PMID- 10695601 TI - Sex-specific trends in validated coronary heart disease rates in southeastern New England, 1980-1991. AB - Although the national decline in coronary heart disease mortality began earlier and was steeper in women relative to men, recent data suggest that the decline in women has slowed. The purpose of this study was to document sex-specific trends in coronary disease morbidity and mortality for the period 1980-1991 in two southeastern New England communities, and to determine whether temporal trends have been similar in men and women aged 35-74 years. Analyses were based on 6,282 validated in-hospital and out-of-hospital coronary disease events ascertained by the retrospective surveillance system of the Pawtucket Heart Health Program. Total (fatal plus non-fatal) coronary disease rates remained stable during this period. The flat trend was the result of an increase in non-fatal hospitalizations and a simultaneous decrease in both in-hospital and out-of hospital mortality. The decline in fatal coronary disease was steeper for men, for both in- and out-of-hospital mortality, although the sex difference was statistically significant only for out-of-hospital deaths. In-hospital case fatality for validated coronary disease declined for both men and women. The steeper decline in coronary disease mortality for men suggests the need for more information regarding sex differential trends in prevention, diagnosis, classification, and treatment. PMID- 10695602 TI - Impact of switching laboratory tests on reported trends in Chlamydia trachomatis infections. AB - Improvements in the sensitivity and specificity of laboratory testing methods for Chlamydia trachomatis infections in recent years have created potential problems with interpreting data on chlamydia prevalence trends. A switch to a more sensitive test can result in an increase in chlamydia positivity even with no increase in the true disease prevalence. To examine the impact of switching laboratory testing methods on chlamydia positivity trends among women, the authors analyzed data from chlamydia screening programs in family planning clinics in two geographic areas of the United States. Data from 7,287 tests performed in Philadelphia, Pennsylvania, indicated a 46% increase in positivity (from 4.1% to 6.0%) when the clinics switched from a nucleic acid probe assay to a ligase chain reaction test. Data from 35,306 tests performed in Oregon and Washington State laboratories showed a 21% increase in positivity (from 3.3% to 4.0%) when clinics switched from a direct immunofluorescent antibody testing procedure to an enzyme immunoassay with negative gray zone confirmation. These increases were within ranges consistent with the variability of the testing methods and occurred primarily in asymptomatic women and in women over age 20 years. Any switch in laboratory testing methods must be considered when interpreting data on chlamydial infection trends. PMID- 10695603 TI - Re: "Controls who experienced hypothetical causal intermediates should not be excluded from case-control studies". PMID- 10695604 TI - Re: "Alcohol intake assessment: the sober facts". PMID- 10695605 TI - Role of Na+-glucose cotransport in jejunal meal-induced absorption. AB - In awake dogs, meal ingestion stimulates the absorption of water and electrolytes from neurovascularly intact jejunal Thiry-Vella loops, even though these loops are isolated from the remainder of the gut. This study was designed to investigate the role of Na+-glucose cotransport in mediating this event. Meal ingestion enhanced absorption when the jejunal lumen was perfused with an isotonic solution containing D-glucose, D-galactose, or 3-O-methylglucose. This response was absent when the perfusate contained mannitol or when phlorizin was added to the D-glucose solution. Mucosa from the jejunal loops was serially biopsied and assayed for brush-border Na+-glucose cotransporter (SGLT1) mRNA and protein expression. Although no changes in SGLT1 mRNA levels were observed, protein levels significantly increased within 30 min following meal ingestion. The time course of SGLT1 protein expression corresponded with that of increased Na+ and water absorption. These results suggest that meal-stimulated jejunal absorption may be mediated through an induction of mucosal SGLT1. PMID- 10695606 TI - Cytokine mRNA expression in mucosal biopsies from German shepherd dogs with small intestinal enteropathies. AB - German shepherd dogs (GSD) are predisposed to enteropathies such as inflammatory bowel disease (IBD) and small intestinal bacterial overgrowth (SIBO). The present study examined the role of cytokines in the immunopathogenesis of both conditions. Duodenal mucosal biopsies were taken from GSDs with small intestinal enteropathies (group 1; N = 16) or control dogs (group 2, N = 12). IL-2, IL-4, IL 5, IL-10, IL-12p40, IFN-gamma, TNF-alpha, and TGF-beta1 mRNA expression was determined by semiquantitative reverse transcriptase polymerase chain reaction. IL-2, IL-5, IL-12p40, TNF-alpha, and TGF-beta1 mRNA expression in group 1 dogs was significantly greater than in group 2 dogs (all P<0.01), but there were no significant differences between dogs with IBD or SIBO. Further, antibiotic treatment in five dogs with SIBO, resulted in reduced TNF-alpha and TGF-beta1 mRNA expression (P<0.05). Such alterations in cytokine mRNA expression suggest heightened immune responses within the duodenal mucosa in GSDs with either SIBO or IBD. PMID- 10695607 TI - Improved screening for intestinal villous atrophy by D-xylose breath test. AB - The hydrogen breath test (H2BT) with D-xylose has proven valid in both early recognition and follow-up of intestinal malabsorption. To further evaluate the specificity of the H2BT with D-xylose in the diagnosis of intestinal malabsorption as compared to the conventional urinary D-xylose test, we analyzed the result in 49 patients referred to our unit with a clinical diagnosis of intestinal malabsorption. These patients had an abnormal 25-g D-xylose H2BT but a normal conventional urinary D-xylose test. Jejunal biopsy with Watson capsule was performed in all patients. H&E staining was prepared from each biopsy specimen, and histological changes were classified according to the Marsh criteria. Jejunal biopsy showed mucosal atrophy in 5 patients (10%), hyperplastic lesion in 11 (22.5%), infiltrative lesion in 14 (28.5%), and normal appearance in 19 (39%). G. lamblia infection was additionally diagnosed in two patients. Histological changes were independent of the presence of diarrhea, weight loss, abdominal pain, or anemia. H2 excretion, assessed as increase over baseline and area under the curve, was similarly independent of the histological pattern. In conclusion, performance of a D-xylose H2BT in patients with a normal urinary test reveals a significant number of patients with intestinal mucosal atrophy who might otherwise remain undiagnosed. PMID- 10695608 TI - Prevalence of electrocardiographic and echocardiographic abnormalities in ambulatory ischemic colitis. AB - The aim of this study was to evaluate the prevalence of cardiac arrhythmia and intracardiac embolic process in ambulatory ischemic colitis. From November 1994 to November 1997, 33 consecutive cases of ambulatory ischemic colitis were detected. This study included 21 women and 12 men with a mean age of 71 years. All patients underwent a cardiovascular investigation including questioning, electrocardiogram, 24-hr ambulatory electrocardiography and transthoracic echocardiography. A prior history of ischemic colitis was found in four cases (12%). Cardiac arrhythmia was detected in eight cases. Transthoracic echocardiography showed an intracardiac process, potentially responsible for a peripheral embolism, in four cases. In conclusion, the aggregate, in 33% of the patients, there was potential cardiac etiology. This suggests that when ambulatory ischemic colitis occurs, it is necessary to perform an exhaustive cardiovascular evaluation similar to those performed in other ischemic diseases. PMID- 10695609 TI - Concerns of patients with inflammatory bowel disease: a review of emerging themes. AB - Idiopathic, chronic inflammatory bowel disease (IBD) refers to two diseases ulcerative colitis (UC) and Crohn's disease (CD). Despite an abundant literature discussing the pathophysiology and treatment of these diseases, little if any empirical studies have focused on patients' subjective experiences with their diseases. The purpose of this paper was to identify and discuss the concerns of individuals with IBD and to suggest that the integration of concerns in clinical management is necessary for a comprehensive understanding of these chronic and debilitating diseases. In addition, case studies were included to highlight the concerns of people with IBD. Our review of the literature identified eight categories of concerns for individuals with IBD. They included loss of energy, loss of control, body image, isolation and fear, not reaching full potential, feeling dirty, and lack of information from the medical community. In conclusion, we argue that the efficacy of treatment for IBD would be greatly improved if psychosocial issues were to be integrated into treatment protocols. PMID- 10695610 TI - Role of psychological stress on IBD onset. PMID- 10695611 TI - Does fasting serum gastrin predict gastric acid suppression in patients on proton pump inhibitors? AB - Proton pump inhibitors (PPIs) block gastric acid secretion and may increase serum gastrin concentration. The aim of this study was to determine whether fasting serum gastrin concentration predicts gastric acid suppression in patients on PPI therapy. Ambulatory pH monitoring with one pH probe in the distal esophagus and a second probe in the stomach was performed in patients with persistent symptoms of GERD despite PPI treatment. Upon completion of pH monitoring, blood was drawn for measurement of fasting serum gastrin concentration. In all, 51 patients were studied: 26 on PPIs, 1 on H2-receptor antagonists, and 24 off acid suppression. Fasting serum gastrin correlated inversely with percent time of gastric pH < 4 for all patients (r = -0.553; P<0.001) and for the subgroup of 26 patients on PPIs (r = -0.435; P = 0.027). In patients on PPIs, an elevated gastrin (> or =100 pg/ml) was associated with gastric pH < 4 for 25+/-7% of the time compared to 54+/-5% when the gastrin was normal (P = 0.004). Therapeutic gastric acid suppression (gastric pH < 4 for <50% of time) was present in 6 of 7 (86%) patients with an elevated fasting serum gastrin, compared with only 8 of 19 (42%) patients with a normal serum gastrin (P<0.05). In conclusion, there is a significant inverse correlation between the fasting serum gastrin concentration and gastric acid profile in patients with GERD. An elevated fasting serum gastrin concentration while on PPI therapy suggests that gastric acid secretion is adequately suppressed. PMID- 10695612 TI - Use of a novel monoclonal antibody in diagnosis of Barrett's esophagus. AB - A novel monoclonal antibody (MAbDAS-1), that specifically reacts with colonic but not small intestinal epithelium, recognizes specialized columnar epithelium (SCE) in the esophagus. The frequency of its reactivity in biopsy specimens of patients with endoscopically suspected Barrett's Esophagus (BE) is examined. Fifty-two biopsy specimens of the distal esophagus from 38 patients were tested by immunoperoxidase method using MAbDAS-1. Fifty-four samples of cardia-type mucosa biopsied from the stomach were used as controls. Results were compared with histology and Alcian blue/high iron diamine (AB/HID). Of the 52 specimens, 29 had glandular epithelium and the rest had only squamous epithelium. Ten were diagnosed to have SCE by histology. All 10 samples reacted with MAbDAS-1 and with Alcian blue. Of the remaining 19 specimens, five also reacted with MAbDAS-1. None of the squamous epithelium and cardia specimens reacted with MAbDAS-1. MAbDAS-1 may detect intestinal metaplasia of the esophagus of colonic phenotype in the absence of histological evidence of SCE. PMID- 10695613 TI - Recurrence of gastric ulcer dependent upon strain differences of Helicobacter pylori in urease B gene. AB - To evaluate the role of different strains of Helicobacter pylori on the recurrence of gastric ulcer, we divided H. pylori into four types (I, II, III, and IV) according to the urease B gene using polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP). The relationship between the recurrence of gastric ulcer and the prevalence of H. pylori types was studied in 32 patients with benign open gastric ulcers using upper gastrointestinal endoscopy. The rate of recurrence was significantly lower in patients with type II than in patients with types I, III, and IV (P<0.05). Using Mongolian gerbils, an animal model of H. pylori infection, we also showed that the occurrence of gastric ulceration following restraint water-immersion stress was significantly lower in type II compared with types I and III. These data indicate that in the context of ulcer recurrence, it is not necessary to eradicate H. pylori during infection with type II. PMID- 10695614 TI - Coexpression of interleukin-8 and inducible nitric oxide synthase in gastric mucosa infected with cagA+ Helicobacter pylori. AB - The cagA-positive Helicobacter pylori strains are thought to be able to induce interleukin-8 expression and to be associated with gastroduodenal diseases. Inducible nitric oxide synthase (iNOS) may be involved in inflammatory pathogenesis. Our aim was to investigate the interrelationships between cagA and the expression of interleukin-8 and iNOS messenger RNAs, and with the type and degree of inflammation in gastric mucosa. In biopsies from 108 Chinese patients, the cagA, 16S rRNA, interleukin-8, and iNOS mRNAs were analyzed using reverse transcription polymerase chain reaction. Specimens infected with cagA-positive strains had significantly more severe infiltration by mononuclear and polymorphonuclear leukocytes and more frequently expressed interleukin-8 and iNOS mRNAs than those infected with cagA-negative strains. iNOS and interleukin-8 mRNAs were significantly more frequently expressed together in the specimens with moderate or severe inflammation than in those with normal mucosa or mild inflammation. Our data suggest that interleukin-8 and excess nitric oxide play important roles in the pathogenesis of H. pylori-associated gastroduodenal diseases. PMID- 10695615 TI - Factors that may affect treatment outcome of triple Helicobacter pylori eradication therapy with omeprazole, amoxicillin, and clarithromycin. AB - Factors affecting Helicobacter pylori eradication rate with omeprazole (OME), clarithromycin (CL), and amoxicillin (AMO) have not been extensively studied. We have investigated the effect of age, sex, smoking, ulcer disease, compliance with therapy, H. pylori colonization density, degree and activity of antral gastritis, the coexistence of corpus gastritis, and the presence of lymphoid follicles on H. pylori eradication rate. We studied 80 consecutive H. pylori-positive patients, with duodenal ulcer (N = 35) or nonulcer dyspepsia (N = 45) treated with OME 20 mg, CL 500 mg, and AMO 1 g, each given twice daily for 10 days. H. pylori was eradicated in 71/80 (88.8%, 95% CI 82-96%) patients. The regimen failed to eradicate the only strain (1.8%, 95% CI 0-5.2%) that was clarithromycin resistant. Multivariate discriminant analysis showed that two histological variables (Wilks lambda = 0.74, chi2 = 23.41, df = 2, P< 0.001), absence of lymphoid follicles in routine gastric biopsies (F = 13.63, P<0.001) and coexistence of antral and body gastritis (F = 13.68, P<0.001), significantly increased H. pylori eradication rate. No other factor examined predicted H. pylori eradication with this regimen. Our data suggest that body gastritis is a positive and presence of lymphoid follicles in routine gastric biopsies is a negative predictive factor of treatment outcome with the omeprazole, clarithromycin, and amoxicillin regime. PMID- 10695616 TI - Effect of pretreatment antibiotic resistance to metronidazole and clarithromycin on outcome of Helicobacter pylori therapy: a meta-analytical approach. AB - Our purpose was to define the effect of pretreatment Helicobacter pylori resistance to metronidazole or to clarithromycin on the success of antimicrobial therapy. We used 75 key words to perform a literature search in MEDLINE as well as manual searches to identify clinical treatment trials that provided results in relation to H. pylori susceptibility to metronidazole and clarithromycin or both during the period 1984-1997 (abstracts were not included). Meta-analysis was done with both fixed- and random-effect models; results were shown using Galbraith's radial plots. We identified 49 papers with 65 arms for metronidazole (3594 patients, 2434 harboring H. pylori strains sensitive to metronidazole and 1160 harboring resistant strains). Metronidazole resistance reduced effectiveness by an average of 37.7% (95% CI = 29.6-45.7%). The variability in the risk difference for metronidazole was 122.0 to -90.6 and the chi-square value for heterogeneity was significant (P<0.001). Susceptibility tests for clarithromycin were performed in 12 studies (501 patients, 468 harboring H. pylori strains sensitive to clarithromycin and 33 harboring resistant strains). Clarithromycin resistance reduced effectiveness by an average of 55% (95% CI = 33-78%). We found no common factors that allowed patients to be divided into subgroups with additional factors significantly associated with resistance. In conclusion, metronidazole or clarithromycin pretreatment resistant H. pylori are the main factors responsible for treatment failure with regimens using these compounds. If H. pylori antibiotic resistance continues to increase, pretherapy antibiotic sensitivity testing might become necessary in many regions. PMID- 10695617 TI - Efficacy of reduced dosage of rabeprazole in PPI/AC therapy for Helicobacter pylori infection: comparison of 20 and 40 mg rabeprazole with 60 mg lansoprazole. AB - Proton pump inhibitor (PPI)- based triple therapy has been a recent trend for treatment of Helicobacter pylori infection, with the PPI-amoxicillin clarithromycin (PPI/AC) regimen being one of the most popular. We have reported the effectiveness of PPI/AC regimens in the Japanese population and have demonstrated that the effectiveness of 40 mg rabeprazole, a recently developed PPI, is similar to that of 40 mg of omeprazole and 60 mg of lansoprazole when used in combination with amoxicillin and clarithromycin. In this study, we focused on whether 20 mg of rabeprazole is effective in our patient population by comparing that dosage with 40 mg of rabeprazole and 60 mg of lansoprazole. In all, 308 H. pylori-infected patients [236 men and 72 women; age (mean +/- SEM) 49.3+/-0.6 years] with peptic ulcer disease (N = 270) or nonulcer dyspepsia (N = 38) were randomly assigned to one of three different PPI/AC regimens for seven days: LAC (N = 104), consisting of lansoprazole 30 mg twice a day, amoxicillin 500 mg three times a day, and clarithromycin 200 mg twice a day; RAC (N = 104), consisting of rabeprazole 20 mg twice a day, amoxicillin 500 mg three times a day, and clarithromycin 200 mg twice a day; and the R1/2AC regimen (N = 100), which included rabeprazole 10 mg twice a day, amoxicillin 500 mg three times a day, and clarithromycin 200 mg twice a day. Cure of the infection was determined by the [13C]urea breath test one month after completion of the treatment. Intention-to-treat based and per-protocol based cure rates for the LAC, RAC, and R1/2AC regimens were 82.7 (95% CI, 74-89) and 88.7% (81-94), 85.6 (77-92) and 89.8% (82-95), and 87.0 (79-93) and 89.7% (82-95), respectively. Although adverse effects were reported by 20.3% of the patients, these affected compliance in only five patients in the RAC and LAC regimens and none in the R1/2AC group. Overall complete compliance was achieved in 94.7% of interviewed patients. In conclusion, the effectiveness of the PPI/AC regimen with 20 mg of rabeprazole is comparable with and even safer than that of 40 mg of rabeprazole and 60 mg of lansoprazole in our patient population. PMID- 10695618 TI - Helicobacter pylori water-soluble surface proteins activate human neutrophils and up-regulate expression of CXC chemokines. AB - To elucidate the mechanisms of the persistent neutrophil recruitment in H. pylori infected gastric mucosa, we evaluated the activation of human neutrophils and CXC chemokine expression in neutrophils by H. pylori water-soluble surface proteins. H. pylori water extract (HPWE) was prepared from a supernatant of the H. pylori suspension in distilled water. After neutrophils were stimulated with HPWE, the mobilization of intracellular free calcium, the expression of lymphocyte function associated antigen-1beta, and the secretion of myeloperoxidase (MPO) were enhanced in the neutrophils. In H. pylori-infected gastric mucosa, transendothelial and transepithelial migration of neutrophils were observed by electron microscopy and mucosal MPO levels were elevated. Up-regulation of the expression of interleukin-8 (IL-8) and growth-related oncogenes (GROs; GROalpha, GRObeta and GROgamma) mRNA and protein in neutrophils by HPWE was demonstrated by quantitative reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. In conclusion, H. pylori-induced neutrophil recruitment may be mediated by CXC chemokines which are expressed by neutrophils activated by H. pylori water-soluble surface proteins. PMID- 10695619 TI - Involvement of reactive oxygen species and nitric oxide in gastric ischemia reperfusion injury in rats: protective effect of tetrahydrobiopterin. AB - The purpose of this study was to examine whether tetrahydrobiopterin (BH4), a cofactor of nitric oxide (NO) synthase, attenuates gastric ischemia-reperfusion injury induced by clamping of the celiac artery. Gastric injury was assessed by a formation of gastric mucosal erosions. The gastric injury was observed at 30 and 60 min after reperfusion following 30-min ischemia and was reduced by superoxide dismutase (SOD), catalase, or NO synthase inhibitors. Therefore, reactive oxygen species (ROS) and NO seem to be implicated in the ischemia-reperfusion injury. Treatment with BH4 reduced the ischemia-reperfusion injury. Pretreatment with sepiapterin, a precursor of BH4, also reduced the ischemia-reperfusion injury with an increase in BH4 content in serum and stomach. Both the increase in BH4 content and the protective effect of sepiapterin were prevented of pretreatment with N-acetylserotonin, an inhibitor of BH4 synthesis. These results suggest that the increase in BH4 content may protect against gastric ischemia-reperfusion injury via reduction of ROS and/or NO toxicity. BH4 might be useful as a therapeutic agent for gastric ischemia-reperfusion injury. PMID- 10695620 TI - Topical prostaglandin E2 protects isolated gastric mucosa against acidified taurocholate-, but not ethanol- or aspirin-induced injury. AB - This study investigates whether topical prostaglandins protect isolated gastric mucosa against injury provoked by acidified "barrier-breaking" agents. Intracellular pH (pHi), apical cell membrane potential (Vcm) and intraepithelial resistances in isolated Necturus antral mucosa were measured using double barreled liquid sensor microelectrodes. Topical PGE, treatment protected the antral mucosa against acidified taurocholate-induced injury, reducing significantly (P<0.05) intracellular acidification (pHi from 7.39+/-0.05 to 7.08+/-0.08 vs. from 7.30+/-0.02 to 6.62+/-0.15), and opposing significantly the changes in Vcm (hyperpolarization followed by depolarization), and completely abolishing the decrease in transmembrane resistance (Rt from 702+/-37 to 723+/-39 Ohms x cm2 vs. from 721+/-34 to 270+/-105 Ohms x cm2). Also the ratio of apical and basolateral membrane resistances (Ra/Rb) remained at a significantly higher level in PGE2-treated tissues. In contrast, PGE2 treatment had no protective influence on the changes of the respective parameters in acidified ethanol or acetylsalicylic acid injured mucosas. Topical prostaglandin E2 protects isolated gastric mucosa against acidified taurocholate, but not against ethanol- or acetylsalicylic acid-induced injury. PMID- 10695621 TI - Protection by intragastric polyethylene glycol 400 in rat stomach against ethanol damage involves alpha2-adrenoceptors. AB - The aim of this study was to investigate the role of alpha2-adrenoceptors in the mechanism of intragastric polyethylene glycol 400 (PEG-400) protection against ethanol-induced gastric mucosal damage. In the injury study, 0.5 hr after subcutaneous control or yohimbine (5 mg/kg), a selective alpha2-adrenoceptor antagonist, rats were treated with intragastric vehicle or PEG-400 (5500 mg/kg). One hour later animals received 96% ethanol (gavage needle), 5 ml/kg, and the rats were killed after another hour. Total lengths of the gastric mucosal lesions were measured by an unbiased observer in a blinded fashion using a binocular magnifier having 5x magnification. In a separate set of experiments, 0.5 hr after subcutaneous control or yohimbine (5 mg/kg) rats received intragastric vehicle or PEG-400 (5500 mg/kg). One hour later gastric mucus volume, gastric juice volume, and gastric acid output in the gastric juice were measured. The protective effect offered by intragastric PEG-400 against ethanol-induced gastric mucosal damage was significantly diminished although not completely abolished by a selective alpha2-adrenoceptor antagonist (yohimbine). Yohimbine also significantly diminished both the basal and PEG-400-stimulated increase in gastric mucus volume. These findings suggest that the protective effect afforded by intragastric PEG-400 against ethanol-induced gastric mucosal damage is partially mediated by alpha2-adrenoceptors, and a mucus-dependent mechanism may be involved. PMID- 10695622 TI - GERD progressing to diffuse esophageal spasm and then to achalasia. AB - The pathophysiology of achalasia is not completely understood. Several reports have suggested that esophageal motility disorders may progress from one type to another. We report a patient with symptoms and esophageal motility findings consistent with gastroesophageal reflux who subsequently developed a diffuse esophageal spasm and then achalasia. We believe this to be the first report showing such a progression in esophageal motility. PMID- 10695623 TI - Expression of tissue inhibitors of metalloproteinases (TIMPs) in gastric cancer. AB - Matrix metalloproteinases (MMPs) are one of the major classes of proteolytic enzymes involved in tumor invasion and metastasis, being inhibited by naturally occurring tissue inhibitors of metalloproteinases (TIMPs). Sixty-five patients who underwent surgery for gastric cancer in 1992 at Chonnam National University Hospital were selected for this study. The primary selection criteria were the availability of formalin-fixed and paraffin-embedded blocks and sufficient clinical follow-up for tumor-specific survival analysis. In this study, we examined the expression of TIMP-1 and TIMP-2 in human gastric cancer tissue by in situ hybridization and immunohistochemistry, and the correlation between their expression and clinicopathological parameters. TIMP-1 and TIMP-2 expressions were detected predominantly in the peritumor stromal cells rather than tumor cells themselves. Immunohistochemical stainings were concordant with the result obtained by in situ hybridization. The intensity of TIMP-1 immunohistochemical stromal staining correlated with tumor stage (P = 0.009) and patient survival (P = 0.025). However, the intensity of TIMP-2 immunohistochemical stromal staining did not correlate with tumor stage (P = 0.339) and patient survival (P = 0.474). The correlation between the increased TIMP-1 expression and cancer stage noted in this study reflects a role of TIMP-1 in predicting the aggressive behavior of gastric cancer. TIMP-2 expression did not correlate with clinicopathological parameters. However, expression of TIMP-1 and the possible additional value of TIMP-2 should be further explored in determining the prognosis of gastric cancer. PMID- 10695624 TI - Circulating p53 antibody in patients with colorectal cancer: relation to clinicopathologic features and survival. AB - The presence of serum anti-p53 antibody has been reported to be associated with survival of patients with breast cancer, ovarian cancer, and hepatocellular carcinoma. To clarify prognostic significance of p53 antibody in colorectal cancer, serum p53 antibody was measured in patients with colorectal cancer. The 89 patients included 71 with colorectal cancer and 18 with colon polyp. An enzyme linked immunosorbent assay was used to detect p53 antibodies in serum. Clinicopathological parameters such as age, sex, degree of differentiation of cancer, location of tumor, liver metastasis, stage classification, Dukes classification, CEA, CA19-9, and immunostaining of p53 and anti-p53 antibody were evaluated as prognostic factors of colorectal cancer. p53 antibody was positive in 18 of 71 (25%) with colorectal cancer, whereas it was positive in only 1 of 18 (6%) with colon polyp. The patients with p53 antibody had higher CEA and CA19-9 levels, higher positive rates of p53 protein expression in cancer cells, and higher liver metastasis rates. The p53 antibody positivity at stage classification I-IIIb/ Dukes classification A-C was significantly lower than that at stage classification IV/Dukes classification D. Overall survival in colorectal cancer patients with p53 antibody was significantly shorter than in those without p53 antibody. A Cox regression analysis showed that liver metastasis, stage classification, Dukes classification, CA19-9, and p53 antibody were significant prognostic factors in colorectal cancer. Serum anti-p53 antibody could serve as one of the prognostic factors in patients with colorectal cancer. PMID- 10695625 TI - Distension-induced myoelectrical dysrhythmia and effect of intestinal pacing in dogs. AB - The aims of this study were to investigate the effect of duodenal distension on intestinal myoelectrical activity and to investigate whether intestinal pacing was able to reverse the effects of distension. Six female hound dogs with four pairs of electrodes on the proximal jejunum were involved in this study. The protocol consisted of 30 min of recording of jejunal myoelectrical activity as baseline and 90 min of recording during distension. Intestinal pacing was performed during the second 30 min of distension. Duodenal distension severely impaired intestinal myoelectrical activity. The percentage of normal slow waves was reduced from 90.8+/-8.4% at baseline to 73.8+/-10.2%, 57.2+/-11.4%, and 53.7+/-16.0% during the first, second and third 30 min of distension (P<0.05, ANOVA). The dominant power was similarly decreased and the minute-by-minute variation of dominant frequency was significantly increased after distension. Intestinal pacing reversed distension-induced dysrhythmia. The percentage of normal slow waves during the 30 min of distension with pacing was significantly higher than the corresponding 30 min of distension without pacing (88.5+/-6.6% vs. 57.2+/-11.4%, P<0.03). It was concluded that intestinal pacing can normalize distension-induced dysrhythmia and has a potential as a future therapeutic modality for intestinal motor disorders. PMID- 10695626 TI - New method for evaluating intestinal contractions in guinea pig by curve fitting. AB - Using a "hybrid logistic function," we attempted to develop a new approach for a quantitative and comprehensive evaluation of the force-time curve of guinea pig gut contractions. We recorded ileum twitch and proximal colon spontaneous isometric longitudinal contractions because of their high regularities. We digitized the force-time curves of both contractions and performed curve fitting to them by hybrid logistic functions with a personal computer. We found that the fitness of these functions to both contractions was excellent. The respective best-fit parameters of these functions were closely correlated with the observed mechanical indexes, all of which are physiologically meaningful. This result suggests the possibility that these parameters can characterize the magnitudes and time courses of F(t) curves of the intestinal contractions. Furthermore, it might be able to show an effect of a pharmacological agent specifically either on the contraction phase, the relaxation phase, or other parameters of each. Therefore, we insisted that the present new approach for evaluating gut motility is promising for clinical application. PMID- 10695627 TI - Prevention of biliary stent occlusion by ursodeoxycholic acid plus norfloxacin: a multicenter randomized trial. AB - We report a prospective randomized multicenter trial that tested the efficacy of combining ursodeoxycholic acid and norfloxacin in the prevention of polyethylene stent clogging in patients with obstructive jaundice due to an unresectable malignancy at the level of the common bile duct. After insertion of a 10-Fr straight polyethylene stent, patients were allocated to receive oral treatment with ursodeoxycholic acid and norfloxacin, or conservative treatment. The primary outcome measure was stent blockage within six months. Thirty-three patients (group I) received ursodeoxycholic acid and norfloxacin, and 29 received conservative treatment (group II). At six months, cumulative stent patency rate did not differ significantly between group I (47+/-11%, mean +/- SE, median 149 days) and group II patients (24+/-10%, mean +/- SE, median 100 days, P = 0.23, log-rank test). Four stents were clogged by ursodeoxycholic acid. Survival did not differ between the two groups. Combined therapy with ursodeoxycholic acid and norfloxacin failed to improve stent patency. Moreover, ursodeoxycholic acid can cause stent obstruction. PMID- 10695628 TI - Expression of transforming growth factor-beta in spontaneous chronic pancreatitis in the WBN/Kob rat. AB - Transforming growth factor-beta1 (TGF-beta1) is suggested to be a mediator of fibrosis in chronic pancreatitis, but the serial change of TGF-beta1 expression in the onset and progression of chronic pancreatitis is still unclear. We investigated the TGF-beta1 expression in the spontaneous chronic pancreatitis model. Four-week-old male WBN/Kob rats were fed with special pellet diet (MB-3) for 20 weeks. TGF-beta1 mRNA in the pancreas was detected by reverse transcription-polymerase chain reaction assay from four weeks, and its expression peaked at 12 weeks when the pancreatic fibrosis first appeared. The localizations of TGF-beta1 mRNA and protein were confirmed in the cytoplasm of pancreatic acinar and ductal cells by in situ hybridization and immunohistochemistry, respectively. Although fibronectin expression peaked at 12 weeks and correlated with that of TGF-beta1, its elevated expression tended to be prolonged. Pancreatic fibrosis peaked at 16 weeks after the peak of TGF-beta1 expression. These results suggest that TGF-beta1 expression may be a trigger of the fibrotic process of chronic pancreatitis in the WBN/Kob rat. PMID- 10695629 TI - Vanishing bile duct syndrome associated with chronic EBV infection. AB - We reported here an adult patient with vanishing bile duct syndrome due to chronic EBV infection. A 22-year-old male was admitted to a nearby hospital complaining of a sore throat and jaundice. He received a high dose of prednisolone for bile stasis of acute viral hepatitis. However, the hepatitis did not improve, and he was transferred to our hospital. He had exhibited jaundice for one year as well as hemophagocytic syndrome and intestinal perforation. Subtotal intestinal resection was successfully performed. Three follow-up biopsied liver specimens indicated vanishing bile duct syndrome. Positive results of EBV-DNA in his serum and mRNA of EBV by in situ hybridization of his liver indicated that massive doses of prednisolone caused chronic EBV infection and vanishing bile duct syndrome. PMID- 10695630 TI - Fecal elastase 1 is not the indirect pancreatic function test we have been waiting for. PMID- 10695631 TI - Giant solitary fibrous tumor of the liver with metastasis to the skeletal system successfully treated with trisegmentectomy. PMID- 10695632 TI - Quantitation of hepatitis C virus RNA in plasma and peripheral blood mononuclear cells of patients with chronic hepatitis treated with interferon-alpha. AB - We quantified hepatitis C virus (HCV) RNA at different times in plasma and peripheral blood mononuclear cells (PBMC) in 51 patients with chronic hepatitis C undergoing interferon-alpha2a (IFN-alpha2a) therapy. HCV RNA loads in plasma correlated with those in PBMC before and during the treatment (P<0.001). After treatment, a sustained response was observed in 19 patients (SR), a response followed by relapse in 9 (RR), and non response in 23 (NR). By univariate analysis PBMC HCV RNA load before treatment was lower in SR than in RR and NR (P = 0.003). In the 9 RR, HCV RNA disappeared in PBMC before or at the same time as in plasma and again became detectable in plasma and PBMC simultaneously or earlier in plasma. These results indicate that quantitation of HCV RNA in PBMC is not a useful parameter for the follow-up of treated patients. PMID- 10695633 TI - Correlation of genotypes and route of transmission with histologic activity and disease stage in chronic hepatitis C. AB - Our objective was to evaluate the histopathological features of chronic hepatitis C of 64 liver biopsies and to correlate this with the route of transmission of hepatitis C virus, the genotype of HCV, and the patient's age. Moderate chronic hepatitis was the most frequently observed (62.5%). Cirrhosis was observed in 14 patients (21.9%) and was more frequently found among patients over 40 years of age (34.3% vs. 6.9%, P = 0.025). The mean histopathological activity index (HAI) was significantly higher in the sporadic (10+/-3.1) than the posttransfusional (7.5+/-3.7) and the intravenous drug use (IVDU) groups (6.3+/-2.8) (P<0.02). Moreover the sporadic group showed more fibrosis (P<0.04) than the posttransfusional group. No liver cirrhosis was found in the IVDU group. The overall prevalence of HCV variants was: 54.7% type 1b, 4.6% type 1a, 37.5% type 2c, 1.6% type 2b, 1.6% type 2. The genotype distribution showed no relation to the HAI, hepatitis activity (grade), and fibrosis (stage) of the liver disease. In conclusion, the sporadic route of transmission of HCV was related to a more severe chronic hepatic disease, a finding that could influence future antiviral therapies. The predominance of HCV type 1b in this study reflects the higher frequency of this variant in our area. Our data suggests that the ultimate consequence of HCV chronic infection depends on patient age rather than on HCV genotype. PMID- 10695634 TI - Lower hepatitis G virus infection prevalence compared to hepatitis B and C virus infection prevalences. AB - To more accurately determine the seroprevalence of hepatitis G virus (HGV) infection, we surveyed antibody to HGV (anti-E2) by enzyme-linked immunosorbent assay (ELISA) and HGV RNA by nested polymerase chain reaction (PCR) in 298 residents of a hepatitis C virus (HCV)-endemic area of Japan and in 225 hemodialysis patients. We then compared these findings with known HCV and hepatitis B virus (HBV) infection prevalences. Anti-E2 and HGV RNA prevalences were 32 (10.7%) and 5 (1.7%) in the residents and 24 (10.7%) and 10 (4.4%) in the hemodialysis patients, respectively. Anti-E2 and HGV RNA concurrence was found in two of the hemodialysis patients. Total HGV marker (anti-E2 and/or HGV RNA) prevalences [37 (12.4%) in residents and 32 (14.2%) in hemodialysis patients], were significantly lower than the prevalences of antibody to HCV (anti-HCV) by ELISA [59 (19.8%) and 96 (42.7%)], and antibody to hepatitis B core antigen (anti HBc) by radioimmunoassay (RIA) [87 (29.2%) and 101 (44.9%)] (P<0.05). The anti HCV prevalence in subjects with total HGV marker was significantly higher than in those without total HGV marker. There was no significant difference in anti-HBc prevalence between those with and without total HGV marker. The viremic rate was highest in HCV infection (HCV RNA by PCR/anti-HCV) (83.2%), with HGV infection (HGV RNA/total HGV marker) (21.7%) intermediate, and HBV infection (hepatitis B surface antigen by RIA/anti-HBc) (5.3%) lowest (P<0.05). These findings indicate that HGV infection was less endemic than HCV and HBV. HGV was eliminated naturally more frequently than HCV infection and less frequently than HBV infection. PMID- 10695635 TI - Spleen enlargement in patients with nonalcoholic fatty liver: correlation between degree of fatty infiltration in liver and size of spleen. AB - Our purpose was to determine if there is an association between nonalcoholic fatty liver and spleen enlargement. Spleen volume was measured by computed tomography (CT) in 32 patients with nonalcoholic fatty liver (23 men and 9 women; age, 41.6+/-12.1, range, 22-69 years) and 34 patients with normal liver (19 men and 15 women; age, 51.1+/-16.2, range, 14-86 years). The values were compared with the patient's demographic data, the liver-to-spleen (L/S) ratio of CT Hounsefield unit measurements, and the results of liver function tests. Diagnosis of fatty liver was made if the L/S ratio was less than 1.0. The mean spleen volume was 73.0+/-24.4 cm3 (range, 21.1-106.1) in normal subjects and 141.2+/ 54.1 cm3 (range, 44.1-267.3) in patients with fatty liver (P<0.0001). Multivariate linear regression analysis identified that only the L/S ratio (P<0.0001) and age (P<0.01) were significantly correlated with spleen volume. Using forward selection stepwise regression, the L/S ratio entered first (beta = 0.634) and age second (beta = -0.293). In conclusion, spleen enlargement was commonly seen in patients with nonalcoholic fatty liver, and the recognition of this association may halt further attempts to evaluate the cause of spleen enlargement. PMID- 10695636 TI - Calcium-channel blocker attenuates Kupffer cell production of cytokine-induced neutrophil chemoattractant following ischemia-reperfusion in rat liver. AB - We investigated the effects of the calcium-channel blocker verapamil hydrochloride on the production of cytokine-induced neutrophil chemoattractant (CINC) following reperfusion injury in rat liver. Ischemia was induced for 30 min by portal vein occlusion. Animals were pretreated with intravenous injection of verapamil hydrochloride (2.5 mg/kg) 5 min before vascular clamp. Verapamil hydrochloride limited increases in the chemoattractant compared with nonpretreated rats. Most cells immunostained for chemoattractant were ED2 positive macrophages in sinusoids. In vitro chemoattractant production by Kupffer cells isolated from animals pretreated with verapamil hydrochloride was significantly lower than by Kupffer cells from nonpretreated animals. Expression of transcripts in liver for chemoattractant peaked 3 hr after reperfusion in nonpretreated animals, while pretreatment with verapamil hydrochloride significantly decreased chemoattractant mRNA levels. In vitro chemoattractant production could be induced in naive Kupffer cells after stimulation with oxygen radicals generated by hypoxanthine and xanthine oxidase, but verapamil hydrochloride prevented these increases. We concluded that the calcium-channel blocker verapamil hydrochloride significantly attenuates chemoattractant release by Kupffer cells after ischemia-reperfusion in the rat liver. PMID- 10695637 TI - Hepatic injury due to troglitazone. PMID- 10695638 TI - Molecular neurobiology for practicing psychiatrists, Part 5: How a leucine zipper can turn on genes: immediate-early genes activate late-gene expression in the brain. PMID- 10695639 TI - The increasing use of polypharmacotherapy for refractory mood disorders: 22 years of study. AB - BACKGROUND: Few studies have approached the subject of polypharmacotherapy systematically. This retrospective review of 178 patients with refractory bipolar disorder or unipolar depression (Research Diagnostic Criteria or DSM-III-R criteria) discharged from the National Institute of Mental Health (NIMH) Biological Psychiatry Branch between 1974 and 1996 was conducted to assess the degree and efficacy of "add-on" pharmacotherapy. METHOD: Following completion of formal structured blinded research protocols, patients entered a treatment phase (often again on a blind basis) in which all agents available in the community could be utilized. Each patient's retrospective life chart and all prospective double-blind nurse- and self-rated NIMH data were reviewed. The overall degree of improvement at discharge was assessed by rating on the Clinical Global Impressions scale (CGI) as modified for bipolar illness (CGI-BP). RESULTS: A 78% improvement rate (moderate or marked on the CGI) was achieved at the time of discharge. There was a significant relationship between number of medications utilized at discharge as a function of discharge date (r = 0.45, p < .0001). The percentages of patients discharged on treatment with 3 or more medications were 3.3% (1974-1979), 9.3% (1980-1984), 34.9% (1985-1989), and 43.8% (1990-1995). No correlation was found between polypharmacy and age (r = -0.03, p = .66). Patients more recently discharged from the NIMH had an earlier age at illness onset, more lifetime weeks depressed, and a higher rate of rapid cycling than patients in the earlier cohorts. CONCLUSION: Increasing numbers of medications in more recent NIMH cohorts were required to achieve the same degree of improvement at hospital discharge. More systematic approaches to the complex regimens required for treatment of patients with refractory mood disorder are clearly needed. PMID- 10695640 TI - Racial variation in antidepressant treatment in a Medicaid population. AB - BACKGROUND: Many studies have found racial and socioeconomic variation in medical care for a variety of conditions. Undertreatment of depression for individuals of all races is a concern, but especially may affect vulnerable populations such as Medicaid recipients and minorities. With this study, we examine racial differences in the antidepressant usage in a Medicaid population. METHOD: Treatment of 13,065 depressed patients (ICD-9-CM criteria) was examined in a state Medicaid database covering the years 1989 through 1994. Treatment differences were assessed in terms of whether an antidepressant was received at the time of the initial depression diagnosis and the type of antidepressant prescribed (tricyclic antidepressants [TCAs] vs. selective serotonin reuptake inhibitors [SSRIs]), using logistic regression techniques. RESULTS: African Americans were less likely than whites to receive an antidepressant at the time of their initial depression diagnosis (27.2% vs. 44.0%, p < .001). Of those receiving an antidepressant, whites were more likely than African Americans to receive SSRIs versus TCAs. These findings remained even after adjusting for other covariates. CONCLUSION: Despite the easy availability of effective treatments, we found that only a small portion of depressed Medicaid recipients receive adequate usage of antidepressants. Within this Medicaid population, limited access to treatment was especially pronounced among African Americans. Racial differences existed in terms of whether an antidepressant was received and the type of medication used. PMID- 10695641 TI - Clozapine treatment in a population of adults with mental retardation. AB - BACKGROUND: There is a paucity of data on the use of clozapine in patients with mental retardation and comorbid psychiatric illness. The authors describe their recent clinical experience using clozapine in treatment-refractory patients with mental retardation and severe psychiatric illness. METHOD: A retrospective review was performed on the records of all patients admitted to a university-affiliated, specialized inpatient psychiatry service who were selected for clozapine therapy from March 1994 through December 1997 (N = 33). Patients had DSM-IV diagnoses of schizophrenia, schizoaffective disorder, bipolar disorder, delusional disorder, or psychotic disorder NOS and were considered treatment resistant. All had deficits in functioning well beyond those expected for their degree of cognitive deficits and adaptive delays. RESULTS: Of 33 initial patients, 26 remained on clozapine therapy for a follow-up duration of 5 to 48 months (mean = 24.8 months). Evaluation at follow-up revealed Clinical Global Impressions-Improvement (CGI-I) scores from 1 to 4 with a mean +/- SD improvement of 2.0 +/- 0.8 (much improved). The mean +/- SD rating of the CGI-Efficacy Index was 5 +/- 2.6 (decided improvement and partial remission of symptoms with no interference from side effects). The 6 patients who were not maintained on clozapine therapy over the study period did not significantly differ from the clozapine group in gender, race, age, side effects, or diagnosis. One patient was lost to follow-up. Side effects were mild and transient with constipation being the most common (N = 10). There were no significant cardiovascular side effects and no seizures. No patients discontinued treatment due to agranulocytosis. CONCLUSION: The current investigation lends support to the conclusion that clozapine appears to be safe, efficacious, and well tolerated in individuals with mental retardation and comorbid psychiatric illness. PMID- 10695642 TI - Venlafaxine versus fluvoxamine in the treatment of delusional depression: a pilot double-blind controlled study. AB - BACKGROUND: Previous studies have reported the efficacy of selective serotonin reuptake inhibitors as monotherapy in the treatment of delusional depression. The clinical efficacy of venlafaxine, a serotonin-norepinephrine reuptake blocker, has been demonstrated in the treatment of patients with moderate-to-severe depression, but, to date, no evidence is available about its use in depressed patients with psychotic features. METHOD: Under double-blind conditions, 28 hospitalized patients who met DSM-IV criteria for major depression, severe with psychotic features, were randomly assigned to receive fluvoxamine or venlafaxine, 300 mg/day, for 6 weeks. Severity was evaluated using the Hamilton Rating Scale for Depression (HAM-D) and the Dimensions of Delusional Experience Rating Scale (DDERS) administered at baseline and every week thereafter. Side effects were also recorded. Clinical response was defined as a reduction of the scores in the 21-item HAM-D to 8 or below and in the DDERS to 0. RESULTS: At study completion, the response rates were 78.6% (N = 11) and 58.3% (N = 7) for fluvoxamine and venlafaxine, respectively. No significant difference was found between drugs (Fisher exact test, p = .40). Analysis of covariance on HAM-D scores did not reveal a significantly different decrease of depressive symptomatology between the 2 treatment groups (p = .14). Treatment response appeared to be unrelated to the demographic and clinical characteristics recorded. The overall safety profile of both fluvoxamine and venlafaxine was favorable. CONCLUSION: The results of this pilot double-blind trial show that fluvoxamine is useful in the treatment of delusional depression and suggest that venlafaxine may also be an effective compound in the treatment of this disorder. The latter finding, although promising, warrants further replication in a larger sample of patients. PMID- 10695643 TI - Effects of light therapy on suicidal ideation in patients with winter depression. AB - BACKGROUND: Recent case reports suggest that some patients with seasonal affective disorder (SAD) may become suicidal after initial treatment with light therapy. This retrospective study sought to determine the effects of light therapy on suicidal ideation in patients with SAD. METHOD: The cases of 191 depressed patients with SAD by DSM-III-R or DSM-IV criteria treated with an open trial of morning light therapy using cool white fluorescent light boxes (2500 lux for 2 hours per day or 10,000 lux for 30 minutes per day) for 2 weeks were retrospectively analyzed. Patients had been rated before and after treatment with the expanded Hamilton Depression Rating Scale (SIGH-SAD). RESULTS: Sixty-seven percent of patients were rated as clinical responders to light therapy. There was significant improvement in the SIGH-SAD suicide item score, with 45% of patients showing a reduction in score. Only 6 patients (3%) had slight worsening of suicide scores. No patients attempted suicide or discontinued light therapy because of emergent suicidality. CONCLUSION: Light therapy relieves suicidal ideation in patients with SAD consistent with overall clinical improvement. Emergence of suicidal ideas or behaviors is very uncommon with light therapy. PMID- 10695644 TI - Antipsychotic drug treatment in first-episode mania: a 6-month longitudinal study. AB - OBJECTIVE: To determine the use of antipsychotics during and following inpatient treatment of patients with a first-episode of mania. METHOD: The 198 subjects available for analysis were 129 consecutively hospitalized first-episode manic patients and 69 nonaffective psychotic patients assessed at admission and at 6 month follow-up postdischarge. Comparisons between the groups were made on frequency, type, and doses of antipsychotics prescribed during and after hospitalization in relation to clinical status. RESULTS: First-episode manic patients were given lower mean +/- SD daily doses of antipsychotics than nonaffective psychotic patients at discharge (163 +/- 132 mg chlorpromazine equivalents [CPZe] vs. 224 +/- 167 mg CPZe, p = .0102), at 6-month follow-up (109 +/- 167 mg CPZe vs. 260 +/- 178 mg CPZe; p = .0001), and if recovered (110 +/- 174 mg CPZe vs. 265 +/- 207 mg CPZe, p = .0014). At 6-month follow-up, 31 (24%) of 129 manic and 24 (35%) of 69 nonaffective psychotic patients continued to receive antipsychotics (NS). There was no difference between the groups in the time to discontinuation of antipsychotic agents. The mean time to drug discontinuation in manic patients was 98 days. CONCLUSION: (1) Antipsychotic doses at discharge and at 6-month follow-up were much lower in manic than in nonaffective psychotic patients, although there was no significant difference in the proportion of patients who continued to receive them 6 months after discharge. (2) The time to discontinuation was independent of clinical outcome. In those who discontinued the antipsychotic agent, the time to discontinuation was more rapid in the manic group than in the nonaffective psychotic patients. PMID- 10695645 TI - Posttraumatic residues of captivity: a follow-up of Israeli ex-prisoners of war. AB - BACKGROUND: This article examines the long-term impact of wartime captivity. METHOD: One hundred sixty-four prisoners of war (POWs) and 189 matched combatants of the 1973 Yom Kippur War filled out a series of questionnaires that assessed posttraumatic stress disorder (PTSD), general psychiatric symptomatology, and social functioning according to DSM-III-R criteria. RESULTS: Almost 2 decades after the war, ex-POWs exhibited higher rates and greater intensity of posttraumatic stress reactions, more general psychiatric symptomatology, and more severe problems in functioning at home, at work, and in the military than did the control group (Israeli veterans who were not POWs). They were also more likely to obtain official disability recognition and to seek psychological help. Their recovery was slower and professional help less effective. In addition, the veterans with PTSD in both groups had high rates of comorbid general psychiatric symptomatology. CONCLUSION: These findings point to the depth, range, and persistence of the stress residuals of wartime captivity. PMID- 10695646 TI - A placebo-controlled trial of cognitive-behavioral therapy and clomipramine in trichotillomania. AB - BACKGROUND: The major treatments reported to be effective in the treatment of trichotillomania are cognitive-behavioral therapy (CBT) with habit reversal and serotonin-norepinephrine reuptake inhibitors such as clomipramine. However, the 2 treatments have not been previously compared with each other. This study examines the efficacy of CBT and clomipramine compared with placebo in the treatment of trichotillomania. METHOD: Twenty-three patients with trichotillomania as determined by the Structured Clinical Interview for DSM-III-R entered and 16 completed a 9-week, placebo-controlled, randomized, parallel-treatment study of CBT and clomipramine. Efficacy was evaluated by the Trichotillomania Severity Scale, the Trichotillomania Impairment Scale, and the Clinical Global Impressions Improvement scale, which were conducted by an independent assessor blinded to the treatment condition. RESULTS: CBT had a dramatic effect in reducing symptoms of trichotillomania and was significantly more effective than clomipramine (p = .016) or placebo (p = .026). Clomipramine resulted in symptom reduction greater than that with placebo, but the difference fell short of statistical significance. Placebo response was minimal. CONCLUSION: Clinicians should be aware of the potential treatments available for trichotillomania. A larger and more definitive study comparing CBT and a serotonin-norepinephrine reuptake inhibitor is indicated. PMID- 10695647 TI - Interpersonal psychotherapy and antidepressant medication: evaluation of a sequential treatment strategy in women with recurrent major depression. AB - BACKGROUND: Few data are available to guide treatment selection in major depression. With increasing pressure to maximize the efficiency and minimize the costs of treatment, it is important to have information that could guide treatment selection or point to treatment strategies that have a high probability of success. METHOD: We used a successive cohort approach to compare 2 highly similar groups of women with recurrent unipolar disorder (DSM-III-R or DSM-IV): one in which the combination of interpersonal psychotherapy (IPT) and pharmacotherapy was initiated at the outset of treatment and a second in which IPT alone was provided first and only those who did not remit with IPT alone were offered the combination treatment. RESULTS: In the group in which the combination was initiated at the outset of treatment (N = 180), the remission rate was 66%, comparable to the remission rate observed in most outpatient treatment studies of major depression. In contrast, among the women in the second cohort who were first treated with IPT alone and only those who did not remit were given combination therapy (N = 159), the remission rate was 79%, significantly greater than that observed in the group that received combination treatment from the outset (chi2 = 6.55, p = .02). CONCLUSION: These results suggest that the strategy of offering IPT to women with recurrent unipolar disorder and, in the absence of remission, adding antidepressant pharmacotherapy can be a highly effective treatment, one that may be particularly attractive to women in the childbearing years. Although slower in its onset of action, this sequential strategy is likely to enable the clear majority of such women to achieve a full remission of depressive symptoms. PMID- 10695648 TI - Gender difference in severity of schizophrenia. PMID- 10695649 TI - Bupropion-amantadine-associated neurotoxicity. PMID- 10695650 TI - Sexual dysfunction on fluvoxamine therapy. PMID- 10695651 TI - Clozapine versus chlorpromazine in geriatric patients. PMID- 10695652 TI - Combination of donepezil and gabapentin for behavioral disorders in Alzheimer's disease. PMID- 10695653 TI - High exposure to neuroleptics in bipolar patients: a retrospective review. AB - BACKGROUND: Acute and long-term use of neuroleptics to treat bipolar disorder remains prevalent despite safety concerns. Neuroleptic-treated patients with bipolar disorder have been reported to have rates of tardive dyskinesia, akathisia, and acute dystonia as high as or higher than patients with schizophrenia. Moreover, the pattern of repeated, intermittent use of neuroleptics in bipolar disorder may increase rather than decrease the risk of tardive dyskinesia. METHOD: Retrospective life charts of 133 treatment-refractory patients with bipolar disorder (diagnosed according to Research Diagnostic Criteria or a clinical interview with the Schedule for Affective Disorders and Schizophrenia-Lifetime Version or the Structured Clinical Interview for DSM-IV Axis I Disorders) admitted to the National Institute of Mental Health (NIMH) were reviewed for prior neuroleptic use, medication exposure, and course of illness variables. Patients' medication response and degree of improvement while at NIMH were also assessed. RESULTS: A total of 72.2% (N = 96) of the bipolar patients examined had exposure to neuroleptics prior to referral to NIMH. Neuroleptic treated patients had a mean of 5.6 neuroleptic trials with a mean duration of 166.4 days for each trial and a dose range of 25 to 960 mg in chlorpromazine equivalents. Life chart data showed that the neuroleptic-exposed and nonexposed bipolar patients were distinguished by 1 course-of-illness variable: increased suicidality in the neuroleptic-treated group. Patients with and without prior neuroleptic exposure experienced the same high degree of improvement at discharge from NIMH. Only 12.5% (N = 12) of the group previously treated with typical neuroleptics (N = 96) required neuroleptics at discharge. CONCLUSION: Our data suggest that the majority of even treatment-refractory bipolar patients can be stabilized without neuroleptics. Given the high risk of tardive dyskinesia and the availability of other novel agents, the routine intermittent use of typical neuroleptics to treat patients with bipolar disorder should be minimized. PMID- 10695654 TI - Immunohematology is 100 years old. PMID- 10695655 TI - The effects of somatostatin and octreotide on experimental and human acute pancreatitis. AB - The role of somatostatin and octreotide for AP has been studied for two decades, yet the data still remain inconclusive. The inconsistencies of the results of experimental studies and clinical trials may stem from the fact that the optimal therapeutic modality has not been determined. Furthermore, although they are similar in structure and physiologic activities, the mechanisms of action and effects of somatostatin and octreotide in AP may be different. Because the data are sparse, most reports, primarily those in the English literature, on the efficacy of somatostatin and octreotide in the management of AP were reviewed. Included are both nonrandomized and prospective, double-blind, clinical trials and studies on the effects of these agents on various experimental models of the disease. The results of the studies on somatostatin and octreotide are presented and discussed separately, with specific reference to the experimental and treatment details. The main focus of the review is the effect of subcutaneous and intravenous administration of octreotide. Analysis of the data suggests that somatostatin could not be recommended for AP and that the efficacy of subcutaneous administration of octreotide is also questionable. Theoretically, intravenous octreotide may be more appropriate for this condition, but recent results with this therapeutic method are limited and contradictory. Studies that would delineate the optimal therapeutical modality and the patient population most likely to respond to the treatment are prerequisite for large-scale clinical trials on the effects of octreotide on human pancreatitis. PMID- 10695656 TI - Gene therapy for chronic granulomatous disease. AB - Recent progress in the development of gene therapy for chronic granulomatous disease (CGD), an inherited immunodeficiency syndrome, is reviewed. This disorder results from defects in any of the four genes encoding essential subunits of respiratory burst oxidase, the superoxide-generating enzyme complex in phagocytic leukocytes. The absence of respiratory burst oxidants results in recurrent bacterial and fungal infections and can also be complicated by the formation of inflammatory granulomas. Although current management, including prophylactic use of antimicrobial agents and interferon-gamma, has significantly improved its prognosis, CGD continues to be associated with significant morbidity and mortality from life-threatening infections and complications. Allogeneic bone marrow transplantation can provide a life-long cure of the disease, but difficulty in finding suitable donors and risks associated with this procedure have limited its application. Recently CGD has emerged as a promising candidate for gene therapy targeted at the hematopoietic system. CGD mouse models have been developed with gene targeting technology, and preclinical studies in these animals with recombinant retroviral vectors have demonstrated the appearance of functionally normal neutrophils and increased resistance against pathogens such as Aspergillus. Although the murine studies have provided a promise of long-term cure of patients by gene transfer, phase I clinical studies in a limited number of patients with CGD with such vectors have yet to produce a clinically relevant number of corrected neutrophils for extended time periods. Efforts are ongoing to improve gene transfer efficiency into human hematopoietic stem/progenitor cells and to achieve better engraftment of the gene-corrected stem cells. PMID- 10695657 TI - Cytokines as targets of immunotherapy in bacterial pneumonia. AB - The generation of a vigorous inflammatory response is essential for the rapid clearance of microbes from the alveolar space. The magnitude of the inflammatory response is tightly controlled by host-derived cytokines, which mediate lung inflammation by serving as leukocyte chemoattractants, leukocyte activating factors, or afferent signals in the induction or regulation of other effector molecules. In this article the role of specific cytokines in lung innate immunity will be reviewed. Future directions regarding the use of specific forms of immunotherapy, including compartmentalized cytokine delivery with gene therapy as adjuvant therapy in the treatment of pneumonia, will be explored. PMID- 10695658 TI - Relationship between Sjogren's syndrome and human T-lymphotropic virus type I infection: follow-up study of 83 patients. AB - We have previously demonstrated a high prevalence of Sjogren's syndrome (SS) in patients with human T-lymphotropic virus type I-associated myelopathy (HAM) in Nagasaki prefecture. The present follow-up study compared the clinical and laboratory findings of SS with or without human T-lymphotropic virus type I (HTLV I) antibody in this endemic area for HTLV-I infection. We investigated the clinical and laboratory manifestations in 83 patients with SS and HAM, including histologic examination of labial salivary glands and the prevalence of SS in patients with HAM. Definite SS was diagnosed in 13 out of 20 patients with HAM when the European Community criteria were used. The density of mononuclear cell infiltration in labial salivary glands was higher in HTLV-I-seropositive patients with SS (including patients with HAM) than in HTLV-I-seronegative patients. The volume of saliva and lacrima determined by the Schirmer or Saxon test was lower than normal but was not different among SS-HTLV-I-seronegative patients, HTLV-I seropositive patients without HAM, and HTLV-I-seropositive patients with HAM. The proportions of patients positive for antinuclear antibody (ANA) and anti-SS-A (Ro) antibody or anti-SS-B (La) antibody were similar in the three groups. However, the low volume of saliva and the frequency of ANA in SS correlated with the degree of mononuclear cell infiltration in labial salivary glands. Our results suggested that HTLV-I infection is related to SS and that laboratory and clinical findings in SS closely correlate with the degree of mononuclear cell infiltration in the salivary glands. PMID- 10695659 TI - Autonomic modulation of heart rate and blood pressure in normotensive offspring of hypertensive subjects. AB - Predominant sympathetic cardiovascular modulation in the hyperkinetic phase of arterial hypertension has been well described. Less information is available on autonomic control in persons with a family history of arterial hypertension. To investigate this question, we selected 61 normotensive subjects (mean age 30.9 +/ 1.8 years) whose mother or father or both had arterial hypertension and 30 normotensive patients (mean age 30.1 +/- 1.4 years) whose parents had not had arterial hypertension (neither mother nor father) to undergo short-term power spectral analysis of RR interval and arterial pressure variabilities. The same recordings were used to determine baroreflex sensitivity or the alpha index by means of the transfer function. Normotensive offspring of hypertensive subjects had higher diastolic blood pressures (P < .05) and left ventricular mass index (P < .05) than did normotensive offspring of non-hypertensive subjects. They also had higher spectral densities of low frequency expressed in normalized units, both for R-R intervals (P < .05) and systolic pressure variabilities (P < .05); they also had a greater ratio of low-frequency to high-frequency powers of R-R interval variability (P < .05). No difference was observed between the two normotensive groups for baroreflex sensitivity. Our spectral data indicate that normotensive persons with a positive family history of arterial hypertension have lower parasympathetic modulation than those with a negative history. In normotensive persons with a family history of arterial hypertension, normal baroreflex sensitivity could be the mechanism that buffers the tendency for pressures to increase. The gradual loss of this regulatory mechanism may favor rising arterial pressures. PMID- 10695660 TI - Decreased response to recall antigens is associated with depressed costimulatory receptor expression in septic critically ill patients. AB - Anti-inflammatory substances are released during septic shock that modulate monocyte function. Decreased monocyte responsiveness to bacterial toxins and decreased expression of human-leukocyte-associated antigen-DR (HLA-DR) have been reported during septic shock and critical illness. Impaired antigen presentation has been inferred from these observations but has not been demonstrated. We assessed antigen presentation and costimulatory molecule expression in 12 age matched control subjects, 10 noninfected critically ill patients (CINS), and 17 critically ill patients with sepsis (CIS). Antigen presentation was assessed by using in vitro lymphocyte 5-bromo-2-deoxyuridine (BrdU) incorporation in response to tetanus toxoid. The expression of HLA-DR and the costimulatory molecules CD28, CD86, and CTLA-4 was assessed by flow cytometry. Serum interleukin-10 (IL-10) was also measured by enzyme-linked immunosorbent assay. Serum IL-10 levels were significantly elevated in CIS patients (91 +/- 38 pg/mL) as compared with levels in control subjects (5 +/- 4 pg/mL)(P < .05). Lymphocyte BrdU incorporation increased by 710% +/- 243% in control subjects but by only 144% +/- 62% in CIS patients and 76% +/- 31% in CINS patients (P < .01 vs control). Monocyte HLA-DR expression, monocyte CD86 expression, and lymphocyte CD28 expression were significantly decreased in CIS patients (P < .01) as compared with control subjects. Conversely, lymphocyte CTLA-4 expression was significantly increased in CIS patients (P < .05 vs control). Monocyte CD86 expression was also significantly decreased in CINS patients as compared with control subjects. These data indicate that antigen presentation is decreased in critically ill patients with sepsis. This appears in part related to decreased expression of HLA-DR and the costimulatory molecules CD86 and CD28. Increased expression of the negative signal receptor CTLA-4 may also impair antigen presentation in patients with sepsis. PMID- 10695661 TI - Inhibition of MIP-1alpha-induced human neutrophil and monocyte chemotactic activity by reactive oxygen and nitrogen metabolites. AB - Peroxynitrite, formed by the reaction between nitric oxide (NO) and superoxide, has been implicated in the pathogenesis of numerous disease processes. Several studies have shown that peroxynitrite-induced protein nitration may compromise enzyme and protein function. We hypothesized that peroxynitrite may regulate cytokine function during inflammation. To test this hypothesis, the neutrophil and monocyte chemotactic responses of macrophage inflammatory protein-1alpha (MIP 1alpha) incubated with and without peroxynitrite were evaluated. Peroxynitrite attenuated neutrophil chemotactic activity (NCA) and monocyte chemotactic activity (MCA) by MIP-1alpha in a dose-dependent manner (P < .05). The inhibitory effects were not significant on NCA and MCA induced by leukotriene B4 or complement-activated serum incubated with peroxynitrite. The reducing agents deferoxamine, dithiothreitol, and exogenous L-tyrosine abrogated the NCA and MCA inhibition by peroxynitrite. Papa-NONOate, an NO donor, or a combination of xanthine and xanthine oxidase to generate superoxide, did not show an inhibitory effect on NCA and MCA induced by MIP-1alpha. In contrast, 3-morpholinosydnonimine (SIN-1), a peroxynitrite generator, elicited a concentration-dependent reduction in NCA and MCA induced by MIP-1alpha. Consistent with its capacity to reduce NCA and MCA, peroxynitrite treatment reduced MIP-1alpha binding to neutrophils and monocytes. Nitrotyrosine was detected in the MIP-1alpha incubated with peroxynitrite. These findings are consistent with nitration of tyrosine by peroxynitrite with subsequent inhibition of MIP-1alpha binding to neutrophils and monocytes and a reduction in NCA and MCA. These data demonstrate that peroxynitrite modulates the inflammatory cell migration by MIP-1alpha, and they suggest that oxidants may play an important role in the regulation of MIP-1alpha induced inflammatory cell chemotaxis. PMID- 10695662 TI - Prevalence of HFE (hemochromatosis gene) mutations in unselected male patients with type 2 diabetes. AB - To assess the prevalence of mutations in the HFE (hemochromatosis) gene in unselected male patients with type 2 diabetes, we examined 220 white men without known diabetes and 220 age-matched white men with type 2 diabetes for mutations in the HFE gene. Nucleotide 845 (C282Y) and 187(H63D) alleles were amplified by polymerase chain reaction (PCR) with lymphocyte DNA. The PCR products were analyzed by restriction enzyme digestion. One of the 220 patients (0.45%) with diabetes was homozygous for the HFE 845A (C282Y) mutation and 25 (11.3%) were heterozygous for the same mutation, of whom 3 (1.3%) were compound heterozygotes also carrying the HFE 187G (H63D) mutation. These frequencies did not differ significantly from the control population without diabetes. There is no evidence that HFE mutations are found in excess in unselected male patients with type 2 diabetes, and there is no indication for a population-based search for an excess of these alleles in type 2 diabetes. PMID- 10695663 TI - 3-Hydroxy-3-methylglutaryl-coenzyme A reductase activity is inhibited by cholesterol and up-regulated by sitosterol in sitosterolemic fibroblasts. AB - Sitosterolemia is an inherited recessive disease characterized by abnormally increased plasma and tissue plant sterol concentrations. Patients hyperabsorb sitosterol. In addition, hepatic, ileal, and mononuclear leukocyte 3-hydroxy-3 methylglutaryl coenzyme A (HMG-CoA) reductase, the rate-controlling enzyme in the cholesterol biosynthetic pathway, is markedly suppressed in this disease. It is still controversial whether the down-regulation is due to accumulated sitosterol, but the effect of sitosterol on HMG-CoA reductase activity has not been studied in sitosterolemic tissues. To investigate whether sitosterol inhibits HMG-CoA reductase activity in sitosterolemia, we measured the enzyme activities in liver and cultured skin flbroblasts from patients. Hepatic HMG-CoA reductase activities in patients were decreased 76% (P < .05) as compared with results in control subjects. In contrast, HMG-CoA reductase activities in sitosterolemic fibroblasts were not decreased as compared with results in control fibroblasts, and the activities in all cells were up-regulated similarly when they were exposed to delipidated medium. Because the cultured sitosterolemic fibroblasts contained only trace amounts of plant sterols, we added 20 microg/mL sitosterol directly to the cell medium. Raising the intracellular sitosterol concentration to 7% of cellular cholesterol level increased HMG-CoA reductase activity 23% (P < .05), while the addition of the same amount of cholesterol to the cells reduced the activity 46% (P < .05). Thus, when sitosterolemic skin fibroblasts were used, it was possible to distinguish between the effects of cholesterol and those of sitosterol on the activity of HMG-CoA reductase. These results suggest that reduced HMG-CoA reductase activity in this disease is caused by secondary effects of unknown regulator(s) other than sitosterol. PMID- 10695664 TI - Pharmacodynamic characterization of hemoglobin-induced vasoactivity in isolated rat thoracic aorta. AB - The origin and mechanism of vasocontraction observed after vascular exposure to acellular Hbs remain controversial. To help resolve the underlying mechanism, we characterized Hb-induced vasoactivities in terms of Hb purity, heme iron oxidation state, and ligand and pharmacodynamic properties. Isolated rat thoracic aortic rings with intact endothelium were suspended in oxygenated Krebs buffer, and isometric tension responses to various test Hb preparations were measured. In norepinephrine tone-enhanced aortic rings, both crude and purified Hbs exhibited similar dose-response characteristics; stroma-free Hb and HbA0, two Hb preparations with disparate purity, were equally potent in inducing vessel ring contraction. Purified Hb preparations significantly attenuated vasodilatory potency of both acetylcholine, an endothelium-dependent NO generator, and glyceryl trinitrate, an endothelium-independent NO generator. With the exception of nitrosylated Hb, ferrous Hbs, oxy Hb, and carbon monoxy Hb elicited contraction, whereas ferric derivatives, met Hb, and cyanomet Hb did not. In addition, NEM-Hb, an Hb with blocked cysteine residues, did not notably attenuate Hb vasoactivity. These results indicate that Hb itself is directly responsible for inducing contraction in the rat thoracic aortic rings. A primary mechanism for the Hb-induced vasoactivity appears to be heme iron inactivation of endothelium-derived NO. Nonheme interaction with endothelial NO does not appear to play a prominent role in this vascular model. In conclusion, Hb elicits dose dependent contraction in isolated rat thoracic aorta with intact endothelium. Vasoactivity of Hbs, however, could greatly vary with heme iron oxidation state, nature of heme ligand, and model vessels used in the evaluation. PMID- 10695665 TI - Inflammation-induced systemic proteolysis of inter-alpha-inhibitor in plasma from patients with sepsis. AB - Inter-alpha-inhibitor (IalphaI) is a human plasma serine proteinase inhibitor. It contains one light peptide chain called bikunin that exerts antiproteinase activity and other antiinflammatory functions. Bikunin is covalently linked to two heavy chains that, after tissular diffusion, stabilize the extracellular matrix. Owing to its negative acute-phase reactant character and its susceptibility to proteolysis, IalphaI has been implicated in the pathophysiology of sepsis. Moreover, IalphaI has been shown to exert a protective effect on a pig model of endotoxic shock. Twenty patients admitted to the intensive care unit (ICU) for a septic syndrome were included in the present study. IalphaI and antithrombin III (ATIII) levels were measured on admission. Sequential measurements of IalphaI could be done in 4 patients. We demonstrate that IalphaI levels are significantly decreased in plasma samples collected on admission from patients with sepsis (59 +/- 32 mg/L vs 241 +/- 70 mg/L; P < .0001). This decrease was greater in severe sepsis and septic shock than in sepsis. Death was not predictable from initiol IalphaI levels. In 2 patients with a favorable course, IalphaI values regularly increased during the ICU stay. By sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by immunoblot analysis and microsequencing, we characterized IalphaI-related components in plasma from several patients; they obviously arise from IalphaI through proteolytic cleavage. Thus, systemic proteolysis and decreased biosynthesis both contribute to the fall in the plasma level of IalphaI. Because IalphaI is very sensitive to proteolysis by polymorphonuclear granulocytes (PMNs) that are stimulated during sepsis, we suggest that IalphaI plasma level would be a useful marker for neutrophil proteinase activity. ATIII, as well as IalphaI, is considered a negative acute phase protein. Because in vitro ATIII is less susceptible than IalphaI to proteolysis by PMNs and because their relative levels weakly correlated, we suggest that an unspecific systemic proteolysis is not significantly involved in the ATIII deficiency occurring in sepsis. PMID- 10695666 TI - Competitive reverse transcription-polymerase chain reaction assay for quantification of human multidrug resistance 1 (MDR1) gene expression in fresh leukemic cells. AB - We have analyzed MDR1 gene expression in 69 clinical samples obtained from 64 patients with leukemic hematologic malignancies by using a competitive reverse transcription-polymerase chain reaction assay with a heterologous competitor RNA. To exclude a false-positive result caused by concomitant normal lymphocytes that physiologically express MDR1, in samples we determined a cut-off value of 8 amol MDR1 transcript per microgram of RNA by simultaneous measurement of rhodamine 123 dye efflux either in lymphocyte or gated leukemic cell populations. Consequently, 23 of 69 samples were concluded to be MDR1-positive in leukemic cells per se. The MDR1 expression rate was significantly correlated with factors such as a history of preceding chemotherapy, elder age of the patient, and certain disease types (eg, leukemia progressed from myelodysplastic syndrome). Moreover, the complete response rate after chemotherapy was significantly higher in MDR1-negative patients than in MDR1-positive patients (52% vs 17%, respectively; P = .01). The assay established will enable the quantification of MDR1 gene expression in blood samples from patients with leukemic hematologic malignancies and will be applicable to clinical laboratories as a routine test. PMID- 10695667 TI - Glucose and lactate concentration determination on single microsamples by Fourier transform infrared spectroscopy. AB - This study is the first to assess the analytic potential of Fourier-transform infrared (FT-IR) spectroscopy in determining exercise-induced metabolic changes, such as glucose and lactate serum concentrations, with single 50 microL blood microsamples. One-hundred ninety-eight capillary blood samples were taken at rest (rest serum) and after rowing exercises at different intensities (exercise serum) to obtain a wide range of lactate concentrations. A quantitative method is described with FT-IR spectroscopy involving only dilution and dessiccation of serum samples. Within serum spectra, an absorption band was strongly specific of glucose (1033 cm(-1)) that allowed the determination of its concentration (r = 0.97; P < .001 with reference values). Once we had substrated measured glucose absorption in serum spectra, one other absorption band seemed to be specific for lactate (1127 cm(-1)), which allowed the determination of the concentration of this metabolite (r = 0.96; P < .001 with reference values). The convenience of a capillary blood sampling with the strong accuracy of FT-IR measurements is of particular interest for medicinal and biologic concerns. PMID- 10695668 TI - Karl Landsteiner: blood groups foreseen? PMID- 10695669 TI - TMC-86A, B and TMC-96, new proteasome inhibitors from Streptomyces sp. TC 1084 and Saccharothrix sp. TC 1094. I. Taxonomy, fermentation, isolation, and biological activities. AB - TMC-86A, B and TMC-96, new 20S proteasome inhibitors with an epoxy-beta aminoketone moiety, were isolated from the fermentation broth of Streptomyces sp. TC 1084 and Saccharothrix sp. TC 1094, respectively. TMC-86A, B and TMC-96 inhibited the chymotrypsin-like and peptidylglutamyl-peptide hydrolyzing activities of 20S proteasome with the following IC50 values: TMC-86A, 5.1 microM and 3.7microM; TMC-86B, 1.1 microM and 31 microM; TMC-96, 2.9 microM and 3.5 microM, respectively. TMC-86A, B and TMC-96 exhibited the weak inhibitory activity against the trypsin-like activity of 20S proteasome with IC50 values of 51 microM, 250 microM, and 36 microM, respectively. They did not inhibit m calpain, cathepsin L, and trypsin at 100 microM, suggesting their high specificity for proteasome. Taxonomy of the producing strains is also described. PMID- 10695670 TI - Production of a family of kinase-inhibiting lactones from fungal fermentations. AB - During the course of our screening for natural products from fungi, extracts of several cultures were found to make a family of related resorcylic acid lactone compounds, which are potent inhibitors of MEK kinase. Comparative and empirical studies of fermentation conditions improved the titers of the compounds of interest. Striking changes in the ratios and amounts of the major and minor compounds in some cases were achieved by manipulations of media composition. PMID- 10695671 TI - Resorcylic acid lactones: naturally occurring potent and selective inhibitors of MEK. AB - A resorcylic acid lactone, L-783,277, isolated from a Phoma sp. (ATCC 74403) which came from the fruitbody of Helvella acetabulum, is a potent and specific inhibitor of MEK (Map kinase kinase). L-783,277 inhibits MEK with an IC50 value of 4 nM. It weakly inhibits Lck and is inactive against Raf, PKA and PKC. L 783,277 is an irreversible inhibitor of MEK and is competitive with respect to ATP. L-783,290, the trans-isomer of L-783,277, was isolated from the same culture and evaluated together with several semi-synthetic resorcylic acid lactone analogs. A preliminary structure-activity relationship is presented. Several independent cell-based assays have been carried out to study the biological activities of these resorcylic acid lactone compounds and a brief result summary from these studies is presented. PMID- 10695672 TI - Funalenone, a novel collagenase inhibitor produced by Aspergillus niger. AB - Funalenone, a phenalene compound that inhibits type I collagenase (MMP-1), was isolated from mycelium of Aspergillus niger FO-5904 by solvent extaction, ODS column chromatography, Sephadex LH-20 column chromatography and reversed phase HPLC. Funalenone inhibited 50% of type I collagenase activity at a concentration of 170 microM, but inhibited 18.3% and 38.7% against 72 kDa and 92 kDa type IV collagenase, respectively, at a concentration of 400 microM. PMID- 10695673 TI - Ferroverdins, inhibitors of cholesteryl ester transfer protein produced by Streptomyces sp. WK-5344. I. Production, isolation and biological properties. AB - Streptomyces sp. WK-5344, a soil isolate, was found to produce structurally related inhibitors of cholesteryl ester transfer protein (CETP). New active compounds, designated ferroverdins B and C, were isolated along with known ferroverdin A from the fermentation broth by solvent extraction, ODS column chromatography and silica gel column chromatography. All ferroverdins showed a dose-dependent inhibitory activity against human CETP. The IC50 values were 21, 0.62 and 2.2 microM for ferroverdins A, B and C, respectively, indicating that ferroverdin B is one of the most potent CETP inhibitors of microbial origin. PMID- 10695674 TI - Ferroverdins, inhibitors of cholesteryl ester transfer protein produced by Streptomyces sp. WK-5344. II. Structure elucidation. AB - The structures of ferroverdins B and C, novel inhibitors of cholesteryl ester transfer protein, were elucidated by spectroscopic studies including various NMR measurements. They are the complex between one Fe2+ and three ligands, that is, two common p-vinylphenyl-3-nitroso-4-hydroxybenzoates and one hydroxy p vinylphenyl-3-nitroso-4-hydroxybenzoate for ferroverdin B and one carboxylic acid p-vinylphenyl-3-nitroso-4-hydroxybenzoate for ferroverdin C. PMID- 10695675 TI - TMC-171A,B,C and TMC-154, novel polyketide antibiotics produced by Gliocladium sp. TC 1304 and TC 1282. AB - Four new antibiotics, TMC-171A (2), B (3), C (4) and TMC-154 (5) have been isolated from the fermentation of fungal strains Gliocladium sp. TC 1304 and TC 1282, respectively. Spectroscopic and degradation studies have shown that TMC 171s and TMC-154 were new members of the TMC-151 class of antibiotics, unique polyketides modified with a D-mannose and a D-mannitol or a D-arabitol. These compounds showed moderate cytotoxicity to various tumor cell lines. PMID- 10695676 TI - Biomolecular-chemical screening: a novel screening approach for the discovery of biologically active secondary metabolites. III. New DNA-binding metabolites. AB - Based on the chemical screening technique, biomolecular-chemical screening has been developed which makes use of two-dimensional TLC analysis of microbial extracts and combines thin-layer chromatography (RP-18) with binding studies towards DNA. In the first dimension the metabolites of the crude microbial extract are separated, and in the second dimension binding properties towards DNA are analysed. An initial screening program with 500 microbial extracts prepared by solid-phase extraction with XAD-16 resin resulted in 17 samples which contained metabolites with significant DNA-binding behavior. Fermentation, isolation and structural characterization led to already known metabolites [phenazine-1,6-dicarboxylate (1), phencomycin (2), 11-carboxy-menoxymycin B (3), soyasaponine I (4), and (8S)-3-(2-hydroxypropyl)-cyclohexanone (5)], as well as to new secondary metabolites. Fermentation of the producing organisms of the new DNA-binding metabolites, ent-8,8adihydro-ramulosin (6). (2R,4R)-4-hydroxy-2-(1,3 pentadienyl)-piperidine (7), (5R)-dihydro-5-pentyl-4'-methyl-4'-hydroxy-2(3H) furanone (8), and seco-4,23-hydroxyoleane-12-en-22-one-3-carboxylic acid (9), as well as isolation, structural characterization, and physico-chemical properties are reported. PMID- 10695677 TI - Presence of genes for beta-lactamases of two different classes on a single plasmid from a clinical isolate of Serratia marcescens. PMID- 10695678 TI - Synthesis and properties of the pyrrole analogs of chloramphenicol. PMID- 10695679 TI - Tylosin derivatives. V. Electrochemical opening of oxirane ring. PMID- 10695680 TI - The total synthesis and absolute structure of antifungal antibiotics(-)-PF1163A and B. PMID- 10695681 TI - Synthesis and biological properties of a novel cephalosporin FR86521 having potent activity against methicillin-resistant Staphylococcus aureus (MRSA) PMID- 10695682 TI - Medical refugees in America. PMID- 10695683 TI - Blood levels of homocysteine and increased risks of cardiovascular disease: causal or casual? AB - BACKGROUND: Accumulating data from epidemiological studies suggest that individuals with elevated blood levels of homocysteine have increased risks of cardiovascular disease. We reviewed the currently available evidence of an association between homocysteine and cardiovascular disease and examined whether the strength of the evidence varies according to study design. METHODS: We used a computerized MEDLINE literature search, 1966 through September 1998, to identify all epidemiological studies that examined the relationship of homocysteine level with risks of coronary heart or cerebrovascular disease. Two measures of plasma homocysteine level and its association with risk of cardiovascular disease were extracted: mean homocysteine level in cases and controls, and relative risk of cardiovascular disease for elevated homocysteine level. RESULTS: A total of 43 studies were reviewed. Most crosssectional and case-control studies indicated higher mean homocysteine levels (either fasting or after methionine load) and/or a greater frequency of elevated homocysteine level in persons with cardiovascular disease as compared with persons without cardiovascular disease. Results of most prospective studies, however, indicated smaller or no association. The few prospective studies that reported a positive association between homocysteine level and risks of cardiovascular disease included patients with preexisting vascular disease. CONCLUSIONS: In contrast to cross-sectional and case-control studies, results of prospective studies indicated less or no predictive ability for plasma homocysteine in cardiovascular disease. Instead, elevated homocysteine level may be an acute-phase reactant that is predominantly a marker of atherogenesis, or a consequence of other factors more closely linked to risks of cardiovascular disease. Randomized trials are necessary to test reliably whether lowering homocysteine levels will decrease risks of cardiovascular disease. PMID- 10695684 TI - Cytokine blockade as a new strategy to treat rheumatoid arthritis: inhibition of tumor necrosis factor. AB - Rheumatoid arthritis (RA) is a common, frequently severe, chronic inflammatory disease. Although the cause of RA remains unknown, recent advances in understanding its pathogenesis have been substantial. Despite the use of a variety of medications, particularly methotrexate, treatment of RA is not fully effective in most patients. Until recently, insights into inflammatory mechanisms in RA had not been successfully translated into novel classes of therapeutic agents. This gap now will likely be bridged in the form of a new strategy for treating RA-cytokine blockade. Although a variety of cytokines are important in the pathogenesis of RA, tumor necrosis factor (TNF) seems to play a pivotal role. Neutralizing TNF in patients with RA, by means of soluble TNF receptors or anti TNF monoclonal antibodies, has proven to be a powerful means of controlling disease activity. Studies are in progress to obtain additional information regarding long-term safety of TNF blockade and its effects on disease progression. PMID- 10695685 TI - Antipyretic therapy: physiologic rationale, diagnostic implications, and clinical consequences. AB - Various treatments have been used to suppress fever since antiquity. Surprisingly, few studies have been performed to ascertain the physiologic consequences of antipyresis and validate the rationale behind such therapy. More importantly, it has not been established conclusively that the benefits of antipyretic therapy outweigh its risks. The present review considers these issues in light of currently available data and formulates guidelines for antipyretic therapy based on these data. PMID- 10695686 TI - The lipid treatment assessment project (L-TAP): a multicenter survey to evaluate the percentages of dyslipidemic patients receiving lipid-lowering therapy and achieving low-density lipoprotein cholesterol goals. AB - OBJECTIVE: To determine the percentage of patients in the multicenter Lipid Treatment Assessment Project receiving lipid-lowering therapy who are achieving low-density lipoprotein cholesterol (LDL-C) goals as defined by National Cholesterol Education Program (NCEP) guidelines. METHODS: Adult patients with dyslipidemia, who had been receiving the same lipid-lowering therapy for at least 3 months, were assessed at investigation sites. Lipid levels were determined once in each patient at the time of enrollment. The primary end point was the success rate, defined as the proportion of patients who achieved their LDL-C target level as specified by NCEP guidelines. RESULTS: A total of 4888 patients from 5 regions of the United States were studied. Of these, 23% had fewer than 2 risk factors for coronary heart disease (CHD) and no evidence of CHD (low-risk group), 47% had 2 or more risk factors and no evidence of CHD (high-risk group), and 30% had established CHD. Overall, only 38% of patients achieved NCEP-specified LDL-C target levels; success rates were 68% among low-risk patients, 37% among high risk patents, and 18% among patients with CHD. Drug therapy was significantly (P< or =.001) more effective than nondrug therapy in all patient risk groups. However, many patients treated with lipid-lowering drugs did not achieve LDL-C target levels. CONCLUSIONS: Large proportions of dyslipidemic patients receiving lipid-lowering therapy are not achieving NCEP LDL-C target levels. These findings indicate that more aggressive treatment of dyslipidemia is needed to attain goals established by NCEP guidelines. PMID- 10695687 TI - Oral anticoagulation treatment in the elderly: a nested, prospective, case control study. AB - BACKGROUND: Whether elderly patients are at increased risk of complications during oral anticoagulant treatment (OAT) is still a matter of debate. METHOD: Bleeding and thrombotic events occurring during OAT in 461 patients, aged 75 years or older when they started OAT, and in 461 patients younger than 70 years, matched for sex, OAT indication, and treating center, were examined in a prospective, multicenter, inception-cohort study. RESULTS: Bleeding rate was 9.9% and 6.6% patient-years in elderly and young patients, respectively (P = .07), and 2.1% and 1.1% for major bleeding (P = .19); 6 and 1 events, respectively, were fatal (all intracranial, relative risk, 6.4; P = .05). In the elderly, bleeding rate was lower (4.5%) for international normalized ratios (INRs) between 2.0 and 2.9; it was higher during the first 90 treatment days (P = .05) and when arterial vascular disease was the indication for OAT (P = .03). Thrombosis rate was 4.2% and 2.5% patient-years in elderly and young patients, respectively (P = .10); however, 13 and 5 events were fatal (relative risk, 2.8; P = .04). Thrombosis rate was lower (1.5%) for INRs between 2.0 and 2.9; it was higher during the first 90 treatment days (P<.001) and 6 of 7 venous events occurred at lower than 2.0 INRs. CONCLUSIONS: A nonsignificant trend was noted toward a higher rate of both bleeding and thrombotic complications in elderly vs matched younger patients. Intracranial bleeding and fatal thrombotic events were significantly more frequent in the elderly. Our results also indicate that lower than 2.0 INRs do not preclude bleeding in the elderly nor offer adequate protection from thrombotic events. Moderate anticoagulation (2.0-3.0 INRs) in elderly patients seems the safest and most effective. PMID- 10695688 TI - Implications of a health lifestyle and medication analysis for improving hypertension control. AB - BACKGROUND: National Health and Nutritional Examination surveys have documented poor rates of hypertension treatment and control, leading to preventable morbidity and mortality. OBJECTIVES: To examine covariation in the medication and health lifestyle beliefs and behaviors of persons with hypertension to identify and profile distinct subgroups of patients. METHODS: A sample of 727 patients with hypertension, weighted to match the 1992 National Health Interview Survey age and sex distribution of patients with hypertension, was interviewed by telephone about their beliefs and behaviors regarding hypertension and its management. Cluster analysis of key variables was used to identify 4 patient types. RESULTS: Subgroups differed significantly. Group A members use an effective mix of medication and health lifestyle regimens to control blood pressure. Group B members are most likely to depend on medication and have high adherence rates. Yet they also have high rates of smoking (29%) and alcohol use (average, 104 times per year) and are less likely to exercise regularly. Group C members are most likely to forget to take medication, are likely to be obese, and find it most difficult to comply with lifestyle changes (except for very low rates of smoking and alcohol use). Group D members are least likely to take medication, most likely to change or stop medication without consulting their physician (20%), most likely to smoke (40%), and least likely to control diet (29%). Group A and B members have better health outcomes than group C and D members. CONCLUSIONS: Optimal management strategies are likely to differ for the 4 patient types. Further research should be conducted to validate these findings on a separate sample and to devise and test tailored management algorithms for hypertension compliance and control. PMID- 10695689 TI - Relation of low body mass to death and stroke in the systolic hypertension in the elderly program. The SHEP Cooperative Research Group. AB - BACKGROUND: There are scant data on the effect of body mass index (BMI) (calculated as weight in kilograms divided by the square of height in meters) on cardiovascular events and death in older patients with hypertension. OBJECTIVE: To determine if low body mass in older patients with hypertension confers an increased risk of death or stroke. PATIENTS: Participants were 3975 men and women (mean age, 71 years) enrolled in 17 US centers in the Systolic Hypertension in the Elderly Program trial, a randomized, double-blind, placebo-controlled clinical trial of lowdose antihypertensive therapy, with follow-up for 5 years. MAIN OUTCOME MEASURES: Five-year adjusted mortality and stroke rates from Cox proportional hazards analyses. RESULTS: There was no statistically significant relation of death or stroke with BMI in the placebo group (P = .47), and there was a U- or J-shaped relation in the treatment group. The J-shaped relation of death with BMI in the treated group (P = .03) showed that the lowest probability of death for men was associated with a BMI of 26.0 and for women with a BMI of 29.6; the curve was quite flat for women across a wide range of BMIs. For stroke, men and women did not differ, and the BMI nadir for both sexes combined was 29, with risk increasing steeply at BMIs below 24. Those in active treatment, however, had lower death and stroke rates compared with those taking placebo. CONCLUSIONS: Among older patients with hypertension, a wide range of BMIs was associated with a similar risk of death and stroke; a low BMI was associated with increased risk. Lean, older patients with hypertension in treatment should be monitored carefully for additional risk factors. PMID- 10695690 TI - Recent experience with Pseudomonas aeruginosa bacteremia in patients with cancer: Retrospective analysis of 245 episodes. AB - BACKGROUND: Pseudomonas aeruginosa bacteremia is a serious and possibly fatal condition in patients with cancer. OBJECTIVES: To ascertain the frequency, demographics, and predisposing factors for P. aeruginosa bacteremia in patients with cancer and to determine the efficacy of various therapeutic regimens. SUBJECTS AND METHODS: Patient records of the Clinical Microbiology Laboratory, The University of Texas, M. D. Anderson Cancer Center, Houston, were reviewed. From January 1, 1991, through December 31, 1995, 245 eligible cases of P. aeruginosa bacteremia were identified. We examined the patient records for the underlying malignant neoplasm and its management, symptoms and signs of infection, culture results of appropriate specimens, antibiotic therapy, and outcome. We also compared our present experience with a previous analysis from this institution covering the period from January 1, 1972, to December 31, 1981. RESULTS: The incidence of P. aeruginosa bacteremia has decreased compared with the previous study (2.8 vs 4.7 cases per 1000 admissions). It was most common in patients with acute leukemia (55 of 1000 registrations), and the frequency in this disease has not changed. Half of the patients were not in the hospital when they developed their infection. The overall cure rate was 80%, which was a significant (P<.001) increase compared with the 62% cure rate in the previous study. In this study, no significant difference in the cure rates was observed between monotherapy with a beta-lactam and combination therapy overall (P = .72), and in patients with shock (P = 1.0) and those with pneumonia (P = .60). The patients' initial neutrophil counts were not of prognostic value; however, the cure rate depended on subsequent changes in neutrophil count during therapy. CONCLUSIONS: The frequency rate of P. aeruginosa bacteremia has decreased in patients with solid tumors but has remained unchanged in patients with acute leukemia. Antibiotic regimens for empirical therapy of neutropenic patients and especially patients with acute leukemia should still provide coverage against P. aeruginosa. PMID- 10695691 TI - Using clinical evaluation and lung scan to rule out suspected pulmonary embolism: Is it a valid option in patients with normal results of lower-limb venous compression ultrasonography? AB - BACKGROUND: In patients with a low clinical probability of pulmonary embolism (PE) and a nondiagnostic lung scan, the prevalence of PE is theoretically very low. We assessed the safety and usefulness of this association for ruling out PE. METHODS: We analyzed data from 2 consecutive cohort management studies performed in 2 university hospitals (Geneva University Hospital, Geneva, Switzerland, and Hospital Saint-Luc, Montreal, Quebec), which enrolled 1034 consecutive patients who came to the emergency department with clinically suspected PE. All patients were submitted to a sequential diagnostic protocol of lung scan, D-dimer testing, lower-limb venous compression ultrasonography (US), and pulmonary angiography in case of inconclusive results of noninvasive workup. RESULTS: The prevalence of PE was 27.6%. Empirical assessment was accurate for identifying patients with a low likelihood of PE (8.2% prevalence of PE in the low clinical probability category). One hundred eighty patients had a low clinical probability of PE and a nondiagnostic lung scan. Among these patients, US showed deep vein thrombosis in 5. Hence, PE could be ruled out by a low clinical probability, a nondiagnostic lung scan, and a normal US in 175 patients (21.5%). The 3-month thromboembolic risk in these patients was low (1.7%; 95% confidence interval, 0.4%-4.9%). CONCLUSIONS: Anticoagulant treatment could be safely withheld in patients with a low clinical probability of PE and a nondiagnostic lung scan, provided that the US is normal. This combination of findings avoided pulmonary angiography in 21.5% of patients with suspected PE in this series. PMID- 10695692 TI - Upper gastrointestinal tract safety profile of alendronate: the fracture intervention trial. AB - OBJECTIVES: To determine whether alendronate sodium treatment is associated with upper gastrointestinal (GI) tract adverse experiences (AEs)-particularly those of the stomach, duodenum, or esophagus-in the Fracture Intervention Trial, and to assess the relationship between alendronate use and upper GI tract events among women at increased risk for these outcomes. DESIGN: Randomized, double-blind, placebo-controlled trial with a mean follow-up of 3.8 years. Women were initially randomized to receive alendronate sodium, 5 mg/d, or placebo. After 2 years, the alendronate sodium dose was increased to 10 mg/d. PARTICIPANTS: A total of 6459 women aged 54 to 81 years recruited from 11 US clinical centers. All participants had low hip bone mineral density. Women with major upper GI tract disease (recent ulcers, upper GI tract bleeding, or use of daily medication for dyspepsia) were excluded. Regular nonsteroidal anti-inflammatory drug users were not excluded. MEASUREMENTS: Self-reported upper GI tract AEs were ascertained by interview every 3 months. Serious upper GI tract AEs were confirmed and classified by review of hospital records and endoscopy reports, if available. Upper GI tract AEs were further analyzed in 2 specified groups-gastroduodenal and esophageal-to examine events that might be related to upper GI tract mucosal irritation. Gastric and duodenal perforations, ulcers, and bleeding events were combined for analysis of these clinically important outcomes. RESULTS: The overall incidence of upper GI tract events was similar in the alendronate and placebo groups (47.5% vs. 46.2%; relative risk [RR], 1.02; 95% confidence interval [CI], 0.95-1.10). The incidence of gastroduodenal perforations, ulcers, and bleeding events was 1.6% in the alendronate group and 1.9% in the placebo group (RR, 0.86; 95% CI, 0.59-1.24). The incidence of nonspecific upper GI tract conditions, such as abdominal pain, dyspepsia, nausea, and vomiting, was also similar in the 2 groups. Esophageal events occurred in 10.0% and 9.4% of patients in the alendronate and placebo groups, respectively (RR, 1.06; 95% CI, 0.91-1.24). Esophagitis not reported as reflux was more common in the alendronate group (0.7%) than in the placebo group (0.4%), but not significantly so (RR, 1.71; 95% CI, 0.90-3.39). Alendronate use was not associated with a significant increase in upper GI tract events among women at increased risk for these events (those aged > or =75 years with previous upper GI tract disease or using nonsteroidal anti inflammatory drugs). CONCLUSION: In these older women, upper GI tract complaints, particularly dyspepsia and abdominal pain, were common, but alendronate treatment was not associated with an increased incidence of upper GI tract events, even in high-risk subgroups. PMID- 10695693 TI - The Colorado thyroid disease prevalence study. AB - CONTEXT: The prevalence of abnormal thyroid function in the United States and the significance of thyroid dysfunction remain controversial. Systemic effects of abnormal thyroid function have not been fully delineated, particularly in cases of mild thyroid failure. Also, the relationship between traditional hypothyroid symptoms and biochemical thyroid function is unclear. OBJECTIVE: To determine the prevalence of abnormal thyroid function and the relationship between (1) abnormal thyroid function and lipid levels and (2) abnormal thyroid function and symptoms using modern and sensitive thyroid tests. DESIGN: Cross-sectional study. PARTICIPANTS: Participants in a statewide health fair in Colorado, 1995 (N = 25 862). MAIN OUTCOME MEASURES: Serum thyrotropin (thyroid-stimulating hormone [TSH]) and total thyroxine (T4) concentrations, serum lipid levels, and responses to a hypothyroid symptoms questionnaire. RESULTS: The prevalence of elevated TSH levels (normal range, 0.3-5.1 mIU/L) in this population was 9.5%, and the prevalence of decreased TSH levels was 2.2%. Forty percent of patients taking thyroid medications had abnormal TSH levels. Lipid levels increased in a graded fashion as thyroid function declined. Also, the mean total cholesterol and low density lipoprotein cholesterol levels of subjects with TSH values between 5.1 and 10 mIU/L were significantly greater than the corresponding mean lipid levels in euthyroid subjects. Symptoms were reported more often in hypothyroid vs euthyroid individuals, but individual symptom sensitivities were low. CONCLUSIONS: The prevalence of abnormal biochemical thyroid function reported here is substantial and confirms previous reports in smaller populations. Among patients taking thyroid medication, only 60% were within the normal range of TSH. Modest elevations of TSH corresponded to changes in lipid levels that may affect cardiovascular health. Individual symptoms were not very sensitive, but patients who report multiple thyroid symptoms warrant serum thyroid testing. These results confirm that thyroid dysfunction is common, may often go undetected, and may be associated with adverse health outcomes that can be avoided by serum TSH measurement. PMID- 10695694 TI - Preventive intervention to reduce sexually transmitted infections: a field trial in the Royal Thai Army. AB - BACKGROUND: During 1991 through 1993, sexually transmitted infections among conscripts in the Royal Thai Army in the upper-northern provinces were common: human immunodeficiency virus (HIV) prevalence at induction was 12%, HIV incidence was 2.4% per year, and incidence of sexually transmitted diseases was 17% per year. We evaluated a behavioral intervention to reduce incident sexually transmitted infections among conscripts inducted into the Thai Army in 1993. METHODS: We developed a preventive intervention that addressed consistent condom use, reducing alcohol consumption and brothel patronage, and improving sexual negotiation and condom skills. Companies were assigned to 1 of 3 groups matched on military mission: 450 men were in the intervention group, 681 were in barracks at the same base but did not receive the intervention (diffusion group), and 414 were in distant camps (controls). Baseline HIV serological testing and behavioral interviews were conducted during basic training in 1993. The intervention was applied for 15 months, and men were followed up at 6-month intervals (with repeated HIV serological testing, sexually transmitted disease assessments, and behavioral interviews) through May 1995. RESULTS: Incident sexually transmitted diseases were 7 times less frequent among men assigned to the intervention than the combined controls (relative risk, 0.15; 95% confidence interval, 0.04-0.55), after adjusting for baseline risk factors (P<.005). There was no diffusion of the intervention to adjacent barracks. The intervention decreased incident HIV by 50% in the intervention group. CONCLUSION: Intensive interventions in structured institutions can successfully reduce risk in settings confronting expanding heterosexual HIV epidemics. PMID- 10695695 TI - Outcomes of intensive care for patients with human immunodeficiency virus infection. AB - BACKGROUND: Intensive care for patients with human immunodeficiency virus is common, costly, and associated with high morbidity. Accurate and up-to-date outcome and prognostic data are needed to effectively counsel patients and to make difficult decisions regarding admission to the intensive care unit. METHODS: We reviewed the medical charts of 394 adults infected with human immunodeficiency virus who received intensive care at San Francisco General Hospital, San Francisco, Calif, from 1992 to 1995, and we performed a multivariate analysis to learn which factors were predictive of poor outcomes. RESULTS: Respiratory failure (47%), sepsis (12%), and neurologic disease (11%) were the most common indications for admission to the intensive care unit. Overall, 63% of the patients survived hospitalization; survival rates were 27%, 18%, 13%, and 11% at 1, 2, 3, and 4 years, respectively. Independent predictors of hospital mortality were low serum albumin level, Acute Physiology Score, mechanical ventilation, and a diagnosis of Pneumocystis carinii pneumonia during admission to the intensive care unit. Low CD4+ cell count, low serum albumin level, and mechanical ventilation predicted poor long-term survival. Of the 121 patients who had a CD4+ cell count less than 50 cells/microL (0.05x10(9)/L) and a serum albumin level less than 25 g/L and required mechanical ventilation, 7% survived for 2.5 years or more after hospital discharge. CONCLUSIONS: In this series, which is the largest to date of patients admitted to the intensive care unit with human immunodeficiency virus infection, we found that long-term survival rates were low. However, even among patients who had multiple risk factors for mortality, a substantial minority survived, with a few patients achieving long-term survival. PMID- 10695696 TI - From profound hypokalemia to life-threatening hyperkalemia: a case of barium sulfide poisoning. AB - We describe a 25-year-old man who was brought to the emergency department with skeletal muscle weakness, respiratory arrest, and rhabdomyolysis, as well as life threatening hyperkalemia, after ingesting a depilatory containing barium sulfide (Magic Shave; Carson Products Co, Savannah, Ga). The findings of his physical examination were significant for hyporeflexia with marked weakness. He was in respiratory distress and required intubation and ventilatory support owing to progressive weakness of the respiratory muscles. His serum potassium level was 1.5 mmol/L. He was treated with intravenous and oral potassium. His serum potassium level peaked at 8.3 mmol/L and his serum creatine kinase level at 8286 IU/L. His acute respiratory weakness resolved with correction of the potassium concentration; his rhabdomyolysis responded well to hydration; and his renal function returned to normal. We also discuss the various pathophysiological findings in this case and compare our patient with another who, despite ingesting a similar amount of the same hair remover, did not develop any of the above complications. PMID- 10695697 TI - The hospitalist movement. PMID- 10695698 TI - Evidence-based medicine: resident preferences for morning report. PMID- 10695699 TI - Celecoxib-induced acute pancreatitis and hepatitis: a case report. PMID- 10695700 TI - A symposium: lipid management summit on improving practice outcome. Introduction. PMID- 10695701 TI - Cholesterol management in the era of managed care. AB - Several large controlled clinical trials have documented that cholesterol lowering causes a marked reduction in major coronary events in patients with established coronary heart disease. Cholesterol lowering thus joins other proven therapies for risk reduction in secondary prevention. The need to include cholesterol-lowering therapy in secondary prevention has been endorsed as a new practice measure in the Health Plan Employer Data Information Set. This endorsement ensures that managed care will get behind the effort to better control cholesterol in patients with coronary heart disease. The next issue is whether managed care will support cholesterol-lowering therapy in primary prevention patients. The patients at highest risk for developing coronary heart disease are those with noncoronary forms of atherosclerotic disease, type 2 diabetes, multiple risk factors, and risk factors plus evidence of advanced subclinical atherosclerosis. Such patients can be said to have coronary heart disease risk equivalents. These patients should be good candidates for aggressive cholesterol management. A strong case can be made for managed-care support for this approach. Support for treatment of patients at lower risk may be open to some question, but the current guidelines of the National Cholesterol Education Program provide a strong rationale for cholesterol management for primary prevention in the medical setting. PMID- 10695702 TI - Rationale and design of the Cardiac Hospitalization Atherosclerosis Management Program at the University of California Los Angeles. AB - Despite clear and consistent clinical-trial evidence that secondary-prevention medical therapies reduce mortality in patients with established coronary artery disease, these therapies are underutilized in patients receiving conventional care. To address this issue, a Cardiac Hospitalization Atherosclerosis Management Program (CHAMP) focused on initiation of aspirin, cholesterol-lowering medication (3-hydroxy-3-methylglutaryl-coenzyme A [HMG-CoA] reductase inhibitor titrated to achieve low-density lipoprotein [LDL] cholesterol < 100 mg/dL), beta-blocker, and angiotensin-converting enzyme (ACE)-inhibitor therapy in conjunction with diet and exercise counseling before hospital discharge in patients with established coronary artery disease was designed and implemented at the University of California Los Angeles (UCLA) Medical Center starting in 1994. This treatment program was based on the hypothesis that initiation of therapy in the hospital setting would result in higher utilization rates both at the time of discharge and during longer-term follow-up. Implementation of this program involved the use of a focused treatment guideline, standardized admission orders, educational lectures by local thought leaders, and tracking/reporting of treatment rates. To assess the impact of the program, treatment rates and clinical outcome were compared in patients discharged in the 2-year periods before and after CHAMP was implemented. Hospital-based treatment protocols such as CHAMP have the potential to significantly increase treatment utilization of therapies previously demonstrated to improve survival and thus substantially improve the outcome of the 2 million patients diagnosed and hospitalized each year with coronary artery disease. PMID- 10695703 TI - Lipid management in a private cardiology practice (the Midwest Heart experience). AB - Emerging evidence that lowering cholesterol levels reduces the incidence of coronary heart disease led Midwest Heart Specialists to establish a lipid clinic in 1985. The physician-directed, nurse-managed program was developed to improve patient adherence to National Cholesterol Education Program (NCEP) guidelines and provide high-quality preventive services while preserving productivity for the busy interventional cardiologist who was the medical director of the program. From 1996 to 1997, Midwest Heart Specialists was one of 140 medical practices to participate in a national Quality Assurance Program (QAP). The purpose of the project was to evaluate the degree of treatment of hyperlipidemia in patients with coronary heart disease. Although the overall physician practice results were significantly better than the national averages, the lipid-clinic results were dramatically more impressive: 100% of the lipid-clinic patients were on lipid lowering therapy and 97% of the patients had a low-density lipoprotein (LDL) cholesterol level documented on the chart. Of these, 71% met their LDL goal, as compared with only 11% nationally. The results of the QAP within the physician practice stimulated the development of a new practice-wide lipid-management system. This new systematic approach to lipid management has improved overall lipid outcomes dramatically. The success of the Cholesterol Management Program has enhanced the reputation of the practice in the community, tied high-risk patients and their families to the practice, and improved the marketability of a full complement of cardiovascular services via Midwest Heart Specialists. PMID- 10695704 TI - Secondary prevention in a cardiology group practice and hospital setting after a heart-care initiative. AB - The American Heart Association (AHA) Consensus Panel Statement for Preventing Heart Attack and Death in Patients with Coronary Disease provides recommendations for the secondary prevention of heart disease in at-risk patients. Blackstone Cardiology Associates of Pawtucket, Rhode Island, undertook an initiative in their practice implementing secondary-prevention guidelines in patients with coronary artery disease. This retrospective study evaluates practice patterns for the management of hyperlipidemia for a cardiology group in an ambulatory and hospital setting after the institution of a physician-supervised, nurse-based disease management program. Practice patterns in patients with established coronary heart disease treated in a lipid center compared with non-lipid-center settings were evaluated. Parameters evaluated included documenting low-density lipoprotein (LDL) cholesterol, presence of lipid-lowering therapy, and achieving the National Cholesterol Education Program II (NCEP II) goal of LDL-cholesterol levels < or =100 mg/dL in patients with preexisting coronary artery disease. A total of 352 patients met inclusion criteria in the lipid-center setting and were compared with 289 non-lipid-center consecutively chosen patients. Age and gender differences were also evaluated. Inpatient medical records from a 254-bed Brown University-affiliated teaching hospital were also evaluated for lipid profile, achievement of NCEP II goal, and use of lipid-lowering medication on admission and discharge. The most recent LDL-cholesterol values of patients followed in the lipid-center and in the non-lipid-center setting of the Blackstone Cardiology Associates were compared. Blackstone Cardiology Associates consists of 4 cardiologists and 4 advanced-practice nurses. Achievement of LDL-cholesterol goal was higher in both the lipid-center and non-lipid-center settings compared with baseline. A smaller percentage of patients at goal in the lipid setting is likely due to referral bias resulting in patients with more difficult-to-manage mixed dyslipidemias and behavior-management issues ending up in the lipid center. There were no apparent sex differences at goal, and more elderly (age > or =65 years) achieved goal in the lipid clinic center. In the non-lipid-center setting, more males were at goal and had a lower mean LDL-cholesterol level. PMID- 10695705 TI - Strategies for implementing lipid-lowering therapy: pharmacy-based approach. AB - A multidisciplinary program was designed to improve patient outcomes after an acute coronary event. The primary objective of the program was that lipid lowering therapy be prescribed at the time of discharge for acute myocardial infarction (AMI) and percutaneous transluminal coronary angioplasty (PTCA) patients. Secondary objectives for this program were (1) a baseline lipid panel within the first 24 hours of admission and (2) documentation of discharge counseling for lipid-lowering therapy in the patient medical record. Improvements were reported for all 3 objectives. For the primary indicator, lipid-lowering therapy prescribed at discharge, the baseline value increased from 40% to 72-81%. The percentage of patients with a lipid panel within 24 hours of admission improved from a baseline of 13% to 38-71%. Overall, 28-77% of patient records contained documentation of lipid-lowering medication counseling after initiation of the program. This information should provide the necessary benchmarking data to maintain competitiveness in the dynamic healthcare environment. Overall, this program provides high-quality, cost-effective health care for the patient with established coronary artery disease. PMID- 10695706 TI - Optimizing treatment of dyslipidemia in patients with coronary artery disease in the managed-care environment (the Rocky Mountain Kaiser Permanente experience). AB - Rocky Mountain Kaiser Permanente has taken aggressive steps to ensure optimal treatment of all modifiable cardiac risk factors, especially low-density lipoprotein (LDL) cholesterol, in patients with coronary artery disease. In this article, we are presenting (1) the basic rationale for our approach, (2) the critical steps translating philosophy into practice, and (3) justification for all health plans to pursue a similar course. The continuum of physician-directed disease management systems that have evolved in our region-one administered by cardiology nurses in the perihospitalization period and the other by pharmacists in the long-term, outpatient setting-is then detailed. Although the relatively short duration that our comprehensive systems have been in place precludes any assessment of their impact on cardiac death, coronary artery disease events, or coronary artery disease procedure rates, the improvements in intermediate surrogate outcomes are promising. Virtually all surveyed patients participating in our management systems have been "very" or "extremely" satisfied with their experience. The LDL-cholesterol screening rate in the approximately 2,500 participants in the programs to date has reached 97%. Of these patients, 84% have LDL cholesterol <130 mg/dL and 48% <100 mg/dL, and only 15% of those few with LDL cholesterol >130 mg/dL (2.5% overall) are currently not receiving lipid-lowering therapy. The proportions of patients on aspirin/antiplatelet and beta-blocker therapy after myocardial infarction are 97% and 92%, respectively. The lipid screening and treatment rates, especially, represent significant improvement from our own baseline, and compare favorably with outcomes from other practice settings. In conclusion, health maintenance organizations have tremendous incentive and the unique opportunity and ability to develop systems to better manage large numbers of individuals with coronary artery disease. PMID- 10695707 TI - Improving patient outcomes by pooling resources (the Texas Heart Care Partnership experience). AB - The morbidity and mortality associated with cardiovascular disease presents an enormous humanistic and economic burden in the United States. In Texas, cardiovascular disease has been the leading cause of death since 1950. Risk factor modification has been targeted in the secondary prevention of cardiovascular disease, including lipid management, smoking cessation, improved control of blood pressure, physical activity, weight management, the use of antiplatelet agents/anticoagulants, angiotensin-converting enzyme (ACE) inhibitors in congestive heart failure, beta blockers after myocardial infarction, and estrogen replacement therapy. The Heart Care Partnership (HCP) is a multifaceted interactive program designed to improve risk-factor management in the secondary prevention of cardiovascular disease through physician education, participation, and consensus development in addition to practice improvement processes and patient education. Development and implementation of the Texas HCP was a joint effort of the Texas Medical Association, the Texas Affiliate of the American Heart Association, and Merck & Co. This program helps hospitals improve the quality of care and outcomes for patients with heart disease. Program resources include educational workshops, quality improvement processes, and patient educational materials. HCP workshops address the treatment gap, define optimal care, and help define institution-specific plans for treating heart disease. Quality-improvement processes provide hospitals with baseline data and tools to improve and measure outcomes over time. The HCP workshops are provided as a combination of lectures, interactive discussions, and small group planning sessions designed to encourage audience participation. Upon completing the HCP program, participants are able to (1) describe the evidence-based medicine supporting secondary prevention of cardiovascular disease; (2) identify and prioritize cardiovascular disease risk factors for secondary prevention; (3) identify barriers to and solutions for implementing secondary prevention; and (4) develop site-based plans for cardiovascular risk-factor modification with definite time lines for implementation ("care maps"). The HCP's initial audit of medical practices indicates that Texas appears to share the same deficiencies in the secondary prevention of cardiovascular disease as the rest of the country. However, improvements can be demonstrated in both the hospital and physician office settings through the HCP. The HCP facilitated the cooperation of the medical community in the state of Texas to work together in a synchronized, communicative manner to decrease coronary events. This partnership represents a watershed event in the history of Texas medicine. It is the first time that such a statewide team approach to address a public health issue has been initiated. In the past, medical organizations within the state have had disparate goals and multiple strategies for achieving them. PMID- 10695708 TI - Quality assessment and lipid management: considerations for computer databases for tracking patients. AB - Improving the quality of lipid management requires an objective assessment of current practice and the ability to monitor whether quality is improved by implementing changes in practice. In a competitive healthcare environment, documentation of quality of care and patient outcomes may be important in securing contracts. It would be almost impossible to perform a meaningful clinical-outcome analysis in a timely fashion without the support of a computerized database. However, evaluating, selecting, and implementing computerized databases can be a daunting task. Before the purchase of a database, the following steps should be performed: (1) consider and prioritize the goals for the computerized database; (2) audit charts to determine whether the existing chart format meets the current guidelines for reimbursement and medical-legal standards; (3) revise the paper chart to improve fulfillment of the goals from step 1; (4) consider the specific clinical environment, including the skill level of personnel using the system, how user-friendly the system is, whether the system is multifunctional, and the costs associated with the software and implementation. We have evaluated 3 types of computerized databases and report their strengths and weaknesses; we also briefly discuss the electronic medical record. PMID- 10695709 TI - On reaching 100. PMID- 10695710 TI - The immunopathogenesis of Miller Fisher syndrome. AB - Over the past decade, remarkable progress has been made in our understanding of the pathogenesis of Miller Fisher syndrome (MFS), a clinical variant of Guillain Barre syndrome (GBS). MFS comprises the clinical triad of ataxia, areflexia and ophthalmoplegia. It is associated with acute-phase IgG antibodies to GQ1b and GT1a gangliosides in over 90% of cases which are highly disease specific. Like GBS, MFS is a post-infectious syndrome following diverse infections, but particular attention has been paid to its association with Campylobacter jejuni enteritis. Serostrains of C. jejuni isolated from infected patients bear ganglioside-like epitopes in their lipopolysaccharide core oligosaccharides, which elicit humoral immune responses exhibiting molecular mimicry with GQ1b/GT1a gangliosides. These antibodies are believed to be the principal cause of the syndrome and physiological studies aimed at proving this have focused on the motor-nerve terminal as a potential site of pathogenic action. This review describes these findings and formulates a pathogenesis model based on our current state of knowledge. PMID- 10695711 TI - Animal models of neuroimmune interactions in inflammatory diseases. AB - Animal models have been used successfully to study various aspects of neural immune interactions. Although different approaches carry certain advantages and disadvantages, current high sensitivity screening and manipulation methods coupled with molecular and genetic approaches can be successfully used to tease out the neural pathways that regulate inflammatory disease and the effects of immune molecules, such as interleukins, on neuronal function and pathology. Newer methodologies that measure gene expression of thousands of genes will in the future add to the ability to evaluate complex systems interactions in whole animal models. This review addresses the advantages and disadvantages of some of these approaches in the context of application to neural-immune interactions. PMID- 10695712 TI - Our shifting understanding of the role of nitric oxide in autoimmune encephalomyelitis: a review. AB - Nitric oxide was first described being produced in inflammatory cells involved in experimental autoimmune encephalomyelitis in 1992. Since then some 45 papers have appeared examining the role of NO in this central nervous system autoimmune inflammatory disease. Of the first 10 papers published all resulted in the interpretation that NO was a pathologic or "bad" molecule in the context of EAE. A few papers then began to appear suggesting that NO may not in fact always be a harmful molecule and by the end of 1997 early 1998, 22 papers suggested a destructive role for the molecule while three suggested it was protective. The past two years have seen a significant increase in reports supporting a protective mechanism for NO in EAE such that as of July 1999, 27 papers suggest a destructive and 15 a protective role for NO with a few uncommitted. This review sets out in a more or less chronological order the studies examining the role of NO in EAE and maps our changing understanding of the molecules role in this CNS inflammatory disease and by inference perhaps multiple sclerosis. PMID- 10695713 TI - The pathogenesis and modulation of the post-treatment reactive encephalopathy in a mouse model of Human African Trypanosomiasis. AB - Drug treatment of late-stage human African Trypanosomiasis (HAT) in which the central nervous system (CNS) is involved may be complicated by a severe post treatment reactive encephalopathy (PTRE) which can be fatal in up to 10% of cases. In order to understand the immunopathogenesis of this complication, an experimental mouse model has been developed that mirrors many of the pathological features of the PTRE in humans, and which allows various anti-inflammatory therapeutic regimes to be evaluated. Following the development of the PTRE in this model a number of cytokines are increased within the CNS including tumour necrosis factor (TNF) alpha, interleukins 1, 4 and 6, and macrophage inflammatory protein (MIP)-1. These cytokines appear at the same time as astrocyte activation which is an early event occurring before the development of the marked meningoencephalitic inflammatory response. The immunosuppressant drug azathioprine prevents but does not reduce the severity of an established PTRE and has a minimal effect on astrocyte activation. The ornithine decarboxylase inhibitor eflornithine prevents the induction, and ameliorates the severity, of the PTRE, and also reduces the degree of astrocyte activation. The Substance P antagonist RP-67,580 ameliorates the severity of an established PTRE, and also reduces astrocyte activation, indicating an important role of SP in the generation of the inflammatory response. Continued use of this mouse model should lead to further enhancement of our understanding of the pathogenesis of the PTRE and to improved drug regimes to prevent and/or treat it. PMID- 10695714 TI - Role of Fas--FasL interactions in the pathogenesis and regulation of autoimmune demyelinating disease. AB - Multiple sclerosis (MS) and experimental autoimmune encephalomyelitis (EAE) represent complex processes that lead to destruction of oligodendrocytes (ODCs) and myelin. T cells are integral to the development of these diseases, but whether T cell-mediated cytolytic mechanisms are involved in the destruction of MHC Class II-negative targets, such as oligodendroglia and myelin, in the CNS is unclear. The primary lytic mechanism employed by CD4+ T cells is Fas-dependent, but can be MHC-unrestricted. Thus, T cell-mediated Fas-FasL interactions could directly contribute to the pathology of EAE and MS. This review summarizes studies from our laboratory and others that implicate Fas-FasL interactions in both the pathogenesis and regulation of demyelinating diseases. PMID- 10695715 TI - Suppressor cells in demyelinating disease: a new paradigm for the new millennium. AB - This brief review highlights investigations conducted over the past three decades concerning the role of suppressor T cells in the regulation of experimental autoimmune encephalomyelitis (EAE). In addition, more recent studies are summarized which suggest that apoptosis of autoreactive T cells is also involved in the regulation of EAE. The possibility that natural killer (NK) cells mediate apoptosis is also considered. PMID- 10695716 TI - Use of peptide ligands to analyze the fine specificity of antibodies against asialo GM1. AB - We recently described clone 10, a monoclonal Fab fragment that binds to asialo GM1 (GA1), and three mutated Abs derived from it that also bind GA1 and have a three to four times increase in avidity. We selected a phage display linear heptapeptide library with these four Abs, and an IgM mAb, 156, which binds to GM1 and GD1b, but not to GA1. Peptides with the same motif, KL/VWQXXX, were selected by clones 10 and the two heavy chain mutants 227 and 109. In contrast, the light chain mutant L3 58 selected an entirely different peptide motif, TFGLQSL. Moreover, a different motif, K/SWTNL/MPP, was selected by mAb 156. Although mAbs clone 10 and its mutants 109, 227 and L3 58 all bind only to GA1, differences in their fine specificity were revealed by binding to peptide ligands. PMID- 10695717 TI - Cytokine, chemokine and chemokine receptor mRNA expression in different strains of normal mice: implications for establishment of a Th1/Th2 bias. AB - The resistance or susceptibility of inbred strains of mice to various pathogens and autoimmune diseases such as EAE has been linked to differences in the balance between cytokines associated with Th1- and Th2-type immune responses. Previous work from this laboratory on the mouse strain specific resistance to mouse adenovirus type I (MAV-1)-induced encephalopathy revealed subtle differences in the transcription rates of several immunologically important molecules that was evident prior to infection. In this study, we show striking differences in cytokine, chemokine and chemokine receptor mRNA expression in the spleens of normal, immunologically naive C57BL/6J, BALB/cJ and SJL/J mice. Messenger RNAs for interferon (IFN)-gamma and the chemokine IFN gamma inducible protein (IP)-10 were preferentially expressed in C57BL/6J spleens, whereas in BALB/cJ spleens mRNAs for lymphotoxin-beta, interferon-beta, transforming growth factor-beta, and the chemokine receptors CCR3 and CXCR4 predominated. A unique profile of chemokine receptors was found in spleens from normal SJL/J mice that correlated with the presence of polymorphisms within the CCR-3 gene. The patterns of gene expression fit well into the Th1/Th2 paradigm for C57BL/6J and BALB/cJ strains and suggest an important role for chemokines, as well as cytokines, in contributing to the genetic basis of the immune response. PMID- 10695718 TI - Pathogenesis of Guillain-Barre syndrome. AB - Recent neurophysiological and pathological studies have led to a reclassification of the diseases that underlie Guillain-Barre syndrome (GBS) into acute inflammatory demyelinating polyradiculoneuropathy (AIDP), acute motor and sensory axonal neuropathy (AMSAN) and acute motor axonal neuropathy (AMAN). The Fisher syndrome of ophthalmoplegia, ataxia and areflexia is the most striking of several related conditions. Significant antecedent events include Campylobacter jejuni (4 66%), cytomegalovirus (5-15%), Epstein-Barr virus (2-10%), and Mycoplasma pneumoniae (1-5%) infections. These infections are not uniquely associated with any clinical subtype but severe axonal degeneration is more common following C. jejuni and severe sensory impairment following cytomegalovirus. Strong evidence supports an important role for antibodies to gangliosides in pathogenesis. In particular antibodies to ganglioside GM1 are present in 14-50% of patients with GBS, and are more common in cases with severe axonal degeneration associated with any subtype. Antibodies to ganglioside GQ1b are very closely associated with Fisher syndrome, its formes frustes and related syndromes. Ganglioside-like epitopes exist in the bacterial wall of C. jejuni. Infection by this and other organisms triggers an antibody response in patients with GBS but not in those with uncomplicated enteritis. The development of GBS is likely to be a consequence of special properties of the infecting organism, since some strains such as Penner 0:19 and 0:41 are particularly associated with GBS but not with enteritis. It is also likely to be a consequence of the immunogenetic background of the patient since few patients develop GBS after infection even with one of these strains. Attempts to match the subtypes of GBS to the fine specificity of anti-ganglioside antibodies and to functional effects in experimental models continue but have not yet fully explained the pathogenesis. T cells are also involved in the pathogenesis of most or perhaps all forms of GBS. T cell responses to any of three myelin proteins, P2, PO and PMP22, are sufficient to induce experimental autoimmune neuritis. Activated T cells are present in the circulation in the acute stage, up-regulate matrix metalloproteinases, cross the blood-nerve barrier and encounter their cognate antigens. Identification of the specificity of these T cell responses is still at a preliminary stage. The invasion of intact myelin sheaths by activated macrophages is difficult to explain according to a purely T cell mediated mechanism. The different patterns of GBS are probably due to the diverse interplay between antibodies and T cells of differing specificities. PMID- 10695719 TI - Highly related immunoglobulin light chain sequences in different multiple sclerosis patients. AB - Although immunoglobulin G and free light (L) chains of oligoclonal origin in cerebrospinal fluid (CSF) are the most common immunologic abnormalities in multiple sclerosis (MS), it is unknown whether homologous CSF L chain sequences are present in different individuals with MS. Using Southern blotting, a particular kappa (kappa) L chain variable region (V) probe was recently found to hybridize to Vkappa cDNA from CSF B cells from almost one half of the MS patients tested but only 10% of normal or other neurologic disease controls [Zhou, S.-R., Maier, C.C., Mitchell, G.W., LaGanke, C.C., Blalock, J.E., Whitaker, J.N., 1998. A cross-reactive idiotope in cerebrospinal fluid cells in multiple sclerosis: further evidence for the role of myelin basic protein. Neurology 50, 411-417.] Here, we report that this likely results from remarkable sequence similarity in certain Vkappa from CSF B cells from different individuals with MS. The high degree of sequence homology even extended to all three complementarity determining regions (CDR) which in part form an antibody combining site. In addition, marked sequence homology was observed between the light chains from the MS patients and those from certain mouse antibodies against myelin basic protein (MBP). The results establish, in principle, that the same or very similar kappa light chain variable regions can be shared between CSF B lymphocytes from different individuals with MS as well as with certain antibodies against MBP. PMID- 10695720 TI - Dendritic cells in experimental allergic encephalomyelitis and multiple sclerosis. AB - The mechanisms by which autoimmune diseases are triggered and by which the activation of autoreactive T cells is initiated and maintained are not yet fully understood. As the most potent antigen presenting cells (APC), and also being responsible for antigen transport as well as primary sensitisation of T cells, dendritic cells (DC) are capable of breaking the state of self-ignorance and inducing aggressive autoreactive T cells. In the development of autoimmune diseases, different types of DC exhibit distinct properties for inducing Th1/Th2 cell responses. Appropriate cytokines can convert immunogenic DC to tolerogenic DC. Utilizing the possibility to promote the tolerogenic effects of DC, a new therapeutic tool might soon become available to treat multiple sclerosis and other autoimmune diseases. PMID- 10695721 TI - Autoimmune maintenance and neuroprotection of the central nervous system. AB - The genesis of immune privilege high in the evolutionary tree suggests that immune privilege is necessary, if not advantageous for the progressive development of the CNS. Upon reaching a certain degree of complexity, it seems as if the CNS was obliged to restrain the immune system from penetrating the blood brain barrier. CNS autoimmunity against myelin proteins is known to be a contributory factor in the pathophysiology of multiple sclerosis and in the animal model of experimental autoimmune encephalomyelitis (EAE) (Wekerle, 1993). Such autoimmunity has therefore been regarded as detrimental and hence obviously undesirable. However, recent findings in our laboratory suggest that T-cell autoimmunity to CNS self-antigens (Moalem et al., 1999), if expressed at the right time and the right place, can do much good in the CNS. We shall review the experiments briefly, and then discuss their implications for our understanding of immune privilege and CNS maintenance after injury. PMID- 10695722 TI - Preferential expansion of autoreactive T lymphocytes from the memory T-cell pool by IL-7. AB - We have developed a new technique that allows us to quantify antigen-specific T cells, and to determine their functional phenotype and origin from naive versus memory populations. Using this methodology, we have characterized a total of 286 T-cell lines specific for myelin basic protein (MBP) and influenza hemagglutinin from 16 multiple sclerosis (MS) patients and nine healthy donors. Our data support the notion that MBP-specific T cells undergo in vivo activation in MS patients and indicate a presence of immune dysregulation that renders MS patients prone to develop autoimmunity. Our methodology offers a way to study antigen specific T-cell characteristics as a surrogate marker in immunotherapy trials. PMID- 10695723 TI - Interleukin-6 expression and regulation in astrocytes. AB - The physiological function of interleukin-6 (IL-6) within the central nervous system (CNS) is complex; IL-6 exerts neurotrophic and neuroprotective effects, and yet can also function as a mediator of inflammation, demyelination, and astrogliosis, depending on the cellular context. In the normal brain, IL-6 levels remain low. However, elevated expression occurs in injury, infection, stroke, and inflammation. Given the diverse biological functions of IL-6 and its expression in numerous CNS conditions, it is critical to understand its regulation in the brain in order to control its expression and ultimately its effects. Accumulating data demonstrate that the predominant CNS source of IL-6 is the activated astrocyte. Furthermore, a wide range of factors have been demonstrated to be involved in IL-6 regulation by astrocytes. In this review, we summarize information concerning IL-6 regulation in astrocytes, focusing on the role of proinflammatory factors, neurotransmitters, and second messengers. PMID- 10695724 TI - IL-10 as a mediator in the HPA axis and brain. AB - Certain functional interactions between the nervous, endocrine, and immune systems are mediated by cytokines. The pro-inflammatory cytokines, interleukin-1 (IL-1) and tumor necrosis factor (TNF) were among the first to be recognized in this regard. A modulator of these cytokines, IL-10, has been shown to have a wide range of activities in the immune system; in this review, we describe its production and actions in the hypothalamic-pituitary-adrenal (HPA) axis. IL-10 is produced in pituitary, hypothalamic, and neural tissues in addition to lymphocytes. IL-10 enhances corticotropin releasing factor (CRF) and corticotropin (ACTH) production in hypothalamic and pituitary tissues, respectively. Further downstream in the HPA axis endogenous IL-10 has the potential to contribute to regulation of glucocorticosteroid production both tonically and following stressors. Our studies and those of others reviewed here indicate that IL-10 may be an important endogenous regulator in HPA axis activity and in CNS pathologies such as multiple sclerosis. Thus, in addition to its more widely recognized role in immunity, IL-10's neuroendocrine activities described here point to its role as an important regulator in communication between the immune and neuroendocrine systems. PMID- 10695725 TI - Autonomic regulation of experimental autoimmune encephalomyelitis in IL-4 knockout mice. AB - The effect of chemical sympathectomy induced with 6-hydroxydopamine (OHDA) on experimental autoimmune encephalomyelitis (EAE) was studied in wild type and IL-4 /- C57BL/6 (B6) mice. When actively sensitized with myelin oligodendrocyte glycoprotein (MOG)35-55 peptide, control B6 mice developed a mild form of EAE with full recovery. The sympathectomized mice developed paralysis with higher maximum disease score and did not recover completely, indicating that the sympathetic nervous system (SNS) down-modulates the process of EAE. Unexpectedly, however, sympathectomy resulted in suppression of EAE in IL-4-/- mice, implying that control of actively induced EAE by the SNS depends on the genetic background of mice. We also induced EAE by passive transfer of MOG35-55-reactive lymph node cells, and this disease was augmented by sympathectomy in both wild type and knockout animals. Further experiments showed that changes in T cell populations and the activity of antigen presenting cells might be responsible for the altered immune response and clinical course after sympathetic ablation. Our studies indicate that the absence of a single cytokine can severely alter nervous-immune system interactions. PMID- 10695726 TI - Peroxisome proliferator-activated receptor gamma agonists protect cerebellar granule cells from cytokine-induced apoptotic cell death by inhibition of inducible nitric oxide synthase. AB - Cerebellar granule cells (CGCs) can express the inducible isoform of nitric oxide synthase (iNOS) in response to inflammatory stimuli. We demonstrate that induction of iNOS in CGCs by bacterial lipopolysaccharide and pro-inflammatory cytokines results in cell death that was potentiated by excess L-arginine and inhibited by the selective iNOS inhibitor, 2-amino-5,6-dihydro-6-methyl-4H-1,3 thiazine. The NO-mediated cell death was accompanied by increased caspase-3-like activity, DNA fragmentation and positive terminal transferase dUTP nick end labeling (TUNEL), suggesting that apoptosis mediates CGC cell death. Incubation of CGCs with the non-steroidal anti-inflammatory drugs (NSAIDs), ibuprofen or indomethacin, or with 15-deoxy-delta12,14 prostaglandin J2 (PGJ2) downregulates iNOS expression and reduces subsequent cell death. Since in other cell types, both NSAIDs and PGJ2 can activate the peroxisome proliferator-activated receptor gamma (PPARgamma) and downregulate cytokine levels and iNOS expression, and since CGCs express PPARgamma in vivo and in vitro, our data suggest that activation of CGC PPARgamma mediates iNOS suppression and reduced cell death. Because PPARgamma is expressed in brains of Alzheimer's Disease (AD) patients, in which neuronal iNOS expression and apoptotic cell death have been described, these results may help explain the basis for the beneficial effects of NSAIDs in AD. PMID- 10695727 TI - Pathogenic autoantibodies to neuronal proteins in neurological disorders. AB - Autoantibodies to acetylcholine receptors and to voltage-gated calcium and potassium channels are thought to be pathogenic in three peripheral neurological disorders: myasthenia gravis, the Lambert Eaton syndrome and acquired neuromyotonia. However, evidence for the role of antibodies in conditions involving the central nervous system, is scanty or unclear. This review describes the ways in which the roles of autoantibodies have been defined in the peripheral diseases, and discusses the more controversial evidence for involvement of autoantibodies in some central disorders such as multiple sclerosis. PMID- 10695728 TI - Immune regulation and CNS autoimmune disease. AB - The central nervous system is a demonstrated target of both clinical and experimental immune mediated disorders. Immune regulatory mechanisms operative at the levels of the systemic immune system, the blood brain barrier, and within the CNS parenchyma are important determinants of the intensity and duration of the tissue directed injury. Convergence of research, involving direct manipulation of specific cells and molecular mediators in animal models and in vitro analysis of human immune and neural cells and tissues, is providing increasing insight into the role of these immune regulatory functions and their potential to serve as therapeutic targets. PMID- 10695729 TI - IFN-beta modulates specific T cell responses in vitro but does not affect Experimental Autoimmune Encephalomyelitis in the SJL mouse. AB - In this study, mouse recombinant IFN-beta was shown to favor PLP139-151-specific Th2 responses in vitro, by inhibiting IFN-gamma production and stimulating IL-4 and IL-10 production. IFN-beta (5000 U/day) failed to prevent the development or severity of EAE induced with PLP139-151. Whereas efficacy of IL-10 was found in the B. pertussis assisted but not in the pertussigen-assisted EAE model, both models appeared insensitive to IFN-beta. Also the combination of (suboptimal) IL 10 and IFN-beta appeared ineffective in inhibiting disease. However, the PLP139 151-specific IL-10 production by T cells from these mice appeared significantly more sensitive to the stimulatory effect of IFN-beta in vitro. It is concluded that despite its Th2 promoting effects, IFN-beta is not effective in inhibiting EAE in this study. PMID- 10695730 TI - The importance of being receptive. AB - Neuroendocrine system and immune system can communicate via the use of soluble mediators like hormones, neurotransmitters and cytokines. The level of mediators secreted by either of these systems creates the milieu in which immune and neuroendocrine responses take place. For adequate communication between the systems, receptors for hormones, neurotransmitters and cytokines are required. This review describes the role of regulated expression and function of receptors for hormones and neurotransmitters within the immune system in neuroendocrine immune communication. PMID- 10695731 TI - Potential mechanisms of interleukin-1 involvement in cerebral ischaemia. AB - Interleukin-1 (IL-1) has pleiotropic actions in the central nervous system. During the last decade, a growing corpus of evidence has indicated an important role of this cytokine in the development of brain damage following cerebral ischaemia. The expression of IL-1 in the brain is dramatically increased during the early and chronic stage of infarction. The most direct evidence that IL-1 contributes significantly to ischaemic injury is that (1) central administration of IL-1beta exacerbates brain damage, and (2) injection or over-expression of interleukin-1 receptor antagonist, and blockade of interleukin-1beta converting enzyme activity reduce, dramatically, infarction and improve behavioural deficit. The mechanisms underlying IL-1 actions in stroke are not definitively elucidated, and it seems likely that its effects are mediated through stimulation and inhibition of wide range of pathophysiological processes. PMID- 10695732 TI - Immune defects observed in patients with primary malignant brain tumors. AB - Malignant glioblastomas (gliomas) account for approximately one third of all diagnosed brain tumors. Yet, a decade of research has made little progress in advancing the treatment of these tumors. In part this lack of progress is linked to the challenge of discovering how glial tumors are capable of both modulating host immune function and neutralizing immune-based therapies. Patients with gliomas exhibit a broad suppression of cell-mediated immunity. The impaired cell mediated immunity observed in patients with gliomas appears to result from immunosuppressive factor(s) secreted by the tumor. This article reviews what has been elucidated about the immune defects of patients harboring glioma and the glioma-derived factors which mediate this immunosuppression. A model involving systemic cytokine dysregulation is presented to suggest how the immune defects arise in these individuals. PMID- 10695733 TI - Neuregulin and erbB receptor expression in normal and diseased human white matter. AB - Human white matter from non-neurologic cases, multiple sclerosis (MS) and other neurologic diseases (OND, inflammatory and non-inflammatory), was subjected to immunocytochemistry and Western blotting for expression of the neuregulin, glial growth factor-2 (GGF2), and its receptors, erbB2, erbB3 and erbB4. GGF2 has previously been shown to have mitogenic effects upon oligodendrocytes in vitro and an enhancing effect upon remyelination in animals with autoimmune demyelination. In all types of human white matter examined, expression of the ligand GGF2 and its three receptors was consistently found on oligodendrocytes, with higher levels being seen in cases of MS. Expression was also seen, albeit at lower levels, on astrocytes and microglial cells, the latter most commonly in MS and OND. In human lymph node tissue, some lymphocytes were positive for erbB2, erbB3 and erbB4. Western blots confirmed the presence of all three receptors in normal, MS and OND white matter. GGF2 and erbB receptor expression did not correlate with areas of remyelination and reactivity occurred throughout the tissue, with some increase in intensity at the edge of MS lesions. Examination of precursor oligodendrocyte immunoreactivity (with anti-PDGF-Ralpha and NG2), revealed widespread expression throughout both normal and diseased white matter. The presence of GGF2 and its receptors on oligodendrocytes and lymphocytes render this cell type a candidate for functional signaling via this pathway, perhaps in relationship to myelinating activity. PMID- 10695734 TI - Neuroendocrine modulation of chronic relapsing experimental autoimmune encephalomyelitis: a critical role for the hypothalamic-pituitary-adrenal axis. AB - Murine relapsing EAE can be profoundly suppressed by restraint stress (RST) administered beginning prior to neuroantigen immunization. This study determined what hormone pathway(s) mediate disease suppression. Our results showed that nadolol (NAD), a beta2-adrenergic antagonist, did not reverse the RST-induced suppression of EAE. However, administration of either RU486 or aminoglutethimide, which block the action of peripheral glucocorticoids, resulted in a partial reversal of EAE suppression. Administration of exogenous corticosterone mimicked the effects of RST, in terms of suppression of EAE, decrease in lymphoid cell numbers and decrease in Thl cytokine production. Therefore, the HPA axis plays a more profound role in the RST-induced suppression of EAE than does the sympathetic nervous system. PMID- 10695735 TI - Molecular pathogenesis of multiple sclerosis. AB - Multiple sclerosis (MS) is best understood as an inflammatory disease of the central nervous system (CNS) white matter characterized by demyelination, focal T cell and macrophage infiltrates, axonal injury and loss of neurological function. Our current understanding invokes proinflammatory cells and mediators that may be triggered by environmental factors to mediate disease in a genetically susceptible host. Five major themes which have been associated with the pathogenesis of MS lesions will be discussed: (1) The differential activation states of myelin-reactive T cells from MS patients vs. normal individuals, (2) the selective expression of chemokines, adhesion molecules and matrix metalloproteinases, (3) the proposed roles of the B7 costimulatory pathway, (4) the proinflammatory cytokines and (5) the role of molecular mimicry. PMID- 10695736 TI - Evidence for gammadelta T cells with a restricted Vgamma6 junctional region in the normal mouse central nervous system. AB - In this study we present evidence that gammadelta T cells are present in the normal mouse central nervous system (CNS). Compared with matching spleen gammadelta T cells, CNS gammadelta T cells expressed only the CD45RBlow phenotype, suggesting that CNS gammadelta T cells belong to the memory cell population. Approximately 20% expressed exclusively the CD8alphabeta heterodimer, consistent with a thymic origin. Gammadelta T cells in both spleen and CNS expressed higher levels of the IL-2rbeta (CD122), as well as Fas and FasL, than alphabeta T cells, suggesting that these cells function as immunoregulatory T cells. RT-PCR analysis showed almost exclusive use of Vdelta6 in the CNS whereas more Vdelta genes were expressed in the spleen. Sequencing of Vdelta6 RT-PCR products demonstrated a polyclonal population of T cells in the spleen but a more clonal population within the CNS. The predominant CNS sequence was found in all animals studied and was also detected in the spleen. From these data we conclude that a selective component of circulating gammadelta T cells traffics through the CNS. Thus, all major populations of lymphocytes can be detected in the normal CNS and as such may play specific roles in the immunological surveillance of that organ. PMID- 10695737 TI - Infections within the peritoneal cavity: a historical perspective. AB - Physicians in antiquity dreaded abdominal infections. Despite the fact that peritonitis was extremely common, reports of successful surgical interventions were only anecdotal before the past century. Medicine's comprehension of the pathophysiology of the peritoneal cavity is still evolving. The history of our understanding of the process could be considered to be as recent as the current literature. Despite this, the mortality rates for patients with secondary peritonitis have fallen in the last century from almost 100 per cent to less than 10 per cent. PMID- 10695738 TI - Prophylactic antibiotics in surgery and surgical wound infections. AB - Wound infection remains a considerable cause of morbidity and mortality among surgical patients, despite the relative success of prophylactic antibiotics. In modern efforts to control healthcare costs while improving the quality of patient care, we must not overlook the basic principles of wound infections and their appropriate treatment. Predisposing factors for the development of surgical wound infection include the creation of a surgical wound, the presence of bacteria, and a susceptible host. The selection of an appropriate antimicrobial drug depends on the identification of the most likely pathogens associated with a given procedure, as well as the expected antibiotic susceptibility of those pathogens. Ideally, a prophylactic antibiotic should achieve high peak tissue concentration at the site of the wound before the first incision and should be maintained until the time of closure. Currently, the administration of prophylactic antibiotics is indicated for contaminated and clean-contaminated wounds. Despite the proven effectiveness of antibiotic prophylaxis, many researchers would argue that contemporary dosing regimens should be reevaluated. The debates concerning the dosage and timing of ideal prophylactic administration are likely to continue. PMID- 10695739 TI - Antibiotic-resistant organism infection. AB - Bacteria possess a remarkable number of ways to become resistant to antibiotics. Antibiotic resistance has become a major problem in the treatment of Gram positive infections. Resistance to methicillin and vancomycin in staphylococci and enterococci has resulted in organisms that are resistant to all known antibiotics. Although it is important to continue to search for newer and more effective antibiotics, it is imperative that we develop a surgical mindset of appropriate antibiotic stewardship. The use of single-dose prophylactic regimens, using narrow-spectrum agents when possible for therapeutic indications, limiting the duration of therapeutic agents appropriately, avoiding the use of vancomycin except when necessary, and adhering to strict infection control measures are all steps that will limit the spread and development of resistant organisms. PMID- 10695740 TI - Diagnosis and treatment of opportunistic infections in immunocompromised surgical patients. AB - The advent of successful therapy for patients who suffer many types of organ dysfunction and failure, malignancies, and acquired immunodeficiency syndrome has led to the concurrent threat of infection due to a wide array of pathogens, particularly opportunistic microbes that rarely cause disease under routine circumstances. Among patients who are subjected to extreme degrees of immunosuppression, almost any type of bacterial, fungal, viral, protozoal, or parasitic organism can exhibit pathogenic potential and lead to devastating consequences for the host. Immunosuppressive drug therapy for the purpose of organ allograft maintenance, cancer chemotherapy, or the human immunodeficiency virus exerts potent effects upon cellular immunity. Therefore, although these groups of patients are more susceptible to all types of infectious disease processes, infections due to those pathogens that require a component of cellular immunity for their eradication, such as fungi and viruses, occur at a higher frequency than that observed among normal individuals. Of critical importance, all types of infections are associated with higher rates of morbidity and mortality in immunosuppressed patients. Currently, improved diagnostic techniques and new treatment modalities have rendered many serious infections, for which suitable therapy previously did not exist, amenable to treatment. Because of the large number of immunosuppressed patients who now lead highly productive lives, it is important for the surgical practitioner to become familiar with the modalities currently available to precisely diagnose and effectively treat opportunistic infections in immunocompromised surgical patients. PMID- 10695741 TI - Sepsis syndrome. AB - A clinical syndrome including fever, leukocytosis, elevated cardiac output, and reduced systemic vascular resistance has been associated with severe infection (i.e., sepsis). However, during the last 15 years, many patients have demonstrated all of the findings that have traditionally been associated with "sepsis" but have not had demonstrated sources of infection. This led to the term "sepsis syndrome" to refer to that population of patients who appeared to have a physiologic and metabolic response associated with, but who did not have, severe infection. More commonly called the systemic inflammatory response syndrome (SIRS), the sepsis syndrome is now associated with the nonspecific systemic activation of the human inflammatory cascade by any of a number of clinical events. The management of the SIRS patient has been ineffective because of incomplete definition of the mechanisms responsible for the syndrome. It is argued that all of the biological mechanisms that are operative in a simple wound and are beneficial are negative for the host when activated systemically. Thus, SIRS is seen in three separate scenarios at present: (1) invasive infection; (2) dissemination of microbes secondary to failure of host defense mechanisms; and (3) severe activation of inflammation by injury, shock, severe soft tissue inflammation, and other noninfectious but proinflammatory events. Newer treatment strategies will need to focus not on the inciting event itself but on better control of the complex responses of the host. PMID- 10695742 TI - Postoperative pneumonia. AB - Despite the advances made in surgical critical care, the diagnosis of one of the most common infections seen in critically ill patients remains a challenge. Ventilator-associated pneumonia is associated with a 20 to 25 per cent mortality rate. There are numerous risk factors for ventilator-associated pneumonia, including underlying disease, prolonged mechanical ventilation, direct lung injury, and shock. The standard clinical criteria for pneumonia are inaccurate. Quantitative cultures of bronchoalveolar lavage effluent are accurate for the diagnosis, and it is safe to base antibiotic therapy on the results of the quantitative cultures. PMID- 10695743 TI - Surgical treatment of biliary tract infections. AB - Despite major advances in surgical and nonsurgical therapy, biliary tract infections remain a significant cause of morbidity and mortality. The two classic biliary tract infections most commonly encountered are acute cholecystitis (either calculous or acalculous) and acute cholangitis. In addition, bile leakage associated with bile duct injuries during laparoscopic cholecystectomy has become a problem not infrequently encountered by surgeons. Acute calculous cholecystitis results from a combination of mechanical, biochemical, and infectious mechanisms, initiated by stone impaction in the cystic duct. After instituting empiric antibiotics, early laparoscopic cholecystectomy should be performed. Although conversion to open cholecystectomy is more common than in chronic cholecystitis, there appears to be no increased morbidity or mortality in that setting. Acute acalculous cholecystitis usually occurs in critically ill patients and may present both a diagnostic and therapeutic dilemma. Aggressive management, however, is warranted, both because of the critical nature of illness in these patients and the high incidence of perforation. Percutaneous cholecystostomy is indicated, particularly in high-risk patients both for diagnosis and treatment. Acute cholangitis results from a combination of bactibilia and biliary obstruction. The majority of patients can be successfully managed with intravenous antibiotics and fluid resuscitation. In those patients in whom initial management is not successful, biliary drainage, which is best accomplished nonoperatively, should be instituted. There is a very limited role for early surgical intervention in acute suppurative cholangitis. Biliary leaks resulting in bile "peritonitis" or bilomas are common sequelae of laparoscopic bile duct injury. Although surgeons may feel it is necessary to operate urgently, delineation of the proximal biliary anatomy via percutaneous transhepatic cholangiography and biliary stent placement is the appropriate first step in management. This procedure will usually control the bile leak and allow delineation of the anatomy and opportune timing of definitive reconstruction. PMID- 10695744 TI - Acute pancreatitis: management of complicating infection. AB - Acute pancreatitis develops precipitously, changing the patient's condition from apparent good health to a critically ill status. Of patients who succumb, 80 per cent die from secondary infection in the pancreas-peripancreatic area. Infection supervenes in the second week or later after onset. Prophylactic antibiotic(s) appear to be helpful in avoiding, delaying, and/or lessening secondary sepsis. Once infection develops, treatment requires open debridement of necrotic material, drainage, and appropriate antibiotic therapy; or mortality will approach 100 per cent. Infecting organisms are commonly Escherichia coli, Klebsiella, Staphylococcus, Enterococcus, Bacteroides, and/or fungi. Antibiotics felt to be preferable for prophylactic therapy include 1) imipenem-cilastatin, 2) a quinolone + metronidazole, and 3) possibly an extended-spectrum penicillin. Treatment should be continued for 2 weeks or until recovery. Because fungus infections are occurring more often, prophylaxis with fluconazole may be warranted. PMID- 10695745 TI - Surgical management of diverticulitis. AB - Diverticular disease, and particularly diverticulitis, has an increasing incidence in Westernized countries because of low-fiber diet. Diverticular disease may be classified as asymptomatic, atypical, acute or uncomplicated, and complicated. Conservative or medical management is usually indicated for acute or uncomplicated diverticulitis, with elective surgical resection generally being recommended after two documented episodes. Complicated diverticulitis, because of the high rate of recurrent problems, is generally managed promptly with sigmoid resection. Sigmoid resection for diverticulitis, under appropriate circumstances, has one of the highest success rates of any of the common gastrointestinal procedures. PMID- 10695746 TI - Evaluation and management of tertiary peritonitis. AB - Tertiary or recurrent peritonitis can occur after any operation for secondary bacterial peritonitis. The major risk factors for the development of tertiary peritonitis include malnutrition, a high Acute Physiology and Chronic Health Evaluation II score, the presence of organisms resistant to antimicrobial therapy, and organ system failure. Most patients with tertiary peritonitis will have fever and leukocytosis, even though other signs of infection may be absent. The management of tertiary peritonitis should include the provision of appropriate physiologic support, the administration of antimicrobial therapy, and operation or intervention to control the source of contamination and to decrease the bacterial load. Antibiotic-resistant organisms and bacteremia are present more commonly and mortality is greater in patients with tertiary peritonitis. Early recognition and effective intervention are critical to achieving a successful outcome. PMID- 10695747 TI - Soft tissue infections. AB - Soft tissue infections are almost routinely the product of direct microbe inoculation through a bridged protective skin. Day of onset and clinical presentation reflect the causative pathogen(s) and course that should be taken in treatment. Exclusive of chronic states, only in the most fulminating cases are culture and antimicrobial drugs of any real value. PMID- 10695748 TI - Management of patients with prosthetic vascular graft infection. AB - Management of patients with infected prosthetic vascular grafts is one of the most difficult challenges faced by the vascular surgeon. Patients often present with nonspecific symptoms, but delay in treatment can lead to life-threatening sepsis and/or hemorrhage. Fortunately, prosthetic vascular graft infection is uncommon, with the incidence varying between 1 and 6 per cent, depending on the location of the graft. Initially, the potentially infected vascular graft should be imaged using either CT or magnetic resonance imaging, with radionuclide studies being reserved for those instances in which imaging studies do not confirm or exclude the diagnosis of infection. Current treatments for prosthetic vascular graft infection include attempted graft preservation, graft removal with in situ graft replacement (using autogenous or new prosthetic grafts), and graft removal with extra-anatomic bypass. Morbidity and mortality associated with treatment, likelihood of long-term limb salvage, and likelihood of persistent or recurrent infection vary among these types of treatment. Therefore, in an individual patient with a prosthetic vascular graft infection, many things must be considered to appropriately determine the treatment most likely to achieve eradication of the infection and long-term limb salvage with the lowest risk. Regardless, with appropriate application of the techniques currently available for treatment of prosthetic vascular graft infection, long-term elimination of infection and limb preservation can be achieved in the great majority of patients with this grave problem. PMID- 10695749 TI - Hepatitis: risks for the surgeon. AB - Six different hepatitis viruses have now been characterized. Hepatitis B and C are the two hepatitis infections that are of greatest concern for surgeons. Hepatitis B and C share several features that have led to this concern. Both are blood-borne infections. Both are associated with chronic infection ultimately leading to cirrhosis, portal hypertension, and hepatocellular carcinoma, and both can be occupational infections for the surgeon after percutaneous injury associated with infected blood. Chronic hepatitis B infection is seen in 1.25 million people in the U.S. It is associated with a transmission rate to healthcare workers of 25 to 30 per cent following a hollow needle stick injury. Five per cent of acute infections result in chronic disease. It can be effectively prevented as an occupational infection by vaccination with the highly effective hepatitis B vaccine. Chronic hepatitis C infection is present in nearly 4 million people in the U.S. It has a lower rate of transmission than hepatitis B following needle stick injury, but it has a 50 to 80 per cent rate of chronic disease after acute infections. There is no vaccine for hepatitis C, and only prevention of blood exposure will avoid the risks of this occupational infection. Other hepatitis viruses are likely to be identified. Prevention of blood exposure, by the better use of barriers in the operating room and modification of surgical techniques, is recommended to prevent occupational infection from both known and unknown blood-borne viruses from the surgical patient. PMID- 10695750 TI - Postoperative sterno-mediastinitis. AB - Postoperative sterno-mediastinitis is a life-threatening complication that occurs in about 0.75 to 1.4 per cent of all open heart operations. The result of treatment largely depends on timely diagnosis and appropriate surgical management. Risk factors for infection should be corrected preoperatively whenever possible. Among other preventive measures, meticulous asepsis, atraumatic surgical technique, preserving the blood supply and the mechanical integrity of the sternum, prevention of sternal instability, and correction of the same if it occurs are the most important. The management of sterno mediastinitis should be tailored to the individual clinical features of the patients. Clearly cases with nonpurulent sternomediastinitis and no soft tissue or bone necrosis (type 1) may be treated with reopening, drainage, sternal stabilization, and primary closure. Virulent infections with tissue necrosis (type II) may be best handled with reopening, several days of open management, and debridement then secondary closure with viable tissue (usually muscle) flaps. Chronic, smoldering infections (type III) are usually managed with debridement and muscle-flap coverage. PMID- 10695751 TI - Acute abdomen and lupus enteritis: thrombocytopenia and pneumatosis intestinalis as indicators for surgery. AB - Bowel symptoms occur often in systemic lupus erythematosus (SLE), but enteric complications in patients on steroid therapy are rare. We report a case of a 14 year-old Mexican girl with SLE on high-dose steroid therapy complicated by abdominal vasculitis and small bowel perforation. Accompanying this serious complication were thrombocytopenia and radiographic changes of pneumatosis intestinalis. These findings suggested necrotizing enteritis and prompted urgent surgery. Four jejunal perforations, pneumatosis intestinalis, and submucosal vasculitis were present in the resected specimen. Persistent SLE activity responded to cyclophosphamide, which is indicated in patients with digestive symptoms who fail to respond to high-dose steroids. PMID- 10695752 TI - Symmetrical peripheral gangrene: a new presentation of an old disease. AB - This report concerns two cases and a review of the literature on the subject of symmetrical peripheral gangrene. Symmetrical peripheral gangrene is defined as symmetrical distal ischemic damage in two or more sites in the absence of major vascular occlusive disease. It occurs in patients who are septic and have disseminated intravascular coagulation and in nonseptic patients who have cardiogenic or hypovolemic shock. The syndrome is devastating and rare, and controlled studies of its etiology and management are lacking. Recommendations are presented for its prevention and treatment. Cooperative multicenter studies may be necessary to obtain valid data about its prevention and management. PMID- 10695753 TI - Diverticulitis and polycystic kidney disease. AB - Patients with adult polycystic kidney disease (PKD) have previously been shown to have an increased incidence of complicated diverticulitis after renal transplantation. The purpose of this study was to assess the risk of diverticulitis in the PKD population. We retrospectively reviewed patients with advanced PKD, defined as end-stage renal failure requiring dialysis. Patients were obtained from a single nephrology group practice between January 1985 and January 1997, or from all patients being evaluated or actively considered for renal transplantation at our institution as of May 1997. The incidence and severity of diverticulitis in these patients was compared with that observed in a similar cohort of patients with end-stage renal disease due to other etiologies. The study population consisted of 184 renal failure patients, 59 with PKD and 125 with other causes of end-stage renal disease. Twelve (20%) patients with PKD had a history of active diverticulitis, whereas only 4 (3%) of the non-PKD controls had diverticulitis (P = 0.0003, Fisher's exact test). Six of the 12 PKD patients required surgical intervention. Patients with renal failure due to PKD experience a significantly higher rate of diverticulitis than do other patients with end stage renal disease. Furthermore, diverticulitis is frequently severe in PKD patients, with 50 per cent requiring surgical intervention. These data suggest that diverticular disease may be an extrarenal manifestation of polycystic kidney disease. PMID- 10695754 TI - Splenic abscess 10 years after splenic trauma: a case report. AB - Splenic abscess is an uncommon complication of splenic trauma. Splenic abscess presents within several months of the trauma. We report a case of a splenic abscess 10 years after trauma and review the current understanding of splenic abscesses. PMID- 10695755 TI - Experience with pneumonia in acutely burned patients requiring ventilator support. AB - Acutely burned patients requiring ventilatory support who developed pneumonia while in the hospital were retrospectively reviewed. The Centers for Disease Control and Prevention (CDC) clinical criteria for pneumonia are based on clinical findings, radiographic findings, and culture data. During an 18-month period, 784 burn patients were admitted. Of these, 145 (18.5%) were placed on ventilators for at least 1 day. Fifty-three (36.6%) patients on ventilators developed acute pneumonia based on CDC criteria. Identification of causative organisms was based on positive cultures from blood or endotracheal aspiration within 3 days of the diagnosis of pneumonia. Thirty-nine patients were diagnosed as having inhalation injury. Forty-seven patients were placed on ventilators before or on the day of admission. Ages ranged from 2 to 82 years (mean, 39). Burn size ranged from 2 to 85 per cent (mean, 29.7%) of total body surface area. The total number of ventilator days was 1310 for the 53 patients, with a mean of 27.7 days. Ten patients had positive blood cultures during the period in which pneumonia was present. Thirty-one different organisms were recovered from blood or tracheal aspirates. The most commonly recovered organism was Pseudomonas aeruginosa. In 30 incidences, polymicrobial cultures were encountered. Initiation of appropriate antimicrobial therapy was begun on the basis of clinical impression and current burn unit experience and revised on the basis of the culture data. Of the 53 patients, 13 (25.5%) died, all while still on ventilators. The other 40 patients survived. Thirty-four were weaned off their ventilators, and 6 were transferred while still on ventilator support. PMID- 10695756 TI - Empyema and restrictive pleural processes after blunt trauma: an under-recognized cause of respiratory failure. AB - Respiratory failure is a common complication among patients sustaining major blunt trauma. This is usually due to the underlying pulmonary injury, pneumonia, or adult respiratory distress syndrome. However, we have frequently found these patients to actually have a pleural process as the cause of their respiratory failure. Our objective was to assess the frequency of empyema and restrictive pleural processes after blunt trauma and their contribution to respiratory failure. We retrospectively reviewed all blunt trauma patients over a 5-year period who required a thoracotomy and decortication for empyema. Twenty-eight patients with blunt trauma required a thoracotomy and decortication for empyema. The most common finding was infected, loculated hemothorax/effusion in 23 patients, whereas 5 had an associated pneumonia. Chest radiographs were nondiscriminating, whereas CT scans in 25 patients showed previously unrecognized fluid collections, air-fluid levels, or gas bubbles. Neither thoracentesis nor placement of additional chest tubes was helpful. Positive cultures were uncommon. Ventilator dependence was present preoperatively in 13 patients who were on the ventilator an average of 13 days preoperatively and only 5.8 days postoperatively. Several patients believed to have adult respiratory distress syndrome were weaned within 72 hours of operation. All patients were ultimately cured. Empyema is an under-recognized complication of blunt trauma and may contribute to respiratory failure and ventilator dependence. Although difficult to diagnose, empyema should be considered in blunt trauma patients with respiratory failure and an abnormal chest radiograph. CT aids in the diagnosis, and the results of surgical treatment are excellent. PMID- 10695757 TI - Abscess of an accessory spleen. AB - Accessory spleens are not infrequent and occur in 11 to 44 per cent of the population with a greater incidence in those with hematological disease. They may remain clinically silent or result in a number of pathologic processes. Abscess of an accessory spleen is rare but must be considered in the differential diagnosis of fever of unknown origin or sepsis in select groups of patients. Computerized tomography is the imaging modality of choice and may also be used in the percutaneous drainage of select cases. Laparoscopic splenectomy in the hands of the experienced laparoendoscopic surgeon is a viable treatment option. PMID- 10695758 TI - Intestinal perforation secondary to Salmonella typhi: case report and review of the literature. AB - The case of a young woman presenting with fever, abdominal distention, and diarrhea is presented. While hospitalized, she developed peritonitis, and a laparotomy was performed emergently. Intraoperative and pathologic examinations are highly suggestive of Salmonella typhi as an etiology for her symptoms and eventual perforation. Salmonella enteritis can be a difficult diagnosis to make, but in most cases it is a self-limited disease process. In a minority of cases, multidrug antibiotic therapy may be required secondary to an increasing prevalence of resistant strains. Patients who perforate require prompt operation to limit morbidity and mortality. Outcome is significantly improved in those patients by directed resection of the affected segment of bowel and by aggressive perioperative care. PMID- 10695759 TI - Temporal aspects of neural coding in the retina and lateral geniculate. AB - The early stages of visual processing provide excellent models for the study of how information is represented in, and processed by, the activity of neurons. The fact that the retina contains both non-spiking and spiking neurons leads us to frame questions about neural coding in a general fashion, rather than in a manner specific either to point processes or continuous signals. In particular, we ask about the role of the statistical structure of the response, the extent to which the neural representation is 'literal', and how information content can be estimated from laboratory data. The broad theme that emerges from a review of experimental data is that each stage of visual processing is accompanied by new features, including adaptive filtering, feedback, rectification and spike generation. These dynamical elements allow an increasingly rich set of strategies for the representation and processing of visual information at retinal and thalamic levels. PMID- 10695760 TI - Pre-synaptic lateral inhibition provides a better architecture for self organizing neural networks. AB - Unsupervised learning is an important ability of the brain and of many artificial neural networks. A large variety of unsupervised learning algorithms have been proposed. This paper takes a different approach in considering the architecture of the neural network rather than the learning algorithm. It is shown that a self organizing neural network architecture using pre-synaptic lateral inhibition enables a single learning algorithm to find distributed, local, and topological representations as appropriate to the structure of the input data received. It is argued that such an architecture not only has computational advantages but is a better model of cortical self-organization. PMID- 10695761 TI - Computational study of experience-dependent plasticity in adult rat cortical barrel-column. AB - We model experience-dependent plasticity in the adult rat S1 cortical representation of the whiskers (the barrel cortex) which has been produced by trimming all whiskers on one side of the snout except two. This manipulation alters the pattern of afferent sensory activity while avoiding any direct nerve damage. Our simplified model circuitry represents multiple cortical layers and inhibitory neurons within each layer of a barrel-column. Utilizing a computational model we show that the evolution of the response bias in the barrel column towards spared whiskers is consistent with synaptic modifications that follow the rules of the Bienenstock, Cooper and Munro (BCM) theory. The BCM theory postulates that a neuron possesses a dynamic synaptic modification threshold, thetaM, which dictates whether the neuron's activity at any given instant will lead to strengthening or weakening of the synapses impinging on it. However, the major prediction of our model is the explanation of the delay in response potentiation in the layer-IV neurons through a masking effect produced by the thresholded monotonically increasing inhibition expressed by either the logarithmic function, h(x) = mu log(1 + x), or by the power function, h(x) = mu x(0.8-0.9), where mu is a constant. Furthermore, simulated removal of the supragranular layers (layers II/III) reduces plasticity of neurons in the remaining layers (IV-VI) and points to the role of noise in synaptic plasticity. PMID- 10695762 TI - How to measure the information gained from one symbol. AB - Information theory provides a powerful framework to analyse how neurons represent sensory stimuli or other behavioural variables. A recurring question regards the amount of information conveyed by a specific neuronal response. Here we show that the commonly used definition for this quantity has a serious flaw: the information accumulated during subsequent observations of neural activity fails to combine additively. Additivity is a highly desirable property, both on theoretical grounds and for the practical purpose of analysing population codes. We propose an alternative measure for the information per observation and prove that this is the only definition that satisfies additivity. The old and the new definitions measure very different aspects of the neural code, which is illustrated with visual responses from a motion-sensitive neuron in the primate cortex. Our analysis allows additional interpretation of several published results, which suggests that the neurons studied are operating far from their information capacity. PMID- 10695763 TI - Natural scene statistics at the centre of gaze. AB - Early stages of visual processing may exploit the characteristic structure of natural visual stimuli. This structure may differ from the intrinsic structure of natural scenes, because sampling of the environment is an active process. For example, humans move their eyes several times a second when looking at a scene. The portions of a scene that fall on the fovea are sampled at high spatial resolution, and receive a disproportionate fraction of cortical processing. We recorded the eye positions of human subjects while they viewed images of natural scenes. We report that active selection affected the statistics of the stimuli encountered by the fovea, and also by the parafovea up to eccentricities of 4 degrees. We found two related effects. First, subjects looked at image regions that had high spatial contrast. Second, in these regions, the intensities of nearby image points (pixels) were less correlated with each other than in images selected at random. These effects could serve to increase the information available to the visual system for further processing. We show that both of these effects can be simply obtained by constructing an artificial ensemble comprised of the highest-contrast regions of images. PMID- 10695764 TI - Effective neural response function for collective population states. AB - Collective behaviour of neural networks often divides the ensemble of neurons into sub-classes by neuron type; by selective synaptic potentiation; or by mode of stimulation. When the number of classes becomes larger than two, the analysis, even in a mean-field theory, loses its intuitive aspect because of the number of dimensions of the space of dynamical variables. Often one is interested in the behaviour of a reduced set of sub-populations (in focus) and in their dependence on the system's parameters, as in searching for coexistence of spontaneous activity and working memory; in the competition between different working memories; in the competition between working memory and a new stimulus; or in the interaction between selective activity in two different neural modules. For such cases we present a method for reducing the dimensionality of the system to one or two dimensions, even when the total number of populations involved is higher. In the reduced system the familiar intuitive tools apply and the analysis of the dependence of different network states on ambient parameters becomes transparent. Moreover, when the coding of states in focus is sparse, the computational complexity is much reduced. Beyond the analysis, we present a set of detailed examples. We conclude with a discussion of questions of stability in the reduced system. PMID- 10695765 TI - Fibrinogen Bbeta14 Arg-->Cys: further evidence for a role in thrombosis. AB - A single base substitution (C-->T) in exon II of the Bbeta fibrinogen gene resulting in an Arg14-->Cys replacement was identified in a young woman with a history of recurrent thrombotic stroke. The patient's plasma showed prolongation of the thrombin and Reptilase times, and plasma fibrinogen, which was low when determined by chronometric assay (Clauss technique) was normal by clot weight. Dysfibrinogenaemia associated with the same mutation was identified in eight family members including two siblings with a history of venous and arterial thrombosis. Fibrin monomer polymerization with thrombin, Reptilase and Agkistrodon contortrix contortrix venom was defective. Polymerization studies revealed a reduced rate of polymerization compared with normal plasma, which improved on cooling from 37 degrees C to 20 degrees C. Plasma viscosity in the affected individuals was normal. Flow cytometric analysis of platelets from the proband and another affected member showed no increase in surface bound fibrinogen. Euglobulin clot lysis time was normal. The same point mutation has been described previously in individuals with thrombosis. This family adds further to the genotype-phenotype correlation of the dysfibrinogenaemias and provides strong evidence for a genuine association of fibrinogen BbetaArg14Cys with thrombosis. The mechanism underlying a causal relationship with the increased incidence of thrombosis remains obscure but a review of related dysfibrinogens suggests that the addition of a free thiol group rather than the loss of the thrombin cleavage site may be important. PMID- 10695766 TI - Influence of factor VIII/von Willebrand complex on the activated protein C resistance phenotype and on the risk for venous thromboembolism in heterozygous carriers of the factor V Leiden mutation. AB - High factor VIII plasma levels have been shown to represent a common increased risk for venous thromboembolism (VTE) and may cause an activated protein C (APC) resistance in the absence of the factor V Leiden mutation, but there are no studies specifically aimed to establish if high factor VIII and von Willebrand factor (vWF) concentrations may influence the APC sensitivity ratio (APC-SR) and increase the risk for VTE in the presence of the factor V Leiden mutation. For this purpose, we performed a retrospective case-control study to investigate the influence of the procoagulant factor VIII (VIII:C) and the antigen of vWF (vWF:Ag) on the normalized APC-SR (n-APC-SR) and on the risk for VTE, in two selected groups of 30 symptomatic (Group I) and 32 asymptomatic (Group II) related heterozygotes for the factor V Leiden mutation. Differences between the two groups (Group I versus Group II) were: n-APC-SR, 0.57+/-0.06 versus 0.63+/ 0.08, P = 0.001; factor VIII:C, 1.49+/-0.42 versus 1.13+/-0.28 IU/ml, P<0.001; vWF:Ag, 1.46+/-0.53 versus 1.26+/-0.32 IU/ml, NS. As a whole (Group I + Group II), Pearson correlation coefficients were: n-APC-SR versus factor VIII:C, r = 0.410, P = 0.001; n-APC-SR versus vWF:Ag, r = -0.309, P = 0.01; factor VIII:C versus vWF:Ag, r = +0.640, P<0.0001. The relative risk for VTE in individuals with the factor VIII:C concentration > 1.5 IU/ml was 2.5 (95% confidence interval 1.6-3.9). We concluded that high factor VIII:C levels, probably in the effect of vWF, play a determinant role in worsening the APC-resistance phenotype and represent a common additional risk factor for VTE in heterozygous carriers of the factor V Leiden mutation. PMID- 10695767 TI - Increased systemic coagulation activity in patients with rheumatic mitral stenosis: assessment of the clinical and echocardiographic determinants. AB - In this study, we aimed to determine systemic coagulation activity in patients with rheumatic mitral stenosis and to define determinants of a possible prethrombotic state. Peripheral venous plasma level of thrombin-antithrombin III complex was measured in 84 consecutive patients with rheumatic mitral stenosis who had no left atrial thrombus by transesophageal echocardiography. The patients had significantly higher thrombin-antithrombin III complex values (mean +/- SD = 9.6+/-15.9 ng/ ml) compared with the healthy subjects (2.1+/-1.8 ng/ml) (P<0.001). Among many clinical and echocardiographic variables, severe mitral regurgitation (odds ratio = 6.7, P<0.001) and left atrial spontaneous echo contrast (odds ratio = 22.8, P<0.001) appeared as significant predictors of the increased systemic coagulation activity in multivariate logistic regression analysis. In conclusion, systemic coagulation activity is increased in the patients with rheumatic mitral stenosis, and coexistence of severe mitral regurgitation and presence of left atrial spontaneous echo contrast are determinants of this increment. PMID- 10695768 TI - A study of the variability seen in the international normalized ratio obtained using different sensitivity thromboplastin reagents on different instrument types. AB - We performed an extensive study to check international normalized ratio (INR) recovery using three different rabbit thromboplastin reagents of different sensitivities [international sensitivity index (ISI) circa 1.1, 1.3 and 2.0] on two different instrument types. Both instrument types used an optical end point: one nephelometric at 660 nm, one photometric at 880 nm. Each thromboplastin reagent in the study had an instrument-specific ISI value assigned to it by the reagent manufacturer. Samples obtained from stable orally anticoagulated patients were tested on four different instruments of each type with the three different sensitivity reagents. Excellent correlation between INR values was seen between all results (r values between 0.97 and 0.99). Regression analysis gave slopes between 0.92 and 1.07, and a maximum deviation of intercept from zero of 0.21. Variations in CV of less than 2% were seen within each instrument type. This study shows that if instrument-specific ISI values are used, comparable INR values can be obtained using reagents of different sensitivity on multiple instruments of the same or different types. This study demonstrates the validity of the INR/ISI system. PMID- 10695769 TI - Hemostatic markers in Japanese patients undergoing anticoagulant therapy under thrombo-test monitoring. AB - The objective of this study was to evaluate several molecular markers of hemostasis in 84 patients with hypercoagulable state, treated with warfarin under thrombo-test (TT) monitoring; TT was expressed as percent of control (TT%). In all patients, the average values of international normalized ratio (INR) of prothrombin time (PT;PT-INR) was 1.68+/-0.49; this increase in PT-INR was not, however, significant in patients under TT% monitoring. There were no thrombotic or severe bleeding complications in these patients during a period of 2 years. Plasma levels of thrombin-antithrombin complex (TAT), plasmin-plasmin inhibitor complex (PPIC), D-dimer, and soluble fibrin monomer (sFM) were slightly increased, suggesting that anticoagulant therapy was not completely effective in our Japanese patients based on the values of TT%. Activated partial thromboplastin time, PT-INR, TT% and protein C activity were significantly correlated with the dose of warfarin; fibrinogen, activated thromboplastin, TAT, PPIC, D-dimer, sFM, protein S and thrombomodulin were not significantly correlated with the dose of warfarin. The PT-INR was negatively correlated with TT%, protein C and protein S, and the correlation between PT-INR and TT-INR was better than that between PT-INR and TT%. The range of TT% was not correlated with the plasma levels of TAT, PPIC, D-dimer or sFM, but the range of PT-INR was correlated with the plasma level of TAT, D-dimer and sFM. The percentage of TAT, D-dimer and sFM within normal range was significantly low in patients with high PT-INR. These finding showed that PT-INR is better than TT% for monitoring the anticoagulant therapy with warfarin, and that TT should be expressed as INR. The values of PT-INR should be more than 1.7 during the anticoagulant therapy with warfarin in Japanese patients with high risk of thrombosis. PMID- 10695770 TI - Chronic fatigue syndrome and/or fibromyalgia as a variation of antiphospholipid antibody syndrome: an explanatory model and approach to laboratory diagnosis. AB - Chronic Fatigue and/or Fibromyalgia have long been diseases without definition. An explanatory model of coagulation activation has been demonstrated through use of the ISAC panel of five tests, including, Fibrinogen, Prothrombin Fragment 1+2, Thrombin/ AntiThrombin Complexes, Soluble Fibrin Monomer, and Platelet Activation by flow cytometry. These tests show low level coagulation activation from immunoglobulins (Igs) as demonstrated by Anti-B2GPI antibodies, which allows classification of these diseases as a type of antiphospholipid antibody syndrome. The ISAC panel allows testing for diagnosis as well as monitoring for anticoagulation protocols in these patients. PMID- 10695771 TI - Idiopathic thrombocytopenic purpura treated with steroid therapy does not prevent acute myocardial infarction: a case report. AB - We report the case of a 65-year-old man affected by idiopathic thrombocytopenic purpura, who developed an acute myocardial infarction after 2 years of steroid therapy. Thrombocytopenia was initially recognized 11 years earlier, and became severe during the past 2 years [platelets (PLTS) 10000-30000/microl]. He was treated with steroids, initially to perform a surgical procedure (prednisone 75 mg/day), subsequently to maintain a platelet count of about 50000/microl (prednisone 12.5 mg/day). After 1 year of treatment, he began to complain about exertional angina and dyspnea. His blood pressure became elevated and cholesterol level raised. The exercise electrocardiogram, previously manifesting ischaemic changes, normalized after 1 month of steroid wash-out; however, steroid therapy was reinstituted (prednisone 5 mg per day). One year later, he suffered an infero lateral non-Q-wave myocardial infarction. It seems likely that the severe coronary atherosclerosis present in our patient developed despite a low platelet count, under the spur of a heavier risk factor profile. Steroid therapy could have had a role as a precipitating agent of the acute event, and the opportunity of alternative treatments is considered. PMID- 10695772 TI - Early occlusion of coronary by-pass associated with the presence of factor V Leiden and the prothrombin 20210A allele: case report. AB - The presence of factor V Leiden mutation and the variation of the prothrombin gene 20210GA have been described as additional risk factors for arterial thrombosis when other acquired or metabolic risk factors are present. We report here a 56-year-old man who developed coronary artery disease since 1980 without any known risk factor and underwent a cardiopulmonary by-pass in 1997. In the first month after surgery, he became symptomatic, and an angiography showed complete occlusion of the grafts and some native coronary arteries. Three months after the second cardiopulmonary by-pass, a thrombophilic state was searched, and plasma levels of lipoprotein (a) (LPa) were measured. The patient is heterozygous for factor V Leiden mutation and has the variation 20210GA of the prothrombin gene and high levels of LPa. These findings induced us to add oral anticoagulation to the aspirin treatment, and the patient is in a good condition 11 months later. PMID- 10695773 TI - The use of recombinant factor VIIa in a patient with severe Glanzmann's thrombasthenia to facilitate insertion of a Port-a-Cath. PMID- 10695774 TI - Lack of association between elevated serum Lp(a) and local thrombus lysability in patients with peripheral arterial occlusive disease. PMID- 10695775 TI - Normalization does not improve between-laboratory agreement but may improve specificity of some assays for activated protein C resistance. PMID- 10695776 TI - Trade wars and their impact on social contracts. PMID- 10695777 TI - Efficacy and effectiveness of five day treatment of uncomplicated falciparum with artemisinin or artesunate in Vietnam. AB - A study on efficacy and effectiveness of artemisinin (total dose of 60 mg/kg) and artesunate (total dose of 12 mg/kg over five days) in treatment of uncomplicated malaria was conducted in highly malaria transmitted areas in Vietnam. 126 uncomplicated malaria cases finished 14 day follow-up. 100% cure rate achieved at day 14 in patients of the efficacy groups received either artemisinin or artesunate, while it was 83% and 93% in patients treated respectively with artemisinin and artesunate of the effectiveness groups. Compliance of the treatment regimens was discussed. PMID- 10695778 TI - The effects of quinine and artesunate treatment on plasma tumor necrosis factor levels in malaria-infected patients. AB - Tumor necrosis factor-alpha (TNF-alpha) is an endogenous mediator of shock and inflammation including malaria. Many lines of evidence suggest that cytoadherence, the life-threatening pathology associated with complicated and cerebral malaria, results from the overproduction of TNF in response to malarial parasite. Quinine has been shown to inhibit TNF synthesis and cytoadherence in vitro suggesting an additional beneficial effect of quinine on its anti-TNF action. On the other hand, artesunate inhibits cytoadherence better than quinine does not suppress TNF production in vitro. The present study compares the effect of artesunate and quinine on TNF levels of malaria-infected patients. Surprisingly, plasma TNF levels increased dramatically after quinine administration but did not increase after artesunate administration. This difference may be explained by previous observations showing that artesunate kills parasites in vitro and clears parasitemias in vivo for more rapidly than quinine. The rapid clearance of plasma TNF in quinine treated patients might be due to the drug's TNF-suppressive activity. PMID- 10695779 TI - Phamacokinetics of a single oral dose of dihydroartemisinin in Vietnamese healthy volunteers. AB - Pharmacokinetics of a 240 mg single dose of oral dihydroartemisinin (DHA) was investigated in 8 healthy (5 males, 3 females) Vietnamese volunteers. Plasma concentrations were measured by high-performance liquid chromatography with electrochemical detection in the reductive mode. The concentration time profile of DHA was fitted with one-compartment model with a lag time. Pharmacokinetics of DHA is comparable between males and females even when adjusted with dosage. The median (range) values of pooled pharmacokinetics of oral DHA were: t(lag) 0.41 (0.09-0.78) hours, t(1/2z) 0.58 (0.17-1.43) hours, t(max) 1.6 (1.1-2.2) hours, Cmax 466 (128-787) ng/ml. Cmax/dosage 97.7 (27.2-124.6) ng/ml, t(1/2z) 2.0 (1.5 3.4) hours, AUC 1867 (420-3535) ng x h/ml, AUC/dosage 364.3 (89.3-559.7) ng x h/ml/dosage, Cl/f 45.8 (30.0-190.0) ml/min/kg, Vz/f 8.0 (5.5-29.9) l/kg. Interindividual variation was large, the coefficients of variation (CV) were 47.8% and 45.3% respectively to AUC and Cmax. The t(max) of DHA formulation was comparable with that of DHA metabolite of artemisinin derivatives. The t(1/2z) was longer and shorter than that of DHA metabolites of oral formulations of artesunate and artemether, respectively. For monotherapeutic regimen(s) of DHA, dosing frequency of at least twice a day is suggested. Combined regimen(s) of DHA with other potent, long half-life antimalarials may also be an alternative approach. PMID- 10695780 TI - Dose findings of dihydroartemisinin in treatment of falciparum malaria. AB - Forty patients with uncomplicated P. falciparum malaria were respectively treated in an open randomized comparative study of dihydroartemisinin tablets given at total doses of 480 mg over 5 days and 640 mg over 7 days in a drug-resistant malaria endemic area in Hainan, China. The result showed that all patients were clinically cured. In 5-day and 7-day groups, the mean fever clearance times (FCT) were 26.1+/-10.2 and 21.1+/-11.8 hours respectively; the mean parasite clearance times (PCT) were 58.7+/-20.9 and 59.4+/-20.9 hours respectively, which showed no significant difference. 28-day follow-ups were accomplished on 39 and 37 cases respectively in two groups, the recrudescence rates were 20.5% (8/39) in 5-day group, while 2.7% (1/37) in 7-day group with significant difference (chi2=4.19, p<0.05). No clinical drug-related side effect was found in two groups during treatment. PMID- 10695781 TI - Use of PCR/DNA probes to identify circumsporozoite genotype of Plasmodium vivax in China. AB - The paper reports the result of identifying cirumsporozoite (CS) genotype of Plasmodium vivax by using PCR/DNA probe labeled with biotin. The sensitivity of this method to detect patient blood samples was 0.2 parasite/microl and also with high specific to P. vivax. CS genes from 52 blood samples collected from patients with P. vivax in Hainan and Yunnan Provinces were amplified by PCR and 49 were positive by gel-e electrophoresis analysis, positive rate was 94%. Then the amplified CS genes further were probed with special oligoprobes (PV210 and PV247) that hybridized with the predominant CS repeat region and the variant CS repeat region. The results showed 46 (88.5%) PV210 positive and 6 (11.5%) PV247 positive; 2 hybridized with both probes. The variant genotype was present only in samples from Yunnan Province. The above results showed that the PCR/DNA probe labeled with biotin was highly sensitive and specific to P. vivax and found a CS variant genotype of P. vivax in Yunnan Province of China. PMID- 10695782 TI - Human antibody isotype responses to Schistosoma japonicum egg antigen. AB - Schistosoma japonicum-infected subjects from Hubei province of China were investigated to determine the class and subclass of the antibody response to soluble egg antigen (SEA), using an enzyme-linked immunosorbent assay. The subjects were 50 acute and 55 chronic cases. In acute cases, the mean OD values for IgA, IgE and IgG3 were very high, while the positive ratios of IgA and IgE were only 78% and 74%, respectively. The positive ratios of IgG, IgM, IgG1, IgG3 and IgG4 were all above 90%. In chronic cases, the mean OD values for IgG, IgG3 and IgG4 were very high, and the positivity rates of IgG, IgG1, IgG3 and IgG4 were all above 90%. Comparing the two study groups, the mean OD values of IgM, IgA, IgE were higher in acute cases than those of chronic cases (p < 0.0001), while the mean OD values of IgG, IgG4 were higher in chronic cases than in acute cases (p < 0.05). The mean OD values of IgG3 in both groups were high and those of IgG2 in both groups were low. PMID- 10695783 TI - Comparative studies on detecting CAg in urine of acute schistosomiasis patients by mAb-RIHA and mAb-DotELISA. AB - Urine was concentrated 20-fold for assay for CAg of Schistosoma japonicum. mAb RIHA and mAb-DotELISA were positive in 78, 31% and 65.06% of cases respectively, of 83 patients with acute schistosomiasis. The false positive rates in 101 healthy controls were 14.85% and 0%, respectively. Cross-reactions (using mAb RIHA) were seen in 16.36% and 14.28% of patients with clonorchiasis, 49 patients with ankylostomiasis, respectively. Corresponding figures for mAb-DotELISA were 0% and 0%. PMID- 10695784 TI - Schistosoma japonicum in the pig: the influence of age on the host/parasite relationship. AB - The current study sought to elucidate a possible association between age and susceptibility to a primary infection with Schistosoma japonicum in pigs. Sixteen Landrace/Yorkshire crossbred specific pathogen-free pigs in three different age groups (group A-C), aged approximately 7, 24 and 37 weeks at the beginning of the experiment, were infected by intramuscular injections of 1,000, 1,500 or 2,400 cercariae, respectively. Fecal egg counts were obtained twice weekly from six to eight weeks post infection (wpi), and the pigs were killed 11 wpi. The number of worms collected were counted and sexed subsequent to perfusion. Tissue egg counts were estimated on samples from the liver. The worm recoveries for group A, B and C were 3.2%, 8.1% and 3.8%, respectively. No differences were observed between the male/female ratios of the three groups. The fecundity parameters, ie, fecal egg counts per mature female and liver egg counts per mature female, showed no significant differences between the three age groups. The results did not indicate any difference in susceptibility between the different age-groups of pigs to a primary infection with S. japonicum. PMID- 10695785 TI - Visceral leishmaniasis presenting as generalized lymphadenopathy in Nepal. PMID- 10695786 TI - Difference of Toxoplasma gondii antibodies between Thai and Austrian pregnant women. AB - Toxoplasma gondii IgG and IgM antibodies of Thai and Austrian pregnant women were studied, in the same laboratory unit, by using the Sabin-Feldman dye test and ISAGA-IgM. In Thai pregnant women, IgG antibody was found in 21.7%, mostly the IgG titers were low and all were negative for IgM antibody. Conversely in Austrian pregnant women, IgG antibody was found in 30.0% with high titer, and there were 19 (6.3%) cases positive for IgM antibody. The seropositivity of T. gondii IgG antibody in Austrian pregnant women was significantly higher than in Thai (p = 0.02) and the titers were much higher. Two possibilities are postulated to explain the data: it may be because Thai women were infected at a younger age than Austrian women, so they were in the chronic infection stage corresponding with their negativity of IgM antibody or it may be due to the difference of strain virulence of T. gondii from the different parts of the world. PMID- 10695787 TI - Baculovirus expression of the major surface antigen of Toxoplasma gondii and the immune response of mice injected with the recombinant P30. AB - The major surface antigen (P30) of the Toxoplasma gondii was expressed by an insect cell culture system infected with recombinant baculovirus. About 750 microg of purified (95% purity) P30 was obtained from a culture of 10(8) insect Sf21 cells. The recombinant P30 was used to immunize mice to induce immune response. Mice injected with the recombinant protein produced specific humoral and cellular immune responses. Immunization with P30 also prolonged the period of survival of mice infected by Toxoplasma. The average survival time of control group is 13.25+/-1.16 days, but are 16.13+/-2.1 days, 19.50+/-3.21 days, 20.38+/ 3.38 days in different immunized groups, respectively. PMID- 10695788 TI - ICT filariasis test: a new screening test for Bancroftian filariasis. AB - Bancroftian filariasis can be detected by using the ICT Filariasis test kit which is composed of specific polyclonal and monoclonal antibodies to Wuchereria bancrofti antigen. Chromatographic reaction with serum or plasma shows a result within 5 minutes. When compared with 454 thick blood films (standard smear method) within the same study, the ICT Filariasis test had sensitivity = 100%, specificity = 96.37%, efficiency = 96.70%, predictive value positive (PVP) = 70.70%, predictive value negative (PVN) = 100%. Compared with 454 membrane filtration technic (MFT), the MFT had sensitivity = 95.10%, specificity = 99.50%, efficiency = 99.12%, PVP = 95.10%, PVN = 99.50%. When we compared capillary tube technic (CAP) with TBF, CAP showed sensitivity = 85.40%, specificity = 100%, efficiency = 98.68%, PVP = 100%, PVN = 98.60%. With the convenience, high sensitivity-efficiency, lack of cross-reactions, no night blood collection, single reagent and rapidity of the test, the ICT Filariasis test can be recommended for screening of Bancroftian filariasis, and is suitable for the confirmation of suspected cases in the field where microscopic diagnosis is not available. PMID- 10695789 TI - The prevalence of Chlamydia trachomatis infection in rural Thai women. AB - A cross sectional study was designed to investigate the prevalence of Chlamydia trachomatis among different groups of rural women in the northeast Thailand. The presence of chlamydial antigens in endocervical swabs was detected by ELISA. The prevalences of Chlamydia trachomatis were 6.8% (31/485), 5.2% (24/466) and 6.7% (12/179) in women attending antenatal, postpartum and family planning clinics respectively. The average prevalences of C. trachomatis among hospital-based and community-based women were 6.1% (67/1,103) and 3.6% (15/411) respectively. In addition, the prevalences of some pathogens including Candida albicans, Trichomonas vaginalis, Treponema pallidum and Neisseria gonorrhoea among hospital based and community-based women were 14.2, 2.8, 0.7, 0.2 and 10.9, 5.1, 2.7, 0.0% respectively. It was concluded that C. trachomatis was a problem of woman's reproductive health. PMID- 10695790 TI - Study of Cyclospora cayetanensis in health care facilities, sewage water and green leafy vegetables in Nepal. AB - Cyclospora cayetanensis, a newly emerging parasite, is endemic in Nepal. A total of 2,123 stool specimens were collected from 3 health care facilities based on clinical symptoms during the period between 1995 to October, 1998. Out of these specimens, cayetanensis oocysts were found in 632 (29.8%). To identify possible sources for Cyclospora infection, drinking water, sewage water, green-leafy vegetables including fecal samples of various animals were collected and examined. The vegetable leaves were washed in distilled water then the washings, sewage water and drinking water were centrifuged and the sediment were examined microscopically. As a result, oocyst of Cyclospora were identified in sewage water and vegetable washings on four different occasions in June, August, October and November. The positive results were also confirmed as C. cayetanensis by development of 2 sporocysts after 2 week incubation period in potassium dichromate. A survey of 196 domestic animals from the same areas demonstrated that two chickens were positive for Cyclospora-like organism and others were negative. Although further studies are needed to clarify the direct link between Cyclospora infection and these sources, the results suggest that sewage water, green leafy vegetables are possible sources of infection and chickens could be possible reservoir host of Cyclospora in Nepal. PMID- 10695791 TI - Soil contamination and infections by soil-transmitted helminths in an endemic village in southern Thailand. AB - The aim of this study was to test the association between soil contamination and infection of the household members by soil-transmitted helminths in dry and rainy seasons. A lake-side community in southern Thailand with a population of 2,340 was studied twice, in the dry season and rainy season of 1995. Fifty households were randomly selected. Soil samples near the latrine, in the yard, at the foot washing area and under the trees were taken and analysed for presence of helminthic eggs. All members of the selected household were interviewed and stool samples obtained. Age-adjusted odds ratios of presence of Ascaris and Trichuris eggs in the household soil for ascariasis and trichuriasis were 10.5 (95% CI 1.5 77.1) and 5.5 (95% CI 2.4-12.7) in dry season and 10.4 (95% CI 2.5-43.8) and 8.3 (95% CI 3.4-20.0) in rainy season. The levels of hookworm eggs detected in the soil were too low to test the association. Soil analysis for eggs of Ascaris and Trichuris may be used to predict infections among the household members but not that for hookworm. PMID- 10695792 TI - Questionnaire survey and prevalence of intestinal helminthic infections in Barru, Sulawesi, Indonesia. AB - A questionnaire survey with parasitological study was carried out on the inhabitants of 4 villages in Barru district, Sulawesi, Indonesia from 1994 to 1995. The questionnaire dealt with life style and sanitary conditions. In 482 houses in the 4 villages, interviews for the items of the questionnaire were conducted with the owner, housekeeper and children of the same family. In Pancana and Lalolang, 37.7% and 50% respectively of man inhabitants surveyed were fishermen, while in Lompo Riaja and Pattappa, 38.6% and 65.5% respectively were farmers. The highest proportion of official workers was 33.7% in Lompo Riaja. Educational level was low; 88.4% in Pancana, 90.4% in Lalolang, 62.1% in Lompo Riaja and 91.2% in Pattappa had elementary or below elementary school education. In Lompo Riaja, 30.8% of the inhabitants graduated from senior high school or university. The percentage of families having their own latrine was 30.3% in Pancana, 13.2% in Lalolang, 31.9% in Pattapa and 60% in Lompo Riaja. The people without latrines usually defecated in rice fields, seaside or riverside. A total of 654 fecal samples was examined by the modified Kato-Katz thick smear method. Five nematode species, Ascaris lumbricoides, Trichuris trichiura, Necator americanus, Strongyloides stercoralis and unidentified Rhabditoids of free-living nature were detected. Cestode, Hymenolepis nana infection was confirmed. All the hookworms examined by the modified Harada-Mori culture technic were Necator americanus. Trichuris infection was most common, followed by hookworm and Ascaris infections, both in young (aged 4-14) and older (aged over 15) age groups. The prevalence of hookworm infection was significantly higher in males than in females of older age. Among the older age group, the prevalence of Trichuris infection was significantly lower in Lompo Riaja, while hookworm infection was the highest in Pattappa. Among all the inhabitants examined for parasite infection, 17.4% had 3 kinds of nematode, Ascaris, Trichuris and hookworm. However, egg counts revealed that most of the inhabitants with Trichuris or hookworm had light infections. The inhabitants with higher education background had significantly lower infection rates of Ascaris and Trichuris. The prevalence of hookworm infection was not significantly different between the inhabitants owning latrine and without it, but the prevalence of Ascaris and Trichuris, differed significantly. PMID- 10695793 TI - Albendazole treatment for Giardia intestinalis infections in school children. AB - A randomized controlled trial, 113 school children with Giardia intestinalis infection were treated with albendazole or tinidazole. Albendazole 400 mg once a day x 3 days and tinidazole 50 mg/kg single dose were given orally to 62 and 51 children, respectively. Parasitological cure was documented when there were > or = 2 times negative stool examination for G. intestinalis at 1-2 weeks after therapy. Thirty-one of 62 (50%) children treated with albendazole and 49 of 51 (96.1%) children treated with tinidazole had parasitological cure (p < 0.001). No major side effects were observed except one case in tinidazole group had severe headache for 30 hours. Albendazole appears to be safe and produced a moderate cure rate for G. intestinalis infection when a 3 day anthelmintic regimen is given. PMID- 10695794 TI - Overcoming the errors of in-house PCR used in the clinical laboratory for the diagnosis of extrapulmonary tuberculosis. AB - Our experiences from 1993 to 1997 in the development and use of IS6110 base PCR for the diagnosis of extrapulmonary tuberculosis in a routine clinical setting revealed that error-correcting processes can improve existing diagnostic methodology. The reamplification method initially used had a sensitivity of 90.91% and a specificity of 93.75%. The concern was focused on the false positive results of this method caused by product-carryover contamination. This method was changed to single round PCR with carryover prevention by uracil DNA glycosylase (UDG), resulting in a 100% specificity but only 63% sensitivity. Dot blot hybridization was added after the single round PCR, increasing the sensitivity to 87.50%. However, false positivity resulted from the nonspecific dot blot hybridization signal, reducing the specificity to 89.47%. The hybridization of PCR was changed to a Southern blot with a new oligonucleotide probe giving the sensitivity of 85.71% and raising the specificity to 99.52%. We conclude that the PCR protocol for routine clinical use should include UDG for carryover prevention and hybridization with specific probes to optimize diagnostic sensitivity and specificity in extrapulmonary tuberculosis testing. PMID- 10695795 TI - Susceptibility to hepatitis A virus infection among chronic liver disease patients and healthy blood donors in Thailand. AB - Due to improvements in socio-economic and sanitation conditions, Thailand has undergone a change from hyperendemicity to intermediate endemicity for hepatitis A virus infection, leaving a large part of the adult population without immunity. At the same time, the country is still highly endemic for hepatitis B and especially in the northeast, hepatitis C virus infection both of which when acquired during infancy or early childhood exhibit a strong tendency to turn towards chronic liver disease, although in particular with hepatitis B virus the asymptomatic carrier state is also rather common. As no cross-immunity exists between any of these viruses, double or triple infections do occur, a situation where previously acquired immunity to HAV becomes crucial as double infections have been shown to take a more severe or even fatal course. In the present study, we investigated 820 HBV- and/or HCV-related chronic liver disease (CLD) patients and 195 blood donors, both groups divided by 10-year age intervals, for the prevalence of anti-HAV. The results showed the same age dependence of immunity for all groups tested as can be expected for an area of intermediate endemicity, in that approximately 50% of those between 21 and 30 years of age had acquired anti-HAV. These findings indicate the immune response to HAV infection not to be altered by chronic infection with either HBV or HCV. Hence, vaccination against HAV should be considered, particularly in anti-HAV-negative patients with CLD. PMID- 10695796 TI - Current status of infection-related gastrointestinal and hepatobiliary diseases in Thailand. AB - The objective of this overview is to assess the present situation with regards to gastrointestinal and hepatobiliary diseases prevailing in Thailand. In that context, special emphasis has been put on those forms of viral hepatitis prevalent in the region, namely, hepatitis A the frequency of which has undergone a change from hyper- to hypoendemic with a resulting decline in naturally acquired immunity; hepatitis B with its tendency to cause chronic liver disease mainly due to asymptomatic infections during early childhood and the impact of mass vaccination programs on its endemicity; hepatitis C which can also lead to chronicity; hepatitis D solely found as a coinfection with hepatitis B; hepatitis E acute cases of which can sporadically be found; hepatitis G encountered in healthy subjects at a prevalence similar to that seen in patients with chronic liver disease and rather more prevalent among people at risk for contracting blood borne agents; finally the novel hepatitis TT virus with a distribution comparable to that of hepatitis G virus and a similarly unclear role as to the etiology of serious liver disease. Particularly in connection with hepatitis B we have examined the situation regarding hepatocellular carcinoma which represents one of the most common malignancies among the Thai population. Cholangiocarcinoma caused by the liver fluke Opisthorchis viverrini is the most common form of liver cancer in the northeastern part of Thailand where an estimated 70% of the population are infested with the parasite. Peptic ulcer caused by Helicobacter pylori constitutes another common gastrointestinal affliction with the overall prevalence of antibodies to the agent amounting to 63 to 74% in patients exhibiting gastroduodenal symptoms. The final part of the paper deals with HIV related gastrointestinal and liver disease and with amebic and pyogenic liver abscesses. PMID- 10695797 TI - A four year review of acute viral hepatitis cases in the east coast of peninsular Malaysia (1994-1997). AB - A total of 1,157 sera from jaundiced patients with clinical and biochemical evidence of liver disease received from government hospital in Kelantan and Terengganu, during the period from 1994 to 1997, were investigated to determine the cause. Hepatitis A virus was found to be the main cause in 26.1% (24/92) of symptomatic clinical hepatitis cases in 1994, 47.8% (63/132) in 1995, 66.4% (613/923) in 1996 and 20% (2/10) in 1997. Sera received in 1996 were also tested for hepatitis B, hepatitis C, hepatitis D and hepatitis E. 1.4% (13/923) anti bodies were found to be positive for HBc IgM indicating recent HBV infection, 5.4% (50/923) for total HCV Ab, 0.9% (8/923) for total HDV Ab and 0.4% (4/923) for anti-HEV IgM. This study shows that HAV is still a major problem in Kelantan and Terengganu, and there is a need to identify effective strategies for prevention and control in these two states. PMID- 10695798 TI - Alpha-L-fucosidase as a serum marker of hepatocellular carcinoma in Thailand. AB - To evaluate the role of serum alpha-L-fucosidase (AFU) in the diagnosis of hepatocellular carcinoma (HCC), we simultaneously studied both AFU activity and alpha-fetoprotein (AFP) level in 60 patients with HCC, 60 patients with cirrhosis and chronic hepatitis each, 30 patients with other liver tumors and 60 healthy subjects. Serum AFU activity in patients with HCC (1,418.62 +/- 575.76 nmol/ml/hr) was significantly higher than that found in cirrhosis (831.25 +/- 261.13 nmol/ml/hr), chronic hepatitis (717.71 +/- 205.86 nmol/ ml/hr) or other tumors (706.68 +/- 197.67 nmol/ml/hr) and in controls (504.18 +/- 121.88 nmol/ml/hr, p < 0.05). With 870 nmol/ml/hr (mean value of controls plus 3 standard deviations) considered as the cut-off point, AFU was more sensitive (81.7 vs 39.1%) but less specific (70.7 vs 99.3%) than AFP at a level of > 400 ng/ml as a tumor marker of HCC. With both markers combined, the sensitivity was improved to as much as 82.6%. AFU activity in HCC patients was correlated to tumor size (r = 0.3529, p = 0.006) but not associated with tumor staging classified by Okuda's criteria (p = 0.1). The AFU activity in the viral hepatitis group (hepatitis B or C) was also significantly higher than in the non-viral group (p = 0.0005). We conclude AFU to be a useful marker, in conjunction with AFP and ultrasonography, for detecting HCC, particularly in patients with underlying viral hepatitis and cirrhosis. PMID- 10695800 TI - The nutritional status of children in resettlement villages in Kelantan. AB - A cross-sectional survey of the nutritional status of children aged 1-10 years old from the Kuala Betis resettlement villages was carried out. A total of 620 children were examined, of which 329 were preschool children and 291 were schoolchildren. The age was determined and anthropometric measurements such as weight, height and MUAC were taken. The nutritional status was assessed by looking at the distributions of the z-scores of weight-for-age (WAZ), height-for age (HAZ) and weight-for-height (WHZ) in relation to the growth charts of the National Center for Health Statistics reference population. It was found that the nutritional status of the Orang Asli children was poor, with a prevalence of 33.7 65.3% underweight, 55.3-74.4% stunting and 4.4-29.7% wasting based on the NCHS reference values. The prevalence of malnutrition among the Malay children was lower, underweight--7.3-34.1%, stunting--9.8-34.1% and wasting--1.7-17.1%. The nutritional status of the Orang Asli children were poorer compared to the Malay children. More preschool Orang Asli children were stunted compared to the Orang Asli schoolchildren. This may be due to the poor economic base of the Orang Asli community during the transformation period after resettlement. A comprehensive primary health care program is essential, especially targeting the preschool Orang Asli children in these resettlement villages. PMID- 10695799 TI - Healthy dietary habits, body mass index, and predictors among nursing students, northeast Thailand. AB - This study aimed to assess body mass index (BMI) of nursing students, and examine the links between health behavior in terms of healthy dietary habits, positive health habits, dieting and BMI. A structured questionnaire was used for obtaining information on dietary habits, positive health habits, demographic characteristic including body weight, and height by administering self-answering questionnaires to all of nursing students in the 1st, 2nd, 3rd, and 4th year-classes of the College of Nursing located in northeast Thailand. Three hundred and eleven female nursing students with an average age of 19.9 (SD = 1.4), had an average BMI of 20.3 kg/m2 (SD = 1.9). Most of the subjects (82.6%) were in the acceptable weight category (BMI > 18.5-24.99 kg/m2), 5.1% underweight (BMI < or = 18.5 kg/m2), and 2.3% overweight. (BMI > or = 25.0 kg/m2). About half of them (50.8-66.2%) practiced healthy dietary habits in terms of avoiding eating fat/cholesterol, enriched fiber foods, while one-fourth practiced daily fruit consumption. Positive health habits in terms of having breakfast, and taking exercise over the last two weeks, were practiced by 49.5% and 59.8%, respectively. Persistent health problem occurred 13.5% amongst the subjects. The univariate analyses revealed significant associations between dieting with the BMI; perception of body size with the BMI; the enriched fiber food consumption with dieting; and the avoidance of fat/cholesterol with dieting. It suggests that the choice of food was predominantly attributable to dieting. Results from multiple logistic regression analysis showed that dietary belief, dieting, and exercise had effects on the strength of the association (p = 0.0191, 0.0024, 0.0165; Odds ratios = 0.97, 2.21, 1.87, respectively). The results and implications are discussed. PMID- 10695801 TI - Causes of death and unintentional injury among schoolchildren in Thailand. AB - Few prospective studies of mortality among children in developing countries have been published. Here we quantify and describe mortality and injury morbidity among a cohort of schoolchildren in rural Southeast Asia. Deaths among a cohort of 40,119 schoolchildren in Thailand were prospectively monitored over a two year period from January 1991. Additionally, data were collected with a questionnaire from a subset of 6,378 children asking them to recall all injuries over a one year period. There were 40 deaths for an annual incidence of 50/100,000. Fifty percent of all deaths were due to injury; 25% to infectious diseases. Sixty percent of the injury deaths were due to motor vehicles and 35% to drowning. Sixty-six percent of the children reported one or more accidents. The leading categories of non-fatal injuries, in decreasing order, were: animal bite, puncture wound, burn, near-drowning, fall from a height. Boys experienced more injuries than girls for almost every type of injury. Injuries are replacing infectious diseases as the most important cause of deaths in developing countries. Additional public health initiatives to reduce childhood accidents may be warranted. PMID- 10695802 TI - Coping with ill health in a rickshaw puller's household in Chittagong, Bangladesh. AB - The correlation between poverty and ill-health is undeniably strong. Ill-health reduces the earning capacity, and increases the risk of families with ill members to drift down the social and economic ladder. In this article, we present a simulation model of how a poor rickshaw puller in Bangladesh copes with illness, in particular tuberculosis (TB). We first analyze the various coping mechanisms that are set in motion when he starts to suffer from tuberculosis; the impact on household assets, income and food intake will be studied. The simulation model is then used to analyse the effects on his household of a specific health intervention, namely the Directly Observer Treatment Short Course (DOTS) treatment. It shows that DOTS offers positive improvements of the overall well being of the household by restoring the working capacity of the rickshaw puller in one treatment course and minimizing lost income. Assets and food consumption would be preserved significantly more in the presence of DOTS, rendering the household both financially and physically less vulnerable. The probability of death of the sick rickshaw puller is also significantly reduced, improving household's welfare over the long run. PMID- 10695803 TI - Presenting features and treatment outcome of 78 Malaysian children with neuroblastoma. AB - To study the distribution of presenting features and their prognostic significance in neuroblastoma treated in a single institution in Malaysia. A retrospective study was made of 78 neuroblastoma cases diagnosed and treated in the University Hospital, Kuala Lumpur, Malaysia between June 1982 and February 1997. Diagnosis was established by standard histological criteria. The presenting features were evaluated for their distribution and prognostic influence. Disease free survival from diagnosis was the outcome variable of interest. The ages ranged from 0.1 to 11 years old (median: 3 years old). The tumor originated from the adrenal glands in 83% and the majority of cases presented in advanced stage (stage III 22%, stage IV 66%). Bone marrow was the commonest site of distant metastasis occurring in 45% of patients. The main presenting signs and symptoms in decreasing order were pallor, fever, abdominal mass, weight loss, and bone/joint pain. Univariate analysis conferred age, initial stage and Hb level as significant prognostic factors. No influence in disease-free survival was found for sex, race, primary site, urinary vanillylmandelic acid level, white cell count and platelet count. Overall 2-year disease-free survival was achieved in 27 (39%) patients. Four patients underwent bone marrow transplant, three of whom achieved 2-year disease-free survival. The results suggest that age, initial stage and hemoglobin level are significant prognostic factors based on univariate analysis. In addition, more Malaysian children presented with adrenal primary site and advanced disease compared to previous reported studies. PMID- 10695804 TI - Intracerebral hemorrhage due to nosocomial aspergillosis following neurosurgery. AB - A unique case of nosocomial aspergillosis following neurosurgery in a 10 year old girl was documented. She presented with intracerebral hemorrhage after three weeks of operation for evacuation of craniopharyngioma. To our knowledge, this is the first reported case of intracerebral hemorrhage due to nosocomial aspergillosis following neurosurgery. PMID- 10695805 TI - Fatal mushroom poisoning caused by Amanita virosa in Thailand. AB - Consumption of toxic mushrooms belonging to the genus Amanita frequently leads to severe gastrointestinal distress followed by acute hepatic failure with a fatal outcome. In Thailand, valuable information as to the locally prevalent poisonous species, the preferred habitat and the management of suspected victims of intoxication is basically non-existent. We report here 5 cases of fatal poisoning with Amanita virosa having occurred in a family residing in the northeast of Thailand who as countless others had enjoyed mushroom gathering as a pasttime. Within 4 to 6 days after ingestion of the mushrooms, all had succumbed to acute hepatic failure with subsequent hepatoencephalopathy. Treatment modalities exist in the form of penicillin and silibinin, or thioctic acid administration followed by plasmapheresis. In cases taking a lethal course apparent from the results of liver biochemistry, liver transplantation is clearly indicated. In order to prevent mushroom poisoning altogether, educating the general population to that end certainly presents the method of choice. PMID- 10695806 TI - Identification of a flavivirus isolated from mosquitos in Chiang Mai Thailand. AB - A virus isolate, ThCAr105/92, from a pool of mosquitos, Culex tritaeniorhynchus, collected in Chiang Mai, Thailand in 1992, appeared to be a member of the genus Flavivirus of the family Flaviviridae, based on the reverse transcription polymerase chain reaction (RT-PCR) using flavivirus cross-reacting primer pairs, electron microscopic examination, and serological tests. However, RT-PCR using Japanese encephalitis (JE) virus-specific primers showed that the isolate was different from JE virus. Sucrose density gradient sedimentation of the virus replicated in C6/36 cells indicated that the virus is relatively unstable in the infected culture fluids at 37 degrees C. Antibody prepared against this virus and a virus seed for the isolate were tested by cross neutralization against a panel of flaviviruses and the results showed that the new isolate was a distinct subtype of Tembusu virus. PMID- 10695807 TI - Genetic and morphological variations in populations of Oncomelania spp in China. AB - Oncomelania snails are the intermediate hoste of Schistosoma japonicum in Asian countries. In order to understand the genetic and morphological variation of Oncomelania snails in mainland China, field snails from 31 localities were collected and investigated by means of allele enzyme electrophoresis and numerical taxonomical technics. Results demonstrated that out of 17 loci examined, seven polymorphic loci were presented. Genetic distance (Nei, 1978) among the populations varied from 0.03 to 0.27. The phenogenetic tree based on UPGMA cluster analysis showed that genetic diversity corresponded to geographic distribution along the Yangtze River, which provided supplementary genetic data about the evolution of Oncomelania spp. A morphological study showed that Mahalanobis' morphological distance ranged from 1.53 to 346.7. Both genetic and morphological data indicated that the diversity among populations of smooth shelled snails was higher than that among populations of ribbed shelled snails. A positive correlation (r = 0.80) between Mahalanobis' morphological distance and genetic distance supports the hypothesis that the different shell phenotypes represent different species or subspecies. PMID- 10695808 TI - Laboratory evaluation of lambda-cyhalothrin a microencapsulated formulation on mosquito nets for control of vector mosquitos. AB - Two formulations of lambda-cyhalothrin (EC-Emulsion concentrate and MC Microencapsulated) were impregnated into bednets made of polyethylene and polyester. The nets were treated at a dosage of 15 mg/m2. For bioassay of insecticidal efficacy, female Anopheles maculatus and Aedes aegypti were exposed to the nets for two minutes and mortality was scored 24 hours later. The nets were also tested after repeated washings with water and with soap and water. Microencapsulated (2.5CS) formulation was more effective than emulsion concentrate (2.5EC) formulation on both net materials--polyethylene and polyester. Repeated washing with water and soap reduces the efficacy of all bednet treatment combinations. Microencapsulated formulation on polyethylene gave best results; it could sustain up to five washes with water and two with soap and water. PMID- 10695809 TI - Current insecticide resistance patterns in mosquito vectors in Thailand. AB - Chemical pesticides are still commonly used in Thailand for control of agricultural pests and disease vectors. Organophosphates, carbamates and synthetic pyrethroids are commonly used for agricultural purposes, whereas synthetic pyrethroids have become more popular and predominate for public health use. The genetic selection of insecticide resistance (whether physiological, biochemical or behavioral) in pests and disease vectors has been extensively reported worldwide (Brown and Pal, 1971). The long-term intensive use of chemical pesticides to control insect pests and disease vectors is often cited as the reason behind the development of insecticide resistance in insect populations. Unfortunately, reliable information on vector resistance patterns to pesticides in Thailand is sparse because of a remarkable shortage of carefully controlled, systematic studies. This review gathers useful information on what is presently known about disease vector resistance to chemical pesticides in Thailand and provides some possible management strategies when serious insecticide resistance occurs. PMID- 10695810 TI - Possible site of action of Kaempferia galanga in killing Culex quinquefasciatus larvae. PMID- 10695811 TI - Influence of IGR treatment on oviposition of three species of vector mosquitos at sublethal concentrations. AB - Sublethal effect of hexaflumuron, an insect growth regulator (IGR), on the oviposition of three species of vector mosquitos. Culex quinquefasciatus, Aedes aegypti and Anopheles stephensi was studied. Significant reduction in oviposition was observed in the females of the above three species derived from fourth instar larvae and pupae exposed to sublethal (EI5 and EI50) doses. The reduction in egg laying is proportional to the dose of exposure and was found to be about twice higher in females of three species exposed to EI50 dose than those exposed to EI5 dose. Among the three species exposed at larval and pupal stages, Ae. aegypti showed maximum reduction in egg laying (29.3-46.6%). Blood feeding was also reduced in females exposed to EI50 dose at larval stage and a positive correlation was demonstrated between the quantity of blood meal taken and the proportion of eggs laid. Significant reduction in the quantum of blood ingested by the treated females may be responsible for the reduced egg laying. PMID- 10695812 TI - A cellular automaton model of cellular signal transduction. AB - On the basis of cellular automata models, a software specifically tailored to model biochemical reactions involved in cellular signal transduction was implemented on a personal computer. Recent data regarding desensitization processes in mouse Leydig cells are used to simulate the underlying reactions of signal transduction. Pretreatment of real Leydig cells with different molecules results in a modification of the signal transduction cascade leading to a diminished response of the cells during subsequent stimulations. The model is capable of simulating the complex behavior of this intracellular second messenger production in a qualitative and semi-quantitative way. The results indicate that quantitative simulations on a molecular level will be possible once appropriate computer hardware is available. The simulations and results of the cellular automaton presented are easily described and comprehended, which make it a useful tool that will facilitate research in cellular signal transduction and other fields covering complex reaction networks. PMID- 10695813 TI - Automated segmentation of multiple sclerosis lesions in multispectral MR imaging using fuzzy clustering. AB - A method is presented for fully automated detection of Multiple Sclerosis (MS) lesions in multispectral magnetic resonance (MR) imaging. Based on the Fuzzy C Means (FCM) algorithm, the method starts with a segmentation of an MR image to extract an external CSF/lesions mask, preceded by a local image contrast enhancement procedure. This binary mask is then superimposed on the corresponding data set yielding an image containing only CSF structures and lesions. The FCM is then reapplied to this masked image to obtain a mask of lesions and some undesired substructures which are removed using anatomical knowledge. Any lesion size found to be less than an input bound is eliminated from consideration. Results are presented for test runs of the method on 10 patients. Finally, the potential of the method as well as its limitations are discussed. PMID- 10695814 TI - Volumetric measurements of right cerebral hemisphere infarction: use of a semiautomatic MRI segmentation technique. AB - The applications of a new segmentation software, Anatomatic, in the evaluation of volumetric measurements of brain infarctions and the new Medimag 3D software in the evaluation of 3D image representation of infarctions are described. These programs are applied to magnetic resonance imaging. The aim of this study is to evaluate the use of these software packages in making accurate volumetric measurements in 40 patients with right cerebral infarctions, in determining the correlations between the quantitated lesions and neurological/neuropsychological dysfunctions and in creating realistic 3D views of the infarctions. Using Anatomatic, reproducible infarction volumes were achieved with ease and within a reasonably fast time. Medimag helped achieve realistic 3D representations of the infarctions. When compared, the semiautomatic segmentation proved to be much faster and yielded higher infarction volumes than the manual segmentation technique. Significantly positive correlations between the infarction volumes and neurological dysfunctions and neuropsychological deficit (neglect) helped to explain the effect of volumes on the clinical status of the patients. PMID- 10695815 TI - Diagnosis, disclosure, and informed consent: learning from parents of children with cancer. AB - PURPOSE: The aim of this study was to learn about and to describe retrospective perceptions of parents of the circumstances of their child's cancer diagnosis and of the informed consent process. METHODS: Professional moderators conducted three focus groups with 22 parents of children with cancer who were eligible for enrollment in a Children's Cancer Group clinical trial research protocol. Each focus group consisted of seven to nine parents and was audiotaped and transcribed. RESULTS: Parents' descriptions of the early phase of their child's illness yielded the following themes: dialogues regarding the diagnosis and treatment options occurred amidst tremendous stress; a sense of constraint and lack of control were common; parents experienced variable degrees of choice regarding their child's participation in a clinical trial; and parents provided suggestions about how to improve the informed consent process. Overall, parents did not verbalize distinctions between their understanding of their child's medical treatment, research participation, and other aspects of their child's cancer experience. CONCLUSIONS: Based on these results, the authors conclude with practical recommendations for health care professionals caring for children with cancer and call for future research about parents' understanding of treatment options, the nature of clinical trials, and experience with the diagnostic and early treatment phase of childhood cancer with larger samples of parents from multiple sites. PMID- 10695816 TI - Immune reconstitution after autologous purged bone marrow transplantation in children. AB - PURPOSE: Immune reconstitution was studied in 30 children who had received purged autologous bone marrow transplantation for neuroblastoma or acute myeloid leukemia (AML). METHODS: Patients with neuroblastoma received high-dose chemotherapy and total body irradiation, and patients with AML received chemotherapy alone. Marrows were purged ex vivo with either antineuroblastoma monoclonal antibodies (neuroblastoma) or 4-hydroperoxycyclophosphamide (AML). Lymphocyte subsets, mitogen stimulation studies, and immunoglobulin levels were studied every 4 months. RESULTS: There were no significant differences between the two groups of patients in lymphocyte number or subsets over time. In both groups, CD2+ and CD4+ cells were below normal in 33% of patients at 12 months. CD4+/CD8+ ratios were below normal for up to 8 months after transplantation and natural killer cells were elevated for up to 2 years in most patients. Median IgG and IgA levels were below the age mean even at 2 years after transplantation, although patients with AML had significantly higher IgG levels at 12 months compared with those with neuroblastoma. Lymphocyte proliferative responses to mitogens were markedly reduced at 4 months but returned to normal at 8 months. Despite the delay in immune reconstitution, there were no life-threatening infections. CONCLUSIONS: There appeared to be little difference in the overall kinetics of immune reconstitution between the children with neuroblastoma, who received total body irradiation and high-dose chemotherapy, and those with AML, who received high-dose chemotherapy alone as their pretransplant preparative regimen. PMID- 10695817 TI - A new sensitive and specific combination of CD81/CD56/CD45 monoclonal antibodies for detecting circulating neuroblastoma cells in peripheral blood using flow cytometry. AB - PURPOSE: Intensive chemoradiotherapy followed by peripheral blood stem cell transplantation has been introduced to treat children with advanced neuroblastoma (NBL). Detection of NBL cells in peripheral blood (PB) is important to prevent reinfusion of NBL cells. Several immunologic methods have been proposed for detecting NBL cells in hematologic samples. The development of a sensitive and specific combination of monoclonal antibodies (MoAbs) for detecting small numbers of NBL cells in PB using flow cytometry remains an important challenge. METHODS: Twenty-one clinical samples from NBL tissues or smears containing NBL cells were examined for reactivity against CD81, CD56, and CD9 using an immunocytochemical technique. The expressions of CD81, CD56, CD9, and antihuman disialoganglioside GD2 MoAb (GD2) in five NBL cell lines were assayed by flow cytometry. For the evaluation of sensitivity, five NBL cell lines were added to normal PB and the detection level of the combination of CD81/CD56/CD45 MoAbs was compared with that of CD9/CD56/CD45 MoAbs (reported previously). One hundred thirty-three normal PB samples were examined to determine the sensitivity and specificity of this method. RESULTS: All NBL cell lines showed strong positivity with CD81 and CD56 MoAb. However, CD9 MoAb was weakly positive against the five NBL cell lines. GD2 MoAb reacted strongly with four NBL cell lines, although almost the entire cell population of the SK-N-SH NBL line failed to bind the GD2 MoAb. In vitro experiments using NBL cell lines demonstrated that tumor cells added to normal PB cells could be detected by flow cytometry using CD81/CD56/CD45 MoAbs even at a concentration of 0.005%. Through comparative studies, the combination of CD81/CD56/CD45 MoAbs was found to be more sensitive and specific than that of CD9/CD56/CD45 MoAbs for detecting small numbers of NBL cells using the above cell lines. CONCLUSIONS: Triple-color flow cytometric analysis using CD81/CD56/CD45 MoAbs is useful for detecting NBL cells in PB. Further studies testing this approach using samples of PB with NBL contamination are needed to test this approach in patients. PMID- 10695818 TI - Early testicular biopsy in males with acute lymphoblastic leukemia: lack of impact on subsequent event-free survival. AB - PURPOSE: Children with acute lymphoblastic leukemia (ALL) who had bulky disease (lymphomatous features) at diagnosis had the highest rate of testicular relapse (20%) of any ALL subgroup on previous Children's Cancer Group (CCG) studies in the late 1980s. To limit curative, but sterilizing, testicular irradiation to those with testicular disease, testicular biopsies were performed to detect occult testicular disease within the first 6 months of treatment. Testicular irradiation then was provided to those with occult disease to increase disease free survival. Identification of those with occult disease was believed to be a factor that would influence ultimate survival in such patients in that era. PATIENTS AND METHODS: One hundred ninety-nine patients had bilateral testicular wedge biopsies performed during the first maintenance therapy phase of the four different chemotherapy regimens. Patients with positive biopsy results were treated with testicular irradiation and continued on therapy. RESULTS: Eleven of 199 biopsy results (5.5%) were judged positive. Patients with positive biopsy results given testicular radiation had a 45% subsequent adverse event rate, compared with 36% for those with a negative biopsy results (P = 0.4). The survival rates for the two groups were similar. The low rate of positive biopsy specimens resulted in discontinuation of the procedure before closure of the study. CONCLUSION: Positive testicular biopsy results early in remission identified patients at a slightly higher risk of subsequent adverse events but did not influence survival. However, because negative biopsy results (94.5%) did not alter the prescribed treatment, the small number of positive biopsy results did not warrant undertaking the procedure in most male patients with ALL, and this procedure was abandoned. PMID- 10695819 TI - Extramedullary myeloid tumors in children: the limited value of local treatment. AB - PURPOSE: To determine the incidence of extramedullary tumors (EMT) in Saudi Arabian children with acute myeloid leukemia, the factors associated with these tumors and the impact of local treatment on local tumor control, complete remission and survival rates. PATIENTS AND METHODS: One hundred children, median age 6 years, who received their primary treatment for acute myeloid leukemia at King Faisal Specialist Hospital and Research Center, from 1983 to 1997 were studied. EMT at diagnosis occurred in 18 (18%) patients at 25 sites. Meningeal leukemia, hepatosplenomegaly, lymph node enlargement, gingival hypertrophy, and cutaneous infiltration were not included in the definition of EMT. With these exclusions, children with EMT were younger than those without EMT (median age, 3.5 v. 7.5 years) and were more likely to have meningeal leukemia at diagnosis (33% v. 10%). The t(8;21) translocation was associated with a 47% EMT incidence compared with 23% without the translocation. Local radiation treatment was given to 16 of 25 (64%) EMT sites. RESULTS: The overall 5-year survival rate for all patients was 28%, and this was not significantly influenced by the drug regimen used, meningeal leukemia at diagnosis, the presence of the (8;21) translocation, M4 and M5 morphology combined, or EMT at diagnosis. Significant differences were observed in the 5-year survival rates for patients who underwent allogeneic bone marrow transplantation (52%; N = 37) and those who attained complete remission (CR) but did not undergo transplantation (21%; N = 44) and those who did not achieve complete remission with initial therapy (5%; N = 19). Systemic and local EMT CR was achieved in 17 of 18 patients with EMT, including 12 patients who underwent radiation treatment and 5 of 6 of those who did not. Isolated relapse was not seen at an EMT site and was not noted at any later stage of the disease. CONCLUSIONS: Permanent local control at sites of EMT was achieved in all patients who attained a bone marrow CR, whether or not the site was irradiated. Local radiation treatment of an EMT site did not appear to contribute to overall CR and survival rates. The use of radiation treatment should be conservative and limited to patients in whom there is a real and immediate threat to vision or renal function or when the spinal cord is compromised. PMID- 10695820 TI - Oral methotrexate for recurrent brain tumors in children: a Pediatric Oncology Group study. AB - PURPOSE: Children with recurrent or progressive central nervous system (CNS) tumors have an unfavorable prognosis. Based on Pediatric Oncology Group (POG) institutional pilot data, low-dose oral methotrexate (MTX) was studied. METHODS: Eight dosages of MTX 7.5 mg/m2 every 6 hours were administered on a weekly schedule for as long as 18 months. Patients in six different brain tumor strata were accrued. RESULTS: The response rates (complete or partial responses) were as follows: astrocytoma 2 of 10, malignant glioma 1 of 19, medulloblastoma 0 of 18, brainstem tumor 0 of 12, ependymoma 1 of 7, and miscellaneous histologic types 0 of 12. The main toxicities, mucositis, myelosuppression, and hepatic transaminase elevation were considered tolerable. CONCLUSION: Low-dose oral MTX showed no significant activity against malignant glioma, medulloblastoma, brainstem tumors, and miscellaneous histologic types. Indeterminate but low response rates were observed in children with astrocytoma and ependymoma. This regimen will not be recommended for front-line therapy. PMID- 10695821 TI - Increased expression of lung resistance-related protein and multidrug resistance associated protein messenger RNA in childhood acute lymphoblastic leukemia. AB - Immunophenotype might be an important indicator for multidrug resistance (MDR) profiles in childhood acute lymphoblastic leukemia (ALL). The authors analyzed the messenger RNA (mRNA) levels of MDR1, multidrug resistance-associated protein (MRP), and lung resistance-related protein (LRP) by reverse transcriptase polymerase chain reaction (RT-PCR) in childhood pre-B ALL, T-cell ALL, and acute nonlymphoblastic leukemia (ANLL). Results showed that MRP and LRP, but not MDR1, mRNAs are overexpressed, particularly in children with pre-B ALL compared with T cell ALL and ANLL tested. In addition, the MRP and LRP mRNA expression levels in initial diagnosis and first relapse samples of one patient with pre-B ALL were similar. Consequently, these preliminary results suggest that the expression of these MDR-related genes in childhood ALL might be regulated differently in a lineage dependent manner. PMID- 10695822 TI - Treatment of iron deficiency anemia and associated protein-losing enteropathy in children. AB - PURPOSE: The aims of this study were to evaluate the response of oral iron treatment in children with iron deficiency anemia (IDA) fed whole cow's milk (WCM) or soy formula; to compare the incidence of fecal blood loss in infants fed WCM and soy formula; and to evaluate the incidence and relation of protein-losing enteropathy (PLE) and IDA by testing serum albumin, fecal blood loss, and fecal alpha1-antitrypsin (alpha1AT). METHODS: Twenty-four children with nutritional IDA were randomly assigned to receive either 16 oz WCM or soy formula daily. Both groups were treated with daily therapeutic oral iron during 12 weeks. Stool specimens for hemoglobin losses were collected at weeks 0, 3, 6, and 12. Levels of serum albumin and fecal alpha1AT were tested at diagnosis and when IDA was corrected. RESULTS: Anemia was corrected in 21 of the 24 children by week 6 or 12. Median fecal hemoglobin losses were not increased in either group at diagnosis or during treatment. Seven of 24 children had PLE at diagnosis with elevated fecal alpha1AT levels of 72 to 381 mg/dL that returned to normal after correction of IDA. Their initial fecal alpha1AT levels averaged 170 mg/ dL at diagnosis and 21 mg/dL after the IDA was corrected. Excessive WCM intake of 30 oz/day or more was present in 63% of the infants. CONCLUSIONS: Treatment of nutritional IDA with oral iron was just as effective with a limited quantity of either WCM or soy formula. Fecal hemoglobin losses were uncommon and did not differ in children at diagnosis or during treatment of IDA. PLE associated with IDA resolves when the IDA is corrected, but differences between children fed WCM or soy formula could not be detected. PMID- 10695823 TI - Self-reported diagnostic and management strategies in childhood idiopathic thrombocytopenic purpura: results of a survey of practicing pediatric hematology/oncology specialists. AB - PURPOSE: To assess current physician self-reported practices regarding initial management of childhood idiopathic thrombocytopenic purpura (ITP) and to determine physician self-reported willingness to participate in randomized clinical trials comparing different initial management strategies. PATIENTS AND METHODS: A questionnaire was mailed in November 1997 to all 720 members of the American Society of Pediatric Hematology/Oncology asking how they would diagnose and manage ITP in children 18 months, 5 years, and 15 years of age who were experiencing either dry purpura (cutaneous hemorrhage only) or wet purpura (active mucous membrane hemorrhage). Specific questions dealt with bone marrow aspiration, hospital admittance, treatment strategy, and specific doses of corticosteroids and intravenous immunoglobulin. RESULTS: The response rate to the questionnaire was 57%. Most respondents indicated they usually perform a bone marrow aspirate when corticosteroids are to be prescribed and administer drug therapy to patients with newly diagnosed ITP with wet or dry purpura. Only 16% of respondents would administer no drug therapy to a child with dry purpura. Intravenous immunoglobulin (IVIG) was preferred to steroids, with anti-D immunoglobulin prescribed less frequently. Hospital admittance often was used for patients with dry purpura and usually recommended for patients with wet purpura. Most respondents expressed willingness to randomize patients with dry purpura to IVIG versus no therapy and those with wet purpura to IVIG versus prednisone as part of a randomized controlled clinical trial. CONCLUSIONS: The self-reported care of the patient with ITP was influenced by the severity of presentation (dry versus wet purpura). Most physicians reported they would administer specific drug treatment in both scenarios. This survey illustrates the diverse diagnostic and management strategies currently used in childhood ITP. Because no one therapeutic approach is predominant and a scientific basis for decision making in childhood ITP has not been developed, future randomized trials are warranted. On the basis of these survey results, such trials are desired by most pediatric hematology/oncology specialists. PMID- 10695824 TI - Hematologic toxicity of sodium valproate. AB - PURPOSE: Sodium valproate is a commonly used anticonvulsant in the management of childhood refractory epilepsy with good response rates and acceptable toxicity. Hepatotoxicity is the most widely recognized toxicity. With the use of higher drug levels to achieve adequate seizure control, hematologic toxicity is being increasingly encountered, and the pediatric hematologist is consulted for these problems in the pre- or perioperative setting. The purpose of this article is to characterize the various hematologic toxicities encountered in a clinical setting and to provide guidelines to assist in the management of these patients. METHODS: A literature review was undertaken to identify the hematologic toxicities of valproate used as monotherapy or polytherapy. Key words used in the search were valproate, hematology, and bleeding. RESULTS: Valproate can cause direct bone marrow suppression leading to aplastic anemia or peripheral cytopenia affecting one or more cell lines. Occasional fatal bone marrow failure, myelodysplasia, and a clinical picture resembling acute promyelocytic leukemia have also been seen. Thrombocytopenia, macrocytosis, neutropenia, and pure red cell aplasia can occur but are not reported to be life-threatening. A bleeding diathesis associated with valproate use may include thrombocytopenia, abnormal platelet function, and acquired von Willebrand disease type I. CONCLUSIONS: Hematologic toxicities of valproate are common, vary in onset and severity, are recurrent, transient, or persistent, and usually occur with a serum valproate level greater than 100 microg/mL. In most situations, even when highly clinically significant, they can be reversed with dosage reduction; drug discontinuation is rarely required. Potential adverse effects such as thrombocytopenia and leukopenia are easily detected by laboratory monitoring, which should be continued indefinitely at least on a quarterly basis. Caution for elective surgery is advised; preoperative coagulation studies should be done, including platelet function studies and von Willebrand factor levels. Perioperative use of DDAVP to increase von Willebrand factor levels and improve platelet function is appropriate in some cases. PMID- 10695825 TI - Prothrombotic abnormalities in children with venous thromboembolism. AB - PURPOSE: The aim of this study was to determine the frequency of acquired or inherited prothrombotic disorders in a pediatric population with venous thromboembolism (VTE). PATIENTS AND METHODS: From May 1992 to April 1998, 56 consecutive children with VTE were prospectively studied at a single center. RESULTS: The median age was 8.4 years (range, 0.1-18 years). There was a male predominance. Fifty (89%) children had thrombosis in the lower venous system. Risk factors were detected in 54 (96%) children. Twenty-one (38%) thrombotic episodes were related to central venous lines. Family history of thrombosis was positive in 13 (23%) patients. In 26 (46%) patients, a prothrombotic disorder was detected. Nine of them had inherited disorders (protein C deficiency, 5 patients; protein S deficiency, 3 patients; Factor V Leiden mutation, 1 patient), and 13 children had acquired disorders (antiphospholipid antibodies, 5 patients; antithrombin deficiency, 8 patients). The remaining four showed combined abnormalities (Factor V Leiden mutation associated with inherited protein S deficiency, 1 patient; acquired antithrombin deficiency, 2 patients and inherited antithrombin deficiency, 1 patient). CONCLUSIONS: In the series, a high percentage of prothrombotic disorders was detected; thus, a complete hemostatic evaluation should be performed in all of the children with VTE whether the patients have one or more risk factors. PMID- 10695826 TI - Osteosarcoma associated with absent thumbs: a report of two cases. AB - An 8-year-old Hispanic boy with a hypoplastic left thumb, absent right thumb, and short stature experienced right leg pain and limp. A right tibial lesion was imaged and found to be osteosarcoma on biopsy. A 6-year-old Hispanic girl with congenitally absent thumbs experienced a pathologic fracture of her left femur after a minor sports injury. The radiologic abnormality seen was diagnosed as osteosarcoma on biopsy. Both patients continue to do well after intensive preoperative and postoperative high-dose chemotherapy and definitive reconstructive limb surgery. Osteosarcoma has been linked to several congenital syndromes in which absent thumbs are a feature. These two patients with absent thumbs and no definable syndrome experiencing osteosarcoma suggest that congenitally absent thumbs might be a risk factor for osteosarcoma in the absence of a syndrome. PMID- 10695827 TI - Renin-producing hepatoblastoma. AB - Renin-producing tumors of extrarenal origin are rare in children. An 8-year-old boy with hepatoblastoma and hypertension associated with a high plasma renin level is reported. After chemotherapy, the plasma renin level normalized and the hypertension spontaneously resolved. The patient underwent surgery, and a right trisegmentectomy of the liver and a partial resection of the second and third segments were performed. The tumor was as shown the source of renin by immunohistochemical study and reverse transcriptase-polymerase chain reaction. PMID- 10695828 TI - Coexistence of lymphoblastic and monoblastic populations with identical mixed lineage leukemia gene rearrangements and shared immunoglobulin heavy chain rearrangements in leukemia developed in utero. AB - Congenital leukemia often provides insight into mechanisms of in utero leukemogenesis. A 10-day-old boy with clinical features of skin nodules, marked hepatosplenomegaly, and subcutaneous bleeding received a diagnosis of congenital leukemia. This patient initially had a dominant B progenitor lymphoblast population and minor monocyte component. Treatment with prednisolone, vincristine, and doxorubicin resulted in a loss of lymphoblast population and a rapid increase and dominance of the monocyte component within 10 days. Complete remission initially was obtained with additional combination chemotherapy with epipodophyllotoxin (VP-16) and cytosine arabinoside (Ara-C), but relapse characterized by a lymphoblastic population in the bone marrow was subsequently observed. The authors hypothesize that the leukemic cells originated from a common B-monocyte lineage stem cell during fetal hematopoiesis. PMID- 10695829 TI - Detecting neuroblastoma using bone marrow aspiration and bone marrow biopsy. PMID- 10695830 TI - Extrarenal Wilms tumor of the left ovary: a case report. PMID- 10695831 TI - Turner syndrome and ganglioneuroma. PMID- 10695832 TI - Wide range of primary liver tumors can be found in patients with familial adenomatous polyposis. PMID- 10695833 TI - Parental psychopathology and reports of the childhood home environment in adults with early-onset dysthymic disorder. AB - In previous studies, patients with dysthymic disorder (DD) have reported significantly more adverse early home environments than patients with episodic major depressive disorder (MDD) and normal controls. However, DD is also associated with increased rates of mood and personality disorders in first-degree relatives, raising the possibility that the DD-early adversity relationship may be due to the confounding effects of parental psychopathology. The present study addressed this issue using a sample of 97 adult outpatients with early-onset DD, 45 adult outpatients with episodic MDD, and 45 normal controls, and their first degree relatives. The early home environment was assessed with semi-structured interviews and self-report inventories. Parental psychopathology was assessed using semi-structured direct and family history interviews, and diagnoses were assigned using the best-estimate procedure. Results indicated that parental mood and personality disorders were strongly associated with probands' reports of early adversity. However, patients with DD continued to differ significantly from patients with episodic MDD and normal controls after controlling for parental psychopathology. PMID- 10695834 TI - Dissociative symptomatology and traumatogenic factors in adolescent psychiatric patients. AB - This study describes the relation of different types of childhood trauma to the degree of dissociative experiences. Subjects were 198 consecutively admitted adolescent psychiatric patients, 11 to 19 years old (89 inpatients and 109 outpatients). All patients completed the Adolescent Dissociative Experiences Scale. A Checklist of Traumatic Childhood Events was filled out by clinicians. The results showed an increase in the degree of dissociative experiences in patients with a history of sexual abuse, physical abuse, neglect, and stressful life events. With the exception of life events, a moderate form of traumatic experience had the same effect on dissociative experiences as severe forms. The strongest effect was found for emotional neglect, which seems to be an important pathogenic risk factor. The results suggest that therapists and researchers should be aware that even less severe forms of abuse and neglect may have a significant impact on the development of dissociative psychopathology in adolescents. PMID- 10695835 TI - Obsessive and compulsive symptoms in schizophrenia: clinical and neurocognitive correlates. AB - Although research suggests that the presence of obsessive and compulsive symptoms in schizophrenia is associated with graver levels of psychosocial dysfunction, it is unclear whether it is also related to clinical features of illness. Accordingly, the present study compared the symptom levels and neurocognitive function of participants with schizophrenia who had and did not have significant obsessive or compulsive symptoms. Analyses of variance revealed that participants with significant levels of either obsessive or compulsive symptoms (N = 21) had higher levels of positive and emotional discomfort symptoms on the Positive and Negative Syndrome Scale (PANSS) and performed more poorly on the Wisconsin Card Sorting Test, a measure of executive function, than participants without obsessions or compulsions (N = 25). ANCOVAs controlling for level of obsessions also revealed that participants with significant levels of compulsions (N = 12) in particular had higher levels of negative and positive symptoms on the PANSS than participants without compulsions (N = 34). The impact of obsessive compulsive phenomena on the course and outcome of schizophrenia is discussed. PMID- 10695836 TI - Soft neurological signs in schizophrenic patients and their nonpsychotic siblings. AB - This study examined neurological signs as familial vulnerability factors to schizophrenia. Fifteen Chinese schizophrenic patients, 21 of their nonpsychotic siblings, and 26 healthy volunteers, matched for age, sex, and education, were assessed by using the Cambridge Neurological Inventory. Results showed that patients and their siblings had significantly higher global neurological impairment than controls. The severity of motor coordination impairment of the siblings was in between patients and controls. This may suggest either patients have higher genetic vulnerability for the neurological abnormality and schizophrenia than their nonpsychotic siblings or the disease can further worsen the preexisting neurological deficit. Disinhibition signs were similar in patients and siblings, but significantly less in controls indicating its familial nature. Extrapyramidal and sensory integration signs were similar in siblings and controls, but significantly more severe in patients, suggesting nonfamilial abnormalities. In conclusion, these findings may imply different etiological origins for different subgroups of neurological signs. PMID- 10695837 TI - Effects of time in the United States and Indian ethnicity on DSM-III-R psychiatric disorders among Mexican Americans in California. AB - The study examines the effects of time in the United States and Indian ethnicity on prevalence of 12 DSM-III-R psychiatric disorders among Mexican Americans in California. In Fresno County, primarily an agricultural area, 3012 participants of Mexican origin (18 to 59 years) were selected under a cluster sampling design and interviewed using a version of the World Health Organization's Composite International Diagnostic Instrument (WHO-CIDI). Lifetime prevalence of any psychiatric disorder was 46.4% for Indians and 32.9% for non-Indians. Alcohol dependence was the most prevalent disorder (Indians = 17.4%, non-Indians = 10.7%). Indians had significantly higher risk of affective disorders (adjusted OR = 2.9) and drug abuse/dependence (adjusted OR = 2.6) compared with non-Indians. Time in the United States was associated with higher risk of lifetime affective disorders and drug abuse/dependence. This effect was more pronounced among Indians. Mexican immigrants are ethnically heterogenous and Indians appear to be more vulnerable to negative effects of exposure to U.S. society. PMID- 10695838 TI - Factors associated with compliance of alcoholics in outpatient treatment. AB - Predictors of compliance with psychosocial treatment were examined in a clinical sample of 150 patients seeking outpatient treatment for alcohol problems. Predictor variables were as follows: a) patient characteristics at the beginning of treatment and b) treatment characteristics of a matching model developed from the Conceptual System Theory. This model was based on the conceptual level of patients and therapists, and on the structure of the treatment. Using logistic regression analysis, we identified predictors of compliance in the motivation phase (first four consultations) and the active treatment phase (the remainder of the agreed treatment period of 12 months). Predictors of compliance in the motivation phase were a) therapists with a high conceptual level, characterized by an attitude toward the patients of empathetic and reflective listening; b) disulfiram; and c) psychopharmacological treatment. The only patient-related predictors of compliance in the active treatment phase were a) age below 40 and b) the presence of family and social problems, both factors predicting noncompliance. The only treatment-related predictor variables were a) the matching of patients with the structure of treatment and b) disulfiram during the first month of treatment, both factors predicting compliance. Two years after the beginning of treatment, those patients who completed the motivation phase as well as the active treatment phase had a significantly better psychosocial function than the noncompliant patients. PMID- 10695839 TI - Atypical Charles Bonnet hallucinations: an elf in the woodshed, a spirit of evil, and the cowboy malefactors. AB - In this article, the authors review the literature on the Charles Bonnet syndrome (CBS), a condition involving visual hallucinations in normal persons with severe sight loss. Attempts to assess the characteristics of this "phantom vision" have resulted in descriptions of a "typical" CBS hallucination. However, the many exceptions to a modal CBS experience cause the authors to postulate two other general categories of CBS hallucinations: a) the atypical sensory/perceptual (ASP), and b) the atypical psychodynamic (APD) hallucinations. Case studies illustrating these two types are provided. Extending the categories of Charles Bonnet hallucinations allows for more specific diagnosis, treatment, and may result in the possibility of greater precision in future research. PMID- 10695840 TI - Prediction of work performance by clinical symptoms and cognitive skills in schizophrenic outpatients. PMID- 10695841 TI - Epidemiology of schizophrenia in northeast Brazil. PMID- 10695842 TI - Societal alternatives to anabolic steroid use. PMID- 10695843 TI - Drugs, sport, and the new millennium. PMID- 10695844 TI - Concussions during the 1997 Canadian Football League season. AB - OBJECTIVE: To examine the incidence and characteristics of concussions for one season in the Canadian Football League (CFL). DESIGN: Retrospective survey. PARTICIPANTS: 289 players reporting to CFL training camp. Of these, 154 players had played in the CFL during the 1997 season. MAIN OUTCOME MEASURES: Based on self-reported symptoms, calculations were made to determine the number of concussions experienced during the previous season, the duration of symptoms, the time for return to play after concussion, and any associated risk factors for concussions. RESULTS: Of all the athletes who played during the 1997 season, 44.8% experienced symptoms of a concussion. Only 18.8% of these concussed players recognized they had suffered a concussion. 69.6% of all concussed players experienced more than one episode. Symptoms lasted at least 1 day in 25.8% of cases. The odds of experiencing a concussion increased 13% with each game played. A past history of a loss of consciousness while playing football and a recognized concussion while playing football were both associated with increased odds of experiencing a concussion during the 1997 season. CONCLUSION: Many players experienced a concussion during the 1997 CFL season, but the majority of these players may not have recognized that fact. Players need to be better informed about the symptoms and effects of concussions. PMID- 10695845 TI - The efficacy of Farabloc, an electromagnetic shield, in attenuating delayed-onset muscle soreness. AB - OBJECTIVE: To assess the hypothesis that Farabloc, a fabric with electromagnetic shielding properties, would attenuate the symptoms, signs, and muscular strength deficit secondary to delayed-onset muscle soreness (DOMS) induced by two exposures to eccentric exercise in humans. DESIGN: Randomized, single-blind, placebo-controlled, crossover trial with two testing stages of 5 days duration separated by a washout period of more than 8 weeks. SETTING: University-based sports medicine center. PARTICIPANTS: Twenty volunteers equally representing untrained male and female subjects. INTERVENTIONS: 20 sets of 10 repetitions of single-leg eccentric knee extensions for 37 minutes with the Biodex dynamometer set at 30 degrees per second were performed on the first day of stage one and stage two to induce DOMS in the quadriceps muscle. Double layers of fabric, either Farabloc or placebo, were wrapped around the thigh of each participant during each stage for 5 days. MAIN OUTCOME MEASURES: Perception of muscle pain, as measured by a visual analog scale (VAS), and strength, as measured by knee extensor torque (EST) with the Biodex dynamometer, were evaluated at 0, 24, 48, 72, and 96 hours. Serum inflammatory markers of muscle damage, including malondialdehyde. creatine phosphokinase, myoglobin, leukocytes, and neutrophils, were assayed at 0, 2, 6, 24, and 48 hours. RESULTS: Repeated-measures analysis of variance was carried out for each of the seven variables to assess differences for fabric, order of treatment, time, and all combinations. Results of VAS and EST and levels of malondialdehyde, creatine phosphokinase, myoglobin, leukocytes, and neutrophils all showed a highly significant effect of Farabloc compared with placebo. This analysis shows that the order of Farabloc or placebo fabric use in stage I and 2 produces different results. This may be caused by a learning effect, but did not alter the overall influence of Farabloc. CONCLUSION: The data indicate that double layers of Farabloc fabric wrapped around the thigh reduces pain and strength loss and serum levels of malondialdehyde, creatine phosphokinase, myoglobin, leukocytes, and neutrophils when untrained human subjects are exposed to eccentric exercise to produce DOMS in the quadriceps. Farabloc shields high-frequency electromagnetic fields, although the results do not indicate how these changes are mediated. Further research is needed to determine the mechanism. PMID- 10695846 TI - Anterior knee pain: a clinical comparison of rehabilitation methods. AB - OBJECTIVE: To compare different types of rehabilitation for anterior knee pain. DESIGN: Prospective, randomized, blinded, and controlled study of 64 participants with anterior knee pain. SETTING: Outpatient rehabilitation clinic and testing laboratory. PARTICIPANTS: Participants were assigned in randomized fashion to three rehabilitation groups: traditional home rehabilitation (n = 20); physical therapy (n = 21); and home rehabilitation with a modified vastus medialis obliquis (VMO) specific straight leg raise (Muncie method; n = 23). INTERVENTIONS AND MAIN OUTCOME MEASURES: Clinical data was obtained at 0, 2, 6, and 12 weeks. Cybex testing was performed at 0, 6, and 12 weeks. RESULTS: Clinical outcome for the Muncie method indicated a statistically significant improvement in subjective pain and functional impairment ratings. Cybex testing in patients using the Muncie method demonstrated a statistically significant improvement in pain-free isometric contractions and maximum voluntary contraction. There were no significant differences between traditional home therapy and physical therapy. CONCLUSION: Findings suggest that the Muncie method results in improved clinical outcome at a lower cost than traditional home and physical therapy and possibly improved VMO/quadriceps muscle balance. Patients with anterior knee pain may benefit from applying the Muncie method in a home therapy program. PMID- 10695847 TI - Professional roller hockey injuries. AB - OBJECTIVE: To study the incidence and types of injuries sustained by professional roller hockey players in practices and games, and to compare these statistics with those from ice hockey. DESIGN: This injury survey used a strict definition of injury, standardized reporting strategies, and diagnosis by a team physician as standards by which to analyze the characteristics of roller hockey injuries. SETTING: The injuries were recorded after the players had been examined by a team physician at the game or practice site or in the physician's office. PARTICIPANTS: During three seasons for one roller hockey team and one season for another team, an average of 22 players per team participated in the study. Due to personnel changes, the team rosters were modified between seasons. Each player injury was included in the study. An injury was defined as any physical impairment caused during a practice or game that eliminated the player from that practice or game or the next day's practice session or contest, or any physical ailment that necessitated a physical examination by the team physicians. MAIN OUTCOME MEASURE: Injury data were categorized and injury rates were calculated. RESULTS: 122 injuries were recorded during four professional roller hockey seasons, resulting in an overall participation injury rate of 14.4 per 1,000 player hours. The game injury rate was 304.9 per 1,000 player hours. The players were 105.1 times more likely to be injured during a game than during practice. Preseason practices produced 4.5 times more injuries than regular season practices. In comparison, sample data from the only other published study of roller hockey injuries and from several studies of ice hockey have indicated game injury rates of 139.0 (roller hockey), 119.0, 96.1, 78.4, 78.8, and 66.0 per 1,000 player hours, respectively. CONCLUSION: Results of this study demonstrate that roller hockey produces a higher rate of both contact and noncontact injuries than ice hockey; this contradicts the findings of the only other published research study on injuries in roller versus ice hockey. This increased incidence of injury may be due in part to the differences in surfaces, and can prove hazardous to even the recreational roller hockey player or in-line skater. PMID- 10695848 TI - The efficacy of magnetic resonance imaging in acute knee injuries. AB - OBJECTIVE: To evaluate the clinical efficacy of magnetic resonance imaging (MRI) of the knee in acute injuries with indeterminate clinical findings, using arthroscopy as a gold standard. DESIGN: A prospective double-blind study was performed. All patients underwent MRI on a 1.5 T magnet using dual spin echo pulse sequences. This was followed by arthroscopy. SETTING: Tertiary care referral center. PATIENTS: Twenty-three patients with an average age of 26 years satisfied the study criteria. Patients had to have been seen by one of two orthopaedic surgeons within 6 weeks of sudden trauma to the knee complicated by a hemarthrosis, clinical assessment of which was equivocal. RESULTS: The respective sensitivity and specificity for MRI of the knee were 90% (18/20) and 67% (2/3) for detecting any anterior cruciate ligament injury, 50% (1/2) and 86% (18/21) for detecting medial meniscal tears, and 88% (7/8) and 73% (11/15) for detecting lateral meniscal tears. MRI also identified injuries that could not be assessed on arthroscopy, including 14 bone bruises, five posterior cruciate ligament tears, nine medial collateral ligament tears, and one lateral collateral ligament tear. The detection of composite injury requiring surgical intervention yielded a sensitivity of 100% (16/16) and a specificity of 71% (5/7). Prospective use of MRI evaluation of the knee could have prevented 22% (5/23) of diagnostic arthroscopic procedures. CONCLUSION: Equivocal clinical findings in patients with acute knee injury should lead to use of MRI in an appropriate clinical setting. To our knowledge a prospective study of the efficacy of MRI of the knee in this patient population has not been reported. In the presence of such inclusion criteria, the results of our study support the use of early MRI to guide further surgical management. PMID- 10695849 TI - Personal exercise habits and counseling practices of primary care physicians: a national survey. AB - OBJECTIVE: Regular physical activity can reduce the incidence and prevalence of many chronic diseases. A vast majority of Americans cite their physician as their primary source of information regarding healthy lifestyle decisions. This study was designed to obtain information about the personal exercise behavior and counseling practices of primary care physicians, to evaluate the relationship between their personal and professional exercise practices, and to determine whether physician specialty is associated with these practices. DESIGN: A cross sectional survey was mailed to a randomly selected sample of primary care physicians in the United States. A questionnaire was used to obtain detailed information on the personal exercise habits, counseling practices, and barriers to counseling of these physicians, regarding both aerobic exercise and strength training. PARTICIPANTS: 298 primary care physicians, comprising 84 family practitioners, 79 pediatricians, 58 geriatricians, and 77 internists. MAIN OUTCOME MEASURES: Frequency of physician exercise, exercise counseling, and relationship between these practices. RESULTS: Physicians who perform aerobic exercise regularly are more likely to counsel their patients on the benefits of these exercises, as are physicians who perform strength training. Pediatricians and geriatricians counsel fewer patients about aerobic exercise than family practitioners and internists. Counseling regarding strength training is less common in all physician groups surveyed, and lowest among pediatricians, of whom 50% did not advise these exercises for any of their patients. Inadequate time was noted by 61% and inadequate knowledge and/or experience by 16% of respondents as the major barriers to counseling regarding aerobic exercise. CONCLUSION: Physicians who exercise are more likely to counsel their patients to exercise. Inadequate time and knowledge/experience regarding exercise are the most common barriers to counseling identified. These findings suggest strategies that might increase physician exercise counseling behavior. PMID- 10695850 TI - Use of diagnostic imaging during the 1997 Canada Summer Games. AB - OBJECTIVE: To document use of diagnostic imaging during a multi-sport games to assist in planning for future such competitions. METHODS: Medical records from the 1997 Canada Summer Games and from the Brandon General Hospital were reviewed. All uses of diagnostic imaging were compiled as were results of the imaging examinations. These data were correlated with demographic information. RESULTS: A total of 80 imaging examinations were performed during the 1997 Canada Summer Games. These were mainly plain radiographs (n = 77), with two nuclear medicine examinations and one computed tomography (CT) scan. Ultrasound and magnetic resonance imaging (MRI) were available but not used. Use of imaging examinations correlated well with the risk category of the sports, and was almost identical between female and male athletes; women accounted for 42.5% of the imaging examinations and 42.7% of the participants. CONCLUSION: These data may be helpful in planning for other multi-sport competitions. The mix of sports is of greater predictive value than the ratio of female to male athletes when predicting the demand for diagnostic imaging services. PMID- 10695851 TI - Diagnosis and prevention of hyponatremia at an ultradistance triathlon. AB - OBJECTIVE: To evaluate a method of medical care at an ultradistance triathlon, with the aim of reducing the incidence of hyponatremia. DESIGN: Descriptive research. SETTING: New Zealand Ironman triathlon (3.8 km swim, 180 km cycle, 42.2 km run). PARTICIPANTS: 117 of 134 athletes seeking medical care after the triathlon (involving 650 race starters). INTERVENTIONS: A prerace education program on appropriate fluid intake was undertaken. The number of support stations was decreased to reduce the availability of fluid. A body weight measurement before the race was introduced as a compulsory requirement, so that weight change during the race could be included in the triage assessment. An on site laboratory was established within the race medical tent. MAIN OUTCOME MEASURES: Numbers of athletes and diagnoses, including the incidence of symptomatic hyponatremia (defined as symptoms of hyponatremia in association with a pretreatment plasma sodium concentration [Na] < 135 mmol/L); weight changes; and changes in [Na]. RESULTS: The common diagnoses in the 117 athletes receiving attention were exercise-associated collapse (27%), musculoskeletal complaints (26%), and dehydration (12%). There was a significant reduction in the number of athletes receiving medical care for hyponatremia, from 25 of the 114 athletes who received care in 1997 (3.8% of race starters) to 4 of the 117 athletes who received care in 1998 (0.6% of race starters). Mean weight change among athletes in the 1998 race was -3.1 kg, compared with -2.6 kg in 1997. CONCLUSION: A preventive strategy to decrease the incidence of hyponatremia, including education on fluid intake and appropriate placement of support stations, was associated with a decrease in the incidence of symptomatic hyponatremia. PMID- 10695853 TI - Fractured tibia and fibula during use of "snowblades": implications of the return of nonrelease bindings. PMID- 10695852 TI - Blood volume, aerobic power, and endurance performance: potential ergogenic effect of volume loading. AB - OBJECTIVE: Blood volume (BV) and hemoglobin concentration ([Hb]) play important roles in oxygen transport. Manipulation of both BV and [Hb] can markedly affect systemic oxygen transport and maximal aerobic capacity (VO2max). However, the role of BV in oxygen transport and aerobic performance is not well understood. It has recently been postulated that an acute expansion of BV using plasma volume (PV), independent of changes in [Hb], may represent a potential ergogenic property. Therefore, the primary objective of this review was to determine the potential ergogenic properties of volume loading. DATA SOURCES: An extensive research of Medline and Sport-Discus along with cross-referencing was conducted. Articles were included on the basis of their relevancy to the purpose of this review. Only articles published in English were included in the analysis. STUDY SELECTION: All study designs using human participants were reviewed. DATA EXTRACTION: A systematic analysis of data regarding the effect of BV on the determination of oxygen transport, aerobic capacity, and endurance performance was conducted. Particular emphasis was given to articles that examined BV as the key independent variable. Articles relevant to the evaluation of the impact of BV on aerobic capacity and endurance performance were reviewed according to the strengths of the individual study designs. RESULTS: Seven investigations evaluated the impact of acute changes in BV, independent of changes in [Hb], on VO2max. Of these investigations, three revealed that acute manipulations of BV result in no change or a slight reduction in VO2max. Three investigations revealed a significant improvement in VO2max after acute PV expansion. One investigation revealed a nonsignificant increase in VO2max after acute PV expansion. Seven investigations evaluated the impact of acute changes in BV, independent of changes in [Hb], on endurance performance: two revealed a significant improvement in endurance performance after acute PV expansion, and five revealed an unchanged or reduced endurance performance after acute PV expansion. The majority of investigations showing an improvement in VO2max and/or endurance performance after acute PV expansion were conducted using untrained individuals. CONCLUSION: Volume loading does not result in an improvement in VO2max and/or endurance performance in endurance athletes. Volume loading in untrained individuals may improve VO2max, but does not bring these individuals to the aerobic capacity of endurance-trained athletes. Also, volume loading generally does not lead to improvement in endurance performance, irrespective of its effects on VO2max. PMID- 10695854 TI - Unusual double clavicle fracture in a lacrosse player. PMID- 10695855 TI - Pes anserinus syndrome and osteoid osteoma. PMID- 10695856 TI - On-field management for the injured football player. PMID- 10695857 TI - Academic medicine careers. PMID- 10695858 TI - Viscosupplementation for osteoarthritis. AB - Viscosupplementation therapy can restore the elastic and viscous properties of synovial fluid and thus recreate the intra-articular joint homeostasis that is disrupted in the degenerative joint. Hyaluronan (hyaluronic acid) products have been developed and used for viscosupplementation therapy in osteoarthritis. Viscosupplementation treatments using these products are well tolerated. Because viscosupplementation therapy is based on the concept of replenishing a normal physiological component of synovial fluid and cartilaginous tissue, exogenous administration of hyaluronic acid has the potential to have few side effects or local or systemic reactions. Viscosupplementation represents an alternative treatment for patients with osteoarthritis in which oral medications and/or surgery are not options or are ineffective. PMID- 10695859 TI - Orthopedic medical malpractice: an attorney's perspective. AB - Orthopedic surgeons are trained to manage problems involving the musculoskeletal system. It would be helpful to identify certain procedures, anatomic areas, or issues related to the physician-patient relationship that could potentially lead to a malpractice lawsuit. Once the problems are identified, steps toward continuing education and physician awareness could be initiated. In this study, we performed a randomized nationwide survey of medical malpractice attorneys to evoke their opinion on these issues. We found that the lumbar spine was the most common anatomic area involved in orthopedic medical malpractice cases, and a physician appearing rushed and uninterested is most likely to be the subject of a lawsuit where a poor physician-patient relationship was a contributing factor. Educational and professional programs are needed to increase the awareness and knowledge of orthopedic malpractice risks, and also to identify potentially preventable problems leading to malpractice litigation. PMID- 10695860 TI - Anterior fibular strut grafting for the treatment of pseudoarthrosis in spondylolisthesis. AB - Ten patients with a failed posterior spinal fusion for symptomatic spondylolisthesis were treated with retroperitoneal anterior lumbosacral interbody fusion. A fibular strut allograft was placed, followed by posterolateral fusion and instrumentation. The mean follow-up was 40 months (range 24-60 months). All patients complained of back pain and leg pain before surgery. All patients achieved solid fusion at L5-S1. One patient developed pseudoarthrosis at L4-5 and improved symptomatically with no postoperative complications. PMID- 10695861 TI - Intraoperative rotational measuring device: a new device to achieve accuracy in corrective osteotomy of malrotated femur. AB - A new derotation device for corrective femoral osteotomy is reported. This device helps eliminate errors of undercorrection and overcorrection in derotational osteotomies of femurs with torsional deformities. A prototype of the intraoperative rotational measuring device was tested on 16 cadaver femurs. This experiment conclusively demonstrated that this simple device is easy to use and a mathematical precision (P < 0.05) is accomplished in rotational corrections by surgical means. PMID- 10695862 TI - Mild gait abnormality and leg discomfort in a child secondary to extradural ganglioneuroma. AB - Ganglioneuromas are benign and slow-growing tumors that most commonly originate from the sympathetic trunk. Ganglioneuromas often decrease in size and rarely require reoperation. Changes in gait or the onset of limb pain without a discernible local cause are indications for investigation of patients for possible intraspinal pathology. We report the case of a 5-year-old boy who presented with seemingly static symptoms, while the slow-growing tumor had enveloped nerve roots and caused bone destruction of the vertebrae. PMID- 10695863 TI - Complications in the management of complex Monteggia-equivalent fractures in adults. AB - We report five adult patients with complex Monteggia-equivalent fractures who were surgically treated, all of whom had significant complications. All ulnar fractures and three radial fractures developed nonunion. Three patients required more than two procedures to achieve bony union, and one required a total elbow arthroplasty. PMID- 10695864 TI - Two-stage extensor tendon reconstruction after composite tissue loss from the dorsum of the hand. AB - Restoration of digital extension after chronic extensor loss has not been detailed extensively in the literature. The present report details an unusual case of composite tissue loss from the dorsum of the hand after a chronic burn wound. After debridement for chronic carpal osteomyelitis and free-tissue transfer were performed, staged wrist fusion and two-stage extensor tendon reconstruction resulted in a stable, pain-free wrist and functional digital extension. The present case illustrates that two-stage extensor tendon reconstruction, when necessary, is indeed feasible. PMID- 10695865 TI - Second-look arthroscopy with removal of bioabsorbable tacks. AB - Eleven years after tearing her anterior cruciate ligament (ACL) (not reconstructed), a 36-year-old dancer reinjured her knee and required arthroscopic ACL reconstruction. At arthroscopy, the medial meniscus had a bucket-handle tear that was repaired by using three bioabsorbable tacks. The ACL was then repaired in the usual manner. Because of persistent posterior knee pain throughout her rehabilitation, we performed "second-look" arthroscopy 14 weeks after reconstruction. The meniscus had healed and was stable; however, tack motion was evident and the tacks were easily removed. Inspection of the tacks showed that the barbs had been resorbed. The patient recovered uneventfully, and pain-free flexion 28 days after surgery was 0 degrees-136 degrees. We believe this to be the first reported case demonstrating the early stages of tack degradation in meniscal repair. PMID- 10695866 TI - Displaced bucket-handle meniscal tear. PMID- 10695867 TI - The aging ovary. AB - During reproductive life, ovarian steroid biosynthesis is gonadotropin dependent and occurs in theca and granulosa cells. In the menopausal ovary, there is atresia of ovarian follicles, with sparing of the androgen-producing theca interstitial cell component. The aging ovary, therefore, produces significantly reduced amounts of estrogen, with continued, though decreased, androgen production. After menopause, ovarian estradiol biosynthesis is minimal, with circulating estrogen being derived principally from peripheral aromatization of ovarian and adrenal androgens. Androgen biosynthesis from the adrenal gland, in addition to that from the ovary, decreases with age. Although ovarian androgen production declines with age, there is not an abrupt decrease as is seen with ovarian estrogen levels at the time of menopause. The biological activity of these steroids, either before or after menopause, depends on the amount of steroid available in the unbound fraction. To this end, sex hormone-binding globulin (SHBG) levels are an important determinant of hormone action. Not only does the concentration of SHBG influence the biological effect of testosterone and estradiol, but these steroids also regulate SHBG concentrations. PMID- 10695868 TI - Hormone receptors and sexuality in the human female. AB - There is only inferential evidence based on observational studies and deductive reasoning that allows us to suppose there are hormone receptors in the human brain that are associated with libido. Data indicate that sexually active pubescent women have higher testosterone levels than matched controls. However, any number of nonhormonal events in puberty could be related to these changes. Another circuitous way to look at hormone libido receptors in the brain is by evaluating patients on birth control pills, in whom sexual activity paradoxically decreases. Sex hormone-binding globulin (SHBG) levels are increased, and, therefore, free testosterone levels seem to be affected by the use of birth control pills. In addition, the progestins in the pill are 19-nortestosterone derivatives, which may have some androgenic effect in their own right. Women on estrogen replacement therapy with the addition of testosterone seem to report increased libido, and there is subjective information that there is an increase in psychological sense of well-being. It has been well documented that females with adrenogenital syndrome are masculinized in utero, indicating imprinting on behavior. Because of the nature of the end point (e.g., increased libido), it is difficult to draw concrete conclusions on the presence of libido receptors in the human brain. Because the outcomes and end points are so soft, it is difficult to come to a consensus, and in fact this topic polarizes opinions of the pure scientist (objective) against the clinician (subjective). PMID- 10695869 TI - Sexuality among older women. AB - The population of the world is living longer. Currently, the average life expectancy is 78.9 and 72 years of age for women and men, respectively. By the year 2030, it is estimated that elderly people will make up approximately 17% of the total United States population. Our responsibility as physicians is, therefore, very clear. The care of this population must provide an acceptable quality of life and allow elderly people to enjoy living. This is where the question of sexuality plays an important part. Hormonal transition encompasses decreased levels of estrogen and testosterone, the latter being associated with decreased sexual libido, sensitivity, and response. Additional genitourinary effects associated with menopause include atrophic changes in the vagina, vulva, urethra, and neck of the bladder. Vaginal atrophy and diminished vaginal lubrication interfere mechanically with sexual comfort and pleasure. In addition to hormonal changes with aging, disease and associated medications may also negatively affect sexuality. Determining the impact of medications, both alone and in combination with others, on quality of life must be considered when providing comprehensive care for elderly patients. PMID- 10695870 TI - Approaches to taking a sexual history. AB - Advances in modern medicine have extended the lives of many people, in both quantity and quality. One of the emerging quality of life factors for many women as they approach and pass the menopause is sexuality. Sexuality is a very important part of physical and emotional health, underscoring the importance of incorporating the sexual history as part of the overall patient history. Although health professionals may experience a degree of anxiety and discomfort in discussing sexual issues, it is nonetheless essential to learn to be comfortable in asking questions about sexuality and in responding to issues that arise from such questioning. Most patients also show some discomfort when discussing their sexual problems. A sensitive, nonjudgmental approach on the part of the physician is essential and may create an atmosphere of security for both patient and physician. It is sometimes helpful to begin the sexual history with basic, open ended questions, thereby allowing for expansion according to the responses received. Dispelling the myth that all older people should have a declining interest in sex may help patients feel less reticent about talking to physicians about sexual matters. PMID- 10695871 TI - Effects of hormone replacement therapy on sexual psychophysiology and behavior in postmenopause. AB - Investigators continue to define the exact relationship between sexual function and changes in hormonal status during menopause. The availability of different preparations that could replace estrogens and androgens has led to many studies of the use of hormone replacement therapy (HRT) for sexual dysfunction. Dyspareunia due to vaginal dryness appears to be most responsive to estrogen replacement therapy (ERT) via restoration of vaginal cells, pH, and blood flow. Progestins, to a certain extent, can oppose these changes and lead to a recurrence of dryness and dyspareunia. ERT has also been reported to enhance sexual desire in a significant percent of women. Although treatment with ERT has been shown to be efficacious for many women, there are others whose sexual difficulties remain unresponsive. There also appears to be a significant subgroup of women whose sexual difficulties respond initially to ERT but who subsequently revert to their initial problems, especially when the problem has been loss of libido. For these women, the addition of androgen has proved helpful. PMID- 10695872 TI - The psychological impact of aging on sexuality and relationships. AB - Aging has a powerful impact on the quality of relationships and sexual functioning. The psychological impact of aging after midlife is a particularly timely topic given improved medical and psychological understanding of sexuality in both women and men, as well as more effective treatment for age-related sexual dysfunctions. It is time to dispel the stereotype of the midlife relationship as the continuation of a traditional heterosexual marriage with grown or almost grown children in order to more effectively address emotional and sexual issues arising in relationships. Regardless of the length or nature of the relationship, however, its quality is enhanced by emotional intimacy, autonomy without too much distance, an ability to manage stress and distractions by external factors, and achieving a satisfying sexual equilibrium. Perception of the quality of the primary relationship and sexuality is influenced by the other factors in a person's life. Thus, the relationship must be examined and issues must be addressed taking these external factors into consideration. Among the most powerful external factors is one's occupation or avocation, as it tends to strongly influence one's sense of identity, self-esteem, and self-worth in all areas of life. To understand and treat effects of aging on sexuality, it is important to address the three components of sexual desire: drive, beliefs/values, and motivation, as well as the sexual equilibrium within the primary relationship. It is also essential to understand how the physiological changes in male and female sexual functioning affect desire and equilibrium. Other health-related changes that occur with aging must be recognized and addressed, including the fact that the oldest of old women will outlive their corresponding male cohort. Treatment implications for these issues are discussed. PMID- 10695873 TI - Contrast enhancement in magnetic resonance imaging using intravenous paramagnetic contrast media: a review. AB - Interest in magnetic resonance (MR) imaging as a major diagnostic tool in veterinary clinical medicine is increasing. Most MR studies are performed with the use of contrast enhancement via intravenous injection of paramagnetic gadolinium-containing contrast media. A vast number of publications are available regarding the use of contrast media in humans. The purpose of this paper is to assist practicing veterinary radiologists in understanding mechanisms of MR contrast enhancement. This paper reviews certain aspects of MR contrast enhancement, including physical, chemical and biologic characteristics of most common MR contrast media targeted primarily at the central nervous system (CNS). Authors also describe processes that explain changes in signal intensity on the MR images. PMID- 10695874 TI - An evaluation of a new radiographic technique utilizing a concave table. AB - In conventional radiography systems, it is apparent that only the area immediately around the central x-ray beam can be evaluated accurately. Consequently in some instances, spinal radiography for example, several exposures are needed at various points along the body to create an accurate image for diagnosis. However, if the film and body part are in a concave shape such that the radius of the curve is equal to the film focal distance, the x-ray beam will penetrate the body and strike the film at two-dimensionally right angles in all areas. Using the spine as an example we found the curved technique had three major advantages over the traditional flat technique: lack of distortion, more uniform beam intensity due to a constant focal film distance, and improved resolution at the periphery of the radiograph because of lack of a cross over effect. It was concluded that an accurate evaluation of larger body parts can be made with minimal distortion utilizing the principles of a curved table technique. PMID- 10695875 TI - Subclinical CT abnormalities in the lumbosacral spine of older large-breed dogs. AB - Computed tomography (CT) of the L5-S3 vertebral levels was performed in six, large-breed dogs presented for problems unrelated to the lumbosacral spine. All dogs were asymptomatic for lumbosacral stenosis on neurologic examination. Breeds included German Shepherd, Golden Retriever, Boxermix and Belgian Malinois. Ages ranged from 5-12 years. Five out of six dogs exhibited CT abnormalities. Among the 18 disc levels examined, the most common findings were idiopathic stenosis, loss of vertebral canal epidural fat, and nerve tissue displacement. Less common abnormalities were vertebral canal or foraminal bone proliferation, loss of intervertebral foramen fat, vertebral canal disc bulging, degenerative articular process joint disease, transitional vertebra, dural ossification, foraminal disc bulging, Schmorl's nodes, calcified extruded disc fragment, and sacroiliac joint osteophytes. Vertebral subluxation was absent in all dogs. Findings indicate that some lumbosacral CT abnormalities may be clinically insignificant, especially in older dogs. PMID- 10695876 TI - Quantitative magnetic resonance imaging of the normal feline cranial abdomen. AB - Magnetic resonance images of the cranial abdomen were acquired from 15 clinically normal cats. All cats had T1-weighted images, 8 cats had T2-images made and 7 cats had T1-weighted post Gd-DTPA images acquired. Signal intensity measurements for T1, T2, and T1 post contrast sequences were calculated for liver, spleen, gallbladder, renal cortex, renal medulla, pancreas, epaxial muscles, and peritoneal fat. On T1-weighted images the epaxial muscle had the lowest signal intensity, followed by renal medulla, spleen, renal cortex, pancreas, liver and fat, respectively. On T2-weighted images, epaxial muscle had the lowest signal intensity followed by liver, spleen, fat, and gallbladder lumen. Calculations of specific organ percent enhancement following contrast medium administration were made and compared with that reported in humans. A brief review of the potential clinical uses of MR in cats is presented. PMID- 10695877 TI - Dynamic magnetic resonance imaging of the normal canine pituitary gland. AB - The pituitary glands of six normal dogs were evaluated using dynamic magnetic resonance imaging. T1 weighted images were obtained every 13 seconds for three minutes of three contiguous slices through the pituitary gland following a bolus intravenous injection of gadolinium-DTPA. Contrast enhancement was seen initially in the region of the pituitary stalk at 52-65 seconds followed by uniform enhancement at 104-143 seconds post injection. This pattern of enhancement was seen in all subjects and is similar to that reported in humans. PMID- 10695878 TI - Radiographic diagnosis: mediastinal parathyroid cyst in a cat. PMID- 10695879 TI - Radiographic diagnosis: thoracic spinal fracture resulting in kyphosis in a horse. PMID- 10695880 TI - Magnetic resonance imaging of otitis media in a dog. AB - Otitis media/interna was diagnosed in a 20-month-old German shepherd with the assistance of magnetic resonance (MR) imaging. The MR images were acquired primarily to exclude a brain lesion responsible for vestibular signs. No brain lesion was detected, but obvious signs of chronic changes in the left bulla and external ear canal were confirmed. Thickening of the epithelium and soft tissue surrounding the external ear canal and a laminated appearance of high and low T2 intensities in the tympanic bulla's mucosa were present. The hypointense lines were suspected to be fibrous tissue, indicating chronic changes. This report suggests that MR imaging may serve as a useful imaging tool for otitis media and that it supplies information not obtained with radiography or computed tomography. PMID- 10695881 TI - Quantitative evaluation of imagent as an abdominal ultrasound contrast medium in dogs. AB - Diagnostic ultrasound currently provides a noninvasive, economical means to evaluate internal organ structure. Methods to increase the amount of diagnostic information obtained from the sonographic examination are being explored. A new area of research has recently focused on the ability of microbubble contrast agents to help provide additional information. A prospective study was undertaken to quantitatively evaluate the amount of gray scale enhancement of ultrasound images of six canine livers, spleens, and left kidneys that occurs after intravenous injection of Imagent. Computerized analysis was used to assess the percent change in brightness of these images. A transient decrease in the percent change in brightness was seen to occur after injection of higher doses of Imagent. This effect was more pronounced in the deeper portions of the organs of interest. Imagent provides a measurable change in the brightness of the organs of interest and would be potentially useful in a clinical setting. PMID- 10695882 TI - The effects of a unilateral ultrasound-guided renal biopsy on renal function in healthy sedated cats. AB - Complications of renal biopsies are well documented except for the change in renal function after a biopsy. Eighteen healthy, adult cats were divided into two groups (n = 9 cats/group). For the measurement of global and split renal function, Group 1 used the renal uptake of 99mTc-DTPA and Group 2 used the renal uptake of 99mTc-MAG3. Scintigraphic data were collected on days (-4), (-3), 0, 1, 2, and 4 post renal biopsy. Using ultrasound guidance, biopsies were taken from the right renal cortex on dO, before acquiring scintigraphic images. P - values less than 0.10 were considered significant due to the limited number of observations. The only statistically significant change (p = 0.08) in global renal function detected was by day following a unilateral renal biopsy. Cats imaged using 99mTc-MAG3 had discernible liver activity. A unilateral, ultrasound guided renal biopsy has minimal effect on renal function in normal, healthy sedated cats. PMID- 10695883 TI - Measurements of hindlimb blood flow recorded using Doppler ultrasound during administration of vasoactive agents in halothane-anesthetized horses. AB - The purpose of the study was to determine the ability of Doppler ultrasound to detect changes in femoral blood flow during pharmacologic manipulation of arterial blood pressure. Doppler ultrasonography was performed in the femoral vessels of six halothane-anesthetized horses before and during administration of phenylephrine HCI and sodium nitroprusside. The time-averaged mean velocity and volumetric flow were calculated. The contour of the velocity waveform was assessed, and the early diastolic deceleration slope (EDDS) and pulsatility index (PI) were calculated. Administration of phenylephrine HCI resulted in increased mean aortic blood pressure (MABP) by 40% (29.3-53.0%). This caused significant decrease in cardiac output (26.8 to 13.5 l/min), femoral arterial velocity (left artery 7.20 to 4.00 cm/s; right artery 5.01 to 3.39 cm/s) and volumetric flow (left artery 556 to 221 ml/min; right artery 397 to 193 ml/min) in the femoral vessels and significant increase in systemic vascular resistance (163 to 433 dyn s/cm5), EDDS (1a: 285 to 468: ra: 250 to 481) and PI (1a: 9.38 to 20.4; ra 17.1 to 29.1). Administration of sodium nitroprusside resulted in a decreased MABP of 27.2% (22.5-33%). This increased cardiac output (20.8 to 32.4 L/min), however, no significant changes were observed in femoral blood flow. Despite obvious changes in the waveform contour, no significant change occurred in EDDS or PI. These results suggest that Doppler ultrasound may be useful for measuring femoral blood flow in anesthetized horses. However, waveform analysis appears to be limited when multiple changes occur in central and peripheral haemodynamics. PMID- 10695884 TI - Determination of canine prostatic volume using transabdominal ultrasonography. AB - The prostate gland from cadavers of 12 intact adult male dogs euthanized less than 3 hour were used to compare prostatic volume measured by ultrasonography to volume measured by water displacement, and to determine specific gravity of the canine prostate. Prostate glands were scanned by transabdominal ultrasonography with a 4-7 MHz curved linear array transducer. The greatest craniocaudal (L), transverse (W), and dorsoventral (D) diameters of the prostate were recorded. Prostatic volume was calculated using formulas for an ellipsoid and for a box. Prostate glands were removed, and the prostate weight was measured and prostatic volume was measured by water displacement. The mean +/- SD specific gravity of the prostate was 1 +/- 0.05 (range = 0.90 to 1.09) g/cm. There were positive correlations (R2 = 0.94) between prostatic volume calculated from ultrasound measurement and measured volume. Measured prostatic volume (VM) can be predicted using the formula: VM = [1/2.6 (L x W x D)] + 1.8 (cm3). PMID- 10695885 TI - Evaluation of plasma time-activity curves of technetium-99m-mebrofenin for measurement of hepatic function in dogs. AB - In this study, plasma time-activity curves of 99mTc-mebrofenin were used to quantify hepatic function in dogs before and after induction of hepatic damage using a hepatotoxic agent. Nine dogs were determined to be healthy on the basis of physical examination, laboratory data and hepatic imaging. Plasma samples were collected 1, 3, 5, 7, 9, 15, 20, 30, 40, 50, and 60 minutes following a peripheral venous injection of 111-222 MBq (3-6 mCi) of 99mTc-mebrofenin. The area under the plasma time-activity curve (AUC) was calculated using two different methods and compared to direct measurement of the hepatic extraction efficiency. First pass hepatic extraction efficiency of 99mTc-mebrofenin was calculated from differential equation analysis of a two-compartment model following mesenteric venous injection of the radiopharmaceutical. In 7 of the original 9 dogs and 2 additional healthy dogs, plasma clearance and hepatic extraction efficiency determination were repeated following induction of hepatic injury by thiacetarsamide (3 mg/kg IV twice daily for 1 day). In one additional dog, hepatic injury was induced using carbon tetrachloride (0.3 ml/kg IP). Plasma time-activity curves of 99mTc-mebrofenin had kinetics of a two compartment model. Area under the curve was highly correlated with hepatic extraction efficiency. The AUC integrated from 1-60 minutes (AUC60) had the best correlation with hepatic extraction efficiency (r2 = 0.978, p < 0.001). A formula for calculation of hepatic extraction efficiency was derived using linear regression analysis: hepatic extraction efficiency = 105.583 - 3.099 x 10(5) x AUC60. Plasma clearance of a peripheral venous injection of 99mTc-mebrofenin is a simple, non-invasive, convenient method to quantify hepatic function which can be performed without a gamma camera. PMID- 10695886 TI - Use of 99mTc-mercaptoacetyltriglycine to evaluate renal function in horses. AB - Ten healthy horses were injected intravenously with 99mTc-MAG3 and the disappearance of radioactivity from the blood was measured. The total body clearance (Cl(B)) and elimination half-life (t1/2(beta)) were 7.9 +/- 1.5 ml/kg/minute and 32.8 +/- 4.1 minutes, respectively. The disappearance of 99mTc MAG3 from the blood of 2 horses with compromised renal function was also measured. The data suggest that 99mTc-MAG3 is a useful and clinically applicable radiopharmaceutical for measurement of effective renal blood flow in the horse. PMID- 10695887 TI - Radiographs presented as part of the 1999 ACVR oral certification examination: large animal imaging elective. PMID- 10695888 TI - Intrathecal baclofen for management of spastic cerebral palsy: multicenter trial. AB - Intrathecal baclofen infusion has demonstrated effectiveness in decreasing spasticity of spinal origin. Oral antispasticity medication is minimally effective or not well tolerated in cerebral palsy. This study assessed the effectiveness of intrathecal baclofen in reducing spasticity in cerebral palsy. Candidates were screened by randomized, double-blind, intrathecal injections of baclofen and placebo. Responders were defined as those who experienced an average reduction of 1.0 in the lower extremities on the Ashworth Scale for spasticity. Responders received intrathecal baclofen via the SynchroMed System and were followed for up to 43 months. Fifty-one patients completed screening and 44 entered open-label trials. Lower-extremity spasticity decreased from an average baseline score of 3.64 to 1.90 at 39 months. A decrease in upper extremity spasticity was evidenced over the same study period. Forty-two patients reported adverse events. Most common reports were hypotonia, seizures (no new onset), somnolence, and nausea or vomiting. Fifty-nine percent of the patients experienced procedural or system-related events. Spasticity in patients with cerebral palsy can be treated effectively by continuous intrathecal baclofen. Adverse events, although common, were manageable. PMID- 10695889 TI - Psychogenic seizures after head injury in children. AB - A total of 148 patients with a history of probable seizure disorder were studied prospectively with long-term video electroencephalography over a 1-year period. Sixteen (11%) were identified with psychogenic seizures. Eleven (69%) of 16 were boys, with a mean age of 10.5 years. Seven (44%) of the 16 had an antecedent history of head injury prior to the development of these episodes; of these patients 85% were boys. Our results suggest that contrary to findings in the adult population, psychogenic seizures can be commonly seen in boys. The prevalence of antecedent head injury suggests that it is a notable risk factor in children as well as adults in the occurrence of psychogenic seizures. PMID- 10695890 TI - Tuberous sclerosis complex and epilepsy: prognostic significance of electroencephalography and magnetic resonance imaging. AB - Tuberous sclerosis complex is a disease that affects many organs, including the central nervous system. Nervous system involvement in the form of hamartomas often results in seizures. In this study we wanted to determine the outcome of epilepsy in tuberous sclerosis complex and determine whether interictal electroencephalograms (EEGs) and hamartoma burden as seen with magnetic resonance imaging (MRI) are predictive of degree of seizure control. The study population consisted of 30 patients. For each patient two sets of EEG and MRI data, separated by at least 12 months, and information on seizure frequency at time of data collection were obtained. Sensitivity, specificity, and positive and negative predictive values of various EEG and MRI findings were determined. Seizure control improved in 20 and worsened in 10 patients. In relation to seizure control, the specificity of an abnormal sleep EEG and the positive predictive value of normal sleep EEG were 100%. MRI and EEG background were neither sensitive nor specific for predicting seizure control. A majority of children with tuberous sclerosis complex can achieve good seizure control. The sleep EEG is helpful in predicting eventual seizure control. PMID- 10695891 TI - De Lange's syndrome: facial features of pediatric neurologic syndromes. PMID- 10695892 TI - Midline developmental anomalies with lipomas in the corpus callosum region. AB - Three children with complete or partial callosal aplasia and intracranial lipoma in the corpus callosum region were investigated. Two lipomas were tubulonodular; one replaced the entire corpus callosum structure. Accompanying anomalies affected the cingulate gyrus, septum pellucidum, and choroid plexus. In one case, diagnosis was made in utero in the 25th gestational week by ultrasonography; in the second case it was made on the first day of life, also by screening ultrasonography. Two children had mild spastic distal diparesis; one complained of chronic headache. Electroencephalography showed no abnormalities; epilepsy anamnesis was negative. Somatosensory and visual evoked potentials showed prolonged conduction in two cases. Surgery was not indicated. Because of the risk of developing epileptic seizures, regular electroencephalographic follow-up investigations are essential. PMID- 10695893 TI - Quantitative morphology of the corpus callosum in children with neurofibromatosis and attention-deficit hyperactivity disorder. AB - Neurofibromatosis-1 is a common autosomal-dominant genetic disorder associated with numerous physical anomalies and an increased incidence of attention-deficit hyperactivity disorder (ADHD). Studies of children with idiopathic ADHD have suggested a link between corpus callosum size and symptom severity. This study examines the contribution of corpus callosum morphology to symptoms of ADHD in children with neurofibromatosis. Eighteen control subjects and 36 children with neurofibromatosis underwent magnetic resonance imaging of the brain. Twelve subjects with neurofibromatosis had evidence of ADHD and 24 did not. Subjects with neurofibromatosis had significantly larger total corpus callosum area and significantly larger regional measurements in three of seven areas. However, there were no differences between the neurofibromatosis alone and neurofibromatosis plus ADHD groups. Increased severity of attention problems was associated with smaller total callosal areas. These results suggest that some features of ADHD in children with neurofibromatosis could be linked to quantifiable differences in brain morphology, but the nature of the genetic mutation in neurofibromatosis suggests that neurochemical effects also could be important. PMID- 10695894 TI - Prospective analysis of strength in spinal muscular atrophy. DCN/Spinal Muscular Atrophy Group. AB - Spinal muscular atrophy is a genetic disorder of the motor neurons that causes profound hypotonia, severe weakness, and often fatal restrictive lung disease. Patients with spinal muscular atrophy present a spectrum of disease from the most severe infantile-onset type, called Werdnig-Hoffmann disease (type 1), associated with a mortality rate of up to 90%, to a late-onset mild form (type 3), wherein patients remain independently ambulatory throughout adult life. Although many clinicians agree that patients with spinal muscular atrophy lose motor abilities with age, it is unknown whether progressive weakness occurs in all patients with spinal muscular atrophy. We present here results of the first prospective study of muscle strength in patients with spinal muscular atrophy. There was no loss in muscle strength as determined by a quantitative muscle test during the observation period. However, motor function diminished dramatically in some patients with spinal muscular atrophy. Explanations for this loss of function could not be determined from our data. Decrease in motor function could be caused by factors other than loss of strength. Therefore, it is not clear from our results whether spinal muscular atrophy is a neurodegenerative disease. We conclude that treatment trials in spinal muscular atrophy should be designed with consideration of the natural history of strength and motor function in this disorder. PMID- 10695895 TI - Developmental brain anomalies in children with attention-deficit hyperactivity disorder. AB - The pathoetiology of attention-deficit hyperactivity disorder (ADHD) has been considered to be neurodevelopmental, yet the timing and processes involved are not clearly identified. Neurodevelopmental brain anomalies have been associated with a variety of psychiatric conditions. However, they have never been evaluated in a population of patients with ADHD. This study was designed to determine the frequency of specific developmental brain anomalies in a group of children with ADHD (n = 85; mean age, 10.9 years) and healthy control children (n = 95; mean age, 11.7 years) by visually inspecting brain magnetic resonance imaging scans. Compared to controls, the ADHD group showed an increase in frequency of two developmental anomalies: (1) gray-matter heterotopia, a neuronal migration anomaly, in 2 of 85 patients versus 0 of 95 controls; and (2) posterior fossa abnormality (excess cerebrospinal fluid in the posterior fossa) in 8 of 85 patients versus 2 of 95 controls. There were no differences in frequency of enlarged cavum septi pellucidi between the two groups. These findings support and extend the idea that ADHD is of developmental origin, and further suggest that the timing of aberrant brain development could be in early gestation. PMID- 10695896 TI - How to construct a neural tube. PMID- 10695897 TI - Hot water epilepsy: a benign and unrecognized form. AB - Hot water epilepsy is a reflex epilepsy. Seizures are provoked by hot water, and result from the association of both cutaneous and heat stimuli. Described mainly in India and Japan, the condition seems to be rare in Europe, where it occurs in young children. We report five infants aged from 6 months to 2 years. They had brief seizures during bathing with activity arrest, hypotonia, and vasoactive modification; clonic movements were observed. A simple treatment-decreasing the bath temperature-can be sufficient. Sometimes an antiepileptic drug is required. Seizure course and psychomotor development are favorable. Hot water epilepsy is a benign form of epilepsy. Its incidence could be underestimated because of confusion with febrile convulsions, vagal fits, or aquagenic urticaria. PMID- 10695898 TI - Alternating hemiplegia of childhood in half-sisters. AB - We present the family of two girls affected with alternating hemiplegia of childhood who were born to the same mother and different fathers. Previous reports suggested mitochondrial dysfunction as an etiologic mechanism for this disorder. Muscle biopsy, including a measurement of the respiratory chain enzymes, performed in one of the sisters showed no mitochondrial abnormalities. The mode of inheritance is not certain, but an autosomal-dominant gene is most likely. PMID- 10695899 TI - Vagal and hypoglossal Bell's palsy. AB - A 7-year-old boy was referred because of a sudden change to nasal speech, dysarthria for words with explosive consonants in speech, and nasal regurgitation of fluids. The symptoms arose over 1 week following a capricious episode of acute asthmatic bronchitis. Physical and neurologic examinations were normal except for a left deviation of the uvula, accompanied by a "curtain" movement of the posterior pharyngeal wall against the opposite side, and a left deviation of the protruded tongue. No vascular, traumatic, infectious, neoplastic, or neurologic causes could be identified. No therapy was administered. Full recovery occurred 4 months later. The diagnosis was idiopathic vagal and right hypoglossal nerve palsy (Bell's palsy). PMID- 10695900 TI - Venlafaxine in children, adolescents, and young adults with autism spectrum disorders: an open retrospective clinical report. AB - Autism is characterized by social deficits, communication and language impairments, narrow restricted interests, repetitive behaviors, inattention, and hyperactivity. While selective serotonin reuptake inhibitors have demonstrated efficacy in treating core symptoms of autism, norepinephrine reuptake inhibitors have demonstrated efficacy in symptoms of attention-deficit hyperactivity disorder (ADHD). An open, retrospective clinical study with venlafaxine evaluated its effect on core symptoms of autism as well as associated features of ADHD. Ten consecutive subjects meeting Diagnostic and Statistical Manual of Mental Disorders, 4th edition (DSM-IV), criteria for an autism spectrum disorder were treated with venlafaxine, initiated at 12.5 mg per day and adjusted on a flexible basis. Six of 10 completers were judged to be sustained treatment responders, by scoring 1 (very much improved) or 2 (much improved) on the Clinical Global Impressions improvement scale. Venlafaxine was effective in low dosages (mean, 24.37 mg/day; range, 6.25 to 50 mg/day) and was well tolerated. Improvement was noted in repetitive behaviors and restricted interests, social deficits, communication and language function, inattention, and hyperactivity. Controlled treatment trials with venlafaxine are warranted in autism spectrum disorders. PMID- 10695901 TI - Fatality from hepatitis A in a child taking valproate. AB - We report an 8-year-old boy with complex partial seizures due to congenital stroke, treated with valproate for more than 3 years (the last 2 years were on monotherapy) with no complications during that period except for transient thrombocytopenia. His sister had uncomplicated hepatitis A. One month later, the patient became jaundiced, went into fulminant hepatic failure, and quickly became encephalopathic despite discontinuation of valproate, aggressive supportive therapy, and treatment with carmitine. He then died. He had positive hepatitis A IgM; other causes for acute hepatitis were ruled out. Liver pathology revealed distended hepatocytes with cholestasis and microvesicular changes. We could find in the literature two other articles on four cases who developed liver failure with hepatitis A while on valproate. All those cases, however, recovered. In our patient a usually benign disease became deadly, probably because of the concomitant use of a hepatotoxic medication. Immunizing, with hepatitis A vaccine, all children on valproate therapy who are living in, or traveling to, endemic areas should be considered and is probably advisable. PMID- 10695902 TI - HyperCKemia as the only sign of McArdle's disease in a child. AB - An asymptomatic 13-year-old boy, who never complained of exercise intolerance or myalgia, was found to have markedly elevated serum creatine kinase (CK) levels during a routine check-up. General physical and neurologic examinations were normal. Surprisingly, histochemical and biochemical analysis of muscle showed myophosphorylase deficiency and genetic analysis showed that the patient was homozygous for the most common mutation encountered in McArdle's disease (R49X). This case illustrates the fuzzy correlation between molecular defect and clinical phenotype in patients with McArdle's disease, and suggests that a thorough study of the muscle biopsy is important in patients with idiopathic hyperCKemia for correct diagnosis and careful follow-up. PMID- 10695903 TI - Oxcarbazepine versus carbamazepine treatment and induction of serum lipid abnormalities. AB - Long-term treatment with carbamazepine is associated with many metabolic changes, including elevations in serum cholesterol, high-density lipoprotein, low-density lipoprotein, and triglyceride levels. Oxcarbazepine has been reported to be a preferable antiepileptic drug, when compared to carbamazepine with respect to its effects on lipid metabolism, especially cholesterol, high-density lipoproteins, and low-density lipoproteins. The case of a 16-year-old girl who developed high serum cholesterol and low-density lipoprotein levels on both of these medications successively, while triglyceride and high-density lipoprotein levels were unaffected, is reported. PMID- 10695904 TI - Operating theatre--the patient is listening. PMID- 10695905 TI - Auditory evoked responses and learning and awareness during general anesthesia. AB - BACKGROUND: There is a major distinction between conscious and unconscious learning. Monitoring the mid-latency auditory evoked responses (AER) has been proposed as a measure to ascertain the adequacy of the hypnotic state during surgery. In the present study, we investigated the presence of explicit and implicit memories after anesthesia and examined the relationships of such memories to the AER. METHODS: We studied 180 patients scheduled for elective surgical procedures. After a thiopental induction, one of four anesthetics were studied: Opioid bolus: 7.5 microg x kg(-1) fentanyl, 70% N2O, with 2.5 microg x kg(-1) supplements as needed (n=100); Opioid infusion: Alfentanil 50 microg x kg( 1) bolus, 1-1.5 microg x kg(-1) x min(-1) infusion, 70% N2O (n=40); Isoflurane 0.3%: Fentanyl 1 microg x kg(-1), 70% N2O, isoflurane 0.3% expired (n=16); Isoflurane 0.7%: Fentanyl 1 microg x kg(-1), 70% N2O, isoflurane 0.7% expired (n=23). AER were recorded before anesthesia, 5 min after surgical incision and then every 30 min until the end of surgery. A tape of either the story of the "Three Little Pigs" or the "Wizard of Oz" was played continuously between the recordings. Explicit memory was assessed postoperatively by tests of recall and recognition, and implicit memory was assessed by the frequency of story-related free associations to target words from the stories, which were solicited twice during a structured interview. RESULTS: Six patients showed explicit recall of intraoperative events: All received the opioid bolus regimen. About 7% of patients reported dreaming during anesthesia. The incidence of picking the correct story that had been presented during anesthesia averaged 49%, i.e., very close to chance level. Overall, priming occurred only at the second association tests for the opioid bolus regimen, for which the frequency of an association to the presented story among those not giving an association to the control story was 26%, which was double the frequency (13%) of an association to the control story among those not giving an association to the presented story. This was significant by McNemar's test, P=0.02. There were significant associations between awareness, priming and AER, e.g., recall was associated with higher Nb amplitudes during anesthesia and priming was associated with shorter wave latencies. CONCLUSIONS: The incidence of awareness in patients anesthetized with nitrous oxide and bolus supplementation was 6%. Thus, this anesthetic technique did not reduce the risk of awareness compared with the use of nitrous oxide alone. Implicit memory occurred during nitrous oxide and bolus supplementation. Recording AER during anesthesia may help to predict awareness and implicit memory, particularly the former. The short contents of most of the dreams which were recalled could hamper future studies in this area. PMID- 10695906 TI - Severe neuroexcitatory symptoms after anaesthesia--with focus on propofol anaesthesia. AB - Delayed neuroexcitatory symptoms after an uneventful anaesthesia are uncommon, although described in many reports. We want to report on two cases. The first patient developed muscle hypertonicity, jerky movements and unconsciousness after an uneventful anaesthesia with propofol, and later the same thing happened after anaesthesia with thiopentone. The second patient developed similar symptoms after an uneventful anaesthesia with propofol, but she never recovered completely after this and is now severely disabled. A search of the literature and the Swedish adverse drug reactions register revealed many similar cases. In both our patients the causal relationship between propofol and the neuroexcitatory symptoms remains uncertain, but we want to alert readers about this possible adverse reaction. PMID- 10695907 TI - Haemodynamic effects of intravenous clonidine on propofol or thiopental induction. AB - The study evaluated the effects of premedication with intravenous clonidine on thiopental or propofol requirements for induction and haemodynamic changes associated with both induction and endotracheal intubation. Clonidine administered intravenously before induction of anaesthesia reduced propofol or thiopental requirements. The association of clonidine and propofol caused, after injection of the induction drug, a decrease in mean arterial pressure which was significantly greater than with thiopental. Moreover, a major haemodynamic stability was registered before and after laryngoscopy in the clonidine thiopental group. These findings might contraindicate the clonidine-propofol combination in patients with cardiovascular disease. PMID- 10695908 TI - Cost comparison between three different general anaesthetic techniques for elective arthroscopy of the knee. AB - INTRODUCTION: We compared three anaesthetic techniques for elective knee arthroscopy with special reference to cost-effectiveness. METHOD: Seventy-five ASA I-II patients having elective arthroscopy of the knee joint were randomised to receive an anaesthetic technique based on propofol, fentanyl for induction followed by sevoflurane in oxygen:nitrous oxide (1:2 l/min) for maintenance of one of two intravenous techniques: propofol alfentanil or propofol-remifentanil infusions in combination with oxygen in air. RESULTS: All patients had an uncomplicated course. No differences were seen with regard to emergence, postoperative pain or emesis or time to discharge. The anaesthetic technique based on sevoflurane was associated with the lowest cost US$ 14.7 as compared to US$ 18 for the propfol/alfentanil and US$ 19.9 for the propofol/remifentanil technique, including both cost for wastage as well as premedication and other fixed drug costs. Looking only at the anaesthetic drugs consumed, the cost per minute was US$ 0.56 for sevoflurane/nitrous oxide as compared to US$ 0.68 and 0.63 per minute for the propofol/alfentanil and proprofol/remifentanil, respectively. When the cost for wastage was taken into account, the difference in mean anaesthetic drug cost was more pronounced: the sevoflurane anaesthetic technique US$ 0.58, the propofol/alfentanil US$ 0.74 and the propofol/remifentanil US$ 0.84 per minute respectively. CONCLUSION: From a cost minimisation point of view, anaesthesia based on sevoflurane in oxygen:nitrous oxide is the technique of choice. PMID- 10695909 TI - Influence of acute normovolaemic haemodilution on the dose-response and time course of action of atracurium. AB - BACKGROUND: Acute normovolaemic haemodilution is a common method to save and avoid homologous blood transfusion during surgery. The aim of this study was to evaluate the influence of acute isovolaemic haemodilution on the dose-response and time-course of action of atracurium. METHODS: We studied 25 patients undergoing acute isovolaemic haemodilution during surgery and 25 patients not receiving haemodilution as a control group. All patients were ASA grade I and aged 18-54 yr. The haemodilution patients underwent major elective plastic surgery with an anticipated surgical blood loss of more than 600 ml, and the control patients underwent elective superficial plastic surgery with an anticipated surgical blood loss of less than 200 ml. Anaesthesia was induced with thiopental 4-6 mg/kg and fentanyl 2-4 microg/kg i.v., and maintained with 60% nitrous oxide in oxygen. After stabilization of anaesthesia, acute isovolaemic haemodilution in the haemodilution group was achieved by drainage of venous blood and an i.v. infusion of lactated Ringer's solution, and 6% dextran, during which haematocrit and haemoglobin decreased from 45.1% to 25.8% and from 147.2 g/l to 91.2 g/l, respectively. When anaesthesia was stable in the control group and haemodilution was completed in the haemodilution group, neuromuscular function was assessed by measuring with accelerography the response of the adductor pollicis muscle to supramaximal train-of-four (TOF) stimuli every 12 s to the ulnar nerve at the wrist via surface electrodes. The dose-response relationships of atracurium in the two groups were determined by the cumulative dose-response technique. RESULTS: The results showed that during haemodilution, the dose response curve of atracurium was shifted to the left in a parallel fashion and the potency of atracurium was increased. In patients undergoing haemodilution, ED50, ED90 and ED95 of atracurium were decreased by 25-33%, and duration of action was increased by 21-48% following administration of the same dose (mg/kg), as compared with patients not undergoing haemodilution. CONCLUSION: We concluded that the patients undergoing acute isovolaemic haemodilution were about 30% more sensitive to neuromuscular blockade of atracurium and had a longer duration after administration of the same dose (microg/kg) than the control patients. Care must be taken with this problem when atracurium is used as a muscle relaxant during acute haemodilution. PMID- 10695910 TI - Nerve stimulation at 0.15 Hz when compared to 0.1 Hz speeds the onset of action of cisatracurium and rocuronium. AB - BACKGROUND: Onset time of specific non-depolarizing muscle relaxants (NDMR), as reported in the literature, varies widely. To test the suggestion of a consensus panel (Copenhagen Consensus Conference 1996) that onset time in muscle relaxant studies depends on stimulation frequency, even in the low frequency range, we examined the onset time of cisatracurium and rocuronium by parallel ulnar nerve stimulation in both upper extremities at 0.1 Hz and 0.15 Hz. METHODS: Thirty patients (ASA class I and II) were included. Onset of action following the administration of 2xED90 of cisatracurium (0.092 mg x kg(-1)) or rocuronium (0.74 mg x kg(-1)) was measured by quantifying the evoked response of the adductor pollicis muscles using mechanomyography. Single twitch stimulations were applied simultaneously to the ulnar nerves of both forearms using a 0.1 Hz and 0.15 Hz stimulation frequency, respectively. RESULTS: Both relaxants showed a significantly (P<0.01) shorter onset time with 0.15 Hz compared to 0.1 Hz stimulation (cisatracurium: 186+/-50 s vs. 233+/-59 s, rocuronium: 73+/-14 s vs. 99+/-23 s; x+/-SD). CONCLUSIONS: Onset time of NDMR depends on the stimulation frequency, even in the low frequency range. When comparing onset studies, the reader must also compare the stimulation rates used by the investigators. We recommend the use of 0.1 Hz single twitch nerve stimulation frequency as the standard for studies of onset profile of NDMR. PMID- 10695911 TI - Composition of the editorial/advisory boards of major English-language anesthesia/critical care journals. AB - BACKGROUND: Publications represent a central part of the research process. An analysis of who is responsible for acceptance of publications in major English language anesthesia/critical care medicine journals was carried out. METHODS: All English-language journals listed in the SCI Journal Citation Reports 1997 under the subheadings Anesthesiology (n=18) and Emergency Medicine & Critical Care (n=16) were analysed with regard to the editorship and the membership of advisory boards listed in the 1998 issues of the journals. The two groups were analysed separately with regard to their country of origin. RESULTS: In the Anesthesiology section, 140 persons were listed as editors and 423 persons were identified as members of the advisory boards. Editors came from 14 different countries, with editors from the USA representing the majority (n=83; 59% of all editors, followed by the UK: n=24; 15% of all editors). Editors from other countries represent only a minority (n=33; 24% of all editors). The advisory boards came from 30 countries and were also dominated by the USA (n=220; 52% of all persons from the advisory boards). In the Emergency Medicine & Critical Care section 159 persons were listed as editors, of whom 119 originated from the USA (75% of all editors). Of the 835 persons listed in the advisory boards, 72% came from the USA, with 37 other countries sharing the remainder (second, UK: 8%; third, Canada: 2.5%). CONCLUSION: Most editors/editorial board members of important Anesthesiology, Emergency and Critical Care journals came from the USA. Other countries play a significantly less influential role even in journals which are characterised as 'International Journals'. PMID- 10695912 TI - Cardiac output measurements during cross-clamping of the descending thoracic aorta in pigs. A comparison between transit-time ultrasound, thermodilution and pulsed Doppler ultrasound. AB - BACKGROUND: Cross-clamping of the descending thoracic aorta (XC) induces an increase in cardiac output (CO). The intention of this study was to evaluate the high CO during XC by the use of clinically available methods (thermodilution and pulsed Doppler ultrasound) compared to transit-time ultrasound flowmetry of the ascending aorta as the gold standard. METHOD: Ten pigs were anaesthetised with ketamine and fentanyl. The descending thoracic aorta was cross-clamped for 30 min, and cardiac output was measured with pulmonary artery thermodilution technique, pulsed Doppler ultrasound on the aortic annulus and transit-time ultrasound flowmetry of the ascending aorta. RESULTS: At 15 min following XC, CO increased from 1.7 l/min to 4.6 l/min measured with transit-time ultrasound (P<0.05). With thermodilution technique, CO increased from 2.6ll/min to 5.7 l/min (P<0.05), and from 2.4 l/min to 6.0 l/min measured with Doppler ultrasound (P<0.05). There was an increase in mean arterial pressure of 81% and heart rate increased 76% (P<0.05). CONCLUSION: XC of the descending thoracic aorta induces an increase in CO of 171%. Thermodilution and pulsed Doppler ultrasound are reliable methods for detecting high cardiac output during thoracic aortic surgery. PMID- 10695913 TI - Coronary and systemic hemodynamic effects of clevidipine, an ultra-short-acting calcium antagonist, for treatment of hypertension after coronary artery surgery. AB - BACKGROUND: The aim was to evaluate the use of clevidipine, a new vascular selective, ultra-short-acting calcium antagonist for blood pressure control after coronary artery bypass grafting (CABG). METHODS: The effects of clevidipine on central hemodynamics, myocardial blood flow and metabolism were studied at two different phases after CABG. In phase 1 (n=13), the hypertensive phase, the effects of clevidipine were compared to those of sodium nitroprusside (SNP) when used to control postoperative hypertension. In phase 2 (n=9), the normotensive phase, a clevidipine dose-response relationship was established. RESULTS: At a target mean arterial pressure (MAP) of 75 mmHg, systemic vascular resistance (SVR) and heart rate (HR) were lower, preload, stroke volume (SV) and pulmonary vascular resistance (PVR) were higher, while there were no differences in myocardial lactate metabolism or oxygen extraction with clevidipine compared to SNP. In the normotensive phase, clevidipine induced a dose-dependent decrease in MAP (-19%), SVR (-27%) and PVR (-15%), accompanied by an increase in SV (10%), but no reflex increase in HR or changes in cardiac preload. Clevidipine caused a direct coronary vasodilation, as indicated by a decrease in myocardial oxygen extraction from 54% to 45%. Myocardial lactate metabolism was unaffected by clevidipine. The blood clearance of clevidipine was 0.05 l x min(-1) x kg(-1), the volume of distribution at steady state was 0.08 l x kg(-1) and the initial and terminal half-lives were <1 min and 4 min, respectively. CONCLUSIONS: Clevidipine rapidly reduced MAP and induced a systemic, pulmonary and coronary vasodilation with no effect on venous capacitance vessels or HR. Clevidipine caused no adverse effects on myocardial lactate metabolism. Clevidipine thus appears suitable to control blood pressure after CABG. PMID- 10695914 TI - Effect of dantrolene in an in vivo and in vitro model of myocardial reperfusion injury. AB - BACKGROUND: In skeletal muscle, dantrolene reduces free cytosolic calcium by inhibiting calcium release from the sarcoplasmic reticulum. A similar effect in ischemic-reperfused heart cells would protect myocardial tissue against reperfusion injury. We tested the hypothesis that dantrolene infusion during reperfusion protects the heart against reperfusion injury. METHODS: Isovolumetric beating rat hearts were subjected to 30 min of ischemia followed by 60 min of reperfusion. Left ventricular (LV) developed pressure (LVDP) and creatine kinase release (CKR) were determined as indices of myocardial performance and cellular injury, respectively. In the treatment groups, dantrolene (25 (DAN25) or 100 (DAN100) micromol l(-1)) was infused during the first 15 min of reperfusion; control hearts received the respective concentration of the vehicle (mannitol (CON25, CON100), each group n=7). To investigate the effects of dantrolene on reperfusion injury in vivo, 18 chloralose-anesthetized rabbits were subjected to 30 min occlusion and 180 min reperfusion of a major coronary artery. LV pressure (LVP), cardiac output (CO), and infarct size were determined. During the last 5 min of ischemia, nine rabbits received 10 mg kg(-1) dantrolene intravenously (DAN). Another nine rabbits received the vehicle (dimethylsulfoxide) and served as controls (CON). RESULTS: In isolated rat hearts, there was no recovery of LVDP in any group. Total CKR during 1 h of reperfusion was 845+/-76 (CON100) and 550+/ 81 U g(-1) dry mass (DAN100, P<0.05). In rabbits in vivo, hemodynamic baseline values were similar between groups (CON vs. DAN: LVP, 99+/-6 (mean+/-SEM) vs. 91+/-6mm Hg, P=0.29; CO, 252+/-26 vs. 275+/-23 ml min(-1), P= 0.53). During coronary artery occlusion, LVP and CO were reduced in both groups (CON: LVP, 89+/ 3%; CO, 90+/-5% of baseline values) and LVP did not recover to baseline values during reperfusion (51+/-5% (CON) vs. 67+/-7% (DAN) of baseline, P=0.10). Infarct size was 41+/-4% of the area at risk in controls and 37+/-6% in dantrolene treated hearts (P=0.59). CONCLUSIONS: Dantrolene reduced CKR, indicating an attenuation of lethal cellular reperfusion injury in isolated rat hearts. However, in the rabbit in vivo, there was no effect on the extent of reperfusion injury after regional myocardial ischemia. PMID- 10695915 TI - In vivo protein synthesis of circulating human T lymphocytes does not respond to a cortisol challenge within 24 h. AB - BACKGROUND: Although immunocompetence is often measured by assessing responsiveness of lymphocytes to mitogenic stimulation in vitro, this approach may not reflect the in vivo situation. The aim of this investigation was to determine in vivo the protein synthesis rate (FSR) in isolated T lymphocytes and to study the effect of a short-term cortisol infusion on FSR. METHODS: Healthy volunteers (n=24) were randomised into 4 groups. A continuous cortisol infusion (6 microg kg(-1) min(-1)) during 6 h was given to groups 1 and 2, whereas groups 3 and 4 served as control groups and received saline infusion. Protein synthesis was studied before and after 6 h of the cortisol/saline infusion (groups 1 and 3) or 24 h after the start of the infusion (groups 2 and 4). FSR was determined in vivo by the flooding method. The isotopic enrichment of phenylalanine in plasma and lymphocyte protein was determined with gas chromatography-mass spectrometry. RESULTS: The FSR in T lymphocytes was 13.6+/-0.9%/24 h as a mean value (+/-SD) of the first determination in 4 groups. There was no significant difference in FSR from the baseline value immediately after the cortisol infusion (group 1: 13.3+/ 1.4%/24 h vs 13.5+/-2.8%/24 h) or 24 h after the start of the infusion (group 2: 13.6+/-0.7%/24 h vs 12.3+/-2.4%/24 h). CONCLUSION: The metabolic activity of circulating T lymphocytes, as reflected by a quantitative measurement of in vivo protein synthesis of human T lymphocytes, was not affected by the increased level of cortisol. PMID- 10695916 TI - Needle design does not affect the success rate of spinal anaesthesia or the incidence of postpuncture complications in children. AB - BACKGROUND: In adults, pencil-point spinal needles are believed to be less traumatic and therefore to be superior compared to cutting-point needles with respect to success rate and postpuncture complications. The aim of this randomised, parallel groups and prospective study was to record the success rate and to evaluate the incidence of complications following spinal anaesthesia with the two types of needles in children. METHODS: We studied 215 children aged 1 to 18 years. A 25-gauge needle was used in children up to 7 years (n=96) and a 27 gauge needle in older children (n=119). During lumbar puncture with either a cutting-point (n=109) or a pencil-point (n=106) spinal needle, we recorded puncture characteristics and the success of cerebrospinal fluid (CSF) aspiration. Hyperbaric bupivacaine 5 mg ml(-1) at a dose of 0.3-0.4 mg kg(-1) was used for the spinal anaesthesia. The incidence of postdural puncture complications was recorded from diaries completed by the children and parents one week after the lumbar puncture. RESULTS: The success rate of the spinal anaesthesia was 97% without difference between the needles. The success rate was higher when the aspiration of CSF was easy compared to if it was difficult (98% vs. 88%, P=0.02). Two hundred and seven diaries were returned (97%). Twenty-four children developed a headache, 8 of which were classified as a postdural puncture headache (PDPH), 6 with the cutting-point needle and 2 with the pencil-point needle (n.s.). Nine children developed signs of transient radicular irritation with no difference between the needles. CONCLUSION: Both types of spinal needles can be used in children, and a free aspiration of CSF results in a high success rate of the spinal block. Postpuncture complications are as common in children as in adults. PMID- 10695917 TI - Comparison of combined spinal epidural anesthesia and epidural anesthesia for cesarean section. AB - BACKGROUND: Epidural anesthesia (EA) is popular for cesarean section, but has some drawbacks such as incomplete block, inadequate muscle relaxation and delayed onset. Combined spinal epidural anesthesia (CSEA) has gained increasing interest as it combines the reliability of a spinal block and the flexibility of an epidural block. We investigated the efficacy of CSEA that combines the main spinal and the supporting epidural anesthesia, comparing with pH-adjusted EA, for cesarean section. METHODS: Sixty-four pregnant women at full term were divided into two groups. Patients in the CSEA group (n=32) were given 1.5-1.6 ml of 0.5% hyperbaric bupivacaine intrathecally, followed by 10 ml of 0.25% plain bupivacaine through the epidural catheter 10 min later. Patients in the EA group (n=32) received 20-25 ml of 2% lidocaine which was already mixed with 0.1 ml of 0.1% epinephrine, 100 g of fentanyl and 1.5 ml of 8.4% sodium bicarbonate. The quality and side effects of surgical anesthesia, neonatal state, and postoperative course were compared between the two groups. RESULTS: In the EA group, 22% (7 cases) complained of intraoperative pain but none in the CSEA group (P=0.011). Muscle relaxation and motor block were much better in the CSEA group (P<0.001 and P=0.011 each). Significantly more women in the EA group had shivering (P=0.001). They also had more nausea and vomiting but the differences were not significant. Not only the time to T4 block (9.7 vs. 18.3 min, mean, P<0.001) but also the stay in the postanesthesia care unit, recovery of sensory and motor block and start of postoperative pain were all significantly shorter in the CSEA group. No one in either group had postdural puncture headache (PDPH). CONCLUSION: We can conclude that, when combining the main spinal and the supporting epidural anesthesia, CSEA has greater efficacy and fewer side effects than the pH-adjusted EA in cesarean sections. PMID- 10695918 TI - Chronic pain after thoracic surgery. PMID- 10695919 TI - General characteristics of Pinus spp. seed fatty acid compositions, and importance of delta5-olefinic acids in the taxonomy and phylogeny of the genus. AB - The delta5-unsaturated polymethylene-interrupted fatty acid (delta5-UPIFA) contents and profiles of gymnosperm seeds are useful chemometric data for the taxonomy and phylogeny of that division, and these acids may also have some biomedical or nutritional applications. We recapitulate here all data available on pine (Pinus; the largest genus in the family Pinaceae) seed fatty acid (SFA) compositions, including 28 unpublished compositions. This overview encompasses 76 species, subspecies, and varieties, which is approximately one-half of all extant pines officially recognized at these taxon levels. Qualitatively, the SFA from all pine species analyzed so far are identical. The genus Pinus is coherently united--but this qualitative feature can be extended to the whole family Pinaceae -by the presence of delta5-UPIFA with C18 [taxoleic (5,9-18:2) and pinolenic (5,9,12-18:3) acids] and C20 chains [5,11-20:2, and sciadonic (5,11,14-20:3) acids]. Not a single pine species was found so far with any of these acids missing. Linoleic acid is almost always, except in a few cases, the prominent SFA, in the range 40-60% of total fatty acids. The second habitual SFA is oleic acid, from 12 to 30%. Exceptions, however, occur, particularly in the Cembroides subsection, where oleic acid reaches ca. 45%, a value higher than that of linoleic acid. Alpha-linolenic acid, on the other hand, is a minor constituent of pine SFA, almost always less than 1%, but that would reach 2.7% in one species (P. merkusii). The sum of saturated acids [16:0 (major) and 18:0 (minor) acids principally] is most often less than 10% of total SFA, and anteiso-17:0 acid is present in all species in amounts up to 0.3%. Regarding C18 delta5-UPIFA, taxoleic acid reaches a maximum of 4.5% of total SFA, whereas pinolenic acid varies from 0.1 to 25.3%. The very minor coniferonic (5,9,12,15-18:4) acid is less than 0.2% in all species. The C20 elongation product of pinolenic acid, bishomo-pinolenic (7,11,14-20:3) acid, is a frequent though minor SFA constituent (maximum, 0.7%). When considering C20 delta5-UPIFA, a difference is noted between the subgenera Strobus and Pinus. In the former subgenus, 5,11-20:2 and sciadonic acids are < or =0.3 and < or =1.9%, respectively, whereas in the latter subgenus, they are most often > or =0.3 and > or =2.0%, respectively. The highest values for 5,11-20:2 and sciadonic acids are 0.5% (many species) and 7.0% (P. pinaster). The 5,11,14,17-20:4 (juniperonic) acid is present occasionally in trace amounts. The highest level of total delta5-UPIFA is 30-31% (P. sylvestris), and the lowest level is 0.6% (P. monophylla). Uniting as well as discriminating features that may complement the knowledge about the taxonomy and phylogeny of pines are emphasized. PMID- 10695920 TI - The biosynthesis of oxylipins of linoleic and arachidonic acids by the sewage fungus Leptomitus lacteus, including the identification of 8R-Hydroxy-9Z,12Z octadecadienoic acid. AB - When the sewage fungus Leptomitus lacteus was grown in liquid culture aerobically and then transferred to medium containing long-chain fatty acids, it produced a number of oxygenated fatty acids. From linoleic acid (18:2n-6), the major metabolite produced was R-8-hydroxy-9Z,12Z-octadecadienoic acid (8R-HODE), with additional quantities of 8,11-di-HODE, 11,16-di-HODE, and 11,17-di-HODE. Other fatty acid derivatives identified included 7-HODE, 10-HODE, and 13-hydroxy octadecamonoenoic acid. Arachidonic acid (20:4n-6) was metabolized primarily to 18- and 19-hydroxy-eicosatetraenoic acids (18- and 19-HETE) also as R enantiomers, along with smaller quantities of 17-HETE, 9-HETE, 14,15-dihydroxy eicosatrienoic acid and 11,12,19-trihydroxy-eicosatrienoic acid. The oxygenated products of long-chain fatty acids, in particular the biosynthesis of 8R-HODE, a compound classified as a precocious sporulation inducer, were similar to those produced by an unrelated fungal species in the Ascomycota, the take-all fungus Gaeumannomyces graminis. As in G. graminis, the biotransformation of linoleate to 8R-HODE was not significantly inhibited by exposure of the organism to CO. This indicated that the enzyme responsible for 8R-HODE biosynthesis in Leptomitus could be similar to that of G. graminis; yet we did not detect 7,8-di-HODE as a product of 18:2n-6 metabolism as in G. graminis. CO did inhibit the biosynthesis of 14,15-di-HETE, 18-HETE, and 19-HETE in L. lacteus, which suggested the involvement of a cytochrome P450-type monooxygenase. The biosynthesis of 8R-HODE from 18:2n-6 was found to occur in certain cell lysates, specifically in low speed (15,000 x g) supernatant, following cell disruption. PMID- 10695921 TI - Fatty acid transport across lipid bilayer planar membranes. AB - The transport of palmitic acid (PA) across planar lipid bilayer membranes was measured using a high specific activity [14C]palmitate as tracer for PA. An all glass trans chamber was employed in order to minimize adsorbance of PA onto the surface. Electrically neutral (diphytanoyl phosphatidylcholine) and charged (Azolectin) planar bilayers were maintained at open electric circuit. We found a permeability to PA of (8.8 +/- 1.9) x 10(-6) cm s(-1) (n = 15) in neutral and of (10.3 +/- 2.2) x 10(-6) cm s(-1) (n = 5) in charged bilayers. These values fall within the order of magnitude of those calculated from desorption constants of PA in different vesicular systems. Differences between data obtained from planar and vesicular systems are discussed in terms of the role of solvent, radius of curvature, and pH changes. PMID- 10695922 TI - Coordinate packaging of newly synthesized phosphatidylcholine and phosphatidylglycerol in lamellar bodies in alveolar type II cells. AB - Methylamine, a weak base, inhibits packaging of newly synthesized phosphatidylcholine (PC) in lamellar bodies in 20-22 h cultured alveolar type II cells, suggesting a role for acidic pH of lamellar bodies. In this study, we tested if (i) the packaging of PC is similarly regulated in freshly isolated type II cells and (ii) methylamine also inhibits the packaging of other surfactant phospholipids, particularly, phosphatidylglycerol (PG). The latter would suggest coordinated packaging so as to maintain the phospholipid composition of lung surfactant. During the short-term metabolic labeling experiments in freshly isolated type II cells, methylamine treatment decreased the incorporation of radioactive precursors into PC, disaturated PC (DSPC), and PG of lamellar bodies but not of the microsomes, when compared with controls. The calculated packaging (the percentage of microsomal lipid packaged in lamellar bodies) of each phospholipid was similarly decreased (approximately 50%) in methylamine-treated cells, suggesting coordinated packaging of surfactant phospholipids in lamellar bodies. Equilibrium-labeling studies with freshly isolated type II cells (as is routinely done for studies on surfactant secretion) +/- methylamine showed that in methylamine-treated cells, the secretion of PC and PG was decreased (possibly due to decreased packaging), but the phospholipid composition of released surfactant (measured by radioactivity distribution) was unchanged; and the PC content (measured by mass or radioactivity) of lamellar bodies was lower, but the PC composition (as percentage of total phospholipids) was unchanged when compared with control cells. We speculate that the newly synthesized surfactant phospholipids, PC, DSPC, and PG, are coordinately transported into lamellar bodies by a mechanism requiring the acidic pH, presumably, of lamellar bodies. PMID- 10695923 TI - Protective effect of oleuropein, an olive oil biophenol, on low density lipoprotein oxidizability in rabbits. AB - On the basis of the results obtained with pilot studies conducted in vitro on human low density lipoprotein (LDL) and on cell cultures (Caco-2), which had indicated the ability of certain molecules present in olive oil to inhibit prooxidative processes, an in vivo study was made of laboratory rabbits fed special diets. Three different diets were prepared: a standard diet for rabbits (diet A), a standard diet for rabbits modified by the addition of 10% (w/w) extra virgin olive oil (diet B), a modified standard diet for rabbits (diet C) differing from diet B only in the addition of 7 mg kg(-1) of oleuropein. A series of biochemical parameters was therefore identified, both in the rabbit plasma and the related isolated LDL, before and after Cu-induced oxidation. The following, in particular, were selected: (i) biophenols, vitamins E and C, uric acid, and total, free, and ester cholesterol in the plasma; (ii) proteins, triglycerides, phospholipids, and total, free, and ester cholesterol in the native LDL (for the latter, the dimensions were also measured); (iii) lipid hydroperoxides, aldehydes, conjugated dienes, and relative electrophoretic mobility (REM) in the oxidized LDL (ox-LDL). In an attempt to summarize the results obtained, it can be said that this investigation has not only verified the antioxidant efficacy of extra virgin olive oil biophenols and, in particular, of oleuropein, but has also revealed a series of thus far unknown effects of the latter on the plasmatic lipid situation. In fact, the addition of oleuropein in diet C increased the ability of LDL to resist oxidation (less conjugated diene formation) and, at the same time, reduced the plasmatic levels of total, free, and ester cholesterol ( 15, -12, and -17%, respectively), giving rise to a redistribution of the lipidic components of LDL (greater phospholipid and cholesterol amounts) with an indirect effect on their dimensions (bigger by about 12%). PMID- 10695924 TI - Lipoproteins modify the macrophage uptake of triacylglycerol emulsion and of zymosan particles by similar mechanisms. AB - The uptake of lipids and formation of foam cells are key events in atherosclerosis and in eruptive xanthomata formation in primary hyperchylomicronemia. Here we have compared the influence of low density lipoprotein (LDL), oxidized LDL (oxLDL), high density lipoprotein (HDL), and delipidated HDL (apoHDL) on the uptake by macrophages of zymosan (an insoluble fraction of yeast cell walls) and of triglyceride-rich emulsion (EM) particles that resemble chylomicrons, but, like zymosan, are equally devoid of protein components. Zymosan internalization is known to occur through unspecific phagocytosis, whereas natural chylomicrons are taken up by several specific lipoprotein receptors. We found that phagocytosis is not promoted as much by oxLDL as by normal LDL. HDL-coated zymosan was found to be inert and apoHDL slightly enhanced phagocytosis. LDL and apoHDL promoted the uptake of EM while oxLDL and HDL significantly inhibited the uptake. Therefore, the data support that HDL, and not apoHDL, particles inhibit EM uptake. We concluded that by using lipoprotein-coated zymosan particles, we could demonstrate different biological effects of LDL, oxLDL, HDL, and apoHDL on macrophage phagocytosis and that this method could be useful to delineate components of the various lipoproteins important for the propagation or inhibition of the formation of foam cells. PMID- 10695925 TI - Dietary docosahexaenoic acid suppresses inflammation and immunoresponses in contact hypersensitivity reaction in mice. AB - This study was designed to examine the immunomodulatory effects of dietary docosahexaenoic acid (DHA) in the absence of eicosapentaenoic acid (EPA). We investigated the effects of feeding dietary DHA ethyl ester (DHA-Et) (97% pure) at levels of 4.8 wt% of the total diet and of feeding EPA ethyl ester (EPA-Et) (99% pure) at 4.8 wt% on the inflammatory response in the challenge phase of the contact hypersensitivity reaction (CHR) in the ears of mice sensitized with 2,4 dinitro-1-fluorobenzene (DNFB). The effect of DHA-Et on T lymphocytes at the CHR site was examined using anti-CD4 antibodies. Furthermore, we examined the cytokines formed at the CHR site on the mRNA level. It was found that 24 h after the challenge, DHA-Et but not EPA-Et reduced the ear swelling. Infiltration of inflammatory cells, in particular, CD4-positive T lymphocytes, into the ears in the challenge phase of CHR was observed. DHA-Et reduced the infiltration of CD4 positive T lymphocytes into the ears. DHA-Et also decreased the expression of interferon-gamma, interleukin (IL)-6, IL-1beta, and IL-2 mRNA in ears. These observations suggest that DHA, but not EPA, may exert an antiinflammatory and immunosuppressive effect. The immunosuppressive effectiveness of fish oil may be attributed mainly to DHA. PMID- 10695926 TI - The metabolism and distribution of docosapentaenoic acid (n-6) in rats and rat hepatocytes. AB - In this study, a new marine oil that contains 45% docosahexaenoic acid (DHA, 22:6n-3) and 13% docosapentaenoic acid (DPA, 22:5n-6) was administered to rats. The metabolism and distribution of DPA in rats was investigated. In experiment 1, the effects of DHA and n-6 fatty acids (linoleic acid, LA; arachidonic acid, AA; and DPA) on AA contents were investigated in vivo. LA group: LA 25%, DHA 30%; LA DPA group: LA 15%, DPA 10%, DHA 35%; LA-AA-DPA group: LA 10%, AA 5%, DPA 10%, DHA 35% were administered to rats for 4 wk. In the liver, the AA content in the LA DPA and LA-AA-DPA groups was significantly higher than in the LA group. The decreased AA contents in the LA group might be caused by DHA administration. Although DHA also was administered in the LA-DPA and LA-AA-DPA groups, the AA contents in these two groups did not decrease. These results suggested that DPA retroconverted to AA, blunting the decrease in AA content caused by DHA administration. To conduct a detailed investigation on DPA metabolism and its relation with AA and DHA, rat hepatocytes were cultured with purified DPA and DHA for 24 h. We discovered the retroconversion of DPA to AA occurred only when AA content was decreased by a high DHA administration; it did not occur when AA content was maintained at a normal level. PMID- 10695927 TI - Relationships between the fatty acid composition of muscle and erythrocyte membrane phospholipid in young children and the effect of type of infant feeding. AB - Muscle membrane fatty acid (FA) composition is linked to insulin action. The aims of this study were to compare the FA composition of muscle and erythrocyte membrane phospholipid in young children; to investigate the effect of diet on these lipid compositions; and to investigate differential incorporation of FA into muscle, erythrocyte and adipose tissue membrane phospholipid, and adipose tissue triglyceride. Skeletal muscle biopsies and fasting blood samples were taken from 61 normally nourished children (45 males and 16 females), less than 2 yr old (means +/- SE, 0.80 +/- 0.06 yr), undergoing elective surgery. Adipose tissue samples were taken from 15 children. There were significant positive correlations between muscle and erythrocyte docosahexaenoic acid (DHA) (r = 0.44, P < 0.0001), total n-3 polyunsaturated fatty acids (PUFA) (r = 0.39, P = 0.002), and the n-6/n-3 PUFA ratio (r = 0.39, P = 0.002). Adipose tissue triglyceride had lower levels of long-chain PUFA, especially DHA, than muscle and erythrocytes (0.46 +/- 0.18% vs. 2.44 +/- 0.26% and 3.17 +/- 0.27%). Breast-fed infants had higher levels of DHA than an age-matched group of formula-fed infants in both muscle (3.91 +/- 0.21% vs. 1.94 +/- 0.18%) and erythrocytes (3.81 +/- 0.40% vs. 2.65 +/- 0.23%). The results of this study show that (i) erythrocyte FA composition is a reasonable index of muscle DHA, total n-3 PUFA, and the n-6/n-3 PUFA ratio; (ii) breast feeding has a potent effect on the FA composition of all these tissues; and (iii) there is a wide range in long-chain PUFA levels in muscle, erythrocytes, and adipose tissue. PMID- 10695928 TI - Effects of different medium-chain fatty acids on intestinal absorption of structured triacylglycerols. AB - To study the effect of the chain length of medium-chain fatty acids on the intestinal absorption of long-chain fatty acids, we examined the lymphatic transport of fat following administration of five purified structured triacylglycerols (STAG) containing different medium-chain fatty acids in the sn 1,3 positions and long-chain fatty acids in the sn-2 position in a rat model. Significant amounts of medium-chain fatty acids were found in lymph samples after intragastric administration of 1,3-dioctanoyl-2-linoleyl-sn-glycerol (8:0/18:2/8:0), 1,3-didecanoyl-2-linoleyl-sn-glycerol, and 1,3-didodecanoyl-2 linoleyl-sn-glycerol. The accumulated lymphatic transport of medium-chain fatty acids increased with increasing carbon chain length. The recoveries of caprylic acid (8:0), capric acid (10:0), and lauric acid (12:0) were 7.3 +/- 0.9, 26.3 +/- 2.4, and 81.7 +/- 6.9%, respectively. No significant differences were observed for the maximal intestinal absorption of linoleic acid (18:2n-6) when the chain length of medium-chain fatty acids at the primary positions was varied, and the absorption of 18:2 and oleic acid (18:1) from 8:0/18:2/8:0 and 1,3-dioctanoyl-2 oleyl-sn-glycerol was similar. We conclude that the chain length of the medium chain fatty acids in the primary positions of STAG does not affect the maximal intestinal absorption of long-chain fatty acids in the sn-2 position in the applied rat model, whereas the distribution of fatty acids between the lymphatics and the portal vein reflects the chain length of the fatty acids. PMID- 10695929 TI - Effects of conjugated linoleic acid isomers on lipid-metabolizing enzymes in male rats. AB - Male weanling Wistar rats (n = 15), weighing 200-220 g, were allocated for 6 wk to diets containing 1% (by weight) of conjugated linoleic acid (CLA), either as the 9c,11 t-isomer, the 10t,12c-isomer, or as a mixture containing 45% of each of these isomers. The five rats of the control group received 1% of oleic acid instead. Selected enzyme activities were determined in different tissues after cellular subfractionation. None of the CLA-diet induced a hepatic peroxisome proliferation response, as evidenced by a lack of change in the activity of some characteristic enzymes [i.e., acyl-CoA oxidase, CYP4A1, but also carnitine palmitoyltransferase-I (CPT-I)] or enzyme affected by peroxisome-proliferators (glutathione S-transferase). In addition to the liver, the activity of the rate limiting beta-oxidation enzyme in mitochondria, CPT-I, did not change either in skeletal muscle or in heart. Conversely, its activity increased more than 30% in the control value in epididymal adipose tissue of the animals fed the CLA-diets containing the 10t,12c-isomer. Conversely, the activity of phosphatidate phosphohydrolase, a rate-limiting enzyme in glycerolipid neosynthesis, remained unchanged in adipose tissue. Kinetic studies conducted on hepatic CPT-I and peroxisomal acyl-CoA oxidase with CoA derivatives predicted a different channeling of CLA isomers through the mitochondrial or the peroxisomal oxidation pathways. In conclusion, the 10t,12c-CLA isomer seems to be more efficiently utilized by the cells than its 9c,11t homolog, though the Wistar rat species appeared to be poorly responsive to CLA diets for the effects measured. PMID- 10695930 TI - Apolipoprotein E polymorphism: automated determination of apolipoprotein E2, E3, and E4 isoforms. AB - Apolipoprotein E (apo E) plays an essential role in lipoprotein metabolism, where it is involved in the clearance of chylomicrons and very low density lipoproteins. Apart from some rare variants, apo E exists in three common isoforms (E2, E3, and E4). The different isoforms have not only been associated with different plasma lipid levels but have also been correlated with certain pathological conditions, such as lipid disorders (dysbetalipoproteinemia, hypercholesterolemia), cardiovascular diseases, and Alzheimer's disease. Here we describe a rapid, automated test for the determination of the most frequent polymorphisms (E2, E3, and E4). This polymerase chain reaction-based test allows the reliable discrimination of all six genotypes. The assay has been developed especially for the nonspecialized routine clinical laboratory by employing an analyzer and chemistry often present in this type of laboratory. Because of its low costs and easy handling, the assay can be performed on a daily basis. PMID- 10695931 TI - Breast-fed infants achieve a higher rate of brain and whole body docosahexaenoate accumulation than formula-fed infants not consuming dietary docosahexaenoate. AB - Docosahexaenoate (DHA) has been increasingly recognized as an important fatty acid for neural and visual development during the first 6 mon of life. One important point of controversy that remains is the degree to which adequate levels of DHA can be acquired from endogenous synthesis in infants vs. what should be provided as dietary DHA. We have approached this problem by a retrospective analysis of published body composition data to estimate the actual accumulation of DHA in the human infant brain, liver, adipose tissue, remaining lean tissue, and whole body. Estimating whether infants can synthesize sufficient DHA required comparison to and extrapolation from animal data. Over the first 6 mon of life, DHA accumulates at about 10 mg/d in the whole body of breast-fed infants, with 48% of that amount appearing in the brain. To achieve that rate of accumulation, breast-fed infants need to consume a minimum of 20 mg DHA/d. Virtually all breast milk provides a DHA intake of at least 60 mg/d. Despite a store of about 1,050 mg of DHA in body fat at term birth and an intake of about 390 mg/d alpha-linolenate (alpha-LnA), the brain of formula-fed infants not consuming DHA accumulates half the DHA of the brain of breast-fed infants while the rest of the body actually loses DHA over the first 6 mon of life. No experimental data indicate that formula-fed infants not consuming DHA are able to convert the necessary 5.2% of alpha-LnA intake to DHA to match the DHA accumulation of breast-fed infants. We conclude that dietary DHA should likely be provided during at least the first 6 mon of life. PMID- 10695932 TI - Glycosaminoglycans in gingival crevicular fluid of patients with periodontal class II furcation involvement before and after guided tissue regeneration. A pilot study. AB - BACKGROUND: The levels of glycosaminoglycans in gingival crevicular fluid (GCF) are good indicators of underlying tissue turnover. We hypothesize that connective tissue elements in GCF may be used as indicators of tissue maturation underneath barrier membranes. Therefore, we investigated the levels of sulfated glycosaminoglycans in GCF at sites before and after guided tissue regeneration (GTR). METHODS: Six patients were selected on the basis of having at least one Class II buccal furcation involvement on a molar tooth. Each molar furcation was treated with the standard GTR surgical protocol using a non-resorbable expanded polytetrafluoroethylene membrane. Gingival crevicular fluid samples were taken at baseline (immediately prior to insertion of the membrane) and at 1, 2, 3, 4, 5, and 6 weeks (immediately prior to removal of the membrane). Glycosaminoglycan levels were determined using an Alcian blue dye detection system. RESULTS: The mean levels of chondroitin sulfate and total sulfated glycosaminoglycans in GCF significantly decreased during the first 4 weeks after GTR surgery. By week 5, the levels began to rise, and by week 6 the levels had returned to baseline levels. CONCLUSIONS: Sulfated glycosaminoglycans can be monitored in GCF at healing GTR sites. It is proposed that this is a useful means of monitoring the status of the regenerating tissues. However, further longitudinal studies are required to assess if the sulfated glycosaminoglycans can be used as indicators of tissue maturation under guided tissue membranes used to treat periodontal defects. PMID- 10695933 TI - Treatment of peri-implant defects with combination growth factor cement. AB - BACKGROUND: The use of growth factor agents in the regeneration of oral tissues is an area of current investigation. Combinations of growth factors have been used synergistically to improve tissue regeneration. The aim of this study was to determine the effects of a combination growth factor cement (GFC) on guided bone regeneration around dental implants. METHODS: A combination of bone morphogenetic protein-2 (BMP-2), transforming growth factor-beta (TGF-beta), platelet-derived growth factor (PDGF), and basic fibroblast growth factor (bFGF) was used in a bioabsorbable, non-hydroxyapatite, calcium phosphate cement. Five adult hound dogs were used to compare the effects of GFC, plain cement, and control (no cement). The right and left second, third, and fourth mandibular premolar teeth were extracted; the implant osteotomies were prepared; and a uniform circumferential gap was prepared 1.5 mm beyond the width of the implant in the coronal half of the osteotomy for cement placement. Titanium machine-polished dental implants were placed in the prepared sites, and coronal defects were treated according to previously randomized, assigned modality. A bioabsorbable collagen membrane was secured over the control site, and the flaps were closed primarily. The dogs were maintained on a soft diet to avoid soft tissue trauma. The dogs were sacrificed at 3 months. The specimens were sectioned, mounted, and stained with Stevenel's blue and van Gieson's picric fuchsin. The bone-to-implant contact and bone 1 mm peripheral to the implant surface were recorded with a computerized microscopic digitizer. RESULTS: The findings of this study indicate a significant effect of GFC on increased bone-to-implant contact and amount of bone per surface area compared with the other treatment modalities (P <0.0009). Plain cement demonstrated slight but nonsignificant increases compared with the control (P>0.05). CONCLUSIONS: GFC increases bone-to-implant contact and bone surface area within peri-implant defects. Further studies may be beneficial to determine the feasibility of its use for other regenerative applications. PMID- 10695934 TI - Persistence patterns of Porphyromonas gingivalis, Prevotella intermedia/nigrescens, and Actinobacillus actinomyetemcomitans after mechanical therapy of periodontal disease. AB - BACKGROUND: The aim of this study was to determine the distribution patterns of Porphyromonas gingivalis, Prevotella intermedia/nigrescens, and Actinobacillus actinomycetemcomitans in periodontitis patients after standard mechanical periodontal therapy, and to determine factors increasing the odds to detect these target organisms in treated sites. METHODS: Eight hundred fifty-two (852) separate subgingival microbial samples were taken from the mesial and distal aspects of every tooth in 17 patients. Target organisms were identified culturally. RESULTS: The 3 microorganisms showed different persistence patterns: P. gingivalis was detected in a high percentage of subjects (59%), but in a low proportion of sites (5.4%). P. intermedia/nigrescens was detected in all subjects except one, and in 40.6% of the tested sites. Only 5 subjects were A. actinomycetemcomitans positive, but 2 of them showed a very high number of positive sites (44% and 75%, respectively). A highly significant relationship was found between a subject's tendency to bleed upon sampling and the number of P. intermedia/nigrescens-positive sites. A significant portion of the variation in frequency of persisting P. gingivalis could be explained by the frequency of persisting pockets deeper than 4 mm. No similar relationship could be established between clinical parameters and A. actinomycetemcomitans. On a site level, the odds of detecting P. gingivalis increased by a factor of 2.47 (P= 0.0001) for every millimeter of residual probing depth; the odds of detecting P. intermedia/nigrescens increased by a factor of 1.84 (P= 0.0001). CONCLUSIONS: If, after standard mechanical periodontal therapy, a large number of sites continue to bleed, one may expect an increased number of sites positive for P. intermedia/ nigrescens. If many deep pockets persist, a greater number of P. gingivalis positive sites can be expected. PMID- 10695936 TI - Enamel matrix-derived protein stimulates attachment of periodontal ligament fibroblasts and enhances alkaline phosphatase activity and transforming growth factor beta1 release of periodontal ligament and gingival fibroblasts. AB - BACKGROUND: Although it is claimed that enamel matrix-derived proteins (EMP) can be used to promote new attachment formation around periodontally involved teeth, the underlying biological mechanism is not understood. It was the aim of the present study to investigate the effects of EMP on the behavior of human periodontal ligament (HPLF) and gingival fibroblasts (HGF) in vitro, with special focus on their attachment properties, the expression of alkaline phosphatase (ALP) activity, the release of transforming growth factor (TGF)beta1, and their proliferative rate. METHODS: Fibroblast populations were obtained from 10 individuals with a healthy periodontium and cultured in chemically defined medium on culture plates coated with EMP, purified collagen type I, or their respective vehicles. Experiments were performed in the absence of serum for periods up to 48 hours. RESULTS: It was shown that HGF barely attached and spread on EMP-coated substrata, whereas HPLF attached and spread within 24 hours. However, when cultured on purified collagen type I, both cell types showed rapid attachment and spreading. Furthermore, the expression of ALP activity was significantly enhanced under the influence of EMP, especially in HPLF. HPLF and HGF both released significantly higher levels of TGFbeta1 in the presence of EMP. EMP did not influence 3H-thymidine incorporation by HPLF and HGF. CONCLUSIONS: Our results indicate that HPLF and HGF respond differently to EMP. A more rapid attachment of HPLF to this substratum might contribute, during the initial stages of periodontal healing, to selective outgrowth and colonization of exposed root surfaces in vivo. PMID- 10695935 TI - The effect of locally delivered controlled-release doxycycline or scaling and root planing on periodontal maintenance patients over 9 months. AB - BACKGROUND: This research report evaluates clinical changes resulting from local delivery of doxycycline hyclate (DH) or traditional scaling and root planing (SRP) in a group of patients undergoing supportive periodontal therapy (SPT). METHODS: In all, 141 patients received either DH (67) or SRP (74) treatment in sites > or =5 mm on one-half of their dentition at baseline and month 4. RESULTS: Clinical results were determined at month 9. Baseline mean probing depth recordings were similar between the two groups (DH = 5.9 mm; SRP = 5.9 mm). Mean month 9 results showed similar clinical results for attachment level gain (DH 0.7 mm; SRP 0.8 mm) and probing depth reduction (DH 1.3 mm; SRP 1.1 mm). Percentage of sites showing > or =2 mm attachment level gain at month 9 was 24.7% in the DH group and 21.2% in the SRP group. Thirty-nine percent (39%) of DH sites and 38% of SRP sites showed > or =2 mm probing depth reduction. When treated sides of the dentition were compared to untreated sides, DH showed a difference in disease activity (> or =2 mm attachment loss) from 19.3% (untreated) to 7.2% (treated); and SRP from 14.3% (untreated) to 8.1% (treated). CONCLUSIONS: Results show that both DH without concomitant mechanical instrumentation and SRP were equally effective as SPT in this patient group over the 9-month study period. PMID- 10695937 TI - Immunolocalizaiton of c-Myc and bcl-2 proto-oncogene products in gingival hyperplasia induced by nifedipine and phenytoin. AB - BACKGROUND: Hyperplastic gingival tissues show the histopathological characteristics of thick parakeratinized squamous epithelia with acanthosis and rete pegs elongated into the lamina propria. However, the pathogenic factors that contribute to the epithelial morphogenesis of this disease are obscure and remain to be studied. METHODS: We immunohistochemically examined the expression of both c-Myc and bcl-2 oncoprotein, which can exert influence on the epithelial morphogenesis and homeostasis, in 12 hyperplastic gingival tissues induced by nifedipine and phenytoin as well as 5 control tissues using avidin-biotin peroxidase complex methods. RESULTS: Four specimens out of 5 nifedipine-induced and 5 out of 7 phenytoin-induced hyperplastic gingival tissues revealed the expression of c-Myc oncoprotein, whereas no significant immunostaining of c-Myc oncoprotein was found in 5 control tissues. The c-Myc oncoprotein-positive cells were observed to be localized in the basal and suprabasal cell layers of the hyperplastic gingival epithelia. Although all of the 12 hyperplastic gingival epithelia showed the distribution of bcl-2 oncoprotein in the basal and suprabasal layer cells, in 5 control epithelia the bcl-2 oncoprotein expression was slight and confined to the basal layer cells. CONCLUSIONS: The results of our present study indicate that the synergistic overexpression of c-Myc and bcl-2 oncoprotein may be related to the pathogenesis of gingival hyperplasia induced by nifedipine and phenytoin, especially to the morphogenesis of hyperplastic epithelia. PMID- 10695938 TI - Levels of leukotriene B4 and platelet activating factor in gingival crevicular fluid in renal transplant patients receiving cyclosporine A. AB - BACKGROUND: Cyclosporine A (CsA) is a potent immunosuppressant effectively used to prevent organ transplant rejection and also to treat several systemic diseases. CsA-induced gingival overgrowth (CsA GO) is the most widely seen side effect of this drug; its pathogenesis is not completely understood. The aim of the present study was to identify the role of leukotriene B4 (LTB4) and platelet activating factor (PAF) in the pathogenesis of CsA GO. METHODS: LTB4 and PAF levels were detected in gingival crevicular fluid (GCF) samples from renal transplant patients receiving CsA therapy and exhibiting CsA GO, from patients with gingivitis and from periodontally healthy subjects. Plaque index, papilla bleeding index, and hyperplastic index were recorded at each study site. GCF samples and clinical data were obtained from: 2 sites exhibiting CsA GO (CsA GO+) and 2 sites not exhibiting CsA GO (CsA GO-) in each CsA-treated patient; 2 diseased sites in each patient with gingivitis; and 2 healthy sites in each subject with clinically healthy periodontium. LTB4 was extracted from the samples by solid-phase method using C18 cartridge and purified by high-performance liquid chromatographic (HPLC) method and analyzed by radioimmunoassay (RIA). PAF was extracted from GCF samples passing through amberlit resin columns, purified by HPLC, and analyzed by RIA. RESULTS: Total amounts of LTB4 and PAF in GCF were higher in CsA GO+ sites compared to the healthy sites from healthy controls. However, the amount of LTB4 and PAF elevation in CsA GO+ sites was not significantly higher than those in diseased sites. Clinical degrees of gingival inflammation were also similar between CsA GO+ and diseased sites. LTB4 and PAF total amounts in GCF were higher in CsA GO+ sites compared to CsA GO- sites in the same subjects, but this difference just failed to reach significance. Similar findings were obtained with concentration data. CONCLUSIONS: The results of this study indicate that CsA therapy does not have a significant effect on GCF LTB4 and PAF levels and that gingival inflammation seems to be the main reason for their elevation. In CsA-treated patients, alterations in LTB4 and PAF levels might play a role in CsA GO through some asyet unknown mechanism. To our knowledge, this is the first report describing the levels of lipid mediators in GCF of CsA-treated patients. We assume that further studies will contribute to the understanding of the pathogenesis of CsA-induced gingival overgrowth. PMID- 10695939 TI - Effect of cigarette smoking on oral elastase activity in adult periodontitis patients. AB - BACKGROUND: We have previously reported that elastase activity in oral fluids is significantly increased in most adult periodontitis patients. In some patients, however, elastase levels remain low despite the presence of deep periodontal pockets. In this study we explored whether or not smoking is related to the unexpected low elastase values in these patients. METHODS: We determined what proportion of the periodontitis patients that showed low oral elastase values were smokers. Paraffin-stimulated saliva or oral rinse samples (3 ml of water, 30 second rinse) were assayed for elastase activity by incubating with 1 mM succinyl alanyl-alanyl-valine-p-nitroanilide for 20 hours at 37 degrees C, and the color formation read with a spectrophotometer. Neutrophil numbers were analyzed by staining the cells in the oral rinse smear samples. RESULTS: In 2 patient groups, one in Helsinki, Finland (n = 46) and the other in Vancouver, British Columbia (n = 25), 63% and 83%, respectively, of the adult periodontitis patients who had one or more pockets > or =6 mm and had low oral elastase values (increase of optical density <0.5) were smokers. Non-smoking periodontitis patients had elevated neutrophil numbers compared to healthy subjects, while the smoking patients showed no significant change. Next we analyzed elastase levels in stimulated whole saliva in a group of smokers (n = 300) and those who had quit smoking (n = 102). Smokers had significantly lower oral elastase levels than former smokers in both advanced and moderate periodontitis groups. In this subject group, 56% of all smokers with periodontitis (at least one pocket > or =6 mm) had oral elastase values less than 0.5 U while only 31% of those patients who had quit smoking had low values. CONCLUSIONS: Cigarette smoking leads to lowered elastase and neutrophil levels in the oral cavity. The oral neutrophil elastase assay, therefore, cannot be used to measure the periodontal status of smokers. PMID- 10695940 TI - Employing a transgenic animal model to obtain cementoblasts in vitro. AB - BACKGROUND: Proper formation of cementum, a mineralized tissue lining the tooth root surface, is required for development of a functional periodontal ligament. Further, the presence of healthy cementum is considered to be an important criterion for predictable restoration of periodontal tissues lost as a consequence of disease. Despite the significance of cementum to general oral health, the mechanisms controlling development and regeneration of this tissue are not well understood and research has been hampered by the lack of adequate in vitro experimental models. METHODS: In an effort to establish cementoblast cell populations, without the trappings of a heterogeneous population containing periodontal ligament (PDL) cells, cells were obtained from the root surface of first mandibular molars of OC-TAg transgenic mice. These mice contain the SV40 large T-antigen (TAg) under control of the osteocalcin (OC) promoter. Therefore, only cells that express OC also express TAg and are immortalized in vitro. Based on results of prior in situ studies, OC is expressed by cementoblasts during root development, but not by cells within the PDL. Consequently, when populations are isolated from developing molars using collagenase/trypsin digestion, only cementoblasts, not PDL cells, are immortalized and thus, will survive in culture. RESULTS: The resulting immortalized cementoblast population (OC/CM) expressed bone sialoprotein (BSP), osteopontin (OPN), and OC, markers selective to cells lining the root surface. These cells also expressed type I and XII collagen and type I PTH/PTHrP receptor (PTH1R). In addition to expression of genes associated with cementoblasts, OC/CM cells promoted mineral nodule formation and exhibited a PTHrP mediated cAMP response. CONCLUSIONS: This approach for establishing cementoblasts in vitro provides a model to study cementogenesis as required to enhance our knowledge of the mechanisms controlling development, maintenance, and regeneration of periodontal tissues. PMID- 10695941 TI - Effect of cigarette smoking on periodontal status of healthy young adults. AB - BACKGROUND: It has been shown that tobacco is a significant risk factor for periodontal disease; however, there have been few studies on young populations where problems of general health can be discounted. The purpose of this study was to examine the influence of tobacco consumption on the periodontal condition of a young, healthy population. METHODS: The study population consisted of 304 young Caucasian males (average age 19.38 +/- 0.72 years) entering the Armed Forces. All the subjects completed a self-administered questionnaire on age, oral hygiene habits, previous dental examinations, and quantity and length of tobacco use. The periodontal examination consisted of the plaque index (PI); periodontal bleeding index (PBI); probing depth (PD); and clinical attachment level (CAL). One- and 2 way ANOVA was used to compare data recorded between smokers and non-smokers. RESULTS: Forty-six percent of subjects reported that they brushed their teeth at least once a day, but only 13% visited a dentist at least once a year. Over half (53%) were habitual smokers, 43% smoking between 5 and 20 cigarettes per day; 39% of the smokers had been smoking for less than 5 years. Mean PI was 31.24 +/- 14.88 (27.19 +/- 15.93 for smokers and 35.78 +/- 12.17 for non-smokers), with significant differences between non-smokers and those who smoked 5 to 20 cigarettes per day (26.85 +/- 16.11, P<0.0001). Mean PBI was 42.29 +/- 8.43 (non smokers 44.67 +/- 6.53 and smokers 40.17 +/- 9.46). Significant differences were found between the PBI of the non-smokers and of those who smoked 5 to 20 cigarettes per day (39.90 +/- 9.64, P <0.0001). There were also differences in the PBI between those who brushed their teeth once (40.53 +/- 9.61) and twice (44.86 +/- 5.9) a day (P<0.0001). Mean PD was 1.62 +/- 0.43 mm (non-smokers 1.56 +/- 0.36 and smokers 1.68 +/- 0.49). Deeper probing depths were recorded among smokers than among non-smokers, with statistically significant differences (P<0.049); statistically significant differences were also found between those who attended (1.49 +/- 0.50) and those who did not attend (1.65 +/- 0.42) regular dental check-ups (P<0.031). Mean CAL 1.75 +/- 0.41 (non-smokers 1.64 +/- 0.32 and smokers 1.82 +/- 0.44). CONCLUSIONS: It may be concluded that, even at such an early age, tobacco consumption affects the periodontal health. It is necessary to inform young smokers of the risk of tobacco use regarding periodontal health. PMID- 10695942 TI - Repeated metronidazole and amoxicillin treatment of periodontitis. A follow-up study. AB - BACKGROUND: The prevailing concept is that little or no clear benefit is derived from antibiotic therapy in chronic periodontitis. Studies to determine the effect of metronidazole plus amoxicillin (M+A) on adult periodontitis are questionable because standard design for clinical trials was usually not used. In addition, there is no information about the effect of M+A as the sole therapy for periodontitis. METHODS: A randomized, triple-blind, controlled clinical trial was used to determine the effect of systemic administration of M+A, as the sole therapy, in progressive adult periodontitis. Forty-six subjects with moderate to advanced adult periodontitis who showed > or =2 mm attachment loss in at least 2 sites in the previous 2 months were entered in the study. Subjects were randomly distributed to a group who received 21 tablets of metronidazole 250 mg plus amoxicillin 500 mg, or to a group receiving a placebo (1 tablet every 8 hours for 1 week). Patients were examined every 2 months for 12 months. The M+A or placebo regimen was repeated at 4 and 8 months. No effort was made to change the oral habits of patients and they received no additional therapy. Differences between groups were assessed using the Mann-Whitney U test. The differences at every 2 month interval within each group were assessed using the ANOVA test. RESULTS: Seven subjects abandoned the study; at 12 months the M+A group had 20 subjects and the placebo group 19. There were no significant differences in the clinical parameters at baseline between the 2 groups. After 2 months and thereafter, the M+A group showed significant clinical improvement while the placebo group showed a progressive deterioration of periodontal status. At 12 months compared to baseline, subjects of the M+A group showed: 1) a significant overall mean attachment gain of 0.43 mm (P = 0.005); 2) a significant decrease of active sites (P< or =0.03); 3) a significant increase of sites gaining attachment level (P< or =0.01); 4) a significant reduction of pocket depth (P< or =0.00006); and 5) a significant decrease in percentage of bleeding on probing sites (BOP) (P< or =0.0005). Significant differences between both groups at all 2-month evaluations were found in overall mean attachment level (P < or =0.000004), in percent of active sites (P< or =0.03), and in percent of BOP sites (P< or =0.02). Sites exhibiting > or =2 mm of attachment loss in 2 successive or alternate evaluations, and periodontal abscess were noticed only in the placebo group. CONCLUSIONS: A 1-week course of systemic M+A every 4 months, as the only therapy, arrests the progression of adult periodontitis and significantly improves the clinical parameters of the disease. PMID- 10695943 TI - Intraoral fluoride releasing device: a new clinical therapy for dentine sensitivity. AB - BACKGROUND: Dentinal sensitivity (DS) occurs frequently in adult populations in western countries. The purpose of this work was to assess the effectiveness of a new intraoral fluoride releasing device (IFRD) in reducing the level of pain in patients with primary or postsurgical dentine sensitivity. METHODS: A total of 49 individuals were selected for this study, 15 of whom had post-periodontal surgery dentine sensitivity and 34 with primary sensitivity. An IFRD was applied to 39, while 10 received a placebo device. All individuals in the control group suffered from primary sensitivity. The IFRD used in this study consists of sodium fluoride encased in an acrylic polymer which releases fluoride at a rate of approximately 0.04 mg/day. All patients were asked to rinse with cold water (10 degrees C) and to indicate the level of pain on a 0 to 10 visual analog scale, 0 equalling "no pain" and 10 "maximum bearable pain." All subjects were evaluated once a week during 4 months. Statistical analysis of dentine sensitivity was performed as a univariate study, in relation to the main factors: age, gender, and primary sensitivity or postsurgical etiology. RESULTS: Symptoms decreased dramatically in all treated patients. The level of sensitivity did not change during the first week after IFRD application, but decreased significantly within the fourth week and remained absent through the duration of the treatment (P <0.01). Difference in sensitivity with respect to different etiology was significant only after 4 weeks (P= 0.01), while there was no statistical difference with respect to age or gender. CONCLUSIONS: This paper is an initial study to evaluate the effectiveness of the IFRD. The method is fast, painless, inexpensive, and it appears to be suitable as a routine treatment. The presented data support the conclusions at this stage and warrant more comprehensive evaluation. PMID- 10695944 TI - Periodontal probing: probe tip diameter. AB - BACKGROUND: Periodontal probing is one of the most common methods used in diagnosing periodontal disease. The purpose of this study was to determine the importance of the diameter of periodontal probing tips in diagnosing and evaluating periodontal disease. METHODS: The literature discussing periodontal probe diameters in human, dog, and monkey studies was reviewed and compared. Tip diameters varied from 0.4 to over 1.0 mm in these studies. Probe advancement between the gingiva and the tooth is determined by the pressure exerted on the gingival tissues and resistance from the healthy or inflamed tissue. The pressure is directly proportionate to the force on the probe and inversely proportionate to the probe tip diameter. The larger probing diameters reduced probe advancement into inflamed connective tissue. This effect of change in probe diameter reduced the pressure in a greater manner than an increase of similar change in probe force. RESULTS: In the studies reviewed, the pressure used to place the probe tip at the base of the periodontal sulcus/pocket was approximately 50 N/cm2 and at the base of the junctional epithelium, 200 N/cm2. A tip diameter of 0.6 mm was needed to reach the base of the pocket. Clinical inflammation did not necessarily reflect the severity of histological inflammation, and the recordings may not illustrate probing depth. Furthermore, probing depth did not identify anatomical locations at the base of the pocket. CONCLUSIONS: Probe tips need to have a diameter of 0.6 mm and a 0.20 gram force (50 N/cm2) to obtain a pressure which demonstrates approximate probing depth. This pressure was needed to measure the reduction of clinical probing depth, which included formation of a long junctional epithelium as a result of therapy. In addition, different forces or diameter tips are needed to measure healthy or inflamed histological periodontal probing depths. PMID- 10695945 TI - Chronic ulcerative stomatitis: a case report. AB - BACKGROUND: Certain mucocutaneous diseases present with painful, ulcerative, or erosive oral manifestations. Chronic ulcerative stomatitis is a newly recognized disease of unknown origin which presents clinically with features of desquamative gingivitis. This report marks only the thirteenth case reported in the world literature. A review of previous reports and studies is presented along with a review of immunofluorescence techniques critical to proper diagnosis. These diseases are difficult to diagnose without the use of immunofluorescence techniques. A 54-year-old Caucasian woman presented with a 2- to 3-year history of stomatitis and dry mouth. METHODS: Direct immunofluorescence revealed a speckled pattern of IgG deposits in the basal one-third of the epithelium, while indirect immunofluorescence confirmed the presence of stratified epithelium specific antinuclear antigen (SES-ANA), both pathognomonic for chronic ulcerative stomatitis. RESULTS: The patient was successfully treated using topical corticosteroid therapy. PMID- 10695946 TI - Gingival metastasis from a medullary thyroid carcinoma: case report. AB - BACKGROUND: Metastatic tumors to the oral cavity are rare, representing about 1% of oral tumors, and they affect jaws more often than the oral soft tissues. METHODS: Fifteen cases of metastases to the jaw bones from thyroid carcinoma were found in a recent review, with no cases located in the oral mucosa. RESULTS: The authors describe the first cases of gingival metastasis from a thyroid medullary carcinoma. CONCLUSIONS: Periodontists must recognize oral soft tissue metastases because they can be the first sign of an undiscovered malignancy, and they can be easily mistaken with several different benign lesions. PMID- 10695947 TI - Primary gingival malignant melanoma. Report of 3 cases. AB - BACKGROUND: Malignant melanoma is rare in the oral cavity and accounts for less than 1% of all melanomas. Nevertheless, the disease can be fatal, and early diagnosis and treatment may improve prognosis dramatically. The purpose of this paper is to report 3 new cases of primary malignant melanoma of the oral cavity arising in the gingiva, and to review the literature regarding intraoral melanoma. METHODS: Three cases are presented. One case was in the right mandibular molar area; the second in the right maxillary canine-premolar area; and the third in the left mandibular canine-premolar region. All patients were treated surgically, with postoperative radiotherapy. RESULTS: The first patient lived for 2 years and the second for 3 years before distant metastases were diagnosed from which they subsequently died. The third patient was lost from follow-up after 18 months. CONCLUSION: Primary oral malignant melanoma is a deadly disease. Early suspicion of this disease will allow prompt treatment and increase the prognosis for these patients. PMID- 10695948 TI - The role of controlled drug delivery for periodontitis. The Research, Science and Therapy Committee of the American Academy of Periodontology. PMID- 10695949 TI - Evidence for distinct attentional bottlenecks in attention switching and attentional blink tasks. AB - E. Weichselgartner and G. A. Sperling (1987), using rapid serial visual presentation (RSVP), estimated that attention could be moved to a new spatial location within 300-400 ms. H. J. Muller and P. M. Rabbit (1989) used a spatial cuing task and found a similar time course for voluntarily redeploying attention. A separate phenomenon known as the attentional blink (AB) also follows a similar time course, yet occurs when participants attend to a single spatial location. The present study found that attention can be shifted more quickly than previously estimated and that part of the deficit observed during searches of spatially distinct RSVP streams is due to an AB. The results support some early and late selection accounts for the temporal dynamics of visual attention and suggest different bottlenecks during visual selection. The implications for visual search and visual processing are discussed. PMID- 10695950 TI - The effects of scene inversion on change blindness. AB - In two experiments, participants searched for a difference between two views of a scene. In Experiment 1, the authors extended the change-blindness findings from previous work by R. A. Rensink, J. K. O'Regan, and J. J. Clark (1997), which used an experimenter-induced global transient, to a less artificial situation in which participants searched for a difference in a pair of photographic images presented simultaneously. To examine the idea that meaning-driven endogenous orienting was responsible for the previously observed advantage for changes in center-of interest items, the authors inverted half of the image pairs. The advantage for center-of-interest items was replicated with upright displays, but it was completely eliminated by inversion, strongly supporting the role of meaning driven endogenous orienting in this task. With flickering displays (Experiment 2), the center-of-interest effect was completely unaffected by inversion. The authors suggest that when change blindness is induced via flicker, scene modifications are typically found by stimulus-driven rather than by meaning driven processes. PMID- 10695951 TI - Selective attention in animal discrimination learning. AB - The traditional approach to the study of selective attention in animal discrimination learning has been to ask if animals are capable of the central selective processing of stimuli, such that certain aspects of the discriminative stimuli are partially or wholly ignored while their relationships to each other, or other relevant stimuli, are processed. A notable characteristic of this research has been that procedures involve the acquisition of discriminations, and the issue of concern is whether learning is selectively determined by the stimulus dimension defined by the discriminative stimuli. Although there is support for this kind of selective attention, in many cases, simpler nonattentional accounts are sufficient to explain the results. An alternative approach involves procedures more similar to those used in human information processing research. When selective attention is studied in humans, it generally involves the steady state performance of tasks for which there is limited time allowed for stimulus input and a relatively large amount of relevant information to be processed; thus, attention must be selective or divided. When this approach is applied to animals and alternative accounts have been ruled out, stronger evidence for selective or divided attention in animals has been found. Similar processes are thought to be involved when animals search more natural environments for targets. Finally, an attempt is made to distinguish these top down attentional processes from more automatic preattentional processes that have been studied in humans and other animals. PMID- 10695952 TI - Basic response time tools for studying general processing capacity in attention, perception, and cognition. AB - One of the more important constructs in the study of attention, perception, and cognition is that of capacity. The authors reviewed some of the common meanings of this construct and proposed a more precise treatment. They showed how the distribution of response times can be used to derive measures of process capacity and to further illustrate how these measures can be used to address important hypotheses in cognition. PMID- 10695953 TI - Summary: let's pay attention to attention. PMID- 10695954 TI - Hormone replacement therapy. Clinical synthesis panel on HRT. PMID- 10695955 TI - Contents of HRT and mechanisms of action. AB - Formulation of HRT preparations has changed very significantly over the last few decades. The problems of unopposed oestrogens are now well recognised. Addition of progestins may overcome these problems in some target tissues. However, there is clear evidence for differential effects of both steroids in different target tissues. It is also vital to be clear on the dose-response to each steroid in each target tissue. Steroids given orally may undergo different metabolism from those given transdermally or subcutaneously. This can mean that, even given dose for dose, there can still be differences in the regulation of, for example, lipid profiles, depending on delivery route. The situation is further complicated by the discovery that there is more than one receptor for each steroid. Steroid receptors are members of the super-family of nuclear receptors. Steroid enters the cell, binds and activates empty receptors, inducing dimerisation and acquisition of high affinity for specific sequences of nucleotides within the hormone-response elements (HRE) of the target genes. Hormone-receptor complex interacts with the HRE to modulate transcription of the gene. The different receptors for each steroid (e.g. oestrogen receptors alpha and beta) help promote the differential responses in different target tissues. Analysis of the overall responses to HRT requires a knowledge of the dose and specificity of each steroid within the formulation, together with an understanding of how each steroid regulates the activities of each sub-group of receptors in each target tissue, allowing for the appropriate metabolism of the primary steroids. Additionally, there will be variations among different individuals. PMID- 10695956 TI - Menopause, HRT and menopausal symptoms. AB - Menopause is defined as the final menstruation, directly preceding the permanent cessation of ovarian follicular function. The transition from the reproductive to the non-reproductive phase of life can take many years, frequently characterised by perimenopausal cycle disorders, vasomotor symptoms (hot flushes and night sweats) and urogenital complaints (vaginal dryness, micturition complaints). In addition to these typical climacteric symptoms other, more atypical symptoms can be present, such as tiredness, irritability and mood swings. Topical and systemic oral and transdermal HRTs are generally very effective as symptomatic treatment, but other non-hormonal options can also be considered. PMID- 10695957 TI - HRT in the prevention and treatment of osteoporosis. AB - Many short term studies have shown that various HRT regimens prevent bone loss at all skeletal sites and normalise biochemical markers at bone turnover. Most, if not all, evidence is derived from cohort and case-control studies. Of concern is the observation that past HRT users are not protected against hip fracture. More attention should be devoted in the future to the long term use of a low-dose combined continuous HRT regimen in late post-menopausal women in order to reduce the burden of osteoporotic fractures. PMID- 10695958 TI - HRT and cardiovascular disease. AB - Conclusions influencing clinical decisions about the effects of hormone replacement therapy (HRT) on cardiovascular disease have undergone major revisions over the last 30 years. Assumptions in the early 1970s that HRT would increase thromboembolic conditions were reversed in the 1980s, when the first formal studies seemed to indicate a cardioprotective effect. The Heart and Estrogen/progestin Replacement Study (HERS) has, however, very recently suggested that HRT does not affect the incidence of coronary heart disease (CHD) one way or the other. These roller-coaster changes of direction have made decisions by doctors and their patients about the long-term use of HRT extremely difficult and need to be replaced by the stability that only further randomised controlled trials (RCTs) can now provide. PMID- 10695959 TI - Use of HRT and the subsequent risk of cancer. AB - At least 20 million women in developed countries are estimated to be currently using hormone replacement therapy (HRT). Almost 100 epidemiological studies have reported on the relationship between the use of HRT and the risk of cancer of female reproductive organs, namely the breast, uterus or ovary. Cancer at these sites is common and there are a priori reasons why the use of hormonal therapy to 'replace' the endogenous production of ovarian hormones after the menopause might increase the risk of these cancers. The available evidence indicates that the risk of breast cancer or endometrial cancer is increased while women are using HRT, the risk increasing with increasing duration of use. Most of the evidence about these cancers relates to use of HRT preparations containing oestrogens alone. The limited evidence about combination therapy, with oestrogens and progestogens, suggests that, compared to oestrogens alone, the effect on the breast is similar, but the effect on the endometrium is diminished, the diminution in risk being greater the more days each month that progestogens are used. The effect of HRT on breast cancer wears off after use ceases and has disappeared largely, if not wholly, within 5 years, whereas the effects on endometrial cancer take longer to wear off, if at all. The breast and endometrial cancers that are diagnosed in HRT users are less aggressive clinically than cancers in never-users but, as yet, there is little reliable information about the relationship between use of HRT and mortality from these cancers. For other cancer sites, the existing data about the effects of HRT are inconclusive. The longer the period of use of HRT, the greater the excess incidence of cancer of the breast and endometrium is likely to be. Use of HRT for short periods of time should have little effect on the incidence of these cancers. The cumulative excess incidence in 1000 women who used HRT for 10 years, beginning at age 50, is estimated to be six for breast cancer, 42 for endometrial cancer in women with an intact uterus using oestrogen therapy alone and about 20 for endometrial cancer in women with an intact uterus using oestrogen-progestogen combinations. The estimate for combined therapy is based on small numbers and may well vary with the type of preparation used. The overall balance between the excess incidence of these cancers and other effects of HRT needs to be evaluated carefully and will require more reliable data than exist at present. PMID- 10695960 TI - HRT and women who have had breast or endometrial cancer. AB - The benefits of oestrogen replacement therapy (ERT) in preventing vasomotor symptoms, cardiovascular disease, osteoporosis, and colon cancer are well documented. Other potential benefits i.e. dementia and macular degeneration are being investigated. Although oestrogen is said to be contraindicated in women previously treated for breast and endometrial cancer, there is no data to support this admonition. Preliminary data would suggest ERT can be used safely in women who have had these cancers. Prospective randomised studies are currently on going in the United States and Europe addressing ERT in previously treated breast and endometrial cancer. Informed consent, patients' desires, and benefit-risk considerations are all part of information the woman needs to make a decision concerning ERT. PMID- 10695961 TI - HRT and dementia. AB - The role of HRT and/or oestrogens in the aetiology, progression and prevention of neurodegenerative disorders is unclear. Various studies have been conducted in cells, animals and humans to better define these relationships. The following is a presentation of data on the role of oestrogens in the two most common dementia syndromes Alzheimer's Disease and vascular dementia. This review examines whether there is support for the hypothesis that the use of oestrogens reduces the risk for dementia and cognitive decline in a variety of experimental models. A literature search was performed using the following key words: oestrogen and dementia, oestrogen and Alzheimer, oestrogen and cognition, oestrogen and learning, and oestrogen and memory. The reference lists for the articles identified using these search strategies were examined. The strongest evidence for a beneficial effect comes from cell and animal studies. These suggest that treatment with oestrogens may influence the pathogenetic processes of AD, mainly by affecting the beta-amyloid metabolism, by promoting the activity of the cholinergic system, by reducing oxidative stress or cardiovascular risk. Observational studies give some support to the hypothesis that HRT may be protective for cognitive decline and dementia in the elderly, but the inherited methodological problems in these studies preclude any conclusions. The treatment trials published thus far have methodological problems that prohibit definite conclusions on the relationship between HRT and cognitive decline and dementia. PMID- 10695962 TI - Indigenous knowledge: an inexhaustible "information bank" for toxin research. PMID- 10695963 TI - Muscarinic toxins: novel pharmacological tools for the muscarinic cholinergic system. AB - Muscarinic receptors are widely spread throughout the body, and are involved in the regulation of fundamental physiological processes, like the modulation of the heart rate, control of motor systems and modulation of learning and memory. In the central nervous system the cholinergic transmission is mainly mediated by muscarinic receptors; there are five subtypes that are all expressed in the brain of mammals (m1-m5). There are regional differences in their concentrations in the brain and more than one subtype is expressed in the same cell. It has been difficult to study their localization and function in vivo due to the lack of ligands that exclusively act on one subtype of the receptor. We studied the action of the muscarinic toxins MT1, MT2 and MT3, from the venom of the snake Dendroaspis angusticeps, on muscarinic receptors, by using the classical muscarinic radioligand 3H-NMS as reporter of the inhibition of its own binding, to either native or cloned receptors. We have also studied the in vivo effects on memory retention of the injection of the toxins into discrete brain regions. The muscarinic toxins appear to be invaluable tools to study receptor pharmacology, physiology and structure/function relationships. They would enable the design of new, more selective, pharmacological agents. PMID- 10695964 TI - Paralytic toxins in a ribbon worm Cephalothrix species (Nemertean) adherent to cultured oysters in Hiroshima Bay, Hiroshima Prefecture, Japan. AB - In 1998, during the surveillance of the toxicity of various marine fouling organisms in Hiroshima Bay, Hiroshima Prefecture, Japan, specimens of the ribbon worm, "himomushi" Cephalothrix sp. (Nemertean) adherent to the shells of cultured oysters hanging onto floating culture rafts were found to contain toxins which showed strong paralytic action in mice throughout the survey period, February to May. The maximum toxicity (as tetrodotoxin, TTX) was 14,734 MU/g whole body. Attempts were made to identify the paralytic toxins in this worm. The "himomushi" toxin (HMT) was extracted from the worm with 80% methanol acidified with acetic acid and the extract defatted with dichloromethane. The aqueous layer was chromatographed on activated charcoal and the unbound and bound toxic fractions were analyzed by high-performance liquid chromatography and gas chromatography mass spectrometry. It was rather unexpectedly revealed from these results that HMT was comprised of TTX, 4-epiTTX, anhydroTTX and three unidentified toxins. To our knowledge, this is the first report of the occurrence of toxic organisms, containing a high concentration of TTX, adherent to cultured bivalves such as oysters. PMID- 10695965 TI - Snake venom modulators of platelet adhesion receptors and their ligands. AB - In thrombosis, platelet aggregation is initiated by a specific membrane glycoprotein (GP) Ib-IX-V complex binding to its adhesive ligand, von Willebrand factor, in the matrix of ruptured atherosclerotic plaques or in plasma exposed to high hydrodynamic shear stress. This process closely resembles normal haemostasis at high shear, where GP Ib-IX-V-dependent platelet adhesion to von Willebrand factor in the injured blood vessel wall initiates platelet activation and integrin alphaIIb beta3 (GP IIb-IIIa)-dependent platelet aggregation. At low shear, other receptors such as those that bind collagen, the integrin alpha2beta1 (GP Ia-IIa) or GP VI, mediate platelet adhesion. Recently, snake venom proteins have been identified that selectively modulate platelet function, either promoting or inhibiting platelet aggregation by targeting GP Ib-IX-V, alpha2beta1, GP VI, alphaIIb beta3, or their respective ligands. Interestingly, these venom proteins typically belong to one of two major protein families, the C type lectin family or the metalloproteinase-disintegrins. This review focuses on recent insights into structure-activity relationships of snake venom proteins that regulate platelet function, and the ways in which these novel probes have contributed in unexpected ways to our understanding of the molecular mechanisms underlying thrombosis. PMID- 10695966 TI - Toxin 2 (PhTx2), a neurotoxic fraction from Phoneutria nigriventer spider venom, causes acute morphological changes in mouse skeletal muscle. AB - Phoneutria nigriventer (Labidognatha, Ctenidae) is a spider found in the warm regions of South America. Bites by this species cause intense local pain, autonomic dysfunction and paralysis. PhTx2, a neurotoxic fraction of the venom of this species, interferes with the physiology of sodium channel function. The present study describes the morphological changes in mouse phrenic nerve and diaphragm muscle after 15, 30, 45 and 60 min of incubation with 1 microg of PhTx2/ml. Light and transmission electron microscopy showed that PhTx2 caused progressive myonecrosis which involved swelling of the sarcoplasmic reticulum, mitochondrial damage, disorganization of the sarcomeres, zones of hypercontracted myofibrils and rupture of the plasma membrane. The intramuscular fascicles of the phrenic nerve showed vacuolated myelinated axons and Schwann cells. The neuromuscular junctions had vesicle-depleted nerve terminals with swollen mitochondria. The axolema was frequently invaginated and sequestered portions of the axoplasm, or was sometimes interrupted at the site of the synaptic gutter. The post-synaptic junctional folds were shallow and disperse. These morphological alterations in the muscle and nerve fibres were similar to those caused by osmotic disturbances and agree with the ability of PhTx2 to increase the permeability of sodium channels. An increase in sodium influx would probably be accompanied by an influx of water and an elevation in the concentration of cytosolic calcium as a result of calcium release by the sarcoplasmic reticulum and/or mitochondria and the entry of extracellular calcium. The morphological effects caused by PhTx2 were comparable to those seen with Phoneutria nigriventer whole venom which is known to activate and to delay the inactivation of sodium channels. We conclude that PhTx2 is probably the main toxic fraction responsible for such morphological alterations. PMID- 10695967 TI - Immunoaffinity purification method for detection and quantification of microcystins in lake water. AB - We have developed a new clean-up method, which consisted of solid-phase extraction on a Sep-Pak PS-2 (styrene-divinylbenzene copolymer) or Excelpak SPE GLF (polymethacrylate) cartridge instead of conventional ODS silica gel and silica gel together with following immunoaffinity purification using anti microcystin-LR monoclonal antibodies. This newly developed method was demonstrated to eliminate co-existing substances and to concentrate microcystins in the lake water. The recoveries from lake water (1 liter) spiked with 100 ng each of microcystins-RR, -YR and -LR were 85.5, 89.2 and 92.2%, respectively, with coefficients of variation of 3.3-7.6%. Only 3 h were required to complete the total procedures starting from the microcystin extraction, the immunoaffinity purification, and the quantification using HPLC. The detection limits for all of the 3 microcystins in lake water were 0.005 microg/l. Applicability of this method has been demonstrated by measuring the concentrations of microcystins in water samples collected from lakes where water blooms occurred, which turned out to be 0.012-0.177 microg/l of total microcystins. PMID- 10695968 TI - Identification of high molecular weight serine-proteases in Loxosceles intermedia (brown spider) venom. AB - High molecular weight serine-proteases have been identified in Loxosceles intermedia (brown spider) venom. The mechanism by which Loxosceles spp venoms cause dermonecrotic injury (a hallmark of loxoscelism) is currently under investigation, but it seems to be molecularly complex and in some instance proteases might be expected to play a role in this skin lesion. In the present investigation, when we submitted L. intermedia venom to linear gradient 3-20% SDS PAGE stained by a monochromatic silver method we detected a heterogeneous protein profile in molecular weight, ranging from 850- to 5-kDa. In an attempt to detect zymogen molecules of proteolytic enzymes, venom aliquots were treated with several exogenous proteases. Among them, trypsin activated two gelatinolytic molecules of 85- and 95-kDa in the venom. In experiments of hydrolysis inactivation using different protease inhibitors for four major class of proteases, we detected that only serine-type protease inhibitors were able to inactivate the 85- and 95-kDa enzymes in the venom. An examination of the 85- and 95-kDa gelatinolytic activities as a function of pH showed that these proteases had no apparent activities at pH below 5.0 and higher than 9.0 and displayed little activity at pH 6.0. with the optimal pH for their activities ranging from 7.0 to 8.0. Evaluation of the functional specificities of the 85- and 95-kDa venom proteases showed that these proteases efficiently degrade gelatin (denatured collagen) but have no proteolytic activity on hemoglobin, immunoglobulin, albumin, librinogen or laminin, suggesting specificity of their proteolytic actions. We describe here two serine-proteases activities in L. intermedia venom probably involved in the harmful effects of the venom. PMID- 10695969 TI - Investigation of the haemodynamic effects of Phoneutria nigriventer venom in anaesthetised rabbits. AB - The haemodynamic alterations induced by the central and peripheral administration of the armed spider (Phoneutria nigriventer) venom (PNV) were investigated in anaesthetised rabbits. The intracerebroventricular injection of increasing doses of PNV (30 and 100 microg/kg) elicited a biphasic cardiovascular response characterised by a brief hypotension (1-3 min) followed by a marked and sustained (more than 30 min) increase in mean arterial pressure (61 +/- 5 and 61 +/- 10%, respectively) and in systemic vascular resistance (135 +/- 21 and 161 +/- 37%) accompanied by mild increases in cardiac contractility. Systemic alterations such as salivation and muscular fasciculation were also observed. At the opposite, the dose of 100 microg/kg of PNV injected intravenously produced only a hypotensive effect (29 +/- 4% decrease in mean arterial pressure) and a decrease in vascular resistance (38 +/- 5%). Nevertheless, a much higher dose of PNV (1 mg/kg) injected intravenously produced a hypertensive response analogous to the one observed upon central administration. The central hypertensive response induced by PNV was not affected by preteating the animals with selective antagonists of receptors of different neurotransmitters or endogenous mediators such as: acethylcoline muscarinic, bradykinin B2, angiotensin II AT1 receptors and also antagonists of the excitatory amino acid receptors of the central nervous system. Nevertheless, the intravenous pretreatment with the selective alpha1-adrenergic receptor antagonist prazosin significantly blunted the excitatory cardiovascular response evoked by the central injection of PNV. It is concluded that PNV can induce central as well as peripheral haemodynamic effects. The central component seems to be mediated by the activation of cardiovascular centres which in turn lead to an increase in the sympathetic outflow to the periphery, whereas the peripheral component can be accounted for either by direct activation of the vascular alpha1-adrenergic receptors or by catecholamine release from the sympathetic nerve endings. PMID- 10695970 TI - Acid-base balance following Tityus serrulatus scorpion envenoming in anaesthetized rats. AB - In the present work the pH and arterial blood gases were measured in fasted and fed male albino rats, weighing 297 +/- 13 g, anaesthetized with urethane (1.4 g/kg, i.p.) before and after injection of T1 fraction from Titys serrulatus scorpion venom, during 60 min. Arterial blood samples were collected at 0, 5, 15, 30 and 60 min for pH, pCO2, pO2, bicarbonate and base-excess analysis. The data showed that the scorpion toxin induced a continuous drop in the blood pH along the time. Hypercapnia and hypoxemia peaking at 30 min and followed by a recovery towards normal values at 60 min were also observed. A pronounced decrease in the blood bicarbonate levels at 60 min and negative base-excess values along with time were evident at 60 min. The comparisons between fasted and fed animals have shown that in the last group the effects of scorpion toxin on the arterial blood gases were less pronounced. We conclude that T1 fraction of Tityus serrulatus scorpion venom induces in anaesthetized rats an acute respiratory acidosis followed by metabolic acidosis. PMID- 10695971 TI - A comparison of different methods to assess the hemorrhagic activity of Bothrops venoms. AB - The hemorrhagic activity of Bothrops (B.) alternatus, B. ammodytoides, B. jararaca, B. jararacussu, B. moojeni and B. neuwiedii venoms from specimens captured in Argentina was assayed after i.d. injection to mice. The hemorrhagic haloes produced by each venom had different color intensities, although no significant differences were observed by measurement of the average diameters or the weight of the excised hemorrhagic haloes. Conversely, important differences were found by measuring the amount of hemoglobin extracted from excised hemorrhagic haloes of similar size produced by different venoms. The relationship between the amount of hemoglobin extracted and the weight of the excised hemorrhagic haloes was linear, with a slope (hemoglobin released per gram of hemorrhagic halo) characteristic for each venom, and proportional to the potency. On this basis, the activity of B. alternatus, B. ammodytoides and B. jararaca is similar, about 1.5 times higher than that of B. jararacussu and B. moojeni venoms and threefold higher than that of B. neuwiedii venom. Thus, measurement of the of hemoglobin released provides additional information in comparative studies, and may be used to assess the antihemorrhagic potency of antivenoms. PMID- 10695972 TI - Inhibition of the hyperalgesic activity of Bothrops jararaca venom by an antibothropic fraction isolated from opossum (Didelphis marsupialis) serum. AB - The antibothropic fraction (ABF) already isolated from Didelphis marsupialis serum, inhibits the haemorrhagic, oedematogenic, myonecrotic and lethal activities of Bothrops jararaca venom (Bjv). The aim of this work was to verify the capability of ABF to inhibit the hyperalgesic activity of Bjv. Intraplantar injection of Bjv induced hyperalgesia in a time- and dose-dependent manner and ABF administered in situ concomitantly with Bjv or i.v. 30 min before venom injection reduced the induced hyperalgesia. This same effect was observed when ABF was intravenously injected at 5 and 15 min after Bjv. Our results show that ABF inhibits also the hyperalgesia induced by Bjv. PMID- 10695974 TI - Bibliography of toxinology. PMID- 10695973 TI - Immunization with liposome-encapsulated Bothrops jararaca venom. AB - The venom of Bothrops jararaca (BjV) snake was encapsulated into liposomes. The toxicity and ability of the resultant liposomes to protect mice were evaluated. No acute toxicity was found in mice, when liposomes were subcutaneously injected whereas the same dose of venom emulsified in Freund's adjuvant caused the mice to die. Immunization with the venom containing liposomes and associated immunostimulants, protected 50% of mice after challenge with BjV (4 x LD50). The hemorrhagic activity induced by BjV was reduced after immunization with liposomes associated with immunostimulants, similarly to immunization with BjV emulsified in Freund's adjuvant. However, mice immunized with BjV:Freund's were better protected against the lethal effects of the venom. PMID- 10695975 TI - The illusion of paradox: commentary on Albrecht, G.L. and Devlieger, P.J. (1998). The disability paradox: high quality of life against all odds. Social Science & Medicine 48, 977-988. PMID- 10695976 TI - Trends in and determinants of mortality in the elderly population of Matlab, Bangladesh. AB - Longitudinal data collected from the Demographic Surveillance System (DSS) in Matlab, a rural area in Bangladesh, are used for determining trends in and determinants of mortality of the elderly population (60 yr and over) in 1974 1996. The old-age mortality rate is high in Matlab, 1.2 times that of Sri Lankan and 1.5 times that of the Swedish elderly population in a comparable period. Mortality among the elderly population declined in 1974 1982, but much less so in 1982-1996. Proportional hazards models were used for examining determinants of mortality in a sample of about 10,000 elderly persons. This multivariate analysis used information on several social and economic variables derived from the 1982 census and mortality data of this population which was followed prospectively in 1982-1992. Marital status was the single most important determinant: widows and widowers had 1.5 to 2 times higher risk of death compared to couples where both husbands and wives were alive. Social support in old age by children also plays a role, especially for women: women living with at least one son or daughter had 18% lower mortality than women living in a household without sons or daughters. Socioeconomic factors are also important. Those who had at least some education or were relatively affluent had lower mortality than those with no education or who were less affluent. PMID- 10695977 TI - Economic development and women's blood pressure: field evidence from rural Mashonaland, Zimbabwe. AB - A survey of 515 non-pregnant women at 12 geographically chosen research sites in rural Mashonaland shows significant differences in mean blood pressure, controlled by age cohorts. Three levels of economic development are identified: (1) the traditional economy on communal lands, with lowest blood pressure, (2) the wage economy in areas of large-scale commercial agriculture, with elevated blood pressure and (3) the wage economy in mining areas, with the highest elevation of blood pressure. The area is dominated by the primate city, Harare, up to distances of 300 km and beyond, from which forces of change and modernization emanate. It is seen that potassium, sodium and the sodium potassium ratio, are distance-related to Harare and that women's blood pressures tend to follow suit. The rise of body sodium in young persons at risk, often accompanied by declining potassium intake and other changes of modernization, suggest that more attention should be focused on rural areas in Africa, now in the throes of economic change. PMID- 10695978 TI - Thinking differently about thinking positive: a discursive approach to cancer patients' talk. AB - There is an extensive social science and psycho-oncology literature on coping with cancer which claims that "thinking positive" is correlated with--and, by extension, causally implicated in--individuals' morbidity and mortality rates, and their overall level of mental health. Drawing on our own data, in which groups of women with breast cancer talk about "thinking positive", this paper interrogates the basis of such claims from a discursive perspective, by challenging the data analyses upon which they are based. We show that previous literature overwhelmingly relies on self-report data, which are taken as offering more or less accurate depictions of speakers' psychological states (i.e. their mental adjustment or coping style). A discursive approach, by contrast, explores talk as a form of action designed for its local interactional context, and pays detailed attention to what statements about "thinking positive" actually mean for speakers in the contexts in which they occur. We show that "thinking positive" functions not as an accurate report of a internal cognitive state, but rather as a conversational idiom, characterised by vagueness and generality, and summarising a socially normative moral requirement; we also show that even those breast cancer patients who report "thinking positive" can also actively resist its moral prescriptions. Finally, we sketch out the implications of our analysis for analyses of cancer patients' talk more generally and for future research on coping with cancer. PMID- 10695979 TI - The effect of patient race and socio-economic status on physicians' perceptions of patients. AB - Despite its potential influence on quality of care, there has been little research on the way physicians perceptions of and beliefs about patients are affected by patient race or socio-economic status. The lack of research in this area creates a critical gap in our understanding of how patients' demographic characteristics influence encounter characteristics, diagnoses, treatment recommendations, and outcomes. This study uses survey data to examine the degree to which patient race and socio-economic status affected physicians' perceptions of patients during a post-angiogram encounter. A total of 842 patient encounters were sampled, out of which 193 physicians provided data on 618 (73%) of the encounters sampled. The results of analyses of the effect of patient race and SES on physician perceptions of and attitude towards patients, controlling for patient age, sex, race, frailty/sickness, depression, mastery, social assertiveness and physician characteristics, are presented. These results supported the hypothesis that physicians' perceptions of patients were influenced by patients' socio-demographic characteristics. Physicians tended to perceive African-Americans and members of low and middle SES groups more negatively on a number of dimensions than they did Whites and upper SES patients. Patient race was associated with physicians' assessment of patient intelligence, feelings of affiliation toward the patient, and beliefs about patient's likelihood of risk behavior and adherence with medical advice; patient SES was associated with physicians' perceptions of patients' personality, abilities, behavioral tendencies and role demands. Implications are discussed in terms of further studies and potential interventions. PMID- 10695980 TI - Doctor-patient communication about drugs: the evidence for shared decision making. AB - The traditional paternalistic model of medical decision-making, in which doctors make decisions on behalf of their patients, has increasingly come to be seen as outdated. Moreover, the role of the patient in the consultation has been emphasised, notably through the adoption of 'patient-centred' strategies. Models that promote patients' active involvement in the decision-making process about treatment have been developed. We examine one particular model of shared decision making [Charles, C., Gafni, A., Whelan, T, 1997. Shared decision-making in the medical encounter: what does it mean? (or it takes at least two to tango). Social Science & Medicine 44, 681-692.]. The model has four main characteristics. These are that (1) both the patient and the doctor are involved, (2) both parties share information, (3) both parties take steps to build a consensus about the preferred treatment and (4) an agreement is reached on the treatment to implement. Focusing on the first two of the four characteristics of the model, we use the findings from a study of 62 consultations, together with interviews conducted with patients and general practitioners, to consider participation in the consultation in terms of sharing information about, and views of, medicines. We found little evidence that doctors and patients both participate in the consultation in this way. As a consequence there was no basis upon which to build a consensus about the preferred treatment and reach an agreement on which treatment to implement. Thus even the first two of the four conditions said to be necessary for shared decision making were not generally present in the consultations we studied. These findings were presented in feedback sessions with participating GPs, who identified a number of barriers to shared decision making, as well as expressing an interest in developing strategies to overcome these barriers. PMID- 10695981 TI - Family planning KAP survey in Gaza. AB - This study explores the reproductive attitudes, contraceptive use, demand for family planning and related topics of a representative sample of the female population of reproductive age resident in a Refugee Camp in the Gaza Strip. A cluster sample of 841 resident women of reproductive age (15-49 years) was interviewed in their homes. Univariate and multivariate statistical analyses were performed using BMDP software. 98% of the interviewees favour family planning and 88% plan to use a contraceptive in the future. However, 52% of the women at risk do not use any contraception because of their husband's opposition, fear of side effects or lack of knowledge. The risk of having seven or more children is positively associated with a woman's low educational level and husband's desire for more than seven children. Despite favourable attitudes regarding family planning, there is ignorance and the prevalence of contraception use is low. There is a gap between fertility preference and achievement. PMID- 10695982 TI - A council of elders: creating a multi-voiced dialogue in a community of care. AB - In an era of 'medical care delivery systems', there is an increasing need for the patient's voice to be heard, for it to be invited, listened to, and taken seriously. This challenge is particularly evident in geriatrics education, a domain of clinical training in which educators and clinicians alike must struggle to overcome adverse attitudes towards the elderly ('ageism'). In this paper we introduce a 'Council of Elders' as an educational innovation in which we invited community elders to function as our 'Senior Faculty', to whom medical residents present their challenging and heartfelt dilemmas in caring for elder patients. In the conversations that ensue, the elders come to function not simply as teachers, but collaborators in a process in which doctors, researchers, and elders together create a community of resources, capable of identifying novel ways to overcome health-related difficulties which might not have been apparent to either group separately. Using the first meeting of the Council as an exemplar, we describe and discuss the special nature of such meetings and also the special preparations required to build a dialogic relationship between participants from very different worlds--different generations, different cultures (including the professional culture and the world of lived experience). Meetings with the council have become a required part of the primary care residency program--a very different kind of 'challenging case conference' in which moral dilemmas can be presented, discussed and reflected upon. It is not so much that elders give good advice in their responses--although they often do--as that they provide life world and value orientation as young residents gain a better sense of the elder's experience and what matters most to them. This project has been particularly worthwhile in addressing the problem of ageism--a way to render visible stereotypes and adverse physician values, with implications for decision-making with the patient, not for the patient. PMID- 10695983 TI - Migration and geographical inequalities in health in Britain. AB - This paper explores the role of migration in creating geographical inequalities in mortality at the district level in Britain for the British Household Panel Study sample--a representative sample of 10264 British residents born after 1890 and enumerated in 1991. Analysis of the mortality rates of migrants showed that male migration accounts for nearly all the differences in mortality rates between districts. The BHPS was then utilised to look at the lifetime socio-economic characteristics of these migrants and to compare men and women. It was found that the health of both men and women moving from high mortality districts to low mortality districts could be explained by advantage over their lifetimes. The small proportion of men and women moving from low mortality districts to high mortality districts represent a very mixed group and their contribution, whilst small, is intriguing, as is the very different mortality rates of men and women in this group. PMID- 10695984 TI - A village treatment center for malaria: community response in Sri Lanka. AB - Early diagnosis and treatment of malaria cases is one of the basic elements of the current global malaria control strategy. In order to provide this service to people in rural areas there is a need for new cost-effective approaches. To ensure that such new approaches are acceptable to the target communities, it is important to know the rationale for people's malaria treatment-seeking behavior. The present study provides insights into the reasons for people's preferences for different types of healthcare facilities and describes variation of these preferences within a rural community in Sri Lanka. The study reports on the experiences with the establishment of a village health facility and its effect on the treatment-seeking behavior of the population. After the introduction of the village treatment center it quickly took over the role of main provider for diagnosis and treatment of malaria from the government facilities. The treatment center did not improve the response time in seeking treatment for young children, but the delay for adults was reduced by 1-2 days. Mothers with small children often preferred the government facilities since they wanted a more qualified opinion than available from the locally recruited staff of the village treatment center. The treatment center significantly reduced the stress and discomfort experienced by the elderly and handicapped segment of the community. The study indicated that the effective catchment area of a village treatment center will be influenced by the degree of initial support from key individuals in the communities, the selection procedure and training of assistants, and the history of the relationships between different villages to be served by the center. The government health services and communities across the dry zone of Sri Lanka could benefit substantially from the establishment of more village treatment centers. To ensure the long-term sustainability of these type of facilities it is necessary to assess the feasibility of charging a user fee and establishing multi purpose clinics. Government policies and administrative procedures will need to be adjusted to make the successful operation of village treatment centers possible. PMID- 10695985 TI - Social factors influencing the acquisition of antibiotics without prescription in Kerala State, south India. AB - We investigated the magnitude of self-medication with antibiotics in a peri-urban area of Southern Kerala State, India and factors influencing this practice. First, a random sample of 400 households was surveyed in one primary health centre area near Trivandrum. We found 69.3% (95% CI = 64.8-73.8) of households had at least one person using a pharmaceutical product during the two-week recall period; antibiotics formed almost 11% of the medicines consumed. Next, pharmacy based interview and observation data were collected from 405 antibiotic purchasers sampled from 11 out of the 12 private pharmacies in the area. Seventy three of these 405 customers purchased antibiotics without a prescription (18%; 95% CI = 14.3-21.7). By combining the household survey and pharmacy observations, we estimate that almost half of 1% (0.41%; 95% CI = 0.24-1.16) of the population, or four people per 1000, is engaged in self-medication using antibiotics in Kerala in any two-week period. Our data show that people least likely to follow this practice are from higher income families, having more education and higher status occupations and receiving the benefits of medical insurance. Conversely, logistic regression analysis indicated that risk of buying antibiotics without a script was associated with education at secondary level or below, the perception that it is expensive to consult a doctor and low satisfaction with medical practitioners. Keralites' self-medication patterns are interpreted broadly using social, cultural, historical and economic perspectives. Solutions to the problem of antibiotic misuse are suggested, proceeding on several fronts: among practitioners, suppliers and marketeers of medicines, and among the population of pharmaceutical consumers themselves. PMID- 10695986 TI - Acute local inflammation potentiates tumor growth in mice. AB - Chronic inflammation in humans has been implicated in the pathogenesis of several types of cancer. In animals, experimentally-induced tumor growth was found to be enhanced at sites of injury. However, a direct demonstration in vivo that an inflammatory agent applied locally at the tumor site can promote a switch into a highly proliferative state of tumor growth, has not yet been documented. The present work was designed to test, in a syngeneic primary tumor model in mice, whether a commonly used inflammatory agent, carrageenan, could cause acceleration of tumor growth and to investigate the cellular mechanisms mediating such a process. Local injection of carrageenan into a tissue site containing tumor cells produced an accelerated rate of tumor growth at that site which was characterized by a decreased percentage of apoptotic cells and an increased proportion of cells at the S and G2/M phases of the cell cycle. The pro-tumorigenic effect of carrageenan is dose-dependent and can be exerted at any time throughout the course of the tumor growth. Furthermore, the effect is prostaglandin-mediated since the cyclooxygenase inhibitor indomethacin totally abrogated it. Experiments with tumors cells in culture have shown that carrageenan actually inhibits cell proliferation as well as increases apoptosis. Thus, the tumor promoting effects of carrageenan in vivo appear to arise not from a direct effect on the tumor cells per se but rather through induction of host-dependent humoral/cellular responses that generate increased levels of prostanoids and pro-inflammatory cytokines that accelerate tumor growth. These data demonstrate for the first time that an acute, local inflammatory stimuli can induce accelerated tumor growth at the affected site and provide further support for a mechanism-based, anti tumorigenic action of anti-inflammatory drugs. PMID- 10695987 TI - Effect of constant light on DMBA mammary tumorigenesis in rats. AB - A study of light, and mammary tumorigenesis was conducted in rats. One-hundred female Sprague-Dawley rats were divided by weight into two groups. One group was exposed to constant light (LL) from 26 days of age, and the second group was exposed to 8 h light and 16 h dark per day (LD). Both groups received an 8 mg dose of a chemical carcinogen, dimethylben-zanthracene (DMBA) at 52 days of age. At 13 weeks post-DMBA, there were significantly fewer mammary tumors in the LL group compared with the LD group. Constant light was clearly demonstrated to have a profound effect on mammary tissue development. Although virgin, the majority of the LL rats (29/50) had gross evidence of lactation at 141 days of age. None of the LD rats (0/50) showed evidence of milk production. These results suggest that constant light not only substantially accelerated mammary gland development, but pushed development of the tissue past the stage normally observed in virgin animals (to the lactation stage). PMID- 10695988 TI - Conserved expression of hepatocyte growth factor activator inhibitor type 2/placental bikunin in human colorectal carcinomas. AB - Hepatocyte growth factor activator inhibitor type 2 (HAI-2) was recently identified as a potent inhibitor of hepatocyte growth factor activator. It was also independently reported as placental bikunin (PB) and as a protein over expressed in pancreatic cancer. The expression of HAI-2/PB was analyzed in human normal colon mucosa, adenomas, and carcinomas. HAI-2/PB mRNA was consistently expressed in the colorectal mucosa. The expression was conserved in the neoplastic colorectal mucosa, and no relationship was found between HAI-2/PB mRNA levels and tumor stages. Moreover, 13 out of 14 colorectal carcinoma cell lines expressed HAI-2/PB mRNA. Immunohistochemically, HAI-2/PB proteins were predominantly stained beneath the apical surface of normal enterocytes. In tumor tissues, rather disarranged intracytoplasmic granular staining was observed. The HAI-2/PB immunoreactivity was well conserved in the colonic adenoma-carcinoma sequence, and this protein may have important unknown function in the intestinal mucosa. PMID- 10695989 TI - Carbonyl reduction of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) by cytosolic enzymes in human liver and lung. AB - The tobacco specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) is a potent pulmonary carcinogen, independent of the route and type of administration. There are competing metabolic activation and detoxification pathways. NNK is activated by alpha-hydroxylation at either the methyl or methylene carbonyl adjacent to the N-nitroso group to yield intermediates that methylate and pyridyloxobutylate DNA. Detoxification of NNK in humans usually occurs via carbonyl reduction to its hydroxy product NNAL, which undergoes glucuronosylation and final excretion. In vitro studies on NNK metabolism have usually been performed with tissue homogenates, microsomal fractions and/or purified microsomal enzymes, but cytosolic metabolism of NNK has been ignored until today. The results of this study demonstrate that cytosolic fractions of human liver and lung also participate in NNK metabolism. We provide evidence that a substantial degree of NNK carbonyl reduction occurs by cytosolic enzymes and that these enzymes may contribute to NNK detoxification in human liver and lung. The relative contribution of cytosolic vs. microsomal NNK carbonyl reduction is nearly identical in liver, whereas it is more than 3-fold higher in lung microsomes compared to lung cytosol. The inhibition profile suggested that mainly carbonyl reductase (EC 1.1.1.184) was active in cytosol of both organs. The expression of carbonyl reductase mRNA in liver and lung was proven by reverse transcription-(RT)-PCR. In conclusion, the results of this study provide the first data on cytosolic enzymes participating in NNK detoxification in human liver and lung. PMID- 10695990 TI - Increased expression of cyclooxygenase-2 protein in rat lung tumors induced by N nitrosobis(2-hydroxypropyl)amine. AB - Expression of cyclooxygenase (COX)-2 protein in preneoplastic and neoplastic lung lesions induced by the administration of 2000 ppm of N-nitrosobis(2 hydroxypropyl)amine (BHP) in the drinking water to Wistar male rats, was examined immunohistochemically. The majority of alveolar/bronchiolar adenomas (ADs) and all adenocarcinomas (ADCs) examined, stained positive or strongly positive for COX-2. In contrast, only a minority of alveolar/bronchiolar hyperplasias demonstrated immunoreactivity and half of the squamous cell carcinomas examined, were only weakly positive. Western blotting analysis also revealed expression of COX-2 protein in the resected ADs and ADCs. These results clearly indicate up regulated expression of COX-2 in lung neoplastic lesions, particularly ADs and ADCs, induced by BHP in rats. PMID- 10695991 TI - Thymidine phosphorylase expression in tumor stroma of uterine cervical carcinomas: histological features and microvessel density. AB - Immunohistochemical staining was performed on 91 cases of uterine neoplasm in order to determine if expression of platelet-derived endothelial cell growth factor/thymidine phosphorylase (TP) correlates with tumor microvessel density (MVD) and histological parameters of uterine carcinomas in tumor cells and in tumor stroma. The sample group consisted of 72 primary invasive squamous cell carcinomas of the cervix (ISC) and 19 cervical intraepithelial neoplasms (CIN) of the uterus. In ISC of the cervix, TP expression in tumor stroma showed a significant correlation with a non-keratinizing histological subtype (P < 0.001) and with an infiltrating invasive pattern (P < 0.001). However, in tumor cells the TP expression showed a higher correlation with a keratinizing histological subtypes (P = 0.009). MVD was significantly higher (P = 0.002) in tumors showing high TP expression in stroma than in tumors with low expression. These findings suggest that the TP expression in stromal cells, rather than in tumor cells, may play a role in promoting microvessel growth in cervical squamous cell carcinoma, and angiogenesis may also have an association with tumor cell invasion. PMID- 10695992 TI - Absence of p51 mutation in human hepatocellular carcinoma. AB - The p51 gene encodes a protein with significant homology to p53. To investigate the involvement of the p51 gene in human hepatocarcinogenesis, mutation analysis of the p51 gene was performed in 54 cases of hepatocellular carcinoma (HCC). No mutations causing amino acid substitutions or frameshifts were found by polymerase chain reaction-single strand conformational polymorphism (PCR-SSCP) analysis of the entire coding region. The result indicated that mutation of the p51 gene does not play a major role in the development of HCC in Japanese patients. Further studies on p51 expression and its functions, including the interaction with p53, are necessary to elucidate the role of the p51 gene in human hepatocarcinogenesis. PMID- 10695993 TI - The significance of thymidine phosphorylase activity in hepatocellular carcinoma and chronic diseased livers: a special reference to liver fibrosis and multicentric tumor occurrence. AB - The role of thymidine phosphorylase (TP), an angiogenic factor, in hepatocellular carcinoma (HCC) remains unclear. The aim of this study was to clarify the significance of TP in HCC. Thirty-seven patients with HCC, who underwent hepatectomy, were included. The TP activity in both cancerous and non-cancerous parts of livers were measured by an enzyme-linked immunosorbent assay. Another 11 patients without HCC were used to evaluate the TP activity in the non-cancerous parts of livers. Both the cancerous and non-cancerous TP activities were clinico pathologically investigated with special reference to the multicentric occurrence of HCCs and the degree of liver fibrosis; consisting of normal, fibrosis and cirrhosis. The TP activity in the cancerous part was 94.6 +/- 70.2 U/mg protein, while that in non-cancerous parts of the liver was 80.9 +/- 48.8 U/mg protein. No significant difference was observed. The TP activity in the cancerous part did not correlate with any clinicopathological variables, such as tumor differentiation, portal vein invasion, intrahepatic metastases and prognosis. However, the TP activity in the non-cancerous parts of the liver correlated with the degree of fibrosis (normal/fibrosis/cirrhosis = 34:74:90 U/ mg protein, respectively). Furthermore, regarding the correlation between TP activity in the non-cancerous parts and the simultaneously multicentric occurrence of HCC, the TP activity in the multicentric group (n = 8; 121 U/mg protein) was significantly higher than that in the non-multicentric group (n = 29; 70 U/mg protein). The TP activity in the non-cancerous parts increased in proportion to the degree of liver fibrosis. Furthermore, it is suggested that the higher TP activity in the non-cancerous part is related to the multicentric occurrence of HCCs. PMID- 10695994 TI - Newly recognized cytotoxic effect of conjugated trienoic fatty acids on cultured human tumor cells. AB - We investigated the cytotoxic effect of conjugated trienoic fatty acids on various human tumor cell lines: DLD- 1, colorectal; HepG2, hepatoma; A549, lung; MCF-7, breast; and MKN-7, stomach. Conjugated linoleic acid (CLA) and conjugated linolenic acid were prepared from linoleic acid (18:2, n-6) and alpha-linolenic acid (18:3, n-3), respectively, by treatment with 6.6% or 21% potassium hydroxide. Spectrophotometric readings at 235 nm for the conjugated diene formation, and at 268 nm for the conjugated triene, were confirmed for the respective conjugated fatty acids. In addition, tung oil (Aleurites fordii) fatty acids consisting principally of a conjugated triene (eleostearic acid, approximately 80% of total fatty acids) were prepared using an alkaline saponification procedure. All tumor cells were incubated for 24 h with 5-100 microM of the conjugated fatty acids, and MTT dye reduction was measured to verify the cell viability. Among the conjugated fatty acids examined, conjugated linolenic acid and tung oil fatty acids exhibited the most intense cytotoxic effects on DLD-1, HepG2, A549, MCF-7 and MKN-7 cells, while CLA was not cytotoxic to the tumor cells. These results demonstrate that conjugated trienoic fatty acids are more cytotoxic to human tumor cells than the conjugated dienoic fatty acid, CLA. PMID- 10695995 TI - p21 expression is a prognostic factor in patients with p53-negative gastric cancer. AB - The expression of p21 and p53 proteins was analyzed by immunohistochemistry in 256 patients with advanced gastric cancer. The results showed that strong, weak and negative expression of p21 were detected in 22.2 (57/256), 68.0 (174/256) and 9.8% (25/256) of the patients, respectively. p53 expression was found in 28.9% (74/256). The expression of p21 was not associated with clinicopathological features. In p53 negative tumors, p21 expression was associated with the survival of patients who underwent curative operations (P = 0.007). The 5-year survival rates were 20.1, 36.6 and 59.8% in patients with p21-negative, -weakly positive and -strongly positive tumors, respectively. In contrast, in p53-positive tumors, prognosis did not differ in spite of p21 expression. Multivariate analysis showed that p21 expression was an independent factor in patients with p53-negative tumors. These results indicate that examination of p21 expression in p53 negative tumors will be useful for estimating the prognosis of patients with advanced gastric cancer. PMID- 10695996 TI - Cytotoxic and antitumor effect of fibrinogen-methotrexate conjugate. AB - In this paper we describe the chemical procedure of fibrinogen-methotrexate (F MTX) conjugate preparation and its in vitro and in vivo antitumor activity. F-MTX conjugates were synthesized in reaction of fibrinogen with MTX N hydroxysuccynimide ester. The conjugates were not cross-linked and were soluble in water. The results of the in vitro and in vivo studies have shown: (1) a lower in vitro cytotoxicity of the F-MTX conjugate as compared with MTX alone; (2) a significantly higher in vivo antitumor activity of the F-MTX conjugate in mice with P388 leukemia as compared with MTX alone; (3) a significantly increased in vivo lethal toxicity of F-MTX as compared with MTX. The results suggest the therapeutic utility of the fibrinogen-methotrexate conjugate and the usefulness of fibrinogen as a chemotherapeutic drug carrier. However, a new effort in the preparation of F-MTX conjugate should be made to decrease its in vivo toxicity. PMID- 10695997 TI - p53 protein detection by the western blotting technique in normal and neoplastic specimens of human endometrium. AB - The aim of the study was to investigate p53 protein expression by the Western blotting technique (estimated by integrated optical density - IOD) in normal (n = 13) and neoplastic (n = 40) human endometrial tissues as well as in a case of uterine carcinosarcoma and in a specimen of the botryoid sarcoma of the uterine cervix. p53 protein levels were correlated with patients' age as well as with conventionally used clinicopathological features of the endometrial neoplasm. A statistically significant difference was noted in p53 levels in the nuclear, but not in the cytoplasmatic, fraction between the normal endometria and endometrial cancer tissues (P < 0.0001). In the neoplastic endometria, nuclear p53 protein expression was higher than in cytoplasmatic fraction, and the difference was significant (P < 0.05). Higher nuclear p53 protein levels correlated with advanced histological grading of endometrioid endometrial carcinomas, but no relationship was noted between p53 protein expression and patients' age, clinical stage, histological type or depth of myometrial invasion. A case of uterine carcinosarcoma and a specimen of a botryoid sarcoma of the uterine cervix expressed nuclear p53 oncoprotein (57 IOD and 89 IOD, respectively). In conclusion, we found a statistically higher nuclear p53 levels in malignant as compared to normal human endometrial specimens by the Western blotting technique. Although there were no significant differences between p53 expression and clinicopathological features of the neoplasm (except poor histological grading), further studies are necessary to evaluate the influence of p53 nuclear/cytoplasmatic levels on the clinical outcome of Polish patients suffering from endometrial cancer. PMID- 10695998 TI - IL-6 enhanced the retinoic acid-induced differentiation of human acute promyelocytic leukemia cells. AB - It has been shown that retinoic acid (RA) induced the expression of interleukin-6 (IL-6) in human acute promyelocytic leukemia HL-60 cells. In the present study, we examined the ability of RA to induce the expression of gp130, the signal transducing receptor component for IL-6, in HL-60 and a RA-supersensitive cell line HL-60/S4. We found that RA induced the expression of gp130, at both the mRNA and protein levels, in HL-60 and HL-60/S4 cells. Interestingly, the induction of gp 130 expression observed in the RA-supersensitive HL-60/S4 cells was much more pronounced than that observed in HL-60 cells. Furthermore, activation of the RA induced gp130 by exogenous IL-6 potentiated the differentiating effects of RA. The synergistic effects observed for IL-6 and RA was also much stronger in HL 60/S4 cells than in HL-60 cells. Our findings suggest that the differentiating effects of RA may partially be mediated by the up-regulation of IL-6/gp130 signaling in HL-60 and HL-60/S4 cells. PMID- 10695999 TI - Inhibitory effect of synthetic interferon inductor Cycloferone on 1,2 dimethylhydrazine-induced colon carcinogenesis in rats. AB - The effect of a synthetic interferon inductor Cycloferone on colon carcinogenesis was firstly studied in rats. Seventy-five 2-month-old outbred female LIO rats were subdivided into three groups and were weekly exposed to 15 s.c. injections of 1,2-dimethylhydrazine (DMH) at a single dose of 7 mg/kg body wt. From the day of the fist injection of DMH rats from group 2 were given weekly i.p. injections of Cycloferone (62.5 mg/kg) until the end of the experiment. DMH-treated rats (group 3) were exposed to weekly i.p. injections of Cycloferone (62.5 mg/kg) starting in the week after the last injection of the carcinogen. Rats from group 1 were exposed to DMH and treated weekly with 0.2 ml i.p. of normal saline. Additional groups of rats were treated weekly with Cycloferone (62.5 mg/kg) or with 0.2 ml of saline. The experiment was ended 6 months after the first injection of DMH. In DMH-treated rats (groups 1, 2 and 3) colon adenocarcinomas developed in 87, 61 and 59%, respectively. The number of colon tumors per tumor bearing rat was 2.5, 1.9 and 1.3 in groups 1, 2 and 3, respectively. Treatment with Cycloferone significantly inhibits carcinogenesis in ascending and descending colon. The incidence of tumors of the rectum was decreased in the group 2 as compared with the group 1. There were no cases of tumors of rectum in rats from group 3. The treatment with Cycloferone alone as well as with normal saline failed to induce any tumors in rats. Thus, our results demonstrated inhibitory effect of Cycloferone on colon carcinogenesis induced by DMH in rats. PMID- 10696000 TI - Diskography: science and the ad hoc hypothesis. PMID- 10696001 TI - Cervical diskography: analysis of provoked responses at C2-C3, C3-C4, and C4-C5. PMID- 10696002 TI - Pontine reversible edema: a newly recognized imaging variant of hypertensive encephalopathy? PMID- 10696003 TI - Radiographic screening for orbital foreign bodies prior to MR imaging: is it worth it? PMID- 10696004 TI - Short-term arteriographic and clinical outcome after cerebral angioplasty and stenting for intracranial vertebrobasilar and carotid atherosclerotic occlusive disease. AB - BACKGROUND AND PURPOSE: The safe performance of percutaneous transluminal cerebral angioplasty for intracranial atherosclerotic lesions requires that the risk of complications, such as acute occlusion or symptomatic dissection, and restenosis be reduced. Our purpose was to assess the effectiveness, safety, and short-term arteriographic and clinical outcome of cerebral angioplasty and stenting (CAS) for intracranial vertebrobasilar and distal internal carotid atherosclerotic occlusive lesions. METHODS: Between March 1998 and November 1998, 10 patients with 12 intracranial atherosclerotic lesions of the vertebrobasilar artery and the distal internal carotid artery underwent treatment with flexible balloon-expandable coronary stents. RESULTS: Although in two of the 10 patients CAS was not successful because of the inability to access the site of arterial stenosis, 10 lesions in eight patients were successfully dilated with stents. No complications occurred during or after the procedure and no neurologic ischemic events or restenoses occurred during the follow-up period. CONCLUSION: CAS appears to be a safe and effective means for treating intracranial atherosclerotic occlusive disease, yielding a favorable arteriographic and clinical outcome. PMID- 10696005 TI - Efficacy of trisacryl gelatin microspheres versus polyvinyl alcohol particles in the preoperative embolization of meningiomas. AB - BACKGROUND AND PURPOSE: Trisacryl gelatin microspheres are a new, commercially available nonabsorbable embolic agent. The purpose of this study was to evaluate their efficacy in the preoperative embolization of meningiomas as compared with polyvinyl alcohol (PVA) particles of various sizes. METHODS: In 30 consecutive patients, trisacryl gelatin microspheres (150-300 microm) were used for the preoperative superselective embolization of meningiomas (group 1). Thirty other consecutive patients had embolization with PVA particles of 45 to 150 microm (n = 15, group 2) and of 150 to 250 microm (n = 15, group 3). Extent of devascularization, intraoperative blood loss, blood transfusion, and hemostasis at the time of surgery were recorded for every patient. The inflammatory reaction, the extent of necrotic areas, and the most distal intravascular location of the embolic agent (arterial, arteriolar, precapillary, capillary) were recorded. RESULTS: There was no significant difference in the extent of angiographic devascularization among the groups. Intraoperative blood loss differed significantly between groups 1 and 2 and groups 1 and 3, but not between groups 2 and 3. The trisacryl gelatin microspheres were located more distally in tumor vessels than were the PVA particles of either size. The extent of intratumoral necrosis was not significantly different between the two embolic agents. In all groups there was a mild inflammatory tissue reaction in the vicinity of the embolic agent. CONCLUSION: Trisacryl gelatin microspheres may be effective in the preoperative embolization of meningiomas, producing significantly less blood loss at surgery than seen with PVA particles of either size, possibly because of the significantly more distal vascular penetration of the microspheres. PMID- 10696006 TI - The retrograde approach: a consideration for the endovascular treatment of aneurysms. AB - BACKGROUND AND PURPOSE: The traditional endovascular approach to a cerebral aneurysm is anterograde, with the embolization and balloon protection catheters introduced via the parent vessel. Unfortunately, this approach may be restrictive, because these catheters cannot always be navigated at an optimal angle into the arterial branch that needs balloon protection or the part of the aneurysm that needs coiling. The purpose of this study was to determine the efficacy of a retrograde approach. METHODS: Twelve patients, seven women and five men, 28 to 65 years old (mean age, 45 years), were treated via the retrograde approach between March 1998 and February 1999. Three patients were treated for acutely ruptured aneurysms following subarachnoid hemorrhage. The rest had asymptomatic, unruptured aneurysms. RESULTS: We were able to accomplish endovascular treatment in 10 cases. In the other two, the attempted retrograde route of access could not be achieved. The treatment afforded complete embolization in nine of the 10 patients. Symptomatic distal clot embolization occurred in one patient who had some residual, albeit improving, deficits at discharge. No other patients worsened with the treatment. There were two intraprocedural aneurysmal ruptures. None of the aneurysms restudied within 6 months (eight of 12) showed evidence of recanalization. CONCLUSION: Our results indicate that it is possible to safely and effectively access a cerebral aneurysm via a retrograde approach. We believe that the anatomic benefits afforded by this technique outweigh the potential risks associated with the catheterization of another major cerebral arterial feeder. PMID- 10696007 TI - Cervical diskography: analysis of provoked responses at C2-C3, C3-C4, and C4-C5. AB - BACKGROUND AND PURPOSE: Previous authors have described the locations of provoked responses to cervical diskography from C3-C4 to C6-C7, but we have found no description of the findings at C2-C3. This study was undertaken to analyze the sensations provoked during cervical diskography at C2-C3 and to compare the results with those provoked at C3-C4 and C4-C5. METHODS: The locations of diskographically provoked responses from 40 consecutive patients who had undergone C2-C3, C3-C4, and C4-C5 diskography were analyzed. Only intensely painful (> or = 7/10) and concordant responses were considered. Disk morphology on MR images and diskograms was also compared with the provoked responses. RESULTS: Eighteen subjects described either unilateral (n = 10) or bilateral (usually asymmetric) (n = 8) concordant pain at the craniovertebral junction in response to C2-C3 diskography. Nine subjects described either unilateral (n = 5) or bilateral (n = 4) neck pain during injection. Cephalalgia or head pain was provoked in 19 subjects, seven bilaterally. Four subjects described either unilateral (n = 3) or bilateral (n = 1) trapezius muscle and/or shoulder pain. Preliminary MR studies were not helpful, as most C2-C3 disks either appeared normal or exhibited nonspecific signs of degeneration. All disks exhibited either fissuring or extradiskal leakage of contrast material at diskography, regardless of the response provoked. CONCLUSION: Diskography at C2-C3 and C3-C4 frequently produces pain sensations in the head, craniovertebral junction, and neck. There is no correlation between C2-C3 disk morphology and the diskographically provoked response. PMID- 10696008 TI - Relationship of Schmorl's nodes to vertebral body endplate fractures and acute endplate disk extrusions. AB - BACKGROUND AND PURPOSE: Literature regarding clinical pain syndromes associated with acute, traumatic Schmorl's nodes (SNs) is limited. Our purpose was to determine whether an SN could be related to a previous traumatic event producing either acute SN or a vertebral endplate fracture. METHODS: Two neuroradiologists independently reviewed initial and follow-up MR examinations of 14 patients with a clinical diagnosis of acute, symptomatic thoracolumbar SNs or vertebral body endplate fractures that evolved into SNs to evaluate marrow edema, signal intensity, margin definition, presence of intravertebral extruded disk material, and pattern of contrast enhancement. RESULTS: Edema of the affected vertebral body, adjacent to an endplate without wedging or collapse, was observed on the initial MR images in all cases. The initial MR images of six (43%) of 14 patients exhibited only edema of the marrow immediately adjacent to the endplate without wedging or collapse. The MR images obtained at the time of follow-up showed subsequent formation of a chronic and eventually asymptomatic SN for all six patients. The initial MR images of eight (57%) of the 14 patients showed the typical appearance of acute SNs with marrow edema of the affected vertebra. The contrast-enhanced images of three patients manifested enhancement of the invaginated disk material in three (100%) of three cases and enhancement of the surrounding vertebral body in one case (33%). Six (43%) of 14 patients had acute typical compression fracture of a vertebral body of at least one additional level. CONCLUSION: Most (57%) of the SNs in this series could be traced to episodes of significant, sudden-onset, localized, nonradiating back pain and tenderness for which the MR images showed SNs surrounded by vertebral body marrow edema. The remaining SNs (43%) were not immediately apparent as SNs and manifested only as vertebral body edema representing endplate fracture but did evolve into classical chronic SNs that follow-up imaging revealed. PMID- 10696009 TI - Registration of three-dimensional MR and CT studies of the cervical spine. AB - A three-dimensional image registration technique for CT and MR studies of the cervical spine was evaluated for feasibility and efficacy. Registration by means of external fiducial markers was slightly more accurate than registration by anatomic landmarks. The interrelationships between bony (eg, neural foramina) and soft tissue structures (eg, nerve roots) in the cervical spine were more conspicuous on registered images than on conventional displays. Registration of CT and MR images may be used to examine more precisely the relationships between bony and soft tissue structures of the cervical spine. PMID- 10696010 TI - Morphologic characteristics of subcortical heterotopia: MR imaging study. AB - BACKGROUND AND PURPOSE: Gray matter heterotopia have been divided into three groups based on clinical and imaging characteristics: subependymal, subcortical, and band heterotopia. Nonetheless, subcortical heterotopia can have variable morphologic findings. The purpose of this study was to perform a morphologic analysis of a series of cases of subcortical heterotopia based on MR images, to correlate the morphologic appearance with clinical characteristics, and to speculate about the embryologic implications of our results. METHODS: The MR imaging studies and clinical records of 24 patients with subcortical heterotopia were retrospectively reviewed. The morphologic findings of the heterotopia were recorded along with presence and type of associated malformations. These results were correlated with available data on development and neurologic status. RESULTS: Analysis revealed that, in six cases, the heterotopia were composed exclusively of multiple nodules, in 13, they appeared primarily as curvilinear ribbons of cortex extending into the white matter, and in five, they had deep nodular regions with curvilinear areas more peripherally. All of the curvilinear regions were contiguous with the cerebral cortex in at least two locations. In eight cases, curvilinear heterotopia contained curvilinear areas of flow void that were thought to be blood vessels; in 10, they contained fluid resembling CSF. No difference in developmental or neurologic manifestations was noted among patients with heterotopia of different morphologic appearances. CONCLUSION: Subcortical heterotopia can have nodular or curvilinear morphologic appearances. Although no difference was found in the clinical conditions of the patients with differing morphologic appearances, additional analysis of these patients or studies of animal models of these malformations may further our understanding of normal and abnormal brain development. PMID- 10696011 TI - Antenatal diagnosis of subependymal heterotopia. AB - Subependymal heterotopia consist of gray matter nodules along the lateral ventricular walls and are associated with epilepsy and other cerebral malformations. Some cases have an X-linked inheritance, and early antenatal diagnosis of affected fetuses is important for appropriate management. We present a case of heterotopia diagnosed by sonography and MR imaging at 23 weeks' gestation and discuss the differential diagnosis, reviewing the evolution and imaging appearances of the germinal matrix and its implications for detection of heterotopia. PMID- 10696012 TI - Magnetoencephalography in children with Landau-Kleffner syndrome and acquired epileptic aphasia. AB - BACKGROUND AND PURPOSE: Landau-Kleffner syndrome (LKS) is epileptiform aphasia acquired during childhood and occurring in children with previously normal language development. The epileptiform activity in these children is thought to result in a functional ablation of eloquent speech areas. The purpose of this study was to investigate the usefulness of magnetoencephalography (MEG) for localizing the source of epileptiform activity in these patients. METHODS: Nineteen patients with acquired aphasia and a suspected diagnosis of LKS were referred for MEG evaluation. Patients ranged in age from 4 to 14 years. Fourteen MEG studies were performed on a 74-channel system, four on a 148-channel whole head system, and one on a 37-channel system. RESULTS: Thirteen of the 19 patients had perisylvian MEG spikes. In 10 of the patients, the spikes were bilateral, and in three they were unilateral. Four other patients had non-sylvian spikes, and two patients had no spikes recorded. The results of MR imaging were normal or noncontributory for all 19 patients. CONCLUSIONS: MEG can play a useful role in evaluating children with LKS and acquired epileptiform aphasia, both in diagnosis and in aiding presurgical localization of epileptiform activity when surgery is being considered. PMID- 10696013 TI - Intraosseous hematoma in a newborn with factor VIII deficiency. AB - We present an unusual case of an intraosseous hematoma in a newborn with a known bleeding disorder. This cephalohematoma was diagnosed shortly after birth, was entirely within the bony skull, and was in fact determined to be an intraosseous hematoma. The initial CT scans showed the unusual appearance and location of the lesion; later scans showed a significant amount of remodeling, with resolution of the hematoma. Although the coagulopathic diagnosis was independent of this finding, a bleeding disorder might be considered in other patients with similar CT findings. PMID- 10696015 TI - Second branchial cleft cysts: variability of sonographic appearances in adult cases. AB - BACKGROUND AND PURPOSE: Previous reports have suggested that second branchial cleft cysts (BCCs) appear on sonograms as well-defined, cystic masses with thin walls and posterior enhancement. Previous CT and MR imaging findings, however, have indicated heterogeneity of these masses, and, in our experience, sonography also shows a similar variable appearance. In this communication, we report the cases of 17 patients with second BCCs and document the variability of sonographic patterns. METHODS: The sonograms of 17 adults with second BCCs were reviewed. Only patients with surgical or cytologic evidence of BCCs were included in this study. The features evaluated were the location, internal echogenicity, posterior enhancement, and presence of septa and fistulous tract. RESULTS: Four patterns of second BCCs were identified: anechoic (41%), homogeneously hypoechoic with internal debris (23.5%), pseudosolid (12%), and heterogeneous (23.5%). The majority (70%) showed posterior enhancement. All were situated in their classical location, posterior to the submandibular gland, superficial to the carotid artery and internal jugular vein, and closely related to the medial and anterior margin of the sternomastoid muscle. Fourteen (82%) of the 17 BCCs had imperceptible walls, and all were well defined. For none of the patients was a fistulous tract revealed by sonography; the presence of internal septations was revealed for three patients. CONCLUSION: As previously suggested by CT and MR imaging findings, sonography reinforces that second BCCs in adults are not simple cysts but have a complex sonographic pattern ranging from a typical anechoic to a pseudosolid appearance. PMID- 10696014 TI - Nodal volume reduction after concurrent chemo- and radiotherapy: correlation between initial CT and histopathologic findings. AB - BACKGROUND AND PURPOSE: The role of concurrent chemoradiation for treatment of head and neck squamous cell carcinoma is expanding. We sought to evaluate the CT appearance of diseased and normal cervical lymph nodes before and after concurrent chemoradiation and to correlate lymph node volume reduction as revealed by CT with histopathologic findings of resected nodes. METHODS: Using concurrent chemoradiation, we treated seven patients with locally advanced head and neck squamous cell carcinoma. Our chemotherapeutic regimen consisted of cisplatin (100 mg/m2 body surface area administered on days 1 through 4 and 29 through 32) and 5-fluorouracil (1000 mg/m2 body surface area, administered on days 1 through 4 and 29 through 32). Radiotherapy was administered twice per day on dosing days 1 through 42 to a total dose of 7200 cGy to the primary tumor and 6000 cGy to the involved lymph nodes. Pre- and post-treatment CT scans were used to calculate lymph node volumes for all CT-positive (size criteria or extracapsular spread or both) diseased nodes (n = 19) and one normal node per patient (n = 7). Volume reduction was determined by CT results and correlated with the histopathologic findings of resected nodes. RESULTS: Average volume reduction (+/- standard error of the mean) for the 19 diseased nodes was 91%+/-4% and for the seven normal nodes was 55%+/-21% (P < .02, two-sided t test). Fifteen of 19 of the diseased lymph nodes showed extracapsular spread before treatment and none of 19 after treatment. The histopathologic findings of resected nodes included persistent tumor in one of the 19 diseased lymph nodes. Six of seven patients remained alive and disease-free, with an average follow-up duration of 24 months. CONCLUSION: Nodal volume reduction of greater than 90% was associated with eradication of tumor as assessed by histopathologic analysis of resected nodes. Serial CT scans obtained both before and after concurrent chemoradiation may be useful for predicting which patients will benefit from adjuvant surgical therapy. PMID- 10696016 TI - Riedel's thyroiditis in multifocal fibrosclerosis: CT and MR imaging findings. AB - Riedel's thyroiditis is a rare disorder of unknown etiology and may be seen isolated or as a part of multifocal fibrosclerosis. It is important to distinguish Riedel's thyroiditis from thyroid carcinoma. Reports about imaging features of Riedel's thyroiditis are limited in the radiologic literature. We describe herein CT and MR imaging features of Riedel's thyroiditis in a case of multifocal fibrosclerosis with previously unreported radiologic observations. PMID- 10696017 TI - Congenital absence of the oval window: radiologic diagnosis and associated anomalies. AB - BACKGROUND AND PURPOSE: In most children with conductive hearing loss, acquired otitis media and/or middle ear effusion are ultimately diagnosed. Congenital conductive hearing loss is a rare condition; absence of the oval window is an unusual pathogenesis for this type of hearing impairment and can be associated with an anomalous horizontal facial nerve canal. Our goal was to describe the imaging features of congenital absence of the oval window, to determine the frequency with which anomalous development of the horizontal facial nerve canal occurs, and to review the developmental error responsible for this malformation. METHODS: Nine temporal bones in seven patients (5 to 36 years old) were found to have an inadequately formed oval window on high-resolution CT scans; seven ears showed complete lack of oval window formation, and two showed partial absence of the oval window. Records were reviewed for clinical information, and images were examined for associated anomalies. RESULTS: Six of nine ears with abnormal oval window formation showed malposition of the horizontal facial nerve canal. In each of these, the canal was abnormally low, overlying the expected location of the oval window; three of the canals lacked a visible bony covering. Seven of the nine ears were found to have a dysplastic or absent stapes. CONCLUSION: Congenital absence of the oval window can be diagnosed on CT studies. In the present series, this anomaly was associated with a grossly aberrant horizontal facial nerve canal in six of nine involved ears. Familiarity with the developmental sequence of oval window formation fosters an understanding of these anomalies. Preoperative recognition is important clinically, as a low facial nerve will block surgical access to the oval window and its presence will alter patient management. PMID- 10696018 TI - Reduced size of the cochlear branch of the vestibulocochlear nerve in a child with sensorineural hearing loss. AB - A 12-year-old female patient presented with unilateral sensorineural hearing loss. Distortion-product otoacoustic emission testing failed to reveal any measurable emissions in the affected side. MR imaging did not reveal labyrinthine malformation. Three-dimensional Fourier transformation-constructive interference in steady-state MR images showed a thin cochlear branch. We speculated that mumps infection or developmental malformation caused the unilateral sensorineural hearing loss. PMID- 10696019 TI - Midline destructive lesions of the sinonasal tract: simplified terminology based on histopathologic criteria. AB - BACKGROUND AND PURPOSE: Destructive lesions of the sinonasal tract, lacking a discernible etiology and referred to as midline destructive disease, have been pathologically classified in accordance with a variety of confusing terms. Development of new pathologic concepts and immunohistochemical techniques has provided a fresh understanding of these lesions, and, as a result, they can be unified into two distinct pathologic groups: Wegener's granulomatosis and non Hodgkin's T-cell lymphoma. METHODS: We retrospectively reviewed the imaging studies and pathologic specimens of seven patients with prior diagnoses included in the midline destructive disease group. The specimens were reviewed by an oral pathologist using currently accepted pathologic criteria and the newly available immunohistochemical markers CD20, CD45, and CD45RO. Lesions were classified as non-Hodgkin's T-cell lymphomas when positive for CD45 and CD45RO and negative for CD20, and as Wegener's granulomatosis in the presence of noncaseating multinucleated giant cell granulomas and necrotizing vasculitis. RESULTS: Three of the lesions were reclassified as Wegener's granulomatosis and four as T-cell lymphomas after applying these pathologic criteria. There were no distinguishing imaging findings between Wegener's granulomatosis and non-Hodgkin's T-cell lymphoma. CONCLUSION: The current pathologic classification for midline destructive disease should be incorporated into the radiologic lexicon and the use of terms from the old classification system, such as idiopathic midline granuloma and lethal midline granuloma, should be abandoned and no longer be used in radiologic reports. PMID- 10696020 TI - A retrospective analysis of spontaneous sphenoid sinus fistula: MR and CT findings. AB - BACKGROUND AND PURPOSE: The sphenoid sinus is rarely implicated as a site of spontaneous CSF fistula. We undertook this study to evaluate the potential etiopathogenesis of spontaneous CSF fistula involving the sphenoid sinus and to review the imaging findings. METHODS: We retrospectively reviewed the imaging findings of 145 cases of CSF fistula from our departmental archives (August 1995 through August 1998). Fifteen (10%) patients had CSF fistulas involving the sphenoid sinus. Eleven (7%) patients had spontaneous CSF fistulas, whereas in four patients, the CSF fistulas in the sphenoid sinus were related to trauma. Of the 11 patients, nine underwent only plain high-resolution CT and MR cisternography. One patient additionally underwent contrast-enhanced CT cisternography, and one other patient underwent MR cisternography only. For each patient, the CSF fistula site was surgically confirmed. The MR imaging technique included T1-weighted and fast spin-echo T2-weighted 3-mm-thick coronal sequences obtained with the patient in the supine position. The plain high-resolution CT study included 3-mm-thick, and sometimes 1- to 1.5-mm-thick, coronal sections obtained with the patient in the prone position. Similar sections were obtained after injecting nonionic contrast material intrathecally via lumbar puncture for the CT cisternographic study. We evaluated each of the 11 patients for the exact site of CSF leak in the sphenoid sinus. We also determined the presence of pneumatization of lateral recess of the sphenoid sinus, orientation of the lateral wall of the sphenoid sinus, presence of arachnoid pits, presence of brain tissue herniation, and presence of empty sella in each of these patients. RESULTS: The exact sites of the CSF fistulas were documented for all 11 patients by using plain high-resolution CT, MR cisternography, or CT cisternography. In nine (82%) patients, the sites of the CSF fistulas were at the junction of the anterior portion of the lateral wall of the sphenoid sinus and the floor of the middle cranial fossa. In the remaining two (18%) patients, the sites of the CSF fistulas were along the midportion of the lateral wall of the sphenoid sinus. Of these 11 patients, one had bilateral sites of the CSF fistula at the junction of the anterior portion of the lateral wall of the sphenoid sinus with the floor of the middle cranial fossa. In nine (82%) patients, the presence of brain tissue herniation was revealed, and this finding was best shown by MR cisternography. Ten (91%) patients had extensive pneumatization of the lateral recess of the sphenoid sinus, with an equal number having outward concave orientation of the inferior portion of the lateral wall of the sphenoid sinus. In seven (63%) patients, the presence of arachnoid pits, predominantly along the anteromedial aspect of the middle cranial fossa, was shown. In seven (63%) patients, empty sella was shown. For comparison, we reviewed the CT studies of the paranasal sinuses in 100 age-matched control subjects from a normal population. Twenty three had extensive lateral pneumatization of the sphenoid sinus along with outward concavity of the inferior portion of the lateral wall. None of these 23 patients had arachnoid pits. CONCLUSION: The sphenoid sinus, when implicated as a site of spontaneous CSF leak, yields a multitude of imaging findings. These are extensive pneumatization of the lateral recess of the sphenoid sinus, outward concave orientation of the inferior portion of the lateral wall of the sphenoid sinus, arachnoid pits, and empty sella. Considering the normative data, we speculate that this constellation of findings could play a role in the etiopathogenesis of spontaneous sphenoid sinus fistulas. Our findings also show the efficacy of noninvasive imaging techniques, such as plain high-resolution CT and MR cisternography, in the evaluation of sphenoid sinus CSF leak. Our data also suggest that spontaneous sphenoid sinus CSF leak is not an uncommon occurrenc PMID- 10696021 TI - CT and MR imaging appearances of an extraosseous calcifying epithelial odontogenic tumor (Pindborg tumor). AB - We herein report a rare case of extraosseous calcifying epithelial odontogenic tumor with local aggressive behavior. CT and MR imaging showed the distinctive appearances of this histologic entity. We briefly discuss the radiologic features of calcifying epithelial odontogenic tumor and the relevant literature. PMID- 10696022 TI - MR characteristics of muslin-induced optic neuropathy: report of two cases and review of the literature. AB - Muslin-induced optic neuropathy is a rarely reported but important cause of delayed visual loss after repair of intracranial aneurysms. Most of the previously reported cases were published before the introduction of MR imaging. We describe the clinical features and MR appearance of two cases of delayed visual loss due to "muslinoma," and compare them with the 21 cases reported in the literature. PMID- 10696023 TI - Extensive intracranial xanthoma associated with type II hyperlipidemia. AB - Xanthomas are associated with a spectrum of medical conditions, most commonly disorders of lipid storage and lipid metabolism. They occur primarily in the subcutaneous tissues, especially along the Achilles tendon and the extensor tendons of the hands. Intracranial xanthomas are extremely rare. We present a case of an extensive xanthoma of the temporal bone in a patient with hyperlipidemia. PMID- 10696025 TI - Effect of voxel position on single-voxel MR spectroscopy findings. AB - BACKGROUND AND PURPOSE: Single-voxel MR spectroscopy is a widely used tool for evaluating brain tumors. Although extensive data are available on the MR spectral appearance of tumors, less is known about the effect of voxel position on the accuracy of single-voxel MR spectroscopy findings. The purpose of this study was to test the hypothesis that the accuracy of single-voxel MR spectroscopy in the categorization of lesions as either tumor or not tumor is dependent on voxel position. METHODS: Fifty single-voxel MR spectra acquired with a fully automated stimulated-echo spectroscopy sequence were reviewed retrospectively in 43 patients with new or previously treated intra-axial brain tumors. Spectra were analyzed for the presence of choline, creatine, N-acetylaspartate (NAA), and lipid/lactate. Choline/creatine and NAA/creatine peak area ratios were assessed qualitatively. Lesions were grouped into one of three categories on the basis of spectral pattern: tumor, not tumor, or indeterminate. Results of MR spectroscopy were compared with the final histopathologic diagnosis. RESULTS: Histologic confirmation was obtained in 19 patients; MR spectra were interpretable in 17 of those. MR spectra correctly categorized nine of 17 lesions (six tumor, three nontumor). All eight misdiagnosed lesions were tumors. When the MR spectroscopy voxel included the enhancing edge of the lesion, the spectra correctly categorized seven of eight lesions (four of five tumors and all three cases of radiation necrosis). When the MR spectroscopy voxel was positioned centrally within the lesion, the spectra correctly reflected histologic outcome in two of nine lesions (all tumors). CONCLUSION: The reliability of single-voxel MR spectroscopy findings is dependent on voxel position. Spectra obtained from voxels at the enhancing edge of a tumor more accurately reflect lesion histopathology than do spectra obtained from the lesion center, even if the centrally placed voxels contain solidly enhancing tissue. PMID- 10696024 TI - Comparison of relative cerebral blood volume and proton spectroscopy in patients with treated gliomas. AB - BACKGROUND AND PURPOSE: Elevated relative regional cerebral blood volume (rCBV) reflects the increased microvascularity that is associated with brain tumors. The purpose of this study was to investigate the potential role of rCBV in the determination of recurrent/residual disease in patients with treated gliomas. METHODS: Thirty-one rCBV studies were performed in 19 patients with treated gliomas. All patients also had proton MR spectroscopy and conventional MR imaging. Regions of abnormality were identified on conventional MR images by two neuroradiologists and compared with rCBV and MR spectroscopic data. Metabolites and rCBV were quantified and compared in abnormal regions. RESULTS: In high-grade tumors, rCBV values were proportional to choline in regions of tumor and nonviable tissue. Although the presence of residual/recurrent disease was often ambiguous on conventional MR images, the rCBV maps indicated regions of elevated vascularity in all low-grade tumors and in 12 of 17 grade IV lesions. Regions of elevated and low rCBV corresponded well with spectra, indicating tumor and nonviable tissue, respectively. CONCLUSION: This study suggests that rCBV maps and MR spectroscopy are complementary techniques that may improve the detection of residual/recurrent tumor in patients with treated gliomas. Compared with the spectra, the rCBV maps may better reflect the heterogeneity of the tumor regions because of their higher resolution. The multiple markers of MR spectroscopy enable better discrimination between normal and abnormal tissue than do the rCBV maps. PMID- 10696026 TI - MR spectroscopy in gliomatosis cerebri. AB - BACKGROUND AND PURPOSE: The diagnosis of gliomatosis cerebri with MR imaging is known to be difficult. We report on the value of MR spectroscopy in the diagnosis, grading, and biopsy planing in eight patients with histopathologically proved gliomatosis cerebri. METHODS: Patients underwent MR imaging and MR spectroscopy (single-voxel point-resolved spectroscopy [PRESS] at 1500/135, and chemical-shift imaging [CSI] PRESS at 1500/135) before open (n = 4) or stereotactic (n = 4) biopsy. In six patients who underwent CSI, biopsy samples were taken from regions of maximally elevated levels of choline/N-acetylaspartate (Cho/NAA). RESULTS: All patients showed elevated Cho/creatine (Cr) and Cho/NAA levels as well as varying degrees of decreased NAA/Cr ratios, which were most pronounced in the anaplastic lesions. In low-grade lesions, there was a maximum Cho/NAA ratio of 1.3, whereas in anaplastic tumors, the maximum Cho/NAA level was at least 2.5. Spectra in two patients with grade III lesions revealed a lactate peak; lactate and lipid signals were seen in two patients with grade IV lesions. Biopsy specimens from regions with maximally elevated levels of Cho/NAA showed dense infiltration of tumor cells. CONCLUSION: MR spectroscopy might be used to classify gliomatosis cerebri as a stable or a progressive disease indicating its potential therapeutic relevance. PMID- 10696027 TI - Dynamic contrast-enhanced MR angiography and MR imaging of the carotid artery: high-resolution sequences in different acquisition planes. AB - BACKGROUND AND PURPOSE: First-pass contrast-enhanced MR angiography has become the technique of choice for studying the carotid bifurcation, but this method has some limitations. We evaluated the clinical utility of performing 3D contrast enhanced MR angiography in the axial plane immediately after performing angiography in the coronal plane. METHODS: Cervical carotid arteries of 80 consecutive patients were studied on a 1.5-T MR imager with phased-array coils. Coronal 3D MR angiography was performed after administering a bolus injection of contrast material (20 mL) with automatic triggering. This was immediately followed by an axial acquisition. We measured carotid diameters on the contrast enhanced MR angiograms as well as on intra-arterial digital subtraction angiograms according to established criteria. We also evaluated original source MR angiograms. RESULTS: Angiograms obtained in the axial plane correlated better with the intra-arterial digital subtraction angiograms than did the coronal angiograms. When first-pass contrast-enhanced MR angiography was incomplete because of a failure of triggering, the second-phase acquisition provided sufficient image quality. Original source images suffered from ring artifacts, low axial resolution, and a low level of soft-tissue visualization. Axial-based source images showed flow-independent contrast filling to the patent lumen with sufficient visualization of plaque morphology, thickened arterial wall, and surrounding disease processes, such as tumors. CONCLUSION: With the addition of a 1-minute second-phase 3D acquisition in a different plane immediately after first pass contrast-enhanced MR angiography, one can obtain a more accurate depiction of the carotid bifurcation, insurance against failure of triggering, and diagnostic source images. PMID- 10696028 TI - The effect of contrast material on transcranial Doppler evaluation of normal middle cerebral artery peak systolic velocity. AB - BACKGROUND AND PURPOSE: Several recent studies have shown that sonographic contrast agents may affect transcranial Doppler evaluation of the arterial peak systolic velocity (PSV). Some investigators reported an increase in PSV, and others reported no change in PSV compared with baseline values. This study was conducted to determine the effect of sonographic contrast agent on PSV measured in normal middle cerebral arteries. METHODS: Continuous spectral Doppler sonography was performed on the right middle cerebral artery of 20 participants with angiographically proven normal intracranial vasculature. Videotaping was performed in each case from the initiation of the administration of contrast medium until the effect of the contrast agent on the PSV subsided. The PSV values were normalized for each participant, were pooled, and were plotted as a function of time. RESULTS: PSV increased in all participants after the administration of contrast material; the mean maximum increase was 24+/-7.4% (mean +/- standard deviation) (range, 15-36%). The mean duration of PSV increase was 320+/-97 s (range, 165-465 s). CONCLUSION: The middle cerebral artery PSV increased substantially after the administration of contrast material. This effect needs to be considered if velocity thresholds developed for disease detection without the use of contrast materials are used when contrast agents are administered. PMID- 10696029 TI - Unusual MR findings of the brain stem in arterial hypertension. AB - MR imaging findings have been reported in only a few cases of severe arterial hypertension. We report two cases of severe paroxysmal arterial hypertension associated with unusual brain stem hyperintensity. The lesions improved dramatically after stabilization of blood pressure, suggesting that edema could be the main cause of the MR imaging-observed hyperintensity. PMID- 10696030 TI - Optimized activation of the primary sensorimotor cortex for clinical functional MR imaging. AB - BACKGROUND AND PURPOSE: One application of functional MR imaging is to identify the primary sensorimotor cortex (M1 and S1) around the central sulcus before brain surgery. However, it has been shown that undesirable coactivation of nonprimary motor areas, such as the supplementary motor area and the premotor area, can interfere with the identification of the primary motor cortex, especially in patients with distorted anatomic landmarks. We therefore sought to design a simple functional MR imaging paradigm for selective activation of the primary sensorimotor cortex. METHODS: Different paradigms using finger tapping for motor activation were examined and compared with respect to the distribution of activated voxels in primary and nonprimary cortical areas. Studies were conducted in 14 healthy volunteers using a blood oxygen level-dependent multislice echo-planar imaging sequence. RESULTS: The most selective activation of the primary sensorimotor cortex was obtained with a paradigm combining right sided finger tapping as the activation condition with left-sided finger tapping as the control condition. Analysis of the signal time course of primary and nonprimary areas revealed that the highly selective primary motor activation was due to it being restricted to contralateral finger movements, as opposed to the nonprimary motor areas, which were activated by ipsilateral, contralateral, and bilateral finger movements alike. CONCLUSION: When performing functional MR imaging to determine the location of the primary sensorimotor cortex, one should compare unilateral voluntary movements as the activation condition with contralateral movements as the control condition to accentuate activation of the primary motor area and to suppress undesirable coactivation of nonprimary motor areas. PMID- 10696031 TI - Cortical/subcortical disease burden and cognitive impairment in patients with multiple sclerosis. AB - BACKGROUND AND PURPOSE: We assessed whether the extent of macro- and microscopic disease in the cortical and subcortical brain tissue, as revealed by MR and magnetization transfer (MT) imaging, correlates with cognitive dysfunction in patients with multiple sclerosis (MS). METHODS: Dual-echo rapid acquisition with relaxation enhancement (RARE), fast fluid-attenuated inversion recovery (fast FLAIR), T1-weighted, and MT MR images of the brain were obtained from 16 MS patients with cognitive impairment and from six without. Impaired and unimpaired patients were similar across demographic and other disease-related variables. Total and cortical/subcortical lesion loads were assessed using RARE, fast-FLAIR, and T1-weighted sequences. In each patient, cortical/subcortical disease was also assessed by means of MT ratio (MTR) histographic analysis. RESULTS: All the impaired patients had multiple hyperintense lesions in the cortical/subcortical regions on both RARE and fast-FLAIR images; two unimpaired patients had such lesions on the RARE images and four had them on the fast-FLAIR images. Total and cortical/subcortical RARE/fast-FLAIR hyperintense and T1 hypointense lesion loads were significantly greater in the group of cognitively impaired patients. Patients with cognitive deficits also had significantly lower MTR histographic values for all the variables. A multivariate regression model showed that average cortical/subcortical brain MTR was the only factor that was significantly associated with cognitive impairment. CONCLUSION: The extent and severity of MS disease in the cortical and subcortical regions significantly influence the cognitive functions of MS patients. MTR histographic findings suggest that subtle changes undetectable by conventional imaging are also important in determining MS cognitive decline. PMID- 10696032 TI - MR imaging of the hippocampus in normal pressure hydrocephalus: correlations with cortical Alzheimer's disease confirmed by pathologic analysis. AB - BACKGROUND AND PURPOSE: MR studies have shown hippocampal atrophy to be a sensitive diagnostic feature of Alzheimer's disease (AD). In this study, we measured the hippocampal volumes of patients with a clinical diagnosis of normal pressure hydrocephalus (NPH), a potentially reversible cause of dementia when shunted. Further, we examined the relationship between the hippocampal volumes and cortical AD pathologic findings, intracranial pressure, and clinical outcomes in cases of NPH. METHODS: We measured hippocampal volumes from 37 patients with a clinical diagnosis of NPH (27 control volunteers and 24 patients with AD). The patients with NPH underwent biopsy, and their clinical outcomes were followed for a year. RESULTS: Compared with those for control volunteers, the findings for patients with NPH included a minor left-side decrease in the hippocampal volumes (P < .05). Compared with those for patients with AD, the findings for patients with NPH included significantly larger hippocampi on both sides. Although not statistically significant, trends toward larger volumes were observed in patients with NPH who had elevated intracranial pressure, who benefited from shunting, and who did not display cortical AD pathologic findings. CONCLUSIONS: Measurements of hippocampal volumes among patients with a clinical diagnosis of NPH have clear clinical implications, providing diagnostic discrimination from AD and possibly prediction of clinical outcome after shunting. PMID- 10696033 TI - Radiologic-pathologic findings in raccoon roundworm (Baylisascaris procyonis) encephalitis. AB - A 13-month-old boy developed eosinophilic meningoencephalitis, retinitis, and a protracted encephalopathy with severe residual deficits. The initial MR examination revealed diffuse periventricular white matter disease, and follow-up images showed atrophy. Brain biopsy, serology, and epidemiologic studies lead to the diagnosis of Baylisascaris procyonis infection, a parasitic disease contracted through exposure to soil contaminated by the eggs of a common raccoon intestinal roundworm. The pathologic, epidemiologic, and imaging features of this disease are herein reviewed. PMID- 10696034 TI - Dynamic CT perfusion to assess the effect of carotid revascularization in chronic cerebral ischemia. AB - We present the case of a female patient who was studied with dynamic contrast enhanced CT perfusion before and after carotid revascularization. Before treatment, there was decreased perfusion in the ipsilateral insula, which was shown to be resolved on the scan obtained 1 day after treatment, indicating the technical success of the revascularization. In the ipsilateral basal ganglia, there was delayed contrast agent clearance from the tissue, which was attributed to vasodilation; after revascularization, there remained a subtle stenotic effect. The observed changes in the dynamic CT perfusion study suggest that this technique may be a useful tool in the evaluation of patients with asymmetrical cerebral blood flow. PMID- 10696035 TI - Cost utility analysis of radiographic screening for an orbital foreign body before MR imaging. AB - BACKGROUND AND PURPOSE: Our purpose was to evaluate the cost-effectiveness of clinical versus radiographic screening for an orbital foreign body before MR imaging. METHODS: Costs of screening were determined on the basis of published reports, disability rating guides, and a practice survey. Base case estimates were derived from published guidelines. A single-state change model was constructed using social cost as the unit of analysis. Sensitivity analysis was performed for each variable. The benefit of screening was avoidance of immediate, permanent, nonameliorable, unilateral blindness. RESULTS: Using base case estimates and a discount rate of zero, we calculated the cost of the current guideline as $328,580 per quality-adjusted life-year saved. Sensitivity analysis identified screening cost as a critical variable. Discount rates and effectiveness of foreign body removal also were found to be important factors. Probability of injury and prevalence of foreign body may impact the analysis. CONCLUSION: Clinical screening before radiography increases the cost effectiveness of foreign body screening by an order of magnitude, assuming base case ocular foreign body removal rates. Asking the patient "Did a doctor get it all out?" serves this purpose. Occupational history by itself is not sufficient to mandate radiographic orbital screening. Current practice guidelines for foreign body screening should be altered. PMID- 10696036 TI - M. Judith Donovan-Post, Fifth President of the American Society of Spine Radiology PMID- 10696037 TI - Breast reduction outcome study. AB - The outcome of 75 consecutive bilateral breast reduction procedures on 73 patients (65 inferior pedicle and 9 superior pedicle with inverted-T closure, and 1 ultrasonic-assisted lipoplasty) is evaluated. Fifty-six surveys at an average follow-up of 35 months, 36 standardized examinations at an average follow-up of 31 months, 52 paired preoperative/postoperative photographs, and 73 chart reviews comprise the data. Mean age at surgery was 32 years. Of the patients surveyed, there were 51 whites, 18 blacks, and 4 other races. The majority of survey respondents reported improvement in breast appearance (93%), self-esteem (85%), posture (84%), and activity level (77%). The percentage of patients free of back pain rose from 9% preoperatively to 59% postoperatively. Some decrease in nipple sensitivity was noted by survey in 34 nipple-areolar complexes (31%), and clinical examination detected decreased sensitivity in 16% of nipples. Thirty-one percent of patients reported improvement in their intimate relationship postoperatively and 65% reported no change. Ninety-five percent felt they had made the right decision in having breast reduction surgery. Physical examination revealed excellent maintenance of shape and impressive fading of surgical scars over the years in the majority of patients. The postoperative aesthetic result depends on several important preoperative factors, including skin tone, breast shape, and degree of ptosis. Adverse sequelae included infection (1.3%), nipple cyanosis (1.3%) but no nipple-areolar necrosis, and wound dehiscence (4.0%) but no skin flap necrosis. Late complications included underresection requiring reoperation (4.0%), fat necrosis (2.7%), hypertrophic scars (6.7%), and pseudoptosis (12.0%). Breast reduction surgery results in a good outcome for most patients, with high patient satisfaction. Patients are accepting of the large T pattern scar that fades surprisingly well with time in exchange for symptomatic relief and substantial improvement in breast size and shape. PMID- 10696038 TI - An alternative technique for managing the lower breast skin during secondary breast reconstruction with a TRAM flap. AB - During secondary breast reconstruction with a transverse rectus abdominis musculocutaneous (TRAM) flap, the native breast skin between the mastectomy scar and inframammary crease is usually left intact, excised, or deepithelialized. The authors have developed transposition flaps utilizing this skin and subcutaneous tissue in selected patients. This technique is most useful in patients who present for secondary reconstruction whose remaining lower breast skin may have contracted or in patients who should have a vertically inset TRAM flap but do not have a wide enough flap relative to the length of the inframammary crease. Although the authors use this technique infrequently due to additional scars placed on the breast, it is a useful technique to add to the armamentarium of the reconstructive surgeon. PMID- 10696039 TI - Augmentation in ptotic and densely glandular breasts: prevention, treatment, and classification of double-bubble deformity. AB - After breast augmentation, separation of breast tissue from the implant is common, especially in patients with well-formed preoperative breasts. This problem is enhanced to a marked deformity in cases of scar contracture with firm, fixed implants. This paper addresses this problem preoperatively and therapeutically in secondary correction of double-bubble and waterfall deformity. The author classifies and explains double-bubble deformity in patients in whom the implant is below the normal crease, with glandular breast tissue superior and anterior to the implant. In "waterfall" deformity (a term suggested by the author), the glandular breast tissue droops over the implant and is inferior and anterior to the implant. Treatment used consists of opening the breast tissue from its posterior surface using radial incisions to accommodate the implant. This allows the two structures--the breast tissue and the implant-to blend as one unit with satisfactory results. The technique is easy to perform and teach. Complications are similar to those of regular breast augmentation. Strangely, radial incisions have not increased complications, and there have been no cases of seroma or hematoma to date. PMID- 10696040 TI - The role of tissue expansion in abdominal wall reconstruction. AB - Abdominal wall reconstruction of ventral hernia defects with loss of visceral domain and inadequate soft-tissue coverage presents a surgical challenge. Four patients with large, skin grafted ventral hernia defects were treated by staged abdominal wall reconstruction. During the initial stage, tissue expanders were placed under the skin and subcutaneous tissue lateral to the defects. After adequate interval expansion, the second stage was performed. The expanders were removed, the visceral contents reduced easily, and the fascia reapproximated with polypropylene mesh. The expanded skin was closed easily over the fascial repair. All four patients were reconstructed successfully without complications. Tissue expansion can restore abdominal domain and allow soft-tissue closure in complicated ventral hernia defects. PMID- 10696041 TI - Use of specialized bone screws for intermaxillary fixation. AB - Fixation of the injured mandible to the maxilla is a proven method of stabilizing mandibular fractures and ensuring proper occlusion. The authors report their results with new specialized intraoral bone screws (IMF Screw System; Howmedica Leibinger, Inc., Carrollton, TX) that are designed for the purpose of achieving intermaxillary fixation (IMF). Nineteen patients were placed into rigid IMF using IMF screws alone. Indications were nondisplaced mandibular fractures; symphyseal, body, and angle fractures; midfacial fractures requiring temporary IMF; and edentulous patients with any of these fracture types and an adequate prosthesis. All procedures were performed with the patient under general anesthesia. The authors found that the operative time was markedly shorter than with standard IMF techniques, patient satisfaction was high, and there were no infections related to the screws. All 19 patients remained in stable, accurate occlusion and had adequate healing. One patient continues to have paraesthesias in the mental nerve distribution after screw removal. Although there is the potential for tooth and nerve injury when screws are placed improperly, the IMF Screw System seems to be a safe and reliable method of achieving secure mandibular fixation. PMID- 10696042 TI - The peroneus brevis muscle flap for lower leg defects. AB - The peroneus brevis is a small muscle with a Mathes-Nahai type II vascular pattern found in the lateral compartment of the leg. It is supplied by branches of the peroneal artery and it maintains its muscular component to the lateral malleolus, allowing it to be transposed to cover small distal third defects. The authors describe their experience with eight peroneus brevis flaps covered with split-thickness skin grafts utilized to cover lateral malleolar fractures with exposed hardware or bone and one case of exposed Achilles' tendon. Seven flaps were successful and one (in a diabetic) underwent partial necrosis, requiring a small fasciocutaneous flap. The peroneus brevis flap provides limited coverage of the distal third of the leg but can be quite useful for problematic wounds of this difficult area. PMID- 10696043 TI - A simplified pneumotachometer for the quantitative assessment of velopharyngeal incompetence. AB - By applying pipe flow rate mechanics, the authors have developed a simple and inexpensive device that can measure oral and nasal airflow and pressure quantitatively. It allows clinicians to document objectively the degree of velopharyngeal incompetence and nasal airway obstruction. This device is a modified U-manometer with a Y-connector to a collection bag. This prototype is being used in the authors' cleft palate clinic and they have found it useful in documenting quantitatively the amount of nasal air escape from either nostril with the corresponding pressure and flow. The primary measurement, which is reproducible and reliable, is the product of a cross-sectional area of oronasal communication and the pressure differential between the oral and nasal cavity. With some modifications, this device can be adapted to estimate the size of the oronasal communication. PMID- 10696045 TI - Exploiting the septum for maximal tip control. AB - Manipulating tip projection and rotation is among the greater challenges in aesthetic surgery. Among common current techniques such as columellar struts and projecting tip grafts, all have considerable failure rates and/or complications. Powerful static and dynamic forces act on the tip, and unfortunately their magnitude and direction vary greatly from the time of surgery when decisions are made to the postoperative period. Traditional techniques have not taken sufficient advantage of the one neighboring stable structure--the septum. Through direct straddling, the medial crura on the septum or the more commonly applicable septal extension graft, tip placement at the end of surgery will vary minimally postoperatively. PMID- 10696044 TI - Lacrimal duct injuries revisited: a retrospective review of six patients. AB - This is a retrospective review of 6 patients treated for lacrimal duct injuries between December 1994 and October 1997. Four patients had inferior canalicular lacerations, one had a superior laceration, and one had a combined inferior and superior laceration. Associated injuries included facial fractures (1 patient), multiple facial lacerations (2 patients), and avulsion of the medial canthus (1 patient). All patients were repaired with Crawford tubes using a semicircular technique. The mean time of lacrimal intubation was 4.2 months (range, 3.6-4.5 months). One tube fell out at an undetermined time during the follow-up period. Visible anatomic restoration was considered excellent in all patients. There were no instances of persistent epiphora or other complications. Although there are several techniques available for treating lacrimal duct injuries in the course of dealing with general facial trauma, the semicircular technique using a Crawford intubation system yielded consistently reliable results with minimal or no complications. PMID- 10696046 TI - Correction of axillary burn scar contracture with the thoracodorsal perforator based cutaneous island flap. AB - Axillary scar contracture is observed frequently after severe burn insult and is usually accompanied by injuries to the adjacent area. Although many therapeutic methods, including skin grafting, Z-plasties, local flaps, island flaps, and free flaps, have been established, each technique has its own advantages and disadvantages in specific situations. The decision regarding which technique to use can only be made after consideration is given to the merits of the individual case. We applied thoracodorsal perforator-based cutaneous flaps to 5 patients with axillary burn scar contractures and damaged adjacent tissues. In 1 patient both axillae were involved. Elevated flaps as large as 11 x 27 cm in size were used. All flaps survived completely even when raised in scar tissue. The donor sites were closed primarily except one, which needed a skin graft. Three patients obtained satisfactory release with more than 160 deg shoulder abduction. In 2 patients, release was incomplete with only 110 deg shoulder abduction, but neither one required a second release. The range of motion in terms of shoulder abduction was improved preoperatively (30-90 deg) to postoperatively (110-170 deg). The thoracodorsal perforator-based cutaneous flap presents a very useful reconstructive method for the treatment of axillary defects. PMID- 10696047 TI - Intralesional laser therapy of extensive hemangiomas in 100 consecutive pediatric patients. AB - The authors have treated 100 consecutive pediatric patients with capillary/cavernous hemangiomas (age range, 1.3 months to 16 years; mean age, 26.6 months; 30 male and 70 female patients) with intralesional laser therapy during a 3-year period. All patients have been followed for a minimum of 6 months after treatment (range, 6-36 months; mean, 18 months). Indications for intralesional laser treatment included interference with vision, blockage of the nose or mouth, ulceration, bleeding, and rapid, uncontrollable growth. The Nd:YAG laser was used in 70 patients, and the Potassium, Titanyl, Phosphate (KTP) laser was used in 30 patients. Fifty-five hemangiomas were in the head and neck region, excluding the orbit; 25 were in the trunk or extremities; 10 were periorbital; and 10 involved multiple sites. Seventy patients (70%) received one treatment, 20 patients (20%) received two treatments, 7 patients (7%) received three treatments, and 3 patients (3%) received four or more treatments. No appreciable differences were noted between treatment with the Nd:YAG and KTP lasers. Forty six patients had more than a 90% reduction in the overall size of the hemangiomas whereas 54 patients had a 50% to 90% reduction in the size of the hemangioma. After maximal reduction in size of the cavernous component was achieved, the external capillary component, found in 68 patients, was treated with a tunable dye laser. Seventy-six patients underwent surgical resection after maximal lesion involution. Residual induration due to lesion fibrosis was treated with local steroid injections in 13 patients. There were four operative complications attributable to intralesional laser therapy. Two patients had residual midfacial weakness, and two patients had punctuated skin burns after intralesional treatment. The authors have found intralesional laser therapy to be a valuable tool in the treatment of large capillary/cavernous hemangiomas, often rendering an inoperable lesion safely resectable, or markedly decreasing the size and functional impact of the lesion. PMID- 10696048 TI - Neo-osseous flaps using demineralized allogeneic bone in a rat model. AB - Surgical reconstruction with revascularized bone grafts can be compromised by donor tissue limitations and may be refined by prefabrication of compound neoflaps using bone substitutes. The principal suitability of demineralized allogeneic bone (DALB) slabs in fabricating neo-osseous flaps based on the inferior epigastric vascular system was studied and compared with neoflaps with autologous bone (AUB). In 45 rats, the histological pattern of bone formation in response to angiogenesis induced by vessel implantation was assessed, and characteristics of vascularization of the neoflap were studied microangiographically at 2, 4, 6, and 8 weeks. Histological techniques included decalcified and nondecalcified sections, as well as intravital polyfluorochrome labeling. Blood flow of the neoflap was also assessed quantitatively using 15 microm microspheres labeled with technetium 99-methylene diphosphate (99-MDP) 8 weeks after flap fabrication. Although the DALB neoflaps showed consistent bone formation and neovascularization, the bone regeneration process was delayed distinctly in comparison with AUB. Microangiographically, however, no differences between the two types of grafts became apparent during all time periods tested. Furthermore, the radioactivity of the DALB neoflap, which means bone blood flow per dry weight, was significantly higher than in AUB grafts and even more than that of intact iliac bone (p = 0.001). The exact meaning of elevated blood flow in DALB and similar degrees of vascularization corresponding to native AUB grafts remains to be determined, but may be a sign of ongoing bone formation resulting in a suitable DALB-containing neo-osseous flap in the long term. The authors findings support that allogeneic bone could be a potential substitute for AUB in creating a prefabricated neo-osseous flap. PMID- 10696049 TI - A new model for experimental tendon adhesions in the chicken. AB - A minimally invasive model using a manual abrader to induce adhesions in the chicken's central digit is described. The flexor synovial sheath and the profundus tendon were abraded with access through small flaps at the level of the proximal and distal phalanges of the avian long toes. The birds were divided into two groups according to the severity of the induced trauma. Group I birds received an abrasion injury and were euthanized to allow biomechanical testing 5 weeks postoperatively. Group II birds had a more severe abrasion and were euthanized similarly and tested 5 weeks after surgery. Results were compared with nonsurgical controls. Long toe function was evaluated weekly by measuring (1) the range of active flexion of each interphalangeal joint, resolved to total angular range; (2) the grasping ability on graded-diameter perches; and (3) the flexion deficit of the long toe. Postmortem biomechanical properties of the adhesions were measured. There was a significant difference between the unoperated controls and abraded digits of both groups in all parameters (p < 0.001). There was, in addition, a marked change in most of the measured parameters between groups I and II. In group I digits the functional and biomechanical deficit was less than group II. In summary, this animal model of long-segment abrasive injury to the tendon and sheath is a simple and reproducible method to generate adhesions and can be used for the evaluation of treatment modalities for adhesion prevention. PMID- 10696050 TI - Skeletal deformities due to tissue expanders: report of two patients. AB - Two patients, burned at ages 2 and 5, developed scars that required multiple reconstructive operations over a period of several years. Tissue expanders were used as part of their reconstructive procedures. After the expanders were removed, skeletal deformity was encountered in the area underlying the expander in each patient. Patient 1 had deformity of the rib cage, and Patient 2 had deformity of the outer table of the skull. No treatment was felt to be indicated. Surgeons should be aware of the possibility of the development of this problem. PMID- 10696051 TI - Replantation of the penis: a patient report. AB - Replantation of the penis is an unusual case in this country and it is unlikely that most plastic and reconstructive surgeons or urologists will see one during their career. A successful repair of a self-inflicted amputation of the penis is presented. The unique anatomy of the penis pertinent to replantation is reviewed, and current concepts and recommendations in performing replantation of the penis are presented. PMID- 10696052 TI - Rare complication of massive hemorrhage in neurofibromatosis with arteriovenous malformation. AB - Neurofibromatosis is rare in the general population. Its clinical manifestations are systemic and variable. The clinical presentation of cutaneous lesions is even more variable. Some patients have giant tumors in the trunk or limbs (so-called "elephant neurofibromatosis"). The pathological findings are diffuse neurofibromatosis of the nerve trunk associated with overgrowth of subcutaneous tissue and skin. The associated vascular malformations make most surgeons hesitant to address them because bleeding to death is possible if the bleeding is not well controlled. According to the authors' experience in treating this complication of neurofibromatosis, they noted that there are three key points to reducing the amount of hemorrhage to a minimal level: (1) hypotensive anesthesia, (2) preliminary sutures around the lesion, (3) ligation of the limited numbers of feeding vessels in the vascular malformation of the neurofibroma. Ligating these pedicles can decrease bleeding during resection of the neurofibroma, as demonstrated in their patient. PMID- 10696053 TI - The vessel loop shoelace technique for closure of fasciotomy wounds. AB - Compartment syndrome of the extremity may occur after severe trauma secondary to fractures, vascular ischemia, crush, or electrical injury. Treatment consists of expedient fasciotomy to avoid permanent injury to muscles or nerves. Management of the wounds postoperatively has consisted traditionally of primary closure, healing by secondary intention, or split-thickness skin grafting to cover defects. The fasciotomy wound may remain substantial secondary to soft-tissue swelling and edema. The authors present an alternative protocol for fasciotomy wound management, consisting of gradual closure with progressive tension using vessel loops. The vessel loops are placed intraoperatively during the compartment release and are attached to the wound margins using standard skin staples. The loops are tightened progressively postoperatively during routine dressing changes, resulting in closure of the wound within 2 weeks. The advantages over split-thickness grafting include avoidance of donor morbidity and better cosmesis. Sporadic case reports using similar techniques have been published in the orthopedic literature with comparable results. The current series includes 37 patients, ages 9 to 48 years, who were treated for open fasciotomy. There were 11 upper extremity and 26 lower extremity wounds treated, all of which were closed within 3 weeks. PMID- 10696054 TI - What's the alternative? PMID- 10696055 TI - Re: Gigantomastia during pregnancy: a case report. PMID- 10696056 TI - The bullet-type fracture: a peculiar type of orbital wall fracture. PMID- 10696058 TI - Expander pressure as a safety factor for expansion. PMID- 10696057 TI - Successful replantation of a completely avulsed scalp. PMID- 10696059 TI - Full-thickness eye lid burn with intact levator function. PMID- 10696060 TI - Muc-5/5ac mucin messenger RNA and protein expression is a marker of goblet cell metaplasia in murine airways. AB - Airway inflammation, hyperreactivity, increased number of goblet cells, and mucus overproduction characterize asthma. Respiratory challenge with ovalbumin (OVA) of sensitized mice has been shown by several laboratories to cause pulmonary pathology similar to that observed in human allergic asthma. Recently, interleukin (IL)-13 has been shown to be a central mediator in this process. Because the airways of healthy mice have few, if any, mucus-producing cells, an increase in the number of these cells likely reflects induction of mucin-gene expression. The purpose of this study was to identify mucin genes induced as a result of airway goblet-cell metaplasia (GCM) in mice sensitized and challenged with OVA or in mice treated with IL-13 alone. BALB/c mice were sensitized by intraperitoneal injection (Days 0, 4, 7, 11, and 14) and intranasal instillation (Day 14) of 100 microg of OVA in saline, and then challenged by intranasal instillation (Days 25, 26, and 27) of the same. IL-13-treated mice received 5 microg of IL-13 by intranasal instillation on three consecutive days. Control mice were given saline alone. All mice were studied 24 h after the last challenge. Histologic analysis of the lungs revealed both a striking peribronchial and perivascular lymphocytic and eosinophilic inflammation and airway GCM in OVA-treated mice, and also airway GCM without inflammation in IL-13 treated mice. Northern blot analysis of lung RNA demonstrated (1) expression of Muc-5/5ac messenger RNA (mRNA) in OVA-treated and IL-13-treated mice, but not in control mice; (2) expression of Muc-1 mRNA at comparable levels in all mice regardless of treatment; and (3) no expression of Muc-2 or Muc-3 mRNA in control or treated mice. Western blot analysis demonstrated the expression of Muc-5/5ac protein (both apomucin and glycosylated mucin) in lung lysates of OVA-treated (but not control) mice, and also the expression of Muc-5/5ac mucins in the bronchoalveolar lavage fluid of OVA-treated and IL-13-treated mice. These findings demonstrate that airway GCM is associated with the induction of pulmonary expression of Muc-5/5ac mRNA and mucin in murine models of allergic asthma. PMID- 10696061 TI - The B7/CD28/CTLA4 T-cell activation pathway. Implications for inflammatory lung disease. PMID- 10696062 TI - B7-1 (CD80) and B7-2 (CD86) have complementary roles in mediating allergic pulmonary inflammation and airway hyperresponsiveness. AB - We examined the roles of B7-1 (CD80) and B7-2 (CD86) in a model of allergic pulmonary inflammation and airway hyperresponsiveness (AHR) by using mice with germline deletions of the B7-1 and/or B7-2 molecules. Multiple parameters of the allergic response were affected to varying degrees by the absence of B7-1 and/or B7-2. Mice lacking both B7-1 and B7-2 had no elevation of serum immunoglobulin E, lack of airway eosinophilia, and no AHR. These same disease parameters were also reduced in mice lacking either B7-1 or B7-2. Lack of B7-1 and/or B7-2 resulted in an increase in T-helper 1 cytokine production. Our observations suggest that whereas B7-2 is quantitatively more significant in the induction of this response, B7-1 and B7-2 may have complementary roles in mediating the development of allergic pulmonary inflammation. PMID- 10696063 TI - Oxygen-independent upregulation of vascular endothelial growth factor and vascular barrier dysfunction during ventilated pulmonary ischemia in isolated ferret lungs. AB - Vascular endothelial growth factor (VEGF) is a potent mediator of endothelial barrier dysfunction, and is upregulated during ischemia in many organs. Because ventilated pulmonary ischemia causes a marked increase in pulmonary vascular permeability, we hypothesized that VEGF would increase during ischemic lung injury. To test this hypothesis, we measured VEGF expression by Northern and Western blot analysis in isolated ferret lungs after 45 (n = 12) or 180 (n = 12) min of ventilated (95% or 0% O(2)) ischemia. Pulmonary vascular permeability, assessed by measurement of osmotic reflection coefficient for albumin (sigma(alb)), was evaluated in the same lungs, as was expression of the transcription factor, hypoxia-inducible factor (HIF)-1alpha. Distribution of VEGF as a function of ischemic time and oxygen tension was also evaluated by immunohistochemical staining in separate groups of lungs (n = 3). VEGF messenger RNA (mRNA) increased 3-fold by 180 min of ventilated ischemia, independent of oxygen tension. VEGF protein increased in parallel to mRNA. Immunohistochemical staining demonstrated the appearance of VEGF protein along alveolar septae after 180 min of hyperoxic ischemia, and after 45 or 180 min of hypoxic ischemia. sigma(alb) was not altered by 45 min of hyperoxic ischemia (0.69+/-0.09 versus 0.50+/-0.12, respectively), but decreased significantly after 180 min of hyperoxic ischemia and after 45 and 180 min of hypoxic ischemia (0.20+/-0.03, 0.26+/-0.08, and 0.23+/-0.03, respectively; P<0.05). HIF-1alpha mRNA increased during both hyperoxic and hypoxic ischemia, but HIF-1alpha protein increased only during hypoxic ischemia. These results implicate VEGF as a potential mediator of increased pulmonary vascular permeability in this model of acute lung injury. PMID- 10696064 TI - Degree of lung maturity determines the direction of the interleukin-1- induced effect on the expression of surfactant proteins. AB - Intra-amniotic interleukin (IL)-1 increases surfactant components in immature fetal lung, whereas high IL-1 after birth is associated with surfactant dysfunction. Our aim was to investigate whether the fetal age influences the responsiveness of surfactant proteins (SPs) to IL-1. Rabbit lung explants from fetuses at 19, 22, 27, and 30 d of gestation and 1-d-old newborns were cultured in serum-free medium in the presence of recombinant human (rh) IL-1alpha or vehicle. The influence of IL-1alpha on SP-A, -B, and -C messenger RNA (mRNA) content was dependent on the conceptional age. In very immature lung on Day 19, rhIL-1alpha (570 ng/ml for 20 h) increased SP-A, -B, and -C mRNA by 860+/-15%, 314+/-108%, and 64+/-17%, respectively. The increase in SP-A mRNA was evident within 4 to 6 h. IL-1alpha increased the SP-A concentration in alveolar epithelial cells and in the culture medium within 20 h. In contrast, at 27 to 30 d of gestation and in newborns, IL-1alpha decreased SP-C, -B, and -A mRNA by means of 64 to 67%, 48 to 59%, and 12 to 15%, respectively. SP-B protein decreased by 45 to 60%. The decrease in mRNA became evident within 8 to 12 h and was dependent on IL-1 concentration. On Day 27, IL-1alpha accelerated the degradation of SP-B mRNA in the presence of actinomycin D. IL-1 did not increase the degradation rate of SP-A mRNA unless both actinomycin D and cycloheximide were added to the explants. The present findings may explain some of the contrasting associations between inflammatory cytokines and lung diseases during the perinatal period. The determinants of the direction of the IL-1 effect on the expression of SPs remain to be identified. PMID- 10696065 TI - Airway hyperresponsiveness and airway obstruction in transgenic mice. Morphologic correlates in mice overexpressing interleukin (IL)-11 and IL-6 in the lung. AB - Understanding the sources of variation in airway reactivity and airflow is important for unraveling the pathophysiology of asthma, obstructive lung disease, and other pulmonary disorders. Transgenic expression of two closely related cytokines in the mouse lung produced opposite effects on these parameters. Interleukin (IL)-6 did not alter basal airways resistance and decreased methacholine responsiveness, whereas IL-11 caused airways obstruction and increased airway responses to methacholine. To clarify these differences we examined histologic sections and used morphometry to compare bronchiolar and parenchymal dimensions in 1- to 2-mo-old transgenic mice expressing IL-6 or IL-11 and littermate control mice. Both transgenic strains showed similar emphysema like airspace enlargement, nodular peribronchiolar collections of mononuclear cells, thickening of airway walls, and subepithelial airway fibrosis. When compared with littermate control mice, the IL-6 mice showed an approximately 50% increase in the caliber of their bronchioles and an increase in airway wall thickness that was in proportion to the increase in the size of their airways. In contrast, the remodeling response was more robust in the IL-11 transgenic mice. It was also seen in airways with normal external and luminal diameters and thus was out of proportion to the caliber of their airways. These results support the hypothesis that structural alterations and resulting caliber changes of respiratory airways can have important effects on airway physiology and reactivity. PMID- 10696066 TI - Diesel exhaust (DE)-induced cytokine expression in human bronchial epithelial cells: a study with a new cell exposure system to freshly generated DE in vitro. AB - We devised a new in vitro cell exposure system to freshly generated diesel exhaust (DE), different from conventional in vitro culture systems, to examine the effects of DE on human epithelial cells. Using this system, we investigated the effects of DE on cytokine gene expressions in BET-1A human bronchial epithelial cells. DE significantly decreased [(3)H]thymidine incorporation into BET-1A cells. DE had a significant stimulatory effect on interleukin (IL)-8 release to a marked degree. IL-8 and transforming growth factor (TGF)-beta1 messenger RNA (mRNA) expression were induced by DE in a time-dependent manner. The gas obtained by filtration of DE alone did not show a sustained increase in IL-8 protein levels and showed no induction of IL-8 mRNA, suggesting that DE particles (DEPs) play an important role in the induction of IL-8 at both mRNA and protein levels. Antioxidants, pyrrolidine dithiocarbamate, and N-acetyl-L cysteine significantly inhibited IL-8 mRNA and protein levels by BET-1A cells. These results indicate that freshly generated DEPs may be important in the induction of cytokines such as IL-8 and TGF-beta1 relevant to allergic airway inflammation. PMID- 10696067 TI - Pulmonary hypoplasia in the myogenin null mouse embryo. AB - Although fetal breathing movements are required for normal lung development, there is uncertainty concerning the specific effect of absent fetal breathing movements on pulmonary cell maturation. We set out to evaluate pulmonary development in a genetically defined mouse model, the myogenin null mouse, in which there is a lack of normal skeletal muscle fibers and thus skeletal muscle movements are absent in utero. Significant decreases were observed in lung:body weight ratio and lung total DNA at embryonic days (E)14, E17, and E20. Reverse transcriptase/polymerase chain reaction, in situ immunofluorescence, and electron microscopy revealed early lung cell differentiation in both null and wild-type lungs as early as E14. However at E14, myogenin null lungs had decreased 5'-bromo 2-deoxyuridine incorporation compared with that of wild-type littermates, whereas at E17 and E20, increased Bax immunolabeling and terminal deoxyribonucleotidyl transferase-mediated dUTP-biotin nick-end labeling staining were detected in the myogenin null mice but not in the wild-type littermates. These observations highlight the importance of skeletal muscle contractile activity in utero for normal lung organogenesis. Null mice lacking the muscle-specific transcription factor myogenin exhibit a secondary effect on lung development such that decreased lung cell proliferation and increased programmed cell death are associated with lung hypoplasia. PMID- 10696068 TI - Nonproteolytic role for the urokinase receptor in cellular migration in vivo. AB - The urokinase receptor (uPAR) binds and localizes urokinase activity at cellular surfaces, facilitating fibrinolysis and cellular migration at sites of tissue injury. uPAR also participates in cellular signaling and regulates integrin dependent adhesion and migration in vitro. We now report evidence that uPAR occupancy regulates cellular migration in vivo in the absence of functional urokinase. Recombinant murine KC (1.5 microg), a potent neutrophil chemoattractant, was delivered to the lungs of wild-type, urokinase-deficient or uPAR-deficient mice 18 h after intraperitoneal injection of 200 microg human immunoglobulin G (IgG) or a fusion protein composed of an amino-terminal receptor binding fragment of urokinase and a human IgG Fc fragment (GFD-Fc). Whole lung lavage for recovery of leukocytes was performed 4 h later. KC treatment resulted in a 100-fold increase in lavage neutrophils. GFD-Fc injection resulted in >50% reduction in neutrophil influx in both wild-type and urokinase-deficient animals but had no effect on uPAR -/- mice. A concomitant reduction in alveolar protein leakage but no change in numbers of circulating neutrophils accompanied this attenuated inflammatory response. The reduction in neutrophil influx induced by GFD-Fc is thus related to uPAR occupancy and yet not due to disruption of uPAR mediated proteolysis. These observations verify that protease-independent functions of uPAR operate in vivo and identify uPAR as a potential target for regulation of inflammatory processes characterized by neutrophil-mediated injury. PMID- 10696069 TI - Modulation of angiotensin II receptor expression during development and regression of hypoxic pulmonary hypertension. AB - Lung vessel muscularization during hypoxic pulmonary hypertension is associated with local renin-angiotensin system activation. The expression of angiotensin II (Ang II) AT1 and AT2 receptors in this setting is not well known and has never been investigated during normoxia recovery. We determined both chronic hypoxia and normoxia recovery patterns of AT1 and AT2 expression and distal muscularization in the same lungs using in situ binding, reverse transcriptase/polymerase chain reaction, and histology. We also used an isolated perfused lung system to evaluate the vasotonic effects of AT1 and AT2 during chronic exposure to hypoxia with and without subsequent normoxia recovery. Hypoxia produced right ventricular hypertrophy of about 100% after 3 wk, which reversed with normoxia recovery. Hypoxia for 2 wk was associated with simultaneous increases (P<0.05) in AT1 and AT2 binding (16-fold and 18-fold, respectively) and in muscularized vessels in alveolar ducts (2. 8-fold) and walls (3.7-fold). An increase in AT2 messenger RNA (mRNA) (P<0.05) was also observed, whereas AT1 mRNA remained unchanged. After 3 wk of hypoxia, muscularization was at its peak, whereas all receptors and transcripts showed decreases (P<0.05 versus hypoxia 2 wk for AT1 mRNA), which became significant after 1 wk of normoxia recovery (P<0.05 versus hypoxia 2 wk). Significant reversal of muscularization (P<0.01) was found only after 3 wk of normoxia recovery in alveolar wall vessels. Finally, the AT1 antagonist losartan completely inhibited the vasopressor effect of Ang II in hypoxic and normoxia-restored lungs, whereas the AT2 agonist CGP42112A had no effect. Our data indicate that in lungs, chronic hypoxia-induced distal muscularization is associated with early and transient increases in AT2 and AT1 receptors probably owing to hypoxia- dependent transcriptional and post-transcriptional regulatory mechanisms, respectively. They also indicate that the vasotonic response to Ang II is mainly due to the AT1 subtype. PMID- 10696070 TI - Pulmonary immune responses during primary mycobacterium bovis- Calmette-Guerin bacillus infection in C57Bl/6 mice. AB - Mechanisms of protective immunity to mycobacterial infection in the lung remain poorly defined. In this study, T-cell subset expansion and cytokine expression in bronchoalveolar spaces, lung parenchyma, and mediastinal lymph nodes of mice infected intratracheally with Mycobacterium bovis-Calmette-Guerin bacillus (BCG) were analyzed in parallel with histopathology and bacterial burden. M. bovis-BCG was cleared rapidly from bronchoalveolar spaces without evidence for persistence. In lung parenchyma bacteria grew during the first 4 wk followed by gradual clearance with less than 0.1% of the original inoculum persisting for more than 8 mo. Clearance of M. bovis-BCG from bronchoalveolar lavage was associated with recruitment of both neutrophils and lymphocytes. Lung CD4(+), CD8(+), and gammadelta T-cell receptor-positive T cells expanded maximally by Week 4, and declined by Week 8 to control values despite bacterial persistence. Both CD4(+) and CD8(+) lung T cells produced interferon (IFN)-gamma in response to M. bovis BCG. Four distinct pathologic states of lung parenchymal infection were noted. Early focal sub-bronchial inflammation with transmigration of cells into airways was followed by diffuse peribronchitis, perivasculitis, and alveolitis with activated macrophages, lymphoblasts, and occasional giant cells. The latter stage corresponded to maximal M. bovis-BCG growth. Resolving infection consisted of small lymphocytes and foamy macrophages, which coincided with decreasing M. bovis BCG colony-forming units, T-cell infiltration, and IFN-gamma expression. A final quiescent phase consisted of residual lymphoid aggregates and perivasculitis associated with persistent spontaneous IFN-gamma production. Bacterial dissemination to lymph node and spleen occurred by Week 4 and declined in parallel to lung. In contrast to lung, IFN-gamma secretion was detected only late despite early expansion of CD4(+) and CD8(+) T cells. By reverse transcriptase/polymerase chain reaction, IFN-gamma and interleukin (IL)-12 p40 messenger RNA (mRNA) in lung paralleled IFN-gamma protein production. Tumor necrosis factor-alpha, IL-4 and IL-10 mRNA expression was not increased during M. bovis-BCG lung infection. Thus, protective immunity to M. bovis-BCG in the lung evolved differently in air space, lung, and lymph node. PMID- 10696071 TI - B-Cell epitope mapping of DNA topoisomerase I defines epitopes strongly associated with pulmonary fibrosis in systemic sclerosis. AB - We hypothesized that B-cell epitope mapping of DNA Topoisomerase I (type-I topoisomerase, or Topo I) may define epitopes strongly associated with pulmonary interstitial fibrosis (PIF) in systemic sclerosis (SSc). B-cell epitope mapping of Topo I was performed using 63 20-mer peptides overlapping by eight residues and spanning the entire length of the Topo I sequence. These peptides, coupled to polystyrene pins, were tested for antibody binding by enzyme-linked immunosorbent assays (ELISAs) using immunoglobulin G fractions from anti-Topo I, anticentromere, anti-U3RNP-positive, and normal sera. Four major epitopes were recognized by anti-Topo I sera, but not from the control sera: WWEEERYPEGIKWKFLEHKG (205-224, epitope I), RIANFKIEPPGLFRGRGNHP (349-368, epitope II), PGHKWKEVRHDNKVTWLVSW (397-416, epitope III), and ELDGQEYVVEFDFLGKDSIR (517 536, epitope IV). Peptide-epitopes were then synthesized in their soluble forms and ELISA systems were developed. Epitopes II to IV are localized at highly exposed sites of the Topo I tertiary structure, whereas epitope I is localized at a less accessible site. In a cohort of 81 patients with SSc with clinical data on the evolution of their disease, patients with antibodies in their sera recognizing at least three of the four epitopes had 3.1 times (P = 0.02) the hazard of developing PIF compared with patients whose sera recognized no epitopes or only one or two of the four epitopes. The discrimination was much stronger than that achieved by the simple determination of Topo I antibodies by counterimmunoelectrophoresis and immunoblot (hazard ratio 1.7, P = 0.30) in the same patients. B-cell epitope mapping of the anti-Topo I response has identified four major epitopes which cumulatively show a strong association with the development of PIF in SSc. PMID- 10696072 TI - Expression of betaig-h3 by human bronchial smooth muscle cells: localization To the extracellular matrix and nucleus. AB - Bronchial smooth muscle cells play a central role in normal lung physiology by controlling airway tone. In addition, airway smooth muscle hyperplasia and hypertrophy are important factors in the pathophysiology of asthma. In this study, expression of betaig-h3, a recently identified component of the extracellular matrix (ECM), was investigated in primary human bronchial smooth muscle (HBSM) cells. Northern blot analysis demonstrated that treatment of cultured HBSM cells with transforming growth factor-beta1 resulted in a 4- to 5 fold increase in the steady-state level of betaig-h3 messenger RNA. Western blot analysis of secreted proteins using monospecific antibodies generated against peptide sequences found in the N- and C-terminal regions of the protein identified several isoforms having apparent mass of 70-74 kD. Immunohistochemical analysis of human lung localized betaig-h3 to the vascular and airway ECM, and particularly to the septal tips of alveolar ducts and alveoli, suggesting that it may have a morphogenetic role. Analysis of cultured HBSM cells identified betaig h3 in both the ECM as well as the cytoplasm, and surprisingly also in the nucleus. These results demonstrate that betaig-h3 is produced by resident lung cells, is a component of lung ECM, and may play an important role in lung structure and function. The presence of this protein in nuclei suggests that it may have regulatory functions in addition to its role as a structural component of lung ECM. PMID- 10696073 TI - Induction and regulation of xenobiotic-metabolizing cytochrome P450s in the human A549 lung adenocarcinoma cell line. AB - Several cytochrome P450 (CYP) enzymes are expressed in the human lung, where they participate in metabolic inactivation and activation of numerous exogenous and endogenous compounds. In this study, the expression pattern of all known xenobiotic-metabolizing CYP genes was characterized in the human alveolar type II cell-derived A549 adenocarcinoma cell line using qualitative reverse transcriptase/polymerase chain reaction (RT-PCR). In addition, the mechanisms of induction by chemicals of members in the CYP1 and CYP3A subfamilies were assessed by quantitative RT-PCR. The expression of messenger RNAs (mRNAs) of CYPs 1A1, 1B1, 2B6, 2C, 2E1, 3A5, and 3A7 was detected in the A549 cells. The amounts of mRNAs of CYPs 1A2, 2A6, 2A7, 2A13, 2F1, 3A4, and 4B1 were below the limit of detection. 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induced CYP1A1 and CYP1B1 mRNAs 56-fold and 2.5-fold, respectively. CYP3A5 was induced 8-fold by dexamethasone and 11-fold by phenobarbital. CYP3A4 was not induced by any of the typical CYP3A4 inducers used. The tyrosine kinase inhibitor genistein and the protein kinase C inhibitor staurosporine blocked TCDD-elicited induction of CYP1A1, but they did not affect CYP1B1 induction. Protein phosphatase inhibitors okadaic acid and calyculin A enhanced TCDD-induction of CYP1B1 slightly, but had negligible effects on CYP1A1 induction. These results suggest that CYP1A1 and CYP1B1 are differentially regulated in human pulmonary epithelial cells and give the first indication of the induction of CYP3A5 by glucocorticoids in human lung cells. These results establish that having retained several characteristics of human lung epithelial cell CYP expression, the A549 lung cell line is a valuable model for mechanistic studies on induction of the pulmonary CYP system. PMID- 10696074 TI - Aquaporin-5 expression, but not other peripheral lung marker genes, is reduced in PTH/PTHrP receptor null mutant fetal mice. AB - Parathyroid hormone-related peptide (PTHrP) and the parathyroid hormone/parathyroid hormone-related peptide (PTH/PTHrP) receptor are important developmental regulators of cell growth and differentiation in some organs. In lung, both the peptide and the receptor are expressed early in development and in alveolar cells in adults. In adult alveolar cells, PTHrP appears to promote the alveolar type II cell phenotype in vitro. Mice carrying null mutations in genes for either receptor or ligand die at birth of respiratory failure. To determine if absence of the PTH/PTHrP receptor alters morphogenesis or cellular differentiation of the distal lung, we analyzed the morphology and gene expression patterns in PTH/PTHrP receptor null mutant mice right before birth and compared them with wild-type and heterozygous null littermates. Using semiquantitative Northern blots, we observed that messenger RNA (mRNA) for aquaporin-5, the type I cell-specific water channel, was markedly decreased. The abundance of other marker mRNAs for type I and type II cell phenotypes, including T1alpha, surfactant proteins, and others, was unaltered. Gross morphology and lung pattern, assessed by in situ hybridization for surfactant protein C, were normal. We conclude therefore that, although signaling through this receptor may influence expression of specific lung genes, it does not play a major role in the general regulation of lung development and growth. PMID- 10696075 TI - Effects of methacholine and uridine 5'-triphosphate on tracheal mucus rheology in mice. AB - We compared the action of methacholine (MCh) and uridine 5'-triphosphate (UTP) with and without pretreatment with the chloride channel blocker 4,4' diisothiocyano-2,2'-stilbenedisulfonate (DIDS) on the transepithelial potential difference (PD), the mucus collection rate (MCR), and tracheal mucus rheology using anesthetized C57BL/6 mice. The cystic fibrosis transmembrane conductance regulator (CFTR) blocker 5-nitro-2-(3-phenylpropylamino)benzoate (NPPB) was also used as a pretreatment for MCh. After collecting baseline mucus for 1.5 h, mucus secretion was stimulated by instilling 5 microl of 10(-2) M MCh or UTP around the upper trachea. There was a significant increase in PD after MCh or UTP stimulation (-21.3+/-2.0 mV MCh versus -14.1+/-1.6 mV control; -25.4+/-2.5 mV UTP versus -19.2+/-1.9 mV control). When UTP administration was preceded by DIDS, PD shifted from -15.2+/-2.9 to -12.0+/-2.2 mV. When MCh was preceded by DIDS or by NPPB, there was no change in PD. There was a significant decrease in mucus rigidity index, logG*, with MCh (2.54+/-0.09 versus 2.99+/-0.14 for control), similar to that previously reported in other species. With UTP, 14 of 16 mice responded in terms of PD becoming more negative, and of these, there was a significant difference in logG* after UTP administration (2.29 +/-0.10 versus 2.57+/-0.10 for control), whereas there was no change in logG* with DIDS administration before UTP. When DIDS administration preceded MCh, there was a diminished but still significant decrease in logG* from control, whereas there was no change in logG* when NPPB was preadministered. The control mucus collection rate was 0.19+/-0.09 mg/h, whereas after MCh stimulation, it increased to 2.83+/-0.78 mg/h. No significant difference was measured in the MCR after either UTP or DIDS+UTP stimulation. DIDS+MCh and NPPB+MCh both resulted in significant increases in MCR, but of a much smaller magnitude than that for MCh alone. We conclude that hypersecretion owing to UTP in C57BL/6 mice is less vigorous than with MCh, reflecting the limited population of Ca(2+)-dependent Cl( ) channels stimulated by UTP P(2) receptors. The action of MCh on tracheal mucus secretion in mice appears to involve both CFTR- and non-CFTR-dependent chloride channels. PMID- 10696076 TI - Pulmonary surfactant metabolism in infants lacking surfactant protein B. AB - Infants with inherited deficiency of pulmonary surfactant protein (SP) B develop respiratory failure at birth and die without lung transplantation. We examined aspects of surfactant metabolism in lung tissue and lavage fluid acquired at transplantation or postmortem from ten infants born at term with inherited deficiency of SP-B; comparison groups were infants with other forms of chronic lung disease (CLD) and normal infants. In pulse/chase labeling studies with cultured deficient tissue, no immunoprecipitable SP-B was observed and an approximately 6-kD form of SP-C accumulated that was only transiently present in CLD tissue. SP-B messenger RNA (mRNA) was approximately 8% of normal in deficient specimens, and some intact message was observed after, but not before, explant culture. Transcription rates for SP-B, assessed by nuclear run-on assay using probes for sequences both 5' and 3' of the common nonsense mutation (121ins2), were comparable in all lungs examined. The minimal surface tension achieved with lavage surfactant was similarly elevated in both deficient and CLD infants (26-31 mN/m) compared with normal infants (6 mN/m). Both SP-B-deficient and CLD infants had markedly decreased phosphatidylglycerol content of lavage and tissue compared with normal lung, whereas synthetic rates for phospholipids, including phosphatidylglycerol, were normal. We conclude that the mutated SP-B gene is transcribed normally but produces an unstable mRNA and that absence of SP-B protein blocks processing of SP-C. Chronic infant lung disease, of various etiologies, reduces surfactant function and apparently alters phosphatidylglycerol degradation. PMID- 10696078 TI - Release of GM-CSF and G-CSF by human arterial and venous smooth muscle cells: differential regulation by COX-2. AB - In addition to their traditional contractile function, vascular smooth muscle cells can be stimulated under inflammatory conditions to release a range of potent biological mediators. Indeed, we and others have shown that human vascular smooth muscle release the colony stimulating factors (CSF) granulocyte macrophage CSF (GM-CSF) and granulocyte-CSF (G-CSF) as well as large amounts of prostaglandins following the induction of cyclo-oxygenase-2 (COX-2), when stimulated with cytokines. Here we demonstrate, for the first time, that co induced COX-2 activity simultaneously suppresses GM-CSF release and potentiates G CSF release by human vascular cells. Moreover, the differential regulation of GM CSF and G-CSF release by COX-2 was mimicked by the prostacyclin (PGI(2)) mimetic, cicaprost. These observations suggest that PGI(2), released following the induction of COX-2, differentially regulates the release of GM-CSF (suppresses) and G-CSF (potentiates) from human vascular cells. PMID- 10696077 TI - Peroxisome proliferator-activated receptors in the cardiovascular system. AB - Peroxisome proliferator-activated receptor (PPAR)s are a family of three nuclear hormone receptors, PPARalpha, -delta, and -gamma, which are members of the steriod receptor superfamily. The first member of the family (PPARalpha) was originally discovered as the mediator by which a number of xenobiotic drugs cause peroxisome proliferation in the liver. Defined functions for all these receptors, until recently, mainly concerned their ability to regulate energy balance, with PPARalpha being involved in beta-oxidation pathways, and PPARgamma in the differentiation of adipocytes. Little is known about the functions of PPARdelta, though it is the most ubiquitously expressed. Since their discovery, PPARs have been shown to be expressed in monocytes/macrophages, the heart, vascular smooth muscle cells, endothelial cells, and in atherosclerotic lesions. Furthermore, PPARs can be activated by a vast number of compounds including synthetic drugs, of the clofibrate, and anti-diabetic thiazoldinedione classes, polyunsaturated fatty acids, and a number of eicosanoids, including prostaglandins, lipoxygenase products, and oxidized low density lipoprotein. This review will aim to introduce the field of PPAR nuclear hormone receptors, and discuss the discovery and actions of PPARs in the cardiovascular system, as well as the source of potential ligands. PMID- 10696079 TI - Inhibition of the hypothalamic-pituitary-adrenal axis in food-deprived rats by a CCK-A receptor antagonist. AB - The circadian activity of the hypothalamic-pituitary-adrenal (HPA) axis is regulated by caloric flow in rats. During the dark cycle, it has been shown that, in fasted rats, the time-course profile of plasma concentrations of adrenocorticotropin (ACTH) and corticosterone parallels the profile of food intake in ad libitum fed animals. Cholecystokinin (CCK) is involved in regulating food intake in rodents. CCK-8 reduces food intake by acting on CCK-A receptors subtype. This work aims at establishing an eventual relationship between the modulatory role of CCK on food intake and its effect on HPA axis activity during fasting. We studied the effect of CCK-A and CCK-B receptor antagonists on food intake during the first period of the dark cycle. Under these conditions we observed that the CCK-A receptor antagonist, SR-27897 (0.3 mg kg(-1)), but not the CCK-B receptor antagonist, L-365260 (1 mg kg(-1)), increases food-intake. In a second series of experiments we observed that the increase of both ACTH and corticosterone plasma level elicited by fasting, was prevented by SR-27897, but not by L-365260. These results indicate that CCK-A receptor blockade during fasting prevents the activation of the HPA axis. PMID- 10696080 TI - Cardiotoxic effects of fenfluramine hydrochloride on isolated cardiac preparations and ventricular myocytes of guinea-pigs. AB - The cardiotoxic effects of fenfluramine hydrochloride on mechanical and electrical activity were studied in papillary muscles, Purkinje fibres, left atria and ventricular myocytes of guinea-pigs. Force of contraction (f(c)) was measured isometrically, action potentials and maximum rate of rise of the action potential (V(max)) were recorded by means of the intracellular microelectrode technique and the sodium current (I(Na)) with patch-clamp technique in the cell attached mode. For kinetic analysis (S)-DPI-201-106-modified Na(+) channels from isolated guinea-pig ventricular heart cells were used. Fenfluramine (1 - 300 microM) produced negative chronotropic and inotropic effects; additional extracellular Ca(2+) competitively antagonized the negative inotropic effect. Fenfluramine concentration-dependently reduced V(max) and showed tonic blockade of sodium channels, shortened the action potential duration in papillary muscles and Purkinje fibres. In cell-attached patches, fenfluramine decreased I(Na) concentration-dependently (10 - 100 microM), frequency-independently (0.1 - 3 Hz; 30 microM). The h(infinity) curve was shifted towards hyperpolarizing direction. At 30 microM, fenfluramine blocked the sodium channel at all test potentials to the same degree, and neither changed the threshold and reversal potentials nor the peak of the curve. No effect on single channel availability, but a significant decrease in mean open times and increase in mean closed times was observed. Mean duration of the bursts decreased and number of openings per record increased with increasing drug concentration. It is concluded that the effect on I(Na) plays an important role in the cardiotoxicity of fenfluramine in addition to primary pulmonary hypertension and valvular disorders. PMID- 10696081 TI - In vivo venodilator action of fenoldopam, a dopamine D(1)-receptor agonist. AB - The effects of the dopamine D(1)-receptor agonist fenoldopam were compared with those of the D(2)-receptor agonist R(-)-propylnorapomorphine and vehicle on mean arterial pressure (MAP), mean circulatory filling pressure (MCFP, the driving force of venous return), arterial resistance (R(a)), venous resistance (R(v)), heart rate (HR) and cardiac output (CO) in groups of thiobutabarbitone anaesthetized rats pre-treated with i.v. injection of mecamylamine (3.7 micromol kg(-1)) and continuously infused with noradrenaline (6.8 nmol kg(-1) min(-1)). The vehicle did not alter any haemodynamic variables. All doses of fenoldopam (0.5, 2 and 16 microgram kg(-1) min(-1)) reduced MAP, R(a) and R(v), and increased CO. At the highest dose, fenoldopam also increased HR and reduced MCFP. All doses of R(-)-propylnorapomorphine (0.5, 2 and 16 microgram kg(-1) min(-1)) increased MAP but did not significantly alter CO, R(v) and MCFP. Both R(a) and HR were increased by the highest dose of R(-)-propylnorapomorphine. Our results indicate that fenoldopam reduces MAP and MCFP, and markedly increases CO through reductions of arterial and venous resistances. The effects of fenoldopam in dilating arterial resistance and capacitance vessels were similar. In contrast, R(-)-propylnorapomorphine elevates MAP through an increase in arterial resistance but has minimal effects on CO, MCFP and venous resistance. Both drugs have a small direct, positive chronotropic action at the highest dose. PMID- 10696082 TI - Pressor response to pulsatile compression of the rostral ventrolateral medulla mediated by nitric oxide and c-fos expression. AB - It has been reported that neurovascular compression of the rostral ventrolateral medulla might be causally related to essential hypertension. Recently, we found that pulsatile compression of the rostral ventrolateral medulla increases sympathetic nerve activity and elevates arterial pressure via activation of glutamate receptors in rats. We also found that increases in sympathetic and cardiovascular activities by microinjection of L-glutamate into the rostral ventrolateral medulla are mediated by c-fos expression-related substance(s) following activation of the nitric oxide-cyclic GMP pathway. Herein, we investigated whether responses to pulsatile compression are mediated by local activation of the nitric oxide-cyclic GMP pathway and/or c-fos expression-related substance(s) in rats. Increases in arterial pressure (15+/-1 mmHg), heart rate (9+/-1 b.p.m.), and sympathetic nerve activity (% change: 8.5+/-1.1%) induced by pulsatile compression were partially but significantly inhibited after local microinjection of a nitric oxide synthase inhibitor, L-N(G)-nitroarginine methyl ester (8+/-2 mmHg, 1+/-1 b.p.m., 4.0+/-1.3%; P<0.05 vs compression without pretreatment) or 7-nitroindazole (7+/-2 mmHg, 2+/-1 b.p.m., 4.0+/-1. 5%; P<0.05), or a soluble guanylate cyclase inhibitor, methylene blue (9+/-1 mmHg, 4+/-1 b.p.m., 4.1+/-1.4%; P<0.05). In addition, increases in arterial pressure, heart rate, and sympathetic nerve activity by pulsatile compression were significantly reduced 6 h after microinjection of antisense oligodeoxynucleotide to c-fos mRNA (2+/-2 mmHg, 2+/-1 b.p.m., 1.0+/-1.0%; P<0.05 vs sense oligodeoxynucleotide). These results suggest that increases in sympathetic and cardiovascular activities induced by pulsatile compression of the rostral ventrolateral medulla are mediated, at least in part, by local activation of the nitric oxide-cyclic GMP pathway and c-fos expression-related substance(s) in rats. PMID- 10696083 TI - Dexamethasone attenuates the depressor response induced by neuropeptide Y microinjected into the nucleus tractus solitarius in rats. AB - An investigation was made of the effect of dexamethasone (Dex) injection into the nucleus tractus solitarius (NTS) on the cardiovascular response to neuropeptide Y in rats. Dex (39 pmol) injected into the NTS inhibited the hypotension and bradycardia caused by NPY (5 pmol) with a short latency (10 min) and a long duration of action (up to 4 h). The rapid inhibition by Dex (39 pmol) of the cardiovascular response to NPY was not blocked by pretreatment with the glucocorticoid receptor blocker, RU38486 (47 or 117 pmol respectively), but was reversed by bicuculline (30 pmol). Microiontophoresis of NPY (0.01 mM, pH 6.5) into the NTS increased the spontaneous firing of the majority (68.4%) of baroreflex-excited cells, but decreased the firing of most (73.7%) baroreflex inhibited cells. In contrast, Dex (0.02 M, pH 6.5) decreased the spontaneous firing of the majority of baroreflex-excited cells (42.1% of normal response) and decreased the inhibition of baroreflex-inhibited cells (47.5% of normal response). The responses of the majority of baroreceptive cells to NPY were blocked by iontophoretic administration of Dex. Dex (200 microM) increased the delayed rectifier outward K+ current by 31.4+/-1.1% (n=5), whereas NPY alone, at a concentration of 1.5 microM, inhibited the current by 28.6+/-0.8% (n=5). In the presence of Dex (200 microM), addition of NPY (1.5 microM) had no effect on the current. In conclusion, NTS-administered-Dex attenuated the cardiovascular response to NPY injected into the same area via a rapid membrane effect, which was mediated by an action on GABA(A) receptors and on the delayed rectifier outward K(+) channel. PMID- 10696084 TI - Molecular identification and pharmacological characterization of adenosine receptors in the guinea-pig colon. AB - The aim of this study is to elucidate the role of adenosine in the motor function of the guinea-pig distal colon.2 To determine whether adenosine A(1) receptors and A(2B) receptors are expressed in the guinea-pig colon, we employed the reverse transcription-polymerase chain reaction (RT - PCR). The gene expression of A(1) receptor and A(2B) receptor was found for the first time in the guinea pig proximal and distal colon.3 Adenosine A(1) agonist N(6)-cyclopentyladenosine (CPA), and A(1)/A(2) agonist 5'-N-ethylcarboxamidoadenosine (NECA) concentration dependently inhibited neurogenic responses to electrical field stimulation (EC(50)=1.07x10(-8) and 2.12x10(-8) M) in the longitudinal muscle, but A(2A) agonist 2-p-(2-carboxyethyl)phenylethylamino-5'-N-ethycarboxamido-ad enosine (CGS21680) had only a slight inhibitory effect (25.9%, 1 microM). A(1) antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX, 10 nM: A(1) selective concentration) antagonized responses to CPA and NECA. Furthermore, the affinity order of antagonists at inhibiting the effect NECA was: DPCPX>8-phenyltheophylline (8-PT: A(1)/A(2) antagonist).3 In the presence of tetrodotoxin (TTX, 0.3 microM), CPA and NECA relaxed myogenic precontraction induced by KCl (50 mM) (EC(50)=1.26x10( 5) and 1.04x10(-5) M, respectively), but CGS21680 (1 microM) did not cause any relaxation. DPCPX did not affect responses to CPA and NECA at a concentration of 10 nM, but a higher concentration (1 microM) of DPCPX and 10 microM of 8-PT antagonized those responses.5 These data lead us to the hypothesis that adenosine may mediate relaxation through two different inhibitory receptor subtypes; A(1) receptors on the enteric neuron and A(2B) receptor on the smooth muscle in the guinea-pig distal colon. PMID- 10696085 TI - Pharmacological characterization of human recombinant melatonin mt(1) and MT(2) receptors. AB - We have pharmacologically characterized recombinant human mt(1) and MT(2) receptors, stably expressed in Chinese hamster ovary cells (CHO-mt(1) and CHO MT(2)), by measurement of [(3)H]-melatonin binding and forskolin-stimulated cyclic AMP (cAMP) production. [3H]-melatonin bound to mt(1) and MT(2) receptors with pK(D) values of 9.89 and 9.56 and B(max) values of 1.20 and 0.82 pmol mg(-1) protein, respectively. Whilst most melatonin receptor agonists had similar affinities for mt(1) and MT(2) receptors, a number of putative antagonists had substantially higher affinities for MT(2) receptors, including luzindole (11 fold), GR128107 (23 fold) and 4-P-PDOT (61 fold). In both CHO-mt(1) and CHO-MT(2) cells, melatonin inhibited forskolin-stimulated accumulation of cyclic AMP in a concentration-dependent manner (pIC(50) 9.53 and 9.74, respectively) causing 83 and 64% inhibition of cyclic AMP production at 100 nM, respectively. The potencies of a range of melatonin receptor agonists were determined. At MT(2) receptors, melatonin, 2-iodomelatonin and 6-chloromelatonin were essentially equipotent, whilst at the mt(1) receptor these agonists gave the rank order of potency of 2-iodomelatonin>melatonin>6-chloromelatonin. In both CHO-mt(1) and CHO MT(2) cells, melatonin-induced inhibition of forskolin-stimulated cyclic AMP production was antagonized in a concentration-dependent manner by the melatonin receptor antagonist luzindole, with pA(2) values of 5.75 and 7.64, respectively. Melatonin-mediated responses were abolished by pre-treatment of cells with pertussis toxin, consistent with activation of G(i)/G(o) G-proteins. This is the first report of the use of [(3)H]-melatonin for the characterization of recombinant mt(1) and MT(2) receptors. Our results demonstrate that these receptor subtypes have distinct pharmacological profiles. PMID- 10696086 TI - Exposure to glibenclamide increases rat beta cells sensitivity to glucose. AB - An increased sensitivity to glucose was observed in islets pre-exposed for 1 h to glibenclamide (0.1 micromol 1(-1)), but not to tolbutamide (100 micromol l(-1)), as indicated by a shift to the left of the dose-response curve (EC(50) at 5.8+/ 0.3 mmol l(-1) glucose vs 10.6+/-0.8 in control islets; n=11, P<0.005). According to this secretory pattern also glucose utilization at 2.5 and 5.0 mmol l(-1) glucose was higher in islets exposed to glibenclamide. Since binding to mitochondria results in an increased enzyme activity, we measured hexokinase (HK) and glucokinase (GK) activity both in a cytosolic and in a mitochondrion-enriched fractions. Cytosolic hexokinase activity was similar in islets exposed to glibenclamide and in control islets but mitochondrial hexokinase activity was significantly increased after exposure to glibenclamide (124+/-7 vs 51+/-9 nmol microgram prot(-1) 90 min(-1), P<0.01), with no change in the enzyme protein content. In contrast, glucokinase activity in the two groups of islets was similar. When in islets < exposed to glibenclamide hexokinase binding to mitochondria was inhibited by the addition of 20 nmol l(-1) dicyclohexylcarbodiimide (DCC), no increase of glucose sensitivity was observed (EC(50) 10.9+/-1.3 mmol l(-1) glucose, n=3, similar to that of control islets). These data indicate that a 1 h exposure to glibenclamide causes the beta cell to become more sensitive to glucose. This increased sensitivity is associated with (and may be due to) an increased hexokinase activity, in particular the mitochondrial-bound, more active, form. This mechanism may contribute to the hypoglycemic action of this drug. PMID- 10696087 TI - Electrophysiological effects of protopine in cardiac myocytes: inhibition of multiple cation channel currents. AB - Protopine (Pro) from Corydalis tubers has been shown to have multiple actions on cardiovascular system, including anti-arrhythmic, anti-hypertensive and negative inotropic effects. Although it was thought that Pro exerts its actions through blocking Ca(2+) currents, the electrophysiological profile of Pro is unclear. The aim of this study is to elucidate the ionic mechanisms of Pro effects in the heart. In single isolated ventricular myocytes from guinea-pig, extracellular application of Pro markedly and reversibly abbreviates action potential duration, and decreases the rate of upstroke (dV/dt)(max), amplitude and overshoot of action potential in a dose-dependent manner. Additionally, it produces a slight, but significant hyperpolarization of the resting membrane potential. Pro at 25, 50 and 100 microM reduces L-type Ca(2+) current (I(Ca,L)) amplitude to 89.1, 61.9 and 45.8% of control, respectively, and significantly slows the decay kinetics of I(Ca,L) at higher concentration. The steady state inactivation of I(Ca,L) is shifted negatively by 5.9 - 7.0 mV (at 50 - 100 microM Pro), whereas the voltage dependent activation of I(Ca,L) remains unchanged. In contrast, Pro at 100 microM has no evident effects on T-type Ca(2+) current (I(Ca,T)). In the presence of Pro, both the inward rectifier (I(K1)) and delayed rectifier (I(K)) potassium currents are variably inhibited, depending on Pro concentrations. Sodium current (I(Na)), recorded in low [Na(+)](o) (40 mM) solution, is more potently suppressed by Pro. At 25 microM, Pro significantly attenuated I(Na) at most of the test voltages (-60 approximately +40 mV, with a 53% reduction at -30 mV. Thus, Pro is not a selective Ca(2+) channel antagonist. Rather, it acts as a promiscuous inhibitor of cation channel currents including I(Ca,L), I(K), I(K1) as well as I(Na). These findings may provide some mechanistic explanations for the therapeutic actions of Pro in the heart. PMID- 10696088 TI - The insulinotropic mechanism of the novel hypoglycaemic agent JTT-608: direct enhancement of Ca(2+) efficacy and increase of Ca(2+) influx by phosphodiesterase inhibition. AB - We examined the effects of the novel hypoglycaemic agent JTT-608 [trans-4-(4 methylcyclohexyl)-4-oxobutyric acid] on insulin secretion using rat pancreatic islets, and analysed the mechanism of its effect. JTT-608 augmented 8.3 mM glucose-induced insulin secretion dose-dependently, and there was a stimulatory effect of 100 microM JTT-608 at both moderate and high concentrations (8.3, 11. 1 and 16.7 mM) of glucose, but not at low concentrations (3.3 and 5. 5 mM). In perifusion experiments, both phases of insulin release were enhanced, and the effect was eliminated 10 min after withdrawal of the agent. In the presence of 200 microM diazoxide and a depolarizing concentration (30 mM) of K(+), there was an augmentation of insulin secretion by 100 microM JTT-608, not only under high levels of glucose but also under low levels, and the effects were abolished by 10 microM nitrendipine. JTT-608 augmented insulin secretion from electrically permeabilized islets in the presence of stimulatory concentrations (0.3 and 1.0 microM) of Ca(2+), and the intracellular Ca(2+) concentration ([Ca(2+)](i)) response under 16.7 mM glucose, 200 microM diazoxide, and 30 mM K(+) was also increased. The cyclic AMP content in the islets was increased by 100 microM JTT 608, and an additive effect to 1 microM forskolin was observed, but not to 50 microM 3-isobutyl-1-methylxanthine (IBMX). JTT-608 inhibited phosphodiesterase (PDE) activity dose-dependently. We conclude that JTT-608 augments insulin secretion by enhancing Ca(2+) efficacy and by increasing Ca(2+) influx. This appears to be a result of the increased intracellular cyclic AMP concentration due to PDE inhibition. PMID- 10696089 TI - ATP suppression of interleukin-12 and tumour necrosis factor-alpha release from macrophages. AB - Immune cell activation releases ATP into the extracellular space. ATP-sensitive P2 purinergic receptors are expressed on immune cells and activation of these receptors alters immune cell function. Furthermore, ATP is metabolized by ectonucleotidases to adenosine, which has also been shown to alter cytokine production. In the present study, we investigated how extracellular ATP affects interleukin (IL)-12 and tumour necrosis factor (TNF)-alpha production in bacterial lipopolysaccharide (LPS)-treated murine peritoneal macrophages and we also examined whether extracellular ATP alters the production of the T helper 1 cytokine interferon (IFN)-gamma. Pretreatment of the peritoneal macrophages with ATP or various ATP analogues decreased both IL-12 and TNF-alpha production induced by LPS (10 microgram ml(-1)). The effect of ATP was partially reversed by cotreatment with adenosine deaminase (0.1 - 1 u ml(-1)), suggesting that the suppressive effect of ATP on cytokine production is, in part, due to its degradation products. Immunoneutralization with an anti-IL-10 antibody demonstrated that although ATP increases IL-10 production, the inhibition of IL 12 and TNF-alpha production is independent of the increased IL-10. The effect of ATP was pretranslational, as it suppressed steady state levels of mRNAs for IL-12 (both p35 and p40). In spleen cells stimulated with either LPS (10 microgram ml( 1)) or anti-CD3 (2 microgram ml(-1)) antibody, ATP suppressed, in a concentration dependent manner, the production of IFN-gamma. These results suggest that extracellular ATP has multiple anti-inflammatory effects and that release of ATP into the extracellular space may play a role in blunting the overactive immune response in autoimmune diseases. PMID- 10696090 TI - Role of nitric oxide and septide-insensitive NK(1) receptors in bronchoconstriction induced by aerosolised neurokinin A in guinea-pigs. AB - The tachykinin, neurokinin A (NKA), contracts guinea-pig airways both in vitro and in vivo, preferentially activating smooth muscle NK(2) receptors, although smooth muscle NK(1) receptors may also contribute. In vitro evidence suggests that NKA activates epithelial NK(1) receptors, inducing the release of nitric oxide (NO) and subsequent smooth muscle relaxation. A number of selective NK(1) receptor agonists have been reported to activate both smooth muscle and epithelial NK(1) receptors, however septide appears only to activate smooth muscle NK(1) receptors. The aim of the present study was to investigate whether NKA-induced bronchoconstriction in guinea-pigs in vivo may be limited by NO release via NK(1) receptor activation, and whether selective NK(1) receptor agonists may activate this mechanism differently. Aerosolized NKA caused an increase in total pulmonary resistance (RL) that was markedly reduced by the NK(2) receptor antagonist, SR 48968, and abolished by the combination of SR 48968 and the NK(1) receptor antagonist, CP-99, 994. The increase in RL evoked by NKA was potentiated by pretreatment with the NO synthase (NOs) inhibitor, L-NAME, but not by the inactive enantiomer D-NAME. Potentiation by L-NAME of NKA-induced increase in RL was reversed by L-Arginine, but not by D-Arginine. Pretreatment with L-NAME did not affect the increase in RL induced by the selective NK(2) receptor agonist, [beta-Ala(8)]NKA(4-10), and by the selective NK(1) receptor agonist, septide, whereas it markedly potentiated the increase in RL caused by a different NK(1) selective agonist, [Sar(9),Met(O(2))(11)]SP. Dose-response curves showed that septide was a more potent bronchoconstrictor than [Sar(9),Met(O(2))(11)]SP to cause bronchoconstriction. Pretreatment with the NK(1) receptor antagonist, CP-96,994, abolished the ability of L-NAME to increase bronchoconstriction to aerosolized NKA. Bronchoconstriction to aerosolized NKA was increased by L-NAME, after pretreatment with the NK(3) receptor antagonist, SR 142801. The present study shows that in vivo bronchoconstriction in response to the aerosolized naturally occurring tachykinin, NKA, is limited by its own ability to release relaxant NO via NK(1) receptor activation. This receptor is apparently insensitive to septide, thus justifying, at least in part, the high potency of septide to cause bronchoconstriction in guinea-pigs. PMID- 10696091 TI - Effect of shear stress on the release of soluble ecto-enzymes ATPase and 5' nucleotidase along with endogenous ATP from vascular endothelial cells. AB - Stimulation of endothelial cells from human umbilical vein by shear stress induced release of endogenous ATP which was accompanied by an extracellular increase in the activity of enzymes degrading both ATP (ATPases) and AMP (5' nucleotidases). The activity of soluble ATPase was progressively increased from 1.62+/-0.27 to 12.7+/-1.0 pmoles ml(-1) h(-1) after 60 min of stimulation by shear stress. The rate of [(3)H]-ATP hydrolysis in the medium was inhibited by the purinergic agents suramin, Reactive blue 2 and pyridoxalphosphate-6-azophenyl 2'4'-disulphonic acid, and remained insensitive to the classic inhibitors of ion pumping and intracellular ATPases. Shear stress also increased the activity of 5' nucleotidase in the medium from 2.0+/-0.5 to 27.2+/-2.8 pmoles ml(-1) h(-1). When shear stress was applied after removal of ecto-5'-nucleotidase by phosphatidylinositol-specific phospholipase C, the release of 5'-nucleotidase was drastically reduced. These results show that soluble ATPase and 5'-nucleotidase which are released during shear stress are not released from an intracellular compartment together with ATP but have an extracellular origin. PMID- 10696093 TI - Endothelium-dependent relaxation followed by contraction mediated by NK(1) receptors in precontracted rabbit intrapulmonary arteries. AB - In the present study, we examined whether substance P (SP) and SP methyl ester (SPME), a selective NK(1) agonist, cause biphasic responses consisting of endothelium-dependent relaxation (EDR) and contraction (EDC) in precontracted rabbit intrapulmonary arteries. In arteries contracted with PGF(2alpha) (2x10(-6) M), SP as well as SPME caused only EDR at low concentration (10(-9) M) and EDR followed by EDC at higher concentrations, indicating the involvement of NK(1) receptors. The SP (10(-8) M)-induced EDR was abolished in arteries moderately contracted by PGF(2alpha) (5x10(-7) M) and the EDC in arteries maximally contracted by PGF(2alpha) (10(-5) M), indicating that EDR and EDC are inversely dependent on preexisting tone. Indomethacin (10(-8) - 10(-6) M), a cyclo oxygenase inhibitor, and ozagrel (10(-8) - 10(-6) M), a TXA(2) synthetase inhibitor attenuated the EDC in the SPME (10(-7) M)-induced biphasic response and markedly potentiated the EDR. AA-861 (10(-8) - 10(-6) M), a 5-lipoxygenase inhibitor, did not affect the EDR or EDC. L-N(G)-nitro-arginine methyl ester (10( 5) - 10(-4) M), a nitric oxide synthase inhibitor, attenuated the EDR and slightly potentiated the EDC. CP-99994 (10(-10) - 10(-8) M), an NK(1) antagonist, attenuated the EDC and potentiated the EDR in the SPME (10(-7) M)-induced biphasic response, while the NK(2) antagonist SR-48968 (10(-9) - 10(-7) M) had no effect. CP-99994 attenuated the SPME (10(-7) M)-induced EDC under EDR-blockade to a greater extent than the EDR under EDC-blockade, indicating that CP-99994 enhanced the EDR component by preferential inhibition of the EDC component. In conclusion, NK(1) agonists caused a biphasic endothelium-dependent response (EDR and EDC) in submaximally precontracted intrapulmonary arteries. The EDC and EDR mediated by NK(1) receptors may play physiological and/or pathophysiological roles in modulation of vascular tone. PMID- 10696092 TI - P(2Y) purinoceptor subtypes recruit different mek activators in astrocytes. AB - Extracellular ATP can function as a glial trophic factor as well as a neuronal transmitter. In astrocytes, mitogenic signalling by ATP is mediated by metabotropic P(2Y) receptors that are linked to the extracellular signal regulated protein kinase (Erk) cascade, but the types of P(2Y) receptors expressed in astrocytes have not been defined and it is not known whether all P(2Y) receptor subtypes are coupled to Erk by identical or distinct signalling pathways. We found that the P(2Y) receptor agonists ATP, ADP, UTP and 2 methylthioATP (2MeSATP) activated Erk and its upstream activator MAP/Erk kinase (Mek). cRaf-1, the first kinase in the Erk cascade, was activated by 2MeSATP, ADP and UTP but, surprisingly, cRaf-1 was not stimulated by ATP. Furthermore, ATP did not activate B-Raf, the major isoform of Raf in the brain, nor other Mek activators such as Mek kinase 1 (MekK1) and MekK2/3. Reverse transcriptase polymerase chain reaction (RT - PCR) studies using primer pairs for cloned rat P(2Y) receptors revealed that rat cortical astrocytes express P(2Y(1)), a receptor subtype stimulated by ATP and ADP and their 2MeS analogues, as well as P(2Y(2)) and P(2Y(4)), subtypes in rats for which ATP and UTP are equipotent. Transcripts for P(2Y(6)), a pyrimidine-preferring receptor, were not detected. ATP did not increase cyclic AMP levels, suggesting that P(2Y(11)), an ATP preferring receptor, is not expressed or is not linked to adenylyl cyclase in rat cortical astrocytes. These signal transduction and RT - PCR experiments reveal differences in the activation of cRaf-1 by P(2Y) receptor agonists that are inconsistent with properties of the P(2Y(1)), P(2Y(2)) and P(2Y(4)) receptors shown to be expressed in astrocytes, i.e. ATP=UTP; ATP=2MeSATP, ADP. This suggests that the properties of the native P(2Y) receptors coupled to the Erk cascade differ from the recombinant P(2Y) receptors or that astrocytes express novel purine-preferring and pyrimidine-preferring receptors coupled to the ERK cascade. PMID- 10696094 TI - The mechanism of bradykinin-induced endothelium-dependent contraction and relaxation in the porcine interlobar renal artery. AB - The mechanism of endothelium-dependent regulation of vascular tone of bradykinin was investigated by simultaneously monitoring the changes in the cytosolic Ca(2+) concentration and the force of smooth muscle in fura-2-loaded strips of the porcine renal artery with endothelium. During phenylephrine-induced sustained contraction, bradykinin (>3x10(-9) M) caused endothelium-dependent triphasic changes in the force of the strips, composed of an initial relaxation, a subsequent transient contraction and a late sustained relaxation. At low concentrations (10(-10) - 10(-9) M), bradykinin caused an endothelium-dependent biphasic relaxation with no contraction. A thromboxane A(2) (TXA(2))/prostaglandin H(2) (PGH(2)) receptor antagonist (10(-5) M ONO-3708) completely inhibited, while a TXA(2) synthase inhibitor (10(-5) M OKY-046) only partially inhibited, the transient contraction induced by bradykinin. Under conditions where the bradykinin-induced contraction was inhibited by ONO-3708 during the phenylephrine-induced contraction, bradykinin induced only a transient relaxation in the presence of N(Omega)-nitro-L-arginine methyl ester (L-NAME). This transient relaxation was inhibited when the precontraction was initiated by phenylephrine plus 40 mM extracellular K(+). The removal of L-NAME from this condition caused a partial reappearance of the initial relaxation and a complete reappearance of the sustained relaxation. In conclusion, bradykinin caused the endothelium-dependent triphasic regulation of vascular tone in the porcine renal artery. The concentrations of bradykinin required to induce a contraction was higher than that required to induce relaxation. Both TXA(2) and PGH(2) were involved in the bradykinin-induced contraction. The initial relaxation was mediated by nitric oxide and hyperpolarizing factors while the sustained relaxation depended on nitric oxide. PMID- 10696095 TI - Oxidative stress and S-nitrosylation of proteins in cells. AB - The effect of prolonged exposure to nitric oxide on enzymes involved in cell metabolism was investigated in T lymphocyte-derived Jurkat and L929 fibroblast human cell lines using a constant concentration of nitric oxide (1.5 microM) released by the nitric oxide donor DETA-NO (0.5 mM). Nitric oxide inhibited immediately the respiration of the cells acting reversibly at complex IV. With time, the inhibition became progressively persistent, i.e. not reversed by trapping of nitric oxide with oxyhaemoglobin, and was preceded by a decrease in the concentration of the intracellular reduced glutathione. This persistent effect of nitric oxide on respiration was due to inhibition of complex I activity which could be reversed by addition of reduced glutathione or by cold light, suggesting that it was due to S-nitrosylation of thiols necessary for the activity of the enzyme. The activity of other enzymes also known to be susceptible to inhibition by S-nitrosylation, i.e. glyceraldehyde-3-phosphate dehydrogenase and glutathione reductase, was progressively decreased by exposure to nitric oxide with a similar time course to that observed for the inhibition of complex I. Furthermore, inhibition of these enzymes only occurred when the concentrations of reduced glutathione had previously fallen and could be prevented by increasing the intracellular concentrations of reduced glutathione. Our results suggest that S-nitrosylation of different enzymes by nitric oxide may occur only if the reducing potential of the cells is impaired. PMID- 10696096 TI - Endothelin-1-induced ET(A) receptor-mediated nociception, hyperalgesia and oedema in the mouse hind-paw: modulation by simultaneous ET(B) receptor activation. AB - Endothelin-1 causes ET(A) receptor-mediated enhancement of capsaicin-induced nociception in mice. We have assessed if this hyperalgesic effect of endothelin-1 is also accompanied by other pro-inflammatory effects, namely nociception and oedema, and characterized the endothelin ET receptors involved. Intraplantar (i. pl.) hind-paw injection of endothelin-1 (0.3 - 30 pmol) induced graded nociceptive responses (accumulated licking time: vehicle, 20. 5+/-3.3 s; endothelin-1 at 30 pmol, 78.1+/-9.8 s), largely confined to the first 15 min. Endothelin-1 (1 - 10 pmol) potentiated ipsilateral capsaicin-induced (0.1 microgram, i.pl.; at 30 min) nociception (vehicle, 40.2+/-2.6 s; endothelin-1 at 10 pmol, 98.4+/-5.8 s, but 30 pmol was inactive), and caused oedema (increase in paw weight 5 min after capsaicin: vehicle, 46.3+/-2.3 mg; endothelin-1 at 30 pmol, 100.3+/-6.1 mg). Selective ET(B) receptor agonists sarafotoxin S6c (up to 30 pmol) and IRL 1620 (up to 100 pmol) were inactive, whereas endothelin-3 (up to 30 pmol) induced only modest oedema. ET(A) receptor antagonists BQ-123 (1 nmol, i.pl. ) or A-127722-5 (6 micromol kg(-1), i.v.) prevented all effects of endothelin-1 (10 pmol), but the ET(B) receptor antagonist BQ-788 (1 or 10 nmol, i.pl.) was ineffective. BQ-788 (10 nmol, i.pl.) unveiled hyperalgesic effects of 30 pmol endothelin-1 and endothelin-3. Sarafotoxin S6c (30 pmol, i.pl.) did not modify endothelin-1-induced (10 pmol) nociception or oedema, but abolished hyperalgesia. Thus, endothelin-1 triggers ET(A) receptor-mediated nociception, hyperalgesia and oedema in the mouse hind-paw. Simultaneous activation of ET(B) receptors by endothelin-1 or selective agonists can limit the hyperalgesic, but not the nociceptive or oedematogenic, effects of the peptide. PMID- 10696097 TI - Nitric oxide-dependent vasodilatation of rabbit femoral artery by beta(2) adrenergic stimulation or cyclic AMP elevation in vivo. AB - Some studies suggest that beta-adrenoceptor-mediated vasorelaxation is in part mediated through nitric oxide (NO) release. We wished to determine the contribution of the L-arginine / NO system to vasodilatation in response to beta adrenoceptor stimulation with isoprenaline or cyclic adenosine-3',5' monophosphate (cyclic AMP) elevation with forskolin and dibutyryl cyclic AMP in vivo, using a rabbit femoral artery constant perfusion model. Baseline femoral artery pressure was similar in rabbits receiving isoprenaline, forskolin or dibutyryl cyclic AMP. Isoprenaline, forskolin and dibutyryl cyclic AMP each decreased femoral artery pressure in a dose-dependent manner. The doses (mol kg( 1)) of isoprenaline, forskolin and dibutyryl cyclic AMP which decreased pressure by 10% from baseline, expressed as a negative logarithm (-log ED(10)) were: 10.0+/-0.2, 9.5+/-0.1 and 4.9+/-0.1 respectively (P<0.0001 for each). Use of beta adrenoceptor subtype-selective antagonists showed that the vascular response to isoprenaline was purely due to stimulation of the beta(2)-adrenoceptor subtype. Injection of 1 micromol kg(-1) N(G)-nitro-L-arginine methyl ester (L-NAME) did not alter baseline pressure. However, it abolished the pressure response to isoprenaline (P<0.0001), and significantly attenuated the pressure responses to forskolin and dibutyryl cyclic AMP: -log ED(10) values for forskolin and dibutyryl cyclic AMP, in the presence of L-NAME, were 7.9+/-0.1 and 3.5+/-0.3 respectively (P<0.0001 for each, as compared with values in the absence of L NAME). These results indicate that beta(2)-adrenergic stimulation and cylic AMP elevation activate the L-arginine/NO system in rabbit femoral artery in vivo, and that NO generation contributes importantly to the changes in vascular tone induced by agents which modulate beta-adrenoceptors or cyclic AMP. PMID- 10696098 TI - Role for endogenous endothelin in the regulation of plasma volume and albumin escape during endotoxin shock in conscious rats. AB - To explore the role of endogenous endothelin (ET) in the regulation of vascular functions, we studied the effects endothelin receptor blockade on blood pressure, plasma volume and albumin escape during endotoxin shock in conscious, chronically catheterized rats. Red blood cell volume and plasma volume were determined by using chromium-51-tagged erythrocytes and iodine-125-labelled albumin, respectively. Intravenous injection of lipopolysaccharide (LPS, 10 mg kg(-1)) resulted in hypotension, haemoconcentration, and increased total-body albumin escape, which is reflected by a 30% reduction in plasma volume. Plasma ET-1 concentrations increased 2.1 fold and 5.4 fold at 30 and 120 min post-LPS, respectively. LPS-induced losses in plasma volume and albumin escape were significantly attenuated by pretreatment of animals with the dual ET(A)/ET(B) receptor antagonist bosentan (17.4 micromol kg(-1), i.v. 15 min prior to LPS) or the ET(A) receptor antagonist FR 139317 (3.8 micromol kg(-1), i.v.) during both the immediate and delayed phases of endotoxin shock. The inhibitory actions of bosentan and FR 139317 were similar. Both antagonists augmented the hypotensive action of LPS. Administration of bosentan or FR 139317 70 min after injection of LPS also attenuated further losses in plasma volume and increases in total body and organ albumin escape rates with the exception of the lung and kidney. These results indicate a role for endogenous endothelin in mediating losses in plasma volume and albumin escape elicited by LPS predominantly through activation of ET(A) receptors, and suggest that by attenuating these events, ET(A) or dual ET(A)/ET(B) receptor blockers may be useful agents in the therapy of septic shock. PMID- 10696099 TI - Modulation of peristalsis by cannabinoid CB(1) ligands in the isolated guinea-pig ileum. AB - The effect of cannabinoid drugs on peristalsis in the guinea-pig ileum was studied. Peristalsis was induced by delivering fluid into the oral end of an isolated intestinal segment. Longitudinal muscle reflex contraction, threshold pressure and threshold volume to trigger peristalsis, compliance of the intestinal wall during the preparatory phase (a reflection of the resistance of the wall to distension) and maximal ejection pressure during the emptying phase of peristalsis were measured. The cannabinoid agonists WIN 55,212-2 (0.3 - 300 nM) and CP55,940 (0.3 - 300 nM) significantly decreased longitudinal muscle reflex contraction, compliance and maximal ejection pressure, while increased threshold pressure and volume to elicit peristalsis. These effects were not modified by the opioid antagonist naloxone (1 microM) and by the alpha adrenoceptor antagonist phentolamine (1 microM). The inhibitory effect of both WIN 55,212-2 and CP55,940 on intestinal peristalsis was antagonized by the cannabinoid CB(1) receptor antagonist SR141716A (0.1 microM), but not by the cannabinoid CB(2) receptor antagonist SR144528 (0.1 microM). In absence of other drugs, the CB(1) receptor antagonists SR141716A (0.01 - 1 microM) and AM281 (0.01 - 1 microM) slightly (approximatively 20%) but significantly increased maximal ejection pressure during the empty phase of peristalsis without modifying longitudinal muscle reflex contraction, threshold pressure, threshold volume to trigger peristalsis and compliance. It is concluded that activation of CB(1) receptors reduces peristalsis efficiency in the isolated guinea-pig, and that the emptying phase of peristalsis could be tonically inhibited by the endogenous cannabinoid system. PMID- 10696100 TI - Pharmacological characterization of small-conductance Ca(2+)-activated K(+) channels stably expressed in HEK 293 cells. AB - Three genes encode the small-conductance Ca(2+)-activated K(+) channels (SK channels). We have stably expressed hSK1 and rSK2 in HEK 293 cells and addressed the pharmacology of these subtypes using whole-cell patch clamp recordings. The bee venom peptide apamin blocked hSK1 as well as rSK2 with IC(50) values of 3.3 nM and 83 pM, respectively. The pharmacological separation between the subtypes was even more prominent when applying the scorpion peptide blocker scyllatoxin, which blocked hSK1 with an IC(50) value of 80 nM and rSK2 at 287 pM. The potent small molecule blockers showed little differentiation between the channel subtypes. The bis-quinolinium cyclophane UCL 1684 blocked hSK1 with an IC(50) value of 762 pM and rSK2 at 364 pM. The antiseptic compound dequalinium chloride blocked hSK1 and rSK2 with IC(50) values of 444 nM and 162 nM, respectively. The nicotinic acetylcholine receptor antagonist d-tubocurarine was found to block hSK1 and rSK2 with IC(50) values of 27 microM and 17 microM when measured at +80 mV. The inhibition by d-tubocurarine was voltage-dependent with increasing affinities at more hyperpolarized potentials. The GABA(A) receptor antagonist bicuculline methiodide also blocked hSK1 and rSK2 in a voltage-dependent manner with IC(50) values of 15 and 25 microM when measured at +80 mV. In conclusion, the pharmacological separation between SK channel subtypes expressed in mammalian cells is too small to support the notion that the apamin-insensitive afterhyperpolarization of neurones is mediated by hSK1. PMID- 10696101 TI - Possible mechanisms underlying the vasodilatation induced by olprinone, a phosphodiesterase III inhibitor, in rabbit coronary artery. AB - The possible mechanisms underlying the vasodilatation induced by olprinone, a phosphodiesterase type III inhibitor, were investigated in smooth muscle of the rabbit coronary artery. Isometric force and membrane potential were measured simultaneously using endothelium-denuded smooth muscle strips. Acetylcholine (ACh, 3 microM) produced a contraction with a membrane depolarization (15. 2+/ 1.1 mV). In a solution containing 5.9 mM K(+), olprinone (100 microM) hyperpolarized the resting membrane and (i) caused the absolute membrane potential level reached with ACh to be more negative (but did not reduce the delta membrane potential seen with ACh, 15.2+/-1.8 mV) and (ii) attenuated the ACh-induced contraction. In a solution containing 30 mM K(+), these effects were not seen with olprinone. Glibenclamide (10 microM) blocked the olprinone-induced membrane hyperpolarization. 4-AP (0.1 mM) significantly attenuated the olprinone induced resting membrane hyperpolarization but TEA (1 mM) had no such effect. Glibenclamide (10 +microM), TEA (1 mM) and 4-AP (0.1 mM), given separately, all failed to modify the inhibitory actions of olprinone on (i) the absolute membrane potential level seen with ACh and (ii) the ACh-induced contraction. It is suggested that olprinone inhibits the ACh-induced contraction through an effect on the absolute level of membrane potential achieved with ACh in smooth muscle of the rabbit coronary artery. It is also suggested that glibenclamide-sensitive, ATP-sensitive K(+) channels do not play an important role in the olprinone induced inhibition of the ACh-induced contraction. PMID- 10696102 TI - Time-dependent block of the slowly activating delayed rectifier K(+) current by chromanol 293B in guinea-pig ventricular cells. AB - The slowly activating delayed rectifier K(+) current (I(Ks)) was recorded in single myocytes dissociated from guinea-pig ventricles and the mechanism underlying the block of I(Ks) by a chromanol derivative, 293B, was investigated. In the presence of 1 - 100 microM 293B, activation phase of I(Ks) was followed by a slower decay during 10 s depolarizing pulses. Both the rate and extent of the decay were increased in a concentration-dependent manner. The relationship between the concentration of 293B and the block showed a Hill's coefficient of approximately 1. The half-inhibitory concentration was approximately 3.0 microM and did not differ significantly at various membrane potentials from +20 to +80 mV. A mathematical model for the 293B block was constructed on the basis of multiple closed and open states for the I(Ks) channels, and the blocking rate was calculated by fitting the model to the original current traces. The blocking rate constant showed a linear function with the 293B concentration, indicating 1 : 1 binding stoichiometry. At +80 mV the blocking rate was 4x10(4) M(-1) s(-1) and the unblocking rate was 0.2 s(-1). The results indicate that 293B is an open channel blocker with relatively smaller blocking rate than those reported so far for time-dependent blockade of various ionic channels. PMID- 10696103 TI - Decreased aortic glutathione levels may contribute to impaired nitric oxide induced relaxation in hypercholesterolaemia. AB - The aim of this study was to determine if the decrease in aortic total glutathione (GSH) levels in hypercholesterolaemia is related to the impairment of relaxation to acetylcholine (ACh) and exogenous nitric oxide (NO). Isometric tension and vascular GSH levels were measured in thoracic aortic rings from rabbits fed for 12 weeks with 0.5% cholesterol diet. Hypercholesterolaemia decreased aortic GSH levels and impaired relaxation to ACh and NO. To determine if GSH depletion impaired the response to NO, normal rabbit thoracic aorta was incubated with 1,3-bis [2-chloroethyl]-1-nitrosourea (BCNU; 0.2 mmol L(-1)), a GSH reductase inhibitor, or diazine-dicarboxylic acid bis [N, N dimethylamide] (diamide; 1 mmol L(-1)), a thiol oxidizing agent. BCNU or diamide decreased aortic GSH levels and impaired ACh and NO-induced relaxation. The effects of diamide on GSH levels and relaxation were partially prevented by co-incubation with GSH ester (GSE; 2 mmol L(-1)). Increasing GSH with GSE significantly enhanced NO-induced relaxation in aorta from both hypercholesterolaemic and normal rabbits, however relaxation of hypercholesterolaemic rabbit aorta was not restored to normal. These data suggest that other factors, perhaps related to the long-term decrease in GSH levels, are responsible for reduced NO bioactivity in hypercholesterolaemia. PMID- 10696104 TI - Role of clot-associated (-derived) thrombin in cell proliferation induced by fibrin clots in vitro. AB - Thrombin is a potent mitogenic agent. Clot-associated thrombin retains its amidolytic and pro-aggregant activity. We therefore studied the ability of fibrin clots to induce proliferation in CCL39 cells (Chinese hamster lung fibroblasts), in the absence and presence of the thrombin inhibitors PPACK, recombinant hirudin (rHV2 Lys47) and heparin:antithrombin III. Fibrin clots incubated for 48 h with CCL39 cells led to significant cell proliferation, which was dependent on the concentration of thrombin used to prepare the clots. Thus, clots prepared with 91 nmol l(-1) thrombin produced a similar proliferation (231+/-21%) to that obtained with 50 nmol l(-1) thrombin in solution (213+/-29%). Rabbit plasma clots led to a 499+/-41% increase in cell number under identical conditions. Fibrin clot-induced cell proliferation was inhibited by all three thrombin inhibitors with no difference in IC(50) values compared to those obtained against thrombin in solution, suggesting that cell proliferation be due to thrombin leaching from the clots. We found a time-dependent increase in thrombin release from the clots attaining a plateau at 24 h (approximately 61% of the total thrombin used in clot formation). Clots separated from the cells using porous cell culture chamber inserts led to similar proliferation to that of clots in contact with the cells. Thus fibrin-clot induced CCL39 proliferation is due to thrombin released from the clots. PMID- 10696105 TI - Postsynaptic 5-hydroxytryptamine(1A) receptor activation increases in vivo dopamine release in rat prefrontal cortex. AB - 5-Hydroxytryptamine (5-HT) plays a role in the regulation of 3, 4 dihydroxyphenylethylamine (dopamine) neurons in the brain, but the precise mechanism of regulation by 5-HT(1A) receptors of dopamine release has not been defined. The present study describes the effect of 5-?3-[[(2S)-1,4-benzodioxan 2ylmethyl]amino]propoxy?-1, 3-benzodioxole HCl (MKC-242), a highly potent and selective 5-HT(1A) receptor agonist, on dopamine release in the prefrontal cortex using microdialysis in the freely moving rat. Subcutaneous injection of MKC-242 (0.3 - 1.0 mg kg(-1)) increased extracellular levels of dopamine in the prefrontal cortex. The effect of MKC-242 in the prefrontal cortex was antagonized by pretreatment with the selective 5-HT(1A) receptor antagonist, N-[2-[4-(2 methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridinyl)cyclohe xanecarboxamide (WAY100635; 1 mg kg(-1), i.p.). Local application of WAY100635 (10 microM) via a microdialysis probe antagonized the effect of systemic MKC-242 in an increasing dopamine release, and locally infused 8-hydroxy-2-(di-n-propylamino)tetralin (10 microM) increased dopamine release in the prefrontal cortex. MKC-242 increased cortical dopamine release in the rats pretreated with 5, 7-dihydroxytryptamine (150 microgram, i.c.v.) that caused an almost complete reduction in cortical 5-HT content. The effect of MKC-242 to increase dopamine release was also observed in the hippocampus, but not in the striatum or nucleus accumbens. Fluoxetine, a selective serotonin reuptake inhibitor, increased dopamine release in the prefrontal cortex, but not in the nucleus accumbens, while buspirone, a 5-HT(1A) receptor agonist, increased dopamine release in both brain regions. The present results indicate that activation of postsynaptic 5-HT(1A) receptors increases dopamine release in a brain region-specific manner. PMID- 10696106 TI - Nitroblue tetrazolium blocks BK channels in cerebrovascular smooth muscle cell membranes. AB - The effects of p-nitroblue tetrazolium (NBT) on large conductance, calcium activated potassium channels (BK channels) in enzymatically dispersed rat cerebrovascular smooth muscle cells (CVSMCs) were examined. Patch clamp methods were employed to record single BK channel currents from inside-out patches of CVMC membrane maintained at 21 - 23 degrees C. When applied to the cytoplasmic face of inside-out membrane patches (internally applied NBT), micromolar concentrations of NBT reversible reduced the mean open time of BK channels, without changing channel conductance. NBT altered the frequency distribution of BK channel open times from a two exponential to a single exponential form. In the absence of NBT, mean channel open time increased on membrane depolarization. In the presence of internally applied NBT, mean channel open became essentially independent of membrane potential. Internally applied NBT also reduced the mean closed time of BK channels when measured at membrane potentials in the range -80 mV to +20 mV. The combined effects of internal NBT on mean open and closed times resulted in the suppression of BK channel open probability when measured at positive membrane potentials. When applied to the external membrane face, micromolar concentrations of NBT reduced mean channel open time progressively as the membrane was hyperpolarized, and also reduced open probability at negative membrane potentials. A model is proposed in which NBT alters channel gating by binding to a site at or near to the cytoplasmic membrane face. Externally applied NBT suppressed BK channel open probability at concentrations which also inhibit nitric oxide synthase (NOS). Therefore, the potential role of potassium channel block in NBT actions previously attributed to NOS inhibition is discussed. PMID- 10696107 TI - Regional modulation of cyclic nucleotides by endothelin-1 in rat pulmonary arteries: direct activation of G(i)2-protein in the main pulmonary artery. AB - The ability of endothelin-1 (ET-1) to modulate the cyclic nucleotides, guanosine 3' 5' cyclic monophosphate (cyclic GMP) and adenosine 3' 5' cyclic monophosphate (cyclic AMP) was assessed in the main elastic pulmonary elastic artery (4 - 5 mm i.d.) and the small muscular pulmonary arteries (150 - 200 micrometer i.d.) of the rat. ET-1 caused an increase in cyclic GMP in the larger vessels but had no effect in the smaller arteries. The increase in cyclic GMP was not dependent on an intact endothelium and was inhibited by the ET(A)-receptor antagonist FR139137 (1 microM). ET-1 caused a decrease in cyclic AMP in the main pulmonary arteries, an effect that was partially blocked by FR139317 but not influenced by the ET(B) receptor antagonist BQ-788 (1 microM) or removal of the vascular endothelium. In contrast, ET-1 caused an increase in cyclic AMP in the small vessels, an effect that was blocked by BQ-788 but unaffected by FR139317. In the main pulmonary arteries, ET-1 caused enhanced incorporation of radiolabelled ADP-ribose by cholera toxin into G(i)2 in the main pulmonary artery, an indicator of its receptor-mediated activation. In summary, we have shown that in the small muscular pulmonary artery of the rat, (where ET(B) mediated vasoconstriction prevails), there is an ET(B)-mediated increase in cyclic AMP with no net effect on cyclic GMP levels. In the large arteries, (where vasoconstriction is mediated via the ET(A) receptor), there is an ET(A)-mediated increase in cyclic GMP (endothelium independent) and an ET(A)-mediated (endothelium independent) decrease in cyclic AMP. PMID- 10696109 TI - Corrigendum. PMID- 10696108 TI - Bioactive beta-bend structures for the antagonist halpha CGRP(8 - 37) at the CGRP(1) receptor of the rat pulmonary artery. AB - The aim of this study was to determine beta-bend structures and the role of the N and C-terminus in the antagonist halpha CGRP(8 - 37) at the rat pulmonary artery CGRP receptor mediating halpha CGRP relaxation. Halpha CGRP(8 - 37) Pro(16) (10( 6) M), with a bend-biasing residue (proline) at position 16, did not antagonize halpha CGRP responses, while a structure-conserving amino acid (alanine(16)) at the same position retained antagonist activity (apparent pK(B) 6.6+/-0.1; 10(-6) M). Halpha CGRP(8 - 37) Pro(19) (10(-6) M), with proline at position 19 was an antagonist (apparent pK(B) 6.9+/-0.1). Incorporation of a beta-bend forcing residue, BTD (beta-turn dipeptide), at positions 19 and 20 in halpha CGRP(8 - 37) (10(-6) M) antagonized halpha CGRP responses (apparent pK(B) 7.2+/-0.2); and BTD at positions 19,20 and 33,34 within halpha CGRP(8 - 37) was a competitive antagonist (pA(2) 7.2; Schild plot slope 1.0+/-0.1). Halpha CGRP(8 - 37) analogues, substituted at the N-terminus by either glycine(8) or des-NH(2) valine(8) or proline(8) were all antagonists (apparent pK(B) 6.9+/-0.1; (10(-6) M), 7.0+/-0.1 (10(-6) M), and pA(2) 7.0 (slope 1.0+/-0.2), respectively); while replacements by proline(8) together with glutamic acid(10,14) in halpha CGRP(8 - 37) (10(-6) M) or alanine amide(37) at the C-terminus of halpha CGRP(8 - 37) (10( 5) M) were both inactive compounds. In conclusion, possible bioactive structures of halpha CGRP(8 - 37) include two beta-bends (at 18 - 21 and 32 - 35), which were mimicked by BTD incorporation. Within halpha CGRP(8 - 37), the N-terminus is not essential for antagonism while the C-terminus may interact directly with CGRP(1) receptors in the rat pulmonary artery. PMID- 10696110 TI - On a classical limit for electronic degrees of freedom that satisfies the Pauli exclusion principle. AB - Fermions need to satisfy the Pauli exclusion principle: no two can be in the same state. This restriction is most compactly expressed in a second quantization formalism by the requirement that the creation and annihilation operators of the electrons satisfy anticommutation relations. The usual classical limit of quantum mechanics corresponds to creation and annihilation operators that satisfy commutation relations, as for a harmonic oscillator. We discuss a simple classical limit for Fermions. This limit is shown to correspond to an anharmonic oscillator, with just one bound excited state. The vibrational quantum number of this anharmonic oscillator, which is therefore limited to the range 0 to 1, is the classical analog of the quantum mechanical occupancy. This interpretation is also true for Bosons, except that they correspond to a harmonic oscillator so that the occupancy is from 0 up. The formalism is intended to be useful for simulating the behavior of highly correlated Fermionic systems, so the extension to many electron states is also discussed. PMID- 10696111 TI - Macroporous polymer foams by hydrocarbon templating. AB - Porous polymeric media (polymer foams) are utilized in a wide range of applications, such as thermal and mechanical insulators, solid supports for catalysis, and medical devices. A process for the production of polymer foams has been developed. This process, which is applicable to a wide range of polymers, uses a hydrocarbon particulate phase as a template for the precipitation of the polymer phase and subsequent pore formation. The use of a hydrocarbon template allows for enhanced control over pore structure, porosity, and other structural and bulk characteristics of the polymer foam. Polymer foams with densities as low as 120 mg/cc, porosity as high as 87%, and high surface areas (20 m(2)/g) have been produced. Foams of poly(l-lactic acid), a biodegradable polymer, produced by this process have been used to engineer a variety of different structures, including tissues with complex geometries such as in the likeness of a human nose. PMID- 10696112 TI - Diabetes-associated mutations in a beta-cell transcription factor destabilize an antiparallel "mini-zipper" in a dimerization interface. AB - Maturity-onset diabetes of the young, a monogenic form of Type II diabetes mellitus, is most commonly caused by mutations in hepatic nuclear factor 1alpha (HNF-1alpha). Here, the dimerization motif of HNF-1alpha is shown to form an intermolecular four-helix bundle. One face contains an antiparallel coiled coil whereas the other contains splayed alpha-helices. The "mini-zipper" is complementary in structure and symmetry to the top surface of a transcriptional coactivator (dimerization cofactor of homeodomains). The bundle is destabilized by a subset of mutations associated with maturity-onset diabetes of the young. Impaired dimerization of a beta-cell transcription factor thus provides a molecular mechanism of metabolic deregulation in diabetes mellitus. PMID- 10696113 TI - Structure of Escherichia coli ribosomal protein L25 complexed with a 5S rRNA fragment at 1.8-A resolution. AB - The crystal structure of Escherichia coli ribosomal protein L25 bound to an 18 base pair portion of 5S ribosomal RNA, which contains "loop E," has been determined at 1.8-A resolution. The protein primarily recognizes a unique RNA shape, although five side chains make direct or water-mediated interactions with bases. Three beta-strands lie in the widened minor groove of loop E formed by noncanonical base pairs and cross-strand purine stacks, and an alpha-helix interacts in an adjacent widened major groove. The structure of loop E is largely the same as that of uncomplexed RNA (rms deviation of 0.4 A for 11 base pairs), and 3 Mg(2+) ions that stabilize the noncanonical base pairs lie in the same or similar locations in both structures. Perhaps surprisingly, those residues interacting with the RNA backbone are the most conserved among known L25 sequences, whereas those interacting with the bases are not. PMID- 10696114 TI - Imidazenil prevention of alprazolam-induced acquisition deficit in patas monkeys is devoid of tolerance. AB - The partial allosteric modulators (PAMs) of gamma-aminobutyric acid-gated Cl(-) current intensities at gamma-aminobutyric acid type A receptors have high affinity but low intrinsic efficacy on benzodiazepine recognition sites. Unlike the full allosteric modulators (FAM), like alprazolam, triazolam, and diazepam, PAMs are virtually devoid of unwanted side effects, including tolerance. Imidazenil (IMD) is a PAM that elicits potent anxiolytic and anticonvulsant actions in rodents and nonhuman primates and retains its anticonvulsant and anxiolytic effects, even in rodents that are tolerant to FAMs. IMD antagonizes the side effects of FAMs in rodents and nonhuman primates. Using patas monkeys and a multiple schedule with repeated acquisition and performance of chain responses, we report that IMD administration for 17 days antagonized without showing tolerance ALP-induced disruption of acquisition. PMID- 10696115 TI - When the shark bites. PMID- 10696116 TI - The use of mushroom glucans and proteoglycans in cancer treatment. AB - Immunoceuticals can be considered as substances having immunotherapeutic efficacy when taken orally. More than 50 mushroom species have yielded potential immunoceuticals that exhibit anticancer activity in vitro or in animal models and of these, six have been investigated in human cancers. All are non-toxic and very well tolerated. Lentinan and schizophyllan have little oral activity. Active Hexose Correlated Compound (AHCC) is poorly defined but has shown early clinical promise. Maitake D-Fraction has limited proof of clinical efficacy to date, but controlled research is underway. Two proteoglycans from Coriolus versicolor - PSK (Polysaccharide-K) and PSP (Polysaccharide-Peptide - have demonstrated the most promise. In Japanese trials since 1970, PSK significantly extended survival at five years or beyond in cancers of the stomach, colon-rectum, esophagus, nasopharynx, and lung (non-small cell types), and in a HLA B40-positive breast cancer subset. PSP was subjected to Phase II and Phase III trials in China. In double-blind trials, PSP significantly extended five-year survival in esophageal cancer. PSP significantly improved quality of life, provided substantial pain relief, and enhanced immune status in 70-97 percent of patients with cancers of the stomach, esophagus, lung, ovary, and cervix. PSK and PSP boosted immune cell production, ameliorated chemotherapy symptoms, and enhanced tumor infiltration by dendritic and cytotoxic T-cells. Their extremely high tolerability, proven benefits to survival and quality of life, and compatibility with chemotherapy and radiation therapy makes them well suited for cancer management regimens. PMID- 10696117 TI - Male infertility: nutritional and environmental considerations. AB - Studies confirm that male sperm counts are declining, and environmental factors, such as pesticides, exogenous estrogens, and heavy metals may negatively impact spermatogenesis. A number of nutritional therapies have been shown to improve sperm counts and sperm motility, including carnitine, arginine, zinc, selenium, and vitamin B-12. Numerous antioxidants have also proven beneficial in treating male infertility, such as vitamin C, vitamin E, glutathione, and coenzyme Q10. Acupuncture, as well as specific botanical medicines, have been documented in several studies as having a positive effect on sperm parameters. A multi-faceted therapeutic approach to improving male fertility involves identifying harmful environmental and occupational risk factors, while correcting underlying nutritional imbalances to encourage optimal sperm production and function. PMID- 10696118 TI - Nutrients and HIV: part two--vitamins A and E, zinc, B-vitamins, and magnesium. AB - There is compelling evidence that micronutrient deficiencies can profoundly affect immunity; micronutrient deficiencies are widely seen in HIV, even in asymptomatic patients. Direct relationships have been found between deficiencies of specific nutrients, such as vitamins A and B12, and a decline in CD4 counts. Deficiencies appear to influence vertical transmission (vitamin A) and may affect progression to AIDS (vitamin A, B12, zinc). Correction of deficiencies has been shown to affect symptoms and disease manifestation (AIDS dementia complex and B12; diarrhea, weight loss, and zinc), and certain micronutrients have demonstrated a direct anti-viral effect in vitro (vitamin E and zinc). The previous article in this series focused on selenium and beta carotene deficiencies in HIV/AIDS. This literature review elucidates how deficiencies of the micronutrients zinc, magnesium, vitamins A, E, and specific B vitamins relate to HIV symptomology and progression, and clearly illustrates the need for nutritional supplementation in HIV disease. PMID- 10696119 TI - Environmental medicine, part one: the human burden of environmental toxins and their common health effects. AB - Chemical compounds ubiquitous in our food, air, and water are now found in every person. The bioaccumulation of these compounds in some individuals can lead to a variety of metabolic and systemic dysfunctions, and in some cases outright disease states. The systems most affected by these xenobiotic compounds include the immune, neurological, and endocrine systems. Toxicity in these systems can lead to immune dysfunction, autoimmunity, asthma, allergies, cancers, cognitive deficit, mood changes, neurological illnesses, changes in libido, reproductive dysfunction, and glucose dysregulation. Chemicals and their effects on these systems are reviewed in this article. Subsequent articles in this series will focus on therapeutic regimens to combat the toxic effects of these and other compounds. PMID- 10696120 TI - Use of neurotransmitter precursors for treatment of depression. AB - Insufficient activity of the neurotransmitters serotonin and norepinephrine is a central element of the model of depression most widely held by neurobiologists today. In the late 1970s and 1980s, numerous studies were performed in which depressed patients were treated with the serotonin precursors L-tryptophan and 5 hydroxytryptophan (5-HTP), and the dopamine and norepinephrine precursors tyrosine and L-phenylalanine. This article briefly reviews the published research on the efficacy of neurotransmitter precursors in treating depression, highlights the findings of studies, and discusses issues regarding the interpretation of those findings. The nature of the studies makes it difficult to draw firm conclusions regarding the efficacy of neurotransmitter precursors for treating depression. While there is evidence that precursor loading may be of therapeutic value, particularly for the serotonin precursors 5-HTP and tryptophan, more studies of suitable design and size might lead to more conclusive results. However, the evidence suggests neurotransmitter precursors can be helpful in patients with mild or moderate depression. PMID- 10696121 TI - Cytogenetics of hepatoblastoma: further characterization of 1q rearrangements by fluorescence in situ hybridization: an international collaborative study. AB - BACKGROUND: Hepatoblastoma (HBT) is the most common hepatic neoplasm in children. This notwithstanding, little is known about pathogenetic factors, such as genetic abnormalities, of importance for the development and progression of this tumor type. To date, only 33 cytogenetically abnormal HBT have been published, and trisomies for chromosomes 2 and 20 have been shown to be the most frequent aberrations. Recently, unbalanced translocations involving proximal 1q have been described in several HBT, suggesting that a pathogenetically important gene maps to 1q. PROCEDURE: Six primary and one recurrent HBT were cytogenetically analyzed after short-term tissue culture. In addition, fluorescence in situ hybridization (FISH) studies, using locus-specific probes, were performed on three of these pediatric HBT as well as on one previously reported adult HBT. RESULTS: Total or partial trisomy 8, gain of chromosome 20, and structural rearrangements of chromosome 1 were detected in three HBT, and overrepresentation of chromosome 2 material was found in two HBT. The adjacent chromosome bands 1q12 and 1q21 were involved in three translocations, t(1;2), t(1;4), and t(1;11), which were all unbalanced and resulted in gain of 1q material. The previously reported adult HBT displayed 1q deletions with breakpoints at 1q12-21. FISH analyses of the 1q rearrangements revealed that all breakpoints were within the heterochromatic region. CONCLUSIONS: These findings provide further support for the importance of trisomies 2, 8, and 20 and rearrangements of 1q in the development of HBT. Furthermore, the consistent localization of breakpoints within the heterochromatic segment of chromosome 1 suggests that the important pathogenetic consequence of 1q abnormalities is the resulting genomic imbalance rather than a specific gene rearrangement. PMID- 10696122 TI - Allogeneic peripheral blood stem cell transplantation in children with hematologic malignancies from HLA-matched siblings. AB - BACKGROUND: Despite the ethical problem of using granulocyte colony-stimulating factor (G-CSF) in normal children, allogeneic peripheral blood stem cell transplantation (PBSCT) might have advantages over allogeneic bone marrow transplantation (BMT). PROCEDURE: Eleven HLA-matched sibling donors aged 2-16 years received 10 microg/kg/day G-CSF for 5 days and underwent apheresis to harvest peripheral blood stem cells (PBSC). PBSC were then cryopreserved until infusion. The 11 corresponding patients aged 8 months to 14 years with high-risk hematological malignancies received busulfan (16 mg/kg or 600 mg/m(2)) and melphalan (210 mg/m(2)) as a preparative regimen. Graft-versus-host disease (GVHD) prophylaxis consisted of cyclosporine and methylprednisolone. RESULTS: All of the donors tolerated G-CSF administration and apheresis procedures. The patients received a median of 5.8 (range 1. 4-11.5) x 10(6)/kg CD34(+) cells, 17.2 (3.8-36.0) x 10(5)/kg colony forming units-granulocyte/macrophage (CFU-GM), and 3.5 (1.4-7.1) x 10(8)/kg CD3(+) cells. All of the patients showed prompt engraftment, with a median time to reach an absolute neutrophil count (ANC) above 0.5 x 10(9)/liter of 10 (9-13) days. Grade I acute GVHD occurred in seven patients (64%), whereas grade II-IV acute GVHD was not seen. Chronic GVHD occurred in four patients (40%) among 10 patients evaluable for chronic GVHD. Three patients showed extensive chronic GVHD. Currently, eight patients (73%) are alive and disease-free for a median follow-up of 775 (103-1,069) days. CONCLUSIONS: Allogeneic PBSCT is feasible in the pediatric population, and PBSC harvest is an alternative to BM harvest in donors who are not eligible for BM harvest. Furthermore, PBSC were successfully collected in pediatric donors with peripheral access. The choice of a stem cell source should be based on the risk/benefit assessment for both patients and donors. PMID- 10696123 TI - Efficacy and safety of radiologically placed gastrostomy tubes in paediatric haematology/oncology patients. AB - BACKGROUND: The treatment of malnutrition, which is of great concern in paediatric haematology/oncology patients, is fraught with problems. The goals of our study were to document the complications and assess the weight gain with gastrostomy tubes (G-tubes) in this population. PROCEDURE: Patient data were acquired by retrospective review of all haematology, oncology, and bone marrow transplant (BMT) patients (n = 44) who received radiologically placed G-tubes at our institution over a 4-year period. RESULTS: Forty-four G-tubes were placed (59% peri-BMT). At tube placement, 55% of patients were malnourished and 45% were nourished. Seventy-five percent of patients had the procedure without general anaesthetic. Localized G-tube-site infection was the most common complication (41%). Major complications occurred in 3 patients; 2 patients experienced localized peritonitis, and 1 patient developed a localized collection of pus in the abdominal wall. There were no G-tube-related deaths. At 1 month after the tube insertion, 39% of patients had gained, 54% maintained, and 7% lost weight. At 3 months, 69% had gained, 28% maintained, and 3% lost weight. There was a statistically significant weight gain from the time of the G-tube placement to both 1 month (P < 0.018) and 3 months (P < 0.0001) after G-tube placement. Patients in all diagnosis categories showed improvement from 1 to 3 months. CONCLUSIONS: We conclude that retrograde tube placement is safe and can frequently be done without general anaesthetic and that G-tube feeding effectively reverses malnutrition and prevents weight loss in this patient population. PMID- 10696124 TI - Complete necrosis induced by preoperative chemotherapy in Wilms tumor as an indicator of low risk: report of the international society of paediatric oncology (SIOP) nephroblastoma trial and study 9. AB - BACKGROUND: The SIOP Nephroblastoma therapeutic protocols include a period of preoperative chemotherapy followed by nephrectomy and a period of postoperative chemotherapy. From the outset, identification of low-risk groups has been an aim of the SIOP Nephroblastoma Trials and Studies. Now that 90% of children with Wilms tumor can be cured, attention is even more focused on the identification of patients who could benefit from less aggressive postoperative therapy, thus minimizing the morbidity and late effects associated with treatment. The prognostic implications of total necrosis in nephroblastoma after chemotherapy have not been investigated hitherto. PROCEDURE: Between November 1, 1987 and June 30, 1993, 599 patients referred to the SIOP-9 Nephroblastoma Trial and Study were preoperatively treated and classified as stages I-IV nonanaplastic Wilms tumor. RESULTS: Of these 599 patients, pathologic examination of the nephrectomy specimen revealed a completely necrotic Wilms tumor (CNWT) with no viable tumor remaining in 59 (10%): these comprised 37 stages I-III and 22 stage IV. Of these patients, 58 (98%) had no evidence of disease at 5 years vs. 90% for the rest of the cohort (P < 0.05). Stages I-III patients represented 63% of CNWT and had a 97% overall survival rate. The only death was related to veno-occlusive disease and occurred in a stage I patient in the month following nephrectomy. Stage IV patients represented 37% of CNWT (vs. only 10% of all other cases of unilateral nonanaplastic Wilms tumor) and had a 100% rate of survival. Children with CNWT were older (mean 59 months vs. 43 months); their tumor at diagnosis was larger and had regressed more significantly at subsequent ultrasound examination. The data also uphold the hypothesis that Wilms tumors of blastemic pattern are most aggressive, but also are extremely responsive to chemotherapy. CONCLUSIONS: Patients with unilateral nonanaplastic WT that showed total necrosis following preoperative chemotherapy had excellent outcome and should benefit from less aggressive postoperative treatment in further trials. Other very responsive tumors, such as Wilms with <10% viable tumor, should also be assessed. PMID- 10696125 TI - Cryopreservation of semen from pubertal boys with cancer. AB - BACKGROUND: The possibility of cryopreservation of semen from adolescents has until now received only little attention. Therefore, we have investigated the possibility of cryopreservation of semen in adolescent boys with cancer. PROCEDURE: Forty-five boys, aged 13-18 years, admitted because of cancer during the period January 1, 1995 to July 31, 1998 were eligible. Semen was obtained after masturbation in the majority of the cases. In three boys, semen was preserved after penile vibration or electroejaculation in general anaesthesia. The semen samples were analysed for concentration, motility, and morphology according to the WHO guidelines. The sample was transferred into straws prior to cryopreservation at 196 degrees C in liquid nitrogen. RESULTS: Twenty-one boys delivered a semen sample for cryopreservation. Four boys were offered and accepted sperm banking but were not able to produce a sample. In 20 cases time did not allow an attempt of sperm banking, the boy was not assessed to be mature enough to deliver a semen sample, or the procedure was not accepted. The boys delivered 1-3 samples, and the total number of spermatozoa ranged from 0-210 millions. Median percentage of motile sperm was 50% (range 9-86%). Semen quality improved with age; however, a 13- year- old boy produced 75 million spermatozoa with 38% motile cells. CONCLUSIONS: Pubertal maturation should be assessed in all boys admitted for cancer, and the possibility of sperm banking should be discussed with the patient and his parents. PMID- 10696126 TI - Inguinal hernia in patients with Ewing sarcoma: a clue to etiology. AB - BACKGROUND: Various congenital anomalies have been associated with childhood cancer, but as yet no anomaly has been consistently found with Ewing sarcoma (ES). Recently a large case-control study of ES patients reported a greater number of hernias in both cases and their sibling controls than in population controls. Most of these hernias were inguinal. Because these anomalies were also reported previously in two case series, we looked for inguinal hernias in a different population of ES patients. PROCEDURE: We abstracted medical records for 306 pathologically confirmed ES/primitive neuroectodermal tumor (PNET) patients seen at NIH between 1960 and 1992. Epidemiological data on demographics and medical conditions were analyzed. The frequency of anomalies was compared to expected rates to calculate relative risk and confidence intervals. RESULTS: Anomalies were present in 67 (22%) cases. A particular anomaly, inguinal hernia, was reported for 13 (5%) NIH cases. Compared to population estimates for white children, the relative risk of inguinal hernia among white NIH cases was 13.3 (95% CI 3.60-34.1) for females and 6.67 (95% CI 2.67-13.7) for males. CONCLUSIONS: The findings of inguinal hernias in some patients with ES suggest that a disruption in normal embryological development occurred. This may provide an important clue to the etiology of ES. We hypothesize that these hernias may relate to an in utero exposure or indicate an underlying genetic disorder. Future studies should carefully evaluate ES families for genetic disease and explore environmental factors. Med. Pediatr. Oncol. 34:195-199, 2000. Published 2000 Wiley-Liss, Inc. PMID- 10696127 TI - Pegaspargase-induced pancreatitis. AB - BACKGROUND: The purpose of this study is to report the incidence of pancreatitis in patients treated with pegaspargase in our hospital during a 2-year period. PROCEDURE: We identified episodes of pancreatitis related to the intramuscular administration of pegaspargase 2,500 IU/m(2) for the treatment of childhood hematological malignancies during a 2-year period (May 1996-April 1998). Patients were evaluated clinically and by sequential serum amylase and lipase determinations and radiographic examinations. For comparison, episodes of pancreatitis in patients who only received native Escherichia coli L-asparaginase were examined during the same time period. RESULTS: Nine children with acute lymphoblastic leukemia (ALL) of 50 (18%) patients who received pegaspargase were diagnosed to have pancreatitis. All had prior therapy with native L-asparaginase. These children developed symptoms consisting of abdominal pain, nausea, vomiting, and decreased appetite within a median of 15 days from the onset of pegaspargase administration. Six patients became symptomatic after their initial dose. Seven patients developed severe or unacceptable toxicity (grades 3 and 4), measured by increased amylase (>2 times normal) and lipase levels or radiographic evidence of pancreatic inflammation or pseudocyst. One patient also developed hyperammonemia and encephalopathy. In contrast, only one out of 52 (1.9%) ALL patients who received native E. coli L-asparaginase during the same time period developed pancreatitis (P= 0.007). CONCLUSION: Clinicians should be aware of a possible higher incidence of pancreatitis associated with pegaspargase. PMID- 10696128 TI - Burkitt lymphoma in the Cote D'Ivoire from 1966 to 1995: a progress report. AB - BACKGROUND AND PROCEDURE: Eighteen years of experience with Burkitt lymphoma (BL), the most common form of childhood cancer in the Ivory Coast, has been reviewed and the epidemiology analyzed. RESULTS: BL is more commonly found in wooded areas than savannahs by a factor of 5:1. Despite good responses to cyclophosphamide-based chemotherapy, chances for survival are often dictated by socioeconomic factors that limit optimal care. CONCLUSIONS: Better public health measures and improved education and socioeconomic conditions are needed if the fruits of earlier and more aggressive therapies are to be gathered. PMID- 10696129 TI - Breast cancer screening in childhood cancer survivors. PMID- 10696130 TI - Diffuse intrinsic brainstem disease with neurologic deterioration: not what it seemed. PMID- 10696131 TI - Catheter-directed thrombolysis in a child with acute lymphoblastic leukemia and extensive deep vein thrombosis. PMID- 10696132 TI - Metastatic appendiceal carcinoid tumor in a child. PMID- 10696133 TI - Successful treatment of vancomycin-resistant Enterococcus sepsis in a neutropenic patient with G-CSF-mobilized granulocyte transfusions. PMID- 10696134 TI - Long-term remission induced by corticosteroids, cyclophosphamide, and methotrexate in a patient with natural killer cell leukemia. PMID- 10696135 TI - Successful rescue by oral cholestyramine of a patient with methotrexate nephrotoxicity: nonrenal excretion of serum methotrexate. PMID- 10696136 TI - Complete regression of choroidal metastases from breast cancer after docetaxel based systemic chemotherapy. PMID- 10696137 TI - Paraneoplastic Cushing syndrome due to adrenal neuroblastoma. PMID- 10696138 TI - Synovial sarcoma mimicking desmoplastic small round-cell tumor: critical role for molecular diagnosis. PMID- 10696139 TI - TGFbeta1 selectively up-regulates CCR1 expression in primary murine astrocytes. AB - Chemokine receptors dictate the cellular responses to chemokines on target cells. Therefore, the regulation of expression of chemokine receptors is likely a crucial point for the regulation of chemokine action. Here we show that CC chemokine receptor 1 (CCR1) expression by primary mouse astrocytes is increased after transforming growth factor beta1 (TGFbeta1) stimulation. TGFbeta1 caused a pronounced up-regulation of CCR1 mRNA in a concentration- and time-dependent manner. TGFbeta1-mediated increase of CCR1 mRNA accumulation resulted in increased CCR1 protein expression and augmented cell migration to a physiological ligand, macrophage inflammatory protein-1alpha (MIP-1alpha). The half life of CCR1 mRNA in the presence and absence of TGFbeta1 stimulation was comparable, suggesting that TGFbeta1-induced CCR1 mRNA accumulation occurred at the transcriptional level. TGFbeta1 did not affect CCR1 mRNA expression in hematopoietic cells, indicating that TGFbeta1 effect on CCR1 expression in primary astrocytes is cell-type specific. This is the first evidence that TGFbeta1 may modulate central nervous system (CNS) inflammation in part by affecting chemokine receptor expression on astrocytes. PMID- 10696140 TI - Inhibition of glutamine synthetase in rabbit pneumococcal meningitis is associated with neuronal apoptosis in the dentate gyrus. AB - Apoptosis of dentate granular cells in the hippocampal formation during bacterial meningitis may be mediated by glutamate toxicity. For this reason, we studied the relationship between glutamine synthetase activity and regional neuronal apoptosis in rabbits with experimental pneumococcal meningitis. The duration of meningitis was 24 h, and the treatment was started 16 h after infection. Significant increases of glutamine synthetase protein concentration (P < 0.05) were found in the frontal cortex of rabbits with meningitis (n = 7) and rabbits with meningitis receiving ceftriaxone treatment (n = 12) as compared to the control animals (n = 14). No significant differences were seen in the hippocampal formation. The enzymatic activity of glutamine synthetase also was elevated in the frontal cortex (P < 0.05), but not in the hippocampal formation of rabbits with meningitis. After intravenous administration of L-methionine sulfoximine (specific inhibitor of glutamine synthetase) in rabbits with meningitis treated with ceftriaxone (n = 10), the concentration of neuron-specific enolase in CSF (P = 0.025) and the density of apoptotic neurons in the dentate gyrus quantified with the in-situ tailing reaction (P = 0.043) were higher than in meningitic animals receiving only ceftriaxone (n = 10). In conclusion, the inability of hippocampal glutamine synthetase to metabolize excess amounts of glutamate may contribute to neuronal apoptosis in the hippocampal formation during meningitis. PMID- 10696141 TI - Synthesis and release of L-serine by rat astroglia-rich primary cultures. AB - L-serine is known to have important functions in the mammalian CNS other than being a constituent of proteins. It is the metabolic precursor of the neuroactive substances D-serine and glycine, serves as a building block for phospholipid biosynthesis and is likely to be a neurotrophic factor. Based on the observation that rat astroglia-rich primary cultures release L-serine into their culture medium, the biosynthesis and release of L-serine in these cultures has been investigated. Release of L-serine is due to both biosynthesis from glucose and to proteolysis. Amino groups for L-serine synthesis originate from transamination of amino acids that are either taken up from the extracellular space or produced intracellularly by proteolysis. The enzymes of the "phosphorylated pathway" of serine biosynthesis, i.e., 3-phosphoglycerate dehydrogenase, phosphoserine aminotransferase and phosphoserine phosphatase are present in rat brain as well as in rat astroglia-rich primary cultures and their specific activities have been determined. The presence of these enzymes indicates the operation of the "phosphorylated pathway" of L-serine biosynthesis in brain. PMID- 10696142 TI - Ion channel expression by astrocytes in situ: comparison of different CNS regions. AB - Patch-clamp recordings were obtained in brain slices from 283 rat astrocytes. The expression of voltage-activated whole-cell currents was compared in four different CNS regions (hippocampus, cerebral cortex, spinal cord, and cerebellum). Our data show that CNS astrocytes do not show significant regional differences in their ion channel complement. With the exception of cerebellar Bergmann glial cells, essentially all astrocytes express a combination of delayed rectifying outward K(+) currents, transient A-type K(+) currents, and small Na(+) currents. Developmentally, an increasing percentage of astrocytes and Bergmann glial cells express inwardly rectifying K(+) currents. We did not observe cells that were passive, i.e., lacking voltage-activated currents. A few cells that appeared "passive" in initial recordings showed voltage-activated K(+) currents after off-line leak subtraction. The heterogeneity observed in the ion channel complement was found to be identical when cell-to-cell variations observed within a given CNS region and between various CNS regions were compared, suggesting a common and fairly stereotypical complement of ion channels in CNS astrocytes. Ion channel expression in Bergmann glial cells differed from that of all other CNS regions studied. These cells typically showed very low input resistances attributable to a significant time- and voltage-independent resting K(+) conductance. However, as with electrophysiologically "passive"-appearing astrocytes, Bergmann glial cells showed expression of delayed rectifying K(+) currents after off-line leak subtraction. Inwardly rectifying K(+) currents were observed in Bergmann glial cells after postnatal day 17. Collectively, our data suggest that all astrocytes contain voltage-gated ion channels that display a common pattern of expression during development. PMID- 10696143 TI - Changes in ion channel expression accompany cell cycle progression of spinal cord astrocytes. AB - Arrest of spinal cord astrocytes at defined stages of the cell cycle clock causes significant changes in the expression of voltage-activated Na(+) and K(+) currents. Arrest of actively proliferating astrocytes in G1/G0 by all-trans retinoic acid induces premature expression of inwardly rectifying K(+) currents (IK(IR)) typically expressed only in differentiated astrocytes. By contrast, arrest in S phase by ara-C or Aphidicolin leads to a greater than twofold increase in "delayed" outwardly rectifying currents (IK(D)) and a concomitant decrease in IK(IR). Pharmacological blockade of IK(D) by TEA and 4AP caused proliferating astrocytes to arrest in G0/G1, suggesting that activity of these channels is required for G1/S checkpoint progression. Conversely, in quiescent astrocytes, inhibition of IK(IR) by 30 microM BaCl(2) led to an increase in astrocyte proliferation and to an increase in the number of cells in S phase from 5% to 26%. These data suggest that a downregulation of K(IR) promotes cell cycle progression through the G1/S checkpoint. Blockade of IK(IR) in actively proliferating cells, however, leads to an accumulation in G2/M, suggesting that reappearance of this current may be critical for progression beyond DNA synthesis. Interestingly, Na(+) currents (INa(+)) are increased greater than fourfold in S phase-arrested cells, yet their pharmacological blockade by TTX has no effect on cell cycle progression. However, the resting membrane potential of S phase-arrested cells increases profoundly, and manipulation of membrane potential by the application of low concentrations of ouabain, or reduction of extracellular potassium, induces the accumulation of quiescent astrocytes in S phase of the cell cycle, suggesting that either depolarization or intracellular sodium, or both, play an important role in promoting astrocyte proliferation. PMID- 10696144 TI - Schwann-like macroglia in adult rat brain. AB - Olfactory ensheathing cells (OECs) share properties with astrocytes and Schwann cells. This study was designed to test the hypothesis that glia with properties similar to those exhibited by OECs might be present in brain areas other than the olfactory bulb. We found tanycytes and pituicytes to express a distinctive set of immunological markers in common with OECs and nonmyelinating Schwann cells, namely low-affinity neurotrophin receptor (p75NTR), O4 antigen, estrogen receptor alpha type, and insulin-like growth factor 1 (IGF-1). The two glial types could be cultured from adult hypothalamus and neurohypophysis, respectively, using the methods developed for olfactory OECs. Both glial types displayed morphologies reminiscent of Schwann cells, in primary culture. Schwann-like central glia presented a preferred growth substrate for dorsal root ganglion neurites and, when making intimate contacts with them, manifested a myelinating phenotype. These combined properties define a type of CNS macroglia that would not fit within conventional central glia types. PMID- 10696145 TI - Inhibition of Muller cell glutamine synthetase rapidly impairs the retinal response to light. AB - It is widely assumed that neurones have sufficient metabolic reserves to allow them to function independently of glial cells for extended periods. The present study investigates the length of time taken before retinal neurones no longer respond normally to light after the inhibition of glial enzymes that are involved in the synthesis of precursors of neuronal glutamate. The glutamine synthetase inhibitor methionine sulfoximine, when injected intraocularly in Wistar rats, caused a time- and dose-dependent suppression of the scotopic electroretinogram b wave. At the highest dosage (40 mM) the b-wave was significantly reduced within 2 min of injection. Because the b-wave is an indicator of neurotransmission in the retina, it is deduced that inhibition of glutamine synthetase rapidly blocks glutamatergic neurotransmission. Immunohistochemistry revealed a depletion of neuronal glutamate and an accumulation of glutamate in Muller glial cells, in a time course that matched the b-wave suppression. The b-wave was quickly restored by injection of glutamine (4 mM). The rapid reduction of glutamatergic transmission after methionine sulfoximine administration challenges the view that neurones have sufficient reserves to allow them to function independently for extended periods; instead, it indicates that glia are essential for the moment-to moment sustenance of neuronal function. PMID- 10696146 TI - Distinct patterns of stimulus-inducible chemokine mRNA accumulation in human fetal astrocytes and microglia. AB - Interferon-gamma-inducible 10 kd protein (IP-10) is an ELR (Glu-Leu-Arg)(-) alpha chemokine with known chemotactic effects on T cells and monocytes, as well as anti-viral, anti-angiogenic, and anti-tumor effects. Previous studies have demonstrated that in cultured rat astrocytes and microglia, stimulation with LPS or virus can induce the expression of IP-10. In this study, we determined the pattern of IP-10 gene induction in primary human microglia and astrocytes by cytokines and LPS using ribonuclease protection assay. The expression of IP-10 mRNA was compared with that of other alpha (IL-8) and beta chemokines. The results showed that in human microglia, IP-10 expression was induced equally potently by LPS, IFNbeta or IFNgamma. "Proinflammatory" cytokines IL-1beta or TNFalpha also induced small amounts of IP-10 mRNA. "Anti-inflammatory" cytokines IL-4, IL-10 and TGFbeta were ineffective in inducing IP-10 in microglia. In human astrocytes, induction of IP-10 mRNA by cytokines was similar to that in microglia. LPS, however, was ineffective in inducing IP-10 in human astrocytes. The monocyte chemoattractant beta-chemokine I-309 mRNA was induced in human astrocytes and microglia by IFNbeta or IFNgamma, or by LPS in microglia, showing a tight co-regulation with IP-10 mRNA expression. In contrast to the potent induction of IP-10 and I-309 by IFNs in human glia, the ELR(+) alpha chemokine IL 8 mRNA was induced by IL-1beta and TNFalpha, and to a lesser extent by IFNbeta in microglia. IFNbeta but not IFNgamma was effective in inducing the expression of beta chemokines MIP-1alpha and MIP-1beta in human microglia, with the levels of mRNA similar to those induced by IL-1beta or TNFalpha. Neither MIP-1alpha nor MIP 1beta mRNAs were induced by any stimulation in human astrocytes. The induction of RANTES mRNA in microglia by IFNbeta, IL-1beta or TNFalpha was variable, showing no to low level expression depending on the case, whereas LPS provided a consistent inducing signal. In astrocytes, only cytokine combinations (IFN + IL 1beta) effectively induced the RANTES mRNA. These results demonstrate that distinct sets of chemokine genes are induced in human glial cells by cytokines and interferons. These results may have wide implications for inflammatory, vascular and neoplastic diseases of the CNS. PMID- 10696147 TI - Sub-population of cultured hippocampal astrocytes expresses neuropeptide Y Y(1) receptors. AB - The expression and pharmacological characterization of neuropeptide Y (NPY) receptors of the Y(1) subtype on cultured hippocampal neurons was reported using radioreceptor assays and immunohistochemical approaches (St-Pierre et al., 1998). The present study aimed to establish the presence of NPY Y(1) receptors on cultured hippocampal astrocytes using similar strategies. Immunocytochemical experiments were carried out using three antisera directed against distinct domains (amino acids sequence 185-203, 198-213 and 355-382) of the Y(1) receptor. Double-labeling experiments and confocal microscopy with these Y(1) receptor antisera demonstrated their recognition of the same sub-population (20%) of GFAP positive astrocytes in culture. The immunostaining seen with all three Y(1) receptor antisera took the form of large irregular clusters distributed throughout cell bodies and processes. Further experiments using radioactive ligands confirmed the presence of NPY receptors on cultured hippocampal astrocytes. Emulsion receptor autoradiography using a newly developed ligand, [(125)I]GR231118 in the presence of PYY, hPP or BIBP3226 (1 microM), pharmacologically established the Y(1) nature of these receptors. Specific [(125)I]GR231118 binding was competed by PYY and the selective Y(1) antagonist BIBP3226 but not by hPP (a Y(4)/Y(5) agonist). Similar autoradiographic labeling patterns were obtained using [(125)I][Leu(31).Pro(34)]PYY (a Y(1)/Y(4)/Y(5) agonist) whereas [(125)I]PYY(3-36) (a Y(2)/Y(5) agonist) failed to generate any specific signal. Hence, rat cultured hippocampal astrocytes express a typical Y(1) receptor without evidence for the presence of Y(2), Y(4) or Y(5) subtypes. These data suggest a preferential regulation by NPY, acting via the Y(1) receptors, of astrocytic function. PMID- 10696148 TI - Lysophosphatidylcholine induces rapid recruitment and activation of macrophages in the adult mouse spinal cord. AB - Lysophosphatidylcholine (LPC) can induce rapid breakdown and removal of myelin from the adult mammalian CNS. In this paper we report the detailed characterization of the immune cell response as well as changes in the expression of cell adhesion molecules and the permeability of the blood-brain barrier after microinjection of LPC into the adult mouse spinal cord. T cells and neutrophils were seen in the spinal cord 6-12 h after LPC injection, but not in PBS-injected mice. Mac-1+ monocytes were also seen at 6 h and 12 h in the white and gray matter of mice injected with LPC and PBS but were significantly greater in the white matter after LPC injections. At later time points LPC induced an increase in the number of activated Mac-1+ macrophages that displayed a variety of morphologies in the white and gray matter. These cells were not present in PBS injected control mice. LPC also induced widespread microglial activation in the white and gray matter. The number of these Mac-1+ microglia reduced drastically at 96 h after LPC injection suggesting that they may have transformed into Mac-1+ phagocytic cells with a different morphology. These LPC-induced changes in immune cells were accompanied by significant increases in VCAM-1+ and ICAM-1+ blood vessels in the spinal cord. In addition, LPC induced a rapid and widespread disruption of the blood-brain barrier, as compared to PBS injected mice. Therefore, LPC can induce an early and transient T cell and neutrophil response in the CNS. These cells likely promote the rapid influx of monocytes followed by widespread and effective activation of macrophages that mediate rapid phagocytosis of myelin debris. PMID- 10696149 TI - Multivariate analysis of diet among three-year-old children and associations with socio-demographic characteristics. The Avon Longitudinal Study of Pregnancy and Childhood (ALSPAC) Study Team. AB - STUDY OBJECTIVE: The study of the whole diet in combination rather than the consumption of individual food items or the intake of specific nutrients could be enlightening. This has been previously performed using principal components analysis (PCA) on adult diets but not on those of children. DESIGN: The frequency of consumption of a range of food items was recorded for 10,139 3-y-old children by their mothers using self-completion postal questionnaires. These children form part of the Avon Longitudinal Study of Pregnancy and Childhood (ALSPAC). METHODS: PCA was performed to identify individual dietary types which were then related to various socio-economic and demographic characteristics. RESULTS: Four distinct dietary components were obtained explaining 23.5% of the total variation in the sample, and the socio-demographic characteristics of the sample were related to them. The first represented a diet based on convenience foods and was associated with younger, less educated mothers and the presence of older siblings. The second was associated with a high consumption of foods currently considered to be healthy and was particularly related to vegetarian mothers and higher education levels. The third component described the established British 'meat and two veg' diet and was associated with girls and children with no older siblings, while the fourth had high loadings for snack and finger foods and was related to socially advantaged conditions and the presence of older siblings. CONCLUSIONS: Identifiable groups of mothers were associated with feeding their child each of the four dietary types, supporting the hypothesis that social, demographic and lifestyle factors relating to the mother have an influence on the early eating patterns of children. This analysis will form a basis for the future study of various childhood outcomes including growth, health and development. SPONSORSHIP: University of Bristol European Journal of Clinical Nutrition (2000) 54, 73-80 PMID- 10696150 TI - Sex Appeal. PMID- 10696151 TI - The relationship of lumbar flexion to disability in patients with low back pain. AB - BACKGROUND AND PURPOSE: Physical therapists routinely assess spinal active range of motion (AROM) in patients with low back pain (LBP). The purpose of this study was to use 2 approaches to examine the relationship between impairment of lumbar spine flexion AROM and disability. One approach relied on the use of normative data to determine when an impairment in flexion AROM was present. The other approach required therapists to make judgments of whether the flexion AROM impairment was relevant to the patient's disability. SUBJECTS: Fifteen physical therapists and 81 patients with LBP completed in the study. METHODS: Patients completed the Roland-Morris Back Pain Questionnaire (RMQ), and the therapists assessed lumbar spine flexion AROM using a dual-inclinometer technique at the initial visit and again at discharge. RESULTS: Correlations between the lumbar flexion AROM measure and disability were low and did not vary appreciably for the 2 approaches tested. CONCLUSION AND DISCUSSION: Measures of lumbar flexion AROM should not be used as surrogate measures of disability. Lumbar spine flexion AROM and disability are weakly correlated, suggesting that flexion AROM measures should not be used as treatment goals. PMID- 10696152 TI - Lower extremity compensations following anterior cruciate ligament reconstruction. AB - BACKGROUND AND PURPOSES: Several studies have demonstrated that patients with knee injury scored within a normal range during one-legged hop tests, yet showed quadriceps femoris muscle weakness with non-weight-bearing isokinetic testing. This study evaluated lower-extremity kinetics while subjects performed a single leg vertical jump (VJ) and a lateral step-up (LSU) in an attempt to explain this phenomenon. SUBJECTS AND METHODS: Using a motion analysis and force platform system, hip, knee, and ankle extension moments of 20 subjects with anterior cruciate ligament (ACL) reconstructions and 20 matched subjects were measured while they performed an LSU and a VJ. RESULTS: An analysis of variance revealed that the knee extension moment of the ACL-reconstructed extremity was lower than that of the uninjured and matched extremities during the LSU, VJ take-off, and VJ landing. However, there was no difference in summated extension moment (hip + knee + ankle) among extremities during the LSU and VJ take-off. The summated extension moment of the ACL-reconstructed extremity during VJ landing was less than that of the uninvolved and matched extremities. CONCLUSION AND DISCUSSION: These results suggest that the hip or ankle extensors may compensate for the knee extension moment deficit. The decrease in summated extension moment in the ACL reconstructed extremity during VJ landing represents inadequate attenuation of landing forces, which may expose the skeleton and joint structures to injury. PMID- 10696153 TI - Lumbar lordosis and pelvic inclination in adults with chronic low back pain. AB - BACKGROUND AND PURPOSE: The causes of lumbopelvic imbalances in standing have been widely accepted by physical therapists, but there is a lack of scientific evidence available to support them. We examined the association between 9 variables and pelvic inclination and lumbar lordosis during relaxed standing. SUBJECTS: Thirty men and 30 women with chronic low back pain (CLBP) for at least 4 months were examined (mean age=54.9 years, SD=9, range=40.4-69.8). METHODS: Multiple linear regression modeling was used to assess the association of pelvic inclination and the magnitude of lumbar lordosis in standing with age, sex, body mass index (BMI), Oswestry Back Pain Disability Questionnaire (ODQ) scores, physical activity level, hip flexor muscle length, abdominal muscle force, and range of motion (ROM) for lumbar flexion and extension. RESULTS: In women, age, BMI, and ODQ scores were associated univariately and multivariately with pelvic inclination. In men, lumbar extension ROM was related univariately to pelvic inclination; age, lumbar extension ROM, and ODQ scores were associated multivariately. Lumbar lordosis was associated univariately with only lumbar extension ROM for women and men. A weak correlation was found between angle of pelvic inclination and magnitude of lumbar lordosis in standing (r=. 31 for women, r=.37 for men). CONCLUSION AND DISCUSSION: The odds ratio of having CLBP is increased if the score on the double-leg lowering test for abdominal muscles exceeds 50 degrees for men and 60 degrees for women. In patients with CLBP, the magnitude of the lumbar lordosis and pelvic inclination in standing is not associated with the force production of the abdominal muscles. PMID- 10696154 TI - Alterations in shoulder kinematics and associated muscle activity in people with symptoms of shoulder impingement. AB - BACKGROUND AND PURPOSE: Treatment of patients with impingement symptoms commonly includes exercises intended to restore "normal" movement patterns. Evidence that indicates the existence of abnormal patterns in people with shoulder pain is limited. The purpose of this investigation was to analyze glenohumeral and scapulothoracic kinematics and associated scapulothoracic muscle activity in a group of subjects with symptoms of shoulder impingement relative to a group of subjects without symptoms of shoulder impingement matched for occupational exposure to overhead work. SUBJECTS: Fifty-two subjects were recruited from a population of construction workers with routine exposure to overhead work. METHODS: Surface electromyographic data were collected from the upper and lower parts of the trapezius muscle and from the serratus anterior muscle. Electromagnetic sensors simultaneously tracked 3-dimensional motion of the trunk, scapula, and humerus during humeral elevation in the scapular plane in 3 handheld load conditions: (1) no load, (2) 2. 3-kg load, and (3) 4.6-kg load. An analysis of variance model was used to test for group and load effects for 3 phases of motion (31(-60(, 61(-90(, and 91(-120(). RESULTS: Relative to the group without impingement, the group with impingement showed decreased scapular upward rotation at the end of the first of the 3 phases of interest, increased anterior tipping at the end of the third phase of interest, and increased scapular medial rotation under the load conditions. At the same time, upper and lower trapezius muscle electromyographic activity increased in the group with impingement as compared with the group without impingement in the final 2 phases, although the upper trapezius muscle changes were apparent only during the 4.6-kg load condition. The serratus anterior muscle demonstrated decreased activity in the group with impingement across all loads and phases. CONCLUSION AND DISCUSSION: Scapular tipping (rotation about a medial to lateral axis) and serratus anterior muscle function are important to consider in the rehabilitation of patients with symptoms of shoulder impingement related to occupational exposure to overhead work. [Ludewig PM, Cook TM. Alterations in shoulder kinematics and associated muscle activity in people with symptoms of shoulder impingement. PMID- 10696155 TI - Pulsed lavage in wound cleansing. PMID- 10696156 TI - The new challenges of mental health nursing research and practice. PMID- 10696157 TI - The biopsychosocial perspective in psychiatric nursing: myth or future reality? PMID- 10696158 TI - [Psychosocial determinants of lithium compliance in patients with bipolar disorder]. AB - Physical, cognitive, and social factors play a central role in the lithium compliance of people with bipolar disorder. However, studies provide only a partial understanding of this phenomenon and there is currently no nursing model that takes into consideration a combination of factors. This study, based on Pender's preventive health beliefs model, was intended to identify the psychosocial determinants of lithium compliance. A random sample (n = 149) of outpatients at a large Montreal psychiatric hospital was used to measure lithium compliance on the basis of 5 criteria: compliance according to the nurse and according to the patient, appointment compliance, and compliance according to two criteria related to hyperuricemia. Polytomous logistic regression analyses were computed by regressing a composite of these criteria on sociodemographic variables and on the variables of the Pender model: susceptibility, seriousness, control over health, motivation to be healthy, perceived benefits and obstacles, and triggering factors. It appears that being female, being elderly, living with a partner, and perceived treatment benefits and obstacles are determining factors in lithium compliance. These results are all the more important in light of Quebec's newly implemented drug insurance plan, which could increase the obstacles to medication. Nurses will have to be increasingly vigilant with respect to these new obstacles and will have to adjust their interventions accordingly. PMID- 10696159 TI - Peer sexual harassment: a barrier to the health of adolescent females? AB - Despite increasing societal concern about sexual harassment in the workplace and in academia, to date sexual harassment has been neglected by nurses as a health issue among adolescents. Sexual harassment includes a wide range of unwelcome sexually oriented and gender-offensive behaviours that contribute to a hostile environment. Although the research is limited and lacking in rigour, early findings, along with evidence abstracted from the workplace-harassment and stress and coping literature, suggest that peer sexual harassment may adversely affect young women's mental and physical health, health-related behaviours, and future relationships. The author makes recommendations for further sexual-harassment research, specific to the adolescent population, based on a conceptual framework derived from the transactional stress and coping literature. PMID- 10696160 TI - Abused women's concerns about safety and the therapeutic environment during psychiatric hospitalization. AB - The purposes of this study were to identify the concerns of women who have a history of abuse regarding safety and the inpatient environment during psychiatric hospitalization, and to identify environmental changes they would like to see. A qualitative design was used to explore the women's concerns through semi-structured interviews. Instruments measuring sexual and physical abuse were administered. Of the 20 women recruited from 3 hospitals, 18 reported a history of sexual and/or physical abuse. One investigator interviewed the participants and one acted as recorder. After each interview, a list of identified concerns was generated; these concerns were raised in the next interview if not spontaneously brought up by the participant. Seventeen women reported feeling unsafe in mixed-gender units and said they would prefer segregated areas for programming and meals; 16 expressed concerns about nighttime routines and the traditional practices of restricting medications and contact with staff at night; 15 considered primary nursing extremely important to feeling understood and safe. The participants said they wanted to be heard and to be included in decision-making. PMID- 10696161 TI - From chaos to order: a nursing-based psycho-education program for parents of children with attention-deficit hyperactivity disorder. AB - A psycho-education program for parents of children with attention-deficit hyperactivity disorder (ADHD) is described. This intervention strategy is based on the nursing theory Modeling and Role Modeling developed by Erickson, Tomlin, and Swain to help people cope with stressors by facilitating the development of self-care knowledge and self-care resources, and by promoting self-care action. Key components of the program are an emphasis on pattern recognition and pattern management, the fostering of insider and outsider knowledge, and an emphasis on the strengths of participants and of their children with ADHD. PMID- 10696162 TI - Mother-loss: recreating relationship and meaning. AB - The purpose of this investigation was to describe adult women's experiences in losing their mother. Using an interpretive phenomenological frame of inquiry, 5 women were purposively selected to share their loss experience. Memories of the mother-daughter relationship were explored, and the meanings the daughters attached to their loss described, in written narratives, 2 in-depth interviews, and 1 group session. Seven themes emerged: Recalling, Holding On, Saying Goodbye, Longings of the Heart, Shifting Patterns of Relationship, Recreating the Dialogue, and Honouring Our Mothers/Ourselves. The loss of one's mother represents the loss of one's first intimate relationship, a relationship that has a unique meaning for daughters because their personal development is profoundly and uniquely shaped by it. This potentially pervasive and transforming life experience is best understood from an in-depth exploration and understanding of the nature of the mother-daughter relationship. Nurses who come to understand the dynamic interaction of grief and development through women's experience of mother loss can more successfully offer their presence, their understanding of the complexity of the mother-daughter relationship, and their skills in bereavement care to facilitate healing and to promote health and personal growth. PMID- 10696163 TI - Measuring the care needs of mothers of children with cancer: development of the FIN-PED. AB - This 2-phase study tested the Family Inventory of Needs-Pediatrics (FIN-PED), a 52-item instrument structured to include 2 subscales, the first measuring the importance of care needs and the second measuring the extent to which needs were met. In Phase I, an expert panel of 6 mothers of children with cancer rated the tool for clarity, apparent internal consistency, and content validity. All items met preset criteria for these assessments. In Phase II, 110 mothers rated the instrument for internal consistency reliability, stability over time, and internal construct validity. Both subscales achieved an estimated internal consistency of 0.94. Evidence of the instrument's stability over time was also achieved. Factor analysis resulted in 4 interpretable factors, suggesting that the tool is multidimensional. PMID- 10696164 TI - A new global resource on the Internet: Canadian-International Nurse Researcher Database (CNRD). PMID- 10696165 TI - Addressing the nursing shortage: researchers and clinicians unite. PMID- 10696166 TI - International nursing: the benefits of working together to improve nursing globally. PMID- 10696167 TI - Predictors of job satisfaction, turnover, and burnout in female and male Jordanian nurses. AB - As health-care systems undergo significant changes, the phenomena of job satisfaction, turnover, and burnout in nurses are of interest to nursing communities throughout the world. The purpose of this research was to examine these phenomena in a population of Jordanian nurses that is constituted of 25% men. This descriptive correlation study involved a sample of 479 nurses (68% female, 32% male) employed in public and military hospitals in Jordan, representing a 77% response rate to a survey. Significant differences were found between men and women for some of the items measured but not for turnover or burnout. However, regression analyses did demonstrate that selected variables impacted differently on men and women for the 3 outcome measures. Implications for nursing in Jordan are discussed. PMID- 10696168 TI - Student and faculty learning styles in a Canadian and a Chilean self-directed, problem-based nursing program. AB - A descriptive comparative study was conducted to identify and compare/contrast the learning styles of nursing faculty and entry-level students in 2 self directed (SDL), problem-based (PBL) nursing programs. The Kolb LSI-1985 was administered to 94 first-year generic students, 63 post-R.N. students, and 22 faculty members in a Canadian university nursing program. A Spanish translation was completed by 37 incoming nursing students and 13 faculty members in a Chilean university. One-way ANOVA analysis of group mean scores showed significant differences among the 4 student groups in the active experimentation learning mode. Post hoc tests confirmed that Chilean students are less likely to be active learners than their teachers or Canadian students, a finding of significance in preparing students to assume self-direction of their learning. Canadian faculty had higher abstract conceptualization scores than Chilean faculty, which has implications for faculty development of educator roles for SDL/PBL. PMID- 10696169 TI - An evaluation of WHA resolution 45.5: health human resource implications. AB - The World Health Assembly approved resolution WHA45.5 in 1992. This paper reports the findings of an evaluation of the implementation of this resolution using a survey technique. A total of 150 WHO Member States responded, for a 79% response rate. Findings suggest that the greatest strides worldwide have been made in education. While the data show that progress has been made at the country level, far more action is needed to strengthen nursing and midwifery if these cost effective resources are to play a decisive role in improving the extent and quality of services, especially as delivered to people in the greatest need. PMID- 10696170 TI - Nurse staffing and patient outcomes: evolution of an international study. AB - Industry-wide health sector reforms in the United States, Canada, and Europe have provided a unique opportunity to examine the effects of hospital restructuring on inpatient nursing care and patient outcomes across an array of settings. Seven interdisciplinary research teams--1 each in Alberta, British Columbia, England, Germany, Ontario, Scotland, and the United States--have formed an international consortium whose aim is to study the effects of such restructuring. Each site has enrolled large numbers of hospitals and nurses to explicate the role that organization of nursing care, a target of hospital restructuring, plays in differential patient outcomes. The study seeks to understand more fully the influence of both nurse staffing and the nursing practice environment on patient outcomes. Discussion of the theoretical foundation, study design, and process of developing the study instruments and measures illustrates the process to date, as well as the feasibility of and opportunities inherent in such an international endeavour. PMID- 10696171 TI - Health in the aftermath of violence: a critical narrative study of children of war and children of battered women. AB - Growing up amid violence has become reality for many children throughout the world. The health effects of this phenomenon have only recently begun to be addressed by researchers. However, there is growing evidence that children who witness violence suffer many of the same outcomes as those who experience violence directly. This critical narrative study examined the understandings and experiences of health and the relationship between violence and health. The sample, aged 10 to 17, comprised 2 groups of witnesses to violence: children of war and children of battered women. Analysis of the data revealed 4 categories: health as the absence of illness, health as a prerequisite for participation in desired activities, health as a holistic and multidimensional phenomenon, and health as a necessity for "getting through the day." While the first 3 ideas are consistent with those of children who have not lived amid violence, the 4th is unique to this population. Although no longer living in violence, the participants continued to face myriad physical and emotional health challenges. However, many also revealed an ability to heal. It is argued that violence and health cannot be separated, that exposure to violence has a profound and lasting influence on children's health beliefs and experiences. This paper addresses long and short-term strategies for intervention. PMID- 10696172 TI - [Supporting the conjugal system during the perinatal period: an experience in participatory research]. AB - This article presents the process and results of a participatory study intended to develop and evaluate preventive interventions for couples in the process of becoming new parents. A total of 21 participants, 4 physicians, 8 couples, and an investigator, studied the interventions using a research approach derived from a constructivist paradigm, the fourth generation evaluation. Employing a family intervention model, the nurse guided and contributed to the investigation. The results enabled physicians to refine their perinatal care and facilitated the couples' adjustment to the arrival of their first child. The interventions, the research process, and the use of a family nursing model are promising for nursing applications. PMID- 10696173 TI - Adapting the CAUSN accreditation process for emerging models of nursing education in Canada. AB - This paper highlights the accreditation issues raised by new and emerging models of baccalaureate nursing education and program delivery in Canada. It suggests ways of adapting the accreditation process to address recent changes. Nursing degree programs now offered by universities include programs at several sites, collaborative programs with partner institutions at multiple sites, and programs offered primarily through distance education. The accreditation program developed by the Canadian Association of University Schools of Nursing (CAUSN) provides a mechanism for monitoring the quality of a nursing education program and promoting the growth of the school that offers the program. Since the decision to undergo accreditation signifies a major commitment on the part of a nursing program, it is essential that the accreditation process be adaptable to meet the needs of evolving nursing education and program delivery models, and that it be fair, equitable, and credible. PMID- 10696174 TI - If values are communal, how is your pursuit of moral health care enhanced by examining the values of persons foreign to your community? PMID- 10696175 TI - Nurse practitioners in developing countries: some ethical considerations. AB - One of the principles of health care ethics is the principle of justice. An important expression of justice is equity. The provision of basic primary health care services to all people is the key to eliminating the gross inequities in health status existing in many countries. For many years nurses in developing countries have 'led the way' in bringing these essential services to poor rural communities, including the diagnosis and treatment of illnesses, and the prescribing and dispensing of medications. Nurses are the most appropriate health workers for this role, but most have not been prepared adequately for it. This is unsafe for patients and puts nurses at legal risk. Justice requires that patients should obtain access to safe health care and that nurses should receive appropriate education. Nurse practitioner programmes are being established to prepare nurses for this advanced practice role, but here again ethical considerations apply. Justice will be served only if nurse practitioner programmes are accessible to the nurses who are most likely to work in medically underserved communities where the need is greatest. PMID- 10696176 TI - The importance of ethics in the clinical supervision of nursing students. AB - This article investigates whether or not an ethical attitude manifests itself in the clinical supervision of nursing students. The data consist of 57 narratives written by nursing students, which were subjected to latent content analysis. The interpretation represents a caring science perspective based on Eriksson's 'caring ethics'. The results showed that some students received good supervision, while others felt hurt and humiliated. The students were of the opinion that they should feel welcome, be allowed to take responsibility and be treated as individuals by their supervisors. Supervision can take a form such that the ethical element comes to the fore in the conduct of the students as well as of their supervisor. Both the students' and the supervisor's disposition permeates all the questions, actions and reflections that form part of supervision, which could help to bring students' ethical sense to maturity. PMID- 10696177 TI - Unravelling the dilemmas within everyday nursing practice. AB - Each day, nurse practitioners are faced with clinical situations and dilemmas that have no obvious right answers. This article sets out the process of ethical mapping as a reflective device to enable practitioners to reflect on dilemmas of practice in order to learn through the experience and inform future practice. Ethical mapping is illustrated around a single experience that an intensive care practitioner shared in an ongoing guided reflection relationship. Within this process the practitioner draws on ethical principles to inform the particular situation, notably autonomy, doing harm, truth telling and advocacy. Through reflection, ethical principles are transcended and assimilated into knowing in practice, enabling the practitioner to become more ethically sensitive in responding to future situations. PMID- 10696178 TI - Shared teaching in health care ethics: a report on the beginning of an idea. AB - In the majority of academic institutions nursing and medical students receive a traditional education, the content of which tends to be specific to their future roles as health care professionals. In essence, each curriculum design is independent of each course. Over the last decade, however, interest has been accumulating in relation to interprofessional and multi-professional learning at student level. With the view that learning together during their student training would not only encourage and strengthen future collaboration in practice settings but also enhance patient care, the University of Dundee decided to run a pilot study to explore shared teaching in ethics between medical and nursing students. This article presents a report on the reasons for selecting health care ethics as a precursor for shared teaching, the educational tool used for the sessions, and the results of student and facilitator evaluation of the short course. Overall, despite problems such as poor attendance by some students, and facilitation and timetable difficulties, most of the feedback from students and facilitators has been positive. In essence the 'idea' has gone from strength to strength and there are now three levels of shared teaching in ethics between nursing and medical students, with plans to include further sessions with students from other disciplines. Within the text, 'health care ethics' will be referred to as 'ethics'; nursing students/nurses encompasses midwifery students/midwives. PMID- 10696179 TI - Elderly Japanese people living in small towns reflect on end-of-life issues. AB - This article, reporting on selected data from a larger study, discusses some responses to end-of-life questions that elderly Japanese people who were living in small towns gave in a questionnaire survey. Japan is now the country with the largest number of elderly people in the world and confronts numerous social and economic questions concerning how best to cope with its older population. Although it is a highly urbanized society, Japan also has large semirural areas. The focus here is on the questions in the survey that sought responses to ethical dimensions of end-of-life issues. The findings demonstrate the strength of traditional values that still exist throughout small towns in Japan. PMID- 10696180 TI - The bed crisis of winter 1995-1996 in the British NHS: an illustration of accountability issues. AB - The aim of this article is to explore the practical complexity of accountability in health care by focusing on a particular crisis affecting one NHS trust in the UK, that of insufficient beds to meet demand. It is presented through the eyes of five middle managers with nursing backgrounds. Although the focus is on their words, their expressions of distress and their awareness of conflict, these lead to a commentary highlighting some of the relationships between theory and practice, policy making and implementation, and, in the final analysis, compulsion and choice. The managers seemed to work within four main patterns of provider accountability: public, professional, pecuniary and personal. These four Ps of accountability created incompatibilities in the accountability process, but the conclusion attempts to draw the threads together to suggest a possible way forward. In order to protect confidentiality, pseudonyms are used for the NHS trust and the interviewees, and some personal details have been disguised. PMID- 10696181 TI - Informal coercion in the physical care of patients suffering from senile dementia or mental retardation. AB - This article discusses under what circumstances patients who are suffering from senile dementia or mental retardation should be submitted to coercive care, who should decide about this kind of coercion, and in what legal framework it should take place. A distinction is drawn between modest (i.e. of moderate degree) and meddlesome coercion. The use of modest coercion is defended. It is argued that medical personnel ought to decide exclusively about the use of modest coercion. However, no law should render legitimate the use of even modest coercion. It is conceded that to prohibit the use of a kind of coercion that is expected to take place is hypocrisy. It is argued that this is, however, an acceptable form of hypocrisy. PMID- 10696182 TI - The meaning of life. PMID- 10696183 TI - Transforming desolation into consolation: the meaning of being in situations of ethical difficulty in intensive care. AB - The purpose of this phenomenological-hermeneutic study was to illuminate the meaning of being in ethically difficult care situations. The participants were 20 enrolled nurses employed in six intensive care units in Sweden. The results reveal a complex human process manifested in relation to one's inner self and the other person, which transforms desolation into consolation through becoming present to the suffering other when perceiving fragility rather than tragedy. The main point of significance here is for all health professionals to create an ethical work environment and strive for praxis that fosters 'at-homeness', which renders us free to transform desolation into consolation. Consolation is of significance in ethics because it makes us available and helps us to fulfil the demands of life, while desolation makes us unavailable to others. PMID- 10696184 TI - Reflections on the meaning of care. AB - Health care is increasingly delivered by using medical technologies and specialized procedures. However, the systems through which it is delivered are coming under attack as lacking in care. Medicine is very capable of treating the human body, but it may be losing its sensitivity towards persons, especially concerning the vulnerability they are experiencing. Nurses are finding that the demands for more efficiency and cost-effective measures do not allow them sufficient time to offer the personal care for which they have always felt responsible. The question of the meaning of care surfaces especially in treating those who are approaching death. The need is to balance the various dimensions of care, keeping its focus on the well-being of persons. PMID- 10696185 TI - Nursing practice: compassionate deception and the Good Samaritan. AB - This article reviews the literature on deception to illuminate the phenomenon as a background for an appraisal within nursing. It then describes nursing as a practice of caring. The character of the Good Samaritan is recommended as indicative of the virtue of compassion that ought to underpin caring in nursing practice. Finally, the article concludes that a caring nurse, responding virtuously, acts by being compassionate, for a time recognizing the prima facie nature of the rules or principles of truth telling. PMID- 10696186 TI - Achieving moral health care: the challenge of patient partiality. AB - Illness and hospitalization are sources of vulnerability; they arguably endow nurses and midwives with the moral obligation to develop caring relationships with patients. Fairness and the equal treatment of patients are central to moral practice; current government publications are giving this political emphasis. This article argues that patient partiality is one factor that may result in insidiously unequal caregiving. Data generated during a qualitative study into professional caring suggest that patient partiality is an accepted part of everyday practice. Factors such as the patient's personality, nurse-patient familiarity and the perceived level of patients' understanding and interest in their illness emerged as possible sources of partiality and influence on practitioners' interactions with patients. The article argues that patient partiality can be managed morally if practitioners develop self-awareness and constantly reflect on the moral integrity of everyday practice. Throughout the article, unless it is stated that specific reference is being made to either nurses or midwives, reference to a nurse or practitioner denotes both. It is also emphasized that no implication is intended that any study participants provided unequal care. Rather, data are utilized solely to generate focused discussion around the concept of patient partiality. PMID- 10696187 TI - Intrusion into patient privacy: a moral concern in the home care of persons with chronic mental illness. AB - The aim of this study was to identify and analyse ethical decision making in the home care of persons with long-term mental illness. A focus was placed on how health care workers interpret and deal with the principle of autonomy in actual situations. Three focus groups involving mental health nurses who were experienced in the home care of persons with chronic mental illness were conducted in order to stimulate an interactive dialogue on this topic. A constant comparative analysis of the transcribed audiotaped sessions identified a central theme that concerned the moral symbolic meaning of 'home'. This reflected the health care workers' conflict between their professional role and their moral role, which they perceived as unclear. PMID- 10696188 TI - Changing values for nursing and health promotion: exploring the policy context of professional ethics. AB - In this article we illustrate, and argue for, the importance of researching the social context of health professionals' ethical agendas and concerns. We draw upon qualitative interview data from 20 nurses working in two occupational health sites, and our discussion focuses mainly upon aspects of the shifting 'ethical context' for those nurses with a health promotion remit who are working in the British National Health Service. Within this discussion we also raise a number of potentially substantive issues, including the risks of colluding in 'double standards', and the tensions between the practitioner and managerial roles in nursing. Overall, we hope to pose questions about the best ways to understand the ethical agency and responsibilities of health professionals. PMID- 10696189 TI - A nursing ethic: the moral voice of experienced nurses. AB - Nursing acts occur in thousands of instances daily, being a major component of professional health care delivery in institutions, communities and homes. It follows that the ethical practice of most nurses is put to the test on an everyday rather than an occasional basis. Hence, within nursing practice there must be a rich and deep seam of reflective interpretation and practical wisdom that is 'embedded' within the experiences of every experienced nurse. This article presents discussion on some of the main findings of a recently completed study on nursing ethics in New Zealand. An interpretation of a nurse's story taken from the study is offered and suggestions are made for nursing ethics education. PMID- 10696190 TI - Teaching nursing ethics by cases: a personal perspective. AB - This article is a reflection on the use of case study material in the teaching of ethics to nursing students. Given the main aims of a course in ethics for nurses and the limited effectiveness of formal moral theory, it seems inevitable that the mainstay of nursing ethics courses will continue to be case study material. This approach has recently been criticized on a number of grounds. The author suggests here that disquiet over teaching ethics in this way should motivate a concern not with whether, but how, teaching by cases is to be undertaken. PMID- 10696191 TI - The ethical issues with which nurses grapple are often brought to a point by polarizing the interests of nurses against those of doctors. PMID- 10696192 TI - The regulation of autonomy in nursing: the Italian situation. AB - We reflect upon the meaning of freedom and autonomy in nursing behaviour, attempting to outline the contemporary situation of nursing in Italy, where the profession is achieving important results after a long period of submission and subordination. The way to real emancipation is not easy, but a statement of law on the one hand--abolishing constraints such as the Mansionario--and professional self-regulation on the other--the recent new Deontological Code--represent a real conquest in that direction. However, no statement of law or deontology can be sufficient without deeper reflection by the profession on itself and its tasks, potentiality, social importance and self-esteem. PMID- 10696193 TI - Moral problems among Dutch nurses: a survey. AB - This article reports on a survey of the moral problems that Dutch nurses experience during their everyday practice. A questionnaire was developed, based on published literature, panel discussions, in-depth interviews and participation observations. The instrument was tested in a pilot study and proved to be useful. A total of 2122 questionnaires were sent to 91 institutions in seven different health care settings. The results showed that nurses were not experiencing important societal issues such as abortion and euthanasia as morally the most problematic, but rather situations such as verbally aggressive behaviour of colleagues towards patients, keeping silent about errors, and medical treatment given against the wishes of patients. Moral problems occurred especially when nurses experienced feelings of powerlessness with regard to the well-being of patients. Moreover, these moral problems proved to be related to institutional organization, leadership, and collaboration with colleagues and other disciplines. Nurses appeared to have a limited awareness of the moral dimensions of their practice. PMID- 10696194 TI - Informed consent for short-stay surgery. AB - This study in the context of short-stay surgery is based on a definition according to which informed consent consists of five elements: consent, voluntariness, disclosure of information, understanding and competence. The data were collected in four district hospitals in southern Finland by using a structured questionnaire. The population consisted of short-stay and one-day surgery patients (n = 107). Data analysis was based on statistical methods. The results indicated some problems in the realization of informed consent. Most commonly, consent was expressed by voluntary admission. Most patients had indicated their voluntary consent by making their decision independently. There were also certain problems with information. The respondents were least well informed about the drawbacks of anaesthesia and about alternative forms of treatment. The patients had not understood all the information they had received; problems of understanding were greatest with information about the advantages and disadvantages of anaesthesia. Problems were also reported with competence. These patients had the most difficulty in assessing the advantages and drawbacks of different treatments and anaesthesia. PMID- 10696195 TI - An Orwellian scenario: court ordered caesarean section and women's autonomy. AB - Between 1992 and 1996, a small number of women in the UK were forced by the courts to undergo caesarean section against their expressed refusal. Analysis of the reported cases reveals the blanket assumption of maternal incompetence and the widespread use of thinly veiled coercion. Such attitudes and practices are themselves frequently compounded by inadequate communication. Medical discretion in such problematic cases seems to err on the side of safety and so appears to favour the life of the fetus over maternal autonomy. Despite current policy's placement of the pregnant woman at the centre of maternity care, obstetricians' concerns appear to lie more with the unborn fetus. In other words, there seems to be a point at which the value of fetal life begins to outweigh, not so much the life of the woman, but her right to self-determination, her plans and her choices. While it is important to acknowledge that these court ordered caesareans represent an unusual extreme within contemporary maternity care in this country, that they have happened brings into sharp relief some of the stereotypical assumptions about women. These are assumptions that underlie much of current medical practice and may compromise or disempower women in other ways during their experience of pregnancy and labour. Using the first and last of the six reported cases as contextual illustrations, this article focuses on the complex interplay of processes that have brought the medical profession to a position in which their own self-conviction and determination to do what they believe is best for their patients has resulted in gross denial of women's autonomy and the use of the law to override pregnant women's refusal of consent. PMID- 10696196 TI - A study of the ethics of induced abortion in Korea. AB - The purposes of this study were to investigate the ethical aspects of induced abortion from the viewpoint of Korean women, and to compare and contrast their ethical considerations and values with the views of western ethical scholars. The two extremes of ethical arguments about induced abortion are pro-life and pro choice. However, the Korean women who participated in this study showed that conflicting ethical values were raised between the principle of caring and the sanctity of life or the principle of respect for the person, not between the right of self-determination and the sanctity of life. The results of the study suggest that it would be better to include the viewpoints of women in any ethical discussion on abortion in order for ethicists and health professionals to understand more fully the dimensions of moral clinical problems and be in a better position to discuss them in a practical manner. PMID- 10696197 TI - Visiting nurses' situated ethics: beyond 'care versus justice'. AB - This article discusses Dutch visiting (district) nurses' moral considerations of their daily work. It is based on an empirical study using extensive semistructured interviews. The study is informed by the theoretical debate on the 'ethics of care' and the 'ethics of justice'. It is argued that this debate easily turns into an unfruitful contest between these two perspectives: which one is best? The results suggest that visiting nurses' moral considerations of their day-to-day work can be described well in terms of an ethic of care. At the same time, however, concepts and issues central to an ethic of justice are also of crucial importance to their considerations. Nurses' ways of managing to combine both perspectives, even in situations of apparent conflict between them, are described. Thus, clues are provided on how the debate on the ethics of care and the ethics of justice may be carried out in a more fruitful way apart from through hierarchically opposing both perspectives. PMID- 10696198 TI - Being in the world of the suffering patient: a challenge to nursing ethics. AB - Ethics in caring is what we actually make explicit through our approach and how we invite the suffering patient into a caring relationship. This phenomenological study investigates suffering and health and how this presupposes a deeper reflection on ethics in caring. The aim was to try to discover, describe and understand how patients experience their life situation three years after undergoing surgery. The theoretical approach is based on central aspects of Eriksson's caritative theory (i.e. the view of the person as body, soul and spirit). The informants were four women and four men aged between 55 and 77 years. The empirical material revealed suffering that was connected with both illness and life. Suffering involves experiences of grief, loneliness and struggling. Health implies a yearning for something beyond the current life situation, a yearning to experience some meaning in life. This leads to an awareness of unplumbed possibilities. Understanding the experiences of individual patients demands of us, as both researchers and nurses, to act, seeking after the scientific truth (i.e. a deeper reflection of the ontological, epistemological and methodological questions). The idea of responsibility helps us to interpret and meet the innermost desires of suffering patients in their true presence. Caritative caring ethics means 'being there', confirming patients' absolute dignity; it is a manifestation of the love that 'just exists'. Compassion requires an inner disposition to go with others to the places where they are weak, vulnerable, lonely and broken. An ontology of caring provides both a starting point and a context for reflection about ethics and the ethical life. PMID- 10696199 TI - Being authentic and being a chameleon: nurse-patient interaction revisited. AB - This paper explores contradictory understandings of nurse-patient interaction arising through an exploration of 'being authentic' and 'being a chameleon'. The concepts arose during a critical praxis research study exploring nurse-patient relationships in the context of life-threatening or terminal illness. Being authentic can be understood as a dominant view in the nursing literature of the nurse-patient relationship, incorporating the value of being genuine. However, we argue that this concept offers only a partial and inadequate framework from which to understand nurse-patient interaction. The paper argues that nurse-patient relationships develop intersubjectively, with both the nurse and the patient choosing to reveal or conceal aspects of themselves in response to their interactions. Intersubjectivity as a concept provided the nurses in this study with a means for understanding how the nurse and the patient each contribute to interactions; nurse-patient relationships being understood as mutually constructed. These ideas are explored in this paper using examples from the nurses' stories, along with the implications raised for nursing practice. PMID- 10696200 TI - Human agency and the process of healing: lessons learned from women living with a chronic illness--'re-writing the expert'. AB - In this paper we examine the notion of human agency in the context of women experiencing a chronic illness. Based on two qualitative studies conducted with Canadian women of Chinese- and Anglo-descent living with diabetes, we unmask the complex power relations inherent in patient-practitioner interactions, and problematize the privileging of healthcare providers as knowers and experts on the patient's body. Specifically, we analyse the meanings that women ascribe to their illness experience. We discuss how women experience the loss of agency in healthcare encounters, how they resist patienthood by reclaiming agency, and how healthcare providers foster agency in their interactions with women. Rather than suggesting that biomedical experts should be 'written out', we propose to rewrite healthcare providers as 'reflexive practitioners' through the construction of transformative knowledge. We argue that praxis-oriented practice, which is based on transformative knowledge, will provide the space for women and healthcare providers to enter into a new dialogue and a relation in which women can sustain a sense of self, and begin the journey of healing. PMID- 10696201 TI - Transcending marginalization in knowledge development. AB - Quality care requires a body of knowledge that reflects the experiences and the responses of the marginalized populations to health and illness, and requires demarginalization of nursing knowledge. We argue the significance of developing an understanding of people who are marginalized, and organize our arguments and discussions into four sections: (i) developing knowledge that is not marginalizing; (ii) developing knowledge about marginalized populations; (iii) integrating nursing knowledge and making it visible; and (iv) the future of research enterprise. We propose that nurses critically consider strategies and processes to deal with and transcend marginalization of populations and of nursing knowledge. PMID- 10696202 TI - The need for psychiatric nursing: towards a multidimensional theory of caring. AB - Contemporary developments in health care have encouraged a review of the professional status of psychiatric nursing. Although research has documented psychiatric nursing activity, little study has been made of the 'need' for psychiatric nursing within a multidisciplinary service. Employing adapted grounded theory methodology, substantive theory was developed concerning the expressed need for psychiatric nursing, by patients, their carers and mental health professionals. The study was based on six sites from England, Eire and Northern Ireland. The study found some consensus across both recipients and providers of mental health care, that the essential feature of nursing (the core category) involved a complex of relationships: 'knowing you, knowing me'. Within that complex, nurses either elected, or were required, to move--or 'toggle'- between three discrete domains of relating: the Ordinary Me (OM); the Pseudo ordinary or Engineered Me (POEM); and the Professional Me (PM). Four internal dimensions involving the nurses' depth of knowing, power, use of time and use of translation distinguished these domains. The emergent theory extends current awareness of the importance of interpersonal relations in nursing. To what extent current health care policy, which emphasizes the promotion of alternative roles for nurses, will challenge this essential focus remains unclear. PMID- 10696203 TI - Kuhse on caring. AB - After a brief description of Helga Kuhse's arguments on the place of caring in moral decision-making in her recent book, Caring: Nurses, Women and Ethics, the author suggests that Kuhse is committed to reasons externalism and to a moderate form of motivational externalism. This compartmentalizes the psychology of moral agency and fails to see caring as the holistic stance toward patients which some nursing theorists have been espousing. The author then goes on to sketch such a holistic conception and to overcome difficulties relating to objectivity, the place of practical reason, and to weakness of will, which the new conception raises. PMID- 10696204 TI - Ideological implications of paradigm discourse. AB - Whilst the notions of paradigm and paradigm shift have become popularized in nursing's scholarly literature, there has been relatively little critical analysis of their impact upon theoretical understanding. In this paper, the authors attempt to deconstruct the ways in which paradigm discourse has been applied in nursing scholarship, looking beyond the claims that are made in the name of paradigm shifts to the apparent motivation underlying those claims. Comparing discourse associated with the paradigm shifts that have emerged in nursing education, research and practice theory, they reveal an inherently divisive purpose to which paradigm language is commonly used, and examine the implications of the discourse for nursing knowledge development. On the basis of this analysis, the authors urge a cautious approach to the extremes of paradigm claims, and argue for a more thoughtful and lively dialogue regarding the interests served by locating ideological positions within paradigm language. PMID- 10696205 TI - Risk and representation: older people and noncompliance. AB - In this paper it is argued that the way in which ageing is portrayed is culturally constructed and historically located. The terminology used to describe older people becomes part of nursing taxonomy but the issue that it is a cultural construct remains hidden. Nurses use histories and records as a means of communication about the condition of the client or patient. The notes mediate the way in which the patient comes to be known as an individual in need of services and help define the way in which nursing care is undertaken. Nursing notes help construct the 'manageable' patient. Noncompliance disrupts this notion of the manageable patient and it challenges medical and nursing dominance of the body. However, noncompliance then becomes reinterpreted as 'risk', which has the effect of extending the clinical gaze from the confines of the body to extra-corporeal spaces. Older people, because of their corporeality, can subvert and resist this clinical gaze. PMID- 10696206 TI - About 'using anthropology to analyse healthcare situations': a response to Monique Skidmore. PMID- 10696207 TI - De(con)struction of nursing work: economic rationalism and regulation. PMID- 10696208 TI - Psychiatric personnel, risk management and the new institutionalism. AB - This article reports the findings of a series of ethnographic research interviews conducted with psychiatric personnel in one region of Tasmania between 1995 and 1997. These interviews formed part of a more wide-ranging project examining changes in the regulatory practices of psychiatric personnel in the light of the professional, media and policy discourses that inform them, especially in relation to the impact of social justice reforms spelt out in recent Australian mental health policy. In discussing the nature of psychiatric work the personnel interviewed returned repeatedly to the themes of safety and risk management. The study presents an analysis of discourses deployed around these themes and argues that concerns over safety and risk are central to the emergence of a new institutionalism in acute in-patient psychiatric services. PMID- 10696209 TI - Foucault and nursing: a history of the present. AB - While a commitment to the development of nursing theory has been a significant force in nursing scholarship, particularly in the US, the authors have noted a recent trend among nurses in different countries to develop Foucauldian interpretations of nursing. The objective of this paper is to identify those publications by nurses that employ a Foucauldian perspective and to provide a useful summative review of these works to date, which illustrate the potential contribution of a Foucauldian reading of nursing. The authors have reviewed 27 publications written by nurses which present a Foucauldian analysis. These publications were issued between 1987 and 1998 in English, Portuguese and German. The most frequent concepts treated in the literature reviewed are power/knowledge, surveillance, discourse, discipline, resistance, docile bodies, clinical gaze, and panopticon. The literature reviewed illustrates that Foucault's concepts can have a profound impact on the way we conceive of nursing as a discipline and as a profession. Nursing care becomes a political event, nursing knowledge contributes to the dissemination of regimes of truth, and nurses, rather than being powerless, are perceived as professionals who exercise power over life in society. A Foucauldian reading of nursing enables nurses to move into a broader interdisciplinary and critical scholarship. PMID- 10696210 TI - Reflection as empowerment? AB - Reflective practice, as an ideal model, is generally espoused as a developmental process to empower practitioners to achieve and sustain effective practice. Yet when reflective practice is accommodated into the real world of everyday practice can this ideal itself be realised? Or will reflective practice be accommodated within existing norms whereby it becomes another technology of surveillance? The paper draws on dialogue taken from a guided reflection session to consider whether reflection can be empowering and to consider those factors which limit this potential. PMID- 10696211 TI - Nursing, social contexts, and ideologies in the early United States birth control movement. AB - Using historical discourse analysis, this study provides a thematic analysis of writings of nursing and birth control as found in The Birth Control Review from 1917 to 1927. The author contrasts this publication with the official journal of the American Nurses Association, the American Journal of Nursing from the same years to explore nursing voices and silences in early birth control stories. In dialogue with social contexts, nursing endeavors and inactivity have played important yet conflicting roles in the birth control movement in the United States. Nursing writings from the early twentieth century reflect eugenic beliefs, national fears of immigrants, and ambivalence about women's roles in society and the home. Nurses simultaneously empowered women to choose when to become pregnant and reinforced nativist and paternalistic views of the poor. PMID- 10696212 TI - Understanding the prattle of praxis. AB - This paper will examine and discuss some of the strengths and weaknesses of conceiving nursing as a form of praxis, encompassing within this, the idea that in order to conceive nursing as a form of praxis, reflection has to be considered a key component. It will be argued that praxis can (and should) become a practical process and that, when applied to one's own nursing practice, changes, reshapes and allows one to discover new meanings (or, draws out the meanings which were always there). Clearly there are many different forms, understandings and applications of the term praxis and this paper will examine some of the tensions and the nexus that exist. These claims will be supported by using personal-professional journal extracts as a catalyst, showing that there is potential for closing the theory-practice gap through more in-depth reflection, and that this examination using reflective techniques will demonstrate that nurses, by using this process develop their own implicit personal nursing theories. Using one's colleagues as a critical resource allows what might be described as ongoing reflection to occur, where critical friends in both theoretical and practice worlds act as a dialectical catalyst for growth and change, moving toward closing the theory-practice gap. PMID- 10696213 TI - Please don't call me 'dear': older women's narratives of health care. AB - This paper foregrounds some of the central themes that a group of older women in writing workshops identified as integral to their health and overall quality of life. In particular it looks at how these women felt about their relationship to professional health care providers, and the role that both play in the maintenance of the women's health and well-being. The research that it is based on comes from narratives and autobiographical stories written by these women, which aimed both to identify the issues and concerns that affect them as older women, and to position them centrally as agents and collaborators in intellectual work. Their challenging words disrupted conventional story lines about the experiences of growing older, and confronted the narrow range of negative images of ageing and the accompanying stereotypes that currently pervade our culture. PMID- 10696214 TI - How to stop the spread of toxocariasis. PMID- 10696215 TI - Assessing a sodium and fluid imbalance. PMID- 10696216 TI - Evaluating nutritional disorders. PMID- 10696217 TI - Action stat. Subarachnoid hemorrhage. PMID- 10696218 TI - Responding to winter emergencies. PMID- 10696219 TI - Nursing's top 10 rewards. PMID- 10696220 TI - Are we overlooking a hidden source of organs? PMID- 10696221 TI - Electrifying news about iontophoresis. PMID- 10696222 TI - Straight talk about MS. PMID- 10696223 TI - New drugs 2000. Part I. PMID- 10696224 TI - Aspergillus. PMID- 10696225 TI - Using intraosseous access in children. PMID- 10696226 TI - Documenting difficult patient encounters. PMID- 10696227 TI - Myths & facts ... about rectal catheters. PMID- 10696228 TI - Suctioning a tracheostomy tube. PMID- 10696229 TI - What's Vietnamese for "crocodile tears"? PMID- 10696230 TI - Sharing. Welcome to the "land of enchantment". PMID- 10696231 TI - Entacapone for Parkinson's disease. PMID- 10696232 TI - Prevention of malaria. PMID- 10696233 TI - Plan B: a progestin-only emergency contraceptive. PMID- 10696234 TI - Nucleolin promotes homologous DNA pairing in vitro. AB - We purified to near homogeneity a previously identified 100 kDa mammalian homologous DNA pairing protein. The purified 100 kDa protein also catalyzed high levels of cell-free homologous DNA recombination activity. This ATP-dependent activity was capable of forming conservative recombinant products between two circular, double-stranded DNA molecules. We were unable to detect any DNA polymerase, DNA ligase, or 5' or 3' exonuclease activity associated with this purified material. The purified 100 kDa protein bound silver nitrate as well as a monoclonal antibody specific for nucleolin. A recombinant protein comprised of the Escherichia coli maltos-ebinding protein fused to the carboxyl-terminal two thirds of human nucleolin possessed homologous DNA pairing activity. These data indicate that the 100 kDa homologous DNA pairing protein is nucleolin. The observation that nucleolin can carry out homologous DNA strand pairing in vitro raises the prospect that it may function similarly in vivo. PMID- 10696235 TI - Suppression of neoplastic transformation and regulation of cytoskeleton by tropomyosins. AB - Down regulation of Tropomyosins (TMs) is a consistent biochemical change observed in many transformed cells. Our previous work has demonstrated that Tropomyosin-1 is an antioncogene and it is a class II tumor suppressor. Using ras-transformed murine fibroblasts (DT cells), we have examined the effects of co-expression of two isoforms of TM on cell morphology, cytoskeleton and tumorigenecity. Enhanced expression of TM1, a suppressor of transformation, along with TM2 which is not a tumor suppressor results in the formation of well-organized microfilaments, a morphology that resembles normal fibroblasts, and suppression of tumorigenecity. Tumor formation in vivo was compatible with the persistence of high-level of TM2, but not TM1. Homodimers of TM1 and TM2 were observed in these cells. Thus, restoration of expression of TM1 and TM2 protein in ras-transformed cells suppresses the transformed phenotype with dramatic re-organization of microfilaments. These data show that TM2 cooperates with TM1 in the reorganization of microfilaments, while TM1 is a suppressor of the transformed phenotype. PMID- 10696236 TI - Inhibition of solid tumor growth by Fas ligand-expressing myoblasts. AB - A major problem with standard treatments of solid tumors such as chemotherapy is that the effects are not localized to the tumor. As a result, normal tissue function is often severely impaired. Here we show that myoblasts from skeletal muscle that have been engineered with retroviral vectors to express Fas ligand (FasL) have potential as site-specific anti-tumor agents. FasL-expression by myoblasts was previously shown to lead to neutrophil-mediated immunodestruction, both of the cells and the surrounding tissue. Moreover, myoblasts expressing FasL induced apoptosis in Fas-expressing human tumor cells in vitro. These findings led us to investigate the possibility that myoblasts expressing FasL could serve as anti-tumor agents acting by both apoptotic and immunological mechanisms. The C57BL/6 lpr/lpr mouse primary myoblasts either expressing or not expressing murine FasL were co-injected with Fas-positive or Fas-negative human rhabdomyosarcoma cells into the tibialis anterior of immunodeficient mice. After 19-31 days, FasL-expressing myoblasts resulted in a marked accumulation of neutrophils and inhibited tumor growth in every case. By contrast, control myoblasts did not prevent significant tumor growth. The status of Fas expression by the tumor tissue in vivo was confirmed by immunostaining tumor sections with antibodies against Fas. Tumor inhibition was observed regardless of the presence or absence of Fas on the tumor cells, suggesting that in vivo, the induction of a neutrophil response is remarkably potent and sufficient to inhibit tumors. PMID- 10696237 TI - Genome exposure and regulation in mammalian cells. AB - A method of measurement of exposed DNA (i.e. hypersensitive to DNase I hydrolysis) as opposed to sequestered (hydrolysis resistant) DNA in isolated nuclei of mammalian cells is described. While cell cultures exhibit some differences in behavior from day to day, the general pattern of exposed and sequestered DNA is satisfactorily reproducible and agrees with results previously obtained by other methods. The general pattern of DNA hydrolysis exhibited by all cells tested consists of a curve which at first rises sharply with increasing DNase I, and then becomes almost horizontal, indicating that roughly about half of the nuclear DNA is highly sequestered. In 4 cases where transformed cells (Raszip6, CHO, HL60 and PC12) were compared, each with its more normal homolog (3T3, and the reverse transformed versions of CHO, HL60 and PC12, achieved by dibutyryl cyclic AMP [DBcAMP], retinoic acid, and nerve growth factor [NGF] respectively), the transformed form displayed less genome exposure than the nontransformed form at every DNase I dose tested. When Ca++ was excluded from the hydrolysis medium in both the Raszip6-3T3 and the CHO-DBcAMP systems, the normal cell forms lost their increased exposure reverting to that of the transformed forms. Therefore Ca++ appears necessary for maintenance of the DNA in the more highly exposed state characteristic of the nontransformed phenotype. LiCl increases the DNA exposure of all transformed cells tested. Dextran sulfate and heparin each can increase the DNA exposure of several different cancers. Colcemid prevents the increase of exposure of CHO by DBcAMP but it must be administered before or simultaneously with the latter compound. Measurements on mouse biopsies reveal large differences in exposure in different normal tissues. Thus, the exposure from adult liver cells was greater than that of adult brain, but both fetal liver and fetal brain had significantly greater exposure than their adult counterparts. Exposure in normal human fibroblasts as revealed by in situ nick translation reveals a nuclear distribution pattern around the periphery, around the nucleoli and in punctate positions in the nuclear interior in parts of both S and G1 phases of the cell cycle. The same exposure pattern is duplicated by the pattern of DNA synthesis in S cells. It would appear that these nuclear regions represent positions of special activity. The previously proposed theory of genome regulation in mammalian cells is supported by these findings. The theory proposes that: a) gene activity requires exposure of the given locus followed by action of transcription factors on the exposed genes; b) the fiber system of the cell (cytoskeleton, nuclear fibers, and extracellular fibers) are required for normal exposure; c) active sites for gene expression and replication consist of the nuclear periphery where differentiation genes particularly are exposed; the nucleoli where at least some housekeeping genes are exposed; and possibly also punctate regions in the interior; d) noncoding sequences play a critical role in genome regulation, possibly including the transport of loci to be activated to appropriate exposure transcriptional and replicating locations. Cancer cells have lost specific differentiation gene activities, at least sometimes because of mutation of appropriate exposure genes; at least some protooncogenes and tumor suppressor genes are responsible for exposure and transport of specific differentiation gene loci to their appropriate exposure sites in the nucleus and for inducing exposure. PMID- 10696238 TI - Suppression of chimpanzee NORS in hamster/chimpanzee hybrid: report on cell line R48-26. AB - We present the first documented NOR suppression in a hybridoma other than man mouse for the hamster-chimpanzee hybrid cell line R48-26. Alu PCR and chromosome painting showed that in this cell line chimpanzee chromosomes 13-15-23 are maintained. NORs on chimpanzee chromosomes 15-23, whose presence was directly verified by FISH with H 28s rDNA, resulted inactive while telomeric rDNA on hamster chromosomes resulted active even if hamster chromosomes presented extensive rearrangements. We observed an all or nothing model in accordance with a model of regulation by selective transcriptional factors. The rearrangements of hamster chromosomes have not involved the location of NORs because they maintain a telomeric position. PMID- 10696239 TI - Chromosomal localization and genomic structure of the human arsenite-stimulated ATPase (hASNA-I). AB - The hASNA-I is a novel human arsenite-stimulated ATPase identified as the human paralogue of the ATPase component of the arsenite efflux system in E. coli. The hASNA-I has distinct biochemical properties and a dual nuclear and cytoplasmic distribution. Immunohistochemical staining showed a distinct pattern of hASNA-I expression in cells within normal tissues, and its overexpression in breast cancer. Recently, the yeast two-hybrid system has identified hASNA-I as a cellular partner of metallothionein II suggesting an additional role in Zn homeostasis and cellular detoxification. This report describes the assignment of hASNA-I to human chromosome 19 by somatic-cell hybrid PCR mapping, the isolation of a chromosome 19-specific cosmid clone, and the genomic structure and exon intron boundaries of hASNA-I. Our results indicate that the coding region of hASNA-I consists of 4 exons spanning 6 kb on band 19q13.3. These data will facilitate molecular analysis of the role of hASNA-I in human disease. PMID- 10696240 TI - The natural history of bladder cancer. Implications for therapy. AB - Transitional cell carcinoma of the bladder is comprised of a variety of cancer diatheses that manifest a spectrum of distinct biologic potentials. Although these diseases have traditionally been classified as "superficial" and "muscle invasive" on the basis of their histologic appearance (depth of penetration of the "bladder wall" and corresponding prognosis) the pathways presumably followed by the various forms of these cancers imply an even greater complexity. These disparate pathways may reflect different events in carcinogenesis, which may determine subsequent development and risk for either recurrence or progression. In addition, biologic activity and malignant potential for each type of cancer may be associated with distinctive molecular and genetic alterations. These considerations may provide an opportunity to expand traditional staging systems in creating molecular profiles that may more precisely characterize the biologic potential of these tumor diatheses. Although there are far more questions than answers concerning how these alterations may effect the natural history of bladder cancer, molecular-based identification of bladder cancer patients at greatest risk for progression may ultimately improve clinical management. PMID- 10696241 TI - Surgical management of noninvasive bladder cancer (stages Ta/T1/CIS). AB - Transurethral removal of bladder cancer is performed for diagnostic and therapeutic reasons. Tissue is returned for histologic determination of tumor grade and stage. For tumors that do not invade the detrusor muscle, resection can be curative although there is a high rate of new tumor occurrence. Surgical removal not only eradicates visible existing lesions but also retrieves material for histologic evaluation. This examination allows a determination of tumor grade as well as stage. Because of the importance of good histologic information in determining prognosis and, possibly, the need for further therapy, the method of surgical removal is important. PMID- 10696242 TI - Urine cytology. It is still the gold standard for screening? AB - Urine cytology remains the gold standard for bladder cancer screening. It is the test against which all others are compared when evaluating potential bladder tumor markers. The answer to whether urine cytology possess the optimal combination of sensitivity and specificity to retain consideration as the best screening device depends on the goals of the clinical practice. Urine cytology has excellent specificity with few false-positive cases. Its overall sensitivity is poor, but this drawback is explained for the most part by poor criteria for identifying well-differentiated, low-grade TCC. The natural history of such lesions is the occurrence of multiple superficial recurrences in 70% to 80% of patients, with only a minority (10% to 15%) progressing to muscle invasive or metastatic disease. Because patients with low-grade TCC are at low risk for progression, they are monitored primarily for the development of a subsequent tumor. One might argue that the detection of new low-grade lesions is of secondary importance to the early detection of disease progression. The performance characteristics of urine cytology in this regard are much improved. Urine cytology often results in the identification of high-grade malignant cells even before a cystoscopically distinguishable gross lesion is present. Routinely diagnosing grade I TCC may be clinically irrelevant. Ancillary techniques to improve the sensitivity of urine cytology have been insufficiently additive to have much clinical value. Several promising bladder tumor markers have been investigated as potential screening tools and are summarized in Table 3. BTA, nuclear matrix proteins, and fibrin/fibrinogen degradation products share lower specificities than urine cytology and may have high rates of false positivity. Telomerase is highly sensitive and highly specific but is not readily available as a point-of-service test. Hyaluronidase and hyaluronic acid are promising prognostic markers, but hyaluronidase does not detect grade I TCC. Early results from studies of this marker await verification. Combining some of these new markers may optimize their performance status, allowing the advantages of one test to correct the shortcomings of another. Likewise, their combination with urine cytology may prove beneficial. Although adding urine cytology has not increased the sensitivity of some point-of-service tests, few studies have addressed the effect on specificity. Until an obvious winner is declared in the race to find a bladder tumor marker, urine cytology will remain the gold standard screening method because of its comfortable familiarity. PMID- 10696243 TI - The utility of fibrin/fibrinogen degradation products in superficial bladder cancer. AB - Fibrin/fibrinogen degradation products are either absent or present at exceedingly low levels in the urine of healthy persons. Although various nonspecific inflammatory conditions of the urinary tract can result in detectable amounts of FDP in the urine, the presence of FDP is far more prevalent in urine from patients with bladder cancer. Urinary FDP levels tend to be higher in patients with tumors of increasing grade and stage. This correlation results in improved sensitivity in detecting more aggressive tumors. Current monoclonal antibody immunoassays are simple, rapid, and inexpensive, and can be performed on urine samples in the clinical setting. The overall accuracy of these immunoassays ranges from 75% to 80% (Table 1), suggesting that the urine FDP test should not be used alone for the surveillance of superficial bladder cancer. When assays for urine FDP are combined with urine cytology, the sensitivity for detecting tumors is improved. Prospective data are needed to determine whether using these tests in combination can safely permit a reduced frequency of endoscopic surveillance. PMID- 10696244 TI - Utility of nuclear matrix protein (NMP22) in the detection of recurrent bladder cancer. AB - This article examines the Nuclear Matrix Protein (NMP22) urine test for recurrent bladder cancer detection. Studies with NMP22 are compared to cytology for sensitivity and specificity. False positives and false negatives do occur, and consequences of these are discussed. Speculations are made regarding possible future uses of this test. PMID- 10696245 TI - HA-HAase urine test. A sensitive and specific method for detecting bladder cancer and evaluating its grade. AB - Hyaluronic acid and HAase are intricately associated with the biology of bladder tumor angiogenesis and metastasis. Tumor-associated HA and HAase are secreted in urine. In G2 and G3 bladder tumors, HA is degraded by HAase, resulting in the generation of angiogenic HA fragments, which, in turn, are secreted in urine. An elevated urinary HA level (> or = 500 ng/mg), indicating a positive HA test, suggests the presence of bladder cancer regardless of tumor grade. The urinary HAase levels correlate with the malignant potential of bladder cancer and are elevated (> or = 10 mU/mg) in the urine of patients with G2 and G3 bladder cancer. Combining the results from the HA and the HAase tests (the HA-HAase test) yields inferences, including the detection of bladder cancer and the evaluation of its grade. The overall 92% sensitivity of the HA-HAase test to detect bladder cancer is higher than the sensitivity of individual tests with little compromise in specificity. The HA-HAase test is equally sensitive for monitoring tumor recurrence. When compared with existing noninvasive tests, the HA-HAase test may be significantly less expensive and more accurate. PMID- 10696246 TI - The application of human complement factor H-related protein (BTA TRAK) in monitoring patients with bladder cancer. AB - The BTA TRAK assay is a quantitative sandwich format enzyme immunoassay performed in a reference laboratory. It measures levels of human complement factor H related protein (hCFHrp) which is similar to human complement factor H. Test characteristics reveal an overall sensitivity of 68% to 77.5% and a specificity of 50%-75%. Retrospective analysis of urine specimens employing hazard analysis suggests that BTA TRAK may have a role as a quantitative diagnostic marker in monitoring bladder cancer patients. Prospective studies are required to validate this application and other uses of the BTA TRAK assay in the management of bladder cancer patients. PMID- 10696247 TI - A critical analysis of the use of p53 as a marker for management of bladder cancer. AB - Delineating the important molecular pathways in carcinogenesis has helped develop and advance the field of molecular diagnosis. Bladder cancer has served as an excellent model in translating some of the advances from the laboratory to clinical settings. Many investigators have examined the use of p53 to help manage patients with bladder cancer who are at high risk of tumor progression. This article reviews the clinical studies that have used p53 as a marker in bladder carcinoma and concludes by determining whether routine assessment of the p53 tumor suppressor gene/protein is indicated at this time. PMID- 10696248 TI - Mutation of cell cycle regulators and their impact on superficial bladder cancer. AB - Early cytogenetic studies in bladder cancer identify regions of chromosomal gain or loss that can be candidate loci for oncogenes and tumor suppressor genes. Oncogenes with potential prognostic significance identified in bladder cancer the RAS family, epidermal growth factor receptor, ERBB-2, MDM2, and cyclin D1. The TP53 gene has been the most thoroughly characterized tumor suppressor gene in bladder cancer, with correlation of TP53 alterations with type of carcinogenic exposure, tumor stage and grade, as well as prognosis. Studies evaluating alterations of the retinoblastoma pathway have identified the retinoblastoma gene, RB, p161NK4A/CDKN2, and E2F-1 as tumor suppressor genes with potential prognostic significance in patients with bladder cancer. Better understanding of the genetic mechanisms underlying bladder tumor development and progression will allow better prevention, diagnosis, and treatment strategies. PMID- 10696249 TI - The potential role of gene therapy in the treatment of bladder cancer. AB - Based on understanding the molecular mechanism of bladder carcinogenesis, cancer gene therapy may well become a novel therapy in the near future. Currently, a viral vector system appears to be a better vehicle to deliver genes into target cells. Exploring different therapeutic strategies has generated promising results from preclinical bladder cancer models. Phase I clinical trials are underway to study the feasibility of this treatment for human bladder cancer patients. However, several potential problems associated with effective gene delivery need to be further refined. PMID- 10696250 TI - Telomerase in human bladder cancer. AB - Biomarkers for human bladder cancer are currently available and more are being developed. However, the ultimate goal of diagnosing bladder cancer consistently in a noninvasive fashion has not yet been achieved. Telomerase is an enzyme that may play a role in maintaining telomere sequences in the ends of chromosomes and its activity may reflect the presence of immortal or cancer cells. In this article, we reviewed the potential applications of telomerase in the diagnosis, monitoring, and treatment of human bladder cancer. PMID- 10696251 TI - An overview of the treatment of superficial bladder cancer. Intravesical chemotherapy. AB - Superficial bladder cancer accounts for approximately 70% to 80% of all newly diagnosed bladder cancers. The vast majority of these cancers are transitional bladder tumors of various histologic grades (I to III). Superficial tumors include carcinoma in situ (CIS), tumors confined to the epithelium (Ta), and superficial tumors that invade the lamina propria (T1) but do not involve superficial muscle layers. The primary treatment for eradication of stage Ta and T1 bladder cancers is transurethral resection of the tumor. Many patients with superficial bladder tumors treated with endoscopic surgery alone have recurrence or tumor progression at some point in their follow-up, and, in these patients, the need for adjuvant treatment becomes a major concern. PMID- 10696252 TI - Current use and questions concerning intravesical bladder cancer group for superficial bladder cancer. AB - Bacille Calmette-Guerin (BCG) is the most effective therapy for CIS of the bladder. Although several series have shown a decrease in recurrence and progression of T1 tumor, this effect is temporary. More than one half of patients with T1 tumors treated with BCG will progress over the longterm. A second course of BCG is indicated after an initial complete response. There is no definitive answer regarding the efficacy of maintenance therapy or the optimum dose of BCG. Randomized trials are needed to address these issues in a more conclusive manner. Phase III trials have shown that mitomycin C can be as effective as BCG in the management of papillary tumors; however, BCG is more effective in patients with CIS and high-risk superficial tumors. PMID- 10696253 TI - Interleukin-12. Opportunities for the treatment of bladder cancer. AB - Despite incomplete understanding of the human immune system, the rapid progress in tumor immunology provides a framework for more effective and safe interventions in the near future. Early approaches in patients with cancer that have focused on the nonspecific and broad stimulation of the immune system by interferons and IL-12 will be replaced by the highly specific stimulation of immune reactions targeting precisely defined tumor antigens. IL-12 has several biologic properties that seem useful in immune therapy for bladder cancer. The striking antitumor responses with IL-12 in preclinical animal models of bladder cancer provide optimism that future clinical trials involving this agent may impact on the risk and mortality associated with this disease. PMID- 10696254 TI - The potential application of Allium sativum (garlic) for the treatment of bladder cancer. AB - Additional studies are needed to identify the active ingredients in Allium Sativum (garlic) that are responsible for the observed antitumor activity and immune stimulation. Garlic seems to detoxify chemical carcinogens and prevent carcinogenesis and can also directly inhibit the growth of cancer cells. Current data suggest that low molecular weight sulfur compounds and protein F4 have immune-stimulation properties. Garlic is reported to stimulate immunity, including macrophage activity, natural killer and killer cells, and LAK cells, and to increase the production of IL-2, TNF, and interferon-gamma. These cytokines are associated with the beneficial Th1 antitumor response, which is characteristic of effective cancer immunotherapies. As is true of BCG, garlic stimulates the proliferation of macrophages and lymphocytes and protects against the suppression of immunity by chemotherapy and ultraviolet radiation. Garlic is clearly not a panacea for cancer, but its broad range of beneficial effects are worthy of serious consideration in clinical trials for the prevention and treatment of cancer. PMID- 10696255 TI - The role of photodynamic therapy in the treatment of recurrent superficial bladder cancer. AB - Photodynamic therapy is an exciting area of research for the treatment of superficial bladder cancer. Significant responses have been seen in patients resistant to standard intravesical treatments. New areas of research are focused on the development of new sensitizers and light distribution methods with less dermal and bladder toxicity. PMID- 10696256 TI - Interferons and bladder cancer. AB - With the introduction of BCG, intravesical instillation of immunotherapeutic agents has become a mainstay of therapy in the treatment of superficial bladder cancer. Interferon is capable of inducing a non-specific cellular and humoral immune response towards tumor cells. It has shown promise in reducing the recurrence and progression rates of superficial bladder cancer. In contrast to BCG, intravesical interferon is associated with minimal side effects and a very low dropout rate. Current research has focused on the use of interferon in combination with immunotherapeutic and cytotoxic drugs. PMID- 10696257 TI - Biological markers in superficial bladder tumors and their prognostic significance. AB - This article reviews the literature on some of the available biomarkers such as p53 and its down-stream effector p21 on superficial bladder tumor biology and their prognostic significance. The role of p53 tumor suppressor gene is controversial in superficial bladder cancer, possibly because analyzing one single effector of a pathway might hide the role of downstream effectors. The aggressiveness of this condition is related to proliferative activity as measured by Ki-67. Further studies are still necessary to draw definitive conclusions about the role of these different biological markers in superficial bladder cancer. PMID- 10696258 TI - Vascular endothelial growth factor and its correlation with superficial bladder cancer recurrence rates and stage progression. AB - Because of the heterogeneous behavior of superficial bladder cancer, the development of additional simple diagnostic and prognostic tests will be invaluable. The authors have demonstrated significantly elevated levels of urinary VEGF in patients with active bladder cancer. The sensitivity and specificity of urinary VEGF for diagnosing primary or recurrent bladder cancer were superior when compared with the results of cytology, which remains the most widely used noninvasive diagnostic investigation. These results and the authors' previous findings at the mRNA and protein level strongly implicate VEGF in the pathogenesis of bladder cancer recurrence and progression. The potential exists for anti-VEGF strategies in the treatment of, or prophylaxis against, recurrent superficial bladder cancer. PMID- 10696259 TI - Application of 1H NMR spectroscopy method for determination of characteristics of thin layers of water adsorbed on the surface of dispersed and porous adsorbents. AB - The paper presents 1H NMR spectroscopy as a perspective method of the studies of the characteristics of water boundary layers in the hydrated powders and aqueous dispergated suspensions of the adsorbents. The method involves measurements of temperature dependence proton signals intensity in the adsorbed water at temperatures lower than 273 K. Free energy of water molecules at the adsorbent/water interface is diminished due to the adsorption interactions causing the water dosed to the adsorbent surface freezes at T < 273 K. Thickness of a non-freezing layer of water can be determined from the intensity of the water signal of 1H NMR during the freezing-thawing process. Due to a disturbing action of the adsorbent surface, water occurs in the quasi-liquid state. As a result, it is observed in the 1H NMR spectra as a relatively narrow signal. The signal of ice is not registered due to great differences in the transverse relaxation times of the adsorbed water and ice. The method of measuring the free surface energy of the adsorbents from the temperature dependence of the signal intensity of non-freezing water is based on the fact that the temperature of water freezing decreases by the quantity which depends on the surface energy and the distance of the adsorbed molecules from the solid surface. The water at the interface freezes when the free energies of the adsorbed water and ice are equal. To illustrate the applicability of the method under consideration the series of adsorption systems in which the absorbents used differed in the surface chemistry and porous structure. In particular, the behaviour of water on the surface of the following adsorbents is discussed: non-porous and porous silica (aerosils, silica gels); chemically and physically modified non-porous and porous silica (silanization, carbonization, biopolymer deposition); and pyrogeneous Al2O3 and aluminasilicas. The effect of preliminary treatment of the adsorbent (thermal, high pressure, wetting with polar and non-polar solvents) on the characteristics of the structurized water layers was discussed. The influence of the adsorbent porous structure on the free energy of the adsorbed water was also studied. The discussion of the obtained results was made. PMID- 10696260 TI - Determination of interfacial tension from crystallization and dissolution data: a comparison with other methods. AB - Methods for the determination of interfacial tension between a solid and a liquid are reviewed including solubility/particle size, crystallization and dissolution kinetics. The use of solubility as a function of particle size, originally put forward by Ostwald and later corrected by Freundlich, may be unjustified for determining interfacial tension at solid-liquid interfaces. The interfacial tension values between solutions and sparingly soluble minerals such as hydroxyapatite, fluorapatite, brushite, octacalcium phosphate, calcium oxalate monohydrate, barium sulfate, calcium sulfate, calcite, and divalent metal fluorides are discussed. A comparison of these results is made with contact angle or wetting measurements. The interfacial tension values obtained from constant composition reaction kinetics are of the same order of magnitude as those determined using a contact angle method involving thin layer wicking techniques. PMID- 10696261 TI - Self-assembly of synthetic glycolipid/water systems. AB - Glycolipids (amphiphiles that bear oligosaccharides as their hydrophilic headgroups) are of importance both scientifically and technically. This review describes recent advances in our understanding of the molecular correlations in phase behavior in aqueous glycolipids over the past several years. In the first part, we discuss how headgroup stereochemistry affects the phase behavior of glycolipids both in two- and three-dimensional systems. In the second part, we discuss the effects of alkyl chain structure and phase behavior of phytanyl chained glycolipid/water systems. The physical properties of glycolipid/water systems depend strongly on the inter-headgroup interactions that are related to such factors as stereochemistry (conformation) and size of headgroups, type of sugar residues involved, alkyl chain structure, etc. Thus, apart from the conventional concept like 'hydrophilic/lipophilic balance', explicit accounts of headgroup interactions are crucial to control the particular glycolipid/water system concerned. This is in marked contrast to the conventional amphiphile/water systems where the inter-headgroup interactions are in most cases simply repulsive. PMID- 10696262 TI - The cleaning of polymer colloids. AB - The current state of knowledge of the cleaning of polymer colloids is reviewed with regard to a wide range of cleaning and characterisation techniques. The type, level and quantity of impurities involved with different polymer latex formulations varies widely. Even for similar formulations, differences in the nature and number of functional groups reported are often a consequence of sometimes subtle differences in the cleaning procedures employed. Not only may surface functionality be affected but also monomer and oligomer extraction procedures may lead to morphological changes in the particles. No single technique alone is likely to be able to remove all impurities. Care is needed to avoid the introduction of new impurities from the equipment, materials and water used as well as possible contamination from atmospheric carbon dioxide, bacteria and fungi. These factors also need to be considered in the storage of latex particle standards. PMID- 10696263 TI - Contaminant transfer during the coextrusion of tri-layer polymer films with a recycled layer. Effect of this transfer on the time of protection of the food. AB - Reusing old polymer packages as new food packages necessitates the coextrusion of tri-layer polymers, the old polymer being located between two virgin polymer layers. As it takes some time for the contaminant initially located in the old polymer, the virgin polymer layer acts upon the food pollution as a functional barrier. With the coextrusion process, the polymer layers are heated up to a high temperature and let cool down in air. During this coextrusion stage, the contaminant diffuses through the package at a fast rate over a short period of time. Then the time of protection of the functional barrier is significantly reduced. The process of the contaminant transfer is especially studied either during the coextrusion stage of the film or in the package-food system at room temperature. PMID- 10696264 TI - Measuring dissociation: comparison of alternative forms of the dissociative experiences scale. AB - The dissociative experiences scale (DES), developed by Bernstein and Putnam (1986), is commonly used to measure dissociation in clinical populations. It is often used with nonclinical populations to assess how levels of dissociation covary with other psychometric measures. When it is used with nonclinical populations, problems arise because the resulting scores can show severe floor effects and often are highly skewed. To remedy these problems, we developed alternative ways of measuring self-reported dissociative experiences. A form of the DES in which people were required to rate how often they have each of 28 experiences compared with other people was superior in avoiding problems of floor effects and skewness. We discuss situations in which this alternative, which we call DES C, is preferred. PMID- 10696265 TI - Word substitution errors in a speeded picture-word task. AB - Using a speeded-naming variant of the picture-word task, we found that word substitution errors can be elicited in the laboratory. In this variant of the task, participants often responded by saying the word instead of the picture's name. Such word substitution errors are interesting because they allow the evaluation of the relative merits of two broad classes of word production models. We obtained evidence in support of interactive models of word production. In addition, the error data presented evidence that speeded naming taps a late, name retrieval process and that picture naming is semantically mediated. PMID- 10696266 TI - Recall or evaluation of chess positions revisited: the relationship between memory and evaluation in chess skill. AB - We extend work by Holding and Reynolds (1982) on recall and problem solving with quasirandom chess positions. We tested 17 chess players on both quasirandom and structured chess positions. Consistent with the earlier study, initial recall of quasirandom chess positions is unrelated to chess skill level, and quality of the move selected in subsequent problem solving is related to skill level. However, recall following problem solving is related to chess skill level. These results support the view that pattern recognition processes underlie superior performance by skilled chess players, contrary to the conclusions of Holding and Reynolds (1982). Mechanisms such as long-term working memory retrieval structures (Ericsson & Kintsch, 1995) or templates (Gobet & Simon, 1996a) could explain the effective encoding of quasirandom positions during problem solving. PMID- 10696267 TI - Hemisphere specialization of Go experts in visuospatial processing. AB - To investigate hemisphere function of experts, Go experts and novices were given two Salthouse-type visuospatial tasks. In Experiment 1, stimuli of 4 digits in 6 cells were projected to the left (LVF) or right visual field (RVF). There was no prominent group difference in identification of digits and locations. In Experiment 2, stimuli of 4 digits in 16 cells were projected to the LVF or RVF. Go experts showed more accurate performance than novices. Both groups showed the same laterality, an RVF advantage, in the number identification. However, in the location identification, Go experts showed no visual field difference, whereas novices showed an RVF advantage. Based on these findings, the relationship between task demand and hemisphere function of experts is discussed. PMID- 10696268 TI - The effect of color intensity and appropriateness on color-induced odor enhancement. AB - Coloring solutions has been shown to increase perceived odor intensity. In Experiment 1, subjects rated the odor intensity of red strawberry and green mint solutions that had four levels of color intensity. Ratings of strawberry odor peaked at the medium color intensity and ratings of mint odor increased monotonically with color intensity. Thus, color-induced odor enhancement can increase with increasing color intensity but need not. Experiment 2 found that the color intensities producing maximum odor enhancement in Experiment 1 are not always the ones perceived as most appropriate for the odorants. Using different odors, Experiment 3 found that color intensity has some influence on the strength of the color-induced odor enhancement and color appropriateness has little. The presence or absence of color in the solution seems to be the most important variable. PMID- 10696269 TI - Toward a causal realist account of causal understanding. AB - Within a limited domain, humans can perceive causal relations directly. The term causal realism is used to denote this psychological hypothesis. The domain of causal realism is in actions upon objects and haptic perception of the effects of those actions: When we act upon an object we cannot be mistaken about the fact that we are acting upon it and perceive the causal relation directly through mechanoreceptors. Experiences of actions upon objects give rise to causal knowledge that can be used in the interpretation of perceptual input. Phenomenal causality, the occurrence of causal impressions in visual perception, is a product of the application of acquired causal knowledge in the automatic perceptual interpretation of appropriate stimuli. Causal realism could constitute the foundation on which all causal perception, judgment, inference, attribution, and knowledge develop. PMID- 10696270 TI - Representation of facial expressions of emotion. AB - Inversion interferes with the encoding of configural and holistic information more than it does with the encoding of explicitly represented and isolated parts. Accordingly, if facial expressions are explicitly represented in the face representation, their recognition should not be greatly affected by face orientation. In the present experiment, response times to detect a difference in hair color in line-drawn faces were unaffected by face orientation, but response times to detect the presence of brows and mouth were longer with inverted than with upright faces, independent of the emergent expression (neutral, happy, sad, and angry). Expressions are not explicitly represented; rather, they and the face configuration are represented as undecomposed wholes. PMID- 10696271 TI - Children's artistic responses to musical intervals. AB - In one experiment, White South African boys drew pictures in response to four musical intervals. In the second, the subjects were of both sexes and drawn from White, urban Black, and rural Black populations. Six intervals were used. Drawing content was similar cross-culturally. Consonances were perceived as generally positive; dissonances, generally negative. There was also an activity dimension. Children in a lower grade drew more concrete pictures than did those in a higher grade, regardless of age. Even young listeners were fairly consistent in their responses. This suggests that perception of musical meaning is a universal rather than culturally based phenomenon. PMID- 10696272 TI - Time course of attention effects with abrupt-onset and offset single- and multiple-element precues. AB - Large differences between the time course of attentional responsiveness to onset single-element precues (onset singles) and to onset multiple-element precues (onset multiples) have suggested differences in the way attention is controlled. In five experiments here, singles presented as offsets produced rapid attention buildup, attentional decay across longer precue-to-target delays, and attentional capture, as do onset singles, suggesting exogenous attentional control; both offset and onset multiples produced gradual onset of attentional effects without subsequent attentional decay, suggesting endogenous attentional control; and onset and offset singles produced higher accuracy than onset and offset multiples. Thus, the dynamic quality of a sudden onset is not sufficient explanation for the exogenous attentional control produced by a single-element peripheral precue. PMID- 10696273 TI - Protopopov's ideas on habit formation and their relation to the Pavlovian theory of higher nervous activity. AB - In 1929 Viktor P. Protopopov began to replicate E. L. Thorndike's animal experiments on habit acquisition. To determine the conditions necessary for habit formation, Protopopov used the natural experiment method, in which dogs encountered environments that prevented them from reaching a stimulus-bait. Not all dogs acquired the behavior necessary for obtaining the bait. Explaining the results within the framework of the Pavlovian theory of higher nervous activity, Protopopov concluded that habits were acquired when an active animal provoked by a bait-stimulus encountered an environmental barrier. The dogs tried a series of phylogenetic behaviors until the stimulus-bait was reached. The latter movements were retained, forming an ontogenetic habit. The dogs also learned not to produce the unsuccessful movements. In accord with the Pavlovian theory, individual differences in habit formation were related to temperament types. A critique of the Thorndikian Law of Effect is provided in terms of the Pavlovian theory of higher nervous activity. PMID- 10696274 TI - Cross-modality priming between odors and odor-congruent words. AB - It is well known that the Stroop effect is subject to influence by same-modality primes, but the possibility of cross-modal priming effects is unclear. Smell is a fundamental sensory system that is assumed to have potent cross-modality priming effects. We might expect the presence of a specific odor to interfere with performance on Stroop cards containing odor-congruent words. Forty participants, half of whom were primed with an unpleasant odor and half with a pleasant odor, were examined with modified Stroop cards containing pleasant and unpleasant descriptive words. A significant Stroop card by odor group interaction was found, indicating that the presence of an odor interferes with the performance on an odor-congruent Stroop card. These findings demonstrate cross-modality priming between olfaction and vision. PMID- 10696275 TI - Retinal location and its effect on the spatial distribution of visual attention. AB - A series of studies tested for distractor compatibility effects across wide target/distractor distances (0.6 degree to 20 degrees of visual angle). The effects of precue condition, constant/varied target location, horizontal/vertical distractor distance, and foveal/peripheral presentation were studied. Results show strong compatibility effects across wide distances when distractors are at peripheral retinal locations. When both stimuli were presented at the same peripheral location in opposite hemifields, compatibility effects were evident within an area of at least 2.5 degrees of visual angle. In contrast, when foveally placed distractors were used, compatibility effects were found primarily with target letters positioned near. The findings suggest that distance effects are not homogeneous across retinal location. PMID- 10696276 TI - Anticipated versus actual reaction to HIV test results. AB - The accuracy of predictions of how people will react to a medical test result is important because it may influence the decision to be tested. We hypothesized that people would overpredict their own long-term reactions to HIV test results (i.e., that they would feel better in response to seropositive results and worse in response to negative results than they expected to). In the first study phase, anticipations of reactions to positive and negative HIV test results were obtained from 50 subjects. In the second phase, postresult reactions were obtained about 5 weeks after subjects learned the results of their tests. The results suggest that people anticipate more distress given a positive result and anticipate less distress given a negative result than they experience. Cautions about the comparability of the 2 samples and recommendations for further research are discussed. PMID- 10696277 TI - Transfer of value from S+ to S- in simultaneous discriminations in humans. AB - When animals learn a simultaneous discrimination, some of the value of the positive stimulus (S+) appears to transfer to the negative stimulus (S-). The present experiments demonstrate that such value transfer can also be found in humans. In Experiment 1 humans were trained on 2 simple simultaneous discriminations, the first between a highly positive stimulus, A (1,000 points); and a negative stimulus, B (0 points); and the second between a less positive stimulus, C (100 points); and a negative stimulus, D (0 points). On test trials, most participants preferred B over D. In Experiments 2 and 3 the value of the 2 original discriminations was equated in training (A[100]B[0] and C[100]D[0]). In Experiment 2 the values of the positive stimuli were then altered (A[1,000]C[0]); again, most participants preferred B over D. In Experiment 3, however, when the values of B and D were altered (B[1,000]D[0]), participants were indifferent to A and C. Thus, the mechanism that underlies value transfer in humans appears to be related to Pavlovian second-order conditioning. Similar mechanisms may be involved in assimilation processes in social contexts. PMID- 10696278 TI - To reverse or not to reverse: when is an ambiguous figure not ambiguous? AB - The role of bottom-up processes in our perception of reversible figures was examined. In Experiment 1 the overlapping squares figure and nonsense reversible figures were used. The effects of adapting subjects for differing durations to an unambiguous version of the figure before presentation of the traditional reversible figure were determined under conditions of varying precision of fixation. In Experiment 2 the research was expanded to other examples of reversible figures. In both experiments, results with two dependent measures (the subject's first percept and the number of reversals reported) were generally consistent with the interpretation of bottom-up processes underlying the adaptation effects. However, the crucial role of stimulus and procedural variables and the differential sensitivity of the two dependent measures was demonstrated. PMID- 10696279 TI - Discriminating memories for actual and imagined taste experiences: a reality monitoring approach. AB - Blindfolded subjects tasted 4 common fruits and imagined the taste of 4 others while focusing on either a few (low sensory detail [SD] or many (high SD) of the fruit's sensory qualities. One week later, subjects judged whether each of 12 fruit names represented a fruit that was previously tasted, imagined tasted, or new (reality monitoring). The major finding was a significant interaction between source (imagined, perceived) and SD level (low, high). Source monitoring was accurate for imagined and perceived fruits in the low SD condition and for perceived fruits in the high SD condition. As predicted, subjects tended to misattribute memories for imagined fruits to perception in the high SD condition. The findings are discussed with reference to the Johnson-Raye reality monitoring model and recent work on memory source confusions. PMID- 10696280 TI - The picture superiority effect: support for the distinctiveness model. AB - The form change paradigm was used to explore the basis for the picture superiority effect. Recognition memory for studied pictures and words was tested in their study form or the alternate form. Form change cost was defined as the difference between recognition performance for same and different form items. Based on the results of Experiment 1 and previous studies, it was difficult to determine the relative cost for studied pictures and words due to a reversal of the mirror effect. We hypothesized that the reversed mirror effect results from subjects' basing their recognition decisions on their assumptions about the study form. Experiments 2 and 3 confirmed this hypothesis and generated a method for evaluating the relative cost for pictures and words despite the reversed mirror effect. More cost was observed for pictures than words, supporting the distinctiveness model of the picture superiority effect. PMID- 10696281 TI - Etiology and pathogenesis of obesity. AB - Obesity results from a greater consumption of energy than is used by the body. As this energy is stored, fat cells enlarge, producing the characteristic pathology of obesity. The pathologic enlargement of fat cells, in turn, produces altered levels of many peptide and nutrient signals that are responsible for the disease we call "obesity." The genetic makeup of human beings, which reflects a long history of relative scarcity of foodstuffs, has run into an age of surfeit, and many people cannot readily adapt. Thus, the increased intake of food does not signal satiety, and there is a gradual increase in energy stores as intake of energy outpaces need as we grow older. Against this background of struggle between nature and nurture, it is possible to identify an increasing number of defects or etiologies that produce obesity. For most patients, however, it is not possible to connect obesity to a specific cause. Leptin deficiency and defects in the leptin receptor both produce human obesity. Defects in the pro opiomelanocortin receptor system, the peroxisome proliferator-activated receptor gamma, the agouti-related peptide, and a few other rare genetic syndromes are also associated with human obesity. Of the genetic causes, Prader-Willi syndrome is the most common. Hypothalamic injury following craniopharyngioma is the most common neuroendocrine cause. Endocrine disorders such as Cushing's disease, polycystic ovary disease, and growth-hormone deficiency can lead to increased body fat. In the modern world, exposure to a high-fat diet predisposes many people to obesity, and this problem is compounded by the low levels of activity now required for daily living. Treatment strategies must be developed against this background. PMID- 10696282 TI - Obesity and its comorbid conditions. AB - Obese patients are at an increased risk for developing many medical problems, including insulin resistance and type 2 diabetes mellitus, hypertension, dyslipidemia, cardiovascular disease, stroke, sleep apnea, gallbladder disease, hyperuricemia and gout, and osteoarthritis. Certain cancers are also associated with obesity, including colorectal and prostate cancer in men and endometrial, breast, and gallbladder cancer in women (1-6). Excess body weight is also associated with substantial increases in mortality from all causes, in particular, cardiovascular disease. More than 5% of the national health expenditure in the United States is directed at medical costs associated with obesity (7). In addition, certain psychologic problems, including binge-eating disorder and depression, are more common among obese persons than they are in the general population (8.9). Finally, obese individuals may suffer from social stigmatization and discrimination, and severely obese people may experience greater risk of impaired psychosocial and physical functioning, causing a negative impact on their quality of life (10). PMID- 10696283 TI - Pharmacotherapy in the treatment of obesity. AB - The incorporation of pharmacotherapy into the comprehensive office-based management of obesity can improve outcomes significantly for patients meeting specific clinical guidelines in whom diet and lifestyle change alone is not successful. With an improved understanding of the mechanisms of action of various pharmacotherapies, it becomes possible for the primary care physician to help obese patients select successful therapeutic approaches and strategies. Some agents work primarily on central nervous system mechanisms related to satiety, while other agents affect metabolism and absorption of nutrients. It is of critical importance not to use pharmacotherapy alone, but rather combined with an aggressive program of diet and lifestyle change. The temptation of patients to find a magic pill that will close the refrigerator door is not likely to lead to a successful result. By gearing patients' expectations to realistic and achievable weight-loss goals, practitioners can reduce the risks of comorbid diseases, enhance ongoing treatment of existing comorbidities, and improve the overall quality of life for the obese patient. PMID- 10696284 TI - The central role of lifestyle change in long-term weight management. AB - Lifestyle change--most notably, modification of eating behavior, physical activity, and psychologic factors like attitudes, goals, and emotions--is the central determinant of whether people will lose weight and maintain the loss. Even when medical intervention appears to be the primary treatment, as with pharmacotherapy, behavior plays the determining role in successful weight loss. For example, the likelihood that a patient will take his or her prescribed medication is influenced by thoughts, attitudes, behaviors, and social environment. The patient who is dissatisfied with the moderate weight loss produced by most treatments, and hence is prone to relapse, is affected by the same factors. Methods are available to help people develop the self-management skills necessary to produce long-term lifestyle change (1-3). As individuals internalize a new set of attitudes and behaviors, resisting old habits and acquiring new ones become easier. Only then will there be any reasonable chance of long-term weight-loss success. PMID- 10696285 TI - Gastric ulceration: response to an unnatural environment. PMID- 10696286 TI - Normal equine gastroduodenal secretion and motility. AB - This article represents an attempt to provide an overview of the current knowledge of equine gastroduodenal secretory and motor activity, with respect to how these functions are controlled and interact. First, the equine gastric mucosal anatomy is discussed in comparison with other monogastric species, with some attention directed at the large nonglandular portion in relation to its function, or lack thereof. Next, control of gastric acid secretion, as assessed by the collection of gastric contents from a cannula or continuous measurement of their changes in pH, is reviewed, pointing out that there appears to be a relatively unobstructed movement of contents between stomach and duodenum in the horse, which can have a strong influence on pH of the gastric contents in particular. Then, methods of evaluating gastroduodenal motility, including recording of myoelectrical activity and changes in intraluminal pressure, and transit of radiolabelled contents from stomach into duodenum, are discussed. Finally, the apparent influence of the gastroduodenal migrating motility complex (MMC) on the composition of fasting gastric contents is introduced to underscore the observation that reflux of duodenal contents into the stomach is probably a common occurrence in the horse, particularly during periods of MMC-related cessation of antral motility. PMID- 10696287 TI - Pathophysiology of peptic disorders in foals and horses: a review. PMID- 10696288 TI - Comparative pathophysiology of nonglandular ulcer disease: a review of experimental studies. AB - Ulceration of the nonglandular, stratified squamous mucosa of the equine and porcine stomach is a common event in both species, although in pigs the fatality rate is significant and it is an economically important disease. Because the barrier function of this mucosa in horses and pigs appears similar, it is probable that similar pathophysiological mechanisms may be responsible for the initiating lesions and reparative events. Recent studies of ulcer pathogenesis in the pig have shown that feed preparation or prolonged fasting can result in disruption of the normal stratification of gastric contents, thereby allowing high concentrations of HCl, pepsin and refluxed bile to mix in the proximal stomach. Conditions simulating those found in vivo have been shown to damage this mucosa in vitro and indicate that luminal products, such as short chain fatty acids and bile salts, which act in synergy with HCl, probably are necessary to induce significant damage to this mucosa. Studies of the equine stomach have shown a similar proximal to distal pH gradient in the fed stomach, a significant duodenal-gastric reflux, and induction of squamous ulcers with fasting, thereby illustrating that similar conditions may be responsible for damage to the equine nonglandular mucosa. PMID- 10696289 TI - Role of duodenal reflux in nonglandular gastric ulcer disease of the mature horse. AB - Gastric contents were sampled in horses via nasogastric tube to determine changes in pH and bile salt concentrations during feeding and fasting periods. The horses were rotated through 4 feeding protocols. (1) hay; (2) hay with twice daily grain meals; (3) and (4) fasting preceded by either hay only or hay and grain. Sequential, hourly samples were collected from 3 horses prepared with gastric cannulas. Horses were fed hay twice daily and grain mix either twice daily or in small aliquots dispensed every 90 min. The horses were sampled during normal feeding or after 14 h of feed deprivation. Gastric pH values varied with time, but there was no significant difference between the feeding protocols or the fasting period on mean pH. Bile salt concentrations in fasted animals averaged 0.23-0.44 mmol/l with individual samples greater than 0.9 mmol/l. The bile salt concentrations in fed animals were consistently below 0.2 mmol/l. The effect of bile salt and acid on the stratified squamous gastric mucosa was tested in vitro. Mucosa, stripped of muscle and serosal layers, was mounted in Ussing chambers and the electrical potential difference (PD) across the tissue recorded. Sodium taurocholate or deoxycholate (0.3 mmol/l, bile salt) and/or HCl were added to the mucosal bathing solutions. The bile salt alone had no significant effect. Addition of acid (pH 2.5) to control tissues caused a decrease in the PD, which recovered within 20 min. Addition of acid to tissues exposed to bile salts resulted in a significant decrease in the PD, which did not recover. We conclude that combinations of bile salts and acid are more injurious to the stratified squamous gastric mucosa of the equine than acid alone. Concentrations of bile salts and acid sufficient to alter the electrolyte transport function of this mucosa can be found in the gastric contents of horses deprived of feed for as little as 14 h. PMID- 10696290 TI - Clinical syndromes of gastric ulceration in foals and mature horses. PMID- 10696292 TI - Induction and maintenance of gastric ulceration in horses in simulated race training. AB - Gastric ulceration is a prevalent condition of racehorses. A number of models of gastric ulceration have been described, but none mimic the conditions of a horse in training. The objectives of this study were to determine whether gastric ulcers could be induced and maintained in a group of horses in simulated race training. In addition, serum cortisol was measured on a weekly basis to investigate the possibility that stress may be important in the pathogenesis of gastric ulceration. Thirty horses used in the trial were fed Bermuda grass hay and 6 kg of a concentrate diet, and exercised 6 days/week at speed over a distance of 1.6-2.4 km. Serum was collected and gastroendoscopic examinations performed on a weekly basis for the duration of the trial. All horses developed moderate to severe ulceration, and ulcers were maintained for the 56 day period of the trial. Only one horse had signs of abdominal discomfort, which resolved with minimal symptomatic treatment and without the use of anti-ulcer medications. Serum cortisol remained within reference ranges for the duration of the trial. Although there was some variation between the weekly examinations, serum cortisol concentrations were decreased from values obtained at the start of the trial. In this study ulcers developed without the administration of nonsteroidal anti inflammatory agents or withholding of feed. This model provides a method to study the condition, and to investigate the effects of medications on the healing of ulcers in racehorses. PMID- 10696291 TI - Cross-sectional study of gastric ulcers of the squamous mucosa in thoroughbred racehorses. AB - Although gastric ulcers have been identified relatively frequently in racing Thoroughbreds, there have been no large scale studies to determine their effect on health and performance. Two hundred and two Thoroughbred horses in active race training were selected by the attending veterinarians for gastro-endoscopic examination. Images of the stomach mucosa were stored in a digitised format for subsequent evaluation. The number of ulcers and a score of severity were determined. Gastric ulceration of the squamous mucosa was identified in 82% of horses. Seventy-three (39%) horses displayed clinical signs consistent with gastric ulceration. Increasing Furr and Murray Score was associated with poor hair coat (P = 0.03), colic (P = 0.03), and increasing serum creatinine concentration (P = 0.029). There were no associations between haematology and serum biochemistry values (other than serum alkaline phosphatase concentration and serum creatinine concentration) and gastric ulceration. Our study confirmed the relatively high incidence of gastric ulceration in Thoroughbred horses involved in active race training. Gastric ulceration is a potential, but rare, cause of overt colic, but may produce more subtle detrimental effects on a horse's condition. It is concluded that the diagnosis of gastric ulceration should be based on an endoscopic examination of the stomach, although future studies are required to elucidate further the aetiology and clinical significance of gastric ulceration. PMID- 10696293 TI - A review of medical treatment for peptic ulcer disease. PMID- 10696294 TI - Effects of intramuscular omeprazole on gastric acid secretion in horses over a twenty-four hour period. AB - The effect of intramuscular (i.m.) omeprazole (0.25 or 1.0 mg/kg bwt; LD and HD), respectively, on volume, total acid output (TAO) and pH of the gastric juice was studied during 24 h in 5 horses with a chronically implanted gastric cannula. Whether secretion in controls was basal or stimulated with pentagastrin (8 micrograms/kg bwt/h), volume (NS) and TAO (P < 0.01, NS) gradually decreased and pH increased (P < 0.05, NS). Omeprazole significantly reduced the average basal TAO by 49 +/- 6% (LD) and 88 +/- 3% (HD) and the stimulated TAO by 64 +/- 2% and 97 +/- 1%. Basal pH in controls was 2.1-4.2 and after omeprazole treatment, pH 2.8-4.1 (LD) and 2.4-6.6 (HD). After stimulation, the corresponding pH values were 2.6-3.3, 3.9-4.9 and 5.4-7.2. The biological availability of omeprazole was 70-80%. Due to the simplicity of the administration technique and the higher biological availability, intramuscular administration may offer a practical and less expensive way of treating gastric ulcers in horses. PMID- 10696295 TI - Comparison of the antisecretory effects of omeprazole when administered intravenously, as acid-stable granules and as an oral paste in horses. AB - The antisecretory activity of omeprazole on gastric acid when administered i.v., intragastrically or per os, was evaluated in 2 female and 3 castrated male horses. Each horse had been prepared with a chronic indwelling gastric cannula. A single i.v. administration of omeprazole (1.5 mg/kg bwt) was effective in abolishing basal and pentagastrin (PG)-stimulated acid secretion. Once daily, nasogastric administration of omeprazole in acid-stable granules for 5 days inhibited acid secretion in a dose-dependent manner: 57% (1.5 mg/kg bwt) and 98% (5.0 mg/kg bwt) reduction of PG-stimulated acid secretion. The degree of inhibition was maintained over a 19 day treatment period with once daily dosing. A prototype oral paste formulation containing either acid-stable omeprazole granules or uncoated omeprazole powder was equipotent when compared to a similar dosage of acid-stable omeprazole granules administered by nasogastric tube. A dose-dependent inhibition was seen with the oral paste formulation containing omeprazole powder: 55% (1.5 mg/kg bwt) and 77% (3.0 mg/kg bwt) reduction of PG stimulated acid secretion after 5 days. Therefore, a paste formulation of omeprazole powder may offer an effective, easily administered, once daily acid inhibitory treatment for gastric ulcer disease in horses. PMID- 10696296 TI - Effect of omeprazole paste on gastric acid secretion in horses. AB - In a multicentre trial, 13 cannulated horses were treated orally once daily with a paste that delivered omeprazole at a dose of 4 and 5 mg/kg bwt in a 2-period crossover design to evaluate steady state gastric acid suppression. In each period, basal (unstimulated) and pentagastrin-stimulated gastric output were evaluated at 5-8 h after 5 doses, at 13-16 h after 10 doses, and at 21-24 h after 15 doses. Baseline data for gastric acid secretion were collected once for each horse in the month prior to initiation of omeprazole treatment. The inhibition of gastric acid secretion relative to baseline values, following treatment with omeprazole, were calculated and expressed as per cent. Pharmacokinetic data were also collected in this trial. At 4 mg/kg bwt, the oral paste formulation of omeprazole inhibited both basal and pentagastrin-stimulated gastric acid secretion by 99% at 5-8 h after treatment and by 83% (basal) and 90% (pentagastrin-stimulated) at 21-24 h. Inhibition following the administration of omeprazole at a dose of 5 mg/kg bwt was not significantly greater than when given at 4 mg/kg bwt. The results from this study could possibly lead to the development of an effective and practical antisecretory treatment of ulcer disease in horses. PMID- 10696297 TI - Safety of omeprazole paste in foals and mature horses. AB - Omeprazole has been shown to promote healing of spontaneously occurring gastric ulcers in horses when administered for 28 days at a dose of 4 mg/kg bwt/day and to prevent recurrence of ulcers in almost all horses when treatment is continued at a dose of at least 2 mg/kg bwt/day. The purpose of the 3 studies reported here was to 1) evaluate the evolution of potential effects of omeprazole paste (GastroGard), at a dose of 20 mg/kg bwt/day (5x the recommended dose) for 91 days in mature Thoroughbred horses; 2) evaluate the safety in young horses of omeprazole paste when dosed at 4 mg/kg bwt/day (1x), 12 mg/kg bwt/day (3x) or 20 mg/kg bwt/day (5x) for 91 days in Tennessee walking horse foals; and 3) evaluate the safety of omeprazole paste when dosed at 40 mg/kg bwt/day (10x) for 21 days in mature Thoroughbred horses. Within each study, horses were allocated randomly to the control or omeprazole paste treatment group. Clinical examinations, serum biochemistry and haematology were performed at regular intervals until necropsy at the end of the study. There were no treatment-related clinical signs in any treated horse and serum biochemistry and haematology were normal. In conclusion, omeprazole paste is safe for use in horses as demonstrated in studies with foals and mature horses. PMID- 10696298 TI - Safety, acceptability and endoscopic findings in foals and yearling horses treated with a paste formulation of omeprazole for twenty-eight days. AB - A paste formulation of the H+,K(+)-ATPase inhibitor omeprazole was evaluated in Thoroughbred foals and yearlings for its safety and acceptability. Twenty foals age 11-16 weeks and 20 yearling horses age 15-17 months were included and gastroscopic examinations performed 1-3 days before and at the end of each trial. Lesions were scored on a scale of 0 to 3 and animals allocated based on endoscopic lesion score and sex, with 4 animals in each of 5 replicates. Dosages of 4 mg omeprazole/kg bwt or sham treatment were administered once daily for 28 days, from a syringe graduated in 50 lb (22.68 kg) increments, the amount of paste administered being rounded up to the nearest corresponding weight in pounds. Acceptability of the paste or sham treatment was assessed and recorded by the individual administering the treatment on the basis of the tolerance or resistance to insertion of the syringe into the mouth, administration of the paste and if the paste was swallowed or actively expelled by the animal. Safety was determined on the basis of daily observation recordings and physical examination findings during and at the conclusion of the trial. Treatment was judged to have been accepted for all 420 doses of omeprazole paste and all 140 sham doses given to foals during the trial and for 418/420 doses of paste and all 140 sham doses given to yearlings. Two doses of paste were entirely rejected by yearlings. On the initial endoscopic examination, lesions were observed in the gastric squamous epithelial mucosa in 4 foals and 3 yearlings, and single small, superficial erosions were seen in the gastric glandular mucosa of 2 foals. On the second examination there were small, superficial erosions in the squamous mucosa in 3 foals and 2 yearlings, multi-focal superficial erosions in 1 foal and 1 yearling, and 1 foal had large areas of erosion extending from the margo plicatus toward the dorsal fundus. No lesions in the glandular mucosa were seen in foals or yearlings. There were no significant differences (P < 0.05) in lesion scores between the beginning and the end of the trials in the omeprazole-treated or sham treated groups of foals or yearlings. A paste formulation of omeprazole, administered at a dose of 4 mg/kg bwt once daily for 28 days, was determined to be highly acceptable to the foals and yearlings we studied, and no adverse effects attributable to the medication were noted. PMID- 10696299 TI - Acceptability of a paste formulation and efficacy of high dose omeprazole in healing gastric ulcers in horses maintained in race training. AB - Gastric ulceration has been found to occur in 80-90% of Thoroughbreds in active race training. Previously, variable success has been reported using mucosal surface protectants and H2 receptor antagonist. Omeprazole, a substituted benzimidazole, has been shown to inhibit gastric acid secretion in both man and animals. Fourteen horses, in active race training and with endoscopic evidence of moderated to severe gastric ulceration were divided into 2 groups: Group 1 (7 horses) were given placebo paste orally once daily for 28 days; Group 2 (7 horses) received 1.54 g active omeprazole in the placebo once daily for 28 days. Logs detailing administration and acceptability of the paste, and the horse's feeding and training regime were maintained by the trainer of each horse. Endoscopic examination of the stomach occurred at the beginning of the trial, and at 13-17 days and 27-31 days following commencement of the trial. Those horses that were free of ulceration on Days 27-31 were reexamined on Days 35-49. Acceptability of the paste, whether with or without active omeprazole, was deemed excellent in all horses except on one occasion, when one horse swallowed the paste following initial mild reluctance. Of the horses given the placebo (Group 1), 3 were withdrawn after the 13-17 day endoscopic examination: 1 horse to be given a H2 receptor antagonist, 1 horse was removed from training due to aryepiglottic entrapment and 1 horse had a greater than 10% fall in bodyweight from the start of the trial. Of the horses given active omeprazole (Group 2), one horses was relocated to another race track following the 13-17 day endoscopic examination. For the horses given placebo (Group 1), there was no change in the severity of ulceration. In contrast, the severity of ulceration in the horses given active omeprazole was significantly reduced at 13-17 days and 27-31 days. In 2 Group 2 horses, ulcers that had been completely eliminated subsequently returned when reexamined at 35-49 days. The results of this study suggest that omeprazole, employing a once daily dosing schedule, is effective at reducing the severity or eliminating gastric ulcers in Thoroughbreds in active race training. PMID- 10696300 TI - Effects of omeprazole paste on healing of spontaneous gastric ulcers in horses and foals: a field trial. AB - A multicentre, blinded, randomised complete-block, field trial was conducted with 140 horses and foals age 4 weeks-28 years to determine if omeprazole paste is effective and safe in promoting healing of spontaneous gastric ulcers under a variety of field conditions and in different breeds and ages of horses. Horses in the study had gastric ulceration as determined by gastroscopy and were divided into replicates of 4 or 5 animals. One horse in each replicate was assigned randomly to receive an empty omeprazole syringe (sham-dosed control) and the remaining horses received omeprazole paste once daily for 28 days. Gastroscopy was repeated at the end of the study. Horses treated with omeprazole had significantly (P < 0.01) more improvement in ulcer scores at the end of the study compared with controls. Ulcers were improved in 32.4 and 99.0% of the control and omeprazole groups, respectively. Ulcers were completely healed in 8.9 and 86.7% of the control and omeprazole groups, respectively. Under typical field conditions, omeprazole was effective at enhancing healing of spontaneous gastric ulcers in horses of a variety of ages and breeds. PMID- 10696301 TI - Efficacy of omeprazole paste in the treatment and prevention of gastric ulcers in horses. AB - Equine gastric ulcer syndrome (EGUS) is very common among performance horses, with a reported prevalence of approximately 90% in racehorses, and also > 50% in foals. Omeprazole, an acid pump inhibitor 5 times more potent than ranitidine, has been used with great success to treat EGUS. This multicentre study of Thoroughbred racehorses with endoscopically verified gastric ulcers was designed to demonstrate the efficacy of an equine oral paste formulation of omeprazole in the treatment and prevention of recurrence of EGUS. Of the 100 horses entered into the study, 25 were sham-dosed for the full 58 days of the study. The remaining 75 horses all received omeprazole paste, 4 mg/kg bwt/day once daily for 28 days. At Day 28, 25 of treated horses continued on this dosing regimen while 25 received a half dose (2 mg/kg bwt once daily) and 25 horses were sham-dosed. By Day 28, gastric ulcers were completely healed in 77% of omeprazole-treated horses, while 92% were significantly (P < 0.01) improved. In contrast, 96% of the sham-dosed horses still had gastric ulcers at Day 28. The improvement was maintained in horses that continued on either a full dose or half dose of omeprazole paste until Day 58. However, in those horses that were removed from omeprazole treatment at Day 28, the incidence and severity of the gastric ulcers at the end of the study were similar to those horses that did not receive the omeprazole paste. This study demonstrates that omeprazole paste, 4 mg/kg bwt per os, once daily, is highly effective in healing gastric ulcers in Thoroughbred racehorses and that either a full dose or half dose of omeprazole paste effectively prevents the recurrence of EGUS. The study also indicates that gastric ulcers in untreated horses did not demonstrate a significant rate of spontaneous healing. PMID- 10696302 TI - Clinical trial to determine the effect of omeprazole given once or twice daily on gastric ulceration. PMID- 10696304 TI - [Eletriptan: decisive advance in migraine therapy] [In Process Citation] PMID- 10696305 TI - [Basel weeks on liver diseases. Falk symposia nos. 114, 115 and 116, Basel, October 20-25 1999. The liver in focus of diagnosis and therapy] [In Process Citation] PMID- 10696303 TI - Treatment of gastric lesions in horses with pectin-lecithin complex. AB - This study compared the study of a pectin-lecithin complex (Pronutrin) on gastric ulcer healing during an 11 day period in 2 groups of 12 horses each. Twenty-four horses suffering from gastric lesions of differing severity were selected from a larger group of clinical cases on the basis of gastroscopic examination. Both gastric mucosal erosions as well as gastric ulcers were found in the 2 mucosal regions (pars nonglandularis and pars glandularis). The gastric mucosal lesions occurred predominantly in the pars nonglandularis in the vicinity of the margo plicatus. The 24 horses were divided equally into a treated group (Group A) and a control group (Group B). Twelve horses in Group A received Pronutrin, in a dose of 300 g/horse/day over a period of 10 days, whereas horses in Group B received no active substance. Gastroscopic examination was performed on Days 0 and 11. The degree of severity of the gastric erosions or gastric ulcers was evaluated independently in the 2 mucosal regions with the aid of a scoring system. Group A horses showed good acceptance of the product and no side effects were recorded. After the 10 day treatment phase, Group A showed a marked reduction in gastric mucosal lesions or disappearance of lesions, while untreated horses showed no change or, even, a deterioration on Day 11. Statistical calculation of efficacy revealed a highly significant reduction in gastric mucosal lesions in the pars nonglandularis and a significant reduction in gastric mucosal lesions in the pars glandularis in the treated horses. It would appear, however, that the treatment period of 10 days was too short, since the gastric mucosal lesions had often not completely healed in all horses. The results of this study in 24 horses with gastric lesions suggest that a pectin-lecithin complex can have a beneficial effect on the healing of gastric ulcers. PMID- 10696306 TI - [Paradigm changes in multiple sclerosis?]. PMID- 10696308 TI - [New perspectives in tumor therapy with erythropoietin]. PMID- 10696307 TI - [Modafinil in narcolepsy. A new therapeutic principle proves successful]. PMID- 10696309 TI - The management of acute angle-closure glaucoma. PMID- 10696310 TI - Acute acquired comitant esotropia. PMID- 10696311 TI - Acute angle closure glaucoma: an evaluation of a protocol for acute treatment. AB - PURPOSE: To report the use of a protocol for the treatment of acute angle closure glaucoma (AACG) and document its effectiveness. METHODS: Following a clinical audit, a formal protocol for the treatment of AACG was introduced in our department. Three and a half years later, the records of 63 consecutive patients were reviewed. A descriptive analysis was performed. RESULTS: At presentation the mean intraocular pressure (IOP) in the affected eye was 56 mmHg. The visual acuity was 6/60 or worse in 68% of patients. The mean duration to achieve adequate IOP control was 3 h (range 1-7 h) and 44% of the patients achieved this without the use of osmotic diuretics. None of the fellow eyes developed AACG prior to peripheral iridotomy. No further medical or surgical treatment was needed in 44%. Further topical treatment alone and surgical treatment were needed in 21% and 35% respectively. Following adequate IOP control, 76% had a visual acuity of 6/24 or better. All who had a final visual acuity of 6/60 or worse had significant non-glaucomatous pathology. At 6 months follow-up, 67% were treatment free and 65% had a normal optic disc. CONCLUSIONS: This study demonstrates that this treatment protocol provided comprehensive and explicit guidance on the emergency treatment of AACG. This resulted in rapid IOP control following presentation and eventual favourable outcome in most cases. PMID- 10696312 TI - Acute acquired comitant esotropia: a prospective study. AB - PURPOSE: To define the clinical characteristics of patients presenting with acute onset esotropia and features suggestive of possible underlying central nervous system pathology. To assess the prognosis for the return of binocular function and to consider the most appropriate management. METHODS: A prospective clinical study was carried out of all patients presenting to the department of paediatric ophthalmology at a university teaching hospital over the period January 1994 to April 1997. Each patient underwent a full ophthalmological examination (including assessment of sensory status). All patients were referred to a paediatric neurologist for examination and CT and/or MRI scan. RESULTS: Ten patients presented during the study period. Uncorrected hypermetropia and/or decompensated monofixation syndrome were the commonest aetiological factors. One patient was found to have a cerebellar tumour. In 5 patients prescription of the full hypermetropic correction alone was sufficient to restore binocularity. Five patients required bilateral medial rectus recession. Binocular function was restored in all cases--in 5 cases with bifoveal fusion. CONCLUSION: Decompensation of a pre-existing phoria or monofixation syndrome appears the commonest aetiology. Prescription of the full hypermetropic correction found at cycloplegic refraction forms an essential part of initial management. No single clinical sign can reliably indicate the rare patient harbouring a tumour. A high index of suspicion should be maintained and neuro-imaging considered in the absence of expected findings such as hypermetropia or fusion potential or in the presence of atypical features or neurological signs. PMID- 10696313 TI - Indocyanine green angiographic findings in idiopathic choroidal neovascularisation. AB - PURPOSE: The authors report the cases of two patients affected with idiopathic choroidal neovascularisation studied with combined fluorescein angiography and indocyanine green (ICG) angiography. In particular the presence of choroidal abnormalities at ICG angiography which could not be detected by fluorescein angiography was studied. METHODS: Both patients underwent a complete systemic and ocular assessment. Fluorescein angiography and ICG angiography were performed in a routine fashion at the time of presentation in both cases and after 14 months in the second patient. RESULTS: Results of the systemic investigations were unremarkable. A distinct dark rim surrounding the choroidal neovascular net was evident until the late phases of ICG angiography despite the presence of subretinal blood. Dilated choroidal vessels were observed beneath the neovascular membrane in both cases. In the first patient a hyperfluorescent area beyond the primary lesion was detected in the affected eye and a distinct leaking subfoveal choroidal venous vessel was found in the fellow eye. The second patient never showed other angiographic alterations either in the affected or in the fellow eye. CONCLUSIONS: ICG angiography has proved to be useful, both to better define and follow up the true extent of the pigment halo (healing response) around the neovascular membrane when subretinal blood and dye leakage at fluorescein angiography prevent its full appreciation, and to rule out other causes of choroidal neovascularisation in young healthy adults associated with either choroidal inflammatory focal lesions or choroidal vascular dynamic or inflammatory alterations. PMID- 10696314 TI - An electrophysiological follow-up study on acquired unilateral nyctalopia. AB - PURPOSE: To describe the clinical picture and electrophysiological findings in acquired unilateral nyctalopia. METHODS: A patient who had acquired unilateral visual loss with normal fundus was followed for a period of 2.5 years with basic ophthalmological examinations including standard electroretinogram and photopic on and off responses. RESULTS: A 46-year-old woman suffered from acquired unilateral nyctalopia. She complained of photopsia and blurred vision in her left eye. The initial examination of the left eye showed 1+ cells in the anterior chamber and a granular appearance in the fovea. After 1 month of treatment she still complained of photopsia in her left eye. Ophthalmoscopy and fluorescein angiography revealed no abnormality in either eye. A bright flash electroretinogram (ERG) in the left eye was a negative shape. Photopic ERG elicited by a 150 ms stimulus showed a depressed b-wave and enhanced a- and d waves in the left eye. CONCLUSIONS: This ERG waveform suggested that the transmission between photoreceptor and on-bipolar cell might be affected by idiopathic retinal disease. PMID- 10696315 TI - Primary vitrectomy for pseudophakic and aphakic retinal detachments. AB - PURPOSE: To evaluate primary vitrectomy for the treatment of pseudophakic and aphakic retinal detachments. Primary vitrectomy may enable better identification of retinal breaks than scleral buckling procedures. METHODS: A prospective study was performed of primary vitrectomy for the treatment of 25 consecutive cases of pseudophakic and aphakic retinal detachment. RESULTS: The primary retinal reattachment rate was 84% (21 eyes). Surgical failure resulted from new/missed retinal breaks (2 eyes) and proliferative vitreoretinopathy (2 eyes). The final retinal reattachment rate with further surgery was 96% (24 eyes). There were 7 macula-on detachments which all retained their presenting visual acuity. A visual acuity of 6/18 or better was achieved by 56% of the 18 macula-off detachments. Visualisation of the peripheral retina was impaired in 17 eyes and procedures to improve visualisation were performed in 7 eyes. Retinal breaks were detected in 16 eyes at surgery that had not been identified pre-operatively. Raised intraocular pressure was the most common complication, affecting 10 eyes in the early post-operative period. CONCLUSIONS: Primary vitrectomy offers certain advantages in the treatment of pseudophakic and aphakic retinal detachments. A controlled study is required to determine whether primary vitrectomy achieves a better outcome than scleral buckling procedures for these retinal detachments. PMID- 10696316 TI - Comparison of sub-Tenon's anaesthesia by different delivery techniques in cataract surgery. AB - PURPOSE: To compare the analgesic effects of three different delivery techniques of sub-Tenon's anaesthesia in cataract surgery by assessing patients' response to the visceral stimulus. METHODS: A prospective, randomised study was conducted on 345 eyes of 345 patients undergoing phacoemulsification and posterior chamber intraocular lens implantation. They received anaesthetic infiltration into the sub-Tenon's space through a conjunctival incision (115 eyes), infiltration into the posterior sub-Tenon's space (retrobulbar space) through a conjunctival incision (114 eyes), or injection into the intra-Tenon's space (subconjunctival space) without making a conjunctival incision (116 eyes). Pain scores were recorded when the anterior chamber was irrigated with an acetylcholine chloride solution to achieve miosis after lens implantation. RESULTS: There were no significant differences in pain scores among the three groups (chi-squared test of homogeneity, p = 0.814). Approximately 10-20% of patients reported slight to severe pain at the time of acetylcholine administration. CONCLUSIONS: The three anaesthetic delivery methods of sub-Tenon's anaesthesia possess similar and reasonable analgesic effects in cataract surgery, but may not block visceral stimuli completely. PMID- 10696317 TI - Ocular alterations in alopecia areata. AB - PURPOSE: To determine the ocular alterations occurring in alopecia areata with regard to the lens and fundus. METHODS: Seventy-five patients with alopecia areata were examined. Seventy healthy control patients unaffected by skin, ocular or systemic disorders were also studied. RESULTS: Symptomless punctate lens opacities were found in 38 (51%) patients, whereas only 2 (3%) control patients had similar lens changes. Fundus alterations were found in 31 (41%) cases of alopecia areata and in only 16 (23%) controls. CONCLUSIONS: These ocular alterations and their prevalence are reported and some theories regarding the possible aetiopathogenetic mechanisms are discussed. PMID- 10696318 TI - Glaucoma awareness and screening uptake in relatives of people with glaucoma. AB - PURPOSE: To assess glaucoma awareness and screening uptake in relatives of people with glaucoma. METHODS: A questionnaire was administered to 52 patients with primary open-angle glaucoma. They were asked about their awareness of glaucoma clustering within families, and the need for glaucoma screening in relatives of glaucoma patients. Patients were asked to identify one or more first-degree relatives, aged over 40 years and thus eligible for free glaucoma screening in the United Kingdom. These relatives were mailed a similar questionnaire. In performing the statistical analysis we corrected for possible clustering within families. The study was approved in advance by our local ethics committee, and all participants were informed of the United Kingdom's free screening service afterwards. RESULTS: Ninety relatives were identified, of whom 70 (78%) returned questionnaires. Only 53% of responding relatives thought they were at increased lifetime risk of developing glaucoma. Though 81% of relatives had been screened, many were screened infrequently. We compared the responses of patients' siblings and patients' offspring. Perceived lifetime glaucoma risk was similar in the two groups, but the (older) siblings had a significantly lower awareness of the free screening service (p = 0.03) and attended for screening less frequently (p = 0.07). Uptake of regular, free glaucoma screening at least every 2 years was 57% among offspring and 30% among siblings (p = 0.005). Because of selection bias (good communicators were more likely to be invited to participate) the true rates of glaucoma awareness and screening uptake are almost certainly lower than this. CONCLUSIONS: Relatives of people with glaucoma should be made more aware of the need for glaucoma screening and encouraged to use the free screening service. Older relatives should be particularly targeted. PMID- 10696319 TI - Ophthalmologists with conjunctivitis: are they fit to work? AB - PURPOSE: To establish current opinion as to whether ophthalmologists with conjunctivitis are fit to work. METHODS: One hundred and sixty ophthalmology units in the United Kingdom were sent a postal survey enquiring about work practices when an ophthalmologist contracts conjunctivitis. RESULTS: One hundred and five replies were received. Twenty-nine per cent of respondents said the ophthalmologist should stay off work while 7% said he or she should continue as usual. There was no concordance as to whether viral or bacterial conjunctivitis posed a greater problem. There is evidence that ophthalmologists have been implicated in the spread of epidemics of viral conjunctivitis but also reports of ophthalmologists who have continued to work with no reported spread of infection. CONCLUSION: In view of the lack of consensus the authors are unable to recommend evidence-based clinical guidelines, but would suggest that use of modern diagnostic laboratory techniques may help in making the decision as to whether to continue at work or not. PMID- 10696320 TI - Comparison of mydriatic efficacy of spray application and drop instillation of tropicamide 1%. AB - PURPOSE: To determine whether the mydriatic efficacy of spray application of tropicamide 1% is comparable to drop instillation of tropicamide 1%, and to compare the ocular discomfort caused by these methods. METHODS: Thirty-four healthy volunteers were randomly assigned to one of two groups, and received either a single drop of tropicamide 1% eye drops or a single puff of tropicamide 1% spray into open eyes. Pupil diameters were measured from anterior segment images taken using a Topcon Imagenet system at baseline and at the fifth, tenth and fifteenth minute after drug administration. Ocular discomfort experienced with each method was also compared. RESULTS: Repeated measures analysis of variance revealed that a statistically significant increase in pupil diameter was achieved with both application methods over time (p < 0.0001), and that there were no statistically significant differences in pupil diameter between the two groups at each time point (p = 0.409). The mean ocular discomfort score for tropicamide 1% spray was 1.45 +/- 0.56, and for tropicamide 1% eye drops was 2.71 +/- 0.67. This difference was statistically significant (p < 0.001). CONCLUSIONS: The mydriatic efficacy of tropicamide 1% spray is similar to that of conventional tropicamide 1% eye drops, and spray application causes less ocular discomfort. PMID- 10696321 TI - Aqueous humour levels of topically applied ciprofloxacin and ofloxacin in the same subjects. AB - PURPOSE: To evaluate aqueous humour levels of topical 0.3% ciprofloxacin and 0.3% ofloxacin in the same subjects. METHODS: Thirty-two bilateral cataractous patients received topical 0.3% ciprofloxacin in one eye and 0.3% ofloxacin in the other eye before each cataract extraction. Eyedrops were repetitively instilled for 6 h. Aqueous humour samples were collected and assayed for drug concentrations by a method described originally by us using high-performance liquid chromatography. RESULTS: Mean aqueous ciprofloxacin and ofloxacin levels were 0.33 +/- 0.04 microgram/ml (mean +/- SEM) and 1.34 +/- 0.14 micrograms/ml respectively (p < 0.0001). CONCLUSION: Ofloxacin level in the aqueous humour is 4 times higher than that of ciprofloxacin in the same subjects. PMID- 10696322 TI - Role of intravitreal dexamethasone in exogenous fungal endophthalmitis. AB - PURPOSE: To determine the effects of intravitreal dexamethasone in patients with exogenous fungal endophthalmitis. METHODS: Twenty cases of culture-proven exogenous fungal endophthalmitis following cataract surgery (11/20) and trauma (9/20) were retrospectively analysed for pre- and postoperative visual acuity, anterior chamber and vitreous inflammation and media clarity. All patients were managed with pars plana vitrectomy with intravitreal amphotericin B and oral ketoconazole with (steroid plus group) or without (steroid minus group) intravitreal dexamethasone. Results were analysed by Fisher's exact test. RESULTS: Following vitrectomy 9 of 20 patients (45%) achieved a visual acuity better than counting fingers at 3 m. No statistically significant difference was observed in anatomical and visual outcome between the steroid plus and steroid minus groups, though the number of patients with favourable visual outcome was greater in the steroid plus group. Rate of clearance of inflammation was better in the steroid plus group (40 +/- 15.5 vs 55 +/- 8.6 days). All patients (6/20) with pre-operative vision better than counting fingers showed good anatomical and visual outcome in both groups. CONCLUSIONS: The results of our retrospective study suggest that steroids may be beneficial in promoting faster clearance of inflammation in fungal endophthalmitis. Sensitivity of the fungi to antifungals, dose and timing of steroid and institution of effective antifungal medication prior to the use of steroids are the essential factors which need to be examined further in a prospective manner. PMID- 10696323 TI - Ocular leishmaniasis. PMID- 10696324 TI - Endoscopic transnasal removal of orbital foreign body. PMID- 10696325 TI - Multiple sclerosis presenting with debilitating Uhthoff's symptom. PMID- 10696326 TI - Congenital oculomotor palsy associated with brainstem hypoplasia. PMID- 10696327 TI - Transient fogging of acrylic (Acrysof) intraocular lenses. PMID- 10696328 TI - Pre-operative hyphaema in Fuchs' heterochromic uveitis. PMID- 10696329 TI - Self-induced cicatricial conjunctivitis. PMID- 10696330 TI - Periorbital necrotising fasciitis in a child. PMID- 10696332 TI - A case of trigeminal schwannoma presenting as Raeder's syndrome in a child. PMID- 10696331 TI - Traumatic prolapse of the globe into the maxillary sinus diagnosed as traumatic enucleation of the globe. PMID- 10696333 TI - Sinonasal melanoma: an unusual cause of proptosis. PMID- 10696334 TI - Recurrent optic disc edema with a macular star (ODEMS) PMID- 10696335 TI - Acute suprachoroidal haemorrhage in a patient treated with streptokinase for myocardial infarction. PMID- 10696336 TI - Valsalva retinopathy associated with blowing balloons. PMID- 10696337 TI - Pedunculated dermolipoma with overlying upper lid coloboma and absent lateral canthus: cause and effect. PMID- 10696338 TI - A case of pseudo-Terson's syndrome. PMID- 10696339 TI - Paroxetine and acute angle-closure glaucoma. PMID- 10696340 TI - Treatment of recurrent hordeolum with Broncasma Berna. PMID- 10696341 TI - Creutzfeldt-Jakob disease presenting with visual disturbance. PMID- 10696342 TI - Indocyanine green angiographic imaging of central retinal artery occlusion with cilioretinal sparing. PMID- 10696343 TI - Bilateral choroidal metastases as the first sign of metastatic gestational choriocarcinoma. PMID- 10696344 TI - Central retinal artery occlusion associated with primary antiphospholipid syndrome. PMID- 10696345 TI - Posterior dislocation of Staar plate haptic silicone lenses following Nd:YAG capsulotomy. PMID- 10696346 TI - Primary orbital Ewing's sarcoma: report of a case and review of the literature. PMID- 10696347 TI - Posterior sub-Tenon's triamcinolone injections in the treatment of uveitis. PMID- 10696348 TI - Genetic predisposition to ocular melanoma. PMID- 10696349 TI - Capsulorhexis phymosis following uncomplicated phacoemulsification surgery. PMID- 10696350 TI - Control of MRSA: too much or too little? PMID- 10696351 TI - From basic science to clinical practice: innovations in cancer therapy. PMID- 10696352 TI - Genetics of cancer: current knowledge and future possibilities. AB - For many years there has been scepticism among clinicians and cancer epidemiologists as to whether cancer was a genetic disease or could be hereditary. It is only in the last 10 years that developments in molecular biology have proven the hereditary nature of a small proportion of certain common cancers. That cancer is now indisputably 'genetic' at the cellular level is beyond dispute. PMID- 10696353 TI - Mechanisms and clinical implications of apoptosis. AB - The importance of controlled cell death in a variety of diseases has now been recognized and apoptosis research has become one of the fastest growing fields in medical science. Scientific advances are rapidly being applied to clinical practice. PMID- 10696354 TI - New developments in radiation treatment. AB - Radiotherapy is a key modality in the treatment of patients with cancer. Recent trials have extended its role. Advances in the technology of radiotherapy delivery and in the molecular basis of radiation treatment offer the prospect of further improvements in treating these patients. PMID- 10696355 TI - BPL plasma products: new source, enhanced products. AB - Bio Products Laboratory (BPL), the main fractionator of plasma-derived medicines, are undergoing a change from UK plasma to plasma from the USA. BPL aim to maintain continuity of supply and a seamless changeover, despite meticulous cleaning of its production plant and months of auditing by both the organization and regulatory authorities. PMID- 10696356 TI - Drug-induced oesophageal injury. AB - Drug-induced oesophageal injury is probably more common than previously thought. It is a major cause of morbidity and in some cases mortality. This article reviews the studies over the years, looking at factors affecting the transit time of pills, injury caused and ways to prevent it, and highlights the salient points with a case study. PMID- 10696357 TI - Neuroaxial blocks and LMWH thromboprophylaxis. AB - In the last 12 months, the Food and Drug Administration issued two documents warning the medical profession about the concurrent use of low molecular weight heparins (LMWHs) and neuroaxial blocks. This article summarizes the American and European experiences with LMWH thromboprophylaxis that could help with risk assessment. PMID- 10696358 TI - What goes wrong at a disaster or major incident? AB - From a receiving hospital's perspective a major incident is 'an accident about to happen'. The potential for chaos is present at every phase. This short paper describes in a chronological fashion the common traps and pitfalls and how to avoid them. PMID- 10696359 TI - Battered woman syndrome. AB - Recent judgments in the Court of Appeal have highlighted the significance of battered woman syndrome. This article describes the origin and features of the syndrome and some of its shortcomings. Medical aspects and legal applications of the concept are discussed. PMID- 10696360 TI - Responding to concerns about childhood emotional abuse. AB - Doctors are increasingly involved in child protection proceedings which consider whether emotional abuse has occurred. This may be a complex and difficult area, hindered by the paucity of consensus opinion and research in this area. This review suggests how doctors can assist in the decision-making process and outlines the management of children and adults who suffer sequelae of emotional abuse. PMID- 10696361 TI - Getting to grips with systematic reviews and meta-analyses. AB - Systematic reviews and meta-analyses now form a major source of evidence on clinical effectiveness. Despite the relative recency of this approach to synthesizing research findings, a reasonable consensus has emerged as to the desirable methodological characteristics of this secondary research. Understanding these basic tenets enables readers to take a balanced view of published review findings. PMID- 10696362 TI - Improving 'new deal' shifts for junior house officers. AB - Since the 'new deal' was introduced, new working patterns such as shift systems have become increasingly common. The current study examines how these shifts might be improved. The results suggest that organizational changes such as increased numbers of ward clerks and support workers can minimize the negative impact of shift systems. However, shifts remain unpopular among junior staff and the authors conclude that new working initiatives must be properly evaluated before their widespread implementation. PMID- 10696363 TI - Spontaneous oesophageal perforation: long known but still not easy to diagnose. PMID- 10696364 TI - Features in radiology: the double bubble. PMID- 10696365 TI - Tumour dislodgement to contralateral lung during double lumen tube insertion. PMID- 10696366 TI - Unnecessary operations: a medical triumph? PMID- 10696368 TI - Diagnosis and imaging of miliary tuberculosis. PMID- 10696367 TI - Diagnosis and imaging of miliary tuberculosis. PMID- 10696369 TI - Future of gene therapy. PMID- 10696370 TI - The three ages of a doctor's income. PMID- 10696371 TI - Emergency caesarean section and suxamethonium apnoea. PMID- 10696372 TI - A morphological and morphometric study of the peripheral process of the human vestibular nerve following posterior cranial fossa neurectomy. AB - Three vestibular nerve specimens removed at transmeatal neurectomy were studied in order to understand better retrograde degeneration and regeneration after vestibular neurectomy in the posterior cranial fossa. In two cases this procedure followed retrolabyrinthine retrosigmoid posterior fossa vestibular neurectomy. The subjects, three patients with Meniere's disease, were compared with one another and two autopsy controls with no known otological problem. The specimens were obtained at the distal end of the internal auditory canal and transversely sectioned. Many collapsed Schwann cell basement membranes were observed. The ratio of small-diameter nerve fibres increased significantly after neurectomy. Onion bulb formation around myelinated nerve fibres with small diameters and Schwann cell proliferation around the soma of vestibular ganglion cells reflected remyelination. We conclude that peripheral processes of vestibular nerve fibres can undergo retrograde degeneration and subsequent regeneration after transection of the central process. PMID- 10696373 TI - Enhancing hearing preservation in endoscopic-assisted excision of acoustic neuroma via the retrosigmoid approach. AB - Surgeons using the operating microscope are able to make use of numerous landmarks described for the lateral limits of dissection to preserve hearing in acoustic neuroma surgery via the retrosigmoid approach. Similar landmarks for hearing preservation described specifically for the endoscopic-assisted technique, are lacking. By analysing computed tomography (CT) scans of temporal bones, it was observed that to reach within 3 mm of the lateral end of the internal auditory meatus (IAM) via a 3 cm retrosigmoid craniotomy, drilling should be up to about 3 mm medial to the opening of the vestibular aqueduct. It was hypothesized that in surgery, by keeping 3 mm medial to the opening of the vestibular aqueduct, the integrity of inner ear structures would be preserved. This hypothesis was tested in 30 temporal bones and was found to be true. In addition, the lateral end of the IAM up to the transverse crest could be viewed by the 30-degree rigid angled endoscope. This landmark could, therefore, be utilized in the endoscopic-assisted technique to predict the optimal amount of bone to be removed at a stage before the internal auditory meatal dura is opened when the intact dura affords added protection to the meatal contents during drilling. Well designed dural flaps on the posterior petrous bone could be created by making a longitudinal incision not more than 7 mm from the superior border of petrous bone and a transverse incision at least 17 mm from sigmoid. These flaps minimize injury to the endolymphatic sac and protect the cochlear nerve and vasculature that when damaged, may result in hearing loss. PMID- 10696374 TI - Paraganglioma as a systemic syndrome: pitfalls and strategies. AB - Tumours of the neuroendocrine system in the head and neck region are mostly paragangliomas of the glomus tympanicum or jugulare, or of the carotid body. The majority of these tumours are benign, and the coexistence of multiple paragangliomas seems to be rare. Pre-operative embolization and surgery are regarded as primary therapy for these tumours. The treatment regimen in any patient depends on age, general health, hearing status and the function of the lower cranial nerves. Several presentations are possible in which paragangliomas occur as systemic disease. 1. Paragangliomas may occur bilaterally, or, in rare cases, in multiple areas. Pre-operative bilateral angiography is of utmost importance. In case of multicentricity, it might be necessary to proceed without, or just with, unilateral surgery for preservation of adjacent structures. In surgery of jugular vein paraganglioma, we usually perform a modified transmastoidal and transcervical approach with preservation of middle-ear structures and the ossicles. As an alternative or supplement to surgery, radiotherapy or definitive embolization may be used in the treatment of paragangliomas. 2. Paragangliomas may occur as multiple endocrine neoplasia (MEN) syndrome combined with medullary thyroid gland carcinoma, and, facultatively, pheochromocytoma. In these cases, endocrinological examination and magnetic resonance imaging (MRI) of the adrenal region, the thorax and the neck are required for an adequate therapeutic strategy. As MEN may be inherited, family history should be evaluated. 3. Paragangliomas can became malignant and metastasize. Thus, cervical lymph node metastases or distant metastases may occur. We recommend the removal of all ipsilateral lymph nodes and their histological examination. PMID- 10696375 TI - Brush cytology and its comparison with histopathological examination in cases of diseases of the nose. AB - Nasal cytology is a simple, easy, bloodless and quite reliable investigation. Although its utility was recognized as early as 1927 by Eyerman, it has not yet gained much popularity. Here, an attempt is made to assess its efficacy for the diagnosis of various inflammatory, benign and malignant conditions in 151 cases of common rhinological problems in the M.Y. Group of Hospitals, Indore MP, India. This method can be used as an adjuvant to clinical diagnosis and histopathological diagnosis as an outpatient department (OPD) procedure with the help of a nasal endoscope, as it was found to be a simple, reliable and time saving procedure for further management. It can also be used as a screening test for the detection of symptomless patients particularly in precancerous lesions. PMID- 10696376 TI - Ultrasound-guided cutting-needle biopsy of the parotid gland. AB - We describe the technique of ultrasound-guided 18 gauge (1.2 mm) needle biopsy in 16 patients with parotid gland lesions. This provides material suitable for histological analysis and can be performed quickly and safely under local anaesthesia. Thirteen of the patients had non-diagnostic blind fine-needle aspiration cytology (FNAC) with a 21 gauge (0.8 mm) needle prior to biopsy. Initial ultrasound was found to be superior to clinical examination in 31 per cent of cases. The ultrasound-guided technique provided a diagnostic specimen in 100 per cent of patients and was helpful where FNAC had been inconclusive. There was a diagnostic accuracy of 100 per cent in the patients who underwent subsequent surgery. This method should be considered when FNAC is non-diagnostic and surgical treatment is being considered. It is particularly useful in patients with diffuse enlargement of the gland and does provide a core of material for accurate assessment of tissue architecture. In this series, nine patients avoided unnecessary surgery. PMID- 10696378 TI - A novel dressing for tracheostomy decannulation. PMID- 10696377 TI - Echography of metastatic nodes treated by radiotherapy. AB - The purpose of this study was to evaluate whether the ultrasonographic appearances of nodal metastases in nasopharyngeal carcinoma (NPC) revert to normal after radiotherapy. Serial ultrasonography was performed in 18 patients with palpable nodal metastases in the neck who underwent radiotherapy for NPC. All patients had a pre-radiotherapy baseline and another study at least one year after radiotherapy. The largest node in each patient was evaluated for any change in the ultrasonographic features following radiotherapy. One year after radiotherapy nodes returned to normal size for their respective areas, the shape of the node and the echogenic hilus also reverted to normal. However, the nodes are more echogenic than nodes in normal subjects. This distinguishes these nodes from nodes not affected by radiotherapy. The histological basis for this observation is presumed to be the result of fibrosis. At first glance the post radiotherapy nodes may resemble normal nodes, however subtle changes within the node and adjacent soft tissues can be recognized on ultrasonography. During long term follow-up, the appearance of nodes at sites previously uninvolved or any change in the appearance of nodes should alert the sonologist to the possibility of recurrence. PMID- 10696379 TI - Gustatory sweating of the external auditory canal. AB - Gustatory sweating of the external auditory canal is extremely rare. A clinical case, that is only the second in the English literature, is presented. The potential pathogenesis and its treatment options are discussed. PMID- 10696380 TI - Erythema multiforme after application of aural Gentisone HC drops. AB - Erythema multiforme is an uncommon acute self-limiting condition characterized by mucocutaneous lesions of varying severity with a variable pattern of recurrence. This is a case of a four-year-old girl who developed erythema multiforme secondary to topical aural application of Gentisone HC drops (hydrocortisone acetate one per cent, gentamicin 0.3 per cent (as sulphate)) prescribed as a treatment for otorrhoea following grommet insertion. The clinical features are described and the literature reviewed. To my knowledge, this is the first reported case in the literature, of erythema multiforme secondary to a topical aminoglycoside/steroid preparation. PMID- 10696381 TI - Carcinoma of the ear: a case report of a possible association with chlorinated disinfectants. AB - In this report we present a case of squamous cell carcinoma developing in a mastoid cavity after prolonged exposure to the chemical disinfectant, Eusol. The efficacy and safety of Eusol and other chloric acid (hypochlorous acid) derivatives in clinical use is debated. PMID- 10696382 TI - Surviving cochlear function in the presence of auditory nerve agenesis. AB - This case reports electrophysiological evidence for cochlear function in a child with radiological evidence of bilateral auditory nerve agenesis or severe hypoplasia. The diagnosis of auditory nerve agenesis was supported by a bilateral atresia of internal auditory canals on computed tomography (CT) scan and magnetic resonance imaging (MRI) absent auditory brainstem responses and absent behavioural responses to sound. Despite the apparent absence of an auditory nerve or spiral ganglion, electrocochleography revealed surviving cochlear function at 70-80 db HL and an abnormal electrocochleographic waveform. This case demonstrates that cochlear function may develop without afferent, or efferent innervation. It also emphasizes that cochlear function may occur in the presence of profound deafness. PMID- 10696383 TI - Delayed deterioration of hearing following bacterial meningitis. AB - Bacterial meningitis is an important cause of acquired sensorineural deafness in childhood. Deafness following meningitis may be progressive. Previous reports have shown deterioration in hearing up to 12 years after the illness. We present two cases of sensorineural deafness following meningitis. Severe to profound sensorineural hearing losses were detected immediately after meningitis in these patients. The hearing subsequently deteriorated in both cases. Deterioration in hearing thresholds occurred 17 years after the illness in one case. In the other patient the hearing got progressively worse three years after meningitis. She subsequently required a cochlear implant. PMID- 10696384 TI - Hearing loss associated with large internal auditory meatus: a report of five paediatric cases. AB - This paper describes the abnormality of a large internal auditory meatus (LIAM). Computed tomography (CT) scans show the otic capsule to be affected by a widened, bulbar internal auditory meatus with loss of or reduction of the bony wall dividing the lateral fundus of the meatus from the cochlea. The vestibule is abnormally dilated. We report five cases of children with LIAM and profound hearing loss. Three of these children are girls and two children were boys. Three had congenital progressive hearing loss, one of these had an accompanying large vestibular aqueduct and dysplasia of the cochlea. Two patients had had meningitis resulting in profound loss. PMID- 10696385 TI - An unusual cause of nasal septum perforation. AB - We present a case where the use of magnetically held earring clips in the nose led to an unfortunate series of events resulting in perforation of the nasal septum. PMID- 10696386 TI - Age-related changes in the epiglottis causing failure of nasal continuous positive airway pressure therapy. AB - At 65 years of age, a former coal miner, now 72-years-old, developed a progressive loss of concentration with daytime sleepiness and sleep disturbances. Work-up in pneumological and medical sleep centres resulted in diagnosis of chronic obstructive pulmonary disease (COPD), borderline obstructive sleep apnoea syndrome and, later, upper airway resistance syndrome. In addition, there was evidence of reduced efficiency of sleep. Neither the initial administration of theophylline nor the later use at night of hyperbaric respiration led to improvement in the patient's symptoms. Instead, the patient developed loud snoring, as well as the inability to sleep while in a lying position. At age 71 years, otorhinolaryngological examination resulted in findings of age-related changes in the epiglottis, that completely blocked the hypopharynx upon inspiration. Polysomnography, which was possible only in a half-seated position, revealed reduction in deep sleep, with a maximum oxygen saturation of 77 per cent at an apnoea-hypopnoea index (AHI) of 4.8. Partial resection of the epiglottis with laser surgery resulted in complete improvement of diurnal drowsiness and reduced stamina. Sleeping in a supine position again became possible. Polysomnography revealed normalization of sleep architecture, but unchanged, low efficiency of sleep. This case underscores the importance of an interdisciplinary approach to the treatment of sleep-related breathing disorders. PMID- 10696387 TI - Severe congenital laryngeal hypoplasia in a 45-year-old man. AB - Congenital laryngeal anomalies are rare. They usually present in infancy and early childhood. We present a case of severe congenital hypoplasia, with multiple anomalies of the laryngeal cartilages in a 45-year-old man. PMID- 10696388 TI - Primary tuberculosis of the posterior oropharyngeal wall. AB - We report a case of primary tuberculosis of the posterior oropharyngeal wall presenting with sore throat, fever and malaise. Pharyngeal tuberculosis is rare and usually occurs with primary pulmonary disease. Primary disease has been reported in small numbers in the nasopharynx and palatine tonsil but never before, to our knowledge, in the posterior oropharyngeal wall. PMID- 10696389 TI - Radiological appearance of primary branchial cleft cyst carcinoma. AB - The hypothesis that primary branchiogenic carcinoma originates from a branchial cleft cyst is controversial. Many reports regarding primary branchiogenic carcinoma failed to provide sufficient evidence to distinguish it from metastatic cervical lymph nodes arising from previously unrecognized primary tumours. The radiological appearance of malignant transformation from a branchial cleft cyst has not been reported previously in the English literature. A radiological study is presented that confirms the primary branchiogenic carcinoma. The management in suspected cases would be wide surgical excision of the tumour including ipsilateral radical neck dissection followed by radiation therapy. PMID- 10696390 TI - Plexiform schwannoma of the cheek. AB - We present a case of subcutaneous plexiform schwannoma, a rare benign peripheral nerve sheath tumour characterized by a multinodular and plexiform growth pattern. A review of the literature was performed to identify the relationship between plexiform schwannoma and neurofibromatosis types 1 and 2, and schwannomatosis. It is also important to distinguish plexiform schwannoma from plexiform neurofibroma, a particular type 1 neurofibromatosis lesion, because of the propensity of the latter for malignant degeneration. PMID- 10696391 TI - Metastatic melanoma of the tonsil. AB - Metastasis to the tonsils from malignant melanoma is rare. This paper describes one such case in a woman with synchronous breast adenocarcinoma and cutaneous malignant melanoma who had a most unusual clinical course. PMID- 10696392 TI - Review of the prevalence of malaria in Zimbabwe with specific reference to parasite drug resistance (1984-96). AB - The response of Plasmodium falciparum malaria to antimalarial drugs, mainly chloroquine, the first-line drug of choice for the treatment of malaria in Zimbabwe is reported here for the period 1984-96. Earlier studies (1982-83) had shown that Zimbabwe was free from drug-resistant falciparum malaria. The first chloroquine-resistant cases of malaria were reported in 1984 in the Zambezi Valley in the north-east of Zimbabwe. Following this report several cases of chloroquine resistance have been reported throughout the malaria-endemic regions of the country thus prompting the Ministry of Health to develop a sustainable national surveillance strategy to monitor, on an annual basis, the spread and extent of P. falciparum resistance to antimalarial drugs available to the National Malaria Control Programme (NMCP). Of all the antimalarial drugs assessed in vivo, only chloroquine and halofantrine have shown resistance, while no resistance in vivo was observed for sulfadoxine-pyrimethamine (Fansidar), quinine and mefloquine. The study shows the need to replace chloroquine with alternative antimalarial drugs in areas where chloroquine resistance is high, and an increase in the drug pool against malaria is also recommended considering that all the alternative antimalarial drugs available to the NMCP have faced resistance in various parts of the world. PMID- 10696393 TI - Royal Society of Tropical Medicine and Hygiene meeting at Manson House, London, 21 January 1999. Malaria control: bednets or spraying? Background and trial in Tanzania. PMID- 10696394 TI - Malaria control: bednets or spraying? Trial in Kwazulu-Natal, South Africa. PMID- 10696395 TI - Malaria control: bednets or spraying? Spray versus treated nets using deltamethrin--a community randomized trial in India. PMID- 10696396 TI - Malaria control: bednets or spraying? Malaria control in the Afghan refugee camps of western Pakistan. PMID- 10696397 TI - Malaria control: bednets or spraying? Summary of the presentations and the discussion. PMID- 10696398 TI - Is the household dog a risk factor for American visceral leishmaniasis in Brazil? PMID- 10696399 TI - Permethrin-treated chaddars and top-sheets: appropriate technology for protection against malaria in Afghanistan and other complex emergencies. AB - Insecticide-treated mosquito nets (ITN) provide excellent protection against malaria; however, they have a number of shortcomings that are particularly evident in politically unstable countries or countries at war: not everyone at risk can necessarily afford a net, nets may be difficult to obtain or import, nets may not be suitable for migrants or refugees sleeping under tents or plastic shelter. There is a need to develop cheaper, locally appropriate alternatives for the most impoverished and for victims of complex emergencies. Afghan women, in common with many Muslim peoples of Asia, wear a veil or wrap known as a chaddar to cover the head and upper body. This cloth doubles as a sheet at night, when they are used by both sexes. A randomized controlled trial was undertaken in which 10% of the families of an Afghan refugee camp (population 3950) in north western Pakistan had their chaddars and top-sheets treated with permethrin insecticide at a dosage of 1 g/m2 while a further 10% had their chaddars treated with placebo formulation. Malaria episodes were recorded by passive case detection at the camp's health centre. From August to November the odds of having a falciparum or vivax malaria episode were reduced by 64% in children aged 0-10 years and by 38% in refugees aged < 20 years in the group using permethrin treated chaddars and top-sheets. Incidence in refugees over 20 years of age was not significantly reduced. The cost of the permethrin treatment per person protected (US$0.17) was similar to that for treating bednets (and cost only 10 20% of the price of a new bednet). An entomological study simulating real-life conditions indicated that host-seeking mosquitoes were up to 70% less successful at feeding on men sleeping under treated chaddars and some were killed by the insecticide. Permethrin-treated top-sheets and blankets should provide appropriate and effective protection from malaria in complex emergencies. In Islamic and non-Islamic countries in Asia, treated chaddars and top-sheets should offer a satisfactory solution for the most vulnerable who cannot afford treated nets. PMID- 10696400 TI - Wide-scale installation of insecticide-treated curtains confers high levels of protection against malaria transmission in a hyperendemic area of Burkina Faso. AB - In a region of Sudanese savannah in Burkina Faso, insecticide-treated curtains were installed in 8 out of 16 zones, each covering an area of about 50 km2. Longitudinal entomological monitoring using CDC light traps was performed in 4 villages (2 intervention, 2 control) over a period of 3 years (including 1 year prior to intervention). In the 3rd year a cross-sectional entomological survey using spray catches was performed in 84 villages (40 intervention). Indoor vector densities in protected houses showed large reductions (P = 0.01). The available data were also consistent with an impact on outdoor and unprotected indoor densities. The proportion of mosquitoes carrying sporozoites was 4.1% in protected villages compared with 11.5% in unprotected villages (P = 0.07). Entomological inoculation rates fell substantially (P = 0.01), reflecting these reductions. The impact of this intervention on mosquito survival appears to have been greater than those in similar trials conducted in the Gambia, Ghana and Kenya in which the intervention was applied over smaller areas. PMID- 10696401 TI - Highland malaria in Uganda: prospective analysis of an epidemic associated with El Nino. AB - Malaria epidemics in African highlands cause serious morbidity and mortality and are being reported more frequently. Weather is likely to play an important role in initiating epidemics but limited analysis of the association between weather conditions and epidemic transmission parameters has been undertaken. We measured entomological variables before and during an epidemic of malaria (which began in February 1998) in a highland region of south-western Uganda and analysed temporal variation in weather data against malaria incidence (estimated from clinic records), mosquito density and entomological inoculation rates (EIR). Indoor resting density of Anopheles gambiae s.l. was positively correlated with malaria incidence (r = 0.68, P < 0.05) despite extremely low vector densities. EIR totalled only 0.41 infectious bites per person during the entire 8-month study period. Rainfall during and following the El Nino event in 1997 was much higher than normal, and rainfall anomaly (difference from the mean) was positively correlated with vector density 1 month later (r = 0.55, P < 0.05). Heavier than normal rainfall associated with El Nino may have initiated the epidemic; the relationship between temperature and transmission parameters remains to be defined. The results from this study indicate that, in this highland population, epidemic malaria may occur at extremely low inoculation rates. PMID- 10696402 TI - Epidemiological surveys confirm an increasing burden of cutaneous leishmaniasis in north-east Brazil. AB - Health service records for north-east Brazil suggest a consistent rise in numbers of cases of cutaneous leishmaniasis due to Leishmania (Viannia) braziliensis over the past decade. In a study site in Pernambuco, prospective, cross-sectional and retrospective epidemiological surveys of infection (a positive Montenegro skin test response) and/or clinical symptoms confirmed a high current force of infection (0.092/year), and an approximately 10-fold increase in transmission during the last 10 years. Cross-sectional analysis indicated that the incidence rate among children (aged < or = 15 years) was lower than that among adult immigrants exposed for similar time periods, but there was no apparent difference in transmission rate according to gender. Coupled with the known behaviour of the local sandfly vector, Lutzomyia whitmani, this suggests that most people are infected outside their houses, rather than either indoors or while visiting remnant rainforest. The estimated proportion of infections which lead to cutaneous lesions (0.81-0.87) is relatively high for L. braziliensis areas. However, an unusually low proportion of clinical infections (0.0042) apparently leads to metastasis. PMID- 10696403 TI - Leishmania (Leishmania) infantum enzymatic variants causing canine leishmaniasis in the Huelva province (south-west Spain). PMID- 10696404 TI - Patterns of concurrent hookworm infection and schistosomiasis in schoolchildren in Tanzania. AB - A cross-sectional study of 6897 schoolchildren in 59 out of the 155 primary schools in Magu District on the shores of Lake Victoria, Tanzania, was undertaken in 1997 to determine the prevalence of single- and multiple-species helminth infection. Schistosoma haematobium, hookworm (primarily Necator americanus) and S. mansoni were the most common helminth species infecting schoolchildren in the district. The prevalences of Ascaris lumbricoides and Trichuris trichiura were negligible (< 1%). Anaemia and stunting were highly prevalent and widespread. Hookworm and S. mansoni occurred more frequently in multiple infections with other helminths than as single-species infections, but triple-species infection was rare. Analysis of the frequency distribution of infection amongst schools showed that prevalences of S. haematobium and hookworm tended to be normally distributed, with medians 75% and 45%, respectively, while the distribution of S. mansoni was markedly skewed such that only 17% schools had a prevalence greater than 20%. An inverse association between S. mansoni and S. haematobium was observed. Geographical information system (GIS) analysis indicated that S. mansoni infection was highly prevalent only along the shore of Lake Victoria, whilst S. haematobium was homogeneously prevalent everywhere except the lakeshore. This pattern appears to reflect the distribution of schistosome species-specific snail intermediate hosts. The results imply that joint treatment for hookworm infection and schistosomiasis would be beneficial throughout the district. PMID- 10696405 TI - The distribution of Ascaris lumbricoides in human hosts: a study of 1765 people in Bangladesh. AB - The Ascaris lumbricoides expelled by 1765 people in a poor urban community in Bangladesh were recovered and counted after the subjects had been treated with pyrantel pamoate. The subjects were divided into 22 classes by age and sex (mean n = 80) to examine how prevalence, mean worm burdens and measures of aggregation of worms varied with age and between the sexes, and to see how a measure of aggregation, k, calculated in 3 ways (by maximum likelihood, from moments, or from the percentage uninfected) compared with an empirical aggregation index (the percentage of subjects who expelled an arbitrary 80% of all worms) and with the proportion who were moderately to heavily infected (defined as > or = 15 worms). The prevalence of infection ranged from 64% to 95%, mean worm burdens ranged from 7 to 23 worms, and k ranged from 0.3 to 1.2. There were significant differences between adult males and females in the prevalence of infection, mean worm burdens and measures of aggregation, differences which are probably driven more by behaviour than immunity. The parameter k was better described in terms of the proportion who were moderately to heavily infected (linear; range 0.15-0.58) than by the empirical aggregation index (non-linear; range 0.30-0.49). PMID- 10696406 TI - Antibodies to Japanese encephalitis virus in human sera collected from Irian Jaya. Follow-up of a previously reported case of Japanese encephalitis in that region. AB - Japanese encephalitis virus (JEV) and other arboviruses are demonstrating an emergence in the southern part of New Guinea Island. JE was previously unknown in this part of the world until 1995 when it was found in the Torres Strait, northern Australia. In this study 96 sera collected from residents of the Timika region of Irian Jaya were tested for antibodies to JEV and related arboviruses by epitope-specific blocking ELISA. Of the 9 sera deemed to be positive for JEV antibodies by ELISA, 5 were collected from persons indigenous to Timika, and who had not travelled to regions where JE is known to be active. This indicates that these individuals were infected with JEV in the Timika area and supports a recent report of a clinical case of JE in this region. Non-immune expatriates visiting or working in the Lowland areas of Irian Jaya and/or Papua New Guinea should consider immunization against JE. Precautions should always be taken to avoid being bitten by any mosquito both in the daytime and at night. PMID- 10696407 TI - Geophagy is common among Luo women in western Kenya. PMID- 10696408 TI - Field evaluation of the OptiMAL rapid malaria diagnostic test during anti malarial therapy in Guyana. PMID- 10696409 TI - An assessment of the accuracy of clinical diagnosis, local microscopy and a rapid immunochromatographic card test in comparison with expert microscopy in the diagnosis of malaria in rural Kenya. PMID- 10696410 TI - Field trial of the direct acridine orange method and ParaSight-F test for the rapid diagnosis of malaria at district hospitals in Dar es Salaam, Tanzania. PMID- 10696411 TI - Application of an enzyme-linked immunosorbent assay to detect antibodies to Onchocerca volvulus on filter-paper blood spots: effect of storage and temperature on antibody decay. PMID- 10696412 TI - Detection of Japanese encephalitis virus antigen in desiccated mosquitoes: an improved surveillance system. PMID- 10696413 TI - Application of PCR for detection of toxigenic Vibrio cholerae O1 in water samples during an outbreak of cholera. PMID- 10696414 TI - Cognitive sequelae of severe malaria with impaired consciousness. AB - Although cerebral malaria is the most common acute encephalopathy arising in children in Africa little is known of its effect upon the longer-term cognitive development of survivors. In Kenya, we compared the performance of 87 survivors of severe malaria with impaired consciousness to matched community controls on a wide range of tasks, not less than 42 months post illness episode. The presence of cognitive impairment was then related to both the pattern of symptoms at the time of the acute illness and the presence of gross neurological impairment on discharge. Significant group differences were found in areas of cognitive functioning suggestive of widespread impairment in the development of the ability to initiate, plan and carry out tasks (the executive functions). On tasks of more discrete cognitive skills (information processing) there were no significant group differences, although impaired performance was found more frequently in the severe malaria group. The odds ratio associated with the development of cognitive impairment following severe malaria with impaired consciousness was found to be 4.48 (95% CI 1.22, 16.47). A combination of 4 signs (coma, hypoglycaemia, seizures, and absence of hyperpyrexia) proved to have greater accuracy than the presence of gross neurological sequelae in predicting cognitive impairment (95% vs 93% specificity, 67% vs 58% sensitivity). PMID- 10696415 TI - Ocular myositis and diffuse meningoencephalitis from Trypanosoma cruzi in an AIDS patient. PMID- 10696416 TI - Helicobacter pylori is not a determinant factor of persistent diarrhoea or malnutrition in Peruvian children. AB - To investigate the role of Helicobacter pylori in childhood diarrhoea, specific IgG antibodies to H. pylori (determined by an ELISA) were sought in 119 infants aged 3-36 months in Peru. Thirty one of the infants had acute diarrhoea (defined as lasting < 72 h and not present in the previous 3 weeks), 67 had persistent diarrhoea (lasting > or = 14 days with no more than 1 intervening diarrhoea-free day) and the remaining 21 had not had diarrhoea in the previous 3 weeks. The children with diarrhoea had been admitted to hospital in Lima for diarrhoea treatment, and the diarrhoea-free children for investigation of possible tuberculosis. Aspirates of duodenal contents and duplicate stool samples were investigated for the presence of bacterial overgrowth and of pathogenic bacteria, viruses and parasites. Anthropometric measurements were also made. There were no statistically significant differences between the prevalence rates of IgG against H. pylori in the children with acute diarrhoea, persistent diarrhoea and without diarrhoea (32%, 43% and 29%, respectively). In addition, H. pylori infection (as evidenced by specific antibodies) had no apparent influence on the presence of small-bowel overgrowth (in 20% of seropositive children compared with 18% of seronegative children) or of pathogens in the stool (in 53% of seropositive children compared with 49% of seronegative children) or on the occurrence of malnutrition in the groups of children considered as a whole. We conclude that H. pylori infection is not associated with acute or persistent diarrhoeal disease, small-bowel overgrowth, stool pathogens or malnutrition in Peruvian children. PMID- 10696417 TI - Histoplasmosis in eastern India: the tip of the iceberg? AB - Systemic histoplasmosis has various clinical presentations and is of especially concern in immunocompromised patients. A high index of suspicion is required for its diagnosis. A total of 38 cases had been reported from India up to 1996. The most frequent occurrence of cases was around Calcutta in eastern India where the previous case was detected 20 years earlier. However, we have diagnosed 5 cases in the past 2 years from eastern India which are reported here. These cases may indicate under-diagnosis and under-reporting of histoplasmosis in India. All 5 patients had disseminated disease with multisystem involvement including 2 with bilateral adrenal enlargement. Two were diabetic and only 1 patient was infected with HIV. PMID- 10696418 TI - A randomized safety and tolerability trial of artesunate plus sulfadoxine- pyrimethamine versus sulfadoxine-pyrimethamine alone for the treatment of uncomplicated malaria in Gambian children. AB - Artemisinin derivatives, such as artesunate, have a short half-life and very rapid anti-malarial activity. Theoretically, using such agents in conjunction with well-established anti-malarial drugs such as sulfadoxine-pyrimethamine may reduce the rate of drug resistance. Such a combination has not previously been used in Africa. We have conducted a pilot safety trial of artesunate (4 mg/kg for 3 days) given with a single dose of sulfadoxine-pyrimethamine (25 mg/kg sulfadoxine) compared to sulfadoxine-pyrimethamine alone among 40 Gambian children with uncomplicated malaria. Both regimens were safe and well tolerated and there were no adverse experiences attributed to the combination. The addition of artesunate resulted in a higher proportion of afebrile children and children with a negative blood film on Day 2, and a reduction in the proportion of gametocyte carriers, when compared to sulfadoxine-pyrimethamine alone. PMID- 10696419 TI - Low-dose quinine for treatment of Plasmodium falciparum malaria in Guinea-Bissau. AB - The recommended dose of 10 mg quinine/kg bodyweight 3 times a day for 7 days for treatment of malaria is so high that many patients experience cinchonism. We have earlier obtained good results with 7 days' treatment with 20 mg Quinimax/kg bodyweight divided into 2 daily doses. In order to identify the lowest effective dose, children with symptomatic malaria were treated with quinine twice a day for 7 days. They were assigned to 1 of 3 groups treated daily with 10 mg/kg, 15 mg/kg, or 20 mg/kg bodyweight, respectively; 42, 46, and 34 children, respectively, received treatment and completed 5 weeks of follow-up. The cumulative percentages of all children with parasitaemia during follow-up on day 28 or before were 33%, 13% and 12%, respectively. Treatment with 10 mg quinine salt/kg daily for 7 days gave a significantly higher rate of recrudescence than did treatment with 15 or 20 mg/kg daily. Thus at least 15 mg of quinine salt/kg bodyweight daily should be recommended for treatment of symptomatic Plasmodium falciparum malaria in Guinea-Bissau. PMID- 10696420 TI - Heterogeneous activity in vitro of vitamin A (retinol) in combination with novel and established antimalarial drugs. PMID- 10696421 TI - Randomized controlled trial of directly observed treatment (DOT) for patients with pulmonary tuberculosis in Thailand. AB - While directly observed treatment (DOT) has been recommended as the standard approach to tuberculosis control, empirical data on its feasibility and efficiency are still scarce. We conducted a controlled trial of DOT at 15 health care facilities at various levels of the government health care system in Thailand. A total of 836 patients diagnosed between August 1996 and October 1997 were randomly assigned to be treated either under DOT or self-supervised using monthly drug supplies (SS). Options for treatment supervisors were health staff, community members or members of the patients' families. Treatment outcomes were compared on the basis of cure, treatment-completion, default and death rates. In both study arms, treatment outcomes were improved compared to pre-study conditions. Cure and treatment-completion rates were significantly higher in the DOT cohort (76% and 84%) than in the SS group (67% and 76%). The benefits of DOT were more pronounced at district and provincial hospitals (DOT cure rate 81% vs. 69% in the SS group), while differences for patients treated at referral centres were non-significant (DOT cure rate 72% vs. 66% in the SS group). No significant differences in outcomes could be observed between patient groups receiving DOT under the various options for treatment supervisors. DOT appears especially suited for treatment at decentralized facilities. While a general focus on programme performance can improve outcomes, DOT provides significant additional benefits. If basic conditions are met, a DOT strategy can be tailored to country specific conditions by exploring multiple observation options, without decreasing its effectiveness. PMID- 10696422 TI - Estimation of the local synthesis of immunoglobulin G (IgG) in the central nervous system of patients with spinal cord schistosomiasis by the IgG index. AB - By analogy with other infections of the central nervous system (CNS), it is believed that schistosomal myeloradiculopathy (SMR) is an entity that may involve a mild-to-moderate degree of impairment of the blood-brain barrier along with intrathecal synthesis of antibodies. The first of these aspects is obvious but the second has not been clearly demonstrated. This study was undertaken in Brazil with the aim of investigating the production of immunoglobulin G (IgG) within the CNS in patients with SMR, by the determination of the cerebrospinal fluid (CSF) IgG index. The study population included 54 patients with SMR, evaluated prospectively. The CSF IgG index was increased in 43 of them (80%). Preliminary results from our laboratory suggest that these antibodies are reactive against Schistosoma mansoni antigens. Thus, this finding also suggests that this index may be useful in the differential diagnosis of SMR. PMID- 10696423 TI - Phlebotomus (Paraphlebotomus) marismortui Theodor, 1947, synonym of P. (Pa.) alexandri Sinton, 1928. PMID- 10696424 TI - HSV gene functions: what have we learned that could be generally applicable to its near and distant cousins? AB - Herpes simplex virus 1 (HSV-1) encodes at least 84 polypeptides to perform two functions: to enable viral replication and to create the environment in which the entry of the virus into host cells, synthesis of virion components, assembly and egress are optimized. Whereas the former are indispensable for viral replication, the latter, numbering 47, can be deleted without a major effect on viral replication in cells in culture. Of particular interest are gene products whose function is either to modify cellular proteins (set 1) or to block entirely their function (set 2). An example of set 1 is the infected cell protein No. 0 (ICP0), a promiscuous transactivator of genes introduced into cells by infection or transfection. In its nuclear phase this protein binds to cyclin D3, extends its life by many hours, and sequesters it in nuclear structures known as ND10. In its cytoplasmic phase, ICP0 binds the translation elongation factor EF-1 delta. Another viral protein, the UL13 protein kinase, hyper-phosphorylates EF-1 delta. ICP0 and the protein kinase stimulate protein synthesis and cause the cell to induce the synthesis of pre-S phase cellular proteins the virus needs for its replication. The gamma 134.5 protein, a prototype of set 2, also has multiple functions. One, mapped at its carboxyl terminus, blocks the effects of double stranded RNA-dependent protein kinase R (PKR) that is activated by all wild-type and mutant viruses examined to date. PKR phosphorylates eIF-2 alpha and shuts off protein synthesis. gamma 134.5 protein binds protein phosphatase 1 and redirects it to dephosphorylate eIF-2 alpha. Although PKR is activated in wild-type infected cells, protein synthesis is unaffected. HSV-1 encodes in addition at least two proteins, ORF O and ORF P that are repressed during productive infection. The ORF P protein localizes in spliceosomes and blocks the synthesis of viral proteins derived from spliced mRNA. The ORF O protein binds ICP4, the major regulatory protein, and prevents it from binding to DNA. The role of ORF O and ORF P proteins in the establishment of latency is uncertain. A significant discovery that has emerged from these studies is that viral proteins can perform several functions that may be totally unrelated. PMID- 10696425 TI - The dynamic herpesvirus DNA genome: the case of MDV-1 and HSV-1. AB - Herpesviruses evolved from an ancestral viral genome that contained five blocks of genes which provide the members of this family of viruses with structural and enzymatic properties. These genes allow the herpesviruses to infect a host by entering into the nuclei of the cells, the site of replication and transcription of the viral DNA. The viral mRNAs are released into the cell cytoplasm where synthesis of enzymatic and structural proteins occurs. The latter proteins are responsible for the formation of the infectious virions. Herpesviruses that were able to adapt to different hosts during the evolution of the species (speciation) had acquired additional genes from transposons or retrotransposons that allowed them to successfully maintain their hold in the specific vertebrate host. The present overview deals with molecular differences between Marek's disease virus type 1 (MDV-1) and herpes simplex virus type 1 (HSV-1) and the specialized genes that differentiate MDV-1 from HSV-1, the promoters of the viral genes that control gene expression and the nuclear localization signals. Dynamic changes in the viral genomes that may occur during viral DNA replication and recombination and their effects on virus pathogenicity and genome evolution will be discussed. PMID- 10696426 TI - Molecular characteristics of very virulent European MDV isolates. AB - Marek's disease virus (MDV) strains with increasing virulence have been reported from many parts of the world. Many of these recent MDV isolates produce an acute early cytolytic disease with high mortality and severe atrophy of the lymphoid organs, thymus and the bursa of Fabricius. Although the degree of the atrophic changes and the virulence of the virus are correlated, the molecular basis of the increased virulence is not known. We examined the characteristics of the disease induced by 3 such MDV isolates, C12/130, MR36 and MR48, isolated from Europe. All the three viruses produce high early mortality and atrophy of the lymphoid organs. As a first step in understanding the determinants of the increased virulence of these isolates, we have compared the sequences of MEQ and the ICP4 genes of these three viruses with that of the published sequences. Some of the amino acid changes seen within the Meq and ICP4 proteins were conserved in all the three isolates and could account for the increased virulence characteristics. PMID- 10696427 TI - MEQ and V-IL8: cellular genes in disguise? AB - One of the hallmarks of oncogenic viruses is their ability to subvert the growth regulation and evade immune response of the host. There are a number of tricks devised by various virus families. Oncogenic herpesviruses often accomplish this by encoding homologs of cellular genes involved in these functions. These viral homologs sometimes are hyperactive forms of their cellular counterparts, which function to overtake the cellular pathways, other times serve as decoys to mask the cellular functions. Marek's disease virus (MDV) carries at least two genes in that category. We have previously described Meq protein (MEQ gene product), a transcriptional factor with homology to proto-oncogenes Jun and Fos in the bZIP domain. Meq dimerizes with Jun or Fos and the Meq/Jun heterodimer is able to transactivate promoters with AP-1 site. We show here that Meq and Jun colocalize in living cells, adding to the physiological significance of the dimer formation. In addition, we present data to show that Meq and Jun can functionally complement each other in cis and in trans, using transformation and transactivation assays. Finally we describe the discovery of an IL8 chemokine homolog, designated as v IL8 (viral IL8) in the MDV genome and discuss its possible function in MDV infection. PMID- 10696428 TI - Comparison of two serotype 1 MDV isolates. AB - We compared the RB1B and T. King (TK) serotype 1 isolates of Marek's disease virus (MDV) in vivo. Body and organ weights, mortality, and lesions indicated that the TK inoculum established early infection more efficiently than RB1B and did greater damage to the bursa of Fabricius and thymus. Subsequent studies showed that the TK inoculum that we used contained chicken infectious anemia virus (CIAV). Therefore, pathogenicity profiles shown here should be interpreted with the presence of CIAV contamination in the TK stock in mind. PMID- 10696429 TI - Establishment of a lymphoblastoid cell line using a mutant MDV containing a green fluorescent protein expression cassette. AB - We have previously described the construction and characterization of mutant Marek's disease viruses (MDVs) having mutations within the unique-short (US) region of the genome that have retained oncogenicity (Anderson et al., 1998; Parcells et al., 1995). We have also reported the characterization of lymphoblastoid cell lines (LBCLs) derived using these mutant viruses (Parcells et al., 1998). These mutant MDVs were constructed using a lacZ expression cassette. Expression of lacZ was found to be constitutive during lytic infection but was found to be tightly repressed in tumors and the derived LBCLs. The construction of these viruses and the analysis of lacZ induction required the use of toxic substrates or antibody staining to detect lacZ expression. We now report the establishment of an MDV lymphoblastoid cell line, MDCC-UA04, that was derived from a tumor induced by an MDV having an insertion of a green fluorescent protein expression cassette into the US2 gene. Like previous mutant-derived LBCLs, expression of the marker cassette is constitutive in lytic infection, but repressed in tumors and in the UA04 cells. UA04 cells express CD3low, CD4, TCR 2low, MHC class II, and CD28 antigens on their surface. The percentage of UA04 cells expressing GFP is generally low (5-7%), but increases markedly within 48 hrs of 5'-iododeoxyuridine (IUdR) treatment (20-30%) in a manner similar to many MDV lytic antigens. Thus, induction of GFP expression in UA04 cells can serve as a non-invasive marker for MDV reactivation from latency. PMID- 10696430 TI - A monoclonal antibody to ICP4 of MDV recognizing ICP4 of serotype 1 and 3 MDV strains. AB - Monoclonal antibodies (MAbs) were prepared against ICP4 of Marek's disease virus (MDV). Mice were inoculated with ICP4 obtained from High-Five insect cells infected with a recombinant baculovirus expressing ICP4. MAbs were selected by enzyme-linked immunosorbent assay (ELISA) using MDV-infected and control chick kidney cells as antigens. One of the MAbs, 5H8, recognized an epitope toward the carboxyl terminus of ICP4 based on staining of reticuloendotheliosis virus transformed cells transfected with full-length and truncated ICP4 constructs. This MAb recognized ICP4 in chicken embryo fibroblasts (CEFs) infected with MDV strains JM16 and HVT but not with SB-1 strain. Using Western blot analysis a protein of 155 kDa was detected in CEFs infected with JM16 and HVT strains. PMID- 10696431 TI - Effect of native chicken interferon on MDV replication. AB - Marek's disease virus (MDV) is an oncogenic alphaherpesvirus. Its specific phosphorylated protein, pp38 has been implicated in MDV oncogenesis. In order to check whether the known anti-viral or anti-proliferative actions of interferon (IFN) are of importance in Marek's disease (MD), chicken embryo fibroblasts (CEFs) were infected with attenuated serotype-1 MDV strain CVI988, or with herpesvirus of turkeys (HVT). Different concentrations of native chicken IFN were added to the cell cultures, prior to their infection. After incubation, MDV plaques were counted. Analysis by flow cytometry for pp38 expression was performed by using three monoclonal antibodies (MAbs) and for HVT by using an anti-glycoprotein B (gB) MAb. Increasing IFN quantities caused a reduction in a stepwise manner of plaque numbers as well as a suppression of pp38 and gB expression in the CVI988- and HVT-infected cells, respectively. PMID- 10696432 TI - Expression of bcl-2 and bcl-x genes in lymphocytes and tumor cell lines derived from MDV-infected chickens. AB - To characterize the molecular events involved in both apoptosis and transformation process induced by Marek's disease virus (MDV), the expressions of the bcl-2 and bcl-x genes, ones of the dominant apoptosis-regulating genes, in Marek's disease (MD) tumor cell lines and cells prepared from MDV-infected chickens were analyzed. The expression of bcl-2 was down-regulated in both CD4+ and CD8+ T cells prepared from MDV-infected chickens at 3 weeks p.i. No bcl-2 transcript was detected in MD tumor-derived MSB1 and MTB1 cell lines, which had been established from primary MD tumors. On the other hand, the bcl-xL transcript whose product can also inhibit apoptosis was expressed in cell lines derived from MD. By the treatment with phorbol 12-myristate 13-acetate (PMA) and ionomycin, normal CD4+ T cells were induced to express bcl-xS which can promote apoptosis, while bcl-xL was constitutively expressed in MD cell lines. Our results suggest that bcl-xL rather than bcl-2 might play an important role in the transformation process by MDV. PMID- 10696433 TI - Inhibition of apoptosis by Marek's disease viruses. AB - Strains of Marek's disease virus (MDV), a herpesvirus, have been shown to augment the development of lymphoid leukosis induced by retroviruses, the avian leukosis virus (ALV) or the reticuloendotheliosis virus. In this study we explored the possibility that the ability to augment lymphoid leukosis may be correlated with the ability of different strains of MDV to block apoptosis. Subclones of the ALV transformed B cell line, DT40, which was free of MDV DNA were infected with either R2/23 strain of MDV-1, SB1 strain of MDV-2, or turkey herpes virus (HVT), a MDV-3. We found that most of the normal DT40 cells and DT40 cells infected with the R2/23 became apoptotic when cultured in serum-reduced medium. By contrast, the frequency of apoptotic cells was greatly reduced in the DT40-SB1 and DT40-HVT subclones. These findings suggest that because the SB-1 strain persists in the ALV-infected cells, it may augment lymphoid leukosis by inhibiting apoptosis and providing a survival advantage to the B cells which have a deregulated myc proto oncogene. PMID- 10696434 TI - Multiple infection of chickens and turkeys with avian oncogenic viruses: prevalence and molecular analysis. AB - The avian herpesvirus, Marek's disease virus (MDV) and several retroviruses, reticuloendotheliosis virus (REV), avian leukosis virus (ALV) (chickens) and lymphoproliferative disease virus (turkeys) are oncogenic and immunosuppressive agents. These viruses were detected either alone, or in various combinations in blood and tumor DNAs of commercial birds using PCR. We present a 5-year retrospective study that included 207 chicken and 52 turkey flocks. Of these, 32 chicken and 18 turkey flocks were negative. Of the positive chicken and turkey flocks 76% and 75%, respectively, had a single, while the rest, 24% and 25%, had a multiple virus infection. In the chickens of the multiple virus-infected flocks, 14% and 17% of the blood and tumor DNAs carry dual MDV and REV and/or ALV sequences, that is about 30% of the PCR-positive, and about 5% of the total DNAs analysed. Multiple virus sequences were detected only in the turkey blood DNAs 11% of 84 samples. Following that quantitation we aimed to analyse the molecular status of the retrovirus sequences in order to determine whether retrovirus sequences were integrated into the herpesvirus genome. We focused on the MDV BamH1-H 132 bp tandem repeat fragment proximity using a combined PCR (cPCR) to identify chimeric PCR products. That included amplification with heterologous combinations of the MDV and retroviral LTR primers. In 13 of 35 DNAs that had both MDV and retrovirus sequences new products were produced. Of 4 MDV + REV chimeric products that were sequenced, one was homologous to the Chicken Repeat element 1 non-LTR type retrotransposon. No evidence for a retrovirus LTR integration was found in the 132 bp repeat proximity, but in two of these products we detected nucleotide stretches of 20 bp and 21 bp with a 70% and 71% homology to the REV-LTR. Also, the amplification of the chimeric products using a retrovirus primer denoted that at least short nucleotide stretches homologous to retroviral LTR primer were present in these DNAs, and that they might resemble ancient retroviral insertions, as previously demonstrated (Isfort et al., 1992). PMID- 10696435 TI - Evidence for Marek's disease in turkeys in Germany: detection of MDV-1 using the polymerase chain reaction. AB - Since 1994 tumorous lesions have been monitored in turkeys on three farms in Germany. On one of these farms, chickens also had tumorous lesions. Affected turkeys were retarded in growth, apathic, pale and almost unable to move. The older the animals got, the more indistinct the clinical signs became. Mortality started at an age of 5 weeks and reached between 20% and 60% by the end of the fattening period of about 20 weeks. The aetiological differential diagnosis includes reticuloendotheliosis (RE), lymphoproliferative disease (LPD), lymphoid leukosis (LL) and Marek's disease (MD). Repeated serological examinations did not establish the presence of antibodies against REV, LLV or MDV-1. Cloacal swabs were negative for LL P27 antigen in ELISA. Solid tumors of various sizes as well as diffuse infiltrations were predominantly seen in the liver, spleen and kidney. Pleomorphic cell infiltration was rarely noted in the plexus brachialis and nervus ischiaticus. Herpesvirus of turkeys (HVT) was the only virus isolated from buffy coat cells derived from affected turkeys in chicken embryo kidney cell (CEK) and chicken embryo fibroblast (CEF) cultures. Use of polymerase chain reaction (PCR) for the amplification of the 132 bp repeat region provided evidence for the presence of MDV-1 DNA in tumor tissue from several diseased turkeys. No evidence was found for the presence of REV. PMID- 10696436 TI - Tissue-specific restriction of latent turkey HVT transcription. AB - Using in situ hybridization, latent turkey herpesvirus (HVT) transcription was examined in lymphoid and/or nonlymphoid tissues. Blood samples were taken for virus isolation from chickens at 7 and 240 days post infection (PI) representing time points for productive and latent turkey herpesvirus infections, respectively. Spleen, thymus, sciatic and brachial nerves from infected chickens were analyzed for latent HVT transcription and HVT glycoprotein B (gB) expression at 240 days PI. Using indirect immunofluorescence, HVT gB expression was not detected in any tissues examined at 240 days PI. HVT genomic fragments from a HVT BamHI library were used as probes in in situ hybridization assays. In the spleen, thymus, sciatic and brachial nerves, latent HVT transcription occurred from the repeat regions flanking the unique long region (TRL and IRL). However, fine mapping of this region revealed a difference in latent HVT transcriptional pattern. A SmaI map of the HVT BamHI-F fragment was made to further fine-map latent HVT transcription. A 1.6 kbp SmaI subfragment hybridized to cells infected with latent HVT in the spleen and thymus. However, the 1.6 kbp SmaI subfragment did not hybridize to cells of the brachial or sciatic nerves. In addition, a 2.0 kbp SmaI subfragment hybridized to cells in the thymus but not in the spleen, sciatic or brachial nerves. The above results suggest that latent turkey herpesvirus exhibits tissue-specific transcription. PMID- 10696437 TI - Identification and characterization of glycoprotein H of MDV-1 GA strain. AB - A 2439 bp open reading frame (ORF) was identified from the DNA sequence of BamHI F and -K2 fragments of Marek's disease virus of serotype 1 (MDV-1) GA strain, which predicts an 813 amino acid polypeptide. This peptide is homologous to HSV-1 gH, and has typical glycoprotein features. There are nine potential N-linked glycosylation sites within the extracellular domain. A fragment of the gH ORF was cloned into pGEX vector in frame with glutathione S-transferase (GST) to produce a GST-gH fusion protein in Escherichia coli. The GST-gH fusion protein was used to develop gH monoclonal and polyclonal antibodies. Expression of gH was detected in duck embryo fibroblasts (DEFs) infected with MDV-1 GA strain by immunofluorescence assay (IFA) with these antibodies. Virus neutralization and plaque-forming inhibition analyses were conducted with the gH antiserum. There were no neutralization and plaque-forming inhibition activities of gH antiserum. Comparison of the DNA sequence of gH gene between GA and RB1B strains of MDV-1 revealed major difference in the upstream control elements of gH ORF. PMID- 10696438 TI - MDV glycoprotein D is expressed in the feather follicle epithelium of infected chickens. AB - Glycoprotein D (gD) and its homologues are essential for many alphaherpesvirus life cycles. A gene encoding a homologue of gD was recently found in the Marek's disease virus (MDV) genome. Interestingly, gD-negative MDV mutants apparently replicate unimpaired in both cell culture and chickens. In this study, we show the expression of the gD homologue of MDV in feather follicle epithelium in infected birds. The gD homologue was detected in a few feather follicles even when most of the follicles were expressing pp38 or gB, other MDV-specific proteins. These observations indicate that the MDV gD homologue is expressed in a very limited set of cells and may be differently regulated. Since feather follicle epithelium of infected birds are the only place where the infectious cell-free MDV virions can be recovered, analysis of the transcriptional regulation of gD may lead to the understanding of the cell-associated nature of MDV. PMID- 10696439 TI - Glycoprotein gD of MDV lacks functions typical for alpha-herpesvirus gD homologues. AB - Glycoprotein D (gD) belongs to family of conserved structural proteins of alpha herpesviruses. During productive infection of cells by herpes simplex virus 1 (HSV-1) gD has several important functions, is involved in virus penetration to and release from infected cells and is one of main targets of neutralizing antibodies. Similar functions are shared also by other alpha-herpesvirus gD homologues. Surprisingly, in previous studies it was found that MDV gD expression could not be detected during infection in vitro using immunological methods. In this study we have analyzed expression of MDV gD and its biological consequences. In vitro expression using rabbit reticulocyte lysate and/or overexpression in transfected cells showed that the second ATG codon is required for synthesis of mature, glycosylated gD. In addition, it was found that gD overexpression is neither toxic for transfected cells nor is involved in membrane fusion. After MDV infection of a proprietary cell line stably transfected with plasmid overexpressing MDV gD, no viral particles could be found in culture. On the other hand, cells overexpressing the MDV gD were sensitive to MDV infection in similar way as parental, non-transfected cells. From our study and results of other authors we propound the following conclusions: (i) MDV gD expression is blocked during in vitro infection at transcription level; (ii) MDV gD is lacking many important functions characteristic for other alpha-herpesvirus gD homologues; (iii) overexpression of single MDV gD does not result in production of mature infectious MDV particles. PMID- 10696440 TI - Construction and characterization of a H19 epitope point mutant of MDV CVI988/Rispens strain. AB - A recombinant virus, CVI/rpp38, was developed from the Marek's disease virus (MDV) CVI988/Rispens vaccine strain. This recombinant was obtained by transfection of CVI988/Rispens-infected chick embryo fibroblasts (CEFs) with plasmid pHA25 DNA containing pp38 gene from GA strain of MDV. Monoclonal antibody (MAb) H19 which reacts with pp38 from GA but not with that from CVI988 was used to screen for recombinant viruses in transfected cell culture plates by immunofluorescent assay (IFA). A positive plaque was isolated, propagated, and purified from cell-free virus particles after sonication of infected CEFs. The mutant CVI/rpp38 was not only reactive with MAb H19 in IFA but also in immunoprecipitation. A 38 kDa protein was immunoprecipitated from the CVI/rpp38 mutant virus but not from parental CVI988 virus. DNA sequence of the mutant virus showed a substitution of G at position 320 by a resulting in a change of an amino acid residue from arginine to glutamine. Comparison of nucleotide sequence of pp38 from strains GA, Md5 and Md11/75c/R2 and CVI988 revealed change to glutamine in this position. The result of this study provides a direct evidence for the location of the identified H19 epitope in pp38. This mutant is potentially useful to further explore the biological function of pp38 and its H19 epitope. PMID- 10696441 TI - Identification and structural analysis of a MDV gene encoding a protein kinase. AB - DNA sequence analysis of the BamHI-C fragment of Marek's Disease Virus (MDV) reveals the presence of a 513 amino acid open reading frame (ORF). This ORF codes for a protein with an estimated M(r) of 58,901. Comparison of the amino acid sequence with those available in the Swiss-Prot database indicates extensive homology with a protein kinase (PK) of herpes simplex virus (HSV) and varicella zoster virus (VZV). In Northern blot hybridization, a transcript of 2.0 kb was detected in MDV (GA strain) infected duck embryo fibroblasts (DEFs). A portion of the ORF was expressed in Escherichia coli as a trpE-fusion protein and used to generate antiserum in New Zealand rabbits. This antiserum specifically detected a protein of 60 kDa in MDV serotype 1, 2 and 3 infected DEFs or chicken embryo fibroblasts (CEFs) by Western blot analysis. This ORF codes for a functional PK. PMID- 10696442 TI - Non-essential loci in the BamHI-I and -F fragments of the HVT FC126 genome. AB - The sequence of BamHI-I fragment of the herpesvirus of turkeys (HVT) FC126 strain DNA was analyzed for the presence of potential open reading frames (ORFs). Four complete (ORFs 2 to 5) and 2 partial ORFs (ORFs 1 and 6) were detected. ORFs 2 and 3 were homologous to the HSV-1 UL55 and the EHV-1 gene 3, respectively. The ORF 6 was already partially sequenced by Smith et al. (Virology 207, 205-216, 1995), and was homologous to a Marek's disease virus (MDV) ORF located in a similar position (ORF 21; Ross et al., Virus Genes 7, 33-51, 1993a). No significant homology was found for the other ORFs. ORF 4 was antisense to ORF 3. Two HVT recombinants having an expression cassette inserted into two intergenic sites were generated and tested for viremia in chickens. Results demonstrated that these 2 intergenic loci are non-essential for in vitro and in vivo HVT replication. A 650 bp deletion in the repeat region flanking UL (TRL and IRL (BamHI-F)) has been identified in some DNA molecules of HVT FC126 strain. This deletion covers the entire truncated pp38 homologous ORF and the N-terminus of a small ORF which has no detectable homology with any known gene. Our results indicate that (1) this genomic region including the HVT pp38 homologue was not essential for in vitro and in vivo growth of HVT, and (2) this deletion had no apparent effect on Marek's disease (MD) protection induced by HVT. PMID- 10696443 TI - Development of a cell line system susceptible to infection with vaccine strains of MDV. AB - Despite reliance on the need to continually prepare fresh cultures of chick embryo fibroblasts (CEFs) to make Marek's disease (MD) vaccines, MD vaccines are the most widely used vaccines in the poultry industry. Preparation of CEF's accounts for approximately 40% of the costs associated with producing MD vaccines. A significant reduction in MD vaccine production costs could be realized if a continuous cell lines were available for MD vaccine production. Recently, we reported development and characterization of a cell line system (OCL) that supports growth and replication of oncogenic serotype 1 Marek's disease virus (MDV). Here we report development of three cell line systems for production of MD vaccine. These cell lines support the growth and replication of attenuated serotype 1 MDV (CVI-OCL), serotype 2 MDV (SB1-OCL) and serotype 3 MDV (HVT-OCL). MDV is maintained in a stable state in the OCL cells and the infected cells can be continuously grown. The vaccines made from these cell lines are safe and protect White Leghorn chickens against challenge with very virulent serotype 1 MDV, similar to traditional vaccines made from CEF cells. These cell line systems can significantly reduce the costs associated with MD vaccine production. Furthermore, the increased stability of MDV and the potential for positive selection of recombinant MDV suggest that OCL will be ideal for production of more effective MDV vaccines using recombinant DNA technology. PMID- 10696444 TI - Vaccination of broilers with HVT expressing an Eimeria acervulina antigen improves performance after challenge with Eimeria. AB - The sequence encoding part of the 100 kDa refractile body protein (Ea1A) from Eimeria acervulina was cloned into the US10 locus of herpesvirus of turkeys (HVT) downstream of LTR promoter. Expression of the fusion protein was shown in vitro. Recombinant HVT showed a delayed and slightly reduced level of viremia compared to the parent strain in SPF chickens as well as in broilers. Effect on the performance of broilers vaccinated with recombinant or parent HVT was measured by challenge at day 24 with a high dose of E. acervulina and E. maxima oocysts. A significant improvement in weight of animals vaccinated with the recombinant HVT was detected at the end of the challenge period. PMID- 10696445 TI - Efficacy of a combination vaccine containing MDV CVI 988 strain and HVT against challenge with very virulent MDV. AB - With the emergence of very virulent Marek's disease virus (MDV) strains, vaccines based on herpesvirus of turkeys (HVT) appear to be not powerful enough to confer full protection, whereas in chicken flocks vaccinated with MDV CVI 988 strain protective immunity sometimes is generated not early enough for full protection. For this reason combination vaccines containing HVT as well as CVI 988 have been developed. In this paper the beneficial effect of combining both types of virus strains in one vaccine for early protection is shown in a vaccination challenge experiment, in which one-day-old chickens were vaccinated with suboptimal dosages of the monovalent vaccines and the same dosages in a combination vaccine. After 5 days the chickens were challenged with a very virulent MDV strain and subsequently observed for a period of approx. 50 days. It appeared that the combination vaccine provided better early protection than the monovalent vaccines. In addition, the combination vaccine was tested as vaccine administered in ovo. It appeared that after in ovo vaccination the vaccine conferred adequate protection against challenge with a very virulent MDV strain, 5 days after hatch, and that protection after in ovo vaccination was similar to that obtained after subcutaneous vaccination with the same combination vaccine. PMID- 10696446 TI - A recombinant fowlpox virus vaccine expressing glycoprotein B gene from CVI988/Rispens strain of MDV: protection studies in different chickens. AB - Recombinant fowlpox virus (rFPV) was constructed to express glycoprotein B (gB) gene from CVI988/Rispens strain of Marek's disease virus (MDV). The rFPV-gB/R alone and in combination with herpesvirus of turkey (HVT) preparations were evaluated for their protective efficacy against challenge with very virulent MDV strains Md5 and RB1B in different chickens. The rFPV-gB/R alone induced protection comparable to that by HVT vaccines in both Ab- SPF chickens and Ab+ production chickens. Significant protective synergism was observed in one of these two types of commercial production chickens when rFPV-gB/R was combined with HVT of either cell-associated or cell-free preparations. Immunogenesis studies showed that rFPV-gB/R, just like conventional vaccines, significantly reduced the level of viremia, splenocytes infection and feather follicle shedding of challenge virus in vaccinated chickens. PMID- 10696447 TI - Temperature-sensitive aqueous microgels. AB - An account of the preparation and characterization of temperature-sensitive aqueous microgels based on poly(N-isopropylacrylamide) was first published in 1986. Since then there has been a steady increase in the number of publications describing preparation, characterization and applications of temperature sensitive microgels. This paper reviews the important developments in the area of temperature-sensitive aqueous microgels over the last decade. Although most of the work involves gels based on poly(N-isopropylacrylamide), other polymers are also considered. Core-shell latex particles exhibiting temperature-sensitive properties are also described. PMID- 10696448 TI - Capillary effects of surfactants at curved interfaces AB - The influence of surfactant on the solvent vapour pressure and equilibrium states in open capillary systems has been considered both in the presence and absence of gravity. The relationships have been derived describing a change in vapour pressure in the course of surfactant adsorption in closed one-phase and two-phase capillary systems. The stability conditions for such systems are discussed as related to surface tension, surface elasticity, surfactant adsorption and concentration, and the interfacial curvature. PMID- 10696449 TI - Which surfactants reduce surface tension faster? A scaling argument for diffusion controlled adsorption AB - Consider the example of surfactant adsorbing from an infinite solution to a freshly formed planar interface. There is an implicit length scale in this problem, the adsorption depth h, which is the depth depleted to supply the interface with the absorbed surfactant. From a mass balance, h can be shown to be the ratio of the equilibrium surface concentration gamma eq to the bulk concentration C infinity. The characteristic time scale for diffusion to the interface is tau D = h2/D, where D is the diffusivity of the surfactant in solution. The significance of this time scale is demonstrated by numerically integrating the equations governing diffusion-controlled adsorption to a planar interface. The surface tension equilibrates within 1-10 times tau D regardless of bulk concentration, even for surfactants with strong interactions. Dynamic surface tension data obtained by pendant bubble method are rescaled using tau D to scale time. For high enough bulk concentrations, the re-normalized surface tension evolutions nearly superpose, demonstrating that tau D is indeed the relevant time scale for this process. Surface tension evolutions for a variety of surfactants are compared. Those with the smallest values for tau D equilibrate fastest. Since diffusion coefficients vary only weakly for surfactants of similar size, the differences in the equilibration times for various surfactant solutions can be attributed to their differing adsorption depths. These depth are determined by the equilibrium adsorption isotherms, allowing tau D to be calculated a priori from equilibrium surface tension data, and surfactant solutions to be sorted in terms of which will reduce the surface tension more rapidly. Finally, trends predicted by tau D to gauge what surfactant properties are required for rapid surface tension reduction are discussed. These trends are shown to be in agreement with guiding principles that have been suggested from prior structure-property studies. PMID- 10696450 TI - On the applicability of Arrhenius plot methods to determine surface energetic heterogeneity of adsorbents and catalysts surfaces from experimental TPD spectra. AB - Recovering adsorption energy distribution from experimental data belongs to most difficult problems of adsorption science. In the case when thermodesorption data are used as a source of information, that difficult problem is overcome by the common use of the Arrhenius plot methods. So, we decided to carry out an extensive model investigation to show, how reliable information concerning the surface energetic heterogeneity is obtained by using the Arrhenius plot methods. Like in our previous publications we have used the Statistical Rate Theory of Interfacial Transport to describe the adsorption/desorption kinetics. Our model investigations showed, that the Arrhenius plot methods, cannot provide reliable information about the surface energetic heterogeneity. Moreover, for strongly heterogeneous surfaces a linear relationship exists between the logarithm of the pre-exponential constant and the adsorption energy, for certain adsorption coverages. That kind of compensation effect has, so far, been ascribed to interactions between the adsorbed molecules. The failure of the popular Arrhenius plot method puts, as an urgent agenda, the development of reliable methods for recovering adsorption energy distribution from the thermodesorption data. PMID- 10696451 TI - Electrical properties of free surface of water and aqueous solutions. AB - The investigations, both experimental and theoretical, concerning the sign and the magnitude of the surface potential drop at the water/air interface and across adsorbed films presented in many papers are surveyed. PMID- 10696452 TI - Coupling of hydrodynamic and electric interactions in adsorption of colloidal particles. AB - The role of the coupling between hydrodynamic and electric interactions in adsorption of colloid particles is reviewed. First the general formulation of the problem of fluid motion connected with the macroscopic suspension flow as well as microscopic particle motion is presented together with the general description of the colloid particle adsorption. Then two limiting cases are considered: motion of uncharged particles and electric interactions of stationary particles. The theoretical consideration are supported by illustrative experimental results. Next the electrokinetic effects in particle adsorption process are considered and the theoretical background for the description of these effects is given. The effect of the electroviscous drag on the particle aggregation and adsorption kinetics is discussed. Also the electrokinetic lift force, which is an example of the non-linear electrokinetic effect, is considered and the possible implications on particle adsorption are examined. In the following part of the review the influence of electro-hydrodynamic coupling on the particle transport to the interface is considered. First the 'inverse salt effect' for the initial adsorption flux when the interface is not covered by adsorbed particles is examined. Then, the electro-hydrodynamic scattering effect which occurs when the interface is partially covered by particles is considered and its experimental evidence concerning the adsorption kinetics and the local structure of adsorbate is considered. Also a novel application of the scattering effect to measure interparticle forces the 'colloid particle collider' is presented and some experimental results obtained using this technique are shown. All results reported in this review suggest strong coupling of the hydrodynamic and electric forces and their pronounced influence on colloid particle adsorption. PMID- 10696453 TI - Surfactant layers at the air/water interface: structure and composition. AB - The use of neutron reflectometry to study the structure and composition of surfactant layers adsorbed at the air/water interface is reviewed. A critical assessment of the results from this new technique is made by comparing them with the information available from all other techniques capable of investigating this interface. PMID- 10696454 TI - Transforming growth factor-beta expression in prostate neoplasia. AB - OBJECTIVE: To understand the role of transforming growth factor (TGF) -beta 1, beta 2 and -beta 3 proteins and TGF-beta type I and II receptors in prostate neoplasia; to determine the correlation between expression of TGF-beta s and their relative receptors in the epithelial and stromal compartments of benign prostatic hyperplasia (BPH), prostatic intraepithelial neoplasia (PIN) and prostate carcinoma; and to determine whether TGF-beta and TGF-beta receptor expression is associated with the grade of tumor differentiation. STUDY DESIGN: Sixty prostate neoplasms were analyzed by immunohistochemistry using anti-TGF beta 1, -beta 2, -beta 3, -beta RI and -beta RII antibodies. RESULTS: TGF-beta and TGF-beta receptor immunoreactivity was more strongly expressed in prostate carcinoma than in PIN and BPH, and TGF-beta type I and type II receptors were less strongly expressed than TGF-beta 1-3 proteins. The difference between epithelial and stromal compartments reached significance (P < .05) for all TGF beta isoforms and related receptors only in BPH, whereas a significant difference was found for TGF-beta protein in all grades of PIN but not for prostate carcinoma tissue. Luminal epithelial cells of BPH and PIN coexpressed all three TGF-beta isoforms and preferentially TGF-beta RII. Conversely, basal epithelial cells stained strongly for TGF-beta 1, -beta 3 and -beta RI but not for TGF-beta 2 and more strongly for TGF-beta RI than -beta RII. Linear regression showed a positive correlation between TGF-beta 1 and -beta 2, between TGF-beta 2 and -beta 3 and between TGF-beta RI and -beta RII proteins in all areas. The epithelium of Gleason score 7 tumors contained significantly higher TGF-beta 2 protein levels than Gleason score 3 and 4, and 5 and 6 tumors (P < .05). CONCLUSION: Stromal and epithelial cells of malignant and nonmalignant prostatic tumors express all three TGF-beta isoforms and their related receptors. These may act as both paracrine and autocrine factors to influence prostate function and the stromal-epithelial cell interaction. TGF-beta and -beta R immunoreactivity noted in basal cells indicates that in BPH and PIN, TGF-beta Rs and signaling pathways remain intact. The overexpression of TGF-beta proteins and underexpression of TGF-beta receptors in prostate cancer could suggest a mechanism for prostate cancer cells to escape the growth inhibitory effect of TGF-beta, thus leading to a more malignant phenotype. PMID- 10696455 TI - Manual versus image analysis estimation of PCNA in breast carcinoma. AB - OBJECTIVE: To compare manual to image analysis estimation of proliferating cell nuclear antigen (PCNA) expression in paraffin sections of breast carcinomas. STUDY DESIGN: Paraffin sections of 51 breast carcinomas were stained with primary antibody to PCNA. Nuclear PCNA expression in 100 randomly selected tumor cells from marked areas was manually graded from 0 to 3. Antigen expression was also calculated by a cell analysis system (CAS-200, Becton Dickinson, Elmhurst, Illinois, U.S.A.) from marked and random microscopic fields. Obtained proliferative index (PI) from both methods was compared. RESULTS: Manually calculated PI correlated strongly with the CAS-200-calculated PI (P < .01). The highest correlation was seen between the CAS-200 PI value and manually calculated PI value using grade 2 and 3 nuclei. A particularly high correlation was noted between the number of positive nuclei and antigen staining area (P < .01) as estimated by the CAS-200. CONCLUSION: Nuclear expression of PCNA and other nuclear antigens can be accurately evaluated by an image analysis system. The speed and objectivity of such machines allow the evaluation of larger parts of tissues and provide more-representative antigen expression profiles. PMID- 10696456 TI - Conceptual comparison of two computer models of corpuscle sectioning and of two algorithms for correction of ploidy measurements in tissue sections. AB - OBJECTIVE: To compare two computer models of corpuscle sectioning and two algorithms for correction of ploidy measurements in tissue sections. STUDY DESIGN: Two models of corpuscle sectioning (the computed corpuscle sectioning program [CCSP] [Analyt Quant Cytol Histol 1997;19:376-386] and the ellipsoid sectioning program [ESP]) were run on a personal computer to generate synthetic corpuscle section data that model the sectioned nuclei in a tissue section. These synthetic data were analyzed by two algorithms for correction of ploidy measurements in tissue sections: the reference curve method (RCM) (Analyt Quant Cytol Histol 1997;19:376-386) and the method of McCready and Papadimitriou (MMP) (Analyt Quant Cytol 1983;5:117-123) for a variety of choices of section thickness and of nuclear section profile selection criteria. RESULTS: Previous recommendations (Analyt Quant Cytol Histol 1999;21:103-112) for optimization of ploidy analysis in tissue sections (selection of only center-containing sections of nuclei in ultrathin sections with a selection bias in favor of elliptical nuclear section profiles) are valid regardless of which corpuscle sectioning model and correction algorithm are employed. Perimeter correction may be desirable or necessary in some cases. The RCM has very significant advantages over the MMP, and the CCSP is more applicable to actual ploidy analysis than is the ESP. CONCLUSION: The RCM always should be used to correct ploidy measurements in tissue sections. The MMP should not be used as the sole method but, when used, should be used with and interpreted in the context of the RCM. PMID- 10696457 TI - DNA ploidy analysis of urinary tract epithelial tumors by laser scanning cytometry. AB - OBJECTIVE: To evaluate a rapid and simple method for DNA content analysis of urinary tract epithelial tumors with laser scanning cytometry (LSC). STUDY DESIGN: The subjects were 25 patients (37 specimens) who underwent surgery for urinary tract epithelial tumors. Tissue specimens of such tumors were frozen immediately after tumor resection and stored at -80 degrees C until used. Touch preparations were made and fixed in ethanol at room temperature. The cell nucleus was stained with propidium iodide solution containing RNase, and DNA ploidy was analyzed by LSC. Nuclear debris and overlapping nuclei were gated out by special statistical filters. In LSC, a normal diploid reference peak was determined by observing lymphocytes morphologically on the computer display of the instrument and/or under the microscope. RESULTS: DNA ploidy could be evaluated in all tumor tissues. The time it took from preparing the tumor specimen to the last measurement was about 40 minutes at the shortest, and measurement of all the specimens was completed within one hour. The coefficient of variation was 2.8 7.8% (mean, 4.4%). All eight specimens (100%) at grade 1 (G1) were DNA diploid, but 20% and 85.7% of the G2 and G3 cells, respectively, were DNA aneuploid. In total, 15 of the 37 specimens were DNA aneuploid. All 17 pTa-pT1 specimens (100%) were DNA diploid, but 100% and 50% of the T2 and T3 tumors, respectively, were DNA aneuploid. CONCLUSION: One can now supplement a morphologic diagnosis with useful information using LSC of touch preparations of tumors obtained at surgery or of imprints of archived, frozen specimens. LSC provides excellent DNA histograms for surgical specimens and has great potential for clinical application in pathology. PMID- 10696458 TI - Value of morphometric nuclear image analysis using the Feulgen reaction in renal cell carcinoma. AB - OBJECTIVE: To evaluate retrospectively the ability of morphometric nuclear image analysis to predict survival in patients with renal cell carcinoma. STUDY DESIGN: The subjects were 40 patients with previously untreated renal cell carcinoma. Pathologic stage was determined using Robson's stage system. Nuclear grade was assigned according to the criteria of Fuhrman et al. We used the Feulgen staining technique, which has been widely used for the histochemical assessment of nuclear DNA content. A minimum of 300 nuclei were analyzed from each subject. Five variables in morphometric nuclear image analysis were measured: nuclear area, nuclear perimeter, nuclear ellipticity, nuclear regularity and DNA content. Cox's proportional hazard model was applied to identify prognostic usefulness with respect to survival time. RESULTS: All nuclear morphometric variables but nuclear regularity correlated with tumor grade. According to univariate survival analyses, Robson stage and nuclear ellipticity revealed a prognosis on survival with statistical significance. After adjustments for age and sex, nuclear ellipticity remained the only significant prognostic factor related to survival (P < .01). The survival rates were relatively high for patients with nuclear ellipticity > 773 as compared to those with nuclear ellipticity < 773 (P < .05). CONCLUSION: These findings indicate that morphometric nuclear image analysis using the Feulgen reaction is a reliable and efficient technique and that nuclear ellipticity is the most discriminating morphometric variable for predicting the prognosis of renal cell carcinoma patients. PMID- 10696459 TI - Nuclear volume estimates in prostatic intraepithelial neoplasia. AB - OBJECTIVE: Prostatic intraepithelial neoplasia (PIN), the most likely precursor of prostatic adenocarcinoma, is divided into two grades, low and high. Pathologists may encounter difficulties in applying these criteria in daily practice. In view of the clinical significance of high grade PIN as strong predictor of carcinoma, the separation of low and high grade PIN plays an important role in patient management. The aim of the present study was to evaluate three-dimensional nuclear size estimation in normal prostatic glands, low and high grade PIN, and prostatic adenocarcinoma as an element in their classification. STUDY DESIGN: We studied 31 formalin-fixed, paraffin-embedded, whole-mounted radical prostastectomy specimens that contained foci of normal prostatic glands, low and high grade PIN, and prostatic adenocarcinoma. Hematoxylin-eosin-stained sections were selected for the stereologic estimation of volume-weighted mean nuclear volume by the "point-sampled intercepts" method. On each focus, an average of six fields of vision were systematically chosen. RESULTS: The quantitative results indicate a significant increase in nuclear volume from normal prostatic glands (mean, 209.0 micron 3; SD, 64.6 micron 3) to low grade PIN, high grade PIN and prostatic adenocarcinoma with increments of 49%, 88% and 109%, respectively (F = 29.1, P < .001). Two-group comparisons (Duncan procedure) showed differences between low and high grade PIN and prostatic adenocarcinoma (P < .01). The difference between high grade PIN and prostatic adenocarcinoma was not significant. CONCLUSION: Three-dimensional estimates of nuclear size discriminate low and high grade PIN. Lack of stereologic differences between high grade PIN and prostatic adenocarcinoma further supports high grade PIN as a precursor of prostatic adenocarcinoma. PMID- 10696460 TI - Phenylbutyrate-induced apoptosis and differential expression of Bcl-2, Bax, p53 and Fas in human prostate cancer cell lines. AB - OBJECTIVE: To assess the mechanisms of action of phenylbutyrate (PB), an investigational chemotherapeutic agent for prostate cancer (PCa), in apoptosis induction in PCa cell lines in vitro. STUDY DESIGN: We analyzed the differential expression of different apoptosis modulators, Bcl-2, Bax, p53 and Fas, for their potential role in PB-induced apoptosis using relative quantitative flow cytometry (FCM). Both androgen-dependent (LNCaP) and androgen-independent (C-4-2, PC-3-PF and DU145) human PCa cell lines were examined. RESULTS: PB induced apoptosis in PCa cell lines in a dose-dependent manner. Fifty percent apoptosis could be induced by 5-10 mM PB. Bcl-2 was down-regulated 30-75% and the Bax:Bcl-2 ratio elevated in apoptotic PCa cell lines regardless of their androgen dependency or p53 status. FCM revealed a heterogeneous stimulatory effect on the expression of Bax and Bcl-2 in PC3-PF cells at 0.5-2.5 mM PB. In a p53-positive cell line (DU145), p53 was repressed by 70% and Fas elevated sixfold with 10 mM PB. CONCLUSION: Our data show that PB-induced PCa apoptosis is associated with the relative repression of Bcl-2 and with up-regulation of Bax and Fas proteins and that this PB-induced apoptosis is independent of p53 and androgen-dependency status of PCa cell lines. PMID- 10696461 TI - Volume-weighted mean nuclear volume. Is this new prognosticator comparable in different institutions? AB - OBJECTIVE: To determine whether volume-weighted mean nuclear volume (MNV) obtained at one institution is comparable to that from other institutions. STUDY DESIGN: MNV calculated from histologic slides obtained at three hospitals- Shizuoka Prefectural Hospital (SPH), Shimada Municipal Hospital (SMH) and Shizuoka City Hospital (SCH)--were compared. Between December 1994 and June 1996, transurethral resection of bladder tumor or transurethral resection of the prostate was performed on 37 patients at SPH and 50 patients at SMH; histologic specimens from 40 cases of bladder tumors, 63 cases of normal bladder mucosa, 28 cases of benign prostatic hyperplasia and 1 case of prostate cancer were obtained. A portion of each specimen obtained at SPH or SMH was carried to SCH, and histologic slides were made at SCH using it. Using the remaining position of each specimen, histologic slides were then prepared at each hospital. Estimates of MNV were made from all histologic slides from each hospital, and the differences in MNV between the hospitals were analyzed. In addition, intraobserver and interobserver reproducibility were analyzed using 50 specimens obtained between December 1994 and August 1995. RESULTS: On linear regression analysis, comparison of MNVs calculated from the histologic slides from SPH and SCH and those calculated from SMH and SCH revealed high correlation coefficients (R = .966 and .966, respectively), and the slope of the regression line did not differ significantly from unity. The paired t test also disclosed no significant difference between MNVs calculated at the two hospitals. Furthermore, the correlation coefficients for intraobserver and interobserver reproducibility of MNV estimates were also high (R = .918 and .949, respectively). CONCLUSION: The results of this study indicate that estimates of MNV are comparable in multiple institutions, and we recommend that they be used to support subjective histologic grading. PMID- 10696462 TI - Neural network application in the discrimination of benign from malignant gastric cells. AB - OBJECTIVE: To investigate the potential value of morphometry and neural networks for the discrimination of benign from malignant gastric lesions. STUDY DESIGN: One thousand cells from 19 cases of cancer, 19 cases of gastritis and 56 cases of ulcer were selected as a training set, and an additional 4,000 cells from the same cases of cancer, gastritis and ulcer were used as a test set. Images of routinely processed gastric smears stained by the Papanicolaou technique were analyzed by a custom-made image analysis system. RESULTS: Application of the neural network gave correct classification in 96% of benign cells and 89% of malignant cells. CONCLUSION: The results indicate that the use of neural networks and image morphometry may offer useful information concerning the potential of malignancy in gastric cells. PMID- 10696463 TI - Cytofluorometric nuclear DNA content analysis of breast tissues after frozen section diagnosis. AB - OBJECTIVE: To establish a suitable method for measurement of nuclear DNA content in breast tissues from frozen storage after frozen section diagnosis. STUDY DESIGN: For fundamental research, rat liver samples preserved in a deep freezer were used. Four protocols were used (1. fixation with 70% ethanol followed by naked nuclei preparation; 2. fixation with 10% neutral buffered formalin followed by naked nuclei preparation; 3. preparation for naked nuclei prior to fixation with 70% ethanol; and 4. preparation for naked nuclei prior to fixation with 70% neutral buffered formalin). For clinical research, 13 separate fresh frozen breast tissue samples were analyzed after frozen section diagnosis. One contained a malignant phyllodes tumor (MPT) consisting of 2 components, benign epithelial cells and malignant stromal cells; 3 were benign tumors containing fibroadenoma; and 9 cases were carcinomas, consisting of 5 scirrhous, 3 papillotubular and 1 mucinous. RESULTS: Protocols 1, 2 and 3 were not suitable methods for our purpose because remaining cytoplasm or cohesive nuclei were observed. In protocol 4 the cytoplasm was completely undetectable, and nuclei were suitably separated for nuclear DNA content measurement. Benign epithelial cell component nuclei presented a diploid pattern, and the malignant stromal cell component nuclei indicated a euploid pattern in MPT. All 3 cases of benign constituents in fibroadenoma showed a diploid pattern, as did the 3 carcinoma cases (1 mucinous, 1 scirrhous and 1 papillary). Four scirrhous and 2 papillary carcinomas showed an aneuploid pattern. CONCLUSION: Our findings show that it is possible to measure nuclear DNA content of human frozen storage tissues after frozen section diagnosis. PMID- 10696464 TI - New concepts and methods of standardizing predictive value, accuracy and incorrect diagnostic rate. AB - OBJECTIVE: To express the value of a diagnostic test under standardized and comparable conditions. STUDY DESIGN: Four new concepts of standardizing positive predictive value (SPPV), standardizing negative predictive value (SNPV), standardizing accuracy (SAc) and standardizing an incorrect diagnostic test were developed. The theoretical positive predictive value (SPPV), theoretical negative predictive value (SNPV), theoretical accuracy (SAc) and theoretical incorrect diagnosis rate (SIDR), which are not affected by a different constituent ratio of disease and nondisease groups and are obtained under the theoretical standard condition that the sample size in the disease group equals that in the nondisease group, were defined. Based on these concepts and the principles and methods of statistics and evaluation of diagnostic tests, corresponding formulas were deduced. RESULTS: The formulas are: SPPV = a(b + d)/[a(b + d) + b(a + c)] = Se/(1 + Se - Sp), SNPV = d(a + c)/[c(b + d) + d(a + c)] = Sp/(1 - Se + Sp), SAc = [a(b + d) + d(a + c)]/[2(a + c)(b + d)] = (Se + Sp)/2, and SIDR = [b(a + c) + c(b + d)]/[2(a + c)(b + d)] = (2 - Se - Sp)/2. Here, a, b, c and d refer to the case numbers of true positives, false positives, false negatives and true negatives; Se and Sp refer, respectively, to sensitivity and specificity. CONCLUSION: SPPV, SNPV, SAc and SIDR are very useful for expressing and evaluating the value of a diagnostic test under standardized and comparable conditions. PMID- 10696465 TI - Studying the growth of cervical carcinoma nests and angiogenesis by immunostaining, quantitation and three-dimensional structural analysis. AB - OBJECTIVE: To analyze the relationship and mutual effect of the growth of cervical carcinoma nests and angiogenesis. STUDY DESIGN: Serial paraffin sections of cervical squamous carcinoma were stained in repeated order with hematoxylin eosin (HE), immunostain for factor VIII-related antigen, type IV collagen and proliferating cell nuclear antigen (PCNA). Three-dimensional reconstructions were made, and the volumes of carcinoma nests, necrosis and microvessels were measured. RESULTS: Two types of cervical carcinoma nests were distinguished on the basis of their growth characteristics and vascularity (groups I and II). Group I nests: The carcinoma cells were proliferating actively, as determined by their morphology and PCNA staining. There were abundant microvessels. Some endothelial sprouts and cords penetrated the nests and then formed networks and new vessels. The volume ratio of microvessels, including sprouts and cords, to the nests was 0.6282:1. Group II nests: The center of these nests underwent necrosis. The peripheral cells were rather small, with no mitosis. The PCNA index was rather low; these nests grew very slowly. There were only a few surrounding microvessels with no endothelial sprouts or cords. The volume ratio of vessels to nest was 0.0657:1. CONCLUSION: Two types of cervical carcinoma nests show a close relationship and mutual effect of the growth of carcinoma nest and angiogenesis. Group I nests grow and develop well, with abundant microvessels. Thus, many tumor cells may be angiogenic and induce angiogenesis; growth of the nests seemed dependent on adequate neovascularization. Group II nests grow slowly, with a few microvessels; the center of the nests undergoes necrosis. The inadequate blood supply must be one of the important causes of necrosis. Considering that there must have been abundant neovascularization during their growth in the past, most of the microvessels must have been obliterated and then reabsorbed to make the remaining vessels so few. PMID- 10696466 TI - Three-dimensional imaging of tumor angiogenesis. AB - OBJECTIVE: To three-dimensionally visualize the microvessel environment of tumor angiogenesis by confocal laser scanning microscopy (CLSM). STUDY DESIGN: To reveal underlying mechanisms of tumor angiogenesis, a 7, 12-dimethylbenz(a) anthracene-induced rat cancer model was used. For demonstrating tumor vasculature, fluorescence injection method (FITC-conjugated gelatin solution) was employed. FITC gelatin was injected into the left ventricle of the rat heart. After complete perfusion, the mammary glands were resected, fixed under ice cold conditions and subjected to immunohistochemistry (IHC) for tumor cells. The LSM 410 (Carl Zeiss, Jena, Germany) was employed on thick sections (300-2,000 microns) to elucidate detailed microvessel networks (MVN) and tumor cells. RESULTS: Tumor vasculature on thick sections was clearly detected by CLSM at the maximum focus depth of 2,000 microns. Three-dimensional (3-D), reconstructed images of normal mammary glands showed regular and linear MVN. In DMBA-induced mammary cancer, vascular density of MVN was markedly increased and showed an anastomosing, irregular MVN pattern. Furthermore, focal segmentation and tortuous, branching patterns of microvessels were also seen. CONCLUSION: Application of the fluorescence injection method and IHC using CLSM was very useful for studying the 3-D relationship between tumor angiogenesis and neoplastic epithelial changes. These results suggest that application of this technique is ideal for studying 3-D imaging of tumor angiogenesis. PMID- 10696467 TI - Evidence for plasmid-mediated chemotaxis of Pseudomonas putida towards naphthalene and salicylate. AB - A naphthalene (Nap) and salicylate (Sal) degrading microorganism, Pseudomonas putida RKJ1, is chemotactic towards these compounds. This strain carries a 83 kb plasmid. A 25 kb EcoRI fragment of the plasmid contains the genes responsible for Nap degradation through Sal. RKJ5, the plasmid-cured derivative of RKJ1, is neither capable of degradation nor is chemotactic towards Nap or Sal. The recombinant plasmid pRKJ3, which contained a 25 kb EcoRI fragment, was transferred back into the plasmid-free wild-type strain RKJ5, and the transconjugant showed both degradation and chemotaxis. The recombinant plasmid pRKJ3 was also transferred into motile, plasmid-free P. putida KT2442. The resulting transconjugant (RKJ15) showed chemotaxis towards both Nap and Sal. Two mutant strains carrying deletions in pRKJ3 (in KT2442) with phenotypes Nap- Sal+ and Nap- Sal-, were also tested for chemotaxis. It was found that the Nap- Sal+ mutant strain showed chemotaxis towards Sal only, whereas the Nap- Sal- mutant strain is non-chemotactic towards both the compounds. These results suggest that the metabolism of Nap and Sal may be required for the chemotactic activity. PMID- 10696468 TI - Diversity in the yeast Cryptococcus albidus and related species as revealed by ribosomal DNA sequence analysis. AB - Evidence accumulated from studies based on physiological, biochemical, and molecular characteristics has pointed to the heterogeneity of the ubiquitous anamorphic basidiomycetous yeast species Cryptococcus albidus (Saito) Skinner, with its current varieties and synonyms. The taxonomic status of this species has not been reappraised because different studies, mostly involving limited numbers of strains, have not been integrated. To assess species diversity within the clade containing Cryptococcus albidus and other phylogenetically related Cryptococcus and Filobasidium species, we determined ribosomal DNA (rDNA) sequences of 69 strains from the 5' end of the 26S gene, D1/D2 region, and in some cases, the non-coding ITS2 region. Analysis of the sequence data together with available physiological, biochemical, and molecular characteristics, showed the segregation of C. albidus into at least 12 species, leading to the elevation of former varieties to the rank of species (C. aerius, C. diffluens), the reinstatement of synonyms (C. liquefaciens, C. terricola), and the proposal of new species (C. arrabidensis, C. chernovii, C. cylindricus, C. oeirensis, C. phenolicus, C. saitoi, C. uzbekistanensis, C. wieringae). The overall analyses of the results argue in favour of the use of rDNA sequence data to improve species delineation when integrated with other available physiological and molecular characteristics. PMID- 10696469 TI - Influence of growth and environmental conditions on cell surface hydrophobicity of Pseudomonas fluorescens in non-specific adhesion. AB - The relative cell surface hydrophobicity (CSH) of 18 soil isolates of Pseudomonas fluorescens, determined by phase exclusion, hydrophobic interaction chromatography (HIC), electrostatic interaction chromatography (ESIC), and contact angle, revealed large degrees of variability. Variation in the adhesion efficiency to Macrophomina phaseolina of the hyphae/sclerotia of these isolates was also examined. Two such isolates with maximum (32.8%; isolate 12-94) and minimum (12%; isolate 30-94) CSH were selected for further study. Early- to mid log exponential cells of these isolates were more hydrophobic than those in stationary phase, and the CSH of these isolates was also influenced by fluctuations in temperatures and pH. Isolate 12-94 exhibited high CSH (32.3%) at 30 degrees C, compared to lower values (28-24%) in the higher temperature range (35-40 degrees C). Increasing concentrations of either Zn2+, Fe3+, K+, and Mg2+ in the growth medium were associated with the increased CSH. Trypsin, pepsin, and proteinase K (75 to 150 micrograms.mL-1) reduced the CSH of isolate 12-94 cells. CSH was reduced, following exposure to DTT, SDS, Triton X-100, or Tween 80. Prolonged exposure of cells to starvation (60 days) also caused a significant decline in CSH. Several protein bands (18, 21, 23, 26 kDa) of the outer cell membrane were absent in 60-day starved cells compared to unstarved cells. In conclusion, our findings demonstrate that CSH of P. fluorescens isolates may contribute to nonspecific attachment/adhesion onto M. phaseolina hyphae/sclerotia, and the efficiency of adhesion is regulated by growth and other environmental conditions. PMID- 10696470 TI - Hydrocarbon-degrading filamentous fungi isolated from flare pit soils in northern and western Canada. AB - Sixty-four species of filamentous fungi from five flare pits in northern and western Canada were tested for their ability to degrade crude oil using gas chromatographic analysis of residual hydrocarbons following incubation. Nine isolates were tested further using radiorespirometry to determine the extent of mineralization of model radiolabelled aliphatic and aromatic hydrocarbons dissolved in crude oil. Hydrocarbon biodegradation capability was observed in species representing six orders of the Ascomycota. Gas chromatography indicated that species capable of hydrocarbon degradation attacked compounds within the aliphatic fraction of crude oil, n-C12-n-C26; degradation of compounds within the aromatic fraction was not observed. Radiorespirometry, using n-[1-14C]hexadecane and [9-14C]phenanthrene, confirmed the gas chromatographic results and verified that aliphatic compounds were being mineralized, not simply transformed to intermediate metabolites. This study shows that filamentous fungi may play an integral role in the in situ biodegradation of aliphatic pollutants in flare pit soils. PMID- 10696471 TI - Three new ascomycetous yeasts from insect-associated arboreal habitats. AB - A new species of Pichia and two new species of Candida are described and were determined to be genetically isolated from all other currently recognized ascomycetous yeasts from their sequence divergence in the species-variable D1/D2 domain of large subunit (26S) ribosomal DNA. The three species were primarily isolated from the frass of wood-boring insects living in pine and spruce trees. The new species and their type strains are the following: Pichia ramenticola NRRL YB-1985 (CBS 8699), mating type alpha (NRRL YB-3835, CBS 8700, mating type a), Candida piceae NRRL YB-2107 (CBS 8701), and Candida wyomingensis NRRL YB-2152 (CBS 8703). Pichia ramenticola and C. piceae assimilate methanol as a carbon source; P. ramenticola is the first known heterothallic ascomycetous yeast to utilize this compound. PMID- 10696472 TI - Characterization and overproduction of the Escherichia coli appA encoded bifunctional enzyme that exhibits both phytase and acid phosphatase activities. AB - The appA gene that was previously shown to code for an acid phosphatase instead codes for a bifunctional enzyme exhibiting both acid phosphatase and phytase activities. The purified enzyme with a molecular mass of 44,708 Da was further separated by chromatofocusing into two isoforms of identical size with isoelectric points of 6.5 and 6.3. The isoforms had identical pH optima of 4.5 and were stable at pH values from 2 to 10. The temperature optimum for both phytase isoforms was 60 degrees C. When heated at different pH values the enzyme showed the greatest thermal resistance at pH 3. The pH 6.5 isoform exhibited K(m) and Vmax values of 0.79 mM and 3165 U.mg-1 of protein for phytase activity and 5.5 mM and 712 U.mg-1 of protein for acid phosphatase, respectively. The pH 6.3 isoform exhibited slightly lower K(m) and Vmax values. The enzyme exhibited similar properties to the phytase purified by Greiner et al. (1993), except the specific activity of the enzyme was at least 3.5-fold less than that previously reported, and the N-terminal amino acid sequence was different. The Bradford assay, which was used by Greiner et al. (1993) for determination of enzyme concentration was, in our hands, underestimating protein concentration by a factor of 14. Phytase production using the T7 polymerase expression system was enhanced by selection of a mutant able to grow in a chemically defined medium with lactose as the carbon source and inducer. Using this strain in fed-batch fermentation, phytase production was increased to over 600 U.mL-1. The properties of the phytase including the low pH optimum, protease resistance, and high activity, demonstrates that the enzyme is a good candidate for industrial production as a feed enzyme. PMID- 10696473 TI - Growth kinetics of Escherichia coli with galactose and several other sugars in carbon-limited chemostat culture. AB - Kinetic models for microbial growth describe the specific growth rate (mu) as a function of the concentration of the growth-limiting nutrient (s) and a set of parameters. A typical example is the model proposed by Monod, where mu is related to s using substrate affinity (Ks) and the maximum specific growth rate (mu max). The preferred method to determine such parameters is to grow microorganisms in continuous culture and to measure the concentration of the growth-limiting substrate as a function of the dilution rate. However, owing to the lack of analytical methods to quantify sugars in the microgram per litre range, it has not been possible to investigate the growth kinetics of Escherichia coli in chemostat culture. Using an HPLC method able to determine steady-state concentrations of reducing sugars, we previously have shown that the Monod model adequately describes glucose-limited growth of E. coli ML30. This has not been confirmed for any other sugar. Therefore, we carried out a similar study with galactose and found steady-state concentrations between 18 and 840 micrograms.L-1 for dilution rates between 0.2 and 0.8.h-1, respectively. With these data the parameters of several models giving the specific growth rate as a function of the substrate concentration were estimated by nonlinear parameter estimation, and subsequently, the models were evaluated statistically. From all equations tested, the Monod model described the data best. The parameters for galactose utilisation were mu max = 0.75.h-1 and Ks = 67 micrograms.L-1. The results indicated that accurate Ks values can be estimated from a limited set of steady-state data when employing mu max measured during balanced growth in batch culture. This simplified procedure was applied for maltose, ribose, and fructose. For growth of E. coli with these sugars, mu max and Ks were for maltose 0.87.h-1, 100 micrograms.L-1; for ribose 0.57.h-1, 132 micrograms.L-1, and for fructose 0.70.h 1, 125 micrograms.L-1. PMID- 10696474 TI - Role of hsfA gene on host-specificity by Bradyrhizobium japonicum in a broad range of tropical legumes. AB - Bradyrhizobium japonicum mutant strain NAD163, containing a 30-kb deletion mutant encompassing the hsfA gene, was inoculated onto a broad range of legume species to test host-specificity. Most legume species formed ineffective nodules except Vigna angularis var. Chibopat and Glycine max var. Pureunkong. A hsfA insertion mutant, BjjC211, gave similar results to strain NAD163, implying that many legume species require HsfA for host-specific nitrogen fixation. To determine whether other genes in the deleted region of NAD163 are also necessary, the hsfA gene was conjugally transferred into the NAD163 mutant. The transconjugant formed effective nodules on the host legume plants, which earlier had formed ineffective nodules with mutant NAD163. Thus, we conclude that the hsfA gene in the 30-kb region is the only factor responsible for host-specific nitrogen fixation in legume plants. PMID- 10696475 TI - Chitinolytic bacteria in the minke whale forestomach. AB - Minke whales consume large amounts of pelagic crustaceans. Digestion of the prey is initiated by indigenous bacteria in a rumen-like forestomach system. A major structural component of the crustacean exoskeleton is chitin, the beta-1,4-linked polymer of N-acetyl-D-glucosamine. The exoskeletons appear to dissolve completely in the non-glandular forestomach. Bacteria in the forestomach fluid of six krill eating minke whales were enumerated and isolated using an anaerobic habitat simulating culture medium. Median viable population densities ranged between 6.0 x 10(6) and 9.9 x 10(9) bacterial cells per mL forestomach fluid. Bacterial isolates (n = 44) cultured from the forestomach fluid of one minke whale mainly resembled strains of Eubacterium (25%), Streptococcus (18%), Clostridium (14%), and Bacteroides (11%). As much as 12% of the bacterial isolates were chitinolytic, while beta-N-acetylglucosaminidase activity was demonstrated in 54% of the isolates, and utilisation of N-acetyl-D-glucosamine was observed in 73%. The chitinolytic isolates resembled strains of Bacteroides, Bacteroidaceae, Clostridium, and Streptococcus. Scanning and transmission electron microscopy of partly digested krill from the minke whale forestomach revealed bacteria close to and inside the chitinous exoskeleton. The bacterial chitinase may act on the chitinous crustacean exoskeletons, thereby allowing other bacteria access to the nutritious soft inner tissues of the prey, and thus initiating its degradation and fermentation. PMID- 10696476 TI - Some probiotic properties of chicken lactobacilli. AB - The beneficial effect of lactobacilli has been attributed to their ability to colonize human and animal gastrointestinal tracts. In this work, adhesion assays with three lactobacillus strains and intestinal fragments obtained from chickens were assessed. Lactobacillus animalis and L. fermentum were able to adhere to three kinds of epithelial cells (crop, small and large intestines) with predominance to small intestine. Among the strains considered, L. fermentum subsp. cellobiosus showed the lowest and L. animalis the highest adhesion ability. Scanning electron microphotographs showing L. animalis and L. fermentum adhering to intestinal cells were obtained. The characterization of L. animalis adhesion indicated that lectin-like structure of this strain has glucose/mannose as specific sugars of binding. However, a calcium requirement was not observed. The adhesion of L. fermentum was reduced by addition of sialic acid or mannose (P < 0.01). These carbohydrates can be involved in the interaction between adhesin and epithelial surface. In this case, the dependence on bivalent cations was demonstrated. Lactobacillus fermentum was effective in reducing the attachment of Salmonella pullorum by 77%, while L. animalis was able to inhibit (90%, 88%, and 78%) the adhesion of S. pullorum, S. enteritidis, and S. gallinarum to host specific epithelial fragments respectively. Our results from this in vitro model suggest that these lactobacilli are able to block the binding sites for Salmonella adhesion. PMID- 10696477 TI - Inhibition of pathogenic Salmonella enteritidis growth mediated by Escherichia coli microcin J25 producing strains. AB - For the first time, microcin-producing strains showing inhibitory activities against enteropathogen Salmonella enteritidis were isolated from poultry intestinal contents. Among the numerous strains isolated, two strains of Escherichia coli, named J02 and J03, showing the greatest activities against S. enteritidis, were studied. Biochemical tests and purification identified the main antagonist compound produced as microcin J25. In order to evaluate the protective potential of E. coli J02 and J03 against S. enteritidis infection, the ability of these strains to inhibit growth of S. enteritidis was investigated in mixed culture. A strong antagonist activity was obtained with a preculture phase of the active strain in minimal medium before incubation with S. enteritidis. In a bioreactor experiment simulating the chicken gastric and intestinal tract environment, a mixture of the two strains E. coli J02 and J03, provided an enhanced inhibitory effect. Microcinogenic strain activities were not affected by bile, pancreatic enzymes addition, or acidic conditions. These results suggest the relevant role of microcin-producing microorganisms in microbial intestinal ecology. To conclude, this study shows that microcin J25 strains could exert a beneficial protective effect against S. enteritidis growth in situ. PMID- 10696478 TI - Removal of silver from photographic wastewater effluent using Acinetobacter baumannii BL54. AB - Acinetobacter baumannii BL54, a silver (Ag) resistant micro-organism was isolated from clinical samples collected at the Armed Forces Medical College hospital in Pune, India. The strain BL54 removed a high quantity of silver (2.85 mg/g biomass) from photographic wastewater effluent. Treatment of the cells with 10 mM EDTA or agitating the culture did not affect the removal process, while altering pH of the wastewater or pre-treating the cells with 0.5 mM 2,4-dinitrophenol (DNP), 20 microM N,N'-dicyclohexylcarbodiimide (DCC), 25 micrograms/mL cefotaxime, and polymyxin-B resulted in considerable decrease in removal of silver by the organism. Dead cells, or a Ags plasmid-cured derivative (BL54.1) removed little silver, which was mainly surface bound. The results, compared with accumulation of Ag by a sensitive culture of Escherichia coli K12 J53.2, suggest that A. baumannii BL54 has good potential for bioremediation of silver from photographic wastewater effluents. PMID- 10696479 TI - 16S rDNA restriction fragment length polymorphism analysis of psychrotrophic vibrios from Japanese coastal water. AB - Restriction fragment length polymorphism (RFLP) analysis was carried out for 136 natural isolates belonging to the family Vibrionaceae. These were collected from inshore areas of Japan, mainly in winter. Twenty-eight 16S rDNA genotypes were obtained by digestion with four restriction endonucleases (HhaI, DdeI, RsaI, and Sau3AI). To estimate the genetic relationships, 53 informative fragments were scored by their presence or absence. A dendrogram was constructed using the unweighted pair group method with the arithmetic averages algorithm. Five RFLP groups (groups I to V) were obtained. Group I corresponded to Vibrio splendidus like strains. It was confirmed that this group was not only found in Otsuchi Bay, but also in broad coastal areas of Japan. Group II strains were not identified as previously known Vibrio species. Group III strains were regarded as members of the Vibrio main group, which is a major phylogenetic group deduced from 16S rRNA gene analysis in the family Vibrionaceae. The RFLP profile indicated that Group IV strains were closely related to V. hollisae. Group V strains showed RFLP patterns which have not been observed previously. From the clustering analysis, it was concluded that group V strains were not Vibrio species. Most of the isolates studied were not identified as previously described species. It suggests that many psychrotrophic vibrios in cold marine environments remain as unknown species. PMID- 10696480 TI - Expression of ptxR and its effect on toxA and regA expression during the growth cycle of Pseudomonas aeruginosa strain PAO1. AB - The expression of the toxA and regA genes in Pseudomonas aeruginosa is negatively regulated by iron at the transcriptional level. We have previously described ptxR, an exotoxin A regulatory gene which appears to enhance toxA expression through regA. In this study, we have tried to determine if ptxR expression correlates with its effect on toxA and regA expression throughout the growth cycle of P. aeruginosa strain PAO1. This was done using Northern blot hybridization experiments (with toxA, regA, and ptxR probes), and ptxR transcriptional fusion studies. To avoid problems related to the presence of multiple copies of ptxR in PAO1, we have constructed a PAO1 strain (PAO1-XR) that carries only two ptxR genes in its chromosome. Our results showed that when PAO1 XR was grown in iron-limited conditions, the increase in exotoxin A activity and the accumulation of toxA mRNA appeared at about mid- to late-exponential phase. A similar increase in the accumulation of regA mRNA was detected. Both regA transcripts, T1 and T2, were enhanced in PAO1-XR. In iron-sufficient medium, neither toxA nor regA mRNA was detected at any time point in the growth cycle of PAO1-XR. In contrast, the accumulation of ptxR mRNA was detected throughout the growth cycle of PAO1-XR under both iron-deficient and iron-sufficient conditions. The presence of iron in the growth medium also had no effect on the level of beta galactosidase activity produced by a ptxR-lacZ fusion in PAO1. These results suggest that (i) the enhancement in toxA expression by ptxR correlates with the enhancement in regA expression; (ii) ptxR affects the expression of the regA P1 and P2 promoters; (iii) ptxR expression precedes its effect on toxA and regA expression; and (iv) unlike toxA and regA, the overall expression of ptxR throughout the growth cycle of PAO1 is not negatively regulated by iron. PMID- 10696482 TI - Effects of iron and manganese availability on growth and production of superoxide dismutase by Streptococcus suis. AB - A complex medium supported good growth of the type strain of Streptococcus suis irrespective of the presence or absence of a high concentration (1 microM) of the iron chelating agent, ethylenediamine di-o-hydroxyphenylacetic acid. Good growth was also obtained using a complex medium that had been treated with Chelex-100 to reduce the iron content, but only if this medium was supplemented with manganese; supplementation with iron had little effect. Collectively, these results indicate that S. suis requires manganese, but not iron, for growth. Polyacrylamide gel electrophoresis of cell extracts followed by activity staining revealed the presence of a single manganese-cofactored superoxide dismutase; activity staining and enzyme assays revealed that manganese availability during growth affected the activity of the superoxide dismutase in cell extracts. The results are discussed with respect to the capacity of S. suis to avoid damage by reactive oxygen species. PMID- 10696481 TI - Isolation and characterization of mini-Tn10 lipopolysaccharide mutants of Actinobacillus pleuropneumoniae serotype 1. AB - Lipopolysaccharide (LPS) has previously been identified as the major adhesin of Actinobacillus pleuropneumoniae involved in adherence to porcine respiratory tract cells. The purpose of the present study was to isolate and characterize mutants in LPS biosynthesis by using a mini-Tn10 transposon mutagenesis system. Seven mutants appeared to possess a rough LPS (among which two had similar Southern blot profiles) while one mutant (#5.1) expressed the high-molecular-mass LPS, but as visualized by Tricine SDS-PAGE, showed an additional band in the core lipid A region. The LPS mutants showed sensitivity to pig serum to various degrees, while the parent strain was serum-resistant. Use of piglet frozen tracheal sections indicated that, surprisingly, the rough LPS mutants adhered similarly or in greater numbers than the parent strain. However, the LPS mutant #5.1 adhered significantly less than the parent strain and was also less virulent in pigs. The gene affected by mini-Tn10 in LPS mutant #5.1 is galU, the structural gene for UTP-alpha-D-glucose-1-phosphate uridylyltransferase, involved in LPS core biosynthesis. Complementation analysis confirmed that the phenotypic characteristics of LPS mutant #5.1 are the result of the inactivation of the galU gene. Our data suggest that although the presence of O-antigen does not seem to be essential, an intact core-lipid A region might be required for adherence of A. pleuropneumoniae to porcine respiratory tract cells. To the best of our knowledge, these mutants represent the first isogenic mutants of A. pleuropneumoniae defective in LPS biosynthetic genes. PMID- 10696484 TI - Effect of carbon source, growth and temperature on the expression of the sec genes of Streptomyces lividans 1326 AB - The mRNA level in sec genes of Streptomyces lividans was studied as a function of growth temperature, glucose effect, and growth using two different carbon sources. Glucose and xylan, a complex hemicellulose, were used as carbon sources for the growth of S. lividans. For both substrates, the mRNA levels of secA, secD, secE, secF, and secY genes were almost constant during the early and log phases, but showed a marked decrease at the beginning of the stationary phase followed by a full recovery of mRNA level in the late stationary phase. This indicates that the sec genes are actively transcribed during the differentiation process. The mRNA level in xylan was generally from 1.5- to 2-fold that in glucose. At growth temperatures of 28 degrees C, 34 degrees C, or 40 degrees C, there was no significant difference in the sec gene mRNA levels. PMID- 10696483 TI - Multiple environmental factors regulate the expression of the carbohydrate selective OprB porin of Pseudomonas aeruginosa. AB - In response to low extracellular glucose concentration, Pseudomonas aeruginosa induces the expression of the outer membrane carbohydrate-selective OprB porin. The promoter region of the oprB gene was cloned into a lacZ transcriptional fusion vector, and the construct was mobilized into P. aeruginosa OprB-deficient strain, WW100, to evaluate additional environmental factors that influence OprB porin gene expression. Growth temperature, pH of the growth medium, salicylate concentration, and carbohydrate source were found to differentially influence porin expression. This expression pattern was compared to those of whole-cell [14C]glucose uptake under conditions of high osmolarity, ionicity, variable pH, growth temperatures, and carbohydrate source. These studies revealed that the high-affinity glucose transport genes are down-regulated by salicylic acid, differentially regulated by pH and temperature, and are specifically responsive to exogenous glucose induction. PMID- 10696485 TI - Antigenic and genetic characterization of a putative hybrid transferrin-binding protein B from Neisseria meningitidis. AB - The transferrin-binding protein Bs (TbpBs) from the bacterium Neisseria meningitidis have been divided into two families according to genetic and antigenic features. TbpB from meningococcal strain B385 showed a molecular mass similar to that exhibited by TbpBs belonging to the high molecular mass family of TbpBs. TbpB was recognized by immunoassay using a specific serum directed against the TbpB of the reference strain for this family (strain M982). It was also recognized by a serum elicited against the TbpB of the reference strain for the low molecular mass family (strain B16B6). The tbpB gene from strain B385 was cloned and sequenced. The highest degree of sequence homology was found to be with the TbpBs belonging to the high molecular mass family, although a region of 14 amino acids that is only present in the TbpB from strain B16B6 was also found. This report illustrates a TbpB that shows hybrid antigenic and genetic behaviour. PMID- 10696486 TI - Welcome 2000: concern for the patient, collaboration with colleagues, and confidence for the future. PMID- 10696487 TI - Canadian psychiatry across 5 decades: from clinical inference to evidence-based practice. AB - OBJECTIVE: To examine the evolution of the epistemological basis of Canadian psychiatry over the last 50 years. METHOD: A comparison of the content of The Canadian Journal of Psychiatry in its early years, then known as the Canadian Psychiatric Association Journal, with publications from recent years, shedding light on broader trends in psychiatric theory and practice. RESULTS: A dramatic change has occurred in Journal content, from clinical inference to an evidence based approach, reflecting stronger biological orientations, as well as empirical psychosocial research. CONCLUSIONS: Epistemological changes in psychiatry are a positive development. The future should bring an even stronger neuroscience base as well as an evidence-based approach to psychotherapy. PMID- 10696488 TI - Psychiatry in the new millennium. AB - The field of psychiatry is experiencing great excitement at this time, much as it was 100 years ago. Current excitement is rooted in the greatly strengthened therapeutics, new models for understanding, and an exponential increase in knowledge of brain function as well as in the opportunity to revise health care delivery. While public expectations of professionals have generally fallen, the role of the healer, which is at the heart of psychiatric practice, has remained high in public regard. Psychiatry has also had to develop new relationships with an active consumer movement. Consumers now appropriately expect to be part of the planning, governance, and evaluation of care. Patients are questioning the research agenda and demand a role in determining the conduct of investigations. This active consumer movement is playing an important role in destigmatizing mental illnesses. Newer, nonmoralistic theories about mental illness and the profession's emphasis on the public trust have also played an important role. The increasing closeness of psychiatry to the rest of medicine has had a greatly beneficial impact, not only on stigma but also on diagnosis and treatment. Care must be taken, however, to see that diagnosis does not become a means to avoiding understanding of people. A welcome recent change has been the reunion between psychiatry and the addictions. This reunion has been facilitated by the development of multifactorial models of care in the mental health field and harm reduction strategies in the addictions. This bodes well for more integrated treatment in the coming years. The strong psychiatric treatments that are now available and those on the horizon also auger well for an exciting period in our field. The excitement is enhanced by recognizing the multiple approaches to care that have been demonstrated to be effective and the need for great investment in research in this country and by developing new partnerships between the profession and public. PMID- 10696489 TI - Ethical issues concerning participants in community surveys of child and adolescent mental disorders. AB - OBJECTIVE: Considering that a literature review yielded limited information on ethical issues concerning participants in child and adolescent mental health community surveys, the authors identify and discuss some of these issues. METHOD: First, the authors present the ethical principles set forth by the National Council on Bioethics in Human Research (NCBHR) and evoked by the 1998 Tri-Council Policy Statement, underscoring their importance as guidelines for establishing ethical standards for research with children. Second, they describe the general objectives and currently preferred methods of child psychiatric surveys. Third, they discuss issues pertaining to the validity and innocuousness of structured interview guides, the limitations of parental authorization and children's assent, and the complexity of clinical interventions based on ethical grounds. CONCLUSIONS: The authors emphasize the importance of developing empirical knowledge regarding the questions raised and bringing the social stakeholders concerned into the debate. PMID- 10696490 TI - Homeless shelter users in the postdeinstitutionalization era. AB - OBJECTIVE: To describe the psychiatric symptomatology and mental health service needs of homeless shelter users in Calgary, Alberta. Data were collected as part of a broad-based community action initiative designed to reduce the problem of homelessness. METHODS: A semistructured interview was conducted with a representative sample of 250 emergency shelter users. Mental health problems were measured through self-reports of 9 psychiatric symptoms known to be related to illnesses prevalent among homeless populations (depression, anxiety, and psychoses). The CAGE alcohol screen was also used. RESULTS: Three-quarters of the sample expressed some symptomatology. About one-third were estimated to have a significant mental health problem. The lifetime prevalence of alcohol abuse was 33.6%. Higher levels of psychiatric symptomatology appeared to relate to a wide range of hardships, personal and public health risks, addictive behaviours, victimization, economic and interpersonal life events, dissatisfaction, and stress. Also, those with significant symptomatology frequently needed mental health care services but often did not know where to access them. CONCLUSIONS: The prevalence of mental health and substance abuse problems within homeless populations is significant and associated with considerable hardship as well as personal and public health risks. PMID- 10696491 TI - Anxiety, significant losses, depression, and irrational beliefs in first-offence shoplifters. AB - OBJECTIVE: To evaluate the relationship among demographic data, anxiety, significant losses, depression, and irrational beliefs reported by first-offence shoplifters. METHOD: One hundred and six adult shoplifters who were first-time offenders completed a self-administered questionnaire. RESULTS: Men and women were equally likely to be arrested for this offence. The majority of offenders were poor and unemployed. Depression, but not anxiety, was the most common psychiatric disorder associated with shoplifting. Subjects with depression presented the greatest number of irrational beliefs related to shoplifting. CONCLUSIONS: The authors suggest 2 categories of shoplifters: those who shoplift through rational choice; and those for whom shoplifting is a response to depression or leads to the fulfillment of some psychological needs. In conclusion, shoplifting does not have a unitary motive, and the clinical implications are that the affective and cognitive aspects of shoplifters' behaviours must be taken into account. PMID- 10696492 TI - The epidemiology of major depression: implications of occurrence, recurrence, and stress in a Canadian community sample. AB - OBJECTIVE: To study the effects of stress process variables on the prevalence of major depressive disorder (MDD) and to explore the factors related to its onset and recurrence, using measures of stress and disorder that are more comprehensive than those in previous studies of depression. METHOD: Data were collected from a randomly selected community sample of 1393 adult respondents in Toronto, Ontario. Depression was measured using the Composite International Diagnostic Interview (CIDI). RESULTS: Bivariate and multivariate analyses examine demographic and stress process correlates of MDD to assess their impact on both occurrence and recurrence. The effects of childhood experiences on lifetime MDD are examined as risk factors for the initial onset of depression. With respect to occurrence, the multivariate analyses tended to show agreement with established findings. Results using a subsample of people with lifetime depression to predict recurrence tended to mirror results using the full sample, except with respect to gender, birthplace, and, to a certain degree, stress differences. Finally, traumas experienced in childhood, as well as early childhood experiences involving parental substance abuse and mental health problems, were significant predictors of the onset of major depressive disorder. CONCLUSION: This research currently represents the only Canadian community study to examine the relationship between stress, social support, coping, and depression using outcome measures that approximate Diagnostic and Statistical Manual of Mental Disorders (DSM) criteria. In addition, it is among the few that use a comprehensive measure of life stress encompassing childhood experiences and current operant stress (both events and chronic problems). PMID- 10696493 TI - The predictive value of dysgraphia and constructional apraxia for delirium in psychiatric inpatients. AB - OBJECTIVE: To evaluate the predictive value of dysgraphia and constructional apraxia for delirium among psychiatric inpatients. METHOD: Data were collected from 2 sources. First, a series of nondelirious psychiatric inpatients that had participated in a previous study was selected to determine the specificity of various indices of dysgraphia and constructional apraxia. Second, a series of 56 psychiatric inpatients with delirium as identified using electronic administrative data and clinical records was selected to evaluate sensitivity. RESULTS: Of the various indices of dysgraphia examined, only a global rating of writing quality and evidence of jagged or angled letter loops were informative clinical signs. The predictive value of constructional apraxia resembled the predictive value of the 2 dysgraphia indices. CONCLUSIONS: Dysgraphia and constructional apraxia are useful clinical signs of delirium in the psychiatric inpatient population. Evaluation of these functions can substantially impact diagnostic decisions where diagnostic uncertainty exists. An evaluation of writing and constructional praxis can be easily incorporated into bedside mental status examinations. PMID- 10696494 TI - A survey of the use of community treatment orders by psychiatrists in Saskatchewan. AB - OBJECTIVE: To determine the pattern of use and satisfaction with community treatment orders (CTOs) by psychiatrists in Saskatchewan. METHOD: All psychiatrists who were licensed to practise by the College of Physicians and Surgeons of Saskatchewan were surveyed by mail in July 1998. RESULTS: The response rate was 72%. The responding psychiatrists were treating 14 patients on CTOs at the time of the survey. Psychiatrists were generally satisfied with the operation of CTOs, though many felt that commitment of only of 3 months before mandatory renewal was too short a period. Almost one-half expected their use of treatment orders to increase. CONCLUSION: While CTOs are used for only a small number of patients in Saskatchewan, they are a clinically useful tool for dealing with a group of otherwise difficult-to-treat patients. PMID- 10696495 TI - Gabapentin treatment for posttraumatic stress disorder. PMID- 10696496 TI - Fatigue and depression in multiple sclerosis. PMID- 10696497 TI - Impact of sildenafil on male erectile disorder due to psychological factors. PMID- 10696498 TI - A home treatment team for individuals with acute mental illnesses. PMID- 10696499 TI - Depression induced by orlistat (Xenical) PMID- 10696500 TI - Electroconvulsive therapy and anticoagulation. PMID- 10696501 TI - Re: The meeting of pain and depression: comorbidity in women. PMID- 10696502 TI - Developing a patient care monitoring system. PMID- 10696503 TI - Increased aggressive, violent, and impulsive behaviour in patients during chronic prolonged benzodiazepine use. PMID- 10696504 TI - Psychostimulant clinical response in fetal alcohol syndrome. PMID- 10696505 TI - Permeable endothelium and the interstitial space of brain. AB - 1. Fenestrated vessels can be reversibly induced in brain by agents that stimulate urokinase production. This plasminogen activator, like vascular endothelial growth factor and metalloproteinases, is secreted by tumor cells and may account for induction of fenestrated vessels. Why only some of the brain's barrier vessels are converted to fenestrated vessels is unknown. 2. The structures responsible for the filtering of solutes by fenestrated vessels may be the same as those of continuous, less permeable vessels: the glycocalyx on the surfaces of the endothelial cells and the subendothelial basal lamina. 3. Solutes leaving the cerebral ventricles immediately enter the interstitial clefts between the cells lining the ventricles. A fraction of a variety of solutes, injected into CSF compartments, is retained by subendothelial basal lamina, from which the solutes may be released in a regulated way. 4. The brain's CSF and interstitial clefts are the conduits for nonsynaptic volume transmission of diffusible signals, e.g., ions, neurotransmitters, and hormones. This type of transmission could be abetted by a parallel, cell-to-cell volume transmission mediated by gap junctions between astrocytes bordering CSF compartments and parenchymal astrocytes. 5. The width and contents of the interstitial clefts in fetal brain permit cell migration and outgrowth of neurites. The contents of the narrower and different interstitial clefts of mature brain permit solute convection but must be enzymatically degraded in order for cells to migrate through it. PMID- 10696506 TI - Inflammatory mediators and modulation of blood-brain barrier permeability. AB - 1. Unlike some interfaces between the blood and the nervous system (e.g., nerve perineurium), the brain endothelium forming the blood-brain barrier can be modulated by a range of inflammatory mediators. The mechanisms underlying this modulation are reviewed, and the implications for therapy of the brain discussed. 2. Methods for measuring blood-brain barrier permeability in situ include the use of radiolabeled tracers in parenchymal vessels and measurements of transendothelial resistance and rate of loss of fluorescent dye in single pial microvessels. In vitro studies on culture models provide details of the signal transduction mechanisms involved. 3. Routes for penetration of polar solutes across the brain endothelium include the paracellular tight junctional pathway (usually very tight) and vesicular mechanisms. Inflammatory mediators have been reported to influence both pathways, but the clearest evidence is for modulation of tight junctions. 4. In addition to the brain endothelium, cell types involved in inflammatory reactions include several closely associated cells including pericytes, astrocytes, smooth muscle, microglia, mast cells, and neurons. In situ it is often difficult to identify the site of action of a vasoactive agent. In vitro models of brain endothelium are experimentally simpler but may also lack important features generated in situ by cell:cell interaction (e.g. induction, signaling). 5. Many inflammatory agents increase both endothelial permeability and vessel diameter, together contributing to significant leak across the blood brain barrier and cerebral edema. This review concentrates on changes in endothelial permeability by focusing on studies in which changes in vessel diameter are minimized. 6. Bradykinin (Bk) increases blood-brain barrier permeability by acting on B2 receptors. The downstream events reported include elevation of [Ca2+]i, activation of phospholipase A2, release of arachidonic acid, and production of free radicals, with evidence that IL-1 beta potentiates the actions of Bk in ischemia. 7. Serotonin (5HT) has been reported to increase blood-brain barrier permeability in some but not all studies. Where barrier opening was seen, there was evidence for activation of 5-HT2 receptors and a calcium-dependent permeability increase. 8. Histamine is one of the few central nervous system neurotransmitters found to cause consistent blood-brain barrier opening. The earlier literature was unclear, but studies of pial vessels and cultured endothelium reveal increased permeability mediated by H2 receptors and elevation of [Ca2+]i and an H1 receptor-mediated reduction in permeability coupled to an elevation of cAMP. 9. Brain endothelial cells express nucleotide receptors for ATP, UTP, and ADP, with activation causing increased blood-brain barrier permeability. The effects are mediated predominantly via a P2U (P2Y2) G protein-coupled receptor causing an elevation of [Ca2+]i; a P2Y1 receptor acting via inhibition of adenyl cyclase has been reported in some in vitro preparations. 10. Arachidonic acid is elevated in some neural pathologies and causes gross opening of the blood-brain barrier to large molecules including proteins. There is evidence that arachidonic acid acts via generation of free radicals in the course of its metabolism by cyclooxygenase and lipoxygenase pathways. 11. The mechanisms described reveal a range of interrelated pathways by which influences from the brain side or the blood side can modulate blood-brain barrier permeability. Knowledge of the mechanisms is already being exploited for deliberate opening of the blood-brain barrier for drug delivery to the brain, and the pathways capable of reducing permeability hold promise for therapeutic treatment of inflammation and cerebral edema. PMID- 10696507 TI - Endothelial vesicles in the blood-brain barrier: are they related to permeability? AB - 1. Macromolecules cross capillary walls via large vascular pores that are thought to be formed by plasmalemmal vesicles. Early hypotheses suggested that vesicles transferred plasma constituents across the endothelial wall either by a "shuttle" mechanism or by fusing to form transient patent channels for diffusion. Recent evidence shows that the transcytotic pathway involves both movement of vesicles within the cell and a series of fusions and fissions of the vesicular and cellular membranes. 2. The transfer of macromolecules across the capillary wall is highly specific and is mediated by receptors incorporated into specific membrane domains. Therefore, despite their morphological similarity, endothelial vesicles from heterogeneous populations in which the predominant receptor proteins incorporated in their membranes define the functions of individual vesicles. 3. Blood-brain barrier capillaries have very low permeabilities to most hydrophilic molecules. Their low permeability to macromolecules has been presumed to be due to an inhibition of the transcytotic mechanism, resulting in a low density of endothelial vesicles. 4. A comparison of vesicular densities and protein permeabilities in a number of vascular beds shows only a very weak correlation, therefore vesicle numbers alone cannot be used to predict permeability to macromolecules. 5. Blood-brain barrier capillaries are fully capable of transcytosing specific proteins, for example, insulin and transferrin, although the details are still somewhat controversial. 6. It has recently been shown that the albumin binding protein gp60 (also known as albondin), which facilitates the transcytosis of native albumin in other vascular beds, is virtually absent in brain capillaries. 7. It seems likely that the low blood brain barrier permeability to macromolecules may be due to a low level of expression of specific receptors, rather than to an inhibition of the transcytosis mechanism. PMID- 10696508 TI - Brain microvascular P-glycoprotein and a revised model of multidrug resistance in brain. AB - 1. P-Glycoprotein is a 170-kDa transmembrane glycoprotein active efflux system that confers multidrug resistance in tumors, as well as normal tissues including brain. 2. The classical model of multidrug resistance in brain places the expression of P-glycoprotein at the luminal membrane of the brain microvascular endothelial cell. However, recent studies have been performed with human brain microvessels and double-labeling confocal microscopy using (a) the MRK16 antibody to human P-glycoprotein, (b) an antiserum to glial fibrillary acidic protein (GFAP), an astrocyte foot process marker, or (c) an antiserum to the GLUT1 glucose transporter, a brain endothelial plasma membrane marker. These results provide evidence for a revised model of P-glycoprotein function at the brain microvasculature. In human brain capillaries, there is colocalization of immunoreactive P-glycoprotein with astrocytic GFAP but not with endothelial GLUT1 glucose transporter. 3. In the revised model of multidrug resistance in brain, P glycoprotein is hypothesized to function at the plasma membrane of astrocyte foot processes. These astrocyte foot processes invest the brain microvascular endothelium but are located behind the blood-brain barrier in vivo, which is formed by the brain capillary endothelial plasma membrane. 4. In the classical model, an inhibition of endothelial P-glycoprotein would result in both an increase in the blood-brain barrier permeability to a given drug substrate of P glycoprotein and an increase in the brain volume of distribution (VD) of the drug. However, in the revised model of P-glycoprotein function in brain, which positions this protein transporter at the astrocyte foot process, an inhibition of P-glycoprotein would result in no increase in blood-brain barrier permeability, per se, but only an increase in the VD in brain of P-glycoprotein substrates. PMID- 10696509 TI - The choroid plexuses and the barriers between the blood and the cerebrospinal fluid. AB - 1. The fluid homeostasis of the brain depends both on the endothelial blood-brain barrier and on the epithelial blood-cerebrospinal fluid (CSF) barrier located at the choroid plexuses and the outer arachnoid membrane. 2. The brain has two fluid environments: the brain interstitial fluid, which surrounds the neurons and glia, and the CSF, which fills the ventricles and external surfaces of the central nervous system. 3. CSF acts as a fluid cushion for the brain and as a drainage route for the waste products of cerebral metabolism. 4. Recent findings suggest that CSF may also act as a "third circulation" conveying substances secreted into the CSF rapidly to many brain regions. PMID- 10696510 TI - Choroid plexus recovery after transient forebrain ischemia: role of growth factors and other repair mechanisms. AB - 1. Transient forebrain ischemia in adult rats, induced by 10 min of bilateral carotid occlusion and an arterial hypotension of 40 mmHg, caused substantial damage not only to CA-1 neurons in hippocampus but also to epithelial cells in lateral ventricle choroid plexus. 2. When transient forebrain ischemia was followed by reperfusion (recovery) intervals of 0 to 12 hr, there was moderate to severe damage to many frond regions of the choroidal epithelium. In some areas, epithelial debris was sloughed into cerebrospinal fluid (CSF). Although some epithelial cells were disrupted and necrotic, their neighbors exhibited normal morphology. This patchy response to ischemia was probably due to regional differences in reperfusion or cellular metabolism. 3. Between 12 and 24 hr postischemia, there was marked restoration of the Na+, K+, water content, and ultrastructure of the choroid plexus epithelium. Since there was no microscopical evidence for mitosis, we postulate that healthy epithelial cells either were compressed together on the villus or migrated from the choroid plexus stalk to more distal regions, in order to "fill in gaps" along the basal lamina caused by necrotic epithelial cell disintegration. 4. Epithelial cells of mammalian choroid plexus synthesize and secrete many growth factors and other peptides that are of trophic benefit following injury to regions of the cerebroventricular system. For example, several growth factors are upregulated in choroid plexus after ischemic and traumatic insults to the central nervous system. 5. The presence of numerous types of growth factor receptors in choroid plexus allows growth factor mediation of recovery processes by autocrine and paracrine mechanisms. 6. The capability of choroid plexus after acute ischemia to recover its barrier and CSF formation functions is an important factor in stabilizing brain fluid balance. 7. Moreover, growth factors secreted by choroid plexus into CSF are distributed by diffusion and convection into brain tissue near the ventricular system, e.g., hippocampus. By this endocrine-like mechanism, growth factors are conveyed throughout the choroid plexus-CSF-brain nexus and can consequently promote repair of ischemia damaged tissue in the ventricular wall and underlying brain. PMID- 10696511 TI - Osmotic opening of the blood-brain barrier: principles, mechanism, and therapeutic applications. AB - 1. Osmotic opening of the blood-brain barrier by intracarotid infusion of a hypertonic arabinose or mannitol solution is mediated by vasodilatation and shrinkage of cerebrovascular endothelial cells, with widening of the interendothelial tight junctions to an estimated radius of 200 A. The effect may be facilitated by calcium-mediated contraction of the endothelial cytoskeleton. 2. The marked increase in apparent blood-brain barrier permeability to intravascular substances (10-fold for small molecules) following the osmotic procedure is due to both increased diffusion and bulk fluid flow across the tight junctions. The permeability effect is largely reversed within 10 min. 3. In experimental animals, the osmotic method has been used to grant wide access to the brain of water-soluble drugs, peptides, antibodies, boron compounds for neutron capture therapy, and viral vectors for gene therapy. The method also has been used together with anticancer drugs to treat patients with metastatic or primary brain tumors, with some success and minimal morbidity. PMID- 10696513 TI - Inequality and children's health. PMID- 10696514 TI - Risk factors for failure to thrive: a population-based survey. AB - AIM: To identify whether differences exist between failure to thrive children and controls in either demographic characteristics or parental rating of their eating and other behaviour. METHODS: As part of an intervention study, 97 children with failure to thrive were identified by population screening and received a standardized assessment by their health visitor at a median age of 15.1 months. This included standard questions to parents concerning their perception of their child's feeding history and behaviour. Their responses were compared with the parents of 28 normally growing children aged 16-18 months, systematically sampled from the same district. RESULTS: Cases had fallen through a mean of 1.69 weight standard deviation score and were markedly underweight for height. The case families had similar levels of deprivation, both to controls and city norms, and only four showed evidence of major neglect. Failure to thrive children had significantly more infancy feeding problems and were introduced to solids and finger foods later than controls; they were significantly more often described as variable eaters, undemanding and shy and less often as hungry. Cases liked most foods, but significantly less so than controls. CONCLUSIONS: This suggests that the role of deprivation and neglect has been overstated and that undemanding behaviour, low appetite and poor feeding skills may contribute to the onset and persistence of failure to thrive. PMID- 10696512 TI - Determinants of passive drug entry into the central nervous system. AB - 1. The blood-brain barriers restrict the passive diffusion of many drugs into the brain and constitute a significant obstacle in the pharmacological treatment of central nervous system diseases and disorders. The degree of restriction they impose is variable, with some lipid-insoluble drugs effectively excluded from the brain, while many lipid-soluble drugs do not appear to be subject to any restriction. 2. The ease with which any particular drug diffuses across the blood brain barrier is determined largely by the number and strength of intermolecular forces "holding" it to surrounding water molecules. By quantifying the molecular features that contribute to these forces, it is possible to predict the in vivo blood-brain barrier permeability of a drug from its molecular structure. Dipolarity, polarizability, and hydrogen bonding ability are factors that appear to reduce permeability, whereas molecular volume (size) and molar refraction are associated with increased permeability. 3. Increasing the passive entry of "restricted" drugs into the central nervous system can be achieved by disrupting the blood-brain barrier (increased paracellular diffusion) or by modifying the structure of "restricted" drugs to temporarily or permanently increase their lipid solubility (increased transcellular permeability). 4. Competitive inhibition of outwardly directed active efflux mechanisms (P-glycoprotein and MRP, the multidrug resistance-related protein) can also significantly increase the accumulation of certain drugs within the central nervous system. PMID- 10696515 TI - Breast-feeding and weight change in newborns in Jamaica. AB - This study was conducted to examine weight change of exclusively breast-fed infants during the first week and through the first 24 days of life, and to evaluate the effect of breast-feeding factors and maternal characteristics on early weight change in the infants. The weights of 21 infants were recorded on day 1 (day of birth), and on days 3, 7, 10, 17, and 24, and the data analysed to evaluate weight change over the period. Multiple regression analysis was used to assess whether birth weight as well as maternal and breast-feeding factors were significant predictors of weight on day 24. Nineteen of the 21 infants gained weight between days 1 and 3, and 20 infants gained weight between days 3 and 7. All infants gained weight over the 24-day period and their weights at day 7 and day 24 were significantly different (P < 0.05 and P < 0.01, respectively) from their birth weights. Multiple linear regression analysis showed that significant (P < 0.01) predictors of weight gain by day 24 included birth weight, mother's educational level, whether the baby cried before feeding, and length of feeding time periods. This is the first study of weight change in the early days and weeks of life of exclusively breast-fed newborn infants in Jamaica. The infants showed significant weight gain during the study period and weight gain was affected by certain maternal and breast-feeding factors. PMID- 10696516 TI - Disclosure of diagnosis and planning for the future in HIV-affected families in Europe. AB - Information relating to disclosure of infection status in families affected by HIV and the existence of plans for the future social care of children with infected parents was collected as part of a larger survey on clinical and psychosocial service use of these families. Parents and alternative carers of HIV affected children in follow-up in 10 paediatric centres from seven European countries were surveyed. A total of 182 questionnaires were returned: most (73%) were completed by parents, of whom 92% were HIV-infected. Of the 226 children cared for by the respondents, most (62%) were HIV-infected. Disclosure of both the child's and the parent's infection status was rare and found to be associated with child's age in both cases. Infected children living with their parents were less likely to know their diagnosis than those living in alternative care. Uninfected parents and carers were significantly more likely to want professional help with disclosing to an infected child than infected parents. Infected parents also face difficult decisions regarding the issue of who will care for their children when they are unable to. Half of the infected parents had made long-term plans for their children's future social care. European parents were more likely to have made such plans than those from elsewhere (mainly Africa) and parents with plans had known about their HIV infection for significantly longer than those without. Increasing numbers of vertically infected children are reaching adolescence as a result of improvements in the management of paediatric HIV infection. As both disclosure and planning for the future social care of HIV affected children have been found to be strongly associated with child's age, the changing epidemiology of paediatric HIV highlights the need for more information on these issues in order to support families more effectively. PMID- 10696517 TI - Drug information leaflets: adolescent and professional perspectives. AB - This study was undertaken to evaluate a range of drug information leaflets from both an adolescent and professional perspective. 72 adolescents were interviewed in focus groups within school settings. Fifteen paediatricians were also interviewed individually. Participants were asked to consider the leaflets under the following categories: initial impact, content and overall suitability for adolescents. Paediatricians were not good judges of which leaflets would appeal to adolescents. They tended to choose the leaflets with comic strip illustrations and those written in a 'cool' or witty style. In contrast the adolescents seemed to prefer a more sober approach with high quality factual information combined with clear illustrations. PMID- 10696518 TI - Physical and personality traits of preschool children in Fuzhou, China: only child vs sibling. AB - Concern about the healthy growth and development of an only child has been voiced since the 1970s, when the Chinese government launched the only child policy. In this study, the physical and personality traits of rural Chinese preschool only children (onlies) whose age ranged from 3 to 6 years old were evaluated. The sample included 197 onlies and 367 children with siblings who came from seven kindergartens in rural areas in Fuzhou, Fujian province. The results showed no statistically significant differences in height, mass or BMI between the onlies and siblings. Regarding the personality traits, the significant difference was that the onlies exhibited more somatic complaints, however, the data didn't indicate any other undesirable personality traits for the onlies. These results suggest that Chinese preschool children grow normally with or without siblings. PMID- 10696519 TI - Parental non-compliance--a paediatric dilemma. A medical and psychodynamic perspective. AB - Considerable resources have been directed towards the recognition and management of child physical and sexual abuse and/or neglect. However, the issue of parental non-compliance is less well defined and under recognized. While outwardly seeking advice, non-compliant parents, especially if anxious, are unable or unwilling to comply with the recommendations made. Conflicts of interest between the parent(s)' and the health professionals' perceptions regarding the best interest of the child may arise. Parental non-compliance is centred around the parents' perception of the child's current problems and its relationship to past problems. Such non-compliance may reflect ignorance or misunderstanding of the clinical situation. Ignorance may be readily addressed if the parents are receptive and trusting. However, non-compliance more commonly arises from the parents' inability to cope emotionally with the stresses surrounding the recommended treatment. Parents may be vulnerable to psychological reactions which inhibit rational thinking. Parental anxieties are best understood in terms of psychological constructs, including 'defences' such as 'denial' and 'splitting', 'repetition compulsion' and the need to 'work through' psychological barriers so that the child's best interest is served. Parental non-compliance can serve to protect the parents from overwhelming fears and anxieties, which if addressed may transform parental defensiveness to co-operation. Extreme parental non-compliance may represent a special form of child abuse where, due to parental psychopathology, parents are unable to consider the child's best interest. Clinical vignettes arising from a consultant private and hospital ambulatory setting will focus on management strategies for successful outcomes. Recommendations offered on ways to reduce the risk of parental non-compliance include building trust, eliciting the aid of a parental partner, and organizing a second opinion, thereby improving the chances of a successful outcome. PMID- 10696520 TI - Cause-specific perinatal death rates, birth weight and deprivation in the West Midlands, 1991-93. AB - OBJECTIVES: To study the relationship between cause-specific perinatal death rates, material deprivation and birthweight among births in 3 consecutive years in the West Midlands Health Region. STUDY DESIGN: Retrospective cohort study. SETTING: West Midlands Health Region (WMHR). STUDY POPULATION: All births (live and stillbirths) to mothers with addresses in the WMHR in 1991, 1992 and 1993. MAIN OUTCOME MEASURES: Cause-specific perinatal death rates--crude and stratified by birthweight. METHODS: Perinatal deaths in the WMHR in 1991-93, collected as part of the national Confidential Enquiry into Stillbirths and Deaths in Infancy, were classified into causes of death by the extended Wigglesworth classification. Crude rates for cause-specific perinatal deaths and rates stratified by birthweight < 2500 g and > or = 2500 g were calculated for each enumeration district (ED) quintile derived by ranking enumeration districts for the whole of the region by Townsend Deprivation Index. Cause-specific rates of death were investigated for a linear trend across ED quintiles. The relative risk of death (most vs least deprived) from specific causes was calculated. Using rates for the least deprived quintile as the reference, deaths from each cause 'attributable' to social inequality were calculated. RESULTS: Positive linear trends in perinatal deaths were noted with increasing deprivation for each specific cause of death except those classified as 'other causes' (Wigglesworth Class E). Relative risk (most vs least deprived) of perinatal death with a congenital anomaly was 1.98 (confidence interval, CI: 1.36,2.89). For deaths related to antepartum events, intrapartum events and immaturity the risks were 1.81 (CI: 1.39,2.38), 1.48 (CI: 1.10,1.98) and 1.92 (CI 1.45,2.56), respectively. Forty three (35.7%) perinatal deaths per year were due to congenital anomalies, 63 (29.7%) antepartum events, 36 (21.9%) intrapartum events and 61 (32.7%) immaturity and these were statistically 'attributable' to social inequality. Cause-specific perinatal death rates for babies < 2500 g showed no correlation with deprivation; however, for babies > or = 2500 g the association with deprivation persisted. CONCLUSIONS: All cause-specific rates except those due to 'other causes' showed a positive linear trend with increasing deprivation. These trends were found for infants born > or = 2500 g but were not seen for low birthweight infants (< 2500 g). Almost 30% of deaths were statistically 'attributable' to social inequality. The results of this study suggest that material deprivation plays an important role in the causal pathway leading to perinatal death and needs to be addressed in preventive programmes aimed at the reduction of perinatal deaths. PMID- 10696521 TI - Alzheimer's disease and inflammation: a review of cellular and therapeutic mechanisms. AB - 1. Of the neurodegenerative diseases that cause dementia, Alzheimer's disease (AD) is the most common. Three major pathologies characterize the disease: senile plaques, neurofibrillary tangles and inflammation. We review the literature on events contributing to the inflammation and the treatments thought to target this pathology. 2. The senile plaques of AD consist primarily of complexes of the beta amyloid protein. This protein is central to the pathogenesis of the disease. 3. Inflammatory microglia are consistently associated with senile plaques in AD, although the classic inflammatory response (immunoglobulin and leucocyte infiltration) is absent. beta-Amyloid fragments appear to mediate such inflammatory mechanisms by activating the complement pathway in a similar fashion to immunoglobulin. 4. Epidemiological studies have identified a reduced risk of AD in patients with arthritis and in leprosy patients treated with anti inflammatory drugs. Longitudinal studies have shown that the consumption of anti inflammatory medications reduces the risk of AD only in younger patients (< 75 years). 5. There is a considerable body of in vitro evidence indicating that the inflammatory response of microglial cells is reduced by non-steroidal anti inflammatory drugs (NSAID). However, no published data are available concerning the effects of these medications on brain pathology in AD. 6. Cyclo-oxygenase 2 enzyme is constitutively expressed in neurons and is up-regulated in degenerative brain regions in AD. Non-steroidal anti-inflammatory drugs may reduce this expression. 7. Platelets are a source of beta-amyloid and increased platelet activation and increased circulating beta-amyloid have been identified in AD. Anti-platelet medication (including NSAID) would prevent such activation and its potentially harmful consequences. 8. Increased levels of luminal beta-amyloid permeabilizes the blood-brain barrier (BBB) and increases vasoconstriction of arterial vessels, paralleling the alterations observed with infection and inflammation. Cerebral amyloidosis is highly prevalent in AD, compromising the BBB and vasoactivity. Anti-inflammatory medications may alleviate these problems. PMID- 10696522 TI - Viral-mediated gene delivery of constitutively activated G alpha s alters vasoreactivity. AB - 1. Decline in beta-adrenoceptor (beta-AR)-mediated function occurs with increasing age, as well as in multiple disease conditions. The mechanisms responsible for this decline include alterations in beta-AR itself, beta-AR coupling proteins, such as G-proteins, or other beta-AR-linked proteins, such as G-protein receptor kinases and/or phosphatases. 2. The present study examines the physiological effects of in vitro transfer of constitutively activated G alpha s (G alpha s-Q227L) to both cultured vascular smooth muscle cells (VSMC) and whole aortic tissue of 6-month-old (adult) animals via a replication-deficient Herpes simplex virus (HSV) vector. These studies were conducted to provide a model for future examination of the role of G alpha s in the age-related decline in beta-AR mediated vasorelaxation. 3. Gene transfer was confirmed by western blotting for specific proteins. Aortic tissue infected with HSV-G alpha s-Q227L had reduced phenylephrine-induced contraction and enhanced isoproterenol-stimulated vasorelaxation. Infection of cultured VSMC with HSV-G alpha s-Q227L increased both basal- and isoproterenol-stimulated cAMP accumulation, whereas forskolin stimulated cAMP production was unchanged. 4. These results implicate G alpha s as a target for further investigation in age-related changes in vascular reactivity and support the use of viral-mediated gene transfer as an effective tool to study adrenergic signal transduction and physiology in vascular tissue. PMID- 10696523 TI - Troponin T in the coronary sinus and percutaneous transluminal coronary angioplasty related myocardial injury. AB - 1. Myocardial injury has been shown to be associated with successful percutaneous transluminal coronary angioplasty (PTCA). The present study was designed to determine whether uncomplicated successful PTCA results in myocardial injury by measuring coronary sinus (CS) cardiac troponin T (cTnT). 2. We measured cTnT in the CS and the femoral vein (FV) in 16 patients with stable angina pectoris who underwent uncomplicated PTCA for stenotic lesions of the left anterior descending artery. Blood samples were drawn from both the CS and FV before and immediately after PTCA and every 4 h for the next 12 h. 3. All patients had chest pain and electrocardiographic ST segment elevation or depression during balloon inflation and higher peak elevation of cTnT in the CS than in the FV (0.054 +/- 0.059 vs 0.036 +/- 0.022 ng/mL; P < 0.05). However, all CS cTnT levels were within the normal range over the 12 h period. 4. The fact that CS cTnT measurements showed no evidence of uncomplicated PTCA-related myocardial injury led us to conclude that uncomplicated successful PTCA does not cause myocardial injury. PMID- 10696524 TI - Sodium sensitivity and sympathetic nervous system in hypertension induced by long term nitric oxide blockade in rats. AB - 1. Pharmacological inhibition of nitric oxide (NO) synthesis is known to produce acute and chronic hypertension in many animal species, but the underlying mechanisms mediating the hypertension are not completely understood. In particular, the pathogenetic roles of sodium sensitivity and the sympathetic nervous system in this model of hypertension are controversial. The present study was designed to test the hypothesis that long-term administration of the NO synthesis inhibitor NG-nitro-L-arginine methyl ester (L-NAME) to male Sprague Dawley rats would produce a sodium-sensitive hypertension and that the enhanced activity of the sympathetic nervous system in this type of hypertension contributes to the sodium sensitivity. 2. NG-Nitro-L-arginine methyl ester was added to drinking fluid for 8 weeks at a concentration of 16 mg/dL. Rats received tap water for the first 4 weeks of the study and were then divided into two groups and placed on either a normal or high sodium intake (ingestion of either tap water or 0.9% NaCl, respectively). Awake systolic blood pressure was measured by the tail-cuff method every week. Urinary excretion rates of the stable NO metabolites and catecholamines during NO synthesis inhibition were examined. 3. Long-term administration of L-NAME produced a marked and sustained elevation in arterial pressure without altering urine flow, or sodium excretion rate. NG-Nitro L-arginine methyl ester-induced hypertension was accompanied by a decreased urinary excretion of the stable NO metabolites NO2- and NO3- and was aggravated when rats drank 0.9% NaCl in place of tap water. Urinary excretion of adrenaline and noradrenaline, but not dopamine, in L-NAME-treated rats increased significantly within the first week of the study compared with control rats. L Arginine (2 g/dL in drinking fluid) completely reversed the elevation of arterial pressure as well as the decrease in urinary NO2- and NO3- excretion and the increased urinary excretion of catecholamines associated with L-NAME treatment by 3 weeks of concomitant administration. 4. These results suggest that long-term inhibition of NO synthesis produces a sodium-sensitive hypertension and that changes in sympathetic nerve activity may, at least in part, contribute to the sodium sensitivity in this type of hypertension. PMID- 10696525 TI - Long-term oestrogen therapy is associated with improved endothelium-dependent vasodilation in the forearm resistance circulation of biological males. AB - 1. The aim of the present study was to determine the effects of long-term oestrogen on resistance vessel reactivity in biological males. 2. Recent studies have demonstrated that long-term oestrogen therapy favourably alters the lipid profile and improves vasodilator function in the conduit arteries of biological males. Whether a similar benefit is exerted on the resistance circulation is not known. Therefore, we examined the effects of long-term oestrogen therapy on skeletal muscle resistance vessel function in biological males and the potential mechanisms by which it may exert its effects. 3. Forearm blood flow (FBF) and resistance were compared in 15 male-to-female transsexuals being prescribed oestrogen, with 14 age-matched healthy males, at rest and in response to the endothelium-dependent nitric oxide (NO) vasodilator acetylcholine (ACh), the endothelium-independent but NO-mediated vasodilator sodium nitroprusside (SNP), the endothelium-independent and non-NO-mediated vasodilator verapamil (VER) and the endothelium-independent vasoconstrictor phenylephrine (PE). 4. Basal blood flows were similar in the two groups. However, the male-to-female transsexuals had a significant upward and leftward shift in FBF responses to ACh compared with males, with a 52% increase in FBF responses at the highest dose of ACh used. Forearm blood flow in transsexuals rose from a mean (+/- SEM) baseline level of 3.02 +/- 0.25 to a maximum of 19.5 +/- 2.59 mL/min per 100 mL forearm tissue (compared with 3.24 +/- 0.41 and 9.43 +/- 1.97 mL/min per 100 mL forearm tissue, respectively, in males) with the highest dose of ACh (+2.73 micrograms/min per 100 mL; P < 0.0005). Forearm vascular resistance was also significantly reduced in transsexuals compared with males (P < 0.05). Vasodilator responses to SNP, VER and PE were similar in both groups. 5. There were no differences observed in total cholesterol and low-density lipoprotein-cholesterol levels. However, male to-female transsexuals had 20% higher high-density lipoprotein-cholesterol levels compared with males (1.57 +/- 0.11 vs 1.26 +/- 0.08 mmol/L, respectively; P < 0.05) and 47% higher triglyceride levels (P < 0.005). Serum testosterone levels (an index of oestrogen therapy) was a predictor of responses to endothelium dependent vasodilation (rs = -0.50; P < 0.01). 6. Long-term oestrogen therapy enhances endothelium-dependent vasodilation in the skeletal muscle microcirculation of biological males. The effects appear to be selective because endothelium-independent vasodilation and vasoconstriction are not altered. PMID- 10696526 TI - Contribution of nitric oxide, cyclic GMP and K+ channels to acetylcholine-induced dilatation of rat conduit and resistance arteries. AB - 1. We compared the effects of inhibiting nitric oxide synthase (NOS), soluble guanylate cyclase (sGC) and K+ channel activation on dilator responses to acetylcholine (ACh) in rat resistance (hindquarters) and conduit arteries (thoracic aorta). 2. In rat perfused hindquarters, the NO synthase inhibitor N omega-nitro-L-arginine (L-NNA; 1 mmol/L) partially inhibited the ACh-induced dilatation and the combination of L-NNA + haemoglobin (Hb; 20 mumol/L), a NO scavenger, did not further affect the response. Exposure to high K+ (30 mmol/L) also inhibited the response to ACh and this response was further reduced by L-NNA + high K+. Surprisingly, when applied alone 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin 1-one (ODQ), an inhibitor of sGC, did not affect responses to ACh, whereas treatment with ODQ + high K+ markedly impaired dilatation. 3. In aortic rings precontracted with phenylephrine (PE; 0.01-1 mumol/L), the maximum relaxation to ACh was significantly reduced by L-NNA (0.1 mmol/L) and further inhibited by L NNA + Hb (20 mumol/L). At 10 mumol/L, ODQ alone inhibited the maximum relaxation to ACh, which was further reduced by ODQ + high K+ (30 mmol/L). High K+ caused a smaller but significant inhibition of ACh-induced relaxation. 4. These results suggest that NO and cGMP play a relatively greater role in ACh-induced dilatation of the aorta compared with the hindquarters resistance vasculature and are consistent with the hypothesis that a non-NO endothelium-derived hyperpolarizing factor (endothelium-derived hyperpolarizing factor; EDHF) makes a relatively greater contribution to dilatation of resistance vessels than in conduit arteries. The data suggest that when sGC is inhibited, a compensatory mechanism involving K+ channel opening by NO can largely maintain ACh-induced vasodilator responses of resistance vessels. Furthermore, when NO synthesis is blocked, a non NO EDHF may play a role in ACh-induced dilatation of the resistance vasculature. PMID- 10696527 TI - Soluble dietary fibre improves insulin sensitivity by increasing muscle GLUT-4 content in stroke-prone spontaneously hypertensive rats. AB - 1. The effects of soluble dietary fibre (psyllium) on peripheral insulin sensitivity and skeletal muscle GLUT-4 protein expression were studied in 12 male stroke-prone spontaneously hypertensive rats (SHRSP) fed a high-caloric diet from 5 to 9 weeks of age. 2. In the psyllium-supplemented group, fasting plasma glucose was significantly reduced and glucose levels following an oral glucose tolerance test were significantly lower than in the cellulose-supplemented group at 30 (P < 0.05) and 60 min (P < 0.01). However, there was no difference in insulin secretion. 3. In the psyllium-supplemented group, skeletal muscle GLUT-4 content was significantly increased in the plasma membrane (P < 0.001), but not in the intracellular membrane. 4. No significant difference was found in phosphatidylinositol 3 (PI3)-kinase activity between cellulose and psyllium diet not only in the basal state but also when stimulated by insulin. 5. These results demonstrate that psyllium increases blood glucose disposal by increasing skeletal muscle plasma membrane GLUT-4 content without PI3-kinase activation. PMID- 10696528 TI - Inhibitory action of ambocarb on voltage-operated sodium channels in rat isolated hippocampal pyramidal neurons. AB - 1. A whole-cell patch-clamp study of the effects of ambocarb, a novel nootropic beta-carboline, on sodium currents in rat acutely isolated hippocampal pyramidal neurons was performed. 2. Ambocarb potently and reversibly suppressed sodium currents in a concentration range of 3-300 mumol/L. The amount of block was dependent on the holding potential, with half-maximal inhibition values being 26 and 94 mumol/L at -80 and -120 mV, respectively. 3. Ambocarb induced a hyperpolarizing shift in the steady state availability curve, which indicates an increase in the proportion of inactivated sodium channels. This action is presumably mediated by promoting the development of inactivation and slowing the recovery of sodium channels from inactivation. 4. Because many neuroprotective drugs were shown to inhibit sodium currents, down-modulation of voltage-operated sodium channels that complements the known positive interaction of ambocarb and other related beta-carbolines with GABAA receptors may provide a promising strategy in the treatment of brain disorders associated with trauma and ischaemia. PMID- 10696529 TI - Effect of chronic lithium administration on endothelium-dependent relaxation in rat aorta. AB - 1. The effects of chronic lithium administration on the relaxant responses of rat thoracic aortic rings in the presence of indomethacin (a cyclo-oxygenase inhibitor) and/or NG-nitro-L-arginine (L-NOARG; a nitric oxide synthase inhibitor) to acetylcholine (ACh) or sodium nitroprusside were investigated in the present study. 2. Acetylcholine produced a concentration-dependent relaxation in vessels precontracted by phenylephrine (PE), while in lithium-treated rats the maximal relaxation was significantly increased. 3. Indomethacin (20 mumol/L) significantly potentiated the ACh-induced relaxation in lithium-treated and control rats. 4. NG-Nitro-L-arginine (1 mumol/L) decreased the ACh-induced relaxation in both control and lithium-treated rats. In contrast, indomethacin (20 mumol/L) reversed the inhibitory effect of L-NOARG. 5. Sodium nitroprusside produced similar concentration-dependent relaxations of vessels from both control and lithium-treated rats, which was not affected by indomethacin. In endothelium denuded rings, indomethacin (20 mumol/L) caused a rightward shift in the concentration-contraction curve to PE. 6. These data support evidence for a possible increase in endothelium-dependent relaxation induced by ACh during long term administration of lithium in rat aortic rings. PMID- 10696530 TI - Salt- and angiotensin II-dependent variations in amiloride-sensitive rectal potential difference in mice. AB - 1. In the rectum and distal nephron, sodium reabsorption is mediated by the amiloride-sensitive epithelial sodium channel (ENaC). The ENaC-mediated sodium transport is electrogenic and creates an amiloride-sensitive transepithelial potential difference (PD). 2. We have evaluated the salt- and angiotensin (Ang)II dependent variations in amiloride-sensitive rectal PD in mice and assessed their relationship with renal sodium handling. 3. Rectal PD was measured in vivo in mice maintained on a medium-, low- or high-sodium diet. On a medium-salt diet, the mean (+/- SEM) amiloride-sensitive PD was larger in the afternoon than in the morning (-26.1 +/- 0.9 and -11.2 +/- 0.7 mV, respectively; P = 0.001), indicating a circadian cyclicity. Rectal PD increased on a low-sodium diet and decreased on a high-sodium diet. 4. Amiloride-sensitive rectal PD correlated significantly with the urinary Na+/K+ ratio (P < 0.001) and with sodium reabsorption in the distal nephron as measured by the lithium clearance technique (P < 0.001). 5. In mice treated with an AngII AT1 receptor antagonist, amiloride-sensitive rectal PD was increased in the afternoon compared with controls (-32.8 +/- 2.0 vs -24.4 +/- 0.9, respectively; P < 0.001). 6. At high doses, AngII decreased the amiloride sensitive rectal PD and this effect was blunted by an AT1 receptor antagonist. 7. These results show the presence of a salt-dependent daily cyclicity of sodium transport in the mouse rectum that follows circadian changes in sodium handling in the distal nephron. Angiotensin II appears to modulate this diurnal pattern of rectal amiloride-sensitive sodium transport. PMID- 10696531 TI - Renal acid-base and sodium handling in hypoxia and subsequent mild metabolic acidosis in foetal sheep. AB - 1. To measure the renal contribution to acid-base homeostasis during hypoxia (not associated with hypercapnia) and in response to the subsequent mild metabolic acidosis and to determine the effects of this hypoxia on the renal handling of sodium, studies were performed in six chronically catheterized foetal sheep (129 138 days gestation) before, during and for 1 h after a 2 h period of hypoxia. 2. Hypoxia was induced in the conscious ewe by infusing nitrogen into the trachea. Foetal arterial oxygen tension fell to 12.0 +/- 0.6 mmHg (P < 0.001). Carbon dioxide tension fell during hypoxia (P < 0.001) and was still somewhat reduced in the recovery period (P < 0.005). Arterial pH fell progressively to 7.19 +/- 0.08 in the recovery period (P < 0.05). Plasma bicarbonate concentrations fell (P < 0.001) and lactate rose (P < 0.001). 3. Urinary pH and the excretion rates of bicarbonate, titratable acid, ammonium and net acid did not change during hypoxia. Ammonium excretion and, hence, generation of new bicarbonate increased in the recovery period (P < 0.05). 4. Renal sodium excretion progressively increased and was greatest after normoxia was restored (P < 0.05). This natriuresis was due to a fall in the reabsorption of sodium by the proximal tubule (P < 0.05). Proximal reabsorption of sodium was directly related to foetal pH (P < 0.0001) and bicarbonate reabsorption (P < 0.001). 5. It was concluded that: (i) the foetal kidneys began to contribute to the maintenance of acid-base balance within the first hour of recovery from a 2 h episode of hypocapnic hypoxia, even though the acidosis was relatively mild; and (ii) a reduction in bicarbonate reabsorption was probably the most important factor that limited sodium reabsorption by the renal tubule during this experiment. PMID- 10696532 TI - Effect of uranyl nitrate-induced renal failure on morphine disposition and antinociceptive response in rats. AB - 1. The aims of the present study were to administer morphine (14.0 mumol/kg, s.c.) to male Hooded Wistar rats and to determine the effect of uranyl nitrate induced renal failure on: (i) the antinociceptive effect of morphine; (ii) the pharmacokinetics of morphine and morphine-3-glucuronide (M3G); and (iii) the relationship between antinociceptive effect and the pharmacokinetics of morphine in plasma and brain. 2. Renal failure was induced by a single s.c. injection of uranyl nitrate and kinetic/dynamic studies were performed 10 days after its administration, when creatinine clearance was 17% of the control group. Antinociceptive effect was measured by the tail-flick method at various times up to 2 h post-drug administration. Concentrations of morphine and M3G in plasma and brain and concentrations of creatinine in urine and serum were determined by specific HPLC methods. 3. After morphine administration, the area under the antinociceptive effect-time curve was decreased by 44% in renal failure rats. There were no differences between control and renal failure rats in: (i) plasma morphine concentration-time curves; (ii) brain morphine concentration-time curves; and (iii) plasma M3G concentration-time curves. Morphine-6-glucuronide was not detected in any plasma or brain sample from rats administered morphine and no M3G was detected in brain. 4. For both control and renal failure rats, the relationships between antinociceptive effect and plasma morphine concentration were characterized by counterclockwise hysteresis loops, probably reflecting a delay for the relatively polar morphine to cross the blood-brain barrier. The relationship between antinociceptive effect and brain morphine concentration in control rats revealed no evidence of acute tolerance and was described by a sigmoidal function. In contrast, the relationship in renal failure rats was characterized by clockwise hysteresis, which is consistent with acute tolerance development. PMID- 10696533 TI - Inhibition of kinin degradation on the luminal side of renal tubules reduces high blood pressure in deoxycorticosterone acetate salt-treated rats. AB - 1. To determine whether the antihypertensive response in deoxycorticosterone acetate (DOCA) salt-treated rats was mediated by kinins on the luminal side of renal tubules or in the circulation, selective urinary kininase inhibitors were administered to normal Brown Norway Kitasato (BN-Ki) rats and kininogen-deficient Brown Norway Katholiek (BN-Ka) rats. 2. Kinins were degraded by neutral endopeptidase (NEP) and carboxypeptidase Y-like kininase (CPY) in urine, but were inactivated mainly by angiotensin-converting enzyme (ACE) in the plasma. 3. Ebelactone B inhibited CPY, while poststatin inhibited CPY and NEP. 4. Daily administration of poststatin (5 mg/kg per day, s.c.) for 3 days reduced blood pressure (BP) in DOCA salt-treated BN-Ki rats, but not in BN-Ka rats. 5. Ebelactone B (5 mg/kg per day, s.c.) also reduced BP in BN-Ki rats, which was accompanied by increased urinary sodium excretion, but had no effect on BP in BN Ka rats. 6. Lisinopril (5 mg/kg per day, s.c.) had no effect on BP in either rat strain. 7. Arterial kinin levels in BN-Ki rats increased significantly (2.2-4.6 pg/mL) with captopril (10 mg/kg, s.c.). However, arterial kinin levels that induced hypotension following the infusion of bradykinin (1000 ng/kg per min, i.v.) were 110-fold higher than endogenous arterial kinin levels attained following captopril. 8. These results suggest that inhibition of kinin degradation on the luminal side of the renal tubules may effectively attenuate hypertension. PMID- 10696534 TI - Effects of resistance training on bone parameters in young and mature rats. AB - 1. Osteoporosis is a major public health problem that is predicted to worsen over the next decade and preventative strategies that increase bone strength have become the focus of substantial research. 2. Although mechanical load is a primary factor in the acquisition and maintenance of skeletal tissue, the type of exercise used and when in life it is most effectively prescribed remain inconclusive. 3. The present study compared 10 weeks of resistance training in both young and mature female Sprague-Dawley rats and measured bone density and body composition by dual energy X-ray absorptiometry and biomechanical properties by three point bending tests of the tibia and femur. 4. No significant differences were observed for any of the bone parameters when comparing exercise and control groups at either age. This was despite using a comparable training protocol to that in humans and using loads of approximately 150% bodyweight. 5. The present study concludes that more intensive work programmes of resistance training or different outcome measures are required when using animal models for skeletal research. PMID- 10696535 TI - Carbonic anhydrase I inhibition by nitric oxide: implications for mediation of the hypercapnia-induced vasodilator response. AB - 1. At present, CO2 is considered to be the most important factor in regulating cerebral blood flow by modification of the interstitial fluid and extracellular pH, but the mechanism by which hypercapnia produces vasodilation is still controversial. In the present paper we investigated the effect of hypercapnia on carbonic anhydrase (CA) activity. We also studied the combined effects of CO2 with either indomethacin or an L-arginine analogue on CA activity. 2. Nine groups of 12 rabbits each were established. Groups 1-4 were ventilated with a mixture of 10% CO2, 21% O2 and 69% N2 for 20, 60, 120 and 180 min. Group 5 rabbits received 15 mg/kg bodyweight, i.v., indomethacin and, after 1 h, were ventilated with a mixture of 10% CO2, 21% O2 and 69% N2 for 2 h. Group 6 animals were ventilated with a mixture of 10% CO2, 21% O2 and 69% N2 for 2 h and then received indomethacin. Group 7 rabbits received 100 mg/kg bodyweight, i.v., NG-monomethyl L-arginine (L-NMMA) and, after 1 h, were ventilated with a mixture of 10% CO2, 21% O2 and 69% N2 for 2 h. Group 8 rabbits were ventilated for 2 h with a mixture of 10% CO2, 21% O2 and 69% N2 and were then administered L-NMMA. Group 9 rabbits received L-NMMA treatment concomitant with ventilation for 2 h with a mixture of 10% CO2, 21% O2 and 69% N2. In all groups, the erythrocyte CA activity was measured, as well as PaCO2 before and after ventilation or treatment. 3. The present study shows that CO2 reduces CA I activity down to complete inhibition and antagonizes the activating effects of indomethacin and L-NMMA on this isozyme. Our data prove that nitric oxide- and prostaglandin-induced CA I inhibition is involved in the vasodilation produced by hypercapnia. These results suggest that, due to subsequent pH changes, CA I is directly implicated in the modulation of vascular processes in the organism. PMID- 10696536 TI - Formation of the aqueous humor. AB - 1. Glaucoma is a worldwide disease affecting approximately 1-2% of the population aged over 35 years in industrial countries and is a major cause of blindness. 2. Glaucoma is usually associated with an increased intraocular pressure reflecting an imbalance between the rate of production of fluid (the aqueous humor) by the ciliary epithelial cells and its drainage from the eye. Therefore, it is important to understand how this secretion is produced. This requires a knowledge of ciliary epithelial cell composition, which has, in the past, proved difficult to obtain in mammalian preparations. 3. We have recently used the technique of electron-probe X-ray microanalysis to determine this composition under a variety of in vitro conditions. 4. Our results have led to a new model for this secretion that emphasizes the potential secretory role of the Na+/K+/2Cl- cotransporter. PMID- 10696537 TI - Comparative effect of flunarizine in two animal models of stroke. AB - 1. A common 'stroke model', the rat middle cerebral artery occlusion preparation, was used to compare photocoagulation with mechanical occlusion. 2. Areas of cerebral infarction and of vascular leakage were compared using these two methods. The general pattern of damage was similar, with central infarcted areas surrounded by larger vascular leakage areas. 3. The two models differed in the extent of their vascular leakage areas and in their response to the cerebroprotective agent flunarizine. 4. We conclude that the two models are not equivalent, probably because they do not damage the vascular endothelium to the same degree. PMID- 10696538 TI - Effect of hypoxia on respiratory activity in the foetus. AB - 1. Breathing movements stop soon after the induction of hypoxia in foetal animals, a response attributed to the active inhibition of the respiratory centres. Separate studies using punctate lesions have identified lateral pontine and thalamic sites in foetal sheep from which respiratory inhibition may arise, but whether either of these loci are actually oxygen sensitive or whether they receive input from other regions responsive to hypoxia, has not been established. 2. FOS immunocytochemistry was used to identify neuronal pools activated by hypoxia in the brain of late-gestation foetal and newborn sheep. FOS-positive cells were found in the pons in regions corresponding to the medial parabrachial and Kolliker-Fuse nuclei and were shown to be catecholaminergic. Neurons in this pontine region were also activated by the piperizine drug almitrine, which, like hypoxia, inhibits respiratory output in the foetus but is a respiratory stimulant in the newborn and adult. Because these pontine neurons do not express FOS protein after challenge with hypoxia or almitrine in the newborn lamb, we argue that they are crucial to the hypoxic inhibition of respiratory activity in foetal life. 3. The role of the placenta in determining these foetal responses and how this may change at birth is discussed. PMID- 10696539 TI - Effects of intra-uterine growth restriction on the control of breathing and lung development after birth. AB - 1. Low birthweight is now recognized as an important risk factor for early postnatal respiratory illness and it is becoming evident that low birthweight can increase the risk for airway dysfunction in children and adults. Our studies have been aimed at determining how low birthweight, resulting from intra-uterine growth restriction (IUGR), affects the control of breathing and the structural and functional development of the lung. 2. We have measured ventilatory responsiveness to progressive hypoxia and progressive hypercapnia during the first weeks after birth in postnatal lambs in which IUGR was induced by chronic placental insufficiency. It was found that the postnatal increase in ventilatory sensitivity to hypoxia observed in control lambs was diminished in low birthweight lambs; in contrast, the sensitivity to hypercapnia was not affected. In other studies, we found that IUGR caused by maternal anaemia led to elevated CO2 levels during sleep and wakefulness. 3. Our findings suggest that the prenatal development of the brain-stem or respiratory chemoreceptors may be affected by intra-uterine factors associated with IUGR, such as foetal hypoxaemia or hypoglycaemia. It is also possible that the structure of respiratory muscles and, hence, their ability to maintain a high level of ventilation may be affected by IUGR. 4. Recently, we studied the influence of IUGR on foetal lung development, in particular its effects on foetal lung liquid, a major determinant of lung growth, as well as alveolar structure and pulmonary surfactant. Lung liquid secretion and volume, in relation to bodyweight, were unaffected; however, there was evidence of structural and functional immaturity in the lungs. In foetuses exposed to IUGR, the air-blood barrier was thicker and, after birth, the diffusing capacity of the lungs for carbon monoxide was lower. In contrast, surfactant protein gene expression was enhanced, particularly in foetuses with high levels of circulating cortisol. 5. Further studies are needed to characterize the effects of specific types of prenatal compromise on postnatal control of ventilation and lung function, to determine mechanisms underlying these effects and to determine the capacity for postnatal recovery. PMID- 10696540 TI - Synaptic control of motoneuron excitability in rodents: from months to milliseconds. AB - 1. Motoneurons (MN) shape motor patterns by transforming inputs into action potential output. This transformation, excitability, is determined by an interaction between synaptic inputs and intrinsic membrane properties. Excitability is not static, but changes over multiple time scales. The purpose of the present paper is to review our recent data on synaptic factors important in the dynamic control of MN excitability over time scales ranging from weeks to milliseconds. 2. Developmental changes in modulation of MN excitability are well established. Noradrenergic potentiation of hypoglossal (XII) MN inspiratory activity in rhythmically active medullary slice preparations from rodents increases during the first two postnatal weeks. This is due to increasing alpha 1 and beta-adrenoceptor excitatory mechanisms and to a decreasing inhibitory mechanism mediated by alpha 2-adrenoceptors. Over a similar period, ATP potentiation of XII inspiratory activity does not change. 3. Motoneuron excitability may also change on a faster time scale, such as between different behaviours or different phases of a behaviour. Examination of this has been confounded by the fact that excitatory synaptic drives underlying behaviour can obscure smaller concurrent changes in excitability. Using the rhythmically active neonatal rat brain-stem-spinal cord preparation, we blocked excitatory inspiratory drive to phrenic MN (PMN) to reveal a reduction in PMN excitability specific to the inspiratory phase that: (i) arises from an inhibitory GABAergic input; (ii) is not mediated by recurrent pathways; and (iii) is proportional to and synchronous with the excitatory inspiratory input. We propose that the proportionality of the concurrent inhibitory and excitatory drives provides a means for phase-specific modulation of PMN gain. 4. Modulation across such diverse time scales emphasizes the active role that synaptic factors play in controlling MN excitability and shaping behaviour. PMID- 10696541 TI - Pattern formation and rhythm generation in the ventral respiratory group. AB - 1. There is increasing evidence that the kernel of the rhythm-generating circuitry for breathing is located within a discrete subregion of a column of respiratory neurons within the ventrolateral medulla referred to as the ventral respiratory group (VRG). It is less clear how this rhythm is transformed into the precise patterns appearing on the varied motor outflows. 2. Two different approaches were used to test whether subregions of the VRG have distinct roles in rhythm or pattern generation. In one, clusters of VRG neurons were activated or inactivated by pressure injection of small volumes of neuroactive agents to activate or inactivate groups of respiratory neurons and the resulting effects on respiratory rhythm and pattern were determined. The underlying assumption was that if rhythm and pattern are generated by neurons in different VRG subregions, then we should be able to identify regions where activation of neurons predominantly alters rhythm with little effect on pattern and other regions where pattern is altered with little effect on rhythm. 3. Based on the pattern of phrenic nerve responses to injection of an excitatory amino acid (DL homocysteate), the VRG was divided into four subdivisions arranged along the rostrocaudal axis. Injections into the three rostral regions elicited changes in both respiratory rhythm and pattern. From rostral to caudal the regions included: (i) a rostral bradypnoea region, roughly associated with the Botzinger complex; (ii) a dysrhythmia/tachypnoea area, roughly associated with the pre-Botzinger complex (PBC); (iii) a second caudal bradypnoea area; and, most caudally, (iv) a region from which no detectable change in respiratory motor output was elicited. 4. In a second approach, the effect of unilateral lesions of one subregion, the PBC, on the Breuer-Hering reflex changes in rhythm were determined. Activation of this reflex by lung inflation shortens inspiration and lengthens expiration (TE). 5. Unilateral lesions in the PBC attenuated the reflex lengthening of TE, but did not change baseline respiratory rhythm. 6. These findings are consistent with the concept that the VRG is not functionally homogenous, but consists of rostrocaudally arranged subregions. Neurons within the so-called PBC appear to have a dominant role in rhythm generation. Nevertheless, neurons within other subregions contribute to both rhythm and pattern generation. Thus, at least at an anatomical level resolvable by pressure injection, there appears to be a significant overlap in the circuitry generating respiratory rhythm and pattern. PMID- 10696542 TI - Cholinergic modulation of respiratory brain-stem neurons and its function in sleep-wake state determination. AB - 1. Shifts in behavioural state are controlled by reciprocal changes in discharge of cholinergic and aminergic groups of brain-stem/pontine neurons. During rapid eye movement (REM) sleep, cholinergic neurons are most active and aminergic neurons are least active. 2. Significant changes occur in the central control of breathing during REM sleep; respiration rate increases in frequency and variability, brain-stem respiratory neuron discharge is generally enhanced and the outputs of some respiratory motor neuron pools are depressed. 3. Hypoglossal motor neurons (HM) control tongue movement and their depression during REM sleep has been implicated in obstructive sleep apnoea. The cellular basis of HM depression has been investigated in vitro and may be due to enhanced activation of cholinergic receptors or decreased activation of aminergic receptors. 4. In vitro preparations that show respiratory rhythmogenesis possess advantages for the investigation of the neurochemical basis of state-dependent changes in respiration. Cholinergic changes in respiratory modulation of HM recorded in rhythmic brain-stem slices from mice depend on the site of activation of cholinergic receptors. PMID- 10696543 TI - Etiologic factors of acute aortic dissection in children and young adults. AB - Current concepts in the pathophysiology and predisposing conditions of acute aortic dissection in children, adolescents, and young adults are presented. Timely diagnosis is required for this life-threatening condition. Most children and adolescents with aortic dissection have congenital cardiovascular anomalies. Certain heritable disorders involving connective tissue also predispose to this disorder. Newer associations include cocaine abuse and weight lifting. To facilitate early diagnosis, the salient physical findings of the known predisposing conditions are reviewed. Clinical presentation and diagnostic imaging of aortic dissection are briefly summarized. Physicians working in an acute care setting, particularly in the emergency room, should be aware of disorders predisposing to acute aortic dissection in the pediatric and young adult population. Practitioners conducting school or college preparticipation sports evaluations can make use of such information in their assessment of risk for sudden death. PMID- 10696544 TI - Applying outpatient protocols in febrile infants 1-28 days of age: can the threshold be lowered? AB - The purpose of this study was to determine the applicability of two accepted outpatient management protocols for the febrile infant 1-2 months of age (Boston and Philadelphia protocols) in febrile infants 1-28 days of age. We retrospectively reviewed charts of patients 1-28 days of age with a temperature greater than or equal to 38.0 degrees C. Criteria from each of the above-cited management protocols were applied to the patients to determine their applicability in screening for serious bacterial infection (SBI). An SBI was defined as bacterial growth in cultures from blood, urine, cerebrospinal fluid (CSF), stool, or any aspirated fluid. Overall, 372 febrile infants were included in the study. Ages ranged from 1 to 28 days of age. The mean age was 15 days. SBI occurred in 45 patients (12%). The mean age of the patients with an SBI was 13 days. Thirty-two infants (8.6%) had a urinary tract infection; 12 (3.2%), bacteremia; five (1.3%), bacterial meningitis; three (0.8%), cellulitis; one (0.3%), septic arthritis; one (0.3%), bacterial gastroenteritis; and one (0.3%), pneumonia. Ten infants had more than one SBI. Of 372 patients, 231 (62%) met the Boston's laboratory low-risk criteria; eight (3.5%) would have been sent home with an SBI with these criteria. Philadelphia's laboratory low-risk criteria would have been met by 186 patients (50%); six (3.2%) would have been sent home with an SBI with these criteria. The negative predictive value of both the Boston and Philadelphia protocols for excluding an SBI was 97%. We conclude that current management protocols for febrile infants 1-2 months of age when applied to febrile infants 1 to 28 days of age would allow 3% of febrile infants less than 28 days of age to be sent home with an SBI. Current guidelines recommending admitting all febrile infants less than 28 days of age should be followed until the outcome of those 3% of febrile infants with an SBI treated as outpatients can be determined. PMID- 10696545 TI - The impact of a pediatric medical home on immunization coverage. AB - This study assessed whether having access to provisions in the American Academy of Pediatrics "medical home" concept was associated with being age-appropriately immunized at 3, 12, and 24 months. Cross-sectional data on 495 Delaware children were collected from June 1994 to June 1995. Immunization status was determined with the Delaware immunization registry. The medical home was not significantly associated with immunization coverage. This study confirms that race, insurance status, maternal education, and family incomes are predictive of having poor immunization outcomes. Simply providing medical homes may not be an effective strategy to improve use of preventive services. PMID- 10696546 TI - Assessing growth patterns--routine but sometimes overlooked. AB - We performed a retrospective chart review for 149 randomly selected well-child encounters to evaluate the frequency with which clinicians plot growth measurements and document growth abnormalities during health maintenance visits. Providers failed to plot at least one measurement of height, weight, and/or head circumference in 31 of 149 encounters (21%, 95% CI = 14.5% to 27.5%). Growth abnormalities in size, velocity, and/or disproportion were not documented in 22 of 40 relevant encounters (55%, 95% CI = 40% to 70%). Overall, 52 of 149 encounters (35%) were associated with an unplotted measurement and/or an undocumented growth abnormality. We feel that both documentation and assessment of growth represent potential areas for quality improvement. PMID- 10696547 TI - Prolonged morbidity due to delays in the diagnosis and treatment of obstructive sleep apnea in children. AB - The inclusion of a query concerning the presence of snoring in a questionnaires used by the Allergy Service of Childrens Hospital Los Angeles (CHLA) uncovered a significant number of patients who were experiencing prolonged and discomforting symptoms owing to previously undiagnosed obstructive sleep apnea (OSA) caused by adenotonsillar hypertrophy. Of 352 patients who were discharged with a diagnosis of OSA and tonsillectomy and/or adenoidectomy at CHLA in 1996-1997, a retrospective study of the first 45 randomly selected patients who agreed to participate in a telephone interview was performed. Analysis revealed that all patients experienced severe and discomforting symptoms with all describing severe or moderate snoring. Other symptoms included chronic mouth breathing (84%), frequent otitis media (64%), sinusitis (56%), sore throat (51%), choking (47%), and daytime drowsiness (42%). Other symptoms included poor school performance, enuresis, poor appetite and/or weight gain, dysphagia, and vomiting. Symptoms began at a mean age of approximately 2 years ("birth"-9 years), and the mean period of time between the development of significant symptoms and OSA was 3.3 years (6 months-13 years). Delay between onset of significant symptoms and surgery was > 1 year in 82% of the patients, > 2 years in 51% of the patients, > 4 years in 31% of the patients, and > 6 years in 13% of the patients. Forty percent of patients were self-referred to an otolaryngologist for treatment despite their primary care physician being aware of the symptoms. These results indicate that patient with OSA experienced prolonged morbidity and delays in treatment, which is probably widespread. Physician, parent, and third-party factors were found to have contributed to the delays in treatment. PMID- 10696548 TI - Anticonvulsant hypersensitivity syndrome vs Kawasaki disease: a challenging clinical diagnosis with therapeutic implications. PMID- 10696549 TI - Congenital rubella syndrome after maternal reinfection. PMID- 10696550 TI - Factitious hyperinsulinemic hypoglycemia in infancy: diagnostic pitfalls. PMID- 10696551 TI - A pilot smoking cessation program run by pediatric respiratory care practitioners for parents. PMID- 10696552 TI - Routine and ritual circumcision is a multi-cultural phenomenon. PMID- 10696553 TI - The trapped penis is a condition in which scarring at the distal prepuce forms a contracture that binds down the shaft of the penis. PMID- 10696555 TI - Photo quiz. Erythema infectiosum. PMID- 10696554 TI - Rectal prolapse in pediatrics. PMID- 10696556 TI - Adolescent striae. PMID- 10696557 TI - Botanical briefs: giant hogweed--Heracleum mantegazzianum Sommier & Levier. PMID- 10696558 TI - Cocaine-induced centrofacial ulceration. PMID- 10696559 TI - Dermatofibrosarcoma protuberans in two patients with acquired immunodeficiency syndrome. AB - Dermatofibrosarcoma protuberans (DFSP) is a locally aggressive cutaneous tumor of intermediate malignancy. Most commonly, it arises as an asymptomatic, indurated plaque on the trunk within which protuberant nodules develop over time. We describe its occurrence in two patients with human immunodeficiency virus, a previously unreported association. The first patient, a 41-year-old woman, complained of painful lesions around the left shoulder that developed within a scar from previous trauma to the area. The second patient, a 50-year-old man, developed a recurrent DFSP within the scar from a previous surgical procedure. Dermatofibrosarcoma protuberans was confirmed in both cases by the histopathologic and immunohistochemical findings. PMID- 10696560 TI - Scleromyxedema: successful treatment of cutaneous and neurologic symptoms. AB - Scleromyxedema is a rare systemic disorder characterized by cutaneous sclerosis and papulosis, accompanied by deposition of mucin in the skin and other organs. We describe a case of scleromyxedema in a 62-year-old man. The cutaneous symptoms of the disorder were preceded by episodes of acute central nervous system dysfunction that included mental confusion, hemiparesis, tremor, and migraine. As the cutaneous symptoms progressed, the patient experienced persistent confusion and difficulty concentrating. Therapy with melphalan and plasmapheresis led to complete resolution of the cutaneous symptoms as well as near-resolution of the neurologic symptoms. This is the first report to describe the successful treatment of the cutaneous symptoms of scleromyxedema accompanied by reversal of chronic neurologic dysfunction. PMID- 10696561 TI - Melanoma in a psoriatic plaque. AB - This is the first reported case of a melanoma in a psoriatic plaque. The clinical, dermoscopic, and histologic features of this case are detailed. A review of the risk of melanoma among patients treated with psoralen-ultraviolet A is presented. PMID- 10696562 TI - Treatment of granuloma faciale with the pulsed dye laser. AB - Granuloma faciale is a chronic benign vasculitis that generally affects the skin of the face. The lesions are commonly refractory to therapy. A patient with long standing granuloma faciale refractory to topical corticosteroid and dapsone therapy had an excellent response to treatment with the pulsed dye laser. PMID- 10696563 TI - Inexpensive digital photography in clinical dermatology and dermatologic surgery. AB - Digital photography can be used to follow clinical improvement in a variety of dermatoses, document pre and postoperative results, and document histologic findings from skin biopsies. Images may be printed as part of text documents and can be filed in conventional medical charts. Images can be transmitted via electronic mail rapidly and to any location. We have found digital photography to be relatively inexpensive and a powerful tool to enhance dermatologic practice. PMID- 10696564 TI - Gingival lesions and nasal obstruction in an immunosuppressed patient post-liver transplantation. AB - Although rare, metastatic hepatocellular carcinoma (HCC) presenting only to the mandible, gingiva, and nasal cavity in patients subsequently found to have primary HCC has been reported. In the age of transplantation, certain HCC patients may receive treatment with an orthotopic liver transplant. Due to the proclivity of HCC for early micrometastases, immunosuppressive therapy can induce significant metastatic lesions. Nasal mass obstruction, gingival lesions, or facial growths in this population must be considered metastatic until proven otherwise. PMID- 10696565 TI - The search for pathogenic T cells and the genetic basis of psoriasis using a severe combined immunodeficient mouse model. AB - The immunologic and genetic bases of psoriasis are under active investigation throughout the world. Rather than pursue the genetic linkage to psoriasis to discover the gene(s) responsible for causing the disease, we have focused on the cellular immunology and basic biology using a novel animal model. We reasoned by identifying specific cellular and molecular abnormalities involved in the biologic responses that initiate lesion formation, that the genes involved in such a pathologic process would lead us to the correct causative DNA-based abnormality that determines disease susceptibility and inheritance. To pursue this line of inquiry, we utilized an animal model in which severe combined immunodeficient (SCID) mice were engrafted with symptomless skin (PN skin), and bona fide psoriatic plaques (PP skin) were created using specific pathogenic T cell subsets. This model can be used experimentally not only to study the mechanism by which PP skin is converted to PN skin, but also to create PP skin from PN skin. The clinical, histologic, immunologic, and pharmacologic validation of this SCID mouse model will be presented. This summary will also highlight the value of such a model, which has recently led to the discovery of previously overlooked types of immunocytes that are associated with induction of psoriatic lesions. Finally, a novel hypothesis linking the immunology and the genetics of psoriasis, based on findings using this animal model, will conclude this presentation. PMID- 10696566 TI - Systemic contact dermatitis caused by oral chromium picolinate. AB - Although nickel is the metal most commonly implicated in systemic contact dermatitis, other metals such as chromate have also been known to cause dermatitis when ingested. Chromium picolinate has been espoused as a nutritional supplement. Allegedly, it helps control blood sugar in diabetes and may reduce cholesterol and blood pressure levels. PMID- 10696567 TI - Definition and measurement of follicle stimulating hormone. AB - FSH has a key role in the development and function of the reproductive system and is widely used both diagnostically and therapeutically in developmental and reproductive medicine. The accurate measurement of FSH levels, in patients for diagnosis and monitoring and in therapeutic preparations for clinical use, is essential for safe and successful treatment. Historically, FSH was defined on the basis of classical in vivo endocrine activity, and early therapeutic preparations were calibrated using in vivo bioassays. There was early recognition that reference preparations were required for calibration if the results from different laboratories were to be comparable. In response to the perceived need, the World Health Organization established the first standard for such preparations in 1959. Subsequent developments in biotechnology have led to recognition that there is no single molecule that can be uniquely defined as FSH, and that FSH can induce a range of biological activities. Several highly purified standards for FSH are now available, but discontinuity and heterogeneity of estimates of FSH activity in terms of these standards made using in vitro assays and binding assays have been noted. It is thus essential that any measurement of FSH include specification both of the standard with which the measured FSH is compared and the assay method used for that comparison. PMID- 10696568 TI - The molecular and genetic basis of fibroblast growth factor receptor 3 disorders: the achondroplasia family of skeletal dysplasias, Muenke craniosynostosis, and Crouzon syndrome with acanthosis nigricans. AB - Achondroplasia, the most common form of short-limbed dwarfism in humans, occurs between 1 in 15,000 and 40,000 live births. More than 90% of cases are sporadic and there is, on average, an increased paternal age at the time of conception of affected individuals. More then 97% of persons with achondroplasia have a Gly380Arg mutation in the transmembrane domain of the fibroblast growth factor receptor (FGFR) 3 gene. Mutations in the FGFR3 gene also result in hypochondroplasia, the lethal thanatophoric dysplasias, the recently described SADDAN (severe achondroplasia with developmental delay and acanthosis nigricans) dysplasia, and two craniosynostosis disorders: Muenke coronal craniosynostosis and Crouzon syndrome with acanthosis nigricans. Recent evidence suggests that the phenotypic differences may be due to specific alleles with varying degrees of ligand-independent activation, allowing the receptor to be constitutively active. Since the Gly380Arg achondroplasia mutation was recognized, similar observations regarding the conserved nature of FGFR mutations and resulting phenotype have been made regarding other skeletal phenotypes, including hypochondroplasia, thanatophoric dysplasia, and Muenke coronal craniosynostosis. These specific genotype-phenotype correlations in the FGFR disorders seem to be unprecedented in the study of human disease. The explanation for this high degree of mutability at specific bases remains an intriguing question. PMID- 10696569 TI - Is estradiol a genotoxic mutagenic carcinogen? AB - The natural hormone 17 beta-estradiol (E2) induces tumors in various organs of rats, mice, and hamsters. In humans, slightly elevated circulating estrogen levels caused either by increased endogenous hormone production or by therapeutic doses of estrogen medications increase breast or uterine cancer risk. Several epigenetic mechanisms of tumor induction by this hormone have been proposed based on its lack of mutagenic activity in bacterial and mammalian cell test systems. More recent evidence supports a dual role of estrogen in carcinogenesis as a hormone stimulating cell proliferation and as a procarcinogen inducing genetic damage. Tumors may be initiated by metabolic conversion of E2 to 4 hydroxyestradiol catalyzed by a specific 4-hydroxylase (CYP1B1) and by further activation of this catechol to reactive semiquinone/quinone intermediates. Several types of direct and indirect free radical-mediated DNA damage are induced by E2, 4-hydroxyestradiol, or its corresponding quinone in cell-free systems, in cells in culture, and/or in vivo. E2 also induces various chromosomal and genetic lesions including aneuploidy, chromosomal aberrations, gene amplification, and microsatellite instability in cells in culture and/or in vivo and gene mutations in several cell test systems. These data suggest that E2 is a weak carcinogen and weak mutagen capable of inducing genetic lesions with low frequency. Tumors may develop by hormone receptor-mediated proliferation of such damaged cells. PMID- 10696570 TI - How do glucocorticoids influence stress responses? Integrating permissive, suppressive, stimulatory, and preparative actions. AB - The secretion of glucocorticoids (GCs) is a classic endocrine response to stress. Despite that, it remains controversial as to what purpose GCs serve at such times. One view, stretching back to the time of Hans Selye, posits that GCs help mediate the ongoing or pending stress response, either via basal levels of GCs permitting other facets of the stress response to emerge efficaciously, and/or by stress levels of GCs actively stimulating the stress response. In contrast, a revisionist viewpoint posits that GCs suppress the stress response, preventing it from being pathologically overactivated. In this review, we consider recent findings regarding GC action and, based on them, generate criteria for determining whether a particular GC action permits, stimulates, or suppresses an ongoing stress-response or, as an additional category, is preparative for a subsequent stressor. We apply these GC actions to the realms of cardiovascular function, fluid volume and hemorrhage, immunity and inflammation, metabolism, neurobiology, and reproductive physiology. We find that GC actions fall into markedly different categories, depending on the physiological endpoint in question, with evidence for mediating effects in some cases, and suppressive or preparative in others. We then attempt to assimilate these heterogeneous GC actions into a physiological whole. PMID- 10696571 TI - Uncovering molecular mechanisms involved in activation of G protein-coupled receptors. AB - G protein-coupled, seven-transmembrane segment receptors (GPCRs or 7TM receptors), with more than 1000 different members, comprise the largest superfamily of proteins in the body. Since the cloning of the first receptors more than a decade ago, extensive experimental work has uncovered multiple aspects of their function and challenged many traditional paradigms. However, it is only recently that we are beginning to gain insight into some of the most fundamental questions in the molecular function of this class of receptors. How can, for example, so many chemically diverse hormones, neurotransmitters, and other signaling molecules activate receptors believed to share a similar overall tertiary structure? What is the nature of the physical changes linking agonist binding to receptor activation and subsequent transduction of the signal to the associated G protein on the cytoplasmic side of the membrane and to other putative signaling pathways? The goal of the present review is to specifically address these questions as well as to depict the current awareness about GPCR structure-function relationships in general. PMID- 10696572 TI - Electrodiagnostic evaluation of traumatic nerve injuries. AB - This article reviews the techniques of motor and sensory nerve conduction studies and needle electromyography methods, which are particularly useful for localizing nerve injuries in upper extremity and hand trauma. Included are details of methods for detecting and quantifying the degree of axon loss and for using this information to make treatment decisions and predict outcomes. The epidemiology and classification of traumatic peripheral nerve injuries, the effects of these injuries on nerve and muscle, and the means by which electrodiagnosis is used to help classify the injury are described. An overview of recovery mechanisms also is presented. PMID- 10696574 TI - Intraoperative monitoring of nerve repairs. AB - These examples illustrate the interplay of the electrophysiologic techniques of SEP, EMG, NCAPs, and nerve conduction studies to achieve the goals of monitoring, guiding, identifying and localizing, and assessing nerve function. Through their successful application, intraoperative neurophysiology moves from solely monitoring to serving as an electrophysiologic aid to the surgeon. Using all of these techniques provides a comprehensive analysis of nerve function. The overall goal is both to prevent intraoperative neural compromise and to optimize postoperative outcome by providing the surgeon with the best possible information regarding the functional condition of the nerves under investigation. PMID- 10696573 TI - MR nerve imaging of the wrist and hand. AB - Peripheral nerve imaging is still in a relatively early phase of development. This article explains a few of the important technical aspects of peripheral nerve imaging and reviews the results of animal experiments that form the foundation for interpreting image results. The clinical discussion focuses on the use of MR imaging for patients with carpal tunnel syndrome, keeping in mind that many aspects of the imaging technique may be applied to other areas. The reliability, diagnostic accuracy, and potential indications for peripheral nerve imaging are discussed. PMID- 10696575 TI - Repair of peripheral nerve defects in the upper extremity. AB - Repair of peripheral nerve defects in the upper extremity using end-to-end coaptation is accomplished by one of four techniques: in situ mobilization, rerouting and transposition, joint positioning, and bone shortening. A key concern is the amount of tension generated when nerves are elongated to overcome a gap defect. The evidence indicates that elongation should be limited to 8% to 10% of the original length to avoid neural ischemia. It should be noted, however, that when repairs are delayed, the vascularity of nerves is increased. As a result, compared with acute injuries, chronic injuries will tolerate the same degree of elongation with less neural ischemia despite increased stiffness. The mesoneural attachments along each end of the nerve may be safely stripped to a distance of 8 to 12 cm when mobilizing the nerve. Larger nerves tolerate greater lengths of mobilization than smaller nerves. The maximum amount of mobilization that does not produce ischemia can be expressed as a ratio of the diameter of the nerve to the length mobilized and its value is 1:45. The amount of nerve mobilization required for a secondary repair may be reduced by the initial application of tension to unrepaired nerves, thereby reducing the amount of retraction. As the interval to repair increases, nerve retraction results in up to a six-fold increase in the gap defect that must be overcome. Finally recommendations exist for the repair of peripheral nerve segmental defects in the acute setting. PMID- 10696576 TI - Techniques for primary nerve repair. AB - This article reviews the anatomy of the peripheral nerve, the pathophysiology of nerve injury, and Wallerian degeneration. It reviews the factors for deciding on immediate or delayed primary nerve repair and discusses the concept of longitudinal excursion of peripheral nerves about joints and the techniques for achieving an appropriate tension-free repair. The techniques of primary nerve repair, epineurial repair, and group fascicular repair are reviewed along with techniques for matching fascicles intraoperatively. PMID- 10696577 TI - Functional results of primary nerve repair. AB - Age and mechanism of injury, both out of the control of the surgeon, seem to be the prime determinants of functional outcome following nerve repair. Careful application of the principles of nerve repair can ensure that the appropriate method of repair--primary, secondary, or nerve graft--is selected, and this may be the best way to increase the chances for a successful result. PMID- 10696578 TI - Techniques for nerve grafting. AB - The nerve graft can be completed using many different techniques. This article elaborates the general principles and many details that can be approached in different ways when dealing with nerve grafting. A review of the reasons for the poor performance of early nerve grafts, as well as the origins of nerve grafts, is included. This article discusses the technical aspects of nerve grafting including free nerve grafting and timing and exposure. PMID- 10696579 TI - Functional outcomes of nerve grafts for the upper and lower extremities. AB - In the early half of this century, nerve grafting was not thought to be a viable option in the management of nerve injuries. With modern techniques, patients now can expect useful recovery of sensory and motor function following nerve grafting of peripheral nerves. By providing standardized and quantifiable data, investigators may combine their clinical series to identify factors that further enhance nerve repair. PMID- 10696580 TI - The physiology of nerve transplantation. AB - As microsurgical techniques improve, attempts to restore function after severe nerve injury place greater demands upon the use of nerve grafts. Viable Schwann cells within these grafts are necessary to maximize nerve regeneration. Progress in decreasing nerve allograft antigenicity and the host response through immunosuppression may provide results comparable to nerve autografts. Although pretreatment methods aimed at reducing allograft antigenicity have yielded inconsistent and overall unsatisfactory results, cryopreservation can maintain Schwann cell viability and shows promise as a means of tissue banking. In addition, tissue typing for MHC I or II antigens and advances in immunosuppressive therapy have rendered encouraging results in experimental models. The use of transplanted nerve allografts remains experimental, yet efforts to understand the host rejection response, advances in immunotherapy, and the development of neurotrophic factors continue to reveal significant benefits over standard treatment methods. PMID- 10696581 TI - Use of nerve conduits in peripheral nerve repair. AB - Studies on nerve conduits for peripheral nerve regeneration have concentrated on the manipulation of various conduit materials to avoid sacrificing native nerve in the clinical situation. With the proliferation of available nerve growth stimulating factors, the focus is shifting experimentally toward molecular biologic manipulation, with the addition of these materials as substrates within the conduit. The clinical use of conduits has concentrated on the use of autogenous tissue, with a few examples of polyglactin (PGA) mesh and silicone. Ultimately, as yet, conduit material does not seem to have a profound effect on outcome. Substrate manipulation has not yet had clinical application. An important problem that remains, both experimentally and clinically, is overriding the size of the maximal gap that can be bridged successfully, as well as obtaining good functional sensory and motor recovery, compared with the use of nerve grafts. Advances in molecular biology may reveal further details about the nerve growth phenomenon, the precise sequencing of the substrate materials that are effective in promoting nerve growth, and when they should be applied. Advances in chemical engineering may provide additional biologically stable materials that have the ability to integrate growth-enhancing agents or factors into the lumen of the conduit. PMID- 10696582 TI - Nerve transfers in the upper extremity. AB - Restoration of extremity function following nerve injury is often unpredictable. Nerve transfers in the upper extremity are important techniques in the management of many types of peripheral nerve injury. The physiologic principles of nerve transfer lead to the indications for use. The elements of planning and execution of common nerve transfers are presented in this article. PMID- 10696583 TI - Partial nerve injuries in the upper extremity. AB - The level of injury of a peripheral nerve is a critical factor that has a great impact on the result of the repair. At the level of the wrist, the median and ulnar nerves have pure motor and sensory fascicular groups. Proximal to the wrist, the motor fascicular groups combine with sensory fascicles and become mixed nerves. Mapping the fascicular orientation with electrical stimulation is indicated for injuries located from the wrist to the distal third of the forearm. Successful application of this technique depends on the level of injury, anesthetic technique, and careful patient selection. Children and patients with other serious coexisting injuries are not candidates for this technique. The depth of anesthesia must provide adequate analgesia while allowing the patient to communicate and cooperate with the surgeon during the procedure. There are few reports in the literature about repair of partially injured nerves in the upper extremities and the comparison of functional outcomes with or without the use of nerve grafts is not easy. Even under ideal operative conditions and with ideal indications, the outcomes are not always satisfactory. Hurst et al reported very good results using end-to-end repair of fascicular groups in their series. Using the rating system of the British Medical Research Council, they reported motor values of 4.0 (normal 5.0), and sensory values of 3.8 (normal 4.0). Kato et al reported very good results in their series of 51 cases with group fascicular end to-end suture using orientation with electrical stimulation. In this series, there were five patients with partial nerve laceration and end-to-end coaptation of the fascicular groups provided very satisfactory outcome. End-to-end repair of the fascicular groups seems to provide better results than repair of the nerve using nerve grafts. It is desired, however, that the nerve gap be less than 2 cm for the application of end-to-end repair of the nerve. PMID- 10696584 TI - End-to-side nerve repair. A review. AB - In recent years, there has been a resurgence of end-to-side peripheral nerve repair. This technique offers a management of a peripheral nerve defect in the absence of a suitable proximal stump. Although numerous animal laboratory investigations demonstrate motor and sensory functional recovery without deleterious effects to the donor nerve, clinical outcomes are yet to be determined. PMID- 10696585 TI - The clip-domain family of serine proteinases in arthropods. PMID- 10696586 TI - Homology modeling of the insect chitinase catalytic domain--oligosaccharide complex and the role of a putative active site tryptophan in catalysis. AB - Knowledge-based protein modeling and substrate docking experiments as well as structural and sequence comparisons were performed to identify potential active site residues in chitinase, a molting enzyme from the tobacco hornworm, Munduca sexta. We report here the identification of an active-site amino acid residue, W145. Several mutated forms of the gene encoding this protein were generated by site-directed mutagenesis, expressed in a baculovirus-insect cell-line system, and the corresponding mutant proteins were purified and characterized for their catalytic and substrate-binding properties. W145, which is present in the presumptive catalytic site, was selected for mutation to phenylalanine (F) and glycine (G), and the resulting mutant enzymes were characterized to evaluate the mechanistic role of this residue. The wild-type and W145F mutant proteins exhibited similar hydrolytic activities towards a tri-GlcNAc oligosaccharide substrate, but the former was approximately twofold more active towards a polymeric chitin-modified substrate. The W145G mutant protein was inactive towards both substrates, although it still retained its ability to bind chitin. Therefore, W145 is required for optimal catalytic activity but is not essential for binding to chitin. Measurement of kinetic constants of the wild-type and mutant proteins suggests that W145 increases the affinity of the enzyme for the polymeric substrate and also extends the alkaline pH range in which the enzyme is active. PMID- 10696587 TI - Structure of the VAP-peptide (BmACP-6.7) gene in the silkworm, Bombyx mori and a possible regulation of its expression by BmFTZ-F1. AB - The VAP-peptide (BmACP-6.7) is a hydrophobic peptide localized in adult cuticle of the silkworm, Bombyx mori. We isolated and characterized the VAP-peptide gene as a useful marker gene to analyze molecular mechanisms of terminal differentiation processes in the adult. The gene is composed of two exons interrupted by one intron. The 5' upstream promoter region is shown to bear a nucleotide sequence similar to the cis-element that is recognized and bound by the Bombyx mori FTZ-F1 protein (BmFTZ-F1). Expression of the BmFTZ-F1 gene preceded expression of the VAP-peptide gene and injection of 20-hydroxyecdysone suppressed the expression of both genes. An in vitro binding assay indicated direct interaction of BmFTZ-F1 with the VAP-peptide gene promoter sequence. Therefore, BmFTZ-F1 is proposed to be a possible factor regulating the stage specific expression of the VAP-peptide gene towards adult life. PMID- 10696588 TI - Isolation and characterization of CRF-related diuretic hormones from the whitelined sphinx moth Hyles lineata. AB - We have isolated and characterized two diuretic hormones (DH), Hylli-DH41 and Hylli-DH30, from extracts of whole heads of the lepidopteran Hyles lineata. We monitored the isolation by measuring the ability of fractions to affect levels of cyclic AMP production by Malpighian tubules of Manduca sexta maintained in vitro. These DH are related to a family of vertebrate neuropeptides which includes sauvagine, corticotropin-releasing factor (CRF), and urotensin I. Both Hylli-DH41 (RMPSLSIDLPMSVLRQKLSLE KERKVQALRAAANRNFLNDI-NH2) and Hylli-DH30 (SFSVNPAVEILQHRYMEKVAQNNRNFLNRV-NH2) show extremely high similarity with two DH from the tobacco hornworm M. sexta. This is not surprising because both H. lineata and M. sexta are sphingid moths. The discovery of these DH provides a third example of two CRF-related DH occurring in one insect species. PMID- 10696589 TI - Calcofluor disrupts the midgut defense system in insects. AB - The insect midgut is generally lined with a unique protective chitin/protein structure, the peritrophic membrane (PM). We demonstrated that in Trichoplusia ni larvae, the majority of PM proteins were assembled with chitin as a consequence of their chitin binding properties. These proteins could be dissociated from the PM in vitro by Calcofluor, a well-known chemical with chitin binding properties. The chitin binding characteristics of PM proteins were confirmed by their high affinity binding in vitro to regenerated chitin. In vivo assays demonstrated that Calcofluor could inhibit PM formation in five lepidopteran insects tested. The inhibition of T. ni PM formation by Calcofluor, was accompanied by increased larval susceptibility to baculovirus infection. Continuous inhibition of PM formation by Calcofluor resulted in retarded larval development and mortality. The destructive effect of Calcofluor on PM formation was demonstrated to be transient and reversible depending on the presence of Calcofluor within the midgut. In addition, degradation of the insect intestinal mucin was observed concurrently with the inhibition of PM formation by Calcofluor. Our studies revealed a potential novel approach to develop strategies for insect control by utilizing chitin binding molecules to specifically target PM formation in a broad range of insect pest species. PMID- 10696590 TI - Low molecular weight serine protease inhibitors from insects are proteins with highly conserved sequences. AB - A low molecular weight protease inhibitor peptide found in ovaries of the desert locust Schistocerca gregaria (SGPI-2), was purified from plasma of the same locust and sequenced. It was named SGCI. It was found active towards chymotrypsin and human leukocyte elastase. SGCI was synthesized using a solid-phase procedure and the sequence of its reactive site for chymotrypsin was determined. Compared with an inhibitor purified earlier from another locust species, the total sequence of SGCI showed 88% identity. In particular, the sequence of the reactive site of these inhibitors was identical. Our search for a closely related peptide in an insect species far removed from locusts, the lepidopteran Spodoptera littoralis, was unfruitful but a different chymotrypsin inhibitor, belonging to the Kazal family, was found whose mass is greater than that of SGCI (20 vs 3.6 kDa). Its N-terminal sequence shares 80% identity with that of a chymotrypsin inhibitor purified earlier from the haemolymph of another lepidopteran. Conservation of the amino acid sequence in the reactive site seems to be an exception among protease inhibitors. PMID- 10696591 TI - Isolation and sequencing of a cDNA encoding for a ribonuclease from the insect Ceratitis capitata. AB - Two overlapping clones encoding for a ribonuclease from six-day-old larvae of the insect Ceratitis capitata (Cc-RNase) have been isolated by immunoscreening a cDNA library and by 5' RACE. The sequence of the Cc-RNase cDNA contains an open reading frame of 414 nucleotides encoding for a precursor protein of 138 amino acids long with a putative signal peptide consisting of 19 amino acids. The calculated M(r) of the mature protein was found to be 13.7 kDa. Multiple alignments of the deduced amino acid Cc-RNase sequence with other ribonucleases revealed an approximate 25% average identity. Despite the low percentage of identity, histidine and lysine residues which are essential for its catalytic activity, were found to be completely conserved. Furthermore, expression of the clone in E. coli resulted in the production of a recombinant product that showed strong immunoreactivity with anti-RNase specific antibodies. These results support the hypothesis that the identified clone encodes for a protein which is a new member of the RNase superfamily. PMID- 10696592 TI - Molecular cloning, nucleotide sequence and gene expression of a cytochrome P450 (CYP6F1) from the pyrethroid-resistant mosquito, Culex quinquefasciatus Say. AB - To analyze cytochrome P450s in the southern house mosquito, Culex quinquefasciatus, we quantified the content of P450s and b5 in larval microsomes of guts and carcasses. Results indicated that content was 30 times higher in guts than in carcasses. A conserved region in the alignment of insect P450 family 6 (CYP6) proteins served as a guide for the synthesis of degenerate oligonucleotide primers to clone P450 cDNAs. Primers were used in the reverse transcription polymerase chain reaction (RT-PCR) of gut mRNA from 4th-instar larvae of the permethrin-susceptible or resistant C. quinquefasciatus. PCR products of ca. 250 base pairs (bp) were cloned, and nucleotide sequences of 35 clones from susceptible and 28 from resistant strains determined. Alignment of the deduced amino acid sequences from these clones showed them to be classifiable into six isoforms. We next screened a cDNA clone (CYP6F1) from a gut cDNA library and determined the nucleotide sequence. Northern blot analysis showed that the CYP6PF1 gene in the permethrin-resistant strain appeared to be expressed more strongly than in the susceptible strain. The deduced amino acid of CYP6F1 showed that it has conserved domains of a membrane-anchoring signal, reductase binding sites, a heme-binding site, ETLR motif and substrate recognition sites in P450s. Phylogenetic analysis showed that CYP6F1 is strongly related to CYP6D1 involved in pyrethroid detoxification. PMID- 10696593 TI - Cloning, sequence analysis and expression in Escherichia coli of the gene encoding a uricase from the yeast-like symbiont of the brown planthopper, Nilaparvata lugens. AB - A urate oxidase (uricase; EC 1.7.3.3) gene of the yeast-like fungal endosymbiont of the brown planthopper, Nilaparvata lugens, was cloned, and sequenced together with its flanking regions. The gene comprised a open reading frame of 987 bp, that was split into two parts by a single 96 bp intron. The encoded uricase was 296 amino acids with 62% sequence identity with that of Aspergillus flavus. The molecular weight deduced was 32,882, and the predicted isoelectric point was 6.06. The symbiont's uricase conserved all the known consensus motifs, except the C-terminal PTS-1, Ser-basic-Leu. The leucine at the third position of PTS-1 was replaced by serine in the C-terminus of the symbiont's uricase. The symbiont's uricase gene was successfully expressed in Escherichia coli, and the product, tagged with histidine residues, was purified. The symbiont's uricase, thus produced, was as active as those from plants and animals, but less active than those from other fungi. PMID- 10696594 TI - Journal of Digital Imaging: the first 12 years and beyond. PMID- 10696595 TI - PACS databases and enrichment of the folder manager concept. AB - Current challenges facing picture archiving and communication systems (PACS) center around database design and functionality. Workflow issues and folder manager concepts such as autorouting, prefetching, hanging protocols, and hierarchical storage management are driven by a properly designed database that ultimately directly impacts the clinical utility of a PACS. The key issues in PACS database design that enable radiologist-friendly, cost-effective, and data secure systems will be discussed, including database difficulties of the DICOM standard, HIS/RIS/PACS (hospital information system/radiology information system) connectivity, and database issues in data acquisition, data dissemination, and data display. PMID- 10696596 TI - Fracture interpretation using electronic presentation: a comparison. AB - The purposes of this study were to determine whether (1) fractures are interpreted differently after digitization and electronic presentation; (2) there are differences in accuracy between screen radiographs and electronic presentation; (3) differences in interpretation are a function of monitor resolution; and (4) differences in interpretation between radiographs and electronic images relate to radiological subspecialty. Forty cases with fractures of varying degrees of subtlety and 35 cases without fractures were interpreted. Radiographs were digitized with 2 different systems and displayed on 3 monitors of different spatial resolution. Four radiologists, with varying experience, were asked to decide whether fractures were present, absent, or they were uncertain. Accuracy of interpretation increased with improved electronic image presentation and monitor resolution. The sensitivity, specificity, and accuracy of fracture detection on System A were 63%, 98%, and 78%, respectively. The results were 72%, 98%, and 84% with System B. System C results were 81%, 97%, and 88% with Lumiscan 75, and 82%, 96%, and 88% with Lumiscan 150. Sensitivity, specificity, and accuracy results of the original radiograph interpretation were 89%, 95%, and 92%. Results were significantly different for System A. No significant differences were found for the other systems compared with film radiographs. System A did not have adequate monitors for interpretation of subtle fractures. Systems B and C were capable of displaying even subtle fractures. Our initial results indicate that interpretation with high-quality 1K x 1K monitors is substantially similar to radiograph interpretation. PMID- 10696597 TI - Clinical input into designing a PACS. AB - The purpose of this study was to evaluate clinical attitudes and expectations in the implementation of a neuroradiology picture archiving and communication system (PACS). A 1-page survey of expectations and clinical attitudes toward a neuroradiology mini-PACS was distributed to 49 full-time faculty members in the departments of neurosurgery, neurology, and otorhinolaryngology at an academic center. Interest in viewing soft-copy images was moderate to very high for over 89% of clinicians. All clinicians were comfortable with phone consultations with radiologists while viewing soft-copy images. Clinicians preferred retrieving images from personal computers over workstations and film libraries by 72.9%, 27.1%, and 0%, respectively. However, 38.5% of surgeons felt the need for hard copy in the operating room. Clinicians estimated that in 18.3% of cases, patients took their in-house films to outside institutions for consultations. Clinicians were enthusiastic about implementing PACS. Although acceptance of soft-copy viewing among clinicians is high, some provision for supplying hard-copy images appears to be necessary. PMID- 10696598 TI - Ultrasound grayscale image compression with JPEG and wavelet techniques. AB - The purpose of the study was to evaluate the effects of lossy compression on grayscale ultrasound images to determine how much compression can be applied while still maintaining images that are acceptable for diagnostic purposes. The study considered how the acquisition technique (video frame-grabber versus directly acquired in digital form) influences how much compression can be applied. For directly acquired digital images, the study considered how text (that is burned into the image) affects the compressibility of the image. The lossy compression techniques that were considered include JPEG and a Wavelet algorithm using set partitioning in hierarchical trees (SPIHT). PMID- 10696599 TI - Low-cost soft-copy display accuracy in the detection of pulmonary nodules by single-exposure dual-energy subtraction: comparison with hard-copy viewing. AB - This study endeavored to clarify the usefulness of single-exposure dual-energy subtraction computed radiography (CR) of the chest and the ability of soft-copy images to detect low-contrast simulated pulmonary nodules. Conventional and bone subtracted CR images of 25 chest phantom image sets with a low-contrast nylon nodule and 25 without a nodule were interpreted by 12 observers (6 radiologists, 6 chest physicians) who rated each on a continuous confidence scale and marked the position of the nodule if one was present. Hard-copy images were 7 x 7-inch laser-printed CR films, and soft-copy images were displayed on a 21-inch noninterlaced color CRT monitor with an optimized dynamic range. Soft-copy images were adjusted to the same size as hard-copy images and were viewed under darkened illumination in the reading room. No significant differences were found between hard- and soft-copy images. In conclusion, the soft-copy images were found to be useful in detecting low-contrast simulated pulmonary nodules. PMID- 10696600 TI - Diagnostic accuracy of film-based, TIFF, and wavelet compressed digital temporomandibular joint images. AB - The purpose of this research was to determine if digitization and the application of various compression routines to digital images of temporomandibular joint (TMJ) radiographs would diminish observer accuracy in the detection of specific osseous characteristics associated with TMJ degenerative joint disease (DJD). Nine observers viewed 6 cropped hard-copy radiographic films each of 34 TMJs (17 radiographic series). Regions of interest measuring 2 in x 2 in were digitized using an 8-bit scanner with transparency adapter at 300 dpi. The images were placed into a montage of 6 images and stored as tagged image file format (TIFF), compressed at 4 levels (25:1, 50:1, 75:1, and 100:1) using a wavelet algorithm, and displayed to the observers on a computer monitor. Their observations regarding condylar faceting, sclerosis, osteophyte formation, erosion, and abnormal shape were analyzed using ROC. Kappa values were determined for relative condylar size and condylar position within the glenoid fossa. Indices were compared using ANOVA at a significance level of P < .05. Although significant and substantial observer variability was demonstrated, there were no statistically significant differences between image modalities, except for condylar position, in which TIFF and wavelet (at all compression ratios) performed better than the original image. For faceting, wavelet 100:1 performed better than radiographic film images. Little actual image file reduction was achieved at compression ratios above 25:1. PMID- 10696601 TI - The Internet as a potential source of information about radiological procedures for patients. AB - The purposes of this study were to determine what percentage of patients in a typical radiology outpatient setting own or have access to a computer with internet capabilities and how many of these patients would find an educational radiology website useful. During a 3-month period, surveys were given to all adult outpatients undergoing computed tomography. The survey asked 4 questions: (1) Do you own a computer?, (2) If you own a computer, does it have Internet access?, (3) If you do not own a computer, do you have access to a computer with Internet capabilities?, and (4) If we provide helpful information regarding preparation for and the conduct of various radiological procedures on the Internet, would you use it? Four hundred surveys were collected. Two hundred one of the respondents (50.3%) owned a computer; 189 of the 201 (94.0%) had Internet access on their computer or had access to another computer with Internet capabilities. One hundred ninety-nine of the 400 respondents (49.8%) did not own a computer, 57 of these (28.6%) had access to a computer and the Internet. Of the 246 of those with Internet access, 205 (83.3%) indicated that they would use a website that provided helpful information regarding radiological examinations. The Internet is an excellent resource for educational information for patients about various radiological procedures. This study showed that 61.5% of patients had access to the Internet, and 83.3% of these would use such a site. It is likely that these numbers will increase with the rapid growth of the Internet and the steadily increasing number of homes with computers. PMID- 10696602 TI - Sensation weighting in comparison and discrimination of heaviness. AB - Participants lifted pairs of successively presented weights and compared them for heaviness, using the constant method with 2, 3, or 6 judgment categories. The standard weight (St) was 100, 200, or 300 g, either roving or fixed within a block. For each St, there were 5 comparison (Co) weights. The lifting orders were St-Co and, with 6 categories, Co-St. Time-order errors were negatively related to St magnitude, particularly with roving St. In terms of Hellstrom's sensation weighting theory, this result was accounted for by a smaller weighting coefficient for the first-presented stimulus than for the second. Time-order errors were negative on average, which was explained as the result of this weighting in conjunction with a low position of the reference level because of light background heaviness. With roving St, the dispersion of the subjective intrapair difference increased with St magnitude, providing evidence for Ekman's law (G. Ekman, 1956, 1959). PMID- 10696603 TI - Representations of motion and direction. AB - In 6 experiments, incidental memory was tested for direction of motion in an old new recognition paradigm. Ability to recognize previously shown directions depended greatly on motion type. Memory for translation and expansion-contraction direction was highly veridical, whereas memory for rotation direction was conspicuously absent. Similar results were obtained in conditions in which motions were illustrated with pictures. Results suggest that explicit representations of direction in long-term memory are not so much related to motion per se as to the consequences of motion, the displacements of objects. Memory for all motions following circular pathways was found to be corrupted by a generic bias to regard the clockwise direction as familiar. Assessment of memory in these cases required disentangling familiarity bias for the clockwise direction from explicit recognition of direction. PMID- 10696604 TI - Perceiving circular heading in noncanonical flow fields. AB - Five experiments examined circular heading perception with optical flows that departed from the canonical form. Noncanonicity was achieved through nonrigidity of the environment (Experiments 1 and 2), oscillations of the point of observation (Experiment 3), and the bending of light (Experiments 4 and 5). In Experiments 1 and 2, perception was impaired more by nonrigidity of the ground plane than by nonrigidity of the medium. In Experiment 3, perception was unimpaired by noncanonical flows induced by the bounce and sway of observer locomotion. In Experiments 4 and 5, perception was not impaired when light paths were distorted by a spherical projection, but perception was impaired when they were distorted by a sine function. Results are discussed in relation to the hypothesis that the information for perceiving heading is the ordinal pattern of optical flow. PMID- 10696605 TI - Phonological competition and cooperation in form-related priming: sequential and nonsequential processes in word production. AB - Phonological competition theory states that competition among discrepant segments of similar words leads to inhibition of high-frequency word-naming responses in form-related priming tasks. If segments are selected sequentially, competition should be greater for begin-related pairs (storage-story), in which discrepant segments are late in the words, than for end-related pairs (glory-story), in which discrepant segments are selected before the shared ones. This pattern was not observed in standard visual priming, probably because of the influence of parallel orthographic input. However, it was observed in a repetitive word-pair production task in which visual input was absent. The findings favor a class of models in which nonsequential activation of phonological content precedes sequential selection of the segments of words to be spoken. PMID- 10696606 TI - Perceptual independence of whole length, partial length, and hand position in wielding a rod. AB - Previous investigations of dynamic touch have shown that, in wielding an occluded rod, the nonvisible perceptions of total rod length, hand position, and fractional rod length above or below the hand are different functions of the eigenvalues and eigenvectors of the inertia tensor. The implication that the 3 perceptions are independent covariants of the inertia tensor of a wielded object was tested with the complete identification experimental procedure using the statistical prescriptions of F. G. Ashby and J. T. Townsend (1986). The confirmed independence is discussed in the context of the generalized psychological or perception-information hypothesis. PMID- 10696607 TI - Unitization during category learning. AB - Five experiments explored the question of whether new perceptual units can be developed if they are useful for a category learning task, and if so, what the constraints on this unitization process are. During category learning, participants were required to attend either a single component or a conjunction of 5 components. Consistent with unitization, the conjunctive task became much easier with practice; this improvement was not found for the single-component task or for conjunctive tasks in which the components could not be unitized. Influences of component organization (Experiment 1), component contiguity (Experiment 2), component proximity (Experiment 3), and number of components (Experiment 4) on practice effects were found. Deconvolved response times (Experiment 5) showed that prolonged practice yielded faster responses than predicted by an analytic model that integrates evidence from independently perceived components. PMID- 10696608 TI - Asymmetries in a unilateral flanker task depend on the direction of the response: the role of attentional shift and perceptual grouping. AB - Four experiments were conducted using a flanker task with 1 distractor appearing either on the left or right side of a central target. Responses were made on a keyboard aligned parallel to the displays. A larger flanker effect was obtained when the distractor was on the same side as the response. Two factors account for this asymmetry. First, when the flanker and target are identical, the 2 form a group that is assigned a spatial tag, creating a form of the Simon effect on the basis of the compatibility between the response keys and the group. Second, preparation of a lateralized response appears to entail a shift of visual attention in the corresponding direction, thus enhancing processing of the flanker on the response side. Consistent with the 2nd hypothesis, participants were more likely to correctly recognize letters that were briefly presented at the distractor position on the same side as the response. PMID- 10696609 TI - A diffusion model account of masking in two-choice letter identification. AB - The diffusion model developed by R. Ratcliff (1978, 1981, 1985, 1988) for 2 choice decisions was applied to data from 2 letter identification experiments. In the experiments, stimulus letters were displayed and then masked, and the stimulus onset asynchrony between letter and mask was manipulated to vary accuracy from near chance to near ceiling. A standard reaction time procedure was used in one experiment and a deadline procedure in the other. Two hypotheses about the effect of masking on the information provided to the decision process were contrasted: (a) The output of perception to the decision process varies with time, so that the information used by the decision process rises and falls, reflecting the stimulus onset and mask onset. (b) The output of perception to the decision is constant over time, reflecting information integrated over the time between the stimulus and mask onsets. The data were well fit by the diffusion model only with the assumption of constant information over time. PMID- 10696610 TI - Performance monitoring in a confusing world: error-related brain activity, judgments of response accuracy, and types of errors. AB - The error-related negativity (ERN) represents a neural response, recorded from scalp electrodes, that is associated with monitoring activities. It is most likely generated in the anterior cingulate cortex (ACC). Measures of the ERN, and of behavioral and perceived accuracy, were obtained from participants while they performed a visual 2-choice reaction time task under degraded stimulus conditions. Irrespective of behavioral accuracy, the amplitude of the ERN (measured at the time of the response) covaried with the perceived inaccuracy of the behavior (measured at the end of the trial). Errors due to premature responding (errors perceived as errors) were associated with large ERNs. Errors due to data limitations (errors about which there was uncertainty) were associated with smaller ERNs. These and other results are consistent with the proposal that performance monitoring, as manifested by the ERN, involves a comparison between representations of the appropriate response and the response actually made. PMID- 10696611 TI - Bias toward regular form in mental shape spaces. AB - The distribution of figural "goodness" in 2 mental shape spaces, the space of triangles and the space of quadrilaterals, was examined. In Experiment 1, participants were asked to rate the typicality of visually presented triangles and quadrilaterals (perceptual task). In Experiment 2, participants were asked to draw triangles and quadrilaterals by hand (production task). The rated typicality of a particular shape and the probability that that shape was generated by participants were each plotted as a function of shape parameters, yielding estimates of the subjective distribution of shape goodness in shape space. Compared with neutral distributions of random shapes in the same shape spaces, these distributions showed a marked bias toward regular forms (equilateral triangles and squares). Such psychologically modal shapes apparently represent ideal forms that maximize the perceptual preference for regularity and symmetry. PMID- 10696612 TI - Effects of word frequency and spelling-to-sound regularity in naming with and without preceding lexical decision. AB - The effects of word frequency and spelling-to-sound regularity were examined using standard naming, standard lexical-decision, go/no-go naming, and go/no-go lexical-decision tasks. In both the standard and go/no-go naming tasks, tasks requiring phonological coding, a significant Frequency x Regularity interaction was observed. That is, the regularity effect was limited to low-frequency words. In the standard and go/no-go lexical-decision tasks, tasks not requiring phonological coding, no Frequency x Regularity interaction was observed. These results indicate not only that the Frequency x Regularity interaction is a product of phonological coding processes but also that these processes are similar in the standard and go/no-go naming tasks. Results are discussed in terms of the dual-route and the parallel distributed processing frameworks. PMID- 10696613 TI - The neighborhood distribution effect in visual word recognition: words with single and twin neighbors. AB - Lexical-decision tasks were used to test the role of neighborhood distribution in visual word recognition. Predictions based on the interactive activation model were generated by running simulations. The data were compared for words with 2 higher frequency neighbors that differed in their neighborhood distribution. The neighbors were "single" when they did not share a neighborhood relationship (e.g., neighbors of flanc: flanc-blanc) or "twin" when they shared a neighborhood relationship (e.g., neighbors of firme: ferme-forme). Results show a facilitatory neighborhood distribution effect on words in Experiments 1 (easy pseudowords) and 3 (difficult pseudowords and easy pseudowords) and on pseudowords in Experiment 2. These data can be accounted for in terms of lexical inhibition in the interactive activation framework. PMID- 10696614 TI - The fast and the slow of skilled bimanual rhythm production: parallel versus integrated timing. AB - Professional pianists performed 2 bimanual rhythms at a wide range of different tempos. The polyrhythmic task required the combination of 2 isochronous sequences (3 against 4) between the hands; in the syncopated rhythm task successive keystrokes formed intervals of identical (isochronous) durations. At slower tempos, pianists relied on integrated timing control merging successive intervals between the hands into a common reference frame. A timer-motor model is proposed based on the concepts of rate fluctuation and the distinction between target specification and timekeeper execution processes as a quantitative account of performance at slow tempos. At rapid rates expert pianists used hand-independent, parallel timing control. In alternative to a model based on a single central clock, findings support a model of flexible control structures with multiple timekeepers that can work in parallel to accommodate specific task constraints. PMID- 10696615 TI - Contributions of automatic and controlled processes to the analysis of hierarchical structure. AB - Three experiments provide evidence that 2 mechanisms, 1 automatic and 1 controlled, produce variations in the efficiency with which local and global forms are processed. Targets are identified faster if they appear at the same level (global or local) as the target on the previous trial. M. R. Lamb, B. London, H. M. Pond, and K. A. Whitt (1998) provided evidence that the beneficial effect of level repetition is due to an automatic process that is outside voluntary control. In the present experiments, pretrial cues informed participants as to the level of the upcoming target. Valid cues benefited performance, whereas invalid cues harmed performance relative to noninformative neutral cues. This was so even when the relation between the cue and the level it signaled was arbitrary, indicating that the cues initiated voluntary shifts of attention. The benefit associated with level repetition, however, was unaffected by the cues. These data suggest that the benefit of level repetition results from a process that is not subject to voluntary control. PMID- 10696616 TI - Not to be and then to be: visual representation of ignored unfamiliar faces. AB - Negative priming, the increase in response time and/or errors to targets previously encountered as distractors, is explained by inhibitory mechanisms that block the access of distractor representations to response systems. The processing of unfamiliar human faces was investigated using negative priming. Observers viewed a row of faces to decide whether 2 target faces were the same or different. Response latencies were longer when 1 or both targets had appeared as distractors on the immediately preceding trial--evidence that never-before seen faces are represented and require inhibition. Response latencies were shorter when face targets had appeared as distractors, either corrupted with high frequency noise or contrast inverted--evidence that representations are facilitated. Finally, response latencies remained unaltered when face targets had appeared as upside-down distractors--evidence that not all distractor representations afford response priming. The visual system indeed represents ignored unfamiliar faces, but blocks these representations only if they vie with targets for the control of action. PMID- 10696617 TI - An auditory repetition deficit under low memory load. AB - Previous studies of the auditory analogue of repetition blindness have led to different conclusions regarding the nature of the effect (e.g., N. Kanwisher & M. C. Potter, 1989; M. Miller & D. MacKay, 1994). In the present study, recall accuracy for repeated elements was examined with lists of 2 or 3 items presented dichotically under high temporal pressure. When this procedure was used, a repetition deficit in recall was obtained for both vowels (Experiment 1) and consonant-vowel syllables (Experiment 2). Further experiments demonstrated that this deficit decreases as the stimulus onset asynchrony between the 2 critical elements increases (Experiment 3) and showed that the effect also occurs for words and not just nonsense syllables (Experiment 4). In all 4 experiments, estimations of guessing biases showed that responses to unrepeated lists were not artificially favored over responses to repeated lists. PMID- 10696618 TI - Hearing shape. AB - In 4 experiments, participants listened to suspended occluded objects set into vibration by a pendular hammer. In Experiment 1, participants provided analogue measures of the heights and widths of 3 rectangular steel plates equal in area and weight. The same report method and rectangular dimensions were used in Experiment 2 with 3 plates each of steel, wood, and Plexiglas. Heights and widths were distinguished and perceived in similar proportions to the actual dimensions regardless of material composition. In Experiments 3 and 4, participants successfully identified circular, triangular, and rectangular plates of a single material (Experiment 3) and of 3 different materials (Experiment 4). Discussion focuses on the dependency of perceived dimensions on the physical properties and linear dimensions of the plates and the acoustic structure determined by the solutions to the 2-dimensional wave equation. PMID- 10696619 TI - Control of rapid aimed hand movements: the one-target advantage. AB - A series of 8 experiments examined the phenomenon that a rapid aimed hand movement is executed faster when it is performed as a single, isolated movement than when it is followed by a second movement (the 1-target advantage). Three new accounts of this effect are proposed and tested: the eye movement hypothesis, the target uncertainty hypothesis, and the movement integration hypothesis. Data are reported that corroborate the 3rd hypothesis, but not the first 2 hypotheses. According to the movement integration hypothesis, the first movement in a series is slowed because control of the second movement may overlap with execution of the first. It is shown that manipulations of target size and movement direction mediate this process and determine the presence and absence of the 1-target advantage. Possible neurophysiological mechanisms and implications for motor control theory are discussed. PMID- 10696620 TI - Human sensitivity to acoustic information from vessel filling. AB - A cylindrical vessel's fundamental resonant frequency increases as it fills with water. In Experiment 1, observers reliably identified water level rising, falling, or not changing. In Experiment 2, observers controlled filling well using only auditory information but less well than with multimodal information. Observers controlled fills to the brim better than to a drinking level, implying anticipation of fullness. In Experiment 3, blind and blindfolded sighted observers filled vessels to the brim using only auditory information. Fills tracked vessel height and flow rate well (R = .93, blind; R = .86, sighted). Experiment 4 tested sensitivity to acoustic time-to-full (TTF), analogous to optical tau. Estimated TTF to 3 fill levels at 3 rates tracked actual TTF (group R > .9; individual median R = .82). Results supported ecological perceptual theory: Changing acoustic information affords adaptive, prospective control of vessel filling. PMID- 10696621 TI - The effect of shared responsibility and competition in perceptual games: a test of a cognitive game-theoretic extension of signal-detection theory. AB - Perceptual decisions are often made in complex social settings in which distinct observers can affect each other. To address such situations, I. Erev, D. Gopher, R. Itkin, and Y. Greenshpan (1995) proposed a formal extension of signal detection theory and a descriptive modification of the extended theory. The current article presents 2 experiments that were designed to test these models in the context of repeated 2-person perceptual safety games. In both experiments, pairs of participants performed a simulation of an industrial-production process under distinct payoff rules. Each participant had to try to produce as much as possible while avoiding costly accidents. In line with the descriptive model's predictions, the results showed a slow adjustment to the incentive structure that can be approximated by a reinforcement learning process among different perceptual cutoff strategies. Providing players with prior information about the game had an initial effect but did not alter the pattern of the results. PMID- 10696622 TI - Attention to overlapping objects: detection and discrimination of luminance changes. AB - Selective attention to 1 of 2 overlapping objects was assessed in a cuing paradigm. Participants detected or identified targets that appeared in 1 of 6 possible target locations (3 on each object). Significant cuing effects for the simple detection of such targets using both reaction time and sensitivity measures of performance were found. Cuing effects were consistently greater when the participants were required to identify some aspect of the target even when the tasks (detection vs. identification) were equated for overall performance level. These differences in cuing effects between tasks were much reduced if the target locations were no longer grouped into 2 objects. It is suggested that identical stimuli can elicit differing attentional mechanisms depending on task type (rather than task difficulty) and that these mechanisms differ in the nature of the representation of the visual world. PMID- 10696623 TI - Assessing the roles of change discrimination and luminance integration: evidence for a hybrid race model of perceptual decision making in luminance discrimination. AB - Many models of perceptual processing assume that participants integrate stimulus evidence over time, for example, random walk models. This class of models is tested in a luminance discrimination paradigm in which the onsets of the stimuli are either instantaneous (stepped) or slowly ramped. The ramped portion of ramped stimuli occurs prior to the stepped stimuli onsets. Consequently, there is more luminance energy in ramped stimuli. Therefore, if participants integrate luminance energy, they should perform better to ramped stimuli. This did not occur in 4 experiments. Participants performed better to stepped stimuli than ramped stimuli in earlier foreperiods and the reverse in later foreperiods. A new model is proposed in which participants monitor both integrated luminance energy and quick temporal changes in luminance, but they do so in a serial fashion. First, participants monitor temporal changes in luminance; later, they monitor integrated luminance energy. PMID- 10696624 TI - Perceptual learning of the detection of features in X-ray images: a functional role for improvements in adults' visual sensitivity? AB - Recent perceptual learning research has found long-term increases in the sensitivity of adults' perceptual systems. The authors examined whether such changes could partly explain improvement on tasks such as perception of medical X ray images. Experiment 1 found experts' sensitivity to low-contrast dots in X rays was better than novices'. Experiment 2 found a direction of luminance contrast-specific improvement in novices' detection of low-contrast dots in X rays as a result of practice. Experiment 3 found a partly specific improvement in novices' detection of low-contrast features in real medical X-rays as a result of practice. Results suggest that experience enhances sensitivity to the critical dimensions of visual analysis for detecting abnormalities in X-ray images. Importantly, they demonstrate a real-world adaptive functional role for the long term flexibility of sensory systems in adulthood. PMID- 10696625 TI - Response selection processes for conjunctive targets. AB - A model is proposed for identification and response selection of cross dimensional conjunctive stimuli. The model assumes that the formation of conjunction representations involves processes similar to those used in response selection for single-feature targets. It predicts that discrimination between conjunctive targets leads to separate competitions in each of the relevant component dimensions and that detection of a predefined single conjunctive target is done at the conjunctive map level. Experiments 1 and 2 support these two sets of predictions. Experiment 3 demonstrates that responses to conjunctions of features within the orientation dimension are qualitatively different from those for cross-dimensional conjunctive targets. It is speculated that line-orientation conjunctions are handled by the visual object-recognition system, whereas cross dimensional conjunctions, as exemplified by the model, may be performed by a different system that is closely associated with response selection processes. PMID- 10696626 TI - Separate features versus one principle: comment on Shimaya (1997). AB - In his article "Perception of Complex Line Drawings," A. Shimaya (1997) proposed a quantitative theory that was designed to predict perceived segmentations and amodal completions of line drawings. Shimaya further evaluated the integrative approach of structural information theory (SIT; R. Van Lier, P. Van der Helm, & E. Leeuwenberg, 1994) to pattern interpretation. It is argued in this comment that Shimaya's evaluation of the SIT approach is based on a misconception of SITs basic assumptions and an inappropriate data analysis. PMID- 10696627 TI - The Ponzo illusion affects grip-force but not grip-aperture scaling during prehension movements. AB - Contextual cues such as linear perspective and relative size can exert a powerful effect on the perception of objects. This fact is demonstrated by the illusory effects that can be induced by such cues (e.g., the Ponzo railway track and Titchener circles illusions). Several recent studies have reported, however, that visual illusions based on such cues have little or no influence on the visuomotor mechanisms used to guide hand action. Furthermore, evidence of this sort has been cited in support of a distinction between visual perception and the visual control of action. In the current study, the authors investigated the effect of the Ponzo visual illusion on the control of hand action, specifically, the scaling of grip force and grip aperture during prehension movements. The results demonstrate that grip force scaling is significantly influenced by the Ponzo visual illusion, whereas the scaling of grip aperture is unaffected by the illusion. PMID- 10696628 TI - Occlusion, symmetry, and object-based attention: comment on Behrmann, Zemel, and Mozer (1998). AB - The validity of M. Behrmann, R. Zemel, and M. Mozer's (1998) finding that object based attention can be directed toward occluded objects is examined in 3 experiments. In M. Behrmann et al.'s (1998) original study, participants made speeded judgments of whether the numbers of bumps attached to 2 arms of an X shape were the same or different. The 2 sets of bumps belonged either to a single object, 2 different objects, or 2 separated parts of an occluded object. Unfortunately, this objecthood manipulation was confounded by the symmetry of the stimuli. Experiment 1 replicated M. Behrmann et al.'s main results using identical stimuli. Experiments 2a and 2b dissociated objecthood from symmetry. The results suggest that the effects of object-based attention found by M. Behrmann et al. are largely due to symmetry. The stimuli used in M. Behrmann et al. are not appropriate for examining the relation between object-based attention and occlusion. PMID- 10696629 TI - Comparison of omega-3 fatty acids and sulfasalazine in ulcerative colitis. AB - Fish oil omega-3 fatty acids exert antiinflammatory effects on patients with ulcerative colitis. However, a comparative study in patients with mild to moderate ulcerative colitis receiving only sulfasalazine or omega-3 fatty acids has not been performed. We sought to detect changes in the inflammatory disease activity with the use of either fish oil omega-3 fatty acids or sulfasalazine in patients with ulcerative colitis. Ten patients (five male, five female; mean age = 48 +/- 12 y) with mild to moderate active ulcerative colitis were investigated in a randomized cross-over design. They received either sulfasalazine (2 g/d) or omega-3 fatty acids (5.4 g/d) for 2 m.o. Disease activity was assessed by clinical and laboratory indicators, sigmoidoscopy, histology, and whole-body protein turnover (with 15N-glycine). Treatment with omega-3 fatty acids resulted in greater disease activity as detected by a significant increase in platelet count, erythrocyte sedimentation rate, C-reactive protein, and total fecal nitrogen excretion. No major changes in protein synthesis and breakdown were observed during either treatment. In conclusion, treatment with sulfasalazine is superior to treatment with omega-3 fatty acids in patients with mild to moderate active ulcerative colitis. PMID- 10696630 TI - Correlation between lung injury score and serum albumin levels in patients at risk for developing acute lung injury. AB - This is a prospective observational study of 100 consecutive patients who were at risk for developing acute lung injury and were admitted into the surgical intensive care unit. We found a highly significant correlation between an increase in serum albumin levels and a fall in lung injury score and vice versa (r = -0.51, P = 0.000). A highly significant association was also found between mortality, fall in serum albumin levels, rise in lung injury score, and a higher simplified acute physiology score at admission (P = 0.000). PMID- 10696631 TI - Structured triacylglycerol emulsion, containing both medium- and long-chain fatty acids, in long-term home parenteral nutrition: a double-blind randomized cross over study. AB - Structured lipid emulsion, an innovative approach in which both medium-chain and long-chain fatty acids are esterified to the same glycerol backbone, has been recently shown to be a safe and efficient way of providing energy to patients requiring parenteral nutrition. As yet, no assessment has been made of its safety and effect on liver functions during long-term treatment. Twenty-two home parenteral nutrition patients with Crohn's disease or short bowel syndrome were enrolled in a double-blind randomized, cross-over study. Twenty patients who completed the study were treated for 4 wk with a structured lipid emulsion and for 4 wk with long-chain triacylglycerol emulsion. Determined every 1 or 2 wk were blood pressure, body weight, respiratory rate, blood count, liver functions, albumin, transferrin, plasma lipids, free fatty acids (FFAs), and, at the end of each treatment period (weeks 4 and 8), plasma dicarboxylic acids and 3-OH-fatty acids. No differences were observed between the groups or within the groups between the two treatments with respect to either clinical safety and adverse event occurrence or laboratory assessments. Plasma dicarboxylic acids and 3-OH fatty acids were similar and within normal range. No alteration of liver function occurred in any of the patients treated with the structured lipid emulsion, whereas two of the patients receiving long-chain triaclyglycerol emulsion developed abnormal liver function, which resolved after switching to the structured lipid emulsion. In conclusion, structured triacylyglycerols containing both medium- and long-chain fatty acids appear to be safe and well tolerated on a long-term basis in patients on home parenteral nutrition, and it may be associated with possible reduction in liver dysfunction. PMID- 10696632 TI - Determinants of energy intake and energy expenditure in HIV and AIDS. AB - To determine the relative importance of various factors in the causation of wasting related to human immunodeficiency virus (HIV), quantitative analysis and linear structural modeling was performed on energy metabolism data collected longitudinally and prospectively from 33 men positive for the human immunodeficiency virus at 105 time points over a 3-y period before the era of highly active antiretroviral therapy. Measured variables included energy intake, total energy expenditure, resting energy expenditure, rate of change in weight, CD4 count, clinical status, appetite, and mood. Derived variables included energy balance, activity-related energy expenditure, and physical activity level. Relative contributions were assessed by linear structural modeling based on multiple regression expressing results as path coefficients for individual relationships. The primary determinant of energy balance was energy intake (r = 0.80). Total energy expenditure made a very minor contribution to energy balance (r = -0.04). Total energy expenditure was primarily determined by activity level (r = 0.91), which itself was negatively related to the presence of opportunistic infection and CD4 count. Energy intake was related to activity level (r = 0.28) and appetite (r = 0.30), which were closely interrelated (r = 0.59). Such linear structural models allow quantitative importance to be apportioned to factors determining weight change in those infected with HIV and represent a powerful tool for future metabolic studies. PMID- 10696633 TI - Effects of supplementation with folic acid and antioxidant vitamins on homocysteine levels and LDL oxidation in coronary patients. AB - Hyperhomocysteinemia is an important cardiovascular risk factor. Serum homocysteine levels are specially dependent on folate nutritional status. In addition, the oxidative modification of low-density lipoproteins (LDLs) in the endothelial microenvironment is a damaging factor that can be modified with fat soluble antioxidant vitamins. The present study was done to assess the effect of a supplementation of folic acid and antioxidant vitamins on homocysteine levels and in vitro LDL oxidation in patients with coronary artery disease. Twenty-three patients with angiographically proven coronary artery disease were given supplements for 15 d consisting of one capsule twice a day of a multivitamin preparation containing 0.65 mg folic acid, 150 mg alpha-tocopherol, 150 mg ascorbic acid, 12.5 mg beta-carotene, and 0.4 microgram vitamin B12. Serum lipids, vitamin and homocysteine levels, and in vitro LDL oxidation were measured before and after the supplementation period. During the supplementation period, serum folate levels increased from 5.0 +/- 1.5 to 10.8 +/- 3.8 ng/mL (P < 0.001), vitamin B12 increased from 317.4 +/- 130.4 to 334.5 +/- 123.8 pg/mL (P < 0.05), and alpha-tocopherol increased from 8.2 +/- 5.1 to 13.7 +/- 7.9 mg/L (P < 0.001). Serum homocysteine levels decreased from 8.7 +/- 4.3 to 6.3 +/- 2.2 mumol/L (P < 0.001). In vitro LDL oxidation decreased from 2.6 +/- 1.1 to 1.6 +/- 1.1 nmol malondialdehyde/mg protein (P < 0.001). In comparing patients with healthy controls, basal levels of folate were lower in the patients, whereas vitamin B12, alpha-tocopherol, and homocysteine levels were similar. No changes in serum lipid levels or body weight were observed. In conclusion, a short-term supplementation with folic acid and antioxidant vitamins can reduce serum homocysteine levels and in vitro LDL oxidation in patients with coronary artery disease. PMID- 10696634 TI - Antioxidant status in vegetarians versus omnivores. AB - Every day, vegetarians consume many carbohydrate-rich plant foods such as fruits and vegetables, cereals, pulses, and nuts. As a consequence, their diet contains more antioxidant vitamins (vitamin C, vitamin E, and beta-carotene) and copper than that of omnivores. Intake of zinc is generally comparable to that by omnivores. However, the bioavailability of zinc in vegetarian diets is generally lower than that of omnivores. Dietary intake of selenium is variable in both groups and depends on the selenium content of the soil. Measurements of antioxidant body levels in vegetarians show that a vegetarian diet maintains higher antioxidant vitamin status (vitamin C, vitamin E, beta-carotene) but variable antioxidant trace element status as compared with an omnivorous diet. To evaluate the antioxidative potential of a vegetarian diet versus an omnivorous diet, more studies are needed in which the total antioxidant capacity is determined rather than the status of a single antioxidant nutrient. PMID- 10696635 TI - Altered tissue electric properties in lung cancer patients as detected by bioelectric impedance vector analysis. AB - Modifications of body composition are frequent in cancer patients. Bioelectric impedance analysis can specifically detect changes in tissue electric properties, which may be associated with outcome. We evaluated the distribution of the impedance vectors from 63 adult male patients with lung cancer, stages IIIB (33 patients) and IV (30 patients), in supportive therapy. Body weight change over the previous 6 m.o. was the same in both groups (stable/increased 36% and decreased in 62%). Patients were compared with 56 healthy subjects matched for gender, age, and body mass index (25 kg/m2). Impedance measurements (standard tetrapolar electrode placement on the hand and foot) were made with 50-kHz alternating currents. The resistance and reactance of the vector components were standardized by the height of the subjects and were plotted as resistance/reactance graphs. The impedance vector distribution was the same in patients with either stage IIIB or IV cancer. The mean vector position differed significantly between cancer patients and control subjects (Hotelling T2 test, P < 0.01) because of a reduced reactance component (i.e., a smaller phase angle) with preserved resistance component in both cancer groups. Patients with a phase angle smaller than 4.5 degrees had a significantly shorter, i.e., 18 m.o., survival. Body weight loss was not significantly associated with survival. In conclusion, impedance vectors from lung cancer patients were characterized by a reduced reactance component. The altered tissue electric properties were more predictive than weight loss of prognosis. PMID- 10696636 TI - Effects of parenteral nutrition supplemented with glutamine or glutamine dipeptides on liver antioxidant and detoxication systems in rats. AB - Our aim was to determine the effects of glutamine or alanyl glutamine parenteral supplementation on the liver oxidant/antioxidant balance and on cytochrome-P450 mediated detoxication in rats. Animals were infused for 5 d with standard total parenteral nutrition (TPN), glutamine-enriched TPN, or alanyl glutamine-enriched TPN. The hepatic concentration of glutathione was reduced, and the levels of thiobarbituric-acid-reactive substances (TBARS) were increased in animals receiving standard TPN. Both glutamine and alanyl glutamine supplementation normalized glutathione, but thiobarbituric-acid-reactive substance concentration was only decreased by ananyl glutamine. This effect was parallel to a partial recovery of the activity of antioxidant enzymes. Cytochrome-P450 liver content, cytochrome-P450-dependent monooxygenases, and antipyrine clearance were not modified by glutamine or alanyl glutamine. Our data suggest a better protection against free radicals by alanyl glutamine supplementation and an absence of effects of both glutamine and alanyl glutamine on liver oxidative metabolism. PMID- 10696637 TI - Extended indications for enteral nutritional support. AB - With the availability of an organized nutrition support team (NST), the use of enteral nutrition (EN) can be extended to patients who would have otherwise received parenteral nutrition (PN). Although the formulation and accuracy of nutrient intake seems easier with PN, it is accepted that EN is cost effective and advantageous and should be used whenever possible. We discuss three cases in which the NST was initially consulted to provide PN. After evaluation by the dietitian, gastrointestinal access was obtained by the NST physician and EN could be initiated. The case studies show that an organized NST consisting of knowledgeable members can extend the use of EN to patients who would have otherwise received PN. The cost saving of such an approach and its efficacy in clinical practice are obvious. PMID- 10696638 TI - Assessment of protein energy malnutrition in older persons, Part II: Laboratory evaluation. AB - A large proportion of chronic diseases affecting older persons can be either prevented or significantly improved by improving nutrition. This places an increased burden on health care professionals caring for older persons. Screening for malnutrition at an early stage allows the intervention to be most successful. History, physical examination, and anthropometric measurements are essential parts of any nutritional evaluation. However, these tools can be highly subjective and rely heavily on the knowledge and experience of the evaluator. Incorporating biochemical measurements in the routine nutritional assessment provides an often-needed objective dimension. Interpreting these measurements must take into consideration the normal biological changes seen with aging. In this article, we review many of the biochemical parameters used in nutritional assessment and their relation to morbidity and mortality, with a special focus on normal changes seen with aging. PMID- 10696639 TI - Can nutritional intervention reduce the incidence of pressure sores? PMID- 10696640 TI - Is leptin involved in the acute anorectic effect of nicotine? PMID- 10696641 TI - Effect of individual fatty acids of omega-6 and omega-3 type on human immune status and role of eicosanoids. PMID- 10696642 TI - Breast milk calcium and phosphorus concentrations. PMID- 10696643 TI - Nutrient intakes and iron status of vegetarians. PMID- 10696644 TI - Not fade away--the glycemic index. PMID- 10696645 TI - Tips on pulmonary aspiration in gastric enteral nutrition. PMID- 10696646 TI - How to provide nutritional support. PMID- 10696647 TI - Actual carcinogens and promoters involved in colon cancer causation and development. PMID- 10696648 TI - [Cerebral activation during movement of both hands. A study with transcranial Doppler]. AB - INTRODUCTION AND OBJECTIVE: In healthy persons, the force carried out by a group of muscles doing bilateral exercise with maximum effort is less than that done during unilateral exercise. The nervous control of movement is probably different in these two cases. Our objective was to study and compare cerebral activation on movement of one and of both hands by means of transcranial Doppler (TCD). MATERIAL AND METHODS: We studied 30 healthy volunteers (19 men and 11 women; average age 65.4 +/- 9.5 years). Using transtemporal TCD we assessed the relative changes in average velocity of flow in both middle cerebral arteries during the exercise of sequential opposition of the fingers of one hand and of both hands. RESULTS: The activity due to the exercise of the hand contralateral to the hemisphere being studied was greater than that due to exercise of both hands together, both on the right side (p < 0.001) and on the left (p < 0.001). CONCLUSIONS: The functional activity of each cerebral hemisphere is not necessarily greater when both hands are exercised than when the contralateral hand is used, and may even be less. The possible increase in activity due to the additional contribution to ipsilateral movement in the first case may be compensated by simultaneous transhemispherical inhibition. PMID- 10696649 TI - [Cocaine and cerebrovascular disease in young adults]. AB - INTRODUCTION AND OBJECTIVE: The use of cocaine has been increasingly associated with cerebrovascular disease specially in young adults. We review the cases of stroke related to cocaine abuse in this group. PATIENTS AND METHODS: We performed a retrospective study between 1989-1998. Data were obtained from the Young Adults Stroke Registry. To investigate the etiology of stroke all patients underwent cardiologic examination, coagulation and neuroimaging tests. RESULTS: We identified 13 patients under 45 years of age with stroke related to cocaine abuse (0.39% of all strokes and 7.60% of the ones in young adults). Mean age in this group was 28.30 years. Eight developed ischemic manifestations (5 infarcts, 2 TIAs and 1 encephalopathy with multiple ischemic lesions), 4 had intraparenchymal hemorrhages and 1 had a subarachnoid hemorrhage. The principal route of administration was intranasal and the time course from cocaine use to stroke ranged from several hours to several years. 61.53% had history of other drug abuse and in 84.61% other risk factors were identified. Angiographic studies demonstrated: arterial occlusions (3 cases), changes consistent with vasospasm (1), segmental narrowing (1) and arterial wall irregularities (1). No aneurysms or arterio-venous malformations were found. The frequency of cocaine-related stroke in young adults has decreased from 8.33% in 1989 to 5% in 1998. CONCLUSIONS: Cocaine is a well known cause of stroke, specially in young adults. In most cases other risk factors can be identified. Multiple overlapping mechanisms may be involved (vasospasm-thrombosis, high blood pressure, embolism. PMID- 10696650 TI - [The coin test]. AB - INTRODUCTION: Normal persons, whatever their level of education, have innate basic mathematical ability, but this is very sensitive to deterioration of cognitive function. The tests currently available do not permit evaluation of this ability in patients who cannot read or have little education. The coin test (CT) was designed with this in mind. OBJECTIVE: To assess the application and use of CT for diagnosis of dementia in a poorly educated population. PATIENTS AND METHODS: We studied 211 persons, 114 with dementia (criteria DSM-IV) and 97 without dementia (50 normal, 20 with deteriorated memory and 27 with cognitive deterioration), and 26 who were unable to read. CT was used in an independent, blind manner with regard to the diagnosis. We calculated the sensitivity (S), specificity (SP), positive (PPV) and negative (NPV) predictive values, correct classifications (CC) and positive likelihood ratio (RV+) for all the results obtained. RESULTS: The groups did not differ with regard to age, sex or studies. The cut off point < or = 7, S = 0.89, SP = 0.92, PPV = 0.92, NPV = 0.88 and CC = 0.90. CONCLUSIONS: The CT can be adapted to any society, is economical, ecological, valid, quick useful and well accepted. It is very sensitive and specific for the diagnosis of dementia. PMID- 10696652 TI - [Paralysis of the nucleus of the third cranial nerve secondary to a mesencephalic hematoma]. AB - INTRODUCTION: The oculomotor disorders due to mesencephalic pathology are very varied. Function of the third cranial nerve may be affected at the level of the nucleus, in the mesencephalic fascicular portion, interpeduncular fossa, pathway anterior to the posterior communicating artery, sinus cavernosus, sphenoid fissure and orbit. Paralysis of the common oculomotor nerve is the least common of these. Generally it is secondary to ischemic, or sometimes to hemorrhagic vascular pathology. CLINICAL CASE: We present the case of a hypertensive woman with an oculomotor nerve syndrome associated with limitation of horizontal gaze of the contralateral eye and reactive head inclination due to a mesencephalic haematoma. CONCLUSION: The three neuro-ophthalmological findings seen in our patient have rarely been described in the literature. We consider the clinical observation of this case and the study of the organization of the oculomotor nerve nucleus to be of interest for publication. PMID- 10696651 TI - [The efficacy of ropinirole in the treatment of chronic insomnia secondary to restless legs syndrome: polysomnography data]. AB - INTRODUCTION: Following the line of our earlier investigations, we made an open pilot study using a polysomnograph to objectively show the efficacy of Ropinirole in the treatment of chronic insomnia secondary to the restless legs syndrome (RLS). PATIENTS AND METHODS: We did polysomnograph studies on 5 patients with the characteristics of primary RLS, according to the criteria of the International Group for the study of RLS, who had had chronic insomnia for over 5 years. We used Spiegel's questionnaires to show subjective improvement. RESULTS: The following parameters were significant between the first night (without treatment) and the second night (with 0.25 mg of Ropinirole) and between night 30 without (treatment) and night 31 (with 0.25 mg of Ropinirole). The results were expressed as average values for the 5 patients with their respective standard deviations. Efficiency of sleep: 62.62% +/- 11.31 as compared with 82.82% +/- 8.80, p = 0.010. After 30 days: 64.88% +/- 17.17 against 88.75% +/- 2.09, p = 0.014. Total time asleep: 295' +/- 51.76 against 397.7 +/- 38.50, p = 0.006. After 30 days: 294.6' +/- 87.40 against 391' +/- 22.38, p = 0.025. Number of periodic movements: 91 +/- 33.59 against 30.8 +/- 35.56, p = 0.011. After 30 days: 189.8 +/- 99.79 against 36.2 +/- 21.56, p = 0.013. All patients had subjective improvement in the parameters of Spiegel's test (p = 0.0005). CONCLUSIONS: The improvement in efficiency of sleep seen on polisomnography, total time slept and reduction in periodic limb movements were statistically significant. The subjective improvement described was also confirmed by Spiegel's questionnaire. PMID- 10696653 TI - [Dysembrioplastic neuroepithelial tumors]. AB - INTRODUCTION: The dysembryoplastic neuroepithelial tumours, first defined by Dumas-Duport in 1988, are characterized histologically by being found in the cerebral cortex and having a histological pattern of multinodular architecture, foci of cortical dysplasia and a specific glioneuronal element. The clinical condition is characterized by seizures with a long evolution. These seizures are usually simple or complex partial seizures, but occasionally become generalized tonic-clonic seizures. Radiological findings on CAT or MR are cystic images localized to the cortex, with a solid component and do not cause displacement. The surgical operation required involves excision of the lesion or lesionectomy. This may be done so as to include 1 cm of the periphery of the lesion. The evolution is excellent and in most cases the seizures disappear. CLINICAL CASES: We present 4 cases of a series of 470 patients with tumours of the nervous system, operated on over the past 10 years in the Hospital del Nino Jesus. The evolution of these 4 cases has been from 1 to 5 years. In all 4 cases lesionectomy was carried out, and the evolution has been excellent (grade I of Engel's classification). CONCLUSIONS: 1. Dysembryoplastic neuroepithelial tumours are solid and cystic, situated in the cerebral cortex, with foci of cortical dysplasia. They are characterized by having a specific glioneuronal element. 2. Clinically they are characterized by crises with a long evolution. 3. The surgical operation involves lesionectomy or excision of the lesion. The evolution is excellent. PMID- 10696654 TI - [Tarsal tunnel syndorme. A report of 3 cases]. AB - INTRODUCTION: The tarsal tunnel syndrome is uncommon. It is a neuropathy of the tibial nerve at the level of the tarsal tunnel. The main symptom is pain of the sole of the foot. On percussion Tinel's sign may be found, with sensory loss all over the sole of the foot, or in parts of it, together with weakness of the intrinsic muscles of the foot. In all three cases nerve conduction studies were done and needle electromyography was also done in two cases, confirming the clinical diagnosis. CLINICAL CASES: The three patients were treated conservatively (rest, vitamin B complex and antiinflammatory analgesics). Two patients improved but one required surgical treatment when medical treatment alone was found not to be satisfactory. At the present time, all three patients are asymptomatic. CONCLUSIONS: The tarsal tunnel syndrome is uncommon but not unimportant. In most patients it presents as a syndrome of intense pain of the sole of the foot, with typical clinical features, usually unilateral. Nerve conduction studies and needle electromyography are particularly useful for confirmation of the diagnosis by localization of the problem. Initially conservative treatment is indicated in all cases. Those which do not respond to this will require surgical intervention. PMID- 10696655 TI - [Chronic eosinophilic meningoencephalitis due to Candida guillermondii]. AB - INTRODUCTION: Eosinophilic meningoencephalitis is characterized by a meningeal syndrome and an eosinophilic reaction of the cerebrospinal fluid (CSF). When these symptoms persist for more than four weeks, it is said to be chronic. Both eosinophilic meningoencephalitis and chronic meningoencephalitis are of multiple aetiologies, including fungi. CLINICAL CASE: We report a case in which chronic meningoencephalitis occurred, with persistence of leukocytes, mainly eosinophils, in the CSF. The patient complained of intense, generalized headache and a temperature of 37 degrees C. On physical examination, the only unusual finding was slight neck stiffness. During the clinical course there were focal motor signs and evidence of intracranial hypertension. On CSF culture Candida guillermondii was grown. Treatment with amphotericin B was given, and the neurological disorder remitted completely. CONCLUSION: Candida guillermondii may be the etiological agent in chronic eosinophilic meningoencephalitis. PMID- 10696656 TI - [Treatment for non-cognitive symptoms in Alzheimer's disease]. AB - OBJECTIVES: To review the classical treatment for non cognitive symptoms in Alzheimer's disease and to estimate the possible future contributions. DEVELOPMENT: The non cognitive symptoms, with a high frequency in dementia, mean a larger clinical severity, an increment of institutionalization and a larger carer's emotional burden. Several treatment frequently used has been reviewed (antipsychotic, antidepressant, antianxiety, anticonvulsive...) and the response for some of this symptoms is relatively narrow and side effects are frequent and intense. New drugs, as cholinesterase inhibitors and cholinergic agonist, that have demonstrated their efficacy for the cognitive symptoms, seem to be also effective for non cognitive ones. CONCLUSION: The relative low effectivity of classical treatment and the frequency and intensity of side effects open new possibilities to cholinesterase inhibitors in the treatment of non cognitive symptoms in Alzheimer's disease. PMID- 10696657 TI - [Neuronal changes induced by degenerative processes in the central nervous system. The influence of normal and pathological aging]. AB - INTRODUCTION: In the last years, an important effort has been made to know which are the causes of the neurodegenerative processes in the Central Nervous System and the morphological changes occurred under pathological situations and/or with normal ageing. DEVELOPMENT: Usually, neurodegenerative disorders have been associated with neuronal loss and reactive gliosis. However, the quantitative studies showed different or although contradictory results. These results are variable depending on the animal model, cerebral area or the technical procedure. However, there are other neuronal changes related with neurodegeneration. One of them is the presence of dark neurons. These neurons have been characterized by their strong staining and their structural and ultrastructural changes. CONCLUSIONS: In the present work we review about these neuronal changes in the literature, and the knowledge about the quantitative changes observed in two degenerative disorders: aging and induced cerebral lesions. In this way, we study normal and pathological neurodegenerative processes. PMID- 10696658 TI - [Symptomatic hemifacial spasm]. PMID- 10696659 TI - [From the genetics to the prevention of stroke]. AB - Genetic risk factors implicated in stroke are reviewed. There is evidence that family history of vascular disease is an independent risk factor for stroke. Twin studies demonstrated that there is a genetic component for stroke. I review the possible pathogenetic relevance of several vascular risk factors, namely dyslipoproteinemia, Lp(a), ApoE, homocystein, and prothrombotic states. Finally, I carry out an overview of genetic monogenic disorders manifesting with embolic stroke, thrombotic stroke or hemorrhagic stroke. This review corroborates that there are many genetic risk factors of stroke, though further studies will be necessary to establish whether or not these factors are pathogenetically independent from acquired factors. PMID- 10696660 TI - [Infection, atherosclerosis and acute ischemic cerebrovascular disease]. AB - The established risk factors for ischemic stroke do not sufficiently explain all clinical and epidemiological features of the disease, such as the winter peak of stroke incidence, the decline of stroke during this century and the time point of cerebral ischemia. A role of infectious disease as stroke risk factor may partly explain above features. Several case-control studies with both hospital and population control groups showed that acute infection within the preceding week and mainly respiratory infection of both viral and bacterial origin increase the risk of cerebral ischemia independent from other risk factors (odds ratio 2.9 14.5). Infection as a risk factor appears to be most important in young age groups. Infection may cause a procoagulant state and thus, trigger thrombosis and cerebral ischemia. There is increasing evidence for chronic infection as stroke risk factor. A case-control study indicated chronic and recurrent bronchitis to increase stroke risk. Two case-control and one cohort study showed that chronic dental infection, mainly parodontitis, is a risk factor for stroke. There are conflicting results on chronic infection with cytomegalovirus and insufficient evidence for a role of Helicobacter pylorii infection in pathogenesis of stroke. Seroepidemiological studies and analyses of carotid plaques indicate a role of Chlamydia pneumoniae in ischemic stroke. However, causality can not yet be inferred from present results. Acute and chronic infectious diseases are treatable and partly preventable conditions. Their recognition as stroke risk factors could therefore be important for stroke prevention. PMID- 10696661 TI - [Ultrasonography of the progression of atherosclerotic plaques]. AB - INTRODUCTION: Severe carotid stenosis is the single most important aetiological factor in focal cerebral ischaemia. Carotid endarterectomy is indicated in stenosis of greater than 80% according to the results of the study ECST. DEVELOPMENT: Since the morbimortality associated with carotid endarterectomy in patients with moderate stenosis is similar to the incidence of stroke in patients treated with drugs, different criteria should be used to quantify stenosis for the detection of patients with a high risk of having a stroke. Ultrasound study (colour duplex) permits one to not only determine the degree of carotid stenosis, based on the characteristics of the flow curve, but also to analyse the structure of the atheromatous plaque. That is to say that one can distinguish between patients with a high risk of stroke and those with a low risk, independently of the degree of carotid stenosis. The ultrasound study has not only shown great precision in identification of the different components of the plaque of atheroma described in anatomopathological studies, but has also made apparent the association between certain characteristics of the atheromatous plaque, the risk of its progression and the risk of stroke. CONCLUSIONS: In this review we consider the ultrasound characteristics of the atheromatous plaque and the risk of stroke associated with it; the different ultrasound patterns and their correlation with anatomopathological findings; we review the mechanisms of progression of atheromatous plaques and finally describe their classification according to the recommendations of the second International Consensus. PMID- 10696662 TI - [Hypolipemic treatment in the prevention of atherosclerotic plaque complications]. AB - INTRODUCTION AND OBJECTIVE: Hypercholesterolemia has been shown to be a definite risk factor for coronary disease, although its relevance in cerebrovascular disease is more controversial. This study reviews the part played by different hypolipemic treatments in primary and secondary prevention of the complications of atherothrombotic diseases, particularly stroke. DEVELOPMENT: We based our study mainly on the HMG-CoA inhibitors (3-hydroxyl 3 methyl glutaryl coenzyme A) reductase, or statins++. This group of drugs acts by inhibiting the synthesis of cholesterol and increasing the expression of LDL-c receptors, achieving a 25-35% lowering of plasma LDL-c levels. In diverse clinical trials they have been shown to have a beneficial effect in the prevention of cardiovascular disease. The results of these studies indicate that, in addition to their purely hypolipemic effect, other anti-atherothrombotic mechanisms are involved. We analyze the main studies on hypolipemic drugs in the primary and secondary prevention of coronary and cardiovascular disease. CONCLUSIONS: The role of statins is clearly defined in reduction of the risk of overall and cardiovascular mortality, and also in reduction of the incidence of cardiovascular incidents in patients with a past history of coronary disease and a cholesterol level over 155 mg/dl. Reduction of the risk from cerebrovascular disease has only been observed in primary prevention studies (patients with a past history of coronary disease). Therefore, we shall have to await the results of the clinical trials currently being carried out to determine the true role of statins in the secondary prevention of cerebrovascular disease. PMID- 10696663 TI - [Dynamics and control of atherosclerotic plaque]. PMID- 10696664 TI - [Epidemiology of multiple sclerosis in Spain. Prevalence and incidence data]. AB - INTRODUCTION: Traditionally, Spain has been considered to be an area with a low prevalence of multiple sclerosis. However, the most recent studies show much higher figures than those previously described. OBJECTIVE: We aim to summarize the current data on prevalence and incidence of the disorder in Spain so as to determine the true situation regarding multiple sclerosis in this country. DEVELOPMENT: We have evaluated all the epidemiological data published on the descriptive epidemiology of multiple sclerosis in Spain, and analyzed the differences in method which might explain the disparity between the figures quoted. We also studied the data on incidence available, mainly in the Alcoi health district, where we carried out the most prolonged incidence study in Spain. CONCLUSIONS: The figures for prevalence given by the most recent studies include Spain in the area of moderate risk. The analysis of prevalence figures seem to indicate that these have varied with time and, at least in some zones, the incidence has increased in recent years. PMID- 10696665 TI - [The epidemiology of cerebrovascular disease in Cuba]. AB - OBJECTIVE AND DEVELOPMENT: We present a synopsis of some of the main achievements of the Cuban Public Health Service in the past 39 years. We emphasize the importance of cerebrovascular disease as a cause of mortality, both in Cuba and in certain selected provinces, together with its relationship with some parameters such as age group. In the second part, we show the main results of a longitudinal or cohort study carried out in the town of Cienfuegos in Cuba. In this the importance of age as a statistically significant risk marker for the development of cerebrovascular disease is clearly seen. We also present the rate of incidence depending on exposure to risk markers, and the estimated number of new cases annually of cerebrovascular disease according to age group and gender in the town of Cienfuegos. We emphasize the need for our Public Health System to continue to deal with this disorder in an integrated manner. PMID- 10696666 TI - [Cerebrovascular mortality in Spain]. AB - INTRODUCTION: Although the mortality due to cerebrovascular disease in Spain has been considerably reduced in the past twenty years, at all ages and in both sexes, it is still the second cause of death in men and first in women. OBJECTIVE: To describe the mortality due to cerebrovascular disease in Spain. MATERIAL AND METHODS: We analyse a time series over a period of 45 years (1951 1995) by age, sex, period of death (calendar year) and cohort of birth, the geographical distribution in the last five year period available (1991-1995), and compared with other industrial countries. RESULTS: The mortality due to cerebrovascular disease in Spain has dropped over the past twenty years. This fall has accelerated over the past five years, mainly in the older age groups. Both effects, period and cohort, are seen in this drop. There is marked male predominance. The geographical distribution is in a north-south pattern. At an international level, mortality due to cerebrovascular disease in Spain is medium low, although in some provinces it is high. CONCLUSIONS: Efforts to reduce the incidence and severity of cerebrovascular disease are essential to reduce mortality. It is necessary to continue investigations as to the true impact of this group of disorders, incidence, gravity and mortality, and the distribution of cardiovascular risk factors in the whole population. PMID- 10696667 TI - [Cerebrovascular morbidity in Spain: incidence and prevalence]. AB - OBJECTIVE: A descriptive study of the morbidity due to cerebrovascular disease in Spain. METHODS: A review of the studies of the incidence and prevalence of cerebrovascular disease in Spain. A search of the literature available through the database MEDLINE. RESULTS: The annual incidence of stroke in Spain is approximately 150 per 100,000 inhabitants. The annual incidence of transient ischaemic accidents is 35-60 per 100,000 in community studies and nearly 300 per 100,000 in door-to-door studies. In the population aged over 65 years, the prevalence of stroke is 4%-8% and the prevalence of transient ischaemic accidents is 2%-3%. CONCLUSION: The incidence and prevalence figures for cerebrovascular disease in Spain are high. PMID- 10696668 TI - [Parkinson disease in Spain: evidence of under-diagnosis and starting points for its reduction]. AB - INTRODUCTION: Parkinson's disease (PD) has been found in all surveys focusing on frequent neurological disorders. In general, sparse geographical variation of PD prevalence was found. OBJECTIVE: Since preliminary data suggest considerable PD underdiagnosis, 69%, 67%, 52%, 29%, we studied potential differences of PD prevalences in Spain. MATERIAL AND METHODS: We obtained age-specific prevalences of PD from scientific media and publications from professional meetings and compared them using stratified analysis. RESULTS: The geographical variation of prevalences at ages 60-89 years was small except among those aged 80-89, being twofold in Cantalejo, Segovia, and approximately 1/3 in Lower Aragon, Teruel. The Cantalejo (door-to-door, 67% de novo diagnoses) versus Lower Aragon (medical services) prevalence ratio was 4/1. CONCLUSION: PD might be considerably underdiagnosed in Spain. PMID- 10696669 TI - [Epidemiology of primary dystonia]. AB - INTRODUCTION: Dystonia is the most difficult disorder of movement to recognize, and in which errors of diagnosis most often occur. DEVELOPMENT: We review the classification of the dystonias and the most important features of the different subdivisions. With regard to epidemiology, the exact incidence and prevalence of primary dystonia are not known. Figures vary considerably depending on the source, method of study and ethnic group of the population studied. In Europe the only study made of focal dystonias was carried out in Segovia, where the prevalence was about 300 persons per million. Finally we discuss some recent genetic findings related to the dystonias, which should stimulate more detailed studies to clarify the genetic basis of focal dystonias. PMID- 10696670 TI - [Motoneuron disease in Spain: differential epidemiological features]. AB - INTRODUCTION AND OBJECTIVES: Little data has been published in Spain on motor neurone disease or amyotrophic lateral sclerosis. This study reports data on incidence and mortality in Cantabria and incidence in Segovia. It discusses and compares them emphasizing certain characteristic features of the epidemiology of the disease in Spain. RESULTS: The incidence of motor neurone disease is higher in males, increases with age, reaches a peak between 70 and 79 years but then drops after the age of 80. There is little difference between incidence and mortality. The incidence of mortality varies greatly with time. CONCLUSIONS: The incidence of motor neurone disease in Spanish studies is average. The mortality is similar to that of other countries, although the pattern varies with age. There was a striking drop in this disease in the 1960s, but since 1970 it has been increasing. PMID- 10696671 TI - [Epidemiology of malignant tumors of the nervous system in Spain]. AB - INTRODUCTION: Increasing incidences or mortality rates from brain malignant tumors have been reported in several countries. OBJECTIVE: This is a review of the studies published by the Cancer Epidemiology Unit of Carlos III Health Institute on incidence and mortality from these tumors in adults and children in Spain. RESULTS: During the period 1952-1986 mortality in adults has increased in both sexes. An ascending effect in cohorts born up to 1920 is detected, probably due to improvements in diagnosis and registration. A positive, progressive, cohort effect in males born post-1920 was detected probably because of a true increase in incidence of brain glioma or brain metastases. The incidence analysis in Navarre and Zaragoza (two Spanish provinces with population-based Cancer Registries working for more than 20 years) shows an increase in all age groups rates, reater among the older age groups. In Navarre we detected a decrease cohort effect in 1978-82 and 1988-91. A more than 50% decrease in mortality is observed among children and adolescent, probably due to treatment improvements. Great geographic differences have been observed in mortality, associated to industry development. Incidence in children, according to the Navarre and Zaragoza Registries, has increased in part due to better diagnostic methods. Incidence is 75% greater in Navarre than in Zaragoza. PMID- 10696672 TI - [Fatigue in multiple sclerosis]. PMID- 10696673 TI - [Epilepsy associated with arteriovenous malformations]. PMID- 10696674 TI - [Anastomoses between the meningeal arteries and the cavernous sinus: the clinical picture, diagnosis and treatment]. AB - The findings of 106 patients with arteriosinus anastomoses formed by the meningeal arteries and cavernous sinus were analyzed. The clinical symptoms, the specific features of the course of this abnormality and the principle of its diagnosis are summarized. Based on angiographic data, 4 types of anastomoses were identified by X-ray and anatomic signs: 1) the anastomoses formed by the meningeal branches of the internal carotid and cavernous sinus (45.4%); 2) those formed by the branches of the external carotid and cavernous sinus (9.3%); 3) the branches of both the internal and external carotids are involved in anastomotic blood supply (38.1%); 4) a combination of carotid-cavernous and arteriosinus anastomoses (7.2%). According to the sources of blood supply, the patients received endovascular treatment (external carotid branch embolization, fistula balloon occlusion), radiation surgery or combined treatment. The differential approach to treating patients with arteriosinus anastomoses at the site of the cavernous sinus yielded good results in 78.9% of cases. PMID- 10696675 TI - [Endoscopic fenestration of median supratentorial cerebrospinal fluid cysts]. AB - Mid-supratentorial liquor cysts are a relatively rare and generally congenital abnormality of the cerebral ventricles and subdural spaces. The data and views available in the literature on rational surgical policy is contradictory. The authors' experience in treating 16 patients was used to consider whether endoscopic techniques can be employed for invasive fenestration of the cysts. The goal of surgery was to remove the masses caused by cystic malformations and their local compression of the brain via fenestration of the walls of the cysts and via communication of their cavities with the ventricles and cisterns. There were solitary cysts in all cases (arachnoidal cysts of the interpedicular cistern and the third ventricle in 9; cysts of the ventricular septum in 4, ependicular cysts of the lateral ventricle in 2, and cysts of the celiac plexus of the third ventricle in other 2 cases, in 1 cases a liquor cyst was located in the midbrain thickness). The clinical picture was characterized by a combination of hypertensive, hydrocephalic and focal symptoms of damages to the hypothalamic and thalamic structures and the adjacent formations of the brain (pyramidal and extrapyramidal disorders, ataxia, chiasmal syndrome, metabolic and endocrine disorders, etc.). In 6 cases these symptoms were persistent despite preimplanted VP anastomosis. Rigid Storz endoscopes (Germany) with an external coat, 6 mm in diameter, and a Codman fibroendoscope (USA), 4 mm in diameter, were employed. Cystic ventriculostomy and cystic ventriculocisternostomies were made in 11 and 6 patients, respectively; one patient underwent endoscopic resection of the walls of an ependymal cyst. In one patient with signs of decreased liquor resorption, endoscopic fenestration was concurrently developed into a ventricle-peritoneal anastomosis. In other 4 anastomosis-dependent patients, the preimplanted mechanically consistent bypass system was left at its site. In 2 of these cases, cystic ventriculostomy was supplemented by ventricular septal fenestration and third-ventricular bottom perforation. Twelve patients were followed up for 6 to 36.5 months (mean 15 months). There has been no information about 6 patients since their discharge. In 12 (66.5%) surgery yielded expected results and the fenestration of cystic walls was followed by their retraction and a steady-state regression of local and/or hypertensive symptoms. In 5 (28%) patients, the complaints and clinical data remained unchanged despite although incomplete but objective cystic relaxation. This was most frequently noted in patients (n = 4) with arachnoidal cysts of the interpedicular cistern and the third ventricle who had endocrine disorders. In one case the operation was stopped due to bleeding. Totally, 5 patients were found to have complications (hemorrhage, ventriculitis). None patient died. Some aspects of indications for endoscopy and surgical techniques are considered. It is concluded that endoscopic internal bypass surgery in patients wit median cystic liquor malformations is the treatment of choice. When equipment is adjusted, fenestration of the membranous walls of these cysts by using an endoscope is reliable and safe. Such patients may be recommended endoscopic technology used as the method of choice. PMID- 10696677 TI - [Our experience with surgical treatment in chronic subdural hematomas]. AB - Intracranial hematomas cause high mortality and 10-15% reoperation. A new surgical policy in this pathology is suggested. Due to the use of this method, blood flows out of hematoma cavity during the operation and postoperative period. This method allows to avoid postoperative complications. PMID- 10696676 TI - [The effect of selective dorsal rhizotomy on motor function in patients with infantile cerebral palsy]. AB - The authors analyze the outcomes of surgical treatment in 15 CP patients with lower paraparesis who had been treated with selective dorsal rhizotomy at the L2 S2 level. All the patients were examined by neurological study, EMG, EEG, visual evoked potential (VEP) recording and motor reaction time estimation. Based on the findings, it is suggested that SPR lumbosacral spinal level may affect cerebral function. Some motor functional changes are associated with this impact. Possible mechanisms of these changes are discussed. PMID- 10696678 TI - [The surgical treatment results in intracranial meningiomas over 40 years]. AB - A comparative study was made of the results of surgical treatment of 292, 290 and 349 patients with intracranial meningioma operated, respectively, in 1946-1966, 1970-1980, 1986-1996. The results were compared by radicality and postoperative lethality. The picture by these two parameters was the following in the above three periods: radicality--69.9%, postoperative lethality--17.8%; radicality- 81%, lethality--15.5%, radicality 72.5%, lethality--5.2%. It is evident that, with time, the results of the surgery were constantly improving. PMID- 10696679 TI - [The role of biochemical processes in the pathogenesis of complicated hydrocephalus in children]. AB - Spinal fluid radical formation levels, malonic dialdehyde concentrations, and intrinsic antioxidative activity were studied in 84 patients with hydrocephalus. The findings suggest that there is a considerable activation of free radical reactions and lipid peroxidation, as well as a reduction in antioxidative activity. These changes were most drastically profound in children with inflammation-complicated hydrocephalus with spinal fluid hemorrhagic changes in particular. Timely correction of impaired energy exchange in children by using antioxidants and nootropics promotes the arrest of inflammation and prevents a number of postoperative complications. PMID- 10696680 TI - [The permeability of the hemato-encephalic barrier and the proteolytic potential of the cerebrospinal fluid in severe craniocerebral trauma]. AB - To study blood-brain barrier permeability and proteolytic changes in in patients with severe brain injury and to evaluate their impact on its course and outcome, the concentrations of albumin, plasminogen (plasmin), alpha 2-macroglobulin, alpha 2-antiplasmin, and alpha 1-antitrypsin were examined in 58 victims by enzyme immunoassay. The control group comprised 20 patients examined for lumbar discal hernia. The studies indicate that early severe brain injury showed blood brain barrier dysfunction whose severity can be detected by the spinal fluid levels of albumin, plasminogen, and alpha 2-macroglobulin. Proteolytic changes in spinal fluid are determined by its albumin, plasminogen (plasmin), alpha 2 macroglobulin, alpha 2-antiplasmin, and alpha 1-antitrypsin concentrations and affect the development of secondary brain lesion and they are of practical value. PMID- 10696681 TI - [The morphological validation for the clinical use of a nerve transplantation method]. AB - The paper summarizes the results of long-term studies made by the Department of Morphology, Institute of Experimental Medicine, Russian Academy of Medical Sciences, and the data available in the literature on the problems of neurografting and histogenesis of nervous tissue in the human prenatal development. It presents data on the conditions that most favourably affect the acceptance and development of neurografts, the integration of their cell elements with the recipient's brain, and the correction of dysfunctions. Emphasis is laid on the choice of a donor's material, the modes of its treatment and insertion, on the determination of grafting sites and scope, on immunosuppression and postoperative drug provision. PMID- 10696682 TI - [The formation of true saccular aneurysms from diverticula of the cerebral arterial walls]. AB - The authors show that aneurysmic diverticula of cerebral vascular walls cam grow in size and to develop, with time, into true saccular aneurysms. Cases of aneurysmic diverticula transformation into aneurysms illustrate dynamic pattern of aneurysmogenesis and formation of this pathology during life. Feasibility of the growth and rupture of the aneurysm in intracranial hemorrhage dictates necessity of follow-up visualizations of cerebral vessels in such patients (contrast angiography, NMR angiography). PMID- 10696683 TI - The use of inhaled corticosteroids in childhood asthma. AB - In the UK around 1 in 5 children will have been diagnosed as having asthma at some stage before they reach 15 years old. The use of inhaled corticosteroids has done much to reduce morbidity in these children and current British Guidelines recommend the use of inhaled corticosteroids in all but mild childhood asthma. However, there are concerns about the long-term safety of such treatment in growing children, and the benefits and risks of their use need careful evaluation. In this article we review the place of inhaled corticosteroids in childhood asthma. PMID- 10696684 TI - Drugs in the peri-operative period: 3--Hormonal contraceptives and hormone replacement therapy. AB - Women taking combined hormonal contraceptives or hormone replacement therapy (HRT) have an increased risk of developing venous thromboembolism, compared to non-users. Surgery itself carries a risk of thromboembolism, the incidence of which varies with the nature of the procedure. In this article, the third in our series on drug therapy in the peri-operative period, we review the thromboembolic effects of oral contraceptives and HRT and consider how these affect management around the time of surgery. PMID- 10696686 TI - Clopidogrel and [symbol: see text] ticlopidine--improvements on aspirin? AB - Clopidogrel (Plavix-Sanofi Winthrop/Bristol-Myers Squibb) and [symbol: see text] ticlopidine (Ticlid-Sanofi Winthrop) are inhibitors of platelet function and are promoted as potential alternatives to aspirin. Clopidogrel is licensed for the secondary prevention of vascular events in patients with established atherosclerotic disease. The manufacturer claims that clopidogrel is "significantly more effective at reducing myocardial infarction, stroke and vascular death" compared to aspirin. Ticlopidine is licensed as an alternative to aspirin for secondary prevention of stroke and coronary complications in patients with intermittent claudication. However, in the UK, ticlopidine is more commonly used with aspirin to prevent complications following insertion of coronary stents during angioplasty. We consider whether the claims for clopidogrel and the current use of ticlopidine are justified. PMID- 10696685 TI - Deflazacort--an alternative to prednisolone? AB - Deflazacort (Calcort--Shire) is an oral corticosteroid licensed for use in adults and children. When deflazacort first became available last year, the manufacturer claimed that the drug had a lower incidence of steroid-induced unwanted effects compared with prednisolone. However, this claim was later withdrawn at the request of the Medicines Control Agency following its review of the cited data. Current promotional material claims simply that deflazacort is "a new choice of oral steroid". Here we discuss whether deflazacort offers any advantages over established corticosteroid therapy. PMID- 10696687 TI - Drugs in the peri-operative period: 1--Stopping or continuing drugs around surgery. AB - Many patients admitted to hospital for an operation will be taking medicines which affect, or are affected by, the drugs used during surgery or by the surgical procedure itself. There are few published data on which to base policy in this situation and doctors generally rely on their own experience. In a series of articles, we will review these issues. In this, our first article, we consider how the events surrounding surgery might affect prescribing and give examples of drugs that need to be stopped or continued. Future articles will cover peri operative drug management in patients: with diabetes mellitus or taking corticosteroids; taking hormone replacement therapy or oral contraceptives; taking drugs for cardiovascular disease. PMID- 10696688 TI - HIV in pregnancy and early childhood. AB - Each year, in the UK, around 300 children are born to women who are infected with HIV. In this article, we review the management of HIV infection in pregnancy and early childhood. This issue is particularly complex as it involves screening for, and the need to treat, HIV in the pregnant woman and her baby, measures to prevent mother-to-baby (vertical) transmission of infection and the use of antiretroviral drugs outside their licensed indications. PMID- 10696689 TI - Drugs in the peri-operative period: 2--Corticosteroids and therapy for diabetes mellitus. AB - In the first article in this series on drug therapy in adults in the peri operative period, we reviewed general management issues. Here we focus on the management of patients taking corticosteroids and those with diabetes mellitus. PMID- 10696690 TI - Hyaluronan or hylans for knee osteoarthritis? AB - In joints affected by osteoarthritis, the synovial fluid's capacity to lubricate and to absorb shock are typically reduced. These changes are partly due to a reduction in the size and concentration of hyaluronic acid (hyaluronan) molecules naturally present in synovial fluid. A new approach in the management of osteoarthritis of the knee is to inject hyaluronan or derivatives of this molecule (hylans) into the joint. We consider the place of two preparations: [symbol: see text]Hyalgan and Synvisc. PMID- 10696692 TI - Reducing long-term complications of type 2 diabetes. AB - Many patients with type 2 (late-onset; non-insulin-dependent) diabetes mellitus present with no symptoms, or become asymptomatic once treatment is started. Consequently, the condition is often regarded as trivial. In reality, the long term outlook is poor, with 30% of patients developing one or more clinical complications within 10 years of diagnosis. Such complications include macrovascular disease (e.g. myocardial infarction, stroke), microvascular disease (e.g. retinopathy, nephropathy) and neuropathy. In this article, we examine the role of hypoglycaemic, antihypertensive and lipid-lowering drugs in preventing these complications. PMID- 10696691 TI - [symbol: see text] Zanamivir for influenza. AB - Every winter, influenza makes many thousands of people ill, prevents them from working and places major demands on health care. In the UK, influenza causes at least 3000-4000 deaths in an average year, mainly in older people with chronic illness, and the figure can far exceed this during an epidemic year. Annual immunisation with inactivated strains of influenza virus, given before the flu season, is the main way of protecting those who would be at high risk if they became infected. [symbol: see text] Zanamivir (Relenza-GlaxoWellcome) is a new inhaled antiviral treatment, licensed for use in people aged 12 years and over who present with symptoms of influenza, when the virus is circulating in the community. The manufacturer claims that treatment with zanamivir "reduces the duration of flu by up to 40% compared with placebo" and, in trials, "resulted in a significant reduction in complications due to flu". The new National Institute for Clinical Excellence has evaluated zanamivir and advised against its use in the NHS for the present. Here, we discuss zanamivir and consider its place in the management of suspected influenza infection. PMID- 10696693 TI - Topical NSAIDs for joint disease. AB - Over 20 proprietary topical preparations of non-steroidal anti-inflammatory drugs (NSAIDs) are now available, both 'over the counter' and on prescription. They are licensed for the treatment of pain due to 'non-serious' arthritic conditions and/or soft tissue injuries. In the early 1990s, we could find little evidence to support the use of these products rather than simple analgesics or oral NSAIDs for acute soft tissue injuries, and even less for chronic inflammatory and degenerative joint diseases. Here, we reconsider the place of topical NSAIDs, concentrating on their use in chronic arthritic conditions. PMID- 10696694 TI - Fractal-based image texture analysis of trabecular bone architecture. AB - Fractal-based image analysis methods are investigated to extract textural features related to the anisotropic structure of trabecular bone from the X-ray images of cubic bone specimens. Three methods are used to quantify image textural features: power spectrum, Minkowski dimension and mean intercept length. The global fractal dimension is used to describe the overall roughness of the image texture. The anisotropic features formed by the trabeculae are characterised by a fabric ellipse, whose orientation and eccentricity reflect the textural anisotropy of the image. Tests of these methods with synthetic images of known fractal dimension show that the Minkowski dimension provides a more accurate and consistent estimation of global fractal dimension. Tests on bone x-ray (eccentricity range 0.25-0.80) images indicate that the Minkowski dimension is more sensitive to the changes in textural orientation. The results suggest that the Minkowski dimension is a better measure for characterising trabecular bone anisotropy in the x-ray images of thick specimens. PMID- 10696695 TI - Interactive image processing system for assessment of cell movement. AB - The study of cancer cell motility is considered to be important in understanding cancer metastasis. The movement behaviour of cells within clustered cell colonies is of particular interest. Changes in cell movement, area and velocity can be an indicator of cell spreading. The aim of the study is to develop and apply a computerised interactive image processing system to quantify the movement of cells within cell clusters. A semi-automatic boundary description method based on two-dimensional rendering is devised. The system is later combined with image processing methods that facilitate the relocation of the cell boundary over time; this forms a new approach to assessing cell movement. These methods are incorporated into a software system, enabling an interactive procedure to define and monitor the movement of single cells in cell clusters from digitised microscope images. Validation of the method shows a maximum error of 10% in defining the area through a cubic spline interpolation. The system is applied to analyse the movement and area of HT115 human colon cancer cells. The system provides tools for the analysis of movement, area and velocity of single cells in cancer cell colonies and may thus be of value in further understanding cancer cell motility. PMID- 10696696 TI - Dual-beam laser illuminator of fluorescence microscope for in vivo microcirculation studies. AB - A new fluorescence intravital microscope of long working distance (39 mm) has been developed for the observation of microcirculation in a wide visual field by designing a simple epi-illumination technique with dual laser beams. Cross illumination, in which a pair of laser beams is symmetrically placed on either side of the objective such that they intersect at the focal plane of the objective, was employed to produce uniform distribution of the incident light in the object plane. In vitro experiments using a fluorescein isothiocyanate dextran (FITC-dextran; molecular weight = 70,000) solution of known concentration confirmed uniform tracer excitation in a wide visual field (approximately 30 mm2), and a linear correlation between fluorescence intensity and tracer concentration (r = 0.999), ranging between 5 mumol l-1 and 25 mumol l-1. In vivo observations in the microcirculation of a hamster cheek pouch indicated that the present technique had the advantage of high contrast compared with the image obtained by bright-field transillumination. This microscope illuminator may prove useful for the evaluation of vascular permeability under physiological and inflammatory conditions, with sufficient quantitative reliability to determine tracer concentrations in all parts of the microvascular network. Furthermore, a long working distance in this technique could have considerable advantages for the application to nail-fold capillaroscopy in humans. PMID- 10696697 TI - Real-time extraction of tissue impedance model parameters for electrical impedance spectrometer. AB - This paper presents a new algorithm for real-time extraction of tissue electrical impedance model parameters from in vivo electrical impedance spectroscopic measurements. This algorithm was developed as a part of a system for muscle tissue ischemia measurements using electrical impedance spectroscopy. An iterative least square fitting method, biased with a priori knowledge of the impedance model was developed. It simultaneously uses both the real and imaginary impedance spectra to calculate tissue parameters R0, R infinity, alpha and tau. The algorithm was tested with simulated data, and during real-time in vivo ischemia experiments. Experimental results were achieved with standard deviations of sigma R0 = 0.80%, sigma R infinity = 0.84%, sigma alpha = 0.72%, and sigma tau = 1.26%. On a Pentium II based PC, the algorithm converges to within 0.1% of the results in 17 ms. The results show that the algorithm possesses excellent parameter extraction capabilities, repeatability, speed and noise rejection. PMID- 10696698 TI - Comparison of optical, electrical, and centrifugation techniques for haematocrit monitoring of dialysed patients. AB - Haematocrits were measured as a function of ultrafiltration in a simulated haemodialysis circuit using bovine blood (plasma conductivity 12 mS cm-1) and hypotonic (8.6 mS cm-1) or hypertonic (16 mS cm-1) dialysates as well as in the absence of dialysate. A comparison was made between measurements by light absorption due to haemoglobin, by impedance in the blood line at 5 kHz using Hanai's model of blood conductivity, by conductivity measurements of blood samples at 1.2 kHz using a conductimeter, by centrifugation of blood samples and by calculations using fluid conservation. The validity of Hanai's model was verified to be satisfactory by direct blood and plasma conductivity measurements. In the absence of ionic transfer the impedance device underestimated the haematocrit by 5 to 7%. This underestimation reached 18% in the case of hypertonic dialysate, but this effect can be minimised if the haematocrit necessary for calibration is measured by centrifugation after 15 min of dialysate circulation when ionic balance is achieved. It was found that the optical method monitors haemoglobin concentration rather than red cell volume changes and is not affected by osmotic red cell swelling in the case of hypotonic dialysate. It can be concluded that the light absorption technique is both more accurate and more convenient to use than impedance. PMID- 10696699 TI - A three-dimensional definition for the flexion/extension and abduction/adduction angles. AB - Flexion/extension and abduction/adduction, two major parameters for the description of joint rotations, are used to define planer anatomical orientations of body segments. These two-dimensional definitions have been used extensively in the biomechanical literature for reporting and representing both two-dimensional and three-dimensional joint rotations. Whether these traditional two-dimensional measurements represent true joint rotations in three dimensions has not been investigated. A quantitative error analysis is presented to show how large an error can be produced in the flexion/extension and abduction/adduction angles when using two-dimensional measurements to represent three-dimensional joint rotations. The results indicate that for an out-of-plane flexion the error in abduction angle measured by previous methods increases with both flexion and initial abduction angle and become very sensitive when the flexion angle exceeds 40 degrees. Although the error can be less than 2 degrees for flexion below 20 degrees when the initial abduction angle is at 30 degrees, it can be as large as 9 degrees for 60 degrees of flexion with an initial 10 degrees of abduction; nearly double the real abduction angle. Therefore, two-dimensional measurements of flexion/extension and abduction/adduction can be erroneous and overestimated for a three-dimensional joint rotation. To overcome the problem new definitions are proposed for the true flexion/extension and abduction/adduction angles as two independent parameters for three-dimensional joint rotation. PMID- 10696700 TI - Registration and geometric modelling of the spine during scoliosis surgery: a comparison study of different pre-operative reconstruction techniques and intra operative tracking systems. AB - During scoliosis instrumentation surgery, it is difficult for surgeons fully to track vertebral motion in 3D, because only the posterior elements of the spine are exposed. Different intra-operative modelling approaches are evaluated using a registration technique that matches intra-operative measurements with a 3D pre operative model of the spine. Two tracking systems (magnetic digitiser and mechanical arm) and two pre-operative reconstruction techniques (multiplanar radiography and CT scan) are sequentially combined to build four intra-operative models. Their accuracy is assessed by comparison with the pre-operative geometry. The most minimally invasive approach (multiplanar radiographic reconstruction and magnetic digitiser) has an accuracy of 5.9 mm in translation, and errors on vertebral rotations are 4.4 degrees, 6.7 degrees and 5.0 degrees in the frontal, sagittal and transverse planes, respectively. With CT scan reconstruction, the accuracy is significantly increased by about 2 mm in translation and as much as 4.5 degrees for vertebral rotations in the sagittal plane. For the mechanical arm, the accuracy is increased by less than 1 mm in translation and 1 degree for vertebral rotations. CT scan is the most accurate reconstruction technique, but its use for long spinal segments is generally not allowed because of the high radiation exposure. Multiplanar radiographic reconstruction may be an alternative solution for long spinal segments when great accuracy is not necessary. Considering the small increase in accuracy and its awkwardness, the use of the mechanical arm may not be appropriate during surgical manoeuvres. PMID- 10696701 TI - In situ measurements of skeletal muscle power output using new capacitive strain gauge. AB - Experiments are described in which a fatigue index is determined for the latissimus dorsi muscle of sheep in situ, using capacitive strain gauges. Parallel experiments for invasive and non-invasive measurements are conducted, measuring global contraction and relaxation rates and shortening duration for paced muscle. The results show that, above one pulse per burst (5 V, 100 microseconds pulsewidth), contraction rates (62 +/- 11 mm s-1) and relaxation rates (50 +/- 7 mm s-1) are constant for unloaded muscle. For one animal, fatigue testing with a 2.5 kg load at six pulses per burst shows shortening rates increasing to a maximum (80 mm s-1) after 30 s and reducing to 5 mm s-1 after 150 s. The decrease in shortening amplitude is used as a fatigue index, log displacement against time. Power output is load dependent, measuring 4.7 W kg-1 with a 2.5 kg load. There is good agreement between the invasive and non-invasive measurements, thus providing a method for monitoring changes in muscle parameters non-invasively during future pacing transformation. PMID- 10696702 TI - Improved accuracy and extended flow range for a Fleisch pneumotachograph. AB - A large linear flow range and a small instrumental dead space volume are incompatible properties for a pneumotachometer (PTM). The linearity of a Fleisch number 2 PTM is studied for flows up to 6 litre s-1 (nominal range 0-2 litre s-1) with various up- and downstream geometries. It is hypothesised that using an array of calibration factors (conductance; flow/pressure), instead of a single calibration factor over the entire flow range, could improve accuracy and also extend the applicable flow range. The conductance against pressure characteristics are calculated with a previously described weighted averaging technique based on multiple strokes from a precision syringe. A single conductance value gives stroke volume errors in the range of -5 to 3% (0-2 litre s-1) and -6 to 11% (0-6 litre s-1) for validation using the same geometry as for calibration. The pressure dependent conductance improves accuracy to within -3% and 1% independent of flow range. However, for validation using a different geometry than for calibration, errors range from -5% to +8%. The degree of non linearity varies between the geometries (range 3-15%) and is highest when using a one-directional valve upstream of the PTM and a Y-shaped connector. In conclusion, a pressure-dependent conductance improves accuracy and can also be used to extend the applicable flow range up to at least three times the nominal flow range. PMID- 10696703 TI - Effect of ambient respiratory noise on the measurement of lung sounds. AB - The effect of ambient sounds, generated during breathing, that may reach a sensor at the chest surface by transmission from mouth and nose through air in the room, rather than through the airways, lungs and chest wall, is studied. Five healthy male non-smokers, aged from 11 to 51 years, are seated in a sound-proof acoustic chamber. Ambient respiratory noise levels are modified by directing expiratory flow outside the recording chamber. Low-density gas (HeO2 = 80% helium, 20% oxygen) is used to modify airway resonances. Spectral analysis is applied to ambient noise and to respiratory sounds measured on the chest and neck. Flow gated average sound spectra are compared statistically. A prominent spectral peak around 960 Hz appears in ambient noise and over the chest and neck during expiration in all subjects. Ambient noise reduction decreases the amplitude of this peak by 20 +/- 4 dB in the room and by 6 +/- 3.6 dB over the chest. Another prominent spectral peak, around 700 Hz in adults and 880 Hz in children, shows insignificant change, i.e. a maximum reduction of 3 dB, during modifications of ambient respiratory noise. HeO2 causes an upward shift in tracheal resonances that is also seen in the anterior chest recordings. Ambient respiratory noise explains some, but not all, peaks in the spectra of expiratory lung sounds. Resonance peaks in the spectra of expiratory tracheal sounds are also apparent in the spectra of expiratory lung sounds at the anterior chest. PMID- 10696704 TI - Expert-system classification of sleep/waking states in infants. AB - This work is part of a project to develop an expert system for automated classification of the sleep/waking states in human infants; i.e. active or rapid eye-movement sleep (REM), quiet or non-REM sleep (NREM), including its four stages, indeterminate sleep (IS) and wakefulness (WA). A model to identify these states, introducing an objective formalisation in terms of the state variables characterising the recorded patterns, is presented. The following digitally recorded physiological events are taken into account to classify the sleep/waking states: predominant background activity and the existence of sleep spindles in the electro-encephalogram; existence of rapid eye movements in the electro oculogram; and chin muscle tone in the electromyogram. Methods to detect several of these parameters are described. An expert system based on artificial ganglionar lattices is used to classify the sleep/waking states, on an off-line minute-by-minute basis. Algorithms to detect patterns automatically and an expert system to recognise sleep/waking states are introduced, and several adjustments and tests using various real patients are carried out. Results show an overall performance of 96.4% agreement with the expert on validation data without artefacts, and 84.9% agreement on validation data with artefacts. Moreover, results show a significant improvement in the classification agreement due to the application of the expert system, and a discussion is carried out to justify the difficulties of matching the expert's criteria for the interpretation of characterising patterns. PMID- 10696705 TI - Assessment of spatial resolution of pace mapping when using body surface potentials. AB - Using computer simulations and statistical methods, the resolution of pace mapping when used in combination with body surface potentials was systematically investigated. In an anatomical model of the human ventricular myocardium, pre excitation sequences were initiated at 69 sites positioned along the atrioventricular (AV) ring and corresponding body surface potential maps (BSPMs) were calculated at 32 leads placed on the anterior torso. For each time after the onset of pre-excitation (every 4 ms to 40 ms) and each root-mean-square (RMS) noise level (5, 10, 20 and 50 microV), BSPMs were cros-correlated and the spatial resolution defined as the largest pacing site separation at which the differences in correlation coefficients were not statistically significant (level p > or = 0.05). The findings indicate that when random RMS noise of 5 microV was added to the simulated BSPMs, average spatial resolution over all 60 sites was at 20 ms after the onset of pre-excitation within 3.5 +/- 0.9 mm. The results provide theoretical evidence that statistical analysis of BSPMs obtained during pace mapping can offer improved means for subcentimetre identification of accessory pathways located along the AV ring. PMID- 10696707 TI - Modelling a parasystolic rhythm in a heart-transplant patient. AB - A parasystole from a heart-transplant patient is analysed using a beat-to-beat RR interval time series obtained from an electrocardiogram (ECG). The dysrhythmia, resulting from the co-existence of two pacemakers, the sinus node and an ectopic focus, presents distinctive regular patterns, with transitions from one pattern to another occurring abruptly. It is shown that the parasystolic rhythm can be simulated by a model involving two oscillators firing at fixed rates, under the restriction that neither is allowed to fire during the other's refractory period. It is found that the structure of the generated RR time series is essentially determined by the ratio of the periods of the two oscillators. In the case of a heart-transplant patient with a small heart-rate variability as a result of heart denervation, the model predicts the RR intervals with an error of less than 6% for an 80-beat sequence. From a physiological point of view, the results imply that the interaction between the two pacemakers in the heart is fairly weak, and hence the parasystole observed in the heart-transplant patient can be modelled as pure parasystole. PMID- 10696706 TI - Phase response curve based model of the SA node: simulation by two-dimensional array of pacemaker cells with randomly distributed cycle lengths. AB - A simulation of the SA node is presented, based on a 2D array (15 x 15) model of randomly distributed pacemaker cells, interacting via a phase response curve (PRC). The model involves only the basic properties that play a direct role in the determination of the SA node rhythm: intrinsic cycle length and PRC. The PRC reflects the 'type' of interaction of each pacemaker cell with the outside world (neighbouring cells, external stimulus, etc.). A major outcome of this study is the demonstration that global dynamics and the degree of 'disorder' of the SA node are strongly affected by the cycle length distribution of the model, as well as spatial inhomogeneity in the cell-to-cell 'electrical' coupling (PRC). Those factors also determine the conduction velocity throughout the SA node and may therefore be responsible for anisotropic conduction. For example, lowering the PRC parameters (d and a) by 25% increases the array activation time from 46 to 126 ms. The model also responds appropriately to a perturbation such as a vagal pulse. This pulse produces a shift of the dominant pacemaker to another site in the array and a transient lengthening of the array cycle length, for example from 312 to 355 ms. PMID- 10696708 TI - Flow dependence and time constant of the change in nitric oxide concentration measured in the vascular media. AB - It has been considered that the concentration of endothelium-derived nitric oxide (NO) in the arterial vascular wall changes in response to flow-induced shear stress. In the present study, using an NO-sensitive electrode, the aim was to directly evaluate the relationship between perfusion rate and NO concentration in the arterial vascular wall. The NO microelectrode (diameter: 100 microns) was inserted into the vascular media of isolated canine femoral arteries, and the vessel was perfused with a Krebs-Henseleit buffer solution. A flow-related change in NO concentration in the vascular media was then evaluated by changing perfusion rate. NO concentration attained a peak value with a first-order time delay by a stepwise increase in perfusion rate, and the peak-level NO concentration was linearly correlated with perfusion rate in each vessel (10-154 pA at 2.1-72.3 ml min-1; n = 7, r2 = 0.89-0.99, p < 0.03). The average time constant for an increase in NO current with a stepwise increase in perfusion rate was 24 +/- 3 s (n = 5). NO production was increased by perfusing a solution containing 1 mmol l-1 L-arginine and was attenuated by 100 mumol l-1 NG-nitro-L arginine, indicating the intactness of the endothelium, proper insertion of the NO electrode and selective detection of NO by the electrode. It is concluded that the NO microelectrode is applicable to NO measurement in the vascular media where NO controls vascular tone and that the concentration of NO in the arterial vascular media changes with perfusion rate in a rate-dependent manner as well as with a time constant of about 24 s for a stepwise increase in flow. PMID- 10696709 TI - Evaluation of Karhunen-Loeve expansion for feature selection in computer-assisted classification of bioprosthetic heart-valve status. AB - This paper analyses the performance of four different feature-selection approaches of the Karhunen-Loeve expansion (KLE) method to select the most discriminant set of features for computer-assisted classification of bioprosthetic heart-valve status. First, an evaluation test reducing the number of initial features while maintaining the performance of the original classifier is developed. Secondly, the effectiveness of the classification in a simulated practical situation where a new sample has to be classified is estimated with a validation test. Results from both tests applied to a reference database show that the most efficient feature selection and classification (> or = 97% of correct classifications (CCs)) are performed by the Kittler and Young approach. For the clinical databases, this approach provides poor classification results for simulated 'new samples' (between 50 and 69% of CCs). For both the evaluation and the validation tests, only the Heydorn and Tou approach provides classification results comparable with those of the original classifier (a difference always < or = 7%). However, the degree of feature reduction is particularly variable. The study demonstrates that the KLE feature-selection approaches are highly population-dependent. It also shows that the validation method proposed is advantageous in clinical applications where the data collection is difficult to perform. PMID- 10696710 TI - Left-ventricular pressure gradients: a computer-model simulation. AB - Both invasive left-ventricular pressure measurements and non-invasive colour M mode echographic measurements have shown the existence of intraventricular pressure gradients (IVPGs) during early filling. The mechanisms responsible for these IVPG cannot be completely explained by the experiments. Therefore a one dimensional numerical model is developed and validated. The model describes filling (both velocities and pressures) along a left ventricular (LV) base-apex axis. Blood-wall interaction in the left ventricle with moving boundaries is taken into account. The computational results for a canine heart indicate that the observed IVPGs during filling are the consequence of a complex interaction between, on the one hand, pressure waves travelling in the LV and, on the other hand, LV geometry, relaxation and compliance. The computational results indicate the pressure dependency of wavespeed (0.77-1.90 m-1 s) for different mean intraventricular pressures (0.88-5.00 mmHg) and IVPGs up to 2 mmHg, independent of the ratio of end systolic volume and equilibrium volume. Increasing relaxation rate not only decreases minimum basal pressure (2.8 instead of 3.6 mmHg) but also has a strong influence on the time delay between the minimum basal and apical pressures (14 ms instead of 49 ms). The results sustain the hypothesis that pressure-wave propagation determines IVPGs and that IVPGs are no proof of elastic recoil. PMID- 10696711 TI - Development and preliminary clinical tests of an impedance sensing VDD recording pacemaker for diagnosis and research. AB - Episodes of serious but infrequently occurring cardiac arrhythmias can be difficult to detect and analyse, even with modern Holter monitoring. A previous diagnostic pacemaker developed by this group provided VVI pacing therapy and recorded intracardiac ECG signals but had no atrial sensing or impedance measuring capability. A new external diagnostic pacemaker system is described that has been developed to assist in diagnosing intermittent arrhythmias by selectively recording intracardiac signals. Unlike other ambulatory monitors, in addition to recording ECG, the device combines VDD pacing therapy with the capability of monitoring and recording intracardiac impedance and pressure waveforms through a temporary intracardiac lead. A PCMCIA memory card allows storage of 48 arrhythmic events of 21 seconds each. Twelve seconds of waveform are retained before the event occurs and nine seconds after. Data retrieval and processing is performed with a PC which reconstructs each waveform for display. The ECG provides data on cardiac rhythm while cardiac function is inferred from the haemodynamic signals. During simulated trials, 14 event types were presented to the system. All events were successfully detected and recorded. During in vivo clinical tests 83 waveform recordings were made. Impedance fluctuations with typical peak-to-peak values of 64 ohms were successfully recorded. PMID- 10696712 TI - Mechanism for polarisation of cardiac tissue at a sealed boundary. AB - If current is flowing in cardiac tissue, and if the myocardial fibres approach a sealed boundary at an angle, then the tissue within a few length constants of the boundary is polarised. This polarisation occurs when the cardiac tissue has different anisotropy ratios in the intracellular and extracellular spaces. This new mechanism of tissue polarisation is demonstrated using a simple, analytical model, and it is shown quantitatively that this polarisation can be nearly as large as that occurring near an electrode. PMID- 10696713 TI - Instrumented staircase for ground reaction measurement. AB - A staircase was developed to record ground reactions during stair climbing at different slopes (inclinations). Each step is instrumented with six strain-gauge based force transducers which allow the measurement of three-dimensional ground reaction force and moment as well as the centre of pressure (COP) location. A specific sensor arrangement permits accurate recording, especially of the COP location. The overall design of the staircase and details of a single instrumented step are presented. Static and dynamic characteristics have been evaluated by different experimental procedures. Preliminary results of ground reaction forces are shown. PMID- 10696714 TI - Atomic force microscopic measurement of the mechanical properties of intact endothelial cells in fresh arteries. AB - Mechanical properties of living endothelial cells in the abdominal aortas and in the medial and lateral wall of aortic bifurcations obtained from rabbits were determined by means of an atomic force microscope (AFM), focusing on the locational differences. Force (F)-indentation (delta) curves of the cells were expressed by an exponential function: F = a(exp(b delta)-1), where a and b are constants. The parameters b and c(= ab) represent the rate of modulus change and initial modulus, respectively. The slope of F-delta curves a and the parameter c were higher in the medial wall than in the other sites, which is attributable to abundant stress fibres in endothelial cells in the medial wall. There were no differences in the parameter b among the three locations. These results indicate that endothelial cells are stiffer in the medial wall of aortic bifurcation than in the other regions. PMID- 10696715 TI - A non-parametric method for the analysis of experimental tumour growth data. AB - Analysis of tumour growth is required to investigate the biology of tumours and to determine the effects of new anti-tumour therapies. A non-parametric mathematical method for the analysis of a set of experimental tumour growth data is described. The method is based on the similarity between time series of tumour size measurements (e.g. tumour volume), similarity being defined as the Euclidean distance between data measured for each tumour at the same time. Subsets of similar time series are found for a given population of tumours. A biologically meaningful parameter H has been derived which is a measure of the scattering of experimental volume samples. The method has been applied to the analysis of the growth of (i) untreated multicellular tumour spheroids obtained with different cell lines and (ii) spheroids treated with cytotoxic drugs (immunotoxins). Results are compared with those previously obtained by applying the classical Gompertz growth model to the analysis of treated and untreated spheroids. PMID- 10696716 TI - Correlation between concentration in urine and in blood of cadmium and lead among women in Asia. AB - The objectives of the present study are to examine if there exists a quantitative relationship between lead in urine (Pb-U) and that in blood (Pb-B), and also between cadmium in urine (Cd-U) and that in blood (Cd-B) among the general populations who are environmentally (and not occupationally) exposed to these elements at various levels. For this purpose, peripheral blood and morning spot urine samples were collected in 1991-1998 from approximately 50 non-smoking adult women each in four cities in south-east Asia and five cities in mainland China, and two cities each in Japan and Korea. The samples were wet-ashed and then analyzed by inductively-coupled plasma mass spectrometry for Pb-B, Cd-B, Pb-U and Cd-U. Measured values were subjected to analysis to detect possible correlation between the pairs of parameters. A significant correlation between Pb-B and Pb-U was observed when the intensity of Pb exposure (as expressed by Pb-B) was relatively high so that the correlation was significant in all cases studied when Pb-B level was, e.g. 50 microg/l or above. It was also observed that the correlation between Cd-B and Cd-U was significant when Cd-B was, e.g. > 1 microg/l. Thus, it is possible to deduce that, in environmental health, Pb-B and Cd-U levels can be estimated on a group basis from Pb-U and Cd-B, respectively, when Pb and Cd exposure levels are relatively high, e.g. with Pb-B and Cd-B of > 50 microg/l and > 1 microg/l. PMID- 10696717 TI - A comparison of heavy metal levels in the kidneys of High Arctic and mainland caribou populations in the Northwest Territories of Canada. AB - Aluminum, nickel, cadmium, mercury and lead levels were measured in the kidney tissue of Banks Island Peary caribou and barren-ground caribou, from the Bluenose herd, of the western Northwest Territories of Canada. Cadmium concentrations of Bluenose caribou were similar to those reported elsewhere for barren-ground caribou and showed a positive correlation with age. Cadmium concentrations of Peary caribou were significantly lower than those of Bluenose caribou regardless of age, were the lowest reported for caribou during winter, and did not show a positive correlation with age. Mercury levels, expressed on a wet weight basis, were similar to those reported for other barren-ground caribou. Mercury levels were significantly higher in Bluenose [mean 10.45 microg g(-1) (dry wt.); S.E.= 0.85; n = 20] than Peary [mean 5.43 microg g(-1) (dry wt.); S.E. = 0.31; n = 20] caribou. Aluminum concentrations for Bluenose and Peary caribou were similar [mean 1.48 microg g(-1) (dry wt.); S.E. = 0.17; n = 20 and mean 1.56 microg g(-1) (dry wt.); S.E.= 0.15; n = 20, respectively), but were considerably lower than those reported for barren-ground caribou elsewhere. Lead and nickel concentrations were low and similar between Bluenose, Peary and other reported barren-ground caribou populations. Higher cadmium and mercury concentrations in Bluenose caribou are consistent with the hypothesis that caribou with a high dietary lichen component have higher contaminant levels. It is unlikely that subsistence harvesters would consume enough kidney during a year to exceed the tolerable intake of cadmium recommended by the WHO. PMID- 10696718 TI - Benthic fluxes of cadmium, lead, copper and nitrogen species in the northern Adriatic Sea in front of the River Po outflow, Italy. AB - Trace heavy metal (Cd, Pb and Cu) and nitrogen species (N-NO3, N-NO2 and N-NH4) fluxes between sediment and water were examined for approximately 4 days, in a coastal marine station located in the northern Adriatic Sea in front of the River Po outflow. An in situ benthic chamber, equipped with electronic devices for monitoring and adjustment of oxygen and pH and with a temperature detector, was used. The benthic chamber experiment enabled study of the temporal trend of metals and nutrients when oxygen concentration varied in a controlled environment. Although particular care was devoted to chamber deposition and parameter control, sediment resuspension occurred at the beginning of the experiment and O2 fluctuations were observed during the course of the experiment. Pb concentration was affected by both resuspension and oxic conditions in bottom water, which prevented determination of any reasonable Pb flux value. Cd and Cu, not influenced by oxygen fluctuations, reached an equilibrium phase in a short period with initial positive fluxes from sediment of 0.68 (S.D. = 0.07) and 6.9 (S.D. = 5.6) pmol cm(-2) h(-1), respectively. With regard to nitrogen species, the highest positive flux was that of N-NH4 (10.5, S.D. = 2.4, nmol cm(-2) h(-1)) whose concentration increased in the chamber, while nitrate concentration (initial flux of -5.7, S.D. = 1.5, nmol cm(-2) h(-1)) immediately decreased after the beginning of the experiment. Nitrite concentration was almost constant throughout the experiment and its flux was generally low (initial flux 0.1, S.D. = 0.9, nmol cm(-2) h(-1)). PMID- 10696719 TI - An assessment of indoor air contaminants in buildings with recreational activity. AB - Indoor air quality measurements were carried out during three concerts and one ice hockey game in three different halls. Gas phase components consisted of CO2, CO, and NO whereas for particulate indicators, measurements of particle mass distributions (0.05-9 microm), particle number distributions (0.75-10 microm), and particle bound polycyclic aromatic hydrocarbons (pPAH) were carried out. The calculated ventilation rates did not meet the ventilation requirements for rooms with occupants who smoke to be perceived as acceptable by 80% of the occupants. Average PM9 (mass of particulate matter with an aerodynamic diameter < 9 microm) concentrations throughout the events ranged from 318 to 2000 microg m(-3). Particle concentrations in the size range < 0.4 microm measured 203-696 microg m( 3), the majority of it being attributed to environmental tobacco smoke (ETS). For particle numbers > 0.75 microm concentrations ranged from 2 x 10(4) to 1.9 x 10(5) particles per l while for pPAH, concentrations from 336 to 990 ng m(-3) were observed. The average event concentrations for the gaseous component CO2 ranged from 1110 to 1700 ppm, for CO 2-3.1 ppm and for NOx 237 ppb. The event to baseline concentration ratios for gaseous components ranged from 1.1 to 4.3 while for particulate indicators generally much greater ratios between 0.7 and 140 were found. Possible health effects inflicted by an exposure based on the measured concentrations of the various parameters are discussed. PMID- 10696720 TI - A new disease-burden method for estimating the impact of outdoor air quality on human health. AB - Urban air quality is a serious problem, with an estimated 40 million people in Europe exposed to exceedences of existing WHO air-quality guidelines, with prospects of further declines in air quality due to projected growth in motor vehicle traffic. Air-quality management strategies, underpinned by legislation are attempting to combat this problem. To support such strategies, assessment of the costs and benefits of remedial measures is required, including an assessment of the impact of urban air quality on human health. This paper describes a disease burden estimation approach, developed to assess 'health gain' from recreational water quality improvement, and its application to urban air quality and incidence of respiratory disease. The method represents an improvement over existing disease-burden estimation techniques applied to air quality, in that by considering the probability density function of pollutant concentrations, improved estimates of exposure and hence disease burden, and also 'health gain' from air-quality improvement, are possible. Estimations of mortality advanced by fine particulate matter (PM10) are presented for five UK cities. Implications of the method for disease burden and air-quality standards are discussed. The utility of integrating the disease-burden assessment model with linked dynamic models of land-use, vehicle movement and pollutant dispersion, as a means to identify remedial strategic planning initiatives, is highlighted. PMID- 10696721 TI - Evaluation on speciation and removal efficiencies of mercury from municipal solid waste incinerators in Taiwan. AB - Two large-scale municipal solid waste incinerators (MWIs) located in Taiwan were selected for conducting flue gas sampling to determine the chemical speciation of mercury by both USEPA Method 29 and Ontario Hydro Method (OHM). In addition, the emission characteristics and removal efficiencies of mercury were evaluated via isokinetic sampling of flue gas upstream and downstream of APCDs. Results indicated that the average removal efficiencies of Hg for MWI-A and MWI-B were 29.56 and 44.70%, respectively. In terms of mercury speciation by USEPA Method 29 and Ontario Hydro Method (OHM), oxidized mercury (Hg2+), in the flue gas was predominant at the inlet of APCD for both incinerators. Less than 30% of mercury in the flue gas existed in the elemental form (Hg0) at APCD inlet. Mercury emitted from the stack also predominated as a form of Hg2+ in MWI-A. Approximately 90% of total mercury emission from the stack existed in the form of Hg2+ for MWI-A. Due to the higher removal efficiency of soluble mercury (Hg2+) in wet scrubber, less total Hg was actually emitted from MWI-B than MWI-A. Regarding the removal efficiency of Hg0 in the flue gas, the APCDs of MWI-A (DSI + FF) had a higher removal efficiency than that of MWI-B (ESP + WS) possibly due to the reduction of Hg2+ which occurred in the wet scrubber. PMID- 10696722 TI - Evaluation of different approaches for modeling effects of acid rain on soils in China. AB - Acid deposition is an environmental problem of increasing concern in China. Acidic soils are common in the southern part of the country and soil acidification caused by acid deposition is expected to occur. Here we test and apply two different approaches for modeling effects of acid deposition and compare results with observed data from sites throughout southern China. The dynamic model MAGIC indicates that, during the last few decades, soil acidification rates have increased considerably due to acid deposition. This acidification will continue if sulfur deposition is not reduced, and if reduced more rapidly than base cation deposition. With the Steady State Mass Balance model (SSMB), and assuming that a molar ratio of Ca2+/Al3+ < 1 in soil water is harmful to vegetation, we estimate a slight probability for exceedance of the critical load for present deposition rates. Results from both modeling approaches show a strong dependence with deposition of base cations as well as sulfur. Hence, according to the models, changes in emission control of alkaline particulate matter prior to sulfur dioxide will be detrimental to the environment. Model calculations are, however, uncertain, particularly because available data on base cation deposition fluxes are scarce, and that model formulation of aluminum chemistry does not fully reproduce observations. An effort should be made to improve our present knowledge regarding deposition fluxes. Improvements to the model are suggested. Our work indicates that the critical loads presented in the regional acid deposition assessment model RAINS Asia are too stringent. We find weaknesses in the SSMB approach, developed for northern European conditions, when applying it to Chinese conditions. We suggest an improved effort to revise the risk parameters for use in critical load estimates in China. PMID- 10696723 TI - PCB and chlorinated pesticide concentrations in swine and bovine adipose tissue in Sweden 1991-1997: spatial and temporal trends. AB - Results from the Swedish control programme regarding organochlorines in food were used to determine time trends of organochlorine concentrations in adipose tissues from swine (4-8 months old) and bovines (non-dairy, 12-36 months) slaughtered between 1991 and 1997. Moreover, possible regional differences in concentrations were studied, as well as differences in concentrations depending on sex and age of the slaughtered animals. Multiple linear regression indicated that the concentrations of PCB, p,p'-DDE, HCB and alpha-HCH decreased by 4-17% per year, suggesting that the decline in organochlorine concentrations in the Swedish environment and biota reported during the 1970s-1990s also has occurred in meat producing animals during the 1990s. The concentrations of PCB, DDE and HCB in bovines and PCB and DDE in swine were 1.4-3.8-fold higher in the southern parts of Sweden than in the northern parts of the country, indicating a regional difference in exposure of the animals. The organochlorine concentrations were higher in bovines than in swine, and declined faster in swine than in bovines. Moreover, the concentrations of CB 153 and p,p'-DDE were similar in bovines, but in swine the average concentrations of the two compounds differed two-fold. Apart from possible species differences in metabolism of organochlorines, this may be due to differences in the age at slaughter between swine and bovines, and differences in husbandry of the animals. In the latter case, swine are generally kept inside during their whole life span, whereas bovines are kept outside grazing during the summer period. Finally, a sex-dependent difference in concentrations was indicated in swine, but not in bovines. Our study shows that a lot of information can be 'extracted' from control program results. PMID- 10696724 TI - Particles and vegetation: implications for the transfer of particle-bound organic contaminants to vegetation. AB - This paper presents a comprehensive review of the mechanisms responsible for the transfer of atmospheric particulate deposition and soil particulate re-suspension onto vegetation. The nature of atmospheric aerosols and dry/wet particulate deposition are reviewed, together with information from the literature on radionuclides as tracers of the air particle/soil particle to vegetation transfer processes. Information from these fields is used to make inferences about the potential significance of these pathways in supplying particle-bound semi volatile organic chemicals (e.g. polycyclic aromatic hydrocarbons, polychlorinated dibenzo-p-dioxins and dibenzofurans, polychlorinated biphenyls) to vegetation. Retention of compounds on particles brought to the above-ground plant surfaces is discussed. In the absence of definitive field/experimental studies, calculations are made drawing on the literature data to estimate the contributions of atmospheric and soil particle-bound organic contaminants to the plant concentration. These show that depending on the site-specific, species specific and compound-specific scenarios considered, particulate-bound inputs may be negligible or may dominate the supply of organic contaminants to the above ground portion of plants. However, field/experimental studies and direct measurements are needed to provide reliable quantitative data on this topic. PMID- 10696725 TI - Arsenic, cadmium, lead, copper and zinc in cattle from Galicia, NW Spain. AB - Knowledge of trace and toxic metal concentrations in livestock is important for assessing the effects of pollutants on domestic animals and contaminant intakes by humans. Metal levels in cattle have been measured in various countries but not in Spain. In this study, the (wet wt.) concentrations of three toxic elements (arsenic, cadmium, lead) and two trace elements (copper, zinc) were quantified in the liver (Li), kidney (Ki), muscle (M) and blood (Bl) of calves (males and females between 6 and 10 months old) and cows (2-16 years old) from Galicia, NW Spain. For the toxic elements, geometric mean concentrations of arsenic in calves (sexes combined) and cows were 10.8 and 10.2 microg/kg (Li), 11.3 and 15.2 microg/kg (Ki), 3.75 and 4.25 microg/kg (M), 3.23 and 2.92 microg/l (Bl). The corresponding cadmium concentrations were 7.78 and 83.3 microg/kg (Li), 54.3 and 388 microg/kg (Ki), 0.839 and 0.944 microg/kg (M), 0.373 and 0.449 microg/l (Bl). Geometric mean concentrations of lead in calves and cows were similarly low and were 33.0 and 47.5 microg/kg (Li), 38.9 and 58.3 microg/kg (Ki), 6.37 and 12.5 microg/kg (M), 5.47 and 12.2 microg/l (Bl). Sex had almost no effect on the amount of toxic metal accumulated except that kidney cadmium concentrations were significantly higher in females than males. Age did influence accumulation; cadmium and lead (but not arsenic) concentrations in most tissues were significantly greater in cows than female calves. For the trace elements, geometric mean copper levels in calf and cow tissues were 49.9 and 36.6 mg/kg (Li), 4.27 and 3.63 mg/kg (Ki), 0.649 and 1.68 mg/kg (M) and 0.878 and 0.890 mg/l (Bl). The corresponding zinc concentrations were 46.3 and 52.5 mg/kg (Li), 14.2 and 20.7 mg/kg (Ki), 47.3 and 52.5 mg/kg (M) and 2.80 and 2.22 mg/l (Bl). Female calves had significantly higher levels than males of muscle zinc and blood copper and zinc. Female calves accumulated more copper but less zinc in the liver and kidneys compared with cows; this may have been associated with the chronic, low level cadmium accumulation observed in cows. Overall, the levels of arsenic, cadmium, lead and zinc in cattle in Galicia do not constitute a risk for animal health. However, up to 20% of cattle in some regions in Galicia had levels of copper in the liver that exceeded 150 mg/kg wet wt. These animals may be at risk from copper poisoning. PMID- 10696726 TI - Quantities and associations of lead, zinc, cadmium, manganese, chromium, nickel, vanadium, and copper in fresh Mississippi delta alluvium and New Orleans alluvial soils. AB - The topic of this study is the effect of anthropogenic metals on the geochemical quality of urban soils. This is accomplished by comparing the metal contents and associations between two alluvial soils of the lower Mississippi River Delta, freshly deposited alluvial parent materials and alluvial soils collected from a nearby urban environment. Fresh alluvium samples (n = 97) were collected from the Bonnet Carre Spillway. The urban alluvial soil samples (n = 4026) were collected from New Orleans and stratified by census tracts (n = 286). The Spillway samples tend to have less Pb and Zn than generally noted for the baseline of natural soils. Except for Mn and V, Spillway alluvium contains significantly less metal than urban soils. For Spillway samples, the median metal content (in microg g( 1)) is 4.7 Pb, 11.1 Zn, 0.7 Cd, 164 Mn, 0.8 Cr, 3.9 Ni, 3.2 V, and 3.9 Cu. For urban soils, the median metal content (in microg g(-1)) is 120 Pb, 130 Zn, 3.2 Cd, 138 Mn, 2.1 Cr, 9.8 Ni, 3.8 V, and 12.7 Cu. Metal associations also differ between Spillway alluvium and urban alluvial soils. Fresh alluvium correlation coefficients between individual metals vary from 0.87 to 0.99 (P < 10(-13)) except for Cr which ranges from 0.57 to 0.68 (P < 10(-7)). The urban soil correlation coefficients for metals and the index value are 0.40-0.98. In urban soils, Pb, Zn, Cr, and Cu are dominant metals and highly associated, with a correlation coefficient ranging from 0.83 to 0.98 (P < 10(-25)). Their strong association justifies the use of GIS to map the integrated soil metal index (sum of the medians of metals by census tract) of New Orleans. Although also positively correlated (0.40-0.68, P < 10(-10)), Cd, Mn, Ni and V differ in their distribution in the city compared to Pb, Zn, Cr and Cu. Overall, significantly higher metal values occur in the inner city and lower values occur in outlying areas. The human health impact of the mixture of metals is not well understood. This study provides empirical data about the mixture and distribution of metals in New Orleans alluvial soils. Given common technical development, especially of traffic flows in cities, similar patterns of soil metals are expected for all US cities and probably international cities as well. Primary prevention of urban metal accumulations is necessary to enhance and sustain the development of urban culture. PMID- 10696727 TI - Heavy metals in wild rice from northern Wisconsin. AB - Wild rice grain samples from various parts of the world have been found to have elevated concentrations of heavy metals, raising concern for potential effects on human health. It was hypothesized that wild rice from north-central Wisconsin could potentially have elevated concentrations of some heavy metals because of possible exposure to these elements from the atmosphere or from water and sediments. In addition, no studies of heavy metals in wild rice from Wisconsin had been performed, and a baseline study was needed for future comparisons. Wild rice plants were collected from four areas in Bayfield, Forest, Langlade, Oneida, Sawyer and Wood Counties in September, 1997 and 1998 and divided into four plant parts for elemental analyses: roots, stems, leaves and seeds. A total of 194 samples from 51 plants were analyzed across the localities, with an average of 49 samples per part depending on the element. Samples were cleaned of soil, wet digested, and analyzed by ICP for Ag, As, Cd, Cr, Cu, Hg, Mg, Pb, Se and Zn. Roots contained the highest concentrations of Ag, As, Cd, Cr, Hg, Pb, and Se. Copper was highest in both roots and seeds, while Zn was highest just in seeds. Magnesium was highest in leaves. Seed baseline ranges for the 10 elements were established using the 95% confidence intervals of the medians. Wild rice plants from northern Wisconsin had normal levels of the nutritional elements Cu, Mg and Zn in the seeds. Silver, Cd, Hg, Cr, and Se were very low in concentration or within normal limits for food plants. Arsenic and Pb, however, were elevated and could pose a problem for human health. The pathway for As, Hg and Pb to the plants could be atmospheric. PMID- 10696728 TI - Comparison of the feasibility of three extraction procedures for trace metal partitioning in sediments from south-west Spain. AB - The feasibility of three sequential extraction schemes (a modification of the Tessier procedure, the scheme proposed by Meguellati and the protocol designed by BCR (now called the Standards, Measurements and Testing Programme, M&T) have been compared to study the distribution of As, Cd, Cr, Cu, Fe, Hg, Mn, Ni, Pb and Zn in sediment samples. The comparison has been performed by analyzing Certified Reference Material (CRM-601), a test material (S-12) and seven sediments from the Odiel Marshes Natural Park (located at the Atlantic coast of southern Spain). Samples were classified as sandy (with low iron oxide and organic matter contents) and clay-silty (with high iron oxide and organic matter contents) sediments. A higher metal mobility, especially under reducing conditions, was more properly assessed using the modified Tessier scheme compared to both the BCR and Meguellati procedures, these two later presenting comparative results for the reducible and residual phases. Significant Hg losses were found using the BCR procedure but the quantification of the acid phase for Cd, Cr and Ni was more reliable than that obtained with the modified Tessier and Meguellati schemes. PMID- 10696729 TI - Iodine-129 in human thyroids and seaweed in China. AB - The concentrations of 129I and the ratios of 129I/127I in normal human thyroids collected in Tianjin, China, and some seaweed samples from the Chinese coast were determined by neutron activation analysis. The mean 129I/127I ratio in these thyroids was found to be 1.13 x 10(-9), which is two orders of magnitude higher than the level of the pre-nuclear era, but one order of magnitude lower than the level in Europe in the post-nuclear era. There is no significant difference between the ratio of 129I/127I in the thyroids for the post-nuclear era from China and other areas, which are considered not to have been directly exposed to 129I emission from a nuclear source, such as Chile, Taiwan and Tokyo. The mean 129I/127I ratio in seaweed from the Chinese coast is 2.35 x 10(-10), approximately two orders of magnitude higher than in seaweed collected in the pre nuclear age, and similar to that from locations without direct exposure to the emission from nuclear installations, influenced only by global fallout. This indicates that the 129I level in China is within the global fallout background level. PMID- 10696730 TI - Atmospheric NO, temperature and ischaemic heart disease. Study in Toulouse and its conurbation. AB - Over a period of 388 days, associations were sought between the occurrence of acute myocardial infarction (AMI), levels of atmospheric nitrogen-containing gases (NO, NO2, NH3), temperature and relative humidity. During this period, there were 282 AMI validated by the WHO-MONICA criteria. Analysis of the data revealed, a decrease in the area, in AMI, when daily nitric oxide levels were above 13 microg m(-3) and when the mean daily temperature was below 13 degrees C. PMID- 10696731 TI - Translational research offers individually tailored treatments for cancer patients. AB - The measurement of the effect of cisplatin on DNA has become feasible with the development of antibodies against DNA adducts. In a phase II dose escalation trial with concomitant radiotherapy and daily cisplatin in lung cancer, we found that patients with high DNA adduct levels measured in the buccal mucosa had a much higher survival rate than patients with a low or undetectable amount of cisplatin-DNA adducts. The use of this assay may therefore allow the selection of individual patients for concomitant treatment with cisplatin and radiotherapy, as has been shown to be effective in randomized trials in patients with lung, head and neck, and cervix malignancies. To predict the response to radiation treatment, assays have been developed for tumor growth potential by measuring the labeling index after intravenous injection of IdUrd or by estimating cyclin D1 expression. Intrinsic radiation sensitivity of human tumors can be estimated by conventional techniques, which are probably too slow or cumbersome for routine use, or with more rapid assays, such as those for chromosome damage with fluorescent probes. These assays should be able to guide us in the adaptation of the individual radiation doses that should be applied and to select patients for an accelerated or hyperfractionated regimen. Pretreatment levels of apoptosis may also be helpful in predicting treatment outcome, although the data so far show inconsistent results. A better understanding of the signal transduction pathways involved in radiation-induced apoptosis may help in the design of studies aimed at modulating the apoptotic response, thereby increasing cell kill. We have recently shown that alkyllysophospholipids, which inhibit mitogenic signaling, induce apoptosis in a variety of tumor cell lines. In combination with ionizing radiation, these compounds cause an enhancement of apoptotic cell kill. This type of a signaling-based intervention could form the basis for new therapeutic strategies. The role of hormonal therapy in breast cancer patients, both in an adjuvant setting and for the treatment of disseminated disease, is becoming increasingly important. The development of a functional assay for the estrogen receptor (ER-FASAY), based on a yeast growth assay, provides a better way than the classical immunohistochemistry assay of estimating abnormal function of the receptor in tumors. These assays are simply examples, illustrating how clinicians could improve the therapeutic outcome for their patients by implementing knowledge obtained in the laboratory in clinical decision making. With further optimization of these assays, this holds the promise for the future that the treatment for each patient can be tailored rationally to the biology of the individual. PMID- 10696732 TI - Should postoperative mammograms be obtained routinely following the surgical excision of a breast cancer? PMID- 10696733 TI - Prevention with tamoxifen or other hormones versus prophylactic surgery in BRCA1/2-positive women: a decision analysis. AB - PURPOSE: Recent randomized controlled trials have shown that tamoxifen and raloxifene may prevent invasive breast cancer. This decision analysis study compares the outcomes of chemoprevention with tamoxifen, raloxifene, or oral contraceptives with the outcomes of prophylactic surgery among women with high risk BRCA1/2 mutations. PATIENTS AND METHODS: We used a simulated cohort of 30 year-old women who tested positive for BRCA1/2 mutations and constructed a Markov model with Monte Carlo simulations, incorporating cumulative breast and ovarian cancer incidence rates from the literature and survival figures from SEER data. We assumed that prophylactic surgery reduces ovarian cancer risk by 45% and breast cancer risk by 90%, that tamoxifen reduces invasive breast cancer risk by 49%, and that raloxifene has similar efficacy and safety in premenopausal and postmenopausal women. We used data obtained from high-risk women by a time trade off questionnaire to calculate quality-adjusted life-years. We based our cost estimates for hospital and ambulatory care on Health Care Financing Administration payments, the SEER-HCFA database, and the Pharmacy Fundamental Reference. RESULTS: In our model, a 30-year-old BRCA1/2+ woman could prolong survival by 0.9 years (95% probability interval, 0.4-1.2 years) by having bilateral oophorectomy, 3.4 years (2.7-3.7 years) by having bilateral mastectomy, and 4.3 years (3.6-4.6 years) by having both procedures instead of surveillance alone. Chemoprevention with tamoxifen and raloxifene increased survival by 1.6 years (1.0-2.1 years) and 2.2 years (1.3-2.8 years), respectively. Chemoprevention yielded more quality-adjusted life-years than did prophylactic surgery, even when treatment was delayed to age 40 or 50 years. All these procedures were cost-effective or cost-saving compared with surveillance alone. DISCUSSION: Our model suggests that although surgery may yield more substantial survival and cost benefits, quality of life issues may make chemoprevention a more attractive option for young women at high genetic risk. PMID- 10696734 TI - Elevations in serum soluble interleukin-2 receptor levels predict relapse in patients with hairy cell leukemia. AB - PURPOSE: Interferon-alfa, 2'-deoxycoformycin, and 2-chlorodeoxy-adenosine (2-CdA) are effective in the management of patients with hairy cell leukemia. These agents produce remissions in most patients, but relapses occur with all three drugs. The optimal means to follow patients for relapse after treatment has not been determined. METHODS: We retrospectively examined serial serum soluble interleukin-2 receptor levels (sIL-2R) and absolute granulocyte counts in eight patients with relapsed hairy cell leukemia. All were treated with 2-CdA at the time of relapse. Serum samples were available at 3- to 6-month intervals from 5 to 9 years before relapse and 2-CdA treatment RESULTS: sIL-2R levels increase only in patients who go on to relapse. sIL-2R levels doubled a mean of 17.1 months (range, 4-36 months) before absolute granulocyte count decreased by 50%. DISCUSSION: Demonstration of a rising serum sIL-2R level in patients with hairy cell leukemia identified those with an increased risk of relapse who need more frequent observation than patients who maintain a stable sIL-2R level. Early intervention may ameliorate the toxicity of salvage therapy because disease related neutropenia may be anticipated. PMID- 10696735 TI - The value of postlumpectomy mammogram in the management of breast cancer patients presenting with suspiciouis microcalcifications. AB - PURPOSE: It is recommended that patients with breast cancer who present with mammographically detected microcalcification should undergo postlumpectomy mammogram with magnification views to ensure adequate removal of all clinically demonstrable disease. The value of postlumpectomy mammogram has not been adequately examined in the literature. This report aims to quantify the value of such a study. MATERIALS AND METHODS: Retrospective review identified 90 breast cancer patients referred to our department between 1992 and 1997 who met all of the following criteria: (1) patients were considered for breast conserving management; (2) patients had suspicious microcalcifications on diagnostic mammograms; (3) the mammographic lesions were thought to be removed entirely on postexcision specimen radiographs; (4) surgical excisions were thought to be adequate on the basis of a review of the histologic pathology reports; and (5) postlumpectomy mammograms with magnification views were obtained. Fifty patients had invasive adenocarcinoma and 40 patients had ductal carcinoma in situ. The margins of last resection were clear, close, or focally involved in 70, 13, and seven patients, respectively. Patient records were reviewed to determine whether postlumpectomy mammograms demonstrated residual microcalcifications. RESULTS: Sixteen patients (17%) were found to have residual microcalcifications on postlumpectomy mammograms. Twelve patients underwent either local re-excision (seven patients) or simple mastectomy (five patients). Re-excision was not performed in four patients. Residual malignant cells were found in eight patients (67% of the re-excision group and 9% of the whole group). Six of these patients had their tumors initially resected with clear margins and the remaining two patients had their tumors initially resected with close margins. CONCLUSIONS: Postlumpectomy mammograms with magnification views detected residual clinical disease in a significant proportion of patients. Our result supports the routine use of this test, even when satisfactory postexcision specimen radiographs and adequate lumpectomy resection margins are obtained. This finding is particularly true for patients with ductal carcinoma in situ. PMID- 10696736 TI - Negative margin status improves local control in conservatively managed breast cancer patients. AB - OBJECTIVE: The purpose of this study was to determine the impact of final pathologic margin status on breast relapse-free survival, distant metastasis-free survival, and overall survival in patients undergoing conservative surgery and radiation therapy for invasive breast cancer. MATERIALS AND METHODS: Between January 1970 and December 1990, 984 patients underwent conservative surgery and radiation therapy at our institution as treatment for invasive breast cancer. After lumpectomy, patients were given radiation therapy to the intact breast with or without treatment to regional nodes with the routine use of electron boost to a total median tumor bed dose of 64 Gy. Pathology reports were available for review in 871 patients. Re-excision was carried out in 294 of these patients. For this analysis, patients were divided into four groups based on final pathologic margin status: negative (n = 278), dose (typically within 2 mm, n = 47), positive (n = 55), or indeterminate (n = 491). RESULTS: There were no significant differences between the groups with respect to age, histology, estrogen and progesterone receptor status, tumor location, or total radiation dose. Patients with negative margins were more likely than those with positive margins to have T1 mammographically detected lesions, to have negative nodal status, and to have undergone re-excision. Patients with positive margins were more likely to receive adjuvant chemotherapy or hormone therapy (P = 0.001). As of July 1998, with a median follow-up of 13 years, the median breast relapse-free survival, distant metastasis-free survival, and overall survival rates at 10 years for the entire cohort of patients were 86%, 81%, and 76%, respectively. Breast relapse-free survival at 10 years was 98% for patients with negative margins versus 98% for those with close margins versus 83% for those with positive margins versus 82% for those with indeterminate margins. There were no significant differences in breast relapse-free survival between patients with negative and dose margins or between patients with positive and indeterminate margins. Although the negative margin status also conferred an overall survival and distant metastasis-free survival advantage, this difference is confounded by the earlier stage of disease in these patients, and margin status did not influence overall survival in multivariate analysis. CONCLUSION: In patients undergoing conservative management of breast cancer, negative margin status significantly improves breast relapse free survival. Close margins appear equivalent to negative margins, and indeterminate margins appear equivalent to positive margins. Adjuvant chemotherapy or hormone therapy did not counteract the adverse impact of positive margin status. Re-excision to obtain dear surgical margins is recommended, even if a radiation boost or adjuvant systemic therapy is planned. PMID- 10696737 TI - Pilot study with weekly chemotherapy for patients with extensive small cell lung cancer: an Eastern Cooperative Oncology Group Study (PA586). AB - PURPOSE: Six of the most active chemotherapy agents in small cell lung cancer were administered sequentially in a weekly fashion in an attempt to optimize the dose and the number of agents received over a 12-week period. The purpose of this study was to estimate the efficacy and to characterize the toxicity of this approach. PATIENTS AND METHODS: Thirty-six patients with extensive-stage small cell lung cancer received weekly treatments with cisplatin and etoposide (weeks 1, 5, and 11), cyclophosphamide (weeks 2, 7, and 10), vincristine (weeks 2, 4, 7, 8, 10, and 12), methotrexate (weeks 3, 6, and 9), and doxorubicin (weeks 4, 8, and 12). Patients achieving a partial response received a second 12-week course. Patients achieving a complete response received prophylactic cranial radiation. RESULTS: Twenty-nine of the 36 patients completed the initial 12-week program over a median of 16 weeks. Hematologic toxicity was most prominent, with two deaths from sepsis and 31 patients having grade 3 or 4 neutropenia The overall response rate was 85%, with 33% of patients achieving a complete response. The median survival was 10.5 months, and the median time to progression was 8.2 months. DISCUSSION: This 12-week program, consisting of administration of six active agents for small cell lung cancer, caused significant myelosuppression that resulted in significant treatment delays and dose reductions. Although a high response rate was achieved, the median overall survival of 10.5 months was not significantly longer than expected from other standard two- to three-drug regimens. PMID- 10696738 TI - Effect of concurrent intra-arterial infusion of platinum drugs for patients with stage III or IV uterine cervical cancer treated with radical radiation therapy. AB - PURPOSE: The purpose of this study was to explore the effect of concurrent intra arterial infusion of platinum drugs in patients with stage III or IV uterine cervical cancer treated with radical radiation therapy. PATIENTS AND METHODS: Thirty-three patients with advanced (stage IIIA, 2; IIIB, 28; IVA, 3) uterine cervical squamous cell carcinoma were randomized into a concurrent intra-arterial infusion of platinum drugs with radiation therapy (IAPRT) group (18 patients) and a radiation therapy alone group (15 patients). After altering intrapelvic blood flow by embolization of the superior and inferior gluteal arteries under pelvic angiography, intra-arterial infusion of platinum drug through catheters inserted into both internal iliac arteries was performed concurrently with radiation therapy. One-shot infusion of cisplatin (100 mg/m2) twice with a 2- to 3-week interval was performed in eight patients, weekly infusion of carboplatin (100 mg/m2) via a reservoir five to six times was performed in four patients, and daily shot of cisplatin (10 mg/body) or 21 days via a reservoir was performed in six patients. Radiation therapy consisted of external-beam irradiation of 50 Gy/25 fractions/5 weeks for the whole pelvis with midline block after 30 Gy and intracavitary high-dose-rate brachytherapy using tandem and ovoids of 24 Gy/4 fractions/4 weeks to point A. RESULTS: The local complete response rate of the IAPRT group was 94% and was significantly higher than that of the radiation therapy group (67%). There were no significant differences in local response in the three drug delivery methods. Two- and 5-year overall survival rates were 54.5% and 44.4% in the IAPRT group, and 74.5% and 50.0% in the radiation therapy group, respectively. There was no significant difference between the two groups. In the IAPRT group, grade 3 or 4 acute bowel complications were seen in 33% of patients, grade 3 or 4 late bowel complications were seen 44%, and grade 3 or 4 myelosuppression was seen in 33%, and these complications were seen more in the IAPRT group than in the radiation therapy group and caused death in some patients. CONCLUSIONS: IAPRT had a better local response than radiation therapy but showed no proof of control over recurrence and had a poorer survival than radiation therapy. There were many local recurrences and distant metastases, contrary to the better first response of the IAPRT group over the radiation therapy group. Complications of the IAPRT group were very severe and made the patient's performance status and prognosis worse than in the radiation therapy group. We need to design some methods to decrease these complications to make use of the good local response acquired with IAPRT. Furthermore, we should re-examine the indication of IAPRT in patients with a large tumor because local recurrence and distant metastasis would be inevitable. PMID- 10696739 TI - Confluent choroidal infiltrates with sarcoidosis. AB - PURPOSE: To report the occurrence of confluent plaquelike choroidal infiltrates in four patients with sarcoidosis. METHODS: The medical records of patients with choroidal plaquelike infiltrates and presumed systemic sarcoidosis seen in the Mayo Clinic were reviewed. RESULTS: The cohort included four patients with confluent plaquelike choroidal infiltrates and systemic sarcoidosis. The most common ophthalmic symptom experienced by the patients was mild blurring of vision. The salient ophthalmic findings were choroidal infiltrates generally unaccompanied by other features frequently seen with sarcoidosis such as periphlebitis, multiple small yellowish choroidal infiltrations, vitreous cellular reaction, and granulomatous anterior uveitis. The choroidal lesions were confluent, yellowish, irregularly thickened infiltrates that frequently radiated peripherally from a peripapillary location in an ameboidlike pattern. Fluorescein angiography demonstrated early hypofluorescence and progressive patchy hyperfluorescence with late staining of the active lesions. Treatment with either systemic or subconjunctival steroids was associated with improvement in vision and partial resolution of the choroidal lesions. In two patients some of the lesions resolved spontaneously and became replaced by areas of chorioretinal atrophy. CONCLUSION: Plaquelike yellowish choroidal infiltrates associated with systemic sarcoidosis may occur in eyes remarkably free of other signs of inflammation. The infiltrates tend to radiate from the region of the optic nerve in a confluent ameboidlike pattern. They generally respond to corticosteroids and may be the first recognized manifestation of systemic sarcoidosis. PMID- 10696740 TI - Choroidal white lesions as an early manifestation of sarcoidosis. AB - PURPOSE: To describe the natural history of a series of patients with fine white choroidal lesions and uveitis of previously unknown cause. METHODS: A retrospective chart review of 11 patients with chronic uveitis and multiple, small (50-100 microm), peripheral white lesions of the choroid was performed using a standardized questionnaire form. RESULTS: Ten of 11 patients were white women with an average age of 62 years. Seven of 11 patients had panuveitis; 4 of 11 patients had vitritis; and 6 of 11 patients had cystoid macular edema. Choroidal white lesions were bilateral in all but one patient. Seven of 11 patients were followed up for more than 1 year. During a 12- to 173-month follow up (mean, 94 months), these patients showed coalescence and atrophy of the white lesions. Initial systemic examination for the cause of the uveitis and white choroidal lesions was negative in all seven patients. With long-term follow-up, sarcoidosis was diagnosed in five of the seven patients followed for more than 1 year. CONCLUSIONS: The pattern of inflammatory white choroidal lesions distributed in the peripheral retina that atrophy and coalesce with time and that are associated with uveitis in middle-aged white women may represent an early form of sarcoidosis. PMID- 10696741 TI - Choroidopathy in patients with sarcoidosis observed by simultaneous indocyanine green and fluorescein angiography. AB - PURPOSE: To examine choroidopathy in patients with sarcoidosis. METHODS: In a prospective clinical study, 10 consecutive patients (20 eyes) with sarcoidosis underwent simultaneous indocyanine green (ICG) and fluorescein angiography (FA) with a double detector of scanning laser ophthalmoscopy. Angiographic findings recorded on videotapes were evaluated, and the relation of ICG angiographic findings with systemic activity and the extent of retinal inflammation was analyzed. RESULTS: Indocyanine green angiographic findings, not evident with fluorescein, were multiple lobular hypofluorescent spots in the posterior pole in 10 eyes of 5 patients, isolated hypofluorescent plaques in 2 eyes of 1 patient, hyperfluorescent spots in 3 eyes of 2 patients, and segmental choroidal vascular wall staining in 6 eyes of 4 patients. The multiple lobular hypofluorescent spots with ICG were observed at a significantly higher rate in eyes with extensive retinal vascular leakage of fluorescein than in eyes with minimal leakage (chi square test, P = 0.0007). CONCLUSIONS: The patients with sarcoidosis showed choroidal abnormalities that could be revealed by ICG angiography, but not by funduscopy or FA. Simultaneous ICG and FA angiography may be useful for examining choroidal lesions in sarcoidosis. PMID- 10696742 TI - Presumed rhegmatogenous retinal detachment in patients with retinoblastoma. AB - PURPOSE: To review the characteristics, treatment, and outcome of presumed rhegmatogenous retinal detachment (RD) in patients with retinoblastoma. METHODS: Descriptive consecutive case series study from 1970 to 1996. RESULTS: Of 45 eyes with retinoblastoma that received various modalities of eye-sparing treatment and adequate follow-up, five developed presumed rhegmatogenous RD after treatment. All five eyes had previous treatment with external beam radiation, four of which also had been treated with cryotherapy. To minimize the potential spread of the malignancy intraoperatively, long disease-free intervals were ensured before scleral buckling surgeries were performed, and special care was taken during subretinal fluid drainage. The RDs were totally reattached in three eyes and partially reattached in the other two. CONCLUSION: Presumed rhegmatogenous RD may occur after external beam radiation or cryotherapy for retinoblastoma. It poses special challenges in management. PMID- 10696743 TI - Repair of late retinal detachment after successful treatment of retinoblastoma. AB - PURPOSE: To analyze the results of vitreous surgery for late retinal detachment (RD) after successful treatment of retinoblastoma. METHODS: The records of all patients with retinoblastoma seen at a single ocular oncology service between 1982 and 1998 were reviewed to identify patients treated for late RD. Previous treatments, characteristics of the RD, surgical techniques used, and visual and anatomic results of the surgery were recorded. RESULTS: Of more than 500 charts reviewed, four patients treated for late RD were identified. All four had received previous, whole-eye, external beam radiotherapy and subsequently required cataract surgery. Other previous treatments included radioactive plaque, cryotherapy, xenon photocoagulation, and chemotherapy. At presentation, some patients had shifting subretinal fluid. None had a tear identifiable preoperatively, but two patients had a definite small slit tear at a tumor edge identified at surgery. One patient had a primary scleral buckle that failed. All patients had vitreous surgery with silicone oil. Average postsurgical follow-up was 30 months. Preoperative visual acuity ranged from 20/80 to light perception and improved postoperatively in two patients. The retina remained completely attached in three patients. CONCLUSIONS: Despite shifting subretinal fluid and no identifiable tear, a rhegmatogenous RD should be considered if it occurs late in patients with otherwise stable, treated retinoblastoma. Tumor reactivation must be excluded carefully. Vitreous surgery can be used to repair the RD successfully and improve vision. PMID- 10696744 TI - Histopathologic changes in retinoblastoma after chemoreduction. AB - PURPOSE: To report the histologic findings in the eyes of two patients with bilateral retinoblastoma who underwent chemoreduction therapy and enucleation of one eye. METHODS: Clinical histories were obtained for both patients. The enucleated eyes were routinely processed and sections were stained with hematoxylin and eosin. RESULTS: The first patient underwent two cycles of carboplatin, vincristine, and etoposide, and the second patient underwent one cycle of carboplatin, vincristine, and etoposide before enucleation. The eyes of both patients exhibited a clinical type 3 regression pattern. Histopathologic examination showed a gliotic mass with interspersed calcifications in one eye and necrotic tumor adjacent to histologically intact retinoblastoma in the other eye. CONCLUSION: Chemoreduction has variable effects on retinoblastoma and the clinical type 3 regression pattern has several histologic counterparts. PMID- 10696745 TI - Optical coherence tomography in the assessment of retinal pigment epithelial tear. AB - PURPOSE: To study the findings of optical coherence tomography (OCT) in retinal pigment epithelial (RPE) tears. METHODS: Sixteen eyes of 16 consecutive patients with age-related macular degeneration complicated by RPE tear were studied using OCT. Fluorescein angiography also was performed. Thirteen eyes were at the acute stage and three eyes were at the scarring stage, still with a recognizable tear. RESULTS: Optical coherence tomography identified an RPE detachment (PED) with focal interruption of the RPE in all cases. Optical coherence tomography always highlighted a peculiar non-dome-shaped profile of the serous PED, as opposed to that of the PED not complicated by an RPE tear. A very intense hyperreflectivity was observed in the OCT scans performed through the retracted RPE. A deep hyperreflectivity under the line corresponding to the RPE was evident in the area of the bare choroid. No choroidal neovascularization could be visualized using OCT, either at the acute or at the scarring stages. CONCLUSIONS: Optical coherence tomography, a noncontact, noninvasive imaging technique, may be a useful tool, complementary to fluorescein angiography, in the clinical assessment of RPE tears. PMID- 10696746 TI - Topical anesthesia in posterior vitrectomy. AB - PURPOSE: To evaluate the efficacy of topical anesthesia as an alternative to peribulbar or retrobulbar anesthesia in posterior vitrectomy procedures. METHODS: Posterior vitrectomy using topical anesthesia (4% lidocaine drops) was performed prospectively in 134 eyes (134 patients) with various vitreoretinal diseases, including severe proliferative diabetic retinopathy (n = 69), vitreous hemorrhage (n = 12), rhegmatogenous retinal detachments (n = 11), epiretinal membranes (n = 10), macular holes (n = 7), dislocated crystalline lens or intraocular lens (n = 6), giant retinal tears (n = 5), intraocular foreign bodies (n = 3), trauma (n = 3), endophthalmitis (n = 3), subfoveal choroidal neovascular membrane (n = 3), and neovascular glaucoma (n = 2). In 26 (19.4%) eyes, posterior vitrectomy was combined with a scleral buckling procedure, and in 84 (62.6%) eyes, argon laser photocoagulation was performed. Preoperative and intraoperative sedation of varying degrees was necessary. Subjective pain and discomfort were graded from 1 (no pain or discomfort) to 4 (severe pain and discomfort). RESULTS: All patients had grade 1 pain and discomfort during most of the procedure. All patients had grade 2 (mild) pain and discomfort during pars plana sclerotomies, external bipolar cautery, and conjunctival closure. The average amount of 4% lidocaine drops needed during each procedure was 0.5 mL. No patient required additional retrobulbar, peribulbar, or sub-Tenon anesthesia. CONCLUSIONS: This technique avoids the risk of globe perforation, retrobulbar hemorrhage, and prolonged postoperative akinesia of the eye. With appropriate case selection, topical anesthesia is a safe and effective alternative to peribulbar or retrobulbar anesthesia in three-port pars plana vitrectomy procedures. PMID- 10696747 TI - The role of preoperative subconjunctival mydricaine and topical diclofenac sodium 0.1% in maintaining mydriasis during vitrectomy. AB - PURPOSE: To establish the role of preoperative subconjunctival mydricaine and diclofenac 0.1% in maintaining mydriasis during vitrectomy. METHODS: Fifty-seven patients were entered into the study. All were given cyclopentolate 1% and phenylephrine 2.5% preoperatively. Each patient was randomly allocated to one of three groups. In Group 1, patients received mydricaine by subconjunctival injection and diclofenac 0.1% topically preoperatively. In Group 2, patients received only subconjunctival mydricaine. Group 3 patients received only topical diclofenac preoperatively. Pupil diameter was measured with calipers before and at the end of the operation. RESULTS: There was no statistically significant difference in the change in pupil size between Groups 2 and 3. In all patients in Group 1 (who received both subconjunctival mydricaine and diclofenac preoperatively), pupil size was either maintained or increased after vitrectomy. This result was statistically significant when compared with the other groups (for Group 1 versus Group 2, P<0.005; for Group 1 versus Group 3, P<4.7x10(-06)). CONCLUSION: Topical diclofenac is useful for maintaining pupil size during vitrectomy only when used in conjunction with subconjunctival mydricaine, especially in patients in whom prolonged surgery is anticipated. PMID- 10696748 TI - Ultrasound biomicroscopy for examination of the sclerotomy site in eyes with proliferative diabetic retinopathy after vitrectomy. AB - PURPOSE: We evaluated the capability of ultrasound biomicroscopy (UBM) to predict fibrovascular proliferation at sclerotomy sites in eyes with postoperative vitreous hemorrhage due to proliferative diabetic retinopathy (PDR). METHODS: Ultrasound biomicroscopy was used for examining the sclerotomy sites in 13 eyes of 11 patients with PDR experiencing postoperative vitreous hemorrhage (PDR group). Thirty-nine sclerotomy sites (all entry sites of each eye) were examined before reoperation, and the UBM images were compared with findings obtained during revision of the vitrectomy. Thirteen eyes of 13 patients undergoing vitrectomy for nondiabetic diseases were used as controls and examined after vitrectomy. RESULTS: The UBM images were classified into the following four categories: A, tent; B, spheroid; C, trapezoid; and N, none. The findings were distributed as follows in the PDR group: category A, 18%; B, 5%; C, 56%; and N, 21 %; and as follows in the control group: category A, 28%; B, 5%; C, 5%; and N, 62%. In the PDR group, 11 of 12 sclerotomy sites disclosing fibrovascular proliferation possessed the trapezoidal image. Mean length of trapezoidal base was 2.49+/-0.97 mm and 1.51+/-0.75 mm in the groups with and without fibrovascular proliferation, respectively (P<0.01). The average relative reflectivity of the trapezoidal image against the sclera was 0.501+/-0.169 in the fibrovascular proliferation group and 0.891+/-0.183 in the fibrous ingrowth group (P<0.01). CONCLUSION: Ultrasound biomicroscopy is useful in detecting fibrovascular proliferation at sclerotomy sites because a large and low reflecting trapezoidal UBM image is highly correlated to its presence. PMID- 10696749 TI - Corneal endothelial cell loss in eyes undergoing lensectomy with and without anterior lens capsule removal combined with pars plana vitrectomy and gas tamponade. AB - PURPOSE: To assess prospectively the comeal endothelial damage associated with pars plana vitrectomy combined with lensectomy and gas tamponade. METHODS: The corneal endothelium was examined with a specular microscope preoperatively and 6 months postoperatively in 42 eyes that underwent pars plana vitrectomy combined with total lensectomy and gas tamponade with SF6 (19 eyes) or C3F8 (23 eyes), and in 12 control eyes that underwent vitrectomy combined with anterior capsule preserved lensectomy and gas tamponade. RESULTS: The mean +/- SD endothelial cell loss was 17.19+/-7.88% in the SF6 tamponade group, 27.54+/-10.57% in the C3F8 tamponade group, and 3.49+/-2.68% in the control group. Significant differences were found in the mean cell loss among these three groups. The degree of cell loss was significantly related to gas tamponade duration in the eyes with total lensectomy. CONCLUSION: These results suggest that ophthalmic surgeons must try to preserve the lens capsule as much as possible in vitrectomy combined with lensectomy and long-acting gas tamponade in order to prevent corneal endothelial damage. PMID- 10696750 TI - Electron immunocytochemical analysis of posterior hyaloid associated with diabetic macular edema. AB - BACKGROUND: Tangential traction in the macula from a thickened posterior hyaloid of the vitreous has been implicated as a cause of diffuse diabetic macular edema. Vitrectomy with peeling of the posterior hyaloid has been shown to reduce retinovascular leakage and improve vision in select patients. We report a clinicopathologic correlation using electron microscopy and electron immunocytochemistry to characterize the membrane infiltrating the posterior hyaloid in two such patients. METHODS: Two patients presented with vision loss associated with diffuse diabetic macular edema and an attached, thickened, and taut posterior hyaloid. The patients underwent vitrectomy with peeling of the posterior hyaloid. The premacular posterior hyaloid specimens then were analyzed by electron microscopy with immunocytochemical staining for cytokeratin and glial fibrillary acidic protein. RESULTS: Both posterior hyaloid specimens contained collagen and a large cellular component. Immunogold labeling showed cells positive for glial fibrillary acidic protein or cytokeratin. With double labeling, no cells expressed both proteins simultaneously. Clinically, both patients had vision improvement and macular edema resolution after surgery. CONCLUSIONS: The thickened, taut posterior hyaloid observed in our patients with diabetic macular edema contained cells of glial and epithelial origin. This cellular infiltration may contribute to abnormal vitreomacular adherence and could play a role in the pathogenesis of macular edema in some patients with diabetes. PMID- 10696751 TI - The efficacy of plasminogen-urokinase combination in inducing posterior vitreous detachment. AB - PURPOSE: To investigate the toxicity of intravitreal plasminogen, urokinase, and their combination, and to evaluate their efficacy in the production of posterior vitreous detachment (PVD) in the rabbit eye. METHODS: Fifty-six albino New Zealand rabbits were examined before and after injection using the indirect ophthalmoscope, slit-lamp biomicroscopy, and electroretinography. Various concentrations of urokinase or recombinant plasminogen or a combination were injected intravitreally into the right eyes of four rabbits for each concentration. The left eyes of the animals served as controls and received 0.1 mL balanced salt solution. Group 1 was injected with pure urokinase (1,000, 5,000, or 10,000 IU); Group 2 with recombinant plasminogen (0.1, 0.4, 1.0, 2.0, 4.0, 8.0, or 16.0 caseinolytic units [CU]); and Group 3 with a combination of 1,000 IU urokinase (highest nontoxic dose) and nontoxic concentrations of plasminogen (0.1, 0.4, 1.0, or 2.0 CU). The animals were killed and the eyes enucleated 15 days after injection. Electron and light microscopy were performed. RESULTS: A concentration of 1,000 IU of urokinase was found to be nontoxic to the retina. Plasminogen concentrations of 2.0 CU or less did not produce retinal toxicity, whereas 4.0, 8.0, and 16.0 CU of plasminogen caused minimal-to-severe inflammatory response in the vitreous without histologic or electroretinographic changes. Neither plasminogen nor urokinase alone was successful in producing PVD. The combination of 1,000 IU of urokinase and 1.0 to 2.0 CU of plasminogen was effective without causing retinal toxicity. CONCLUSION: Posterior vitreous detachment can be produced in the rabbit eye using a combination of plasminogen and urokinase. PMID- 10696752 TI - Intraocular melanoma spread to regional lymph nodes: report of two cases. AB - PURPOSE: To report two cases of regional lymphatic spread of primary uveal melanoma. METHODS: The clinical records of two patients who underwent enucleation for uveal melanoma and later developed regional lymph node metastases were reviewed. One of the two eyes was initially treated with proton beam irradiation. Histologic sections of the enucleated eyes and excised lymph nodes were examined. RESULTS: The melanomas arose in the choroid and ciliary body of the two patients and spread to regional lymph nodes 2 years after enucleation. The choroidal melanoma recurred after irradiation, diffusely infiltrated the uveal tract, and extended into the conjunctiva via an emissary canal. The ciliary body melanoma spread through the trabecular meshwork to the conjunctiva. CONCLUSIONS: Choroidal and ciliary body melanoma may rarely exhibit regional lymph node metastasis. This mode of metastasis may occur after extraocular spread and invasion of conjunctival lymphatics. PMID- 10696753 TI - Vitrectomy update for macular traction in ocular toxocariasis. AB - PURPOSE: To study the results of modern vitrectomy in traction and combined traction-rhegmatogenous retinal detachment involving the macula in cases of ocular toxocariasis. METHODS: This was a cohort study of patients seen in different institutions in the United States. Ten eyes of 10 patients were studied. Vitrectomy was performed in all eyes, combined with membrane removal, scleral buckle, fluid-gas exchange, silicone oil, or lensectomy in certain cases. The anatomic and visual results of surgery were reviewed. RESULTS: Ten eyes from 10 patients ranging in age from 2 to 33 years (median, 6 years) were reviewed. Follow-up ranged from 3 months to 8 years (median, 2 years). All eyes achieved macular attachment following surgery; vision improved in 5 (50%) eyes, and was unchanged in 5 (50%). Histologic specimens from six eyes were reviewed, and revealed combinations of fibrous tissue, eosinophils, plasma cells, lymphocytes, and giant cells. One specimen revealed an encysted Toxocara canis organism. CONCLUSION: Inflammation created in response to Toxocara larvae may lead to traction retinal detachment of the macula. Vitreoretinal surgery has a good chance of reattaching the macula and improving vision. PMID- 10696754 TI - Diagnostic and therapeutic challenges. PMID- 10696755 TI - Diagnostic and therapeutic challenges. PMID- 10696756 TI - Macular hole following ruptured retinal arterial macroaneurysm. PMID- 10696757 TI - Dense premacular hemorrhage from a retinal macroaneurysm treated by argon laser. PMID- 10696758 TI - Central serous chorioretinopathy associated with a carcinoma of the adrenal cortex. PMID- 10696759 TI - Morphometric comparison of subfoveal neovascular membranes and postoperative retinal pigment epithelial defects in age-related macular degeneration. PMID- 10696760 TI - An easy method to remove retinal biopsy specimens from the eye. PMID- 10696761 TI - External diaphanoscopic illuminator: a new device for visualization in pars plana vitrectomy. PMID- 10696762 TI - Pseudo-silicone oil shine on the anterior surface of ectropion uveae. PMID- 10696763 TI - Profiles in toxicology. Harold Carpenter Hodge (1904-1990). PMID- 10696764 TI - Lessons learned in applying the U.S. EPA proposed cancer guidelines to specific compounds. AB - An expert panel was convened to evaluate the U.S. Environmental Protection Agency's "Proposed Guidelines for Carcinogen Risk Assessment" through their application to data sets for chloroform (CHCl3) and dichloroacetic acid (DCA). The panel also commented on perceived strengths and limitations encountered in applying the guidelines to these specific compounds. This latter aspect of the panel's activities is the focus of this perspective. The panel was very enthusiastic about the evolution of these proposed guidelines, which represent a major step forward from earlier EPA guidance on cancer-risk assessment. These new guidelines provide the latitude to consider diverse scientific data and allow considerable flexibility in dose-response assessments, depending on the chemical's mode of action. They serve as a very useful template for incorporating state-of-the-art science into carcinogen risk assessments. In addition, the new guidelines promote harmonization of methodologies for cancer- and noncancer-risk assessments. While new guidance on the qualitative decisions ensuing from the determination of mode of action is relatively straightforward, the description of the quantitative implementation of various risk-assessment options requires additional development. Specific areas needing clarification include: (1) the decision criteria for judging the adequacy of the weight of evidence for any particular mode of action; (2) the role of mode of action in guiding development of toxicokinetic, biologically based or case-specific models; (3) the manner in which mode of action and other technical considerations provide guidance on margin-of-exposure calculations; (4) the relative roles of the risk manager versus the risk assessor in evaluating the margin of exposure; and (5 ) the influence of mode of action in harmonizing cancer and noncancer risk assessment methodologies. These points are elaborated as recommendations for improvements to any revisions. In general, the incorporation of examples of quantitative assessments for specific chemicals would strengthen the guidelines. Clearly, any revisions should retain the emphasis present in these draft guidelines on flexibility in the use of scientific information with individual compounds, while simultaneously improving the description of the processes by which these mode-of action data are organized and interpreted. PMID- 10696765 TI - Quantitative evaluation of alternative mechanisms of blood disposition of di(n butyl) phthalate and mono(n-butyl) phthalate in rats. AB - Phthalate esters are ubiquitous, low-level environmental contaminants that induce testicular toxicity in laboratory animals. The diester is rapidly metabolized in the gut to the monoester, which causes the testicular toxicity. Several physiologically based pharmacokinetic (PBPK) model structures have been evaluated for di(2-ethylhexyl) phthalate (DEHP) and mono(2-ethylhexyl) phthalate (MEHP). The objective of this study was to test these PBPK models for a less lipophilic phthalate diester, di(n-butyl) phthalate (DBP), and monoester, mono(n-butyl) phthalate (MBP). Alternate models describing enterohepatic circulation, diffusion limitation, tissue pH gradients (pH trapping), and a simpler, flow-limited model were evaluated. A combined diffusion-limited and pH trapping model was also tested. MBP tissue:blood partition coefficients were similar when determined either experimentally by a nonvolatile, vial equilibration technique or algorithmically. All other parameters were obtained from the literature or estimated from MBP blood concentrations following intravenous or oral exposure to DBP or MBP. A flow-limited model was unable to predict MBP blood levels, whereas each alternative model had statistically better predictions. The combined diffusion-limited and pH trapping model was the best overall, having the highest log-likelihood function value. This result is consistent with a previous finding that the pH trapping model was the best model for describing DEHP and MEHP blood dosimetry, though it was necessary to extend the model to include diffusion limitation. The application of the pH trapping model is a step toward developing a generic model structure for all phthalate esters, though more work is required before a generic structure can be identified with confidence. Development of a PBPK model structure applicable to all phthalate esters would support more realistic assessments of risk to human health from exposure to one or more members of this class of compounds. PMID- 10696766 TI - Analysis of respiratory exchange of methanol in the lung of the monkey using a physiological model. AB - A physiologically based pharmacokinetic (PBPK) model was developed for the monkey, to account for fractional systemic uptake of inhaled methanol vapors in the lung. Fractional uptake of inhaled [14C]-methanol was estimated using unreported exhaled breath time course measurements of [14C]-methanol from the D.C. Dorman et al. (1994, Toxicol Appl Pharmacol. 128, 229-238) lung-only exposure study. The cumulative amount of [14C]-methanol exhaled was linear with respect to exposure duration (0.5 to 2 h) and concentration (10 to 900 ppm). The model estimated that forty to eighty-one percent of the of inhaled [14C]-methanol delivered to the lung was taken into systemic circulation in female Cynomolgus monkeys exposed for two h to 10-900 ppm of [14C]-methanol. There was no apparent trend between the percent of inhaled [14C]-methanol absorbed systemically and the [14C]-methanol exposure concentration. Model simulations were conducted using a single saturable Michaelis-Menten equation with Vmaxc, the metabolic capacity set to 15.54 mg/kg/h and Km, the affinity constant, to 0.66 mg/l. The [14C]-methanol blood concentrations were variable across [14C]-methanol exposure groups and the PBPK model tended to over-predict systemic clearance of [14C]-methanol. Accounting for fractional uptake of inhaled polar solvents is an important consideration for risk assessment of inhaled polar solvents. PMID- 10696767 TI - Biotransformation and kinetics of excretion of ethyl tert-butyl ether in rats and humans. AB - Ethyl tert-butyl ether (ETBE) may be used in the future as an additive to gasoline to increase oxygen content and reduce tailpipe emissions of pollutants. Therefore, widespread human exposure may occur. To contribute to the characterization of potential adverse effects of ETBE, its biotransformation was compared in humans and rats after inhalation exposure. Human volunteers (3 males and 3 females) and rats (5 males and 5 females) were exposed to 4 (4.5+/-0.6) and 40 (40.6+/-3.0) ppm ETBE for 4 h in a dynamic exposure system. Urine samples from rats and humans were collected for 72 h at 6-h intervals, and blood samples were taken in regular intervals for 48 h. In urine, ETBE and the ETBE-metabolites tert butanol (t-butanol), 2-methyl-1,2-propane diol, and 2-hydroxyisobutyrate were quantified; ETBE and t-butanol were determined in blood samples. After the end of the exposure period to inhalation of 40-ppm ETBE, blood concentrations of ETBE were found at 5.3+/-1.2 microM in rats and 12.1+/-4.0 microM in humans. The ETBE blood concentrations, after inhalation of 4-ppm ETBE, were 1.0+/-0.7 microM in rats and 1.3+/-0.7 microM in humans. ETBE was rapidly cleared from blood. After the end of the 40-ppm ETBE exposure period, the blood concentrations of t-butanol were 13.9+/-2.2 microM in humans and 21.7+/-4.9 microM in rats. After 4-ppm ETBE exposure, blood concentrations of t-butanol were 1.8+/-0.2 microM in humans and 5.7+/-0.8 microM in rats. t-Butanol was cleared from human blood with a half-life of 9.8+/-1.4 h in humans after 40-ppm ETBE exposure. In urine samples from controls and in samples collected from the volunteers and rats before the exposure, low concentrations of t-butanol, 2-methyl-1,2-propane diol, and 2 hydroxyisobutyrate were present. In the urine of both humans and rats exposed to ETBE, the concentrations of these compounds were significantly increased. 2 Hydroxy-isobutyrate was recovered in urine as the major excretory product formed from ETBE; t-butanol and 2-methyl-1,2-propane diol were minor metabolites. All metabolites of ETBE excreted with urine were rapidly eliminated in both species after the end of the ETBE exposure. Excretion half-lives for the different urinary metabolites of ETBE were between 10.2 and 28.3 h in humans and 2.6 and 4.7 h in rats. The obtained data indicate that ETBE biotransformation and excretion are similar for rats and humans, and that ETBE and its metabolites are rapidly excreted by both species. Between 41 and 53% of the ETBE retained after the end of the exposure was recovered as metabolites in the urine of both humans and rats. PMID- 10696768 TI - Hypochlorous acid-mediated activation of N-acetylbenzidine to form N'-(3' monophospho-deoxyguanosin-8-yl)-N-acetylbenzidine. AB - Hypochlorous acid (HOCl), a chemically reactive oxidant, is an important component of the inflammatory response and may contribute to carcinogenesis. This study assessed the possible activation of N-acetylbenzidine (ABZ) by HOCI to form a specific DNA adduct, N'-(3'-monophospho-deoxyguanosin-8-yl)-N-acetylbenzidine. HOCl was incubated with 0.06 mM 3H-ABZ, and transformation assessed by HPLC. Similar results were observed at pH 5.5 or 7.4. A linear increase in transformation was observed from 0.025 to 0.1 mM HOCl with up to 80% of ABZ changed. Approximately, 2 nmoles of HOCI oxidized 1 nmole of ABZ. N-oxidation products of ABZ metabolism, such as N'-hydroxy-N-acetylbenzidine, were not detected. Oxidation of ABZ was prevented by taurine, DMPO, glutathione, and ascorbic acid, whereas mannitol was without effect. Results are consistent with a radical mechanism. In the presence of 2'-deoxyguanosine 3'-monophosphate (dGp), a new product (dGp-ABZ) was observed. The same adduct was observed with DNA. dGp ABZ was found to be quite stable (>80% remaining) at 70 degrees C in pH 5.5 (60 min) and 7.4 (240 min). Electrospray mass spectrometry indicated that dGp-ABZ was N'-(3'-monophospho-deoxyguanosin-8-yl)-N-acetylbenzidine, and this was confirmed by NMR. 32P-postlabeling in combination with TLC and HPLC determined that the adduct made by either HOCl or prostaglandin H synthase oxidation of ABZ in the presence of dGp or DNA was dGp-ABZ. Thus, HOCI activates ABZ to form dGp-ABZ and may be responsible for the presence of this adduct in peripheral white blood cells from workers exposed to benzidine. Reaction of ABZ with HOCl provides an easy, convenient method for preparing dGp-ABZ. PMID- 10696769 TI - Responses of transgenic mouse lines p53(+/-) and Tg.AC to agents tested in conventional carcinogenicity bioassays. AB - The haplo-insufficient p53 knockout (p53+/-) and zetaglobin v-Ha-ras (Tg.AC) transgenic mouse models were compared to the conventional two rodent species carcinogen bioassay by prospectively testing nine chemicals. Seven of the chemicals classified as carcinogens in the conventional bioassay induced tumors in the liver or kidneys of B6C3F1 mice, and one (pentachlorophenol) also induced tumors in other tissues. Only three chemicals, furfuryl alcohol, pyridine, and pentachlorophenol, induced tumors in rats. The tumorigenic effect of pyridine was seen in F344 rats but not in Wistar strain rats. None of the chemicals induced tumors in the p53+/- transgenic mice, which is consistent with the absence of genotoxicity of these chemicals. Only two of the seven nongenotoxic carcinogens were positive in the Tg.AC model (lauric acid diethanolamine and pentachlorophenol). These results show that these transgenic models do not respond to many chemicals that show strain- or species-specific responses in conventional bioassays. PMID- 10696770 TI - DNA-damaging effects of genotoxins in mixture: modulation of covalent binding to DNA. AB - Modulation of DNA adduct formation by pre-existing adducts was examined in synthetic oligonucleotides and genomic DNA (calf thymus); genotoxins studied were N-acetoxy-acetylaminofluorene (N-AcO-AAF), aminofluorene (AF), aflatoxin B1-8,9 epoxide (AFB1-8,9-epoxide), and dimethylsulfate (DMS). Oligodeoxynucleotides containing either guanine-C8-AAF (Gua-C8-AAF) or Gua-C8-AF adducts and a neighboring unadducted guanine (G) (target G), located 1, 2, or 4 nucleotides from the adduct, were reacted, as single- (ss) or double-stranded (ds) substrates, with dimethylsulfate (DMS) or AFB1-8,9-epoxide. A modified Maxam Gilbert technique showed that the presence of the AAF adduct lowered the extent to which AFB1-8,9-epoxide, but not DMS, reacted with target G. Binding of AFB1 8,9-epoxide to the target G was attenuated (> or =5-fold) when the target was located immediately adjacent to an AAF, but not AF, adduct in ds-DNA. Reaction with AFB1-8,9-epoxide increased when the target G was located 2 or 4 nucleotides from the AAF adduct. Pretreatment of calf thymus DNA with AAF (0-1.8% nucleotides modified) reduced levels of Gua-N7-AFB1 adducts formed after subsequent treatment with AFB1-8,9-epoxide. Pretreatment of calf thymus DNA with AFB1 did not alter levels of adducts formed after subsequent treatment with N-AcO-AAF. The supposition that aflatoxin B1-binding to DNA may be altered by conformational changes in the helix, due to the presence of a pre-existing AAF adduct, is supported by the absence of an effect by AF and confirmation of local denaturation of the oligomer helix by use of chemical probes hydroxylamine and diethylpyrocarbonate. Nonetheless, the importance of changes in the nucleophilicity of neighboring nucleotides and local steric effects cannot be ruled out. PMID- 10696771 TI - Re-evaluation of the 2-year chloroform drinking water carcinogenicity bioassay in Osborne-Mendel rats supports chronic renal tubule injury as the mode of action underlying the renal tumor response. AB - Chloroform, generally regarded as a non-genotoxic compound, is associated with the induction of liver and/or kidney tumors in laboratory mice and rats. In particular, chloroform produced renal tubule tumors in low incidence in male Osborne-Mendel rats when administered by corn-oil gavage or in the drinking water. There is a lack of data on intermediate endpoints that may be linked to renal cancer development in this strain of rat, in contrast to mice. Specifically, evidence linking chloroform-induced liver and kidney tumors in mice with cytotoxicity and regenerative cell proliferation is very strong, but weak in the rat. In the present study, kidney tissue from a carcinogenicity bioassay of chloroform in Osborne-Mendel rats was re-evaluated for histological evidence of compound-induced cytotoxicity and cell turnover. All rats treated with 1800 ppm (160 mg/kg/day, high-dose group) in the drinking water for 2 years and half the rats treated with 900 ppm (81 mg/kg/day) had mild to moderate changes in proximal convoluted tubules in the mid to deep cortex indicative of chronic cytotoxicity. Tubule alterations specifically associated with chronic chloroform exposure included cytoplasmic basophilia, cytoplasmic vacuolation, and nuclear crowding consistent with simple tubule hyperplasia. Occasional pyknotic cells, mitotic figures in proximal tubules, and prominent karyomegaly of the renal tubule epithelium were present. These alterations were not present in control groups or at the 200-ppm (19 mg/kg/day) or 400-ppm (38 mg/kg/day) dose levels. This new information adds substantially to the weight of evidence that the key events in chloroform-induced carcinogenicity in rat kidney include sustained cellular toxicity and chronic regenerative hyperplasia. PMID- 10696772 TI - Comparative hepatocarcinogenicity of hexachlorobenzene, pentachlorobenzene, 1,2,4,5-tetrachlorobenzene, and 1,4-dichlorobenzene: application of a medium-term liver focus bioassay and molecular and cellular indices. AB - Of the twelve different chlorobenzene isomers, a thorough evaluation of carcinogenicity has only been assessed on monochlorobenzene, 1,2-, and 1,4 dichlorobenzene, and hexachlorobenzene. In the studies presented here, we measured the ability of 1,4-dichlorobenzene (DCB), 1,2,4,5-tetrachlorobenzene (TeCB), pentachlorobenzene (PeCB), and hexachlorobenzene (HCB) to promote glutathione S-transferase pi (GSTP1-1) positive preneoplastic foci formation in rat liver, following diethylnitrosamine (DEN) initiation. The results from these studies show that TeCB, PeCB, and HCB all promote the formation of GSTP1-1 positive foci and that DCB does not. The numbers and area of foci were greatest following HCB promotion, and TeCB and PeCB were approximately equal in their promoting ability. Levels of hepatic CYP1A2, CYP2B1/2, non-focal GSTP1-1, and c fos were measured in response to treatment with the 4 chlorobenzene isomers, as were reduced glutathione (GSH) and oxidized glutathione (GSSG) levels. Results from these studies show that induction of CYP1A2 and CYP2B1/2 have correlation with both the presence and degree of GSTP1-1 foci promotion by the 4 chlorobenzenes. Alterations in GSH and GSSG levels were similar in PeCB- and TeCB treated animals in that GSSG levels were significantly decreased, whereas HCB and DCB did not have this effect, although HCB treatment led to a significant increase in GSH levels. We conclude that induction of CYP1A2 or CYP2B1/2 by chlorobenzene isomers may indicate promotional ability, and that this property might be exploited to predict the hepatocarcinogenicity of other chlorobenzene isomers. PMID- 10696773 TI - Lipopolysaccharide and the trichothecene vomitoxin (deoxynivalenol) synergistically induce apoptosis in murine lymphoid organs. AB - Human exposure to Gram-negative bacterial lipopolysaccharide (LPS) is common and may have an important influence on chemical toxicity. LPS has been shown previously to enhance synergistically the toxicity of trichothecene mycotoxins. Because either of these toxin groups alone characteristically target lymphoid organs at high doses, we evaluated the effects of coexposure to subthreshold doses of Salmonella typhimurium LPS and vomitoxin (VT) administered by intraperitoneal injection and oral gavage of B6C3F1 mice, respectively, on apoptosis in lymphoid tissues after 12-h exposure. The capacity of LPS (0.5 mg/kg body weight) and VT (25 mg/kg body weight) to act synergistically in causing apoptosis in thymus, spleen, and Peyer's patches was suggested by increased internucleosomal DNA fragmentation in whole cell lysates as determined by gel electrophoresis. Following terminal deoxynucleotidyl transferase (TdT)-mediated fluorescein-dUTP nick end-labeling (TUNEL) of tissue sections, a dramatic enhancement of fluorescence intensity indicative of apoptosis was observed in thymus, spleen, Peyer's patches, and bone marrow from coexposed animals as compared to those given the agents alone. Evaluation of hematoxylin and eosin stained tissue sections of treatment mice revealed the characteristic features of lymphocyte apoptosis, including marked condensation of nuclear chromatin, fragmentation of nuclei, and formation of apoptotic bodies in tissues from mice. Combined treatment with VT (25 mg/kg body weight) and LPS (0.5 mg/kg body weight) significantly increased (p<0.05) the amount of apoptotic thymic and splenic tissue as compared to the expected additive responses of mice receiving either toxin alone. When apoptosis was examined in cell suspensions of thymus, spleen, Peyer's patches, and bone marrow by flow cytometry in conjunction with propidium iodide staining, the percentage of apoptotic cells was significantly increased (p<0.05) in cotreatment groups as compared to the additive responses to LPS and VT given alone. The results provide qualitative and quantitative evidence for the hypothesis that LPS exposure markedly amplifies the toxicity of trichothecenes and that the immune system is a primary target for these interactive effects. PMID- 10696774 TI - Analysis of benzo[a]pyrene partitioning and cellular homeostasis in a rat liver cell line. AB - The uptake and subcellular partitioning of benzo[a]pyrene (BaP) were examined in a rat-liver cell line (Clone 9) using confocal and multiphoton microscopy. Following a 16-h treatment, intracellular accumulation of BaP increased with increasing concentration, and cytoplasmic BaP fluorescence reached saturation at 10 microM. Analysis of the kinetics of BaP uptake at this concentration indicated that BaP is rapidly partitioned into all cytoplasmic membranes within several min, although saturation was not reached until 4 h. Based upon the rapid uptake of BaP into membranes, the chronology of changes in gap junction-mediated intercellular communication (GJIC), plasma membrane potential (PMP), and steady state levels of intracellular Ca2+ in relation to the time-course for induction of microsomal ethoxyresorufin-0-deethylase (EROD) activity were examined. EROD activity in Clone 9 cells treated for 16 h increased with increasing concentrations of BaP and reached the highest levels at 40 microM BaP. In addition, kinetic analysis of EROD activity in Clone 9 cells treated with 10 microM BaP indicated that significant induction of EROD activity was not detected before 3 h, and it reached maximal levels by 16 h of treatment at this concentration. Both GJIC and PMP were directly affected by the partitioning of BaP into cellular membranes. The most sensitive index of BaP-induced changes in membrane function was GJIC which revealed a 25% suppression in cells exposed to 0.4 microM BaP for 16 h. Kinetic analysis revealed that suppression of GJIC occurred within 15 min of exposure of cells to 10 microM BaP, whereas significant suppression of PMP was not detected prior to 30-min exposure at this concentration. Elevation of basal Ca2+ level was also detected simultaneously with PMP at this dose. These data suggest that early changes in cellular membrane functions occur prior to detectable induction of EROD activity, although basal metabolic activation of BaP may contribute to these changes. PMID- 10696775 TI - Induction of unscheduled DNA synthesis in primary human urothelial cells by the mycotoxin ochratoxin A. AB - Ochratoxin A (OTA) is a widespread contaminant in human staple food. Exposure of humans to this mycotoxin is a matter of concern because OTA is a known rodent carcinogen. As the urothelium is one target tissue of this mycotoxin, primary cultured human urothelial cells (HUC) from adults and children were used to analyze the induction of unscheduled DNA synthesis (UDS) by OTA. HUC were isolated from the ureters or renal pelves of two nephrectomized adults and of two children with ureteropelvic junction stenosis and cultured under serum-free conditions. After a confluency of 70-80% was reached, cell proliferation was suppressed by arginine-deficient medium (ADM), and UDS was assessed autoradiographically by 3H-thymidine incorporation upon exposure to OTA (10-2000 nM), ethyl methanesulfonate (EMS, 5 mM, positive control), or dimethyl sulfoxide (DMSO, 0.2%, solvent control). In control cultures the level of UDS was low. Exposure to EMS resulted in an induction of UDS (2-to 5-fold compared to control), thus allowing the sensitive detection of repair resulting from induction of DNA lesions in all four specimens, and demonstrating that repair of EMS-induced DNA lesions can take place under the chosen culture conditions. In two HUC cultures derived from adults, a significant induction of UDS was observed in the concentration range of 50-500 nM OTA. The highest fraction of cells in repair (CIR) was found at 50 nM OTA for the HUC from the older male (50% CIR). The maximum response in the other specimens from the adult female and the 7-year old boy were seen at OTA concentrations of 500 and 250 nM, respectively. In contrast to all other specimens, no significant induction of UDS by OTA was found in the HUC cultures derived from an infant's urothelium. Signs of cytotoxicity were observed above 500 nM OTA in all cultures. The varying susceptibility toward OTA observed in vitro may hint at varying predispositions of individuals in vivo. PMID- 10696776 TI - Examination of selected food additives and organochlorine food contaminants for androgenic activity in vitro. AB - In order to produce a reporter gene assay for androgenic chemicals, a constitutive expression vector coding for the human androgen receptor and a reporter construct containing the firefly luciferase coding sequence under transcriptional control of the androgen responsive MMTV promoter were cotransfected into the androgen-insensitive human PC-3 prostate carcinoma cell line and stable transfectants selected. One colony of transfectants, PC-3 LUCAR+, was characterized further. 5alpha-Dihydrotestosterone (DHT) enhanced luciferase activity in a linear fashion for up to 3 days of culture. The Kd for DHT activation was within the range of 25.0-60.0 pM (r2 values >0.95). Flutamide competitively inhibited DHT activation (mean Ki value of 0.89 microM). Progesterone, estradiol, dexamethasone, and hydrocortisone were weak agonists (100-fold less effective than DHT) and diethylstilbestrol was without effect. The effects of organochlorine food contaminants (0, 0.1, 1.0, and 10.0 microM) on luciferase activity in PC-3 LUCAR+ cells were determined after exposure to the chemical for 18 h in the presence and absence of DHT (50 pM). 1,1-dichloro-2,2 bis(p-chlorophenyl)-ethylene (p,p'-DDE) induced luciferase activity in the absence of DHT (100 microM p,p'-DDE equivalent to 50 pM DHT), but in the presence of DHT (50 pM), p,p'-DDE acted antagonistically. 2,3,7,8-Tetrachlorodibenzo-p dioxin, kepone, butylated hydroxyanisole, and butylated hydroxytoluene all partially inhibited activation by DHT (50 pM) but alone had little or no effect. Toxaphene at 10 microM induced luciferase activity in the absence of DHT but decreased cell viability. Alpha- and delta-Hexachlorocyclohexanes (HCH) at 10 microM antagonized the DHT effect, but beta-HCH and gamma-HCH mirex, photomirex, oxychlordane, cis- and trans-nonachlor were without effect. Thus, of the chemicals tested, some interact with the human androgen receptor in vitro as agonists, others as antagonists, and some as partial agonists/antagonists. PMID- 10696777 TI - Comparative evaluation of a 5-day Hershberger assay utilizing mature male rats and a pubertal male assay for detection of flutamide's antiandrogenic activity. AB - A 5-day Hershberger assay utilizing mature male rats and a pubertal male assay were evaluated for the ability to detect antiandrogenic compounds such as flutamide, an androgen receptor antagonist. Six days after the operation, implantation with two silicon capsules containing testosterone (T) (30 mg/capsule) in castrated rats provided the ventral prostate and seminal vesicle weights as well as serum T and luteinizing hormone (LH) levels equivalent to those of the controls (non-castrated, non-implanted rats). Castrated rats implanted with two T-capsules (6 rats/dose) were treated by gavage for 5 days with vehicle (0.5% carboxymethylcellulose) or flutamide (0.15, 0.6, 2.5, or 10 mg/kg/day). Flutamide produced significant decreases in weights of the seminal vesicles and the levator ani plus bulbocavernosus muscles (> or =0.6 mg/kg/day) and ventral prostate (> or =2.5 mg/kg/day), and an increase in serum LH levels (> or =2.5 mg/kg/day), but no changes in serum T levels. When age-matched intact male rats were treated with 10-mg/kg/day flutamide, a significant increase in serum T levels was observed concomitant with a tendency of increased LH. The organ weights were also decreased; however, the changes were less than those in the castrated, T-implanted rats. Immature intact male rats (10 rats/dose) were treated for 20 days with flutamide (0, 0.15, 0.6, 2.5, or 10 mg/kg/day). Flutamide produced significant decreases in weights of the seminal vesicles, ventral prostate, and levator ani plus bulbocavernosus muscles at 2.5 and 10 mg/kg/day. Serum LH levels, but not T levels, were increased at 10 mg/kg/day. Statistical significance of some of these changes was not observed in the 6 animals/dose examined. Our findings support that the Hershberger assay, in the current conditions, is the most sensitive among the assays examined and a useful short-term screening method for the detection of antiandrogenic compounds. PMID- 10696778 TI - Atrazine disrupts the hypothalamic control of pituitary-ovarian function. AB - The chloro-S-triazine herbicides (i.e., atrazine, simazine, cyanazine) constitute the largest group of herbicides sold in the United States. Despite their extensive usage, relatively little is known about the possible human-health effects and mechanism(s) of action of these compounds. Previous studies in our laboratory have shown that the chlorotriazines disrupt the hormonal control of ovarian cycles. Results from these studies led us to hypothesize that these herbicides disrupt endocrine function primarily through their action on the central nervous system. To evaluate this hypothesis, we examined the estrogen induced surges of luteinizing hormone (LH) and prolactin in ovariectomized Sprague-Dawley (SD) and Long-Evans hooded (LE) rats treated with atrazine (50-300 mg/kg/day, by gavage) for 1, 3, or 21 days. One dose of atrazine (300 mg/kg) suppressed the LH and prolactin surge in ovariectomized LE, but not SD female rats. Atrazine (300 mg/kg) administered to intact LE females on the day of vaginal proestrus was without effect on ovulation but did induce a pseudopregnancy in 7 of 9 females. Three daily doses of atrazine suppressed the estrogen-induced LH and prolactin surges in ovariectomized LE females in a dose dependent manner, but this same treatment was without effect on serum LH and prolactin in SD females. The estrogen-induced surges of both pituitary hormones were suppressed by atrazine (75-300 mg/kg/day) in a dose-dependent manner in females of both strains evaluated after 21 days of treatment. Three experiments were then performed to determine whether the brain, pituitary, or both organs were the target sites for the chlorotriazines. These included examination of the ability of (1) the pituitary lactotrophs to secrete prolactin, using hypophyosectomized females bearing pituitary autotransplants (ectopic pituitaries); (2) the synthetic gonadotropin-releasing hormone (GnRH) to induce LH secretion in females treated with high concentrations of atrazine for 3 days; and (3) atrazine (administered in vivo or in vitro) to suppress LH and prolactin secretion from pituitaries, using a flow-through perifusion procedure. In conclusion, the results of these studies demonstrate that atrazine alters LH and prolactin serum levels in the LE and SD female rats by altering the hypothalamic control of these hormones. In this regard, the LE female appeared to be more sensitive to the hormone suppressive effects of atrazine, as indicated by the decreases observed on treatment-day 3. These experiments support the hypothesis that the effect of atrazine on LH and prolactin secretion is mediated via a hypothalamic site of action. PMID- 10696779 TI - In vivo acetylcholinesterase inhibition, metabolism, and toxicokinetics of aldicarb in channel catfish: role of biotransformation in acute toxicity. AB - The carbamate pesticide, aldicarb, demonstrates significant acute toxicity in mammals, birds, and fish through the inhibition of acetylcholinesterase (AChE), and may present high potential for exposure of aquatic organisms during periods of runoff. Toxicity studies have shown that channel catfish are less sensitive to the acute toxic effects of aldicarb than are rainbow trout or bluegill. An earlier in vitro study suggests that the aldicarb resistance in catfish may be related to a low level of bioactivation to the potent aldicarb sulfoxide. The current study examines the toxicity, AChE inhibition, plasma kinetics, and in vivo metabolism of aldicarb in channel catfish. A 48-h LC50 of 9.7 mg/l was determined for juvenile channel catfish. Mortality was accompanied by dramatic loss of brain AChE. Further characterization of tissue-level effects suggests that muscle AChE plays a causal role in mortality. Aldicarb was metabolized in channel catfish to aldicarb sulfoxide, along with the formation of minor hydrolytic products. The toxicokinetics of aldicarb in catfish are bi compartmental with rapid elimination (t1/2 = 1.9 h). Plasma AChE was inhibited in a pattern similar to that of the elimination of total aldicarb-derived compounds. A comparison of aldicarb uptake between catfish and rainbow trout showed no difference in compound absorbed in 24 h. The pattern of in vivo metabolism, however, was quite different between these species. Rainbow trout produce significantly more hydrolytic derivatives and have a 3-fold higher aldicarb sulfoxide to aldicarb ratio at 3 h. These data give strength to the hypothesis that a slower rate of bioactivation in the catfish (vs. rainbow trout) is acting as a protective mechanism against the acute toxicity of aldicarb. PMID- 10696780 TI - Serum alters the uptake and relative potencies of halogenated aromatic hydrocarbons in cell culture bioassays. AB - The effects of many chemicals on cellular processes are governed by their ability to enter the cell, which is in turn a function of the composition of the cell's external environment. To examine this relationship, the effect of serum in cell culture medium on the bioavailability of cytochrome P450 1A (CYP1A)-inducing compounds was determined in PLHC-1 (Poeciliopsis lucida hepatocellular carcinoma) cells. The presence of 10% calf serum in the medium increased the EC50 (effective concentration to achieve 50% maximal response) for induction of ethoxyresorufin O deethylase (EROD) activity by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) 20-fold as compared to treatment in serum-free medium. Measurement of [3H]TCDD uptake and Ah receptor binding indicated that the apparent difference in potencies was a result of decreased bioavailability in the presence of serum, effectively reducing the concentration of TCDD within the cells. Induction of EROD and CYP1A protein in response to treatment with each of three coplanar polychlorinated biphenyls (PCB congeners 77, 126, and 169) was similarly affected by serum, although the magnitude varied among inducers and assays. Relative potencies (calculated as EC50TCDD / EC50PCB) for EROD induction by the three PCBs were significantly higher in the absence of serum. However, serum showed no significant effect on the relative potencies for CYP1A protein induction. These results demonstrate that measured inducing potencies, and relative potencies for EROD induction, by halogenated aromatic hydrocarbons are strongly dependent on the composition of culture medium, which can lead to artificial differences in comparisons among cell types. PMID- 10696781 TI - Differential binding affinities of PCBs, HO-PCBs, and aroclors with recombinant human, rainbow trout (Onchorhynkiss mykiss), and green anole (Anolis carolinensis) estrogen receptors, using a semi-high throughput competitive binding assay. AB - A comparative study was undertaken to assess the ability of 44 polychlorinated biphenyls (PCBs), 9 hydroxylated PCBs (HO-PCBs), and 8 aroclors at concentrations ranging from 1 nM to 10 microM to compete with [3H]17beta-estradiol (E2) for binding to bacterially expressed fusion proteins using a semi-high throughput competitive-binding assay. The fusion proteins consisted of the D, E, and F domains of human (alpha), cloned reptilian (Anolis carolinensis) and recloned rainbow trout (Onchorhynkiss mykiss) estrogen receptors (ER) linked to the glutathione S-transferase (GST) protein. GST-hERalphadef (human), GST-aERdef (reptile) and GST-rtERdef (rainbow trout) fusion proteins exhibited high affinity for E2 with dissociation constants (Kd) of 0.4+/-0.1 nM, 0.7+/-0.2 nM, and 0.6+/ 0.1 nM, respectively. Of the 44 PCBs examined, only PCBs 104, 184, and 188 effectively competed with [3H]E2 for binding to the GST-rtERdef protein with IC50 values ranging from 0.4-1.3 microM. In contrast, these same congeners only caused a 30% displacement of [3H]E2 in GST-hERalphadef and GST-aERdef proteins. Several additional congeners were found to bind to the GST-rtERdef fusion protein, although the degree of interaction varied among congeners. Among the HO-PCBs, 2',3',4',5'-tetrachloro-4-biphenylol and 2,6,2',6'-tetrachloro-4-biphenylol bound to all three fusion proteins with IC50 values ranging from 0.1-0.3 microM. Dimethyl sulphoxide (DMSO) concentrations of 20% significantly increased the ability of PCBs 104, 184, and 188 to compete with [3H]E2 for binding to the GST ERdef fusion proteins, whereas at 20% DMSO, a significant reduction in saturable binding was observed. These results demonstrate that ERs from different species exhibit differential ligand preferences and relative binding affinities for PCBs, which can be dramatically affected by DMSO concentration. PMID- 10696782 TI - Role of the mitochondrial membrane permeability transition (MPT) in rotenone induced apoptosis in liver cells. AB - Rotenone inhibits spontaneously and chemically induced hepatic tumorigenesis in rodents through the induction of apoptosis. However, the mechanism for the induction of apoptosis by rotenone has not been defined. Mitochondrial dysfunction, in particular the induction of the mitochondrial membrane permeability transition (MPT), has been implicated in the cascade of events involved in the induction of apoptosis. Inhibition of the mitochondrial electron transport chain reduces the mitochondrial transmembrane potential (delta(psi)m), which may induce the formation of the mitochondrial permeability transition pore and the subsequent MPT. Fluorescent microscopy of Hoechst 33258-stained WB-F344 cells, a rat-liver cell line, was utilized to examine the effect of the mitochondrial respiratory chain inhibitor, rotenone (0.5-5 microM), atractyloside (5-10 microM), and cyclosporin A (2.5-10 microM) on apoptosis. A time- and concentration-dependent increase in liver cell apoptosis was observed following treatment with rotenone and atractyloside (11.7- and 7.7-fold, respectively, over solvent control). Cotreatment with 7.5- and 10 microM-cyclosporin A for 12 h inhibited the apoptogenicity of 5-microM rotenone treatment. A similar effect was observed following cyclosporin A cotreatment with atractyloside. Rotenone induced a rapid increase in apoptosis (within 20 min of treatment). By 2 h of treatment, the morphological appearance of apoptosis was similar to that observed in cultures treated continuously with rotenone for 12 h. Inhibition studies demonstrated that cyclosporin A prevented apoptosis if the exposure to it occurred prior to the 20-min threshold necessary to induce apoptosis by rotenone. Mitochondrial function was examined by staining with the mitochondrial membrane potential (delta(psi)m)-sensitive fluorochrome, MitoTracker Red (CMXRos) and confirmed utilizing cytofluorometric analysis of DiOC6(3)-stained cells. Rotenone (5.0-microM) and atractyloside (5.0-microM) reduced the percent of CMXRos or DiOC6(3)-positive (delta(psi)m-positive) liver cells within 15 min and throughout the duration of the study (6 h) to approximately 65-80% and 50-80% of control. However, cotreatment with concentrations of cyclosporin A that inhibited the apoptogenicity of rotenone and atractyloside prevented the rotenone- and atractyloside-induced reduction of the delta(psi)m. Therefore, the apoptogenic effect of rotenone and atractyloside appears to occur rapidly (within 20 min) and is irreversible once mitochondrial damage occurs. The inhibition of the rotenone- and atractyloside-induced apoptosis and mitochondrial dysfunction by cyclosporin A suggests the MPT may be involved in the induction of apoptosis by rotenone. PMID- 10696783 TI - Cytotoxicity of 1,2-epoxynaphthalene is correlated with protein binding and in situ glutathione depletion in cytochrome P4501A1 expressing Sf-21 cells. AB - Naphthalene is metabolized by several cytochrome P-450 (CYP) monooxygenases to 1,2-epoxynaphthalene. However, the subsequent interactions of the epoxide with macromolecules in the cells, and the significance of these interactions to cellular injury, are not well characterized. Additionally, CYP1A1, which can metabolize naphthalene to 1,2-epoxynaphthalene, may be induced by a number of xenobiotics. Yet, the in situ interaction between naphthalene and CYP1A1 alone, without the influence of other xenobiotic metabolizing enzymes, has not been examined. Using a model eukaryotic expression system capable of over-expressing recombinant CYP1A1, we found that naphthalene was toxic to cells expressing CYP1A1 in a dose- (LC50: 0.3 mM) and time-dependent (LT50: 12 h) manner. Naphthalene treatment of CYP1A1-expressing cells resulted in a 47% decrease in cellular glutathione (GSH) levels. Pretreatment with ethyl ester GSH, a GSH analog, protected CYP1A1-expressing cells such that viability was 30% greater than for cells treated with naphthalene alone. Cytotoxicity was strongly correlated (r2: 0.96) with covalent binding of cellular proteins. Alkaline permethylation techniques showed that cysteinyl-SH groups of cellular proteins are a nucleophilic target of the epoxide metabolite. These results suggest that, in the absence of other pathways, naphthalene is modified by CYP1A1 to 1,2 epoxynaphthalene, which subsequently binds cellular sulfhydryl groups on proteins and GSH. PMID- 10696784 TI - Mercury chloride activates c-Jun N-terminal kinase and induces c-jun expression in LLC-PK1 cells. AB - In response to various environmental stresses including heavy metals, the c-Jun N terminal kinase (JNK) is phosphorylated and then it phosphorylates c-Jun protein. In the present study, effects of mercury chloride (HgCl2) on JNK signalling pathway were examined in LLC-PK1 cells. When exposed to 10 or 20 microM HgCl2, the level of phosphorylated JNK and the activity of JNK assayed in vitro using glutathione-S-transferase-c-Jun as substrate increased markedly. The level of phosphorylated JNK increased 30 min after HgCl2 exposure and remained elevated even at 8 h. On the other hand, no changes were found in the total amount of JNK protein. Consistent with the activation of JNK, c-Jun proteins phosphorylated on Ser63 and Ser73 were accumulated in cells exposed to HgCl2. Concomitantly, the levels of c-jun mRNA and c-Jun protein were elevated. When compared to other heavy metal compounds such as manganese chloride, zinc chloride, cadmium chloride, and lead chloride, HgCl2 could phosphorylate JNK more markedly. Neither intracellular Ca2+ nor sulfhydryl groups appeared to play a major role in the activation of JNK by HgCl2 exposure in this porcine renal epithelial cell line. PMID- 10696785 TI - Induction of cell-proliferation hormesis and cell-survival adaptive response in mouse hematopoietic cells by whole-body low-dose radiation. AB - Hormesis and a cytogenetic adaptive response induced by low-dose radiation (LDR) have been extensively documented. However, few studies have investigated the induction of an adaptive response by LDR for cell survival in vitro. In the present study, we investigated whether LDR could induce hormesis in hematopoietic cells and the adaptive response of these cells to subsequent high-dose radiation induced cytotoxic effects. Mice were exposed in whole-body to 0 (as control), 0.05, 0.25, 0.50, 0.75, and 1.00 Gy of X-rays. They were killed 12, 24, 48, and 72 h later to observe the stimulating effect of LDR on total bone marrow cells per femur and bone marrow progenitor, colony-forming unit-granulocyte-macrophage (CFU-GM). Exposure to 0.5 Gy of X-rays resulted in significantly stimulating effects on both parameters with a maximum effect at 48 h, showing a cell proliferation hormesis. In the next experiment, mice were irradiated by 0.5 Gy X rays as an adaptive exposure (D1), and 6, 12, 24, 48, and 72 h later, they were exposed to 6 Gy X-rays as a challenging exposure (D2). Forty-eight h after D2, cytotoxic effects were analyzed using peripheral blood cells (red blood cells, white blood cells, and platelets) and bone marrow cells (total bone marrow cells of the femur, and bone marrow progenitors such as CFU-GM and erythroid burst forming unit, BFU-E). An adaptive response to D2-induced cytotoxic effect, named as the cell-survival adaptive response, was found in both peripheral blood cells and bone marrow cells when D1 and D2 exposures were given at intervals of 24-48 h. These results suggested that LDR could induce both cell-proliferation hormesis and cell-survival adaptive response to subsequent high-dose radiation in bone marrow cells. It may be of potential importance, if this phenomenon is confirmed clinically, since it may be applied to reduce the adverse effect of radiotherapy. PMID- 10696786 TI - An assessment of neurotoxicity of aroclors 1016, 1242, 1254, and 1260 administered in diet to Sprague-Dawley rats for one year. AB - As part of a comparative chronic toxicity/oncogenicity study of different Aroclors (1016, 1242, 1254, and 1260), neurotoxicity was assessed in male and female Sprague-Dawley rats using functional observational battery (FOB) and motor activity tests, and histopathologic evaluation of selected nervous system tissues. Doses varied by Aroclor and ranged from 25 to 200 ppm in the diet. Animals were evaluated prior to initiation of dosing and at 13, 26, 39, and 52 weeks of exposure. Clinical signs, body weights, and feed consumption were evaluated weekly. Data analysis of FOB and motor activity results revealed several instances where Aroclor-treated groups were different from control. However, these were considered incidental, as they lacked any consistent dose- or time-related pattern that would suggest Aroclor-induced neurotoxicity. The nonremarkable findings during each of the four assessments were supported by the absence of any treatment-related clinical signs or mortality. Decreased body weight gain was evident in the male 100 ppm Aroclor 1254 dose group and in all female Aroclor 1254 dose groups late in the study (when a linear relationship was assumed between body weight and time), correlating with decreased feed consumption. Although a variety of incidental, spontaneous, degenerative changes were found in nervous tissue evaluated histopathologically, these changes were seen with similar incidence and severity in treated and control groups. No lesions were found that could be attributed to Aroclor-related neurotoxicity. In summary, 52 weeks of exposure to Aroclors 1016, 1242, 1254, or 1260 mixed in the diet did not yield any functional or morphologic changes indicative of PCB induced neurotoxicity. PMID- 10696787 TI - Effects of prenatal aflatoxin B1 exposure on behaviors of rat offspring. AB - The effects of prenatal aflatoxin B1 (AFB) exposure on eight behavioral parameters in Jcl:Wistar rat offspring were assessed. Pregnant rats were injected subcutaneously with 0.3 mg/kg/day of AFB dissolved in dimethylsulfoxide on days 11-14 (Group A) or 15-18 (Group B) of gestation. Controls received the vehicle similarly on days 11-18 of gestation. Before weaning, the offspring were examined using the cliff avoidance response (5 days of age), the negative geotaxis reflex (7 days), and swimming development (6, 8, and 10 days). After weaning, animals were examined using the rotarod test (5 weeks of age), the open field test (6 weeks), a conditioned avoidance learning test (14 weeks), an underwater T-maze test (15 weeks), and a reproduction test (16 weeks). The preweaning offspring in the AFB-A group showed significantly lower success rates than controls in cliff avoidance responses. In swimming development, the offspring in the AFB-A group had significantly lower scores than controls for swimming direction. In the rotarod test, the AFB-A group remained on the rod for a significantly shorter time than the controls at 15 rpm on both the first and second trial days. The avoidance performance of the rats in AFB-A and AFB -B groups was significantly poorer than that of controls. These results indicate that prenatal exposure to AFB produced a delay of early response development, impaired locomotor coordination, and impaired learning ability in the offspring of rats exposed to AFB during middle pregnancy, and the early gestational exposure appears to produce more effects than latter exposure. PMID- 10696788 TI - Postulated human sperm count decline may involve historic elimination of juvenile iodine deficiency: a new hypothesis with experimental evidence in the rat. AB - Human sperm count studies, historic dietary iodination, and an animal model where neonatal goitrogen administration causes unprecedented testis enlargement, together suggest an hypothesis relevant to the postulated fall in human sperm counts. We present the hypothesis with a supporting study extending the model to include iodine deficiency. In a one-generation rat reproduction study, dams were fed an iodine sufficient (control, 200 ppb I) or deficient (low iodine diet [LID], <20 ppb I) diet from prebreeding through weaning, when male offspring were divided into three groups: 1) controls from iodine sufficient dams, 2) neonatal LID (NLID) from the LID dams, fed control diet postweaning, and 3) chronic LID (CLID) from LID dams, fed a moderate LID (40 ppb I) postweaning. F1 males were euthanized on postnatal day (PND) 133+/-1. Each of the three diet groups comprised two subgroups in which testicular parameters were evaluated: 1) daily sperm production (DSP), sperm motility, morphology, and histopathology, and 2) Sertoli cell and round spermatid morphometry. In the first subgroup, NLID and CLID testes weights were 8.5% and 14.0% heavier than their unusually heavy controls (3.921 g; historical control mean approximately 3.5 g), with proportional DSP increases. Sperm motility, morphology, and testis histopathology were unaffected. In the morphometry subgroup, respective increases in NLID and CLID rats included testes weights (+28.6% and +20.3%), Sertoli cells (+24.8% and +23.9%), and round spermatids (+20.4% and +15.8%). The results indicate that neonatal iodine deficiency can significantly increase spermatogenic function in rats, and support our hypothesis concerning human sperm counts. PMID- 10696789 TI - Acute administration of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in pregnant Long Evans rats: association of measured tissue concentrations with developmental effects. AB - Prenatal exposure to TCDD interferes with fetal development at doses lower than those causing overt toxicity in adult animals. Exposure to TCDD during development produces alterations in the reproductive system of the developing pups- delayed puberty and reduced sperm counts in males and malformations in the external genitalia of females. The objectives of this study were to determine maternal and fetal tissue concentrations of TCDD after acute exposure and whether these tissue concentrations can be used to estimate the intensity of the developmental abnormalities reported by other laboratories. Pregnant Long Evans rats received a single, oral dose of 0.05, 0.20, 0.80, or 1.0 microg [3H]-TCDD/kg on gestation day (GD) 15, and maternal and fetal tissue concentrations of TCDD were measured on GD16 and GD21. On GD16, maternal liver contained the greatest amount of TCDD (30-47% administered dose). One day after administration of 0.20 microg TCDD/kg on GD15, there were 13.2 pg TCDD/g present in an individual fetus. This concentration is associated with delayed puberty and decreased epididymal sperm counts in male pups as well as malformations in the external genitalia of females. For the responses studied, tissue concentration measured during a critical period of gestation adequately predicts the intensity of the response. In addition, there was a strong correlation between fetal body burden and maternal body burden on GD16. A dose of 0.05 microg TCDD/kg resulted in maternal body burdens of 30.6+/-3.1 and 26.6+/-3.1 ng TCDD/kg on GD16 and GD21, respectively. In conclusion, low-level TCDD exposure during the perinatal stage of life can produce adverse effects within the developing pups and that tissue concentration measured during a critical period is the appropriate dose metric to predict adverse reproductive and developmental effects. PMID- 10696790 TI - The reproductive and developmental toxicity of the antifungal drug Nyotran (liposomal nystatin) in rats and rabbits. AB - Nyotran is a liposomally encapsulated i.v. formulation of the antifungal polyene nystatin. This drug was evaluated in a series of reproductive toxicity studies, according to the guidelines outlined by the International Conference on Harmonization (ICH). A fertility and early embryonic development study (SEG I) and a prenatal and postnatal development (SEG III) study were conducted in rats, and embryo-fetal development (SEG II) studies were conducted in rats and rabbits. Nyotran was administered iv in all studies. In SEG I and SEG III, rats were administered daily doses of 0.5, 1.5, or 3.0 mg/kg Nyotran. In both studies, parental mortality and toxicity in the 3.0 mg/kg dose group necessitated the lowering of the high dose to 2.0 mg/kg/day. Parental toxicity, in the form of decreased body weights, decreased food consumption, and piloerection were also observed at the 1.5 mg/kg/day dose level in the SEG I and SEG III studies. Despite the parentally toxic doses in the SEG I study, there was no effect of Nyotran on F0 male or female fertility or early embryonic development of F1 offspring. In the SEG III study, lactational body weights of the F1 generation were decreased at all Nyotran dose levels. There was no effect on pre-wean developmental landmarks, but post-wean development was affected by Nyotran administration at all dosage levels. Preputional separation was delayed in the 1.5 and 3.0/2.0 mg/kg/day F1 offspring, auditory startle function was decreased in F1 females at all dose levels, and motor activity was decreased in male F1 offspring at all dose levels. However, there were no treatment-related effects on the subsequent mating of the F1 generation and resulting F2 offspring. In SEG II studies, rats and rabbits were also administered 0.5, 1.5, or 3.0 mg/kg/day of Nyotran during gestation. The high dose in these SEG II studies was not lowered, as the maternal animals were able to tolerate the shorter duration of dosing. Maternal effects in rabbits were observed only in the high-dose group and were limited to decreased food consumption and decreased absolute and relative liver weight. Decreased food consumption in high-dose dams and clinical weight loss in some animals at the mid- and high-dose levels evidenced maternal toxicity in rats. Nyotran did not have any effect on Caesarian section parameters in either rats or rabbits and no effect on the incidence of fetal malformations in rabbits. A statistically significant increase in mild hydrocephaly, observed in 4 rat fetuses, was seen at the highest dose level of 3.0 mg/kg/day. The biological significance and relationship to Nyotran treatment of this finding is not clear. This finding may represent a change in the background incidence or a change in the pattern of responsiveness of this strain of rat fetus to the test chemical. Toxicokinetic data were also collected in the SEG II rabbit and rat studies for comparison to human exposures. In both species, systemic exposure to the nystatin at effective antifungal concentrations was demonstrated. The systemic exposures in rats and rabbits were, however, considerably less than have been reported in humans administered clinical doses of 2 or 4 mg/kg/day Nyotran. Thus, humans tolerate higher dosages and systemic exposures of Nyotran relative to rats and rabbits and there is no margin of safety in either dosage level or systemic exposure to drug. Given this lack of a margin of safety and the effects on postnatal development in F1 rats, caution should be exercised when using this drug in females of childbearing potential. PMID- 10696791 TI - The effect of heterogeneity of lung structure on particle deposition in the rat lung. AB - Differences in particle deposition patterns between human and rat lungs may be attributed primarily to their differences in breathing patterns and airway morphology. Heterogeneity of lung structure is expected to impact acinar particle deposition in the rat. Two different morphometric models of the rat lung were used to compute particle deposition in the acinar airways: the multiple-path lung (MPL) model (Anjilvel and Asgharian, 1995, Fundam. Appl. Toxicol. 28, 41-50) with a fixed airway geometry, and the stochastic lung (SL) model (Koblinger and Hofmann, 1988, Anat. Rec. 221, 533-539) with a randomly selected branching structure. In the MPL model, identical acini with a symmetric subtree (Yeh et aL, 1979, Anat. Rec. 195, 483-492) were attached to each terminal bronchiole, while the respiratory airways in the SL model are represented by an asymmetric stochastic subtree derived from morphometric data on the Sprague-Dawley rat (Koblinger et al., 1995, J. Aerosol. Med. 8, 7-19). In addition to the original MPL and SL models, a hybrid lung model was also used, based on the MPL bronchial tree and the SL acinar structure. Total and regional deposition was calculated for a wide range of particle sizes under quiet and heavy breathing conditions. While mean total bronchial and acinar deposition fractions were similar for the three models, the SL and hybrid models predicted a substantial variation in particle deposition among different acini. The variances of acinar deposition in the MPL model were consistently much smaller than those for the SL and the hybrid lung model. The similarity of acinar deposition variations in the two latter models and their independence on the breathing pattern suggests that the heterogeneity of the acinar airway structure is primarily responsible for the heterogeneity of acinar particle deposition. PMID- 10696792 TI - Lack of differential sensitivity to cholinesterase inhibition in fetuses and neonates compared to dams treated perinatally with chlorpyrifos. AB - Pregnant Sprague-Dawley rats were exposed to chlorpyrifos (CPF; O,O-diethyl-O [3,5,6-trichloro-2-pyridinyl] phosphorothioate) by gavage (in corn oil) from gestation day (GD) 6 to postnatal day (PND) 10. Dosages to the dams were 0 (control), 0.3 (low), 1.0 (middle) or 5.0 mg/kg/day (high). On GD 20 (4 h post gavage), the blood CPF concentration in fetuses was about one half the level found in their dams (high-dose fetuses 46 ng/g; high-dose dams 109 ng/g). CPF oxon was detected only once; high-dose fetuses had a blood level of about 1 ng/g. Although no blood CPF could be detected (limit of quantitation 0.7 ng/g) in dams given 0.3 mg/kg/ day, these dams had significant inhibition of plasma and red blood cell (RBC) ChE. In contrast, fetuses of dams given 1 mg/kg/day had a blood CPF level of about 1.1 ng/g, but had no inhibition of ChE of any tissue. Thus, based on blood CPF levels, fetuses had less cholinesterase (ChE) inhibition than dams. Inhibition of ChE occurred at all dosage levels in dams, but only at the high-dose level in pups. At the high dosage, ChE inhibition was greater in dams than in pups, and the relative degree of inhibition was RBC approximately plasma > or = heart > brain (least inhibited). Milk CPF concentrations were up to 200 times those in blood, and pup exposure via milk from dams given 5 mg/kg/day was estimated to be 0.12 mg/kg/day. Therefore, the dosage to nursing pups was much reduced compared to the dams exposure. In spite of exposure via milk, the ChE levels of all tissues of high-dosage pups rapidly returned to near control levels by PND 5. PMID- 10696793 TI - Mercury induces regional and cell-specific stress protein expression in rat kidney. AB - Cells respond to physiologic stress by enhancing the expression of specific stress proteins. Heat-shock proteins (hsps) and glucose-regulated proteins (grps) are members of a large superfamily of proteins collectively referred to as stress proteins. This particular stress-protein response has evolved as a cellular strategy to protect, repair, and chaperone other essential cellular proteins. The objective of this study was to evaluate the differential expression of four hsps in the renal cortex and medulla during experimental nephrotoxic injury using HgCl2. Male Sprague-Dawley rats received single injections of HgCl2 (0.25, 0.5, or 1 mg Hg/kg, i.v.). At 4, 8, 16, or 24 h after exposure, kidneys were removed and processed for histopathologic, immunoblot, and immunohistochemical analyses. Nephrosis was characterized as minimal or mild (cytoplasmic condensation, tubular epithelial degeneration, single cell necrosis) at the lower exposures, and progressed to moderate or severe (nuclear pyknosis, necrotic foci, sloughing of the epithelial casts into tubular lumens) at the highest exposures. Western blots of renal proteins were probed with monoclonal antibodies specific for 4 hsps. In whole kidney, Hg(II) induced a time- and dose-related accumulation of hsp72 and grp94. Accumulation of hsp72 was predominantly localized in the cortex and not medulla, while grp94 accumulated primarily in the medulla but not cortex. The high, constitutive expression of hsp73 did not change as a result of Hg(II) exposure, and it was equally localized in cortex and medulla. Hsp90 was not detected in kidneys of control or Hg-treated rats. Since hsp72 has been shown involved in cellular repair and recovery, and since Hg(II) damage occurs primarily in cortex, we investigated the cell-specific expression of this hsp. Hsp72 accumulated primarily in undamaged distal convoluted tubule epithelia, with less accumulation in undamaged proximal convoluted-tubule epithelia. These results demonstrate that expression of specific stress proteins in rat kidney exhibits regional heterogeneity in response to Hg(II) exposure, and a positive correlation exists between accumulation of some stress proteins and acute renal cell injury. While the role of accumulation of hsps and other stress proteins in vivo prior to or concurrent with nephrotoxicity remains to be completely understood, these stress proteins may be part of a cellular defense response to nephrotoxicants. Conversely, renal tubular epithelial cells that do not or are unable to express stress proteins, such as hsp72, may be more susceptible to nephrotoxicity. PMID- 10696794 TI - Metabolism and toxicity of trichloroethylene and S-(1,2-dichlorovinyl)-L-cysteine in freshly isolated human proximal tubular cells. AB - Trichloroethylene (Tri) caused modest cytotoxicity in freshly isolated human proximal tubular (hPT) cells, as assessed by significant decreases in lactate dehydrogenase (LDH) activity after 1 h of exposure to 500 microM Tri. Oxidative metabolism of Tri by cytochrome P-450 to form chloral hydrate (CH) was only detectable in kidney microsomes from one patient out of four tested and was not detected in hPT cells. In contrast, GSH conjugation of Tri was detected in cells from every patient tested. The kinetics of Tri metabolism to its GSH conjugate S (1,2-dichlorovinyl)glutathione (DCVG) followed biphasic kinetics, with apparent Km and Vmax values of 0.51 and 24.9 mM and 0.10 and 1.0 nmol/min per mg protein, respectively. S-(1,2-dichlorovinyl)-L-cysteine (DCVC), the cysteine conjugate metabolite of Tri that is considered the penultimate nephrotoxic species, caused both time- and concentration-dependent increases in LDH release in freshly isolated hPT cells. Preincubation of hPT cells with 0.1 mM aminooxyacetic acid did not protect hPT cells from DCVC-induced cellular injury, suggesting that another enzyme besides the cysteine conjugate beta-lyase may be important in DCVC bioactivation. This study is the first to measure the cytotoxicity and metabolism of Tri and DCVC in freshly isolated cells from the human kidney. These data indicate that the pathway involved in the cytotoxicity and metabolism of Tri in hPT cells is the GSH conjugation pathway and that the cytochrome P-450-dependent pathway has little direct role in renal Tri metabolism in humans. PMID- 10696795 TI - Western blot analysis for nitrotyrosine protein adducts in livers of saline treated and acetaminophen-treated mice. AB - The hepatic centrilobular necrosis produced by the analgesic/antipyretic acetaminophen correlates with metabolic activation of the drug leading to its covalent binding to protein. However, the molecular mechanism of the toxicity is not known. Recent immunohistochemical analyses using an antinitrotyrosine antiserum indicated that nitrotyrosine protein adducts co-localized with the acetaminophen-protein adducts in the centrilobular cells of the liver. Nitration of proteins is believed to occur by peroxynitrite, a substance formed by the rapid reaction of superoxide with nitric oxide. Nitric oxide and superoxide may be formed by activated Kupffer cells or by other cells. Because we were unable to successfully utilize the commercial antiserum in Western blot analyses of liver fractions, we developed a new antiserum. With our antiserum, liver fractions from saline-treated control and acetaminophen-treated mice were successfully analyzed for nitrated proteins. The immunogen for this new antiserum was synthesized by coupling 3-nitro-4-hydroxybenzoic acid to keyhole limpet hemocyanin. A rabbit immunized with this adduct yielded a high titer of an antiserum that recognized BSA nitrated with peroxynitrite. Immunoblot analysis of nitrated BSA indicated that nitrotyrosine present in a protein sample could be easily detected at levels of 20 pmoles. Immunohistochemical analyses indicated that nitrotyrosine protein adducts were detectable in the centrilobular areas of the liver. Immunoblot analysis of liver homogenates from both saline-treated and acetaminophen-treated mice (300 mg/kg) indicate that the major nitrotyrosine protein adducts produced have molecular weights of 36 kDa, 44 kDa, and 85 kDa. The 85-kDa protein stained with the most intensity. The hepatic homogenates of the acetaminophen- treated mice showed significantly increased levels of all protein adducts. PMID- 10696796 TI - Acute, nontoxic cadmium exposure inhibits pancreatic protease activities in the mouse. AB - Toxic effects of cadmium on liver, kidney, lung, and testes have been well established in experimental animals and in cell model systems. However, little is known about the effect of cadmium on pancreas, though the pancreas has been reported to accumulate high concentrations of cadmium. Therefore, in this study we examined the effects of cadmium on the pancreas of mice. A single sc injection of 1 mg Cd/kg to mice had no obvious toxic effects on the liver, kidney, and pancreas at both 1 and 5 days after cadmium treatment. Within the pancreas, however, the activities of trypsin, chymotrypsin, and carboxypeptidase A were significantly decreased at 1 day after cadmium treatment, whereas the activity of carboxypeptidase B was not changed. All pancreatic enzyme activities returned to the control levels by 5 days after cadmium treatment. The concentrations of cadmium in pancreas were very similar at 1 and 5 days after cadmium treatment, indicating a stable deposition of the metal. The concentration of zinc in pancreas was markedly increased at 5 days after cadmium treatment. In order to more fully examine the inhibitory effects of cadmium on these protease activities in pancreas, the direct effects of cadmium on purified proteases were studied in vitro. Contrary to the results in vivo, cadmium increased the activity of purified trypsin in a concentration-dependent manner. Consistent with the in vivo results, the activity of purified carboxypeptidase A was decreased by cadmium treatment in a concentration-dependent fashion in vitro. The activities of chymotrypsin and carboxypeptidase B did not change by the cadmium exposure in vitro. The enhanced activity of trypsin by cadmium was returned to the control levels by subsequent treatment with EDTA, indicating that enhancement was reversible. In addition, the zinc normally contained in purified carboxypeptidase A and carboxypeptidase B was released by the cadmium treatment. These results indicate that cadmium inhibits protease activities within the pancreas in vivo at doses that do not induce overt hepatic, renal, or pancreatic toxicity. Based on in vitro study, the decreases seen in trypsin and chymotrypsin activities might be based on indirect effects of cadmium, whereas the decreases in carboxypeptidase A are probably due to the direct inhibition by the metal. PMID- 10696797 TI - Functional alterations in the olfactory bulb of the staggerer mutant mouse. AB - Putative alterations of the functional activity in the staggerer mutant mouse olfactory bulb neuronal network have been studied by recording odor induced evoked field potentials (EFP) in the mitral cells layer. In standard conditions, the main feature observed in mutants was a significant increase in latency preceding the functional response of the mitral cells to the odorant. In these animals, all parameters of the average EFP were widely modified when compared with those recorded in wild mice. Amplitudes and most of the duration of the EFP phases were significantly decreased. Functional alterations were discussed according to the structural disorganization previously described in staggerer mutant mouse olfactory bulb. PMID- 10696798 TI - Loss of tyrosine hydroxylase immunoreactivity in dendrites is a sensitive index of kainic acid-induced damage in rat substantia nigra neurons in vivo. AB - An early indicator of damage to substantia nigra dopamine neurons in vitro is loss of dendrites that precedes loss of the cell body. To investigate dendritic damage in vivo, rats were treated for 1 day or 1 week with kainic acid (KA; 5 or 10 mg/kg i.p.), the brain fixed and substantia nigra (SN) dopamine neurons and their dendrites labeled using an antibody to tyrosine hydroxylase (TH). KA (10 mg/kg) produced seizures initially and resulted in significant loss of TH immunoreactivity in dendrites of dopamine neurons 1 week, but not 1 day, after a single injection. Daily injections of 5 mg/kg KA, which did not produce seizures, resulted in more extensive dendritic damage. The findings indicate that loss of dendritic staining is a sensitive index of damage to SN dopamine neurons in vivo. PMID- 10696799 TI - Glucose metabolism in the rat frontal cortex recovered without the recovery of choline acetyltransferase activity after lesioning of the nucleus basalis magnocellularis. AB - We measured the cerebral metabolic rate of glucose (CMRglc) by using positron emission tomography (PET) with [18F]fluorodeoxyglucose (FDG) and the choline acetyltransferase (ChAT) activity at 3 days and 3 months after destruction of the nucleus basalis magnocellularis (NBM). Although the frontal ChAT activity remained 20% lower than that of controls even at 3 months post-lesioning, the frontal CMRglc, which was reduced by 40% at 3 days, returned to normal at 3 months, namely CMRglc recovered with time without the recovery of ChAT activity with time. Since glucose metabolism reflects mainly presynaptic neuronal activity, we speculate that presynaptic rearrangement may have some relation to the recovery of CMRglc. PMID- 10696800 TI - Two distinctive antinociceptive systems in rats with pathological pain. AB - A common obstacle in clinical management of pathological pain is the poor response to opioid analgesics. We now report that delta9-tetrahydrocannabinol (delta9-THC)-induced antinociception remained effective in rats with pathological pain. The selective central cannabinoid receptor antagonist SR141716A, but not the generic opioid receptor antagonist naloxone, blocked the delta9-THC antinociception. Moreover, there is no cross-tolerance between the antinociceptive effects of morphine and delta9-THC in pathological pain states. The results indicate that delta9-THC antinociception is both effective and independent of opioid receptors in rats with pathological pain. Thus, the cannabinoid analgesic system may be superior to opioids in alleviating intractable pathological pain syndromes. PMID- 10696801 TI - The intra-spine electric force can drive vesicles for fusion: a theoretical model for long-term potentiation. AB - We have estimated the intensity of intra-spine electric fields triggered by stimulation of excitatory spine synapses. We show that this electric force can cause fast electrophoretic movement of negatively charged vesicles which bring new postsynaptic receptors and membrane for insertion during the induction of long-term potentiation (LTP). Due to the direction of an intra-spine electric field, movement of vesicles is electrophoretically directed along the longitudinal spine axis towards the spine head. Thus, the number of fused vesicles may be proportional not only to the increased calcium concentration within the spine head during the induction of LTP but also to the magnitude of electric force which drives vesicles towards the postsynaptic membrane. PMID- 10696802 TI - Developmental expression of excitatory amino acid transporter 5: a photoreceptor and bipolar cell glutamate transporter in rat retina. AB - Excitatory amino acid transporter 5 (EAAT5) is a retina-specific glutamate transporter which has an associated chloride conductance. Thus it is comparable in its functional properties to the glutamate transport systems previously described in photoreceptors and some bipolar cells. We have raised antibodies to the carboxyl- and amino-terminal regions of EAAT5. Labeling for both of these antisera was developmentally regulated: weak labeling appeared in photoreceptors around P7; by P10 strong labeling was present in photoreceptors and by P21 a population of bipolar elements were also weakly labeled. In adult retinae both antisera heavily immunolabeled all photoreceptors as well as a heterogeneous population of bipolar cell somata and their proximal axonal processes: synaptic terminals of these cells were also labeled after partial proteolytic digestion of the tissues. The positions and morphology of these terminals suggests that they are the terminals of both rod and cone rod bipolar cells. We conclude that in rat retina, EAAT5 is a photoreceptor and bipolar cell glutamate transporter. PMID- 10696803 TI - Dynamic mediolateral activation of the pyramidal cell population in human somatosensory 3b area can be visualized by magnetic recordings. AB - Dynamic mediolateral activation in the pyramidal cell population of the human somatosensory 3b area was studied non-invasively. Somatosensory evoked fields (SEFs) were recorded over the hand area contralateral to the right median nerve stimulation at the wrist. Localization of an equivalent current dipole (ECD) for N20m primary cortical response was successively calculated for periods of 2.4 ms around N20m peak where the signal-to-noise ratio exceeded 100 and the spatial resolution was approximately 0.3 mm. The ECD moved toward the anterior, lateral and inferior direction for a distance of 8.7 mm at a propagation velocity of 3.6 m/s. This direction was orthogonal to the orientation of apical dendrites of the pyramidal neurons in area 3b and parallel to the surface of the posterior bank of the central sulcus. These findings suggest that the sequential mediolateral activation of the pyramidal cells in the somatosensory cortex is mediated by horizontal connections running parallel to the cortical surface. PMID- 10696804 TI - Genetic polymorphisms of human melatonin 1b receptor gene in circadian rhythm sleep disorders and controls. AB - Recent studies suggest that melatonin 1b (Mel1b) receptor, as well as melatonin 1a (Mel1a) receptor, is involved in the modulation of circadian rhythms in mammals. Mutational analysis was performed in the entire coding region of the human Mel1b receptor gene using genomic DNA from sleep disorder subjects. We have identified two missense mutations, G24E and L66F. However, neither is likely to be associated with sleep disorders in our study population. One of the subjects with non-24-h sleep-wake syndrome carries missense mutations in both the Mel1a and Mel1b receptor genes. PMID- 10696805 TI - Artificial lighting conditions and melatonin alter motor performance in adult rats. AB - Entrained circadian rhythms may modulate many behavioral activities of animals and humans. In the present study, we examined whether lighting conditions and melatonin treatment participate behaviorally in the entrainment of circadian rhythms in the rodent. In experiment one, Sprague-Dawley rats were introduced to the Rotorod test apparatus at nighttime or daytime and either with the lights on (4 lux) or in the dark. During nighttime tests, the exposure of rats to dark or light condition did not alter mean rev./min or length of times spent on the Rotorod. Interestingly, during daytime tests, animals exposed to light condition displayed significantly reduced mean rev./min (7.95 +/- 1.68), as well as length of time on the Rotorod (41.07 +/- 3.45 s) compared with their performance in the dark condition (mean rev./min, 11.16 +/- 1.52; length of time spent on the Rotorod, 66.94 +/- 6.15 s). In experiment two, treatment with melatonin (1.5 mg/kg, orally administered at 1 h prior to testing) in animals introduced to the daytime test with exposure to light condition, restored the rev./min (12.90 +/- 1.26) and the time spent on the Rotorod (63.21 +/- 2.73 s) to near normal levels. Thus, we demonstrated here that exposure of nocturnal animals to their preferred dark condition and treatment with melatonin could enhance motor coordination. PMID- 10696806 TI - Autoradiographic distribution of peripheral benzodiazepine receptors in the retina of the albino rabbit, Lepus cunicula. AB - The distribution of peripheral benzodiazepine receptors (PBRs) in the retina of the albino rabbit, Lepus cunicula, was studied by autoradiography using [3H] PK11195, a isoquinoline carboxamide, as a tracer. Autoradiograms obtained by directly placing the slides containing the retina sections on tritium-sensitive film provide evidence for the presence of PBRs in rabbit retina. Furthermore, the dark field examination of photomicrographs taken from autoradiograms showed two dense horizontal bands corresponding to the outer and inner photoreceptor segments, and to the inner plexiform layer. The retinal regions where [3H] PK11195 binding was more dense are rich in mitochondria, suggesting that as in other neuronal tissues, retinal PBRs are involved in the mitochondrial activity. PMID- 10696807 TI - Exploring a critical parameter of timing in the mouse cerebellar microcircuitry: the parallel fiber diameter. AB - Since the conduction velocity of the parallel fibers is a critical parameter for a theory of timing in the cerebellar cortex, we set out to quantify the diameter of these axons on an ultrastructural level. The overall mean of the fiber diameter was 0.18 microm. Our results confirm that the parallel fibers of the upper molecular layer are significantly thinner than those of the lower layers. Nevertheless, the difference of about 0.02 microm determined by this study is surprisingly small. In addition, the distribution of the fiber diameters of the upper layers differed slightly, but significantly from a normal distribution, partly on account of a positive skew and a positive kurtosis excess. In summary, the results show that there are fewer differences between the parallel fibers of different levels of the molecular layer than previously assumed and that these differences do not contradict a theory of timing in the cerebellar cortex. PMID- 10696808 TI - Micturition and defensive behaviors are controlled by distinct neural networks within the dorsal periaqueductal gray and deep gray layer of the superior colliculus of the rat. AB - Electrical stimulation of the dorsal periaqueductal gray (DPAG) or the deep gray layer of the superior colliculus (DGSC) of rats placed in an open-field elicited either a display of tense immobility, accompanied by exophthalmus and/or defecation and micturition, or running and jumping responses. Threshold curves of each response were obtained for each structure by the logistic fitting of accumulated response frequencies. DPAG and DGSC threshold curves were compared by likelihood-ratio coincidence tests. The output of micturition was significantly higher following the stimulation of DPAG (P < 0.0005). In contrast, no differences were found for the remaining responses. These data support previous studies in anaesthetized cats suggesting the critical involvement of DPAG in the control of micturition. Furthermore, they also suggest that topographically distinct neural networks within the DPAG and DGSC control micturition and the other defensive behaviors. PMID- 10696809 TI - Muscle sympathetic nerve activity during handgrip and post-handgrip muscle ischemia after exposure to simulated microgravity in humans. AB - To examine the effect of 6 degrees head-down bed rest (HDBR) on vasomotor sympathetic responses to isometric forearm exercise, 16 healthy male subjects aged 20-36 years performed voluntary isometric handgrip (HG) at 30% of maximal voluntary contraction until fatigue, followed by 2 min of post-handgrip muscle ischemia (PHGMI) with 250 mmHg of cuff inflation, before and after 14 days of HDBR. Time to fatigue and maximal voluntary HG force were not affected by HDBR. Pre-exercise baseline muscle sympathetic nerve activity (MSNA, measured by microneurography), heart rate (measured by electrocardiogram) and mean blood pressure (measured by Portapres) increased after HDBR. Increases in MSNA were similar during HG but significantly lower during PHGMI (P < 0.01) after HDBR. Responses of heart rate and mean blood pressure during HG and PHGMI were not affected by HDBR. These results suggest that the magnitude of muscle metaboreflex during isometric forearm exercise might be attenuated after 14 days of simulated microgravity. PMID- 10696810 TI - Effects of repeated phencyclidine treatment on serotonin transporter in rat brain. AB - Phencyclidine (PCP) is known to be an inhibitor of serotonin (5-HT) uptake and to increase serotonergic activity. The development of tolerance to serotonergic stereotyped behaviors induced by repeated PCP treatment and changes of 5-HT transporters were examined. Backpedalling was significantly reduced in frequency following 14 days PCP treatment (7.5 mg/kg per day). Furthermore, repeated PCP treatment decreased the equilibrium dissociation constant (Kd) of [3H]paroxetine binding to 5-HT transporters in whole brain excluding the cerebellum without any change of maximum number of binding sites (Bmax). Single treatment with PCP failed to change binding parameters. These results indicate that repeated PCP treatment causes tolerance in serotonergic stereotyped behavior and increases affinity of 5-HT transporters for [3H]paroxetine binding. The increased affinity of 5-HT transporters could represent compensatory responses to chronic inhibition of 5-HT uptake by PCP. PMID- 10696811 TI - Activity-dependent enhancement of miniature excitatory postsynaptic current amplitude and its modulation by protein kinase C in cultured rat sympathetic neurons. AB - A high K+ solution increased the frequency of miniature excitatory postsynaptic currents (MEPSCs) and intracellular Ca2+ concentration ([Ca2+]i) in cultured rat sympathetic neurons. Repetition or continuation of high K+ treatment increased MEPSC amplitude, acetylcholine-induced currents and the averaged rise in [Ca2+]i per single MEPSC. The enhancement of MEPSCs lasted over 30 min and was inhibited by intracellular BAPTA and phorbol ester, but not by atropine. The results suggest that repeated Ca2+ entry through the channel pore of nicotinic acetylcholine receptor enhances the efficacy of its opening and the activation of protein kinase C inhibits the enhancement. PMID- 10696812 TI - Reduced activation of midline frontal areas in human elderly subjects: a contingent negative variation study. AB - Contingent negative variation (CNV) was recorded from electrodes F7, F3, Fz, F4, F8, T7, C3, Cz, C4, T8, P7, P3, Pz, P4 and P8 in 19 young (mean age: 23 years) and 15 elderly (mean age: 66 years) healthy right-handed subjects, using a S2 choice paradigm. Young subjects showed early peak negativity shortly after the warning stimulus over mid-frontal areas, whereas for the remaining electrodes the negativity increased continuously. The amplitude of the early CNV was selectively reduced in elderly subjects over midline but not lateral frontal areas. We conclude that the activation of frontal midline areas as pre-supplementary motor area or anterior cingulate might be impaired in higher age. PMID- 10696813 TI - No influence of adrenergic receptor polymorphisms on schizophrenia and antipsychotic response. AB - The adrenergic system plays an important role in psychiatric disorders such as depression and schizophrenia. Antagonism of the adrenergic receptor subtypes alpha1A and alpha2A has been found to have an antipsychotic effect. Genetic mutations in these receptors could be related to the alterations in the adrenergic system observed in psychiatric patients and to failure to respond to drug antagonism. We have studied one polymorphism in the alpha1A-adrenergic receptor (Arg492Cys) and two polymorphisms in the promoter region of the alpha2A adrenergic receptor (-1291-C/G and -261-G/A) in a sample of clozapine-treated schizophrenic patients and controls. No clear differences were observed between the different groups suggesting that these polymorphisms did not play an important role in the aetiology of the disorder or in determining antipsychotic response. PMID- 10696814 TI - A magnetoencephalographic study of brain activity related to recognition memory in healthy young human subjects. AB - Neural activity associated with recognition memory was investigated using magnetencephalography (MEG) in healthy young subjects. At sensor sites overlying frontal and temporoparietal cortices, magnetic evoked fields (MEFs) revealed a difference between studied and unstudied stimuli, which onset about 400 ms following stimulus onset and lasted about 600 ms. MEG yielded reliable source information revealing the activity of three independent dipoles, located in the right medial temporal lobe (MTL), the right inferior frontal and the left inferior parietal cortices. Our findings suggest that neural activity underlying recognition memory from both superficial and deep brain structures can be monitored by MEG. PMID- 10696815 TI - Increase of fragmented DNA transport in apical dendrites of gerbil CA1 pyramidal neurons following transient forebrain ischemia by mild hypothermia. AB - Mild hypothermia (38 degrees C) accelerated transport of fragmented DNA in apical dendrites of the gerbil CA1 pyramidal neurons and increased dendrite-terminal fragmented DNA pooling in the apoptotic process following transient forebrain ischemia. The specific DNA fragmentation after the ischemic insult in gerbil hippocampus was examined by in situ nick-end-labeling method, and fluorescence DNA detection technique by DAPI was also performed. There is a precise temperature dependence for the migration of fragmented DNA from nuclei into apical dendrites of CA1 pyramidal cells during apoptosis following transient forebrain ischemia. Increase of fragmented DNA pooling is highly temperature sensitive, occurring at 38 degrees C, while at 39 degrees C there is a marked decrease in DNA pooling. PMID- 10696816 TI - Crisis phenomena after stroke reflected in an existential perspective. AB - The study gains a deeper understanding of crisis phenomena emerging after stroke and focuses on these phenomena viewed in an existential perspective. Ten stroke victims narrated their experiences of their new life situation in open-ended interviews, conducted during the first few months after discharge. The participants were analyzed using a phenomenological-hermeneutic approach. This analysis disclosed an extremely distressing situation related to the individuals' struggle to manage in various dimensions of life. The phenomena were intertwined in a complex way and the critical interpretation involved a transcendence to the existential dimension of life. The situation was metaphorically depicted as "a struggle in the darkness" in a "boundary situation," where the issues ultimately touched on life and death, fate and future, meaning and meaninglessness. The study indicates the significance of existential issues pervading the seemingly concrete struggle to manage life after stroke. PMID- 10696817 TI - Grandparents in the United States and the Republic of China: a comparison of generations and cultures. AB - Grandparent behaviors in the United States and the Republic of China are examined to identify curriculum themes for helping them adjust to their changing role. The 3,286 non-consanguineous subjects included Chinese (n = 751), African Americans (n = 777), Caucasian Americans (n = 1,086), and Mexican Americans (n = 672). Analyses were performed using 1) Generation with three levels (grandparent, parent, and grandchild) and 2) Culture with four levels (Chinese, African American, Caucasian American, and Mexican American). The results revealed significant differences in perceptions about grandparents across cultures as well as between generations within cultures. All three generations reported grandparent strengths and needs. Specific guidelines and curriculum topics are recommended for education to support grandparent development. PMID- 10696818 TI - Stereotypes of the elderly in magazine advertisements 1956-1996. AB - The globalization of American culture is increasing as various media target an international market. This article reports the results of a study examining trends in the stereotyping of the elderly in print advertisements appearing from 1956 to 1996 in U.S. magazines. Results show that the percentage of elderly portrayals in print ads has decreased. There has been relatively little overall stereotyping of elderly, with only 4 percent of the sample depicting negative stereotypes. Nevertheless, there has been an increasing percentage of negative stereotypes and a decreasing percentage of positive stereotypes. Results are analyzed in relationship to marketing trends and the social impact on aging. PMID- 10696819 TI - Personality change in adulthood: loci of change and the role of interpersonal process. AB - This study examined personality change in two domains--dispositional tendencies (emotion traits) and characteristic adaptations (views of self) over eight years in a sample of older adults (M = 63.4). Stability coefficients for anxiety, depression, interest, anger, anger-in, anger-out, and aggression ranged from .47 to .75; only anger-out showed significant change over the eight years. On the other hand, respondents reported moderate changes in perspectives, goals, personality, feelings, and ways of relating and the ratings of outside informants were significantly correlated with self-reports of change for all but goals. Personality change was associated with positive and negative interpersonal life events of an intimate nature such as marriage, divorce, and death of loved ones that took place over the past eight years, and was not associated with other high and low points in lives involving careers, changes in residence, and more distant social relationships. PMID- 10696820 TI - Effects of four trichothecene mycotoxins on activation marker expression and cell proliferation of human lymphocytes in culture. AB - Four trichothecene mycotoxins--the type A trichothecenes T2-toxin and diacetoxyscirpenol and the type B trichothecenes nivalenol and deoxynivalenol- were studied. The effects of these mycotoxins on the expression of the sequentially expressed activation markers CD69, CD25, and CD71 and on proliferation of human lymphocytes were studied in culture with a duration of up to 72 h. All the examined toxins affected activation marker expression in a similar way. After 6 h, the CD69 expression was lower in exposed cultures compared to controls. After 24 and 48 h of exposure, an increased frequency of cells expressing CD69 was found in exposed cultures, indicating a delay in downregulation of CD69 expression. Stimulation of CD25 expression was observed for doses below the IC50 value, while suppression was found for higher doses. The pattern was different from that detected for CD69 expression, in that an increased expression of CD25 never occurred after exposure to the highest concentration of the toxin, and in that no stimulatory effects were found after 48 h of exposure, indicating that the response was inhibited and not delayed. The effects of toxin exposure on CD71 expression were in many respects similar to the effects on CD25 expression. We conclude that the trichothecene mycotoxins investigated in this study inhibited the cell cycle in a similar way and exert their main antiproliferative action rather early in the cell cycle, before or in conjunction with CD25 expression. PMID- 10696821 TI - The role of Ca2+ and hemodynamics in the action of diltiazem on hepatic energy metabolism. AB - Diltiazem causes vasoconstriction in the liver when present at high concentrations, an action that is strictly Ca2+-dependent. Diltiazem is also active on energy metabolism. This toxic action could be partly a consequence of hemodynamic effects. In the absence of Ca2+, the hemodynamic effects are no longer present and, consequently, Ca2+-free experiments are useful for distinguishing between hemodynamics-dependent and hemodynamics-independent effects. The experimental system used was the hemoglobin-free perfused rat liver from fed and fasted rats. Diltiazem was infused at various concentrations in the presence and absence of Ca2+. Several metabolic parameters were measured: lactate and pyruvate production (glycolysis), glycogenolysis, oxygen uptake, gluconeogenesis, and the cellular levels of lactate, pyruvate, glucose, AMP, ADP, and ATP. The effects of diltiazem can be divided into three groups: (1) Effects that are strictly dependent on the Ca2+-mediated hemodynamic action. This group comprises inhibition of oxygen uptake at all concentrations (50-500 micromol/L) inhibition of lactate, pyruvate, and glucose release at high concentrations; the decrease in cellular ATP; the increase in cellular AMP; and the cellular accumulation of glucose and lactate. (2) Effects that are independent of the hemodynamic action. The most relevant effect of this type is inhibition of gluconeogenesis. (3) Effects that are influenced by Ca2+ but are independent of the hemodynamic effects. This is the typical case of lactate and glucose release from endogenous glycogen, whose stimulation by low diltiazem concentrations is more pronounced in the presence of Ca2+ than in its absence. PMID- 10696822 TI - Modulation of a fibrotic process induced by transforming growth factor beta-1 in dermal equivalents. AB - Why initially normal wound healing sometimes shifts toward an impaired cicatrization is poorly understood. Collagen gels with incorporated fibroblasts constitute valuable in vitro models to study mechanisms of connective tissue reorganization. Such 1-week-old, partially contracted normal dermal equivalents were treated with concentrations of TGF-beta1 ranging from 1 to 10 ng/ml. The cytokine was applied in a single dose or four times at regular intervals, over a 2-week period. Dose-dependent activation of fibroblasts was observed after treatment. The cytokine induced a myofibroblastic transformation of dermal cells, significantly enhanced the process of dermal contraction, and stimulated synthesis of such proteins as cellular fibronectin, tenascin and smooth-muscle actin. Our approach is more informative than models using pathological or pretreated dermal cells, since it demonstrates newly induced modulation of fibrotic transformation in an initially normal dermal equivalent. This in vitro assay will enable us to study mechanisms involved in the shift between normal and impaired fibrotic transformation during wound healing. PMID- 10696823 TI - Fas-mediated apoptosis is attenuated in preneoplastic GST-P-positive hepatocytes isolated from diethylnitrosamine-treated rats. AB - Previous reports have indicated that apoptosis is selectively decreased in enzyme altered foci (EAF) in the livers of rats treated with a carcinogen. Here we have investigated the effects of an anti-Fas antibody (anti-Fas Ab) on EAF cells in vitro. Hepatocytes were isolated from rats treated repeatedly with diethylnitrosamine (DEN), whose livers contained glutathione S-transferase P (GST P)-positive EAF. Subsequently, primary cultures of GST-P-positive and GST-P negative hepatocytes were established and exposed to anti-Fas Ab. Anti-Fas Ab (4 microg/ml) preferentially induced apoptosis in GST-P-negative cells. Furthermore, GST-P-positive cells were shown to be resistant to p53-mediated apoptosis. We conclude that EAF hepatocytes are resistant to Fas-mediated apoptosis in vitro. This lack of response may explain the selective decrease in apoptosis in EAF. PMID- 10696825 TI - Differential gliotoxicity of organotins. AB - On the basis of reports that astrocytes play an important role in the neurotoxicity of trimethyltin (TMT), we investigated the sensitivity of astrocytes to TMT and compared it to triethyltin (TET), a neurotoxic analog with a different in vivo specificity. The gliotoxicity of these two compounds was further compared to that of tributyltin (TBT) and triphenyltin (TPT), two purportedly nonneurotoxic organotin compounds. The time and concentration components of organotin toxicity were determined by measuring lactate dehydrogenase (LDH) release and formazan production from dimethylthiazolyldiphenyltetrazolium bromide (MTT). A TMT concentration of 100 micromol/L did not elevate extracellular LDH until 48 h after exposure, while signs of toxicity were not seen at 72 h for concentrations less than 10 micromol/L. Extracellular LDH activity increased 24 h after exposure to concentrations of TET, TBT, and TPT as low as 2.5 micromol/L. TMT was the only organotin to produce a delayed cytotoxicity, requiring both higher concentrations and more time to produce discernible toxicity. In contrast with TBT and TPT, the toxicity of the two neurotoxic organotins (TMT and TET) produced an early increase in MTT reduction. The distinct pattern of toxicity for TMT does not explain its selective in vivo toxicity, but the lack of sensitivity of astrocytes to this organotin also does not rule out more subtle changes in these cells that could disrupt normal glial/neuronal interactions. PMID- 10696824 TI - Photosensitization of lymphoblastoid cells with phthalocyanines at different saturating incubation times. AB - Photodynamic therapy of cancer is a promising treatment based on the tumor specific accumulation of photosensitizers followed by irradiation with visible light which induces tumor cell death. The effect of different preincubation times on the photosensitization efficiency of the phthalocyanines AlPc and AlPcS4 was investigated in lymphoblastoid CCRF-CEM cells under conditions that allow maximal uptake of the sensitizers. First, the time course for the uptake of AlPcS4 and AlPc by CCRF-CEM cells and by the pheochromocytoma PC12 cells was compared. The uptake of AlPcS4 by CCRF-CEM cells was not significantly different after 6 h or 24 h incubation, but the photosensitization efficiency of the phthalocyanine was much higher when a 24 h preincubation period was used, with a fluence rate of 5 mW/cm2. However, for a fluence rate of 10 mW/cm2, the photosensitization efficiency of AlPcS4 was almost completely independent of the preincubation time (6 h vs. 24 h) with the phthalocyanine. When the cells were preincubated with 1 micromol/L AlPc for 10 min or 6 h, which allows the same accumulation of sensitizer by the cells, no significant effect of the incubation time on the photodynamic inactivation of CCRF-CEM cells was observed, with fluence rates of 5 mW/cm2 or 10 mW/cm2, for different light doses. Confocal fluorescence microscopy studies did not reveal differences in the localization of the phthalocyanines after maximal uptake was reached. The results show that the preincubation time with AlPcS4, after the maximal uptake is reached, affects cell growth to an extent depending on the fluence rate used, and this effect was not due to a major redistribution of the sensitizer during incubation. However, this was not observed when AlPc was used. PMID- 10696826 TI - Dexamethasone represses 3,5,3'-triiodothyronine-stimulated expression of intercellular adhesion molecule-1 in the human cell line ECV 304. AB - The effect of the thyroid hormone L-3,5,3'-triiodothyronine (T3) on the expression of ICAM-1, a cell surface glycoprotein playing a pivotal role in inflammatory responses, was investigated. In ECV 304 cells, T3 (30 nmol/L) markedly increased ICAM-1 protein expression, with a peak after 24 h of treatment. ECV 304 human cells express both alpha1 and beta1 T3 receptors. In an attempt to understand the molecular mechanisms leading to the induction of the ICAM-1 gene by T3, we have studied the effects of a synthetic glucocorticoid, dexamethasone, and of a sesquiterpene lactone, parthenolide, on this T3 stimulated expression of ICAM-1. Our results demonstrate a repressive effect of dexamethasone and parthenolide on the expression of the protein ICAM-1 stimulated by T3. Both anti-inflammatory compounds interfere with this T3-mediated pathway in a dose-dependent manner. PMID- 10696827 TI - The responsiveness of the Hamilton Depression Rating Scale. AB - In clinical studies of antidepressants, the Hamilton Depression Rating Scale (HAMD) total score has been the gold standard instrument for establishing and comparing the efficacy of new treatments. However, the HAMD is a multidimensional measure, which may reduce its ability to detect differences between treatments, in particular, changes in core symptoms of depression. Two meta-analyses were conducted to compare the responsiveness of the HAMD total score with several published unidimensional subscale scores based upon core symptoms of depression. The first compared the above instrument's ability to detect differences between fluoxetine and placebo across eight studies involving over 1600 patients. The second analysis involved four studies and over 1200 patients randomized to tricyclic antidepressants and placebo. In both meta-analyses, the unidimensional core subscales outperformed the HAMD total score at detecting treatment differences. The implications of this on sample sizes and power for clinical studies will be discussed. In fact, studies based on the observed effect sizes from the core subscales would require approximately one-third less patients than studies based on the HAMD total score. Effect sizes from each individual HAMD item will also be presented to help explain the differences in responsiveness between the scales. PMID- 10696828 TI - Discrepancy between subjective and objective ratings for negative symptoms. PMID- 10696829 TI - Evidence of a schizotypy subtype in OCD. AB - OCD patients represent a heterogeneous mix of clinical phenotypes, likely reflecting a wide range of genetic vulnerabilities. In other medical illnesses, neurobiologically-based traits with a genetic component that are associated with the target disorder have been successfully used to detect patients with a specific genetic liability to disease. The overlap between symptoms of OCD and Schizophrenia suggested that schizotypal traits could have the potential to distinguish a relatively homogeneous subtype of OCD. We obtained schizotypy scores for 119 affected adult probands who met lifetime criteria for DSM-IV OCD. Five subscales from the Structured Interview of Schizotypy were used to assess ideas of reference, suspiciousness, magical thinking, illusions and psychotic like thought. Selected for their obvious face validity with the cardinal signs of schizophrenia, Cronbach's alpha suggested that these subscales also provided a reliable measure of positive sign schizotypy (0.83). Fifty percent of our OCD sample had mild to severe positive schizotypy signs. t- and chi2 tests of significance suggested seven variables that distinguished OCD patients with schizotypy, including earlier age of onset, greater number of comorbid diagnoses and increased rates of learning disability, aggressive and somatic obsessions and counting and arranging compulsions. Three of these seven variables, including learning disabilities, counting compulsions and history of specific phobia, significantly increased the odds of schizotypy among patients with lifetime OCD. These findings enhanced the validity of the schizotypy construct in OCD. Whether this schizotypy subtype can distinguish a subgroup of patients with relatively homogeneous genetic characteristics waits further investigation. PMID- 10696830 TI - Elevated concentration of N-CAM VASE isoforms in schizophrenia. AB - Neural cell adhesion molecule (N-CAM) is a cell recognition molecule, four major isoforms (180, 140, 120, and 105-115 kDa) of which are present in brain. N-CAM has several roles in cellular organization and CNS development. Previously we have found an elevation in CSF N-CAM 120 kDa in the CSF of patients with schizophrenia, bipolar disorder, and depression. We now report an increase in the variable alternative spliced exon (VASE), a 10 amino acid sequence inserted into the fourth N-CAM domain, in the CSF of patients with schizophrenia, but not in bipolar disorder or depression. VASE-immunoreactive (VASE-ir) bands were measured in CSF from patients with schizophrenia (n = 14), bipolar disorder I (n = 7), bipolar disorder II (n = 9), unipolar depression (n = 17) and matched controls (n = 37) by Western immunoblotting. Three VASE-ir bands were distinguished in lumbar CSF corresponding to heavy (165 kDa), medium (155 kDa) and low (140 kDa) MW. A logarithmic transformation was applied to the VASE protein units and analyzed with a MANOVA. There was a 51% and 45% increase in VASE heavy (p = 0.0008) and medium (p = 0.04) MW protein, respectively, in patients with schizophrenia as compared with normal controls. Current neuroleptic treatment in patients with schizophrenia had no effect on CSF VASE concentrations. VASE concentration correlated significantly with behavioral ratings in patients with schizophrenia but not affective disorders. Thus, VASE immunoreactivity is increased in schizophrenia but not in affective disorders. These results provide further evidence of an abnormality of N-CAM protein in chronic schizophrenia and suggest differences between schizophrenia and affective disorders in regulation of N-CAM. PMID- 10696831 TI - Effects of (S)-ketamine on striatal dopamine: a [11C]raclopride PET study of a model psychosis in humans. AB - Administration of the N-methyl-D-aspartate (NMDA) antagonist S-ketamine in normals produces a psychosis-like syndrome including several positive and negative symptoms of schizophrenic disorders (Abi-Saab WM, D'Souza DC, Moghaddam B, Krystal JH. The NMDA antagonist model for schizophrenia: promise and pitfalls. Pharmacopsychiatry 1998;31:104-109). Given the clinical efficacy of dopamine (DA) D2 receptor antagonists in the treatment of positive symptoms, it is conceivable that S-ketamine-induced psychotic symptoms are partially due to a secondary activation of dopaminergic systems. To date, animal and human studies of the effects of NMDA antagonists on striatal DA levels have been inconsistent. The present study used positron emission tomography (PET) to determine whether a psychotomimetic dose of S-ketamine decreases the in vivo binding of [11C]raclopride to striatal DA D2 receptors in humans (n = 8). S-ketamine elicited a psychosis-like syndrome, including alterations in mood, cognitive disturbances, hallucinations and ego-disorders. S-ketamine decreased [11C]raclopride binding potential (BP) significantly in the ventral striatum ( 17.5%) followed by the caudate nucleus (-14.3%) and putamen (-13.6%), indicating an increase in striatal DA concentration. The change in raclopride BP in the ventral striatum correlated with heightened mood ranging from euphoria to grandiosity. These results provide evidence that the glutamatergic NMDA receptor may contribute to psychotic symptom formation via modulation of the DA system. PMID- 10696832 TI - Minor physical anomalies in schizophrenia: cognitive, neurological and other clinical correlates. AB - Minor physical anomalies (MPAs) are minor congenital malformations which are found with significantly increased frequency among both patients with schizophrenia and their siblings, suggesting the effect of early developmental disturbance in their families. The aim of this study was to explore the relationship between these signs of early dysmorphogenesis and cognitive and neurological dysfunction in the patients and their siblings as well as the clinical characteristics of the patients. Sixty patients with schizophrenia, 21 nonpsychotic siblings and 75 normal comparison subjects were studied. Increased rates of cognitive and neurological dysfunction and high MPA scores were found in both the patients and their siblings. High rates of MPAs were not significantly related to cognitive or neurological dysfunction in the patients or siblings, or to premorbid history or other characteristics of the clinical disease process in the patients. These results suggest that MPAs are possibly markers of general early neuromaldevelopment rather than markers of a specific cognitive/neurological or clinical subtype of schizophrenia. PMID- 10696833 TI - An EEG approach to the neurodevelopmental hypothesis of schizophrenia studying schizophrenics, normal controls and adolescents. AB - Based on an integrative brain model which focuses on memory-driven and EEG state dependent information processing for the organisation of behaviour, we used the developmental changes of the awake EEG to further investigate the hypothesis that neurodevelopmental abnormalities (deviations in organisation and reorganisation of cortico-cortical connectivity during development) are involved in the pathogenesis of schizophrenia. First-episode, neuroleptic-naive schizophrenics and their matched controls and three age groups of normal adolescents were studied (total: 70 subjects). 19-channel EEG delta-theta, alpha and beta spectral band centroid frequencies during resting (baseline) and after verbal stimuli were used as measure of the level of attained complexity and momentary excitability of the neuronal network (working memory). Schizophrenics compared with all control groups showed lower delta-theta activity centroids and higher alpha and beta activity centroids. Reactivity centroids (centroid after stimulus minus centroid during resting) were used as measure of update of working memory. Schizophrenics showed partial similarities in delta-theta and beta reactivity centroids with the 11-year olds and in alpha reactivity centroids with the 13-year olds. Within the framework of our model, the results suggest multifactorially elicited imbalances in the level of excitability of neuronal networks in schizophrenia, resulting in network activation at dissociated complexity levels, partially regressed and partially prematurely developed. It is hypothesised that activation of age- and/or state-inadequate representations for coping with realities becomes manifest as productive schizophrenic symptoms. Thus, the results support some aspects of the neurodevelopmental hypothesis. PMID- 10696834 TI - Abnormal visual event-related potentials in obsessive-compulsive disorder without panic disorder or depression comorbidity. AB - Visual event-related potentials and spline map topography during a discriminative response task (DRT) were studied in 8 obsessive-compulsive disorder (OCD) patients without comorbidity for panic disorder or depression and in 12 age matched controls. In the DRT task (like in a go/no-go task) the subject had to press a button when the target stimuli appeared and had to retain the response when the non-target stimulus appeared (vertical bars were intermixed with an equal probability of horizontals). OC patients had greater N1 latency than controls and their N1 and P3 amplitude was larger for the target stimuli, but not for non-target stimuli. In the normals, non-target stimuli (no-go task) produced a larger activation than target stimuli (go task). In the OCD patients the target stimuli produced the same large activation as the non-target. These findings are consistent with theories that consider OCD to be an attentional disorder deriving from a misallocating of cognitive resources. Moreover, spline map topography confirmed that P3 hyperactivation is localised principally on the frontal lobes. PMID- 10696835 TI - Reduced body fat in long-term followed-up female patients with anorexia nervosa. AB - We aimed to evaluate both body composition and serum leptin levels in females with a past history of anorexia nervosa (AN) adjusted for their current body mass index (BMI). Twenty-three females with a past history of AN were followed-up 10 years after inpatient treatment and compared to 23 female controls of a similar age range matched for BMI on a one to one basis. Serum leptin levels were assessed and percent body fat (%BF) was determined via bioelectric impedance analysis. Differences of both %BF and leptin levels between cases and controls were tested under the hypothesis that cases have lower %BF and lower serum leptin levels than the controls. %BF was indeed lower in the cases compared to the controls (p < 0.05). However, differences in leptin levels between both groups just failed significance (p = 0.051). We conclude that body composition differs between long-term followed-up patients with AN and BMI- and gender-matched controls. Based on the finding that the former patients reported being more physically active, we assume that the higher physical activity levels in recovered patients with AN underlie the lower %BF. PMID- 10696836 TI - Premedication, preparation, and surveillance. AB - Once again the staggering variation in IV sedation practice between different countries is highlighted. This year the "to sedate or not sedate" debate focuses on colonoscopy. Several papers on the use of Propofol are reviewed. It remains this authors' opinion that propofol is an anaesthetic agent to be used by (or at least in the presence of) an anaesthetist. Informed consent and the question of what to do if a patient withdraws consent halfway through the procedure are discussed. Predictably further recent papers on the relative merits of midazolam and diazepam are presented plus another report on the use of flumazenil in the recovery period. The use of 3% hydrogen peroxide solution to aid the visualization of acutely bleeding gastro-duodenal lesions is presented in two papers along with a discussion of its possible mode of action. The use of antispasmodics to aid colonoscopy is further discussed: this year concentrating on the use of hyoscyamine sulphate (as opposed to hyoscine butylbromide, the preferred agent in the UK). The patients receiving hyoscyamine sulphate had significantly shorter caecal intubation times, better sedation and easier colonic insertion. The "downside" was drug-induced tachycardia and the authors caution against the widespread use of this drug until this situation is further clarified. The subject of hypoxaemia at the time of gastroscopy, colonoscopy and ERCP was reviewed last year and further papers are presented in which the incidence of various levels of hypoxia are given. In anaesthetic circles it would be considered totally unacceptable to allow a patient's oxygen saturation to fall below 85 %, and yet we continue to have papers reporting its incidence. This level of desaturation is potential dangerous and the routine use of supplemental oxygen would greatly reduce this unneccessary risk to patients. PMID- 10696837 TI - Reflux disease and Barrett's esophagus. AB - Gastroesophageal reflux disease (GERD) is still an important clinical problem. Continuing efforts are being made to establish a classification of the condition that would allow improved communications for both clinical and research purposes. In medical treatment, the trends are toward proton-pump inhibitor therapy at all stages of GERD, calling into question the role of endoscopy for tailoring individual therapy. Arguments against the use of H. pylori eradication therapy in GERD have gained importance. Surgeons are continuing to report excellent results with fundoplication, but careful studies are needed to prove whether antireflux surgery is really capable of saving costs, as its proponents claim. Barrett's esophagus is still a topic of lively interest. Since there is no method of primary prevention, endoscopy has a crucial role in detecting affected patients and guiding them toward one of the various surveillance strategies--which are not yet clearly established. The debate over short-segment Barrett's esophagus, and especially over "microscopic" Barrett's esophagus (at the squamocolumnar junction), has not yet been resolved. However, there is now less doubt that GERD is a condition associated with a substantially higher risk for the development of esophageal adenocarcinoma. Given this risk of malignant transformation, there is continuing competition between different ablation techniques; however, careful data from much larger populations will be needed before ablation reaches the stage of broad clinical application. Until specific guidelines become available, patients with Barrett's esophagus should receive endoscopic follow-up until it can be ascertained which individuals are at risk for cancer and require ablation of Barrett's mucosa. PMID- 10696838 TI - Ulcers and gastritis. AB - Once again this year, developments in the field of ulcers and gastritis have been entirely dominated by findings relating to Helicobacter pylori. However, interest in H. pylori can be expected to decline, since the prevalence of the infection is rapidly decreasing in the developing world - to the point that many gastroduodenal ulcers are now unrelated to H. pylori A further reason for declining interest is disappointment regarding the role of H. pylori in dyspepsia. The expected reduction in the endoscopic workload produced by screening for H. pylori or cagA positivity has so far not occurred. The exact role of the damage caused by nonsteroidal anti-inflammatory drugs in H. pylori infected stomachs remains controversial. Equally controversial is the magnitude of the increased reflux symptomatology and reflux disease observed after H. pylori infection has been cured in patients with ulcer disease. Eradication therapy is largely dominated by proton-pump inhibitor triple therapies, although the efficacy of these is declining. Bismuth triple therapy and quadruple therapy continue to be valid alternatives. PMID- 10696839 TI - Ulcers and nonvariceal bleeding. AB - Although peptic ulcer continues to be the commonest cause of acute upper gastrointestinal bleeding, obscure haemorrhage from the Dieulafoy malformation and haemobilia must be considered, and may be amenable to endoscopic therapy. The patients who are at the highest risk of rebleeding and death are elderly, in shock at presentation, and have major stigmata of recent haemorrhage (SRH). The endoscopist must identify SRH, and identification may be made easier by washing the area with hydrogen peroxide. The natural history of SRH has been defined. There is wide interobserver variation in the interpretation of SRH, and there is probably therefore little value in the endoscopist describing subtle appearances. Although the value of endoscopic haemostatic therapy is established, it has still not been taken up by all institutions. Endoscopic injection of fibrin glue into the bleeding ulcer is a logical and relatively easy approach, and a systematic histological study of resected ulcers has shown that this does not adversely affect the ulcer healing process. Thermal therapies such as argon plasma coagulation and the heater probe have comparable efficacy. Although a combination of injection and thermal treatments may be logical, there are only trends suggesting that this is better than monotherapy. Nevertheless, the gold probe continues to be used in clinical practice. Experiments in an animal model of gastric bleeding suggest that the gold probe is effective, and that the version with a wide-gauge needle is best. Haemoclips may stop acute upper gastrointestinal bleeding from a range of sources. Patients who rebleed after initial endoscopic haemostasis have a tenfold increase in the risk of death. An important study from Hong Kong suggests that repeat endoscopic treatment after rebleeding has comparable morbidity and mortality to a policy of urgent surgery without endoscopic repeat intervention. PMID- 10696840 TI - Colon polyps and cancer. AB - Several important studies on screening for colorectal neoplasia were published in 1998-99, including average-risk as well as high-risk groups, and different strategies such as endoscopy, fecal occult blood tests, and other more specific markers of neoplasia. Some of these studies included the use of interventions other than initial screening and polypectomy; a few dealt with diagnostic methods in symptomatic patients, and the new diagnostic tool of virtual colonoscopy was highlighted by several authors. Endoscopic treatment for large adenomas and early colorectal cancer has become more widespread, but clearly it has limitations. Current discussion of the topic of colorectal polyps and cancer is largely based on the concept of the adenoma-carcinoma sequence, which is thought to be the most probable pathogenesis for colorectal cancer. PMID- 10696841 TI - Inflammatory bowel disease. AB - This article reviews important papers on inflammatory bowel disease published between May 1998 and June 1999. It does not review every aspect of treatment, but focuses on the effects of anti-tumor necrosis factor antibodies on the inflammatory lesions. The new information summarized includes: the role of bacteria and the modulating effects of probiotics; the frequency of appendiceal orifice inflammation in ulcerative colitis; progress in imaging based on endoscopic ultrasonography, magnetic resonance imaging, and leukocyte scintigraphy; frequency and treatment of massive hemorrhage, viral superinfection, and persistent perineal sinus; and the pathogenesis, detection, and treatment of dysplasia and cancer. PMID- 10696842 TI - Small-bowel endoscopy. AB - During the last year, several interesting publications have further confirmed the role of enteroscopy in clinical practice. Of particular interest have been various articles concerning the use of two-way enteroscopy in large series of patients with obscure gastrointestinal bleeding or with radiological abnormalities. Reports continue to appear describing a high incidence of gastroduodenal or colonic lesions that were missed or misinterpreted during previous upper and lower endoscopies in patients with recurrent obscure bleeding. By contrast, there have been few outcome studies on patients with obscure bleeding, and the conclusions reached are not in full agreement. Other important publications have stressed the value of enteroscopy in selected cases of chronic unexplained diarrhea, for diagnosing small-bowel lesions caused by nonsteroidal anti-inflammatory drugs, and in identifying small-bowel tumors. In addition to numerous reports on intraoperative enteroscopy, the results of initial experience with laparoscopically assisted enteroscopy have also been reported. PMID- 10696843 TI - Endoscopic ultrasonography. AB - Endosonography is primarily a diagnostic imaging modality, but new therapeutic applications are being developed. Miniprobe technology (ultrasound catheter probes) has led to new clinical applications. The staging of gastrointestinal cancers remains the major accepted indication for endosonography. Other conditions, such as chronic pancreatitis, portal hypertension, and inflammatory bowel diseases, are also being evaluated with endosonographic imaging. PMID- 10696845 TI - Minimal standard terminology in digestive endoscopy. PMID- 10696844 TI - Diagnostic laparoscopy. AB - Diagnostic laparoscopy continues to play an important role in the accurate evaluation of patients with abdominal disorders. Combined with laparoscopic ultrasound, it is highly accurate in the staging of intra-abdominal malignancies, and it is superior to transcutaneous ultrasonography and computed tomography. Other important applications include the evaluation of patients with acute and chronic abdominal pain, acute abdomen, peritonitis, and blunt and penetrating abdominal trauma. Laparoscopy now rests firmly in the hands of surgeons. The majority of last year's papers originated from departments of surgery; papers on laparoscopy in hepatic disorders are sorely missing in this year's review. PMID- 10696846 TI - A new device to facilitate the endoscopic treatment of oesophageal variceal bleeding. PMID- 10696847 TI - Abdominal pain and hyperamylasaemia due to gastric pancreatic heterotopia: endoscopic diagnosis and treatment. PMID- 10696848 TI - Acute renal insufficiency after colon cleansing. PMID- 10696849 TI - Treatment of persisting bilioma using endoscopic retrograde cholangiography and doxycycline. PMID- 10696850 TI - Novel use of Hegar's dilators in gastrostomy tube reinsertion. PMID- 10696851 TI - Does atopic disease start in foetal life? PMID- 10696852 TI - Systemic corticosteroid treatment for seasonal allergic rhinitis: a common but poorly documented therapy. PMID- 10696853 TI - Inhaled and nasal corticosteroids: safety aspects. PMID- 10696854 TI - Nasal provocation test in the diagnosis of natural rubber latex allergy. AB - BACKGROUND: Natural rubber latex (NRL) allergy in workers using rubber gloves has been an occupational health problem for the last 10 years. In the case of the occupational agents, clinical history may be far from conclusive; hence, appropriate provocation should be carried out. The objective was to evaluate the usefulness of the nasal challenge test in the diagnosis of allergic rhinitis in subjects occupationally exposed to NRL. METHODS: A single-blind, placebo controlled study was conducted in 16 nurses with respiratory symptoms (bronchial asthma and/or rhinitis) related to NRL exposure as well as positive skin prick test (SPT) response to NRL. The controls were nine nurses with asthma and/or perennial rhinitis unrelated to NRL exposure; six atopic patients not occupationally exposed to NRL, with asthma and/or perennial rhinitis; and six healthy subjects. All the controls had negative results of SPT with NRL. Patients with a history of anaphylaxis or positive results of RAST to NRL were not considered in the study. The "nasal pool" technique was used to evaluate the cellular response and changes in protein level and ECP concentration in nasal washings after topical provocation with allergen or placebo. RESULTS: A significant increase was noted in eosinophil and basophil number, albumin/total protein ratio, and ECP level only in NRL SPT-positive patients subjected to nasal challenge with NRL. Neither bronchial nor systemic reactions were found after the nasal provocation with NRL. CONCLUSIONS: The nasal challenge test appears to be useful for diagnosing occupational rhinitis in NRL-sensitized patients. PMID- 10696855 TI - Allergenicity of major cow's milk and peanut proteins determined by IgE and IgG immunoblotting. AB - BACKGROUND: Specific IgG antibodies are frequently observed in food-allergic patients. However, the allergen-fraction specificity of IgG antibodies in relation to IgE antibodies is not well defined. Our aim was to determine the IgE and IgG antibody profile to major cow's milk and peanut-antigen fractions in food allergic patients and tolerant individuals. METHODS: Sera were collected from 10 patients allergic to cow's milk and 10 patients allergic to peanuts, as well as from 20 control subjects. Cow's milk and peanut proteins were migrated on SDS PAGE and immunoblotted for IgE, IgG, and IgG4 antibodies. Food-specific IgE concentrations were measured by CAP System FEIA, and IgG and IgG4 concentrations by ELISA. RESULTS: In food-allergic children, similar fraction-specific IgE, IgG, and IgG4 antibody-binding profiles to the major cow's milk or peanut antigens were found. In nonallergics, the presence of fraction-specific IgG antibodies was mostly dependent on regular ingestion of the food. The presence of specific antibody on immunoblots correlated with their quantitative measurement. The mean value for specific IgE in cow's milk-allergic patients was 450 +/- 1,326 IU/ml, and 337 +/- 423 IU/ml in peanut-allergic patients. Specific IgG antibody values in milk-allergic patients were not different (median OD 1.5, range 0.3-2.3) from controls (median OD 1, range 0.2-1.8). However, in peanut-allergic patients, IgG concentrations were significantly higher than in controls (OD 1.2 [0.5-1.3] vs 0.5 [0.3-0.7]; P< 0.01). CONCLUSIONS: Similar fraction-specific IgE and IgG antibody profiles in allergic individuals suggest a common switching trigger involving both isotypes. Intrinsic allergenicity might explain identical IgG antibody fraction specificity in nonallergics and in allergics. The presence of IgG antibodies in nonallergics was related to regular ingestion of the food. PMID- 10696856 TI - Diagnosis of allergic bronchopulmonary aspergillosis (ABPA) in cystic fibrosis. AB - BACKGROUND: The diagnosis of allergic bronchopulmonary aspergillosis (ABPA) in cystic fibrosis (CF) patients may be difficult to establish because ABPA shares many characteristics with coexisting atopy or other lung infections in these patients. This study aimed to evaluate the sensitivity and specificity of various paraclinical parameters in the diagnosis of ABPA in patients with CF. METHODS: Accumulated data from a 5-year period in 238 CF patients were used to divide patients into two groups designated the ABPA group (n=26) and the non-ABPA group (n = 35). Patients in both groups were colonized with Aspergillus fumigatus (Af.), but only the ABPA group consistently demonstrated specific IgE antibodies and specific precipitins. Patients without A. fumigatus colonization were not assigned to either of these groups (n = 177). By this selection as the true diagnosis, 10 patients were selected from the ABPA group and 10 patients from the non-ABPA group. RESULTS: The groups were comparable as to age, sex, lung function (P=0.6), and presence of chronic Pseudomonas aeruginosa infection (P>0.1). No significant difference between the groups in unspecific atopic parameters such as eosinophil count (P=0.9) or eosinophil cationic protein (ECP) in sputum, plasma, or serum (P=0.9, P=0.59, and P = 0.9, respectively) was demonstrated. Total IgE was significantly higher in the ABPA group (P<0.01). The groups were comparable in skin prick test (SPT) positivity to a standard panel of aeroallergens (pollen, dander, molds, and mites) (P>0.2). Statistically significantly higher levels in the ABPA group were demonstrated in specific IgE to Af. (P < 0.05), SPT positivity to Af. (P < 0.02), and Af. precipitins (P < 0.05). Histamine release (HR) to Af. tended to be higher (P=0.075) in the ABPA group. Specific IgE to Af. was determined by Magic Lite (ML), CAP, and Maxisorp (in-house RAST). The CAP level was one to two classes higher than the ML level; however, the results were comparable (r=0.66, P<0.005). IgE to Af. measured by CAP was the test which offered the highest positive predictive value (PPV) and negative predictive value (NPV). Optimal diagnostic cutoff levels for the diagnosis of ABPA were determined: class 2 for HR to Af., 200 kIU/l for total IgE, and 3.5 (titer) for precipitating antibodies to Af., and class 2 for IgE to Af. (by CAP System). CONCLUSIONS: Unspecific atopy markers were of limited value for the diagnosis of ABPA. Patients with ABPA do not seem to be more atopic to other aeroallergens than non-ABPA patients. The most valid parameters for the diagnosis of ABPA in CF are SPT to Af., IgE to Af. in combination with precipitating antibodies to Af., and/or total IgE. PMID- 10696857 TI - Monoclonal antibody-based method to quantify Gly m 1. Its application to assess environmental exposure to soybean dust. AB - BACKGROUND: The demonstration that some asthma epidemics have been caused by allergens of soybean-hull dust prompted us to develop a two-site ELISA, suitable for the quantification of the major allergen (Gly m 1), to be used for the prevention of new episodes. METHODS: BALB/c mice were injected with Gly m 1 purified from soybean hulls. After fusion and screening, 10 monoclonal antibodies (mAbs) were obtained that were shown to be specific for Gly m 1 Two of them (6G1 as the capture antibody; 1G10 as the tracer) were selected to develop a quantitative two-site ELISA for the indoor and outdoor determination of Gly m 1. RESULTS: The two-site ELISA developed is very sensitive, with a detection limit of less than 0.2 ng/ml and a practical working range of 0.4-10 ng/ml. The assay is also highly reproducible with an intra-assay coefficient of variation of 3.5% and an interassay coefficient of variation of 12.5%. The method was applied to measure the concentration of Gly m 1 in air-sampler filters and in house-dust samples. Our results illustrate that there is a good correlation between the content of Gly m 1 in a number of samples and the allergenic activity as measured by ELISA inhibition. CONCLUSIONS: A specific and sensitive method is presented that can be used for the quantification of Gly m 1. The application of this method may allow the establishment of risk limits for soybean dust, and thus may contribute to the control of environmental contamination and to the prevention of new asthma epidemics. PMID- 10696858 TI - Occurrence of dog, cat, and mite allergens in public transport vehicles. AB - BACKGROUND AND METHODS: Helsinki City Transport buses, trams, and underground trains carry 687,000 passengers on a weekday. Of the passengers, 0.13% travel with a pet. We interviewed passengers and measured allergen levels in vehicles to determine what difficulties allergens cause to passengers with allergy and asthma. RESULTS: Of 2,021 interviewed passengers, 14% complained about inconvenience caused by pets, usually health problems. Of 324 adult passengers with allergy or asthma, 53% had experienced symptoms in public transport; the corresponding figure for 75 children was 32%. The median concentration of the main dog allergen, Can f 1, in dust from seats and floors in public transport vehicles was 2,400 ng per g of dust (range 20-8,500 ng/g). For the main cat allergen, Fel d 1, the median was 870 ng/g (range 3-2,600 ng/g). These levels can be regarded as low or moderate, and they cause symptoms in sensitive persons. Concentrations of mite allergens were undetectable or low. Allergen levels were lower in vehicles where pets were not allowed than in vehicles where pets were allowed, lower in dust from uncovered seats than in dust from seats with a covering, and lower after cleaning vehicle floors and seats than before cleaning. CONCLUSIONS: Dog and cat allergens are present in public transport vehicles in Helsinki at levels that cause symptoms in sensitive persons. Prohibiting pets would probably bring only a modest reduction in levels, as few pets are carried, and much allergen contamination comes from passengers' clothes. PMID- 10696859 TI - Long-term effects of specific immunotherapy, administered during childhood, in asthmatic patients allergic to either house-dust mite or to both house-dust mite and grass pollen. AB - In a retrospective study, asthmatic patients allergic to either house-dust mite (HDM) (Dermatophagoides pteronyssinus) (n = 34) or to both HDM and grass pollen (GP) (n = 14), and who were treated with specific immunotherapy (SIT) during childhood (mean duration of SIT: 61 +/- 9.70 months), were re-evaluated in early adulthood after mean cessation of SIT for 9.3 +/- 2.76 years. The results were compared to those of a control group of asthmatic patients (n = 42) with comparable asthma features, who were treated with appropriate antiasthmatic drugs during childhood, but who never received SIT. Re-evaluation was carried out with a standardized questionnaire, skin prick tests (SPT), and lung-function assessments. At the time of re-evaluation, the mean age in the SIT-treated group was 23.1 +/- 3.50 years; in the control group, it was 22.7 +/- 3.40 years. At re evaluation, the risk of frequent asthmatic symptoms was three times higher in the control group than in the SIT-treated group (prevalence ratio: 3.43; P = 0.0006). The frequent use of antiasthmatic medication was also more pronounced in the control group, although the difference was not statistically significant (P=0.38). Lung-function parameters and results of SPT with HDM were comparable in both groups. It is concluded that SIT has long-term effects on asthmatic symptoms in young adults. PMID- 10696860 TI - Nonatopic wheezy children have reduced interferon-gamma. AB - Viruses cause asthmatic exacerbations in schoolchildren. We tested the hypothesis that children who wheezed with viral respiratory tract infections secrete higher levels of the type 1 cytokine interferon-gamma (IFN-gamma) in the peripheral circulation than children who had never wheezed. Blood was taken from 13 children (eight atopic) with episodic wheeze and 11 controls. CD4 and CD8 cells were separated from peripheral blood mononuclear cells and stimulated with phorbol 12 myrisate 13-acetate (PMA) and ionomycin for 24 h. IFN-gamma, IL-4, and IL-5 were measured in the supernatant by ELISA. IFN-gamma production by CD4 and CD8 cells was lower in children with a history of wheeze (CD4, P = 0.046; CD8, P = 0.037). These children were then analysed according to atopic status. CD4 and CD8 IFN gamma production in nonatopic wheezy children was reduced (CD4, P=0.009; CD8, P=0.003). IFN-gamma production by atopic wheezy children was lower than by controls, but the differences were not significant (CD4, P = 0.2831; CD8, P = 0.1372). CD8 IL-5 was lower in children who wheezed (P=0.012). Release of IL-4 and IL-5 by CD4 cells did not differ between the three groups. We propose that defective IFN-gamma secretion by CD4 and CD8 cells may contribute to viral induced wheeze in nonatopic children. PMID- 10696861 TI - House-dust-mite allergen concentrations (Der f 1) and mold spores in apartment bedrooms before and after installation of insulated windows and central heating systems. AB - BACKGROUND: It has been hypothesized that changes in heating systems and insulation of homes in developed countries have generated an indoor climate favorable to organisms that excrete allergens inducing sensitization and allergic disease. The purpose of this study was to determine the influence of the installation of highly insulated windows and central heating systems on indoor climate, and mite-allergen (Der f 1) and mold spore concentrations. METHODS: The bedrooms of 98 apartments were examined before and 7 months (mean) after installation of insulated windows and central heating systems. The air-exchange rate, temperature, and humidity were measured. In settled dust on carpets and mattresses, the number of colony-forming mold spores and the Der f 1 concentration were determined. The inhabitants completed a questionnaire about their lifestyles and housing conditions. RESULTS: The air-exchange rate decreased from geometric mean 0.73 to 0.52 per hour (P=0.029). Temperature (mean 13.4 vs 17.5 degrees C, P<0.001), and absolute humidity (mean 4.6 g vs 6.2 g H2O/kg air, P<0.001) increased. Relative humidity remained nearly unchanged (mean 47.6 vs 49.1%). Der f 1 concentrations on carpets (geometric mean 0.65 vs 1.28 microg/g dust, P < 0.001) and mattresses (geometric mean 1.56 vs 2.40 microg/g, P=0.002) increased. Among the fungi that were analyzed, only the thermotolerant species Aspergillus fumigatus increased (geometric mean 20 vs. 60 colony-forming units/g carpet dust, P = 0.02). CONCLUSIONS: The findings of this study suggest that the installation of insulated windows and central heating systems is associated with an increase of Der f 1 concentrations in carpet and mattress dust and of A. fumigatus in carpet dust in apartment bedrooms. PMID- 10696862 TI - Exposure to formaldehyde and phenol during an anatomy dissecting course: sensitizing potency of formaldehyde in medical students. AB - BACKGROUND: The sensitizing potency of formaldehyde and phenol during anatomy dissecting was investigated. The objective was to determine whether exposure induces specific IgE or IgG against formaldehyde-albumin or phenol-albumin. METHODS: In 27 medical students, specific IgE against formaldehyde-albumin by RAST plus ELISA and specific IgE against phenol-albumin by ELISA were assessed. In addition, specific IgG against formaldehyde-albumin was assessed in 23 students. Symptoms before and during dissecting were assessed, and indoor formaldehyde and phenol were measured. RESULTS: Mean indoor formaldehyde was 0.265 +/- 0.07 mg/m3, and mean indoor phenol was 4.65 +/- 2.96 mg/m3. Specific IgE/IgG against formaldehyde-albumin was not found at the beginning. Four students developed specific IgE against formaldehyde-albumin (RAST classes of > or =2.0), and all four also had specific IgE in the ELISA, but IgG against formaldehyde-albumin was not found. Specific IgE against phenol-albumin was not seen. Itch and paresthesia of the hands (P<0.00001), dizziness (P<0.008), burning eyes (P<0.01), headache, sneezing, epistaxis, gingival bleeding, oral or pharyngeal itch, and shortness of breath were experienced. CONCLUSIONS: Formaldehyde exposure during dissecting may induce specific IgE, but not IgG, against formaldehyde-albumin. Sensitization did not correlate with symptoms. PMID- 10696863 TI - Successful treatment of occupational allergy to bumblebee venom after failure with honeybee venom extract. AB - BACKGROUND: Immediate-type allergies to bumblebee stings occur infrequently. Previous studies have demonstrated a high degree of cross-reactivity between honeybee venom (HBV) and bumblebee venom (BBV). It has been proposed that venom immunotherapy (VIT) with HBV is a therapeutic alternative for patients with BBV allergy. METHODS AND RESULTS: We present two cases of occupational immediate-type allergies to BBV. Although both nonatopic patients had a negative personal history of previous allergic reactions to honeybee sting, specific IgE antibodies and a positive intradermal reaction to HBV were detected. Despite VIT with HBV, the two developed another severe allergic reaction after incidental bumblebee stings. VIT with BBV, using in one patient a rush protocol and in the other a "conventional" regimen, with escalating doses of 0.01-100 microg of BBV, was performed. Before and during the VIT, the course of IgE and IgG specific antibodies to BBV was analyzed, demonstrating a significant decrease of BBV-IgE and an increase of BBV-IgG. The effectiveness of the treatment was also proven by an in-hospital sting challenge with a live bumblebee. CONCLUSIONS: Our data demonstrate that cross-immunotherapies with HBV do not protect BBV-allergic patients sufficiently. We conclude that BBV-allergic patients should be treated with BBV. A "rush" VIT with BBV is a safe alternative to a "conventional" protocol. PMID- 10696864 TI - IgE-mediated anaphylactic reaction to imipenem. PMID- 10696865 TI - Pyrethroid syndrome in an animal keeper. PMID- 10696866 TI - Anaphylactoid reaction to patch testing with ammonium persulfate. PMID- 10696867 TI - Inhalant allergy in the United Arab Emirates. PMID- 10696868 TI - Asthma to Gammarus shrimp. PMID- 10696869 TI - Immediate-type allergy caused by poppy seed. PMID- 10696870 TI - Chlorendic anhydride allergy. PMID- 10696871 TI - Role of cytosine deaminase and beta-alanine-pyruvate transaminase in pyrimidine base catabolism by Burkholderia cepacia. AB - A determination of the possible role of the salvage enzyme cytosine deaminase or beta-alanine-pyruvate transaminase in the catabolism of the pyrimidine bases uracil and thymine by the opportunistic pathogen Burkholderia cepacia ATCC 25416 was undertaken. It was of interest to learn whether these enzymes were influenced by cell growth on pyrimidine bases and their respective catabolic products to the same degree as the pyrimidine reductive catabolic enzymes were. It was found that cytosine deaminase activity was influenced very little by cell growth on the pyrimidines tested. Using glucose as the carbon source, only B. cepacia growth on 5-methylcytosine as a nitrogen source increased deaminase activity by about three fold relative to (NH4)2SO4-grown cells. In contrast, the activity of beta-alanine pyruvate transaminase was observed to be at least double in glucose-grown ATCC 25416 cells when pyrimidine bases and catabolic products served as nitrogen sources instead of (NH4)2SO4. Transaminase activity in the B. cepacia glucose grown cells was maximal after the strain was grown on either uracil or 5 methylcytosine as a nitrogen source compared to (NH4)2SO4-grown cells. A possible role for beta-alanine-pyruvate transaminase in pyrimidine base catabolism by B. cepacia would seem to be suggested from the similarity in how its enzyme activity responded to cell growth on pyrimidine bases and catabolic products when compared to the response of the three reductive catabolic enzymes. PMID- 10696872 TI - Molecular analyses of the IGS & ITS regions of rDNA of the psychrophilic yeasts in the genus Mrakia. AB - Species of the genus Mrakia are currently classified as synonyms based on molecular sequence analyses of the large sub-unit ribosomal DNA (LrDNA). Physiological and protein electrophoretic studies, however, reveal possible species differences. To clarify this discrepancy, we undertook molecular sequence analyses of the internal transcribed spacer (ITS) and intergenic spacer (IGS) regions of rDNA from the four psychrophilic Mrakia species and the psychrophilic yeast, Cryptococcus curiosus. Identical ITS sequences were found between C. curiosus, M. nivalis and M. frigida. Although, M. stokesii and M. gelida displayed identical ITS and IGS sequences, their sequences differed from the other three species by 2.3% and 38%, respectively. The results suggest that M. stokesii is a synonym of M. gelida, whereas M. nivalis is a synonym of M. frigida. Sequence differences (1.9%) observed in the IGS region indicates that C. curiosus is a distinct strain of M. frigida. PMID- 10696873 TI - Adaptive response to cold temperatures and characterization of cspA in Salmonella typhimurium LT2. AB - Salmonella typhimurium is a major foodborne microbial pathogen which primarily contaminates poultry products causing salmonellosis in humans. S. typhimurium LT2 cultures, when transferred from 37 degrees C to 5 degrees C or 10 degrees C, showed an initial lag period in growth with an approximate generation time of 10 25 h. Western blot assay using E. coli CS7.4 antibody and analysis of radiolabeled total cellular proteins from S. typhimurium cultures after exposure to 10 degrees C or 5 degrees C showed elevated expression of a major cold shock protein, CS7.4. Identification of a decreased level of CS7.4 at 37 degrees C suggests that the expression of this protein may require a large temperature downshift. Putative regulatory protein binding segment on the 5'-untranslated region referred as 'Fragment 7' in S. typhimurium exhibited a 90.6% and a 56.25% nucleotide sequence identity when compared with the Fragment 7 of E. coli and S. enteritidis, respectively. The differences in the nucleotide sequence within the Fragment 7 between S. typhimurium and S. enteritidis may explain the differential expression of CspA at 37 degrees C. The nucleotide sequence of the open reading frame of S. typhimurium cspA gene showed a single base difference at 816 bp position from a G to a C which altered the amino acid residue from a glycine to an alanine. In addition to CspA, an elevated expression of a 105 kDa, and decreased expression of 6 proteins were evidenced when cultures of S. typhimurium were exposed to 10 degrees C or 5 degrees C. Differential expression of the CspA and other proteins in S. typhimurium following exposure to cold temperatures suggest that adaptation and continued growth and survival at cold temperatures in this pathogen may be aided by these cold-responsive proteins. PMID- 10696874 TI - A role for Lewis a antigens on salivary agglutinin in binding to Streptococcus mutans. AB - Streptococcus mutans is a major etiological agent in dental caries. Salivary agglutinin is one of the main salivary components binding to S. mutans. To learn more about the interaction of salivary agglutinin with S. mutans, parotid, submandibular, sublingual and palatal saliva samples were incubated with S. mutans suspension. Both depleted saliva samples and bacterial extracts were analyzed by SDS-PAGE and immunoblotting. Salivary agglutinin was present in all types of glandular saliva and in all cases bound to S. mutans, also to PC337C, a P1 mutant of S. mutans. Agglutinin was separated by SDS-PAGE under reducing and non-reducing conditions and then transferred to nitrocellulose. Non-reduced agglutinin bound S. mutans, but reduced agglutinin did not. Adhesion of S. mutans to agglutinin-coated microplates was inhibited by amine-containing components, 1 M NaCl or KCl and EDTA. Adhesion decreased with decreasing pH with no adhesion below pH 5.0. These data suggest that calcium-dependent electrostatic interactions play a role in binding. By immunoblotting was demonstrated that blood group antigens and Lewis antigens were present on agglutinin. Synthetic blood group antigens and Lewis antigens covalently coupled to polyacrylamide were tested for binding to S. mutans. Only Le(a)(Gal beta 1,3(Fuc alpha 1,4)GlcNAc) bound to S. mutans, whereas the blood group antigens Le(b), Le(x), Le(y), H1, H2, A, B and sialylated Le(a) did not. Lea without galactose (Fuc alpha 1,4GlcNAc) still bound to S. mutans, but Le(a) without fucose (Gal beta 1,3GlcNAc) did not. Binding of agglutinin to S. mutans was not inhibited by Le(a). In conclusion, S. mutans can bind to Le(a) carbohydrate epitopes in which the fucose is an essential residue. Le(a) carbohydrate epitopes are present on salivary agglutinin but play no major role in binding. PMID- 10696875 TI - Production of polyesters consisting of medium chain length 3-hydroxyalkanoic acids by Pseudomonas mendocina 0806 from various carbon sources. AB - Pseudomonas mendocina strain 0806 was isolated from oil-contaminated soil and found to produce polyesters consisting of medium chain length 3-hydroxyalkanoates (mclPHAs). The monomers of mclPHAs contained even numbers of carbon atoms, such as 3-hydroxyhexanoate (HHx or C6), 3-hydroxyoctanoate (HO or C8), and/or 3 hydroxydecanoate (HD or C10) as major components when grown on many carbon sources unrelated to their monomeric structures, such as glucose, citric acid, and carbon sources related to their monomeric structures, such as myristic acid, octanoate, or oleic acid. On the other hand, PHA containing both even and odd numbers of hydroxyalkanoates (HA) monomers was synthesized when the strain was grown on tridecanoic acid. The molar ratio of carbon to nitrogen (C/N) had a significant effect on PHA composition: the strain produced PHAs containing 97-99% of HD monomer when grown in a glucose ammonium sulfate medium of C/N<20, and 20% HO, and 80% of the HD monomer when growth was conducted in media containing C/N>40. It was demonstrated that the HO/HD ratio in the polymers remained constant in media with a constant C/N ratio, regardless of the glucose concentration. Up to 3.6 g/L cell dry weight containing 45% of PHAs was produced when the strain was grown for 48 h in a medium containing 20 g/L glucose with a C/N ratio of 40. PMID- 10696876 TI - Pichia sporocuriosa sp. nov., a new yeast isolated from rambutan. AB - A strain of a hitherto undescribed yeast species with a unique ascospore morphology was isolated from rambutan (Nephelium lappaceum). A description of the new species, Pichia sporocuriosa, is given. PMID- 10696877 TI - The isolation and classification of Candida mokoenaii sp. nov. A new yeast isolate from South African soil. AB - A new xylanase producing yeast species was isolated from South African soil. A description of the new species, Candida mokoenaii is given. PMID- 10696878 TI - Significance of gaseous NO for ammonia oxidation by Nitrosomonas eutropha. AB - Nitrification by the obligately lithoautotrophic ammonia oxidizer Nitrosomonas eutropha was significantly inhibited when nitric oxide was removed from the culture medium by means of intensive aeration and turbulence. Nearly complete recovery of ammonia oxidation could be achieved by adding 100 ppm NO to the supplied air. Inhibition of ammonia oxidation occurred also upon addition of the NO binding agent 2,3-Dimercapto-1-propane-sulfonic acid (DMPS). Recovery of ammonia oxidation occurred within 3 h in the presence of 100 ppm NO and within 76 h in the absence of externally added NO. In co-cultures of N. eutropha and the NO detoxifying bacterium Pseudomonas PS88, hardly any nitrification was detectable and release of NO was extremely low when the heterotroph was provided with an organic substrate. When cells of Pseudomonas PS88 were added to a mixotrophically nitrifying culture of N. eutropha the release of NO decreased drastically upon the addition and ammonia oxidation ceased. These results confirm for the first time the significance of NO in the course of ammonia oxidation by N. eutropha. PMID- 10696879 TI - Strain variability and the effects of organic compounds on the growth of the chemolithotrophic bacterium Thiobacillus ferrooxidans. AB - The effects of naturally-occurring organic compounds on ferrous iron oxidation by the bacterium Thiobacillus ferrooxidans were examined with a view to using these compounds to treat or prevent acid mine/rock drainage. The compounds glucose, cellobiose, galacturonic acid, and citric acid were added to the growth medium of five different strains of the bacterium and growth studies were done to determine whether or not strain differences existed with respect to organic compound sensitivity. The effects of these compounds were compared to the effects of sodium dodecyl sulfate (SDS) an anionic detergent. Each of the compounds tested had an inhibitory effect on the strains of the bacterium and sensitivity to these compounds was strain dependent. All strains appeared to be equally susceptible to SDS. Inhibitory concentrations ranged from 70 mM to >280 mM for glucose, 7.5 mM to 150 mM for cellobiose, 20 mM to 230 mM for galacturonic acid, and 50 mM to 130 mM for citric acid. SDS effectively inhibited iron oxidation for all strains at a concentration of 0.3 mM, the lowest concentration tested. Some naturally occurring organic compounds, therefore, might be candidates for the growth control of T. ferrooxidans. PMID- 10696880 TI - Molecular cloning of Saccharomyces cerevisiae MGC1/YDR473c gene which is essential for cell growth. AB - Saccharomyces cerevisiae haploid cells undergo morphological changes in response to mating pheromones, a- and alpha-factors, during sexual conjugation. As a first step to elucidate the mechanism, I had previously identified the mgc1 mutation which affected the morphogenesis induced by mating pheromones. The mutation had been designated mgc1 for morphogenesis control. In the present study I cloned the MGC1 gene. Sequencing analysis indicates that the MGC1 gene corresponds to the YDR473c gene. The MGC1 gene was shown to be essential for cell growth and required for the transition from the G1 to S phase of cell cycle. Protein-protein interaction of Mgc1 protein was shown by using yeast two-hybrid system. Mgc1 protein was also proposed to be localized in the nucleus in yeast cells. PMID- 10696881 TI - Re-examination of some species of the genus Geotrichum Link: Fr. AB - The nutritional physiology and the growth rate of thirty-four strains representing species of Geotrichum without known teleomorph states were examined. From twenty-seven strains the mol% G+C were calculated from the DNA melting curves. The first derivatives of the melting curves of seven strains, including the type strain of Geotrichum clavatum, demonstrated the presence of two peaks, 12% away from each other; the remaining strains showed only a single broad peak. DNA homology values among strains of the former group were high, indicating their conspecificity. The strains of the latter group could be subdivided into six DNA homology groups, four of which could be identified with recognized species and two may represent novel taxa. A combined key of Geotrichum and its teleomorph states Galactomyces and Dipodascus is presented. PMID- 10696882 TI - Analysis of genetic variability within the genus Petromyces. AB - Phenotypic and genotypic features of three teleomorphic species, Petromyces alliaceus, P. albertensis and P. muricartus and some related anamorphic Aspergillus species were compared. The dendrogram based on carbon source utilisation data revealed a close relationship between P. muricarus and the A. ochraceus strains examined. P. alliaceus and P. albertensis strains were very closely related to each other. A dendrogram with similar topology was obtained by analysing sequences of the intergenic transcribed spacer regions of representatives of these species. P. alliaceus and P. albertensis strains could only be distinguished by the random amplified polymorphic DNA technique. These strains possibly represent a single species closely related to Aspergillus section Flavi, while the anamorph of P. turicatus is a member of Aspergillus section Circumdati. Our results indicate that Aspergillus section Circumdati is in need of taxonomic revision. PMID- 10696883 TI - The specific transport system for lysine is fully inhibited by ammonium in Penicillium chrysogenum: an ammonium-insensitive system allows uptake in carbon starved cells. AB - The regulation exerted by ammonium and other nitrogen sources on amino acid utilization was studied in swollen spores of Penicillium chrysogenum. Ammonium prevented the L-lysine, L-arginine and L-ornithine utilization by P. chrysogenum swollen spores seeded in complete media, but not in carbon-deficient media. Transport of L-[14C]lysine into spores incubated in presence of carbon and nitrogen sources was fully inhibited by ammonium ions (35 mM). However, in carbon derepressed conditions (growth in absence of sugars, with amino acids as the sole carbon source) L-[14C]lysine transport was only partially inhibited. Competition experiments showed that L-lysine (1 mM) inhibits the utilization of L-arginine, and vice versa, L-arginine inhibits the L-lysine uptake. High concentrations of L ornithine (100 mM) prevented the L-lysine and L-arginine utilization in P. chrysogenum swollen spores. In summary, ammonium seems to prevent the utilization of basic amino acids in P. chrysogenum spores by inhibiting the transport of these amino acids through their specific transport system(s), but not through the general amino acid transport system that is operative under carbon-derepression conditions. PMID- 10696884 TI - Standards for anal sphincter replacement. AB - PURPOSE: Anal sphincter replacement offers a new treatment option for patients with severe refractory fecal incontinence or for those who require abdominoperineal resection for localized malignancy. The purpose of this study was to review the current status of anal sphincter replacement, formulate a consensus statement regarding its current use, and outline suggestions for future development. METHODS: Four areas of interests were selected: indications for sphincter replacement, continence scoring and quality of life, choice of therapy, and dissemination of new technology. A questionnaire regarding these issues was developed and circulated to working party members; its results served as the basis for this consensus document. RESULTS: Both electrically stimulated skeletal muscle neosphincter and artificial anal sphincter are options for patients with end-stage fecal incontinence. Electrically stimulated skeletal muscle neosphincter is also appropriate for reconstruction after surgical excision of the anorectum in selected cases. Avoidance of complications requires strict attention to sterile technique, prophylactic antibiotics, and deep venous thrombus prophylaxis. A standardized scoring system is proposed that evaluates both continence and evacuation. Quality of life is a critical endpoint for assessing sphincter replacement, and use of The American Society of Colon and Rectal Surgeons incontinence-specific quality-of-life instrument is recommended. As the efficacy of sphincter replacement becomes proven, dissemination of the technique should occur in a controlled manner to ensure adequate surgeon training, minimization of complications, and optimization of results. CONCLUSIONS: Sphincter replacement by electrically stimulated skeletal muscle neosphincter and artificial anal sphincter provide a continent option for patients with end-stage fecal incontinence and those requiring abdominoperineal resection. The guidelines offered in this document are intended to facilitate the controlled and safe development and acceptance of these new techniques. PMID- 10696885 TI - Ischiorectal fossa block decreases posthemorrhoidectomy pain: randomized, prospective, double-blind clinical trial. AB - PURPOSE: Hemorrhoidectomy can be associated with severe pain in the immediate postoperative period. The aim of this study was to assess the efficacy of a preemptive local anesthetic, ischiorectal fossa block, in the reduction of pain and analgesic requirements after hemorrhoidectomy. METHODS: All patients were suitable for an established day surgery hemorrhoidectomy protocol. Immediately before surgery patients were randomly assigned either to receive (Group 1) or not receive (Group 2) the local anesthetic block. All other aspects of surgery and anesthesia were standardized. Nursing staff assessed pain at 30 minutes and 2, 4, and 24 hours postoperatively using a visual analog scale (1-10, where 1 represented no pain and 10 represented the worst pain imaginable). Analgesic requirements also were recorded at these times. Both the patients and the nursing staff were blinded to which local anesthetic protocol had been used. RESULTS: Twenty patients were enrolled in the trial. Ten patients were randomly assigned to Group I and ten to Group 2. Mean pain scores for Group 1 (anal block) at 0.5, 2, 4, and 24 hours were 1.5, 1.8, 2.1, and 2.5, respectively, compared with Group 2, with mean pain scores of 3.4, 3.4, 3.9, and 5.1. These differences were statistically significant. Patients in Group 1 used less analgesia in the first 24 hours postoperatively than those in Group 2. CONCLUSION: The use of a preemptive local anesthetic, ischiorectal fossa block, is associated with a significant decrease in pain and analgesia requirements after hemorrhoidectomy. PMID- 10696886 TI - Validation of a questionnaire to assess fecal incontinence and associated risk factors: Fecal Incontinence Questionnaire. AB - PURPOSE: Although fecal incontinence is a topic of considerable importance, there are no validated self-report measures of fecal incontinence available. The aim of this study was to develop a questionnaire to measure fecal incontinence and its risk factors in the community. METHOD: The reliability and concurrent validity of the fecal incontinence questionnaire were measured by test-retest procedures in a population of clinic patients. The questionnaire was created for a sixth-grade reading level, with large print. Ninety-four adult patients were surveyed. Thirty four patients repeated the questionnaire through the mail. Forty-one patients were independently retested over the telephone by a physician to assess concurrent validity. Nine patients refused retest, and ten patients did not respond to a second contact. RESULTS: The fecal incontinence questionnaire was well understood and well accepted. Reliability (overall median kappa, 0.68; interquartile range, 0.03-1) and validity (overall median kappa, 0.59; interquartile range, 0.27-1) were acceptable for the mailed retest and the telephone retest, respectively. The presence of fecal incontinence as measured by questionnaire was greatly increased when compared with physician history in clinical records; only 3 percent of patients reported no fecal incontinence on the questionnaire when the clinic chart had documented this problem. CONCLUSION: Our initial results indicated that this new self-report questionnaire is a useful tool for assessing the presence of fecal incontinence in the population and has greater sensitivity compared with a standard physician interview. Specific attention should be given to identifying fecal incontinence and associated symptoms during history taking. PMID- 10696887 TI - Molecular staging of colorectal cancer: K-ras mutation analysis of lymph nodes upstages Dukes B patients. AB - PURPOSE: Multiple attempts have been made to improve the clinical/pathologic staging system of Dukes to focus adjuvant therapy decisions. The purpose of this study was to determine whether K-ras mutational status of regional nodes in patients with Dukes B2 colorectal cancer could be used to stage their disease more accurately. METHODS: Using formalin-fixed, paraffin-embedded archival material, tumor samples were screened for K-ras mutations using a mutation specific polymerase chain reaction method, followed by gel electrophoresis in a 96-well array. Patients with Dukes B2 tumors that have mutations in codon 12 or 13 of the K-ras gene were identified. RESULTS: Mutational analysis of the lymph nodes from these patients revealed an 80 percent (16/20) incidence of the same mutations in regional lymph nodes. None of the four patients with mutation-free nodes developed recurrence compared with 37.5 percent (6/16) with K-ras positive lymph nodes. CONCLUSIONS: The data suggest that patients with Dukes B2 colorectal cancers that have mutations in codon 12 or 13 of the K-ras gene are at high risk for the development of nodal metastases. Mutational analysis of the lymph nodes identifies high-risk patients who should be considered for adjuvant chemotherapy. Therefore, K-ras mutational analysis should be considered for molecular staging of colorectal cancer. PMID- 10696888 TI - Effect of Morphine and incision length on bowel function after colectomy. AB - PURPOSE: Return of bowel function remains the rate-limiting factor in shortening postoperative hospitalization of patients with colectomies. Narcotics are most commonly used in the management of postoperative pain, even though they are known to affect gut motility. Narcotic use has been felt to be proportional to the length of the abdominal incision. The aim of this study was to determine whether return of bowel function after colectomy is directly related to narcotic use and to evaluate the effect of incision length on postoperative ileus. METHODS: A prospective evaluation of 40 patients who underwent uncomplicated, predominantly left colon and rectal resections was performed. Morphine administered by patient controlled analgesia was the sole postoperative analgesic. The amount of morphine used before the first audible bowel sounds, first passage of flatus and bowel movement, and incision length were recorded. Spearman correlation coefficients were calculated between all variables. RESULTS: The strongest correlation was between time to return of bowel sounds and amount of morphine administered (r = 0.74; P = 0.001). There were also significant correlations between morphine use and time to report of first flatus (r = 0.47; P = 0.003) and time to bowel movement (r = 0.48; P = 0.002). There was no relationship between incision length and morphine use or incision length and return of bowel function in the total group. CONCLUSIONS: Return of bowel sounds, reflecting small-intestine motility after colectomy, correlated strongly with the amount of morphine used. Similarly, total morphine use adversely affects colonic motility. Because no relationship with incision length was found, efforts to optimize the care of patients with colectomies should be directed less toward minimizing abdominal incisions and more toward diminishing use of postoperative narcotics. PMID- 10696889 TI - Anal sphincter injuries from stapling instruments introduced transanally: randomized, controlled study with endoanal ultrasound and anorectal manometry. AB - PURPOSE: Injury sustained from the transanally introduced stapling technique was assessed by comparison with biofragmentable anastomotic ring anastomosis, which excluded anal manipulation. METHODS: A randomized, controlled trial was conducted on consecutive patients undergoing sigmoid colectomy (where pelvic nerve injury was avoided). A bowel function questionnaire was administered six months after surgery. Anorectal manometry and endoanal ultrasonography were performed preoperatively and at six months postoperatively. The observers were blinded to the randomization. RESULTS: There were 18 patients in the transanally introduced stapling technique group and 17 patients in the biofragmentable anastomotic ring group, with no differences in age, gender, Dukes staging, and follow-up. Three of the transanally introduced stapling technique patients had occasional liquid soiling, which was absent in biofragmentable anastomotic ring patients. Mean change in resting anal pressures was also significantly impaired when compared with patients with biofragmentable anastomotic ring (P = 0.007). Endosonographic internal sphincter fragmentation was found in five transanally introduced stapling technique patients but none after biofragmentable anastomotic ring anastomosis (P = 0.046). Internal sphincter fragmentation was associated with the impaired resting pressures (P = 0.007). External sphincter deficiencies were found after transanally introduced stapling technique in two patients (biofragmentable anastomotic ring = 0), and these were associated with the soiling (P = 0.005). CONCLUSIONS: The transanally introduced stapling technique may result in anal sphincter defects and impaired anal pressures when assessed at six months of follow-up. PMID- 10696890 TI - Glyceryl trinitrate for chronic anal fissure--healing or headache? Results of a multicenter, randomized, placebo-controled, double-blind trial. AB - PURPOSE: Internal anal sphincterotomy for treating chronic anal fissure can irreversibly damage anal continence. Reversible chemical sphincterotomy may be achieved by anal application of glyceryl trinitrate ointment (nitric oxide donor), which has been reported to heal the majority of patients with anal fissure by inducing sphincter relaxation and improving anodermal blood flow. This trial aimed to further clarify the role of glyceryl trinitrate in the treatment of chronic anal fissure. METHODS: A total of 132 consecutive patients from nine centers were randomly assigned to receive 0.2 percent glyceryl trinitrate ointment or placebo twice daily for at least four weeks. The severity of pain and maximum anal resting pressure were measured before and after one week of treatment. Anodermal blood flow was measured before and after application of glyceryl trinitrate or placebo in ten patients. RESULTS: The study was completed by 119 patients (59 glyceryl trinitrate and 60 placebo), matched for gender, age, duration of symptoms, duration of treatment, site of fissure, previous attempts to treat, pain score, and maximum anal resting pressure. Twenty-nine patients (49.2 percent) healed after glyceryl trinitrate and 31 patients (51.7 percent) healed after placebo (P = not significant). Pain score fell significantly in both groups, in addition to maximum anal resting pressure. Anodermal blood flow improved significantly in seven patients receiving glyceryl trinitrate, but not in the three receiving placebo. Twenty-three patients (33.8 percent) experienced headache and 4 (5.9 percent), orthostatic hypotension after glyceryl trinitrate. CONCLUSION: This trial fails to demonstrate any superiority of topical 0.2 percent glyceryl trinitrate treatment vs. a placebo, although the effects of glyceryl trinitrate on anodermal blood flow and sphincter pressure are confirmed. This finding, together with the high incidence of side-effects, should discourage the use of this treatment as a substitute for surgery in chronic anal fissure. PMID- 10696891 TI - Use of bioresorbable membrane (sodium hyaluronate + carboxymethylcellulose) after controlled bowel injuries in a rabbit model. AB - PURPOSE: Patients in whom enterolysis is performed are at high risk for recurrence of adhesions and for injury during adhesiolysis. Therefore, the aim of this study was to assess the safety of sodium hyaluronate-based bioresorbable membrane (Seprafilm) after myotomy and enterotomy. METHODS: A total of 60 rabbits underwent laparotomy with equal distribution to one of three groups: creation of either three repaired, or three unrepaired myotomies, or three repaired enterotomies. Thus, a total of 180 defects were created in the same anatomic positions. One-half of the animals in each group had the surface of the myotomies or enterotomies covered by Seprafilm. Fourteen days later, after complete absorption of Seprafilm, the presence of intra-abdominal abscess, adhesions, and the integrity of the suture line were evaluated by a surgeon blinded to the use of Seprafilm and by a standard radiographic isobaric contrast study. Statistical analysis was done by use of Fisher's exact test; significance was set at P < 0.05. RESULTS: The incidence of adhesions in the repaired myotomy group were 2 (6.6 percent) and 9 (30 percent) in the Seprafilm and control (nonSeprafilm) groups, respectively (P < 0.05); in the unrepaired myotomy group, 2 (6.6 percent) and 10 (33 percent) in the Seprafilm and control groups, respectively (P < 0.05); and in the enterotomy group, 28 (94 percent) and 29 (97 percent) in the Seprafilm and control groups, respectively (P = not significant). A single phlegmon occurred in the myotomy group at a Seprafilm site (1.6 (1/60) vs. 0 percent, P = not significant). There were no leaks in this group. In the enterotomy group, the incidence of phlegmons was 33 percent (10/30) in the Seprafilm group, whereas it was 27 percent (8/30) in the nonSeprafilm group (P = not significant). The incidence of leaks was 6.6 (2/30) and 10 percent (3/30) in the Seprafilm and nonSeprafilm group, respectively (P = not significant). CONCLUSION: The use of Seprafilm at the sites of myotomies significantly reduced the incidence of adhesions. Effectiveness at the enterotomy site may have been attenuated by a greater inflammatory response. Importantly, Seprafilm did not increase septic mortality in any group. PMID- 10696892 TI - Evaluation and outcome of the delorme procedure in the treatment of rectal outlet obstruction. AB - PURPOSE: This study was designed to assess the results of the Delorme procedure in the treatment of patients with rectal outlet obstruction. METHODS: A descriptive retrospective study from October 1989 to October 1997 was undertaken. Thirty-four patients with an abnormal defecography documenting rectal outlet obstruction caused by internal rectal prolapse or a combination of internal rectal prolapse and rectocele were included in the study. RESULTS: Thirty-four patients (33 females) ages 35 to 82 (mean, 61.4) years were followed up for the duration of the study (mean follow-up, 43 months). Twenty-six patients (76.4 percent) reported a good to excellent overall result after the Delorme procedure. Eight patients (23.6 percent) reported fair to poor results. Symptomatic improvement was observed in 89.7 percent for patients who had incomplete evacuation, and in 88.5 percent of patients who had constipation. There was improvement in 78.6 percent of patients with bleeding per rectum, in 92.9 percent of patients with straining, and in 82.4 percent of patients with the need to manually assist in defecation by pushing in the perineum or vagina. Discontinuation of laxative use after the procedure was reported by 66.7 percent of patients. Improvement in the patients with some degree of incontinence was seen in 33.3 percent. Twelve patients (35.3 percent) experienced one or more complications. The procedure was performed in an outpatient setting in 71 percent of the patients. CONCLUSIONS: The Delorme procedure for the treatment of rectal outlet obstruction can be done with minimal morbidity, short hospital stay often in an outpatient setting, with good functional results, and with an overall patient satisfaction above 75 percent. PMID- 10696893 TI - Learning curve of transrectal ultrasound. AB - PURPOSE: Transrectal ultrasound is the most accurate means of assessing the degree of invasion for rectal neoplasms. A learning curve for performing and interpreting these studies exists, but it is unknown how long or steep it is. We reviewed our initial results with transrectal ultrasound to determine our accuracy and to define the learning curve. METHODS: All patients undergoing transrectal ultrasound during our initial 30 months of experience were included. Each patient was staged with transrectal ultrasound and, after resection, the histopathologic stage was compared with transrectal ultrasound staging. The accuracy of transrectal ultrasound was calculated at intervals as experience was gained. RESULTS: A total of 42 examinations were performed on 41 neoplasms in 41 patients. Comparison between transrectal ultrasound and the pathologic stage could be made in 36 studies. Overall accuracy of degree of wall invasion was 78 percent. Overstaging occurred with eight neoplasms, and one lesion was understaged. Accuracy of transrectal ultrasound staging improved with time: 58 percent of the initial 12 studies were staged correctly compared with 87.5 percent accuracy in the remaining 24 examinations (P = 0.048). CONCLUSION: A definite learning curve was apparent. We conclude that transrectal ultrasound is a relatively simple procedure to learn and, once a moderate degree of experience is gained, should be routinely incorporated into the evaluation of rectal neoplasms. PMID- 10696894 TI - Preliminary experience in management of fecal incontinence caused by internal anal sphincter injury. AB - PURPOSE: Isolated injuries of the internal anal sphincter can cause fecal incontinence. With the advent of ultrasound, which accurately delineates the anatomy of the anal sphincters, internal sphincter injuries can be diagnosed more precisely. The purpose of this study was to evaluate the outcome of direct repair of isolated internal anal sphincter defects. METHODS: Eight patients (6 males; median age, 37 years) with clinically and sonographically proved internal anal sphincter defects were the subject of this study. Patients had different degrees of incontinence that failed to respond to medical treatment. All patients had their sphincters repaired by direct apposition using coated Vicryl 2-0 stitches. A strict postoperative regime that avoided stretch of the sphincter for one month was adopted. RESULTS: At a median follow-up period of 15 months, continence improved in all patients, and two achieved full continence. None of the patients wore pads. Mean continence score improved significantly from 4 to 12 and 11 at 6 and 12 postoperative months, respectively (P < 0.0001, paired t-test). CONCLUSION: Despite the limited number of patients and the short follow-up, the preliminary results of repair of isolated internal sphincter defects are satisfactory. PMID- 10696895 TI - Diagnosing enteroceles using dynamic magnetic resonance imaging. AB - PURPOSE: Enteroceles are in part difficult to detect but a frequent finding in pelvic floor disorders. The aim of this study was to evaluate magnetic resonance colpocystorectography in the diagnosis of enteroceles. METHODS: In this prospective study 11 volunteers and 55 patients with pelvic floor descent were examined. In addition to magnetic resonance colpocystorectography, a dynamic cystoproctography was performed on 34 patients. Opacification of organs was used. An enterocele was assessed in relationship to the pubococcygeal reference line (magnetic resonance colpocystorectography) or the width of the rectovaginal space (dynamic cystoproctography). A clinical gynecologic examination served as reference. RESULTS: The clinical examination diagnosed an enterocele in 43, magnetic resonance colpocystorectography in 49, and dynamic cystoproctography in 14 cases. Magnetic resonance colpocystorectography further subdivided the enteroceles according to their contents (mesenteric fat or fluid, 12; small bowel, 32, large bowel, 3; and rectosigmoidocele, 2). Magnetic resonance colpocystorectography proved statistically significantly superior to dynamic cystoproctography (15 cases) and the reference. Sensitivity and specificity of magnetic resonance colpocystorectography were 100 percent each. It was able to reveal clinically missed enteroceles as being peritoneoceles associated with a rectocele or a uterovaginal prolapse (10 cases). CONCLUSION: Magnetic resonance colpocystorectography is a promising method for diagnosis of enteroceles, because hernial canal, sac, and contents are reliably identified. PMID- 10696896 TI - Assessment of ileal pouch inflammation by single-stool calprotectin assay. AB - PURPOSE: Assessment of inflammation within the ileal pouch to establish a diagnosis of "pouchitis" requires both pouch endoscopy and biopsy because there can be a poor correlation between macroscopic and histologic assessments of inflammation. A simplified diagnostic test would be of clinical advantage. Calprotectin is a stable myelomonocytic protein, measurable in feces. It quantitatively relates to inflammation within the gastrointestinal tract. This study was designed to compare single and 24-hour stool measurements of calprotectin in patients with and without evidence of ileal pouch inflammation with endoscopic, histologic, and immunohistochemical indices. METHODS: Twenty four-hour stool collections were made in ileal pouch patients, 9 with and 15 without (7 with ulcerative colitis and 8 with familial polyposis coli) evidence of pouch inflammation. First-morning stool concentration and total 24-hour calprotectin were quantified by use of a single step enzyme-linked immunosorbent assay. Biopsies from the reservoir were taken for conventional histology and scoring of intraepithelial neutrophil infiltrate. Cells positive for CD3, CD45RO, CD14, and CD15 within the lamina propria were quantified by use of immunohistochemistry. RESULTS: The mean first-morning stool calprotectin concentration correlated with the 24-hour level (r = 0.91; P = <0.0001). The median single-stool calprotectin concentrations were 39 mg/l, 4 mg/l, and 8.5 mg/l (normal range, 0.2-10 mg/l) in patients with inflamed, noninflamed ulcerative colitis, and familial adenomatous polyposis, respectively. All nine patients with endoscopic and histologic evidence of pouch inflammation had raised stool calprotectin. Two of 15 patients without evidence of pouch inflammation had abnormal stool calprotectin. Single-stool calprotectin concentration correlated with the percentage of mature granulocytes (CD15; r = 0.46; P = 0.04) and activated macrophages (CD14; r = 0.65; P = 0.006), but not memory T cells (CD45RO; r = -0.05; P = 0.4) within the lamina propria. CONCLUSION: Single first morning stool calprotectin levels provide a quantitative measure of pouch inflammation, which may be helpful in the diagnosis and assessment of pouchitis. PMID- 10696897 TI - Concurrent expressions of metallothionein, glutathione S-transferase-pi, and P glycoprotein in colorectal cancers. AB - PURPOSE: Because the status of the inherent drug-resistance of colorectal cancers remains obscure, human colorectal cancers with no neoadjuvant therapy were retrospectively investigated regarding the expression of three drug-resistant proteins: metallothionein, glutathione S-transferase-pi, and P-glycoprotein. METHODS: Paraffin-embedded tissues of 130 colorectal cancers (Dukes A, 20; B, 49; C, 41; D, 20) obtained by surgical resections from 1982 to 1989 were used. The three proteins were immunostained by the streptavidin-biotin complex method. The immunostaining was judged to be positive if more than 5 percent of cells showed positive staining by use of cell analysis system. The data were compared with clinicopathologic features (Dukes A-D) and patients' prognosis (Dukes AC). RESULTS: Metallothionein, glutathione S-transferase-pi, and P-glycoprotein were positively expressed in 91 (70 percent), 30 (23 percent), and 98 (75 percent), respectively. A total of 120 (86 percent) expressed at least one drug-resistant protein. No intergroup differences were observed between positive and negative expressions of the proteins and their clinicopathologic features except tumor location. Rectal cancers positively expressed P-glycoprotein and three proteins more frequently. Twenty-six (20 percent), 65 (50 percent), and 21 (16 percent) cancers positively expressed one, two, and three proteins, respectively. The disease-free survival rates of patients with Dukes A through C cancer with positive staining for one, two, and three proteins were 100, 94, and 83 percent (at 1 year); 100, 72, and 51 percent (at 3 years); and 94, 66, and 38 percent (at 5 years), respectively (Kaplan-Meier with log-rank test; P = 0.016). In the multivariate Cox analysis, age, Dukes stage, tumor size, and glutathione S transferase-pi were independent prognostic factors. CONCLUSIONS: The patients with concurrent expression of drug-resistant proteins in their cancers had worse prognoses. Examining drug-resistant proteins in colorectal cancers may be useful in selecting adjuvant chemotherapy and in predicting prognosis more accurately. PMID- 10696898 TI - How should patients 80 years of age or older with colorectal carcinoma be treated? Long-term and short-term outcome and postoperative cytokine levels. AB - PURPOSE: The aim of this study was to determine how extensive resection affects operative morbidity, mortality, and long-term survival in elderly patients with colorectal cancer. METHODS: A total of 119 patients 80 years of age or older were given a diagnosis of colorectal carcinoma at our hospital between 1985 and 1997. Eleven patients who did not undergo surgery were excluded. The remaining 108 patients underwent laparotomy and were reviewed. Serum levels of interleukin-6 were measured perioperatively in 22 patients to assess the degree of operative stress. RESULTS: Potentially curative resection was performed in 64 (88.9 percent) of the 72 patients in the active performance status group and 13 (36.1 percent) of the 36 patients in the sedentary performance status group (P < 0.001). The in-hospital mortality rate was 8.3 percent in group the active performance status group and 38 percent in the sedentary performance status group (P = 0.007). Patients in the sedentary performance status group and those who underwent emergency operations had higher levels of IL-6 than patients in the active performance status group or those who underwent elective operations. CONCLUSIONS: Preoperative performance status, operative curability, and tumor stage have a significant impact on outcome in patients with colorectal cancer who are 80 years of age or older. Knowledge of early postoperative response of IL-6 is useful in predicting postoperative mortality and morbidity in this subgroup of patients. PMID- 10696899 TI - Computer-based inpatient medical record in colorectal surgery: pilot study. AB - PURPOSE: Clinical guidelines and care maps are important tools for improving quality of care and reducing costs. However, problems of quantity, quality, and accessibility of data recorded in the inpatient medical record have not been solved by the implementation of clinical pathways. Variance or "charting by exception" improves legibility, in part. The aim of the present study was to design a computer-based medical record on a database platform to provide legible notes within a clinical guideline and variance charting framework. METHODS: A computerized database program was written, integrating pre-established clinical guidelines into a user-friendly interface according to modification of the charting by exception principles. Patient care guidelines were provided for each postoperative day. After an initial debugging process by entering data from old charts of patients, the software was installed and its function was evaluated on selected patients. The charting time was compared with the standard charting method. Functionality and user friendliness were assessed. RESULTS: After a brief introduction of ten minutes, all users were able to use the software without difficulties. It was found to be functional and user friendly. The charting time was shorter for the computer-based inpatient medical record compared with the charting time of the standard charts. Because all daily notes were printed on standardized forms on a laser printer, legibility was excellent. CONCLUSIONS: The results of this pilot study suggest that the idea of computer-based inpatient medical record integrating an on-line inpatient medical record in a database platform is feasible. Further development and integration with other hospital information systems and the other health-care providers is required. PMID- 10696900 TI - Audit of single-stage proctocolectomy for Crohn's disease: postoperative complications and recurrence. AB - PURPOSE: This study was undertaken to review our overall experience of single stage proctocolectomy for Crohn's disease. METHODS: One hundred three patients who underwent single-stage proctocolectomy for Crohn's disease between 1958 and 1997 were reviewed. Factors affecting the incidence of recurrence were examined using a multivariate analysis. RESULTS: Principal indications for proctocolectomy were chronic colitis (49 percent), acute colitis (37 percent), and anorectal disease (14 percent). The commonest postoperative complication was delayed perineal wound healing (n = 36; 35 percent), followed by intra-abdominal sepsis (17 percent) and stomal complications (15 percent). In 23 patients the perineal wound healed between three and six months after proctocolectomy, whereas in 13 patients the wound remained unhealed for more than six months. There were two hospital deaths (2 percent) caused by sepsis. The 5-year, 10-year, and 15-year cumulative reoperation rates for small-bowel recurrence were 13, 17, and 25 percent, respectively, after a median follow-up of 18.6 years. From a multivariate analysis, factors affecting reoperation rate for recurrence were gender (male; hazard ratio 2.4 vs. female; P = 0.03) and age at operation (< or =30 years; hazard ratio 2.6 vs. >30 years; P = 0.04). The following factors did not affect the reoperation rate: duration of symptoms, smoking habits, associated perforating disease, coexisting small-bowel disease, postoperative complications, and medical treatment. CONCLUSIONS: Proctocolectomy for Crohn's disease is associated with a high incidence of complications, particularly delayed perineal wound healing. Proctocolectomy carries a low recurrence rate in the long term. However, young male patients are at high risk of recurrence. PMID- 10696901 TI - Radiation-induced total regression of a highly recurrent giant perianal condyloma: report of case. AB - We report a case of a highly recurrent giant perianal condyloma, or Buschke Lowenstein tumor, which was successfully treated by telecobalt therapy. We conclude that radiation therapy is an optional treatment modality for the management of giant perianal condylomata in selected cases. PMID- 10696902 TI - Formalin instillation for ischemic proctitis with unrelenting hemorrhage: report of a case. AB - A case report of an elderly male with multiple medical problems and hemorrhagic, ischemic proctitis is presented. The proctitis was refractory to all other medical options but responded to topical instillation of 4 percent formalin. PMID- 10696903 TI - Pseudomembranous colitis: report of a severe case with unusual clinical signs in a young nurse. AB - We describe the case of a young and otherwise healthy nurse who developed pseudomembranous colitis ten days after receiving oral clindamycin for dental infection. Her clinical course was particularly stormy and was characterized by severe diarrhea and vomiting, profuse ascites, pleural effusion, abdominal tenderness, peritoneal irritation, and systemic toxicity. The Clostridium difficile assay was negative on two occasions. Features compatible with pseudomembranous colitis were seen at sigmoidoscopy, and the diagnosis was confirmed by biopsies. PMID- 10696904 TI - Laparoscopic right colectomy: five-step procedure. AB - The advanced laparoscopic skills required for laparoscopic resection of the colon and rectum have precluded wide dissemination of this procedure. By applying certain key principles, laparoscopic right hemicolectomy can be made simple, reproducible, easy to teach, easy to learn, and cost-effective. PMID- 10696905 TI - Tolerance controls encephalitogenicity of alphaB-crystallin in the Lewis rat. AB - The myelin-associated protein, alphaB-crystallin, is considered a candidate autoantigen in multiple sclerosis (MS). In the present study, we examined the potential of alphaB-crystallin to induce experimental autoimmune encephalomyelitis (EAE) in Lewis rats. Attempts to induce EAE with either bovine, rat or murine alphaB-crystallin or alphaB-crystallin peptides consistently failed. Immunization with either autologous rat or murine alphaB-crystallin did not trigger any antigen-specific T cell response. Immunization with bovine alphaB crystallin or a synthetic peptide representing the cryptic epitope 49-64 did trigger T cell responses but these failed to crossreact with autologous rat alphaB-crystallin. Examination of lymphoid tissues of the Lewis rat revealed constitutive expression of alphaB-crystallin in thymus, spleen, and peripheral lymphocytes. Our data show that in Lewis rats, constitutive lymphoid expression of alphaB-crystallin is associated with a state of nonresponsiveness to autologous alphaB-crystallin that effectively controls the development of EAE in response to this myelin antigen. PMID- 10696906 TI - Differential regulation of neurotrophin expression by mitogens and neurotransmitters in mouse lymphocytes. AB - In this study, we examined the expression of neurotrophins in mouse lymphocytes and the regulation of their expression by mitogens and neurotransmitters. We found that mixed splenocytes as well as T and B lymphocytes expressed mRNA for all the neurotrophins examined. Differential regulation of the neurotrophins was obtained upon stimulation of the cells. Thus, LPS increased the expression of NGF, BDNF and NT-3 in splenocytes and B cells, whereas Con-A increased the mRNA of NT-3 and NT-4 in T cells and NGF expression in splenocytes. The neurotransmitter substance P and the beta-adrenergic agonist, isoproterenol induced an increase in the expression of NGF. Our results suggest an important role for the different neurotrophins in the function of the immune system and point to a bi-directional interaction between neurotrophins and neurotransmitters in this system. PMID- 10696907 TI - Effect of transforming growth factor-beta1 on microglial MHC-class II expression. AB - In the present report, the effects of IFN-gamma and transforming growth factor beta1 (TGF-beta1) on major histocompatibility complex class II (MHC-II) gene expression in isolated mouse brain microglial cells, in the MH-S macrophage cell line and in the primary mouse macrophage cultures were examined. IFN-gamma is a potent inducer of MHC-II gene and this induction was further elevated in microglia by TGF-beta1, while TGF-beta1 inhibited IFN-gamma, induction in macrophages. The enhancing effect of TGF-beta1 was also detected in microglia at the protein level. Transient transfection of microglia with 5' deletional mutants of the MHC-II IAalpha promoter linked to the chloramphenicol acetyltransferase reporter gene demonstrated that TGF-beta1 acts at the transcriptional level to enhance the MHC-II expression induced by IFN-gamma. PMID- 10696908 TI - Alterations in T lymphocyte activity following chemical sympathectomy in young and old Fischer 344 rats. AB - In aged Fischer 344 (F344) rats, sympathetic noradrenergic (NA) innervation of the spleen is markedly diminished compared with young rats. To determine if diminished NA innervation can still provide functional signals to splenic T cells, young (3 months old) and old (17 months old) F344 rats were treated with the NA-selective neurotoxin, 6-hydroxydopamine (6-OHDA) to destroy peripheral NA nerve fibers. In 3-month-old rats, no alterations in spleen cell Con A-induced T cell proliferation, IL-2 or IFN-gamma production were observed up to 15 days after sympathectomy, when splenic NE was maximally depleted. By 21 days post sympathectomy, when NE levels had partially recovered, Con A-induced proliferation and IFN-gamma production, but not IL-2 production, were reduced in sympathectomized animals. After day 21 post-sympathectomy, no alterations in T cell functions were observed in sympathectomized animals. In 17-month-old rats, spleen cell Con A-induced proliferation and IL-2 production were reduced 5 days after sympathectomy in the absence of changes in CD5+ T cells or IFN-gamma production. Desipramine pretreatment, to block 6-OHDA uptake and prevent sympathectomy, completely blocked the 6-OHDA-induced effects, demonstrating that the destruction of NA nerve fibers is required. After day 5 post-sympathectomy, no sympathectomy-induced alterations in Con A-induced T cell functions were observed in old animals. These differences between young and old rats demonstrate that old animals are more susceptible to loss of sympathetic NA innervation, perhaps because compensatory mechanisms are limited. The sympathectomy-induced reduction in T cell proliferation indicates that splenic NA innervation in old animals, though diminished, can exert a positive regulatory influence on T lymphocyte function. Further study of sympathetic neural-immune interactions in the aged rat may provide a means to improve T cell responsiveness in aging. PMID- 10696909 TI - Beta7 integrins contribute to demyelinating disease of the central nervous system. AB - A role for alpha4 integrins in different forms of the multiple sclerosis-like disease experimental autoimmune encephalomyelitis (EAE) has been demonstrated, but the individual contributions of alpha4beta1, alpha4beta7, and the related alphaEbeta7 integrin have not been determined. The P7 integrins alpha4beta7 and alphaEbeta7 play a central role in chronic inflammation, mediating the trafficking, entry, and/or adhesion of lymphocytes in the inflamed pancreas and gut, and their ligands MAdCAM-1, VCAM-1 and E-cadherin are expressed on brain endothelial cells and/or on microvessels in the inflamed central nervous system. Here, we show that an antibody directed against the beta7 subunit greatly attenuates a non-remitting form of EAE, induced by adoptive transfer of myelin oligodendrocyte peptide (MOG35-55)-stimulated T cells. Combinational treatment with both anti-beta7 and alpha4 integrin subunit antibodies led to more rapid and complete remission than that obtained with anti-alpha4 antibody alone, potentially implicating a role for alphaEbeta7 in disease progression. Remission correlated with the down-regulation of the vascular addressins VCAM-1. MAdCAM-1, and ICAM-1 on cerebral blood vessels. Attenuated forms of disease were induced by adoptive transfer of either wild-type encephalitogenic T cells to beta7-deficient gene knockout mice, or of beta7-/-encephalitogenic T cells to wild-type recipients. The former finding indicates that beta7 + ve recruited cells contribute to disease progression. Thus alpha4beta1, alpha4beta7, and alphaEbeta7 integrins may all play a contributory role in the progression of chronic forms of demyelinating disease, and together with their ligands could represent potential targets for improved treatment of some forms of multiple sclerosis. PMID- 10696910 TI - The effect of acute immobilization stress on the abundance of corticotropin releasing factor receptor in lymphoid organs. AB - We have previously found a dramatic increase of corticotropin-releasing factor receptor (CRF-R1) production in splenic neutrophils of male C57BL/6N mice after application of an immunological stimulus. We demonstrate here that immobilization, a predominantly psychological stress, exhibited a similar effect. Shortly after 90 min of immobilization, the number of splenic CRF-RI+ cells was transiently increased by nearly 8-fold, while it was reduced in thymus and unchanged in lymph nodes. The CRF-R1+ cells were detected by an affinity-purified polyclonal antibody directed against the N-terminus of CRF-R1, and identified as neutrophils, eosinophils or their immature precursors on the basis of their nuclear shapes, Wright-Giemsa staining and colocalization of CRF-R1 with the ER MP58 antigen. PMID- 10696911 TI - Chemokines are produced in the brain early during the course of experimental African trypanosomiasis. AB - African trypanosomiasis is characterized by progressive central nervous system (CNS) involvement. Using single and double immunohistochemistry, we evaluated the induction of alpha- and beta-chemokines in brains of Sprague-Dawley rats infected with Trypanosoma brucei brucei (T. b. brucei) and identified their cellular source. The results showed high production of MIP-2, RANTES and MIP-1alpha and to a lower extend MCP-1 in infected animals compared to controls. MIP-2, RANTES and MIP-1alpha were produced early by astrocytes and microglia and later by macrophages and T-cells. These findings suggest that chemokines may contribute to the immunopathogenesis that occurs in the CNS early during infections. PMID- 10696912 TI - 1,25-dihydroxyvitamin D3 treatment decreases macrophage accumulation in the CNS of mice with experimental autoimmune encephalomyelitis. AB - Sunlight, which is required for vitamin D biosynthesis, may be protective in multiple sclerosis (MS), due to the immunoregulatory functions of 1,25 dihydroxyvitamin D3 (1,25-(OH)2D3), the hormonally active vitamin D metabolite. This hypothesis provided the impetus for the experiments reported here investigating mechanisms whereby 1,25-(OH)2D3 may inhibit murine experimental autoimmune encephalomyelitis (EAE). Severe EAE was induced, 1,25-(OH)2D3 or mock treatment was administered, and clinical disease, histopathological disease, and encephalitogenic cells in the central nervous system (CNS) were analyzed within 24-72 h of the treatment. The mock-treated mice remained paralyzed (stage 3 EAE) while most hormone-treated animals regained the partial use of both hind limbs (stage 2 EAE) within 72 h of treatment. A histopathological examination showed the hormone-treated mice had a 50% decrease in white matter and meningeal inflammation at 72 h post treatment. A flow cytometric analysis of cell surface markers on spinal cord cells recovered 24 h post treatment showed the mock treated mice with EAE had about 7.0 +/- 2.3 million Mac-1+ cells/cord, whereas the hormone-treated mice had about 2.1 +/- 2.6 million Mac-1+ cells/cord, which was not significantly different from the unmanipulated control mice. Otherwise, the flow cytometric analysis detected no significant differences between the groups with respect to CD4+ or CD8+ T cells or B cells or macrophages in draining lymph nodes or spinal cords. These results are discussed with regard to possible fates for the 5 million Mac-1+ cells that were rapidly lost from the inflamed CNS in the 1,25-(OH)2D3-treated mice, and the possible beneficial effect of hormone treatment in resolving acute MS. PMID- 10696913 TI - Melatonin is responsible for the nocturnal increase observed in serum and thymus of thymosin alpha1 and thymulin concentrations: observations in rats and humans. AB - This paper shows that melatonin regulates both thymosin alpha1 and thymulin production as well as the expression of the prothymosin alpha gene. The results revealed the following facts: (a) The concentrations of thymosin alpha1 in both serum and thymus of rat showed a nyctohemeral profile with peak values late at night and basal values during the day. The concentrations of thymulin in rat serum also showed a 24-h rhythm with an increase in their values at night. This rhythmical character for thymosin alpha1, and thymulin was also found in the human serum. (b) Rats injected with melatonin during the day exhibited a significant increase in the concentrations of both peptides. Moreover, continuous light exposure on the animals at daytime and pinealectomy cause a decrease in thymosin a1 and thymulin concentrations with regards to those found in control rats. (c) Melatonin regulates the expression of the prothymosin alpha gene, analyzed by Northern blot. These results suggest that melatonin may be involved in the regulation of immune functions by increasing the thymic peptides production. PMID- 10696914 TI - Copolymer 1 inhibits experimental autoimmune uveoretinitis. AB - Copolymer 1 (Cop 1) inhibits experimental allergic encephalomyelitis induced by a variety of myelin proteins, but has been found ineffective so far in inhibiting other experimental autoimmune diseases such as diabetes or arthritis. Here, we report for the first time that Cop I inhibits the development of experimental autoimmune uveoretinitis, induced in mice by interphotoreceptor retinoid-binding protein (IRBP). Pooled data of three experiments showed that treatment with Cop 1, at 0.5 mg/mouse, reduced the disease severity by 53% ( p = 0.0002). Cop 1 treatment also inhibited the proliferation and the production of cytokines by lymph node cells in response to IRBP and moderately reduced the antibody response to this antigen. The possible mechanisms of EAU inhibition by Cop 1 are discussed. PMID- 10696915 TI - Oligodendroglia are protected from antibody-mediated complement injury by normal immunoglobulins ("IVIg"). AB - High-dose intravenous immunoglobulin (IVIg) treatment has become a promising immune therapy that can modulate the immune system at several levels, including the complement cascade. In relation to inflammatory demyelinating disease, there is some clinical evidence for the suppression of disease activity by IVIg, while a role in promoting remyelination after experimental myelin damage has been described. Antibody and complement deposition have been implicated in the immune attack in some cases of multiple sclerosis (MS), and to investigate the mechanisms of action of IVIg, we studied the effect of IVIg using the model of complement-mediated cell injury on oligodendroglia in vitro. There was no effect on direct complement lysis of the oligodendroglial cell line CG4, but antibody dependent complement damage was inhibited in a dose-dependent manner by IVIg. These results were confirmed with primary cultures of oligodendrocyte precursor cells (OPC) and oligodendrocytes. The addition of excess C1, C3, and C4 did not influence the inhibitory effect of IVIg, implying that binding of these complement components does not play a role, in contrast to other experimental models of complement damage. F(ab')2 immunoglobulin fragments were at least partially responsible for the effect. We conclude that IVIg may be protective in antibody-mediated complement injury of oligodendrocytes and their progenitors, and that this effect is likely to be mediated via antibody binding, rather than interference with complement activation. Inhibition of inflammatory mechanisms, as opposed to a direct effect on remyelinating cells, may underlie the role of IVIg in promoting myelin repair in experimental models. PMID- 10696916 TI - Reduction of both pro- and anti-inflammatory cytokines after 6 months of interferon beta-1a treatment of multiple sclerosis. AB - Treatment of multiple sclerosis (MS) with interferon beta (IFNbeta) reduces relapse rate, magnetic resonance imaging (MRI) activity and progression of disability. It has been suggested that this beneficial effect is paralleled by an inhibition of proinflammatory cytokines such as interferon gamma (IFNgamma) and tumor necrosis factor alpha (TNFalpha) and an induction of anti-inflammatory cytokines such as interleukin-4 (IL-4) and interleukin-10 (IL-10). In this study, we record a reduced number of spontaneously IFNgamma mRNA-expressing cerebrospinal fluid mononuclear cells (CSF-MC) and IFNgamma, TNFalpha and IL-10 mRNA-expressing peripheral blood mononuclear cells (PBMC) after 6 months of IFNbeta-1a treatment, paralleled by a decreased purified protein derivate (PPD) stimulated and unstimulated IFNgamma secretion by PBMC. These effects were not apparent after 2 weeks of treatment, and IFNbeta-1a induced IFNgamma production by naive PBMC in vitro. We did not record increased numbers of IL-4 mRNA expressing CSF-MC or PBMC, increased plasma IL-10 levels, increased numbers of IgG, A or M secreting plasma cells or in vitro induction of IL-10 production by IFNbeta-1a. We conclude that long-term cytokine modulation by IFNbeta-1a differs from acute effects and that downregulation of both pro- and anti-inflammatory cytokines, rather than a shift in the cytokine profile, is apparent after 6 months of IFNbeta-1a treatment of MS patients. PMID- 10696917 TI - The hemodynamic effects of cocaine during acute controlled hemorrhage in conscious rats. AB - BACKGROUND: Cocaine is often associated with trauma; however, little is known about how its use alters the response to blood loss. The effect of cocaine on hemodynamics following acute hemorrhage was studied in a rat model. METHODS: Following baseline measurements, rats were administered either intravenous cocaine, or saline as a control. Both groups then underwent arterial catheter hemorrhage of 30% of total blood volume. Outcome variables include blood pressure, heart rate, hematocrit, pH, PCO2, PO2, and serum bicarbonate. RESULTS: Following hemorrhage, blood pressure decreased in both groups but the hypotension was significantly greater in the saline group than the intravenous cocaine group at 0 and 5 minutes posthemorrhage. Heart rate was increased significantly for the intravenous cocaine group compared to the saline group starting at 15 minutes postcocaine and lasting for the next 25 minutes. No difference was noted for hematocrit, pH, PO2, or serum bicarbonate. CONCLUSION: Although transient, cocaine blunted the hypotensive response to acute controlled hemorrhage and resulted in tachycardia. PMID- 10696918 TI - Fumigant-related illnesses: Washington State's five-year experience. AB - OBJECTIVE: Exposure to fumigants may have severe or persistent health effects. Washington State's fumigant-related illnesses were reviewed to better understand the circumstances surrounding exposure and resultant health effects. METHODS: Fumigant-related illnesses reported to and investigated by the Washington State Department of Health were reviewed. Illnesses considered by Department of Health to be definitely, probably, or possibly related to pesticide exposure were then analyzed. RESULTS: From 1992-1996, 39 (3.3%) of 1192 definite, probable, or possible cases of pesticide-related illnesses involved exposures to fumigants. Fumigant exposures during this period were to aluminum phosphide (15), methyl bromide (12), metam-sodium (9), and zinc phosphide (3). Symptoms included respiratory problems and eye and/or skin irritation for the majority of exposures, and no deaths were reported. The nature of exposure for these cases included exposure to applicators (17), reentry into a fumigated structure (9), improper storage or disposal (6), reentry into treated agricultural fields (4), drift from treated fields (2), and other (1). CONCLUSIONS: Review of fumigant exposures should be used to prevent future events through continued enforcement of established regulations and training of applicators. PMID- 10696919 TI - A nationwide survey of the management of unintentional-low dose tricyclic antidepressant ingestions involving asymptomatic children: implications for the development of an evidence-based clinical guideline. AB - BACKGROUND: The triage of unintentional tricyclic and cyclic antidepressant ingestions involving children <6 years seems based on single cases or small studies. Walsh, in describing 2 cases involving 15-20 mg/kg ingestions, recommended hospitalizing all children ingesting tricyclic and cyclic antidepressants. OBJECTIVE: To evaluate the patterns of triage for pediatric tricyclic and cyclic antidepressants practiced by regional poison control centers nationwide, and to determine the amount ingested (mg/kg) that resulted in referral to the emergency department, including the recommended duration of observation time for asymptomatic children. Second, to analyze the cost implications, as well as the need for a practice guideline based on severity stratification. METHODS: We sent a survey to 44 major regional poison control centers. We reviewed Health Care Financing Administration's tricyclic and cyclic antidepressants management related costs. RESULTS: Thirty centers responded (68%). Eighty-seven percent of all centers send children, regardless of dose ingested, to the emergency department. Four out of the 30 recommended observation based on dose in mg/ kg ingested (range >1.5-5). Recommended observation times in the emergency department varied between 6-24 hours. Twenty-seven (90%) Poison Control Centers recommended 6 hours (although one recommended doing so only after administering activated charcoal). One recommended 6-12 hours of observation and 2 Poison Control Centers recommended 24-hour observation. Only 1 center recommended obtaining tricyclic and cyclic antidepressant plasma levels. DISCUSSION: In our review of the literature, the lowest toxic dose reported was 6.7 mg/kg. This is consistent with our Poison Control Center data over the past 5 years where no child was toxic at doses <5 mg/kg. While only 13% of the centers surveyed utilize a stratification strategy to triage pediatric tricyclic and cyclic antidepressant ingestions, the current referral patterns support evaluation based on pharmacokinetics, not worst case incidents. CONCLUSION: This survey demonstrates that most children with tricyclic and cyclic antidepressant ingestions will be sent to the emergency department, regardless of the amount ingested. A prospective study is needed to determine the probable dose of tricyclic and cyclic antidepressant ingestions that requires observation at a health care facility. PMID- 10696920 TI - Inadequate stocking of antidotes in Taiwan: is it a serious problem? AB - OBJECTIVE: Insufficient hospital stock of a variety of poisoning antidotes is a worldwide problem. In an attempt to establish an antidote storage and distribution system for the response of the various poisoning accidents, we conducted a nationwide survey to characterize the current availability of selected antidotes and their anticipated need in Taiwan. MATERIALS AND METHODS: A questionnaire was mailed to 834 hospitals to gather information on the availability, anticipated need, and preferred purchase policy of 20 selected antidotes. A survey on the availability of cyanide antidote in 523 cyanide handling facilities and their neighboring hospitals was also conducted. RESULTS: Hospitals of different size and service levels had a statistically significant difference in response rates. Except for pyridoxine, the availability and anticipated need for antidotes also varied significantly among different hospital groups. We found that physostigmine, cyanide antidote kit, BAL, EDTA, methylene blue, Vipera Russell formosensis antivenin, and botulism antitoxin were not available in most (>90%) hospitals. Interestingly, these antidotes are also among the most needed antidotes. Most hospitals preferred a government-ordered purchase of antidotes. In the survey of cyanide-processing facilities, a response rate of 24.1% was obtained and only 9.3% of these 107 facilities that both replied to the questionnaire and continued handling cyanide products had stocked cyanide antidote. It is noteworthy that cyanide antidote was also frequently lacking in the neighboring hospitals. CONCLUSIONS: The appropriate storage of antidotes in hospitals or workplaces in rural areas is instrumental in the timely treatment of certain poisonings, while nationwide unavailability is the critical problem. Raising awareness of the importance of antidotes by education, regular review of antidote storage, distribution plans, and appropriate legislation might provide solutions. PMID- 10696921 TI - Sources of information for acute poisoning in accident and emergency departments in Dublin, Ireland. AB - BACKGROUND: Access by accident and emergency staff to up-to-date information on poisoning is essential for optimal management of acute poisoning. Apart from the National Poisons Information Centre, other information sources can be used. The objectives of the study were to identify sources of information on acute poisoning in accidents and emergencies and satisfaction with their use. METHODS: In a cross-sectional survey of medical staff of accidents and emergencies in Dublin in 1997, data were collected by interviewer-administered questionnaire. RESULTS: All 11 accidents and emergencies participated, with a staff response rate of 95%. One hundred and twenty-eight respondents were included. Ninety-seven percent had managed cases of poisoning (median 3 per week). The National Poisons Information Centre had been used by 93% of respondents, textbooks by 80%, paper database by 63%, and disc database (CD-ROM) by 10%. Of those managing cases, the National Poisons Information Centre would always be contacted by 23% and by 53% in most cases. The National Poisons Information Centre and CD-ROMs were rated the most useful sources of information. Information provided by the National Poisons Information Centre was considered sufficient by 98% of respondents. Thirty-three percent considered that advice should always be confirmed by fax. Limitations with the National Poisons Information Centre were described by 55% (e.g., manual transcription), with textbooks (e.g., limited content) by 83%, with paper databases (e.g., incompleteness) by 85%, and with CD-ROMs (e.g., time-consuming) by 54%. CONCLUSION: For the optimal management of acute poisoning, direct access to computerized information databases in accidents and emergencies combined with telephone access to the National Poisons Information Centre is required, with information available in hard copy. PMID- 10696922 TI - Cerebellar dysfunction in chronic toluene abuse: beneficial response to amantadine hydrochloride. AB - CASE REPORT: A 21-year-old man who had sniffed toluene since the age of 13 presented with a 4-year history of progressive cerebellar dysfunction and visual deterioration. The patient's condition did not improve despite 5 months of abstinence. Magnetic resonance imaging revealed cerebral atrophy and hypointensity signals in the white matter and bilaterally in the globus pallidus, thalamus, red nucleus, and substantia nigra. Amantadine hydrochloride therapy (100 mg/d, then 200 mg/d) resulted in dramatic improvement of his cerebellar and visual symptoms. PMID- 10696923 TI - Carbofuran-induced delayed neuropathy. AB - BACKGROUND: Although carbamates have been widely used in the world for many years, carbamate-induced delayed neuropathy is rare. We report what appears to be delayed neuropathy caused by poisoning with carbofuran, a cholinesterase inhibiting carbamate, although the certainty of diagnosis is somewhat limited by the lack of a sural nerve biopsy and spinal fluid examination. CASE REPORT: A 23 year-old man attempted suicide by ingesting 100 mL of carbofuran (2,3-dihydro-2,2 dimethyl-7-benzofuranyl methylcarbamate). After recovering from acute cholinergic toxicity, he had notable paresthesia in his lower limbs and difficulty walking. Electrophysiologic findings revealed sensorimotor neuropathy. Recovery began at 1 week and continued for 4 months. A similar delayed neuropathy has been described with carbamate, 1-naphthyl N-methylcarbamate, and m-tolyl methylcarbamate, but not with carbofuran insecticides. PMID- 10696924 TI - Intermediate syndrome after malathion ingestion despite continuous infusion of pralidoxime. AB - CASE REPORT: A 33-year-old female ingested an unknown quantity of malathion in a suicide attempt. Cholinergic signs consistent with severe organ, phosphate intoxication developed and were treated within 6 hours of ingestion. Intravenous atropine and a continuous infusion of pralidoxime (400 mg/h) were administered. Prolonged depression of plasma and red blood cell cholinesterases were documented. Despite an initial clinical improvement and the presence of plasma pralidoxime concentrations exceeding 4 microg/mL, the patient developed profound motor paralysis consistent with the diagnosis of Intermediate Syndrome. In addition to the dose and frequency of pralidoxime administration, other factors including persistence of organophosphate in the body, the chemical structure of the ingested organophosphate, and the time elapsed between ingestion and treatment may limit the effectiveness of pralidoxime as an antidote in organophosphate ingestions. This case study suggests that these factors should be taken into account in assessing the risk of Intermediate Syndrome after intentional organophosphate ingestions. PMID- 10696925 TI - Fatal cardiovascular collapse following acute colchicine ingestion. AB - BACKGROUND: A previously published prognostic rule predicts 100% survival after ingestion of cochicine doses less than 0.5 mg/kg and 100% mortality after ingestion of more than 0.8 mg/kg. This rule inaccurately predicted survival in a recent case. CASE REPORT: We present a case of fatal colchicine poisoning in an adult who ingested a maximum of 39.6 mg of colchicine (0.40 mg/kg). He subsequently developed hypotension which was refractory to fluid resuscitation and infusion of vasopressors. He died of cardiovascular collapse approximately 35 hours after ingestion. DISCUSSION: Fatal outcomes are possible even with colchicine doses less than 0.5 mg/kg. Physicians caring for colchicine-poisoned patients must be prepared for the possibility of acute cardiovascular collapse and ventricular dysrhythmias regardless of the reported dose of colchicine. PMID- 10696926 TI - Massive ibuprofen ingestion with survival. AB - OBJECTIVE: To report a massive, 100 g ibuprofen ingestion in an adolescent, with survival. CASE REPORT: The patient developed coma, metabolic acidosis, and mild thrombocytopenia, but improved rapidly with supportive care. Renal function remained normal and no gastrointestinal bleeding occurred. CONCLUSIONS: Massive ingestion of ibuprofen may result in a variable picture with some elements of significant toxicity, but supportive care usually results in survival without sequelae. PMID- 10696927 TI - Pulse steroid therapy in adult respiratory distress syndrome following petroleum naphtha ingestion. AB - CASE REPORT: A suicide attempt by a 23-year-old woman involved ingestion of 1000 mL of petroleum naphtha. Early chemical pneumonitis was complicated by life threatening, diffuse interstitial lung consolidation with pneumatoceles. Pulse steroid therapy beginning on day 17 was associated with remarkable resolution of interstitial consolidation, although an enlarging secondarily infected pneumatocele ruptured to produce a bronchopleural fistula. Thoracic surgery and antibiotic therapy resulted in improvement of the patient's respiratory condition, and she was discharged with no residual respiratory symptoms. High dose corticosteroid therapy appears to be a useful addition to aggressive supportive treatment in late adult respiratory distress syndrome following hydrocarbon ingestion. PMID- 10696928 TI - Toxicity of Passiflora incarnata L. AB - BACKGROUND: Herbal medicines may have significant adverse effects which are not suspected or recognized. CASE REPORT: A 34-year-old female developed severe nausea, vomiting, drowsiness, prolonged QTc, and episodes of nonsustained ventricular tachycardia following self-administration of a herbal remedy, Passiflora incarnata L., at therapeutic doses. The possible association of symptoms with passiflora was not recognized for several days. She required hospital admission for cardiac monitoring and intravenous fluid therapy. CONCLUSIONS: Passiflora incarnata was associated with significant adverse effects in this patient. It is important to ask specifically about the use of herbal medicines in patients with undiagnosed illnesses. PMID- 10696929 TI - Hepatotoxicity from castor bean ingestion in a child. AB - BACKGROUND: Castor beans contain ricin, one of the most toxic substances known. A biphasic toxicity is described consisting of acute and potentially fatal gastroenteritis followed by damage to the viscera several days later. However, the most current review of the literature states that the above delayed toxicity has not been reported. CASE REPORT: We report a 20-month-old girl with no gastrointestinal symptoms who developed reversible hepatotoxicity beginning 48-72 hours after the ingestion of castor beans. CONCLUSION: It seems prudent to follow for several days any patients who actually chewed castor beans before ingestion. PMID- 10696930 TI - Payment deferred: strychnine poisoning in Nicaragua 65 years ago. AB - BACKGROUND: In 1933 in Leon, Nicaragua, a 22-year-old woman died after an acute convulsive illness in which she experienced trismus, opisthotonos, and hyperpyrexia. Three years later her husband, Oliverio Castaneda, was convicted of her murder and that of 2 other people in the same city. METHODS: We went to Nicaragua to investigate documents involved with that case and evaluate whether the verdict of murder by strychnine was substantiated by the data. We present the results of the investigation and provide information about the practice of medicine, pharmacy, and toxicology early in this century. RESULTS: The clinical picture in all 3 cases suggests strychnine poisoning. The clinical, toxicological, and circumstantial evidence is strong and implicates Castaneda as a murderer and strychnine as the weapon. CONCLUSION: We conclude that Oliverio Castaneda was the probable perpetrator of three 1933 strychnine murders in Leon and that he may have previously used strychnine to kill others in Nicaragua and neighboring countries. PMID- 10696931 TI - Does nebulized corticosteroid therapy have an effect on ammonia-induced pulmonary injury? PMID- 10696932 TI - Corticosteroid inhalation therapy has no proven effect on gas exchange or airway pressure levels in ammonia-induced lung injury. PMID- 10696933 TI - Measuring cancer care. PMID- 10696934 TI - Basic life support: extending and integrating teaching in the Australian community. PMID- 10696935 TI - In-hospital mortality and associated complications after bowel surgery in Victorian public hospitals. AB - BACKGROUND: The purpose of the present paper was to determine the mortality rate and associated complications after large bowel resection and anastomosis in Victorian public hospitals. METHODS: A retrospective analysis of data from the Victorian Inpatient Minimum Database (VIMD) was undertaken. The data were collected from all Victorian public hospitals performing hemicolectomy and anterior resection (resection of the rectum with anastomosis) from 1987/88 to 1995/96. RESULTS: A total of 11036 patients underwent hemicolectomy or anterior resection in the time period studied, there being a 7% increase in the rate of operations performed over the 9 years. Two-thirds of these operations were for carcinoma of the large bowel. The anastomotic leak rate of 4.5% fell slightly but the in-hospital mortality rate of 6.5% did not change over the study period. The total morbidity recorded (mainly major complications) was 24.6%. The patients most at risk of death were the elderly with pre-existing cardiac or respiratory disease undergoing an emergency operation. CONCLUSIONS: Notwithstanding some inaccuracies of coding and reporting, the morbidity and mortality for surgery of the large intestine remains high, largely due to the comorbidities of the patients, although certain technical complications such as leakage of an anastomosis after anterior resection are still associated with a significantly increased risk of death. Consideration should be given to the routine use of high dependency nursing units for these high-risk patients after major colorectal surgery, and support from physicians to reduce morbidity and mortality from associated medical conditions worsened by surgery. PMID- 10696936 TI - A population-based study of the incidence, mortality and outcomes in patients following surgery for colorectal cancer in Western Australia. AB - BACKGROUND: The literature contains many reports on the management of colorectal cancer from single institutions or groups of specialist surgeons. But there are few data on community-wide patterns of treatment or the outcomes of colorectal surgery. The aim of the present study was to use a population-based linked database to assess the trends in colorectal cancer incidence and mortality in Western Australia (WA) in the period 1982-95, and to evaluate the outcomes following surgical care. METHODS: A population-based linked database was used to relate the cancer registry, hospitalization and mortality records of all patients with a diagnosis of colorectal cancer in WA during 1982-95. Data on surgical treatment and postoperative morbidity and mortality in this group of patients were available only in 1988-95. Patient records were selected using the international classification for diagnosis and procedure codes pertaining to colorectal cancer and surgery. Incidence and mortality trends in colon and rectal cancers were estimated by Poisson regression regression of age-standardized rates, and relative survival analysis was used to compare patient survival with the general population. RESULTS: During the 14-year period, 9673 patients presented with a diagnosis of colorectal cancer. The sex distribution of patients with colon cancer was evenly divided, but rectal cancer was more common in men (ratio 1:4). The mean age at diagnosis was 67.8 years (SD: 12.7). During the study period there was a significant increase in the standardized incidence rate of rectal cancer in men, and in the mortality rates from colon cancer in women. The overall crude 5-year survival was 57%. Large bowel resections were performed on 71% of patients with an in-hospital postoperative mortality of 4.2%. CONCLUSION: Colorectal cancer is a continuing major cause of morbidity and mortality in WA. The present study demonstrated increases in the incidence rate of rectal cancer in men and in the mortality rate from colon cancer in women in the period 1982-95. PMID- 10696937 TI - Cut or paste? The use of glyceryl trinitrate paste in the treatment of acute and chronic anal fissure. AB - BACKGROUND: Anal fissure unresponsive to conservative measures such as stool softeners frequently requires surgical intervention. The present study describes the use of glyceryl trinitrate (GTN) in the treatment of acute and chronic anal fissure. METHODS: Eighteen consecutive patients with anal fissure were treated with 0.5% GTN paste in soft white paraffin applied twice daily to the anus. These patients were followed at regular intervals to assess symptom control, rate of healing, adverse effects and recurrence rate. RESULTS: Two patients were lost to follow-up. Twelve of the remaining 16 were cured. Of these, symptomatic relief was obtained for most within 2 days, and for all within 1 week. No patient required cessation of treatment due to adverse effects. Treatment failed in four of 16 patients. Two of these patients subsequently underwent successful surgical procedures, and two patients (while not completely cured) had sufficient symptomatic relief to decide against surgery. CONCLUSIONS: The use of GTN paste in the treatment of acute and chronic anal fissure may be a safe and effective modality that can be considered as first-line treatment for this condition. PMID- 10696938 TI - Open Nissen fundoplication and highly selective vagotomy as a treatment for gastro-oesophageal reflux disease. AB - BACKGROUND: Open Nissen fundoplication has been the most common surgical treatment of gastro-oesophageal reflux disease (GORD). The present paper describes the symptomatic result, and quantifies the acid reduction achieved by open Nissen fundoplication combined with highly selective vagotomy (HSV) in a consecutive case series. METHODS: A study of 106 patients undergoing open Nissen fundoplication and HSV for GORD was performed between 1988 and 1996. A history consistent with reflux was obtained and the diagnosis confirmed with ambulatory oesophageal pH studies and endoscopy. Postoperative pH studies were performed, and peri-operative and late complications were recorded. A standard questionnaire was sent out to patients postoperatively to assess the efficacy of surgery. RESULTS: Pre-operative pH studies were performed on 104 (98%) patients, and on 97 (91.5%) following surgery. There was a highly significant improvement in all parameters of the pH study postoperatively. All symptoms, including bloating and dysphagia, improved significantly postoperatively, except flatulence, which was exacerbated. The majority of patients were very satisfied with their outcome, 82% rating the operation from 80 to 100% successful. Complications were rare and there was no mortality. CONCLUSIONS: Open Nissen fundoplication and HSV is an effective method of treating GORD, producing an improvement in symptoms and in ambulatory pH studies. PMID- 10696939 TI - Prevalence of Helicobacter pylori infection in 160 patients with Barrett's oesophagus or Barrett's adenocarcinoma. AB - BACKGROUND: The role of Helicobacter pylori infection in the development of Barrett's oesophagus and its complications is uncertain. The aim of the present study was to determine the importance of H. pylori infection in this disease by comparing the frequency of oesophageal and gastric H. pylori infection in a group of patients with Barrett's oesophagus or adenocarcinoma, with the frequency of infection in a control group without Barrett's disease. METHODS: The study group included 160 patients (123 male, 37 female; mean age: 61.2 years) who were classified (according to the highest grade pathological lesion in the oesophagus) as having Barrett's intestinal metaplasia (IM; 88 patients), Barrett's oesophagus with low-grade dysplasia (LGD; 28 patients), high-grade dysplasia (HGD; five patients), Barrett's indefinite for dysplasia (n = 4), and Barrett's adenocarcinoma (33 patients). A total of 91 of these patients had gastric antral specimens available for study. The control group consisted of 214 consecutive, prospectively enrolled symptomatic patients (122 male, 92 female; mean age: 57.2 years) who underwent upper gastrointestinal endoscopy and in whom Barrett's oesophagus or Barrett's adenocarcinoma was not found. A modified Warthin-Starry method was used to detect H. pylori infection. RESULTS: Oesophageal H. pylori infection was found in eight of 160 (5%) patients with Barrett's oesophagus or Barrett's adenocarcinoma. Holicobacter pylori organisms in the oesophagus were found only on non-intestinalized cardiac or oxyntocardiac mucosa. All patients with oesophageal H. pylori infection and an antral biopsy available for study had antral H. pylori infection. Gastric antral H. pylori infection was significantly less prevalent in patients in the Barrett's study group (15/91, 16.5%) than in the non-Barrett's control group (67/214, 31.3%; Fisher's exact test, P = 0.01). Patients from the control group with an endoscopic diagnosis of duodenal ulcer, gastric ulcer, gastritis, or duodenitis had a significantly higher prevalence of infection compared with the Barrett's group, but there was no difference in the infection prevalence in patients in the Barrett's group and patients with reflux oesophagitis, hiatal hernia, no endoscopic abnormality, or any other diagnosis. CONCLUSIONS: Oesophageal H. pylori infection is uncommon in patients with Barrett's IM, dysplasia, or adenocarcinoma, and may be restricted to non intestinalized columnar epithelium. Gastric H. pylori infection may have a protective effect for the development of Barrett's oesophagus. PMID- 10696940 TI - Supratrigonal cystectomy and ileocystoplasty in management of interstitial cystitis. AB - BACKGROUND: Interstitial cystitis is a chronic non-infectious inflammatory disease of the bladder of unknown aetiology which is characterized by irritative voiding symptoms and suprapubic pain related to bladder filling. Surgical treatment is indicated in severely symptomatic patients when medical therapies have failed, usually after a period of several years. The authors' experience with a modified technique of ileocystoplasty following supratrigonal cystectomy performed in five patients with interstitial cystitis is presented here. METHODS: A modified technique of bladder augmentation using ileum following supratrigonal bladder resection is described. RESULTS: All patients experienced relief from their symptoms. No patient had residual bladder pain and urinary frequency settled down in all. Bladder capacity was increased significantly. Three patients voided spontaneously postoperatively and two required clean intermittent self catheterization. CONCLUSIONS: Supratrigonal cystectomy and ileocystoplasty can be a satisfactory option in refractory cases of interstitial cystitis. A simplified technique of ileal bladder construction that provides satisfactory bladder capacity is presented. Most urologists are familiar with ileal surgery, having used the ileum as a conduit after cystectomy for urinary diversion. PMID- 10696941 TI - A protocol of early spiral computed tomography for the detection of stones in patients with renal colic has reduced the time to diagnosis and overall management costs. AB - BACKGROUND: The recent use of spiral computed tomography (CT) without contrast for the diagnosis of acute flank pain has been shown to be highly sensitive and specific for the detection of urolithiasis. This method has not, however, been evaluated for its contribution to savings in management costs. The present study aims to evaluate the cost savings gained by instituting a protocol of early spiral CT to investigate these patients. METHODS: The records of 200 patients presenting to the Accident and Emergency Department (A&E) with acute flank pain during two periods were retrospectively reviewed. The first period was before the spiral CT protocol was instituted and the second was after. Cost analyses between the two periods were performed. RESULTS: After the spiral CT protocol, 72 versus 31 patients had a definitive diagnosis prior to discharge from hospital. The time taken to diagnosis was also significantly shorter after the protocol implementation (6.3 vs. 16.8 h). This resulted in a shorter time spent in the A&E, and hence bed cost savings. Radiological costs were reduced by 22%, but the major cost saving was made by a reduction in time spent in A&E (44%). CONCLUSIONS: The implementation of a protocol of early spiral CT for patients with suspected renal colic has led to earlier definitive diagnosis and shorter hospital stays. This is associated with a significant reduction in costs associated with managing this condition. PMID- 10696942 TI - Factors that influence length of stay after appendicectomy in children. AB - BACKGROUND: The length of hospital stay following appendicectomy in children at Christchurch Hospital has decreased in recent years. The aim of the present study was to identify those factors that contributed to this change. METHODS: A retrospective review of children admitted to Christchurch Hospital between 1994 and 1998 inclusive who underwent appendicectomy for suspected appendicitis was conducted. Data recorded included standard demographic information, symptom duration, operative details, analgesia, antibiotics, pathology, complications and postoperative length of stay (LOS). RESULTS: Postoperative LOS decreased significantly during the period reviewed across all degrees of appendiceal inflammation, from a mean of 70.5 to 50.1 h. The main determinant of postoperative hospital stay was the severity of the appendiceal inflammatory process. Other factors that influenced LOS included surgical approach (open vs. laparoscopic), use of intra-operative local anaesthesia, type and mode of postoperative analgesia, and age of the child. Longer duration of antibiotic use and symptom duration of greater than 24 h were associated with a longer LOS, primarily as a reflection of the severity of inflammation of the appendix. Factors that appeared to have little or no influence included gender and the experience of the surgeon. CONCLUSION: The severity of the inflammatory process appeared to be the main determinant of postoperative hospital LOS; advanced appendicitis with abscess formation or peritonitis was associated with the longest LOS, irrespective of the surgical approach, although the LOS after appendicectomy was reduced by a laparoscopic approach. Intra-operative local anaesthesia during open appendicectomy reduced hospital stay, probably because it reduced the need for postoperative narcotics. Early diagnosis (< 24 h) was associated with a shorter postoperative LOS for acutely inflamed appendices. PMID- 10696943 TI - Risk factors for surgical wound infection and bacteraemia following coronary artery bypass surgery. AB - BACKGROUND: There has been no consensus from previous studies of risk factors for surgical wound infections (SWI) and postoperative bacteraemia for patients undergoing coronary artery bypass graft (CABG) surgery. METHODS: Data on 15 potential risk factors were prospectively collected on all patients undergoing CABG surgery during a 12-month period. RESULTS: Of 693 patients, 62 developed 65 SWI using the Centres for Disease Control definition: 23 were sternal wound infections and 42 were arm or leg wound infections at the site of conduit harvest. There were 19 episodes of postoperative bacteraemia. Multivariate analysis revealed that: (i) diabetes, obesity and previous cardiovascular procedure were independent predictors of SWI; and (ii) obesity was an independent risk factor for postoperative bacteraemia. CONCLUSIONS: These findings suggest that improved diabetic control and pre-operative weight reduction may result in a decrease in the incidence of SWI. But further prospective studies need to be undertaken to examine (i) whether the increased SWI risk in diabetes occurs with both insulin- and non-insulin-requiring diabetes, and whether improved peri operative diabetes control decreases SWI; and (ii) what degree of obesity confers a risk of SWI and postoperative bacteraemia, and whether pre-operative weight reduction, if a realistic strategy in this patient group, results in a decrease in SWI. PMID- 10696944 TI - Risk factors for peri-operative stroke complicating carotid endarterectomy: selective analysis of a prospective audit of 1000 consecutive operations. AB - BACKGROUND: The aim of the present study was to investigate the role of potential clinical risk factors in the causation of peri-operative stroke associated with carotid endarterectomy. With the change in carotid endarterectomy practice from the use of a shunt to high-dose thiopental for cerebral protection (a previously undocumented method), it was essential to identify accurately the causes of all perioperative strokes. METHODS: A prospective audit was undertaken of 1000 carotid endarterectomies in which the causes and pathology of all peri-operative strokes were documented. The roles of advanced age, female gender, hypertension, previous stroke, contralateral carotid stenosis >70%, and contralateral carotid occlusion as potential causes of peri-operative stroke were defined. Results were statistically analysed using odds ratio and Fisher's exact test. RESULTS: None of the potential risk factors was statistically significant for peri-operative stroke. Female gender was associated with a significant risk of peri-operative stroke due to operative site thrombosis. Complications at the endarterectomy site were the commonest cause of stroke. CONCLUSIONS: Prospective audit is a useful tool for identifying causes of peri-operative stroke and indicating the need for modifications to surgical clinical management which might improve outcomes for carotid endarterectomy. PMID- 10696945 TI - Skin wrinkling for the assessment of sympathetic function in the limbs. AB - BACKGROUND: Wrinkling of the skin of the palm and sole is considered to be dependent on the presence of intact sympathetic nervous activity. Loss of sympathetic integrity could be simply and usefully assessed by the absence of wrinkling. To test this hypothesis, the skin wrinkle test was compared with the starch-iodine sweat test and sympathetic skin response (SSR) in patients with abnormal sympathetic function. METHODS: The three tests were carried out in 34 patients (68 limbs) undergoing temporary or permanent disruption of the sympathetic chain to upper or lower limbs. Included in this group were six diabetics undergoing chemical or surgical sympathectomy, lumbar epidural infusions following vascular surgery, and patients for whom sympathectomy was being considered. Sensitivity and specificity analysis and predictive values of the wrinkling response and the starch-iodine test were related to the SSR as the standard. RESULTS: The wrinkle test showed a sensitivity of 97% and specificity of 95%, and bore good correlation to the SSR. The starch-iodine test showed sensitivity of 55% and specificity of 93%. A hypothesis for the mechanism of wrinkling based on the observations of the present study is proposed. CONCLUSION: The wrinkle test is a reliable test of sympathetic function, is inexpensive and is easy to perform at the bedside. The sweat gland myo-epithelial cells and absence of sebum could play an important role in the wrinkling response. It can be used to select patients who will benefit from sympathectomy, and can adequately evaluate sympathetic blockade. PMID- 10696946 TI - Robert Liston and vascular surgery. AB - Robert Liston was a general surgeon who dealt with all conditions that were presented to him, and in his practice saw a number of cases that involved the vascular system. These were mainly wounds of vessels, aneurysms and vascular tumours. His writings on the subject have been reviewed with some examples of the cases he had to treat. PMID- 10696947 TI - Verapamil reduces intimal hyperplasia in a sheep carotid artery patch graft model. AB - BACKGROUND: The current study investigated the effect of verapamil on the development of intimal hyperplasia (IH) using a sheep model. METHODS: A gelatin sealed Dacron patch graft was implanted into the left common carotid artery in 40 adult Merino sheep. Sheep were then randomly allocated to four groups. Group 1 were controls given no treatment (n = 10). Groups 2-4 were treated with intravenous verapamil 0.5 mg/kg per day in two divided doses for different lengths of time. Group 2 (n = 10) received treatment for 1 week; group 3 (n = 10) received treatment for 2 weeks and group 4 (n = 10) received treatment for 4 weeks. All sheep were killed at 4 weeks and the grafted segments of artery were harvested for IH assessment by an image analysis system. RESULTS: The IH index from the three groups treated with verapamil was significantly less than that of the control (group 1,0.287+/-0.077: group 2,0.205+/-0.064, P<0.05; group 3, 0.193+/-0.059, P<0.01; group 4,0.171+/-0.046, P<0.01). CONCLUSION: The present study suggests that verapamil inhibits the development of IH even when treatment is given for only 1 week. PMID- 10696948 TI - The effect of fundoplication on the motility of the canine lower oesophageal sphincter. AB - BACKGROUND: A canine model was used to define whether Nissen fundoplication inhibits gastro-oesophageal reflux by inhibiting transient lower oesophageal sphincter relaxations (TLOSR) or by creating a pressure barrier at the gastro oesophageal junction. METHODS: Four surgical models were studied pre-operatively and postoperatively. These were: (i) the surgical mobilization required for fundoplication (sham fundoplication, n = 5); (ii) a standard fundoplication (n = 4); (iii) anterior and posterior myotomy of the lower oesophageal sphincter (LOS; cardiomyotomy, n = 4); and (iv) combined cardiomyotomy and fundoplication (n = 4). Each operative procedure was assessed for its effect on the incidence of TLOSR and gas reflux events, the mean LOS pressure and the LOS pressure profile during swallow events. RESULTS: Sham fundoplication reduced the rate of evoked TLOSR in response to gaseous gastric insufflation from 9.8+/-1.6/h (mean +/- SEM) to 5.4 +/-1.5/h. The mean LOS pressure was reduced from 25.1+/-2.6 to 18.5+/-2.1 mm Hg but nadir LOS pressure during swallowing was not altered. Nissen fundoplication virtually abolished evoked TLOSR from 10.4+/-1.2/h to 0.4+/-0.4/h, increased mean basal LOS pressure from 19.8+/-2.1 to 27.0+/-1.1 mm Hg and increased the nadir pressure on swallowing from 3.4+/-1.0 mm Hg to 14.4+/-1.0 mm Hg. Cardiomyotomy was associated with a near continuous leakage of gas across a chronically hypotensive LOS. Cardiomyotomy reduced the resting LOS pressure from 14.7+/-1.2 mm Hg to 2.3+/-1.0 mm Hg. Cardiomyotomy with fundoplication was associated with no loss of LOS competence. No gas venting episodes occurred either by passive leakage or by TLOSR. Cardiomyotomy with fundoplication was associated with a fall in mean LOS pressure from 14.3+/-1.5 mm Hg to 7.1+/-1.8 mm Hg but no LOS relaxation occurred during swallowing. CONCLUSION: Nissen fundoplication is highly effective in preventing reflux across a normal or chronically hypotensive LOS. Fundoplication results in a constant, measurable pressure barrier at the lower end of the oesophagus that is not due to a change in intrinsic LOS tone. Following fundoplication TLOSR are prevented by the constant low-pressure barrier. PMID- 10696949 TI - Practice parameters for the management of colonic cancer I: surgical issues. Recommendations of the Colorectal Surgical Society of Australia: comment. PMID- 10696950 TI - Is the port site really at risk? Biology, mechanisms and prevention: a critical view: comment. PMID- 10696951 TI - Lipohyperplasia of the ileo-caecal valve causing appendicitis. PMID- 10696952 TI - Open heart surgery in a patient with liver cirrhosis and thrombocytopenia. PMID- 10696953 TI - Peripheral renal angiomyolipoma with extension into the renal vein. PMID- 10696954 TI - Fistula between the hip and a diverticular abscess after revision total hip replacement. PMID- 10696955 TI - Hemoglobin and erythrocyte indices during normal pregnancy and postpartum in 206 women with and without iron supplementation. AB - BACKGROUND: The aim was to define reference values for hemoglobin, hematocrit and erythrocyte indices, i.e. erythrocyte count, mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), mean corpuscular hemoglobin concentration (MCHC), in normal pregnancy and after a normal delivery in non-iron-supplemented and iron supplemented women. METHODS: Two hundred and six healthy Danish women included at 9-18 weeks of gestation were allocated to treatment with placebo tablets (n=107) or tablets containing 66 mg iron (n=99). Blood samples were obtained at inclusion, every fourth week during gestation, and 8 weeks postpartum. RESULTS: All hematologic indices were significantly lower in placebo-treated than in iron treated women. In placebo-treated women, the 5th percentile for hemoglobin was 110 g/L in the 1st trimester; in the 2nd trimester it was 105 g/L in the first and the second, and 103 g/L in the last third; in the 3rd trimester, it was 102 g/L in the first, 100 g/L in the second, and 101 g/L in the last third; postpartum it was 113 g/L. In iron-treated women, the 5th percentile for hemoglobin was 111 g/L in the 1st trimester; in the 2nd trimester it was 109 g/L in the first, 106 g/L in the second, and 103 g/L in the last third; in the 3rd trimester, it was 105 g/L in the first and second, and 110 g/L in the last third; postpartum it was 123 g/L. CONCLUSIONS: Hematologic reference values should be derived from iron replete women. We suggest that the lowest critical hemoglobin value in iron-treated pregnant women should be 110 g/l (6.8 mmol/L) in the 1st trimester, and 105 g/L (6.5 mmol/L) in the 2nd and 3rd trimester. PMID- 10696956 TI - A hypothesis to explain the occurence of inner myometrial laceration causing massive postpartum hemorrhage. AB - BACKGROUND: Inner myometrial lacerations were found in three patients who developed uncontrollable postpartum massive bleeding despite the usual treatment for uterine atony. Because all the patients suffered from hemorrhage shock and their medical status deteriorated, their uteri were surgically removed to stop bleeding. After removal, one of them died. Postpartum hemorrhage was caused by inner myometrial laceration. We hypothesized a cause of inner myometrial laceration, using the three resected uteri, an assumed model of the uterine body, and 34 women. METHODS: The subjects were 37 women, of whom three were patients with inner myometrial laceration, 23 were women without inner myometrial laceration who underwent cesarean section, and 11 were women in the first stage of labor. The three resected uteri were examined both macroscopically and microscopically. We measured the thickness of the wall of the uterine muscle at the widest point of the uterine corpus and the thickness of the myometrial wall at a transverse section of the uterine cervix, as well as the radius of the inner lumen at the widest point of the uterus in 23 women during cesarean section. We also measured the thickness of the myometrial wall at the widest point of the uterine corpus in 11 women at the end of the first stage of labor during ultrasonic examination. The data were then used to estimate the stress on the uterine muscle. RESULTS: The stress on the uterine cervix was stronger than that on the uterine corpus during labor. When the stress on the uterine muscle is stronger than a specific value, inner myometrial lacerations develop on the right and/or left side of the uterine cervix. These lacerations may involve large vessels. CONCLUSIONS: We have discovered another cause of postpartum hemorrhage which we have named inner myometrial laceration. These lacerations appeared to result from a strong stress on the uterine cervix caused by an abnormal rise in intrauterine pressure during labor. PMID- 10696957 TI - Perinatal deaths in a Norwegian county 1986-96 classified by the Nordic-Baltic perinatal classification: geographical contrasts as a basis for quality assessment. AB - BACKGROUND: Quality assessment of perinatal care can be carried out by classifying perinatal deaths. In the following we have analyzed the geographical contrasts in perinatal deaths according to the Nordic-Baltic perinatal death classification in a sparsely populated Norwegian county. MATERIAL AND METHODS: All stillbirths (> or =28 weeks of gestation) and neonatal deaths (gestational age > or =22 weeks; death < or =28 days) in 1986-96 from Nordland county (240,000 inhabitants) were classified. For comparison the county was geographically divided into six general local hospital areas and one central hospital area. RESULTS: The classification showed a well acceptable inter and intra observer variation. One hundred and seventy-one stillbirths and 155 neonatal deaths were analyzed. The death rate (pr 1,000 births) for single, non-malformed, antenatal stillbirths was higher in the central hospital area than in the local hospital areas (3.22 vs. 2.02). The death rate for extreme preterm infants (22-27 weeks of gestation) was on the other hand higher in the local hospital areas (2.45 vs. 1.05). One of the general local hospital areas was singled out with an especially high neonatal death rate among extreme preterm infants. This was to some extent explained by the death of extreme preterm twins and triplets. CONCLUSION: The Nordic-Baltic perinatal death classification system is a consistent and reproducible tool also for studying perinatal death in restricted geographical areas. The observed contrasts in perinatal deaths were used as basis for programs aimed at improving perinatal care. The observation of an unexplained increased number of antenatal stillbirths in the central hospital area resulted in a program for prospective recording and better characterization of the placenta and umbilical cord. Proposals for a better antenatal program preventing extreme preterm birth of twins for the whole county has been launched. In utero transfer to a hospital with a neonatal intensive care unit seems crucial in improving the prognosis for these infants. PMID- 10696958 TI - The influence of stress and state anxiety on the outcome of IVF-treatment: psychological and endocrinological assessment of Swedish women entering IVF treatment. AB - BACKGROUND: Comparing stress levels in women entering IVF treatment with those of fertile controls as well as relating these levels to the outcome of IVF. METHODS: State anxiety and personality profiles as well as stress hormones were studied in 22 normally menstruating women entering IVF treatment for tubal infertility. Their personality profiles as well as state anxiety scores measured before entering IVF treatment were related to the outcome of treatment. Twenty-two fertile women served as controls. Stress markers were serum prolactin and cortisol. These were estimated by radioimmunoassay. The psychological evaluation included the Karolinska Scales of Personality (KSP) and state anxiety as measured by the STAI questionnaire. Basal FSH on cycle day 3 and E2 and P4 AUC during the luteal phase were evaluated as hormonal predictors for the outcome of IVF treatment. RESULTS: Comparison of the personality profiles of the two groups, showed that infertile women had significantly higher scores of suspicion (p>0.05), guilt (p>0.05), and hostility (p>0.01), but lower somatic anxiety (0.05) and indirect aggression (0.05) than fertile controls. The infertile women also had significantly higher levels of prolactin and cortisol throughout the menstrual cycle. Serum cortisol, prolactin and FSH levels on cycle day 3 did not differ between the women who conceived after IVF treatment and those who did not conceive. However, significant differences were found in E2 and P4 AUC (p>0.01) in the luteal phase between those women who became pregnant and those who failed. There was a trend (p<0.06) toward higher state anxiety levels among the women who did not succeed in becoming pregnant after IVF treatment. CONCLUSIONS: The main findings suggest that infertile women have a different personality profile in terms of more suspicion, guilt and hostility as compared to the fertile controls, perhaps as a response to their infertility. Their stress levels in terms of circulating prolactin and cortisol levels were elevated compared to the fertile controls. Psychological stress may affect the outcome of IVF treatment since state anxiety levels among those who did not achieve pregnancy were slightly higher than among those who became pregnant. PMID- 10696959 TI - In vitro fertilization in patients with ovarian endometriomas. AB - OBJECTIVE: The objective of the study was to establish whether operative treatment of recurrent ovarian endometriosis improves the prognosis of in vitro fertilization. METHODS AND MATERIAL: A retrospective analysis of one hundred endometriosis patients admitted to Tampere University Hospital for IVF treatment. Forty-five patients had an ovarian endometrioma during IVF treatment, 36 of the cases being recurrences after a previous operation. Fifty-five patients had ovarian endometriomas operated without recurrence. The patient groups with or without endometriosis did not differ in age, duration of infertility, sperm parameters, amount of gonadotropins required per oocyte and number of retrieved oocytes. RESULTS: The patients with ovarian endometriosis had more embryos (mean 3.9) than women without endometriomas (mean 2.8) (p<0.05) and the respective pregnancy rates per IVF cycle were 38% and 22%. Patients with endometriomas had a live birth rate of 27% compared with 20% in women with no endometriomas. CONCLUSIONS: The presence of a small endometrioma does not reduce the success of IVF treatment. PMID- 10696960 TI - The frequency of salpingitis and ectopic pregnancy as epidemiologic markers of Chlamydia trachomatis. AB - BACKGROUND: To study the incidence of non-gonococcal salpingitis, gonococcal salpingitis and ectopic pregnancy in a defined population over a 28-year period on the assumption that the frequency of salpingitis and ectopic pregnancy may indirectly illustrate the epidemiological pattern of Chlamydia trachomatis. DESIGN: A retrospective epidemiological study. SETTING: University hospital with an urban catchment area. PATIENTS: Five thousand two hundred and thirty-three patients admitted to the hospital between 1969 and 1996 with a diagnosis of ectopic pregnancy, non-gonococcal salpingitis, or gonococcal salpingitis. RESULTS: The frequencies of both non-gonococcal and gonococcal salpingitis increased steeply early in the period under study, rising to a peak in the early 1970s, then decreasing throughout the period except for the last 3 years when a slight increase was seen again. The frequency of ectopic pregnancy showed a steady increase, peaking in the late 1980s and early 1990s and then declining at the end of the study period. While the introduction of more sensitive pregnancy tests and programs for assisted fertility would increase the rate of ectopic frequency the decline during the 'nineties cannot be accounted for in this way. The peak of salpingitis cases in the early 'seventies seems to be mirrored exactly by the peak of ectopic pregnancies fifteen years later in the late 'eighties. CONCLUSION: The frequencies of salpingitis and of ectopic pregnancy can probably be used to estimate the incidence of preceding Chlamydia trachomatis. Thus the incidence of C. trachomatis has probably declined since the early 'seventies like that of N. gonorrheae. PMID- 10696961 TI - Perioperative morbidity of gynecological laparoscopy. A prospective monocenter observational study. AB - BACKGROUND: To study the morbidity rate of gynecological laparoscopy and to the most influential variables. METHODS: We conducted a prospective observational study from January 1st 1992 to December 31st 1998 in a single tertiary care center. It concerned patients who underwent gynecological laparoscopic surgery performed by seniors and residents. We have prospectively recorded patients characteristics, indications for laparoscopy, leading diagnosis, main operative procedures, post-operative course, surgical and anesthetic incidents and accidents. Complications were defined as any event that would modify the usual course of laparoscopy or of the post-operative period. RESULTS: One thousand and thirty-three laparoscopies were included. 80.1% of the procedures were major or advanced laparoscopies. The overall complication rate was 3%, with a laparotomy rate of 1.2%. About half of those complications (54.8%) occurred during the installation of laparoscopy. Veress needle and first trocar insertion accounted for 23.5% of those accidents (0.3% of the procedures) and suprapubic trocar insertion for 76.5%. Hemorrhages constituted almost all of the complications occurring during the operative stage (80%). The risk increased with the level of surgery and decreased with surgeon's experience. Prior abdominal surgery had no significant effect on the overall morbidity rate. Post-operative and anesthetic complications were rare. The overall complication rate as well as the laparotomy rate were stable all along the course of the study. CONCLUSIONS: Complication rate of gynecological laparoscopy is not negligible. Efforts should be made to lower the complications induced by the installation of laparoscopy, especially for secondary trocars. PMID- 10696962 TI - The long-term outcome of retropubic urethrocystopexy (sutures and fibrin sealant) and pubococcygeal repair. AB - BACKGROUND: To evaluate the results of retropubic urethrocystopexy (with sutures and fibrin sealant) and pubococcygeal repair five to seven years postoperatively. MATERIALS AND METHODS: Thirty women with genuine stress urinary incontinence were subjected to retropubic urethrocystopexy (n=30) and 15 women to pubococcygeal repair (n= 15). The preoperative assessment included both subjective and objective methods. The results evaluated three months, one year and five to seven years after the surgical treatment. RESULTS: One year after surgery 71% of the women in the urethrocystopexy group reported that they were continent, compared with only 43% five to seven years after surgery. In the pubococcygeal repair group 80% were continent at one-year follow-up, compared with 60% at the long term follow-up. According to pad test 79% of the women in the urethrocystopexy group had ceased leaking urine at minimal activity and 64% at maximal activity five to seven years after surgery. However, in the pubococcygeal repair group the corresponding percentage was 71% under both conditions. Intravesical pressure and Body Mass Index increased significantly in the whole group but urethra conductance and maximal urine flow decreased only in the urethrocystopexy group five to seven years after the surgical treatment. CONCLUSIONS: Accurate assessment of the results of any surgical treatment of stress urinary incontinence is difficult. During long term follow-up period significant changes may occur among the women, e.g. menopause and increase of Body Mass Index both predisposing to urinary incontinence. PMID- 10696963 TI - The role of pretreatment squamous cell carcinoma antigen in predicting nodal metastasis in early stage cervical cancer. AB - PURPOSE: To evaluate whether the presence of pelvic lymph node metastasis can be predicted by pretreatment squamous cell carcinoma antigen (SCC-Ag) levels in early stage squamous cervical carcinoma. MATERIALS AND METHODS: Between 1994 and 1998, 284 patients with stage Ib and IIa cervical squamous cell carcinoma undergoing radical hysterectomy had preoperative SCC-Ag determination. The correlation between clinicopathological findings on SCC-Ag levels were examined. The Mann-Whitney U test was used to statistically analyze differences between node positive and negative patients. Multiple regression analysis and a multiple logistic model were employed to examine the effect of clinicopathological findings on SCC-Ag levels. RESULTS: Of the 284 patients, 56 patients were found to have nodal metastasis. Median serum levels and 90% ranges of SCC-Ag were 0.74 microg/l (0.5-7.8) in the 228 nodal negative patients and 4.33 microg/l (0.5 48.5) in the 56 nodal positive patients (p<0.001). Lymph node metastasis and tumor size were found to have a significant impact on SCC-Ag levels. Around 86% of the patients with SCC-Ag levels below 8 microg/l showed no nodal metastasis, while about 65% of the patients with serum levels above 8 microg/l exhibited nodal metastasis. Multivariate analyses confirmed that only lymph node metastasis had a significant impact on the SCC-Ag levels exceeding 8 microg/l. CONCLUSION: For predicting nodal metastasis preoperatively, SCC-Ag levels greater than 8 microg/ l can be considered a high-risk zone for nodal metastasis. PMID- 10696964 TI - A prospective study of the discrete fascial defect rectocele repair. AB - BACKGROUND: The aim of this study is to describe the results of the discrete fascial defect rectocele repair with special emphasis on dyspareunia after the operation. METHODS: Sixty-seven women underwent rectocele repair from October 1997 to January 1999. Repair was limited to reapproximation of discrete defects in the rectovaginal fascia when possible and a small perineorrhaphy. Each patient was evaluated as to whether or not a discrete defect was present and the location of the fascial defect. Outcome measures were complications, recurrent prolapse after 3 months, changes in defecation problems and dyspareunia after the operation. RESULTS: In 96% (64/67) of the patients a discrete defect was present. Before the operation 40% (27/67) reported problems with evacuation of the rectum and 12% (8/67) dyspareunia or problems with intercourse because of the prolapse. The patients were evaluated 3 months after the operation. Only three patients still complained of evacuation problems and only two patients with prior dyspareunia still had problems, which were cured after a further 3 months. Two patients had de novo dyspareunia but in only one patient was an anatomical defect found. CONCLUSIONS: The discrete defect rectocele repair offers an anatomical correction of the rectocele, which alleviates the symptoms but, most important, does not give the woman dyspareunia. Long term results of the operation are awaited. PMID- 10696965 TI - Secondary infertility due to retained fetal bones--a diagnostic dilemma. PMID- 10696966 TI - Creation of a vagina by repeated coital dilatation in four teenagers with vaginal agenesis. PMID- 10696967 TI - Postpartum pneumopericardium. PMID- 10696968 TI - Primary psoas abscess during pregnancy. PMID- 10696969 TI - Unusual presentation of acute abdomen in a syndrome of double uterus, unilaterally imperforated double vagina, and ipsilateral renal agenesis. PMID- 10696970 TI - Diffuse malignant mesothelioma of the uterus. PMID- 10696971 TI - Defining the limits of public health. PMID- 10696972 TI - Evidence of gene-environment interaction in development of tuberculosis. PMID- 10696973 TI - Can growth hormone therapy cause diabetes? PMID- 10696974 TI - What happens when mutant neuroserpins get into bad shape. PMID- 10696975 TI - Serum aminotransferase concentration as evidence of hepatocellular damage. PMID- 10696976 TI - Prospects for lung-cancer screening. PMID- 10696977 TI - Patients' attitudes to psychiatric hospital admission. PMID- 10696978 TI - Population requirement for adult critical-care beds: a prospective quantitative and qualitative study. AB - BACKGROUND: The provision of adult critical-care facilities is not based on a rational or scientific assessment of need. We aimed to define the numbers of adult critical-care beds required for a population of 500000. METHODS: In five hospitals in Wales, UK, we classified patients who might be suitable for critical care in intensive-care or high-dependency units. On every 12th day for 1 calendar year, we counted the numbers of such patients admitted in a defined geographical population. A panel of ten intensivists made consensus decisions about whether individual patients were in the appropriate unit. The data were used to predict the numbers of beds and units required for the population. FINDINGS: 4058 patients were suitable for critical care, of whom 3028 lived in the study area. 56.4% were in general wards, 22.3% in high-dependency units, and 21.3% in intensive-care units. The mean risk of death was 22.0% and the in-hospital death rate 17.3%. According to the masked consensus, 41.3% of patients required high dependency beds and 21.5% intensive-care beds. Mean risk of death increased from general wards (14.7%) to high-dependency units (19.2%) to intensive care (37.0%). Based on the consensus decisions, the average daily requirement of intensive-care beds was 21 and of high-dependency beds 43; to meet needs 95% of times required 30 and 55 beds, respectively, in a single critical-care unit. INTERPRETATION: We estimated, scientifically, numbers of adult critical-care beds required to meet population needs. Studies are necessary periodically to track changes in admissions requiring critical care. PMID- 10696979 TI - Effect of recent thymic emigrants on progression of HIV-1 disease. AB - BACKGROUND: The concentration of T-cell receptor-rearrangement excision DNA circles (TREC) in peripheral-blood T cells is a marker of recent thymic emigrant alphabeta T cells. We studied the predictive ability of measurements of TREC for clinical outcome in HIV-1-infected individuals. METHODS: We measured TREC in peripheral-blood mononuclear cells with a real-time PCR assay. We studied 131 Greek participants in the Multicenter Hemophilia Cohort Study who had known HIV-1 seroconversion dates. The prognostic value of baseline TREC, CD4 T-cell count, and HIV-1 RNA concentration was assessed by Kaplan-Meier and Cox's regression analysis. FINDINGS: Four participants had progressed to AIDS by first blood sampling. Among the remaining 127 individuals, the median value of TREC per 10(6) cells was 6900 (IQR 2370-15604). Baseline TREC values were lower in the 53 who progressed to AIDS than in those who did not (geometric mean 2843 [95% CI 1468 5504] vs 6560 [4723-9113] per 10(6) cells; p=0.017). The relative hazard of AIDS, adjusted for plasma viral load, CD4 T-cell count, and age at seroconversion was 1.44 (95% CI 1.04-2.01; p=0.031) per ten-fold increase in TREC; that for death was 1.52 (1.12-2.06; p=0.007). The adjusted relative hazards of death were 2.91 (1.91-4.44; p<0.001) per ten-fold increase in plasma HIV-1 RNA load and 1.20 (1.04-1.38; p=0.014) per 100-cell decrease in CD4 T-cell count. INTERPRETATION: The concentration of TREC in the peripheral T-cell pool complements HIV-1 RNA load and CD4 T-cell count in predicting the rate of HIV-1 disease progression. Recent thymic emigrants have a role in the pathogenesis of HIV-1 disease. PMID- 10696980 TI - Interferon alfa-2b, colchicine, and benzathine penicillin versus colchicine and benzathine penicillin in Behcet's disease: a randomised trial. AB - BACKGROUND: Sight-threatening eye involvement is a serious complication of Behcet's disease. Extraocular complications such as arthritis, vascular occlusive disorders, mucocutaneous lesions, and central-nervous-system disease may lead to morbidity and even death. We designed a prospective study in newly diagnosed patients without previous eye disease to assess whether prevention of eye involvement and extraocular manifestations, and preservation of visual acuity are possible with combination treatments with and without interferon alfa-2b. METHODS: Patients were randomly assigned 3 million units interferon alfa-2b subcutaneously every other day for the first 6 months plus 1.5 mg colchicine orally daily and 1.2 million units benzathine penicillin intramuscularly every 3 weeks (n=67), or colchicine and benzathine penicillin alone (n=68). The primary endpoint was visual-acuity loss. Analysis was by intention to treat. FINDINGS: Significantly fewer patients who were treated with interferon had eye involvement than did patients who did not receive interferon (eight vs 27, relative risk 0.21 [95% CI 0.09-0.50], p<0.001). Ocular attack rate was 0.2 (SD 0.62) per year with interferon therapy and 1.02 (1.13) without interferon therapy (p=0.0001). Visual acuity loss was significantly lower among patients treated with interferon than in those without interferon (two vs 13, relative risk 0.13 [95% CI 0.03-0.60], p=0.003). Arthritis episodes, vascular events, and mucocutaneous lesions were also less frequent in patients treated with interferon than in those not receiving interferon. No serious side-effects were reported. INTERPRETATION: Therapy with interferon alfa-2b, colchicine, and benzathine penicillin seems to be an effective regimen in Behcet's disease for the prevention of recurrent eye attacks and extraocular complications, and for the protection of vision. PMID- 10696981 TI - Incidence of diabetes mellitus and impaired glucose tolerance in children and adolescents receiving growth-hormone treatment. AB - BACKGROUND: Growth hormone (GH) contributes to insulin resistance, but whether children treated with GH are at increased risk of diabetes has not been established. We undertook a retrospective analysis of data from an international pharmacoepidemiological survey of children treated with GH to find out the incidence of impaired glucose tolerance and types 1 and 2 diabetes mellitus. METHODS: Reports to the survey of abnormal glucose metabolism were investigated and classified. The incidence and age-distribution of type 1 diabetes were compared with values from a model of reference data. The incidence of type 2 diabetes was compared with data from two reports of children not treated with GH. FINDINGS: 85 (0.36%) of 23333 children were reported with abnormal glucose metabolism. After investigation, 43 had confirmed glucose disorders (11 with type 1 diabetes, 18 with type 2 diabetes, and 14 with impaired glucose tolerance). The incidence and age at diagnosis of type 1 diabetes in children treated with GH did not differ from expected values. The incidence of type 2 diabetes was 34.4 cases per 100000 years of GH treatment which was six-fold higher than reported in children not treated with GH. Type 2 diabetes did not resolve after GH therapy was stopped. INTERPRETATION: GH treatment did not affect the incidence of type 1 diabetes mellitus in any age group. We postulate that the higher than expected incidence of type 2 diabetes mellitus with GH treatment may be an acceleration of the disorder in predisposed individuals. PMID- 10696982 TI - Community care and criminal offending in schizophrenia. AB - BACKGROUND: The introduction of community care in psychiatry is widely thought to have resulted in more offending among the seriously mentally ill. This view affects public policy towards and public perceptions of such people. We investigated the association between the introduction of community care and the pattern of offending in patients with schizophrenia in Victoria, Australia. METHODS: We established patterns of offending from criminal records in two groups of patients with schizophrenia over their lifetime to date and in the 10 years after their first hospital admission. One group was first admitted in 1975 before major deinstitutionalisation in Victoria, the second group in 1985 when community care was becoming the norm. Each patient was matched to a control, by age, sex, and place of residence to allow for changing patterns of offending over time in the wider community. FINDINGS: Compared with controls, significantly more of those with schizophrenia were convicted at least once for all categories of criminal offending except sexual offences (relative risk of offending in 1975=3.5 [95% CI 2.0-5.5), p=0.001, in 1985=3.0 [1.9-4.9], p=0.001). Among men, more offences were committed in the 1985 group than the 1975 group, but this was matched by a similar increase in convictions among the community controls. Those with schizophrenia who had also received treatment for substance abuse accounted for a disproportionate amount of offending. Analysis of admission data for the patients and the total population of admissions with schizophrenia showed that although there had been an increase of 74 days per annum spent in the community for each of the study population as a whole, first admissions spent only 1 more day in the community in 1985 compared with 1975. INTERPRETATION: Increased rates in criminal conviction for those with schizophrenia over the last 20 years are consistent with change in the pattern of offending in the general community. Deinstitutionalisation does not adequately explain such change. Mental-health services should aim to reduce the raised rates of criminal offending associated with schizophrenia, but turning the clock back on community care is unlikely to contribute towards any positive outcome. PMID- 10696983 TI - Influence of vitamin D deficiency and vitamin D receptor polymorphisms on tuberculosis among Gujarati Asians in west London: a case-control study. AB - BACKGROUND: Susceptibility to disease after infection by Mycobacterium tuberculosis is influenced by environmental and host genetic factors. Vitamin D metabolism leads to activation of macrophages and restricts the intracellular growth of M. tuberculosis. This effect may be influenced by polymorphisms at three sites in the vitamin D receptor (VDR) gene. We investigated the interaction between serum vitamin D (25-hydroxycholecalciferol) concentrations and VDR genotype on susceptibility to tuberculosis. METHODS: This study was a hospital based case-control analysis of Asians of Gujarati origin, a mainly vegetarian immigrant population with a high rate of tuberculosis. We typed three VDR polymorphisms (defined by the presence of restriction endonuclease sites for Taq1, Bsm1, and Fok1) in 91 of 126 untreated patients with tuberculosis and 116 healthy contacts who had been sensitised to tuberculosis. Serum 25 hydroxycholecalciferol was recorded in 42 contacts and 103 patients. FINDINGS: 25 hydroxycholecalciferol deficiency was associated with active tuberculosis (odds ratio 2.9 [95% CI 1.3-6.5], p=0.008), and undetectable serum 25 hydroxycholecalciferol (<7 nmol/L) carried a higher risk of tuberculosis (9.9 [1.3-76.2], p=0.009). Although there was no significant independent association between VDR genotype and tuberculosis, the combination of genotype TT/Tt and 25 hydroxycholecalciferol deficiency was associated with disease (2.8 [1.2-6.5]) and the presence of genotype ff or undetectable serum 25-hydroxycholecalciferol was strongly associated with disease (5.1 [1.4-18.4]). INTERPRETATION: 25 hydroxycholecalciferol deficiency may contribute to the high occurrence of tuberculosis in this population. Polymorphisms in the VDR gene also contribute to susceptibility when considered in combination with 25-hydroxycholecalciferol deficiency. PMID- 10696984 TI - Transient ischaemic attack in a young woman. PMID- 10696985 TI - Rise in gonorrhoea in London, UK. London Gonococcal Working Group. AB - We report an alarming 34.9% overall increase in the number of gonococcal infections diagnosed during the same 3 month study period during 3 years. Between 1997 and 1998, an increase of 12.1% was detected, with a larger increase between 1998 and 1999, of 20.4%. PMID- 10696986 TI - Rapid loss of superoxide dismutase activity during antigen-induced asthmatic response. AB - Loss of superoxide dismutase activity occurs within minutes of an acute asthmatic response to segmental antigen instillation into the lung of individuals with atopic asthma. Decreased activity undoubtedly contributes to airway inflammation and injury through increased formation of reactive oxygen and nitrogen species, and suggests that enrichment of lung antioxidants is therapeutic for asthma. PMID- 10696987 TI - Primary chemotherapy and conservative surgery for vaginal yolk-sac tumour. Maligne Keimzelltumoren Study Group. AB - Vaginal yolk-sec tumours are usually incurable unless radical surgery is done. We have shown, however, that, neoadjuvant cisplatin-based chemotherapy with conservative surgery is effective in the management of these tumours, and results in a good survival rate, unlike germ-cell tumours of other origin. PMID- 10696988 TI - Molecular classification of the dementias. AB - Most patients with frontotemporal dementia do not have taupathology as shown by immunohistochemistry. The use of the term tauopathy to classify frontotemporal dementia is inappropriate. PMID- 10696989 TI - Extensive vitiligo after ganciclovir treatment of GvHD in a patient who had received donor T cells expressing herpes simplex virus thymidine kinase. AB - The use of donor T cells expressing a suicide gene for destruction of graft versus-host effector cells provides a tool for alloreactivity modulation. We describe a case of extensive vitiligo that developed after ganciclovir treatment of cutaneous chronic graft-versus-host disease in a patient who had received donor T lymphocytes expressing herpes simplex virus thymidine kinase at the time of transplantation. PMID- 10696991 TI - Internet addiction: genuine diagnosis or not? PMID- 10696990 TI - Exposure to airborne dust contaminated with pesticide in the Aral Sea region. AB - The Aral Sea region is one of the world's foremost ecological disaster zones and there is increasing local concern for the health of millions of people living in this region. We have found that dust deposition rates across eastern Turkmenistan are among the highest in the world and that the dust is contaminated with pesticide. PMID- 10696992 TI - Hong Kong debates where to draw the line with passive euthanasia. PMID- 10696993 TI - HIV-contamination scare adds to US gene researchers' woes. PMID- 10696994 TI - Irish media revelations prompt revised post-mortem guidelines. PMID- 10696995 TI - UK government starts to lose patience with General Medical Council. PMID- 10696996 TI - Angiotensin II receptor antagonists. AB - Blockade of the renin-angiotensin system began as a way of studying the pathogenesis of cardiovascular disease with specific pharmacological probes. Oral activity, achieved by shortening the original peptide structures, transformed the probes into therapeutic agents, the angiotensin-converting enzyme (ACE) inhibitors. However, ACE is a non-specific target for blocking the renin angiotensin enzymatic cascade. The availability of orally active drugs turned ACE inhibition into a therapeutic breakthrough but more specific blockade always seemed desirable. This goal has now been achieved with the orally active angiotensin II receptor antagonists; six are on the market and more are under development. This new class of drugs is equal in efficacy to ACE inhibitors, at least in hypertensive patients. Trials now underway will demonstrate whether angiotensin II receptor antagonists can prevent target-organ damage and reduce cardiovascular morbidity and mortality. If they do, these compounds might one day replace ACE inhibitors. PMID- 10696997 TI - COX-1, COX-2, and COX-3 and the future treatment of chronic inflammatory disease. AB - A new generation of non-steroidal anti-inflammatory drugs has been described that selectively targets the inducible isoform of cyclo-oxygenase, cyclo-oxygenase 2 (COX-2). This isoform is expressed at sites of inflammation, which has led to the speculation that its inhibition could provide all the benefits of current nonsteroidal anti-inflammatory drugs, but without their major side-effects on the gastrointestinal system (which are due to inhibition of COX-1). We have shown that COX-2 (identified by use of specific antibodies) is induced during the resolution of an inflammatory response, inhibition of COX-2 resulting in persistence of the inflammation due to the prevention of the synthesis of a range of anti-inflammatory prostanoids. We propose that there is a third isoform of this enzyme family, COX-3, a proposal that will have implication for the prescription of both existing and new generation anti-inflammatory drugs, and might represent a new therapeutic target. PMID- 10696998 TI - A matter of choice. PMID- 10696999 TI - What does STOP-2 tell us about management of hypertension? PMID- 10697000 TI - What does STOP-2 tell us about management of hypertension? PMID- 10697001 TI - What does STOP-2 tell us about management of hypertension? PMID- 10697002 TI - What does STOP-2 tell us about management of hypertension? PMID- 10697003 TI - What does STOP-2 tell us about management of hypertension? PMID- 10697004 TI - What does STOP-2 tell us about management of hypertension? INSIGHT Steering Committee. PMID- 10697005 TI - Contrast agent confusion. PMID- 10697006 TI - Selection of metastatic tumour phenotypes. PMID- 10697007 TI - Oesophageal involvement in pemphigus vulgaris. PMID- 10697008 TI - Bornavirus isolates of human origin. PMID- 10697009 TI - Salvage of critically ischaemic limbs. PMID- 10697010 TI - Salvage of critically ischaemic limbs. PMID- 10697011 TI - Ursodeoxycholic acid for primary biliary cirrhosis. PMID- 10697012 TI - Promiscuity and the abuse of stigma. PMID- 10697013 TI - Promiscuity and the abuse of stigma. PMID- 10697014 TI - A neonatal hypothermia indicator. PMID- 10697015 TI - Peer review. PMID- 10697016 TI - Palliative medicine and modern cancer care. PMID- 10697017 TI - Organization of services and nursing care: hospice and palliative medicine. AB - The health care industry is changing and nursing case management is an integral part of restructured care in many institutions. Health care organizations must evaluate services and outcomes. The terminally ill comprise a large portion of patients in any health care delivery system. Hospitals that provide formal cancer care services need to evaluate where palliative care and hospice fit. Shifts in patient care will be evident due to changes in demographics, payor initiatives, and technological advances. Providing care for patients with advanced disease and the role of nursing have evolved over the past 10 years. One important area that has not changed is the passion and caring evident in the nurse's everyday practice. PMID- 10697018 TI - Clinical evaluation in advanced cancer. AB - This chapter will outline a general approach to symptom assessment, using the interdisciplinary approach to pain as a model. Due to the implications of cognitive impairment for treatment compliance, consent, and caregiver burden, assessment of cognitive function will be reviewed in detail. Problem areas in assessment are identified, along with aids to improve assessment and emphasize the key contribution of the nurse. We will also focus on measurement issues from both clinical and research perspectives. PMID- 10697019 TI - Common symptoms in advanced cancer. AB - The relief of physical and psychological symptoms is an essential part of palliative care. Advanced cancer is an acute process; because the clinical picture changes rapidly, symptoms must be reassessed regularly, and a careful history is essential. Defining the relationship of the symptoms to the disease can defuse fear and encourage a sense of control in patients and their families. We review the pathophysiology, causes, prevalence, consequence, and management of common symptoms in advanced cancer. PMID- 10697020 TI - Common complications of advanced cancer. AB - Complications due to cancer and its treatment are common and increase in incidence and severity as the disease progresses. Central nervous system complications affect 15% to 20% of patients, and up to 75% have bone metastases at some point during the disease process. Endocrine abnormalities include hypercalcemia, adrenal insufficiency, and inappropriate antidiuretic syndrome. Hematologic disorders, malignant effusions, and gastrointestinal (GI) problems may cause significant morbidity. PMID- 10697021 TI - Pharmacological management of cancer pain. AB - Given modern techniques of pain assessment and management, it is now possible to be optimistic about cancer pain control. Assessment of cancer pain must include information about the site(s) of pain, pathophysiology, pain severity, and quantification of analgesic responses. Correct diagnosis of common pain patterns including breakthrough and incident pain are essential. The principles of analgesic use are well defined. The concept of rescue dosing in safe analgesic titration and management of breakthrough/incident pain is a key concept. Individualization of opioid dosing is important and this is facilitated by a number of dosing strategies. Choice of specific opioid is often less important than correct dosing, as side effects are similar among the commonly prescribed drugs. Anticipations and management of common side effects improve the therapeutic index. Alternate routes of administration are important, usually because of loss of the oral route of administration. Misunderstandings about opioids are common and patient and family education paramount. Adjuvant analgesics are necessary for good pain control, but have important differences in indications, usage, and side effects compared with opioids. First-rate pain management is a basic professional and humanitarian responsibility of the skilled clinical oncologist. PMID- 10697022 TI - The cancer anorexia-cachexia syndrome. AB - The cancer anorexia-cachexia syndrome is one of the most common causes of death among cancer patients and is present in 80% at death. It is a complex example of metabolic chaos effecting protein, carbohydrate, and fat metabolism. Tumors produce both direct and indirect abnormalities, resulting in anorexia and weight loss. The disease burden does not correlate with the degree of cachexia. Although there is no treatment to control or reverse the process, appetite stimulants may promote increased food intake and enhance quality of life. PMID- 10697023 TI - Symptom control in advanced cancer: important drugs and routes of administration. AB - Aggressive symptom control is a vital component of palliative medicine. Frequently both physicians and patients focus on pain control, forgetting the broader issues of symptom control. Pain and other symptoms are inextricably linked. Common symptoms include constipation, nausea and vomiting, insomnia, anorexia, weight loss, and cough. All oncologists should be familiar with the indications, doses, and unwanted effects of drugs commonly indicated for symptom control. This article will discuss some drugs presently available to achieve good symptom control. At the correct dose and dosing schedule, these agents can have a significant impact on quality of life. As in all areas of medicine, it is best to know the benefits and unwanted effects of a few drugs, rather than randomly prescribing different agents for similar clinical situations. This is rational prescribing. While the list presented here is not exhaustive, it does reflect core drugs currently available in the United States. PMID- 10697024 TI - The dying cancer patient. AB - Cancer patients often die with serious unrelieved symptoms causing a distressing death for them and needless added suffering for their families. Many physicians have not been trained to care for the dying patient. This chapter reviews the common symptoms and describes the methods to control them and support the patient and family through this difficult time. These symptoms are so characteristic of the dying process that all physicians should recognize them, be skilled in providing appropriate care, and prepare for problems that may arise. PMID- 10697026 TI - Bioethics: communication and decision-making in advanced disease. AB - Effective communication is the cornerstone of excellence in patient care, and breakdown in communication is a common problem leading to requests for bioethics consultation. In palliative medicine, issues involving end-of-life decisions inherently involve many potential ethical concerns. Ensuring good communication among the physician, health care professionals, patient, and family will facilitate care, and avoid ethical problems. This chapter will address communication with patients, families, and health care professionals. Common ethical concerns such as withholding and withdrawing treatment modalities, medical futility, euthanasia, and assisted suicide will be discussed. PMID- 10697025 TI - Palliative radiotherapy. AB - Radiotherapy is often used for the palliative treatment of advanced cancer. Although each cancer is unique in its histology and natural history, there are several scenarios that repeatedly challenge physicians. Specifically, skeletal metastases, spinal cord compression, brain metastasis, bronchial obstruction, and vena cava obstruction are regularly encountered. This review will discuss the diagnosis and treatment of these common important situations. PMID- 10697027 TI - Psychosocial care in advanced cancer. AB - In palliative care, the focus is management of major symptoms and complications, and psychosocial support of the patient and family. Approaching the end of life, the patient's needs move beyond physical care to include the psychological, social, and spiritual dimensions. The main psychosocial interventions are counseling, education, and practical services directed at the needs identified by the multidimensional/multidisciplinary assessments. We will present the roles of the various team members and methods of psychosocial assessment. PMID- 10697028 TI - Gemcitabine-containing regimens in non-small cell lung cancer: are these standard therapies or not? PMID- 10697029 TI - Combination chemotherapy with paclitaxel plus carboplatin versus paclitaxel plus gemcitabine in inoperable non-small cell lung cancer: a phase III randomized study. Preliminary results. Hellenic Cooperative Oncology Group. AB - Gemcitabine plus paclitaxel and paclitaxel plus carboplatin are active and well tolerated in patients with advanced non-small cell lung cancer, showing similar rates of response and survival. The Hellenic Cooperative Oncology Group conducted a randomized phase III trial comparing gemcitabine plus paclitaxel with paclitaxel plus carboplatin. Patients were randomly assigned to two groups. Group A received paclitaxel 200 mg/m2 plus carboplatin (area under the curve = 6) on day I. Group B received paclitaxel in identical fashion to group A plus gemcitabine 1,000 mg/m2 on days I and 8 every 3 weeks. A minimum of two cycles and a maximum of six cycles was allowed. To date, 127 eligible patients (63 in group A and 64 in group B) have been randomized; the median follow-up time is 4.6 months. Preliminary results suggest that both combinations can be given in full doses and are well tolerated. Grade 3/4 neutropenia was mild but more prominent in group A (10% v 3%, respectively) while thrombocytopenia was not significant for either group. Moreover, severe neurotoxicity, hepatotoxicity, or cardiac toxicity has not been observed in the vast majority of patients in either group. Although patients in group B experienced higher response rates (37.5%) than those in group A (21.8%), the difference between the groups was not statistically significant. Definite conclusions about this study cannot be made until more data are available. PMID- 10697030 TI - Phase II trial evaluating triplet chemotherapy using gemcitabine, paclitaxel, and carboplatin in the treatment of patients with advanced non-small cell lung cancer. AB - The purpose of this study was to evaluate the combination of gemcitabine, paclitaxel, and carboplatin in patients with advanced non-small cell lung cancer. Previously untreated patients with stage IIIB or IV non-small cell lung cancer were enrolled into this trial. Sixty-nine patients from the phase II portion and eight patients from the phase I portion were treated with gemcitabine 1,000 mg/m2 intravenously on days I and 8, paclitaxel 200 mg/m2 as a 1-hour infusion on day 1, and carboplatin at an area under the curve of 5.0 intravenously on day 1. Treatment courses were repeated every 21 days. The phase II component of the study was completed at 13 community-based practices in the Minnie Pearl Cancer Research Network. Thirty-four of 71 fully evaluable patients had an objective response (48%, two complete and 32 partial responses). Twenty-five patients (35%) were stable and 12 (17%) progressed. The median response duration was 6 months (range, 3 to 14 months) and the median survival was 9.9 months, with 1- and 2 year survival rates of 47% and 21%, respectively. The combination of gemcitabine, paclitaxel, and carboplatin has been shown to be safe and effective; thus, this three-drug regimen will be compared with a standard two-drug regimen, paclitaxel/carboplatin, in a phase III study. PMID- 10697031 TI - The gemcitabine/epirubicin/paclitaxel combination in advanced breast cancer. AB - The purpose of this study was to determine the response rate of the gemcitabine/epirubicin/paclitaxel combination (GET) and its feasibility as induction chemotherapy before high-dose consolidation treatment in patients with metastatic breast cancer. Patients received gemcitabine 1,000 mg/m2 on days I and 4, epirubicin 90 mg/m2 on day 1, and paclitaxel 175 mg/m2 on day I every 3 weeks for up to eight courses. After six courses of GET, responding patients or those with stable disease entered a high-dose chemotherapy program. All 36 enrolled patients were evaluated for toxicity and response. The GET combination was well tolerated, with myelosuppression the being most common toxicity; grade 4 neutropenia was reported in 56% of patients. The overall response rate was 89% (95% confidence interval, 73.4% to 96.9%), with a 28% complete response rate. The high-dose chemotherapy program resulted in a response rate of 92% and a complete response rate of 44%. As a result of the promising activity demonstrated in this phase II study with GET and following high-dose chemotherapy, three related studies are planned: an in vitro study evaluating the possible synergism of paclitaxel and gemcitabine, a phase III study comparing GET with epirubicin/paclitaxel in metastatic breast cancer, and a phase II trial evaluating GET in patients with operable breast cancer. PMID- 10697032 TI - Paclitaxel/gemcitabine administered every two weeks in advanced breast cancer: preliminary results of a phase II trial. AB - The purpose of this study was to evaluate the toxicity and efficacy of paclitaxel/gemcitabine administered every 2 weeks in the first-line treatment of advanced breast cancer. Forty-three chemonaive patients with histologically confirmed metastatic breast carcinoma were enrolled. Patients received paclitaxel 150 mg/m2 followed by gemcitabine 2,500 mg/m2, both on day I of a 14-day cycle, for a maximum of eight cycles. Thirty-four patients were evaluable for toxicity; 38 were evaluable for efficacy. The median age at the time of diagnosis was 54 years, the median performance status was 90, and the median number of lesions was three. Most patients (71%) had received prior adjuvant therapy. Grade 3 and 4 toxicity was limited to leukocytes (14% and 18%, respectively). Grade 3 toxicities (5% each) were thrombocytopenia, nausea and vomiting, elevation of aspartate transaminase, neurosensory, and constipation. One patient had neutropenia and fever. The objective response rate was 68% (21% complete response and 47% partial response); 18% had stable disease and 13% had partial disease. The preliminary evaluation of paclitaxel/gemcitabine given as a 2-week schedule to patients with untreated advanced breast carcinoma shows encouraging activity and a favorable toxicity profile. PMID- 10697033 TI - Phase I study of the gemcitabine/oxaliplatin combination in patients with advanced solid tumors: a preliminary report. AB - The purpose of this study was to determine the maximum tolerated dose and evaluate the dose-limiting toxicities of the gemcitabine/oxaliplatin combination in patients with advanced-stage solid tumors. Of the 48 enrolled patients, 35% had first-line, 23% had second-line, and 42% had third-line therapy. Patients received escalating doses of 1,000 to 1,600 mg/m2 gemcitabine on days I and 8 plus 60 to 120 mg/m2 oxaliplatin on day 8, every 21 days. The dose-limiting toxicities (first nine dose levels) were grade 4 neutropenia, grade 3 asthenia, and grade I to 3 neutropenia or thrombocytopenia. One hundred fifty-three cycles have been administered without any febrile neutropenia or toxic death. Grade 3-4 neutropenia and grade 3 thrombocytopenia were noted in 13 (9%) and seven (5%) cycles, respectively. Nonhematologic toxicity has been mild; the most common was grade 2 to 3 asthenia occurring in 29% of cycles. Among 30 evaluable patients, four partial responses have been observed (response rate, 13%). In this phase I study, the maximum tolerated dose of this drug combination has not yet been reached, although gemcitabine up to 1,600 mg/m2 (days I and 8) plus oxaliplatin up to 120 mg/m2 (day 8) was active and well tolerated, warranting further evaluation in phase II studies. PMID- 10697034 TI - Gemcitabine in bladder cancer. AB - Systemic chemotherapy is the only current modality that provides the potential for long-term survival in patients with advanced or metastatic transitional cell carcinoma of the urothelium. The methotrexate/vinblastine/doxorubicin/cisplatin combination is currently considered to be the standard treatment of this disease, with overall response rates of up to 72% and survival durations of approximately I year. With the addition of gemcitabine to the chemotherapeutic arsenal, approaches to bladder cancer are changing significantly. When administered as a single agent, gemcitabine produces response rates of 23% to 28% in both pretreated and chemonaive patients with an excellent toxicity profile. When combined with cisplatin, overall response rates increase to 66% with maintained tolerability. Other new gemcitabine combinations with agents such as taxanes or carboplatin also appear promising; however, these preliminary phase II data must be confirmed in the phase III setting. Effective management of transitional cell carcinoma also involves understanding the mechanisms of resistance to treatment, including the implications of the expression of p-glycoprotein, p53 proteins, and several other biochemical predictors of outcome, as well as overcoming resistance. Growth factors may enable us to maximize drug delivery. PMID- 10697035 TI - The role of gemcitabine in the treatment of ovarian cancer. AB - Epithelial ovarian cancer remains the number one gynecologic killer in the Western world. The standard therapeutic approach for patients with advanced-stage epithelial ovarian cancer has been cytoreductive surgery followed by combination chemotherapy. Despite improvements in outcome associated with carboplatin/ paclitaxel-based chemotherapy, most patients with advanced ovarian cancer are not cured by this combination. Gemcitabine has been shown to be an active agent in the treatment of patients with recurrent ovarian cancer. The response rate of approximately 19% noted with single-agent gemcitabine is associated with acceptable toxicity. In addition, gemcitabine can be combined with other active agents, including carboplatin and paclitaxel. Clinical trials will be performed to evaluate whether a three-drug combination including gemcitabine will be superior to treatment with the standard approach of carboplatin plus paclitaxel. PMID- 10697036 TI - Ifosfamide: actual data, new insights, and persisting questions. PMID- 10697037 TI - Ifosfamide and irinotecan in solid tumors: a phase I dose-finding trial. AB - The combination of ifosfamide and irinotecan was tested in a dose-finding study. The preliminary results of the combination in this phase I study did not show any major toxicity that could help to define the dose-limiting toxicity. The escalation continues even after nine levels; the study is therefore ongoing. The main toxicity is gastrointestinal, with mild nausea, vomiting, and diarrhea. There was some other irrelevant toxicity, which was easily manageable with the usual supportive therapy. When responses were evaluated, stable disease was found in some cases, suggesting some activity for the combination. PMID- 10697038 TI - Ifosfamide-based drug combinations: preclinical evaluation of drug interactions and translation into the clinic. AB - Ifosfamide is an alkylating antineoplastic agent with documented activity against a variety of solid tumor types, most notably lung cancer, testicular cancer, and sarcoma. Ifosfamide has been included in various drug combination protocols, usually on an empirical basis. To gather more insight into the mechanisms that underlie these drug interactions and to develop guidelines for further improvement of clinical combination protocols, we performed a broad preclinical evaluation program of ifosfamide-based combination regimens using isobologram analysis of drug interactions. In established cisplatin-sensitive and cisplatin refractory ovarian carcinoma cell lines, a schedule-dependent drug interaction between paclitaxel and activated hydroperoxy-ifosfamide (4-OOH-IF) could be demonstrated. When both drugs were given for 2 hours, simultaneous exposure or the sequence of paclitaxel followed by 4-OOH-IF were additive or synergistic. In contrast, application of 4-OOH-IF before paclitaxel resulted in pronounced antagonism. Based on the sequence-dependent synergistic interactions a phase I trial was initiated with paclitaxel given on day 1 and ifosfamide given on days 2 to 5 in patients with cisplatin-refractory ovarian cancer. Four dose levels were evaluated in 18 patients. The maximum tolerated dose was paclitaxel 175 mg/m2 on day 1 and ifosfamide 2,000 mg/m2 on days 2 to 5, with central nervous system toxicity and nephrotoxicity being dose-limiting. The recommended dose for further evaluation of this combination was paclitaxel 175 mg/m2 on day 1 and ifosfamide 1,500 mg/m2 on days 2 to 5. Although all patients were heavily pretreated with multiple agents, nine of 18 patients achieved an objective response. Ifosfamide also has been shown to reduce cellular glutathione content; thus, a series of experiments evaluated the ability of activated cyclophosphamide or ifosfamide to deplete cellular glutathione in vitro. It was demonstrated that glutathione depletion is dose- and time-dependent, with 4-OOH-IF leading to a more pronounced suppression of cellular glutathione compared with 4-OOH-Cy. The decrease in cellular glutathione content was maximal at 2 hours after drug treatment; however, cellular glutathione levels returned to normal within 24 hours. When 4 OOH-IF was combined in vitro with cisplatin, schedule-dependent interactions again became obvious. The highest antitumor activity was seen when both drugs were given concurrently; sequential application with 4-OOH-IF given before cisplatin resulted in antagonism. Since adequate glutathione levels are necessary for multidrug resistance protein (MRP) function, glutathione depletion might lead to reversal of MRP-mediated drug resistance. Preliminary data showed that 4-OOH IF significantly decreases glutathione concentrations in MRP-expressing human HT1080/DR4 sarcoma cells, leading to maximum steady-state reduction after a 90 min exposure to 4-OOH-IF. Taken together the data reported here demonstrate that in vitro ifosfamide may potentiate the antitumor activity of a variety of cytotoxic agents and therefore merits further clinical evaluation in drug combinations (eg, taxanes, anthracyclines). PMID- 10697039 TI - Ifosfamide- and paclitaxel-based treatment of relapsed and refractory lymphoma. AB - Several trials of new salvage therapies for relapsed and refractory non-Hodgkin's lymphoma are based on ifosfamide and paclitaxel. These programs variably Induce clinical remissions and prepare patients for stem cell and bone marrow transplantations. Ifosfamide-containing regimens are also being evaluated in the treatment of newly diagnosed patients. However, no data exist with regard to paclitaxel-containing regimens outside salvage settings. This report reviews the effects of these compounds in the treatment of relapsed and refractory non Hodgkin's lymphoma and proposes likely directions for future study. PMID- 10697040 TI - Chemotherapy with paclitaxel, ifosfamide, and cisplatin for the treatment of squamous cell cervical cancer: the experience of Monza. AB - The medical treatment of squamous cell cervical carcinoma is receiving increasing attention. Cisplatin and ifosfamide are known effective drugs. Paclitaxel has been tested with interesting results in cervical cancer. We evaluated the toxic effects and the antitumor activity of a multiagent regimen that included paclitaxel, ifosfamide, and cisplatin (TIP) in two different settings: bulky and locally advanced cervical cancer and recurrent-persistent disease. Treatment consisted of paclitaxel 175 mg/m2 given over 3 hours on day 1, cisplatin 50 mg/m2 (75 mg/m2 in 24 patients), ifosfamide 5 g/m2 and mesna 5 g/m2 given on day 2, and mesna 3 g/m2 given on day 3. In the neoadjuvant setting, the course was repeated every 3 weeks for three courses. Unless there was progression of disease or reason to avoid surgery, all patients were scheduled for radical hysterectomy and pelvic lymphadenectomy. Thirty-eight patients with locally advanced cervical cancer were studied: 11 women achieved a clinical complete response, 21 had a partial response, five had stable disease, and one had disease progression. Among the 34 patients who underwent surgery, six had a pathologically documented complete response, seven had an optimal partial response (only microscopic residual disease), 19 had a suboptimal partial response, and two stable diseases. Grade 3-4 neutropenia was recorded in 71% of patients, grade 3-4 thrombocytopenia in 10.5%, and grade 2 peripheral neuropathy in 2.6%. With a median follow-up period of 22 months for the patients who remain alive, 28 women are alive without recurrence and five are alive with persistent/recurrent disease. Five patients have died of disease. In the salvage setting, 45 women with persistent-recurrent disease after primary treatment were treated; 31 of these women had received prior radiation. In the salvage setting we observed 15 clinical complete responses, 15 partial responses, nine stable diseases, and six disease progressions. The objective response rate was 66.6%. Ten complete responders underwent subsequent surgery and seven had pathologic complete response (two in radiated areas). The response rate was 52% in radiated areas and 75% in nonradiated areas. The median survival time is 6 months for the nonresponders, 9+ month for the partial responders, and 13+ months for the complete responders. The most relevant side effect was myelotoxicity, with 91% of patients experiencing grade 3-4 myelotoxicity. One woman had life-threatening toxicity. This regimen yields a high response rate with acceptable toxicity and should be prospectively compared with other regimens. The high rate of pathologic complete and optimal responses might impact positively on survival, but only a longer follow-up period will allow objective assessment of this impact. The specific roles of paclitaxel and ifosfamide in this regimen remain to be fully understood. PMID- 10697041 TI - Sequential chemoradiation therapy with vinorelbine, ifosfamide, and cisplatin in stage IIIB non-small cell lung cancer: a phase II study. AB - Meta-analysis has demonstrated survival benefit for patients with stage IIIB non small cell lung cancer treated with sequential chemoradiotherapy versus radiotherapy alone. The introduction of chemotherapy as part of a multimodality approach has improved the outcome in this poor prognostic subset of cancer patients. In the present phase II study we evaluated the safety and activity of a new cisplatin-based three-drug regimen consisting of vinorelbine/ifosfamide/cisplatin (VIP) followed by curative thoracic irradiation in 28 patients with stage IIIB non-small cell lung cancer. Patients received vinorelbine 25 mg/m2 on days 1 and 8, ifosfamide 3 g/m2 on day 1 (with mesna), and cisplatin 80 mg/m2 on day 1 every 3 weeks. After three courses of induction chemotherapy, patients with objective response or stable disease were eligible for thoracic radiotherapy. Twenty-six of the 28 patients received at least three courses of chemotherapy and were evaluable for response. The response rate to induction VIP was 58% (15 of 26 patients; one complete response and 14 partial responses). Seven patients had disease stabilization and four progressed during chemotherapy. Radiation treatment started from 4 to 6 weeks after the end of chemotherapy with standard fractionation (200 cGy/day, 5 fractions/wk/6 wk). Eighteen of 22 patients started thoracic irradiation; 14 completed the treatment plan, reaching the total dose of 60 Gy. The most relevant acute and late toxicities of radiotherapy were grade 3 dysphagia and pneumonitis in two patients and grade 3 lung fibrosis in six patients. By comparing the tumor volumes before and after radiation treatment we observed six clinical remissions, three stable diseases, and five local progressions. The first site of recurrence was local in 10 of 18 patients (56%), distant in seven patients (38.8%), and both local and distant in one patient. Median progression-free survival and overall survival for the patients treated with radiotherapy (18 patients) were 14 months (range, 4 to 36 months) and 26 months (range, 7 to 54+ months), respectively; the 1- and 2 year survival rates were 61% and 52%. Curative thoracic radiotherapy was well tolerated after VIP induction chemotherapy; it reduced residual tumor volume in six patients. PMID- 10697042 TI - Paclitaxel (Taxol) combination therapy for resistant germ cell tumors. AB - Rob Paclitaxel (Taxol; Bristol-Myers Squibb Company Princeton, NJ) showed antitumor activity in patients with germ cell tumors resistant to combination cisplatin-containing chemotherapy and was included in two risk-directed first line combination salvage programs. Patients with relapsed germ cell tumors originating from a testis primary site were treated in a phase I/II trial with conventional-dose paclitaxel, ifosfamide, and cisplatin. Fifteen of 21 evaluable patients (71%) achieved a complete or partial response with normal serum tumor markers; 12 patients (57%) remain progression free with a median follow-up period of 15 months. Accrual to this trial continues to further define efficacy and toxicity. In a second trial, patient with cisplatin-resistant germ cell tumors and unfavorable prognostic features (prior incomplete response or an extragonadal primary site) were treated with a dose-intensive program consisting of rapid recycling of paclitaxel plus ifosfamide followed by carboplatin plus etoposide with stem cell support. Twenty-one of 37 assessable patients (57%) achieved a complete response and two (5%) achieved a partial response with negative serum tumor markers; 15 (41%) of these patients remain in durable response with a median follow-up period of 30 months. PMID- 10697043 TI - Paclitaxel (Taxol)/ifosfamide-based chemotherapy in patients with recurrent or metastatic squamous cell carcinoma of the head and neck. AB - Squamous cell carcinoma of the head and neck (SCCHN) is a functionally and cosmetically devastating tumor, and its treatment and the economic costs associated with aggressive treatment are substantial. Even with aggressive standard local surgery, radiotherapy, or both, locally advanced tumors recur in approximately two thirds of patients and the prognosis for those whose disease recurs or metastasizes is dismal. Newer chemotherapeutic agents such as ifosfamide and taxanes (paclitaxel [Taxol; Bristol-Myers Squibb Company, Princeton, NJ] and docetaxel) have shown higher response rates than those seen with conventional agents and renew hope of prolonged survival, improved quality of life, and greater convenience. We recently conducted two consecutive phase II studies using paclitaxel/ifosfamide/cisplatin and paclitaxel/ifosfamide/carboplatin for patients with recurrent or metastatic SCCHN. These two regimens yielded high response rates (including complete responses) and appear to be promising therapies for SCCHN. This report describes our experience with these two regimens, including a comparison of their toxic effects. PMID- 10697045 TI - Healthy People 2010 initiative launched. PMID- 10697044 TI - A piece of my mind. Attention must be paid. PMID- 10697046 TI - New center director states "complementary" agenda. PMID- 10697047 TI - Whose news is it, anyway? PMID- 10697048 TI - Picture this: smoking kills. PMID- 10697049 TI - From the Centers for Disease Control and Prevention. HIV/AIDS among racial/ethnic minority men who have sex with men--United States, 1989-1998. PMID- 10697050 TI - From the Centers for Disease Control and Prevention. Guidelines for surveillance, prevention, and control of West Nile virus infection--United States. PMID- 10697051 TI - Risk of HIV transmission through breastfeeding. PMID- 10697052 TI - Clinical diagnosis of carpal tunnel syndrome. PMID- 10697054 TI - Clinical diagnosis of carpal tunnel syndrome. PMID- 10697053 TI - Clinical diagnosis of carpal tunnel syndrome. PMID- 10697055 TI - Clinical diagnosis of carpal tunnel syndrome. PMID- 10697056 TI - Ventilator-induced lung injury. PMID- 10697057 TI - Computed tomography for predicting complications of lumbar puncture. PMID- 10697058 TI - Leprosy in the eastern United States. PMID- 10697059 TI - Streptococcus equinus endocarditis in a woman with pulmonary histiocytosis. PMID- 10697060 TI - Estrogen replacement therapy for treatment of mild to moderate Alzheimer disease: a randomized controlled trial. Alzheimer's Disease Cooperative Study. AB - CONTEXT: Several reports from small clinical trials have suggested that estrogen replacement therapy may be useful for the treatment of Alzheimer disease (AD) in women. OBJECTIVE: To determine whether estrogen replacement therapy affects global, cognitive, or functional decline in women with mild to moderate AD. DESIGN: The Alzheimer's Disease Cooperative Study, a randomized, double-blind, placebo-controlled clinical trial conducted between October 1995 and January 1999. SETTING: Thirty-two study sites in the United States. PARTICIPANTS: A total of 120 women with mild to moderate AD and a Mini-Mental State Examination score between 12 and 28 who had had a hysterectomy. INTERVENTIONS: Participants were randomized to estrogen, 0.625 mg/d (n = 42), or 1.25 mg/d (n = 39), or to identically appearing placebo (n = 39). One subject withdrew after randomization but before receiving medication; 97 subjects completed the trial. MAIN OUTCOME MEASURES: The primary outcome measure was change on the Clinical Global Impression of Change (CGIC) 7-point scale, analyzed by intent to treat; secondary outcome measures included other global measures as well as measures of mood, specific cognitive domains (memory, attention, and language), motor function, and activities of daily living; compared by the combined estrogen groups vs the placebo group at 2, 6, 12, and 15 months of follow-up. RESULTS: The CGIC score for estrogen vs placebo was 5.1 vs 5.0 (P = .43); 80% of participants taking estrogen vs 74% of participants taking placebo worsened (P = .48). Secondary outcome measures also showed no significant differences, with the exception of the Clinical Dementia Rating Scale, which suggested worsening among patients taking estrogen (mean posttreatment change in score for estrogen, 0.5 vs 0.2 for placebo; P = .01). CONCLUSIONS: Estrogen replacement therapy for 1 year did not slow disease progression nor did it improve global, cognitive, or functional outcomes in women with mild to moderate AD. The study does not support the role of estrogen for the treatment of this disease. The potential role of estrogen in the prevention of AD, however, requires further research. PMID- 10697061 TI - Efficacy and safety of the oral neuraminidase inhibitor oseltamivir in treating acute influenza: a randomized controlled trial. US Oral Neuraminidase Study Group. AB - CONTEXT: Previous studies have shown oseltamivir, a neuraminidase inhibitor, to be effective in preventing influenza and treating experimental influenza. OBJECTIVE: To evaluate the efficacy and safety of oseltamivir in the treatment of naturally acquired influenza infection. DESIGN: Randomized, placebo-controlled, double-blind study conducted January through March 1998. SETTING: Sixty primary care and university health centers throughout the United States. PARTICIPANTS: A total of 629 healthy nonimmunized adults aged 18 to 65 years with febrile respiratory illness of no more than 36 hours' duration with temperature of 38 degrees C or more plus at least 1 respiratory symptom and 1 constitutional symptom. INTERVENTIONS: Individuals were randomized to 1 of 3 treatment groups with identical appearing pills: oral oseltamivir phosphate, 75 mg twice daily (n = 211) or 150 mg (n = 209) twice daily, or placebo (n = 209). MAIN OUTCOME MEASURES: Duration and severity of illness in individuals infected with influenza. RESULTS: Two individuals withdrew before receiving medication and were excluded from further analyses. A total of 374 individuals (59.6%) were infected with influenza. Their duration of illness was reduced by more than 30% with both oseltamivir, 75 mg twice daily (median, 71.5 hours; P < .001), and oseltamivir, 150 mg twice daily (median, 69.9 hours; P = .006), compared with placebo (median, 103.3 hours). Severity of illness was reduced by 38% (median score, 597 score hours; P < .001) with oseltamivir, 75 mg twice daily, and by 35% (median score, 626 score-hours; P < .001) with oseltamivir, 150 mg twice daily, vs placebo (median score, 963 score-hours). Oseltamivir treatment reduced the duration of fever and oseltamivir recipients returned to usual activities 2 to 3 days earlier than placebo recipients (P < or = .05). Secondary complications such as bronchitis and sinusitis occurred in 15% of placebo recipients compared with 7% of combined oseltamivir recipients (P = .03). Among all 629 subjects, oseltamivir reduced illness duration (76.3 hours and 74.3 hours for 75 mg and 150 mg, respectively, vs 97.0 hours for placebo; P = .004 for both comparisons) and illness severity (686 score-hours and 629 score-hours for 75 mg and 150 mg, respectively, vs 887 score-hours for placebo; P < .001 for both comparisons). Nausea and vomiting occurred more frequently in both oseltamivir groups (combined, 18.0% and 14.1%, respectively; P = .002) than in the placebo group (7.4% and 3.4%; P < .001). CONCLUSIONS: Our data suggest that oral oseltamivir treatment reduces the duration and severity of acute influenza in healthy adults and may decrease the incidence of secondary complications. PMID- 10697062 TI - Trends in the prescribing of psychotropic medications to preschoolers. AB - CONTEXT: Recent reports on the use of psychotropic medications for preschool-aged children with behavioral and emotional disorders warrant further examination of trends in the type and extent of drug therapy and sociodemographic correlates. OBJECTIVES: To determine the prevalence of psychotropic medication use in preschool-aged youths and to show utilization trends across a 5-year span. DESIGN: Ambulatory care prescription records from 2 state Medicaid programs and a salaried group-model health maintenance organization (HMO) were used to perform a population-based analysis of three 1-year cross-sectional data sets (for the years 1991, 1993, and 1995). SETTING AND PARTICIPANTS: From 1991 to 1995, the number of enrollees aged 2 through 4 years in a Midwestern state Medicaid (MWM) program ranged from 146,369 to 158,060; in a mid-Atlantic state Medicaid (MAM) program, from 34,842 to 54,237; and in an HMO setting in the Northwest, from 19,107 to 19,322. MAIN OUTCOME MEASURES: Total, age-specific, and gender-specific utilization prevalences per 1000 enrollees for 3 major psychotropic drug classes (stimulants, antidepressants, and neuroleptics) and 2 leading psychotherapeutic medications (methylphenidate and clonidine); rates of increased use of these drugs from 1991 to 1995, compared across the 3 sites. RESULTS: The 1995 rank order of total prevalence in preschoolers (per 1000) in the MWM program was: stimulants (12.3), 90% of which represents methylphenidate (11.1); antidepressants (3.2); clonidine (2.3); and neuroleptics (0.9). A similar rank order was observed for the MAM program, while the HMO had nearly 3 times more clonidine than antidepressant use (1.9 vs 0.7). Sizable increases in prevalence were noted between 1991 and 1995 across the 3 sites for clonidine, stimulants, and antidepressants, while neuroleptic use increased only slightly. Methylphenidate prevalence in 2- through 4-year-olds increased at each site: MWM, 3-fold; MAM, 1.7-fold; and HMO, 3.1-fold. Decreases occurred in the relative proportions of previously dominant psychotherapeutic agents in the stimulant and antidepressant classes, while increases occurred for newer, less established agents. CONCLUSIONS: In all 3 data sources, psychotropic medications prescribed for preschoolers increased dramatically between 1991 and 1995. The predominance of medications with off-label (unlabeled) indications calls for prospective community-based, multidimensional outcome studies. PMID- 10697063 TI - Incidence of cervical squamous intraepithelial lesions in HIV-infected women. AB - CONTEXT: Women infected with human immunodeficiency virus (HIV) are at increased risk for cervical squamous intraepithelial lesions (SILs), the precursors to invasive cervical cancer. However, little is known about the causes of this association. OBJECTIVES: To compare the incidence of SILs in HIV-infected vs uninfected women and to determine the role of risk factors in the pathogenesis of such lesions. DESIGN: Prospective cohort study conducted from October 1,1991, to June 30, 1996. SETTING: Urban clinics for sexually transmitted diseases, HIV infection, and methadone maintenance. PARTICIPANTS: A total of 328 HIV-infected and 325 uninfected women with no evidence of SILs by Papanicolaou test or colposcopy at study entry. MAIN OUTCOME MEASURE: Incident SILs confirmed by biopsy, compared by HIV status and risk factors. RESULTS: During about 30 months of follow-up, 67 (20%) HIV-infected and 16 (5%) uninfected women developed a SIL (incidence of 8.3 and 1.8 cases per 100 person-years in sociodemographically similar infected and uninfected women, respectively [P<.001]). Of incident SILs, 91% were low grade in HIV-infected women vs 75% in uninfected women. No invasive cervical cancers were identified. By multivariate analysis, significant risk factors for incident SILs were HIV infection (relative risk [RR], 3.2; 95% confidence interval [CI], 1.7-6.1), transient human papillomavirus (HPV) DNA detection (RR, 5.5; 95% CI, 1.4-21.9), persistent HPV DNA types other than 16 or 18 (RR, 7.6; 95% CI, 1.9-30.3), persistent HPV DNA types 16 and 18 (RR, 11.6; 95% CI, 2.7-50.7), and younger age (<37.5 years; RR, 2.1; 95% CI, 1.3-3.4). CONCLUSIONS: In our study, 1 in 5 HIV-infected women with no evidence of cervical disease developed biopsy-confirmed SILs within 3 years, highlighting the importance of cervical cancer screening programs in this population. PMID- 10697064 TI - A 3-level prognostic classification in septic shock based on cortisol levels and cortisol response to corticotropin. AB - CONTEXT: The hypothalamic-pituitary-adrenal axis is a major determinant of the host response to stress. The relationship between its activation and patient outcome is not known. OBJECTIVE: To evaluate the prognostic value of cortisol levels and a short corticotropin stimulation test in patients with septic shock. DESIGN AND SETTING: Prospective inception cohort study conducted between October 1991 and September 1995 in 2 teaching hospital adult intensive care units in France. PARTICIPANTS: A total of 189 consecutive patients who met clinical criteria for septic shock. INTERVENTION: A short corticotropin stimulation test was performed in all patients by intravenously injecting 0.25 mg of tetracosactrin; blood samples were taken immediately before the test (T0) and 30 (T30) and 60 (T60) minutes afterward. MAIN OUTCOME MEASURES: Twenty-eight-day mortality as a function of variables collected at the onset of septic shock, including cortisol levels before the corticotropin test and the cortisol response to corticotropin (delta max, defined as the difference between T0 and the highest value between T30 and T60). RESULTS: The 28-day mortality was 58% (95% confidence interval [CI], 51%-65%) and median time to death was 17 days (95% CI, 14-27 days). In multivariate analysis, independent predictors of death (P < or = .001 for all) were McCabe score greater than 0, organ system failure score greater than 2, arterial lactate level greater than 2.8 mmol/L, ratio of PaO2 to fraction of inspired oxygen no more than 160 mm Hg, cortisol level at T0 greater than 34 microg/dL and delta max no more than 9 microg/dL. Three groups of patient prognoses were identified: good (cortisol level at T0 < or = 34 microg/dL and delta max > 9 microg/dL; 28-day mortality rate, 26%), intermediate (cortisol level at T0 34 microg/dL and delta max < or = 9 microg/dL or cortisol level at T0 > 34 microg/dL and delta max > 9 microg/dL; 28-day mortality rate, 67%), and poor (cortisol level at T0 > 34 microg/dL and delta max < or = 9 microg/dL; 28-day mortality rate, 82%). CONCLUSION: Our data suggest that a short corticotropin test has a good prognostic value and could be helpful in identifying patients with septic shock at high risk for death. PMID- 10697065 TI - An 80-year-old man with memory loss. PMID- 10697066 TI - A 75-year-old man with congestive heart failure, 1 year later. PMID- 10697067 TI - Estrogen and Alzheimer disease: plausible theory, negative clinical trial. PMID- 10697068 TI - Expanding the treatment options for influenza. PMID- 10697069 TI - Psychotropic drug use in very young children. PMID- 10697070 TI - Withdrawing very low-burden interventions in chronically ill patients. PMID- 10697072 TI - JAMA Patient Page: cervical cancer. PMID- 10697071 TI - Decisions to withdraw life-sustaining treatment: a moral algorithm. PMID- 10697073 TI - Neuroprotective strategies for basal ganglia degeneration: Parkinson's and Huntington's diseases. AB - There are three main mechanisms of neuronal cell death which may act separately or cooperatively to cause neurodegeneration. This lethal triplet of metabolic compromise, excitotoxicity, and oxidative stress causes neuronal cell death that is both necrotic and apoptotic in nature. Aspects of each of these three mechanisms are believed to play a role in the neurodegeneration that occurs in both Parkinson's and Huntington's diseases. Strategies to rescue or protect injured neurons usually involve promoting neuronal growth and function or interfering with neurotoxic processes. Considerable research has been done on testing a large array of neuroprotective agents using animal models which mimic these disorders. Some of these approaches have progressed to the clinical arena. Here, we review neuroprotective strategies which have been found to successfully ameliorate the neurodegeneration associated with Parkinson's and Huntington's diseases. First, we will give an overview of the mechanisms of cell death and the background of Parkinson's and Huntington's diseases. Then we will elaborate on a range of neuroprotective strategies, including neurotrophic factors, anti excitotoxins, antioxidants, bioenergetic supplements, anti-apoptotics, immunosuppressants, and cell transplantation techniques. Most of these approaches hold promise as potential therapies in the treatment of these disorders. PMID- 10697074 TI - Unraveling of important neurobiological mechanisms by the use of pure, fully differentiated neurons obtained from adult animals. AB - An important, and often overlooked, problem in the neurochemical approach to neurobiological problems is that analysis of tissue involves almost always a heterogeneous population of cells (neurons, glia and other types of tissue cells). The use of cell cultures has obvious limitations such as that they derive from embryonic or immediately postnatal animals; in addition, the cell culture conditions most certainly are quite different from the real tissue environment for the nerve cells. We underline here an alternative strategy, which is not new, but which, in our view, has already given formidable contributions to neurobiological studies and still is giving results of great importance. This is the technique proposed and used since the late fifties and early sixties by the senior author (H. Hyden). The method involves the isolation of the big vestibular neurons from the adult rabbit vestibular nucleus. The neurons, fully differentiated and performing a precisely defined function, are obtained rapidly and completely free from surrounding glial cells. The separate microbiochemical study of these cells and their surrounding glia has yielded already in 1962, the information that modifications in gene expression are associated with plastic modifications of the function of the relevant neurons, which take place in the behavioral event of learning. Another important concept was formulated in the same time period following determination of the activities of energy metabolism related enzymes separately in vestibular neurons and their glia under vestibular stimulation. This is the concept that, under increased functional activity glia increases its anaerobic metabolism and passes then on the resulting metabolites to the neurons for aerobic metabolism. Both these concepts (RNA and memory; metabolic cooperation between glia and neurons) are nowadays widely accepted. In addition, this approach with pure big nerve cells has allowed in recent years the discovery of a novel mechanism for chloride extrusion in these cells. This mechanism utilizes structures similar to GABA activated chloride channels in cyclic modifications resulting in the final extrusion of chloride ions. The energy for the process is provided by a protein phosphorylation step. Future approaches are warranted such as the possibility of recognizing by RT-PCR specific neuronal mRNAs and their modification in expression in relation to function and plastic modifications (learning). Another possible interesting application appears to be the recognition of the mRNAs for GABA(A) receptor subunits expressed here in these neurons in relation to the physiological and pharmacological characteristics of these native neuronal GABA(A) receptors. PMID- 10697075 TI - Vacuum pack technique of temporary abdominal closure: a 7-year experience with 112 patients. AB - BACKGROUND: Temporary abdominal wound closure after celiotomy for trauma is often desirable. The ideal method of temporary closure should allow rapid closure, easy maintenance, and allow reexploration and wound repair with minimal tissue damage. Over the past 7 years, we have successfully used a vacuum closure system (the vacuum pack) for temporary management of the open abdomen. METHODS: Medical records of trauma patients undergoing exploratory celiotomy from April of 1992 to February of 1999 were reviewed. Demographic data as well as indications for open abdominal management and complications of open-abdominal management were collected. RESULTS: Two hundred sixteen vacuum packs were performed in 112 trauma patients. Of the 216 vacuum packs placed, 2.8% were placed for increased intra abdominal pressure, 5.3% for inability to achieve tension-free fascial closure, 20% for damage control, 55% for reexploration, and 16.7% for a combination of factors. Sixty-two patients (55.4%) went on to primary closure and 25 patients (22.3%) underwent polyglactin mesh repair of the defect followed by wound granulation and eventual skin grafting. Twenty-two patients (19.6%) died before abdominal closure was attempted. Five patients (4.5%) developed enterocutaneous fistulae. Five patients (4.5%) developed intra-abdominal abscesses. There were no eviscerations. Three patients (2.7%) required further explorations after abdominal closure. Overall mortality rate was 25.9%, none related to the vacuum pack. CONCLUSIONS: The vacuum pack is the temporary abdominal wound closure of choice in patients undergoing open abdominal management at our institution. Primary closure is achieved in the majority of patients with a low rate of complication. The technique is simple and easily mastered. Technical complications are rare and easily repaired. PMID- 10697076 TI - Is routine arteriography mandatory for penetrating injury to zone 1 of the neck? Zone 1 Penetrating Neck Injury Study Group. AB - OBJECTIVE: Surgical dogma dictates that the evaluation of all penetrating zone 1 neck injuries must include arteriography to reliably exclude arterial injury requiring operation. This study was done to determine whether patients with normal findings at physical examination (PE) and on chest radiographs (CXR) really do require arteriography to identify occult, surgically important arterial injuries. METHODS: All penetrating zone 1 neck injuries in five Level I trauma centers over a 10-year period were reviewed retrospectively. Data collected included demographics, results of PE, CXR findings, other diagnostic studies done, injuries identified, need for operation, and operative findings. Arterial injury was defined as any injury to the aorta or brachiocephalic, subclavian, vertebral, or carotid arteries found on arteriography, duplex, or at operation. RESULTS: Of 138 patients studied, there were 28 arterial injuries. Of the total group of 138 patients, 36 patients had normal findings at PE and on CXR. None of these 36 patients had an arterial injury. The negative predictive value of normal PE and CXR together is 100% in this series. CONCLUSIONS: Patients with penetrating wounds to zone I who have no evidence of vascular injury on PE and who have normal findings on CXR may not require routine arteriography. Further study is needed to confirm these findings. PMID- 10697077 TI - Chlorpromazine modulates cytokine expression in the liver and lung after burn injury and endotoxemia. AB - BACKGROUND: Previous data from our laboratory have demonstrated that alterations in cytokine production occur in the lung and liver as the result of a two-hit model of injury, i.e., burn with subsequent endotoxin administration. The purpose of this study was to determine whether the phenothiazine derivative chlorpromazine would alter cytokine production in a sequential model of injury. METHODS: By using a sublethal burn/endotoxemia model, B2D6F1 mice (n = 40) were assigned to two groups and subjected to a 15% full-thickness burn. Three days after burn injury, one group (BURN/ETX) received 2.5 mg/kg Escherichia coli endotoxin intraperitoneally, and the other group (CPZ) received 4 mg/kg chlorpromazine 1 hour before the administration of 2.5 mg/kg E. coli endotoxin intraperitoneally. At selected time points, the animals were killed and lung and liver were removed and processed for protein and total RNA. Northern blots and enzyme-linked immunosorbent assays were used to assess the production of tumor necrosis factor-alpha, macrophage inflammatory protein-1alpha, and interleukin 10. RESULTS: Chlorpromazine significantly reduced tumor necrosis factor-alpha mRNA and protein expression in the liver. Macrophage inflammatory protein-1alpha mRNA was reduced by chlorpromazine in both liver and lung. Interleukin-10 production was not altered by chlorpromazine. CONCLUSION: The reduction of tumor necrosis factor-alpha and macrophage inflammatory protein-1alpha by chlorpromazine in the liver and lungs may have potential as a pharmaceutical agent that may dampen the inflammatory response in a model of sequential injury. PMID- 10697078 TI - Complications of prone ventilation in patients with multisystem trauma with fulminant acute respiratory distress syndrome. AB - INTRODUCTION: Prone ventilation improves oxygenation in selected patients with acute respiratory distress syndrome (ARDS). However, prone positioning of critically ill patients with multiple invasive lines and tubes is potentially dangerous. Trauma patients, in particular, may require special consideration because of skeletal fixation devices or prior operative procedures. Our objective was to critically evaluate our experience with prone positioning in patients with severe postinjury ARDS. METHODS: Injured patients admitted to our Level I trauma center who developed ARDS were prospectively identified. Serial lung injury severity and pulmonary mechanical data, as well as complications of prone ventilation were recorded. RESULTS: During the 12-month period ending August of 1998, nine patients with postinjury ARDS were treated with prone ventilation because of hypoxemia refractory to other ventilatory strategies. All patients suffered blunt trauma. Their mean age was 29 +/- 4.5 years; seven patients were men. The average Injury Severity Score was 26 +/- 5; and, at the time of prone positioning, the mean Lung Injury Score was 3.5. The mean PaO2/FIO2 ratio increased from 75 +/- 7 to 147 +/- 27 with prone ventilation (p < 0.05, paired t test); and in six patients, the FIO2 could be decreased. Four major complications occurred (44%). One patient experienced a midline abdominal wound dehiscence. Severe facial or upper chest wall pressure necrosis developed in two patients, despite extensive padding and careful attention to skin care. The fourth patient sustained a cardiac arrest immediately after prone positioning. CONCLUSION: Prone ventilation in postinjury patients with ARDS may improve oxygenation but has the potential for significant complications. Careful consideration is required before prone positioning in this subset of patients. PMID- 10697079 TI - Outcomes of trauma patients who survive prolonged lengths of stay in the intensive care unit. AB - OBJECTIVE: To examine a subgroup of severely injured patients spending > or = 3 weeks in the intensive care unit (ICU) and to determine their disposition and eventual functional outcome. METHODS: A retrospective review of our trauma registry and medical records over a 7-year period (January of 1991 to December of 1997) identified 115 patients with ICU length of stay (LOS) > or = 3 weeks. Variables selected included age, length of stay, injury severity score, injuries, disposition, and charges. Functional independence measures (FIM) were obtained in patients requiring inpatient rehabilitation and a written questionnaire (Rand 36 item Health Survey) was mailed to all patients alive at discharge. RESULTS: Mean ICU length of stay of the 115 patients was 36 days (range, 21-106 days); mean age, 49 years (range, 4-89 years); 73 patients (63%) were males, 42 patients (37%) were females. Overall mortality was 22% (n = 25). The remaining 90 patients survived to discharge with the following disposition: rehabilitation facility 60% (n = 54), home with temporary disability 22% (n = 20), nursing home 8% (n = 7), home with permanent disability 4% (n = 4), transferred 6% (n = 5). Mean hospital charge was $193,000 (range, $77,000-$528,000). No variable or combination could predict outcome except age. Elderly patients (age > or = 75, n = 24) had an overall mortality of 42% (n = 10). Eight of 14 survivors fulfilled admission criteria and entered our rehabilitation facility. The remaining six elderly patients either went to nursing homes or were permanently disabled. Complete FIM scores were available on 47 of 54 patients who went to rehabilitation facility. The mean rehabilitation facility admission FIM score was 52, indicating either complete dependence or the need for moderate assistance. After they had remained at the rehabilitation facility for a mean of 48 days (range, 7-278 days), patients' FIM scores improved to a mean of 86, signifying minimal contact assistance or supervision only. Three-month follow-up FIM scores continued to improve to a mean of 101, a score denoting complete independence. Elderly patients within the rehabilitation facility fared as well as the younger group. For the Rand-36 survey, 47 of 90 patients or family members were contacted. Twelve patients died since discharge, leaving 35 patients to complete the survey. Despite excellent FIM scores, overall mean health was only fair to good, with limitations to activity and lack of energy cited as the main problems. CONCLUSION: Despite tremendous resource utilization, the majority of trauma patients with prolonged ICU stays can eventually return to varying degrees of functional daily living and independence, but not to preinjury levels. A subgroup of severely injured elderly patients had a significantly higher mortality rate. However, elderly survivors that entered our rehabilitation facility fared as well as the younger patients. PMID- 10697080 TI - Submental endotracheal intubation: an alternative to tracheotomy in patients with midfacial and panfacial fractures. AB - BACKGROUND: The submental route for endotracheal intubation has been proposed as an alternative to tracheotomy in the surgical management of patients with maxillofacial trauma. The purpose of this study was to review our experience with this procedure. METHODS: Medical records of 25 patients who had surgical reduction of midfacial or panfacial fractures while securing their airway with submental intubation were reviewed. After standard orotracheal intubation, a passage was created by blunt dissection with a hemostat clamp through the floor of the mouth in the submental area. The proximal end of the orotracheal tube was pulled through the submental incision. Surgery was completed with minimal interference from the endotracheal tube. At the end of surgery, the tube was pulled back to the usual oral route. RESULTS: Mean duration of surgery was 7.9 hours (range, 2-16 hours). Mean duration of postoperative mechanical ventilation was 5.2 days (range, 1-24 days). Fourteen of these patients required prolonged (>24 hours) postoperative mechanical ventilation because of associated injuries. Two patients later required a tracheotomy because of prolonged respiratory failure. One patient died of multiple organ failure. One complication of the submental intubation was observed: a superficial infection of the submental wound. CONCLUSION: Submental intubation is a simple technique associated with a low morbidity. It is an attractive alternative to tracheotomy in the surgical management of selected cases of maxillofacial trauma. PMID- 10697081 TI - Translocation of endotoxin and acute-phase proteins in malleolar fractures. AB - BACKGROUND AND OBJECTIVE: Translocation of endotoxins was demonstrated for multiple injury but not for minor trauma such as isolated malleolar fractures. Major trauma leads to substantial changes in the plasma concentration of acute phase proteins. However, isolated malleolar fractures are minor trauma. The objective of this study was to elucidate the kinetics of endotoxemia and the ability of plasma to inactivate endotoxin of patients operated on malleolar fractures and to demonstrate the early time course of the acute-phase proteins C reactive protein, transferrin, alpha1-acid glycoprotein, haptoglobin, and interleukin-6 and to correlate them with the amount of endotoxemia. METHODS: Thirty patients with malleolar fractures were operated on within 6 hours after injury. Blood was collected immediately after admission and regularly up to 96 hours after surgery. RESULTS: Preoperative endotoxin plasma levels were increased compared with that of healthy individuals (0.05 +/- 0.017 vs. 0.02 EU/mL). Endotoxemia peaked 0.5 hours after the surgical procedure at 0.096 +/- 0.03 (p < 0.05 vs. healthy) and decreased to almost normal values after 24 hours. The ability of the plasma to inactivate endotoxin was significantly reduced after the surgical procedure compared with normal subjects (recovery, 0.17 +/- 0.028 EU/mL vs. 0.04 +/- 0.01 EU/mL; p < 0.05). Plasma interleukin-6 peaked 0.5 hours postoperatively (114 +/- 11 pg/mL, p < 0.05 vs. healthy), decreasing thereafter. C-Reactive protein peaked at 45 +/- 5 mg/mL (p < 0.05) 48 hours after injury. Transferrin decreased significantly postoperatively (2.41 +/- 0.12 mg/mL vs. pre OP 2.65 +/- 0.1 mg/mL) and remained on this level for 96 hours. Both, alpha1-acid glycoprotein and haptoglobin increased postoperatively until day 4 (0.78 +/- 0.06 mg/mL to 1.15 +/- 0.08 mg/mL and 1.51 +/- 0.12 mg/mL to 3.24 +/- 0.22 mg/mL). There was no correlation between endotoxemia and the concentrations of the acute phase proteins and interleukin-6. CONCLUSION: Surgery for malleolar fractures is associated with temporary endotoxemia and temporary reduced endotoxin inactivation capacity of the plasma. The injury and the surgical procedure leads to substantial changes in the plasma concentrations of acute-phase proteins. The relation between endotoxemia and acute-phase response is not dose dependent. PMID- 10697082 TI - Antibiotic-impregnated autogenic cancellous bone grafting is an effective and safe method for the management of small infected tibial defects: a comparison study. AB - OBJECTIVE: Bone grafting plays an important role in reconstructing infected tibial nonunions. The effects of antibiotic-impregnated bone grafting in infection elimination and bone incorporation was reported in this retrospective study. METHODS: Ninety-six patients treated for infected tibial nonunions were evaluated. These patients were managed with local antibiotic bead therapy and staged antibiotic-impregnated autogenous cancellous bone graft or pure autogenous cancellous bone graft. Patients were randomized to antibiotic-impregnated bone grafting or bone grafting-only groups on the basis of whether the admission date was odd or even. Patients were divided into two groups (antibiotic-impregnated bone grafting group and pure cancellous bone grafting group), according to the procedure used in preparing the bone grafts. The antibiotic-impregnated bone grafting group included 37 men and 9 women whose average age was 36 years (range, 17 to 72 years). The average follow-up period was 4.8 years. By using the Cierny Mader staging classification of chronic osteomyelitis, 32 of 46 patients (70%) were stage 4A, and 14 of 36 patients (30%) were stage 4B. The pure cancellous bone grafting group included 39 men and 11 women whose average age was 37 years (range, 18 to 72 years). The average follow-up period was 4.5 years (range, 4 to 6 years). Thirty-nine of 50 patients (78%) were stage 4A, and 11 of 50 patients (22%) were stage 4B. The bone defects in both groups ranged from 2 to 4 cm. RESULTS: Wound healing and bony union were achieved in the antibiotic-impregnated bone grafting group. Only two patients had recurrent infections. The infection arrest rate was 95.6%. However, 9 of 50 patients in the pure cancellous bone grafting group had recurrent infections. The infection arrest rate was 82%. The antibiotic-impregnated bone grafting group had significantly superior results (95.6% vs. 82% chi2 test, p < 0.05) in infection elimination than the pure cancellous bone grafting group. CONCLUSION: After 4 to 6 years of follow-up, our results suggest that the use of impregnating antibiotics have no adverse effects on autogenic cancellous bone graft incorporation and could help to eliminate infection effectively. PMID- 10697083 TI - Treatment of femoral shaft fractures in children by intramedullary Kirschner wires. AB - BACKGROUND: We present a retrospective analysis of a case series to evaluate closed intramedullary Kirschner wire (K-wire) fixation as a surgical technique in the treatment of femoral shaft fractures in children. METHODS: Fifty-three femoral shaft fractures (at various levels) were fixed by using closed intramedullary K-wires. The patient was placed supine on an orthopedic traction table. Under fluoroscopic control, two K-wires (2.5-3.5 mm thick) were introduced from distal metaphysis to the proximal metaphysis, one each, from medial and lateral cortices. In three distal fractures, K-wires were inserted antegrade from the proximal to distal metaphysis through the lateral cortex. Early mobilization and weight bearing was allowed. Mean hospital stay was 6.5 days, and K-wires were removed after a mean of 5.6 months. The study included seven open fractures as well. RESULTS: Sound unions were achieved between 6 and 10 weeks without any significant complication. CONCLUSION: Closed intramedullary K-wire fixation for femoral shaft fractures in children is a simple surgical technique that has excellent clinical and functional results. PMID- 10697084 TI - Occult hypoperfusion is associated with increased morbidity in patients undergoing early femur fracture fixation. AB - BACKGROUND: The presence of persistent occult hypoperfusion (OH) is associated with higher morbidity and mortality rates after trauma. Early femur fracture fixation in trauma patients with multiple injuries is associated with decreased morbidity and mortality. Association of OH and incidence of postoperative complications after intramedullary (IM) fixation in patients with femur fractures was investigated. METHODS: A retrospective study design was used. All patients with femur fractures admitted to the trauma service of a Level I trauma center between January 1, 1995, and August 1, 1998, who were older than 18 years of age and who had IM fracture fixation within 24 hours of admission and serum lactate determinations on admission and at proscribed intervals, were included in the study. Patients with lactic acid levels > or = 2.5 mmol/L were determined to have OH. No patients had clinical signs of shock (hypotension, tachycardia, decreased urine output) on transfer to the operating room. Complete resuscitation was defined as a lactic acid level < 2.5 mmol/L. Patients were divided into two groups based on presence/absence of OH determined from the lactic acid level immediately before surgery. The incidence of all postoperative organ complications was recorded, and complication rates were compared between groups. Total hospital costs were also compared. RESULTS: One hundred seventy-seven patients with femur fractures were admitted to the trauma service during this period. Seventy-nine patients met initial criteria for inclusion in the study. Further review excluded 32 patients. Occult hypoperfusion was present in 20 patients before early IM fixation (group 2). Twenty-seven patients were completely resuscitated before early IM fixation (group 1). Injury Severity Scores were similar in both groups. Group 2 had 35 complications in 20 patients, and group 1 had 11 complications in 27 patients. A significant difference was found in incidence of postoperative complications in group 1 (20%) versus group 2 (50%). Group 2 also had a significantly higher proportion of postoperative infections than group 1 (72% vs. 28%, respectively) and higher total hospital costs ($46,469 vs. $23,139). CONCLUSION: The presence of OH in trauma patients undergoing early IM fixation of a femur fracture is associated with a twofold higher incidence of postoperative complications. Clinical judgment, not surgical dogma, should guide the timing of IM fixation in these patients. Identifying and correcting OH through relatively simple resuscitative measures may be advantageous in reducing morbidity in the patient with multiple injuries. PMID- 10697085 TI - Deep vein thrombosis prophylaxis in hip fractures: a comparison of the arteriovenous impulse system and aspirin. AB - BACKGROUND: A prospective, randomized controlled trial was used to compare the efficacy of the arteriovenous (AV) impulse system and aspirin in reducing venous thrombosis after fracture to the femoral neck. METHODS: A total of 143 patients underwent hemiarthroplasty, after which 70 patients were treated with the AV pump and a second group of 73 patients were commenced on 325 mg of aspirin. Duplex ultrasound was used to assess both proximal and distal venous thrombi on days 7 to 10. Calf and thigh circumferences were also measured. RESULTS: Thrombi developed in seven of the patients treated with aspirin and in four patients treated with the AV pump. No statistically significant difference could be established (p = 0.109). There was a significant reduction in both calf (p = 0.003) and thigh (p = 0.002) swelling in the group treated with the AV pump. Neither treatment group was a significant predictor of a poorer outcome by using logistical regression analysis (p = 0.258). CONCLUSIONS: Both aspirin and the AV pump are effective in reducing thromboembolic events after hemiarthroplasty of the hip. PMID- 10697086 TI - Psychosocial support in rehabilitation after orthopedic injuries. AB - OBJECTIVE: Several studies have shown that psychosocial factors play a significant role in the recovery process after injuries. The aim of this study was to investigate whether psychosocial support would have a beneficial effect on outcome. METHODS: A total of 151 patients with orthopedic injuries were randomized into an intervention group and a control group. The intervention group was offered a psychosocial support program during the early phase of rehabilitation. RESULTS: One year after the injury, patients in the control group had an excess risk of having psychiatric complaints compared with patients in the intervention group (odds ratio = 2.74). They also reported a poorer quality of life according to Short Formula 36 Health Survey subscores for General Health (odds ratio = 2.3) and Vitality (odds ratio = 2.45). The length of the sick leave period did not differ between the groups. CONCLUSION: Psychosocial support during the early phase of rehabilitation after orthopedic injuries may have a beneficial effect on outcome when measured as quality of life. PMID- 10697087 TI - Validation of the International Classification of Diseases 10th Edition-based Injury Severity Score (ICISS). AB - OBJECTIVE: To compare the predictive power of International Classification of Diseases 10th Edition (ICD-10)-based International Classification of Diseases 9th Edition-based Injury Severity Score (ICISS) with Trauma and Injury Severity Score (TRISS) and ICD-9CM-based ICISS in the injury severity measure. METHODS: ICD-10 version of survival risk ratios was derived from 47,750 trauma patients from 35 emergency centers for 1 year. The predictive power of TRISS, the ICD-9CM-based ICISS and ICD-10-based ICISS were compared in a group of 367 severely injured patients admitted to two university hospitals. The predictive power was compared by using the measures of discrimination (disparity, sensitivity, specificity, misclassification rates, and receiver operating characteristic curve analysis) and calibration (Hosmer-Lemeshow goodness-of-fit statistics), all calculated by logistic regression procedure. RESULTS: ICD-10-based ICISS showed a lower performance than TRISS and ICD-9CM-based ICISS. When age and Revised Trauma Score were incorporated into the survival probability model, however, ICD-10-based ICISS full model showed a similar predictive power compared with TRISS and ICD 9CM-based ICISS full model. ICD-10-based ICISS had some disadvantages in predicting outcomes among patients with intracranial injuries. However, such weakness was largely compensated by incorporating age and Revised Trauma Score in the model. CONCLUSION: The ICISS methodology can be extended to ICD-10 horizon as a standard injury severity measure in the place of TRISS, especially when age and Revised Trauma Score were incorporated in the model. For patients with intracranial injuries, the predictive power of ICD-10-based ICISS was relatively low because of differences in the classifying system between ICD-10 and ICD-9CM. PMID- 10697088 TI - Severe contusion of the femoral vessels in rats alters tissue oxygenation and microvascular blood flow regulation in the skeletal muscles of the limb. AB - BACKGROUND: Severe contusion of an artery often presents clinical problems in that it affects flow distal to the injury. However, the effect of a contusion on the microvascular flow regulation in the distal part of the limb is still largely unknown. METHODS: A multipoint microelectrode technique was used to assess both tissue oxygenation (PtO2) and microflow (hydrogen clearance) on the skeletal muscle surface in a standard contusion injury to the femoral vessels in rats. RESULTS: A significant increase in and an altered distribution of (PtO2) as well as a reduction in and altered distribution of microflow on the muscle surface distal to the injury was found in all animals (n = 27) compared with the uninjured control leg. These findings could not be reproduced experimentally by sympathectomy or when the adjacent skeletal muscle alone was injured. CONCLUSION: The results suggest that the changes observed distal to the injury are of vascular origin, possibly as a result of endothelial damage at the site of the contusion. PMID- 10697089 TI - Arterial repair with synthetic patch by using titanium clips. AB - BACKGROUND: Vascular closure staple (VCS) clips made of titanium were originally developed for microvascular anastomoses. There is limited experience with their applicability to vascular reconstruction in larger vessels. This study compares VCS clips to standard sutures in arterial repair using a synthetic patch. METHODS: In an experimental study with pigs, two sequential 10-mm abdominal aortotomies were allocated randomly to synthetic patch (polytetrafluoroethylene) repair with VCS clips or continuous 6-0 polypropylene sutures. Angiographic, macroscopic, and microscopic results were assessed after 2 months. RESULTS: There were no significant differences in the patency rate, vessel diameter at the repair site, or healing indices. The mean (SD) clamp time was 8.7 (3.0) minutes for clip repair and 14.3 (7.4) minutes for suture repair (p = 0.04), and the times required for the vessel reconstruction were 5.3 (1.3) and 9.3 (3.0) minutes, respectively (p = 0.009). CONCLUSION: Patched arterial repair with VCS clips is faster than sutured reconstruction with comparable results after 2 months of follow up. PMID- 10697090 TI - Similarities between civilian gunshot wounds to the head and nongunshot head injuries. AB - BACKGROUND: This investigation compared the cerebral pathophysiologic status of gunshot wounds to the head (GSWH) with that of severe head injury of other causes (non-GSWH). METHODS: Data were collected prospectively from 71 GSWH and 541 non GSWH patients. The two groups had similar demographic characteristics and injury severities. Cerebral metabolic parameters for each patient were averaged for the entire period of monitoring. These per-patient averages were compared between GSWH and non-GSWH groups. RESULTS: Median intracranial pressure was 21.4 mm Hg in GSWH patients vs. 16.7 mm Hg in non-GSWH patients (p < 0.001). Mean arterial pressures were similar, but the higher intracranial pressure in GSWH patients produced a lower median cerebral perfusion pressure. Cerebral blood flow, cerebrovascular resistance, cerebral metabolic rate of oxygen, average jugular venous oxygen saturation, and number of jugular venous desaturations did not differ significantly between the groups. Three-month outcome was death in 43% of GSWH patients and 32% of non-GSWH patients, persistent vegetative state or severe disability in 33% and 32%, respectively, and moderate disability or good recovery in 24% and 36%, respectively. These outcomes were not significantly different (p = 0.11). CONCLUSION: GSWH patients suffer global cerebral metabolic disturbances that are at least as severe as those seen in non-GSWH patients with injuries of comparable severity. This selected population of GSWH patients may enjoy outcomes comparable to those of non-GSWH patients if they are treated by the same aggressive protocols. PMID- 10697091 TI - Preliminary experience with postmortem computed tomography in military penetrating trauma. AB - BACKGROUND: Postmortem examination serves as a tool for confirmation of clinical diagnosis, "quality" assurance, and education. In Israel, mostly because of religious reasons, most families withhold their permission to perform autopsies. To obtain objective information regarding the death of soldiers, the Israel Defense Forces Medical Corps started in September of 1997 to perform postmortem computed tomographic (PMCT) scans. The purpose of our study is to determine what information can be obtained from the PMCT scans. METHODS: In a period of 16 months, 27 soldiers were killed in low-intensity conflicts and PMCT was obtained in 22 cases. Medical data obtained from the field medical care providers were collected and compared with PMCT results. RESULTS: Several examples of patients whose death was determined at the scene either before any medical intervention or after initiation of resuscitative treatment are shown in our study and compared with the clinical impression of the treating physician. Two examples of autopsy results are compared with PMCT results. Gas was detected in various parts of the circulatory system in many cases. The significance of this finding, described in our study for the first time, needs further investigation. CONCLUSION: PMCT scanning has limits in detecting superficial injuries and injuries of the extremities and determining the exact route of fragments through body tissues in penetrating military trauma. It also cannot serve as a tool for examining ammunition or the protection provided by various armors. However, it can provide a substantial amount of evidence that, when reviewed with the clinical information obtained from the physician at the scene, can help in assessing the treatment given at the field and point toward the probable cause of death. PMID- 10697092 TI - Kiting injuries: report of two cases and discussion. PMID- 10697093 TI - Retained cardiac missile: an unusual case report. PMID- 10697094 TI - Evidence of bacterial translocation in fatal hemorrhagic pancreatitis. PMID- 10697095 TI - Fatal subclavian artery transection from isolated clavicle fracture. PMID- 10697096 TI - Painful nonunion of multiple rib fractures managed by operative stabilization. PMID- 10697097 TI - Transected subscapular artery in a transmediastinal gunshot wound presenting as a hemothorax: treatment with embolotherapy. PMID- 10697098 TI - Occlusion of an aberrant right hepatic artery, originating from the superior mesenteric artery, secondary to blunt trauma. PMID- 10697099 TI - Microvascular repair of jejunal and ileal vessels for near complete mesenteric avulsion after seat-belt injury. PMID- 10697100 TI - Drainage tube perforation of the stomach: an exceptionally rare complication. PMID- 10697101 TI - Bilateral adrenal hemorrhage in blunt abdominal trauma. PMID- 10697102 TI - Bilateral rupture of multicystic kidneys after blunt abdominal trauma. PMID- 10697103 TI - Indirect carotid-cavernous sinus fistula after shotgun injury. PMID- 10697104 TI - Basal fracture of the skull and lower (IX, X, XI, XII) cranial nerves palsy: four case reports including two fractures of the occipital condyle--a literature review. PMID- 10697105 TI - Combined midline-transverse surgical approach for severe blunt injuries to the right liver. AB - BACKGROUND: Although nonoperative treatment has been a major advance in the management of liver trauma, emergency surgery is still required for unstable patients. Severe hepatic lesions located in the right lobe, notably juxtahepatic venous injuries, are difficult to access and still carry a high mortality. METHODS: We describe a surgical approach for severe blunt injury to the right liver by a combined midline-transverse incision. This techniques allows simple, easy, and rapid mobilization and compression of the liver to control bleeding. RESULTS: This technique was used in 10 patients with blunt liver trauma, with grade III (n = 2), IV (n = 5), and V (n = 3) injuries. Mean intraoperative blood transfusion required was 21 units. Six patients underwent mandatory anatomic resection, three patients were treated by hepatic suture, and one patient was treated by packing. This patient developed brain death after surgery and was the only mortality. CONCLUSION: This technique is efficient and less cumbersome than shunting approaches. PMID- 10697106 TI - Communicating with the family: the risks of medical radiation to conceptuses in victims of major blunt-force torso trauma. AB - BACKGROUND: Trauma surgeons must balance the risk and benefits of diagnostic radiographic procedures on potentially pregnant patients and should know the range and likelihood of effects that radiation might have on pregnancy. METHODS: We present guidelines for assessing such radiation risks. Knowledge of a patient's pregnancy status and an estimate of radiation dose to the conceptus (low, < 10 mGy [milligray]; intermediate, 10-250 mGy; high, > 250 mGy) allow provisional assessment of radiation-induced risks. RESULTS: Dose estimates may be estimated at 2 mGy per exposure (radiographs), 5 mGy per slice (computed tomography), and 10 mGy per minute of fluoroscopy, when the conceptus is within the x-ray field. A formal radiation exposure assessment is appropriate when provisional estimates exceed 10 mGy. CONCLUSION: A simple qualitative dose assessment can inform clinical decisions and guide appropriate triage to more formal quantitative assessment. PMID- 10697107 TI - The search for an optimal end point of resuscitation. PMID- 10697108 TI - Race is of very limited importance in determining the outcome after trauma. PMID- 10697109 TI - In vitro elution of antibiotic from antibiotic-impregnated biodegradable calcium alginate wound dressing. PMID- 10697110 TI - The evaluation of the usefulness of alternate devices for difficult and/or emergency intubation. PMID- 10697111 TI - Optimal prosthesis for acute replacement of the abdominal wall. PMID- 10697112 TI - Engineering cells for glucose-sensitive production of insulin: toward genetic reconstruction. PMID- 10697113 TI - Deletion of the adenoviral E1b-19kD gene enhances tumor cell killing of a replicating adenoviral vector. AB - Replicating adenoviral vectors are a promising new modality for cancer treatment and clinical trials with such vectors are ongoing. Targeting these vectors to cancer cells has been the focus of research. However, even if perfect targeting were to be achieved, a vector still must effectively kill cancer cells and spread throughout the bulk of the tumor. The adenoviral E1b-19kD protein is a potent inhibitor of apoptosis and may therefore compromise the therapeutic efficacy of an adenoviral vector. In this study we have investigated if an E1b-19kD gene deletion could improve the ability of a replicating adenoviral vector to spread through and kill cancer cells. In several lung cancer cell lines an E1b-19kD deleted virus (Ad337) induced substantially more apoptosis than did a wild-type virus (Ad309), and tumor cell survival was significantly reduced in three of four cell lines. In addition, the apoptotic effects of cisplatin or paclitaxel were augmented by Ad337, but inhibited by wild-type virus. The number of infectious virus particles in the supernatant of infected cells was increased with Ad337 compared with wild-type virus, indicating enhanced early viral release. Ad337, in contrast to Ad309, induced significantly larger plaques after infection of A549 cells. This well-described large plaque phenotype of an E1b-19kD mutant virus is likely the result of early viral release and enhanced cell-to-cell viral spread. Loss of E1b-19kD function caused only minor cell line-specific increase or decrease in viral yield. We conclude that deletion of the E1b-19kD gene may enhance the tumoricidal effects of a replicating adenoviral vector. PMID- 10697114 TI - Antiviral activity of an intracellularly expressed single-chain antibody fragment directed against the murine leukemia virus capsid protein. AB - We have addressed the possibility that intracellularly expressed miniantibodies directed against the viral capsid protein can be used as antiretroviral agents in gene transfer experiments. R187 is a rat monoclonal antibody that has been reported to recognize the MuLV p30gag capsid polypeptide. We report here that it also binds to the Pr65gag precursor polyprotein. R187 has been cloned and expressed in the form of a single-chain variable fragment (scFv) that shows the same binding specificity as the parental antibody. When expressed intracellularly, the R187 scFv favors the production of viral particles showing reduced infectivity. It, however, exerts no detectable protective effect against infection. This was observed both when using replication-incompetent MuLV-derived vector and replication-competent wild-type MuLV. Although the intimate mechanism of the inhibition is not clear, this work raises the possibility that gene engineering of anti-capsid protein scFvs may offer an additional lead for gene therapy of severe retrovirus-linked diseases. PMID- 10697115 TI - Engineering of a glucose-responsive surrogate cell for insulin replacement therapy of experimental insulin-dependent diabetes. AB - Glucose responsiveness in the millimolar concentration range is a crucial requirement of a surrogate pancreatic beta cell for insulin replacement therapy of insulin-dependent diabetes. Novel insulin-secreting GK cell clones with millimolar glucose responsiveness were generated from an early-passage glucose unresponsive RINm5F cell line. This line expressed constitutively both the K(ATP) channel and the GLUT2 glucose transporter; but it had a relative lack of glucokinase. Through overexpression of glucokinase, however, it was possible to generate glucose-responsive clones with a glucokinase-to-hexokinase ratio comparable to that of a normal pancreatic beta cell. This aim, on the other hand, was not achieved through overexpression of the GLUT2 glucose transporter. Raising the expression level of this glucose transporter into the range of rat liver, without correcting the glucokinase-to-hexokinase enzyme ratio, did not render the cells glucose responsive. These glucokinase-overexpressing RINm5F cells also stably maintained their molecular and insulin secretory characteristics in vivo. After implantation into streptozotocin diabetic immunodeficient rats, glucokinase overexpressing cells retained their insulin responsiveness to physiological glucose stimulation under in vivo conditions. These cells represent a notable step toward the future bioengineering of a surrogate beta cell for insulin replacement therapy in insulin-dependent diabetes mellitus. PMID- 10697116 TI - Modulation of the inflammatory properties and hepatotoxicity of recombinant adenovirus vectors by the viral E4 gene products. AB - Liver toxicity and inflammation were assessed in C57BL/6, CBA, and BALB/c mice injected intravenously with a series of recombinant adenoviruses deleted simultaneously in E1/E3, in E1/E3/E2A, or in E1/E3/E4. All vectors were either devoid of transgenes or carried in E1 the human CFTR cDNA under the control of the CMV promoter. Injection of the E1/E3-deleted vector induced a significant liver dystrophy and inflammatory responses that were accompanied by an increased serum transaminase concentration. The vector toxicity remained elevated on additional deletion of the E2A gene and was further enhanced when hCFTR was expressed. In contrast, additional deletion of E4 led to a reduction in hepatotoxicity, suggesting an active role of E4 gene products in liver injury. However, deletion of E4 also led to a loss of transgene expression. To identify the individual E4 product(s) involved in liver toxicity and in the regulation of transgene expression, a series of isogenic E1/E3-deleted vectors, with or without the hCFTR transgene, and containing various combinations of functional E4 open reading frames (ORFs), were evaluated in vitro and in vivo. We demonstrate that liver injury was markedly reduced with vectors containing either ORF3 alone or ORF3,4 while vectors containing ORF4, ORF6,7 or ORF3,6,7 still displayed elevated hepatotoxicity and inflammatory responses. Moreover, transgene expression was restored when ORF3,4 or ORF3,6,7 was retained in the vector. These results highlight the importance of the E4 gene products in the design of improved in vivo gene transfer vectors. PMID- 10697117 TI - Continuous erythropoietin delivery by muscle-targeted gene transfer using in vivo electroporation. AB - It has been demonstrated that gene transfer by in vivo electroporation of mouse muscle increases the level of gene expression by more than 100-fold over simple plasmid DNA injection. We tested continuous rat erythropoietin (Epo) delivery by this method in normal rats, using plasmid DNA expressing rat Epo (pCAGGS-Epo) as the vector. A pair of electrodes was inserted into the thigh muscles of rat hind limbs and 100 microg of pCAGGS-Epo was injected between the electrodes. Eight 100 V, 50-msec electric pulses were delivered through the electrodes. Each rat was injected with a total of 400 microg of pCAGGS-Epo, which was delivered to the medial and lateral sides of each thigh. The presence of vector-derived Epo mRNA at the DNA injection site was confirmed by RT-PCR. The serum Epo levels peaked at 122.2 +/- 33.0 mU/ml on day 7 and gradually decreased to 35.9 +/- 18.2 mU/ml on day 32. The hematocrit levels increased continuously, from the preinjection level of 49.5 +/- 1.1 to 67.8 +/- 2.2% on day 32 (p < 0.001). In pCAGGS-Epo treated rats, endogenous Epo secretion was downregulated on day 32. In a control experiment, intramuscular injection of pCAGGS-Epo without subsequent electroporation did not significantly enhance the serum Epo levels. These results demonstrate that muscle-targeted pCAGGS-Epo transfer by in vivo electroporation is a useful procedure for the continuous delivery of Epo. PMID- 10697118 TI - Development of a self-inactivating, minimal lentivirus vector based on simian immunodeficiency virus. AB - In contrast to oncoviruses, lentiviruses do not require target cell division for integration into the host genome. Lentiviral vectors can therefore expand the spectrum of target cells susceptible to retroviral gene transfer. To analyze whether vectors based on simian immunodeficiency viruses (SIVs) could be used for gene transfer, a three-plasmid vector-packaging system was developed, in which Gag-Pol and the vector itself are of SIV origin, while Env is derived either from SIV, amphotropic murine leukemia virus (MuLV), or the G glycoprotein of vesicular stomatitis virus (VSV-G). To increase the safety of the SIV vector system, a self inactivating SIV vector was constructed. After optimization of the SIV gag-pol expression plasmid, a minimal SIV vector, which contained only SIV sequences present on the multiply spliced nef transcript, could still be produced at titers of 2 x 10(5) infectious units/ml. Growth-arrested cells could be transduced with this vector even if vif, vpr, vpx, and nef had been deleted from the packaging construct and the vector. PMID- 10697119 TI - Additional transduction events after subretinal readministration of recombinant adeno-associated virus. AB - Subretinal delivery of recombinant adeno-associated virus (rAAV) results in a systemic humoral response in the adult immunocompetent mouse. We characterized this response and determined whether it is possible to readminister rAAV to the subretinal space despite the presence of antibodies to the vector. A systemic humoral response to rAAV capsid proteins was induced by either unilateral subretinal injection or by intradermal administration of 1 x 10(9) infectious units of rAAV carrying the cDNA encoding green fluorescent protein (GFP), rAAV GFP. Experiments were performed in cohorts of adult C57BL/6 mice. Assessment of systemic humoral response to viral capsid protein was performed through enzyme linked immunoabsorbent assay (ELISA) and infection inhibition studies of serum samples 3 weeks after virus delivery. The rAAV-GFP virus was readministered by subretinal injection. GFP expression after subretinal administration was evaluated ophthalmoscopically throughout the course of the experiment and histologically at the termination of the experiment. We observed significant systemic humoral responses to viral capsid protein after subretinal delivery of rAAV. Intradermal injection resulted in a larger humoral response (with a higher percentage of neutralizing antibodies) than subretinal injection. Additional transduction events were observed after readministration of rAAV despite the presence of strong humoral response to the vector. PMID- 10697120 TI - Effective use of donor MHC class I gene therapy in organ transplantation: prevention of antibody-mediated hyperacute heart allograft rejection in highly sensitized rat recipients. AB - Immunologically sensitized recipients present one of the most critical problems in clinical organ transplantation today, since preformed antibodies rapidly destroy donor tissue expressing specific MHC class I antigens (Ag). Therefore, sensitized patients are either unable to receive a compatible organ, or experience a prolonged waiting period. In this study we examined the effectiveness of donor MHC class I gene therapy in preventing hyperacute rejection (HR) of rat heart allografts in passively sensitized recipients. Our gene therapy strategy to address this problem is based on the phenomenon that liver transplants, which resist antibody-mediated HR, produce soluble MHC class I Ag capable of neutralizing preformed antibodies and suppressing the immune response. To mimic this "liver effect," we used liposomes to transfect cultured recipient (Lewis-RT1.Al) hepatocytes with plasmid DNA encoding the soluble donor MHC class I Ag, RT1.Aa. Control or RT1.Aa-transfected hepatocytes were implanted intrasplenically into Lewis recipients 1 day prior to heterotopic ACI (RT1.Aa) heart transplantation and injection of 6 ml of anti-ACI hyperimmune serum (HIS). Results showed that nearly all recipients receiving ACI-specific HIS and control hepatocytes experienced HR, while none of the recipients receiving HIS and hepatocytes expressing soluble RT1.Aa developed HR. Furthermore, active immunosuppression by soluble RT1.Aa was evidenced by prolongation of allograft survival, compared with controls not receiving HIS. In summary, soluble donor-MHC class I Ag gene therapy can prevent antibody-mediated destruction associated with HR. Future development of a similar strategy in humans may significantly improve the results of clinical organ transplantation in immunologically sensitized recipients. PMID- 10697121 TI - Enhancement of transgene expression by cotransfection of oriP plasmid with EBNA-1 expression vector. AB - We have attempted to develop a system for specific enhancement of transgene expression, which has been one of the most important issues in human gene therapy. When an Epstein-Barr virus (EBV) nuclear antigen 1 (EBNA-1) expression vector, pCMV-trEBNA-1, was cotransfected with an origin of latent viral DNA replication (oriP)-harboring plasmid, poriP-CMV-luciferase, luciferase gene expression was up to 20 times greater than in the absence of EBNA-1. This enhancement was regulated mainly at the transcriptional level and was dependent on the oriP sequence and the amount of EBNA-1. However, cointroduction of poriP CMV-luciferase with purified recombinant EBNA-1 inhibited luciferase gene expression whereas no inhibition was observed when pCMV-luciferase was cointroduced with recombinant EBNA-1. We also introduced poriP-CMV-luciferase into mouse liver via the use of HVJ (hemagglutinating virus of Japan)-liposomes. By 10 days after transfer, luciferase gene expression was decreased to low levels. We then introduced pCMV-trEBNA-1 to mouse liver via HVJ-liposomes on day 10. Luciferase gene expression was reactivated, whereas no reactivation was detected by the injection of EBNA-1 expression plasmid into liver injected with pCMV-luciferase lacking the oriP sequence. Thus, cotransfection of oriP-harboring expression vector with EBNA-1 expression plasmid should be promising for human gene therapy, although the safety of the system must be investigated thoroughly. PMID- 10697122 TI - Liver sinusoidal endothelial cells are not permissive for adenovirus type 5. AB - Adenoviral vectors are known to transduce hepatocytes in normal liver tissue with high efficiency. The aim of this study was to investigate whether sinusoidal endothelial cells, which separate hepatocytes from the bloodstream in the sinusoidal lumen, are permissive for infection by adenoviruses. We show here that microvascular liver sinusoidal endothelial cells are not infected by adenovirus type 5 in vivo or in vitro unless high MOIs are used. In contrast, macrovascular endothelial cells from aorta are efficiently infected by adenovirus type 5. In addition, Kupffer cells, similar to sinusoidal endothelial cells, are not infected by adenovirus type 5. Liver sinusoidal endothelial cells do not express the integrin receptor alpha(v)beta3, which is required for efficient infection by adenoviruses. Our results demonstrate that hepatocytes are the main cell population of the liver that is infected by adenovirus type 5. PMID- 10697123 TI - A phase I study of intralesional administration of an adenovirus vector expressing the HSV-1 thymidine kinase gene (AdV.RSV-TK) in combination with escalating doses of ganciclovir in patients with cutaneous metastatic malignant melanoma. PMID- 10697124 TI - Is it time for separate pediatric hepatology and liver transplant training programs and special certification or qualification? PMID- 10697125 TI - Is there an infectious etiology to abdominal pain in children? PMID- 10697126 TI - Secretin: cure or snake oil for autism in the new millennium? (response) PMID- 10697127 TI - Secretin: real therapeutic potential (response) PMID- 10697128 TI - Nutritional implications of replacing bovine milk fat with vegetable oil in infant formulas. PMID- 10697129 TI - Are all human milks created equal? Variation in human milk oligosaccharides. PMID- 10697130 TI - EGG like EKG: are we there yet? PMID- 10697131 TI - Use of intrahepatic chemotherapy to treat advanced pediatric hepatic malignancies. AB - BACKGROUND: To evaluate the effect of intrahepatic arterial chemotherapy (IAC) on children with primary hepatic malignancies. METHOD: A nonrandomized inception cohort of 11 pediatric patients was referred for treatment of advanced primary hepatic malignancies at Children's Hospital of Pittsburgh. None of the patients was a candidate for resection before the initiation of IAC. Tumor response to treatment was observed by determining serum alpha-fetoprotein (AFP) levels and by abdominal computed tomographic scan. The patients received hepatic artery infusions of cisplatin and/or doxorubicin. The last five also received gelfoam embolization. RESULTS: Eight of 11 patients had multiple IAC treatments. Eight patients had AFP-producing tumors, and five of the eight had dramatic reductions in serum levels after IAC treatment. Five of the 11 patients underwent successful orthotopic liver transplantation after receiving IAC therapy, and the five explanted specimens showed varying degrees of tumor necrosis. One-year survival in patients in the authors' center is 67% for those with hepatoblastoma and 40% for those with hepatocellular carcinoma. Three-year survival is 60% and 30% for patients with hepatoblastoma and hepatocellular carcinoma, respectively. CONCLUSION: Intrahepatic arterial chemotherapy therapy can halt the progression and possibly down-stage advanced pediatric hepatic malignancies. This therapy can also be used as a successful adjunct in altering a patient's chance for successful liver transplantation. PMID- 10697132 TI - Imaging changes in the pancreas in cystic fibrosis: a retrospective evaluation of 55 cases seen over a period of 9 years. AB - BACKGROUND: Pathologic changes of the pancreas have been observed as early as the recognition of the disease termed initially "cystic fibrosis of the pancreas". Atrophy of the gland and its fatty infiltration were considered as usual features. The aim of this study was to follow-up the evolution of cystic fibrosis pancreas and to define its successive stages in correlation with the clinical, biochemical, and imaging findings. METHODS: Fifty-five patients were followed up during 9 years. The patients' genetic backgrounds were systematically performed. Blood lipase levels were analyzed systematically at each consultation of the patients and in the event of bouts of abdominal pains. Imaging using mainly echograms and tomodensitometric scans were regularly performed: echograms every 6 months, and tomodensitometric scans every 1 to 2 years. Magnetic resonance imaging was performed in four patients. RESULTS: Five groups of patients were identified on the basis of tomodensitometric scan findings: normal pancreas (n = 4), incomplete lipomatosis of the pancreas (n = 9), complete lipomatosis of the pancreas (n = 23), cystic pancreas (n = 5), macrocystic pancreas (n = 1), atrophic pancreas (n = 13). Pancreas exocrine function was not correlated with findings. Forty episodes of pancreatitis were observed in seven patients. They had bouts of abdominal pain and elevation of lipase levels. Five of these patients were composite heterozygotes (D508/other). Incomplete lipomatosis represents an intermediate stage leading toward complete lipomatosis or toward atrophy after pancreatitis. CONCLUSIONS: Studies of pancreatic function should be performed routinely in cystic fibrosis, especially in pancreatic sufficiency or in patients with normal pancreas images. Acute pancreatitis should be diagnosed and properly identified to be differentiated from other acute abdominal syndromes occurring in cystic fibrosis. PMID- 10697134 TI - Impaired accommodation of the proximal stomach in children with recurrent abdominal pain. AB - BACKGROUND: A new ultrasonographic method was applied in children with recurrent abdominal pain, to study accommodation of the proximal stomach to a meal. METHODS: After an overnight fast, 20 patients with recurrent abdominal pain (age, 7-14 years) and 23 healthy control subjects (age, 7-13 years), were scanned by a 5-MHz transducer positioned in the epigastrium, to monitor the size of the proximal stomach before and after a test meal of meat soup. RESULTS: Children with recurrent abdominal pain had a significantly smaller sagittal area of the proximal stomach at 10 and 20 minutes after the meal than in healthy control subjects (P = 0.01 for both) and significantly higher emptying fraction of the proximal stomach at 10 minutes after the meal than in healthy control subjects (P = 0.02). There was no significant difference in emptying of the distal stomach between the patients and healthy control subjects. Children with recurrent abdominal pain experienced more symptoms (pain, bloating) in response to the test meal than did healthy control subjects. CONCLUSION: The results support the view that recurrent abdominal pain in children may be a motility disorder that can be detected in the proximal stomach as an impairment of adaptive relaxation in response to a meal. This new ultrasonographic method may become a valuable diagnostic tool in patients with recurrent abdominal pain. PMID- 10697133 TI - Long-term outcome after partial external biliary diversion for intractable pruritus in patients with intrahepatic cholestasis. AB - BACKGROUND: Chronic intrahepatic cholestasis is associated with severe pruritus that is often refractory to maximal medical management and leads to significantly impaired quality of life. The hypothesis in this study was that partial external biliary diversion (PEBD) can substantially improve intractable pruritus secondary to intrahepatic cholestasis with subsequent improvement of functional quality of life. METHODS: Parents' and/or patients' clinical rating of pruritus, growth percentiles, biochemical parameters, and liver biopsies performed before and after surgery were compared in a retrospective medical record review. RESULTS: Eight children underwent PEBD from 1990 through 1997. Complete follow-up data were available for seven patients. Before surgery, all patients had intense pruritus, which was not responsive to maximal medical therapy. Specimens obtained in preoperative liver biopsies showed moderate (n = 1), minimal (n = 6), or no (n = 1) portal fibrosis. After PEBD, all patients received ursodeoxycholic acid (10 15 mg/kg/dose two to three times daily) until resolution of pruritus. Of the seven patients with complete follow-up data, six had complete resolution of pruritus and sustained resolution up to 8 years after surgery. The patient with mild to moderate residual pruritus was the youngest to undergo PEBD. Growth improved from below the 5th percentile before surgery to the 5th through the 25th percentiles for five of six patients with more than 6 years' follow-up. All families reported improved quality of life, defined by school attendance and ability to resume normal activity with peers. There has been no clinical evidence of progression of liver disease. CONCLUSION: Partial external biliary diversion is effective in the long-term treatment of pruritus refractory to medical therapy and provides a favorable outcome in a select group of patients with chronic intrahepatic cholestasis without cirrhosis. PMID- 10697135 TI - Toxaphenes and chlorinated naphthalenes in adipose tissue of children. AB - BACKGROUND: Chlorinated hydrocarbons are ingested by humans in food and accumulate in adipose tissue. At the University Kinderklinik, Mannheim, previously unknown substances have been found in children (e.g., the pesticide toxaphene and chlorinated naphthalenes). These substances have been widely used for industrial purposes in the past. Samples from West and East Germany; Saratov, Russia; and Almaty, Kazakhstan were examined to determine whether these substances are ubiquitous. METHODS: After Soxhlet extraction, the extracts were cleaned up using a liquid chromatographic technique. Measurement was performed by gas chromatography-mass spectrometry using negative chemical ionization in the single-ion-monitoring mode. RESULT: In specimens from all cities, toxaphene congeners Parlar 26 and Parlar 50 and six chlorinated naphthalenes were traced. Highest median load of toxaphene was 1.97 microg/kg for Parlar 26 and 2.36 microg/kg for Parlar 50 in Stralsund, East Germany. For chlorinated naphthalenes, the median was highest in Mannheim, West Germany, with 12.0 microg/kg. CONCLUSION: These findings show that monitoring these toxic substances remains necessary. Even though the use and as a consequence the amount of chlorinated hydrocarbons were reduced, these substances have by no means disappeared from the environment. PMID- 10697136 TI - Magnetic resonance imaging to distinguish the type and severity of pediatric inflammatory bowel diseases. AB - BACKGROUND: The distinction between ulcerative colitis and Crohn's disease is important, because treatment options and clinical course may vary. Magnetic resonance imaging (MRI) allows noninvasive transmural assessment of the intestine and may facilitate differentiation of ulcerative colitis from Crohn's disease. The objective of this prospective study was to determine whether MRI differentiates Crohn's disease from ulcerative colitis in children as effectively as colonoscopy with mucosal biopsies. METHODS: Fifteen patients underwent colonoscopy with biopsies followed by abdominal MRI. The MRI diagnosis, determined by two radiologists independently completing a standardized form was compared with the gastroenterologic diagnosis. RESULTS: After colonoscopy and review of histology, Crohn's disease was diagnosed in nine patients, ulcerative colitis in five, and indeterminate colitis in one, who was excluded from study. Agreement of the MRI diagnosis with the gastroenterologic diagnosis was 4 of 4 (100%) for ulcerative colitis, 4 of 10 (40%) for Crohn's disease considering both radiologists, and 5 of 10 (50%) for Crohn's disease for each radiologist individually. Percentage of enhancement by MRI did not correlate with the severity of inflammation determined at endoscopy among the patients with Crohn's disease (r = -0.3, P = 0.366). There was agreement on severity of inflammation in three of four patients with ulcerative colitis. CONCLUSIONS: Current MRI interpretation of inflammatory bowel disease did not adequately recognize Crohn's disease in children. Therefore, colonoscopy with biopsy remains the most accurate tool for determining the type and severity of inflammatory bowel disease in children and adolescents. PMID- 10697137 TI - Transpyloric enteral nutrition reduces the complication rate and cost in the critically ill child. AB - BACKGROUND: Studies in adults have shown that transpyloric enteral nutrition (TEN) is useful in certain patients who cannot tolerate oral or gastric feeding. This study was conducted to compare TEN with parenteral nutrition (PN) in critically ill pediatric patients. METHODS: A retrospective descriptive study conducted in the pediatric intensive care unit of a tertiary pediatric referral center. All patients in the pediatric intensive care unit (PICU) receiving PN and/or TEN from January 1993 through December 1996 were included in the study. RESULTS: Two hundred forty patients (14.6% of all patients admitted to the PICU) received PN and/or TEN (168 exclusively PN, 21 exclusively TEN, and 51 a combined regimen). The number of patients receiving PN and duration of PN declined significantly from 1993 (65 patients, 703 days) through 1996 (48 patients, 395 days). This was mirrored by the increase in the number of patients receiving TEN and duration of TEN. The incidence of complications (hyperglycemia, hypertriglyceridemia, and cholestasis) was higher in the PN group. There was no difference in the incidence of hospital-acquired infection or mortality between the two groups. The cost of TEN was lower than that of PN, with an estimated annual saving of $5,422. CONCLUSIONS: Transpyloric enteral nutrition is a suitable method of nutritional support for critically ill pediatric patients. It has fewer complications and a lower cost than PN. PMID- 10697138 TI - Variability of human milk neutral oligosaccharides in a diverse population. AB - BACKGROUND: A complex array of free oligosaccharides is a distinctive compositional feature of human milk. Although these oligosaccharides have been studied for several years, their variability and distribution have not been systematically studied, and their nutritional and functional roles have not been elucidated. This report describes a study in which a large number of human milk samples were analyzed for the presence and content of nine neutral oligosaccharides. The resultant data were used to probe for distribution trends by donor groups and stage of lactation. METHODS: Milk samples from 435 women residing in 10 countries were analyzed using a simple preparation procedure, gel filtration, and high-performance anion-exchange chromatography. RESULTS: All samples contained structures based on lacto-N-neotetraose and lacto-N-tetraose. This contrasts with the fucosyloligosaccharides tested, none of which was detected in 100% of the samples. Unexpected distribution trends were observed. For example, 100% of the samples from Mexico (n = 156) contained 2' fucosyllactose, whereas only 46% of the samples from the Philippines (n = 22) contained this structure. Concentration ranges for the analyzed oligosaccharides revealed quantitative and qualitative distribution trends. CONCLUSIONS: The oligosaccharide composition of human milk varied among samples. The geographical origin of the donors was one of the factors that accounted for this variability. This can be explained by genetically determined traits that are not uniformly distributed. Results indicated that further systematic studies are needed to ascertain the effect of other factors, such as lactation stage or diet. PMID- 10697139 TI - Electrogastrography versus gastric emptying scintigraphy in children with symptoms suggestive of gastric motility disorders. AB - BACKGROUND: Cutaneous electrogastrography is a method of recording gastric electrical activity. Abnormalities of the electrogastrogram have been described in a variety of disorders. The purpose of the study was to correlate the electrogastrograms of children with vomiting and dyspepsia with the results of radionucleotide gastric emptying studies. METHODS: Nine patients (5-16 years old) with gastrointestinal symptoms of vomiting and/or abdominal pain were studied. The electrogastrogram was recorded using surface electrodes for 30 minutes in the fasting state and for 120 minutes after a radioisotope-labeled solid meal. Gastric emptying was simultaneously monitored for 120 minutes. The postprandial change in dominant power (power ratio: postprandial/fasting dominant power), percentages of normal slow wave, bradygastria, and tachygastria were recorded and analyzed. RESULTS: The patients were divided into two groups. The first group (four patients; five studies) had normal gastric emptying, whereas the second group (five patients) had delayed emptying (half-life, >90 minutes). The median power ratio in the first group was 1.69 and in the second group was 2.78; the difference was not statistically significant (P = 0.90). The median difference in slow wave percentages in the fasting and postprandial periods was 0.99 in the first group and 0.73 in the second group; again, the difference was not statistically significant (P = 0.27). CONCLUSIONS: Although it is a method of assessing gastric myoelectrical activity and gastric motility disorders, electrogastrogram does not correlate with nuclear scintigraphic gastric emptying studies in children. PMID- 10697140 TI - Efficacy of bismuth-based triple therapy in children with abdominal pain and Helicobacter pylori gastritis. AB - BACKGROUND: To evaluate the effect of a therapeutic regimen of 7 days versus 14 days on the clinical manifestations of Helicobacter pylori gastritis in children. METHODS: Ninety children (age 2-19 years) who had abdominal pain and/or recurrent vomiting were determined to have H. pylori gastritis by endoscopy, histology, and a Giemsa stain positive for H. pylori. The patients were randomized to receive amoxicillin, metronidazole, and bismuth subcitrate for 7 days (group A; 45 children) or 14 days (group B; 45 children) and were observed clinically for 19 +/- 11.5 months. Resolution of all abdominal and gastrointestinal symptoms was considered a good response. RESULTS: A good response was obtained in 36 (80%) children from group A, and in 37 (82%) from group B. A recurrence of symptoms occurred in four (11%) of the responders from group A, and in six (15.2%) from group B. CONCLUSIONS: A 7-day course of bismuth-based triple therapy for H. pylori gastritis in children appears to be clinically as effective as a 14-day regimen. The feasibility of a shorter therapeutic regimen may enhance patient compliance and provide a better chance of clinical benefit. PMID- 10697141 TI - Long-term outcome of chronic hepatitis B in adolescents or young adults in follow up from childhood. AB - BACKGROUND: It has not yet been defined whether children with chronic hepatitis B are likely to develop severe liver disease in the future. The purpose of this study was to evaluate the evolution of chronic hepatitis B acquired in childhood. METHOD: Fifty-two children in the age range of 0 to 15 years who were positive for hepatitis B surface antigen and hepatitis B e antigen in serum for at least 6 months were enrolled in this study. In the majority of the 52 children, hepatitis B virus infection was acquired by perinatal transmission. All 52 showed abnormal liver function test findings for more than 6 months before enrollment, and the subjects were followed up longitudinally for 3 to 22 years (mean, 11 years). They are now more than 15 years of age (15-27 years old). RESULTS: During the follow up period, 26 (50%) children had spontaneous seroconversion to anti-hepatitis B e. Serum levels of alanine aminotransferase normalized in these 26 children. In one child of these children, hepatocellular carcinoma developed at the age of 21 years, 16 years after seroconversion, although his liver function profiles remained normal. The other 26 children remained hepatitis B e antigen positive, most with unchanged biochemical features. Sixteen (62%) children among these 26 children were treated with interferon-alpha. Eleven (69%) children had seroconversion to anti-hepatitis B e within the first year after the cessation of therapy. Hepatocellular carcinoma developed in 1 of these 11 children at the age of 16 years, 6 years after interferon therapy. Thus, hepatocellular carcinoma developed in two children in an anti-hepatitis B e positive phase. CONCLUSION: All children carrying hepatitis B surface antigen should be observed carefully to monitor the possible development of hepatocellular carcinoma, especially in the antihepatitis B e-positive phase after spontaneous seroconversion or even after interferon treatment. PMID- 10697142 TI - Helicobacter pylori infection in children: a consensus statement. European Paediatric Task Force on Helicobacter pylori. PMID- 10697143 TI - Why don't we bud? PMID- 10697144 TI - Could oligosaccharide supplementation promote gut colonization with a beneficial flora in preterm infants? PMID- 10697145 TI - Extrapulmonary IMT generally occur in children and young adults. PMID- 10697146 TI - A fatal small dose of phosphate enema in a young child with no renal or gastrointestinal abnormality. PMID- 10697147 TI - Tissue transglutaminase: does the key fit the celiac lock? PMID- 10697148 TI - Constipation and intolerance to cow's milk. PMID- 10697149 TI - Quality of life of patients with chronic hepatitis C virus infection. PMID- 10697150 TI - Maternal cigarette smoking during pregnancy and the risk of having a child with cleft lip/palate. AB - Maternal cigarette smoking during pregnancy as a risk factor for having a child with cleft lip/palate has been suggested by several epidemiologic studies. However, most of these studies contained small sample sizes, and a clear association between these two factors could not be established. The U.S. Natality database from 1996 and a case-control study design were used to investigate the association between maternal smoking during pregnancy and having a child with cleft lip/palate. The records of 3,891,494 live births from the 1996 U.S. Natality database were extracted to obtain cleft lip/palate cases and random controls. The National Center for Health Statistics collects maternal and newborn demographic and medical data from the birth certificates of all 50 states. New York (excluding New York City), California, Indiana, and South Dakota did not collect smoking data, and the data from these states were excluded from the analysis. A total of 2207 live births with cleft lip/palate cases were identified, and 4414 controls (1:2 ratio) were randomly selected (using the SAS program) from live births with no congenital defects. Odds ratios and 95 percent confidence intervals were determined from logistic regression models, adjusting for confounding variables, including maternal demographic and medical risk factors. A significant association was found between any amount of maternal cigarette use during pregnancy and having a child with cleft lip/palate [unadjusted odds ratio 1.55 (1.36, 1.76), p < 0.001]. Univariate analysis showed that maternal education level, age, race, and maternal medical conditions (diabetes and pregnancy-associated hypertension) were potential confounders. After adjusting for these confounders, the odds ratio remained significant [Mantel-Haenszel odds ratio 1.34 (1.16, 1.54), p < 0.001]. To determine the dose response of cigarette smoking during pregnancy, the cigarette consumption per day was divided into four groups: none, 1 to 10, 11 to 20, and 21 or more. A dose response relationship was found when comparing each smoking category with the no smoking reference group: 1.50 (1.28, 1.76), 1.55 (1.23, 1.95), and 1.78 (1.22, 2.59), respectively. This means that increased cigarette smoking during pregnancy resulted in increased odds of having a child with cleft lip/palate. This is the largest study to date to test the association between maternal cigarette smoking during pregnancy and having a newborn with cleft lip/palate. The significant trend in the dose-response relationship strongly suggests the association of smoking tobacco and this common congenital deformity. These results emphasize the public health risks associated with smoking during pregnancy. To prevent this devastating craniofacial anomaly, educational initiatives should be considered that will alert expectant mothers to the association between smoking during pregnancy and the occurrence of cleft lip/palate. PMID- 10697151 TI - Progression of facial asymmetry in hemifacial microsomia. AB - Hemifacial microsomia is a common craniofacial anomaly, variably affecting structures derived from the first and second pharyngeal arches. Correction of the skeletal deformity in children has been advocated to improve growth potential and reduce secondary deformity. However, contrary reports have suggested that facial asymmetry in hemifacial microsomia does not increase with growth; therefore, skeletal correction can be postponed, even until adolescence. The purpose of this study was to test the hypothesis that facial asymmetry in hemifacial microsomia is progressive. This is a retrospective evaluation of 67 patients with untreated hemifacial microsomia. The patients were categorized as: group I (mandible type I, IIa), n = 38, and group II (mandible type IIb, III), n = 29. Pretreatment posterior-anterior cephalometric radiographs were used to analyze asymmetry by measuring the angle between the true horizontal and the following planes: piriform rim, maxillary occlusal plane, and intergonial angle. Angular measurements were averaged for patients in the deciduous (<6 years), mixed (> or =6<13 years), and permanent dentition (> or =13 years). In group I, angle piriform rim, maxillary occlusal plane, and intergonial angle increased from 7.0, 4.3, and 4.4 to 8.4, 6.6, and 6.1 degrees, respectively [mean age, 4.1 (deciduous) to 8.6 (mixed) to 21.0 (permanent) years]. In group II, angle piriform rim, maxillary occlusal plane, and intergonial angle increased from 9.5, 6.2, and 5.3 to 11.7, 7.6, and 8.0 degrees, respectively [mean age, 3.4 (deciduous) to 8.0 (mixed) years]. These data demonstrate that hemifacial microsomia is progressive and underscores the importance of early surgical correction of mandibular asymmetry in this disorder. PMID- 10697152 TI - Cleft lip nose correction with onlay calvarial bone graft and suture suspension in Oriental patients. AB - To correct the secondary cleft lip nose deformity in Oriental patients, many alar cartilage mobilization and suspension techniques have been developed. However, these techniques have critical limitations. One of the limitations is the suspension vector, and another is suspension power. The suspension vector is from inferior to superior and from the deformed alar cartilage to the normal alar cartilage. Thus, the vector is not suitable for normal nasal tip projection. The suspension power is not satisfactory because Oriental people have underdeveloped, thin alar cartilages and thick skin. So, the suspended, deformed alar cartilage may relapse and pull the normal alar cartilage to the deformed side. To overcome these limitations, the authors use the cantilever calvarial bone graft for tip projection; it also serves as a strong, rigid framework for cartilage and soft tissue suspension. Using these techniques, the authors can create normal nasal tip projection and a normal looking nasal aperture. PMID- 10697153 TI - Otoplasty: anterior scoring technique and results in 500 cases. AB - Corrective otoplasty is a commonly performed procedure to change the shape of the auricular cartilage. Many techniques use permanent sutures to maintain the cartilage folding, whereas other techniques rely on cartilage incisions (partial thickness or full thickness). At this institution, a cartilage cutting and anterior scoring technique has been used for more than 30 years with pleasing results. The surgical techniques published in the past have been reviewed and compared with the procedure used at this institution to point out the advantages, disadvantages, and differences of these various techniques. Also reviewed were 500 consecutive cases operated on under local or general anesthesia between January of 1993 and December of 1995 to determine the incidence of early and late complications. The patients were contacted by mail to return for a follow-up examination or answer a questionnaire, at least 2 years after the procedure. Early complications were bleeding in 13 cases (2.6 percent) and hematoma in 2 cases (0.4 percent). There were no infections or ear necrosis. A small cutaneous wound was present on the anterior skin in three patients (0.6 percent), and there was one wound dehiscence (0.2 percent). Late complications were keloids in two cases and inclusion cysts in three cases. Residual deformity was noted in 22 cases and asymmetry in 28 cases. Secondary surgery was performed in six cases. The questionnaire was answered by 387 patients (77.4 percent response rate): pain when the ear is touched was present in 22 cases (5.7 percent), hypesthesia in 15 cases (3.9 percent), occasional cutaneous irritation in 38 cases (9.8 percent), asymmetry in 71 cases (18.4 percent), and abnormal ear shape in 17 cases (4.4 percent). Twenty-nine patients (7.5 percent) also noted that the ear was more sensitive to cold or touch. The satisfaction rate was 94.8 percent: very satisfied, 74 percent; satisfied, 20.8 percent; dissatisfied, 4.2 percent; and very dissatisfied, 1 percent. These results were compared with other published series of complications and late results after otoplasty; the complication rates are similar or lower in this study. Therefore, it can be concluded that the cartilage cutting and anterior scoring technique otoplasty is a safe procedure with a high patient-parent-surgeon satisfaction rate. PMID- 10697154 TI - The influence of airbag and restraining devices on the patterns of facial trauma in motor vehicle collisions. AB - According to the National Highway Traffic Safety Administration (1990), there were more than 3 million motor vehicle collisions severe enough to lead to significant injury or fatality. Airbags may prevent brain and facial injury caused by these accidents. To date, however, no study has focused primarily on the correlation between facial injuries and the use of airbags and restraining devices. A retrospective analysis was performed on motor vehicle collision data submitted to the Pennsylvania Trauma Outcome Study database from 1990 through 1995. Criteria for submission to the database included admission to the intensive care unit, death during hospitalization, hospitalization for >72 hours, or transfer to or from the receiving hospital. There were 15,450 patients who sustained facial trauma (identified by ICD-9 codes) and were analyzed for patterns of injury and the presence or absence of protective devices. Protective devices were categorized into four groups: airbag alone, airbag with seatbelt, seatbelt or car seat without airbag, and no restraining devices. Statistical analysis was performed using chi-squared test of association. For contingency tables with small expected frequencies, Fisher's exact test was used. There were 9408 male and 6042 female subjects, with a mean age of 38 years (range, 3 to 98 years). There were 11,672 drivers and 3778 passengers. Airbags were deployed in 429 instances. In 276 of these cases, additional restraint was provided with a seatbelt. Airbags were not deployed in 4866 cases when a seatbelt or a car seat was used. In 10,155 cases, no restraining device was employed. There was significantly more facial trauma in patients without protective devices (p < 0.001). Drivers sustained significantly fewer facial fractures when airbags were used, either alone or in combination with a seatbelt (p < 0.001); however, there was no difference in the number of facial lacerations. Among passengers, airbags provided protection from lacerations (p < 0.001) but had no impact on the incidence of facial fractures. In collisions in which airbags were deployed, the use of a seatbelt provided no additional protection from facial fractures or lacerations. In summary, the use of any protective device decreased the incidence of facial fractures and lacerations sustained in motor vehicle collisions (p < 0.001). Airbags provided the best protection of all currently available devices. PMID- 10697155 TI - Risk factors contributing to symptomatic plate removal in maxillofacial trauma patients. AB - This study analyzed the fate of plates used to correct maxillofacial injuries and defined risk factors that eventually resulted in plate removal. The outpatient clinic files of 108 patients treated with rigid internal fixation after maxillofacial trauma were reviewed. Study variables included age, sex, trauma circumstances, diagnosis, type of fracture, approach to the facial skeleton, presence of teeth in the line of fracture, plate material, site of plates, and reasons for plate removal. Of 204 plates used for fixation, 44 plates (22 percent) were removed. When all factors were considered together, only fracture diagnosis (mandibular body and angle) and plate location (mandibular body and angle) were statistically significant. Only when each factor was considered separately, the approach to the facial skeleton (intraoral) and the type of fracture (comminuted and compound fractures) were statistically associated with plate removal. Selection of favorable plate location, the extraoral approach, and vigilant infection control may reduce plate removal in patients with maxillofacial injuries. Special attention should be given to compound and comminuted fractures of the mandibular body and angle. PMID- 10697156 TI - Histologic results of neuronal anastomosis of the microvascular latissimus dorsi transplant. AB - Latissimus dorsi transplants have little neuronal regenerative capacity without neuronal anastomosis. Histologic differences between transplants with and without neuronal anastomosis and two distinct types of neurotization are highlighted in this study. Eighteen patients after tumor resection and defect coverage with a latissimus dorsi transplant were examined preoperatively and postoperatively by means of a biopsy for histologic examination. The number of fascicles, degree of scarring, myelinization, and fibrosis were examined. All patients had a mean of 11.8 fascicles preoperatively. Patients without neuronal anastomosis showed an average of 5.0 fascicles, patients with nerve anastomosis to the cutaneous branches of the intercostal nerve showed an average of 6.2 fascicles, and patients with anastomosis to the thoracodorsalis nerve showed an average of 9.2 fascicles postoperatively. In cases of nerve anastomosis, a lesser degree of fibrosis was found, together with good myelinization. Neuronal reanastomosis led to more vital neuronal structures in the postoperative histologic specimen. The highest density of fascicles was found in the case of the well-vascularized thoracodorsalis nerve. PMID- 10697157 TI - Breast-feeding after inferior pedicle reduction mammaplasty. AB - The breast-feeding practices of a series of postpartum women, who had undergone prior reduction mammaplasty by means of an inferior pedicle approach, are reported in this retrospective study. Also identified are the factors that influenced the decision to breast-feed postoperatively. From a patient pool of 544 individuals who elected to have reduction mammaplasty between 1984 and 1994 (age range, 15 to 35 years), 334 could be contacted and interviewed by means of telephone by using a standardized questionnaire. Successful breast-feeding was defined as the ability to feed for a duration equal to or greater than 2 weeks. Seventy-eight patients had children after their breast reduction surgery. Fifteen of the 78 patients (19.2 percent) breast-fed exclusively, 8 (10.3 percent) breast fed with formula supplementation, 14 (17.9 percent) had an unsuccessful breast feeding attempt, and 41 (52.6 percent) did not attempt breast-feeding. Of the 41 patients not attempting to breast-feed, 9 patients did so as a direct consequence of discouragement by a health care professional. Further reasons for feeding with supplementation, having an unsuccessful attempt, and not attempting to breast feed are presented. Of the 78 women who had children postoperatively, a total of 27 were discouraged from breast-feeding by medical professionals with only 8 of the 27 (29.6 percent) subsequently attempting, despite this recommendation. In comparison, 26 patients were encouraged to breast-feed; nineteen (73.1 percent) of them did subsequently attempt breast-feeding. This rate is statistically significant by using a chi2 test with 1 df(p = 0.0016). Postpartum breast engorgement and lactation was experienced by 31 of the 41 patients not attempting to breast-feed. Of these 31 patients, 19 believed that they would have been able to breast-feed due to the extent of breast engorgement and lactation experienced. Given the use of an inferior flap mammaplasty technique and patient encouragement, the possibility for breast-feeding after reduction mammaplasty exists. This prevalence falls near the breast-feeding rate found in the population not having undergone breast surgery, according to an article in the Canadian Journal of Public Health. PMID- 10697158 TI - Breast cancer stage at diagnosis and survival among patients with prior breast implants. AB - Longstanding concern exists regarding the potential for women with breast implants to experience delayed detection of breast cancer. Furthermore, survival among cosmetic breast implant patients who subsequently develop breast cancer is a concern. Since 1976, this institution has monitored cancer incidence in a cohort of 3182 women who underwent cosmetic breast augmentation between 1959 and 1981. The distributions of stage at diagnosis and survival of the 37 women who subsequently developed in situ or invasive breast cancer were compared with the observed population distributions. The distribution of stage at diagnosis for cosmetic breast implant patients who subsequently developed breast cancer was virtually identical to that of all breast cancer patients in Los Angeles County who were of the same age and race, and were diagnosed during the same time period. Furthermore, the 5-year survival rate of the 37 patients did not differ from that which would be expected based on rates established by the U.S. National Cancer Institute's Surveillance, Epidemiology, and End Results (SEER) program. These results suggest that cosmetic breast implant patients are not at increased risk of delayed detection of breast cancer, nor do they suffer a poorer prognosis when breast cancer does occur. Although the number of breast cancer patients in this study is small, the results are highly consistent with the existing epidemiologic evidence related to breast cancer detection and survival among breast implant patients. Although breast implant patients should continue appropriate breast cancer screening behavior, there seems to be no cause for alarm. PMID- 10697159 TI - Implementation and evaluation of a clinical pathway for TRAM breast reconstruction. AB - Among strategies recently proposed to reduce practice variation, promote quality, and control costs in health care delivery, the concept of the clinical pathway has received considerable attention. Because transverse rectus abdominis musculocutaneous (TRAM) breast reconstruction is a common and often costly intervention, this institution sought to evaluate cost and quality outcomes of a clinical pathways program for this procedure. The TRAM reconstruction clinical pathway was implemented in April of 1996 to standardize postoperative care in this patient population. Outcomes of consecutive pathway cases for the first 14 months of the program were assessed in a retrospective cohort design, by using all nonpathway TRAM cases from the 18 months immediately before pathway implementation as controls. Outcomes assessed included length of hospital stay, postoperative complications, total postoperative charges, and total postoperative costs in relative value units. Data on these dependent variables were collected from hospital charts and billing records. The effects of pathway implementation on the outcomes of interest were analyzed by using analysis of covariance to control for potential confounding by other independent variables, including surgical site (unilateral versus bilateral reconstructions), technique (pedicle versus free TRAMs), timing (immediate versus delayed reconstructions), and patient age. Finally, a comparison of variances in the outcomes of interest between the two groups was analyzed by using an Ftest. For all statistical tests, p values of < or = 0.05 were considered significant. Twenty-nine patients were treated in the TRAM pathway group, whereas the control population included 40 nonpathway patients. After implementation of the TRAM pathway, length of stay decreased from 6.0 to 5.2 days; total postoperative charges were reduced from $8587 to $7744; and total postoperative relative value unit utilization declined from 1686 to 1104. Analysis of covariance showed that the decreases in length of hospital stay and relative value units in the TRAM pathway were statistically significant (p = 0.05 and p = 0.007, respectively). By contrast, no significant increase in complications was observed after pathway implementation. Variability in the TRAM pathway group, as measured by SD, decreased significantly for both length of hospital stay (p = 0.039) and relative value units (p = 0.023). Implementation of the TRAM reconstruction clinical pathway resulted in significant declines in length of hospital stay and total costs. These decreases in resource utilization had no significant effect on postoperative complication rates. Although additional research is needed to further assess the impact of clinical pathways, this approach offers considerable promise for improving the cost-effectiveness of health care. PMID- 10697160 TI - Inframammary fold: a histologic reappraisal. AB - The inframammary fold is a defining element in the shape and structure of the female breast. It should be preserved whenever possible in ablative procedures and recreated accurately when the breast is reconstructed after mastectomy. To date, no accurate anatomic description of this essential structure exists. Previous studies have suggested that the fold is produced by a supporting ligament running from the dermis in the fold region to a variety of locations on the rib cage. This clinic's experience with mastectomy, augmentation mammaplasty, and breast reconstruction does not support the existence of a ligamentous structure. To define the structure of the inframammary fold, 10 female and 2 male cadavers were studied. The anterior chest wall was removed en bloc and frozen in orthostatic position. Parasagittal sections were made of the inframammary fold with the chest wall intact. After decalcification of the ribs and routine histologic preparation, thin sections were stained with Gomori's trichrome. On light microscopic examination, no demonstrable ligamentous structure of dense regular connective tissue could be identified in the fold region in any of the 12 specimens. Superficial and deep fascial layers were uniformly observed anterior to the pectoralis major and serratus anterior muscles. The superficial fascia was connected to the dermis in the fold region in a variety of configurations. In some cases, the deep fascia fused with the superficial fascia and dermis at the fold level. In other cases, bundles of collagen fibers arising from the superficial fascial layer were found to insert into the dermis at the inframammary fold, slightly inferior to it, or both. These bundles were observed consistently in sections from the sternum to the middle axillary line. They were distinct from Cooper's suspensory ligaments, which are seen more superiorly in the glandular tissue. PMID- 10697161 TI - A comparison of thinning and conventional free-flap transfers to the lower extremity. AB - This study compares the application of conventional free flaps and thinning flaps to the lower extremities. Thirty patients whose skin and soft tissue of the lower extremities had been reconstructed were divided into two groups: a conventional flap group, reconstructed using conventional free flaps (15 cases), and a thinning flap group, reconstructed using thinning flaps (15 cases). Postoperative complications, long-term results, and revisional surgery were studied in the two groups. Although survival after surgery was the same in both, in the conventional flap group, 11 patients required secondary revisional surgery, the excessive bulk of the flap resulting in poor aesthetics and difficulty in wearing shoes. The conventional flap group also required longer treatment. In the thinning flap group, only 5 of 15 patients received secondary revisional surgery. As a reconstruction material for the lower extremities, thinning flaps are both aesthetically and functionally superior to conventional bulky flaps. PMID- 10697162 TI - Soleus-fibula free transfer in lower limb reconstruction. AB - Free-fibula transfer has been widely used since 1975. Many modifications have been described; one of them, association of the lateral part of the soleus muscle to the fibula, is reported here through a 14-case series. This composite flap is intended for extensive defects of the lower limbs involving bone and soft tissues. The flap is considered by the authors to be reliable, with a constant vascularization. A 20-cm length offibula may be harvested associated either with the lateral part of the soleus muscle or with the whole muscle. Moreover, the soleus muscle represents a vascular security inasmuch as it preserves both medullar and periosteal bone supply. Fourteen cases have been performed by the authors since 1978 and could be reviewed with a minimum 2-year follow-up. Average length of bone defect was 12 cm, and average length offibula harvested was 18.6 cm. Soft-tissue defect was always associated and ranged from 8 x 4 cm to 20 x 30 cm. The fibula was harvested with the lateral part of the soleus muscle in 10 cases and with the whole soleus muscle in 4 cases. One total treatment failure was reported and was related to intimal degenerative lesions on veins used for arteriovenous bypass. In other patients, mean time for bone healing was 11 months. Patients could walk again, on average, 17 months after reconstruction. Sequelae at the donor site were minimal. PMID- 10697163 TI - Comparison between sensitive and nonsensitive free flaps in reconstruction of the heel and plantar area. AB - In this study, 12 cases of reconstruction of the heel and plantar area since 1982 are reviewed. Six nonsensate muscle free flaps and six sensate fasciocutaneous flaps were used, respectively. Categories assessed were the time interval for return to daily living activities, sensation to light touch, pinprick, Semmes Weinstein monofilament test of the reconstructed area for sensory evaluation; and results of pedograms (maximal pressure, pressure distribution, and total contact area of the plantar surface). Follow-up periods were between 2 and 14 years, with an average of 6 years. Better sensory results and early return to daily living activities were observed in the sensate flap group, but the defects were smaller in this group. Despite the slightly longer time to return to daily living activities and worse sensory results, long-term follow-up showed that patients with nonsensate flaps had no difficulty in performing living activities if they continued to be careful and to use some kind of protective shoes. The results of the pedogram analyses were similar between the two groups with regard to total contact area of the reconstructed foot in relation to the healthy foot. Pressure values of the reconstructed areas in sensate flaps were found to be close to pressure values in the same weight areas of the normal foot. The differences between pressure values of the sensate and nonsensate flaps were statistically significant (p < 0.001). Therefore, in reconstruction of the weight-bearing surface of the foot, each case should be evaluated individually. The reconstructive method should be chosen according to the location and soft-tissue requirements of the defect. PMID- 10697164 TI - Fist position for skin grafting on the dorsal hand: II. Clinical use in deep burns and burn scar contractures. AB - The fundamental problem in all types of hand burns is a loss of skin and subsequent deformities. The goal of skin grafting on the dorsal hand is to graft a sufficient amount of skin, as much as the original amount, and to restore normal hand function without secondary deformities. The safe, or Michigan, position commonly has been used for immobilizing the hand. However, this position is to protect hand function rather than to provide for adequate skin grafting. This institution has developed a new hand position (the fist position) for grafting the greatest amount of skin on the dorsal side of the hand. In the fist position, the hand is positioned flexing all joints of the wrist and the fingers and maximally stretching the dorsal surface of the hand before skin grafting. Ten hands with deep second- or third-degree burn (n = 6) and burn scar contracture (n = 4) of the dorsal hand in eight patients were treated with split-thickness skin grafting after immobilizing in the fist position. The burns and contractures involved nearly the total area of the dorsal hand. The hand was kept in the fist position for 7 to 9 days after skin grafting. Excellent functional and cosmetic results were observed in all cases during the follow-up period of 6 months to 2 years. Complications resulting from hand immobilization for a short period did not occur. The fist position may be a proper hand position for skin grafting to reconstruct the dorsal hand. PMID- 10697165 TI - Use of the rectus abdominis muscle for abdominal stoma sphincter construction: an anatomical feasibility study. AB - Permanent fecal abdominal stomas significantly decrease quality of life. Previous attempts to create continent stomas by using dynamic myoplasty procedures have resulted in disappointing outcomes, primarily owing to denervation atrophy of the muscle flap that was used in the creation of the sphincter and because of muscle fatigue resulting from continuous electrical stimulation that is received by the flap to force contraction. On the basis of these problems, we designed two separate studies: an anatomical study addressing flap denervation and a functional study addressing muscle fatigue. The present study addresses the first topic and was designed to develop a rectus abdominis muscle flap into a sphincter that was anatomically situated to create a stoma while preserving as much innervation as possible. In 24 rectus abdominis muscles of human cadavers, the neurovascular anatomy was defined, then the anatomical feasibility of two different muscle flap configurations was considered. The flaps investigated were the peninsula flap and island flap designs, with both using the most caudal segment of the rectus abdominis muscle in construction of the sphincter. Neither flap design required the killing of a nerve for stoma sphincter creation, resulting in minimal muscle denervation. The conclusion of our comparison was that the above, in conjunction with other features of the island flap design, such as muscle overlap after sphincter formation and abdominal wall positioning of the sphincter, made the island flap design better suited to stoma sphincter construction. PMID- 10697166 TI - Modified penis lengthening surgery: review of 52 cases. AB - A modified surgical method for penis lengthening was, for the first time, set up in this laboratory. The procedure involves covering the dissected corpus cavernosum with either a scrotal flap or a skin graft after releasing the superficial ligament and even some deep suspensory ligament. The advantage of the scrotal flap is emphasized to cover the wound, and a V-Y suture was made to avoid the traction. The results, both in appearance and increased length, were satisfactory in 52 cases. Among the 52 patients, 39 suffered from congenital short penis and 13 from traumatic injuries. The significance and the blood supply of the penis are discussed. PMID- 10697167 TI - Treatment modalities for hypospadias cripples. AB - Hypospadias cripples can be defined as patients with remaining functional complications after previous hypospadias repair. A retrospective follow-up study was performed on the long-term results of a group of 94 patients disabled by hypospadias. The records of 94 patients showed that they presented with the following problems: 82 had a major meatal dystopia (87 percent), 43 (46 percent) had residual curvature of the penile body, 19 (20 percent) showed meatal stenosis, and only 5 (5 percent) had one or more fistulas. The techniques used to solve these problems were circumferential advancement of penile skin, dorsal transposition flap of preputial skin, distally based transposition flap of penile skin, and full-thickness skin graft. Between one and nine operations were needed to achieve the desired result (mean and median of two operations). The complications after these procedures were 11 fistulas in nine patients, meatal stenosis caused by tight scarring in six patients, and a residual curvature after an orthoplasty that had to be released once before a urethroplasty could be performed. Forty-three men were seen at long-term follow-up (range, 2 to 25 years; mean, 12 years). Functional complaints that were seen included spraying at micturition (5 patients, 12 percent), dribbling (6 patients, 14 percent), and deviation of urinary stream (7 patients, 16 percent). No patients complained of painful miction, hesitation, or straining. At physical examination, 4 patients had a residual curvature (three of which were mild without functional problems), 5 had a skin surplus, 1 presented with a fistula after an operation in another hospital, and 13 had a penile torsion. Only 6 patients had a penile torsion greater than 10 degrees, which was evenly distributed to the left and right. There was no correlation between any functional complaint and the presence of a physical abnormality. PMID- 10697168 TI - Wall Street's assessment of plastic surgery--related technology: a clinical and financial analysis. AB - Many plastic surgeons develop technologies that are manufactured by Wall Street financed companies. Others participate in the stock market as investors. This study examines the bioengineered skin industry to determine whether it integrates clinical and financial information as Wall Street tenets would predict, and to see whether the financial performance of these companies provides any lessons for practicing plastic surgeons. In efficient markets, the assumptions on which independent financial analysts base their company sales and earnings projections are clinically reasonable, the volatility of a company's stock price does not irrationally differ from that of its industry sector, and the buy/sell recommendations of analysts are roughly congruent. For the companies in this study, these key financial parameters were compared with a benchmark index of 69 biotech companies of similar age and annual revenues (Student's t test). Five bioengineered skin companies were included in the study. Analysts estimated that each company would sell its product to between 24 and 45 percent of its target clinical population. The average stock price volatility was significantly higher for study companies than for those in the benchmark index (p < 0.05). Similarly, buy/sell recommendations of analysts for the study companies were significantly less congruent than those for the benchmark companies (p < 0.05). These results indicate clinically unrealistic projections for market penetration, significantly high price volatility, and significantly high discordance among professional analysts. In all cases, the market is inefficient-an unusual finding on Wall Street. A likely explanation for this market failure is a cycle of poor clinical correlation when assigning sales projections, which in turn leads to price volatility and discordance of buy/sell recommendations. This study's findings have implications for plastic surgeons who develop new technology or who participate in the equities markets as investors. Plastic surgeons who develop new medical devices or technology cannot universally depend on the market to drive clinically reasonable financial performance. Although inflated sales estimates have benefits in the short term, failure to meet projections exacts severe financial penalties. Plastic surgeons who invest in the stock market, because of their unique clinical experience, may sometimes be in the position to evaluate new technologies and companies better than Wall Street experts. Well timed trades that use this expertise can result in opportunities for profit. PMID- 10697169 TI - Craniofacial shortening by contraction osteogenesis: an experimental model. AB - Application of gradual external forces to correct craniofacial deformities challenges many procedures in conventional craniomaxillofacial surgery. Distraction osteogenesis is replacing traditional osteotomies for correction of patients with craniomaxillofacial deficiencies. However, the reverse concept, contraction osteogenesis, has yet to be established for patients with craniomaxillofacial excesses. The purpose of this investigation is to demonstrate the contraction osteogenesis phenomenon applied in a controlled animal model during the craniofacial growth period. Twenty-six 26-day-old rabbits were assigned to one of four groups: 0, control; 1, pin control (pin insertion); 2, no contraction (pins and contraction device application, without active contraction); and 3, contraction (pin insertion, contraction device application, and active contraction). An external fixator was placed across the incisive maxillary suture, and the effects after 4.5 weeks of contraction at a rate of 0.5 mm twice a week were compared with control groups. The results were assessed by craniometric and cephalometric measurements and by histologic examination. Gross alterations were evident in the contraction group, characterized by midface anteroposterior shortening, maxillary regression, snout deviation, and anterior crossbite. Histologic examination of the contraction group demonstrated a significant increase in osteoblastic activity. Contraction osteogenesis is a new treatment concept in craniofacial development and may offer therapeutic opportunities for shortening skeletal structures without the need of osteotomies, thus taking advantage of the potential of craniofacial growth and remodeling. PMID- 10697170 TI - Bone morphogenetic protein excipients: comparative observations on poloxamer. AB - Clinicians await the availability of synthetic bioimplants that will replace the need for autogeneic bone grafts in bone reconstructive surgery. For more than a decade, researchers have evaluated delivery vehicles for the tissue morphogen bone morphogenetic protein. The object of this investigation was to measure induced bone development when bone morphogenetic protein was delivered by human tendon collagen, human demineralized bone matrix, hydroxyapatite, a composite of human tendon collagen and human demineralized bone matrix (tendon collagen + demineralized bone matrix), Poloxamer 407, and a composite of human demineralized bone matrix and Poloxamer 407. Sixty-three adult male Swiss Webster mice (Harlan Sprague-Dawley, Indianapolis, Ind.) received 126 implants. The animals were divided into seven groups of nine animals, depending on carrier (six carriers plus the positive control group) used. Each animal received a bone morphogenetic protein-enhanced carrier in one hindquarter muscle mass, with the contralateral leg being implanted with the carrier alone. Implants were evaluated by quantitative radiomorphometry validated by histologic methods. Radiographically, no significant differences were identified among any of the implants evaluated (p > 0.05). Histomorphometric analysis demonstrated that Poloxamer 407 was significantly (p < 0.05) better at delivering bone morphogenetic protein than the other carriers involved in this investigation. The new bone developed in a tubular or spherical shape. Interaction of endogenous and exogenous delivery systems seems to be essential for optimal transmission of bone morphogenetic protein. The importance of the excipient to deliver bone morphogenetic protein and develop a bone morphogenetic protein concentration gradient has been emphasized by other investigators and confirmed by our research on poloxamer. With further research on the physicochemical mechanisms of localization and transmission of bone morphogenetic protein, it may be possible to avoid hazardous operations with autogeneic bone. PMID- 10697171 TI - Differential expression of matrix metalloproteinases and their tissue-derived inhibitors in cutaneous wound repair. AB - Wound extracellular matrix is a key regulator of cell adhesion, migration, proliferation, and differentiation during cutaneous repair. The amount and organization of normal wound extracellular matrix are determined by a dynamic balance among overall matrix synthesis, deposition, and degradation. Matrix metalloproteinases (MMPs) are one family of structurally related enzymes that have the collective ability to degrade nearly all extracellular matrix components. The MMPs are broadly categorized into collagenases, gelatinases, stromelysins, and membrane-type MMPs by their substrate specificity. The aim of this study was to characterize the temporal changes in mRNA profiles for rat collagenase [matrix metalloproteinase-1 (MMP-1)], gelatinase A (MMP-2), matrilysin (MMP-7), gelatinase B (MMP-9), and membrane type 1-MMP (MT1-MMP), as well as tissue inhibitor of metalloproteinases-1 (TIMP-1), TIMP-2, and TIMP-3 during the inflammatory, granulation, and early remodeling phases of excisional skin repair. Eight full-thickness skin wounds were made on the backs of each rat (7-mm2 wounds; 16 rats; n = 128 wounds). Two animals at a time were reanesthetized, and all eight wounds on each animal were excised at 12 and 24 hours and at 2, 3, 5, 7, 10, and 14 days after injury. Six wounds from each animal were excised for RNA isolation, whereas two wounds were excised for histology. Controls consisted of nonwounded skin from identical locations in four animals. Total RNA from each time point was isolated and relative mRNA quantitation performed by using reduced-cycle reverse transcription-polymerase chain reaction. Correct polymerase chain reaction product amplification was confirmed by probing the blotted polymerase chain reaction product with a 32P labeled oligonucleotide specific for a given MMP or TIMP. We demonstrated that the majority of MMP and TIMP mRNA induction and peak expression coincided temporally with the well-characterized inflammatory and granulation stages of repair. In conclusion, there is a distinct pattern of MMP and TIMP expression during normal excisional wound repair. PMID- 10697172 TI - Initially alloperfused autograft: an experimental study with rat groin free flap. AB - The purpose of this experimental study was to investigate the effects of alloperfusion on autografts. The authors designed an experimental model to study flap viability during and after alloperfusion. They performed 23 free groin flap transfers on 46 rats. The flap was transferred as an autograft; however, the anastomosis was performed as an allograft procedure. After the operation, animals were held together on a table for observation. The rats were randomly divided into two groups depending on whether or not a steroid was given. They were fed by hand. At the end of the various waiting periods, the pedicles were divided, and animals were separated from each other. The survival of the flap and the animal was observed. No flaps survived in the untreated group. Seven flaps in the steroid-treated group that had their connections cut before 140 hours also did not survive. Five flaps that had their pedicles cut after 140 hours survived. Results are presented and discussed. PMID- 10697174 TI - Modulation of peripheral nerve regeneration: a tissue-engineering approach. The role of amnion tube nerve conduit across a 1-centimeter nerve gap. AB - A new type of a biodegradable nerve graft conduit material, the amnion tube, has been developed in our laboratory. To test the tube in the peripheral nerve regeneration process, it was initially applied across a 1-cm sciatic nerve gap in rats and was compared with other nerve conduit materials. We used male Sprague Dawley rats as our animal model. The experiment included 66 rats that were randomly assigned into five groups: autograft (n = 17), amnion tube (n = 19), silicone tube (n = 20), no repair (n = 7), and sham group (n = 3). The process of peripheral nerve regeneration was evaluated at 2, 4, 10, and 17 weeks following injury and repair by using morphologic and functional assessments of the outcome of nerve regeneration in each animal. Nerve regeneration across the amnion tube nerve conduit was comparable with that seen in autograft and superior to that of the silicone group. A uniform nerve tissue was seen filling and crossing the amnion conduit, and the regenerated nerve from the proximal stump reached the distal end and was undifferentiated from the normal nerve tissues. At 4 months, the amnion tube biodegraded and no longer could be identified and differentiated from the nerve tissues. The amnion tube animal group showed a number of axons very close to that in the nerve autograft group (37,157 versus 33,054). Functional recovery at a 2- to 4-week interval was significantly statistically higher only in the amnion tube animal group (p = 0.01). However, the improvement disappeared between 10 and 17 weeks. In conclusion, the amnion tube is a potential ideal nerve conduit material secondary to its unique characteristics: it contains important neurotropic factors, is biodegradable, provokes a very weak immune response, is semiflexible, is readily available, and is easily manufactured into different sizes and diameters. PMID- 10697173 TI - Prevention of ischemia-reperfusion injury in a rat skin flap model: the role of mast cells, cromolyn sodium, and histamine receptor blockade. AB - The objective of this study was to examine the role of mast cells and their principal product, histamine, in ischemia/reperfusion injury. Cromolyn sodium, diphenhydramine, and cimetidine were administered to ischemic flaps just before reperfusion and evaluated for flap survival, mast cell count, neutrophil count, and myeloperoxidase levels. Epigastric island skin flaps were elevated in 49 rats; they were rendered ischemic by clamping the artery for 10 hours. Thirty minutes before reperfusion, the rats were treated with intraperitoneal saline (n = 11), cimetidine (n = 11), diphenhydramine (n = 11), or cromolyn sodium (n = 10). Flap survival was evaluated at 7 days. Neutrophil counts, mast cell counts, and myeloperoxidase levels were evaluated 12 hours after reperfusion. Flap necrosis in the sham group of animals (n = 6) was 0.0 percent, as expected, whereas the control group (saline-treated animals) had 47.3+/-33.4 percent necrosis. Animals treated with diphenhydramine and cimetidine demonstrated a significant decrease in flap necrosis to 17.7+/-8.8 percent and 19.4+/-14.7 percent, respectively. This protective effect was not seen with cromolyn sodium (44.3+/-35.6 percent). Both neutrophil and mast cell counts were significantly decreased in flaps from antihistamine-treated and sham animals versus both saline and cromolyn sodium-treated groups. The administration of diphenhydramine and cimetidine before reperfusion can significantly reduce the extent of flap necrosis and the neutrophil and mast cell counts caused by ischemia/reperfusion. This protective effect is not seen with cromolyn sodium. The protective effect of antihistamines on flap necrosis might be related to the decrease in neutrophils and, possibly, mast cells within the flap. PMID- 10697175 TI - Sequential segmental neuromuscular stimulation: an effective approach to enhance fatigue resistance. AB - Electrical stimulation of skeletal muscle flaps is used clinically in applications that require contraction of muscle and force generation at the recipient site, for example, to assist a failing myocardium (cardiomyoplasty) or to reestablish urinary or fecal continence as a neo-sphincter (dynamic graciloplasty). A major problem in these applications (muscle fatigue) results from the nonphysiologic manner in which most of the fibers within the muscle are recruited in a single burst-like contraction. To circumvent this problem, current protocols call for the muscle to be put through a rigorous training regimen to transform it from a fatigue-prone to a fatigue-resistant state. This process takes several weeks during which, aside from becoming fatigue-resistant, the muscle loses power and contraction speed. This study tested the feasibility of electrically stimulating a muscle flap in a more physiologic way; namely, by stimulating different anatomical parts of the muscle sequentially rather than the entire muscle all at once. Sequential segmental neuromuscular stimulation (SSNS) allows parts of the muscle to rest while other parts are contracting. In a paired designed study in dogs (n = 7), the effects of SSNS on muscle fatigability and muscle blood perfusion in gracilis muscles were compared with conventional stimulation: SSNS on one side and whole muscle stimulation on the other. In SSNS, electrodes were implanted in the muscles in such a way that four separate segments of each muscle could be stimulated separately. Then, each segment was stimulated so that part of the muscle was always contracted while part was always resting. This type of stimulation permitted sequential yet continuous force generation. Muscles in both groups maintained an equal amount of continuous force. In SSNS muscles, separate segments were stimulated so that the duty cycle for any one segment was 25, 50, 75, or 100 percent, thus varying the amount of work and rest that any segment experienced at any one time. With duty cycles of 25, 50, and 75 percent, SSNS produced significantly (p < 0.01) enhanced resistance to fatigue. In addition, muscle perfusion was significantly (p < 0.01) increased in these sequentially stimulated muscles compared with the controls receiving whole muscle stimulation. It was concluded that SSNS reduces muscle fatigue and enhances muscle blood flow during stimulation. These findings suggest that using SSNS in clinical myoplasty procedures could obviate the need for prolonged training protocols and minimize problems associated with muscle training. PMID- 10697176 TI - Manipulation of callus after linear distraction: a "lifeboat" or an alternative to multivectorial distraction osteogenesis of the mandible? AB - Despite impressive results, distraction osteogenesis of the mandible is still compromised by difficulties with vector control. Because of the action of the masticatory muscles, the gonion angle has the tendency to open, resulting in an open bite. We report two patients, aged 10 and 12 years, who developed a severe open bite during mandibular distraction. As a salvage procedure, manual shaping of the soft regenerate was done immediately after distraction. Uneventful bony consolidation was observed, which resulted in anatomically shaped gonion angles. The fact that a regenerate created by distraction osteogenesis can be molded to virtually any shape offers interesting perspectives for the correction of complex mandibular deformities. PMID- 10697177 TI - Lymphatic mapping for Merkel cell carcinoma. AB - Merkel cell trabecular carcinoma of the skin has a prognosis poorer than expected for a small skin lesion. An early diagnosis and prompt treatment can contribute to improve survival in cases of this aggressive skin tumor. A wide local excision is indicated for localized disease. Elective lymph node dissection is controversial. The authors report a rare case of Merkel cell tumor treated with wide local excision and sentinel lymph node biopsy. PMID- 10697178 TI - Free flap to the arteria peronea magna for lower limb salvage. AB - A 36-year-old woman sustained an amputation of her right leg at the thigh level and a degloving injury of her left foot and ankle region in an accident during a suicide attempt. Primarily, her left foot was covered with a split skin graft, resulting in a soft-tissue defect at the medial malleolus and at the calcaneus bone. Reconstruction was planned with a free latissimus dorsi muscle flap. Preoperative examinations revealed an arteria peronea magna with a hyperplastic peroneal artery solely providing arterial blood supply to the foot. The arteria peronea magna divided into two branches proximal to the upper ankle joint, replacing the dorsal pedis artery and the medial plantar artery. Tibial posterior and tibial anterior arteries were hypoplastic-aplastic. Microvascular end-to-end anastomoses of the flap vessels to the medial branch ("medial plantar artery") of the arteria peronea magna and its concomitant vein at the medial malleolar bone level were successfully performed. The postoperative course was uneventful. Four weeks postoperatively, the patient started walking assisted by a prosthesis on her right thigh stump. This experience demonstrates that even in a case of arteria peronea magna, free flap surgery for lower limb salvage is a reliable and worthwhile method. PMID- 10697179 TI - New surgical technique for macrostomia repair with two triangular flaps. AB - Our new surgical procedure with two triangular flaps for macrostomia repair allows us to achieve all three therapeutic goals, including formation of symmetric lips and commissures of the mouth, reconstruction of the orbicularis muscle of mouth to restore labial function, and reconstruction of the commissure of the mouth with a natural looking contour. Furthermore, the position of the commissure of the mouth can be adjusted intraoperatively according to the extent of macrostomia. As reported here, our method provides very satisfactory clinical results and is relatively easy to perform. Thus, we believe that our method can serve as a standard for the surgical treatment of macrostomia. PMID- 10697180 TI - Modified towel-clamp technique to effect reduction of displaced mandible fractures. AB - We report a technique that uses a modified standard towel clamp, allowing a single surgeon to perform and maintain an anatomic reduction of displaced mandible fractures simultaneous with the application of internal fixation. The reduced convergent angle of the modified towel clamp allows bicortical engagement of the clamp, which prevents comminution of the fracture or the outer cortex of the mandible. Additionally, the modification allows the clamp to engage the bone with less exposure than conventional towel clamps. In our clinical experience of treating more than 100 mandible fractures a year, this technique proves superior to others described in the literature. PMID- 10697181 TI - Skin-stretching device for intraoperative primary closure of radial forearm flap donor site. AB - The use of a skin-stretching device to close radial forearm donor-site defects in patients being treated for intraoral cancer is described. Intraoperative application of the device achieved primary closure of the donor site during the time course of the reconstructive procedure with no requirement for additional operative time. The device was used successfully in seven patients with two instances of marginal skin necrosis and delayed healing but good long-term functional and cosmetic outcomes in all cases. PMID- 10697182 TI - Umbilical reconstruction with an inverted C-V flap. AB - We have created a method for umbilical reconstruction with satisfactory results. The C-V flap developed for nipple reconstruction was used in an inverted fashion. The inverted C-V flap can produce a satisfactory reconstruction of umbilical structures, especially the ring. PMID- 10697183 TI - Widely split latissimus dorsi muscle flaps for reconstruction of long soft-tissue defects in lower extremities. AB - Although the latissimus dorsi is one of the largest and longest muscles in the human body, it is still sometimes inadequate for reconstruction of a soft-tissue defect of extensive length and dimension. Eight patients with such lower limb defects were treated with latissimus dorsi muscles split into two hemiflaps sequentially linked, one after the other like a chain. Six transfers were completely successful, one required reexploration for arterial occlusion, and two hemiflaps had a partial loss that could be managed by touching up the skin graft. The average split sequential-link muscle was 42 cm in length. Although two patients had a partial loss, we consider that the widely split single latissimus dorsi muscle can still be used reliably to reconstruct a long slender defect, or two separate, longitudinally located, medium-sized defects in the same leg. PMID- 10697184 TI - A new and safer anastomosis technique in cross-leg free flap procedure using the dorsalis pedis arterial system. PMID- 10697185 TI - Evaluation and treatment of BRCA-positive patients. AB - The recent discovery of the breast cancer-associated genes BRCA1 and BRCA2 has now made it possible to identify individuals who are at a very high risk for the future development of breast cancer. To some extent, however, society has fallen victim to its molecular genetic technology. The significance of these discoveries to the detection, treatment, and prevention of breast cancer cannot be overstated. Nevertheless, the appropriate administration and interpretation of BRCA genetic testing and the treatment of BRCA-positive patients remain controversial issues. Complexities of BRCA testing require that such genetic screening be restricted to selected high-risk patients and that test results be interpreted by a knowledgeable molecular geneticist. Although no medical prophylaxis has been demonstrated to be of benefit in BRCA-positive patients, recent evidence suggests that a prophylactic mastectomy, with or without reconstruction, is a reasonable treatment option that substantially reduces cancer risk. PMID- 10697186 TI - Endoscopically assisted "components separation" for closure of abdominal wall defects. AB - The repair of ventral hernia defects of the abdominal wall challenges both general and plastic surgeons. Ventral herniation is a postoperative complication in 10 percent of abdominal surgeries; the repair of such defects has a recurrence rate as high as 50 percent. The "components separation" technique has successfully decreased the recurrence rates of ventral abdominal hernias. However, this technique has been associated with midline dehiscence and a prolonged postoperative stay at the authors' institutions. The purpose of this study was to determine whether endoscopically assisted components separation could minimize operative damage to the vasculature of the abdominal wall and decrease postoperative wound dehiscence. The study group consisted of seven patients who underwent endoscopically assisted components separation; the control group consisted of 30 patients who underwent open components separation. The two groups were similar regarding demographic data and defect size. The endoscopic group had a higher initial success rate than the open group (100 versus 77 percent). Recurrence rates were not significantly different between the two groups. However, the endoscopically assisted components separation patients had fewer postoperative and long-term complications. In the authors' experience, endoscopically assisted components separation has proved to be a safe and effective method for the repair of complicated and recurrent midline ventral hernias. PMID- 10697187 TI - The separation of anatomic components technique for the reconstruction of massive midline abdominal wall defects: anatomy, surgical technique, applications, and limitations revisited. AB - Reconstruction of massive abdominal wall defects has long been a vexing clinical problem. A landmark development for the autogenous tissue reconstruction of these difficult wounds was the introduction of "components of anatomic separation" technique by Ramirez et al. This method uses bilateral, innervated, bipedicle, rectus abdominis-transversus abdominis-internal oblique muscle flap complexes transposed medially to reconstruct the central abdominal wall. Enamored with this concept, this institution sought to define the limitations and complications and to quantify functional outcome with the use of this technique. During a 4-year period (July of 1991 to 1995), 22 patients underwent reconstruction of massive midline abdominal wounds. The defects varied in size from 6 to 14 cm in width and from 10 to 24 cm in height. Causes included removal of infected synthetic mesh material (n = 7), recurrent hernia (n = 4), removal of split-thickness skin graft and dense abdominal wall cicatrix (n = 4), parastomal hernia (n = 2), primary incisional hernia (n = 2), trauma/enteric sepsis (n = 2), and tumor resection (abdominal wall desmoid tumor involving the right rectus abdominis muscle) (n = 1). Twenty patients were treated with mobilization of both rectus abdominis muscles, and in two patients one muscle complex was used. The plane of "separation" was the interface between the external and internal oblique muscles. A quantitative dynamic assessment of the abdominal wall was performed in two patients by using a Cybex TEF machine, with analysis of truncal flexion strength being undertaken preoperatively and at 6 months after surgery. Patients achieved wound healing in all cases with one operation. Minor complications included superficial infection in two patients and a wound seroma in one. One patient developed a recurrent incisional hernia 8 months postoperatively. There was one postoperative death caused by multisystem organ failure. One patient required the addition of synthetic mesh to achieve abdominal closure. This case involved a thin patient whose defect exceeded 16 cm in width. There has been no clinically apparent muscle weakness in the abdomen over that present preoperatively. Analysis of preoperative and postoperative truncal force generation revealed a 40 percent increase in strength in the two patients tested on a Cybex machine. Reoperation was possible through the reconstructed abdominal wall in two patients without untoward sequela. This operation is an effective method for autogenous reconstruction of massive midline abdominal wall defects. It can be used either as a primary mode of defect closure or to treat the complications of trauma, surgery, or various diseases. PMID- 10697188 TI - The importance of cosmetic plastic surgery education: an evolution. PMID- 10697189 TI - Transconjunctival orbital fat repositioning: transposition of orbital fat pedicles into a subperiosteal pocket. AB - Rejuvenation of the lower eyelid complex is based on the principle that the contour changes characterizing aging involve not only prolapse of orbital fat but also descent of the cheek tissues, resulting in accentuation of the orbital rim and tear trough groove. When a deep groove is present along the orbital rim in the area of the tear trough deformity, it is advantageous, rather than removing orbital fat, to reposition the fat over the orbital rim through the opened arcus marginalis onto the superior face of the maxilla. Orbital fat repositioning can be accomplished through a transconjunctival approach. The arcus marginalis is exposed and incised, and a subperiosteal pocket is created over the superior face of the maxilla. The subperiosteal pocket shape and location are customized based on the desired location of the orbital fat pedicle; often the origins of the levator superioris labialis and the levator alae nasi muscles are partially dissected. Medial and central fat pedicles are created and rotated over the orbital rim into the subperiosteal pocket. A 6-0 polypropylene externalized sutured is used to fixate the fat pedicle in position. The suture can be removed after 3 to 5 days. Twenty-four patients were followed clinically after orbital fat repositioning, with follow-up ranging from 6 to 30 months. Although the fat pedicle undergoes some variable resorption, the viability of the graft, the texture and contour of the repositioned fat after a healing period of 1 to 2 months, and the excellent patient acceptance are indicative of the viability of orbital fat repositioning. PMID- 10697190 TI - An examination of the phenol-croton oil peel: Part III. The plastic surgeons' role. AB - In Part II, the author focused on the lay peelers' history and success using croton oil-containing phenol peeling recipes. In Part III, the author reviews what was known or should have been known about croton oil and phenol as physicians became keenly interested in face peeling in the mid-1950s. The lay peelers recognized that croton oil was a critical ingredient of the so-called phenol peel while physicians focused on phenol without recognizing the intense cytotoxic effect of croton resin. Physicians have persisted in this systematic error for 40 years. Both dermatology and plastic surgery have shown a remarkable credulity about phenol's action and lack of curiosity about croton oil's action. A hitherto unreferenced and unknown croton oil-containing formula of Adolph Brown, patented in 1959, has been unearthed, preceding Litton's and Baker's formulas in time. The recollections of Litton, Baker, Truppman, and Georgiade shed some light on the interaction between them and the lay peelers and how the formulas were transferred. Other plastic surgeons probably acquired the same knowledge, used it in their practices, but chose not to draw attention to it. None of the physicians credited the lay peelers. Brown, Litton, and Baker each could have published a complete formula, but only Baker did. However, his formula was vastly stronger than the lay formulas but, nevertheless, came to dominate medical peeling for the next 35 years because of its simplicity of preparation. A review of the peel literature reveals many oft-repeated but unsupported dogmas regarding the mode of action of phenol, which have obscured our understanding of the phenol-croton oil peel. Animal studies exist that refute these dogmas, e.g., (1) lesser concentrations of phenol wound more deeply; and (2) phenol has an all or-nothing action. As well, studies from the early 1980s showed that the presence of croton oil caused much deeper burns than phenol alone. Suggested areas of research that could solve the conundrum of the phenol-croton oil peel are presented. PMID- 10697191 TI - The changing role of platysma in face lifting. AB - Since the 1970s, surgical procedures on platysma muscle, aiming to achieve better results in face lifting, became popular and turned out to be an important surgical step for the plastic surgeon. Many plastic surgeons have contributed to the topic throughout these years, as several articles on the subject have been published. Articles dealing with platysma muscle still bring great interest among plastic surgeons. My concern with platysma muscle began in the mid-1970s and since then has grown continuously. I have steadily been studying the importance of the platysma muscle in the surgeries for facial rejuvenation, involving its anatomy, the techniques proposed, the results obtained, the recommendations for the best surgical procedure for each patient, and the complications. My experience and studies regarding platysma muscle, and the contributions I have brought into this field, are thoroughly described and discussed in this article. PMID- 10697192 TI - Botulinum toxin A for the treatment of facial hyperkinetic wrinkle lines in Koreans. AB - There are a number of different causes for facial wrinkle lines, such as aging, gravity, and chronic pulling of mimetic muscles on the face. Among these, pulling by mimetic muscles on the skin not only involves facial expression but also has a great role in forming facial wrinkle lines as a result of repetitive action, such as dynamic or hyperkinetic wrinkle lines. Botulinum toxin A is currently being used for eliminating facial hyperkinetic wrinkles by causing paralysis of the underlying mimetic muscles. Because there are some histologic differences between Asians and Caucasians, such as thick dermis and more abundant collagen fiber, etc., the chronic pulling by mimetic muscles on the skin is expected to affect facial wrinkles differently. Therefore, the purpose of this study was to determine the efficacy of botulinum toxin A injection in eliminating facial hyperkinetic wrinkle lines among Korean patients. This study included 38 patients and 59 injection sessions from January of 1996 to April of 1997. We used Botox containing 100 U. Toxin was diluted with 4 ml of sterile normal saline and yielded 2.5 U for each 0.1 cc. A dose of 5 to 10 U was used in each muscle. Ages ranged from 26 to 56 years. There were 33 women and 5 men included in this study. Thirty-two of the patients were followed from 3 months up to 12 months after injections. The number of injection sessions that were performed on each patient was as follows: one session, 23 patients; two sessions, 10 patients; three sessions, 4 patients; four sessions, 1 patient. The number of injections per target site among these 38 patients was as follows: lateral canthal area, 33; glabellar area, 9; forehead, 9; nasal dorsum, 5. The most common duration of effective response was about 4 months, but in eight patients the period was over 5 months. After the response, complete recovery took about 1 or 2 months. Two patients felt unsatisfied, 5 patients felt slightly improved, and 25 patients retained only a slight line and were satisfied with the results. None of the patients experienced complete removal of wrinkle lines. Adverse effects included altered facial looks or appearances, mild local swelling, and ecchymosis at the injection sites. No systemic side effects were noted. Based on these results, the injection of botulinum toxin A seems to be an effective method of eliminating wrinkle lines on the upper third of the face in Korean patients, and it was a simple and effective nonsurgical procedure. PMID- 10697193 TI - External ultrasonic lipoplasty: an effective method of fat removal and skin shrinkage. AB - External ultrasonic lipoplasty is an effective method for the removal of localized fat and the fat due to moderate obesity. The ultrasound is externally applied and transmitted through the skin surface. The acoustic waves are selectively absorbed by previously injected tumescent fluid and fat. It is the combination of this acoustic wave and ultrasound-induced fluid streaming that facilitates fat removal. Multiple, 2.5- to 3.7-mm incisions are made, through which standard thin (2.3 to 3.5 mm) suction lipectomy cannulae are used to aspirate the emulsified fat and oil. A major advantage of this procedure is that superficial subdermal liposuction can be used safely, which enhances the thoroughness of fat removal and the contraction of the overlying skin. A total of 160 consecutive patients successfully underwent this procedure. Recovery was rapid, and patients returned to full activities within 24 to 48 hours. The skin remained soft, with minimal to no bruising throughout the entire postoperative period. The problems seen with internal ultrasonic liposuction, such as end hits and skin burns, were avoided. The large incisions required for internal ultrasound liposuction were not necessary. Previous models of externally applied ultrasound support current observations of the safety of external ultrasound lipoplasty. Only one small seroma was seen. External ultrasound lipoplasty is a safe, effective, and low-cost method of ultrasound-assisted removal of localized fat and the fat due to moderate obesity. Physician and patient satisfaction is high. PMID- 10697195 TI - Conflicts of interest in medical writing and the concept of disclosure. PMID- 10697194 TI - Crushed cartilage grafts over alar dome reduction in open rhinoplasty. PMID- 10697196 TI - Extracranial diffuse neurofibroma with intracranial extension. PMID- 10697197 TI - Transconjunctival upper blepharoplasty. PMID- 10697198 TI - Transpalpebral approach to the corrugator supercilii and procerus muscles. PMID- 10697199 TI - A new reshaped nostril retainer. PMID- 10697200 TI - Asymmetric incision for open rhinoplasty in cleft lip nasal deformity. PMID- 10697201 TI - Problems in the assessment of results after surgery for cleft lip nasal deformity. PMID- 10697202 TI - Sensory recovery in latissimus dorsi transplant after joining the thoracodorsal nerve to sensory nerves. PMID- 10697203 TI - Removal of lipomas by liposuction. PMID- 10697204 TI - The use of composite biodegradable skin graft and artificial skin for burn reconstruction. PMID- 10697205 TI - Successful use of the ultrapulse CO2 laser for the deepithelialization of TRAM flaps in breast reconstruction. PMID- 10697206 TI - Postoperative noise in breast implants. PMID- 10697207 TI - Postoperative "slosh". PMID- 10697208 TI - Explantation. PMID- 10697209 TI - A plea for more science regarding articles on breast implant capsular contracture. PMID- 10697210 TI - Operating on cubital tunnel syndrome without electrodiagnostics. PMID- 10697211 TI - Operating on cubital tunnel syndrome without electrodiagnostics. PMID- 10697213 TI - Use of disposable tubing in suction-assisted lipectomy. PMID- 10697212 TI - Local subcutaneous heparin as treatment for venous insufficiency in replanted digits. PMID- 10697214 TI - Skim milk masquerades as cream. PMID- 10697215 TI - Blood supply to the gluteus maximus flap. PMID- 10697216 TI - Embryology of corpus spongiosum and glans penis in hypospadias. PMID- 10697217 TI - Expression of alternative flap designs derived by different combinations of distal foot parts. PMID- 10697218 TI - Prophylactic antibiotics in plastic and reconstructive surgery. PMID- 10697219 TI - Medical wars: fighting corporate socialized medicine. PMID- 10697220 TI - The regulation of MR examinations in Germany: a threat to scientific and technical progress for MR in Europe? PMID- 10697221 TI - Application of LCModel for quality control and quantitative in vivo 1H MR spectroscopy by short echo time STEAM sequence. AB - The linear combination of model spectra (LCModel) calculation of a parameter for long-term quality control, kT, was introduced, representing the ratio of the temporal and nominal intensities of CH3 groups of lactate and acetate in a quality control phantom. This procedure is a part of the quality assurance of the scanner using fully automatic measurement and calculation of kT parameters, and utilizing Shewhart regulation control charts for continuous evaluation of the magnetic resonance (MR) scanner setting. The application of the kT parameter for the correction of in vivo data increases the precision of molar concentration determination by about 4%. This was tested by the quantitative in vivo MR determination of the molar concentrations of 13 prominent metabolites (N acetylaspartate (NAA), N-acetylaspartylglutamate, creatine and phosphocreatine (Cr), choline-containing compounds (Cho), myo-inositol, scyllo-inositol, gamma aminobutyric acid, glutamine, glutamate, glucose, lactate, alanine, taurine) in the white matter and hippocampus of the brain in groups of volunteers, using a short echo time stimulated echo acquisition mode sequence (echo time = 10 ms) and the LCModel technique. The repeatability of the measurement of prominent metabolites such as NAA, Cr and Cho was found to be around 10% (relative standard deviation, n = 6); precision in a group of volunteers (n = 20 and 28, respectively) was in the range of approximately 13-20%. For other metabolites, which are measured with a lower signal-to-noise ratio, the precision can be much lower. PMID- 10697222 TI - Assessment of prosthetic aortic valve performance by magnetic resonance velocity imaging. AB - OBJECTIVES: Magnetic resonance (MRI) velocity mapping was used to evaluate non invasively the flow profiles of the ascending aorta in normal volunteers and in patients with an aortic (mechanical) valve prosthesis. BACKGROUND: In patients with artificial aortic valves the flow profile in the ascending aorta is severely altered. These changes have been associated with an increased risk of thrombus formation and mechanical hemolysis. METHODS: Velocity profiles were determined 30 mm distal to the aortic valve in six healthy volunteers and seven patients with aortic valve replacement (replacement within the last 2 years) using ECG triggered phase contrast MRI. Peak flow, mean flow and mean reverse flow were measured in intervals of 25 ms during the entire heart cycle. Systolic reverse flow, end-systolic closing and diastolic leakage volume were calculated for all subjects. RESULTS: Peak flow velocity during mid-systole was significantly higher in patients with valvular prosthesis than in normals (mean + SD, 1.9 +/- 0.4 m/s vs. 1.2 +/- 0.03 m/s, P < 0.001) with a double peak and a zone of reversed flow close to the inner (left lateral) wall of the ascending aorta of the patients. Closing volume was significantly larger in patients than in controls (-3.3 +/- 1.2 ml/beat vs. -0.9 +/- 0.5 ml/beat; P < 0.001). There was reverse flow during systole in valvular patients amounting to 15.7 +/- 6.7% of total cardiac output compared to 2.3 +/- 1.2% in controls (P < 0.001). Diastolic mean flow was negative in patients after valve replacement but not in controls (-11.0 +/- 15.2 ml/beat vs. 6.8 +/- 3.2 ml/beat; P < 0.01). CONCLUSIONS: The following three major quantitative observations have been made in the present study: (1) Mechanical valve prostheses have an increased peak flow velocity with a systolic reverse flow at the inner (left lateral) wall of the ascending aorta. (2) A double peak flow velocity pattern can be observed in patients with bileaflet (mechanical) prosthesis. (3) The blood volume required for leaflet closure and the diastolic leakage blood volume are significantly higher for the examined bileaflet valve than for native heart valves. PMID- 10697223 TI - Assessment of arterial blood flow characteristics in normal and atherosclerotic vessels with the fast Fourier flow method. AB - The purpose of this study was to scrutinize the ability of magnetic resonance imaging (MRI)-performed measurements to compare arterial flow patterns in patients with peripheral arterial occlusive disease (PAOD), healthy volunteers (HV) and endurance athletes (EA). MRI blood flow data were partially repeated with Doppler ultrasound (DUS) with a view to a methodical comparison. Additionally, pulse wave velocity was assessed with the MUFF technique. For this purpose, MRI-performed flow measurements were performed in the common femoral artery in 21 patients with PAOD, in 34 HV and in 12 EA. The analysis included maximum flow velocities (MFV), velocity/time profile (VTP), pulse wave velocity (Vpulse), and vessel diameter (VD). In addition, MFV and VD were observed by DUS in most individuals. The results revealed a significant change regarding arterial blood flow characteristics in patients compared with HV and EA, with respect to the span between the peak positive and negative blood flow velocity in the femoral artery. The pulse wave velocity in patients was markedly elevated compared with healthy individuals. Furthermore, a complete, characteristic change in the VTP could be observed in patients. The methodical comparison between DUS and MRI showed a good correlation. Multi-slice Fourier flow data have indicated markedly increased pulse wave velocity in PAOD patients. Changes in the arterial blood flow can be clearly observed with MRI. In the future, this might offer a noninvasive possibility not only for the evaluation of the stage of the disease, but also for the detection of early, pre-clinical stages of atherosclerosis. PMID- 10697224 TI - Real-time volume rendering of MRCP: clinical applications. AB - MR-cholangiopancreatography (Signa Contour 0.5T; GE/Medical Systems, Milwaukee, WI) data sets of 156 patients, obtained with a 2D T2-weighted FSE sequence, in the coronal plane, were volume rendered (Advantage Windows 3.1; GEMS) independently by two radiologists, that were asked to define the range of signal intensities in which the signal of the pancreaticobiliary system was included and to rank the quality of native images and volume renderings. Patients had biliary stones (n = 47), inflammatory ampullary stenoses (n = 18), pancreatic tumors (n = 12), surgical bilio-enteric anastomoses (n = 19), ampullary carcinomas (n = 2), pancreatic duct stone (n = 1), cholangiocarcinoma (n = 3) and normal pancreaticobiliary tree (n = 54). Good quality volume renderings of the bile ducts were obtained for at least a maximum diameter of 1.5 mm. The quality rank agreement between volume rendering and native images was excellent (k = 0.94). The correlation between the observers for the setting the signal intensity range was excellent and statistically significant (P < 0.001). The correlation between the observers for the time of volume rendering was not statistically significant. Biliary stones could be displayed in 32/47 (68%) cases. The pancreatic duct stones was displayed as well. Inflammatory ampullary stenoses were detected in all cases (100%). In case of pancreatic tumors, cholangiocarcinomas and ampullary carcinomas volume rendering allowed to identify the site of stenosis. In surgical bilio-enteric anastomoses volume rendering was helpful to display the residual biliary tract, the site of anastomosis and the enteric tract. Volume rendering could be a reliable and efficient tool for the study of the anatomy and pathological changes of the pancreaticobiliary tract. PMID- 10697225 TI - Liposomes as fatty acids carriers in isolated rat liver: effect on energy metabolism and on isolated mitochondria activity. AB - The effects of fatty acids (FA)-carrier, egg-lecithin liposomes (LIPO) as alternative to BSA, on ATP, glycogen and glucose contents in isolated perfused liver of fed rats were non-invasively studied using 31P/13C nuclear magnetic resonance (NMR). Oxidative phosphorylation was studied in isolated mitochondria from the same liver consecutively to the NMR experiments. ATP content decreased slowly and ATP turnover was similar during the perfusion with saline solution (KHB) or LIPO. However, LIPO induced an enhancement of respiratory control ratio in isolated mitochondria. Tissue glycogen and glucose content decreased when FA (linoleate or linolenate) were perfused with defatted BSA (3%) or LIPO (600 mg/l) whereas glucose excretion level was unchanged and lactate excretion tended to increase, reflecting changes in the cytosolic redox state and/or an enhancement of glycolysis. Addition of FA (0.5 or 1.5 mM) to LIPO caused a dramatic fall in liver ATP, a mitochondrial uncoupling and an impairment of the phosphorylation activity. Perfusion with FA (1.5 mM) carried by BSA significantly increased the ATP degradation without change of mitochondrial function. Owing to the higher affinity of BSA than LIPO for FA, these latter could be more easily released from complex LIPO-FA, increasing their uncoupling effect. Hence, the FA concentrations have to be largely decreased from the above currently used concentrations to avoid this effect. It will then be possible to minimize the effector action of FA and to study their more specific metabolic function as fuel. It was concluded that LIPO were appropriate carriers to study the different metabolic effects of FA. PMID- 10697226 TI - Is absolute noninvasive temperature measurement by the Pr[MOE-DO3A] complex feasible. AB - Recently, the feasibility of the praseodymium complex of 10-(2-methoxyethyl) 1,4,7,10-tetraaza-cyclododecane-1,4,7-tr iacetate (Pr[MOE-DO3A]) for non-invasive temperature measurement via 1H spectroscopy has been demonstrated. Particularly the suitability of the complex for non-invasive temperature measurements including in vivo spectroscopy without spatial resolution as well as first spectroscopic imaging measurements at low temporal resolution (> or = 4 min) and high temporal resolution (breath hold, approximately 20 s) has been shown. As of today, calibration curves according to the particular experimental conditions are necessary. This work aims to clarify whether the Pr[MOE-DO3A] probe in conjunction with 1H-NMR spectroscopy allows non-invasive absolute temperature measurements with high accuracy. The measurement results from two different representative media, distilled water and human plasma, show a slight but significant dependence of the calibration curves on the surrounding medium. Calibration curves in water and plasma were derived for the temperature dependence of the chemical shift difference (F) between Pr[MOE-DO3A]'s OCH3 and water with F = -(27.53 +/- 0.04) + (0.125 +/- 0.001) x T and F = -(27.61 +/- 0.02) + (0.129 +/- 0.001) x T, respectively, with F in ppm and T in degrees C. However, the differences are minuscule even for the highest spectral resolution of 0.001 ppm/pt, so that they are indistinguishable under practical conditions. The estimated temperature errors are +/- 0.18 degrees C for water and +/- 0.14 degrees C for plasma and with that only slightly worse than the measurement accuracy of the fiber-optical temperature probe (+/- 0.1 degrees C). It can be concluded that the results obtained indicate the feasibility of the 1H spectroscopy method in conjunction with the Pr[MOE-DO3A] probe for absolute temperature measurements, with a maximum accuracy of +/- 0.2 degrees C. PMID- 10697227 TI - Posterior interbody fusion using laminectomy bone and transpedicular screw fixation in the treatment of lumbar spondylolisthesis. AB - BACKGROUND: Laminectomy bone is used widely in posterolateral lumbar fusion, but not interbody fusion. No prospective evaluation of interbody fusion using bone grafts from the posterior neural arch in spondylolisthesis has been found in the literature. We prospectively studied series of patients operated on for lumbar spondylolisthesis to evaluate clinical improvement and bony fusion. METHODS: Forty-six patients were operated on for lumbar spondylolisthesis using a simplified one-stage posterior procedure. The whole mobile dorsal segment of the vertebral arch was taken out in one piece and the bone was used for interbody fusion. Fixation was performed with transpedicular screws and rods using transverse connectors. RESULTS: After an average follow-up time of 27.3 months, 87% of the patients could be considered to have an excellent or good clinical outcome. The rate of successful fusion was 95.7%. No noteworthy complications occurred. CONCLUSION: Laminectomy bone seems to be optimal for posterior interbody fusion and together with transpedicular rigid fixation the long-term clinical and radiological results are convincingly good. The method is advisable even for severe spondylolisthesis. PMID- 10697228 TI - Increased MRI signal intensity in association with myelopathy and cervical instability: case report and review of the literature. AB - BACKGROUND: Increased T2-weighted signal intensity in patients with cervical myelopathy has been extensively reviewed in the literature. A variety of etiologies with similar MRI appearances have been described; attempt at correlation of MRI findings with clinical presentation and outcomes after treatment has led to a limited consensus. METHODS: We present a case of cervical myelopathy with associated hyperintense T2-weighted signal characteristics, secondary to cervical spondylosis and instability. RESULTS: Rapid resolution of radiographic abnormalities after surgical decompression and fusion was noted. Clinical improvement did not parallel radiographic resolution. CONCLUSION: These findings are important in considering the pathophysiology of MRI changes in cervical myelopathy. PMID- 10697229 TI - Elective neck clipping for unruptured aneurysms in elderly patients. AB - BACKGROUND: With the recent advancements of neuroimaging techniques, the number of unruptured aneurysms diagnosed in elderly patients has increased. However, the surgical indications in this special subgroup have not been studied critically. The purposes of this study were to analyze the results of elective neck clipping surgery for unruptured aneurysms in the elderly and to elucidate the surgical indications. METHODS: From 1985 to 1997, 96 patients, aged 70 years or older, with 103 unruptured cerebral aneurysms underwent elective neck clipping. There were 67 females and 29 males. Their ages ranged from 70 to 86, with a mean of 73.3 years. Seventy-five aneurysms were asymptomatic and 28 were symptomatic. The aneurysms were located on the internal carotid artery (46.6%), middle cerebral artery (35.9%), anterior cerebral artery (16.5%), and basilar artery (1.0%). RESULTS: The surgical outcome was a good recovery in 75 patients (78.1%), mild deficits in 12 (12.5%), severe deficits in 4 (4.2%), and death in 5 (5.2%). Recovery from preoperative symptoms with improved quality of life was seen in 22 (78.6%) of the 28 symptomatic cases. Multiple regression analysis showed that increase in the size of aneurysms and location on the middle cerebral artery and internal carotid artery were significantly related to a poor outcome. The causes of the five deaths were hemorrhagic infarction, systemic infection, and myocardial infarction. CONCLUSION: Surgery for elective neck clipping of unruptured aneurysms in the elderly should be considered in symptomatic patients with simple aneurysms that can be clipped without the use of temporary clips. PMID- 10697230 TI - Risks of surgery for patients with unruptured intracranial aneurysms. AB - BACKGROUND: With the widespread use of less invasive imaging tools, such as magnetic resonance angiography and computed tomographic angiography, unruptured cerebral aneurysms are found much more often than in the past. This retrospective study was undertaken to determine the risk factors for surgical intervention in a patient with an unruptured intracranial aneurysm. METHODS: Over a 5-year period, 1,558 patients with intracranial aneurysms underwent surgery at our center. Of these, 310 patients (20%) with unruptured aneurysms were included in this study. RESULTS: Out of 310 patients with unruptured aneurysms, 292 (95%) had a favorable outcome, and only one patient (0.3%) with a giant vertebral artery aneurysm died. Aneurysm size larger than 15 mm and location of the aneurysm in the posterior circulation were independent risk factors associated with less favorable outcomes. Patients with a single aneurysm had a better outcome than did patients with multiple aneurysms. CONCLUSION: Our results support the contention that surgical treatment of unruptured intracranial aneurysms carries a low risk of morbidity and mortality and may improve the outcome in patients harboring cerebral aneurysms by preventing the devastating effects of subarachnoid hemorrhage. Aneurysm size, location, and number were risk predictors for surgical morbidity in patients with unruptured aneurysms. PMID- 10697231 TI - Thalamic cavernous malformations. AB - BACKGROUND: Only few anecdotal reports and small series of thalamic cavernous malformations have been reported. It follows that the clinical behavior and management are poorly understood; in particular, experiences with the surgical treatment of these lesions are scarce. METHODS: The clinical course, treatment, and outcome of 12 patients (10 females and 2 males, mean age 36 years) with symptomatic cavernous malformations of the thalamus are reviewed. Eight patients (66%) presented with cerebral hemorrhage, one with progressive neurological deficit and three with hydrocephalus/increased intracranial pressure; associated venous anomalies were found in three cases. Treatment consisted of radical surgery in four cases with progressive neurological decline or recurrent disabling hemorrhage, radiosurgery (one case), evacuation of a chronic satellite hematoma (one case), ventriculoperitoneal shunt for hydrocephalus (one case) and observation (five cases). Operative treatment in four cases included transcallosal, trigonal, and occipital interhemispheric approaches. RESULTS: In the surgical group, one patient died, two improved after operation, and one remained the same. Of the patients not operated on radically, one had recurrent hemorrhage 4 months after radiosurgery, one remains stable 8 years after ventriculoperitoneal shunt, and one 3 years after aspiration of a satellite hematoma. Five other patients presenting with thalamic hemorrhage were treated conservatively; recurrent hemorrhage occurred in two cases at 1 month and at 2 years, leaving a mild residual deficit in both cases. Overall, rehemorrhage occurred in four cases (50%) at a mean interval of 18 months after the first bleeding; the annual hemorrhage rate was 6.1%. CONCLUSIONS: Thalamic malformations are more likely to be symptomatic from small hemorrhages compared with lesions in the cerebral hemispheres; progressive growth may also occur with third ventricle invasion or caudal extension to the midbrain. Their high-risk location deters heavy-handed management, but they should not be left long untreated. Both surgery and radiosurgery have been used in the management of thalamic cavernomas reported in the literature. Definite surgical indications include progressive neurological decline and recurrent hemorrhages of malformations abutting the ventricular surface or the posterior incisural space; the complex anatomy of the deep venous system and the association with unexpected venous anomalies complicates the removal of these lesions. PMID- 10697232 TI - Surgical treatment of primary midbrain gliomas. AB - BACKGROUND: The treatment of brainstem gliomas remains controversial. This article focuses on surgical results. METHODS: The authors retrospectively analyzed 35 patients with primary midbrain gliomas who were treated at Beijing Neurosurgical Institute from 1986 to 1997. The diagnosis was verified by histological examination. RESULT: The incidence of midbrain glioma was 10.3% (35/340) in our patients with brain stem tumors. The 35 gliomas were classified into three therapeutic groups by their locations: 7 were located in the tectal region, 8 in the aqueductal region, and 20 in the tegmental region. All of the patients underwent microsurgical treatment based on a minimally invasive approach. The operation took the form of total resection in 19 cases, subtotal resection in 12, and partial resection in 4. The operative mortality was 0. With a mean follow-up of 28 months (range, 6-48 months), 65.7% (23/35) of patients could live independently. CONCLUSION: The volume and location of midbrain tumors were highly correlated with outcome. The resection of as much tumor as possible was optimal for the treatment of midbrain gliomas and radiotherapy after operation was beneficial to patients. PMID- 10697233 TI - Selection of surgical approaches for small acoustic neurinomas. AB - BACKGROUND: The purpose of this study was to evaluate the results of surgery for small acoustic neurinomas at our institute via the middle cranial fossa and retrosigmoid approaches, and to determine the indications for each approach. METHODS: Fifty-three patients with unilateral tumors less than 2 cm in diameter were studied. Surgery was performed via the middle cranial fossa approach in 36 tumors and via the retrosigmoid approach in 17 tumors. RESULTS: The hearing preservation rate was 68% (36/53) in all patients, 93% (14/15) in patients with intracanalicular tumors, 79% (15/19) in patients with tumors less than 1 cm in diameter, and 43% (7/19) in patients with tumors between 1 and 2 cm in diameter. The facial nerve function was excellent or good in 80% (42/53), 74% (11/15), 84% (16/19), and 78% (15/19), respectively. Among the 19 patients with tumors between 1 and 2 cm in diameter, the frequencies of hearing preservation and of excellent or good facial nerve function (47% and 87%, respectively) in the 15 patients approached via the retrosigmoid approach were higher than those (0% and 50%, respectively) in the four patients approached via the middle cranial fossa approach. CONCLUSIONS: We conclude that tumors smaller than 2 cm should be removed because preservation of hearing as well as facial nerve function may be possible in almost all of these patients. Tumors larger than 1 cm should be surgically treated through the retrosigmoid approach. PMID- 10697234 TI - Calvarial metastasis of a paraganglioma. Case report and review of the literature. AB - BACKGROUND: Metastasis of a paraganglioma (PRG) to the calvarium is very rare. In this paper, the case of a 25-year-old male with metastasis of a PRG to the frontoparietal bone is described. CASE DESCRIPTION: The patient presented with bulging on the left side of the head, headache, and weight loss. Magnetic resonance imaging (MRI) revealed a mass lesion in the left frontoparietal region that had destroyed both the external and internal table of the bone, extending under the skin and above the dura mater. After a frontoparietal craniotomy the tumor was removed totally. Histopathological examination revealed the "Zellballen," which are pathognomonic for a PRG. Systemic examination and radiological investigation revealed no primary tumor source. CONCLUSION: Metastasis of a PRG to the calvarium is possible; radical removal of the tumor will provide a cure. PMID- 10697235 TI - A keyhole middle fossa approach to large cholesterol granulomas of the petrous apex. AB - OBJECTIVE: In this article we review our surgical experience in a series of eight patients with large cholesterol granulomas of the petrous apex extending into the cerebellopontine angle. METHODS: All lesions, four primary and four recurrent, were studied with magnetic resonance imaging (MRI), and computed tomography (CT). The patients underwent pre- and postoperative audiographic testing. A keyhole middle fossa approach was used in all cases. RESULTS: There was no mortality. Surgery was complicated in one case by a subgaleal hematoma and in another by a transitory increase of a preexisting facial palsy. In five cases the granuloma was totally resected, whereas in the remaining three small remnants of the pseudocapsule were left in place. At follow-up (12-90 months), three patients were asymptomatic. In the remaining five patients, trigeminal neuralgias had subsided. Palsies of the VIth cranial nerve recovered more consistently than those of the VIIth. Hearing was unchanged postoperatively. So far, there has been no clinical or radiological evidence of a recurrence. CONCLUSION: Large cholesterol granulomas of the petrous apex can be effectively treated through a keyhole middle fossa approach. Despite its contained size the approach allows a rather large exposure of the granuloma. The resection of these lesions carries a low risk of compromising the facial or hearing function of the patient. Small remnants of the capsule, left in place to avoid potential complications, seem not to affect the long-term outcome of the patients, provided the cavity in the petrous bone is adequately ventilated. PMID- 10697236 TI - Cerebral atrophy in Cushing's disease. AB - BACKGROUND: Cushing's disease causes significant pathological changes throughout the body as a result of elevated cortisol levels. Very few systematic investigations have focused on the morphologic effects of hypercortisolism on the central nervous system. The validity of using premature cerebral atrophy as a diagnostic tool for Cushing's disease remains unknown. METHODS: This study includes 63 patients with Cushing's disease who were evaluated and treated at the University of Virginia Medical Center. Radiologists randomly compared these individuals with age- and sex-matched controls in a blinded protocol, assessing the degree of cerebral atrophy on computed tomography and magnetic resonance scans. RESULTS: Patients with Cushing's disease showed significant premature atrophy when compared with controls. This trend continued after subdividing the groups based on age and duration of symptoms except in the following groups: age greater than 60, duration of symptoms less than 1 year, and symptoms lasting between 4-5 years. CONCLUSIONS: Excluding the three aforementioned groups, the hypercortisolemic state manifested in patients with Cushing's disease promotes the premature development of cerebral atrophy, which can be identified on routine radiologic imaging and may assist in the clinical diagnosis of the condition. PMID- 10697237 TI - Absence of peritumoral Crooke's change is associated with recurrence in surgically treated Cushing's disease. AB - BACKGROUND: Pituitary surgery is the standard treatment for Cushing's disease but is complicated by a recurrence rate that ranges from 5.9 to 27%. Whereas some recurrences may be due to technical or anatomical factors resulting in subtotal resection of adenoma, clinical relapse after total tumor resection is a well documented occurrence. The factors leading to such recurrences are unknown. METHODS: With the hypothesis that the pathology of the nontumoral adenohypophysis is important in predicting relapse, we undertook a study to determine if the absence of Crooke's change (CC), thought to be an indicator of nontumoral corticotroph inhibition, was associated with unexpected clinical recurrence. Twenty-one patients with Cushing's disease, with gross total resection of intrasellar corticotroph microadenoma, were reviewed independently by 2 neuropathologists for the presence of CC in adjacent adenohypophysis. All tumors were stained with H&E, PAS/orange-G and immunohistochemistry for ACTH. Clinical relapse was determined by chart reviews and defined as serum ACTH > 15 pg/ml, clinically Cushingoid, and/or radiographic evidence of recurrent tumor. RESULTS: Seven of 21 patients recurred; 3 did not have CC in their initial resection specimen. All 3 of these patients had unexpected recurrences at 6 to 48 months post-op. Two patients with CC recurred at one year follow-up, 1 after 4 years and 1 after 5 years. All specimens from patients with long-term cure (follow-up from 9-72 months) contained CC. In this study, the absence of CC in peritumoral adenohypophysis was associated with unexpected recurrence of Cushing's disease (p = 0.0214). CONCLUSIONS: We conclude that absence of CC in peritumoral adenohypophysis may be of some assistance in predicting recurrence of Cushing's disease after adequate resection of intrasellar microadenoma. PMID- 10697238 TI - Neurosurgery of the peripheral nervous system: entrapment syndromes of the brachial plexus. PMID- 10697239 TI - Multiple brain abscesses caused by Salmonella typhi: case report. AB - BACKGROUND: Focal intracranial infections caused by Salmonella species are uncommon. The authors report a case of multiple brain abscesses caused by Salmonella typhi. CASE DESCRIPTION: A 2-month-old girl was admitted to the hospital because of diarrhea, vomiting, fever, and poor feeding. Neurological examination revealed cervical hyperextension and absence of sucking and Moro reflexes. During the next 20 hours she developed complex partial seizures with secondary generalization and alternated irritability with drowsiness. Investigation showed hemoglobin 6.3 g/dl; white blood cell count of 19500/mm3 with a marked shift to the left. The analysis of the cerebrospinal fluid revealed white cell count of 1695/mm3, lymphocytes 61%, protein 300 mg/dl and glucose 6 mg/dl. The patient was treated for acute gastroenterocolitis, sepsis, and meningitis. Blood culture taken on the day of admission showed gram-negative bacilli, later identified as S. typhi. Computed tomography scan demonstrated a lesion in the right parietal lobe compatible with a brain abscess. Follow-up computed tomography after 7 days showed several other lesions with the same features. Surgical drainage of the right parietal lesion was performed on the 13th day, through a burr hole. The patient was discharged 5 weeks after admission without neurological deficit. CONCLUSION: Bacteremia, sepsis, and meningitis are relatively common in children with Salmonella infection but intracranial abscesses are very rare. Surgical drainage combined with prolonged antibiotic therapy (drug of choice: chloramphenicol) is the best treatment for Salmonella brain abscesses. The possibility of intracranial infection should be considered in patients with Salmonellosis and neurological dysfunction. PMID- 10697240 TI - Presphenoidal marrow changes in the pediatric skull base. PMID- 10697241 TI - Thoughts about Professor Ramamurthi and senior neurosurgeons: when should they retire? PMID- 10697242 TI - Horner's syndrome and sixth nerve palsy. PMID- 10697243 TI - NF2 in monozygotic twins. PMID- 10697244 TI - NF2 in monozygotic twins. PMID- 10697246 TI - Changes in mRNA of protein inhibitor of neuronal nitric oxide synthase following facial nerve transection. AB - Protein inhibitor of neuronal nitric oxide synthase (PIN) is reported as the protein inhibiting neuronal nitric oxide synthase (nNOS) activity by preventing dimerization of nNOS. It was also reported that PIN inhibits the activity of all nitric oxide synthase (NOS) isozymes. We examined the effects of facial nerve transection on PIN mRNA and NOS expression by in situ hybridization for PIN mRNA and nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d) staining. PIN mRNA was initially expressed and transiently increased from 3 to 5 days and returned to the basal level at 7 days after axotomy in the motoneurons of the facial nucleus. NADPH-d-positive motoneurons were found from 7 days post operation in the facial nucleus. These results suggest that PIN may interact with NOS from 7 days post-operation. PMID- 10697245 TI - Constitutive and inducible nitric oxide synthases after dynorphin-induced spinal cord injury. AB - It has recently been demonstrated that selective inhibition of both neuronal constitutive and inducible nitric oxide synthases (ncNOS and iNOS) is neuroprotective in a model of dynorphin (Dyn) A(1-17)-induced spinal cord injury. In the present study, various methods including the conversion of 3H-L-arginine to 3H-citrulline, immunohistochemistry and in situ hybridization are employed to determine the temporal profiles of the enzymatic activities, immunoreactivities, and mRNA expression for both ncNOS and iNOS after intrathecal injection of a neurotoxic dose (20 nmol) of Dyn A(1-17). The expression of ncNOS immunoreactivity and mRNA increased as early as 30 min after injection and persisted for 1-4 h. At 24-48 h, the number of ncNOS positive cells remained elevated while most neurons died. The cNOS enzymatic activity in the ventral spinal cord also significantly increased at 30 min 48 h, but no significant changes in the dorsal spinal cord were observed. However, iNOS mRNA expression increased later at 2 h, iNOS immunoreactivity and enzymatic activity increased later at 4 h and persisted for 24-48 h after injection of 20 nmol Dyn A(1-17). These results indicate that both ncNOS and iNOS are associated with Dyn-induced spinal cord injury, with ncNOS predominantly involved at an early stage and iNOS at a later stage. PMID- 10697247 TI - A morphological analysis of the motor neuron degeneration and microglial reaction in acute and chronic in vivo aluminum chloride neurotoxicity. AB - The monthly intracisternal inoculation of aluminum chloride (AlCl3) to young adult New Zealand white rabbits induces motor neuron degeneration marked by intraneuronal neurofilamentous aggregates similar to that observed in amyotrophic lateral sclerosis (ALS). However, in contrast to ALS, this process occurs in the experimental paradigm in the absence of a glial response. In addition, whereas ALS is a fatal disorder, the cessation of aluminum exposure leads to both clinical and neuropathological recovery. Because microglia can influence neuronal regeneration, we have examined the effect of both acute and chronic aluminum exposure on microglial activation in vivo. We have studied microglial morphology in young adult New Zealand white rabbits receiving either single (1000 microg) or repeated sublethal (100 microg monthly) intracisternal inoculums of AlCl3. In addition, rabbits receiving 1000 microg AlCl3 inoculums were studied following an unilateral sciatic axotomy 48 h prior to the AlCl3 exposure. Our studies demonstrate that microglial activation in vivo is inhibited by AlCl3 exposure, and that a correlation exists between the extent of microglia suppression and the potential for recovery. This suggests that microglial activation is an important determinant of neuronal injury. PMID- 10697248 TI - A protein encoded by an alternative splice variant of choline acetyltransferase mRNA is localized preferentially in peripheral nerve cells and fibers. AB - Central cholinergic systems have been visualized by immunohistochemistry using antibodies to choline acetyltransferase (ChAT). Peripheral cholinergic cells and fibers, however, have been hardly detectable with most of these antibodies. This phenomenon suggests that a different form of ChAT may exist in peripheral tissues. Here we report two types of mRNA for ChAT expressed by alternative splicing in rat pterygopalatine ganglion. One is exactly identical with ChAT mRNA reported in the central nervous system (ChAT of a common type; cChAT). The other lacks exons 6, 7, 8 and 9, which was detected only in the pterygopalatine ganglion (ChAT of a peripheral type; pChAT). The peculiarity of pChAT in chemical structure, possessing a splice joint of the exons 5 and 10, led us to produce rabbit antisera against a recombinant peptide of 41 amino acids which spans over the splice joint. On Western blots using a successfully obtained antiserum, an intense band of about 50 kDa, corresponding to the expected molecular weight of pChAT, was detected in the pterygopalatine ganglion but not in the brain. Immunohistochemistry using the antiserum failed to reveal positive staining of known brain cholinergic structures, while it permitted us to observe peripheral, probably cholinergic, nerve cells and fibers including those in the pterygopalatine ganglion and enteric nervous system. PMID- 10697249 TI - The effect of electrolytic thalamic lesions on the NADPH-diaphorase activity of neurons of the laterodorsal tegmental and pedunculopontine nuclei in rats. AB - Cholinergic neurons of the mesopontine complex have extensive ascending projections to the forebrain: the laterodorsal tegmental nucleus extensively innervates the anterior thalamus, the anteroventral nucleus in particular, whereas the pedunculopontine nucleus has widespread projections to both the thalamus and extrapyramidal structures. Most of their neurons express nitric oxide synthase (NOS) activity. Following electrolytic lesions of the anteroventral thalamic nucleus, nicotinamide adenine dinucleotide phosphate diaphorase (NADPHd) activity in neurons of the laterodorsal tegmental nucleus changed drastically. The intensity of NADPH-diaphorase staining increased in laterodorsal tegmental neurons ipsilateral to the lesion side, but decreased contralaterally. The intensity of the NADPH-diaphorase staining of neurons of the pedunculopontine nucleus, however, remained unchanged bilaterally. After partial lesions of the anteroventral thalamic nucleus a similar effect was noted. In contrast, large electrolytic lesions involving other thalamic nuclei or extrapyramidal structures did not change the number of NADPH-diaphorase neurons or their intensity of staining in the laterodorsal tegmental nuclei. These data show that electrolytic lesions of target areas can lead to an upregulation of NOS expression in the parent cell bodies, provided that there is no wide collateralization as found for the pedunculopontine nucleus. PMID- 10697250 TI - Adult asthma prevalence, morbidity and mortality and their relationships with environmental and medical care factors in Singapore. AB - We have conducted a series of studies on adult asthma in Singapore that describe the prevalence, morbidity and mortality and their relationships with environmental and medical care factors. There was no evidence of a temporal increase of mortality from 1976 to 1995 for adults. The prevalence rate of asthma is 2.4% in men and 2.0% in women. There is considerable morbidity among asthmatics, corticosteroids are under-used, and patients' knowledge and self management skills is poor. Increased morbidity is significantly associated with current keeping of pets, current smoking, and the patients' knowledge and self management skills. Occupational exposure contributes up to a third of asthma morbidity. Malays and Indians have higher rates of asthma mortality and morbidity than Chinese. They have greater exposures to airborne allergens from keeping rugs or carpets, and pets. Malays experience the most morbidity from asthma, but make less use of health services, and receive less medical attention, than Indians or Chinese. PMID- 10697251 TI - The epidemiology of atopic dermatitis at a tertiary referral skin center in Singapore. AB - Atopic dermatitis is a common chronic, relapsing, pruritic ecematous skin condition with a predilection for the flexural areas and occurs in patients with a personal or family history of atopy. The aim of this study is to describe the profile of atopic dermatitis seen at the National Skin Centre in Singapore. A retrospective chart review was conducted of all the patients with atopic dermatitis seen during the first six months of 1994. There were 492 patients whose ages ranged from one month to 74 years with an equal sex ratio. The prevalence was 2%. The onset of the disease occurred before the age of 10 years in 61.2% of patients. In 13.6% of the patients, the onset was after the age of 21 years. Two hundred and fifty-four patients (52%) had "pure" atopic dermatitis without concomitant respiratory allergies. Two hundred and thirty-eight patients (48%) suffered from a "mixed" type, with 23% having allergic rhinitis, 12% having asthma and 13% having both asthma and allergic rhinitis. Two hundred and thirty one patients (47%) had at least one first-degree family member with atropy: atopic dermatitis (17%), asthma (15%) and allergic rhinitis (15%). Most of the patients, 416 (84.5%), had subacute eczema at presentation. Ichthyosis vulgaris was present in 38 patients (8%) and pityriasis alba in 13 patients (3%). The most common infective complication was bacterial infection (impetiginized eczema, folliculitis, cellullitis) present in 95 patients (19%) followed by viral infections (eczema herpeticum, viral warts and molluscum contagiosum) in 17 patients (3%). Allergies were noted in 43 patients (9%) based on the history given. The most common was drug allergies (penicillin and co-trimoxazole) in 28 patients followed by food allergies in 11 patients. Common aggravating factors reported include heat, sweating, stress, thick clothing and grass intolerance. Most patients could be controlled with a fairly simple regimen of moisturizers, topical steroids and antibiotics for acute flares. Short courses of systemic steroids were used in 78 patients (16%). Three patients were treated with phototherapy, Two on UVAB and one on PUVA. The pattern of atopic dermatitis in Singapore is similar to that reported in the Western literature except for a lower prevalence and a significant proportion of adult onset atopic dermatitis. PMID- 10697252 TI - Factors associated with increased respiratory symptoms among asthmatic children in Singapore. AB - Asthma is a common cause of childhood morbidity. The objective of the present study was to evaluate the factors associated with increased asthma morbidity among asthmatic children in Singapore. A cohort of primary school children (n = 6,404, aged 6-13 years) were evaluated using the American Thoracic Society and the Division of Lung Diseases of the National Heart, Lung and Blood Institute, USA (ATS-DLD) respiratory questionnaire. A total of 2,222 of 6,404 children (34.8%) was found to have reported symptoms of wheezing. Of these, 899/2,222 (40.5%) reported symptoms of "increased asthma morbidity". This was associated with the younger age group, male sex and higher socio-economic status. In addition, concurrent or past allergies were strongly associated with increased asthma morbidity, while premature birth and a history of prior childhood respiratory illnesses and Infections were predictive of greater asthma morbidity. No association was found between increased morbidity and presence of domestic pets, parental smoking, childcare attendance, and the season of birth. PMID- 10697253 TI - Pediatric asthma quality of life questionnaire: validation in children from Singapore. AB - "Quality of Life" is a multidimensional measure encompassing the physical, emotional and social functioning of the child. The asthma specific questionnaire contains 23 questions (items) in three areas (domains) of activity, symptoms and emotions. The objective of the present study was to validate the Paediatric Asthma Quality of Life Questionnaire "PAQLQ"(copyright 1991 McMaster University). If the questionnaire is valid, a change in the child's asthma will be accompanied by a change in the "Quality of Life" questionnaire score. The questionnaire was administered twice over four weeks and the child's asthma status was assessed concurrently. Two groups were thus identified; Group A = unchanged asthma, Group B = changed asthma. Forty-seven children, aged 7 to 14 years, completed the study. Reliability of the questionnaire shows an intraclass-correlation coefficient of only 0.71. Cross-sectional construct validity was demonstrated by a significant correlation between the whole questionnaire and the clinical asthma score (p<0.001) but not in the separate domains. Longitudinal construct validity was also demonstrated by the significant correlation between change in the total questionnaire score, but not separate domains, with change in the child's asthma score (p<0.05). Responsiveness was shown by a significant difference in the magnitude of the change in the questionnaire score between the two groups (p<0.001), but again not in the separate domains. It was concluded that the questionnaire was validated as a whole but not in as convincing a manner, as has been done by others, and we are therefore in a position to advise caution in its application in our population. PMID- 10697254 TI - Quality of life of patients with perennial allergic rhinitis: preliminary validation of the Rhinoconjunctivitis Quality of Life Questionnaire in Singapore. AB - Though sufferers of perennial allergic rhinitis do not die from their ailment, they endure years of chronic nose disease that Interferes with many important aspects of their lives. A rhinitis-specific instrument to gauge the quality of life of patients with this disease was published in 1991. Here, we validated the Rhinoconjunctivitis Quality of Life Questionnaire (RQLQ) for use in English speaking patients with perennial allergic rhinitis. We established that the RQLQ distinguishes between patients and control, demonstrates internal consistency and is sensitive to change. This study suggests that the RQLQ can be used to assess the quality of life of patients with perennial allergic rhinitis in Singapore. PMID- 10697255 TI - An evaluation of mattress encasings and high efficiency particulate filters on asthma control in the tropics. AB - The effect of two allergen avoidance modalities, Allergy Control Covers (ACC) and High Efficiency Particulate Filters (HEPA) on asthma control in children were evaluated. This was an open study involving 24 dust mite sensitive asthmatic children. Following a 4 week run-in period, the subjects were randomly allocated to use mattresses fitted with ACC (n = 6), HEPA filters in their bedrooms (n = 12) or act as controls (n = 6) for a study duration of 4 months. Measurements of the major Dermatophagoides spp. mite allergens, Der p 1 and Der f 1, levels in dust samples obtained from mattresses were made at baseline, 1, 2 and 4 months post implementation. Daily symptom scores including morning and evening peak flow readings, and monthly spirometry and exercise bronchoprovocation tests were carried out Our results showed that dust mite allergen levels in mattresses fell at 1 and 2 months post implementation in the ACC group (p<0.05). In contrast, no decrease in allergen levels was seen in the HEPA and control group. At the end of the 16 weeks, only the ACC group showed improvement in FEV1 and reduction in diurnal peak expiratory flow rate (p<0.05). Improvement in mean symptom scores was also observed for both the ACC and HEPA groups, but not the control groups (p<0.05). Although the numbers in this study were small, the results Indicate that the effectiveness on mite exposure barrier covers was short-lived, and the improvement in asthma control though documented was not obvious. PMID- 10697256 TI - Healthy adults demonstrate less skin reactivity to commercial extracts of commonly ingested food than to D. farinae. AB - The aim of this study is to determine the skin reactivity of healthy Oriental adults to commercial extracts of commonly ingested food and the house dust mite D. farinae, a common local aeroallergen. D. farinae and 18 food extracts were skin prick tested on adults without any personal history of atopic diseases and food allergy. The extracts of food not consumed by any subject on religious or personal grounds were not tested for that individual. A total of 103 healthy adults who fulfilled the selection criteria were skin prick tested. There were 35 males and 68 females. Their mean age was 29 years (SD +/- 7.5) with a range of 19 to 49 years. Sixty-eight percent were Chinese, 12.6% Malay, 12.6% Indian and 6.8% other Oriental races. Fifty-four (52.4%) were positive for D. farinae while only 12 (11.7%) were positive for at least one food extract The food extract that gave the most number of positive reactions was shellfish mix (5/102, 4.9%). A family history of atopy did not have any significant correlation with the results of skin test. It was concluded that healthy adults demonstrate less skin reactivity to extracts of commonly ingested food than to D. farinae. PMID- 10697257 TI - Sensitization to Blomia tropicalis and dermatophagoides pteronyssinus-a comparative study between Singapore and Taiwan. AB - Blomia tropicalis (Bt) and Dermatophagoides pteronyssinus (Dp) are the predominant domestic mites species in Singapore and Taiwan. This study aims to characterize and compare the mite sensitization profiles in both countries. Skin prick tests were performed on 203 Singaporeans with Dp and Bt crude extracts. In vitro IgE and IgG4 reactivity to extracts and specific allergens (Der p 1, Der p 2 Der p 5 and Blo t 5) were determined by immunoassays. Approximately 91% of the tested Singaporeans were skin test positive for both Bt and Dp. Both populations share similar frequencies of in vitro IgE reactivity to all the allergens tested, but they differ in the pattern and magnitude of allergen sensitization. Although Der p 1, Der p 2 and Blo t5 are major sensitizing allergens in both countries, Blo t 5 is a more potent one in Singapore, probably reflecting the high level of exposure to Bt. The unique major Bt and Dp allergens should be included for precise diagnosis and effective immuno-therapeutic treatment of mite allergy in both countries. PMID- 10697258 TI - Culture of Blomia tropicalis and IgE immunoblot characterization of its allergenicity. AB - Blomia tropicalis is an important triggering factor for allergic diseases such as asthma, rhinitis and atopic dermatitis in tropical and subtropical regions, which climate favours the growth of this species. Our previous mite fauna study revealed that Blomia tropicalis is the most dominant species present in Singapore house dust The main objective of this study is to establish a mass culture of Blomia tropicalis for further characterization of the antigenic and molecular properties of this mite. Approximately one gram of mites could be obtained for every 300-gram of culture medium by culturing under natural condition with a mean annual temperature of 30 degrees C and a mean relative humidity of 80%, and harvested by modified Tullgren funnel. Allergen characterization by IgE immunoblot analysis with crude mite extracts showed some IgE reactivity differences between Blomia tropicalis mite extract from Singapore and Colombia. The possible reasons for these findings are the quality and source of the mite protein extracts used, or selective differences in the population under evaluation. Further, the atopic sera tested showed differences in the pattern and Intensity of IgE immunoblot reactivity to crude extracts of Blomia tropicalis and Dermatophagoides pteronyssinus, the other highly prevalent mite in Singapore. These data support the existence of species-specific allergens. In conclusion, we have been successful in setting up B. tropicalis mass cultures and have prepared extracts of high allergenicity. PMID- 10697259 TI - Asthma management: evidence based studies and their implications for cost efficacy. AB - This review attempts to infer a cost-effective strategy for the management of bronchial asthma based on evidence from randomized controlled trials. Acute severe asthma should be treated with short-acting inhaled beta-agonists followed by a short course of oral steroids. Decisions on hospital admission should be made within 1 to 2 hours and prolonged treatment in emergency departments avoided. A comprehensive educational and drug optimizing program will prevent chronic illness and relapse. Educational programs should be brief but intensive, supervised by asthma specialists and incorporate self monitoring of symptoms plus written action plans. Peak expiratory flow monitoring should not be mandated for all patients. Inhaled corticosteroids (ICS) are the most cost-effective drugs for the long term prevention of asthma. ICS should be started at low doses. If the symptoms of asthma are not well controlled by moderate doses of ICS, high dose ICS treatment should be avoided and add on medication prescribed instead. Oral bronchodilators are less expensive add on medication than long-acting inhaled beta-agonists. PMID- 10697260 TI - Food allergy in children-the Singapore story. AB - The study of food allergy in Singaporean children is still in its infancy. Confusion and misunderstanding is common among the public. Even so, we have found certain unique features regarding food allergy among Singaporean children. "Bird's nest" has been shown to be the most common cause of anaphylaxis requiring medical attention. This allergen has not been described before. Peanuts and tree nuts are extremely uncommon causes of anaphylaxis, unlike the West. However, the pattern of sensitization to foods in children as shown by skin prick test is similar to other Western populations. The reasons for the difference between the profile foods responsible for anaphylactic reactions in our population and those of the western population, despite the similarity in sensitization profiles, are still unclear. PMID- 10697261 TI - Contact allergy in Singapore. AB - Contact allergy, viz. allergic contact dermatitis, photo-allergic contact dermatitis and contact urticaria, is a well-studied sub-specialty of dermatology in Singapore. Over the years, numerous studies and anecdotal reports on the subject have been published in both international and local refereed journals. This article reviews the epidemiological data on patch testing and photo-patch testing in Singapore. It also summarizes published clinical reports on important contact allergens that are found in both non-occupational and occupational setting. PMID- 10697262 TI - The significance and technical aspects of quantitative measurements of inflammatory mediators in allergic rhinitis. AB - The pathophysiology of allergic rhinitis induced by various inhalant allergens through an IgE mediated mechanism, has been well demonstrated. The participation of many important inflammatory cells and mediators released by these cells in the human nasal allergic reaction provides insight into the relationship between the responsiveness to allergen exposure and nasal symptoms of allergic rhinitis. This paper summarizes our previous studies on some important mediators in the nasal secretions of atopic patients during different phases after nasal allergen challenge and during natural allergen exposure. The microsuction technique proves to be an especially useful and reliable nasal sampling method permitting quantitative analysis of important mediators such as histamine, tryptase, leukotriene C4 and eosinophil cationic protein in nasal secretions. The measurement of these mediators during allergic reactions provides accurate data on the activity of some important inflammatory cells (i.e., mast cells, basophils, and eosinophils) and their responses to therapy. PMID- 10697263 TI - Effects of inhibitors of the tyrosine kinase signaling cascade on an in vitro model of allergic airways. AB - It has been shown that activation of protein tyrosine kinases (PTKs) is the earliest detectable signaling response to FcepsilonRI cross-linking on mast cells. Following tyrosine kinase activation, a family of mitogen-activated protein kinases (MAPKs) was found to be activated as well. Activation of this PTK signaling cascade will lead to mast cell degranulation. This review summarizes our recent studies on the role of PTK signaling cascade in an in vitro guinea pig model of allergic asthma using PTK inhibitors, genistein and tyrphostin 47, and MAPK kinase inhibitor, PD098059. Inhibitors of the PTK and MAPK signaling pathways significantly attenuated the ovalbumin (OVA)-induced bronchial anaphylactic contraction and enhanced relaxation of constricted airways, respectively, and substantially blocked the release of histamine and peptidoleukotrienes from chopped lung preparations induced by OVA. Based upon their substantial inhibitory effects on the Schultz-Dale reaction, further examination on the potential anti-asthmatic effects of PTK cascade inhibitors in an in vivo model of allergic asthma is recommended. PMID- 10697264 TI - Ethnic differences in genetic susceptibility to atopy and asthma. AB - The genetics of asthma and atopy is complex, but can be approached by studies of both candidate genes and mapping of susceptibility loci. Genetic factors conferring susceptibility to disease may vary among ethnic groups. We present our experience with some candidate gene studies for asthma and atopy and susceptibility locus mapping for linkage to chromosome 5q. PMID- 10697265 TI - Value of the mesothelium-associated antibodies thrombomodulin, cytokeratin 5/6, calretinin, and CD44H in distinguishing epithelioid pleural mesothelioma from adenocarcinoma metastatic to the pleura. AB - Until recently, the standard approach of most laboratories in distinguishing epithelioid pleural mesothelioma from metastatic adenocarcinoma has been a negative result from a panel of adenocarcinoma-associated antibodies. However, several "mesothelium-associated" antibodies have been proposed as useful in this situation, and we have applied four of these putative mesothelioma markers- thrombomodulin, cytokeratin 5/6, calretinin, and CD44H--to a series of 61 epithelioid pleural mesotheliomas and 63 metastatic adenocarcinomas with known primary sites (lung = 19; breast = 21; ovary = 6; colon = 10; kidney = 4; uterus, epididymis, pancreas = 1 case each). Of the mesotheliomas, 55 of 61 (90%) stained for thrombomodulin, 56 of 61 (92%) for cytokeratin 5/6, 47 of 51 cases (92%) were positive for calretinin, and 39 of 43 (91%) were positive for CD44H. Of the metastatic adenocarcinomas, 12 of 63 (19%) cases were positive for thrombomodulin, 9 of 63 (14%) were positive for CK5/6, and 27 of 60 (45%) were positive for CD44H. With calretinin, only 1 case of 59 (2%) showed positive nuclear staining. All four antibodies stained reactive mesothelium; thrombomodulin also stained endothelium; and CD44H variably stained lymphocytes, macrophages, and fibroblasts. We conclude that all four antibodies show high sensitivity for epithelioid mesothelioma, but only calretinin (98%), cytokeratin 5/6 (86%), and thrombomodulin (81%) show sufficient specificity for practical use in this situation. PMID- 10697266 TI - Extranodal extension in lymph node-positive prostate cancer. AB - Evaluation of extranodal tumor extension may provide prognostic information for patients with epithelial malignancies. However, its importance for the patient who has prostate cancer with regional lymph node metastasis requires further investigation and clarification. This study was performed to evaluate the prognostic significance of extranodal extension (ENE) in a large series of node positive patients. The study group included 212 node-positive patients who were treated by bilateral pelvic lymphadenectomy, radical retropubic prostatectomy, and androgen deprivation between 1987 and 1992 at the Mayo Clinic. ENE was defined as cancer perforating through the lymph node capsule into perinodal tissue. Nodal cancer volume was measured by the grid method. Univariate and multivariate risk ratios (RR) for distant metastasis-free and cancer-specific survival were estimated using the Cox proportional model. The mean follow-up was 6.3 years (median, 6.1 years). Distant metastasis-free and cancer-specific survival at 5 years for all patients was 91% and 95%, respectively. ENE was found in 126 of 212 patients (59%). The presence of ENE was not significantly associated with distant metastasis-free (RR = 1.6; 95% confidence interval [CI], 0.7 to 3.9) or cancer-specific survival (RR = 2.2; 95% CI, 0.7 to 6.8). Among 98 patients with a single positive node, there was no significant difference in distant metastasis or cancer-specific survival according to the presence of ENE (P = .88 and P = .36, respectively). After adjusting for Gleason score, DNA ploidy, and ENE, only nodal cancer volume was significantly associated with adverse distant metastasis-free (RR = 1.9; 95% CI, 1.5 to 2.8) and cancer specific survival (RR = 1.4; 95% CI, 1.1 to 1.9). Our data indicate that the presence of ENE is not associated with unfavorable survival in patients with node positive prostate cancer treated by radical retropubic prostatectomy, bilateral pelvic lymphadenectomy, and androgen deprivation therapy. In contrast, nodal cancer volume was predictive of distant metastasis-free survival and cancer specific survival. PMID- 10697267 TI - Androgen receptors: a marker to increase sensitivity for identifying breast cancer in skin metastasis of unknown primary site. AB - Metastatic lesions to the skin may present a dilemma in the identification of the primary site. Breast carcinoma, metastatic to the skin, that is negative for estrogen receptors (ERs) and/or progesterone receptors (PRs) may be mimicked by a number of other metastatic lesions. In the present study, 16 formalin-fixed and paraffin-embedded infiltrating ductal carcinomas metastatic to the skin, which were ER-/PR-, ER-/PR+, or ER+/PR-; 5 metastatic lesions to the skin from primary lesions other than breast cancer; and 5 eccrine tumors were examined for immunoreactivity to the androgen receptor. The majority of the metastatic breast lesions (82%) exhibited immunopositivity for androgen receptor, whereas the metastatic skin lesions from primary lesions other than breast cancer and the eccrine tumors were immunonegative. Thus, androgen receptor immunohistochemistry could serve as a marker to increase sensitivity for identifying breast cancer in skin metastasis of unknown primary sites. PMID- 10697268 TI - Metastatic minimally invasive (encapsulated) follicular and Hurthle cell thyroid carcinoma: a study of 34 patients. AB - Most studies that have examined minimally invasive, encapsulated, follicular carcinoma (FC) or Hurthle cell carcinomas (HCs) have contained only a few metastatic neoplasms. We studied 34 patients with a single, minimally invasive, metastatic FC or HC and compared them with 38 patients with similar, nonmetastatic FCs or HCs. The numbers of incomplete capsular penetration (neoplasm into but not through the capsule), complete capsular penetration (neoplasm through the capsule), and vascular invasion foci were quantified. The median number (three), range, and distribution of complete capsular penetration and vascular invasion foci were similar in the nonmetastatic and metastatic carcinomas. All of the metastatic FCs and HCs had at least one vascular invasion or complete capsular penetration focus. Sixty-two percent of the metastatic carcinomas had two to four complete capsular penetration foci, and 60% had two to four vascular invasion foci. Two metastatic neoplasms had incomplete capsular penetration but had one and two vascular invasion foci, respectively. One tumor had no vascular invasion but had four complete capsular penetration foci. No metastatic neoplasms had incomplete capsular penetration only. There were no differences in the number of vascular invasion or complete capsular penetration foci between metastatic and nonmetastatic FCs and HCs and between metastatic FCs and HCs. Most metastatic neoplasms had vascular space invasion and complete capsular penetration. The number of complete capsular penetration or vascular invasion foci was not associated with the initial site of metastasis or the interval between the surgery and the metastasis. PMID- 10697269 TI - Discordance of p53 mutations of synchronous colorectal carcinomas. AB - It is unclear whether synchronous multiple tumors arise from multicentric or monoclonal origins. To verify the multicentric origin of synchronous colorectal carcinomas at a genetic level, immunohistochemical and molecular techniques were used to determine the p53 alterations in individual lesions of synchronous colorectal carcinomas. This study was based on a total of 32 colorectal tumors from 16 patients. Twenty-one of the 32 (66%) advanced tumors examined had positive staining for p53. Single-strand conformation polymorphism and polymerase chain reaction direct sequencing were carried out for exons 5 to 8 of p53. All cases had p53 mutations in one or more tumors of synchronous lesions. In nine patients in this series, individual lesions were found to carry a different mutated codon of the p53 gene. In the other seven patients, a p53 mutation was found in one tumor but not in another. These results indicate discordance of the mutation pattern of p53 in individual lesions of multiple colorectal carcinomas and support the idea that most synchronous colorectal carcinomas are genetically distinguishable and are multicentric in origin. We also confirmed the high frequency of p53 mutations in left-sided (71%) and rectal (91%) carcinomas, rather than right-sided (43%; P = .04) carcinomas, suggesting that the molecular mechanism of synchronous colorectal carcinomas might differ between right- and left-sided tumors in the same patient. PMID- 10697270 TI - Immunohistochemical expression of chromogranins A and B, prohormone convertases 2 and 3, and amidating enzyme in carcinoid tumors and pancreatic endocrine tumors. AB - Although chromogranin A (CgA) is widely distributed in neuroendocrine tumors, the distribution of chromogranin B (CgB) has not been elucidated. Hormones produced by tumors are sometimes prohormones and not necessarily bioactive hormones. Prohormones have to be processed into bioactive peptides by prohormone convertases (PCs), and some of them have to be amidated by peptidylglycine a amidating monooxygenase (PGM). Whether PCs and PGM are present or not in tumors may explain why some tumors are functioning and some are nonfunctioning. We investigated 45 carcinoids and 16 pancreatic endocrine tumors. Of the carcinoids, CgA was expressed in most of the tumors, except for the rectal and ovarian carcinoids, which expressed CgB strongly. The expressions of PC2, PC3, and PGM were 31%, 100%, and 87%, respectively. In the pancreatic tumors, CgA was expressed in all tumors, whereas CgB was not expressed in any tumor. The expressions of PC2, PC3, and PGM were 63%, 88%, and 63%, respectively. PC3 was expressed in all of the functioning tumors but not in two of the four nonfunctioning tumors. PC2 and PGM were not expressed in three of the four nonfunctioning tumors. In conclusion, expression of CgA and CgB was different depending on the tumor location. High frequency of PCs and PGM may explain why even nonfunctioning tumors produce some inconspicuous peptides. PMID- 10697271 TI - Influence of histochemical and immunohistochemical stains on polymerase chain reaction. AB - The polymerase chain reaction (PCR) analysis of DNA extracted from tissue sections can be applied to a variety of research and diagnostic protocols. To analyze selectively the specific areas of tissue, a direct microdissection of histochemically or immunohistochemically stained sections, if satisfactory for PCR, is helpful. However, the influence of various staining methods on PCR has been poorly investigated. In this study, paraffin sections of formalin-fixed lymph node samples were histochemically stained with Mayer's hematoxylin, eosin Y, methyl green, or May-Grunwald solution and immunostained for CD45 using 3,3' diaminobenzidine (DAB), DAB with cobalt ion (DAB-Co), or new fuchsin as the chromogen. In addition, unstained sections were treated with trypsin, microwave, or pressure cooker, the techniques frequently used in immunostains for antigen unmasking. DNA was extracted from each section, and the PCR efficiency in amplifying a 110 bp portion of the beta-globin gene was evaluated by two parameters: the cycle count in which the first visible band was obtained (CYCLE(min)) and the maximum amount of PCR products (CONC(max)). The hematoxylin stain showed a significantly prolonged CYCLE(min) (P < .01) and lower CONC(max) (P < .05) in comparison with unstained and untreated control sections. The May Grunwald stain showed a prolonged CYCLE(min) (P < .01), although the CONC(max) was not significantly different from that of the control (P = .051). The eosin and methyl green stains showed no effects against PCR. In immunostains, the DAB Co method showed a lower CONC(max) (P < .05), whereas the CYCLE(min) was not prolonged. The DAB and new fuchsin methods had no untoward effects. Antigen unmasking treatments showed deteriorating effects on PCR. The trypsin treatment significantly prolonged the CYCLE(min) (P < .01), and the PCR amplification did not reach the "plateau" level with a maximum of 60 cycles. The PCR efficiency was worse in microwave or pressure cooker treatment, with neither CYCLE(min) nor CONC(max) being obtained. When target areas from sections for subsequent PCR amplification are microdissected, methyl green is most suitable as a dye for nuclear staining. The immunohistochemical visualization with DAB or new fuchsin yields no unfavorable effects. A successful PCR amplification may not be expected in sections that are pretreated in a microwave oven or pressure cooker. PMID- 10697272 TI - Sequential observation of liver cell regeneration after massive hepatic necrosis in auxiliary partial orthotopic liver transplantation. AB - The morphogenesis of hepatocytes after massive hepatic necrosis to recovery through liver cell regeneration has not been fully understood. Sequential biopsies were performed on the native liver of a 22-year-old man who underwent auxiliary partial orthotopic liver transplantation 1 month after fulminant hepatitis. Auxiliary partial orthotopic liver transplantation was successful, and the biopsy samples permitted us to examine the regenerating process of hepatocytes after massive necrosis. At the time of auxiliary partial orthotopic liver transplantation (postoperative day 0), 95% of hepatocytes were lost and a few ductules were found in the portal areas. The ductules stained with cytokeratin 19. At postoperative day 7, the ductules began to increase in size and number and became dilated over a period of 1 month, when individual hepatocytes with clear cytoplasm appeared from the ductules. As the differentiation of hepatocytes increased, the expression of cytokeratin 19 was found to decrease. From 2 to 3 months, all of the ductules were transformed into hepatocytes, and they began to form round cell clusters. From 3 to 6 months, the round cell clusters became organized into trabecula with fibrosis. From 6 to 12 months, a lobular architecture was established, and by 14 months, the necrotic liver was fully recovered to normal. This study by examination of sequential biopsies demonstrates the progression of the regenerating process from total hepatic necrosis to complete recovery. PMID- 10697273 TI - Overexpression of BCL-2, BCL-X, and BAX in primary central nervous system lymphomas that occur in immunosuppressed patients. AB - In contrast to primary central nervous system lymphomas (PCNSLs) that occur in immunocompetent patients, most of those that occur in immunosuppressed patients are associated with Epstein-Barr virus (EBV). BCL-2-related proteins either block or promote cell death, forming homo- or heterodimers with each other. LMP-1, EBV latent protein, has been shown to upregulate BCL-2 and BCL-XL. This observation suggests that these proteins may be involved in the transformation process of EBV infected cells. Twenty-three cases of PCNSLs were studied: 12 of the patients were immunosuppressed, and 11 were immunocompetent. For all cases, we collected clinical information, histologic data, and immunophenotype and tested for the presence of EBV (EBER-1, LMP-1). Apoptosis was assessed by the TdT-mediated dUTP biotin nick-end labeling method and quantified by image analysis. In three cases, electron microscopy was performed. The BCL-2 family proteins (BCL-2, BCL-X, MCL1, and BAX) and p53 expression were studied by immunohistochemistry on paraffin slides. All cases were classified as diffuse large B-cell lymphomas. PCNSLs in immunosuppressed patients were characterized by EBV association, necrosis, important gliosis, and numerous macrophages. There was no significant difference between the two groups regarding the TdT-mediated dUTP-biotin nick-end labeling staining (P = .08). In contrast, PCNSLs in immunosuppressed patients were shown to express high levels of BCL-2, BCL-X, and BAX in more than 80% of tumor cells in 7, 10, and 11 cases, respectively. In immunocompetent patients, only one case showed a high level of BCL-2 expression in more than 80% of the cells, whereas BCL-X and BAX were overexpressed in two cases. These differences are significant (P < .05). In contrast, there was no significant difference between the two groups in MCL-1 expression. Besides EBV association and necrosis, PCNSLs related to immunosuppression are characterized by an overexpression of BCL-2-related proteins, without dramatically modifying their susceptibility for apoptosis. PMID- 10697274 TI - Hepatitis B virus-related nephropathy and lupus nephritis: morphologic similarities of two clinical entities. AB - We compared the clinicopathologic features of 22 patients with hepatitis B virus related membranous nephropathy, all with detectable glomerular hepatitis B e antigen, and of 26 patients with lupus nephritis class V. Both groups of patients similarly presented with heavy proteinuria or nephrotic syndrome; however, the patients with hepatitis B virus-related membranous nephropathy, who were predominantly male, did not have the extrarenal manifestations and autoantibodies seen in systemic lupus erythematosus. The glomerular lesions in both clinical entities were similar and at times indistinguishable, demonstrating polyclonal immunoglobulins and polytypic complements in similar subepithelial ultrastructural distribution. No morphologic feature, single or combined, carrying a high positive predictive value for the diagnosis of either nephritis was identified. Lesions such as hematoxyphil bodies and fingerprint dense deposits, distinctive of systemic lupus erythematosus, were rarely found. At the time of biopsy, when systemic lupus erythematosus is not clinically suspected, the diagnosis between hepatitis B virus-related membranous nephropathy and lupus nephritis may be difficult or impossible to differentiate, especially in geographic areas where both lupus nephritis and hepatitis B surface antigen carriers are common. This study focused on the use of specific monoclonal antisera to detect glomerular hepatitis B virus antigens, which contribute to the diagnosis of hepatitis B virus-related nephritis. PMID- 10697275 TI - CMV and transplant-related coronary atherosclerosis: an immunohistochemical, in situ hybridization, and polymerase chain reaction in situ study. AB - Accelerated graft coronary atherosclerosis is the main obstacle to long-term survival in patients who have had a heart transplant. A possible involvement of the human cytomegalovirus (HCMV) in this type of coronary atherosclerosis has been postulated by many authors but has not been definitively demonstrated. In an attempt to clarify the role of HCMV infection in the pathogenesis of this complication, we looked for in situ antigens or DNA of HCMV in 30 coronary artery segments obtained at necropsy from patients who had undergone orthotopic cardiac transplantation at the Sao Paulo Heart Institute. We tried to correlate these HCMV markers with the presence of inflammation and/or atherosclerosis in histologic sections. The patients were grouped as follows: GI, less than 170 days of graft survival and absent/mild atherosclerosis; GII, more than 170 days of graft survival and absent/mild atherosclerosis; GIII, more than 170 days of graft survival and severe/moderate atherosclerosis (170 days was the shortest graft survival time associated with atherosclerosis). The search for HCMV genome and antigens in the coronary artery sections was performed using immunohistochemistry, in situ hybridization, and polymerase chain reaction in situ techniques. Immunohistochemistry and in situ hybridization revealed no evidence of HCMV in all 30 cases. Polymerase chain reaction in situ revealed scarce HCMV-positive lymphocytes in two cases (one each from GI and GIII) located in the adventitial layer. These findings preclude a direct role for the HCMV in the pathogenesis of accelerated graft coronary atherosclerosis. However, the possibility of an indirect effect of the virus, such as an immune-mediated inflammatory response by the host that increases the expression of histocompatibility antigens, leading to tissue injury, cannot be excluded. PMID- 10697276 TI - Vascular endothelial growth factor receptor-3 (VEGFR-3): a marker of vascular tumors with presumed lymphatic differentiation, including Kaposi's sarcoma, kaposiform and Dabska-type hemangioendotheliomas, and a subset of angiosarcomas. AB - Recently, a novel monoclonal antibody to vascular endothelial growth factor receptor 3 (VEGFR-3), a tyrosine kinase receptor expressed almost exclusively by lymphatic endothelium in the adult, has been shown to react with a small number of cases of Kaposi's sarcoma (KS) and cutaneous lymphangiomas. We sought to extend these studies to a large number of well-characterized vascular neoplasms to evaluate diagnostic uses of this antibody and to determine whether it defines them in a thematic fashion. Formalin-fixed, paraffin-embedded sections from 70 vascular tumors were immunostained with antibodies to VEGFR-3 von Willebrand factor (vWF), and CD31. Anti-VEGFR-3 was positive in 23 of 24 KS, 8 of 16 angiosarcomas (AS), 6 of 6 kaposiform hemangioendotheliomas, 4 of 4 Dabska tumors, and 2 of 13 hemangiomas. Positively staining angiosarcomas were characterized either by a prominent lymphocytic component, a hobnail endothelial cell similar to that encountered in the Dabska tumor, or spindled areas resembling KS. No VEGFR-3 expression was noted in any cases of epithelioid hemangioendothelioma, pyogenic granuloma, littoral angioma, or stasis dermatitis. vWF expression was seen in 10 of 13 KS; 13 of 14 AS; 4 of 5 kaposiform hemangioendotheliomas; and all Dabska tumors, hemangiomas, lymphangiomas, epithelioid hemangioendotheliomas, vascular malformations, stasis dermatitis, and splenic littoral angiomas. CD31 expression was present in 12 of 13 KS, 13 of 14 AS, and in all other cases. Expression of VEGFR-3 is a very sensitive marker of KS, kaposiform, and Dabska-type hemangioendotheliomas, suggesting that all show at least partial lymphatic endothelial differentiation. Expression of VEGFR-3 does not reliably discriminate KS from AS. However, the expression of VEGFR-3 by certain AS having Kaposi-like areas, a prominent lymphocytic infiltrate, or hobnail endothelium may define subset(s) having phenotypic, if not pathogenetic and biologic, differences. PMID- 10697277 TI - Pathologic features, proliferative activity, and cyclin D1 expression in Hurthle cell neoplasms of the thyroid. AB - Making a histologic distinction between Hurthle cell adenomas and carcinomas sometimes may be difficult. We analyzed a series of Hurthle cell lesions to determine whether specific histologic features and expression of Ki67 and cyclin D1 could be useful in distinguishing Hurthle cell adenomas from carcinomas. Formalin-fixed, paraffin-embedded tissues from 128 Hurthle cell neoplasms, including 59 adenomas; 55 carcinomas; and 14 tumors classified as neoplasms of uncertain malignant behavior (UMB), which had equivocal capsular invasion but no vascular invasion, were analyzed for expression of Ki67 and cyclin D1 by immunostaining. The distribution of immunoreactivity for Ki67 with antibody MIB-1 was analyzed by quantifying the percentage of positive nuclei that was expressed as the labeling index. None of the patients with adenomas or UMB tumors developed recurrent or metastatic disease after a mean follow-up of 7.8 and 7.9 years, respectively. Of the 55 patients with Hurthle cell carcinoma, 19 were associated with metastatic disease, 13 of whom died with disease. No patient with a Hurthle cell carcinoma without vascular invasion developed metastatic disease. The mean tumor size for Hurthle cell carcinomas (4.8 cm) was significantly larger than that of Hurthle cell adenomas (3.1 cm) or UMB tumors (3.7 cm). No patient with a Hurthle cell tumor smaller than 3.5 cm developed metastatic disease, even when vascular invasion was present. The Ki67 labeling index in Hurthle cell carcinomas (10.0 +/- 1.2) was 3-fold higher than in Hurthle cell adenomas (3.2 +/- 0.3). The Ki67 labeling index in the UMB group was 5.0 +/- 0.7. Cyclin D1 showed diffuse nuclear staining in 1 of the 59 (1.7%) Hurthle cell adenomas, in 10 of the 55 (18%) Hurthle cell carcinomas, and in none of the UMB tumors. In summary, analyses of the cell cycle proteins Ki67 and cyclin D1 in Hurthle cell thyroid neoplasms indicate that these markers may assist in distinguishing some Hurthle cell carcinomas from adenomas. Among the Hurthle cell carcinomas, large tumor size and vascular invasion are associated with clinically aggressive tumors. Our study also suggests that Hurthle cell neoplasms with only equivocal capsular invasion and no vascular invasion should behave in a benign manner. PMID- 10697278 TI - The World Health Organization classification of hematological malignancies report of the Clinical Advisory Committee Meeting, Airlie House, Virginia, November 1997. AB - Since 1995, the European Association of Pathologists and the Society for Hematopathology have been developing a new World Health Organization (WHO) classification of hematologic malignancies. The classification includes lymphoid, myeloid, histiocytic, and mast cell neoplasms. The WHO project involves 10 committees of pathologists, who have developed lists and definitions of disease entities. A Clinical Advisory Committee of international hematologists and oncologists was formed to ensure that the classification will be useful to clinicians. A meeting was held in November 1997 to discuss clinical issues related to the classification. The WHO has adopted the Revised European-American Classification of Lymphoid Neoplasms, published in 1994 by the International Lymphoma Study Group, as the classification of lymphoid neoplasms. This approach to classification is based on the principle that a classification is a list of "real" disease entities, which are defined by a combination of morphology, immunophenotype, genetic features, and clinical features. The relative importance of each of these features varies among diseases, and there is no one "gold standard." The WHO classification has applied the principles of the Revised European-American Classification of Lymphoid Neoplasms to myeloid and histiocytic neoplasms. The classification of myeloid neoplasms recognizes distinct entities defined by a combination of morphology and cytogenetic abnormalities. The Clinical Advisory Committee meeting, which was organized around a series of clinical questions, was able to reach a consensus on most of the questions posed. The questions and the consensus are discussed in detail in this article. Among other things, the Clinical Advisory Committee concluded that clinical grouping of lymphoid neoplasms was neither necessary nor desirable. Patient treatment is determined by the specific type of lymphoma, with the addition of grade within the tumor type, if applicable, and clinical prognostic factors such as the international prognostic index. The experience of developing the WHO classification has produced a new and exciting degree of cooperation and communication between oncologists and pathologists from around the world. This should facilitate progress in the understanding and treatment of hematologic malignancies. PMID- 10697279 TI - Correspondence re: Sherman ME, Tabbara SO, Scott DR, Kurman RJ, Glass AG, Manos MM, et al. "Ascus, rule out HSIL": cytologic features, histologic correlates, and human papillomavirus detection. Mod Pathol 1999;12:335-42. PMID- 10697280 TI - Correspondence re: Basolo F, Baloch ZW, Baldanzi A, Miccolo P, Livolsi VA: Usefulness of ultrafast Papanicolaou-stained scrape preparations in intraoperative management of thyroid lesions. Mod Pathol 1999;12:653-7. PMID- 10697281 TI - The epidemiology of contact transmission: beyond Semmelweis. PMID- 10697282 TI - A prolonged outbreak of Pseudomonas aeruginosa in a neonatal intensive care unit: did staff fingernails play a role in disease transmission? AB - OBJECTIVES: To describe an outbreak of Pseudomonas aeruginosa bloodstream infection (BSI) and endotracheal tube (ETT) colonization in a neonatal intensive care unit (NICU), determine risk factors for infection, and make preventive recommendations. DESIGN: A 15-month cohort study followed by a case-control study with an environmental survey and molecular typing of available isolates using pulsed-field gel electrophoresis. SETTING AND PATIENTS: Neonates in the NICU of a university-affiliated children's hospital. INTERVENTIONS: Improved hand washing and restriction of use of long or artificial fingernails. RESULTS: Of 439 neonates admitted during the study period, 46 (10.5%) acquired P aeruginosa; 16 (35%) of those died. Fifteen (75%) of 20 patients for whom isolates were genotyped had genotype A, and 3 (15%) had genotype B. Of 104 healthcare workers (HCWs) from whom hand cultures were obtained, P aeruginosa was isolated from three nurses. Cultures from nurses A-1 and A-2 grew genotype A, and cultures from nurse B grew genotype B. Nurse A-1 had long natural fingernails, nurse B had long artificial fingernails, and nurse A-2 had short natural fingernails. On multivariate logistic regression analysis, exposure to nurse A-1 and exposure to nurse B were each independently associated with acquiring a BSI or ETT colonization with P aeruginosa, but other variables, including exposure to nurse A-2, were not. CONCLUSION: Epidemiological evidence demonstrated an association between acquiring P aeruginosa and exposure to two nurses. Genetic and environmental evidence supported that association and suggested, but did not prove, a possible role for long or artificial fingernails in the colonization of HCWs' hands with P aeruginosa. Requiring short natural fingernails in NICUs is a reasonable policy that might reduce the incidence of hospital-acquired infections. PMID- 10697283 TI - Morphological heterogeneity of the GABAergic network in the suprachiasmatic nucleus, the brain's circadian pacemaker. AB - GABA (gamma-amino-butyric acid) is the predominant neurotransmitter in the mammalian suprachiasmatic nucleus (SCN), with a central role in circadian time keeping. We therefore undertook an ultrastructural analysis of the GABA containing innervation in the SCN of mice and rats using immunoperoxidase and immunogold procedures. GABA-immunoreactive (GABA-ir) neurons were identified by use of anti-GABA and anti-GAD (glutamic acid decarboxylase) antisera. The relationship between GABA-ir elements and the most prominent peptidergic neurons in the SCN, containing vasopressin-neurophysin (VP-NP) or vasoactive intestinal polypeptide (VIP), was also studied. Within any given field in the SCN, approximately 40-70% of the neuronal profiles were GABA-ir. In GABA-ir somata, immunogold particles were prominent over mitochondria, sparse over cytoplasm, and scattered as aggregates over nucleoplasm. In axonal boutons, gold particles were concentrated over electron-lucent synaptic vesicles (diameter 40-60 nm) and mitochondria, and in some instances over dense-cored vesicles (DCVs, diameter 90 110 nm). GABA-ir boutons formed either symmetric or asymmetric synaptic contacts with somata, dendritic shafts and spines, and occasionally with other terminals (axo-axonic). Homologous or autaptic connections (GABA on GABA, or GAD on GAD) were common. Although GABA appeared to predominate in most neuronal profiles, colocalisation of GABA within neurons that were predominantly neuropeptide containing was also evident. About 66% of the VIP-containing boutons and 32% of the vasopressinergic boutons contained GABA. The dense and complex GABAergic network that pervades the SCN is therefore comprised of multiple neuronal phenotypes containing GABA, including a wide variety of axonal boutons that impinge on heterologous and homologous postsynaptic sites. PMID- 10697284 TI - A descriptive and comparative lectin histochemical study of the vomeronasal system in pigs and sheep. AB - The accessory olfactory bulb (AOB) is the primary target of the sensory epithelium of the vomeronasal organ (VNO), and thus constitutes a fundamental component of the accessory olfactory system, which is involved in responses to behaviour-related olfactory stimuli. In this study we investigated the characteristics of the AOB, VNO, vomeronasal nerves (VNNs) and caudal nasal nerve (CdNN) in pigs and sheep, species in which olfaction plays a key behavioural role both in the neonatal period and in adulthood. The patterns of staining of the AOB by the Bandeiraea simplicifolia and Lycopersicon esculentum lectins were the same in the 2 species, whereas the Ulex europeus and Dolichos biflorus lectins gave different patterns. In both species, lectin staining of the AOB was consistent with that of the VNNs, while the CdNN did not label any of the structures studied. The entire sensory epithelium of the pig was labelled by Ulex europeus and Lycopersicum esculentum lectins, and all 4 lectins used labelled the mucomicrovillar surface of the sensory epithelium in sheep. PMID- 10697285 TI - The relationship between accessory foramina and tumour spread on the medial mandibular surface. AB - The medial cortical surface of the mandible can be involved by tumour infiltration from the floor of the mouth. A detailed study of spread via accessory foramina through the edentulous alveolar crest has been previously undertaken, but no similar study has been carried out for the medial surface. In order to gain further appreciation of the mode of tumour spread, a study of the number and distribution of accessory foramina on the medial mandibular surface was performed on 89 mandibles. The number of foramina varied greatly from specimen to specimen. In the ascending ramus above the inferior dental foramen, 3 mandibles showed no foramina; the condylar section possessed the greatest proportion followed by the sigmoid and the coronoid. On the rest of the medial surface below the inferior dental foramen, all specimens showed at least 1 accessory foramen; the greatest concentration was in the middle third along the path of the inferior dental canal, followed by the upper third and the lower third section. Accessory foramina were repeatedly present at certain dedicated sites. The medial facing wall of the inferior dental foramen was found to be the commonest dedicated site (98.3%) followed by foramina on either side of the genial tubercles (71.9%), the digastric fossa (71.9%) and the median foramen above the genial tubercles (64%). The findings of this study are in keeping with the current observation that the lower border is least commonly involved in tumour spread. In view of the presence of accessory foramina along the inferior dental canal and especially on the medial facing wall of the inferior dental foramen, it is imperative to preclude tumour spread in this region prior to undertaking the conservative rim resection procedure. Medial to the symphysis the alveolar mucosa dips down almost to the level of the dedicated foramina in the vicinity of the genial tubercles. As a general rule the attached muscle forms a barrier to tumour spread except in the later stages, however, in irradiated mandibles resistance to spread has been previously reported to be diminished. Under these circumstances, it is possible that the numerous accessory foramina reported in this study could facilitate a direct pathway into the cancellous bone. PMID- 10697286 TI - The metanephros of the quail embryo. Developmental expression of carbonic anhydrase investigated by multiple approaches. AB - The expression of carbonic anhydrase (CA) in the quail metanephros was investigated during embryonic development. The immunohistochemical localisation of the isoenzymes CAII and CAIII was compared with the distribution of enzyme activity visualised by a histochemical cobalt-precipitation procedure. The developmental profile of CA activity was also evaluated by means of a biochemical method. The occurrence of a moderate and diffuse CAII immunostaining from the first developmental appearance of the metanephros anlage testified to an early expression of carbonic anhydrase. This finding is discussed in relation to the involvement of the enzyme in the morphogenetic mechanisms leading to the establishment both of cell polarity and epithelial phenotype. CA expression in the renal sites that are positive in adults proved to be developmentally regulated. In the collecting duct system, enzyme activity could not be identified until the time of hatching. No CA was detected at any stage examined at the sites where, in adults, enzyme occurrence has previously been interpreted as a membrane associated CA isoform. The differentiating renal tubules displayed no CAIII immunoreactivity. It can be argued that the bulk of the enzyme activity in the embryonic metanephros is due to the cytosolic isoenzyme CAII. PMID- 10697287 TI - Effects of ageing on the insertion zones of the human vocal fold. AB - The vocal ligaments insert at the anterior and posterior commissures of the larynx. These structures fulfil biomechanical functions, balancing the different elastic moduli of tendon, cartilage or bone and undergo age-related changes that may be responsible for voice changes with increasing age. The aim of this study was to analyse the insertion structures of the vocal ligaments by means of macroscopic, histological, immunohistochemical and electron-microscopic methods and to draw conclusions from age-related structural changes on a functional basis. Investigations were carried out on the larynges of 22 males and 15 females (aged 1-95 y). In adolescence, the insertion zone of the vocal ligament tendon, a dense network of connective tissue rich in sulphated glycosaminoglycans at the thyroid cartilage, is characterised by a layer between tendon and cartilage comparable to fibrocartilage. The insertion zone lacks a perichondrium. Collagen fibrils of the vocal ligament tendon penetrate directly into the thyroid cartilage. In the insertion area, the chondrocytes are surrounded by collagen fibrils, which show positive reactivity to antibodies against type I and type III collagen. Sulphated glycosaminoglycans are integrated between the collagen fibrils. In the area of the posterior glottis, elastic cartilage rests like a cap on the hyaline base of the arytenoid cartilage. There is no distinctive border between the structures. With increasing age, ossification of the laryngeal skeleton occurs, involving hyaline cartilage at the posterior glottis and hyaline and fibrocartilage at the anterior commissure. At the same time, a loss or sulphated glycosaminoglycans is observed inside the vocal ligament tendon. Advanced ossification of the laryngeal skeleton, particularly in the area of the commissures, an increasing loss of glycosaminoglycans in the vocal ligament tendon and changes in the elastic tissue reduce the elastic modulus between tendon, cartilage and bone, thus 'stiffening' the insertion zones, which could be one factor among others favouring voice changes with advancing age. PMID- 10697288 TI - Endothelial glycoconjugates: a comparative lectin study of the brain, retina and myocardium. AB - There is evidence that the endothelial cell (EC) glycocalyx is a significant determinant of vascular permeability, acting as a charge-size filter to permeant molecules. We have therefore examined its oligosaccharide composition in 3 classes of microvessel with differing permeabilities. EC in rat brain, retina and myocardium were labelled with a panel of lectins and subjected to a semiquantitative analysis. Surprisingly, no substantial differences were evident for any lectin labelling between the 3 microvessel types despite their marked morphophysiological diversity. In particular, all showed substantial sialic acid expression, with Maackia amurensis (MAA) labelling sialic acid in an alpha2-3 linkage to beta-galactose and Sambucus nigra (SNA) recognising sialic acid in an alpha2-6 linkage to beta-galactose. Arachis hypogaea (PNA) binding after neuraminidase digestion indicated the presence of Gal beta1-3GalNAc attached to terminal sialic acid. The results therefore show that the sequences NeuNAc alpha2 3Gal beta1-3GalNAc and NeuNAc alpha2-6Gal beta1-3GalNAc are strongly expressed in the 3 microvessel types irrespective of their permeability properties. This homogeneity suggests that these lectin ligands may be involved in a common set of EC functions, e.g. cell:cell and cell:matrix interactions. However, we cannot rule out the possibility that glycocalyx differences may exist between vessels in the paracellular cleft which may alter its filtration properties. PMID- 10697289 TI - Human retinal astroglia. A comparative study of adult and the 18 month postnatal developmental stage. AB - The immunohistochemical location of glial fibrillary acidic protein (GFAP) was used to study the state of maturation of retinal astrocytes from an 18-mo-old infant and to compare it with the situation in the adult. Infant astrocytes showed intense GFAP immunoreactivity in the perikarya and possessed spindle-like enlargements in their processes, while in the adult immunoreactivity in the perikarya was scarce and the spindle-like enlargements were not evident. Two types of astrocyte were observed in adult and child retinas: elongated and star shaped. In the adult, the star-shaped type tend to be more stylised and to have longer processes than in the infant. In the infant, numerous astrocyte cell bodies were observed over vessels, while in the adult these were scarce. In the infant, the star-shaped astrocytes made up a honeycomb plexus, but this was not fully developed. These results suggest that at 18 mo of postnatal development the retinal astrocytes are still increasing and growing into the astroglial structure found in adults. PMID- 10697290 TI - Structure and mineralisation density of antler and pedicle bone in red deer (Cervus elaphus L.) exposed to different levels of environmental fluoride: a quantitative backscattered electron imaging study. AB - The structure and relative degree of mineralisation of antler and pedicle bone of yearling red deer stags exposed either to low or high levels of environmental fluoride were determined by digital quantitative backscattered electron (BSE) imaging. Bone fluoride content (BFC) in antlers (845 +/- 86 mg F-/kg ash, arithmetic mean +/- S.E.M.) and pedicles (1448 +/- 154 mg F-/kg ash) of deer from a highly fluoride polluted area in North Bohemia (Czech Republic) were significantly higher (P < 0.001) than those of controls from uncontaminated regions in West Germany (antlers: 206 +/- 41, pedicles: 322 +/- 52 mg F-/kg ash). Mean (56.5 +/- 4.5%) and maximum (84.9 +/- 2.1%) mineralised bone area of the control antlers significantly (P < 0.05 and P < 0.001, respectively) exceeded the corresponding values for the N. Bohemian deer (43.3 +/- 1.3 and 73.3 +/- 1.9%, respectively), while the pedicles from the 2 groups did not differ significantly. In the pooled antler samples (n = 18), negative correlations existed between BFC and mean (r(s) = -0.62, P < 0.01) as well as maximum (r(s) = -0.69, P < 0.01) mineralised bone area. Morphological imaging revealed a decreased width and an increased porosity of the antler cortex in the N. Bohemian specimens. Mean (148.5 +/- 1.7) and maximum (154.2 +/- 1.7) BSE-signal intensities (= grey levels; range between a monobrominated (grey level 0) and a monoiodinated (grey level 255) dimethacrylate resin standard) of the antlers from the controls were significantly higher than those of the N. Bohemian deer (140.7 +/- 2.1 and 145.7 +/- 2.2, respectively; P < 0.05 for both comparisons). In the pooled antler samples, negative correlations between BFC and mean (r(s) = -0.51, P < 0.05) as well as maximum (r(s) = -0.52, P < 0.05) BSE-signal intensities were observed. No significant differences in mineralisation density parameters were found for the 2 pedicle samples, and BFC and mineralisation density of the pooled pedicles were uncorrelated. Morphological imaging revealed bone mottling (denoting increased remodelling activity) and frequent occurrence of apparently increased osteocyte lacunae in some of the pedicles from the N. Bohemian deer. It is concluded that the reduced amount of mineralised bone in, and the lower mineralisation density of, the N. Bohemian antlers resulted from a fluoride induced disturbance of bone mineralisation. The rapid growth of antlers leads both to a high mineral demand and a high rate of fluoride uptake during antlerogenesis. This, and the limited lifespan of antlers, which does not allow for a compensation of a delay in the onset or progression of the mineralisation process, renders antler bone particularly susceptible to fluoride. Antlers are therefore considered a useful model for studying fluoride effects on bone formation. Furthermore, analysis of cast antlers enables a noninvasive monitoring of environmental pollution by fluorides. PMID- 10697291 TI - Quantitative analysis of the anatomy of the epineurium of the canine recurrent laryngeal nerve. AB - The purpose of this investigation was to determine the amount of epineurium surrounding the recurrent laryngeal nerve (RLN) compared with a limb nerve, that to flexor hallicus longus (NFHL). Nerve samples were obtained from 10 adult dogs and studied using scanning electron microscopy and light microscopy to measure the relative proportion of epineurium and the relative proportions of adipose and collagenous tissue comprising the epineurium in both nerves. Significantly greater relative epineurial cross-sectional areas and adipose content were found in the RLN than in the NFHL. Based on observations on noncranial peripheral nerves, the findings indicate that the RLN is better protected against deformational forces associated with compression than stretching forces. The RLN may not be structured well for successful reinnervation after injury. The patterns observed for adipose tissue in RLN epineurial tissue appeared unique compared with those previously reported in peripheral nerves. The primary role associated with adipose tissue is to 'package' the nerve for protection. The RLN is considered to be a vital nerve in the body, as are other cranial nerves. The large proportions of adipose tissue in the epineurium may relate to the importance of protecting this nerve from injury. PMID- 10697292 TI - Elimination of atretic follicles from the mouse ovary: a TEM and immunohistochemical study in mice. AB - We examined numerous ovarian follicles from 32-35 d virgin mice by transmission electron microscopy and light microscopic immunohistochemistry. No macrophages were seen, but various stages of apoptotic granulosa cells were encountered. Presumably a granulosa cell or its debris in an advanced stage of apoptosis was destined to be phagocytosed by adjacent normal-looking granulosa cells. Other granulosa cells of normal appearance were seen in the region of the zona pellucida in contact with and apparently phagocytosing atrophic oocytes. Such granulosa cells were characterised by the presence of gap junctions with other cells and frequently contained annular gap junctions in the cytoplasm. To confirm the lack of involvement of macrophages in the process of follicular atresia and elimination, specially prepared ovarian sections were incubated with antimouse macrophage monoclonal antibodies (F4/80, Mac-1, Mac-2). None of the follicles examined showed positive immunoreactivity with these antibodies. Atretic follicles may shrink and eventually disappear from the ovary as a result of repeated apoptosis and phagocytosis by granulosa cells. There is no evidence for the presence or involvement of macrophages in the atretic follicles, at least in prereproductive mice as examined. PMID- 10697293 TI - Relationship of decrease in fecundity with advancing age to structural changes in mouse endometrium. AB - The aim of this study was to determine whether a relationship exists between decrease in fecundity and structural changes in the antimesometrial endometrium of the mouse. Fecundity was calculated as the number of animals showing a placental sign/number of copulated animals x 100 (%). Structural changes in the endometrium were examined by electron microscopy. A negative correlation between age and fecundity was found. Fecundity was 50% at 7 mo of age. At this age, amorphous material appeared in the region between the basement membrane deep to the luminal epithelium and the subepithelial cells. This material was sometimes attached to the basement membrane. It increased in amount with advancing age, as fecundity decreased. The structure of the uterine luminal epithelial cells did not alter with age. The results indicated that decrease in fecundity with advancing age is correlated with the appearance of amorphous material beneath the basal lamina of the endometrial epithelium. It is suggested that this could impair communication between the luminal epithelium and the endometrial stroma, which plays an important role in implantation. PMID- 10697294 TI - Anastomosis at the level of the elbow joint connecting the deep, or normal, brachial artery with major arterial variations of the upper limb. PMID- 10697295 TI - DbetaH-immunoreactive subepithelial nerves in the vas deferens of prostate cancer patients. PMID- 10697296 TI - The transitional zone and CNS regeneration. AB - Most nerves are attached to the neuraxis by rootlets. The CNS-PNS transitional zone (TZ) is that length of rootlet containing both central and peripheral nervous tissue. The 2 tissues are separated by a very irregular but clearly defined interface, consisting of the surface of the astrocytic tissue comprising the central component of the TZ. Central to this, myelin sheaths are formed by oligodendrocytes and the supporting tissue is astrocytic. Peripheral to it, sheaths are formed by Schwann cells which are enveloped in endoneurium. The features of transitional nodes are a composite of those of central and peripheral type. The interface is penetrated only by axons. It is absent at first. It is formed by growth of processes into the axon bundle from glial cell bodies around its perimeter. These form a barrier across the bundle which fully segregates prospectively myelinated axons. Rat spinal dorsal root TZs have been used extensively to study CNS axon regeneration. The CNS part of the TZ responds to primary afferent axon degeneration and to regenerating axons in ways which constitute a satisfactory model of the gliotic tissue response which occurs in CNS lesions. It undergoes gliosis and the gliotic TZ tissue expands distally along the root. In mature animals axons can regenerate satisfactorily through the endoneurial tubes of the root but cease growth on reaching the gliotic tissue. The general objective of experimental studies is to achieve axon regeneration from the PNS through this outgrowth and into the dorsal spinal cord. Since immature tissue has a greater capacity for regeneration than that of the adult, one approach includes the transplantation of embryonic or fetal dorsal root ganglia into the locus of an extirpated adult ganglion. Axons grow centrally from the transplanted ganglion cells and some enter the cord. Other approaches include alteration of the TZ environment to facilitate axon regeneration, for example, by the application of tropic, trophic, or other molecular factors, and also by transplantation of cultured olfactory ensheathing cells (OECs) into the TZ region. OECs, by association with growing axons, facilitate their extensive regeneration into the cord. Unusually, ventral motoneuron axons may undergo some degree of unaided CNS regeneration. When interrupted in the spinal cord white matter, some grow out to the ventral rootlet TZ and thence distally in the PNS. The DRTZ is especially useful for quantitative studies on regeneration. Since the tissue is anisometric, individual parameters such as axon numbers, axon size and glial ensheathment can be readily measured and compared in the CNS and PNS environments, thereby yielding indices of regeneration across the interface for different sets of experimental conditions. PMID- 10697297 TI - Hypertension and dementia: can antihypertensive treatment preserve cognitive function? PMID- 10697298 TI - The costs of not treating hypertension. PMID- 10697299 TI - Mortality, risk indicators, mode and place of death and symptoms of angina pectoris in the five years after coronary artery bypass grafting in patients with and without a history of hypertension. AB - AIM: To describe mortality, risk indicators for death, place and mode of death, and symptoms of angina pectoris among survivors in the 5 years after coronary artery bypass grafting (CABG) in patients with and without a history of hypertension. METHODS: All patients in western Sweden who underwent CABG without concomitant valve surgery and without previously performed CABG between June 1988 and June 1991. RESULTS: A total of 1997 patients were included in the analysis, 740 (37%) of whom had a history of hypertension. Patients with no history had a 5 year mortality of 12.4%. The corresponding relative risk for hypertensives was 1.4 (95% CI 1.1-1.8). Risk factors for death appeared similar in patients with and without a history of hypertension. Patients with hypertension had an increased risk of death in hospital and an increased risk of a non-cardiac death. Among survivors after 5 years, patients with a history of hypertension tended to have a higher prevalence of symptoms equivalent to angina pectoris. CONCLUSIONS: Patients with a history of hypertension have an increased risk of death in the 5 years after CABG. Risk factors for death appear similar in patients with and without a history of hypertension. Patients with hypertension have a particularly increased risk of death in hospital and of death judged as non-cardiac. PMID- 10697300 TI - Influence of non-steady state during isoglycemic hyperinsulinemic clamp in hypertension. A LIFE substudy. AB - We wanted to investigate whether time to steady state was reached within 2 h of insulin infusion during isoglycemic hyperinsulinemic clamp, comparing the glucose uptake index (M/IG) with Bergman's insulin sensitivity index (Sip). We performed a 2-h oral glucose tolerance test and a 3-h isoglycemic hyperinsulinemic clamp in 26 young, healthy subjects and 43 elderly patients with unmedicated essential hypertension and left ventricular hypertrophy. The 3-h Sip correlated strongly with the 2-h M/IG in the patients (r = 0.88, p < 0.001) as well as in the healthy subjects (r = 0.96, p < 0.001) with relatively narrow limits of agreement in the patients. However, during the third hour of insulin infusion, M/IG (10.0 vs 12.21(2) x kg(-1) x min(-1) x mmol(-1), p < 0.001) as well as Sip (7.1 vs 9.41(2) x kg(-1) x min(-1) x mmol(-1), p < 0.001) increased significantly in the patients, but not in the healthy subjects. Because the 2-h M/IG correlated strongly with the 3-h Sip with relatively narrow limits of agreement, it is a good measure of insulin sensitivity. However, a 2-h clamp results in lower insulin sensitivity values in elderly, hypertensive patients due to the fact that steady state is not reached, demonstrating a higher prevalence of insulin resistance in such a population. PMID- 10697301 TI - Effects of selective angiotensin II type 1 receptor blockade with losartan on arterial compliance in patients with mild essential hypertension. AB - It has been suggested that antihypertensive drugs should not only decrease blood pressure, but also improve large artery compliance. The aim of the present study was to examine the effect of losartan, a selective angiotensin II type 1 receptor antagonist, on parameters reflecting arterial compliance. In a randomized, double blind cross-over study, 16 patients with mild essential hypertension were examined after 4 weeks of treatment with placebo/losartan. The effect on finger plethysmographic arterial pulse curves were quantified by computing the relative height of the dicrotic notch, and pulse wave velocity was estimated by measurements of the time delay from the start of the QRS-complex (electrocardiogram) to the foot of the plethysmographic pulse wave. Compared with placebo, losartan reduced the relative height of the dicrotic notch from 55% (SD 12) to 47% [14] (p < 0.01), and pulse wave velocity from 9.3 m/sec to 8.7 m/sec (p < 0.05). The supine blood pressure decreased from 146/89 mmHg to 134/82 mmHg (p < 0.01). There was no correlation between the effects on blood pressure and the effects on the arterial compliance parameters, suggesting that losartan exerted an effect on arterial compliance beyond its effect on blood pressure. PMID- 10697302 TI - Age-related differences in blood pressure and heart rate responses to changes in body position: results from a study with serial measurements in the supine and standing positions in 30-, 50- and 60-year-old men. AB - This population-based study presents the blood pressure and heart rate responses to sudden changes in body position in representative groups of men aged 30 (n = 50), 50 (n = 44) and 60 (n = 69) years, using an unbiased method for non-invasive blood pressure measurements. Blood pressure and heart rate were measured every minute during three 7-min periods in the supine, standing and again supine positions. Whereas there was an initial decrease in systolic blood pressure upon standing in men aged 50 and 60 years, an increase was seen in the 30-year-olds. The diastolic blood pressure increased in all age groups, but less in the older compared to the younger men. In all age groups, the changes in systolic blood pressure upon standing were transient, while the changes in the diastolic blood pressure lasted during the entire observation period. The heart rate increased to a similar extent upon standing in all age groups. No symptomatic hypotension was observed. After resuming the supine position, both blood pressure and heart rate returned towards the levels initially recorded. This population-based study confirms previous observations in selected subjects of age-related attenuation in blood pressure response to change in body position. The study also shows that blood pressure and heart rate are rapidly stabilized upon standing up. PMID- 10697303 TI - Increased forearm blood flow during glucose clamp is related neither to insulin sensitivity nor to hyperinsulinemia in borderline hypertensive young men. AB - It is controversial whether raised insulin within the physiological concentration range increases forearm blood flow (FBF). The aim of the present study was therefore to examine the effect of the isoglycemic hyperinsulinemic glucose clamp procedure on FBF and to relate the increase to the glucose disposal rate (GDR), i.e. insulin sensitivity. Borderline hypertensive young men were examined with the clamp technique or received saline infusion, and FBF was measured using plethysmography. It is of particular interest to study this group of subjects because their GDR correlates to a number of metabolic and hemodynamic variables, and these subjects hyperreact to stressful stimuli. There was no correlation between deltaFBF during clamp and GDR (r = -0.002, p = 0.99, n = 28). While serum insulin increased from 107 +/- 5 to 628 +/- 31 pmol/l in the hyperinsulinemic group and remained unchanged (135 +/- 11 vs 116 +/- 11 pmol/l) in the saline group, FBF increased from 3.5 +/- 0.3 to a maximum of 5.1 +/- 0.4 ml/min/100 ml (p < 0.001, n = 28) and from 2.8 +/- 0.5 to a maximum of 4.5 +/- 0.5 ml/min/100 ml (p = 0.01, n = 8), respectively. The increase in FBF (delta%) was similar in the two groups (p = 0.9). Thus, we could not demonstrate any relationship between insulin sensitivity and increments in FBF during hyperinsulinemic glucose clamp in borderline hypertensive young men. The moderate increases in FBF during insulin infusion with serum concentrations within the physiological range seem to be time-dependent and not caused by hyperinsulinemia. PMID- 10697304 TI - Similar central hemodynamics in salt-sensitive and salt-resistant hypertensive patients. AB - Salt may be involved in the pathogenesis of essential hypertension but no agreement has been reached on how salt might exert its blood pressure control. One reason for the conflicting results could be differences in response to changes in salt intake--i.e. between salt-sensitive and salt-resistant subjects. Hypertension reflects a hemodynamic disturbance: mainly an increase in total peripheral resistance. In order to determine whether central hemodynamics is different in salt-sensitive and salt-resistant essential hypertension, a study was carried out on 37 patients aged 31-63 years with mean casual blood pressure 165/104 mmHg. Based on an increase in ambulatory 24-h mean blood pressure of > or = 10% after one week of dietary salt loading (260 mmol NaCl/24 h) following a one week salt depletion period (60 mmol NaCl/24 h), 7 patients (19%) were classified as salt sensitive and 30 patients (81%) as salt resistant. Before the salt sensitivity test, while patients were on their habitual salt intake (160 mmol NaCl/24 h), central hemodynamics (intra-arterial pressure, cardiac output by dye dilution, heart rate by electrocardiogram, and total peripheral resistance) was examined at rest and during bicycle exercise. None of the central hemodynamic variables were different between the two groups, despite a marked difference in blood pressure response to one week of salt loading between the salt-sensitive and the salt-resistant groups (27/9 mmHg vs -2/1 mmHg). Furthermore, no statistically significant differences were observed in neurohumoral variables or echocardiographic indices of left ventricular dimensions between the two groups. Owing to the invasive hemodynamic procedure, central hemodynamics was not restudied during high- or low salt intake. It is concluded that there is no difference in central hemodynamics in salt-sensitive and salt-resistant hypertensive patients when they are on their habitual salt diet. PMID- 10697306 TI - Health economics and orthopaedics. AB - It is becoming widely accepted that research which considers only the outcome and not the costs associated with new technologies in health care, is of limited value in making decisions about the use of scarce resources. Economic evaluation is becoming a standard feature of clinical research but many published economic evaluations fall short of best practice in their methodology. We have described the essential features of economic evaluation, using published studies in orthopaedics, in order to try to improve the ability of orthopaedic surgeons to read, understand and appraise such studies critically, and to encourage them to consider including economic evaluation in future investigations. PMID- 10697305 TI - Candesartan cilexetil in hypertension: effects of six weeks' treatment on haemodynamics, baroreceptor sensitivity and the renin-angiotensin-aldosterone system. AB - The effects of the angiotensin II receptor blocker candesartan cilexetil on systemic and forearm haemodynamics and baroreceptor sensitivity were evaluated in this randomized, placebo-controlled, double-blind, crossover study. After a 4 week placebo run-in period, 22 patients with essential hypertension (diastolic blood pressure 100-114 mmHg) were randomized to receive either candesartan cilexetil 16 mg or placebo once daily for 6 weeks. At the end of each period, 24 h after the last dose, invasive haemodynamic assessments were performed. Simultaneously, the plasma renin activity and plasma concentrations of angiotensin II, aldosterone and catecholamines were measured. Compared to placebo, candesartan cilexetil significantly reduced mean arterial pressure by 8 mmHg (95% CI: 2.6; 12.3), while cardiac output, stroke volume and heart rate were unchanged. Forearm vascular resistance was reduced by 1 mmHg x ml(-1) x L x min (CI: 0.3; 2.3). The baroreceptor sensitivity was not influenced, but a change in the set-point was noted. Plasma renin activity and angiotensin II concentrations were increased, while the aldosterone concentration was significantly reduced. Plasma catecholamine concentrations were unaffected. In conclusion, 6 weeks' treatment with candesartan cilexetil 16 mg o.d. induced systemic and forearm vasodilatation and a reduction in blood pressure without compromising cardiac performance. The plasma concentration of aldosterone was reduced. PMID- 10697307 TI - Accuracy of references in the orthopaedic literature. PMID- 10697308 TI - The Sofield-Millar operation in osteogenesis imperfecta. A modified technique. AB - We have reviewed the results of the Sofield-Millar operation on 58 long bones in ten patients. If more than three osteotomies were undertaken the time to union of the bone was significantly prolonged (p < 0.001) with significant thinning of the bone (p < 0.02). We have used a modified technique in order to minimise surgical trauma and devascularisation of the bone. The rod is introduced under the control of an image-intensifier. Small surgical exposures are made only at the sites of corrective wedge osteotomies. The number of osteotomies is kept to the minimum. PMID- 10697309 TI - Test of stability as an aid to decide the need for osteotomy in association with open reduction in developmental dysplasia of the hip. AB - After open reduction for developmental dysplasia of the hip (DDH), a pelvic or femoral osteotomy may be required to maintain a stable concentric reduction. We report the clinical and radiological outcome in 82 children (95 hips) with DDH treated by open reduction through an anterior approach in which a test of stability was used to assess the need for a concomitant osteotomy. The mean age at the time of surgery was 28 months (9 to 79) and at the latest follow-up, 17 years (12 to 25). All patients have been followed up until closure of the triradiate cartilage with a mean period of 15 years (8 to 23). At the time of open reduction before closure of the joint capsule, the position of maximum stability was assessed. A hip which required flexion with abduction for stability was considered to need an innominate osteotomy. If only internal rotation and abduction were required, an upper femoral derotational and varus osteotomy was carried out. For a 'double-diameter' acetabulum with anterolateral deficiency, a Pemberton-type osteotomy was used. A hip which was stable in the neutral position required no concomitant osteotomy. Overall, 86% of the patients have had a satisfactory radiological outcome (Severin groups I and II) with an incidence of 7% of secondary procedures for persistent dysplasia including one hip which redislocated. The results were better (p = 0.04) in children under the age of two years. Increased leg length on the affected side was associated with poor acetabular development and recurrence of joint dysplasia (p = 0.01). The incidence of postoperative avascular necrosis was 7%. In a further 18%, premature physeal arrest was noted during the adolescent growth spurt (Kalamchi-MacEwen types II and III). Both of these complications were also associated with recurrence of joint dysplasia (p = 0.01). Studies with a shorter follow-up are therefore likely to underestimate the proportion of poor radiological results. PMID- 10697310 TI - Vascularised rib graft defects of the diaphysis of the humerus in children. A report of two cases. AB - In our practice sequestration of the shafts of long bones in children because of acute osteomyelitis continues to be a problem. Conventional procedures for bone grafting are likely to fail. Vascularised grafts with microvascular anastomosis are technically demanding with a high rate of failure. Transfer of the rib on its vascular pedicle to achieve anterior fusion in the thoracic spine is now well established and the length of the pedicle available is adequate to allow grafting of a diaphyseal defect in the humerus. We describe the successful use of this procedure in two patients. PMID- 10697311 TI - Proximal entry for intramedullary nailing of the tibia. The risk of unrecognised articular damage. AB - The risk of articular penetration during tibial nailing is well known, but the incidence of unrecognised damage to joint cartilage has not been described. We have identified this complication in the treatment of tibial fractures, described the anatomical structures at risk and examined the most appropriate site of entry for tibial nailing in relation to the shape of the bone, the design of the nail and the surgical approach. We studied the relationship between the intra articular structures of the knee and the entry point used for nailing in 54 tibiae from cadavers. The results showed that the safe zone in some bones is smaller than the size of standard reamers and the proximal part of some nails. The structures at risk are the anterior horns of the medial and lateral menisci, the anterior part of the medial and lateral plateaux and the ligamentum transversum. This was confirmed by observations made after nailing 12 pairs of cadaver knees. A retrospective radiological analysis of 30 patients who had undergone tibial nailing identified eight at risk according to the entry point and the size of the nail. Unrecognised articular penetration and damage during surgery were confirmed in four. Although intramedullary nailing has been shown to be a successful method for treating fractures of the tibia, one of the most common problems after bony union is pain in the knee. Unrecognised intra articular injury of the knee may be one cause of this. PMID- 10697312 TI - Knee instability after injury to the anterior cruciate ligament. Quantification of the Lachman test. AB - We have measured anterior and posterior displacement in 563 normal knees and 487 knees with chronic deficiency of the anterior cruciate ligament (ACL). We performed stress radiography using a simple apparatus which maintained the knee at 20 degrees of flexion while a 9 kg load was applied. There was no significant difference in posterior translation dependent on the condition of the ACL. Measurement of anterior translation in the medial compartment proved to be more reliable than in the lateral compartment for the diagnosis of rupture of the ACL, with better specificity, sensitivity and predictive values. We have classified anterior laxity based on the differential anterior translation of the medial compartment and identified four grades in each of which we can further distinguish four subgrades for laxity of the lateral compartment. Within each of these subgroups, either internal or external rotation may dominate and sometimes there is a major translation of both compartments. Radiological evaluation of displacement of the knee in 20 degrees of flexion provides conclusive evidence of rupture of the ACL. A detailed study of pathological displacement is the basis for a classification of laxity. It is then possible to decide for each type of laxity, the surgical treatment which is specifically adapted to the lesion, and to define a reference value for judging outcome. PMID- 10697313 TI - Intraoperative heparin in addition to postoperative low-molecular-weight heparin for thromboprophylaxis in total knee replacement. AB - The administration of heparin during operation has been reported to enhance the efficacy of thromboprophylaxis in patients undergoing total hip replacement. We have performed a small pilot study in which intraoperative doses of heparin were given in addition to the usual postoperative thromboprophylaxis with enoxaparin in 32 patients undergoing total knee replacement. The primary endpoint was deep vein thrombosis (DVT) as demonstrated by bilateral venography on 6 +/- 2 days after operation. Sixteen patients developed DVT; in two the thrombosis was proximal as well as distal and in one the occurrence was bilateral. There was one major haemorrhage. These results are similar to those obtained with the use of postoperative thromboprophylaxis with enoxaparin alone. They do not provide support for the initiation of a larger randomised trial of this approach to management. PMID- 10697314 TI - Ultrasound-guided needle biopsy of primary bone tumours. AB - Needle biopsy is an established technique for the histological diagnosis of bone tumours, usually guided by fluoroscopy or CT. Surface lesions and aggressive tumours which have extended through the cortex are also amenable to imaging with ultrasound (US). We have assessed the diagnostic accuracy of US-guided Trucut needle biopsy in a consecutive series of patients referred to a Bone Tumour Unit with suspected primary bone tumours. Of 144 patients (83 men, 61 women; mean age 34.7 years) referred over a period of two years, 63 were considered suitable for US-guided biopsy. This was based on the presence of a relatively large extraosseous component, seen typically in osteosarcoma and malignant round-cell tumours. The results of needle biopsy were compared with those of surgical biopsy. The diagnostic accuracy was 98.4%, with only a single failed biopsy. Thus, in a selected group of patients, US is a very reliable technique of guidance for percutaneous needle biopsy of bone tumours. PMID- 10697315 TI - Experience in the treatment of dedifferentiated chondrosarcoma. AB - Dedifferentiated chondrosarcoma is a rare, highly malignant variant of chondrosarcoma in which a high-grade spindle-cell sarcoma coexists with a lower grade chondroid tumour. We have reviewed our experience with this neoplasm in 22 patients, all of whom were treated using modern oncological principles of planned resection and chemotherapy. Despite this the median survival was under nine months and only 18% were alive at five years. Those patients who received chemotherapy, and in whom wide margins of excision were achieved at operation, did best. It is essential to have an accurate preoperative diagnosis in order to plan treatment which may offer a better prospect of cure. PMID- 10697316 TI - Radical surgery for the solitary bony metastasis from renal-cell carcinoma. AB - We carried out excision of a solitary bony metastasis from renal-cell carcinoma in 25 patients in the hope that this would produce a prolonged disease-free interval. Two patients had excisions only, five had amputations and 18 had excision and endoprosthetic replacement. The one-, three- and five-year cumulative survival rates were 88%, 54% and 13%, respectively. There were three complications. One patient developed a local recurrence and three had problems related to the endoprosthesis. We recommend radical excision of a solitary bony metastasis from renal-cell carcinoma to achieve local control of the tumour for the remainder of the patient's life. PMID- 10697317 TI - The medial approach for operative release of post-traumatic contracture of the elbow. AB - We treated post-traumatic contracture of the elbow in 13 consecutive patients (14 elbows) by operative release. Through a single medial approach, the posterior oblique bundle of the medial collateral ligament was resected, followed by posterior and anterior capsulectomies. An additional lateral release through a separate incision was required in only four elbows. The results were assessed at a mean interval of 57 months after operation. Before surgery active extension lacked 43 degrees which improved to 17 degrees after operation. Active flexion before operation was 89 degrees, which improved to 127 degrees. The mean arc of movement increased from 46 degrees to 110 degrees. All 14 elbows showed scarring of the posterior oblique bundle of the medial collateral ligament. Neither the interval from injury to operative release nor the age of the patient affected the results. A medial approach is useful to reveal and excise the pathological changes in the medial collateral ligament. It is a safe and effective route through which to correct post-traumatic contracture of the elbow. PMID- 10697318 TI - The effectiveness of turnbuckle splinting for elbow contractures. AB - We have treated 22 patients with an elbow contracture using a static progressive turnbuckle splint for a mean of 4.5 +/- 1.8 months. All had failed to improve with supervised physiotherapy and splinting. The mean range of flexion before splintage was from 32 +/- 10 degrees to 108 +/- 19 degrees and afterwards from 26 + 10 (p = 0.02) to 127 +/- 12 degrees (p = 0.0001). A total of 11 patients gained a 'functional arc of movement,' defined as at least 30 degrees to 130 degrees. In eight patients movement improved with turnbuckle splinting, but the functional arc was not achieved. Six of these were satisfied and did not wish to proceed with surgical treatment and two had release of the elbow contracture. In three patients movement did not improve with the use of the turnbuckle splint and one subsequently had surgical treatment. Our findings have shown that turnbuckle splinting is a safe and effective treatment which should be considered in patients whose established elbow contractures have failed to respond to conventional physiotherapy. PMID- 10697319 TI - Anatomical reduction of intra-articular fractures of the distal radius. An arthroscopically-assisted approach. AB - We treated 31 intra-articular fractures of the distal radius by arthroscopically assisted reduction and percutaneous fixation with Kirschner (K-) wires. Tears of the triangular fibrocartilage (58 %), scapholunate (85 %) and lunotriquetral (61%) instability and osteochondral lesions (19%) were also treated. A total of 26 patients was independently reviewed at an average of 19 months. The mean pain score was 1.3/10, the range of movement 79% and the grip strength 90% of the contralateral wrist. Using the New York Orthopaedic Hospital score, 88% were graded excellent to good. On follow-up radiographs, 65% had no step and 31% had a step of < or =1 mm. Pain was significantly related to the size of the step. There was a significant difference in the incidence of persistent scapholunate diastasis and the Leibovic and Geissler grade (p < 0.01): I (0%), II (0%), III (42%) and IV (100%). We recommend anatomical reduction and acceptance of a step of <1 mm since the size of the step is related to the incidence of pain. PMID- 10697320 TI - Bone mineral density of the radius in patients with Colles' fracture. AB - To ascertain whether patients with Colles' fracture should be investigated for osteoporosis and the risk of future fractures, we measured the bone mineral density of the distal radius of the other arm in 31 women patients and compared the results with those of a control group of 289 normal women. We divided the patients into two groups, those younger than 66 years and those older. In 25 patients we found values for bone mineral density which were lower than one standard deviation below the mean value for their age. Younger patients had a deficit greater than that expected for their ages. We believe that women with Colles' fracture should be evaluated routinely for osteoporosis, particularly if they are under 66 years of age. PMID- 10697321 TI - Dermofasciectomy in the management of Dupuytren's disease. AB - Dupuytren's disease may present with well-defined subcutaneous cords or as more diffuse disease with involvement of the skin. Fasciectomy is the procedure commonly carried out for the full range of disease, but is associated with rates of recurrence of up to 66%. We reviewed 143 rays in 103 patients undergoing dermofasciectomy for diffuse disease with involvement of the skin. We found recurrence in 12 rays (8.4% of rays; 11.6% of patients) during a mean follow-up of 5.8 years, eight as cords and four as nodules. We suggest that dermofasciectomy is a better method of disease control than fasciectomy for the more diffuse type of disease with involvement of the skin. PMID- 10697322 TI - The image intensifier as an operating table--a dangerous practice. PMID- 10697323 TI - The Stanmore total hip replacement. A 22-year follow-up. AB - From a series of 135 patients (146 prostheses) who had had primary hip replacement in 1975 and 1976 we reported the outcome at ten years in 83 surviving patients in 1988 and that at 15 years in 44 surviving patients in 1994. Now, 22 years after the operation, we have reviewed the 21 patients who are still alive. Nineteen (20 hips) of these 21 patients (22 hips) with a mean age of 85.7 years still had their original prosthesis. Most patients were satisfied with the result, although the level of activity in many was reduced because of increasing age and other medical problems. The stem was stable in all 20 hips. Only one cup was definitely loose. Wear was observed in 40% of the cups but this was not a clinical problem. At the 22-year follow-up the cumulative survival rate of the prosthesis was 85%, of the stem 91% and of the cup 88%. Since 1975, 11 (7.5%) of the original 146 prostheses have been revised. PMID- 10697324 TI - Impaction bone grafting in revision hip surgery. A high incidence of complications. AB - We have reviewed retrospectively 68 revisions of the femoral component in arthroplasties of the hip in 65 patients, using impaction bone grafting, at a median of three years (1 month to 6 years). We employed the cemented Exeter X Change technique in 36 patients and the uncemented Bi-Metric allografting method in 32. The 37 bone defects were grade 3 or grade 4 on the Endo-Klinik classification. The Mayo hip score improved from a mean of 32 (SD +/- 18) to 62 (SD +/- 15). Most (25) of the 34 complications occurred in grade-3 and grade-4 defects; nine were intraoperative diaphyseal fractures and eight fractures of the greater trochanter. All the fractures united. The risk of intraoperative fracture was prevented by supporting the bone with wires in 16 hips, with reinforcement mesh in 18 and by a plate in six. Early migration of the stem of more than 10 mm during the first year indicated rotational instability; it occurred in three cases. In difficult revision cases with large defects of the femoral bone, bone impaction techniques carry a high risk of complications. PMID- 10697325 TI - Loosening of the cup after low-friction arthroplasty in patients with acetabular protrusion. The importance of the position of the cup. AB - Between 1972 and 1990, we performed 168 primary low-friction arthroplasties in 125 patients with acetabular protrusion. Twelve hips were lost to follow-up within eight years and eight which became infected were excluded from the final study. Of the 148 hips remaining, 62 with a mild protrusion were classified as group 1, 54 with moderate or severe protrusion as group 2 and, after 1985, 32 with moderate and severe protrusion which required bone grafts as group 3. The mean follow-up was 18.3 years (3 to 24) for group 1, 17.4 years (8 to 22) for group 2 and ten years (8 to 13) for group 3. There were 31 revisions of the cup, 12 in group 1 and 19 in group 2. According to the Kaplan-Meier analysis the overall rates at 20 years were 21 +/-10.79% in group 1 and 37 +/- 11.90% in group 2. There have been 43 radiological loosenings: 22 in group 1, 21 in group 2 and none so far in group 3, at ten years. The overall loosening rates at 20 years were 42 +/-14.76% in group 1 and 49 +/- 19.50% in group 2. The grafts were well incorporated in all group-3 hips, and the bone structure appeared normal after one year. The distance between the centre of the head of the femoral prosthesis and the approximate true centre of the femoral head was less in group 3 than in groups 1 and 2 (p < 0.01). According to the Cox proportional-hazards regression this was the single most important factor in loosening of the cup (odds ratio 1.11; 95% CI 1.05 to 1.18/mm). Better results were obtained in moderate and severe protrusions reconstructed with bone grafting than in hips with mild protrusion which were not grafted. PMID- 10697326 TI - Graded compression stockings in elective orthopaedic surgery. An assessment of the in vivo performance of commercially available stockings in patients having hip and knee arthroplasty. AB - We recruited 89 patients who had hip or knee replacements to assess the performance of below-knee graded compression stockings. The pressure gradients generated by the stockings were measured and all patients had venography of the ipsilateral leg. We found that 98% of stockings failed to produce the 'ideal' pressure gradient (+/- 20%) of 18, 14 and 8 mmHg from the ankle to the knee, while 54% produced a 'reversed gradient' on at least one occasion during the course of the study. The overall rate of deep-venous thrombosis was 16.7%. Stockings which produced reversed gradients were associated with a significantly higher incidence of deep-venous thrombosis (p = 0.026) than those with the correct gradient (25.6% v 6.1%). This suggests that the performance of graded compression stockings can be improved if reversed pressure gradients are detected and prevented. PMID- 10697327 TI - Periprosthetic bone remodelling around a prosthesis for distal femoral tumours. Measurement by dual-energy X-ray absorptiometry (DEXA). AB - We used dual-energy x-ray absorptiometry (DEXA) to evaluate the extent of periprosthetic bone remodelling around a prosthesis for distal femoral reconstruction, the Kotz modular femoral tibial replacement (KMFTR; Howmedica, Rutherford, New Jersey). A total of 23 patients was entered into the study which had four parts: 1) 17 patients were scanned three times on both the implant and contralateral legs to determine whether the precision of DEXA measurements was adequate to estimate bone loss surrounding the anchorage piece of the KMFTR; 2) in 23 patients the bone mineral density (BMD) in different regions of interest surrounding the diaphyseal anchorage was compared with that of the contralateral femur at the same location to test whether there was consistent evidence of loss of BMD adjacent to the prosthetic stem; 3) in 12 patients sequential studies were performed about one year apart to compare bone loss; and 4) bone loss was compared in ten patients with implants fixed by three screws and in 13 without screws. The mean coefficients of variation (SD/mean) for the 17 sets of repeated scans ranged from 2.9% to 7.8% at different regions of interest in the KMFTR leg and from 1.4% to 2.5% in the contralateral leg. BMD was decreased in the KMFTR leg relative to the contralateral limb and the percentage of BMD loss in general increased as the region of interest moved distally in the femur. Studies done after one year showed no consistent pattern of progressive bone loss between the two measurements. The ten patients with implants fixed by screws were found to have a mean loss of BMD of 42% in the most distal part of the femur, while the 13 without screw fixation had a mean loss of 11%. DEXA was shown to have adequate precision to evaluate loss of BMD around the KMFTR. This was evident relative to the contralateral leg in all patients and generally increased in the most distal part of the femur. In general, it stabilised between two measurements taken one year apart and was greater surrounding implants fixed by cross-locking screws. PMID- 10697328 TI - Treatment with ibandronate preserves bone in experimental tumour-induced bone loss. AB - Cancer-induced bone diseases are often associated with increased bone resorption and pathological fractures. In recent years, osteoprotective agents such as bisphosphonates have been studied extensively and have been shown to inhibit cancer-related bone resorption in experimental and clinical studies. The third generation bisphosphonate, ibandronate (BM 21.0955), is a potent compound for controlling tumour osteolysis and hypercalcaemia in rats bearing Walker 256 carcinosarcoma. We have studied the effect of ibandronate given as an interventional treatment on bone strength and bone loss after the onset of tumour growth in bone. Our results suggest that it is capable of preserving bone quality in rats bearing Walker 256 carcinosarcoma cells. Since other bisphosphonates have produced comparable results in man after their success in the Walker 256 animal models our findings suggest that ibandronate may be a powerful treatment for maintaining skeletal integrity in patients with metastatic bone disease. PMID- 10697329 TI - Growth factors improve muscle healing in vivo. AB - Injury to muscles is very common. We have previously observed that basic fibroblast growth factor (b-FGF), insulin growth factor type 1 (IGF-1) and nerve growth factor (NGF) are potent stimulators of the proliferation and fusion of myoblasts in vitro. We therefore injected these growth factors into mice with lacerations of the gastrocnemius muscle. The muscle regeneration was evaluated at one week by histological staining and quantitative histology. Muscle healing was assessed histologically and the contractile properties were measured one month after injury. Our findings showed that b-FGF, IGF and to a less extent NGF enhanced muscle regeneration in vivo compared with control muscle. At one month, muscles treated with IGF-1 and b-FGF showed improved healing and significantly increased fast-twitch and tetanus strengths. Our results suggest that b-FGF and IGF-1 stimulated muscle healing and may have a considerable effect on the treatment of muscle injuries. PMID- 10697330 TI - Parathyroid hormone (1-34) increases the density of rat cancellous bone in a bone chamber. A dose-response study. AB - Intermittent treatment with parathyroid hormone I(PTH) has an anabolic effect on both intact cancellous and cortical bone. Very little is known about the effect of the administration of PTH on the healing of fractures or the incorporation of orthopaedic implants. We have investigated the spontaneous ingrowth of callus and the formation of bone in a titanium chamber implanted at the medioproximal aspect of the tibial metaphysis of the rat. Four groups of ten male rats weighing approximately 350 g were injected with human PTH (1-34) in a dosage of 0, 15, 60 or 240 microg/kg/day, respectively, for 42 days from the day of implantation of the chamber. During the observation period the chamber became only partly filled with callus and bone and no difference in ingrowth distance into the chamber was found between the groups. The cancellous density was increased by 90%, 132% and 173% in the groups given PTH in a dosage of 15, 60 or 240 microg/kg/day, respectively. There was a linear correlation between bone density and the log PTH doses (r 2= 0.6). Our findings suggest that treatment with PTH may have a potential for enhancement of the incorporation of orthopaedic implants as well as a beneficial effect on the healing of fractures when it is given in low dosages. PMID- 10697331 TI - The influence of stiffness of the fixator on maturation of callus after segmental transport. AB - The treatment of large bony defects by callus distraction is well accepted, but the duration of treatment is long and the rate of complications increases accordingly. We have examined the effect of the stiffness of the axial fixator on reducing the time for maturation of callus. We created a mid-diaphyseal defect of 15 mm in the metatarsal bone in sheep and stabilised it with a ring fixator. After four days a bony segment was transported for 16 days at 1 mm per day. After 64 days the animals were divided into four groups, three with axial interfragmentary movement (IFM) of 0.5, 1.2 and 3.0 mm, respectively, and a control group. The 3.0 mm IFM group had the smallest bone density (p = 0.001) and area of callus and the largest IFM after 12 weeks; it also had typical clinical signs of hypertrophic nonunion. The most rapid stiffening of the callus was in the 0.5 mm group which had the smallest IFM (p = 0.04) after 12 weeks and radiological signs of bridging of the defect. These results indicate that suitable dynamic axial stimulation can enhance maturation of distraction callus when the initial amplitude is small, but that a large IFM can lead to delayed union. PMID- 10697332 TI - Outcome of Charnley total hip replacement across a single health region in England. PMID- 10697333 TI - Outcome of Charnley total hip replacement across a single health region in England. PMID- 10697334 TI - Biomechanical comparison of fixation of type-I fractures of the lateral tibial plateau. PMID- 10697335 TI - Planar anteversion of the acetabular cup as determined from plain anteroposterior radiographs. PMID- 10697336 TI - Questions raised in anaesthesia debate. PMID- 10697337 TI - Questions raised in anaesthesia debate. PMID- 10697338 TI - Women and the world of dentistry. AB - It was only in 1895 that the first woman dentist in the UK graduated from Edinburgh Dental School, and a further 17 years until a women was granted a dental qualification from The Royal College of Surgeons of England. At around this time cartoons began to appear, flippantly depicting women to be working in a profession regarded by many as masculine. Over the following years women dentists became more accepted, although as recently as the 1960's women were encouraged to enter certain branches of the profession where it was thought that they would be most useful. Government publications of this era encouraged women dentists to join the Maternity and Child Welfare Service and the School Health Service. It was felt that this work would be particularly suitable for them and that child patients would react more favourably to women dentists. PMID- 10697339 TI - Chairside options for the treatment of complete denture problems associated with the atrophic (flat) mandibular ridge. AB - This article outlines a number of simple clinical steps which should enable general dental practitioners to diagnose and treat the majority of complete denture problems in patients with atrophic mandibular ridges. The guiding principle is the reduction of forces transmitted to the denture-bearing area via the lower denture. Methods of optimising the size of the denture bases and/or the occlusal tables are discussed and illustrated. PMID- 10697340 TI - A national survey of infant feeding in Asian families: summary of findings relevant to oral health. AB - In 1994 the Department of Health commissioned a survey of early feeding practices within the Asian community. The survey conducted by the Office for National Statistics (ONS) was designed to assess from a nationally representative sample of infants from the Bangladeshi, Indian and Pakistani communities from across England their feeding practices from birth to 15 months. A small sample of White infants were also included in the sample although they cannot be considered as nationally representative of the majority White population in England. Data was collected from 2382 mothers in total. The survey was carried out between 1994 and 1996 and was published in 1997. This article will summarise the main findings relevant to oral health. These include patterns of breastfeeding, use of bottles, care and advice on infant feeding, weaning practices and drinks consumption. Implications for oral health promotion are considered. PMID- 10697341 TI - Perceived risk of future pathology associated with pathology-free third molars: a comparison of oral and maxillofacial surgeons and family dentists. AB - OBJECTIVE: To examine and compare practitioners' judgements of risk of future pathology associated with pathology-free disease asymptomatic third molars. SUBJECTS: 10 oral and maxillofacial surgeons and 18 family dentists (90% male) with experience ranging from 5-28 years. METHOD: Participants were presented with periapical radiographs of 36 asymptomatic, disease-free mandibular third molars and were informed of the age and sex of the patients and the degree of eruption of the third molars. Participants were asked to assess likelihood of future pathology in general, and more specifically, likelihood of root resorption, pericoronitis, periodontitis, cystic change and neoplasia if the third molar was left in situ. RESULTS: There was significant variation between the 28 raters but not between the two groups. Excepting assessment of future cystic change, there was no evidence that oral and maxillofacial surgeons and family dentists rated the 36 cases in consistently different ways. CONCLUSIONS: Practitioners varied very considerably in their judgment of the risks of pathology associated with asymptomatic disease-free third molars. Specialisation, did not account for this variation. PMID- 10697342 TI - An investigation of the relative efficacy of Buckley's Formocresol and calcium hydroxide in primary molar vital pulp therapy. AB - OBJECTIVE: To compare the clinical and radiological outcomes following two different, single visit vital pulp therapy techniques, in cariously exposed primary molar teeth. SETTING: A paediatric dental clinic within the Dental Hospital, Newcastle upon Tyne, UK. SUBJECTS: Fifty two child patients were sequentially enrolled in the clinical investigation, 26 males and 26 females with an age range of 3.3-12.5 years. Primary molar teeth requiring vital pulp therapy were randomly allocated to either the formocresol group (F) or the calcium hydroxide group (C). The total number of teeth treated was 84. DESIGN: Recruitment was on the basis of strict inclusion criteria. Coronal pulp amputation was prescribed only in teeth with vital, cariously exposed pulp tissue. Treatment was undertaken between October 1994 and December 1996. All cases were reviewed using predefined clinical and radiological criteria. The statistical tests used were logistic regression of a triple nested data structure, chi-squared analysis of equality of treatment and probability of success with relation to subject age. RESULTS: Eighty-four cariously exposed primary molars required vital pulp therapy. Forty six (55%) teeth were included in the F group and 38 (45%) allocated to the C group. Five teeth were lost to follow-up, leaving 79 teeth: forty four (56%) in group F and 35 (44%) in group C. Eighty four percent (37/44) of teeth treated with formocresol and 77 percent (27/35) treated with calcium hydroxide were classed as clinically and radiographically successful at the cut-off date, December 1997, after a mean clinical review of 22.5 months (range 6.1-38.5 months) and a mean radiographic review of 18.9 months (range 1.3-36.9 months). CONCLUSION: This investigation confirms the clinical efficacy of a one-fifth dilution of Buckley's Formocresol as an agent in pulp treatment of cariously exposed, vital primary molar teeth. However, calcium hydroxide in its pure, powder form is a clinically acceptable alternative when combined with strict selection criteria for this method of restorative care. There was a statistically insignificant difference in successful clinical and radiological outcome between the two treatment groups. Success was unrelated to the duration of time taken to achieve haemostasis and the presence or absence of bleeding after placement of the medicament. PMID- 10697343 TI - The value of patient feedback in the audit of TMJ arthroscopy. AB - AIM: This study assessed the patients' and clinicians' perception of the outcome of temporomandibular joint arthroscopy. METHOD: All patients who underwent TMJ arthroscopy for both diagnostic and therapeutic purposes over a 6-year period were sent a questionnaire that asked about various symptoms attributable to the TMJ. Additionally a review of the clinical notes was performed. RESULTS: 83 patients underwent arthroscopy to 127 temporomandibular joints. The mean follow up was 3.6 years. 55% of patients assessed their jaw function as being effective, jaw movement, pain control, and overall satisfaction were satisfactory in 37%, 57%, and 48% of cases respectively. The clinicians' assessment revealed that 45% of patients had no joint tenderness, 74% of patients were able to open to > 35 mm and 74% of patients were free of any joint noise. 66% of patients were prepared to undergo a second procedure if indicated. CONCLUSION: Overall, 50% of patients seemed to view arthroscopy favourably although many patients still felt that jaw opening was restricted. The outcome was not related to the position and reducibility of the disc at surgery and other variables may be responsible. The disparity between the clinical evaluation and the patients' perception of effectiveness emphasises the importance of patient feedback. PMID- 10697344 TI - Development and preliminary evaluation of an instrument designed to assess dental students' communication skills. AB - The aim of this study was to develop, and assess the inter-observer reliability of an instrument for evaluating dental students' communication skills. Methods used were process-tracking of interactions between experienced practitioners and patients, development of the instrument and its simultaneous use by two researchers observing 43 third year dental students prior to communication skills training. The results found that the instrument was appropriate for the purpose for which it was designed, and was easy to utilise. There were no significant differences between observers' total scores. Item-specific weighted kappa scores showed almost perfect agreement between observers for all but four of the 31 items. The lowest interobserver weighted kappa score was for the measurement of eye contact (k = 0.60). In conclusion, assessment of communication skills is now a necessity in the undergraduate curriculum. Preliminary analysis of an instrument of communication skills in the dental surgery indicates that it may be possible to do this reliably. PMID- 10697345 TI - Where is UK general dental practice going? AB - Dentistry is a fundamental part of healthcare; without it a large proportion of the population would suffer severe detriment to their general health. However, in order to achieve our professional aims we have to pursue the business of dentistry. PMID- 10697346 TI - Trying to keep it simple. PMID- 10697347 TI - National Alliance for Equity in Dental Health: report of the 3rd Annual Symposium on Inequalities in Dental Health held at the House of Commons on 2 November 1998. PMID- 10697348 TI - Preference based measurements in dentistry: a review of the literature and recommendations for research. AB - Clinical outcomes are used routinely in dental clinical practice to determine whether or not a patient has exhibited improvement following treatment. While these measures can be useful in comparing therapies in disease specific terms, they do not incorporate outcomes which may be of interest to patients. Preference based outcome measurements, on the other hand, take into account an individual's life style, overall well being and economic resources. There is a number of preference based measures available from the medical field, many of which have been adapted for use in a dental setting. This paper outlines the strengths and weaknesses of these preference based measures, using examples from the dental literature when available. Particular emphasis is placed on an economic tool known as contingent valuation or 'willingness to pay' as a potential technique in the measurement of dental preferences. PMID- 10697349 TI - The Xerostomia Inventory: a multi-item approach to measuring dry mouth. AB - OBJECTIVE: To develop a valid multi-item method of measuring the symptoms of xerostomia which includes the wide range of xerostomia symptoms in a single quantitative measure. DESIGN: A combination of qualitative and quantitative approaches. SETTING: A cohort study in South Australia. PARTICIPANTS: Older people aged 65 years or more who were taking part in the South Australian Dental Longitudinal Study. MEASURES: Xerostomia symptoms were evaluated using a multi item inventory format and, for comparison purposes, a standard single dry-mouth question. Resting whole-salivary flow rate was estimated using the 'spit' method. RESULTS: Xerostomia and flow-rate data were available for 636 individuals. Factor analysis revealed the presence of a discrete xerostomia dimension, represented by 11 items whose responses were summated to give a single Xerostomia Inventory (XI) scale score. This had a very low correlation with resting flow rate but a much stronger, positive correlation with the standard dry-mouth question responses. CONCLUSIONS: The XI shows adequate content and concurrent validity, and appears to be a promising advance on previous approaches to xerostomia symptomatology although further testing is required. PMID- 10697350 TI - Caries experience and sucrose availability: an analysis of the relationship in the United Kingdom over fifty years. AB - OBJECTIVE: A previous study suggested that the most likely explanation for the rise and subsequent fall in caries in children in England and Wales during the last 50 years was the concurrent increase and then reduction in the sugar challenge to the population, mitigated after the early 1970s by the preventive effect of fluoride toothpaste. The current objective was to quantify the relationship between sucrose available for consumption annually since 1948 and caries experience at 5 and 12 years. METHOD: In a retrospective, longitudinal ecological study, cross-sectional mean dmft values at 5 years and DMFT values at 12 years in England from 1948 to 1968 and England and Wales from 1973 to 1996/97, from three series of standardised surveys, were regressed on data for the relevant years on sucrose available for consumption in the UK. RESULTS: For sucrose consumption and 5-year-old dmft, Spearman's rho was +0.62 (P < 0.05) while for 12-year-old DMFT the value was +0.84 (P < 0.001). For the 12 year age group, Pearson's coefficient could also be calculated (r = +0.87; P < 0.001). CONCLUSION: For several reasons caution should be used in interpreting these findings. Nevertheless they do suggest a strong positive correlation over time between dmft/DMFT and sucrose availability nationally. PMID- 10697351 TI - The dental health of pre-school children in a deprived urban community in Glasgow. AB - OBJECTIVE: This study investigated the dental health status of pre-school children in a deprived urban community in Greater Glasgow. The aim was to gather baseline data to support the need for a multi-agency dental health programme for this age group and against which trends in dental health could be measured over time. METHOD: A defined deprived community was identified and an area profile compiled. Children attending the five nursery schools in and around the area were examined using the standardised criteria adopted by BASCD/SHBDEP. RESULTS: Two hundred and forty-eight children were examined representing 75.8% of those on the nursery rolls. Caries prevalence and mean dmft rose from 64% and 3.14 for three to three and a half-year-old children to 86% and 6.14 for four and a half- to five-year-old children. This latter figure was higher than the Scottish and Health Board averages for five-year-old children (2.93 and 3.5 respectively). Those from the most deprived postcode sector had significantly worse dental health than those resident in other areas (mean dmft = 6.50 compared with 3.77). They also had significantly more unrestorable lesions. Overall, the Care Index (ft/dmft x 100) was 3.03 which is less than the Scottish average of 7.8. CONCLUSION: The dental health of nursery school children in and around the Possilpark area of Glasgow is worse than both the Scottish and health board averages for five-year-old children. Those resident in the most deprived sector of the community have significantly worse dental health. The main components of dmft were untreated decay and missing teeth. The Care Index was low. PMID- 10697352 TI - Views of parents and head teachers on the school dental screening service in a north of England city. AB - OBJECTIVE: The study was designed to explore parents' and head teachers' understanding of and attitudes to the process, objectives and perceived outcomes of a school dental screening. BASIC RESEARCH DESIGN: Qualitative methods were used as a source for questionnaire development and questionnaires were subsequently used to test the strength of agreement with the qualitative findings. PARTICIPANTS: Head teachers and parents of a sample of 5- and 10-year old children attending primary schools in Sunderland. RESULTS: Questionnaire responses from 83 (79%) of the head teachers and 934 (82%) parents of 5- and 10 year-old children showed that the exercise was well accepted by the majority. Gaps were identified in many aspects of communication between the Community Dental Service, schools and parents. CONCLUSIONS: It is recommended that, taking into account the views of all involved, the objectives of the exercise should be clearly defined and then clearly communicated. Evaluation should follow, measuring the extent to which the defined objectives are met. In this way the process will make a more positive and relevant contribution to oral health and the provision of primary dental care. PMID- 10697353 TI - Dental beliefs, knowledge and behaviour of Chinese people in the United Kingdom. AB - OBJECTIVE: This study explored oral health beliefs, knowledge and behaviour among a sample of United Kingdom Chinese. DESIGN: A quota sample of Chinese people, stratified by age and gender, were interviewed by trained and standardised Chinese interviewers using a piloted, validated semi-structured questionnaire. SUBJECTS: One hundred and fifty-six Chinese people--with similar number of teenagers, younger adults and older people--resident in the North East of England. SETTING: Chinese communities. OUTCOME MEASURE: Reported dental knowledge, beliefs and behaviours. RESULTS: Regardless of gender and age, the majority of respondents believed that it was natural for people to lose all their teeth in old age. Less than half were convinced that they would be able to keep their own teeth for life. The majority of the sample considered that they were susceptible to dental diseases, the consequences of which were thought to be serious. Approximately half presumed that dental diseases were preventable, although the aetiology of dental caries, periodontal disease and tooth loss was poorly understood. While 94% claimed to brush their teeth as part of routine dental care, dental visiting and dietary restriction of sugar intake were reported only in 61% and 30% of the sample respectively. Inter-generational differences were marked; older people tended to have a fatalistic attitude and were least likely to attend the dentist. CONCLUSIONS: A low level of dental awareness was found among the UK Chinese. In order to facilitate effective health promotion and treatment services, the extent of Chinese people's traditional oral health beliefs and behaviour must be taken into account. PMID- 10697354 TI - Job satisfaction amongst rural medical aides providing emergency oral health care in rural Tanzania. AB - OBJECTIVE: To investigate to what extent the rural medical aides (RMAs) in Tanzania were satisfied with their new (added) role of providing emergency oral health care services, and to analyse factors influencing job satisfaction amongst them. DESIGN: Cross-sectional survey using a self-administered job satisfaction questionnaire. SETTING AND SUBJECTS: All 40 RMAs providing emergency oral health care in rural health centres and dispensaries in Mbeya and Tanga regions, Tanzania. MAIN OUTCOME MEASURE: RMAs ratings of their overall satisfaction with the job of providing emergency oral health care. RESULTS: Overall, 95% of the RMAs were satisfied with providing emergency oral health care. Patient relations, personal time and stress were significantly correlated with overall job satisfaction in providing emergency oral health care. CONCLUSIONS: The RMAs' newly added role of providing emergency oral health care does not seem to generate problems with job satisfaction. PMID- 10697355 TI - Dental caries experience among school children in St. Vincent and The Grenadines: report of the first national oral health survey. AB - OBJECTIVE: To determine the prevalence and severity of dental caries among school children of St. Vincent and The Grenadines. Also, to establish baseline data on dental caries and determine the extent to which the present oral health care system is meeting dental needs. BASIC RESEARCH DESIGN: National cross-sectional survey utilising the World Health Organization's pathfinder methods. Dental examinations were conducted from October to November 1991 by three trained examiners. PARTICIPANTS: A total of 1648 students, 21% of school children in St. Vincent and The Grenadines, were sampled through multistage systematic sampling. OUTCOME MEASURES: Dental caries was diagnosed clinically according to the World Health Organization's diagnostic criteria. RESULTS: Caries prevalence in the permanent dentition was 69.4%, ranging from 68 to 73% according to gender and geographic location. In the primary dentition caries prevalence was 76.6%. Mean DMFT for the survey population was 2.69, dmft was 3.25, while the DMFT scores for gender and location types varied from 2.39 to 3.25. DMFT at 12 years was 3.25. No difference in caries prevalence was observed between gender in the permanent dentition but prevalence was significantly different in the primary dentition being higher among boys. Prevalence was significantly higher in the urban population relative to the rural population. The decayed component constituted 92% of DMFT and 91% of dmft. CONCLUSION: Moderate caries severity and high levels of untreated decay were found in both primary and permanent teeth. The results emphasise the need for continuous surveillance and for appropriate intervention and prevention programmes. PMID- 10697356 TI - The dental caries experience of 5-year-old children in the United Kingdom. Surveys co-ordinated by the British Association for the Study of Community Dentistry in 1997/98. AB - OBJECTIVE: This paper reports the results of standardised clinical caries examinations of 176,781 5-year-old children from across the United Kingdom. These 1997/98 co-ordinated surveys are the latest in a series which seek to monitor the dental health of children and to assess the delivery of dental services. METHOD: The criteria and conventions of the British Association for the Study of Community Dentistry were used. Representative samples were drawn from participating health authorities and boards and caries was diagnosed at the caries into dentine threshold using a visual method without radiography or fibre optic transillumination. RESULTS: The results again demonstrated a wide variation in prevalence across the UK, with mean values for d3mft for the current English regions (of the National Health Service) and the other UK 'territories' ranging from 1.02 in the West Midlands to 2.92 in Northern Ireland. Mean d3mft across the UK was 1.68 (d3t = 1.18, mt = 0.26, ft = 0.23). Overall, 43% of children had evidence of caries experience (d3mft > 0), although the means ranged between 33% (West Midlands) and 63% (Northern Ireland). The distribution of caries was highly skewed. Thus the UK mean caries experience for those with disease was 3.94, as opposed to the overall mean of 1.68. Trends over time demonstrate a modest improvement of 8.7% in overall d3mft for Great Britain since 1995/96, compared to the small improvement seen two years ago and the deterioration seen four years previously. Both dt and mt have fallen while ft remained unchanged. The care index has increased in all but one region/territory (14% in 1997/8, compared to 12% in 1995/6). Regional/country means for 1997/8 ranged from 9-23%. This indicator has not, however, regained the levels seen in the past. CONCLUSION: There has been some improvement in the dental health of 5-year-old children. Overall, the provision of operative care for those with dentinal decay has also improved; however, significant groups remain within the population of 5-year-old children who have dental disease and who are in need of dental care. PMID- 10697357 TI - Some experience of using Super Profiles in oral health studies of children and adults in West Yorkshire. PMID- 10697358 TI - The sound and the fury: an update on the role of high-dose chemotherapy and autologous stem cell salvage for both metastatic and high-risk breast cancer. PMID- 10697359 TI - Perforation of jejunal diverticulum: case report and review of literature. AB - We report the case of a 90-year-old woman, previously diagnosed with jejunal and colonic diverticula, who presented with left lower quadrant abdominal pain suggesting either colonic diverticulitis or ischemic colitis. A computed tomography scan revealed a perforated jejunal diverticulum with abscess formation. The patient promptly was treated surgically without complications. A review of the literature indicates the rarity of perforation of jejunal diverticula and the difficulty of early diagnosis. We discuss the etiology, pathogenesis, diagnosis, and management of this rare entity. It is important for primary care physicians to be familiar with this disease. Delay in work-up often results in catastrophic consequences. PMID- 10697360 TI - The Laffer curve and HMOs: or, is managed care on the slippery downhill slope? PMID- 10697361 TI - Emerging drugs of abuse in Connecticut. AB - During the past decade, several new drugs of abuse have emerged as public health concerns in Connecticut. These include ketamine, gamma-hydroxybutyrate, methylphenidate (Ritalin), and "Illy." Two trends stand out when one looks at these new drugs of abuse. First, drugs that have legitimate medical indications are being increasingly diverted for illicit purposes. Second, much of the information needed to acquire, synthesize, and use these drugs can be found on the Internet. While none of these newer agents is abused to the extent of heroin or cocaine, all of them have the potential to cause serious morbidity, and occasionally death. The goal of this article is to make Connecticut physicians aware of these emerging drugs of abuse. Emphasis is placed on recognizing the signs and symptoms produced by these drugs and on managing the complications that are associated with their use. PMID- 10697363 TI - An interview with Dr. Gerald Labriola. Interview by H. David Crombie, with Tim Norbeck. PMID- 10697362 TI - Optimizing oral dosing and monitoring of noncardiac toxicities with chronic amiodarone therapy. PMID- 10697364 TI - Airlift to Baghdad. PMID- 10697365 TI - [Apoptosis in pathologic processes of digestive organs]. PMID- 10697366 TI - [Enteropathic amyloidosis: clinical features, place among other forms of amyloidosis]. PMID- 10697367 TI - [Present-day infectious endocarditis (part II)]. PMID- 10697368 TI - [Blood lipids and intensity of free radical oxidant processes in elderly patients with ischemic heart disease on antiatherogenic vegetarian diet]. AB - The authors studied the effects of balanced antiatherogenic vegetarian diet enriched with soya bean products on blood lipids and intensity of free radical oxidant processes in elderly patients with ischemic heart disease. 45 patients with dyslipoproteinemia type IIA or IIB were examined for hemodynamic parameters, lipid spectrum and intensity of free radical oxidation. The diet promoted a trend to normalization of central hemodynamics, significantly reduced the level of atherogenic lipids in blood, improved free radical lipid peroxidation and activity of nonenzymatic antioxidant defence. PMID- 10697369 TI - [Circadian index as indicator of stable organization of heart circadian rhythm]. AB - The author proposes to use circadian index (CI) as a method of evaluation of major structure of the circadian heart rhythm profile in Holter monitoring. CI is calculated as a ratio of awaking (7 a.m. to 22 p.m.) mean heart rate to sleep (23 p.m. to 6 a.m.) mean heart rate. CI was estimated by the literature data on 7648 healthy subjects and patients aged 2 to 79 years. In healthy subjects CI was stable irrespective of the sex and age. Normal CI makes up 1.33 +/- 0.05. All the cardiovascular patients exhibited increasing rigidity of the rhythm with the disease progression and deterioration of the prognosis. In patients with diabetes mellitus with vegetopathy rigidity of the circadian rhythm grows with aggravation of vegetopathy. In neurological patients free of vegetopathy CI was normal. Intensification of the heart rhythm profile was seen in patients with high sensitivity to catecholamines. PMID- 10697370 TI - [Echocardiographic evidence of heart state in patients with hypothyreosis]. AB - Echocardiographic examination of 41 patients has demonstrated that mitral prolapse (MP) of the 1st and 2nd degree is frequently associated with latent or apparent hypothyroidism and occasionally with prolapse of the tricuspid and/or aortic valve. As a rule, MP has a silent course without regurgitation and is diagnosed at ultrasonography. Its correction is made by replacement with thyroid hormones. Beta-adrenoblockers for MP treatment are contraindicated in hypothyroidism. MP complications (sudden death, thromboembolism, infectious endocarditis) are rare in patients with hypothyroidism. Thyroid insufficiency sometimes is accompanied with hydropericardium which is symptomless and is detected only at echocardiography. PMID- 10697371 TI - [Application of high information technologies in predicting outcomes of cardiovascular diseases]. AB - The authors review their studies conducted for many years on cardiological prognostication in the context of the republican program "Cardiology" performed in the Udmurt Republic. The lines of the research and results obtained are described. PMID- 10697372 TI - [Pain sensitivity in patients with rheumatoid arthritis]. AB - The paper deals with specific characteristics of pain sensitivity, types of anxiety disorders in patients with rheumatoid arthritis and their correlation. PMID- 10697373 TI - [Aspirin vs ticlid and their combination in patients with acute myocardial infarction]. AB - ADP-induced platelet aggregation was studied in 28 patients with myocardial infarction randomized, at admission, into three groups. Conventional therapy with heparin and antianginal drugs was combined with aspirin (250 mg/day, n = 9), tiklid (500 mg/day, n = 9), tiklid (500 mg/day) + aspirin (250 mg/day, n = 10) in group 1, 2 and 3, respectively. Tiklid diminished platelet aggregation more effectively than aspirin on the disease day 7 and 21. Tiklid + aspirin combination suppresses platelet aggregation more than monotherapy with either of the drugs, provides insignificant attenuation of postinfarction angina but is associated with a high risk of hemorrhagic complications. PMID- 10697374 TI - [Effects of metabolic therapy on intracellular level of antioxidant system in elderly patients with ischemic heart disease]. AB - Patients with ischemic heart disease (angina pectoris of functional class II and III) whose mean age was 67.4 +/- 1.67 years were divided into 3 randomized groups. 15 control patients received standard treatment. 30 and 15 patients of group 1 and 2, respectively, received standard treatment plus amino acid composition (100 mg 3 times a day sublingually) or preductal (20 mg 3 times a day)--groups 1 and 2, respectively. The treatment lasted 21 days. Before the treatment and on its day 1, 7 and 21 measurements were made of glutathione level, activity of glutathion-dependent enzymes, catalase, Cu, Zn-superoxide dismutase and malonic dialdehyde in red cells. The amino acid composition was found to raise the level of antioxidant system and suppress lipid peroxidation. Preductal raised the activity of Cu, Zn-superoxide dismutase and had an unbalanced effect on the antioxidant system. PMID- 10697375 TI - [Cardiac stimulator implantation in atrioventricular blocks complicating acute myocardial infarction: terms and indications]. PMID- 10697376 TI - [Abdominal pains]. PMID- 10697377 TI - [Diagnostic difficulties in tuberculous pneumonia]. PMID- 10697378 TI - [On the problem of diseases association]. PMID- 10697379 TI - [Medical terms from West Europe and their role in present-day Russian medical terminology, other disputable terminological problems]. PMID- 10697381 TI - [In Process Citation] PMID- 10697382 TI - [In Process Citation] PMID- 10697380 TI - [Medical professionals among A.S.Pushkin's friends (part I)]. PMID- 10697383 TI - The New Jersey Medicine interview. Harvey A. Holzberg. PMID- 10697384 TI - Medical science is the key to success in the 21st century. PMID- 10697385 TI - Improving pneumococcal vaccination rates for elderly patients. AB - Last year, the pneumococcal vaccination rate of 32% in our elderly clinic population was, like that of the state of New Jersey, well below the Healthy People 2000 target of 60% to 80%. During a nine-month period in which intervention strategies, including education and standing orders for nurses, were implemented, we achieved an improved rate of 63%. PMID- 10697386 TI - Teen-friendly services. Advocates for successful teen contraception. AB - Teenagers seeking contraceptive advice need teen-friendly services that are accessible, free or low-cost, confidential, and convenient. From the teen's perspective, the picture is quite simple: "I need help now, I don't want my parents to know about it, I have no money, and I'm not sure what to do." It is the responsibility of the clinical staff to walk new patients through the steps of becoming a successful health care consumer, keeping in mind that they have never acted independently in this area before. PMID- 10697387 TI - Communicating with patients who have hearing loss. AB - Access to health care services is of paramount interest to all New Jerseyans. For people with varying degrees of hearing loss, affordable health insurance, while important, is not the only barrier to quality health care. Doctors are required by law to ensure effective communication between themselves and their deaf or hard-of-hearing patients. PMID- 10697388 TI - Tea and health. AB - Tea is a pleasant, popular, socially accepted, economical, and safe drink that is enjoyed every day by hundreds of millions of people across all continents. Tea also provides a dietary source of biologically active compounds that help prevent a wide variety of diseases. It is the richest source of a class of antioxidants called flavonoids and contains many other beneficial compounds such as vitamins and fluoride. A growing body of evidence suggests that moderate consumption of tea may protect against several forms of cancer, cardiovascular diseases, the formation of kidney stones, bacterial infections, and dental cavities. Future research needs to define the actual magnitude of health benefits, establish the safe range of tea consumption associated with these benefits, and elucidate potential mechanisms of action. PMID- 10697389 TI - Changes in protein, carbohydrate, and fat metabolism with aging: possible role of insulin. AB - Age is associated with modifications of body composition, i.e., an increase in body fat mass and a decrease in protein mass. Because insulin controls substrate disposal and production, these changes could theoretically be related to changes in either insulin action or secretion on the various substrates. On the basis of available evidence, insulin action on whole-body amino acid and protein metabolism seems not to be impaired in the aged. Decreased synthesis of contractile and mitochondrial proteins in muscle, associated with decreased gene expression, was described in humans. Decreased physical activity apparently represents an important factor responsible for decreased muscle protein synthesis and mass in the elderly. Exercise in the elderly may acutely revert these changes, although its chronic effects are still uncertain. In addition, the possible interaction between insulin and exercise in the maintenance of muscle mass needs to be specifically investigated in aged people. Higher free fatty acid (FFA) absolute flux and oxidation rates were observed in healthy elderly subjects in both the fasting state and following hyperinsulinemia, but not when normalized over fat mass. This suggests that FFA kinetics reflect the established changes in fat mass. Insulin sensitivity on glucose metabolism is usually normal in the aged, despite subtle impairments in insulin secretion, hepatic uptake, and onset of action. Finally, data support the operation of the Randle cycle (i.e., inverse relationships between fat and glucose oxidation) in the elderly. PMID- 10697390 TI - Warfarin use and fracture risk. AB - Two recent studies examined the association between chronic use of warfarin, a vitamin K antagonist, and fracture rate among older women. Whereas one study reported no association, the other reported a significantly higher risk for vertebral and rib fractures among warfarin users compared with nonusers. The effect of vitamin K antagonists on age-related bone loss continues to be controversial. PMID- 10697391 TI - Discovery of the ceruloplasmin homologue hephaestin: new insight into the copper/iron connection. AB - Approximately 75 years ago Hart and colleagues discovered that copper deficiency impaired mammalian iron metabolism. Discovery of hephaestin identifies a critical new component of the copper and iron connection in mammals. Hephaestin appears to be a multicopper oxidase required for efficient export of iron from the intestine. PMID- 10697392 TI - Micronutrient shortfalls in young children's diets: common, and owing to inadequate intakes both at home and at child care centers. AB - Numerous studies documented low intakes of iron, zinc, calcium, and other micronutrients in young children. A recent report suggests that intakes are low in both home food and food provided at day care centers. Most of the young children in that study could not have obtained adequate intakes of key micronutrients without major dietary changes. Is it time to recommend routine multivitamin/mineral supplementation for all young children? PMID- 10697393 TI - [Contemporary rules for diagnosing hypertension]. PMID- 10697394 TI - [Insulin binding to erythrocyte receptors in patients with essential hypertension]. AB - Recently many researchers have described the presence of insulin resistance and hyperinsulinemia in a substantial number of patients with essential hypertension. Reduced insulin binding to the receptors may play important role in development of insulin resistance in these patients. The study was aimed to assess the value of insulin binding to erythrocyte receptors in the patients with essential hypertension and compare to values in healthy persons. Additional purpose was the evaluation of insulin degradation by erythrocytes in patients with essential hypertension. 23 patients with essential hypertension (BMI 22.7 +/- 3.2) and 21 healthy persons (with BMI value 23.3 +/- 2.9) were studied. In all examined individuals the blood glucose and blood insulin concentrations were determined, insulin binding to erythrocyte receptors and insulin degradation by erythrocytes were measured by the method of Gambhir and al. Insulin concentration was significantly higher in patients with essential hypertension than in healthy subjects. We demonstrated a statistically significant positive correlation between body weight and insulin concentration in blood serum only in healthy people. Insulin binding to the receptors of red blood cells was significantly stronger (p < 0.001) in healthy persons than in patients with essential hypertension (0.972 +/- 0.395 pg 10(11) RBC and 0.446 +/- 0.14 pg 10(11) RBC respectively). In patients with hypertension insulin binding to receptors of red blood cells does not depend on body weight and insulin concentration in blood serum. Values of insulin degradation by erythrocyte in patients with essential hypertension and healthy persons were not significantly different. It seems that decreased insulin binding to insulin receptors is an important mechanism of insulin resistance patients with essential hypertension. PMID- 10697395 TI - [Levels of tumor necrosis factor (TNF-alpha) and interleukin 6 (IL-6) in serum of patients with acute myocardial infarction]. AB - In acute myocardial infarction may increase the synthesis of cytokines, which can enlarge the myocardial lesion owing to their direct toxic action on myocytes or induction of inflammatory changes that lead to myocardiofibrosis. All this may quickening the appearance of congestive heart failure after myocardial infarction. The aim of the study was examination of tumor necrosis factor (TNF alpha) and interleukin 6 (IL-6) plasma levels in patients with acute myocardial infarction and analysis of correlation between concentrations of these cytokines and myocardial lesions during infarction. The study was made in 94 patients admitted to the Department of Cardiology with acute myocardial infarction (AMI). Of these, 40 were women aged from 41 to 85 (mean 67 years) and 54 were men aged from 39 to 86 (mean 63 years). Anterior AMI was diagnosed in 40 patients, inferior AMI was diagnosed in 54 patients. 63 patients underwent the thrombolytic therapy, reperfusion appeared in 45 patients, 24 patients were excluded from the thrombolytic therapy. Control group consisted of 28 healthy persons aged from 35 to 76 (mean 61 years). Blood samples for determination of TNF-alpha and IL-6 plasma levels were taken just after admission prior to the treatment. Then patients were taken streptokinase or tissue-type plasminogen activator with typical doses. Blood samples for determination of cytokines were obtained in 3. and 7. day after treatment. TNF-alpha and IL-6 plasma levels were determined with radioimmunological assay. Creatine kinase activity were monitored in patients with AMI as well as ejection fraction was checked in echocardiography in 3. and 7. day after treatment. We showed increased plasma levels of TNF-alpha and IL-6 in patients with AMI with maximum in 3. day of infarction. Concentrations of cytokines were higher in patients with anterior AMI than in patients with inferior AMI. In anterior infarction concentrations of cytokines were significantly lower after thrombolytic therapy with reperfusion than after treatment without reperfusion. There is a correlation between infarct size and concentrations of TNF-alpha and IL-6. PMID- 10697396 TI - [DQA1 and DQB1 HLA genes as markers of insulin-dependent diabetes mellitus in the Polish population]. AB - The second-generation screen of human genome has confirmed that HLA region genes play a key role in the susceptibility to insulin-dependent diabetes mellitus. The aim of the present study was to estimate the frequency of chosen alleles of DQA1 and DQB1 genes in the patients with insulin-dependent diabetes mellitus and their first degree relatives in comparison to the healthy population in the north eastern region of Poland. HLA typing of DQA1 and DQB1 alleles of the HLA region was performed by "phototyping" PCR-SSP method. The highest predisposition to IDDM in the population of the north-eastern region of Poland was associated with DQB1*0302 allele and DQB1*02-DQA1*0301 or DQB1*0302-DQA1*0301 haplotypes, while the dominant protection was connected with DQB1*0602, DQB1*0603 and DQB1*0301 alleles. The high frequency of protective DQB1*0602 and DQB1*0603 alleles and the low percentage of "diabetogenic" DQB1*0302 genes in the healthy control population of north-estern region of Poland may suggest their dominant influence on the relatively low incidence of IDDM in this region. The relatively high percentage of the first degree relatives of IDDM patients with pancreatic autoantibodies but with protective alleles observed in our study, which significantly decreases the risk of IDDM, suggests the necessity of DQB1*0602-03 measurements in such subjects for the better IDDM risk assessment. PMID- 10697397 TI - [Stem cell factor (SCF) in diagnosis and monitoring of non-small-cell lung cancer]. AB - Lung cancer, of which non-small-cell lung cancer (NSCLC) constitutes about 80%, is the greatest cause of cancer-related death worldwide. Serum tumour markers may be helpful in early diagnosis, in the initial assessment of the progress of the disease and in monitoring of the tumour growth or tumour volume reduction. Recent studies have focused on a new family of markers--hematopoietic growth factors. Some clinical investigations have shown autologous production of stem cell factor (SCF) in various human cell lines derived from lung cancer and the expression of SCF mRNA in these lines. In this study, the serum level of SCF was measured using a sensitive sandwich ELISA system in 34 patients with non-small-cell lung cancer before and 10, 30, 90, 180 and 270 days after operation. Additionally common accepted tumour markers such as CEA and CYFRA 21.1 were also assayed. Preoperative level of SCF was increased in cancer patients in comparison to the normal sera. Concentrations of SCF and CYFRA 21.1 were decreased on 10th day, but CEA on 30th day after surgical treatment, although upon comparison of pre- and postoperative tumour markers serum levels significant difference was observed for SCF and CYFRA 21.1 (p < 0.05). Levels of SCF were increased in 79%, CEA in 62% and CYFRA 21.1 in 51%. The diagnostic sensitivity of SCF were related to the stage of the disease and the combined use of two markers increased the sensitivity compared with the use of only one. These results suggest that SCF may be useful in the diagnostic and monitoring of patients with NSCLC. PMID- 10697398 TI - [Insulin resistance in offspring of type 2 diabetic patients]. AB - Diabetes mellitus is the disease with heterogeneous aetiology. Among the causes of hyperglycaemia the insulin resistance with it's genetical background is mentioned. The aim of the study was the assessment of insulin resistance in healthy offspring of type 2 diabetic patients as well as assessment whether the coexistence of nephropathy in parents has an impact on insulin resistance in offspring. 56 subjects with positive familial history of diabetes type 2 divided into 2 groups were admitted. Subgroup A1 30 subjects (mean age 33.0 +/- 8.5 years) consisted of those who had familial history of diabetes without nephropathy and subgroup A2 26 subjects (mean age 33.0 +/- 6.5 years) with familial history of diabetic nephropathy. Control group consisted of 30 healthy volunteers without familial history. Euglycemic hyperinsulinemic clamp test was performed in all subjects studied. Tissue glucose uptake (TGU) was significantly lower while fasting insulinemia In0 was significantly higher in A1 and A2 groups when compared to controls (respectively TGU 5.6 +/- 2.2, 6.3 +/- 2.5 and 9.5 +/- 2.2 mg/kg/min p < 0.005, In0 19.4 +/- 8.3, 20.8 +/- 8.9 and 11.4 +/- 6.0 p < 0.001). No differences in TGU and In0 when compared A1 vs. A2 group were found. In-depth analysis did not show any differences in relation on whether diabetes was inherited from father's or mother's side. It was also shown that BMI did not interfere on insulin resistance in patients with positive familial history of diabetes. We conclude that insulin resistance has the genetical background and that insulin resistance and nephropathy are inherited separately. PMID- 10697399 TI - [The usefulness of MIBI scintigraphy for postoperative monitoring of patients with thyroid cancer]. AB - Metoxyizobutyloizonitrile labelled with technetium 99mTc is a radio pharmaceutical that was shown to accumulate in benign and cancerous thyroid tissue. As it can be applied without thyroid hormone withdrawal this gave a stimulus to the investigations on its usefulness in diagnostic and follow up procedures for thyroid cancer patients. The goal of this study is to evaluate the efficacy and benefit of 99mTc-MIBI whole body scintigrams in post surgery follow up of patients with differentiated thyroid cancer. One hundred and twenty eight 99mTc MIBI scintigraphy were performed and evaluated. Sensitivity of MIBI scans was the highest for bone metastases--79%. Good results were also obtained for lymph node metastases (sensitivity--73%, specificity--90%). In case of lung metastases the sensitivity and specificity were 21% and 94% respectively. Sensitivity of detection of clinically apparent recurrent disease in thyroid bed was 70% and specificity of visualization 78%. Results of our study demonstrate that 99mTC-MIBI is valuable tool in follow up of thyroid cancer patients, but can not replace 131I scintygraphy. PMID- 10697400 TI - [Results of promoting fibrinolytic therapy for myocardial infarction in the Katowice Province from 1996-1998]. AB - The action promoting fibrinolytic treatment of myocardial infarction and questionnaire-based follow up of the results was carried out over a four million people area. 5262 questionnaires, in which physicians answered a number of questions, were received. The poll revealed that 79% of patients received fibrinolytic treatment. The authors estimate the overall percent of patients, treated in hospital wards, at about 27.7%. The mortality rate in the group of patients, who received streptokinase, was 8.61% and in the group of patients, who did not receive the treatment, was 16.28%. The signs of reperfusion were present in 60.38% of patients. The authors conclude that the increase in the number of patients administered fibrinolytic treatment should be aimed, and the results obtained in this group of patients come close to published data. Exercise tests are performed in too small number of myocardial infarction patients and only little proportion of them undergoes further coronary angiography. PMID- 10697401 TI - [Hypoglycemia during the course of chronic renal failure]. AB - In two patients with end stage renal failure and not treated diabetes spontaneous hypoglycaemiaes were observed. The lowest levels of glucose in blood serum were: 1.9 mmol/L and 1.16 mmol/L. These levels were accompanied by symptoms of severe neuroglycopenia. Despite of intensive pharmacological treatment recurrent hypoglycaemia episodes appeared for 34 hours in one case and 32 hours in the other. Authors discussed pathophysiological processes leading to hypoglycaemia in end stage renal failure patients. It seems that disturbances in renal gluconeogenesis together with lower degradation of insulin played the key role in creating hypoglycaemia in those two patients. PMID- 10697402 TI - [Human ehrlichiosis]. PMID- 10697403 TI - [CD5 and CD6 molecules--similarities and differences]. PMID- 10697404 TI - [Treatment of symptoms in the course of neutropenia]. PMID- 10697405 TI - [Drug treatment of heart failure--what's new after the era of angiotensin converting enzyme inhibitors]. PMID- 10697406 TI - ATP-dependent restriction enzymes. AB - The phenomenon of restriction and modification (R-M) was first observed in the course of studies on bacteriophages in the early 1950s. It was only in the 1960s that work of Arber and colleagues provided a molecular explanation for the host specificity. DNA restriction and modification enzymes are responsible for the host-specific barriers to interstrain and interspecies transfer of genetic information that have been observed in a variety of bacterial cell types. R-M systems comprise an endonuclease and a methyltransferase activity. They serve to protect bacterial cells against bacteriophage infection, because incoming foreign DNA is specifically cleaved by the restriction enzyme if it contains the recognition sequence of the endonuclease. The DNA is protected from cleavage by a specific methylation within the recognition sequence, which is introduced by the methyltransferase. Classic R-M systems are now divided into three types on the basis of enzyme complexity, cofactor requirements, and position of DNA cleavage, although new systems are being discovered that do not fit readily into this classification. This review concentrates on multisubunit, multifunctional ATP dependent restriction enzymes. A growing number of these enzymes are being subjected to biochemical and genetic studies that, when combined with ongoing structural analyses, promise to provide detailed models for mechanisms of DNA recognition and catalysis. It is now clear that DNA cleavage by these enzymes involves highly unusual modes of interaction between the enzymes and their substrates. These unique features of mechanism pose exciting questions and in addition have led to the suggestion that these enzymes may have biological functions beyond that of restriction and modification. The purpose of this review is to describe the exciting developments in our understanding of how the ATP dependent restriction enzymes recognize specific DNA sequences and cleave or modify DNA. PMID- 10697407 TI - DNA polymerase of the T4-related bacteriophages. AB - The DNA polymerase of bacteriophage T4, product of phage gene 43 (gp43), has served as a model replicative DNA polymerase in nucleic acids research for nearly 40 years. The base-selection (polymerase, or Pol) and editing (3'-exonuclease, or Exo) functions of this multifunctional protein, which have counterparts in the replicative polymerases of other organisms, are primary determinants of the high fidelity of DNA synthesis in phage DNA replication. T4 gp43 is considered to be a member of the "B family" of DNA-dependent DNA polymerases (those resembling eukaryotic Pol alpha) because it exhibits striking similarities in primary structure to these enzymes. It has been extensively analyzed at the genetic, physiological, and biochemical levels; however, relationships between the in vivo properties of this enzyme and its physical structure have not always been easy to explain due to a paucity of structural data on the intact molecule. However, gp43 from phage RB69, a phylogenetic relative of T4, was crystallized and its structure solved in a complex with single-stranded DNA occupying the Exo site, as well as in the unliganded form. Analyses with these crystals, and crystals of a T4 gp43 proteolytic fragment harboring the Exo function, are opening new avenues to interpret existing biological and biochemical data on the intact T4 enzyme and are revealing new aspects of the microanatomy of gp43 that can now be explored further for functional significance. We summarize our current understanding of gp43 structure and review the physiological roles of this protein as an essential DNA-binding component of the multiprotein T4 DNA replication complex and as a nucleotide-sequence-specific RNA-binding translational repressor that controls its own biosynthesis and activity in vivo. We also contrast the properties of the T4 DNA replication complex to the functionally analogous complexes of other organisms, particularly Escherichia coli, and point out some of the unanswered questions about gp43 and T4 DNA replication. PMID- 10697408 TI - The peripheral myelin protein 22 and epithelial membrane protein family. AB - The peripheral myelin protein 22 (PMP22) and the epithelial membrane proteins (EMP-1, -2, and -3) comprise a subfamily of small hydrophobic membrane proteins. The putative four-transmembrane domain structure as well as the genomic structure are highly conserved among family members. PMP22 and EMPs are expressed in many tissues, and functions in cell growth, differentiation, and apoptosis have been reported. EMP-1 is highly up-regulated during squamous differentiation and in certain tumors, and a role in tumorigenesis has been proposed. PMP22 is most highly expressed in peripheral nerves, where it is localized in the compact portion of myelin. It plays a crucial role in normal physiological and pathological processes in the peripheral nervous system. Progress in molecular genetics has revealed that genetic alterations in the PMP22 gene, including duplications, deletions, and point mutations, are responsible for several forms of hereditary peripheral neuropathies, including Charcot-Marie-Tooth disease type 1A (CMT1A), Dejerine-Sottas syndrome (DDS), and hereditary neuropathy with liability to pressure palsies (HNPP). The natural mouse mutants Trembler and Trembler-J contain a missense mutation in different hydrophobic domains of PMP22, resulting in demyelination and Schwann cell proliferation. Transgenic mice carrying many copies of the PMP22 gene and PMP22-null mice display a variety of defects in the initial steps of myelination and/or maintenance of myelination, whereas no pathological alterations are detected in other tissues normally expressing PMP22. Further characterization of the interactions of PMP22 and EMPs with other proteins as well as their regulation will provide additional insight into their normal physiological function and their roles in disease and possibly will result in the development of therapeutic tools. PMID- 10697409 TI - Translational frameshifting: implications for the mechanism of translational frame maintenance. AB - The ribosome rapidly translates the information in the nucleic sequence of mRNA into the amino acid sequence of proteins. As with any biological process, translation is not completely accurate; it must compromise the antagonistic demands of increased speed and greater accuracy. Yet, reading-frame errors are especially infrequent, occurring at least 10 times less frequently than other errors. How do ribosomes maintain the reading frame so faithfully? Geneticists have addressed this question by identifying suppressors that increase error frequency. Most familiar are the frameshift suppressor tRNAs, though other suppressors include mutant forms of rRNA, ribosomal proteins, or translation factors. Certain mRNA sequences can also program frameshifting by normal ribosomes. The models of suppression and programmed frameshifting describe apparently quite different mechanisms. Contemporary work has questioned the long accepted model for frameshift suppression by mutant tRNAs, and a unified explanation has been proposed for both phenomena. The Quadruplet Translocation Model proposes that suppressor tRNAs cause frameshifting by recognizing an expanded mRNA codon. The new data are inconsistent with this model for some tRNAs, implying the model may be invalid for all. A new model for frameshift suppression involves slippage caused by a weak, near-cognate codon.anticodon interaction. This strongly resembles the mechanism of +1 programmed frameshifting. This may mean that infrequent frameshift errors by normal ribosomes may result from two successive errors: misreading by a near-cognate tRNA, which causes a subsequent shift in reading frame. Ribosomes may avoid phenotypically serious frame errors by restricting apparently innocuous errors of sense. PMID- 10697410 TI - Syndromes associated with Homo sapiens pol II regulatory genes. AB - The molecular basis of human characteristics is an intriguing but an unresolved problem. Human characteristics cover a broad spectrum, from the obvious to the abstract. Obvious characteristics may include morphological features such as height, shape, and facial form. Abstract characteristics may be hidden in processes that are controlled by hormones and the human brain. In this review we examine exaggerated characteristics presented as syndromes. Specifically, we focus on human genes that encode transcription factors to examine morphological, immunological, and hormonal anomalies that result from deletion, insertion, or mutation of genes that regulate transcription by RNA polymerase II (the Pol II genes). A close analysis of abnormal phenotypes can give clues into how sequence variations in regulatory genes and changes in transcriptional control may give rise to characteristics defined as complex traits. PMID- 10697411 TI - Topoisomerase II as a target for anticancer drugs: when enzymes stop being nice. AB - Topoisomerase II is an essential enzyme that plays a role in virtually every cellular DNA process. This enzyme interconverts different topological forms of DNA by passing one nucleic acid segment through a transient double-stranded break generated in a second segment. By virtue of its double-stranded DNA passage reaction, topoisomerase II is able to regulate DNA over- and underwinding, and can resolve knots and tangles in the genetic material. Beyond the critical physiological functions of the eukaryotic enzyme, topoisomerase II is the target for some of the most successful anticancer drugs used to treat human malignancies. These agents are referred to as topoisomerase II poisons, because they transform the enzyme into a potent cellular toxin. Topoisomerase II poisons act by increasing the concentration of covalent enzyme-cleaved DNA complexes that normally are fleeting intermediates in the catalytic cycle of topoisomerase II. As a result of their action, these drugs generate high levels of enzyme-mediated breaks in the genetic material of treated cells and ultimately trigger cell death pathways. Topoisomerase II is also the target for a second category of drugs referred to as catalytic inhibitors. Compounds in this category prevent topoisomerase II from carrying out its required physiological functions. Drugs from both categories vary widely in their mechanisms of actions. This review focuses on topoisomerase II function and how drugs alter the catalytic cycle of this important enzyme. PMID- 10697412 TI - The biological properties and evolutionary dynamics of mammalian LINE-1 retrotransposons. AB - Mammalian LINE-1 (L1) elements belong to the superfamily of autonomously replicating retrotransposable elements that lack the long terminal repeated (LTR) sequences typical of retroviruses and retroviral-like retrotransposons. The non LTR superfamily is very ancient and L1-like elements are ubiquitous in nature, having been found in plants, fungi, invertebrates, and various vertebrate classes from fish to mammals. L1 elements have been replicating and evolving in mammals for at least the past 100 million years and now constitute 20% or more of some mammalian genomes. Therefore, L1 elements presumably have had a profound, perhaps defining, effect on the evolution, structure, and function of mammalian genomes. L1 elements contain regulatory signals and encode two proteins: one is an RNA binding protein and the second one presumably functions as an integrase replicase, because it has both endonuclease and reverse transcriptase activities. This work reviews the structure and biological properties of L1 elements, including their regulation, replication, evolution, and interaction with their mammalian hosts. Although each of these processes is incompletely understood, what is known indicates that they represent challenging and fascinating biological phenomena, the resolution of which will be essential for fully understanding the biology of mammals. PMID- 10697413 TI - Regulation of the mammalian alcohol dehydrogenase genes. AB - This review focuses on the regulation of the mammalian medium-chain alcohol dehydrogenase (ADH) genes. This family of genes encodes enzymes involved in the reversible oxidation of alcohols to aldehydes. Interest in these enzymes is increased because of their role in the metabolism of beverage alcohol as well as retinol, and their influence on the risk for alcoholism. There are six known classes ADH genes that evolved from a common ancestor. ADH genes differ in their patterns of expression: most are expressed in overlapping tissue-specific patterns, but class III ADH genes are expressed ubiquitously. All have proximal promoters with multiple cis-acting elements. These elements, and the transcription factors that can interact with them, are being defined. Subtle differences in sequence can affect affinity for these factors, and thereby influence the expression of the genes. This provides an interesting system in which to examine the evolution of tissue specificity. Among transcription factors that are important in multiple members of this gene family are the C/EBPs, Sp1,USF, and AP1, HNF-1, CTF/NF-1, glucocorticoid, and retinoic acid receptors, and several as-yet unidentified negative elements, are important in at least one of the genes. There is evidence that cis-acting elements located far from the proximal promoter are necessary for proper expression. Three of the genes have upstream AUGs in the 5' nontranslated regions of their mRNA, unusual for mammalian genes. The upstream AUGs have been shown to significantly affect expression of the human ADH5 gene. PMID- 10697414 TI - Transcriptional regulation by cyclic AMP-responsive factors. AB - In eukaryotes, transcriptional regulation on stimulation of the adenylate cyclase signaling pathway is mediated by a family of cyclic AMP-responsive nuclear factors, including CREB, CREM, and ATF-1. These factors contain the basic domain/leucine zipper motifs and bind as dimers to cAMP-responsive elements (CREs). The activation function of CRE-binding proteins is modulated by phosphorylation by several kinases and is mediated by coactivators such as CBP and p300. Activation might also be independent of CBP and phosphorylation in some specific cell types, such as male germ cells, wherein the protein ACT confers a powerful activation function to CREM. The inducible cAMP early repressor (ICER) protein is the only inducible member of this family. The induction of this powerful repressor is likely to be important for the transient nature of cAMP induced gene expression. CRE-binding proteins have been found to play an important role in the physiology of the pituitary gland, in regulating spermatogenesis, in the response to circadian rhythms, and in the molecular basis of memory. PMID- 10697415 TI - Hormonal disorders as a consequence of antineoplastic therapy in children. PMID- 10697416 TI - The gastrointestinal mast cell in health and disease. AB - Mast cells are metachromatic cells found widely throughout the body. In gastrointestinal tract they reside particularly in mucosa having close contact with external environment. Their certain role in health and disease remains unclear. Mast cells seem to be involved in lots of allergenic and non-allergenic inflammatory events taking place throughout the gastrointestinal tract including IgE-dependent hypersensitivity reaction, gastritis with or without Helicobacter pylori infection, Crohn's disease, ulcerative colitis, irritable bowel syndrome. Mast cell involvement in certain inflammatory processes in gastrointestinal tract were reviewed. PMID- 10697417 TI - Preparation of gastric mucin "STP-domains" using pronase digestion and chemical deglycosylation. AB - In this work we present a method of mucin protein backbone fragments preparation, which are rich in serine, threonine and proline amino acid residues. The purified native gastric mucin was reduced, the pronase digested and chemically deglycosylated. The obtained apomucin preparations contained slight quantities of residual carbohydrates (0-1%). In O-glycosylation reaction in vitro, with the participation of GalNAc-transferase, the amounts of incorporated [14C]GalNAc to apomucin fractions were about twofold greater in relation to the deglycosylated non-digested mucin subunits. We also demonstrate the usefulness of immunoblotting technique of apomucin preparations detection. PMID- 10697418 TI - The lichen flora of Suprasl as the health resort. AB - This study presents the actual lichen flora of Suprasl. 68 species have been collected in the investigated area. Among these, 53 are epiphytic species, 12 are epilithic species and 17--epiksylithic species. On the basis of lichens scale, Suprasl was classified to 6th bioindicative zone, where air pollution of SO2 was 30-40 mg/m3. PMID- 10697419 TI - An assessment of the structure of perception in people with bruxism. AB - The aim of the study was to assess the structure of perception in people with bruxism. 50 people with a diagnosis of bruxism (B), in whom primary evidence of oclusal disharmony and temporomandibular disorders were excluded, were included in the study. The control group (C) consisted of 50 people with no evidence of parafunctional abnormalities of the chewing mechanism. Shalit's diagram was used to assess the structure of perception. The primary indices: discriminability (D), articulation (A), affective load+ (AL+) and affective load- (AL-) and secondary indices: reduction (R), emotionality (E), intensity (I) were derived from Shalit's diagram. A significant reduction in the mean value of the primary indices D and AL+ was noted, as well as in the secondary indices E and I in people with bruxism, in comparison to the control group. Assessment of the structure of perception in people with bruxism through the analysis of the primary and secondary indices, makes it possible to predict behavior which has a negative effect on the individual's own health and provides the opportunity for changing such behavior to be more pro-health oriented. PMID- 10697420 TI - Morphological parameters of blood platelets before and after renal artery embolization in patients with renal cancer. AB - Embolization consists in the occlusion of renal artery and its ramifications, which induces acute renal infarction. Renal artery embolization, causing necrosis, stimulates cellular reactions within and around the neoplasm. The interaction of neoplastic cells with blood platelets is of great importance in the development of neoplastic disease. Neoplastic cells exert a stimulatory effect on blood platelets. During activation, platelets change in shape and secrete certain substances. The aim of the present study was to evaluate morphological parameters of blood platelets in renal cancer patients before and after renal artery embolization. 45 patients suffering from renal cancer were examined (22 women and 23 men) before and after embolization. Diagnosis was established basing on the patients' history, clinical picture, and ultrasonographic, urographic and angiographic examinations. Tumour advancement was defined as T2 and T3. The study has demonstrated that embolization leads to an increase in platelet count and in the percentage of "megathrombocytes". The changes observed in the study may indirectly testify to the involvement of the thrombopoietic system in the course of neoplastic disease. PMID- 10697421 TI - Hyphomycetes in the ice of water reservoirs. AB - The presence of 111 Hyphomycetes species was noted in the water obtained from melting ice from four water bodies. The following fungi were recorded for the first time from Poland: Acrodictys martinii, A. peruamazonensis, Beltraniella peruamazonica, Blodgettia borneti, Cylindrocarpon aequatoriale, Diplocladiella appendiculata, Clavariopsis azlanii, Fusariella candida, Fusticeps laevisporus, Helicoma vaccinii, Heliscus submersus, Helminthosporium bigenum, Isthmolongispora geniculata, Lateriramulosa uniinflata, Lemonniera centrosphaera, Menispora amazonensis, Microstella pluvioriens, Nectria flavo-lanata, Paraarthrocladium amazonense, Paradactylella peruviana, Phaeodactylium acutisporum, Phialogeniculata multiseptata, Pseudohansfordia dimorpha, Pseudospiropes subliferus, P. lotorus, Pyricularia peruamasonica, Scolecobasidium fusarioideum, Scolecosporiella amazonensis, Stemphyliomma tambopataense, S. terricola, Taeniolina deightonii, Triscelophorus magnificus and Veronaea botryosa. PMID- 10697422 TI - Activity of lysosomal proteases in the liver and in the plasma from rats intoxicated with methanol. AB - The activity of lysosomal proteolytic enzymes (cathepsin A, B, C, D and E) in cytosol and in the whole homogenate of the liver and in the blood plasma from rats intoxicated with 1.5, 3.0 and 6.0 g methanol/kg b.w. was measured 6, 12 and 24 h and 2, 5 and 7 days after the intoxication. The activity of all proteases was increased in the cytosol from 12 h to 5 days of intoxication, whereas the activity of these enzymes was decreased in the whole liver homogenate during the same time. The magnitude of the decrease in proteolytic activity in the whole homogenate of the liver depended on the amino acid active center of the enzyme. The greatest decrease was observed for sulfhydryl and hydroxyl proteases and smaller one for carboxyl proteases. The proteases activity in the plasma was increased from 12 h to 5 days after methanol intoxication. These results suggest that during methanol intoxication the cellular and lysosomal membranes are impaired and proteases are translocated into the blood. However, changes in proteases activities and proteases distribution within the hepatocytes may lead to disturbances in the catabolism of cell proteins and to destruction of liver cells. PMID- 10697423 TI - Morphological changes in the liver of rats intoxicated with methanol. AB - On the basis of morphological examinations in light and electron microscope, the evaluation of methanol influence on the liver of rats was conducted. The examination was carried out in the group of 36 rats that were given a single dose of methanol (1.5 g/kg b.w.) into the stomach through a gastric tube. The liver was taken from rats under the ether anaesthesia after 6, 12, and 24 hours as well as after 2, 5, and 7 days of methanol administration. Results showed that methanol intoxication caused visible changes in the examined organ. Only 6 h after intoxication, lobular peripheral hepatocytes presented characteristic features of vacuolar degradation persisting up to 48 h. Since the second day of intoxication, many cells with double nuclei were found more frequently than in controls. Single hepatocytes or small hepatocytic clusters with the features of deliquescent necrosis could be seen after 5 and 7 days of examination. All animals intoxicated with methanol showed distinct weakness of glycogen reaction. The loss of glycogen resources was highest at 24 h after methanol administration. The results indicate, that methanol causes morphological changes in the rat liver and that intensification of these changes depends on the time after intoxication. PMID- 10697424 TI - Coagulative and fibrinolytic activity in parietal thrombus of aortic aneurysm. AB - Lumen of aortic aneurysm is usually filled with parietal thrombus. Behaviour of the parietal thrombus is determined by the ratio of coagulation factors to factors of fibrinolytic system. Activity of some factors of coagulation and fibrinolysis in the parietal thrombus of aortic aneurysm was determined using coagulative, fibrinolytic and caseinolytic tests. Retracted, blood clot was a comparative material. Tissue factor activity in the parietal thrombus of the aneurysm was above threefold higher and antiheparin activity was nearly twice higher in comparison to the blood clot. Activity of plasminogen activators in the parietal thrombus was higher than in the blood clot. The parietal thrombus contained fourfold more of the plasminogen. Antiplasmin activity in the thrombus was above twofold lower than in the blood clot. High activity of the tissue factor and substances neutralizing heparin may intensify the thrombus growth. Yet the thrombotic tendency may be balanced by a high activity of plasminogen activators and plasminogen. PMID- 10697425 TI - Enzymuria in antibiotic therapy of acute infections. AB - Activities of N-acetyl-beta-D-glucosaminidase (NAG), its isoenzyme NAG-B, alanylaminopeptidase (AAP), elastase and trypsin inhibitor (alpha 1 PI) were evaluated as markers of nephrotoxicity and inflammation in acute infections treated with various antibiotics (vancomycin, netilmicin, pefloxacine, cefoperazone and imipenem). PMID- 10697426 TI - Late posttraumatic epilepsy in patients with an alcoholic problem treated surgically for posttraumatic chronic subdural hematomas. AB - The frequency and time of late posttraumatic epilepsy in 34 patients (6 women and 28 men) was evaluated on the basis of a three-year observation and performed examinations (CT of the head and EEG). These patients were treated surgically in the years 1989-1994 in the Department of Neurosurgery of the Medical Academy of Bialystok due to chronic posttraumatic subdural hematoma. On the basis of environmental and family interviews it was concluded that 41.1% of the patients operated systematically abuses alcohol. Within 14-30 minutes following the surgery, seizures of late posttraumatic epilepsy took place in 20.6% of patients. They took place over eight times more frequently in the case of patients with alcohol problems that in the ones without such problems. PMID- 10697427 TI - The concentration of alpha-1-antichymotrypsin in blood serum of women in labour. AB - Concentration of alpha-1-antichymotrypsin (A-1-ACT) in blood serum of parturient women was determined. The investigation was conducted in 33 women bearing eutrophic newborns and 36 women bearing hypotrophic newborns. The control group consisted of 30 healthy non-pregnant women in reproductive age. Concentrations of A-1-ACT were determined using the radial immunodiffusion method according to Mancini et al. Maximal concentration of A-1-ACT determined in group I was three times higher than minimal concentration. Maximal concentration of A-1-ACT determined in group II was two times higher than minimal concentration. In the control group, the difference between minimal and maximal concentrations of A-1 ACT was inconsiderable. The lack of statistically significant differences between these three groups suggests that labour stress does not influence serum concentrations of this inhibitor. The importance of A-1-ACT in the placental tissue may be connected with immunological mechanisms that assure development and maintenance of pregnancy. PMID- 10697428 TI - Effects of clobazam and vitamin E on the lipid peroxidation in the rat brain after electroconvulsive shock. AB - The effect of vitamin E and clobazam on lipid peroxides [LP] in the rat brain and the pattern of electroshock-induced seizures were assessed. Significant increase in the concentration of brain LP at the peak of seizures was found. Both vitamin E and clobazam reduced the levels of LP in the rat brain after electroshock. Clobazam combined with vitamin E inhibited markedly formation of LP in the rats with electroshock-induced seizures. Vitamin E augmented anticonvulsant effect of clobazam though itself it had not exhibited any anticonvulsant effect in this model of seizures. The action of two drugs combined resulted in reducing the intensity and the duration of seizures, and only minimal seizures were observed. In our opinion the obtained results possess some interesting clinical aspect They suggest that the combined treatment with clobazam and vitamin E of epileptic patient may decrease the intensity of epileptic seizures due to inhibition of LP formation. PMID- 10697429 TI - The influence of post-traumatic syndrome in determining capacity to return to work after closed head injury. AB - Neuropsychological deficits following closed head injury are responsible for a significant part of the post-traumatic syndrome. Nonetheless such deficits frequently elude standard neurological examination and head injured subjects are not recognised as suffering from any impairments, despite these having a significant effect on their ability to return to work. As a result, subjects are refused disability status, resulting in a large number of litigation appeals. The criteria for determining disability status after head injury are often inadequate, as they do not take into account the post-traumatic syndrome, and medical experts, if they are sensitive to these issues are left in a quandary as to how to justify their decisions before the courts. Equally, psychological assessment, where it is included, is frequently limited to the examination of intellectual functions and memory, which are not always grossly disturbed in these cases. It is recommended that neuropsychological assessments should form part of routine practice in the evaluation of outcome in closed head injury and the test instruments be of adequate sensitivity to measure the deficits occurring after head injury. However, it is necessary to bear in mind that these subjective symptoms, which in reality hamper subjects' ability to return to work, may be the result of disruption of fine cognitive functions, which are inaccessible to currently available test methods. PMID- 10697430 TI - The alterations of CD11A expression on peripheral blood lymphocytes/monocytes and CD62L expression on peripheral blood lymphocytes in Graves' disease and type 1 diabetes. AB - There is increasing evidence that the alterations of function and/or level of adhesion molecules play a key role in the pathogenesis of autoimmune diseases, such as Graves' disease or type 1 diabetes. The aim of the present study was to evaluate the expression of lymphocyte function-associated antigen 1 alpha (LFA-1 alpha, CD11a) and L-selectin (CD62L) molecules on peripheral mononuclear cells in Graves disease and type 1 diabetes in comparison to healthy controls, since they were shown to play an important role in lymphocytes and/or monocytes migration into the organs affected by immune process and are suggested to contribute to the pathogenesis of Graves disease and type 1 diabetes. The percentages of monocytes/lymphocytes expressing LFA-1 alpha antigen and lymphocytes expressing L selectin antigen and the fluorescence intensity of the studied molecules were measured by flow cytometry. At the onset of both autoimmune diseases the percentage of highly CD11a positive lymphocytes and the mean fluorescence intensity were statistically higher than in the healthy controls and patients with Graves' disease after thyreostatic therapy. The fluorescence intensity of LFA-1 alpha on monocytes was also increased in type 1 diabetic patients, but not in Graves' disease. The analysis of CD62L antigen expression on peripheral blood lymphocytes revealed decreased percentages of L-selectin positive cells in patients with Graves' disease (before and after treatment) and insulin-dependent diabetes in comparison to the controls. Our study suggests that the alterations of the expression of CD11a and/or CD62L molecules on peripheral blood lymphocytes could be the markers of ongoing autoimmune process in Graves disease and type 1 diabetes. PMID- 10697431 TI - Serum pro- and anti-inflammatory cytokines in patients with Graves' disease with ophthalmopathy during treatment with glucocorticoids. AB - The aim of the study was to estimate serum pro-inflammatory cytokines concentration: TNF alpha and IL-6 as well as anti-inflammatory cytokine levels: IL-10 and IL-1ra in patients with Graves' disease and ophthalmopathy before and during glucocorticoid therapy. Serum cytokines were detected in three groups of subjects: 20 patients with Graves' disease without ophthalmopathy (GD), 17 patients with clinical symptoms of ophthalmopathy (GO) (CAS > or = 3, anamnesis of GO > or = 1 yr) and 24 healthy volunteers. Glucocorticoid therapy consisted of intravenous infusions of methyloprednisolone (MP) (2 series, 3 grams each time) and subsequent treatment with oral prednisone (60 mg per day) in a tapering schedule. The serum samples were collected 24 hours before MP, 24 hours after MP, 12 +/- 2 days of treatment with prednisone and at the end of glucocorticoid therapy. The levels of TNF alpha, IL-6, IL-10 and IL-1ra in the serum were determined by the ELISA method. The statistical significance was estimated by the Mann-Whitney U-test. Serum IL-6 level was increased in patients with GO in comparison to the controls and did not change significantly after immunosuppressive treatment. We did not find any significant differences of serum TNF alpha between the studied groups and glucocorticoids did not change its level significantly. Serum IL-10 was elevated significantly both in patients with GD and GO in comparison to the control group. IL-10 levels increased significantly after glucocorticoids. IL-1ra level was significantly higher in patients with GO. In the GO group we found an increase of IL-1ra after MP treatment and its gradual decline during prednisone therapy. CONCLUSIONS: 1. The production of both: pro- and anti-inflammatory cytokines is increased in infiltrative Graves' ophthalmopathy 2. Efficient glucocorticoid therapy may be related to its influence on anti-inflammatory cytokines: IL-10 and IL-1ra. PMID- 10697432 TI - Effect of streptozotocin diabetes on fatty acid content and composition of the heart lipids in the rat. AB - The aim of the present study was to examine the effect of diabetes on long chain fatty acid content and composition in the heart lipids. The experiments were carried out on male Wistar rats. Diabetes was induced by intravenous administration of streptozotocin. Samples of the blood and the left ventricle were taken. Lipids were extracted and separated into different fractions. The following fractions were examined: free fatty acids, diacylglycerols, triacylglycerols and phospholipids. The fatty acids from each fraction were identified and quantified by means of gas-liquid chromatography. It was found that diabetes resulted in an almost four-fold elevation in the content of the free fatty acid fraction in the heart, whereas the plasma concentration of free fatty acids increased only two-fold. The diabetes induced changes in the content of particular acids in the fraction of free fatty acid in the heart did not reflect changes in their concentration in the plasma. The content of total di- and triacylglycerol fatty acids also markedly increased in diabetes. In both the compounds, the elevation in the content of individual acids, with the exception of myristic and palmitoleic acid reflected roughly the elevation in their concentration in the plasma. There were, however, several differences in the percentage composition of fatty acids between the two groups. In the fraction of phospholipids, the content of myristic, palmitic, stearic and linoleic acids remained stable, whilst the content of palmitoleic acid was reduced and the content of arachidonic acid was elevated. It is concluded that insulin deficiency results in marked changes in the endogenous lipid fatty acid content of the heart. These changes are not directly related to alterations in the supply of individual acids through the plasma. PMID- 10697433 TI - Flow cytometric examination of DNA ploidy, p53 and bcl-2 expression in primary squamous epithelial pulmonary carcinoma. AB - The aim of this study was answering the question whether cytofluorimetric determining of DNA ploidy, p53 and bcl-2 expression in tumour cells may enhance the diagnosis of squamous epithelial lung carcinoma. Samples of neoplastic tissue were taken from 11 operated patients. DNA staining was performed with propidium iodide. p53 and bcl-2 expression was evaluated by using monoclonal antibodies. By assessing of DNA ploidy we observed increased proliferative activity and, at the same time, high rate of the cell necrosis. In the analysis of p53 protein the presence of antigen was observed on average in 7.1 +/- 2.4, 15.1 +/- 3.9, and 19.8 +/- 4.1% of normal cells of the pulmonary tissue, the peripheral part of the tumour and cells of the central part of the tumour, respectively. bcl-2 was found on average in 7.3 +/- 2.2, 9.9 +/- 3.1, and 12.4 +/- 3.19% of the above cells, respectively. These results show that the technique of cytofluorimetric evaluation of p53 and bcl-2 proteins can be useful in diagnosis of pulmonary cancer in addition to classic histological and immunohistochemical methods. PMID- 10697434 TI - Prolidase as a prodrug converting enzyme. III. Synthesis of proline analogues of melphalan and theirs susceptibility to the action of prolidase. AB - The feasibility to targeting prolidase as an antineoplastic prodrug-converting enzyme has been examined. The synthesis of proline analogues of melphalan (well known antineoplastic agent) conjugated through imido-bond (potential target for prolidase action) has been performed. One of the compounds, N-[[[[(S) carboxy]pyrrolidin-1-yl]carbonyl]methyl]-4-[bis(2-chloro ethyl) amino]-2 phenylalanine, was found as very good prolidase substrate with susceptibility over 2 fold higher compared to standard, endogenous its substrate--Gly-L-Pro. It suggests that targeting of prolidase as a proline analogue of melphalan converting enzyme may serve as a novel strategy in therapy of neoplastic diseases. PMID- 10697435 TI - Synthetic analogues of netropsin and distamycin. V. Synthesis of a carbocyclic bis-lexitropsin as potential DNA cleaving agent. AB - A carbocyclic analogue of bis-distamycin, in which N-methylpyrroles were replaced by two trisubstituted benzene moieties joined by tetramethylenedioxy linkage, has been prepared. This compound bearing photosensitive benzotriazole moieties at the N-terminal residues. The synthetic method for the preparation of the bis benzotriazolyllexitropsin was accomplished in twelve steps starting from 2 hydroxy-5-phenylazobenzoic acid. PMID- 10697436 TI - Phagocytic activity of platelets in patients with myocardial infarction. AB - The aim of our present study was to analyse the dynamics of changes in the phagocytic activity of platelets with myocardial infarction according to the treatment applied. Twenty patients with acute myocardial infarction and thirty healthy subjects were examined. Platelet count, phagocytic activity of platelets and the phagocytic index were determined. We observed reduced phagocytic activity of platelets in the first 24 hours of infarction, which may be due to exhaustion of most active platelets in a thrombus and microaggregates. A transitory increase in the phagocytosis activity of platelets observed on the third day of infarction may be caused by the release of large, more active platelets from splenic pool. On the eight day of infarction, the phagocytic activity of platelets became normalized in both experimental groups. Our study indicates that together with platelet activation the phagocytic activity gets reduced being a likely response to the thrombocytopenic factor. This can be confirmed by the increased number of platelets on the 8th day. Our results suggest a slightly attenuated phagocytic ability of platelets, irrespective of the treatment applied. The reduction seems to result from the effect of platelets exhaustion on the procoagulative activity in patients with myocardial infarction. PMID- 10697437 TI - Analysis of recently activated, memory and naive lymphocyte T subsets in the peripheral blood of patients with Graves' disease and insulin-dependent diabetes mellitus. AB - It has been recently suggested that the expression of two different isofoms of tyrosine phosphatase antigen (CD45RA and CD45RO) could differentiate T cells into autoreactive and immunoregulatory subsets, which play a crucial role in the autoimmunity process. The aim of the present study was to evaluate the differences between the distribution of memory, naive and recently activated T cells (co-expressing both CD45RA and CD45RO antigens) in the peripheral blood of patients with newly diagnosed Graves' disease and insulin-dependent diabetes mellitus in comparison to healthy controls. The study was carried out in 3 groups: 18 patients with Graves' disease, 16 subjects with a recent onset of type 1 diabetes mellitus and 16 healthy, age and sex matched volunteers. At the onset of both autoimmune diseases the percentage of naive CD+ cells were lower than in the control group and in patients with Graves' disease treated with methimazole. The analysis of CD8+ lymphocyte subsets in the peripheral blood revealed higher levels of CD8+CD45RO+ cells in the newly diagnosed Graves' disease in comparison to the controls, and significant decline in the percentage of memory CD8+ and CD8+CD45RA+CD45RO+ lymphocytes after thyreostatic treatment. The number of CD4+ T lymphocytes co-expressing CD45RA and CD45RO antigens were statistically higher in the patients with recently diagnosed insulin-dependent diabetes mellitus. The abnormal distribution of naive, memory and "transient" T cell subsets in the peripheral blood at the onset of Graves' disease and diabetes type 1 suggest their role in the development of autoimmunity. The significant alterations of lymphocyte T subsets in the peripheral blood of patients with Grave's disease after thyreostatic therapy in comparison to the newly diagnosed subjects supports the immunomodulatory effect of methimazol treatment. PMID- 10697438 TI - Significance of some tumor markers in differential diagnosis of ovarian tumor. AB - The levels of CA-125, CA 72-4 and CEA were evaluated in serum of patients with various histological types and clinical stages of ovarian carcinoma and of benign ovarian tumor. An increased CA-125 was found in 8% patients with benign ovarian tumor. The antigens CA 72-4 and CEA exhibit values below the cut-off limit. CA 125 value was significantly elevated in patients with serous and CA 72-4 with ovarian mucinous carcinoma. An increase of CA-125 was found in 81% of patients with serous, 60% with endometrioid and 30% with ovarian mucinous carcinoma. An increase of CA 72-4 was found in 76% of patients with mucinous, 54% with serous and 45% with ovarian endometrioid carcinoma. Insignificant rise of CEA value was found in 8% of patients with serous, 10% with endometrioid and 15% with ovarian mucinous carcinoma. The frequency of increased concentration of antigens level showed a trend to significant increase and a correlation with the clinical stage of disease. The observations allow to conclude that the determination of CA 72-4 in the serum may be useful in differentiation diagnostics of benign and malignant ovarian tumors. The combined determination of CA-125 and CA 72-4 is useful in differentiation of histological types of ovarian tumors and their clinical stage. PMID- 10697439 TI - Aryltriazole C-nucleosides with affinity with certain DNA sequences. I. Synthesis of 2-(3-nitrophenyl)-4-(beta-D-erythrofuranosyl)-1,2,3-triazole. AB - The paper describes the synthesis of 2-(3-nitrophenyl)-4-(beta-D erythrofuranosyl)-1,2,3-triazole. This substance is a key substrate for synthesis of C-nucleosides, having a potential affinity with the G-C base pair. PMID- 10697440 TI - Plasmapheresis in the treatment of tropical malaria. AB - A case of tropical malaria imported from Kenya was described. Plasmodium falciparum (an etiological agent of this disease) is often drug resistant, therefore pharmacological treatment of the patients was supplemented with plasmapheresis. Full recovery has been achieved. It is suggested that plasmapheresis may be recommended for a complex therapy of severe forms of tropical malaria. PMID- 10697441 TI - Efficacy of interferon therapy on patients with HCV infection in dependence from the time of treatment. AB - Among 45 patients with chronic hepatitis C the efficiency of interferon alpha 2b treatment was evaluated. The efficiency of interferon therapy was determined after 6 months of treatment in the group of 28 patients (group I) treated for 6 months and 17 patients (group II) treated for 12 months. The side effects were investigated with respect to such a factor as time of treatment. 11 (39%) patients from group I had eliminated HCV RNA and 14 (50%) patients had normalized AlAT levels in the serum. Five (29%) patients from group II eliminated HCV RNA and 8 (47%) patients had normalized AlAT levels in the serum. The flu-like syndrome, thrombocytopenia, vision disorder, depression and somnolence were most often the side effects observed in treated patients. Proportionally to time of treatment vision disorder, depression and somnolence increased. PMID- 10697442 TI - [Asthma and chronic obstructive lung disease. Prevention and treatment]. PMID- 10697443 TI - [Allergen-specific immune therapy in the treatment of asthma]. AB - OBJECTIVES: 1. To identify all published randomised controlled trials of allergen specific immunotherapy in asthma. 2. To estimate the overall efficacy of allergen specific immunotherapy upon asthmatic symptoms, medication requirements, lung function, nonspecific bronchial hyperreactivity (BHR) and allergen specific BHR. SEARCH STRATEGY: A search of the asthma database by the Cochrane Airways Group at St. Georges Hospital Medical School, London identified 660 nonunique citations with the keywords Immunotherapy* or Hyposensitive or Desensiti*. This database included all studies published up to 1997 with the keywords Asthma or Wheez* from the Medline, Embase and Cinahl databases, together with other studies identified by handsearching. SELECTION CRITERIA: The review was restricted to randomised controlled trials (RCT). Only studies which focussed upon asthma were included. Allergen specific immunotherapy was defined as the subcutaneous administration of extracts of house dust mites, pollens, animal danders or moulds, chemically modified allergoids or antigen-antibody complexes. Although placebo controlled trials were methodologically stronger, studies which administered house dust or other relatively antigenically inactive preparations to the control group were also considered. Double blinded trials were preferred, but single blind and open studies were also reviewed for possible inclusion. At least one of the following clinical outcomes had to be reported: asthmatic symptoms, asthma medication requirements, lung function, nonspecific BHR or allergen specific BHR. Inclusion of studies in the review was decided by a simple majority of all three reviewers, who independently read the methods sections of papers identified by the search strategy and applied the stated criteria. Quality assessment was performed by 2 reviewers, who independently assessed the concealment of allocation. DATA COLLECTION AND ANALYSIS: The comparisons were: Allergen immunotherapy v placebo, Allergen immunotherapy v antigenically inactive control, House dust v placebo and Allergen immunotherapy v untreated control. These comparisons were performed separately for each outcome, whenever these results were reported. Outcome data were extracted and entered into RevMan 3.0.1 for statistical analysis. Categorical outcomes were analysed as odds ratios (OR) and 95% confidence intervals (95% CI) calculated by Peto's method. Continuous outcomes were analysed as standardised mean differences (SMD). Fixed effects models were used to obtain summary statistics for the overall efficacy of allergen immunotherapy and x2 tests were performed to assess heterogeneity between studies. MAIN RESULTS: Fifty four randomised controlled trials published between 1954 and 1997 satisfied the inclusion criteria. There were 25 studies reporting immunotherapy for mite allergy, 13 studies of pollen allergy, eight studies of animal dander allergy, two studies of allergy to the mould Cladosporium and six studies which attempted simultaneous immunotherapy for multiple aeroallergens. Concealment of allocation was assessed as clearly adequate in only 11 studies. The adequacy or otherwise of 40 studies could not be determined from the details published in the papers. Only three studies used a clearly inadequate method for concealment of allocation. There was a significant overall improvement in asthma symptom scores following immunotherapy (combined SMD -0.52; 95% -0.70 to -0.35). Patients randomised to immunotherapy were also significantly less likely to report a deterioration in asthma symptoms than those randomised to placebo (OR 0.27; 95% CI 0.21 to 0.35). Asthma medication requirements were significantly reduced (SMD -0.51; 95% CI 0.74 to -0.28). Patients randomized to immunotherapy were also significantly less likely to require medication than those randomised to placebo (OR 0.28; 95% CI 0.19 to 0.42). There was no overall improvement in lung function following immunotherapy and marked hete PMID- 10697444 TI - [Quality assurance of drug therapy for patients with asthma. Health economic analysis]. AB - The cost-effectiveness of a community pharmacy based programme for therapeutic outcomes monitoring of asthma patients' drug therapy is evaluated. Five hundred asthma patients, aged 16-60 and treated in primary care, with moderate to severe asthma, 31 community pharmacies and 139 general practitioners participated in the study. The total programme costs, costs of drugs, health care resource costs and indirect costs were evaluated together with the effects of the programme on: asthma symptoms status, days of sickness, quality of life, satisfaction with health care, peak-flow (PEF), inhalation technique and knowledge. The evaluation of the programme shows it to be cost-effective with cost-effectiveness ratios between 0.18 and 0.56. The pay off time for the programme is 23 months (range 9 64 months in the sensitivity analysis). It is concluded that the community pharmacist can contribute to identify and solve drug-related problems in a cost effective way with positive impact on asthma patients health, clinical and psycho social outcomes, even though the program is time consuming and intensive. PMID- 10697445 TI - [The course of pulmonary function in adults with asthma. The Osterbro study]. AB - We studied the course of forced expiratory volume in one second (FEV1) in adults with self-reported asthma using data from a longitudinal epidemiological study of the general population, The Copenhagen City Heart Study. The study was conducted over a period of 15 years with three measurements of lung function. The data base consisted of 17,506 men and women including 1.095 participants with asthma. The unadjusted FEV1 decline in subjects with asthma was 38 ml/year compared to 22 ml/year in nonasthmatics. Similarly, the statistical analysis showed that the FEV1 normalised by height (FEV1/height2) was significantly poorer in subjects with asthma compared to nonasthmatics (p < 0.001). Smoking contributed significantly to lung function decline regardless of asthma status (p < 0.001). In a sample of the general population, adults with self-reported asthma have a significantly faster decline of ventilatory function than nonasthmatics. PMID- 10697446 TI - [Occurrence of allergens on hospital premises]. AB - The occurrence of allergens from the house-dust mites Der p 1, Der f 1 and Der m, and from dogs (Can f 1) and cats (Fel d 1) was assessed in Viborg Hospital. Three hundred samples collected in a standardized manner were analysed for allergens by ELISA technique. In only one dust sample was the total occurrence of mite allergens marginally above the sensitization threshold level of 2,000 ng mite allergens/g dust. For Fel d 1 a threshold level for sensitization or symptoms of 8,000 ng Fel d 1/g dust has been proposed; none of the dust samples contained this concentration. A low occurrence of Can f 1 was found. One dust sample contained 8,902 ng Can f 1, while the remainder exhibited lower concentrations. Efficient cleaning and adequate ventilation can reduce allergens in public buildings, but it is impossible to remove all allergens from upholstered furniture. Avoidance of such furniture in wards and outpatient departments which receive allergic patients might be considered. PMID- 10697447 TI - [Long-term effect of inhaled budesonide in patients with mild to moderate chronic obstructive lung disease. The Osterbro Study]. AB - We compared the effect of inhaled budesonide with placebo on decline in lung function and respiratory symptoms in a three-year study of patients with chronic obstructive pulmonary disease (COPD). We used a parallel-group, randomized, double-blind, placebo-controlled design, nested in an ongoing epidemiological survey. Patients were non-asthmatic subjects with a decreased ratio between forced expiratory volume in one second (FEV1) and vital capacity (VC); i.e., FEV1/VC < or = 0.7. All included patients had an FEV1 which was irreversible to both inhaled terbutaline and prednisolone. Two hundred and ninety patients were randomized to receive either budesonide, 1200 mcg. daily for six months followed by 800 mcg. daily for 30 months, or placebo for 36 months. Patients had a mean age of 59 years and their mean FEV1 was 2.37 liters or 86% of predicted. Crude FEV1 declines were 41.8 ml/year in the placebo group and 45.1 ml/year in the budesonide group. Using a regression model in the intention-to-treat population, patients in the placebo group had an FEV1 decline of 49.1 ml/year in contrast to 46.0 ml/year in the budesonide group; the estimated difference 3.1 ml/year (95% confidence interval--12.8-19.0) was statistically insignificant, p = 0.70. No effect of inhaled budesonide was seen on respiratory symptoms or number of exacerbations. These findings question the role of longterm inhaled corticosteroids in the treatment of mild-moderate COPD. PMID- 10697448 TI - [Rehabilitation of patients with chronic obstructive lung disease. Are two exercise sessions per week sufficient?]. AB - Several studies of patients with chronic obstructive pulmonary disease (COPD) have shown that pulmonary rehabilitation three to seven times a week improves exercise performance and well being. This study investigates feasibility, effect and economic aspects of a programme consisting of two sessions a week. Twenty four patients were randomized to rehabilitation and twenty-one to placebo. In an outpatient setting patients were assigned to an eight-week programme of exercise plus education twice a week (Exercise group) or conventional community cares (Placebo group). Seven patients did not complete the rehabilitation. The characteristics of the thirty-eight COPD patients at baseline (mean +/- SD): forced expiratory volume in one second (FEV-1) 1.1 +/- 0.4 L, six-minute walk distance (6MWD) 413 +/- 75 m. Rehabilitation resulted in an insignificant improvement in well being and the 6MWD (29 m ?95% confidence interval: -8-66 m?. Rehabilitation session twice a week for eight weeks had no effect in patients with moderate COPD. PMID- 10697449 TI - [Intramedullary tuberculous abscess in a patient with liver transplantation]. AB - Recipients of solid-organ transplants are, due to immunosuppressive treatment, disposed to opportunistic infections including tuberculosis. We report a rare case of acute intramedullary tuberculous abscess in a Danish male recipient of a liver transplant. The problems in diagnostics, medical and surgical treatment are discussed. PMID- 10697450 TI - [Hypertrophic pulmonary osteoarthropathy. A paraneoplastic syndrome associated with lung cancer]. AB - The case report presents a 52-year-old woman. The patient was a smoker, but did not have any pulmonary symptoms. Four months before admission she developed pains in her knee, ankle, elbow and finger joints. Bone scintigraphy showed periosteal new bone formation along the femur and tibia and increased tracer uptake in the hands. These findings were diagnosed as HPOA. Chest X-ray revealed a lung tumour. PMID- 10697451 TI - [Asthma caused by methylene-diphenyl-diisocyanate cast in a nurse]. AB - A case of a 35 year-old female nurse without atopic disposition is presented. For one year (1990-1991), she worked in an emergency room, applying synthetic casts containing MDI 0-3 times daily. She developed rhinitis, itchy eyes and nightly wheezing, during employment in the emergency room, with subsequent serious asthma attacks in 1992 and 1996. Just before the last attack, the patient's husband had used insulation foam containing MDI. A specific bronchial provocation test was performed with MDI-based synthetic cast material. The patient developed an asthma attack after seven hours, with a 48% drop in FEV1, suggesting that MDI is the causative agent. PMID- 10697452 TI - [Plasmodium falciparum malaria]. PMID- 10697453 TI - [Plasma homocysteine as a risk factor for apoplexy]. PMID- 10697454 TI - [Vigabatrin and visual fields defects]. PMID- 10697455 TI - [Syncope--a challenge in general practice]. PMID- 10697456 TI - [Triple test screening is not harmless]. PMID- 10697457 TI - [Treatment of stable chronic obstructive lung disease with inhaled corticosteroid]. PMID- 10697458 TI - [Allergen-specific immunotherapy--vaccination against allergy]. PMID- 10697459 TI - [Adverse effects in short-term steroid therapy. Which adverse effects can be expected during daily administration of 20-40 mg prednisolone for a period 1-3 weeks?]. PMID- 10697461 TI - Chagas disease, Chile. PMID- 10697460 TI - [Glucocorticoids treatment of exacerbation of chronic obstructive lung diseases]. AB - Corticosteroids are widely used as first-line drugs in the treatment of patients with exacerbation of COPD. Until recently the scientific evidence has been scarce and generally taken from the treatment of asthma. In this article we interpret the results of the available prospective studies on corticosteroid treatment in exacerbation of COPD and suggest guidelines regarding the clinical handling of COPD patients in matter of dose, form of administration and duration of treatment. PMID- 10697462 TI - Thiomersal as a vaccine preservative. AB - Thiomersal poses a theoretical low risk of neurodevelopmental toxicity in infants. The known risk of morbidity and mortality from vaccine-preventable diseases and of contaminated multidose vaccine vials far outweigh any potential risk posed by thiomersal. However, with the weight of public opinion against the use of mercury of any sort, WHO and other agencies have begun the process of reducing and removing thiomersal from vaccines. In the short term (the next 3 years), modifications to existing strategies will result in a reduction in exposure to thiomersal. Over the long term (beyond 3 years), efforts will be focused on new vaccine-delivery technologies, alternative preservatives and combination vaccines, further reducing and eventually, perhaps, eliminating thiomersal from vaccines. PMID- 10697463 TI - Women's satisfaction with primary care: a new measurement effort from the PHS National Centers of Excellence in Women's Health. AB - This paper describes efforts by the National Centers of Excellence in Women's Health to develop a woman-specific primary care satisfaction instrument suitable for quality improvement and research. PMID- 10697464 TI - An HMO survey on mass customization of healthcare delivery for women. AB - A telephone survey of 1,000 randomly selected women members of Kaiser Permanente examined preferences for care delivery. The majority of women under age 55 (80%) were interested in scheduling evening or Saturday appointments, and half (50%) of them were willing to switch doctors for this option. Although most (57%) said that physician gender 'did not matter,' women who preferred to see a female physician but were seeing a male were significantly less satisfied than women whose preferences were matched. Half (51%) of women were open to receiving health education in group classes. Information on when care is preferred, by whom, and in what setting sets the stage for mass customization strategies. PMID- 10697465 TI - Chinese women: behaviors and attitudes toward breast cancer education and screening. AB - A study eliciting Chinese Women's Attitudes and behaviors toward breast cancer screening to identify and overcome barriers to providing access to health promotion information. PMID- 10697466 TI - Childhood sexual abuse and preterm labor in adulthood: an endocrinological hypothesis. AB - Anecdotal reports link adverse pregnancy outcomes, such as preterm delivery, to women with histories of childhood sexual abuse. Although little research has been conducted on this subject, we provide an overview of known health effects of violence against women and posit a biological explanation for adverse pregnancy outcomes among this population. Specifically, we hypothesize that early traumatic experiences of childhood sexual abuse may activate corticotropin releasing hormone (CRH) gene expression in the brain, and a vulnerability to elevated CRH gene expression in the placenta. Those traumatized by early abuse may be more susceptible to stress vis a vis CRH dysregulation during a major psychosocial stressor, such as pregnancy. Elevated CRH has been associated with preterm labor. PMID- 10697467 TI - The Commonwealth Fund 1998 Survey of Women's Health. PMID- 10697468 TI - Methodology and criteria in the evaluation of dental implants. AB - This study was designed to develop an inexpensive, reproducible method for studying the reaction of the tissues of the oral cavity to the endosseous implant. Thirty-six guinea pigs, three materials (Teflon, Vitallium, and Titanium 6Al-4V), two observation periods (two and 12 weeks), and two implant designs (one with exposure to the oral cavity and one without exposure to the oral cavity) were used. The inflammatory response was significantly greater in the exposed implants than in the unexposed implants. In the implants that were exposed 12 weeks, there was a strong interrelationship between severe inflammation, bacteria, and epithelial invagination. These factors are significant causes for failures of implants. PMID- 10697469 TI - Response of subcutaneous connective tissue to materials and drugs: a simplified technique. AB - The procedure that uses the response of subcutaneous connective tissue to screen dental materials and drugs will continue to be favored because of its simplicity, economy, reliability, and usefulness. The techniques of subcutaneous injection and of implantation of capsules by surgical means have many disadvantages. A modified technique is suggested; it has the ease of injection and the precision of implantation and it overcomes the difficulties inherent in both techniques. PMID- 10697470 TI - Corrosion of endodontic silver cones in humans: a scanning electron microscope and X-ray microprobe study. AB - Analyses with the scanning electron microscope and the X-ray microprobe were performed on 13 silver cones removed from 12 patients. The cones showed changes that ranged from surface dulling to black corrosion and pitting. Sulfur and chlorine were detected at the apical end of the cones and in the biopsy specimens of periapical tissues. PMID- 10697471 TI - The effect of serial preparation versus nonserial preparation on tissue removal in the root canals of extracted mandibular human molars. AB - Traditional methods of root canal preparation were compared with a serial type of preparation that included the use of rotary instruments. The serial type of preparation method proved more effective in debriding the root canals than did the traditional methods. PMID- 10697472 TI - The effect of preparation procedures on original canal shape and on apical foramen shape. AB - With the use of clear casting resin, simulated curved canals were created so that canal preparation procedures could be directly visualized and compared. Regardless of the type of enlarging instrument or the technique used, undesirable characteristics were produced in all preparations that would make canal filling difficult. To modify the typical preparations, alteration of the enlarging flutes, use of rasping rather than rotation of the instruments, and a flaring technique are recommended. PMID- 10697473 TI - Tricalcium phosphate as an adjunct to apical closure in pulpless permanent teeth. AB - Tricalcium phosphate was effective in inducing apical closure in human permanent pulpless teeth with flaring apices. However, it was not more effective than the calcium hydroxide that was used as a control. PMID- 10697474 TI - A comparative study of apical leakage with endodontic implant stabilizers. AB - The apical seals of 75 extracted teeth that were treated with three methods of endodontic implant stabilizers were compared with the seals of 25 teeth filled with silver cones and 25 teeth filled with laterally condensed gutta-percha. Apical leakage was measured with the use of methylene blue dye solution. Statistical analysis showed that there were significant differences between the apical leakage obtained from the teeth treated with endodontic implant stabilizers and those teeth filled with silver cones and with laterally condensed gutta-percha. PMID- 10697475 TI - The sporicidal activity of glass bead sterilizers. AB - Endodontic glass bead sterilizers were evaluated for sporicidal effectiveness. Small metal endodontic instruments contaminated with Bacillus subtilis spores were effectively sterilized when the instruments were exposed for 15 seconds at a temperature of 218 C. Nonmetal objects such as cotton pellets and paper points could not be sterilized because they charred and decomposed under the required temperature conditions. PMID- 10697476 TI - Povidone-iodine and isopropyl alcohol as disinfectants in preparation for endodontics. AB - The clinical effectiveness of povidone-iodine as a disinfectant in teeth isolated by a rubber dam was compared to the effectiveness of isopropyl alcohol. The results showed no statistically significant difference between povidone-iodine or isopropyl alcohol for disinfecting intact enamel surfaces. When a scrub-type application of povidone-iodine was used, preliminary removal of plaque with pumice did not increase its effectiveness. Both agents were significantly less effective at 15 minutes after application than they were at 90 seconds after application. PMID- 10697477 TI - The use of transparent teeth in the teaching of preclinical endodontics. AB - Demineralized teeth, which were cleared and hardened in xylene, were used in the teaching of preclinical endodontics. The teeth appeared well suited for the purpose. Of 39 students attending the course, 30 students thought that the use of transparent teeth had facilitated the learning of endodontic techniques. PMID- 10697478 TI - Scanning electron microscopy of cells from periapical lesions. AB - Examination of lymphocytes from peripheral blood with the scanning electron microscope (SEM) has shown differences between B cells and T cells on the basis of their surface architecture. This study was initiated to determine whether the cellular components of periapical lesions could be identified with the use of similar criteria. Cells were dispersed from lesions by aspiration of fragments of tissue through syringe needles of decreasing diameters. The liberated cells were filtered on silver-coated Flotronic membranes and examined under the SEM. Lymphocytes, macrophages, epithelial cells, and mast cells were observed in granulomas and cysts. Most of the lymphocytes had smooth surfaces similar to that of T cells; others had villous projections similar to that of B cells. Epithelial nests were seen in the cyst linings while the cyst fluid was rich in lymphocytes. These findings suggest that SEM examination of periapical lesions can be a useful adjunct in studying cellular composition and possible immunological reactions in these tissues. PMID- 10697479 TI - Blood-lead levels of monkeys treated with a lead-containing (N2) root canal cement: a preliminary report. AB - The effect of a root canal filling material on the level of lead in blood of rhesus monkeys was examined by anodic stripping voltometry. Blood-lead levels after root canal treatment with N2 cement were elevated when compared to preoperative controls. Lead 210 was incorporated into the leadfree N2 cement to identify the source of lead. Analyses of blood samples for 210Pb indicated that the lead originated from the filling material. PMID- 10697480 TI - Postdebridement retention of endodontic reagents: a quantitative measurement with radioactive isotope. AB - Forty freshly extracted human teeth were biomechanically prepared for endodontic obturation using a radioisotopically labeled EDTA-urea peroxide-Carbowax compound. The amount of radiolabeled mixture remaining in the canals after through cleansing was about 3.8% of that originally applied. The amount of residue did not decrease with reinstrumentation and irrigation. PMID- 10697481 TI - Long-distance bactericidal and fungicidal effectiveness of parachlorophenol and Formalin on Streptococcus faecalis and Candida albicans. AB - The "long-distance" bactericidal and fungicidal effectiveness of several concentrations of parachlorophenol, 25% Formalin, and 2% glutaraldehyde, all in 80% alcohol solution, has been tested in vitro on Streptococcus faecalis and Candida albicans. A concentration of at least 10% parachlorophenol in 80% alcoholic solution has the same long-distance bactericidal and fungicidal effectiveness as 25% Formalin in 80% alcoholic solution. PMID- 10697482 TI - Formation of mineralized scar tissue induced by implants containing collagen calcium phosphate gel. AB - The ability of a collagen-calcium phosphate gel to induce physiologic closure of subcutaneous polyethylene tube implants was tested. The openings of several tubes that had been filled with the gel were occluded by scars of mineralized connective tissue. Differentiation of cells into palisading fibroblasts and elaboration of a linear collagen matrix at the tissue-gel interface was seen. PMID- 10697483 TI - A combined post, core, and endodontic endosseous implant. Case report. AB - A hyperextended chrome-cobalt post and core also used as an endodontic endosseous implant served to stabilize a short mobile root and to provide for placement of a porcelain veneer metal crown. The technique is not complicated and does not require instruments other than those for routine endodontic treatment and for the construction of the post and core. PMID- 10697484 TI - Workshop of Advanced Endodontic Programs. Group 1. A. Recruiting, selecting, and evaluating faculty. PMID- 10697485 TI - The role of research in advanced endodontic programs. PMID- 10697486 TI - Corrosion of silver cones in bone: a scanning electron microscope and microprobe analysis. AB - Sections of silver cones were implanted in tibial bone wounds in 36 Sprague Dawley rats. Groups of six rats each were killed at monthly intervals, and the specimens were examined by scanning electron microscopy and by X-ray microanalysis. The silver cones had corroded rapidly, but they were well tolerated by the tissues. Elements of silver, chlorine, and sulfur were found in the tissues adjacent to the implants. PMID- 10697487 TI - Frequency, location, and direction of the lateral, secondary, and accessory canals. AB - Observation of 1,140 transparent teeth of adult humans was made to verify the frequency, location, and direction of the accessory, secondary, and lateral canals located at the radicular-apical area, at the body of the root, and in the base of the root. In 27.4% of the teeth studied, some type of ramification was observed; these ramifications were usually located in the apical area of the root. The premolars and molars showed the greatest variety of ramifications. PMID- 10697488 TI - Clinical, radiographic, and histological study of endodontically treated retained roots to preserve alveolar bone. AB - Endodontically treated, submerged roots in two Macaca speciosa monkeys were studied clinically, radiographically, and histologically. The roots were successfully covered by soft tissue, except in two areas. In several sites, radiographic evidence of bone formation was observed; this was confirmed by histologic examination. Bone formation coronal to the submerged roots was not a predictable occurrence. Even though epithelium and inflammation commonly occurred over the amputation sites, their presence did not seem to affect bone formation. PMID- 10697489 TI - Antibiotic sensitivities of enterococci isolated from treated root canals. AB - Enterococci that persisted in debrided, medicated root canals were tested by the Kirby-Bauer procedure for sensitivity to various antibiotics. The 50 strains tested were uniformly sensitive to ampicillin and vancomycin. More than 90% were also sensitive to erythromycin. Varying degrees of sensitivity and resistance were noted to bacitracin, cephaloridine, cephalothin, chloramphenicol, gentamicin, and tetracycline. All organisms were either partly or wholly resistant to clindamycin; penicillin; streptomycin; and sulfadiazine, sulfamerazine, and sulfamethazine (triple sulfas). PMID- 10697490 TI - American Society for Neurochemistry 31st annual meeting. Chicago, Illinois, USA. March 26-29, 2000. Abstracts. PMID- 10697491 TI - American College of Cardiology 49th annual scientific session. Anaheim, California, USA. March 12-15, 2000. Abstracts. PMID- 10697492 TI - Cumulative author, subject and title index, volumes 100-138. PMID- 10697493 TI - Overexpression of p21Cip1 or p27Kip1 in the promyelocytic leukemia cell line HL60 accelerates its lineage-specific differentiation. AB - The process of terminal differentiation is associated with exit from the cell cycle and loss of the proliferative potential of cells. The cyclin-dependent kinase inhibitors (CDIs) play critical roles in check-point functions during the cell cycle and as inhibitors of cell proliferation. Loss of their activities can impair development and differentiation and contribute to the uncontrolled proliferation characteristic of cancer cells. When the promyelocytic leukemia cell line HL60 is induced to differentiate in vitro, by a variety of agents, cellular levels of the CD1 proteins p21Cip1 and p27Kip1 are increased. To further address the roles of these two proteins in differentiation, we have overexpressed either a human p21Cip1 or p27Kip1 construct in HL60 cells. The overexpression of p21Cip1 accelerated both the monocytic and granulocytic differentiation of HL60 cells triggered by TPA or DMSO, respectively. The accelerated and more dramatic induction of differentiation seen in the p21Cip1 overexpressors was associated with a more rapid reduction of CDK2 kinase-associated activity, increased levels and more rapid dephosphorylation of the Rb protein, and increased levels of the cyclin D3 protein. Stable overexpression of p27Kip1 also enhanced TPA-induced differentiation of HL60 cells. These studies provide direct evidence that the increased expression of p21Cip1 and p27Kip1 play a causal role in the process of terminal differentiation of HL60 cells. Therefore, agents that enhance the expression of one or both of these proteins might be useful in therapy by enhancing the terminal differentiation of leukemia cells. PMID- 10697494 TI - Antisense human neuroglia related cell adhesion molecule hNr-CAM, reduces the tumorigenic properties of human glioblastoma cells. AB - BACKGROUND: Human Nr-CAM (Neuroglia related Cell Adhesion Molecule) is over expressed in glioblastoma multiforme tissue (GMT) as compared to normal brain tissue (NBT). MATERIALS AND METHODS: We transfected a human glioblastoma cell line (2020-CRL) with a vector that overexpresses antisense hNr-CAM using a CMVpromoter. RESULTS: Antisense hNr- CAM caused reduction in the native hNr-CAM expression, changed cell morphology, reduced the cell proliferation rate and lengthening of the cell cycle. Furthermore, antisense hNr-CAM overexpression in these cells caused extensive reduction in the number of soft agar colonies and invasion through extra cellular matrix (ECM) gel in vitro. Subcutaneous injection of antisense hNr-CAM overexpressing glioblastoma cells into nude mice caused complete inhibition of tumor formation as compared to vector only transfected cells. Intra-tumoral inoculation of antisense hNr-CAM expressing plasmid also caused slow tumor growth in nude mice in vivo. CONCLUSION: On the basis of these results, we conclude that hNr-CAM is a valid target for potential gene therapy of glioblastoma tumors. PMID- 10697495 TI - In vivo nuclear uptake of a vitamin D analog (OCT) in different tumor cell populations of FA-6 cancer xenograft in nude mice by receptor autoradiography. AB - 1 alpha, 25-dihydroxyvitamin D3 [1 alpha, 25(OH)2D3] and its analogs have been shown to repress the production of parathyroid hormone-related peptide (PTHrP) in tumors, which is a major factor causing humoral hypercalcemia associated with various cancers. Since vitamin D analogs may be applicable to the treatment of cancer patients, the present study was undertaken to examine whether OCT, an analog with little calcemic activity, is incorporated into tumor tissues, and to identify cellular and subcellular sites of its specific uptake and retention. [26 3H]OCT was injected i.v. into nude mice inoculated with a human pancreatic carcinoma cell line (FA-6). At 1 hour after the injection, intracellular concentration of radioactivity was visualized by receptor (thaw-mount) autoradiography. The results indicate a heterogeneous distribution of radioactivity in nuclei of certain large cancer cells as well as in single or clustered small and elongated cells within the tumor, while connective tissue cells outside of the tumor remained free of nuclear labeling. The data suggest that OCT acts selectively at the genome of cancer cells during a certain maturational stage and also of a second population of small fibroblast-like cells that may have been transplanted with the tumor or are host-derived. PMID- 10697496 TI - Mitochondrial DNA influences radiation sensitivity and induction of apoptosis in human fibroblasts. AB - BACKGROUND: We aimed to assess the role of mitochondrial DNA (mtDNA) in ionizing irradiation. MATERIAL AND METHODS: We examined three human fibroblast cell lines either lacking mtDNA (rho 0 cell), carrying mutant mtDNA (syn cell) or normal mtDNA (rho + cell). Cell survival curves were generated with colony formation and dye exclusion tests. Cell cycle analysis and apoptosis assay were performed by flow cytometry. RESULTS: The rho 0 cell line showed a stronger resistance to irradiation than the other two cell lines when assessed by the colony formation assay. In the dye exclusion test, the rho 0 cells revealed a superior cell viability among the cell lines after 5 and 8 Gy irradiation. The rho 0 cells underwent apoptosis at lower rates than the other two cell lines after 2, 5 and 8 Gy irradiation, and showed no alterations in the cell cycle distribution among them. CONCLUSION: mtDNA plays an important role in radiation sensitivity and induction of apoptosis. PMID- 10697497 TI - The detection of microscopically disseminated cancer cells in the abdominal cavity by intraoperative lavage cytology combined with an immunocytochemical method in gastric cancer. AB - This study was conducted to clarify the possible role of the immunocytochemical examination of intraoperative lavage cytology in gastric cancer. The expression of CA19-9, STN, SLX and CEA in tissues were examined in 70 patients with advanced gastric cancer who underwent gastric resection. The tissue sections were processed with the hematoxylin and eosin staining and immunostaining using the avidin-biotin-peroxidase complex (ABC) method. Fifty one patients underwent the lavage cytology. The cytologic samples were stained by the conventional Papanicolau method and ABC immunocytochemical method. Expression of CEA was detected at obviously higher frequency than those of the 3 carbohydrate antigens. The method combined with 4 antibodies increased the detection rate to 97.2%. Conventional lavage cytology was positive in 16 out of 51 patients. The diagnosis of class III in four patients was changed to class V through the immunocytochemical examination. The immunocytochemical examination of lavage cytology is very useful to verify the microscopically disseminated cancer cells in gastric cancer. PMID- 10697498 TI - Correlation of TGF beta 1 overexpression with down-regulation of proliferation inducing molecules in HPV-11 transformed human tissue xenografts. AB - The epidemiologic association of human papillomavirus (HPV) infection with dysplasia and cervical cancer is well established. Transforming growth factor beta 1 (TGF beta 1) has regulatory effects on a broad spectrum of cell types and is a growth inhibitory protein for epithelial cells. To examine the phenotype of experimentally generated, HPV-11 transformed human tissues, we looked at expression of TGF beta 1 and a number of proliferation-enhancing molecules which are known to be regulated by TGF beta 1, including bcl-2, c-myc, c-Ha-ras, c-jun and NFkB. HPV-11 transformed xenografts showed up-regulation of TGF beta 1 expression and down-regulation of the expression levels of bcl-2, c-myc, c-Ha ras, c-jun and NFkB. These results suggest that TGF beta 1 may exert antiproliferative effects on HPV-11 transformed papillomas by down-regulating different proliferation-enhancing molecules. PMID- 10697499 TI - Transforming growth factor beta 1 (TGF beta 1) down-regulates expression and function of proliferation-inducing molecules in HPV-transformed cells. AB - The epidemiologic association of human papillomavirus (HPV) infection with dysplasia and cervical cancer is well established. Transforming growth factor beta 1 (TGF beta 1) is a growth inhibitory protein for epithelial cells. To examine the phenotype of HPV-transformed cells, we examined expression of TGF beta 1 and a number of cellular proliferation-enhancing molecules which are known to be regulated by TGF beta 1, including bcl-2, c-jun and NFkB. Previous studies had identified significant induction of TGF beta 1 and concomitant down regulation of other growth stimulatory molecules in experimental papillomas. We used HPV-16 and -18 transformed cell lines. The HPV-16 transformed cells showed down-regulation of bcl-2 and NFkB as well as NFkB function upon TGF beta 1 treatment. The results suggest that TGF beta 1 may exert antiproliferative effects on some HPV-transformed cells by down-regulating expression and function of different proliferation-enhancing molecules. It is uncertain if this function is virus type specific and/or related to state of tumor cell progression. PMID- 10697500 TI - Amplified increase in signal transduction activity in cancer cells. AB - AIM: To elucidate the behavior of 1 phosphatidylinositol 4,5 bisphosphate (PIP2) 5 phosphatase (PIP2 5-Pase, EC 3.1.3.36) and 1 phosphatidylinositol 4 phosphate (PIP) 4-phosphatase (PIP 4-Pase) in cancer cells, the steady state activities of PIP2 5-Pase and PIP 4-Pase were determined in the particulate fractions of rat liver and in a spectrum of rat hepatomas of different growth rates and malignancy. METHODS: A standard method was developed using exogenous PIP2 or PIP as substrates. RESULTS AND DISCUSSION: One half of the maximum activities was reached at about 0.3 mM of the substrates and at 0.1 to 0.2 mM magnesium chloride. The optimum concentrations of Triton X-100 and cetyltrimethyl ammonium bromide were 0.25% (w/v) and 1 mM, respectively. PIP2 5-Pase and PIP 4-Pase activities were linear with time for 20 min and proportional with protein concentrations up to 22 micrograms per 50 microliters reaction mixture. In rat liver the steady state activities of PIP2 5-Pase and PIP 4-Pase were 1849 to 1881 and 1394 to 1670 nmol/h/mg protein. In three rat hepatomas the enzyme activities decreased to 50-51% and 33-42%, respectively. These results and our earlier data showing increased 1 phosphatidylinositol 4 kinase (EC 2.7.1.67), PIP 5-kinase (EC 2.7.1.68) and phospholipase C, PIP2 phosphodiesterase (EC 3.1.4.11) activities provide evidence of a transformation linked amplified increased capacity in signal transduction activity in cancer cells. The discovery and documentation of this integrated imbalance in regulation in phosphoinositide metabolism in cancer cells are in line with our observations in cancer cells for enzymes of purine and pyrimidine metabolism. The amplified increase in signal transduction capacity in cancer cells provides novel targets for the development of anticancer drugs. PMID- 10697501 TI - Potential use of radiolabeled antisense oligonucleotides in oncology. AB - Antisense oligodeoxynucleotides (ODNs) have great promise for therapy. Oligomers labeled with gamma-emitting radioelements have great promise for diagnosis. These radiolabeled oligomers can be used to identify the presence of a particular messenger RNA through simple external detection of radioactivity by means of scintigraphy. This review evaluates the progress in the development of antisense oligodeoxynucleotides for non-invasive imaging of chemoresistance. As nominated by H.N. Wagner at the final session of Nuclear Medicine congress in Denver, the deoxyribonucleic acid is "the molecule of the millennium". PMID- 10697502 TI - Chemoprevention of carcinogen-induced mammary tumorigenesis by the hybrid polar cytodifferentiation agent, suberanilohydroxamic acid (SAHA). AB - Hybrid Polar Cytodifferentiation (HPC) agents represent a novel class of anticancer compounds which act by inducing terminal differentiation and/or apoptosis rather than by cytotoxic action. Among these are HPC agents such as hexamethylenebisacetamide (HMBA) and more potent 2nd generation hybrid/polar compounds such as suberanilohydroxamic acid (SAHA). As of the present, most studies on HPC agents have focused on cancers of the hematopoietic system rather than solid epithelial tumors. The objective of the present study therefore was to assess the chemopreventive action of these two related compounds in the N methylnitrosourea (NMU)-induced rat mammary tumor model. Female Sprague-Dawley rats were fed diets containing 450 and 900 ppm, SAHA and 1000 and 2000 ppm HMBA, starting one week prior to NMU administration and continued for a period of 18 weeks. Mammary tumor development was monitored by palpation throughout the study, and at termination tumor incidence, number, multiplicity, latency and volume were determined. Weight gain was measured biweekly throughout the study. The salient results were as follows: SAHA at 900 ppm reduced NMU-induced mammary tumor incidence by 40%, total tumors by 66%, mean tumor multiplicity by 43% and mean tumor volume by 78%, with no detectable toxic side effects. HMBA exerted no tumor inhibiting effects at either concentration. This study represents the first demonstration that an HPC agent, namely SAHA, can inhibit the development of a chemically-induced, solid, epithelial tumor, at a relatively low dose (approximately 13 mgs/rat/day) without untoward side effects. PMID- 10697503 TI - GM1 inhibits early signaling events mediated by PDGF receptor in cultured human glioma cells. AB - Binding of platelet-derived growth factor receptor (PDGF) to its receptor (PDGFR) activates its receptor tyrosine kinase which autophosphorylates tyrosine residues. The p85 regulatory subunit of phosphatidylinositol 3-kinase (PI 3 kinase) binds to specific phosphotyrosines on PDGFR-beta and through the associated p110 catalytic subunit of PI 3-kinase catalyzes the formation of lipids that are involved in intracellular signaling. We examined if GM1 affects interactions between PDGFR-beta and specific proteins involved in PDGFR-mediated signaling. U-1242 MG cells were studied under different growth conditions using immunoprecipitation and Western Blot analysis. PDGF-stimulated the association of PDGFR-beta with p85, ras GTPase-activating protein and PLC gamma. GM1 decreased these associations in parallel with decreased tyrosine phosphorylation of PDGFR. PDGF augmented the activity of PI 3-kinase associated with PDGFR-beta, and this was attenuated by GM1. However, GM1 did not alter SH2 domains of p85. GM1 probably inhibits PDGF-induced signaling proteins with PDGFR-beta by inhibiting phosphorylation of specific tyrosines on the receptor which bind to SH2-domains on signaling proteins. PMID- 10697504 TI - Cytosolic p53 protein and serum p53 autoantibody evaluation in breast cancer. Comparison with prognostic factors. AB - Mutations of the TP53 gene induce the production of an abnormal protein (p53) with a prolonged half-life allowing its detection by monoclonal antibodies and leading to a development of serum p53 autoantibodies. We have quantified the protein p53 in 196 cytosols of primary breast cancer tissues, by an immunoluminometric method and searched for p53 autoantibodies in the sera of 101 patients, by an immunoenzymatic assay. The median value has been chosen as cut off (0.26 ng/mg protein). 18.4% of tumors had a p53 level < 0.10 ng/mg protein, and 8.2% had a p53 level > or = 2.5 ng/mg protein (range: 0.43.37). We found a significant difference between p53 distribution (p = 0.003) and median level (p = 0.001) in ductal and lobular carcinomas. The p53 median levels were significantly different between the grade III versus the grade I tumors (p = 0.01) and versus the grade II tumors (p = 0.008). An inverse correlation was obtained between p53 levels and ER (p = 0.003) alone or with PR (p = 0.006). p53 autoantibodies were detected in 7.9% of cases (8/101). This p53 study shows correlations with some poor prognostic factors, but would require further evaluation to better define the patient groups with poor prognosis. The p53 detection autoantibodies is neither correlated with the p53 cytosolic assay nor with prognostic factors of breast cancer, and should therefore not be used for patient the selection of the patient groups with poor prognosis. PMID- 10697505 TI - Inhibition of tumor growth by L-deprenyl involves neural-immune interactions in rats with spontaneously developing mammary tumors. AB - L-deprenyl, a monoamine oxidase-B inhibitor, has been shown to reverse the age related decline in sympathetic noradrenergic innervation and immune function in old rats and enhance T cell and NK cell activity in tumor-bearing rats. The objective of the present study was to examine whether deprenyl treatment of old female rats with mammary tumors could augment sympathetic nervous system and immune responses to inhibit the tumor growth. Female Sprague-Dawley rats with spontaneous mammary tumors were administered 0, 2.5 mg, or 5.0 mg/kg body weight (BW)/day deprenyl for i.p. 9 weeks. Tumor diameter, tumor number and body weight were measured throughout the treatment period. At the end of the treatment period, norepinephrine (NE) concentration, interferon-gamma production (IFN gamma), Con A-induced T lymphocyte proliferation, and percentage of T and B lymphocytes and natural killer cells were measured in the spleen, and the concentrations of monoamines were measured in the medial basal hypothalamus. Relative to saline-treated controls, treatment with deprenyl reduced tumor growth, increased NE concentration, IFN-gamma production and percentage of the CD8+ T lymphocytes in the spleen. In the medial basal hypothalamus, deprenyl treatment increased the concentrations of catecholamines and indoleamine. These results suggest that the anti-tumor effects of deprenyl on spontaneous rat mammary tumors may be achieved via neural-immune signaling in the spleen and medial basal hypothalamus. PMID- 10697506 TI - Antitumor activity and modified immunoregulation associated with IFN-gamma treatment of RG2 gliomas. AB - BACKGROUND: Interferon-gamma (IFN-gamma) is a key cytokine that upregulates molecules that participate in the processing and presentation of antigen, and effectively induces various immune regulatory factors. IFN-gamma also has cytotoxic or antitumor activities against some tumors in humans as well as animals. In this study, we evaluated the antitumor activity of recombinant rat IFN-gamma (rrIFN-gamma) on RG2 gliomas, and its immunological effect on tumor sites. MATERIALS AND METHODS: Rats with RG2 gliomas were intracarotidly treated by rrIFN-gamma with or without RMP-7, which has been reported to selectively increase the transport of cytokines to tumor tissue. To evaluate its immunological effect on tumor sites, an immunohistochemical study was performed using monoclonal antibodies against MHC class-1, CD8 and ED2 antigens after rrIFN gamma treatment. RESULTS: Intracarotid treatment with high rrIFN-gamma (2.4 x 10(5) U/kg) significantly increased survival (p < 0.02), however, the combined use of RMP-7 and rrIFN-gamma did not result in a further increase. Although the immunohistochemical study showed no clear increase in the staining of MHC class-1 or CD8 antigens on tumors following rrIFN-gamma treatment, immunostaining for ED2 antigen, which is known to be expressed on perivascular cells with MHC class-2 antigen, revealed a clear increase in the number of infiltrating positive cells within the tumors. CONCLUSION: Intracarotid rrIFN-gamma treatment can increase survival in rat glioma models through a mechanism in which antigen presentation is enhanced, and such effects are not always further increased by combination with RMP-7. PMID- 10697507 TI - Expression of the MAGE 3 gene product in squamous cell carcinomas of the head and neck. AB - BACKGROUND: The melanoma antigen (MAGE) 3 gene may be a useful tumor specific marker since it is expressed in a variety of cancers. MATERIALS & METHOD: The expression and intracellular location of MAGE 3 gene product were investigated in 40 squamous cell carcinomas, 2 tumor lines, 20 benign diseases, and 20 normal tissues of the head and neck. Immunohistochemical staining with anti-MAGE 3 mAb 57B was conducted from fresh frozen specimens. Correlations between MAGE 3 expression and clinicopathological parameters were also evaluated. RESULTS: The MAGE 3 gene product was detected in squamous cell carcinomas (18/40, 45%) and in tumor cell lines (2/2, 100%), but not in benign diseases and normal tissues. No significant correlation was drawn between MAGE 3 expression and clinical parameters including clinical stages and metastasis. CONCLUSION: These results show MAGE 3 antigen could represent a potential target for immunotherapy in head and neck squamous cell carcinomas. PMID- 10697508 TI - Expression of the multidrug resistance protein (MRP1) in breast cancer. AB - BACKGROUND: The multidrug resistance protein (MRP1) is expressed in human breast carcinomas but its clinical significance remains unclear. The aim of the present study was to determine the clinical significance of MRP1 in breast cancer patients. MATERIALS AND METHODS: MRP1 expression of primary carcinomas from 100 breast cancer patients was immunohistochemically determined by means of the monoclonal antibodies QCRL-1/QCRL-3. RESULTS: MRP1 was negative in 20 (20%) and positive in 80 (80%) breast carcinomas. MRP1 expression was more frequent in both estrogen receptor-negative carcinomas and progesterone receptor-negative carcinomas (p = 0.1 in both cases), but was independent of tumor size and lymph node involvement. Patients with MRP1-negative carcinomas had prolongations of overall survival (p = 0.01 for death due to any cause, p = 0.04 for breast cancer related death) and disease-free survival (p = 0.07) as compared to those with MRP1-positive carcinomas. Also in subsets of patients (negative lymph nodes; positive lymph nodes; positive estrogen receptor; T1/T2 tumors), overall survival was longer for patients with MRP1-negative carcinomas. In univariate Cox regression analyses, MRP1 positivity was associated with relative risks of 4.9 (95% CI 1.2-20.6; p = 0.03) for death due to any cause, 6.4 (95% CI 0.9-48.0; p = 0.07) for breast cancer-related death and 3.5 (95% CI 0.8-14.9; p = 0.09) for relapse. In multivariate Cox regression analyses, MRP1 positivity had relative risks of 5.1 (95% CI 1.2-21.7; p = 0.03) for death due to any cause, 6.5 (95% CI 0.8-50.1; p = 0.07) for breast cancer-related death and 3.4 (95% CI 0.8-15.1; p = 0.1) for relapse. CONCLUSIONS: Our results suggest that MRP1 might be an important factor in breast cancer indicating excellent prognosis for patients with MRP1-negative carcinomas. PMID- 10697509 TI - Expression of the lung resistance protein (LRP) in primary breast cancer. AB - BACKGROUND: To determine the clinical significance of the lung resistance protein (LRP) in breast cancer, we have studied its expression in primary breast carcinomas (n = 99) and assessed the association of this expression with clinical parameters of the patients. MATERIALS AND METHODS: LRP expression was immunohistochemically determined by means of the monoclonal antibody LRP-56 on frozen tumor sections. RESULTS: LRP expression was negative in 12%, low in 20%, intermediate in 47% and high in 21% of the carcinomas. LRP expression was independent of age of the patients, histology, tumor grade, estrogen receptor as well as progesterone receptor status, tumor size and lymph node involvement. Kaplan-Meier analyses revealed that both overall survival and disease-free survival were independent of the degree of LRP expression. CONCLUSIONS: LRP is frequently expressed in breast carcinomas, but is neither associated with known prognostic factors, nor a prognostic factor by itself. PMID- 10697510 TI - HPV33 DNA in premalignant and malignant breast lesions in Chinese and Japanese populations. AB - The relationship between human papillomavirus (HPV) infection and breast cancer is controversial. In this study, paraffin-embedded tissue blocks prepared from 72 patients with benign, premalignant or malignant mammary lesions were randomly collected from the Shanghai region of China and Tokushima in Japan. DNA specimens extracted from all tissues were amplified by the polymerase chain reaction (PCR) using HPV16, 18 and 33 primers. Southern blot hybridization showed 19 cases to be positive for HPV33 DNA: The positive rate for HPV33 DNA in Chinese (41.7%) was significantly higher than in Japanese (11.1%) (P < 0.01): The positive rate for HPV33 DNA in invasive ductal carcinoma (34.1%) was higher than in benign or borderline mammary lesions (5%) (P < 0.02). There were no statistically significant difference among the relationship of the nuclear grade of breast cancers with HPV33 DNA-positivity. This is the first report of a positive correlation between HPV33 DNA and breast lesions in Chinese and Japanese populations. These results suggest that the infection by HPV33, but not HPV 16 or HPV 18, may be involved in breast hyperplastic lesions, especially breast cancer, in humans. PMID- 10697511 TI - Analysis of caspases that are activated during apoptosis in leukemia U937 cells in response to geranylgeraniol. AB - Affinity labeling showed that active caspases with molecular masses of 20 kDa, 19 kDa, and 17 kDa were formed upon treatment of human leukemia U937 cells with GGO. These caspases are quite similar to those activated by treatments with other apoptosis-inducers, such as VP16 and camptothecin, suggesting that similar caspases, such as caspases 3 and 6, are activated during apoptosis in U937 cells that is induced by a variety of apoptosis-inducing stimuli. An inhibitor of caspases, Z-Asp-CH2DCB, inhibited DNA fragmentation in response to GGO in vivo by blocking the cleavage of 20-kDa to 17-kDa peptides. This cleavage is catalyzed by caspase 3 itself or by a caspase-3-like protease. In contrast, other inhibitors of caspases such as Z-DEVD-FMK and Z-VAD-FMK, inhibited the processing of the caspase 3 precursor p32 to 20-kDa and 17-kDa peptides, a result which suggests that these inhibitors inhibited other upstream caspases. Treatment of U937 cells with GGO resulted in the release of cytochrome c from mitochondria prior to DNA fragmentation and the release of cytochrome c was inhibited by Zn2+ ions and by a chelator of Ca2+ ions but not by inhibitors of caspases such as Z-Asp-CH2DCB or Z VAD-FMK. These results suggest that intracellular free Ca2+ ions, and some caspases that are inhibited by Zn2+ ions, but not by Z-Asp-CH2DCB or Z-VAD-FMK are necessary for the release of cytochrome c that is caused by the treatment with GGO. PMID- 10697512 TI - Isolation and culture of ovarian tumour cells, cytological and cell survival evaluation. AB - The purpose of this study was to evaluate the reproducibility and reliability of the fluorometric microculture cytotoxicity assay (FMCA). Emphasis was placed on obtaining pure tumour cell cultures which were subjected to careful cytological evaluation. Preparations of 39 ovarian tumours, malignant, borderline and benign were made, of which 37 were successfully cultured. In 34 of the 37 tumour cell cultures, the epithelial cell fraction was > 90%, and in 30 of 39 cultures the epithelial cell fraction was > 95%. Transportation within 24 h and the 72 h incubation did not change the yield or epithelial cell fraction. There was a linear relationship between fluorescence and the number of viable cells. The fluorescence increased with time, making only comparisons within each assay plate possible. The sensitivity of the method makes it possible to perform many analyses on a small amount of material. The method also makes it possible to study cells derived from all stages of the disease, including benign tumours. PMID- 10697513 TI - Plasmid elimination and immunomodulation by 3-benzazepines in vitro. AB - For studying the mechanisms of biological activity on 3-benzazepines, antimicrobial effect, F'lac plasmid elimination activity (a plasmid curing effect on F'lac plasmid) and antibody-dependent cellular cytotoxicity (ADCC) test were performed. A weak antiplasmid effect was found at sub-inhibitory concentrations. A combination of [KF4] with verapamil [2] did not alter the ineffectivity, however, [KF4] could inhibit the antiplasmid effect of promethazine, as compared to the control (promethazine alone) plasmid curing effect. A competition between promethazine and [KF4] might exist in plasmid elimination effect. ADCC activity of human leukocytes was enhanced by KF1, KF2, KF3, DA and NE at 1.0 microgram/mL concentrations. The majority of 3-benzazepines [KS02, KM57, KN50, KE04, KI10, KP80] was ineffective for plasmid curing, however, inhibited the ADCC reaction, but they did not show a real dose-dependent effect. PMID- 10697514 TI - Comparative ability of ibuprofen and N-(4-hydroxyphenyl)retinamide to inhibit development of rat mammary adenocarcinomas associated with differential inhibition of gene expression of cyclooxygenase isoforms. AB - A rodent model of carcinogen-induced mammary tumorigenesis was used to determine the comparative growth inhibitory effects of dietary administration of either 1000 mg/kg of the non-steroidal antiinflammatory drug (NSAID) ibuprofen or 1.5 mmol/kg of the synthetic retinoid N-(4-hydroxyphenyl)-retinamide (4-HPR). In addition, the effects of these compounds on gene expression and protein production of the two isoforms of the cyclooxygenase (COX) gene which are responsible for prostaglandin production were examined. Experimental diets were provided to rats beginning at 7 days prior to administration of a single intragastric dose of 15 mg dimethylbenz[a]anthracene (DMBA) and diets were provided ad libitum until the study was terminated at 16 weeks later. Ibuprofen significantly decreased levels of gene expression of both COX-1 and COX-2 (p < 0.01). Although dietary 4-HPR did significantly diminish levels of COX-1 gene expression (p < 0.01) in rat mammary adenocarcinomas, this synthetic retinoid did not significantly inhibit COX-2 gene expression. COX-1 protein was localized to endothelial cells, infiltrating inflammatory cells, and tumor cells, while COX-2 protein was detected primarily within tumor cells. Although ibuprofen was more effective in inhibiting COX-2 gene expression than 4-HPR, ibuprofen and 4-HPR were equally effective in inhibiting development of carcinogen-induced mammary adenocarcinomas. PMID- 10697515 TI - Characterization of the anticancer activity of DW2282, a new anticancer agent. AB - DW2282 [(S)-(+)-4-phenyl-1-[N-(4-aminobenzoyl) indoline-5-sulfonyl]-4,5-dihydro-2 imidazolone].hydrochloride] was derived from diarylsulfonylurea and was identified as a prominent new anticancer agent. We examined the characteristics of DW2282 activity on the proliferation of human lung carcinoma cells, A549 and human leukemic cells, K562. DW2282 effectively inhibited cancer cell proliferation in vitro. Colony forming assay and viability tests demonstrated that DW2282 is a cytotoxic agent rather than a cytostatic agent. The isotope uptake test exhibited that DW2282 inhibited or inactivated protein synthesis. Also, under conditions which cause RNA or protein synthesis inhibition, by co treatment with actinomycin D or cycloheximide, reduced the anticancer effects of DW2282. This means that the cytotoxicity of DW2282 depends partially on RNA or protein synthesis and proteins affected by DW2282 may inactivate or alter the process of the synthesis of another protein. DW2282 activity was highly diminished in the presence of colcemid, a metaphase spindle blocker. This result suggests that DW2282 may be related to the cell cycle. After exposure to DW2282, morphologically apoptotic cells appeared in A549 cells and fragmented DNA was detected in K562 cells. It demonstrated that apoptosis is one of the mechanisms by which DW2282 inhibits the proliferation of A549 and K562 cells. PMID- 10697516 TI - Differential binding of toxic lectins from Viscum album L., ML I and ML III, to human lymphocytes. AB - The killing capacity of extracts from Viscum album L., widely used as an adjuvant in complementary cancer therapy, is dependent on the content of toxic proteins, especially the mistletoe lectins (ML). Although one may expect a homogeneous distribution of 'receptors' for these proteins on the cell surface, the sensitivity of cells to the ML-mediated cytotoxicity obviously differs, as the galNAc-binding ML III in contrast to the gal-binding ML I selectively killed CD8+ lymphocytes with a 'memory' phenotype (CD62Llo), while CD19+ B cells remained almost unaffected. B cells hardly bind ML III but did bind the gal-specific ML I. In accordance with these observations, in leukaemic B cells from patients with B chronic lymphocytic leukaemia and the human IgE-secreting myeloma cell line U-266 a strong induction of apoptosis-associated mitochondrial Apo2.7 molecules was observed after treatment with ML I and less effectively by ML III, while in the leukaemic T cell line Molt-4 both ML were strong inductors of apoptosis. In the light of these findings, the possible impact of ML I- and ML III-rich mistletoe extracts in the treatment of B cell neoplasia has to be carefully investigated. PMID- 10697517 TI - Effect of paclitaxel pretreatment on radiation-induced p53-dependent apoptosis. AB - The aim of this study was to investigate the effect of paclitaxel on radiation induced p53-dependent apoptosis. A human ependymoblastoma transplanted to nude mice was used. They were treated with paclitaxel (40 mg/kg), irradiation (2 Gy), or a combination of both. Apoptosis was markedly increased in the irradiation group. p53 protein expression was well correlated with the frequency of radiation induced apoptosis. There was only a slight increase in apoptosis in the paclitaxel group, with little p53 protein expression. In the combination group, the frequency of apoptosis varied with the time intervals, and the group irradiated 12 h after paclitaxel administration showed much less apoptosis than the irradiation group. The Ki-67 labeling index in the paclitaxel group was always higher than before administration. The present study indicates that p53 dependent apoptosis was frequently induced in the human tumor in vivo by irradiation, but not by paclitaxel alone. When combined with irradiation, the timing affected the frequency of apoptosis and the degree of p53 protein expression. PMID- 10697518 TI - Multidrug resistance due to impaired DNA cleavage in a VP-16-resistant human leukemia cell line. AB - We established a VP-16-resistant line of human leukemia cells, K562/VP-H2, derived from K562 cells. K562/VP-H2 cells were 44-fold more resistant to VP-16 than were K562 cells. K562/VP-H2 cells were also resistant to doxorubicin, daunorubicin and mitoxantrone, but showed little or no resistance to vincristine, aclarubicin, idarubicin, idarubicinol, cytosine arabinoside, cis-platinum or camptothecin. K562/VP-H2 cells did not over-express P-glycoprotein or multidrug resistance protein, and showed intracellular accumulation of VP-16 similar to that in K562 cells. While the expressions of topoisomerase II-alpha gene and topoisomerase II-beta gene, or catalytic activity in nuclear extract of K562/VP H2 cells were similar to that of K562 cells, the VP-16 induced DNA cleavage was reduced in K562/VP-H2 cells compared to K562 cells, suggesting that the reduction of topoisomerase II-mediated DNA cleavage through qualitative alteration of topoisomerase II may be the main mechanism of acquired multidrug resistance for K562/VP-H2 cells. The K562/VP-H2 cell line is an interesting model for studying resistance to antileukemia drugs targeting topoisomerase II. PMID- 10697519 TI - Histone H1 kinase activity in ovulated oocytes. AB - The level of kinase activity of cdkl is known to be high during metaphase of the two meioses. In this experiment, histone H1 kinase activity (which is known to reflect cdk1 activity) was assayed in BALB/c mouse ovulated oocytes at various timepoints after ovulation. Histone H1 kinase activity in ovulated oocytes was stable up to 37 hours after ovulation. After that time, histone H1 kinase activity significantly decreased suggesting that cdkl might be degraded after this period of time if the ovulated oocyte is not fertilised. PMID- 10697520 TI - Reverse transformation of human mammary carcinoma cells. AB - Exposure of cells derived from human mammary carcinoma cell line, MaTu, to daunorubicin started a selection process which reproducibly gave rise to sublines with different phenotypes. One subline exhibited a fibroblast-like morphology (MaTu/c7), while others retained the epitheloid phenotype of the parental cells (MaTu/p). Among the latter was clone 8 (MaTu/c8) which displayed piling-up structures not seen in MaTu/p cells. Striking differences were detected on immunocytochemistry using the anti-cytokeratin 19 antibody A53-B/A2 which positively reacted with cells from MaTu/c7, but not with those of MaTu/c8 and MaTu/p. In contrast, the anti-blood group H 2 antibody A46-B/B10 positively stained cells from MaTu/c8 and MaTu/p, but not those of MaTu/c7. Assays for tumorigenicity in nude mice demonstrated that MaTu/c7 is far less tumorigenic than MaTu/p, while MaTu/c8 showed a pattern distinguishing it from MaTu/p cells. Cross-resistance assays showed decreasing drug resistance in the order MaTu/c8 > MaTu/c7 > MaTu/p. These data suggest drug-induced differentiation with reversion of the neoplastic phenotype in MaTu/c7 and some form of malignant progression in MaTu/c8. This model system may be helpful for understanding cancer development, especially its relation to differentiation. PMID- 10697521 TI - Development of a very sensitive luminescence assay for the measurement of paclitaxel and related taxanes. AB - A rapid and very sensitive enzyme immunoassay was developed for the measurement of paclitaxel and related taxanes in crude extracts of Taxus sp., in human serum and in culture medium of paclitaxel-producing microorganisms such as Erwinia taxi. For the ELISA, paclitaxel was chemically modified by the introduction of an amine to enable coupling with biotin. The presence of paclitaxel or related taxanes competitively inhibited the binding of paclitaxel-biotin to anti-taxane monoclonal antibody. This method detected paclitaxel in concentrations as low as 33 pM; the affinity of the antibody was higher for paclitaxel than for cephalomanine, baccatin and DAB. The sensitivity of this assay makes it useful for estimating the paclitaxel and taxanes content of Taxus sp. extracts, monitoring the paclitaxel serum level of paclitaxel treated patients and in other biological fluids. PMID- 10697522 TI - De novo deletions of p53 gene and wild-type p53 correlate with acquired cisplatin resistance in human osteosarcoma OST cell line. AB - BACKGROUND: Initial p53 status is a useful determinant of chemoresistance or chemosensitivity of primary tumors, however, it remains unclear whether p53 status is a critical chemoresistant marker in tumors that acquire drug-resistance after the initiation of chemotherapy. We investigated the relationship between p53 status and the development of resistance to cisplatin in osteosarcoma cell lines. MATERIALS AND METHODS: Cisplatin-sensitive human osteosarcoma OST cells and acquired cisplatin-resistant OST/R cells derived from OST cells were used. Single-strand conformation polymorphism (SSCP) analysis of exons 5 to 8, and immunohistochemistry using anti-p53 antibodies were analyzed to detect mutations of p53. Fluorescence in situ hybridization (FISH) and enzyme immunoassay (EIA) were performed to detect deletions of p53. RESULTS: SSCP and immunohistochemistry revealed that both cell lines had wild-type p53 gene and protein. However, in OST/R cells, genomic instability of chromosome 17 and de novo deletion of the p53 gene located in chromosome 17p were detected by FISH. The constitutive levels of wild-type p53 protein measured by EIA were significantly lower in OST/R cells than in OST cells. Furthermore, p53 induction was lost in OST/R cells after cisplatin exposure. CONCLUSIONS: De novo deletions of the p53 gene and wild-type p53 were associated with the acquisition of cisplatin-resistance in osteosarcoma. PMID- 10697523 TI - Effects of long-term administration of UFT plus leucovorin on colorectal tumors induced with 1,2-dimethylhydrazine in rats. AB - Thymidylate synthase and thymidine kinase are key enzymes involved in the de novo and salvage pathways for pyrimidine nucleotide synthesis, respectively. Thymidylate synthase is inhibited by 5-fluorodeoxyuridine monophosphate, forming an inactive ternary complex with intracellular folate. We investigated the effects of 1-(2-tetrahydrofuryl)-5-FU plus uracil (UFT) with or without leucovorin on 1,2-dimethylhydrazine-induced rat colorectal carcinomas. Thirty week administration of UFT with or without leucovorin markedly suppressed both colorectal carcinogenesis and tumor growth, resulted in the increase of thymidylate synthase inhibition and the decrease of thymidine kinase activity in the tumor cells. These results indicate that the combination of UFT with leucovorin could be useful in the development of pre- and post-operative adjuvant chemotherapy programs. PMID- 10697524 TI - Effects of a low dose leucovorin with 5-fluorouracil derivative on colorectal tumors induced with 1,2-dimethylhydrazine in rats. AB - Thymidylate synthase, which is a key enzyme involved in the de novo pathway for pyrimidine nucleotide synthesis, is inhibited by 5-fluorodeoxyuridine monophosphate, forming an inactive ternary complex with intracellular folate. We investigated the effect of a 5-fluorouracil derivative (UFT) with or without low dose leucovorin on the number of 5-fluorodeoxyuridine monophosphate binding sites, thymidine kinase activity and intracellular folate concentration in 1,2 dimethylhydrazine-induced rat colorectal carcinomas. A 10-day administration of UFT with or without leucovorin enhanced the thymidine kinase activity and the number of 5-fluorodeoxyuridine monophosphate binding sites, with an increase of thymidylate synthase mRNA expression. Thymidylate synthase inhibition was slightly increased as the intracellular folate concentration increased. These results indicate that thymidylate synthase inhibition increases when the intracellular folate is exogenously supplemented and maintained at an adequate concentration. PMID- 10697525 TI - Effect of 4-hydroxynonenal, a product of lipid peroxidation, on natural cell mediated cytotoxicity. AB - Lipid peroxidation of cell membrane yields a variety of final products whose a quantitatively important component is 4-hydroxynonenal (HNE). Previous studies performed in our laboratory suggest that HNE may play a physiological role in the control of cellular proliferation and/or differentiation. This appears to be further supported by our recent findings showing that pre-treatment of K562 cells with a physiological concentration of HNE leads to a marked reduction of susceptibility of NK cells. The observed regulatory effects of HNE on tumor cell growth and susceptibility to natural immune resistance, led us to try to better understand the immunotoxicological properties of this aldehyde. The present study analyses the effects of HNE on NK-mediated cytotoxicity. Treatment of MNC as effector cells with concentrations of HNE ranging from 0.001 to 1 microM for 1 h, did not produce noticeable effects on NK activity. Therefore, this aldehyde at physiological concentrations is able to differentiate tumor cells and to down regulate target susceptibility to NK effectors from one side. On the other side, it is not able to modify the efficiency of the NK function. Moreover, HNE concentrations higher than 1 microM showed significant and concentration dependent inhibition of NK activity. However, this effect is reversible and can be antagonized, at least in part, by treatment of effector cells with HNE in combination with beta-interferon. PMID- 10697526 TI - Chemosensitivity testing of primary tumor cells from gastric cancer patients with liver metastasis can identify effective antitumor drugs. AB - The liver metastasis of gastric carcinoma is resistant to conventionally available treatment. Twenty patients with liver metastasis of gastric cancer were treated by arterial drug infusion using a reservoir and seven cases were treated with systemic chemotherapy. The resected primary gastric cancer specimen was used for chemosensitivity assay with 3-(4,5-dimethylthiazol-2yl)-2,5-diphenyl-2H tetrazolium bromide (MTT) endpoint, and the patients were treated without reference to the results of the chemosensitivity assay. The mean survival period was assessed according to the histology of the primary lesion, the grade of liver metastasis and, the presence of peritoneal dissemination. No significant differences were observed in the primary tumor histology and grade of liver metastasis, but the survival period of the patients with liver metastasis and peritoneal dissemination was significantly shorter than that of the patients without peritoneal dissemination. Nine patients were treated with drugs that were effective in the chemosensitivity assay, and their responses included two complete responses and two partial responses; these patients showed a significantly prolonged survival period compared with patients treated with drugs that were not effective in the assay. The chemosensitivity assay is useful for evaluating the effectiveness of antitumor agents against liver metastasis of gastric cancer. PMID- 10697527 TI - Acquisition of multidrug resistance in osteosarcomas, analyzed by doxorubicin binding assay, and histologic response to chemotherapy. AB - BACKGROUND: The mechanism by which multidrug resistance is acquired in human osteosarcomas remains unclear. In this study, we analyzed the changes in doxorubicin (DOX) binding ability (%DB), showing chemosensitivity before and after chemotherapy with DOX and cisplatin (CDDP), and evaluated the histologic response of human osteosarcomas to chemotherapy. MATERIALS AND METHODS: Eight osteosarcomas were analyzed. %DB by the DOX binding assay was measured in fresh tumor tissues at biopsy and at resection after preoperative chemotherapy. The histologic response to chemotherapy was also evaluated. RESULTS: Changes in %DB before and after chemotherapy were classified into 4 patterns; A: high (> 80%) to low (< 80%), B: high to high, C: low to low, D: low to high. The two tumors, classified as type A and B were responders (> 90% necrosis), whereas the 6 tumors of type C and D were classified as non-responders (< 90% necrosis). Based on these data, we speculated that the fraction of cell populations with different chemosensitivities to DOX and CDDP varied and found that type B tumors have no population of resistant cells, whereas type C tumors have a large fraction of resistant cells. CONCLUSION: We concluded that osteosarcomas are composed of mixed cell populations with different chemosensitivities to anticancer agents and also that survival of multidrug resistant cell populations may be the most important mechanism in acquisition of multidrug resistance of osteosarcomas after chemotherapy. PMID- 10697528 TI - Morphology of the designed biodegradable cisplatin microsphere. AB - We observed the morphological changes in microspheres containing cisplatin (CDDP) embedded in poly-d,l,-lactic acid (PLA) and polyethylene glycol acid (CDDP-PPMS). The in vitro release of CDDP from CDDP-PPMS continued for more than four weeks. Scanning electron microscopy revealed that though the particles of CDDP-PPMS were spherical and superficially smooth, small pores with a superficial irregularity appeared six months later. CDDP-PPMS particles were found in the stomata of the omentum after intraperitoneal administration. When CDDP-PPMS was administered after Yoshida sarcoma cells were intraperitoneally transplanted, CDDP-PPMS was observed histologically in the necrotic tissues of the omentum. These experimental results confirmed the release of CDDP from CDDP-PPMS and the histological efficacy of CDDP-PPMS in the omentum against peritoneal metastasis. PMID- 10697529 TI - Unsaturated fatty acid feeding prevents the development of acute hepatitis in Long-Evans cinnamon (LEC) rats. AB - To investigate the effects of dietary alpha-linolenic acid (18:3, n-3; alpha-LNA) and linoleic acid (18:2, n-6; LA) on the development of hereditary hepatitis, we compared incidences and grades of acute hepatitis between the Long-Evans cinnamon (LEC) rats fed with safflower oil-supplemented diet and perilla oil-supplemented diet. Both safflower and perilla oil supplemented diets reduced the incidence of hepatitis and significantly prolonged its onset as compared to the non supplemented conventional diet. No significant difference was observed between safflower and perilla oil diets in the rats of incidence of hepatitis. At the age of 16 weeks, just before the onset of hepatitis, serum levels of transaminase (AST, ALT) and concentration of copper in rats fed with both test diets were significantly reduced as compared with that of rats fed alpha-linolenate and linoleate have an inhibitory effect on the development of hepatitis in LEC rats due to the prevention of serum copper elevation. PMID- 10697530 TI - Dibutyryl cyclic AMP-induced enhancement of tissue inhibitor of metalloproteinases-3 expression and its possible relation to the invasive activity of the human hepatoma cell line PLC/PRF/5. AB - The effects of dibutyryl cyclic AMP (DBcAMP) on tissue inhibitor metalloproteinase (TIMP) expression were studied in the human hepatoma cell line PLC/PRF/5 with relation to the invasive activity of the cells. Messenger RNA expression levels of metalloproteinases (MMP)-2 and 9, and TIMP-1, 2 and 3 in the cells were determined by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). MMP-9 and TIMP-2 mRNA expression were not detectable in the cells with or without DBcAMP treatment. Relative MMP-2 and TIMP-1 mRNA expression levels in the cells were not affected by DBcAMP, whereas TIMP-3 mRNA expression was enhanced by DBcAMP. This stimulatory effect of DBcAMP on TIMP-3 expression was confirmed at the mRNA and intracellular protein levels by Northern and Western blotting, respectively. Invasive activity of PLC/PRF/5 cells was determined using the in vitro Matrigel (extracellular matrix extract) invasion model. DBcAMP inhibited the invasive activity of the cells, and this effect was eliminated by addition of an antisense oligonucleotides corresponding to TIMP-3 mRNA. These results suggested that TIMP-3 expression is enhanced by DBcAMP and may play a role in inhibition of the invasive activity of hepatoma cells. PMID- 10697531 TI - Dibutyryl cyclic AMP-induced enhancement of RB protein degradation in human hepatoma cells. AB - Dibutyryl cyclic AMP (DBcAMP) was previously reported to enhance the down regulation of the retinoblastoma (RB) protein during G1 phase in proliferating primary rat hepatocytes, but to inhibit their entry into S phase and RB phosphorylation. In the present study, DBcAMP was also found to enhance the down regulation of RB protein in the human hepatoma cells PLC/PRF/5 after hydroxyurea induced synchronization at G1/S phase. One hour after synchronization, CPP32 activity was detected in the cells and was further enhanced in the presence of DBcAMP. CPP32-specific cleavage of the RB protein was also detected and enhanced by the addition of DBcAMP in a dose-dependent manner. DNA analysis by flow cytometry after serum starvation-induced synchronization at G0/G1 phase revealed that DBcAMP elicited an apoptotic peak after the S phase. Based on these findings, DBcAMP was suspected of inducing apoptosis by RB protein degradation during G1/S transition and thereby inhibit the growth of PLC/PRF/5 cells. Under serum-deficient culture conditions, addition of the CPP32 inhibitor DEVD or the ICE inhibitor YVAD enhanced cell growth but did not abolish the DBcAMP-induced growth inhibition. On the other hand, antisense oligodeoxynucleotides against Bcl 2 mRNA showed a growth inhibitory effect on PLC/PRF/5 cells, but did not show an additive effect on the DBcAMP-induced growth inhibition. DBcAMP itself inhibited bcl-2 protein expression. DBcAMP-induced growth inhibition may be mediated by different mechanisms, including apoptosis. PMID- 10697532 TI - Human megakaryocyte polyploidization is associated with a decrease in GPIIIA expression. AB - Megakaryocytes are platelet forming cells and are characterized by polyploidization, a phenomenon by which nuclear division occurs without corresponding cytoplasmic separation. Among the markers allowing to identify megakaryocytes, glycoprotein (GP) IIIa with GPIb and GPIIb are the most important. Using GPIIIa as a marker to recognize megakaryocytes in the bone marrow, we have estimated GPIIIa expression by flow cytometry in megakaryocyte populations from normal individuals and from patients with chronic myelogenous leukemia, immune thrombocytopenic purpura or polycythemia vera. We showed that the expression of GPIIIa is decreasing during megakaryocyte polyploidization in normal and pathological situations. PMID- 10697533 TI - Specific H-Ras minisatellite alleles in breast cancer susceptibility. AB - Mutations in BRCA1 and BRCA2 genes account for the majority of familial aggregation of breast and ovarian cancers but other common genes in the population with low penetrance should be also involved in susceptibility to breast cancer. The H-ras minisatellite, located downstream of H-ras oncogene, is considered to be a likely candidate. Previous findings have estimated that as many as 1 in 11 cancers of the breast might be attributed to this region, but other studies observed inconsistent results. We propose to elucidate the potential role of H-ras locus in breast cancer, by looking at somatic alterations occurring in tumor DNAs such as the instability or the loss of heterozygosity (LOH) and by determining a potential correlation between constitutional specific H-ras alleles and clinical and/or pathological characteristics. DNA was extracted from 123 sporadic breast tumors and matched peripheral blood lymphocytes. 143 DNA samples from of peripheral blood lymphocytes from healthy donors served as a control population. The allelic diversity was determined by polymerase chain reaction analysis. Rare H-ras alleles were found to be present in about 9% of breast cancer patients while they were detected in only 1.4% of lymphocytes from healthy donors (P = 0.0044). Therefore, the risk of breast cancer is increased in patients with one or two rare alleles (odd ratio = 7.14 and 95% confidence interval = 1.94-22.27). Analyses of somatic alterations in tumor DNA have shown the lost of one allele, in general the longest, in 6.7% informative cases and an instability to H-ras locus in 6.5% tumors that appeared as a size increase of one of the two alleles. No correlation of rare H-ras alleles with clinicopathological parameters was found. Our results demonstrated an association of rare H-ras alleles with breast cancer and suggest that minisatellite H-ras may be considered as an informative marker for the breast cancer risk. PMID- 10697534 TI - Inhibition of epigallocatechin gallate-induced apoptosis by CoCl2 in human oral tumor cell lines. AB - Epigallocatechin gallate (EGCG) induced apoptotic cell death in two human oral tumor cell lines (HSC-2, HSG), as judged by TUNEL method which detects DNA nick. Furthermore, the cytoplasm of EGCG-treated HSG cells was stained by M30 monoclonal antibody, which detects the degradation product of cytokeratin by activated caspase. The apoptosis-inducing activity of EGCG was significantly reduced by millimolar concentrations of CoCl2. CoCl2 also inhibited the cytotoxic activity of sodium ascorbate, gallic acid and curcumin, but not that of sodium-5, 6-benzylidene-L-ascorbate (SBA). This suggests that SBA, an antitumor agent, induces cell death by a different mechanism from that of other antioxidants used in this study. The possible role of CoCl2 for cell survival was discussed. PMID- 10697535 TI - Expression of activated c-erbB-2 oncogene induces sensitivity to cisplatin in human gallbladder adenocarcinoma cells. AB - Overexpression of the c-erbB-2/HER-2/neu protooncogene which encodes for the tyrosine kinase receptor p185neu, has been observed frequently in cisplatin resistant human tumors, such as colorectal, breast, and non-small-cell lung cancers, and is known to induce resistance to cisplatin (CDDP) in vitro. To confirm a direct relationship between erbB-2 expression and CDDP resistance, we examined the role of erbB-2 in the cellular sensitivity to cisplatin using erbB-2 transfected HAG-1 human gallbladder adenocarcinoma cell lines. Three out of four cell lines, which stably expressed ErbB-2 protein (p185neu), did not show CDDP resistance but acquired sensitivity to cisplatin, compared to non-transfected cells. This chemosensitivity appears to be inversely correlated with the abundance of p185neu. Although the mechanism still remains unclear, these results suggest that sensitivity to CDDP in erbB-2 expressed cells may vary, depending on the cell type. PMID- 10697536 TI - A new analytical method for the detection of anti-tumor promoters using flow cytometry. AB - Flow cytometry analysis was applied to the measurement of Epstein-Barr virus (EBV) induction which is used as a short term assay for anti-tumor promoters. The data obtained by measurement with flow cytometry were parallel with those of the fluorescence microscopic method. Flow cytometry is rapid, quantitative and should be applicable for the EBV activating test. PMID- 10697537 TI - Relevance of ploidy related parameters for prognosis in malignant fibrous histiocytomas. AB - Ploidy associated parameters were determined in 161 malignant fibrous histiocytomas (MFH) by DNA image cytometry. The rate of euploid cases in MFH with histopathological grade I was 42%, grade II 16% and grade III 6%. Univariate analyses showed that cases with a low percentage of diploid cells (< 40%) and cases with high rates of aneuploid cells between 4c and 8c (> 9%), octaploid cells (> 0%) and 16-ploid cells (> 0%), high 2.5c (> 60%) and 5c (> 16%) exceeding rates and cases with high DNA indices of the first (> 1.08) and the second (> 3.78) peak had significantly reduced survival rates. Multivariate analyses with Cox regression demonstrated that in addition to grade, only the percentage of aneuploid cells between 4c and 8c, the 5c exceeding rate and the DNA index of the first peak had independent prognostic impact. It was concluded that besides ploidy status and grading, ploidy associated parameters are suited for prediction of survival in MFH. PMID- 10697538 TI - Antiproliferative activity in vitro of side-chain analogues of calcitriol against various human normal and cancer cell lines. AB - The antiproliferative in vitro activity of side-chain modified analogues of 1,25 dihydroxyvitamin D3 was examined in order to select compounds with potential antitumour activity. Analogues PRI-1906, PRI-1907, PRI-1909, PRI-2191, PRI-2192, PRI-2193 and PRI-2194 were examined for their antiproliferative activity in vitro against a spectrum of various human cancer cell lines using the MTT technique. In addition, analogues PRI-1906 and PRI-2191 were screened against cells of human leukaemia HL-60 line and against normal human skin fibroblasts. Calcitriol and these two analogues revealed strong antiproliferative activity against these two targets with maximal growth inhibition of 68% for HL-60 cells and of 60% for fibroblasts, and this effect was dose dependent. All analogues tested, except PRI 1909, revealed antiproliferative activity against human carcinoma cell lines of breast origin applied, namely against T47D and MCF-7. The maximal growth inhibition of 49% for T47D cell line and 39% for MCF-7 line was observed, and this effect was dose dependent. The inhibitory doses of the analogues tested were compared with the indices for calcitriol. Analogue PRI-1906 revealed the strongest antiproliferative activity against these four target cell lines (HL-60, fibroblasts, MCF-7, and T47D). The novel analogues of calcitriol, similarly to calcitriol, appeared to be not active against other human cancer cell lines tested (including those originated from lung, colon, prostate, urinary bladder, ovary, pancreas, stomach and kidney) revealing an antiproliferative activity not exceeding 20%. The mechanism of the observed antiproliferative effect of calcitriol and its analogues in vitro remains unclear, however, it may be related to their effect on cell differentiation. The appearance of antigen CD14 and CD11b expression after exposure to calcitriol and its new analogues confirmed their effect on cell differentiation. PMID- 10697539 TI - Dietary beta-carotene and astaxanthin but not canthaxanthin stimulate splenocyte function in mice. AB - The in vivo modulatory effect of beta-carotene, astaxanthin and canthaxanthin on lymphocyte function was investigated. Female BALB/c mice (8 wk old) were fed a basal diet containing 0, 0.1% or 0.4% beta-carotene, astaxanthin or canthaxanthin for 0, 2 or 4 wk (n = 8/diet/period). Splenic lymphocytes were isolated and mitogen-stimulated proliferation, IL-2 production and lymphocyte cytotoxicity were assessed. Body weight and feed intake were not different among dietary treatments. Plasma carotenoids were undetectable in unsupplemented mice but concentrations of the respective carotenoids were elevated in mice fed 0.1 or 0.4% beta-carotene (0.22 and 0.39 mumol/L), astaxanthin (16.4 and 50.2 mumol/L) and canthaxanthin (5.00 and 7.02 mumol/L) respectively. Mice fed both dietary levels of beta-carotene and astaxanthin had enhanced phytohemagglutinin-induced lymphoblastogenesis compared to unsupplemented mice (P < 0.03). No treatment difference was detected with concanavalin A- or lipopolysaccharide-induced lympho proliferation nor with IL-2 production (P < 0.05). Astaxanthin (0.1%) also enhanced lymphocyte cytotoxic activity (P < 0.08). In contrast, canthaxanthin did not significantly influence any of the lymphocyte functions measured. Results indicate that beta-carotene and astaxanthin but not canthaxanthin exert enhanced splenic lymphocyte function in mice. PMID- 10697540 TI - Effects of flavonoids on the growth and cell cycle of cancer cells. AB - In this study, we investigated the cytotoxicities of flavone (F01), 3 hydroxyflavone (F02), 6- hydroxyflavone (F03), 7-hydroxyflavone (F04), 3,6 dihydroxyflavone (F05), 5,7-dihydroxyflavone (F06) and 5,6,7-trihydroxyflavone (F07) to human cancer cells including P- glycoprotein (Pgp)-expressing HCT15 cells and its multidrug resistant subline, HCT15/CL02 cells. We also examined the effects of those flavonoids on the cell cycle of these cancer cells. HCT15/CL02 cells did not reveal resistance to all the flavonoids tested in comparison with HCT15 cells. In cell cycle analysis, all the flavonoids tested, except F01 and F04, reduced the G0/G1 population of SF295 cells at growth inhibitory concentrations, and increased G2/M (F02, F03 and F06) or S (F05 and F07) populations. In addition, F02 and F03 decreased the G2/M and G0/G1 population, and increased the S and G2/M population in HCT15 cells, respectively. Meanwhile, in HCT15/CL02 cells, F02 and F03 decreased the G0/G1 populations and increased the S population. In conclusion, we deemed that the flavonoids tested had diverse cytotoxic mechanisms, and exerted their cell growth inhibitory or killing activity by distinctive ways in different cells. PMID- 10697541 TI - Improving the cytometric detection of doxorubicin resistance in osteosarcoma cells by determining cellular doxorubicin/DNA ratio. AB - To study the influence of cellular DNA content on the accumulation and efficacy of doxorubicin (DOX), we characterized P-glycoprotein (Pgp)-positive and negative murine osteosarcoma cell clones that had a different DNA index. Statistical analysis demonstrated that the cytotoxic effects of DOX correlated significantly with the ratio of intracellular DOX accumulation divided by the cellular DNA content (DOX/DNA ratio) (P = 0.001), but not with the intracellular DOX accumulation (P = 0.16). We also tested this relationship for Pgp-negative human osteosarcoma cell lines with a different DNA ploidy, and found that these Pgp-negative cell lines all had similar DOX/DNA ratios. These results indicate that the DOX/DNA ratio is a determinant for the effects of DOX in osteosarcoma cells, regardless of their Pgp status and DNA ploidy. Thus, consideration of the cellular DNA content as well as the intracellular DOX accumulation is important to accurately detect DOX resistance. PMID- 10697542 TI - Mucins as immunogenic targets in cancer. AB - Abnormal mucins are overexpressed by many malignant adenocarcinomas. The variation in their expression and glycosylation makes certain immunodominant peptide core epitopes available for immunological recognition. We reviewed the structural differences between "native" mucins and those detected on malignant cells, a knowledge of which would aid in the design of better anti-mucin vaccines for use in several carcinomas. We also considered the character of inducible anti MUC1 immune responses reported from recent animal experiments as well as the role of MUC1-transgenic animal models in understanding mucin immunoreactivity. We concluded that the provocation of an anti-MUC1 immune response may be used for the development of a vaccine strategy, which holds promise for therapeutic antineoplastic interventions. PMID- 10697543 TI - The plant alkaloid usambarensine intercalates into DNA and induces apoptosis in human HL60 leukemia cells. AB - Usambarensine is a plant alkaloid isolated from the roots of Strychnos usambarensis collected in Central Africa. This bis-indole compound displays potent antiamoebic activities and shows antigardial, antimalarial and cytotoxic effects. Usambarensine is highly toxic to B16 melanoma cells and inhibits the growth of leukemia and carcinoma cells. To date, the molecular basis for its diverse biological effects remains totally unknown. However, its capacity to inhibit nucleic acids synthesis in melanoma cells, on the one hand, and its structural analogy with DNA-binding pyridoindole plant alkaloids recently studied (cryptolepine and matadine), on the other hand, suggested that usambarensine could also bind to DNA. Consequently, we studied the strength and mode of binding to DNA of usambarensine by means of absorption, circular and linear dichroism. The results of the optical measurements indicate that the alkaloid effectively binds to DNA and behaves as a typical intercalating agent. Biochemical experiments indicated that, in contrast to cryptolepine and matadine, usambarensine does not interfere with the catalytic activity of topoisomerase II. Human HL60 leukemia cells were used to assess the cytotoxicity of the alkaloid and its effect on the cell cycle. Usambarensine treatment is associated with a loss of cells in the G1 phase accompanied with a large increase in the sub-G1 region which is characteristic of apoptotic cells. The DNA of usambarensine treated cells was severely fragmented and the proteolytic activity of DEVD caspases is enhanced. Usambarensine is thus characterized as DNA intercalator inducing apoptosis in leukemia cells. PMID- 10697544 TI - Early effects of retinoic acid on proliferation, differentiation and apoptosis in non-small cell lung cancer cell lines. AB - BACKGROUND: Retinoids represent a potentially useful class of drugs in the chemoprevention and treatment of cancer, due to their ability to regulate cell proliferation and differentiation. However, there is controversy in the literature about the effects of all-trans retinoic acid (ATRA) in non- small cell lung cancer (NSCLC). In this study we examined the effects of ATRA on apoptotic death in NSCLC. MATERIALS AND METHODS: Cell proliferation was determined by thymidine incorporation in cultured NSCLC cells. DNA fragmentation was measured in NSCLC cell lines as a marker of apoptosis. The expression of keratinocyte transglutaminase and cytokeratin 10 were measured as markers of squamous differentiation. RESULTS: ATRA inhibited cell proliferation, and induced markers of both apoptosis and squamous differentiation after 24-48 hrs of treatment. CONCLUSIONS: These results indicate the possibility of the early growth inhibitory and apoptotic effects of ATRA in NSCLC which may result in selection of ATRA-resistant cells. PMID- 10697545 TI - Is placental tissue protein 17b/TIP47 a new factor in cervical cancer genesis? AB - We identified novel members of the placental tissue protein 17 (PP17) protein family which consists of different-size variants of PP17; cDNAs of PP17a and PP17b variants have also been cloned and sequence analyzed. By Western-blot analysis in cervical carcinoma tissues we found overexpression of PP17b. Compared to healthy controls a mean five-fold increase in the amount of PP17b was also detected in the sera of untreated cervical carcinoma patients, which declined after radical operations. In our recent findings, in sera of inoperable cervical carcinoma patients, we also found elevated PP17b levels which did not change after irradiation. By Northern-blot analyses we confirmed PP17b overexpression in cervical carcinoma tissues and also the alternative splicing of PP17 mRNAs in various normal human tissues. Presently, the amino acid sequence of TIP47--a mannose-6-phosphate receptor cargo selection device--turned out to be identical to that of PP17b. Due to its oncodevelopmental function, PP17b/TIP47 is more than likely to be connected to HSV-2 infection, which is probably one of the main etiopathogenic agents of cervical carcinoma along with the HPV virus, and may open a new trend in the research of pathological processes in human uterine cervical cancer. PMID- 10697546 TI - Metexyl (4-methoxy-2,2,6,6-tetramethylpiperidine-1-oxyl) as an oxygen radicals scavenger and apoptosis inducer in vivo. AB - A stable nitroxide radical named Metexyl (4-methoxy-2,2,6,6-tetramethylpiperidine 1-oxyl) was synthesized and its antioxidant and antitumor properties were investigated and compared with these of another nitroxide derivatives previously designed in our laboratories. Three experimental models were used: xanthine/xanthine oxidase system, pulse radiolysis and experimental rat cancer (Yoshida Sarcoma) in vivo. In this work we measured the rate constant of the reactions of Metexyl with enzymatically generated O2.- or radiolytically produced .OH. For comparison, the reactions of non radical derivative (4-acetamide-2,2,6,6 tetramethylpiperidinium acetate) or nitroxide Tempace (4-acetamide-2,2,6,6 tetramethylpiperidine-1-oxyl) with the above mentioned reactive oxygen radicals were also studied. The comparative ability of Metexyl to act as an inducer of apoptosis in vivo was also investigated in pharmacological test. The ring substituent (-OCH3) at position 4 of the Metexyl molecule had significant influence on its properties as antioxidant and apoptosis inducer. The results in this study suggest that Metexyl is a promising nitroxide antioxidant, which can induce apoptosis of tumor cells in vivo, thus providing a base for its further investigations in vitro and pharmacological applications. PMID- 10697547 TI - Irradiation induced clonogenic cell death of human malignant glioma cells does not require CD95/CD95L interactions. AB - BACKGROUND: Radiotherapy is the single most effective therapy for malignant gliomas. Targeting the CD95 apoptotic pathway is a promising experimental approach to these neoplasms. Here, we asked whether irradiation modulates CD95 mediated apoptosis of human malignant glioma cells in vitro. MATERIALS AND METHODS: LN-18, LN-229 and T98G human malignant glioma cell lines were irradiated with dosages from 0-8 Gy and treated with CD95L (CD95 ligand). CD95 expression was assessed by flow cytometry. Caspase activity was determined by DEVD cleavage. Cytotoxic effects were assessed by crystal violet staining of cells in a 96-well plate assay. Clonogenic cell death was determined by a standard colony forming assay. RESULTS: We find that (i) CD95L-induced apoptosis, but not irradiation induced clonogenic cell death, involves caspase 3 activation and is blocked by the viral caspase inhibitor, crm-A. (ii) Irradiation does not modulate CD95 expression either in p53 wild-type or in p53 mutant glioma cell lines, and does not enhance CD95L-evoked caspase 3 activity or CD95L-induced clonogenic cell death. CONCLUSIONS: We conclude that endogenous CD95/CD95L interactions are not involved in radiation-induced clonogenic cell death and that the killing cascades of CD95L and irradiation are independent in human malignant glioma cells. PMID- 10697548 TI - Elevation of glutathione in RAW 264.7 cells by low-dose gamma-ray irradiation and its responsibility for the appearance of radioresistance. AB - We examined the relationship between the induction of glutathione (GSH) level in macrophage-like RAW 264.7 cells by a low (adapting) dose gamma-rays and the cell damage caused by a lethal dose of gamma-rays at various intervals after the adapting dose. The reduced glutathione (GSH) level increased soon after exposure of the cells to 25 cGy of gamma-rays, peaked between 3 hr and 6 hr, and returned almost to the zero time (0 hr) level by 24 hr post-irradiation. Cell damage was assessed by measuring the 3H-thymidine (3H-TdR) incorporation into cellular DNA. gamma-Ray irradiation produced a dose-dependent cell damage in RAW cells, causing about 40% and 60% inhibition of 3H-TdR incorporation into DNA at 1.0 Gy and 2.0 Gy, respectively, as compared with non-irradiated cells. Treatment with the adapting dose of 25 cGy at 1 hr or 24 hr before the lethal irradiation was ineffective. However, pre-irradiation with 25 cGy at 3 hr or 6 hr prior to lethal irradiation inhibited the decrease of 3H-TdR incorporation into DNA, indicating a protective effect. GSH exogenously added to the medium also inhibited the cell damage induced by lethal doses of gamma-Rays in a dose-dependent manner. These results indicate that the induction of endogenous GSH in living cells immediately following low-dose gamma-Ray irradiation is at least partially responsible for the appearance of radioresistance to a subsequent lethal dose of radiation, and may make it possible to use higher doses of radiation in radiotherapy for tumor patients. PMID- 10697549 TI - Immunohistochemistry of DNA fragmentation factor in human stomach and colon: its correlation to apoptosis. AB - DNA fragmentation factor (DFF) is an important factor in the pathway leading to apoptosis, which is activated by caspase-3 and is involved in the formation of nuclear DNA fragments. DFF is a heterodimic protein of 40kDa and 45kDa that becomes activated when DFF is cleaved by caspase-3. Of the two enzymatically cleaved fragments of DFF, it is the 40kDa fragment (DFF40) that is the active component of DFF and is responsible for triggering chromatin condensation when incubated with nuclei. However, the topological correlation between apoptosis and DFF expression in human tissues has not been examined. Therefore, in this study, we first immunolocalized DFF in non-neoplastic mucosa, hyperplastic polyp, adenoma and carcinoma of human stomach and colon. We then examined apoptosis in serial tissue sections. Labeling index (LI) of DFF and TUNEL positive cells in the same areas of serial tissue sections were obtained using computer-assisted image analysis. In the stomach, the DFF LI in non-neoplastic mucosa (9.8 +/- 5.0%, n = 3) and carcinoma (18.2 +/- 3.6, n = 3) were significantly lower than that of hyperplastic polyp (73.3 +/- 9.2%, n = 3) and adenoma (66.5 +/- 18.3%, n = 3) [p < 0.0001]. In colon, the DFF LI in non-neoplastic mucosa (10.2 +/- 6.4%, n = 3) was significantly lower than that of hyperplastic polyp (56.0 + 34.7%, n = 3) [p = 0.0013] and adenoma (30.1 +/- 16.3%, n = 3) [p = 0.0037]. Cells positive for DFF were much more widely distributed than TUNEL positive cells in both non pathologic and pathologic mucosa of human stomach and colon. Notably, DFF positive cells were present beneath the TUNEL positive cells in non-pathological gastric and colonic epithelium. In addition, there was a significant positive correlation between DFF and TUNEL LIs in human stomach and colon [p < 0.0001]. These results suggest that DFF may be involved in the process of apoptosis in human gastric and colonic mucosa. PMID- 10697550 TI - Involvement of p38 MAP kinase in apoptotic and proliferative alteration in human colorectal cancers. AB - Mitogen-activated protein kinases (MAPKs) have been thought to be involved in tumor development, and recently implicated in the regulation of apoptosis. We assessed the activation of p38 MAPK and extracellular signal-regulated kinases (ERKs) in human colorectal adenocarcinoma by immunoblotting with antibodies raised against each activated form. We also assessed the alteration of proliferative and apoptotic states, and analyzed the association of p38 MAPK with these alterations. The incidence of p38 MAPK activation in the cancer nucleus was significant (p = 0.0215), while ERKs activation was not significant. Proliferating cell nuclear antigen (PCNA)-labeling index and apoptotic index were increased in all cancer tissues. These findings suggested that p38 MAPK was constitutionally activated and was associated with increased proliferative and apoptotic states in colorectal cancers. Serum-deprivation-induced apoptosis in colonic adenocarcinoma cell line was significantly inhibited by SB203580, a specific p38 MAPK inhibitor, suggesting that apoptosis was mediated by the p38 MAPK pathway. PMID- 10697551 TI - MHC-class-I expression in human breast cancer correlates with nuclear localization of the 90 kDa heat-shock-protein. AB - Breast cancer cells frequently exhibit a reduction in expression of major histocompatibility-complex (MHC) class I proteins which blocks cytotoxic T lymphocyte (CTL) mediated apoptosis. Recent studies indicate that the 90 kD heat shock-protein (HSP90) plays a major role in the transfer of antigenic peptides to the MHC class I complex. HSP90 is a molecular chaperone which is involved in signal transduction and regulation of apoptosis. Since HSP90 is described to be elevated in breast cancer, its relationship with MHC class I expression was investigated. Using immunohistochemistry we analyzed the expression and localization of HSP90 and MHC class I in 17 human breast tumors. Positive correlation (p < 0.025) between strong nuclear staining for HSP90 and high MHC class I expression was observed. In tumors with reduced MHC class I expression, no nuclear localization of HSP90 was detectable. These findings lead to the hypothesis that tumor cells with high MHC class I expression and susceptibility to CTL action may escape apoptosis by a mechanism which involves increased nuclear HSP90. PMID- 10697552 TI - Immunological properties of tumor cells genetically modified to secrete interleukin-2. AB - The immunological properties of interleukin-2 (IL-2) gene-transduced tumor cells were investigated in mice. A murine ovarian cancer cell line, OVHM, was retrovirally transduced with the human IL-2 gene (OVHM/IL-2) and the neomycin resistance gene (OVHM/Neo). OVHM/IL-2 cells continuously secreted IL-2 detected by ELISA using an antibody specific for human IL-2, and by a bioassay using an IL 2-reactive cell line (CTLL-2). When OVHM cells were inoculated subcutaneously into syngeneic B6C3F1 mice, OVHM/IL-2 cells but not parental (OVHM/P) or OVHM/Neo cells were regressed even though their rate of in vitro growth was comparable. This was not observed in nude mice, indicating the involvement of T lymphocytes in the regression of OVHM/IL-2 cells. The survival of mice inoculated intraperitoneally with OVHM/IL-2 cells was prolonged compared with those inoculated with OVHM/P cells. In irradiation experiments, IL-2 secretion by irradiated OVHM/IL-2 cells was retained for at least 5 days, although in vitro growth and in vivo tumorigenicity of irradiated OVHM/IL-2 cells were completely diminished. When mice were challenged with viable OVHM/P cells, survival of mice previously immunized with irradiated OVHM/IL-2 cells was prolonged compared to those immunized with irradiated OVHM/P cells, indicating a vaccine property of irradiated OVHM/IL-2 cells. Moreover, survival of mice with established ascites was improved upon injection of irradiated OVHM/IL-2 cells, indicating a therapeutic potential of OVHM/IL-2 cells. Tumor cells genetically engineered to secrete IL-2 may therefore be promising candidates for tumor vaccines and may provide a new mode of cancer immunotherapy. PMID- 10697553 TI - Radiation and hypothermia: changes in DNA supercoiling in human diploid fibroblasts. AB - The influence of hypothermia (2 degrees, 15 degrees and 28 degrees C) upon the effect of X-irradiation on chromatin from human diploid fibroblast cells (AG1518) was studied using the fluorescent halo assay. Rewinding of supercoils was inhibited in a dose-dependent manner when cells were irradiated with 4, 8 or 16 Gy. This inhibition of rewinding was reduced when cells were irradiated at subnormal temperatures compared with cells irradiated at 37 degrees C. One hour's preincubation at low temperature did not influence rewinding. When AG1518 cells were irradiated at 37 degrees C in the presence of the radical scavenger DMSO (0.5 M), the radiation-induced damage was reduced. No additional protection of DMSO in hypothermic cells (2 degrees C) was found, possibly indicating that OH radical-mediated effects are more temperature dependent. These results are similar to those recently found for the malignant MCF-7 cell line. PMID- 10697554 TI - Development of a syngenic brain-tumor model resistant to chloroethyl-nitrosourea using a methylguanine DNA methyltransferase cDNA. AB - Chloroethyl-nitrosourea (CENU) is one of the most potent chemotherapeutic agents for brain tumors. However, acquired resistance to this drug has become a serious problem in the treatment of brain tumor patients. The main mechanism of the resistance is a recruitment of the O6-methylguanine-DNA- methyltransferase (MGMT) in tumor cells. Many approaches, including treatment with enzyme-depletions, antibodies, antisenses, and a ribozyme, have been reported to successfully overcome the resistance. In order to evaluate these approaches properly, we designed a syngenic rat brain-tumor model resistant to CENU. The 9L rat gliosarcoma cells were retrovirally transduced with MGMT cDNA and stereotactically implanted into the brain parenchyma. In this model, rats inoculated with resistant cells died significantly earlier than did rats with control cells after treatment with CENU. Because of the limited intracranial space, the animals presented a restricted survival. Since the survival was sensitive and reproducible, this system may have a role in the evaluation of approaches to drug-resistant brain-tumors. PMID- 10697555 TI - Enhancement of mitomycin C efficiency by vitamin C, E-acetate and beta-carotene under irradiation. A study in vitro. AB - Using E.coli bacteria (AB 1157) and leukemia cells (HL 60) as a model for in vitro studies it was established that the efficiency of mitomycin C (MMC) can be influenced in the presence of antioxidant vitamins. This synergistic effect of the vitamins C, E-acetate and beta-carotene on MMC activity is rather strong for E.coli bacteria under irradiation (15 and 50 Gy) in the presence of air. Vitamin C contributes more efficiently to the MMC-activity in leukemia cells than the other two vitamins. The effect is explained by a cascade electron transfer process from the vitamins to MMC, where vitamin C is acting as a major electron source. These results might be of importance in cancer therapy. PMID- 10697556 TI - Measurement of mRNA expression for a variety of cytokines and its receptors in bone marrows of patients with myelodysplastic syndromes. AB - BACKGROUND: Myelodysplastic syndromes (MDS) are a group of disorders characterized by ineffective and dysplastic haemopoiesis. Previous studies in the lab have shown extensive apoptosis and high levels of transforming growth factor (TGF-beta) and tumor necrosis factor (TNF-alpha) in the stromal layer of MDS bone marrow. The current study focuses on the cytokines expressed in the bone marrow parenchymal cells. MATERIALS AND METHODS: Bone marrow aspirate from 5 normal donors and 26 patients with myelodysplastic syndromes were examined for mRNA expression of tumor necrosis factor alpha (TNF-alpha), macrophage colony stimulating factor (M-CSF), Flt-3 Ligand (Flt-3L), Flt-3 receptor(Flt-3 rec), interleukin 1 beta (IL 1 beta) and interleukin 1 receptor antagonist (IL-1 ra). RESULTS: Comparison of 26 MDS marrows with 5 normals showed a significantly higher value for Flt-3 rec and IL 1 beta (p = 0.031 and p = 0.031) in the former, while only Flt-1 beta rec was considerably higher (p = 0.016) in newly diagnosed patients. In previously diagnosed group, Flt-3 rec (p = 0.001), TNF-alpha (p = 0.04) and IL-1 beta (p = 0.016) were higher than normal while there was no statistically significant difference in the newly versus previously diagnosed MDS cases: CONCLUSION: mRNA expression of all six cytokines measured were considerably higher in MDS when compared to normal and that these levels tend to increase with disease duration. The precise source of these cytokines as well as their role in MDS pathogenesis remains to be determined, but this study confirms our previous reports that there is no dearth of cytokines in these bizarre myelosuppressive states. PMID- 10697557 TI - Effect of interleukin-2 on CD95 (Fas/APO-1) expression in fresh chronic lymphocytic leukaemia and mantle cell lymphoma cells: relationship to ex vivo chemoresponse. AB - Cells from B-cell chronic lymphocytic leukaemia (CLL) and mantle cell lymphoma (MCL) rarely express the CD95 (Fas/APO-1) antigen. Our aims were to determine whether CD95 levels (assessed by flow cytometry) in fresh neoplastic CD19+/CD5+ B lymphocytes from CLL and MCL could be upregulated using clinically relevant doses of interleukin-2 (IL-2), and to compare this with their ex vivo cytotoxic drug sensitivity (assessed by DiSC assay). CD95-expression was absent/negligible in 13/14 CLL and 7/7 MCL specimens. Following culture (without IL-2) CD95 was expressed on dying CD5+ B-lymphocytes but only on live cells in 2/15 cases. In live cells from 2 CLL specimens, IL-2 caused up-regulation of CD95 and was associated with ex vivo drug resistance. Clinically relevant doses of IL-2 had pleiotropic effects on CD95-levels in fresh CLL cells, but did not induce CD95 in MCL cells. The link between CD95-induction and ex vivo drug resistance may point to a clinically resistant subset of CLL patients. PMID- 10697558 TI - Targeting head and neck cancer by GM-CSF-mediated gene therapy in vitro. AB - BACKGROUND: The prognosis for patients with squamous cell carcinoma of the head and neck (SCCHN) has remained poor during the last decades, emphasizing the need for new treatment modalities. Consequently, the objective of our study was to evaluate the feasibility and efficacy of cytokine-mediated gene therapy in SCCHN in vitro. MATERIALS AND METHODS/RESULTS: The SCCHN cell line PCI-1 was transduced by lipofection with a plasmid encoding the human granulocytemacrophage colony stimulating factor (GM-CSF). Transfection of PCI-1 resulted in the production of significant amounts of GM-CSF as tested by ELISA. Enhanced proliferation of a GM CSF sensitive cell line, TF-1, after incubation with supernatants of GM-CSF transduced tumor cells demonstrated the release of biological active GM-CSF from these PCI-1 cells. In addition, GM-CSF-secreting PCI-1 cells enhanced antitumor cytotoxicity of allogeneic peripheral blood mononuclear cells, as tested in 24h MTT-cytotoxicity assays. CONCLUSIONS: Our data demonstrate the feasibility and efficacy of GM-CSF-mediated stimulation of the antitumor immune response against SCCHN in vitro and may help to define new strategies in the treatment of this malignancy. PMID- 10697559 TI - Confocal spectral imaging analysis of intracellular interactions of mitoxantrone at different phases of the cell cycle. AB - It is suggested that the cytotoxicity of anticancer agent mitoxantrone (MITOX) is related to a complex combination of molecular interactions which lead to slowing of S phase traverse and arresting of cells in G2 phase of the cell cycle or even to an apoptosis at high concentration of MITOX. Here intracellular molecular interactions of MITOX were visualised and studied using the confocal spectral imaging technique in synchronised K562 cells. Localisation, quantitative distributions of MITOX in the polar environment, MITOX bound to hydrophobic cellular structures (MITOXphob), nucleic acid-related complexes of MITOX (MITOXNA) and relative distributions of naphthoquinoxaline (NQX) metabolite and intrinsic cellular fluorescence of porphyrins were measured within cytoplasmic and nuclear compartments (chromosomes) of the G2, S, and M cells treated with 10 or 2 microM of MITOX for 1 hour. Colocalisation of MITOX, NQX metabolite and sites of intrinsic cellular fluorescence indicates an accumulation of MITOX within or near mitochondria. One may suppose that due to high concentration MITOX can compete with natural substrates for binding to the enzymes thus affecting the normal functioning of a mitochondria. A remarkable redistribution of MITOX and its complexes occurs in the M cells. In particular, a prominent amount of MITOX is associated with the surface of chromatids but not with the cytoplasmic structures in M cells. At the present time the exact location of the sites of MITOX accumulation in the M cells is not known. It is thought to be some cytoskeleton/microtubule structures associated directly with the chromosomes. Selective labelling of particular cytoskeleton structures and/or proteins in MITOX treated cells is in the progress now and the question will be addressed using the CSI technique. PMID- 10697560 TI - Cytotoxicity of L-cycloserine against human neuroblastoma and medulloblastoma cells is associated with the suppression of ganglioside expression. AB - BACKGROUND: Human neuroblastoma and medulloblastoma express abnormal ganglioside patterns especially GD2 and GM2 which are important for tumour growth. We tested the effects of L-cycloserine (L-CS), a potent inhibitor of synthesis of glycosphingolipids, on the growth, viability and expression of GD2 and GM2 in neuroblastoma and medulloblastoma cells. MATERIALS AND METHODS: The cytotoxic effects of L-CS were tested using the MTT dye reduction assay on four neuroblastoma (IMR-32, SK-N-SH, UKF-NB-2 and UKF-NB-3), two medulloblastoma (D283 and D341) and normal human fibroblasts and epithelial cell lines. In some experiments cytotoxicity of L-CS was tested in the presence of exogenous GD2 and GM2. The expression of GD2 and GM2 was analysed by flow cytometry. The antitumoral effects of L-CS in vivo were assessed on established xenografts of UKF-NB-3 or D283 cells in athymic (nude) mice using systemic administration of the drug (150 mg/kg intraperitoneally, once per day on 20 consecutive days). RESULTS: In vitro experiments revealed that L-CS was toxic for tumour cells at concentrations ranging from 0.5 to 20 micrograms/ml without any significant effects on normal fibroblasts and epithelial cells. L-CS treatment of UKF-NB-3 and D283 cells significantly inhibited expression of GD2 and GM2. The addition of exogenous GD2 and GM2 to culture medium partially prevented cytotoxic effects of L-CS on tumour cells. In vivo treatment resulted in complete tumour regression of UKF-NB-3 xenografts whereas growth of D283 xenografts was reduced by 60%. CONCLUSIONS: L-CS is a selective antitumoral agent for neuroblastoma and medulloblastoma cells with the ability to reduce expression of tumour associated gangliosides. In vivo experiments suggest that L-CS may be effective drug for treatment of neuroblastoma and medulloblastoma. PMID- 10697561 TI - Effect of extracellular NADH on human tumor cell proliferation. AB - We investigated the antiproliferative effects of extracellular nicotinamide adenine dinucleotide against human malignant CaCo-2 (colon carcinoma), Hep-2 (laryngeal carcinoma), MCF-7 (breast carcinoma), CaSki (cervix carcinoma) cell lines, as well as against murine fibrosarcoma and normal human embryonal fibroblast (HEF). NADH was very potent in the growth inhibition of murine fibrosarcoma and human Hep-2 cells, regardless of the dose applied. During the observed period (4 or 5 days) only one dose of NADH was sufficient in reducing the growth rate for up to 92%. It had no effect on the growth of other cell lines tested. The identification of DNA-fragmentation and p53 and Ki-67 genes expression suggest that the mechanism of NADH action is different from disregulation of genes considered as check-points in cell cycle. PMID- 10697562 TI - Subtractive hybridization and differential screening identified two genes differentially expressed after induction of in vitro (atypical) terminal differentiation in the NSCLC-N6 cell line by a marine substance (bistramide K). AB - Non-small-cell lung carcinoma is generally refractory to chemotherapy. The difficulties that arise in the treatment of this type of tumor make it necessary to develop new therapeutic strategies. Previous work done in our laboratory showed that a marine substance named bistramide K induced in vitro (atypical) terminal differentiation of NSCLC-N6 cell line. This activity is linked to a growth arrest of NSCLC-N6 cell line and an irreversible block at the G1 phase of the cell cycle (G1DT). In order to identify the genes that could be expressed after the treatment by the drug, we constructed a subtractive cDNA library with enriched mRNA extracted from BK-treated NSCLC-N6. After differential hybridization and DNA sequencing, we identified two sequences. The sequence identified for the clone 8 showed strong homology to the sequence of the ribosomal protein L35A. The sequence identified for the clone 4 did not show any homology with known sequences in official gene data banks. PMID- 10697563 TI - Allogeneic class I major histocompatibility complex gene transfer in murine neuroblastoma in vivo. AB - In vivo intratumoral gene transfer of allogeneic class I major histocompatibility complex (MHC) genes augments the immune response against weak tumor antigens. In this study, mice inoculated with the allogeneic MHC molecule (H-2Kb), had transduced-murine neuroblastoma C1300S3 cells showed prolonged survival relative to non-transduced or neo transduced tumors (p < 0.005). Interestingly, direct in vivo gene transfer of H-2Kb plasmid DNA complexed with HVJ-liposomes into S3 tumors was highly efficient, resulting in transduction of 8% of the interstitial cells within the tumor but rarely within tumor cells. Regression of established tumors and prolonged survival occurred in 50% of mice injected with H-2Kb, in contrast to no tumor regression in mice receiving control plasmid (p < 0.005). This study concludes that interstitial cells could serve as an important target of intratumoral gene transfer, and further that HVJ-liposome complexes could be a vehicle for in vivo gene transfer. PMID- 10697564 TI - Induction of apoptosis in human gastric adenocarcinoma cells by two novel organoselenium compounds, TS-2 and TS-6. AB - Two new organoselenium compounds, 4-ethyl-4-hydroxy-2-p-tolyl-4H-5,6-dihydro-1,3 selenazine (TS-2) and 4-hydroxy-4-methyl-6-propyl-2-p-tolyl-4H-5,6-dihydro-1,3 selenazine++ + (TS-6) were investigated for their inhibitory effect on the growth of 8 human tumor cell lines, including stomach, lung, prostate and colon cancer cell lines, in vitro. Both TS-2 and TS-6 exhibited the strongest cytotoxicity against a gastric adenocarcinoma (TMK-1) among 8 human tumor cell lines, and their IC50 were 2.38 microM and 2.78 microM, respectively. Twenty-four-h exposure of TMK-1 cells to TS-2 or TS-6 (0.1-100 microM) in the absence of serum led to a concentration-dependent increase of cytotoxicity. Exposure of TMK-1 cells to TS-2 or TS-6 (5, 10 or 20 microM) in the presence of 5% serum resulted in significant inhibition of TMK-1 cell proliferation in a concentration- and time-dependent manner. Morphological changes including shrinkage of the nucleus, and DNA ladder fragmentation, which are considered to be the typical features of apoptosis, were observed in TMK-1 cells in response to TS-2 or TS-6. Furthermore, the caspase-3 activity in TMK-1 cells treated with TS-2 or TS-6 was also found to be increased in a time-dependent manner, in parallel with the induction of DNA fragmentation. Taken together, the results of the present study suggest that the inhibition of growth of TMK-1 cells by TS-2 and TS-6 is mediated by the induction of apoptosis. PMID- 10697565 TI - Effect of glutathione-modulating compounds on platinum compounds-induced cytotoxicity in human glioma cell lines. AB - The relation between the effect of glutathione(GSH)-modulating compounds and platinum compounds (Cisplatin, Nedaplatin)-induced cytotoxicity was investigated. Pretreatment of human glioblastoma (T98G, U87MG) and glioma (KG1C) cell lines with L-buthionine-[S,R]-sulfoximine, which decrease the intracellular GSH concentration, remarkably increased their sensitivity against platinum compounds, whereas pretreatment with N-acetyl-L-cysteine, which increase the intracellular GSH concentration, only marginally protected the cells from the cytotoxic effect of platinum compounds. The results suggest that platinum compounds-induced cytotoxicity can be modified by GSH-modulating compounds in glioblastoma and glioma cell lines. PMID- 10697566 TI - Tumor histopathology following new sensitizers: dithiaporphyrin- and sulfoxaporphyrin-mediated photodynamic therapy. AB - BACKGROUND: Our main aim was to evaluate tumor histopathology following new sensitizer-mediated photodynamic therapy (PDT). MATERIALS AND METHODS: In order to complete our studies we decided to use photosensitizers, i.e. dithiaporphyrin (DTP) and sulfoxaporphyrin (OXA) in combination with halogen lamp irradiation of presensitized tumors. The doses of sensitizers were: 2.5, 5.0, 7.5 and 10.0 mg/kg of body weight and total light doses were: 50, 100 and 150 J/sq.cm at the selected wavelength. Following such a treatment we have evaluated tumor necrosis of BFS1 fibrosarcoma growing on BALB/c mice. Together with tumor necrosis evaluation we have examined skin response to photodynamic treatment. RESULTS: We have found that both new sensitizers caused significant tumor damage at no skin alterations. The induction of tumor necrosis seemed to be dose dependent, i.e. higher photodynamic doses (sensitizer dose x light dose) resulted in more severe damage to the tumors than the lower doses. CONCLUSION: Our study showed that BFS1 fibrosarcoma is highly sensitive to PDT after application of new sensitizers. Both compounds can be considered as potent tumor photosensitizers in future clinical trials. PMID- 10697567 TI - Expression and relationship between topoisomerase I and II alpha genes in tumor and normal tissues in esophageal, gastric and colon cancers. AB - DNA topoisomerases are known to be nuclear enzymes that are important targets of topoisomerase I (topo I) and topoisomerase II (topo II) inhibitors in cancer chemotherapy. We investigated the mRNA expression of topo I and topo II alpha genes as assessed by northern blot analysis in tumor and the adjacent normal tissues of esophageal, gastric and colon cancers. The surgical specimens consisted of 18 tumor tissues and the adjacent normal tissues including 6 esophageal cancers, 6 gastric cancers and 6 colon cancers. We found that the mRNA expression of topo I gene was not significantly different between tumor and normal tissues in 18 surgical specimens, whereas the mRNA expression of topo II alpha gene in the all types of tumors was significantly higher than that of the adjacent normal tissues. Furthermore, the mRNA expression of topo II alpha gene in tumor and adjacent normal tissues was correlated with the S-phase population in cell cycle. Of great importance was the significant relationship between mRNA expression of topo I and topo II alpha genes in tumor and normal tissues was found in esophageal and colon cancers (p < 0.05), except in gastric cancers. These results indicate that the rationale in tumor specific chemotherapy with topo II inhibitors was based on the finding of its higher expression of topo II alpha gene in tumors than that of normal tissues, an important target of topo II inhibitors, and suggest that the sequential chemotherapy targeting topo I and topo II enzymes by modulating topo II alpha expression by topo I inhibitors might be more effective in esophageal and colon cancers, in terms of their relationship between topo I and topo II alpha expression in tumor cells. PMID- 10697568 TI - Taxotere activates transcription factor AP-1 in association with apoptotic cell death in gastric cancer cell lines. AB - We investigated whether a novel mitotic inhibitor, taxotere can activate the transcription factor AP-1 in association with apoptotic cell death in 8 gastric cancer cell lines. Apoptotic cell death was analyzed by DNA ladder formation assay, and AP-1 binding activity was assessed by gel mobility-shift assay. The activation of AP-1 binding was induced in accordance with the sensitivity to taxotere in gastric cancer cell lines. The relationship between the increase of AP-1 binding and the formation of internucleosomal DNA ladders induced by taxotere was significant (p < 0.05). Furthermore, the activation of AP-1 binding was induced following the treatment of taxotere in a dose and -time dependent manner. Although the sensitivity to taxotere was correlated with the formation of internucleosomal DNA ladders in apoptotic cell death, its sensitivity was not influenced by their p53 genomic states in gastric cancer cell lines. Rather, the activation of AP-1 binding was correlated with the induction of a growth arrest and DNA damage inducible gene, gadd153 (p < 0.05). These results indicate that the activation of AP-1 binding by taxotere seems to be an important factor in determining its sensitivity in association with internucleosomal DNA ladders, and suggest that the induction of gadd153 gene could be a downstream target of AP-1 regulated genes involved in signal transduction pathways leading to apoptosis in gastric cancer cells. PMID- 10697569 TI - The effect of red wine and its components on growth and proliferation of human oral squamous carcinoma cells. AB - Epidemiologic evidence indicates that red wine may contain phenolic compounds which protect against heart disease and cancer. Resveratrol and quercetin are wine phenolics which possess antioxidant and antimutagenic effects. Resveratrol at 10 and 100 microM induced significant dose-dependent inhibition in human oral squamous carcinoma cell (SCC-25) growth and DNA synthesis. Quercetin exhibited a biphasic effect, stimulation at 1.0 and 10 microM and minimal inhibition at 100 microM in cell growth and DNA synthesis. Combining 50 microM of resveratrol with 10, 25 and 50 microM of quercetin resulted in gradual and significant increase in the inhibitory effect of the two compounds. Diluted red wine which contained only 1.6 microM of each of resveratrol and quercetin had significantly more inhibitory effect on cell growth, DNA synthesis and changes in cell morphology than each compound alone or in combination. We conclude that: (i) Resveratrol by itself or a combination of resveratrol and quercetin are effective inhibitors of SCC-25 growth and DNA synthesis. (ii) The presence of other wine phenolic phytochemicals enhance significantly the effect of resveratrol and quercetin on inhibition of cancer cell growth and DNA synthesis. PMID- 10697570 TI - Expression of p53 and pRb in bladder and prostate cancers of patients having both cancers. AB - Evidence has been presented that tumor suppressor genes p53 and Rb play a crucial role in the development of both human prostate and bladder cancer. Patients with either cancer are at an increased risk for developing the other malignancy as compared to the general population. The purpose of the present study was to investigate whether there is abnormal expression of these two suppressor proteins in both the bladder and prostate cancers of the same patient. The expression of p53 and pRb in bladder and prostate cancer specimens obtained from 15 patients having both cancers was studied using immunohistochemical staining with antibodies against these proteins. The expression of p53 and pRb in both bladder and prostate cancers of the same patient was congruent in 8 of 15 cases (53%) for p53 and 9 of 15 cases (60%) for pRb. The significance of these findings warrants further investigations. PMID- 10697571 TI - Transcription of 2'-deoxy-2'-fluoro-modified and phosphorothioate-modified RNA templates by HIV-1 reverse transcriptase. AB - RNA templates yield the corresponding DNA in the presence of human immunodeficiency virus reverse transcriptase (HIV-1 RT). The purpose of this study was to determine whether RNA that was modified with either 2'-deoxy-2' fluoro analogs or with internucleotide phosphorothioate linkages could serve as templates for HIV-1 RT. Modified RNA that contained either 2'-deoxy 2'-fluoro pyrimidine nucleoside analogs or internucleotide phosphorothioate diester linkages 5'- to pyrimidine nucleosides were enzymatically synthesized and tested for template activity with recombinant HIV-1 RT. RNA that was modified with either 2'-deoxy-2'-fluorouridine or with internucleotide phosphorothioate linkages 5'- to pyrimidines yielded full length HIV-1 reverse transcription products, with complete fidelity in transcription. RNA that was modified with 2' deoxy-2'-fluorocytidine, either alone or in combination with 2'-deoxy-2' fluorouridine, did not function as templates for HIV-1 RT, under the conditions reported here. The ability of 2'-deoxy-2'-fluoro-modified and phosphorothioate modified RNA to serve as template for the RNA-dependent DNA polymerase of HIV-1 RT has not hitherto been reported. PMID- 10697572 TI - Synergistic effect of gemcitabine and irinotecan (CPT-11) on breast and small cell lung cancer cell lines. AB - Gemcitabine (2'-2'-difluorodeoxycytidine, dFdC), an analog of deoxycytidine, is an antineoplastic agent with clinical activity against several types of cancer. Irinotecan (CPT-11), a topoisomerase I inhibitor, is a drug with a broad spectrum of anticancer activity. Since these drugs have different mechanisms of cytotoxicity and dose-limiting toxicity profiles, preclinical combination studies were performed on the MCF-7 breast cancer and the SCOG small cell lung cancer (SCLC) cell lines. Both gemcitabine and CPT-11 as single agents were effective growth inhibitors in these cell lines. Isobologram analysis revealed for the first time that the combination of these drugs exerted synergy over a wide range of concentrations in MCF-7 and SCOG cells. Moreover, combination index (CI) analysis revealed that at low concentrations, combinations of gemcitabine and CPT 11 show a synergistic growth inhibitory effect on MCF-7 cells. However, in SCOG cells CI analysis showed synergy at concentrations of gemcitabine and CPT-11 greater than 1 microM but antagonism at combination concentrations less than 1 microM. These preclinical cytotoxicity data provide an experimental basis for conducting clinical trials using combinations of gemcitabine and CPT-11, especially in patients with breast and lung cancers. PMID- 10697573 TI - Expression of Po66-CBP, a type-8 galectin, in different healthy, tumoral and peritumoral tissues. AB - A monoclonal antibody, Po66, recognized an antigen named Po66 carbohydrate binding protein (PO66-CBP), which was homologous to the galectin-8 protein. Two additional isoforms previously not described were characterized. The aim of this study was to compare the expression of Po66-CBP and its isoforms in different healthy, tumoral and peritumoral tissues and at last to determine the localization of the protein in tumors and distant tissues. MATERIALS AND METHODS: Reverse transcriptase PCR of Po66-CBP was performed on total RNA extract from eleven healthy and eleven tumoral and peritumoral tissue specimens. Antibody Po66 was used to localize the protein in the tumors and distant tissues by an immunohistochemistry method. RESULTS: Po66-CBP was expressed by half of the healthy tissues. One of the isoforms, the last often present in healthy tissues, was found in all tumoral and peritumoral tissues studied. Immunohistochemistry evidenced a gradient of protein expression in normal cells depending on the vicinity of tumoral tissue. Po66-CBP expression was modified in cancerous tissue, suggesting the implication of galectins in carcinogenesis. PMID- 10697574 TI - The induction of apoptosis in human melanoma, breast and ovarian cancer cell lines using an essential oil extract from the conifer Tetraclinis articulata. AB - The cytotoxic effect of conifer Tetraclinis articulata essential oil (TAEO) on a number of human cancer cell lines and peripheral blood lymphocytes was assessed at various concentrations and time exposures. The cytotoxic effect showed the hallmarks of apoptosis confirmed by a variety of techniques including flow cytometry, an apoptosis- specific marker combined to fluorescent staining and DNA laddering. All cell lines tested were inhibited in a dose-dependent fashion and within a contact time of less than eight hours for the higher concentrations. Melanoma, breast and ovarian cancer cells gave IC50s of around 80 micrograms/ml whilst the IC50s on peripheral blood lymphocytes was almost double this value. We conclude that the essential oil contains components that are effective at inducing apoptosis. The advantages of using a mixture of monoterpenes (C10) as present in an EO over a single component, are discussed. PMID- 10697575 TI - Bcl-2 and Bax proteins are nuclear matrix associated proteins. AB - Bcl-2 and Bax proteins are implicated in the regulation of apoptosis. Nuclear matrix has been demonstrated to be associated with a vast array of functional and regulatory properties of cells. NuMA is one member of a class of nuclear matrix proteins that resides in both the nucleus and mitotic apparatus. The nuclear lamins appear to form a thin fibrous structure immediately underlying the inner nuclear membrane of eukaryotic cell nuclei. The association of bcl-2 and Bax protein with nuclear matrix in glioblastoma cell line U343 was studied by confocal microscopy and Western blotting. Confocal microscopic images display that bcl-2 was localized at the peripheral of the nuclear matrix and Bax protein was located in the nuclear matrix. Western blotting detected a 26 kDa bcl-2 band and a specific band of Bax at around 66 kDa in nuclear matrix proteins. Our results suggest that bcl-2 and Bax proteins are nuclear matrix associated proteins. PMID- 10697576 TI - In vitro antitumor effect of topoisomerase-I inhibitor, CPT-11, on freshly isolated human gastric and colorectal cancer. AB - CPT-11 is a comptothecin analogue which has shown a broad spectrum of strong antitumor effect against various cancers, including gastroenterological malignancies. In the present study, the antitumor effect of CPT-11 was determined by MTT assay for freshly isolated human gastric and colorectal cancer cells, especially highly purified tumor cells. Twenty-three patients with gastric cancer, and 32 patients with colorectal cancer were enrolled in this study. Three gastric and 3 colonic cancer cell lines were used to study the antitumor effect of CPT-11, and freshly isolated cancer cells from 3 patients with gastric cancer were investigated. The in vitro antitumor effect was tested by MTT assay, and showed % inhibition rate. CPT-11 and SN-38 showed the antitumor effect as a dose dependent matter for human gastric and colorectal tumor cells in vitro. From the results of chemosensitivity for freshly isolated gastric and colorectal tumor cells, antitumor effect of SN-38 was as strong as other conventional anticancer agents. It was demonstrated that the MTT assay was appropriate for the analysis of the antitumor effects of CPT-11 and SN-38, and that CPT-11 may be a worthwhile choice as an anticancer agent against gastric and colorectal cancer. PMID- 10697577 TI - Interaction between antioxidants and hydroquinone/bisphenol. AB - We have investigated the stability of semiquinone radical (SQ .) produced from the mixture of hydroquinone (HQ) and bisphenol A (BPA) in the presence of various antioxidants, some of which induce apoptosis in tumor cell lines. BPA produced no detectable amount of ESR signal. The acceleration of HQ oxidation by BPA results in the production of SQ .. The BPA-enhanced SQ . production may occur through either SQ. regeneration by electron transfer from BPA working as an antioxidant or the O2- scavenging activity of BPA. SQ . intensity of HQ/BPA was reduced by lower concentrations of sodium ascorbate, epigallocatechin, gallate and quercetin, but significantly enhanced by gallic acid. The initial SQ . was replaced by the radicals of antioxidants except lignin and salicylic acid, at higher concentrations, due to their radical-radical coupling or dismutation process. Among these compounds, gallic acid enhanced the radical intensity of SQ . to the greatest extent. The results suggest that antioxidants from foods suppress the activity of xenobiotics such as HQ, however, they produce their own radical at higher concentrations, suggesting their bimodal actions. The present study demonstrates the usefulness of ESR spectroscopy for the interaction between HQ/BPA and antioxidants. PMID- 10697578 TI - bcl-2 protein expression in cervical intraepithelial neoplasia: no evidence of a prognostic significance in mild and moderate lesions. AB - BACKGROUND: The bcl-2 proto-oncogene codes for a protein which appears to block apoptosis. In our study, we examined bcl-2 protein expression in cervical squamous metaplasia, cervical intraepithelial neoplasia (CIN) and microinvasive squamous carcinoma with the aim of identifying a relationship between bcl-2 protein expression and neoplastic development and progression. MATERIALS AND METHODS: Cervical bioptic samples were obtained from 86 white women, selected consecutively from our Colposcopic Service from January 1993 to June 1994, because of abnormal pap- smear suspicious for cervical dysplasia and/or human papilloma virus (HPV) infection. Upon histologic evaluation, 41 women had CIN, 23 cervical condyloma, and 22 squamous metaplasia. Ten patients with microinvasive squamous carcinoma, matched for age and demographic characteristics, were selected from our series of invasive cervical carcinomas and immunohistochemically analyzed. The expression of primary tumor bcl-2 protein was immunohistochemically evaluated by antihuman bcl-2 monoclonal antibody (diluted 1:100, Dako, Copenhagen, Denmark) on formalin-fixed paraffin-embedded tissue. Positive staining was expressed as a percentage of positive cells per 1000 counted dysplastic cells for each case. RESULTS: Bcl-2 immunostaining was found in all the 22 squamous metaplasias, limited to the basal layer. Nineteen of the 41 CINs (46%) were bcl-2 immunoreactive, and 2 of the 10 microinvasive carcinomas (20%). By analysing CIN lesions, the bcl-2 protein showed a striking increase in the rate of positivity with increasing severity of CIN; the bcl-2 protein expression in CINs III was significantly higher than for CINs I, CINs II or microinvasive carcinomas (P = 0.03, P = 0.02, and P = 0.03 respectively). No relationship was observed between bcl-2 immunostaining and HPV infection. bcl-2 protein expression was not useful for predicting CIN I and II evolution, although the rate of persistence/progression was higher in bcl-2 positive dysplasias (7 of 9 cases, 78%) than in negative ones (13 of 21 cases, 62%) (p = 0.88). CONCLUSIONS: Based on these results, it seems possible that the increase in bcl-2 expression in higher grade of CINs implies an increasing protection against programmed cell death, but also the induction of genetic instability in dysplastic epithelial cells and a greater capacity to evolve into invasive carcinoma. PMID- 10697579 TI - Effect of vitamins A, C and E on normal and HPV-immortalized human oral epithelial cells in culture. AB - There is experimental and epidemiological evidence that antioxidant vitamins can inhibit carcinogenesis. Since immortalization by Human Papilloma Virus (HPV) is one possible early step towards carcinogenesis in oral epithelia, we studied the differential effect of vitamins A, C and E on HPV-immortalized oral epithelial cells (IHGK) as compared to the normal counterpart. The dose response was determined by morphology, cell cycle by flow cytometry, and growth curve by cell number. The optimum dose in terms of inhibitory effect vs. toxicity was determined for each vitamin by morphology. Optimum doses were: vitamin A--1.4 x 10(-5) M, vitamin C--10(-3) M, and vitamin E--10(-6) M for both HPV-immortalized and normal cells. Growth curve showed reduction of proliferation by all three vitamins, with vitamins A and E more effective than C for both cell types. Flow cytometry showed that vitamins A and E reduced the percentage of cells at G2 phase of cell cycle and indicated arrest in the S phase. This effect was greatest in the immortalized cells with a 50% and 35% decrease of G2 for vitamins A and E respectively, whereas the normal counterpart showed a 48% decrease for A and a 12% increase for E. By organotypic culture, the morphology was not markedly different between the vitamin-treated and the control cells, except for a slight increase in the keratinization of normal cells with vitamin A. Also noted was a reduction in number of cell layers from five layers or more for controls to only one or two for vitamin E. In conclusion, we have demonstrated that the antioxidant vitamins inhibit proliferation, and show a preferential effect on IHGK cells. PMID- 10697580 TI - What is cancer? Theories on carcinogenesis. PMID- 10697581 TI - Toward a philosophy of cancer research. AB - Every theory and all research assumes, either explicitly or implicitly, a scientific philosophy. Making the philosophy of cancer research explicit is likely to improve our understanding both of what we know and what we still need to discover. The implications for cancer research of three different philosophies of science are therefore outlined here: 1) reduction-mechanism; 2) holism; and 3) complementarity. We show that each of these philosophies leads to a different notion of causation, a different expectation of what represents a valid explanation of cancer, and thus to different problems that are addressed by different types of experiments. We conclude that the development of an appropriate philosophy of science is not only a relevant but necessary element in research on carcinogenesis. PMID- 10697582 TI - History of cancer research in nosological perspective. AB - Contrary to conventional wisdom progress in cancer research has not followed a linear course, but rather a torturous capricious path, often guided by the prevailing concepts of disease (nosology). Various theories appear, disappear and reappear all the time. Periods with rapid progression alternate with periods of stagnation. Really new ideas emerge rarely in oncology. Most of the so-called new ideas are modifications of older concepts i.e. old wine in new skins. Moreover existing oncological theories brought under the umbrella of a new nosological paradigm may change drastically and often get a totally new meaning. As a consequence older theories may linger on over centuries but may be modified to such an extent that the original concepts are barely recognisable. Progress in oncology is characterised by periods of rapid progress alternating with periods of apparent stagnation which with hindsight were nevertheless periods in which new concepts were in statu nascendi. The history of oncology provides still a source of inspiration, since older ideas may be worth reconsidering. PMID- 10697583 TI - Morphology of human carcinogenesis. AB - Carcinogenesis, the process by which normal cells become neoplastic, has been extensively studied in tissue cultures and animal models. Screening and periodic examination of various organs with the use of modern technology provides the opportunity of directly studying human carcinogenesis. Following is the description of morphologic changes accompanying the stages of carcinogenesis in the most common human tumors, the lung, breast, uterine cervix, colon/rectum, and urinary bladder. PMID- 10697584 TI - The role of genomic instability in human carcinogenesis. AB - Neoplastic cells typically possess numerous genomic lesions, which may include sequence alterations (point mutations, small deletions, and insertions) and/or gross structural abnormalities in one or more chromosomes (large-scale deletions, rearrangements, gene amplifications). Based upon this general observation, it has been suggested that cancer cells are genetically unstable, and that acquisition of genomic instability may represent an early step in the process of carcinogenesis and a general feature of many human tumors. Numerous studies have appeared that characterize the nature and frequency of occurrence of various molecular lesions in human tumors, and significant progress has been made towards the elucidation of the molecular mechanisms that govern genetic stability in normal cells and genetic instability in neoplastic cells. In this review, we examine the evidence that genomic instability plays a significant role in the genesis of various human tumors. Furthermore, we consider the possible molecular pathways to tumorigenesis in humans and how different forms of genetic instability may impact upon these pathways. PMID- 10697585 TI - Genomic instability: first step to carcinogenesis. AB - Multiple genetic alterations are commonly observed in human cancers. It has been suggested that inactivation of DNA repair pathways, which leads to an increased mutation rate and chromosomal instability, can initiate and accelerate the neoplastic process. Such a causality has been shown for DNA mismatch repair and Hereditary Nonpolyposis Colorectal Cancer (HNPCC), and evidence is accumulating that several other DNA repair pathways are frequently inactivated in different cancer types. In addition to genetic alterations, perturbations in DNA methylation patterns (epigenetic changes), which include both local hypermethylation and genome-wide hypomethylation, are frequently observed early in tumorigenesis. Therefore, genomic instability including genetic and/or epigenetic alterations may be the first step in carcinogenesis. Knowledge of these biochemical mechanisms are likely to lead to more effective cancer diagnosis and therapy. PMID- 10697586 TI - Chromosomal abnormalities: detection and implications for cancer development. AB - The occurrence of chromosomal abnormalities is a common theme in carcinogenesis. A large proportion of tumours which have been characterized at the cytogenetic level carries numerical and/or structural aberrations. Numerical alterations may include acquisition or loss of specific chromosomes or dramatic changes in overall ploidy levels. Structural aberrations may include DNA amplifications or deletions, inversions and translocations. Many chromosomal alterations occur in a non-random fashion and may be subdivided in to primary and secondary, according to their timing of occurrence. Primary chromosomal abnormalities usually occur at the early stages of tumourigenesis and are often encountered as sole cytogenetic abnormalities. Secondary chromosomal abnormalities are usually associated with more advanced stages of tumour development. In recent years several chromosomal abnormalities could be correlated with specific gene alterations, thus providing insights into the molecular mechanisms underlying tumourigenesis. The biological consequences imparted by other chromosomal changes such as numerical changes are, however, less clear. By using recently developed molecular techniques for chromosome characterization, so-called molecular cytogenetics, our perception on cancer cytogenetics is rapidly changing through the disclosure of hitherto unknown (specific) chromosomal abnormalities. PMID- 10697587 TI - Tumor suppressor genes (TSG). AB - In the last two decades the field of tumor suppressor genes (TSG) has developed from a novel epidemiology-based concept into one of the pillars of cancer research. New TSG are being discovered and although at a painstakingly slower pace, their functions are also being unravelled. These breakthroughs have not only brought about immediate benefits to clinical medicine such as genetic predictive testing for familial cancer syndromes, but have also introduced new strategies against cancer including early intervention, prevention and promising anti-cancer drugs. Despite that, we are still far from fully understanding the functional complexity of TSG with each discovery leading to even more queries. New developments that challenge the paradigm of TSG have also surfaced warranting verification and there is still a lack of consensual criteria and systematic cataloging for TSG. With the advent of high throughput sequencing and powerful bio-informatics, it is expected that the field of TSG will be expanding at an even faster pace. This review is to give a summary of the history, new concepts and recent advancements in the field of TSG in the hope that some of these concepts and works can be evaluated, expanded or even challenged. PMID- 10697588 TI - Oncogenes. AB - Oncogenes are gain-of-function mutations of normal regulatory genes or proto oncogenes. Originally discovered in retroviruses initiating a variety of animal and avian cancers, oncogenes are believed to be important contributors to human carcinogenesis. Proto-oncogenes are altered by point mutation, amplification or rearrangement. Structural alteration of proto-oncogenes leads to a quantitative or qualitative expression change of the corresponding protein product. Oncoproteins subvert signal transduction pathways at the cell surface, in the cytosol and/or in the nucleus. Together with other oncoproteins or in the absence of tumor suppressor gene products, oncogenes contribute to human cancer formation by supporting accelerated proliferation, de-regulating cell cycle control or blocking apoptosis. PMID- 10697589 TI - Human tumor viruses. AB - Tumor viruses play an important role for the development of a substantial fraction of human malignancies, including common cancers, such as carcinomas of the cervix uteri, hepatocellular carcinomas, or lymphomas. In the recent past, much progress has been made in elucidating the molecular mechanisms by which human tumor viruses contribute to cellular growth deregulation and carcinogenesis. The picture emerges that different tumor viruses target similar cellular pathways for growth deregulation but, in addition, also have unique properties contributing to oncogenesis. Malignant transformation typically requires additional genetic alterations of the host cell, to which tumor viruses can contribute by destabilizating the cellular genome. PMID- 10697590 TI - Role of p53 and apoptosis in carcinogenesis. AB - Carcinogenesis is a process that converts normal cells from controllable to uncontrollable growth. Coordinate regulation of the rates of cell proliferation and cell death is an important determinant in maintenance of homeostasis. Loss of control of this balance is central to the development of cancer. This loss may be due to genetic alteration in either growth promoting genes resulting in constitutive activation or negative growth regulating genes such as tumor suppressor genes. Recent advances in studying the molecular mechanisms related to the etiology of cancer have provided further understanding of these pathways. Earlier studies have been primarily concerned with cell proliferation resulting from activation of oncogenes. However, many recent studies have focused on the induction of cell death. The recognition of the importance of apoptosis, a distinct mode of cell death, in maintenance of genomic stability was further prompted by studying the mechanism of the tumor suppressor gene product p53, as well as many other oncogenes and tumor suppressor genes. For example, many oncogenes appear to act as potent inducers of apoptosis through activation of p53 dependent apoptosis pathways. Therefore, one possible mechanism for tumor suppression involves activation of apoptosis pathways in cells at risk of neoplastic transformation. These studies have provided extensive knowledge of the signal transduction pathways in response to genotoxic stress and promoted mechanistic research related to the apoptosis pathways. These studies also provide a perfect explanation that p53 is a key element in maintaining genomic stability and loss of the p53 function is a common event during carcinogenesis. This chapter will mainly focus on the role of apoptosis in carcinogenesis. In particular, I will summarize recent studies related to the mechanisms of p53 and its role in this process. PMID- 10697591 TI - Cancer cell cycle control. AB - The cell cycle is controlled by multiple mechanisms on which exogenous and endogenous stimuli converge. The pathways governing the different cell cycle phases are central for the cells' decisions when to commit to DNA synthesis and proliferation versus growth arrest, DNA repair or apoptosis. Consequently, these pathways incorporate various oncogenes and tumor suppressors and are therefore a central target for genetic alterations in human cancers. These events may ultimately lead to aberrant cell proliferation and increased genetic instability. Unraveling these regulatory networks provides an important insight into the balance of normal and cancerous cell proliferation and is pivotal for the design of novel anticancer strategies. PMID- 10697592 TI - Genetic, epigenetic, dysgenetic and non-genetic mechanisms in tumorigenesis. II. Further delineation of the rate limiting step. AB - A not yet understood phenomenon in carcinogenesis is presented by the enormous species-specific and age-specific differences in cellular susceptibility to malignant transformation. Murine cells are 100,000 times fold more susceptible than human cells. A previous review article suggested the promotion phase as the key for this difference. Presently, a further analysis of literature data indicates that promotion results in the dedifferentiation of cells, which, in initiated cells, leads to an imbalanced regenerative growth of stem-cell-like cells. As aging results in the loss of adaptive functions and in a decreased response to growth stimuli, dedifferentiation of old cells is supposed to increase the imbalance of the regenerative growth for initiated cells and consequently to enhance the chance for immortalization of these cells. As immortalization is accompanied by chromosomal dysgenetics, malignant conversion by the appearance of secondary genetic events during multistage carcinogenesis is not just a matter of random chance for additional mutations, but is determined to some degree by the condition of the initiated target cell and to the species to which the target cell belongs. PMID- 10697593 TI - Growth regulatory factors and carcinogenesis: the roles played by transforming growth factor beta, its receptors and signaling pathways. AB - It is widely recognized that growth factors play critical roles in cell proliferation and differentiation. In the early 1980s, several members of the transforming growth factor-beta (TGF-beta) superfamily were identified and subsequently shown to play important roles in many diseases, in particular cancer. Efforts to understand how TGF-beta exerts its effects led to identification of TGF-beta-receptors and several downstream signaling pathways activated by this family of growth factors. This review provides an overview of TGF-beta-receptors and its downstream signaling pathways. As part of this discussion, this review indicates that inactivation of TGF-beta-receptors or components of their signaling pathways is often a target during carcinogenesis and that mutations or altered expression at any step along this complex, growth regulatory pathway can lead to aberrant cell proliferation. Lastly, the Cancer Genome Anatomy Project is briefly discussed, in particular how it may help to identify aberrant growth factor expression during carcinogenesis and improve the diagnosis of cancer patients. PMID- 10697594 TI - Tumor stroma. AB - The accumulated evidence indicates that tumor stroma with its cells and cell products plays a much more active and important role than previously believed. Growth factors and cytokines produced by macrophages and other cells are crucial for stroma formation and angiogenesis. Lytic enzymes provided by stromal cells may be essential for invasion. TNF and other inflammatory mediators may be operative in the systemic effects of tumors, e.g. cachexia. All these effects may come about through the action of soluble substances produced by tumor cells or by more intimate interactions. There is no evidence that stromal cells are directly involved in carcinogenesis--i.e. the cellular transformation to produce the malignant cell. On the other hand, stromal cells and other components of the interstitia are instrumental in tumorigenesis--i.e. the development of a real malignant tumor from its start on the cellular or subcellular level. In one way of looking at it, the stromal cells, e.g. macrophages may be considered as "slaves", kept to carry out certain functions, synthesize essential substances e.g. growth factors that the tumor cells do not have the capacity or the degree of finely tuned machinery to produce. The objective of immunomodulation should then be to create a "slave uprising", to make the macrophages and other cells turn against their masters, stop producing growth factors and start producing harmful factors that would lead to the elimination of the malignant growth. The first target of immunomodulation in tumor disease should probably be local malignancies where no effective treatment exists today- and selected cases of metastatic prevention (181, 182). PMID- 10697595 TI - The roles of telomeres and telomerase in cellular immortalization and the development of cancer. AB - Normal human cells have a limited lifespan in culture called the Hayflick limit. Recent studies have indicated that telomere shortening is one of the important meters utilized by cells to determine the Hayflick limit, and that activation of a mechanism to maintain telomere length is essential for cells to become immortal. It is generally believed that cells must have a means to maintain telomeres in order to progress to malignancy. Most cancers do this by activating an enzyme called telomerase which adds telomeric repeats to the telomere ends. Recently, expression of this enzyme has been shown to extend the lifespan of cells. This review discusses the research that led to the discovery of telomerase, the characteristics of telomerase complex, and how recent and future advances in the telomerase field may lead to better diagnostic and treatment protocols for many different cancer types. PMID- 10697596 TI - The role of oval cells and gap junctional intercellular communication in hepatocarcinogenesis. AB - The role of oval cells, and Gap Junctional Intercellular Communication (GJIC) in hepatic differentiation and neoplasia is controversial. Oval cells accumulate in great number when hepatocyte regeneration is blocked following massive hepatotoxicity or after treatment with some hepatocarcinogens. This suggests oval cells are facultative stem cells or close progeny of liver stem cells that are activated only under specific conditions. Studies with oval cell lines clearly indicate that they can differentiate into hepatocytes and that neoplastic derivatives of oval cells can produce hepatocellular and biliary neoplasms. Because hepatocytes express Cx32 and biliary cells express Cx43, the differentiation of oval cells into hepatocytes or In addition, because Cx32 hemichannels and Cx43 hemichannels cannot form heterotypic patent channels, the type of connexin expressed by the differentiating oval cell will determine whether it communicates with hepatocytes or biliary epithelial cells, respectively. This communication may be necessary for the further differentiation and regulated growth of the differentiating oval cells and impairment of this GJIC may contribute to the formation of hepatocellular and cholangiocellular neoplasms. The type of connexin expressed may also determine the susceptibility of the differentiating oval cells to the various types of rodent liver tumor promoters. Thus, three major points have been developed here. First, Cx32 or Cx43 expression and GJIC with hepatocytes or biliary epithelial cells, respectively, may determine the final differentiated fate of oval cells. Secondly, blocked GJIC may determine whether oval cells progress to hepatocellular or cholangiocellular carcinoma. Lastly, the ability of tumor promoters to block Cx32 or Cx43-mediated GJIC in differentiating oval cells may determine whether these agents promote the formation of hepatocellular or cholangiocellular carcinomas. Thus, GJIC may be the key factor in the differentiation of oval cells and blocked GJIC may promote their neoplastic transformation in a lineage-specific manner. In this chapter, we have outlined several new hypotheses on the role of oval cells and GJIC in hepatocarcinogenesis. We hope that other investigators will consider our ideas, but realize these views will be contentious to many. Our intent, however, was to stimulate discussion and debate, even argument, because truth often arises amidst controversy and may be found in the most peculiar places. PMID- 10697597 TI - Spontaneous remission of cancer--a thyroid hormone dependent phenomenon? AB - Spontaneous remission (SR) of neoplasia is a rare biological event. Very few reports provide evidence for an eliciting event or mechanism. The only case in the literature of SR of lung cancer following myxedema coma is suggested to have been an instance of thyroid hormone deprivation-induced total tumor apoptosis. Review of the collective data suggests that the thyroid hormones modulate pleiotropic neoplasia--abetting mechanisms and that hypothyroidism may enhance the predisposition of neoplasms to spontaneous and therapy induced regression by lowering thresholds for apoptosis. PMID- 10697598 TI - A 4-mutation model of carcinogenesis for tumour suppressor genes. AB - Human life can be described as the clonal development of a fertilized egg. During cell proliferation in this clone among others cancer-specific mutations arise. Supposing that about 10(16) cells are produced in a lifetime; that the mutation frequency is about 10(-6) per gene per cell generation; that cells can go through about 60 cell divisions; and that cancer arises about once in a lifetime, carcinogenesis is likely to depend on 4 cancer-specific mutations. The advantage of a 4-mutation model over a multimutational model is that it implies far more predictions that can be tested. The 4-mutation model accurately predicts the age/incidence relation of tumors and the prevalence of cancer in the various organs. It further explains the increased tumor incidence in persons with an inherited cancer gene; and the well-known histological stages of carcinogenesis. PMID- 10697599 TI - The role of molecular discreteness in normal and cancerous growth. AB - The physicochemical events that underlie biological processes are inevitably either/or events. Either a growth factor molecule binds to a cell, or it doesn't. Either a site on a cyclin molecule is phosphorylated, or it isn't. Either a regulatory molecule binds to a DNA sequence, or it doesn't. These molecular either/or events lead to cellular either/or events. Either a cell divides, or it doesn't. Either a cell dies, or it doesn't. Either a cell turns on a particular gene, or it doesn't. Either a tumor cell stays where it is, or it forms a distant metastasis. By considering biological processes as the macroscopic aggregate results of these many individual microscopic either/or events, we can gain considerable insight into both normal and cancerous growth. In fact, as will be outlined here, such discrete modeling may allow us to see how the normal cellular populations of the body can grow to predictable sizes, at predictable times, and to predictable shapes. Such modeling can also allow us to gain insight into how normal cellular populations may become cancerous cellular populations. Indeed, such an approach allows us do a sufficiently good job of imitating the growth and spread of tumors as to be able to make estimates the most effective ways to both detect and treat cancer. PMID- 10697600 TI - A simple stochastic model of development and carcinogenesis. AB - Game theory can be a powerful tool for generating testable hypotheses concerning biological systems. We present a simple game that has many features analogous to a developing cellular system. The game mimics random turning on of genes in cells. Despite the randomness explicit in the mechanism, the game nonetheless results in deterministic outcomes that are extremely resistant to perturbation. Analysis of the types of mistakes or rule changes that are necessary in order to alter the outcome of the game suggests that there are a very limited number of mechanisms by which the differentiation process can result in tumor formation or carcinogenesis. The most significant causes of altered outcomes are alterations in gene order or number, and alteration of the rule by which the gene sequences are traversed. These alterations correspond to chromosomal defects or rearrangements, changes in chromosomal number, and changes in the "orders" delivered by regulatory genes. Notably, most common mistakes, which correspond to simple forms of mutations, have no effect on the outcome of the game, suggesting that mutation is relevant to tumorigenesis and carcinogenesis only to the extent that it results in altered "rules" for reading other gene sequences. PMID- 10697601 TI - Cell damage, aging and transformation: a multilevel analysis of carcinogenesis. AB - There is a vast scientific literature on cancer which no individual can hope to assimilate. To put it mildly, cancer is an enormously complex biological problem that needs to be considered at multiple levels to achieve reasonable understanding. I describe details of work at the cellular level of "spontaneous" neoplastic transformation in culture to point out parallels with the development of cancer in the organism. One of the most surprising relations is that inhibition of cell growth in a transformation-competent population by long term confluence or acutely lowered serum concentration is a strong enhancer of neoplastic change. The transformation is preceded and accompanied by heritable damage to the entire cell population as expressed among progeny cells in a heterogeneous reduction in growth rate at low density as well as delayed reproductive death in some of the cells. The picture bears resemblance to the relation in vivo between local atrophy in the stomach and prostate and cancer in those organs, as well as the relation between tissue damage and cancer in relatively quiescent organs such as pancreas, urinary bladder, etc. Such damage accumulates with age, as does an increasingly permissive local environment for tumor growth. The most common genetic changes found in tumors are large chromosomal deletions. These are precisely the changes that have been found by somatic cell geneticists to cause heritable reduction in growth rate and delayed reproductive death in mutagen-treated and carcinogen-treated somatic cells. Thus, there is a convergence of findings related to cancer in culture and in the organism. The results in cell culture help to focus attention on proximal mechanisms of malignant cell behavior in the organism. However, they are incomplete without considering fundamental aspects of biological behavior such as Elsasser's first principle of ordered heterogeneity in which there can be regularity in the large where there is heterogeneity in the small. The order that controls heterogeneity is weakened with age and contributes to the origin and progression of disordered growth. PMID- 10697602 TI - How aneuploidy may cause cancer and genetic instability. AB - It has been difficult to find a common cause for the many and complex phenotypes of cancer such as dedifferentiation, invasiveness, abnormal morphology, growth rate and metabolism, genetic instability, progression to malignancy, cellular heterogeneity of phenotypes and karyotypes, and clonal origin despite heterogeneity. Over 100 years ago aneuploidy, an abnormal balance of chromosomes, was proposed to cause cancer. However, the aneuploidy hypothesis has since been abandoned, in favor of the gene mutation hypothesis, because it could not offer conventional explanations for cancer-specific phenotypes. For example, the aneuploidy hypothesis seemed unable to (i) explain the genesis of abnormal, cancer-specific phenotypes, (ii) reconcile the heterogeneous karyotypes with the clonal origin of cancers, (iii) explain aneuploidy in non-cancerous cells, and (iv) explain how carcinogens would cause aneuploidy. Here we introduce new evidence that aneuploidy offers a simple, coherent explanation of all cancer specific phenotypes: (i) Congenital and experimental aneuploidy is now known to generate abnormal phenotypes, such as Down syndrome in humans and cancer in animals. (ii) Based on metabolic control analysis, we have derived equations that correlate degrees of aneuploidy with the resulting phenotype abnormalities. These equations suggest that aneuploidy must exceed a certain threshold to generate cancer-specific phenotypes. Therefore, we propose that multistep carcinogenesis corresponds to multiple steps of aneuploidization. (iii) Aneuploidy is also sufficient to explain cancer-specific, karyotypic instability. Since aneuploidy imbalances the highly balance-sensitive components of the spindle apparatus it destabilizes symmetrical chromosome segregation. This autocatalytic instability is the reason why cancers have heterogeneous karyotypes, but are clonal for aneuploidy. Progression to malignancy corresponds to selection of ever more aggressive karyotypic variants. (iv) Both non-genotoxic and genotoxic carcinogens can cause aneuploidy by physical or chemical interaction with mitosis proteins. We conclude that aneuploidy offers a mechanism of phenotype alteration which- above a certain threshold--is sufficient to cause all cancer-specific phenotypes, and is independent of gene mutation. PMID- 10697603 TI - Pernicious cachexia: a different view of cancer. AB - Despite intensive effort to cure breast cancer, treatment generally fails, as evidenced by the age adjusted mortality from breast cancer. For 60 years, breast cancer mortality remained virtually constant. As treatment failed to improve the life prospect of the average patient, it is based on false premises, e.g., Halsted's hypothesis, according to which the tumor is the only threat to the patient. Yet there is more to cancer than just the tumor. Two hallmarks of cancer, cachexia, and paraneoplasia, are usually ignored, since it is assumed that they are caused by the tumor. But, what if it is the other way around, and cancer is first of all a cachexia accompanied by a tumor? At least this could explain why in most cancers treatment fails. Cancer is a chronic systemic disease with local manifestations. Like arteriosclerosis, that is also systemic and manifested solely by its local manifestations, e.g., stroke and myocardial infarction. In the same way as treatment of an ailing heart does not cure the underlying arteriosclerosis, tumor removal does not cure cancer, since being "metabolically" systemic. It is proposed here that carcinogens deplete a vital substance and induce a metabolic deficiency that ends in cachexia. In order to survive, the organism grows a protective organ, the tumor, that replenishes the missing substance. During pre-clinical phase of cancer, deficiency is slight and compensated even by a minute tumor. With time it gets worse and the tumor has to grow more and more in order to make up for the loss, causing pain and secondary damage to vital functions. The patient seeks help and the disease starts its clinical course. When deficiency worsens, the patient becomes cachectic and dies. Such a metabolic relationship exists in pernicious anemia, that illustrates how a tumor might be protective. Cancer is viewed here as pernicious cachexia induced by the loss of a vital metabolite and compensated by the tumor. Until the discovery of the missing substance, treatment ought to preserve the tumor and alleviate its secondary manifestations. PMID- 10697604 TI - Evaluation: complementarity and contradictoriness of cancer theories. A genetic perspective. AB - In this Chapter we evaluate complementarity and contradictoriness regarding theories and data of carcinogenesis described in this issue. Most theories and data are compatible with a multimutation model of carcinogenesis. There are a few authors having severe criticism regarding this mainstream. From a view of philosophy of science such criticism is valuable and this type of papers deserves careful evaluation. Zajicek has the most serious criticism. He argues that cachexia, due to the absence of essential molecules, induces the tumor which tries to produce these missing essential molecules. So, in his view, cachexia causes cancer instead of cancer cachexia. The implication is that cachexia should be treated. Duesberg argues that cancer is due to an imbalance of chromosomes rather than to cancer specific mutations. A few points and implications seem important: (a) Duesberg does not really object to a multimutation model; (b) he wants to defend the view that cancer can also be due to chromosomal imbalance, and (c) cancer due to chromosomal imbalance cannot be inherited, in contrast to cancer based on a mutation. PMID- 10697605 TI - Complementarity and contradiction in cancer research: the role of hierarchies in carcinogenesis. AB - The essays on carcinogenesis in this volume tend to fall into groups according to the level of complexity at which the theories focus. Many investigators focus on events and processes involving genetic mutations. Others, however, identify events or processes at the chromosomal, cellular, tissue, or organsimal level as being of primary importance. While such different approaches to carcinogenesis may, at first, seem to be contradictory, they may be seen to be complementary in light of the general theory of hierarchical organization. Hierarchy theory, which is a development of general systems theory, describes the manner in which organized processes are formed by subunits that have unique properties of their own but which can acquire new properties through their interactions. Hierarchy theory suggests that carcinogenesis can only be understood as a set of interactions between organizational processes at every level from genetic to organismal. PMID- 10697606 TI - Epilogue: what we don't know about cancer. AB - Despite billions of dollars expended since the "war on cancer" was declared more than 25 years ago, cancer continues to be a leading cause of death worldwide, and its precise pathogenesis remains elusive. Moreover, current treatments, even when successful, are grounded in extensive operations, high energy irradiation, and toxic chemotherapy, which carry substantial short- and long-term sequelae. It is, therefore, appropriate to reconsider current concepts of cancer biology in terms of the vast ignorance that surrounds this ancient scourge. From the perspective of a Curriculum on Medical Ignorance, this epilogue surveys questions raised by authors of this monograph, other experts, and patients afflicted with cancer and related disorders. Further, the focus is on basic biological, clinical, and societal implications of carcinogenesis theories as well as the nature and process of scientific inquiry in the context of the general phenomenon of ignorance in medicine. PMID- 10697607 TI - Overexpression of MPS antigens by squamous cell carcinomas of the head and neck: immunohistochemical and serological correlation with FDG positron emission tomography. AB - Survival from advanced primary or recurrent Squamous Cell Carcinoma (SCC) of the head and neck (H&N) is poor. More accurate detection of primary tumors and recurrence may provide ways to improve survival. No standard serum tumor marker is routinely used for surveillance of SCC-H&N. In this paper, we evaluated the performance characteristics of the MPS-H tumor marker test for the quantitative measurement of "MPS-H" heat-generated immunoreactive proteins and assessed the clinical utility of this marker in the detection and monitoring of SCC-H&N. In approximately 92% of the subjects having no evidence of SCC-H&N, the MPS-H levels were lower than 15 ng/mL. In 76% of patients having SCC-H&N at various stages (T1 T4), the MPS-H level was > 15 ng/mL (range: 20-200 ng/mL). In addition, we found a statistically significant correlation between PET positive cases and high MPS-H serum levels in SCC-H&N patients with recurrent disease. These results suggest that MPS-H may provide an initial screening test that would allow for selective PET imaging in these patients. Furthermore, we found that there was greater expression of MPS-1 in tumors of higher histological grades. Thus, in tumors with more histological aggressiveness there is more MPS-1, indicating the potential usefulness of this marker in prognosis for SSC-H&N. Considering the immunohistochemical, serological, and FDG-PET data presented here, and the compelling need to expedite the early diagnosis of primary and recurrent epithelial malignancies of the head and neck, we are further evaluating the system of MPS antigens in a large patient population as a tool for the early serologic and histologic diagnosis of SCC-H&N. PMID- 10697608 TI - Inhalation therapy using a streptococcal preparation (OK-432) against bronchioloalveolar carcinoma of the lung. AB - OBJECTIVE: In order to inhibit the occurrence of airborne metastasis of bronchioloalveolar cell carcinoma (BAC), we tried to activate alveolar macrophages by the inhalation of aerosolized OK-432, which is a heat and penicillin-treated lyophilized preparation of the Su strain of Streptococcus pyogenes. METHODS AND RESULTS: Alveolar macrophages were obtained from resected specimens of lung cancer patients and cultured for 24 h in the presence of various concentrations of OK-432 (0.001-1 KE/ml). The cytotoxic activity against a lung cancer cell line was augmented in a dose dependent manner and reached a plateau level at 0.1 KE/ml of OK-432. Furthermore, the alveolar macrophages produced various cytokines, i.e., IL-1, TNF-alpha, and IL-6 after 72 h cultivation with 0.1 KE/ml of OK-432. Based on the in vitro experiments, six patients with intrapulmonary metastatic BAC were therefore treated by the inhalation of aerosolized OK-432. All 6 patients inhaled aerosolized OK-432 (0.1 KE/ml) twice a day for 5 days. The inhalation therapy regimen was repeated either weekly or monthly unless the tumor markedly progressed. No adverse events were observed in any patients. Either an augmentation of antitumor cytotoxicity or TNF alpha production by the alveolar macrophages was observed in the two of three patients examined. CONCLUSION: OK-432 inhalation therapy was found to be safe and thus warrants further investigation to determine its clinical effectiveness for BAC. PMID- 10697609 TI - Cyclin D1 is a possible predictor of sensitivity to chemoradiotherapy for esophageal squamous cell carcinoma. AB - BACKGROUND: Chemoradiotherapy is currently performed on patients with advanced esophageal squamous cell carcinoma (SCC). The preoperative administration revealed that the patients who responded well to chemoradiotherapy had favorable outcomes, whereas the poor responders conversely resulted in worse prognosis. The aim of this study was to identify molecular markers predicting sensitivity to chemoradiotherapy prior to this treatment. MATERIALS AND METHODS: Our clinical protocol for chemoradiotherapy for esophageal SCC were enrolled in 34 patients comprising 20 patients who underwent surgery after neoadjuvant chemoradiotherapy and 14 patients who were treated chemoradiotherapy without surgery. The expressions of cyclin D1, p53 and Ki-67 were investigated immunohistochemically in biopsy specimens obtained before the treatment from all 34 patients. The immunoreactivities were compared with responsiveness to chemoradiotherapy as evaluated by macroscopic or microscopic method. RESULTS: The mean rate of primary tumor reduction as estimated by esophagography was 75.3% in the cyclin D1 negative group whereas 42.7% reduction rate was observed in the cyclin D1 positive group. The difference in the reduction rate between cyclin D1 positive and negative groups was statistically significant (p = 0.0025). The immunoreactivities of p53 and Ki-67 did not show a significant correlation with responsiveness to chemoradiotherapy. In neoadjuvant group, patients with cyclin D1-positive tumors showed significantly worse overall survival than patients with cyclin D1-negative tumors (p = 0.0380). CONCLUSIONS: Among 34 patients with esophageal SCC, differences in the responsiveness to chemoradiotherapy were correlated with cyclin D1 immunoreactivity assessed in the biopsy specimens. Thus the cyclin D1 protein may be a useful predictor of sensitivity to concurrent chemoradiotherapy for esophageal SCC. PMID- 10697610 TI - Hras1 VNTR alleles as susceptibility markers for lung cancer: relationship to microsatellite instability in tumors. AB - PURPOSE: To further evaluate lung cancer risk associated with rare Hras1 VNTR alleles and possible biological mechanisms. MATERIALS AND METHODS: The Hras1 VNTR was genotyped in 295 lung cancer patients and 500 healthy controls by PCR and high resolution electrophoresis. Microsatellite alterations were examined in 168 tumors by PCR and capillary electrophoresis. RESULTS: 35 Hras1 VNTR alleles were found, of which 24 were defined as rare. A relative risk of 3.3 (95% CI; 1.9-6.0) associated with rare alleles was obtained using the total groups. Increased risk was significant both for males and females. When a matched control group was used, a relative risk of 12.7 (95% CI; 1.7-93.9) was calculated for individuals with rare alleles at the Hras1 VNTR locus. A low frequency of microsatellite alterations was observed (4.7%) in lung tumors. The frequency of altered microsatellite loci was higher among patients with rare Hras1 VNTR alleles than among patients with common alleles. CONCLUSION: Rare Hras1 VNTR alleles are associated with lung cancer risk, and a genetic mechanism which increases allelic diversity may be involved. PMID- 10697611 TI - Selective use of preoperative radiotherapy in the treatment of cancer in the lower two thirds of the rectum. AB - PURPOSE: The authors review the result of the selection of patients with a low rectal cancer for pre-operative radiotherapy. METHODS: The selection was based on the findings of digital examination eventually combined with surgical staging consisting of bimanual palpation during a staging laparotomy or "trial" operation. This selection was used to divide the patients into three groups: one where local radicality could be expected from primary surgery (group 1), one with deeply infiltrating, but mobile tumours requiring 10 x 3 Gy pre-operative radiotherapy (group 2) and one with fixed or borderline resectable tumours requiring protracted pre-operative radiotherapy with 55-59 Gy (group 3). One hundred and one patients were eligible for this study. A resection aiming for pelvic radicality was carried out in 94 patients: primary resection in 38 (group 1), surgery subsequent to 10 x 3 Gy pre-operative radiotherapy in 20 (group 2) and 55-59 Gy in 36 (group 3). RESULTS: The calculated risk of local recurrence at 5 years was 15% (95% C.I. 4-27) for group 1, 8% (95% C.I. 0-20) for group 2 and 30% (95% C.I. 16-44) for group 3. The calculated 5 years survival for the 3 groups was respectively 60%, 49% and 39%. CONCLUSION: The overlap in local recurrence rate between the three groups suggests a substantial downgrading by this approach of selective use of pre-operative radiotherapy in the patients with the most advanced tumour. Notwithstanding recent improvements of imaging techniques there still is a place for the staging laparotomy in the selection of the treatment strategy for advanced rectal cancers. PMID- 10697612 TI - 37 kiloDalton oncofetal antigen protein and immature laminin receptor protein are identical, universal T-cell inducing immunogens on primary rodent and human cancers. AB - Based upon positive immunologic comparisons, protein and cDNA sequencing, and in vitro and in vivo immunogenicity studies we propose that carcinomas, sarcomas and lymphomas/leukemias of rodents and humans share 37 kDa Onco-Fetal Antigen [OFA] as a T and B-lymphocyte stimulating, universal tumor specific transplantation antigen [UTSTA]. In the past four years, biochemical studies from several laboratories studying laminin receptor protein and immunological studies of OFA from our laboratories independently converged. OFA protein and immature or precursor Laminin Receptor Protein [iLRP] are > 99% identical proteins based on amino acid and cDNA sequencing and immunobiology studies summarized here. Acquired expression of 37 kDa OFA/iLRP enables malignant tumor cells to penetrate laminin tissue and vessel barriers. 37 kDa OFA/iLRP activates precursor thymic anti-OFA/iLRP specific cytotoxic T cell which kill emerging pretumor cells. Our reported findings also demonstrate that OFA/iLRP can function to induce specific immunoregulatory CD8 T-suppressor cells secreting IL-10 which impair effector T cell killing of emerging tumor cells non-specifically and thereby facilitate tumor progression. Potential applications of OFA/iLRP detection in early cancer formation, for monitoring patient T cell subclass responses to OFA/iLRP as a predictor of tumor progression, and the use of OFA/iLRP peptides for specific anti-tumor immunotherapy are presented. PMID- 10697613 TI - Thymidine phosphorylase expression in invasive carcinoma: correlations with clinicopathologic variables and in vitro chemosensitivity to 5-fluorouracil. AB - BACKGROUND: Thymidine phosphorylase (TP) is critical for angiogenic activity, but little information is available on the relationship between TP expression and other Clinicopathologic variables. Furthermore, the relationship between TP and chemosensitivity is still debated. MATERIALS AND METHODS: The expression of TP was examined in 116 primary breast carcinomas and in vitro chemosensitivity was assessed in 67 of them by histoculture drug response assay (HDRA) using 5 fluorouracil (5-FU). RESULTS: Tumor cell TP expression was significantly inversely correlated with histological grade and positively correlated with Bcl-2 expression, but no association with other tumor variables was found. Unexpectedly, TP immunoreactivity was not related to 5-FU chemosensitivity. CONCLUSION: The present results suggest that TP is important for remodeling the existing vasculature early in tumor development and intraductal extension, expressions of TP and Bcl-2 are tightly linked and TP status can not generally predict chemotherapeutic sensitivity for 5-FU as a single molecular marker. PMID- 10697614 TI - Subcutaneous administration of interferon alpha and gamma in patients with metastatic renal cell carcinoma. AB - BACKGROUND: Renal cell carcinoma (RCC) is relatively resistant to both chemotherapy and radiotherapy. Response and survival of treatment with Interferon alpha (IFN-alpha) and Interferon-gamma (IFN-gamma) were evaluated in patients with metastatic RCC. PATIENTS AND METHODS: Thirty-one patients with confirmed RCC were included in this study. Fifteen of 31 patients received injection of IFN alpha and IFN-gamma three times a week. IFN-gamma was infused subcutaneously by microinfusion pump. Sixteen received IFN-alpha alone more than three times a week. RESULTS: The overall response rate was 20.0% in the IFN-alpha and IFN-gamma group, and 12.5% in the IFN-alpha alone group. Long lasting stabilization of the disease (more than; 12 months) was seen in 92.3% of CR, PR or SD in the IFN-alpha and IFN-gamma group, as compared with 71.4% in the IFN-alpha alone group. Both groups differed significantly in survival rate from the first treatment with IFN (p < 0.05). CONCLUSIONS: A long lasting stabilization of the disease can be expected in patients who were treated with our regimen of IFN-alpha and IFN gamma. PMID- 10697615 TI - Urinary gonadotropin peptide (UGP) and serum CA 125 in gynaecologic practice, a clinical prospective study. AB - BACKGROUND: Beta human chorionic gonadotropin (beta-hCG) is expressed in human fetal tissue and cancer cells of various histologic types. It is degraded to the beta-core fragment (beta cf-hCG) which is concentrated in urine, and is known as urinary gonadotropin peptide (UGP). The objective of this study was to assess 1) the value of urinary gonadotropin peptide (UGP) as a single test and the combination of UGP with CA 125 as a diagnostic test in predicting the benign or malignant origin of gynecologic disease, 2) the influence of surgical removal of the tumor on the levels of UGP and CA 125, 3) the influence of the urinary concentration on the UGP levels in relation to the test results. PATIENTS, MATERIALS, METHODS AND STATISTICS: Serum and urine were collected from 31 gynecological patients with malignant and non-malignant disease, preoperatively, and 1 week and 6 weeks after surgery. Optimal cut-off levels were determined by Receiver Operating Characteristic-curves (ROC). Sensitivity (SENS), specificity (SPEC), positive (PPV) and negative predictive value (NPV) and overall test accuracy (ACC) for their ability to discriminate benign from malignant masses were calculated. Logistic regression analysis was performed to calculate the contribution of CA 125, UGP and UGP/creatinine (UGP/creat) to a model predicting malignancy. RESULTS: The optimal cut-off level for UGP was found 1 fmol/l, for UGP/creat 1.33 fmol/mg creatinine and for CA 125 100 kU/L. The distribution of the urinary creatinine values varied considerably (median = 8.3 mmol/l, range 0.6 25.8 mmol/l). The correlation coefficient (r) between log UGP and log CA 125 was 0.44 (p = 0.001) and between log UGP/creat and log CA 125 0.53 (p < 0.0001). CONCLUSIONS: UGP may be used as a tumor maker in gynecological disease. However, CA 125 as single test discriminates malignant from benign disease better than UGP or UGP/creat. In a logistic model the combination of CA 125 with UGP and UGP/creat predicts the benign or malignant character in 89% of the study population. Significant changes in UGP and UGP/creat levels were seen after removal of benign tumors, however, this was not found in ovarian cancer patients. Correction of the UGP values for the urinary concentration improved the results slightly. PMID- 10697616 TI - The clinical value of the Enzymun-Test for total and free PSA--a multicentre evaluation. AB - This study examined the clinical relevance of the determination of free PSA (f PSA) in addition to total PSA (t-PSA) in 6 study groups. PATIENTS AND METHODS: Both total PSA- and free PSA-values of sera samples obtained pretherapeutically from 455 patients with carcinoma (PCA) and 680 patients with benign hyperplasia of the prostate (BPH) were analyzed by means of Enzymun-Test PSA and Enzymun-Test Free PSA (Boehringer Mannheim GmbH, Germany). RESULTS: At 95% specificity (true negative test results), a cutoff value of 13.6 [micrograms/L] was obtained for total PSA (34 patients with BPH [5%] were above this value). For this cutoff value we calculated a sensitivity (true positive test results) of 44%. Using the same criteria for the ratio Q = f-PSA:t-PSA a cutoff of 0.13 was found again at a specificity of 95%. In a second step only patients with total PSA values below the cutoff level of 13.6 [micrograms/L]) were considered. Out of these patients 26 of 646 with BPH and 108 of 257 with PCA were below the above mentioned ratio (Q = 0.13). Considering both steps (total PSA and Q) 306 patients with PCA were detected correctly and 60 patients with BPH would have been biopsied unnecessarily. CONCLUSION: High total PSA levels are a very good indicator for the presence of prostate cancer. There is still concern to improve the differentiation of the diagnosis between BPH and PCA, when an intermediate or low value (< or = 95% specificity) is observed. The determination of the ratio is only useful in this range. It is more powerful at discriminating between PCA and BPH than t-PSA alone and may contribute to a reduction in unnecessary invasive techniques. PMID- 10697617 TI - Two cases of osteosarcoma occurring as second malignancy of childhood cancer. AB - We report on two patients in whom osteosarcoma occurred as second malignancy of childhood cancer. One patient had a malignant teratoma and the other adrenocortical carcinoma as the primary cancer. The emergence of cancer in cured cases and long-term survival cases of childhood cancer may result in an increase in the number of osteosarcomas seen in adolescence occurring as second malignancy. Anti-cancer drugs in large does were used for the treatment of a malignant teratoma in the former. These anti-cancer drugs may be involved in the occurrence of the second malignancy. In the latter, the patient has the germ-line mutation of p53 tumor suppressor gene, so genetic factors are presumably related to the occurrence of the second malignancy. PMID- 10697619 TI - Breast cancer metastatic to the kidney. AB - We report a case of renal tumor secondary to a breast cancer occurring 16 years after radical mastectomy. This is the sixth report case of renal metastasis from breast cancer of a 51-year-old woman. Percutaneous biopsy of the renal tumor confirmed the diagnosis during the follow-up. The patient was treated with chemotherapy and is alive 8 months after diagnosis. Previously, cases like our case showed long interval from mastectomy to diagnosis of metastasis. PMID- 10697618 TI - The effect of orchiectomy on lung cancer survival. AB - Gender is an independent prognostic factor for survival from lung cancer with the relative risk of lung cancer death for men compared to women being 1.3. This observation remains unexplained but in vitro data suggests a stimulatory effect of androgens on lung cancer growth. We hypothesized that androgen deprivation improves survival in men with lung cancer compared to hormonally intact men with lung cancer. On the basis of age, race and region, we matched 44 men with lung cancer and bilateral orchiectomy with 88 men who had lung cancer and no history of orchiectomy and compared their survival. Since hazards were non-proportional we chose the generalized gamma model to describe the survival function. The survival was significantly different between the two groups (p = 0.0048) with the orchiectomy group having a better two year overall survival. This retrospective study suggests that androgen depletion may significantly improve the short term survival of men with lung cancer but further, prospective investigation is required for confirmation. PMID- 10697620 TI - The role of surgery in renal cell carcinoma with solitary metachronous metastasis to contralateral adrenal gland. AB - We report a case of renal cell carcinoma with solitary metachronous metastasis to contralateral adrenal gland occurring 9 months after radical nephrectomy. The patient was treated with a contralateral adrenalectomy and is alive for 87 months. The literature was reviewed and 5-year survival of solitary metachronous metastasis to contralateral adrenal gland was 60%. Follow-up duration of our case was the longest in the literature. It is suggested that the solitary contralateral adrenal gland metastasis of renal cell carcinoma should be resected since there is no effective treatment of metastatic renal cell carcinoma. Good prognosis may be then and the good be expected. PMID- 10697622 TI - Increased survival in brain metastatic patients treated with stereotactic radiotherapy, omega three fatty acids and bioflavonoids. AB - Stereotactic radiotherapy represents a method to effectively treat brain metastases with high precision and with high doses. Few acute toxicities are associated with stereotactic radiotherapy, however delayed reactions may occur and after six months, 20% of patients can develop radionecrosis. To avoid this adverse effect, in patients with metastases localized in critical brain areas, a supplementation of Omega three fatty acids and bioflavonoids has been used. At the end of 1997, we initiated a series of retrospective studies to test the efficacy of stereotactic radiotherapy on 405 patients, and the prognostic importance on survival of various variables among which this type of supplementation. From the comparison of various survival curves with the Cox multivariate analysis, it emerged that the patients using this supplementation had a decreased risk ratio and an improvement in survival time. A decreased number of radionecrosis was noted. We suggest their use as radioprotectors. PMID- 10697621 TI - Serum type I collagen degradation markers, ICTP and CrossLaps, are factors for poor survival in lung cancer. AB - We investigated the prognostic value of the serum markers of type I collagen synthesis (PINP and PICP) and degradation (ICTP and CrossLaps) in 143 lung cancer patients with a local or locally advanced disease or a metastatic disease. The mean values of ICTP, CrossLaps, PINP and PICP were significantly higher in patients with bone metastases than in those without metastases or with only soft tissue metastases. The patients with ICTP < or = 5.0 micrograms/l or CrossLaps < or = 5000 pmol/l had a better prognosis. The histopathological type, the site of metastases or the stage of the disease had no influence on these results. In multivariate regression analysis, both ICTP and CrossLaps in contrast to PINP or PICP, were prognostic factors for poor survival in lung cancer patients. ICTP, CrossLaps, sedimentation rate, hemoglobin and AFOS reached separately weaker, but statistically significant values as predictors of survival with stage and operation. PMID- 10697623 TI - Efficacy of 51Cr-EDTA clearance to tailor a carboplatin therapeutic regimen in ovarian cancer patients. AB - AIM: The aim of this study was to evaluate the efficacy of 51Cr-EDTA clearance to tailor the carboplatin dose in two different therapeutic regimens of advanced epithelial ovarian cancer. MATERIALS AND METHODS: 14 patients entered the study, eight treated by carboplatin (C) alone and six by C and paclitaxel (P). The dose of C was calculated from the Calvert formula [DOSE(mg) = desired AUC x (GFR + 25)] based on the Glomerular filtration rate (GFR) figure; in our protocol desired Area under the curve (AUC) figure was 5 mg/ml x min. The method used to calculate the GFR requires only 4 blood samples taken in the late part of the disappearance plasmatic curve and conjugates accuracy to an acceptable clinical compliance. RESULTS: In only 5 courses a significant hematological toxicity (HT) was present (4 courses grade 2, 1 course grade 3); it was necessary to delay only 2 courses; no treatment was discontinued because of HT. CONCLUSION: We concluded that there is no summation toxicity of C and P if administered simultaneously and that the assessment of GFR by 51Cr-EDTA clearance is an optimal tool to predict an acceptable toxicity. PMID- 10697624 TI - The role of adjuvant immunotherapy after radical nephrectomy and prognostic factors in pT3N0M0 renal cell carcinoma. AB - Five-year overall survival after radical nephrectomy in pT3N0M0 renal cell carcinoma is 35-50%. In light of immunotherapy, which has shown some activity in advanced diseases with increasing efficacy in limited metastatic invasion, we decided to explore the theoretical advantage of adjuvant immunotherapy in radically resected stage pT3N0M0 renal cell carcinoma. We studied several factors including tumor size, nuclear grade, mean nuclear area and expression of p53 protein to find out which factor is concerned with disease progression. A total of 10 patients with pT3N0M0 RCC who received radical nephrectomy from February 1992 to April 1999 were randomly assigned to receive treatment with either interferon-alpha alone or interferon-alpha plus vinblastine. Eight patients with pT3N0M0 RCC who received only radical nephrectomy from January 1984 to February 1993 were analyzed and the results were compared with the first group. Six out of 10 (60%) patients in the adjuvant immunotherapy group are alive with no evidence of disease. Metastases were documented in 4 patients (40%) with a median interval to progression of 17.5 months. All of them died of tumor. In the surgery only group, 5 out of 8 patients (62.5%) are still alive with no evidence of disease. Two patients (25%) developed distant metastases and both of them died of tumor. The median progression interval was 11 months. There were no statistical differences in time to progression and survival rate between the two groups. In the univariate analysis using a log-rank test, the expression of p53 protein seemed to be associated with shorter survival (p = 0.0591). However, in the multivariate analysis using Cox's proportional hazard model, no parameter had significant independent prognostic value. We concluded that adjuvant immunotherapy did not improve the survival of patients with pT3N0M0 RCC. Furthermore, we failed to find significant prognostic factors in patients with pT3N0M0 RCC. PMID- 10697625 TI - Nonspecific cross-reacting antigen-50/90 (NCA-50/90) as a new tumor marker. AB - BACKGROUND: The nonspecific cross-reacting antigen-50/90 (NCA-50/90) is a glycoprotein antigen which shares some antigenic determinants with carcinoembryonic antigen (CEA). No definite clinical value has been established for the measurement of NCA-50/90 in cancer patients. METHODS: We established and evaluated a chemiluminescent enzyme immunoassay (CLEIA) specific for NCA-50/90 using two monoclonal antibodies. RESULTS: No significant reactivity with 4 purified related antigens including CEA was found in the NCA-50/90 CLEIA. Serum samples (n = 572) from patients with malignant (n = 326) as well as from healthy individuals (n = 246) were analyzed by the NCA-50/90 CLEIA and by the established ACCESS CEA assay. The average sensitivity of NCA-50/90 for malignant disease was 40.8%, compared to 45.4% of CEA. Relatively high positive rates of NCA-50/90 were observed in sera from patients with lung cancer (72.0%), hepatoma (62.5%), pancreatic cancer (47.6%), breast cancer (35.6%), and colorectal cancer (34.5%). About 15% of patients with malignant disease were positive only for NCA-50/90. The levels of NCA-50/90 and CEA in sera from patients with malignant disease correlated only poorly. CONCLUSIONS: The present study suggests that increases in blood levels of NCA-50/90 occur in a population of cancer patients which is different from those with elevated levels of CEA and that NCA-50/90 might be useful for NCA-50/90-positive, but CEA-negative patients. PMID- 10697626 TI - The significance of body weight change in non-Hodgkins lymphoma. AB - BACKGROUND: The purpose of this study was to evaluate the effect of weight changes after completion of chemotherapy on the prognosis and survival of patients with intermediate and high grade non-Hodgkin's lymphoma. MATERIALS AND METHODS: A retrospective analysis of data on patients from the TCOG T1488 protocol, a phase II study using CHOP in the treatment of intermediate and high grade lymphoma. From September, 1988 to December 1994, 138 adult patients had complete weight data for analysis. Weight gain in lymphoma patients after therapy significantly correlated with improved survival (Logrank test p = .0031). In patients with initial B symptoms, weight gain after therapy correlated with survival (Logrank test p = .0039), female patients (odds ratio = 6.2) were less likely to gain weight on treatment. CONCLUSION: Weight gain after chemotherapy for lymphoma is a significant positive prognostic factor for survival. PMID- 10697627 TI - Genetics and lung carcinoma in Okinawa Prefecture, Japan. AB - BACKGROUND: Although the incidence of all cancers in Okinawa is the lowest in Japan, that of lung cancer is high. This study was performed to clarify the underlying mechanism of this tendency. MATERIALS AND METHODS: Family histories of the lung cancer patients in Okinawa, p53 mutation, microsatellite alterations, and titers of serum anti-p53 antibodies were examined. RESULTS: The number of patients who had relatives with some malignancies in relatives was low in Okinawa, but lung cancer was frequently observed in their relatives. Overexpression of p53 protein was frequently observed in squamous cell carcinoma (SCC) than in adenocarcinoma (AD), and in smokers than in non-smokers. Anti-p53 antibodies were detected in 17.4%. The incidence of loss of heterozygosity at D3S643 and at IFNA were higher in SCC than in AD. CONCLUSIONS: Lung cancer was frequently observed in relatives of lung cancer patients. Pulmonary SCC had different genetic alterations compared with pulmonary AD in Okinawa. PMID- 10697628 TI - Clinical benefit and response in patients with gastric cancer to weekly 24-hour infusion of high-dose 5-fluorouracil (5-FU) and leucovorin (LV). AB - BACKGROUND: A prospective study evaluated the efficacy and correlation of different outcome measurements, including the WHO response criteria and clinical benefit (CB), to weekly high-dose 5-FU and LV for patients with advanced gastric cancer. PATIENTS AND METHODS: Thirty-nine chemotherapy-naive patients were enrolled from Sep. 1996 to Oct. 1997. The treatment consisted of a 24-hour continuous infusion of 5-FU 2600 mg/m2 & LV 150 mg weekly for 6 weeks with a subsequent 2-week break. The responses were evaluated by CB and WHO criteria at the end of the 8th week, then at 8-week intervals. RESULTS: There were 21 male and 18 female patients with a median age of 56 years. The median Karnofsky performance score was 70%. Thirty-six patients were evaluable for WHO criteria, and 12 (33.3%) had partial response, 12 (33.3%) had stable disease and 12 (33.3%) had progressive disease. Twenty-one of the 35 (60%) evaluable patients for CB were found to have a positive response. There was a significant correlation between WHO response and CB. The median survival was 10.5 months for CB responders, while the median survival among the CB non-responders was 5 months only. CONCLUSIONS: This study found that this regimen yielded a 60% CB, despite a 33% WHO response rate. Improvement in CB resulted in an improvement in survival as well as the correlation between CB and WHO response, and suggested the value of CB as an alternative indicator for clinical response. PMID- 10697629 TI - Thymidylate synthase levels as a therapeutic and prognostic predictor in breast cancer. AB - 5-Fluorouracil (5-FU) is a commonly used adjuvant therapeutic drug in treating breast cancer. 5-FU is metabolically converted to 5-fluorouracil-2'-deoxyuridine 5'-monophosphate-(FdUMP) which is believed to inhibit DNA synthesis in neoplastic cells by forming a tightly bound ternary complex with thymidylate synthase (TS). In the present study, we examined the possible relationship between TS levels and clinico-pathologic and prognostic features in breast disease. Mean TS levels of 2.9 pmol/g, 6.1 pmol/g, and 23.1 pmol/g were obtained in cases of benign breast disease (3 cases), primary breast cancer (115 cases), and recurrent tumors (4 cases), respectively. In breast cancer, mean TS levels significantly correlated with S-phase fraction (SPF), DNA polymerase a and lymphatic invasion. Thus, TS levels in breast cancer significantly reflected cell proliferation and malignancy. Regarding the survival rate, patients with TS values above 10 pmol/g showed an unfavorable prognosis. The effectiveness of adjuvant 5-FU derivatives chemotherapy was reflected in a higher disease-free survival rate in node (+) cases showing TS levels between 5 and 10 pmol/g (p < 0.1), but not in node (-) cases. In conclusion, TS levels in neoplastic tissues of the breast were highest in recurrent tumors, followed by those in primary cancer, benign breast disease and in breast cancer which reflected proliferative activity. Breast cancers with extremely high TS levels were accompanied by an unfavorable prognosis; however, those with moderately high TS levels tended to respond to adjuvant chemotherapy with 5-FU derivatives. PMID- 10697630 TI - Smoking and smoking-related lung cancer in female patients. AB - Smoking is the primary factor responsible for the epidemic of squamous cell lung cancer (SqLC) and small cell lung cancer (SCLC), however, there have been few studies which reported the relationship between smoking and SCLC in female patients. To examine the relationship between smoking and SCLC and SqLC in female patients, a retrospective chart review was performed in 877 patients with lung cancer diagnosed at Tsukuba University Hospital. In both SCLC and SqLC, the proportions of non-smokers were higher in female patients than those of male patients (p = 0.0001, p = 0.0001, respectively). A significant difference in smoking history was found between female and male patients who were diagnosed with SqLC and SCLC (p = 0.0001). In female patients, active smoking seems less responsible for the occurrence of smoking-related lung cancer compared with male patients, which suggests passive smoking influence the development of smoking related lung cancer, especially in female patients. PMID- 10697631 TI - Molecular remission induced by fractionated dose-escalating donor leukocyte infusion without graft-versus-host disease in a patient with chronic myelogenous leukemia relapsed after allogeneic bone marrow transplantation. AB - A 39-year-old male with CML who relapsed 5 years and 8 months after allogeneic bone marrow transplantation achieved complete molecular remission following fractionated dose-escalating donor leukocyte infusions. Acute or chronic graft versus-host disease (GVHD) did not occur and the patient remained asymptomatic throughout treatment. Since no prophylaxis against GVHD was administered, this case indicated that the graft-versus-leukemia effect is entirely separate from GVHD in certain conditions. PMID- 10697632 TI - Thymidylate synthetase and dihydropyrimidine dehydrogenase levels in gastric cancer. AB - The measurement of thymidylate synthetase (TS) and dihydropyrimidine dehydrogenase (DPD) enzymatic activities and mRNA levels in tumors may be useful in predicting tumor sensitivity to 5-fluorouracil (5-FU). Forty-one patients with advanced gastric cancer gave informed consent and were enrolled in this study. Biopsy specimens of gastric cancer were obtained preoperatively through gastrofiberscopy and used to determine TS and DPD mRNA levels. We also measured TS and DPD enzymatic activities and mRNA levels in surgically resected gastric cancer samples, as well as in adjacent normal gastric mucosa. TS and DPD activities were measured using the TS-binding assay and a radioenzymatic assay, respectively, while mRNA levels were measured by semi-quantitative reverse transcription-PCR (RT-PCR) co-amplified with glyceraldehyde-3-phosphate dehydrogenase (GAPDH) as an internal standard. In resected tumor specimens, TS and DPD activities ranged from 7.1 to 176.6 fmol/mg protein and from 3.6 to 99.8 pmol/min/mg protein, respectively, while TS and DPD mRNA levels ranged from 0.50 to 21.12 and from 0.014 to 7:22, respectively. There were no significant correlations between TS/DPD levels and other clinicopathological factors, except for low DPD mRNA levels in undifferentiated carcinoma. Both TS activity and mRNA levels were significantly higher in tumor tissues compared to normal adjacent mucosa. In contrast, there was no significant difference between tumoral and non tumoral DPD activity, although tumor tissue showed significantly lower DPD mRNA levels than non-tumoral tissue. High tumoral TS mRNA levels in preoperative biopsy specimens from patients with stage III/IV was associated with poor survival outcome after surgery compared with patients with low tumoral TS mRNA levels. In contrast, DPD levels had no influence on prognosis. We conclude that high tumoral TS levels and low tumoral DPD mRNA may indicate the selective cytotoxicity of 5-FU on gastric cancer, and that tumoral TS mRNA levels may be a prognostic factor for patients with stage III/IV gastric cancer. PMID- 10697633 TI - Portal venous gas associated with splenic abscess secondary to colon cancer. AB - We report a successfully treated case accompanied by portal venous gas, which was associated with splenic abscess due to penetration of colon cancer. In June, 1998, a 67-year-old Japanese man was referred to our hospital because of a continuous fever over 40 degrees C and portal venous gas detected by computed tomography (CT). CT revealed low density areas in the spleen and wall thickening of the descending colon next to the spleen. Barium-enema examination demonstrated an extrinsic filling defect in the splenic flexure of the colon. Splenectomy, resection of the pancreatic tail and left hemicolectomy were performed Histopathological studies showed moderately differentiated adenocarcinoma, which made a fistula at the bottom of the ulceration to the spleen. The postoperative course was uneventful. The portal venous gas was likely to have resulted from a bacterial infection in the portal venous systems secondary to the splenic abscess. PMID- 10697634 TI - Adoptive immunotherapy of advanced solid tumors: an eight year clinical experience. AB - BACKGROUND: Adoptive immunotherapy (AI) of cancer, based upon the injection of in vitro manipulated autologous lymphocytes is still in an experimental phase. Our group started different clinical trials of AI in early 1990, and, at present, some specific targets for this approach seem to have been identified. PATIENTS AND METHODS: 296 patients with solid tumors (melanoma, kidney carcinoma, non small-cell lung cancer, mesothelioma, neoplastic pleural effusion, and liver cancer) were treated with either locoregional or systemic adoptive immunotherapy (AI) using both LAK and TIL cells in combination with s.c. rIL-2. RESULTS: The surgery/AI combination resulted in good clinical results, characterized by enhanced survival and long lasting disease free periods in a significant number of patients. CONCLUSIONS: AI seems to be efficacious in the treatment of melanoma, lung and hepatic cancers. Further studies will expand the application of the treatment to other malignancies. PMID- 10697635 TI - Cisplatin-carboplatin-gemcitabine or ifosfamide-gemcitabine in advanced non-small cell lung carcinoma: two pilot studies. AB - Gemcitabine has been demonstrated active in non-small cell lung cancer (NSCLC). The objective of this trial was to evaluate the feasibility of combinations of gemcitabine (1 g/m2 dl,8,15) with cisplatin (60 mg/m2 dl) and carboplatin (200 mg/m2 dl) (CCG; n = 12) or ifosfamide (4.5 g/m2 dl) (IG;n = 4) in patients with advanced NSCLC, in order to prepare a phase III randomised trial. Toxicity, mainly haematological, was tolerable. It consisted in neutropenia (IG) and both thrombopenia and neutropenia (CCG). The administration of carboplatin according to the AUC (AUC = 3) resulted in a significant reduction of haematological toxicity. A good number of responses were documented. These acceptable results urged our group to compare these regimens to the combination of cisplatin, carboplatin and if osfamide. PMID- 10697636 TI - Ovarian cancer-induced immunosuppression: relationship to tumor necrosis factor alpha (TNF-alpha) release from ovarian tissue. AB - Cytokines have been reported to be potential biological markers of, disease status in cancer patients. Tumor necrosis factor-alpha (TNF-alpha) is a key cytokine released from monocytes and macrophages. TNF-alpha is involved in essential biological functions such as immunoregulation, modulation of cell growth and differentiation. In this work, the role of TNF-alpha release in ovarian cancer patients was investigated. Fifty-five patients with ovarian cancer and 20 controls of matched age and parity were included in this study. TNF-alpha concentrations were measured in sera and cytosolic fractions of both groups. The results demonstrated a significant increase in TNF-alpha concentrations among patients compared to the control subjects (P < 0.001). Furthermore, a non significant increase (P = 0.05, was observed between the different types (serous, Mucinous, and endometrioid) of epithelial ovarian cancers. Also TNF-alpha concentrations did not correlate with the disease stage. Moreover, immunohistochemical analysis of tissue specimens stained for TNF-alpha was positive in malignant lesions and negative for the normal ovarian tissue. These findings confirmed the TNF-alpha kinetics obtained by ELISA assays. Interestingly, TNF-alpha levels were also elevated in culture supernatants of PBMC stimulated by cytosolic fractions from malignant ovarian tissues. Blastogenic assays using cytosolic fractions from malignant ovarian specimens to stimulate healthy donor peripheral blood mononuclear cells (PBMC) showed a marked decrease in 3H-thymidine uptake compared to the cells stimulated by normal cytosols. To establish a cause-effect relationship between TNF-alpha release and inhibition of cell proliferation, the experiments showed that 3H-thymidine uptake by PBMC was markedly inhibited by recombinant human TNF-alpha (rh TNF-alpha) and that inhibition was significantly reversed when TNF-alpha monoclonal antibody was added to the cells. The data presented in this work indicate that TNF-alpha may play an important role in the biology of ovarian cancer and hence, tumor pathogenesis. PMID- 10697637 TI - Serum chromogranin A in monitoring metastatic prostate cancer patients. AB - In a number of studies the important role of neuroendocrine structures during the development of aggressive prostate cancer has been reported. We have recorded serum Chromogranin A and PSA values (CIS biointernational, France) in responders and nonresponders to both hormone therapy (Mab) and chemohormonal treatment (Estracyt) using (BPH) subjects as controls. In BPH patients (12 subjects) the mean CgA serotest value (+/- SD) was 23.3 + 13.9 ng/ml (range 7.2-55.9). In responders (24) and nonresponders (14) to Mab, respective values were 39.5 + 18.3 (7.6-78.4) and 214.8 + 250.3 (9.9-1084.3), while in responders (19) and nonresponders (12) to Estracyt, the mean CgA level was 47.6 +/- 22.7 (4.4-101.2) and 366.7 +/- 291.4 (82.0-925.7). In all patients osseous metastates were detected. Statistical P-values were > 0.05 in the correlation between both responders to hormonal therapy and Estracyt (P = 0.194) and nonresponders to these two therapies (P = 0.179). All other recorded P-values were < 0.01. In some nonresponders to Estracyt (3/12) the normal total PSA value together with %FPSA well below 20 indicated contribution of anaplastic tumor. Cessation of Estracyt therapy in 4/14 responders caused the sharp elevation of CgA serotest level. Accordingly, Estracyt may control the activity of CgA positive structure(s). In accordance with these preliminary data, we advocate the CgA serotest assessment in candidates for hormonal therapy as well as during follow-up of pharmacological treatment. PMID- 10697638 TI - Percutaneous implantation of arterial Port-a-cath via trans-subclavin access. AB - BACKGROUND: Hepatic artery infusion is the best choice of treatment for colorectal liver metastases, but it could be suggested for other hepatic tumors or locally advanced pancreatic cancer. The need of a laparotomy for the positioning of the arterial catheter has been the limiting factor for the diffusion of regional treatments. MATERIALS AND METHODS: 170 patients suffering from primary or secondary liver tumours and pancreatic or bile ducts cancer, underwent the positioning of intra-arterial hepatic part-a-cath by a transcutaneous subclavian access in local anaesthesia. In 163 patients, a catheter was placed into the hepatic artery, 4 into the splenic and 3 into the gastroduodenal artery. RESULTS: The procedure was performed successfully in all patients. We observed 5 aneurysms of the subclavian artery and 9 thrombosis of the hepatic artery. Only in 7 patients was arterial infusion suspended for technical complications. We observed 10.6% of dislocation, but dislodged catheters were always moved again into the hepatic artery. CONCLUSIONS: The development of percutaneous techniques of arterial port-a-cath implantation could enlarge the indication of regional chemotherapy. PMID- 10697639 TI - Epidemiology of HIV/AIDS in children. AB - HIV infection has been a major cause of morbidity and mortality since the first cases of AIDS among children were reported in 1982 in the United States. Considerable advances, especially in the past 5 years, in the understanding of the pathogenesis, diagnosis, treatment, monitoring, and prevention of HIV infection in children have changed the rate of pediatric HIV infection in the United States. Efforts to maximally decrease perinatal HIV transmission in the United States are ongoing. Physicians must try to prevent HIV infection among women, especially adolescents. PMID- 10697640 TI - Update on perinatal HIV transmission. AB - Over the past decade, much progress has been made in understanding the risk factors and timing of perinatal HIV transmission. Even more impressive have been the successful clinical trials with antiretrovirals, such as ZDV, ZDV-3TC, and nevirapine, that demonstrated significant reductions in the risk for infant infection. Within the United States and Europe, these trial results have led to rapid implementation and dramatic decreases in new perinatal HIV cases since 1994. An immediate challenge is to rapidly translate the short-course antiretroviral trial results with ZDV and nevirapine into public health policy and practice in resource-poor settings, where almost 600,000 neonates continue to become infected by mother-infant HIV transmission each year. Physicians must also test strategies to further decrease the risk for infant HIV infection during the breast-feeding period. PMID- 10697641 TI - Diagnosis of HIV infection in children. AB - Many advances have been made in the area of HIV diagnostics. Commercially available virologic assays are sensitive and specific for the early detection of HIV in perinatal infection. The timing of the transmission of HIV from mother to child (in utero, at the time of birth, or postnatally by breast-feeding) is a critical consideration in the appropriate diagnosis of infants. Several algorithms can be used to define early infection and the potential timing of acquisition of infection that combine different assays and timing of specimens. The use of virologic assays, including HIV DNA PCR and HIV RNA detection methods and culture, can define and rule out infection in infants less than 18 months of age. Serologic diagnostic methods, including HIV ELISA, immunofluorescence, and western blot assays, can be used to diagnose infants more than 18 months of age, when transplacental antibody has disappeared in uninfected HIV-exposed infants. The challenge of the early and accurate diagnosis of perinatally HIV-exposed infants is the use of new assays to detect different HIV subtype infections that are prevalent in developing countries. Rapid, simple, and inexpensive serologic and virologic assays are being developed for worldwide use. PMID- 10697642 TI - Viral and immunopathogenesis of vertical HIV-1 infection. AB - High-level viral replication is the primary determinant of CD4 depletion or disease development in HIV-1--infected children. The developing immune system of infants might allow for more efficient viral replication and less efficient immune containment of viral replication. Advances in the understanding of the pathogenesis of vertical HIV-1 infection suggest that the use of potent combination regimens to control HIV-1 replication offers the best opportunity to prevent or reverse the sequelae of HIV-1 infection. PMID- 10697643 TI - Opportunistic infections and other clinical manifestations of HIV disease in children. AB - As the decade draws to a close, physicians can be cautiously optimistic about the prevention and treatment of opportunistic infections in children with HIV disease. As more children receive therapy with powerful antiretroviral regimens, fewer are likely to be at risk for opportunistic pathogens. The widespread use of protease inhibitor combination therapies has already resulted in a dramatic decrease in morbidity and mortality in the population of HIV-infected adults. The same effect has been seen at pediatric care centers throughout the United States. Clinicians caring for HIV-infected children are now considering the safety of discontinuing prophylactic therapies for children with sustained immunologic improvement on antiretroviral therapy. For children who remain at risk, prophylactic regimens for PCP and MAC have been shown to decrease the risk for these infections. Preventive regimens for several other opportunistic infections are also available. The understanding of the pathogenesis of HIV and many of the opportunistic pathogens has led to the development of a variety of efficacious therapies for these infections. Despite these advances, physicians can anticipate that HIV-infected children will continue to develop opportunistic infections and other related complications. Some children fail to respond to antiretroviral therapies, whereas others are unable to tolerate the complex medication regimens. Prophylactic therapies are not 100% protective and, despite improved treatments, few opportunistic infections are cured. Most require lifelong maintenance therapy in the absence of immune reconstitution. Drug interactions, complex dosing schedules, adverse side effects, and high costs further limit the efficacy of these therapies. The prophylaxis, diagnosis, and treatment of opportunistic infections are likely to remain integral components of HIV care for the near and distant future. PMID- 10697644 TI - Organ-specific manifestations of HIV disease in children. AB - The clinical manifestations of HIV disease in children affect multiple organ systems. The severity of each manifestation varies by organ system and can be related in many cases to multifactorial causes, namely HIV replication in affected tissue, concomitant opportunistic infection of the organ, effect of concurrent immunodeficiency or autoimmune mechanisms on the organ, or adverse end organ drug effect (primary HIV therapy or prophylaxis regimens). More information is needed to understand the pathogenesis of the systemic effect of HIV on different organ systems, especially the CNS. Most clinicians hope that advances in therapeutic interventions for primary HIV will halt the progression of the organ-specific manifestations that have been outlined in this article, but such potent therapies will probably have their own unique and new effects on HIV infected organ systems. Vigilance for organ-specific manifestations in the era of HAART is imperative to provide the best clinical outcome for HIV-infected children. PMID- 10697646 TI - Antiretroviral therapy of HIV infection in children. AB - Much progress has been made in the therapy of pediatric HIV infection, which has been transformed from a usually fatal disease into that of a chronic disease model. Early, aggressive therapy with the goal of complete suppression of viral replication (undetectable plasma virus) should be the therapeutic goal, but this new, more hopeful environment has been created at the cost of complexity and compromises in quality of life. The rapid pace of new developments and therapeutic complexities argue strongly for care in specialized centers or, at least, frequent consultation. Efforts are ongoing to develop simpler, more effective therapeutic regimens that suppress and ultimately eradicate infection and that stimulate immune reconstitution. PMID- 10697645 TI - Medical management of HIV disease in children. AB - Significant advances have been made in the understanding of the pathophysiology of HIV infection since the beginning of the epidemic. This knowledge has translated into the development of new therapies for HIV and opportunistic infections, laboratory advances in monitoring viral and immune status, and a better understanding of factors affecting patient outcome. Concomitantly, significant progress has been made in the medical management of children with HIV infection in the past 5 years. The number of children reported with AIDS in the United States is decreasing, and efforts are shifting from caring for children with advanced immunosuppression and severe opportunistic infections to early HAART, maintenance of the immune system, and prevention of opportunistic infections. Primary care physicians are now more involved and informed in the care of HIV-infected patients. Although published data are limited, physicians who have been working with this population have observed a dramatic improvement in the quality of life and length of survival of these patients. Unfortunately, this progress is not shared by developing countries where resources are minimal and antiretroviral agents are commonly unavailable. Although efforts to develop a vaccine to prevent HIV infection are ongoing, progress has been slow. Education and awareness continue to be the most powerful weapons against HIV. PMID- 10697647 TI - HIV and AIDS in adolescents. AB - HIV infection in adolescents continues to challenge health care providers, policy makers, and advocates for youth. Primary care providers working with parents of adolescents and at-risk youth are in a unique position to identify or help develop HIV prevention and care programs that address many needs. Effective interventions are those that move beyond moralism to realism and a willingness to engage youth and their families. Youth at high risk for HIV should be identified and engaged in primary care as soon as possible. HIV-infected youth need intensive individual and group interventions to keep themselves healthy and reduce transmission to others. Incumbent on all providers is to make adolescents' services visible, flexible, affordable, confidential, culturally appropriate, and available for all youth. PMID- 10697648 TI - Caring for the child and family with HIV disease. AB - Caring for children with HIV infection is a much more optimistic process than in the beginning of the epidemic. Antiretroviral therapies are available, and additional drugs are receiving approval from the US Food and Drug Administration. Cautious optimism must be tempered with an understanding that living with the disease is a complicated and daunting process for these children and their families. Although scientific knowledge and medical treatments are moving forward, the social and environmental uncertainties remain for families. Comprehensive care is a balance of health care services and supportive, community based services offered in a compassionate manner. PMID- 10697649 TI - Management of HIV-infected children in the home and institutional settings. Care of children and infections control in schools, day care, hospital settings, home, foster care, and adoption. AB - The likelihood of high-risk pediatric exposure to HIV infection, other than perinatal exposure, has been shown to be low in most cases, and HIV PEP should be considered on a case-by-case basis. Generic considerations in the management of children who have become HIV infected emphasizes the principles of inclusion, maintaining confidentiality of a child's HIV status, and notifying those who need to know about the HIV status to care properly for the child or adolescent. Although appropriate infection-control precautions are applicable for all children and for many pathogens, children especially HIV-infected children, exposed to such pathogens, must be managed in a timely fashion. In many cases, recommendations that are applicable in one setting are applicable in others. Some exceptions apply, including infection-control precautions in hospitals versus other settings. A few additional considerations have been made for special settings and activities, including adoption, foster care, athletics, summer camp, and other recreational activities. PMID- 10697650 TI - Prevention of HIV infection in children. AB - The threshold of a new century is an opportune time to review advances in the prevention of HIV infection in children. In the United States, progress in the ability to virtually eliminate perinatal HIV transmission that was unthinkable just a few years ago has been achieved. Clinicians providing care to pregnant women should educate and counsel women about HIV and strongly recommend that they be tested. They should also counsel HIV-infected women about the means available to substantially decrease the risk for HIV transmission to their infants (e.g., antiretroviral drug use, avoidance of breast-feeding, elective C-section, encouraging pregnant women to use barrier methods during sexual intercourse, and to discontinue injection drug use). This article has highlighted some of the remaining challenges that constitute barriers to achieving maximal decrease of HIV infection in children. Studies conducted in resource-poor countries have added greatly to the understanding of vertical transmission of HIV, and they are now leading to practical and affordable approaches to reduce vertical HIV transmission world-wide. The results of this research must lead to coordinated public health action and a global political commitment to extend the benefits of antiretroviral drug prophylaxis that now exist widely in the United States to more resource-poor countries. PMID- 10697651 TI - [The histology of the primary tumor and metastatic behavior. The implications for the radio-oncologist with examples]. AB - BACKGROUND: In the past, little attention has been given to the relationship of the histology of a primary tumor to its possible subsequent pattern of metastasis. The knowledge of these relations, however, is of importance for the radio-oncologist for several reasons. MATERIALS AND METHODS: A review on the relation between the histology of a primary tumor and the pattern of subsequent metastasis has been worked out. The review is based on references personally collected during the past 25 years, a MEDLINE screen (1988 to 1997) and own analyses related to the topic. RESULTS: Strong relationships seem to exist between the histology of certain primary tumors and their subsequent pattern of metastasis. Examples demonstrate those relations which are presently known about the main solid tumors of the upper part of the body and the skin. Some implications for the radio-oncologist were pointed out. CONCLUSIONS: Besides a better understanding of a disease, the use of these data may be helpful for treatment planning and decision findings in conformal radiotherapy. Moreover, the data are of considerable importance to the follow-up of patients: imaging procedures may be more accurately applied in the areas with a high probability of recurrence or metastasis. PMID- 10697652 TI - Effect and toxicity of endoluminal high-dose-rate (HDR) brachytherapy in centrally located tumors of the upper respiratory tract. AB - AIM: To assess effect and toxicity of high-dose-rate afterloading (HDR) alone or in combination with external beam radiotherapy (EBRT) in centrally located tumors of the upper respiratory tract. PATIENTS AND METHODS: From 1987 to 1996, 55 patients were treated. Twenty-one patients (group A1: 17 non-small-cell lung cancer [NSCLC], A2: 4 metastases from other malignancies) were treated using HDR alone due to a relapse after external beam irradiation. In 34 previously untreated and inoperable patients (group B1: 27 NSCLC, B2: 7 metastases from other malignancies) HDR was given as a boost after EBRT (30 to 60 Gy, median 50). HDR was carried out with a 192Ir source (370 GBq). The brachytherapy dose (group A: 5 to 27 Gy, median 20; B: 10 to 20 Gy, median 15) was prescribed to 1 cm distance from the source axis. A distanciable applicator was used in 39/55 patients. RESULTS: In group A1, a response rate (CR, PR) of 53% (group B1: 77%) was reached. The median survival (Kaplan-Meier) was 5 months in group A1 (B1: 20 months). The 1-, 3- and 5-year local progression free survival rates (Kaplan Meier) were 66% (15%), 52% (0%), and 37% (0%) in group B1 (group A1). Prognostic favorable factors in group B1 were a tumor diameter < 20 mm, the lack of radiological mediastinal involvement, a complete remission, and a Karnofsky performance status > 70. Grade-1 or 2 toxicity (RTOG/EORTC) occurred in 0% in group A and in 6% in group B. We observed no Grade-3 or 4 toxicity. Complications caused by persistent or progressive local disease occurred in 3 patients in group A (fatal hemorrhage, tracheomediastinal fistula, hemoptysis) and in 2 patients in group B (fatal hemorrhage, hemoptysis). CONCLUSIONS: HDR brachytherapy is an effective treatment with moderate side effects. In combination with external beam irradiation long-term remissions can be reached in one third of the patients. PMID- 10697653 TI - Combined radiochemotherapy with docetaxel in patients with unresectable locally advanced head and neck tumors. AB - BACKGROUND: As the treatment with docetaxel in metastatic head and neck cancer resulted in an encouraging response rate, the following phase-I study examined the effects of a combined radiochemotherapy with weekly docetaxel in patients with inoperable advanced head and neck tumors. PATIENTS AND METHODS: Six patients with Stage IV head and neck cancer were included into the study. Within the treatment regimen the primary tumor and the involved lymph nodes were irradiated up to a total dose of 70 Gy, the non involved cervical and supraclavicular lymph nodes received 50 Gy in conventional fractionation. Simultaneously docetaxel was given 1 hour before radiotherapy. The initial dose was 15 mg/m2. RESULTS: A dose escalation was impossible because of several dose limiting toxicities (NCI-CTC) already in the first dose level. Two patients showed skin reactions Grade 4, 2 patients pulmonary complications Grade 4, 2 patient neurologic side effects Grade 3 and 1 a thrombocytopenia Grade 3. The response rate resulted in 3 complete and 1 partial remission, 1 death, 1 patient was not evaluable. CONCLUSION: Unexpectedly already in the first dose level several dose limiting toxicities were evaluated. For that reason the treatment scheme is not feasible. PMID- 10697654 TI - Prognostic value of hemoglobin concentrations in patients with advanced head and neck cancer treated with combined radio-chemotherapy and surgery. AB - PURPOSE: Hemoglobin levels are currently the focus of interest as prognostic factors in patients with head and neck cancer. Most published clinical trials have confirmed hemoglobin to possess a significant influence on survival in patients treated with radiotherapy. In our study we have investigated the prognostic value of hemoglobin in a combined modality schedule. PATIENTS AND METHODS: Forty-three patients with advanced head and neck tumors were treated with combined radio-chemotherapy. The therapy comprised 2 courses of induction chemotherapy with ifosfamide (1,500 mg/m2, day 1 to 5) and cisplatin (60 mg/m2, day 5) followed by hyperfractionated accelerated radiotherapy with a total dose of only 30 Gy. Surgery involved tumor resection and neck dissection. RESULTS: The 1-year overall survival rate and the 2-year survival rate were 79% and 56%, respectively. The 1- and 2-year recurrence-free survival rates were 68% and 49%, respectively. Prognostic factors with an impact on survival were seen in tumor size (T3 vs T4, p = 0.0088), response to radio-chemotherapy at the primary site (no vital tumor rest vs vital tumor rest, p = 0.045), response to lymph node radio-chemotherapy (no vital tumor cells vs vital tumor cells, p = 0.013) and level of hemoglobin after radio-chemotherapy (Hb > or = 11.5 g/dl vs < 11.5 g/dl, p = 0.0084). CONCLUSION: In our study hemoglobin level after radio-chemotherapy was identified for the first time to be also a significant prognostic factor (univariate analysis) in head and neck cancer patients who underwent combined radio-chemotherapy. Besides chemotherapy plus low-dose irradiation achieved similar results in comparison with radical resection and high-dose radiotherapy at least for the first 2 years after therapy. Relapsing disease could be treated with 1 additional course of radiotherapy which is supposed to be well tolerated. PMID- 10697655 TI - Comparison of radiation effects of gadolinium and boron neutron capture reactions. AB - PURPOSE: Cell survival assays were performed to evaluate the effects of radiations released during neutron capture reactions by gadolinium-157, boron-10 and by the combination of both. MATERIALS AND METHODS: Single cell suspensions with or without Gd-157 and/or B-10 were exposed to thermal neutrons produced by the Kyoto University reactor, and standard cell survival curves were obtained. RESULTS: Under the same molarity, cytocidal effects were 1.5 times greater for Gd 157 than for boron when compared at 10% survival levels. The presence of B-10 enhanced the radiation effect of Gd-157 neutron capture by 1.2-fold, suggesting that cells were not sufficiently irradiated as a result of neutron fluency attenuation by the presence of excess neutron capture agents in the medium. CONCLUSIONS: When an equal number of atoms were present, Gd-157 was effective as B-10 when exposed to an equal number of thermal neutrons. However, there was no benefit observed in the combination of Gd-157 and B-10 for neutron capture therapy. Further studies are needed to determine optimal Gd-157 and B-10 concentrations as a function of tumor dimension. PMID- 10697656 TI - [Improved tumor contrast and delineation in the stereotactic radiotherapy planning of cerebral gliomas and metastases with contrast media-supported FLAIR imaging]. AB - BACKGROUND: FLAIR MR imaging has shown to be a valuable imaging modality in pathologic lesions of the brain including intra-axial brain tumors. The aim of the study was to assess the value of a FLAIR technique in the planning process of stereotactic radiotherapy in patients with cerebral gliomas and metastases. PATIENTS AND METHODS: Thirty-five patients with cerebral gliomas and 12 patients with a total of 39 cerebral metastases were examined by T2/PD-weighted fast spin echo, fast FLAIR prior and after contrast and contrast enhanced T1-weighted spin echo using identical slice parameters. The images were evaluated by using quantitative and qualitative criteria. Quantitative criteria were tumor-to background and tumor-to-cerebrospinal fluid contrast and contrast-to-noise. The qualitative evaluation was performed as a multireader analysis concerning lesion detection, lesion delineation and image artifacts. RESULTS: In the qualitative evaluation (Table 3 and 6), all readers found the fast FLAIR images to be superior to fast spin-echo in the exact delineation of cerebral tumors (p < 0.001) and the delineation of enhancing and non enhancing tumor parts. Fast FLAIR was superior in the delineation of cortically located and small lesions but was limited in lesions adjacent to the ventricles. Fast FLAIR provided a significantly better tumor-to-CSF contrast and tumor-to-CSF contrast-to-noise (p < 0.001) (Tables 1, 2a, 2b, 4, 5). The tumor-to-background contrast and tumor-to background contrast-to-noise of the fast FLAIR images were lower than that of T2 weighted spin-echo images but were significantly increased after the application of contrast media. FLAIR images had more image artifacts, but the image interpretation was not influenced. CONCLUSIONS: FLAIR MR imaging was found to be a valuable sequence in the planning protocol of stereotactic radiotherapy. The concurrent presentation of enhancing and non enhancing tumor tissue on contrast enhanced fast FLAIR imaging enables to use a single imaging sequence in the treatment protocol. This enables to load a reduced image amount into the radiotherapy planning software, is therefore time saving and reduces potential errors. PMID- 10697657 TI - [The inadequacy of computed tomography for the assessment of patients with esophageal carcinomas after neoadjuvant radiochemotherapy]. PMID- 10697658 TI - [Can a total dosage of 30 Gy be adequate for patients with anal canal carcinomas after excision biopsy and subsequent radiochemotherapy?]. PMID- 10697659 TI - [The efficacy of surgical salvage therapy in patients with locally uncontrolled anal carcinoma following sphincter-preserving treatment]. PMID- 10697660 TI - [A chemical inhibitor of p53 that protects mice from the side effects of cancer treatment]. PMID- 10697661 TI - Novel approaches to the treatment of nausea and vomiting. AB - Nausea and vomiting are debilitating symptoms complicating many clinical conditions. Conventional antiemetic agents act as muscarinic, histamine, and dopamine receptor antagonists in the central nervous system. In a retrospective analysis, tricyclic antidepressant drugs demonstrated efficacy in long-term treatment of functional nausea. Some cases of vomiting result from impaired gastrointestinal motor activity. Agents which act on gastric serotonin (5-HT4), dopamine, and motilin receptors accelerate gastric emptying and relieve symptoms in gastroparesis. Recent investigations suggest that some patients with refractory gastroparesis may benefit from gastric electrical pacing. The treatment of acute chemotherapy-induced emesis was revolutionized by 5-HT3 receptor antagonists; however, these agents are less efficacious in delayed vomiting. Neurokinin (NK-1) receptor antagonists show promise in treating delayed chemotherapy-evoked emesis. Furthermore, animal studies indicate a broad spectrum of action for NK-1 antagonists in treating diverse causes of nausea and vomiting. The cyclic vomiting syndrome is characterized by discrete episodes of relentless vomiting separated by asymptomatic intervals and is associated with migraine headaches. Antimigraine therapies including the 5-HT1D receptor agonists sumatriptan reduce the severity of cyclic vomiting attacks. Investigations into these and other novel treatments may provide important advances in the care of difficult cases of nausea and vomiting resulting from disparate illnesses. PMID- 10697663 TI - Therapeutic options in patients with Barrett's esophagus. AB - The rising incidence of esophageal adenocarcinoma has focused attention on the only known risk factor: Barrett's esophagus. Practice guidelines for the diagnosis, surveillance and management of Barrett's esophagus were published recently. Although the ultimate goal in the management of this premalignant condition would be the permanent elimination of Barrett's mucosa, current therapeutic options are limited or still in the investigational stages. This review summarizes current medical and surgical treatment options and introduces endoscopic ablative modalities currently under investigation. PMID- 10697662 TI - Biliary complications of orthotopic liver transplantation. AB - Biliary complications are a common cause of morbidity following orthotopic liver transplantation. Complications involving the biliary tree occur after 6-34% of all liver transplants performed, usually within the first 3 months after transplantation. Bile leaks and biliary strictures are the most common biliary complications, but sphincter of Oddi dysfunction, hemobilia, and biliary obstruction from stones, sludge, or casts have also been described. The risk of specific biliary complications is related to the type of biliary reconstruction performed at the time of transplantation. In this article, we review the major types of biliary reconstruction and their associated biliary complications. Specific risk factors for the development of biliary complications are outlined. Finally, the management of biliary complications is discussed, with an emphasis on the role of endoscopic therapy. PMID- 10697664 TI - Adenocarcinoma of the esophagogastric junction. AB - Adenocarcinoma of the esophagogastric junction (EGJ) has increased rapidly in incidence in the latter half of the twentieth century. The increase in incidence has affected white men between the ages of 40 and 60 disproportionately. Understanding the etiology and improving treatment requires careful classification of EGJ tumors. A recent consensus conference recognized three types of EGJ adenocarcinomas: distal esophageal, cardia, and subcardia gastric. Distal esophageal adenocarcinomas are associated with Barrett's esophagus. Helicobacter pylori infection may play a role in some adenocarcinomas of the subcardia, but the association is unproven. Therapy for all types of EGJ tumors is surgical, but multimodal forms of treatment are commonly used because of the advanced stage at which these tumors often present. Several endoscopic options exist for primary therapy of early-stage tumors and for palliation. PMID- 10697665 TI - Update in medical therapy in inflammatory bowel disease: a clinician's view. AB - Recent advances in the therapy of inflammatory bowel disease specifically directed against the inflammatory and immune mechanisms include an impressive and often overwhelming cornucopia of anti-inflammatory agents, immunomodulators, antibiotics, biologicals, topical therapies, nicotine, heparin, and nutritional supplements. The interface of one drug regimen into another may lead to confounding and often confusing programs of treatment. This review will attempt to offer a perspective of care and an update of specific remedies, but the aim is practicality and usefulness, not encyclopedic detail. PMID- 10697666 TI - Congenital esophageal stenosis: clinical and endoscopic features in adults. AB - BACKGROUND: Congenital esophageal stenosis (CES) is an uncommon anomaly that reportedly rarely goes undiagnosed until adulthood. One variant of CES includes patients with multiple cartilaginous rings described usually in the mid-distal esophagus. METHODS: Ten patients with CES seen over the past 7 years were interviewed and their clinical and endoscopic records reviewed. RESULTS: Eight patients were male and age at time of diagnosis ranged from 21 to 75 years. Meat impaction was the presenting symptom in 8 patients and 3 patients reported a relapsing history. Intermittent solid food dysphagia over extended duration (10 40 years) was reported in all patients with an estimated onset of symptoms at a mean age of 27 years (11-52 years). Endoscopically, all patients had segmental esophageal stenosis (length 2-8 cm, mean = 4.7 cm) due to 'trachea-like' multiple submucosal rings. Pseudodiverticulosis and distal reflux esophagitis were evident in 1 patient. Nine of 10 patients had no macroscopic esophagitis. Dilatation was performed by balloon insufflation (18 mm in 5 patients, 15 mm in 3 patients, 12 mm followed by 15 mm in a patient with a tight stricture) and by Savary dilators in 1 patient, without any complications. No patient had recurrent meat impaction on follow-up (1-7 years, mean = 3 years) after education about the condition. CONCLUSION: (1) We suspect CES is an under-recognized cause for intermittent, long-standing dysphagia in adults. (2) Food impaction is a frequent initial presentation. Recognition of CES is critical for appropriate patient education and planning. PMID- 10697667 TI - Achalasia presenting with hydropneumothorax. PMID- 10697668 TI - Active esophageal variceal bleeding treated with band ligation. PMID- 10697669 TI - Chromoendoscopy facilitates the identification of adenomatous polyps arising on the ileocecal valve. PMID- 10697670 TI - Pharmacist knowledge of common herbal preparations. PMID- 10697671 TI - Pharmacokinetic/pharmacodynamic modeling of theophylline in patients with different degrees of airway obstruction. PMID- 10697672 TI - Simulated microgravity impairs vascular contractility: role of nitric oxide dependent vasodilator mechanisms. PMID- 10697673 TI - Simulated microgravity-induced vascular hyporesponsiveness: role of signal transduction. PMID- 10697674 TI - The relationship between reactivity of metabolites of pyrrolizidine alkaloids and extrahepatic toxicity. AB - Pyrrolizidine alkaloids (PAs) are a large group of structurally similar toxins. In animals, including man, they are hepatotoxic and in some cases pneumo- and neurotoxic. PAs are metabolized by the liver P450 system to reactive dehydroalkaloid (DHA) intermediates. PA toxicity is a result of alkylation of macromolecules by DHAs. We have measured the relative reactivity of a series of semi-synthetic DHAs by recording the rate at which they alkylate a model nucleophile, 4-(p-nitrobenzyl)pyridine. Rate data fit mono- or biexponential equations. Rank order of reactivity for the macrocyclic and open ester DHAs was the same as those measured for DHA hydrolysis. The reaction with 4-(p nitrobenzyl)pyridine was easier to follow, however, as rates of reaction can easily be controlled by temperature or level of acid catalysis, and the final product can be measured colorimetrically. DHAs of the primarily hepatotoxic alkaloids, retrorsine and seneciphylline, were more reactive than DHAs of monocrotaline and trichodesmine, which additionally produce pneumo- and neurotoxicity, respectively. This suggests that DHAs with greater stability (longer half-lives) are able to survive long enough to reach target organs downstream form the liver. We believe that differences in PA metabolism and the nature of toxicity ultimately produced are in part related to differences in reactivity of the primary toxic intermediate, the DHA. PMID- 10697675 TI - Dose-regimen dependent caffeic acid prevention of acute liver damage. PMID- 10697676 TI - Nitric oxide and prostaglandin interactions in the relaxant responses of rat aorta: influence of pregnancy. PMID- 10697677 TI - Segmental differences in rat aorta contraction induced by phenylephrine in aortic rings. PMID- 10697678 TI - Potentiation by L-cysteine of N-methyl-D-aspartate receptor: effects on intracellular free Ca2+ in cultured cerebellar granule cells. PMID- 10697679 TI - Anticonvulsant taurine derivatives modify taurine and GABA release in the mouse hippocampus. PMID- 10697680 TI - Effect of glycine on hemoglobin glycation in diabetic patients. PMID- 10697681 TI - Prevention of gastroduodenal injury induced by NSAIDs with low-dose misoprostol. PMID- 10697682 TI - Inhibition of hemoglobin glycation with glycine in induced diabetes mellitus in rats. PMID- 10697683 TI - Effect of 5-HT-moduline, an endogenous peptide, in associative learning. PMID- 10697684 TI - Inhibition of leukocyte rolling by submandibular gland peptide-T (SGP-T). PMID- 10697685 TI - Evidence for spare beta-adrenoceptors in the rabbit heart. PMID- 10697686 TI - Cardiovascular actions of vanadate in the genetically obese Zucker rat. PMID- 10697687 TI - Endothelium-dependent alterations in responses of the mesenteric vascular bed in the streptozotocin diabetic rat. PMID- 10697688 TI - Mechanism for pentobarbital-induced enhancement of synaptic activity may be similar to that for long-term potentiation (LTP) in the mammalian CNS. PMID- 10697689 TI - Activation of adenosine A1 receptors facilitates the analgesic effect of ketorolac and ketorolac-caffeine combinations in the rat. PMID- 10697690 TI - Purinergic and calcium-mediated enhancement of NGF-induced neurite expression in PC12 cells. PMID- 10697691 TI - Dynamic control of the release of a hepatic insulin-sensitizing substance. PMID- 10697692 TI - Tyrosine is detrimental to the biological activity of submandibular gland peptide T (SGP-T). PMID- 10697693 TI - Interaction of clozapine and other antipsychotic drugs with human alpha 1 adrenergic receptor subtypes. PMID- 10697694 TI - The histological effect of the human chorionic gonadotropin and luteinizing hormone-releasing hormone on experimental cryptorchidism in rats. PMID- 10697695 TI - Morphological changes produced by acute prenatal exposure to ethanol on the immunoreactive vasoactive intestinal polypeptide cells of the suprachiasmatic nucleus of the rat. PMID- 10697696 TI - The effect of streptokinase-streptodornase administration on histological damage following testicular torsion. PMID- 10697697 TI - Expression of P2y and P2x receptors in cultured human microglia. PMID- 10697698 TI - Cardiomyopathic changes in streptozotocin-induced diabetes. PMID- 10697699 TI - Design of mimetics of gonadotropin releasing hormone (GnRH) with effects on proliferation of breast cancer cells. PMID- 10697700 TI - Analgesic effect of sodium diclofenac plus vitamin B complex in the PIFIR model. PMID- 10697701 TI - Cervical plexus block with ropivacaine for neck surgery. PMID- 10697702 TI - Modulation of phospholipid metabolism, leukotrienes and prostaglandin synthesis by local anesthetic agents. PMID- 10697703 TI - Protein calorie malnutrition induces decreased macrophage NF-kappa B expression. PMID- 10697704 TI - Continuous thoracic epidural ropivacaine drips for multiple rib fractures. PMID- 10697705 TI - Successful treatment of reflex sympathetic dystrophy by bier block with lidocaine and clonidine. PMID- 10697706 TI - Association between cephalosporin therapy and subsequent nosocomial enterococcal colonization in surgical patients. PMID- 10697707 TI - Blockade of neuronal sodium channels by the antiepileptic drugs phenytoin, carbamazepine and sodium valproate. PMID- 10697708 TI - Studies on muscarinic M2 receptors in smooth muscle. AB - Histamine after M3 receptor alkylation with 4-DAMP mustard does not serve simply as a spasmogen but facilitates visualization of the M2-mediated contraction induced by oxotremorine M. We speculate that M3 receptor activation has a similar role in untreated tissue. The facilitation appears to involve protein kinase C. The results with propranolol suggest that the phospholipase D pathway may also be important in the development of the M2-mediated response. PMID- 10697709 TI - Inflammatory mediator-induced modulation of TTX-R INa: an underlying mechanism of inflammatory hyperalgesia. PMID- 10697710 TI - A case study in pharmacogenomics: leptin preclinical and clinical development. PMID- 10697711 TI - Beta 3-adrenoceptors: their role and regulation in the gastrointestinal tract. PMID- 10697712 TI - Current status of P2X receptors: distribution and pathophysiological roles. PMID- 10697713 TI - Developing targets for drug discovery using genome-wide gene trapping and ultra sensitive beta-lactamase reporters. PMID- 10697714 TI - Pharmacology--a unique discipline for unique questions? AB - This article discusses the nature and responsibilities of the biomedical subject known as pharmacology. It will attempt to show how pharmacology is a very special subject that is uniquely positioned to ask fundamental questions about drugs, such as how they produce their actions, and how they may be used more effectively in man. Furthermore, pharmacology is a most important subject to the pharmaceutical industry as it is pharmacology, combined with medicinal chemistry and toxicology, that is responsible for the introduction of the vast majority of new drugs. Thus, pharmacologists are to be found in universities, research institutes and industry. In universities, pharmacologists are concerned more with discovering new drug-like entities in the body and natural world and in understanding how current drugs work. In industry, pharmacologists seek new drugs. Over the last decade there have been many discoveries in molecular biology, the new biology. The enthusiasm and revolutionary spirit which greets such discoveries leads to an assumption by some that pharmacology in its present form has been rendered redundant. This article refutes such a conclusion and shows how pharmacology has unique attributes and approaches that are as important now as they always were. Indeed, it argues for improvements in the understanding of pharmacology and how it remains a unique and special subject that should be nurtured. PMID- 10697715 TI - The future of pharmacology in the 20th century: a 19th century prediction. PMID- 10697716 TI - The future of pharmacology in Canada. PMID- 10697717 TI - Mexican pharmacology at the dawn of the new millennium: achievements and challenges. PMID- 10697718 TI - Pharmacology in the 21st century in the United States of America. PMID- 10697719 TI - Academic pharmacologists: meeting the educational needs of graduate programs and the health care professions. PMID- 10697720 TI - A brief history of pharmacology, therapeutics and scientific thought. PMID- 10697721 TI - Mossbauer studies on laser evaporated iron atoms and their reactions with oxygen in argon matrices. AB - Laser-evaporated iron atoms were isolated in low-temperature Ar matrices and their chemical reactions with oxygen were investigated by means of Mossbauer spectroscopy. Reactions of iron atoms with oxygen produce FeO, Fe(O2), FeO3, (O2)FeO2 and OFeO isolated in the Ar matrices and their yields vary depending on the concentration of oxygen. Similarly, FeO and Fe(O2) were obtained by the reaction of iron atoms with N2O. Infrared spectroscopy and molecular orbital calculations were applied to support their assignments. PMID- 10697722 TI - Sorption and desorption of Eu and Yb on alumina: mechanisms and effect of fulvic acid. AB - The effects of pH, ionic strength and FA (fulvic acid) on the sorption and desorption of Eu(III) and Yb(III) on alumina were respectively investigated by using batch technique and radiotracers 152 + 154Eu and 169Yb. The distribution coefficients for sorption and desorption of Eu on alumina at pH 4.4, 4.6 and 5.7 in 1 mol/l NaCl solutions as a function of solid phase concentration were determined in the presence or absence of FA. The effects of pH, FA and ionic strength on the distribution coefficients for sorption and desorption of Yb on alumina were determined in 0.01-2.0 mol/l NaNO3. It was found that pH and FA influenced the sorption of Eu(III) and Yb(III) on alumina greatly. A surface hydrolysis model can satisfactorily and qualitatively explain the observations on bare alumina. The competition among the complexations of surface free hydroxyl groups, soluble and sorbed fulvic acids can satisfactorily and qualitatively explain the observations on the coated alumina. PMID- 10697723 TI - Tritium in [15O]water, its identification and removal. AB - The present investigation was undertaken to identify the long-lived radionuclide and its chemical forms existing in [15O]water which was synthesized from 15O produced by the nuclear reaction 14N(d,n)15O, and to develop a method for its removal to facilitate radioactive waste disposal. The long-lived nuclide was identified as tritium based on a comparison of its physical half-life and the energy spectrum of beta-rays with those of tritium. The major chemical form of tritium in the target gas was estimated to be molecular hydrogen. The tritium radioactivity was completely removed without a serious loss occurring to the yield of [15O]water by passing the irradiated target gas over a heated palladium catalyst followed by a calcium chloride column before the final synthesis of the [15O]water. This provided a practical way of removing tritium from the [15O]water. PMID- 10697724 TI - Preparation of 124I solutions after thermodistillation of irradiated 124TeO2 targets. AB - The 4.15-d radionuclide 124I is produced via the nuclear reaction 124Te(d, 2n) 124I by irradiation of 96% enriched 124TeO2 with 14 MeV deuterons, followed by thermodistillation. In order to minimise the loss of 124I, the quartz distillation tube was fitted to a stainless steel helix capillary trap directly behind the end of the furnace. Using this device, distillation yields of more than 80% were routinely obtained, and the activity was concentrated in markedly less than 100 microL solution. The 124I produced by this method proved to be useful for labelling proteins and IUdR. PMID- 10697725 TI - Cation exchange separation of trace amounts of 203Hg and 181Hf. AB - A radiochemical method of separation of 203Hg from 181Hf is described. The method involves separation by Dowex 50W- X 8, 100-200 mesh, cation exchange resin using 1 M hydrochloric acid as the eluant. The chemical yield for the separation of mercury is > 85% and the decontamination factor is > 10(4). The 181Hf can be eluted from the column by 4 M HCl. PMID- 10697726 TI - Considerations for choice of a kinetic fig. of merit in process radiation chemistry for waste treatment. PMID- 10697727 TI - A new approach for extension of the identification period of irradiated cellulose containing foodstuffs by EPR spectroscopy. AB - We report on the possibility of identification by EPR spectroscopy of some irradiated cellulose-containing foodstuffs, at a relatively long time after the irradiation, when the characteristic EPR spectral lines of the cellulose free radical have essentially disappeared. In such cases rather expensive and time consuming methods (e.g. thermoluminescence analysis) have to be applied. The present communication demonstrates with some pre-irradiated spices, dried medicinal and sweet herbs that simply heating the samples to 60 degrees C for one hour leads to a significant (50% or more) decrease of the EPR intensity of the remaining central line of the samples. For comparison, the loss in the intensity of the same line upon heating non-irradiated samples at 60 degrees C for one hour was only about 10%. This inexpensive new procedure will extend the post irradiation period in which EPR can be used for distinguishing irradiated from non-irradiated samples, of certain cellulose-containing foodstuffs. PMID- 10697728 TI - Automatic synthesis of L-[beta-11C]amino acids using an immobilized enzyme column. AB - We have developed a system for the automatic synthesis of L-[beta-11C]amino acids for i.v. injection by means of enzyme-mediated reactions from 11CO2 via 11CH3I and D,L-[beta-11C]alanine as labeled intermediates. This system, which incorporates an ultrafilter cartridge sterilized by electron beam irradiation and a column packed with immobilized enzymes, was effective for eliminating enzymes and endotoxins that may contaminate the product. Using this system, 1.3 +/- 0.5 GBq of 5-hydroxy-L-[beta-11C]tryptophan with a radiochemical purity of 97.1 +/- 0.6% and a specific activity of 39.6 +/- 8 GBq/mumol a pH value of 4 could be obtained in about 32 min (n = 3, at EOS). No endotoxin, enzyme, or bacteria was detected in the product. L-[beta-11C]dihydroxyphenylalanine (L-[beta-11C]DOPA) was also synthesized using this system. PMID- 10697729 TI - A system for continuous production and infusion of [15O]H2O for PET activation studies. AB - A system for continuous production and infusion of [15O]H2O has been designed for Positron Emission Tomography brain activation studies. The infusion system consists of two Horizon Nxt infusion pumps, a four-port-two-position valve and a sterile 50 ml vial. The line and the back check valve between the furnace and the reservoir were heated in order to reduce vapor condensation in the line. The variation of the production of [15O]H2O was < 1%. The activity delivered as measured by scanner counts varied < 2% during the steady state period. The system has been demonstrated to be capable of delivering activity over a wide range of conditions. PMID- 10697730 TI - Production of 105Rh-EDTMP and its bone accumulation. AB - 105Rh has favorable physical characteristics as a radiotherapeutic nuclide. Carrier-free 105Rh can be produced by the neutron activation of 104Ru followed by beta decay of 105Ru. It was clarified that carrier-free 105Rh can be produced in quantities and the purity necessary for chemical and clinical investigations of its use as a nuclide for radiotherapy. 105Rh-EDTMP was simply obtained from 105Rh3+ and EDTMP by heating for 30 min in boiling water, giving a radiochemical yield of > 99%. Dissociation of radioactivity assessed by paperchromatography was negligible for up to 5 days after its preparation. In animals, 105Rh-EDTMP showed rapid blood clearance and selective uptake in the bone. Hence, 105Rh-EDTMP is thought to be a promising therapeutic agent for the treatment of pain due to bone metastases. PMID- 10697731 TI - Synthesis and structural characterization of a Re(V) complex with a 2N1S donor and the radiochemical behavior of its Tc analog. AB - A rhenium(V) complex with 2-(2-pyridylmethylthio)-aniline has been synthesized and characterized by UV-VIS absorption, IR, MS and X-ray crystallography. The complex contains a ReO3+ moiety and the amine compound, acting as a monobasic tridentate(NSN) ligand. The chemical behavior of its technetium analog has been studied radioanalytically by solvent extraction, TLC, electrophoresis and HPLC using 99mTc tracer. PMID- 10697732 TI - Efficient HPLC separation of [11C]beta-CFT or [11C]beta-CIT from an N-desmethyl precursor on a semipreparative reversed phase ODS column. AB - HPLC separation of either [11C]beta-CFT or [11C]beta-CIT from the N-desmethyl precursor greatly depended on the ODS column used and the pH of the phosphate buffer in the mobile phase. The separation was accomplished on the semipreparative reversed phase ODS column without end-capping treatment over a pH ranging from 6.4 to 9.2, but failed on the end-capped ODS column. This suggests that the presence of residual silanol groups on the ODS is an important factor for the separation. PMID- 10697733 TI - EPR and TL correlation in some powdered Greek white marbles. AB - Thermoluminescence of white powdered marble samples, chosen to display different EPR spectra, were studied. Two peaks at 280 degrees C and 360 degrees C can be observed among the TL glow curves while the EPR spectra exhibit two signals: the A signal with g perpendicular = 2.0038 and g parallel = 2.0024 due to the SO3- centre and the B one with g1 = 2.0005; g2 = 2.0001; g3 = 1.9998 due to mechanical powder reduction (drilling). Owing to heating and simultaneous experiments, a correlation have been established: the 280 degrees C TL peak is associated to the A signal and thus to the SO3- centre and the 360 degrees C TL peak is caused by mechanical treatment corresponding to the B EPR signal. PMID- 10697734 TI - Uptake of 137Cs by fresh water fish. AB - The uptake and discharge rates of 137Cs by fresh water fish at different radionuclide concentrations have been studied. A dual compartment model was used to fit the experimental data. The discharge rates have been found to be negligible for the duration of the experiment of 10 d. The uptake rates were independent of radionuclide concentrations for a particular type of fresh water fish and were different for different types of fish. The uptake rates of carp, tilapia and snakehead were 1.58, 1.66 and 2.23, in unit of 10(-6) h-1, respectively. It was also estimated that the consumption of fresh water fish, even if the water were contaminated as much as that in the Chernobyl accident, leads to negligible latent cancer fatality to the Hong Kong population. PMID- 10697735 TI - Dosimetric characterisation of a new production of MgB4O7:Dy,Na thermoluminescent material. AB - Thermoluminescent dosimetric characteristics of MgB4O7:Dy,Na are presented. MgB4O7:Dy,Na is a newly prepared TL material with attractive features for dosimetric applications, such as near-tissue equivalence, simple glow curve, high sensitivity and low fading. The dosimetric properties of this material examined in this study include glow curve shape, TL sensitivity, annealing procedure, photon dose response, minimum detectable dose, precision of TL measurements, reproducibility, energy response and fading characteristics. PMID- 10697736 TI - Analysis of volatile radiolysis products in gamma-irradiated LDPE and polypropylene films by thermal desorption-gas chromatography-mass spectrometry. AB - Low-molecular-weight ('volatile') radiolysis products of low-density polyethylene (LDPE) and polypropylene (PP) films were investigated by thermal desorption-(TDS) GC-MS after absorbed doses of up to 25 kGy. The films produce fingerprint chromatograms with highly characteristic patterns of groups of radiation-induced peaks; these are mainly hydrocarbons, aldehydes, ketones, and carboxylic acids with concentrations (after 25 kGy) ca one order of magnitude below that of the residual hydrocarbons (oligomers). PP additionally produces very substantial amounts of three degradation products of phenol-type antioxidants. The low molecular-weight (MW) radiolysis products are retained for considerable times in LDPE films and they are retained in PP films much longer than had been expected. Besides product identification, the following topics are addressed: Effects of the absorbed dose and the desorption temperature; comparison of several commercial (proprietary) films; high-temperature thermal desorption; the question whether TDS analyzes radiation-induced artifacts rather than genuine products; the possible existence of cyclic radiolysis products; the possibility of identifying an LDPE film as irradiated after a dose of only 1 kGy; and atypical trace fragments of antioxidants. Finally, the geometry and efficiency of the thermal desorption system is briefly discussed, and the implications of our findings for irradiation detection and for the safety of irradiated materials are considered. PMID- 10697737 TI - Accounting for 222Rn loss during oven drying for the immediate laboratory gamma ray spectroscopy of collected soil samples. AB - Drying soil samples in an oven to remove water alters the 222Rn emanation rate. Measurements of the oven drying 222Rn emanation rate from soil were made with a continuous radon monitor and the degree of 222Rn disequilibrium was quantified by laboratory gamma-ray spectroscopy. This paper presents a disequilibrium correction where the 226Ra activity in oven-dried soil samples is inferred from immediate laboratory gamma-ray spectroscopy of 214Bi before 222Rn and its decay progeny reach secular equilibrium. PMID- 10697738 TI - PIXE study of Cuban quaternary paleoclimate geological samples and speleothems. AB - PIXE elemental analysis of sediments, speleothems, and other geological formations related to the karst of the Sierra de San Carlos is presented. The similarity of the elemental composition of the sediments studied, as well as the alluvial regime which created them, indicate their common origin at each location. The Sr/Ca concentration ratio of a stalactite indicates that the average atmospheric temperature 12,000 and 18,000 years B.P. was colder than that of 6000 years B.P. PMID- 10697739 TI - Natural radionuclide distribution in soils of Gudalore, India. AB - The concentration of primordial radionuclides in soil samples of Gudalore Taluk in the Udagamandalam district has been measured from the gamma ray spectrum of the soil. The mean activities of 232Th, 238U and 40K are 75.3 +/- 44.1, 37.7 +/- 10.1 and 195.2 +/- 85.1 Bq kg-1 dry weight, respectively. The average outdoor absorbed dose rate in air at a height of 1 m above ground is 74.3 +/- 27.8 nGy h 1, corresponding to an annual effective dose equivalent of 455.6 microSv. The dose equivalent ranges from 168.3 to 1250.5 microSv. The results have been compared with other global radioactivity measurements and evaluations. PMID- 10697740 TI - 36th Annual meeting of the Society for Cryobiology, jointly with France Cryo and the Society for Low Temperature Biology. Marseille, France, July 12-15, 1999. Abstracts. PMID- 10697741 TI - The B chromosome system of Trypoxylon (Trypargilum) albitarse (Hymenoptera, Sphecidae) 1. Banding analysis. AB - Karyotypic analyses of 366 specimens of the solitary wasp Trypoxylon (Trypargilum) albitarse collected from ten populations in the municipalities of Vicosa and Porto Firme (Minas Gerais, Southeastern Brazil), revealed the presence of two morphological types of supernumerary (B) chromosomes. C-banding and fluorochrome banding suggest that the B chromosomes of T. albitarse may have originated from heterochromatin breaks within the standard (A) chromosome complement. PMID- 10697742 TI - Occurrence and antibiotic sensitivity of Enterobacteriaceae isolated from a group of Jordanian patients with community acquired urinary tract infections. AB - The type and antibiotic sensitivity of urinary tract pathogens may differ in various communities. Of 207 isolates recovered from midstream urine specimens collected from a group of patients with community acquired urinary tract infections (UTI), 86% were species of Enterobacteriaceae. The most frequently recovered pathogens were Escherichia coli (82%), Klebsiella spp. (7.3%), Proteus spp. (6.2%), Enterobacter spp. (3.4%) and Citrobacter spp. (1.1%). High rates of resistance were found against ampicillin (95%), tetracycline (86%), carbenicillin (84%), trimethoprim/sulphamethoxazole (48%), and amoxycillin/clavulanic acid (45%). For the antibiotics tobramycin, aztreonam, ceftriaxone and gentamicin 7% of the isolates were resistant, while resistance varied from 9-18% for amikacin, ciprofloxacin, norfloxacin, nalidixic acid and cefuroxime. The incidence of UTI caused by Enterobacteriaceae was three times higher in females than in males, particularly in young and middle age groups (< or = 19 and 20-39 years). PMID- 10697743 TI - Effect of crude seaweed extracts on seed germination, seedling growth and some metabolic processes of Vicia faba L. AB - Crude extracts from three green seaweeds (Cladophora dalmatica, Enteromorpha intestinalis, Ulva lactuca) and the three red algae (Corallina mediterranea, Jania rubens, Pterocladia pinnate) were prepared. Their effects on germination, growth of seedlings, chlorophyll content and other metabolic activities of Vicia faba were investigated. The crude extract of C. dalmatica showed maximal activity, and it increased seed germination, length of main root and shoot systems and the number of lateral roots. All the crude extracts of seaweed increased protein content in both root and shoot systems, total soluble sugars and chlorophyll content in leaves. The cytokinin content of the green algae was higher than that in red algae. Growth of seedlings of V. faba was stimulated but to different degrees. PMID- 10697744 TI - A comparative study of chromosome morphology among the nine annual species of Cicer L. AB - Thirty-six accessions, representing the full complement of all the nine annual Cicer L. species, viz C. arietinum, C. reticulatum, C. echinospermum, C. pinnatifidum, C. judaicum, C. bijugum, C. chorassanicum, C. yamashitae and C. cuneatum, were subjected to karyotype analysis for the first time in a single comprehensive study. The detailed karyotype of C. chorassanicum was also investigated for the first time. A 12 h cold water pretreatment and 13 min 60 degrees C 1 N HCl hydrolysis confirmed a somatic chromosome number of 2n = 16 in all the species. Within species interchromosomal size variation was observed to be quite large in C. arietinum, C. reticulatum and C. echinospermum, but not in the remaining six species. Individual chromosome size ranged from 3.77 microns in C. echinospermum to 1.32 microns in C. arietinum while the haploid genome length ranged from 20.65 microns in C. echinospermum to 14.92 microns in C. cuneatum. Ample rearrangement of chromatin among chromosomes within a species was implied to have played a role in Cicer genome evolution. The nine species were classified in two groups based on karyotypic similarity, with the first group comprising the inter-crossable species C. arietinum, C. reticulatum and C. echinospermum, while the remaining species forming the second group. The first group species are also genetically close to each other as deduced by other morphological, biochemical and DNA based studies. Circumstantial evidence has lead to the speculation that perhaps karyotypic similarity and interspecific crossability are positively related to each other. PMID- 10697746 TI - [Treatment recommendations for type 2 diabetes. Agence Francaise de Securite Sanitaire des Produits de Sante (AFSSAPS)]. PMID- 10697745 TI - Vascular endothelial growth factor and basic fibroblast growth factor evaluation in blood serum of patients with hormonally active and inactive adrenal gland tumours. AB - The vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) levels of 36 patients with adrenal gland tumours were analysed. The mean age of patients was 43 years (29-67 years), and there were 25 women (69.4%) and 11 men (31.6%). In 34 patients adrenalectomy was performed and in two cases lesions were considered inoperable. In all cases VEGF and bFGF were measured preoperatively and in all operated patients the level of VEGF was measured at 1 month postoperatively. A statistically significant increase in VEGF levels before surgery in comparison with the controls was recorded in all patients with adrenal tumours. No correlation between the size of a tumour and VEGF levels was observed. The serum level of VEGF decreased in patients after surgical removal of the tumour, no matter which type of tumour, with the exception of a patient showing a recurrence of cortex cancer. A statistically significant decrease was found only in patients operated on for cortex cancers and hormonally active and inactive cortex and medulla inactive benign tumours. The postoperative recurrence of the malignant tumour may be preceded by an increase in plasma VEGF levels. Such correlations were not found with bFGF. PMID- 10697747 TI - Anionic mucoadhesive polymers as auxiliary agents for the peroral administration of (poly)peptide drugs: influence of the gastric juice. AB - The incorporation of (poly)peptide drugs in mucoadhesive polymers is a promising strategy for their peroral administration. In this study, the protective effect of various polymers toward an artificial gastric fluid and the influence of an enteric coating on the adhesive properties have been investigated. Tablets containing 30 mg of carbomer (C934P), neutralized carbomer (NaC934P), or sodium carboxymethylcellulose (NaCMC), 0.1 mg of the model protein peroxidase, and 19.9 mg of mannitol were incubated at 37 degrees C for 2.5 hr with a simulated gastric fluid with and without pepsin. All polymers--although anionogenic--displayed quick swelling behavior in the acid milieu, leading to an unintended protein release. Moreover, pepsin is capable of penetrating into the polymeric carrier systems, thereby rapidly degrading the embedded protein. Enteric coating, on the other hand, leads to strongly reduced adhesive properties. Only NaC934P tablets coated with polymethacrylate containing 9% triethylcitrate displayed no significant (p < .05) reduction in adhesive strength. Results give essential basic information for the development of peroral (poly)peptide dosage forms based on mucoadhesive polymers. PMID- 10697748 TI - Peroral administration of enzymes: strategies to improve the galenic of dosage forms for trypsin and bromelain. AB - In this study, we investigated the presystemic metabolism of trypsin and bromelain and the influence of these proteolytic enzymes on the mucus layer covering the gastrointestinal (GI) epithelia. In vitro studies demonstrated that 77.3% +/- 4.0% (mean +/- SD, n = 3) of trypsin is autodegraded within 2 hr, whereas autodegradation of bromelain was negligible. In contrast to the metabolization of bromelain by all pancreatic serine proteases, trypsin is only degraded to some extent by elastase. Both therapeutically used enzymes remained stable after incubation with an excised porcine mucosa, demonstrating that proteolysis caused by brush border membrane-bound enzymes is negligible. Trypsin and bromelain were highly mucolytic active, thereby reducing the diffusion barrier based on the mucus gel layer. Strategies to improve the galenic of dosage forms for trypsin and bromelain include the use of bioadhesive polymers such as hydroxyethylcellulose or slightly modified chitosan-EDTA, providing strongly improved stability of these enzymes toward proteolytic degradation in vitro. The given information represents a good starting point to improve the galenic of dosage forms for orally administered proteolytic enzymes. PMID- 10697749 TI - Ruggedness study of HPLC peptide mapping for the identity of a drug compound: a chemometrics approach. AB - A statistically more reliable approach than the traditional visual inspection of peptide maps to identify a drug compound is to generate a set of reference standards from a designed experiment that incorporates many possible factors that affect variation of peptide mapping. In fact, the experiment can be done for a ruggedness study as part of a high-performance liquid chromatography (HPLC) method validation. Once the ruggedness is proved with the study, those articles in the experiment may form a set of reference standards, and future articles can be compared to the set later to prove identity. A quantitative analysis of the ruggedness study can be done using a chemometrics approach, principal component analysis (PCA). The analysis is used to reduce the many channels of peptide maps to a few manageable dimensions. The scores projected onto the reduced dimensions are used to test factor effects of the ruggedness study. As a by-product, the analysis provides visual inspection of the set of articles in the experiment for any outliers and anomalies. PMID- 10697750 TI - Chemometrics approach to the determination of polymorphism of a drug compound by infrared spectroscopy. AB - A chemometrics approach, multivariate calibration in particular, was used to determine the polymorphism of a drug compound based on Fourier transform infrared (FTIR) spectroscopy. The partial least-squares projection to latent structure makes use of all of the data, and the latent variables created by the method make use of hidden or partially separated peaks for quantitation. This paper illustrates the usefulness of the partial least-squares multivariate calibration method as an efficient tool to determine the polymorphism of a drug. Also, the analysis suggests the use of information from the modeling as diagnostic tools to gain more insight from the data. In particular, the diagnostic tools allow an analyst to assess design characteristics and any shortcomings of a calibration experiment for the polymorphism of a drug compound. PMID- 10697751 TI - Measurement of the adhesive force of fine particles on tablet surfaces and method of their removal. AB - The adhesion force of fine particles on the surface of tablets was measured by a centrifugal force and impact separation method. A Finededuster (FDD) was employed to remove fine particles from the tablet surface. The centrifugal force and impact separation method was suggested to be effective for measuring the adhesive forces between particles and the tablet surface, and effective disjoining force in the FDD could be estimated by comparison of the results obtained using these two methods. The FDD showed high removal efficiency regardless of how many tablets were processed at the same time. In either of these methods, critical particle size was about 10-20 microns, and larger particles were removed more efficiently. This critical particle size was similar to that observed for other mechanical properties of powders, such as angle of repose and flowability. We simulated particle residual percentage under various operation conditions by ANN (artificial neural network) analysis and multiple regression analysis. This simulation enabled us to predict how the efficiency of particle removal is affected by the interaction of the experimental and material factors. PMID- 10697752 TI - Influence of pharmacotechnical design on the interaction and availability of norfloxacin in directly compressed tablets with certain antacids. AB - Norfloxacin is a fluorquinolone that can interfere with antacids that contain aluminum and magnesium salts by complexation and modification of its solubility, which reduces its absorption and may lead to therapeutic failures. The purpose of this work was to evaluate the effect of the pharmaceutical design on this interaction and to develop a formulation of norfloxacin tablets in which this process could be avoided. Norfloxacin tablets were designed in 28 formulations. The interaction was studied in terms of in vitro dissolution behavior (USP 23, apparatus 2) in simulated gastric fluid with different doses of four commercially available antacid preparations. It was observed that dissolution rates were markedly reduced in the presence of all antacids studied. This phenomenon was practically avoided with some formulations of norfloxacin tablets in which a disintegrant (sodium starch glycolate or crospovidone) was included. These results indicated that the chelation among metal ions and norfloxacin could be affected by the delivering ability of the drug in the tablet. It was demonstrated that the pharmacotechnical design could modify an interaction process. Some formulations of tablets, in which the reduced dissolution rates in the presence of nonsystemic antacids in vitro was practically avoided, were developed by direct compression. PMID- 10697753 TI - Hot-melt coating technology. I. Influence of Compritol 888 Ato and granule size on theophylline release. AB - The aim of this work was to study the influence of theophylline granule size and the percentage of Compritol 888 Ato on in vitro drug release from granules and tablets. The granules were coated in a fluidized bed apparatus. The dissolution profiles of these granules differed from those of granules coated with classical agents, and there were also differences between the various sieve fractions studied. Drug release was characterized by a rapid-release phase, followed by a slow-release phase. Results indicate that theophylline release can be controlled by controlling granule size. Inspection of the appearance of the tablets at the end of the dissolution test revealed that all tablets containing Compritol 888 Ato remained intact. This indicated that the Compritol 888 Ato used in the tablet formulation created an inert matrix through which the drug diffused. It was found that the Higuchi relationship of linear square root of time was the best model to describe the release kinetics of the drug from tablets. This also confirmed that a matrix diffusion-controlled mechanism was operative. Given the difference between the dissolution profiles of the granules and the tablets, it was concluded that this matrix is formed during compression. PMID- 10697754 TI - Variation of composition of an enteric formulation based on Kollicoat MAE 30 D. AB - Using a formulation described previously with Kollicoat MAE 30 D as the film forming agent, the effect of variations in plasticizer type and quantity and talc concentration on the preparation and processing of spray-coating suspensions and the properties of isolated films and film-coated caffeine tablets prepared using them was investigated. In the preparation and processing of spray-coating suspensions, the plasticizers polyethylene glycol (PEG) 400, PEG1500, and TEC (triethyl citrate) tended to coagulate at all concentrations investigated, while Cremophor RH 40 coagulated above 10% (expressed as a percentage of the mass of the film-forming agent used). Analogous preparations using propylene glycol (PG), PEG6000, and Lutrol F 68, on the other hand, were found to be stable at all concentrations. The instability was not caused by the Kollicoat MAE 30 D polymer dispersion as such, but by interactions between the finely dispersed pigments and other formulation ingredients. Equivalent nonpigmented preparations are stable and do not coagulate. With all the plasticizers investigated, the minimum film forming temperature (MFT) fell, albeit to differing degrees, as the amount of plasticizer increased. Similarly, the tensile strength of isolated films declined as plasticizer concentration increased, while the reverse was true as regards their elongation at break. Whereas neither the subsequent disintegration time nor the rate of release of active ingredient at pH 6.8 was significantly affected by the various plasticizer additives, the different film-coated tablet formulations with a core containing a powerful disintegrant exhibited varying degrees of permeability to simulated gastric fluid. With PEG6000, permeability increased as the plasticizer concentration increased, while Lutrol F 68 provided an optimum barrier at 20%, and PG provided a good barrier between 10% and 30%. No gastroresistance was obtained with TEC at 10%. Only the best plasticizer formulations were used in the trials with different talc concentrations, namely, those formulations with 20% PEG6000, 20% Lutrol F 68, 20% PG, and 10% PG. When talc was added, the MFT rose, reaching its maximum at 13% talc (as a proportion of the film-forming agent). In the test for gastroresistance, film-coated caffeine tablets without talc absorbed distinctly more acid than those containing talc. Above 27% talc, the acid resistance improved only insignificantly. On the other hand, during this test, only a maximum of 3% of the active ingredient was released into the gastric juice. Of the variants investigated, the formulation with 20% PG and 27% talc performed best. PMID- 10697755 TI - A preliminary study on intravenous infusion of sodium eicosapentaenoate. AB - Eicosapentaenoic acid (EPA) and arachidonic acid (AA), made into sodium salt solution (50 micrograms/ml), were used for intravenous infusion. In a preclinical study in dogs, Na-EPA lowered the activity of transminases (glutamic pyruvic transaminase [GPT], glytamic oxaloacetic transaminase [GOT]); however, Na-AA increased the activity of GPT and GOT. In the clinical study, the numbers of leukocytes and lymphocytes of volunteers increased and remained at that level for 3 to 5 days after intravenous infusion. The study indicated that an intravenous infusion of Na-EPA may have anti-inflammatory and immunomodulatory effects. PMID- 10697756 TI - Effect of application volume of ethanol-isopropyl myristate mixed solvent system on permeation of zidovudine and probenecid through rat skin. AB - Permeation of zidovudine (AZT) and probenecid from an ethanol-isopropyl myristate (IPM) mixed system through rat skin was studied in a finite system. Several volume sizes of the ethanol-IPM mixed systems containing AZT and probenecid, both as suspensions, were applied on the skin of the hairless rat using a vertical glass cell, and the fractions of the drugs permeated in 8 hr Q%,8 hr were determined. For the systems containing 40% ethanol, the Q%,8 hr value decreased with the reduction of volume of the system applied, and the decreasing profile was similar to that calculated on the assumption that the permeability of the drug does not change with the volume of the sample applied. On the other hand, in the systems containing 10% or 20% ethanol, the Q%,8 hr value showed a maximum when a specific volume of the sample was applied. Therefore, the effect of sample volume on the Q%,8 hr value was different between the 40% ethanol-IPM system and the 10% or 20% ethanol-IPM system. Following pretreatment of the skin with 0.105 ml/cm2 of drug-free 40% ethanol-IPM for 2 hr, several volume sizes of 10% ethanol IPM systems containing the drugs were applied on the skin to explain why the different profiles were observed in the system containing 10% or 20% ethanol. The results for pretreated skin suggest that the amount of ethanol in the systems with low ethanol concentration and small application volume is too small to exert an effect that enhances permeation of the drugs. In those systems, the integrated effect of ethanol on the skin would be important for the enhancing effect. Total volume, as well as concentration, of an enhancer should be set precisely in designing an efficient transdermal delivery system. PMID- 10697757 TI - Comparison of the effect of lipid A analog E5531 and the lipid A from Escherichia coli on phospholipid membrane properties. AB - The effect of the lipid A analog E5531 on the phospholipid membrane was compared with that of the lipid A from Escherichia coli (EC). E5531 decreased the phase transition temperature of dipalmitoylphosphatidylcholine (DPPC) membrane and increased the fluidity and micropolarity. On the other hand, the effect of EC on the membrane was contradictory. These results suggested that the reason for the difference of biological effects of these two lipid A would be caused by the differences from the effect on the cell membranes. PMID- 10697758 TI - Influence of surfactants in aqueous-based polymeric dispersions on the thermomechanical and adhesive properties of acrylic films. AB - Good adhesion between a polymeric film and the surface of a solid substrate is critical to the performance of coated pharmaceutical products. Previous research has shown that tablet wettability by an organic-based cellulosic solution could predict the extent of film-tablet adhesion. Using an aqueous-based acrylic polymeric dispersion, the current study investigated the relationship between film adhesion and tablet wettability. Up to 10% (w/w based on dry polymer weight) polysorbate 80 or sorbitan monooleate was incorporated into the film-coating formulations. While the contact angle between the polymeric dispersion and the tablet surface was dependent on the type and concentration of surfactants added to the coating formulation, no correlation between tablet wettability and polymer adhesion could be established. The addition of surfactants to formulations containing the hydrophobic plasticizer tributyl citrate (TBC) caused lowering of the glass transition temperature of the polymer. Increased force of adhesion, elongation at adhesive failure, and adhesive toughness, however, were noted only in the TBC-plasticized films containing polysorbate 80. These findings demonstrate that our understanding of the mechanisms involved in film-tablet adhesion is still quite limited. PMID- 10697759 TI - Formulation optimization technique based on artificial neural network in salbutamol sulfate osmotic pump tablets. AB - The aim of this study was to develop a formulation optimization technique in which an artificial neural network (ANN) was incorporated; 30 kinds of salbutamol sulfate osmotic pump tablets were prepared, and their dissolution tests were performed. The amounts of hydroxypropyl methylcellulose (HPMC), polyethylene glycol 1500 (PEG1500) in the coating solution, and the coat weight were selected as the causal factors. Both the average drug release rate v for the first 8 hr and the correlation coefficient r of the accumulative amount of drug released and time were obtained as release parameters to characterize the release profiles. A set of release parameters and causal factors was used as training data for the ANN, and another set of data was used as test data. Both sets of data were fed into a computer to train the ANN. The training process of the ANN was completed until a satisfactory value of error function E for the test data was obtained. The optimal formulation produced by the technique gave the satisfactory release profile since the observed results coincided well with the predicted results. These findings demonstrate that an ANN is quite useful in the optimization of pharmaceutical formulations. PMID- 10697760 TI - Nonionic surfactant vesicles (niosomes) of cytarabine hydrochloride for effective treatment of leukemias: encapsulation, storage, and in vitro release. AB - Niosome vesicles of cytarabine hydrochloride were prepared by a lipid hydration method that excluded dicetylphosphate. The sizes of the vesicles obtained ranged from 600 to 1000 nm, with the objective of producing more blood levels in vivo. The study of the release of drug from niosomes exhibited a prolonged release profile as studied over a period of 16 hr. The drug entrapment efficiency was about 80% with Tween 80, Span 60 and Tween 20; for Span 80, it was 67.5%. The physical stability profile of vesicular suspension was good as studied over a period of 4 weeks. PMID- 10697761 TI - Consolidation behavior of an experimental, cross-linked polyalkyl ammonium polymer. AB - The effect of various factors (i.e., particle size, lubricant, moisture, and excipients) on the tableting properties of DMP 504 powder, an experimental, cross linked polyalkyl ammonium polymer, was studied using an instrumented single-punch tablet press. The results indicate that plastic deformation is the primary consolidation mechanism for DMP 504. Lubrication of DMP 504 with magnesium stearate resulted in negative interaction in compactibility. The increase in tablet hardness with increase in water content of DMP 504 (up to 2.5%) could be attributed to the lubricating effect of water. Increasing the water content above the optimum moisture range (i.e., 2.5% to 4.0%) caused a drastic reduction in tablet crushing strength due to the hydrodynamic resistance. A mixture of DMP 504 with microcrystalline cellulose or starch led to a positive interaction with respect to compactibility. A deviation in tablet strength from the linear interpolated value did not correspond to a deviation in tablet thickness. The improved compactibility for the mixture of DMP 504 and microcrystalline cellulose or starch is not related directly to the facilitated densification. PMID- 10697762 TI - New Ultraviolet spectrophotometric method for the estimation of nimesulide. AB - Two simple and accurate ultraviolet (UV) spectrophotometric methods with better detection range for estimation of nimesulide in pure form and in solid dosage form were developed in the present studies using 50% v/v and 100% v/v acetonitrile as the solvent system. The linearity range of nimesulide in both the methods was found to be 10-50 micrograms/ml at a lambda max of 300 nm. The linear regression equations obtained by the least-square regression method are Abs = 1.33 x 10(-1).Conc + 1.89 x 10(-1) in 50% v/v acetonitrile and Abs = 1.05 x 10( 1).Conc + 1.14 x 10(-1) in 100% v/v acetonitrile. The detection limit as per the error propagation theory was found to be 0.46 microgram/ml and 1.04 micrograms/ml, respectively, in 50% v/v and 100% v/v acetonitrile. The developed methods were employed with high degree of precision and accuracy for the estimation of total drug content in three commercial tablet formulations of nimesulide. The results of the analysis were validated statistically and by recovery studies. PMID- 10697763 TI - Are family physicians keeping up with the rapid changes in medicine? PMID- 10697764 TI - Understanding anger in residents. PMID- 10697765 TI - Including the patient in student presentations. PMID- 10697766 TI - Physician expression of empathy and positiveness to Hispanic and non-Hispanic white patients during medical encounters. AB - BACKGROUND: This study examined the extent to which physicians expressed empathy and positiveness to Hispanic and non-Hispanic white patients during primary care visits. METHODS: Twenty-seven family practice and internal medicine resident physicians at the University of New Mexico Health Sciences Center were audiotaped in 1995 with 427 adult patients who were fluent in English or Spanish. The tapes were reviewed and organized to measure how frequently physicians expressed empathy and positiveness to patients. RESULTS: Physicians expressed empathy at equal rates to Hispanic and non-Hispanic white patients. When examining only Hispanic patients, physicians were significantly more likely to express empathy to patients who they knew better. Physicians expressed positiveness to non Hispanic white patients more often than to Hispanic patients. When examining only Hispanic patients, physicians were more likely to express positiveness to patients who they knew better, who rated their health better, and who were more educated. When examining only non-Hispanic white patients, physicians were more likely to express positiveness to older and male patients than to younger and female patients. Also, female and younger physicians were significantly more likely to express positiveness to non-Hispanic white patients than male and older physicians were. CONCLUSIONS: Our findings illustrate that the resident physicians expressed empathy equally well to Hispanic and non-Hispanic white patients but that resident physicians need further training on how to express positiveness to patients from different ethnic backgrounds, especially Hispanic patients. PMID- 10697767 TI - Surgical practice of primary care physicians in a rural state: implications for curriculum design. AB - BACKGROUND: We surveyed practicing primary care physicians to help determine surgical practice patterns of primary care physicians in a rural state. The information obtained can be used to make surgical curriculum decisions for generalist medical students and primary care residents. METHODS: We developed a questionnaire in which practicing primary care physicians were asked to rate, on a 5-point Likert scale, the importance of 145 areas of surgical knowledge and 48 areas of clinical skills to their practice. Responses were rank ordered by the mean ratings for each individual item. The questionnaire was sent to all 876 primary care physicians in the home state of the institution. RESULTS: The survey response rate was 61% (n = 534). The most highly ranked items and procedures included acute otitis media, sinusitis, gastroesophageal reflux disease, pharyngitis, urinary tract infection, performance of abdominal exam, history and physical, daily progress notes, ear canal cleaning, and ability to write admission orders. The lowest ranked items included transplantation, infertility, amputations, performance of tracheostomy, venous cutdown, and cricothyrotomy. CONCLUSIONS: Information regarding the surgical practice patterns of practicing primary care physicians can be used to develop a surgical curriculum for medical students and primary care residents. PMID- 10697768 TI - Student religiosity and attitudes toward religion in medicine at a private Catholic medical school. AB - BACKGROUND AND OBJECTIVES: This study examined attitudes of medical students at a private Catholic medical school toward religion in medical education and practice and the relationship of these attitudes to medical student religiosity. METHODS: Surveys were mailed to first- and second-year medical students at Saint Louis University. The survey concerned attitudes about the integration of religious issues into the medical school curriculum and clinical practice and the personal importance of religion in the student's life (i.e., religiosity). RESULTS: The response rate was 61% (188/308). Nearly half of the students supported the introduction of religious studies into the medical curriculum, primarily through electives and modelling during clinical clerkships. Students with a higher level of personal religiosity were more likely to advocate training and participation in religious inquiry and behavior in the medical clinic. CONCLUSIONS: A significant minority of medical students at this Catholic university supported attention to religious issues in the medical school curriculum. The percentage might be lower at medical schools with no religious affiliation. The data indicate that students' religiosity is associated with their support for religious inquiry with patients and for the inclusion of religious issues in the medical school curriculum. PMID- 10697769 TI - An analysis of trends, perceptions, and use patterns of electronic medical records among US family practice residency programs. AB - BACKGROUND AND OBJECTIVES: This study intended to quantify electronic medical record (EMR) use in family practice residencies, associate program characteristics with EMR use, and identify perceptions and issues about the use of EMRs. METHODS: A survey was mailed to all 454 US family practice residency programs, with a 72% response rate. The survey, which was pretested and revised, was designed to identify benefits, problems, perceptions, and trends regarding the use of EMRs. RESULTS: Fifty-five of 329 programs (17%) were using an EMR, while 10 (3%) had used an EMR but discontinued. Programs in the South reported the highest EMR use (21%, 21/99), and those in the North Central region reported the lowest use (11%, 11/102). EMR use was highest in university settings (19%, 15/81), programs offering fellowships (26%, 24/92), new programs (36%, 18/48), and programs that require research (22%, 20/91). Of the 329 programs that responded, 43% (143 programs) reported having information systems (IS) committees. Of the 55 programs currently using EMRs, 78% had at least one full time equivalent IS technician. Of programs that discontinued use, software inadequacy was the most frequently cited reason (40%, 4/10). Programs that had never used EMR systems (n = 264) were more likely than those that had used EMRs (n = 65) to favorably perceive EMRs with respect to 1) meeting program requirements (44% versus 34%), 2) documenting improved patient care (65% versus 43%), 3) providing a reliable research database (94% versus 55%), and 4) documenting resident experience (92% versus 53%). Of the 264 (80%) programs that had never used an EMR, 172 (65%) plan to implement one. CONCLUSIONS: EMR use is low among US family practice residency programs, but some success in implementation of EMRs has been achieved. Based on the responses to this survey, use will likely increase from 55 of 329 programs (17%) to 153 of 329 (47%) by 2000. PMID- 10697770 TI - Using handheld computers to document family practice resident procedure experience. AB - BACKGROUND AND OBJECTIVES: We examined the use of inexpensive handheld computers in documenting resident procedures. With a handheld computer, data is entered at the time of the procedure, eliminating the problem of double entry. METHODS: Connectivity and ease of use were important factors considered when choosing a handheld computer. All residents received a handheld computer for data entry. Residency staff downloaded the data to a desktop computer. At the same time, data useful to residents was placed on their devices. The process of generating individual and program reports required 2 hours of staff time each month. Survey data regarding use and acceptance by residents was collected. RESULTS: Eighty eight percent of residents collected data on their handheld computer. Those residents responding to a survey felt that the handheld computer was "very useful," and 73% reported daily use. Initial costs were $310 per resident. CONCLUSIONS: Handheld computers streamlined the collection of procedure data for family practice residents. Handheld computers assisted in producing timely and useful procedural reports for both residents and the residency program. Additional uses of handheld computers were beneficial to the program and the residents. PMID- 10697771 TI - German family physicians' attitudes toward care of involuntarily childless patients. AB - BACKGROUND AND OBJECTIVES: Many family physicians regard fertility counseling out of their scope of practice, although key elements in the care of involuntarily childless couples fall within the theoretical framework of family practice. This study analyzed the doctors' value system concerning the care of infertile patients and whether a personal interview leads to a greater sensitivity toward fertility issues. METHODS: We conducted 57 baseline and 51 follow-up interviews with family physicians in the area of Gottingen, Germany. We performed quantitative and qualitative analyses. RESULTS: During the baseline interview, all family physicians placed involuntary childlessness within the domain of fertility specialists or regarded it as patients' private matter. Fourteen family physicians (27%) considered fertility counseling more important at the follow-up interview than at the time of the baseline interview. Judgmental views of infertile couples could be detected in both interviews. More than one third of the family physicians assumed a connection between the patients' childlessness and their personal behavior or way of living. Although the majority (73%) of the family physicians regarded involuntary childlessness as a disease and considered assisted conception techniques as legitimate, a recommendation for fee reimbursement for fertility services was rejected by more than half of the physicians. CONCLUSIONS: Most German family physicians do not consider that care of involuntarily childless couples is within or appropriate to their scope of practice. PMID- 10697772 TI - Family practice training in Nepal. AB - BACKGROUND: This article describes the health care system in Nepal and the only existing family practice (general practice) training program in that country. The majority of doctors in Nepal still have no residency training, and a specialist focus pervades. The efforts of some leading educators in Nepal led to establishment of a family practice training program in 1982, and the program now enrolls 12 residents per year, half of whom are from India. Major obstacles in education, financing, and policy must be addressed before family practice can have a meaningful presence or effect on the health care in Nepal. PMID- 10697773 TI - Family medicine: return to counterculture? PMID- 10697774 TI - Orchestral musings. PMID- 10697775 TI - Report of the European Network of Forensic Science Institutes (ENSFI): formulation and testing of principles to evaluate STR multiplexes. AB - This paper describes a collaborative exercise organised under the auspices of the European Network of Forensic Science Institutes (ENFSI). The purpose of this EU (European Union) funded group is to carry out research to enable STR loci to be compared between European laboratories, ultimately leading to the formation of a pan-European database. Accordingly, an exercise was designed to evaluate a prototype STR multiplex system manufactured by Applied Biosystems (ABD). Each laboratory was sent 12 samples to analyse along with a multiplex kit. Of specific interest was the definition of parameters to define the efficiency of the system. Stutter, split allelic peaks (differing by one base), pull-up, heterozygous balance and between locus balance were all objectively measured. Once the important parameters are defined it is possible to directly compare performances of different multiplexes and the different laboratories carrying out the tests. Since the multiplex used was a prototype system, this exercise cannot be regarded as a proficiency test. PMID- 10697776 TI - Portuguese population and paternity investigation studies with a multiplex PCR- the AmpFlSTR Profiler Plus. AB - A Portuguese Caucasian population of 146 unrelated individuals was studied. DNA samples were amplified by multiplex PCR for D3S1358, vWA, FGA, D8S1179, D21S11, D18S51, D5S818, D13S317 and D7S820 using the AmpFlSTR Profiler Plus PCR Amplification Kit (Perkin-Elmer). All loci met Hardy-Weinberg expectations. Forensic statistical parameters were according to those obtained by other authors. Statistical differences were observed concerning three loci when comparing the Portuguese Caucasian population and an Italian Caucasian population, although these differences mainly concern the less frequent alleles. Eighty-three paternity investigation cases were analysed. Exclusions in between three and nine loci were observed in all the 23 exclusion cases obtained. Most of the non-exclusion cases had probability of paternity > 99.9%. Two cases with an isolated genetic incompatibility between the alleged father and the child were detected, which may indicate probable mutation cases. These results demonstrate that the AmpFlSTR Profiler Plus is a suitable multiplex for paternity investigation in the Portuguese population. PMID- 10697777 TI - The acetabulum: sex assessment of prehistoric New Zealand Polynesian innominates. AB - With increasing urban development in New Zealand, prehistoric Polynesian skeletal remains are frequently being recovered. Since such material must often be reinterred quickly, it has become important that the sex of individuals be determined from the remains in a relatively short time. For this purpose, discriminant function analysis was utilised for sex determination of prehistoric adult New Zealand Polynesian innominates (21 male and 35 female). Maximum diameter of the acetabulum was measured and subjected to SPSS direct discriminant function analysis. Accuracy of sex determination ranged from 85.2% to 86.2%. Reduction in error over random assignment by sex ranged from 70% to 72%. The two discriminant functions derived will provide a useful tool for the assessment of human remains in the forensic and archaeological context because they incorporate a single measurement which can be taken on incomplete bones. PMID- 10697778 TI - Noscapine as an adulterant in illicit heroin samples. AB - In this short report the evidence is given (based on the analyses of 22 case samples) that noscapine can be used as an adulterant in illicit heroin samples. In this context, the appearance of illicit heroin samples characterised by a high noscapine content (up to 61%) and a high noscapine/whole morphine ratio (up to 3.5) is highlighted. All samples discussed in the paper (132) were seized in Slovenia, in the period from 1997 to 1999 and were analysed by gas chromatography mass spectrometry. PMID- 10697779 TI - Blood strontium concentration related to the length of the agonal period in seawater drowning cases. AB - The levels of ventricular blood strontium (Sr) from 70 seawater drowning victims were compared with their diagnosis of drowning based mainly on certain criteria selected from their autopsy report. From this comparison, intervals of either the difference of Sr concentration between the left and the right ventricle blood (LVSr-RVSr) or the Sr concentration in the left ventricle blood (LVSr), appear to be related to different time-lapses of the agonal period of drowning. In the aim to diagnose drownings, intervals of both LVSr-RVSr and LVSr were proposed to characterize three different agonal periods in seawater drowning cases: instantaneous death (ID), fast vital-submersion drowning (FVSD) and common vital submersion drowning (CVSD). PMID- 10697780 TI - Suicide of a body packer. AB - The smuggling of illicit drugs by means of body packing has become a common problem at European airports. Europe is considered to be the fastest growing market for cocaine worldwide, and the air route is the most frequently used method of trafficking cocaine. Smuggling illicit drugs by use of body packing is considered to be a high toxicological hazard because of the risk of leakage or a package bursting. We report about the first case of suicide of a body packer by re-ingesting the content of excreted cocaine packages. The consequence of this case is that the death scene investigation and autopsy assessments in case of a body packer's death should always consider the possibility of re-ingested packages. Detention personnel should be instructed by forensic and criminalistic experts to take preventive measures. PMID- 10697781 TI - A frozen newborn infant: froth in the air-passage after thawing. AB - We performed an autopsy on a frozen newborn infant who was found in a freezer at 18 degrees C. After thawing, froth emerged from the nostrils and was present in the trachea. Sometimes froth may be seen in the air-passage in cases of strangulation and drowning. In our case, however, there was neither proof of asphyxia due to strangulation nor drowning. The existence of the froth indicates that the infant was probably in a state of respiratory distress before death. Histologic findings of the lung showed that the infant did not suffer from respiratory disorders such as respiratory distress syndrome. Karyopyknosis and vacuolation of the keratinocytes, shrinkage of the hepatocytes, dilatation of the sinusoid, spaces between heart muscle fibers and deep staining of the nuclei and hemolysis were characteristic in our case. This case shows that froth persists in the internal air-passage for a long time as a result of freezing. Moreover, the froth in the air-passage, along with the findings of the lungs, demonstrates that the newborn infant was born alive. PMID- 10697782 TI - Bacterial meningitis--problems on many fronts. PMID- 10697783 TI - The impact of HIV on meningitis as seen at a South African Academic Hospital (1994 to 1998). AB - BACKGROUND: The increase in HIV infections in South Africa is alarming. The aim of this prospective 4-year study was to evaluate the rising incidence of HIV related admissions due to meningitis at the Pretoria Academic Hospital (PAH) adult neurology ward and to investigate the spectrum of meningitis during this time. PATIENTS AND METHODS: Adults with meningitis presenting at the PAH neurology ward from March 1994 through February 1998 were included. HIV antibody status was determined and patients were assigned to five categories: bacterial, tuberculous, viral and cryptococcal meningitis, as well as an uncertain category. RESULTS: Over the 4-year study period 141 patients with meningitis were seen. Of these, 44 were HIV-positive (31%), with TB meningitis occurring in 16 (36%), cryptococcal meningitis in 22 (50%) and acute bacterial meningitis in three (7%). In the first 2 years of the study, 14% of patients were HIV positive; this figure rose to 44% in the 3rd year, and 57% in the final year. The spectrum of meningitis also changed: bacterial meningitis remained relatively stable at about 25% of the total; TB meningitis almost doubled from 16% in the 1st year to 31% in the last year of the study; viral meningitis initially occurred in 8% of patients and later in 3% of cases, while cryptococcal meningitis showed the most significant increase from 6% of cases in 1994/5 to 31 and 26% respectively in the last 2 years of the study. CONCLUSION: Over a 4-year period the HIV epidemic was responsible for a marked shift in the spectrum of meningitis towards chronic infections such as TB and cryptococcal meningitis at the PAH. PMID- 10697784 TI - Mixed infection in adult bacterial meningitis. AB - 12 adult patients suffering from bacterial meningitis caused by mixed infection were identified at Kaohsiung Chang Gung Memorial Hospital over a period of 13 years (1986-1998), and they accounted for 6.5% (12/184) of our culture-proven adult bacterial meningitis. The 12 cases included seven males and five females, aged 17-74 years. Six of the 12 cases had community-acquired infections and the other six had nosocomially-acquired infections. Ten of the 12 cases had associated underlying diseases, with head trauma and/or neurosurgical procedure being the most frequent. Both gram-negative and gram-positive pathogens were identified in these 12 cases with gram-negative pathogens outnumbering the gram positive ones. The implicated pathogens, starting with the most frequent, included Enterobacter species (Enterobacter cloacae, Enterobacter aerogenes), Klebsiella species (Klebsiella pneumoniae, Klebsiella oxytoca), Escherichia coli, Staphylococcus species (Staphylococcus aureus, Staphylococcus haemolyticus), Pseudomonas aeruginosa, Acinetobacter baumannii, Enterococcus, Serratia marcescens, Citrobacter diversus, Proteus mirabilis, Streptococcus viridans and Neisseria meningitidis. Six of the 12 cases were found to have multi-antibiotic resistant strains, which included E. cloacae in one, A. baumannii in one, K. pneumoniae in one and S. aureus in three. The management of these 12 cases included appropriate antibiotics and neurosurgical procedures including shunt revision. Despite the complexity of implicated pathogens and the high incidence of emergence of resistant strains, the overall mortality rate (8.3%, 1/12) was not higher than that in adult bacterial meningitis. However, complete recuperation was difficult in adult patients with mixed bacterial meningitis. PMID- 10697785 TI - Release of teichoic and lipoteichoic acids from 30 different strains of Streptococcus pneumoniae during exposure to ceftriaxone, meropenem, quinupristin/dalfopristin, rifampicin and trovafloxacin. AB - The release of teichoic acids (TA) and lipoteichoic acids (LTA) from 30 different strains of Streptococcus pneumoniae during exposure to ceftriaxone, meropenem, quinupristin/dalfopristin, rifampicin and trovafloxacin at concentrations of 10 micrograms/ml and of the respective MIC was determined by an enzyme immunoassay. At 10 micrograms/ml the most rapid and intense release was detected during treatment with the beta-lactam antibiotics ceftriaxone and meropenem, the lowest release was seen with rifampicin and quinupristin/dalfopristin. Trovafloxacin delayed the release of TA/LTA. The maximum concentrations of TA/LTA, however, during trovafloxacin treatment were almost as high as those during exposure to ceftriaxone and meropenem. During exposure to the MIC, ceftriaxone, meropenem, rifampicin and trovafloxacin released significantly higher amounts of TA/LTA than during exposure to 10 micrograms/ml (p < 0.01). Only quinupristin/dalfopristin released small amounts of TA/LTA at the low and high concentration. In conclusion, at high concentrations antibiotics that do not affect the bacterial cell wall released less pro-inflammatory compounds from S. pneumoniae than ceftriaxone and meropenem. This may be of value in the treatment of meningitis and sepsis. PMID- 10697786 TI - Mixed infection with two types of hepatitis C virus is probably a rare event. AB - A search for the simultaneous presence of two hepatitis C virus (HCV) types in sera of a group of chronically infected intravenous drug users, hemodialysis patients and hemophiliacs from Sweden and Russia was performed with two genotyping methods based on the use of type-specific primers from core and NS4 regions of the viral genome. An important feature of NS4 based assay is that type specific primers are used in both rounds of nested PCR, thus providing the possibility of the identification not only of the abundant type, but also of the minor HCV type present in a particular serum. The experiments, however, did not reveal the simultaneous presence of two or more HCV types in any of the 40 samples. These results suggest that the frequency of mixed infections in serum with different HCV types is very low even in high-risk groups, at least in the geographic region studied. PMID- 10697787 TI - Epidemiology of candidemia--a nationwide survey in Israel. AB - Bloodstream infections with Candida are often lethal and have been reported to be increasing in frequency. The current retrospective study describes the magnitude and epidemiological characteristics of candidemia in all western-type hospital facilities in Israel in 1994. Comprehensiveness of the data from the reporting hospitals was checked by cross-study of the data from the infectious diseases records and from the hospitalization records. Vital status of all reported cases was evaluated 1 year after the diagnosis. Data on 298 newly diagnosed cases of candidemia were received from 14 of the 18 general hospitals in Israel. The proportion of candidemia in the Israeli hospitals ranged from 0.1 to 0.01% of all admissions, with a mean of 0.05%. The incidence of candidemia differed significantly between the wards from 4-5/10,000 in general surgery and internal medicine wards to about 60/10,000 and 80/10,000 in intensive care and preterm units, respectively. Of all detected cases 53.6% were Candida albicans. Another nine specific species of Candida (mainly Candida parapsilosis, Candida tropicalis and Candida glabrata) were detected, with major differences between the various hospitals. The species of Candida differed significantly by sex and age. Of the cases of candidemia 21.5% died within 30 days of the isolation of the pathogen. The one-year mortality rate was 31.9%. Species-specific 30-day mortality rate was highest for C. glabrata. Throughout the analysis, C. glabrata emerged as a unique cause of candidemia, producing higher mortality, appearing at a younger age and predominating among females. PMID- 10697788 TI - GB virus C/hepatitis G virus infections in traumatologic outpatients, chronic non A-E hepatitis and extrahepatic malignancies. AB - GB virus C/hepatitis G virus (GBV-C/HGV) is a recently discovered flavivirus of still unknown pathogenic relevance. We examined traumatologic outpatients to determine GBV-C/HGV viremia for further epidemiological studies, as blood donors hitherto used as controls represent healthy individuals without risk factors. Anti-GBV-C/HGV antibodies were detectable in 13.2% (95% confidence interval [CI] 9.3-18.2) and GBV-C/HGV RNA was detectable in 4.5% (95% CI 2.4-8.2) of the outpatients. In chronic non-A-E hepatitis patients GBV-C/HGV viremia was detectable at a significantly higher level of 16.1% (95% CI 6.1-34.5), while the prevalence of anti-GBV-C/HGV antibodies was 12.9% (95% CI 4.2-30.8). The rate of GBV-C/HGV viremia in patients with malignant diseases (different types of tumors, blood recipients were excluded) was 12.5% (95% CI 8.4-18.1), a significant elevation compared to traumatologic outpatients. The seroprevalence in the tumor group was 22.1% (95% CI 16.7-28.6), also significantly elevated. Thus, there are two messages. Firstly, testing for GBV-C/HGV may be a useful extension of the diagnostic procedure of viral hepatitis. Secondly, common risk factors or etiologic relations of GBV-C/HGV and extrahepatic malignancies should be discussed. PMID- 10697789 TI - Significance of respiratory syncytial virus (RSV) infection in the 1st year of life. AB - BACKGROUND: In this study we investigated the frequency, symptoms and predisposing factors of respiratory syncytial virus (RSV) infection during the 1st year of life in infants with obstructive airway disease in comparison with infants without airway disease. PATIENTS: We enrolled 216 infants in their 1st year of life, who were hospitalized because of obstructive airway disease. As an age- and sex-balanced control group, we examined 133 infants hospitalized for other reasons than airway disease. METHOD: A deep pharyngeal swab was taken from all infants and immediately examined for the presence of RSV antigen by using an enzyme immunoassay (Directigen). Patient data were surveyed by a questionnaire. RESULTS: The frequency of RSV infections among infants with obstructive airway disease (34.3%; n = 74) differed significantly from the control group (15%; n = 20; p < 0.01). The frequency of RSV-infected infants with obstructive airway disease decreased with age ranging from 39.1% in trimenon I to 29.0% in trimenon IV. This trend was not observed in the control group. With respect to clinical symptoms and risk factors, there were no differences between RSV-infected versus noninfected infants. CONCLUSION: RSV is an important agent causing lower obstructive airway disease (34.3% of all patients). There are no specific symptoms that can be used for diagnosing RSV infection. In order to prevent other patients on the ward from contracting nosocomial RSV infection and in the light of therapeutic options, one should test newly admitted patients presenting with symptoms of an obstructive airway disease for RSV antigen. On a ward with high risk patients, we would recommend the use of an RSV test for all new patients. PMID- 10697790 TI - Seroprevalence of Chlamydia pneumoniae antibodies in a young adult population sample living in Verona. European Community Respiratory Health Survey (ECRHS) Verona. AB - The aim of the study was to evaluate the prevalence of antibodies to Chlamydia pneumoniae in a random population sample of 369 young adults (aged 20-44 years), living in Verona, Italy. IgG and IgM titers were measured by micro immunofluorescence. IgG antibodies, greater or equal to 16, were found in 104/177 (58.8%) men and 76/192 (39.6%) women (p < 0.001). No relationship was found between IgG seropositivity, age, social class, education and family size. Factors positively associated with IgG seropositivity included smoking (p < 0.001), occupational status (employed vs unemployed: p = 0.02; students vs unemployed: p < 0.01) and living area (suburban [65.0%] vs urban area [45.3%]: p = 0.03). The geometric mean of IgG titers was higher in students (GM: 26.05) than in both employed (GM: 11.02) and unemployed persons (GM: 4.80) (p < 0.01 and p < 0.001, respectively). IgG titres > or = 512 and/or IgM titers > or = 16 (suggestive of a recent C. pneumoniae infection) were found in 39 subjects (10.6%). Recent infection was more frequent in spring (14.9%), with no significant variation in the other seasons (mean prevalence 6.7%) (p < 0.01). Recent infection was also associated with cigarette smoking. On the other hand, no significant association was found between respiratory symptoms and serologic evidence of recent infection. IN CONCLUSION: 1) the prevalence of antibodies to C. pneumoniae in young adults from Verona is similar to that found in European countries, and therefore, in Europe, it seems not related to latitude or climate; 2) male sex, tobacco smoking, employment status and living in a suburban area are independent risk factors of infection; 3) the infection is subclinical in most cases. PMID- 10697791 TI - Specific in vitro proliferative immune responses and lymphokine production in Ethiopian children with and without tuberculosis. AB - We investigated the profile of some in vitro parameters of cellular immune responses in non-HIV-infected Ethiopian children and young adults with and without tuberculosis (TB) as compared to healthy Ethiopian and non-Ethiopian controls. The in vitro proliferative responses of peripheral blood mononuclear cells (PBMC) to purified protein derivative (PPD) were determined in 15 Ethiopian children and young adults with TB, 12 healthy Ethiopian children who were contacts of TB patients, 20 Ethiopian children without contact with TB and ten non-Ethiopian controls. All TB patients and contacts had a positive Mantoux skin test. The PBMC proliferative response to PPD of the Ethiopian children with TB was significantly higher than that of the Ethiopian children without TB, while all Ethiopian children demonstrated stronger proliferative response as compared to non-Ethiopian healthy controls. Interleukin 2 (IL-2), interferon gamma (IFN gamma), interleukin 4 (IL-4) and interleukin 6 (IL-6) were measured by ELISA assays performed on the supernatant of PPD-stimulated and non-stimulated PBMC cultures of seven Ethiopian children with TB, ten Ethiopian children without TB and eight non-Ethiopian controls. IFN-gamma and IL-4 were undetectable and IL-2 levels in unstimulated supernatants were low in all groups. PPD stimulation induced a significant rise in IL-2 levels in Ethiopians with TB as compared to all other groups. There was no increase above baseline in IL-6 levels in any group studied. CONCLUSIONS: Ethiopian children with TB exhibit a strong cellular immune response as expressed by Mantoux tests and lack of stimulation of IL-4 and IL-6 production. This pattern suggests a Th1 type effective cellular immune response to mycobacteria in a cohort of young Ethiopians with TB. PMID- 10697792 TI - Retroperitoneal abscess and bacteremia due to Mycoplasma hominis in a polytraumatized man. AB - We report a case of a retroperitoneal abscess due to Mycoplasma hominis in a young polytraumatized man who developed septicemia under treatment with rifampin and flucloxacillin. M. hominis was recovered from blood cultures as well as from the abscess near the left iliac spine. After 10 days of therapy with clindamycin the patient improved, and intraoperatively taken swabs were culture negative but still positive by PCR. PMID- 10697793 TI - Pyomyositis of the thigh due to Prevotella melaninogenica. AB - Pyomyositis is an uncommon infection in temperate climates, however, it is being more frequently reported among patients with diabetes or malignancy, or those who are immunocompromised. It is predominantly caused by Staphylococcus aureus, and rarely by Bacteroides species. Pyomyositis due to Prevotella melaninogenica has not previously been reported. We describe an elderly patient with pyomyositis of the thigh due to P. melaninogenica which was successfully treated by surgical incision and drainage in combination with metronidazole therapy. PMID- 10697794 TI - Double prosthetic valve endocarditis caused by Streptococcus pneumoniae. AB - Infective endocarditis (IE) caused by Streptococcus pneumoniae is a rare disease. Only eight cases of pneumococcal prosthetic valve endocarditis have been described in the literature. In this report we describe the first case of pneumococcal endocarditis involving two prosthetic heart valves. The patient had pneumonia as the probable portal of entry but no predisposing conditions for invasive pneumococcal disease. Our case also illustrates the importance of transesophageal echocardiography (TEE) for the early diagnosis of IE and a timely decision for cardiac surgery. PMID- 10697795 TI - Persistent wound infection after herniotomy associated with small-colony variants of Staphylococcus aureus. AB - A small-colony variant (SCV) of Staphylococcus aureus was cultured from a patient with a persistent wound infection (abscess and fistula) 13 months after herniotomy. The strain was nonhemolytic, nonpigmented and grew only anaerobically on Schaedler agar. As it was coagulase-negative, it was initially misidentified as a coagulase-negative Staphylococcus. In further analysis, however, the microorganism was shown to be an auxotroph that reverted to normal growth and morphology in the presence of menadione and hemin (Schaedler agar) and could be identified as a SCV of Staphylococcus aureus. Surgery and antibiotic treatment of the patient with flucloxacillin and rifampicin for 4 weeks resulted in healing of the chronic wound infection. PMID- 10697796 TI - Pulmonary tuberculosis presenting with cutaneous leukocytoclastic vasculitis. AB - This case report deals with a rare association: tuberculosis and cutaneous leukocytoclastic vasculitis. The patient was a 36-year-old man with no significant past medical problems. He presented with a palpable purpura on both legs, low-grade fever, cough and expectoration, progressive dyspnea due to a massive left pleural effusion and a symmetric swelling on his ankles and wrists. Skin biopsy yielded a histological diagnosis of leukocytoclastic vasculitis and the primary diagnosis was only achieved after performing a pleural biopsy, which unequivocally showed the presence of Mycobacterium tuberculosis. This case shares many features with the few cases already reported in the medical literature. Possible pathogenic mechanisms are reviewed and discussed in detail. PMID- 10697797 TI - Decreased susceptibility to extended-spectrum cephalosporins of a penicillin susceptible Streptococcus pneumoniae in meningitis. AB - A 69-year-old woman was admitted to the hospital with meningitis due to Streptococcus pneumoniae. The strain was susceptible to penicillin but intermediate to cefotaxime. In Europe the decrease of susceptibility generally pertains more to penicillin than to cefotaxime. Such a strain is perhaps a forewarning of the existence of high-level cephalosporin-resistant strains. Despite the possible detection of the resistance by oxacillin disk, it underlines the need to determine the MICs of different beta-lactams without delay and to choose the most efficient treatment. PMID- 10697798 TI - Linezolid, critical characteristics AB - In spite of a constantly expanding information base with the oxazolidinones generally and linezolid specifically, we have elected here to focus on the key characteristics of linezolid. Linezolid is the first member of a new class of antimicrobial agents, the oxazolidinones, to be tested in humans in Phase I, Phase II and Phase III clinical trials. The oxazolidinones have a novel mechanism of action in that they inhibit initiation complex formation in bacterial protein synthesis and, consistent with a novel mechanism of action, they do not exhibit cross-resistance with existing antibacterial agents. Importantly, resistance development as measured in the laboratory occurs very slowly, there is no evidence of rapid resistance development. The spectrum of oxazolidinone activity is principally gram-positive and in vitro studies demonstrate that linezolid is equivalent to vancomycin in vitro. Linezolid is orally as well as intravenously active and orally administered linezolid is as efficacious in mouse models of bacterial disease as is subcutanously administered vancomycin against appropriate pathogens. The exceptional oral behavior of linezolid in mouse models is readily explained by the observation that oral linezolid is 100% bioavilable and that administration of 400- and 600-mg doses of linezolid in humans results in blood level curves which predict that linezolid will be very well suited for bid dosing. Additionally, the blood level concentrations are in significant and very comfortable excess of the MIC90 concentrations for the important gram-positive pathogens for the bulk of the dosing interval. PMID- 10697799 TI - 192nd and 193rd meetings of the Dutch Ophthalmological Society. March 1998 and March 1999. Abstracts. PMID- 10697800 TI - Cysteine protease inhibitory activity and levels of salivary cystatins in whole saliva of periodontally diseased patients. AB - The 3 human salivary cystatins S, SA and SN are multifunctional proteins that possess a cysteine protease inhibitory property, but their ability to act as such is very different (SN > SA >> S). One form, S, also appears to possess antibacterial properties towards the bacterium Porphyromonas gingivalis, often associated with periodontal diseases. In this study we measured the total cystatin inhibitory activity and the levels of each salivary cystatin in the whole saliva of 8 periodontally diseased patients and 2 groups of control subjects (n = 6 and n = 10). The total cystatin inhibitory activity and the total salivary cystatin concentration in the periodontally diseased patients were found to be lower than the controls (p < or = 0.005). The concentration of S was depleted to levels that would not allow it to be an effective antibacterial agent, and the concentration of SA, although depleted in some cases, was still present at sufficient levels to allow it to act as an effective physiological inhibitor of cathepsin L. The concentration of cystatin SN was also depleted in the periodontally diseased patients, but was still present in sufficient quantities to act as an effective physiological cysteine protease inhibitor of cathepsins H and L. In comparison, the concentration of all 3 salivary cystatins in the control subjects were sufficient to enable these proteins to be both effective physiological cysteine protease inhibitors and antibacterial agents. PMID- 10697801 TI - Comparison of gingival and peripheral blood T cells among patients with periodontitis suggests skewing of the gingival T cell antigen receptor V beta repertoire. AB - The present study investigated the expression of different variable regions of T cell receptor beta-chain (V beta) among functional subsets of T cells, i.e. CD45RO+ (activated/memory), CD4+ and CD8+ in gingiva and peripheral blood of patients with periodontitis. Gingival tissue specimens (n = 25) and peripheral blood were procured from 18 patients with periodontitis during periodontal surgery or extraction. Single-cell suspensions of gingival tissues were made by enzymatic digestion. These cells were immunofluorescently labeled with a panel of monoclonal antibodies specific for 18 TCR V beta regions, in concert with markers for various T cell subsets. The cells were then analyzed with 3-color multivariate flow cytometry. Results demonstrated that a significantly higher proportion of T cells in gingiva expressed V beta 5.2 (0.0005), V beta 6 (0.0007) and V beta 9 (0.003) regions compared to those in peripheral blood. Comparison of CD45RO+ (activated/memory) and CD45RO- (naive) subsets of gingival T cells revealed differences in the expression of TCR V beta regions. V beta 5.2 expression was significantly higher among CD45RO+ gingival T cells (p = 0.004), whereas V beta 14 expression was elevated among the CD45RO- subset relative to peripheral blood (p = 0.008). Analysis of TCR V beta region expression among CD4+ and CD8+ subsets did not reveal any statistically significant differences between gingiva and peripheral blood, although some V beta regions approached significance. Collectively, these results demonstrate that the T cell repertoire in the gingival compartment differs significantly from that in the peripheral blood. Furthermore, since the skewing of TCR V beta was observed among naive, as well as activated/memory T cells, it is likely that both developmental and environmental factors are influential in shaping the gingival TCR repertoire in patients with periodontitis. Elucidation of the cause of the skewed expression of T cell receptors in gingiva can provide insights into the specificity of T cells in periodontitis. PMID- 10697802 TI - Alteration in expression of MMP-1 and MMP-2 but not TIMP-1 and TIMP-2 in hereditary gingival fibromatosis is mediated by TGF-beta 1 autocrine stimulation. AB - Hereditary gingival fibromatosis (HGF) is characterized by an excess accumulation of extracellular matrix (ECM) resulting in a generalized and fibrotic enlargement of the gingiva. To investigate some of the regulatory features of this condition, gingival fibroblasts from normal gingiva (NG) and HGF were examined for the expression and production of matrix metalloproteinases (MMPs) and their inhibitors, tissue matrix metalloproteinases inhibitor (TIMPs). Our results, obtained from 2 different assays, semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR) and enzymography, clearly demonstrated that the expression and production of MMP-1 and MMP-2 was significantly lower in fibroblasts from HGF than from NG. Interestingly, TIMP-1 and TIMP-2 expression from NG cells was shown to be slightly higher to those from HGF. Addition of antibodies against transforming growth factor-beta 1 (TGF-beta 1), which is produced in greater amounts by HGF fibroblasts, resulted in a slight increase in MMP-1 and a decrease in MMP-2 expression, whereas TIMP-1 and TIMP-2 expressions were unaffected. These patterns of expression and production suggest that enhanced TGF-beta 1 production reduce the proteolytic activities of HGF fibroblasts, which favor the accumulation of ECM. PMID- 10697803 TI - Molecular genetics and nomenclature of proteases of Porphyromonas gingivalis. AB - The strategies used by bacterial pathogens to establish and maintain themselves in the host represent one of the fundamental aspects of microbial pathogenesis. Characterization of these strategies and the underlying molecular machinery offers new opportunities both to our understanding of how organisms cause disease in susceptible individuals and to the development of novel therapeutic measures designed to undermine or interfere with these determinants of successful survival. With respect to the microbial aetiology of the periodontal diseases, a growing body of evidence suggests that the proteolytic enzymes of Porphyromonas gingivalis represent key survival and, by extrapolation, virulence determinants of this periodontal bacterium. This in turn has led to international efforts to characterize these enzymes at the gene and protein level. Approximately 20 protease genes of P. gingivalis with different names and accession numbers have been deposited in the gene databases and a correspondingly heterogeneous nomenclature system is employed for the products of these genes in the literature. However, it is evident, through comparison of these gene sequences and through gene inactivation studies, that the genetic structure of the proteases of this organism, particularly those with specificity for arginyl and lysyl peptide bonds, is less complicated than originally thought. The major extracellular and surface associated arginine specific protease activity is encoded by 2 genes which we recommend be designated rgpA and rgpB (arg-gingipains A & B). Similarly we recommend that the gene encoding the major lysine specific protease activity is designated kgp (lys-gingipain). These three genes, which account for all the extracellular/surface arginine and lysine protease activity in P. gingivalis, belong to a family of sequence-related proteases and haemagglutinins. PMID- 10697804 TI - The in vitro effects of cetyltrimethylammonium naproxenate on oral and pharyngeal microorganisms of various ecological niches. AB - The purpose of this study was to determine the in vitro susceptibility to cetyltrimethylammonium naproxenate for various aerobic and anaerobic micro organisms responsible for oral and pharyngeal diseases by assessing the minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs) or minimum fungicidal concentrations (MFCs) and by determining kill-times. The MICs of cetyltrimethylammonium naproxenate for 46 tested strains (25 reference strains and 21 clinical isolates) ranged from 8 to 500 micrograms/ml. The MIC was found to be 31.25 micrograms/ml for 36% of the reference strains. Even lower MIC values (15.63 micrograms/ml) were observed for some anaerobic strains, for Haemophilus influenzae and for Candida tropicalis. MIC and MBC values corresponded for the majority of strains tested while the MFC for C. tropicalis and C. albicans was much higher. Only 9.5% of the clinical isolates gave a MIC value of 31.25 micrograms/ml. Enterococcus faecalis, Streptococcus pyogenes and Staphylococcus aureus showed MIC at 62.5 micrograms/ml. The MIC and MBC values among the isolates were comparable, while the MFC value for the yeasts was greater. A concentration of 125 micrograms/ml of cetyltrimethylammonium naproxenate inhibited the growth of all bacteria, except Enterobacteriaceae and Pseudomonaceae, and yeasts. Cetyltrimethylammonium naproxenate shows very rapid kill-time for S. sanguis (0"), and rapid (15") for S. pyogenes, S. dysgalactiae and S. mutans and for Moraxella catarrhalis, while a longer kill-time was necessary for the other microbes tested. PMID- 10697805 TI - Prostaglandin E2 receptors of the EP2 and EP4 subtypes downregulate tumor necrosis factor alpha-induced intercellular adhesion molecule-1 expression in human gingival fibroblasts. AB - Prostaglandin E2 (PGE2) exerts its biological actions via EP receptors, which are divided into 4 subtypes of EP1, EP2, EP3 and EP4. In the present study, we investigated whether PGE2 regulated intercellular adhesion molecule-1 (ICAM-1) expression in human gingival fibroblasts (HGF) stimulated with tumor necrosis factor-alpha (TNF alpha) and if so, which subtype(s) of PGE2 receptors was involved. Exogenous addition of PGE2 to HGF inhibited ICAM-1 expression elicited by TNF alpha in a concentration-dependent manner. Treatment of HGF with indomethacin, a cyclo-oxygenase inhibitor, had no effect on TNF alpha-elicited ICAM-1 expression, although indomethacin completely inhibited PGE2 production enhanced by TNF alpha. Next, we examined which subtype(s) of the 4 EP receptors modulated the ICAM-1 expression elicited by TNF alpha, using subtype-specific agonists and antagonists. 11-deoxy-PGE1, a selective EP2/EP4 agonist, inhibited TNF alpha-elicited ICAM-1 expression as potently as PGE2, while butaprost, a selective EP2 agonist, was somewhat less effective than PGE2. AH23848B, an EP4 antagonist, antagonized the inhibitory effect of TNF alpha-elicited ICAM-1 expression by PGE2. Sulprostone, an EP1/EP3 agonist, and ONO-AP-324, an EP3 agonist, were inert to TNF alpha-elicited ICAM-1 expression. As EP2 and EP4 receptors are linked to elevation of intracellular cyclic AMP (cAMP), the effect of dibutyryl cAMP and 8-bromo-cAMP, cAMP analogs, on TNF alpha-elicited ICAM-1 expression was examined. Both the agents downregulated ICAM-1 expression in TNF alpha-stimulated HGF. From these data, we suggest that PGE2 downregulates TNF alpha-induced ICAM-1 expression in HGF, via EP2 and EP4 receptors by cAMP dependent signaling pathways, which may result in control of inflammatory and immunological responses in periodontal disease. PMID- 10697806 TI - Role playing in smoking prevention: use caution. PMID- 10697807 TI - Mother calls for more outreach. PMID- 10697808 TI - Youth risk behavior surveillance. National Alternative High School Youth Risk Behavior Survey, United States, 1998. AB - Alternative high schools serve approximately 280,000 students nationwide who are at high risk for failing or dropping out of regular high school or who have been expelled from regular high school because of illegal activity or behavioral problems. Such settings provide important opportunities for delivering health promotion education and services to these youth and young adults. However, before this survey, the prevalence of health-risk behaviors among students attending alternative high schools nationwide was unknown. The Youth Risk Behavior Surveillance System (YRBSS) monitors the following six categories of priority health-risk behaviors among youth and young adults: behaviors that contribute to unintentional and intentional injuries; tobacco use; alcohol and other drug use; sexual behaviors that contribute to unintended pregnancy and sexually transmitted diseases (STDs) (including human immunodeficiency virus [HIV] infection); unhealthy dietary behaviors; and physical inactivity. The national Alternative High School Youth Risk Behavior Survey (ALT-YRBS) is one component of the YRBSS; it was conducted in 1998 to measure priority health-risk behaviors among students at alternative high schools. The 1998 ALT-YRBS used a three-stage cluster sample design to produce a nationally representative sample of students in grades 9-12 in the United States who attend alternative high schools. The school response rate was 81.0%, and the student response rate was 81.9%, resulting in an overall response rate of 66.3%. This report summarizes results from the 1998 ALT-YRBS. The reporting period is February-May 1998. In the United States, 73.6% of all deaths among youth and young adults aged 10-24 years results from only four causes--motor vehicle crashes, other unintentional injuries, homicide, and suicide. Results from the 1998 ALT-YRBS demonstrate that many students at alternative high schools engage in behaviors that increase their likelihood of death from these four causes. During the 30 days preceding the survey, 51.9% had ridden with a driver who had been drinking alcohol, 25.1% had driven a vehicle after drinking alcohol, 32.9% had carried a weapon, 64.5% had drunk alcohol, and 53.0% had used marijuana. During the 12 months preceding the survey, 15.7% had attempted suicide, and 29.0% had rarely or never worn a seat belt. Substantial morbidity among school-aged youth and young adults also results from unintended pregnancies and STDs, including HIV infection. ALT-YRBS results indicate that in 1998, a total of 87.8% of students at alternative high schools had had sexual intercourse, 54.1% of sexually active students had not used a condom at last sexual intercourse, and 5.7% had ever injected an illegal drug. Among adults aged > or = 25 years, 66.5% of all deaths result from two causes--cardiovascular disease and cancer. Most risk behaviors associated with these causes of death are initiated during adolescence. In 1998, a total of 64.1% of students at alternative high schools had smoked cigarettes during the 30 days preceding the survey, 38.3% had smoked a cigar during the 30 days preceding the survey, 71.2% had not eaten > or = 5 servings of fruits and vegetables during the day preceding the survey, and 81.0% had not attended physical education (PE) class daily. Comparing ALT-YRBS results with 1997 national YRBS results demonstrates that the prevalence of most risk behaviors is higher among students attending alternative high schools compared with students at regular high schools. Some risk behaviors are more common among certain sex and racial/ethnic subgroups of students. ALT YRBS data can be used nationwide by health and education officials to improve policies and programs designed to reduce risk behaviors associated with the leading causes of morbidity and mortality among students attending alternative high schools. PMID- 10697809 TI - Preventing eating and body image problems in children and adolescents using the Health Promoting Schools Framework. AB - This paper outlines the Health Promoting Schools Framework and how it may be implemented in schools for preventing eating and body image problems. Discussion focuses on the efficacy of preventive school-based strategies, and on the safest and most successful interventions. The Framework encompasses three major areas of intervention in the school and community: 1) School curriculum, teaching, and learning; 2) School ethos, environment, and organization; and 3) School-community partnerships and services. Suggested strategies for implementing the Framework are outlined. A case study of how a girls high school adapted the new approach for dealing with the problem of eating and body image problems is presented. PMID- 10697810 TI - Adolescents' use of school-based health clinics for reproductive health services: data from the National Longitudinal Study of Adolescent Health. AB - Offering reproductive health services to students through school-based clinics (SBCs) may be a valuable public health strategy. Using data from the National Longitudinal Study of Adolescent Health, this report describes adolescents' use of SBCs for family planning and STD-related services. Of more than 1,200 students receiving reproductive health services in the year preceding the survey, 13.3% received family planning services from a SBC and 8.9% received STD-related services. Rural residence, no driver's license, younger age, and minority ethnicity increased the likelihood of using a SBC for family planning services. Rural residence, minority ethnicity, male gender, having a physical exam from a SBC, and less perceived parental approval of sex increased the likelihood of using a SBC for STD-related services. Further research should determine factors that increase adolescents' acceptance of reproductive health services from a SBC. PMID- 10697811 TI - The effectiveness of tobacco control in California schools. PMID- 10697812 TI - The "Shaqweeta Fake" talk show. PMID- 10697813 TI - American Academy of Pediatrics sponsors media matters. PMID- 10697814 TI - Secondary ion images of the rodent brain. AB - Sections of biologic tissue obtained from laboratory rodents are prepared and analyzed by secondary ion mass spectrometry. The intensity of phosphocholine secondary ions is used to identify anatomical features of the brain from secondary ion images and to evaluate the effectiveness of procedures developed. Secondary ion emission of phosphocholine (m/z 184), is found to be abundant and its intensity is heterogeneous. Effects of sample thickness are addressed. Correspondence between conventional optical images of stained tissue and secondary ion images shows that successive ion images may be used to produce a three-dimensional map of the brain, i.e., an atlas. PMID- 10697815 TI - Collisionally activated dissociations of aminocyclitol-aminoglycoside antibiotics and their application in the identification of a new compound in tobramycin samples. AB - Several aminocyclitol-aminoglycoside antibiotics have been studied by tandem mass spectrometry. Glycosidic bond cleavages were the major reactions in the low energy collisionally activated decomposition (CAD) of the protonated antibiotics. Only the glycoside residing on the C6-O of the 2-deoxystreptamine was observed to undergo significant decomposition at the C2-C3 and O-C1 bonds. The comprehension of the CAD of known aminoglycosides aided in the identification of an unknown impurity in tobramycin. The unknown compound was initially detected by reverse phase high-performance liquid chromatography following dinitrofluorobenzene derivatization of the amino groups. The molecular weight of the dinitrobenzene derivative measured by LC mass spectrometry (MS) led to the detection of two isomeric impurities in tobramycin by LC-MS using an amino column. Their CAD spectra were subsequently acquired by LC-MS/MS. One of the two compounds was determined to be a known compound, 6"-O-carbamyltobramycin with the carbamyl group substituted on the glycoside residing on the C6-O of 2-deoxystreptamine. The fragmentation pattern of the other compound was interpreted as that the unknown was also a carbamyltobramycin. The carbamyl group was, however, substituted on 2-deoxystreptamine. It was speculated that the carbamyl group was substituted at the C1 amino group. This compound, to our knowledge, has not been reported before. PMID- 10697816 TI - Mechanistic studies of multipole storage assisted dissociation. AB - The degree and onset of fragmentation in multipole storage assisted dissociation (MSAD) have been investigated as functions of several hexapole parameters. Strict studies of hexapole charge density (number of ions injected) and hexapole storage time were made possible by placing a pulsed shutter in front of the entrance to the mass spectrometer. The results obtained show that the charge density is the most critical parameter, but also dependencies on storage time, radio-frequency (rf) -amplitude, and pressure are seen. From these data, and from simulations of the ion trajectories inside the hexapole, a mechanism for MSAD, similar to the ones for sustained off-resonance irradiation (SORI), and for low energy collisionally induced dissociation in the collision multipole of a triple quadrupole mass spectrometer, is proposed. It is believed that, at higher charge densities, ions are pushed to larger hexapole radii where the electric potential created by the rf field is higher, forcing the ions to oscillate radially to higher amplitudes and thereby reach higher (but still relatively low) kinetic energies. Multiple collisions with residual gas molecules at these elevated energies then heat up the molecules to their dissociation threshold. Further support for this mechanism is obtained from a comparison of MSAD and SORI spectra which are almost identical in appearance. PMID- 10697817 TI - In-source decay of hyperbranched polyesteramides in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. AB - Hyperbranched polyesteramides (DA2), prepared from hexahydrophthalic anhydride (D) and diisopropanolamine (A) have been characterized, by use of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS), field desorption (FD)-MS, and electrospray ionization (ESI)-MS. MALDI of polyesteramides produces protonated molecules. The spectra show a complex chemical composition distribution and end-group distribution which are mainly composed of two series of homologous oligomers DnA(n)+1 - mH2O and DnA(n) - mH2O, where m = 1-2. Signals from protonated molecules DnAn+1 and DnAn are almost absent in the MALDI spectrum, whereas these ions are responsible for the base peak of DnA(n)+1 - mH2O and DnA(n) - mH2O (m = 1-2) clusters in the ESI spectrum. The absence of -OH end-groups signals in the MALDI spectrum is due to a metastable decay of protonated DnA(n)+1 and DnAn ions in the ion source of the MALDI mass spectrometer prior to ion extraction. In-source decay results in the formation of protonated lower DnA(n)+1 - mH2O and DnA(n) - mH2O oligomers and their corresponding neutrals, leading to wrong conclusions concerning the relative end-group distribution as a function of the degree of polymerization and the chemical composition. PMID- 10697818 TI - Noncovalent association phenomena of 2,5-dihydroxybenzoic acid with cyclic and linear oligosaccharides. A matrix-assisted laser desorption/ionization time-of flight mass spectrometric and X-ray crystallographic study. AB - D-Glucose and 19 glucose derivatives were investigated by positive and negative ion matrix assisted laser desorption/ionization time-of-flight mass spectrometry using 2,5-dihydroxybenzoic acid (DHB) as the matrix. The set of substrates includes oligomers of amylose and cellulose, native alpha-, beta-, and gamma cyclodextrin, and chemically modified beta- and gamma-cyclodextrins. These analytes were chosen to modulate molecular weight, polarity, and capability of establishing noncovalent interactions with guest molecules. In the negative-ion mode, the DHB matrix gave rise to charged multicomponent adducts of type [M + DHB - H]- (M = oligosaccharide) selectively for those analytes matching the following conditions: (i) underivatized chemical structure and (ii) number of glucose units > or = 4. In the positive-ion polarity, only some amylose and cellulose derivatives and methylated beta-cyclodextrins provided small amount of cationized adducts with the matrix of type [M + DHB + X]+ (X = Na or K), along with ubiquitous [M + X]+ ions. The results are discussed by taking into account analyte-matrix association phenomena, such as hydrogen bond and inclusion phenomena, as a function of the molecular structure of the analyte. The conclusions derived by mass spectrometric data are compared with the X-ray diffraction data obtained on a single crystal of the 1:1 alpha-cyclodextrin - DHB noncovalent adduct. PMID- 10697819 TI - Electrospray ionization Fourier transform ion cyclotron resonance mass spectrometric analysis of the recombinant human macrophage colony stimulating factor beta and derivatives. AB - The potential of electrospray ionization (ESI) Fourier transform ion cyclotron mass spectrometry (FTICR-MS) to assist in the structural characterization of monomeric and dimeric derivatives of the macrophage colony stimulating factor beta (rhM-CSF beta) was assessed. Mass spectrometric analysis of the 49 kDa protein required the use of sustained off-resonance irradiation (SORI) in-trap cleanup to reduce adduction. High resolution mass spectra were acquired for a fully reduced and a fully S-cyanylated monomeric derivative (approximately 25 kDa). Mass accuracy for monomeric derivatives was better than 5 ppm, after applying a new calibration method (i.e., DeCAL) which eliminates space charge effects upon high accuracy mass measurements. This high mass accuracy allowed the direct determination of the exact number of incorporated cyanyl groups. Collisionally induced dissociation using SORI yielded b- and y-fragment ions within the N- and C-terminal regions for the monomeric derivatives, but obtaining information on other regions required proteolytic digestion, or potentially the use of alternative dissociation methods. PMID- 10697820 TI - Derivatization of protonated peptides via gas phase ion-molecule reactions with acetone. AB - The protonated [M + H]+ ions of glycine, simple glycine containing peptides, and other simple di- and tripeptides react with acetone in the gas phase to yield [M + H + (CH3)2CO]+ adduct ion, some of which fragment via water loss to give [M + H + (CH3)2CO - H2O]+ Schiff's base adducts. Formation of the [M + H + (CH3)2CO]+ adduct ions is dependent on the difference in proton affinities between the peptide M and acetone, while formation of the [M + H + (CH3)2CO - H2O]+ Schiff's base adducts is dependent on the ability of the peptide to act as an intramolecular proton "shuttle." The structure and mechanisms for the formation of these Schiff's base adducts have been examined via the use of collision induced dissociation tandem mass spectrometry (CID MS/MS), isotopic labeling [using (CD3)2CO] and by comparison with the reactions of Schiff's base adducts formed in solution. CID MS/MS of these adducts yield primarily N-terminally directed a- and b-type "sequence" ions. Potential structures of the b1 ion, not usually observed in the product ion spectra of protonated peptide ions, were examined using ab initio calculations. A cyclic 5 membered pyrrolinone, formed by a neighboring group participation reaction from an enamine precursor, was predicted to be the primary product. PMID- 10697821 TI - Rapid confirmation/quantitation of cocaine and benzoylecgonine in urine utilizing high performance liquid chromatography and tandem mass spectrometry. AB - A rapid, but sensitive and selective method for confirmation and quantitation of benzoylecgonine (BZE) and cocaine (COC) in urine by fast-gradient liquid chromatography/tandem mass spectrometry (LC/MS/MS) is described. The chromatographic separation was performed on a reversed phase column employing fast-gradient techniques. Matrix prepared standards, blanks, and QC's were filtered then aliquots were transferred into a 96 well plate. Injection volumes of 25 microL were made onto the analytical column, with the flow diverted from the atmospheric pressure ionization source for the first 0.5 min of the analysis. Simultaneous multiple reaction monitoring (MRM) of three discrete transitions for each compound were used to identify BZE and COC. Quantitation was achieved utilizing the most prominent parent-daughter transition and internal standard calibration techniques. The coefficients of variation (CV) for the analysis of these drugs ranged from 0.6% to 6.8% at a concentration of 150 ng/mL (n = 155). This method suggests that fast-gradient LC/MS/MS may be suitable for routine confirmation of immunoassay cocaine-positive samples. PMID- 10697822 TI - hSKCa3: no association of the polymorphic CAG repeat with bipolar affective disorder and schizophrenia. AB - hSKCa3 is a neuronal small conductance calcium-activated potassium channel, which contains a polyglutamine tract, encoded by a polymorphic CAG repeat in the gene. Since an association between longer alleles of this CAG repeat and bipolar disorder or schizophrenia has been reported, we genotyped the polymorphic CAG repeat in 91 German family trios of patients with bipolar disorder I and used the transmission disequilibrium test (TDT) to test for association. Applying a dichotomized model (< or = 19 or > 19 CAG triplets), we found no evidence for an association of longer alleles with bipolar disorder (TDT = 0.75, P = 0.386). Regarding the whole range of alleles, there was no preference in transmitting the larger of the two observed alleles from parents to the affected offspring. In parallel we performed an independent case-control study on German patients with bipolar disorder and schizophrenia. Again we did not detect an overrepresentation of longer CAG repeats in patients. Thus, our data do not support the hypothesis that longer CAG repeats in the hSkCa3 gene contribute to the susceptibility for bipolar disorder and schizophrenia. PMID- 10697823 TI - A study of linkage disequilibrium between polymorphic loci for monamine oxidases A and B in schizophrenia. AB - Two X-linked microsatellites, (AC) n repeats at the monoamine oxidase (MAO)-A locus and (TG)n repeats at the MAO-B locus, were studied in 140 unrelated Caucasian male patients with schizophrenia and 91 unrelated Caucasian male controls. Among these subjects, we totally typed out nine alleles for the (AC) n repeats and eight alleles for the (TG) n repeats by using a PCR-based procedure. Allelic frequencies of either (AC) n repeats or (TG) n repeats were not found to be significantly different between patients and controls. However, a significant excess of the (AC)18/(TG)23 haplotype with a relative risk of 4.05 (95%; CI 1.15 14.26) was observed in patients with schizophrenia (Fisher's P = 0.011). The coefficient of linkage disequilibrium (delta) for the (AC)18/(TG)23 haplotype was 0.019 in schizophrenic patients and -0.046 in control subjects, respectively. The latter reached statistical significance (chi 2 = 6.02; df = 1; P < 0.02). The present findings suggest that linkage disequilibrium between polymorphic loci for human MAO-A and MAO-B may be associated with schizophrenia, and the (AC)18/(TG)23 haplotype may render an individual more vulnerable to such an illness. PMID- 10697824 TI - Lack of evidence for association between the COMT locus and schizophrenia. AB - Family-based studies have been conducted with restriction fragment length polymorphism (RFLP) analysis for testing association between polymorphisms for the catechol-O-methyltransferase (COMT) locus and schizophrenia in 49 Caucasian nuclear families consisting of fathers, mothers and offspring affected with schizophrenia. The present results did not support the hypothesis that the COMT gene might play an important role in predisposing an individual to a genetic risk for schizophrenia. Neither did we find a significant association of the COMT locus with violent behaviour in schizophrenia. Nevertheless, there may be a susceptibility gene for schizophrenia in a distinct region from the COMT locus on chromosome 22q, as a genome scan has suggested recently. PMID- 10697825 TI - Two single nucleotide polymorphisms in the promoter region of the human phenylethanolamine N-methyltransferase PNMT gene. PMID- 10697826 TI - Association between the dopamine D3 receptor gene locus (DRD3) and unipolar affective disorder. AB - Dopamine neurotransmission has been implicated in the pathophysiology of schizophrenia and, more recently, affective disorders. Among the dopamine receptors, D3 can be considered as particularly related to affective disorders due to its neuroanatomical localization in the limbic region of the brain and its relation to the serotoninergic activity of the CNS. The possible involvement of dopamine receptor D3 in unipolar (UP) major depression was investigated by a genetic association study of the D3 receptor gene locus (DRD3) on 36 UP patients and 38 ethnically matched controls. An allelic association of DRD3 (Bal I polymorphism) and UP illness was observed, with the Gly-9 allele (allele '2', 206/98 base-pairs long) being more frequent in patients than in controls (49% vs 29%, P < 0.02). The genotypes containing this allele (1-2 and 2-2) were found in 75% of patients vs 50% of controls (P < 0.03, odds ratio = 3.00, 95% CI = 1.12 8.05). The effect of the genotype remained significant (P < 0.02) after sex and family history were controlled by a multiple linear regression analysis. These results further support the hypothesis that dopaminergic mechanisms may be implicated in the pathogenesis of affective disorder. More specifically, the '2' allele of the dopamine receptor D3 gene seems to be associated with unipolar depression and can be considered as a 'phenotypic modifier' for major psychiatric disorders. PMID- 10697827 TI - No evidence for linkage of schizophrenia to DXS7 at chromosome Xp11. AB - There have been claims that a gene on the X chromosome may contribute to susceptibility to schizophrenia. Crow (1988) initially proposed that such a gene might lie in the pseudoautosomal region, but when evidence that weakened this hypothesis accumulated, he proposed that a susceptibility locus might be present elsewhere on the sex chromosomes instead. DeLisi et al. (1994) found a small nonsignificant positive lod score between the marker DXS7 and schizophrenia, but other failed to replicate this finding. Another study reported by Crow and DeLisi's group was also weakly positive for this marker (Dann et al., 1997). This locus was then investigated in a collaborative study by Laval et al. (1997), which produced a nonparametric lod score of 2.44. Using a sample of 17 pedigrees from Britain and Iceland, we have also tested the hypothesis of linkage between DXS7 and schizophrenia. The 17 families were selected from a larger sample on the basis of an absence of male-to-male transmission for schizophrenia. These families were originally selected for having multiple cases of schizophrenia within them and for having no cases of bipolar affective disorder. We genotyped subjects for a marker at DXS7 and performed classical lod score and model-free linkage analysis using broad and narrow definitions of affection with schizophrenia. We found strongly negative lod scores and no evidence for linkage using model-free analysis. Therefore, this study does not support the hypothesis of linkage of schizophrenia to DXS7, and the evidence for a susceptibility locus on this part of the X chromosome is weakened. PMID- 10697828 TI - Chromosome 18p11 and linkage to bipolar disorder revisited. PMID- 10697829 TI - The last and the next hundred years of malariology. AB - The founding fathers of malariology combined scientific originality, perseverance in research, strong characters, breadth of interest and social concern. A hundred years later research and understanding has made immense progress but the world still bears a huge burden of malaria. For the next century research requires both more specialism and a holistic range if it is to be used in control, requiring multidisciplinary team work. Environmental changes and interventions produce a dynamic and changing pattern of malaria, not the static one of the past. From the original parasite life cycle, research has analysed a series of other cycles at electron microscope, biochemical and genome levels on decreasing size scales and quantitative epidemiological cycles for control. Recent additions to these concepts have been stage-specific antigens, cycles of disease rather than parasites alone, considering populations of parasites rather than just cases, and also genetic variation in each component of the parasite-human host-vector triad. In this volume there emerges for the first time a coherent overall picture of the biomedical aspects of basic malariology as the interacting population genetics of malaria parasites, anophelines and people. This provides a coherent model for the new century dealing with the great biological malaria problems of drug resistance, vaccine development, insecticidal and net control and can feed, with socio-economic work, into the gathering renewal of control efforts. New work on large-scale changes of malaria in space and time enables us to be precise about effects of local and global environmental changes to predict epidemics. Future research will be as much about linking these different scales of understanding as control will be about linking different levels of the health system. The grim situation in poor holoendemic countries also requires practical support of the type that the founders of malariology were involved in. A coherent understanding needs to feed into the new control efforts, from Roll Back Malaria onwards, for the next century. PMID- 10697830 TI - The malariology centenary. AB - The year 1898 was one of the most significant years in the history of malariology. One hundred years later scientists gathered at the Accademia Nazionale dei Lincei, Rome, to commemorate the Malariology Centenary. This paper provides a short overview of some of the key developments and discoveries in malaria research which took place at the end of the 19th century. The major contributions of Alphonse Laveran, Patrick Manson, Ronald Ross, Battista Grassi and a number of scientists of the Italian School of Malariology to the understanding of the transmission of malaria by Anopheles mosquitoes are described. This paper also highlights the importance of an historical perspective in furthering our understanding of the 'Malaria Challenge after One Hundred Years of Malariology'. PMID- 10697831 TI - Malaria diseases and parasites. AB - The milestones in the discovery of malaria parasites and their relationships with malaria diseases are presented and discussed with particular reference to the contribution of the Italian scientists. Laveran's discovery (1880) of the malaria parasite produced some schepticism among the Roman scientists who were under the influence of Tommasi-Crudeli, the discoverer of the supposed Bacillus malariae. However, Marchiafava and Celli confirmed soon Laveran's observations and, between 1883 and 1885, improved the description of the parasite adding important details. They described, then, the aestivo-autumnal tertian fever as a distinct disease from the 'primaverile' or benign tertian. This work influenced Golgi who went on to analyse the features that distinguish the benign tertian parasite from that of the quartan. The fact that in North Italy the aestivo-autumnal tertian fever was hardly ever found, whereas it was common in the Roman Campagna and the Pontin marshes, explains why it was Celli and Marchiafava and later Bignami and Bastianelli, and Marchiafava and Bignami--but not Golgi--who were committed to work on this pernicious form of malaria. By the early 1890s the Italian scientists came to define the three malaria parasites, presently known as Plasmodium vivax, P. malariae, and P. falciparum, and to associate them with precise anatomo-pathological and clinical features. By the middle 1890s the Italian school was prepared to contribute also to the discovery of the mosquito cycle in human malaria, clearly hypothesized by Bignami in 1896 and experimentally proved in 1898 by Bignami, Bastianelli and Grassi. PMID- 10697832 TI - The concept of specificity and the Italian contribution to the discovery of the malaria transmission cycle. AB - At the dawn of the 20th century, the change in the scientific, political and cultural attitudes towards malaria was the result of the discovery of the theoretical simplicity of the malaria transmission cycle and of the possibility to interrupt it, by avoiding the contacts between people and mosquitoes. The 'mosquito hypothesis', suggested already in the 1880s, had to be included into a coherent scientific theory, in which a fundamental part was played by the concept of specificity. The paper analyses the Italian contribution to this scientific change and the epistemological aspects of the debate between Ronald Ross and Battista Grassi about their respective role in the discovery of the human malaria transmission cycle. This debate has been often interpreted in sociological or psychological terms. However, behind the dispute there is a different definition of what is a scientific explanation in biological sciences and in particular in parasitology. This point is made clear by the analysis of four different theoretical problems implied in the discovery of the transmission cycle: the concept of specificity, the comparative method in parasitology, the specificity of the life-cycle of parasites and vectors, and the role of the analogical reasoning in science and medicine. PMID- 10697833 TI - Ronald Ross and the malaria-mosquito cycle. AB - This essay examines briefly Ronald Ross's research on malaria in India between 1895 and 1899, during which time he elucidated the role of the Anopheles mosquito in the transmission of the disease. During his crucial experimental work in 1898, he used birds, human malaria being relatively uncommon in Calcutta. This article is based largely on the correspondence between Ross and Patrick Manson, which has just been edited. PMID- 10697834 TI - Evolution of Plasmodium and the recent origin of the world populations of Plasmodium falciparum. AB - We have investigated the evolution of Plasmodium parasites by analyzing DNA sequences of several genes. We reach the following conclusions: (1) The four human parasites, P. falciparum, P. malariae, P. ovale, and P. vivax are very remotely related to each other, so that their evolutionary divergence predates the origin of the hominids; several of these parasites became associated with the human lineage by lateral transfer from other hosts. (2) P. falciparum diverged from P. reichenowi about 8 million years ago, consistently with the time of divergence of the human lineage from the apes; a parsimonious inference is that falciparum has been associated with humans since the origin of the hominids. (3) P. malariae is genetically indistinguishable from P. brasilianum, a parasite of New World monkeys; and, similarly. (4) P. vivax is genetically indistinguishable from the New World monkey parasite P. simium. We infer in each of these two cases a very recent lateral transfer between the human and monkey hosts, and explore alternative hypotheses about the direction of the transfer. We have also investigated the population structure of P. falciparum by analyzing 10 genes and conclude that the extant world populations of this parasite have evolved from a single strain within the last several thousand years. The extensive polymorphisms observed in the highly repetitive central region of the Csp gene, as well as the apparently very divergent two classes of alleles at the Msa-1 gene, are consistent with this conclusion. PMID- 10697835 TI - The malaria genome sequencing project: complete sequence of Plasmodium falciparum chromosome 2. AB - An international consortium has been formed to sequence the entire genome of the human malaria parasite Plasmodium falciparum. We sequenced chromosome 2 of clone 3D7 using a shotgun sequencing strategy. Chromosome 2 is 947 kb in length, has a base composition of 80.2% A + T, and contains 210 predicted genes. In comparison to the Saccharomyces cerevisiae genome, chromosome 2 has a lower gene density, a greater proportion of genes containing introns, and nearly twice as many proteins containing predicted non-globular domains. A group of putative surface proteins was identified, rifins, which are encoded by a gene family comprising up to 7% of the protein-encoding gene in the genome. The rifins exhibit considerable sequence diversity and may play an important role in antigenic variation. Sixteen genes encoded on chromosome 2 showed signs of a plastid or mitochondrial origin, including several genes involved in fatty acid biosynthesis. Completion of the chromosome 2 sequence demonstrated that the A + T-rich genome of P. falciparum can be sequenced by the shotgun approach. Within 2-3 years, the sequence of almost all P. falciparum genes will have been determined, paving the way for genetic, biochemical, and immunological research aimed at developing new drugs and vaccines against malaria. PMID- 10697836 TI - Evolution of the human genome under selective pressure from malaria: applications for control. AB - Research on the molecular basis for resistance of humans to malaria has been vigorous during the last 10 years, with new discoveries and extension of work from previous decades. Much of the work has important implications both for understanding pathogenesis and for applications for control of disease. PMID- 10697837 TI - Genetic dissection of Plasmodium falciparum blood infection levels and other complex traits related to human malaria infection. AB - There is accumulating evidence of host genetic control in malaria infection and, in humans, some genes have been associated with severe malaria. Nevertheless, other important genes controlling blood infection levels, malarial disease and immune responses are likely to be identified. In this paper, we focus on segregation and linkage analyses of blood infection levels in an urban population living in Burkina Faso. We found evidence of a complex genetic control and a linkage to chromosome 5q31-q33. The identification of genes controlling complex traits related to malaria infection should be helpful in understanding protective mechanisms and the relationship between infection, malaria attacks and severe malaria. PMID- 10697838 TI - Molecular evolution of coding and non-coding regions in Plasmodium. AB - Recurrence analysis provides a useful tool for the characterisation of oligonucleotide usage along genomic tracts. While coding regions are characterised by a low-recurrence regimen (except in the case of intragenic repeats) introns and intergenic regions exhibit a high density of recurring oligos, and appear to be correlated from the point of view of oligonucleotide preference. By comparing homologous loci in Plasmodium falciparum and P. berghei, it can be seen that introns and intergenic regions, though exhibiting very low sequence similarity, do not drift without constraints, but maintain a consistent use of the same oligos in the two species. PMID- 10697839 TI - Natural selection on apical membrane antigen-1 of Plasmodium falciparum. AB - The Apical Membrane Antigen-1 (AMA-1) is a protein localized in the apical organelles of the merozoite, one of the stages in the life cycle of malaria parasites (Plasmodium spp.) that infects the vertebrate host. This antigen, which is encoded by a single polymorphic locus, plays a role in evading immune detection and mediating invasion into target host cells. We found evidence of positive Darwinian selection on immunogenic regions of P. falciparum AMA-1 favoring genetic diversity in the T-cell epitopes and in regions likely to interact with host antibodies. These results support the hypothesis that polymorphism at the AMA-1 locus in maintained by balancing selection arising from host immune recognition. PMID- 10697840 TI - Complexities in the analysis of cryptic taxa within the genus Anopheles. AB - Grassi's discovery one hundred years ago brought to light the puzzle of anophelism without malaria in Europe. With the discovery of the European Anopheles maculipennis complex the puzzle was solved but the 'species problem' has not gone away. Meaningful epidemiologic studies and effective vector control programs depend upon efficient methods for discriminating among the major vectors, lesser vectors and non-vectors of ubiquitous anopheline sibling species complexes. We now have a variety of techniques for identifying cryptic species, ranging from crossing studies through morphological, cytogenetic, allozyme and repetitive DNA-based strategies. However, cytogenetic and molecular data can also be used to infer evolutionary relationships among cryptic taxa. This approach has been crucial to understanding the biology of the vector, and may illuminate the speciation process and the human impact upon this process. Nevertheless, the analysis of cryptic taxa has proven unexpectedly complex. Studies of An. funestus and An. gambiae reveal conflicts among classes of markers and between different genomic locations. The data are consistent with a model of speciation in which gene flow may still occur in parts of the genome, and they suggest that caution should be exercised in the interpretation of results from small numbers of loci, only one type of marker, and markers located in specific genomic regions such as chromosomal inversions. PMID- 10697841 TI - Population structure, speciation, and introgression in the Anopheles gambiae complex. AB - We review here what is known about the population structure and evolutionary dynamics of members of the Anopheles gambiae complex with emphasis on the situation in West Africa. First, the importance of the 2nd chromosome inversion polymorphism is demonstrated especially in adaptation to levels of aridity, a major environmental variable in Africa. This affects the distribution of karyotypes on both a macro- and micro-geographic scale as well as temporally. Such differentiation leads to karyotypes being differentially effective transmitters of malaria and differentially susceptible to indoor residual spraying of insecticides. Second, we review the evidence that cryptic taxa, especially in An. gambiae s.s., exist. This observation stems from both karyotype studies and molecular studies. It is abundantly clear that West African populations of An. gambiae s.s. are often not panmictic units, with premating factors evidently acting to maintain distinct genetic forms. Third, we review phylogenetic studies that have revealed the presence of introgression between the two most important vectors, An. gambiae and An. arabiensis. This is most evident for the 2nd chromosome inversions. This interpretation of phylogenetic data is consistent with a direct laboratory study indicating inversions in this chromosome are stably maintained in back-crossed populations. All of this information has led to the view that members of the An. gambiae complex are highly variable with an abundance of adaptive genetic variation. This presents a significant challenge to vector control programs designed to reduce malaria in sub-Saharan Africa. PMID- 10697842 TI - Molecular identification of chromosomal forms of Anopheles gambiae sensu stricto. AB - The Afrotropical malaria vectors Anopheles gambiae sensu stricto and An. arabiensis, responsible for more than 3/4 of the world Plasmodium falciparum inoculations, are members of the Anopheles gambiae complex, a group consisting of six sibling species. The nominal species (An. gambiae s.s.) is by far the most anthropophilic vector and its adaptation to man and his environment involves further ongoing speciation processes. This fact is shown by the existence of a number of incipient taxonomic units characterised by different chromosomal arrangements derived from the presence of polymorphic paracentric inversions. This speciation process is centered in West Africa, where five so-called 'chromosomal forms' have been described, designated with a non-Linnean nomenclature: Forest, Bissau, Savanna, Bamako, and Mopti. Studies on the distribution and the ecology of these incipient species have highlighted specific adaptations to eco-ethological parameters, which might reflect on their relative efficiency as malaria vectors. Cytogenetic analysis, in spite of some inherent difficulties, has proved to be a powerful tool for the identification of An. gambiae sibling species and the individual chromosomal forms. Yet, modern molecular tools are now available, providing alternative faster low-cost technologies, and we discuss here their relative merits. PMID- 10697843 TI - Are chromosomal inversions induced by transposable elements? A paradigm from the malaria mosquito Anopheles gambiae. AB - Chromosomal rearrangements abound in nature and can be studied in detail in organisms with polytene chromosomes. In Drosophila and in Anopheline mosquitoes most speciation processes seem to be associated with the establishment of chromosomal rearrangements, particularly of paracentric inversions. It is not known what triggers inversions in natural populations. In the laboratory inversions are commonly generated by X-rays, mutagens or after the activity of certain transposable elements (TEs). The Anopheles gambiae complex is comprised of six sibling species, each one characterized by the presence of fixed paracentric inversions on their chromosomes. Two of these, An. gambiae s.s. and An. arabiensis, are the most important vectors of human malaria and are structured into sub-populations, each carrying a characteristic set of polymorphic chromosomal inversions. We have cloned the breakpoints of the naturally occurring polymorphic inversion In(2R)d' of An. arabiensis. Analysis of the surrounding sequences demonstrated that adjacent to the distal breakpoint lies a transposable element that we called Odysseus. Characteristics of Odysseus' terminal region and its cytological distribution in different strains as well as within the same strain indicate that Odysseus is an actively transposing element. The presence of Odysseus at the junction of the naturally occurring inversion In(2R)d' suggests that the inversion may be the result of the TEs activity. Cytological evidence from Drosophila melanogaster has also implicated the hobo transposable element in the generation of certain Hawaiian endemic inversions. This picture supports the hypothesis of the important role of TEs in generating natural inversions. PMID- 10697844 TI - Genetics and evolution of parasites and hosts: some comments. AB - We provide a brief commentary on aspects of the analysis of the genetics and evolution of malaria parasites. Any attempt to understand the nature and manifestations of an infectious disease requires an understanding of the genetics of both pathogen and host. The outcome of a malaria infection, i.e. whether it is asymptomatic, mild, severe or causes cerebral malaria, is due to a complex interaction between the products of parasite and host genes. In general terms, genes in the parasite determine its ability to infect the host, its virulence, etc., while host genes will determine resistance or susceptibility to infection. More than this, however, genetics is about the spread of genes in populations, how they mutate and recombine to produce novel genotypes, and how the parasite and its hosts co-evolve with changing environments. This is a complex subject, and we present some discussion of a few aspects of its analysis. PMID- 10697845 TI - Immune regulation of protection and pathogenesis in Plasmodium falciparum malaria. AB - The immune mechanisms whereby malaria parasites are eliminated by the human host or how they may avoid the immune response are poorly understood. Individuals living in malaria-endemic areas gradually acquire immunity. It is well established that this immunity involves both cell-mediated and humoral mechanisms and that T cells are the major regulators in both these events. The existence of functionally distinct P. falciparum-specific CD4+ T-cell subsets in humans has been shown in several studies. However, in contrast to what is the case in murine models there is no definitive link between the activation of various T cells and the course of human P. falciparum blood-stage infection. In the present paper we will review recent findings which illustrate how the balance between functionally different T-cell subsets affects the development of malaria immunity but also may contribute to its pathogenicity. An example of the latter is the deposition of IgE-containing immune complexes in small vessels, probably leading to local overproduction of tumor-necrosis factor (TNF), a pathogenic factor in malaria. PMID- 10697846 TI - Plasmodium differentiation in the mosquito. AB - The essential passage of the malarial parasite through a mosquito vector results in major population bottlenecks in parasite numbers. The volume of the bloodmeal ingested by the female mosquito is 1-2 microliters. This may contain from 1 to 10(5) gametocytes. Of these, it is normal for just 12 to become macrogametes; 5-6 become ookinetes, and 2 develop into oocysts 2-7 days later. Of the 16,000 sporozoites produced from these two oocysts just 10-20 are inoculated by the malaria-infected female mosquito each time she probes when taking a subsequent bloodmeal. These significant population bottlenecks suggest that parasite differentiation is severely constrained by the environment in the mosquito, and therefore by the interactions between the parasite and the vector. This review will describe parasite differentiation in the mosquito and try to highlight the more important interactions between the parasite, the bloodmeal and the mosquito, attempting to identify those interactions which are essential to parasite differentiation, and those where the mosquito may be mounting effective strategies against this important pathogen. The potential exploitation of these interactions as possible mechanisms for intervention will be discussed. PMID- 10697847 TI - Genome plasticity and sexual differentiation in Plasmodium. AB - Spontaneous subtelomeric deletions of Plasmodium chromosomes have been observed both in natural infections and in laboratory maintained parasites. In the latter case, functions dispensable for asexual parasite multiplication and encoded at the extremities of the chromosomes are easily lost. In particular, spontaneous subtelomeric deletions have been characterised which affect gametocytogenesis both in Plasmodium berghei maintained in laboratory animals and in Plasmodium falciparum propagated in in vitro cultures. In order to identify these genetic determinants, and, potentially, other genes located subtelomerically, we designed a transfection system able to induce and select for controlled, site-specific subtelomeric deletions. PMID- 10697848 TI - Sex ratio adjustment in Plasmodium gallinaceum. AB - The sex ratio of the avian malaria parasite, Plasmodium gallinaceum, was examined during the course of infection in its natural host, the chicken. Infections can have two possible outcomes: death of the host resulting from anaemia or self-cure and survival. In lethal infections the sex ratio remained female biased throughout, whereas in self-curing infections, the sex ratio became progressively less female biased. We examined the consequences of altering sex ratio for parasite transmission success using a theoretical fertilisation model and hypothesise that an immune response specifically effective against the male gametes would provide a selective explanation for the observed sex ratio adjustment. Previous studies have demonstrated that there is an efficient anti gamete antibody response as an infection is cleared by the host. We performed in vitro mosquito infection studies comparing mosquito infection rates with naive serum replacement and showed that decomplemented serum from curing infections is transmission blocking, whereas serum from lethal infections is not. We discuss aspects of the malaria parasite-host interaction which might provide the selective pressure for such observed sex ratio adjustment. PMID- 10697849 TI - Transmission stages of Plasmodium: does the parasite use the one same signal, provided both by the host and the vector, for gametocytogenesis and sporozoite maturation? AB - Among the microorganisms that strictly depend upon other organisms (hosts or vectors) for achieving their life cycle, protozoan and metazoan parasites have been often primarily distinguished through the major pathogenic processes they could induce. A variety of different mechanisms linked to parasitism can indeed systemically (e.g. Plasmodium falciparum) or locally (e.g. Toxoplasma gondii) induce important alterations of tissue homeostasis. But more than obvious pathogenicity, it is the capacity to be transmitted that is essential for parasite survival and there is increasing evidence that certain parasites can achieve their life cycle to the point of transmission in the absence of clinically detectable processes. For this, constitutive microenvironments of the host or vector can be exploited. Moreover, parasites are sometimes able to highjack effectors of the host's immune response towards conditioning the microenvironments which are permissive to differentiation of transmissible developmental stages. Based on a few examples taken from studies on the transmission stages of Leishmania, Toxoplasma and Plasmodium, we have here attempted to formulate a few hypothesis on the biology of the transmission stages of P. falciparum, i.e. on gametocytogenesis and sporozoite maturation. As discussants, we may have been somewhat dwarfed by issues evoked by the organizers of this meeting in the title of the session, i.e. 'Vector-parasite-man interactions'!... In reaction, we may have taken refuge in somewhat over selective comments, biased by the objects of our personal research.... PMID- 10697850 TI - Genetics in the study of mosquito susceptibility to Plasmodium. AB - Within the past several years, a number of powerful genetic and genomic tools have been developed for use in research on the African malaria vector Anopheles gambiae. While these tools have been developed with a broad range of potential applications in mind, they have been particularly useful in advancing the effort to clone a set of An. gambiae genes that enable a refractory strain of this mosquito to encapsulate and kill a wide variety of different malaria parasites to which this mosquito is normally fully susceptible. This paper describes the latest progress in this map-based cloning research, which involves the collaborative contributions of a number of different laboratories in Europe and the United States. PMID- 10697851 TI - How does Anopheles gambiae kill malaria parasites? AB - The malaria parasite's lifecycle in the mosquito vector Anopheles gambiae involves several translocations within and between tissues during which large parasite losses have been documented. Interestingly, during the critical transition stages of midgut invasion and relocation of sporozoites from the oocysts to the salivary glands the mosquito innate immune system is activated. These defense reactions could, at least partially, be responsible for the parasite killing in the mosquito. This important question is now being approached by the dissection of the mosquito innate immune system as well as genetic and genomic studies. PMID- 10697852 TI - Host-parasite intimacy: how do mosquito defense reactions affect Plasmodium sporogonic development? AB - Here are a few sundry reflections and questions stimulated by the talks and discussions in session 3 of the Malariology Centenary Conference 1998 on the 'intimacy' that has been established between Plasmodium and Anopheles. The degree of 'intimacy' achieved in a vectorial system seems to correlate to the difficulties incurred when one attempts to interrupt parasite development in the insect host. The main questions addressed in this essay are as follows: Are antimicrobial peptides the most efficient effector molecules to be used to block sporogonic development of malaria parasites? Are the mosquito defense reactions elicited by Plasmodium invasion deleterious, advantageous or neutral to parasite development? The purpose in asking these questions is to put in perspective the direction of research in this area and to generate new ideas. PMID- 10697853 TI - Genetic basis of encapsulation response in Anopheles gambiae. AB - The phenomenon of encapsulation of invading organisms is widespread in insects. Co-evolution has produced an intricate balance between the immune responses of the host and immune-suppressive (or immune-evading) properties of the parasite. Genome-wide genetic mapping revealed different loci in Anopheline mosquitoes were involved in melanotic encapsulation of different malaria parasites. Certain isolates of human malaria parasites can still suppress or avoid the immune response from refractory mosquitoes. Similar interactions with parasitoids were observed in Drosophila melanogaster. Species-specific encapsulation locus was identified for two parasitoids, respectively, and virulent strain of parasitoid can suppress the immune system of an otherwise resistant fruitfly. It is believed that the encapsulation loci in both mosquitoes and fruitfly may encode gene products that function at the early stages of parasite/parasitoid recognition or immediate signaling events. Future research on membrane receptor molecules and their roles in insect immunity will yield interesting insights into mosquito parasite interactions. PMID- 10697854 TI - Genetic and dietary adaptation to malaria in human populations. AB - Plasmodial invasion places a severe oxidant stress on parasitized erythrocytes which can result in red cell damage and removal within the reticuloendothelial system or lysis, thus interrupting the parasitic cycle. The basis of a number of genetic adaptations to malaria--including the hemoglobin variants, the thalassemias, and glucose-6-phosphate dehydrogenase deficiency--is an increased sensitivity of the variant erythrocytes to the oxidant stress of plasmodial parasitization. It is suggested that dietary adaptations of traditional cusines in human populations living in areas where malaria is endemic augment this oxidant stress. It appears that there are three components of this adaptive dietary pattern in most tropical populations: the consumption of 'oxidant fuels', moderate to high iron intake, and limitation of dietary antioxidant intake or storage. It is argued that this dietary pattern maximizes iron-mediated free radical production in parasitized erythrocytes and thus provides a form of diet mediated antimalarial prophylaxis. African pastoral populations that are heavy consumers of milk appear to manifest a different adaptive pattern involving low intakes of para-aminobenzoic acid, vitamin E, and iron. Periodic food restriction may also contribute to this antimalarial effect. PMID- 10697855 TI - Different response to Plasmodium falciparum in west African sympatric ethnic groups: possible implications for malaria control strategies. AB - The comparison of malaria indicators among populations with different genetic backgrounds and uniformly exposed to the same parasite strains, is one of the approaches to the study of human heterogeneities in the response to the infection. The results of our comparative studies conducted in Burkina Faso, West Africa, showed consistent interethnic differences in Plasmodium falciparum infection rates, malaria morbidity, prevalence and levels of antibodies to various P. falciparum antigens, and genetic background. The differences in the immune response were not explained by the entomological observations which indicated substantially uniform exposure to infective bites. The presence in the same epidemiological context of individuals characterized by different immune reactivity to malaria represents an ideal opportunity to study the possible relationships between the baseline level of anti-malaria immunity of a population and the protective efficacy of control measures based on the reduction of transmission. In spite of similar reduction of entomological inoculation rates obtained by permethrin-impregnated curtains, ethnic- and age-dependent efficacy was observed. These studies demonstrate the existence of marked interethnic differences in the susceptibility to P. falciparum malaria, probably involving the genetic regulation of humoral immune responses. These differences should be considered in the development of anti-malaria vaccines and in the evaluation and application of malaria control strategies. PMID- 10697856 TI - 15(S)-hydroxyeicosatetraenoic acid (15-HETE), a product of arachidonic acid peroxidation, is an active component of hemozoin toxicity to monocytes. AB - Several studies have shown that human and murine hemozoin-fed phagocytes are functionally impaired. Unpurified hemozoin contains unspecifically attached unsaturated fatty acids such as arachidonic and linolenic acids. The presence in unpurified hemozoin of large quantities of ferric heme with small amounts of free iron makes hemozoin a generator of oxidative radicals capable of forming lipoperoxides or other breakdown products from polyunsaturated fatty acids. Here we show that delipidized hemozoin had reduced toxicity to monocytes. Phorbol myristate acetate (PMA)-elicited burst was poorly affected by delipidized hemozoin (ca. 17% and 21% burst inhibition by delipidized hemozoin vs ca. 75% and 65% burst inhibition by native hemozoin at 20 min or 17 h post-phagocytosis, respectively). Analysis of the lipid fraction isolated from native hemozoin by HPLC and chiral-phase HPLC showed equimolar amounts of 15(R)- and 15(S)-HETE (HETE, 15-hydroxy-6,8,11,13-eicosatetraenoic acid), most likely by-products of non-enzymatic peroxidation of arachidonic acid. The biologically active isomer, 15(S)-HETE, the product of 15-lipoxygenase, is a powerful mediator of inflammation and the effector of a large number of bioactions. 15(R,S)-HETE was found in native hemozoin (0.24 millimole/mole hemozoin heme), in supernatants of hemozoin-fed monocytes (87 nMol) and in hemozoin-fed monocytes (9.6 microMol). Approximately 84% of 15-HETE attached to hemozoin was in the esterified form. A large preponderance of esterified over free 15-HETE was also noted in supernatants of hemozoin-fed monocytes and in hemozoin-fed monocytes. In the latter cells, remarkable levels of the substance were attained. A dose-dependent curve of inhibition of PMA-elicited oxidative burst was observed. Assuming homogenuous distribution of 15-HETE in hemozoin-fed monocytes, 15(S)-HETE concentrations measured in hemozoin-fed monocytes (8 muMol) would bring about ca. 85% inhibition of PMA-elicited burst. In conclusion, derivatives of lipoperoxidation of unsaturated fatty acids such as 4-hydroxynonenal, 15-HETE and others now under study, appear to be relevant causes of hemozoin toxicity. PMID- 10697857 TI - Genetic resistance to malaria, oxidative stress and hemoglobin oxidation. AB - I describe a model which posits the molecular basis of some malaria-resistance genes in the interaction between oxidized hemoglobin and membrane components. The model is supported by a considerable body of evidence which indicates that erythrocytes of genetically protected individuals (carriers of sickle cell trait, alpha- and beta-thalassemia, and G6PD deficiency) are susceptible to the increase of oxidation of hemoglobin following H2O2 release in the host cell by Plasmodium falciparum. I suggest that the irreversible interaction between oxidized hemoglobin and the red cell membrane could trigger mechanisms that: (i) reduce invasion of erythrocytes by the falciparum parasite; (ii) impair parasite survival and development within the cell; (iii) accelerate infected erythrocyte clearance by phagocytosis. PMID- 10697858 TI - A fine balance between oxidised and reduced haem controls the survival of intraerythrocytic plasmodia. AB - The polymerisation of haemoglobin-derived ferri-protoporphyrin IX (Fe(III)PPIX) to haemozoin (malaria pigment) is a crucial process for intraerythrocytic plasmodia to prevent haem toxicity. It is also the target of in-use antimalarial drugs and newer compounds. This reaction and the inhibition thereof can be reproduced and studied in vitro. PMID- 10697859 TI - Mosquito behavioural aspects of vector-human interactions in the Anopheles gambiae complex. AB - The behaviour of two of the most anthropophilic malaria vectors in the world, Anopheles gambiae Giles and An. arabiensis Patton, is revisited with respect to recent studies on their host preferences and the chemical ecology of host seeking. Issues are discussed in relation to the ways anthropophily may have arisen in the complex, and the opportunities the study of olfaction and host seeking behaviour offers to malaria control in Africa. PMID- 10697860 TI - Review of intra-host models of malaria. AB - Intra-host models of malaria (and some related models of trypanosomiasis) are reviewed. A first section gives a short description of the different models, their purposes and the authors' conclusions. A second section discusses some common issues, including intra-host populations, the intra-host basic reproduction number (R0) and growth rates, density regulation mechanisms (including acquired immunity), and the models' behaviour compared to that of Plasmodium falciparum in man. PMID- 10697861 TI - The molecular genetic approach to malarial pathogenesis and immunity. AB - It is poorly understood why some malarial infections are fatal while others resolve without complications. Host genetic factors are partly responsible. More than ten specific susceptibility determinants have already been defined, including both structural and regulatory polymorphisms of erythyrocytes and of the immune system, and it is likely that many more have yet to be discovered. A vast number of DNA polymorphisms, scattered throughout the human genome, cause individual variation in probably all immunological and biochemical processes. Advances in DNA technology offer the prospect of screening thousands of candidate genes for association with susceptibility to severe malaria in large multicentre case-control and family-based studies. Saturation mapping of candidate gene regions, combined with cellular and molecular analysis of disease-associated polymorphisms, is essential for understanding the functional basis of the genetic associations that such an exercise will generate. This information will pinpoint critical molecular pathways in immunity and pathogenesis and may lead to fundamentally new strategies for treatment and prevention of severe malaria. PMID- 10697862 TI - Malaria transmission and morbidity. AB - Stable malaria endemicity is maintained over a wide range of transmission intensities in sub-Saharan Africa. This paper considers variations in the clinical manifestations and their consequences with differences in transmission intensity. Epidemiological approaches to malarial disease have concentrated on two clinical syndromes, severe malarial anaemia and cerebral malaria. Within an area the mean age of children with severe malarial anaemia is always lower than that of those with cerebral malaria. In areas of higher malaria transmission children, on average, encounter malaria at a younger age and the mean age of clinical cases is lower. Malarial anaemia tends therefore to be relatively more important under high transmission settings and cerebral malaria tends to gain in importance under lower transmission settings. In a number of studies the total load of malaria morbidity, whether measured as none severe malaria in the community or as severe malaria admitted to hospital, is low under stable low transmission conditions but is at its highest under moderate intensities of transmission. Thereafter it reaches a plateau, or even falls, at the highest transmission intensities. It is not known whether the same is true for mortality in communities living under different transmission settings. Possible implications for changes in patterns of morbidity and mortality following interventions which lower malaria transmission are discussed. It is concluded that such interventions should play an important role in integrated malaria control programmes but that these should involve concomitant introduction of other interventions, in order to minimise the possible risks of a reduced effect as the immune response of the population re-equilibrates in the face of reduced challenge. PMID- 10697863 TI - Consequences of multiple infection with Plasmodium falciparum in an area of high endemicity. AB - Most Plasmodium falciparum infections occur in partially immune hosts in highly endemic areas. In such situations, many hosts are simultaneously infected with multiple parasite genotypes, which must lead to intense competition between different parasite populations. We here summarise a series of studies of multiple infection, mostly using polymerase chain reaction-restriction fragment polymorphism (PCR-RFLP) genotyping of the highly polymorphic msp-2 gene. These indicate that chronic infections, characteristic of the partially immune host, appear to protect against super-infecting parasites. This protection is not seen in infants. A consequence is that selection for fast-growing (virulent) parasites, occurs mainly in the youngest, immunologically naive, hosts. The normal situation for P. falciparum is one in which the host is partially immune, and competition between parasite genotypes in this situation is not expected to result in selection for virulence. PMID- 10697864 TI - Severe malaria in Burkina Faso: urban and rural environment. AB - The age distribution and the clinical patterns of severe malaria (SM) were compared in patients from urban areas characterized by relatively low transmission, and from rural areas where the mean inoculation rates are at least twenty fold higher. The mean age of the urban and rural patients was 4.8 +/- 3.0 and 2.2 +/- 1.9 respectively (p < 0.000). The prevalence of coma was higher in the urban subsample (53.6 vs 28.9%, p << 0.000) while that of severe anemia (hemoglobin < 5 g/dl) was higher in rural patients (47.4 vs 14.8%, p < 0.000). Our data, in line with previous results obtained comparing rural areas characterized by different inoculation rates, show that the epidemiological context influences the clinical presentation of SM. PMID- 10697865 TI - Plasmodium falciparum clinical malaria: lessons from longitudinal studies in Senegal. AB - Development of new antimalaria strategies and particularly vaccines, needs an in depth understanding of the relationships between transmission, infection, immunity, morbidity and mortality. The intensive and longitudinal collection of entomological, parasitological and clinical data from the Senegalese populations of Dielmo (250-300 inhabitants), exposed to a perennial and intense transmission (about 200 infective bites/person/year) and of Ndiop (300-350 inhabitants) exposed to a seasonal transmission (about 20 infective bites/person/year), allows to respond to many questions about this subject. The acquisition of an antimalaria immunity as one gets older appears to reduce parasite density, complexity of infection, risk of new patent infection after a suppressive treatment but does not reduce the prevalence (as assessed by PCR) of infection which is commonly chronic and asymptomatic. The existence of a pyrogenic threshold effect of parasitaemia allows the individual diagnosis of malaria attacks. P. falciparum genotyping suggests that successive malaria attacks are due to distinct recently inoculated parasite populations that multiply initially without restriction, a dominant population is generally responsible of the clinical manifestations and all new populations do not trigger systematically attacks. The initial intensity of clinical manifestations does not differ perceptibly among children and adults, is not related to the duration of the attacks, does not allow the distinction between several types of attacks, is not predictive of their severity, and the clearance of parasites and manifestations is longer among youngest persons. The risk of malaria attacks is lower as one gets older and among carriers of AS haemoglobin, is higher when transmission increases and during pregnancy up to three months after delivery, and vary between children. The risk of malaria attack per infective bite is negatively related to the intensity of transmission. Because of their high sensitivity in malaria case detection, this type of small community-based studies are powerful and useful for the identification of protective immunological mechanisms as well as for testing rapidly and cheaply the clinical efficacy of any intervention such as antimalarial vaccines and drug therapy or prophylaxis. As a lot of vaccine candidates and drug combinations will be screened or tested in the perspective of the 'Roll-Back Malaria' programme, more attention must be given to longitudinal studies of this type. PMID- 10697866 TI - Malaria in migrants. AB - An increasing proportion of malaria cases in Italy is observed in immigrants revisiting their country of origin, but little specific research work has been carried out in this field. All malaria cases occurring from 1990 to 1998 at the Reference Clinic for Infectious and Tropical Diseases in Brescia were prospectically evaluated to compare clinical outcome in migrant and non-immune cases. No difference was observed between parasitaemia at diagnosis and time to clearance of peripheral parasitaemia. Clinical presentation was milder in migrants than in non-immunes, with an OR for severe malaria of 0.27 (c.i. = 0.09 0.84) (p = 0.01). Fever clearance time was significantly shorter in migrants (3.0 days, SD = 1.2) than in non-immunes (4.3 days, SD = 1.7) (p < 0.001). Among immigrants, the proportion of severe cases was higher in residents since 2 years or less (12.5%) compared to residents since 2 to 5 years (3.3%) and residents since more than 5 years (0.9%) (p = 0.02). The proportion of malaria cases who had used chemoprophylaxis was significantly lower among immigrants (30/272, 11.0%) compared to non-immunes (41/74, 55.4%) (p < 0.001). In a population based malaria KAP analysis among 504 migrants from malaria endemic countries, correct knowledge of malaria risk was reported by 351 (69.5%). Of 170 subjects who reported at least one visit back to the home country, 30 (17.6%) had sought pre travel advice, 24 (14.1%) had started chemoprophylaxis and 7 (4.1%) had completed it during the last visit. Of 140 migrants who failed to seek pre-travel advice, 73 (52%) were unaware of malaria risk, 56 (40%) did not know how to protect themselves, and 11 (8%) refused to use protective measures. Migrants account for a significant proportion of imported malaria cases in industrialised countries. Clinical presentation is milder compared to non-immune subjects. The proportion of migrants who adopt malaria protective measure while returning home is very low, due to both unawareness of risk and inappropriateness of medical advice. PMID- 10697867 TI - The complexity of the malaria vectorial system in Africa. AB - The malaria vectorial system in Africa is very complex. Five very efficient vector species transmit malaria: Anopheles gambiae, An. arabiensis, An. funestus, and the sometimes overlooked An. nili and An. moucheti. This paper focuses on morphological, behavioural and genetic differences observed among populations within each vector species. It emphasises that future strategies for controlling vectors should take into account this heterogeneity. PMID- 10697868 TI - Diversity of malaria in rice growing areas of the Afrotropical region. AB - It is well known that 'in many instances the rice agrosystem perfectly fits the ecological requirements of pathogens or vectors' and in fact 'malaria, schistosomiasis and Japanese encephalitis are important vector-borne diseases associated with rice production in developing countries' (IRRI, 1987). In spite of these fears, rice cultivation has been on the increase in the African region in response to demographic and economic pressures. However, although rice fields provide suitable breeding places for Anopheles mosquitoes and rice cultivation leads to an increase in the biting rates, the species which are adapted to these sites are not the same in all parts of Africa. Several examples illustrate this phenomenon: An. funestus in the rice fields of Madagascar, An. pharoensis in saline water rice fields in the delta of the Senegal river, An. arabiensis in northern Cameroon and Burundi, An. gambiae Mopti form in the Kou Valley (Burkina Faso) and An. gambiae Savanna form in the rice fields of Kafine near Bouake (Cote d'Ivoire). The vectorial capacities of these species are not the same and malaria inoculation rates are not necessarily increased in the riceland agroecosystem. The consequences for malaria of introducing rice cultivation depend on the situation before its introduction: it could be worsened in unstable malaria areas but not in stable malaria areas. Therefore, sound epidemiological and entomological knowledge are needed before causing any environmental modifications for agricultural purposes and there should be regular monitoring to avoid any outbreak. PMID- 10697869 TI - The clay feet of the malaria giant and its African roots: hypotheses and inferences about origin, spread and control of Plasmodium falciparum. AB - Grassi's allegory of the fragile feet of clay of the malaria giant applies particularly to Plasmodium falciparum marginal populations in temperate climates such as those that spread within the last three thousand years in the Mediterranean area through their close association with non diapausing vectors of the Anopheles maculipennis complex. The winter survival of the vector and the successful completion of the sporogonic cycle depended on the availability of the house environment to the mosquito. The fragility of the parasite's cycle became especially evident with the crucial impact of indoor-sprayed residual insecticides resulting in very rapid malaria eradication. The malaria giant showed to possess much more solid feet in the Tropics where P. falciparum eventually reached an exceptionally stable endemicity in sub-Saharan Africa due to a vectorial system which produces inoculation rates far higher than the minimum necessary to saturate human populations. This very high transmissibility resulting from recent human-dependent speciation processes in Afrotropical Anopheles mosquitoes (namely the emergence in the Neolithic period of specifically anthropophilic taxa in the An. funestus and An. gambiae complexes) had probably a key influence on the origin of the modern P. falciparum from an ancestral, less pathogenic, taxon. It is hypothesised that under the prevalence of multiple inoculation during epidemic flashes, a fast growing, aggressive strain responsible for acute, short-lived infections was selected. This quickly replaced the ancestral taxon and spread all over the world taking advantage of previous Anopheles radiation and of the demographic expansion following the agricultural revolution. Dealing with the African 'roots' of the malaria giant means to face both the exceptional stability of the parasite cycle and the risk of disrupting the human natural response with unsustainable interventions. Most efforts should be concentrated in the support and improvement of the available anti-disease strategies whereas the development of effective anti-infection strategies should be mainly pursued through reasonably diversified research programmes and well monitored pilot interventions within a long-term perspective. The aim is to discover and test new anti-malarial tools which either alone or, more probably, integrated with the available measures, could allow the interruption of transmission. In most Afrotropical hyper- and holoendemic zones, once given full priority to strengthening disease control, P. falciparum eradication appears the only realistic anti-infection objective, unless it is demonstrated that progress can be achieved in these epidemiological zones through sustainable and cost-effective intermediate steps based on the reduction of infection prevalence. PMID- 10697870 TI - Management of severe malaria. AB - The case fatality of WHO-defined 'severe falciparum malaria' remains unacceptably high, at 10-20%. However, a gradual decline in case fatality in adults and children treated in hospitals may reflect use of improved regimens of antimalarial chemotherapy and increased awareness of important complications of the disease. The development of severe, perhaps inevitably-fatal, malaria might be prevented by early appropriate chemotherapy of uncomplicated disease. At the most peripheral levels of the health service, suppository formulations of artemisinin derivatives can be administered even to patients who are vomiting or prostrated. At dispensaries, clinics or hospitals, where intramuscular or intravenous administration of antimalarial drugs is possible, quinine and artemisinin derivatives are the treatments of choice. There is growing evidence of the safety and efficacy of the quinine loading dose and of the use of artemether and artesunate, based on large, randomised, controlled clinical studies. No safe and effective form of prophylactic ancillary treatment has yet emerged. Results of studies of antipyretics, anticonvulsants (phenobarbitone), anticytokine/anti-inflammatory agents (anti-TNF antibodies, pentoxifylline, dexamethasone), iron chelators and hyperimmune sera have been disappointing. Only blood transfusion and treatment of respiratory, circulatory and renal failure are of obvious benefit. New ideas are needed, based on what is known of the pathophysiology of severe disease. PMID- 10697871 TI - What can the residents of malaria endemic countries do to protect themselves against malaria? AB - The incidence of malaria may vary substantially between adjacent communities and within an individual community, even in areas of high malaria transmission. Analysis of the factors responsible for these local variations in the incidence of malaria may identify potential control measures. Factors shown to be associated with local protection against malaria in some situations include house position, house design, the use of insect repellents and mechanical barriers such as bednets and curtains. The efficacy of insecticide treated nets and curtains in preventing mortality and morbidity from malaria, at least in the short-term, has been demonstrated convincingly. However, other measures of personal protection have not been evaluated in large trials which have clinical malaria as their endpoint. Such trials are needed to see if new malaria control tools can be identified that will assist current international efforts to improve malaria control, especially in Africa. The millions of non-immune travellers who visit malaria endemic areas each year need to protect themselves against malaria and the ways in which they can do this most effectively have been studied extensively. However, less attention has been paid to the local population of malaria endemic areas. What steps can they adopt to provide personal protection against malaria and how effective are these measures? Clues to which measures might be effective can come from study of the reasons for local variations in the incidence of malaria. PMID- 10697872 TI - Delaying antimalarial drug resistance with combination chemotherapy. AB - Resistance to antimalarial drugs arises when spontaneously occurring mutants with gene mutations or amplifications which confer reduced drug susceptibility are selected, and are then transmitted. Simultaneous use of two or more antimalarials with different modes of action and which therefore do not share the same resistance mechanisms will reduce the chance of selection, because the chance of a resistant mutant surviving is the product of the parasite mutation rates for the individual drugs, multiplied by the number of parasites in an infection that are exposed to the drugs. The artemisinin derivatives are very active antimalarials, which produce large reductions in parasite biomass per asexual cycle, and reduce malaria transmissibility. To date no resistance to these drugs has been reported. These drugs therefore make particularly effective combination partners. This suggests that antimalarial drugs should not be used alone in treatment, but always in combination, as in the treatment of tuberculosis or HIV, and that the combination should include artemisinin or one of its derivatives. PMID- 10697873 TI - Redox metabolism in glucose-6-phosphate dehydrogenase deficient erythrocytes and its relation to antimalarial chemotherapy. AB - Experiments in glucose-6-phosphate dehydrogenase (G6PD) deficient erythrocytes parasitized by Plasmodium falciparum proved that depletion of glutathione increased fluxes of reactive oxygen species and was detrimental to the parasite at various sites and developmental stages. Chloroquine is also considered an inducer of oxidant damage due to its role in preventing heme polymerization. Recently it has been found that GSH prevents cellular damage by degrading the toxic heme. Consequently, we suggest that the use of combinations of chloroquine and depletors of GSH would be highly efficient for the chemotherapy of malaria. PMID- 10697874 TI - Studies on anti-folate antimalarials in east Africa. AB - Chlorproguanil-dapsone (CD) appears to be a promising anti-folate combination (Amukoye et al., 1997) to substitute for pyrimethamine-sulfadoxine (PS), which has a long half-life and against which there is resistance in several Plasmodium falciparum populations including the highly endemic lowland area near Muheza, Tanzania (Trigg et al., 1997). PMID- 10697875 TI - Insecticide resistance in malaria vectors: a new approach to an old subject. AB - The application of biochemical and molecular biological techniques to the study of insecticide resistance has revolutionized our understanding of the underlying genetic basis of resistance. Using the examples of glutathione S-transferase and esterase-based metabolic insecticide resistance, three different routes via which increased insecticide detoxication can be achieved are elucidated. An understanding of these molecular pathways opens up new avenues for manipulating mosquito populations to restore insecticide susceptibility to the vectors. PMID- 10697876 TI - Current distribution of a pyrethroid resistance gene (kdr) in Anopheles gambiae complex from west Africa and further evidence for reproductive isolation of the Mopti form. AB - In the field, the kdr mutation, involved in pyrethroid resistance, has been found widely distributed in the Savanna form of Anopheles gambiae s.s., but never in wild populations of the Mopti form or An. arabiensis, even in areas where both occur in sympatry with resistant Savanna populations. Under laboratory conditions, Mopti and Savanna forms were fully able to interbreed and the kdr mutation was transmissible from one form to the other. Both forms appeared to be exposed to pyrethroid selection pressure in the field. The absence of the kdr mutation in the Mopti form and the total lack of Mopti-Savanna heterozygotes in field populations provides further evidence of a pre-copulatory barrier to gene flow between these two forms. Molecular markers, including kdr, are powerful tools for studying population genetics and circulation of resistance genes, and should be used through an integrated approach for a better understanding of the speciation process. PMID- 10697877 TI - The kdr pyrethroid resistance gene in Anopheles gambiae: tests of non-pyrethroid insecticides and a new detection method for the gene. AB - The organophosphate pirimiphos-methyl and the carbamate carbosulfan were evaluated in comparison to the pyrethroid alphacypermethrin and the 'near pyrethroid' etofenprox against pyrethroid resistant Anopheles gambiae and Culex spp. in an experimental hut station located in central Cote d'Ivoire. Bednets were impregnated with the above mentioned compounds and randomly allocated to the huts. On 40 consecutive mornings, after sleepers had occupied the huts overnight, mosquitoes were collected from the huts, identified and scored as live or dead (including delayed mortality). An. gambiae s.l. that had been collected were tested for the presence of the kdr allele in heterozygous or homozygous form. Both non-pyrethroid treatments caused very high mortality, whereas mortality with alpha-cypermethrin and etofenprox generally did not differ from the levels observed with untreated control nets in this experiment. The nets had holes cut in them and there was considerable bloodfeeding on the sleepers, which was only significantly reduced for An. gambiae by carbosulfan and alpha-cypermethrin. PCR genotyping suggested that there was selection for the kdr resistance allele by the pyrethroid treated nets. Organophosphates and carbamates may therefore present an alternative to be used on bednets especially in areas of pyrethroid resistance, but the safety of these insecticides will have to be carefully considered. PMID- 10697878 TI - Re-emerging malaria in the WHO European region: control priorities and constraints. AB - Malaria has been identified by some of the countries in the European Region of the World Health Organization as a priority, due to the re-emergence of the problem. This paper aims to present the situation of indigenous malaria in the Region and the strategy to be adopted to roll back malaria. PMID- 10697879 TI - Malaria control priorities and constraints. AB - Each year, there are still between 300-500 million clinical cases of malaria and over one million deaths due to the disease, 90% of which occur in Africa south of the Sahara. In all continents, malaria risk is highest in remote rural areas where poverty abounds, population densities are low and the quality and coverage of the health services are poor or in existent. A sustained impact on the malaria burden can only be achieved through the cost-effective use of current tools, by including malaria in health sector development and inter-sectorial action, by mobilizing malaria control within communities and by investing in new and more effective tools. This paper highlights some of the constraints faced by countries in controlling malaria and outlines the priority activities that are being carried out to address these constraints within both communities and the health services. It aims to be set the scene for the papers of this Centenary book which address some of these issues in more detail. PMID- 10697880 TI - Opportunities, problems and perspectives for malaria control in sub-Saharan Africa. AB - Environments conducive to high malaria transmission and widespread poverty are at the roots of the 'malaria giant', which affects 46 countries in Africa. The recent interest in and momentum of work on malaria, in endemic countries and the international community, is unprecedented and opens new perspectives for controlling the disease. Significant steps included: (i) the allocation of US$20 million by WHO for accelerated implementation of malaria control in 34 African countries in 1997-98; (ii) the Declaration on Malaria by the Heads of States of the Organization of African Unity and the establishment of the African Initiative for Malaria Control in 1997; (iii) the concomitant mobilisation of the research community in the Multilateral Initiative on Malaria; (iv) the G8 Summit in 1998 in Birmingham asking for higher commitment to malaria control, particularly in Africa; and (v) the Roll Back Malaria initiative set as a WHO priority project in 1998. However, experiences have proved the alarming 'resilience' of the malaria system in Africa, showing devastating consequences when malaria returns to the original levels after intensive control is interrupted. Effective malaria control in Africa requires long-term action, firmly rooted in the social development of the country. PMID- 10697881 TI - Prevention and control of malaria epidemics. AB - Malaria epidemics have recently occurred in many areas of the world, particularly in the irregular fringe, along the limits of distribution of malaria endemicity, whether the limiting factors are temperature (latitude or altitude) or relative humidity (deserts), which were the scene of the major epidemics of the past. A review is made of the current approaches to epidemic prevention and control in line with the global malaria control strategy, of which it constitutes an essential element. The different components are discussed, including the identification and study of epidemic prone areas, the search for indicators of epidemic risk, their monitoring, and the early detection and control of actual epidemics. The potential implementation of preventive and control activities depends, nevertheless, on the degree of preparedness of the health services to detect alarm signals, the real time left for action and their capacity of implementation within that time. Recent developments in geographical information systems (GIS) and satellite derived meteorological information offer most useful tools for precise and timely epidemic forecasting, although it should be recognised that such information is only useful if based on a real understanding of their local significance. There remains an urgent need to develop and support local capabilities for the ground truthing of satellite information and for translating it into preventive and control actions. PMID- 10697882 TI - Current scenario of malaria in India. AB - The Indian National Malaria Eradication Programme (NMEP) is reporting 2.5 to 3 million malaria cases, and about 1,000 malaria deaths annually. Malaria in the northeastern states is stable and in the peninsular India unstable. There are six major and three minor malaria vectors, of which Anopheles culicifacies transmits malaria in rural areas and An. stephensi in the towns. Other vectors are of local importance. Plasmodium vivax is the dominant infection and accounts for 60-65% cases whereas P. falciparum contributes 30-35% cases. Field operations to control malaria are impeded by resistance and/or exophilic vector behavior, parasite resistance to antimalarial drugs, operational problems in spraying, failure to search breeding of mosquitoes at weekly intervals, staff shortages and financial constraints. Resurgent malaria invaded new ecotypes created by green revolution, industrial growth and urban development resulting in paradigm shift towards man made malaria. NMEP has launched a world bank-assisted enhanced malaria control project with primary emphasis to protect 62.2 million high risk population in 7 states. PMID- 10697883 TI - Overview of malaria control in the Americas. AB - The malaria endemic countries of the Americas have adopted in 1992 the WHO Global Malaria Control Strategy whose difficulties of implementation have been compounded to a major reform in the health sector, as the countries adjust to conform to financial scarcity and new economic policies. Most countries of the Region have reoriented its control program from eradication of malaria to the elimination of malaria mortality and morbidity. The Region has advanced towards these objectives having already reduced its mortality by 60% and is now in the final stages of applying new tools to control transmission and rapidly advance to reduce the incidence of malaria in the Americas. PMID- 10697884 TI - Italian Development Cooperation: the commitment for the struggle against malaria in Africa. AB - The Italian Development Cooperation (DGCS) support the health reform process in Developing Countries, with the aim to provide populations in greatest need with access to decentralized health services. DGCS acts in close coordination with the donor community, United Nations' system and the World Bank, in agreement with sector-wide approach (SWAP) for health sector development. Since malaria control in endemic countries is a relevant component of the health system, DGCS is actively involved in the struggle against malaria in sub-Saharan Africa, supporting control activities and research capability strengthening. The following African countries are presently receiving bilateral support for antimalaria activities: Burkina Faso, Centre de Lutte contre le Paludisme in Ouagadougou; Ethiopia, community-based malaria control in Tigray; Eritrea, malaria control at national level in the framework of the Public Health and Rehabilitation Programme for Eritrea (PHARPE) initiative; Madagascar, malaria surveillance at national level; Tanzania, feasibility study for the support to the national malaria control programme. Support is provided by technical/financial assistance involving Italian academic and research institutions. On the multilateral channel, DGCS has provided regular contribution for WHO's work in malaria control and participates to the WHO Roll Back Malaria initiative. A new commitment to malaria is the trilateral joint scientific endeavour USA-Italy-Burkina Faso for the development and field testing of a candidate vaccine suitable for African populations. PMID- 10697885 TI - Community-based malaria control in Tigray, northern Ethiopia. AB - Community-based control activities have been a major component of the Tigray regional malaria control programme since 1992. A team of 735 volunteer community health workers treat on average 60,000 clinical malaria cases monthly during the high malaria transmission season. Ensuring access for the rural population to early diagnosis and treatment has contributed to a significant decrease in death rate in under-five children at the village level from 1994 to 1996. Mapping and geographic information system (GIS) technologies have been introduced to support planning for control by assessment of community-based coverage. With further development, GIS will be used in stratification, and to assess the impact of water resources development on malaria transmission and intensity. PMID- 10697886 TI - Control of epidemic malaria on the highlands of Madagascar. AB - The Malagashy national malaria control programme ('Programme National de Lutte contre le Paludisme', PNLP) has been developing, since 1996, an epidemiological early warning system for malaria epidemics in the Central Highlands with the support of the Italian Development Cooperation. The system is based on the monitoring of malaria morbidity (clinical diagnosis) in 536 peripheral health centres (CSB) of the Highlands. The intervention area corresponds to 27 districts of the Antananarivo and Fianarantsoa provinces (4.7 million inhabitants) and spans around 100,000 square km. For each CSB a monthly warning threshold, defined as the 1993-1996 monthly mean number of malaria cases plus two standard deviations, was established. Three levels of epidemic alert have been defined according to the number of times the cases of presumptive malaria surpassed the threshold and according to the reported presence of severe malaria cases. The surveillance system relies also on the monitoring, in district hospitals of the Highlands, of the Plasmodium falciparum infection rate among clinically diagnosed malaria cases. A total of 185,589 presumptive malaria cases, corresponding to a 42/1000 malaria incidence, were recorded in 1997 by the surveillance system. During the same year 184 alerts of 2nd degree were reported. During 1998 173,632 presumptive malaria cases corresponding to a 38/1000 incidence were reported and 207 alerts of 2nd degree were detected; 75 of these alerts were investigated with ad hoc surveys and 3 initial malaria epidemics identified and controlled. Out of 6884 presumptive malaria cases diagnosed in the district hospitals during 1997 1998, only 835 (12.1%) have been confirmed by microscopy (P. falciparum 81.7%, P. vivax 15.0%, P. malariae 2.5%, P. ovale 0.2%, mixed infections 0.6%); 22.4% of these infections were imported cases from coastal endemic areas. The efficiency of the system in monitoring the trend of malaria morbidity and in the rapid detection and response to malaria epidemics is still being evaluated. PMID- 10697888 TI - Chemotherapeutic malaria control as a selective primary health care activity in the Solomon Islands. AB - Malaria control by chemotherapy has been established in rural villages of Guadalcanal, the Solomon Islands, following field trials. As a selective primary health care activity, mobile unit teams visited villages once or twice a year to detect malaria positives and gave chloroquine and primaquine to treat the infection and interrupt the transmission. On site diagnosis was by the use of acridine orange fluorescent staining or the ICTPf commercial diagnostic kit. To avoid possible haemolytic crises, a new single step screening method of G6PD deficiency was introduced. This approach has been accepted well by villagers and proved to be an efficient and feasible control method even in remote rural villages with endemic malaria transmission. Epidemiological modelling of the situation predicts reduction of prevalence in five years. PMID- 10697887 TI - Widespread distribution of insecticide-impregnated curtains reduces child mortality, prevalence and intensity of malaria infection, and malaria transmission in rural Burkina Faso. AB - The results of the first two years of implementation of a large scale trial of insecticide-treated curtains in Burkina Faso are summarised in this presentation. The trial was conducted in a highly malarious area and involved a population of slightly less than 100,000, distributed in 158 villages over an area of almost 1000 km2. A remarkable impact on entomological parameters (Anopheles density, sporozoite rate, entomological inoculation rate) was accompanied by a relatively modest reduction of parasitological indices (prevalence and density of Plasmodium falciparum). All-cause mortality in children 0.5 to 5 year old showed over two years a 15% decline. The authors believe that the wide surface of the protected zone and the almost total coverage achieved in the intervention villages were the major determinants of the observed reduction of transmission. A conclusive interpretation of the mortality results requires a further follow-up of the study population. PMID- 10697889 TI - Malaria prevention by vector control: effectiveness of insecticidal methods. AB - House-spraying with residual insecticides continues to be the most popular and cost-effective means of malaria prevention and control. It remains to be seen whether insecticide treated nets will prove to be more economical and sustainable. PMID- 10697890 TI - Can the resurgence of malaria be partially attributed to structural adjustment programmes? AB - Structural Adjustment Programmes (SAPs) (restrictive fiscal and monetary policies and extensive public sector reforms) were implemented in many developing countries during the 1980s. Their socio-economic impact on the poor has been widely discussed. Antimalarial efforts could have been affected by this economic reform process, and the resurgence of malaria during the past twenty years could be partially attributed to the socio-economic hardship generated by these programmes. PMID- 10697891 TI - Implementation of malaria control. AB - Global trends of infant and child mortality have decreased over the last 30 years, while the proportion of malaria deaths has progressively increased due to the deteriorating situation in sub-Saharan Africa. The Global Malaria Control Strategy promoted by WHO has encountered several obstacles to its implementation. Early diagnosis and prompt treatment can reduce malaria mortality, but there is still low investment on safe and effective modalities of care delivery at the periphery, where most of the malaria burden exists. Selective vector control (indoor residual spraying and insecticide-treated nets) plays a significant role outside Africa, but its wider use is limited by cost/affordability problems and operational issues (supply, delivery and logistics). Alternative methods such as environmental management and biological control are cost-effective only under very specific epidemiological situations. In most countries forecasting, early detection and containment of malaria epidemics is deficient, and there is separation between the research and control communities, particularly in Africa. Involvement of the internal agencies, strategic investments in capacity building and institutional networking are needed to strengthen capacity for malaria and research in the countries. The major responsibility is to guide the expenditure made by the communities (which far out-weigh the limited share of national health budgets) towards the most cost-effective approaches to reduce malaria mortality and morbidity. PMID- 10697892 TI - Pre-erythrocytic malaria vaccine: mechanisms of protective immunity and human vaccine trials. AB - In order to provide a rational basis for the development of a pre-erythrocytic malaria vaccine we have aimed at: (a) elucidating the mechanisms of protection, and (b) identifying vaccine formulations that best elicit protection in experimental animals and humans. Based on earlier successful immunization of experimental animals with irradiated sporozoites, human volunteers were exposed to the bites of large numbers of Plasmodium falciparum or P. vivax infected irradiated mosquitoes. The result of this vaccine trial demonstrated for the first time that a pre-erythrocytic vaccine, administered to humans, can result in their complete resistance to malaria infection. However, since infected irradiated mosquitoes are unavailable for large scale vaccination, the alternative is to develop subunit vaccines. The human trials using irradiated sporozoites provided valuable information on the human immune responses to pre erythrocytic stages and studies on mice an excellent experimental model to characterize protective immune mechanisms. The circumsporozoite protein, the first pre-erythrocytic antigen identified, is present in all malaria species, displaying a similar structure, with a central region of repeats, and two conserved regions, essential for parasite development. Most pre-erythrocytic vaccine candidates are based on the CS protein, expressed in various cell lines, microorganisms, and recently the corresponding DNA. We and others have identified CS-specific B and T cell epitopes, recognized by the rodent and human immune systems, and used them for the development of synthetic vaccines. We used synthetic peptide vaccines, multiple antigen peptides and polyoximes, for immunization, first in experimental animals, and recently in two human safety and immunogenicity trials. We also report here on our work on T cell mediated immunity, particularly the protection of mice immunized with viral vectors expressing CS-specific cytotoxic CD8+ T cell epitopes, and the striking booster effect of recombinant vaccinia virus. To what degree CD8+ T cells, and/or other T cells specific for sporozoites and/or liver stage epitopes, contribute to pre erythrocytic protective immunity in humans, remains to be determined. PMID- 10697893 TI - Malaria vaccines: triumphs or tribulations? AB - A safe and effective malaria vaccine will greatly facilitate efforts to control the global spread of malaria. This paper discusses the conceptual framework for developing malaria vaccines and some of the difficulties that the various approaches face. It emphasizes the role of pre-erythrocytic malaria vaccines, which are designed to protect against malaria infection, rather than simply prevent clinical disease. It describes recent encouraging results in human subjects with the RTS,S vaccine, a promising pre-erythrocytic malaria vaccine candidate. PMID- 10697894 TI - Merozoite surface protein 1, immune evasion, and vaccines against asexual blood stage malaria. AB - There is an urgent need for a vaccine against malaria and proteins on the surface of the merozoite are good targets for development as vaccine candidates because they are exposed to antibody. However, it is possible that the parasite has evolved mechanisms to evade a protective immune response to these proteins. Merozoite surface protein 1 (MSP-1) is a candidate for vaccine development and its C-terminal sequence is the target of protective antibody. MSP-1 is cleaved by proteases in two processing steps, the second step releases the bulk of the protein from the surface and goes to completion during successful red blood cell invasion. Antibodies binding to the C-terminus of Plasmodium falciparum MSP-1 can inhibit both the processing and erythrocyte invasion. Other antibodies that bind to either the C-terminal sequence or elsewhere in the molecule are 'blocking' antibodies, which on binding prevent the binding of the inhibitory antibodies. Blocking antibodies are a mechanism of immune evasion, which may be based on antigenic conservation rather than diversity. This mechanism has a number of implications for the study of protective immunity and the development of malaria vaccines, emphasising the need for appropriate functional assays and careful design of the antigen. PMID- 10697895 TI - Epidemiological considerations for malaria reduction by transmission blocking vaccination. AB - Outside of the temperate regions, malaria transmission continues throughout much of the world in a distribution which is not very different to that of one hundred years ago. However, with the notable exception of Africa sub Sahara, the morbidity and mortality due to malaria has generally been reduced to very low levels by comparison with earlier times. In a broad sense the malaria problem today falls into two distinct compartments, 1) how to deal with the remaining problem of malaria in the affected areas outside of sub Saharan Africa and 2) how to manage the, currently, much greater problem of malaria-related morbidity and mortality in Africa sub Sahara. Malaria control campaigns of the past have always placed great emphasis on reducing malaria inoculation rates in the affected populations. This may seem entirely logical, and is, indeed, an absolute requirement where eradication of malaria from an endemic area is the goal. There can, nevertheless, be dangers as well as benefits associated with reducing malaria inoculation rates in previously endemic populations. I discuss here the epidemiological issues which should be taken into account in this respect. I then examine the role that vaccination to reduce malaria inoculation rates in endemic populations--malaria transmission blocking vaccination--could play in malaria control. PMID- 10697896 TI - Plasmodium falciparum CS C-terminal fragment: preclinical evaluation and phase I clinical studies. AB - Preclinical evaluation of synthetic peptides corresponding to the C-terminal regions of the circumsporozoite (CS) protein in various Plasmodia showed that these preparations were immunogenic and safe upon injection in various animal models. Additionally, the corresponding peptide from Plasmodium falciparum was widely recognized by sera and PBL obtained from semi-immune adults living in malaria endemic areas. Moreover, the CS C-terminal peptide derived from P. berghei conferred protection upon challenge with live sporozoites in mice. A GLP preparation of the synthetic peptide corresponding to residues 282-383 of the Pf CS, NF-54 strain is currently evaluated in a open, non-randomized, Phase I human trial. Data obtained after the second antigen injection show that the malaria vaccine Pf CS 282-383 is safe, well tolerated and gives rise to high antibody titre, CD4+ and CD8+ lymphocyte responses. PMID- 10697897 TI - Thrombospondin related adhesive protein (TRAP), a potential malaria vaccine candidate. AB - We have investigated whether naturally induced immunity to Plasmodium falciparum thrombospondin related adhesive protein contributes to protection against malaria in humans. We have carried out a case control study in children living in an endemic region of West Africa to reveal associations between PfTRAP seroprevalence and the risk of cerebral malaria. Sera collected from the case and control groups were analysed by ELISA to compare their serum reactivity against PfTRAP, the circumsporozoite protein and the merozoite surface protein 1. Children with uncomplicated malaria had a significantly higher PfTRAP seroprevalence when compared to children with cerebral malaria. The risk of developing cerebral malaria appeared to depend on the reciprocal relationship between sporozoite inoculation rates and humoral immunity against PfTRAP. Our results suggest that naturally induced humoral immunity against PfTRAP contributes to the development of protection against severe malaria. Experimentally induced immunity against TRAP in different rodent models has consistently proven to elicit a high degree of protection against malaria. This together with the functional properties of TRAP and data describing CD4 and CD8 epitopes for PfTRAP indicate that this molecule could increase the protective efficiency of available sporozoite malaria vaccines. PMID- 10697898 TI - Progress in the development of malaria vaccines: context and constraints. AB - Major technical advances in the field of vaccine development have culminated in an impressive array of prototype vaccines that may well provide 'proof of principle' that vaccines against all life-cycle stages may induce a degree of protection against malaria. As the mechanisms responsible for protection against this disease are not known, and vaccines for populations at greatest risk will be applied in the presence of ongoing infection and a degree of concomitant immunity, it is essential for us to learn from the 'experiments of nature' about acquired and ongoing immunity in order to determine when and how these vaccines may be applied. Successful interventions with chemoprophylaxis or vector control have provided obvious lessons and highlight the importance of recognising the lack of correlation between infection, clinical disease and mortality. Vaccines inducing sterile immunity raise concerns about rebound mortality in populations who will undoubtedly be re-challenged later in life, hence the need to review supplementary or alternative strategies for reducing disease through immune responses to toxins or molecules inducing pathology by adherence to host endothelium. Following antigen selection there are many challenges in choosing methods of antigen delivery and adjuvants, and measuring vaccine efficacy. A successful vaccine would need to be delivered through a national programme in the context of implementation of a wide range of components required for an effective control strategy. PMID- 10697899 TI - Vaccine efficacy, and immunity affecting transmission. AB - New candidate malaria vaccines being developed experimentally can only be assessed properly after they have been through the full range of clinical trials and extended follow-up. Defining vaccine efficacy, especially that of multi component vaccines, is difficult, in particular because the outcome is determined by finely defined epitope specificity. Immune responses can have contrasting effects, some protective, some promoting infection, but concerns that interventions that reduce transmission from a high to a lower intensity will lead to greater risks of severe malaria may not have taken into account all of the many factors that determine the outcome of infection. PMID- 10697901 TI - New tools for malaria therapy. AB - In the absence of vaccines, new chemotherapies are needed urgently to help in the prevention and control of malaria. A number of strategies are being followed world-wide in attempts to discover and develop them: (a) resurrecting 'forgotten' molecules; (b) developing new formulations of existing products; (c) finding ways to bypass known toxicological limitations of existing products; (d) looking for combinations of existing products; (e) discovering molecules which reverse the resistance phenotype; (f) identifying 'old' chemical entities (OCEs) for new, antimalarial, indications; (g) discovering new chemical entities (NCEs) directed towards already exploited biological targets; (h) discovering NCEs directed to novel biological targets. Examples of such strategies are given below, together with an indication of their advantages and limitations. PMID- 10697900 TI - Forecasting and prevention of epidemic malaria: new perspectives on an old problem. AB - There is a clear need for improved epidemic malaria surveillance mechanisms in areas prone to the disease. Epidemiological surveillance systems are rarely able to provide information in a sufficiently timely manner for adequate epidemic response. This is especially true in African countries where surveillance is poorly developed, and particularly so in remote regions of unstable malaria such as desert fringes. There is long standing evidence linking climatic variability and epidemic risk. The last ten years have seen significant developments in Environmental Information System (EIS) for a range of natural resource management purposes. The routine information products from these systems have been shown to be both spatially and temporally related to malaria transmission indicators across the African continent. EIS may therefore provide a useful and cost effective input to epidemic malaria control planning and response. PMID- 10697902 TI - The development of and perspectives for genetic engineering of malaria parasites. AB - The genetic manipulation of malaria parasites is a rapidly emerging technology that offers great promise for the investigation of many aspects of infection. Currently it is possible to transform avian, rodent, primate as well as human parasites, the latter three on a stable, drug selectable basis. This review focuses on the history of the development of the technology, current abilities and future perspectives. PMID- 10697903 TI - Controlling malaria transmission with genetically-engineered, Plasmodium resistant mosquitoes: milestones in a model system. AB - We are developing transgenic mosquitoes resistant to malaria parasites to test the hypothesis that genetically-engineered mosquitoes can be used to block the transmission of the parasites. We are developing and testing many of the necessary methodologies with the avian malaria parasite, Plasmodium gallinaceum, and its laboratory vector, Aedes aegypti, in anticipation of engaging the technical challenges presented by the malaria parasite, P. falciparum, and its major African vector, Anopheles gambiae. Transformation technology will be used to insert into the mosquito a synthetic gene for resistance to P. gallinaceum. The resistance gene will consist of a promoter of a mosquito gene controlling the expression of an effector protein that interferes with parasite development and/or infectivity. Mosquito genes whose promoter sequences are capable of sex- and tissue-specific expression of exogenous coding sequences have been identified, and stable transformation of the mosquito has been developed. We now are developing the expressed effector portion of the synthetic gene that will interfere with the transmission of the parasites. Mouse monoclonal antibodies that recognize the circumsporozoite protein of P. gallinaceum block sporozoite invasion of mosquito salivary glands, as well as abrogate the infectivity of sporozoites to a vertebrate host, the chicken, Gallus gallus, and block sporozoite invasion and development in susceptible cell lines in vitro. Using the genes encoding these antibodies, we propose to clone and express single-chain antibody constructs (scFv) that will serve as the effector portion of the gene that interferes with transmission of P. gallinaceum sporozoites. PMID- 10697904 TI - Model systems to evaluate the use of transgenic haematophagous insects to deliver protective vaccines. AB - Insect vector control has proved an effective method for reducing the transmission of disease-causing organisms to human populations in many tropical countries. We are interested in employing direct genetic manipulation of insect vector genomes to use them in beneficial ways so as to have a profound and long lasting effect on disease transmission. Our research focuses on assessing whether haematophagous insects may be used as a means to deliver protective proteins, such as an antimalarial vaccine, when they take a blood meal. The progress which has been made towards assessing this concept using a number of model systems is described. PMID- 10697905 TI - Biological problems with the replacement of a vector population by Plasmodium refractory mosquitoes. AB - Attempts are being made to backcross into Anopheles gambiae s.s. the gene(s) which cause zoophily in Anopheles quadriannulatus. Such a backcrossed strain might be preferable to a Plasmodium-refractory strain as a basis for genetic control because a refractory strain could select for evasion of refractoriness in the wild Plasmodium population. The species composition of the malaria vector population in several Tanzanian villages was overwhelmingly An. gambiae s.s. in a normal rainy season, but consisted of four species, all proved by ELISA and/or PCR to carry P. falciparum sporozoites, at the time of the heavy rains associated with El Nino. Thus any scheme, for malaria transmission control by replacement of vectors by genetically-manipulated non-vectors, would have to be able to replace more than one species. PMID- 10697906 TI - Salivary gland-specific gene expression in the malaria vector Anopheles gambiae. AB - Molecular studies on the tissue-specific gene expression in the salivary glands of Anopheles gambiae may provide useful tools for the development of new strategies for the control of the most efficient malaria vector in the sub Saharan Africa. We summarize here the results of a recent investigation focused on the isolation of secreted factors and putative receptors from the salivary glands of An. gambiae. Using the Signal Sequence Trap technique we have identified the first cDNAs specifically expressed in the An. gambiae salivary glands. Among these, four are exclusively expressed in female glands and encode factors presumably involved in blood-feeding, whereas two other cDNAs seem to be expressed both in male and in female glands and are likely implicated in sugar feeding. Homologues of genes previously identified in the yellow fever mosquito Aedes aegypti, like the apyrase and D7, as well as novel salivary gland-specific cDNAs, were identified. The isolation and characterization of promoter sequences from the corresponding genes may prove useful for the expression of anti parasitic agents in the salivary glands of transgenic mosquitoes. PMID- 10697907 TI - Malaria entomology: can genomics help? AB - The field of genomics has advanced over the last decades to the forefront of molecular genetic research. Many genomes, including that of yeast, have already been sequenced and the determination of the genome sequence of several higher organisms is now within sight, including the complete DNA sequence of Homo sapiens. Here I review the state of the Anopheles gambiae genomic research and I present the current plans for an 'Anopheles Genome Project'. An understanding of the structure and function of the vector's genome may ultimately provide better tools for the control of the malaria mosquito. PMID- 10697908 TI - Malaria control priorities and constraints. AB - The capacity to prioritize correctly actions in malaria control depends on good knowledge not only of the epidemiology of the disease in the area but also of the behaviour of the people. Health policy makers frequently believe that the people already know enough about malaria and there is no need to commit further resources on finding out what the people actually know and do about the disease in order to modify their wrong habits. One of the pressing priorities for malaria control in Africa is therefore changing the attitude of malaria control policy makers. Considering the constraints to malaria control it is stressed that the health budget is usually below a level sufficient to finance an effective health care system. This is further compounded by inequities in the allocation of funds between health care institutions located in the urban areas compared with those located in the rural areas. Another important constraint is lack of manpower suitably trained to undertake the various elements of the global malaria control strategy. Finally, a very well known constraint is the unavailability of effective drugs at the locations where they are needed. PMID- 10697909 TI - The Multilateral Initiative on Malaria: co-ordination and co-operation in international malaria research. AB - The Multilateral Initiative on Malaria (MIM) is an international alliance of organisations and individuals. It aims to maximise the impact of scientific research against malaria, through strengthening research capacity in Africa, promoting global collaboration and co-ordination, and increasing available resources. Since its establishment in 1997, the initiative has generated a remarkable level of enthusiasm and activity. Many new scientific partnerships have been established, enabled by enhanced communications and novel funding mechanisms. Dovetailing of research activities with control programmes is also improving. The challenges posed by malaria remain great, however, and in order to achieve a sustainable impact it will be crucial for the research community to capitalise on what has been achieved to date and to maintain the momentum for action well into the next millennium. This article is a personal view contributed by the Wellcome Trust as the nominated co-ordinator for MIM during 1998 and a leading international funder of malaria research. It aims to explain how the novel malaria initiative operates, to summarise some of its key outcomes, and to set out the perspectives for the future. PMID- 10697910 TI - Roll Back Malaria. AB - Roll Back Malaria is an initiative intended to halve the suffering caused by this disease by 2010. The initiative is being developed as a social movement. Action is directed by national authorities backed by a global partnership which consists of development agencies, banks, private sector groups and researchers. The World Health Organization, the World Bank, UNICEF and UNDP founded the partnership in October 1998. The WHO has established a new Cabinet Project, and a WHO-wide strategy and workplan, to support the partnership. High quality, practical, consistent and relevant technical advice is made available through networks of experts based in research, academic, and disease control institutions, particularly those in endemic countries. The initiative also supports research and development of new products and tools to control malaria. Implementation of Roll Back Malaria began with a series of in-country consultations in 1998, followed by sub-regional consensus building and inception meetings. The current period is one of momentum building at country level during which national authorities are developing their own strategies with the global partners. It is anticipated that, during the year 2000, RBM movements will become active in at least 30 countries. PMID- 10697911 TI - Malaria research: back to the future. AB - One hundred years ago, Ross and Grassi provided us with the biological basis and fifty years ago, the human right declaration highlighted the moral responsibility of preventing death and reducing morbidity due to malaria. Yet one million children still die every year from the disease. The diversity in the parasite host interaction surely represents a major obstacle, but modern scientific technology offers possibilities, at least partly or temporarily, to overcome the inherent ability of the parasite to evade chemotherapy and the immune system. True partnerships, North-South and others, represent the only way to respond to the scientific challenges. A strong commitment of different sectors in the global community to scientific achievements is necessary also with regards to malaria, a disease of the poor. PMID- 10697912 TI - The malaria challenge in the 21st century: perspectives for Africa. AB - Malaria control has had little success in Africa despite the achievements in malaria research. It is time to put more emphasis on sustainable control measures through local commitment to diagnose and treat malaria in order to prevent illness and death. This goal can be best achieved through basic health care centers, schools and safe water supplies to rural areas. Complementary actions through research and international support will be strongly needed. PMID- 10697913 TI - The need to 'maintain the rage'. AB - Personal reflections are presented on the need to ensure that appropriate attention is given to malaria. The importance of political and financial commitments is stressed and the investment in people is considered a major priority. PMID- 10697914 TI - Hepatitis B virus infection among pregnant women delivering at Harare Maternity Hospital, Harare Zimbabwe, 1996 to 1997. AB - OBJECTIVE: To determine the prevalence of hepatitis B virus (HBV) carrier and infectivity status among pregnant women delivering at Harare Maternity Hospital. DESIGN: A serological survey study of pregnant women admitted for labour and delivery. SETTING: Harare Maternity Hospital, Harare, Zimbabwe between June 1996 and June 1997. SUBJECTS: A random sample of 1,000 women, delivering at the hospital during the study period agreed to participate in the study. Serum samples were available for 984 women. MAIN OUTCOME MEASURES: HBV carriage status was determined by the presence of hepatitis B surface antigen (HBsAg) by enzyme immunoassay (EIA). Maternal HBV infectivity status was determined by testing all HBsAg positive women for the presence of hepatitis e surface antigen (HBeAg) using EIA. RESULTS: Overall 246 (25%) women were identified as carriers of HBV (95% confidence interval 22 to 28%). The frequency of HBV carriers did not vary with maternal age, parity or marital status. Only a positive prior history of spontaneous abortion was associated with an increased prevalence of HBV carriage status. Eight of the 246 (3.3%) women identified as HBV carriers tested positive for HBeAg. Hence, 0.8% of the entire study population was found to be at high risk of transmitting HBV to their newborns. CONCLUSIONS: Our results demonstrate a high prevalence of HBV carriage among women giving birth at Harare Maternity Hospital. None of the demographic variables studied were important predictors of HBV carriage status. The high carriage rate and low infectivity rates suggest that HBV infection is likely to be acquired by horizontal, rather than by vertical means of transmission. Given the scarcity of financial resources, routine testing of mothers for HBsAg may not be feasible. Our results suggest, however, that mass vaccination of all infants, irrespective of maternal HBV carriage status, may be the most effective approach to HBV prevention and control in Zimbabwe. PMID- 10697915 TI - A comparison of nutritional indices of children in Chitungwiza, Zimbabwe, with the international reference standard. AB - OBJECTIVES: To examine the hypothesis that, the fifth, 50th and 95th percentiles of the weights and heights of primary school children of Chitungwiza Municipality, (a town 30 km south west of Harare, Zimbabwe), did not differ from those of the NCHS reference population of children. DESIGN: A descriptive cross sectional study. SETTING: Chitungwiza Municipality. SUBJECTS: Primary school children aged five to 16 years. MAIN OUTCOME MEASURES: Height for age < 90%, weight for height < 80% and the comparability of mean weights and heights between the study children and the NCHS reference children. RESULTS: Low rates were found for height for age < 90% (stunting) and for weight for height < 80% (wasting) among the Chitungwiza children, 3.5% (95% CI 2.8%, 4.7%) and 1.9% (CI 0.9%, 3%), respectively. The differences between age and sex matched pairs of the sample mean heights and reference mean heights, and of the sample mean weights and reference mean weights at the fifth, 50th and 95th percentiles, were significant. Chitungwiza children consistently dropped below the NCHS mean weight and height for all three percentiles. CONCLUSION: This study has demonstrated that stunting and wasting is low among Chitungwiza primary school children but that the spread of their heights and weights lies lower than the spread of the heights and weights of the NCHS reference children. We recommend that wider cross sectional and longitudinal anthropometric assessments in a nation wide sample of primary school children be carried out to shed more light on the growth potential of Zimbabwean children. PMID- 10697916 TI - Knowledge and practices of family planning in Zimbabwe. AB - OBJECTIVE: To assess the level of knowledge and use of family planning in Zimbabwe. DESIGN: Cross sectional study. SETTING: All eight provinces and two major cities in Zimbabwe. SUBJECTS: Women of child bearing age (15 to 49 years, 6,083 respondents). MAIN OUTCOME MEASURES: Number of live births, knowledge of contraceptive methods, previous, current and intention for future use of contraceptives, method related problems. RESULTS: The contraceptive prevalence rate was 59.6% (CI 95% 58.4 to 60.9). The median number of live births was two (Q1 = 1, Q3 = 4) among all women, and seven (Q1 = 6, Q3 = 8) among women aged 40 to 49 years. Of 6,083 women interviewed, 5,849 (96.2%) knew at least one method of modern family planning, and 4,743 (78.0%) had ever used contraceptive in their life. Health concerns were the main reason for both discontinuation (28.5%) and postponement (22.8%) of contraceptive use. CONCLUSIONS: As compared to the 1991 Mother and Child Health Survey, knowledge and coverage of family planning services have improved further, and the introduction of injectable contraceptives has proved a success. Areas which need attention include the groups with high parity that remain under served, the low knowledge and use of condoms as a contraceptive, and the high level of health concerns among current and potential users. PMID- 10697917 TI - Malignant colorectal tumours in patients 30 years and below: a review of 35 cases. AB - OBJECTIVE: To review malignant colorectal tumour arising in patients 30 years and below. DESIGN: Retrospective cross sectional descriptive study. SETTING: Jos University Teaching Hospital, Jos, Nigeria. SUBJECTS: A total of 35 patients 30 years and below. MAIN OUTCOME MEASURES: Occurrence of malignant colorectal tumours tends to be higher among men and women over age 65 years and more common among Whites than Blacks. The disease rarely presents in the young population and the prognosis is usually unfavourable. This may be due to a delay in the diagnosis because colorectal cancer is not usually considered first in this age group. INTERVENTIONS: 15 patients had abdomino-perineal excision of the rectum, two right hemicolectomy, one left hemicolectomy, two anterior resection and six had colostomy and biopsy. RESULTS: Altogether, 149 patients were treated for large bowel cancer. From then, 35 (23.6%) given a yearly incidence of 3.5 were 30 years old or younger. The mean age was 25 (STD +/- 6) years, while the M:F ratio was 1.2:1. Weight loss, bloody mucoid diarrhoea, tenesmus and an anorectal mass were common clinical features present for more than six months. The rectum was involved in 24 patients (68.6%) and adenocarcinoma either well/moderately well differentiated or poorly differentiated was the predominant histological type. All the cancers except four were advanced at first presentation and treatment was merely palliative with only 30% of those treated and followed up still alive at six months. Complications of surgery were considered minor with the exception of the pelvic abscesses and deaths were due to the effects of the disease. CONCLUSIONS: This study illustrates that colorectal cancer is not rare, as it was previously believed. Presentation is commonly late and prognosis poor. In this age group, malignant colorectal tumours should frequently be considered in the differential diagnosis of bowel symptoms. The importance of the prudence of the general practitioner is thus emphasized. PMID- 10697918 TI - Traumatic intracranial aneurysms following penetrating stab wounds to the head: two unusual cases and review of the literature. AB - Two patients with rare complications of traumatic intracranial aneurysms following penetrating cranial stab wounds are described. One patient had a good outcome despite a secondary rupture of a traumatic proximal middle cerebral artery aneurysm, while the second patient had a traumatic basilar bifurcation artery aneurysm. To our knowledge neither the survival from a secondary rupture of a traumatic intracranial aneurysm, nor the development of a basilar bifurcation aneurysm secondary to a transcranial stab wound has been described previously. Furthermore, this is the first report of the technique of deep hypothermic cardiac arrest utilized to treat a traumatic false aneurysm. Traumatic intracranial aneurysms are a rare clinical entity, most often diagnosed after rupture and often resulting in fatal haemorrhage. A high index of suspicion needs to be maintained when managing patients with transcranial stab wounds. Early surgical intervention improves outcome by preventing initial aneurysmal rupture or rebleeding. PMID- 10697919 TI - Not everything acid-fast is Mycobacterium tuberculosis--a case of Nocardia. AB - We report a case of a 47 year old woman who presented with a history of motor convulsions and a three month history of an increasingly painful and progressively enlarging mass on the right side of her back. Neurological examination revealed generalised wasting and a right sided hemiplegia. A biopsy of the mass was taken for microbiology which reported growing branching gram positive rods after three days of incubation. A mycological diagnosis of Nocardia asteroides was made. An MRI scan revealed extensive infiltration of the fungal mass into extending from the base of the skull to fifth cervical vertebra. PMID- 10697920 TI - Current treatment and future prospects for the management of acute coronary syndromes. AB - The impact of ischaemic heart disease on the burden of cardiovascular disease continues to escalate worldwide, although international statistics suggest a levelling off in Western world, in the less industrialised parts of the world the effects of this disease are only beginning to be documented, nonetheless, rapid advances have been made in the diagnosis and management of the acute coronary syndromes (the term which encompasses the protean clinical manifestations of the ischaemic process). The therapeutic strategies discussed in this article cover two broad subjects that have been found to be critical in the evolution of the disease:- i. interfering with the haemostatic balance by retarding the thrombotic process; ii. modifying local and systemic vasoconstricting stimuli. PMID- 10697921 TI - Hypertension in pregnancy--1. AB - This review is aimed at clinicians working in a country like Zimbabwe, with limited health care resources. The management of the condition includes early detection, control of blood pressure, monitoring for maternal and foetal complications with timely delivery by the most appropriate route. PMID- 10697922 TI - The history of the Central African Journal of Medicine 1953 to 1999. AB - The Central African Journal of Medicine Company was founded in 1953 and registered in 1954 in accordance with the then existing company act. Its purpose was to assist medical personnel in central Africa find a place to publish the results of their research endeavours as well as an avenue to disseminate their clinical observation and updates. Since its first publication 46 years ago, to the present, the journal has attracted research papers from as far afield as Nigeria in West Africa, China, Hong Kong, the middle east and all the SADC states. PMID- 10697923 TI - Outbreaks of measles in communication with low vaccine coverage. PMID- 10697924 TI - Promptly reported diagnoses of HIV infection reached highest UK total in 1999. PMID- 10697925 TI - AIDS and HIV infection in the United Kingdom: monthly report. PMID- 10697926 TI - Nonobstructing exostoses of the external auditory canal. PMID- 10697927 TI - Endoscopic view of the lateral nasal wall following sinonasal surgery. PMID- 10697928 TI - Reactive swelling and cyst of the vocal fold. PMID- 10697929 TI - Hemorrhagic labyrinthitis. PMID- 10697930 TI - Electronystagmography in a patient with Meniere's syndrome. PMID- 10697931 TI - Low-dose oral methotrexate management of patients with bilateral Meniere's disease. AB - In this retrospective clinical trial, we evaluated the effectiveness of low-dose oral methotrexate in the management of bilateral Meniere's disease of immune mediated origin. At our tertiary-care referral center, we evaluated ten men and eight women who had longstanding bilateral Meniere's disease that had been unresponsive to traditional conservative medical management. Sixteen of these patients had steroid-responsive bilateral Meniere's disease. Two patients had contraindications to steroids, but their clinical and laboratory evaluations were consistent with an immune-mediated process. Patients were treated with 7.5 to 20 mg/week of oral methotrexate. The mean duration of treatment was 16.7 months (range: 8 to 35), with a mean followup of 2 years (range: 9 mo to 5 yr). Changes in clinical symptoms (vertigo, hearing loss, tinnitus, and aural fullness), audiometric changes, and side effects of therapy were evaluated. Vertigo resolved in 14 patients (78%), was substantially alleviated in three patients (17%), and remained unchanged in one patient (6%). Hearing improved in five patients (28%) and stabilized in seven patients (39%). Tinnitus and aural fullness resolved or was relieved in 11 of 17 (65%) and 13 of 14 (93%) patients, respectively. Side effects were minimal and reversible. We conclude that low-dose oral methotrexate is effective and safe for treating bilateral Meniere's disease of immune-mediated origin. In this study, methotrexate alleviated vertiginous symptoms and improved or stabilized hearing in most patients. Low-dose methotrexate can be considered for patients with immune-mediated bilateral Meniere's disease when long-term treatment is required or when a steroid or cyclophosphamide is contraindicated. PMID- 10697932 TI - Osteoradionecrosis of the temporal bone in nasopharyngeal carcinoma after radiotherapy: a case report. AB - Osteoradionecrosis of the temporal bone after external-beam radiotherapy for nasopharyngeal carcinoma is not uncommon following a long posttreatment interval. We describe the case of a man who had experienced this complication 13 years after he had undergone such radiotherapy. His condition resolved after removal of dead bone from the external auditory canal, followed by antibiotic therapy and periodic aural toileting. PMID- 10697933 TI - Cutaneous metastatic lung cancer: literature review and report of a tumor on the nose from a large cell undifferentiated carcinoma. AB - Cutaneous metastatic disease is a prognostically important diagnosis. We report the case of a 64-year-old man who had an uncommon histologic type of lung cancer- a large cell undifferentiated carcinoma, which was metastatic to the skin of the nose. The relative frequency of cutaneous metastasis is similar to that of primary cancers. Cutaneous disease as the first sign of metastasis is most often seen in cancer of the lung. However, its appearance as a large tumor on the nose, which was observed in this case, is unusual. PMID- 10697934 TI - Dislocation and hypertrophy of the medial head of the clavicle: an unusual late complication of radical neck dissection. AB - We report three cases of a rare late complication of neck dissection: anterior dislocation and hypertrophy of the sternal head of the clavicle manifesting as a hard lump in the lower neck. We describe the mode of presentation, etiology, and methods of prevention. PMID- 10697935 TI - Gustatory sweating syndrome of the submandibular gland. AB - Gustatory sweating syndrome involving the submandibular gland is rare. We present a case of a patient who experienced this syndrome 5 years after undergoing submandibular gland resection. Our patient was satisfied simply with an explanation of the disorder and reassurance. But in cases where further intervention is sought, medical and surgical options are available and should be individualized for the patient. PMID- 10697936 TI - A clinical comparison of outpatient and standard myringoplasty. AB - This prospective study compared the surgical outcomes of 35 patients who underwent myringoplasty--16 who were treated as outpatients and 19 who were admitted as inpatients. The outpatient technique involved a free skin graft, with the temporalis fascia placed as an underlay graft. The inpatients underwent the standard myringoplasty procedure. Postoperatively, 14 of the 16 outpatients (87.5%) and 17 of the 19 inpatients (89.5%) were completely healed within 2 weeks. The results of this study indicate that outpatient myringoplasty with a free skin graft is as safe and effective as standard myringoplasty for most patients. PMID- 10697937 TI - Nasal foreign body: removal of an open safety pin from the left nostril. AB - We describe the case of a woman who presented with an open safety pin lodged in her left nostril. An attempt to remove the pin with the patient under local anesthesia was not successful. Removal was eventually accomplished in the operating room with the patient under general anesthesia. PMID- 10697939 TI - Oral and maxillofacial surgery advances in implant dentistry. PMID- 10697938 TI - Biomaterials and biomechanics of oral and maxillofacial implants: current status and future developments. AB - Research in biomaterials and biomechanics has fueled a large part of the significant revolution associated with osseointegrated implants. Additional key areas that may become even more important--such as guided tissue regeneration, growth factors, and tissue engineering--could not be included in this review because of space limitations. All of this work will no doubt continue unabated; indeed, it is probably even accelerating as more clinical applications are found for implant technology and related therapies. An excellent overall summary of oral biology and dental implants recently appeared in a dedicated issue of Advances in Dental Research. Many advances have been made in the understanding of events at the interface between bone and implants and in developing methods for controlling these events. However, several important questions still remain. What is the relationship between tissue structure, matrix composition, and biomechanical properties of the interface? Do surface modifications alter the interfacial tissue structure and composition and the rate at which it forms? If surface modifications change the initial interface structure and composition, are these changes retained? Do surface modifications enhance biomechanical properties of the interface? As current understanding of the bone-implant interface progresses, so will development of proactive implants that can help promote desired outcomes. However, in the midst of the excitement born out of this activity, it is necessary to remember that the needs of the patient must remain paramount. It is also worth noting another as-yet unsatisfied need. With all of the new developments, continuing education of clinicians in the expert use of all of these research advances is needed. For example, in the area of biomechanical treatment planning, there are still no well-accepted biomaterials/biomechanics "building codes" that can be passed on to clinicians. Also, there are no readily available treatment-planning tools that clinicians can use to explore "what-if" scenarios and other design calculations of the sort done in modern engineering. No doubt such approaches could be developed based on materials already in the literature, but unfortunately much of what is done now by clinicians remains empirical. A worthwhile task for the future is to find ways to more effectively deliver products of research into the hands of clinicians. PMID- 10697940 TI - Current status of dental implants: a periodontal perspective. PMID- 10697941 TI - Implant prosthodontics: current perspective and future directions. PMID- 10697942 TI - Implants and components: entering the new millennium. PMID- 10697943 TI - Experimental alveolar ridge augmentation by distraction osteogenesis using a simple device that permits secondary implant placement. AB - The purpose of this study was to develop an improved technique of alveolar ridge augmentation by distraction osteogenesis using distraction screws, and to investigate tissue reactions to titanium implants at the distraction site. The left mandibular premolars were extracted from 6 adult dogs. After 12 weeks, a box shaped osteotomy of the alveolar bone was carried out, and distraction devices were placed on the transport and base segments. After a 7-day latency period, the alveolar bone was augmented by 7 mm vertically at a rate of 1.0 mm/day. Just after distraction, these devices were replaced with dental implants for fixation of the transport segment and bone formation of the distraction site. Histologic and radiographic evaluations were made at 8 and 12 weeks after distraction. Vertical augmentation averaged 6.1 mm after 12 weeks of consolidation. It was possible to lengthen the alveolar bone without great difficulty, and good bone formation was recognized in the distraction site. Greater integration between the implant and the distracted bone was observed at 12 weeks after distraction than at 8 weeks. Distraction osteogenesis was successfully applied to alveolar ridge augmentation by this improved technique, and the implants osseointegrated in the augmented ridge. PMID- 10697944 TI - Influence of bicortical or monocortical anchorage on maxillary implant stability: a 15-year retrospective study of Branemark System implants. AB - The present study evaluated implant survival and marginal bone loss in maxillae over a 15-year follow-up period as a function of either monocortical or bicortical implant anchorage. Of 207 standard Branemark implants (10 mm in length) followed, 110 implants were judged to be monocortically anchored and 97 as bicortically anchored. The bicortically anchored implants failed nearly 4 times more often than the monocortical ones. Implant fractures accounted for over 80% of the observed failures and were found to affect the bicortical group almost 3 times more often. As tentative explanations, induction of increased stress and bending forces resulting from possible prosthetic misfit, presence of unfavorable arch relationships, or high occlusal tables in combination with bicortically anchored implants have been suggested, all indicating an overambitious fixation of the bicortical anchorage. Total marginal bone loss was low over the 15-year period and close to identical for the 2 groups, suggesting that the mode of cortical anchorage did not have any clinically significant influence on marginal bone remodeling. PMID- 10697945 TI - Measurement of misfit at the implant-prosthesis interface: an experimental method using a coordinate measuring machine. AB - Measurement of misfit at the implant-prosthesis interface is a difficult procedure. One factor common to all methods that attempt to measure 3-dimensional distortion to the micron level is the difficulty in providing verifiably consistent reference points between individual measurement sets. Consequently, the majority of studies use a relative distortion model in which the coordinate reference system is integral to the framework, thus limiting the value of the data gathered. In the method described, the datum plane and the coordinate reference system were set up external to the framework and could be re established between measurement sets in a verifiable manner. PMID- 10697946 TI - Tissue regeneration adjacent to titanium implants placed with simultaneous transposition of the inferior dental nerve: a study in dogs. AB - Transposition of the inferior alveolar nerve was performed in an experimental dog model. Four adult greyhounds were used in the study. Surgical transposition of the nerve was made bilaterally, and 3 implants were placed on each side while the nerve was lateralized. On one side, the nerve was repositioned in contact with the implants, while on the contralateral side a resorbable membrane was positioned between the implant surface and the neurovascular bundle. Histologic section after 4 months of healing showed an intimate contact between implants and nerve tissue in all cases without an interpositional membrane, in contrast to cases with membranes. Histomorphometric measurements of the distance between the implants and the nerve tissue showed that the membrane side had a considerably larger distance between the implant and the nerve, although not with concomitant bone formation. PMID- 10697947 TI - Treatment of peri-implantitis defects with autogenous bone grafts: six-month to 3 year results of a prospective study in 17 patients. AB - As part of an ongoing prospective study, the treatment of peri-implantitis defects using autogenous bone grafts was evaluated. This present report is based on data from 25 ITI screw implants in 17 patients with progressive peri-implant tissue destruction during the maintenance phase. Treatment of these lesions included raising flaps, removal of the surrounding granulation tissue, and air polishing of the implant surface. Subsequently, corticocancellous bone grafts or particulate bone were placed into the peri-implant osseous defects, and the flaps were sutured around the cervical segment of the implants, allowing for transmucosal healing. Two of the 25 cases resulted in a negative outcome of the procedure. One of the transplants had to be removed 40 days after augmentation because of flap dehiscence and graft mobility. In another patient, the healing period was uneventful until the re-entry surgery, but when the site was reopened, the total graft volume was resorbed. The primary therapeutic success at re-entry surgery evaluated by intraoperative measurements resulted in a median defect depth reduction of 6.9 to 0.7 mm (P = .001), corresponding to a bone repair of 90%. The change in defect width was 1.9 mm (P = .002, repair 100%). A positive result of the reconstructive therapy has been observed during a re-evaluation time of up to 3 years. Median marginal bone loss was reduced from 6.2 to 2.3 mm after 2 and 3 years, respectively. The median vertical bone resorption of 4.5 mm was completely repaired. The crevicular fluid volume, a parameter of the level of marginal inflammation, along with probing depths and attachment levels, were reduced to a physiologic rate. The implant observation period until the first appearance of the lesion seems to be crucial to the effectiveness of the therapy. Early failures appearing within the first 2 years after implant placement showed a more stable therapeutic result over time. PMID- 10697948 TI - Subjectivity and unconsciousness. AB - Concepts of the unconscious were crucial to both Jung's and Freud's thinking. Psychoanalytic and analytical psychological views of the unconscious are compared and contrasted, and both are critically reviewed. It is suggested that we need to revise our conceptualization so as to take better account of the role of the analyst's expectations and inferences, and therefore of his or her subjectivity, whenever he or she makes a clinical judgement that unconscious mental processes are in operation. Some technical implications of a revised definition of unconsciousness are considered, especially indications for self-disclosure by an analyst of his or her own experience of events within the treatment. PMID- 10697949 TI - Opening the mind to reality. AB - The author shows, through the use of clinical material, how an early failure in love can give rise to a severely crippling superego. The experience of a hateful relation with the mother is not simply internalized as a persecuting internal object, but is grafted onto the very roots of superego formation. As a result, the development of other parts of the psyche are affected--specifically the relation between the ago and self and the development of sexuality. The alienation between ego and self impairs thinking and the perception of external reality, which is modified and denied in the service of maintaining a pathological superego. By allowing the patient's hateful feelings to come out in the transference, without making him feel guilty, he is then able to risk expressing his loving feelings without the fear of rejection or abandonment. Through this process, the pathological superego can be dismantled and a more benign superego constructed. PMID- 10697950 TI - Theatres of the psyche. AB - Fragments of the analytic voyage of a forty year old patient, haunted by the idea of her death since infancy when confronted with the anguish that accompanies the discovery of a potentially death dealing illness. In three years of intensive analytic work, her way of functioning psychically changed dramatically (her flight from imaginative life, total disaffectation of her emotional life, psychic deafness regarding her body and its messages...). These discoveries led to my analysand's resuming her psychoanalytic voyage in these words: 'Even if I am destined to die of this cancer, at least I shall have lived'. PMID- 10697951 TI - Response to Joyce McDougall. The triumph of compassion over mourning. PMID- 10697952 TI - Substantive unconscious and adjective unconscious: the contribution of Wilfred Bion. AB - The author first provides her readers with a brief summary of some of Freud's ideas, as found throughout his work, on the notion of 'unconscious'. The notion of unconscious as noun is contrasted to the idea of unconscious as adjective, this latter being proposed as a quality, or a state, ever temporary, dynamic, and subject to the constant changes going on in the individual's internal psychic world, as well as to external conditions. After presenting some considerations, the author then contrasts the Kleinian model of the mind to the Freudian, and Wilfred Bion's contribution is discussed at some length. Within Bion's conception of psychic functioning, the model of 'dream' is highlighted and, in this regard, clarifications are sought regarding Bion's view of the unconscious. To conclude, a brief and superficial approximation to the work of Carl Jung is touched upon, although the author admits to knowing little of Jung's positions. PMID- 10697953 TI - Transference and dream in illness: waxing psyche, waning body. AB - In times of change, crisis, and illness, the excited points of an individual's personal history are reactivated within the transference and may also be noted by observing countertransference. When there are anomalies in the emotional and imaginal circle of the therapeutic relationship, there is occasion for repetition and/or a transformative opening. In some cases, there is simultaneous treatment of severe developmental fixations and compulsions, and issues of individuation. Images may emerge both from the personal field and from the collective and archetypal imagination. These may be expressions of interpersonal experience, intrapsychic dynamics, and physical as well as psychic state. PMID- 10697954 TI - Philosophical assumptions in Freud, Jung and Bion: questions of causality. AB - The historical development of concepts of causality in philosophy is described. Since the Enlightenment and the growth of science, exponents of the two most important concepts, determinism and teleology, have been in conflict. At the inception of psychoanalysis at the end of the nineteenth century this conflict was particularly intense. It was the cause of the first major schism in psychoanalysis between Jung and Freud. Psychoanalytic theorists have continued to disagree over this issue. Post-modernist philosophy has abolished all metaphysics and therefore called into question concepts of psychic causality. Parallel to, but uninfluenced by this development, Bion has developed a psychoanalytic conceptualization which may be seen as transcending causality. The clinical and theoretical implications of these developments are described. PMID- 10697955 TI - Enzymatic corrections for cells derived from Fabry disease patients by a recombinant adenovirus vector. AB - Fabry disease is an X-linked inherited metabolic disorder caused by a deficiency of alpha-galactosidase (alpha-gal), resulting in the accumulation of ceramide trihexoside (CTH) in body fluids and in many organs and tissues. We constructed a recombinant adenovirus with a human alpha-gal cDNA (AxCAG alpha-gal), and transfected this vector to skin fibroblasts from Fabry patients. Transfected cells expressed high amounts of alpha-gal in their cytoplasm, and a high level of alpha-gal activity was detected in the medium. The accumulated CTH in the fibroblasts disappeared 3 days after infection. The secreted alpha-gal also eliminated the accumulated CTH from uninfected patient's cells. The enzyme may be taken up through mannose-6-phosphate receptors, as the addition of mannose-6 phosphate to the medium completely inhibited the uptake of the enzyme. The infected cells continued to express alpha-gal for more than 10 days. These results suggest that AxCAG alpha-gal could be used as enzyme replacement gene therapy for Fabry disease. PMID- 10697956 TI - Genomic organization and chromosomal mapping of ELKS, a gene rearranged in a papillary thyroid carcinoma. AB - We recently isolated a novel cDNA, designated ELKS, that was fused to RET cDNA in a papillary thyroid carcinoma. Its encoded polypeptide sequence was rich in glutamic acid (E), leucine (L), lysine (K), and serine (S), and was characterized by the presence of nine alpha-helical coiled-coil domains consisting of periodic heptad repeats. We have now cloned the entire structure of the human ELKS gene from within a 700-kb genomic region represented by overlapping bacteriophage P1 derived artificial chromosome (PAC) and bacterial artificial chromosome (BAC) clones, and localized it to chromosomal band 12p13.3 by fluorescence in situ hybridization. The gene is approximately 500 kb long, with 19 exons and 18 introns; the transcription initiation site within exon 1 is separate from the initiation codon (in exon 2). Analysis of the exon/intron structure revealed that introns interrupt the coding sequence in such a way that many functional segments of the protein are encoded by distinct exons. Exon 1 encodes the 5' non-coding region; exons 2, 3, 6, 7, 8, 9, 11, 14, and 15 encode the nine coiled-coil domains. Exons 17-19 constitute the 3' non-coding region. Analysis of the region immediately upstream of exon 1 showed that it was extremely rich in G/C nucleotides and contained multiple Sp-1 and AP2 binding sequences. The ELKS-RET gene fusion rearrangement we had observed in a papillary thyroid carcinoma occurred between intron 10 of the ELKS gene and intron 11 of RET. PMID- 10697957 TI - Molecular cloning and characterization of two novel genes on chromosome 8p21.3. AB - Through large-scale sequencing of genomic DNA from human chromosome 8p22-p21.3 we have isolated two novel genes, designated GK1 and G5. Their predicted products showed no significant similarity to any known proteins in public databases. A comparison of GK1 cDNA sequences, which encode a 1270-amino-acid protein, with corresponding genomic DNA sequences revealed that this gene consists of 15 exons and spans an approximately 113-kb genomic region. Northern blot analysis revealed ubiquitous expression of 7.0- and 4.4-kb transcripts; in addition, we detected a 5.0-kb skeletal muscle-specific transcript and a 4.0-kb transcript specifically expressed in heart and pancreas. Computer and immunocytochemical analyses of a GK1 Green fluolesent protein (GFP) fused construct indicated that the gene product, which contains putative leucine-zipper domains, was likely to be a mitochondrial protein. The other novel gene, G5, expressed four transcripts (4.2, 2.2, 1.7, and 1.0-kb) ubiquitously; the longer three transcripts, which differed only in the 3'-non coding region, encoded identical 397-amino-acid peptides. The G5 gene consists of 14 exons and spans approximately 52 kb of genomic DNA; its deduced 397-amino acid product appears to contain coiled-coil domains and a proline-rich region, and to be located in cytoplasm. PMID- 10697958 TI - Transduction of fibroblasts and CD34+ progenitors using a selectable retroviral vector containing cDNAs encoding arylsulfatase A and CD24. AB - Metachromatic leukodystrophy (MLD) is an autosomal recessive, inherited, lysosomal storage disease caused by a deficiency in arylsulfatase A (ASA). This disease is characterized by progressive demyelination leading to severe neurological symptoms. Allogenic bone marrow transplantation at an early stage of clinical course is only effective treatment currently available. Accordingly the corrective transfer of the ASA gene into hematopoietic stem cells is thought to be an important option for curative treatment for MLD. We have recently developed a selectable vector system based on ex vivo sorting of transduced cells (Migita et al. 1995). In this study, we applied this selectable system for development of MLD gene therapy. A bicistronic retroviral vector containing ASA cDNA and CD24 cDNA as a selectable marker gene was constructed. This vector was successfully transduced on fibroblasts from MLD patients, ASA activity was increased 7-fold compared to normal untransduced cells. PCR Southern analysis of hematopoietic colonies showed that transduction efficiency of CD34+ cells was 11-22%. However, after fluorescence-activated cell sorting using anti-CD24 antibody, 75-100% of colonies became vector positive. The sorting raised the ASA activity several fold compared to untransduced CD34+ progenitors. These results suggest that a bicistronic ASA vector containing a CD24 selectable marker could be a useful component of gene therapy for MLD. PMID- 10697959 TI - Novel polymorphisms of the AP-2 gene (6p24): analysis of association with schizophrenia. AB - The transcription factor activator protein 2 (AP-2) gene is a possible candidate gene for schizophrenia, since it maps near D6S470, a marker on chromosome 6p24 that provided evidence of linkage to schizophrenia. In the present study we analyzed the promoter region and the whole coding region of the human AP-2 gene in order to identify genetic variations that may lead to the modification of AP-2 expression or the alteration of protein function, contributing to schizophrenia or particular schizophrenic phenotypes. Genomic DNA was isolated from the whole blood samples of 87 unrelated schizophrenics and 100 healthy controls. Polymerase chain reaction (PCR) was performed, using 15 primer sets that spanned the promoter region and the whole coding region, and amplified products were screened by single-strand conformational polymorphism (SSCP) analysis. Aberrant SSCP patterns were analyzed by direct sequencing. Three novel polymorphisms were found in the promoter region; two relatively common (-90G-->C, -803G-->T) and one rare (-1769G-->A). Polymorphic status at both loci suggested strong linkage disequilibrium between the -90G and -803G alleles, and between the -90C and -803T alleles. Although no significant differences in genotypic and allelic frequencies at the -90 and -803 loci were found between patients and controls, significant differences in the distribution of genotypes at the -90 (P = 0.008) and -803 (P = 0.037) loci were observed in patients with an episodic course compared with controls. However, the difference for the -803 locus was not significant after Bonferroni correction for multiple comparisons. Our data provided no direct evidence of an association between schizophrenia and the polymorphisms of the AP 2 gene, although the positive result at the -90 locus in schizophrenics with an episodic course is potentially interesting. PMID- 10697960 TI - cDNA cloning of a novel human gene NAKAP95, neighbor of A-kinase anchoring protein 95 (AKAP95) on chromosome 19p13.11-p13.12 region. AB - A-kinase anchoring protein 95 (AKAP95) is a nuclear protein which binds to the regulatory subunit (RII) of cyclic adenosine monophosphate (cAMP)-dependent protein kinase (PKA) and to DNA. A novel nuclear human gene which shares sequence homology with the human AKAP95 gene was identified by a nuclear transportation trap method. By polymerase chain reaction (PCR)-based analysis with both a human/rodent monochromosomal hybrid cell panel and a radiation hybrid panel, the gene was mapped to the chromosome 19p13.11-p13.12 region between markers WI-4669 and CHLC.GATA27C12. Furthermore, alignment with genomic sequences revealed that the gene and human AKAP95 resided tandemly only approximately 250 bp apart from each other. We designated this gene as neighbor of AKAP95 (NAKAP95). The exon intron structure of NAKAP95 and AKAP95 was conserved, indicating that they may have evolved by gene duplication. The predicted protein product of the NAKAP95 gene consists of 646 amino acid residues, and NAKAP95 and AKAP95 had an overall 40% similarity, both having a potential nuclear localizing signal and two C2H2 type zinc finger motifs. The putative RII binding motif in AKAP95 was not conserved in NAKAP95. A reverse transcription coupled (RT)-PCR experiment revealed that the NAKAP95 gene was transcribed ubiquitously in various human tissues. PMID- 10697961 TI - cDNA cloning, expression profile, and genomic structure of human and mouse RNF10/Rnf 10 genes, encoding a novel RING finger protein. AB - RING finger (C3HC4-type zinc finger) is a variant zinc finger motif present in a new family of proteins including transcription regulators. A new member of the RING finger protein family was identified through a mouse expressed sequence tag (EST) database search, and its full-length cDNA was isolated from a mouse brain full length-enriched cDNA library. The gene was designated as Rnf10, for RING finger protein 10. The cDNA clone consists of 3110 nucleotides and encodes an open reading frame (ORF) of an 804-amino acid protein. A database search revealed that human KIAA0262 protein (accession number, D87451) has strong homology to mouse Rnf10. To confirm that mouse Rnf10 is the homolog or an isolog of human KIAA0262, a human RNF10 cDNA was cloned in our hands from a fetal brain cDNA pool. The newly isolated cDNA contained an ORF for 811 amino acids which had almost identical structure to mouse Rnf10 protein, indicating that the human ORF codes for RNF10 protein. This finding was also supported by comparative chromosome mapping in which both genes were localized in a conserved linkage homology region between mouse and human. Comparison of the RNF10 and KIAA0262 proteins revealed that both were transcribed from the same gene and that the longer RNF10 ORF would be the authentic form. The complete genomic organization of RNF10 was determined to consist of 17 exons spanning at least 40kb in the genome. PMID- 10697963 TI - A family with hydrocephalus as a complication of cerebellar hemangioblastoma: identification of Pro157Leu mutation in the VHL gene. AB - Various mutations in the VHL gene on chromosome 3p25-26 are responsible for von Hippel-Lindau (VHL) syndrome. We report on a Japanese VHL family in which two of the three affected members developed acute occlusive hydrocephalus that necessitated emergency surgery for ventricular shunt or drainage. Direct sequencing and restriction fragment length polymorphism analysis identified a germline missense mutation, Proline-to-Leucine, caused by a C-to-T transition at the second nucleotide of codon 157. PMID- 10697962 TI - Isolation and characterization of a human cDNA encoding a protein homologous to the 7.2-kDa protein (subunit X) of bovine ubiquinol-cytochrome C reductase. AB - Through large-scale sequencing of clones randomly selected from a library of human cDNAs, we have isolated a novel human gene termed hUQCR10. Its open reading frame encodes 63 amino acids that share 88.5% identity with the sequence of bovine ubiquinol-cytochrome C reductase 7.2-kDa protein (subunit X). A single 0.6 kb transcript was expressed in all human tissues examined, but was particularly abundant in heart and skeletal muscle, tissues that consume a large amount of oxygen. The gene product therefore may play a significant role in the cellular respiratory system. In support of this hypothesis, our immunohistochemical analysis revealed that the hUQCR10 protein is located in mitochondria. A homology search using computer programs determined the chromosomal localization of the gene at 22q12. PMID- 10697964 TI - Identification of two novel mutations of the carnitine/acylcarnitine translocase (CACT) gene in a patient with CACT deficiency. AB - Carnitine/acylcarnitine translocase (CACT) transports acylcarnitines into mitochondria in exchange for free carnitine, and is therefore an essential component within the fatty acid beta-oxidation pathway. CACT deficiency is an autosomal recessive disease caused by a mutation of the CACT gene. We have identified two novel mutations of the CACT gene in a patient with CACT deficiency. The first, a deletion mutation (146 del T), leads to premature termination and results in a very immature CACT protein. The second, a splicing mutation (261-10T > G), results in either skipping of exons 3 and 4, or of exon 3 alone, and leads to truncation of the protein. Each of these mutations is hypothesized to destroy the function of the CACT protein. We propose that each of these mutations of the CACT gene play a causative role in the disease. PMID- 10697965 TI - Identification of a novel Tru9 I polymorphism in the human vitamin D receptor gene. AB - We found a novel Tru9 I restriction polymorphism in intron 8 of the vitamin D receptor (VDR) gene in healthy French Caucasians. It corresponds to a substitution of A for G at nucleotide +443 bp from the end of exon 8. The allelic frequency of G and A in 151 unrelated subjects was 0.894 and 0.106, respectively. This polymorphism is located in the reverse primer binding site of primers that have been frequently used in the literature to genotype a BsmI restriction polymorphism. The presence of the Tru9I A allele may result in allele drop-out when the BsmI restriction fragment length polymorphism (RFLP) is genotyped with the original set of primers. This novel Tru9I polymorphism may be useful for analysis of the VDR gene. PMID- 10697966 TI - Identification of a novel single nucleotide polymorphism (SNP) in the human organic cation transporter-like 2-antisense (ORCTL2S) gene. AB - We found a single nucleotide polymorphism (SNP) in exon 3 of the human organic cation transporter-like 2-antisense (ORCTL2S) gene: a base substitution A266G which was confirmed by direct sequencing. Heterozygosity of the polymorphic alleles was 0.45 in a Japanese population. This polymorphism will be useful in the allelic expression analysis of the ORCTL2S gene. PMID- 10697967 TI - Mutation analysis of two Japanese patients with Fanconi-Bickel syndrome. AB - Fanconi-Bickel syndrome (FBS), or glycogen storage disease type XI, is a rare autosomal recessive disorder characterized by hepatorenal glycogen accumulation, Fanconi nephropathy, and impaired utilization of glucose and galactose. Recently, this disease was elucidated to link mutations in the glucose transporter 2 (GLUT2) gene. Only three mutations in three FBS families have been reported. Therefore, it is important to elucidate mutations in the GLUT2 gene in FBS by answering the question of whether the syndrome is a single gene disease. In this report, we describe two patients in two unrelated families clinically diagnosed with FBS. No mutation in the entire protein coding region of the GLUT2 gene was detected in patient 1, which suggested that no mutation existed in the GLUT2 gene, or that some mutations had affected the expression of the GLUT2 gene. In patient 2, a novel homozygous nonsense mutation (W420X, Trp at codon 420 to stop codon) was detected. These results support the correlation between GLUT2 gene mutation and FBS syndrome. However, many patients must be analyzed to determine whether other genes are involved in FBS. PMID- 10697969 TI - The instep plantar fasciotomy for chronic plantar fasciitis. A retrospective review. AB - A retrospective study was conducted on the use of the instep plantar fasciotomy for the treatment of recalcitrant plantar fasciitis. A total of 83 patients (94 feet) were analyzed. The average postoperative follow-up time was 20.9 months. Surgery was deemed successful 93.6% of the time, and in 95.7% of cases, the patient would recommend the procedure to someone with the same condition. The main complications were scarring (9.6%), medial arch or heel pain (7.5%), cramping in the arch (6.4%), lateral column pain (5.3%), aching or pain across the dorsal midfoot (5.3%), and burning or tingling of the ball of the foot (5.3%). PMID- 10697968 TI - The potential benefits of advanced therapeutic modalities in the treatment of diabetic foot wounds. AB - This article discusses the advantages and disadvantages of primary wound healing as compared with primary amputation in individuals with chronic diabetic foot wounds. The authors review the potential benefits of vascular surgical procedures and advanced dressings, including two of the most promising modalities in modern wound care: growth factors and bioengineered skin. In this era of cost-conscious health-care administration, it is incumbent on the practitioner to consider not only the basic science of wound care, but also the economic aspect of treatment rendered. These various interventions, dressings, growth factor delivery systems, and new modalities could significantly reduce healing time, thereby reducing the risk of infection, hospitalization, and amputation while improving quality of life. If so, they may be truly cost-effective. PMID- 10697970 TI - The effectiveness of gait plates in controlling in-toeing symptoms in young children. AB - A range of patient-oriented and practitioner-oriented outcomes were used to evaluate the efficacy of "gait plate" shoe inlays in controlling symptoms associated with in-toeing in otherwise healthy children. For 18 in-toeing children, parents completed a preintervention questionnaire. Then, during randomized trials, foot placement angle was measured both with and without gait plate inlays in the children's footwear. After the children had worn the gait plates for 1 month, a simple questionnaire was used to rate parental satisfaction with a range of factors associated with control of symptoms. The use of gait plate inlays resulted in a small but statistically significant reduction in the amount of in-toeing as measured by foot placement angle. Gait plates reduced the reported frequency of tripping in 14 of the 18 cases. The reported parental satisfaction was high or very high in all but one case, suggesting that this intervention warrants further investigation as an alternative to "observational management" for symptomatic in-toeing. PMID- 10697971 TI - Freiberg's infraction in a collegiate heptathlete. AB - In 1914, Freiberg described a condition of infraction of the second metatarsal head. At the time, he considered direct trauma to be the etiologic agent, but he later disproved this by demonstrating a case without a history of trauma. This article describes a case of Freiberg's infraction in a female collegiate heptathlete and the effects of conservative treatment of her condition with custom-made orthoses. PMID- 10697972 TI - Mycetoma. AB - Subcutaneous fungal infections are relatively uncommon in the lower extremity. Mycetoma begins as painless papules or nodules that increase in size and progresses to involve the connective tissue. Diagnosis is based on biopsy, with definitive identification of the organism needed for effective treatment. Treatment consists of antifungal medications and surgical debridement. This article provides an overview of this disorder and reports on a case of recurrent mycetoma in a 70-year-old woman. PMID- 10697973 TI - 1999 Arthritis Survey. American Podiatric Medical Association. AB - This report presents the results of analyses of statistical data from 1,114 members of the American Podiatric Medical Association (APMA) who responded to the 1999 Arthritis Survey, conducted from July through August 1999. The purpose of the survey was to determine the extent and methods of treatment of patients with arthritis of the foot or ankle by doctors of podiatric medicine. PMID- 10697974 TI - Wound-care resources on the Internet. AB - The Internet offers many resources in the area of wound and ulcer care that are of potential interest to podiatric physicians and students. This article provides an overview of World Wide Web sites that contain factual information, management guidelines, and illustrations pertaining to various aspects of wound and ulcer care. Web sites that emphasize preventive care are also reviewed. Because the prudent use of antimicrobial therapy is an important part of wound care, a few sites that offer antibiotic information are described. PMID- 10697975 TI - Paronychia in patients receiving antiretroviral therapy for human immunodeficiency virus infection. PMID- 10697976 TI - A simple cure for Morton's neuralgia. PMID- 10697977 TI - Avulsion fracture of the plantar lateral base of the first metatarsal. PMID- 10697978 TI - Retained digital foreign body after a pellet gun injury. AB - A symptomatic foreign body embedded in the human body can be a frustrating problem for physician and patient alike. A unique case of a retained foreign object resulting from a pellet gun injury has been presented. Although the course of treatment in this case was uncomplicated, it is important to understand the complexities of the human body's response to foreign bodies. PMID- 10697979 TI - [Glucocorticoid-induced osteoporosis]. AB - Chronic glucocorticoid treatment is the most frequent cause of secondary osteoporosis. The direct inhibitory effect of glucocorticoids on osteoblasts results in decreased bone formation. Increased osteoclastic bone resorption due to low concentrations of gonadal steroid hormones and glucocorticoid-induced direct suppression of intestinal calcium absorption also contribute to the decrease of bone mass in these patients. Bone loss is rapid, particularly in the first months of glucocorticoid therapy. Bone mineral density of the lumbar spine and proximal femur should be measured in patients who are starting chronic therapy with glucocorticoids. Although glucocorticoid-induced osteoporosis is a severe and nowadays partially preventable disorder, osteoporosis prophylaxis is only rarely prescribed to these patients. Recent randomized, controlled trials proved the therapeutic effects of hormone replacement therapy, as well as of bisphosphonates and active vitamin D analogs in primary and secondary prevention of glucocorticoid-induced osteoporosis. PMID- 10697980 TI - [Dermatomyositis: clinical study of 34 patients]. AB - Clinical data of 34 patients with DM, who have been treated during the years 1971 and 1998 were evaluated. 79% of the patients (27 patients) were female, 21% of them (7 patients) were male. 59% of the patients (20 pts) were between the ages of 41 and 50 years. The characteristic heliotrop rash were observed in 26 patients, Gottron's papules in 20 patients, poikiloderma in 2 patients, calcification, ulcers, Raynaud syndrome in 1 patient. 3 of the 34 patients presented with strongly itching erythematopapulosus symptoms, most prominently on the scalp. Cardiac involvement were present in 10 patients (29%), lung involvement in 8 patients (23%), gastrointestinal complaints in 11 patients, dysphagia, dysphonia in 4 patients, joint pain in 5 patients. Overlap syndrome- scleroderma-dermatomyositis, SLE-dermatomyositis--was present in 2 patients. 9 of the 34 patients (26%) suffered from malignant tumours: gastric, breast, lung, epipharynx carcinoma, malignant melanoma. 13 of the 34 patients have been treated with corticosteroids together with immunosuppressor agents, in most cases azathioprin was administered. Cyclosporin was given in 7 cases, chloroquine in 2 cases. PMID- 10697981 TI - [Problems of treating burn injuries in Hungary in the light of advancements during the last half century]. AB - Until the middle of 20th century the burnt patients were treated on dermatological departments, and skin transplantations were performed after the regular conservative wound care too late and rarely. By reason of the medical experiences of the 2nd World War special departments were founded for the burn injured patients, it happened likewise in our country in 1953 as well. The specialists recognized that the burn injury causes a general disease (multiple organ failure), moreover only an aggressive surgical tactics can product any good results. The author presents the progress and problems of Hungarian burn treatment since 1953. A significant part of the burnt patients has been treated firstly not in burn departments, and sometimes the first therapy is insufficient, because the national medical training for cure of burnt patients is deficient. For this reason the author gives a simple treatment guide for the first two postburn days, and finally he summarizes the conditions of advance. PMID- 10697982 TI - [Clinical experience with raloxifene]. AB - Selective receptor modulators (SERMs) are drugs which act via the estrogen receptors and possess tissue specific estrogenic or anti-estrogenic properties. The bone and cardiovascular effects of SERMs are estrogen-like, while they have an effect as estrogen antagonist in the mammary tissues. Raloxifene is the first representative of selective estrogen receptor modulators which does not cause estrogenic effects in the uterus. Based on numerous recently completed controlled clinical trials, the authors characterize the clinical features of raloxifene to assess its therapeutic potency. PMID- 10697983 TI - [A case of retroperitoneal pancreatic abscess spreading to the femoral region]. AB - Authors report the case of a 47 years old male patient with acute femoral abscess. The examinations made with urgency found proceeding acute pancreatitis, fluid collection in the right pleural cavity, exceeding fluid collections in the retroperitoneum and right paracolic region. The inflammatory infiltrate and collection in the upper third of the right thing seemed to be in connection with the proceeding pancreatitis. Because of the process endangering also the viability of the limb and severe septic state, an acute operation was performed, in the course of which extension of the retroperitoneal abscess to the thing was observed. After abdominal oncotomy expanded to thigh and inguinal region, lavage, drainage operations, therapy with wide spectrum followed by aimed antibiotics general condition of the patient improved. On the 18 postoperative day a new feverish state manifested. Image forming examinations showed newer purulent collections in the abdominal cavity and on the thigh in addition to the previous abscesses. Because of this repeated exposure was necessary. After the second operation the patient recovered without complaint and further complications. Authors think the case worth attention because of the extensive and unusual localization of abscesses formed beside the relatively discrete abdominal complaints. PMID- 10697984 TI - [Genetic background of alcoholic liver disease]. AB - The genetic background of alcoholic behaviour and alcoholic liver diseases has long been the target of intense research. The current knowledge of this very interesting topic is herewith summarized with special emphasis on findings and facts which might have clinical significance including results of family studies, gene polymorphisms of enzyme families of alcohol metabolism, cytokines as well as HLA antigens. PMID- 10697985 TI - [Psychosomatic dermatologic symptoms]. AB - The explanations of the psychosomatic dermatological symptoms are based on the dysfunction of the emotional and autonomic nervous system. In the first instance the dermatological symptoms of psychic origin are examined by the general practitioner, than the dermatologist and finally after negative findings--the psychiatrist. Behind the non-improving, itching and scratched eczema sometimes was found either Ekbom-syndrome (often diagnosed as gerontological delusions of parasitosis) or therapy-resistant allergy (which conceals depression). The authors present condensed case--stories (about delusions of parasitosis and allergic syndromes) proving the excellent results achieved using citalopram and in the case of Ekbom-syndrome--risperidone therapy. PMID- 10697986 TI - [Diabetic ketoacidosis in childhood]. AB - Diabetic ketoacidosis is the most serious acute complication of insulin-dependent diabetes mellitus and the most frequent reason for hospital admission of diabetic children. The most frequent cause of death of these patients is also the diabetic ketoacidosis. The mortality rate of the disease has not changed since the seventies (1-2%). In this work, the data of 89 patients with diabetic ketoacidosis were analyzed. These patients were admitted to the 1. Department of Pediatrics of the Semmelweis University of Medicine between 1992-1997. The data (metabolic parameters, the causes of ketoacidosis and the length of hospital stay) of previously known diabetic children was compared with the data of previously unknown diabetic children. Our patients were divided in 2 groups: serious (n = 11), and mild-to-moderate (n = 48) acidosis. Their laboratory findings, their intravenous infusion-, and insulin demand and the length of their hospital stay were compared. The state of consciousness at their hospitalisation and the concomitant complications were also examined. Significant difference was found only in the duration of intravenous insulin administration (with the exception of pH and BE, of course). There was no relationship between the seriousness of the disease and the duration of hospital treatment. It is noteworthy that even the previously known diabetic children with the shortest hospitalization spent more than 7 days at the department. PMID- 10697987 TI - [Diffuse alveolar hemorrhage in systemic lupus erythematosus]. AB - Of the 120 systemic lupus erythematosus (SLE) patients treated by the authors, two have developed diffuse alveolar haemorrhage. The authors' objective is to present this rare, but severe manifestation. Patients 1 and 2 were 66- and 22 year old women, respectively. Both had SLE with multi-organ involvements including diffuse proliferative lupus nephritis. Before the diagnosis of the disease, both patients had experienced pneumonitis that resolved on corticosteroid treatment. Soon after the diagnosis, respiratory failure, haemoptoea and acute anaemia developed, accompanied by a rapid deterioration in the general condition. Chest radiographs revealed bilateral, diffuse, alveolar infiltrates. The pulmonary haemorrhage temporarily ceased in response to corticosteroid treatment, but both patients later died in consequence of active SLE and mixed bacterial and fungal sepsis. Post mortem examination demonstrated fibrosing alveolitis and alveolar bleeding in Patient 1, and an immune complex deposition-induced alveolocapillary inflammation with alveolar haemorrhage in Patient 2. Diffuse alveolar haemorrhage is a life-threatening manifestation of SLE. Its onset may be preceded by episodes of pneumonitis resolving on corticosteroid treatment. An active diagnostic workup, intensive observation and aggressive immunosuppressive treatment are the cornerstones of the management. The early detection and the active treatment of secondary infections are obligatory. The authors consider the most difficult challenge to be the optimum coordination of the above treatment modalities. PMID- 10697988 TI - [History of medicine on the Internet]. PMID- 10697989 TI - [Harvey Cushing and the Hungarians]. PMID- 10697990 TI - State medical board takes up challenge of scope of practice issues. PMID- 10697991 TI - Public-private partnership discussed as means to take lead in alleviating medical errors. PMID- 10697992 TI - OSHA revises bloodborne pathogens compliance directive. PMID- 10697993 TI - How much progress against cancer? PMID- 10697994 TI - Narrowing the margin of error. AB - The patient had been feeling remarkably well 24 hours post-operatively. She remained on intravenous feeding and medication, although she was ambulatory. That afternoon, she realized something was wrong. She was experiencing an overall, generalized weakness, difficulty breathing, and wheezing. In response to her breathing problems, she had used her Serevent (once in the morning) and albuterol inhalers (every four hours), but had achieved no relief. It wasn't until 10:00 p.m. that evening, when she questioned whether or not she had received her dose of prednisone that day which she had been on prior to surgery, that the problem was identified. Despite a detailed history of medication changes, which had been recorded and reported to the anesthesia department prior to surgery, the prednisone dose had been missed. PMID- 10697995 TI - Congestive heart failure project launched. PMID- 10697996 TI - Selecting the right computer system for your practice. PMID- 10697997 TI - Campaign tackles family violence epidemic. PMID- 10697998 TI - [Clinical symptoms of and diagnostic possibilities for hypophyseal adenoma in horses]. AB - Hirsutism was the most often observed symptom in horses with a pituitary gland tumor and was present in all 13 examined horses. Other symptoms were atrophy of muscles (n = 10), hyperhidrosis (n = 8), polyuria/polydipsia (n = 5), bulging or supraorbital fat (n = 3), polyphagia (n = 2), apathy (n = 2) and seizures (n = 2). Laminitis was the most frequently observed concurrent disease (n = 8). Hyperglycaemia (mean, 9.9 +/- 3.71 mmol/l) in 13 horses and glucosuria (median, 55 [range, 2-55] mmol/l) in 7 horses were the most important laboratory results. The dexamethasone suppression test was positive in all tested horses (n = 9) 20 h after administration of dexamethasone. The pituitary gland tumor was visible in every case underwent computed tomography (n = 7). From these results it can be concluded that a pituitary gland tumor can be suspected based on typical clinical signs. Hyperglycaemia and glucosuria support the preliminary diagnosis and a positive dexamethasone suppression test allows a final diagnosis. PMID- 10697999 TI - [Reassessment of the herd consultation in facilities with accumulated abortions in cattle]. AB - One hundred and ninety-eight dairy herds in which an abortion problem had been investigated by laboratory examination alone or in conjunction with a farm visit were reassessed via a questionnaire at least 1 year after the initial investigation. The overall response rate to the questionnaire was 78%, but more owners responded after a farm visit. One hundred and twenty-five questionnaires (63%) could be evaluated. Significantly more (p = 0.01) complete or tentative diagnoses were made in herds that were visited. The abortion problem had resolved in 88%, 79% and 88% of herds with a complete, a tentative and no diagnosis, respectively. In 65% of herds, the suggested control measures were followed by the attending veterinarian or the owner. Resolution of the abortion problem was more common in herds in which the proposed control measures were followed, in herds in which a complete diagnosis had been made, and in herds that had been visited during the initial investigation. The success in resolving the abortion problem, that could be attributed to the initial investigation, was 21% when a complete diagnosis had been made and 11% when only a tentative diagnosis had been made. Most (67%) herd owners would favour an accompanying farm visit, as opposed to laboratory examinations alone, if another abortion problem occurred, but only if a considerable portion of the cost could be deferred. PMID- 10698000 TI - [An unusual case from paternity testing in dogs]. AB - We are solving disputed paternities in purebred dogs using microsatellite analysis. An interesting case involved four puppies for which pedigrees were already issued by the Swiss Kennel Club. The investigation had to be carried out without a sample from the stated sire. Even though the relationship of this sire could not be determined conclusively, comparison of microsatellite alleles within this family, including an alleged half-brother of the four puppies, showed clearly that the breeder had made contradictory statements and that false recordings of matings had occurred. The pedigrees for these four puppies were withdrawn and sanctions against the breeder were imposed. PMID- 10698001 TI - A cluster of Ser/Thr residues at the C-terminus of mu-opioid receptor is required for G protein-coupled receptor kinase 2-mediated desensitization. AB - To investigate the functional role of G protein-coupled receptor kinases (GRK) in homologous desensitization of the mu-opioid receptor, human embryonic kidney (HEK) 293 cells, which express a significant level of GRK2, were stably transfected with the cDNA encoding the rat mu-opioid receptor. Wild-type mu opioid receptors developed homologous desensitization after 30 min pretreatment with DAMGO ([D-Ala2,N-methyl-Phe4,Gly-ol5]enkephalin), a specific mu-opioid receptor agonist. The ability of mu-opioid receptors to develop homologous desensitization was greatly impaired following the transfection of a cDNA fragment encoding the GRK2(495-689) polypeptide, which is believed to block Gbetagamma-mediated transduction events including the membrane translocation and activation of GRK2. The mu(Cdelta45) receptor, a deletion mutant that lacks 45 C terminal amino acids, failed to exhibit homologous desensitization after 30 min pretreatment of DAMGO. The mu(Cdelta41) receptor, which differs from the mu(Cdelta45) receptor by having four more Ser/Thr residues (Thr354Ser355Ser356Thr357), developed GRK2-mediated desensitization. These results suggest that homologous desensitization of rat mu-opioid receptors results from the activation of GRK2 and that a cluster of Ser/Thr residues (Thr354Ser355Ser356Thr357) at the intracellular carboxyl tail plays an important role in GRK2-mediated mu-opioid receptor desensitization. PMID- 10698002 TI - Acute and chronic activation of the mu-opioid receptor with the endogenous ligand endomorphin differentially regulates adenylyl cyclase isozymes. AB - While acute activation of G(i/o)-coupled receptors leads to inhibition of adenylyl cyclase, chronic activation of such receptors produces an increase in cyclic AMP accumulation, particularly evident upon withdrawal of the inhibitory agonist. This phenomenon has been referred to as adenylyl cyclase superactivation and is believed to play an important role in opiate addiction. Nine adenylyl cyclase isozymes have been recently identified and shown by us to be differentially regulated by acute and chronic inhibitory receptor activation. Using COS-7 cells cotransfected with various adenylyl cyclase isozymes, we examined here whether the endomorphins (the most recently discovered of the four classes of endogenous opioid peptides, and which interact selectively with the mu receptor) are able to induce inhibition/superactivation of representatives from the various adenylyl cyclase isozyme classes. Here, we show that adenylyl cyclase types I and V were inhibited by acute endomorphin application and superactivated upon chronic exposure, while adenylyl cyclase type II was stimulated by acute and "superinhibited" by chronic endomorphin exposure. These results show that the endomorphins are capable of regulating adenylyl cyclase activity and that different adenylyl cyclase isozymes respond differently to these endogenous ligands. PMID- 10698004 TI - Topographical evaluation of behavioural phenotype in a line of mice with targeted gene deletion of the D2 dopamine receptor. AB - The phenotype of spontaneous and dopamine D2-like agonist-induced behaviour was assessed topographically in a line of mice with targeted gene deletion of the D1 receptor. An ethologically-based, rapid time-sampling behavioural check-list technique was used to resolve and quantify all behaviours in the natural repertoire of the mouse. Relative to wildtypes [D2+/+], D2-null [D2-/-] mice evidenced over a 1 h period of initial exploration modest but significant reductions in locomotion, grooming, rearing free and rearing to wall; rearing seated, sniffing, sifting and stillness were not altered. Individual elements of behaviour habituated similarly over a 6 h period for both genotypes. The dose dependent induction of stereotyped sniffing and ponderous locomotion by the D2 like agonist RU 24213 (0.1-12.5 mg/kg) in wildtypes was essentially absent in D2 null mice. The ethogram of spontaneous behaviour in D2-null mice was characterised by only modest reductions in, and topographical shifts between, certain individual elements of behaviour. Essential abolition of D2-like agonist responsivity in D2-null mice vis-a-vis considerable preservation of spontaneous behavioural topography suggests compensatory processes subsequent to developmental absence of the D2 receptor that are able to sustain function under naturalistic, tonic conditions but not during phasic challenge. PMID- 10698003 TI - Dopamine-opioid interactions in the rat striatum: a modulatory role for dopamine D1 receptors in delta opioid receptor-mediated signal transduction. AB - Dopaminergic and opioidergic systems interact in the striatum to modulate locomotor and motivated behaviors. The present study investigated the molecular interactions of these two systems by determining the role of dopamine D1 and D2 receptors in the modulation of opioid receptor-mediated signal transduction. Male Fischer rats were injected daily for 10 days with either saline, the D1 receptor agonist SKF 82958, the D2 receptor agonist quinpirole, or both SKF 82958 and quinpirole. Administration of SKF 82958 alone or together with quinpirole attenuated the ability of the delta receptor agonist D-Pen2,D-Pen5-enkephalin (DPDPE) to inhibit adenylyl cyclase activity in the caudate putamen and nucleus accumbens. Quinpirole administration alone had no effect. The efficacy and potency of the mu opioid receptor agonist D-Ala2,N-Me-Phe4,Gly-ol5-enkephalin (DAMGO) to inhibit adenylyl cyclase activity was unaltered following administration of either dopamine receptor agonist. Administration of SKF 82958 had no affect on delta receptor binding, forskolin-stimulated adenylyl cyclase activity, or G protein/adenylyl cyclase coupling. However, the ability of DPDPE to stimulate binding of [35S]GTPgammaS to inhibitory G proteins was attenuated in animals that received SKF 82958. These results suggest that repeated activation of D1 receptors attenuates the functional coupling of delta opioid receptors with adenylyl cyclase due to decreased coupling between delta receptors and G proteins. PMID- 10698005 TI - Effects of chronic delta9-tetrahydrocannabinol on rat midbrain dopamine neurons: an electrophysiological assessment. AB - Delta-9-tetrahydrocannabinol (delta9-THC), the principal psychoactive ingredient in marijuana elicits a variety of physiological effects in animals and humans, and with repeated exposure tolerance develops to most of its effects. However, studies in humans found that tolerance did not occur to the pleasurable marijuana "high". Since ventral tegmental dopamine neurons play a pivotal role in drug reinforcement and reward, and possibly in the euphorigenic quality of marijuana, the present study sought to determine whether tolerance develops to the neurophysiological response elicited in these neurons by delta9-THC. Using single unit extracellular recordings the activity of midbrain ventral tegmental (VTA) and substantia nigra pars compacta (SNpc) dopamine neurons was measured in animals that had received twice-daily injections of 5 mg/kg delta9-THC for 14 days. Cannabinoid-induced changes in body temperature, locomotion, and catalepsy were also assessed in the same animals. After 2 weeks tolerance had developed to delta9-THC-induced hypothermia, catalepsy and reduction in locomotor activity. In naive animals and in animals that had received twice-daily vehicle injections for 14 days, delta9-THC increased VTA neuronal firing by 52% and 46%, respectively, while SNpc neurons showed increases of 23% and 30%, respectively. Following chronic cannabinoid treatment, however, SNpc neurons were significantly less responsive to delta9-THC with a maximum increase in rate of only 3%, while VTA neurons continued to show a robust increase in firing rate (+45%) when challenged with THC. These results suggest that VTA and SNpc dopamine neurons develop a differential response to delta9-THC following long-term cannabinoid exposure. This finding may be relevant to the observation that in humans tolerance occurs to many of marijuana's physiological effects but not to its euphorigenic actions. PMID- 10698006 TI - Methamphetamine-induced striatal dopamine neurotoxicity and cyclooxygenase-2 protein expression in BALB/c mice. AB - The expression of cyclooxygenase-2 (COX-2) and striatal dopamine (DA) depletion in BALB/cAnNcrj (BALB/c) mice following a neurotoxic dose of methamphetamine (METH) was investigated. METH-treatment (4 mg/kg x 4, 2 h intervals, s.c.) induced a significant hyperthermia and a persistent depletion of striatal DA levels 72 h after the treatment. COX-2, a marker of the cytotoxic effect of inflammation and oxidative stress and thiobarbituric acid (TBA) were significantly induced in the striatum 72 h after the METH-treatment, but not in the hippocampus. These results suggest that COX-2 may participate in METH-induced neurotoxicity in striatum. PMID- 10698007 TI - Prevention of cocaine-induced convulsions and lethality in mice: effectiveness of targeting different sites on the NMDA receptor complex. AB - N-methyl-D-aspartate (NMDA) receptors appear to be involved in the behavioral toxic effects of cocaine. Therefore, different classes of NMDA receptor antagonists were compared for their ability to attenuate cocaine-induced convulsions and lethality in male, Swiss Webster mice. The mice were pre-treated (i.p.) with vehicle or an antagonist from one of the following classes: NMDA/glycine site antagonist (7-chlorokynurenic acid, ACEA-1021, ACEA-1031, ACEA 1328, DCQX, R(+)-HA-966), competitive antagonist (CPP, D-AP7), channel blocker (MK-801, memantine), or allosteric modulator (ifenprodil, CP-101,606, Co 101022, haloperidol). After a 15 min pre-treatment period, the mice were administered a convulsive (60 mg/kg, i.p.) or lethal (125 mg/kg, i.p.) dose of cocaine, equivalent to the calculated ED/LD97 values. Pre-treatment with competitive or NMDA/glycine site antagonists dose-dependently attenuated cocaine-induced convulsions and lethality (P<0.05). Pre-treatment with channel blockers or allosteric modulators of the NMDA receptor protected against cocaine-induced convulsions (P<0.05), but were ineffective or less effective than the competitive and glycine site antagonists in preventing death. The glutamate release inhibitor riluzole failed to prevent both the convulsions and lethality induced by cocaine. Significantly, post-treatment with NMDA/glycine site antagonists (ACEA-1021, ACEA 1031, ACEA-1328) after a cocaine overdose prevented death in a significant number of animals. The data suggest that NMDA receptors are involved in the pathophysiology of a cocaine overdose. PMID- 10698008 TI - Intra-striatal infusion of D-amphetamine induces hydroxyl radical formation: inhibition by MK-801 pretreatment. AB - Recent evidence suggests that free radicals can be produced in the brain following systemic administration of repeated or high doses of D-amphetamine (AMPH). However, it has been proposed that the toxic effects of AMPH are mostly secondary to AMPH-induced hyperthermia, and agents that protect against AMPH neurotoxicity do so by blocking AMPH-induced hyperthermia or causing hypothermia. In this study, we examined the effects of AMPH on the formation of hydroxyl radicals (*OH) following its infusion into the rat striatum via a microdialysis probe. We found that intra-striatal perfusion of AMPH (10 microM) caused an increased formation of hydroxyl radicals but did not raise the core temperatures of the rats. Pretreatment with the NMDA antagonist MK-801 (0.5 mg/kg) attenuated hydroxyl radical production elicited by AMPH infusion, although core body temperatures in AMPH-treated rats were not significantly altered. Additionally, infusion of AMPH in the striatum increased extracellular dopamine concentration and this effect was potentiated by MK-801 pretreatment. Thus, these results demonstrate that direct infusion of AMPH in the striatum induces hydroxyl radical production without causing hyperthermia, and also imply that activation of glutamate NMDA receptors mediates, at least in part, AMPH-induced hydroxyl radical formation in the rat striatum. PMID- 10698009 TI - Regulation of gamma-aminobutyric acid (GABA) release in cerebral cortex in the gamma-hydroxybutyric acid (GHB) model of absence seizures in rat. AB - Gamma-hydroxybutyric acid (GHB) has the ability to induce absence seizures. The precise way in which GHB causes seizures remains unclear, but GABA(B)- and/or GHB mediated presynaptic mechanisms within thalamocortical circuitry may play a role. In the present study, we determined the basal and K+-evoked release of GABA and glutamate in the superficial laminae of frontal cortex during GHB-induced absence seizures. Our data indicate that both the basal and K+-evoked release of GABA were significantly decreased in laminae I-III of frontal cortex at the onset of GHB-induced absence seizures. The appearance and disappearance of the observed changes in basal and K+-evoked extracellular levels of GABA correlated with the onset and offset of absence seizures. In contrast, neither the basal nor the K+ evoked release of glutamate was altered in superficial laminae of cerebral cortex at any time during the absence seizures. Intracortical perfusion of the GABA(B) receptor antagonists, CGP 35348 and phaclofen as well as the GHB receptor antagonist, NCS 382 attenuated GHB-mediated changes in the basal and K+-evoked release of GABA. These data suggest that GHB induces a selective decrease in the basal and depolarization-induced release of GABA in cerebral cortex, and further, that this action of GHB may play a role in the mechanism by which GHB induces absence seizures. PMID- 10698010 TI - Brain allopregnanolone regulates the potency of the GABA(A) receptor agonist muscimol. AB - Allopregnanolone (ALLO), a potent positive-allosteric modulator of the action of GABA at GABA(A) receptors, is synthesized in the brain from progesterone by the sequential action of two enzymes: 5alpha-reductase and 3alpha hydroxysteroidoxidoreductase. The concentration of ALLO in various parts of the mouse brain varies substantially, from 15 pmol/g in the olfactory bulb, to approximately 6 pmol/g in the frontoparietal cortex, and 2.7 pmol/g in the cerebellum. The systemic administration of 48 micromol/kg of the Type I and Type II 5alpha-reductase inhibitor, (17beta)-17-[bis (1-methylethyl) amino carbonyl)] androsta-3, 5-diene-3-carboxylic acid (SKF 105,111), reduced brain ALLO content by 80-90% in 30 min; the rate constant (k) of ALLO decrease in each brain area can be utilized to establish the rate of ALLO biosynthesis, which is higher in the olfactory bulb (62 pmol/g/h) than in the frontoparietal cortex (24 pmol/g/h) or cerebellum (11 pmol/g/h). The duration of the righting reflex loss elicited by the potent GABA(A) receptor agonist muscimol was reduced in SKF 105,111-treated ALLO-depleted mice. SKF 105,111 treatment had no effect on muscimol metabolism or on brain levels of pregnenolone and progesterone; however, the brain levels of 5alpha-DHP, the precursor of ALLO, were also decreased. Administration of ALLO at a dose of 15 micromol/kg i.p. by itself did not alter the muscimol-induced loss of the righting reflex; but it completely blocked the effect of SKF 105,111. To elucidate the possible molecular mechanism by which a decrease of brain ALLO content can shorten the duration of the righting reflex loss elicited by muscimol, we patch-clamped neocortical pyramidal neurons of mice pretreated with SKF 105,111 or vehicle, and studied the efficiency of muscimol in eliciting Cl- currents. The current amplitude was significantly smaller in neurons from SKF 105,111-treated mice, especially at lower doses (0.1-1 microM) of muscimol, and the muscimol dose-response (0.1-10 microM) relationship displayed cooperativity (nH=1.4). These data suggest that ALLO synthesized in brain plays an important physiological permissive role in the modulation of GABA-gated Cl- channel function. PMID- 10698011 TI - GABA(B) receptor antagonists elevate both mRNA and protein levels of the neurotrophins nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) but not neurotrophin-3 (NT-3) in brain and spinal cord of rats. AB - In this study we show that single, physiologically-active and non-convulsive doses of the three GABA(B) receptor antagonists CGP 36742, CGP 56433A and CGP 56999A increase NGF and BDNF mRNA levels by 200-400% and protein levels by 200 250% in rat neocortex, hippocampus as well as spinal cord. In all areas examined the increase in NGF protein preceded that of BDNF. Peak levels of both neurotrophins are transient and occur between 24 and 72 h, depending on the region. In contrast, NT-3 protein concentrations in the neocortex and hippocampus were decreased significantly to 50% of control values within 48-96 h. The decrease in the spinal cord was less than 30% and did not reach significant levels. These data clearly demonstrate that GABA(B) receptor antagonists induce a specific neurotrophin expression in the central nervous system at physiologically relevant doses, as opposed to the extreme conditions of seizure paradigms. The results are in line with the concept that neuronal neurotrophin synthesis and release in brain are controlled by afferent nerve activity. GABA(B) receptor antagonists could therefore be a valuable new approach to selectively increase endogenous neurotrophin levels in the central nervous system. PMID- 10698012 TI - Serotonergic regulation of mRNA expression of Arc, an immediate early gene selectively localized at neuronal dendrites. AB - Arc (activity regulated, cytoskeleton associated protein) is an effector immediate early gene that is selectively localized in the neuronal dendrites. Elevation of brain 5-HT by the combined administration of the monoamine oxidase inhibitor, tranylcypromine (TCP, 5 mg/kg, i.p.), and the 5-HT precursor L tryptophan (L-TP, 100 mg/kg, i.p.), increased Arc mRNA abundance in the cingulate, orbital, frontal and parietal cortices as well as in the striatum but a reduction was observed in the CA1 region of the hippocampus. The 5-HT releasing agent p-chloroamphetamine (PCA, 5 mg/kg, s.c.) also increased Arc mRNA in the cortical and striatal areas. Depleting brain 5-HT with the tryptophan hydroxylase inhibitor, p-chlorophenylalanine (pCPA, 300 mg/kg, i.p. for two days), on the other hand, significantly attenuated the increase in Arc mRNA induced by tranylcypromine and L-tryptophan (TCP/L-TP). Pretreatment with the 5-HT2 receptor antagonist ketanserin (2 mg/kg, i.p.) significantly attenuated the effect of TCP/L-TP in the cortex but only partially in striatum and did not affect the reduction in the CA1 region. The 5-HT2 agonist DOI (0.2, 1 and 2 mg/kg, i.p.) dose-dependently increased Arc mRNA abundance in cortical areas with a pattern similar to that of TCP/L-TP and PCA. DOI, however, had much weaker effects on Arc mRNA in the striatum and did not have any significant effect in the CA1, CA3 and the dentate gyms (DG) of the hippocampus. Pretreatment with ketanserin completely blocked the effect of DOI on Arc expression. These data suggest that Arc mRNA expression can be induced in the cortex by increases in extracellular 5-HT and that 5-HT2 receptors play a major part in mediating such effects. Additional 5-HT receptors as well as other neurotransmitters may also be involved, particularly in the striatum and in CA1 subfield of the hippocampus. Overall, our data suggest that expression of Arc mRNA is highly responsive to changes in brain 5-HT functions, and may provide a sensitive marker of postsynaptic 5-HT2(2A and 2C) receptor functions. PMID- 10698013 TI - Activation of 5-HT2A receptors impairs response control of rats in a five-choice serial reaction time task. AB - The present experiments investigated the effects of agents acting at serotonin (5 HT)-2 receptors on the performance of rats in a choice serial reaction time (5 CSRT) task in order to examine the role of 5-HT2 receptors in the modulation of attention and response control. The results indicate that DOI, [(+/-)-1-(2,5 dimethoxy-4-iodophenyl)-2-aminopropane hydrochloride; 0.05, 0.1 and 0.2 mg/kg, subcutaneously], a 5-HT(2A/2C) agonist, slightly impaired the choice accuracy of the well performing rats and markedly increased their premature responding. DOI (0.05 and 0.1 mg/kg) had no effect on the latency to collect earned food pellets or to respond correctly, indicating that these lower doses of DOI did not reduce motivation for the food reward in this task. The selective effect of a low dose of DOI (0.1 mg/kg) on premature responding was completely blocked by ketanserin (0.2 mg/kg), a 5-HT2A antagonist, and ritanserin (0.3 mg/kg), a 5-HT(2A/2C) antagonist, but only partially blocked by a high dose of SER082 (1.0 mg/kg), a 5 HT2C antagonist. In contrast to DOI, mCPP, [1-(3-phenyl)piperazine; 0.05 and 0.15 mg/kg], a 5-HT2C agonist, had no effect on choice accuracy or premature responding, but it reduced behavioral activity and/or arousal as indicated by the decreased number of trials completed and increased the probability of omissions. SER082 (1.0 mg/kg) blocked the effects of mCPP on performance. These data suggest that the overactivation of 5-HT2A receptors impairs response control in a 5-CSRT task, whereas the overactivation of 5-HT2C receptors can affect behavioral activity and/or arousal state of the animals for this food rewarded task. PMID- 10698014 TI - P2Y receptors contribute to ATP-induced increases in intracellular calcium in differentiated but not undifferentiated PC12 cells. AB - ATP-induced Ca2+ transients were examined in individual PC12 cells of a well defined clone, before and after treatment with nerve growth factor (NGF) to induce a neurone-like phenotype. Using reverse transcriptase PCR these cells were found to express mRNA for several P2 receptors. In undifferentiated cells the ATP induced Ca2+ response was entirely dependent on Ca2+ influx, could not be mimicked by UTP, alpha,beta-methylene ATP or dibenzoyl ATP or be blocked by pyridoxalphosphate-6-azophenyl-2',4'-disulphonic acid (PPADS). ATP had no significant effect on levels of cyclic AMP or inositol 1,4,5-trisphosphate (InsP3). These results suggest that in undifferentiated PC12 cells ATP mainly acts on a P2X receptor, possibly the P2X4 subtype. After treatment with NGF for 7 days the ATP response was increased and partially sensitive to PPADS. A component of the ATP-induced Ca2+ increase was due to mobilisation of intracellular Ca2+ stores and another to capacitative Ca2+ entry. UTP caused an increase in intracellular Ca2+, and InsP3 formation could be stimulated by ATP and UTP. ATP also caused a small increase in cyclic AMP, but this was abolished in the presence of indomethacin. Thus, after NGF treatment ATP acts partially via a P2Y receptor, possibly the P2Y2 subtype. PMID- 10698015 TI - Gender differences in the effect of rivastigmine on brain cholinesterase activity and cognitive function in rats. AB - This study compared the effect of rivastigmine on cholinesterase (ChE) activity in different brain regions, heart, skeletal muscle and plasma and on the cognitive impairment induced by scopolamine (0.5 mg/kg) in male and female rats. Rats were injected s.c. with saline or rivastigmine (0.75-2.5 mg/kg) or physostigmine (0.05 mg/kg) and killed 30-120 min later. Amelioration of scopolamine-induced memory deficits by rivastigmine (0.75 mg/kg) was assessed in the Morris water maze. There were no gender differences in spatial memory or basal ChE activity in the brain or other organs. Rivastigmine (0.75 and 1.5 mg/kg) and physostigmine (0.05 mg/kg) caused significantly greater ChE inhibition in females than in males (P<0.01) in the cerebral cortex, hippocampus and striatum, but not in the periphery 30 and 60 min after injection. Rivastigmine was also more effective in antagonising the scopolamine-induced spatial memory impairment in female than in male rats. Ovariectomy did not affect the degree of enzyme inhibition by rivastigmine in any brain area. Orchidectomy completely abolished the difference in enzyme inhibition. It is concluded that a testicular hormone suppresses the effect of rivastigmine, by reducing the amount of drug reaching the brain or its interaction with ChE. PMID- 10698016 TI - Melatonin reduces oxidative neurotoxicity due to quinolinic acid: in vitro and in vivo findings. AB - The in vivo and in vitro effects of melatonin on quinolinic acid-induced oxidative damage in rat brain were determined. The concentrations of malonaldehyde and 4-hydroxyalkenals were assayed as an index of oxidatively damaged lipid. In in vitro experiments, the increase in malonaldehyde and 4 hydroxyalkenals concentrations induced by quinolinic acid were concentration dependent and time-dependent. The accumulation of products of lipid peroxidation induced by quinolinic acid were very significantly reduced by melatonin in a concentration-dependent manner. Additionally, at the highest concentrations of melatonin used in quinolinic acid treated homogenates, it reduced the levels of oxidatively damaged lipid products below those measured in control homogenates (no quinolinic acid or melatonin). When quinolinic acid (200 mg/kg) was intraperitonally injected into 11-day-old rats, lipid peroxidation in the brain was significantly increased 24 hours later compared to levels in control rats. When melatonin (10 mg/kg) was injected i.p. 30 min before and 4 and 20 hours after the administration of quinolinic acid, the increased lipid peroxidation induced by quinolinic acid was significantly reduced. Likewise, neurobehavioral signs associated with quinolinate administration were attenuated by melatonin. These results show that both in vitro and in vivo pharmacological levels of melatonin confer protection against quinolinic acid-induced oxidative toxicity in the brain. The findings also indicate that melatonin may be pharmacologically useful in combatting quinolinic neurotoxicity which is associated with several acute and chronic neurodegenerative neurological diseases. PMID- 10698017 TI - Combination of low dose ethanol and caffeine protects brain from damage produced by focal ischemia in rats. AB - Caffeine and ethanol are two commonly overused psychoactive dietary components. The purpose of this study was to assess the effects of acute, chronic, oral (p.o.) and intravenous (i.v.) caffeine, ethanol and their combination on infarct volume following focal ischemia in rats. Rats received treatment either p.o. 3 h and 1 h before, or by i.v. infusion for 2.5 h beginning 30-180 min after, ischemia. There were six acute treatment groups. (1) oral dH2O (control); (2) oral caffeine (10 mg/kg); (3) oral ethanol (0.65 g/kg total); (4) oral ethanol plus caffeine; (5) intravenous saline; and (6) intravenous ethanol (0.65 g/kg) plus caffeine (10 mg/kg) in saline. A 7th group received oral ethanol plus caffeine for three weeks prior to ischemia. After 3 h of left MCA/CCA occlusion and 24 h reperfusion, infarct volume was determined. Control animal infarct volume was 102.4+/-42.0 mm3. Oral caffeine alone had no effect (122.4+/-30.2 mm3). Oral ethanol alone exacerbated infarct volume (177.2+/-27.8 mm3). Oral caffeine plus ethanol almost entirely eliminated the damage (17.89+/-10.41 mm3). When i.v. treatment with ethanol plus caffeine was initiated at 30, 60, 90 and 120 minutes post-ischemia the infarct volume was reduced by 71.7%, 49.8%, 64.8% and 47.1%, respectively. Chronic daily oral ethanol plus caffeine prior to ischemia eliminated the neuroprotection seen with acute treatment. These studies indicate that ethanol, which by itself aggravates cerebral ischemia, and caffeine, when combined together immediately before or for 2 h after focal stroke, reduces ischemic damage. PMID- 10698018 TI - Signaling regulation for synergistic effects of substance P and insulin-like growth factor-1 or epidermal growth factor on corneal epithelial migration. AB - PURPOSE: In a previous report we showed that substance P (SP) and insulin-like growth factor-1 (IGF-1) or epidermal growth factor (EGF) synergistically stimulate corneal epithelial migration. In this study, we used an organ culture system of rabbit cornea to identify which signal transduction system affects corneal epithelial migration. METHODS: Rabbit corneal blocks were cultured in TC 199 culture medium containing various reagents for 24 hours. After the end of cultivation, the length of the path of epithelial migration was measured. RESULTS: Acting alone, protein kinase C (PKC) inhibitors, calphostin C and H-7, each reduced the length of epithelial migration. Tyrosine kinase (TK) inhibitors, genistein and herbimycin A, also acted individually to inhibit epithelial migration. The synergistic stimulatory effects of SP and IGF-1 on corneal epithelial migration were eliminated when PKC inhibitors or TK inhibitors were added. The synergistic effect of SP and EGF was eliminated by TK inhibitors, but only partly suppressed by PKC inhibitors. CONCLUSIONS: These results suggest that the synergistic effect of SP and EGF might require a TK pathway, and that the synergistic effect of SP and IGF-1 might require both PKC and TK pathways. PMID- 10698019 TI - Electron microscopic study on the development of precapsular layer in eyes with exfoliation syndrome. AB - PURPOSE: To search for a pathogenic mechanism for the formation of the precapsular layer on the anterior lens surface in pre-exfoliative eyes. METHODS: We examined anterior lens capsules obtained during surgery from 38 patients (control, 16; exfoliation suspect, 10; exfoliation, 12) by transmission electron microscopy. RESULTS: A precapsular layer was found in 5 of 16 controls and 7 of 10 exfoliation suspects. It was composed mainly of microfibrils 5-8 nm in diameter. Degenerated zonular fragments were occasionally found interspersed in, and sometimes merged with microfibrils of the precapsular layer. CONCLUSION: Zonular fibers might contribute to the formation of the precapsular layer in pre exfoliation stages. PMID- 10698020 TI - Disturbance of electrolyte balance in vitreous of chicks with form-deprivation myopia. AB - PURPOSE: To investigate the changes in the electrolyte and protein concentrations in the vitreous of 3-week-old chicks with form-deprivation myopia (FDM). METHODS: FDM was induced in 2-day-old male white leghorn chicks by covering the left eye with a translucent plastic goggle and leaving the right eye uncovered to serve as control. After 19 days the animals were euthanized, and the axial dimensions of the eyes were measured with a caliper in an unfixed condition. The liquid vitreous and aqueous humor were removed by paracentesis, and blood was collected from the jugular vein. Sodium, potassium, and chloride concentrations were determined using ion-selective electrodes. Calcium and phosphate concentrations were determined by colorimetric assays using orthocresol phthalein complexone and bacterial xanthine oxidase, respectively. RESULTS: The concentrations of potassium and phosphate were decreased, whereas chloride concentration was increased in the vitreous of the FDM eyes (P < .01). Sodium and calcium concentrations were similar to those in the control eyes. No significant changes in the concentration of electrolytes were observed in the aqueous humor. No significant differences were found in the protein concentrations in the liquid vitreous, gel vitreous, and aqueous humor. CONCLUSIONS: Form-deprivation induced a significant increase of the volume of the liquid vitreous in the eye of the FDM chick. The increased liquid vitreous of the myopic eye was accompanied by an alteration of the electrolyte balance, by a mechanism that has not yet been clarified. PMID- 10698021 TI - Amplitude decrease of photopic ERG b-wave at higher stimulus intensities in humans. AB - PURPOSE: The b-wave of the human photopic electroretinogram (ERG) elicited by a short-flash increases in amplitude with increasing stimulus intensities at lower stimulus levels, but then decreases at higher stimulus intensities. The purpose of the present study was to explore this phenomenon in more detail, using short- and long-flash stimuli. METHODS: The intensity-response functions of the b-wave elicited by short- and long-flashes were compared from threshold to higher stimulus intensities in 5 normal subjects. Short- and long-flash ERGs were elicited under rod-saturating background levels using white light-emitting diodes built into a contact lens electrode. RESULTS: Whereas the amplitude of the short flash b-wave decreased at higher intensities, the amplitude of the long-flash ERG b-wave did not decrease but plateaued. The long-flash ERG d-wave or OFF-response decreased at higher stimulus levels as did the short-flash elicited b-wave. CONCLUSIONS: Because it is widely accepted that the b-wave and the OFF-response d wave interact to produce a single positive response, our results suggest that the decrease in the b-wave amplitude at high stimulus intensity is caused by the decrease of the d-wave at the higher stimulus intensities. PMID- 10698022 TI - Corneal changes in spondyloepiphyseal dysplasia tarda. AB - BACKGROUND: A new type of corneal opacity with prominent corneal nerve fibers as an ocular complication of spondyloepiphyseal dysplasia tarda (SEDT). CASE: A 58 year-old woman, diagnosed with SEDT at 5 years of age, underwent a complete ophthalmological examination. OBSERVATIONS: The patient had no complaints and no history of eye disease. No relatives were reported to have suffered from SEDT. Slit-lamp examination disclosed a diffuse opacity in the central cornea in both eyes, which was localized in the middle to deep stroma. Dot opacities in the central and paracentral cornea were located in the middle of the stroma in both eyes. Optically clear regions were observed in the peripheral cornea of both eyes. More interestingly, corneal nerve fibers were visible passing from the limbus to the central cornea in both eyes. RESULTS: The etiology of the corneal opacities of this patient with SEDT is unknown. However, collagen and proteoglycan abnormalities in the skin of patients with SEDT have been reported. Therefore, such abnormalities may also be present in the cornea and these alterations may lead to corneal complications. PMID- 10698023 TI - Efficacy and safety of latanoprost eye drops for glaucoma treatment: a 1-year study in Japan. AB - PURPOSE: To evaluate the intraocular pressure (IOP)-lowering effect and safety of latanoprost, a prostaglandin analogue, in patients with primary open-angle glaucoma or ocular hypertension. METHOD: One hundred and twenty-four Japanese patients with primary open-angle glaucoma or ocular hypertension were enrolled in this open-labeled study and were treated with 0.005% latanoprost once daily for 1 year. RESULTS: At all follow-up visits there was a significant (P < .001) reduction in IOP compared with the baseline value. After 1 year, the IOP was reduced by 5.4 +/- 2.9 (mean +/- SD) mm Hg from a baseline value of 23.5 +/- 2.2 mm Hg. No evidence of an upward drift in the IOP was observed during the treatment period. The most frequently reported adverse ocular events were mild conjunctival hyperemia and iris pigmentation. Very few adverse systemic events were observed. CONCLUSIONS: Latanoprost eye drops showed a marked and stable IOP lowering effect during the 1-year treatment period. Furthermore, latanoprost was well-tolerated and should be a valuable contribution to the management of glaucoma. PMID- 10698024 TI - Association between watershed zone and visual field defect in normal tension glaucoma. AB - PURPOSE: To evaluate the association between the watershed zone and glaucomatous optic nerve head (ONH) damage. METHODS: We performed indocyanine green fluorescence angiography with a scanning laser ophthalmoscope in 54 eyes of 27 patients with normal tension glaucoma (NTG). RESULTS: We identified 7 eyes of 8 patients (14.8%) with a watershed zone not including the ONH, 32 eyes of 20 patients (59.3%) with the watershed zone partially including the ONH, and 10 eyes of 14 NTG patients (25.9%) with the watershed zone including the ONH. Of the 27 NTG patients, 10 patients (37.0%) had different types in each eye. CONCLUSIONS: In these patients, the mean deviation of visual field indices was greater in the eye with the watershed zone, which included a larger part of the ONH than in the contralateral eye. Conversely, the eye with the greater mean deviation had a watershed zone that included a larger part of the ONH. The location of the watershed zone appeared to influence the progression of the visual field defect. PMID- 10698025 TI - Angle recess area decreases with age in normal Japanese. AB - PURPOSE: To investigate prospectively the relationships between anterior chamber angle configuration and refractive error, axial length, age, and body height in the Japanese. METHODS: We studied 65 eyes of 65 subjects (30 men, 35 women) who were either patients at the Ophthalmology Department of Mie University or volunteers. The 65 subjects underwent a complete eye examination, A-scan biometry, and ultrasound biomicroscopy. Images were exported to an IBM-compatible personal computer in PCX format. The angle recess area (ARA) was measured using a software program of our own design. RESULTS: The ARA decreased with age in all quadrants of all eyes. In older individuals, the angle in the superior quadrant was significantly narrower than in the other quadrants. The ARA correlated directly with anterior chamber depth (P < .001), axial length (P < .001), and body height (P = .003), and inversely with age (P < .001) and refractive error (P = .003) in pairwise analysis. Multivariate analysis revealed a significant association between ARA and anterior chamber depth (P < .001), axial length (P = .016), and younger age (P = .043). CONCLUSIONS: The anterior chamber area narrows with age, especially in the superior quadrant. Narrowing of the angle in Japanese is associated with older age, shorter axial length, and shallower anterior chamber depth. We hypothesize that because of the increasing prevalence of axial myopia in younger Japanese, angle-closure glaucoma could become less common in Japan in the future. PMID- 10698026 TI - Cataract development after trabeculectomy with mitomycin C: a 1-year study. AB - PURPOSE: To measure cataract changes quantitatively in the crystalline lens after trabeculectomy with mitomycin C and to identify associated clinical factors. METHODS: Forty-one consecutive trabeculectomy patients (41 eyes) were enrolled in this prospective study. The enrollment criteria were: phakic eye, no history of intraocular surgeries or inflammation, and no corneal opacification. The transparency of the lens was measured and analyzed by an anterior segment analysis system preoperatively and 1, 3, and 12 months postoperatively. Cataract development was correlated with clinical factors using a multivariate regression analysis. RESULTS: During the first postoperative month, changes occurred primarily in the anterior segment of the lens, but later, deeper lenticular areas also started to show an increase in light scattering intensity. Multivariate regression analyses demonstrated that some clinical factors were associated with cataract development. CONCLUSIONS: Slight cataractous changes develop after trabeculectomy with mitomycin C as early as 1 month postoperatively, and they gradually increase in extent and in intensity during the next 11 months. PMID- 10698027 TI - Comparison of diclofenac and fluorometholone in preventing cystoid macular edema after small incision cataract surgery: a multicentered prospective trial. AB - PURPOSE: To compare a nonsteroidal topical solution (0.1% diclofenac) to a steroidal topical solution (0.1% fluorometholone) in preventing cystoid macular edema (CME) and disruption of the blood-aqueous barrier. METHODS: A multicentered, prospective clinical trial was performed on eyes undergoing phacoemulsification followed by implantation of a foldable acrylic intraocular lens by the envelope technique. The presence and degree of cystoid macula edema (CME) was determined by fluorescein angiography. A breakdown of the blood-aqueous barrier was determined by laser flare-cell photometry. RESULTS: Five weeks after surgery, CME was present in 3 of 53 eyes (5.7%) receiving diclofenac and in 29 of 53 eyes (54.7%) receiving fluorometholone. This difference was statistically significant (P < .001). The amount of flare in the anterior chamber at 3 days, 1, 2, 5, and 8 weeks after surgery was also significantly lower (P < .01-P < .001) in the diclofenac group. The degree of flare at 3 days, 1, 2, 5, and 8 weeks after surgery was significantly higher in eyes with CME (P < .001). CONCLUSIONS: These findings suggest that diclofenac effectively prevents CME following cataract surgery and that CME is closely related to the breakdown of the blood aqueous barrier. PMID- 10698028 TI - Mobility of hydroxyapatite orbital implant covered with autologous sclera. AB - PURPOSE: To evaluate the mobility of a hydroxyapatite implant covered with autologous sclera from the enucleated globes of patients with severe atrophy of the eyeball or eyelid retraction. METHODS: This implant was used in seven patients with phthisis bulbi. We measured the movement of the implant by photographic analysis of the anterior orbit and by using a strain gauge. RESULTS: At 1-3 years after surgery, neither infection nor prolapse of the implant had occurred in any of the patients. The implant remained stable in the orbit; the extraocular muscles sutured to the sclera of the implant were functioning satisfactorily, and the implants showed great conjugate mobility to the ocular movement of the healthy eye. On photographic analysis of the anterior orbit, the adducting efficiency of the implant was 92.6 +/- 3.3%; the abducting efficiency was 85.9 +/- 5.4%; the supraducting efficiency was 84.9 +/- 5.6%, and the infraducting efficiency was 90.9 +/- 3.9%. The mean tugging weight, as determined using a strain gauge, was 344.2 +/- 29.2 g for adduction, and 327.6 +/- 33.4 g for abduction. These values corresponded to 90.4 +/- 4.4% and 89.5 +/- 5.3% of the respective movements of the healthy eye. CONCLUSIONS: Fitting an artificial eye to the peg of this implant did not greatly impair the movement of the implant, and its mobility was greater than that of the artificial eye of the controls in which a semi-integrated magnetic implant, previously available, had been used. This new technique makes it possible to wear an artificial eye earlier than with other prosthetic procedures. PMID- 10698029 TI - Present status of ophthalmological care for diabetic patients in Japan. AB - PURPOSE: To investigate the present status of diabetic care provided by ophthalmologists working in hospitals and private clinics in Japan. METHODS: Questionnaires were mailed to 315 ophthalmologists. There was a return rate of 73%. RESULTS: Problems identified in the clinics were: (1) many diabetic patients who complain chiefly of ophthalmological symptoms voluntarily request their first ophthalmological examination; (2) appropriate cooperation between ophthalmologists and physicians is not established; (3) assessing the level of blood glucose control is difficult; and (4) scheduling of appointments is inadequate. Moreover, (1) inadequate handling of patients who discontinue their ophthalmological examinations, and (2) the lack of an established patient education program were seen as problems common to both hospitals and clinics. CONCLUSIONS: For the resolution of these problems, comprehensive countermeasures should be developed urgently by medical associations, medical administrators, and other relevant entities. PMID- 10698030 TI - The relationship between visual disability and visual scores in patients with retinitis pigmentosa. AB - PURPOSE: To evaluate the relationship between visual disability and visual scores in patients with retinitis pigmentosa. METHODS: The relationship between visual disability and visual scores (visual acuity and visual field) was investigated in 93 patients with retinitis pigmentosa. The visual disability of each patient was evaluated using a questionnaire (a total of 35 questions, in 7 sections, regarding daily life). The reproducibility and validity of the data obtained by the questionnaire had been established by a similar investigation in glaucoma patients. Mean (+/-SD) age of patients was 52.6 +/- 15.1 years, the mean visual acuity of the logarithm of the minimum angle of resolution (log10MAR) was 0.5 +/- 0.4, and the mean deviation of visual field with the Humphrey Field Analyzer program 30-2 was -22.0 +/- 10.9 dB. RESULTS: The visual acuity of log10MAR in the better eye (r = 0.66 to 0.81) and the mean sensitivity within the central 10 degrees of the visual field (r = -0.76 to -0.62) had a definite relationship to the visual disability index of each section and their sum (P < .0001). This relationship was also confirmed in multiple regression analysis, which showed a high correlation coefficient (R2 = 0.57 to 0.77, P < .0001). CONCLUSIONS: The retinal sensitivity within the central 10 degrees and the visual acuity of log10MAR in the better eye had a significant influence on a patient's daily life. We suggest that in patients with retinitis pigmentosa, visual disability in daily life can be precisely evaluated with the retinal sensitivity within the central 10 degrees and the visual acuity in the logarithm of the minimum angle of resolution in the better eye. PMID- 10698031 TI - A possible mechanism of X-ray-induced injury in rat lens. AB - PURPOSE: X-ray and other radiation can cause cataract, but the pathogenic mechanism is largely unknown. The aim of this study was to investigate the accumulation of iron in the x-ray-exposed rat lens and its relationship to lens injury. METHODS: Fifty male Wistar rats were divided randomly into five groups of 10. Groups 2 and 4 rats were sham-exposed, groups 3 and 5 were x-ray-treated, and group 1 served as control. X-ray radiation and sham exposure were performed in a similar manner. After 10 and 30 days of exposure, a lens from each rat in groups 2 and 3, and 3 and 5, respectively, were analyzed by flame atomic absorption technique for the assessment of metal content. RESULTS: Significantly decreased zinc and increased iron and calcium concentrations were detected in the lens samples of groups 3 and 5 compared with groups 2 and 4 and controls. Similar results were obtained comparing groups 5 and 3. CONCLUSIONS: We propose that x ray exposure may cause toxic cell injury of the rat lens via Fenton metals catalyzed damage. Initial lens membrane damage in the radiolytic phase may permit the access of iron resulting in lens damage. PMID- 10698032 TI - Role of the vitreous in age-related macular degeneration. AB - PURPOSE: To investigate the relationship between posterior vitreous detachment (PVD) and age-related macular degeneration (AMD). METHODS: The condition of the vitreous was examined by slit-lamp funduscopy and ultrasonography in 93 eyes of 50 patients with AMD (exudative or dry) and 100 eyes of 50 controls. RESULTS: There was complete PVD in 31 of the 93 eyes (33.3%) of 50 patients with AMD and the posterior vitreous was attached in 62 of these eyes (66.6%). In the control group, in 50 eyes (50%) of 50 subjects there was posterior vitreous detachment. The prevalence of PVD in eyes with macular degeneration was significantly lower (P < .05). There was no statistically significant difference between the exudative and the nonexudative groups in respect to PVD. CONCLUSIONS: PVD may have a protective role against the development of AMD. Chronic vitreomacular traction and/or continuous exposure to free radicals and cytokines may possibly be one of the causes of AMD in eyes with attached vitreous. PMID- 10698033 TI - The influence of different types of antibodies on in vitro fertilization results. AB - PROBLEM: The influence of anti-sperm (ASA), anti-phospholipid (APA), and antizonal (AZA) antibodies on in vitro fertilization (IVF) results and the need for intracytoplasmic sperm injection (ICSI) were assessed. METHOD OF STUDY: Forty four couples with infertility of immunologic origin were investigated. ASA in serum and ovulatory mucus were studied by a tray agglutination test (TAT) and indirect mixed anti-globulin reaction test (MAR) test, AZA were studied by passive hemagglutination and commercial enzyme-linked immunosorbent assay (ELISA; BioGen, Germany), and APA were tested by ELISAs in immunoglobulin isotypes IgG and IgM. RESULTS: Because of failed or very low fertilization after standard IVF in the previous cycle, ICSI had to be used in five out of 15 cases with ASA (33.3%), in 16 out of 18 couples with AZA (89.4%), and in only one case if APA were present (9%). Clinical pregnancy rate was 60% in cases with ASA, 38.5% with AZA, and 27.3% per embryo transfer (ET) if APA were detected. CONCLUSIONS: Immunologic infertility can be treated by IVF with very good results. The most important group are women with AZA, in whom IVF ICSI without any delay is recommended. PMID- 10698034 TI - Interleukin-1 levels in the supernatant of conditioned media of embryos grown in autologous endometrial coculture: correlation with outcome after in vitro fertilization. AB - PROBLEM: To determine if interleukin (IL)-1 produced by autologous endometrial coculture (AECC) was associated with outcome in patients with a history of multiple in vitro fertilization (IVF) failures. METHOD OF STUDY: The conditioned media (CM) from AECC cells exposed or non-exposed to human embryos was analyzed for IL-1. RESULTS: Embryos grown on AECC demonstrated a significant improvement in number of blastomeres and fragmentation (frag) when compared to embryos grown in conventional media without ECC (6.4 +/- 1.3 vs. 5.5 +/- 1.2 blastomeres and 14.6 +/- 9.3%, vs. 18.4 +/- 9.8% frag; P< 0.008 and 0.003, respectively). When IL 1alpha and IL-1beta were undetectable in the CM, the embryos grown in ECC were of improved quality as compared to the embryos grown only in conventional media. Conversely, IL-1ra levels in the CM were positively associated with embryo quality. Exposure or non-exposure to an embryo did not result in differing levels of IL-1alpha, IL-1beta, or IL-1ra in the CM. IL-1beta levels were negatively associated with clinical pregnancy outcome (3.3 pg/mL (pregnant, n = 12) vs. 27.1 pg/mL (not pregnant, n = 17); P = 0.008, Mann Whitney U-test). IL-1alpha and IL 1ra levels were not associated with outcome. CONCLUSIONS: We have demonstrated a significant improvement in blastomere number and frag with ECC. The presence of IL-1beta in the CM was negatively associated with embryonic development and clinical pregnancy. The presence of IL-1alpha in the CM was negatively associated with embryonic development and the presence of IL-1ra in the CM was positively associated with embryonic development. Whether IL-1beta itself interferes with successful outcome after embryo transfer or if it is a marker for undetected endometritis in the biopsy specimens remains to be determined. PMID- 10698035 TI - Relationship between ovarian stimulation regimen and cytokine concentration in follicular fluid and their effect on fertilization and pregnancy outcome of patients undergoing ICSI program. AB - PROBLEM: The aims of this study were to evaluate the presence of insulin-like growth factor (IGF)-I, platelet-derived growth factor (PDGF), and epidermal growth factor (EGF) in pre-ovulatory follicular fluid (FF) in patients undergoing ovarian hyperstimulation for intra-cytoplasmic sperm injection (ICSI) treatment, to determine the differences between the concentrations of these cytokines in relation to ovarian stimulation regimens, and to find the relationship between these parameters and estradiol 17-beta, progesterone, and luteinizing hormone (LH) concentration in serum, as well as ICSI outcome. METHOD: IGF-I and PDGF were measured in the FF of 85 patients. The IGF-I levels were measured by radioimmunoassay, whereas the concentrations of PDGF and EGF were measured by enzyme-linked immunosorbent assay technique, using commercially available kits. RESULTS: IGF-I (0.42 +/- 0.09 ng/mL), PDGF (307.3 +/- 274.5 pg/mL), and EDF (8.88 +/- 6.4 pg/mL) were present in pre-ovulatory FF in patients undergoing ovarian hyperstimulation for ICSI treatment. The mean concentration of IGF-I in the follicle-stimulating hormone (FSH) group was significantly higher (P = 0.036) than that found in the human menopausal gonadotrophin (hMG)/FSH group, whereas no significant difference in the mean concentrations of PDGF (P = 0.58) and EGF was shown between all investigated groups. CONCLUSION: Controlled ovarian stimulation regimens affect only IGF-I levels in FF and the cytokine concentrations of all investigated groups, in turn, showed no correlation either with steroid hormones in serum or ICSI outcome. PMID- 10698036 TI - CA-125 levels are related to the likelihood of pregnancy after in vitro fertilization and embryo transfer. AB - PROBLEM: To assess the relationship between serum cancer antigen (CA)-125 concentrations and the likelihood of pregnancy after in vitro fertilization/embryo transfer (IVF ET). METHOD OF STUDY: A prospective longitudinal follow-up study was conducted in 44 IVF patients receiving luteinizing hormone-releasing hormone (LHRH) suppression, followed by human menopausal gonadotrophin (hMG). There were no indications of endometriosis. Progesterone or human chorionic gonadotrophin (hCG) were given as luteal support. Serum samples were taken just before oocyte pick-up (OPU), 14 days after ET and, after the establishment of a clinical pregnancy, on day ET + 21. Samples were stored at -80 degrees C until analysis. CA-125 was measured using a commercially available enzyme immunoassay (IMx CA-125; Abbott, North Chicago, IL, USA). The signed rank (paired samples) and Wilcoxon tests were used for statistical analysis. RESULTS: There were no differences in CA-125 concentrations at the time of OPU between pregnant (n = 18) and non-pregnant (n = 26) IVF patients. After OPU. the CA-125 concentrations rose both in pregnant (P<0.0001) and in non pregnant (P <0.001) patients. This rise was greater after successful implantation, and 14 days after ET, higher CA-125 concentrations were found in the pregnant patients (P< 0.01). CONCLUSIONS: Differences in serum CA-125 concentrations between pregnant and non-pregnant IVF patients appear to be related to the likelihood of successful implantation. PMID- 10698037 TI - No cyclicity in serum vascular endothelial growth factor during normal menstrual cycle but significant luteal phase elevation during an in vitro fertilization program. AB - PROBLEM: To compare changes in serum vascular endothelial growth factor (VEGF) levels during normal and in vitro fertilization (IVF) cycles. METHOD OF STUDY: Ten healthy women with ovulatory cycles and 37 infertile women participating in an IVF program were followed by frequent serum samples and with VEGF measurements throughout their cycles. RESULTS: Serum VEGF remained unchanged during the normal menstrual cycle, whereas the IVF program participants showed elevations in serum VEGF in the luteal phase of the cycle. When data from controls and patients were pooled, redundant midluteal VEGF level correlated with progesterone and with peak follicular phase estrogen level. The midluteal VEGF level in the IVF cycles was associated with body mass index (P < 0.01) and progesterone level (P < 0.05) by multiple regression. The 14 women conceiving tended to have higher VEGF levels than those failing to become pregnant. CONCLUSIONS: The IVF program was associated with increased synthesis of VEGF either in the ovaries, endometrium, or at other sites and this may be of significance for the outcome of IVF. PMID- 10698038 TI - Activated protein C resistance and factor V Leiden mutation can be associated with first-as well as second-trimester recurrent pregnancy loss. AB - PROBLEM: To examine whether the occurrence of activated protein C resistance (APCR) and factor V Leiden mutation differs in women with first- compared to women with second-trimester unexplained recurrent pregnancy loss. METHOD OF STUDY: Seventy eight consecutive women with two or more unexplained post embryonic recurrent pregnancy losses and 139 fertile women with at least one successful pregnancy and no abortions were prospectively investigated for APCR and the factor V Leiden mutation. No women were pregnant at the time of investigation. APCR was defined as APC sensitivity ratio (APC SR) of < or = 2.0. All patients with an APC SR < or = 2.4 were investigated for the factor V Leiden mutation. Women in this study were divided into three groups. Group A included only women with a history of recurrent first-trimester embryonic loss (37 women) and Group B included women with second-trimester abortions with or without additional first-trimester abortions (41 women). Group C included the controls (139 women). RESULTS: APCR and factor V Leiden mutations were significantly more prevalent in all recurrent pregnancy loss patients in this study as compared to controls. 38%(30/78) and 19%(15/78) in contrast to 8% (11/139) and 6% (8/139), respectively. All three groups in the study were comparable regarding age, parity, and number of living children, whereas Groups A and B were also comparable regarding gravidity. Mean APC SRs were significantly higher in Group C as compared to Groups A and B. The incidence of APCR was significantly higher in Groups A and B, as compared to controls, 27 and 49% in contrast to 8%, respectively. Moreover, the incidence of the factor V Leiden mutation was significantly higher in Groups A and B as compared to Group C, 16 and 22% as distinct from 6%, respectively. The incidence of APCR was higher in Group B as compared to Group A, 49% in contrast to 27%, with borderline significance: however, the factor V Leiden mutation did not significantly differ between the two groups. CONCLUSIONS: APCR and factor V Leiden are associated with unexplained recurrent pregnancy loss. The occurrence of APCR and factor V Leiden seems to be linked to post-embryonic first-trimester as well as second-trimester recurrent pregnancy loss. The significance of acquired, non-heritable APCR in recurrent fetal loss patients, especially in the second-trimester aborters, is still to be determined. PMID- 10698039 TI - Prednisone and aspirin improve pregnancy rate in patients with reproductive failure and autoimmune antibodies: a prospective study. AB - PROBLEM: The study was conducted to investigate the efficacy of prednisone and aspirin in autoantibody seropositive patients with repeated in vitro fertilization embryo transfer (IVF ET) failure. METHODS OF STUDY: The study group comprised 52 consecutive patients seropositive for non-organ-specific autoantibodies, i.e., anti-cardiolipin antibodies (ACA), anti-nuclear antibodies (ANA), anti-double-stranded (ds) DNA, rheumatoid factor (RF), and lupus anti coagulant (LAC). These patients were treated with prednisone, 10 mg per day, and aspirin, 100 mg per day, starting 4 weeks before induction of ovulation in 52 IVF cycles. RESULTS: The clinical pregnancy rate per cycle was 32.7% (17/52). No increased incidence of pregnancy complications, including premature labor, gestational diabetes mellitus, and pregnancy-induced hypertension, were found. CONCLUSIONS: Combined treatment of prednisone for immunosuppression and aspirin as an anti-thrombotic agent, starting before ovulation induction, may improve pregnancy rate in autoantibody seropositive patients who have had repeated IVF-ET failures. PMID- 10698040 TI - Involvement of serum and lipopolysaccharide in the production of interleukin-1- and interleukin-6-like molecules by human sperm cells. AB - PROBLEM: To examine the capacity of sperm cells from fertile and infertile men to secrete interleukin (IL)-6, and the involvement of serum factors and lipopolysaccharide (LPS) in the regulation of IL-6 and IL-1 production by sperm cells. METHODS: Swim-up sperm cells from fertile (donors) and oligoteratoasthenospermic (OTA)-infertile men were incubated with or without 5% fetal calf serum (FCS) and LPS (10 microg/mL) for 2-24 hr. After incubation, IL-6 and IL-1 bioactivities were measured in supernatants and lysates by specific bioassays (B9 cell proliferation assay and 1A-5 system, respectively). RESULTS: IL-6- and IL-1-like activities were observed to be produced by swim-up sperm cells from both study groups. Stimulation of swim-up sperm cells with either LPS or FCS or both together did not affect their capacity to produce IL-1. However, LPS, but not serum increased the secretion levels of IL-6 by swim-up sperm cells. CONCLUSIONS: Swim-up sperm cells from both study groups constitutively produce IL 6 and IL-1, and serum components did not affect this capacity. However, LPS was shown to increase the capacity of swim-up sperm cells of both study groups to secrete IL-6, but not IL-1. Cytokines may be involved in the physiology and pathophysiology of sperm functions and, thus, may affect male fertility. PMID- 10698041 TI - Effects of interferon-gamma on secretion of vascular endothelial growth factor by endometrial stromal cells. AB - PROBLEM: The aim of this study was to clarify the physiological effects of interferon (IFN)-gamma on secretion of vascular endothelial growth factor (VEGF) by endometrial stromal (ES) cells. METHOD OF STUDY: ES cells were obtained from human uterine endometrium by enzymic digestion and filtration. The effects of IFN gamma on production of VEGF by ES cells were examined by analyzing VEGF mRNA expression with Northern blotting analysis and by assaying VEGF protein. RESULTS: IFN-gamma inhibited VEGF mRNA and protein expression by ES cells in a dose dependent manner. In ES cells treated with IFN-gamma, VEGF production was not significant until 6 hr of incubation and was significantly affected after 6 hr of incubation, but decreased significantly after 12 to 48 hr. IFN-gamma also suppressed VEGF mRNA expression by ES cells. CONCLUSIONS: ES cells produce VEGF, which may contribute to endometrial neovascularization and proliferation. IFN gamma may play an important role in regulating VEGF production by ES cells. PMID- 10698042 TI - Decreased expression of mac25 mRNA in uterine leiomyomata compared with adjacent myometrium. AB - PROBLEM: The pathogenesis of uterine leiomyomata is still unclear. Recently it has been suggested that mac25 plays a tumor-suppressive role in various tumors. The aims of this study were to evaluate a possible involvement of mac25 in the growth of leiomyoma and in the mechanism of a gonadotropin-releasing hormone agonist (GnRHa) inducing shrinkage of leiomyoma. METHODS OF STUDY: Mac25 mRNA transcript was measured by Northern blot in total RNA extracted from the paired specimens of leiomyoma and adjacent myometrium from untreated patients (n = 25) and from leiomyoma specimens from GnRHa-pretreated patients (n = 10). RESULTS: Mac25 mRNA expression was significantly lower in large leiomyoma (more than 150 cm3 in volume) than in adjacent myometrium and small leiomyoma (less than 120 cm3 in volume) from untreated patients. There was no difference in this expression between the proliferative and secretory phases of the menstrual cycle. Leiomyoma from GnRHa-pretreated patients had mac25 gene expression levels similar to myometrium and small leiomyoma from untreated patients. CONCLUSIONS: Mac25 may be involved in the growth of uterine leiomyoma and the action of GnRHa may, in part, be mediated by mac25. PMID- 10698043 TI - In vitro expression of proalpha1(I) collagen mRNA by human pre-osteoclastic cells. AB - In vitro studies have demonstrated that the extracellular matrix modulates the cell phenotype. In the present study we have investigated in vitro proalpha1(I) collagen mRNA expression in a human pre-osteoclastic cell line (FLG 29.1 cells) in basal condition and after various stimuli. In addition, in order to evaluate the effect of cell-cell interactions on collagen type I mRNA expression, we have cultured the human pre-osteoclastic cells FLG 29.1 with either the human osteoblast-like cell line Saos-2 or the bovine bone endothelial cell line BBE. We showed that the FLG 29.1 cells express proal (I) collagen mRNA, whose expression is modulated by phorbol esters (TPA). Co-culturing FLG 29.1 cells with either Saos-2 or BBE cells induced decrease of proalpha1(I) collagen mRNA expression. PMID- 10698044 TI - DHEA-S levels in hypopituitaric patients with severe GH deficiency are strongly reduced across lifespan. Comparison with IGF-I levels before and during rhGH replacement. AB - Both IGF-I and DHEA-S undergo an age-related decrease and their decrease could be involved in age-related changes in body composition, structure functions and metabolism. On the other hand, it is well known that mean IGF-I levels are clearly reduced in hypopituitaric patients with GH deficiency (GHD) while data about dehydroepiandrosterone sulfate (DHEA-S) levels in hypopituitarism are scanty. We evaluated DHEA-S and IGF-I levels and their relationship in 90 patients with panhypopituitarism (HYPOPIT) with severe GHD [49 women and 41 men; age, mean+/-SE: 47.9+/-1.49 yr, range: 20-80 yr, BMI: 26.4+/-0.6 kg/m2; 21 with childhood-onset (CO) and 69 with adult-onset (AO) HYPOPIT]. DHEA-S and IGF-I levels were also evaluated in 24 HYPOPIT with GHD after 3-month recombinant human GH (rhGH) replacement. Data in HYPOPIT were compared with those in a large group of healthy controls (NS, 233 women and 103 men, aged 20-80 yr; all subjects were within +/-15% of their ideal body weight). In NS both DHEA-S levels and IGF-I were gender-independent while showed a strong, inverse correlation with age (r= 0.6; p<0.001 and r=-0.56; p<0.0001, respectively). Nevertheless, no relationship was found between DHEA-S and IGF-I levels in NS. In HYPOPIT, age-adjusted mean DHEA-S and IGF-I levels were clearly lower than those in NS (2.3+/-0.4 vs 16.0+/ 0.7 microg/l, p<0.005; 71.1 +/- 4.5 vs 170+/-4.7 microg/l, p<0.005). IGF-I levels in CO-HYPOPIT were lower (p<0.01) than those in AO-HYPOPIT (49.6+/-4.8 vs 77.0+/ 5.4 microg/l), while DHEA-S levels were similar in both subgroups (2.6+/-0.7 vs 2.3+/-0.4 microg/l). In HYPOPIT both DHEA-S and IGF-I were independent of age and gender while there was a trend toward a positive association between each other (r=0.45; p<0.003). Analyzing individual levels in HYPOPIT with respect to age adjusted normal ranges, IGF-I levels were below normal in 84, 62 and 0% between 20-40, 40-60 and 60-80 yr, respectively. On the other hand, DHEA-S levels were below normal in 84, 86 and 67% between 20-40, 40-60 and 60-80, respectively. In HYPOPIT rhGH treatment strikingly increased IGF-I levels (150+/-3.2 vs 85.3+/-4.1 microg/l, p<0.005) while did not modify DHEA-S levels (1.7+/-0.2 vs 1.6+/-0.2 microg/l). In conclusion, our results demonstrate that DHEA-S and IGF-I are negatively and independently associated to age in physiological conditions but not in hypopituitaric patients in whom both are strikingly reduced. Both DHEA-S and IGF-I levels in HYPOPIT show some overlap with those in normal subjects; thus the assay of these parameters is not diagnostic for hypopituitarism. DHEA-S reduction in HYPOPIT does not depend on IGF-I as indicated also by evidence that GH replacement restores IGF-I but does not modify DHEA-S levels. PMID- 10698045 TI - Short-term pre-surgical treatment with somatostatin analogues, octreotide and lanreotide, in acromegaly. AB - Eighteen patients with symptoms of active acromegaly were treated with somatostatin analogues for 4 weeks before surgery. Both before and after the treatment, levels of growth hormone (GH), prolactin (PRL), insulin growth factor I (IGF-I), luteotropin (LH), folliculostimulin (FSH) and subunit alpha of glycoprotein hormones were estimated. Glucose tolerance test, magnetic resonance imaging (MRI) examination, sight acuity and field of vision tests were also performed. The same tests were performed on ten control patients with clinically and biochemically active acromegaly, subjected to surgery but not treated with somatostatin analogues. In six patients treated with somatostatin analogues GH levels decreased significantly to less than 5 ng/ml and in two patients remained elevated while in 10 patients GH level decreased and ranged from 6.1 to 42.9 ng/ml. In 13 patients we observed a decrease in IGF-I to normal levels (<400 ng/dl) and in 3 patients we noted a decrease to levels slightly higher than normal. There was also a slight decrease in alpha subunit concentration. In the glucose inhibition test 4 patients demonstrated normalized GH levels. In patients with elevated PRL and TSH levels, treatment with somatostatin analogues induced their decrease. No changes were observed in levels of LH and FSH. After therapy MRI examination disclosed a decrease in tumor volume in two patients (by 20 and 25%, respectively) and no changes in tumor size in 16 patients. The two patients with a decreased tumor volume also showed normalized glucose tolerance tests. All patients manifested an improved clinical condition. Neurosurgeons disclosed a decreased tumor consistency which greatly facilitated surgical procedure. Our studies documented favourable effects of somatostatin analogues on the assayed hormone levels, and on the general condition of the patients as well as on the course of the surgical procedure itself. PMID- 10698046 TI - Biological action of keratinocyte growth factor in BeWo cells, a human choriocarcinoma cell line. AB - Previously, we reported that the expression of keratinocyte growth factor (KGF) is enhanced in secretory phase endometrial and decidual cells in early pregnancy as compared with the expression of KGF in proliferative phase endometrial cells, in humans. In order to clarify the role of KGF in embryo-endometrial interaction, we analyzed the in vitro effect of KGF on the human chorionic gonadotropin (hCG) secretion and on DNA synthesis in chorionic villi which are in close contact with the endometrium/decidua in the early stage of pregnancy. In this study, we used the BeWo cell line, a human choriocarcinoma cell line that possesses the biological features of secreting various placental hormones including hCG. Furthermore, we investigated the expression of KGF receptor (KGF-R) in these cells. KGF significantly stimulated hCG secretion in cultured BeWo cells but did not affect [3H]-thymidine incorporation. KGF-R mRNA was detected in BeWo cells by reverse transcriptase-polymerase chain reaction. These results suggest that the expression of KGF, which is induced in endometrial/decidual cells by progesterone, plays an important role in the embryo-endometrial/ decidual interaction by stimulating hCG secretion rather than affecting cell proliferation. PMID- 10698047 TI - Pituitary gonadal axis and child rate in males with classical 21-hydroxylase deficiency. AB - Though appropriate glucocorticoid substitution therapy should abolish both cortisol deficiency and adrenal androgen excess in patients with 21-hydroxylase deficiency (21-OHD), the long-term outcome is not always satisfactory. There are several reports on low adult height in both male and female patients, and impaired fertility has been reported in females with 21-OHD. There are only few reports on gonadal function of adult male patients with 21 -OHD. In this study, we calculated the child rate of all the 29 diagnosed adult Finnish males with classical 21-OHD and compared it with the mean child rate of the whole Finnish male population with equal age distribution. Sixteen males with 21-OHD and their age-matched healthy controls were further examined in a cross-sectional study. Auxology and pituitary gonadal axis were examined in both patients and controls. Testicular ultrasonography of the patients was also performed. The mean child rate of the 29 males with 21-OHD was 0.07 which was significantly lower (p<0.001) than that in the Finnish male population of the same age (0.34). In the cross sectional study, males with 21-OHD had serum testosterone, inhibin B, LH and FSH concentrations comparable to those of healthy controls and reference values. Serum DHEA-S concentrations were remarkably low, even in the undersubstituted males with 21-OHD (p<0.001, compared with the healthy controls). In the patient group, serum inhibin B concentration did not correlate with serum FSH concentration. Adrenal rest tumors of the testicles were found in two undersubstituted males with 21-OHD. In conclusion, our study suggests normal pituitary and gonadal function but reduced child rate in adult males with 21-OHD. This might be explained by suboptimal psychosocial adaptation to the chronic disease. However, the patients in this study were young and the final child rate may become essentially higher. PMID- 10698048 TI - Lack of effects of recombinant human growth hormone in a child with a complex cardiovascular malformation and dilated cardiomyopathy. AB - Recent studies have suggested the beneficial effects of GH treatment in patients with dilated cardiomyopathy. We have treated with recombinant human growth hormone (rhGH) a 6-year-old female with a complex congenital heart defect (severe tricuspid hypoplasia and malposition of the great arteries), who developed a progressive dilated cardiomyopathy of unknown etiology. rhGH treatment (0,1 U/kg/day, for 3 months) did not improve cardiac function, nor clinical symptoms, although we have no clear explanations for this. However, a trial with rhGH may be offered to children with dilated cardiomyopathy and waiting for heart transplantation. PMID- 10698049 TI - Effect of long-term treatment with recombinant human growth hormone on erythropoietin secretion in an anemic patient with panhypopituitarism. AB - We studied the effect of treatment with recombinant human GH in an anemic patient with panhypopituitarism in which hemoglobin (Hb) concentration remained as low as 11.0 g/dl in spite of appropriate replacement with thyroid and adrenocortical hormones. Recombinant human GH was subcutaneously and constantly infused for 12 months using a portable syringe pump at a rate of 0.25 U/kg/week. After the treatment with human GH plasma erythropoietin (EPO) levels increased from 12.2 to 25.1 mIU/ml, with a concomitant increase of Hb concentration to 13.6 g/dl. When the administration of human GH was interrupted, both plasma EPO levels and Hb concentrations decreased. There was a close correlation between plasma GH and EPO levels before and during the human GH administration (y=2.444x+1 3.423, r=0.641, p<0.05). Plasma GH levels were well correlated with Hb concentrations before and during human GH administration (y=0.529x+11.313, r=0.690, p<0.01). Plasma IGF4 levels were also correlated with Hb concentrations (y=0.007x+10.874, r=0.832, p<0.001), but not with plasma EPO levels. These findings suggest that GH treatment may be useful in anemic patients with panhypopituitarism. PMID- 10698050 TI - Double adenoma of the pituitary: a somatotroph adenoma colliding with a gonadotroph adenoma. AB - Pituitary collision tumors are rare. They may create difficult diagnostic problems and their histogenesis is not clear. We report here an unusual case of a somatotroph adenoma colliding with a gonadotroph adenoma.The 64-year-old man had clinical acromegaly. His blood growth hormone level was elevated and magnetic resonance imaging demonstrated a pituitary tumor. The surgically removed sellar mass was investigated by histology, immunocytochemistry and electron microscopy. Morphologic study revealed a collision tumor; one was a somatotroph adenoma, the other a gonadotroph adenoma. Authors call attention to the difficulties in clinical, imaging and pathological diagnosis. Detailed morphologic studies are needed to establish the presence of two distinct tumors composed of two different cell types. PMID- 10698051 TI - Autoimmune thyroid disease and breast cancer: a chance association? PMID- 10698052 TI - Is it possible to predict diabetic kidney disease? PMID- 10698053 TI - Obesity and hypertension. PMID- 10698054 TI - Effect of radioiodine therapy on the presence of autoantibodies against human thyroid and eye muscle fractions. PMID- 10698055 TI - Ferritin concentrations in patients with differentiated thyroid cancer. PMID- 10698056 TI - A. Mantegna "The bridal room" (La Camera degli Sposi). PMID- 10698057 TI - Mapping the visual recognition memory network with PET in the behaving baboon. AB - By means of a novel 18F-fluoro-deoxyglucose PET method designed for cognitive activation imaging in the baboon, the large-scale neural network involved in visual recognition memory in the nonhuman primate was mapped for the first time. In this method, the tracer is injected in the awake, unanesthetized, and unrestrained baboon performing the memory task, and brain imaging is performed later under light anesthesia. Brain maps obtained during a computerized trialunique delayed matching-to-sample task (lists of meaningless geometrical patterns and delay > 9 seconds) were statistically compared pixel-by-pixel to maps obtained during a specially designed visuomotor control task. When displayed onto the baboon's own anatomic magnetic resonance images, foci of significant activation were distributed along the ventral occipitotemporal pathway, the inferomedial temporal lobe (especially the perirhinal cortex and posterior hippocampal region), and the orbitofrontal cortex, consistent with lesion, single unit, and autoradiographic studies in monkeys, as well as with activation studies in healthy humans. Additional activated regions included the nucleus basalis of Meynert, the globus pallidus and the putamen. The results also document an unexpected left-sided advantage, suggesting hemispheric functional specialization for recognition of figural material in nonhuman primates. PMID- 10698058 TI - 7-Nitroindazole impedes erythrocyte flow response to isovolemic hemodilution in the cerebral capillary circulation. AB - The role of nitric oxide (NO) in the mechanism of hemodilution-induced cerebral hyperemia is unclear. Based on findings in hypoxemia, the authors hypothesize that NO of neuronal origin contributes to an increase in velocity of erythrocytes in the cerebral microcirculation during anemia produced by isovolemic hemodilution. The change in erythrocyte velocity in cerebrocortical capillaries was assessed by intravital fluorescence video microscopy. A closed cranial window was implanted over the frontoparietal cortex of barbiturate-anesthetized, ventilated adult rats. Erythrocytes were labeled in vitro with fluorescein isothiocyanate and infused intravenously, and their velocity in subsurface capillaries was measured by frame-to-frame image tracking. Arterial blood was withdrawn in increments of 2 mL and replaced by serum albumin; arterial blood pressure was maintained at control level with an infusion of methoxamine. Erythrocyte velocity increased progressively, reaching 215% of baseline, as arterial hematocrit was reduced from 45% to 17%. Pretreatment of a separate group of rats with 7-nitroindazole (20 mg/kg intraperitoneally), a relatively selective inhibitor of neuronal NO synthase, abolished the increase in velocity at hematocrits greater than 20%, but the maximum velocity attained at the lowest hematocrit was similar to that in the control group. The results suggest that NO from neuronal source may contribute to the increase in capillary erythrocyte flow during moderate isovolemic hemodilution. PMID- 10698059 TI - Measurement of striatal and extrastriatal dopamine D1 receptor binding potential with [11C]NNC 112 in humans: validation and reproducibility. AB - To evaluate the postulated role of extrastriatal D1 receptors in human cognition and psychopathology requires an accurate and reliable method for quantification of these receptors in the living human brain. [11C]NNC 112 is a promising novel radiotracer for positron emission tomography imaging of the D1 receptor. The goal of this study was to develop and evaluate methods to derive D1 receptor parameters in striatal and extrastriatal regions of the human brain with [11C]NNC 112. Six healthy volunteers were studied twice. Two methods of analysis (kinetic and graphical) were applied to 12 regions (neocortical, limbic, and subcortical regions) to derive four outcome measures: total distribution volume, distribution volume ratio, binding potential (BP), and specific-to-nonspecific equilibrium partition coefficient (k3/k4). Both kinetic and graphic analyses provided BP and k3/k4 values in good agreement with the known distribution of D1 receptors (striatum > limbic regions = neocortical regions > thalamus). The identifiability of outcome measures derived by kinetic analysis was excellent. Time-stability analysis indicated that 90 minutes of data collection generated stable outcome measures. Derivation of BP and k3/k4 by kinetic analysis was highly reliable, with intraclass correlation coefficients (ICCs) of 0.90+/-0.06 (mean +/- SD of 12 regions) and 0.84+/-0.11, respectively. The reliability of these parameters derived by graphical analysis was lower, with ICCs of 0.72+/-0.17 and 0.58+/ 0.21, respectively. Noise analysis revealed a noise-dependent bias in the graphical but not the kinetic analysis. In conclusion, kinetic analysis of [11C]NNC 112 uptake provides an appropriate method with which to derive D1 receptor parameters in regions with both high (striatal) and low (extrastriatal) D1 receptor density. PMID- 10698060 TI - The suitability of [11C]-alpha-methyl-L-tryptophan as a tracer for serotonin synthesis: studies with dual administration of [11C] and [14C] labeled tracer. AB - The tracer [11C]-alpha-methyl-L-tryptophan (alphaMTP) has been used to measure brain serotonin synthesis rates with positron emission tomography (PET). To address questions about the accuracy of the kinetic model, [14C]alphaMTP was used to directly measure conversion to [14C]-alpha-methyl-serotonin (alphaM5HT) in monkeys that had been previously studied with PET and [11C]alphaMTP. Four male, fasted, isoflurane-anesthetized rhesus monkeys were studied with [11C]alphaMTP and PET. Immediately after the initial 3-hour scan, a second dose of [11C]alphaMTP was coinjected with 1 mCi of [14C]alphaMTP, and additional PET data were collected. Approximately 90 minutes after the second alphaMTP administration, the animals were killed with an overdose of phenobarbital, and brain samples from 21 regions were taken and analyzed by HPLC. Minimal conversion of alphaMTP to alphaM5HT occurred; HPLC analysis of 14C radioactivity showed that greater than 96% of the total counts were in fractions corresponding to the alphaMTP peak. Brain concentrations of serotonin, tryptophan, 5-hydroxyindole-3 acetic acid, and alphaMTP also were determined fluorometrically using external quantification. Patlak plots generated from PET images acquired over 3 hours showed no time period of linear increase, and final slopes were not significantly different from zero, consistent with the finding of minimal conversion to [14C]alphaM5HT. These data indicate that in the 3-hour period after injection, [11C]alphaMTP is acting predominantly as a tracer of tryptophan uptake, not serotonin synthesis. PMID- 10698061 TI - Evaluation of serotonergic transporters using PET and [11C](+)McN-5652: assessment of methods. AB - [11C](+)McN-5652 is an established positron emission tomography tracer used to assess serotonergic transporter density. Several methods have been used to analyze [11C](+)McN-5652 data; however, no evaluation of candidate methods has been published in detail yet. In this study, compartmental modeling using a one tissue compartment model (K1, k2"), a two-tissue compartment model (K1 to k4), and a noncompartmental method that relies on a reference region devoid of specific binding sites were assessed. Because of its low density of serotonergic transporters, white matter was chosen as reference. Parameters related to transporter density were the total distribution volume DV" (= K1/k2", one tissue compartment), DVtot, (=K1/k1' (1 + k3/k4), two tissue compartments), and Rv (= k3'/k4, noncompartmental method). The DV", DVtot, and Rv values extended over a similar range and reflected the known pattern of serotonergic transporters. However, all parameters related to transporter density were markedly confounded by nonspecific binding. With regard to K1, the one-tissue compartment model yielded markedly lower values, which were, however, more stable. The minimal study duration needed to determine stable values for the distribution volume was approximately 60 minutes. The choice of the method to analyze [11C](+)McN-5652 data depends on the situation. Parametric maps of Rv are useful if no information on K1 is needed. If compartmental modeling is chosen, both the one- and the two tissue compartment models have advantages. The one-tissue compartment model underestimates K1 but yields more robust values. The distribution volumes calculated with both models contain a similar amount of information. None of the parameters reflected serotonergic transporter density in a true quantitative manner, as all were confounded by nonspecific binding. PMID- 10698062 TI - Assessment of hemispheric language lateralization: a comparison between fMRI and fTCD. AB - The cerebral blood flow velocity (CBFV) in the basal arteries during a word generation task was assessed by functional transcranial Doppler ultrasonography (fTCD) and by functional magnetic resonance imaging (fMRI). The study investigates how event-related CBFV modulations in the middle cerebral artery (MCA) relate to regional cerebral blood flow (rCBF) changes. Both fMRI and fTCD were used in 13 subjects (7 men, 6 women, aged 21 to 44 years). The maximum difference of relative CBFV changes between the left and right MCA during the word-generation task was used as the language laterality index (LIfTCD). For the fMRI examination during the nearly identical language task, the corresponding index was defined by LIfMRI = 100(N(L) - N(R))/(N(L) + N(R)), where N(L) and N(R) refer to the numbers of voxels activated in the left and right hemisphere, respectively. The evoked CBFV changes expressed by LIfTCD and the corresponding laterality index, LIfMRI, estimated by fMRI showed a close linear relation (regression analysis: r = 0.95, p < 0.0001). The results of this study demonstrate that language-related velocity changes in the MCAs relate to rCBF increases in a linear fashion. Since the laterality indices assessed by fMRI and fTCD are in such close agreement both techniques can therefore be used in a complementary way. PMID- 10698063 TI - Measurement of cerebral blood flow volume by extracranial sonography. AB - Measurement of global cerebral blood flow (CBF) using extracranial duplex sonography has been described, but normal values are still lacking. In 85 healthy adults (median age 43.4 years, range 20 to 80 years), CBF was determined by duplex sonographic examination of both internal carotid arteries and vertebral arteries. The mean global CBF was 630+/-97 mL/min. Global CBF declined with age; sex did not influence the total CBF. When measurements were repeated, the intraindividual variability was low. This noninvasive sonographic measurement of CBF is reproducible, and values correspond closely to those obtained with positron emission tomography and magnetic resonance imaging. PMID- 10698064 TI - Determinants of cerebral fractional oxygen extraction using near infrared spectroscopy in preterm neonates. AB - Cerebral fractional oxygen extraction (FOE) represents the balance between cerebral oxygen delivery and consumption. This study aimed to determine cerebral FOE in preterm infants during hypotension, during moderate anemia, and with changes in the PaCO2. Three groups of neonates were studied: stable control neonates (n = 43), anemic neonates (n = 46), and hypotensive neonates (n = 19). Cerebral FOE was calculated from the arterial oxygen saturation measured by pulse oximetry, and cerebral venous oxygen saturation was measured using near infrared spectroscopy with partial jugular venous occlusion. Mean +/- SD cerebral FOE was similar in control (0.292+/-0.06), anemic (0.310+/-0.08; P = 0.26), and hypotensive (0.278+/-0.06; P = 0.41) neonates. After anemic neonates were transfused, mean +/- SD cerebral FOE decreased to 0.274+/-0.05 (P = 0.02). There was a weak negative correlation with the hemoglobin concentration (n = 89, r = 0.24, P = 0.04) but not with the hemoglobin F fraction (n = 56, r = 0.24, P = 0.09). In the hypotensive neonates, there was no relationship between cerebral FOE and blood pressure (n = 19, r = 0.34, P = 0.15). There was a significant negative correlation between cerebral FOE and PaCO2 within individuals (n = 14, r = -0.63, P = 0.01), but there was no relationship between individuals (n = 14, r = 0, P = 1). Cerebral FOE was not significantly altered in neonates with either mild anemia or hypotension. There were, however, changes in cerebral FOE when physiological changes occurred over a relatively short period: Cerebral FOE decreased after blood transfusion and increased with decreasing PaCO2. As no change in cerebral FOE was seen during hypotension, it was speculated that cerebral oxygen delivery may have been maintained by cerebral blood flow autoregulation. PMID- 10698065 TI - Oxygen dependency of cerebral oxidative phosphorylation in newborn piglets. AB - Changes in hemoglobin oxygenation and oxidation state of the CuA centre of cytochrome oxidase were measured with full spectral near infrared spectroscopy simultaneously with phosphorus metabolites using nuclear magnetic resonance 31P spectroscopy at high time resolution (10 seconds) during transient anoxia (FiO2 = 0.0 for 105 seconds) in the newborn piglet brain. During the onset of anoxia, there was no change in either phosphocreatine (PCr) concentration or the oxidation state of the CuA centre of cytochrome oxidase until there was a substantial fall in cerebral hemoglobin oxygenation, at which point the CuA centre reduced simultaneously with the decline in PCr. At a later time during the anoxia, intracellular pH decreased rapidly, consistent with a fall in cerebral metabolic rate for O2 and reduced flux through the tricarboxylic acid cycle. The simultaneous reduction of CuA and decline in PCr can be explained in terms of the effects of the falling mitochondrial electrochemical potential. From these observations, it is concluded that, at normoxia, oxidative phosphorylation and the oxidation state of the components of the electron transport chain are independent of cerebral oxygenation and that the reduction in the CuA signal occurs when oxygen tension limits the capacity of oxidative phosphorylation to maintain the phosphorylation potential. PMID- 10698066 TI - Effects of variations in interstimulus interval on activation-flow coupling response and somatosensory evoked potentials with forepaw stimulation in the rat. AB - In functional neuroimaging studies, the hemodynamic response to functional activation is used as a surrogate marker for neuronal activity, typically in response to task paradigms that use periodic stimuli. With use of a model system of electrical forepaw stimulation in rats (n = 14) with laser-Doppler (LD) monitoring of cerebral blood flow (CBF) changes in the somatosensory cortex, the effects of variations in the interstimulus interval (ISI) on the hemodynamic response to periodic stimuli were examined. A characteristic peak flow response was seen for 4-second stimuli and a peak and plateau response were seen for all 8 second stimuli regardless of ISI. However, both the amplitude of the LD(CBF) response and the integrated response were significantly reduced for shorter ISIs, whereas the baseline flow was not altered. Somatosensory evoked potential responses were also recorded in some rats (n = 8) and remained unchanged for the various ISIs for a particular stimulus duration. These results suggest that the decrease in the LD(CBF) responses observed with shorter ISIs likely represents a refractoriness of the hemodynamic response and not neuronal function. These results may have important implications for the optimization and interpretation of functional activation paradigms that use periodic stimuli. PMID- 10698067 TI - Muscarinic--but not nicotinic--acetylcholine receptors mediate a nitric oxide dependent dilation in brain cortical arterioles: a possible role for the M5 receptor subtype. AB - Increases in cortical cerebral blood flow are induced by stimulation of basal forebrain cholinergic neurons. This response is mediated in part by nitric oxide (NO) and reportedly involves both nicotinic and muscarinic receptors, some of which are possibly located in the vessel wall. In the present study, the vasomotor response(s) elicited by acetylcholine (ACh) on isolated and pressurized bovine and/or human intracortical penetrating arterioles were investigated, and pharmacological characterization of the receptor involved in this response was carried out. Acetylcholine (10(-11) to 10(-4) mol/L) dose dependently dilated bovine and human intracortical arterioles at spontaneous tone (respective pD2 values of 6.4+/-0.3 and 7.2+/-0.3 and E(Amax) of 65.0+/-26.8 and 43.2+/-30.1% of the maximal dilation obtained with papaverine) and bovine arterioles after preconstriction with serotonin (pD2 = 6.3+/-0.1, E(Amax) = 80.0+/-17.9% of induced tone). In contrast, nicotine (10(-8) to 10(-4) mol/L) failed to induce any vasomotor response in bovine vessels whether at spontaneous or at pharmacologically induced tone. Application of the nitric oxide synthase (NOS) inhibitor Nomega-nitro-L-arginine (L-NNA; 10(-5) mol/L) elicited a gradual constriction (approximately 20%) of the arterioles, indicating the presence of constitutive NO release in these vessels. Nomega-Nitro-L-argigine (10(-5) to 10( 4) mol/L) also significantly blocked the dilation induced by ACh. The muscarinic ACh receptor (mAChR) antagonists pirenzepine, 4-DAMP, and AF-DX 384 dose dependently inhibited the dilatation induced by ACh (10(-5) mol/L) with the following rank order of potency: 4-DAMP (pIC50 = 9.2+/-0.3) >> pirenzepine (pIC50 = 6.7+/-0.4) > AF-DX 384 (pIC50 = 5.9+/-0.2). These results suggest that ACh can induce a potent, dose-dependent, and NO-mediated dilation of bovine and/or human intracortical arterioles via interaction with an mAChR that best corresponds to the M5 subtype. PMID- 10698068 TI - Dynamics of regional brain metabolism and gene expression after middle cerebral artery occlusion in mice. AB - The evolution of brain infarcts during permanent occlusion of the middle cerebral artery (MCA) was studied in mice using multiparametric imaging techniques. Regional protein synthesis and the regional tissue content of ATP were measured on adjacent cryostat sections at increasing intervals after vascular occlusion ranging from 1 hour to 3 days. The observed changes were correlated with the expression of the mRNA of hsp70, c-fos, c-jun, and junB, as well as the distribution of DNA double-strand breaks visualized by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labelling (TUNEL). One hour after MCA occlusion, the tissue volume with suppressed protein synthesis was distinctly larger than that in which ATP was depleted. With ongoing ischemia time, the ATP depleted area gradually expanded and, within 1 day, merged with the region of suppressed protein synthesis. Expression of hsp70 mRNA occurred mainly in the penumbra (defined as the region of suppressed protein synthesis but preserved ATP), peaking at 3 hours after vascular occlusion. Expression of the immediate early genes c-jun, c-fos, and junB increased both in the penumbra and the periinfarct normal tissue already at 1 hour after vascular occlusion, with slightly different regional and temporal patterns for each of these genes. DNA fragmentations were clearly confined to neurons; they appeared after 1 day in the infarct core (defined as the region of suppressed ATP) and never were detected in the penumbra. The late appearance of TUNEL after infarcts had reached their final size and the absence in the penumbra points against a major pathogenetic role of apoptosis. Permanent MCA occlusion in mice thus produces a gradually expanding infarct, the final size of which is heralded by the early inhibition of protein synthesis. PMID- 10698069 TI - Graded reduction of cerebral blood flow in rat as detected by the nuclear magnetic resonance relaxation time T2: a theoretical and experimental approach. AB - The ability of transverse nuclear magnetic resonance relaxation time, T2, to reveal acutely reduced CBF was assessed using magnetic resonance imaging (MRI). Graded reduction of CBF was produced in rats using a modification of Pulsinelli's four-vessel occlusion model. The CBF in cerebral cortex was quantified using the hydrogen clearance method, and both T2 and the trace of the diffusion tensor (Dav = 1/3TraceD) in the adjacent cortical tissue were determined as a function of reduced CBF at 4.7 T. A previously published theory, interrelating cerebral hemodynamic parameters, hemoglobin, and oxygen metabolism with T2, was used to estimate the effects of reduced CBF on cerebral T2. The MRI data show that T2 reduces in a U-shape manner as a function of CBF, reaching a level that is 2.5 to 2.8 milliseconds (5% to 6%) below the control value at CBF, between 15% and 60% of normal. This reduction could be estimated by the theory using the literature values of cerebral blood volume, oxygen extraction ratio, and precapillary oxygen extraction during compromised CBF. Dav dropped with two apparent flow thresholds, so that a small 11% to 17% reduction occurred between CBF values of 16% to 45% of normal, followed by a precipitous collapse by more than 20% at CBF below 15% of normal. The current data show that T2 can be used as an indicator of acute hypoperfusion because of its ability to indicate blood oxygenation level dependent phenomena on reduced CBF. PMID- 10698070 TI - Cerebral metabolism of lactate in vivo: evidence for neuronal pyruvate carboxylation. AB - The cerebral metabolism of lactate was investigated. Awake mice received [3 13C]lactate or [1-13C]glucose intravenously, and brain and blood extracts were analyzed by 13C nuclear magnetic resonance spectroscopy. The cerebral uptake and metabolism of [3-13C]lactate was 50% that of [1-13C]glucose. [3-13C]Lactate was almost exclusively metabolized by neurons and hardly at all by glia, as revealed by the 13C labeling of glutamate, gamma-aminobutyric acid and glutamine. Injection of [3-13C]lactate led to extensive formation of [2-13C]lactate, which was not seen with [1-13C]glucose, nor has it been seen in previous studies with [2-13C]acetate. This formation probably reflected reversible carboxylation of [3 13C]pyruvate to malate and equilibration with fumarate, because inhibition of succinate dehydrogenase with nitropropionic acid did not block it. Of the [3 13C]lactate that reached the brain, 20% underwent this reaction, which probably involved neuronal mitochondrial malic enzyme. The activities of mitochondrial malic enzyme, fumarase, and lactate dehydrogenase were high enough to account for the formation of [2-13C]lactate in neurons. Neuronal pyruvate carboxylation was confirmed by the higher specific activity of glutamate than of glutamine after intrastriatal injection of [1-14C]pyruvate into anesthetized mice. This procedure also demonstrated equilibration of malate, formed through pyruvate carboxylation, with fumarate. The demonstration of neuronal pyruvate carboxylation demands reconsideration of the metabolic interrelationship between neurons and glia. PMID- 10698071 TI - Plasminogen activation in focal cerebral ischemia and reperfusion. AB - In focal cerebral ischemia the plasminogen-plasmin system plays a role in the fibrinolysis of vessel-occluding clots and also in the proteolysis of extracellular matrix components, which potentially contributes to brain edema and bleeding complications. The authors investigated the plasminogen activation after middle cerebral artery occlusion with and without reperfusion (reperfusion intervals 9 and 24 hours) in rats by histologic zymography and compared areas of increased plasminogen activation to areas of structural injury, which were detected immunohistochemically. After 3 hours of ischemia, increased plasminogen activation was observed in the ischemic hemisphere. The affected area measured 5.2%+/-8.5% and 19.4%+/-30.1% of the total basal ganglia and cortex area, respectively. Reperfusion for 9 hours after 3 hours of ischemia led to a significant expansion of plasminogen activation in the basal ganglia (68.8%+/ 42.2%, P < 0.05) but not in the cortex (43.0%+/-34.6%, P = 0.394). In the basal ganglia, areas of increased plasminogen activation were related to areas of structural injury (r = 0.873, P < 0.001). No such correlation was found in the cortex (r = 0.299, P = 0.228). In this study, increased plasminogen activation was demonstrated early in focal cerebral ischemia. This activation may promote early secondary edema formation and also secondary hemorrhage after ischemic stroke. PMID- 10698072 TI - Interplay between the gamma isoform of PKC and calcineurin in regulation of vulnerability to focal cerebral ischemia. AB - Protein phosphorylation and dephosphorylation mediated by protein kinases and protein phosphatases, respectively, represent essential steps in a variety of vital neuronal processes that could affect susceptibility to ischemic stroke. In this study, the role of the neuron-specific gamma isoform of protein kinase C (gammaPKC) in reversible focal ischemia was examined using mutant mice in which the gene for gammaPKC was knocked-out (gammaPKC-KO). A period of 150 minutes of unilateral middle cerebral artery and common carotid artery (MCA/CCA) occlusion followed by 21.5 hours of reperfusion resulted in significantly larger (P < 0.005) infarct volumes (n = 10; 31.1+/-4.2 mm3) in gammaPKC-KO than in wild-type (WT) animals (n = 12; 22.6+/-7.4 mm3). To control for possible differences related to genetic background, the authors analyzed Balb/cJ, C57BL/6J, and 129SVJ WT in the MCA/CCA model of focal ischemia. No significant differences in stroke volume were detected between these WT strains. Impaired substrate phosphorylation as a consequence of gammaPKC-KO might be corrected by inhibition of protein dephosphorylation. To test this possibility, gammaPKC-KO mice were treated with the protein phosphatase 2B (calcineurin) inhibitor, FK-506, before ischemia. FK 506 reduced (P < 0.008) the infarct volume in gammaPKC-KO mice (n = 7; 24.6+/-4.6 mm3), but at this dose in this model, had no effect on the infarct volume in WT mice (n = 7; 20.5+/-10.7 mm3). These results indicate that gammaPKC plays some neuroprotective role in reversible focal ischemia. PMID- 10698073 TI - Hypoxic but not ischemic neurotoxicity of free radicals revealed by dynamic changes in glucose metabolism of fresh rat brain slices on positron autoradiography. AB - Dynamic changes in the regional cerebral glucose metabolic rate induced by hypoxia/reoxygenation or ischemia/reperfusion were investigated with a positron autoradiography technique. Fresh rat brain slices were incubated with [18F]2 fluoro-2-deoxy-D-glucose ([18F]FDG) in oxygenated Krebs-Ringer solution at 36 degrees C, and serial two-dimensional time-resolved images of [18F]FDG uptake in the slices were obtained. In the case of loading hypoxia (oxygen deprivation)/pseudoischemia (oxygen and glucose deprivation) for various periods of time, the net influx constant (K) of [18F]FDG at preloading and after reoxygenation/pseudoreperfusion (post-loading) was quantitatively evaluated by applying the Patlak graphical method to the image data. Regardless of the brain region, with hypoxia lasting > or =20 minutes, the postloading K value was decreased compared with the unloaded control, whereas with pseudoischemia of < or =40 minutes, approximately the same level as the unloaded control was maintained. Next, the neuroprotective effect against hypoxia/pseudoischemia loading induced by the addition of a free radical scavenger or an N-methyl-D-aspartate (NMDA) antagonist was assessed by determining whether a decrease in the postloading K value was prevented. Whereas with 20-minute hypoxia, both agents exhibited a neuroprotective effect, in the case of 50-minute pseudoischemia, only the NMDA antagonist did so, with the free radical scavenger being ineffective. These results demonstrate that hypoxia causes irreversible neuronal damage within a shorter period than ischemia, with both free radicals and glutamate suggested to be involved in tandem in the neurotoxicity induced by hypoxia, whereas glutamate alone is involved in ischemic neurotoxicity. PMID- 10698074 TI - Differing effects of copper,zinc superoxide dismutase overexpression on neurotoxicity elicited by nitric oxide, reactive oxygen species, and excitotoxins. AB - Overexpression of Cu,Zn superoxide dismutase (SOD1) reduces ischemic injury in some stroke models but exacerbates injury in a neonatal stroke model and in other settings. The current study used a SOD1 transgenic (SOD1-Tg) murine cortical culture system, derived from the same mouse strain previously used for the stroke models, to identify conditions that determine whether SOD1 overexpression in neurons is protective or detrimental. The nitric oxide (NO) donors S-nitroso-N acetylpenicillamine, spermine-NONOate, and diethylamine-NONOate produced less death in SOD1-Tg neurons than in wild-type neurons (p < 0.01). Also, NO produced markedly less 3-nitrotyosine in SOD1-Tg cells. In contrast, the superoxide generator menadione produced significantly greater death and nearly twice as much 2'7'-dichlorofluorescein fluorescence in SOD1-Tg neurons than in wild-type neurons, suggesting increased peroxide formation in the SOD1-Tg cells. No significant difference was observed in the vulnerability of the two cell types to H2O2, the product of the SOD reaction. Overexpression of SOD1 also had no effect on neuronal vulnerability to glutamate, N-methyl-D-aspartate, or kainate. These observations suggest that SOD1 overexpression can reduce neuronal death under conditions where peroxynitrite formation is a significant factor, but may exacerbate neuronal death under conditions of rapid intracellular superoxide formation or impaired H2O2 disposal. PMID- 10698075 TI - Experimental closed head injury: analysis of neurological outcome, blood-brain barrier dysfunction, intracranial neutrophil infiltration, and neuronal cell death in mice deficient in genes for pro-inflammatory cytokines. AB - Cytokines are important mediators of intracranial inflammation following traumatic brain injury (TBI). In the present study, the neurological impairment and mortality, blood-brain barrier (BBB) function, intracranial polymorphonuclear leukocyte (PMN) accumulation, and posttraumatic neuronal cell death were monitored in mice lacking the genes for tumor necrosis factor (TNF)/lymphotoxin alpha (LT-alpha) (TNF/LT-alpha-/-) and interleukin-6 (IL-6) and in wild-type (WT) littermates subjected to experimental closed head injury (total n = 107). The posttraumatic mortality was significantly increased in TNF/LT-alpha-/- mice (40%; P < 0.02) compared with WT animals (10%). The IL-6-/- mice also showed a higher mortality (17%) than their WT littermates (5.6%), but the difference was not statistically significant (P > 0.05). The neurological severity score was similar among all groups from 1 to 72 hours after trauma, whereas at 7 days, the TNF/LT alpha-/- mice showed a tendency toward better neurological recovery than their WT littermates. Interestingly, neither the degree of BBB dysfunction nor the number of infiltrating PMNs in the injured hemisphere was different between WT and cytokine-deficient mice. Furthermore, the analysis of brain sections by in situ DNA nick end labeling (TUNEL histochemistry) at 24 hours and 7 days after head injury revealed a similar extent of posttraumatic intracranial cell death in all animals. These results show that the pathophysiological sequelae of TBI are not significantly altered in mice lacking the genes for the proinflammatory cytokines TNF, LT-alpha, and IL-6. Nevertheless, the increased posttraumatic mortality in TNF/LT-alpha-deficient mice suggests a protective effect of these cytokines by mechanisms that have not been elucidated yet. PMID- 10698076 TI - Development of an in situ mouse brain perfusion model and its application to mdr1a P-glycoprotein-deficient mice. AB - An in situ mouse brain perfusion model predictive of passive and carrier-mediated transport across the blood-brain barrier (BBB) was developed and applied to mdr1a P-glycoprotein (Pgp)-deficient mice [mdr1a(-/-)]. Cerebral flow was estimated from diazepam uptake. Physical integrity of the BBB was assessed with sucrose/inulin spaces; functional integrity was assessed with glucose uptake, which was saturable with a Km of approximately 17 mmol/L and Vmax of 310 mmol x 100 g(-1) x min(-1). Brain uptake of a Pgp substrate (colchicine) was significantly enhanced (two- to fourfold) in mdr1a(-/-) mice. These data suggest that the model is applicable to elucidating the effects of efflux transporters, including Pgp, on brain uptake. PMID- 10698077 TI - Induction of angiopoietin and Tie receptor mRNA expression after cerebral ischemia-reperfusion. AB - The angiopoietin/Tie receptor system may contribute to angiogenesis and vascular remodeling by mediating interactions of endothelial cells with smooth muscle cells and pericytes. The temporal expression of angiopoietin-1 (Angpo-1), angiopoietin-2 (Angpo-2), Tie-1, and Tie-2 mRNA was studied in a focal cerebral ischemia model in rats. The cDNA fragments obtained from reverse transcription polymerase chain reaction amplification were cloned and used as a probe to detect individual genes. Northern blot analysis showed a delayed increase of a 4.4-kb Angpo-1 transcript for up to 2 weeks after ischemia, eightfold higher than the values of the sham-operated controls. A biphasic expression of a 2.4-kb Angpo-2 transcript was noted, peaking at 24 hours (6.4-fold) and 2 weeks (4.6-fold) after ischemia. The expression of Tie-2 mRNA (4.3 kb), a receptor for Angpo-1, and Tie 1 mRNA (4.3 kb) also increased starting 24 hours after reperfusion and remained elevated for up to 2 weeks after ischemia. The temporal profiles of the expression of these genes were different from those of other angiogenic genes such as basic fibrobast growth factor/fibroblast growth factor receptor and vascular endothelial growth factor/vascular endothelial growth factor receptor and proteolytic enzymes (tissue-type plasminogen activator and urokinase plasminogen activator) and their inhibitors (plasminogen activator inhibitor-1). The expression patterns of these genes could be related to progressive tissue liquefaction and neovascularization after ischemia in this stroke model. Differential expression of these angiogenesis genes suggests the involvement of complex regulatory mechanisms that remain to be characterized. PMID- 10698078 TI - Evidence for apoptosis after intercerebral hemorrhage in rat striatum. AB - The overall hypothesis that cell death after intracerebral hemorrhage is mediated in part by apoptotic mechanisms was tested. Intracerebral hemorrhage was induced in rats using stereotactic infusions of 0.5 U of collagenase (1-microL volume) into the striatum. After 24 hours, large numbers of TUNEL-positive stained cells with morphologies suggestive of apoptosis were present in the center and periphery of the hemorrhage. Double staining with Nissl and immunocytochemical labeling with antibodies against neuronal nuclei and glial fibrillary acidic protein suggested that these TUNEL-positive cells were mostly neurons and astrocytes. Electrophoresis of hemorrhagic brain extracts showed evidence of DNA laddering into approximately 200-bp fragments. Western blots showed cleavage of the cytosolic caspase substrate gelsolin. The density of TUNEL-positive cells at 24 and 48 hours after hemorrhage was significantly reduced by treatment with the broad-spectrum caspase inhibitor zVADfmk. It was unlikely that apoptotic changes were due to neurotoxicity of injected collagenase because TUNEL-positive cells and DNA laddering were also obtained in an alternative model of hemorrhage where autologous blood was infused into the striatum. Furthermore, equivalent doses of collagenase did not induce cell death in primary neuronal cultures. These results provide initial evidence that apoptotic mechanisms may mediate some of the injury in brain after intracerebral hemorrhage. PMID- 10698079 TI - Reduction in water and metabolite apparent diffusion coefficients during energy failure involves cation-dependent mechanisms. A proton nuclear magnetic resonance study of rat cortical brain slices. AB - Proton (1H) nuclear magnetic resonance (NMR) diffusion spectroscopy was used to assess apparent diffusion coefficients (ADCs) in rat brain slices. Aglycemic hypoxia caused reductions in the ADC of N-acetylaspartate (NAA) (0.15 to 0.09 x 10(-3) mm2/s) and "slow" diffusion coefficient (D2) of tissue water (0.51 to 0.37 x 10(-3) mm2/s), together with a 32+/-11% increase in tissue water volume, attributable to tissue swelling. The ADC and D2 reductions were diminished, however, by removing external Ca2+, and under 10 mmol/L Mg2+, normoxic diffusion coefficients persisted until 40 minutes of hypoxia. The data suggest that the shift of water into the intracellular space alone cannot satisfactorily explain the reduced cerebral diffusion upon energy failure and that external Mg2+ and Ca2+ play crucial modulatory roles. PMID- 10698080 TI - Glutamate does not mediate acute neuronal damage after spreading depression induced by O2/glucose deprivation in the hippocampal slice. AB - This study argues that, in contrast to accepted excitotoxicity theory, O2/glucose deprivation damages neurons acutely by eliciting ischemic spreading depression (SD), a process not blocked by glutamate antagonists. In live rat hippocampal slices, the initiation, propagation, and resolution of SD can be imaged by monitoring wide-band changes in light transmittance (i.e., intrinsic optical signals). Oxygen/glucose deprivation for 10 minutes at 37.5 degrees C evokes a propagating wave of elevated light transmittance across the slice, representing the SD front. Within minutes, CA1 neurons in regions undergoing SD display irreversible damage in the form of field potential inactivation, swollen cell bodies, and extensively beaded dendrites, the latter revealed by single-cell injection of lucifer yellow. Importantly, glutamate receptor antagonists do not block SD induced by O2/glucose deprivation, nor do they prevent the resultant dendritic beading of CA1 neurons. However, CA1 neurons are spared if SD is suppressed by reducing the temperature to 35 degrees C during O2/glucose deprivation. This supports previous electrophysiologic evidence in vivo that SD during ischemia promotes acute neuronal damage and that glutamate antagonists are not protective of the metabolically stressed tissue. The authors propose that the inhibition of ischemic SD should be targeted as an important therapeutic strategy against stroke damage. PMID- 10698081 TI - Tissue distribution of histo-blood group antigens. AB - The introduction of immunohistochemical techniques and monoclonal antibodies to specific carbohydrate epitopes has made it possible to study in detail the tissue distribution of histo-blood group antigens and related carbohydrate structures. The present paper summarizes the available data concerning the histological distribution of histo-blood group antigens and their precursor structures in normal human tissues. Studies performed have concentrated on carbohydrate antigens related to the ABO, Lewis, and TTn blood group systems, i.e. histo-blood group antigens carried by type 1, 2, and 3 chain carrier carbohydrate chains. Histo-blood group antigens are found in most epithelial tissues. Meanwhile, several factors influence the type, the amount, and the histological distribution of histoblood group antigens, i.e. the ABO, Lewis, and saliva-secretor type of the individual, and the cell- and tissue type. Oligosaccharides with blood-group specificity are synthesized by the stepwise action of specific gene-encoded glycosyltransferases. In general, this stepwise synthesis of histo-blood group antigens correlates with cellular differentiation. The H and the Se genes both encode an al-2fucosyltransferase, which is responsible for the synthesis of blood group antigen H from precursor disaccharides. A new model for the participation of the Se/H-gene-encoded glycosyl transferases in synthesis of terminal histo blood group antigens in human tissues is proposed; the type and degree of differentiation rather than the embryologic origin determines whether it is the H or the Se gene-encoded transferases that influence expression of terminal histo blood group antigens in tissues. PMID- 10698082 TI - NO and de novo mammalian angiogenesis: further evidence that NO inhibits bFGF induced angiogenesis while not influencing VEGF165-induced angiogenesis. AB - Using the non-surgical rat mesenteric window angiogenesis assay, we studied the systemic effect of (i) the nitric oxide (NO)-releasing vasodilator isosorbide-5 mononitrate (ISMN) and (ii) the NO-synthase inhibitor L-NAME on angiogenesis induced by the intraperitoneal injection of bFGF and VEGF165. The response was assessed objectively and quantitatively by microscopic morphometry and image analysis in terms of the vascularized area (VA; a measurement of microvessel spatial extension), the microvascular length (MVL; a composite measurement of microvessel density), the total microvascular length (TMVL=VA x MVL), the number of microvessel segments per unit tissue volume (No. MS), the length of the microvessel segments (Le. MS) and the degree of microvessel tortuosity (MVT). Additional architectural features of the network were assessed in terms of variables introduced here: the number of microvessel branching points per unit tissue volume (No. BP), the index of interconnecting microvessel loop formation (In. LF), the index of microvessel intersection (In. IS), the number of microvessel sprouts per unit tissue volume (No. SP) and their length (Le. SP). In bFGF-mediated angiogenesis, L-NAME significantly, augmented angiogenesis, whereas ISMN significantly inhibited angiogenesis. By contrast, neither L-NAME nor ISMN affected the angiogenic response to VEGF165. PMID- 10698083 TI - Application of the polymerase chain reaction in determination of recombinant retrovirus titers as fifty percent endpoints. AB - Retroviral vectors constitute the most efficient system to deliver and integrate foreign genes into mammalian cells. One of the most laborious routine assays in the application of retroviral-mediated gene transfer is the determination of viral titers of vector producer cell lines. Traditionally, determination of virus titer involves the testing of culture medium from individual packaging cell lines for the ability to transfer drug resistance to susceptible cells - a process that can easily take up to 14 days. It is generally agreed that this method is cumbersome. We sought to develop PCR-based protocols that would significantly simplify and shorten this procedure. Using PCR and primers specific for the Neoregion of the MLV-derived vector LeGSN, we determined 1. the proviral integration in target cells, and 2. the viral nucleic acid (RNA or DNA) content of the vector stock. Results were compared with those using the conventional method. We found that these specific PCR-based procedures were indeed useful for rapid determination of viral titers as well as for quick screening for high-titer vector-producing cell clones and successful transduction of target cells. PMID- 10698084 TI - Identification of a Salmonella typhimurium genomic region involved in invasion of HeLa and Henle-407 epithelial cells. AB - To identify the invasion determinant, a cosmid library was constructed by cloning a genomic library of Salmonella typhimurium 82/6915 into a cosmid vector, pLA2917. A genomic region involved in invasion of cultured HeLa and Henle-407 cells was subcloned into plasmid pGEM-7Z. E. coli strain DH1 carrying pSV6235 consisting of a S. typhimurium 4.6 kb genomic region in pGEM-7Z showed invasion of cultured HeLa and Henle-407 cells. Nested sequential deletions were introduced into the 4.6 kb genomic region of pSV6235. The E. coli recombinants which contained less than 1.5 kb deletions from the 5' end (SmaI site) of the genomic region invaded the cells as effectively as DH1 (pSV6235). The invasion of the recombinants carrying over 2.0 kb deletions from the end of pSV6235 was significantly inactivated compared to DH1 (pSV6235). Restriction enzyme analysis showed that the 3.1 kb fragment from the 3' end of the 4.6 kb genomic region was distinguished from the Salmonella pathogenicity I genes of S. typhimurium such as the inv, spa, and hil regions showing invasion of the cultured eukaryotic cells. PMID- 10698085 TI - Serum gastrin and gastrin-immunoreactive cells in the antral mucosa of patients with alcoholic liver disease. AB - BACKGROUND/AIMS: Hypergastrinaemia has been reported in liver cirrhosis; meanwhile, it is unclear whether it is associated with an increase in gastrin cell function. The serum gastrin concentration and the number of gastrin cells in antral biopsies were studied in patients with alcoholic liver disease. METHODS: Immunocytochemical and quantification techniques were used to localize and determine the number of gastrin cells. RESULTS: Slight non-significantly higher serum gastrin values were observed in the alcoholic liver disease patients compared with controls, but the individual variation within the groups was considerable. The frequency of gastrin cells did not differ between groups. However, the size of the gastrin cell nuclei was larger in patients with liver disease than in controls, indicating increased cellular activity. CONCLUSIONS: Alcoholic liver disease, with a disturbed liver function, influences the gastrin cells. The observed alterations may reflect the effect of alcohol and/or malnutrition, or may be secondary to the influence of liver disease on other regulatory peptides. PMID- 10698086 TI - Detection of Mycobacterium-specific interferon-gamma-producing human T lymphocytes by flow cytometry. AB - Flow cytometry has proven to be a useful tool for the investigation of cytokine synthesis by selected cell subpopulations. While most reports have used mitogen stimulation or long-term cultures with antigen, we describe here a novel method to allow the detection of rare mycobacterial antigen-specific cytokine synthesizing cells within one day. The most important feature of this method is the use of an FITC-conjugated isotype-matched control antibody to identify and exclude cells which fluoresce non-specifically. With this technique, we demonstrate interferon-gamma (IFN-gamma) staining in 785 cells per 1 x 10(5) T cells counted, in mycobacterial antigen-stimulated peripheral blood mononuclear cells from a BCG-vaccinated subject. In comparison, only 14 IFN-gamma-staining T cells were seen in the cultures not stimulated by mycobacterial antigen. Less than 10 cells per 1 x 10(5) T cells are stained by an irrelevant control antibody. Specific responses are detectable after 12 h of in vitro culture, and peak at 24 h. In volunteer health care workers, IFN-gamma staining correlated with IFN-gamma production using a published ELISPOT assay (r=0.927). IFN-gamma staining was also higher in PBMC from mantoux skin test-positive volunteers, compared to cells from skin test-negative subjects (p=0.0045). Flow cytometry following short-term culture can thus be used for enumeration of antigen-specific IFN-gamma synthesizing cells. PMID- 10698087 TI - Detection of vancomycin resistance genes combined with typing of Enterococci by means of multiplex PCR amplification and multiple primer DNA sequencing. AB - A multiplex PCR assay for the detection of vancomycin resistance (van) genes in enterococci was established. Primers targeting the 16S rRNA gene were included in the reaction mixture. Multiple-primer DNA sequencing of the PCR products provided species identification through partial nucleotide sequences of 16S rRNA genes, as well as confirmation of the correct identification of vanA, vanB, vanC-1, and vanC-2/3 genotypes. Thirty-nine enterococcal clinical isolates and type strains were examined for the presence of vancomycin resistance determinants. Twelve other isolates from a clinical reference collection (some of them having vanA, vanB, vanC-1, or vanC-2/3 genotypes) were used as controls. Hybridization and partial DNA sequence analysis of multiplex PCR products revealed that none of the clinical isolates had a vanA genotype and only one had a vanB genotype. vanC-1 was found in three clinical isolates, and vanC-2/3 in one. Results obtained with the reference and type strains were in agreement with earlier results. PMID- 10698088 TI - Sumatriptan increases the proliferation of peripheral blood mononuclear cells from HIV-infected individuals and healthy blood donors in vitro. AB - HIV infection is characterized by the loss of CD4+ T cells as well as the loss of T-cell function, leading to severe immunodeficiency. The proliferative capacity of T cells measured in vitro as responses to antigens and mitogens is severely reduced during HIV infection. An increased level of the intracellular second messenger adenosine 3',5'-cyclic monophosphate (cAMP) has been shown to cause impaired proliferative capacity of peripheral blood mononuclear cells (PBMC) from HIV-infected individuals in vitro. Sumatriptan, a 5HT1d receptor agonist, inhibits the activity of adenylyl cyclases, the enzymes responsible for regulation of the intracellular levels of cAMP. In a preliminary study sumatriptan increased the proliferative responses of PBMC to a polyclonal activator in vitro in 9 of 10 HIV-seropositive individuals (p=0.007), and in 7 of 9 healthy blood donors (p=0.05). This was probably due to a decrease in the intracellular level of cyclic AMP. PMID- 10698089 TI - Transfusion iron overload in adults with acute leukemia: manifestations and therapy. AB - BACKGROUND: Hepatic dysfunction occurs commonly among persons successfully treated for acute leukemia, and iron overload is a possible cause of hepatic and other abnormalities in these patients. METHODS: We identified 5 adults 40+/-13 (mean +/- S.D.) years of age who developed transfusion iron overload in association with successful chemotherapy of de novo acute leukemia. None had evidence of hemochromatosis, viral hepatitis, or other primary hepatic disorders. RESULTS: The mean serum ferritin concentration of our patients was 1531+/-572 ng/mL. Other abnormalities associated with transfusion iron overload were increased stainable iron in bone marrow macrophages, elevated serum concentrations of hepatic enzymes, hyperpigmentation, hyperferremia, elevated iron saturation of serum transferrin, and increased stainable iron in Kupffer cells and hepatocytes. Rheumatologic, endocrinologic, or cardiac abnormalities attributable to iron overload were not observed. Therapeutic phlebotomy was initiated after chemotherapy; recovery of hemoglobin concentrations after phlebotomy permitted weekly treatment in each case. This yielded an average of 28+/-9 units per patient (range 15-35 units). Abnormalities associated with transfusion iron overload resolved after iron depletion therapy. The mean leukemia-free survival of our patients is 96+/-36 months (range 40-125 months). CONCLUSIONS: Our data and those of others suggest that 15 to 20% of adults who are long-term survivors of acute leukemia develop iron overload, often with hepatic abnormalities. Iron overload is a relatively common sequela to successful management of acute leukemia in adults for which routine evaluation should be performed and for which therapeutic phlebotomy should be used as treatment. PMID- 10698090 TI - Immunohistochemical localization of modified C-reactive protein antigen in normal vascular tissue. AB - BACKGROUND: The prototypic acute phase reactant, C-reactive protein (CRP), is a serum soluble, cyclic pentameric protein, the concentration of which increases markedly within hours of any tissue-damaging, inflammatory event. However, upon dissociation of its pentameric quaternary structure, CRP subunits undergo a spontaneous and irreversible conformational change. The resulting molecule, termed modified CRP or mCRP, has reduced aqueous solubility and a propensity to aggregate into a matrix-like lattice structure. METHODS: Using monoclonal antibodies, normal human tissues were immunohistochemically screened for the presence of CRP as well as mCRP antigens. RESULTS: Significant levels of mCRP were detected in the walls of blood vessels associated with normal human tissues. These data indicate that mCRP is a naturally occurring form of CRP and that it is a tissue-based rather than serum-based molecule. SIGNIFICANCE: This report describes the localization of a stable form of CRP, mCRP, in blood vessels associated with normal human tissues. PMID- 10698091 TI - Coronary artery disease risk predicted by insulin resistance, plasma lipids, and hypertension in people without diabetes. AB - BACKGROUND: It has been shown that insulin resistance syndrome, including glucose intolerance, dyslipidemia, and hypertension, is frequently associated with coronary artery disease (CAD). However, their relative contributions and predictive power in the development of CAD are still unclear, particularly in persons without diabetes. METHOD: We examined these risk factors between 96 patients without diabetes but with angiographically documented CAD and 96 age-, sex-, and body mass index-matched healthy control subjects. Fasting plasma lipoprotein, glucose, and insulin concentrations in response to a 75-g oral glucose tolerance test were determined, and insulin sensitivity was measured by the insulin suppression test. RESULTS: Patients with CAD had significantly higher values of fasting glucose, glucose and insulin responses to oral glucose tolerance test, total cholesterol, low-density lipoprotein (LDL) cholesterol, and triglyceride and decreased high-density lipoprotein (HDL) cholesterol concentrations compared with those of healthy people (P < 0.02-0.001). Although the steady-state plasma insulin values were similar in both groups, the steady state plasma glucose (SSPG) concentrations were significantly higher in patients with CAD (12.2+/-0.4 versus 8.1+/-0.4 mmol/L, P < 0.001) compared with healthy subjects. When HDL < 0.9 mmol/L, LDL cholesterol > or = 4.1 mmol/L, triglyceride > or = 2.3 mmol/L, SSPG > or = 10.5 mmol/L, and presence of hypertension were defined as separate risk factors for CAD, significantly higher odds-ratio values were observed in patients with CAD compared with healthy people. From logistic multiple regression analysis, SSPG was the strongest risk, followed by lowered HDL cholesterol, elevated triglyceride and LDL cholesterol, and hypertension, to predict CAD. These 5 factors accounted for 36% of total risk for development of CAD in persons without diabetes. CONCLUSIONS: Patients without diabetes with CAD have abnormal glucose metabolism, hyperinsulinemia, and insulin resistance. Degree of insulin resistance (SSPG values), plasma lipid values, and history of hypertension together accounted for one third of all risk for CAD, although degree of insulin resistance was the strongest risk factor. PMID- 10698092 TI - Neurally mediated hypotension in fatigued Gulf War veterans: a preliminary report. AB - BACKGROUND: Many patients with chronic fatigue syndrome (CFS) have neurally mediated hypotension when subjected to head-up tilt, suggesting autonomic nervous system dysfunction. Some Gulf War veterans have symptoms similar to CFS. Whether they also tend to have neurally mediated hypotension is unknown. METHODS: We performed 3-stage tilt-table testing on 14 Gulf War veterans with chronic fatigue, 13 unfatigued control Gulf War veterans, and 14 unfatigued control subjects who did not serve in the Gulf War. Isoproterenol was used in stages 2 and 3 of the tilt protocol. RESULTS: More fatigued Gulf War veterans than unfatigued control subjects had hypotensive responses to tilt (P < 0.036). A positive response to the drug-free stage 1 of the tilt was observed in 4 of 14 fatigued Gulf War veterans versus 1 of 27 unfatigued control subjects (P < 0.012). Heart rate and heart rate variation during stage 1 was significantly greater in the fatigued group (P < 0.05). CONCLUSION: We conclude that more fatigued Gulf War veterans have neurally mediated hypotension than unfatigued control subjects, similar to observations in CFS. Autonomic nervous system dysfunction may be present in some fatigued Gulf War veterans. PMID- 10698094 TI - Candidiasis may induce glossodynia without objective manifestation. AB - BACKGROUND: The causes of glossodynia in the absence of objective abnormalities range widely and differential diagnosis of glossodynia is very difficult. METHODS: Based on the examination results of peripheral blood, stimulated and nonstimulated salivary flow rate (SFR), glossal pain threshold, and C. albicans cell culture and the response to treatment, we identified the cause of vague pain of the tongue in 98 patients who lacked objective findings and identified candidiasis as the cause of glossodynia in 26 patients. RESULTS: These patients revealed hyposalivation and decreased glossal pain thresholds and C. albicans cell overgrowth. Pain thresholds in the painful portion (54.6+/-2.9 degrees C) were significantly decreased compared with those in the painless portion (57.7+/ 3.4 degrees C) (P < 0.05) and the pain thresholds were largely increased after treatment (57.2+/-1.6 degrees C). Nonstimulated SFR before treatment was lower than that of age- and gender-matched healthy people, although stimulated SFR was decreased only slightly. C. albicans cell overgrowth was detected by the number of C. albicans colonies that formed in Sabouraud's agar plate (539.3+/ 198.4/dish). After the subsidence of glossal pain by mouth washing with a 3% amphotericin B solution, the C. albicans colonies were decreased to 31.5+/ 19.3/dish, which was almost same as the control level, 14.1+/-8.4/dish. CONCLUSION: These results indicate that candidiasis in conjunction with hyposalivation may induce pain in the tongue without manifestation of objective abnormalities. PMID- 10698093 TI - Propylthiouracil reduces xenograft tumor growth in an athymic nude mouse prostate cancer model. AB - METHODS: Several anecdotal reports indicate that cancer may occasionally remain in a dormant state for prolonged periods in patients with hypothyroidism. Once the hypothyroid state is recognized and supplementation therapy with thyroid hormones is initiated, disease progression occurs. In this experiment, 6-n-propyl 2-thiouracil (PTU) was added to the water of athymic nude mice. The animals were subsequently inoculated with cells from a human prostate cancer cell line. RESULTS: The growth rate of subcutaneously implanted prostate xenografts was significantly slower in mice treated with PTU compared with mice that did not receive PTU. In a separate experiment, tritiated thymidine incorporation assays were performed in DU145 and PC3 human prostate cancer cells with and without PTU. No significant differences were observed, indicating that PTU did not exert any antitumor effect in vitro. CONCLUSIONS: Our study demonstrates that PTU inhibits the growth of human prostate tumors in nude mice via an indirect effect. This antitumor effect may be caused by hypothyroidism. This is the first in vivo study suggesting potential therapeutic applications for thyroid hormone manipulations in human cancer of the prostate. Further studies will determine growth kinetics of xenotransplanted prostate cancer in vitro and in vivo. PTU-induced hypothyroidism may be further explored in conjunction with other antineoplastic therapy. PMID- 10698095 TI - Beyond diagnosis: patient mix and challenges to patient care in ambulatory training sites. AB - BACKGROUND: Educational experiences in ambulatory medicine vary by site. PURPOSE: To evaluate variations in patient mix and challenges in patient care at 3 ambulatory training sites. METHOD: Patients (n=123) receiving care at a VA medical center (VA), an urban academic medical center's primary care center (PCC), and a community group practice (CGP) were evaluated. RESULTS: Patients at the VA (n=55), compared with those at the PCC (n=44) and the CGP (n=24), were older, more likely to be male, and white (all P<0.05). Patients at the VA and the PCC reported difficulty with functional and health status more frequently than those at the CGP (all P<0.05). Common medical diagnoses varied across sites and comorbidity scores were > or = 2 in 48% of VA subjects compared with 16% at the PCC and 29% at the CGP (P<0.05). Challenges most frequently cited were administrative issues at the VA (44%), patient-physician communication at the PCC (39%), and medical decisions at the CGP (50%) (P<0.05). CONCLUSIONS: Ambulatory training sites can differ greatly with respect to characteristics beyond diagnosis. Plans for increased and improved ambulatory training of internal medicine residents should include attention to these issues to ensure exposure to an adequate patient spectrum. PMID- 10698096 TI - Use of intraluminal stents in the treatment of carotid, renal, and peripheral arterial disease. PMID- 10698098 TI - Rapid communication: Cervical Chlamydia trachomatis in women at low risk for infection. AB - BACKGROUND: Evidence suggests that the bacterium Chlamydia trachomatis can cause asymptomatic genital infection in persons at risk for acquisition of the organism. We employed 2 independent molecular screening systems to assess such inapparent cervical chlamydial infections in low-risk female patients attending general (non-STD) clinics in 2 locations. METHODS: Three hundred seventy-five cervical swab samples were obtained in duplicate from patients attending a general women's clinic (278 samples) and a colposcopy clinic (97 samples). One set of samples from the general clinic was screened by a highly-specific molecular hybridization system, using a probe targeting the chlamydial 16S ribosomal RNA; the other set was screened with the use of the Chlamydiazyme test. Samples from the colposcopy clinic were screened using a sensitive and specific polymerase chain reaction (PCR) assay system targeting chlamydia; the duplicates were assayed by direct fluorescent antibody assay (DFA). RESULTS: Of the 278 patients screened by RNA-directed hybridization, 6.5% were positive for C. trachomatis, in contrast to screening of duplicate samples via Chlamydiazyme, which indicated that 3.6% were infected. PCR-based screening of the additional 97 patients gave a positivity rate of 17.5% for the organism, whereas DFA on duplicate samples from this group showed only 7.5% positive. CONCLUSIONS: These observations suggest that the level of asymptomatic cervical C. trachomatis infection is significant even in women who are at low risk for such infections; the data also indicate that results from standard laboratory screening for chlamydia should be viewed with caution. PMID- 10698097 TI - Metastatic prostate cancer (with prostate-specific antigen of 9996) presenting as obstructive jaundice. AB - A 78-year-old man admitted with clinical jaundice and pelvic pain had a total bilirubin level of 6.56 mg/dL, an alkaline phosphatase level of 855 U/L, and a prostate specific antigen (PSA) level of 9996 ng/mL. A computed tomogram demonstrated marked retroperitoneal, peripancreatic, periceliac, and periaortic lymphadenopathy. A bone scan revealed increased radiolabeled technetium uptake in the pelvis, vertebral column, parietooccipital region, ribs, and appendiceal skeleton. A biopsy of one pelvic lesion revealed metastatic prostate cancer. This man's obstructive jaundice and bone pain had a dramatic response to treatment with a gonadotropin-releasing hormone analog (leupro lide) and antiandrogen (bicalutamide). All bone pair and clinical signs of jaundice disappeared in 1 week His total bilirubin decreased to 0.84 mg/dL by 2 weeks His PSA values reflected this clinical response, decreasing to 4022 ng/mL in 1 week, 2680 ng/dL after 2 weeks and 1028 ng/mL after 1 month of the above therapy. PMID- 10698099 TI - Acute fulminant lactic acidosis complicating metastatic cholangiocarcinoma. AB - A patient with cholangiocarcinoma, metastatic to the liver and lungs, developed acute fulminant lactic acidosis in the absence of overt hepatic failure, sepsis, hypoxia, or circulatory failure. Despite extensive tumor replacement of hepatic parenchyma, no acid-base disorder was present during initial evaluation. The onset of acute lactic acidosis was temporally associated with the development of otherwise asymptomatic episodes of intermittent atrial arrhythmias. Once established, lactic acidosis was inexorably progressive, despite resolution of arrhythmias. Extensive areas of acute necrosis within the large hepatic metastases were demonstrated on postmortem examination, suggesting that local tissue ischemia, precipitated by cardiac arrhythmias, lead to excessive lactic acid production. PMID- 10698100 TI - Genetic and biochemical manipulations of the small ribosomal subunit from Thermus thermophilus HB8. AB - Crystals of the small ribosomal subunit from Thermus thermophilus diffract to 3A and exhibit reasonable isomorphism and moderate resistance to irradiation. A 5A MIR map of this particle shows a similar shape to the part assigned to this particle within the cryo-EM reconstructions of the whole ribosome and contains regions interpretable either as RNA chains or as protein motifs. To assist phasing at higher resolution we introduced recombinant methods aimed at extensive selenation for MAD phasing. We are focusing on several ribosomal proteins that can be quantitatively detached by chemical means. These proteins can be modified and subsequently reconstituted into depleted ribosomal cores. They also can be used for binding heavy atoms, by incorporating chemically reactive binding sites, such as -SH groups, into them. In parallel we are co-crystallizing the ribosomal particles with tailor made ligands, such as antibiotics or cDNA to which heavy atoms have been attached or diffuse the latter compounds into already formed crystals. PMID- 10698101 TI - Structural properties of hybrid triplex of polycation deoxyribonucleic S methylthiourea (DNmt) strands with a complementary DNA strand, probed by nanosecond molecular dynamics. AB - The replacement of phosphodiester linkages of the polyanion DNA with S methylthiourea linkers provides the polycation deoxyribonucleic S-methylthiourea (DNmt). Molecular dynamics studies to 1,220 ps of the hybrid triplex formed from octameric DNmt strands d(Tmt)8 with a complementary DNA oligomer strand d(Ap)8 have been carried out with explicit water solvent and Na+Cl- counterions under periodic boundary conditions using the CHARMM force field and the Ewald summation method. The Watson-Crick and Hoogsteen hydrogen-bonding patterns of the A/T tracts remained intact without any structural restraints for triplex structures throughout the simulation. The duplex portion of the triplex structure equilibrated at a B-DNA conformation in terms of the helical rise and other helical parameters. The dynamic structures of the DNmt x DNA x DNmt triplex were determined by examining histograms from the last 800 ps of the dynamics run. These included the hydrogen-bonding pattern (sequence recognition), three centered bifurcating occurrences, minor groove width variations, and bending of tracts for the hybrid triplex structures. Together with the Watson-Crick hydrogen bondings, the strong Hoogsteen hydrogen-bondings, the partially maintained three centered bifurcatings in the Watson-Crick pair, and the medium-strength three centered bifurcatings in the Hoogsteen pair suggest that the hybrid triplex is energetically favorable as compared to a duplex with similar base stacking, van der Waals interactions, and helical parameters. This is in agreement with our previously reported thermodynamic study, in which only triplex structures were observed in solution. The bending angle measured between the local axis vectors of the first and last helical axis segments is about 20 degrees for the Watson Crick portion of the averaged structure. Propeller twist (associated with three centered hydrogen-bonding) up to -30 degrees, native to DNA AT base pairing, was also observed for the triplex structure. The sugar pseudorotation phase angles and the ring rotation angles for the DNA strand are within the C3'-endo domain and C2'-endo domain for the DNmt strand. Water spines are observed in both minor and major grooves throughout the dynamics run. The molecular dynamics simulations of the structural properties of DNmt x DNA x DNmt hybrid triplex is compared to the DNG x DNA x DNG hybrid triplex (In DNG the -O-(PO2-)-O- linkers in DNA is replaced by -NH-C(=N+H2)-NH-). PMID- 10698102 TI - The impact of abasic sites on DNA flexibility. AB - We use internal coordinate molecular mechanics calculations to study the impact of abasic sites on the conformation and the mechanics of the DNA double helix. Abasic sites, which are common mutagenic lesions, are shown to locally modify both the groove geometry and the curvature of DNA in a sequence dependent manner. By controlled twisting and bending, it is also shown that these lesions modify the deformability of the duplex, generally increasing its flexibility, but again to an extent which depends on the nature of the abasic site and on the surrounding base sequence. Both the conformational and mechanical influence of this type of DNA damage may be significant for recognition and repair mechanisms. PMID- 10698103 TI - Structural polymorphism of oligo(dC) with mixed alpha,beta-anomeric backbone. AB - Oligonucleotides with mixed alpha,beta-anomeric backbone have been proposed recently for the recognition of random DNA sequence via new triplex motif (Doronina and Behr, Chem. Soc. Reviews 26, 63-71 (1997)). In the present work we examined alpha- and beta- anomers of cytidine as possible candidates to recognize AT and TA base pairs of the double stranded DNA. The binding properties of beta oligo(dC) were studied on a series of synthetic oligodeoxynucleotides by UV absorbtion spectroscopy, measurements of bound EtBr fluorescence polarization, circular dichroism (CD) and non-denaturing gel electrophoresis. The UV thermal denaturation, polarization studies and CD experiments with three stranded oligonucleotide 5'-((dCalpha) (dCbeta))5-L-(dAT)5-L-(dAT)5 (L = triethyleneglycol linker) and other oligonucleotide models showed that the formation of semiprotonated oligocytidilic complexes takes place at low temperatures and neutral pH, rather than folding of the clip into intramolecular triplex. The low temperature transition was observed in denaturation profiles of any oligonucleotide containing beta- or mixed alpha,beta- cytidine stretches at the concentration of 1 microM. Self-association of alpha,beta-oligo(dC) was additionally confirmed by the appearance of two CD bands (at 290 and 265 nm) characteristic of CC+ base pairs. Despite the effective ability of alpha,beta oligo(dC) to form self-associates, we succeeded in targeting 30-bp AT containing random DNA duplex by a 30-nt alpha,beta-oligocytidilate as evidenced by non denaturing gel electrophoresis. A complete binding of the duplex was observed at a 5-fold excess of the third strand at 15 degrees C. Along with the formation of the three-stranded complex, self-association of mixed backbone oligo(dC) strands occurred. PMID- 10698104 TI - Optimum DNA curvature using a hybrid approach involving an artificial neural network and genetic algorithm. AB - In the present paper, a hybrid technique involving artificial neural network (ANN) and genetic algorithm (GA) has been proposed for performing modeling and optimization of complex biological systems. In this approach, first an ANN approximates (models) the nonlinear relationship(s) existing between its input and output example data sets. Next, the GA, which is a stochastic optimization technique, searches the input space of the ANN with a view to optimize the ANN output. The efficacy of this formalism has been tested by conducting a case study involving optimization of DNA curvature characterized in terms of the RL value. Using the ANN-GA methodology, a number of sequences possessing high RL values have been obtained and analyzed to verify the existence of features known to be responsible for the occurrence of curvature. A couple of sequences have also been tested experimentally. The experimental results validate qualitatively and also near-quantitatively, the solutions obtained using the hybrid formalism. The ANN GA technique is a useful tool to obtain, ahead of experimentation, sequences that yield high RL values. The methodology is a general one and can be suitably employed for optimizing any other biological feature. PMID- 10698105 TI - Dynamics of a double stranded DNA oligomer: mode-coupling diffusion approach and reduced rigid fragment models. AB - The local dynamics of a double stranded DNA fragment [d(CpGpCpApApApTpTpTpGpCpG)]2 of twelve base pairs is obtained to second order in the mode-coupling expansion of the Smoluchowski diffusion theory. The DNA is considered a fluctuating three-dimensional (3D) structure undergoing rotational diffusion. The starting structure for the calculations is the B canonical structure of the fragment, while the fluctuations are evaluated using molecular dynamics simulations, with the ensemble averages approximated by time averages along a trajectory of length 1.5 ns. The rotational dynamics of the bonds along the double strands are calculated and compared to experimental NMR relaxation rates of different 13C along the sequence: R(Cz), R(Cxy) and R(Hz-->Cz). For a fluctuating 3D structure the mode-coupling diffusion theory is found to be in good agreement with several relative characteristics of the experimental relaxation parameters, while motivations are given for the few differences which are due mainly to poor statistics or to inaccuracies in the diffusion model. With a view to application to larger DNA fragments, discussion is dedicated to the validity of reducing the number of degrees of freedom in the double helix statistics by grouping the atoms in rigid fragments (e.g. the backbone atoms, the sugar atoms and the base atoms of each nucleotide). Consideration is given to the effect on local dynamics properties of reduced descriptions that include only three or four rigid bodies per nucleotide as well as five rigid bodies per base pair. It is found that in general these approximations almost uniformly produce slight increase in the correlation time pattern, which grows as the rigidity in the model increases. The relative effects on the dynamic pattern for the most accurate rigid body models are modest. The errors in C1' and C5' mobilities are more significant if C5' is included in the backbone rigid body. These results offer new tools to analyse NMR relaxation behaviour and new perspectives in studying the role of dynamics in biological macromolecules. PMID- 10698106 TI - Atomic force and electron microscopy of high molecular weight circular DNA complexes with synthetic oligopeptide trivaline. AB - Intramolecular compact structures formed by high molecular weight circular superhelical DNA molecules due to interaction with synthetic oligopeptide trivaline (1) were studied by atomic force and electron microscopy. Three DNA preparations were used: plasmids pTbol, pRX10 and cosmid 27,877, with sizes 6,120 bp, 10,500 bp and 44,890 bp respectively. Plasmid pTbo1 and pRX10 preparations along with monomers contained significant amount of dimers and trimers. Main structures in all preparations observed were compact particles, which coincide in their appearance and compaction coefficient (3,5-3,7) with triple rings described earlier. The size and structure characteristics of triple rings and other compact particles on atomic force images in general coincide with those obtained by EM (2). AFM (3) images allow to get additional information about the ultrastructural organization and arrangement of DNA fibers within the compact structures. Along with triple rings in pTbol and pRX10-TVP complexes significant amount of compact structures were observed having the shape of two or three compact rings attached to each other by a region of compact fibre. Basing on the data of contour length measurements and the shape of the particles it was concluded that these structures were formed due to compaction of dimeric and trimeric circular DNA molecules. Structures consisting of several attached to each other triple rings were not found for pTbol, pRX10 monomers or cosmid preparations--TVP complexes where only single triple rings were observed. The conclusion is made that initiation of compact fibre formation within the circular molecules depends on the primary structure and for dimeric or trimeric circular molecules two or three compaction initiation points are present, located in each monomer unit within one circular DNA molecule. The nucleotide sequence dependent compaction mechanism providing independent compaction of portions of one circular molecule can be of interest for understanding of DNA compaction processes in vivo. PMID- 10698107 TI - Melting of cross-linked DNA IV. Methods for computer modeling of total influence on DNA melting of monofunctional adducts, intrastrand and interstrand cross-links formed by molecules of an antitumor drug. AB - A theoretical method is developed for calculation of melting curves of covalent complexes of DNA with antitumor drugs. The method takes into account all the types of chemical modifications of the double helix caused by platinum compounds and DNA alkylating agents: 1) monofunctional adducts bound to one nucleotide; 2) intrastrand cross-links which appear due to bidentate binding of a drug molecule to two nucleotides that are included into the same DNA strand; 3) interstrand cross-links caused by bidentate binding of a molecule to two nucleotides of different strands. The developed calculation method takes into account the following double helix alterations at sites of chemical modifications: 1) a change in stability of chemically modified base pairs and neighboring ones, that is caused by all the types of chemical modifications; 2) a change in the energy of boundaries between helical and melted regions at sites of chemical modification (local alteration of the factor of cooperativity of DNA melting), that is caused by all the types of chemical modifications, too; 3) a change in the loop entropy factor of melted regions that include interstrand cross-links; 4) the prohibition of divergence of DNA strands in completely melted DNA molecules, which is caused by interstrand cross-links only. General equations are derived, and three calculation methods are proposed to calculate DNA melting curves and the parameters that characterize the helix-coil transition. PMID- 10698109 TI - Docking of peptide-T onto the D1 domain of the CD4 receptor. AB - Peptide T (pepT) is a segment of the human immunodeficiency virus (HIV) envelope protein gp120. The peptide competitively binds to the CD4 receptor of a subset of peripheral T lymphocytes and inhibits binding of gp120. Previous studies of this laboratory allowed the assessment of a bioactive form of the peptide and a pharmacophore for the peptide-receptor interaction. In the present study the proposed bioactive form of pepT and its (4-8) segment, the smallest pepT fragment shown to retain full activity, were docked onto the D1 domain of the CD4 receptor. The bioactive conformation of the peptides complements well a cleft on the surface of the CD4 receptor, shown to be the attachment site of gp120 from site directed mutagenesis experiments. These studies provide an improved description of the ligand-receptor pharmacophore. PMID- 10698108 TI - 15N-enriched 5-fluorocytosine as a probe for examining unusual DNA structures. AB - A new method is presented for the synthesis of oligonucleotides containing 15N enriched 5-fluorocytosine (FC). Due to the reduced pK of FC, the amino protons of an unpaired FC residue may be observed at lower values of solution pH. The labeled FC residue has been placed as a template base at a model DNA replication fork. The amino protons of the FC residue have been identified in isotope-edited NMR spectra. Data is presented for a template FC residue unpaired, paired with guanine, and mispaired with adenine. These studies demonstrate the utility of labeled FC in examining unusual DNA structures. PMID- 10698110 TI - Modeling the correlation functions of conformational motions in proteins. AB - A first principles calculation of the correlation function for conformational motion (CM) in proteins is carried out within the framework of a microscopic model of a protein as a heterogeneous system. The fragments of the protein are assumed to be identical hard spheres undergoing the CM within their conformational potentials about some mean equilibrium positions assigned by the tertiary structure. The memory friction function (MFF) for the generalized Langevin equation describing the CM of the particle is obtained on the basis of the direct calculation which is feasible for the present model of the protein due to the existence of a natural large parameter, viz. the ratio of the minimal distance between the mean equilibrium positions of the particles (approximately 7A) to the amplitude of their CM (<1A). A relationship between the MFF and the correlation functions of the CM of the particles is derived which makes their calculation to be a self-consistent mathematical problem. The general analysis of the MFF is exemplified by a simple model case in which the mean equilibrium positions of the particles form a regular lattice so that the correlation functions for all particles are the same. In this case the MFF is shown to be an infinite series of the powers of the auto-correlation function whose coefficients are independent on temperature. The latter is a result of the abstraction of the interaction potential by that of hard spheres which actually corresponds to the high temperature limit. On the examples of cubic and triangular lattices the coefficients are shown to be non-negative values which increase with the increase of the packing density of the particles and quickly tend to zero with the increase of their index. Thus the MFF can be approximated by a polynomial of the correlation function and the resulting mathematical equation is analogous to the one from the dynamic theory of liquids. The correlation function of the CM is obtained by numerical solution of the equation. At realistic packing densities for proteins it exhibits transparent non-exponential decay and includes two relaxation processes: the first one on the intermediate timescale (tens of picoseconds) and the second on the long timescale (its characteristic time is about tens of nanoseconds at small values of the friction coefficient and increases by orders of the magnitude with the increase of the latter). Thus the present approach provides the microscopic basis for previous phenomenological models of cooperative dynamics in proteins. PMID- 10698111 TI - Topology prediction of Brucella abortus Omp2b and Omp2a porins after critical assessment of transmembrane beta strands prediction by several secondary structure prediction methods. AB - In order to propose a reliable model for Brucella porin topology, several structure prediction methods were evaluated in their ability to predict porin topology. Four porins of known structure were selected as test-cases and their secondary structure delineated. The specificity and sensitivity of 11 methods were separately evaluated. Our critical assessment shows that some secondary structure prediction methods (PHD, Dsc, Sopma) originally designed to predict globular protein structure are useful on porin topology prediction. The overall best prediction is obtained by combining these three "generalist" methods with a transmembrane beta strand prediction technique. This "consensus" method was applied to Brucella porins Omp2b and Omp2a, sharing no sequence homology with any other porin. The predicted topology is a 16-stranded antiparallel beta barrel with Omp2a showing a higher number of negatively charged residue in the exposed loops than Omp2b. Experiments are in progress to validate the proposed topology and the functional hypotheses. The ability of the proposed consensus method to predict topology of complex outer membrane protein is briefly discussed. PMID- 10698112 TI - Cytochrome P450 catalyzed nitric oxide synthesis: a theoretical study. AB - Similar to nitric oxide synthase (NOS) cytochrome P450 isoforms (e.g. 3A and 4E) can produce nitric oxide from arginine. Although the active site of both proteins contains a protoporphyrin IX unit having an axial cystein ligand, their effectiveness in the synthesis of NO differs significantly. Now the molecular basis of this functional difference was investigated. A homology model for cytochrome P450 3A4 was refined and compared to the X-ray structure of iNOS. We found the active site of iNOS to be more readily accessible for the substrate than that of P450. Docking calculations were performed using the Monte Carlo conformational analysis technique on all internal and external degrees of freedom of arginine and active site residues as well. The lowest energy conformation of the cytochrome P450 3A4-substrate complex was compared to the high resolution X ray structure of the iNOS-arginine complex. Comparison of substrate orientations revealed that arginine binds in a similar conformation in both enzymes. In contrast to iNOS we found, however, that in P450 partially negative propionate side chains of protoporphyrin IX are located on the opposite side of the heme plane. As a result of this and the absence of other negatively charged residues the distal (substrate binding) side of P450 should be less negative than that of NOS and therefore its affinity toward the partially positive arginine is reduced. Comparison of molecular electrostatic potentials calculated within the active site of the proteins supports this proposal. Reduced affinity in combination with limited substrate access might be responsible for the less effective NO synthesis of cytochrome P450 observed experimentally. PMID- 10698113 TI - Cloning of a neoteleost (Oreochromis mossambicus) pro-opiomelanocortin (POMC) cDNA reveals a deletion of the gamma-melanotropin region and most of the joining peptide region: implications for POMC processing. AB - A signature feature of tetrapod pro-opiomelanocortin (POMC) is the presence of three melantropin (MSH) coding regions (alpha-MSH, beta-MSH, gamma-MSH). The MSH duplication events occurred early during the radiation of the jawed vertebrates well over 400 million years ago. However, in at least one order of modern bony fish (subdivision Teleostei; order Salmoniformes; i.e. salmon and trout) the gamma-MSH sequence has been deleted from POMC. To determine whether the gamma-MSH deletion has occurred in other teleost orders, a POMC cDNA was cloned from the pituitary of the neoteleost Oreochromis mossambicus (order Perciformes). In O. mossambicus POMC, the deletion is more extensive and includes the gamma-MSH sequence and most of the joining peptide region. Because the salmoniform and perciform teleosts do not share a direct common ancestor, the gamma-MSH deletion event must have occurred early in the evolution of the neoteleost fishes. The post-translational processing of O. mossambicus POMC occurs despite the fact that the proteolytic recognition sequence, (R/K)-Xn-(R/K) where n can be 0, 2, 4, or 6, a common feature in mammalian neuropeptide and polypeptide hormone precursors, is not present at several cleavage sites in O. mossambicus POMC. These observations would indicate that either the prohormone convertases in teleost fish use distinct recognition sequences or vertebrate prohormone convertases are capable of recognizing a greater number of primary sequence motifs around proteolytic cleavage sites. PMID- 10698114 TI - Characterization of C-terminal-truncated urinary metabolites of a recombinant hirudin in rats. AB - Although it has been reported that hirudin was excreted in urine mainly as its nonmetabolized form in humans, dogs, and rabbits, no report has been published about the molecular nature of urinary metabolites in rats. We found that nonmetabolized hirudin could not be detected in rat urine after its i.v. administration and that urinary metabolites of recombinant hirudin CX-397 consisted of at least the following six C-terminal-truncated peptides: CX-397(1 49), CX-397(1-50), CX-397(1-51), CX-397(1-52), CX-397(1-54), and CX-397(1-55), in the ratio of roughly 11, 51, 3, 11, 19, and 5%, respectively. In conclusion, the urinary metabolism of recombinant hirudin in rats is different from that in humans, dogs, and rabbits, suggesting that the handling of hirudin in rat kidney is unique among them. PMID- 10698115 TI - The modulation of skeletal muscle contraction by FMRFamide-related peptides of the locust. AB - The external ventral protractor muscle of the VIIth abdominal segment, M234, is a skeletal muscle that possesses receptors that recognize a range of FMRFamide related peptides and application of these peptides results in an increase in the amplitude of neurally evoked contractions with little or no effect on basal tonus. FLRFamide itself has the same biologic activity as the extended peptides, whereas truncation to LRFamide or RFamide results in a peptide with no biologic activity. The receptors recognizing these extended FLRFamides, which include SchistoFLRFamide, seem to be different from the SchistoFLRFamide receptors found on locust oviduct visceral muscle. SchistoFLRFamide and the non-peptide mimetic, benzethonium chloride (Bztc), increase the frequency and amplitude of miniature endplate potentials, increase the amplitude of neurally evoked excitatory junction potentials, and result in a hyperpolarisation of resting membrane potential. Bztc, however, also abolishes the active membrane response that may explain its ability to decrease neurally evoked contractions. PMID- 10698116 TI - Chronic exposure to antibodies directed against anti-opiate peptides alter delta opioid receptor levels. AB - The development of addictive states in response to chronic opioid use may be regulated partially by the release of endogenous peptides. These anti-opiate peptides (AOP) are secreted or released into the CNS and produce diverse actions that counterbalance the effects of prolonged opiate exposure. Though the mechanism(s) by which these peptides exert their physiological properties remain largely unknown, there is some indication that AOP's modulate opioid receptor levels. In this study, we investigated the effects of chronically infused alpha melanocyte stimulating hormone (alpha-MSH), dynorphin(1-8) (DYN(1-8)), dynorphin A (DYNA), and NPFF antibodies on delta-opioid receptor expression in rat brains. Quantitative autoradiographic experiments revealed that antibodies directed against alpha-MSH and DYNA produced significant increases in delta receptor levels in the caudate, claustrum, and cingulate cortex of the rat brain. Conversely, NPFF monoclonal antibodies caused significant decreases in the caudate, nucleus accumbens, olfactory tubercle, and cingulate cortex. These results suggest that the density of delta-opioid receptors is affected by changes in the levels of the anti-opioid peptides in the extracelluar fluid in the rat brain. PMID- 10698117 TI - Effects of neonatal treatment with Tyr-MIF-1 and naloxone on the long-term body weight gain induced by repeated postnatal stressful stimuli. AB - Stressful stimuli repeatedly applied during the first postnatal weeks can induce body weight gain in the mouse during adulthood. This effect can be prevented by injecting naloxone concomitantly with stress. The peptides belonging to the Tyr MIF-1 family have a great modulating activity on numerous stress-induced phenomena. The aim of the present work was to compare the effect of repeated neonatal injections of Tyr-MIF-1 or naloxone on the long-term body weight gain induced by a stressing procedure applied daily during the first three weeks of life. The results indicate that although naloxone blocked the development of the stress-induced effects, Tyr-MIF-1 potentiated them. PMID- 10698118 TI - Reduction of pentylenetetrazol-induced convulsions by the octadecaneuropeptide ODN. AB - Intracerebroventricular injection of the octadecaneuropeptide ODN in mouse, at doses of 12.5-1000 ng, reduced the percentage of convulsing animals and increased the latency of convulsions elicited by pentylenetetrazol (50 mg/kg, intraperitoneal [i.p.]). ODN also reduced the percentage of mortality induced by pentylenetetrazol (100 mg/kg, i.p.). The COOH-terminal octapeptide fragment of ODN was approximately equally effective but acted more rapidly than ODN to reverse the convulsant effect of pentylenetetrazol. ODN (100 ng, intracerebroventricular [i.c.v.]) increased the convulsion latency and reduced the percentage of animals that convulsed after the administration of the inverse agonist of benzodiazepine receptors DMCM (13 mg/kg, i.p.), whereas the benzodiazepine receptor antagonist flumazenil (1 mg/kg, subcutaneously) abrogated the protective effect of ODN (100 ng, i.c.v.) on pentylenetetrazol-induced convulsions. ODN (100 ng, i.c.v.) also reduced the percentage of DBA/2J mice displaying audiogenic convulsions. In contrast, ODN did not reduce the percentage of mice displaying tonic or clonic convulsions when electrical interauricular stimulations were applied. It is concluded that ODN, or more likely a proteolytic fragment derived from ODN, reduces pentylenetetrazol-induced convulsions through activation of central-type benzodiazepine receptors. PMID- 10698119 TI - Anxiogenic effects of substance P and its 7-11 C terminal, but not the 1-7 N terminal, injected into the dorsal periaqueductal gray. AB - The dorsal periaqueductal gray matter (DPAG) is one of the main output regions of the brainstem for the expression of defense reaction. Recent findings implicating neurokinins in the expression of fear or anxiety-like behaviors, have stimulated interest in the participation of these neuropeptides in the generation of aversive states in the dorsal periaqueductal gray matter. Analyses of traditional measures of the behavior of rats submitted to the elevated plus-maze test in this laboratory have shown that microinjections of substance P (SP) into the DPAG produce anxiogenic-like effects. The present study employs an ethological analysis of the behavior of animals in this test to investigate the involvement of substance P (SP) and its C- and N- fragments (7-11 and 1-7) in the expression of the different aspects of fear upon injection into the DPAG. To this end, rats were implanted with a cannula in the DPAG and injected one week later with 35 and 70 pmol of either substance P, or C- or N- SP fragments and tested immediately afterwards in the elevated plus-maze. The results show that SP and its C terminal fragment, produced increases in scanning, stretched attend posture, head dipping and flat-back approach, whereas the fragment N terminal produced only an increase in rearing. Therefore, the effects of SP and its C terminal fragment were associated to risk assessment behavior, whereas those of N terminal fragment were related to vertical exploratory activity. The results indicate that SP produces anxiogenic effects through activation of neural substrates of aversion in the DPAG and that this effect is probably related to its C terminal fragment. PMID- 10698120 TI - Mouse pancreatic polypeptide modulates food intake, while not influencing anxiety in mice. AB - This study was designed to investigate the effects of synthetic mouse pancreatic polypeptide (mPP) on feeding and anxiety in mice. The intracerebroventricular (i.c.v.) injection of mPP (0.003-3 nmol) dose-dependently increased food intake. A significant increase was observed 20 min after i.c.v. injection and continued for 4 h. The intraperitoneal (i.p.) injection of mPP (0.03-30 nmol) dose dependently decreased food intake. A significant decrease was observed 20 min after i.p. injection and continued for 4 h. In the elevated plus maze test, the i.c.v. injection of mPP (0.003-3 nmol) did not affect anxiety behavior. These results suggest that mPP modulates food intake and the Y4 receptor in the brain may contribute to the regulation of feeding, whereas appearing not to influence anxiety in mice. PMID- 10698121 TI - Decreased transport of leptin across the blood-brain barrier in rats lacking the short form of the leptin receptor. AB - Leptin is produced in adipose tissue in the periphery, but its satiety effect is exerted in the CNS that it reaches by a saturable transport system across the blood-brain barrier (BBB). The short form of the leptin receptor has been hypothesized to be the transporter, with impaired transport of leptin being implicated in obesity. In Koletsky rats, the splice variant that gives rise to the short form of the leptin receptor contains a point mutation that results in marked obesity. We studied the transport of leptin across the BBB in Koletsky rats and found it to be significantly less than in their lean littermates. By contrast, Sprague-Dawley rats matched in weight to each of these two groups showed no difference in the blood-to-brain influx of leptin. HPLC showed that most of the leptin crossing the BBB in rats remained intact and capillary depletion showed that most of the leptin reached the parenchyma of the brain. The results indicate that the short form of the leptin receptor is involved in the transport of leptin across the BBB. PMID- 10698122 TI - Differential distribution of orexin-A and orexin-B immunoreactivity in the rat brain and spinal cord. AB - The orexins are recently identified appetite-stimulating hypothalamic peptides. We used immunohistochemistry to map orexin-A and orexin-B immunoreactivity in rat brain, spinal cord, and some peripheral tissues. Orexin-A- and orexin-B immunoreactive cell bodies were confined to the lateral hypothalamic area and perifornical nuclei. Orexin-A-immunoreactive fibers were densely distributed in the hypothalamus, septum, thalamus, locus coeruleus, spinal cord, and near the ventricles, but absent from peripheral sites investigated. In contrast, orexin-B immunoreactive fibers were distributed sparsely in the hypothalamus. Orexin cells are strategically sited to contribute to feeding regulation, but their widespread projections suggest that orexins have other physiological roles. PMID- 10698123 TI - Is adrenomedullin a causal agent in some cases of type 2 diabetes? AB - The study of two populations with a recent onset of type 2 diabetes showed that a subset of the patients had higher levels of adrenomedullin (AM) than the rest of the diabetics. In this subset, physiological elevations of AM might have triggered the disease in predisposed individuals. Diabetics showed higher levels of AM than healthy controls. In addition, glycemia was measured in diabetic rats after injection of saline, AM, or antiAM antibody. AM elevated glycemia, whereas the antibody reduced circulating glucose to normal. These results suggest that manipulation of AM levels could represent a new approach in the management of diabetes for the appropriate individuals. PMID- 10698124 TI - Distribution, functional role, and signaling mechanism of adrenomedullin receptors in the rat adrenal gland. AB - Adrenomedullin (ADM) is a hypotensive peptide, highly expressed in the mammalian adrenal medulla, which belongs to a peptide superfamily including calcitonin gene related peptide (CGRP) and amylin. Quantitative autoradiography demonstrated the presence of abundant [125I]ADM binding sites in both zona glomerulosa (ZG) and adrenal medulla. ADM binding was selectively displaced by ADM(22-52), a putative ADM-receptor antagonist, and CGRP(8-37), a ligand that preferentially antagonizes the CGRP1-receptor subtype. ADM concentration-dependently inhibited K+-induced aldosterone secretion of dispersed rat ZG cells, without affecting basal hormone production. Both ADM(22-52) and CGRP(8-37) reversed the ADM effect in a concentration-dependent manner. ADM counteracted the aldosterone secretagogue action of the voltage-gated Ca2+-channel activator BAYK-8644, and blocked K+- and BAYK-8644-evoked rise in the intracellular Ca2+ concentration of dispersed ZG cells. ADM concentration-dependently raised basal catecholamine (epinephrine and norepinephrine) release by rat adrenomedullary fragments, and again the response was blocked by both ADM(22-52) and CGRP(8-37). ADM increased cyclic-AMP release by adrenal-medulla fragments, but not capsule-ZG preparations, and the catecholamine response to ADM was abolished by the PKA inhibitor H-89. Collectively, the present findings allow us to draw the following conclusions: (1) ADM modulates rat adrenal secretion, acting through ADM(22-52)-sensitive CGRP1 receptors, which are coupled with different signaling mechanisms in the cortex and medulla; (2) ADM selectively inhibits agonist-stimulated aldosterone secretion, through a mechanism probably involving the blockade of the Ca2+ channel-mediated Ca2+ influx; (3) ADM raises catecholamine secretion, through the activation of the adenylate cyclase/PKA signaling pathway. PMID- 10698125 TI - Epithelin mRNA expression in polycystic kidney disease. AB - Epithelins are polypeptides that are preferentially expressed in epithelial cells and modulate growth. Epithelin expression is predominant in tissues of epithelial origin such as the kidney, spleen, lung, placenta, and colon. Because polycystic kidney disease involves abnormal proliferation of the proximal and/or distal tubule epithelial cells, we investigated epithelin mRNA expression in polycystic kidneys of mice homozygous for the mutation. Epithelin mRNA was highly expressed in the polycystic kidneys of homozygous mice when compared with the heterozygotes or wild type controls. A study on the time course of epithelin expression indicated that epithelin mRNA expression paralleled cyst formation and progression of the disease. A 2-fold increase in expression was observed at Day 15, a stage when cystic changes were first visible. This increase in expression was also observed at Day 21, a stage of maximum disease pathology, which ultimately results in the death of the animal. In situ hybridization localized epithelin mRNA predominantly to the epithelial cell layer surrounding the cysts. The high levels of epithelin in epithelial cells suggest a role in renal epithelial cell proliferation and cyst formation in polycystic kidney disease. PMID- 10698126 TI - Conformation and vasoreactivity of VIP in phospholipids: effects of calmodulin. AB - The purpose of this study was to determine the conformation and vasorelaxant effects of vasoactive intestinal peptide (VIP) self-associated with sterically stabilized phospholipid micelles (SSM) and whether calmodulin modulates both of these processes. Circular dichroism spectroscopy revealed that VIP is unordered in aqueous solution at room temperature but assumes appreciable a helix conformation in SSM. This conformational transition was amplified at 37 degrees C and by a low concentration of calmodulin (0.1 nM). Suffusion of VIP in SSM elicited significant time- and concentration-dependent potentiation of vasodilation relative to that elicited by aqueous VIP in the in situ hamster cheek pouch (P < 0.05). This response was significantly potentiated by calmodulin (0.1 nM). Collectively, these data indicate that exogenous calmodulin interacts with VIP in SSM to elicit conformational transition of VIP molecule from a predominantly random coil in aqueous environment to alpha helix in SSM. This process is associated with potentiation and prolongation of VIP-induced vasodilation in the in situ peripheral microcirculation. PMID- 10698127 TI - Localization of immunoreactive lamprey gonadotropin-releasing hormone in the rat brain. AB - A highly specific antiserum against lamprey gonadotropin-releasing hormone (GnRH) was used to localize 1-GnRH in areas of the rat brain associated with reproductive function. Immunoreactive 1-GnRH-like neurons were observed in the ventromedial preoptic area (POA), the region of the diagonal band of Broca and the organum vasculosum lamina terminalis, with fiber projections to the rostral wall of the third ventricle and the organum vasculosum lamina terminalis. Another population of 1-GnRH-like neurons was localized in the dorsomedial and lateral POA, with nerve fibers projecting caudally and ventrally to terminate in the external layer of the median eminence. Other fibers apparently projected caudally and circumventrically to terminate around the cerebral aqueduct in the mid-brain central gray. By using a highly specific antiserum directed against mammalian luteinizing hormone-releasing hormone (m-LHRH), the localization of the LHRH neuronal system was compared to that of the 1-GnRH system. There were no LHRH neurons in the dorsomedial or the lateral region of the POA that contained the 1 GnRH neurons. As expected, there was a large population of LHRH neurons in the ventromedial POA associated with the diagonal band of Broca and organum vasculosum lamina terminalis. In both of these regions, there were many more LHRH neurons than 1-GnRH neurons and the LHRH neurons extended more dorsally and laterally than the 1-GnRH neurons. The LHRH neurons seemed to project to the median eminence in the same areas as those that were innervated by the 1-GnRH neurons. Absorption studies indicated that 1-GnRH cell bodies were eliminated by adding 1 microg of either 1-GnRH-I or 1-GnRH-III, but not m-LHRH to the antiserum before use. Fibers were largely eliminated by the addition of 1 microg 1-GnRH-III to the antiserum. No chicken GnRH-II neurons or nerve fibers could be visualized by immunostaining. Because the antiserum recognized GnRH-I and GnRH-III equally, we have visualized an 1-GnRH system in rat brain. The results are consistent with the presence of either one or both of these peptides within the rat hypothalamus. Because 1-GnRH-I has only weak nonselective gonadotropin-releasing activity, whereas 1-GnRH-III is a highly selective releaser of follicle-stimulating hormone, and because 1-GnRH neurons are located in areas known to control follicle-stimulating hormone release selectively, our results support the hypothesis that 1-GnRH-III, or a closely related peptide, may be mammalian follicle-stimulating hormone-releasing factor. PMID- 10698128 TI - Are prions a relic of an early stage of peptide evolution? AB - The rather unique properties of prions and their presence in very different kinds of living species suggest that this type of molecule was created at a very early stage of evolution and may even represent a relic from a time where peptide evolution was ongoing and RNA/DNA did not yet exist. A comparison of the most frequently occurring amino acid sequences in known prions with the sequences preferentially formed in the salt-induced peptide formation reaction, the most simple mechanism enabling the formation of peptides under primitive earth conditions, shows a remarkable coincidence that strongly supports this hypothesis. PMID- 10698129 TI - Melanin-concentrating hormone (MCH) modifies memory retention in rats. AB - The purpose of the present study was to evaluate the possible effect of melanin concentrating hormone (MCH) on learning and memory by using the one-trial step down inhibitory avoidance test in rats. The peptide was infused into hippocampus, amygdala, and entorhinal cortex. MCH caused retrograde facilitation when given at 0 or 4 h post-training into hippocampus, but only at 0 h into amygdala. From these results, it seems that MCH modulates memory early after training by acting on both the amygdala and hippocampus and, 4 h after training, on the hippocampus. PMID- 10698131 TI - Endogenous opiates: 1998. AB - This paper is the twenty-first installment of our annual review of research concerning the opiate system. It summarizes papers published during 1998 that studied the behavioral effects of the opiate peptides and antagonists, excluding the purely analgesic effects, although stress-induced analgesia is included. The specific topics covered this year include stress; tolerance and dependence; eating and drinking; alcohol; gastrointestinal, renal, and hepatic function; mental illness and mood; learning, memory, and reward; cardiovascular responses; respiration and thermoregulation; seizures and other neurologic disorders; electrical-related activity; general activity and locomotion; sex, pregnancy, and development; immunologic responses; and other behaviors. PMID- 10698130 TI - Attenuated hypotensive response to proadrenomedullin N-terminal 20 peptide in pregnant rats: modulation by steroid hormones. AB - The hypotensive effect of proadrenomedullin N-terminal 20 peptide (PAMP) was examined in conscious pregnant (8, 14, and 20 days of pregnancy) and nonpregnant rats. Intravenous administration of PAMP (3-60 nmol/kg) produced a dose-dependent depressor response in both pregnant and nonpregnant rats. However, the maximum decrease in blood pressure was significantly attenuated in pregnant rats in mid- and late-gestation (14 and 20 days), but not in early gestation (8 days), than in nonpregnant rats. In ovariectomized rats, the depressor responses in 17beta estradiol (E2)-treated, progesterone (P)-treated, and E2+P-treated rats were significantly attenuated compared with the control rats. We also demonstrated that treatment of sex hormones reduces the depressor response to PAMP in 8-day pregnant rats. In addition, we showed that treatment of sex hormone receptor antagonists partially prevents the attenuation of the depressor response to PAMP in 20 day pregnant rats. These findings suggested that the hypotensive response to PAMP was more attenuated in pregnant rats in mid- and late-gestation than in nonpregnant rats, and that the changes in depressor response that occur at term in pregnant rats may be mediated by sex hormones. PAMP may play some important role in cardiovascular regulation during pregnancy. PMID- 10698132 TI - Neural science: a century of progress and the mysteries that remain. PMID- 10698133 TI - Chloral hydrate versus midazolam for sedation of children for neuroimaging: a randomized clinical trial. AB - OBJECTIVE: The comparative safety and efficacy of chloral hydrate and midazolam for sedation of children has not been adequately studied. METHODS: In a double blind randomized trial, at a single university hospital, we enrolled 40 children, ages 2 months to 8 years, in an out-patient neuroimaging study. Children judged to require sedation were enrolled during a 14-month period ending August 1995. They received identically appearing liquids of equal volume of either chloral hydrate (75 mg/kg, maximum 2 g) or midazolam (0.5 mg/kg, maximum 10 mg) by mouth. Children were monitored for changes in arterial blood pressure, oxygen saturation, pulse, respiration and anxiety. Efficacy was judged by evaluating the child's ability to complete the intended scan. Supplemental dosing was administered to children who were judged inadequately sedated 30 minutes after the initial medication. RESULTS: Interim analysis demonstrated a significant sedation failure rate. Of 40 enrolled children, 33 completed the protocol. Efficacy was significantly improved for the chloral hydrate group for both ability to perform the scan, chloral hydrate = 11/11 (100%, 95% CI = 72-100) vs. midazolam = 11/22 (50%, 95% CI = 29-71), and the need for supplementary dosing, chloral hydrate = 1/11 (9%, 95% CI = 0-26) vs midazolam = 12/22 (55%, 95% CI = 34 76), P<0.05. Mean duration of sedation was not significantly different. No physiological deterioration occurred and no oxygen administration was required. CONCLUSIONS: We conclude that, in these doses, oral chloral hydrate may provide more effective sedation than midazolam for brief neuroimaging studies in young children. PMID- 10698134 TI - Comparison of intubation skills between interfacility transport team members. AB - OBJECTIVE: To compare intubation skill level and success rate between interfacility transport team members. DESIGN: Prospective collection of data. SETTING: University affiliated children's hospital interfacility transport team. PATIENTS: One hundred thirty-two pediatric patients (age range 4 days to 11 years) intubated prior to transport by a specialized team. INTERVENTIONS: None. METHODS: Prospective data was gathered from June 1992 November 1996. In 3616 transports reviewed, 132 intubations were performed by the team at the referring facility. Patient ages ranged from 4 days to 11 years with a mean age of 23 months. We compared resident physicians and respiratory care practitioners (RCPs) to a standard threshold of 1 attempt per successful intubation. An attempt was defined as passage of the endotracheal tube into the oropharynx in an effort to pass it through the vocal cords. Patients were sedated and paralyzed for the procedures. The physicians were 2nd and 3rd year pediatric or emergency medicine residents. They received intubation training in Pediatric Advanced Life Support (PALS), Neonatal Resuscitation Program (NRP), and during rotations through neonatal and pediatric intensive care units. RCPs had an average of 3.5 years of experience overall on the transport team. They received training primarily on mannequins and written tests while in school. They were certified in PALS and NRP and required to participate in annual skill laboratories, which consisted of mannequin intubations and a written examination. RESULTS: The results showed the RCPs to have greater overall success as well as greater success of intubation on first attempt compared to the resident physicians. CONCLUSION: In our experience, RCPs on the interfacility transport team were very successful in performing endotracheal intubations and were more successful than resident physicians. RCPs are established members of not only the transport team, but also the intensive care units and, therefore, should be considered qualified to routinely perform endotracheal intubations in those settings as well. PMID- 10698135 TI - Pediatric emergency department nurses' perspectives on fever in children. AB - BACKGROUND: Fever is the most common complaint of children seen in a Pediatric Emergency Department (PED). Since pediatric emergency nurses commonly educate parents on fever management, this study sought to examine their knowledge base regarding fever in children. METHODS: Through convenience sampling, pediatric emergency registered nurses working at one of four PEDs were surveyed using a self-administered questionnaire containing 10 open-ended questions pertaining to fever in children. RESULTS: Eighty-eight pediatric emergency registered nurses (median experience 8.0 years, range 3 months to 28 years) were surveyed. The median temperature considered by pediatric emergency nurses to be a fever was 38.0 degrees C (100.4 degrees F) with a range of 37.2 degrees C (99.0 degrees F) to 38.9 degrees C (102.0 degrees F), while the median temperature considered to be dangerous to a child was 40.6 degrees C (105.0 degrees F) with a range of 38.0 degrees C (100.4 degrees F) to 41.8 degrees C (107.0 degrees F). Eleven percent was not sure what temperature constituted a fever while 31% was not sure what temperature would be dangerous to a child. Fifty-seven percent considered seizures the primary danger to a febrile child while 29% stated permanent brain injury or death could occur from a high fever. Sixty percent chose acetaminophen as first line treatment while 7% stated alcohol or tepid water baths were also acceptable treatment options. Thirty-eight percent stated that a different medication should be added if a child was still febrile 1 hour after initial treatment while 31% would not use additional medication. Eighteen percent stated it was dangerous for a child to leave the PED if still febrile. CONCLUSION: Fever phobia and inconsistent treatment approaches occur among experienced pediatric emergency registered nurses. These phobias and inconsistencies subsequently could be conveyed to parents. In order to assure accurate parental education, PEDs should educate their medical team regarding the management of fever in children. PMID- 10698136 TI - Pediatric advanced life support training of pediatricians in New Jersey: cause for concern? AB - OBJECTIVE: The Pediatric Advanced Life Support (PALS) course teaches the fundamental basics for pediatric emergency care, and it is recommended that all physicians, nurses, and paramedics who care for children complete training and refresher courses on a regular basis. The purpose of this study was to determine how many pediatricians in general practice participated in PALS courses in the first 3 years since its introduction in New Jersey. METHODS: A questionnaire was sent to all PALS training centers in New Jersey that administered the course from 1990 through 1993. The questionnaire was designed to determine the number of physicians trained; their specialty, and their practice setting. The questionnaire and follow-up telephone interviews focused on the perceptions of course coordinators as to why primary care pediatricians did or did not take PALS courses, and their recommendations for improving pediatrician participation. RESULTS: Two PALS training centers provided courses for only 1 year and did not maintain records of their students. A total of 3652 individuals completed training in the remaining 11 centers. Only 649 of these students were physicians. The largest groups of physicians who completed training were Emergency Medicine physicians (248) and Pediatric residents (175). Forty-two students were pediatricians in general office-based practice, which represents a crude rate of only 0.81% of New Jersey American Academy of Pediatrics (AAP) members. Training center coordinators offered several opinions for these findings. CONCLUSIONS: The majority of those students who participated in PALS training were not physicians. Pediatricians in general office practice accounted for a small percentage of those who could have participated. Further research should be conducted to determine attitudes toward PALS training and the barriers that exist to the office-based pediatrician participating in PALS training. PMID- 10698137 TI - The safety of etomidate for emergency rapid sequence intubation of pediatric patients. AB - OBJECTIVE: To determine whether pediatric patients given etomidate for rapid sequence intubation (RSI) in the ED develop clinically important hypotension or adrenal insufficiency. METHODS: Retrospective review of 100 consecutive patients younger than age 10 years given etomidate for RSI in the ED at two academic medical centers. Data were abstracted from ED and in-patient medical records. Clinically important hypotension was defined as a decrease in systolic blood pressure (BP) measurement to below one standard deviation (SD) of mean normal for age. Clinically important adrenal insufficiency was defined as the need for exogenous corticosteroid replacement for suspected adrenal insufficiency at any time during hospitalization. RESULTS: BP measurements before and within 20 minutes after etomidate administration for RSI were recorded on 84 intubations (84%). The mean change in BP between pre-intubation and post-intubation measurements was a decrease of 1 mmHg (95% CI: -6 mm Hg to +7 mm Hg, P = 0.83). When expressed as a percentage of normal BP for age, the mean change in BP was a decrease of 1% (95% CI: -7% to +6%, P = 0.82). Four patients (4.8%; 95% CI: 1.3 11.7%) had a systolic BP decrease to below one SD of mean normal for age. Fourteen patients received corticosteroids during hospitalization, but none (0/99, 95% CI: 0-3.7%) for suspected adrenal insufficiency. CONCLUSIONS: We found no evidence of clinically important adrenocorticoid suppression and a low incidence of clinically important hypotension when using etomidate for emergent pediatric RSI. Because other induction agents may also result in hypotension, prospective comparison studies are needed to further evaluate the safety of etomidate in this patient population. PMID- 10698138 TI - History and radiographic findings associated with clinically suspected radial head subluxations. AB - OBJECTIVES: To determine: 1) physician practices regarding the use of radiographs for radial head subluxations (RHS), 2) the prevalence of missed fractures in children with a clinical diagnosis of RHS, 3) the relative risk of a fracture with a nonclassic history for mechanism of injury for RHS, and 4) radiographic findings associated with RHS that are difficult to reduce. METHODS: This study began with a physician survey that addressed the integration of radiographs into the management of RHS. We subsequently conducted a prospective randomized trial with a consecutive sampling of children less than 6 years of age who presented to one of 2 urban pediatric emergency departments and 2 suburban pediatric urgent care centers with a clinical diagnosis of RHS. After informed consent was obtained, reduction was undertaken with a maximum of four attempts (two by hyperpronation and two by supination/flexion), 15 minutes apart. Failure to reduce the RHS resulted in the procurement of a radiograph of the elbow. At the conclusion of the study, all radiographs were evaluated by a radiologist blinded to the diagnosis. Patients receiving radiographs were contacted 2 weeks after discharge for verification of the diagnosis. RESULTS: Eighty-four percent of 224 physicians returned completed surveys. Fifty-six percent reported using radiographs for failed reduction attempts. In the second phase of the study, 136 patients were enrolled prospectively: 127 were reduced successfully and 9 patients failed attempts at reduction. Of the nine patients receiving radiographs: four had fractures (prevalence of 2.9% with 95% confidence interval (CI) = 0.8-7.4), two had no radiographic findings and normal function on follow up, and three had isolated posterior fat pads on radiograph and normal function on follow-up. The relative risk of a fracture in children with a nonclassic history defined as any mechanism other than "pull" was 1.200 (95% CI = 0.441 3.264); the relative risk was 1.886 (95% CI = 0.680-5.231) when defining a nonclassic history as any mechanism other than "pull" or "fall." CONCLUSIONS: 1) Physicians tend to order radiographs for elbow injuries they initially perceive to be radial head subluxations when attempts at reduction fail. 2) In our study, fractures in children who presented with the classic flexed elbow/pronated wrist position were rare. 3) The relative risk of a fracture in children with a nonclassic history for mechanism of injury was not significant. 4) An isolated finding of a posterior fat pad in a child with RHS that is difficult to reduce was not associated with a fracture in our small sample of children with radiographic findings. PMID- 10698139 TI - Signs and symptoms of carbamazepine overdose in young children. AB - OBJECTIVE: To examine common signs and symptoms of mild to moderate carbamazepine (CBZ) overdose in young children. METHODS: The medical records of previously healthy children admitted to the pediatric departments for acute accidental CBZ poisoning during the years 1993-1998 were evaluated retrospectively. Information was retrieved on serum CBZ levels, signs and symptoms on admission and during hospitalization, ECG findings, and chemical laboratory test. RESULTS: There were 14 exposed children all under the age of 5 years. These children accidentally took CBZ prescribed for a family member. The diagnosis of CBZ poisoning in seven children was unknown on admission because of inadequate history and was revealed only on toxicology screen. Nystagmus and drowsiness occurred in 8 of the 14 children, nystagmus and ataxia in 4 children, and drowsiness and tachycardia in another 2 children. The peak CBZ serum levels in these children ranged from 18 microg/ml to 32 microg/ml, mean + SD; 25 microg/ml + 4.64 microg/ml (therapeutic range: 5-10 microg/ml). CONCLUSION: Based on a certain group of young pediatric patients with mild to moderate CBZ poisoning, it is concluded that, nystagmus is the most common sign of this overdose. Other common signs are drowsiness and ataxia. The presence of nystagmus and CNS depression of unknown etiology, in a young child should suggest the possibility of CBZ toxicity. PMID- 10698140 TI - Unsuspected splenic rupture in a neonate. AB - Serious intra-abdominal injuries in neonates are very rare. In addition, the signs and symptoms of hemoperitoneum caused by bleeding from solid viscera are vague and nonspecific and often are not recognized before the onset of hypovolemic shock or death. In this report, we describe a 2-day-old infant who presented with shock and pallor who had a ruptured spleen, presumably from birth. We also review the literature and the importance of recognizing this injury in the emergency department setting. PMID- 10698141 TI - Isolated first rib fracture in a high school lacrosse player. PMID- 10698142 TI - Non-Q wave acute myocardial infarction in acute meningococcemia in a 10-year-old girl. AB - INTRODUCTION: Children with acute meningococcemia may have impaired myocardial function resulting in low cardiac output despite normal intravascular volume. Severe meningococcal infection has been associated with acute interstitial myocarditis, endocarditis, and pericarditis, but not with myocardial infarction. CASE: We present the case of a 10-year-old girl with positive family history for premature myocardial infarction who sustained an acute myocardial infarction temporally related to meningococcemia. DISCUSSION: This is the first pediatric case of non-Q wave acute myocardial infarction associated with purpura fulminans in meningococcemia. Similarly, the association of high troponin I levels and meningococcemia has not been described previously. Although, the patient's genetic predisposition for myocardial infarction might have been a potential contributing factor, there was no angiographic evidence of coronary artery disease in this patient. Thereby, other factors related to shock, endotoxin, microthrombi of meningococcemia, and their treatment might have been also contributing. We propose possible mechanisms for this rare but serious complication of meningococcemia and review the literature. PMID- 10698143 TI - Central nervous system manifestations of an ibuprofen overdose reversed by naloxone. AB - Ibuprofen overdose is usually characterized by GI upset, dizziness, and mild sedation. On rare occasions, severe complications such as respiratory failure, metabolic acidosis, renal failure, coma, and death have been reported in both adults and children. Presently, treatment of acute ibuprofen intoxication with complications requires supportive therapy until the symptoms resolve over 24 to 48 hours. We report the case of an 11-month-old female infant with a depressed level of consciousness after ingestion of ibuprofen whose mental status markedly improved with administration of naloxone. PMID- 10698145 TI - Point-of-care testing: a critical review. PMID- 10698144 TI - A 4-week-old infant with idiopathic pulmonary hemorrhage. PMID- 10698146 TI - Yohimbine. PMID- 10698147 TI - Hypertension discovered during health fair week. PMID- 10698148 TI - Pediatric emergency medicine: legal briefs. PMID- 10698149 TI - Nasal wash technique for nasal foreign body removal. AB - Nasal foreign bodies are seen commonly both in the office and pediatric emergency department setting. There have been a number of strategies described for their removal. We describe the "nasal wash" as a technique for nasal foreign body removal in the following three case reports. The "nasal wash" has been used in many pediatric vaccine studies as a method to collect mucus and relies on simple equipment readily available in the office setting. PMID- 10698150 TI - Antibiotic conundrum. PMID- 10698151 TI - Viral infections, allergy and autoimmunity: a complex, but fascinating link. PMID- 10698152 TI - Good or evil: CD26 and HIV infection. AB - Acquired immune deficiency syndrome (AIDS) is an incurable disease at present and so many efforts to conquer this disease are being made around the world. In studies of human immunodeficiency virus (HIV) infection and the disease progression, it has been reported that T cells expressing CD26 are preferentially infected and depleted in HIV-infected individuals. CD26 is a widely distributed 110 kDa cell-surface glycoprotein with known dipeptidyl peptidase IV (DPPIV) activity in its extracellular domain. This ectoenzyme is capable of cleaving N terminal dipeptides from polypeptides with either proline or alanine residues in the penultimate position. On human T cells, CD26 exhibits the co-stimulatory function and plays an important role in immune response via its ability to bind adenosine deaminase (ADA) and association with CD45. Recent studies have been stripping the veil from over the relationship between CD26 and HIV infection. Susceptibility of cells to HIV infection is correlated with CD26 expression, and HIV transactivator Tat and envelope protein gp120 are reported to interact with CD26. These observations indicate that CD26 is closely involved in HIV cell entry and that CD26-mediated T cell immune response is suppressed. In addition, it has been demonstrated that the anti-HIV and chemotactic activities of RANTES (regulated on activation, normal T cell expressed and secreted) and stromal cell derived factor-1 (SDF-1) are controlled with the DPPIV activity of CD26. Thus, the regulation of the function of chemokines by CD26/DPPIV appears to be essential for lymphocyte trafficking and infectivity of HIV strains. PMID- 10698153 TI - Hepatitis C virus replication and pathogenesis. AB - The mechanism involved in the development of persistent hepatitis C virus (HCV) infection and the pathogenesis remain unclear. The present review is an accumulation of evidence gathered to date. In addition, it discusses the system developed to characterize HCV. PMID- 10698154 TI - Virus-induced immune dysregulation as a triggering factor for the development of drug rashes and autoimmune diseases: with emphasis on EB virus, human herpesvirus 6 and hepatitis C virus. AB - There are a considerable amount of empirical and theoretic medical literature regarding the possible role of viruses in the development of drug rashes and autoimmune diseases: under these conditions, interactions of viruses with the immune system would serve as an accelerating factor of disease pathogenesis. Recent reports have provided evidence to indicate that immune responses against infections with Epstein Barr (EB) virus and human herpesvirus 6 (HHV-6), which are lymphotropic members of the herpes virus group, not only aid the direct elimination of the virus but also contribute to a favorable milieu for the initiation or acceleration of drug rashes. Viruses that can persist for the lifetime of the host despite strong immune responses against them, such as EB virus and hepatitis C virus (HCV), would be also relevant to the pathogenesis of autoimmune diseases. HCV has been reportedly associated with a wide variety of dermatoses that, in common, show histologically the lichenoid tissue reaction. Although porokeratosis that manifests lichenoid histopathological features had long been regarded as being associated with immunosuppression, we found that HCV could act as trigger for the development of porokeratosis during states of immunosuppression. Thus, the main purpose of this review is to describe recent work on the etiology of drug rashes and autoimmune disease with special reference to viral infections. PMID- 10698155 TI - Cutaneous lymphoproliferative disorders associated with Epstein-Barr virus infection: a clinical overview. AB - Epstein Barr virus (EBV) infection is implicated in various kinds of neoplasms including certain types of cutaneous T or natural killer (NK) cell proliferative disorders. Although a pathogenic role of EBV infection is not clear, some EBV gene products expressed during a latency phase were found to have biological properties leading to cellular gene expression and immortalization. Furthermore, EBV can use an array of strategies to evade host immune responses, and maintain the latent infection. EBV-associated cutaneous lymphoproliferative disorders are prevalent in Asia, and less frequent in western countries where infectious mononucleosis is common in adolescents and young adults. This review introduces recent advances on the mechanism of EBV infection, highlighting unique clinicopathologic manifestations of EBV-associated cutaneous lymphoproliferative disorders. PMID- 10698156 TI - Current understanding of cytomegalovirus infection in immunocompetent individuals. AB - Human cytomegalovirus (CMV) is a member of the herpes family of viruses. After primary infection, it undergoes latency/persistence. Significant progress has been made in the last few years in detecting CMV. The most available approach to the diagnosis of CMV infection is the direct detection of CMV antigen in nuclei of peripheral blood leukocytes, an assay known as pp65 direct antigenemia test. CMV infection is well controlled in the immunocompetent hosts; however, there are various immunological changes in immune function during and after recovery from CMV infections. Characteristic changes in lymphocyte subsets occur during CMV infection, mainly involving expansion and activation of CD8+ T lymphocytes and NK cells. On the other hand, CMV has an array of immune escape strategies for establishing a life long latent state: CMV inhibits major histocompatibility complex (MHC) class I expression within infected cells and impairs IFN-gamma induced MHC class II-dependent antigen presentation by macrophages; it can also encode proteins that can interfere with the presentation of viral peptide antigens to T cells. While cutaneous manifestations of CMV seen in immunocompromised patients have been extensively reported, those in adult immunocompetent individuals have received relatively little attention: in this setting the primary CMV infection appears as CMV mononucleosis. At the time of occurrence of the mononucleosis syndrome, a variety of extracutaneous and cutaneous manifestations occur. These clinical symptoms are not the direct consequence of proliferation of CMV in given tissues but indicative of the immunological response toward CMV. The incidence of the appearance of eruptions in CMV mononucleosis is variable. Certain drugs given in the early stage of this disease play an important role in the development of eruption, just as with the ampicillin rashes in the Epstein-Barr virus mononucleosis. Although the mechanism by which drugs trigger the development of rashes in patients with CMV mononucleosis is unknown, it is assumed that CMV is likely to be a potential amplifier of drug rashes induced by activation of drug-specific T cells. By improving methods for detection of CMV, we can recognize that many types of eruptions other than CMV mononucleosis could be induced by primary infection or reactivation of CMV. PMID- 10698157 TI - HHV-6, 7 and their related diseases. AB - Human herpesvirus 6 (HHV-6) and human herpesvirus 7 (HHV-7) are relatively recently discovered beta-herpesvirus. They are prevalent in the human population. Primary infection of HHV-6 has been associated with exanthem subitum and febrile illness. Little information is known about the clinical characteristics of primary infection with HHV-7, although some cases of exanthem subitum have been linked to it. HHV-6 has been recently recognized as an opportunistic pathogen in patients with HIV infection and in transplant recipients. The techniques now available to detect these two viruses remain limited, though putative roles for HHV-6 and HHV-7 in several diseases linked to viral infection have been reported. This report reviews the current knowledge of HHV-6 and HHV-7 biology and their pathogenesis. PMID- 10698158 TI - What can spermatogonial transplants teach us about male reproductive biology? PMID- 10698160 TI - Rhythmic bursts of calcium transients in acute anterior pituitary slices. AB - Endocrine cells isolated from the anterior pituitary fire intracellular Ca2+ ([Ca2+]i) transients due to voltage-gated Ca2+ entry. However, the patterns of [Ca2+]i transients within the glandular parenchyma of the anterior pituitary are unknown. Here we describe, using real-time confocal laser microscopy, several spontaneous patterns of calcium signaling in acute pituitary slices prepared from male as well as cycling and lactating female rats. Forty percent of the cells demonstrated a spontaneous bursting mode, consisting of an active period of [Ca2+]i transients firing at a constant frequency, followed by a rest period during which cells were either silent or randomly active. The remaining recordings from endocrine cells either demonstrated random [Ca2+]i transients or were silent. These rhythmic bursts of [Ca2+]i transients, which required extracellular calcium, were detected in lactotrophs, somatotrophs, and corticotrophs within the acute slices. Of significance was the finding that the bursting mode could be adjusted by hypothalamic factors. In slices prepared from lactating rats, TRH recruited more bursting cells and finely adjusted the average duty cycle of [Ca2+]i bursts such that cells fired patterned bursts for approximately 70% of the recording period. Eighty-six percent of these cells were lactotrophs. Thus, the rhythmic [Ca2+]i bursts and their tuning by secretagogues may provide timing information that could encode for one or more cellular functions (e.g. exocytosis and/or gene expression) critical for the release of hormones by endocrine cells in the intact gland. PMID- 10698159 TI - Transforming growth factor-beta3 stimulates lactotrope cell growth by increasing basic fibroblast growth factor from folliculo-stellate cells. AB - Recently, we have shown that transforming growth factor-beta3 (TGFbeta3) mediates estradiol's mitogenic action in primary cultures of mixed anterior pituitary cells. In some cell types, TGFbeta isoforms stimulate cell proliferation via a paracrine mechanism by increasing growth stimulatory peptide growth factors. Whether such a mechanism exists in pituitary cell culture was examined in the studies presented here. The data demonstrate that unlike the response of lactotropes in mixed pituitary cultures, cultures of enriched lactotropes, obtained by Percoll gradient separation, did not proliferate in response to TGFbeta3 treatment. The lactotropic cells of the RC-4B/C cell line, a cell line that contains all of the hormone-secreting cell types of the anterior pituitary but is devoid of folliculo-stellate (FS) cells, did not proliferate in response to TGFbeta3 unless RC-4B/C cells were cocultured with FS cells. Enriched lactotropes cocultured with FS cells also demonstrated a proliferative response to TGFbeta3. Media collected from FS cells treated with TGFbeta3 stimulated the proliferation of lactotropes in enriched cultures. TGFbeta3 increased the release of basic fibroblast growth factor from FS cells. Immunoneutralization of basic fibroblast growth factor in FS cell-conditioned medium inhibited the growth stimulatory action on lactotropes. These data provide evidence for a novel mechanism of TGFbeta3 action involving cell-to-cell interaction in the anterior pituitary between lactotropes and FS cells during estrogen-induced mitogenesis. PMID- 10698161 TI - Epidermal growth factor secreted from submandibular salivary glands interferes with the lipolytic effect of adrenaline in mice. AB - We had described that epidermal growth factor (EGF) interfered with the lipolytic effect of catecholamines in isolated adipocytes. Since catecholamines stimulate the release of EGF from submandibular salivary glands to blood plasma in male mice, we studied whether EGF affected also the lipolytic response to adrenaline in whole animals. We studied the effect of adrenaline in sialoadenectomized and sham-operated mice receiving or not a high dose of EGF following adrenaline injection. There was no difference in plasma EGF concentration between sham operated and sialoadenectomized animals receiving saline. After adrenaline administration plasma EGF increased by 20-fold in sham-operated but did not increase in sialoadenectomized mice. Indeed, the increase was much higher (more than 100-fold) in mice receiving exogenous EGF. The effect of adrenaline on plasma concentration of both glycerol and nonesterified fatty acids was higher as lower was plasma EGF concentration. Isolated adipocytes obtained from sham operated or sialoadenectomized mice had identical lipolytic response to adrenaline. The lipolytic response of adipocytes to isoproterenol was decreased by addition of EGF. To study whether the interference with the in vivo lipolytic effect of adrenaline had further metabolic consequences, we measured plasma beta hydroxybutyrate concentration in plasma. There was no difference in the response to adrenaline between sham-operated and sialoadenectomized mice in spite of the difference in plasma nonsterified fatty acid concentration. Studies in isolated hepatocytes indicated that ketogenesis run at near maximal rate in this range of substrate concentration. These results suggest that EGF in the physiological range decreases the lipolytic effect of adrenaline but does not compromise further metabolic events like the enhancement of ketogenesis. PMID- 10698162 TI - Autosomal dominant growth hormone (GH) deficiency type II: the Del32-71-GH deletion mutant suppresses secretion of wild-type GH. AB - Familial isolated GH deficiency type II is an autosomal dominant form of short stature, associated in some families with mutations that result in missplicing to produce del32-71-GH, a protein that cannot fold normally. The mechanism by which this mutant suppresses the secretion of wild-type GH encoded by the normal allele is not known. Coexpression of del32-71-GH with wild-type human GH in transient transfections of the neuroendocrine cell lines GH4C1 and AtT20 suppressed accumulation of wild-type GH. The suppression of wild-type GH accumulation by del32-71-GH was a posttranslational effect on wild-type GH caused by decreased stability, rather than decreased synthesis, of wild-type GH. Coexpression of del32-71-GH with human PRL did not suppress accumulation of PRL, indicating that there was not a general suppression of secretory pathway function. Accumulation of del32-71-GH protein was not necessary for the suppression of wild-type GH, because del32-71-GH did not accumulate in the neuroendocrine cell lines in which suppression of accumulation of wild-type GH was observed. Del32-71-GH did accumulate in transfected COS and CHO cells, but did not suppress the accumulation of wild-type GH in these cells. These studies suggest that del32-71 GH may cause GH deficiency in somatotropes of heterozygotes expressing both wild type and del32-71-GH by decreasing the intracellular stability of wild-type GH. PMID- 10698163 TI - Influence of the in vivo calcium status on cellular calcium homeostasis and the level of the calcium-binding protein calreticulin in rat hepatocytes. AB - Little attention has been given to the consequences of the in vivo calcium status on intracellular calcium homeostasis despite several pathological states induced by perturbations of the in vivo calcium balance. The aim of these studies was to probe the influence of an in vivo calcium deficiency on the resting cytoplasmic Ca2+ concentration and the inositol-1,4,5-trisphosphate-sensitive Ca2+ pools. Studies were conducted in hepatocytes (a cell type well characterized for its cellular Ca2+ response) isolated from normal and calcium-deficient rats secondary to vitamin D depletion. Both resting cytoplasmic Ca2+ concentration and Ca2+ mobilization from inositol-1,4,5-trisphosphate-sensitive cellular pools were significantly lowered by calcium depletion. In addition, Ca deficiency was shown to significantly reduce calreticulin messenger RNA and protein levels but calcium entry through store-operated calcium channels remained unaffected, indicating that the Ca2+ entry mechanisms are still fully operational in calcium deficiency. The effects of calcium deficiency on cellular calcium homeostasis were reversible by repletion with oral calcium feeding alone or by the administration of the calcium-regulating hormone 1,25-dihydroxyvitamin D3, further strengthening the tight link between extra- and intracellular calcium. These data, therefore, challenge the currently prevailing hypothesis that extracellular Ca2+ has no significant impact on cellular Ca2+ by demonstrating that despite the large Ca2+ gradient between extra- and intracellular Ca2+ concentrations, calcium deficiency in vivo significantly alters the hormone-sensitive cellular calcium homeostasis. PMID- 10698164 TI - Thyroid-specific gene expression is differentially influenced by intracellular glutathione level in FRTL-5 cells. AB - Alteration of the redox potential has been proposed as a mechanism influencing gene expression. Reduced glutathione (GSH) is one of the cellular scavengers involved in the regulation of the redox potential. To test the role that GSH may play in thyroid cells, we cultured a differentiated rat thyroid cell strain (FRTL 5) in the presence of L-buthionine-(S,R)-sulfoximine (BSO). BSO affects GSH synthesis by irreversibly inhibiting gamma-glutamylcysteine synthetase (EC 6.3.2.2), a specific enzyme involved in GSH synthesis. BSO-treated FRTL-5 cells show a great decrease in the GSH level, whereas malondialdehyde increases in the cell culture medium as a sign of lipid peroxidation. In these conditions the activity of two thyroid-specific promoters, thyroglobulin (Tg) and thyroperoxidase (TPO), is strongly reduced in transient transfection experiments. As both Tg and TPO promoters depend upon the thyroid-specific transcription factors, thyroid-specific transcription factor-1 (TTF-1) and Pax-8 for full transcriptional activity, we tested whether reduction of GSH concentration impairs the activity of these transcription factors. After BSO treatment of FRTL 5 cells, both transcription factors fail to trans-activate the respective chimerical targets, C5 and B-cell specific activating protein promoters, containing, respectively, multimerized TTF-1- or Pax-8-binding sites only as well as the Tg and TPO natural promoters. Northern analysis revealed that endogenous Tg messenger RNA (mRNA) expression is also reduced by BSO treatment, whereas endogenous TPO expression is not modified. Furthermore, the Pax-8 mRNA steady state concentration does not change in BSO-treated cells, whereas TTF-1 mRNA slightly decreases. Immunoblotting analysis of FRTL-5 nuclear extracts does not show significant modification of the Pax-8 concentration in BSO-treated cells, whereas a decrease of 25% in TTF-1 protein is revealed. Furthermore, BSO treatment decreases the DNA-binding activity to the respective consensus sequence of both transcription factors. Finally, different mechanisms seem to act on TTF-1 and Pax-8 functional impairment in BSO-treated cells. Indeed, with a lowered GSH concentration, the overexpressed Pax-8 still activates transcription efficiently, whereas, on the contrary, the overexpressed TTF-1 does not recover its transactivation capability when the respective chimerical target sequences are used (C5 and BSAP). When the natural Tg and TPO promoter sequences are used, overexpression of Pax-8 parallels the effect on both promoters observed using the chimeric target sequences, whereas overexpression of TTF-1 increases TPO promoter transcriptional activity only. PMID- 10698165 TI - Differential cloning of growth hormone-regulated hepatic transcripts in the aged rat. AB - It has been suggested that aging or at least some of its symptoms are related to a physiological decline in GH levels with age. This study was performed to elucidate age-related changes in GH-dependent effects at the level of gene expression. Through the application of complementary DNA representational difference analysis (RDA) we have identified gene products that are reduced during aging in rat liver. The expression of these genes was restored upon GH treatment. Results from reverse Northern and ribonuclease protection analysis confirmed that the RDA products were truly differentially expressed. In addition to well characterized GH-regulated genes, including CYP2C12, CYP2C13, and alpha2u globulin, we demonstrate the differential expression of at least 11 genes previously not known to be under GH control. Several hepatic transcripts encoding enzymes and receptors involved in the metabolism of protein, carbohydrates, and lipids were identified. Other RDA products consisted of transcripts encoding proteins involved in ATP synthesis, detoxification of reactive oxygen species, or immune responses. This list of GH-regulated genes in the old rat may shed further light on the action and mechanism behind the positive effects of GH on, for example, body composition and the immune system that have been observed in different animal and human studies. PMID- 10698166 TI - Insulin-mediated cell proliferation and survival involve inhibition of c-Jun N terminal kinases through a phosphatidylinositol 3-kinase- and mitogen-activated protein kinase phosphatase-1-dependent pathway. AB - We previously reported that long term treatment with insulin led to sustained inhibition of c-Jun N-terminal kinases (JNKs) in CHO cells overexpressing insulin receptors. Here we investigated the signaling molecules involved in insulin inhibition of JNKs, focusing on phosphatidylinositol 3-kinase (PI 3-K) and mitogen-activated protein kinase phosphatase-1 (MKP-1). In addition, we examined the relevance of JNK inhibition for insulin-mediated proliferation and survival. Insulin inhibition of JNKs was mediated by PI 3-K, as it was blocked by wortmannin and LY294002 and required the de novo synthesis of a phosphatase(s), as it was abolished by orthovanadate and actinomycin D. MKP-1 was a good candidate because 1) insulin stimulation of MKP-1 expression correlated with insulin inhibition of JNKs; 2) insulin stimulation of MKP-1 expression, like insulin inhibition of JNKs, was mediated by PI 3-K; and 3) the transient expression of an antisense MKP-1 RNA reduced the insulin inhibitory effect on JNKs. The overexpression of a dominant negative JNK1 mutant increased insulin stimulation of DNA synthesis and mimicked the protective effect of insulin against serum withdrawal-induced apoptosis. The overexpression of wild-type JNK1 or antisense MKP-1 RNA reduced the proliferative and/or antiapoptotic responses to insulin. Altogether, these results demonstrate that insulin inhibits JNKs through a PI 3-K- and MKP-1-dependent pathway and provide evidence for a key role for JNK inhibition in insulin regulation of proliferation and survival. PMID- 10698167 TI - Early hyperplastic renal growth after uninephrectomy in adult female rats. AB - The early, accelerated remnant kidney growth following uninephrectomy (UNX) occurs through alternate mechanisms in juvenile and adult male rats, which may govern the type of renal growth that occurs after UNX. Early compensatory renal growth (CRG) in the adult male rat is GH dependent, but independent of changes in the renal insulin-like growth factor I (IGF-I) system. In contrast, CRG is GH independent in the juvenile male rat, but is associated with significant increases in the renal IGF-I system, and hyperplastic kidney growth. The few studies that examined early CRG in female animals suggest that remnant kidney growth is less than that observed in males, and there is a hyperplastic component, indicating potential gender differences. Whether these differences result from alternate growth mechanisms is unknown. The purpose ofthe present study was to determine the rate, type, and potential mechanism of early remnant kidney growth in adult female rats after UNX. GH levels were determined in conscious, sham-operated, and UNX adult female Wistar rats 24 h postsurgery. Unlike previous findings in adult male UNX rats, pulsatile GH levels were not elevated in UNX female rats. When GH release was suppressed using an antagonist to GH-releasing factor, remnant kidney growth was not different from that in saline/UNX remnant kidneys (25.7+/-4.8% vs. 27.7+/-2.1%, respectively, at 48 h post-UNX). This GH-independent CRG was associated with significant hyperplastic growth in both adult andjuvenile female remnant kidneys, as determined by bromodeoxyuridine incorporation and increases in total DNA. Also associated with the mitogenic growth in the adult female were significant 2- to 4-fold increases in remnant kidney IGF-I receptor gene expression, which occurred in the presence and absence of pulsatile GH secretion. Lastly, the growth rate of adult female remnant kidneys was not different from that observed in male remnant kidneys at these early time points (0.21+/-0.02 vs. 0.20+/-0.02 g at 24 h, and 0.26+/-0.02 vs. 0.30+/-0.03 g at 48 h in female and male remnant kidneys, respectively; P = NS). Thus, in female rats, the initial phase of CRG is GH independent, but is associated with significant increases in remnant kidney IGF-I receptor gene expression and hyperplastic renal growth. This, in addition to previous findings, indicates that there are sex differences in early CRG after UNX. Moreover, the findings confirm that the mechanism governing the initial phase of CRG appears to be a critical determinant for significant hyperplastic remnant kidney growth. PMID- 10698168 TI - Identification of SP3 as a negative regulatory transcription factor in the monocyte expression of growth hormone. AB - A number of studies from different laboratories clearly show that cells of the immune system produce a GH molecule indistinguishable from that produced in the pituitary. A more recent finding from our studies suggests that monocytes use the same first exon and promoter sequence for the expression of lymphocyte GH as that reported for the expression of pituitary GH. In this report we have extended these results by determining that two members of the SP family of transcription factors, SP1 and SP3, bind to the region at -138/-133 bp containing a GGGAGG motif. Confirmation that this region of the monocyte GH promoter-bound SP1 and SP3 was accomplished using electrophoretic mobility shift assays with SP1 consensus and mutant probes as well as specific antibodies to SP1 and SP3. Selective mutation of the SP1/SP3 site increased basal transcription by 73%, indicating that this site is important in transcriptional inhibition. Overexpression of SP1 had no demonstrable effect on the GH promoter, whereas overexpression of SP3 caused inhibition of expression in P-388 monocyte cells. Cotransfection of P-388 cells with overexpression vectors for both SP1 and SP3 transcription factors also resulted in inhibition of basal expression. Transfection experiments in Drosophila SL-2 cells overexpressing SP1 and/or SP3 suggest that both factors repress the basal expression of GH promoter luciferase constructs and that the effect together was additive. Taken together, the results demonstrate that basal expression of monocyte GH may be negatively regulated by SP3. PMID- 10698169 TI - Role of G protein-coupled receptor kinases in glucose-dependent insulinotropic polypeptide receptor signaling. AB - The glucose-dependent insulinotropic polypeptide receptor (GIPR) is a member of class II G protein-coupled receptors. Recent studies have suggested that desensitization of the GIPR might contribute to impaired insulin secretion in type II diabetic patients, but the molecular mechanisms of GIPR signal termination are unknown. Using HEK L293 cells stably transfected with GIPR complementary DNA (L293-GIPR), the mechanisms of GIPR desensitization were investigated. GIP dose dependently increased intracellular cAMP levels in L293 GIPR cells, but this response was abolished (65%) by cotransfection with G protein-coupled receptor kinase 2 (GRK2), but not with GRK5 or GRK6. Beta arrestin-1 transfection also induced a significantly decrease in GIP-stimulated cAMP production, and this effect was greater with cotransfection of both GRK2 and beta-arrestin-1 than with either alone. In betaTC3 cells, expression of GRK2 or beta-arrestin-1 attenuated GIP-induced insulin release and cAMP production, whereas glucose-stimulated insulin secretion was not affected. GRK2 and beta arrestin-1 messenger RNAs were identified by Northern blot analysis to be expressed endogenously in betaTC3 and L293 cells. Overexpression of GRK2 enhanced agonist-induced GIPR phosphorylation, but receptor endocytosis was not affected by cotransfection with GRKs or beta-arrestin-1. These results suggest a potential role for GRK2/beta-arrestin-1 system in modulating GIP-mediated insulin secretion in pancreatic islet cells. Furthermore, GRK-mediated receptor phosphorylation is not required for endocytosis of the GIPR. PMID- 10698170 TI - Identification of the lipophilic factor produced by macrophages that stimulates steroidogenesis. AB - Macrophages are known to release a lipophilic factor that stimulates testosterone production by Leydig cells. This macrophage-derived factor (MDF) is thought to be physiologically relevant, because removal of macrophages from the testis results in altered testosterone secretion and reduced fertility. The purpose of the present study was to purify this factor, elucidate its chemical structure, and determine whether it is both present in the testis and acts when injected intratesticularly. Culture media from testicular and peritoneal macrophages were extracted with ether, and the organic phase was sequentially purified on C18, silica, and cyano-HPLC columns. MDF was detected using a rat Leydig cell bioassay, with testosterone secretion being the end point. Purified material and crude ether extracts were analyzed by gas chromatography/mass spectrometry and nuclear magnetic resonance spectroscopy. The time of elution of MDF from both testicular and peritoneal macrophages was identical on all three HPLC columns. A single peak was observed when MDF, obtained from the final HPLC column, was analyzed by gas chromatography. The MS fragmentation pattern of purified material from both peritoneal and testicular macrophages was identical to that of a reference preparation of 25-hydroxycholesterol. Also, the nuclear magnetic resonance spectrum of MDF was similar to that of authentic 25-hydroxycholesterol. When 25-hydroxycholesterol was subjected to the identical purification scheme as MDF, it was found to elute at the same times as MDF on all three columns and elicited activity in the Leydig cell bioassay as expected. Control medium purified identically did not contain 25-hydroxycholesterol or have biological activity. Ether extracts of testis contained 25-hydroxycholesterol, indicating that this compound is present under physiological conditions. Similarly, when 25 hydroxycholesterol was injected into the testis of adult rats, testosterone production was increased within 3 h. Taken together, these data indicate that the lipophilic factor produced by macrophages that stimulates steroidogenesis is 25 hydroxycholesterol. PMID- 10698171 TI - Calnexin and calreticulin binding to human thyroperoxidase is required for its first folding step(s) but is not sufficient to promote efficient cell surface expression. AB - Human thyroperoxidase (hTPO) is a type I transmembrane-bound heme-containing glycoprotein that catalyzes the synthesis of thyroid hormones. In a previous study we stably expressed hTPO in Chinese hamster ovary cells and observed that after the synthesis, only 20% of the hTPO molecules were recognized by a monoclonal antibody (mAb 15) directed against a conformational structure, and that only 2% were able to reach the cell surface. In the present study it was proposed to determine how calnexin (CNX) and calreticulin (CRT) contribute to the folding of hTPO. Sequential immunoprecipitation was performed using anti-CNX or anti-CRT followed by anti-hTPO antibodies, and the results showed that CNX and CRT were associated with hTPO. Inhibiting the interactions between CNX or CRT and hTPO using castanospermine greatly reduced the first step(s) in the hTPO folding process. Under these conditions, the half-life of this enzyme was greatly reduced (2.5 vs. 17 h in the control experiments), and hTPO was degraded via the proteasome pathway. This reduced the rate of hTPO transport to the cell surface. Overexpression of CNX or CRT into the hTPO-CHO cells was found to enhance the first hTPO folding step(s) by 20-60%, but did not increase the level of hTPO present at the cell surface. All in all, these findings provide evidence that CNX and CRT are crucial to the first step(s) in hTPO folding, but that interactions with other molecular chaperones are required for the last folding steps to take place. PMID- 10698172 TI - Involvement of the Sst1 somatostatin receptor subtype in the intrahypothalamic neuronal network regulating growth hormone secretion: an in vitro and in vivo antisense study. AB - Five somatostatin (SRIH) receptors (sst1-5) have been cloned. Recent anatomical evidence suggests that sst1 and sst2 may be involved in the central regulation of GH secretion. Given the lack of specific receptor antagonists, we used selective antisense oligodeoxynucleotides (ODNs) to test the hypothesis that one or both of these subtypes are involved in the intrahypothalamic network regulating pulsatile GH secretion. In mouse neuronal hypothalamic cultures the proportion of GHRH neurons coexpressing sst1 or sst2 messenger RNAs (mRNAs) was identical. In contrast, sst1 mRNAs were more often present than sst2 in SRIH-expressing neurons. Firstly, sst1 antisense ODN in vitro treatment abolished sst1, but not sst2, receptor modulation of glutamate sensitivity and decreased sst1, but not sst2, mRNAs. The reverse was true after treatment with sst2 antisense. Sense ODNs did not alter the effects of SRIH agonists. In a second series of experiments, nonanaesthetized adult male rats were infused for 120 h intracerebroventricularly with ODNs. Only the sst1 antisense ODN diminished the amplitude of ultradian GH pulses without modifying their frequency. In parallel, sst1 antisense ODN strongly diminished sst1 immunoreactivity in the anterior periventricular nucleus and median eminence, as well as sstl periventricular nucleus mRNA levels. The effectiveness of the sst2 antisense ODN was attested by the inhibition of hypothalamic binding of [125I]Tyr0-D-Trp8-SRIH. Scrambled ODNs had no effect on GH secretion or on sst mRNAs or SRIH binding levels. These results favor a preferential involvement of sst1 receptors in the intrahypothalamic regulation of GH secretion by SRIH. PMID- 10698173 TI - Modulation of endocrine systems and food intake by green tea epigallocatechin gallate. AB - Green tea polyphenols, especially the catechin, (-)-epigallocatechin gallate (EGCG), have been proposed as a cancer chemopreventative based on a variety of laboratory studies. For clear assessment of the possible physiological effects of green tea consumption, we injected pure green tea catechins ip into rats and studied their acute effects on endocrine systems. We found that EGCG, but not related catechins, significantly reduced food intake; body weight; blood levels of testosterone, estradiol, leptin, insulin, insulin-like growth factor I, LH, glucose, cholesterol, and triglyceride; as well as growth of the prostate, uterus, and ovary. Similar effects were observed in lean and obese male Zucker rats, suggesting that the effect of EGCG was independent of an intact leptin receptor. EGCG may interact specifically with a component of a leptin-independent appetite control pathway. Endocrine changes induced by parenteral administration of EGCG may relate to the observed growth inhibition and regression of human prostate and breast tumors in athymic mice treated with EGCG as well as play a role in the mechanism by which EGCG inhibits cancer initiation and promotion in various animal models of cancer. PMID- 10698174 TI - Major hypercorticism is an endocrine feature of ewes with naturally occurring scrapie. AB - The aim of this study was to identify the origin of scrapie-induced hypercortisolism. Cortisol and ACTH kinetics and production rate were measured in 14 ewes (6 healthy and 8 scrapie-affected). It was shown that cortisol plasma clearance remained unmodified but that cortisol production rate and plasma concentrations of free cortisol were increased by a factor of 5, whereas the total cortisol plasma concentrations were only doubled. The apparent discrepancy between adrenal secretion rate and the corresponding total cortisol plasma levels was attributable to the scrapie-induced lower corticosteroid-binding globulin (CBG) binding capacity, which altered the ratio of free-to-bound cortisol. The secretion rate of ACTH from diseased ewes was increased by a factor of 1.5, in comparison with healthy ewes, and 4 of the 8 scrapie-affected ewes exhibited a decreased response to a low dexamethasone suppression test. The administration of tetracosactide induced a 2-fold increase in the cortisol production in diseased ewes, compared with that of healthy ewes, but the pituitary sensitivity to ovine CRF was not modified by the prion disease. In conclusion, natural scrapie displays a syndrome of hypercorticism associated with increased ACTH secretion, hyperresponsiveness of the adrenals, and lower CBG binding capacity, which leads to overexposure to CBG-free cortisol. PMID- 10698175 TI - Suppression of luteal angiogenesis in the primate after neutralization of vascular endothelial growth factor. AB - Manipulation of angiogenesis may have a profound effect on female reproductive function, but this has not yet been demonstrated by direct experiment in species with ovulatory cycles similar to those in women. To investigate whether angiogenesis could be inhibited in the primate corpus luteum, and the consequences of such inhibition on luteal function, marmosets were treated with an antibody to vascular endothelial growth factor (VEGF). Treatment commenced at the time of ovulation and was continued for 3 days (early luteal group) or 10 days (midluteal group). Bromodeoxyuridine was used to label proliferating cells, being administered 1 h before collecting ovaries from control and treated animals in the early or midluteal phase. Ovarian sections were stained using an antibody to bromodeoxyuridine, and a proliferation index was obtained; endothelial cell quantification was performed using factor VIII as an endothelial cell marker. Intense proliferation in the early luteal phase was suppressed by anti-VEGF treatment. This resulted in blockade of development of the normally extensive capillary bed, as in the animals treated until the mid-luteal phase the numbers of endothelial cells were reduced. The hormone-producing cells remained largely unaltered in the posttreatment corpus luteum, although the presence of lipid accumulation, and small pockets of cells showing basophilia and nuclear condensation were observed. Significantly, luteal function, as judged by secretion of progesterone, was markedly compromised by the treatment, being reduced by 60% in comparison with controls. It is concluded that VEGF-mediated angiogenesis is an essential component of luteal function in primates and therefore has the potential to be regulated. PMID- 10698176 TI - Responsiveness of the ovine gonadotropin-releasing hormone receptor gene to estradiol and gonadotropin-releasing hormone is not detectable in vitro but is revealed in transgenic mice. AB - Although the ability of estradiol to enhance pituitary sensitivity to GnRH is established, the underlying mechanism(s) remain undefined. Herein, we find that approximately 9,100 bp of 5' flanking region from the ovine GnRH receptor (oGnRHR) gene is devoid of transcriptional activity in gonadotrope-derived cell lines and is not responsive to either estradiol or GnRH. In stark contrast, this same 9,100 bp promoter fragment directed tissue-specific expression of luciferase in multiple lines of transgenic mice. To test for hormonal regulation of the 9,100-bp promoter, ovariectomized transgenic females were treated with a GnRH antiserum alone or in combination with estradiol. Treatment with antiserum alone reduced pituitary expression of luciferase by 80%. Pituitary expression of luciferase in animals receiving both antiserum and estradiol was approximately 50 fold higher than animals receiving antiserum alone. The estradiol response of the -9,100-bp promoter was equally demonstrable in males. In addition, a GnRH analog (D-Ala-6-GnRH) that does not cross-react with the GnRH antiserum restored pituitary expression of luciferase in males passively immunized against GnRH to levels not different from castrate controls. Finally, treatment with both estradiol and D-Ala-6-GnRH increased pituitary expression of luciferase to a level greater than the sum of the individual treatments suggesting synergistic activation of the transgene by these two hormones. Thus, despite the complete absence of transcriptional activity and hormonal responsiveness in vitro, 9,100 bp of proximal promoter from the oGnRHR gene is capable of directing tissue specific expression and is robustly responsive to both GnRH and estradiol in transgenic mice. To begin to refine the functional boundaries of the critical cis acting elements, we next constructed transgenic mice harboring a transgene consisting of 2,700 bp of 5' flanking region from the oGnRHR gene fused to luciferase. As with the -9,100 bp promoter, expression of luciferase in the 2,700 lines was primarily confined to the pituitary gland, brain and testes. Furthermore, the passive immunization-hormonal replacement paradigms described above revealed both GnRH and estradiol responsiveness of the -2,700-bp promoter. Thus, 2,700 bp of proximal promoter from the oGnRHR gene is sufficient for tissue specific expression as well as GnRH and estradiol responsiveness. Given the inability to recapitulate estradiol regulation of GnRHR gene expression in vitro, transgenic mice may represent one of the few viable avenues for ultimately defining the molecular mechanisms underlying estradiol regulation of GnRHR gene expression. PMID- 10698177 TI - Role of the Y1 receptor in the regulation of neuropeptide Y-mediated feeding: comparison of wild-type, Y1 receptor-deficient, and Y5 receptor-deficient mice. AB - Neuropeptide Y (NPY) increases food intake through the action of hypothalamic NPY receptors. At least six subtypes of NPY, peptide YY (PYY), and pancreatic polypeptide (PP) receptors have been identified in mice. Although the involvement of Y1 and Y5 receptors in feeding regulation has been suggested, the relative importance of each of these NPY receptors and the participation of a novel feeding receptor are still unclear. To address this issue, we generated a Y1 receptor-deficient (Y1-/-) and a Y5 receptor-deficient (Y5-/-) mouse line in which we directly compared the orexigenic effects of NPY and its analogs after intracerebroventricular (icv) administration. The icv NPY-induced food intake was remarkably reduced in Y1-/- mice, but was not significantly altered by inactivation of the Y5 receptor. The Y1 receptor therefore plays a dominant role in NPY-induced feeding. Stimulation of feeding by moderately selective Y5 agonists [PYY-(3-36), human PP, and bovine PP] was reduced in Y5-/- mice, although food intake did not decrease to vehicle control levels. These results indicate that the Y5 receptor functions as one of the feeding receptors. In addition, the finding that Y5-preferring agonists still induce food intake in Y5 /- mice suggests a role for another NPY receptor(s), including the possibility of novel NPY receptors. Surprisingly, despite the limited efficacy of PYY-(3-36) and PPs at the Y1 receptor, food consumption induced by these agonists was significantly diminished in Y1-/- mice compared with that in wild-type controls. These observations suggest that the feeding stimulation induced by NPY and its analogs may be directly or indirectly modulated by the action of the Y1 receptor. We conclude that multiple NPY receptors, possibly including the novel feeding receptor, are involved in the feeding response evoked by NPY and its analogs. Among them, the Y1 receptor plays a key role in NPY-induced feeding in mice. PMID- 10698178 TI - Differences in gonadotropin-releasing hormone-induced calcium signaling between melatonin-sensitive and melatonin-insensitive neonatal rat gonadotrophs. AB - The sensitivity of GnRH-stimulated calcium signaling to melatonin, in a subpopulation of neonatal gonadotrophs, is supposed to be attributable to melatonin receptors. However, it is not yet known whether the intracellular pathway for GnRH action in melatonin-sensitive cells is the same as in melatonin insensitive cells. By monitoring intracellular Ca2+ changes as an outward current carried through apamin-sensitive Ca2+-activated K+ channels, we compared GnRH induced calcium responses in these two subpopulations of neonatal gonadotrophs. GnRH induced various oscillatory, as well as nonoscillatory, responses in both cell types that was not related to melatonin sensitivity. Melatonin-sensitive GnRH-induced responses could be clearly distinguished according to the pharmacological properties of their latency. The latency increased in zero extracellular Ca2+ or with the addition of nifedipine, staurosporine, and ryanodine. This effect was only rarely observed in melatonin-insensitive cells. This indicates that there are two pathways for initiation of GnRH-induced calcium signaling in neonatal gonadotrophs. The first pathway is mediated by inositol 1,4,5,-trisphosphate production, whereas the second involves extracellular calcium entry through voltage-dependent L-type Ca2+ channels, protein kinase C activation, and Ca2+ release from a ryanodine-sensitive store, which may coactivate Ca2+ release from an inositol 1,4,5,-trisphosphate-sensitive store. Only the second mechanism is accessible to inhibition by melatonin. PMID- 10698179 TI - Effects of pharmacological and nonpharmacological treatments on thyroid hormone metabolism and concentrations in rat brain. AB - The activities of the 5'I-deiodinase (5'D-I), 5'II deiodinase (5'D-II) and 5III deiodinase (5D-III) isoenzymes and tissue concentrations of thyroxine (T4) and triiodothyronine (T3) were measured in up to 10 regions of the rat brain after acute and subchronic nonpharmacological (sleep deprivation, 12 h fasting, 14 days' calorie-reduced diet) and pharmacological (ethanol, haloperidol, clozapine, lithium, carbamazepine, desipramine, fluoxetine, tranylcypromine, and mianserin) treatments. All of these treatments induced significant and sometimes dramatic changes in 5'D-II activities and tissue concentrations of thyroid hormones and, to a lesser extent, in 5D-III activity. The activity of 5'D-I remained unaffected. The results revealed a surprising specificity for each type of treatment in terms of the isoenzyme and hormone affected, the direction of the change, the brain region affected and the time of day. The changes in thyroid hormone concentrations frequently failed to correspond in any way to those in deiodinase activities and unexpected effects such as inhibition of both 5'D-II and 5D-III were seen, indicating that there may be additional pathways of iodothyronine metabolism in the CNS. In conclusion, particularly 5'D-II activity and thyroid hormone concentrations in the CNS are highly sensitive to many different kinds of influence that may induce changes in neuronal activity. However, these changes in deiodinase activities do not ensure stable tissue concentrations of T3, but were, on the contrary, in most cases accompanied by marked changes T3 levels in the tissue. The implications of these findings for the physiological role of thyroid hormones in the CNS are discussed. PMID- 10698180 TI - Reduced body weight, adipose tissue, and leptin levels despite increased energy intake in female mice lacking acylation-stimulating protein. AB - Acylation-stimulating protein (ASP) is a potent lipogenic protein produced by adipocytes. In vitro studies have shown that ASP increases triglyceride synthesis and glucose transport in both murine and human adipocytes. Our initial study indicated that complement C3-deficient (-/-) mice (and, therefore, ASP deficient) demonstrated altered dietary postprandial triglyceride clearance. In the present study we examined the phenotype of female mice longitudinally on different diets. Female C3(-/-) mice on both low (10% of energy) and high (40% of energy) fat diets displayed an average reduction in total body weight of 10.1+/-0.5% (P < 0.0003, by ANOVA) compared with the C3(+/+) littermates. Reductions in white adipose tissue mass accounted for most of this weight difference (59% reduction; P < 0.01 on low fat diet). Plasma leptin levels were significantly reduced in C3( /-) mice on both high (P < 0.001) and low fat diets (P < 0.01). This reduction was significant even after adjusting for the reduced body weight and body fat (P < 0.001). Leptin reductions in the C3(-/-) were greater on the high fat diet and were associated with increased food intake (18+/-2% increase; P < 0.001). Furthermore, there was a decrease in basal glucose levels and basal insulin levels [12.8% decrease in glucose at 14 weeks (HF; P < 0.05) and 41% decrease in insulin at 26 weeks (HF; P < 0.05)]. These in vivo experiments demonstrate that female mice lacking ASP have marked alterations of body weight, adiposity, plasma leptin, and plasma insulin levels. Decreased adiposity and leptin levels occurred in the ASP-deficient animals despite increased energy intake, suggesting that energy expenditure was elevated in these animals. Thus, ASP appears to have an important role in the regulation of energy balance in mice. PMID- 10698181 TI - Prostaglandins mediate the endotoxin-induced suppression of pulsatile gonadotropin-releasing hormone and luteinizing hormone secretion in the ewe. AB - Five experiments were conducted to test the hypothesis that PGs mediate the endotoxin-induced inhibition of pulsatile GnRH and LH secretion in the ewe. Our approach was to test whether the PG synthesis inhibitor, flurbiprofen, could reverse the inhibitory effects of endotoxin on pulsatile LH and GnRH secretion in ovariectomized ewes. Exp 1-4 were cross-over experiments in which ewes received either flurbiprofen or vehicle 2 weeks apart. Jugular blood samples were taken for LH analysis throughout a 9-h experimental period. Depending on the specific purpose of the experiment, flurbiprofen or vehicle was administered after 3.5 h, followed by endotoxin, vehicle, or ovarian steroids (estradiol plus progesterone) at 4 h. In Exp 1, flurbiprofen reversed the endotoxin-induced suppression of mean serum LH concentrations and the elevation of body temperature. In Exp 2, flurbiprofen prevented the endotoxin-induced inhibition of pulsatile LH secretion and stimulation of fever, reduced the stimulation of plasma cortisol and progesterone, but did not affect the rise in circulating tumor necrosis factor alpha. In Exp 3, flurbiprofen in the absence of endotoxin had no effect on pulsatile LH secretion. In Exp 4, flurbiprofen failed to prevent suppression of pulsatile LH secretion induced by luteal phase levels of the ovarian steroids progesterone and estradiol, which produce a nonimmune suppression of gonadotropin secretion. In Exp 5, flurbiprofen prevented the endotoxin-induced inhibition of pulsatile GnRH release into pituitary portal blood. Our finding that this PG synthesis inhibitor reverses the inhibitory effect of endotoxin leads to the conclusion that PGs mediate the suppressive effects of this immune/inflammatory challenge on pulsatile GnRH and LH secretion. PMID- 10698182 TI - An increased intraovarian synthesis of nerve growth factor and its low affinity receptor is a principal component of steroid-induced polycystic ovary in the rat. AB - A form of polycystic ovary (PCO) resembling some aspects of the human PCO syndrome can be induced in rats by a single injection of estradiol valerate (EV). An increase in sympathetic outflow to the ovary precedes, by several weeks, the appearance of cysts, suggesting the involvement of a neurogenic component in the pathology of this ovarian dysfunction. The present study was carried out to test the hypotheses that this change in sympathetic tone is related to an augmented production of ovarian nerve growth factor (NGF), and that this abnormally elevated production of NGF contributes to the formation of ovarian cysts induced by EV. Injection of the steroid resulted in increased intraovarian synthesis of NGF and its low affinity receptor, p75 NGFR. The increase was maximal 30 days after EV, coinciding with the elevation in sympathetic tone to the ovary and preceding the appearance of follicular cysts. Intraovarian injections of the retrograde tracer fluorogold combined with in situ hybridization to detect tyrosine hydroxylase (TH) messenger RNA-containing neurons in the celiac ganglion revealed that these changes in NGF/p75 NGFR synthesis are accompanied by selective activation of noradrenergic neurons projecting to the ovary. The levels of RBT2 messenger RNA, which encodes a beta-tubulin presumably involved in slow axonal transport, were markedly elevated, indicating that EV-induced formation of ovarian cysts is preceded by functional activation ofceliac ganglion neurons, including those innervating the ovary. Intraovarian administration of a neutralizing antiserum to NGF in conjunction with an antisense oligodeoxynucleotide to p75 NGFR, via Alzet osmotic minipumps, restored estrous cyclicity and ovulatory capacity in a majority of EV-treated rats. These functional changes were accompanied by restoration of the number of antral follicles per ovary that had been depleted by EV and a significant reduction in the number of both precystic follicles and follicular cysts. The results indicate that the hyperactivation of ovarian sympathetic nerves seen in EV-induced PCO is related to an overproduction of NGF and its low affinity receptor in the gland. They also suggest that activation of this neurotrophic-neurogenic regulatory loop is a component of the pathological process by which EV induces cyst formation and anovulation in rodents. The possibility exists that a similar alteration in neurotrophic input to the ovary contributes to the etiology and/or maintenance of the PCO syndrome in humans. PMID- 10698183 TI - Intraovarian excess of nerve growth factor increases androgen secretion and disrupts estrous cyclicity in the rat. AB - A single injection of estradiol valerate induces a form of cystic ovary resembling some aspects of the human polycystic ovarian syndrome. Preceding the development of follicular cysts, there is an increase in intraovarian synthesis of nerve growth factor (NGF) and the low affinity NGF receptor (p75 NGFR). Selective blockade of NGF actions and p75 NGFR synthesis in the ovary restored estrous cyclicity and ovulatory capacity in estradiol valerate-treated rats, suggesting that an increase in NGF-dependent, p75 NGFR-mediated actions within the ovary contributes to the development of cystic ovarian disease. We have tested this hypothesis by grafting NGF-producing neural progenitor cells into the ovary of juvenile rats that have been induced to ovulate precociously by a single injection of PMSG. The NGF-producing cells, detected by their content of immunoreactive p75 NGFR material, were found scattered throughout the ovary with some of them infiltrating the granulosa cell compartment of large, precystic follicles. Ovarian NGF content was 2-fold higher than in the ovary of rats receiving control cells. Estrous cyclicity was disrupted, with the animals showing prolonged periods of persistent estrus, and an almost continuous background of vaginal cornified cells at other phases of the estrous cycle. Morphometric analysis revealed that the presence of NGF-producing cells neither reduced the total number of corpora lutea per ovary nor significantly increased the formation of follicular cysts. However, the ovaries receiving these cells showed an increased incidence of precystic, type III follicles, accompanied by a reduced number of healthy antral follicles, and an increased size of both healthy and atretic follicles. These changes in follicular dynamics were accompanied by a selective increase in serum androstenedione levels. The results show that an abnormally elevated production of NGF within the ovary suffices to initiate several of the structural and functional alterations associated with the development of follicular cysts in the rat ovary. PMID- 10698184 TI - Subunit composition and pharmacological characterization of gamma-aminobutyric acid type A receptors in frog pituitary melanotrophs. AB - The frog pars intermedia is composed of a single population of endocrine cells directly innervated by gamma-aminobutyric acid (GABA)ergic nerve terminals. We have previously shown that GABA, acting through GABA(A) receptors, modulates both the electrical and secretory activities of frog pituitary melanotrophs. The aim of the present study was to take advantage of the frog melanotroph model to determine the relationship between the subunit composition and the pharmacological properties of native GABA(A) receptors. Immunohistochemical labeling revealed that in situ and in cell culture, frog melanotrophs were intensely stained with alpha2-, alpha3-, gamma2-, and gamma3-subunit antisera and weakly stained with a gamma1-subunit antiserum. Melanotrophs were also immunolabeled with a monoclonal antibody to the beta2/beta3-subunit. In contrast, frog melanotrophs were not immunoreactive for the alpha1-, alpha5-, and alpha6 isoforms. The effects of allosteric modulators of the GABA(A) receptor on GABA activated chloride current were tested using the patch-clamp technique. Among the ligands acting at the benzodiazepine-binding site, clonazepam (EC50, 5 x 10(-9) M), diazepam (EC50, 10(-8) M), zolpidem (EC50, 3 x 10(-8) M), and beta-carboline 3-carboxylic acid methyl ester (EC50, 10(-6) M) were found to potentiate the whole cell GABA-evoked current in a dose-dependent manner. Methyl-6,7-dimethoxy-4 ethyl-beta-carboline-3-carboxylate (IC50, 3 x 10(-5) M) inhibited the current, whereas Ro15-4513 had no effect. Among the ligands acting at other modulatory sites, etomidate (EC50, 2 x 10(-6) M) enhanced the GABA-evoked current, whereas 4'-chlorodiazepam (IC50, 4 x 10(-7) M), ZnCl2 (IC50, >5 x 10(-5) M), and furosemide (IC50, >3 x 10(-4) M) depressed the response to GABA. PK 11195 did not affect the GABA-evoked current or its inhibition by 4'-chlorodiazepam. The results indicate that the native GABA(A) receptors in frog melanotrophs are formed by combinations of alpha2-, alpha3-, beta2/3-, gamma1-, gamma2-, and gamma3-subunits. The data also demonstrate that clonazepam is the most potent, and zolpidem is the most efficient positive modulator of the native receptors. Among the inhibitors, 4'-chlorodiazepam is the most potent, whereas ZnCl2 is the most efficient negative modulator of the GABA(A) receptors. The present study provides the first correlation between subunit composition and the functional properties of native GABA(A) receptors in nontumoral endocrine cells. PMID- 10698185 TI - Modulation of guanosine triphosphatase activity of G proteins by arachidonic acid in rat Leydig cell membranes. AB - Previous results from our group have indicated that arachidonic acid decrease cAMP production through a modification of heterotrimeric G proteins. In the present study, we have characterized the high affinity GTPase activity present in Leydig cell membranes and its regulation by fatty acids. The high-affinity GTPase activity, measured as [gamma32P] GTP hydrolysis rate, was both time and protein concentration dependent. Arachidonic acid elicited a dose-dependent inhibition of enzyme activity with an IC50 = 26.7+/-1.1 microM. The existence of only two double bonds in linoleic acid is reflected by a decrease in its inhibitory activity (IC50 = 34+/-2.3 microM). Saturated fatty acids showed no effect at this level. The kinetic analysis as interpreted by Lineweaver-Burk plots, indicated that 50 microM arachidonic acid had no effect on the apparent affinity for GTP, but resulted in a 40% decreases in the maximal velocity of the reaction. Arachidonic acid modulation of GTPase activity was not attenuated by blocking eicosanoid metabolism with inhibitors of 5'-lipoxygenase, cyclooxygenase, or epoxygenase P-450. The addition of arachidonic acid to pertussis toxin-treated membranes had no effect on the enzyme activity, indicating that arachidonic acid does not modify the GTPase activity present in Galphas protein. However, ADP ribosylation with cholera toxin followed by arachidonic acid treatment led to a further 40% inhibition when compared with cholera toxin treatment alone. These results allowed us to postulate that arachidonic acid inhibits the GTPase activity of Gi protein family. To further analyze the mechanism of arachidonic acid inhibition of GTPase activity, the effect of arachidonic acid on the [35S]GTPgammaS binding was studied. No effect of this fatty acid on GTP binding was found. Combining our previous results with those found here, we can conclude that arachidonic acid maintains Gi proteins in their active state, which in turn inhibit adenylate cyclase and results in decrease cAMP levels. PMID- 10698186 TI - Thrombospondin and osteopontin bind to insulin-like growth factor (IGF)-binding protein-5 leading to an alteration in IGF-I-stimulated cell growth. AB - Insulin-like growth factor (IGF)-binding protein-5 (IGFBP-5) has been shown to bind to extracellular matrix (ECM) with relatively high affinity, but the ECM components that mediate this interaction have not been identified. These studies show that radiolabeled IGFBP-5 specifically coprecipitates with two ECM proteins, thrombospondin-1 (TSP-1) and osteopontin (OPN). As TSP-1 binds avidly to heparin, as does IGFBP-5, the effect of glycosaminoglycans on the TSP-1/IGFBP-5 interaction was analyzed. Heparan and dermatan sulfate inhibited binding, whereas heparin increased binding. Chondroitin sulfate A and B had no effect. In contrast, both heparin and heparan sulfate significantly inhibited the OPN-IGFBP 5 interaction and chondroitin sulfate A, B, and C had no effect. To determine the region of IGFBP-5 that was involved in each interaction, synthetic peptides that spanned several regions of IGFBP-5 were tested for their capacity to competitively inhibit coprecipitation. A peptide that contained the amino acids between positions 201 and 218 resulted in 76% and 86% inhibition of binding to TSP-1 and OPN, respectively. Three other synthetic peptides that spanned regions ofIGFBP-5 with several charged residues had no effect. IGFBP-5 mutants that contained substitutions for basic residues in the 201-218 region were tested for their ability to bind to TSP-1 or OPN. A mutant with substitutions for amino acids at positions R201 and K202 and a mutant with substitutions for K211, R214, K217, and R218 had the greatest reduction in binding to TSP-1. Mutants containing substitutions for R214 alone and the combined K217A, R218A mutant had the greatest reductions in OPN binding. When the smooth muscle cell growth response to these components was assessed, IGF-I plus IGFBP-5 or the combination of TSP-1 or OPN with IGF-I potentiated the IGF-I effect. The addition of IGFBP-5 to these combinations resulted in further significant growth stimulation. Both OPN and TSP 1 specifically bind to IGFBP-5 with high affinity. These interactions may be important for concentrating intact IGFBP-5 in extracellular matrix and for modulating the cooperative interaction between the IGF-I receptor and integrin receptor signaling pathways. PMID- 10698187 TI - Pregnancy-dependent changes in cell signaling underlie changes in differential control of vasodilator production in uterine artery endothelial cells. AB - During pregnancy, the uterine vasculature shows a marked increase in vasodilator production [prostacyclin (PGI2) and nitric oxide (NO)] in response to a number of agonists including angiotensin II (AII) and ATP. As a consequence vascular resistance is kept low, and uterine blood flow is maximized to meet the needs of the growing fetus. Studies of the molecular basis underlying this change in control of endothelial NO and PGI2 production have been hampered by the lack of availability of a suitable cell model. To that end we have developed and characterized a new ovine uterine artery endothelial cell (UAEC) culture model derived from nonpregnant (NP) or pregnant (P) ewes. Endothelial cells were isolated from pregnant (120-130 days; n = 6) and nonpregnant (n = 4) ewes and maintained in primary culture. Endothelial cells at passage 4 showed uniform expression of endothelial nitric oxide synthase (eNOS; an endothelial marker) as well as AII type 1 receptor and growth factor receptors and uniform uptake of acetylated low density lipoprotein (a property of endothelial cells not shared by fibroblasts or vascular smooth muscle cells), thus demonstrating cell purity. Expressions of eNOS, cyclooxygenase-1, PGI2 synthase, cytosolic phospholipase A2, AII type 1 receptor, and growth factor receptors are also maintained at passage 4. Mitogenesis is maintained in response to basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), and vascular endothelial growth factor (VEGF) in both NP-UAEC and P-UAEC. The differential production of vasodilators by NP-UAEC and P-UAEC is maintained in a manner similar to that previously reported in vivo. Thus, P-UAEC make NO in response to AII, ATP, bFGF, EGF, and VEGF, whereas NP-UAEC make NO in response to bFGF, EGF, and VEGF only. Similarly, P UAEC make PGI2 in response to AII, ATP, bFGF, and VEGF, whereas NP-UAEC make PGI2 only in response to ATP and VEGF. As both cytosolic phospholipase A2 and eNOS may be regulated by both Ca2+ and protein kinases, we investigated the effects of these agonists on Ca2+ mobilization and ERK-1/2 phosphorylation. ATP consistently elevates Ca2+ levels in both P-UAEC and NP-UAEC. All other agonists were without acute (0-4 min) effect on Ca2+ in P-UAEC or NP-UAEC. In contrast, all agonists stimulated an acute (10 min) phosphorylation of ERK-1/2 in P-UAEC, whereas only EGF stimulated activation in NP-UAEC. P-UAEC production of PGI2 by agonists of both heptahelical receptors and growth factor receptors correlates closely with ERK-2 phosphorylation alone. For NO, this correlation holds for heptahelical receptor agonists, but additional signaling pathways are also implicated for bFGF and VEGF. In contrast, in NP-UAEC the lack of ERK-2 phosphorylation in response to all agonists other than EGF, and the dissociation between NO or PGI2 production and ERK-2 phosphorylation suggest that alternate pathways play a predominant role. PMID- 10698188 TI - Two putative GATA motifs in the proximal exon 1 promoter of the rat insulin-like growth factor I gene regulate basal promoter activity. AB - The insulin-like growth factor I gene is transcribed from two promoters, which direct synthesis of alternative first exons (exon 1 and exon 2) in insulin-like growth factor I messenger RNAs (mRNAs). An exon 1 promoter construct extending from +75 to +192 (the most upstream exon 1 start site was designated as +1) showed significant promoter activity in C6, OVCAR-3, and SK-N-MC cells. Within the +75 to +192 region, there are two perfect matches to the consensus binding site for GATA transcription factor, at +108 (GATA-A) and at +183 (GATA-B). Mutations of the GATA-A or GATA-B sequences resulted in slight (or no) effect on exon 1 promoter activity in both C6 and OVCAR-3 cells. However, mutation of the GATA-A sequence stimulated exon 1 promoter activity by 68% in SK-N-MC cells. Mutation of the GATA-B sequence inhibited exon 1 promoter activity by 4.4-fold in SK-N-MC cells. Electrophoretic mobility shift assays showed that there were nuclear proteins in SK-N-MC cells capable of specifically binding to the GATA-A and GATA-B elements and that this binding was GATA-sequence specific. GATA-2, GATA-3, and GATA-4 are the only GATA proteins that have been reported to be expressed in neurons. None of the antibodies against these three GATA proteins were capable of inhibiting or supershifting the bands formed by the nuclear proteins and oligonucleotides containing GATA-A or GATA-B elements. A GATA-1 expression vector was used to perform cotransfection experiments. The GATA-A mutation abolished the stimulatory effect of the GATA-1 factor on promoter activity. In contrast, the GATA-B mutation enhanced the stimulatory effect of GATA-1 protein. Anti-GATA-1 antibody was also incapable of inhibiting or supershifting the bands formed by the nuclear proteins and oligonucleotides containing the GATA-A or GATA-B elements. In conclusion, the GATA-A element seems to bind an inhibitory endogenous factor(s) in SK-N-MC cells, whereas the GATA-B element may bind a stimulatory factor(s). These factors seem to be related to GATA transcription factors but are immunologically distinct from GATA-2, GATA-3, or GATA-4. GATA-1 has the potential to transactivate the exon 1 promoter through the GATA-A element but is unlikely to be the endogenous protein binding to the GATA-A or the GATA-B motifs in SK-N-MC cells. PMID- 10698189 TI - Substrate-induced down-regulation of human type 2 deiodinase (hD2) is mediated through proteasomal degradation and requires interaction with the enzyme's active center. AB - Type 2 iodothyronine deiodinase (D2) catalyzes the first step in thyroid hormone action, the deiodination of T4 to T3. Endogenous D2 activity is posttranslationally regulated by substrate that accelerates its degradation through the ubiquitin-proteasome pathway. To understand how D2 activity correlates with D2 protein during its normal decay and rT3-induced down regulation, HEK-293 cells, transiently expressing human D2, were labeled with Na75SeO3 and then treated with 100 microM cycloheximide (CX), 30 nM rT3, and/or 10 microM MG132, a specific proteasome inhibitor, for 2-4 h. D2 protein and enzyme activity changed in parallel, disappearing with a half-life of 2 h in the presence of CX, or 1 h when CX + rT3 were combined. Treatment with MG132 blocked these effects. We created selenocysteine (Sec) 133 to cysteine (Cys) or alanine (Ala) D2 mutants, without changing Sec 266. The CysD2 activity and protein levels were also parallel, with a similar half-life of approximately 2 h, whereas the rT3-induced D2 down-regulation required approximately 1000-fold higher rT3 concentration (30 microM) due to a proportionally higher Michaelis constant of CysD2. In similar experiments, the AlaD2 mutant retained the short half-life but was not catalytically active and not susceptible to rT3-accelerated degradation. We conclude that substrate-induced loss of D2 activity is due to proteasomal degradation of the enzyme and requires interaction with the catalytic center of the protein. PMID- 10698190 TI - Leukemia inhibitory factor expression and regulation within the testis. AB - Leukemia inhibitory factor (LIF) is a pleiotropic cytokine known to control the proliferation and survival of stem cells including primordial germ cells and gonocytes. This led us to study the origin and regulation of testicular LIF. The LIF transcript was detected in the rat testis by RT-PCR from 13.5 days postcoitum until adulthood. LIF expression was investigated further in vitro in seven different highly purified testicular cell populations using RT-PCR and bioassays combined with neutralization experiments. LIF was found to be produced by peritubular cells and, to a much lesser extent, by the other testicular somatic cell types. No LIF was detected in meiotic and postmeiotic germ cell-conditioned medium, and only low levels of LIF were detected in spermatogonia-conditioned medium. Large amounts of bioactive LIF were measured in testicular lymph. While LIF production was greatly enhanced in presence of serum, lipopolysaccharide, and TNFalpha further increased this production in peritubular and Sertoli cells, and human CG enhanced Leydig cell LIF production. In conclusion, peritubular cells are the principal source of testicular LIF, probably accounting for its high concentration in the lymph. Given the proliferative effect of LIF on immature germ cells, we suggest that peritubular LIF plays an important role in the regulation of testicular function. PMID- 10698191 TI - Impaired basal and restraint-induced epinephrine secretion in corticotropin releasing hormone-deficient mice. AB - CRH is thought to play a role in responses of the adrenocortical and adrenomedullary systems during stress. To investigate the role of CRH in stress induced secretions of corticosterone and epinephrine, we subjected wild-type (WT) and CRH-deficient (knockout, KO) mice to restraint, and analyzed plasma corticosterone, plasma catecholamines, and adrenal phenylethanolamine N methyltransferase (PNMT) gene expression and activity before and during 3 h of restraint. Plasma corticosterone increased over 40-fold in WT mice, but minimally in CRH KO mice. Adrenal corticosterone content tended to increase in CRH KO mice, although to levels 5-fold lower than that in WT mice. CRH KO mice had significantly lower plasma epinephrine and higher norepinephrine than WT mice at baseline, and delayed epinephrine secretion during restraint. Adrenal PNMT messenger RNA content in CRH KO mice tended to be lower than that in WT mice, though the degree of induction was similar in both genotypes. PNMT enzyme activity was significantly lower in CRH KO mice. Pharmacological adrenalectomy abolished restraint-induced corticosterone secretion and PNMT gene expression in WT mice, consistent with an absolute requirement of glucocorticoids for PNMT gene expression. We conclude that glucocorticoid insufficiency in CRH KO mice leads to decreased basal and restraint-induced plasma epinephrine and adrenal PNMT gene expression and enzyme activity. PMID- 10698192 TI - Increased and persistent circulating insulin-like growth factor II in neonatal transgenic mice suppresses developmental apoptosis in the pancreatic islets. AB - In rats, a proportion of pancreatic beta-cells are deleted by apoptosis in the second week of postnatal life and replaced by endocrine cell neogenesis from pancreatic ductal epithelium. This coincides with a reduction in pancreatic insulin-like growth factor II (IGF-II) expression, and IGF-II has been shown to act as a beta-cell survival factor in vitro. To examine whether IGF-II regulates beta-cell apoptosis in vivo, an IGF-II transgenic mouse model was used in which mouse IGF-II is overexpressed in skin, gut, and uterus driven by a keratin promoter, so that circulating IGF-II is retained postnatally. Mice were killed between postnatal days 7 and 26, and the pancreas was examined histologically. Apoptotic cells were visualized by the terminal deoxynucleotidyltransferase mediated deoxy-UTP nick end labeling method, and proliferating cells were examined by immunohistochemistry for proliferating cell nuclear antigen. In nontransgenic mice, serum IGF-II was absent by 26 days, but mean (+/-SEM) values were 45+/-9 ng/ml (n = 5) in transgenic animals. A 2- to 3-fold rise in islet cell apoptosis was seen in normal animals between days 11 and 16, but this was substantially decreased in IGF-II transgenic mice (day 11; control, 12+/-1%; transgenic, 6+/-1%; P < 0.01; n = 5). Consequently, islets from IGF-II transgenic mice had a significantly greater mean area from days 11-16, but the proportions of beta- and alpha-cells and circulating insulin levels were not changed. Islet cell DNA synthesis was increased in transgenic mice on days 13 and 16. The total islet number per section did not alter. The results show that a persistent presence of circulating IGF-II postnatally alters endocrine pancreatic ontogeny in the mouse and largely prevents the wave of developmental apoptosis that precipitates beta-cell turnover in neonatal life. PMID- 10698193 TI - Pituitary adenylate cyclase-activating polypeptide (PACAP) and PACAP-receptor type 1 expression in rat and human placenta. AB - Pituitary adenylate cyclase-activating polypeptide (PACAP), the new hypophysiotropic factor member of the vasoactive intestinal peptide (VIP)/secretin/glucagon/GHRH family of neuropeptides, exerts its biological action by interacting with both PACAP-selective type I receptors (PAC1) and type II receptors (VPAC1), which bind both PACAP and VIP. The placenta is a site of production of hypophysiotropic factors that participate in the control of local hormone production, as well as the respective hypothalamic-pituitary neurohormones. In the present study, we show the expression of PACAP gene and irPACAP distribution within rat and human placental tissues, by means of RT-PCR and immunohystochemical experiments. In both rat and human placenta, we evaluated the expression of PAC1 gene by Northern hybridization analysis performed with a 32P-labeled 706 nt complementary DNA probe, derived from the full-length coding region of the rPAC1 complementary DNA. The results of these experiments demonstrate the presence, in both human and rat placenta, of a 7.5-kb transcript similar in size to those detected in the ovary, brain, and hypothalamus. Alternative splicing of two exons occurs in human and rat PAC1 gene generating splice variants with variable tissue-specific expression. To ascertain which of the splice variants were expressed in placental tissue we performed RT-nested PCR using primers flanking the insertion sequence termed hip/hop cassette in rat or SV1/SV2 box in human gene. Electrophoretic analysis of the PCR products showed a different pattern of expression of messenger RNA splicing variants in human and rat placenta. In particular, the rat placenta expresses the short PAC1 receptor (PAC1short), the rPAC1-hip or hop (which are indistinguishable with the primers used), and the rPAC1-hip-hop, whereas the human placenta expresses only the PAC1SV1 (or SV2) variant, structurally homologous to the rat PAC1 hip (or hop). Sequence analysis of the human PCR-amplified PAC1 variant was therefore carried out and revealed that human placenta only expresses the PAC1SV2 isoform. The presence and characterization of PACAP binding sites was then investigated in human placenta by radioligand binding studies performed on crude membrane preparation using [125I]PACAP27 as tracer. Scatchard analysis of the binding results revealed the presence of two binding sites, one with high affinity and low capacity (Kd 0.33+/-0.04 nM; Bmax 36.9+/-12.1 fmol/mg protein) and one with low affinity and high capacity (Kd 24+/-6.9 nM, Bmax 9.3+/-0.19 pmol/mg protein). The relative potencies of PACAP-related peptides for inhibition ofradioligand binding were: PACAP27 > or = PACAP38 > VIP, whereas GHRH and other unrelated peptides, such as CRH and beta-endorphin, did not inhibit [125I]PACAP27 binding. In conclusion, in this study, we provide evidence for the expression of PACAP within rat and human placenta. We also demonstrate that both human and rat placenta express the PAC1 gene and that the human tissue has binding sites for PACAP. These findings may suggest a role for PACAP in the regulation of placental physiology through autocrine and/or paracrine mechanisms. PMID- 10698194 TI - Meiotic arrest and germ cell apoptosis in androgen-binding protein transgenic mice. AB - The fundamental role of androgen-binding protein (ABP) in spermatogenesis remains obscure after nearly 25 yr since its first characterization. In the present investigation, we used a transgenic mouse model that overexpresses rat ABP to examine the potential involvement of this protein in the regulation of processes occurring during spermatogenesis. Specifically, homozygous or heterozygous transgenic mice were analyzed in terms of spermatogenic progression, DNA fragmentation pattern, and germinal cell ploidy status. All animals homozygous for transgenic ABP exhibited an increased accumulation of primary spermatocytes and cells at metaphase with abnormal morphology and localization within the seminiferous epithelium. Analysis of DNA fragmentation by in situ techniques and agarose gel electrophoresis provided evidence for an increased occurrence of apoptosis in the transgenic animals, principally involving pachytene spermatocytes and cells at metaphase. Flow cytometric analysis of the DNA content of isolated germ cells revealed a reduction in the number of haploid cells, an increase in the number of tetraploid cells, and the appearance of a hypotetraploid cell population, consistent with degenerating primary spermatocytes. In mice heterozygous for the transgene, the effects were less prominent, and the degree to which spermatogenesis was compromised correlated with the levels of ABP messenger RNA in individual animals. The present results are interpreted to suggest that ABP can act as a modulator of spermatogenesis by regulating completion of the first meiotic division of primary spermatocytes. PMID- 10698195 TI - Gonadal steroids regulate the number and activational state of mast cells in the medial habenula. AB - While mast cells in connective tissues have long been associated with allergic reactions, it is now clear that they are also present within the central nervous system under normal physiological conditions. The mast cell population increases 10-fold in the medial habenular region of the brain within 2 h after pairing in doves. The first study explored whether this increase was due to exposure to gonadal steroids. Light microscopic immunocytochemistry indicates an increased number of brain MC following exposure to either testosterone (T) or dihydrotestosterone (DHT) in the male, or 17beta estradiol (E) in the female, but not in cholesterol-treated controls. Thus, the increased habenular MC population is produced by gonadal hormones in the absence of sexual behavior, is not sexually dimorphic, and does not require aromatization of androgen. In the next study, MC activational state was determined using electron microscopy. Cells were categorized into five states: (I) resting; (II) initiation of degranulation; (III) fully degranulated; (IV) piecemeal secretion; and (V) resynthesizing. Hormone treatment (T, DHT, or E) resulted in a significant increase in the percent of cells in activated states. MC granules contain a wide range of biologically active molecules. The release of these granule contents into the neuropil of the central nervous system is likely to have wide ranging effects at multiple levels including vascular permeability and neuronal excitability. In that steroid treatment is known to result in such effects, the present demonstration of a hormonally induced shift in MC secretory state is one avenue by which these effects are mediated. PMID- 10698196 TI - Local regulation of gonadotroph function by pituitary gonadotropin-releasing hormone. AB - Cultured rat pituitary cells and immortalized pituitary gonadotrophs (alphaT3-1 cells) express specific messenger RNA transcripts for GnRH and exhibit positive immunostaining for the GnRH peptide. Each cell type released GnRH during both static culture and perifusion, albeit in lesser amounts than cultured hypothalamic cells and GT1-7 neurons. In perifused pituitary cells, exposure to a GnRH agonist stimulated the release of GnRH as well as LH. In contrast, treatment with a GnRH receptor antagonist or with GnRH antiserum decreased basal LH release. In pituitary cell cultures, a small proportion of gonadotrophs exhibited high amplitude and low frequency baseline Ca2+ oscillations in the absence of GnRH stimulation. Such spontaneous oscillations were comparable to those induced by picomolar concentrations of GnRH and could be abolished by treatment with a GnRH antagonist. These in vitro findings indicate that locally produced GnRH causes low level activation of pituitary GnRH receptors, induces spontaneous intracellular Ca2+ oscillations, and contributes to basal LH secretion in cultured pituitary cells. In vivo, such autocrine or paracrine actions of pituitary-derived GnRH could provide a mechanism for the maintenance of optimal responsiveness of the gonadotrophs to pulses of GnRH arising in the hypothalamus. The presence and actions of GnRH in the anterior pituitary gland, the major site of expression of GnRH receptors, suggest that local regulatory effects of the neuropeptide could supplement the primary hypothalamic mechanism for the control of episodic gonadotropin secretion. PMID- 10698197 TI - The renal expression of transforming growth factor-beta isoforms and their receptors in acute and chronic experimental diabetes in rats. AB - Transforming growth factors-beta (TGF-beta) are fibrogenic factors that have been strongly implicated in the development of diabetic nephropathy. Our aim was to use two animal models [the streptozotocin (STZ)-induced diabetic rat and the genetically prone biobreeding (BB) rat] to fully characterize the responses of the renal TGF-beta system in both short- and long-term diabetes. In this study changes in the entire renal TGF-beta system, at both protein and messenger RNA (mRNA) levels, have been characterized using the techniques of immunocytochemistry, Western blotting, and ribonuclease protection assay. We also used Western blotting of pro-collagen-I C-peptide to demonstrate that the rate of fibrogenesis was highest over the first 2 weeks of diabetes. TGF-beta1, TGF beta2, and receptor mRNA and protein were detected in the control nondiabetic kidney. It was found that dramatic and dynamic changes occur in all parts of the renal TGF-beta axis in both models of experimental diabetes, but TGF-beta2 and TGF-betaRII proteins were the predominant responsive element, particularly during the acute phase of disease. For example, during the acute phase of disease (0-30 days), although renal TGF-beta1 mRNA levels were elevated, no increases in the corresponding protein were detected in the kidney. By contrast, in the absence of changes in TGF-beta2 mRNA levels, twice as much TGF-beta2 protein was measured in the kidney by day 30 of STZ-induced diabetes compared with day 0 controls analyzed by Western blotting (P < 0.05), and the protein was localized both to the nuclei and cytoplasm of glomerular cells, analyzed by immunocytochemistry. In addition, three times as much TGF-betaRII protein was found by day 90 of STZ induced diabetes compared with day 0 controls, making this the most responsive receptor type. These results suggest that the entire TGF-beta axis has a role in the etiology of kidney fibrosis and could be manipulated therapeutically to preserve kidney function. PMID- 10698198 TI - Involvement of insulin-like growth factors in early T cell development: a study using fetal thymic organ cultures. AB - The expression of insulin-like growth factor (IGF) and IGF receptor genes was investigated by RT-PCR during ontogeny of the murine thymus. IGF-1, IGF-1R, M6P/IGF-2R genes are expressed in the thymus both in fetal and postnatal life, whereas IGF-2 messenger RNAs (mRNAs) decline after birth but are still detectable on the seventh week. By in situ hybridization, IGF-2 transcripts were located in the outer cortex and medulla of the postnatal thymus, and on the whole surface ofthe epithelial-like network in the fetal thymus. The effects of anti-IGFs and IGF-receptors neutralizing Abs on the generation of pre-T cell subpopulations were then investigated using fetal thymic organ cultures (FTOC). FTOC treatment with an anti-IGF-2 mAb, an anti-IGF-1R mAb, or an anti-M6P/IGF-2R polyclonal Ab induced a blockade of T cell differentiation at the CD4-CD8- stage, as shown by a significant increase in the percentage of CD4-CD8- cells and a decrease in the percentage of CD4+CD8+ cells. Moreover, anti-IGF-2 Ab treatment induced an increase in CD8+ cells suggesting that thymic IGF-2 might have a role in determining differentiation into the CD4 or CD8 lineage. Anti-IGF-1 Ab treatment decreased the proportion in CD4-CD8- cells and increased the frequency in CD4+CD8+. FTOC treatment with anti-(pro)insulin did not exert any significant effect on T cell development. These data indicate that the intrathymic IGF mediated signaling plays an active role in the early steps of T cell differentiation during fetal development. PMID- 10698199 TI - Pituitary adenylate cyclase-activating polypeptide gene expression regulated by a testis-specific promoter in germ cells during spermatogenesis. AB - Pituitary adenylate cyclase-activating polypeptide (PACAP) is a member of the glucagon-related family of hormones that is widely expressed in various tissues. The PACAP messenger RNA (mRNA) and protein is expressed at high levels in the germ cells of the testis, where it locally activates cAMP-coupled receptors located in the somatic Sertoli cells. The PACAP mRNA expressed specifically in the testis is shorter than the mRNA expressed in hypothalamus and includes 127 nucleotides of novel sequence at the 5'-end, suggesting a different start site of transcription in the testes and the utilization of a tissue-specific promoter. Here we present evidence that a single PACAP gene uses a testis-specific promoter to express a mRNA containing a unique exon located 13.5 kb upstream from the first coding exon. As determined by RT-PCR analysis of testis mRNA, the expression of the first testis-specific exon is relatively specific for the testis, as no PACAP mRNA containing the testis-specific first exon was detected in hypothalamic mRNAs. The promoter for the testis-specific PACAP gene was cloned, and a start site for transcription was mapped by primer extension. The testis-specific promoter sequence directs germ cell-specific expression upon transfection of promoter-transcriptional reporter plasmids to populations of testicular cells in vitro and upon expression of a promoter-reporter transgene in mice. Analyses of PACAP gene expression during the spermatogenic cycle, accomplished by RT-PCR of segments of isolated seminiferous tubules, identified intense expression in the postmeiotic round spermatids during developmental stages I-VIII. These observations establish the existence of a specialized PACAP gene promoter whose activity is highly regulated during the spermatogenic cycle. PMID- 10698200 TI - Osteoblast-derived cells express functional glucose-dependent insulinotropic peptide receptors. AB - Glucose-dependent insulinotropic peptide (GIP) is a 42-amino acid peptide synthesized and secreted from endocrine cells in the small intestine. The role of GIP in coupling nutrient intake and insulin secretion, the incretin effect, is well known. We report that GIP receptor messenger RNA and protein are present in normal bone and osteoblast-like cell lines, and that high affinity receptors for GIP can be demonstrated by [125I]GIP binding studies. When applied to osteoblast like cells (SaOS2), GIP stimulated increases in cellular cAMP content and intracellular calcium, with both responses being dose dependent. Moreover, administration of GIP results in elevated expression of collagen type I messenger RNA as well as an increase in alkaline phosphatase activity. Both of these effects reflect anabolic actions of presumptive osteoblasts. These results provide the first evidence that GIP receptors are present in bone and osteoblast like cells and that GIP modulates the function of these cells. PMID- 10698201 TI - Chimeric and point-mutated receptors reveal that a single glycine residue in transmembrane domain 6 is critical for high affinity melatonin binding. AB - To delineate domains of high affinity melatonin receptors that are essential for melatonin binding, we generated chimeras between the human Mel1a melatonin receptor and the melatonin-related orphan H9 receptor. The latter receptor displays no high affinity melatonin binding. The chimeric receptors were transiently expressed in COS-7 cells and analyzed by radioligand binding using 2 [126I]iodomelatonin ([125I]Mel). Replacement of individual transmembrane domains (TMs) of the Mel1a receptor by the corresponding H9 helixes revealed that TM6 plays a critical role in ligand binding. Substitution of H9-TM6 into the Mel1a receptor abolished any detectable [125I]Mel binding, whereas the remaining TMs could be readily exchanged without affecting ligand binding. Subsequent site directed mutagenesis showed that glycine 20 in TM6 of the Mel1a receptor occupies an important position in the binding site. Thus, the mutation of glycine 20 to threonine, the corresponding H9 residue, severely reduced the receptor's affinity for melatonin. Furthermore, the double mutation of alanine 14 to cysteine and of glycine 20 to threonine in TM6 completely eliminated high affinity [125I]Mel binding. This strongly suggests that molecular modifications in TM6 that involve glycine 20 lead to steric incompatibilities in the binding pocket that prohibit high affinity melatonin binding. PMID- 10698202 TI - HE2beta and HE2gamma, new members of an epididymis-specific family of androgen regulated proteins in the human. AB - HE2 is an epididymis-specific sperm-binding secretory protein. We isolated a family of HE2-related complementary DNAs from a human caput/corpus library. The transcripts code for identical 71-amino acid N-termini and different C-termini, and 5'- and 3'-untranslated regions. Compared with the original HE2, HE2beta and HE2gamma proteins have a 25-amino acid deletion near the C-terminus, and HE2gamma isoforms have a second deletion. These frame-shifting deletions result in C termini differing in length, amino acid sequence, including number of cysteines, and isoelectric point. Identical sequences and deletion start and stop points indicate the HE2 isoforms are derived from alternative splicing of 8 or more exons of a single gene. Northern hybridization revealed that the 0.9-kb messenger RNA (mRNA) is most abundant in human caput; there is much less of it (20%) in corpus and little (<5%) in cauda. In castrated Macaca mulatta, HE2 mRNA decreased to 10% of sham-operated levels. Testosterone replacement maintained HE2 mRNA 3- to 5-fold higher than castrate levels, indicating its androgen dependence. Immunohistochemical staining revealed that the beta1 form is highly expressed in principal cells of the initial segment and caput. It is secreted into the lumen and binds to the sperm surface in the postacrosomal and neck regions. The beta2 form is expressed in principal cells primarily in efferent ducts. PMID- 10698203 TI - Effects of gamma-aminobutyric acid(A) receptor manipulation on migrating gonadotropin-releasing hormone neurons through the entire migratory route in vivo and in vitro. AB - GnRH neurons originate in the nasal compartment and migrate along vomeronasal fibers over the cribiform plate to the forebrain. Previously, we found gamma aminobutyric acid (GABA) present in GnRH neurons during development. To clarify the influence of GABA across the entire GnRH migration route, we examined the effects of muscimol and bicuculline (GABA(A) agonist and antagonist) in vivo and in vitro, maintaining the integrity of the nasal-forebrain connection. For in vivo experiments, mice were administered muscimol, bicuculline, or vehicle on days 10-15 of pregnancy and were killed on embryonic day 15 (E15). For in vitro experiments, 250-microm parasagittal slices of whole heads of E13 mice were incubated with muscimol, bicuculline, or vehicle for 2 days. Muscimol inhibited GnRH cell migration and decreased extension of GnRH fibers. Bicuculline treatment led to a disorganized distribution of GnRH cells in the forebrain and a concomitant dissociation of GnRH cells from fibers of guidance. These results suggest that GABA's influence on GnRH development changes as the cells move out of the nasal compartment and extend processes toward the median eminence. PMID- 10698204 TI - Activin A-induced HepG2 liver cell apoptosis: involvement of activin receptors and smad proteins. AB - A balance between cell proliferation and apoptosis is important for regulating normal liver function. Proteins of the transforming growth factor-beta superfamily are known to be important mediators of apoptosis in the liver. In this study we demonstrate that activin A potently induces apoptotic cell death in a hepatoma cell line, HepG2 cells. To determine the roles of activin receptors and downstream signaling proteins in activin A-induced apoptosis in these cells, the activin signaling pathway was analyzed using the transcription of an activin responsive reporter gene, p3TP-Lux, as an assay. Although individual activin receptors had little effect on transcriptional activity, coexpression of an activin type I receptor and a type II receptor significantly increased both basal and activin-induced transcriptional activation, with the combination ofreceptors IB and IIB being the most potent. Similarly, expression of individual Smad proteins had only a modest effect on reporter gene activity, but the combination of Smad2 and Smad4 strongly stimulated transcription. Activin signaling induced a rapid relocation of Smad2 to the nucleus, as determined using a green fluorescence protein-Smad2 fusion protein. In contrast, green fluorescence protein-Smad4 remained localized to the cytoplasm unless it was coexpressed with Smad2. In agreement with the transcriptional response assays, overexpression or suppression of activin signaling components in HepG2 cells altered apoptosis. Overexpression of receptors IB and IIB or Smad proteins 2 and 4 stimulated apoptosis, whereas dominant negative mutant forms of the activin type IIB receptor or Smad2 blocked activin-stimulated apoptosis. These studies suggest that signaling from the cell surface to the nucleus through Smad proteins is a required component of the activin A-induced cell death process in liver cells. PMID- 10698205 TI - Spermatogenic cells do not require estrogen receptor-alpha for development or function. AB - Estrogen receptors alpha (ERalpha) and beta (ERbeta) are ligand-dependent transcription factors and members of the nuclear hormone receptor superfamily encoded by separate genes. Male mice homozygous for a mutation in the gene encoding ERalpha are infertile. To determine whether germ cells or somatic cells require ERalpha, germ cells were transplanted from donor males homozygous for the mutation (ERalpha-/-) to testes of wild-type (ERalpha+/+) recipient mice depleted of germ cells. The recipients served as "surrogate fathers" for the infertile ERalpha-/- males. When mated to wild-type females, the recipients sired offspring heterozygous for the mutation (ERalpha+/-) and carrying the coat-color marker of the ERalpha-/- donor mice. These studies show that male germ cells do not require ERalpha for development or to function in fertilization, and imply that male ERalpha-/- mice are infertile due to disruption of estrogen action within somatic cells of the male reproductive system. PMID- 10698206 TI - Regulation of the Na,K-pump by leptin in 3T3-L1 fibroblasts. AB - Leptin, the product of the obesity (ob) gene, controls energy intake and expenditure primarily by actions on the central nervous system. However, recently it has become apparent that leptin also elicits a growing and diverse array of effects on peripheral tissues. The Na,K-pump is an electrogenic plasma membrane protein which actively extrudes 3Na+ ions and imports 2K+ ions per molecule of ATP hydrolysed. The pump is responsible for the maintenance of the electrochemical potential of all cells, which in turn drives all ion-coupled transport mechanisms. In this study we use 3T3-L1 fibroblasts to show that leptin inhibits Na,K-pump activity, as assessed by ouabain-sensitive 86Rb+ uptake. Inhibition of the Na,K-pump correlated with increased serine phosphorylation of the catalytic Na,K-pump alpha1 subunit. Upon investigation of leptin-stimulated signalling pathways using specific pharmacological inhibitors, only wortmannin prevented inhibition of the Na,K-pump by leptin. Moreover, leptin stimulated phosphotyrosine-associated PI 3-kinase activity in these cells. In summary, leptin was found to inhibit Na,K-pump activity, likely via PI 3-kinase. We propose that this effect may have wide ranging cardiovascular and metabolic implications and perhaps explain physiological effects of the hormone such as natriuresis. PMID- 10698207 TI - Nuclear receptor coactivators facilitate vitamin D receptor homodimer action on direct repeat hormone response elements. AB - Vitamin D receptor (VDR) is a ligand-dependent transcription factor that regulates target gene expression. Although VDR forms stable heterodimer complex with retinoid X receptors (RXRs) on vitamin D-response elements (VDREs), it is still not clear whether VDR/RXR heterodimers are the only VDR complexes responsible for vitamin D-mediated gene transcription. In this report, we analyzed the effect of nuclear receptor coactivators (SRC-1 and TRAM-1) on VDR homodimer and VDR/RXR heterodimer formation by electrophoretic mobility shift assay. We found that VDR forms stable homodimers after interaction with the coactivators on a VDRE (DR+3). Of particular note, DR+4 and DR+5 hormone-response elements (HREs) may also support such interactions. Cotransfection experiments revealed further that the coactivators enhance ligand-induced VDR transcription on these elements. Our studies suggest the important role of VDR homodimers, in addition to VDR/RXR heterodimers, in vitamin D-induced transactivation. Thus, specific coactivator-VDR interactions on HREs may determine target gene transactivation. PMID- 10698208 TI - Nonlymphoid intraepidermal mononuclear cell collections (pseudo-Pautrier abscesses): a morphologic and immunophenotypical characterization. AB - We evaluated the incidence, morphology, and immunophenotype of intraepidermal collections of mononuclear cells (ICMC) in a large number of inflammatory dermatosis and cutaneous lymphomas. ICMC appeared as small to large aggregates of cells, showing a morphology variable from monocytes to obvious dendritic cells, admixed with rare lymphocytes. ICMC were recognized in the epidermis or within hair follicle epithelium, and were either loosely or compactly arranged. ICMC were identified in 124 of 1,248 skin biopsies (9.9%) of inflammatory or lymphoid infiltrates, and were particularly frequent in spongiotic (43.4%) and in lichenoid dermatitis (10%), whereas they were rarely found in nonspecific superficial dermatitis (3.8%) and in psoriasis (4.7%). ICMC were also frequent in cutaneous T-cell lymphoma (13.3%), where they mimicked Pautrier abscesses. The ICMC forming cells showed a unique phenotype: the majority of them expressed CD1a and S-100, and lacked CD14, similar to mature Langerhans cells, but they were also strongly labeled by anti-CD11b, anti-CD36, and anti-CD68. Moreover, a subpopulation of them expressed CD83, an antigen that is usually absent on Langerhans cells. The occurrence of ICMC is a rather frequent, although hitherto poorly studied, phenomenon, occurring in several dermatosis, but particularly frequent in spongiosis-associated skin reactions. The cells within ICMC are represented by dendritic cells and dendritic cell precursors, whose phenotype indicates their derivation from circulating monocytes and differentiation into mature Langerhans cells. PMID- 10698210 TI - Expression of CD34 in sclerotic ("plywood") fibromas. AB - CD34 antigen is expressed in normal human skin on endothelium, in spindle cells located around adnexal structures, and in a subset of interstitial cells in the reticular dermis. CD34 expression has also been identified in a number of fibrohistiocytic neoplasms, such as dermatofibrosarcoma protuberans and solitary fibrous tumors of soft tissue. CD34 expression has not previously been described in sclerotic, or "plywood" fibromas. Here presented are three lesions from three patients, in which histologic examination revealed a well-circumscribed dermal nodule composed of spindled cells with focal nuclear pseudo-inclusions. There was extensive fibrosis with hypocellular, storiform areas, characteristic of sclerotic fibroma. The spindled cells strongly expressed CD34, but not factor XIIIa or markers of melanocytic, neural, or muscular differentiation. A diagnosis of Cowden syndrome was considered in one of the cases. These cases provide evidence that CD34 expression can occur in sclerotic fibromas, either solitary or associated with Cowden syndrome. When diagnosing a sclerotic fibroma, one should comment in the report regarding the possibility of Cowden syndrome. PMID- 10698209 TI - Comparative immunophenotypic study of lichen sclerosus: epidermotropic CD57+ lymphocytes are numerous--implications for pathogenesis. AB - To characterize the immunophenotype of inflammatory cells in lichen sclerosus (LS), we performed a comparative case control study using one- and two-color immunohistochemistry and the nitro blue tetrazolium (NBT) reaction. Study material consisted of 100 biopsies from patients with LS or from 12 control groups consisting of inflammatory, scarring, and depigmenting cutaneous disorders. In addition, fresh tissue was sampled from four vulvectomy specimens for NBT testing. The typical inflammatory infiltrate of LS contained numerous epidermotropic CD3+, CD8+, CD57+ cells, increased intraepidermal HLA-DR+ cells, and a dermal infiltrate rich in CD8+, CD57+, HLA-DR+, and CD68+ inflammatory cells. Comparing LS to the 12 control groups, epidermotropic CD57+ lymphocytes independently predicted LS (P = 0.006, logistic regression, multivariate analysis). Among the 12 control groups, only specimens of the inflammatory stage of morphea exhibited numerous dermal CD57+ lymphocytes. Two-color immunohistochemistry confirmed the CD3+/CD8+CD57+ and CD3+/ CD8+/CD57+HLA-DR+ epidermotropic and dermal lymphocytic phenotypes and the dermal macrophage CD68+HLA-DR+ phenotype. In LS, the NBT reaction revealed evidence of superoxide production associated with CD68+HLA-DR+ cells. Expansion of CD8+CD57+lymphocytes is associated with viral infections, autoimmune disease, malignancies, and transplantation and is suspected to be the result of chronic excessive antigen challenge. In these pathologic states, CD8+CD57+ lymphocytes (as terminally differentiated, antigen-specific T cells) participate in the suppression of cytolytic activity to limit tissue damage. In LS, activated macrophages and lymphocytes indicate persistent antigen-driven inflammation. LS's numerous CD8+CD57+ lymphocytes may be either the mediators or the consequence of its hallmark sclerosis. PMID- 10698211 TI - Solitary sclerotic fibroma of the skin: degenerated sclerotic change of inflammatory conditions, especially folliculitis. AB - Two cases showing changes of sclerotic fibroma developed in association with an inflammatory process, especially folliculitis. The lesion in the first case showed a well-circumscribed, nonencapsulated nodule in the dermis, which consisted of a perifollicular fibrotic area and a peripheral sclerotic area. In addition to the usual findings of sclerotic fibroma, spindle cells were heavily infiltrated in a storiform and fascicular pattern around the degenerated hair follicle, suggestive of dermatofibroma. The lesion in the second case showed the typical findings of sclerotic fibroma in association with folliculitis and hair follicle remnants. Our observations suggest that solitary sclerotic fibroma of the skin may be a degenerated or sclerotic end stage of other fibrous conditions, such as dermatofibroma, and that it may be induced by inflammation, especially folliculitis. PMID- 10698212 TI - Comparison of S-100 versus hematoxylin and eosin staining for evaluating dermal invasion and peripheral margins by desmoplastic malignant melanoma. AB - Desmoplastic malignant melanoma (DMM) is a rare and locally aggressive variant of malignant melanoma, which is difficult to diagnose clinically and microscopically. A retrospective study of 21 DMM were collected during 7 years, representing 1.7% of melanomas. By comparing S-100 and hematoxylin and eosin (H & E) staining in DMM, we sought to determine whether S-100 staining offered a more accurate means of assessing dermal and neural invasion, tumor thickness, and peripheral margins. Eleven cases were excluded because the tumor extended past the deep margin. Six cases that were stained with S-100 showed a greater tumor thickness than by H & E (difference of 0. 13 to 2.79 mm). In two cases, the tumor thickness was greater by using H & E than S-100, and there was no difference in the remaining two cases. Of the eight cases in which the peripheral margins were positive by S-100, neoplastic cells were only apparent at those margins in four cases when examined with H & E. The remaining two cases showed negative margins by both stains. Clinical follow-up was obtained from nine cases, and none had recurrence of melanoma. Particularly for hypocellular and amelanotic tumors, S 100 staining proved to be a valuable adjunct in determining the extent of the tumor at the peripheral margins. PMID- 10698213 TI - Folliculotropic T cells in regressive basal cell carcinoma of skin. AB - The histologic features of regression may be found in some basal cell carcinomas (BCCs), and it is known that T-cell infiltrates have a significant role in host defense against this tumor. We examined 945 hair follicles (HFs) adjacent to 150 regressing BCCs of skin for the presence of inflammatory infiltrates and compared the results against 315 HFs in 50 samples of normal skin. Focal T-cell infiltrates localized mainly to the upper portion of the HFs were found in 14.5% of the follicles adjacent to regressing BCCs. A statistically significant increase of inflammation in HFs was observed in BCCs with active regression compared with BCCs with inactive and mixed regression (P < 0.05). An increase in the number of HFs involved by T lymphocytes was also found in regressing BCCs compared to normal skin ( P < 0.00005). These data suggest that the damage to the follicles is concordant with active regression of BCCs. We speculate that the immune-mediated regression of BCCs is not only specifically directed to the cells of the tumor but may also induce activated lymphocytes with cytotoxic capability to cross react with the follicular epithelium. PMID- 10698214 TI - Cellulite: from standing fat herniation to hypodermal stretch marks. AB - There are glaring discrepancies in the microanatomical descriptions of cellulite in the literature. We revisited this common skin condition in women with a microscopic examination of 39 autopsy specimens. A control group consisted of 4 women and 11 men showing no evidence of cellulite. The lumpy aspect of the dermohypodermal interface appeared to represent a gender-linked characteristic of the thighs and buttocks without being a specific sign of cellulite. Incipient cellulite identified by the mattress phenomenon was related to the presence of focally enlarged fibrosclerotic strands partitioning the subcutis. Such strands possibly serve as a physiologic buttress against fat herniation limiting the outpouching of fat lobules on pinching the skin. These structures might represent a reactive process to sustained hypodermal pressure caused by fat accumulation. Full-blown cellulite likely represents subjugation of the hypertrophic response when connective tissue is overcome by progressive fat accumulation. Histologic aspects reminiscent of stretch marks are identified within the hypodermal strands, resulting in clinical skin dimpling. PMID- 10698215 TI - Of epithelium and nerve. PMID- 10698216 TI - Canine cutaneous and systemic histiocytosis: reactive histiocytosis of dermal dendritic cells. AB - Canine histiocytic proliferative disorders include reactive diseases such as cutaneous and systemic histiocytosis and neoplastic diseases such as cutaneous histiocytoma and localized and disseminated histiocytic sarcoma (malignant histiocytosis). Their etiology and pathogenesis are unknown. Canine cutaneous and systemic histiocytosis target the skin and subcutis and have similar clinical behavior. Systemic histiocytosis also affects other organ systems. Clinicopathologic and phenotypic features of canine cutaneous and systemic histiocytosis were examined in this study. Canine cutaneous (18 cases) and systemic (26 cases) histiocytosis were characterized by angiocentric, pleocellular accumulations consisting of CD1+, CD11c+, MHCII+, CD4+, and Thy-1+ (CD90) activated dermal dendritic antigen-presenting cells (APC) with admixed CD3+, CD8+, TCRalphabeta+ T lymphocytes, and neutrophils. Hence, canine cutaneous and systemic histiocytosis represent two clinical manifestations of a reactive proliferation of dermal dendritic cells. Cultures and special stains failed to identify infectious agents. Canine reactive histiocytoses respond to immunosuppressive therapy (cyclosporine A or leflunomide). Therefore, immune dysregulatory mechanisms are likely to be involved. Spontaneous reactive histiocytoses are frequently seen in dogs, and they constitute an excellent model to study pathologic mechanisms involved in reactive proliferations of dermal dendritic APC. PMID- 10698217 TI - Canine mast cell tumors express stem cell factor receptor. AB - c-kit protooncogene encodes a type III transmembrane receptor kinase, the stem cell factor receptor, or KIT. The ligand of the KIT. stem cell factor, is a cytokine that stimulates mast cell growth and differentiation. We have studied immunohistochemically KIT expression in 23 canine mast cell tumors (MCTs), 10 histiocytomas, 5 malignant melanomas, and in 2 cell lines derived from mast cells (HMC-1, human and C2, canine). As expected, KIT was detected both in the human mast cell leukemia cell line (HMC- ) and in the canine mastocytoma cell line C2. In normal canine skin, KIT expression was confined to mast cells. All canine MCTs expressed KIT, although the intensity of the staining reaction varied considerably among the 23 neoplasms. Grade III tumors showed the highest expression of KIT, whereas grade I tumors showed the lowest expression of KIT. Two patterns of KIT expression were detected in mast cells. In normal canine mast cells and in some neoplastic mast cells, KIT appeared mainly on the cell membrane. However, in many canine MCTs, KIT is accumulated in the cytoplasm, usually near the cell nucleus. The meaning of these two patterns is not clear. Expression of KIT could not be detected immunohistochemically in any of the other neoplasias investigated. According to our results, it can be concluded that most, if not all, canine MCT express KIT. Furthermore, there is an inverse correlation between the degree of differentiation and the expression of KIT. Moreover, according to our results, KIT can be used as a reliable immunohistochemical marker for canine mast cells and undifferentiated mast cell tumors. PMID- 10698218 TI - Phakomatous choristoma: a case report and review of the literature. AB - Phakomatous choristoma is a rare congenital lesion of the eyelid that can be clinically and/or histologically mistaken for a cyst, cutaneous adnexal neoplasm, or an ocular adnexal oncocytoma. Only 13 such cases have been previously described, mostly in the English language ophthalmic literature. Zimmerman reported the first case in 1971 and proposed the lesion to be of lenticular anlage origin, a theory that has been widely accepted. We report an additional case occurring in an 8-week-old male infant with a firm nodule of the right lower eyelid that was present since birth. A 15 x 12 x 2 mm circumscribed solid nodule with a homogenously white cut surface was surgically excised. Histologically, this lesion was comprised of cuboidal cells forming cystically dilated and irregularly branched ducts and cords within a densely fibrotic stroma. Also present were eosinophilic basement membranelike material, psammoma body-like calcifications and intraluminal degenerated ghost cells. The immunohistochemical profile of the epithelial cells included strong immunoreactivity for vimentin, focal weak staining for S-100, and negative staining for cytokeratin, epithelial membrane antigen, synaptophysin, and chromogranin. The irregularity of the ducts and cords of epithelial cells within the densely fibrotic stroma resembled an infiltrative neoplasm of cutaneous adnexal or lacrimal duct origin. However, the site of involvement, the peculiar basement membrane material, ghost cells, and immunohistochemical profile were features that helped to distinguish phakomatous choristoma from an infiltrative carcinoma. The correct identification of this lesion is essential to avoid an aggressive surgical excision, thus sparing the eyelid and lacrimal system. The purpose of this article is to bring attention to this rare entity, because it has not been described in either the dermatology or dermatopathology literature and furthermore, is not mentioned in any of the major dermatopathology texts. PMID- 10698219 TI - Clear cell syringoid carcinoma: an ultrastructural and immunohistochemical study. AB - Syringoid carcinoma (syringoid "eccrine" carcinoma or eccrine epithelioma) is a rare cutaneous tumor with some controversy regarding its correct definition. It may also be difficult to differentiate from its benign counterpart (syringoma), other adnexal carcinomas, and cutaneous metastasis from adenocarcinomas. We present a case of a syringoid carcinoma of the clear cell variant complemented with an immunohistochemical and ultrastructural study, the latter revealing cytoplasmic accumulation of glycogen and presence of intercellular and intracellular lumina in clear tumor cells, as well as diverse hallmarks of malignancy (i.e., perineural invasion, tumor necrosis, and deep invasion). Clear tumor cells showed cytoplasmic and membranous immunoreactivity to epithelial membrane antigen, carcinoembryonic antigen, keratins, and S-100. Our ultrastructural and immunohistochemical results support the ductal differentiation of the glycogen-filled clear cell tumor population. PMID- 10698220 TI - Intraepidermal Merkel cell carcinoma with no dermal involvement. AB - Cutaneous Merkel cell carcinoma (MCC) typically involves the dermis. Less than 10% of MCC have epidermal involvement. Only one MCC confined exclusively to the epidermis has been previously reported but was not recognized until the lesion recurred with typical MCC in the dermis. We present a case of a wholly intraepidermal pagetoid MCC without dermal involvement in a 74-year-old man with a 2.0-cm solitary verrucous papule on the left index finger. The initial biopsy and complete excision specimens showed marked epidermal hyperplasia, focal prominent squamous cell atypia, and MCC with florid pagetoid spread through the epidermis. There was no evidence of tumor within the dermis. The pagetoid MCC tumor cells showed diffuse cytoplasmic staining with antibodies to cytokeratin 20, and negative staining for chromogranin, neurofilament, S-100, vimentin, HMB45, leukocyte common antigen, and CD3. The cell of origin of MCC is still debated. The existence of an entirely intraepidermal variant of MCC would lend support to the view that MCC is a neoplastic expression of Merkel cells in at least some cases. Dermal-based MCC is a high-grade primary cutaneous neoplasm, but MCC confined exclusively to the epidermis may have a better prognosis. PMID- 10698221 TI - Benign cephalic histiocytosis progressing into juvenile xanthogranuloma: a non Langerhans cell histiocytosis transforming under the influence of a virus? AB - Benign cephalic histiocytosis (BCH) is best understood as a form of non Langerhans cell histiocytosis, specifically as an early mononuclear variant of juvenile xanthogranuloma (JXG). However, the progression of BCH into JXG in the same patient has only been reported once before. We describe the case of a 2-year old girl with asymptomatic, large, ill-defined infiltrated flat plaques over both cheeks, in addition to isolated papules. A punch biopsy of a plaque revealed dermal infiltration by vacuolated and scalloped histiocytes positive for CD68 KP 1, and that lacked expression of CD1a and S-100 protein, favoring macrophages over Langerhans cells. Electron microscopy study showed comma-shaped intracytoplasmic bodies in the histiocytic cells leading to the diagnosis of BCH. One year later, after an episode of varicella-zoster infection, the flat plaques over the cheeks became large reddish-yellow nodules, and in a second biopsy appeared to progress to JXG. Virus-related mechanisms of progression are discussed. PMID- 10698222 TI - Tumorlike eosinophilic granuloma of the skin. AB - In 1952, Kuske reported on a patient with a peculiar tumor on the dorsum of the right hand; histological analysis revealed a dense dermal infiltrate with numerous eosinophils. Not aware of any similar case report in the literature, he coined the descriptive term "tumor-like eosinophilic granuloma of the skin." In 1995, a 55-year-old white man with cancer of the prostate presented with a 4 month history of two reddish-brown, solid skin tumors on his left forearm and on the right side of his abdomen, respectively. Histologic examination revealed a dense, superficial and deep, tumorlike dermal inflammatory infiltrate consisting mainly of eosinophils as well as neutrophils and in part epithelioid, in part foamy histiocytes. Flame figures were absent. Immunohistochemical analysis was negative for S-100 protein, whereas sporadic cells in the infiltrate were CD1a positive and many mononuclear-histiocytic cells reacted with MAC 387. Stains as well as cultures for bacteria, mycobacteria, and fungi were negative. The descriptive diagnosis of tumorlike eosinophilic granuloma of the skin was made. Seven weeks after prostatectomy, both tumors resolved spontaneously and so far has not recurred. In our opinion, this is the second report of Kuske's tumorlike eosinophilic granuloma of the skin. Perhaps tumorlike eosinophilic granuloma of the skin, eosinophilic ulcer of the mucosa, and transient eosinophilic nodulomatosis should be considered a mucocutaneous reaction pattern as is seen in cats. In humans, hypersensitivity reactions or atopy could emerge as an etiological link. PMID- 10698223 TI - Presternal bronchogenic sinus with predunculated lymphoid aggregate. AB - Congenital cranial, cervical, and upper thoracic sinuses are rare conditions that historically have been classified according to their location and/or pathology. However, published reports of bronchogenic or branchial anomalies are on the increase, and the traditional defining characteristics--location and histopathology--are proving to be less reliable. We describe the pathologic and clinical findings of a congenital presternal pedunculated lesion with a sinus, and review the literature to describe its proper classification. PMID- 10698224 TI - Solitary fibrous tumor of the vagina. AB - We report of a solitary fibrous tumor (SFT) of the vagina and discuss the differential diagnosis. This is the first SFT documented, to our knowledge. SFTs should be included in the differential diagnosis of fibroblastic, myofibroblastic, and neural lesions of the skin, subcutaneous tissue, and mucosa and can be distinguished from other spindle cell neoplasms at those sites. PMID- 10698225 TI - Puzzling follicular germinative proliferations. PMID- 10698226 TI - Histopathologic features of progression in Mediterranean and immunodeficiency related Kaposi sarcoma. PMID- 10698227 TI - Syringocystadenoma papilliferum mimicking breast carcinoma. PMID- 10698228 TI - Predictors of improvement in left ventricular ejection fraction with carvedilol for congestive heart failure. AB - BACKGROUND: Beta-blocker therapy has been reported to improve survival and left ventricular ejection fraction (LVEF) in the setting of congestive heart failure (CHF). The magnitude and predictors of improved LVEF are unclear. METHODS: A total of 295 patients were enrolled in the study. Inclusion criteria were LVEF <35% at baseline and symptomatic (New York Heart Association class II to IV) CHF despite treatment with at minimum an angiotensin-converting enzyme inhibitor. Carvedilol was initiated at 3.125 mg twice daily and titrated to a target dose of 25 or 50 mg twice daily, depending on the patient's weight. Paired pretreatment baseline and 9 months with treatment follow-up quantitative LVEFs (assessed by resting radionuclide ventriculograms) were obtained in 161 (55 %) of the patients. RESULTS: LVEF improved from 25% +/- 6% at baseline to 36%+/-12% at follow-up (P<.001). Mean change in LVEF (deltaLVEF) was greater for nonischemic cardiomyopathy (NICM) (+14.5+/-2 LVEF points) than ischemic cardiomyopathy (deltaLVEF +/- 7.6+/-10 EF points, P = .001). The deltaLVEF was > or =21 LVEF points in 30% of the NICM group versus 10% of the ischemic cardiomyopathy group. Conversely, the deltaLVEF was unchanged to minimally improved (< or =5 LVEF points) in 21% of the NICM group versus 52% of the ischemic cardiomyopathy group. Multivariable analysis identified NICM and recent onset of congestive heart failure as correlates of improved LVEF. CONCLUSIONS: Carvedilol significantly improved LVEF, especially in patients with NICM and those with recent onset of CHF. PMID- 10698229 TI - Quantitative thallium-201 and technetium 99m sestamibi tomography at rest in detection of myocardial viability in patients with chronic ischemic left ventricular dysfunction. AB - BACKGROUND: This study was designed to determine the most effective quantitative threshold for thallium-201 and technetium 99m sestamibi uptake on tomographic imaging after rest injection for the detection of myocardial viability in patients with chronic myocardial infarction. METHODS AND RESULTS: Thallium and sestamibi cardiac tomography at rest was performed in 43 patients with chronic myocardial infarction and impaired left ventricular (LV) function undergoing coronary revascularization. In all patients, echocardiography and radionuclide angiography were performed at baseline and repeated 12 months later to evaluate recovery of regional LV function and LV ejection fraction, respectively. Optimal threshold cutoff points to separate reversible from irreversible dysfunction were determined by receiver operating characteristic analysis. When all dysfunctional segments were considered, the best cutoff point in the identification of reversible LV dysfunction for both thallium and sestamibi activity was 67%. When only akinetic or dyskinetic segments were considered, the best cutoff point in the identification of reversible LV dysfunction was 58% for thallium and 55% for sestamibi. In these segments, the area under the receiving operating characteristic curves constructed for thallium and sestamibi activity were 0.74+/ 0.05 and 0.75+/-0.04, respectively (P = not significant). LV ejection fraction was 33%+/-7% at baseline and increased to 37%+/-7% after revascularization (P<.0001). A significant relation between the number of akinetic or dyskinetic but viable myocardial segments and revascularization-induced changes in LV ejection fraction was observed for both thallium (r = 0.60, P<.0001) and sestamibi (r = 0.64, P<.0001) imaging. CONCLUSIONS: In patients with chronic myocardial infarction, quantitative analysis of thallium and sestamibi activity on tomographic imaging at rest predicts recovery of regional and global LV dysfunction after revascularization procedures. The most effective quantitative threshold for detecting reversible LV dysfunction is comparable for thallium and sestamibi tomographic imaging. However, the optimal cutoff point is different for both tracers when all dysfunctional segments are considered or when the analysis is focused only on segments with more severe functional impairment (i.e., akinetic or dyskinetic segments). PMID- 10698230 TI - Relation between thallium-201/iodine 123-BMIPP subtraction and fluorine 18 deoxyglucose polar maps in patients with hypertrophic cardiomyopathy. AB - BACKGROUND: Clinical studies have shown discrepancies in the distribution of thallium-201 and iodine 123-beta-methyl-iodophenylpentadecanoic acid (BMIPP) in patients with hypertrophic cardiomyopathy (HCM). Myocardial uptake of fluorine 18 deoxyglucose (FDG) is increased in the hypertrophic area in HCM. METHODS AND RESULTS: We examined whether the distribution of a Tl-201/BMIPP subtraction polar map correlates with that of an FDG polar map. We normalized to maximum count each Tl-201 and BMIPP bull's-eye polar map of 6 volunteers and obtained a standard Tl 201/BMIPP subtraction polar map by subtracting a normalized BMIPP bull's-eye polar map from a normalized Tl-201 bull's-eye polar map. The Tl-201/BMIPP subtraction polar map was then applied to 8 patients with HCM (mean age 65+/-12 years) to evaluate the discrepancy between Tl-201 and BMIPP distribution. We compared the Tl-201/BMIPP subtraction polar map with an FDG polar map. In patients with HCM, the Tl-201/BMIPP subtraction polar map showed a focal uptake pattern in the hypertrophic area similar to that of the FDG polar map. By quantitative analysis, the severity score of the Tl-201/BMIPP subtraction polar map was significantly correlated with the percent dose uptake of the FDG polar map. CONCLUSION: These results suggest that this new quantitative method may be an alternative to FDG positron emission tomography for the routine evaluation of HCM. PMID- 10698231 TI - Angiotensin-converting enzyme inhibitor therapy affects myocardial fatty acid metabolism after acute myocardial infarction. AB - BACKGROUND: Angiotensin-converting enzyme (ACE) inhibitor therapy has an early mortality benefit in unselected patients with acute myocardial infarction (AMI). However, the effects of ACE inhibition on myocardial fatty acid metabolism in this patient population have not been studied. We tested the hypothesis that ACE inhibitor therapy improves myocardial fatty acid metabolism and decreases mortality rate in patients after AMI. METHODS: Forty-two patients after first anterior AMI and primary angioplasty were randomly assigned to titrated oral enalapril (n = 24) or placebo therapy (n = 18). Iodine 123-labeled 15-(p iodophenyl)-3-(R,S)-methylpentadecanoic acid (BMIPP) single photon emission computed tomography imaging was performed an average of 4.8 days after AMI and 1 month after AMI. BMIPP abnormalities were quantified as a severity index by a polar map. RESULTS: There were no significant changes in baseline characteristics, cardiac function, and angiographic findings between patients in the enalapril group and patients in the placebo group. However, BMIPP severity index from acute phase to chronic phase was significantly decreased in the enalapril-treated group (118+/-48 to 82+/-36, P<.05), but not in the placebo group (123+/-65 to 115+/-58, P not significant). CONCLUSION: ACE inhibition therapy improved myocardial fatty acid metabolism and regional left ventricular function in patients after anterior AMI. BMIPP single photon emission computed tomography findings imply that this better outcome may be attributable to an improvement of cellular function with ACE inhibitors. PMID- 10698232 TI - Assessment of perfusion, function, and myocardial metabolism after infarction with a combination of low-dose dobutamine tetrofosmin gated SPECT perfusion scintigraphy and BMIPP SPECT imaging. AB - BACKGROUND: Tetrofosmin gated single photon emission computed tomography (SPECT) allows simultaneous assessment of regional myocardial perfusion, global and regional left ventricular function, and function at rest and during pharmacologic intervention. SPECT with fatty acid analogues, such as beta-methyl-iodophenyl pentadecanoic acid (BMIPP), can be used to monitor metabolic changes induced by myocardial ischemia. In this work, the results of both studies obtained in patients with recent myocardial infarction are integrated. METHODS: Twenty patients underwent tetrofosmin and BMIPP scintigraphy with a 3-head camera. Two consecutive tetrofosmin gated SPECT acquisitions were performed 60 minutes after administration of technetium-99m tetrofosmin (925 MBq) at rest (3x20 stops of 9 s; matrix 64x64 over 360 degrees . One acquisition was made at rest, and the second was made during dobutamine infusion (10 microg/kg/min). Regional functional abnormalities were quantified and expressed as wall thickening severity (WTsev) in arbitrary units. Left ventricular ejection fraction and volumes were assessed with the Cedars Sinai algorithm. BMIPP imaging started 20 minutes after iodine 123-BMIPP (150 MBq) administration at rest (3x32 stops of 60 s; matrix 64x64 over 360 degrees; medium energy collimators). Tracer uptake was scored according to a 25-segment model. RESULTS: Sixteen of 18 patients had regional functional abnormalities at baseline (average WTsev 13.7 units). The WTsev score at baseline correlated well with the degree of residual perfusion. During dobutamine infusion, WTsev did not change (from 23.4 to 23.6 units) in 5 patients; it decreased (from 16.1 to 5.9 units) in 11 patients; and it increased (from 13.0 to 22.3 units) in 3 patients. An increase or decrease in WTsev during dobutamine infusion was associated with the presence of a considerable amount of BMIPP mismatched myocardium, whereas no change in WTsev was preferentially associated with a BMIPP matched pattern and perfusion defects with a higher severity score. CONCLUSION: Immediately after infarction, the severity of regional dysfunction at rest correlated well with the perfusion defect severity. Improvement in regional function during dobutamine administration is associated with less severe perfusion defects and a considerable amount of BMIPP mismatched myocardium, both suggesting viability. PMID- 10698233 TI - Evaluation of three rhenium-188 candidates for intravascular radiation therapy with liquid-filled balloons to prevent restenosis. AB - BACKGROUND: Intravascular brachytherapy is an effective method for inhibiting coronary restenosis after percutaneous transluminal coronary angioplasty. A new concept for preventing restenosis is the use of a liquid-filled balloon containing a beta-ray-emitting radioisotope. Generator-produced rhenium-188 (Re 188) is a good candidate for intravascular brachytherapy. However, in the unlikely event of balloon rupture, release of Re-188 perrhenate may cause a high radiation dose to the thyroid and stomach. In this study, we compared the biodistributions of three Re-188 preparations (Re-188 perrhenate, Re-188 pentetic acid [DTPA], and Re-188 MAG3) to assess the radiation dose to organs in a rat model that mimicked balloon rupture. METHODS AND RESULTS: After injection of Re 188 preparations intravenously, rats were killed at 10 minutes, 30 minutes, 60 minutes, 2 hours, and 6 hours (n = 5/group). Tissue concentrations were calculated and expressed as percent injected dose per gram or per milliliter. In addition, urine excretion and thyroid gland uptake were evaluated in rats (n = 5/group) with a gamma camera after administration of 37 MBq (1 mCi) of each agent. Our data showed all 3 agents were excreted primarily via urine. In the Re 188 MAG3 group, 82% was excreted within 1 hour, but in the Re-188 perrhenate group, only 28% was excreted. The biodistribution data for these agents revealed that radioactivity levels in the stomach and the thyroid gland were high in the perrhenate group but low in the Re-188 DTPA and Re-188 MAG3 groups. The concentration levels in other tissues including lung, liver, testis, muscle, and blood were low throughout this study for all 3 agents. The thyroid radiation values were 0.163, 0.0167, and 0.00728 mGy/MBq for Re-188 perrhenate, Re-188 DTPA, and Re-188 MAG3, respectively. The stomach radiation values were 0.127 mGy/MBq for Re-188 perrhenate, 0.013 mGy/MBq for Re-188 DTPA, and 0.0104 mGy/MBq for Re-188 MAG3. CONCLUSIONS: In the event of balloon rupture, the release of Re 188 MAG3 or Re-188 DTPA results in lower radiation doses than release of Re-188 perrhenate, especially to the thyroid gland and the stomach. PMID- 10698234 TI - Severe regional ischemia alters coronary flow reserve in the remote perfusion area. AB - BACKGROUND: Clinical and experimental studies suggest that coronary flow reserve (CFR) may be abnormal in regions remote from myocardial infarction. We sought to determine the possible relation among stenosis severity, ischemic dysfunction, and impairment of CFR in remote regions. METHODS AND RESULTS: In 7 open-chest dogs, acute graded left circumflex (LCX) ischemia was created and maintained based on measurement of the transstenotic (aortic-distal LCX) pressure gradient (measured in millimeters of mercury). Regional thickening was assessed with sonomicrometers. Regional myocardial flow was assessed at rest with radiolabeled microspheres. Doppler flow probes were placed on proximal LCX and left anterior descending (LAD) arteries to measure resting flow and CFR in response to intracoronary injection of adenosine (36 microg). These parameters were assessed under baseline conditions and during transstenotic gradients of 10, 20, 30, and 40 mm Hg. Increasing LCX stenosis severity caused progressive impairment of LCX CFR: baseline (2.22+/-0.10), stenosis 10 (1.80+/-0.06), stenosis 20 (1.56+/ 0.08), stenosis 30 (1.30+/-0.04), and stenosis 40 (1.17+/-0.06) (P<.01 vs. baseline). Remote LAD CFR was not altered by mild to moderate LCX stenosis (baseline [2.33+/-0.19]; stenosis 10 [2.30+/-0.25]; stenosis 20 [2.15+/-0.26]). However, critical LCX stenosis producing mild to moderate reduction in thickening in the ischemic region was associated with a significant impairment of LAD CFR: stenosis 30 (1.90+/-0.26) and stenosis 40 (1.80+/-0.22) (P<.01 vs. baseline). These changes in remote CFR persisted after correction for changes in the rate pressure product. CONCLUSION: In an acute canine model of progressive LCX coronary stenosis, CFR was impaired in both ischemic and remote nonischemic regions in association with mild to moderate ischemic-induced regional myocardial dysfunction. Thus pharmacologic vasodilation provoked only mild heterogeneity in CFR in the presence of a critical LCX stenosis as a result of concurrent reduction of LAD CFR. This phenomenon warrants further clinical and experimental investigation because it may affect detection of flow heterogeneity during acute ischemia (which induced myocardial dysfunction). PMID- 10698235 TI - Cardiotoxicity of doxorubicin and other anthracycline derivatives. PMID- 10698236 TI - Cardiac signal transduction. PMID- 10698237 TI - Use of myocardial perfusion imaging to assess viability. PMID- 10698238 TI - The role of cardiac imaging in optimizing therapy in heart failure. PMID- 10698239 TI - Acute isolated right ventricular infarction: rest technetium 99m tetrofosmin myocardial perfusion imaging in a patient with chest pain. PMID- 10698240 TI - Missed opportunities to assess sexually transmitted diseases in U.S. adults during routine medical checkups. AB - OBJECTIVES: Although sexually transmitted diseases (STDs) cause tremendous health and economic burdens in our society, awareness and knowledge regarding STDs remain poor among health care providers. To examine missed opportunities for STD related counseling, diagnosis and treatment, we investigated how frequently U.S. adults reported being asked about STDs by their health care providers during routine checkups. METHODS: We analyzed the responses of 3390 adults aged 18-64 who reported having a routine checkup during the past year in the 1994 U.S. National Health Interview Survey (NHIS), a nationally representative survey. We used a logistic model to determine factors that were independently associated with the likelihood of being asked about STDs during the checkup. RESULTS: Only 28% (+/-0.9%) of respondents reported being asked about STDs during their last routine checkup. Persons were significantly more likely (p<0.05) to be asked about STDs if they were aged under 45, male, single, had a household income under the federal poverty level, or were insured by a health maintenance organization, public coverage or by no plan rather than by a fee-for-service arrangement. CONCLUSIONS: Only about one quarter of U.S. adults reported being asked about STDs during routine checkups. Routine checkups in which these issues are not discussed may represent missed opportunities for STD prevention. Persons presenting for routine care can be counseled, screened and, if infected, can be treated. Interventions are needed at the patient, provider, and community levels to increase the opportunities to assess STD risk, to counsel, to diagnose, and to treat infections during routine checkups. PMID- 10698241 TI - Selective screening for chlamydial infection: which criteria to use? AB - BACKGROUND: Screening sexually active women for Chlamydia trachomatis is necessary to detect asymptomatic infections. Selective screening is a common strategy because universal screening is too costly in many settings. In order to guide local programs in the choice of selective screening criteria, we examined the performance of previously proposed screening criteria for C. trachomatis. METHODS: A clinic-based, cross-sectional study was conducted in public family planning and sexually transmitted disease (STD) clinics in ten counties in North Carolina. Women (n = 4471 in family planning and n = 2201 in STD clinics) undergoing pelvic examination were enrolled consecutively. Nine sets of screening criteria, including age alone, were compared using sensitivity, specificity, number of tests required and receiver-operator characteristic (ROC) analysis. All women underwent testing with ligase chain reaction assay of cervical specimens to identify C trachomatis infection. RESULTS: The prevalence of C. trachomatis was 7.8% and 11.0% in family planning and STD clinics, respectively. The sensitivities of published criteria ranged from 0.50 to 0.97. Specificities ranged from 0.05 to 0.66. In family planning clinics, the best performing criteria would detect 84% of infections while screening 51% of women. In STD clinics, the same criteria would detect 83% of infections but require testing 67% of women. Testing women aged < or =22 would detect 77% of infections in family planning and 74% of infections in STD clinics, while testing 51% and 48% of the women, respectively. CONCLUSIONS: When site-specific criteria cannot be developed, age alone is an acceptable strategy for selective screening for chlamydial infection. PMID- 10698242 TI - Increasing influenza and pneumococcal immunization rates: a randomized controlled study of a senior center-based intervention. AB - BACKGROUND: Immunizations decrease morbidity from influenza and pneumococcal infections. Immunization levels remain below desired levels despite clinic-based and public education efforts. This paper describes a randomized, controlled trial of a senior center-based program, which used peer-to-peer outreach to increase pneumococcal and influenza immunization rates among an urban senior population. METHODS: Seniors were randomized to intervention or control groups. The intervention group received educational brochures mailed with reply cards to report immunization status, telephone calls from senior volunteers to unimmunized participants, and computerized immunization tracking. Immunization rates were obtained before and after the intervention by self-report. RESULTS: Among participants without prior pneumococcal immunization, the pneumococcal immunization rate among the intervention group (52.0%; 95% CI = 46.6%-57.4%) was significantly higher than that of the control group (30.9%; 95% CI = 26.6%-35.2%) (rate ratio = 1.68; 95% CI = 1.40-2.03). Among those without influenza immunization in the prior year, significantly more (50.0%; 95% CI = 40.0%-60.0%) were immunized against influenza in the intervention group than in the control group (23.0%; 95% CI = 15.2%-33.3%) (rate ratio = 2.17; 95% CI = 1.42-3.31). Among those with influenza immunization in the prior year, the rate ratio was 1.04 (95% CI = 1.01-1.07). CONCLUSIONS: The intervention increased both influenza and pneumococcal immunization rates to high levels, suggesting that further progress in increasing adult immunization coverage is possible. PMID- 10698243 TI - The impact of computer-generated messages on childhood immunization coverage. AB - INTRODUCTION: Recent evaluations of computer-generated reminder/recall messages have suggested that they are an inexpensive, labor-saving method of improving office visitation rates of childhood immunization providers. This study assesses the sustained impact of computer-generated messages on immunization coverage during the first two years of life. DESIGN: Randomized, controlled trial. SETTING: County health department in the Denver metropolitan area. STUDY PARTICIPANTS: Children (n = 1227) 60 to 90 days of age who had received the first dose of diphtheria-tetanus-pertussis (DTP) and/or poliovirus vaccines. INTERVENTION: Households of children were randomized into four groups to receive: telephone messages followed by letters (Group A); telephone messages alone (Group B); letters only (Group C); or no notification (Group D). Households in the intervention groups (A, B, and C) received up to five computer-generated telephone messages and/or up to four letters each time their children became due for immunization(s). MAIN OUTCOME MEASURE: Immunization series completion at 24 months of age. RESULTS: Children whose families were randomized to receive any of the interventions were 21% more likely to have completed the immunization series by 24 months of age than were children randomized into the control group (49.2% vs 40.9%; RR [rate ratio] = .21; CI [confidence interval] = 1.01, 1.44). While not statistically significant, children in Group A were 23% more likely to complete their immunization series by 24 months of age than those in the control group (50.2% vs 40.9%; RR = 1.23; CI = 1.00, 1.52). No differences were detected among the intervention groups. The costs per additional child completing the series by 24 months of age in Group A was $226 ($79 after start-up costs were discounted). CONCLUSION: Computer-generated contacts, either by phone or by mail (or both combined), used each time vaccines become due, are efficacious in increasing immunization coverage of children under 2 years of age. PMID- 10698244 TI - Impact of the sequential poliovirus immunization schedule: a demonstration project. AB - OBJECTIVE: Researchers for this project evaluated compliance with the sequential poliovirus immunization schedule that uses inactivated poliovirus vaccine (IPV) for the first 2 doses of the polio immunization series, and assessed immunization coverage rates before and after implementation of this schedule at 6 public health clinics serving 1 county in Georgia. DESIGN: Immunization histories for 3 birth cohorts of infants were compared: (1) the baseline cohort, born January 1 through June 30, 1995; (2) the evaluation cohort, born January 1 through June 30, 1997, after implementation of the schedule change; and (3) the dose-3 cohort, born August 1 through November 30, 1996 (i.e., old enough to be eligible for a third dose of poliovirus vaccine following implementation of the sequential schedule). RESULTS: Following implementation of the new poliovirus immunization recommendations, 94% (534 of 567) of infants who received their first dose of poliovirus vaccine by age 3 months received IPV. Among these infants, 99.6% (532 of 534) were also up to date (UTD) for first doses of diphtheria and tetanus toxoids and acellular pertussis vaccine (DTP1/DTaP1), 99.6% (532 of 534) were UTD for first doses of hemophilus influenza type b (Hib 1), and 98.6% (527 of 534) had received at least one dose of Hepatitis B. Among infants visiting the clinics for their first or second dose of poliovirus vaccine, DTaP/DTP, and/or Hib, 76% received 3 or 4 simultaneous injections. In the dose-3 cohort, 78% (145 of 185) of infants who received a third dose of poliovirus vaccine had received 2 doses of IPV and 1 dose of oral poliovirus vaccine. CONCLUSIONS: Compliance with the recommended use of IPV for the first 2 poliovirus immunization doses as part of the sequential schedule was very high in this low-income and ethnically diverse population. Furthermore, the need for additional injections did not impede the delivery of recommended childhood immunizations. PMID- 10698245 TI - Occupational needlestick injuries in a metropolitan police force. AB - OBJECTIVES: Police officers are at risk of bloodborne diseases through needlestick injuries but few studies have addressed this problem. The purpose of this study was to assess the risk of needlestick injuries in law enforcement officers and to determine predictors of injuries and reporting rates. DESIGN: An anonymous, voluntary questionnaire was distributed to 1738 active-duty, metropolitan police officers. The survey included the number of needlestick injuries ever experienced, how often these were reported, activities at the time of injury and attitudes toward injuries. RESULTS: Of the 803 respondents (46.2% of survey population), 29.7% had at least one needlestick injury, and 27.7% of this group had two or more. Risk factors included evening shifts, pat-down searches, patrol duties, male gender and less experience. Only 39.2% sought medical attention for these injuries. CONCLUSIONS: Needlestick injuries occur with considerable frequency in this group of law enforcement personnel, suggesting an increased risk of becoming infected with bloodborne pathogens, including hepatitis B, hepatitis C and HIV. PMID- 10698246 TI - Self-reported childhood sexual and physical abuse and adult HIV-risk behaviors and heavy drinking. AB - CONTEXT: Although studies of clinical samples have identified links between childhood abuse, especially sexual abuse, and adult health-risk behaviors, the generalizability of these findings to the population and the relative importance of different types of abuse in men and women are not known. OBJECTIVE: To estimate the risk of self-reported adult HIV-risk behaviors and heavy drinking that is associated with self-reported childhood histories of physical and/or sexual abuse for men and women in a general-population sample, after controlling for age and education. A second objective is to determine whether, among women, early and chronic sexual abuse is associated with heightened risk compared to later or less extensive abuse. DESIGN: A population-based telephone survey, the 1997 Washington State Behavioral Risk Factor Surveillance System (BRFSS), asked a representative sample of adults whether they had ever been physically or sexually abused in childhood, and if so, the age at first occurrence and number of occurrences. The survey also asked about levels of alcohol use and, for those under 50 years, about HIV-risk behaviors. PARTICIPANTS: Three thousand four hundred seventy-three English-speaking non-institutionalized civilian adults in Washington State. MAIN OUTCOME MEASURES: Self-reported HIV-risk behaviors in the past year and heavy drinking in the past month. RESULTS: We identified associations between reported abuse history and each health-risk behavior that we examined. For women, early and chronic sexual abuse (occurring without nonsexual physical abuse) was associated with more than a 7-fold increase in HIV-risk behaviors (odds ratio [OR], 7.4; 95% confidence intervals [CI] 2.4 to 23.5); and any sexual abuse, combined with physical abuse, was associated with a 5-fold increase in these risk behaviors (OR, 5.0; 95% CI, 2.2 to 11.5). For women, only combined sexual and physical abuse was associated with heavy drinking (OR, 6.2; 95% CI, 2.2 to 16.9). Physical abuse alone was not associated with either health risk behavior for women. For men, any sexual abuse was associated with an 8-fold increase in HIV-risk behaviors (OR, 7.9; 95% CI, 1.8 to 35.1). Physical abuse alone was associated with a 3-fold increase in risk of HIV-risk behaviors (OR, 3.2; 95% CI, 1.3 to 7.9) and a similar increase in risk of heavy drinking (OR, 3.2; 95% CI, 1.8 to 5.5). Although only 29% of the women and 19% of the men who were asked about HIV-risk behaviors reported any history of childhood abuse, these accounted for 51% and 50% of those reporting HIV-risk behaviors, respectively. For heavy drinking the corresponding figures were 25% of the women and 23% of the men reporting any abuse, who accounted for 45% and 33% of those reporting heavy drinking, respectively. CONCLUSIONS: Efforts to prevent or remediate adult health-risk behaviors should consider the possibility of a history of childhood abuse, as one third to one half of those reporting HIV-risk behaviors or heavy drinking in a general-population survey also reported childhood abuse. PMID- 10698247 TI - Preventive dentistry: practitioners' recommendations for low-risk patients compared with scientific evidence and practice guidelines. AB - INTRODUCTION: The purpose of this article is to compare published evidence supporting procedures to prevent dental caries and periodontal disease, in low risk patients, with the actual preventive recommendations of practicing dentists. METHODS: Methods included (1) a survey questionnaire of general dentists practicing in western New York State concerning the preventive procedures they would recommend and at what intervals for low-risk children, young adults, and older adults; and (2) review of the published, English-language literature for evidence supporting preventive dental interventions. RESULTS: The majority of dentists surveyed recommended semiannual visits for visual examination and probing to detect caries (73% to 79%), and scaling and polishing to prevent periodontal disease (83% to 86%) for low-risk patients of all ages. Bite-wing radiographs were recommended for all age groups at annual or semiannual intervals. In-office fluoride applications were recommended for low-risk children at intervals of 6 to 12 months by 73% of dentists but were recommended for low risk older persons by only 22% of dentists. Application of sealants to prevent pit and fissure caries was recommended for low-risk children by 22% of dentists. Literature review found no studies comparing different frequencies of dental examinations and bite-wing radiographs to determine the optimal screening interval in low-risk patients. Two studies of the effect of scaling and polishing on the prevention of periodontal disease found no benefit from more frequent than annual treatments. Although fluoride is clearly a major reason for the decline in the prevalence of dental caries, there are no studies of the incremental benefit of in-office fluoride treatments for low-risk patients exposed to fluoridated water and using fluoridated toothpaste. CONCLUSIONS: Comparative studies using outcome end points are needed to determine the optimal frequency of dental examinations and bite-wing radiographs for the early detection of caries, and of scaling and polishing to prevent periodontal disease in low-risk persons. There is no scientific evidence that dental examinations, including scaling and polishing, at 6 month intervals, as recommended by the dentists surveyed in this study, is superior to annual or less frequent examinations for low-risk populations. There is also no evidence that in-office fluoride applications offer incremental benefit over less costly methods of delivering fluoride for low-risk populations. PMID- 10698248 TI - The Fort McMurray Demonstration Project in Social Marketing: theory, design, and evaluation. AB - CONTEXT: The Fort McMurray Demonstration Project in Social Marketing is a multifaceted program that applies the techniques of social marketing to health and safety. This paper describes the origins of the project and the principles on which it was based. VENUE: Fort McMurray, in the province of Alberta, Canada, was selected because the community had several community initiatives already underway and the project had the opportunity to demonstrate "value added." CONCEPT: The project is distinguished from others by a model that attempts to achieve mutually reinforcing effects from social marketing in the community as a whole and from workplace safety promotion in particular. DESIGN: Specific interventions sponsored by the project include a media campaign on cable television, public activities in local schools, a community safety audit, and media appearance by a mascot that provides visual identity to the project, a dinosaur named "Safetysaurus." The project integrated its activities with other community initiatives. MAIN OUTCOME MEASURES: The evaluation component emphasizes outcome measures. A final evaluation based on injury rates and attitudinal surveys is underway. RESULTS: Baseline data from the first round of surveys have been compiled and published. In 1995, Fort McMurray became the first city in North America to be given membership in the World Health Organization's Safe Community Network. PMID- 10698249 TI - The Armed Forces Epidemiological Board: its impact on communicable diseases with a special tribute to Abram S. Benenson, MD. PMID- 10698250 TI - American College of Preventive Medicine public policy on needle-exchange programs to reduce drug-associated morbidity and mortality. AB - OVERVIEW: Based on a review of the current literature and recommendations, the American College of Preventive Medicine presents a public policy statement on needle-exchange programs. PMID- 10698251 TI - Notes from the Association of Teachers of Preventive Medicine: vaccine risk/benefit communication project. PMID- 10698252 TI - Bioterrorism and the public health response. PMID- 10698253 TI - It's a small world after all. PMID- 10698254 TI - STD risk assessment and chlamydia screening: what's missing? PMID- 10698255 TI - Highlights of National Conference on Violence and Reproductive Health. PMID- 10698256 TI - Nucleocytoplasmic protein transport and recycling of Ran. AB - The active transport of proteins into and out of the nucleus is mediated by specific signals, the nuclear localization signal (NLS) and nuclear export signal (NES), respectively. The best characterized NLS is that of the SV40 large T antigen, which contains a cluster of basic amino acids. The NESs were first identified in the protein kinase inhibitor (PKI) and HIV Rev protein, which are rich in leucine residues. The SV40 T-NLS containing transport substrates are carried into the nucleus by an importin alpha/beta heterodimer. Importin alpha recognizes the NLS and acts as an adapter between the NLS and importin beta, whereas importin beta interacts with importin alpha bound to the NLS, and acts as a carrier of the NLS/importin alpha/beta trimer. It is generally thought that importin alpha and beta are part of a large protein family. The leucine rich NES containing proteins are exported from the nucleus by one of the importin beta family molecules, CRM1/exportin 1. A Ras-like small GTPase Ran plays a crucial role in both import/export pathways and determines the directionality of nuclear transport. It has recently been demonstrated in living cells that Ran actually shuttles between the nucleus and the cytoplasm and that the recycling of Ran is essential for the nuclear transport. Furthermore, it has been shown that nuclear transport factor 2 (NTF2) mediates the nuclear import of RanGDP. This review largely focuses on the issue concerning the functional divergence of importin alpha family molecules and the role of Ran in nucleocytoplasmic protein transport. PMID- 10698257 TI - TPA induced expression and function of human connexin 26 by post-translational mechanisms in stably transfected neuroblastoma cells. AB - Connexin 26 (Cx26) has been proposed to be a tumor suppressor gene and its expression may modulate development, cell growth and differentiation in various tissues, including the brain. 12-O-tetradecanoylphorbol-13-acetate (TPA) may serve as either tumor promoter (in mammary gland amd skin) or as a differentiating agent (in neuroblastoma and leukemic cells) and may also modulate expression, function and phosphorylation of gap junctions. In this study, to determine the effects of TPA on Cx26 expression and its function in neuroblastoma, we transfected N2A mouse neuroblastoma cells (which are gap junction deficient) with the coding region of human Cx26 gene (which lacks TPA response elements) and examined the changes of expression and function of Cx26 following 10 nM TPA treatment. Individual clones of transfectants stably expressed distinct levels of exogenous Cx26 as judged by Northern and Western blots, immunocytochemistry and electrophysiological recordings. Cx26 channels displayed unitary conductances of about 140-155 pS. Increase of Cx26 expression following TPA treatment was markedly observed using immunocytochemistry and Western blots of membrane fractions although it was not detected in Northern or Western blots of whole cells. This increase in Cx26 expression in the plasma membrane was accompanied by an increase of function as evidenced in measurements of junctional conductance. These results suggest that induction of exogenous Cx26 in neuroblastoma cells by TPA treatment is controlled by post-translational mechanisms. PMID- 10698258 TI - Extensive degradation of mutant-type D123 protein is responsible for temperature sensitive proliferation inhibition in 3Y1tsD123 cells. AB - A temperature-sensitive mutant of 3Y1, 3Y1tsD123, reversibly arrested in G1 phase of cell cycle at the restrictive temperature of 39.8 degrees C, shows a single amino acid exchange in the D123 protein. In this study, we found that the D123 protein level in 3Y1tsD123, which was 1/8 of that in 3Y1 compared at the permissive temperature of 33.9 degrees C, lowered to 1/4 after a shift to the restrictive temperature. During inhibition of protein synthesis with cycloheximide, the D123 protein level in 3Y1tsD123 decreased markedly depending on the incubation temperature, compared with that in 3Y1, indicating that the lowered levels of D123 protein in 3Y1tsD123 are due to its degradation. Unexpectedly, 2 stably temperature-resistant clones were isolated after transfection of SV-3Y1tsD123 (SV40-transformed 3Y1tsD123, which shows cell death instead of G1 arrest at the restrictive temperature) with the cDNA of the mutant type (3Y1tsD123-derived) D123 protein. The D123 protein in both clones degraded extensively at both temperatures, suggesting that the overexpression of the mutant-type D123 protein exceeds its degradation. Both temperature-resistant clones contained higher levels of D123 protein at the restrictive temperature than did SV-3Y1tsD123 at the permissive temperature. We concluded that the lowered D123 protein level at the restrictive temperature induces the temperature sensitive characteristics of 3Y1tsD123 and SV-3Y1tsD123. PMID- 10698259 TI - Nitric oxide induces apoptosis via Ca2+-dependent processes in the pancreatic beta-cell line MIN6. AB - An excessive production of nitric oxide (NO) in response to cytokines has been shown to be the major cause of the destruction of islet beta-cells associated with type 1 (insulin-dependent) diabetes mellitus. The NO-induced beta-cell death is the typical apoptosis. In the present study, we show evidence that supports a tight link between NO, Ca2+, protease and apoptosis in beta-cells. Three different NO donors, SNAP, NOR3 and NOC7, induced apoptosis in a beta-cell line, MIN6 cells, in a concentration-dependent manner. SNAP at 200 microM increased cytosolic Ca2+ concentration ([Ca2+]i) and induced apoptosis. The SNAP-induced apoptosis was blocked by a Ca2+ chelator, BAPTA-AM, and by an inhibitor of a Ca2+ dependent protease, calpain. In conclusion, an excessive NO production induces apoptosis, wherein an increase in [Ca2+]i and resultant activation of calpain play a key role. PMID- 10698260 TI - Both low and high concentrations of staurosporine induce G1 arrest through down regulation of cyclin E and cdk2 expression. AB - Staurosporine has been reported to cause arrest of cells in G1 phase at low concentration and in G2 phase at high concentration. This raises the question of why the effects of staurosporine on the cell cycle depend on the applied concentration. In order to verify these multiple functions of staurosporine in Meth-A cells, we used cyclin E as a landmark of G1/S transition, cyclin B as a landmark of G2/M transition and MPM2 as a hallmark of M phase. We found that staurosporine arrested cells in G1 phase at a low concentration (20 nM) and in G2/M phase at a high concentration (200 nM). However, 200 nM staurosporine increased the expression of cyclin B and cdc2 proteins, suggesting that the cells progressed through the G2/M transition, and increased the expression of MPM2 protein, indicating that the cells entered M phase. Moreover, 200 nM staurosporine increased the expression of p53 and p21 proteins and inhibited the expression of cyclin E and cdk2 proteins, suggesting that the cells were arrested in the G1 phase of the next cycle. Morphological observation showed similar results as well. These data suggest that the G2/M accumulation induced by 200 nM staurosporine does not reflect G2 arrest, but rather results from M phase arrest, followed by progression from M phase to the G1 phase of the next cycle without cytokinesis, and finally arrest of the cells in G1 phase. PMID- 10698261 TI - Participation of a cathepsin L-type protease in the activation of caspase-3. AB - A previous paper from this laboratory reported the activation of a caspase-3-like protease by a digitonin-treated lysosomal fraction [FEBS Lett. 435, 233-236, 1998]. In this study, we examined the effects of specific inhibitors of lysosomal cysteine proteases, such as cathepsins B, S, and L, on the activation of caspase 3 to find out which cathepsin is responsible for the activation. Pro-caspase-3 in the cytosol was cleaved by a lysosomal protease(s) contained in the supernatant of a digitonin-treated crude mitochondrial fraction containing lysosomes (ML) and the cleaved product was detected by Western blotting using anti-caspase-3 antibody. The activation of caspase-3 by the lysosomal protease(s) was pH dependent and the optimum pH for activation was pH 6.6-6.8. This activation was not inhibited by CA-074, a specific inhibitor of cathepsin B, but was strongly inhibited by CLIK-066 and CLIK-181, specific inhibitors of cathepsin L. The inhibitory effect of CLIK-060, a specific inhibitor of cathepsin S, was very weak. Furthermore, the activation of caspase-3 was enhanced by addition of purified cathepsin L only in the presence of the supernatant of the digitonin treated ML. These results suggested that a cathepsin L-type protease activity might participate in the activation mechanism of caspase-3 in the presence of the supernatnat from the ML. PMID- 10698262 TI - Folding initiation sites and protein folding. AB - The paper discusses the role of local structural preferences of protein segments in the folding of proteins. First a short overview of the local, secondary structures detected in peptides, protein fragments, denatured proteins and early folding intermediates is given. Next the discussion of their role in protein folding is presented based on recent literature and data obtained in our laboratory. In conclusion it is pointed out that, during folding, local structures populated at low levels in denatured state may facilitate the crossing of the folding transition state barrier, and consequently accelerate the rate limiting step in folding. However, the data show that this effect does not follow simple rules. PMID- 10698263 TI - Peptide nucleic acids and their structural modifications. AB - Peptide (polyamide) analogues of nucleic acids (PNAs) make very promising groups of natural nucleic acid (NA) ligands and show many other interesting properties. Two types of these analogues may be highlighted as particularly interesting: the first, containing a polyamide with alternating peptide/pseudopeptide bonds as its backbone, consisting of N-(aminoalkyl)amino-acid units (type I), with nucleobases attached to the backbone nitrogen with the carboxyalkyl linker; and the second, containing a backbone consisting of amino-acid residues carrying the nucleobases in their side chains (type II). So far, these two groups have been studied most intensively. The paper describes main groups of peptide nucleic acids, as well as various other amino acid-derived nucleobase monomers or their oligomers, which were either studied in order to determine their hybridisation to nucleic acids, or only discussed with respect to their potential usefulness in the oligomerisation and nucleic acids binding. PMID- 10698264 TI - Protein inhibitors of serine proteinases. AB - Serine proteinases and their natural protein inhibitors belong to the most intensively studied models of protein-protein recognition. Protein inhibitors do not form a single group but can be divided into about 20 different families. Global structures of proteins representing different inhibitor families are completely different and comprise alpha-helical proteins, beta-sheet proteins, alpha/beta-proteins and different folds of small disulfide-rich proteins. Three different types of inhibitors can be distinguished: canonical (standard mechanism) inhibitors, non-canonical inhibitors, and serpins. The canonical inhibitor binds to the enzyme through the exposed and convex binding loop, which is complementary to the active site of the enzyme. The mechanism of inhibition in this group is consistently very similar and resembles that of an ideal substrate. Non-canonical inhibitors, originating from blood sucking organisms, specifically block enzymes of the blood clotting cascade. The interaction is mediated through inhibitor N-terminus which binds to the proteinase forming a parallel beta-sheet. There are also extensive secondary interactions which provide an additional buried area and contribute significantly to the strength and specificity of recognition. Serpins are major proteinase inhibitors occurring in plasma. Similarly to canonical inhibitors, serpins interact with their target proteinases in a substrate-like manner. However, in the case of serpins, cleavage of a single peptide bond in a flexible and exposed binding loop leads to dramatic structural changes. PMID- 10698265 TI - Coordination chemistry of glutathione. AB - The metal ion coordination abilities of reduced and oxidized glutathione are reviewed. Reduced glutathione (GSH) is a very versatile ligand, forming stable complexes with both hard and soft metal ions. Several general binding modes of GSH are described. Soft metal ions coordinate exclusively or primarily through thiol sulfur. Hard ones prefer the amino acid-like moiety of the glutamic acid residue. Several transition metal ions can additionally coordinate to the peptide nitrogen of the gamma-Glu-Cys bond. Oxidized glutathione lacks the thiol function. Nevertheless, it proves to be a surprisingly efficient ligand for a range of metal ions, coordinating them primarily through the donors of the glutamic acid residue. PMID- 10698266 TI - Molecular modeling of the oxytocin receptor/bioligand interactions. AB - Oxytocin is a nonapeptide hormone (CYIQNCPLG-NH2, OT), controlling labor and lactation in mammalian females, via interactions with specific cellular membrane receptors (OTRs). The native hormone is cyclized via a 1-6 disulfide and its receptor belongs to the GTP-binding (G) protein-coupled receptor (GPCR) family, also known as heptahelical transmembrane (7TM) or serpentine receptors. Using a technique combining multiple sequence alignments with available experimental constraints, a reliable OTR model was built. Subsequently, the OTR complexes with a selective agonist [Thr4,Gly7]OT, a selective cyclohexapeptide antagonist L 366,948 and oxytocin itself were modeled and relaxed using a constrained simulated annealing (CSA) protocol. All three ligands seem to prefer similar modes of binding to the receptor, manifested by repeating receptor residues which directly interact with the ligands. Those involved in the three complexes are putative helices: TM3: R113, K116, Q119, M123; TM4: Q171, and TM5: I201 and T205. Most of them are the equivalent residues/positions to those found in our earlier studies, regarding related vasopressin V2 receptor/bioligand interactions. PMID- 10698267 TI - Hexahistidine (His6)-tag dependent protein dimerization: a cautionary tale. AB - Nickel nitrilotriacetic acid (Ni2+-NTA) immobilization of hexahistidine (His6) tagged proteins has become one of the most commonly used methods of affinity chromatography. Perhaps the greatest utility of this protein purification method stems from the general belief that His-tagged proteins (comprised of His6) are little affected in their activities or efficiencies, while alterations in specificity are unexpected. Although this is certainly true in many instances, we present a case in which the biochemical properties of proteins being studied were fundamentally altered due to the presence of His-tags. We carried out these studies using variants of the pi(30.5) protein of plasmid R6K, a DNA binding protein which negatively regulates plasmid replication. Pi(30.5) can bind DNA containing a target sequence (TGAGR) arranged either asymmetrically (direct repeats) in the gamma origin, or symmetrically in inverted half-repeats (IR's) in the operator of its own gene, pir. Importantly, dimers of pi protein bind to an IR; this property allows researchers to quickly assess whether different regulatory variants of pi proteins exhibit altered dimerization properties. For example, pi(30.5) containing a single amino-acid substitution, F107S (pi200(30.5)), has been shown to be monomeric in solution and dimers were not observed bound to IR's. Here we demonstrate that the presence of a His-tag partially restores the ability of pi200(30.5) to dimerize in solution and bind to an IR in dimeric form. This report sends an important message that (other) proteins containing His-tags may differ from their wild type counterparts in dimerization/oligomerization properties. PMID- 10698268 TI - Modelling of insulin receptor tyrosine kinase in its active form: a case study for validation of theoretical methods. AB - An active form of an insulin receptor tyrosine kinase (IRK) catalytic core was modelled based on its experimentally known inactive form and the active form of a serine/threonine kinase, protein kinase A (PKA). This theoretical model was compared with the crystallographic structure of the active form of IRK reported later. The structures are very similar, which shows that all the most important features and interactions have been taken into account in the modelling procedure. The elaborated procedure can be applied to other tyrosine kinases. This would allow designing of a wide class of tyrosine kinase inhibitors, very important potential anti-cancer and/or anti-viral drugs. PMID- 10698269 TI - Structural polymorphism of telomeres studied by photon correlation spectroscopy. AB - Photon Correlation Spectroscopy (PCS) was used to study the dynamics and structure of Tetrahymena telomeric sequence d(5'-TGGGGT-3')4. Two different modes were observed, corresponding to the following structures: intermolecular (tetramolecular) G-quadruplex and intramolecular (monomeric) G-quartet. Experimental values of translational diffusion coefficients DT were obtained for each structural form. The value of DT for the monomer equals to 1.4 x 10(6) (cm2/s), while for the tetramolecular structure, to 0.8 x 10(6) (cm2/s). The relative weight concentrations of these two forms were analyzed versus the concentration of NaCl varied from 10 mM to 500 mM. The values of experimentally determined diffusion coefficients were compared with those calculated assuming the "bead model" and with the atomic coordinates from the NMR and X-ray crystallographic data. For both structures the experimental and calculated values of DT were in reasonable agreement. In the entire NaCl concentration range studied, the contribution of the relative weight concentration of the monomeric telomere form changed from 85% for 10 mM NaCl to 60% for 500 mM NaCl. PMID- 10698270 TI - Fluorescence decay time distribution analysis of cyclic enkephalin analogues. Influence of the solvents and configuration of amino acids in position 2 and 3 on changes in conformation. AB - The lifetime distribution calculations were applied to study the influence of configuration of amino-acid residues in positions 2 and 3 on changes in conformation of the peptide chain of cyclic analogues of enkephalins containing a fluorescence energy donor and acceptor in different solvents. In all the solvents studied the lifetime distributions were bimodal. This testified to the presence of two families of conformations. In this paper the relationship between the population of each conformation and configuration of the residues in position 2 and 3, and the solvent used is discussed. PMID- 10698271 TI - Construction and optimisation of a computer model for a bacterial membrane. AB - The main steps in the construction of a computer model for a bacterial membrane are described. The membrane has been built of 72 lipid molecules, 54 of which being 1-palmitoyl-2-oleoyl-sn-glycerol-3-phosphatidylethanolamine (POPE) and 18- 1-palmitoyl-2-oleoyl-sn-glycerol-3-phosphatidyl-rac-glycerol (POPG) molecules (thus in the proportion of 3:1). The membrane was hydrated with 1955 water molecules (approximately 27 water molecules per lipid). To neutralise the electronic charge (-e) on each POPG molecule, 18 sodium ions (Na+) were added to the membrane close to the POPG phosphate groups. The atomic charges on the POPE and POPG headgroups were obtained from ab initio quantum mechanical restrained electrostatic potential fitting (RESP) (Bayly et al., 1993, J. Phys. Chem. 97, 10269) using the GAMESS program at the 6-31G* level (Schmidt et al., 1993, J. Comput. Chem. 14, 1347). The model constructed in this way provided an initial structure for subsequent molecular dynamics simulation studies intended to elucidate the atomic level interactions responsible for the structure and dynamics of the bacterial membrane. PMID- 10698272 TI - Conformational analysis of a novel cyclic enkephalin analogue using NMR and EDMC calculations. AB - Conformational space of a novel cyclic enkephalin analogue, cyclo(N(epsilon),N(epsilon')-carbonyl-D-Lys2,Lys5)enkephalin amide, was exhaustively examined. A large number of conformations was selected and clustered into families on the basis of their structure and energy. For representative conformations ROESY spectra were generated and their linear combination was fitted to the spectra measured in water and Me2SO-d6. This procedure yielded an ensemble of most populated conformations of the peptide in the two solvents. PMID- 10698273 TI - NMR measurements of proton exchange between solvent and peptides and proteins. AB - Scope and limitations of the NMR based methods, equilibration and magnetization transfer, for measuring proton exchange rates of amide protons in peptides and proteins with water protons are discussed. Equilibration is applied to very slow processes detected by hydrogen-deuterium exchange after a solute is dissolved in D2O. Magnetization transfer allows to study moderately rapid processes in H2O. A number of precautions should be undertaken in order to avoid systemic errors inherent in the magnetization transfer method. PMID- 10698274 TI - Factors improving the accuracy of determination of 15N relaxation parameters in proteins. AB - A number of factors at all stages of data processing which affect the accuracy of determination of 15N relaxation parameters in 15N-labeled proteins is discussed. Methods which allow to improve accuracy of the determined parameters are presented using data obtained for Cucurbita maxima trypsin inhibitor. PMID- 10698275 TI - A comparative CD and fluorescence study of a series of model calcium-binding peptides. AB - Lanthanide-saturated peptides analogous to calcium-binding loops of EF-hand proteins can be used to stabilize the alpha-helical structure of peptide or protein segments attached to their C-termini. To study conformational properties of such loop-containing hybrids it is necessary to produce them in bacteria. In peptides obtained in this way the helix will be destabilized by the negatively charged C-terminal alpha-carboxyl groups. We propose to block them by the homoserine lactone. The results presented in this paper indicate that the presence of the lactone even at the C-terminus of the loop does not have any negative effect on the loop helix-nucleation ability. On the other hand, the presence of the alpha-NH3+ at the loop N-terminus leads to a drop of metal binding constant and loss of the rigid structure of the alpha-helical segment of the loop. The alpha-amino group separated by one glycine residue from the loop N terminus should also be avoided because it perturbs the conformation of the N terminal part of the loop and may reduce the loop affinity to lanthanide ions. PMID- 10698276 TI - Inhibition of plasminogen activator inhibitor release in endothelial cell cultures by antisense oligodeoxyribonucleotides with a 5'-end lipophilic modification. AB - A series of conjugates containing residues of lipophilic alcohols covalently bound to 5' end of oligodeoxyribonucleotides targeted against human plasminogen activator inhibitor (PAI-1) mRNA was synthesized via the oxathiaphospholane approach. The highest anti-PAI-1 activity in EA.hy 926 endothelial cell cultures was found for conjugates containing menthyl or heptadecanyl groups linked with an oligonucleotide complementary to a segment of human PAI-1 mRNA. The phosphodiester antisense oligonucleotides, which otherwise exhibit only limited anti-PAI-1 activity, were found to be more active than phosphorothioate oligonucleotides when conjugated to lipophilic alcohol residues. For menthyl conjugates an evidence of antisense mechanism of inhibition was found. PMID- 10698277 TI - Interferon gamma bound to endothelial cells is phosphorylated by ecto-protein kinases. AB - The presence of protein kinase activity and its phosphorylated products has been demonstrated on the outer surface of the plasma membrane of endothelial cells. Extracellular phosphorylation was detected by incubation of primary endothelial cells (HUVEC's) and endothelial cell line EA.hy 926 with [gamma-32P]ATP. The reaction products were subjected to SDS/PAGE, autoradiography and scanning densitometry. Under the experimental conditions, five proteins with apparent molecular masses of 19, 23, 55, 88, and 110 kDa were prominently phosphorylated in both types of cells. Phosphorylation of the 19 kDa protein was the most rapid reaching maximum after 60 s and then the protein became dephosphorylated. Ecto protein kinases responsible for the surface labeling of membrane proteins were characterized by using (a) protein kinase C inhibitors: K-252b, chelerythrine chloride, and [Ala113] myelin basic protein (104-118), (b) protein kinase A inhibitor Kemptide 8334, and (c) casein kinase II inhibitor 5,6-dichloro-1-beta-D ribofuranosyl benzimidazole (DRB). Stimulation of endothelial cells with tumor necrosis factor alpha (TNF alpha) and interferon gamma (IFN gamma) is associated with 20-80% reduction of extracellular phosphorylation of all membrane proteins. IFN gamma bound to membrane receptors becomes rapidly phosphorylated. Only in the case of IFN gamma it was associated with the appearance of a strongly phosphorylated band of 17 kDa corresponding to IFN gamma itself. Phosphorylation of this 17 kDa exogenous substrate was prevented by an ecto-kinase inhibitor K 252b. The existence of ecto-phosphoprotein phosphatase activity in endothelial cells was evidenced by testing the effect of microcystin LR--a membrane impermeable reagent that inhibits both PP-1 and PP-2a phosphoprotein phosphatases. The extent of phosphorylation of 19 kDa and 110 kDa phosphoproteins significantly increased in the presence of microcystin. Our results suggest the presence of at least two ecto-kinase activities on endothelial cells that may play a significant role(s) in the regulation of cytokines function. PMID- 10698278 TI - Nitric oxide mediates the mitogenic effects of insulin and vascular endothelial growth factor but not of leptin in endothelial cells. AB - The regulation of vascular wall homeostasis by nitric oxide (NO) generated by endothelium is being intensively studied. In the present paper, the involvement of NO in the vascular endothelial growth factor (VEGF), insulin or leptin stimulated proliferation of human endothelial cells (HUVEC) was measured by [3H]thymidine or bromodeoxyuridine incorporation. VEGF and insulin, but not leptin, increased NO generation in HUVEC, as detected with ISO-NO electrode. Proliferation of HUVEC induced by leptin was not changed or was higher in the presence of N(omega)-nitro-L-arginine methyl ester (L-NAME) a nitric oxide synthase (NOS) inhibitor. In contrast, L-NAME blunted the proproliferative effect of VEGF and insulin. Furthermore, we demonstrated that, in human arterial smooth muscle cells (hASMC) transfected with endothelial NOS (eNOS) gene, the generation of biologically active VEGF protein was NO-dependent. Inhibition of NO generation by L-NAME decreased the synthesis of VEGF protein and attenuated HUVEC proliferation induced by conditioned media from transfected hASMC. Endothelium derived NO seems to participate in VEGF and insulin, but not leptin, mitogenic activity. Additionally, the small amounts of NO released from endothelial cells, as mimicked by eNOS transfection into hASMC, may activate generation of VEGF in sub-endothelial smooth muscle cells, leading to increased synthesis of VEGF protein necessary for turnover and restitution of endothelial cells. PMID- 10698279 TI - Anticoagulative effect of pepsin. AB - Anticoagulative effect of pepsin is observed in vitro when its concentration is 36 microM and higher. This effect is due to inhibition of fibrin monomer polymerization. Protamine abolishes anticoagulative effect of pepsin. Pepsin does not influence platelet aggregation induced by ADP and collagen. PMID- 10698280 TI - Molecular analysis of hemophilia B in Poland: 12 novel mutations of the factor IX gene. AB - We examined the molecular basis of factor IX deficiency in 53 unrelated Polish patients with hemophilia B. Heteroduplex analysis and direct sequencing of polymerase chain reaction (PCR) products were applied to identify the gene defect. Forty-three different point mutations were detected in the factor IX gene of 47 patients. There were 29 missense mutations, 9 nonsense mutations, 4 splice site mutations and 1 point mutation in the promoter region. Twelve mutations were novel. The results of this study emphasize a very high degree of heterogeneity of hemophilia B. PMID- 10698281 TI - Carbohydrate-deficient glycoprotein syndromes. AB - Carbohydrate-deficient glycoprotein syndromes are rare, multisystemic diseases, typically with major nervous system impairment, that are caused by hypo- and unglycosylation of N-linked glycoproteins. Hence, a biochemical evidence of this abnormality, like hypoglycosylation of serum transferrin is essential for diagnosis. Clinically and biochemically, six types of the disease have been delineated. Three of them are caused by deficiencies of the enzymes that are required for a proper glycosylation of lipid--(dolichol) linked oligosaccharide (phosphomannomutase or phosphomannose isomerase or alpha-glycosyltransferase), and one results from a deficiency of Golgi resident N acetylglucosaminyltransferase II. In addition one variant of the disease has been reported as due to a defective biosynthesis of dolichol iself. The diseases are heritable but genetics has been established for only two types. Therapy, based on administration of mannose to patients is currently under investigation. It benefits patients with deficiency of phosphomannose isomerase. Taking into account the complexity of N-linked glycosylation of proteins more of the disease variants is expected to be found. PMID- 10698282 TI - Isoenzymes of N-acetyl-beta-hexosaminidase. AB - Biological significance, structure and posttranslational processing of N-acetyl beta-hexosaminidase isoenzymes are described. Clinical application of N-acetyl beta-hexosaminidase is also reviewed. PMID- 10698283 TI - Application of aqueous hydrazine solution for beta-elimination of O-glycans from gastric mucin. AB - O-Glycans from pig gastric mucin were released by beta-elimination in 0.2 M triethylamine and 50% aqueous hydrazine solution. The released glycan hydrazides were isolated using Centricon 10 separators, brought to their reducing form and reductive by labelled with p-aminobenzoic acid ethyl ester (ABEE). Labelled products were fractionated into neutral and acid fractions on a Bio-Gel P4 column, calibrated with a mixture of dextran oligosaccharides, labelled according to the same procedure. PMID- 10698284 TI - Structure of yellow lupine genes coding for mitotic cyclins. AB - Cell cycle progression in eukaryotes is controlled by complexes of p34 protein kinases and cyclins. For the first time in plants, we have established the sequence of four yellow lupine mitotic cyclin B1 genes. Their coding regions and expression pattern were also characterised recently. Structure of all the four lupine genes is similar: they consist of nine exons and eight introns, analogously located, except Luplu;CycB1;3 lacking 7th intron. Analysis of 5' regulatory sequences of two of them showed that both comprise M-specific activators (MSA), common to plant genes induced in late G2 and early M. Putative repressor binding sites CDE/CHR found in animal G2-specific promoters can also be detected in lupine genes. Controlling region of Luplu;CycB1;4 gene that is highly activated by IAA, contains up to 7 auxin response elements, while insensible to IAA Luplu;CycB1;4 gene have no such motifs. Further studies should be undertaken to determine precisely the functions of putative regulatory elements in the expression of lupine mitotic cyclins. PMID- 10698285 TI - Ras protein of the slime mold Physarum polycephalum is farnesylated in vitro. AB - Physarum Ppras1 protein was efficiently prenylated by prenyltransferases of spinach. Surprisingly in spite of the C-terminal sequence (CLLL) specific for geranylgeranylation the protein was preferentially farnesylated. Consequences of this observation are discussed. PMID- 10698286 TI - Characterization of DNA sequences localized 11 to 17.5 kb upstream of the chicken embryonic pi-globin gene. AB - We have analyzed the DNA fragment localized about 11 to 17.5 kb upstream of the chicken alpha-globin gene domain (the fragment was designed as alpha-0). The nucleotide sequence of its 3.3 kb-long 5' part was established and interactions with nuclear matrix proteins were studied. The DNA region localized about 16 kb upstream of the embryonic pi-globin gene showed high affinity to nuclear matrices in vitro. Two palindromes and a cluster of inverted repeats were co-localized in the same region. The whole 6.6 kb alpha-0 fragment decreased the activity of linked CAT reporter gene when transfected into chicken erythroblastoid cells. PMID- 10698287 TI - Mutagenic specificity of imidazole ring-opened 7-methylpurines in M13mp18 phage DNA. AB - The most abundant lesion formed in DNA upon modification with methylating agents 7-methylguanine, under alkaline conditions is converted into 2,6-diamino-4 hydroxy-5N-methyl-formamidopyrimidine (Fapy-7MeGua). We have previously shown that treatment of dimethylsulfate methylated DNA with NaOH creates mutagenic base derivatives leading to a 60-fold increase in the frequency of A-->G transitions and a 2-3-fold increase of G-->T and G-->C transversions. We have analyzed which lesions lead to these mutations. We compared mutagenic spectra in the lacZ gene of M13mp18 phage DNA modified with dimethylsulfate and NaOH after selective elimination of damaged bases from molecules used for transfection into SOS induced E. coli. Partial elimination of Fapy-7MeGua from phage DNA performed by its digestion with formamidopyrimidine-DNA glycosylase resulted in a 2-3-fold decrease of G-->T and G-->C transversions. Selective depurination of methylated bases (9 h, 37 degrees C, pH 7.0) resulting in almost complete loss of 7MeAde as demonstrated by HPLC analysis of [3H]MNU alkylated phage DNA used as a probe, caused a dramatic, 9-fold decrease of A-->G transitions. Alkali-catalysed rearrangement of 7MeAde was followed by HPLC analysis of [3H]MNU alkylated poly(A) and poly(dA). After incubation of these oligonucleotides in NaOH, 7MeAde disappeared from both chromatograms, but only in polyA, 2 new peaks migrating with retention time different from that of 1MeAde, 3MeAde or 7MeAde were detected, suggesting formation of two rotameric forms of Fapy-7MeAde as observed for Fapy-7MeGua. Thus the miscoding lesion, giving rise to A-->G transitions derived from 7MeAde was Fapy-7MeAde. Fapy-7MeGua was at least an order of magnitude less mutagenic, but in SOS-induced cells it gave rise to G-->T and G- >C transversions. PMID- 10698288 TI - Annexin VI: an intracellular target for ATP. AB - Annexin VI (AnxVI), an Ca2+- and phospholipid-binding protein, interacts in vitro with ATP in a calcium-dependent manner. Experimental evidence indicates that its nucleotide-binding domain which is localized in the C-terminal half of the protein differs structurally from ATP/GTP-binding motifs found in other nucleotide-binding proteins. The amino-acid residues of AnxVI directly involved in ATP binding have not been yet defined. Binding of ATP to AnxVI induces changes in the secondary and tertiary structures of protein, affecting the affinity of AnxVI for Ca2+ and, in consequence, influencing the Ca2+-dependent activities of AnxVI: binding to F-actin and to membranous phospholipids, and self-association of the annexin molecules. These observations suggest that ATP is a functional ligand for AnxVI in vivo, and ATP-sensitive AnxVI may play the role of a factor coupling vesicular transport and calcium homeostasis to cellular metabolism. PMID- 10698289 TI - RT-PCR and Northern blot analysis in search for a putative Paramecium beta adrenergic receptor. AB - RT-PCR and Northern blot analysis were performed in order to search for a putative beta-adrenergic receptor (beta-AR) in Paramecium using several beta2 adrenergic-specific molecular probes. Under strictly defined RT-PCR conditions DNA species of expected molecular size about 360 bp were generated with the primers corresponding to the universal mammalian beta2-AR sequence tagged sites (located within the 4th and the 6th transmembrane regions of the receptor). This RT-PCR product hybridized in Southern blot analysis with the oligonucleotide probe designed to the highly conservative beta2-AR region involved in G-proteins interaction and located within the amplified region. Northern hybridization was performed on Paramecium total RNA and mRNA with human beta2-AR cDNA and two oligonucleotide probes: the first included Phe 290 involved in agonist binding (Strader et al., 1995) and the second was the backward RT-PCR primer. All these probes revealed the presence of about 2 kb mRNA which is consistent with the size of beta2-AR transcripts found in higher eukaryotes. PMID- 10698290 TI - Effect of thyroid hormone on the myosin heavy chain isoforms in slow and fast muscles of the rat. AB - The myosin heavy chain (MHC) was studied by biochemical methods in the slow twitch (soleus) and two fast-twitch leg muscles of the triiodothyronine treated (hyperthyroid), thyroidectomized (hypothyroid) and euthyroid (control) rats. The changes in the contents of individual MHC isoforms(MHC-1, MHC-2A, MHC-2B and MHC 2X) were evaluated in relation to the muscle mass and the total MHC content. The MHC-1 content decreased in hyperthyreosis, while it increased in hypothyreosis in the soleus and in the fast muscles. The MHC-2A content increased in hyperthyreosis and it decreased in hypothyreosis in the soleus muscle. In the fast muscles hyperthyreosis did not affect the MHC-2A content, whereas hypothyreosis caused an increase in this MHC isoform content. The MHC-2X, present only in traces or undetected in the control soleus muscle, was synthesised in considerable amount in hyperthyreosis; in hypothyreosis the MHC-2X was not detected in the soleus. In the fast muscles the content of MHC-2X was not affected by any changes in the thyroid hormone level. The MHC-2B seemed to be not influenced by hyperthyreosis in the fast muscles, whereas the hypothyreosis caused a decrease of its content. In the soleus muscle the MHC-2B was not detected in any groups of rats. The results suggest that the amount of each of the four MHC isoforms expressed in the mature rat leg muscles is influenced by the thyroid hormone in a different way. The MHC-2A and the MHC-2X are differently regulated in the soleus and in the fast muscles; thyroid hormone seems to be necessary for expression of those isoforms in the soleus muscle. PMID- 10698291 TI - An isotopic assay of dUTPase activity based on coupling with thymidylate synthase. AB - A new rapid, sensitive and convenient procedure is presented allowing determination of dUTPase activity. With [5-(3)H]dUTP used as the substrate, dUTPase, converts it to the corresponding monophosphate and is coupled with thymidylate synthase-catalyzed reaction, resulting in tritium release from [5 (3)H]dUMP. Following charcoal absorption of the labeled nuleotides, radioactivity of tritiated water is determined. The new assay was tested to show comparable results with a previously described assay, based on measuring dUTPase-catalyzed [5-(3)H]dUMP production. PMID- 10698292 TI - Congenital dyserythropoietic anemias. AB - The congenital dyserythropoietic anemias (CDAs) are an uncommon and heterogeneous group of disorders characterized by markedly ineffective erythropoiesis and, usually, striking dysplastic changes in erythroblasts. Each of the three originally described forms, designated CDA types I to III, is defined by the presence of distinctive morphologic (including ultrastructural) abnormalities in erythroblasts. CDA type II is also characterized by a marked reduction in polylactosamine structures associated with the erythrocyte membrane glycoprotein, band 3 (detected by sodium dodecyl sulfate polyacrylamide gel electrophoresis), and, usually, a positive result on the acidified serum lysis test. The course of CDA is often complicated by cholelithiasis. Even patients who have not had transfusions sometimes develop substantial iron overload. Recent studies have extended our knowledge on the clinical manifestations of CDA types I and III and have revealed the existence of forms of CDA distinct from types I to III. Information is now available on the chromosomal localization of the genes involved in CDA types I and II and in the Swedish cases of CDA type III. A few patients with CDA type I have been treated with interferon-alpha2, with a good response. PMID- 10698293 TI - Zebrafish: a genetic approach in studying hematopoiesis. AB - The zebrafish (Danio rerio) has emerged in recent years as an exciting animal model system for studying vertebrate organ development and, in particular, the development of the hematopoietic system. The combined advantages of developmental biology and genetic screens for mutations in zebrafish have provided insights into early events in hematopoiesis and identified several genes required for normal blood development in vertebrates. As a result of the large-scale mutagenesis screens for developmental mutants, several zebrafish mutants with defects in blood development have been recovered. This review discusses how these blood mutations in zebrafish have given new perspectives on hematopoietic development. PMID- 10698294 TI - Diamond-Blackfan anemia. AB - Diamond Blackfan anemia is a rare congenital hypoplastic anemia that usually presents early in infancy. Congenital anomalies, in particular of the head and upper limbs, are present in about 25% of reported patients. The disease is characterized by a moderate to severe macrocytic anemia, occasional neutropenia or thrombocytosis, a normocellular bone marrow with erythroid hypoplasia, and an increased risk of developing leukemia. Recent genetic studies have led to the identification of mutations in the ribosomal protein RPS19 in approximately 25% of sporadic and familial cases, a second gene on chromosome 8p, and evidence for an additional locus (or loci). The pathogenesis is unknown. The majority of patients respond to prednisone, and often erythropoiesis can be maintained with low doses of the drug. Both remissions and increased resistance to steroid treatment can occur. Patients who do not respond to treatment are usually transfusion dependent, although responses to high dose steroid, androgen, and interleukin-3 have been observed. Bone marrow transplantation can be curative. PMID- 10698295 TI - Bone marrow transplantation for hemoglobinopathies. AB - In patients with hemoglobinopathy who are treated by allogeneic matched sibling bone marrow transplantation before the onset of disease-associated organ damage, long term, disease-free survival currently stands at approximately 90%, and transplant-associated mortality is 5% or less. Less toxic nonmyeloablative conditioning regimens that have the potential to reduce procedure-related mortality to even lower levels are under active investigation. Expansion of the donor pool by use of unrelated matched donors awaits improvement in HLA-typing methodology. PMID- 10698296 TI - Stroke prevention in sickle cell disease. AB - Neurological complications, especially stroke, have long been recognized in sickle cell disease. Advances in care have increased the life expectancy of such patients, and recent information has better established the epidemiology of stroke. Prevention of stroke in children has been established in a clinical trial. Silent brain lesions revealed by MRI are common and are associated with impairments of cognitive function. Transfusion remains the primary mode of prevention and treatment for stroke, although interest is increasing in hydroxyurea; however, there are no data regarding its efficacy. PMID- 10698297 TI - Clinical and hematologic aspects of hemoglobin E beta-thalassemia. AB - Hemoglobin E beta-thalassemia is an important cause of childhood chronic disease in Southeast Asia. It is characterized by the presence of hemoglobin E and F, and the amount of hemoglobin E ranges from 35% to 75%. The patients are generally classified as having thalassemia intermedia because they have inherited a beta thalassemia allele and hemoglobin E, which acts as a mild beta+-thalassemia. However, a remarkable variability in the clinical expression, ranging from a mild form of thalassemia intermedia to transfusion-dependent conditions, is observed. Severe hemoglobin E beta-thalassemia may have clinical features of thalassemia major. Phenotypes of thalassemia major can be predicted from the early onset of clinical symptoms and the requirement of regular blood transfusion from infancy for survival. Coinheritance of alpha-thalassemia alleviated the severity of beta thalassemia disease in patients with at least one allele of mild beta-thalassemia genotype. PMID- 10698298 TI - Oxidation and erythrocyte senescence. AB - Direct macrophage recognition of an externalized phosphatidylserine signal on senescent erythrocytes is a process of erythrophagocytic clearance that is in line with the general clearance process of all other circulating cells that become apoptotic. Advances in deciphering this process suggest that oxidation of the erythrocyte's hemoglobin, the salient target of the free radicals encountered in the circulatory environment, may drive subsequent steps. The progressive accumulation of oxidized hemoglobin covalently bound to the membrane skeleton not only disrupts membrane organization but also threatens eventual phospholipid oxidation via a calcium-promoted quasi-lipoxygenase activity. The emergence on the cell surface of a threshold concentration of oxidized phospholipids, principally phosphatidylserine, signals recognition by the CD36 macrophage receptor. PMID- 10698299 TI - New insights into erythrocyte membrane organization and microelasticity. AB - The erythrocyte membrane's ability to withstand the stresses of circulation has its origins in various levels of structural organization. Central to this membrane's structure-function relationships is a quasi-two-dimensional meshwork of spectrin-actin-protein 4.1 that imparts a resilence to the overlying plasma membrane. New insights into the nonlinear microelasticity of this substructure are being provided by experiments that range from elegant atomic force microscopy tests of single spectrin chains to patterned photobleaching of the micropipette deformed network. Breakthroughs in atomic level structure determinations are further complemented by emerging biophysical studies of transgenically engineered mice lacking specific erythrocyte membrane proteins. Recent theoretical efforts (computational approaches most notably) also have begun to correlate molecular scale aspects of structure with mechanical measures. All of this recent activity in the biophysics of erythrocyte structure-function is certain to challenge and refine some of the most basic tenets in cell membrane structure-function. PMID- 10698300 TI - New insights into functions of erythroid proteins in nonerythroid cells. AB - This article presents new insights into potential roles that three erythrocyte cytoskeletal proteins, protein 4.1, ankyrin, and spectrin, may play in nonerythroid nucleated cells. Each of these proteins is encoded by several closely related genes characterized by complex alternative splicing of their pre mRNA, thus resulting in the cellular expression of a broad repertoire of isoforms that can adopt tissue- and cell-specific distribution. This could account for the presence of skeletal networks in intracellular organelles such as lysosomes, the Golgi apparatus, or the nucleus. In addition to providing structural support to cell membranes, these cytoskeletal proteins regulate the functions of various transmembrane proteins they interact with, in particular ion channels, as well as the activity of membrane-bound enzymes. Thus, they appear to be key players in major unsuspected cell functions such as protein sorting, dynamics of nuclear architecture during mitosis, or regulation of signal transduction pathways. PMID- 10698301 TI - Bibliography. Current world literature. Erythrocytes and their disorders. PMID- 10698302 TI - Regulation of Src-family protein tyrosine kinase transcription during lymphocyte ontogeny. AB - The distribution and quantity of cellular signaling elements influence response patterns to a variety of stimuli. As protein tyrosine phosphorylation is a requisite event induced by a majority of surface receptors, and protein tyrosine kinases of the src-family (src-PTKs) act as proximal transducers for many hematopoietic receptors, we have designed a quantitative RT-PCR assay to measure src-family PTK expression during critical stages of lymphocyte ontogeny. With this assay we demonstrate that the distal promoter element regulating expression of lck, a src-PTK essential for T-cell development and activation, is similarly regulated during ontogeny of T and B cells. However, lck transcript abundance is drastically reduced in B lineage cells, suggesting that transcriptional elements influencing lck promoter activity are modulated in these cells. Moreover, although transcripts encoding the src-PTK fyn accumulate at 0.1% of lck mRNA levels in thymocytes, diminished activity of the lck distal promoter in the B cell background brings lck and fyn transcript levels to near equivalence in this population. Importantly, transcripts arising from the lck distal promoter element and the fyn locus are similarly upregulated during developmental transitions associated with antigen-receptor expression in both B and T cells. These findings suggest that although the magnitude of lck and fyn expression is differentially regulated in B and T cells, expression at these loci is similarly developmentally programmed during ontogeny of both lymphocyte lineages. PMID- 10698303 TI - Post-translational modifications of immunoglobulin G: a mouse IgG variant that lacks the entire CH1 domain. AB - In the present study, we characterized the post-translational modifications of a short-chain variant of mouse IgG2a that lacks the entire CH 1 domain. The short chain IgG2a and its proteolytic fragments were subjected to electrospray ionization- and fast atom bombardment-mass spectrometric analyses. It has been demonstrated that approximately 14% of the heavy chain of the short-chain IgG2a is O-glycosylated with a disaccharide of Ga1-GalNAc- at Thr220A in the hinge region. while the Oglycosylation does not occur in its parent IgG2a molecule. Two additional modifications have been detected at the C-termini of both the heavy and light chains of the short-chain IgG2a. Biological significance of the post translational modifications of the short-chain IgG2a variant is briefly discussed. PMID- 10698304 TI - Activation and tyrosine phosphorylation of protein kinase C delta in response to B cell antigen receptor stimulation. AB - Activation of the B cell through antigen receptor (BCR) crosslinking is known to initiate a prominent tyrosine kinase cascade and lipid second messenger production through the activation of phospholipase Cgamma and phosphatidylinositol 3' kinase. In this study, we demonstrate that protein kinase C delta (PKCdelta) responds to crosslinking of the BCR by becoming activated and tyrosine phosphorylated within 30s of stimulation. PKC6 activation was dependent primarily on phosphatidylinositol 3' kinase, and this in turn was dependent on an upstream tyrosine phosphorylation event. Inhibition of PKCdelta activation by blocking phosphatidylinositol 3' kinase was also accompanied by a decrease in its tyrosine phosphorylation, suggesting that PKCdelta must be activated in order to become tyrosine phosphorylated. Inhibition of phospholipase C activation had an insignificant effect on the activation of PKCdelta, however it attenuated the tyrosine phosphorylation of PKCdelta. This suggests a distinct role for phospholipase C in the regulation of PKCdelta. This report describes a role for PKCdelta in response to the combined signals originated by the BCR. PMID- 10698305 TI - Polymorphism of the human TNF-alpha promoter--random variation or functional diversity? AB - As more and more polymorphisms are discovered in the genes encoding cytokines, a crucial question is whether this polymorphism has any functional effect. One of the most widely studied cytokine genes in this respect is the gene encoding human TNF-alpha. Much of the literature investigating the issue of whether TNF-alpha promoter polymorphisms have any functional effect on TNF-alpha transcription or influence disease susceptibility appears to report negative results, giving the appearance and leading some authors to conclude that polymorphism at this locus is functionally silent and exists only because of linkage disequilibrium with selectable HLA alleles. This review presents a new analysis of the available data which suggests that polymorphism in the TNF-alpha promoter is not randomly distributed and therefore that it most likely does have some functional and selectable effect. Further, a comparison of available data suggests that there is more consensus in the literature than may at first appear to be the case. PMID- 10698306 TI - SHP-1 variant proteins are absent in motheaten mice despite presence of splice variant transcripts with open reading frames. AB - Motheaten mice have a mutation that causes abnormal splicing of the SHP-1 gene producing transcripts that are out of frame. Thus, motheaten mice cannot produce normal SHP-1 protein. However, we have found that the SHP-1 locus in normal mice is expressed as multiple in-frame splice variant transcripts. We report here that the motheaten SHP-1 gene is likewise expressed in multiple spliced forms, two of which are in frame. One of these two variants, which is also present in normal mice, lacks the exon containing the motheaten mutation and is therefore expected to encode an active phosphatase with only one of the two SH2 domains of SHP-1. We showed that both of these variants produce phosphatases with a higher specific activity than SHP-1, suggesting that motheatein mice are not SHP-1 null. The possibility that motheaten mice produce disregulated phosphatases offered a simple explanation for the puzzling observation that substrates of SHP-1 are hypo phosphorylated in motheaten mice. We tested this by measuring for SHP-1 protein and activity in motheaten macrophages. However, we did not detect specific activity, and found that one of these variant proteins was unstable. These findings likewise suggest that little or no SHP-1 variant proteins exist in normal cells. PMID- 10698307 TI - Differential regulation of germinal center genes, BCL6 and SWAP-70, during the course of MAIDS. AB - Germinal centers (GC) are the sites of antigen-driven B cell switch recombination, V(D)J gene hypermutation, and selection to generate high-afinity CD38+ memory B cells. A marked expansion of GC associated with hypergammaglobulinemia followed by complete disruption of normal splenic architecture and a striking drop in immunoglobulin levels are prominent features of the murine retrovirus-induced immunodeficiency syndrome, MAIDS. B cell lymphomas are frequent in long-term infected mice. Normal GC formation is critically dependent on a number of genes including the transcription factor, Bcl6. Deregulated expression of BCL6 protein has been implicated in the development of human and mouse B cell lymphomas. Another nuclear protein, SWAP 70, has been identified as a subunit of the protein complex, SWAP, that recombines switch regions in vitro. To develop a fuller understanding of B cell biology in MAIDS, we examined the characteristics of BCL6, SWAP-70, CD38, and peanut agglutinin (PNA)-staining cells during the course of the disease. The levels of both nuclear proteins increased rapidly until 6-8 weeks after infection. During this time frame, BCL6 was expressed at highest levels in the usually rare CD4+ Thyl- T cell subset as well as in B cells. At later times. BCL6 levels dropped to undetectable levels while SWAP-70 levels continued to increase. Changes in the levels of either protein could not be ascribed to transcriptional regulation. PNA-reactive cells decreased in concert with BCL6 while CD38 staining increased with SWAP-70. These results demonstrate that progression of MAIDS results in the massive accumulation of B cells with the morphology of secretory cells that behave like post-GC cells for expression of BCL6 and CD38, and for PNA staining but with abnormally high-level expression of SWAP-70. PMID- 10698308 TI - Non-anaphylactic combination of partially deleted fragments of the major house dust mite allergen Der f 2 for allergen-specific immunotherapy. AB - Allergen-specific immunotherapy, in which repeated injections of allergens over prolonged periods are used to induce tolerance, has proven an effective treatment of allergy. A major side effect of allergen-specific immunotherapy is anaphylactic reaction. House dust mite allergens are major causative factors associated with various allergic diseases. Der f 2 is the major house dust mite allergen composed of 129 amino acid residues. Analysis using deletion mutants of Der f 2 suggested that T-cell epitopes of Der f 2 were multiple in mite-allergic patients. We found that some IgE epitopes were renatured by dialysis of a mixture of two denatured C- and N-terminal deletion mutants, 1-112 and 85-129 in 13 patients out of 14. On the other hand, IgE binding activity was negative in the separately dialyzed fragments and their mixture in each patient tested. Furthermore, we demonstrated that neither of the two separately prepared polypeptides induced in vivo skin prick test reactivity. These findings are important for improvement of T-cell targeting allergen-specific immunotherapy and development of monovalent IgE haptens. The use of combinations of overlapping non anaphylactic fragments of allergen covering all of the T-cell epitopes achieves the removal of IgE reactivity, the cause of harmful anaphylactic reactions, without affecting the T-cell reactivity essential for immunotherapy, offering potentially safer and more effective treatment for allergic disease. PMID- 10698309 TI - A human early B-cell factor-like protein participates in the regulation of the human CD19 promoter. AB - CD19 is a functional component of the B-cell receptor complex where it acts as a modulator of the cellular response to surface immunoglobulin signaling. The gene is expressed from early B-cell developmental stages until the mature B-cell stage, but not in the plasma cell. The human CD19 promoter has been suggested to be regulated by the B-cell specific activator protein. BSAP, the Erg transcription factor and unidentified factors interacting with a GC rich binding site denoted PyG. In this report we present data suggesting that one of the PyG interacting factors is related to mouse early B-cell factor (EBF). Recombinant mouse EBF binds to the PyG site with an affinity about 3-fold lower than to the EBF binding site from the mouse mb-1 promoter in electrophoretic mobility shift assays. Furthermore, the PyG box binds to a factor in nuclear extracts from human B-cell lines that also interact with the mouse mb-1 promoter EBF binding site. Mutation of the PyG box impaired binding of the factor and the function of a minimal CD19 promoter in human cells of the B lineage, but not in Jurkat T or non lymphoid HeLa cells. In addition to this, murine EBF was able to activate a wild type but not a PyG mutant human CD19 promoter 7-fold upon transient co transfection in HeLa cells. Thus, we suggest that a human homologue of mouse EBF participate in transcriptional regulation of the human CD19 promoter. PMID- 10698310 TI - CDR substitutions of a humanized monoclonal antibody (CC49): contributions of individual CDRs to antigen binding and immunogenicity. AB - One of the major obstacles in the successful clinical application of monoclonal antibodies has been the development of host immune responses to murine Ig constant and variable regions. While the CDR grafting of MAbs may alleviate many of these problems, the potential remains that one or more murine CDRs on the human Ig backbone of a "humanized" MAb may still be immunogenic. Studies were undertaken employing a MAb of potential clinical utility, CC49, to define those CDRs that are essential for antigen binding and those that may be immunogenic in humans. We previously developed a humanized CC49 (HuCC49) by grafting the MAb CC49 hypervariable regions onto frameworks of human MAbs. To identify those CDRs essential for binding, a panel of variant HuCC49 MAbs was generated here by systematically replacing each of the murine CDRs with their human counterparts. The relative affinity constant of each variant was determined. Serum from a patient who received murine CC49 was used to determine the potential immunogenicity of each CDR in humans. The serum was shown to react with the anti CC49 variable region. Results showed that patients' anti-idiotypic responses are directed mainly against LCDR3 and moderately against LCDR1 and HCDR2. These studies demonstrate for the first time that variants containing individual CDR substitutions of a humanized MAb can be constructed, and each CDR can be defined for the two most important properties for potential clinical utility: antigen binding and immunogenicity. PMID- 10698311 TI - Protein-protein interaction of the Ro-ribonucleoprotein particle using multiple antigenic peptides. AB - Protein protein interactions play a significant role in maintaining the structural and functional integrity of the cell. We used multiple antigen peptides (MAPs) to analyze such interactions within the Ro (or SSA) ribonucleoprotein complex. Our data showed that 60 kD Ro and La colocalize in the nucleus of the cell. Previous data have indicated that 60 kD Ro and La co-exist via interactions with the hYRNAs. We were interested to see whether 60 kD Ro and La interact with each other through protein protein interactions. MAPs were produced with sequences derived from the autoepitopes of 60 kD Ro. When used in agarose immunodiffusion certain MAPs formed precipitin lines specifically with Ro and La antigens. Used in affinity chromatography the Ro MAPs purified the Ro ribonucleoprotein particle from lymphocyte extract. Solid phase immunoassay and surface plasmon resonance (SPR) confirmed the observations obtained with agarose diffusion. Using SPR, kinetic analyses gave an apparent affinity constant of about 1 x 10(7) M(-1) for Ro-MAP-60 kD Ro interactions. The autoantigens Ro and La are specific targets in autoimmune diseases, particularly systemic lupus erythematosus (SLE) and Sjogren's syndrome, and are known to exist together as a complex with hYRNAs. The present data indicate that there are protein-protein interactions between Ro and La. PMID- 10698312 TI - A potent neutralizing monoclonal antibody can discriminate amongst IFNgamma from various primates with greater specificity than can the human IFNgamma receptor complex. AB - A monoclonal antibody (AF2) generated against recombinant human interferongamma (IFNgamma) exhibited potent IFNgamma neutralizing activity and prevented human IFNgamma from binding to the cell surface IFNgamma receptor complex. The AF2 antibody also neutralized IFNgamma from higher primates (superfamily Hominoidea) but did not react with IFNgamma from rhesus or other primates in the suborder Anthropoidea IFNgamma from all primates tested, however, could signal via the human IFNgamma receptor complex, as indicated by the ability to upregulate the level of MHC class II molecule expression on the surface of a responsive human cell line. We cloned and sequenced the IFNgamma gene from chimpanzee, gorilla, orangutan, and gibbon, and compared those with the previously reported IFNgamma sequences of human, rhesus, baboon and marmoset. This comparison revealed that, of the primate IFNgammas that were not reactive with AF2, rhesus IFNgamma was most homologous to human IFNgamma, differing at only nine amino acids and containing a one amino acid deletion. Comparing the sequence of human IFNgamma with that of rhesus IFNgamma suggested residues of the human IFNgamma molecule that were involved in the formation of the epitope recognized by the AF2 antibody. Constructing human/rhesus chimeric IFNgamma molecules, combined with site-directed mutagenesis of both human and rhesus IFNgamma revealed that this epitope was dependent upon two non-contiguous amino acids that are juxtaposed in the tertiary structure of IFNgamma. The determinant recognized by AF2 antibody resides in a portion of IFNgamma that is proximal to, but distinct from the surface that interacts with the IFNgamma receptor. Therefore, this neutralizing monoclonal antibody reacts with a conformational determinant that distinguishes primate IFNgammas serologically, but not functionally. PMID- 10698313 TI - Sedimentation equilibrium analysis of recombinant mouse FcRn with murine IgG1. AB - The interaction of mouse IgG1 or IgG1-derived Fc fragment with recombinant, insect cell expressed mouse FcRn has been analyzed using sedimentation equilibrium. This results in a model for the interaction in which the two binding sites for FcRn on Fc or IgG1 have significantly different affinities with macroscopic binding constants of < 130 nM and 6 microM. This data indicates the formation of an asymmetric FcRn:Fc (or IgG1):FcRn complex which is consistent with earlier suggestions that for this form of recombinant FcRn, binding to IgG1 or Fc does not result in a symmetric 2:1 complex in which both binding sites are equivalent. PMID- 10698314 TI - The immunome. PMID- 10698315 TI - Immunostimulatory properties of the Leishmania infantum heat shock proteins HSP70 and HSP83. AB - Emerging evidence indicates that the heat shock proteins (HSPs), a set of highly evolutionary conserved proteins, are playing essential roles in both normal processes of the immune system and specific immune responses. In a previous work, we demonstrated that the Leishmania infantum HSP70 possesses remarkable immunostimulatory properties. In the present work, we have extended the study to another HSP from this parasite, the HSP83. We show that this protein also has an adjuvant effect to an accompanying protein by stimulation of the humoral response when both proteins are fused and co-administered to BALBjc mice. The analysis of the IgG isotypes, IgG1 and IgG2a, indicated that the immunisations with the Leishmania HSPs, mainly the HSP70, potentiate a Thl-type response. It was found that the amino-terminal domain of the HSP70, the most evolutionary conserved region of the molecule, maintains the ability to stimulate the humoral response, whereas the carboxyl-terminal domain does not have a similar effect. Unexpectedly, we found that the L. infantum HSP70 and HSP83 recombinant proteins stimulated the proliferation of spleen cells from unprimed BALB/c mice. Remarkably, this proliferation was abolished either by thermal denaturing of the proteins or by using specific antibodies. The use of the T-cell inhibitor cyclosporin A in the splenocytes proliferation assays suggested that both T- and non-T-cells are stimulated by the Leishmania HSPs. These findings may be relevant for therapeutic and prophylactic applications. PMID- 10698316 TI - A search within the IL-1 type I receptor reveals a peptide with hydropathic complementarity to the IL-1beta trigger loop which binds to IL-1 and inhibits in vitro responses. AB - In previous research, we were able to demonstrate that a seven amino acid residue peptide (VITFFSL), designed as an antisense peptide of the beta-bulge trigger loop region of interleukin 1beta (IL-1beta) (QGEESND; residues 48-54 [mature protein sequence]), was able to interact with IL-1 specifically and inhibit the response to IL-1 in an in vitro bioassay. The evidence was consistent with a specific interaction ocurring between antisense peptide and the trigger loop region. On the basis that antisense peptides are able to interact with their corresponding sense peptide sequences as a result of their mutually complementary hydropathic profiles (Fassina G., Verdoliva, A., Cassani, G., Melli, M., 1994. Binding of type I IL-1 receptor fragment 151-162 to IL-1. Growth Factors 10, 99 106; Maier, C.C., Moseley, H.N.B., Zhou, S., Whitaker, J.N., Blalock, J.E., 1994. Indentification of interactive determinants on idiotypic-anti-idiotypic antibodies through comparison of their hydropathic profiles. Immunomethods 5, 107 113), we devised a computer program (FINDH) to search the amino acid residue sequence of interleukin-1 type 1 receptor (IL-1 R1) for peptide motifs possessing hydropathic complementarity to the trigger loop sequence. The most complementary "best-fit peptide" motif (LITVLNI) was located in the third extracellular domain of IL-1 R1. A best-fit peptide corresponding to this motif was synthesised and found to bind to IL-1beta as well as inhibit the response to IL-1 in two independent in vitro bioassays (monitoring IL-1 dependent serum amyloid A synthesis and IL-1 dependent alkaline phosphatase activity, respectively). A second peptide motif (VIEFITL) was identified and the corresponding peptide synthesised along with a reordered version (LTILINV) of the best fit peptide. Both failed to bind measurably with IL-1beta or inhibit the response to IL-1 in the two bioassays. This best fit peptide behaved very similarly, in terms of IL-1 binding and inhibition behaviour, to the original trigger loop antisense peptide. Reference to the recently released X-ray crystal structure of IL-1beta and the IL1-R1 extracellular domain shows that the best fit peptide motif in IL-1 R1 is not apparantly interacting with the IL-1 trigger loop, although both are close in space. The intriguing possibility exists that the best fit peptide motif could represent an alternative site for IL-1beta receptor interaction which has not thus far been identified. PMID- 10698317 TI - Effect of pressure on antigen-antibody complexes: modulation by temperature and ionic strength. AB - Changes in the equilibrium binding affinity of antigen-antibody complexes subjected to hydrostatic pressures of about 2000 bar provide a potential means for the separation and recovery under mild conditions of biological molecules from immunoadsorbents or immunosensors. We have investigated the ability of temperature and ionic strength to modulate the pressure sensitivity of several antigen-antibody complexes in solution. For two different protein:monoclonal antibody complexes (BSA:9.1 and HEWL:HyHEL-10) exhibiting pressure-induced dissociation (positive association volume), we find little temperature dependence to the association volume. For another complex (digoxigenin:26-10) exhibiting pressure-induced association, the association volume increases with temperature, which, via a Maxwell relation, indicates that enthalpic changes drive the pressure effect. An increase in ionic strength decreases the affinity of binding the HEWL:HyHEL-5 complex, which contains several salt bridges. At low ionic strengths (<0.3 M), no pressure dependence of the free energy of association is observed, but at higher ionic strengths, significant pressure-induced association is observed, suggesting that positive contributions to the association volume provided by the salt bridges are counterbalanced by other (e.g., aromatic stacking) interactions that lead to negative association volumes. These results suggest that ionic strength may be used to modulate the pressure sensitivity of antigen antibody complexes, which may be useful in designing processes that exploit this phenomenon for immunoseparations. PMID- 10698318 TI - Junctional diversity in Xenopus immunoglobulin light chains. AB - Xenopus cDNA sequences encoding the homolog of mammalian kappa (kappa) light (L) chains were isolated from isogenic tadpole and adult individuals to investigate whether there existed stage-specific immunoglobulin L chain expression and somatic diversification. In the course of these studies rearrangements to a sixth J(L) gene segment and a pseudogene (J(L)psi) were found, and it is suggested that the order of these gene segments with respect to the L chain constant (C) region exon is: J(L)6-J(L)1-J(L)2-J(L)3-J(L)4-J(L)5-J(L)psi-C(L). The cDNA junctional diversity was analyzed; few N and P regions were found and almost all the CDR3 were 9 codons in length. There were restricted patterns of recombination site resolution, and this is attributed to some constraint in JL coding end processing. PMID- 10698319 TI - Hypomethylation is necessary but not sufficient for V(D)J recombination within a transgenic substrate. AB - Although an inverse correlation between CpG methylation and V(D)J recombination has been demonstrated for both artificial substrates and endogenous genes, it is not known whether all hypomethylated targets are competent to rearrange or if other factors are required. We have created several artificial V(D)J recombination substrate transgenes whose methylation can be controlled by breeding into different genetic backgrounds. A transgene which contains the immunoglobulin heavy chain intronic enhancer rearranges efficiently in B lymphocytes when the transgene loci are unmethylated. When the same loci become methylated, upon breeding into a different mouse strain, no rearrangement can be detected. A similar transgene, but lacking the enhancer, also shows no evidence of V(D)J recombination when it is methylated. Even when this enhancerless transgene is hypomethylated, however, no V(D)J recombination can be detected in B lymphocytes. Thus, hypomethylation is required to permit V(D)J recombination but not all hypomethylated targets are capable of recombination. The results may indicate that the immunoglobulin enhancer is required for the assembly of factors involved in V(D)J recombination. PMID- 10698320 TI - Identification and characterisation of human Junctional Adhesion Molecule (JAM). AB - It is widely believed that migrating immune cells utilise the intercellular junctions as routes of passage, and in doing so cause the transient disruption of junctional structures. Thus there is much interest in the molecules that have been identified at cell-cell contact points and their potential involvement in the control of leukocyte diapedesis. In this report we describe the human orthologue to Junctional Adhesion Molecule (JAM), a recently identified member of the immunoglobulin superfamily expressed at intercellular junctions (Martin Padura et al., 1998). The human protein shares a highly conserved structure and sequence with the murine protein. However it is distinct in that it is constitutively expressed on circulating neutrophils, monocytes, platelets and lymphocyte subsets. This broad expression pattern is similar to another IgSF molecule, CD31, expressed at intercellular junctions, and may indicate further complexities in the control of leukocyte/ endothelial interactions. PMID- 10698321 TI - Structural differences among monoclonal antibodies with distinct fine specificities and kinetic properties. AB - The mAbs HyHEL-8, HyHEL-26 (HH8, and HH26, respectively) recognize epitopes on hen egg-white lysozyme (HEL) highly overlapping with the structurally defined HH10 epitope, while the structurally related XRPC-25 is specific for DNP and does not bind HEL. All four Abs appear to use the same Vk23 germ line gene, and all but HH8 use the same VH36-60 germ line gene. Of the three anti-HEL Abs, the sequences of HH26 variable regions are closest to those encoded by the respective germ line sequences. HH8 utilizes a different member of the VH36-60 gene family. Thus, the same Vk and VH genes, combined with somatically derived sequence differences, are used to recognize the unrelated Ags HEL and DNP. In contrast, different VH36-60 germ line genes are used to bind the same antigen (e.g. HH8 vs HH10 and HH26). While the affinities of HH10, HH8, and HH26 for HEL vary by less than 10-fold, their affinities for mutated Ag vary over several orders of magnitude. Analyses of Fab binding kinetics with natural species variants and site-directed mutants of lysozyme indicate that these cross-reactivity differences reflect the relative sensitivities of both the association and dissociation rates to antigenic mutation: HH8 has relatively mutation-insensitive association and dissociation rates, HH10 has a relatively mutation-sensitive association rate but more variable dissociation rates, and HH26 has variable association and dissociation rates. Only a few amino acid differences among the antibodies produce the observed differences in the robustness of the association and dissociation rates. Our results suggest that association and dissociation rates and mutation sensitivity of these rates may be independently modulated during antibody repertoire development. PMID- 10698322 TI - The paediatric clinic: disability and the family. PMID- 10698323 TI - Aetiology in severe and mild mental retardation: a population-based study of Norwegian children. AB - The aetiology of mental retardation (MR) was studied in a population-based series of Norwegian children derived from 30 037 children born between 1980 and 1985. The study included 178 children, 79 with severe MR (SMR) (IQ<50) and 99 with mild MR (MMR) (IQ 50 to 70). Aetiology was divided into two main groups: biopathological and unspecified. The biopathological group comprised 96% of SMR and 68% of MMR, and was subdivided into prenatal (70% and 51%), perinatal (4% and 5%), and postnatal damage (5% and 1%), and a group of undetermined timing of the damaging event (18% and 11%). Single-gene disorders accounted for 15 of the 63 children with genetic disorders, including X-linked recessive in six. During the course of the study, at least 27 (15%) children had their aetiological diagnosis revised. Gestational age <32 weeks, birthweight <1500 g, and Apgar scores 0 to 2 at 1 and 5 minutes implied a significantly increased risk of MR, but contributed to only 4% of the children in the study. Decreased birthweight (1500 to 2499 g) and Apgar scores 3 to 6 at 1 and 5 minutes showed increased probability of MR. Despite extensive investigations, 4% of SMR and 32% of MMR were not identified with any biological markers and were considered as unspecified MR, several most probably representing the lower end of the normal IQ distribution in the population. PMID- 10698325 TI - Kinaesthetic acuity in adolescent boys: a longitudinal study. AB - The Kinaesthetic Sensitivity Test (KST) was used to measure the development of kinaesthetic acuity in adolescent boys. Thirty boys were tested longitudinally, at intervals of 6 months, between the ages of 11 1/2 and 14 years. A second group of 20 boys was tested at the ages of 14 and 16 1/2 years. The findings were compared with existing normative data on 5- to 12-year-old children and young adults, and they indicated improvement in kinaesthetic acuity with age. Although the age effect is statistically significant only in the older group, confidence intervals show that the rate of improvement in both groups is comparable to improvement between the ages of 5 and 12 years. The reliability of the test is rather poor. The conclusion is that kinaesthetic development continues throughout adolescence. Further, development is quite robust and detectable even with a fairly unreliable measurement instrument. However, individual assessments should be interpreted with caution. PMID- 10698324 TI - Psychomotor development and minor anomalies in children exposed to antiepileptic drugs in utero: a prospective population-based study. AB - The aim of this study was to assess psychomotor development, using Griffiths' test, and the incidence of minor anomalies at birth in children who had been exposed to antiepileptic drugs (AEDs) in utero. The study sample comprised 100 children of mothers who were treated with AEDs during pregnancy and 100 matched control children. Women with epilepsy were recruited from a pregnant urban population (450 000 inhabitants). The lowest possible dose of the fewest AEDs to maintain seizure control was used. Drug levels were controlled on a monthly basis. The children were assessed at 9 months of age. The study children had a significant increase in the number of minor anomalies (31 compared with 18 control children; odds ratio 2.4, CI 1.15 to 5.02, P=0.02 McNemars test), and an increased number of facial anomalies after carbamazepine exposure (11 compared with six control children). Drug exposure did not influence the Griffiths' score at 9 months of age. Even a meticulous AED treatment strategy during pregnancy increases the number of minor anomalies. However, treatment with AEDs does not necessarily influence short-term psychomotor development. PMID- 10698326 TI - Children with neurological disorders do not always need fundoplication concomitant with percutaneous endoscopic gastrostomy. AB - Whether antireflux surgery should be routinely performed at the time of gastrostomy in children with neurological disorders is debatable because of the risk of gastroesophageal reflux. Some argue that these children should be screened for occult gastroesophageal reflux as this will determine the need for fundoplication. This study retrospectively examines outcome in 29 children with neurological disorders who underwent percutaneous endoscopic gastrostomy (PEG) without concomitant fundoplication. Children were included if they had no clinical evidence of severe gastroesophageal reflux before PEG insertion. The median age of children at PEG insertion was 5.6 years (range 1.1 to 18.0). The children were followed for a median of 2.6 years (range 0.4 to 4.9). Insertion of PEG was technically impossible in two children; and an asymptomatic gastrocolic fistula in another child led to subsequent tube removal. Fourteen of the 26 remaining children developed symptomatic gastroesophageal reflux after PEG; five of these showed no reflux on pH monitoring prePEG. Control of symptoms was achieved by medical intervention in 12, but two required fundoplication. Our findings indicate that in the child with neurological disabilities without symptoms indicating severe gastroesophageal reflux, fundoplication is unlikely to be necessary as a consequence of PEG insertion. We conclude that routine investigation for gastroesophageal reflux in the child without severe vomiting can be avoided and the number of antireflux procedures reduced. PMID- 10698327 TI - Visual function in school-aged children born before 29 weeks of gestation: a population-based study. AB - The aim of this study was to assess visual function, including visual perception, in a geographically-based population of school-aged children, with a median age of 7.2 years (range 5.1 to 9.3 years), born before 29 weeks of gestation to mothers living in Goteborg, Sweden. Fifty-one preterm children participated in the study, six of whom had known brain lesions. Visual acuity, visual fields, stereoacuity, and visual perception were tested. The Test of Visual Perceptual Skills Revised (TVPS-R, Gardner 1996) was used to measure visual perception, and the results were compared with those of 50 term (control) subjects. Six percent of the preterm children were visually impaired, with a visual acuity of less than 0.3 (6/18), while 42% of all the preterm children and 34% of those without known brain lesions had a total score below the 5th centile of the reference material for the test, compared with 14% of the control subjects. In conclusion, visual perceptual problems seem to be common among very preterm children and should be screened for and assessed before the children start school. PMID- 10698328 TI - Perceptions of mainstreaming: a systems approach. AB - This paper examines the perceptions of mainstreaming among six young people (mean age, 15.5 years; range 13 to 18 years) with physical disabilities, their parents, and seven teaching staff of one 'designated' school in the United Kingdom. The study used semistructured interviews to allow the participants to talk about their experiences of mainstreaming in their own words. The results were then analysed and formed the basis for a multiperspective case study. The analysis revealed that the processes involved that exclude the child with a physical disability from mainstream school life are highlighted. They appear to be subtle and also specific to individual children and families. PMID- 10698329 TI - Analgesic effects of botulinum toxin A: a randomized, placebo-controlled clinical trial. AB - Postoperative pain in children with spastic cerebral palsy (CP) is often attributed to muscle spasm and is difficult to manage using opiates and benzodiazepines. Adductor-release surgery to treat or prevent hip dislocation in children with spastic CP is frequently performed and is often accompanied by severe postoperative pain and spasm. A double-blinded, randomized, placebo controlled clinical trial of 16 patients (mean age 4.7 years) with a mainly spastic type of CP (either diplegic or quadriplegic in distribution) was used to test the hypothesis that a significant proportion of postoperative pain is secondary to muscle spasm and, therefore, might be reduced by a preoperative chemodenervation of the target surgical muscle by intramuscular injection of botulinum toxin A (BTX/A). Compared with the placebo, BTX/A was found to be associated with a reduction in mean pain scores of 74% (P<0.003), a reduction in mean analgesic requirements of approximately 50% (P<0.005), and a reduction in mean length of hospital admission of 33% (P<0.003). It was concluded that an important component of postoperative pain in the patient population is due to muscle spasm and this can be managed effectively by preoperative injection with BTX/A. These findings may have implications for the management of pain secondary to muscle spasm in other clinical settings. PMID- 10698330 TI - Foix-Chavany-Marie (anterior operculum) syndrome in childhood: a reappraisal of Worster-Drought syndrome. AB - Foix-Chavany-Marie syndrome (FCMS) is a distinct clinical picture of suprabulbar (pseudobulbar) palsy due to bilateral anterior opercular lesions. Symptoms include anarthria/severe dysarthria and loss of voluntary muscular functions of the face and tongue, and problems with mastication and swallowing with preservation of reflex and autonomic functions. FCMS may be congenital or acquired as well as persistent or intermittent. The aetiology is heterogeneous; vascular events in adulthood, nearly exclusively affecting adults who experience multiple subsequent strokes; CNS infections; bilateral dysgenesis of the perisylvian region; and epileptic disorders. Of the six cases reported here, three children had FCMS as the result of meningoencephalitis, two children had FCMS due to a congenital bilateral perisylvian syndrome, and one child had intermittent FCMS due to an atypical benign partial epilepsy with partial status epilepticus. The congenital dysgenetic type of FCMS and its functional epileptogenic variant share clinical and EEG features suggesting a common pathogenesis. Consequently, an increased vulnerability of the perisylvian region to adverse events in utero is discussed. In honour of Worster-Drought, who described the clinical entity in children 40 years ago, the term Worster-Drought syndrome is proposed for this unique disorder in children. PMID- 10698331 TI - How does the brain learn language? Insights from the study of children with and without language impairment. AB - Neurobiological studies have generated new ways of thinking about development of brain structure and function. Development involves more than just growth from simple to complex structures. The initial over-abundance of neurons and synaptic connections is subsequently pruned of those that are non-functional. In addition, as behavioural and cognitive functions emerge and become automatized, the underlying brain representations are reorganized. In this paper, I shall argue that these different modes of neurodevelopmental change provide a useful metaphor for examining language acquisition. It will be argued that language acquisition can involve learning to ignore and inhibit irrelevant information, as well as forming new ways of representing complex information economically. Modular organization is not present from the outset, but develops gradually. This analysis suggests a new way of assessing specific language impairment (SLI). There has been much debate as to whether children with SLI lack specific modular components of a language processing system. I propose instead that these children persist in using inefficient ways of representing language. Finally, I consider what we know about the neurobiological basis of such a deficit. There is mounting evidence that children with SLI have subtle structural anomalies affecting the language areas of the brain, which are largely genetically determined. We should not, however, conclude that the language difficulties are immutable. PMID- 10698332 TI - Serial magnetoencephalographic observations of a normally developing infant during sleep? PMID- 10698333 TI - An integrated map of the human regulator of complement activation (RCA) gene cluster on 1q32. PMID- 10698334 TI - FHL-1/reconectin and factor H: two human complement regulators which are encoded by the same gene are differently expressed and regulated. AB - FHL-1/reconectin and factor H are two human complement regulators which are encoded by a single gene. FHL-1/reconectin contains the first 7 of 20 SCR protein domains of factor H and has four unique residues attached to its C-terminal end. The overlapping region of 445 amino acids explains the related complement regulatory functions of the two proteins. However, unique biological functions have also been reported for FHL-1/reconectin, such as cell adhesion and binding to microbial surfaces. Both proteins are synthesised and secreted by the liver. Extrahepatic synthesis occurs in a wide variety of cells, e.g. in monocytes, fibroblasts or neuronal cells. Unexpectedly, FHL-1/reconectin and factor H exhibit distinct expression patterns. This is also observed in disease situations such as in rheumatoid arthritis or malignancies. In rheumatoid arthritis a potentially protective role is suggested by the local synthesis of both FHL 1/reconectin and factor H in synovial fibroblasts and their induction by the anti inflammatory agent dexamethasone and the cytokine IFN-gamma, but not by TNF alpha. FHL-1/reconectin is overexpressed in certain tumor cells such as glioblastoma, conferring an exceptional resistance to such cells against complement mediated lysis. Although FHL-1/reconectin and factor H are encoded by a single gene, regulated by the same gene promoter and initiate transcription at the same start site, their transcripts are differently regulated. The putative control levels, which are responsible for this complex regulation, include transcript elongation, RNA processing, alternative splicing and differential poly(A) site selection. PMID- 10698335 TI - The C3b/C4b receptor (CR1, CD35) on erythrocytes: methods for study of the polymorphisms. AB - This report is devoted to methodologies used in analyzing the C3b/C4b receptor (CR1, CD35) on erythrocytes (E), its soluble form, the CRI structural or allotype polymorphism, and CR1 density polymorphism. In primates E CR1 serves as the main system for processing and clearance of complement opsonized immune complexes (IC). CR1 copy numbers decrease with aging of E in normal individuals. Erythrocyte CR1 is also decreased in pathological conditions such as systemic lupus erythematosus (SLE), HIV infection, certain hemolytic anemias, and many other conditions featuring immune complexes. Consequently, CRI on E has an important physiological role in immune complex handling and has interesting alterations in disease. PMID- 10698336 TI - How are immune complexes bound to the primate erythrocyte complement receptor transferred to acceptor phagocytic cells? AB - Immune complexes (IC) bound to the primate erythrocyte (E) complement receptor (CR1) are cleared from the circulation of primates and localized to phagocytic cells in the liver and spleen without E destruction. IC can be bound to E CRI either via C3b opsonization or with cross-linked mAb complexes (heteropolymers, HP) which contain a mAb specific for CRI and a mAb specific for an antigen. The long-term goal of our work is to apply the HP system to the treatment of human diseases associated with blood-borne pathogens. This review discusses the mechanism by which the E-bound IC are transferred to acceptor cells. Our studies in animal models as well as our in vitro investigations indicate that IC transfer is rapid (usually >90% in 10 min) and does not lead to lysis or phagocytosis of the E. Experiments with specific inhibitors and the use of IC prepared with Fab' fragments suggest that transfer depends mainly upon recognition by Fc receptors on the acceptor cell. Moreover, we find that IC release from the E is associated with a concerted loss of CR1, and is followed by uptake and internalization of the IC by the acceptor cell. We suggest that recognition and binding of the E bound IC substrates by Fc receptors allows close contact between the E and acceptor cells, which in turn facilitates proteolysis of E CR1, presumably by a macrophage-associated protease. After proteolysis, the released IC are internalized by the macrophages. PMID- 10698337 TI - Assembly and regulation of the complement amplification loop in blood: the role of C3b-C3b-IgG complexes. AB - Amplification of complement activation in blood and serum starts on multi-protein complexes that act as precursors of an alternative C3 convertase. Among these covalently linked C4b-, C3b-, and IgG-containing complexes C3b-C3b-IgG complexes represent the major species containing C3b and IgG. Recent work on their purification and characterization is discussed. Special emphasis is placed on the arrangement of ester bonds in these complexes and their dual type of partial protection from inactivation. Partial protection from inactivation is mediated by properdin which binds to these complexes in the complete absence of any other complement protein. High dose IgG, known to stimulate inactivation of these complexes, appears to lower properdin binding in a process that also involves factor H. Properdin stimulates factor B binding to these complexes and renders them far better precursors of a C3 convertase than C3b. The available information allows a suggestion for a new scheme on how the amplification loop is assembled and regulated in blood and serum. PMID- 10698338 TI - The covalent interaction of C3 with IgG immune complexes. AB - Antigens (Ags) are converted into immune complexes (antigen-antibody complexes, IC) as soon as they encounter their specific antibodies (Abs). In fluids containing complement, the process of IC formation and fixation of complement components occur simultaneously. Hence, the formation of Ag-Ab-complement complexes is the normal way of eliminating Ags from a host. C3b-C3b-IgG covalent complexes are immediately formed on interaction of serum C3 with IgG-IC. These C3b-C3b dimers constitute the core for the assembly of C3/C5-convertase on the IC, which are subsequently converted into iC3b-iC3b-IgG by the complement regulators. These complexes are detected on SDS-PAGE by two bands of molecular composition, C3alpha65-C3alpha43 (band A) and C3alpha65-heavy chain of the Ab (band B), which correspond to C3b-C3b and C3b-IgG covalent interaction respectively, and that identify opsonized IC (C3b-IC). C3b can attach to Fab and Fc regions of the Ab molecule with similar efficiency. The presence of multiple C3b binding regions on IgG is considered an advantageous characteristic that facilitates the elimination of Ags in the form of C3b(n)-IC. Ab molecules on the IC recognize the Ag, and also serve as a very good acceptor for C3b binding. In this way, Ags, even if they have no acceptor sites for C3b, can be efficiently processed and removed. When C3 is activated in serum by IC or other activators, secondary C3b-IgG covalent complexes are generated, with bystander monomeric circulating IgG, and thus constitute, physiological products of complement activation. These complexes gain importance when IgG concentration is extremely high as in cases of infusion of intravenous IgG (IVIG) in several pathologies. The covalent attachment of activated complement C3 (C3b, iC3b, C3 d,g) to Ags or IC links innate and adaptative immunity by targeting Ags to different cells of the immune system (follicular dendritic cells, phagocytes, B cells). Hence C3b marks Ags definitively, from the earliest contact with the innate immune system until their complete elimination from the host. PMID- 10698339 TI - Activities of the MBL-associated serine proteases (MASPs) and their regulation by natural inhibitors. AB - There has been rapid progress in determining the mechanism by which complement is activated by the complex formed between Mannose-Binding Lectin and its associated proteases (MASPs). MBL and the MASPs are of low abundance, but are similar to the more abundant C1q-C1r2s2 complex (C1), which has been extensively investigated. In this review we summarise recent findings on MBL-MASPs' structure. enzymic activity and regulation, and compare MBL-MASPs with C1. PMID- 10698340 TI - Properdin deficiency: molecular basis and disease association. PMID- 10698341 TI - A critical evaluation of the putative role of C3adesArg (ASP) in lipid metabolism and hyperapobetalipoproteinemia. AB - The acylation stimulating protein, ASP is a small, basic serum protein capable of stimulating triglyceride synthesis in cultured fibroblasts and adipocytes. Sequence analysis of ASP has shown that ASP is identical to C3adesArg the inactive fragment of the complement anaphylatoxin peptide, C3a. It has been proposed that C3adesArg (ASP) can be generated by mature adipocytes secreting the three complement proteins: complement protein C3, factor B and factor D (adipsin). There have also been indications that adipocytes may express a specific C3adesArg (ASP)-receptor that is distinct from the recently cloned C3a receptor. This suggests that C3adesArg (ASP) acts as an adipocyte autocrine and that it plays a central role in the metabolism of adipose tissue. Based on these observations a hypothesis for the etiology of hyperapobetalipoproteinemia (hyperapoB) has been proposed. Hyperapobetalipoproteinemia (hyperapoB), is a familial lipoprotein disorder characterized by increased hepatic secretion of very low density lipoprotein (VLDL) and low density lipoprotein (LDL) particles. If C3adesArg (ASP) function in the adipose tissue is impaired, a reduced rate of triglyceride synthesis will follow, generating an increased flux of fatty acids to the liver. In response to an increased flow of fatty acids, the liver will increase its production of VLDL particles yielding the phenotype of hyperapoB. This review critically assesses this hypothesis and the potential role of C3adesArg (ASP) as a major determinant for triglyceride synthesis in the light of data collected in vitro and in vivo. PMID- 10698342 TI - Expression of the anaphylatoxin C5a receptor in non-myeloid cells. AB - C5, a 74 amino acid peptide cleaved from the complement protein C5, represents the most potent anaphylatoxin and possesses inflammatory as well as immunomodulatory activities. In the past, expression of the receptor for the anaphylatoxin C5a (C5aR) has been thought to be restricted to cells of myeloid origin. However, recent evidence suggests that the C5aR is constitutively expressed in non-myeloid cells including epithelial, endothelial and smooth muscle cells in the human liver and lung. These findings are contrasted by results from our laboratory which demonstrated that in the normal human liver and lung C5aR expression is detectable exclusively in macrophages and macrophage derived cells (Kupffer cells). Interestingly, we found evidence that C5aR expression may be inducible in epithelial cells as C5aR mRNA was observed in vivo in human keratinocytes of the inflamed but not of the normal skin. Herein we review the work of our laboratory and others on the expression of the C5aR in various human non-myeloid cells types. A better understanding of the expression patterns of this important anaphylatoxin receptor may provide new insights in the pathophysiological role of C5a in vivo. PMID- 10698343 TI - Neutrophil priming by cytokines and vitamin D binding protein (Gc-globulin): impact on C5a-mediated chemotaxis, degranulation and respiratory burst. AB - At the site of acute inflammation, leukocytes are confronted with multiple mediators which are expected to modulate each other with respect to cell responses to the individual ligand. Previous contact of neutrophils with pro inflammatory cytokines, such as TNF-alpha or GM-CSF, or with the vitamin D binding protein (Gc-globulin) leads to the alteration of either multiple or rather distinct C5a-mediated neutrophil functions. Gc-globulin, the transport protein for 25-(OH)-D3, serves selectively as a cochemotactic factor for C5a/Ca(des)Arg. In contrast, TNF-alpha and GM-CSF, previously shown to modulate FMLP-induced neutrophil responses, are able to reduce C5a-mediated neutrophil chemotaxis, but augment their degranulation and respiratory burst activity. Cytokine priming was shown to be accompanied by a down-regulation of C5a receptors (CD88) whereas vitamin D binding protein had no impact on the level of neutrophil C5a receptors. C5a itself diminishes chemotaxis as well as degranulation and oxidative burst in response to a second dose of the same ligand (homologous desensitization). A similar effect, termed heterologous desensitization, occurs, if cell responses to a given mediator (e.g. to C5a) are reduced or even abolished upon the activation of another receptor of the same G protein coupled chemoattractant receptor subfamily (e.g. receptors for FMLP or IL 8). In concert with C5a, certain molecules may either augment chemotaxis or shift neutrophil effector functions from migration to exocytosis, an essential step within the sequence of events in a coordinated inflammatory response. PMID- 10698344 TI - On the role of complement and Fc gamma-receptors in the Arthus reaction. AB - The contribution of either the complement system or the activation of Fc receptors for IgG (FcyRs) to the inflammatory response in immune complex (IC) disease is puzzling. A series of studies has been performed in mice with engineered deficiencies of either FcgammaRs, the complement components C3, C4 or the C5a receptor. In addition, different C5-deficient mice strains have been evaluated. Mice with gene targeted disruption of the gamma-subunit, which mediates surface expression and signal transduction of the high affinity Fc receptor type I for IgG (FcgammaRI), the low affinity receptor Fc receptor type III for IgG (FcgammaRIII) and the high affinity receptor type I for IgE (IgepsilonRI), showed an impaired inflammatory response in the reverse passive Arthus reaction in skin, peritoneum and lung. These data suggest, that the activation of FgammaRs is the initial event triggering the inflammatory cascade in IC disease. On the other hand, C5aR deficient mice are either protected from tissue injury induced by ICs, as in the lung, or the degree of the inflammatory response is markedly attenuated, as in peritoneum and skin. A detailed analysis of data obtained with the different knock-out strains revealed that both the activation of the complement system as well as the activation of different effector cells via FcgammaRs contribute to the inflammatory sequelae leading to tissue destruction in IC disease. The relative contributions of FcgammaRI or FcgammaRIII and the main effector cells through which these receptors mediate their effector functions are tissue dependent. The activation of the C5a receptor pathway appears to be the prominent contribution of the complement system. PMID- 10698345 TI - Complement activation and inhibition in experimental models of arthritis. AB - Complement activation has been implicated as a pathological process in a number of inflammatory and autoimmune disorders including chronic rheumatoid arthritis (RA). Animal models of experimental arthritis have been widely used to investigate the pathogenesis of RA and also in the development of novel therapies. Many of these models are complement-dependent and both incidence and progression of disease can be influenced by complement inhibition. In certain situations, local inhibition is of greater therapeutic benefit than systemic decomplementation. An increasing awareness and availability of a wide range of naturally occurring complement regulatory proteins can now offer a more targeted approach to complement inhibition while the availability of novel engineering strategies has also improved the efficiency of this process. The success of complement inhibition in the experimental models described should offer a novel therapeutic approach to the treatment of human inflammatory arthritis. PMID- 10698346 TI - The contrasting mechanisms of serum resistance of Neisseria gonorrhoeae and group B Neisseria meningitidis. AB - Neisseria gonorrhoeae and Neisseria meningitidis have evolved intricate mechanisms to evade complement-mediated killing. Sialylation of gonococcal lipooligosaccharide (LOS) results in conversion of previously serum sensitive strains to unstable serum resistance, which is mediated by factor H binding. Porin (Por) is also instrumental in mediating stable serum resistance in gonococci. The 5th loop of certain gonococcal PorlAs binds factor H, which efficiently inactivates C3b to iC3b. Factor H glycan residues may be essential for factor H binding to certain Por1A strains. Por1A strains can also regulate the classical pathway by binding to C4b-binding protein (C4bp) probably via the 1st loop of the Por molecule. Certain serum resistant Por1 B strains can also regulate complement by binding C4bp through a loop other than loop 1. Purified C4b can inhibit binding of C4bp to Por 1B, but not Por1A, suggesting different binding sites on C4bp for the two Por types. Unlike serum resistant gonococci, resistant meningococci have abundant C3b on their surface, which is only partially processed to iC3b. The main mechanism of complement evasion by group B meningococci is inhibition of membrane attack complex (MAC) insertion by their polysaccharide capsule. LOS structure may act in concert with capsule to prevent MAC insertion. Meningococcal strains with Class 3 Por preferentially bind factor H, suggesting Class 3 Por acts as a receptor for factor H. PMID- 10698347 TI - Complement resistance of tumor cells: basal and induced mechanisms. AB - Clinical and experimental studies have suggested that complement may play a role in tumor cytotoxicity. However, the efficiency of complement-mediated tumor cell lysis is hampered by various protective mechanisms, which may be divided into two categories: basal and induced mechanisms. The basal mechanisms are spontaneously expressed in cells without a need for prior activation, whereas the induced mechanisms develop in cells subjected to stimulation with cytokines, hormones, drugs or with sublytic doses of complement and other pore-formers. Membrane associated complement regulatory proteins, such as CD55 (DAF, Decay-Accelerating Factor), CD46 (MCP, Membrane Cofactor Protein), CD35 (CR1, Complement Receptor type 1) and CD59, which serve as an important mechanism of self protection and render autologous cells insensitive to the action of complement. appear to be over-expressed on certain tumors. Furthermore, tumor cells secrete several soluble complement inhibitors. Tumor cells may also express proteases that degrade complement proteins, such as C3, or ecto-protein kinases which can phosphorylate complement components, such as C9. Besides this basal resistance, nucleated cells resist, to some extent, complement damage by removing the membrane attack complexes (MAC) from their surface. Several biochemical pathways, including protein phosphorylation, activation of G-proteins and turnover of phosphoinositides have been implicated in resistance to complement. Calcium ion influx and activation of protein kinase C (PKC) and of mitogen-activated protein kinase (MAPK) have also been demonstrated to be associated with the complement induced enhanced resistance to lysis. The complete elucidation of the molecular mechanisms involved in basal and induced tumor cell resistance will enable the development of strategies for interfering with these evasion mechanisms and the use of the cytotoxic complement system against tumor cells. PMID- 10698348 TI - Complement activation following oxidative stress. AB - It is clear that complement plays an important role in the inflammatory process following oxidative stress in cellular and animal models. Clinical trials underway with novel complement inhibitors will establish the potential therapeutic benefit of complement inhibition in human disease. For as much as we understand about the role of complement in disease states, many questions remain. How is complement activated on endothelial cells following oxidative stress? What is the ligand for MBL on endothelial cells following oxidative stress? Will inhibition of MBL provide tissue protection to the extent observed with other complement inhibitors such as sCR1 or anti-C5 mAbs? These questions and more will undoubtedly be answered in the next millennium. PMID- 10698350 TI - Strategies for targeting complement inhibitors in ischaemia/reperfusion injury. AB - A transplanted organ suffers inherently from an ischaemic insult and subsequent reperfusion injury. The severity of such early events is thought to influence the success of the transplant procedure, not only in the immediate post-transplant period, but also to predispose the graft to both acute and chronic rejection. In this paper, we review the influence of the complement system upon ischaemia,reperfusion injury. The recognition of the involvement of complement has led to novel strategies to try to modulate ischaemia/reperfusion injury, some of which we have summarized. Finally, we note our own strategy to target complement inhibition in ischaemic tissues. PMID- 10698349 TI - Complement activation and atherosclerosis. AB - Atherosclerosis is an inflammatory disease mediated through the action of monocyte/macrophages, complement and T-lymphocytes. C5a and monocyte chemotactic factor released during complement activation in the arterial wall may participate in the initial monocyte recruitment. Assembly of C5b-9 on cells of the arterial wall may also induce cell lysis. On the other hand, sublytic assembly of C5b-9 on smooth muscle cells (SMC) and endothelial cells (EC) induces cell activation and proliferation. Analysis of mitogen activated protein kinases (MAPK) pathways induced by C5b-9 in aortic SMC revealed that extracellular signal regulated kinase (ERK) 1, c-jun NH2-terminal kinase (JNK) 1, and p38 MAPK are all activated by C5b-9. ERK1 activity was inhibited by wortmannin suggesting that ERK1 pathway is activated through phosphatidyl inositol -3 (PI 3-) kinase. Sublytic C5b-9 assembly on the plasma membrane was also able to activate Janus kinase (JAK) 1, signal transducer and activator (STAT) 3 and STAT4 in EC. JAK1 but not STAT3 activation induced by C5b-9 is dependent on Gi protein activation. New evidence accumulated during the last decade support the role of complement activation in both initiation and progression of the atherosclerotic lesions. Complement system activation is a major component of the chronic inflammatory process associated with atherosclerosis. PMID- 10698351 TI - The role of complement in transplantation. AB - The complement system contributes critically to the barrier to transplantation of cells and organs. In the case of tissues and organs transplanted between individuals of the same species, that is in allotransplantation, the barrier posed by complement is seemingly eclipsed by the barrier posed by cellular immune responses. In the case of cells and organs transplanted between individuals of disparate species, that is xenotransplantation, the complement system has been thought to pose a nearly insurmountable barrier. With our understanding on how the complement system contributes to rejection, it is now clear that the complement system is more important in allotransplantation and more forgiving in xenotransplantation than was previously thought. PMID- 10698352 TI - Estrogenic/antiestrogenic and scavenging properties of (E)- and (Z)-resveratrol. AB - Resveratrol, natural compound found in grapes and wine, has been reported to have a variety of health benefit properties. Based on the structural similarity to the synthetic estrogen diethylstilbestrol, we investigated estrogenic/antiestrogenic effects on human breast cancer cell lines, MCF-7 and MVLN, and scavenging properties using DPPH of both (E)- and (Z)-isomers. Both isomers increased the in vitro growth of MCF-7 cell lines at medium concentrations (10 and 25 microM) whereas the low concentrations (0.1 and 1 microM) had no effect and the high concentration (50 microM) decreased the cell growth and was cytotoxic. The 25 microM (E)-isomer alone was able to reduced the proliferation induced by the estradiol. Low concentrations of (E)- and (Z)-resveratrol (0.1 and 1 microM) and medium concentration 10 microM (Z)-resveratrol did not interfere with the estrogen receptor. In contrast, medium concentrations of (E)-resveratrol (10 and 25 microM) and (Z)-resveratrol (25 microM) functioned as superagonists of estradiol. Whatever the model used, MCF-7 or MVLN cell lines, (Z)-resveratrol was less effective than (E)-resveratrol. Extinction of DPPH and Fe(III) reduction experiments showed that both isomers of resveratrol could act as free radicals scavengers or pro-oxidant compounds. The properties of low concentrations of resveratrol raise the possibility that structure-function studies could lead to the development of more selective estrogen receptor agonists and antagonists, which could be useful as a therapeutic agent. PMID- 10698353 TI - Effects of the radical scavenger AVS on behavioral and BBB changes after experimental subarachnoid hemorrhage. AB - Free radicals are important contributors to the global brain dysfunction that follows subarachnoid hemorrhage (SAH). We evaluated the effects of hydroxyl radical scavenger AVS [(+/-)-N,N'-propylenedinicotinamide; Nicaraven] after experimental SAH on rodent behavioral deficits (employing a battery of well characterized assessment tasks over a 2-day observation period) and blood-brain barrier (BBB) permeability changes two days after SAH (quantifying the microvascular alterations according to the extravasation of protein-bound Evans Blue using a spectrophotofluorimetric technique) in dose-response and time-window experiments. Groups of 10 rats were injected with 400 microl of autologous blood into the cisterna magna, and followed by intravenous continuous infusion of saline or 0.1, 03 or 1 mg/kg/min of AVS beginning within 5 minutes or 6 or 12 hours after SAH. The results were compared with sham-operated saline-treated and with SAH saline-treated animals. AVS significantly ameliorated performances on Beam Balance (p < 0.01) and decreased BBB permeability changes in frontal, temporal, parietal, occipital and cerebellar cortices and subcortical and cerebellar nuclei and brainstem (p < 0.01), but did not significantly affect changes in Beam Walking. This study demonstrates the neuroprotective effects of AVS when administered after experimental SAH in rats. These effects were dose dependent and, moreover, were evident within the therapeutic window of 6-12 hours after SAH. These results reinforce the concept of a participation of reactive oxygen intermediates in the cerebral dysfunction following SAH. PMID- 10698354 TI - Antinociceptive properties of the methanolic extract and two triterpenes isolated from Epidendrum Mosenii stems (Orchidaceae). AB - The antinociceptive effect of the methanolic extract (ME) and two triterpenes isolated from E. mosenii (Orchidaceae) has been investigated in chemical and thermal models of nociception in mice. The ME of E. mosenii (0.3-30 mg kg(-1), i.p. or 50-400 mg kg(-1), p.o.) produced dose-related, significant and long lasting (4 to 6 h) inhibition of acetic acid-induced abdominal constriction, with ID50 values of 3.9 and 137.0 mg kg(-1), respectively. Pholidotin and 24 methylenecycloartenol isolated from E. mosenii (0.1-3.0 mg kg(-1), i.p.) also produced marked and dose-related inhibition of acetic acid-induced pain, with ID50 values of 0.9 and 1.1 mg kg(-1). However, these compounds and the ME were about 3- to 13-fold more potent at the level of ID50 than diclofenac when assessed in acetic acid-induced abdominal constriction. The ME of E. mosenii in the same range of doses produced dose-related inhibition of both phases of formalin-induced licking, with mean ID50 values for the first and the second phases of 0.9, 122.0 mg kg(-1) and 0.7, 258.0 mg kg(-1), respectively by i.p. or p.o. routes. In addition, the ME (0.3-30 mg kg(-1), i.p., or 50-400 mg kg(-1), p.o.) also caused dose-related inhibition of capsaicin-induced neurogenic pain with mean ID50 values of 5.2 and 130.0 mg kg(-1), respectively. Treatment of animals with naloxone (5 mg kg(-1), i.p.) completely reversed the antinociceptive effect caused by morphine (5 mg kg(-1), s.c.) and that caused by ME of E. mosenii (1 mg kg(-1), i.p.) when assessed against either phase of the formalin-induced pain. Furthermore, when assessed in the hot-plate test, ME (100 mg kg(-1), i.p.) and morphine (10 mg kg(-1), s.c.) caused significant increase in response latency. However, ME given daily for to 7 consecutive days did not develop tolerance to itself nor did it induce cross-tolerance to morphine. Taken together these data demonstrate that the ME of E. mosenii elicited pronounced antinociception, when assessed by i.p. or p.o. routes, against several models of pain. Its actions involve, at least in part, an interaction with opioid system, seeming no to be related with a non-specific peripheral or central depressant actions. Finally, the active principle(s) responsible for the antinociceptive action of E. mosenii is likely related to the presence of the triterpenes. PMID- 10698355 TI - Effects of U-50,488H and U-50,488H withdrawal on catecholaminergic neurons of the rat hypothalamus. AB - Previous report from our laboratory showed that morphine produces a stimulatory effect of hypothalamic noradrenaline (NA) turnover concurrently with enhanced pituitary-adrenal response after its acute injection and during withdrawal. In the present work we have studied the effects of acute and chronic administration of the kappa agonist U-50,488H as well as the influence of U-50,488H withdrawal on the activity of hypothalamic NA and dopamine (DA) neurons and on the activity of hypothalamic-pituitary-adrenal (HPA) axis. A single dose of U-50,488H (15 mg/kg i.p.) significantly increased hypothalamic NA and decreased DA turnover at the time of an enhanced corticosterone release. Rats rendered tolerant to the kappa agonist by administration of U-50,488H twice a day for 4 days showed no changes in corticosterone secretion. Additionally, a decrease in both hypothalamic MHPG (the cerebral NA metabolite) production and NA turnover was observed, whereas DOPAC concentration and DA turnover were enhanced, which indicate the development of tolerance towards the neuronal and endocrine actions of U-50,488H. After naloxone (3 mg/kg s.c.) administration to U-50,488H-tolerant rats, we found neither behavioural signs of physical dependence nor changes in hypothalamic catecholaminergic neurotransmission. In addition, corticosterone secretion was not altered in U-50,488H withdrawn rats. Present data clearly indicate that tolerance develops towards the NA turnover accelerating and DA turnover decreasing effect of U-50,488H. Importantly and by contrast to mu agonists, present results demonstrate that U-50,488H withdrawal produce no changes in hypothalamic catecholamines turnover or in corticosterone release (an index of the hypothalamus-pituitary-adrenal activity), which indicate the absence of neuroendocrine dependence on the kappa agonist. As has been proposed, this would suggest that the mu and the kappa receptor be regulated through different cellular mechanisms, as kappa agonists have a lower proclivity to induce dependence. PMID- 10698356 TI - Charcoal suspension for tumor labelling modifies macrophage activity in mice. AB - We have previously developed a charcoal suspension for injection into human breast cancers in order to facilitate their location during surgery. We observed that charcoal particles were ingested by intra and peritumoral macrophages, some of which carried the particles at some distance from the injection site. We studied the influence of the formulation parameters of the charcoal suspension for intratumoral injection on in vitro and in vivo activation and in vivo mobilization of mouse peritoneal macrophages after intra-peritoneal injection of 2 mL of each preparation. The influence of the charcoal origin (peat vs wood), granulometry, suspension vehicle (water for parenteral injection, vs saline), concentration and excipients were studied. Micronized peat charcoal in water for injection at the highest studied concentration reduced macrophage activation in vitro and in vivo. However, macrophage mobilization was weaker than after thioglycolate injection and did not seem to be charcoal dose-dependent. The additives incorporated in the charcoal suspension led in vivo to increased peritoneal macrophage activation and mobilization (mannitol, and glucose), only increased activation (polysorbate 80 and pluronic F68) or mobilization (dextran 40, egg lecithin, and cabosil), or inhibited both activation and mobilization (cremophor EL). PMID- 10698357 TI - In vivo effects of chronic treatment with [MET5]-enkephalin on hematological values and natural killer cell activity in athymic mice. AB - The role of endogenous opioids in immunological mechanisms was examined by subjecting athymic (nu/nu) mice to chronic injections of the opioid agonist [Met5]-enkephalin (MET) or continuous opioid receptor blockade with naltrexone (NTX). After 8 days of treatment, neither excess peptide nor deprivation of opioids from receptors had any effect on body weight, spleen index (spleen to body weight ratio), total and differential white blood cell counts, and natural killer (NK) cell activity in peripheral blood or splenic lymphocytes. At 28 days, chronic treatment with MET or NTX had no effect on any of these parameters with the exception of an elevation from controls in NK cell activity in peripheral blood in mice receiving NTX, and subnormal NK cell activity related to splenic lymphocytes in the MET group. These results suggest that chronic exposure to an opioid agonist, or persistent opioid receptor blockade, have little influence on a variety of immunological properties in athymic mice, suggesting that native opioids such as MET do not play a marked role in defense mechanisms in the athymic mouse. PMID- 10698358 TI - Acetylcholinesterase activity in chronic renal failure. AB - Twenty healthy subjects and 39 Chronic Renal Failure patients (CRF-patients) maintained on chronic hemodialysis were used in this investigation to study the changes in acetylcholinesterase (AChE) activity of red blood cells (RBCs). The CRF-patients were all undergoing hemodialysis treatment. AChE activity from the CRF-patients was determined before and after dialysis. An additional objective was to study the effect of chronic renal failure on human red blood cell aging. Blood samples were drawn from controls and CRF-patients in tubes containing EDTA or sodium heparin as an anticoagulant. Red blood cells were purified to avoid interference with monocytes, reticulocytes and leukocytes. The purified RBCs were subfractionated into young (y) (1.08-1.09), mid (m) (1.09-1.11) and old (o) (1.11 1.12) percoll density (g/mL) fractions using a discontinous percoll gradient. The mean +/- SD AChE per gram hemoglobin (U/g Hgb) activities in whole blood (WB), purified human red blood cells (PRBCs), young human red blood cells (y-RBCs), mid age human red blood cells (m-RBCs) and old human red blood cells (o-RBCs) in CRF patients were 31.2+/-3.43, 29.3+/-3.26, 30.4+/-3.91, 25.1+/-5.25, 17.1+/-6.02 in females and 29.8+/-5.39, 28.8+/-5.29, 28.7+/-5.29, 23.7+/-5.39 and 16.0+/-5.60 in males. AChE activity from CRF-patients were higher than that found in the control subjects. The aging of human RBCs in both the controls and CRF-patients showed a progressive reduction in AChE activity. AChE activity of RBCs from female CRF patients were significantly higher (p < 0.05) than that of the female control subjects. The RBCs isolated from male CRF-patients showed a higher AChE activity than control males, but a significant difference was only observed with the mid age-cells. These studies further indicate that AChE activity remained insignificantly different in the various density based age subfractions of RBCs of both CRF-patients and controls. PMID- 10698359 TI - Prokinetic effect of black tea on gastrointestinal motility. AB - The gastrokinetic effects of hot water extract of black tea [Camellia sinensis, (L) O. Kuntze (Theaceae)] on gastrointestinal motility were studied both in vivo and in vitro. The extract significantly accelerated the gastrointestinal transit (GIT) in vivo in mice. These facilitatory effect was reduced after pretreatment with atropine, hemicholinium-3, morphine, indomethacin, McN-A-343 and L-arginine. In guinea pig ileum, the extract facilitated the peristaltic reflex in response to pressures in normal preparation. The black tea extract and L-NMMA (nitric oxide synthase inhibitor) significantly reduced the electrical field stimulated nonadrenergic, noncholinergic (NANC) relaxation of isolated rat fundal strips. The extract markedly enhanced the tonic ('hump') responses to transmural stimulation in longitudinal muscle of guinea pig ileum which was unaltered in the presence of atropine. These findings suggest a cholinergic involvement and a partial role of prostaglandin and nitric oxide in the mechanism of action of black tea extract on gastrointestinal motility. To determine the effective constituents in black tea responsible for this activity, the effect of black tea polyphenols on GIT were also studied. Thearubigin fraction (but not theaflavin) accelerated GIT significantly which suggests its involvement in the prokinetic effect of black tea. PMID- 10698360 TI - Effects of some isoxazolpyrimidine derivatives on nitric oxide and eicosanoid biosynthesis. AB - The inhibitory effect of some isoxazolpyrimidine derivatives on iNOS and COX-2 endotoxin induction in mouse peritoneal macrophages has been studied. Three of these compounds inhibited nitrite and PGE2 accumulation in a concentration dependent-manner at microM range. None of these active compounds affected iNOS, COX-2, COX-1 or PLA2 activities, although some reduced iNOS or COX-2 expression. Besides, no effect was observed on human neutrophil inflammatory responses (LTB4 biosynthesis and superoxide or elastase release). Active compounds were assayed by oral administration in the mouse air pouch model, where they inhibited nitrite accumulation without affecting PGE2 levels or leukocyte migration. PMID- 10698361 TI - Effect of St. John's wort (Hypericum perforatum) on cytochrome P-450 2D6 and 3A4 activity in healthy volunteers. AB - The effects of the herb St. John's wort (Hypericum perforatum), a purported antidepressant, on the activity of cytochrome P-450 (CYP) 2D6 and 3A4 was assessed in seven normal volunteers. Probe substrates dextromethorphan (2D6 activity) and alprazolam (3A4 activity) were administered orally with and without the co-administration of St. John's wort. Urinary concentrations of dextromethorphan and dextrorphan were quantified and dextromethorphan metabolic ratios (DMRs) determined. Plasma samples were collected (0-60 hrs) for alprazolam pharmacokinetic analysis sufficient to estimate tmax, Cmax, t 1/2, and AUC. Validated HPLC methods were used to quantify all compounds of interest. No statistically significant differences were found in any estimated pharmacokinetic parameter for alprazolam or DMRs. These results suggest that St. John's wort, when taken at recommended doses for depression, is unlikely to inhibit CYP 2D6 or CYP 3A4 activity. PMID- 10698362 TI - Evidence that gingko biloba extract does not inhibit MAO A and B in living human brain. AB - Extracts of Ginkgo biloba have been reported to reversibly inhibit both monoamine oxidase (MAO) A and B in rat brain in vitro leading to speculation that MAO inhibition may contribute to some of its central nervous system effects. Here we have used positron emission tomography (PET) to measure the effects of Ginkgo biloba on human brain MAO A and B in 10 subjects treated for 1 month with 120 mg/day of the Ginkgo biloba extract EGb 761, using [11C]clorgyline and [11C]L deprenyl-D2 to measure MAO A and B respectively. A three-compartment model was used to calculate the plasma to brain transfer constant K1 which is related to blood flow, and lambdak3, a model term which is a function of the concentration of catalytically active MAO molecules. Ginkgo biloba administration did not produce significant changes in brain MAO A or MAO B suggesting that mechanisms other than MAO inhibition need to be considered as mediating some of its CNS effects. PMID- 10698363 TI - Association between the functional polymorphism of catechol-O-methyltransferase gene and alcohol consumption among social drinkers. AB - BACKGROUND: A common functional genetic polymorphism in the catechol-O methyltransferase (COMT) gene (Val158 Met) results in 3- to 4-fold differences in COMT enzyme activity and dopamine inactivation rate. Previous studies have shown that type I alcoholism is more common among subjects with low activity COMT genotype (LL), compared with high activity (HH) or heterozygotic (LH) genotypes. METHODS: We studied alcohol consumption and the COMT genotype in middle-aged Finnish men (n 896), who represented an unselected ethnically homogenous population sample and reported using alcohol during the past year. Average alcohol use in pure ethanol (grams per week) was compared between subjects with LL genotype and subjects with LH or HH genotypes. RESULTS: Men with LL genotype (30% of all subjects) reported 27% higher weekly alcohol consumption compared with the two other genotype groups (p < 0.05). The difference remained statistically significant after a multivariate adjustment for sociodemographic factors and prior or existing diseases (p = 0.031). CONCLUSIONS: The results indicate that COMT polymorphism may contribute significantly to alcohol intake not only in alcoholics but also in a general male population. PMID- 10698364 TI - Differential effects of ethanol on insulin-like growth factor-I receptor signaling. AB - BACKGROUND: Activation of the insulin-like growth factor I receptor (IGF-IR) by its ligands IGF-I and IGF-II induces cell proliferation and protects against apoptosis. Ethanol inhibits IGF-IR tyrosine autophosphorylation, which subsequently interferes with the activation of key downstream signaling mediators including insulin-receptor substrate-1, phosphatidylinositol 3-kinase, and mitogen-activated protein (MAP) kinase. The ethanol-induced inhibition of IGF-IR signaling reduces mitogenesis and enhances apoptosis. In the current study, we demonstrate that the antiproliferative action of ethanol can be modulated by differential sensitivity of the autophosphorylation of the IGF-IR to ethanol. METHODS: A series of subclones was generated from 3T3 cells that express the human IGF-IR. RESULTS: There was considerable variability in the ability of ethanol to inhibit IGF-I-dependent IGF-IR tyrosine autophosphorylation and MAP kinase activation, despite equivalent IGF-IR expression. The IGF-IR was completely resistant to a high concentration of ethanol (150 mM) in several subclones. The sensitivity of IGF-IR autophosphorylation to ethanol correlated directly with the inhibition of IGF-I-mediated MAP kinase activation and cell proliferation. Resistant subclones exhibited features of the transformed phenotype including high MAP kinase activity, partial loss of contact inhibition, and the development of foci at confluency. The IGF-IR isolated from ethanol resistant cells was similarly resistant to ethanol in autophosphorylation reactions in vitro, whereas ethanol inhibited the autophosphorylation of IGF-IR obtained from sensitive cells. CONCLUSIONS: Our findings are the first to demonstrate the modulation of ethanol sensitivity of a tyrosine kinase receptor, and they provide a molecular basis for differential effects of ethanol on cell proliferation. PMID- 10698365 TI - Neuropsychological deficits in sober alcoholics: influences of chronicity and recent alcohol consumption. AB - BACKGROUND: The relationships between severity of neuropsychological (NP) deficits and quantity and duration of alcoholic drinking remain controversial. Eckardt et al. (1998) proposed that NP deficits can be observed only if chronicity of alcohol abuse equals or exceeds 10 years. In this study we tested the hypothesis of Eckardt et al. and reexamined the relationship of NP performance and alcohol consumption. METHODS: One hundred sixty-two alcoholics and 165 controls completed a NP test battery at least 3 weeks after the alcoholics attained sobriety. Chronicity varied from 4 to 9 years for 55 alcoholics and from 10 to 33 years for the remaining 107. RESULTS: Compared to controls, both groups of alcoholics were impaired on the Shipley Vocabulary and Abstraction tests and on two versions of the Digit Symbol test, but there was no difference between the two alcoholic groups on any measure. Regression analyses that controlled for age and education showed that chronicity predicted less than 0.5% of the variance on NP measures. By contrast, a measure of recent alcohol consumption, the Quantity-Frequency Index, contributed significantly (approximately 5% of the variance) to the prediction of alcoholics' NP performance. CONCLUSIONS: These data provide weak support for a dose effect relationship between degree of NP impairment and level of alcoholic drinking in the past 6 months but no evidence for an influence of chronicity. PMID- 10698366 TI - Involvement of nicotinic receptors in alcohol self-administration. AB - BACKGROUND: Alcohol and nicotine, in the form of tobacco, are commonly co-abused. Nicotinic receptors also have been implicated in alcohol action. We designed the present study to examine the possible involvement of nicotinic receptors in alcohol self-administration. METHODS AND RESULTS: Pretreatment with lower doses (0.1-0.4 mg/kg) of nicotine, administered acutely or chronically, did not affect alcohol consumption, whereas a higher dose (0.8 mg/kg) initially suppressed alcohol consumption but stimulated alcohol consumption on repeated treatment. We observed the same pattern of nicotine effects on alcohol self-administration using an operant procedure. A dose of 0.8 mg/kg of nicotine initially suppressed operant responding for alcohol. Such suppression of alcohol self-administration was more pronounced during the first 20 min of the 60 min operant session. Responding for alcohol in the nicotine treated group, however, was significantly increased above the saline treated group by the 5th day of treatment. Mecamylamine, a noncompetitive nicotinic receptor antagonist, reduced alcohol consumption, whereas dihydro-beta-erythroidine (DHbetaE), a competitive nicotinic receptor antagonist, did not modify alcohol consumption. CONCLUSIONS: The stimulation of alcohol intake induced by nicotine treatment and the suppression of alcohol intake induced by mecamylamine provide evidence for the involvement of nicotinic receptors in alcohol consumption and/or self-administration. The failure of DHbetaE to reduce alcohol consumption, however, suggests that ethanol nicotine interaction is mediated by other nicotinic receptor subtypes rather than alpha4beta2 receptor subtype, or that mecamylamine acts through a nonnicotinic mechanism. PMID- 10698367 TI - Neurocognitive functioning of adolescents: effects of protracted alcohol use. AB - BACKGROUND: The present study examined associations between alcohol involvement in early to middle adolescence and neuropsychological (NP) functioning. METHODS: Alcohol-dependent adolescents (n = 33) with over 100 lifetime alcohol episodes and without dependence on other substances were recruited from alcohol/drug abuse treatment facilities. Comparison (n = 24) adolescents had no histories of alcohol or drug problems and were matched to alcohol-dependent participants on age (15 to 16 years), gender, socioeconomic status, education, and family history of alcohol dependence. NP tests and psychosocial measures were administered to alcohol dependent participants following 3 weeks of detoxification. RESULTS: Alcohol dependent and comparison adolescents demonstrated significant differences on several NP scores. Protracted alcohol use was associated with poorer performance on verbal and nonverbal retention in the context of intact learning and recognition discriminability. Recent alcohol withdrawal among adolescents was associated with poor visuospatial functioning, whereas lifetime alcohol withdrawal was associated with poorer retrieval of verbal and nonverbal information. CONCLUSIONS: Deficits in retrieval of verbal and nonverbal information and in visuospatial functioning were evident in youths with histories of heavy drinking during early and middle adolescence. PMID- 10698368 TI - Carbohydrate-deficient transferrin in relation to the menopausal status of women. AB - BACKGROUND: Carbohydrate-deficient transferrin (CDT) has been reported as an excellent marker for male alcohol abuse. Little is known about its validity among women, in whom rather conflicting data concerning the efficiency of the CDT marker and its biochemical mechanism have been reported. Moreover, it is not clear why the reference ranges are different for women (0 to 26 Units per liter) and men (0 to 20 Units per liter). METHODS: In this population-based study, we examined the normal CDT values measured by CDTect in 331 healthy female teetotalers, randomly selected from a large cohort. They were divided into four groups: premenopausal women (n = 76), perimenopausal women (n = 86), postmenopausal women (n = 84), and users of estrogens/progestagens (n = 85). RESULTS: The mean of the CDT value in the premenopausal group (15.2 Units per liter) was significantly higher than the mean in the postmenopausal group (13.6 Units per liter; p < 0.016). In pre- and perimenopausal women, higher CDT levels were associated with the last period of menstruation; for women menstruating less than 1 month ago versus longer ago, the mean serum CDT value was 15.4 vs. 13.0 Units per liter (p < 0.01). CONCLUSIONS: The premenopausal state seems to increase serum levels of CDT, probably due to the amount and frequency of blood loss during the menstrual period, and should be considered when interpreting CDT values in women. PMID- 10698369 TI - Test characteristics of carbohydrate-deficient transferrin and gamma glutamyltransferase in alcohol-using perimenopausal women. AB - BACKGROUND: The application of biochemical markers to detect heavy alcohol use in women has shown disappointing results until now. We evaluated carbohydrate deficient transferrin (CDT) by the CDTect method and gamma-glutamyltransferase (GGT) in a large cohort of alcohol-using perimenopausal women studied primarily for osteoporosis. METHODS: CDT and GGT were measured in 431 women aged 46 to 54 years, who were selected from a large cohort (n = 8503) of pre-, peri-, and postmenopausal women. Their alcohol intake was known from questionnaires and face to-face interviews. Three groups were constructed for statistical analysis: those drinking on average less than 7 alcoholic drinks per week (n = 103), those drinking 7 to 34 per week (n = 280), and those drinking at least 35 per week (n = 48). RESULTS: The mean values of CDT and GGT of the three groups increased with an increasing alcohol intake, but there was a poor correlation between CDT and GGT in the complete study group (r = 0.3). The specificities of CDT and GGT were comparable, 83% and 78%, respectively. The sensitivities for CDT and GGT were 30% and 50%, respectively. A logistic regression model could assign, overall, 77% of the women correctly in relation to their alcohol intake: 43% of the women drinking at least 35 drinks per week and 92% of the women drinking less than 7 drinks per week. CONCLUSIONS: The test characteristics of both GGT and CDT are not good enough to be used as biochemical markers for detecting heavy alcohol use in women. The use of a logistic regression model offers an advantage, because both numeric values of CDT and GGT are taken into account instead of arbitrary cutoff values. PMID- 10698370 TI - The Alcohol Use Disorders Identification Test (AUDIT) and carbohydrate-deficient transferrin (CDT) in a routine workplace health examination. AB - BACKGROUND: Only a few studies on workplaces have examined the Alcohol Use Disorders Identification Test (AUDIT) or carbohydrate-deficient transferrin (CDT) as screening instruments for the early identification of elevated and risky levels of alcohol consumption. The purpose of this study was to compare the performances of AUDIT, CDT, and gamma-glutamyltransferase (GGT) in a routine health examination (alcohol screening) in the workplace. METHODS: The study, carried out over 16 months in a large workplace in the transport sector, was part of an on-going controlled study. Employees who came to the company health service for a routine health examination were offered the opportunity to undergo an alcohol screening and check their alcohol habits. RESULTS: Of the 570 subjects who participated, 105 (18.4%) screened positive according to AUDIT, CDT, or both. Only 7.6% of the persons who screened positive did so according to both instruments. If GGT had been included as a screening instrument, the proportion of positive results would have increased to 22.0%. If we had only used AUDIT in the screening process, the proportion of positives would have fallen by nearly half. CONCLUSIONS: The present findings suggest that AUDIT and CDT are complementary instruments for alcohol screening in a routine workplace health examination, and each has value for identifying a different segment of the risky drinking population. PMID- 10698371 TI - Evidence of a structural effect for alcohol outlet density: a multilevel analysis. AB - BACKGROUND: Ecological studies reveal that alcohol-related outcomes tend to occur in high alcohol outlet density neighborhoods. The ecological design of these studies limits the interpretation of the findings in terms of the level of the effect. The effect of alcohol outlet density could be related to greater individual access to alcohol, an individual level effect, or to the grouping of drinkers by neighborhood, a structural effect at the neighborhood level. METHODS: To differentiate between individual and neighborhood level possibilities, we conducted a multilevel study. Individual distance to the closest alcohol outlet was the individual level measure of the effect of alcohol outlet density, whereas the mean distance to the closest alcohol outlet for all individuals within a census tract was the neighborhood level measure for the effect of alcohol outlet density. We analyzed telephone surveys of 2604 telephone households within 24 census tracts stratified by poverty status and alcohol outlet density. Individual distance to alcohol outlets, age, sex, race/ethnicity, and level of education were entered as individual level covariates, and their corresponding aggregated means were entered as census tract level covariates (i.e., mean distance to outlets, mean age, percentage male, percentage Black, mean education). RESULTS: Analysis of variance revealed that 16.2% of the variance in drinking norms and 11.5% of the variance in alcohol consumption were accounted for at the census tract level. In multivariate hierarchical analysis, individual distance to the closest alcohol outlet was unrelated with drinking norms and alcohol consumption, whereas mean distance to the closest alcohol outlet demonstrated a negative relation with drinking norms (betae = -5.50+/-2.37) and with alcohol consumption (betae = -0.477+/-0.195); that is, the higher the mean distance to the closest alcohol outlet, the lower the mean drinking norms score and mean level of alcohol consumption. CONCLUSIONS: The findings suggest that the effect of alcohol outlet density on alcohol-related outcomes functions through an effect at the neighborhood level rather than at the individual level. Problem drinkers tend to be grouped in neighborhoods, an effect predicted by alcohol outlet density. PMID- 10698372 TI - Polyenylphosphatidylcholine protects against alcohol but not iron-induced oxidative stress in the liver. AB - BACKGROUND: We reported before that, in baboons, the alcohol-induced oxidative stress in the liver is associated with depletion of dilinoleoylphosphatidylcholine [the major component of polyenylphosphatidylcholine (PPC)] and that both can be corrected by the administration of PPC, but we did not determine whether this protection extended to iron-induced oxidative stress. METHODS: To compare the effects of PPC on alcohol- and iron-induced hepatic oxidative stress, 56 Sprague Dawley male rats were pair-fed nutritionally adequate liquid diets containing ethanol (36% of energy) or isocaloric carbohydrate and PPC (3 mg/ml) or safflower oil (2.73 mg/ml), with or without 5 mg/ml carbonyl iron for 2 months. Markers of oxidative stress (4-hydroxynonenal and reduced glutathione), antioxidants (vitamin E, ubiquinol-9, and ubiquinol-10), and phosphatidylcholine (PC) species were assessed by HPLC and/or gas chromatography/mass spectrometry. RESULTS: Alcohol feeding increased hepatic 4-hydroxynonenal 3-fold and decreased glutathione by 19%, ubiquinol-10 by 53%, and PC species containing arachidonate (palmitoyl- and stearoylarachidonoylphosphatidylcholines by 24% and 21%, respectively) and total phospholipids by 14%. PPC feeding prevented the rise of 4-hydroxynonenal, restored glutathione, and increased the hepatic content of dilinoleoylphosphatidylcholine and of some other PC carrying polyunsaturated fatty acids. Administration of iron alone increased hepatic iron, doubled 4 hydroxynonenal and glutathione, whereas it decreased vitamin E, ubiquinol-9, total phospholipids, and several polyunsaturated PC. Alcohol given with iron further exacerbated the hepatic oxidative stress, as documented by the increase of 4-hydroxynonenal and the decrease in glutathione and ubiquinols-10. PPC did not prevent this oxidative stress, although it increased hepatic glutathione. Hepatic dilinoleoylphosphatidylcholine content was comparable with and without dietary iron. CONCLUSIONS: PPC prevents the alcohol-induced oxidative stress but only in the absence of iron overload. PMID- 10698373 TI - Attenuation of alcohol-induced apoptosis of hepatocytes in rat livers by polyenylphosphatidylcholine (PPC). AB - BACKGROUND: Alcohol consumption increases apoptosis of hepatocytes. This effect appears to be mediated by the induction of hepatic cytochrome P-4502E1(CYP2E1) and its generation of free radicals, which results in an enhanced lipid peroxidation that initiates apoptosis. Because polyenylphosphatidylcholine (PPC), a soybean extract rich in polyunsaturated phosphatidylcholines, decreases the induction of ethanol-specific CYP2E1 and opposes oxidative stress, we hypothesized that PPC supplementation may attenuate hepatocyte apoptosis caused by ethanol ingestion. METHODS: Twenty-eight male Sprague Dawley rats were pair fed Lieber-DeCarli liquid diets containing 36% of energy as alcohol or an isocaloric amount of carbohydrate for 28 days. Half of the rats were given PPC (3 g/liter), whereas the other half received the same amount of linoleate (as safflower oil) and of choline as the bitartrate. An additional dose of alcohol (3 g/kg) was given intragastrically 90 min before the livers were removed. We assessed apoptosis in formalin-fixed, paraffin-embedded liver sections by using the TUNEL (terminal transferase dUTP nick end labeling) assay. Apoptotic hepatocytes were identified by positive TUNEL staining in conjunction with condensation of nucleoplasm or margination of chromatin. In each rat, 20,000 to 60,000 hepatocytes were counted by light microscopy by using Image-Pro Plus computer software, and the incidence of apoptosis was expressed as the percentage of total hepatocytes. RESULTS: Alcohol feeding resulted in a 4.5-fold increase in apoptosis of hepatocytes compared to pair-fed control rats; PPC supplementation decreased the alcohol-induced apoptosis to less than half. No difference in the incidence of apoptosis between the control and PPC-supplemented rats was found in the absence of alcohol. Apoptosis was distributed randomly in the liver lobules of the rats fed the control diet, whereas the alcohol-induced apoptosis was significantly increased in the perivenular area. PPC supplementation strikingly reduced this effect. CONCLUSIONS: PPC attenuates alcohol-induced apoptosis of hepatocytes; this effect may provide a mechanism for PPC's protection against liver injury, possibly in association with its antioxidative action via the down regulation of ethanol-mediated CYP2E1 induction. PMID- 10698374 TI - Maternal ethanol exposure is associated with decreased plasma zinc and increased fetal abnormalities in normal but not metallothionein-null mice. AB - BACKGROUND: Ethanol profoundly affects fetal development, and this is proposed to be due primarily to a transient fetal zinc (Zn) deficiency that arises from the binding of Zn by metallothionein (MT) in the maternal liver. Zn homeostasis and fetal outcome were investigated in normal (MT+/+) and metallothionein-null (MT-/ ) mice in response to ethanol exposure. METHODS/RESULTS: Mice were treated with saline or ethanol (0.015 m/g intraperitoneally at 0 and 4 hr) on day 8 of gestation (Gd8), and the degree of fetal dysmorphology was assessed on Gd18. The incidence of external abnormalities was significantly increased in offspring from MT+/+ dams exposed to ethanol, where 27.4% of fetuses were affected. MT-/- ethanol-, MT+/+ saline-, and MT-/- saline-treated dams had fetuses in which the frequencies of abnormalities were 2.2, 6.4, and 6.9%, respectively. To investigate Zn homeostasis, nonpregnant mice were killed at intervals over 16 hr after ethanol injection. Liver MT concentrations in MT+/+ mice were increased 20 fold by 16 hr, with a significant elevation evident by 4 hr, whereas liver Zn levels were also significantly increased by 2 hr and maintained for 16 hr. In parallel with these changes, plasma Zn concentrations in MT+/+ mice decreased by 65%, with minimum levels of 4.5+/-0.3 micromol/liter at 8 hr. Conversely, MT-/- mice exhibited increased plasma Zn concentrations, with peak values of 20.8+/-0.3 observed at 4 hr. CONCLUSION: These findings link the teratogenic effect of ethanol to the induction of maternal MT and the limitation of fetal Zn supply from the plasma. PMID- 10698375 TI - Acute effects of ethanol on kainate receptors in cultured hippocampal neurons. AB - BACKGROUND: Kainate receptors are a subclass of ionotropic glutamate receptors that regulate excitability and mediate synaptic transmission and plasticity in the hippocampus. The acute effects of ethanol on these receptors are not completely understood. METHODS: The acute effects of ethanol on pharmacologically isolated kainate receptor-mediated currents were studied in cultured hippocampal neurons obtained from neonatal rats. Whole-cell patch-clamp electrophysiological techniques were used for these studies. LY303070 (GYKI-53784), a potent AMPA (alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid) receptor-selective noncompetitive antagonist, was used to isolate kainate currents. RESULTS: Kainate receptor-mediated currents corresponded to 7% of the total non-N-methyl-D aspartate (non-NMDA) currents in these neurons and were reduced to 24% of control values in the presence of 15 microM lanthanum. These kainate receptor-mediated currents were significantly inhibited by ethanol concentrations of 50 mM or more. Under our recording conditions, ethanol inhibited non-NMDA receptor- and NMDA receptor-mediated currents to a similar extent as kainate receptor-mediated currents. Western blot analysis indicated that glutamate receptor-5 and -6/7 subunits, and kainic acid-2 subunits are expressed in these cultured hippocampal neurons. CONCLUSIONS: The present results suggest that kainate receptors are important targets for the actions of ethanol in the central nervous system. PMID- 10698376 TI - Parent ratings of behavior in children with heavy prenatal alcohol exposure and IQ-matched controls. AB - BACKGROUND: Behavioral disturbances are well documented in children with heavy prenatal alcohol exposure. However, the degree to which these disturbances are related to factors other than alcohol, such as general intellectual functioning or socioeconomic status, is not known. METHODS: Using the Child Behavior Checklist, parent-rated behaviors of children with histories of heavy prenatal alcohol exposure were compared with those of a control group matched by age, sex, socioeconomic status, ethnicity, and verbal IQ score. Using this same questionnaire, children with fetal alcohol syndrome were compared with children with heavy prenatal alcohol exposure that did not meet the criteria for fetal alcohol syndrome classification. RESULTS: Data were analyzed by multivariate analyses of covariance. In the comparison of children with and without a history of prenatal alcohol exposure, significant differences were found on the competence, problem, and summary scales (all p < 0.05). For the secondary comparison between the fetal alcohol syndrome and the heavy prenatal alcohol exposure groups, there were no significant differences on any of the scales (allp > 0.10). CONCLUSIONS: These results suggest that prenatal alcohol exposure results in the significant and profound impairment of parent-rated behaviors and that these deficits are not explained entirely by the presence or absence of facial dysmorphology, general intellectual functioning, or demographic factors. PMID- 10698377 TI - Adolescents are not adults: developmental considerations in alcohol users. AB - Much of the work in adolescent substance abuse assessment and treatment has been a direct transport from tools and modalities used in adult substance use populations. There was a consensus among symposium participants that developmental issues are important in assessment, evaluation, and treatment of adolescents with substance use disorders. These issues directly impact outcome at all levels. Presentations from the symposium may be helpful for conceptualizing the problems of adolescent substance use as well as formulating strategies for future research. Information from the symposium may be viewed as a springboard for future research and clinical intervention in adolescent substance abuse. PMID- 10698378 TI - Developmental perspectives on risk and vulnerability in alcoholic families. PMID- 10698379 TI - Cytoplasmic cytokines in the T cells of chronic alcoholics. PMID- 10698380 TI - Repeated ethanol withdrawal delays development of focal seizures in hippocampal kindling. PMID- 10698381 TI - What is inherited in the predisposition to alcoholism: new model or more muddle? PMID- 10698382 TI - Internal mammary artery spasm: is calcium the culprit? PMID- 10698383 TI - Effects of calcium chloride on grafted internal mammary artery flow after cardiopulmonary bypass. AB - OBJECTIVE: To examine the effects of calcium chloride (CaCl2) administration on blood flow through the grafted left internal mammary artery (IMA) after cardiopulmonary bypass (CPB). DESIGN: Single-arm prospective study. SETTING: University-affiliated hospital operating room. PARTICIPANTS: Twenty adult patients scheduled for coronary artery bypass graft surgery with IMA graft. INTERVENTIONS: IMA flow was measured noninvasively with a laser Doppler flow probe placed around the IMA, and measurements were recorded for 10 seconds and averaged. After separation from CPB under stable hemodynamics, baseline IMA flow was measured. CaCl2, 15 mg/kg, was administered intravenously over 1 minute. Blood pressure, left atrial pressure, heart rate, and IMA flow were then measured at 1, 5, and 10 minutes. Coronary perfusion pressure and IMA vascular resistance were calculated. MEASUREMENTS AND MAIN RESULTS: After CaCl2 administration, IMA blood flow significantly decreased from baseline at 1, 5, and 10 minutes (from 28+/-9 mL/min to 19+/-8 mL/min, 22+/-6 mL/min, and 25+/-4 mL/min), with gradual return toward baseline over time. Blood pressure, coronary perfusion pressure, and IMA vascular resistance significantly increased at 1 and 5 minutes after CaCl2. Left atrial pressure and heart rate remained unchanged. No systolic regional wall motion abnormalities were detected on transesophageal echocardiography. CONCLUSIONS: CaCl2, administered as a bolus dose after separation from CPB, transiently but significantly reduces IMA flow and can potentially trigger vasospasm, increasing the risk for myocardial ischemia or infarction in susceptible patients. Further studies are needed to determine whether this effect also occurs with nitrosodilators or phosphodiesterase inhibitors. PMID- 10698384 TI - Effects of milrinone versus epinephrine on grafted internal mammary artery flow after cardiopulmonary bypass. AB - OBJECTIVE: To compare changes on grafted internal mammary artery (IMA) flow after cardiopulmonary bypass in response to the administration of milrinone or epinephrine. DESIGN: Prospective and randomized. SETTING: University-affiliated hospital. PARTICIPANTS: Twenty consenting, adult patients undergoing CABG. INTERVENTIONS: Patients were randomized to receive either milrinone, 50 microg/kg, or epinephrine, 0.03 microg/kg/min, immediately after cardiopulmonary bypass. IMA flow was measured with a laser Doppler flow probe before and after the administration of either drug. MEASUREMENTS AND MAIN RESULTS: Baseline grafted IMA flow was similar for both groups (milrinone, 38+/-14 mL/min; epinephrine, 33+/-10 mL/min). In patients who received milrinone, flow increased by 24% to 50+/-17 mL/min, p<0.05; whereas with epinephrine, it remained essentially unchanged (33+/-10 v. 31+/-11 mL/min). CONCLUSIONS: This study confirms that the vasodilatory effect of milrinone on the IMA is also present after its anastomosis, whereas low-dose epinephrine exhibits neither beneficial nor adverse effects. It is suggested that in the absence of excessive vasodilation, milrinone should be considered as a first-line inotrope after coronary artery bypass graft surgery, to achieve an increase in contractility and IMA artery flow. PMID- 10698385 TI - A comparison of inhaled nitric oxide and milrinone for the treatment of pulmonary hypertension in adult cardiac surgery patients. AB - OBJECTIVE: To investigate the relative effects of milrinone and nitric oxide on pulmonary and systemic hemodynamic responses in cardiac surgery patients with a history of pulmonary hypertension. DESIGN: Prospective and randomized. SETTING: University hospital. PARTICIPANTS: Forty-five adult cardiac surgery patients. INTERVENTIONS: Cardiac surgery patients with pulmonary hypertension were randomly assigned to one of three study groups: Group 1 patients (n = 15) were treated with intravenous milrinone on separation from cardiopulmonary bypass, group 2 patients (n = 15) with 20 ppm of inhaled nitric oxide, and group 3 patients (n = 15) with 40 ppm of inhaled nitric oxide. Heart rate, right ventricular ejection fraction, and pulmonary vascular resistance were measured throughout the perioperative period at specific data points. MEASUREMENTS AND MAIN RESULTS: There were no significant differences in demographics, anesthesia, surgery, or baseline hemodynamics among the groups. The group receiving 40 ppm nitric oxide had a significantly higher (p<0.05) right ventricular ejection fraction on arrival in the intensive care unit (40% v. 30% for the milrinone group and 33% for the nitric oxide 20 ppm group). The milrinone group required significantly more phenylephrine in the intensive care unit (p<0.05). CONCLUSIONS: Treatment of pulmonary hypertension in adult cardiac surgery patients with inhaled nitric oxide compared with milrinone is associated with lower heart rates, higher right ventricular ejection fraction, and a lower requirement for treatment with vasopressor agents. PMID- 10698386 TI - Adenosine with cold blood cardioplegia during coronary revascularization. AB - OBJECTIVE: To investigate whether adenosine in association with blood cardioplegia results in more rapid cardiac arrest or improved myocardial protection. DESIGN: A prospective, randomized, placebo-controlled double-blind clinical study. SETTING: Operative and intensive care units in a university hospital, Finland. PARTICIPANTS: Forty patients undergoing primary, elective coronary revascularization. INTERVENTION: Adenosine as a bolus dose, 12 mg intravenously, was given immediately before the induction of blood cardioplegia. MEASUREMENTS AND MAIN RESULTS: There were nonsignificantly higher serial serum values of CK (MB) (p = 0.33), troponin-T (p = 0.23), and troponin-I (p = 0.10) in the adenosine group. There were no differences between the groups in arrest time, blood pressure decrease, or lactate extraction. CONCLUSIONS: The adenosine regimen used in this study did not cause more rapid arrest with blood cardioplegia. The effect on cardioprotection was insignificant. PMID- 10698387 TI - Adenosine for cardioplegic induction: a comparison with St Thomas solution. AB - OBJECTIVE: To determine if quicker cardiac standstill obtained by adding adenosine to potassium crystalloid cardioplegia translated into better myocardial preservation and cardiac function in the early postoperative period compared with the same cardioplegia without adenosine. DESIGN: A prospective study. SETTING: Cardiac center of a teaching institute. PARTICIPANTS: Sixty consecutive patients with left main vessel or triple-vessel disease undergoing coronary artery bypass surgery under moderate hypothermia. INTERVENTIONS: The study comprised two groups of patients. Group N (n = 15) was the control group, given St Thomas cardioplegic solution after aortic cross-clamping, without adenosine; whereas group A (n = 45) received 250 microg/kg of adenosine into the aortic root after aortic cross clamping, followed by the same St Thomas cardioplegia as in group N. The two groups were otherwise similar in all aspects of perfusion management. MEASUREMENTS AND MAIN RESULTS: Time taken to achieve cardiac standstill after aortic cross-clamping was significantly greater, 18.7+/-3.1 seconds, in the control group compared with the adenosine group, 3.4+/-0.9 seconds (p<0.001). The quicker arrest of the adenosine group led to better postoperative function, in the form of higher cardiac index (p<0.01), lower filling pressures (pulmonary artery wedge pressure) (p<0.05), and lower mean pulmonary artery pressure (p<0.05) at 6 hours. In the adenosine group, only 3 of 45 (6.6%) patients had elevated creatine phosphokinase (CPK) (MB) values greater than 50 U/L over preoperative CPK values compared with 3 of 15 (20%) in the control group (p<0.01). CONCLUSIONS: Injection of 250 microg/kg of adenosine into the aortic root before administration of cold crystalloid St Thomas cardioplegia solution after cross-clamping, in patients with severe coronary artery disease, produces significantly faster cardiac standstill, better myocardial preservation, and better cardiac function in the early postoperative period. PMID- 10698388 TI - Bypass flow, mean arterial pressure, and cerebral perfusion during cardiopulmonary bypass in dogs. AB - OBJECTIVE: To determine if normal cardiopulmonary bypass (CPB) pump flows maintain cerebral perfusion in the context of reduced mean arterial pressure at 33 degrees C. DESIGN: A prospective investigation. SETTING: Animal CPB research laboratory. PARTICIPANTS: Seven dogs that underwent CPB. INTERVENTIONS: Seven dogs underwent CPB at 33 degrees C using alpha-stat management and a halothane, fentanyl-midazolam anesthetic. Cerebral blood flow was measured using the sagittal sinus outflow technique. After control measurements at 70 mm Hg, cerebral physiologic values were determined under four conditions in random order: (1) mean arterial pressure of 60 mm Hg achieved by a reduction in pump flow, (2) mean arterial pressure of 60 mmHg determined by partial opening of a femoral arterial-to-venous reservoir shunt, (3) mean arterial pressure of 45 mm Hg by reduced pump flow, and (4) mean arterial pressure of 45 mm Hg by shunt. A 9F femoral arterial-to-venous reservoir shunt was controlled by a screw clamp. MEASUREMENTS AND MAIN RESULTS: Except for the controlled variables of mean arterial pressure and bypass flow, physiologic determinants of cerebral blood flow (temperature, PaCO2 and hematocrit) did not differ under any of the CPB conditions. Pump flow per se was not a determinant of cerebral perfusion. Cerebral blood flow and cerebral oxygen delivery did not differ with changes in pump flow if mean arterial pressure did not differ. Cerebral blood flow depended on mean arterial pressure under all pump flow conditions, however. CONCLUSIONS: Over the range of flows typical in adult CPB at 33 degrees C, pump flow does not have an effect on cerebral perfusion independent of its effect on mean arterial pressure. A targeted pump flow per se is not sufficient to maintain cerebral perfusion if mean arterial blood pressure is reduced. PMID- 10698391 TI - Respiratory jugular venodilation: a new landmark for right internal jugular vein puncture in ventilated patients. AB - OBJECTIVE: To report a new technique for right internal jugular vein puncture using respiratory jugular venodilation as a landmark for vein location. DESIGN: Prospective observational study. SETTING: Single community hospital. PARTICIPANTS: Two hundred patients undergoing right internal jugular vein cannulation under general anesthesia. INTERVENTIONS: Catheter placement was attempted using respiratory jugular venodilation as the primary landmark. When it was not applicable, an alternative technique using the carotid pulse as a landmark was used. MEASUREMENTS AND MAIN RESULTS: Visibility of the venodilation, the number of needle passes, the success rate, and the incidence of arterial puncture were analyzed. Respiratory jugular venodilation was observed in 158 patients (79%). In this group of patients, the jugular vein was cannulated at the first attempt in 83.5% of patients, and arterial puncture occurred in one patient (0.6%). In the remaining 42 patients (21%) lacking the visible venodilation, catheter placement was accomplished at the first attempt in 42.9% of patients (p<0.01 v. the venodilation-visible group), and 4 patients (9.5%) suffered arterial puncture (p<0.01). The overall incidence of arterial puncture was 2.5%. The success rate of cannulation (within four needle passes and no arterial puncture) was 98.1% in the venodilation-visible patients and 73.8% in the others (p<0.01), with the overall success rate of 93%. CONCLUSIONS: Respiratory jugular venodilation can be identified in a large proportion of ventilated patients. This experience suggests that respiratory jugular venodilation could be favorably used as the primary landmark for right internal jugular vein puncture in anesthetized patients. PMID- 10698390 TI - Total intravenous anesthesia with ketamine for pediatric interventional cardiac procedures. AB - OBJECTIVE: To evaluate the safety and efficacy of ketamine in pediatric patients undergoing interventional cardiac procedures. DESIGN: A retrospective clinical study. SETTING: A single, tertiary referral center. PARTICIPANTS: Patients (n = 107) undergoing interventional cardiac procedures between July 1996 and July 1998. INTERVENTIONS: Each patient received a bolus of ketamine, 1 mg/kg intravenously, followed by an infusion of 50 to 75 microg/kg/min for the duration of the procedure. MEASUREMENTS AND MAIN RESULTS: Hemodynamic and respiratory parameters were noted. All patients were breathing spontaneously. Average infusion dose of ketamine was 51.40+/-3.54 microg/kg/min (mean +/- standard deviation). Increases in heart rate and mean arterial pressure by more than 20% from baseline values were seen in 10 and 9 patients, respectively. Transient apnea and excessive salivation were seen in two patients each. Excessive movement of extremities was seen in six patients. There were no episodes of unpleasant dreams or hallucinations. There were two deaths (1.9%) related to the interventional procedures. CONCLUSION: The technique described is a simple, safe, and effective method for anesthetizing children in the cardiac catheterization laboratory for interventional procedures. PMID- 10698392 TI - Assessment of systematic use of intraoperative transesophageal echocardiography during cardiac surgery in adults: a prospective study of 203 patients. AB - OBJECTIVE: To determine the usefulness of systematic intraoperative transesophageal echocardiography in a cardiac surgical unit. DESIGN: Open prospective observational survey. SETTING: University Hospital. PARTICIPANTS: Consecutive adult patients (n = 203) undergoing elective or urgent cardiac operations. MEASUREMENTS AND MAIN RESULTS: Pre-cardiopulmonary bypass imaging yielded unsuspected findings in 26 patients (12.8%) and changed the planned surgery in 22 patients (10.8%). Transesophageal echocardiography modified the diagnosis in eight patients (17%) operated on for mitral valvulopathy, in seven patients (15.5%) with aortic valvular disease, in four patients (4.6%) with coronary artery disease, in five patients operated on for thoracic aorta diseases regardless of their localization (18.5%), and in two miscellaneous cases. On the basis of the data obtained from the transesophageal echocardiography carried out at the end of cardiopulmonary bypass, an immediate reintervention was required in five cases (2.5%). CONCLUSIONS: It is concluded that systematic intraoperative transesophageal echocardiography significantly affected decision making in this cardiac surgical unit. Its routine use in all cardiac surgical patients is recommended. PMID- 10698389 TI - Continuous infusion of remifentanil and target-controlled infusion of propofol for patients undergoing cardiac surgery: a new approach for scheduled early extubation. AB - OBJECTIVE: To assess hemodynamic stability, postoperative pain management, and the control and timing of early extubation of a total intravenous anesthetic technique using propofol target-controlled infusion (TCI) and remifentanil in cardiac surgery. DESIGN: Prospective study. SETTING: University hospital. PARTICIPANTS: Fifty patients scheduled for elective cardiac surgery. INTERVENTIONS: Premedication consisted of oral midazolam, 0.1 mg/kg. Anesthesia was induced with propofol TCI at a target concentration of 1.5 to 2 microg/mL; remifentanil, 1 microg/kg; and rocuronium. Anesthesia was maintained with propofol at the same target concentration and remifentanil titrated between 0.25 and 1 microg/kg/min. Thirty minutes before the end of surgery, a 0.1-mg/kg bolus of morphine was administered intravenously. Postoperative sedation was achieved by maintaining the propofol infusion until the patient was deemed ready for extubation. Postoperative pain relief was evaluated using a visual analog scale. The intervals between arrival in the intensive care unit, spontaneous ventilation, and extubation were recorded. MEASUREMENTS AND MAIN RESULTS: Included in this study were 36 men and 14 women (American Society of Anesthesiologist = III; New York Heart Association = II) scheduled for cardiac surgery. All patients remained hemodynamically stable throughout the perioperative period. Thirty-seven patients were successfully extubated during the first 4 postoperative hours. Spontaneous breathing was achieved at a mean interval of 15+/-5 minutes after propofol discontinuation. The mean interval to extubation was 163+/-45 minutes after arrival in the intensive care unit. Extubation was performed 48+/-12 minutes after patients were considered ready to awaken. During spontaneous ventilation, 36 patients received additional boluses of morphine (mean, 2.5+/-1 mg). Subsequently, all patients achieved a visual analog scale less than 40 mm. CONCLUSION: The combination of remifentanil and propofol TCI resulted in hemodynamic stability and good postoperative analgesia. This technique allows physicians to schedule the time of extubation in patients undergoing cardiac anesthesia. PMID- 10698394 TI - Hypoxemia from an atrial septal defect 7 days after blunt thoracic trauma. PMID- 10698393 TI - Renal effects of amino acid infusion in cardiac surgery. AB - OBJECTIVE: To evaluate effects of amino acids on renal function and oxygen consumption and the role of individual amino acids on renal blood flow (RBF) changes. DESIGN: Prospective, randomized, controlled study. SETTING: Operating room in cardiothoracic surgery department, university hospital. PARTICIPANTS: Twenty-two male patients submitted to elective first-time coronary artery bypass surgery. INTERVENTIONS: A catheter was placed in the left renal vein for thermodilution RBF measurements and blood sampling. In 11 patients, a balanced mixed amino acid infusion was infused (200 mL/hr) for 30 minutes immediately after the operation. MEASUREMENTS AND MAIN RESULTS: RBF and glomerular filtration rate increased during amino acid infusion compared with the control group. Renal oxygen consumption increased in the amino acid group and correlated with the increase in RBF (r = 0.70, p<0.001). Amino acid infusion induced two- to fourfold increases in plasma concentrations of individual amino acid concentrations and promoted renal extraction of aspartate, glutamate, glycine, and histidine. No correlation was observed between arterial concentration or uptake of individual amino acids and RBF. CONCLUSIONS: The increase in RBF from a mixed amino acid infusion was associated with increased glomerular filtration rate and renal consumption of oxygen. Changes in RBF of a mixed amino acid infusion could not be linked to plasma level or renal uptake of any individual amino acids. PMID- 10698395 TI - Low-dose surfactant instillation during extracorporeal membrane oxygenation therapy in a patient with adult respiratory distress syndrome and secondary atelectasis after chest contusion. PMID- 10698397 TI - Coronary artery bypass graft surgery in a patient with atypical Klippel-Trenaunay syndrome. PMID- 10698398 TI - Early postoperative bacteremia following cardiac surgery after recent surgery for colon cancer. PMID- 10698396 TI - Placement of an automatic implantable cardioverter-defibrillator in a 6-month-old infant: anesthetic management. PMID- 10698399 TI - The patient with cardiac trauma. PMID- 10698400 TI - Case 1--2000. Unilateral lung edema during anesthesia for reconstructive surgery of the trachea after caustic agent ingestion. PMID- 10698402 TI - Con: epiaortic scanning is not routinely necessary for cardiac surgery. PMID- 10698401 TI - Pro: epiaortic scanning is routinely necessary for cardiac surgery. PMID- 10698403 TI - An unusual cause of right-leg ischemia. PMID- 10698404 TI - Hypotension after coronary artery bypass surgery. PMID- 10698405 TI - Hemodynamic monitoring in patients undergoing off-pump coronary artery bypass graft surgery using the octopus tissue stabilizer: left atrial pressure as a gold standard. PMID- 10698406 TI - Sudden hypotension after cardiopulmonary bypass. PMID- 10698407 TI - Awareness during cardiac surgery. PMID- 10698408 TI - Blood levels of propofol during induction of anesthesia. PMID- 10698409 TI - The role of lung scintigraphy in the diagnosis of nephrotic syndrome with pulmonary embolism. AB - PURPOSE: Patients with nephrotic syndrome (NS) have an increased tendency to develop thrombosis and even to progress to pulmonary embolism (PE). This study was performed to determine the incidence of PE in NS with severe hypoalbuminemia and to investigate the possible role of ventilation-perfusion (V/Q) lung scans to evaluate these patients. METHODS: Eighty-nine patients with NS (serum albumin concentration < 2 g/dl) and risk factors for PE were studied. In all patients, the probability that PE would develop was assessed based on the results of V/Q lung scans (Xe-133 for ventilation and Tc-99m MAA for perfusion imaging). The lung scans were judged using the modified Prospective Investigation of Pulmonary Embolism Diagnosis criteria. In 25 (28%) patients whose lung scans showed an intermediate or low probability, but for whom there was a strong clinical indication of PE, pulmonary angiography was performed. The patients' clinical symptoms and signs on initial examination were observed. Additional examinations included electrocardiograms, chest radiography, and hematochemical tests such as albumin, blood urea nitrogen, creatinine, cholesterol, triglycerides, fibrinogen, antithrombin III, prothrombin time, and activated partial thromboplastin time. RESULTS: Based on the findings of lung scans, 19 (21%) of the patients were categorized as having a high probability of PE. However, pulmonary angiography found that 10 (11%) other patients had PE despite having lung scan findings categorized as intermediate or low probability of PE. Except for plasma fibrinogen and antithrombin III levels, neither the clinical symptoms and signs, electrocardiogram findings, chest radiograph results, nor values of hematochemical testing were consistent with the occurrence of PE in these 29 patients. CONCLUSION: The results of this study suggest that PE is not a rare complication in patients with NS, and is usually clinically silent. In this series, the occurrence of PE did not appear to be always correlated with the clinical or hematochemical severity of NS, except for the association with elevated levels of fibrinogen and antithrombin III. When treating the clinical symptoms of patients with NS, physicians should be alert to the possible complication of PE. Serial V/Q lung scans may provide valuable clues in the evaluation of these patients. PMID- 10698411 TI - Clinical production of pharmaceutical grade Tc-99m dextran 70 for lymphoscintigraphy. AB - Lymphoscintigraphy is an established means to determine the lymphatic drainage patterns from malignant tissues and edematous extremities. Unfortunately, there are no commercially available dedicated radiopharmaceuticals labeled for use in lymphoscintigraphy studies. The authors report a simple way to extemporaneously compound pharmaceutical-grade Tc-99m dextran 70 to meet this need. Pharmaceutical quality Tc-99m dextran 70 injection is prepared by a simple rapid method from drug intermediates that are marketed as parenteral drug products. The radiolabeling process yields a product of high radiochemical purity, with good in vitro and in vivo stability. The authors illustrate the use of this product in a patient with melanoma to show the lymphatic drainage pattern before surgery. The method described permits rapid compounding of Tc-99m dextran 70 injection from drug components that are intended for parenteral administration. Tc-99m dextran 70 provides the option of performing lymphoscintigraphy in any clinical nuclear medicine setting. PMID- 10698410 TI - Complementary nature of radiotracer parathyroid imaging and intraoperative parathyroid hormone assays in the surgical management of primary hyperparathyroid disease: case report and review. AB - PURPOSE: This article illustrates the complementary nature of preoperative radionuclide parathyroid imaging and intraoperative rapid parathyroid hormone (PTH) assays in primary hyperparathyroid disease. The authors review the literature on these procedures and compare this protocol and its cost effectiveness with those of the classic four-gland exploration. MATERIALS AND METHODS: Preoperative parathyroid imaging with Tc-99m MIBI and intraoperative rapid PTH assays were performed at the time of neck exploration. RESULTS: One of two parathyroid adenomas seen on radionuclide images would have been missed if the authors had relied solely on the initial decrease in PTH assay value to a normal level. CONCLUSIONS: Tc-99m MIBI imaging and intraoperative rapid PTH assays are complementary; when used together, they lessen the likelihood that abnormal parathyroid glands will be overlooked. This experience and that of others suggest these combined procedures are cost-effective. PMID- 10698412 TI - Floating gallbladder: a questionable prelude to torsion: a case report. AB - A 55-year-old woman had recurrent bouts of low substernal and epigastric pain radiating into the interscapular region. A hepatobiliary scan initially showed what was believed to be a dilated common bile duct and nonvisualization of the gallbladder. A delayed image obtained after having the patient move about revealed the presence of a filled gallbladder and normal common bile duct. The combination of recurrent pain with this scintigraphic picture may be representative of a floating gallbladder or an incomplete torsion with spontaneous detorsion. This case is presented to describe the scintigraphic appearance of a mobile gallbladder that may be prone to volvulus and to emphasize the importance of obtaining decubitus or oblique views at the end of a hepatobiliary study in selected cases of unusual findings. PMID- 10698413 TI - Unilateral acute renal cortical necrosis: correlative imaging. AB - Bilateral acute cortical necrosis is a rare form of acute renal failure characterized by necrosis of the renal cortex and sparing of the medulla. Little information on the imaging presentation of bilateral acute renal cortical necrosis is available. The enhanced CT appearance is pathognomonic and diagnostic. The unilateral presentation of acute cortical necrosis is extremely rare, and no imaging methods have been described. The authors chose to apply scintigraphic evaluation to this unique condition complementary to CT to confirm the diagnosis. Mercaptoacetylglycine (T3) was selected to assess tubular damage, in contrast to the pure glomerular agent DTPA. Evidence of some tubular function and clear delineation of the shrunken kidney was found. Conversely, in the DTPA study the kidney was not visualized. A DMSA scan was performed for assessment of viability of the renal cortex and showed a photopenic halo around the small area of the viable cortex of the upper pole. The halo sign represents a cortical loss. The visualization of the upper pole as evidence of cortical viability as a consequence of collateral blood flow from capsular vessels was seen on angiography. Radiographic and scintigraphic correlation of this rare condition may be an effective means to confirm the diagnosis and to establish the extent of involvement. However, contrast CT remains the preferred method in the diagnosis of acute cortical necrosis. PMID- 10698414 TI - Anterior choroidal artery infarction presenting as a progressive cognitive deficit. AB - PURPOSE: The authors describe a patient in whom neuroimaging using Tc-99m HMPAO SPECT, F-18 fluorodeoxyglucose (F-18 FDG) coincidence imaging, and magnetic resonance imaging (MRI) identified an anterior choroidal artery infarction. Neuroimaging played a critical role in confirming this diagnosis, because the patient had symptoms of progressive cognitive decline and satisfied the National Institute of Neurological and Communicative Disorders and Stroke-Alzheimer's Disease and Related Disorders Association criteria for Alzheimer's disease (AD). METHODS: Tc-99m HMPAO brain SPECT was performed using a triple-head gamma camera. F-18 FDG scanning was obtained 40 minutes after intravenous injection of 5 mCi F 18 FDG using a coincidence camera. A brain MRI scan was performed using a 1.5 Tesla scanner. RESULTS: Tc-99m HMPAO SPECT showed focal hypoperfusion to the right parahippocampal cortex. F-18 FDG coincidence imaging showed a more extensive reduction in glucose metabolism compared with SPECT. The MRI scan confirmed the presence of a small segmental choroidal artery infarction. The Tc 99m HMPAO and F-18 FDG scans were not consistent with AD. CONCLUSIONS: This case illustrates the value of the regional cerebral blood flow SPECT for evaluating memory impairment in the elderly. Decreased regional cerebral blood flow to the posterior temporoparietal region is consistent with AD, whereas regional cerebral blood flow diminution in a vascular territory is consistent with vascular dementia. In this case, the patient was clinically diagnosed with AD, and SPECT was performed to establish the baseline regional cerebral blood flow before the cholinesterase inhibitor donepezil was administered. An infarction was diagnosed on the regional cerebral blood flow brain SPECT scan, which was later confirmed by MRI. Infarctions of the parahippocampal cortex may resuft in memory impairment, which can appear clinically similar to AD. PMID- 10698415 TI - Lymphoscintigraphy using larger colloid particles may enhance visualization of the sentinel node in breast cancer: a case report. AB - Lymphoscintigraphy along with a gamma-detecting probe has been applied successfully to breast cancer patients to localize the sentinel node during surgery, with a 5% false-negative rate. The authors report a case of stage II breast cancer. The sentinel node was not visualized on the initial lymphoscintigraphy with peritumoral injection of 37 MBq (1 mCi) 10 ml Tc-99m colloidal rhenium sulfide (Tc-99m ReS colloid; average particle size, 100 nm). However, the sentinel node was visualized in the left lateral view of the second lymphoscintigraphy with peritumoral injection of 37 MBq (1 mCi) 10 ml Tc-99m ReS colloid (average particle size, 500 nm). Lymphoscintigraphy using the larger colloid particles may enhance visualization of a sentinel node in breast cancer. PMID- 10698416 TI - The role of Tc-99m (V) DMSA scintigraphy in the evaluation of superscan on bone scintigraphy. AB - PURPOSE: This study evaluated the biodistribution of Tc-99m (V) DMSA in patients with superscans on bone imaging and defined its role in differentiating the underlying cause. METHODS: Nine patients (five with metastatic and four with metabolic bone disease) with classical superscans were entered into the study. All patients had the necessary radiologic and biochemical studies and a final diagnosis was reached accordingly. Tc-99m (V) DMSA scintigraphy was performed 1 week after Tc-99m MDP whole-body bone imaging. RESULTS: In four of five patients with widespread skeletal metastases, Tc-99m (V) DMSA scan showed diffusely increased bone uptake. In the remaining patient, the Tc-99m (V) DMSA scan showed a normal distribution pattern. All patients with metabolic bone disease had increased bone uptake on Tc-99m (V) DMSA scans. CONCLUSION: Tc-99m (V) DMSA shows increased bone uptake in patients having a superscan appearance in metastatic or metabolic bone disease. Tc-99m (V) DMSA imaging may play a role in the evaluation of patients with equivocal bone scan findings for a superscan. PMID- 10698417 TI - Tc-99m albumin scintigraphy to monitor the effect of treatment in protein-losing gastroenteropathy. AB - PURPOSE: Tc-99m albumin scintigraphy is a noninvasive method for detecting protein-losing gastroenteropathy. METHODS: Seven patients with protein-losing gastroenteropathy were evaluated with Tc-99m albumin scintigraphy. In addition, Tc-99m albumin scintigraphy was used to monitor the effect of treatment in five of these seven patients. The scintigraphic results were compared with serum albumin levels. RESULTS: All seven patients had positive results in the pretreatment images. Changes in scintigraphic findings in the five treated patients correlated well with changes in serum albumin level. CONCLUSION: Tc-99m albumin scintigraphy is useful not only for diagnosing protein-losing gastroenteropathy but also for monitoring the effect of treatment. PMID- 10698418 TI - Diagnostic value of TI-201 and three-phase bone scintigraphy for bone and soft tissue tumors. AB - PURPOSE: Although TI-201 is highly sensitive for detecting bone and soft-tissue tumors, its uptake is not specific for malignant lesions. This study assessed the differentiation of malignant and benign lesions and evaluated the sensitivity, specificity, and accuracy of TI-201 imaging and three-phase bone scans. MATERIALS AND METHODS: Forty bone and soft-tissue tumors (16 malignant and 24 benign) were evaluated. TI-201 static images were acquired 10 minutes (early) and 2 hours (delayed) after injection of the radionuclide. Within 14 days, three-phase bone scintigraphy was performed using Tc-99m HMDP with the patient in the same position. The count ratio of the lesion compared with the normal contralateral or adjacent site (L:N ratio) was measured. RESULTS: With TI-201 scintigraphy, mean (+/- SD) values of early and delayed L:N ratios were 3.36 +/- 1.25 and 2.88 +/- 1.20, respectively, in malignant lesions; and 1.88 +/- 1.14 and 1.48 +/- 0.76, respectively, in benign lesions. TI-201 accumulation in benign lesions was significantly less than that of malignancies on early and delayed images. However, an overlap of both ratios between malignant and benign lesions was seen. No such significance was detected on three-phase bone scintigraphy (L:N ratios of malignant and benign tumors were 2.57 +/- 1.22 and 2.24 +/- 2.11, respectively, for blood flow imaging; 2.41 +/- 0.78 and 2.26 +/- 1.54, respectively, for blood pool imaging; and 2.80 +/- 2.10 and 2.89 +/- 4.55, respectively, for bone imaging). When we assumed that the tumor was malignant when the delayed TI-201 L:N ratio exceeded the blood pool phase L:N ratio with bone scintigraphy, the sensitivity rate was 81%, specificity rate was 100%, and accuracy rate was 93%. CONCLUSIONS: TI-201 imaging for bone and soft-tissue tumors was better than three phase bone scintigraphy alone but was not good enough to clearly differentiate malignant lesions from benign ones. TI-201 scintigraphy, performed in combination with three-phase bone scintigraphy, may be superior to either one of the two imaging procedures alone for bone and soft-tissue tumor diagnosis. PMID- 10698419 TI - Long-term follow-up of myocardial perfusion and metabolic images in a patient with hypertrophic cardiomyopathy. PMID- 10698420 TI - Leg vein uptake of TI-201 in a patient with venous thrombosis in the lower extremity. PMID- 10698421 TI - Melanotic schwannoma mimicking myositis ossificans. PMID- 10698422 TI - Huge necrotic liver metastases in advanced pancreatic carcinoma visualized on bone scans. PMID- 10698423 TI - Incidental detection of skeletal uptake on sestamibi cardiac images in a patient with previously undiagnosed multiple myeloma. PMID- 10698424 TI - Peritoneal linear uptake of Ga-67 caused by tuberculous peritonitis. PMID- 10698425 TI - Sestamibi retention in reactive lymph node hyperplasia: a cause of false-positive parathyroid localization. PMID- 10698426 TI - Cerebral blood flow and glucose metabolism in an infant with Sturge-Weber syndrome. PMID- 10698427 TI - Staging an esophageal carcinoma by F-18 fluorodeoxyglucose whole-body positron emission tomography. PMID- 10698428 TI - Incidental visualization of an esophageal prosthesis on Tc-99m DISIDA cholescintigraphy. PMID- 10698429 TI - Bone marrow scan using Tc-99m-labeled anti-granulocyte antibody to evaluate hematopoiesis in osteomyelofibrosis. PMID- 10698430 TI - Demonstration of spinal osteomyelitis with Ga-67 citrate, Tc-99m MDP, and Tc-99m ciprofloxacin with provisionally negative results on MRI. PMID- 10698431 TI - Uptake of Tc-99m human immunoglobulin in malignant abdominal lymphoma. PMID- 10698432 TI - Thoracic kyphosis results in photopenia in the upper thoracic and lumbar spine. PMID- 10698433 TI - Sentinel nodes. Interval nodes, lymphatic lakes, and accurate sentinel node identification. PMID- 10698434 TI - Current readings in nuclear medicine. PMID- 10698435 TI - Structure-activity relationship studies of chloromethyl ketone derivatives for selective human chymase inhibitors. AB - Based on the SAR study of a classical chloromethyl ketone derivative, Z-PheCH2Cl 1, a series of compounds were synthesized. Among all the derivatives, compound 21 was found to be a potent human chymase inhibitor with no inhibitory activity against human leukocyte cathepsin G. PMID- 10698436 TI - Preparation of 5-(2,6-dideoxy-2-fluoro-alpha-L-talopyranosyloxy)-6-hydroxynap htho[2,3- f]quinoline-7,12-dione (FT-Alz), a new-type, potentially antitumor substance with various biological activities. AB - The title compound (6), its structure being imaginatively created, has been prepared through coupling of alizarine blue (2), a classical dye, and 3,4-di-O acetyl-2,6-dideoxy-2-fluoro-alpha-L-talopyranosyl bromide (3). Compound 6 has considerably higher and different antitumor activity from that of doxorubicin or its analogue (10), and, further, has properties to reverse multidrug resistance (by P-glycoprotein), to inhibit topoisomerase II, and to induce apoptosis. PMID- 10698437 TI - Synthesis and preliminary biological evaluation of [3H]-MRE 3008-F20: the first high affinity radioligand antagonist for the human A3 adenosine receptors. AB - The synthesis and the preliminary biological evaluation of the first high affinity radioligand antagonist for the human A3 adenosine receptor, named [3H] MRE 3008-F20 are reported. [3H]-MRE 3008-20 bound human A3 receptors expressed in CHO cells with K(D) and Bmax value of 0.82 +/- 0.08 nM and 297 +/- 28 fmol/mg of protein, respectively. [3H]-MRE 3008-F20 represents a useful tool for a further characterization of A3 adenosine receptor subtype. PMID- 10698438 TI - N-(sulfonamido)alkyl[tetrahydro-1H-benzo[e]indol-2-yl]amines: potent antagonists of human neuropeptide Y Y5 receptor. AB - [3a,4,5,9b-Tetrahydro-1H-benzo[e]indol-2-yl]amines were prepared via reductive amination and concomitant cyclization of alpha-cyanomethyl-beta-aminotetralins. N acylation with omega-sulfonamido-carboxylic acids and subsequent reduction afforded a series of N-(sulfonamido)alkyl[tetrahydro-1H-benzo[e]indol-2 yl]amines, which bound to the human neuropeptide Y Y5 receptor with nanomolar affinity. PMID- 10698439 TI - Amido-(propyl and allyl)-hydroxybenzamidines: development of achiral inhibitors of factor Xa. AB - The design, synthesis and SAR of amido-(propyl and allyl)-hydroxybenzamidine coagulation factor Xa inhibitors is described. These achiral inhibitors are selective for fXa vis a vis structurally related serine proteases and are readily prepared in 6-7 linear steps. The most potent member 9j (fXa Ki = 0.75 nM) is selective (>1000-fold) and an effective anticoagulant in mammalian plasma. PMID- 10698440 TI - The discovery of potent cRaf1 kinase inhibitors. AB - A series of benzylidene-1H-indol-2-one (oxindole) derivatives was synthesized and evaluated as cRaf-1 kinase inhibitors. The key features of the molecules were the donor/acceptor motif common to kinase inhibitors and a critical acidic phenol flanked by two substitutions. Diverse 5-position substitutions provided compounds with low nanomolar kinase enzyme inhibition and inhibited the intracellular MAPK pathway. PMID- 10698441 TI - Structure-activity relationships of carboxymethylpullulan-peptide-doxorubicin conjugates--systematic modification of peptide spacers. AB - A series of carboxymethylpullulan (CMPul)-doxorubicin (DXR) conjugates bound by peptide spacers of different compositions and lengths were prepared and evaluated for their in vivo antitumor effects. Systematic study of the peptide spacers indicated that CMPul-DXR conjugates bound via appropriate dipeptide spacers were more potent than DXR. PMID- 10698442 TI - Mechanistic studies on type I and type II dehydroquinase with (6R)- and (6S)-6 fluoro-3-dehydroquinic acids. AB - (6R)- and (6S)-6-Fluoro-3-dehydroquinic acids are shown to be substrates for type I and type II dehydroquinases. Their differential reactivity provides insight into details of the reaction mechanism and enables a novel enzyme-substrate imine to be trapped on the type I enzyme. PMID- 10698443 TI - Chemical and enzymatic modifications of integric acid and HIV-1 integrase inhibitory activity. AB - Integric acid (1), an acyl eremophilane sesquiterpenoid, was identified as an inhibitor of HIV-1 integrase, the enzyme responsible for provirus entry into the host cell nucleus and integration in to the host genome. Chemical and enzymatic modification of integric acid led to the preparation of several selective chemical derivatives of integric acid. Preparation, HIV-1 inhibitory activity, and the structure-activity relationship against coupled and strand transfer assays are described. It appears that most of the groups present in the natural product are required for inhibition of HIV-1 integrase strand transfer activity. In contrast, inhibition of 3' processing activity is less stringent suggesting distinct SAR for the two integrase reactions. PMID- 10698444 TI - Chemical synthesis and cytotoxicity of some azinomycin analogues devoid of the 1 azabicyclo[3.1.0]hexane subunit. AB - A series of compounds related to the left-hand domain of the azinomycins have been made and evaluated for cytotoxic activity against a small panel of human tumour cell lines. The epoxide ring is shown to be essential for biological activity. Cytotoxicity is also shown to be sensitive to changes in the substitution pattern on the aromatic ring and the amide group. PMID- 10698445 TI - Exploring the chiral space within the active site of alpha-thrombin with a constrained mimic of D-Phe-Pro-Arg--design, synthesis, inhibitory activity, and X ray structure of an enzyme-inhibitor complex. AB - An indolizidinone motif with strategically placed substitutents was designed and synthesized as a constrained mimic of D-Phe-Pro-Arg. Low nanomolar inhibition of alpha-thrombin validates the design elements in this inhibitor which also exhibits a 20-fold selectivity for thrombin versus trypsin. An X-ray crystal structure of the inhibitor with alpha-thrombin shows the expected interactions with key amino acids within the active site and some notable changes in positions. PMID- 10698446 TI - Recognition and inhibition of HIV integrase by a novel dinucleotide. AB - The viral enzyme, HIV integrase, is involved in the integration of viral DNA into host cell DNA. In the quest for a small nucleotide system with nuclease stability of the internucleotide phosphate bond and critical structural features for recognition and inhibition of HIV-1 integrase, we have discovered a conceptually novel dinucleotide, pIsodApdC, which is a potent inhibitor of this key viral enzyme. PMID- 10698447 TI - 1-[2-(Diphenylmethoxy)ethyl]-2-methyl-5-nitroimidazole: a potent lead for the design of novel NNRTIs. AB - A novel family of non-nucleoside reverse transcriptase inhibitors (NNRTIs) active at submicromolar concentrations was discovered. The new derivatives are 1-[2 (diarylmethoxy)ethyl]imidazoles bearing substituents both at benzene and imidazole rings. The most potent derivatives were those having nitro and methyl groups as substituents in the imidazole ring. Among 10 test derivatives compound 6d was found to be as potent as nevirapine and was selected as a lead for further studies. PMID- 10698448 TI - N-formyl hydroxylamine containing dipeptides: generation of a new class of vasopeptidase inhibitors. AB - Four primary zinc-binding pharmacophores (thiols, carboxylates, phosphorus acids, and hydroxamates) have been utilized in generating inhibitors of zinc metalloproteases such as ACE, NEP, the MMPs, and ECE. Although compounds which inhibit the activity of both ACE and NEP (vasopeptidase inhibitors, VPIs) have been reported which incorporate a thiol, carboxylate, or phosphorus acid pharmacophore, the generation of hydroxamate based vasopeptidase inhibitors has remained elusive. Herein we report the first potent vasopeptidase inhibitors which were generated from the incorporation of conformationally restricted dipeptide mimetics to an N-formyl hydroxylamine zinc-binding group. Compounds such as 13c and 13d are among the most potent in this series, exhibiting in vitro activity comparable to other classes of inhibitors. PMID- 10698449 TI - Synthesis and evaluation of daunorubicin-paclitaxel dimers. AB - Bifunctional, heterodimeric compounds were synthesized to test their ability to create polyvalent arrays between DNA and microtubules in cells. Each dimer was examined for the capacity to bind to microtubules and for cytotoxicity against MES-SA and MES-SA/Dx5 cell lines. PMID- 10698450 TI - Bioisosteric modification of PETT-HIV-1 RT-inhibitors: synthesis and biological evaluation. AB - Bioisosteric substitution of the thiourea (3, 5, 7, 9) and urea (10) moiety of PETT compounds with sulfamide (1), cyanoguanidine (2, 4) and guanidine (6, 8) functionalities, and replacement of the phenethyl group with benzoylethyl group (compounds 11-20) have been studied. Synthesis and antiviral activities are described. PMID- 10698451 TI - Synthesis of immunoreagents for detection of deoxypyrrololine, a cross-link of bone collagen. AB - Two immunogens (5,6) and two probes (fluorescent 7 and chemiluminescent 8) were prepared from benzyl ester (-)-10. These immunoreagents (5,6 and 7,8) are useful for detection of collagen cross-link (+)-deoxypyrrololine (Dpl, 4), and for development of assays for osteoporosis. PMID- 10698452 TI - 3-Imidazolylmethylaminophenylsulfonyltetrahydroquinolines, a novel series of farnesyltransferase inhibitors. AB - Design, synthesis and structure-activity relationship of a series of 3 imidazolylmethylaminophenylsulfonyltetrahydroquinolines as farnesyltransferase inhibitors are presented. A working pharmacophore of inhibiting farnesyltransferase by this series of inhibitors is proposed. PMID- 10698453 TI - A novel class of apical sodium co-dependent bile acid transporter inhibitors: the 2,3-disubstituted-4-phenylquinolines. AB - A series of 2,3-disubstituted-4-phenylquinolines were prepared and were found to inhibit the apical sodium co-dependent bile acid transporter (ASBT). Alkyl and ester substitution at the 3-position showed comparable activities while substitution at the 2-position was much more sensitive to the nature of the substituent. The synthesis and in vitro potency data are presented for this novel class of compounds. PMID- 10698454 TI - Probing the extended binding determinants of oligosaccharyl transferase with synthetic inhibitors of asparagine-linked glycosylation. AB - Protein glycosylation is associated with many critical biological processes. In connection with studies on the mechanism of asparagine-linked glycosylation by the enzyme oligosaccharyl transferase, we have prepared peptide inhibitors that interact with the enzyme at nanomolar concentrations. Herein we describe efforts directed toward improving the binding properties of these inhibitors. PMID- 10698455 TI - Synthesis and enzyme inhibitory activities of a series of lipidic diamine and aminoalcohol derivatives on cytosolic and secretory phospholipases A2. AB - We have synthesised some lipidic diamines and aminoalcohols and examined their behaviour as inhibitors of secretory and cytosolic PLA2. Some structure-activity relationships considerations have been deduced. Compound 14 was a potent and selective inhibitor of cPLA2 and compound 4 showed a dual inhibitory profile against both types of PLA2 while no cytotoxicity at 10 microM on human neutrophils or on murine macrophage line was observed for both. PMID- 10698456 TI - The influence of modified purine bases on the stability of parallel DNA. AB - The stability of the parallel-stranded (ps) DNA duplexes is increased when the dA residues are replaced by the 7-substituted 7-deaza-2'-deoxyadenosine derivatives 3a,b or the dG-residues by the 8-aza-7-deazapurine 2'-deoxynucleosides 6 and 7a,b. Also the N-7-glycosylated adenine 5 forms stable base pairs in ps-DNA while it destabilizes oligonucleotide duplexes with antiparallel chain orientation. The presence of a 2-amino group as in compound 4b is critical for the DNA-structure, leading to a much greater destabilization of the ps-hybrids than of aps-DNA. PMID- 10698457 TI - Synthesis and study of a new adenine-acridine tandem, inhibitor of exonuclease III. AB - A new heterodimer adenine-chain-acridine containing a mixed amido-guanidinium linker chain was synthesized. To achieve the synthesis a new method of introduction of aminoalkyl chain at position 9 of adenine was designed. The heterodimer interacts specifically with the abasic sites in DNA and inhibits the major base excision repair enzyme in Escherichia coli, Exonuclease III. PMID- 10698458 TI - Design, synthesis and biological evaluation of 7-azatricyclodecanes: analogues of cocaine. AB - The synthesis and biological activity of a series of azatricyclodecane analogues of cocaine are described. All compounds studied in this series exhibit nanomolar potency and good selectivity for the serotonin transporter versus the dopamine transporter. PMID- 10698460 TI - Synthesis and antitumor activities of 5-methyl-1- and 2-[[2 dimethylaminoethyl]amino]-aza-thiopyranoindazoles. AB - The synthesis of 1- and 2-substituted aza-benzothiopyranoindazoles has been accomplished. The comparisons of the in vitro antitumor activities of the 2 substituted analogues with the benzothiopyranoindazole chemotypes indicate that the positioning of the nitrogen atom at C-9 (9-aza analogue 4d) leads to a substrate with potent antitumor activity. The 1-substituted aza benzothiopyranoindazoles, in comparison with the corresponding 2-substituted analogues, exhibit a much lower potency. PMID- 10698459 TI - The de novo design and synthesis of cyclic urea inhibitors of factor Xa: optimization of the S4 ligand. AB - In this report refinements to the S4 ligand group leads to compound 19, an inhibitor of fXa with good potency in vitro and an improved pharmacokinetic profile in rabbit. The X-ray crystallographic study of a representative analogue confirms our binding model for this series. PMID- 10698461 TI - Novel C-4 heteroaromatic kainoid analogues: a parallel synthesis approach. AB - New C-4 thiazole 4, 5 and aminothiazole 6, 7 kainoid analogues were efficiently synthesised in five steps from commercially available (-)-alpha-kainic acid I and exhibited strong binding to the kainate receptors. A reactive alpha-bromoketone 10 was generated and reacted with thioamides and thioureas to form thiazole and aminothiazole heterocycles 11-14. Deprotection gave the new kainoid amino acids 4 7 in excellent yield. PMID- 10698462 TI - Prevalence of asthma, rhinitis, and eczema in Northern Thai children from Chiang Mai (International Study of Asthma and Allergies in Childhood, ISAAC). AB - The Phase One International Study of Asthma and Allergies in Childhood (ISAAC) was developed for geographical and temporal comparisons of the prevalence and severity of asthma and allergic diseases in childhood within and between countries. In Thailand, a study was carried out in Bangkok and this is a further study undertaken in Chiang Mai. Using the ISAAC questionnaire, schoolchildren of two age groups were studied: 6-7 years old (n = 3,828) and 13-14 years old (n = 3,927). The data were entered and analyzed using the Epi-info computer program. The prevalence over the past 12 months, in 6-7 and 13-14 years olds, respectively, are as follows; wheezing, 5.5% and 12.6%; rhinitis, 18.5% and 38.3%; rash at flexural areas, 12.9% and 10.7%. The prevalence of these three conditions was found to be close to, but slightly lower than, that from Bangkok, except for eczema in older children. PMID- 10698463 TI - The effect of allergen immunotherapy on in vitro IL-4 and IFN-gamma production by peripheral mononuclear cells in house dust-sensitive Chinese patients with bronchial asthma. AB - The IFN-gamma produced by Th1 cells and IL-4 produced by Th2 cells are two most important cytokines in the regulation of IgE production. House dust immunotherapy has been tried in the treatment of house dust-sensitive Chinese asthmatic patients with good results. We examined the influence of such treatment on in vitro IL-4 and IFN-gamma production by peripheral blood mononuclear cells in house dust-sensitive asthmatic patients. Allergen immunotherapy in house-dust sensitive asthmatic patients can significantly decrease IL-4 production from peripheral mononuclear cells (p<0.05). The production levels of IL-4 in patients without treatment had higher levels than those in patients with hyposensitization (p<0.01). Such therapy also have some effect on promotion of IFN-gamma production in asthmatic patients. In conclusion, immunotherapy with house dust may have the potential ability to shift the Th1/Th2 balance of immune response to allergens and to create a favorable cytokine microenvironment to suppress the allergic reaction in the asthmatic airway. PMID- 10698464 TI - Comparative study of the pharmacokinetic characteristics of slow release theophylline oral preparations in Thai children with persistent asthma. AB - Significant differences in the rate and extent of absorption exist between slow release theophylline (SRT) preparations. The pharmacokinetic characteristics of Xanthium were compared with those of Theo-Dur in twelve Thai children with stable persistent asthma by randomized, double blind, crossover study. Serum theophylline concentrations (STCs) were determined by fluorescence polarization immunoassay. The pharmacokinetic parameters were estimated by using a computer program (Topfit 2.0). The STCs, at steady state after different doses, were predicted by using the modified Wagner-Nelson Equation. The mean resident time (MRT) and apparent T1/2 were significantly larger for Xanthium, but the Cmax and AUC0-infinity of Xanthium were significantly lower than those of Theo-Dur. The Frel of Xanthium was 80.1% relative to Theo-Dur. The appropriate dosing interval of both preparations for Thai children was twice a day. PMID- 10698465 TI - Nasal endoscopic findings in patients with perennial allergic rhinitis. AB - Nasal endoscopy was carried out in 83 patients with perennial allergic rhinitis to evaluate endonasal anatomic variation and to find the correlation between the symptoms of patients and the endoscopic findings. All of the patients had nasal symptoms, 7.2% of the patients were runner, 7.2% were blocker and 85.6% were both. 86.75% of the patients had allergy-related symptoms, i.e. throat symptoms (73.5%), sinus headache (50.6%), and smell disturbance (10.8%). 95.2% of patients had abnormal endoscopic findings, i.e. deviated nasal septum (72.3%), abnormal middle turbinate (49.4%), narrowing of the entrance into the frontal recess (30.1%), septal spur (25.3%), obstruction of the entrance into the frontal recess (19.3%), nasal polyps (15.7%), mucopurulent discharge (14.5%), inferior turbinate hypertrophy (10.8%), abnormal uncinate process (9.6%), abnormal ethmoid bullae (7.2%), and enlargement of aggar nasi cells (2.4%). There was no significant correlation between each symptom and each endoscopic finding. However, there was a significant correlation between sinus headache and all of the combined abnormal endoscopic findings (P<0.05). These findings suggested that variations in endonasal anatomy was not by itself a pathology or a cause of symptoms. However, a combination of these variations may narrow the cleft of the ostiomeatal unit and cause contact area or stenosis, which predisposed patients to persistent symptoms, recurrent infection or resistance to therapy in patients with perennial allergic rhinitis. The endoscope might be a very useful tool for allergists, immunologists, and rhinologists, who work in the nose to deal with these cases. PMID- 10698466 TI - Serum eosinophil cationic protein determination in asthmatic children-effect of different collecting tubes used for blood sampling. AB - We compared the effect of using 2 different serum collecting tubes, serum separation tubes (SST, with clot activator and gel barrier) and conventional glass tubes (with no additives), on circulating levels of eosinophil cationic protein (ECP) in asthmatic children and controls. The serum ECP values obtained from both SST and glass tubes were significantly higher in asthmatic children than in corresponding controls. ECP values were higher in serum samples using SST than in those using glass tubes (P<0.01), while no difference was found between the two in controls. ECP levels correlated with peripheral eosinophil counts, for SST samples and glass tube samples alike. The difference in ECP levels between these two tubes also correlated with circulating eosinophil counts (r = 0.62, P = 0.004) After 18-hour storage at room temperature, the ECP values increased significantly in samples obtained from asthmatic children. No difference in ECP values between SST samples and glass tube samples was found for 18 hour samples. Thus, ECP levels obtained from SST samples and glass tube samples, though reliable, should not be directly compared, especially in asthmatic children with eosinophilia. PMID- 10698467 TI - Comparison of counter immunoelectrophoresis with immunoblotting for detection of anti-extractable nuclear antigen antibodies in systemic lupus erythematosus. AB - Anti-extractable nuclear antigen (ENA) antibodies were assayed by counter immunoelectrophoresis (CIE) and immunoblotting in patients with systemic lupus erythematosus (SLE). We found the two methods showed good concordance rates, the lowest being 67% for anti-SS-A. Immunoblotting was more sensitive in detecting anti-Sm, anti-SS-B and anti-PCNA (proliferating cell nuclear antigen); CIE was more sensitive for anti-nRNP and anti-SS-A. Overall, the prevalence of these anti ENA antibodies in SLE was increased by 9-20% if immunoblotting was used in addition to CIE. Sera specific for the 52 kDa peptide of the SS-A antigen (anti 52kDa SS-A) were better detected by immunoblotting. Anti-PCNA antibody was found in 6.3% of SLE patients and was associated with active disease and hemolytic anemia. The positive rate of anti-Sm was 9% by CIE and 23.7% by immunoblotting and this antibody was a specific marker for SLE using either method. It was concluded that using immunoblotting in addition to CIE, the overall sensitivity of detection of anti-ENA antibodies in SLE was increased and clinically useful antibodies such as anti-52kDa SS-A and anti-PCNA could be detected. PMID- 10698468 TI - Antineutrophil cytoplasmic antibody (ANCA) and rapidly progressive crescentic glomerulonephritis in Thai population. AB - The impact of vasculitis as a cause of primary rapidly progressive crescentic glomerulonephritis (RPGN) was examined in patients with Thai ethnic by antineutrophil cytoplasmic antibody (ANCA) test. Thirty patients found in a six years study period were included. Patients' mean age was 34.8+/-16.4 years. Mean crescent score was 86.2+/-22.9%. ANCA proved positive in fifteen patients. This helps to differentiate vasculitis associated (ANCA positive) from nonvasculitis (ANCA negative) RPGN. Incidence of immune complex type RPGN (46.6%) is higher than the Caucasians while the incidence of antiglomerular basement membrane antibody (anti-GBM disease) is much lower. More vasculitis patients were treated with cyclophosphamide (n = 11) than the nonvasculitis group (n = 2). Mean renal survival time of ANCA and non-ANCA associated patients were 26.69 and 14.16 months, respectively. Renal survival of all patients is significantly worse if associated with a high entry creatinine (>6 mg/dl). Our results show that vasculitis associated RPGN is not an uncommon disease in the Thai population and can be recognized initially by ANCA test. PMID- 10698469 TI - Delayed administration of G-CSF in both mobilization and post transplantation in autologous peripheral blood stem cell transplantation. AB - Granulocyte colony stimulating factors (G-CSFs) play a very important role in the current technique of stem cell transplantation. The conventional timing of administration of G-CSF in both mobilization and post transplantation has been right after chemotherapy or right after transplantation. We have studied the effects of timing of administration of G-CSF in 21 patients who had autologous stem cell transplantation for breast cancer, lymphoma or nasopharyngeal cancer. Their stem cells were mobilized by chemotherapy followed by G-CSF, which were given on day +1 or day +5 after chemotherapy. The median peak percentage of CD34 positive cells harvested using both technique were 1.88 and 0.48% respectively. After transplantation, G-CSF were given on day +1 or day +6 after stem cell infusion until neutrophil recovery. The time until bone marrow recovery was significantly longer in the group with delayed administration of G-CSF (10 days versus 8 days). However, there was no difference in duration of neutropenic fever or hospital stay after transplantation. The transplantation outcome was also unaffected. We therefore concluded that G-CSF can be given in the delayed fashion in both mobilizing and post transplantation settings without jeopardizing the outcome and this would result in a significant cost saving. PMID- 10698470 TI - Cytomegalovirus infection in immunocompromised children in Thailand. AB - Cytomegalovirus (CMV) constitutes the most widespread cause of congenital and perinatal viral infections on a global scale, exceeding a 90%-prevalence among blood donors in Thailand. The present study was aimed at determining the prevalence of CMV in the sera of 113 immunocompromised children by means of serology, as well as polymerase chain reaction using nested primers. Our results showed anti-CMV IgG, i.e. latent infection, prevalent in an age-dependent manner irrespective of the disorder underlying the children's immunocompromised condition, whereas anti-CMV IgM was equally prevalent throughout all age groups and disease patterns and, therefore, unreliable as a marker. Detection of serum CMV DNA by PCR represented the most exact diagnostic parameter by far, indicating active infection long before clinical symptoms may appear. In conclusion, based on the high prevalence of latent CMV infection among the general population in Thailand, we recommend especially the sera of immunocompromised patients be examined for the presence of viral DNA by means of PCR in order to provide clinical guidelines for their proper management. PMID- 10698471 TI - Combined immunoprecipitation and agglutination for the detection of the heterodimeric molecule: human chorionic gonadotropin as a study model. AB - Double-particle reversed passive hemagglutination (RPHA) was performed for the detection of an intact heterodimeric form of human chorionic gonadotropin (intact hCG) composed in Profasi hCG (P-hCG). This technique relied on two mixed types of human O RBC, which were individually coated with two distinct monoclonal antibodies that recognized alpha or beta subunit of hCG, i.e. ALC-1 and BEL-5, respectively. The positive hemagglutination result was achieved by this technique. However, in the BEL-5 coated single-particle control system, positive results for both P-hCG and beta subunit hCG solution were realized. The occurrence of beta-multimer hCG was a causative molecule revealed by the hemagglutination inhibition technique. Thereby, the novel method called "combined immunoprecipitation and agglutination" was developed to overcome this problem. The free beta subunit together with the betamultimer hCG were eliminated from other forms presented in P-hCG after using the ALC-1 coated particles. The precipitated particles, which captured the heterodimer hCG molecule, reacted further with soluble BEL-5 to subsequently form a trellis. A positive result was obtained only with P-hCG, but not with beta subunit hCG or hLH. This study is inferable as a model for the detection of heterodimeric molecule by an elementary method. PMID- 10698472 TI - Perspectives on surrogate end points in the development of drugs that reduce the risk of cancer. AB - This paper proposes a scientific basis and possible strategy for applying surrogate end points in chemopreventive drug development. The potential surrogate end points for cancer incidence described are both phenotypic (at the tissue, cellular, and molecular levels) and genotypic biomarkers. To establish chemopreventive efficacy in randomized, placebo-controlled clinical trials, it is expected that in most cases it will be critical to ensure that virtually all of the biomarker lesions are prevented or that the lesions prevented are those with the potential to progress. This would require that both the phenotype and genotype of the target tissue in agent-treated subjects, especially in any new or remaining precancers, are equivalent to or show less progression than those of placebo-treated subjects. In the National Cancer Institute chemoprevention program, histological modulation of a precancer (intraepithelial neoplasia) has thus far been the primary phenotypic surrogate end point in chemoprevention trials. Additionally, we give high priority to biomarkers measuring specific and general genotypic changes correlating to the carcinogenesis progression model for the targeted cancer (e.g., progressive genomic instability as measured by loss of heterozygosity or amplification at a specific microsatellite loci). Other potential surrogate end points that may occur earlier in carcinogenesis are being analyzed in these precancers and in nearby normal appearing tissues. These biomarkers include proliferation and differentiation indices, specific gene and general chromosome damage, cell growth regulatory molecules, and biochemical activities (e.g., enzyme inhibition). Serum biomarkers also may be monitored (e.g., prostate-specific antigen) because of their accessibility. Potentially chemopreventive drug effects of the test agent also may be measured (e.g., tissue and serum estrogen levels in studies of steroid aromatase inhibitors). These initial studies are expected to expand the list of validated surrogate end points for future use. Continued discussion and research among the National Cancer Institute, the Food and Drug Administration, industry, and academia are needed to ensure that surrogate end point-based chemoprevention indications are feasible. PMID- 10698473 TI - Is cadmium a cause of human pancreatic cancer? AB - Little is known about the etiology of pancreatic cancer, which is an important cause of cancer mortality in developed countries. We hypothesize that exposure to cadmium is a cause of pancreatic cancer. Cadmium is a nonessential metal that is known to accumulate in the human pancreas. The major risk factors for pancreatic cancer (increasing age, cigarette smoking, residence in Louisiana, and occupations involving exposure to metalworking and pesticides) are all associated with increased exposure to cadmium. Our meta-analysis of cohorts with high exposure to cadmium is also consistent with an increased risk of pancreatic cancer (standardized mortality ratio = 166; 95% confidence interval, 98-280; P = 0.059). Cadmium can cause the transdifferentiation of pancreatic cells, increases in the synthesis of pancreatic DNA, and increases in oncogene activation. Thus, cadmium is a plausible pancreatic carcinogen. The cadmium hypothesis provides a coherent explanation for much of the descriptive epidemiology of pancreatic cancer and suggests new avenues for analytical research. PMID- 10698474 TI - Genetic polymorphism of cytochrome P450-1B1 and risk of breast cancer. AB - Cytochrome P450-1B1 (CYP1B1) is a major enzyme catalyzing the formation of genotoxic 4-hydroxyestradiol. This enzyme is also involved in the activation of polycyclic aromatic hydrocarbons and heterocyclic aromatic amines, mammary carcinogens in experimental animals. CYP1B1 is genetically polymorphic, and the variations in the CYP1B1 gene may be related to the risk of breast cancer. We evaluated this hypothesis among 186 breast cancer cases and 200 age-matched controls as part of a large population-based case-control study conducted in urban Shanghai during 1996 to 1998. Genomic DNA from cases and controls was analyzed for genetic polymorphism in codon 432 (Val-->Leu) of the CYP1B1 gene using a PCR-RFLP-based assay. The frequency of the Leu allele was 53% in cases and 46% in controls (P = 0.06). Compared with those with the Val/Val genotype, women with the Leu/Leu genotype had a 2.3-fold [95% confidence interval (CI), 1.2 4.5] elevated risk of breast cancer after adjusting for potential confounding variables. This positive association was more pronounced among postmenopausal women (Odds ratio, 3.1; 95% CI, 1.0-9.1) than premenopausal women (OR, 1.9; 95% CI, 0.8-4.3). Elevated risks of breast cancer associated with homozygosity for the Leu allele were observed in virtually all subgroups of women defined by major risk factors for breast cancer. The results from this study were consistent with recent findings from in vitro and animal experiments implicating a potentially important role of CYP1B1 in the etiology of human breast cancer. PMID- 10698475 TI - Association of diet and mammographic breast density in the Minnesota breast cancer family cohort. AB - Mammographic breast density is a significant risk factor for breast cancer. The present report analyzes the association of breast density and dietary factors in 1508 women in a historical cohort study of breast cancer families in Minnesota. Diet was assessed by a semiquantitative food frequency questionnaire. Percent breast density was estimated visually by a radiologist experienced in mammography. The association of percent breast density with quartiles of energy adjusted dietary intakes was examined in analysis of covariance models adjusting for potential confounding effects of age, body mass index, and other covariates as well as correcting for familial correlation. Analyses were performed on all women combined and were also stratified by menopausal status. Among premenopausal women, percent breast density was positively associated with intakes of polyunsaturated fat, polyunsaturated:saturated fat ratio, and vitamins C and E and was inversely associated with saturated fat and total dairy intake. Among postmenopausal women, vitamin B12 was linearly associated with increased breast density. The positive associations for vitamin C and B12 were attributable to supplement intake only. There was a suggestive positive trend between breast density and daily alcohol consumption in both premenopausal and postmenopausal women. After adjustment for other sources of alcohol, only wine intake among postmenopausal women was significant such that white wine showed a positive association and red wine an inverse association with percent breast density. There was no association with other examined dietary factors. The cross-sectional differences in breast density across levels of dietary factors were small in magnitude but may have implications for breast cancer risk. PMID- 10698476 TI - Risk and aggressiveness of breast cancer in relation to plasma organochlorine concentrations. AB - Several organochlorines identified as "hormone mimics" were proposed as possible risk factors for breast cancer. We conducted a case-control study to assess breast cancer risk and disease aggressiveness in relation to plasma concentrations of several organochlorine compounds. Plasma lipid concentrations of 11 chlorinated pesticides and 14 polychlorinated biphenyl congeners were measured in 315 women newly diagnosed with breast cancer, 219 hospital-based controls, and 307 population controls from the Quebec City area (Canada). Concentrations of hormonally active organochlorines or their surrogates were compared between cases and controls as well as between groups of cases defined according to tumor size and axillary-lymph-node involvement. We found similar levels of organochlorines in cases and controls and no relationship between the relative risk of breast cancer and organochlorine exposure. However, the probability of lymph-node invasion among cases increased with exposure to 1,1 dichloro-2,2-bis(4-chlorophenyl)ethylene [p,p'-DDE; odds ratio, 2.54; 95% confidence interval (CI), 1.20-5.35; between the highest and the lowest tertiles]. Furthermore, p,p'-DDE exposure was associated with a dose-related increased relative risk of exhibiting both lymph-node involvement and a large tumor. Indeed odds ratio raised to 2.33 (95% CI, 0.94-5.77) for the second tertile relative to the first tertile and reached 3.51 (95% CI, 1.41-8.73) for the third tertile relative to the first tertile. Similar associations were noted with beta-hexachlorocyclohexane, oxychlordane, and transnonachlor. We conclude that exposure to persistent, hormonally active organochlorines during adulthood is not associated with breast cancer risk. The possibility that some organochlorines and especially p,p'-DDE may increase breast cancer aggressiveness deserves further attention. PMID- 10698478 TI - Comparative polymerase chain reaction analysis of c-myc amplification on archival breast fine-needle aspiration materials. AB - The oncogene c-myc is a key regulator of cell cycle progression (from G1 to S phase). The amplification of c-myc can either induce cell proliferation or apoptosis. As a part of our ongoing effort to develop methods for multiple tumor marker analysis, this study was carried out to determine whether biomarkers such as c-myc amplification could be analyzed on genetic materials collected from archival fine-needle aspiration (FNA) smears. A novel comparative PCR analysis was used to analyze c-myc amplification semiquantitatively. Genomic DNA was prepared using cells obtained from archival FNA materials that had undergone quantitative fluorescence image analysis (QFIA) for other biomarkers. Of the 72 cases selected from 1995 for this study, 53 had an adequate amount of DNA for analysis. A novel comparative PCR analysis was used to analyze c-myc amplification quantitatively. For each batch of experiments, DNA from the high c myc expressing cells, HL-60, and DNA from the low expressing cells, K562, were served as positive and negative controls, respectively. c-myc amplification was observed in 16 (94.1%) of 17 malignant lesions, 5 (41.7%) of 12 proliferative breast diseases with nuclear atypia, and 4 (16.7%) of 24 other benign lesions (fibroadenoma or fibrocystic disease). The overall difference of c-myc expression among these groups was highly significant by chi2 analysis (P = 0.0002). We conclude that multiple phenotypic markers and genotypic markers may be combined in a risk assessment biomarker profile on small FNA samples that can be obtained on multiple occasions relatively noninvasively from the patient. The results of this study suggest that c-myc amplification may be a biomarker of breast cancer risk. However, additional large, prospective studies are needed to confirm the current observation. PMID- 10698477 TI - Risk of female breast cancer associated with serum polychlorinated biphenyls and 1,1-dichloro-2,2'-bis(p-chlorophenyl)ethylene. AB - This case-control study was designed to investigate the relationship between polychlorinated biphenyls (PCBs) and 1,1-dichloro-2,2'-bis(p chlorophenyl)ethylene (DDE) and breast cancer risk in Connecticut. Cases were incident breast cancer patients who were either residents of Tolland County or who had a breast-related surgery at the Yale-New Haven Hospital in New Haven County. Controls were randomly selected from Tolland County residents or from patients who had newly diagnosed benign breast diseases or normal tissue at Yale New Haven Hospital. A total of 475 cases and 502 controls had their serum samples analyzed for PCBs and DDE in 1995-1997. The age- and lipid-adjusted geometric mean serum level of DDE was comparable between the cases (460.1 ppb) and controls (456.2 ppb). The geometric mean serum level of PCBs was also comparable between cases (733.1 ppb) and controls (747.6 ppb). After adjustment for confounding factors, odds ratios of 0.96 (95% confidence interval, 0.67-1.36) for DDE and 0.95 (95% confidence interval, 0.68-1.32) for PCBs were observed when the third tertile was compared with the lowest. Further stratification by parity, lactation, and menopausal and estrogen receptor status also showed no significant association with serum levels of DDE or PCBs. The results by PCB congener groups also showed no major increased risk associated with any of the congener groups. Our study does not support the hypothesis that DDE and PCBs, as encountered through environmental exposure, increase the risk of female breast cancer. PMID- 10698479 TI - Association of the myeloperoxidase -463G-->a polymorphism with lung cancer risk. AB - Myeloperoxidase, which is released from neutrophils in response to various pulmonary insults including tobacco smoke, is suspected to play a carcinogenic role in the lung. A G-to-A substitution polymorphism in the promoter region of the MPO gene has been suggested in in vitro studies to decrease gene transcription. We tested the association of this polymorphism with lung cancer in a population-based case-control study of 323 cases and 437 controls of Caucasian, Japanese, or Native Hawaiian ancestry in Hawaii. We found a marked difference in the frequency of the variant A allele among Caucasians (26%), Japanese (17%), and Hawaiians (13%). Overall, the variant allele was somewhat less frequent in cases than controls (P = 0.13). Individuals with the A/A genotype were found to be at a 50% decreased risk compared to those with two G alleles (95% confidence interval, 0.2-1.3). Although not statistically significant, this inverse association was suggested in both sexes and two of the three ethnic groups studied. Heterozygotes were at no decreased risk. Further work needs to clarify the functional relevance of the A allele in vivo and to confirm the inverse association of the A/A genotype with lung cancer in large epidemiological studies. PMID- 10698480 TI - GSTM1, GSTT1, GSTP1, CYP1A1, and NAT1 polymorphisms, tobacco use, and the risk of head and neck cancer. AB - Squamous cell carcinoma of the head and neck (SCCHN), including the oral cavity, pharynx, and larynx, provides an ideal tumor model to investigate gene environment interaction. We conducted a hospital-based case-control study including 182 cases with newly diagnosed SCCHN and 202 controls with nonneoplastic conditions of the head and neck that required surgery. Lifetime tobacco use and risk of SCCHN were evaluated in relation to the polymorphisms of GSTM1, GSTT1, GSTP1, CYP1A1, and NAT1. The main effects of genotype were associated with a slightly increased risk of SCCHN for GSTP1 [age-, race-, and sex-adjusted odds ratio (OR), 1.2; confidence interval (CI), 0.8-1.9], GSTT1 (OR, 1.2; CI, 0.7-2.3), and NAT1 (OR, 1.1; CI, 0.7-1.7). The joint effects of genotype combinations showed some excess risk for the combination of the GSTM1 null genotype and the CYP1A1 Ile/Val polymorphism (OR, 2.6; CI, 0.7-10.3). The analysis of the joint effects (interaction) of the "at-risk" genotypes and tobacco use did not reveal any interaction on either the multiplicative or additive scale for GSTM1, GSTP1, or CYP1A1. However, there was a suggestion of an interaction on the additive scale between the pack-years of tobacco use and the GSTT1 null genotype. The combined heterozygote and homozygote NAT1*10 genotypes also had a suggestive interaction with tobacco smoking history. The results of this study suggest a possible gene-environment interaction for certain carcinogen metabolizing enzymes, but larger studies that fully evaluate the interaction are needed. PMID- 10698481 TI - Blood levels of organochlorines before and after chemotherapy among non-Hodgkin's lymphoma patients. AB - Several small studies suggest a link between environmental exposure to organochlorine compounds and risk of non Hodgkin's lymphoma (NHL). Because NHL is uncommon, studies of the topic often use a population-based case-control design, in which cases generally are enrolled after treatment has begun. If chemotherapy affects blood levels of organochlorines, exposure will be misclassified and findings distorted. To determine whether chemotherapy alters serum levels of organochlorines in NHL cases, we compared serum samples before and after treatment in 22 cases diagnosed with NHL between March 1994 and August 1995 and enrolled in a clinical trial at the United States National Cancer Institute's Clinical Center. The time difference between pretreatment and posttreatment samples ranged from 15 to 27 months with an average of 20 months. Laboratory analyses were conducted in blinded pretreatment and posttreatment pairs of the subjects. Pretreatment and posttreatment organochlorine serum levels were compared using Pearson correlation coefficient (r) and paired t test. The pretreatment and posttreatment serum levels were highly correlated for 1,1 dichloro-2,2-bis(p-chlorophenyl)ethylene (DDE) and polychlorinated biphenyls (PCBs) PCB-138, PCB-153, PCB-156, and total PCBs (ranging from 0.78 to 0.93). Serum levels of all of these organochlorines significantly decreased between initiation and completion of chemotherapy, 25% for total PCB (P = 0.0044), 28% for DDE (P = 0.0014), 25% for PCB-138 (P = 0.0053), 27% for PCB-153 (P = 0.0031), and 29% for PCB-156 (P = 0.045). Neither weight change nor lipid change was correlated with changes in chemical levels. There was no association between the length of time between blood draws and changes in chemical levels. Our data raise the possibility that lymphoma treatment depresses serum organochlorine levels. PMID- 10698482 TI - Pancreatic cancer and serum organochlorine levels. AB - Occupational exposure to p,p'-dichlorodiphenyltrichloroethane (DDT) has been associated with increased pancreatic cancer risk. We measured organochlorine levels in serum obtained at the study enrollment from 108 pancreatic cancer cases and 82 control subjects aged 32-85 years in the San Francisco Bay Area between 1996 and 1998. Cases were identified using rapid case-ascertainment methods; controls were frequency-matched to cases on age and sex via random digit dial and random sampling of Health Care Financing Administration lists. Serum organochlorine levels were adjusted for lipid content to account for variation in the lipid concentration in serum between subjects. Median concentrations of p,p' dichlorodiphenyldichloroethylene (DDE, 1290 versus 1030 ng/g lipid; P = 0.05), polychlorinated biphenyls (PCBs; 330 versus 220 ng/g lipid; P<0.001), and transnonachlor (54 versus 28 ng/g lipid; P = 0.03) were significantly greater among cases than controls. A significant dose-response relationship was observed for total PCBs (P for trend <0.001). Subjects in the highest tertile of PCBs (> or =360 ng/g lipid) had an odds ratio (OR) of 4.2 [95% confidence interval (CI) = 1.8-9.4] compared to the lowest tertile. The OR of 2.1 for the highest level of p,p'-DDE (95% CI = 0.9-4.7) diminished (OR = 1.1; 95% CI = 0.4-2.8) when PCBs were included in the model. Because pancreatic cancer is characterized by cachexia, the impact of this on the serum organochlorine levels in cases is difficult to predict. One plausible effect of cachexia is bioconcentration of organochlorines in the diminished lipid pool, which would lead to a bias away from the null. To explore this, a sensitivity analysis was performed assuming a 10-40% bioconcentration of organochlorines in case samples. The OR associated with PCBs remained elevated under conditions of up to 25% bioconcentration. PMID- 10698483 TI - Association between polycyclic aromatic hydrocarbon-DNA adduct levels in maternal and newborn white blood cells and glutathione S-transferase P1 and CYP1A1 polymorphisms. AB - Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous environmental pollutants; a number are carcinogenic. Metabolic polymorphisms may modulate susceptibility to PAH-induced DNA damage and carcinogenesis. This study investigates the relationship between PAH-DNA adduct levels (in maternal and newborn WBCs) and two polymorphisms: (a) an MspI RFLP in the 3' noncoding region of cytochrome P4501A1 (CYP1A1); and (b) an A-->G transition in nucleotide 313 of glutathione S transferase P1 (GSTP1), resulting in an ile105val substitution. CYP1A1 catalyzes the bioactivation of PAH; the CYP1A1 MspI RFLP has been associated with cancer of the lung. GSTP1 catalyzes the detoxification of PAH; the val allele has greater catalytic efficiency toward PAH diol epoxides. The study involves 160 mothers and their newborns from Poland. Regression models controlled for maternal smoking and other confounders. No association was seen between maternal adduct levels and either polymorphism, separately or combined. However, adduct levels were higher among newborns with the CYP1A1 MspI restriction site (heterozygotes and homozygotes combined) compared with newborns lacking the restriction site (P = 0.06). Adducts were higher among GSTP1 ile/val and ile/ile newborns compared with GSTP1 val/val newborns (P = 0.08). Adduct levels were 4-fold higher among GSTP1 ile/ile newborns having the CYP1A1 restriction site compared with GSTP1 val/val newborns who lacked the CYP1A1 restriction site (P = 0.04). This study demonstrates a significant combined effect of phase I and phase II polymorphisms on DNA damage from PAHs in fetal tissues. It illustrates the importance of considering interindividual variation in assessing risks of transplacental exposure to PAHs. PMID- 10698484 TI - Etiology of hepatocellular carcinoma in Italian patients with and without cirrhosis. AB - We performed a case-control study to assess the role of hepatitis B virus (HBV), hepatitis C virus (HCV), GB virus C/hepatitis G virus (HGV), TT virus, alcohol intake, and tobacco smoking as risk factors for hepatocellular carcinoma (HCC) in the presence or absence of cirrhosis. We prospectively recruited 174 patients with a first diagnosis of HCC admitted to the main hospitals in Brescia, North Italy. On the basis of histological, clinical, and radiological criteria, the presence of cirrhosis was established in 142 cases, excluded in 21 cases, and remained undefined in 11 cases. Among the HCC cases without cirrhosis, a histological picture of normal liver was found in a single patient, chronic viral hepatitis was found in 11 patients, alcoholic hepatitis was found in 5 patients, nonspecific reactive hepatitis was found in 3 patients, and hemochromatosis was found in 1 patient. As controls, we also included 610 subjects unaffected by hepatic diseases and admitted to the same hospitals as cases. The odds ratios for having HCC according to positivity for HCV RNA, HBsAg and/or HBV DNA, and alcohol intake > 80 g/day (95% confidence interval) were as follows, in the presence and absence of cirrhosis, respectively: (a) 33.5 (17.7-63.4) and 19.7 (6-64.8) for HCV RNA; (b) 17.6 (9.0-34.4) and 20.3 (5.7-72.6) for HBsAg; and (c) 5.5 (3.1-9.7) and 4.6 (1.5-13.8) for alcohol intake. No association was found with HGV or TT virus infections or tobacco. This study has shown that most HCC cases arising in the area are due to HBV, HCV, or alcohol intake, in both the presence and absence of cirrhosis. PMID- 10698485 TI - NAT1*10 and NAT1*11 polymorphisms and breast cancer risk. AB - Several recent epidemiological studies examined the association of N acetyltransferase (NAT) 1 and 2 genotypes and breast cancer risk. Taken together, these studies do not support a strong role for the most common NAT alleles in etiology of breast cancer. Only one study estimated odds ratios (ORs) for the relatively rare NAT1*11 allele: a strong positive association for the NAT1*11 allele and breast cancer was reported, as well as strong combined effects for NAT1*11-containing genotypes and two environmental factors, smoking and red meat consumption. To further address the association of NAT1*11 and breast cancer, an analysis was performed using previously collected data from the Carolina Breast Cancer Study, a population-based, case-control study conducted in North Carolina. The OR for NAT1*11-containing genotypes and breast cancer was 0.5 (95% confidence interval, 0.2-1.3) among white women; ORs were not calculated among African Americans because only one participant exhibited the NAT1*11 allele. There was no evidence for combined effects of NAT1*11 and smoking. Unfortunately, the results of both studies of NAT1*11 are imprecise and lack sufficient statistical power to address fully the potential contribution of NAT1*11 to breast cancer. These results illustrate that the limitations imposed by sample size, as well as incomplete knowledge of biological function, need to be considered when planning and interpreting studies of genetic polymorphisms and environmental exposures. PMID- 10698486 TI - Duration of gestation and prostate cancer risk in offspring. AB - This large population-based nested case-control study investigated the importance of perinatal characteristics as risk factors for prostate cancer in later life in a cohort of men who were born between 1889 and 1941 in Stockholm, Sweden. Eight hundred and thirty-four prostate cancer cases over 18 years of age and of singleton birth were identified from the cohort between 1958 and 1994. For each case, singleton males born live to the first four mothers admitted after the case's mother were selected as potential controls; 1880 eligible controls were included in the study. For each study subject, we obtained data on mother's parity, pre-eclampsia or eclampsia before delivery, age at delivery, and socioeconomic status, as well as child's birth length and weight, placental weight, and gestational age. Odds ratio (OR) estimates and 95% confidence intervals (CIs) were derived from logistic regression analyses. We found no statistically significant differences between cases and controls with respect to maternal age, socioeconomic status, or parity. Birth weight, birth length, and placental weight were also not significantly related to prostate cancer risk. Pregnancy toxemia (OR = 0.33; 95% CI, 0.07-1.45) and longer gestation age were associated with a reduced risk of prostate cancer; the OR estimate was 0.94 (95% CI, 0.89-0.99) for each 1-week prolongation of the duration of gestation. Our results suggest that birth size indicators are not important risk factors for prostate cancer in later life. In addition, our data on gestation age indicate that the late in utero environment may be as important as the early in utero environment in the modulation of prostate cancer risk in offspring. PMID- 10698487 TI - The p53 Arg72Pro polymorphism, human papillomavirus, and invasive squamous cell cervical cancer. AB - A. Storey et al. [Nature (Lond.), 393: 229-234, 1998)] reported a 7-fold increased risk of cervical cancer associated with having an Arg/Arg polymorphism at codon 72 of p53 compared with the Pro/Arg heterozygotes (odds ratio, 7.4; 95% confidence interval, 2.1-29.4). Complementary in vitro studies suggested that the HPV E6 oncoprotein more readily targets the arginine form, as opposed to the proline form, of p53 for degradation. We investigated the impact of this polymorphism in a population-based case-control study of invasive cervical cancer. Using a PCR assay to detect the p53 codon 72 polymorphism, we tested blood samples from 111 women with invasive squamous cell cancer of the cervix identified by a population-based registry and 164 random-digit telephone-dialed controls. The distribution of the genotype among control women was 38% heterozygous, 7% proline homozygous, and 55% arginine homozygous, and among the cases was 38%, 6%, and 56%, respectively. There was no increased risk of squamous cell invasive cervical cancer associated with homozygosity for the arginine allele (odds ratio, 1.0; 95% confidence interval, 0.6-1.7). Furthermore, there was no modification of this result by human papillomavirus (HPV) DNA status of the tumor, age, or smoking status. Among controls, there was no association between the polymorphism and HPV-16 L1 seropositivity. However, among case subjects, the codon 72 polymorphism may be related to HPV 16L1 seropositivity status. PMID- 10698488 TI - Validity of self-reported colorectal cancer screening behavior. AB - End points for trials promoting cancer screening are often based on self-reported screening behavior. This study was designed to evaluate and optimize the reliability of a computer-assisted telephone interview for collecting self reported colorectal cancer screening behavior. Cases who had received a fecal occult blood test (FOBT), flexible sigmoidoscopy, and/or colonoscopy, and controls who had no record of colorectal screening were identified among 40-75 year-old members of the Denver Kaiser Permanente Health Care Program and were contacted by telephone. Sensitivities and specificities of self-reported screening were calculated by comparison of subjects' recall with Kaiser Permanente records. The questionnaire was revised based upon results of the pilot phase of the study. Using the revised questionnaire, the sensitivity of self reported screening was 96.2% for the FOBT, 94.9% for flexible sigmoidoscopy, 88.7% for colonoscopy, and 96.2% for either endoscopic screening test. The specificity of self-reported screening was 85.9% for the FOBT, 92.2% for flexible sigmoidoscopy, 96.8% for colonoscopy, and 92.0% for either endoscopic screening test. No marked differences in the accuracy of the self-reports were detected as a function of gender, age, ethnicity, or family history of colorectal cancer of the participants. Self-reports of colon cancer screening behavior can be reliably used as end points for intervention trials when carefully phrased questions are used. PMID- 10698489 TI - HPV E6 targeted degradation of the discs large protein: evidence for the involvement of a novel ubiquitin ligase. AB - The Discs Large (DLG) tumour suppressor protein is targeted for ubiquitin mediated degradation by the high risk human papillomavirus (HPV) E6 proteins. In this study we have used a mutational analysis of E6 in order to investigate the mechanism by which this occurs. We first show that the differences in the affinities of HPV-16 and of HPV-18 E6 proteins for binding DLG is reflected in their respective abilities to target DLG for degradation. A mutational analysis of HPV-18 E6 has enabled us to define regions within the carboxy terminal half of the protein which are essential for the ability of E6 to direct the degradation of DLG. Mutants within the amino terminal portion of E6 which have lost the ability to bind the E6-AP ubiquitin ligase, as measured by their ability to degrade p53, nonetheless retain the ability to degrade DLG. Significant levels of DLG degradation are also obtained using wheat germ extracts which lack E6-AP. Finally, we show that the transfer of the DLG binding domain onto the low risk HPV-6 E6 confers DLG binding activity to that protein and, most significantly, allows HPV-6 E6 to target DLG for degradation. These results indicate that E6 mediated degradation of DLG does not involve the E6-AP ubiquitin ligase and, in addition, shows that the high and low risk HPV E6 proteins most likely share a common cellular intermediary in the ubiquitin pathway. PMID- 10698490 TI - Overexpression of the wild-type p53 gene inhibits NF-kappaB activity and synergizes with aspirin to induce apoptosis in human colon cancer cells. AB - The tumor suppressor gene p53 is a potent transcriptional regulator of genes which are involved in many cellular activities including cell cycle arrest, apoptosis, and angiogenesis. Recent studies have demonstrated that the activation of the transcriptional factor nuclear factor kappaB (NF-kappaB) plays an essential role in preventing apoptotic cell death. In this study, to better understand the mechanism responsible for the p53-mediated apoptosis, the effect of wild-type p53 (wt-p53) gene transfer on nuclear expression of NF-kappaB was determined in human colon cancer cell lines. A Western blot analysis of nuclear extracts demonstrated that NF-kappaB protein levels in the nuclei were suppressed by the transient expression of the wt-p53 in a dose-dependent manner. Transduced wt-p53 expression increased the cytoplasmic expression of I kappaB alpha as well as its binding ability to NF-kappaB, thus markedly reducing the amount of NF kappaB that translocated to the nucleus. The decrease in nuclear NF-kappaB protein correlated with the decreased NF-kappaB constitutive activity measured by electrophoretic mobility shift assay. Furthermore, parental cells transfected with NF-kappaB were better protected from cell death induced by the wt-p53 gene transfer. We also found that the wt-p53 gene transfer was synergistic with aspirin (acetylsalicylic acid) in inhibiting NF-kappaB constitutive activity, resulting in enhanced apoptotic cell death. These results suggest that the inhibition of NF-kappaB activity is a plausible mechanism for apoptosis induced by the wt-p53 gene transfer in human colon cancer cells and that anti-NF-kappaB reagent aspirin could make these cells more susceptible to apoptosis. PMID- 10698491 TI - RAS oncogene activation induces proliferation in normal human thyroid epithelial cells without loss of differentiation. AB - Neoplastic transformation of rodent thyroid epithelial cell lines by mutant RAS genes has been widely studied as an experimental model of oncogene-induced loss of tissue-specific differentiation. However, separate evidence strongly implicates RAS mutation as an early event in human thyroid tumour development at a stage prior to loss of differentiation. To resolve this controversy we examined the short- and long-term responses of normal human thyroid epithelial cells to mutant RAS introduced by micro-injection and retroviral transduction respectively. In both cases, expression of RAS at a level sufficient to induce rapid proliferation did not lead to loss of differentiation as shown by expression of cytokeratin 18, E-cadherin, thyroglobulin, TTF-1 and Pax-8 proteins. Indeed, RAS was able to prevent, and to reverse, the loss of thyroglobulin expression which occurs normally in TSH-deficient culture medium. These responses were partially mimicked by activation of RAF, a major RAS effector, indicating involvement of the MAP Kinase signal pathway. The striking contrast between the effect of mutant RAS on differentiation in primary human, compared to immortalized rodent, epithelial cultures is most likely explained by the influence of additional co-operating abnormalities in the latter, and highlights the need for caution in extrapolating from cell line data. PMID- 10698492 TI - EAP1/Daxx interacts with ETS1 and represses transcriptional activation of ETS1 target genes. AB - ETS1 is a member of the evolutionarily conserved family of ets genes, which are transcription factors that bind to unique DNA sequences, either alone or by association with other proteins. In this study, we have used the yeast two-hybrid system to identify an ETS1 interacting protein. The ETS1 N-terminal amino acid region was used as bait and an interaction was identified with the Daxx protein, referred to as EAP1 (ETS1 Associated Protein 1)/Daxx. This interactin has been shown to exist in yeast and in vitro. EAP1/Daxx and ETS1 are co-localized in the nucleus of mammalian cells. The region in EAP1/Daxx which specifically binds to ETS1 is located within its carboxy terminal 173 amino acid region. The ETS1 interaction region is located within its N-terminal 139 amino acids and is referred as the Daxx Interaction Domain (DID). The DID appears to be conserved in several other ets family members, as well as in other proteins known to interact with Daxx. The EAP1/Daxx interacts with both isoforms of ETS1, p51-ETS1 and p42 ETS1. Interaction of EAP1/Daxx with ETS1 causes the repression of transcriptional activation of the MMP1 and BCL2 genes. The interaction domains of both ETS1 and EAP1/Daxx are required for this repression and deletion of either domain abolishes this activity. PMID- 10698493 TI - Mapping of the 7q31 subregion common to the small chromosome 7 derivatives from two sporadic papillary renal cell carcinomas: increased copy number and overexpression of the MET proto-oncogene. AB - Molecular cytogenetic analysis of several sporadic papillary renal cell carcinomas and of their xenografts in immunodeficient mice had previously allowed us to delimit a minimal overrepresented region of chromosome 7 shared by all of them to band 7q31. We have refined the location of the overlapping region to the junction of the subbands 7q31.2 and 7q31.3 by reverse painting with two differently labelled probes prepared from the small chromosome 7 derivatives microdissected from the cells of two distinct tumours. This small region was shown to contain the MET proto-oncogene, present at three to four copies per cell as determined by Southern blot analysis. The increased copy number of the MET gene was found to be associated with its overexpression at the mRNA level. However, no change in MET copy number or expression level was observed in the cells from two xenografted tumours serially transplanted into immunodeficient mice, as compared to those from the corresponding initial tumours. Our results indicate that expression of the MET proto-oncogene above a critical threshold is required for the maintenance of the tumorigenic phenotype of at least some papillary renal cell carcinomas, but does not further increase during tumour progression. PMID- 10698494 TI - Constitutive expression of the DNA-binding domain of Ets1 increases endothelial cell adhesion and stimulates their organization into capillary-like structures. AB - We previously reported that the Ets1 transcription factor is expressed in endothelial cells during angiogenesis both in normal and pathological development. We analyse here the effects of the stable expression of an Ets transdominant negative mutant (Ets1-DB), consisting in an Ets1 protein lacking its transactivation domain. A retrovirus containing the Ets1-DB sequence fused to an IRES-Neo sequence was designed and used to infect brain capillary (IBE) and aorta (MAE) mouse endothelial cell lines. Cells expressing this Ets1 mutant were examined for proliferation, migration and adhesion. Consistent changes were observed on cell morphology, with increased spreading and modifications in the organization of the cytoskeleton, and increased cell adhesion. We investigated the ability of endothelial cells to organise into capillary-like structures using three-dimensional gels. On Matrigel, all endothelial cell lines formed a cord like network within 24 h, with an increased ability of Ets1-DB cells to spread on this substrate. In long term cultures, IBE cells expressing Ets1-DB showed a higher capacity to form branched structures; this effect was potentiated by FGF2. These results demonstrate a role of the Ets transcription factors in the regulation of the adhesive and morphogenetic properties of endothelial cells. PMID- 10698495 TI - Sensitivity to myc-induced apoptosis is retained in spontaneous and transplanted lymphomas of CD2-mycER mice. AB - To study the effects of the Myc oncoprotein in a regulatable in vivo system, we generated lines of transgenic mice in which a tamoxifen inducible Myc fusion protein (c-mycER) is expressed under the control of the CD2 locus control region. Activation of the Myc oncoprotein resulted in both proliferation and apoptosis in vivo. Lines with a high transgene copy number developed spontaneous lymphomas at low frequency, but the tumour incidence was significantly increased with tamoxifen treatment. Surprisingly, we found that cellular sensitivity to Myc induced apoptosis was retained in tumours from these mice and in most lymphoma cell lines, even when null for p53. Resistance to Myc-induced apoptosis could be conferred on these cells by co-expression of Bcl-2. However, acquired resistance is clearly not an obligatory progression event as sensitivity to apoptosis was retained in transplanted tumours in athymic mice. In conclusion, lymphomas arising in CD2-mycER mice retain the capacity to undergo apoptosis in response to Myc activation and show no phenotypic evidence of the presence of an active dominant inhibitor. PMID- 10698496 TI - H-, K- and N-Ras inhibit myeloid leukemia cell proliferation by a p21WAF1 dependent mechanism. AB - Mutated ras genes are frequently found in human cancer. However, it has been shown that oncogenic ras inhibits growth of primary cells, through pathways involving p53 and the cell cycle inhibitors p16INK4a and p19ARF. We have analysed the effect of the ectopic expression of the three mammalian ras genes on the proliferation of K562 leukemia cells, which are deficient for p53, p16INK4a, p15INK4b and p19ARF genes. We have found that high expression levels of both wild type and oncogenic H-, K- and N-ras inhibit the clonogenic growth of K562 cells. Induction of H-rasV12 expression in K562 transfectants retards growth and this effect is accompanied with an increase of p21WAF1 mRNA and protein levels. Furthermore, p21WAF1 promoter is activated potently by oncogenic ras and less pronounced by wild-type ras. This induction is p53-independent since a p21WAF1 promoter devoid of the p53 responsive elements is still activated by Ras. Finally, inhibition of p21WAF1 expression by an antisense construct partially overcomes the growth inhibitory action of oncogenic H-ras. Altogether, these results indicate that the antiproliferative effect of ras in myeloid leukemia cells is associated to the induction of p21WAF1 expression and suggest the existence of p19ARF and p16INK4a-independent pathways for ras-mediated growth inhibition. PMID- 10698497 TI - EGR-1 enhances tumor growth and modulates the effect of the Wilms' tumor 1 gene products on tumorigenicity. AB - The Wilms' tumor 1 gene (WT1) encodes a transcription factor of the zinc-finger family and is homozygously mutated or deleted in a subset of Wilms' tumors. Through alternative mRNA splicing, the gene is expressed as four main polypeptides that differ by a stretch of 17 amino acids just N-terminal of the four zinc-fingers and three amino acids between zinc fingers 3 and 4. We have previously shown that expression of the WT1(-/-) isoform, lacking both inserts, increases the tumor growth rate of the adenovirus-transformed baby rat kidney (AdBRK) cell line 7C3H2, whereas expression of the WT1(-/+) isoform, lacking the 17aa insert, strongly suppresses the tumorigenic phenotype. In the present study we show that expression of these splice variants does not affect the tumorigenic potential of the similar AdBRK cell line, 7C1T1. In contrast to the 7C3H2 cell line, this AdBRK cell line expresses high endogenous levels of EGR-1 (early growth response-1) protein, a transcription factor structurally related to WT1. Ectopic expression of EGR-1 in the 7C3H2 AdBRK cells significantly increases their in vivo growth rate and nullifies the tumor suppressor activity of the WT1( /+) protein. Furthermore, we find that EGR-1 levels are elevated in some Wilms' tumors. These data are the first to show that EGR-1 overexpression causes enhanced tumor growth and that WT1 and EGR-1 exert antagonizing effects on growth regulation in baby rat kidney cells, which might reflect the situation in some Wilms' tumors. PMID- 10698498 TI - v-Abl utilizes multiple mechanisms to drive G1/S progression in fibroblasts. AB - Transformation of 3T3 fibroblasts by the v-Abl tyrosine kinase replaces mitogenic and adhesion signals normally required for cell cycle progression. A 3T3 cell line conditionally transformed with v-Abl has been used to study v-Abl's effects on cell cycle in the context of either serum depletion or absence of adhesion signals. We show that E2F-dependent mRNAs, encoding proteins required for cell cycle progression, are induced by v-Abl. In addition, we identify two previously unknown targets of v-Abl signaling: (1) cyclin D1 and D2 mRNAs are induced upon v Abl activation; and (2) the CDK inhibitor p27 is decreased upon v-Abl activation. PMID- 10698499 TI - Oncogenic epidermal growth factor receptor mutants with tandem duplication: gene structure and effects on receptor function. AB - A number of epidermal growth factor receptor (EGFR) deletion mutants have been identified in gliomas, in which the EGFR gene is frequently amplified and rearranged. We have previously characterized the structure of a gene in A-172 human glioma cells that encodes a 190-kDa EGFR mutant with tandem duplication of the tyrosine kinase (TK) and calcium-mediated internalization (CAIN) domains. Here we describe a 185-kDa tandem duplication mutant (TDM) that is expressed in KE and A-1235 glioma cells, along with certain functional characteristics of the mutants. The corresponding transcripts in KE and A-1235 cells contain 1053 additional nucleotides representing an in-frame duplication of exons 18 through 25 which encode the entire TK region and a portion of the CAIN domain. As with duplication of the entire TK/CAIN region (exons 18-26) in A-172 cells, duplication of exons 18-25 is associated with a specific genomic rearrangement between flanking introns. Involved introns contain homology to recombination signal sequence (RSS) heptamers present in the V(D)J region of the T lymphocyte receptor gene. In defined medium, both oncogenic TDM are constitutively autophosphorylated and inefficiently downregulated. High-affinity binding is reduced in EGFR.TDM/18-26, although the t1/2 of receptor internalization is not prolonged. PMID- 10698500 TI - Human papillomavirus E7 proteins stimulate proliferation independently of their ability to associate with retinoblastoma protein. AB - Studies on human papillomavirus type 16 have demonstrated that the product of the early gene, E7, plays a key role in the immortalization and malignant transformation of the host cell. Several of the biological activities of HPV16 E7 are mediated by inactivation of the members of the pocket protein family, pRb, p107 and p130. In this study, we have characterized the in vitro properties of five E7 proteins from benign and malignant HPV types (10, 32, 48, 54, 77). We show that these E7 proteins associate with pRb and p107 with different efficiencies. All E7s increased the proliferative rate of immortalized rodent fibroblasts cultured in 10% calf serum containing medium. This property is completely independent of their ability to associate with the pocket proteins. Furthermore, all E7s, except HPV10 E7, stimulate G1/S progression and activated the cyclin E and cyclin A promoter in the absence of growth factors. This activity also does not correlate with the E7-efficiency of binding the pocket proteins. Together these data provide evidence that different E7s alter the regulation of the cell cycle by diverse mechanism(s). Finally, this comparative analysis of the different E7 proteins demonstrates that the oncogenicity of a HPV type is not determined by the ability of E7 to associate with the pocket proteins. PMID- 10698501 TI - Functional impairment of p73 and p51, the p53-related proteins, by the human T cell leukemia virus type 1 Tax oncoprotein. AB - We have previously demonstrated that the human T-cell leukemia virus type 1 (HTLV 1) Tax oncoprotein represses the trans-activation function of p53 tumor suppressor protein. Recently, several proteins with sequence homology to p53 have been identified. In this study, we demonstrated that Tax represses the trans activation functions of p73alpha, p73beta, and p51A, the p53-related proteins, as well as p53. Moreover, a mutant Tax of coactivator CBP-binding site (K88A), which activated NF-kappaB but not CREB pathway, could not repress the p73 nor p51 trans activation functions, indicating that CBP-binding domain of Tax is essential for the suppression of their functions. Using proteins of Gal4-fused N-terminal region of p73 and p51, we showed that Tax-mediated inactivation of p73 or p51 requires for their N-terminal trans-activation domains. Furthermore, only the putative N-terminal trans-activation domains of them did not have enough transcriptional activities and their adjacent regions are essential for their full trans-activation, suggesting the existence of their second trans-activation subdomains. Thus, HTLV-1 Tax inactivated the p53-related proteins through their N terminal trans-activation domains. PMID- 10698502 TI - p73 transcriptional activity increases upon cooperation between its spliced forms. AB - The p53 homologue p73 efficiently activates p53-responsive genes. The well documented over-expression of p73 spliced forms in a wide variety of tumor types promoted us to elucidate the mechanisms underlying p73-mediated transcription. Using the luciferase reporter gene driven by Mdm2-minimal promoter in p53 null cells, we demonstrate that the weak transcriptional activity mediated by p73alpha was increased by the mutant form p73beta292, which by itself is transcriptionally inactive. Similarly, cooperation between p73beta and an inactive form of p73alpha increased p73beta-mediated transcriptional activities. Conversely, p73beta elicited a silencing effect on a gain of function mutant, p53(281), which by itself mediated efficient transactivation of the MDR promoter. Neither anisomycin nor actinomycin D altered p73-mediated transcriptional activities, whereas sorbitol profoundly inhibited them through a rapid proteasome-dependent degradation of p73. Our observations point to plausible scenarios in which p73, through cooperation between p73 spliced forms and suppression of gain of function mutant p53 may elicit changes in the transcription of p53 target genes that play key roles in cell growth and death. PMID- 10698503 TI - Somatic mutations and genetic polymorphisms of the PPP1R3 gene in patients with several types of cancers. AB - Recently, we found nonsense and missense mutations of the PPP1R3 (protein phosphatase 1, regulatory subunit 3) gene in diverse human cancer cell lines and primary lung carcinomas, indicating that PPP1R3 functions as a tumor suppressor in human carcinogenesis. In this study, to assess the prevalence of PPP1R3 mutations in human primary cancers and the genetic diversity of the PPP1R3 gene in the human population, somatic mutations and genetic polymorphisms in the PPP1R3 gene were examined in 137 pairs of cancerous and non-cancerous tissues of patients with cancers of colon, ovary, and liver. Five somatic mutations including two missense mutations were detected in three cancerous tissues consisting of two colorectal carcinomas and one ovarian carcinoma. Five novel single nucleotide polymorphisms (SNPs) associated with the substitution of amino acids were also identified in cancer patients, in addition to five known nonsynonymous SNPs, including three previously reported ones as having an impact on the susceptibility to insulin resistant disorders. Differences in the activities and properties of multiple PPP1R3 proteins, which are produced in human cells due to variable somatic mutations and genetic polymorphisms in the PPP1R3 gene, can be involved in human carcinogenesis and susceptibility to diseases. PMID- 10698504 TI - Mutant envelope residues confer a transactivation function onto N-terminal sequences of the v-Rel oncoprotein. AB - The retroviral oncoprotein v-Rel is a member of the Rel/ NF-kappaB family of transcription factors. v-Rel has multiple changes as compared to the proto oncoprotein c-Rel, and these changes render v-Rel highly oncogenic in avian lymphoid cells. Previous results have shown that three mutant residues in the eleven helper virus-derived Envelope (Env) amino acids (aa) at the N-terminus of v-Rel are required for its full oncogenicity. In this report, we show that these mutant Env aa also enable sequences in the N-terminal half of v-Rel to activate transcription in yeast and chicken cells, under conditions where the analogous sequences from c-Rel either do not or only weakly activate transcription. Removal of the Env aa from v-Rel or site-directed mutations that revert the three mutant residues to the residues present in the Rev-A helper virus Env protein abolish this transactivation ability of v-Rel. Addition of mutant Env aa onto c-Rel is not sufficient to fully restore the transactivation function; other sequences in the N-terminal half of v-Rel are needed for full transactivating ability. A C terminally-truncated form of NF-kappaB p100 (p85), produced in HUT-78 human leukemic cells, also activates transcription in yeast, under conditions where the normal p52 and p100 proteins do not. Furthermore, transcriptional activation by p85 in yeast is likely to occur through N-terminal sequences. Taken together, these results are consistent with a model in which transactivation by N-terminal Rel Homology (RH) domain sequences in oncogenic Rel family proteins is influenced by sequences outside the RH domain. PMID- 10698505 TI - HoxA9-mediated immortalization of myeloid progenitors requires functional interactions with TALE cofactors Pbx and Meis. AB - Specific Hox genes are implicated in leukemic transformation, and their selective genetic collaboration with TALE homeobox genes, Pbx and Meis, accentuates their oncogenic potential. The molecular mechanisms underlying these coordinate functions, however, have not been characterized. In this study, we demonstrate that HoxA9 requires its Pbx interaction motif as well as its amino terminus to enhance the clonogenic potential of myeloid progenitors in vitro. We further show that HoxA9 forms functional trimeric DNA binding complexes with Pbx and Meis-like proteins on a modified enhancer. DNA binding complexes containing HoxA9 and TALE homeoproteins display cooperative transcriptional activity and are present in leukemic cells. Trimeric complex formation on its own, however, is not sufficient for HoxA9-mediated immortalization. Rather, structure-function analyses demonstrate that domains of HoxA9 which are necessary for cellular transformation are coincident with those required for trimer-mediated transcriptional activation. Furthermore, the amino terminus of HoxA9 provides essential transcriptional effector properties and its requirement for myeloid transformation can be functionally replaced by the VP16 activation domain. These data suggest that biochemical interactions between HoxA9 and TALE homeoproteins mediate cellular transformation in hematopoietic cells, and that their transcriptional activity in higher order DNA binding complexes provides a molecular basis for their collaborative roles in leukemogenesis. PMID- 10698506 TI - High expression of Survivin, mapped to 17q25, is significantly associated with poor prognostic factors and promotes cell survival in human neuroblastoma. AB - Survivin (SVV) is a family member of inhibitor of apoptosis proteins (IAPs) and its expression is cell cycle regulated. The gene is mapped to chromosome 17q25, the region of which is frequently gained in advanced stages of neuroblastoma (NBL). However, the role of SVV in NBL is poorly understood. Here we studied the clinical and biological role of SVV in NBL. A 1.9 kb SVV transcript was expressed in all of 9 NBL cell lines at higher levels than those in adult cancer cell lines. In 34 primary NBLs, high levels of SVV expression was significantly associated with age greater than 12 months (two sample t-test: P= 0.0003), advanced stages (P = 0.0136), sporadic tumors (P= 0.0027) and low levels of TrkA expression (P = 0.0030). In NBL cell lines, SVV mRNA expression was dramatically down-regulated in CHP134 and IMR32 cells undergoing apoptosis after treatment with all-trans retinoic acid (RA) or serum deprivation. It was only moderately decreased in cells (SH-SY5Y and CHP901) undergoing RA-induced differentiation. On the other hand, in proliferating NBL cells or RA-treated SK-N-AS line which is refractory to RA, the SVV mRNA remained at steady state levels or rather up regulated. Furthermore, transfection of SVV into CHP134 cells induced remarkable inhibition of the RA-induced apoptosis. Collectively, our results suggest that high expression of SVV is a strong prognostic indicator for the advanced stage neuroblastomas, and that it could be one of the candidate genes for the 17q gain. PMID- 10698507 TI - Tandem-duplicated Flt3 constitutively activates STAT5 and MAP kinase and introduces autonomous cell growth in IL-3-dependent cell lines. AB - We have recently identified an internal tandem duplication of the human Flt3 gene in approximately 20% of acute myeloid leukemia (AML) cases. In the present study, the wild-type and the mutant Flt3 genes were transfected into two IL-3-dependent cell lines, 32D and BA/F3 cells. Mutant Flt3-transfected cells exhibited autonomous growth while wild-type Flt3-transfected cells with the continuous stimulation of Flt3 ligand exhibited a minimal proliferation. Cells expressing mutant Flt3 showed constitutive activation of STAT5 and MAP kinase. In contrast, Flt3 ligand stimulation caused rapid activation of MAP kinase but not STAT5 in cells expressing wild-type Flt3. Finally, we found constitutive activation of MAP kinase and STAT5 in all clinical samples of AML patients with mutant Flt3. Our study shows the significance of internal tandem duplication of Flt3 receptors for leukemia cell expansion. PMID- 10698508 TI - Dissociation of p53-mediated suppression of homologous recombination from G1/S cell cycle checkpoint control. AB - The tumor suppressor p53 is considered as the guardian of the genome which is activated following genotoxic stress. In many cell types, p53 mediates G1 cell cycle arrest as the predominant cellular response. Inactivation of wild-type p53 leads to loss of G1/S checkpoint control and to genomic instability, including increased spontaneous homologous recombination (HR). To determine whether regulation of the G1/S checkpoint is required for suppression of HR, we assessed recombination events using a plasmid substrate that stably integrated into the genome of p53-null mouse fibroblasts. Exogenous expression of a temperature sensitive p53 protein (Ala135 to Val), which had lost trans-activation function and could not regulate G1/S transition when in mutant conformation, reduced HR rates to the same extent as wild-type p53. Furthermore, a p53 construct with an alternatively-spliced carboxy terminus also retained this ability in the absence of both activities, G1/S control and non-sequence specific DNA binding as mediated by the carboxy terminus. Our data dissociate regulation of HR by p53 from its role as a cell cycle checkpoint protein. The results support a model which extends p53's role as a guardian of the genome to include transactivation independent regulatory functions in DNA repair, replication and recombination. PMID- 10698509 TI - Neu differentiation factor/heregulin induction by hepatocyte and keratinocyte growth factors. AB - Hepatocyte growth-factor (HGF) is a potent, widely produced, pleiotropic mediator of mesenchymal-epithelial interaction. In a study of changes in gene expression initiated by HGF in Balb/MK keratinocytes, we observed the induction of Neu differentiation factor (NDF) mRNA (also known as heregulin, or HRG). Further characterization of the regulation of NDF expression in Balb/MK keratinocytes revealed potent induction by keratinocyte growth factor (KGF) and epidermal growth factor (EGF), but not by HGF/NK2, an alternative HGF isoform with motogenic but not mitogenic or morphogenic activities. Sustained treatment (8 h) of Balb/MK cells with KGF stimulated secretion of mature NDF protein into the culture medium, and Balb/ MK cells treated with purified recombinant NDF protein showed increased DNA synthesis. We also found evidence of NDF induction in two models of tissue repair in mice: in full-thickness skin wounds, following locally increased KGF production, and in kidney after partial hepatectomy, following elevation of circulating HGF levels. These results reveal that mesenchymally derived HGF and KGF can activate autocrine NDF signaling in their epithelial targets, and suggest that this mechanism contributes to the coordination of stages of wound repair, and possibly development, where these growth factors act in concert to direct epithelial proliferation, morphogenesis and differentiation. PMID- 10698510 TI - Tissue and cell-specific expression of the p53-target genes: bax, fas, mdm2 and waf1/p21, before and following ionising irradiation in mice. AB - The mechanisms by which the p53 tumour suppressor protein would, in vivo, co ordinate the adaptive response to genotoxic stress is poorly understood. p53 has been shown to transactivate several genes that could be involved in two main cellular responses, growth arrest and apoptosis. To get further insight into the tissue-specific regulation of p53 transcriptional activity, we performed an extensive study looking at the expression of four well characterized p53 responsive genes, before and after gamma-irradiation in p53 wild-type (p53+/+) and p53-deficient (p53-/-) mice. The waf1, bax, fas and mdm2 genes were chosen for their different potential roles in the cellular response to stress. Our data demonstrate the strict p53-dependence of mRNA up-regulation for bax, fas and mdm2 in irradiated tissues and confirm such findings for waf1. They further highlight complex levels of regulatory mechanisms that could lead, in vivo, to selective transcriptional activation of genes by p53. In addition, our results provide arguments for the involvement of p53 in the basal mRNA expression of the four genes in some organs. Finally, in situ expression of Bax and p21Waf-1 protein suggests, at least in lymphoid organs, a direct correlation between selective p53 target gene expression and a particular response of a cell to ionising radiation. PMID- 10698511 TI - Anti-sense oligonucleotide of p21(waf1/cip1) prevents interleukin 4-mediated elevation of p27(kip1) in low grade astrocytoma cells. AB - Elevation of the cyclin-dependent kinase (cdk) inhibitor, p27(kip1) is necessary for Interleukin (IL)-4-mediated growth arrest of human low grade astrocytoma (RTLGA) cells and occurs at 24 h of treatment. Pathways involved in IL4 alteration of p27(kip1) are unknown, however. Here we investigated whether other cdk inhibitors contributed to the actions of IL-4 on RTLGA cells. By 12 h of IL-4 treatment, both cdk4 and cdk2 kinase activities against the retinoblastoma protein (pRb) were reduced and nuclear entry of pRb was prohibited. Twelve-hour cdk complexes contained elevated p21(waf1/cip1) but not p27(kip1), p15(ink4B) or p16(ink4A). IL-4 increased p21(waf1/cip1) but not p27(kip1) mRNA levels, and stimulated luciferase activity of a p21(waf1/cip1) promoter-luciferase reporter. In p53-mutant WITG3 cells, IL-4 did not alter p21(waf1/cip1) mRNA and promoter luciferase activity or p27(kipl) protein, suggesting a need for functional p53. STAT6 phosphorylation by IL-4, however, occurred in both p53-mutant WITG3 and p53 functional RTLGA cells. Pre-treatment of RTLGA with anti-sense but not missense p21(waf1/cip1) oligonucleotide prior to IL-4: (a) restored cdk activities; (b) reduced cdk4-associated p21(waf1/cip1) levels; (c) prevented p27(kipl) elevation; and (d) reversed growth arrest. These results are the first to suggest that p21(waf1/cip1) is essential for IL-4-mediated elevation of p27(kip) and growth arrest of astrocytoma cells. PMID- 10698512 TI - Involvement of p27Kip1 in G1 arrest by high dose 5 alpha-dihydrotestosterone in LNCaP human prostate cancer cells. AB - The cell cycle is governed by cyclin dependent kinases (cdks), which are activated by binding of cyclins, inhibited by cdk inhibitors and regulated by phosphorylation and dephosphorylation. Exposure to high dose dihydrotestosterone (DHT) inhibits population growth of the human prostate carcinoma cell line, LNCaP. To determine the mechanism of growth arrest by high dose DHT, we assayed the changes in cell cycle profile and the cell cycle regulators that mediate these effects. Treatment of asynchronously growing LNCaP cells with 100 nM DHT caused a G1 arrest. The proportion of cells in S phase fell from 22 to 2%, while the G1 fraction rose from 74 to 92% by 24 h. Loss of phosphorylation of the retinoblastoma protein was noted and cdk4 and cyclin E/ cdk2 activities fell. Inhibition of these G1 cyclin dependent kinases was not due to loss of either cyclin or cdk proteins nor to increases in the cdk inhibitors p16INK4A and p21CiP1. p21Cip1 protein levels remained constant, and cyclin E-associated p21CiP1 fell, suggesting that p21CiP1 is not relevant to this form of cyclin E/cdk2 inhibition. Of note, total p27KiP1 levels and cyclin E-associated p27Kip1 increased as cells arrested and the amount of the CAK activated cdk2 bound to cyclin E decreased. p27KiP1 immunodepletion experiments demonstrated that the DHT mediated increase in p27Kip1 was sufficient to fully saturate and inhibit target cyclin E/ cdk2. The inhibition of cyclin E/cdk2 by p27Kip1 contributes to G1 arrest of LNCaP following high dose DHT. p27KiP1 may be a key effector of androgen dependent growth modulation in prostate cancer cells. PMID- 10698513 TI - Inhibition of H-Ras transformation by the PTEN/MMAC1/TEP1 tumor suppressor gene. AB - The human PTEN/MMAC1/TEP1 (PTEN) tumor suppressor gene encodes a phosphatase with specificity towards the D3 phosphate of phosphatidylinositides. PTEN mutations have been reported in the endometrioid type of uterine tumors which are associated with frequent activations of the Ras oncogenes. In this study, we report the ability of PTEN to potently inhibit H-Ras induced morphological transformation and anchorage-independent growth in NIH3T3 cells. This novel activity of PTEN was correlated more with its ability to suppress the phosphatidylinositol 3-kinase (PI3-K)-dependent signaling cascade, but not the mitogen-activated protein kinase (MAPK) pathway. To define the minimal region in PTEN protein that is responsible for this anti-oncogenic activity, a panel of carboxyl-terminal truncation mutants was generated. While deletions of 4 and 33 amino acids do not have marked effects, removal of up to 68 amino acids drastically reduced the ability of PTEN to inhibit Ras transformation. The propensity of these mutants to suppress Ras transformation is correlated with their relative ability to dephosphorylate inositol (1,3,4,5)-tetrakisphosphate in vitro, and to suppress Akt kinase activity in cultured cells. In addition, we have evidence to suggest that the C-terminal region of PTEN contributes to the stability of the encoded gene product. PMID- 10698514 TI - [Lys61]N-Ras is able to induce full activation and nuclear accumulation of Cdk4 in NIH3T3 cells. AB - The elements of the cell cycle regulatory machinery activated by the oncogenic form of Ras, [Lys61]N-Ras, have been analysed in NIH3T3 cells. We demonstrate that [Lys61]N-Ras expression is able to induce full cdk4 activation. As already reported, oncogenic Ras expression was sufficient to induce cyclin D1 and p21cip1 expression and their association with cdk4. Furthermore, serum-starved [Lys61]N Ras NIH3T3 cells showed nuclear accumulation of cyclin D1 and cdk4 not observed in serum-starved NIH3T3 cells. This accumulation of cdk4 into the cell nucleus observed in serum-starved [Lys61]N-Ras NIH3T3 cells was inhibited by a microinjection of neutralizing anti-Ras antibodies. Thus, active [Lys61]N-Ras was a sufficient signal to induce nuclear accumulation of cyclin D1/cdk4, leading to its full activation. Transfection of [Lys61]N-Ras NIH3T3 cells with an inactive form of MEK or their treatment with PD 98059, showed that nuclear translocation of cdk4 was MEK dependent. Interestingly, cells constitutively expressing [Lys61]N-Ras did not inactivate pRb and did not proliferate in the absence of serum. This may be due to the fact that although association of cdk2 with cyclin E and the translocation of those complexes to the nucleus were achieved, [Lys61]N Ras expression was not sufficient to induce cdk2 activation. The high levels of p27(kip1) that were found in cyclin E/cdk2 complexes may be responsible for the inability of oncogenic Ras to activate this kinase. In consequence, oncogenic alterations that lead to a decrease in p27kip1 bound to cyclin E may cooperate with Ras to induce full cdk2 activation, pRb inactivation and thus cell proliferation. PMID- 10698515 TI - Epstein-Barr virus EBNA3C can disrupt multiple cell cycle checkpoints and induce nuclear division divorced from cytokinesis. AB - Expression of EBNA3C is essential for the immortalization of B cells by EBV in vitro and, in co-operation with activated ras, EBNA3C has oncogenic activity in primary rodent fibroblasts. This suggested that this viral oncoprotein might disrupt the cyclin/CDK-pRb-E2F pathway, which regulates cell cycle progression at the restriction point (R-point) in G1 of the proliferation cycle. An assay was established in which transfected EBNA3C-positive cells could be sorted and simultaneously analysed for their distribution in the cell cycle. This revealed that in NIH3T3 fibroblasts compelled to arrest by serum-withdrawal, EBNA3C induces nuclear division that is often divorced from cytokinesis and so produces bi- and multinucleated cells. This was confirmed using the ecdysone-inducible system for expression of EBNA3C in human U2OS cells and by microinjection of expression vectors into NIH3T3 and U2OS. Further analysis revealed that in the inducible system, EBNA3C expression inhibits the accumulation of p27(K1P1) but not the dephosphorylation of pRb. Experiments using the microtubule destabilizing drug nocodazole, showed that EBNA3C could abrogate the mitotic spindle checkpoint. PMID- 10698516 TI - KFC, a Ste20-like kinase with mitogenic potential and capability to activate the SAPK/JNK pathway. AB - The Sterile-20 (Ste20) family of serine-threonine kinases has been implicated in the activation of the stress-activated protein kinase pathways. However, the physiological role has remained ambiguous for most of the investigated mammalian Ste20's. Here we report the cloning of a novel Ste20-like kinase, from chicken embryo fibroblast (CEF) cells, which we have named KFC, for Kinase From Chicken. The 898 amino acid full-length KFC protein contains an amino-terminal kinase domain, an adjacent downstream serine-rich region, and a C-terminal tail containing a coiled-coil domain. Here we show that the coiled-coil domain of KFC negatively regulates the intrinsic kinase activity. We have also identified a splice variant of KFC in which there is a 207 nucleotide in-frame deletion. This deletion of 69 amino acids encompasses the serine-rich region. These two isoforms, called KFCL, for full-length, and KFCS for spliced (or short) form, not only differ in structure, but also in biological properties. Stable CEF cells overexpressing KFCL, but not KFCS, have a significant increase in growth rate when compared to parental cells. This mitogenic effect is the first such reported for this family of kinases. Finally, we found that KFC, when activated by truncation of the regulatory C-terminus, has a specific activation of the stress activated protein kinase (SAPK/JNK) pathway. PMID- 10698517 TI - Role of the ATPase domain of the Cockayne syndrome group B protein in UV induced apoptosis. AB - Cockayne syndrome (CS) is a human autosomal recessive disorder characterized by many neurological and developmental abnormalities. CS cells are defective in the transcription coupled repair (TCR) pathway that removes DNA damage from the transcribed strand of active genes. The individuals suffering from CS do not generally develop cancer but show increased neurodegeneration. Two genetic complementation groups (CS-A and CS-B) have been identified. The lack of cancer formation in CS may be due to selective elimination of cells containing DNA damage by a suicidal pathway. In this study, we have evaluated the role of the CSB gene in UV induced apoptosis in human and hamster cells. The hamster cell line UV61 carries a mutation in the homolog of the human CSB gene. We show that both human CS-B and hamster UV61 cells display increased apoptotic response following UV exposure compared with normal cells. The increased sensitivity of UV61 cells to apoptosis is complemented by the transfection of the wild type human CSB gene. In order to determine which functional domain of the CSB gene participates in the apoptotic pathway, we constructed stable cell lines with different CSB domain disruptions. UV61 cells were stably transfected with the human CSB cDNA containing a point mutation in the highly conserved glutamic acid residue in ATPase motif II. This cell line (UV61/ pc3.1-CSBE646Q) showed the same increased apoptosis as the UV61 cells. In contrast, cells containing a deletion in the acidic domain at the N-terminal end of the CSB protein had no effect on apoptosis. This indicates that the integrity of the ATPase domain of CSB protein is critical for preventing the UV induced apoptotic pathway. In primary human CS B cells, the induction and stabilization of the p53 protein seems to correlate with their increased apoptotic potential. In contrast, no change in the level of either p53 or activation of mdm2 protein by p53 was observed in hamster UV61 cells after UV exposure. This suggests that the CSB dependent apoptotic pathway can occur independently of the transactivation potential of p53 in hamster cells. PMID- 10698518 TI - Cofactor competition between the ligand-bound oestrogen receptor and an intron 1 enhancer leads to oestrogen repression of ERBB2 expression in breast cancer. AB - Overexpression of the ERBB2 proto-oncogene in breast tumours, which occurs in 25 30% of patients, correlates with poor prognosis. In oestrogen receptor (ER) positive breast epithelial cells oestrogens reduce ERBB2 mRNA and protein levels, an effect that is reversed in the presence of anti-oestrogens such as tamoxifen and ICI 182780. Our previous studies have shown that the major effect of oestrogen on ERBB2 expression is at the level of transcription and that this is mediated through a region within the ERBB2 first intron which can act as an oestrogen-suppressible enhancer in ER positive breast cells. In vitro footprinting of the smallest DNA fragment that retained full activity revealed four transcription factor binding sites. We report here that two of these sites are recognized by AP-2 proteins and the other two are bound by a variety of bZIP factors, including CREB and ATFI, with a major complex containing ATFa/ JunD. However, by using ER mutants it is clear that repression occurs essentially off the DNA. Indeed, the essential domain of the ER responsible for repression of the ERBB2 enhancer is a region termed AF2 which is required for the ligand-dependent association of non-DNA binding cofactors. We further demonstrate that one of these ER cofactors, SRC-1, can relieve oestrogen repression of the ERBB2 enhancer and conclude that these data fit with a model whereby the ER and the ERBB2 enhancer compete for this limiting, non-DNA binding cofactor. Thus, in oestrogenic conditions SRC-1 preferentially binds to the ER which effectively sequesters it thereby reducing enhancer activity, but in antioestrogenic media the cofactor is released from the ER and is therefore available to activate the ERBB2 enhancer. PMID- 10698519 TI - Activation of beta-catenin in epithelial and mesenchymal hepatoblastomas. AB - Wnt/beta-catenin signaling is frequently activated in cancer cells by stabilizing mutations of beta-catenin or loss-of-function mutations of the APC tumor suppressor gene. We have analysed the role of beta-catenin in the pathogenesis of hepatoblastoma (HB), an embryonic liver tumor occurring mainly in children under 2 years of age. Sequence analysis of the beta-catenin NH2-terminal domain in 18 epithelial and mixed HBs revealed missense mutations in the GSK3beta phosphorylation motif or interstitial deletions in 12 tumors (67%). In the remaining cases, no truncating mutation of APC could be evidenced. Immunohistochemical analysis of beta-catenin in 11 HBs demonstrated nuclear/cytoplasmic accumulation of the protein in all tumors analysed, with predominant nuclear beta-catenin immunostaining in undifferentiated cells. Membranous beta-catenin localization was preserved only in fetal-type tumoral hepatocytes and was associated with E-cadherin expression. Moreover, we show that beta-catenin is aberrantly overexpressed in a large spectrum of tumor components, including hepatocyte-like cells at various differentiation stages and heterologous elements such as squamous, osteoid and chrondroid tissues, and in occasional other mesenchymally-derived cells. These data strongly suggest that activation of beta-catenin signaling is an obligatory step in HB pathogenesis, and raise the possibility that it interferes with developmental signals that specify different tissue types at early stages of hepatic differentiation. PMID- 10698520 TI - Differential effect of subcellular localization of communication impairing gap junction protein connexin43 on tumor cell growth in vivo. AB - There is a large body of evidence suggesting the connexin gap junction proteins appear to act as tumor suppressors, and their tumor inhibitory effect is usually attributed to their main function of cell coupling through gap junctions. However, some cancer cells (e.g. the rat bladder carcinoma BC31 cell line) are cell-cell communication proficient. Using specific site-directed mutagenesis in the third membrane-spanning (3M) domain of connexin43 (Cx43), we abolished the intrinsic gap junction intercellular communication (GJIC) in BC31 cells either by closing the gap junctional channels or by disruption of the transport of connexin complexes to the lateral membrane. Clones of BC31 cells transfected with a dominant negative Cx43 mutant giving rise to gap junctional channels, permeable only for a small tracer (neurobiotin), displayed accelerated growth rate in vivo, showing the critical role of selective gap junctional permeability in the regulation of cell growth in vivo. The use of other dominant-negative mutants of Cx43 also suggested that the effect of impaired communication on the tumorigenicity of cancer cells depends on the subcellular location of connexin. Inhibition of intrinsic GJIC in BC31 cells by sequestering of Cx protein inside the cytoplasm, due to expression of dominant-negative transport-deficient Cx43 mutants, did not significantly enhance the growth of transfectants in nude mice, but occasionally slightly retarded it. In contrast, augmentation of GJIC in BC31 cells by forced expression of wild-type Cx43, or a communication-silent mutant, fully suppressed tumorigenicity of these cells. Overall, these results show that cell coupling is a strong, but not the sole, mechanism by which Cx suppresses growth of tumorigenic cells in vivo; a GJIC-independent activity of Cx proteins should be considered as another strong tumor-suppressive factor. PMID- 10698521 TI - Interleukin-2 expression in human carcinoma cell lines and its role in cell cycle progression. AB - Human carcinomas were shown to express mRNA and protein for IL-2R alpha, beta and gamma chains. Recently, human carcinomas were also shown to constitutively express protein and mRNA for IL-2 in vivo and in vitro. Here we report that the expression levels of cytoplasmic IL-2 as well as IL-2Rbeta- and gamma-chain in human carcinoma cells change during the cell cycle progression. Carcinoma cells synchronized in the G2/M phase of the cell cycle expressed significantly more intracytoplasmic IL-2 as well as IL-2Rbeta and gamma proteins than tumor cells in the G0/G1 phase. The level of mRNA for IL-2 was 5-10-fold higher in the M phase than in the G0/G1-phase, as shown by quantitative competitive RT-PCR. Expression of the cyclin-dependent kinase (CDK) inhibitor p27kip1 in these carcinoma cells was found to be high in the G0/G1 phase, nearly absent in the S phase, and it increased again in the G2/M phase of the cell cycle. In synchronized cells, the decrease in p27 expression coincided with high levels of expression of IL-2. Using the IL-2 specific antisense oligonucleotide to block synthesis of endogenous IL-2 in tumor cells, we observed increased levels of p27 as well as p21. The antisense oligonucleotides specific for p27 or p21 blocked expression of these proteins but not of IL-2. Thus, endogenous IL-2 is important in regulating expression of p27 as well as p21 and, therefore, in controlling cell cycle progression of tumor cells, while its own expression remains independent of the CDK inhibitors. PMID- 10698522 TI - A conditionally-active form of MEK1 results in autocrine tranformation of human and mouse hematopoietic cells. AB - The Raf/MEK/MAP kinase cascade plays a critical role in transducing growth signals from activated cell surface receptors. Using deltaMEK1:ER, a conditionally-active form of MEK1, we demonstrate the ability of this dual specificity protein kinase to abrogate the cytokine-dependency of the human and murine hematopoietic cells lines TF-1, FDC-P1 and FL5.12. Cytokine-independent cells were obtained from TF-1, FDC-P1 and FL5.12 cells at frequencies of 2.5 x 10(-3), 5 x 10(-5) and 10(-7) respectively, indicating that not all cells expressing deltaMEK1:ER were factor-independent. In general, cells that were converted to a cytokine-independent phenotype displayed a higher level of MAP kinase activity in response to deltaMEK1:ER activation than those that remained cytokine-dependent. deltaME-K1:ER-responsive cells could be maintained long-term in the presence of beta-estradiol as well as the estrogen-receptor antagonist 4 Hydroxy-Tamoxifen and the anti-estrogen ICI 164383. Removal of hormone led to the rapid cessation of cell growth in a manner similar to that observed when cytokine is withdrawn from the parental cells. Treatment of deltaMEKI:ER-responsive cells with a specific and selective inhibitor, PD98059, prevented growth in response to beta-estradiol. GM-CSF mRNA transcripts were detected in the MEK1-responsive cells indicating that the activated deltaMEK1:ER may induce a pathway leading to autocrine proliferation. Treatment of MEK1-responsive cells with an anti-GM-CSF antibody, but not a control antibody, suppressed cell growth. The cell lines described here will be useful for elaborating the ability of the MAP kinase pathway to regulate cell proliferation in hematopoietic cells. PMID- 10698523 TI - GSK-3beta-dependent phosphorylation of adenomatous polyposis coli gene product can be modulated by beta-catenin and protein phosphatase 2A complexed with Axin. AB - Axin forms a complex with adenomatous polyposis coli gene product (APC), glycogen synthase kinase-3beta (GSK-3beta), and beta-catenin through different binding sites and downregulates beta-catenin. GSK-3beta-dependent phosphorylation of APC (1211-2075) which has the Axin-binding site was facilitated by Axin, but that of APC-(959-1338) which lacks the Axin-binding site was not. Axin-(298-506) or Axin (298-832), which has the GSK-3beta- and beta-catenin- but not APC-binding sites, did not enhance GSK-3beta-dependent phosphorylation of either APC-(1211-2075) or APC-(959-1338). Furthermore, beta-catenin stimulated the phosphorylation of APC (959-1338) and APC-(1211-2075) by GSK-3beta in the presence of Axin. Consistent with these in vitro observations, expression of beta-catenin or Axin in COS cells promoted an SDS gel band shift of APC. These results indicate that APC complexed with Axin is effectively phosphorylated by GSK-3beta and that beta-catenin may modulate this phosphorylation. In addition, the heterodimeric form of protein phosphatase 2A (PP2A) directly bound to Axin, and PP2A complexed with Axin dephosphorylated APC phosphorylated by GSK-3beta. Taken together, these results suggest that GSK-3beta-dependent phosphorylation of APC can be modulated by beta catenin and PP2A complexed with Axin. PMID- 10698524 TI - DCC and SMAD4 alterations in human colorectal and pancreatic tumor dissemination. AB - Chromosome 18q is lost a high proportion of colorectal and pancreatic cancers. Three candidate tumor suppressor genes, DCC, Smad4 and Smad2 have been identified in this chromosome region. DCC and Smad4 aberrations have been previously identified in pancreatic and colorectal tumors. The aim of this study was to compare the presence of concurrent genetic aberrations in DCC and neighboring Smad4 and Smad2 genes during colorectal and pancreatic distal dissemination. We have used a panel of orthotopically implanted colorectal and pancreatic xenografts and corresponding metastases. We have shown that while LOH at DCC locus occurred at a similar frequency in both tumors, diminished DCC protein expression was exclusively present in colorectal tumors harboring intragenic DCC LOH. In contrast, in pancreatic xenografts loss of DCC protein and mRNA expression was restricted to metastases. Smad4 gene aberrations were detected at a similar frequency in both tumors and were selected for during distal dissemination. Acquisition of alterations in both genes occurred independently. Our results suggest that both DCC and Smad4 contribute to pancreatic and colorectal distal dissemination. However, the role of DCC may differ between both tumor types. PMID- 10698525 TI - Neoplastic transformation by Notch is independent of transcriptional activation by RBP-J signalling. AB - Signalling through the transmembrane receptor Notch is triggered by ligand binding, which induces the proteolytic cleavage of the Notch protein. This cleavage generates an intracellular fragment of the Notch protein (Notch-IC), which translocates into the nucleus and modifies transcription of target genes through its association with the RBP-J protein. Thus, the isolated Notch-IC protein represents the constitutively activated receptor. We have performed a deletion analysis of Notch IC in order to identify the transferable transactivation domain of Notch-IC and the minimal domain of Notch-IC required for RBP-J dependent transactivational activation. Functionally, Notch-IC has been linked to cell fate decision in development and oncogenesis in vivo. In vitro, Notch-IC can cooperate in neoplastic transformation of baby rat kidney cells with the adenoviral E1A protein. We have defined the minimal domain of Notch-IC required for E1A cotransformation. This domain, consisting of the ankyrin repeats of Notch-IC only, can neither activate RBP-J dependent transcription nor does it carry a transactivation domain. Therefore, the ankyrin repeat domain of Notch-IC might trigger novel pathways relevant for transformation but unrelated to RBP-J signalling. PMID- 10698526 TI - Glutamic acid mutagenesis of retinoblastoma protein phosphorylation sites has diverse effects on function. AB - The retinoblastoma tumor suppressor gene (Rb) has many functions within the cell including regulation of transcription, differentiation, apoptosis, and the cell cycle. Regulation of these functions is mediated by phosphorylation at as many as 16 cyclin-dependent kinase (CDK) phosphorylation sites in vivo. The contribution of these sites to the regulation of the various Rb functions is not well understood. To characterize the effect of phosphorylation at these sites, we systematically mutagenized the serines or threonines to glutamic acid. Thirty five mutants with different combinations of modified phosphorylation sites were assayed for their ability to arrest the cell cycle and for their potential to induce differentiation. Only the most highly substituted mutants failed to arrest cell cycle progression. However, mutants with as few as four modified phosphorylation sites were unable to promote differentiation. Other mutants had increased activity in this assay. We conclude that modification of Rb phosphorylation sites can increase or decrease protein activity, that different Rb functions can be regulated independently by distinct combinations of sites, and that the effects of modification at any one site are context dependent. PMID- 10698527 TI - Analysis of mechanisms involved in the prevention of gamma irradiation-induced apoptosis by hGM-CSF. AB - Human granulocyte-macrophage colony-stimulating factor (hGM-CSF) induces proliferation and sustains viability of the mouse interleukin (IL)-3 dependent lymphoid cell line BA/F3 expressing the hGM-CSF receptor. Caspase-3 like enzyme activity and DNA fragmentation were augmented by depletion of this factor from the cell, and exposure to gamma irradiation accelerated kinetics of these events. Anti gamma irradiation-induced apoptosis occurred through various mutant GM-CSF receptors and only the box1 region was essential while the C terminal region, including tyrosine residues which are required for MAPK cascade activation, was dispensable. Consistent with this notion, the addition of PD98059 had no effect on this activity thereby indicating that activation of MAPK is not essential for the activity. As expected, gamma irradiation increased p53 protein and bax mRNA levels and the presence of hGM-CSF dramatically modulated bax/bcl-X(L) ratio. The PI-3K specific inhibitor wortmannin did not affect hGM-CSF dependent anti gamma irradiation induced apoptosis nor bcl-X(L) induction, thus bcl-X(L) but not PI-3K pathway seems to be involved in hGM-CSF dependent anti gamma irradiation-induced apoptosis. It is well documented that the boxl region is essential for GM-CSF dependent activation of JAK2 and JAK2 specific inhibitor AG490 suppressed anti gamma, irradiation-induced apoptosis by hGM-CSF. An artificial JAK2 activating molecule in which extracellular and the transmembrane of beta(c) fused with whole JAK2 can sustain BA/F3 cells survival and proliferation mIL-3 independently, but these cells are susceptible to gamma irradiation. Furthermore GyrB/Jak2, which can activate STAT5 but not the MAPK cascade nor survival of BA/F3 cells, also could not prevent gamma irradiation-induced apoptosis. Although JAK2 is essential for hGM-CSF dependent anti gamma irradiation-induced apoptosis, it appeared that JAK2 does not seem sufficient for the activity. PMID- 10698528 TI - Motility and invasion are differentially modulated by Rho family GTPases. AB - Cell migration in vivo often requires invasion through tissue matrices. Currently little is known regarding the signaling pathways that regulate cell invasion through three-dimensional matrices. The small GTPases Cdc42, Rac and Rho are key regulators of actin cytoskeletal and adhesive structures. We now show that expression of dominant negative forms of either Cdc42, Rac or Rho inhibited PDGF BB-stimulated Rat1 fibroblast invasion into 3D collagen matrices, indicating that the activity of each of these GTPases is necessary for cell invasion. In contrast, only Rac activation was required for PDGF-BB-stimulated locomotion across a planar substrate in the Boyden chamber. Interestingly, PDGF-induced invasion was also strongly inhibited by expression of constitutively active forms of Cdc42 or Rho, and to a lesser extent by constitutively active Rac. We also show that constitutively active V12-Rac significantly stimulated basal Rat1 fibroblast invasion, independent of PI-3-kinase activity, and that this effect was suppressed by the effector mutant V12/H40-Rac. These results suggest that cellular invasion may require an optimal level of activation of Cdc42, Rho and Rac, and that migration and invasion are differentially modulated by Rho family GTPases. PMID- 10698529 TI - E6 proteins from diverse cutaneous HPV types inhibit apoptosis in response to UV damage. AB - In addition to their role in anogenital cancer, human papillomaviruses (HPVs) are also involved in the development of a range of cutaneous lesions. HPV types 5 and 8 are associated with the development of skin cancers in individuals with Epidermodysplasia verruciformis (EV). A broad spectrum of HPV types are also commonly found in non-melanoma skin cancers in immunocompromised individuals, such as organ transplant recipients. The skin cancers in EV and immunocompromised patients occur predominantly at body sites exposed to ultra violet (UV) radiation, pointing to a key role for UV in their development. Here we show that the E6 protein from a range of cutaneous HPV types effectively inhibits apoptosis in response to UV damage. This occurs in both p53 null and wild type cells and does not require p53 degradation. PMID- 10698530 TI - Development and validation of a high-performance liquid chromatographic stability indicating method for the analysis of Synercid in quality control, stability and compatibility studies. AB - A gradient high-performance liquid chromatographic (HPLC) method was developed for the analysis of Synercid freeze-dried powder in routine quality control, stability and compatibility studies. This method is suited for a simultaneous assay of drug substances and impurities. The method was validated for precision, reproducibility, linearity, accuracy and limits of detection. The robustness study that was performed according to an experimental design is described. PMID- 10698531 TI - An LC/MS/MS method for the quantitation of MTIC (5-(3-N-methyltriazen-1-yl) imidazole-4-carboxamide), a bioconversion product of temozolomide, in rat and dog plasma. AB - A sensitive and selective HPLC/electrospray ionization tandem mass spectrometric (LC/ESI/MS/MS) method for the quantitative determination of MTIC (5-(3-N methyltriazen-1-yl)-imidazole-4-carboxamide), a pharmacologically active hydrolysis product of temozolomide, was developed and validated over a linear range from 10 to 400 ng ml(-1) in dog plasma and from 10 to 500 ng ml(-1) in rat plasma. This HPLC method utilized small plasma volumes (70 microl), rapid sample processing, and isocratic elusion conditions to achieve sensitive and selective MS/MS detection. Samples were processed and analyzed one at a time every 4.5 min in order to compensate for the inherent instability of MTIC. Both MTIC and the internal standard DTIC [5-(3,3'-N,N'-dimethyltriazen-1-yl)-imidazole-4 carboxamide] were quantitated in the positive ion, selected reaction monitoring (SRM) mode. The lower limit of quantitation (LLOQ) was 10 ng ml(-1) in the plasma from both species. Inter-assay accuracy and precision of all calibration standards and quality control (QC) samples were within +/- 11 and 12%, respectively, with the exception of the LLOQ in rat plasma (17%). The validated method was used to determine the time dependent plasma concentration of MTIC in rats and dogs following a single oral dose of temozolomide. The standard curve and the quality control data indicate that the method performed acceptably throughout the sample analysis period. PMID- 10698532 TI - Peak fronting in reversed-phase high-performance liquid chromatography: a study of the chromatographic behavior of oxycodone hydrochloride. AB - Severe peak asymmetry--fronting--was observed for oxycodone during elution at 30 degrees C from a C18 HPLC column using a mobile phase consisting of 14.9% MeOH, 84.5% 0.05 M KH2PO4 (pH 3.0), 0.5% MTBE, and 0.1% TEA. Investigation using deuterium-labeled oxycodone and analysis by LC/MS showed that gem diol and hemiketal adducts of oxycodone formed as a result of the equilibrium addition of water and methanol to the C-6 ketone on oxycodone. As a result of slow equilibrium kinetics at room temperature in aqueous solution, the gem-diol and methyl hemiketal eluted as an unresolved broad band in front of the oxycodone peak. Decreasing the column temperature to 0 degrees C decreased the rates of interconversion and allowed the resolution and separation of these species from each other and from oxycodone. Increasing the column temperature to 60 degrees C increased the rates of interconversion with the result that the three species eluted as a single, homogenous peak with greatly improved peak symmetry. PMID- 10698533 TI - Systematic screening approach for chiral separations of basic compounds by capillary electrophoresis with modified cyclodextrins. AB - A simple, systematic method was developed for rapidly screening potential capillary electrophoresis (CE) separation conditions for small, amine-containing enantiomers. During method development, 39 pairs of enantiomers were investigated and partial or complete separation was achieved in every case. Baseline resolution was achieved by these initial screening conditions in over half of the cases. The screening strategy uses a bare fused silica capillary and a pH 2.5 amine-modified phosphate buffer containing one of the selected cyclodextrins (CD): dimethyl-beta-CD, hydroxypropyl-beta-CD, hydroxypropyl-alpha-CD, hydroxypropyl-gamma-CD and sulfated-beta-CD. An additional set of compounds have been screened by this approach to demonstrate the validity of the method. The paper outlines the experimental work carried out to develop the screen and describes how one might implement it for a new compound. PMID- 10698534 TI - Shorter development of immunoassay for drugs: application of the novel RIMMS technique enables rapid production of monoclonal antibodies to ranitidine. AB - High affinity, specific murine monoclonal antibodies have been produced for ranitidine using the novel RIMMS (repetitive immunizations, multiple sites) technique. We demonstrate that this technique can be employed to produce high affinity monoclonal antibodies to drug haptens in approximately 1 month; whereas, conventional techniques typically require 3-9 months. Polyclonal antiserum development typically requires at least 6 months. Consequently, RIMMS has a clear impact allowing reagent antibodies to be available earlier in the drug development process. Isotyping studies demonstrated that the developed antibodies are either IgG1 or IgG2b immunoglobulins which confirms that the technique produces class-switched, affinity matured reagent antibodies. The most promising monoclonal antibody for quantitative applications afforded similar sensitivity, by competitive ELISA, to the established sheep polyclonal anti-ranitidine sera. The calibration range, estimated as the limits between the asymptotic regions of calibration graphs, is 0.5-41.2 ng ranitidine per well. Specificity studies indicated that the monoclonal antibody afforded superior selectivity, yielding only 4.1% cross-reactivity with the ranitidine sulphoxide metabolite; the corresponding value for the antiserum was 8.6%. Both reagents had similar cross reactivities with the N-oxide metabolite. PMID- 10698535 TI - Solution conformation of model polypeptides with the use of particle beam LC/FT IR spectrometry and electrospray mass spectrometry. AB - Solution conformations of the polypeptides beta endorphin (beta-END) and a cysteine peptide (CYSP) were investigated with the use of particle beam LC/FT-IR spectrometry. Gradient elution HPLC with mobile phases that contained acetonitrile with 0.1% TFA (v/v) and 0.1% aqueous TFA (v/v) were used. The conformations of both polypeptides were studied in 0.9% sodium chloride injection USP, 5% dextrose in water injection USP and sterile water for injection USP. Additional conformational studies over a pH range of 2-10, temperatures of 25, 50, 75 and 100 degrees C and after storage for 24 h were investigated. The studies indicated that the two polypeptides did not behave similarly under identical conditions. It was observed that both beta-END and CYSP had slightly different conformations in the various parenteral solutions. It was also shown that the conformation of CYSP changed with both pH and temperature while beta-END was conformationally stable to both temperature and pH. The identity of the peptides and the conformationally sensitive charge-state intensities of the peptides were investigated with electrospray ionization mass spectrometry (ESI/MS). The combination of IR and MS data allowed an estimation of solution effects on the conformations of the model polypeptides. PMID- 10698536 TI - Determination of indinavir, potassium sorbate, methylparaben, and propylparaben in aqueous pediatric suspensions. AB - A gradient, reversed phase, HPLC method was developed for simultaneous analysis of potassium sorbate, methylparaben, propylparaben, and indinavir in aqueous suspensions that contain a proprietary orange flavoring and Magnasweet sweetener enhancer (MacSanrews and Forbes Company, Magnasweet product brochure). The chromatographic separation is performed on an Eclipse XDB-C8 column using a gradient run with an analysis time of 35 min. The mobile phase consists of acetonitrile and acetonitrile:citrate buffer, pH 4.0 (20:80 v/v). The method successfully separates the three preservatives, indinavir (active ingredient), the orange flavoring, the Magnasweet species, and the indinavir lactone degradate. Recovery, linearity, and precision results for the three preservatives and indinavir are described. The method applies to two types of formulations: Xanthan Gum suspension and NanoSystems suspension. PMID- 10698537 TI - HPLC assay of Lidocaine in plasma with solid phase extraction and UV detection. AB - A sensitive and reliable method based on solid-phase extraction and reversed phase liquid chromatography was developed and validated for the quantitation of Lidocaine (Lid) in dog plasma. Phenacemide was used as an internal standard (IS) in the extraction which employed C18 solid-phase extraction cartridges. The washing and eluting solutions were 2 ml acetonitrile-pH 9.0 phosphate buffer (10:90 v/v) and 0.5 ml acetonitrile-pH 4.0 phosphate buffer (40:60 v/v). respectively. The eluent obtained from the cartridge was directly analyzed on a reversed-phase ODS column with UV detection at 210 nm. A clean chromatogram and high sensitivity were achieved at this wavelength. The mobile phase was acetonitrile and pH 5.9 phosphate buffer (20:80 v/v). The retention times were 6.4 and 7.2 min for Lid and IS, respectively, at a flow rate of 1.0 ml min(-1). The mean absolute recovery was 96.6% (n = 9) with a CV of 3.8% for Lid and 81.7% with CV of 2.5% (n = 3) for IS. The limit of quantitation was 20 ng ml(-1), with the intra- and inter-day precisions (n = 5) of 4.4 and 3.4%, respectively, and the intra- and inter-day accuracies (n = 5) of -4.3 and -5.0%, respectively. For the analyses of Lid in spiked plasma samples at 20, 100 and 200 ng ml(-1), the overall mean intra- and inter-day precisions (n = 15) were 3.9 and 4.9%, respectively, and the overall mean intra- and inter-day accuracies (n = 15) were 3.7 and -4.6%, respectively. The correlation coefficients for calibration plots in the range 20-1000 ng ml(-1) in plasma were typically higher than 0.998. The suitability of the method was demonstrated by the study in a beagle dog receiving a low intravenous dose of Lid. PMID- 10698538 TI - Cleaning level acceptance criteria and a high pressure liquid chromatography procedure for the assay of Meclizine Hydrochloride residue in swabs collected from pharmaceutical manufacturing equipment surfaces. AB - A method using pharmacologically based and visual limit of detection criteria to determine the acceptable residue level for Meclizine Hydrochloride (MH) on pharmaceutical manufacturing equipment surfaces after cleaning is described. A formula was used in order to determine the pharmacologically safe cleaning level for MH. This level was termed as specific residual cleaning Level (SRCL) and calculated to be 50 microg 100 cm(-2). The visual limit of detection (VLOD) was determined by spiking different levels of MH on stainless steel plates and having the plates examined by a group of observers. The lowest level that could be visually detected by the majority of the observers, 62.5 microg 100 cm(-2), was considered as the VLOD for MH. The lower of the SRCL and VLOD values, i.e. 50 microg 100 cm(-2), was therefore chosen as the cleaning acceptance criterion. A sensitive reversed-phase HPLC method was developed and validated for the assay of MH in swabs used to test equipment surfaces. Using this method, the mean recoveries of MH from spiked swabs and '180-Grit' stainless steel plates were 87.0 and 89.5% with relative standard deviations (RSD) of +/- 3.3 and +/- 2.4%, respectively. The method was successfully applied to the assay of actual swab samples collected from the equipment surfaces. The stability of MH on stainless steel plates, on cleaning swabs and in the extraction solution was investigated. PMID- 10698539 TI - High performance liquid chromatographic analysis of the pharmacologically active quinones and related compounds in the oil of the black seed (Nigella sativa L.). AB - An HPLC method for quantifying the putative pharmacologically active constituents: thymoquinone (TQ), dithymoquinone (DTQ), thymohydroquinone (THQ), and thymol (THY), in the oil of Nigella sativa seed is described. Extraction of the constituents from the oil was carried out using C18 PrepSep mini columns followed by quantification of the recovered constituents by HPLC on a reversed phase muBondapak C18 analytical column, using an isocratic mobile phase of water:methanol:2-propanol (50:45:5% v/v) at a flow rate of 2 ml min(-1). UV detection was at 254 nm for TQ, DTQ, and THY, and at 294 nm for THQ. The above four compounds were separated with good resolution, reproducibility, and sensitivity under these conditions. This analytical method was used to quantify the above four constituents in a commercial sample of N. sativa seed oil, and provides a good quality control methodology for the pharmacologically active components in this widely used natural remedy. PMID- 10698540 TI - Comparison of free solution capillary electrophoresis and size exclusion chromatography for quantitating non-covalent aggregation of an acylated peptide. AB - There are few methods available for the rapid and precise quantitation of non covalent aggregation. The very methods used to measure the aggregation can easily disrupt the weak forces holding an aggregate together. This paper describes the novel application of free solution capillary electrophoresis (CE) for the quantitation of a biologically inactive non-covalent aggregate of C8GLIP (Des amino-histidine-7-arginine-26 N(epsilon)-octanoyl-lysine-34-human glucagon-like insulinotropic peptide), an acylated peptide. The CE results are compared to a more traditional approach using size exclusion chromatography (SEC). Under the conditions explored in this paper, SEC showed a significantly slower apparent rate of aggregation than CE. This is due to the disruption of the aggregate during the SEC process. The cause of the disruption is complex and is potentially related to the separation process itself, on-column dilution effects, and/or interactions of the aggregate with the column packing or SEC components. Analysis times and dilution are greatly reduced by CE, and, because there is no potentially interactive stationary phase and because both the protein and the walls of the capillary are negatively charged, potential disaggregation due to surface interactions is reduced. Thus, CE is shown to be superior to SEC for this peptide in that disruption of the aggregate is minimized. PMID- 10698541 TI - Determination of the endothelin receptor antagonist ABT-627 and related substances by high performance liquid chromatography. AB - The determination of the endothelin (ET) antagonist receptor ABT-627 (I) and related substances is performed by HPLC. I is determined in bulk drug substance and drug formulation using isocratic conditions and an Inertsil ODS-2 column. The determination is stability indicating and detector response is linear from 24 to 118 microg ml(-1) (33-164% of assay level). Intermediate precision for the determination ranged from +/- 0.60 to +/- 1.9% RSD. The measurement is accurate, with quantitative recovery of I from the formulation placebo. Related substances in I and formulated I are determined using the same chromatographic conditions, with a gradient elution profile to elute impurities having varying relative polarities. The detector response for related substances determination is linear for I from 0.60 to 17.8 microg ml(-1) (0.05-1.5% of assay level) with the limit of detection and quantitation estimated at 0.01 and 0.05%, respectively. Comparable precision was obtained in drug substance and drug formulation (RSD values +/- 3.7 to +/- 12% and +/- 5.5 to 16.9%, respectively for impurities ranging from 0.05 to 0.30%). The quantitated impurities agreed well for the same lot of I when assayed as a bulk substance and after the formulation into a drug product. PMID- 10698542 TI - Simultaneous analysis of dextran-methylprednisolone succinate, methylprednisolone succinate, and methylprednisolone by size-exclusion chromatography. AB - An analytical HPLC method is reported for simultaneous measurement of low (1.0 100 microg ml(-1)) concentrations of dextran-methylprednisolone succinate (DEX MPS) and its degradation products methylprednisolone hemisuccinate (MPS) and methylprednisolone (MP). The analytes were detected at 250 nm after resolution using a size exclusion column with a mobile phase of KH2PO4 (10 mM): acetonitrile (3:1) and a flow rate of 1 ml min(-1). The resolution of MP and MPS peaks was substantially affected by the pH of the mobile phase; while MP and MPS co-eluted at pH 3.4, they were baseline-resolved at pH > or = 5. Linear relationships (r > or = 0.997) were found between the detector response and the concentrations of the analytes (1.0-100 microg ml(-1) for MP and MPS and 2.5-100 microg ml(-1) for DEX-MPS). Intra- and inter-run error (< 13%) and precision (CV of < or = 6%) data indicated that the assay could accurately and precisely quantitate all three components in the examined concentration range. The application of the assay to determination of degree of substitution, purity, and stability of DEX-MPS was also demonstrated. PMID- 10698543 TI - Method development and validation for the HPLC assay (potency and related substances) for 20 mg paroxetine tablets. AB - A reversed phase high performance liquid chromatographic (HPLC) method was developed and validated for use as a stability indicating assay (potency and related substances) of paroxetine in paroxetine hydrochloride 20 mg tablets. Assay samples were extracted at a paroxetine concentration of 0.4 mg ml(-1) utilizing mobile phase as the extraction solvent. The chromatographic conditions employed a C18 column (Inertsil, 5 microm, 15 cm x 4.6 mm), isocratic elution with 10 mM 1-decane sulfonic acid sodium salt containing 10 mM sodium phosphate monobasic (pH 3.0)-ACN (60:40, v/v) and ultraviolet (UV) detection at 235 nm. PMID- 10698544 TI - Development and validation of a reversed-phase liquid chromatographic method for analysis of estradiol valerate and medroxyprogesterone acetate in a tablet formulation. AB - A simple and accurate liquid chromatographic method was developed for estimation of estradiol valerate and medroxyprogesterone acetate in pharmaceuticals. Drugs were chromatographed on a reverse phase C18 column, using a mixture (30:70) of ammonium nitrate buffer and acetonitrile and eluants monitored at a wavelength of 280 nm. Solution concentrations were measured on a weight basis to avoid the use of an internal standard. The method was statistically validated for its linearity, accuracy, precision and selectivity. Due to its simplicity and accuracy, the authors believe that the method may be used for routine quality control analysis. It does not require any specific sample preparation except the use of a column guard before the analytical column and suitable prefilter attached to the syringe prior to injection. PMID- 10698545 TI - High-performance liquid chromatographic assay of terbinafine hydrochloride in tablets and creams. PMID- 10698546 TI - Simultaneous determination of cinnarizine and domepiridone maleate from tablet dosage form by reverse phase ion pair high performance liquid chromatography. AB - A new simple, precise, rapid and selective reverse phase ion pair high performance liquid chromatography (HPLC-RP) method has been developed for the simultaneous determination of cinnarizine (CINN) and domepiridone maleate (DOME) from tablets using acetonitrile-methanol-water-0.1 N sulfuric acid (37:10:48:5 v/v/v/v) containing sodium lauryl sulfate (0.01 M), as a mobile phase and a Machery Nagel nitrile column (10 microns, 25 cm x 4.0 mm i.d.) as the stationary phase. The flow of mobile phase through the column was kept at 1.0 ml min(-1) through out the analysis. Detection was carried out using a UV detector at 225 nm. The retention times for CINN and DOME were 4.73 and 9.41 min, respectively. The linearity range and percentage recoveries for CINN and DOME were 4 1000 and 60-750 microg ml(-1) and 99.90 and 99.60%, respectively. PMID- 10698547 TI - Spectrophotometric and spectrofluorometric methods for the assay of lisinopril in single and multicomponent pharmaceutical dosage forms. AB - Simple and sensitive methods are described for the assay of lisinopril in tablets. The first method (A) is based on the reaction of the drug with chloranil in aqueous solution of pH 9.5 to give yellow colour measured at 346 nm. The second method (B) is based upon the interaction of lisinopril with dichlone resulting in the formation of an intense purple colour measured at 580 nm. The third method (C) depends on the reaction of the drug with acetylacetone and formaldehyde to form a coloured condensation product measured at 356 nm and also has a strong fluorescence at 475 nm (lambda(ex) 410 nm). This method is extended to determine lisinopril in binary mixtures with hydrochlorothiazide. The last method (D) depends on measuring the first and second derivative spectra of lisinopril. Moreover, the derivative method is used as stability-indicating method where lisinopril can be determined in presence of its degradation products. The proposed methods proved to be suitable for a rapid quality control of commercial dosage forms. The results obtained were precise and accurate. PMID- 10698548 TI - Quantitative analysis of R-84760, a selective kappa-opioid receptor agonist, in plasma by liquid chromatography with electrospray ionization tandem mass spectrometry. AB - A sensitive method for monitoring R-84760, a selective kappa-opioid receptor agonist, in plasma using liquid chromatography with electrospray ionization tandem mass spectrometry was explored. R-84760 and internal standard (I.S., d8-R 84760) were extracted from human or various animal plasmas with ethyl acetate. The analysis was performed by the selected reaction monitoring method, and the precursor-product combinations of m/z 399-328 for R-84760 and m/z 407-328 for I.S. were chosen for quantification. The calibration curve was linear in the range 5-500 pg/ml, and the limit of quantification was 5 pg/ml using 1 ml of human plasma. Pharmacokinetic studies of R-84760 in rats, dogs, and monkeys were performed by this method. The plasma concentration of unchanged form after administration at a trace dosage amount was able to be monitored. Interspecies correlations of pharmacokinetic parameters obtained in animals were utilized to estimate pharmacokinetic behavior in humans. The results showed that it is possible to perform pharmacokinetic studies on R-84760 by this quantitative analysis. PMID- 10698549 TI - Reduction of Cys36-Cys42 and Cys64-Cys74 disulfide bonds in recombinant human granulocyte colony stimulating factor. AB - The Cys36-Cys42 and Cys64-Cys74 disulfide bonds in recombinant methionyl human granulocyte colony-stimulating factor were reduced to sulfhydryls with dithiothreitol or mercury. Both reduction reactions are dependent on the pH. The reduction reaction with dithiothreitol increased in rate with increasing pH; between pH 7-9 and above pH 10.5 this increase was less than in other regions. These observations are explained by repulsive forces between dithiothreitol and regions in granulocyte colony-stimulating factor which intensify in these pH regions. The hydroxyl catalysis causes the overall increase in k(obs) in the pH region studied. The reduction of the disulfides with mercury is, as could be expected from the Nernst equation for disulfide reduction, also pH dependent: the half-wave potential decreases with increasing pH as predicted by theory. PMID- 10698550 TI - Enzyme inhibition XI: glutamate decarboxylase activity relationship with the reaction products as determined by the colorimetric and radioisotopic methods. AB - The relationship of glutamate acid decarboxylase (GAD) activity with the reaction products was developed. It was based on incubating sodium glutamate substrate (S) with GAD enzyme (E) when the enzyme-substrate-complex (ES) product was obtained along with gamma aminobutyric acid (GABA) and unreacted sodium glutamate. The reaction products were separated by paper chromatography. The ES, GABA and S products were sprayed with ninhydrin reagent when ninhydrin-colored-complex (NCC) was formed on the paper chromatogram. The products were extracted with 75% ethanol containing 0.5% cupric sulfate. The NCC absorption readings of ES and S products were measured by a spectrophotometer. A standard curve was prepared by plotting absorption readings against different concentrations of sodium glutamate. This curve was the basis of determining GAD activity of E. coli and C. welchii. It was observed that NCC absorption of ES and S products was directly related with the enzyme activity. The qualities of ES and S products in the reaction mixture increased as the enzyme activity increased with the incubation time. On the other hand, some products in the reaction mixture decreased in the presence of an inhibitor of GAD activity. The relationship of reaction products with GAD activity was also established by the radioisotopic method. The results obtained by the chromatographic separation of products followed by the spectrophotometric method of determining GAD activity is a simple, safe and less expensive compared to the other methods. PMID- 10698551 TI - Analysis of leucine enkephalin by high-performance liquid chromatography using enzymatic derivatization by tyrosinase and electrochemical or fluorescence detection. AB - Two tyrosine specific, HPLC methods with either electrochemical (HPLC-ED) or fluorescence (HPLC-FL) detection are described for leucine-enkephalin (LE) in cerebrospinal fluid (CSF). Both approaches involve the hydroxylation of the Tyr1 moiety of LE by mushroom tyrosinase. Production of a catechol permitted the selective clean-up of the analyte using boronate gels and produced species which were more readily oxidizable than the LE. The controlled oxidation of the catechol to corresponding chinones enabled the reaction with 1,2-diamino-1,2 diphenylethane (DPE) and subsequent quantification by HPLC-FL. The HPLC-ED and HPLC-FL yielded limits of detection for LE in CSF of 360 and 500 fmol per injection, respectively. Inter-day variability of calibration curve samples was lower than 20% for the HPLC-ED with slightly higher variability for the HPLC-FL assay. PMID- 10698552 TI - Foundations of chemical microscopy, 2 derivatives of primary phenylalkylamines with 5-nitrobarbituric acid. AB - 5-Nitrobarbituric acid (dilituric acid) was extensively used with great success as a chemical microscopic reagent for the qualitative identification of primary phenylalkylamines. This methodology was based on the characterization of observed crystal morphologies, since a unique crystal habit could be associated with each adduct product. To understand the scientific foundations which permitted chemical microscopy to function as a useful analytical technique, the products formed between dilituric acid and a series of primary phenylalkylamines were characterized using polarizing optical microscopy, powder X-ray diffraction, thermal analysis, and solid-state nuclear magnetic resonance. It was deduced that the origins of the different crystal morphologies associated with each of the crystalline adducts arose from the ability of the systems to form differing structural types and/or hydrates upon crystallization. The degree of hydration in the crystalline phenylalkylamine adducts appeared to increase as additional carbon atoms were added between the aromatic ring and the terminal amine group of the aliphatic sidechain. PMID- 10698553 TI - Analysis of benzalkonium chloride and its homologs: HPLC versus HPCE. AB - Benzalkonium chloride (BAK) is a mixture of alkylbenzyldimethylammonium chloride homologs with n-C,2H25, n-C,4H29, and n-C16H33 comprising a major portion of the alkyl groups present. An analytical method for BAK must differentiate and quantitate the homologs in the BAK mixture. Reversed-phase high performance liquid chromatography (HPLC) separates compounds based on their affinity for a nonpolar column, which is a direct correlation to the compounds' polarity. High performance capillary electrophoresis (HPCE), however, separates compounds in an electric field according to their charge and size. The BAK homologs are suitable for separation by either of these methods because their polarity and sizes differ significantly. The HPLC method employed a mobile phase of 60% acetonitrile and 40% 0.1 M sodium acetate buffer pH 5 pumped at 1.0 ml min(-1), a 4.6 x 250 mm cyano column with 5 microm packing, and UV detection at 254 nm. The HPCE method utilized a run buffer of 30% acetonitrile and 70% 0.05 M sodium phosphate pH 3.06, a 50 microm x 20 cm open silica capillary, 7.5 kV electric field and UV detection at 214 nm. Both HPLC and HPCE demonstrated good linearity in the range of 0.025 to 0.8 mg ml(-1) with r2 values of approximately 0.99. The HPLC method produced good separation of the homolog peaks with a total analysis time of 25 min. HPCE run time was less than 5 min and demonstrated good separation of the three homologs. The HPLC method, however, was superior to HPCE in the areas of sensitivity and precision. The HPLC has been extensively used in the routine quantitation and qualitation of benzalkonium chloride concentrations in various products; however, long analysis times make this method inefficient. The HPCE method produced comparable results to the HPLC method but with much shorter analysis times. An HPCE analysis method, as presented here, may prove to be a much more useful and efficient method for the analysis of benzalkonium chloride and its homologs. PMID- 10698554 TI - Kinase receptor activation (KIRA): a rapid and accurate alternative to end-point bioassays. AB - We have developed a novel strategy for a rapid bioassay that is accurate, precise, sensitive, and high capacity. It is capable of quantifying ligand bioactivity by measuring ligand-induced receptor tyrosine kinase activation in terms of receptor phosphorylation. The assay, termed a 'kinase receptor activation' or KIRA, utilizes two separate microtiter plates, one for ligand stimulation of intact cells, and the other for receptor capture and phosphotyrosine ELISA. The assay makes use of either endogenously expressed receptors or stably transfected receptors with a polypeptide flag. KIRA assays for the ligands IGF-I and NGF were compared to their corresponding endpoint bioassays (3T3 cell proliferation for IGF-I and PC12 cell survival for NGF). The KIRA assays showed excellent correlation with the more classical endpoint bioassays. Further, they were highly reproducible, minimizing the requirement for repeat assays. The KIRA assay format has great potential as a rapid, accurate and precise bioassay, both for potency determination as well as stability-indicating analyses. PMID- 10698555 TI - Automation of metabolic stability studies in microsomes, cytosol and plasma using a 215 Gilson liquid handler. AB - A 215 Gilson liquid handler was used to automate enzymatic incubations using microsomes, cytosol and plasma. The design of automated protocols are described. They were based on the use of 96 deep well plates and on HPLC-based methods for assaying the substrate. The assessment of those protocols was made with comparison between manual and automated incubations, reliability and reproducibility of automated incubations in microsomes and cytosol. Examples of the use of those programs in metabolic studies in drug research, i.e. metabolic screening in microsomes and plasma were shown. Even rapid processes (with disappearance half lives as low as 1 min) can be analysed. This work demonstrates how stability studies can be automated to save time, render experiments involving human biological media less hazardous and may be improve inter-laboratory reproducibility. PMID- 10698556 TI - Determination of a novel bile acid sequestrant in rodent diet by near-infrared spectroscopy. AB - DMP 504 is a high molecular weight polymer currently under development by The DuPont Merck Pharmaceutical Company as a novel bile acid sequestrant to lower serum cholesterol. To assess its safety, DMP 504 is incorporated into rodent diet for oral administration to rats and mice. An analytical method was developed to determine the accuracy and homogeneity of the blends. Since a physical separation or extraction of DMP 504 from the diet was not feasible, near-infrared spectroscopy (near-IR) was employed. The near-IR method provides accurate and precise results for blends containing 1.5-8.0% of DMP 504. Comparison of results at the 1.5% level with a cholate binding referee method is also presented. Both methods provided equivalent results for the 1.5% level. PMID- 10698557 TI - A phosphate binding assay for sevelamer hydrochloride by ion chromatography. AB - Sevelamer hydrochloride is a cross-linked polymeric amine; it is the active ingredient of Renagel capsules. Renagel is indicated for the control of hyperphosphatemia in patients with end-stage renal disease. An in vitro phosphate binding assay is required to measure the drug's efficacy. The assay developed for this purpose involves mixing the drug (polymer) with a solution of known phosphate concentration, filtering off the polymer-phosphate complex, and quantitating the unbound phosphate concentration by ion chromatography. The binding capacity, reported as mmol of phosphate bound g of polymer(-1), is calculated from the calculated amount of bound phosphate and the weight of polymer used. The method has been validated for accuracy, precision, linearity, range, and ruggedness. PMID- 10698558 TI - Analysis of a residual diamine in a pharmaceutical polymer using solid phase extraction with analysis by gas chromatography mass spectrometry. AB - A method was developed for the analysis of 4,4'-methylenebiscyclohexylamine (DMDA) and 4,4'-methylenedicyclohexylisocyanate (DMDI) in a pharmaceutical polymer. The DMDA was extracted from the polymer with either buffer (0.1 M potassium phosphate pH 3.1) and the extract was passed through a SCX solid phase extraction cartridge. It was eluted from the cartridge with methanolic ammonia and then converted to its heptafluorobutyramide (HFB) derivative prior to analysis by gas chromatography-negative chemical ionisation mass spectrometry (GC MS) in the negative ion chemical ionisation (NICI) mode. It was not possible to directly measure DMDI and it was thus analysed by selecting extraction conditions such that it would decompose to DMDA. The quantification of the residues in the polymer was based on the method of standard additions since this gave a better indication of the recovery from the complex matrix. PMID- 10698559 TI - Identification of pharmaceutical excipients using NIR spectroscopy and SIMCA. AB - Soft independent modelling of class analogy (SIMCA) is applied to identify near infrared (NIR) spectra of ten excipients used in the pharmaceutical industry. For each class at least 15 excipient samples were collected for the data base, considering different batches and occasionally various suppliers. Therefore the data of the classes are not always homogeneous. The performance of the original SIMCA method, which is usually described in the literature and also applied by the users, carried out at two confidence levels, 95 and 99%, on original data, SNV (standard normal variate transformation) and second derivative pre-processed data, is discussed. Reasons for the rejection rates are given. No objects were assigned to a wrong class using SIMCA. PMID- 10698560 TI - Determination of tungsten in bulk drug substance and intermediates by ICP-AES and ICP-MS. AB - A quick and sensitive method has been developed and validated for the determination of tungsten in bulk drug substance and intermediates using either Inductively Coupled Plasma Atomic Emission Spectrometry (ICP-AES) or Inductively Coupled Plasma Mass Spectrometry (ICP-MS). Sample preparation is by direct dissolution with a 80:20 (v/v) concentrated nitric acid:deionized water mixture and avoids labor intensive and potentially hazardous digestion techniques. Excellent agreement was found between ICP-AES and ICP-MS results and between Merck results and Microwave Induced Plasma Mass Spectrometry (MIP-MS) results provided by an independent raw material vendor. PMID- 10698561 TI - Comparison between micellar electrokinetic chromatography and HPLC for the determination of Betamethasone Dipropionate, Clotrimazole and their related substances. AB - The complete separation of a composite mixture that consisted of Betamethasone Dipropionate (BMD), Clotrimazole and their derivatives in a pharmaceutical dosage form was achieved within 15 min using sodium dodecyl sulfate (SDS) micellar electrokinetic chromatography (MEKC). For the MEKC separations, electrophoretic media consisting of SDS-phosphate buffer and various concentrations of alcohols or acetonitrile were used. The optimal condition for separating BMD, Clotrimazole and their analogues was found to be 50 mM SDS-15% acetonitrile-5% butanol at pH 7.2. The results demonstrated that the method was valid for the quantitation of BMD, Clotrimazole and analogues with selectivity and precision comparable to that of High-Performance Liquid Chromatography (HPLC). PMID- 10698562 TI - Analysis of various nucleosides in plasma using solid phase extraction and high performance liquid chromatography with UV detection. AB - The National Cancer Institute (NCI) has screened many nucleosides for antiviral activity to the HIV-1 virus. Drugs demonstrating antiviral activity are tested in animal models to evaluate their toxicity and pharmacokinetic characteristics. These drugs are subsequently evaluated for efficacy in human clinical trials. Sensitive analytical methodology is needed to quantify nucleosides in plasma and other biological matrices in support of these studies. Battelle has modified and validated a reversed phase high-performance liquid chromatography (HPLC) method for several of these nucleosides that could be easily adapted for similar compounds. Methods have been validated for 6-chloro-2',3'-dideoxyguanosine (6ClddG), 6-chloro-2',3'-dideoxyinosine (6ClddI) and their primary metabolites 2',3'-dideoxyguanosine (ddG) and 2',3'-dideoxyinosine (ddI) in both rat and dog plasma containing EDTA. The method has also been validated for 2'-fluoro-2',3' dideoxyara-adenosine (betaFlddA) and its primary metabolite 2'-beta fluorodideoxyinosine (betaFddI) in rat plasma containing heparin. Calibration plasma standards were prepared over ranges of 0.1-10 microg ml(-1) for betaFlddA and betaFddI, 0.1-50 microg ml(-1) for 6ClddG and ddG, and 0.25-50 microg ml(-1) for 6ClddI and ddI in plasma containing 4 microg ml(-1) pentostatin. The addition of pentostatin to the plasma samples inhibits in-vitro deamination of the drug after collection. Quality control (QC) standards were prepared containing the appropriate anticoagulant and 4 microg ml(-1) pentostatin at concentrations within each of the bracketed calibration ranges in plasma. These methods have been successfully applied to plasma samples generated during various animal studies. PMID- 10698563 TI - Liquid chromatographic determination of hippuric acid for the evaluation of ethacrynic acid as angiotensin converting enzyme inhibitor. AB - A rapid, simple and interference-free method is described to evaluate the inhibitory effects of organic compounds on the activity of angiotensin converting enzyme irrespective of their acid-base properties. The assay is based on the high performance liquid chromatographic separation of the synthetic substrate hippuryl L-histidyl-L-leucine, the hydrolysis product hippuric acid and the test compound. Using the new method, the diuretic drug ethacrynic acid was found to act as an inhibitor for the enzyme in a non competitive mode. PMID- 10698564 TI - Bioanalytical applications of polyion-sensitive electrodes. AB - Recent developments in the design and bioanalytical applications of polyion sensitive electrodes (PSEs) are reviewed. The general electrochemical principles governing the potentiometric response of such polymer membrane-based devices are summarized and new directions for the use of these novel sensors are detailed. These new directions include basic fundamental studies aimed at determining the thermodynamics of polyion extraction into ion exchanger-doped polymeric membranes, new methods to quantitate the anticoagulant drug heparin in whole blood via titrations with polycationic protamine, selective assays of protease activities (and inhibitors of such activities) using natural and synthetic polyionic peptides as substrates, and novel homogeneous immunoassay schemes based on potentiometric polyion detection. PMID- 10698565 TI - Chemical profiles and monitoring dynamics at an individual nerve cell in Planorbis corneus with electrochemical detection. AB - The identified dopamine cell of Planorbis corneus is described as a model system to study neurotransmitter storage and dynamics. Techniques developed with this model system include capillary electrophoresis with electrochemical detection and microelectrochemistry at single cells. These techniques provide a powerful combination to examine single cell neurochemistry. Whole cell and cytoplasmic dopamine concentrations have been quantified with capillary electrophoresis. Additionally, this technique has been used to profile amino acids and to quantify two compartments of neurotransmitter in a single cell. Individual exocytosis events have been monitored at the cell body of the dopamine cell of P. corneus with microelectrodes. In this case, two different types of vesicles have been identified based on the amount of transmitter released. The psychostimulant, amphetamine, has been shown to selectively affect the amount of dopamine in these vesicles with lower to higher doses affecting the larger to smaller vesicle types, respectively. Microelectrochemistry at single nerve cells has also been used to demonstrate reverse transport of dopamine across the cell membrane and to suggest a role of this process in the molecular mechanism of amphetamine. PMID- 10698566 TI - Electrochemical monitoring of biogenic amine neurotransmission in real time. AB - Three techniques, constant-potential amperometry, high-speed chronoamperometry, and fast-scan cyclic voltammetry, have been used extensively to investigate the rapid events associated with neurotransmission. These techniques vary in sensitivity, chemical resolution and temporal resolution. Amperometry provides the best temporal resolution but little chemical resolution. Fast-scan cyclic voltammetry provides both good temporal and chemical resolution, while high-speed chronoamperometry offers good temporal resolution and moderate chemical resolution. The amount of chemical information which is needed for a neurochemical measurement depends upon the sample. For single cells, secondary methods, such as HPLC and capillary electrophoresis, offer extensive chemical information about the contents of a cell. With this information, chemical information is not needed during the electrochemical measurement. Therefore, amperometry is employed to obtain the greatest temporal resolution. However, when using more complex biological samples, such as brain slices or in vivo implantation, there is a greater demand for chemical resolution provided by the electrochemical measurement. To bolster results, further confirmation is sought from anatomical, physiological and pharmaceutical evidence. Within this review, the three electrochemical techniques are outlined and compared. Examples are then provided of measurements which have been made in the three predominant biological samples which have been studied: single cells, brain slices and intact animals. PMID- 10698567 TI - Amperometric biosensors for clinical and therapeutic drug monitoring: a review. AB - The coupling of enzymes and electrode transducers permits the rapid and simple determination of endogeneous compounds and therapeutic drugs in clinical samples. New developments in the operation, miniaturization and microfabrication of electrochemical biosensors offer exciting possibilities for biomedical and pharmaceutical analysis. This review focuses on the current state of amperometric enzyme electrodes for biomedicine, with emphasis on recent advances, challenges and trends. PMID- 10698568 TI - Chemical and biochemical sensors based on advances in materials chemistry. AB - The advance of materials chemistry has influenced the design of analytical sensors, especially those using spectroscopic or electrochemical methods for generating the signal. New methods of immobilizing enzymes, chromophores, and electron-transfer catalysts have resulted from initiatives in materials science. Systems based on sol-gel chemistry are especially noteworthy in this regard, but other important materials for chemical and biochemical sensors include zeolites, organic polymers, and various conducting composites. Applications cited include determinations of inorganic ions, gases, neurotransmitters, alcohols, carbohydrates, amino acids, proteins, and DNA. PMID- 10698569 TI - Electrochemical determination of carbohydrates: enzyme electrodes and amperometric detection in liquid chromatography and capillary electrophoresis. AB - In recent years, electrochemical detection (EC) methods have become increasingly important for the determination of carbohydrate compounds in a variety of biological and pharmaceutical samples. In this work, recent advances in the design and application of EC approaches are reviewed, with the goal of providing the non-electrochemist with a basic understanding of the most important EC approaches to carbohydrate detection and an overview of their current applications. Two specific EC detection strategies are considered in detail: enzyme electrodes and electrodes used for HPLC or capillary electrophoresis detection. PMID- 10698570 TI - Development of sub-micron patterned carbon electrodes for immunoassays. AB - Sub-micron sized domains of a carbon surface are derivatized with antibodies using biotin/avidin technology. These sites are spatially-segregated from, and directly adjacent to, electron transfer sites on the same electrode surface. The distance between these electron transfer sites and enzyme-loaded domains are kept to a minimum (e.g. less than a micron) to maintain the high sensitivity required for the measurement of enzyme-linked cofactors in an enzyme-linked immunoassay (ELISA). This is accomplished through the use of photolithographic attachment of photobiotin using an interference pattern from a UV laser generated at the electrode surface. This allows the construction of microscopic arrays of active ELISA sites on a carbon substrate while leaving other sites underivatized to facilitate electron transfer reactions of redox mediators; thus maximizing sensitivity and detection of the enzyme mediator. The carbon electrode surface is characterized with respect to its chemical structure and electron transfer properties following each step of the antibody immobilization process. The characterization of specific modifications of micron regions of the carbon surface requires analytical methodology that has both high spatial resolution and sensitivity. We have used fluorescence microscopy with a cooled CCD imaging system to visualize the spatial distribution of enzyme immobilization sites (indicated by fluorescence from Texas-Red labeled antibody) across the carbon surface. The viability of the enzyme attached to the surface in this manner was demonstrated by imaging the distribution of an insoluble, fluorescent product. PMID- 10698571 TI - Redox hydrogel based bienzyme electrode for L-glutamate monitoring. AB - Amperometric bienzyme electrodes based on coupled L-glutamate oxidase (GlOx) and horseradish peroxidase (HRP) were constructed for the direct monitoring of L glutamate in a flow injection (FI)-system. The bienzyme electrodes were constructed by coating solid graphite rods with a premixed solution containing GlOx and HRP crosslinked with a redox polymer formed of poly(1-vinylimidazole) complexed with (osmium (4-4'-dimethylbpy)2 Cl)II/III. Poly(ethylene glycol) diglycidyl ether (PEGDGE) was used as the crosslinker and the modified electrodes were inserted as the working electrode in a conventional three electrode flow through amperometric cell operated at -0.05 V versus Ag?AgCl (0.1 M KCl). The bienzyme electrode was optimized with regard to wire composition, Os-loading of the wires, enzyme ratios, coating procedure, flow rate, effect of poly(ethyleneimine) addition, etc. The optimized electrodes were characterized by a sensitivity of 88.36 +/- 0.14 microA mM(-1) cm(-2), a detection limit of 0.3 microM (calculated as three times the signal-to-noise ratio), a response time of less than 10 s and responded linearly between 0.3 and 250 microM (linear regression coefficient = 0.999) with an operational stability of only 3% sensitivity loss during 8 h of continuous FI operation at a sample throughput of 30 injections h(-1). PMID- 10698572 TI - A new enzyme electrode for quantification of salicylic acid in a FIA system. AB - This work presents an amperometric biosensor incorporated into a flow configuration comprising salicylate hydroxylase that catalyses the irreversible hydroxylation of salicylate to catechol in the presence of NADH and molecular oxygen, and tyrosinase that further oxidises catechol giving o-quinone which is electrochemically reduced at -100 mV vs. Ag/AgCl yielding catechol and entering the catalytic oxidation and electrochemical reduction cycling which results in signal amplification and, consequently, low limits of detection. Additional incorporation of glucose dehydrogenase in the enzymatic sequence results in regeneration of NADH provided that glucose is present in the carrier stream and incorporation of a dialysis membrane provides operational stability to the biosensor. The analytical characteristics of this catalytic and electrochemical transduction sequence in a FIA system are: a limit of detection of 3.5 10(-6) M (S/N = 3), a sensitivity of 22.6 nA microM(-1) cm(-2), no loss of response at least after 5 h of continuous operation, and a sample frequency of 15 h(-1). Monitoring of salicylate after ingestion of 500 mg of acetylsalicylic acid has been followed in non-pretreated urine samples and the amount of salicylate in several drugs has been also successfully quantified. PMID- 10698573 TI - Using entropies of reaction to predict changes in protein stability: tyrosine-67 phenylalanine variants of rat cytochrome c and yeast Iso-1 cytochromes c. AB - Using the voltammetric method of square-wave voltammetry, a direct electrochemical examination was made of the wild type and Tyr67Phe mutant of both rat cytochrome c and yeast iso-1-cytochrome c. In addition to determining the equilibrium reduction potential (E0') for each cytochrome, the entropy of reaction, deltaS0'(Rxn)(deltaS0'(Rxn) = S0'(Red) - S0'(Ox)), for the reduction process was determined via the non-isothermal method. Having determined deltaS0'(Rxn) and E0', deltaH0' was calculated. For rat cytochrome c, it was found that deltaS0'(Rxn) = -43 J mol(-1) K(-1) for the wild type and -53 J mol( 1) K(-1) for the Tyr67Phe variant, with the deltaH0' for both the wild type and variant nearly identical, indicating that the changes in reduction potential and probably stability are due to changes in deltaS0'(Rxn). In contrast the measured deltaS0'(Rxn) for yeast iso-1-cytochrome c demonstrated significant changes in both entropic and enthalpic contributions in going from wild type to mutant cytochrome c. The entropy of reaction provides information regarding the relative degree of solvation, and very likely the degree of compactness, of the oxidized state versus the reduced state of the redox protein. A thermodynamic scheme and stability derivation are presented that show how the entropies of reaction of wild type versus variant cytochromes contribute to and predict changes in stability in going from oxidized to reduced protein. For yeast iso-1-cytochrome c, the thermodynamically predicted change in stability was very close to the experimentally observed value, based on previous differential scanning calorimetric stability measurements. While such data is not available for rat cytochrome c, consideration of the enormously increased local stability of the rat oxidized cytochrome c variant predicts that the reduced rat variant will be even more stable than the already stabilized oxidized variant. PMID- 10698574 TI - Electrochemical behaviour of cytochrome c at electrically heated microelectrodes. AB - The structural changes in cytochrome c with temperature have been been followed using a recently developed electrically-heated microelectrode sensor. Differential pulse voltammetry was used to perform electrochemical measurements of cytochrome c oxidation at different temperatures at heated bare gold electrodes contained in phosphate-buffered cytochrome c solution at room temperature. The voltammetric response shows the onset of unfolding and a marked dependence of the signal on electrode temperature. This augurs well for applications of heated electrodes as local probes in the study of the temperature dependence of electron transfer processes of other redox proteins, avoiding problems of bulk deterioration. PMID- 10698575 TI - Direct electrochemical measurement of nitric oxide in vascular endothelium. AB - The endothelium plays a critical role in maintaining vascular tone by releasing vasoconstrictor and vasodilator substances. Endothelium-derived nitric oxide (NO) is a vasodilator rapidly inactivated by superoxide and by Fe(II) and Fe(III), all found in significant quantities in biological systems. Thus due to the short life of NO in tissue (t1/2 = 3-6 s), in situ quantification of NO is a challenging problem. We designed the present study to perform direct measurements of nitric oxide using the electrochemical porphyrinic sensor. The most significant advantages of this sensor is small size (0.5-8 microm), rapid response time (0.1 1 ms), and low detection limit (10(-9) mol l(-1)). The porphyrinic sensor was used for in vitro and in vivo measurements of NO in an isolated single cell or tissue. Effects of hypertension, endotoxemia, and ischemia/reperfusion on the release of NO and/or its interaction with superoxide (O2-) were delineated. In the single endothelial cell (rabbit endocardium), NO concentration was highest at the cell membrane (950 +/- 50 nmol l(-1)), decreasing exponentially with distance from cell, and becoming undetectable at distances beyond 50 microm. The endothelium of spontaneously hypertensive rats (SHR) released 35% less NO (580 +/ 30 nmol l(-1)) than that of normotensive rats (920 +/- 50 nmol l(-1)), due to the higher production of O2- in SHR rats. Endothelial NO synthase (eNOS) generated NO (140 +/- 20 nmol l(-1)) in lung during the acute phase (first 10-15 min) of endotoxemia, followed by production of NO by inducible NOS. High production of O2- was observed during the entire period of endotoxemia. Ischemia (lower limb of rabbit) caused a significant increase of NO peaking at 15 min and decreasing thereafter, also due to O2- production. PMID- 10698576 TI - Capillary electrochemical enzyme immunoassay (CEEI) for phenobarbital in serum. AB - A competitive heterogeneous capillary enzyme immunoassay with electrochemical detection has been developed for phenobarbital in serum. The oxidized primary antibody was attached covalently to the modified interior surface of a microcapillary (22 microl). The competition between analyte phenobarbital and alkaline phosphatase labeled phenobarbital for a limited number of antibody binding sites was complete in 1.5 h. The enzymatic product (p-aminophenol) from the catalytic conversion of the substrate (p-aminophenyl phosphate) was detected by amperometric flow injection analysis. The calibration curve for phenobarbital had a detection limit of 30 microg l(-1) (2.8 pmoles or 0.65 ng) and a range of 30-3000 microg l(-1). The assay could be used to determine the phenobarbital serum concentration in a 4 microl clinical serum sample without pretreatment. PMID- 10698577 TI - Feasibility studies of simultaneous multianalyte amperometric immunoassay based on spatial resolution. AB - A multianalyte immunoassay concept based on the geometric separation of different analyte-specific antibodies has been demonstrated. The assay and amperometric detection are done in a cell with two working electrodes controlled at the same potential, and the amperometric signal at each electrode is monitored. The distance between any two adjacent electrodes in this prototype is 2.5 mm, and during the course of amperometric measurement, the product formed at one electrode does not reach the other working electrode within 20 min after the addition of enzyme substrate. Thus, the method relies on the spatial resolution between the different antibodies being such that measurements are taken before cross-interference due to diffusion can occur. Identical enzyme labels (alkaline phosphatase, ALP) and substrates (p-aminophenyl phosphate, PAPP) are used for all analytes. Alkaline phosphatase-conjugated rat anti-mouse IgG was immobilized by passive adsorption. Our studies showed that this concept is feasible and can be applied to the simultaneous measurement of multiple analytes. PMID- 10698578 TI - Improving measurement stability and reproducibility of potentiometric sensors for polyions such as heparin. AB - The growing importance of polymer membrane-based potentiometric polyion sensors in biomedical research and clinical measurements has brought up the question of how accurate and reproducible these sensors are. Indeed, recent research has revealed that these sensors behave quite differently than classical ion-selective electrodes. This paper explores ways to improve measurement reproducibility and long term potential stability by considering the unique pseudo steady-state response mechanism of the polyion sensors developed so far. Heparin may be stripped out of the phase boundary membrane surface with a high sample NaCl concentration and this characteristic is used to modify the calibration procedure in order to avoid memory effects. It is also attempted to reduce long term potential drifts by continuously stripping heparin out of the membrane at the membrane-inner filling solution side. PMID- 10698579 TI - Preconcentration and voltammetry of mercury on a functionalized sol-gel thin film modified glassy carbon electrode. AB - A functionalized sol-gel thin film modified glassy carbon electrode has been prepared for the determination of mercury. The tetrasulfide in the sol-gel matrix has a high affinity for mercury species. The chemically integrated functional groups in the sol-gel precursor ensure a high chemical and mechanical stability of the modified electrodes, and therefore a stable and reproducible analytical performance. By medium exchange anodic stripping voltammetry, this modified electrode allows reliable, low cost and interference-free determination of mercury. PMID- 10698580 TI - Transport studies of beta-lactam antibiotics and their degradation products across electrified water/oil interface. AB - An electrochemical method for quantifying beta-lactam antibiotics (cephalexin and ampicillin) and their hydrolysis products is described. Cyclic voltammetry at the water/nitrobenzene interface in a four-electrode system was used. The zwitterionic compounds were ionized to the necessary electrochemical form by pH adjustment. The pH change, however, resulted also in hydrolysis of the antibiotics. Hydrolysis products were characterized across UV-vis spectrum. The various hydrolysis products as well as the ionized antibiotics were studied in voltammetric transfer from water to nitrobenzene using the method of the interface between two immiscible electrolyte solutions (ITIES). It was concluded that this electrochemical method is suitable for the quantification of beta lactam antibiotics and their hydrolysis products. PMID- 10698581 TI - Electrochemical behavior of 3-chloro-2,4-pentanedione in the presence of cobalt salen. AB - We have studied the catalytic two-electron reduction of 3-chloro-2,4-pentanedione by cobalt(I) salen electrogenerated at a glassy carbon cathode in acetonitrile containing tetramethylammonium tetrafluoroborate. When cobalt(I) salen is electrogenerated at -0.65 V (a potential that is 30 mV more negative than the peak potential for the reversible one-electron reduction of cobalt(II) salen), the carbon-chlorine bond of 3-chloro-2,4-pentanedione is catalytically cleaved to form 2,4-pentanedion-3-ate; this anion can be protonated either by adventitious water or by a deliberately added proton donor to produce 2,4-pentanedione, or the anion can be trapped with iodoethane to give 3-ethyl-2,4-pentanedione. However, when cobalt(I) salen is electrogenerated at -0.40 V (a potential at which the rate of generation of cobalt(I) salen is relatively small), the 2,4-pentanedion-3 ate, resulting from the catalytic two-electron cleavage described above, can deprotonate unreduced starting material to form 3-chloro-2,4-pentanedion-3-ate and 2,4-pentanedione. In further work, we have found that 2,4-pentanedion-3-ate can be oxidized directly to form the corresponding radical which couples to yield 3,4-diacetyl-2,5-hexanedione. Chemically produced 2,4-pentanedion-3-ate reacts with electrogenerated cobalt(III) salen to give a dionylcobalt(III) salen species which undergoes a one-electron reduction to liberate cobalt(II) salen and the dionate. In addition, cobalt(II) salen reacts with molecular oxygen to give cobalt(III) salen and superoxide, and the latter reduces 3-chloro-2,4 pentanedione to form chloride ion, the 2,4-pentanedion-3-yl radical, and molecular oxygen. PMID- 10698583 TI - Amperometric determination of xanthine and hypoxanthine at carbon electrodes. Effect of surface activity and the instrumental parameters on the sensitivity and the limit of detection. AB - The performance of active graphite and carbon fiber surfaces produced by different mechanical/electrochemical methods of surface activation has been investigated in the amperometric determinations of xanthine and hypoxanthine under physiologically relevant conditions. The electrodes showed better limits of detection (LOD) when used with differential techniques with a capability of discriminating the analytical signal from the background. Square wave voltammetry and cyclic voltammetry showed the most sensitive response. Electrochemically activated carbon fiber ultramicroelectrodes showed the highest sensitivity (58 A M(-1) cm(-2)) and the LOD in the 200 nM range was observed at the rough pyrolytic graphite electrodes by square wave voltammetry. The results demonstrate the feasibility of the development of new electroanalytical methods for the determination of oxypurines in biological samples. PMID- 10698582 TI - A method for evaluating chemical selectivity of agonists for glutamate receptor channels incorporated in liposomes based on an agonist-induced ion flux measured by ion-selective electrodes. AB - A new method for evaluating chemical selectivity of agonists for the NMDA subtype of glutamate receptor (GluR) channels is described. The method is based on the magnitude of Ca2+ release from GluR-incorporated liposomes, which is measured by a Ca2+ ion-selective electrode with a thin-layer mode. The partially purified GluRs from rat whole brain were reconstituted into Ca2+-loaded liposomes. Small aliquots (each 50 microl) of the proteoliposomes, in the presence of an antagonist DNQX for blocking non-NMDA subtype, were subjected to potentiometric measurements of Ca2+ release under stimulation by three kinds of agonists, i.e. NMDA, L-glutamate and L-CCG-IV. The amount of the Ca2+ ion flux through the GluR channel induced by the agonists was found to increase in the order of NMDA < L glutamate < L-CCG-IV, which was consistent with that of binding affinity of the agonists toward the NMDA subtype. However, the range of selectivity of the relevant agonists was much smaller compared with results based on binding affinities. The present method provides physiologically more relevant values for the agonist selectivity of GluRs as compared to that of the conventional binding assay in the sense that the selectivity is based on the very magnitude of Ca2+ flux through the NMDA receptor, i.e. the extent of signal transduction by a given agonist. The evaluation of agonist selectivity based on Na+ release was also investigated by using a Na+ ion-selective electrode, but agonist-induced Na+ release was not detected, because of low permeability of Na+ through the NMDA subtype. PMID- 10698584 TI - Detection of hydrazine, methylhydrazine and isoniazid by capillary electrophoresis with a 4-pyridyl hydroquinone self-assembled microdisk platinum electrode. AB - Capillary electrophoresis (CE)/electrochemical detection (EC) for the simultaneous detection of hydrazine, methylhydrazine, and isoniazid has been developed with a 4-pyridyl hydroquinone self-assembled microdisk platinum electrode. Such an electrode has very high catalytic ability for hydrazines and they could be detected even at 0.0 V. The responses for hydrazine, methylhydrazine, and isoniazid are linear over 3 orders of detected concentration and of magnitude of 0.2-400 microM, 0.2-400 microM, 0.5 microM-2 mM, with correlation coefficients of 0.9998, 0.9991, and 0.9982, respectively. And they could be detected to levels of 0.1, 0.1 and 0.2 microM, respectively. This modified electrode was found to be very stable and reproducible when continuously used as detector for capillary electrophoresis for period of at least 4 weeks with no apparent loss of response. PMID- 10698585 TI - Pulsed electrochemical detection of sulfur-containing antibiotics following high performance liquid chromatography. AB - Pulsed electrochemical detection (PED) following reversed-phase chromatography has been applied to the direct detection of sulfur-containing antibiotics, specifically, penicillins, cephalosporins, and lincomycin. The compounds are detected sensitively and selectively without the need for derivatization. Integrated pulsed amperometric detection (IPAD) yields limits of detection lower than UV detection for these compounds. Detection limits using an optimized IPAD waveform are typically 10 ppb or less. The high selectivity of PED for thiocompounds reduces sample preparation. This work is applied to the determination of penicillin and related analogues in various pharmaceutical formulations/preparations, including a chicken feed. PMID- 10698586 TI - Electrochemical detection of catecholamines at sub-5 fg levels by redox cycling. AB - Two simple modifications to a commercially available thin layer electrochemical detector cell permitted the attainment of ultra-low detection levels of two neurotransmitter catecholamines. An ESA model 5041 analytical cell was modified with a glassy carbon embedded ceramic composite electrode to allow the use of a thin 12 microm gasket. Also a capillary HPLC column was connected directly to the detector cell using a 30 microm i.d. fused silica capillary. These modifications permitted the extensive redox cycling of the electrochemically reversible catecholamines. The ensuing amplified analytical signal allowed the detector cell to achieve efficiencies of 1300%. This resulted in a mass limit of detection of 4 fg and a concentration limit of detection of 116 pM for dopamine with an S/N of 3. PMID- 10698587 TI - Development of a liquid chromatographic method for picomole determination of S sulfocysteine in trifluoroacetic acid extracts of neonatal rat brain. AB - Neonatal Sprague Dawley rat brain tissue was extracted with methanol, acetonitrile, acetic acid and trifluoroacetic acids (TFA). Among the extractants tested, 0.1 M TFA gave the highest recovery, 73.4 +/- 5.2% (slope of regression of 'added' vs. 'found' and standard error of the slope) of S-sulfocysteine (SSC). The poorest recovery of SSC was found with acetonitrile and 90% methanol extractions (less than 10%). Possible reasons for the low recoveries have been explored. The recovery of SSC from aqueous standards in 0.1 M TFA is 92 +/- 5%. Detection of picomole quantities of SSC has been demonstrated with a combination of the optimized extraction procedures and our previously developed detection system. Supernatant of rat brain homogenate (0.10 M TFA as extractant) was evaporated to dryness in a vacuum centrifuge. Residues were reconstituted with deionized water. Samples were separated on a reversed phase column. The mobile phase was 20 mM aqueous acetate buffer (pH 5.2) containing 0.40 mM cetyl trimethylammonium p-toluene sulfonate and 2 vol.% methanol. Electrochemical detection used dual series gold-mercury amalgam electrodes. For the first time, S sulfocysteine was detected in normal neonatal rat brain. Its concentration is 0.99 +/- 0.25 pmol/mg brain tissue. The results indicate that TFA, rarely reported an an extractant, efficiently recovers SSC from rat brain tissues. PMID- 10698588 TI - Natural history of the classical form of primary growth hormone (GH) resistance (Laron syndrome). AB - A description of the clinical, biochemical and endocrinological features of the classical form of the syndrome of primary growth hormone (GH) resistance (Laron syndrome) is presented including the progressive changes during follow-up from infancy into adulthood. The main diagnostic features are: severe growth retardation, acromicria, small gonads and genitalia, and obesity. Serum GH levels are elevated and insulin-like growth factor-I (IGF-I) values are low and do not rise upon stimulation by exogenous hGH. The pathogenesis of this syndrome is due to various molecular defects from exon deletion to nonsense, frameshift, splice and missense mutations in the GH receptor (GH-R) gene or in its post-receptor pathways. PMID- 10698589 TI - Partial growth hormone insensitivity. AB - Partial growth hormone (GH) insensitivity can be defined as the clinical and biochemical features of IGF-I deficiency without GH deficiency and in the absence of the dysmorphic features of Laron syndrome. There is good evidence that this form of GH insensitivity exists, both in the context of severe GH resistance, and also in some patients with idiopathic short stature. The series of GH insensitivity patients in the European study shows a spectrum of clinical and biological defects, with several patients at the milder end of the spectrum having normal facies. The report of the presence of heterozygous mutations of the GH receptor in patients with idiopathic short stature has been confirmed by documentation of dominantly inherited mutations in familial short stature. Molecular screening in our unit of a group of 31 children with idiopathic short stature and normal GHBP, failed to identify mutations of the intracellular domain of the GH receptor. Consequently, although partial GH insensitivity is a proven entity, the clinical and biochemical identification of patients with GH resistance should precede molecular analysis. The analysis of individual patients and their families is more likely to reveal mutations, rather than a strategy of blanket molecular screening. PMID- 10698590 TI - Experience with insulin-like growth factor I (IGF-I) treatment of growth hormone insensitivity syndrome (GHIS). PMID- 10698591 TI - Loss of function mutations of the GnRH receptor: a new cause of hypogonadotropic hypogonadism. AB - The association of hypogonadotropic hypogonadism with anosmia defines Kallmann's syndrome. The gene of the X-linked form of this syndrome has been cloned and several mutations described. However, the relatively small number of hypogonadotropic hypogonadic patients with Kallmann's gene defects supports the hypothesis that other genes may be involved. Idiopathic hypogonadotropic hypogonadism (IHH) is not associated with anosmia. The GnRH gene was excluded as a candidate gene in IHH since no abnormality was found in several patients. The action of the GnRH is mediated through a G-protein coupled receptor present in the cell membrane of gonadotropes. The GnRH receptor was thus another candidate gene. Recently, we described the first patient with partial hypogonadotropic hypogonadism without anosmia caused by loss of function mutations of the GnRH receptor. We compare this first family with a new family presenting complete hypogonadotropic hypogonadism and a variable degree of gonadotrope deficiency in the affected kindred, and discuss genotype-phenotype correlation. PMID- 10698592 TI - ACTH resistance syndromes. AB - Inherited adrenocorticotropin (ACTH) insensitivity syndromes comprise a group of rare diseases in which resistance to ACTH is either the sole feature or associated with other symptoms. This review focuses on two autosomal recessive disorders, familial glucocorticoid deficiency (FGD) (MIM*202200) and the triple A syndrome (MIM*231550), which have at least three different molecular aetiologies. In FGD, several missense mutations within the coding region of the ACTH receptor (MC2-R) have been identified in some, but not all patients, and segregation analyses and functional studies in a Y6 cell expression system confirmed that these mutations cause the disease. Some cases of FGD are not linked to the MC2-R locus on chromosome 18p11.2 suggesting genetic heterogeneity. The triple A syndrome is clinically characterized by the triad of adrenal insufficiency, achalasia and alacrima and a variety of neurological symptoms. After excluding several candidate genes we mapped this syndrome to a 6 cM interval on chromosome 12q13 with no indication for genetic heterogeneity. The identification of the gene(s) causing FGD without mutations in the MC2-R and causing the triple A syndrome may reveal novel aspects in cell signalling and neuroendocrinology. PMID- 10698593 TI - Pathology of the TSH receptor. AB - Gain of function and loss of function mutations of the TSH receptor have been implicated in the pathogenesis of various thyroid diseases. Gain of function mutations, when somatic, are the first cause of autonomous nodules; when germline, they are responsible for hereditary non-autoimmune toxic thyroid hyperplasia and for some cases of sporadic congenital hyperthyroidism. A subset of mutations modifying the receptor selectivity have recently been found to be involved in the pathogenesis of familial gestational hyperthyroidism. These mutations are of great interest for understanding the mechanism of receptor activation. Loss of function mutations of the TSH receptor are responsible for different phenotypes ranging from asymptomatic resistance to TSH to overt congenital hypothyroidism. PMID- 10698594 TI - Hormone resistance caused by mutations in G proteins and G protein-coupled receptors. AB - G proteins couple receptors for many hormones to effectors that regulate second messenger metabolism. Several endocrine disorders have been shown to be caused by either loss or gain of function mutations in G proteins or G protein-coupled receptors. Pseudohypoparathyroidism (PHP), the first described example of a hormone resistance disorder, is characterized by renal resistance to parathyroid hormone (PTH) proximal to generation of the second messenger, cAMP. In PHP Ia there is more generalized hormone resistance (PTH, TSH, gonadotropins) and associated abnormal physical features, Albright hereditary osteodystrophy (AHO). Subjects with PHP Ib are normal in appearance and resistant exclusively to PTH. Germline loss of function mutations have been identified in the Gs-alpha gene in PHP Ia, and recent evidence suggests that the Gs-alpha gene is paternally imprinted in a tissue-specific manner. In PHP Ib, several studies have excluded PTH receptor gene mutations, and the molecular basis has not yet been defined. PMID- 10698595 TI - GH transcription factors. AB - The pituitary transcription factor Pit-1 is expressed during the later differentiation stages of anterior pituitary development and Pit-1 mutations have been identified as the cause of a combined pituitary hormone deficiency (CPHD) for GH, prolactin and TSH. Mutations within the human Pit-1 gene can either impair the DNA binding of this transcription factor, or while leaving DNA binding capabilities unimpaired, decrease its function within the transactivation complex. Approximately half of all patients with this phenotype do not show any defect within the Pit-1 gene. Prop-1, a recently discovered transcription factor of anterior pituitary development, seemed a likely candidate for such mutations. Prop-1 mutations, however, have been found so far to induce a combined pituitary hormone deficiency for GH, prolactin, TSH and gonadotropins. We describe here a group of patients with isolated and combined pituitary hormone deficiencies who were screened for Pit-1 and Prop-1 mutations to characterize the phenotypic spectrum of defects within these two genes. PMID- 10698596 TI - Development of the pituitary and its abnormalities. AB - The embryology of the pituitary gland and its normal development is described, including the importance of Pit-1. Congenital abnormalities of the hypothalamo pituitary region are discussed. PMID- 10698597 TI - Management of puberty in growth hormone deficient children. AB - The pubertal growth spurt accounts for approximately one-eighth of adult height and is regulated by complex hormonal interactions involving the somatotropic and gonadal axes. The observation that children with growth hormone deficiency (GHD) may fail to achieve an appropriate pubertal growth spurt led to the development of strategies to optimize GH therapy during puberty. In one strategy the dosage of GH is increased during puberty to support pubertal growth and in keeping with the physiological increase in serum levels of the hormone seen at that age. A different approach is to combine a GnRH analog (GnRHa) to GH to stop pubertal development, delaying epiphyseal fusion and prolonging peripubertal growth. Both strategies require caution. As regards the first strategy, too high doses of GH may shorten the pubertal time for growth; we found a small, nonsignificant, improvement in final height by increasing the dose by less than half. Preliminary results on the second strategy are more encouraging. However, manipulation of puberty should be limited to selected patients who show a statural height SDS for bone age unfavorable in terms of height prognosis. PMID- 10698598 TI - Changes in body composition during 12 months after discontinuation of growth hormone therapy in young adults with growth hormone deficiency from childhood. AB - Little is known about the velocity of changes in body composition after discontinuation of GH therapy at final height. The aim of this study was to describe the changes of fat distribution in male and female GH deficient young adults during the first year after discontinuation of GH therapy. Ten Dutch GH deficient young adults who had reached final height were retested and confirmed to be still GH deficient. These preliminary results demonstrate that the greatest gain in subcutaneous fat, measured by skinfold thickness, was observed in the first 3 months after GH withdrawal. Intra-abdominal fat was measured by CT scan at 0 and 12 months study time. The mean gain in intra-abdominal fat after 12 months was dramatically high (48%). We conclude that the subcutaneous and intra abdominal fat mass increased dramatically in young GH deficient Dutch adults with GH deficiency, after discontinuation of therapy, especially in the first three months. This indicates that GH therapy should be restarted as soon as possible, after reconfirming the diagnosis of GH deficiency in adults. This will reduce the risk for these patients of diseases associated with overweight, such as cardiovascular diseases. PMID- 10698599 TI - Gonadotropin treatment of hypogonadotropic hypogonadal adolescents. AB - Testosterone substitution, needed for normal physical development in male hypogonadal adolescents, does not induce testicular growth. We treated 37 hypogonadal adolescents with gonadotropins (hCG/hMG), to obtain complete virilization during the first two years of treatment, to avoid psychological sequellae and to allow normal sexual development. Testicular volume increased significantly during therapy (from 1.98 +/- 1.2 to 9 +/- 3.3 ml), while testosterone rose from 0.26 +/- 0.04 to 5.3 +/- 0.8 ng/ml, with worse results in adolescents with cryptorchidism. hCG/hMG treatment had a better outcome than testosterone during the induction of puberty, avoiding psychological problems induced by atrophic testes. Further long term studies are necessary to evaluate whether early hCG/hMG treatment facilitates later spermatogenesis even in patients with cryptorchidism. PMID- 10698600 TI - Growth hormone treatment of patients with Prader-Willi syndrome. Swedish Growth Hormone Advisory Group. AB - Several studies have now been published on the effect of one or several years of growth hormone treatment on growth and body composition of children with Prader Willi syndrome. The majority of the patients have responded with greatly increased growth rate, decreased body fat and increased muscle volume. Many of these children seem to have a functional growth hormone deficiency, probably secondary to their hypothalamic dysfunction. Further studies are needed to establish the long-term effect of this treatment on somatic and psychological well-being. PMID- 10698601 TI - Body composition during GH treatment in Prader-Labhardt-Willi syndrome. AB - Prader-Labhardt-Willi syndrome (PLWS) is a model to study GH secretion, body composition and consequences of GH therapy. Twenty-seven patients were studied by dual-energy X-ray absorptiometry (DXA) and were each compared to two age- and sex matched controls (obese and normal weight). Fat mass (FM) was significantly greater in PLWS than in patients with simple obesity; lean body mass (LM) and bone mineral content (BMC) were significantly lower compared to both controls. The peculiar body composition of PLWS patients seems to be similar to that found in GH deficiency. In six PLWS children treated with GH, LM increased after 6 months (p<0.02) up to 12 months (p<0.03); FM decreased in 5/6 patients. Obese adult PLWS patients treated with GH for 6 months showed a reduction in adiposity; LM increased significantly only in the leg compartment. Abdominal CT scan did not show a significant reduction of intrabdominal fat area. In conclusion, GH therapy might improve final stature and exert a positive influence on body composition in patients with PLWS. PMID- 10698602 TI - Developing the new professorate. PMID- 10698603 TI - Responses of baccalaureate and graduate programs to the emergence of choice in nursing accreditation. AB - Specialized accreditation in nursing is a widely recognized and respected hallmark of program quality. The advent of a second specialized accrediting agency for baccalaureate and higher degree programs in nursing prompted a survey of these programs to determine their choice of nursing accreditation agency, factors influencing their choice, their perceptions of the value added by nursing accreditation, and the difficulties encountered with the accreditation process. These study variables and the relationships between choice of accrediting agency and types of degree-granting nursing education programs offered by the institution, agency membership in the National League of Nursing (NLN) or the American Association of Colleges of Nursing (AACN), expected date of next accreditation visit, geographic region, public versus private status, and type of institution (Carnegie classification) were analyzed. Findings revealed that nearly a quarter (24%) of respondents intend to continue with the NLN Accrediting Commission (NLNAC), whereas 30% indicated they have already switched to the Commission on Collegiate Nursing Education (CCNE) or intend to do so prior to their next accreditation cycle. However, nearly a quarter (24%) of respondents said they plan to be accredited by both agencies for the immediate future, and 21% indicated they are still undecided. Study findings suggest an end to single source accreditation, and the beginning of a new market-oriented approach. PMID- 10698604 TI - Evaluation of graduates of an associate degree nursing program. AB - Approximately 58% of the RN nursing programs in the United States are associate degree programs (ADN). The curricula of these programs are designed to prepare graduates to competently provide direct patient care. The purpose of this descriptive comparative study was to evaluate the nursing performance of ADN graduates as perceived by graduates, faculty, and employers. In the first phase of the study, graduates and faculty completed the Six-Dimension Scale of Nursing Performance (6-DSNP) at the time of graduation. Six months later, the graduates again were asked to complete the same questionnaire along with their employers. Findings indicated a significant difference in the planning and evaluation performance of graduates at graduation and 6 months later as perceived by the graduates. Graduates rated themselves significantly higher in all areas of performance when compared to faculty and employers. There was no significant difference in the perception of graduate performance between faculty and employers. This outcome assessment provides needed data on perceptions of ADN programs. PMID- 10698605 TI - Wellness Wednesdays: health promotion and service learning on campus. AB - The Wellness Wednesdays program has shown itself to be an effective method for providing community-based service learning opportunities in a setting that is convenient for participants, students, and faculty. As the program continues to grow, it will provide opportunities for interdisciplinary collaboration with other health-related departments on campus. It also will provide a setting for faculty and student research in multiple areas related to management, health promotion, disease prevention, and health behavior motivation. PMID- 10698606 TI - Teaching feminist group process within a nursing curriculum. AB - Nurse educators are challenged to develop emancipatory teaching approaches that will create opportunities for students to develop their own praxis. In particular, the authors faced the challenge of teaching feminist group process within a curriculum based on phenomenology, feminism, and critical social theory. In this article, we discuss the challenges and rewards of teaching nursing and other students about feminist process through the creation of experiential learning opportunities. In addition, we highlight recommendations based on our own praxis. PMID- 10698607 TI - Teaching cancer principles to undergraduate students. AB - Cancer is the second leading cause of death in the United States. Most of the United States population will have some exposure to cancer during their lifetime. Medical knowledge becomes more advanced daily. Rapidly expanding knowledge specific to cancer etiology, diagnosis, and treatment is no exception. The consumer is at risk of misunderstanding this information at the time of the crisis because of the emotional, physical, and financial involvement. Gaining knowledge about cancer prior to such involvement may allow a better understanding if cancer care is required in the future. The Honors Oncology Course titled Learning to Live with Cancer was developed at the College of Nursing at Brigham Young University. It was developed to teach nursing and non-nursing students basic cancer principles, provide information regarding diagnosis and treatment of the disease, and allow students to work with cancer patients in the community to understand the impact of cancer on the patients, families, significant others, and the community at large. PMID- 10698608 TI - A comparison of self-reported cultural competency skills among two groups of nursing students: implications for nursing education. AB - This study was designed to examine self-reported cultural competency skills of second-semester junior-level nursing students toward clients from culturally diverse backgrounds. The purpose of this study was to ascertain if the addition of an innovative cultural sensitivity intervention facilitated greater self perceived cultural competency skills when compared with the traditional method of incorporating cultural diversity into a junior-level clinical course. The Ethnic Competency Skills Assessment Inventory was used to collect data from participants attending a university in an urban midwestern county. Significant differences were noted between the pretest scores and posttest scores. Pretest scores were significantly lower than posttest scores for both groups. Nurse educators must examine further the differences in learning experiences related to cultural diversity that may account for these differences. PMID- 10698609 TI - Attitudes of RN students toward obese adult patients. PMID- 10698610 TI - A survey of health policy content in Canadian graduate programs in nursing. PMID- 10698611 TI - Nurses' knowledge of diabetes. PMID- 10698612 TI - The development of conceptual categories of attention during the elementary school years. AB - The purpose of this study was to explore the development of children's conceptual understanding of attention by focusing on their awareness of various subtypes of attention. Adults, fifth graders (mean age = 11.16 years), and third graders (9.16 years) participated in a Similarity Judgment task in which they rated the similarity of pairs of cognitive scenarios involving various attentional processes. These similarity judgments were submitted to an extended similarity tree analysis (EXTREE; J. E. Corter & A. Tversky, 1986) and the interpretations of the clusters and features that emerged were confirmed by participants in an Attribute Rating task. Three changes were found with increasing age: (1) Less emphasis was placed on the surface features of the scenarios, (2) more attentional subtypes were identified, and (3) more emphasis was placed on the role of effort in attention. We conclude that during elementary school and beyond children came to understand that intentional, effortful cognitive processes can mediate between the external world and our conscious experiences of it. PMID- 10698613 TI - The ability to activate and inhibit speeded responses: separate developmental trends. AB - When children grow older they respond faster and are less susceptible to interference caused by task-irrelevant information. These observations suggested the hypothesis that a global mechanism may account for developmental change in the speed of responding and that inhibitory function may underlie the ability to activate speeded responses. The current study examined these issues by comparing the performance of 4 age groups (5-, 8-, and 11-year-olds and young adults) on a battery of 6 speeded performance tasks, 4 of which required the inhibition of response activation. An analysis of reaction and inhibition times supported a hypothesis of generalized developmental changes in response activation, but revealed a less pronounced development of inhibition. A nonselective mechanism of response inhibition seems to be fully developed during early childhood. PMID- 10698614 TI - Phonology acquired through the eyes and spelling in deaf children. AB - Hearing and deaf children, ranging in age from 6 years 8 months to 14 years 4 months, and matched for general spelling level, were required to spell high frequency and low-frequency words. Of interest was performance in relation to degree of exposure to Cued Speech (CS), which is a system delivering phonetically augmented speechreading through the visual modality. Groups were (a) hearing children, (b) deaf children exposed early and intensively to CS at home (CS Home), and (c) deaf children exposed to CS later and at school only (CS-School). Most of the spelling productions of hearing children as well as of CS-Home children were phonologically accurate for high-frequency as well as for low frequency words. CS-School children, who had less specified phonological representations, made a lower proportion of phonologically accurate spellings. These findings indicate that the accuracy of phonological representations, independent of the modality (acoustic versus visual) through which spoken language is perceived, determines the acquisition of phonology-to-orthography mappings. Analyses of the spelling productions indicate that the acquisition of orthographic representations of high precision depends on fully specified phonological representations. PMID- 10698615 TI - Abrogation of oral tolerance by feeding encapsulated antigen. AB - Results from previous studies have indicated that suppression of immune responses cannot be abrogated once oral tolerance has been established. Using a new methodology, OVA was encapsulated and fed to tolerized BDF1 mice. Results from these studies indicated that although IgG2a titers remained low, total IgG and IgG1 antibody titers were no longer suppressed compared to controls. Furthermore, in vitro splenocyte proliferation was not significantly suppressed in tolerized mice fed encapsulated OVA. To determine whether oral tolerance could be abrogated in other strains of mice, BALB/c mice were tolerized and fed encapsulated OVA. Results from these studies indicated that IgG2a as well as IgG1 and total anti OVA antibody titers were no longer suppressed. Although splenocyte proliferation did remain significantly suppressed in these mice, IFN-gamma and IL-4 levels were no longer decreased. To the best of our knowledge, this is the first time that abrogation of an established oral tolerance has been demonstrated. PMID- 10698616 TI - Cell-specific inhibition of inducible nitric oxide synthase activation by leflunomide. AB - The influence of a novel immunomodulating drug, leflunomide, on iNOS-dependent nitric oxide (NO) production in rodent macrophages and fibroblasts was investigated. Leflunomide's active metabolite A77 1726 caused a dose-dependent decrease of NO production in IFN-gamma-treated L929 fibroblasts. The observed effect was cell-specific, as well as stimulus-specific, since A77 1726 did not affect NO production in IFN-gamma-stimulated murine peritoneal macrophages or db cAMP-treated L929 cells. A77 1726 reduced expression of IFN-gamma-induced iNOS and IRF-1 mRNA in L929 cells, while iNOS enzymatic activity remained unchanged. Specific inhibitor of MAP kinase kinase (MEK), PD98059, but not unselective protein kinase inhibitor genistein, completely mimicked cell-type-specific and stimulus-specific NO-inhibitory action of leflunomide. Therefore, the recently described inhibition of MEK/MAP pathway by leflunomide could present a possible mechanism for its suppression of iNOS activation in L929 fibroblasts. Finally, a similar inhibitory effect of A77 1726 on both NO production and iNOS mRNA expression was observed also in IFN-gamma + LPS-activated murine and rat primary fibroblasts. PMID- 10698617 TI - Localization of pepstatin's inhibitory action during Fc-mediated antibody internalization: possible implications for antibody-mediated viral transmission. AB - Antibody internalization via Fc receptors is an important cellular mechanism, possibly facilitating the entry of antigenic peptides or viral particles into cells when specific antibodies are present at the periphery. Using an experimental model of trophoblast cells, we have shown that anti-p21(ras) monoclonal antibodies can use IFN-gamma-induced surface Fcgamma receptors to enter the cell. This entry of anti-p21(ras) antibodies ultimately inhibits IFN gamma-mediated class II antigen induction. Since there may be obvious and inevitable harmful aspects of this mechanism, during which Fc-mediated viral particle or autoantigen transport may occur, we concentrated efforts on defining a potent inhibitor able to eliminate such uptake. The results presented here show that the protease inhibitor pepstatin A efficiently inhibits Fcgamma receptor induction by IFN-gamma and also blocks the endocytic pathway followed by an antibody when it enters the cell at the level of early endosomal compartments. We thus postulate that the use of pepstatin A, because of its inhibition of autoantigen presentation or viral transmission, including that of HIV, may find important applications in therapeutic protocols. PMID- 10698618 TI - Single expression of CD45RC and RT6 in correlation with T-helper 1 and T-helper 2 cytokine patterns in the rat. AB - Previous studies involving the function and development of peripheral T cells have proposed that, in the rat, CD4(+)CD45RC(+)RT6(-) and CD4(+)CD45RC(-)RT6(+) T cell subsets may represent Th1 and Th2 cells, respectively. Here we tested this hypothesis directly by analyzing frequencies of IFN-gamma- and IL-4-producing cells in these two subpopulations using ELISPOT assays. We found that the CD4(+)CD45RC(-)RT6(+) subset showed higher numbers of IL-4-producing cells than the CD4(+)CD45RC(+)RT6(-) subset and, though less pronounced, that the latter demonstrated higher numbers of IFN-gamma producers. Therefore, we conclude that our results provide evidence for the existence of phenotypically defined Th1 and Th2 cells in the rat. This is supported by the finding that the ratios of IFN gamma/IL-4 and CD45RC/RT6 correlated positively among various rat strains. Finally, rat strains susceptible to induction of a Th1-mediated autoimmune disease showed the highest CD45RC/RT6 ratio, whereas the reverse was true for strains susceptible to a Th2-mediated autoimmune disease. PMID- 10698619 TI - Alpha 6 beta 1 integrin (VLA-6) mediates leukocyte tether and arrest on laminin under physiological shear flow. AB - Polymorphonuclear leukocytes (PMN) were perfused over extracellular matrix protein substrates under laminar shear flow. Under shear below 1.5 dyn/cm(2), many PMN tethered to immobilized laminin but not to fibronectin or vitronectin. Almost all the tethered PMN immediately arrested on laminin. The number of tethered PMN was mostly abrogated by mAbs to integrin alpha 6 or beta 1 chains at concentrations of more than 5 microg/ml. Addition of the two mAbs together produced no further inhibition compared with each mAb alone. In contrast, none of the mAbs to alpha 2, alpha 3, and beta 4 chains showed significant inhibition, indicating that PMN tethering to laminin is mostly dependent on alpha 6 beta 1 integrin. The addition of 10-100 ng/ml IL-8 in the assay medium before perfusion partially reduced PMN tethering to laminin. Stimulation with IL-8 also induced detachment of some tethered PMN within 30 s. Thus, IL-8 partially weakens the adhesiveness of alpha 6 beta 1 integrin on PMN in flow conditions. PMID- 10698620 TI - T lymphocytes and neutrophil granulocytes differ in regulatory signaling and migratory dynamics with regard to spontaneous locomotion and chemotaxis. AB - Chemotactic migration of T lymphocytes and neutrophil granulocytes within a three dimensional collagen matrix is distinct from spontaneous, matrix-induced migration concerning dynamic parameters and regulatory intracellular signaling. Both spontaneous T lymphocyte locomotion and stromal-cell-derived factor-1 (SDF 1)-induced chemotaxis-involved protein tyrosine kinase (PTK) activity, whereas only SDF-1-induced migration was protein kinase C (PKC) dependent. Spontaneous locomotion of neutrophil granulocytes was independent of PKC and PTK activity, but formyl-methionyl-leucyl-phenylalanine-induced migration involved PKC activity. In addition, the microtubule cytoskeleton was not changed after induction of chemotaxis in both cell types. T lymphocytes had a well-developed microtubule cytoskeleton with the microtubule organizing center located in the uropod, whereas neutrophil granulocytes revealed a clustered tubulin distribution at the leading edge of the migrating cell. Therefore, differences of the microtubule cytoskeleton might contribute to differences in locomotion between T lymphocytes and neutrophil granulocytes but not to differences between spontaneous locomotion and chemotaxis. PMID- 10698621 TI - Autocrine activation-induced cell death of T cells by human peripheral blood monocyte-derived CD4+ dendritic cells. AB - Mature T cells activated by antigen (Ag)-presenting cells are subject to various downmodulatory processes designed to maintain T cell homeostasis. Here we describe experiments in which mature T cells were subjected to apoptosis following stimulation with CD4(+) dendritic cells (DCs) during Ag presentation. The proliferative response of allogeneic T cells was increased by DCs at stimulator to responder (S/R) ratios ranging from 10(-3) to 1, whereas this response was decreased at S/R ratios ranging from 2 to 10. Allogeneic T cells stimulated with DCs at an S/R ratio of 5 underwent apoptosis, whereas this event was not observed in allogeneic T cells stimulated with DCs at an S/R ratio of 0.5. Stimulation of T cells with DCs at an S/R ratio of 5 induced a higher level of expression of CD95 ligand (CD95L) than stimulation of T cells cultured with DCs at an S/R ratio of 0.5, whereas similar levels of expression of CD28 and CD154 were observed in both cells. The abortive proliferation of mature T cells stimulated with DCs was prevented by blocking the CD95-CD95L system. Our results suggest that the CD4(+) DCs play counterregulatory roles in dictating T cell responses during Ag presentation. PMID- 10698622 TI - Activation-induced programmed cell death of nonspecific cytotoxic cells and inhibition by apoptosis regulatory factors. AB - Nonspecific cytotoxic cells (NCC) are the teleost equivalent of mammalian lymphokine-activated natural killer cells. The cytotoxic activities of NCC are enhanced by stress-activated serum factors (SASF) present in tilapia acute-phase serum. In the present study purified NCC and xenogeneic target HL-60 tumor cells and nuclei were distinguishable in mixtures determined by flow cytometry. NCC activated by target HL-60 cells undergo activation-induced programmed cell death (AIPCD) during 12- to 16-h killing assays as shown by Annexin-V binding and nuclear DNA fragmentation results. Annexin-V binding studies also demonstrated that NCC kill HL-60 cells by an apoptotic mechanism. NCC are protected from AIPCD by 4-h preincubation in 50% SASF. Pretreatment also produced more than a fourfold increase in NCC cytotoxicity (effector/target (E:T) ratio = 100:1). In the absence of SASF preincubation, the percentage of apoptotic NCC increased from 8 to 91% at E:T ratios of 1:0 and 1:1, respectively. Kinetic studies (E:T = 10:1) demonstrated that the percentage of NCC exhibiting HL-60-dependent AIPCD increased between 0.1 and 12 h and then decreased inversely with total cell necrosis over the next 60 h. Preincubation of NCC with SASF protected NCC from AIPCD for over 72 h. Crosslinkage of the NCCRP-1 receptor with monoclonal antibody (mab) 5C6 produced AIPCD between 1 and 100 microg/mL mab concentrations. Preincubation with SASF completely protected NCC from mab 5C6-dependent AIPCD. SASF-mediated protection of NCC from AIPCD was dependent upon divalent cations, as demonstrated by increases in DNA hypoploidy of 38, 67, and 88% following preincubation in the presence of 10, 100, and 1000 microM EDTA, respectively. SASF also protected NCC from glucocorticoid- (i. e., dexamethasone) induced apoptosis. Combined, these results demonstrated that NCC activity is down regulated by AIPCD. Release of SASF into the peripheral circulation may prevent negative regulation of NCC by AIPCD by increasing recycling capacity. Results are discussed in the context of the effects of acute stressors on innate immunity. PMID- 10698623 TI - A cruciform structural transition provides a molecular switch for chromosome structure and dynamics. AB - The interaction between specific sites along a DNA molecule is often crucial for the regulation of genetic processes. However, mechanisms regulating the interaction of specific sites are unknown. We have used atomic force microscopy to demonstrate that the structural transition between cruciform conformations can act as a molecular switch to facilitate or prevent communication between distant regions in DNA. Cruciform structures exist in vivo and they are critically involved in the initiation of replication and the regulation of gene expression in different organisms. Therefore, structural transitions of the cruciform may play a key role in these processes. PMID- 10698624 TI - Site-specific recombination in human cells catalyzed by phage lambda integrase mutants. AB - Phage lambda Integrase (Int) is the prototype of the so-called integrase family of conservative site-specific recombinases, which includes Cre and FLP. The natural function of Int is to execute integration and excision of the phage into and out of the Escherichia coli genome, respectively. In contrast to Cre and FLP, however, wild-type Int requires accessory proteins and DNA supercoiling of target sites to catalyze recombination. Here, we show that two mutant Int proteins, Int h (E174 K) and its derivative Int-h/218 (E174 K/E218 K), which do not require accessory factors, are proficient to perform intramolecular integrative and excisive recombination in co-transfection assays inside human cells. Intramolecular integrative recombination is also detectable by Southern analysis in human reporter cell lines harboring target sites attB and attP as stable genomic sequences. Recombination by wild-type Int, however, is not detectable by this method. The latter result implies that eukaryotic co-factors, which could functionally replace the prokaryotic ones normally required for wild-type Int, are most likely not present in human cells. PMID- 10698625 TI - Identifying the protein folding nucleus using molecular dynamics. AB - Molecular dynamics simulations of folding in an off-lattice protein model reveal a nucleation scenario, in which a few well-defined contacts are formed with high probability in the transition state ensemble of conformations. Their appearance determines folding cooperativity and drives the model protein into its folded conformation. Amino acid residues participating in those contacts may serve as "accelerator pedals" used by molecular evolution to control protein folding rate. PMID- 10698626 TI - HPV oncoprotein E6 is a structure-dependent DNA-binding protein that recognizes four-way junctions. AB - E6 is an oncoprotein implicated in cervical cancers, produced by "high-risk" human papillomaviruses. E6 is thought to promote tumorigenesis by stimulating cellular degradation of the tumour suppressor p53, but it might display other activities. Sequence similarity was recently detected between E6 and endonuclease VII, a protein of phage T4 that recognizes and cleaves four-way DNA junctions. Here, we purified recombinant E6 proteins and demonstrated that high-risk E6 s bind selectively to four-way junctions in a structure-dependent manner. Several residues in the C-terminal zinc-binding domain, the region of E6 similar to endonuclease VII, are necessary for the junction-binding activity. E6 binds to the junction as a monomer. Comparative electrophoresis shows that E6-bound junctions migrate in an extended square conformation. Magnesium inhibits the electrophoretic migration of the complexes but does not seem to influence their formation at equilibrium. This work is the first demonstration of specific binding of purified active E6 to a well-characterized DNA ligand, and suggests new modes of action of E6 in oncogenesis. PMID- 10698627 TI - Computational identification of cis-regulatory elements associated with groups of functionally related genes in Saccharomyces cerevisiae. AB - AlignACE is a Gibbs sampling algorithm for identifying motifs that are over represented in a set of DNA sequences. When used to search upstream of apparently coregulated genes, AlignACE finds motifs that often correspond to the DNA binding preferences of transcription factors. We previously used AlignACE to analyze whole genome mRNA expression data. Here, we present a more detailed study of its effectiveness as applied to a variety of groups of genes in the Saccharomyces cerevisiae genome. Published functional catalogs of genes and sets of genes grouped by common name provided 248 groups, resulting in 3311 motifs. In conjunction with this analysis, we present measures for gauging the tendency of a motif to target a given set of genes relative to all other genes in the genome and for gauging the degree to which a motif is preferentially located in a certain distance range upstream of translational start sites. We demonstrate improved methods for comparing and clustering sequence motifs. Many previously identified cis-regulatory elements were found. We also describe previously unidentified motifs, one of which has been verified by experiments in our laboratory. An extensive set of AlignACE runs on randomly selected sets of genes and on sets of genes whose upstream regions contain known transcription factor binding sites serve as controls. PMID- 10698628 TI - Crystal structure of the DNA polymerase processivity factor of T4 bacteriophage. AB - The protein encoded by gene 45 of T4 bacteriophage (gene 45 protein or gp45), is responsible for tethering the catalytic subunit of T4 DNA Polymerase to DNA during high-speed replication. Also referred to as a sliding DNA clamp, gp45 is similar in its function to the processivity factors of bacterial and eukaryotic DNA polymerases, the beta-clamp and PCNA, respectively. Crystallographic analysis has shown that the beta-clamp and PCNA form highly symmetrical ring-shaped structures through which duplex DNA can be threaded. Gp45 shares no sequence similarity with beta-clamp or PCNA, and sequence comparisons have not been able to establish whether it adopts a similar structure. We have determined the crystal structure of gp45 from T4 bacteriophage at 2.4 A resolution, using multiple isomorphous replacement. The protein forms a trimeric ring-shaped assembly with overall dimensions that are similar to those of the bacterial and eukaryotic processivity factors. Each monomer of gp45 contains two domains that are very similar in chain fold to those of beta-clamp and PCNA. Despite an overall negative charge, the inner surface of the ring is in a region of positive electrostatic potential, consistent with a mechanism in which DNA is threaded through the ring. PMID- 10698629 TI - The high resolution crystal structure of green abalone sperm lysin: implications for species-specific binding of the egg receptor. AB - Abalone sperm lysin is a 16 kDa acrosomal protein used by sperm to create a hole in the egg vitelline envelope. Lysins from seven California abalone exhibit species-specificity in binding to their egg receptor, and range in sequence identity from 63 % to 90 %. The crystal structure of the sperm lysin dimer from Haliotis fulgens (green abalone) has been determined to 1.71 A by multiple isomorphous replacement. Comparisons with the structure of the lysin dimer from Haliotis rufescens (red abalone) reveal a similar overall fold and conservation of features contributing to lysin's amphipathic character. The two structures do, however, exhibit differences in surface residues and electrostatics. A large clustering of non-conserved surface residues around the waist and clefts of the dimer, and differences in charged residues around these regions, indicate areas of the molecule which may be involved in species-specific egg recognition. PMID- 10698630 TI - The 1.3 A crystal structure of a biotin-binding pseudoknot and the basis for RNA molecular recognition. AB - A pseudoknot-containing aptamer isolated from a pool of random sequence molecules has been shown previously to represent an optimal RNA solution to the problem of binding biotin. The affinity of this RNA molecule is nonetheless orders of magnitude weaker than that of its highly evolved protein analogs, avidin and streptavidin. To understand the structural basis for biotin binding and to compare directly strategies for ligand recognition available to proteins and RNA molecules, we have determined the 1.3 A crystal structure of the aptamer complexed with its ligand. Biotin is bound at the interface between the pseudoknot's stacked helices in a pocket defined almost entirely by base-paired nucleotides. In comparison to the protein avidin, the aptamer packs more tightly around the biotin headgroup and makes fewer contacts with its fatty acid tail. Whereas biotin is deeply buried within the hydrophobic core in the avidin complex, the aptamer relies on a combination of hydrated magnesium ions and immobilized water molecules to surround its ligand. In addition to demonstrating fundamentally different approaches to molecular recognition by proteins and RNA, the structure provides general insight into the mechanisms by which RNA function is mediated by divalent metals. PMID- 10698631 TI - Crystal structure of Lysbeta(1)82-Lysbeta(2)82 crosslinked hemoglobin: a possible allosteric intermediate. AB - The crystal structure of human hemoglobin crosslinked between the Lysbeta82 residues has been determined at 2.30 A resolution. The crosslinking reaction was performed under oxy conditions using bis(3, 5-dibromosalicyl) fumarate; the modified hemoglobin has increased oxygen affinity and lacks cooperativity. Since the crystallization occurred under deoxy conditions, the resulting structure displays conformational characteristics of both the (oxy) R and the (deoxy) T states. beta82XLHbA does not fully reach its T-state conformation due to the presence of the crosslink. The R-state-like characteristics of deoxy beta82XLHbA include the position of the distal Hisbeta63 (E7) residue, indicating a possible reason for the high oxygen affinity of this derivative. Other areas of the molecule, particularly those thought to be important in the allosteric transition, such as Tyrbeta145 (HC2) and the switch region involving Proalpha(1)44 (CD2), Thralpha(1)41 (C6) and Hisbeta(2)97 (FG4), are in intermediate positions between the R and T-states. Thus, the structure may represent a stabilized intermediate in the allosteric transition of hemoglobin. PMID- 10698632 TI - Towards a complete description of the structural and dynamic properties of the denatured state of barnase and the role of residual structure in folding. AB - The detailed characterization of denatured proteins remains elusive due to their mobility and conformational heterogeneity. NMR studies are beginning to provide clues regarding residual structure in the denatured state but the resulting data are too sparse to be transformed into molecular models using conventional techniques. Molecular dynamics simulations can complement NMR by providing detailed structural information for components of the denatured ensemble. Here, we describe three independent 4 ns high-temperature molecular dynamics simulations of barnase in water. The simulated denatured state was conformationally heterogeneous with respect to the conformations populated both within a single simulation and between simulations. Nonetheless, there were some persistent interactions that occurred to varying degrees in all simulations and primarily involved the formation of fluid hydrophobic clusters with participating residues changing over time. The region of the beta(3-4) hairpin contained a particularly high degree of such side-chain interactions but it lacked beta structure in two of the three denatured ensembles: beta(3-4) was the only portion of the beta-structure to contain significant residual structure in the denatured state. The two principal alpha-helices (alpha1 and alpha2) adopted dynamic helical structure. In addition, there were persistent contacts that pinched off core 2 from the body of the protein. The rest of the protein was unstructured, aside from transient and mostly local side-chain interactions. Overall, the simulated denatured state contains residual structure in the form of dynamic, fluctuating secondary structure in alpha1 and alpha2, as well as fluctuating tertiary contacts in the beta(3-4) region, and between alpha1 and beta(3-4), in agreement with previous NMR studies. Here, we also show that these regions containing residual structure display impaired mobility by both molecular dynamics and NMR relaxation experiments. The residual structure was important in decreasing the conformational states available to the chain and in repairing disrupted regions. For example, tertiary contacts between beta(3-4) and alpha1 assisted in the refolding of alpha1. This contact-assisted helix formation was confirmed in fragment simulations of beta(3-4) and alpha1 alone and complexed, and, as such, alpha1 and beta(3-4) appear to be folding initiation sites. The role of these sites in folding was investigated by working backwards and considering the simulation in reverse, noting that earlier time-points from the simulations provide models of the major intermediate and transition states in quantitative agreement with data from both unfolding and refolding experiments. Both beta(3-4) and alpha1 are dynamic in the denatured state but when they collide and make enough contacts, they provide a loose structural scaffold onto which further beta-strands pack. The beta-structure condenses about beta(3-4), while alpha1 aids in stabilizing beta(3-4) and maintaining its orientation. The resulting beta-structure is relatively planar and loose in the major intermediate. Further packing ensues, and as a result the beta-sheet twists, leading to the major transition state. The structure is still expanded and loops are not well formed at this point. Fine-tuning of the packing interactions and the final condensation of the structure then occurs to yield the native state. PMID- 10698633 TI - Energetic and structural interactions between delta-dendrotoxin and a voltage gated potassium channel. AB - Dendrotoxin proteins isolated from Mamba snake venom block potassium channels with a high degree of specificity and selectivity. Using site-directed mutagenesis we have identified residues that constitute the functional interaction surfaces of delta-dendrotoxin and its voltage-gated potassium channel receptor. delta-Dendrotoxin uses a triangular patch formed by seven side-chains (Lys3, Tyr4, Lys6, Leu7, Pro8, Arg10, Lys26) to block K(+) currents carried by a Shaker potassium channel variant. The inhibitory surface of the toxin interacts with channel residues at Shaker positions 423, 425, 427, 431, and 449 near the pore. Amino acid mutations that interact across the toxin-channel interface were identified by mutant cycle analysis. These results constrain the possible orientation of dendrotoxin with respect to the K(+) channel structure. We propose that dendrotoxin binds near the pore entryway but does not act as a physical plug. PMID- 10698634 TI - Conformational changes preceding decapsidation of bromegrass mosaic virus under hydrostatic pressure: a small-angle neutron scattering study. AB - The stability of bromegrass mosaic virus (BMV) and empty shells reassembled in vitro from purified BMV coat protein was investigated under hydrostatic pressure, using solution small-angle neutron scattering. This technique allowed us to monitor directly the dissociation of the particles, and to detect conformational changes preceding dissociation. Significant dissociation rates were observed only if virions swelled upon increase of pressure, and pressure effects became irreversible at very high-pressure in such conditions. At pH 5.0, in buffers containing 0.5 M NaCl and 5 mM MgCl(2), BMV remained compact (radius 12.9 nm), dissociation was limited to approximately 10 % at 200 MPa, and pressure effects were totally reversible. At pH 5.9, BMV particles were slightly swollen under normal pressure and swelling increased with pressure. The dissociation was reversible to 90 % for pressures up to 160 MPa, where its rate reached 28 %, but became totally irreversible at 200 MPa. Pressure-induced swelling and dissociation increased further at pH 7.3, but were essentially irreversible. The presence of (2)H(2)O in the buffer strongly stabilized BMV against pressure effects at pH 5.9, but not at pH 7.3. Furthermore, the reversible changes of the scattered intensity observed at pH 5.0 and 5.9 provide evidence that pressure could induce the release of coat protein subunits, or small aggregates of these subunits from the virions, and that the dissociated components reassociated again upon return to low pressure. Empty shells were stable at pH 5.0, at pressures up to 260 MPa. They became ill-shaped at high-pressure, however, and precipitated slowly after return to normal conditions, providing the first example of a pressure-induced conformational drift in an assembled system. PMID- 10698635 TI - Microsecond to minute dynamics revealed by EX1-type hydrogen exchange at nearly every backbone hydrogen bond in a native protein. AB - A previous comprehensive analysis of the pH dependence of native-state amide hydrogen (NH) exchange in turkey ovomucoid third domain (OMTKY3) yielded apparent opening and closing rate constants (k(op) and k(cl)) at 14 NH groups involved in global conformational changes. This analysis has been extended to 18 additional slowly exchanging NH groups. Quench-flow experiments were performed to monitor NH exchange in native OMTKY3 from neutral to very alkaline pH ( approximately 12) conditions. Above pH 10 the mechanism of exchange switched from one governed by a rapid equilibrium preceding the chemistry of exchange (i.e. EX2 exchange), to one where exchange was limited by the rate of opening (i.e. EX1 exchange). Kinetics of solvent exposure are now known for nearly all backbone NH groups in native OMTKY3, yielding rate constants that span five orders of magnitude, 0.004 to 200 s(-1). PMID- 10698636 TI - Combination of threading potentials and sequence profiles improves fold recognition. AB - Using a benchmark set of structurally similar proteins, we conduct a series of threading experiments intended to identify a scoring function with an optimal combination of contact-potential and sequence-profile terms. The benchmark set is selected to include many medium-difficulty fold recognition targets, where sequence similarity is undetectable by BLAST but structural similarity is extensive. The contact potential is based on the log-odds of non-local contacts involving different amino acid pairs, in native as opposed to randomly compacted structures. The sequence profile term is that used in PSI-BLAST. We find that combination of these terms significantly improves the success rate of fold recognition over use of either term alone, with respect to both recognition sensitivity and the accuracy of threading models. Improvement is greatest for targets between 10 % and 20 % sequence identity and 60 % to 80 % superimposable residues, where the number of models crossing critical accuracy and significance thresholds more than doubles. We suggest that these improvements account for the successful performance of the combined scoring function at CASP3. We discuss possible explanations as to why sequence-profile and contact-potential terms appear complementary. PMID- 10698637 TI - Theoretical study on mutation-induced activation of the luteinizing hormone receptor. AB - Here, three-dimensional model building and molecular dynamics simulations of the luteinizing hormone receptor have been employed to generate hypotheses about the molecular mechanisms underlying the activation of the receptor induced by naturally occurring activating mutations. The comparative analysis of the wild type receptor and of 16 constitutively active or inactive mutants has been instrumental in inferring the structural/dynamic features which could characterize the inactive and the active forms of the receptor. These features have been also employed for predicting the functional behavior of new receptor mutants. The results of this study might provide a structural framework to interpret the pathological effects induced by mutations of the luteinizing hormone receptor. In addition, the proposed theoretical model could be useful for engineering new mutations or ligands able to modulate receptor function. PMID- 10698638 TI - Paramyxovirus F1 protein has two fusion peptides: implications for the mechanism of membrane fusion. AB - Viral fusion proteins contain a highly hydrophobic segment, named the fusion peptide, which is thought to be responsible for the merging of the cellular and viral membranes. Paramyxoviruses are believed to contain a single fusion peptide at the N terminus of the F1 protein. However, here we identified an additional internal segment in the Sendai virus F1 protein (amino acids 214-226) highly homologous to the fusion peptides of HIV-1 and RSV. A synthetic peptide, which includes this region, was found to induce membrane fusion of large unilamellar vesicles, at concentrations where the known N-terminal fusion peptide is not effective. A scrambled peptide as well as several peptides from other regions of the F1 protein, which strongly bind to membranes, are not fusogenic. The functional and structural characterization of this active segment suggest that the F1 protein has an additional internal fusion peptide that could participate in the actual fusion event. The presence of homologous regions in other members of the same family suggests that the concerted action of two fusion peptides, one N-terminal and the other internal, is a general feature of paramyxoviruses. PMID- 10698639 TI - Immunoglobulin superfamily proteins in Caenorhabditis elegans. AB - The predicted proteins of the genome of Caenorhabditis elegans were analysed by various sequence comparison methods to identify the repertoire of proteins that are members of the immunoglobulin superfamily (IgSF). The IgSF is one of the largest families of protein domain in this genome and likely to be one of the major families in other multicellular eukaryotes too. This is because members of the superfamily are involved in a variety of functions including cell-cell recognition, cell-surface receptors, muscle structure and, in higher organisms, the immune system. Sixty-four proteins with 488 I set IgSF domains were identified largely by using Hidden Markov models. The domain architectures of the protein products of these 64 genes are described. Twenty-one of these had been characterised previously. We show that another 25 are related to proteins of known function. The C. elegans IgSF proteins can be classified into five broad categories: muscle proteins, protein kinases and phosphatases, three categories of proteins involved in the development of the nervous system, leucine-rich repeat containing proteins and proteins without homologues of known function, of which there are 18. The 19 proteins involved in nervous system development that are not kinases or phosphatases are homologues of neuroglian, axonin, NCAM, wrapper, klingon, ICCR and nephrin or belong to the recently identified zig gene family. Out of the set of 64 genes, 22 are on the X chromosome. This study should be seen as an initial description of the IgSF repertoire in C. elegans, because the current gene definitions may contain a number of errors, especially in the case of long sequences, and there may be IgSF genes that have not yet been detected. However, the proteins described here do provide an overview of the bulk of the repertoire of immunoglobulin superfamily members in C. elegans, a framework for refinement and extension of the repertoire as gene and protein definitions improve, and the basis for investigations of their function and for comparisons with the repertoires of other organisms. PMID- 10698640 TI - Frequency-selective quantification of biomedical magnetic resonance spectroscopy data. AB - In this paper the possibility of obtaining accurate estimates of parameters of selected peaks in the presence of unknown or uninteresting spectral features in biomedical magnetic resonance spectroscopy (MRS) signals is investigated. This problem is denoted by frequency-selective parameter estimation. A new time-domain technique based on maximum-phase finite impulse response (FIR) filters is presented. The proposed method is compared to a number of existing approaches: the application of a weighting function in the time domain, frequency domain fitting using a polynomial baseline, and the time-domain HSVD filter method. The ease of use and low computational complexity of the FIR filter method make it an attractive approach for frequency-selective parameter estimation. The methods are validated using simulations of relevant (13)C and (31)P MRS examples. PMID- 10698641 TI - Three-dimensional (13)C-spectroscopic imaging in the isolated infarcted rat heart. AB - Acquisition weighted (13)C-spectroscopic imaging with three spatial dimensions is demonstrated in the isolated, perfused rat heart. Experiments were performed at 11.75 T with a home-built double resonant (13)C-(1)H probehead. Three-dimensional chemical shift imaging was used to obtain (1)H-decoupled (13)C-spectra in 96 microl voxels in about 58 min. Acquisition weighting significantly reduced signal contamination and improved image quality, with no penalty in sensitivity. As a first application, infarcted hearts were studied during perfusion with [2-(13)C] sodium acetate. The extent of the incorporation of the (13)C-label into glutamate allows us to distinguish intact and infarcted myocardium. Chemical shift images show a homogeneous glutamate distribution in intact tissue, but a negligible amount in the infarction scar. PMID- 10698642 TI - Diffusion-weighted imaging of bacteria colonies in the STRAFI plane. AB - Imaging colonies of bacteria in water suspension using NMR requires that the water inside the bacteria can be differentiated from the surrounding water. This is commonly carried out by using diffusion-weighted pulsed field gradient techniques. However, it is also possible to use the diffusion sensitivity inherent in stray field imaging (STRAFI). In STRAFI, the subject to be imaged is normally moved along the axis of a superconducting magnet so that it passes through the sensitive slice. However, by moving the sample in the transverse direction and by using a long copper strip in place of a surface induction coil, a diffusion-weighted one-dimensional projection profile can be obtained across the sensitive slice. Profiles from a water phantom and from a bacteria suspension show convincing discrimination between intracellular and extracellular water. PMID- 10698643 TI - Improved Shinnar-Le Roux algorithm. AB - Selective excitation pulses are widely used in magnetic resonance imaging in order to excite predetermined slices of the body under examination. Such pulses are optimally designed by means of the Shinnar-Le Roux algorithm. In this paper, we show that under minimal assumptions, the complexity and computing cost of the original Shinnar-Le Roux algorithm can be drastically reduced. We further propose an improved version of the algorithm, involving only real quantities, which is both easier to implement and faster to execute, so it is suitable for implementation at the hardware level, in the context of a real-time fully digital magnetic resonance transceiver. PMID- 10698644 TI - Decay of dipolar order in diamagnetic and paramagnetic proteins and protein gels. AB - Magnetic relaxation in solids may be complicated by the creation and loss of dipolar order at finite rates. In tissues the molecular and spin dynamics may be significantly different because of the relatively high concentration of water. We have applied a modified Jeneer-Broekaert pulse sequence to measure dipolar relaxation rates in both dry and hydrated protein systems that may serve as magnetic models for tissue. In lyophilized and dry serum albumin, the dipolar relaxation time, T(1D) is on the order of 1 ms and is consistent with earlier reports. When hydrated by deuterium oxide, the dipolar relaxation times measured were on the order of tens of microseconds. When paramagnetic centers are included in the protein, the Jeneer-Broekaert echo decay times became the order of the decay time for transverse magnetization, i.e., the order of 10 micros or less. In the hydrated or paramagnetic systems, the dipolar relaxation times are too short to require inclusion in the quantitative analysis of magnetization transfer experiments. PMID- 10698645 TI - 17O-decoupled (1)H spectroscopy and imaging with a surface coil: STEAM decoupling. AB - (17)O-decoupled (1)H spin-echo imaging has been reported as a means of indirect (17)O detection, with potential application to measurement of blood flow and metabolism. In its current form, (17)O decoupling requires large RF amplitudes and a 180 degrees refocusing pulse, complicating its application in volume and surface coils, respectively. To overcome this problem, we have developed an (17)O decoupled proton stimulated echo sequence ("STEAM decoupling") to allow (17)O detection with a surface coil. A high B(1) amplitude is easily generated, allowing complete decoupling of (17)O and (1)H. Slice-selective, (17)O-decoupled (1)H imaging is readily performed and the sequence is easily adapted for localized spectroscopy. Intrinsic correction for variations in B(1) and further compensation for B(1) inhomogeneity are discussed. PMID- 10698646 TI - Cross-correlated relaxation for the measurement of angles between tensorial interactions. AB - Theory,experimental aspects, and use in structure calculation of cross-correlated relaxation rates measured on zero- and double-quantum coherences in liquid state NMR are presented. The relative size of the interaction depends on the projection angle between the two tensorial interactions. The tensorial interaction can be either a dipolar interaction or a chemical shift anisotropy relaxation mechanism (CSA). Effects of additional sources of relaxation on the cross-correlated relaxation rates are analyzed. Also, an easy-to-use formalism is given to manipulate different cross-correlated relaxation interactions. The application addresses measurement of the backbone angle psi in a protein by measuring dipole((15)N-(1)H)-dipole((13)C(alpha)-(1)H(alpha)) and CSA((15)N) dipole((13)C(alpha)-(1)H(alpha)) cross-correlated relaxation rates. It is shown that ambiguities due to the 3 cos(2)θ-1 dependence of one cross-correlated relaxation rate can be overcome by measuring additional cross-correlated relaxation rates. The use of cross-correlated relaxation rates is demonstrated in structure calculations. PMID- 10698647 TI - Small volume flow probe for automated direct-injection NMR analysis: design and performance. AB - A detailed characterization of an NMR flow probe for use in direct-injection sample analysis is presented. A 600-MHz, indirect detection NMR flow probe with a 120-microl active volume is evaluated in two configurations: first as a stand alone small volume probe for the analysis of static, nonflowing solutions, and second as a component in an integrated liquids-handling system used for high throughput NMR analysis. In the stand-alone mode, (1)H lineshape, sensitivity, radiofrequency (RF) homogeneity, and heat transfer characteristics are measured and compared to conventional-format NMR probes of related design. Commonly used descriptive terminology for the hardware, sample regions, and RF coils are reviewed or defined, and test procedures developed for flow probes are described. The flow probe displayed general performance that is competitive with standard probes. Key advantages of the flow probe include high molar sensitivity, ease of use in an automation setup, and superior reproducibility of magnetic field homogeneity which enables the practical implementation of 1D T2-edited analysis of protein-ligand interactions. PMID- 10698648 TI - A new class of contrast agents for MRI based on proton chemical exchange dependent saturation transfer (CEST). AB - It has been previously shown that intrinsic metabolites can be imaged based on their water proton exchange rates using saturation transfer techniques. The goal of this study was to identify an appropriate chemical exchange site that could be developed for use as an exogenous chemical exchange dependent saturation transfer (CEST) contrast agent under physiological conditions. These agents would function by reducing the water proton signal through a chemical exchange site on the agent via saturation transfer. The ideal chemical exchange site would have a large chemical shift from water. This permits a high exchange rate without approaching the fast exchange limit at physiological pH (6.5-7.6) and temperature (37 degrees C), as well as minimizing problems associated with magnetic field susceptibility. Numerous candidate chemicals (amino acids, sugars, nucleotides, heterocyclic ring chemicals) were evaluated in this preliminary study. Of these, barbituric acid and 5, 6-dihydrouracil were more fully characterized with regard to pH, temperature, and concentration CEST effects. The best chemical exchange site found was the 5.33-ppm indole ring -NH site of 5-hydroxytryptophan. These data demonstrate that a CEST-based exogenous contrast agent for MRI is feasible. PMID- 10698649 TI - High-field, multifrequency EPR spectroscopy using whispering gallery dielectric resonators. AB - High-field/high frequency EPR spectroscopy measurements are shown. Experiments were carried out at 240- and 316-GHz frequencies. The employed apparatus uses a novel combination of far infrared molecular lasers and of probehead exploiting dielectric resonators working in the whispering gallery modes. This approach constitutes a relatively simple method of multifrequency EPR spectroscopy and opens appealing perspectives in high-sensitivity EPR spectroscopy up to the THz regime. PMID- 10698650 TI - Simultaneous spectral editing for gamma-aminobutyric acid and taurine using double quantum coherence transfer. AB - Conventional double quantum (DQ) editing techniques recover resonances of one metabolite at a time and are thus inefficient for monitoring metabolic changes involving several metabolites. A DQ coherence transfer double editing sequence using a dual-band DQ coherence read pulse is described here. The sequence permits simultaneous spectral editing for two metabolites with similar J coupling constants in a single scan. Simultaneous editing for taurine and gamma aminobutyric acid (GABA) is demonstrated using solution phantoms and rat brain tissue. Selectivity of the double editing sequence for the target metabolites is as good as that achieved using conventional DQ editing which selects each metabolite individually. With experimental parameters of the double editing sequence chosen to optimize GABA editing, the sensitivity for GABA detection is the same as that with GABA editing only, while the sensitivity for taurine detection is decreased slightly compared to that with taurine editing only. PMID- 10698651 TI - Diffusion measurements using the nonlinear stimulated echo. AB - The nonlinear stimulated echo that is generated by a sequence of three radiofrequency pulses, 90 degrees-tau(1)-90 degrees-tau(2)-45 degrees, in high magnetic fields (or at low temperatures) in the presence of pulsed or steady field gradients can be applied for measurements of the diffusion coefficient. Corresponding test experiments are reported. Steady gradients can be used without knowledge of the relaxation times. Remarkably the attenuation of the nonlinear stimulated echo by diffusion is substantially stronger than in the case of the ordinary stimulated echo. PMID- 10698652 TI - The Bloch equations in high-gradient magnetic resonance force microscopy: theory and experiment. AB - We report theory and observations of paramagnetic resonance in a measured field gradient of 44,000 T per meter by the technique of magnetic resonance force microscopy (MRFM). Resonance was induced in a dilute solid solution of diphenylpicrylhydrazyl in polystyrene at 77 and 10 K by an amplitude-modulated microwave field. This modulated the force between resonant sample spins and a micrometer-scale SmCo magnetic tip on a force microscope cantilever. The force signals were typically of order 10 fN, and were detected above a thermal noise floor of 80 aN per root hertz at 10 K, equivalent to a magnetic moment noise of 200 micro(B) per root hertz of bandwidth. Resonance saturation was readily observed. Starting with the Bloch equations, we derived simple analytic expressions for the predicted cantilever signal amplitudes and T(1)-dependent phase lags, valid at low microwave power levels. For power levels below saturation, the data were in good agreement with the Bloch equation predictions, while above saturation the measured force increased more slowly with power than predicted. Several ESR mechanisms which might lead to non-Bloch dynamics in the MRFM environment are reviewed. Spin-relaxation mechanisms are also reviewed. A detailed description of the experimental apparatus is offered. PMID- 10698653 TI - Spin dynamics of Carr-Purcell-Meiboom-Gill-like sequences in grossly inhomogeneous B(0) and B(1) fields and application to NMR well logging. AB - The spin dynamics for Carr-Purcell-Meiboom-Gill-like sequences is analyzed in grossly inhomogeneous B(0) and B(1) fields. This problem is important for many applications, especially when the bandwidth of the signal is excitation limited. Examples include stray-field NMR or inside-out NMR probes used in well logging. The amplitudes of the first few echoes exhibit a characteristic transient behavior but quickly approach a smooth asymptotic behavior. For simple Hamiltonians without scalar or dipolar couplings, the evolution of a refocusing subcycle for a given isochromat is described by a rotation. Simple expressions for the signal of the Nth echo are derived in terms of these effective rotations that have a simple geometrical interpretation. It is shown that the asymptotic behavior is controlled by the direction of the axis of these effective rotations and the signal is dominated by magnetization "spin-locked" to the rotation axis. The phase of the signal is independent of the details of the field inhomogeneities. Relaxation in inhomogeneous fields leads to a signal decay that is in general nonexponential with an initial decay rate that is a weighted sum of T(-1)(1) and T(-1)(2). At long times, the echo amplitudes decay to a finite value. Phase cycling eliminates this offset. The effect of diffusion is also studied. This analysis has been applied to an inside-out NMR well logging apparatus. Good quantitative agreement is found between measurements and calculations that are based on the measured B(0) and B(1) field maps. PMID- 10698654 TI - Spin dynamics of polarization inversion spin exchange at the magic angle in multiple spin systems. AB - Polarization inversion spin exchange at the magic angle (PISEMA) [J. Magn. Reson. A 109, 270 (1994)] is an important experiment in NMR structural characterization of membrane proteins in oriented lipid bilayers. This paper presents a theoretical and experimental study of the spin dynamics in PISEMA to investigate the line-narrowing mechanism. The study focuses on the effect of neighboring protons on the spin exchange of a strongly coupled spin pair. The spin exchange is solved analytically for simple spin systems and is numerically simulated for many-spin systems. The results show that the dipolar couplings from the neighboring protons of a strongly coupled spin pair perturb the spin exchange only in the second order, therefore it has little contribution to the linewidth of PISEMA spectra in comparison to the separated-local-field spectra. The effects from proton resonance offset and the mismatch of the Hartmann-Hahn condition are also discussed along with experimental results using model single-crystal samples. PMID- 10698655 TI - Dielectric resonator-based side-access probe for muscle fiber EPR study. AB - We present a novel dielectric resonator (DR)-based resonant structure that accommodates aqueous sample capillaries in orientations that are either parallel (i.e., side-access) or perpendicular to the direction of an external (Zeeman) magnetic field, B(0). The resonant structure consists of two commercially available X-band DRs that are separated by a Rexolite spacer and resonate in the fundamental TE(01delta) mode. The separator between the DRs is used to tune the resonator to the desired frequency and, by appropriately drilled sample holes, to provide access for longitudinal samples, notably capillaries containing oriented, spin-labeled muscle fibers. In contrast to the topologically similar cylindrical TE(011) cavity, the DR-based structure has distinct microwave properties that favor its use for parallel orientation of lossy aqueous samples. For perpendicular orientation of a dilute (6.25 microM) aqueous solution of IASL spin label, the S/N ratio was at least one order of magnitude better for the side access DR-based structure than for a standard TE(102) cavity. EPR spectra acquired for maleimide spin-labeled myosin filaments also revealed ca. 10 times better S/N ratio than those obtained with a standard TE(102) cavity. For the side access DR with sample capillaries oriented either parallel or perpendicular to the external magnetic field, the Q- and filling factors are in good agreement with the theoretical estimates derived from the distribution of magnetic (H(1)) and electric (E(1)) components. PMID- 10698656 TI - Multiple-rotor-cycle 2D PASS experiments with applications to (207)Pb NMR spectroscopy. AB - Thetwo-dimensional phase-adjusted spinning sidebands (2D PASS) experiment is a useful technique for simplifying magic-angle spinning (MAS) NMR spectra that contain overlapping or complicated spinning sideband manifolds. The pulse sequence separates spinning sidebands by their order in a two-dimensional experiment. The result is an isotropic/anisotropic correlation experiment, in which a sheared projection of the 2D spectrum effectively yields an isotropic spectrum with no sidebands. The original 2D PASS experiment works best at lower MAS speeds (1-5 kHz). At higher spinning speeds (8-12 kHz) the experiment requires higher RF power levels so that the pulses do not overlap. In the case of nuclei such as (207)Pb, a large chemical shift anisotropy often yields too many spinning sidebands to be handled by a reasonable 2D PASS experiment unless higher spinning speeds are used. Performing the experiment at these speeds requires fewer 2D rows and a correspondingly shorter experimental time. Therefore, we have implemented PASS pulse sequences that occupy multiple MAS rotor cycles, thereby avoiding pulse overlap. These multiple-rotor-cycle 2D PASS sequences are intended for use in high-speed MAS situations such as those required by (207)Pb. A version of the multiple-rotor-cycle 2D PASS sequence that uses composite pulses to suppress spectral artifacts is also presented. These sequences are demonstrated on (207)Pb test samples, including lead zirconate, a perovskite-phase compound that is representative of a large class of interesting materials. PMID- 10698657 TI - Burst imaging: rotation artifacts and how to correct them. AB - The effect of coherent rotational motion on images acquired with the ultrafast single-shot spin-echo Burst sequence has been analyzed. Previous experience has demonstrated that sample rotation during Burst experiments has the potential to cause severe image artifacts. In this paper we show that no distortions are visible when the readout gradient is parallel to the rotation axis, but that there is a very distinctive behavior for the case of the rotation axis orthogonal to the imaging plane. The mathematical expression that describes the resulting signal is presented and is used as a basis for a method of correcting the k-space data. The conditions under which undistorted images may be recovered are discussed. It is shown that there is an asymmetry, dependent on the rotation direction, in both the manifestation of the artifact and the range of angular velocities over which one can correct the images. Data from an agar gel phantom rotating at a known rate are used to show how the theory is successful at reconstructing images, with no free parameters. The range of angular velocities over which correction is possible depends on the timing parameters of the pulse sequence, but for these data was -0.016 < omega less, similar 0.1 revolutions/s. Volunteer experiments have confirmed that the theory is applicable to patient motion and can correct motional distortion even when the exact rate is not known a priori. By optimizing the reconstruction to restore a known sample geometry/aspect ratio, an estimate of the rotation angular frequency is obtained with a precision of +/-10%. PMID- 10698658 TI - Development and use of a virtual NMR facility. AB - We have developed a "virtual NMR facility" (VNMRF) to enhance access to the NMR spectrometers in Pacific Northwest National Laboratory's Environmental Molecular Sciences Laboratory (EMSL). We use the term virtual facility to describe a real NMR facility made accessible via the Internet. The VNMRF combines secure remote operation of the EMSL's NMR spectrometers over the Internet with real-time videoconferencing, remotely controlled laboratory cameras, real-time computer display sharing, a Web-based electronic laboratory notebook, and other capabilities. Remote VNMRF users can see and converse with EMSL researchers, directly and securely control the EMSL spectrometers, and collaboratively analyze results. A customized Electronic Laboratory Notebook allows interactive Web-based access to group notes, experimental parameters, proposed molecular structures, and other aspects of a research project. This paper describes our experience developing a VNMRF and details the specific capabilities available through the EMSL VNMRF. We show how the VNMRF has evolved during a test project and present an evaluation of its impact in the EMSL and its potential as a model for other scientific facilities. All Collaboratory software used in the VNMRF is freely available from www.emsl.pnl.gov:2080/docs/collab. PMID- 10698659 TI - Evaluation of cross-correlation effects and measurement of one-bond couplings in proteins with short transverse relaxation times. AB - Various strategies are described and compared for measurement of one-bond J(NH) and J(NC') splittings in larger proteins. In order to evaluate the inherent resolution obtainable in the various experiments, relaxation rates of (15)N (1)H(N) coupled and heteronuclear decoupled resonances were measured at 600- and 800-MHz field strengths for both perdeuterated and protonated proteins. A comparison of decay rates for the two (15)N-?H(N)? doublet components shows average ratios of 4.8 and 3.5 at 800- and 600-MHz (1)H frequency, respectively, in the perdeuterated proteins. For the protonated proteins these ratios are 3.2 (800 MHz) and 2.4 (600 MHz). Relative to the regular HSQC experiment, the enhancement in TROSY (15)N resolution is 2.6 (perdeuterated; 800 MHz), 2.0 (perdeuterated; 600 MHz), 2.1 (protonated; 800 MHz), and 1.7 (protonated; 600 MHz). For the (1)H dimension, the upfield (1)H(N)-?(15)N? component on average relaxes slower than the downfield (1)H(N)-?(15)N? component by a factor of 1.8 (perdeuterated; 800 MHz) and 1.6 (perdeuterated; 600 MHz). The poor resolution for the upfield (15)N-?(1)H? doublet component in slowly tumbling proteins makes it advantageous to derive the J(NH) splitting from the difference in frequency between the narrow downfield (15)N doublet component and either the (1)H decoupled (15)N resonance or the peak position in an experiment which J-scales the frequency of the upfield doublet component but maintains some of the advantages of the TROSY experiment. PMID- 10698660 TI - Omega-space adaptive acquisition technique for magnetic resonance imaging from projections. AB - An omega-space adaptive acquisition technique for MRI from projections is presented. It is based on the evaluation of the information content of a set composed of four initial projections, measured at angles 0 degrees, 45 degrees, 90 degrees, and 135 degrees, followed by the selection of new angles where the information content is maximum. An entropy function is defined on the power spectrum of the projections that is useful for evaluating the information content of each projection. The method makes it possible to reduce the total acquisition time with little degradation of the reconstructed image and it adapts to the arbitrary shape of the sample. For this reason, it can be particularly useful in those applications where acquisition from projections is strongly recommended to save acquisition time, such as functional MRI, imaging of species having very short T(2), or angiography. The method has been tested both on simulated data and on experimental data collected by a commercial MRI apparatus. The method has also been compared to the regular acquisition method, that is, the standard acquisition method in MRI from projections. PMID- 10698661 TI - Volume selective detection by weighted averaging of constant tip angle scans. AB - We propose a method to improve the sensitivity in volume selective detection based on the CARVE excitation sequence (I. Sersa and S. Macura, J. Magn. Reson. 135, 466-477 (1998)) which consists of signal acquisition with constant tip angle excitation and a short phase-encoding gradient pulse. Volume selectivity is achieved using the weighted average of a number of scans whose weights and gradient steps are determined by the shape of the excitation profile. The method is particularly useful for broadband volume selective detection of insensitive spins where the volume selection can be merged with the standard signal averaging process, without compromising the excitation bandwidth or sensitivity. PMID- 10698662 TI - Temperature-dependent chemical shift and relaxation times of (23)Na in Na(4)HTm[DOTP]. AB - We describe the characterization of a (23)Na temperature-dependent chemical shift and relaxation rates in the complex, Na(4)HTm[DOTP]. This is the first characterization of a (23)Na temperature-dependent chemical shift in a nonmetallic sample. The (23)Na temperature-dependent chemical shift coefficient is approximately -0. 5 PPM/ degrees C for both an aqueous solution and a 6% agarose gel of this compound. This is 50 times the magnitude of the temperature dependent chemical shift coefficient of water protons. The relaxation times, T(1), T(2f), and T(2s) increased by 0.1, 0.01, and 0.05 ms/ degrees C, respectively. Applications of these unique properties for designing an MRI technique for monitoring heat deposition in tissue and tissue phantoms are discussed. PMID- 10698663 TI - Pure absorption-mode spectra using a modulated RF mixing period in MQMAS experiments. AB - Different approaches to obtain pure absorption-mode lineshapes in MQMAS experiments employing a train of 180 degrees phase-alternating pulses for the multiple-quantum to single-quantum mixing period are investigated. Four pulse sequences, which achieve this by using either the shifted-echo approach or the hypercomplex approach with symmetric coherence transfer pathways, are presented and their improved lineshape- and sensitivity-performance is experimentally demonstrated by (87)Rb MQMAS of RbNO(3). Compared to the original modulated-rf mixing sequence, sensitivity enhancements by factors up to 1.3 are obtained with the sequences described here. PMID- 10698664 TI - Sensitivity gain by simultaneous acquisition of two coherence pathways: the HNCA(+) experiment. AB - In most multidimensional nuclear magnetic resonance experiments a single and distinct coherence transfer pathway is selected by phase cycling or by pulsed field gradients. It was shown that simultaneously exploiting more than one coherence transfer pathway could increase the overall sensitivity of NMR experiments. However, sensitivity enhancement schemes described to date introduce additional delays in the pulse schemes, resulting in considerable decrease of the expected sensitivity gain when applied to biomolecules due their fast transverse relaxation. A novel sensitivity enhancement principle which increases sensitivity of an experiment by simultaneously exploiting two completely independent coherence pathways in a single NMR pulse scheme is presented in this paper. As an example an improved HNCA experiment, the HNCA(+), is presented, which combines the "out-and-back" coherence transfer pathway used in HNCA with an "out-and-stay" experiment, analogous to HCANH, without adding any time periods compared to the conventional HNCA pulse sequence. The applicability of the HNCA(+) was theoretically evaluated with regard to different sizes of peptides or proteins, which showed that the experimental time can be reduced twofold in ideal cases. The application of this novel experiment to a 7-kDa protein showed a 20% sensitivity gain of HNCA(+) when compared to conventional HNCA. PMID- 10698665 TI - Simple and accurate determination of global tau(R) in proteins using (13)C or (15)N relaxation data. AB - In the study of protein dynamics by (13)C or (15)N relaxation measurements different models from the Lipari-Szabo formalism are used in order to determine the motion parameters. The global rotational correlation time tau(R) of the molecule must be estimated prior to the analysis. In this Communication, the authors propose a new approach in determining an accurate value for tau(R) in order to realize the best fit of R(2) for the whole sequence of the protein, regardless of the different type of motions atoms may experience. The method first determines the highly structured regions of the sequence. For each corresponding site, the Lipari-Szabo parameters are calculated for R(1) and NOE, using an arbitrary value for tau(R). The chi(2) for R(2), summed over the selected sites, shows a clear minimum, as a function of tau(R). This minimum is used to better estimate a proper value for tau(R). PMID- 10698666 TI - 1H dynamic nuclear polarization in supercritical ethylene at 1.4 T. AB - (1)H dynamic nuclear polarization (DNP) has been measured in supercritical ethylene in the pressure range 60-300 bar in an external field of 1.4 T. A single cell sapphire tube was used as a high-pressure cell, and powdered 1,3 bisdiphenylene-2-phenyl allyl (BDPA) free radicals were added and distributed at the wall of the cell. At all pressures the dominant DNP mechanism was a positive Overhauser enhancement, caused by proton-electron contact interactions at the fluid/solid radical interface. The observed enhancements varied from 12 at 60 bar to 17 at 300 bar. Besides the Overhauser enhancement, small solid state and thermal mixing enhancements also were observed, indicating that part of the ethylene was adsorbed at the radical surface for a prolonged time. The impacts of the experimental conditions on the Overhauser enhancement factors are discussed, and enhancements of at least 40-60 are estimated when the EPR saturation factor and the leakage factor become maximal. These data indicate that DNP-enhanced NMR has the potential of extending the impact of NMR in research areas involving supercritical fluids. PMID- 10698667 TI - Acute effects of nicotine infusion on platelets in nicotine users with normal and impaired renal function. AB - The role of platelets in cardiovascular disease associated with smoking is becoming more established, but the effects of nicotine on platelets are unclear. Nicotine therapy is used for smoking cessation in both health and disease. Consequently, the effects of nicotine on platelets are of particular significance in disorders such as renal disease, which is associated with defective platelet function, increased cardiovascular morbidity, and altered nicotine metabolism. Thus, the aim of the present study was to investigate the acute effects of nicotine infusion (NI) on platelets in seven healthy subjects (HS) and seven patients with renal failure (RF). All subjects were nicotine users and had refrained from using nicotine for 36 h before NI. Blood was collected before, immediately after, and 2 h after NI. The plasma concentrations of nicotine and its main metabolite cotinine were determined by gas chromatography. Platelet responsiveness was assessed by aggregometry and flow cytometry in whole blood (P selectin surface expression, fibrinogen- and von Willebrand factor-binding), P selectin expression in isolated platelets, and immunoassays of platelet release (beta-thromboglobulin, platelet factor 4, and soluble P-selectin) and nitric oxide (NO) products. The plasma levels of cotinine, but not nicotine, were significantly higher in RF compared to HS at all time points. In both groups, collagen-induced platelet aggregation was restrained immediately after NI, when the plasma concentration of nicotine was maximal, and was restored after 2 h. Two hours after NI, activation-dependent P-selectin surface expression in isolated platelets increased in both groups. This increased platelet responsiveness occurred simultaneously with a significant increase of plasma cotinine and a decrease of NO products. Thus, the present study suggests that nicotine, directly or through some secondary mechanism or metabolite, only slightly potentiates some of the platelet responses. Renal failure appears not to influence the effects of nicotine on platelets. PMID- 10698668 TI - Interference with growth hormone stimulation of hepatic cytochrome P4502C11 expression in hypophysectomized male rats by 3-methylcholanthrene. AB - Cytochrome P450 2C11 (CYP2C11) is a sexually dimorphic liver enzyme whose expression is regulated by the male pulsatile pattern of growth hormone (GH) secretion. Hepatic CYP2C11 expression is down-regulated by polycyclic aromatic hydrocarbons such as 3-methylcholanthrene (MC). An attractive hypothesis as to the mechanism of CYP2C11 down-regulation by aromatic hydrocarbons is the disruption of normal GH signaling by exposure to these compounds. To evaluate the effects of MC on the ability of GH to stimulate hepatic CYP2C11 expression, our approach was to employ GH replacement in male Fischer 344 rats made GH-deficient by hypophysectomy (hypx). Groups of hypx rats received the following treatments: vehicle; GH alone (twice daily, 125 microg/kg sc, days 1-6); MC alone (20 mg/kg gavage, days 1, 3, and 5); and both GH and MC. Rats were euthanized on day 7. As a positive control response, pronounced induction of hepatic CYP1A1 apoprotein was observed in all MC-treated rats. CYP2C11 expression in hypx rats receiving GH alone was increased at the mRNA, apoprotein, and catalytic activity (testosterone 16alpha-hydroxylation) levels, with mRNA and apoprotein levels approaching that of intact male rats. The inability of GH to fully restore CYP2C11 catalytic activity was attributed to the lowered NADPH-cytochrome P450 reductase apoprotein and catalytic activity observed in all hypx rats. CYP2C11 expression in hypx rats receiving both GH and MC was significantly lower at the mRNA, apoprotein, and catalytic activity levels than that observed in hypx rats treated with GH alone, but significantly higher at the mRNA, apoprotein, and catalytic activity levels than that observed in vehicle-treated hypx rats and in hypx rats treated with MC alone. These data suggest that MC interferes with the ability of GH to stimulate CYP2C11 expression. Thus, disruption of GH signaling by aromatic hydrocarbons may represent a mechanism contributing to the suppression of CYP2C11 gene expression. PMID- 10698669 TI - Effects of polychlorinated dibenzofurans, biphenyls, and their mixture with dibenzo-p-dioxins on ovulation in the gonadotropin-primed immature rat: support for the toxic equivalency concept. AB - Previous studies have shown that 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), 1,2,3,7,8-pentachlorodibenzo-p-dioxin (PeCDD), and 1,2,3,4,7, 8-hexachlorodibenzo p-dioxin (HxCDD), and their equipotent mixture block ovulation, reduce ovarian weight gain and alter preovulatory hormone levels in a similar manner. The objective of the current experiment was to investigate the effect of other structurally related compounds such as chlorinated furans and biphenyls on ovulation and related hormonal endpoints. The gonadotropin-primed immature female rat model was used to study the effect of 2,3,4,7, 8-pentachlorodibenzofuran (PeCDF), 3,3',4,4',5-pentachlorobiphenyl (PeCB), and 2,2',5,5' tetrachlorobiphenyl (TCB) and their mixture with polychlorinated dibenzo-p dioxins (PCDDs) on ovulation. Rats were dosed on Day 23 of age at 0900 h with individual congeners (PeCDF, PeCB, TCB) or a mixture of five compounds, which included TCDD, PeCDD, HxCDD, in addition to PeCDF and PeCB. Equine choronic gonadotropin (eCG; 5 IU) was injected 24 h later to induce follicular development. Blood and ovaries were harvested, and ovarian weights determined at various times after eCG. Serum concentrations of 17beta-estradiol (E(2)), progesterone (P(4)), luteinizing hormone (LH), and follicle-stimulating hormone (FSH) were determined by radioimmunoassay. At 72 h after injection of eCG, the number of ova shed was measured by irrigating the ova from oviducts. The slopes of the dose-responses for inhibition of ovulation generated by the individual PeCDF, PeCB, and/or their mixture with PCDDs were similar. PeCDF, PeCB, and the mixture increased serum concentrations of E(2) at 72 h after eCG injection, the day of expected ovulation; in contrast, serum P(4) and FSH were decreased at that same time point. Only the high doses of TCDD, PeCDF, and PeCB blocked LH and FSH surges at 58 h after eCG. The ovarian histology revealed that the effects of PeCDF, PeCB, and the mixture were very similar to those of PCDDs, consisting of ova in large preovulatory follicles and a lack of or reduced number of corpora lutea. Parallel dose-responses of the individual congeners (PeCDF and PeCB) and their equipotent mixture with PCDDs support the toxic equivalency (TEQ) concept for the blockage of ovulation. Thus, PCDDs, PCDFs, and PeCBs appear to block ovulation by the same or a very similar mechanism of action. PMID- 10698670 TI - Rat testicular Src: normal distribution and involvement in ethylene glycol monomethyl ether-induced apoptosis. AB - Kinase activities were previously proposed to be central to germ cell apoptosis induced by ethylene glycol monomethyl ether (EGME) and its active metabolite methoxyacetic acid (MAA). We evaluated the role of tyrosine kinase pp60(c-src) in control and EGME-treated adult rat testis in vivo, as well as in vitro using cultured adult rat seminiferous tubules treated with MAA. In normal testicular tissue, immunoreactivity of Src was mostly detected in Sertoli cell cytoplasm and reached the maximum level around the lumen at spermiation. Src localization was confirmed by immunostaining of cocultures of Sertoli and germ cells and was further confirmed by electron microscopic observation that immunoreactivity was predominant in Sertoli cell cytoplasm as well as occasionally at the Sertoli/germ cell junctions. A single dose of 200 mg/kg EGME induced an increase of Src immunoexpression in both epithelium and interstitium in rat testis. Eight hours after treatment, an intensive immunostaining of Src began to be observed specifically in the cytoplasm of the dying spermatocytes. The apoptotic changes were replicated by exposure of 5 mM MAA in the adult rat seminiferous tubule culture model. Furthermore, spermatocyte degeneration was significantly prevented by cotreatment with 0.1 microM geldanamycin, 10 microM herbimycin A, or 10 microM PP2, which are inhibitors of Src activity. These data collectively suggest that pp60(c-src) mediates Sertoli-germ cell interaction in physiological events, and may play an important role in EGME/MAA-induced germ cell apoptosis. PMID- 10698671 TI - Preclinical evaluation of the cardiotoxicity of taxane-anthracycline combinations using the model of isolated perfused rat heart. AB - Paclitaxel strongly potentiates the cardiotoxicity of doxorubicin in the clinical setting. In this study, we aimed (1) to determine whether this potentiation could be reproduced in an ex vivo model and, if so, (2) to select drugs and protocols that did not cause this potentiation. The effect of paclitaxel and docetaxel on the cardiotoxicity induced by doxorubicin and epirubicin was studied using the model of isolated perfused rat heart. Cardiac performances were evaluated after several combination protocols administered every 2 days over a period of 12 days, and anthracycline concentrations in the heart and liver were determined on Day 12. When administered simultaneously, paclitaxel strongly potentiated the cardiotoxicity of doxorubicin ex vivo, and this effect was not due to Cremophor EL, the solvent used in the formulation of paclitaxel. The potentiation of anthracycline cardiotoxicity could be avoided by the replacement of doxorubicin by epirubicin, and/or of paclitaxel by docetaxel. Cardiotoxic potentiation was also avoided by the introduction of a 24-h lag time between the repetitive injections of doxorubicin and docetaxel. The concentration of doxorubicin and its cardiotoxic metabolite, doxorubicinol, in the heart and liver was not significantly altered by the taxanes, but that of epirubicin was increased twofold both in the heart and the liver. These results show that the potentiation of doxorubicin-induced cardiotoxicity by paclitaxel can be reproduced with an ex vivo model, and that it is not related to an increase in tissue concentration of the drug or active metabolite. Our model, therefore, may be useful for the selection of anthracycline-containing protocols with no increased risk of cardiotoxicity for the patients. PMID- 10698672 TI - 1,2-Dichlorobenzene-mediated hepatocellular oxidative stress in Fischer-344 and Sprague-Dawley rats. AB - 1,2-Dichlorobenzene (1,2-DCB) is a potent hepatotoxicant in male Fischer 344 (F 344) rats but not in Sprague-Dawley (SD) rats. While Kupffer cell-dependent oxidative stress plays a role in the progression of 1,2-DCB-mediated liver injury, we hypothesize that initiation of liver injury is due to oxidative events within the hepatocyte. This study compared hepatocellular oxidative stress marked by glutathione disulfide (GSSG) and glutathione (GSH) production in either bile, liver, or isolated hepatocytes of F-344 and SD rats following 1,2-DCB administration. Hepatic GSH concentrations were depleted at a greater rate in F 344 than in SD rats within 12 h of 1,2-DCB administration (3.6 mmol/kg ip). In bile, GSSG concentrations were threefold greater in F-344 rats compared to SD rats by 9 h of 1,2-DCB treatment. Moreover, 1-aminobenzotriazole but not gadolinium chloride pretreatment blocked the rise in biliary GSSG concentrations following 1,2-DCB administration. In in vitro studies, isolated hepatocytes of F 344 rats had a 15% increase in cellular GSSG concentrations following 1 h of 1,2 DCB (3.55 nmol) exposure, while GSH decreased 22% by 6.5 h compared to controls. In contrast, isolated SD hepatocytes exposed to 1,2-DCB had no increase in GSSG and only an 8% reduction in GSH. Furthermore, parameters of lipid peroxidation were increased in F-344 rats and not in SD rats. Collectively, these data suggest that hepatocellular oxidative stress is dependent upon bioactivation and the enhanced oxidative stress in the F-344 rat may explain its susceptibility to 1,2 DCB compared to the SD rat. PMID- 10698673 TI - Comparative developmental toxicities of aliphatic nitriles: in vivo and in vitro observations. AB - The effects on embryonic development of a series of eight saturated (acetonitrile, propionitrile, and n-butyronitrile) and unsaturated (acrylonitrile, methacrylonitrile, allylnitrile, cis-2-pentenenitrile, and 2 chloroacrylonitrile) nitriles were compared in vitro using the whole embryo culture system. Day 10 rat embryos were cultured for 46 h in rat serum in the presence of either of these chemicals. All the tested chemicals produced concentration-dependent decreases in growth and differentiation and increases in the incidences of morphologically abnormal embryos. A wide range of embryotoxic potency was observed, with 2-chloroacrylonitrile and acetonitrile at the extremes (lowest effect levels of 50 microM and 40 mM, respectively). No common pattern could be drawn for all the eight nitriles tested in vitro, although there were some similarities between the malformations elicited by propionitrile and n butyronitrile or between those elicited by the five unsaturated nitriles. Presence of a rat hepatic microsomal fraction and NADPH in the culture medium enhanced the embryotoxic effects of the five unsaturated nitriles tested but had no effects on saturated nitriles embryotoxicity. In addition to these in vitro experiments, pregnant rats were given a single oral dose of each compound on Day 10 of gestation and the embryos were evaluated on Day 12 of gestation, i.e., at a time of development corresponding to the developmental stage at the end of the whole embryo culture. All the nitriles investigated produced the characteristic defects developed by embryos exposed to sodium cyanide in utero or in culture. Our results provide further evidence that maternal production of cyanide may contribute to the developmental toxicity of saturated and unsaturated nitriles and suggest that distinct metabolites derived from microsomal metabolism of unsaturated nitriles may also play a role. PMID- 10698674 TI - Pharmacokinetic data support pharmacologically induced embryonic dysrhythmia as explanation to Fetal Hydantoin Syndrome in rats. AB - New studies suggest that the teratogenicity of phenytoin (PHT) is linked to its membrane-stabilizing pharmacological action via the rapid component of the delayed rectified potassium channel (lkr), resulting in embryonic cardiac dysrhythmia during a restricted sensitive period. In order to further elucidate this theory, PHT was administered to Sprague-Dawley rats on gestation day (GD) 11 with either a single dose of 150 or 100 mg/kg ip or 150 mg/kg po and developmental toxicity at term (GD 21) was studied. In satellite animals blood samples were withdrawn (0.5-24 h after dose) and total and free maternal plasma concentrations of PHT were measured. Pharmacokinetic data correlated well with pregnancy outcome data. At 150 mg/kg ip high concentrations of long duration (C(max) 240 microM and AUC 5300 microMhl(-1) - total) and marked developmental toxicity (embryonic death, decreased fetal weights, and orofacial clefts) were observed. After 100 mg/kg ip (C(max) 150 microM, AUC 2600 microMhl(-1) - total) only slight developmental toxicity (decreased fetal weights) was recorded and after 150 mg/kg po the plasma concentrations were even lower (C(max) 63 microM and AUC 1100 microMhl(-1) - total) and no adverse effects at all were observed. In separate experiments the effect of different concentrations of PHT on the embryonic heart was studied by adding PHT to GD 11 rat embryos cultured in vitro or by culturing GD 11 embryos from exposed dams. The decrease in heart rates was 3, 16, and 32% after culture with 50, 100, and 200 microM of PHT, respectively. After maternal administration of 150 mg/kg ip or po, the embryonic heart rate in vitro decreased by 25 and 7%, respectively, compared to controls. Altogether the results suggest that the development toxicity of PHT is caused by concentration dependent induction of embryonic dysrhythmia and hypoxia related damage. PMID- 10698675 TI - In vitro covalent binding of nafenopin-CoA to human liver proteins. AB - Endogenous fatty acyl-CoAs play an important role in the acylation of proteins. A number of xenobiotic carboxylic acids are able to mimic fatty acids, forming CoA conjugates and acting as substrates in pathways of lipid metabolism. In this study nafenopin, a substrate for human hepatic fatty acid-CoA ligases, was chosen as a model compound to study xenobiotic acylation of human liver proteins. (3)H nafenopin (+/- unlabeled palmitate) or (14)C-palmitate (+/- unlabeled nafenopin) were incubated for up to 120 min at 37 degrees C with ATP, CoA, and homogenate protein (1 mg/ml) from four individual human livers. Nafenopin covalently bound to proteins was detectable in all human livers and increased with time. Nafenopin adduct formation was directly proportional to nafenopin-CoA formation (r = 0.985, p < 0.05). Attachment of nafenopin to proteins involved both thioester and amide linkages with 76 and 24% of adducts formed with proteins > 100 and 50-100 kDa, respectively. Protein acylation by palmitate was also demonstrated. Palmitate significantly inhibited nafenopin-CoA formation by 29% but had no effect on nafenopin-CoA-mediated protein acylation. In contrast, nafenopin significantly inhibited protein palmitoylation by palmitoyl-CoA. This is the first study to demonstrate a direct relationship between xenobiotic-CoA formation, acylation of human liver proteins, and inhibition of endogenous palmitoylation. The ability of xenobiotics to acylate tissue proteins may have important biological consequences including perturbation of endogenous regulation of protein localization and function. PMID- 10698676 TI - Cotinine and nicotine inhibit each other's calcium responses in bovine chromaffin cells. AB - Cotinine is the major metabolite of nicotine. It has some biological activity, but its pathophysiological effects are largely unclear. We studied whether cotinine initiates calcium transients or affects those induced by nicotine. In bovine adrenal chromaffin cells labeled with the fluorescent calcium indicator Fura 2, cotinine (0. 32-3.2 mM) concentration-dependently increased the intracellular Ca(2+) concentration ([Ca(2+)](i)). The effect was abolished by omitting extracellular Ca(2+) during the stimulations. Also nicotinic receptor channel blockers hexamethonium (10 microM-1 mM) and chlorisondamine (100 microM), as well as a competitive nicotinic receptor antagonist dihydro-beta-erythroidine (10-100 microM), inhibited the response. Cotinine (0.32-3.2 mM) preincubation for 2 min inhibited both the nicotine-induced and the cotinine-induced increases in [Ca(2+)](i). Also nicotine (3.2-10 microM) inhibited the cotinine-induced increase in [Ca(2+)](i). Tetrodotoxin (1 microM) and thapsigargin (1 microM) pretreatments did not affect the responses to cotinine, while 300 nM nimodipine partially inhibited the cotinine-induced increase in [Ca(2+)](i). The results indicate that cotinine has nicotine-like effects on chromaffin cells. It may also desensitize the nicotinic cholinergic receptors, possibly by acting as a low affinity agonist at these receptors. PMID- 10698677 TI - Regional modification of [(3)H]Ethynylbicycloorthobenzoate binding in mouse brain GABA(A) receptor by endosulfan, fipronil, and avermectin B(1a). AB - [(3)H]Ethynylbicycloorthobenzoate ([(3)H]EBOB), a high affinity radioligand for the noncompetitive blocker site of the GABA(A) receptor, is used here for quantitative autoradiography to determine regional binding in mouse brain and the effects on this binding of administering toxic doses of endosulfan, fipronil, and avermectin B(1a) (AVM). Animals were euthanized 4-8 min after 1 LD50 or 2 LD50 doses of the two channel blockers and 32 min after 1 LD50 or 4 LD50 doses of the channel activator AVM. Specific binding of [(3)H]EBOB was determined for 20 microm brain sections as the difference in labeling on incubation with 2 nM [(3)H]EBOB either alone (total binding) or with 5 microM alpha-endosulfan (nonspecific binding). The highest specific labeling was observed for layers I and IV of the cerebral cortex, the globus pallidus, and the medial septal nucleus/nucleus of the vertical limb of the diagonal band. Dose-dependent inhibition by endosulfan was highest in the nucleus accumbens and least in the cerebellum and periaqueductal gray matter. Fipronil had much less effect on binding even at severely toxic doses. AVM increased [(3)H]EBOB binding in most regions and was the only one of the three agents inhibiting in vitro [(3)H]strychnine binding to the glycine receptor. In summary, the noncompetitive blocker site was strongly inhibited with dose dependence and regional selectively by alpha-endosulfan but was generally poorly inhibited or activated by fipronil and was activated by avermectin. PMID- 10698678 TI - Catabolites of cholesterol synthesis pathways and forskolin as activators of the farnesoid X-activated nuclear receptor. AB - The nuclear receptors are a family of transcriptional mediators that, upon activation, bind DNA and regulate gene transcription. Among these receptors, the farnesoid X-activated receptor (FXR) has recently been identified as one activated by bile acids and farnesol. To investigate the potential of other sterols to activate FXR, as well as to examine relevant relationships among identified activators of FXR, the current study used a mammalian cell transcription assay to quantify and compare activation potential. In addition to the classical bile acids deoxycholate (DCA) and chenodeoxycholate (CDCA), FXR was shown to be transcriptionally active in the presence of the androgen catabolites 5alpha-androstan-3alpha-ol-17-one (androsterone) and 5beta-androstan-3alpha-ol-17 one (etiocholanolone), as well as the sterol bronchodilatory drug forskolin. Conversely, cholesterol and several other key precursors to the androgens and bile acids were either not active or only slightly active. Furthermore, it was observed that the bile acid ursodeoxycholate (UDCA) could inhibit DCA and CDCA activation of FXR in a manner parallel to its ability to antagonize DCA and CDCA induction of apoptosis. By far, the most efficacious activator of FXR was forskolin. Interestingly, although it is classically viewed as an initiator of the adenylate cyclase/protein kinase A (PKA) pathway, PKA inhibition did not inhibit forskolin's activation of FXR nor was cyclic AMP (cAMP) able to stimulate FXR-mediated transcription. These data would suggest that forskolin acts as a ligand for FXR rather than as a secondary activator of FXR and could have important implications with respect to its potential toxicity and pharmacological use. PMID- 10698679 TI - Monomethylarsonous acid (MMA(III)) is more toxic than arsenite in Chang human hepatocytes. AB - Methylation has been considered to be the primary detoxication pathway of inorganic arsenic. Inorganic arsenic is methylated by many, but not all animal species, to monomethylarsonic acid (MMA(V)), monomethylarsonous acid (MMA(III)), and dimethylarsinic acid (DMA(V)). The As(V) derivatives have been assumed to produce low toxicity, but the relative toxicity of MMA(III) remains unknown. In vitro toxicities of arsenate, arsenite, MMA(V), MMA(III), and DMA(V) were determined in Chang human hepatocytes. Leakage of lactate dehydrogenase (LDH) and intracellular potassium (K(+)) and mitochondrial metabolism of the tetrazolium salt XTT were used to assess cytotoxicity due to arsenic exposure. The mean LC50 based on LDH assays in phosphate media was 6 microM for MMA(III) and 68 microM for arsenite. Using the assay for K(+) leakage in phosphate media, the mean LC50 was 6.3 microM for MMA(III) and 19.8 microM for arsenite. The mean LC50 based on the XTT assay in phosphate media was 13.6 microM for MMA(III) and 164 microM for arsenite. The results of the three cytotoxicity assays (LDH, K(+), and XTT) reveal the following order of toxicity in Chang human hepatocytes: MMA(III) > arsenite > arsenate > MMA(V) = DMA(V). Data demonstrate that MMA(III), an intermediate in inorganic arsenic methylation, is highly toxic and again raises the question as to whether methylation of inorganic arsenic is a detoxication process. PMID- 10698681 TI - Alpha-synuclein overexpression promotes aggregation of mutant huntingtin. AB - Protein aggregates are a neuropathological feature of Huntington's disease and Parkinson's disease. Mutant huntingtin exon 1 with 72 CAG repeats fused to enhanced green fluorescent protein (EGFP) forms hyperfluorescent inclusions in PC12 cells. Inclusion formation is enhanced in cells co-transfected with EGFP huntingtin-(CAG)(72) and alpha-synuclein, a major component of Parkinson's disease aggregates. However, alpha-synuclein does not form aggregates by itself, nor does it appear in huntingtin inclusions in vitro. PMID- 10698682 TI - The thermostabilizing domain, XynA, of Caldibacillus cellulovorans xylanase is a xylan binding domain. AB - We show that the N-terminal 'thermostabilizing domain' (TSD) of the xylanase, XynA, from the thermophilic bacterium Caldibacillus cellulovorans also acts as a xylan binding domain. Affinity electrophoresis experiments show that this TSD selectively binds soluble xylan and binds weakly to hydroxyethylcellulose. Based on this, and previously reported evidence, we propose that xylanase-associated TSDs are xylan binding domains. PMID- 10698683 TI - Nuclear targeting of the beta isoform of type II phosphatidylinositol phosphate kinase (phosphatidylinositol 5-phosphate 4-kinase) by its alpha-helix 7. AB - Type II phosphatidylinositol phosphate kinases (PIPkins) have recently been found to be primarily phosphatidylinositol 5-phosphate 4-kinases, and their physiological role remains unclear. We have previously shown that a Type II PIPkin [isoform(s) unknown], is localized partly in the nucleus [Divecha, Rhee, Letcher and Irvine (1993) Biochem. J. 289, 617-620], and here we show, by transfection of HeLa cells with green-fluorescent-protein-tagged Type II PIPkins, that this is likely to be the Type IIbeta isoform. Type IIbeta PIPkin has no obvious nuclear localization sequence, and a detailed analysis of the localization of chimaeras and mutants of the alpha (cytosolic) and beta PIPkins shows that the nuclear localization requires the presence of a 17-amino-acid length of alpha-helix (alpha-helix 7) that is specific to the beta isoform, and that this helix must be present in its entirety, with a precise orientation. This resembles the nuclear targeting of the HIV protein Vpr, and Type IIbeta PIPkin is apparently therefore the first example of a eukaryotic protein that uses the same mechanism. PMID- 10698680 TI - The PI3K-PDK1 connection: more than just a road to PKB. AB - Phosphoinositide 3-kinases (PI3Ks) generate specific inositol lipids that have been implicated in the regulation of cell growth, proliferation, survival, differentiation and cytoskeletal changes. One of the best characterized targets of PI3K lipid products is the protein kinase Akt or protein kinase B (PKB). In quiescent cells, PKB resides in the cytosol in a low-activity conformation. Upon cellular stimulation, PKB is activated through recruitment to cellular membranes by PI3K lipid products and phosphorylation by 3'-phosphoinositide-dependent kinase-1 (PDK1). Here we review the mechanism by which PKB is activated and the downstream actions of this multifunctional kinase. We also discuss the evidence that PDK1 may be involved in the activation of protein kinases other than PKB, the mechanisms by which this activity of PDK1 could be regulated and the possibility that some of the currently postulated PKB substrates targets might in fact be phosphorylated by PDK1-regulated kinases other than PKB. PMID- 10698684 TI - Rabaptin4, a novel effector of the small GTPase rab4a, is recruited to perinuclear recycling vesicles. AB - The small GTPase rab4a is associated with early endocytic compartments and regulates receptor recycling from early endosomes. To understand how rab4a mediates its function, we searched for proteins which associate with this GTPase and regulate its activity in endocytic transport. Here we identified rabaptin4, a novel effector molecule of rab4a. Rabaptin4 is homologous with rabaptin5 and contains a C-terminal deletion with respect to rabaptin5. Rabaptin4 preferentially interacts with rab4a-GTP and to a lesser extent with rab5aGTP. We identified a rab4a-binding domain in the N-terminal region of rabaptin4, and two binding sites for rab5, including a novel N-terminal rab5a-binding site. Rabaptin4 is a cytosolic protein that inhibits the intrinsic GTP hydrolysis rate of rab4a and is recruited by rab4a-GTP to recycling endosomes enriched in cellubrevin and internalized indocarbocyanine-3 (Cy3)-labelled transferrin. We propose that rabaptin4 assists in the docking of transport vesicles en route from early endosomes to recycling endosomes. PMID- 10698685 TI - Hepatic expression, synthesis and secretion of a novel fibrinogen/angiopoietin related protein that prevents endothelial-cell apoptosis. AB - Using degenerate PCR we isolated a cDNA encoding a novel 406- and 410-amino acid protein from human and mouse embryonic cDNAs and have designated it 'hepatic fibrinogen/angiopoietin-related protein' (HFARP). The N-terminal and C-terminal portions of HFARP contain the characteristic coiled-coil domains and fibrinogen like domains that are conserved in angiopoietins. In human and mouse tissues, HFARP mRNA is specifically expressed in the liver. HFARP mRNA and protein are mainly present in the hepatocytes. HFARP has a highly hydrophobic region at the N terminus that is typical of a secretory signal sequence and one consensus glycosylation site. Recombinant HFARP expressed in COS-7 cells is secreted and glycosylated. HFARP protein is present not only in the hepatocytes, but also in the circulating blood. Recombinant HFARP acts as an apoptosis survival factor for vascular endothelial cells, but does not bind to Tie1 or Tie2 (endothelial-cell tyrosine kinase receptors). These results suggest that HFARP may exert a protective function on endothelial cells through an endocrine action. PMID- 10698686 TI - Organization and chromosomal localization of the human ECEL1 (XCE) gene encoding a zinc metallopeptidase involved in the nervous control of respiration. AB - ECEL1 (endothelin-converting enzyme-like 1; previously known as XCE) is a putative zinc metalloprotease that was identified recently on the basis of its strong identity with endothelin-converting enzyme. Although the physiological function of ECEL1 is unknown, inactivation of the corresponding gene in mice points to a critical role of this protein in the nervous control of respiration. In the present study we have characterized the human ECEL1 gene. It was located to region q36-q37 of chromosome 2 and shown to be composed of 18 exons spanning approx. 8 kb. The structure of the ECEL1 gene displays some striking similarities with those of genes of related metallopeptidases, supporting the hypothesis that they are all derived from a common ancestor. A short phylogenetic study describing the relationship between the various members of this gene family is also presented. PMID- 10698687 TI - Molecular decipherment of Rho effector pathways regulating tight-junction permeability. AB - We reported recently that the activation of RhoA induced an increase in transepithelial electrical resistance (TER). To clarify effectors of Rho for this RhoA-induced regulation of tight-junction permeability, we introduced two effector-loop mutants of constitutively active RhoA(V14), RhoA(V14/L40) and RhoA(V14/C42), into Mardin-Darby canine kidney cells in an isopropyl beta-D thiogalactoside-inducible expression system. RhoA(V14) and the two effector-loop mutants interacted in vitro with the Rho-binding domain of Rho-associated kinase, ROKalpha. Next we examined two parameters of Rho functions, stress-fibre formation and TER elevation, induced by RhoA(V14). Stress-fibre formation was induced by RhoA(V14/C42) but not by RhoA(V14/L40). On the other hand, TER elevation was induced by neither RhoA(V14/L40) nor RhoA(V14/C42). RhoA-associated kinase inhibitor, Y-27632, inhibited both stress-fibre formation and TER elevation induced by RhoA(V14). These results demonstrated that RhoA-induced regulation of tight-junction permeability is mediated by Rho-associated kinase and at least one other unidentified effector, the coupling to RhoA being disrupted by mutation at position 40 or 42 in the effector loop. PMID- 10698688 TI - Histidine to aspartate phosphotransferase activity of nm23 proteins: phosphorylation of aldolase C on Asp-319. AB - nm23 genes have been implicated in the suppression of tumour metastasis and cell motility; however, the biochemical mechanisms for these suppressions are not known. We have previously described the transfer of phosphate from the catalytic histidine residues of nm23 proteins to an aspartic or a glutamic residue on one or more 43 kDa proteins in detergent extracts of bovine brain membranes. To gain a better understanding of this transferase activity, we partly purified this 43 kDa protein and identified aldolases A and C as the major 43 kDa proteins present in the preparation. Aldolase was purified from brain cytosol; its phosphorylation by rat liver nm23 proteins and by recombinant human nm23-H1 was examined. The site of phosphorylation was identified as Asp-319 on aldolase C. The equivalent residue on aldolase A, a glutamic residue, was not phosphorylated. Aldolase C was rapidly phosphorylated by wild-type nm23-H1 but was not phosphorylated, or was phosphorylated very slowly, by either nm23-H1(P96S) or nm23-H1(S120G), mutants of nm23-H1 that do not suppress cell motility. This is the first identification of a protein that is phosphorylated on an aspartic residue by nm23 proteins. The sequence around Asp-319 of aldolase C has some similarities to those around the histidine residues on ATP-citrate lyase and succinic thiokinase that are phosphorylated by nm23 proteins. PMID- 10698689 TI - Two phospholipase A2 inhibitors from the plasma of Cerrophidion (Bothrops) godmani which selectively inhibit two different group-II phospholipase A2 myotoxins from its own venom: isolation, molecular cloning and biological properties. AB - Myotoxic phospholipases A(2) (PLA(2)s; group II) account for most of the muscle tissue damage that results from envenomation by viperid snakes. In the venom of the Godman's viper (Cerrophidion godmani, formerly Bothrops godmani), an enzymically active PLA(2) (myotoxin I) and an inactive, Lys-49 variant (myotoxin II) induce extensive muscle damage and oedema. In this study, two distinct myotoxin inhibitor proteins of C. godmani, CgMIP-I and CgMIP-II, were purified directly from blood plasma by selective binding to affinity columns containing either myotoxin I or myotoxin II, respectively. Both proteins are glycosylated, acidic (pI=4) and composed of 20-25-kDa subunits that form oligomers of 110 kDa (CgMIP-I) or 180 kDa (CgMIP-II). In inhibition studies, CgMIP-I specifically neutralized the PLA(2) and the myotoxic, oedema-forming and cytolytic activities of myotoxins I, whereas CgMIP-II selectively inhibited the toxic properties of myotoxin II. N-terminal amino acid sequence analysis and sequencing of cDNAs encoding the two inhibitors revealed that CgMIP-I is similar to gamma-type inhibitors, which share a pattern of cysteine residues present in the Ly-6 superfamily of proteins, whereas CgMIP-II shares sequence identity with alpha type inhibitors that contain carbohydrate-recognition-like domains, also found in C-type lectins and mammalian PLA(2) receptors. N-terminal sequencing of myotoxin I revealed a different primary structure from myotoxin II [De Sousa, Morhy, Arni, Ward, Diaz and Gutierrez (1998) Biochim. Biophys. Acta 1384, 204-208], which provides insight into the nature of such pharmacological specificity. PMID- 10698690 TI - Evidence for regulation of NF-kappaB by poly(ADP-ribose) polymerase. AB - The DNA-binding activity of NF-kappaB in nuclear extracts of poly(ADP-ribose) polymerase (PARP)-defective mutant L1210 cell clones was markedly increased and was inversely correlated with the PARP content in these cells. The DNA-binding activity of NF-kappaB in a clone with the lowest PARP content (Cl-3527, contained 6% of PARP of wild type cells) was about 35-fold of that of the wild-type cells, whereas the change in the DNA-binding activity of AP-1 and SP-1 in the mutant was relatively small or not so significant. Transfection of a PARP-expressing plasmid to the mutant cells decreased the abnormally high levels of NF-kappaB complexes, especially p50/p65(Rel A) complex, to near the normal level. Moreover, poly(ADP ribosyl)ation of nuclear extracts in vitro suppressed the ability of NF-kappaB to form a complex with its specific DNA probe by approx. 80%. Further analysis with purified recombinant NF-kappaB proteins revealed that both rp50 and rMBP-p65 (Rel A) proteins, but not rGST-IkappaB, could be poly(ADP-ribosyl)ated in vitro and that the modification resulted in a marked decrease in the DNA-binding activity of rMBP-p65, whereas a slight activation was observed in rp50. Poly(ADP ribosyl)ated p65/NF-kappaB was detected in the cytosol of wild type L1210 cells by immunoblotting with anti-poly(ADP-ribose) and anti-p65 antibodies. Taken together, these results strongly suggest that PARP is involved in the regulation of NF-kappaB through the protein modification. PMID- 10698691 TI - Fibre-type specific modification of the activity and regulation of skeletal muscle pyruvate dehydrogenase kinase (PDK) by prolonged starvation and refeeding is associated with targeted regulation of PDK isoenzyme 4 expression. AB - Using immunoblot analysis with antibodies raised against recombinant pyruvate dehydrogenase kinase (PDK) isoenzymes PDK2 and PDK4, we demonstrate selective changes in PDK isoenzyme expression in slow-twitch versus fast-twitch skeletal muscle types in response to prolonged (48 h) starvation and refeeding after starvation. Starvation increased PDK activity in both slow-twitch (soleus) and fast-twitch (anterior tibialis) skeletal muscle and was associated with loss of sensitivity of PDK to inhibition by pyruvate, with a greater effect in anterior tibialis. Starvation significantly increased PDK4 protein expression in both soleus and anterior tibialis, with a greater response in anterior tibialis. Starvation did not effect PDK2 protein expression in soleus, but modestly increased PDK2 expression in anterior tibialis. Refeeding for 4 h partially reversed the effect of 48-h starvation on PDK activity and PDK4 expression in both soleus and anterior tibialis, but the response was more marked in soleus than in anterior tibialis. Pyruvate sensitivity of PDK activity was also partially restored by refeeding, again with the greater response in soleus. It is concluded that targeted regulation of PDK4 isoenzyme expression in skeletal muscle in response to starvation and refeeding underlies the modulation of the regulatory characteristics of PDK in vivo. We propose that switching from a pyruvate-sensitive to a pyruvate-insensitive PDK isoenzyme in starvation (a) maintains a sufficiently high pyruvate concentration to ensure that the glucose- >alanine-->glucose cycle is not impaired, and (b) may 'spare' pyruvate for anaplerotic entry into the tricarboxylic acid cycle to support the entry of acetyl-CoA derived from fatty acid (FA) oxidation into the tricarboxylic acid cycle. We further speculate that FA oxidation by skeletal muscle is both forced and facilitated by upregulation of PDK4, which is perceived as an essential component of the operation of the glucose-FA cycle in starvation. PMID- 10698692 TI - Characterization of AMP-activated protein kinase gamma-subunit isoforms and their role in AMP binding. AB - The AMP-activated protein kinase (AMPK) cascade plays an important role in the regulation of energy homeostasis within the cell. AMPK is a heterotrimer composed of a catalytic subunit (alpha) and two regulatory subunits (beta and gamma). We have isolated and characterized two isoforms of the gamma subunit, termed gamma2 and gamma3. Both gamma2 (569 amino acids) and gamma3 (492 amino acids) have a long N-terminal domain which is not present in the previously characterized isoform, gamma1. As with gamma1, mRNA encoding gamma2 is widely expressed in human tissues, whereas significant expression of gamma3 mRNA was only detected in skeletal muscle. Using isoform-specific antibodies, we determined the AMPK activity associated with the different gamma isoforms in a number of rat tissues. In most tissues examined more than 80% of total AMPK activity was associated with the gamma1 isoform, with the remaining activity being accounted for mainly by the gamma2 isoform. Exceptions to this were testis and, more notably, brain where all three isoforms contributed approximately equally to activity. There was no evidence for any selective association between the alpha1 and alpha2isoforms and the various gamma isoforms. However, the AMP-dependence of the kinase complex is markedly affected by the identity of the gamma isoform present, with gamma2 containing complexes having the greatest AMP-dependence, gamma3 the lowest, and gamma1 having an intermediate effect. Labelling studies, using the reactive AMP analogue 8-azido-[(32)P]AMP, indicate that the gamma subunit may participate directly in the binding of AMP within the complex. PMID- 10698693 TI - Neutral sphingomyelinase activity dependent on Mg2+ and anionic phospholipids in the intraerythrocytic malaria parasite Plasmodium falciparum. AB - Sphingolipid metabolism and metabolites are important in various cellular events in eukaryotes. However, little is known about their function in plasmodial parasites. Here we demonstrate that neutral sphingomyelinase (SMase) involved in the sphingomyelin (SM) catabolism is retained by the intraerythrocytic parasite Plasmodium falciparum. When assayed in a neutral pH buffer supplemented with Mg(2+) and phosphatidylserine, an activity for the release of the phosphocholine group from SM was detected in parasite-infected, but not in uninfected, erythrocyte ghosts. The SMase activity in the parasite-infected erythrocyte ghosts was enhanced markedly by anionic phospholipids including unsaturated but not saturated phosphatidylserine. Mn(2+) could not substitute for Mg(2+) to activate SMase in parasite-infected erythrocyte ghosts, whereas both Mn(2+) and Mg(2+) activated mammalian neutral SMase. The specific activity level of SMase was higher in isolated parasites than in infected erythrocyte ghosts; further fractionation of lysates of the isolated parasites showed that the activity was bound largely to the membrane fraction of the parasites. The plasmodial SMase seemed not to hydrolyse phosphatidylcholine or phosphatidylinositol. The plasmodial SMase, but not SM synthase, was sensitive to scyphostatin, an inhibitor of mammalian neutral SMase, indicating that the plasmodial activities for SM hydrolysis and SM synthesis are mediated by different catalysts. Our finding that the malaria parasites possess SMase activity might explain why the parasites seem to have an SM synthase activity but no activity to synthesize ceramide de novo. PMID- 10698694 TI - Mapping the catalytic pocket of phospholipases A2 and C using a novel set of phosphatidylcholines. AB - A set of radioiodinatable phosphatidylcholines (PCs) derivatized with the Bolton Hunter reagent (BHPCs) was synthesized to probe the substrate recognition and activity of phospholipases. A common feature of this series is the presence of a bulky 4-hydroxyphenyl group at the end of the fatty acyl chain attached to position sn-2. The distance between the end group and the glycerol backbone was varied by changing the length of the intervening fatty acyl chain (3-25 atoms). Except for the shortest, this chain includes at least one amide linkage. The usefulness of this series of substrates as a molecular ruler was tested by measuring the hydrolytic activities of Naja naja naja phospholipase A(2) (PLA(2)) and Bacillus cereus phospholipase C (PLC) in Triton X-100 micelles. The activity of PLA(2) proved to be highly dependent on the length of the fatty acyl chain linker, the shorter compounds (3-10 atoms) being very poor substrates. In contrast, the PLC activity profile exhibited much less discrimination. In both cases, PCs with 16-21 atom chains at position sn-2 yielded optimal activity. We interpret these findings in terms of fatty acyl chain length-related steric hindrance caused by the terminal aromatic group, affecting the activity of PLA(2) and, to a smaller extent, that of PLC. This notion agrees with the more extended recognition of aliphatic chains inside the narrow channel leading to the catalytic site in the former case. Molecular models of these substrates bound to PLA(2) were built on the basis of the crystallographic structure of Naja naja atra PLA(2) complexed with a phospholipid analogue. Docking of these substrates necessarily requires the intrusion of the bulky 4-hydroxyphenyl group inside the binding pocket and also the failure of the amide group to form hydrogen bonds inside the hydrophobic substrate channel. PMID- 10698695 TI - Mechanism of an antibody-catalysed allylic isomerization. AB - The catalytic antibody 4B2, which was generated against a substituted amidine 1, catalyses the allylic isomerization of beta, gamma-unsaturated ketones with an acceleration factor (k(cat)/k(uncat)) of 1.5x10(3). On the basis of the 'bait and switch' strategy, it was reasoned that the positively charged hapten could elicit, by charge complementarity, an acidic residue (Asp or Glu) in the antibody binding site in the right position to catalyse this proton transfer reaction. The pH dependence curve of k(cat)/K(m) shows a bell-shaped feature with an optimum at approx. pH 4.5. By cloning and sequencing the light and heavy chains of the 4B2 antibody, we confirmed the presence of several Asp and Glu residues in the complementarity-determining region loops. The antibody catalyses the alpha-proton exchange on the same substrates, demonstrating the involvement of a dienol intermediate in the reaction mechanism. Kinetic studies with (2)H-NMR provide evidence that alpha-proton abstraction is stereospecific. Whether the process involves one or two acid/base residues in this simple proton transfer or whether it is a concerted mechanism is discussed. PMID- 10698696 TI - Antizyme inhibitor is rapidly induced in growth-stimulated mouse fibroblasts and releases ornithine decarboxylase from antizyme suppression. AB - Ornithine decarboxylase (ODC) catalyses the first step in the synthesis of the polyamines putrescine, spermidine and spermine. The polyamines are essential for cell growth, but at elevated levels they may be tumorigenic, toxic, or may induce apoptosis. Therefore, ODC activity is highly regulated. It is induced when cells are stimulated to grow, and it is subjected to feedback inhibition by the polyamines. By causing ribosomal frameshifting, polyamines induce the synthesis of antizyme, a 23-kDa protein, which binds to ODC, inhibits its activity and promotes its degradation by the 26 S proteasome. Antizyme, in turn, is inhibited by antizyme inhibitor (AZI). We describe the cloning of a mouse AZI cDNA, encoding a protein with high homology to mouse ODC. Using purified recombinant proteins, we show that AZI (which has no ODC activity) can release enzymically active ODC from antizyme suppression in vitro. We also show that ODC reactivation takes place in mouse fibroblasts upon transient transfection with an AZI expressing plasmid construct. Finally we demonstrate that the AZI mRNA content of mouse fibroblasts increases significantly within an hour of growth stimulation, i.e. much earlier than ODC transcripts. Our results indicate that induction of AZI synthesis may represent a means of rescuing ODC molecules that have been inactivated and tagged for degradation by antizyme, when culture conditions improve and polyamine production is needed for cell growth and proliferation. PMID- 10698697 TI - Neutral amino acid transporter ASCT2 displays substrate-induced Na+ exchange and a substrate-gated anion conductance. AB - The neutral amino acid transporter ASCT2 mediates electroneutral obligatory antiport but at the same time requires Na(+) for its function. To elucidate the mechanism, ASCT2 was expressed in Xenopus laevis oocytes and transport was analysed by flux studies and two-electrode voltage clamp recordings. Flux studies with (22)NaCl indicated that the uptake of one molecule of glutamine or alanine is accompanied by the uptake of four to seven Na(+) ions. Similarly to the transport of amino acids, the Na(+) uptake was mediated by an obligatory Na(+) exchange mechanism that depended on the presence of amino acids but was not stoichiometrically coupled to the amino acid transport. Other cations could not replace Na(+) in this transport mechanism. When NaCl was replaced by NaSCN in the transport buffer, the superfusion of oocytes with amino acid substrates resulted in large inward currents, indicating the presence of a substrate-gated anion channel in the ASCT2 transporter. The K(m) for glutamine derived from these experiments is in good agreement with the K(m) derived from flux studies; it varied between 40 and 90 microM at holding potentials of -60 and -20 mV respectively. The permeability of the substrate-gated anion conductance decreased in the order SCN(-)>>NO(3)(-)>I(-)>Cl(-) and also required the presence of Na(+). PMID- 10698698 TI - Kinetic analysis of the internalization and recycling of [3H]TRH and C-terminal truncations of the long isoform of the rat thyrotropin-releasing hormone receptor 1. AB - The C-terminal tail of the long splice variant of the rat thyrotropin-releasing hormone (TRH) receptor-1 (TRHR-1L) comprises around 93 amino acids. A series of C terminal truncations was constructed and expressed transiently in HEK-293 cells. The extent of steady-state internalization of these in response to [(3)H]TRH was dependent upon the degree of truncation. Little effect was produced by deletion of the C-terminal to 50 amino acids, although there was a substantial decrease in the extent of internalization by deletion to 45-46 amino acids. The rate of internalization of TRHR-1L in response to ligand was substantially decreased by the acid-wash procedures often used in the analysis of cellular distribution of receptors with peptide ligands, and thus an alternative procedure using a Mes containing buffer was employed in the present study. Apart from a truncation anticipated to eliminate post-translational acylation of the re-ceptor, which altered both the association and dissociation rates of [(3)H]TRH, the kinetics of ligand binding were unaffected by C-terminal truncation. Equally, the rate of recycling to the plasma membrane of internalized receptors was unaffected by C terminal truncation. Although the extent of internalization of the full-length receptor was impaired by pre-exposure of cells to TRH, this was not true of C terminal truncation mutants, which displayed limited steady-state internalization ratios. A mutant with a substantial C-terminal deletion also displayed decreased functional desensitization compared with the full-length receptor. PMID- 10698700 TI - Characterization of MAX.3 antigen, a glycoprotein expressed on mature macrophages, dendritic cells and blood platelets: identity with CD84. AB - MAX.3 is a monoclonal antibody that preferentially reacts with mature macrophages (MAC), monocyte-derived dendritic cells, megakaryocytes and platelets. In this study, we describe the characterization, purification and identification of the MAX.3 antigen. Immunoprecipitation and SDS/PAGE revealed different molecular masses of MAX.3 antigen in MAC (60-90 kDa) and platelets (58-64 kDa), whereas a similar size (45 kDa) was observed in both cell types after digestion with N glycosidase F. Lectin affinity and sequential treatment with different glycosidases suggests complex type glycosylation of MAX.3 antigen in MAC and hybrid type glycosylation in platelets. Amino acid sequencing led to the identification of a corresponding cDNA clone and showed its identity to the sequence of the CD84 antigen, a member of the CD2 family of cell surface molecules. MAX.3/CD84 was further studied by immunohistochemistry and a variable expression was found on tissue MAC, confirming this antigen to be mainly a marker for MAC in situ. PMID- 10698699 TI - Identification of copper/zinc superoxide dismutase as a nitric oxide-regulated gene in human (HaCaT) keratinocytes: implications for keratinocyte proliferation. AB - Recent studies have demonstrated an induction of expression of inducible nitric oxide synthase that is associated with several inflammatory diseases of the skin. To define the mechanisms of action of nitric oxide (NO) in the skin, we attempted to identify genes that are regulated by NO in keratinocytes. Using the human keratinocyte cell line HaCaT as a model system, we identified a Cu/Zn superoxide dismutase (SOD) that was strongly induced by high concentrations (500 microM) of NO-donating agents ?S-nitrosoglutathione, sodium nitroprusside and (Z)-1-[2-(2 aminoethyl)-N-(2-ammonioethyl) amino] diazen-1-ium-1,2 -diolate (DETA-NO)?, but not by serum or by single recombinant growth factors and inflammatory cytokines or by treatment with superoxide anions. Furthermore, endogenously produced NO increased the expression of Cu/Zn SOD mRNA in keratinocytes. Moreover, treatment of HaCaT cells with NO was associated with a biphasic effect on cell proliferation, because low doses (100 microM) of different NO donors (S nitrosoglutathione and DETA-NO) mediated a proliferative signal to the cells, whereas high concentrations (500 microM) were cytostatic. To determine a possible correlation between the close regulation of Cu/Zn SOD expression and proliferation by NO in keratinocytes, we established a cell line (psp1CZ1N) carrying a human Cu/Zn SOD cDNA under the control of a ponasterone-inducible promoter construct. Ponasterone-induced overexpression of Cu/Zn SOD caused a cytostatic effect in proliferating psp1CZ1N cells. We therefore suggest that the up-regulation of Cu/Zn SOD expression by NO establishes an inhibitory mechanism on keratinocyte proliferation. PMID- 10698701 TI - Deuterium-labelled isotopomers of 2-C-methyl-D-erythritol as tools for the elucidation of the 2-C-methyl-D-erythritol 4-phosphate pathway for isoprenoid biosynthesis. AB - Escherichia coli synthesizes its isoprenoids via the mevalonate-independent 2-C methyl-D-erythritol 4-phosphate (MEP) pathway. The MC4100dxs::CAT strain, defective in deoxyxylulose-5-phosphate synthase, which is the first enzyme in this metabolic route, exclusively synthesizes its isoprenoids from exogenous 2-C methyl-D-erythritol (ME) added to the culture medium. The fate of the hydrogen atoms in the MEP pathway was followed by the incorporation of [1,1-(2)H(2)]ME and [3,5,5,5-(2)H(4)]ME. The two C-1 hydrogen atoms of ME were found without any loss in the prenyl chain of menaquinone and/or ubiquinone on the carbon atoms derived from C-4 of isopentenyl diphosphate (IPP) and on the E-methyl group of dimethylallyl diphosphate (DMAPP), the C-5 hydrogen atoms on the methyl groups derived from IPP C-5 methyl group and the Z-methyl group of DMAPP. This showed that no changes in the oxidation state of these carbon atoms occurred in the reaction sequence between MEP and IPP. Furthermore, no deuterium scrambling was observed between the carbon atoms derived from C-4 and C-5 of IPP or DMAPP, suggesting a completely stereoselective IPP isomerase or no significant activity of this enzyme. The C-3 deuterium atom of [3,5,5,5-(2)H(4)]ME was preserved only in the DMAPP starter unit and was completely missing from all those derived from IPP. This finding, aided by the non-essential role of the IPP isomerase gene, suggests the presence in E. coli of two different routes towards IPP and DMAPP, starting from a common intermediate derived from MEP. PMID- 10698703 TI - Involvement of lipoxygenase in lysophosphatidic acid-stimulated hydrogen peroxide release in human HaCaT keratinocytes. AB - Although it is now recognized that low levels of reactive oxygen species (ROS) are required for the mitogenic response, mitogen-induced signalling pathways that regulate ROS generation in non-phagocytic cells remain largely uncharacterized. Using a real-time assay for measuring hydrogen peroxide (H(2)O(2)) formation, we analysed H(2)O(2) release in human HaCaT keratinocytes in response to lysophosphatidic acid (LPA), a mitogen for keratinocytes. LPA rapidly increased H(2)O(2) release in HaCaT cells. Unlike LPA-induced mitogen-activated protein (MAP) kinase activation, LPA-stimulated H(2)O(2) release was independent of the tyrosine kinase activity of the epidermal growth factor (EGF) receptor. Calcium chelators, phospholipase A(2) inhibitors, and lipoxygenase inhibitors effectively blocked LPA-stimulated H(2)O(2) release, whereas cyclooxygenase inhibitors were without effect. Addition of 5-lipoxygenase products 5-hydroperoxyeicosatetraenoic acid (5-HPETE) and leukotriene B(4), but not 5-hydroxyeicosatetraenoic acid (5 HETE) and leukotriene C(4), restored LPA-stimulated H(2)O(2) release in cells treated with the lipoxygenase inhibitors nordihydroguaiaretic acid and Zileuton. These results suggest that the lipoxygenase products 5-HPETE and leukotriene B(4) are required for LPA-stimulated H(2)O(2) release in HaCaT cells. PMID- 10698702 TI - Different Ca2+-releasing abilities of sperm extracts compared with tissue extracts and phospholipase C isoforms in sea urchin egg homogenate and mouse eggs. AB - A soluble phospholipase C (PLC) from boar sperm generates InsP(3) and hence causes Ca(2+) release when added to sea urchin egg homogenate. This PLC activity is associated with the ability of sperm extracts to cause Ca(2+) oscillations in mammalian eggs following fractionation. A sperm PLC may, therefore, be responsible for causing the observed Ca(2+) oscillations at fertilization. In the present study we have further characterized this boar sperm PLC activity using sea urchin egg homogenate. Consistent with a sperm PLC acting on egg PtdIns(4,5)P(2), the ability of sperm extracts to release Ca(2+) was blocked by preincubation with the PLC inhibitor U73122 or by the addition of neomycin to the homogenate. The Ca(2+)-releasing activity was also detectable in sperm from other species and in whole testis extracts. However, activity was not observed in extracts from other tissues. Moreover recombinant PLCbeta1, -gamma1, -gamma2, delta1, all of which had higher specific activities than boar sperm extracts, were not able to release Ca(2+) in the sea urchin egg homogenate. In addition these PLCs were not able to cause Ca(2+) oscillations following microinjection into mouse eggs. These results imply that the sperm PLC possesses distinct properties that allow it to hydrolyse PtdIns(4,5)P(2) in eggs. PMID- 10698704 TI - Protective effect of thioredoxin upon NO-mediated cell injury in THP1 monocytic human cells. AB - Although NO has been postulated to play important roles in host defences, it is potentially damaging for exposed cells, including for the macrophages producing the NO. Thus a network of radical acceptors and enzymes is thought to play an important redox-buffering role to protect cells against NO-mediated injury. We examined the properties of the redox systems superoxide dismutase (SOD)/catalase, glutathione (GSH) and thioredoxin (Trx), in regulating the viability of two human monocytic cell lines (THP1 and U937) exposed to the NO-generating compound diethylene triamine-nitric oxide (DETA-NO). We observed that NO-induced cytotoxic effects were time- and dose-dependent towards the two cell lines. After vitamin induced differentiation in vitro with retinoic acid (RA) and 1,25-dihydroxy vitamin D(3) (VD), termed RA/VD, we observed that THP1 RA/VD cells became more resistant to NO-mediated cytotoxicity whereas the susceptibility of U937 cells was not modified. Using Western blotting and reverse-transcriptase PCR methods, we observed that gene transcription and protein expression of Trx and thioredoxin reductase were significantly increased upon RA/VD treatment and differentiation in THP1 cells. By contrast, SOD/catalase and GSH redox state remained unmodified. Finally, a stable transfectant THP1 line overexpressing Trx was found to be more resistant than THP1 control cells that were untransfected or transfected with an empty plasmid, when exposed to DETA-NO in vitro. In conclusion, we observed an inverse correlation between cell susceptibility to NO damaging effects and Trx expression, suggesting that the Trx system may have important preventative capacities towards NO-mediated cellular injury in monocytic macrophage cells. PMID- 10698705 TI - Allosteric regulation of neuronal nitric oxide synthase by tetrahydrobiopterin and suppression of auto-damaging superoxide. AB - The underlying mechanisms regulating the activity of the family of homodimeric nitric oxide synthases (NOSs) and, in particular, the requirement for (6R) 5,6,7,8-tetrahydro-L-biopterin (H(4)Bip) are not fully understood. Here we have investigated possible allosteric and stabilizing effects of H(4)Bip on neuronal NOS (NOS-I) during the conversion of substrate, L-arginine, into L-citrulline and nitric oxide. Indeed, in kinetic studies dual allosteric interactions between L arginine and H(4)Bip activated recombinant human NOS-I to increase L-arginine turnover. Consistent with this was the observation that H(4)Bip, but not the pterin-based NOS inhibitor 2-amino-4,6-dioxo-3,4,5,6,8,8a,9,10 octahydrooxazolo[1, 2-f]-pteridine (PHS-32), caused an L-arginine-dependent increase in the haem Soret band, indicating an increase in substrate binding to recombinant human NOS-I. Conversely, L-arginine was observed to increase in a concentration-dependent manner H(4)Bip binding to pig brain NOS-I. Secondly, we investigated the stabilization of NOS quaternary structure by H(4)Bip in relation to uncoupled catalysis. Under catalytic assay conditions and in the absence of H(4)Bip, dimeric recombinant human NOS-I dissociated into inactive monomers. Monomerization was related to the uncoupling of reductive oxygen activation, because it was inhibited by both superoxide dismutase and the inhibitor N(omega) nitro-L-arginine. Importantly, H(4)Bip was found to react chemically with superoxide (O(2)(-.)) and enzyme-bound H(4)Bip was consumed under O(2)(-.) generating conditions in the absence of substrate. These results suggest that H(4)Bip allosterically activates NOS-I and stabilizes quaternary structure by a novel mechanism involving the direct interception of auto-damaging O(2)(-.). PMID- 10698706 TI - GDP dissociation inhibitor D4-GDI (Rho-GDI 2), but not the homologous rho-GDI 1, is cleaved by caspase-3 during drug-induced apoptosis. AB - Different cytotoxic drugs induce cell death by activating the apoptotic programme; a family of cysteinyl aspartate proteases named caspases has been shown to be involved in the initiation as well as the execution of this kind of cell death. In the present study, cleavage of D4-GDI (Rho-GDI 2), an abundant haemopoietic-cell GDP dissociation inhibitor for the Ras-related Rho family GTPases, was demonstrated after treatment of BJAB Burkitt-like lymphoma cells with taxol or epirubicin. The cleavage of D4-GDI occurred simultaneously with the activation of caspase-3 but preceded DNA fragmentation and the morphological changes associated with apoptotic cell death. By using high-resolution two dimensional gel electrophoresis it was shown that this cleavage is specific: whereas the level of the homologous protein Rho-GDI 1 was not significantly altered during drug-induced apoptosis and in cytochrome c/dATP-activated cellular extracts, D4-GDI disappeared owing to proteolytic cleavage. Inhibitor experiments with Z-DEVD-fmk (in which Z stands for benzyloxycarbonyl and fmk for fluoromethyl ketone) and microsequencing of the D4-GDI fragment revealed that this occurs at the caspase-3 cleavage site. Our results strongly suggest the differential regulation of the homologous GDP dissociation inhibitors Rho-GDI 1 and D4-GDI during drug-induced apoptosis by proteolysis mediated by caspase-3 but not by caspase-1. Owing to their crucial role as modulators of Rho GTPases, this might in turn have a significant impact on the mechanisms that induce the cytoskeletal and morphological changes in apoptotic cells. PMID- 10698707 TI - Altered toxicity of the prion protein peptide PrP106-126 carrying the Ala(117)- >Val mutation. AB - The inherited prion diseases such as Gerstmann-Straussler-Scheinker syndrome (GSS) are linked to point mutations in the gene coding for the cellular isoform of the prion protein (PrP(C)). One particular point mutation A117V (Ala(117)- >Val) is linked to a variable pathology that usually includes deposition of neurofibrillary tangles. A prion protein peptide carrying this point mutation [PrP106-126(117V)] was generated and compared with a peptide based on the normal human sequence [PrP106-126(117A)]. The inclusion of this point mutation increased the toxicity of PrP106-126 which could be linked to an increased beta-sheet content. An assay of microtubule formation in the presence of tau indicated that PrP106-126 decreased the rate of microtubule formation that could be related to the displacement of tau. PrP106-126 carrying the 117 mutation was more efficient at inhibiting microtubule formation. These results suggest a possible mechanism of toxicity for protein carrying this mutation via destabilization of the cytoskeleton and deposition of tau in filaments, as observed in GSS. PMID- 10698708 TI - Cyclolinteinone, a sesterterpene from sponge Cacospongia linteiformis, prevents inducible nitric oxide synthase and inducible cyclo-oxygenase protein expression by blocking nuclear factor-kappaB activation in J774 macrophages. AB - We investigated the effect of cyclolinteinone, a sesterterpene from Caribbean sponge Cacospongia linteiformis, on inducible NO synthase (iNOS) and cyclo oxygenase-2 (COX-2) protein expression in lipopolysaccharide (LPS)-stimulated J774 macrophages. Incubation of J774 cells with LPS (1 microgram/ml) caused an increase of both iNOS and COX-2 protein expression, which was prevented in a concentration-dependent fashion by cyclolinteinone (12.5, 25 and 50 microM). Electrophoretic mobility-shift assay indicated that cyclolinteinone blocked the activation of nuclear factor-kappaB (NF-kappaB), a transcription factor necessary for either iNOS or COX-2 induction. Cyclolinteinone also blocked disappearance of I(kappa)B-alpha from cytosolic fraction and nuclear translocation of NF-kappaB subunits p50 and p65. These results show that cyclolinteinone down-regulates iNOS and COX-2 protein expression by inhibiting NF-kappaB activation and suggest that it may represent a novel anti-inflammatory compound capable of controlling the excessive production of prostaglandins and nitric oxide occurring in several inflammatory diseases. PMID- 10698709 TI - Cloning and characterization of a novel nuclease from shrimp hepatopancreas, and prediction of its active site. AB - Approximately 95% of the amino acid sequence of a shrimp (Penaeus japonicus) nuclease was derived from protease-digested peptides. A 1461-base cDNA for the nuclease was amplified and sequenced with degenerate primers based on the amino acid sequence and then specific primers by 3' and 5' RACE (rapid amplification of cDNA ends). It contains an open reading frame encoding a putative 21-residue signal peptide and a 381-residue mature protein. The N-terminus of the enzyme is pyroglutamate, deduced from composition and matrix-assisted laser desorption ionization-time-of-flight MS analyses, and confirmed by a glutamine residue in the cDNA sequence. The enzyme has 11 Cys residues, forming five intramolecular disulphides. The eleventh Cys residue was linked to a thiol compound with an estimated molecular mass of between 500 and 700 Da. A sequence similarity search revealed no homologous proteins but residues 205-255 shared a conserved active site motif within a distinct group of nucleases. His(211) in this conserved motif was shown to be very important in catalysis by site-specific modification with (14)C-labelled iodoacetate. The shrimp nuclease, previously designated DNase I, does indeed possess a low level of hydrolytic activity towards RNA in the presence of Mg(2+) and Ca(2+). The conservation of functionally important residues during distant evolution might imply that the catalytic mechanisms are similar in these nucleases, which should be classified in one subfamily. Finally, an active-site structure for shrimp nuclease was proposed on the basis of published structural data and the results of mutational and biochemical analyses of Serratia nuclease. PMID- 10698710 TI - Genomic organization of three novel toxins from the scorpion Buthus martensi Karsch that are active on potassium channels. AB - The cDNA and genomic DNA of three novel toxins from the scorpion Buthus martensi Karsch that are active on K(+) channels, designated BmKTX (where KTX is kaliotoxin), BmTX1 and BmTX2, were cloned and sequenced. On the basis of their known amino acid sequences, gene-specific primers for 3' and 5' rapid amplification of cDNA ends (RACE) were designed and synthesized. By overlapping the two partial cDNA sequences obtained by 3' and 5' RACE, their full-length cDNA sequences were completed. BmKTX encodes a signal peptide of 22 amino acid residues and a mature toxin of 38 residues, whereas BmTX1 and BmTX2 encode signal peptides of 20 and 21 residues respectively and a mature toxin of 38 residues for each. Their cDNA-deduced amino acid sequences were totally consistent with those determined except that the C-terminus of BmKTX had an additional Gly residue, which was removed during post-translational processing and was indispensable for the amidation of its C-terminal Lys residue. In addition, the first deduced amino acid for both BmTX1 and BmTX2 is Gln instead of pyro-Glu in the reported toxins, which obviously also undergoes post-translational processing. The genomic DNA species of these three toxins were also amplified by PCR, then cloned and sequenced. They all consisted of two exons disrupted by a small single intron. All of these introns were inserted within the signal peptides at position -6 for BmKTX and at position -5 for both BmTX1 and BmTX2 upstream of the mature toxins, and consisted of 87, 87 and 80 bp respectively. PMID- 10698711 TI - Alternative splicing for the alpha1 subunit of soluble guanylate cyclase. AB - Soluble guanylate cyclase (sGC), the receptor for nitric oxide, is a heterodimer consisting of alpha and beta subunits. We investigated the mRNA species for the alpha(1) subunit in human brain, heart, artery and immortalized B-lymphocytes. Three mRNA species were identified in these tissues. The major mRNA species contained the full expression sequence of the alpha(1) subunit. Two other types of mRNA were detected in which 5' sequences were deleted by splicing (506-590 and 412-590). Each of these deletions included the predicted translation start site, indicating that translation of these two alternatively spliced RNA species does not result in the production of full-length alpha(1) subunits. The relative amounts of the two mRNA species with deletions of the translation start site differed significantly between cell lines of immortalized B-lymphocytes from different individuals. sGC enzymic activity was significantly decreased in cellular extracts from cell lines with high proportions of mRNA species containing the deletion 506-590 when compared with extracts from cell lines that contained mostly mRNA without this deletion. PMID- 10698712 TI - Latexin, a carboxypeptidase A inhibitor, is expressed in rat peritoneal mast cells and is associated with granular structures distinct from secretory granules and lysosomes. AB - Latexin, a protein possessing inhibitory activity against rat carboxypeptidase A1 (CPA1) and CPA2, is expressed in a neuronal subset in the cerebral cortex and cells in other neural and non-neural tissues of rat. Although latexin also inhibits mast-cell CPA (MCCPA), the expression of latexin in rat mast cells has not previously been confirmed. In the present study we examined the expression and subcellular localization of latexin in rat peritoneal mast cells. Western blot and reverse-transcriptase-mediated PCR analyses showed that latexin was contained and expressed in the rat peritoneal mast cells. Immunocytochemically, latexin immunofluorescence was localized on granular structures distinct from MCCPA-, histamine- or cathepsin D-immunopositive granules. Immunoelectron microscopy revealed that latexin was associated with a minority population of granules. The latexin-associated granules were separated from MCCPA- or histamine containing granules on a self-generating density gradient of polyvinylpyrrolidone coated silica-gel particles (Percoll). Treatments with high ionic strength and heparinase released latexin from the granules, suggesting that latexin is non covalently associated with a heparin-like component of the granules. MCCPA and histamine were released from the mast cells after non-immunological and immunological stimulation with compound 48/80, A23187 and anti-IgE antibody, whereas latexin was not released. These results show that latexin is synthesized in rat peritoneal mast cells and suggest that it is associated with a unique type of intracellular granules distinct from MCCPA- and histamine-containing secretory granules and lysosomes. PMID- 10698713 TI - Analysis of the cellular functions of PTEN using catalytic domain and C-terminal mutations: differential effects of C-terminal deletion on signalling pathways downstream of phosphoinositide 3-kinase. AB - The tumour suppressor protein, PTEN (phosphatase and tensin homolog deleted on chromosome 10), is a phosphatase that can dephosphorylate tyrosine-containing peptides, Shc, focal adhesion kinase and phosphoinositide substrates. In cellular assays, PTEN has been shown to antagonize the PI-3K-dependent activation of protein kinase B (PKB) and to inhibit cell spreading and motility. It is currently unclear, however, whether PTEN accomplishes these effects through its lipid- or protein-phosphatase activity, although strong evidence has demonstrated the importance of the latter for tumour suppression by PTEN. By using a PTEN G129E (Gly(129)-->Glu) mutant that has lost its lipid phosphatase activity, while retaining protein phosphatase activity, we demonstrated a requirement for the lipid phosphatase activity of PTEN in the regulation of PKB activity, cell viability and membrane ruffling. We also made a small C-terminal deletion of PTEN, removing a putative PDZ (PSD95, Dlg and ZO1)-binding motif, with no detectable effect on the phosphatase activity of the protein expressed in HEK293 cells (human embryonic kidney 293 cells) assayed in vitro. Surprisingly, expression of this mutant revealed differential requirements for the C-terminus in the different functional assays. Wild-type and C-terminally deleted PTEN appeared to be equally active in down-regulating PKB activity, but this mutant enzyme had no effect on platelet-derived growth factor (PDGF)-induced membrane ruffling and was only partially active in a cell viability assay. These results stress the importance of the lipid phosphatase activity of PTEN in the regulation of several signalling pathways. They also identify a mutation, similar to mutations that occur in some human tumours, which removes the effect of PTEN on membrane ruffling but not that on PKB. PMID- 10698714 TI - Functional CB1 cannabinoid receptors in human vascular endothelial cells. AB - Cannabinoid CB1 receptor mRNA was detected using reverse transcription-polymerase chain reaction (RT-PCR) in endothelial cells from human aorta and hepatic artery and in the ECV304 cell line derived from human umbilical vein endothelial cells. CB1 receptor-binding sites were detected by the high-affinity antagonist radioligand [(125)I]AM-251. In ECV304 cells, both the highly potent synthetic cannabinoid agonist HU-210 and the endogenous ligand anandamide induce activation of mitogen-activated protein (MAP) kinase, and the effect of HU-210 was completely blocked, whereas the effect of anandamide was partially inhibited by SR141716A, a selective CB1 receptor antagonist. Transfection of ECV304 cells with CB1 receptor antisense, but not sense, oligonucleotides caused the same pattern of inhibition as SR141716A. This provides more definitive evidence for the involvement of CB1 receptors in MAP kinase activation and suggests that anandamide may also activate MAP kinase via an additional, CB1 receptor independent, SR141716A-resistant mechanism. The MAP kinase activation by anandamide in ECV304 cells requires genistein-sensitive tyrosine kinases and protein kinase C (PKC), and anandamide also activates p38 kinase and c-Jun kinase. These findings indicate that CB1 receptors located in human vascular endothelium are functionally coupled to the MAP kinase cascade. Activation of protein kinase cascades by anandamide may be involved in the modulation of endothelial cell growth and proliferation. PMID- 10698715 TI - Molecular mechanisms of contraction-regulated cardiac glucose transport. AB - Insulin and contraction are the most important regulators of glucose utilization in cardiac muscle. In contrast with insulin, the intracellular signalling elements of contraction have remained unexplored. In the present studies, adult rat ventricular cardiomyocytes were electrically stimulated to perform rhythmic contractions to permit the determination of potential sites of convergence of contraction and insulin signalling to glucose transport. The participation of phosphoinositide 3-kinase (PI-3K) in Ca(2+)- and contraction-stimulated 3-O methylglucose transport was suggested by the great sensitivity of this process towards the PI-3K inhibitors wortmannin and LY294002 and by the presence of PI-3K activity in anti-phosphotyrosine immunoprecipitates from contracted cells. Initial signalling events of insulin action, including receptor kinase activation, the tyrosine phosphorylation of insulin receptor substrate (IRS)-1 and IRS-2 and the recruitment of PI-3K to IRS-1 and IRS-2, were found not to be involved in contraction-mediated signalling. However, immunoprecipitation of p85alpha revealed a markedly enhanced tyrosine phosphorylation of an unknown co precipitated 200 kDa protein in response to both stimuli. It is concluded that contraction-regulated cardiac glucose transport involves the activation of PI-3K in response to upstream signalling pathways different from that of insulin. PMID- 10698716 TI - Internal-ribosome-entry-site functional activity of the 3'-untranslated region of the mRNA for the beta subunit of mitochondrial H+-ATP synthase. AB - Translation in vitro of the mammalian nucleus-encoded mRNA for the beta subunit of mitochondrial H(+)-ATP synthase (beta-mRNA) of oxidative phosphorylation is promoted by a 150 nt translational enhancer sequence in the 3'-untranslated region (3' UTR). Titration of the eukaryotic initiation factor eIF4E with cap analogue revealed that translation of capped beta-mRNA was pseudo-cap independent. The 3' UTR of beta-mRNA stimulates the translation of heterologous uncapped mRNA species, both when the 3' UTR is placed at the 3' end and at the 5' end of the transcripts. The 3' UTRs of the alpha subunit of mitochondrial H(+) ATP synthase (alpha-F1-ATPase) and subunit IV of cytochrome c oxidase (COX IV) mRNA species, other nucleus-encoded transcripts of oxidative phosphorylation, do not have the same activity in translation as the 3' UTR of beta-mRNA. On dicistronic RNA species, the 3' UTR of beta-mRNA, and to a smaller extent that of COX IV mRNA, is able to promote the translation of the second cistron to a level comparable to the activity of internal ribosome entry sites (IRESs) described in picornavirus mRNA species. These results indicate that the 3' UTRs of certain mRNA species of oxidative phosphorylation have IRES-like functional activity. Riboprobes of the active 3' UTRs on dicistronic assays formed specific RNA protein complexes when cross-linked by UV to proteins of the lysate, suggesting that cytoplasmic translation of the mRNA species bearing an active 3' UTR is assisted by specific RNA-protein interactions. PMID- 10698717 TI - Molecular cloning and expression of novel sulphotransferase-like cDNAs from human and rat brain. AB - The sulphotransferase (SULT) gene family is involved with the conjugation of many small drugs, xenobiotics and endogenous compounds. In this report, we describe the cloning and expression of novel cDNAs from human and rat brain, which are structurally related to the SULTs. These cDNAs have been termed 'brain sulphotransferase-like' (BR-STL), because of their similarity to the SULTs and their selective expression in brain tissue. The proteins encoded by the human and rat BR-STL cDNAs (hBR-STL-1 and rBR-STL cDNA respectively), denoted as hBR-STL and rBR-STL, are 98% identical and 99% similar in sequence. The hBR-STL-1 cDNA contains an 852-nt open reading frame encoding a 284-amino-acid protein with a calculated molecular mass of 33083 Da. Northern-blot analyses of RNA isolated from eight human tissues indicate that the hBR-STL message is selectively expressed in brain. Characterization of different brain regions showed that message levels were highest in cortical brain regions. rBR-STL message levels were relatively low in brains of 1-day-old male and female rats, but increased to adult levels in RNA from 7-day-old rats, and remained at that level in adult animals. The hBR-STL and rBR-STL cDNAs were expressed in both Escherichia coli and Sf9 insect cells in the presence or absence of an N-terminal histidine affinity tag (His-tag). Polyclonal antibodies were raised in chickens against purified His-tagged hBR-STL, and were used to detect the presence of rBR-STL in adult male and female rat brain cytosol. The high degree of sequence conservation, and the selective localization of the BR-STL message in brain, suggest an important function in the central nervous system. PMID- 10698718 TI - Reviews in environmental health, 2000. PMID- 10698719 TI - Imprinted genes as potential genetic and epigenetic toxicologic targets. AB - Genomic imprinting is an epigenetic phenomenon in eutherian mammals that results in the differential expression of the paternally and maternally inherited alleles of a gene. Imprinted genes are necessary for normal mammalian development. This requirement has been proposed to have evolved because of an interparental genetic battle for the utilization of maternal resources during gestation and postnatally. The nonrandom requisite for monoallelic expression of a subset of genes has also resulted in the formation of susceptibility loci for neurobehavioral disorders, developmental disorders, and cancer. Since imprinting involves both cytosine methylation within CpG islands and changes in chromatin structure, imprinted genes are potential targets for dysregulation by epigenetic toxicants that modify DNA methylation and histone acetylation. PMID- 10698720 TI - Mechanisms underlying Children's susceptibility to environmental toxicants. AB - An important public health challenge has been the need to protect children's health. To accomplish this goal, the scientific community needs scientifically based child-specific risk assessment methods. Critical to their development is the need to understand mechanisms underlying children's sensitivity to environmental toxicants. Risk is defined as the probability of adverse outcome and when applied to environmental risk assessment is usually defined as a function of both toxicity and exposure. To adequately evaluate the potential for enhanced health risks during development, both child-specific factors affecting toxicity and exposure need to be considered. In the first section of this article, example mechanisms of susceptibility relevant for toxicity assessment are identified and discussed. In the second section, examples of exposure factors that help define children's susceptibility are presented. Examples of pesticide research from the newly funded Child Health Center at the University of Washington will be given for illustration. The final section discusses the importance of putting these considerations of children's susceptibility into an overall framework for ascertaining relevancy for human risk assessment. PMID- 10698721 TI - Genetic susceptibility to lead poisoning. AB - Major strides have been taken in the regulation of lead intoxication in the general population, but studies using genetic markers of susceptibility to environmental toxicants raise the question of whether genes can make certain individuals more vulnerable to environmental toxins such as lead. At least three polymorphic genes have been identified that potentially can influence the bioaccumulation and toxicokinetics of lead in humans. The first gene to be discussed in this review is the gene coding for delta-aminolevulinic acid dehydratase (ALAD), an enzyme of heme biosynthesis, that exists in two polymorphic forms. The resulting isozymes have been shown to affect the blood and bone lead levels in human populations. The effects of ALAD in lead intoxication have also been studied in laboratory mice that differ in the genetic dose for this enzyme. The second gene reviewed here is the vitamin D receptor (VDR) gene. The VDR is involved in calcium absorption through the gut and into calcium-rich tissues such as bone. Recent findings suggest that VDR polymorphism may influence the accumulation of lead in bone. Finally, the third gene to be discussed here that may influence the absorption of lead is the hemochromatosis gene coding for the HFE protein. The presence of mutations in the HFE gene leads to hemochromatosis in homozygotic individuals. Because of the associations between iron and lead transport, it is possible that polymorphisms in the HFE gene may also influence the absorption of lead, but this has not yet been studied. More studies will be needed to define the role of these genes in lead intoxication. PMID- 10698723 TI - Molecular epidemiology studies on occupational and environmental exposure to mutagens and carcinogens, 1997-1999. AB - Molecular epidemiology is a new and evolving area of research, combining laboratory measurement of internal dose, biologically effective dose, biologic effects, and influence of individual susceptibility with epidemiologic methodologies. Biomarkers evaluated were selected according to basic scheme: biomarkers of exposure--metabolites in urine, DNA adducts, protein adducts, and Comet assay parameters; biomarkers of effect--chromosomal aberrations, sister chromatid exchanges, micronuclei, mutations in the hypoxanthine-guanine phosphoribosyltransferase gene, and the activation of oncogenes coding for p53 or p21 proteins as measured on protein levels; biomarkers of susceptibility--genetic polymorphisms of genes CYP1A1, GSTM1, GSTT1, NAT2. DNA adducts measured by 32P postlabeling are the biomarker of choice for the evaluation of exposure to polycyclic aromatic hydrocarbons. Protein adducts are useful as a biomarker for exposure to tobacco smoke (4-aminobiphenyl) or to smaller molecules such as acrylonitrile or 1,3-butadiene. Of the biomarkers of effect, the most common are cytogenetic end points. Epidemiologic studies support the use of chromosomal breakage as a relevant biomarker of cancer risk. The use of the Comet assay and methods analyzing oxidative DNA damage needs reliable validation for human biomonitoring. Until now there have not been sufficient data to interpret the relationship between genotypes, biomarkers of exposure, and biomarkers of effect for assessing the risk of human exposure to mutagens and carcinogens. PMID- 10698722 TI - The influence of nutrition on methyl mercury intoxication. AB - This article reviews progress in the research of methyl mercury (MeHg) and nutrient interactions during the past two decades. Special emphasis is placed on the following three major areas: a) effects on kinetics, b) effects on toxicity, and c) possible mechanisms. Dietary information is not usually collected in most epidemiologic studies examining of the effects of MeHg exposure. However, inconsistency of the MeHg toxicity observed in different populations is commonly attributed to possible effects of dietary modulation. Even though the mechanisms of interaction have not been totally elucidated, research in nutritional toxicology has provided insights into the understanding of the effects of nutrients on MeHg toxicity. Some of this information can be readily incorporated into the risk assessment of MeHg in the diets of fish-eating populations. It is also clear that there is a need for more studies designed specifically to address the role of nutrition in the metabolism and detoxification of MeHg. It is also important to collect more detailed dietary information in future epidemiologic studies of MeHg exposure. PMID- 10698724 TI - Sunscreens, skin photobiology, and skin cancer: the need for UVA protection and evaluation of efficacy. AB - Sunscreens are ultraviolet radiation (UVR)-absorbing chemicals that attenuate the amount and nature of UVR reaching viable cells in the skin. They are selected and tested for their ability to prevent erythema. No sunscreen prevents photodamage, as it has been demonstrated that suberythemal doses of UVR cause a variety of molecular changes (including DNA damage) in these cells. Furthermore, the spectrum of UVR reaching viable cells is altered by topically applied sunscreen. In this review, the basic aspects of sunscreens and skin photobiology are reviewed briefly. Although there can be no question concerning the efficacy of sunscreens for the prevention of erythema, questions remain because of the possible cumulative effects of chronic suberythemal doses and the increased exposure of skin cells to longer UVR wavelengths. The current major issue surrounding sunscreens involves their ability to protect skin cells against the effects of UVA radiation. These UVA effects may be direct damage (base oxidations) or effects on the skin immune system, yet there is no uniformly accepted method for the evaluation of UVA protection. This review is focused primarily on the latter topic covering action spectra that implicate the need for UVA protection. In addition, in vivo and in vitro methods proposed for the evaluation of candidate sunscreen formulations of UVA protective ability are reviewed. Finally, revisions in the terminology used to describe the protection afforded by sunscreens are suggested. It is proposed that SPF ("sun" protection factor) be renamed "sunburn" protection factor and that "critical wavelength" be designated "long wave index." PMID- 10698726 TI - Health effects of residence near hazardous waste landfill sites: a review of epidemiologic literature. AB - This review evaluates current epidemiologic literature on health effects in relation to residence near landfill sites. Increases in risk of adverse health effects (low birth weight, birth defects, certain types of cancers) have been reported near individual landfill sites and in some multisite studies, and although biases and confounding factors cannot be excluded as explanations for these findings, they may indicate real risks associated with residence near certain landfill sites. A general weakness in the reviewed studies is the lack of direct exposure measurement. An increased prevalence of self-reported health symptoms such as fatigue, sleepiness, and headaches among residents near waste sites has consistently been reported in more than 10 of the reviewed papers. It is difficult to conclude whether these symptoms are an effect of direct toxicologic action of chemicals present in waste sites, an effect of stress and fears related to the waste site, or an effect of reporting bias. Although a substantial number of studies have been conducted, risks to health from landfill sites are hard to quantify. There is insufficient exposure information and effects of low-level environmental exposure in the general population are by their nature difficult to establish. More interdisciplinary research can improve levels of knowledge on risks to human health of waste disposal in landfill sites. Research needs include epidemiologic and toxicologic studies on individual chemicals and chemical mixtures, well-designed single- and multisite landfill studies, development of biomarkers, and research on risk perception and sociologic determinants of ill health. PMID- 10698725 TI - Assessing the potential carcinogenic activity of magnetic fields using animal models. AB - We update our 1997 publication by reviewing 29 new reports of tests of magnetic fields (MFs) in six different in vivo animal models of carcinogenesis: 2-year, lifetime, or multigeneration exposure studies in rats or mice; and promotion/progression models (rat mammary carcinoma, rat liver focus, mouse skin, several models of human leukemia/lymphoma in rats and mice, and brain cancer in rats). Individual experiments are evaluated using a set of data quality criteria, and summary judgments are made across multiple experiments by applying a criterion of rough reproducibility. The potential for carcinogenicity of MFs is discussed in light of the significant body of carcinogenesis data from animal bioassays that now exists. Excluding abstracts, approximately 80% of the 41 completed studies identified in this and our previous review roughly satisfy data quality criteria. Among these studies, the criterion for independent reproducibility is not satisfied for any positive results but is satisfied for negative results in chronic bioassays in rats and mice and for negative results in both promotion and co-promotion assays using the SENCAR mouse skin model. Results of independent replication studies using the rat mammary carcinoma model were conflicting. We conclude that long-term exposure to continuous 50- or 60-Hz MFs in the range of 0.002-5 mT is unlikely to result in carcinogenesis in rats or mice. Though results of most promotion/progression assays are negative, a weak promoting effect of MFs under certain exposure conditions cannot be ruled out based on available data. PMID- 10698728 TI - Noise exposure and public health. AB - Exposure to noise constitutes a health risk. There is sufficient scientific evidence that noise exposure can induce hearing impairment, hypertension and ischemic heart disease, annoyance, sleep disturbance, and decreased school performance. For other effects such as changes in the immune system and birth defects, the evidence is limited. Most public health impacts of noise were already identified in the 1960s and noise abatement is less of a scientific but primarily a policy problem. A subject for further research is the elucidation of the mechanisms underlying noise-induced cardiovascular disorders and the relationship of noise with annoyance and nonacoustical factors modifying health outcomes. A high priority study subject is the effects of noise on children, including cognitive effects and their reversibility. Noise exposure is on the increase, especially in the general living environment, both in industrialized nations and in developing world regions. This implies that in the twenty-first century noise exposure will still be a major public health problem. PMID- 10698727 TI - Cyanobacterial toxins: removal during drinking water treatment, and human risk assessment. AB - Cyanobacteria (blue-green algae) produce toxins that may present a hazard for drinking water safety. These toxins (microcystins, nodularins, saxitoxins, anatoxin-a, anatoxin-a(s), cylindrospermopsin) are structurally diverse and their effects range from liver damage, including liver cancer, to neurotoxicity. The occurrence of cyanobacteria and their toxins in water bodies used for the production of drinking water poses a technical challenge for water utility managers. With respect to their removal in water treatment procedures, of the more than 60 microcystin congeners, microcystin-LR (L, L-leucine; R, L-arginine) is the best studied cyanobacterial toxin, whereas information for the other toxins is largely lacking. In response to the growing concern about nonlethal acute and chronic effects of microcystins, the World Health Organization has recently set a new provisional guideline value for microcystin-LR of 1.0 microg/L drinking water. This will lead to further efforts by water suppliers to develop effective treatment procedures to remove these toxins. Of the water treatment procedures discussed in this review, chlorination, possibly micro /ultrafiltration, but especially ozonation are the most effective in destroying cyanobacteria and in removing microcystins. However, these treatments may not be sufficient during bloom situations or when a high organic load is present, and toxin levels should therefore be monitored during the water treatment process. In order to perform an adequate human risk assessment of microcystin exposure via drinking water, the issue of water treatment byproducts will have to be addressed in the future. PMID- 10698729 TI - Marine algal toxins: origins, health effects, and their increased occurrence. AB - Certain marine algae produce potent toxins that impact human health through the consumption of contaminated shellfish and finfish and through water or aerosol exposure. Over the past three decades, the frequency and global distribution of toxic algal incidents appear to have increased, and human intoxications from novel algal sources have occurred. This increase is of particular concern, since it parallels recent evidence of large-scale ecologic disturbances that coincide with trends in global warming. The extent to which human activities have contributed to their increase therefore comes into question. This review summarizes the origins and health effects of marine algal toxins, as well as changes in their current global distribution, and examines possible causes for the recent increase in their occurrence. PMID- 10698730 TI - Infectious disease and worldwide declines of amphibian populations, with comments on emerging diseases in coral reef organisms and in humans. AB - Many populations of amphibians are declining on all six continents on which they occur. Some causes of amphibian declines, such as habitat destruction, direct application of xenobiotics, and introduction of predators or competitors, are clearly attributable to human activities. Infectious disease appears to be the direct cause of mass amphibian die-offs in relatively undisturbed areas of the world where anthropomorphic environmental disruption is minimal. In these cases, it is not yet clear whether these epizootics result from the natural evolution of new pathogens or from environmental changes that promote the emergence of pathogenic forms and/or that weaken the immune defenses of amphibians. Because some aspects of pathogen-related amphibian mass mortalities are similar to outbreaks of new diseases in humans and coral reef organisms, amphibian declines may be part of a much larger pattern than previously appreciated. PMID- 10698731 TI - The catalytic pathway of cytochrome p450cam at atomic resolution. AB - Members of the cytochrome P450 superfamily catalyze the addition of molecular oxygen to nonactivated hydrocarbons at physiological temperature-a reaction that requires high temperature to proceed in the absence of a catalyst. Structures were obtained for three intermediates in the hydroxylation reaction of camphor by P450cam with trapping techniques and cryocrystallography. The structure of the ferrous dioxygen adduct of P450cam was determined with 0.91 angstrom wavelength x rays; irradiation with 1.5 angstrom x-rays results in breakdown of the dioxygen molecule to an intermediate that would be consistent with an oxyferryl species. The structures show conformational changes in several important residues and reveal a network of bound water molecules that may provide the protons needed for the reaction. PMID- 10698732 TI - Mid-Pleistocene Acheulean-like stone technology of the Bose basin, South China. AB - Stone artifacts from the Bose basin, South China, are associated with tektites dated to 803,000 +/- 3000 years ago and represent the oldest known large cutting tools (LCTs) in East Asia. Bose toolmaking is compatible with Mode 2 (Acheulean) technologies in Africa in its targeted manufacture and biased spatial distribution of LCTs, large-scale flaking, and high flake scar counts. Acheulean like tools in the mid-Pleistocene of South China imply that Mode 2 technical advances were manifested in East Asia contemporaneously with handaxe technology in Africa and western Eurasia. Bose lithic technology is associated with a tektite airfall and forest burning. PMID- 10698733 TI - Isotope fractionation and atmospheric oxygen: implications for phanerozoic O(2) evolution AB - Models describing the evolution of the partial pressure of atmospheric oxygen over Phanerozoic time are constrained by the mass balances required between the inputs and outputs of carbon and sulfur to the oceans. This constraint has limited the applicability of proposed negative feedback mechanisms for maintaining levels of atmospheric O(2) at biologically permissable levels. Here we describe a modeling approach that incorporates O(2)-dependent carbon and sulfur isotope fractionation using data obtained from laboratory experiments on carbon-13 discrimination by vascular land plants and marine plankton. The model allows us to calculate a Phanerozoic O(2) history that agrees with independent models and with biological and physical constraints and supports the hypothesis of a high atmospheric O(2) content during the Carboniferous (300 million years ago), a time when insect gigantism was widespread. PMID- 10698734 TI - Natural NaAlSi(3)O(8)-hollandite in the shocked sixiangkou meteorite AB - The hollandite high-pressure polymorph of plagioclase has been identified in shock-induced melt veins of the Sixiangkou L6 chondrite. It is intimately intergrown with feldspathic glass within grains previously thought to be "maskelynite." The crystallographic nature of the mineral was established by laser micro-Raman spectroscopy and x-ray diffraction. The mineral is tetragonal with the unit cell parameters a = 9.263 +/- 0.003 angstroms and c = 2.706 +/- 0.003 angstroms. Its occurrence with the liquidus pair majorite-pyrope solid solution plus magnesiowustite sets constraints on the peak pressures that prevailed in the shock-induced melt veins. The absence of a calcium ferrite structured phase sets an upper bound for the crystallization of the hollandite polymorph near 23 gigapascals. PMID- 10698735 TI - Green, catalytic oxidation of alcohols in water AB - Alcohol oxidations are typically performed with stoichiometric reagents that generate heavy-metal waste and are usually run in chlorinated solvents. A water soluble palladium(II) bathophenanthroline complex is a stable recyclable catalyst for the selective aerobic oxidation of a wide range of alcohols to aldehydes, ketones, and carboxylic acids in a biphasic water-alcohol system. The use of water as a solvent and air as the oxidant makes the reaction interesting from both an economic and environmental point of view. PMID- 10698736 TI - Translation of the edited mRNA for cytochrome b in trypanosome mitochondria. AB - The type of RNA editing found in the kinetoplast-mitochondria of trypanosomes and related protozoa, involving uridylate insertions and deletions, creates translatable messenger RNAs (mRNAs) out of nonsense pre-edited RNAs by correcting encoded defects that vary from simple frameshifts to large "cryptic" regions. However, any evidence for translation of these mRNAs in the kinetoplast has been missing for decades. We identified a kinetoplast-encoded protein, apocytochrome b, whose mRNA is edited in the 5' region. The determined amino-terminal sequence of the protein coincides with the predicted sequence derived from the edited region, demonstrating that the cognate apocytochrome b mRNA is translated into a functional protein. This finding represents the first direct evidence for a functional translation system in the kinetoplasts. PMID- 10698737 TI - A potassium channel protein encoded by chlorella virus PBCV-1. AB - The large chlorella virus PBCV-1, which contains double-stranded DNA (dsDNA), encodes a 94-codon open reading frame (ORF) that contains a motif resembling the signature sequence of the pore domain of potassium channel proteins. Phylogenetic analyses of the encoded protein, Kcv, indicate a previously unidentified type of potassium channel. The messenger RNA encoded by the ORF leads to functional expression of a potassium-selective conductance in Xenopus laevis oocytes. The channel blockers amantadine and barium, but not cesium, inhibit this conductance, in addition to virus plaque formation. Thus, PBCV-1 encodes the first known viral protein that functions as a potassium-selective channel and is essential in the virus life cycle. PMID- 10698738 TI - Requirement of the prolyl isomerase Pin1 for the replication checkpoint. AB - The peptidyl-prolyl isomerase Pin1 has been implicated in regulating cell cycle progression. Pin1 was found to be required for the DNA replication checkpoint in Xenopus laevis. Egg extracts depleted of Pin1 inappropriately transited from the G2 to the M phase of the cell cycle in the presence of the DNA replication inhibitor aphidicolin. This defect in replication checkpoint function was reversed after the addition of recombinant wild-type Pin1, but not an isomerase inactive mutant, to the depleted extract. Premature mitotic entry in the absence of Pin1 was accompanied by hyperphosphorylation of Cdc25, activation of Cdc2/cyclin B, and generation of epitopes recognized by the mitotic phosphoprotein antibody, MPM-2. Therefore, Pin1 appears to be required for the checkpoint delaying the onset of mitosis in response to incomplete replication. PMID- 10698739 TI - Requirement of the inositol trisphosphate receptor for activation of store operated Ca2+ channels. AB - The coupling mechanism between endoplasmic reticulum (ER) calcium ion (Ca2+) stores and plasma membrane (PM) store-operated channels (SOCs) is crucial to Ca2+ signaling but has eluded detection. SOCs may be functionally related to the TRP family of receptor-operated channels. Direct comparison of endogenous SOCs with stably expressed TRP3 channels in human embryonic kidney (HEK293) cells revealed that TRP3 channels differ in being store independent. However, condensed cortical F-actin prevented activation of both SOC and TRP3 channels, which suggests that ER-PM interactions underlie coupling of both channels. A cell-permeant inhibitor of inositol trisphosphate receptor (InsP3R) function, 2-aminoethoxydiphenyl borate, prevented both receptor-induced TRP3 activation and store-induced SOC activation. It is concluded that InsP3Rs mediate both SOC and TRP channel opening and that the InsP3R is essential for maintaining coupling between store emptying and physiological activation of SOCs. PMID- 10698740 TI - An ultrasensitive bacterial motor revealed by monitoring signaling proteins in single cells. AB - Understanding biology at the single-cell level requires simultaneous measurements of biochemical parameters and behavioral characteristics in individual cells. Here, the output of individual flagellar motors in Escherichia coli was measured as a function of the intracellular concentration of the chemotactic signaling protein. The concentration of this molecule, fused to green fluorescent protein, was monitored with fluorescence correlation spectroscopy. Motors from different bacteria exhibited an identical steep input-output relation, suggesting that they actively contribute to signal amplification in chemotaxis. This experimental approach can be extended to quantitative in vivo studies of other biochemical networks. PMID- 10698741 TI - Salmonella pathogenicity island 2-dependent evasion of the phagocyte NADPH oxidase. AB - A type III protein secretion system encoded by Salmonella pathogenicity island 2 (SPI2) has been found to be required for virulence and survival within macrophages. Here, SPI2 was shown to allow Salmonella typhimurium to avoid NADPH oxidase-dependent killing by macrophages. The ability of SPI2-mutant bacteria to survive in macrophages and to cause lethal infection in mice was restored by abrogation of the NADPH oxidase-dependent respiratory burst. Ultrastructural and immunofluorescence microscopy demonstrated efficient localization of the NADPH oxidase in the proximity of vacuoles containing SPI2-mutant but not wild-type bacteria, suggesting that SPI2 interferes with trafficking of oxidase-containing vesicles to the phagosome. PMID- 10698742 TI - Resonant formation of DNA strand breaks by low-energy (3 to 20 eV) electrons. AB - Most of the energy deposited in cells by ionizing radiation is channeled into the production of abundant free secondary electrons with ballistic energies between 1 and 20 electron volts. Here it is shown that reactions of such electrons, even at energies well below ionization thresholds, induce substantial yields of single- and double-strand breaks in DNA, which are caused by rapid decays of transient molecular resonances localized on the DNA's basic components. This finding presents a fundamental challenge to the traditional notion that genotoxic damage by secondary electrons can only occur at energies above the onset of ionization, or upon solvation when they become a slowly reacting chemical species. PMID- 10698743 TI - Dual signaling regulated by calcyon, a D1 dopamine receptor interacting protein. AB - The synergistic response of cells to the stimulation of multiple receptors has been ascribed to receptor cross talk; however, the specific molecules that mediate the resultant signal amplification have not been defined. Here a 24 kilodalton single transmembrane protein, designated calcyon, we functionally characterize that interacts with the D1 dopamine receptor. Calcyon localizes to dendritic spines of D1 receptor-expressing pyramidal cells in prefrontal cortex. These studies delineate a mechanism of Gq- and Gs-coupled heterotrimeric GTP binding protein-coupled receptor cross talk by which D1 receptors can shift effector coupling to stimulate robust intracellular calcium (Ca2+i) release as a result of interaction with calcyon. The role of calcyon in potentiating Ca2+ dependent signaling should provide insight into the D1 receptor-modulated cognitive functions of prefrontal cortex. PMID- 10698744 TI - Epitopes involved in antibody-mediated protection from Ebola virus. AB - To determine the ability of antibodies to provide protection from Ebola viruses, monoclonal antibodies (mAbs) to the Ebola glycoprotein were generated and evaluated for efficacy. We identified several protective mAbs directed toward five unique epitopes on Ebola glycoprotein. One of the epitopes is conserved among all Ebola viruses that are known to be pathogenic for humans. Some protective mAbs were also effective therapeutically when administered to mice 2 days after exposure to lethal Ebola virus. The identification of protective mAbs has important implications for developing vaccines and therapies for Ebola virus. PMID- 10698745 TI - Leaching of Escherichia coli O157:H7 in diverse soils under various agricultural management practices. AB - Application of animal manures to soil as crop fertilizers is an important means for recycling the nitrogen and phosphorus which the manures contain. Animal manures also contain bacteria, including many types of pathogens. Manure pathogen levels depend on the source animal, the animal's state of health, and how the manure was stored or treated before use. Rainfall may result in pathogen spread into soil by runoff from stored or unincorporated manure or by leaching through the soil profile. Steady rainfall consisting of 16.5 mm h(-1) was applied to 100 mm disturbed soil cores that were treated with manure and inoculated with Escherichia coli O157:H7 strain B6914. The level of B6914 in leachate was near the inoculum level each hour for 8 h, as was the level of B6914 at several soil depths after 24 h, indicating that there was a high rate of growth. Bacterial movement through three different types of soil was then compared by using disturbed (tilled) and intact (no-till) soil cores and less intense rainfall consisting of 25.4 mm on 4 consecutive days and then four more times over a 17 day period. Total B6914 levels exceeded the inoculum levels for all treatments except intact clay loam cores. B6914 levels in daily leachate samples decreased sharply with time, although the levels were more constant when intact sandy loam cores were used. The presence of manure often increased total B6914 leachate and soil levels in intact cores but had the opposite effect on disturbed soil cores. Ammonia and nitrate levels correlated with B6914 and total coliform levels in leachate. We concluded that tillage practice, soil type, and method of pathogen delivery affect but do not prevent vertical E. coli O157:H7 and coliform transport in soil and that soluble nitrogen may enhance transport. PMID- 10698746 TI - Overexpression of trigger factor prevents aggregation of recombinant proteins in Escherichia coli. AB - To examine the effects of overexpression of trigger factor (TF) on recombinant proteins produced in Escherichia coli, we constructed plasmids that permitted controlled expression of TF alone or together with the GroEL-GroES chaperones. The following three proteins that are prone to aggregation were tested as targets: mouse endostatin, human oxygen-regulated protein ORP150, and human lysozyme. The results revealed that TF overexpression had marked effects on the production of these proteins in soluble forms, presumably through facilitating correct folding. Whereas overexpression of TF alone was sufficient to prevent aggregation of endostatin, overexpression of TF together with GroEL-GroES was more effective for ORP150 and lysozyme, suggesting that TF and GroEL-GroES play synergistic roles in vivo. Although coexpression of the DnaK-DnaJ-GrpE chaperones was also effective for endostatin and ORP150, coexpression of TF and GroEL-GroES was more effective for lysozyme. These results attest to the usefulness of the present expression plasmids for improving protein production in E. coli. PMID- 10698747 TI - Simultaneous enhancement of thermostability and catalytic activity of phospholipase A(1) by evolutionary molecular engineering. AB - The thermal stability and catalytic activity of phospholipase A(1) from Serratia sp. strain MK1 were improved by evolutionary molecular engineering. Two thermostable mutants were isolated after sequential rounds of error-prone PCR performed to introduce random mutations and filter-based screening of the resultant mutant library; we determined that these mutants had six (mutant TA3) and seven (mutant TA13) amino acid substitutions. Different types of substitutions were found in the two mutants, and these substitutions resulted in an increase in nonpolar residues (mutant TA3) or in differences between side chains for polar or charged residues (mutant TA13). The wild-type and mutant enzymes were purified, and the effect of temperature on the stability and catalytic activity of the enzymes was investigated. The melting temperatures of the TA3 and TA13 enzymes were increased by 7 and 11 degrees C, respectively, compared with the melting temperature of the wild-type enzyme. Thus, we found that evolutionary molecular engineering was an effective and efficient approach for increasing thermostability without compromising enzyme activity. PMID- 10698748 TI - Genetic analysis of chromosomal regions of Lactococcus lactis acquired by recombinant lytic phages. AB - Recombinant phages are generated when Lactococcus lactis subsp. lactis harboring plasmids encoding the abortive type (Abi) of phage resistance mechanisms is infected with small isometric phages belonging to the P335 species. These phage variants are likely to be an important source of virulent new phages that appear in dairy fermentations. They are distinguished from their progenitors by resistance to Abi defenses and by altered genome organization, including regions of L. lactis chromosomal DNA. The objective of this study was to characterize four recombinant variants that arose from infection of L. lactis NCK203 (Abi(+)) with phage phi31. HindIII restriction maps of the variants (phi31.1, phi31.2, phi31.7, and phi31.8) were generated, and these maps revealed the regions containing recombinant DNA. The recombinant region of phage phi31.1, the variant that occurred most frequently, was sequenced and revealed 7.8 kb of new DNA compared with the parent phage, phi31. This region contained numerous instances of homology with various lactococcal temperate phages, as well as homologues of the lambda recombination protein BET and Escherichia coli Holliday junction resolvase Rus, factors which may contribute to efficient recombination processes. A sequence analysis and phenotypic tests revealed a new origin of replication in the phi31.1 DNA, which replaced the phi31 origin. Three separate HindIII fragments, accounting for most of the recombinant region of phi31.1, were separately cloned into gram-positive suicide vector pTRK333 and transformed into NCK203. Chromosomal insertions of each plasmid prevented the appearance of different combinations of recombinant phages. The chromosomal insertions did not affect an inducible prophage present in NCK203. Our results demonstrated that recombinant phages can acquire DNA cassettes from different regions of the chromosome in order to overcome Abi defenses. Disruption of these regions by insertion can alter the types and diversity of new phages that appear during phage-host interactions. PMID- 10698749 TI - Gene cloning and nucleotide sequencing and properties of a cocaine esterase from Rhodococcus sp. strain MB1. AB - A strain of Rhodococcus designated MB1, which was capable of utilizing cocaine as a sole source of carbon and nitrogen for growth, was isolated from rhizosphere soil of the tropane alkaloid-producing plant Erythroxylum coca. A cocaine esterase was found to initiate degradation of cocaine, which was hydrolyzed to ecgonine methyl ester and benzoate; both of these esterolytic products were further metabolized by Rhodococcus sp. strain MB1. The structural gene encoding a cocaine esterase, designated cocE, was cloned from Rhodococcus sp. strain MB1 genomic libraries by screening recombinant strains of Rhodococcus erythropolis CW25 for growth on cocaine. The nucleotide sequence of cocE corresponded to an open reading frame of 1,724 bp that codes for a protein of 574 amino acids. The amino acid sequence of cocaine esterase has a region of similarity with the active serine consensus of X-prolyl dipeptidyl aminopeptidases, suggesting that the cocaine esterase is a serine esterase. The cocE coding sequence was subcloned into the pCFX1 expression plasmid and expressed in Escherichia coli. The recombinant cocaine esterase was purified to apparent homogeneity and was found to be monomeric, with an M(r) of approximately 65,000. The apparent K(m) of the enzyme (mean +/- standard deviation) for cocaine was measured as 1.33 +/- 0.085 mM. These findings are of potential use in the development of a linked assay for the detection of illicit cocaine. PMID- 10698750 TI - Inactivation of isocitrate lyase leads to increased production of medium-chain length poly(3-hydroxyalkanoates) in Pseudomonas putida. AB - Medium-chain-length (mcl) poly(3-hydroxyalkanoates) (PHAs) are storage polymers that are produced from various substrates and accumulate in Pseudomonas strains belonging to rRNA homology group I. In experiments aimed at increasing PHA production in Pseudomonas strains, we generated an mcl PHA-overproducing mutant of Pseudomonas putida KT2442 by transposon mutagenesis, in which the aceA gene was knocked out. This mutation inactivated the glyoxylate shunt and reduced the in vitro activity of isocitrate dehydrogenase, a rate-limiting enzyme of the citric acid cycle. The genotype of the mutant was confirmed by DNA sequencing, and the phenotype was confirmed by biochemical experiments. The aceA mutant was not able to grow on acetate as a sole carbon source due to disruption of the glyoxylate bypass and exhibited two- to fivefold lower isocitrate dehydrogenase activity than the wild type. During growth on gluconate, the difference between the mean PHA accumulation in the mutant and the mean PHA accumulation in the wild type strain was 52%, which resulted in a significant increase in the amount of mcl PHA at the end of the exponential phase in the mutant P. putida KT217. On the basis of a stoichiometric flux analysis we predicted that knockout of the glyoxylate pathway in addition to reduced flux through isocitrate dehydrogenase should lead to increased flux into the fatty acid synthesis pathway. Therefore, enhanced carbon flow towards the fatty acid synthesis pathway increased the amount of mcl PHA that could be accumulated by the mutant. PMID- 10698751 TI - Influence of Acanthamoeba castellanii on intracellular growth of different Legionella species in human monocytes. AB - Previous studies using a murine model of coinhalation of Legionella pneumophila and Hartmannella vermiformis have shown a significantly enhanced intrapulmonary growth of L. pneumophila in comparison to inhalation of legionellae alone (J. Brieland, M. McClain, L. Heath, C. Chrisp, G. Huffnagle, M. LeGendre, M. Hurley, J. Fantone, and C. Engleberg, Infect. Immun. 64:2449-2456, 1996). In this study, we introduce an in vitro coculture model of legionellae, Mono Mac 6 cells (MM6) and Acanthamoeba castellanii, using a cell culture chamber system which separates both cell types by a microporous polycarbonate membrane impervious to bacteria, amoebae, and human cells. Whereas L. pneumophila has shown a maximal 4-log-unit multiplication within MM6, which could not be further increased by coculture with Acanthamoeba castellanii, significantly enhanced replication of L. gormanii, L. micdadei, L. steigerwaltii, L. longbeachae, and L. dumoffii was seen after coculture with amoebae. This effect was seen only with uninfected amoebae, not with Legionella-infected amoebae. The supporting effect for intracellular multiplication in MM6 could be reproduced in part by addition of a cell-free coculture supernatant obtained from a coincubation experiment with uninfected A. castellanii and Legionella-infected MM6, suggesting that amoeba-derived effector molecules are involved in this phenomenon. This coculture model allows investigations of molecular and biochemical mechanisms which are responsible for the enhancement of intracellular multiplication of legionellae in monocytic cells after interaction with amoebae. PMID- 10698752 TI - Copper induction of laccase isoenzymes in the ligninolytic fungus Pleurotus ostreatus. AB - Pleurotus ostreatus is a white rot basidiomycete that produces several extracellular laccase isoenzymes, including phenol oxidase A1b (POXA1b), POXA2, and POXC. POXC was the most abundant isoenzyme produced under all of the growth conditions examined in this study. Copper was the most efficient inducer of laccase activity among the putative inducers tested. The amounts of all of the previously described laccase isoenzymes increased substantially in copper supplemented cultures. Under these conditions expression of POX isoenzymes was regulated at the level of gene transcription. It is worth noting that poxa1b mRNA was the most abundant induced transcript at all of the growth times analyzed, and the amount of this mRNA increased until day 7. The discrepancy between the poxa1b transcript and protein amounts can be explained by the presence of a high level of the protein in P. ostreatus cellular extract, which indicated that the POXA1b isoenzyme could be inefficiently secreted and/or that its physiological activity could occur inside the cell or on the cell wall. Moreover, the POXA1b isoenzyme behaved uniquely, as its activity was maximal on the second day of growth and then decreased. An analysis performed with protease inhibitors revealed that the loss of extracellular POXA1b activity could have been due to the presence of specific proteases secreted into the copper-containing culture medium that affected the extracellular POXA1b isoenzyme. PMID- 10698753 TI - Biochemical and molecular characterization of a laccase from Marasmius quercophilus. AB - The basidiomycete Marasmius quercophilus is commonly found during autumn on the decaying litter of the evergreen oak (Quercus ilex L.), a plant characteristic of Mediterranean forest. This white-rot fungus colonizes the leaf surface with rhizomorphs, causing a total bleaching of the leaf. In synthetic liquid media, this white-rot fungus has strong laccase activity. From a three-step chromatographic procedure, we purified a major isoform to homogeneity. The gene encodes a monomeric glycoprotein of approximately 63 kDa, with a 3.6 isoelectric point, that contains 12% carbohydrate. Spectroscopic analysis of the purified enzyme (UV/visible and electron paramagnetic resonance, atomic absorption) confirmed that it belongs to the "blue copper oxidase" family. With syringaldazine as the substrate, the enzyme's pH optimum was 4.5, the optimal temperature was 75 degrees C, and the K(m) was 7.1 microM. The structural gene, lac1, was cloned and sequenced. This gene encodes a 517-amino-acid protein 99% identical to a laccase produced by PM1, an unidentified basidiomycete previously isolated from wastewater from a paper factory in Spain. This similarity may be explained by the ecological distribution of the evergreen oak in Mediterranean forest. PMID- 10698754 TI - Succession of microbial communities during hot composting as detected by PCR single-strand-conformation polymorphism-based genetic profiles of small-subunit rRNA genes. AB - A cultivation-independent technique for genetic profiling of PCR-amplified small subunit rRNA genes (SSU rDNA) was chosen to characterize the diversity and succession of microbial communities during composting of an organic agricultural substrate. PCR amplifications were performed with DNA directly extracted from compost samples and with primers targeting either (i) the V4-V5 region of eubacterial 16S rRNA genes, (ii) the V3 region in the 16S rRNA genes of actinomycetes, or (iii) the V8-V9 region of fungal 18S rRNA genes. Homologous PCR products were converted to single-stranded DNA molecules by exonuclease digestion and were subsequently electrophoretically separated by their single-strand conformation polymorphism (SSCP). Genetic profiles obtained by this technique showed a succession and increasing diversity of microbial populations with all primers. A total of 19 single products were isolated from the profiles by PCR reamplification and cloning. DNA sequencing of these molecular isolates showed similarities in the range of 92.3 to 100% to known gram-positive bacteria with a low or high G+C DNA content and to the SSU rDNA of gamma-Proteobacteria. The amplified 18S rRNA gene sequences were related to the respective gene regions of Candida krusei and Candida tropicalis. Specific molecular isolates could be attributed to different composting stages. The diversity of cultivated bacteria isolated from samples taken at the end of the composting process was low. A total of 290 isolates were related to only 6 different species. Two or three of these species were also detectable in the SSCP community profiles. Our study indicates that community SSCP profiles can be highly useful for the monitoring of bacterial diversity and community successions in a biotechnologically relevant process. PMID- 10698755 TI - Nickel availability and hupSL activation by heterologous regulators limit symbiotic expression of the Rhizobium leguminosarum bv. viciae hydrogenase system in Hup(-) rhizobia. AB - A limited number of Rhizobium and Bradyrhizobium strains possess a hydrogen uptake (Hup) system that recycles the hydrogen released from the nitrogen fixation process in legume nodules. To extend this ability to rhizobia that nodulate agronomically important crops, we investigated factors that affect the expression of a cosmid-borne Hup system from Rhizobium leguminosarum bv. viciae UPM791 in R. leguminosarum bv. viciae, Rhizobium etli, Mesorhizobium loti, and Sinorhizobium meliloti Hup(-) strains. After cosmid pAL618 carrying the entire hup system of strain UPM791 was introduced, all recipient strains acquired the ability to oxidize H(2) in symbioses with their hosts, although the levels of hydrogenase activity were found to be strain and species dependent. The levels of hydrogenase activity were correlated with the levels of nickel-dependent processing of the hydrogenase structural polypeptides and with transcription of structural genes. Expression of the NifA-dependent hupSL promoter varied depending on the genetic background, while the hyp operon, which is controlled by the FnrN transcriptional regulator, was expressed at similar levels in all recipient strains. With the exception of the R. etli-bean symbiosis, the availability of nickel to bacteroids strongly affected hydrogenase processing and activity in the systems tested. Our results indicate that efficient transcriptional activation by heterologous regulators and processing of the hydrogenase as a function of the availability of nickel to the bacteroid are relevant factors that affect hydrogenase expression in heterologous rhizobia. PMID- 10698756 TI - Expression of outer membrane proteins in Escherichia coli growing at acid pH. AB - It is generally accepted for Escherichia coli that (i) the level of OmpC increases with increased osmolarity when cells are growing in neutral and alkaline media, whereas the level of OmpF decreases at high osmolarity, and that (ii) the two-component system composed of OmpR (regulator) and EnvZ (sensor) regulates porin expression. In this study, we found that OmpC was expressed at low osmolarity in medium of pH below 6 and that the expression was repressed when medium osmolarity was increased. In contrast, the expression of ompF at acidic pH was essentially the same as that at alkaline pH. Neither OmpC nor OmpF was detectable in an ompR mutant at both acid and alkaline pH values. However, OmpC and OmpF were well expressed at acid pH in a mutant envZ strain, and their expression was regulated by medium osmolarity. Thus, it appears that E. coli has a different mechanism for porin expression at acid pH. A mutant deficient in ompR grew slower than its parent strain in low-osmolarity medium at acid pH (below 5.5). The same growth diminution was observed when ompC and ompF were deleted, suggesting that both OmpF and OmpC are required for optimal growth under hypoosmosis at acid pH. PMID- 10698757 TI - Frequency and biodiversity of 2,4-diacetylphloroglucinol-producing bacteria isolated from the maize rhizosphere at different stages of plant growth. AB - A Pseudomonas 2,4-diacetylphloroglucinol (DAPG)-producing population that occurred naturally on the roots, in rhizosphere soil of Zea mays and in the nonrhizosphere soil was investigated in order to assess the microbial diversity at five stages of plant growth. A total of 1,716 isolates were obtained, and 188 of these isolates were able to produce DAPG. DAPG producers were isolated at each stage of plant growth, indicating that the maize rhizosphere is colonized by natural DAPG producers throughout development. The frequency of DAPG producers was very low in the first stage of plant growth and increased over time. An analysis of the level of biodiversity of the DAPG producers at the species level was performed by comparing the AluI restriction patterns of the 16S ribosomal DNAs (rDNAs) amplified by PCR from 167 isolates. This comparison allowed us to cluster the isolates into four amplified rDNA restriction analysis (ARDRA) groups, and the main group (ARDRA group 1) contained 89.8% of the isolates. The diversity of the 150 isolates belonging to ARDRA group 1 was analyzed by the random amplified polymorphic DNA (RAPD) technique. An analysis of RAPD patterns by a molecular variance method revealed that there was a high level of genetic diversity in this population and that the genetic diversity was related to plant age. Finally, we found that some of the DAPG producers, which originated from all stages of plant growth, had the same genotype. These DAPG producers could be exploited in future screening programs for biocontrol agents. PMID- 10698758 TI - Increase in bacterial community diversity in subsurface aquifers receiving livestock wastewater input. AB - Despite intensive studies of microbial-community diversity, the questions of which kinds of microbial populations are associated with changes in community diversity have not yet been fully solved by molecular approaches. In this study, to investigate the impact of livestock wastewater on changes in the bacterial communities in groundwater, bacterial communities in subsurface aquifers were analyzed by characterizing their 16S rDNA sequences. The similarity coefficients of restriction fragment length polymorphism (RFLP) patterns of the cloned 16S ribosomal DNAs showed that the bacterial communities in livestock wastewater samples were more closely related to those in contaminated aquifer samples. In addition, calculations of community diversity clearly showed that bacterial communities in the livestock wastewater and the contaminated aquifer were much more diverse than those in the uncontaminated aquifer. Thus, the increase in bacterial-community diversity in the contaminated aquifer was assumed to be due to the infiltration of livestock wastewater, containing high concentrations of diverse microbial flora, into the aquifer. Phylogenetic analysis of the sequences from a subset of the RFLP patterns showed that the Cytophaga-Flexibacter Bacteroides and low-G+C gram-positive groups originating from livestock wastewater were responsible for the change in the bacterial community in groundwater. This was evidenced by the occurrence of rumen-related sequences not only in the livestock wastewater samples but also in the contaminated-groundwater samples. Rumen-related sequences, therefore, can be used as indicator sequences for fecal contamination of groundwater, particularly from livestock. PMID- 10698759 TI - Molecular analysis of the pmo (particulate methane monooxygenase) operons from two type II methanotrophs. AB - The particulate methane monooxygenase gene clusters, pmoCAB, from two representative type II methanotrophs of the alpha-Proteobacteria, Methylosinus trichosporium OB3b and Methylocystis sp. strain M, have been cloned and sequenced. Primer extension experiments revealed that the pmo cluster is probably transcribed from a single transcriptional start site located 300 bp upstream of the start of the first gene, pmoC, for Methylocystis sp. strain M. Immediately upstream of the putative start site, consensus sequences for sigma(70) promoters were identified, suggesting that these pmo genes are recognized by sigma(70) and negatively regulated under low-copper conditions. The pmo genes were cloned in several overlapping fragments, since parts of these genes appeared to be toxic to the Escherichia coli host. Methanotrophs contain two virtually identical copies of pmo genes, and it was necessary to use Southern blotting and probing with pmo gene fragments in order to differentiate between the two pmoCAB clusters in both methanotrophs. The complete DNA sequence of one copy of pmo genes from each organism is reported here. The gene sequences are 84% similar to each other and 75% similar to that of a type I methanotroph of the gamma-Proteobacteria, Methylococcus capsulatus Bath. The derived proteins PmoC and PmoA are predicted to be highly hydrophobic and consist mainly of transmembrane-spanning regions, whereas PmoB has only two putative transmembrane-spanning helices. Hybridization experiments showed that there are two copies of pmoC in both M. trichosporium OB3b and Methylocystis sp. strain M, and not three copies as found in M. capsulatus Bath. PMID- 10698760 TI - Effects of pH and trace minerals on long-term starvation of Leuconostoc mesenteroides. AB - Laboratory experiments have definitively shown that exopolymer-producing bacteria have the potential to modify the flow of fluids in oil reservoirs to enhance oil production. Once injected into the reservoir, they will be subjected to a wide range of pH values and to starvation resulting from nutrient depletion. For successful field implementation it is necessary to have a fundamental understanding of these effects on the viability of bacteria. This paper addresses the effects of pH and trace minerals on cell viability of Leuconostoc mesenteroides during carbon source depletion. Two different carbon sources were used to grow cells before transferring the cells to starvation conditions: sucrose and a combination of glucose and fructose. These substrates were chosen because L. mesenteroides produces a significant amount of water-insoluble exopolymers (dextran) under sucrose-fed conditions, which may enhance cell survival under harsh conditions. The effects of dextran on the cell viability were tested at different pH values with and without trace minerals. The rate of cell death followed an exponential-decay law for different values of the solution pH. The optimal solution pH for survival was pH 5, whereas cells died rapidly at pH 3 and below and at pH 13 and above. The sucrose-fed cells showed a greater viability than cells fed glucose and fructose for all pH ranges tested. The results indicated that water-insoluble exopolymers help cells survive for longer periods of time under starvation conditions. The effects of trace minerals on cell culturability were tested at two pH values, 4.5 and 7. For both cases, cells showed a greater culturability (smaller decay rate constant) in the presence of trace minerals than without trace minerals. It was also found that the effects of trace minerals on cell culturability were greater for glucose-fructose-fed cells than for sucrose-fed cells. The Michaelis pH function theory was used for comparing the relationships between the cell decay rate and pH. PMID- 10698761 TI - Prevalence of Lyme disease Borrelia spp. in ticks from migratory birds on the Japanese mainland. AB - Borrelia sp. prevalence in ticks on migratory birds was surveyed in central Japan. In autumn, a total of 1,733 birds representing 40 species were examined for ticks. A total of 361 ticks were obtained from 173 birds of 15 species, and these ticks were immature Haemaphysalis flava (94.4%), Haemaphysalis longicornis, Ixodes columnae, Ixodes persulcatus, Ixodes turdus, and an unidentified Ixodes species. Of these, 27 juveniles of H. flava on Turdus pallidus, Turdus cardis, or Emberiza spodocephala, 2 juveniles of I. persulcatus on T. pallidus, and 1 female H. flava molted from a T. pallidus-derived nymph were positive for the presence of Borrelia by Barbour-Stoenner-Kelly culture passages. In spring, a total of 16 ticks obtained from 102 birds of 21 species were negative for the spirochete. Isolates from 15 ticks were characterized by 5S-23S rRNA intergenic spacer restriction fragment length polymorphism analysis; all isolates were identified as Borrelia garinii with pattern B/B' based on the previous patterning. According to the intergenic spacer sequences, 2 of 15 isolates, strains Fi14f and Fi24f, were highly similar to B. garinii strains 935T of Korea and ChY13p of Inner Mongolia, China, respectively. These findings indicate that Lyme disease-causing B. garinii may have been introduced to Japan by migratory birds from northeastern China via Korea. Additionally, a case of transstadial transmission of B. garinii from nymph to adult H. flava suggests that the infected H. flava may transmit Borrelia to large animals. PMID- 10698762 TI - Multiplex PCR for detection and identification of lactococcal bacteriophages. AB - Three genetically distinct groups of Lactococcus lactis phages are encountered in dairy plants worldwide, namely, the 936, c2, and P335 species. The multiplex PCR method was adapted to detect, in a single reaction, the presence of these species in whey samples or in phage lysates. Three sets of primers, one for each species, were designed based on conserved regions of their genomes. The c2-specific primers were constructed using the major capsid protein gene (mcp) as the target. The mcp sequences for three phages (eb1, Q38, and Q44) were determined and compared with the two available in the databases, those for phages c2 and bIL67. An 86.4% identity was found over the five mcp genes. The gene of the only major structural protein (msp) was selected as a target for the detection of 936 related phages. The msp sequences for three phages (p2, Q7, and Q11) were also established and matched with the available data on phages sk1, bIL170, and F4-1. The comparison of the six msp genes revealed an 82. 2% identity. A high genomic diversity was observed among structural proteins of the P335-like phages suggesting that the classification of lactococcal phages within this species should be revised. Nevertheless, we have identified a common genomic region in 10 P335-like phages isolated from six countries. This region corresponded to orfF17 orf18 of phage r1t and orf20-orf21 of Tuc2009 and was sequenced for three additional P335 phages (Q30, P270, and ul40). An identity of 93.4% within a 739 bp region of the five phages was found. The detection limit of the multiplex PCR method in whey was 10(4) to 10(7) PFU/ml and was 10(3) to 10(5) PFU/ml with an additional phage concentration step. The method can also be used to detect phage DNA in whey powders and may also detect prophage or defective phage in the bacterial genome. PMID- 10698763 TI - High-affinity maltose binding and transport by the thermophilic anaerobe Thermoanaerobacter ethanolicus 39E. AB - Thermoanaerobacter ethanolicus is a gram-positive thermophile that produces considerable amounts of ethanol from soluble sugars and polymeric substrates, including starch. Growth on maltose, a product of starch hydrolysis, was associated with the production of a prominent membrane-associated protein that had an apparent molecular weight of 43,800 and was not detected in cells grown on xylose or glucose. Filter-binding assays revealed that cell membranes bound maltose with high affinity. Metabolic labeling of T. ethanolicus maltose-grown cells with [(14)C]palmitic acid showed that this protein was posttranslationally acylated. A maltose-binding protein was purified by using an amylose resin affinity column, and the binding constant was 270 nM. Since maltase activity was found only in the cytosol of fractionated cells and unlabeled glucose did not compete with radiolabeled maltose for uptake in whole cells, it appeared that maltose was transported intact. In whole-cell transport assays, the affinity for maltose was approximately 40 nM. Maltotriose and alpha-trehalose competitively inhibited maltose uptake in transport assays, whereas glucose, cellobiose, and a range of disaccharides had little effect. Based on these results, it appears that T. ethanolicus possesses a high-affinity, ABC type transport system that is specific for maltose, maltotriose, and alpha-trehalose. PMID- 10698764 TI - Detection of DNA damage in prokaryotes by terminal deoxyribonucleotide transferase-mediated dUTP nick end labeling. AB - Numerous agents can damage the DNA of prokaryotes in the environment (e.g., reactive oxygen species, irradiation, and secondary metabolites such as antibiotics, enzymes, starvation, etc.). The large number of potential DNA damaging agents, as well as their diverse modes of action, precludes a simple test of DNA damage based on detection of nucleic acid breakdown products. In this study, free 3'-OH DNA ends, produced by either direct damage or excision DNA repair, were used to assess DNA damage. Terminal deoxyribonucleotide transferase (TdT)-mediated dUTP nick end labeling (TUNEL) is a procedure in which 3'-OH DNA ends are enzymatically labeled with dUTP-fluorescein isothiocyanate using TdT. Cells labeled by this method can be detected using fluorescence microscopy or flow cytometry. TUNEL was used to measure hydrogen peroxide-induced DNA damage in the archaeon Haloferax volcanii and the bacterium Escherichia coli. DNA repair systems were implicated in the hydrogen peroxide-dependent generation of 3'-OH DNA ends by the finding that the protein synthesis inhibitors chloramphenicol and diphtheria toxin blocked TUNEL labeling of E. coli and H. volcanii, respectively. DNA damage induced by UV light and bacteriophage infection was also measured using TUNEL. This methodology should be useful in applications where DNA damage and repair are of interest, including mutant screening and monitoring of DNA damage in the environment. PMID- 10698765 TI - Degradation and mineralization of high-molecular-weight polycyclic aromatic hydrocarbons by defined fungal-bacterial cocultures. AB - This study investigated the biodegradation of high-molecular-weight polycyclic aromatic hydrocarbons (PAHs) in liquid media and soil by bacteria (Stenotrophomonas maltophilia VUN 10,010 and bacterial consortium VUN 10,009) and a fungus (Penicillium janthinellum VUO 10, 201) that were isolated from separate creosote- and manufactured-gas plant-contaminated soils. The bacteria could use pyrene as their sole carbon and energy source in a basal salts medium (BSM) and mineralized significant amounts of benzo[a]pyrene cometabolically when pyrene was also present in BSM. P. janthinellum VUO 10,201 could not utilize any high molecular-weight PAH as sole carbon and energy source but could partially degrade these if cultured in a nutrient broth. Although small amounts of chrysene, benz[a]anthracene, benzo[a]pyrene, and dibenz[a,h]anthracene were degraded by axenic cultures of these isolates in BSM containing a single PAH, such conditions did not support significant microbial growth or PAH mineralization. However, significant degradation of, and microbial growth on, pyrene, chrysene, benz[a]anthracene, benzo[a]pyrene, and dibenz[a,h]anthracene, each as a single PAH in BSM, occurred when P. janthinellum VUO 10,201 and either bacterial consortium VUN 10,009 or S. maltophilia VUN 10,010 were combined in the one culture, i.e., fungal-bacterial cocultures: 25% of the benzo[a]pyrene was mineralized to CO(2) by these cocultures over 49 days, accompanied by transient accumulation and disappearance of intermediates detected by high-pressure liquid chromatography. Inoculation of fungal-bacterial cocultures into PAH-contaminated soil resulted in significantly improved degradation of high-molecular-weight PAHs, benzo[a]pyrene mineralization (53% of added [(14)C]benzo[a]pyrene was recovered as (14)CO(2) in 100 days), and reduction in the mutagenicity of organic soil extracts, compared with the indigenous microbes and soil amended with only axenic inocula. PMID- 10698766 TI - Fusarium species from nepalese rice and production of mycotoxins and gibberellic acid by selected species. AB - Infection of cereal grains with Fusarium species can cause contamination with mycotoxins that affect human and animal health. To determine the potential for mycotoxin contamination, we isolated Fusarium species from samples of rice seeds that were collected in 1997 on farms in the foothills of the Nepal Himalaya. The predominant Fusarium species in surface-disinfested seeds with husks were species of the Gibberella fujikuroi complex, including G. fujikuroi mating population A (anamorph, Fusarium verticillioides), G. fujikuroi mating population C (anamorph, Fusarium fujikuroi), and G. fujikuroi mating population D (anamorph, Fusarium proliferatum). The widespread occurrence of mating population D suggests that its role in the complex symptoms of bakanae disease of rice may be significant. Other common species were Gibberella zeae (anamorph, Fusarium graminearum) and Fusarium semitectum, with Fusarium acuminatum, Fusarium anguioides, Fusarium avenaceum, Fusarium chlamydosporum, Fusarium equiseti, and Fusarium oxysporum occasionally present. Strains of mating population C produced beauvericin, moniliformin, and gibberellic acid, but little or no fumonisin, whereas strains of mating population D produced beauvericin, fumonisin, and, usually, moniliformin, but no gibberellic acid. Some strains of G. zeae produced the 8-ketotrichothecene nivalenol, whereas others produced deoxynivalenol. Despite the occurrence of fumonisin-producing strains of mating population D, and of 8-ketotrichothecene producing strains of G. zeae, Nepalese rice showed no detectable contamination with these mycotoxins. Effective traditional practices for grain drying and storage may prevent contamination of Nepalese rice with Fusarium mycotoxins. PMID- 10698767 TI - pH regulation of pectate lyase secretion modulates the attack of Colletotrichum gloeosporioides on avocado fruits. AB - Growth of Colletotrichum gloeosporioides in pectolytic enzyme-inducing medium (PEIM) increased the pH of the medium from 3. 8 to 6.5. Pectate lyase (PL) secretion was detected when the pH reached 5.8, and the level of secretion increased up to pH 6.5. PL gene (pel) transcript production began at pH 5.0 and increased up to pH 5.7. PL secretion was never detected when the pH of the inducing medium was lower than 5.8 or when C. gloeosporioides hyphae were transferred from PL-secreting conditions at pH 6.5 to pH 3.8. This behavior differed from that of polygalacturonase (PG), where pg transcripts and protein secretion were detected at pH 5.0 and continued up to 5.7. Under in vivo conditions, the pH of unripe pericarp of freshly harvested avocado (Persea americana cv. Fuerte) fruits, resistant to C. gloeosporioides attack, was 5.2, whereas in ripe fruits, when decay symptoms were expressed, the pericarp pH had increased to 6.3. Two avocado cultivars, Ardit and Ettinger, which are resistant to C. gloeosporioides attack, had pericarp pHs of less than 5.5, which did not increase during ripening. The present results suggest that host pH regulates the secretion of PL and may affect C. gloeosporioides pathogenicity. The mechanism found in avocado may have equivalents in other post-harvest pathosystems and suggests new approaches for breeding against and controlling post-harvest diseases. PMID- 10698768 TI - Selective inhibition of the oxidation of ferrous iron or sulfur in Thiobacillus ferrooxidans. AB - The oxidation of either ferrous iron or sulfur by Thiobacillus ferrooxidans was selectively inhibited or controlled by various anions, inhibitors, and osmotic pressure. Iron oxidation was more sensitive than sulfur oxidation to inhibition by chloride, phosphate, and nitrate at low concentrations (below 0.1 M) and also to inhibition by azide and cyanide. Sulfur oxidation was more sensitive than iron oxidation to the inhibitory effect of high osmotic pressure. These differences were evident not only between iron oxidation by iron-grown cells and sulfur oxidation by sulfur-grown cells but also between the iron and sulfur oxidation activities of the same iron-grown cells. Growth experiments with ferrous iron or sulfur as an oxidizable substrate confirmed the higher sensitivity of iron oxidation to inhibition by phosphate, chloride, azide, and cyanide. Sulfur oxidation was actually stimulated by 50 mM phosphate or chloride. Leaching of Fe and Zn from pyrite (FeS(2)) and sphalerite (ZnS) by T. ferrooxidans was differentially affected by phosphate and chloride, which inhibited the solubilization of Fe without significantly affecting the solubilization of Zn. PMID- 10698769 TI - Highly ordered vertical structure of Synechococcus populations within the one millimeter-thick photic zone of a hot spring cyanobacterial mat. AB - A variety of contemporary techniques were used to investigate the vertical distribution of thermophilic unicellular cyanobacteria, Synechococcus spp., and their activity within the upper 1-mm-thick photic zone of the mat community found in an alkaline siliceous hot spring in Yellowstone National Park in Wyoming. Detailed measurements were made over a diel cycle at a 61 degrees C site. Net oxygenic photosynthesis measured with oxygen microelectrodes was highest within the uppermost 100- to 200-microm-thick layer until midmorning, but as the day progressed, the peak of net activity shifted to deeper layers, stabilizing at a depth of 300 microm from midday throughout the afternoon. Examination of vertical thin sections by bright-field and autofluorescence microscopy revealed the existence of different populations of Synechococcus which form discrete bands at different vertical positions. Denaturing gradient gel electrophoresis analysis of PCR-amplified 16S rRNA gene segments from horizontal cryosections obtained at 100 microm-thick vertical intervals also suggested vertical stratification of cyanobacterial, green sulfur bacterium-like, and green nonsulfur bacterium-like populations. There was no evidence of diel migration. However, image analysis of vertical thin sections revealed the presence of a narrow band of rod-shaped Synechococcus cells in which the cells assumed an upright position. These upright cells, located 400 to 800 microm below the surface, were observed only in mat samples obtained around noon. In mat samples obtained at other time points, the cells were randomly oriented throughout the mat. These combined observations reveal the existence of a highly ordered structure within the very thin photic zone of this hot spring microbial mat, consisting of morphologically similar Synechococcus populations that are likely to be differentially adapted, some co occurring with green sulfur bacterium-like populations, and all overlying green nonsulfur bacterium-like populations. PMID- 10698770 TI - Reduction of Fe(III), Mn(IV), and toxic metals at 100 degrees C by Pyrobaculum islandicum. AB - It has recently been noted that a diversity of hyperthermophilic microorganisms have the ability to reduce Fe(III) with hydrogen as the electron donor, but the reduction of Fe(III) or other metals by these organisms has not been previously examined in detail. When Pyrobaculum islandicum was grown at 100 degrees C in a medium with hydrogen as the electron donor and Fe(III)-citrate as the electron acceptor, the increase in cell numbers of P. islandicum per mole of Fe(III) reduced was found to be ca. 10-fold higher than previously reported. Poorly crystalline Fe(III) oxide could also serve as the electron acceptor for growth on hydrogen. The stoichiometry of hydrogen uptake and Fe(III) oxide reduction was consistent with the oxidation of 1 mol of hydrogen resulting in the reduction of 2 mol of Fe(III). The poorly crystalline Fe(III) oxide was reduced to extracellular magnetite. P. islandicum could not effectively reduce the crystalline Fe(III) oxide minerals goethite and hematite. In addition to using hydrogen as an electron donor for Fe(III) reduction, P. islandicum grew via Fe(III) reduction in media in which peptone and yeast extract served as potential electron donors. The closely related species P. aerophilum grew via Fe(III) reduction in a similar complex medium. Cell suspensions of P. islandicum reduced the following metals with hydrogen as the electron donor: U(VI), Tc(VII), Cr(VI), Co(III), and Mn(IV). The reduction of these metals was dependent upon the presence of cells and hydrogen. The metalloids arsenate and selenate were not reduced. U(VI) was reduced to the insoluble U(IV) mineral uraninite, which was extracellular. Tc(VII) was reduced to insoluble Tc(IV) or Tc(V). Cr(VI) was reduced to the less toxic, less soluble Cr(III). Co(III) was reduced to Co(II). Mn(IV) was reduced to Mn(II) with the formation of manganese carbonate. These results demonstrate that biological reduction may contribute to the speciation of metals in hydrothermal environments and could account for such phenomena as magnetite accumulation and the formation of uranium deposits at ca. 100 degrees C. Reduction of toxic metals with hyperthermophilic microorganisms or their enzymes might be applied to the remediation of metal-contaminated waters or waste streams. PMID- 10698771 TI - Flow cytometry for determination of the efficacy of contact lens disinfecting solutions against Acanthamoeba spp. AB - Flow cytometric analyses of cellular staining with fluorescent viability dyes and direct microscopic observations of methylene blue exclusion were compared for evaluation of the effects of a chlorhexidine gluconate-based contact lens disinfectant solution and a polyhexamethylene biguanide solution against cysts and trophozoites of Acanthamoeba castellanii and Acanthamoeba polyphaga. The flow cytometric procedure with propidium iodide (used to stain dead cells) indicated that more than 90% of trophozoites of both species (inocula of 10(5) to 10(6)/ml) at 22 degrees C lost their viability after 4 h of exposure to chlorhexidine. When propidium iodide was used in combination with fluorescein diacetate (for live cells), the apparent number of propidium iodide-stained cells was reduced, but the relative efficacies of the two biguanide solutions appeared unchanged from those evident with the single dyes; the chlorhexidine solution was more effective than the polyhexamethylene biguanide solution. Similar data were obtained with the more cumbersome methylene blue exclusion procedure. Flow cytometric analyses provided a statistically reproducible and rapid procedure for determining the relative antiamoebal efficacies of the disinfecting solutions. PMID- 10698772 TI - Bacterial reductive dissolution of crystalline Fe(III) oxide in continuous-flow column reactors. AB - Bacterial reductive dissolution of synthetic crystalline Fe(III) oxide-coated sand was studied in continuous-flow column reactors in comparison with parallel batch cultures. The cumulative amount of aqueous Fe(II) exported from the columns over a 6-month incubation period corresponded to (95.0 +/- 3.7)% (n = 3) of their original Fe(III) content. Wet-chemical analysis revealed that only (6.5 +/- 3.2)% of the initial Fe(III) content remained in the columns at the end of the experiment. The near-quantitative removal of Fe was visibly evidenced by extensive bleaching of color from the sand in the columns. In contrast to the column reactors, Fe(II) production quickly reached an asymptote in batch cultures, and only (13.0 +/- 2.2)% (n = 3) of the Fe(III) oxide content was reduced. Sustained bacterial-cell growth occurred in the column reactors, leading to the production and export of a quantity of cells 100-fold greater than that added during inoculation. Indirect estimates of cell growth, based on the quantity of Fe(III) reduced, suggest that only an approximate doubling of initial cell abundance was likely to have occurred in the batch cultures. Our results indicate that removal of biogenic Fe(II) via aqueous-phase transport in the column reactors decreased the passivating influence of surface-bound Fe(II) on oxide reduction activity, thereby allowing a dramatic increase in the extent of Fe(III) oxide reduction and associated bacterial growth. These findings have important implications for understanding the fate of organic and inorganic contaminants whose geochemical behavior is linked to Fe(III) oxide reduction. PMID- 10698773 TI - Prevention of yeast spoilage in feed and food by the yeast mycocin HMK. AB - The yeast Williopsis mrakii produces a mycocin or yeast killer toxin designated HMK; this toxin exhibits high thermal stability, high pH stability, and a broad spectrum of activity against other yeasts. We describe construction of a synthetic gene for mycocin HMK and heterologous expression of this toxin in Aspergillus niger. Mycocin HMK was fused to a glucoamylase protein carrier, which resulted in secretion of biologically active mycocin into the culture media. A partial purification protocol was developed, and a comparison with native W. mrakii mycocin showed that the heterologously expressed mycocin had similar physiological properties and an almost identical spectrum of biological activity against a number of yeasts isolated from silage and yoghurt. Two food and feed production systems prone to yeast spoilage were used as models to assess the ability of mycocin HMK to act as a biocontrol agent. The onset of aerobic spoilage in mature maize silage was delayed by application of A. niger mycocin HMK on opening because the toxin inhibited growth of the indigenous spoilage yeasts. This helped maintain both higher lactic acid levels and a lower pH. In yoghurt spiked with dairy spoilage yeasts, A. niger mycocin HMK was active at all of the storage temperatures tested at which yeast growth occurred, and there was no resurgence of resistant yeasts. The higher the yeast growth rate, the more effective the killing action of the mycocin. Thus, mycocin HMK has potential applications in controlling both silage spoilage and yoghurt spoilage caused by yeasts. PMID- 10698774 TI - Growth of Escherichia coli O157:H7 in bruised apple (Malus domestica) tissue as influenced by cultivar, date of harvest, and source. AB - Four of five apple cultivars (Golden Delicious, Red Delicious, McIntosh, Macoun, and Melrose) inoculated with Escherichia coli O157:H7 promoted growth of the bacterium in bruised tissue independent of the date of harvest (i.e., degree of apple ripening) or the source of the apple (i.e., tree-picked or dropped fruit). Apple harvest for this study began 4 September 1998 and ended 9 October, with weekly sampling. Throughout this study, freshly picked (<2 days after harvest) McIntosh apples usually prevented the growth of E. coli O157:H7 for 2 days. Growth of E. coli O157:H7 did occur following 6 days of incubation in bruised McIntosh apple tissue. However, the maximum total cell number was approximately 80-fold less than the maximum total cell number recovered from Red Delicious apples. When fruit was stored for 1 month at 4 degrees C prior to inoculation with E. coli O157:H7, all five cultivars supported growth of the bacterium. For each apple cultivar, the pH of bruised tissue was significantly higher and degrees Brix was significantly lower than the pH and degrees Brix of undamaged tissue regardless of the source. In freshly picked apples, changes in the pH did not occur over the harvest season. Bruised Golden Delicious, McIntosh, and Melrose apple tissue pHs were not significantly different (tree-picked or dropped), and the degrees Brix values of McIntosh, Macoun, and Melrose apple tissue were not significantly different. Single-cultivar preparations of cider did not support growth of E. coli, and the cell concentration of inoculated cider declined over an 11-day test period. The rate of decline in E. coli cell concentration in the McIntosh cider was greater than those in the other ciders tested. The findings of this study suggested that the presence of some factor besides, or in addition to, pH inhibited E. coli growth in McIntosh apples. PMID- 10698775 TI - Characterization and determination of origin of lactic acid bacteria from a sorghum-based fermented weaning food by analysis of soluble proteins and amplified fragment length polymorphism fingerprinting. AB - The group that includes the lactic acid bacteria is one of the most diverse groups of bacteria known, and these organisms have been characterized extensively by using different techniques. In this study, 180 lactic acid bacterial strains isolated from sorghum powder (44 strains) and from corresponding fermented (93 strains) and cooked fermented (43 strains) porridge samples that were prepared in 15 households were characterized by using biochemical and physiological methods, as well as by analyzing the electrophoretic profiles of total soluble proteins. A total of 58 of the 180 strains were Lactobacillus plantarum strains, 47 were Leuconostoc mesenteroides strains, 25 were Lactobacillus sake-Lactobacillus curvatus strains, 17 were Pediococcus pentosaceus strains, 13 were Pediococcus acidilactici strains, and 7 were Lactococcus lactis strains. L. plantarum and L. mesenteroides strains were the dominant strains during the fermentation process and were recovered from 87 and 73% of the households, respectively. The potential origins of these groups of lactic acid bacteria were assessed by amplified fragment length polymorphism fingerprint analysis. PMID- 10698776 TI - Cyt1A from Bacillus thuringiensis synergizes activity of Bacillus sphaericus against Aedes aegypti (Diptera: Culicidae). AB - Bacillus sphaericus is a mosquitocidal bacterium recently developed as a commercial larvicide that is used worldwide to control pestiferous and vector mosquitoes. Whereas B. sphaericus is highly active against larvae of Culex and Anopheles mosquitoes, it is virtually nontoxic to Aedes aegypti, an important vector species. In the present study, we evaluated the capacity of the cytolytic protein Cyt1A from Bacillus thuringiensis subsp. israelensis to enhance the toxicity of B. sphaericus toward A. aegypti. Various combinations of these two materials were evaluated, and all were highly toxic. A ratio of 10:1 of B. sphaericus to Cyt1A was 3, 600-fold more toxic to A. aegypti than B. sphaericus alone. Statistical analysis showed this high activity was due to synergism between the Cyt1A toxin and B. sphaericus. These results suggest that Cyt1A could be useful in expanding the host range of B. sphaericus. PMID- 10698777 TI - Genetic and biochemical diversity among isolates of Paenibacillus alvei cultured from Australian honeybee (Apis mellifera) colonies. AB - Twenty-five unique CfoI-generated whole-cell DNA profiles were identified in a study of 30 Paenibacillus alvei isolates cultured from honey and diseased larvae collected from honeybee (Apis mellifera) colonies in geographically diverse areas in Australia. The fingerprint patterns were highly variable and readily discernible from one another, which highlighted the potential of this method for tracing the movement of isolates in epidemiological studies. 16S rRNA gene fragments (length, 1,416 bp) for all 30 isolates were enzymatically amplified by PCR and subjected to restriction analysis with DraI, HinfI, CfoI, AluI, FokI, and RsaI. With each enzyme the restriction profiles of the 16S rRNA genes from all 30 isolates were identical (one restriction fragment length polymorphism [RFLP] was observed in the HinfI profile of the 16S rRNA gene from isolate 17), which confirmed that the isolates belonged to the same species. The restriction profiles generated by using DraI, FokI, and HinfI differentiated P. alvei from the phylogenetically closely related species Paenibacillus macerans and Paenibacillus macquariensis. Alveolysin gene fragments (length, 1, 555 bp) were enzymatically amplified from some of the P. alvei isolates (19 of 30 isolates), and RFLP were detected by using the enzymes CfoI, Sau3AI, and RsaI. Extrachromosomal DNA ranging in size from 1 to 10 kb was detected in 17 of 30 (57%) P. alvei whole-cell DNA profiles. Extensive biochemical heterogeneity was observed among the 28 P. alvei isolates examined with the API 50CHB system. All of these isolates were catalase, oxidase, and Voges-Proskauer positive and nitrate negative, and all produced acid when glycerol, esculin, and maltose were added. The isolates produced variable results for 16 of the 49 biochemical tests; negative reactions were recorded in the remaining 30 assays. The genetic and biochemical heterogeneity in P. alvei isolates may be a reflection of adaptation to the special habitats in which they originated. PMID- 10698778 TI - Identification and characterization of a bile acid 7alpha-dehydroxylation operon in Clostridium sp. strain TO-931, a highly active 7alpha-dehydroxylating strain isolated from human feces. AB - Clostridium sp. strain TO-931 can rapidly convert the primary bile acid cholic acid to a potentially toxic compound, deoxycholic acid. Mixed oligonucleotide probes were used to isolate a gene fragment encoding a putative bile acid transporter from Clostridium sp. strain TO-931. This DNA fragment had 60% nucleotide sequence identity to a known bile acid transporter gene from Eubacterium sp. strain VPI 12708, another bile acid-7alpha-dehydroxylating intestinal bacterium. The DNA (9.15 kb) surrounding the transporter gene was cloned from Clostridium sp. strain TO-931 and sequenced. Within this larger DNA fragment was a 7.9-kb region, containing six successive open reading frames (ORFs), that was encoded by a single 8.1-kb transcript, as determined by Northern blot analysis. The gene arrangement and DNA sequence of the Clostridium sp. strain TO-931 operon are similar to those of a Eubacterium sp. strain VPI 12708 bile acid-inducible operon containing nine ORFs. Several genes in the Eubacterium sp. strain VPI 12708 operon have been shown to encode products required for bile acid 7alpha-dehydroxylation. In Clostridium sp. strain TO-931, genes potentially encoding bile acid-coenzyme A (CoA) ligase, 3alpha-hydroxysteroid dehydrogenase, bile acid 7alpha-dehydratase, bile acid-CoA hydrolase, and a bile acid transporter were similar in size and exhibited amino acid homology to similar gene products from Eubacterium sp. strain VPI 12708 (encoded by baiB, baiA, baiE, baiF, and baiG, respectively). However, no genes similar to Eubacterium sp. strain VPI 12708 biaH or baiI were found in the Clostridium sp. strain TO-931 bai operon, and the two putative Eubacterium sp. strain VPI 12708 genes, baiC and baiD, were arranged in one continuous ORF in Clostridium sp. strain TO-931. Intergene regions showed no significant DNA sequence similarity, but primer extension analysis identified a region 115 bp upstream from the first ORF that exhibited 58% identity to a bai operator/promoter region identified in Eubacterium sp. strain VPI 12708. These results indicate that the gene organization, gene product amino acid sequences, and promoters of the bile acid inducible operons of Clostridium sp. strain TO-931 and Eubacterium sp. strain VPI 12708 are highly conserved. PMID- 10698779 TI - Initial colonization, nutrient supply, and fungal activity on leaves decaying in streams. AB - Aquatic hyphomycetes dominate leaf decomposition in streams, and their biomass is an important component in the diet of leaf-eating invertebrates. After 2 weeks of exposure in a first-order stream, maple leaf disks had low levels of fungal biomass and species diversity. Spore production by aquatic hyphomycetes also was low. Subsets of these disks were left in the stream for another 3 weeks or incubated in defined mineral solutions with one of three levels of nitrate and phosphate. Stream disks lost mass, increased ergosterol levels and spore production, and were colonized by additional fungal species. External N and P significantly stimulated mass loss, ergosterol accumulation, and spore production of laboratory disks. On disks incubated without added N and P, ergosterol levels declined while conidium production continued, suggesting conversion of existing hyphal biomass to propagules. In all other treatments, approximately equal amounts of newly synthesized biomass were invested in hyphae and conidia. Net yield (fungal biomass per leaf mass lost) varied between 1% (in the laboratory, without added N or P) and 31% (decay in stream). In most treatments, the three aquatic hyphomycete species that dominated spore production during the first 2 weeks in the stream also produced the largest numbers of conidia in the following 3 weeks. Principal-component analysis suggested two divergent trends from the initial fungal community established after 2 weeks in the stream. One culminated in the community of the second phase of stream exposure, and the other culminated in the laboratory treatment with the highest levels of N and P. The results suggest that fungal production in streams, and, by extension, production of invertebrates and higher tropic levels, is stimulated by inorganic N and P. PMID- 10698780 TI - Medium-chain fatty acids affect citrinin production in the filamentous fungus Monascus ruber. AB - During submerged culture in the presence of glucose and glutamate, the filamentous fungus Monascus ruber produces water-soluble red pigments together with citrinin, a mycotoxin with nephrotoxic and hepatoxic effects on animals. Analysis of the (13)C-pigment molecules from mycelia cultivated with [1-(13)C]-, [2-(13)C]-, or [1, 2-(13)C]acetate by (13)C nuclear magnetic resonance indicated that the biosynthesis of the red pigments used both the polyketide pathway, to generate the chromophore structure, and the fatty acid synthesis pathway, to produce a medium-chain fatty acid (octanoic acid) which was then bound to the chromophore by a trans-esterification reaction. Hence, to enhance pigment production, we tried to short-circuit the de novo synthesis of medium-chain fatty acids by adding them to the culture broth. Of fatty acids with carbon chains ranging from 6 to 18 carbon atoms, only octanoic acid showed a 30 to 50% stimulation of red pigment production, by a mechanism which, in contrast to expectation, did not involve its direct trans-esterification on the chromophore backbone. However, the medium- and long-chain fatty acids tested were readily assimilated by the fungus, and in the case of fatty acids ranging from 8 to 12 carbon atoms, 30 to 40% of their initial amount transiently accumulated in the growth medium in the form of the corresponding methylketone 1 carbon unit shorter. Very interestingly, these fatty acids or their corresponding methylketones caused a strong reduction in, or even a complete inhibition of, citrinin production by M. ruber when they were added to the medium. Several data indicated that this effect could be due to the degradation of the newly synthesized citrinin (or an intermediate in the citrinin pathway) by hydrogen peroxide resulting from peroxisome proliferation induced by medium-chain fatty acids or methylketones. PMID- 10698781 TI - Biomarker evidence for widespread anaerobic methane oxidation in Mediterranean sediments by a consortium of methanogenic archaea and bacteria. The Medinaut Shipboard Scientific Party. AB - Although abundant geochemical data indicate that anaerobic methane oxidation occurs in marine sediments, the linkage to specific microorganisms remains unclear. In order to examine processes of methane consumption and oxidation, sediment samples from mud volcanoes at two distinct sites on the Mediterranean Ridge were collected via the submersible Nautile. Geochemical data strongly indicate that methane is oxidized under anaerobic conditions, and compound specific carbon isotope analyses indicate that this reaction is facilitated by a consortium of archaea and bacteria. Specifically, these methane-rich sediments contain high abundances of methanogen-specific biomarkers that are significantly depleted in (13)C (delta(13)C values are as low as -95 per thousand). Biomarkers inferred to derive from sulfate-reducing bacteria and other heterotrophic bacteria are similarly depleted. Consistent with previous work, such depletion can be explained by consumption of (13)C-depleted methane by methanogens operating in reverse and as part a consortium of organisms in which sulfate serves as the terminal electron acceptor. Moreover, our results indicate that this process is widespread in Mediterranean mud volcanoes and in some localized settings is the predominant microbiological process. PMID- 10698782 TI - Reactivation of insertionally inactivated Shiga toxin 2 genes of Escherichia coli O157:H7 caused by nonreplicative transposition of the insertion sequence. AB - IS1203v is an insertion sequence which has been found in inactivated Shiga toxin 2 genes of Escherichia coli O157:H7. We analyzed the transpositional mechanism of IS1203v in order to investigate whether the Shiga toxin 2 genes inactivated by IS1203v could revert to the wild type. When the transposase activity of IS1203v was enhanced by artificial frameshifting, IS1203v was obviously excised from the Shiga toxin 2 gene in a circular form. The IS1203v circle consisted of the entire IS1203v, but an extra 3-bp sequence (ATC) intervened between the 5' and 3' ends of IS1203v. The extra 3-bp sequence was identical to a direct repeat which was probably generated upon insertion. Moreover, we detected the Shiga toxin 2 gene with a precise excision of IS1203v. In the wild-type situation, the transposition products of IS1203v could be observed by PCR amplification. These results show that IS1203v can transpose in a nonreplicative manner and that the Shiga toxin gene inactivated by this insertion sequence can revert to the wild type. PMID- 10698783 TI - Selected bacterial strains protect Artemia spp. from the pathogenic effects of Vibrio proteolyticus CW8T2. AB - In this study Vibrio proteolyticus CW8T2 has been identified as a virulent pathogen for Artemia spp. Its infection route has been visualized with transmission electron microscopy. The pathogen affected microvilli and gut epithelial cells, disrupted epithelial cell junctions, and reached the body cavity, where it devastated cells and tissues. In vivo antagonism tests showed that preemptive colonization of the culture water with nine selected bacterial strains protected Artemia juveniles against the pathogenic effects. Two categories of the selected strains could be distinguished: (i) strains providing total protection, as no mortality occurred 2 days after the experimental infection with V. proteolyticus CW8T2, with strain LVS8 as a representative, and (ii) strains providing partial protection, as significant but not total mortality was observed, with strain LVS2 as a representative. The growth of V. proteolyticus CW8T2 in the culture medium was slowed down in the presence of strains LVS2 and LVS8, but growth suppression was distinctly higher with LVS8 than with LVS2. It was striking that the strains that gave only partial protection against the pathogen in the in vivo antagonism test showed also a restricted capability to colonize the Artemia compared to the strains providing total protection. The in vivo antagonism tests and the filtrate experiments showed that probably no extracellular bacterial compounds were involved in the protective action but that the living cells were required to protect Artemia against V. proteolyticus CW8T2. PMID- 10698784 TI - Transcriptional analysis of the tutE tutFDGH gene cluster from Thauera aromatica strain T1. AB - The denitrifying strain T1, identified as Thauera aromatica, is able to grow with toluene serving as its sole carbon source. Previous work identified two genes, tutD and tutE, that are involved in toluene metabolism. Two small open reading frames, tutF and tutG, which may also play a role in toluene metabolism, were also identified. The present work examines the transcriptional organization and regulation of these toluene utilization genes. Northern analysis indicates that the four genes are organized into two operons, tutE and tutFDG, and that both operons are regulated in response to toluene. Primer extension analysis has identified major transcriptional start sites located 177 bp upstream of the tutE translational start and 76 bp upstream of the tutF translational start. Furthermore, a fifth gene, tutH, has been identified immediately downstream of tutG. It is transcribed from the same start site as tutFDG and is predicted to code for a 286-amino-acid protein with a calculated molecular mass of about 31,800 Da. The TutH protein is predicted to have an ATP/GTP binding domain and is similar to the NorQ/NirQ family of proteins. PMID- 10698785 TI - Antagonistic activity of Lactobacillus acidophilus LB against intracellular Salmonella enterica serovar Typhimurium infecting human enterocyte-like Caco-2/TC 7 cells. AB - To gain further insight into the mechanism by which lactobacilli develop antimicrobial activity, we have examined how Lactobacillus acidophilus LB inhibits the promoted cellular injuries and intracellular lifestyle of Salmonella enterica serovar Typhimurium SL1344 infecting the cultured, fully differentiated human intestinal cell line Caco-2/TC-7. We showed that the spent culture supernatant of strain LB (LB-SCS) decreases the number of apical serovar Typhimurium-induced F-actin rearrangements in infected cells. LB-SCS treatment efficiently decreased transcellular passage of S. enterica serovar Typhimurium. Moreover, LB-SCS treatment inhibited intracellular growth of serovar Typhimurium, since treated intracellular bacteria displayed a small, rounded morphology resembling that of resting bacteria. We also showed that LB-SCS treatment inhibits adhesion-dependent serovar Typhimurium-induced interleukin-8 production. PMID- 10698786 TI - Quantitative use of fluorescent in situ hybridization to examine relationships between mycolic acid-containing actinomycetes and foaming in activated sludge plants. AB - The formation of viscous foams on aeration basins and secondary clarifiers of activated sludge plants is a common and widespread problem. Foam formation is often attributed to the presence of mycolic acid-containing actinomycetes (mycolata). In order to examine the relationship between the number of mycolata and foam, we developed a group-specific probe targeting the 16S rRNA of the mycolata, a protocol to permeabilize mycolata, and a statistically robust quantification method. Statistical analyses showed that a lipase-based permeabilization method was quantitatively superior to previously described methods (P << 0.05). When mixed liquor and foam samples were examined, most of the mycolata present were rods or cocci, although filamentous mycolata were also observed. A nested analysis of variance showed that virtually all of the measured variance occurred between fields of view and not between samples. On this basis we determined that as few as five fields of view could be used to give a statistically meaningful sample. Quantitative fluorescent in situ hybridization (FISH) was used to examine the relationship between foaming and the concentration of mycolata in a 20-m(3) completely mixed activated sludge plant. Foaming occurred when the number of mycolata exceeded a certain threshold value. Baffling of the plant affected foaming without affecting the number of mycolata. We tentatively estimated that the threshold foaming concentration of mycolata was about 2 x 10(6) cells ml(-1) or 4 x 10(12) cells m(-2). We concluded that quantitative use of FISH is feasible and that quantification is a prerequisite for rational investigation of foaming in activated sludge. PMID- 10698787 TI - Quantification of Hyphomicrobium populations in activated sludge from an industrial wastewater treatment system as determined by 16S rRNA analysis. AB - The bacterial community structure of the activated sludge from a 25 million-gal per-day industrial wastewater treatment plant was investigated using rRNA analysis. 16S ribosomal DNA (rDNA) libraries were created from three sludge samples taken on different dates. Partial rRNA gene sequences were obtained for 46 rDNA clones, and nearly complete 16S rRNA sequences were obtained for 18 clones. Seventeen of these clones were members of the beta subdivision, and their sequences showed high homology to sequences of known bacterial species as well as published 16S rDNA sequences from other activated sludge sources. Sixteen clones belonged to the alpha subdivision, 7 of which showed similarity to Hyphomicrobium species. This cluster was chosen for further studies due to earlier work on Hyphomicrobium sp. strain M3 isolated from this treatment plant. A nearly full length 16S rDNA sequence was obtained from Hyphomicrobium sp. strain M3. Phylogenetic analysis revealed that Hyphomicrobium sp. strain M3 was 99% similar to Hyphomicrobium denitrificans DSM 1869(T) in Hyphomicrobium cluster II. Three of the cloned sequences from the activated sludge samples also grouped with those of Hyphomicrobium cluster II, with a 96% sequence similarity to that of Hyphomicrobium sp. strain M3. The other four cloned sequences from the activated sludge sample were more closely related to those of the Hyphomicrobium cluster I organisms (95 to 97% similarity). Whole-cell fluorescence hybridization of microorganisms in the activated sludge with genus-specific Hyphomicrobium probe S G-Hypho-1241-a-A-19 enhanced the visualization of Hyphomicrobium and revealed that Hyphomicrobium appears to be abundant both on the outside of flocs and within the floc structure. Dot blot hybridization of activated sludge samples from 1995 with probes designed for Hyphomicrobium cluster I and Hyphomicrobium cluster II indicated that Hyphomicrobium cluster II-positive 16S rRNA dominated over Hyphomicrobium cluster I-positive 16S rRNA by 3- to 12-fold. Hyphomicrobium 16S rRNA comprised approximately 5% of the 16S rRNA in the activated sludge. PMID- 10698789 TI - Survival and epiphytic fitness of a nonpathogenic mutant of Xanthomonas campestris pv. glycines. AB - Xanthomonas campestris pv. glycines is the causal agent of bacterial pustule disease of soybeans. The objective of this work was to construct a nonpathogenic mutant derived from the pathogenic wild-type strain YR32 and to evaluate its effectiveness in preventing growth of its parent on the soybean phyllosphere. A mini-Tn5-derived transposon was used to generate nonpathogenic mutants. Southern hybridization and pulsed-field gel electrophoresis confirmed the presence of a single transposon in each of the nonpathogenic mutants. One of the nonpathogenic mutants, M715, failed to induce a hypersensitive response in tomato leaves. An ice nucleation gene (inaZ) carried in pJL1703 was introduced into strain YR32 as a reporter gene to demonstrate that the presence of M715 could reduce colonization of the soybean phyllosphere by YR32. de Wit serial replacement analysis showed that M715 competed equally with its wild-type parental strain, YR32. Epiphytic fitness analysis of YR32 in the greenhouse indicated that the population dynamics of strains YR32, YR32(pJL1703), and M715 were similar, although the density of the mutant was slightly less than that of its parent. The M715 mutant was able to survive for 16 days after inoculation on soybean leaves and maintained population densities of approximately 10(4) to 10(5) cells g (fresh weight) of leaf(-1). Therefore, M715 shows promise as an effective biocontrol agent for bacterial pustule disease in soybeans. PMID- 10698788 TI - Identification of polyphosphate-accumulating organisms and design of 16S rRNA directed probes for their detection and quantitation. AB - Laboratory-scale sequencing batch reactors (SBRs) as models for activated sludge processes were used to study enhanced biological phosphorus removal (EBPR) from wastewater. Enrichment for polyphosphate-accumulating organisms (PAOs) was achieved essentially by increasing the phosphorus concentration in the influent to the SBRs. Fluorescence in situ hybridization (FISH) using domain-, division-, and subdivision-level probes was used to assess the proportions of microorganisms in the sludges. The A sludge, a high-performance P-removing sludge containing 15.1% P in the biomass, was comprised of large clusters of polyphosphate containing coccobacilli. By FISH, >80% of the A sludge bacteria were beta-2 Proteobacteria arranged in clusters of coccobacilli, strongly suggesting that this group contains a PAO responsible for EBPR. The second dominant group in the A sludge was the Actinobacteria. Clone libraries of PCR-amplified bacterial 16S rRNA genes from three high-performance P-removing sludges were prepared, and clones belonging to the beta-2 Proteobacteria were fully sequenced. A distinctive group of clones (sharing >/=98% sequence identity) related to Rhodocyclus spp. (94 to 97% identity) and Propionibacter pelophilus (95 to 96% identity) was identified as the most likely candidate PAOs. Three probes specific for the highly related candidate PAO group were designed from the sequence data. All three probes specifically bound to the morphologically distinctive clusters of PAOs in the A sludge, exactly coinciding with the beta-2 Proteobacteria probe. Sequential FISH and polyphosphate staining of EBPR sludges clearly demonstrated that PAO probe-binding cells contained polyphosphate. Subsequent PAO probe analyses of a number of sludges with various P removal capacities indicated a strong positive correlation between P removal from the wastewater as determined by sludge P content and number of PAO probe-binding cells. We conclude therefore that an important group of PAOs in EBPR sludges are bacteria closely related to Rhodocyclus and Propionibacter. PMID- 10698790 TI - Propachlor removal by Pseudomonas strain GCH1 in an immobilized-cell system. AB - A bacterial strain capable of growing on propachlor (2-chloro-N isopropylacetanilide) was isolated from soil by using enrichment and isolation techniques. The strain isolated, designated GCH1, was classified as a member of the genus Pseudomonas. Washed-cell suspensions of strain GCH1 accumulated N isopropylacetanilide, acetanilide, acetamide, and catechol. Pseudomonas strain GCH1 grew on propachlor with a generation time of 4.2 h and a rate of substrate utilization of 1.75 +/- 0.15 micromol h(-1). Gene expression did not require induction but was subject to catabolite expression. Acetanilide was a growth substrate with a yield of 0.56 +/- 0.02 mg of protein micromol(-1). GCH1 strain cells were immobilized by adsorption onto a ceramic support and were used as biocatalysts in an immobilized cell system. Propachlor elimination reached 98%, with a retention time of 3 h and an initial organic load of 0.5 mM propachlor. The viability of immobilized cells increased 34-fold after 120 days of bioreactor operation. PMID- 10698791 TI - Selected chitinase genes in cultured and uncultured marine bacteria in the alpha- and gamma-subclasses of the proteobacteria. AB - PCR primers were patterned after chitinase genes in four gamma-proteobacteria in the families Alteromonadaceae and Enterobacteriaceae (group I chitinases) and used to explore the occurrence and diversity of these chitinase genes in cultured and uncultured marine bacteria. The PCR results from 104 bacterial strains indicated that this type of chitinase gene occurs in two major groups of marine bacteria, alpha- and gamma-proteobacteria, but not the Cytophaga-Flavobacter group. Group I chitinase genes also occur in some viruses infecting arthropods. Phylogenetic analysis indicated that similar group I chitinase genes occur in taxonomically related bacteria. However, the overall phylogeny of chitinase genes did not correspond to the phylogeny of 16S rRNA genes, possibly due to lateral transfer of chitinase genes between groups of bacteria, but other mechanisms, such as gene duplication, cannot be ruled out. Clone libraries of chitinase gene fragments amplified from coastal Pacific Ocean and estuarine Delaware Bay bacterioplankton revealed similarities and differences between cultured and uncultured bacteria. We had hypothesized that cultured and uncultured chitin degrading bacteria would be very different, but in fact, clones having nucleotide sequences identical to those of chitinase genes of cultured alpha-proteobacteria dominated both libraries. The other clones were similar but not identical to genes in cultured gamma-proteobacteria, including vibrios and alteromonads. Our results suggest that a closer examination of chitin degradation by alpha proteobacteria will lead to a better understanding of chitin degradation in the ocean. PMID- 10698792 TI - Reductive cleavage of demeton-S-methyl by Corynebacterium glutamicum in cometabolism on more readily metabolizable substrates. AB - Corynebacterium glutamicum is able to biotransform demeton-S-methyl, an organophosphorus compound, during cometabolism with more readily metabolizable substrates. Among the cosubstrates used, fructose is the growth substrate that is most favorable for demeton-S-methyl biotransformation. The reaction mechanism of demeton-S-methyl biotransformation involves reductive cleavage of an S-C bond, which leads to accumulation of dimethyl thiophosphate in the culture medium. PMID- 10698793 TI - A new Shiga toxin 2 variant (Stx2f) from Escherichia coli isolated from pigeons. AB - We have isolated Shiga toxin (Stx)-producing Escherichia coli (STEC) strains from the feces of feral pigeons which contained a new Stx2 variant gene designated stx(2f). This gene is most similar to sltIIva of patient E. coli O128:B12 isolate H.I.8. Stx2f reacted only weakly with commercial immunoassays. The prevalence of STEC organisms carrying the stx(2f) gene in pigeon droppings was 12.5%. The occurrence of a new Stx2 variant in STEC from pigeons enlarges the pool of Stx2 variants and raises the question whether horizontal gene transfer to E. coli pathogenic to humans may occur. PMID- 10698794 TI - Atypical polyphosphate accumulation by the denitrifying bacterium Paracoccus denitrificans. AB - Polyphosphate accumulation by Paracoccus denitrificans was examined under aerobic, anoxic, and anaerobic conditions. Polyphosphate synthesis by this denitrifier took place with either oxygen or nitrate as the electron acceptor and in the presence of an external carbon source. Cells were capable of poly-beta hydroxybutyrate (PHB) synthesis, but no polyphosphate was produced when PHB-rich cells were incubated under anoxic conditions in the absence of an external carbon source. By comparison of these findings to those with polyphosphate-accumulating organisms thought to be responsible for phosphate removal in activated sludge systems, it is concluded that P. denitrificans is capable of combined phosphate and nitrate removal without the need for alternating anaerobic/aerobic or anaerobic/anoxic switches. Studies on additional denitrifying isolates from a denitrifying fluidized bed reactor suggested that polyphosphate accumulation is widespread among denitrifiers. PMID- 10698795 TI - Inhibitory effects of collagen on the PCR for detection of Clostridium perfringens. AB - It is essential to identify specific food components that inhibit PCR in order to increase the sensitivity of the PCR method for rapid detection of pathogens contaminating a food. We found that collagen, a major component of several foods, inhibited PCR. The inhibitory action of collagen on PCR could be partially reversed by adjusting the concentration of magnesium ion in the reaction mixture and by the use of various DNA extraction methods to remove the collagen from the DNA. Also, the source of thermostable DNA polymerase was affected by the presence of collagen. These results suggest the need to optimize the extraction and assay conditions for rapid detection of enterotoxigenic Clostridium perfringens by PCR with respect to the kind of food being analyzed. PMID- 10698796 TI - Development of a nonantibiotic dominant marker for positively selecting expression plasmids in multivalent Salmonella vaccines. AB - We report the novel application of a herbicide-resistance-based dominant marker for the positive selection of expression plasmids in Salmonella serovar vaccines. The beta-lactamase gene of the plasmid pTETnir15, which expresses fragment C of tetanus toxin (TetC), has been replaced with the bar gene marker. The new plasmid pBAT1 can be positively selected in vitro within Salmonella serovars in the presence of the herbicide DL-phosphinothricin. The expression of TetC remains unaltered, and the Salmonella enterica serovar Typhimurium vaccine strain is stable and immunogenic in vivo. PMID- 10698797 TI - Transient growth requirement in Bacillus subtilis following the cessation of exponential growth. AB - During an investigation of the parameters controlling mutations in Bacillus subtilis we observed that this bacterium exhibits a transient growth requirement for two nonessential amino acids (glutamic acid and isoleucine) during a type of postexponential growth on a minimal medium. PMID- 10698798 TI - Cloning, sequencing, and expression of the pyruvate carboxylase gene in Lactococcus lactis subsp. lactis C2. AB - A functional pyc gene was isolated from Lactococcus lactis subsp. lactis C2 and was found to complement a Pyc defect in L. lactis KB4. The deduced lactococcal Pyc protein was highly homologous to Pyc sequences of other bacteria. The pyc gene was also detected in Lactococcus lactis subsp. cremoris and L. lactis subsp. lactis bv. diacetylactis strains. PMID- 10698799 TI - A flow cytometry method for rapid detection and enumeration of total bacteria in milk. AB - Application of flow cytometry (FCM) to microbial analysis of milk is hampered by the presence of milk proteins and lipid particles. Here we report on the development of a rapid (/= 0.98) between the FCM assay and the more conventional methods of plating and direct microscopic counting was achieved. Raw milk data showed a significant correlation (P < 0.01) and a good agreement (r = 0.91) between FCM and standard plate count methods. The detection limit of the FCM assay was Phe mutation abrogates PTPalpha-Src binding, dephosphorylation of pTyr527 (although not of other substrates), and neoplastic transformation by overexpressed PTPalpha in vivo. We suggest that pTyr789 enables pTyr527 dephosphorylation by a pilot binding with the Src SH2 domain that displaces the intramolecular pTyr527-SH2 binding. Consistent with model predictions, we find that excess SH2 domains can disrupt PTPalpha-Src binding and can block PTPalpha-mediated dephosphorylation and activation in proportion to their affinity for pTyr789. Moreover, we show that, as predicted by the model, catalytically defective PTPalpha has reduced Src binding in vivo. The displacement mechanism provides another potential control point for physiological regulation of Src-family signal transduction pathways. PMID- 10698939 TI - Identification of a pocket in the PDK1 kinase domain that interacts with PIF and the C-terminal residues of PKA. AB - The 3-phosphoinositide-dependent protein kinase-1 (PDK1) phosphorylates and activates a number of protein kinases of the AGC subfamily. The kinase domain of PDK1 interacts with a region of protein kinase C-related kinase-2 (PRK2), termed the PDK1-interacting fragment (PIF), through a hydrophobic motif. Here we identify a hydrophobic pocket in the small lobe of the PDK1 kinase domain, separate from the ATP- and substrate-binding sites, that interacts with PIF. Mutation of residues predicted to form part of this hydrophobic pocket either abolished or significantly diminished the affinity of PDK1 for PIF. PIF increased the rate at which PDK1 phosphorylated a synthetic dodecapeptide (T308tide), corresponding to the sequences surrounding the PDK1 phosphorylation site of PKB. This peptide is a poor substrate for PDK1, but a peptide comprising T308tide fused to the PDK1-binding motif of PIF was a vastly superior substrate for PDK1. Our results suggest that the PIF-binding pocket on the kinase domain of PDK1 acts as a 'docking site', enabling it to interact with and enhance the phosphorylation of its substrates. PMID- 10698940 TI - Differential regulation of gene expression by insulin and IGF-1 receptors correlates with phosphorylation of a single amino acid residue in the forkhead transcription factor FKHR. AB - The transcription factor FKHR is inhibited by phosphorylation in response to insulin and IGF-1 through Akt kinase. Here we show that FKHR phosphorylation in hepatocytes conforms to a hierarchical pattern in which phosphorylation of the Akt site at S(253), in the forkhead DNA binding domain, is a prerequisite for the phosphorylation of two additional potential Akt sites, T(24) and S(316). Using insulin receptor-deficient hepatocytes, we show that T(24) fails to be phosphorylated by IGF-1 receptors, suggesting that this residue is targeted by a kinase specifically activated by insulin receptors. Lack of T(24) phosphorylation is associated with the failure of IGF-1 to induce nuclear export of FKHR, and to inhibit expression of a reporter gene under the transcriptional control of the IGF binding protein-1 insulin response element. We propose that site-specific phosphorylation of FKHR is one of the mechanisms by which insulin and IGF-1 receptors exert different effects on gene expression. PMID- 10698941 TI - RNA recognition by a Staufen double-stranded RNA-binding domain. AB - The double-stranded RNA-binding domain (dsRBD) is a common RNA-binding motif found in many proteins involved in RNA maturation and localization. To determine how this domain recognizes RNA, we have studied the third dsRBD from Drosophila Staufen. The domain binds optimally to RNA stem-loops containing 12 uninterrupted base pairs, and we have identified the amino acids required for this interaction. By mutating these residues in a staufen transgene, we show that the RNA-binding activity of dsRBD3 is required in vivo for Staufen-dependent localization of bicoid and oskar mRNAs. Using high-resolution NMR, we have determined the structure of the complex between dsRBD3 and an RNA stem-loop. The dsRBD recognizes the shape of A-form dsRNA through interactions between conserved residues within loop 2 and the minor groove, and between loop 4 and the phosphodiester backbone across the adjacent major groove. In addition, helix alpha1 interacts with the single-stranded loop that caps the RNA helix. Interactions between helix alpha1 and single-stranded RNA may be important determinants of the specificity of dsRBD proteins. PMID- 10698942 TI - Dishevelled phosphorylation, subcellular localization and multimerization regulate its role in early embryogenesis. AB - Dishevelled (Dsh) induces a secondary axis and can translocate to the membrane when activated by Frizzleds; however, dominant-negative approaches have not supported a role for Dsh in primary axis formation. We demonstrate that the Dsh protein is post-translationally modified at the dorsal side of the embryo: timing and position of this regulation suggests a role of Dsh in dorsal-ventral patterning in Xenopus. To create functional links between these properties of Dsh we analyzed the influence of endogenous Frizzleds and the Dsh domain dependency for these characteristics. Xenopus Frizzleds phosphorylate and translocate Xdsh to the membrane irrespective of their differential ectopic axes inducing abilities, showing that translocation is insufficient for axis induction. Dsh deletion analysis revealed that axis inducing abilities did not segregate with Xdsh membrane association. The DIX region and a short stretch at the N-terminus of the DEP domain are necessary for axis induction while the DEP region is required for Dsh membrane association and its phosphorylation. In addition, Dsh forms homomeric complexes in embryos suggesting that multimerization is important for its proper function. PMID- 10698943 TI - Hormone activation induces nucleosome positioning in vivo. AB - The mouse mammary tumor virus (MMTV) promoter is induced by glucocorticoid hormone. A robust hormone- and receptor-dependent activation could be reproduced in Xenopus laevis oocytes. The homogeneous response in this system allowed a detailed analysis of the transition in chromatin structure following hormone activation. This revealed two novel findings: hormone activation led to the establishment of specific translational positioning of nucleosomes despite the lack of significant positioning in the inactive state; and, in the active promoter, a subnucleosomal particle encompassing the glucocorticoid receptor (GR) binding region was detected. The presence of only a single GR-binding site was sufficient for the structural transition to occur. Both basal promoter elements and ongoing transcription were dispensable. These data reveal a stepwise process in the transcriptional activation by glucocorticoid hormone. PMID- 10698944 TI - Loss of FBP function arrests cellular proliferation and extinguishes c-myc expression. AB - The c-myc regulatory region includes binding sites for a large set of transcription factors. The present studies demonstrate that in the absence of FBP [far upstream element (FUSE)-binding protein], which binds to the single-stranded FUSE, the remainder of the set fails to sustain endogenous c-myc expression. A dominant-negative FBP DNA-binding domain lacking effector activity or an antisense FBP RNA, expressed via replication-defective adenovirus vectors, arrested cellular proliferation and extinguished native c-myc transcription from the P1 and P2 promoters. The dominant-negative FBP initially augmented the single stranded character of FUSE; however, once c-myc expression was abolished, melting at FUSE could no longer be supported. In contrast, with antisense FBP RNA, the single-stranded character of FUSE decreased monotonically as the transcription of endogenous c-myc declined. Because transcription is the major source of super coiling in vivo, we propose that by binding torsionally strained DNA, FBP measures promoter activity directly. We also show that FUSE is predicted to behave as a torsion-regulated switch poised to regulate c-myc and to confer a higher order regulation on a large repertoire of factors. PMID- 10698945 TI - Structure of the RXR-RAR DNA-binding complex on the retinoic acid response element DR1. AB - The 9-cis retinoic acid receptor (retinoid X receptor, RXR) forms heterodimers with the all-trans retinoic acid receptor (RAR) and other nuclear receptors on DNA regulatory sites composed of tandem binding elements. We describe the 1.70 A resolution structure of the ternary complex of RXR and RAR DNA-binding regions in complex with the retinoic acid response element DR1. The receptors recognize identical half-sites through extensive base-specific contacts; however, RXR binds exclusively to the 3' site to form an asymmetric complex with the reverse polarity of other RXR heterodimers. The subunits associate in a strictly DNA dependent manner using the T-box of RXR and the Zn-II region of RAR, both of which are reshaped in forming the complex. The protein-DNA contacts, the dimerization interface and the DNA curvature in the RXR-RAR complex are distinct from those of the RXR homodimer, which also binds DR1. Together, these structures illustrate how the nuclear receptor superfamily exploits conformational flexibility and locally induced structures to generate combinatorial transcription factors. PMID- 10698946 TI - Transcription-dependent R-loop formation at mammalian class switch sequences. AB - Immunoglobulin class switching is mediated by recombination between switch sequences located immediately upstream of the immunoglobulin constant heavy chain genes. Targeting of recombination to particular switch sequences is associated temporally with transcription through these regions. We recently have provided evidence for inducible and stable RNA-DNA hybrid formation at switch sequences in the mouse genome that are mechanistically important for class switching in vivo. Here, we define in vitro the precise configuration of the DNA and RNA strands within this hybrid structure at the Smicro, Sgamma3 and Sgamma2b mouse switch sequences. We find that the G-rich (non-template) DNA strand of each switch sequence is hypersensitive to probes throughout much of its length, while the C rich (template) DNA strand is essentially resistant. These results demonstrate formation of an R-loop, whereby the G-rich RNA strand forms a stable heteroduplex with its C-rich DNA strand counterpart, and the G-rich DNA strand exists primarily in a single-stranded state. We propose that the organized structure of the R-loop is essential for targeting the class switch recombination machinery to these sequences. PMID- 10698947 TI - Intimate adhesion of Neisseria meningitidis to human epithelial cells is under the control of the crgA gene, a novel LysR-type transcriptional regulator. AB - PilC1, a pilus-associated protein in Neisseria menin- gitidis, is a key element in initial meningococcal adhesion to target cells. A promoter element (CREN, contact regulatory element of Neisseria) is responsible for the transient induction of this gene upon cell contact. crgA (contact-regulated gene A) encodes a transcriptional regulator whose expression is also induced upon cell contact from a promoter region similar to the CREN of pilC1. CrgA shows significant sequence homologies to LysR-type transcriptional regulators. Its inactivation in meningococci provokes a dramatic reduction in bacterial adhesion to epithelial cells. Moreover, this mutant is unable to undergo intimate adhesion to epithelial cells or to provoke effacing of microvilli on infected cells. Purified CrgA is able to bind to pilC1 and crgA promoters, and CrgA seems to repress the expression of pilC1 and crgA. Our results support a dynamic model of bacteria cell interaction involving a network of regulators acting in cascade. CrgA could be an intermediate regulator in such a network. PMID- 10698948 TI - Cap-dependent deadenylation of mRNA. AB - Poly(A) tail removal is often the initial and rate-limiting step in mRNA decay and is also responsible for translational silencing of maternal mRNAs during oocyte maturation and early development. Here we report that deadenylation in HeLa cell extracts and by a purified mammalian poly(A)-specific exoribonuclease, PARN (previously designated deadenylating nuclease, DAN), is stimulated by the presence of an m(7)-guanosine cap on substrate RNAs. Known cap-binding proteins, such as eIF4E and the nuclear cap-binding complex, are not detectable in the enzyme preparation, and PARN itself binds to m(7)GTP-Sepharose and is eluted specifically with the cap analog m(7)GTP. Xenopus PARN is known to catalyze mRNA deadenylation during oocyte maturation. The enzyme is depleted from oocyte extract with m(7)GTP-Sepharose, can be photocross-linked to the m(7)GpppG cap and deadenylates m(7)GpppG-capped RNAs more efficiently than ApppG-capped RNAs both in vitro and in vivo. These data provide additional evidence that PARN is responsible for deadenylation during oocyte maturation and suggest that interactions between 5' cap and 3' poly(A) tail may integrate translational efficiency with mRNA stability. PMID- 10698949 TI - Functional interaction between RAFT1/FRAP/mTOR and protein kinase cdelta in the regulation of cap-dependent initiation of translation. AB - Hormones and growth factors induce protein translation in part by phosphorylation of the eukaryotic initiation factor 4E (eIF4E) binding protein 1 (4E-BP1). The rapamycin and FK506-binding protein (FKBP)-target 1 (RAFT1, also known as FRAP) is a mammalian homolog of the Saccharomyces cerevisiae target of rapamycin proteins (mTOR) that regulates 4E-BP1. However, the molecular mechanisms involved in growth factor-initiated phosphorylation of 4E-BP1 are not well understood. Here we demonstrate that protein kinase Cdelta (PKCdelta) associates with RAFT1 and that PKCdelta is required for the phosphorylation and inactivation of 4E-BP1. PKCdelta-mediated phosphorylation of 4E-BP1 is wortmannin resistant but rapamycin sensitive. As shown for serum, phosphorylation of 4E-BP1 by PKCdelta inhibits the interaction between 4E-BP1 and eIF4E and stimulates cap-dependent translation. Moreover, a dominant-negative mutant of PKCdelta inhibits serum-induced phosphorylation of 4E-BP1. These findings demonstrate that PKCdelta associates with RAFT1 and thereby regulates phosphorylation of 4E-BP1 and cap-dependent initiation of protein translation. PMID- 10698950 TI - Charged tmRNA but not tmRNA-mediated proteolysis is essential for Neisseria gonorrhoeae viability. AB - tmRNA, through its tRNA and mRNA properties, adds short peptide tags to abnormal proteins, targeting these proteins for proteolytic degradation. Although the conservation of tmRNA throughout the bacterial kingdom suggests that it must provide a strong selective advantage, it has not been shown to be essential for any bacterium. We report that tmRNA is essential in Neisseria gonorrhoeae. Although tagging per se appears to be required for gonococcal viability, tagging for proteolysis does not. This suggests that the essential roles of tmRNA in N.gonorrhoeae may include resolving stalled translation complexes and/or preventing depletion of free ribosomes. Although derivatives of N.gonorrhoeae expressing Escherichia coli tmRNA as their sole tmRNA were isolated, they appear to form colonies only after acquiring an extragenic suppressor(s). PMID- 10698951 TI - Essential interaction between the fission yeast DNA polymerase delta subunit Cdc27 and Pcn1 (PCNA) mediated through a C-terminal p21(Cip1)-like PCNA binding motif. AB - Direct interaction between DNA polymerase delta and its processivity factor proliferating cell nuclear antigen (PCNA) is essential for effective replication of the eukaryotic genome, yet the precise manner by which this occurs is unclear. We show that the 54 kDa subunit of DNA polymerase delta from Schizosaccharomyces pombe interacts directly with Pcn1 (PCNA) both in vivo and in vitro. Binding is effected via a short sequence at the C-terminus of Cdc27 with significant similarity to the canonical PCNA binding motif first identified in the mammalian p21(Cip1) protein. This motif is both necessary and sufficient for binding of Pcn1 by Cdc27 in vitro and is essential for Cdc27 function in vivo. We also show that the Pcn1 binding motif in Cdc27 is distinct from its binding site for Cdc1, the 55 kDa B-subunit of polymerase delta, and present evidence that Cdc27 can bind to Pcn1 and Cdc1 simultaneously. Finally, we show that Cdc27 performs at least two distinct essential functions, one of which is independent of Pcn1 binding. PMID- 10698952 TI - Crystal structure of NAD(+)-dependent DNA ligase: modular architecture and functional implications. AB - DNA ligases catalyze the crucial step of joining the breaks in duplex DNA during DNA replication, repair and recombination, utilizing either ATP or NAD(+) as a cofactor. Despite the difference in cofactor specificity and limited overall sequence similarity, the two classes of DNA ligase share basically the same catalytic mechanism. In this study, the crystal structure of an NAD(+)-dependent DNA ligase from Thermus filiformis, a 667 residue multidomain protein, has been determined by the multiwavelength anomalous diffraction (MAD) method. It reveals highly modular architecture and a unique circular arrangement of its four distinct domains. It also provides clues for protein flexibility and DNA-binding sites. A model for the multidomain ligase action involving large conformational changes is proposed. PMID- 10698953 TI - Escherichia coli promoter opening and -10 recognition: mutational analysis of sigma70. AB - The opening of specific segments of DNA is required for most types of genetic readout, including sigma70-dependent transcription. To learn how this occurs, a series of single point mutations were introduced into sigma70 region 2. These were assayed for duplex DNA binding, DNA opening and DNA double strand-single strand fork junction binding. Band shift assays for closed complex formation implicated a series of arginine and aromatic residues within a minimal 26 amino acid region. Permanganate assays implicated two additional aromatic residues in DNA opening, known to form a parallel stack of the type that can accept a flipped out base. Substitution for either of these aromatics had no effect on duplex probe recognition. However, when a single unpaired -11 nucleotide is added to the probe, the mutants fail to bind appropriately to give heparin resistance. A model for DNA opening is presented in which duplex recognition by regions 2.3, 2.4 and 2.5 of sigma positions the pair of aromatic amino acids, which then create the fork junction required for stable opening. PMID- 10698954 TI - Cell cycle regulator phosphorylation stimulates two distinct modes of binding at a chromosome replication origin. AB - In Caulobacter crescentus, the global response regulator CtrA controls chromosome replication and determines the fate of two different cell progenies. Previous studies proposed that CtrA represses replication by binding to five sites, designated [a-e], in the replication origin. We show that phosphorylated CtrA binds sites [a-e] with 35- to 100-fold lower K(d) values than unphosphorylated CtrA. CtrA phosphorylation stimulates two distinct modes of binding to the replication origin. Phosphorylation stimulates weak intrinsic protein-protein cooperation between half-sites and does not stimulate CtrA-P binding unless protein-DNA contacts are made at both half-sites. CtrA phosphorylation also stimulates cooperative binding between complete sites [a] and [b]. However, binding to each of the other CtrA-binding sites [c], [d] and [e] is completely independent and suggests a modular organization of replication control by CtrA. We therefore propose a model where a phosphorelay targets separate biochemical activities inside the replication origin through both cooperative and independent CtrA-binding sites. PMID- 10698955 TI - Tailed duplex DNA is the preferred substrate for Rad51 protein-mediated homologous pairing. AB - The repair of potentially lethal DNA double-stranded breaks (DSBs) by homologous recombination requires processing of the broken DNA into a resected DNA duplex with a protruding 3'-single-stranded DNA (ssDNA) tail. Accordingly, the canonical models for DSB repair require invasion of an intact homologous DNA template by the 3'-end of the ssDNA, a characteristic that the bacterial pairing protein RecA possesses. Unexpectedly, we find that for the eukaryotic homolog, Rad51 protein, the 5'-end of ssDNA is more invasive than the 3'-end. This pairing bias is unaffected by Rad52, Rad54 or Rad55-57 proteins. However, further investigation reveals that, in contrast to RecA protein, the preferred DNA substrate for Rad51 protein is not ssDNA but rather dsDNA with ssDNA tails. This important distinction permits the Rad51 proteins to promote DNA strand invasion using either 3'- or 5'-ends with similar efficiency. PMID- 10698956 TI - UV damage causes uncontrolled DNA breakage in cells from patients with combined features of XP-D and Cockayne syndrome. AB - Nucleotide excision repair (NER) removes damage from DNA in a tightly regulated multiprotein process. Defects in NER result in three different human disorders, xeroderma pigmentosum (XP), trichothiodystrophy (TTD) and Cockayne syndrome (CS). Two cases with the combined features of XP and CS have been assigned to the XP-D complementation group. Despite their extreme UV sensitivity, these cells appeared to incise their DNA as efficiently as normal cells in response to UV damage. These incisions were, however, uncoupled from the rest of the repair process. Using cell-free extracts, we were unable to detect any incision activity in the neighbourhood of the damage. When irradiated plasmids were introduced into unirradiated XP-D/CS cells, the ectopically introduced damage triggered the induction of breaks in the undamaged genomic DNA. XP-D/CS cells thus have a unique response to sensing UV damage, which results in the introduction of breaks into the DNA at sites distant from the damage. We propose that it is these spurious breaks that are responsible for the extreme UV sensitivity of these cells. PMID- 10698957 TI - Compositional bias and mimicry toward the nonself proteome in immunodominant T cell epitopes of self and nonself antigens. AB - We investigated whether and how molecular mimicry affects the shaping of the helper T cell repertoire. We implemented an algorithm that measures the probability of mimicry between epitopes of known immunogenicity and self or nonself proteomes. This algorithm yields 'similarity profiles', which represent the probability of matching between all contiguous overlapping peptides of the antigen under examination and those in the proteome(s) considered. Similarity profiles between helper T cell epitopes (of self or microbial antigens and allergens) and human or microbial SWISSPROT collections were produced. For each antigen, both collections yielded largely overlapping profiles, demonstrating that self-nonself discrimination does not rely on qualitative features that distinguish human from microbial peptides. However, epitopes whose probability of mimicry with self or nonself prevails are, respectively, tolerated or immunodominant and coexist within the same (auto-)antigen regardless of its self/nonself nature. Epitopes (on self and nonself antigens) can cross-stimulate T cells at increasing potency as their similarity with nonself augments. Mimicry, rather than complicating self-nonself discrimination, assists in the shaping of the immune repertoire and helps define the defensive or autoreactive potential of a T cell. Being a predictor of epitope immunogenicity, it bears relevance to vaccine design. PMID- 10698958 TI - Amelioration of accelerated diabetic mesangial expansion by treatment with a PKC beta inhibitor in diabetic db/db mice, a rodent model for type 2 diabetes. AB - Activation of protein kinase C (PKC) is implicated as an important mechanism by which diabetes causes vascular complications. We have recently shown that a PKC beta inhibitor ameliorates not only early diabetes-induced glomerular dysfunction such as glomerular hyperfiltration and albuminuria, but also overexpression of glomerular mRNA for transforming growth factor beta1 (TGF-beta1) and extracellular matrix (ECM) proteins in streptozotocin-induced diabetic rats, a model for type 1 diabetes. In this study, we examined the long-term effects of a PKC beta inhibitor on glomerular histology as well as on biochemical and functional abnormalities in glomeruli of db/db mice, a model for type 2 diabetes. Administration of a PKC beta inhibitor reduced urinary albumin excretion rates and inhibited glomerular PKC activation in diabetic db/db mice. Administration of a PKC beta inhibitor also prevented the mesangial expansion observed in diabetic db/db mice, possibly through attenuation of glomerular expression of TGF-beta and ECM proteins such as fibronectin and type IV collagen. These findings provide the first in vivo evidence that the long-term inhibition of PKC activation in the renal glomeruli can ameliorate glomerular pathologies in diabetic state, and thus suggest that a PKC beta inhibitor might be an useful therapeutic strategy for the treatment of diabetic nephropathy. PMID- 10698959 TI - Radiolabeling revisited: metabolic labeling with (35)S-methionine inhibits cell cycle progression, proliferation, and survival. AB - Metabolic labeling of cells with low-energy beta-emitting radioisotopes such as [(35)S]methionine is often used to follow the biosynthesis, maturation, and degradation of proteins in vivo. Such techniques have generally been assumed to be relatively nonperturbing to the cell. The results presented here indicate that metabolic labeling of cells with [(35)S]methionine under standard experimental conditions can inhibit cell progression into mitosis, cause cell cycle arrest, inhibit cell proliferation in both short-term and colony-forming assays, alter cell morphology, and induce apoptosis over the course of several days. These results thus suggest the need for caution in interpretation of studies using such methods, especially if the experiments rely on the normal progression of the cell cycle or are intended to monitor events occurring in a normally proliferating cell. PMID- 10698960 TI - Diazoxide down-regulates leptin and lipid metabolizing enzymes in adipose tissue of Zucker rats. AB - We have previously reported that attenuation of hyperinsulinemia by diazoxide (DZ), an inhibitor of glucose-mediated insulin secretion, increased insulin sensitivity and reduced body weight in obese Zucker rats. These findings prompted us to investigate the effects of DZ on key insulin-sensitive enzymes regulating adipose tissue metabolism, fatty acid synthase (FAS), and lipoprotein lipase (LPL), as well as on circulating levels of leptin. We also determined the direct effects of diazoxide on FAS in 3T3-L1 adipocytes. Seven-week-old female obese and lean Zucker rats were treated with DZ (150 mg/kg/d) or vehicle (C, control) for a period of 6 wk. Changes in plasma parameters by DZ include significant decreases in triglycerides, free fatty acids, glucose, and insulin, consistent with our previous reports. DZ obese rats exhibited lower plasma leptin levels (P<0.03) compared to their C animals. DZ significantly reduced adipose tissue FAS activity in both lean (P<0.0001) and obese (P<0.01) animals. LPL mRNA content was also decreased significantly in DZ-treated obese animals (P<0.009) as compared to their respective controls without a significant effect on lean animals. The possibility that DZ exerted a direct effect on adipocytes was further tested in cultured 3T3-L1 adipocytes. Although diazoxide (5 microM) alone did not change FAS activity in cultured 3T3-L1 adipocytes, it significantly attenuated insulin's effect on FAS activity (P<0.001). We demonstrate that DZ regulates key insulin sensitive enzymes involved in regulation of adipose tissue metabolism. These findings suggest that modification of insulin-sensitive pathways can be therapeutically beneficial in obesity management. PMID- 10698961 TI - Reduced amount of the glucose-regulated protein GRP94 in skeletal myoblasts results in loss of fusion competence. AB - We previously showed that skeletal myocytes of the adult rabbit do not accumulate the endoplasmic reticulum glucose-regulated protein GRP94, neither constitutively nor inducibly, at variance with skeletal myocytes during perinatal development (5). Here we show that C2C12 cells up-regulate GRP94 during differentiation and, similarly to primary cultures of murine skeletal myocytes, specifically display GRP94 immunoreactivity on the cell surface. Stable transfection of C2C12 cells with grp94 antisense cDNA shows lack of myotube formation in clones displaying >40% reduction in GRP94 amount. The same result is obtained after in vivo injection of grp94-antisense myoblasts. Conversely, GRP94 overexpression is accompanied by accelerated myotube formation. Analyses of BrdU incorporation, p21 nuclear translocation, and muscle-gene expression show that muscle differentiation is not apparently affected in grp94-antisense clones. In contrast, cell-surface GRP94 is greatly reduced in grp94-antisense clones, as shown by immunocytochemistry and precipitation of cell-surface biotinylated proteins. Thus, cell-surface expression of GRP94 is necessary for maintenance of fusion competence. Furthermore, differentiating C2C12 cells grown in the presence of anti-GRP94 antibody show decreased myotube number suggesting that cell-surface GRP94 is directly involved in myoblast fusion process. PMID- 10698962 TI - Glycine-gated chloride channels in neutrophils attenuate calcium influx and superoxide production. AB - Recently, it was demonstrated that liver injury and TNF-alpha production as a result of endotoxin (lipopolysaccharide, LPS) were attenuated by feeding animals a diet enriched with glycine. This phenomenon was shown to be a result of, at least in part, activation of a chloride channel in Kupffer cells by glycine, which hyperpolarizes the cell membrane and blunts increases in intracellular calcium concentrations ([Ca(2+)](i)) similar to its action in the neuron. It is well known that hepatotoxicity due to LPS has a neutrophil-mediated component and that activation of neutrophils is dependent on increases in [Ca(2+)](i). Therefore, the purpose of this study was to determine if glycine affected agonist induced increases in [Ca(2+)](i) in rat neutrophils. The effect of glycine on increases in [Ca(2+)](i) elicited either by the bacterial-derived peptide formyl methionine-leucine-phenylalanine (FMLP) or LPS was studied in individual neutrophils using Fura-2 and fluorescence microscopy. Both FMLP and LPS caused dose-dependent increases in [Ca(2+)](i), which were maximal at 1 microM FMLP and 100 microgram/ml LPS, respectively. LPS increased intracellular calcium in the presence and absence of extracellular calcium. Glycine blunted increases in [Ca(2+)](i) in a dose-dependent manner with an IC(50) of approximately 0.3 mM, values only slightly higher than plasma levels. Glycine was unable to prevent agonist-induced increases in [Ca(2+)](i) in chloride-free buffer. Moreover, strychnine (1 microM), an antagonist of the glycine-gated chloride channel in the central nervous system, reversed the effects of glycine (1 mM) on FMLP- or LPS stimulated increases in [Ca(2+)](i). To provide hard evidence for a glycine-gated chloride channel in the neutrophil, the effect of glycine on radioactive chloride uptake was determined. Glycine caused a dose-dependent increase in chloride uptake into neutrophils with an ED(50) of approximately 0.4 mM, an effect also prevented by 1 microM strychnine. Glycine also significantly reduced the production of superoxide anion from FMLP-stimulated neutrophils. Taken together, these data provide clear evidence that neutrophils contain a glycine-gated chloride channel that can attenuate increases in [Ca(2+)](i) and diminish oxidant production by this important leukocyte. PMID- 10698963 TI - Human mutations in glucose 6-phosphate dehydrogenase reflect evolutionary history. AB - Glucose 6-phosphate dehydrogenase (G6PD) is a cytosolic enzyme encoded by a housekeeping X-linked gene whose main function is to produce NADPH, a key electron donor in the defense against oxidizing agents and in reductive biosynthetic reactions. Inherited G6PD deficiency is associated with either episodic hemolytic anemia (triggered by fava beans or other agents) or life-long hemolytic anemia. We show here that an evolutionary analysis is a key to understanding the biology of a housekeeping gene. From the alignment of the amino acid (aa) sequence of 52 glucose 6-phosphate dehydrogenase (G6PD) species from 42 different organisms, we found a striking correlation between the aa replacements that cause G6PD deficiency in humans and the sequence conservation of G6PD: two thirds of such replacements are in highly and moderately conserved (50-99%) aa; relatively few are in fully conserved aa (where they might be lethal) or in poorly conserved aa, where presumably they simply would not cause G6PD deficiency. This is consistent with the notion that all human mutants have residual enzyme activity and that null mutations are lethal at some stage of development. Comparing the distribution of mutations in a human housekeeping gene with evolutionary conservation is a useful tool for pinpointing amino acid residues important for the stability or the function of the corresponding protein. In view of the current explosive increase in full genome sequencing projects, this tool will become rapidly available for numerous other genes. PMID- 10698964 TI - Effects of fatty acids on uncoupling protein-2 expression in the rat heart. AB - Fatty acids are thought to play a role in the activity of uncoupling proteins (UCP) and have been shown to regulate the expression of genes encoding proteins involved in fatty acid handling. Therefore, we investigated whether fatty acids, which are the main substrates for the heart, affect rat cardiac UCP-2 expression in vivo and in vitro. After birth, when the contribution of fatty acid oxidation to cardiac energy conversion increases, UCP-2 expression enhanced rapidly. In the adult heart, however, UCP-2 mRNA levels did not alter during conditions associated with either enhanced (fasting, diabetes) or decreased (hypertrophy) fatty acid utilization. Exposure of neonatal cardiomyocytes and embryonic rat heart-derived H9c2 cells to fatty acids (palmitic and oleic acid) for 48 h strongly induced UCP-2 expression. Stimulation of neonatal cardiomyocytes with triiodothyronine also increased UCP-2 mRNA levels, though only in the presence of fatty acids. Ligands specific to the fatty acid-activated transcription factor PPARalpha, but not to PPARgamma, acted as inducers of cardiomyocyte UCP-2 expression. It is concluded that fatty acids promote UCP-2 expression in neonatal cardiomyocytes, which might explain the rapid increase in UCP-2 mRNA in the postnatal heart. However, UCP-2 mRNA levels in the adult heart appear to be insensitive to changes in cardiac fatty acid handling under various pathological conditions. PMID- 10698965 TI - Differential regulation of three thyroid hormone-responsive matrix metalloproteinase genes implicates distinct functions during frog embryogenesis. AB - Matrix metalloproteinases (MMPs) are a family of Zn(2+)-dependent extracellular proteases capable of degrading various proteinaceous components of the extracellular matrix (ECM). They are expressed in developmental and pathological processes such as postlactation mammary gland involution and tumor metastasis. Relatively few studies have been carried out to investigate the function of MMPs during embryogenesis and postembryonic organ development. Using Xenopus development as a model system, we and others have previously isolated three MMP genes as thyroid hormone response genes. They have distinct temporal and organ specific regulations during thyroid hormone-dependent metamorphosis. We demonstrate here that three MMPs-stromelysin-3 (ST3), collagenases-3 (Col3), and collagenases-4 (Col4)-also have distinct spatial and temporal expression profiles during embryogenesis. Consistent with earlier suggestions that ST3 is a direct thyroid hormone response gene whereas Col3 and Col4 are not, we show that precocious overexpression of thyroid hormone receptors in the presence of thyroid hormone lead to increased expression of ST3, but not Col3. Furthermore, our whole mount in situ hybridizations reveal a tight but distinct association of individual MMPs with tissue remodeling in different regions of the animal during embryogenesis. These results suggest that ST3 is likely to play a role in ECM remodeling that facilitate apoptotic tissue remodeling or resorption, whereas Col3 and Col4 appear to participate in connective tissue degradation during development. PMID- 10698967 TI - Effect of ABC transporters on HIV-1 infection: inhibition of virus production by the MDR1 transporter. AB - The MDR1 multidrug transporter P-gp (P-glycoprotein) is an efflux pump that extrudes diverse hydrophobic drugs and peptides from cells. Since the entry of HIV-1 into cells involves an initial interaction of the viral gp41 hydrophobic peptide with the plasma membrane, a potential effect of P-gp on HIV-1 infectivity was explored. Virus production was greatly decreased when P-gp was overexpressed at the surface of a continuous CD4(+) human T-leukemic cell line (12D7) infected with HIV-1(NL4-3), a T-tropic molecular clone of HIV-1. P-gp overexpression did not significantly alter the surface expression or distribution of either the HIV 1 receptor CD4 or the coreceptor CXCR4. Reduction of HIV-1 infectivity in P-gp expressing cells occurred both during the fusion of viral and plasma membranes and at subsequent step(s) in the HIV-1 life cycle. PMID- 10698966 TI - P-glycoprotein-overexpressing multidrug-resistant cells are resistant to infection by enveloped viruses that enter via the plasma membrane. AB - The multidrug resistance gene product P-glycoprotein confers drug resistance to tumor cells by acting as a transporter that blocks the entry into the cell of a great variety of drugs and hydrophobic peptides. In this study we find that in drug-resistant cells, the insertion of the influenza virus fusion protein (hemagglutinin-2) into the plasma membrane is blocked and that the fusion of the viral envelope with the plasma membrane of these cells is impaired. Multidrug resistant cells display significant resistance to infection by envelope viruses that invade cells by fusion with the plasma membrane, but not to infection by pH dependent viruses that penetrate cells by fusion with endocytic vesicles. These observations suggest that multidrug resistance phenomena may protect cells from infection by a large group of disease-causing viruses that includes human immunodeficiency virus, herpes simplex virus, and some cancer-inducing retroviruses. PMID- 10698968 TI - Best5: a novel interferon-inducible gene expressed during bone formation. AB - Regulation of bone formation is important in the pathogenesis of many conditions such as osteoporosis, fracture healing, and loosening of orthopedic implants. We have recently identified a novel rat cDNA (best5) by differential display PCR that is regulated during osteoblast differentiation and bone formation in vitro and in vivo. Expression of best5 mRNA is induced in cultures of osteoblasts by both interferon-alpha (IFN-alpha) or IFN-gamma. Whereas IFN-alpha induced a rapid, transient induction of best5 expression peaking at 4-6 h poststimulation, IFN-gamma elicited a more prolonged induction of best5 expression, which remained elevated 48 h poststimulation. A polyclonal antibody generated to a peptide derived from the best5 coding region recognized a 27 kDa protein on Western blot analysis of osteoblast lysates. We localized BEST5 protein in osteoblast progenitor cells and mature osteoblasts in sections of rat tibiae and in sections of bones loaded in vivo to induce adaptive bone formation. Best5 may therefore be a fundamental intermediate in the response of osteoblasts to stimuli that modulate proliferation/differentiation, such as interferons or mechanical loading. These findings highlight the close interactions between the immune system and bone cells and may open new therapeutic avenues in modulating bone mass. PMID- 10698969 TI - Oxidized LDLs alter the activity of the ubiquitin-proteasome pathway: potential role in oxidized LDL-induced apoptosis. AB - Oxidized low-density lipoproteins (oxLDL) play a role in the genesis of atherosclerosis. OxLDL are able to induce apoptosis of vascular cells, which is potentially involved in the formation of the necrotic center of atherosclerotic lesions, plaque rupture, and subsequent thrombotic events. Because oxLDL may induce structural modifications of cell protein and altered proteins may impair cell viability, the present work aimed to evaluate the extent of protein alterations, the degradation of modified proteins through the ubiquitin proteasome system (a major degradative pathway for altered and oxidatively modified proteins) and their role during apoptosis induced by oxLDL. This paper reports the following: 1) oxLDL induce derivatization of cell proteins by 4 hydroxynonenal (4-HNE) and ubiquitination. 2) Toxic concentrations of oxLDL elicit a biphasic effect on proteasome activity. An early and transient activation of endogenous proteolysis is followed rapidly by a subsequent decay (resulting probably from the 26S proteasome inhibition) and followed later by the inhibition of the 20S proteasome (as assessed by inhibition of sLLVY-MCA hydrolysis). 3) Specific inhibitors of proteasome (lactacystin and proteasome inhibitor I) potentiated considerably the toxicity of oxLDL (nontoxic doses of oxLDL became severely toxic). The defect of the ubiquitination pathway (in temperature-sensitive mutants) also potentiated the toxicity of oxLDL. This suggests that the ubiquitin-proteasome pathway plays a role in the cellular defenses against oxLDL-induced toxicity. 4) Dinitrophenylhydrazine (DNPH), an aldehyde reagent, prevented both the oxLDL-induced derivatization of cell proteins and subsequent cytotoxicity. Altogether, the reported data suggest that both derivatization of cell proteins (by 4-HNE and other oxidized lipids) and inhibition of the proteasome pathway are involved in the mechanism of oxLDL induced apoptosis. PMID- 10698970 TI - Specific phosphorylation of Torpedo 43K rapsyn by endogenous kinase(s) with thiamine triphosphate as the phosphate donor. AB - 43K rapsyn is a peripheral protein specifically associated with the nicotinic acetylcholine receptor (nAChR) present in the postsynaptic membrane of the neuromuscular junction and of the electrocyte, and is essential for its clustering. Here, we demonstrate a novel specific phosphorylation of 43K rapsyn by endogenous protein kinase(s) present in Torpedo electrocyte nAChR-rich membranes and identify thiamine triphosphate (TTP) as the phosphate donor. In the presence of Mg(2+) and [gamma-(32)P]-TTP, 43K rapsyn is specifically phosphorylated with a (32)P-half-maximal incorporation at approximately 5-25 microM TTP. The presence of TTP in the cytosol and of 43K rapsyn at the cytoplasmic face of the postsynaptic membrane, together with TTP-dependent phosphorylation of 43K rapsyn without added exokinases, suggests that TTP dependent-43K-rapsyn phosphorylation may occur in vivo. In addition, phosphoamino acid and chemical stability analysis suggests that the residues phosphorylated are predominantly histidines. Inhibition of phosphorylation by Zn(2+) suggests a possible control of 43K rapsyn phosphorylation state by its zinc finger domain. Endogenous kinase(s) present in rodent brain membranes can also use [gamma-(32)P] TTP as a phosphodonor. The use of a phosphodonor (TTP) belonging to the thiamine family but not to the classical (ATP, GTP) purine triphosphate family represents a novel phosphorylation pathway possibly important for synaptic proteins. PMID- 10698971 TI - Endothelial preconditioning by transient oxidative stress reduces inflammatory responses of cultured endothelial cells to TNF-alpha. AB - Brief episodes of ischemia can render an organ resistant to subsequent severe ischemia. This 'ischemic preconditioning' is ascribed to various mechanisms, including oxidative stress. We investigated whether preconditioning exists on an endothelial level. Human umbilical vein endothelial cells (HUVECs) were transiently confronted with oxidative stress (1 mM H(2)O(2), 5 min). Adhesion molecules ICAM-1 and E-selectin and release of cytokines IL-6 and IL-8 to subsequent stimulation with TNF-alpha (2.5 ng/ml, 4 h) were measured (flow cytometry and immunoassay), as were nuclear translocation of the transcription factor NFkappaB (Western blotting, confocal microscopy) and redox status of HUVECs (quantification of glutathione by HPLC). TNF-alpha elevated IL-6 in the cell supernatant from 8.8 +/- 1 to 41 +/- 3 pg/ml and IL-8 from 0.5 +/- 0. 03 to 3 +/- 0.2 ng/ml. ICAM-1 was increased threefold and E-selectin rose eightfold. Oxidative stress (decrease of glutathione by 50%) reduced post-TNF-alpha levels of IL-6 to 14 +/- 3 and IL-8 to 1 +/- 0.2; the rise of ICAM-1 was completely blocked and E-selectin was only doubled. The anti-inflammatory effects of preconditioning via oxidative stress were paralleled by reduction of the translocation of NFkappaB on stimulation with TNF-alpha, and antagonized by the intracellular radical scavenger N-acetylcysteine. 'Anti-inflammatory preconditioning' of endothelial cells by oxidative stress may account for the inhibitory effects of preconditioning on leukocyte adhesion in vivo. PMID- 10698972 TI - Response of keratinocytes from normal and psoriatic epidermis to interferon-gamma differs in the expression of zinc-alpha(2)-glycoprotein and cathepsin D. AB - Psoriasis is a T cell-mediated inflammatory disease characterized by hyperproliferation and by aberrant differentiation. We found cathepsin D and zinc alpha(2)-glycoprotein, two catalytic enzymes associated with apoptosis and desquamation, to be present in the stratum corneum of the normal epidermis but absent from the psoriatic plaque. Psoriasis is characterized by an altered response to interferon-gamma (IFN-gamma), including the induction of apoptosis in normal but not in psoriatic keratinocytes, often with opposite effects on gene expression of suprabasal proteins. We found that IFN-gamma binding and signaling were attenuated in psoriasis: The IFN-gamma receptor, the signal transducer and activator of transcription STAT-1, and the interferon regulatory factor IRF-1 were strongly up-regulated by IFN-gamma in normal keratinocytes, but not in psoriatic ones. IFN-gamma strongly up-regulated the expression of the catalytic enzymes cathepsin D and zinc-alpha(2)-glycoprotein in normal keratinocytes but down-regulated them in psoriatic ones; the reverse was true of the apoptotic suppressor bcl-2. We believe that the aberrant response to IFN-gamma plays a central role in the pathophysiology of psoriasis, particularly the disruption of apoptosis and desquamation. PMID- 10698973 TI - The anti-inflammatory peptides, antiflammins, regulate the expression of adhesion molecules on human leukocytes and prevent neutrophil adhesion to endothelial cells. AB - Antiflammin-1 and antiflammin-2 are nonapeptides corresponding to the region of highest similarity between glucocorticoid-inducible proteins lipocortin-1 and uteroglobin. We have studied whether antiflammins could affect expression of adhesion molecules on human leukocytes and coronary artery endothelial cells (HCAEC) and binding of neutrophils (PMNs) to HCAEC. Although neither antiflammin 1 nor antiflammin-2 affected expression of adhesion molecules on resting PMNs, monocytes, and lymphocytes in whole blood, they attenuated changes in L-selectin and CD11/CD18 expression evoked by platelet-activating factor or interleukin-8 with IC(50) values of 4-20 micromol/l. The maximum inhibition was similar to those seen with human recombinant lipocortin-1 (100 microgram/ml). Unlike dexamethasone (100 nmol/l), the antiflammins had little effect on LPS-stimulated expression of E-selectin and ICAM-1 on HCAEC. Consistently, culture of HCAEC with dexamethasone, but not with antiflammins, decreased PMN binding to endothelial cells. Preincubation of PMNs with antiflammins markedly decreased their adhesion to LPS-activated HCAEC. Inhibition of adhesion was additive with function blocking anti-E-selectin and anti-L-selectin antibodies, but was not additive with anti-CD18 antibody. These results show that antiflammins inhibit PMN adhesion to HCAEC by attenuating activation-induced up-regulation of CD11/CD18 expression on leukocytes, and suggest that antiflammins may represent a novel therapeutic approach in blocking leukocyte trafficking in host defense and inflammation. PMID- 10698974 TI - STAG3, a novel gene encoding a protein involved in meiotic chromosome pairing and location of STAG3-related genes flanking the Williams-Beuren syndrome deletion. AB - Chromatin rearrangements in the meiotic prophase are characterized by the assembly and disassembly of synaptonemal complexes (SC), a protein structure that stabilizes the pairing of homologous chromosomes in prophase. We report the identification of human and mouse cDNA coding for stromalin 3 (STAG3), a new mammalian stromalin member of the synaptonemal complex. The stromalins are a group of highly conserved proteins, represented in several organisms from yeast to humans. Stromalins are characterized by the stromalin conservative domain (SCD), a specific motif found in all proteins of the family described to date. STAG3 is expressed specifically in testis, and immunolocalization experiments show that STAG3 is associated to the synaptonemal complex. As the protein encoded by the homologous gene (Scc3p) in Saccharomyces cerevisiae was found to be a subunit of a cohesin complex that binds chromosomes until the onset of anaphase, our data suggest that STAG3 is involved in chromosome pairing and maintenance of synaptonemal complex structure during the pachytene phase of meiosis in a cohesin like manner. We have mapped the human STAG3 gene to the 7q22 region of chromosome 7; six human STAG3-related genes have also been mapped: two at 7q22 near the functional gene, one at 7q11.22, and three at 7q11.23, two of them flanking the breakpoints commonly associated with the Williams-Beuren syndrome (WBS) deletion. Since the WBS deletion occurs as a consequence of unequal meiotic crossing over, we suggest that STAG3 duplications predispose to germline chromosomal rearrangement within this region. PMID- 10698975 TI - Microfabrication of an analog of the basal lamina: biocompatible membranes with complex topographies. AB - A microfabrication approach was used to produce novel analogs of the basal lamina with complex topographic features. A test pattern of ridges and channels with length scales (40 to 310 micrometer) similar to the invaginations found in a native basal lamina was laser machined into the surface of a polyimide master chip. Negative replicates of the chip were produced using polydimethylsiloxane silicone elastomer and these replicates were used as templates for the production of thin ( approximately 21 micrometer) membranes of collagen or gelatin. The resulting membranes had a complex topography of ridges and channels that recapitulated the features of the master chip. To demonstrate their utility, these complex membranes were laminated to type I collagen sponges and their surfaces were seeded with cultured human epidermal keratinocytes to form a skin equivalent. The keratinocytes formed a differentiated and stratified epidermis that conformed to the features of the microfabricated membrane. The topography of the membrane influenced the differentiation of the keratinocytes because stratification was enhanced in the deeper channels. Membrane topography also controlled the gross surface features of the skin equivalent; infolds of the epidermis increased as channel depth increased. These novel microfabricated analogs of the basal lamina will help to elucidate the influence of topography on epithelial cell proliferation and differentiation and should have applications in the tissue engineering of skin equivalents as well as other basal lamina containing tissues. PMID- 10698976 TI - Efficiencies of translation in three reading frames of unusual non-ORF sequences isolated from phage display. AB - An unusual nucleotide sequence, called H10, was previously isolated by biopanning with a random peptide library on filamentous phage. The sequence encoded a peptide that bound to the growth hormone binding protein. Despite the fact that the H10 sequence can be expressed in Escherichia coli as a fusion to the gene III minor coat protein of the M13 phage, the sequence contained two TGA stop codons in the zero frame. Several mutant derivatives of the H10 sequence carried not only a stop codon, but also showed frameshifts, either +1 or -1 in individual isolates, between the H10 start and the gene III sequences. In this work, we have subcloned the H10 sequence and three of its derivatives (one requiring a +1 reading frameshift for expression, one requiring a -1 reading frameshift, and one open reading frame) in gene fusions to a reporter beta-galactosidase gene. These sequences have been cloned in all three reading frames relative to the reporter. The non-open reading frame constructs gave (surprisingly) high expression of the reporter (10-40% of control vector expression levels) in two out of the three frames. A site-directed mutant of the TGA stop codon (to TTA) in the +1 shifter greatly reduced the frameshift and gave expression primarily in the zero frame. By contrast, a site-directed mutant of the TGA in the -1 shifter had little effect on the pattern of expression, and alteration of the first TGA (of two) in H10 itself paradoxically reduced expression by half. We believe these phenomena to reflect a translational recoding mechanism in which ribosomes switch reading frames or read past stop codons upon encountering a signal encoded in the nucleotide sequence of the mRNA, because both the open reading frame derivative (which has six nucleotide changes from parental H10) and the site-directed mutant of the +1 shifter, primarily expressed the reporter only in the zero frame. PMID- 10698977 TI - Experimental biology 2000 april 15-18, 2000 san diego, calif PMID- 10698978 TI - Resolution of the pathways of poliovirus type 1 transmission during an outbreak. AB - An outbreak of poliomyelitis with 20 cases occurred in Israel, Gaza, and the West Bank from October 1987 to October 1988. The wild type 1 poliovirus associated with the outbreak was most closely related to viruses found in the Nile Delta. The epidemiologic links among patients involved in the outbreak and patients with community-acquired infections during the outbreak were inferred from the evolutionary relationships among isolates of the outbreak virus. Complete VP1 sequences (906 nucleotides) were determined for 12 clinical and 4 sewage isolates. A total of 58 nucleotide differences were found among the 16 isolates; 74% of all substitutions were synonymous third-position transitions. An evolutionary tree, representing both the pathways of VP1 sequence evolution and the inferred chains of virus transmission during the outbreak, was constructed under the assumption that each substitution had occurred only once. The combined epidemiologic and molecular data suggest that a single founder strain was introduced into Israel from the vicinity of Gaza in the fall of 1987. Poliovirus circulation was apparently localized to southern communities during the winter and spread north by the following summer into the Hadera subdistrict of Israel, where it radiated via multiple chains of transmission into other communities in northern Israel and the West Bank. The close sequence matches (>99%) between clinical and sewage isolates from the same communities confirm the utility of environmental sampling as a tool for monitoring wild poliovirus circulation. PMID- 10698979 TI - Identification by 16S rRNA gene analyses of a potential novel mycobacterial species as an etiological agent of canine leproid granuloma syndrome. AB - PCR amplifications of the 16S rRNA gene were performed on 46 specimens obtained from 43 dogs with canine leproid granuloma syndrome to help determine its etiology. Sequence capture PCR was applied to 37 paraffin-embedded specimens from 37 dogs, and nested PCR was attempted on DNA from 9 fresh tissue specimens derived from 3 of the 37 aforementioned dogs and from an additional 6 dogs. Molecular analyses of the paraffin-embedded tissues and fresh tissue specimen analyses were performed at separate institutions. PCR products with identical sequences over a 350-bp region encompassing variable regions 2 and 3 of the 16S rRNA gene were obtained from 4 of 37 paraffin-embedded specimens and from all 9 specimens of fresh tissue originating from 12 of the 43 dogs. Identical sequences were determined from amplicons obtained from paraffin-embedded and fresh specimens from one dog. The consensus DNA sequence, amplified from paraffin embedded tissue and represented by GenBank accession no. AF144747, shared highest nucleotide identity (99.4% over 519 bp) with mycobacterial strain IWGMT 90413 but did not correspond exactly to any EMBL or GenBank database sequence. With a probe derived from the V2 region of the novel canine sequence, reverse cross blot hybridization identified an additional four paraffin-embedded specimens containing the same novel sequence. In total, molecular methodologies identified the proposed novel mycobacterial sequence in 16 of 43 dogs with canine leproid granuloma syndrome, indicating that the species represented by this sequence may be the principal etiological agent of canine leproid granuloma syndrome. PMID- 10698980 TI - Accelerated detection and identification of mycobacteria with MGIT 960 and COBAS AMPLICOR systems. AB - An automated cultivation system for mycobacteria, the MGIT 960 system (MGIT system), was compared in the clinical routine with two variants of Lowenstein Jensen (L-J) medium. A total of 152 isolates were recovered from 2,015 specimens: 139 (91%) with the MGIT system and 127 (84%) with L-J media (P = 0.05). These included 68 isolates of Mycobacterium tuberculosis, of which 88% grew in the MGIT system and 93% grew in L-J media (P = 0.389), and 84 isolates of mycobacteria other than M. tuberculosis (MOTT), of which 94% grew in the MGIT system and 76% grew in L-J media (P = 0.003). More M. avium complex isolates were detected in the MGIT system (n = 65) than in L-J media (n = 50) (P = 0.001). Growth in the MGIT system was detected in 2 weeks for 78% of the isolates, whereas growth was detected in the two L-J media for 17 and 25% of the isolates, respectively. The mean times to detection of M. tuberculosis were 12 days in the MGIT system and 20 days in L-J media, and for M. avium complex the mean times to detection were 8 and 22 to 25 days, respectively. The contamination rates were similar (8.7 to 8.9%) in all media. A commercial amplification system (COBAS AMPLICOR) was evaluated for its ability to rapidly identify M. tuberculosis, M. avium, and M. intracellulare directly from 393 samples in MGIT system broth. A correct PCR result, as evaluated by culture or clinical data, was obtained for 96% of the samples, with inhibition being detected for 2% of the samples. Of the 89 results positive for M. tuberculosis, 91% were regarded as true positive, 8% were regarded as inconclusive, and 2% were considered false positive. For results positive for M. avium and M. intracellulare, 97 and 79%, respectively, were regarded as true positive. Increased rapidity and enhanced isolation of MOTT were obtained with the MGIT system. COBAS AMPLICOR was suitable for rapid identification of these three common pathogens from MGIT system broth. PMID- 10698981 TI - Genotyping of Mycoplasma pneumoniae clinical isolates reveals eight P1 subtypes within two genomic groups. AB - Three methods for genotyping of Mycoplasma pneumoniae clinical isolates were applied to 2 reference strains and 21 clinical isolates. By a modified restriction fragment length polymorphism (RFLP) analysis of PCR products of the M. pneumoniae cytadhesin P1 gene, 5 subtypes were discriminated among 13 P1 type 1 strains and 3 subtypes were discriminated among 8 P1 type 2 strains. Sequence analysis of the 16S-23S rRNA gene spacer region and part of the 23S rRNA gene revealed one nucleotide difference in the intergenic spacer region in 3 of the 21 isolates. In the 23S rRNA gene sequence of the 8 P1 type 2 strains an extra adenosine was present, but it was absent from the 13 P1 type 1 strains. On the basis of M. pneumoniae genome sequence data, primers were designed to amplify large interrepeat fragments by long PCR, and these fragments were subsequently analyzed by RFLP analysis. Only two types, long PCR types 1 and 2, could be discriminated among the M. pneumoniae isolates. All P1 type 1 strains were assigned to long PCR type 1, and all P1 type 2 strains were assigned to long PCR type 2. These data obtained by three independent typing methods thus confirm the existence of two distinct M. pneumoniae genomic groups but expand the possibility of strain typing on the basis of variations within their P1 genes. PMID- 10698982 TI - Evaluation of a capacitance method for direct antifungal susceptibility testing of yeasts in positive blood cultures. AB - The feasibility of using a capacitance method (CM) for direct antifungal susceptibility testing of yeasts in positive blood cultures was evaluated. The CM used the same test conditions as those recommended by the National Committee for Clinical Laboratory Standards. After direct inoculation of positive culture broths into module wells (Bactometer; bioMerieux, Inc., Hazelwood, Mo.), the end point determination was made by monitoring the capacitance change in the culture broths with Bactometer. The MIC of amphotericin B was the lowest concentration at which yeast growth was completely inhibited, while the MICs of ketoconazole, flucytosine, and fluconazole were the concentrations at which a >/=80% reduction in capacitance change was observed. The MICs of the four drugs against each blood isolate obtained on subculture plates were also determined by the macrodilution method. For 51 positive blood cultures tested, the percent agreement (+/-2 log(2) dilutions) between the CM and the macrodilution method were as follows: amphotericin B (98%), ketoconazole (92%), flucytosine (84%), and fluconazole (96%). The CM was further used for breakpoint susceptibility testing of fluconazole (8 and 64 microg/ml) and flucytosine (4 and 32 microg/ml) against yeasts in positive blood cultures. After testing of 74 specimens by the CM, flucytosine and fluconazole produced one (1.4%) major error and two (2.8%) minor errors, respectively. All yeasts that displayed resistance to flucytosine or fluconazole were detected within 24 h after direct inoculation of the positive broths into Bactometer. The CM may be useful for the rapid detection of antifungal resistance in positive blood cultures containing yeasts. PMID- 10698983 TI - Identification of the major Spanish clones of penicillin-resistant pneumococci via the Internet using multilocus sequence typing. AB - Multilocus sequence typing was used to characterize isolates of the major Spanish clones of penicillin-resistant and multiple-antibiotic-resistant Streptococcus pneumoniae. Isolates of the multidrug-resistant Spanish serotype 23F clone and serotype variants of this clone either had identical allelic profiles or their allelic profiles differed from this typical allelic profile at only one of the seven housekeeping loci. Similarly, isolates of the Spanish serotype 6B and 14 clones and the penicillin-resistant serotype 9V clone (and serotype variants of this clone) each had the same allelic profiles or profiles that differed at a single locus. Multilocus sequence typing therefore allows resistant pneumococci to be assigned to the Spanish clones if they have the typical allelic profile of the clone or if their profiles differ from that profile at a single locus. A few resistant isolates that had allelic profiles typical of that of a Spanish clone or whose profiles differed from that of the typical profile at only a single locus possessed penicillin-binding protein pbp1a, pbp2b, or pbp2x genes that differed from those that are characteristic of the clone. In most cases these isolates could be assigned as variant members of the clone. Since almost all serotype 9V isolates have very similar genotypes, independently emerging penicillin-resistant clones of this serotype will inevitably appear to be similar by molecular typing procedures. Analysis of the pbp genes, in addition to multilocus sequence typing (or any other molecular typing procedure), is therefore required to assign isolates unambiguously to the penicillin-resistant Spanish serotype 9V clone. PMID- 10698984 TI - Serological and virological characterization of clinically diagnosed cases of measles in suburban Khartoum. AB - Measles continues to be a major childhood disease in terms of global morbidity and mortality. In the main areas of its endemicity the only available means of diagnosis are based on clinical criteria: the presence of a maculopapular rash and fever accompanied by cough, coryza, and/or conjunctivitis. We have studied 38 clinically diagnosed cases of measles in Khartoum, Sudan, by means of serology, reverse transcriptase PCR (RT-PCR) on throat swabs and virus isolation from lymphocytes. On the basis of serology, 28 patients were diagnosed as having an acute measles virus (MV) infection, while in 10 cases the clinical symptoms proved to have other causes. It was shown that in cases with low serum immunoglobulin M (IgM) levels, an additional measurement of IgG or virus neutralizing antibodies was necessary to discriminate between patients with an acute MV infection sampled during an early stage of the disease and patients who had experienced an MV infection in the more distant past. The serological laboratory diagnosis was validated by an MV-specific RT-PCR: for all confirmed measles cases tested a fragment of the correct size which hybridized with a third MV-specific primer could be amplified, while all serologically negative cases were also RT-PCR negative. MV could be isolated from 17 out of 23 of the serologically confirmed cases, demonstrating that virus isolation is less reliable as a diagnostic tool than serology or RT-PCR. This study stresses the urgent need for a rapid diagnostic field test for measles. PMID- 10698985 TI - Molecular analysis of CAP59 gene sequences from five serotypes of Cryptococcus neoformans. AB - The nucleotide sequences of CAP59 genes from five serotypes of Cryptococcus neoformans were analyzed for their phylogenetic relationships. Approximately 600 bp genomic DNA fragments of the CAP59 gene were amplified from each isolate by PCR and sequenced. The CAP59 nucleotide sequences of C. neoformans showed more than 90% similarity among the five serotypes. By phylogenetic analysis, their sequences were divided into three clusters: serotypes A and AD, serotypes B and C, and serotype D. In addition, the results of reduced amino acid sequences were similar to the nucleotide sequence data. These data revealed that serotype AD was genetically close to serotype A rather than serotype D, although it had been considered to be a mixed type of serotype A and D by serological analysis. Furthermore, the nucleotide sequences of the serotype B and C isolates of C. neoformans were very similar to each other. These results indicated that serotype B and C isolates belonging to C. neoformans var. gattii were genetically homogeneous and closely related. The molecular analysis of the CAP59 gene will provide useful information for the differentiation of serotypes of C. neoformans and for an understanding of their phylogenetic relationships. PMID- 10698986 TI - Early aqueous humor analysis in patients with human ocular toxoplasmosis. AB - To evaluate the diagnostic sensitivity of a panel of laboratory tests for ocular toxoplasmosis performed at the time of presentation, paired samples of aqueous humor and serum were collected from 49 consecutive episodes of ocular toxoplasmosis with a clinical course of less than 3 weeks. Total immunoglobulin G (IgG) and Toxoplasma gondii-specific IgG, IgM, and IgA were quantified by enzyme linked immunosorbent assay. The avidity of T. gondii-specific IgG was determined, and DNA extracted from aqueous humor was amplified for detection of a glycoprotein B gene sequence of T. gondii. The diagnosis was confirmed for 73% (36 of 49) of the patients; this rate rose to 79.5% if data from a later analysis of aqueous humor derived from five of the negative patients were included. The analysis of serum (detection of T. gondii-specific IgM and analysis of consecutive serum samples) alone did not contribute to the diagnosis. Calculation of local antibody production lacked diagnostic sensitivity when it was determined less than 3 weeks after the manifestation of clinical symptoms (28 of 49 patients [57%]), but this rose to 70% after an analysis of a second aqueous humor sample. The antibody avidity index attained diagnostic significance in only 8 of 43 instances (19%), and T. gondii DNA was amplified from no more than 6 of 39 (16%) aqueous humor samples. However, T. gondii-specific IgA was found within the aqueous humors of 11 of 43 patients (26%); measurement of the T. gondii-specific IgA level thus contributed substantially to the diagnostic sensitivity of the laboratory tests. PMID- 10698987 TI - PCR detection of granulocytic ehrlichiae in Ixodes ricinus ticks and wild small mammals in western Switzerland. AB - The presence of granulocytic ehrlichiae was demonstrated by PCR in Ixodes ricinus ticks and wild small mammals in Switzerland in two areas of endemicity for bovine ehrlichiosis. Six ticks (three females and three nymphs) (1.4%) of 417 I. ricinus ticks collected by flagging vegetation contained ehrlichial DNA. A total of 201 small mammals from five species, wood mouse (Apodemus sylvaticus), yellow-necked mouse (Apodemus flavicollis), earth vole (Pitymys subterraneus), bank vole (Clethrionomys glareolus), and common shrew (Sorex araneus), were trapped. The analysis of I. ricinus ticks [corrected] collected on 116 small mammals showed that nine C. glareolus voles and two A. sylvaticus mice hosted infected tick larvae. In these rodents, granulocytic ehrlichia infection was also detected in blood, spleen, liver, and ear samples. Further examinations of 190 small mammals without ticks or with noninfected ticks showed the presence of ehrlichial DNA in spleen and other tissues from six additional C. glareolus, three A. flavicollis, and one S. araneus mammals. This study suggests that A. sylvaticus, A. flavicollis, S. araneus, and particularly C. glareolus are likely to be natural reservoirs for granulocytic ehrlichiae. Partial 16S rRNA gene sequences of granulocytic ehrlichiae from ticks and rodents showed a high degree of homology (99 to 100%) with granulocytic ehrlichiae isolated from humans. In contrast, groESL heat shock operon sequence analysis showed a strong divergence (approximately 5%) between the sequences in samples derived from rodents and those derived from samples from questing ticks or from other published ehrlichia sequences. Dual infections with granulocytic ehrlichia and Borrelia burgdorferi were found in ticks and small mammals. PMID- 10698988 TI - Multilocus sequence typing for characterization of methicillin-resistant and methicillin-susceptible clones of Staphylococcus aureus. AB - A multilocus sequence typing (MLST) scheme has been developed for Staphylococcus aureus. The sequences of internal fragments of seven housekeeping genes were obtained for 155 S. aureus isolates from patients with community-acquired and hospital-acquired invasive disease in the Oxford, United Kingdom, area. Fifty three different allelic profiles were identified, and 17 of these were represented by at least two isolates. The MLST scheme was highly discriminatory and was validated by showing that pairs of isolates with the same allelic profile produced very similar SmaI restriction fragment patterns by pulsed-field gel electrophoresis. All 22 isolates with the most prevalent allelic profile were methicillin-resistant S. aureus (MRSA) isolates and had allelic profiles identical to that of a reference strain of the epidemic MRSA clone 16 (EMRSA-16). Four MRSA isolates that were identical in allelic profile to the other major epidemic MRSA clone prevalent in British hospitals (clone EMRSA-15) were also identified. The majority of isolates (81%) were methicillin-susceptible S. aureus (MSSA) isolates, and seven MSSA clones included five or more isolates. Three of the MSSA clones included at least five isolates from patients with community acquired invasive disease and may represent virulent clones with an increased ability to cause disease in otherwise healthy individuals. The most prevalent MSSA clone (17 isolates) was very closely related to EMRSA-16, and the success of the latter clone at causing disease in hospitals may be due to its emergence from a virulent MSSA clone that was already a major cause of invasive disease in both the community and hospital settings. MLST provides an unambiguous method for assigning MRSA and MSSA isolates to known clones or assigning them as novel clones via the Internet. PMID- 10698989 TI - Comparison of different PCR approaches for typing of Francisella tularensis strains. AB - In this study, we evaluated three PCR methods for epidemiological typing of Francisella tularensis: repetitive extragenic palindromic element PCR (REP-PCR), enterobacterial repetitive intergenic consensus sequence PCR (ERIC-PCR), and random amplified polymorphic DNA (RAPD) assay with both M13 and T3-T7 primers. The analysis was performed with 40 strains of F. tularensis isolated from hares, humans, ticks, and a vole. On the basis of the combination of REP, ERIC, and RAPD fingerprints, F. tularensis strains were divided into 17 genetic groups (designated A to Q), and one Francisella novicida strain was classified in group R. The F. novicida strain is of special concern, since previous genetic methods have been unable to clearly distinguish between F. tularensis and F. novicida. The F. tularensis isolates recovered from hares were included in groups A to J, M, and P; those recovered from humans were included in groups A, D, G, J, L, O, and N; those isolated from ticks were included in groups B and Q; and that recovered from a vole was in group K. The diversities calculated for the 40 F. tularensis isolates, according to Simpson's index, were 0.14 for REP-PCR, 0.52 for ERIC-PCR, 0.39 for RAPD assay with the M13 primer (RAPD/M13-PCR), and 0.65 for RAPD/T3-T7-PCR, and the diversity increased up to 0.90 when ERIC-PCR, RAPD/M13-PCR, and RAPD/T3-T7-PCR were combined. Our results suggest that although limited genetic heterogeneity among F. tularensis strains was observed, this small variation is enough to validate the PCR methods used in this study and their combinations, because they can provide safe, useful, and rapid tools for the typing of F. tularensis. PMID- 10698990 TI - Prevalence and characterization of Shiga toxin-producing Escherichia coli isolated from cattle, food, and children during a one-year prospective study in France. AB - During a 1-year survey of Shiga toxin-producing Escherichia coli (STEC) prevalence in central France, 2,143 samples were investigated by PCR for Shiga toxin-encoding genes. A total of 330 (70%) of 471 fecal samples collected from healthy cattle at the Clermont-Ferrand slaughterhouse, 47 (11%) of 411 beef samples, 60 (10%) of 603 cheese samples, and 19 (3%) of 658 stool specimens from hospitalized children with and without diarrhea were positive for the stx gene(s). A STEC strain was isolated from 34% (162 of 471) of bovine feces, 4% (16 of 411) of beef samples, 1% (5 of 603) of cheese samples, and 1.5% (10 of 658) of stool specimens. Of the 220 STEC strains isolated, 34 (15%) harbored the stx(1) gene, 116 (53%) harbored the stx(2) gene, and 70 (32%) carried both the stx(1) and stx(2) genes. However, 32 (14.5%) were not cytotoxic for Vero cells. The eae gene, found in 12 (5%) of the 220 strains, was significantly associated with the stx(1) gene and with isolates from children. Sequences homologous to ehxA were found in 102 (46%) of the 220 strains. Thirteen serotypes, OX3:H2, O113:H21, O113:H4, OX3:H21, O6:H10, OX178:H19, O171:H2, O46:H38, O172:H21, O22:H16, O91:H10, O91:H21, and O22:H8, accounted for 102 (55%) of 186 typeable isolates, and only one strain (0.5% of the 186 STEC isolates from cattle), belonged to the O157:H7 serotype. We showed that the majority of the STEC isolates from cattle, beef, and cheese were not likely to be pathogenic for humans and that the STEC strains isolated from children in this study were probably not responsible for diarrheal disease. Finally, the strains associated with hemolytic-uremic syndrome in the same geographical area were shown to belong to particular subsets of the STEC population found in the bovine reservoir. PMID- 10698991 TI - Multiplex PCR for detection of genes for Staphylococcus aureus enterotoxins, exfoliative toxins, toxic shock syndrome toxin 1, and methicillin resistance. AB - A multiplex PCR assay for detection of genes for staphylococcal enterotoxins A to E (entA, entB, entC, entD, and entE), toxic shock syndrome toxin 1 (tst), exfoliative toxins A and B (etaA and etaB), and intrinsic methicillin resistance (mecA) was developed. Detection of femA was used as an internal positive control. The multiplex PCR assay combined the primers for sea to see and femA in one set and those for eta, etb, tst, mecA, and femA in the other set. Validation of the assay was performed using 176 human isolates of Staphylococcus aureus. This assay offers a very specific, quick, reliable, and inexpensive alternative to conventional PCR assays used in clinical laboratories to identify various staphylococcal toxin genes. PMID- 10698992 TI - Failure of an automated blood culture system to detect nonfermentative gram negative bacteria. AB - During a 1-year study we observed that both aerobic and anaerobic blood culture bottles from patients were negative by the BacT/Alert system during a 7-day incubation period. However, upon subcultivation of negative bottles, growth of Pseudomonas aeruginosa was detectable. In an attempt to explain this observation, aerobic BacT/Alert Fan bottles were seeded with a defined inoculum (0.5 McFarland standard; 1 ml) of Escherichia coli, Klebsiella pneumoniae, Serratia marcescens, P. aeruginosa, Stenotrophomonas maltophilia, or Acinetobacter baumannii. Half of the inoculated bottles were loaded into the BacT/Alert system immediately, and the remainder were preincubated for 4, 8, 16, and 24 h at 36 degrees C. With preincubation all bottles seeded with the Enterobacteriaceae signaled positive during the next 1.5 h. Organisms in bottles seeded with the nonfermentative species P. aeruginosa and A. baumannii remained undetected by the BacT/Alert system for 7 days. S. maltophilia was detected if the preincubation time was equal or less than 8 h. Without preincubation all bottles seeded with the Enterobacteriaceae or nonfermentative species signaled positive. Since nonfermentative species seem to enter a state of bacteriostasis within the preincubation period, we reasoned that an unknown factor is consumed. Accordingly, a smaller inoculum should allow the detection of nonfermentative species, even after preincubation, and serial dilutions of P. aeruginosa were detected in preincubated bottles. In this case preincubated bottles signaled positive faster than bottles without preincubation. We conclude that all bottles from clinical settings should be subcultured prior to loading to avoid false negatives. An alternative may be preincubation at room temperature. PMID- 10698993 TI - Identification and population structure of Burkholderia stabilis sp. nov. (formerly Burkholderia cepacia genomovar IV). AB - The Burkholderia cepacia complex currently comprises five genomic species, i.e., B. cepacia genomovar I, B. multivorans (formerly known as B. cepacia genomovar II), B. cepacia genomovar III, B. cepacia genomovar IV, and B. vietnamiensis (also known as B. cepacia genomovar V). In the absence of straightforward diagnostic tests for the identification of B. cepacia genomovars I, III, and IV, the last two genomic species were not formally classified as novel Burkholderia species (genomovar I contains the type strain and therefore retains the name B. cepacia). In the present study, we describe differential biochemical tests and a recA gene-based PCR assay for the routine identification of strains currently known as B. cepacia genomovar IV and propose formal classification of this organism as Burkholderia stabilis sp. nov. B. stabilis can indeed be differentiated from all other B. cepacia complex strains by the absence of beta galactosidase activity, from strains of B. cepacia genomovars I and III and B. vietnamiensis by the inability to oxidize sucrose, and from B. multivorans by the lack of growth at 42 degrees C. In addition, analysis with the recA gene-derived primers BCRG41 (5'-ACCGGCGAGCAGGCGCTT-3') and BCRG42 (5'-ACGCCATCGGGCATGGCA-3') specifically allows the detection of B. stabilis strains in a conventional PCR assay. Examination of a set of 21 B. stabilis strains by means of random amplified polymorphic DNA analysis and pulsed-field gel electrophoresis typing suggested that the genome of this organism is highly conserved, which is in sharp contrast to the generally accepted genomic diversity, variability, and plasticity among B. cepacia strains. PMID- 10698994 TI - Imipenem resistance of enterobacter aerogenes mediated by outer membrane permeability. AB - Multidrug-resistant Enterobacter aerogenes strains are increasingly isolated in Europe and especially in France. Treatment leads to imipenem resistance, because of a lack of porin. We studied the evolution of resistance in 29 strains isolated from four patients during their clinical course. These strains belonged to the prevalent epidemiological type observed in France in previous studies (C. Bosi, et al., J. Clin. Microbiol. 37:2165-2169, 1999; A. Davin-Regli et al., J. Clin. Microbiol. 34:1474-1480, 1996). They also harbored a TEM-24 extended-spectrum beta-lactamase-coding gene. Thirteen strains were susceptible to gentamicin and resistant to imipenem and cefepime. All of the patients showed E. aerogenes strains with this resistance after an imipenem treatment. One patient showed resistance to imipenem after a treatment with cefpirome. Twelve of these 13 strains showed a lack of porin. Cessation of treatment with imipenem for three patients was followed by reversion of susceptibility to this antibiotic and the reappearance of porins, except in one case. For one patient, we observed three times in the same day the coexistence of resistant strains lacking porin and susceptible strains possessing porin. The emergence of multidrug-resistant E. aerogenes strains is very disquieting. In our study, infection by E. aerogenes increased the severity of the patients' illnesses, causing a 100% fatality rate. PMID- 10698995 TI - An antigen capture enzyme-linked immunosorbent assay reveals high levels of the dengue virus protein NS1 in the sera of infected patients. AB - We describe the development of a capture enzyme-linked immunosorbent assay for the detection of the dengue virus nonstructural protein NS1. The assay employs rabbit polyclonal and monoclonal antibodies as the capture and detection antibodies, respectively. Immunoaffinity-purified NS1 derived from dengue 2 virus infected cells was used as a standard to establish a detection sensitivity of approximately 4 ng/ml for an assay employing monoclonal antibodies recognizing a dengue 2 serotype-specific epitope. A number of serotype cross-reactive monoclonal antibodies were also shown to be suitable probes for the detection of NS1 expressed by the remaining three dengue virus serotypes. Examination of clinical samples demonstrated that the assay was able to detect NS1 with minimal interference from serum components at the test dilutions routinely used, suggesting that it could form the basis of a useful additional diagnostic test for dengue virus infection. Furthermore, quantitation of NS1 levels in patient sera may prove to be a valuable surrogate marker for viremia. Surprisingly high levels of NS1, as much as 15 microg/ml, were found in acute-phase sera taken from some of the patients experiencing serologically confirmed dengue 2 virus secondary infections but was not detected in the convalescent sera of these patients. In contrast, NS1 could not be detected in either acute-phase or convalescent serum samples taken from patients with serologically confirmed primary infection. The presence of high levels of secreted NS1 in the sera of patients experiencing secondary dengue virus infections, and in the context of an anamnestic antibody response, suggests that NS1 may contribute significantly to the formation of the circulating immune complexes that are suspected to play an important role in the pathogenesis of severe dengue disease. PMID- 10698996 TI - Epidemiology of astrovirus infection in young children hospitalized with acute gastroenteritis in Melbourne, Australia, over a period of four consecutive years, 1995 to 1998. AB - The incidence of astrovirus infection in children less than 5 years of age hospitalized with acute gastroenteritis in Melbourne, Australia, from 1995 to 1998 was determined. Astrovirus was detected in 40 of 449 specimens tested by Northern hybridization, and astrovirus infection was confirmed by reverse transcription-PCR with or without culture in CaCO-2 cells. This represented 3.0% (40 of 1, 327) of all children tested for enteric pathogens, including viral, bacterial, and parasitic pathogens, over the survey period. The incidences of astrovirus infection in each year were 4.4% (1995), 2. 2% (1996), 3.9% (1997), and 1.4% (1998). In 1995 and 1997, the incidences of astrovirus infection were greater than the incidence of infection with all individual bacterial pathogens and were either greater than or equal to the incidence of adenovirus infection. Astrovirus exhibited an unusual biennial winter peak of incidence that correlated with a greater incidence of serotype 1 virus and an increased rate of hospitalization of children aged 6 to 12 months. Uncommon (serotype 2 and 4) and rare (serotype 8) serotypes were detected during the survey period. Genetic analysis of ORF2 (which encodes the astrovirus capsid precursor) of Melbourne isolates showed nucleotide sequence variation from year to year. This was not accompanied by significant amino acid substitutions. However, geographical variation was apparent by comparison of Melbourne astrovirus isolates with prototype strains identified in the United Kingdom. PMID- 10698997 TI - Evaluation of the Oricult-N dipslide for laboratory diagnosis of vaginal candidiasis. AB - The Oricult-N semiquantitative dipslide (Orion Diagnostica, Espoo, Finland) was evaluated for the laboratory diagnosis of vaginal candidiasis. It was compared with broth culture (Vagicult; Orion Diagnostica). Oricult-N was positive for 14.5% of 124 symptomatic patients and 12% of 50 asymptomatic controls. The results for broth cultures were 17 and 22%, respectively. Thus, the test group and the control group did not differ significantly by either method. High vaginal yeast counts (>/=10(5) CFU/ml) were detected by Oricult-N in 7% of patients and in 0% of controls, but both groups harbored low numbers of yeasts. An accurate quantitative cutoff point separating a level of yeast associated with infection from vaginal yeast carriage could not be defined in the study. Nevertheless, the easy semiquantitation allowed by the Oricult-N method could be helpful because, especially in low-count carriers of Candida, other potential causes of vaginal symptoms should be considered. The Oricult-N method was technically simple and could be applied in primary health care. Further studies are required, however, before Oricult-N can be recommended as a routine diagnostic tool. PMID- 10698998 TI - Simultaneous detection of multiplex-amplified human immunodeficiency virus type 1 RNA, hepatitis C virus RNA, and hepatitis B virus DNA using a flow cytometer microsphere-based hybridization assay. AB - The feasibility of performing a multiplex assay for the detection of human immunodeficiency virus type 1 (HIV-1) and hepatitis C virus (HCV) RNAs and hepatitis B virus (HBV) DNA is demonstrated. This assay is based (i) on the coamplification of a 142-bp fragment from the gag region of the HIV-1 genome and a 142-bp HIV-1 quantitation standard fragment, a 244-bp fragment from the 5' noncoding region of the HCV genome, and a 104-bp fragment from the pre-C and C gene regions of the HBV genome, using three sets of specific primers; (ii) on the capacity of these four biotinylated PCR products to hybridize to their specific oligonucleotide probe-coated microspheres; and (iii) on the ability of the flow cytometer to discriminate between distinct fluorescent-microsphere categories. Absence of cross-hybridization between the unrelated oligonucleotide probes and PCR products generated by the multiplex reverse transcription-PCR (RT-PCR) and the highly sensitive detection method allowed us to assess unambiguously the HIV 1 viral load and the infectious status of 35 serologically well-established clinical samples and 20 seronegative blood donor plasma samples tested. The results indicate that multiplex RT-PCR and flow cytometer microsphere-based hybridization assays, when combined, provide a rapid, sensitive, and specific method for the quantitation and detection of the major viral agents of infectious diseases in a single plasma sample. PMID- 10699000 TI - Evaluation of the discriminatory powers of the Dienes test and ribotyping as typing methods for Proteus mirabilis. AB - A total of 63 clinical isolates of Proteus mirabilis collected over a 19-month period were typed by the Dienes test and ribotyping. Ribotyping was performed using the fully automated RiboPrinter Microbial Characterization System (Qualicon, Wilmington, Del.). Isolates that were indistinguishable by the Dienes test and/or ribotyping were characterized further by pulsed-field gel electrophoresis (PFGE). Most of the isolates represented unique strains as judged by the Dienes test and ribotyping. Forty isolates represented 40 different ribotypes and Dienes types. The remaining 23 isolates were grouped into 13 Dienes types, 12 ribotypes, and 14 PFGE types. The index of discrimination was 0.980 for the Dienes test, 0.979 for ribotyping, and 0.992 for PFGE. Both the Dienes test and ribotyping are useful methods for identifying individual strains of P. mirabilis. The Dienes test is simple, inexpensive, and easy to perform. It can be performed in virtually any laboratory and should be used in the initial epidemiologic characterization of P. mirabilis isolates. PMID- 10698999 TI - Identification of Helicobacter pylori and other Helicobacter species by PCR, hybridization, and partial DNA sequencing in human liver samples from patients with primary sclerosing cholangitis or primary biliary cirrhosis. AB - Helicobacter pylori was identified in human liver tissue by PCR, hybridization, and partial DNA sequencing. Liver biopsies were obtained from patients with primary sclerosing cholangitis (n = 12), primary biliary cirrhosis (n = 12), and noncholestatic liver cirrhosis (n = 13) and (as controls) normal livers (n = 10). PCR analyses were carried out using primers for the Helicobacter genus, Helicobacter pylori (the gene encoding a species-specific 26-kDa protein and the 16S rRNA), Helicobacter bilis, Helicobacter pullorum, and Helicobacter hepaticus. Samples from patients with primary biliary cirrhosis and primary sclerosing cholangitis (11 and 9 samples, respectively) were positive by PCR with Helicobacter genus-specific primers. Of these 20 samples, 8 were positive with the 16S rRNA primer and 9 were positive with the 26-kDa protein primer of H. pylori. These nine latter samples were also positive by Southern blot hybridization for the amplified 26-kDa fragment, and four of those were verified to be H. pylori by partial 16S rDNA sequencing. None of the samples reacted with primers for H. bilis, H. pullorum, or H. hepaticus. None of the normal livers had positive results in the Helicobacter genus PCR assay, and only one patient in the noncholestatic liver cirrhosis group, a young boy who at reexamination showed histological features suggesting primary sclerosing cholangitis, had a positive result in the same assay. Helicobacter positivity was thus significantly more common in patients with cholestatic diseases (20 of 24) than in patients with noncholestatic diseases and normal controls (1 of 23) (P = <0.00001). Patients positive for Helicobacter genus had significantly higher values of alkaline phosphatases and prothrombin complex than Helicobacter-negative patients (P = 0.0001 and P = 0.0003, respectively). Among primary sclerosing cholangitis patients, Helicobacter genus PCR positivity was weakly associated with ulcerative colitis (P = 0.05). Significant differences related to blood group or HLA status were not found. PMID- 10699001 TI - Evaluation of the indirect hemagglutination assay for diagnosis of acute leptospirosis in Hawaii. AB - Timely diagnosis of leptospirosis is important to ensure a favorable clinical outcome. The definitive serologic assay, the microscopic agglutination test (MAT), requires paired sera and is not useful for guiding early clinical management. The only screening test approved for use in the United States, the indirect hemagglutination assay (IHA), has not undergone extensive field evaluation. To assess the performance of the leptospirosis IHA in Hawaii, serum from patients evaluated for leptospirosis between 1992 and 1997 were tested with the IHA at the Hawaii State Laboratories Division and with the MAT at the Centers for Disease Control and Prevention. Leptospirosis was considered confirmed by a fourfold rise in MAT titer and/or a positive culture. A total of 92 (41%) of 226 specimens from 114 persons with confirmed leptospirosis were found positive by IHA. Only 18 (15%) of 119 specimens obtained within 14 days of onset were IHA positive, compared to 74 (69%) of 107 specimens collected more than 14 days after onset (P <0.001). Repeat testing ultimately resulted in 78 (68%) of the confirmed cases having at least one specimen found positive by IHA. Thirteen different presumptive infecting serogroups were identified among 251 specimens with an MAT titer of >/=200 and obtained from persons with confirmed or probable leptospirosis. Fifty (68%) of 73 specimens with Icterohaemorrhagiae as the presumptive infecting serogroup were found positive by IHA, compared to 44 (47%) of 93 specimens with Australis as the presumptive infecting serogroup (P, 0.01). The IHA test was positive for 3 (1%) of 236 specimens from 154 persons without leptospirosis. The sensitivity of the leptospirosis IHA in Hawaii was substantially below figures reported previously, particularly early in the course of illness, limiting its usefulness for diagnosing acute infection. Since the presumptive infecting serogroup affected IHA results and the prevalence of serovars varies with geography, the performance of the IHA should be assessed locally. More sensitive leptospirosis screening tests are needed in Hawaii. PMID- 10699002 TI - Touchdown enzyme time release-PCR for detection and identification of Chlamydia trachomatis, C. pneumoniae, and C. psittaci using the 16S and 16S-23S spacer rRNA genes. AB - Three touchdown enzyme time release (TETR)-PCR assays were used to amplify different DNA sequences in the variable regions of the 16S and 16S-23S spacer rRNA genes specific for Chlamydia trachomatis, Chlamydia pneumoniae, and Chlamydia psittaci as improved tests for sensitive diagnosis and rapid species differentiation. The TETR-PCR protocol used 60 cycles of amplification, which provided improved analytical sensitivity (0.004 to 0.063 inclusion-forming unit of Chlamydia species per PCR). The sensitivity of TETR-PCR with primer set CTR 70 CTR 71 was 96.7%, and the specificity was 99.6%, compared to those of the AMPLICOR PCR for the detection of C. trachomatis in vaginal swab samples. TETR PCR for C. pneumoniae with primer set CPN 90-CPN 91 was 90% sensitive and 93.3% specific compared with a nested PCR with primer set CP1/2-CPC/D for clinical respiratory samples. TETR-PCR for C. psittaci with primer set CPS 100-CPS 101 showed substantial agreement with cell culturing (kappa, 0.78) for animal tissue samples. Primer sets were then combined into a single multiplex TETR-PCR test. The respective 315-, 195-, and 111-bp DNA target products were precisely amplified when DNA from each of the respective Chlamydia species or combinations of them was used. Multiplex chlamydia TETR-PCR correctly identified one strain of each of the 15 serovars of C. trachomatis, 22 isolates of C. pneumoniae, and 20 isolates of C. psittaci. The primer sets were specific for each species. No target products were amplified when DNA from C. pecorum or a variety of other microorganisms was tested for specificity. TETR-PCR with primers selected for specific sequences in the 16S and 16S-23S spacer rRNA genes is a valuable test that could be used either with individual primers or in a multiplex assay for the identification and differentiation of Chlamydia species from culture isolates or for the detection of chlamydiae in clinical samples. PMID- 10699003 TI - Novel diagnostic algorithm for identification of mycobacteria using genus specific amplification of the 16S-23S rRNA gene spacer and restriction endonucleases. AB - A novel genus-specific PCR for mycobacteria with simple identification to the species level by restriction fragment length polymorphism (RFLP) was established using the 16S-23S ribosomal RNA gene (rDNA) spacer as a target. Panspecificity of primers was demonstrated on the genus level by testing 811 bacterial strains (122 species in 37 genera from 286 reference strains and 525 clinical isolates). All mycobacterial isolates (678 strains among 48 defined species and 5 indeterminate taxons) were amplified by the new primers. Among nonmycobacterial isolates, only Gordonia terrae was amplified. The RFLP scheme devised involves estimation of variable PCR product sizes together with HaeIII and CfoI restriction analysis. It yielded 58 HaeIII patterns, of which 49 (84%) were unique on the species level. Hence, HaeIII digestion together with CfoI results was sufficient for correct identification of 39 of 54 mycobacterial taxons and one of three or four of seven RFLP genotypes found in Mycobacterium intracellulare and Mycobacterium kansasii, respectively. Following a clearly laid out diagnostic algorithm, the remaining unidentified organisms fell into five clusters of closely related species (i.e., the Mycobacterium avium complex or Mycobacterium chelonae-Mycobacterium abscessus) that were successfully separated using additional enzymes (TaqI, MspI, DdeI, or AvaII). Thus, next to slowly growing mycobacteria, all rapidly growing species studied, including M. abscessus, M. chelonae, Mycobacterium farcinogenes, Mycobacterium fortuitum, Mycobacterium peregrinum, and Mycobacterium senegalense (with a very high 16S rDNA sequence similarity) were correctly identified. A high intraspecies sequence stability and the good discriminative power of patterns indicate that this method is very suitable for rapid and cost-effective identification of a wide variety of mycobacterial species without the need for sequencing. Phylogenetically, spacer sequence data stand in good agreement with 16S rDNA sequencing results, as was shown by including strains with unsettled taxonomy. Since this approach recognized significant subspecific genotypes while identification of a broad spectrum of mycobacteria rested on identification of one specific RFLP pattern within a species, this method can be used by both reference (or research) and routine laboratories. PMID- 10699004 TI - Multicenter evaluation of the AMPLICOR and automated COBAS AMPLICOR CT/NG tests for detection of Chlamydia trachomatis. AB - The fully automated COBAS AMPLICOR CT/NG and semiautomated AMPLICOR CT/NG tests were evaluated in a multicenter trial for the ability to detect Chlamydia trachomatis infections. Test performance compared to that of culture was evaluated for 2,236 matched endocervical swab and urine specimens obtained from women and for 1,940 matched urethral swab and urine specimens obtained from men. Culture-negative, PCR-positive specimens that tested positive in a direct fluorescent-antibody test or in a confirmatory PCR test for an alternative target sequence were resolved as true positives. The overall prevalences of chlamydia were 2.4% in women and 7.2% in men. The COBAS AMPLICOR and AMPLICOR formats yielded concordant results for 98.1% of the specimens. With the infected patient as the reference standard, the resolved sensitivities of COBAS AMPLICOR were 89.7% for endocervical swab specimens, 89.2% for female urine specimens, 88.6% for male urethral swab specimens, and 90.3% for male urine specimens. When results were analyzed as if only a single test had been performed on a single specimen type, the resolved sensitivity was always higher. The resolved specificities of PCR were 99.4% for endocervical swab specimens, 99.0% for female urine specimens, 98.7% for male urethral swab specimens, and 98.4% for male urine specimens. The internal control revealed that 2.4% of the specimens were inhibitory when initially tested. Nevertheless, valid results were obtained for 98.6% of the specimens because 59.1% of the inhibitory specimens were not inhibitory when a second aliquot was tested. The COBAS AMPLICOR and AMPLICOR CT/NG tests for C. trachomatis exhibited equally high sensitivity and specificity with both urogenital swab and urine specimens and thus are well suited for screening for C. trachomatis infection. PMID- 10699005 TI - Quantitative and cost comparison of ultrasensitive human immunodeficiency virus type 1 RNA viral load assays: Bayer bDNA quantiplex versions 3.0 and 2.0 and Roche PCR Amplicor monitor version 1.5. AB - Quantification of human immunodeficiency virus type 1 (HIV-1) RNA as a measure of viral load has greatly improved the monitoring of therapies for infected individuals. With the significant reductions in viral load now observed in individuals treated with highly active anti-retroviral therapy (HAART), viral load assays have been adapted to achieve greater sensitivity. Two commercially available ultrasensitive assays, the Bayer Quantiplex HIV-1 bDNA version 3.0 (bDNA 3.0) assay and the Roche Amplicor HIV-1 Monitor Ultrasensitive version 1.5 (Amplicor 1.5) assay, are now being used to monitor HIV-1-infected individuals. Both of these ultrasensitive assays have a reported lower limit of 50 HIV-1 RNA copies/ml and were developed from corresponding older generation assays with lower limits of 400 to 500 copies/ml. However, the comparability of viral load data generated by these ultrasensitive assays and the relative costs of labor, disposables, and biohazardous wastes were not determined in most cases. In this study, we used matched clinical plasma samples to compare the quantification of the newer bDNA 3.0 assay with that of the older bDNA 2.0 assay and to compare the quantification and costs of the bDNA 3.0 assay and the Amplicor 1.5 assay. We found that quantification by the bDNA 3.0 assay was approximately twofold higher than that by the bDNA 2.0 assay and was highly correlated to that by the Amplicor 1.5 assay. Moreover, cost analysis based on labor, disposables, and biohazardous wastes showed significant savings with the bDNA 3.0 assay as compared to the costs of the Amplicor 1.5 assay. PMID- 10699006 TI - Genome-sequence-based fluorescent amplified-fragment length polymorphism analysis of Mycobacterium tuberculosis. AB - The whole-genome fingerprinting technique, fluorescent amplified-fragment length polymorphism (FAFLP) analysis, was applied to Mycobacterium tuberculosis. Sixty five clinical isolates were analyzed to determine the value of FAFLP as a stand alone genotyping technique and to compare it with the well-established IS6110 typing system. The genome sequence of M. tuberculosis strain H37Rv (S. T. Cole et al., Nature 393:537-544, 1998) was used to model computer-generated informative primer combination(s), and the precision and reproducibility of FAFLP were evaluated by comparing the results of in vitro and computer-generated experiments. Multiplex FAFLP was used to increase resolving power in a predictable and systematic fashion. FAFLP analysis was broadly congruent with IS6110 typing for those strains with multiple IS6110 copies. It was also able to resolve an epidemiologically unlinked group of strains with only one copy of IS6110; up to 10% of clinical isolates may fall into this category. For certain epidemiological investigations, it was concluded that a combination of FAFLP and IS6110 typing would give higher resolution than would either alone. FAFLP data were digital, precise, reproducible, and suitable for rapid electronic dissemination, manipulation, interlaboratory comparison, and storage in national or international epidemiological databases. Because FAFLP samples and analyzes base substitution across the genome as a whole, FAFLP could generate new information about the microevolution of the M. tuberculosis complex. PMID- 10699007 TI - Evaluation of autoSCAN-W/A and the Vitek GNI+ AutoMicrobic system for identification of non-glucose-fermenting gram-negative bacilli. AB - The autoSCAN-W/A (W/A; Dade Behring Microscan Inc., West Sacramento, Calif.) and Vitek AutoMicrobic System (Vitek AMS; bioMerieux Vitek Systems, Inc., Hazelwood, Mo.) are both fully automated microbiology systems. We evaluated the accuracy of these two systems in identifying nonglucose-fermenting gram-negative bacilli. We used the W/A with conventional-panel Neg Combo type 12 and Vitek GNI+ identification systems. A total of 301 isolates from 25 different species were tested. Of these, 299 isolates were identified in the databases of both systems. The conventional biochemical methods were used for reference. The W/A correctly identified 215 isolates (71. 4%) to the species level at initial testing with a high probability of >/=85%. The Vitek GNI+ correctly identified 216 isolates (71.8%) to the species level at initial testing with a high probability of >/=90%. After additional testing that was recommended by the manufacturer's protocol, the correct identifications of the W/A and Vitek GNI+ improved to 96.0 and 92.3%, respectively. The major misidentified species were Sphingomonas paucimobilis and Agrobacterium radiobacter in the W/A system and Acinetobacter lwoffii, Chryseobacterium indologenes, and Comamonas acidovorans in the Vitek GNI+ system. The error rates were 4.0 and 7.6%, respectively. The overall accuracy for both systems was above 90% if the supplemental tests were applied. There was no significant difference in accuracy (P > 0.05) between the two systems. PMID- 10699008 TI - Detection of "Candidatus Helicobacter suis" in gastric samples of pigs by PCR: comparison with other invasive diagnostic techniques. AB - Recently, a new 16S ribosomal DNA-based PCR assay was developed for the specific detection of "Candidatus Helicobacter suis" (former "Gastrospirillum suis") in porcine gastric samples. In the present study, this PCR assay was compared to three other invasive diagnostic methods (rapid urease test, immunohistochemistry, histologic analysis by Giemsa staining). Antral stomach samples from 200 slaughterhouse pigs from Belgium and The Netherlands were examined. Bacterial presence was determined in 77% (154 of 200) of the samples by PCR in combination with Southern blot hybridization, 56% (111 of 200) of the samples by immunohistochemistry, 61% (122 of 200) of the samples by urease testing (20 h postinoculation [p.i.]), 36% (71 of 200) of the samples by urease testing (3 h p.i.), and 33% (65 of 200) of the samples by Giemsa staining. The intrinsic specificity of the PCR assay was assessed by Southern blot analysis with an "Candidatus H. suis"-specific probe and sequencing of PCR products. Interassay sensitivity and specificity values were assessed for each test by pairwise comparisons between tests. Agreement between tests was evaluated by calculating Cohen's kappa coefficient. From that analysis, the PCR assay was considered the most reliable benchmark. Microscopic detection of immunohistochemically labeled or Giemsa-stained "Candidatus H. suis" cells in stomach sections proved to be highly specific (100%) but relatively insensitive (72 and 42%, respectively) compared to the PCR assay. A longer incubation time of the urease test improved its sensitivity considerably (74 versus 55%) but was accompanied by a loss of specificity (72 versus 93%). In conclusion, we found the "Candidatus H. suis" specific PCR assay to be a sensitive and reliable diagnostic method for the detection of "Candidatus H. suis" in the stomachs of pigs and could prove to be a valuable tool for further epidemiological studies both for "Candidatus H. suis"- and for "Helicobacter heilmannii" type 1-related research. PMID- 10699009 TI - Molecular characterization of Staphylococcus sciuri strains isolated from humans. AB - We previously characterized over 100 Staphylococcus sciuri isolates, mainly of animal origin, and found that they all carried a genetic element (S. sciuri mecA) closely related to the mecA gene of methicillin-resistant Staphylococcus aureus (MRSA) strains. We also found a few isolates that carried a second copy of the gene, identical to MRSA mecA. In this work, we analyzed a collection of 28 S. sciuri strains isolated from both healthy and hospitalized individuals. This was a relatively heterogeneous group, as inferred from the different sources, places, and dates of isolation and as confirmed by pulsed-field gel electrophoresis analysis. All strains carried the S. sciuri mecA copy, sustaining our previous proposal that this element belongs to the genetic background of S. sciuri. Moreover, 46% of the strains also carried the MRSA mecA copy. Only these strains showed significant levels of resistance to beta-lactams. Strikingly, the majority of the strains carrying the additional MRSA mecA copy were obtained from healthy individuals in an antibiotic-free environment. Most of the 28 strains were resistant to penicillin, intermediately resistant to clindamycin, and susceptible to tetracycline, erythromycin, and gentamicin. Resistance to these last three antibiotics was found in some strains only. The findings reported in this work confirmed the role of S. sciuri in the evolution of the mechanism of resistance to methicillin in staphylococci and suggested that this species (like the pathogenic staphylococci) may accumulate resistance markers for several classes of antibiotics. PMID- 10699011 TI - A simplified method for testing Bordetella pertussis for resistance to erythromycin and other antimicrobial agents. AB - Present methods of antimicrobial susceptibility testing of Bordetella pertussis are time consuming and require specialized media that are not commercially available. We tested 52 isolates of B. pertussis for resistance to erythromycin, trimethoprim-sulfamethoxazole, chloramphenicol, and rifampin by agar dilution with Bordet-Gengou agar (BGA) containing 20% horse blood (reference method), Etest using BGA and Regan-Lowe agar without cephalexin (RL-C), and disk diffusion using BGA and RL-C. The organisms tested included four erythromycin-resistant isolates of B. pertussis from a single patient, a second erythromycin-resistant strain of B. pertussis from an unrelated patient in another state, and 47 nasopharyngeal surveillance isolates of B. pertussis from children in the western United States. The results of agar dilution testing using direct inoculation of the organisms suspended in Mueller-Hinton broth were within +/-1 dilution of those obtained after overnight passage of the inoculum in Stainer-Scholte medium, which is the traditional method of testing B. pertussis. The Etest method produced MICs similar to those of the agar dilution reference method for three of the four antimicrobial agents tested; the trimethoprim-sulfamethoxazole results were lower with Etest, particularly when the direct suspension method was used. Most of the Etest MICs, except for that of erythromycin, were on scale. Disk diffusion testing using RL-C medium was helpful in identifying the erythromycin resistant strains, which produced no zone of inhibition around the disk; susceptible isolates produced zones of at least 42 mm. Thus, the antimicrobial susceptibility testing of B. pertussis can be simplified by using the Etest or disk diffusion on RL-C to screen for erythromycin-resistant isolates of B. pertussis. PMID- 10699010 TI - Recombinant antigens to detect Toxoplasma gondii-specific immunoglobulin G and immunoglobulin M in human sera by enzyme immunoassay. AB - We have evaluated the diagnostic utility of eleven Toxoplasma gondii recombinant antigens (P22 [SAG2], P24 [GRA1], P25, P28 [GRA2], P29 [GRA7], P30 [SAG1], P35, P41 [GRA4], P54 [ROP2], P66 [ROP1], and P68) in immunoglobulin G (IgG) and IgM recombinant enzyme-linked immunosorbent assays (Rec-ELISAs). Following an initial evaluation, six recombinant antigens (P29, P30, P35, P54, P66, and P68) were tested in the IgG and IgM Rec-ELISAs with four groups of samples which span the toxoplasmosis disease spectrum (negative, chronic infection, acute infection, and recent seroconversion). Our results suggest that the combination of P29, P30, and P35 in an IgG Rec-ELISA and the combination of P29, P35, and P66 in an IgM Rec ELISA can replace the tachyzoite antigen in IgG and IgM serologic tests, respectively. The relative sensitivity, specificity, and agreement for the IgG P29-P30-P35 Rec-ELISA were 98.4, 95.7, and 97.2%, respectively. The resolved sensitivity, specificity, and agreement for the IgM P29-P35-P66 Rec-ELISA were 93.1, 95.0, and 94. 5%, respectively. Relative to the tachyzoite-based immunocapture IgM assay, the IgM P29-P35-P66 Rec-ELISA detects fewer samples that contain IgG antibodies with elevated avidity from individuals with an acute toxoplasmosis. PMID- 10699012 TI - Efficacy of antimicrobial treatments and vaccination regimens for control of porcine reproductive and respiratory syndrome virus and Streptococcus suis coinfection of nursery pigs. AB - Seventy-six, crossbred, porcine reproductive and respiratory syndrome virus (PRRSV)-free pigs were weaned at 12 days of age and randomly assigned to seven groups of 10 to 11 pigs each. Pigs in group 1 served as unchallenged controls. Pigs in groups 2 to 7 were challenged intranasally with 2 ml of high-virulence PRRSV isolate VR-2385 (10(4.47) 50% tissue culture infective doses per 2 ml) on day 0 of the study (30 days of age). Seven days after PRRSV challenge, pigs in groups 2 to 7 were challenged intranasally with 2 ml of Streptococcus suis serotype 2 (10(8.30) CFU/2 ml). Group 2 pigs served as untreated positive controls. Antimicrobial treatments included daily intramuscular injection with 66,000 IU of procaine penicillin G per kg of body weight on days 8 to 10 (group 3), drinking water medication with 23.1 mg of tiamulin per kg during days 8 to 10 (group 4), and daily intramuscular injection of 5.0 mg of ceftiofur hydrochloride per kg on days 8 to 10 (group 5). Vaccination regimens included two intramuscular doses of an autogenous killed S. suis vaccine (group 6) prior to S. suis challenge or a single 2-ml intramuscular dose of an attenuated live PRRSV vaccine (group 7) 2 weeks prior to PRRSV challenge. Mortality was 0, 63, 45, 54, 9, 40, and 81% in groups 1 to 7, respectively. Ceftiofur treatment was the only regimen that significantly (P < 0. 05) reduced mortality associated with PRRSV and S. suis coinfection. The other treatments and vaccinations were less effective. We conclude that ceftiofur administered by injection for three consecutive days following S. suis challenge was the most effective regimen for minimizing disease associated with PRRSV and S. suis coinfection. PMID- 10699013 TI - Comparison of a new neuraminidase detection assay with an enzyme immunoassay, immunofluorescence, and culture for rapid detection of influenza A and B viruses in nasal wash specimens. AB - The performance of a new, rapid, easy-to-perform assay based on neuraminidase enzyme activity for detection of influenza virus types A and B was compared to detection by culture, indirect immunofluorescence, and enzyme immunoassay in 479 nasal wash specimens from children with respiratory infections. Compared to isolation of influenza virus by culture, the neuraminidase assay had a sensitivity of 70.1%, specificity of 92.4%, positive predictive value of 76.3%, and negative predictive value of 89.9%. There was a higher sensitivity for the detection of influenza A virus (76.4%) than for influenza B virus (40.9%). Indirect immunofluorescence showed a sensitivity of 59.8% and specificity of 97% compared to culture isolation for detection of influenza A and B viruses. Enzyme immunoassay showed a sensitivity of 89.7% and specificity of 98.1% for the detection of influenza A alone. The quality of the nasal wash specimen had a significant effect on the detection of influenza virus by all of the assays. A strong response of the neuraminidase assay was more likely to represent a culture confirmed influenza infection. This new rapid neuraminidase assay was useful for the detection of influenza A and B viruses in nasal wash specimens. PMID- 10699014 TI - Single-tube balanced heminested PCR for detecting Mycobacterium tuberculosis in smear-negative samples. AB - In order to achieve more sensitive and specific results for the rapid diagnosis of tuberculosis, we have developed a new method, named balanced heminested PCR, which avoids the inconvenience of asymmetric amplification and has the advantages of single-tube heminested PCR. This was achieved by replacing the outer primer that participates in both rounds of amplification in the standard heminested technique by another primer containing the sequence of the inner primer attached at its 5' end. When both techniques were tested for the IS6110 target of Mycobacterium tuberculosis complex in 80 smear-negative culture-positive sputum samples and 60 control samples, the results showed 100% specificity for both techniques and sensitivities of 60 and 75% for heminested PCR and balanced heminested PCR, respectively (P = 0.02). In conclusion, the balanced heminested technique shows a higher sensitivity than that of the standard heminested, and it could be applied to any PCR by attaching the inner primer at the 5' end of the opposite outer primer. Thus, the balanced heminested technique provides a target for the inner primer in both strands, avoiding asymmetric amplification and thereby resulting in a more efficient amplification, and, in practice, a higher sensitivity without loss of specificity and with a minimum risk of cross contamination. PMID- 10699015 TI - Comparison of classic and molecular approaches for the identification of untypeable enteroviruses. AB - Members of the family Picornaviridae are the most common viruses infecting humans, and species in several genera also infect a wide variety of other mammals. Picornaviruses have traditionally been classified by antigenic type, based on a serum neutralization assay. However, this method is time-consuming and labor-intensive, is sensitive to virus aggregation and antigenic variation, and requires a large number of antisera to identify all serotypes, even when antiserum pools are used. We developed generic reverse transcription (RT)-PCR primers that will amplify all human enterovirus serotypes, as well as many rhinoviruses and other picornaviruses, and used RT-PCR amplification of the VP1 gene and amplicon sequencing to identify enteroviruses that were refractory to typing by neutralization with pooled antisera. Enterovirus serotypes determined by sequencing were confirmed by neutralization with monospecific antisera. Of 55 isolates tested, 49 were of known enterovirus serotypes, two were rhinoviruses, and four were clearly picornaviruses but did not match any known picornavirus sequence. All four untyped picornaviruses were closely related to one another in sequence, suggesting that they are of the same serotype. RT-PCR, coupled with amplicon sequencing, is a simple and rapid method for the typing and classification of picornaviruses and may lead to the identification of many new picornavirus serotypes. PMID- 10699016 TI - Species identification and subtyping of Ureaplasma parvum and Ureaplasma urealyticum using PCR-based assays. AB - There is good evidence that the organism currently known as Ureaplasma urealyticum should be divided into two species-U. parvum (previously U. urealyticum biovar 1) and U. urealyticum (previously U. urealyticum biovar 2). In this study, we designed a series of primers, targeting the 16S rRNA gene and 16S rRNA-23S rRNA intergenic spacer regions, the urease gene subunits, and the 5' ends of the multiple-banded antigen (MBA) genes, to identify and subtype these Ureaplasma species. All of the species-specific primer pairs could distinguish the two species, but only subtype-specific primer pairs targeting the MBA genes could distinguish subtypes within each species. U. parvum was separated into three subtypes, represented by serovars 1, 3/14, and 6. U. urealyticum was also separated into three subtypes by PCR and/or direct sequencing. Subtype 1 consisted of serovars 2, 5, 8, and 9; subtype 2 contained serovars 4, 10, 12, and 13; and subtype 3 contained serovars 7 and 11. A selection of primer pairs was used to identify and subtype 78 clinical ureaplasma isolates from vaginal swabs of pregnant women and to identify and subtype ureaplasmas directly in 185 vaginal swabs in which they had been previously detected. U. parvum was identified in 228 (87%) of 263 isolates or specimens, and U. urealyticum was identified in 50 (19%) (both were present in 6%). Serovars 3/14 (48%) and 1 (43%) were most common among U. parvum isolates, and subtypes 2 (62%) and 1 (34%) were most common among U. urealyticum isolates. This new PCR-based typing system will facilitate future studies of the relationship between individual Ureaplasma species or subtypes and human disease. PMID- 10699017 TI - Molecular characterization of Cryptosporidium isolates obtained from human immunodeficiency virus-infected individuals living in Switzerland, Kenya, and the United States. AB - A total of 22 Cryptosporidium isolates from human immunodeficiency virus-infected patients from Kenya, Switzerland, and the United States were examined at three genetic loci: the 18S ribosomal DNA, HSP-70, and acetyl coenzyme A synthetase genes. Four distinct Cryptosporidium genotypes were identified: (i) the Cryptosporidium parvum "human" genotype, (ii) the C. parvum "cattle" genotype, (iii) Cryptosporidium felis, and (iv) Cryptosporidium meleagridis. This is the first report of C. meleagridis in a human host. These results and those of others indicate that immunocompromised individuals are susceptible to a wide range of Cryptosporidium species and genotypes. Future studies are required to understand the full public health significance of Cryptosporidium genotypes and species in immunocompromised hosts. PMID- 10699018 TI - Plasmodium falciparum histidine-rich protein 2-based immunocapture diagnostic assay for malaria: cross-reactivity with rheumatoid factors. AB - Recently introduced rapid nonmicroscopic immunocapture assays for the diagnosis of malaria infection are being evaluated for their sensitivity and specificity in various epidemiological settings. A Plasmodium falciparum histidine-rich protein 2 (PfHRP-2)-based assay (ICT) and a Plasmodium-specific lactate dehydrogenase test (OptiMAL) were evaluated for their specificities in different groups of patients who tested negative for malaria infection by microscopy. The patients were selected from different disease groups: rheumatoid arthritis, hepatitis C, toxoplasmosis, schistosomiasis, and hydatid disease. One hundred thirty-three of the 225 patients were positive for rheumatoid factor. Thirty-five of the 133 (26%) rheumatoid factor-positive patients gave a false-positive reaction with the ICT assay, but only 4 of these gave false-positive reactions with the OptiMAL test. Thirty-three of the 35 false-positive specimens became negative when repeat tested with the ICT assay after absorbing out the rheumatoid factor activity. Our study shows that the PfHRP-2-based ICT assay gave a false-positive reaction in 26% of the patients who had rheumatoid factors, but were negative for malaria by microscopy. PMID- 10699019 TI - Novel screening method for urine cultures using a filter paper dilution system. AB - We have developed a novel method for urine culture for office practice based on the use of filter paper as a solid-phase dilution device. Filtration dilutes and spreads the inoculum onto a solid culture surface. Experiments were conducted to determine the optimum inoculum size, microbial permeability through filter papers, and ability to exclude vaginal epithelial cells. The filter paper dilution system was compared to the standard streak method to detect bacteriuria in specimens submitted to the diagnostic laboratory. The sensitivity and specificity of the filter paper dilution system for detection of high-count (>/=10(4) CFU/ml) gram-negative bacteriuria in 487 urine specimens were 98.2 and 97.4%, respectively. The sensitivity and specificity for gram-positive bacteriuria in 404 urine specimens were 91.2 and 99.2%, respectively. Low-count gram-negative bacteriuria (<10(4) CFU/ml) was detected by the filter paper dilution system in five of nine specimens (55.6%). In addition, the filter paper dilution system was able to detect gram-negative bacteria in 12 of 41 (29.3%) mixed cultures. Lactobacillus and Gardnerella organisms in urine specimens were excluded by the filter paper dilution system. Only three of eight Candida sp. isolates were detected at counts of >/=10(4) CFU/ml. The system has good storage properties and can be inoculated at the point of source without the need for refrigeration or preservatives. It should be a useful screening method for office practice, where members of the family Enterobacteriaceae and staphylococci cause most infections. Standard culture methods are preferred for hospital diagnostic microbiology laboratories, where there is a need to detect yeasts and fastidious microorganisms and to isolate individual colonies from mixed cultures. PMID- 10699020 TI - Detection and identification of human parainfluenza viruses 1, 2, 3, and 4 in clinical samples of pediatric patients by multiplex reverse transcription-PCR. AB - We describe a multiplex reverse transcription-PCR (m-RT-PCR) assay that is able to detect and differentiate all known human parainfluenza viruses (HPIVs). Serial dilution experiments with reference strains that compared cell culture isolation and m-RT-PCR showed sensitivities ranging from 0.0004 50% tissue culture infective dose (TCID(50)) for HPIV type 4B (HPIV-4B) to 32 TCID(50)s for HPIV-3. As few as 10 plasmids containing HPIV PCR products could be detected in all cases. When 201 nasopharyngeal aspirate specimens from pediatric patients hospitalized for lower respiratory illness were tested, m-RT-PCR assay detected 64 HPIVs (24 HPIV-3, 23 HPIV-1, 10 HPIV-4, and 7 HPIV-2), while only 42 of them (21 HPIV-1, 14 HPIV-3, 6 HPIV-2, and 1 HPIV-4 isolates) grew in cell culture. Our m-RT-PCR assay was more sensitive than either cell culture isolation or indirect immunofluorescence with monoclonal antibodies for the detection of HPIV infections. Also, HPIV-4 was more frequently detected than HPIV-2 in this study, suggesting that it may have been underestimated as a lower respiratory tract pathogen because of the insensitivity of cell culture. PMID- 10699021 TI - Acquisition and colonization stability of Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis in children. AB - The presence of Porphyromonas gingivalis has been shown to be a risk factor for periodontitis in adults, and Actinobacillus actinomycetemcomitans has been implicated as a pathogen in early-onset periodontitis. Both species have been shown to establish stable colonization in adults. In cross-sectional studies, both A. actinomycetemcomitans and P. gingivalis have been detected in over one third of apparently healthy children. Information on the stability of colonization with these organisms in children could help to elucidate the natural history of the development of periodontitis. For this purpose, samples previously collected from a cohort of 222 children between the ages of 0 and 18 years and previously examined for the presence of P. gingivalis with a PCR-based assay were examined for the presence of A. actinomycetemcomitans. It was detected in 48% of subjects and, like P. gingivalis, was found at similar frequencies among children of all ages (P = 0.53), suggesting very early initial acquisition. One hundred one of the original subjects were recalled after 1 to 3 years to determine the continuing presence of both A. actinomycetemcomitans and P. gingivalis. The prevalence of both species remained unchanged at resampling. However, in most children both species appeared to colonize only transiently, with random concordance between the results of the first and second sampling. Stability of colonization was unrelated to age for A. actinomycetemcomitans, but P. gingivalis was more stable in the late teenage years. PMID- 10699023 TI - Rapid, efficient detection and drug susceptibility testing of Mycobacterium tuberculosis in sputum by microscopic observation of broth cultures. The Tuberculosis Working Group in Peru. AB - Inexpensive, rapid, and reliable methods of detecting infection by and drug susceptibility of Mycobacterium tuberculosis (MTB) are crucial to the control of tuberculosis. The novel microscopic observation broth-drug susceptibility assay (MODS) detects early growth of MTB in liquid medium, allowing more timely diagnosis and drug susceptibility testing. Sputum samples from hospitalized patients in Peru were analyzed by using stains, culture, and PCR. Sensitivity of MODS (92%) compared favorably with the most sensitive of the other culture methods (93%). Sputum samples positive for tuberculosis were tested for susceptibility to isoniazid and rifampin with the microwell alamar blue assay (MABA) and MODS. In 89% of cases, there was concordance between MODS and MABA. Of the diagnostic and susceptibility testing methods used, MODS yielded results most rapidly (median, 9.0 and 9.5 days, respectively). MODS is a rapid, inexpensive, sensitive, and specific method for MTB detection and susceptibility testing; it is particularly appropriate for use in developing countries burdened by significant infection rates and increasing numbers of multiple-drug-resistant cases. PMID- 10699022 TI - Isolation and molecular characterization of Clostridium difficile strains from patients and the hospital environment in Belarus. AB - Toxigenic Clostridium difficile is the most common etiologic agent of hospital acquired diarrhea in developed countries. The role of this pathogen in nosocomial diarrhea in Eastern Europe has not been clearly established. The goal of this study was to determine the prevalence of C. difficile in patients and the hospital environment in Belarus and to characterize these isolates as to the presence of toxin genes and their molecular type. C. difficile was isolated from 9 of 509 (1.8%) patients analyzed and recovered from 28 of 1,300 (2. 1%) environmental sites cultured. A multiplex PCR assay was used to analyze the pathogenicity locus (PaLoc) of all isolates, and strain identity was determined by an arbitrarily primed PCR (AP-PCR). The targeted sequences for all the genes in the PaLoc were amplified in all C. difficile strains examined. A predominantly homogeneous group of strains was found among these isolates, with five major AP PCR groups being identified. Eighty-three percent of environmental isolates were classified into two groups, while patient isolates grouped into three AP-PCR types, two of which were also found in the hospital environment. Although no data on the role of C. difficile infection or epidemiology of C. difficile-associated diarrhea (CDAD) in this country exist, the isolation of toxigenic C. difficile from the hospital environment suggests that this pathogen may be responsible for cases of diarrhea of undiagnosed origin and validates our effort to further investigate the significance of CDAD in Eastern Europe. PMID- 10699024 TI - Comparison of two commercial methods for measurement of cytomegalovirus load in blood samples after renal transplantation. AB - A cohort of 77 renal transplant recipients was prospectively studied for comparison of two commercially available cytomegalovirus (CMV) load assays, i.e., the COBAS AMPLICOR CMV Monitor test (Amplicor), using plasma samples, and the Murex Hybrid Capture System (HCS), using whole blood. The manufacturer of the HCS assay changed the version of the test from 1.0 (HCS-1) to 2.0 (HCS-2) after the first 37 patients had been tested. Despite the differences in principle and type of specimen used, the Amplicor and HCS assays gave comparable results. The regression line correlating the HCS-1 assay to the Amplicor assay was similar to that correlating the HCS-2 assay to the Amplicor assay. The HCS results could be converted to Amplicor-equivalent units by using linear-regression equations [log(10) HCS-1 result = 0.49 (log(10) Amplicor result) + 2.58, and log(10) HCS-2 result = 0.61 (log(10) Amplicor result) + 2.18]. The HCS-2 assay appeared to have the lowest detection limit, followed by the Amplicor assay and then the HCS-1 assay. When a sliding scale of cutoff values in Amplicor-equivalent units (>1,000, >2,500, >6,000, >16,000, >40,000, and >100,000 copies/ml) was applied to diagnose CMV disease, similar patterns of sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) were observed with the Amplicor and HCS assays. A cutoff value of >40,000 copies/ml has a low sensitivity (Amplicor, 29.4%; HCS, 41.2%) but is specific (Amplicor, 96.7%; HCS, 93.3%) and can be used for the differential diagnosis of CMV disease (PPV, 71.4% [Amplicor] or 63. 6% [HCS]; NPV, 82.9% [Amplicor] or 84.8% [HCS]). A lower cutoff value of >1,000 copies/ml improves the sensitivity (Amplicor, 76.5%; HCS, 82.4%) and has a high NPV (Amplicor, 91.8%; HCS, 94.2%) but, due to the low PPV (Amplicor, 46.2%; HCS, 56%), is useful only for exclusion of CMV disease. PMID- 10699025 TI - Clonal and spontaneous origins of fluconazole resistance in Candida albicans. AB - The genotypes and susceptibilities to fluconazole of 78 strains of the human pathogenic yeast Candida albicans were compared. The strains comprised two sets of samples from Durham, N.C.: one from patients infected with the human immunodeficiency virus (HIV) and the other from healthy volunteers. For each strain, the MIC of fluconazole was determined by the standard National Committee for Clinical Laboratory Standards protocol. Genotypes were determined by PCR fingerprinting with five separate primers. The analysis revealed little evidence for genotypic clustering according to HIV status or body site. However, a small group of fluconazole-resistant strains isolated from patients infected with HIV formed a distinct cluster. In addition, two fluconazole-resistant strains were isolated from individuals who never took fluconazole, one from a patient infected with HIV and the other from a healthy person. The results suggest both clonal and spontaneous origins of fluconazole resistance in C. albicans. PMID- 10699027 TI - Two liquid medium systems, mycobacteria growth indicator tube and MB redox tube, for Mycobacterium tuberculosis isolation from sputum specimens. AB - Two manual liquid medium systems, the Mycobacteria Growth Indicator Tube (MGIT) and MB Redox tube systems, were evaluated in comparison to the radiometric BACTEC 460 semiautomated system for recovery of Mycobacterium tuberculosis from sputum specimens. The highest level of recovery, from a total of 77 culture-positive specimens, occurred with the BACTEC-460 system (92.2%), followed by the MB Redox tube (80.5%) and the MGIT (63.6%) systems. The shortest time to detection was observed also among the cultures in BACTEC-460: a mean of 12 days to a growth index (GI) of 10 and 15 days to a GI of 500. The mean times for the other systems were 16 days for the MB Redox tube system and 17.4 days for the MGIT system. The proportion of cultures grown after more than 3 weeks of incubation was only 2.8 or 8.4% in BACTEC-460 (for a GI of 10 or 500) but 17.7% in MB Redox and 22.5% in MGIT. Despite these differences in comparison to the BACTEC-460 system and some differences between the MGIT and MB Redox tube systems, either of the two manual liquid medium systems presents a reasonable alternative to the BACTEC-460 system, especially for laboratories with a limited workload, and a valuable element in the laboratory protocol, in conjunction with solid media, for obtaining rapid detection of growth from about 80% of culture-positive specimens and for better overall recovery of M. tuberculosis. PMID- 10699026 TI - Diagnostic and public health dilemma of lactose-fermenting Salmonella enterica serotype Typhimurium in cattle in the Northeastern United States. AB - The presence of lactose-fermenting Salmonella strains in clinical case materials presented to microbiology laboratories presents problems in detection and identification. Failure to detect these strains also presents a public health problem. The laboratory methods used in detecting lactose-fermenting Salmonella enterica serotype Typhimurium from six outbreaks of salmonellosis in veal calves are described. Each outbreak was caused by a multiply-resistant and lactose fermenting strain of S. enterica serotype Typhimurium. The use of Levine eosin methylene blue agar in combination with screening of suspect colonies for C8 esterase enzyme and inoculation of colonies into sulfide-indole-motility medium for hydrogen sulfide production was particularly effective for their detection. A hypothesis for the creation of lactose-fermenting salmonellae in the environment is presented. It is proposed that the environment and husbandry practices of veal raising barns provide a unique niche in which lactose-fermenting salmonellae may arise. PMID- 10699028 TI - Detection of a previously unamplified spacer within the DR locus of Mycobacterium tuberculosis: epidemiological implications. AB - Spoligotyping, a method based on the variability of distribution of the 43 inter direct repeat (DR) spacers of Mycobacterium tuberculosis and Mycobacterium bovis BCG, is useful to study the molecular epidemiology of bovine and human tuberculosis. Recently, a major family of M. tuberculosis clinical isolates named the Haarlem family, which did not contain spacers 31 and 33 to 36, was reported in a multicenter study. Independently, a data bank containing all the published spoligotypes showed that the two most prevalent spoligotypes in the world differed only by the presence or absence of spacer 31. A careful analysis of the DR locus sequence led us to hypothesize that spacer 31 may not have been amplified in some isolates with the primer sets DRa and DRb currently used for spoligotyping. Consequently, a modified spoligotyping method based on different combinations of the 36-bp DR and IS6110 primers was devised that was able to discriminate between the left and the right parts of the DR locus and demonstrated the presence of the previously unamplified spacer 31 for some of the clinical isolates. By analogy, we suggest that a single-spacer difference in some epidemiologically linked cases of tuberculosis may simply arise due to the insertion of an extra copy of IS6110 within the DR locus, leading to its asymmetrical disruption and subsequent lack of the DRa or DRb targets. The influence of the IS6110 preferential insertion sites within the DR locus on spoligotyping results should be further investigated. PMID- 10699030 TI - Growth of Cowdria ruminantium, the causative agent of heartwater, in a tick cell line. AB - The tick-borne rickettsia Cowdria ruminantium has been propagated continuously for over 500 days in the Ixodes scapularis tick cell line IDE8 by using the Gardel isolate from bovine endothelial cells as an inoculum. Infection of the tick cells was confirmed by PCR, karyotyping, electron microscopy, and reinfection of bovine cells. PMID- 10699029 TI - Molecular identification and epidemiological tracing of Pasteurella multocida meningitis in a baby. AB - We report a case of Pasteurella multocida meningitis in a 1-month-old baby exposed to close contact with two dogs and a cat but without any known history of injury by these animals. 16S rRNA gene sequencing of the isolate from the baby allowed identification at the subspecies level and pointed to the cat as a possible source of infection. Molecular typing of Pasteurella isolates from the animals, from the baby, and from unrelated animals clearly confirmed that the cat harbored the same P. multocida subsp. septica strain on its tonsils as the one isolated from the cerebrospinal fluid of the baby. This case stresses the necessity of informing susceptible hosts at risk of contracting zoonotic agents about some basic hygiene rules when keeping pets. In addition, this study illustrates the usefulness of molecular methods for identification and epidemiological tracing of Pasteurella isolates. PMID- 10699031 TI - Development of a hypersensitive detection method for human parvovirus B19 DNA. AB - A new detection method for human parvovirus B19 DNA was established using PCR coupled with a hybridization protection assay. The amplified product was detected using acridinium ester-labeled DNA probes. By this method, a few copies of B19 DNA were detected in human serum albumin. PMID- 10699032 TI - Etiology of sexually transmitted infections among street-based female sex workers in Dhaka, Bangladesh. AB - An etiological study of sexually transmitted infections (STIs) was conducted among female sex workers (FSWs) in Dhaka, Bangladesh. Endocervical swab and blood samples from 269 street-based FSWs were examined for Neisseria gonorrhoeae, Chlamydia trachomatis, and Trichomonas vaginalis as well as for antibodies to Treponema pallidum and herpes simplex virus 2 (HSV-2). Sociodemographic data and data regarding behavior were also collected. A total of 226 of the 269 FSWs (84%) were positive for the STI pathogens studied. Among the 269 FSWs, 35.5% were positive for N. gonorrhoeae, 25% were positive for C. trachomatis, 45.5% were positive for T. vaginalis, 32.6% were seropositive for T. pallidum, 62.5% were seropositive for HSV-2, and 51% had infections with two or more pathogens. PMID- 10699033 TI - Use of calibrated viral load standards for group M subtypes of human immunodeficiency virus type 1 to assess the performance of viral RNA quantitation tests. AB - Optimal management of human immunodeficiency virus type 1 (HIV-1) disease requires accurate quantitation of viral RNA concentrations in plasma. Evidence for increasing geographic intermixing of HIV-1 subtypes makes equivalent quantitation of all subtypes essential. The performances of six quantitative viral RNA tests are described, for the first time, with calibrated viral isolates of diverse subtypes. PMID- 10699034 TI - Epidemiological analysis of non-M-typeable group A Streptococcus isolates from a Thai population in northern Thailand. AB - Infection with group A streptococci (GAS) can lead to the development of severe postinfectious sequelae such as rheumatic fever (RF). In Thailand, RF and rheumatic heart disease (RHD) remain important health problems. More than 80% of GAS circulating in this population are non-M antigen typeable by conventional M serotyping methods. In this study, we determine the M protein sequence types of GAS isolates found in northern Thailand. The emm genes from 53 GAS isolates, collected between 1985 and 1995 from individuals with pharyngitis, impetigo, acute RF (ARF), RHD, or meningitis as well as from individuals without infections, were amplified by PCR and sequenced. Thirteen new sequence types that did not show homology to previously published sequences were characterized. Six of these sequence types could be isolated from both skin and throat sites of impetigo and pharyngitis/ARF patients, respectively. In many cases we could not specifically differentiate skin strains or throat strains that could be associated with ARF or acute glomerulonephritis. Antigenic variations in the emm gene of the isolates investigated, compared to published M protein sequences, were predominantly due to point mutations, small deletions, and insertions in the hypervariable region. One group of isolates with homology to M44 exhibited corrected frameshift mutations. A new M type isolated from an RHD patient exhibited nucleotide sequence corresponding to the N terminus of M58 and the C terminus of M25, suggesting that recombination between the two types may have occurred. This study provided epidemiological data relating to GAS endemic to northern Thailand which could be useful for identification of vaccine candidates in a specific region of endemicity. PMID- 10699035 TI - Rapid discrimination of monkey B virus from human herpes simplex viruses by PCR in the presence of betaine. AB - A PCR method to amplify DNA segments of the glycoprotein G gene of monkey B virus (BV) was achieved by adding betaine to the PCR mixture, in spite of the high G+C content of this gene. No product was obtained when DNA of human herpes simplex viruses (HSVs) was used as the template under the same conditions. Thus, this PCR method is useful in discriminating BV from HSVs. PMID- 10699037 TI - Otitis externa associated with Malassezia sympodialis in two cats. AB - The lipid-dependent species Malassezia sympodialis was isolated from two cats with otitis externa. To our knowledge, this is the first report of the isolation of lipid-dependent species of the genus Malassezia associated with skin disease in domestic animals. PMID- 10699036 TI - Molecular characterization of Brucella strains isolated from marine mammals. AB - Recently, gram-negative bacteria isolated from a variety of marine mammals have been identified as Brucella species by conventional phenotypic analysis. This study found the 16S rRNA gene from one representative isolate was identical to the homologous sequences of Brucella abortus, B. melitensis, B. canis, and B. suis. IS711-based DNA fingerprinting of 23 isolates from marine mammals showed all the isolates differed from the classical Brucella species. In general, fingerprint patterns grouped by host species. The data suggest that the marine mammal isolates are distinct types of Brucella and not one of the classical species or biovars invading new host species. In keeping with historical precedent, the designation of several new Brucella species may be appropriate. PMID- 10699038 TI - Detection of Giardia lamblia and Cryptosporidium parvum antigens in human fecal specimens using the ColorPAC combination rapid solid-phase qualitative immunochromatographic assay. AB - The ColorPAC Giardia/Cryptosporidium (Becton Dickinson) is a solid-phase qualitative immunochromatographic assay that detects and distinguishes between Giardia lamblia and Cryptosporidium parvum in human stool. Agreement between the Alexon-Trend ProSpecT Giardia Rapid EIA and the ColorPAC assay was 166 of 172 (96.5%). Agreement between the Alexon-Trend ProSpecT Cryptosporidium Rapid EIA and the ColorPAC assay was 169 of 171 (98.8%). No cross-reactions were seen with other parasites or human cells. PMID- 10699039 TI - Multiple clones within multidrug-resistant Salmonella enterica serotype Typhimurium phage type DT104. The Greek Nontyphoidal Salmonella Study Group. AB - Six distinct clones were present among Greek multidrug-resistant Salmonella enterica serotype Typhimurium phage type DT104, since isolates belonging to resistance phenotypes including the ACSSuT (ampicillin, chloramphenicol, streptomycin, sulfonamides, and tetracycline) core could be distinguished with respect to their pulsed-field gel electrophoresis patterns, int1 integron structures, and presence or absence of antibiotic resistance genes ant(3'')-Ia, pse-1, and tem-1. PMID- 10699040 TI - Simple latex agglutination assay for rapid serodiagnosis of human leptospirosis. AB - A newly developed latex agglutination assay for the detection of genus-specific Leptospira antibodies in human sera was evaluated. The assay is performed by mixing, on an agglutination card, serum with equal volumes of stabilized antigen coated, dyed test and control latex beads and is read within 2 min. The latex agglutination test was evaluated with groups of serum samples from patients with leptospirosis and control patients from Hawaii, the Seychelles, Thailand, and The Netherlands. The mean overall sensitivity was 82.3%, and the mean overall specificity was 94.6%. The assay is easy to perform and does not require special skills or equipment. The reagents have a long shelf life, even at tropical temperatures. Together, these factors make the assay suitable for use even at the peripheral level of a health care system as a rapid screening test for leptospirosis. PMID- 10699041 TI - Experimental inoculation with human granulocytic Ehrlichia agent derived from high- and low-passage cell culture in horses. AB - We report the successful infection throughout intravenous inoculation with low and high passage of in vitro-grown human granulocytic ehrlichiosis (HGE) agent in horses. Differences in disease severity but not in incubation time, hematological changes, PCR detection, ehrlichial load, seroconversion time, and titer range were noted between horses infected with a low and a high passage of in vitro grown HGE agent. PMID- 10699042 TI - Comparative evaluation of nine different enzyme-linked immunosorbent assays for determination of antibodies against Treponema pallidum in patients with primary syphilis. AB - Nine different enzyme-linked immunosorbent assays (ELISAs) with a sonicate or recombinant proteins of Treponema pallidum as antigen have been evaluated comparatively by testing 52 highly selected sera from patients with primary syphilis, all negative in the microhemagglutination test for T. pallidum (MHA TP). Eight tests exhibited greater sensitivity (48.5 to 76.9%) than the commonly used Venereal Disease Research Laboratory test (44.2%). Higher sensitivity could be related to (i) the volume and dilution of the serum, (ii) the design of the assay (capture and competitive tests showed higher sensitivity than sandwich based assays), and (iii) the ability to detected specific immunoglobulin M antibodies. The specificity of the ICE Syphilis and the Enzygnost Syphilis tests was 99.5 and 99.8%, respectively, as determined by routine testing of 2, 053 unselected sera in comparison with the MHA-TP test. ELISAs tested offered high sensitivity in patients with primary syphilis; however, recommendations to use these tests as screening assays do need further data on specificity and reactivity in late stages of the disease. PMID- 10699043 TI - Report of the first human case of lobomycosis in the United States. AB - We describe the first human case of lobomycosis caused by Lacazia loboi in a 42 year-old white male resident of Georgia. The patient had traveled to Venezuela 7 years earlier, where he had planned to rappel down Angel Falls in Canaima. Although he never actually rappelled the falls, he did walk under the falls at least three times, exposing himself to the high water pressures of the falls. He noticed a small pustule with surrounding erythema developing on the skin of his right chest wall. The lesion gradually increased in size and had an appearance of a keloid. For cosmetic reasons, the patient sought medical treatment to remove the lesion. After an uncomplicated excision of the lesion, the patient recovered completely. The excised tissue was fixed in formalin for pathologic examination. Tissue sections stained by hematoxylin and eosin, periodic acid-Schiff stain, and Gomori methenamine silver stain procedures showed numerous histiocytes, multinucleated giant cells, and numerous globose or subglobose, lemon-shaped cells producing multiple blastoconidia connected by narrow tube-like connectors and catenate chains of various lengths characteristic of L. loboi. PMID- 10699044 TI - Rapid subtyping of dengue virus serotypes 1 and 4 by restriction site-specific PCR. AB - We previously reported a simple subtyping method, restriction site-specific PCR (RSS-PCR), for dengue virus serotypes 2 and 3; here we describe its application for subtyping dengue virus serotypes 1 and 4. Three major RSS-PCR types were observed for dengue virus serotype 1 and two types were observed for dengue virus serotype 4, in agreement with previous strain classifications based on sequence analysis. Because of its simplicity, this method is amenable to rapid subtyping and application to epidemiological studies of dengue in countries where dengue is endemic. PMID- 10699045 TI - Outbreak of infections caused by Pseudomonas aeruginosa producing VIM-1 carbapenemase in Greece. AB - Resistance to imipenem and meropenem was observed in 211 (16.5%) isolates of Pseudomonas aeruginosa recovered in a Greek university hospital during 1996 to 1998. In six isolates selected from throughout this period, high-level resistance to both carbapenems (MICs >/= 128 microg/ml) was associated with production of the class B beta-lactamase VIM-1. bla(VIM)-bearing isolates belonged to serotype O:12 and were indistinguishable by pulsed-field gel electrophoresis. PMID- 10699046 TI - Helminthic transmission and isolation of Ehrlichia risticii, the causative agent of Potomac horse fever, by using trematode stages from freshwater stream snails. AB - We report successful helminthic transmission of Ehrlichia risticii, the causative agent of Potomac horse fever, using trematode stages collected from Juga yrekaensis snails. The ehrlichial agent was isolated from the blood of experimentally infected horses by culture in murine monocytic cells and identified as E. risticii ultrastructurally and by characterization of three different genes. PMID- 10699047 TI - Genetic and antigenic evidence supports the separation of Hepatozoon canis and Hepatozoon americanum at the species level. AB - Recognition of Hepatozoon canis and Hepatozoon americanum as distinct species was supported by the results of Western immunoblotting of canine anti-H. canis and anti-H. americanum sera against H. canis gamonts. Sequence analysis of 368 bases near the 3' end of the 18S rRNA gene from each species revealed a pairwise difference of 13.59%. PMID- 10699048 TI - Potential errors in recognition of Erysipelothrix rhusiopathiae. AB - Here we describe four isolations of Erysipelothrix rhusiopathiae associated with polyarthralgia and renal failure, septic arthritis, classic erysipeloid, and peritonitis. Although the biochemical identification was straightforward in each case, recognition presented a challenge to the clinical microbiologist, since in three cases E. rhusiopathiae was not initially considered due to unusual clinical presentations, in two cases the significance might not have been appreciated because growth was in broth only, and in one case the infection was thought to be polymicrobic. Because the Gram stain can be confusing, abbreviated identification schemes that do not include testing for H(2)S production could allow E. rhusiopathiae isolates to be misidentified as Lactobacillus spp. or Enterococcus spp. in atypical infections. PMID- 10699049 TI - Emergence of serotype G9 human rotaviruses in Australia. PMID- 10699050 TI - Detecting low penetrance genes in cancer: the way ahead. AB - The search for the genes responsible for many complex genetic diseases is well under way and has already been successful in some cases. The study of cancer as a complex genetic disease has lagged behind other conditions, largely because of particular problems that are associated with malignant disease. Cancer also, however, presents specific opportunities for gene identification, which are not found in many other diseases. While the methods of genetic mapping and gene cloning used for other complex diseases will be applied to cancer, these must almost certainly be complemented by other methods, such as the study of somatic mutations, cancer associated phenotypes, and modifier genes for Mendelian cancers. Here, we review the strategies available for identifying cancer predisposition genes of low and moderate penetrance. PMID- 10699051 TI - Diagnostic analysis of the Rubinstein-Taybi syndrome: five cosmids should be used for microdeletion detection and low number of protein truncating mutations. AB - Rubinstein-Taybi syndrome (RTS) is a malformation syndrome characterised by facial abnormalities, broad thumbs, broad big toes, and mental retardation. In a subset of RTS patients, microdeletions, translocations, and inversions involving chromosome band 16p13.3 can be detected. We have previously shown that disruption of the human CREB binding protein (CREBBP or CBP) gene, either by these gross chromosomal rearrangements or by point mutations, leads to RTS. CBP is a large nuclear protein involved in transcription regulation, chromatin remodelling, and the integration of several different signal transduction pathways. Here we report diagnostic analysis of CBP in 194 RTS patients, divided into several subsets. In one case the mother is also suspect of having RTS. Analyses of the entire CBP gene by the protein truncation test showed 4/37 truncating mutations. Two point mutations, one 11 bp deletion, and one mutation affecting the splicing of the second exon were detected by subsequent sequencing. Screening the CBP gene for larger deletions, by using different cosmid probes in FISH, showed 14/171 microdeletions. Using five cosmid probes that contain the entire gene, we found 8/89 microdeletions of which 4/8 were 5' or interstitial. This last subset of microdeletions would not have been detected using the commonly used 3' probe RT1, showing the necessity of using all five probes. PMID- 10699052 TI - Mutation analysis in glutaric aciduria type I. AB - Glutaric aciduria type 1 (GA1), resulting from the genetic deficiency of glutaryl CoA dehydrogenase (GDH), is a relatively common cause of acute metabolic brain damage in infants. Encephalopathic crises may be prevented by carnitine supplementation and diet, but diagnosis can be difficult as some patients do not show the typical excretion of large amounts of glutaric and 3-hydroxyglutaric acids in the urine. We present a rapid and efficient denaturing gradient gel electrophoresis (DGGE) method for the identification of mutations in the glutaryl CoA dehydrogenase (GCDH) gene that may be used for the molecular diagnosis of GA1 in a routine setting. Using this technique, we identified mutations on both alleles in 48 patients with confirmed GDH deficiency, while no mutations were detected in other patients with clinical suspicion of GA1 but normal enzyme studies. There was a total of 38 different mutations; 27 mutations were found in single patients only, and 21 mutations have not been previously reported. Fourteen mutations involved hypermutable CpG sites. The commonest GA1 mutation in Europeans is R402W, which accounts for almost 40% of alleles in patients of German origin. GCDH gene haplotypes were determined through the analysis of polymorphic markers in all families, and three CpG mutations were associated with different haplotypes, possibly reflecting independent recurrence. The high sensitivity of the DGGE method allows the rapid and cost efficient diagnosis of GA1 in instances where enzyme analyses are not available or feasible, despite the marked heterogeneity of the disease. PMID- 10699053 TI - Butyrylcholinesterase K variant is genetically associated with late onset Alzheimer's disease in Northern Ireland. AB - Alzheimer's disease (AD) is a progressive neurodegenerative disorder that has been associated, sometimes controversially, with polymorphisms in a number of genes. Recently the butyrylcholinesterase K variant (BCHE K) allele has been shown to act in synergy with the apolipoprotein E epsilon4 (APOE epsilon4) allele to promote risk for AD. Most subsequent replicative studies have been unable to confirm these findings. We have conducted a case-control association study using a clinically well defined group of late onset AD patients (n=175) and age and sex matched control subjects (n=187) from the relatively genetically homogeneous Northern Ireland population to test this association. The BCHE genotypes of patients were found to be significantly different from controls (chi(2)=23.68, df=2, p<<0.001). The frequency of the K variant allele was also found to differ significantly in cases compared to controls (chi(2)=16.39, df=1, p<<0.001) leading to an increased risk of AD in subjects with this allele (OR=3.50, 95% CI 2. 20-6.07). This risk increased in subjects 75 years and older (OR=5. 50, 95% CI 2.56-11.87). At the same time the APOE epsilon4 associated risk was found to decrease from 6.70 (95% CI 2.40-19.04) in 65-74 year olds to 3.05 (95% CI 1.34 6.95) in those subjects 75 years and older. However, we detected no evidence of synergy between BCHE K and APOE epsilon4. The results from this study suggest that possession of the BCHE K allele constitutes a significant risk for AD in the Northern Ireland population and, furthermore, this risk increases with increasing age. PMID- 10699054 TI - Age and sex based genetic locus heterogeneity in type 1 diabetes. AB - BACKGROUND: Two genome scans for susceptibility loci for type 1 diabetes using large collections of families have recently been reported. Apart from strong linkage in both studies of the HLA region on chromosome 6p, clear consistent evidence for linkage was not observed at any other loci. One possible explanation for this is a high degree of locus heterogeneity in type 1 diabetes, and we hypothesised that the sex of affected offspring, age of diagnosis, and parental origin of shared alleles may be the bases of heterogeneity at some loci. METHODS: Using data from a genome wide linkage study of 356 affected sib pairs with type 1 diabetes, we performed linkage analyses using parental origin of shared alleles in subgroups based on (1) sex of affected sibs and (2) age of diagnosis. RESULTS: Among the results obtained, we observed that evidence for linkage to IDDM4 on chromosome 11q13 occurred predominantly from opposite sex, rather than same sex sib pairs. At a locus on chromosome 4q, evidence for linkage was observed in sibs where one was diagnosed above the age of 10 years and the other diagnosed below 10 years of age. CONCLUSIONS: We show that heterogeneity tests based on age of diagnosis, sex of affected subject, and parental origin of shared alleles may be helpful in reducing locus heterogeneity in type 1 diabetes. If repeated in other samples, these findings may assist in the mapping of susceptibility loci for type 1 diabetes. Similar analyses can be recommended in other complex diseases. PMID- 10699055 TI - A model protocol evaluating the introduction of genetic assessment for women with a family history of breast cancer. AB - Randomised controlled trials allow comparisons to be made between different models of service delivery, but have not been used in the field of clinical genetics. With the advent of clinical governance, the evidence provided by such trials will be increasingly important in informing and shaping clinical genetics practice. The TRACE project (Trial of genetic assessment in breast cancer) is a randomised controlled trial of genetic assessment for women who are at increased risk of breast cancer because of their family history. The absence of cancer genetics service provision in Wales before this study gave a window of opportunity in which this important trial could be conducted. The present paper describes how TRACE will provide crucial evidence regarding the psychosocial as well as resource implications of adding individualised genetic assessment, genetic counselling, and (where appropriate) gene testing to typical advice and surveillance from a hospital breast clinic. In addition, it is anticipated that TRACE will represent a model for future trials of service delivery in the increasing number of complex genetic disorders where evidence on the economic implications of screening and management is currently limited. PMID- 10699056 TI - Why do women attend familial breast cancer clinics? AB - The increasing demand for genetic assessment for familial breast cancer has necessitated the development of cancer genetics services. However, little is known about the factors motivating the client population likely to approach these services. A cross sectional questionnaire survey of 1000 women with a family history of breast cancer was conducted to identify self-reported reasons for attending a familial breast cancer clinic and possible differences in the characteristics of women who were attending for diverse reasons. Before attendance at clinic, 833 women completed a baseline questionnaire (83% response rate). Women who gave personal risk (n=188), awareness of a family history (n=120), risk to family members (n=84), reassurance (n=69), genetic testing (n=65), breast screening (n=46), or prevention (n=39) as their main reason for attending were compared on demographic and medical variables, and on psychological variables including general anxiety, cancer worry, perceived risk, and attitudes towards prophylactic surgery and genetic testing. Important differences in the psychological characteristics of these groups were found, which were unrelated to reported family history. In particular, women who primarily wanted genetic testing felt extremely vulnerable to developing breast cancer, were more likely to be considering prophylactic surgery, and perceived fewer limitations of testing. Those who primarily wanted reassurance were highly anxious about the disease. We recommend that cancer genetics services take into consideration the informational and psychological needs and concerns of their client group. PMID- 10699057 TI - Guidelines for a genetic risk based approach to advising women with a family history of breast cancer. UK Cancer Family Study Group (UKCFSG). AB - A family history of breast cancer has long been recognised as a significant risk factor for breast cancer. Quantifying that risk has been approached in publications and practically in a number of different ways. Increasingly regional genetics departments are called upon to help clarify guidelines for referral of women with a family history of breast cancer for genetic testing and to clarify breast cancer risk for women seeking early mammographic screening. This paper represents the current consensus guidelines from the UK Cancer Family Study Group and discusses some of the difficulties surrounding genetic risk estimation. PMID- 10699058 TI - MEMORANDUM FOR: science writers and editors on the journal press list: adding radiotherapy to chemotherapy does not improve disease-free or overall survival of rectal cancer patients PMID- 10699059 TI - MEMORANDUM FOR: science writers and editors on the journal press list: limited potential for human papillomavirus (HPV) testing for women with low-grade pap smears PMID- 10699060 TI - Defining the optimal therapy for rectal cancer. PMID- 10699061 TI - Emerging technologies and cervical cancer. PMID- 10699062 TI - Can HER2 status predict response to cancer therapy? PMID- 10699063 TI - Britain debates validity of guideline development. PMID- 10699064 TI - Stat bite: Black/white differences in ALL incidence in U.S. children. PMID- 10699065 TI - Clinical trials: finding balance in randomization. PMID- 10699066 TI - Cancer treatment during pregnancy: there's strength in numbers for researchers. PMID- 10699067 TI - NIH launches PubMed Central. PMID- 10699068 TI - Preclinical and clinical development of cyclin-dependent kinase modulators. AB - In the last decade, the discovery and cloning of the cyclin-dependent kinases (cdks), key regulators of cell cycle progression, have led to the identification of novel modulators of cdk activity. Initial experimental results demonstrated that these cdk modulators are able to block cell cycle progression, induce apoptotic cell death, promote differentiation, inhibit angiogenesis, and modulate transcription. Alteration of cdk activity may occur indirectly by affecting upstream pathways that regulate cdk activity or directly by targeting the cdk holoenzyme. Two direct cdk modulators, flavopiridol and UCN-01, are showing promising results in early clinical trials, in which the drugs reach plasma concentrations that can alter cdk activity in vitro. Although modulation of cdk activity is a well-grounded concept and new cdk modulators are being assessed for clinical testing, important scientific questions remain to be addressed. These questions include whether one or more cdks should be inhibited, how cdk inhibitors should be combined with other chemotherapy agents, and which cdk substrates should be used to assess the biologic effects of these drugs in patients. Thus, modulation of cdk activity is an attractive target for cancer chemotherapy, and several agents that modulate cdk activity are in or are approaching entry into clinical trials. PMID- 10699069 TI - Randomized trial of postoperative adjuvant chemotherapy with or without radiotherapy for carcinoma of the rectum: National Surgical Adjuvant Breast and Bowel Project Protocol R-02. AB - BACKGROUND: The conviction that postoperative radiotherapy and chemotherapy represent an acceptable standard of care for patients with Dukes' B (stage II) and Dukes' C (stage III) carcinoma of the rectum evolved in the absence of data from clinical trials designed to determine whether the addition of radiotherapy results in improved disease-free survival and overall survival. This study was carried out to address this issue. An additional aim was to determine whether leucovorin (LV)-modulated 5-fluorouracil (5-FU) is superior to the combination of 5-FU, semustine, and vincristine (MOF) in men. PATIENTS AND METHODS: Eligible patients (n = 694) with Dukes' B or C carcinoma of the rectum were enrolled in National Surgical Adjuvant Breast and Bowel Project (NSABP) Protocol R-02 from September 1987 through December 1992 and were followed. They were randomly assigned to receive either postoperative adjuvant chemotherapy alone (n = 348) or chemotherapy with postoperative radiotherapy (n = 346). All female patients (n = 287) received 5-FU plus LV chemotherapy; male patients received either MOF (n = 207) or 5-FU plus LV (n = 200). Primary analyses were carried out by use of a stratified log-rank statistic; P values are two-sided. RESULTS: The average time on study for surviving patients is 93 months as of September 30, 1998. Postoperative radiotherapy resulted in no beneficial effect on disease-free survival (P =.90) or overall survival (P =.89), regardless of which chemotherapy was utilized, although it reduced the cumulative incidence of locoregional relapse from 13% to 8% at 5-year follow-up (P =.02). Male patients who received 5 FU plus LV demonstrated a statistically significant benefit in disease-free survival at 5 years compared with those who received MOF (55% versus 47%; P =.009) but not in 5-year overall survival (65% versus 62%; P =.17). CONCLUSIONS: The addition of postoperative radiation therapy to chemotherapy in Dukes' B and C rectal cancer did not alter the subsequent incidence of distant disease, although there was a reduction in locoregional relapse when compared with chemotherapy alone. PMID- 10699070 TI - Viral vector delivery in solid-state vehicles: gene expression in a murine prostate cancer model. AB - BACKGROUND: Although there are increasingly more clinical trials involving gene therapy, efficient gene transfer remains a major hurdle to success. To enhance the efficiency of delivery of viral vectors in gene therapy protocols, we evaluated the effect of various matrices to act as a vehicle for recombinant virus during intratumoral injection. METHODS: The ability of several vehicles (catgut spacer, polyglycolic acid, chromic catgut, and gelatin sponge matrix) to deliver the canarypox virus ALVAC to the cells of the murine prostate cancer cell line RM-1 was studied in vitro and in vivo. ALVAC recombinants encoding the murine cytokines interleukin 2 (IL-2), interleukin 12 (IL-12), and tumor necrosis factor-alpha (TNF-alpha) were used to assess enhancement of antitumor activity after intratumoral inoculation. Confirmatory experiments were conducted by use of another mouse prostate cancer cell line, RM-11, and a mouse bladder cancer cell line, MB-49. All statistical tests were two-sided. RESULTS: The gelatin sponge matrix proved to be the most effective solid-state vehicle for delivering viral vectors to cells in culture. In addition, this matrix statistically significantly enhanced expression of ALVAC-delivered reporter genes in tumor models when compared with fluid-phase delivery of virus (P =.037 for the RM-1 model and P =.03 for the MB-49 model). Statistically significant growth inhibition of established tumors was observed when a combination of the three recombinant ALVAC viruses expressing IL-2, IL-12, and TNF-alpha was delivered with the matrix in comparison with 1) fluid-phase intratumoral injection of the ALVAC recombinants, 2) no treatment, or 3) treatment with parental ALVAC (all P<.05). CONCLUSIONS: Viral vector delivery in a solid-state vehicle resulted in improved recombinant gene expression in vivo and translated to greater inhibition of tumor growth in an immunotherapy protocol for heterotopic tumor nodules. The efficient delivery of reporter genes described herein may prove useful in many solid tumor gene therapy protocols. PMID- 10699071 TI - Population-based, case-control study of HER2 genetic polymorphism and breast cancer risk. AB - BACKGROUND: Alterations of the HER2 (also known as erbB-2 or neu) proto-oncogene have been implicated in the carcinogenesis and prognosis of breast cancer. A polymorphism at codon 655 (GTC/valine to ATC /isoleucine [Val(655)Ile]) in the transmembrane domain-coding region of this gene has been identified and may be associated with the risk of breast cancer. We evaluated this hypothesis in a subgroup of women who participated in a large-scale, population-based, case control study of breast cancer in Shanghai, China. METHODS: Genomic DNA from 339 patients with breast cancer and 361 healthy control subjects was examined for the Val(655)Ile polymorphism with a polymerase chain reaction-restriction fragment length polymorphism-based assay. All study subjects completed a structured questionnaire during an in-person interview. All P values are from two-sided tests. RESULTS: We found that 25.1% of the case patients and 21.7% of the control subjects were heterozygous for the Val allele and 3.2% of the case patients and 0. 3% of the control subjects were homozygous for this allele (P =.005). Compared with women with the Ile/Ile genotype, women who had the Ile/Val or Val/Val genotype had an elevated risk of breast cancer (odds ratio [OR] = 1.4; 95% confidence interval [CI] = 1.0-2.0; P =. 05) after adjustment for age, educational level, study period, history of breast fibroadenoma, leisure physical activity, and age at first live birth. The risk was elevated even more among women who were homozygous for the Val allele (OR = 14.1; 95% CI = 1.8-113.4). The association was more pronounced among younger women (45 years). The adjusted OR associated with the Val allele was 1.7 (95% CI = 1.1-2.6) for younger women and 1.0 (95% CI = 0.5-1.9) for older women. CONCLUSIONS: Results of this study suggest that polymorphisms of the HER2 gene may be important susceptibility biomarkers for breast cancer risk, particularly among younger women. PMID- 10699072 TI - A ligand of peroxisome proliferator-activated receptor gamma, retinoids, and prevention of preneoplastic mammary lesions. AB - BACKGROUND: Chemoprevention of breast cancer is an active area of investigation. Recent in vivo and in vitro studies have shown that thiazolidinediones (e.g., troglitazone) and retinoids are able to inhibit the growth of breast cancer cells. Troglitazone mediates its action via peroxisome proliferator-activated receptor gamma (PPARgamma). We evaluated the ability of troglitazone, alone or in combination with retinoids, to prevent the induction of preneoplastic lesions by 7,12-dimethylbenz[a]anthracene (DMBA) in a mouse mammary gland organ culture model. METHODS: Mammary glands of BALB/c mice were treated with DMBA (2 microg/mL) to induce preneoplastic lesions in organ culture. Effects of troglitazone, all-trans-retinoic acid (retinoic acid; ligand for retinoic acid receptor [RAR] alpha), and LG10068 (ligand for retinoid X receptors [RXRs]), singly or in combination, on the development of lesions were evaluated. Expression of retinoid receptors (RARalpha and RXRalpha) and PPARgamma was determined by western blot analysis. Statistical significance was determined by generalized chi-squared analysis using the GENCAT software program and Bonferroni correction. All P values are two-sided. RESULTS: Troglitazone (at 10(-5) M) or retinoic acid (at 10(-6) M) markedly inhibited the development of mammary lesions (both P values <.05); however, together they did not enhance the effectiveness of the other. In contrast, LG10068 (at 10(-7) M or 10(-8) M) alone had very little ability to inhibit development of these lesions, but a combination of LG10068 (at 10(-8) M) and troglitazone (at 10(-5) M or 10(-6) M) almost completely inhibited (by 85% and 100%, respectively; both P values <. 05) the development of mammary lesions. The expression of PPARgamma and RXRalpha remained unchanged with the various treatments, whereas the expression of RARalpha was substantially reduced after treatment with the combination of retinoic acid and troglitazone. CONCLUSIONS: To our knowledge, this is the first report showing the possibility of a PPARgamma ligand having chemopreventive activity. Furthermore, an RXR selective retinoid, LG10068, appears to enhance this activity. PMID- 10699073 TI - Joint effect of diet and environmental tobacco smoke on risk of lung cancer among nonsmokers. PMID- 10699074 TI - Ca2+ store dynamics determines the pattern of activation of the store-operated Ca2+ current I(CRAC) in response to InsP3 in rat basophilic leukaemia cells. AB - 1. The relationship between the amplitude of the store-operated Ca2+ ICR AC and intracellular inositol 1,4,5-triphosphate (InsP3) concentration is complex. In rat basophilic leukaemia (RBL-1) cells dialysed with high intracellular Ca2+ buffer, the relationship is supra-linear with a Hill coefficient of 12 and resembles an apparent 'all-or-none' phenomenon. The non-linearity seems to arise from InsP3 metabolism. However, it is not clear which InsP3-metabolising pathway engenders the non-linear behaviour nor whether ICRAC is always activated to its maximal extent by InsP3. 2. Using the whole-cell patch clamp technique, we dialysed RBL-1 cells with different concentrations of the InsP3 analogue InsP3-F. InsP3-F is broken down by Ins(1,4,5)P3 5-phosphatase but is not a substrate for Ins(1,4,5)P3 3-kinase. The relationship between InsP3-F and ICRAC amplitude was supra-linear and very similar to that with InsP3 but was distinct from the graded relationship seen with the non-metabolisable analogue Ins2,4,5P3. 3. In the presence of high intracellular Ca2+ buffer, InsP3-F activated ICRAC to its maximal extent. With moderate Ca2+ buffer, however, sub-maximal ICRAC could be obtained to a maximal InsP3-F concentration. Nevertheless, the relationship between the amplitude of ICRAC and InsP3-F concentration was still supra-linear. 4. Submaximal ICRAC in response to InsP3-F in the presence of moderate Ca2+ buffer was due to partial depletion of the stores, because the size of the current could be increased by thapsigargin. 5. The data suggest that first Ins(1,4,5)P3 5-phosphatase is an important factor which contributes to the non linear relationship between InsP3 concentration and the amplitude of ICRAC and second, InsP3 does not always activate ICRAC to its maximal extent. At moderate buffer strengths, submaximal ICRAC is evoked by maximal InsP3. However, the supra linear relationship between InsP3 concentration and amplitude of the current still holds. PMID- 10699075 TI - Tyrosine phosphorylation modulates the osmosensitivity of volume-dependent taurine efflux from glial cells in the rat supraoptic nucleus. AB - 1. In the supraoptic nucleus, taurine, selectively released in an osmodependent manner by glial cells through volume-sensitive anion channels, is likely to inhibit neuronal activity as part of the osmoregulation of vasopressin release. We investigated the involvement of various kinases in the activation of taurine efflux by measuring [3H]taurine release from rat acutely isolated supraoptic nuclei. 2. The protein tyrosine kinase inhibitors genistein and tyrphostin B44 specifically reduced, but did not suppress, both the basal release of taurine and that evoked by a hypotonic stimulus. Inhibition of tyrosine phosphatase by orthovanadate had the opposite effect. 3. The tyrosine kinase and phosphatase inhibitors shifted the relationship between taurine release and medium osmolarity in opposite directions, suggesting that tyrosine phosphorylation modulates the osmosensitivity of taurine release, but is not necessary for its activation. 4. Genistein also increased the amplitude of the decay of the release observed during prolonged hypotonic stimulation. Potentiation of taurine release by tyrosine kinases could serve to maintain a high level of taurine release in spite of cell volume regulation. 5. Taurine release was unaffected by inhibitors and/or activators of PKA, PKC, MEK and Rho kinase. 6. Our results demonstrate a unique regulation by protein tyrosine kinase of the osmosensitivity of taurine efflux in supraoptic astrocytes. This points to the presence of specific volume-dependent anion channels in these cells, or to a specific activation mechanism or regulatory properties. This may relate to the particular role of the osmodependent release of taurine in this structure in the osmoregulation of neuronal activity. PMID- 10699076 TI - An intracellular ATP-activated, calcium-permeable conductance on the basolateral membrane of single renal proximal tubule cells isolated from Rana temporaria. AB - 1. The following study describes the properties of a non-selective cation channel, which has a unit conductance below the resolving power of the single channel technique, located on the basolateral membrane of single proximal tubule cells isolated from frog kidney. The conductance was examined using cell attached, inside-out and outside-out patches. Due to the small single channel magnitude, macroscopic patch currents were measured. 2. Addition of 2 mM ATP to the intracellular surface of excised patches activated an outwardly rectifying conductance (MCANS): outward (Gout) and inward (Gin) conductances increased by 46.8 +/- 6.7 and 11.6 +/- 2.1 pS, respectively (n = 29). MCANS was more selective for cations than anions, with a cation:anion selectivity ratio of 10.1 +/- 1.7 (n = 7), but did not discriminate between Na+ and K+. It was more selective for Ca2+ over Na+, with a Ca2+:Na+ selectivity ratio of 4. 49 +/- 0.69 (n = 7). 3. In cell attached patches addition of 100 microM strophanthidin to the bath increased both Gout and Gin. However this increase in conductance was absent in the presence of Gd3+, which inhibits MCANS. 4. These data suggest that single proximal tubule cells isolated from frog kidney contain an ATP-activated, non-selective cation conductance. The conductance does not discriminate between Na+ and K+, but is more selective for Ca2+ over Na+. Considering the prevailing electrochemical gradients for these ions, functional activation of the conductance would be expected to lead to a rise in intracellular Ca2+. MCANS is linked to the activity of the Na+, K+-ATPase and may therefore provide a link between the ATPase and K+ channel activity in the basolateral membrane and form an integral part of the pump-leak mechanism in transporting epithelia. PMID- 10699077 TI - The chloride channel ClC-2 contributes to the inwardly rectifying Cl- conductance in cultured porcine choroid plexus epithelial cells. AB - 1. The contribution of ClC-2 protein to the inwardly rectifying Cl- conductance in cultured porcine choroid plexus epithelial cells was investigated using Western analysis and whole-cell current recordings. 2. Inwardly rectifying currents were elicited by hyperpolarizing voltage at a potential more negative than -50 mV in the presence of intracellular protein kinase A (PKA). The relative halide selectivity estimated from the shift in the reversal potential (Erev) was I- > Br- > Cl- > F-. 3. Extracellular vasoactive intestinal peptide (VIP) activated the same currents in a dose-dependent manner with a half-maximal concentration of 167.3 nM. H-89 (a PKA inhibitor) interfered with the current activation by VIP. 4. The Cl- channel was inhibited by external Cd2+, Ba2+or H+, but only weakly inhibited by known Cl- channel blockers including glibenclamide, NPPB, DIDS and anthracene-9-carboxylic acid (9AC). 5. A specific antibody to ClC 2 detected a 79 kDa protein in porcine choroid plexus cells, which was reduced in cells treated with antisense oligodeoxynucleotide for ClC-2. Both PKA and VIP failed to activate the inwardly rectifying Cl- currents in cells transfected with the antisense oligodeoxynucleotide, while they activated the currents in cells transfected with GFP alone or the control oligodeoxynucleotide randomized from antisense oligonucleotide. 6. It is concluded that ClC-2 protein contributes to the inwardly rectifying Cl- conductance in porcine choroid plexus epithelial cells. PMID- 10699078 TI - Ca2+ and cAMP-activated Cl- conductances mediate Cl- secretion in a mouse renal inner medullary collecting duct cell line. AB - 1. The nature of Cl- conductance(s) participating in transepithelial anion secretion by renal inner medullary collecting duct (IMCD, mIMCD-K2 cell line) was investigated. 2. Extracellular ATP (100 microM) stimulated a transient increase in both whole-cell Cl- conductance and intracellular free Ca2+. In contrast, ionomycin (10-100 nM) caused a sustained increase in whole-cell Cl- conductance. Pre-loading cells with the Ca2+ buffer BAPTA abolished the ATP-dependent responses and delayed the onset of the increase observed with ionomycin. 3. The Ca2+-activated whole-cell Cl- current stimulated by ATP (peak) and ionomycin (maximal) displayed (i) a linear steady-state current-voltage relationship and (ii) time and voltage dependence with slow activation at +80 mV and slow inactivation at -80 mV. In BAPTA-loaded cells, ionomycin-elicited whole-cell currents exhibited pronounced outward rectification with time-dependent activation/inactivation. 4. Ca2+-activated and forskolin-activated Cl- conductances co-exist since ATP activation of whole-cell current occurred during a maximal stimulation by forskolin in single cell recordings. 5. In IMCD epithelial layers, ATP and ionomycin stimulated an inward short circuit current (Isc) dependent upon basal medium Na+ and Cl-/HCO3- but independent of the presence of apical bathing medium Na+ and Cl-/HCO3-. This was identical to forskolin stimulation and consistent with transepithelial anion secretion. 6. PCR amplification of reverse-transcribed mRNA using gene-specific primers demonstrated expression of both cystic fibrosis transmembrane conductance regulator (CFTR) mRNA and Ca2+-activated Cl- channel (mCLCA1) mRNA in mIMCD-K2 cells. 7. Ca2+ and forskolin-activated Cl- conductances participate in anion secretion by IMCD. PMID- 10699079 TI - Stoichiometry of Na+-Ca2+ exchange in inside-out patches excised from guinea-pig ventricular myocytes. AB - 1. The stoichiometry (nx) of cardiac Na+-Ca2+ exchange was examined by measuring the reversal potential of the Na+-Ca2+ exchange current (INa-Ca) in large inside out patches, 'macro patches', excised from intact guinea-pig ventricular cells. 2. Cytoplasmic application of Na+ (Na+i) or Ca2+ (Ca2+i) induced INa-Ca which showed properties similar to INa-Ca in the giant membrane patch. The outward INa Ca was depressed by an exchanger inhibitory peptide, XIP. 3. The reversal potential of the XIP-sensitive current indicated that nx was approximately 4 (3.6 4.2) at 9-40 mM Na+i, and nx tended to increase as Na+i was increased. Proteolysis by trypsin did not significantly affect the stoichiometry. Similar results were obtained from the reversal potential of INa-Ca that was induced by application of both Na+i and Ca2+i. 4. At 0.1 microM Ca2+i, nx was approximately 4 (3.7-4. 4) at 6-25 mM Na+i and tended to increase as Na+i was increased. When Ca2+i was changed from 0.1 to 1 and 1000 microM at constant 50 mM Na+i, the value was approximately 4 (3.6-4.4). 5. When the extracellular Na+ (Na+o) and Ca2+ (Ca2+o) concentrations were varied in the presence of 25 or 9 mM Na+i and 1 microM Ca2+i, nx was almost constant ( approximately 4) over the range 0.3-20 mM Ca2+o and 10-145 mM Na+o. 6. These results indicated that the stoichiometry of Na+-Ca2+ exchange is different from generally accepted 3Na+:1Ca2+, and suggested that the stoichiometry is either 4Na+:1Ca2+ or variable depending on Na+i and Ca2+i. PMID- 10699080 TI - Paradoxical block of the Na+-Ca2+ exchanger by extracellular protons in guinea pig ventricular myocytes. AB - 1. The Na+-Ca2+ exchange is a major pathway for removal of cytosolic Ca2+ in cardiac myocytes. It is known to be inhibited by changes of intracellular pH that may occur, for example, during ischaemia. In the present study, we examined whether extracellular protons (pHo) can also affect the cardiac exchange. 2. Na+ Ca2+ exchange currents (INa-Ca) were recorded from single adult guinea-pig ventricular myocytes in the whole-cell voltage-clamp configuration while [Ca2+]i was simultaneously imaged with fluo-3 and a laser-scanning confocal microscope. To activate INa-Ca, intracellular Ca2+ concentration jumps were generated by laser flash photolysis of caged Ca2+ (DM-nitrophen). 3. Exposure of the cell to moderately and extremely acidic conditions (pHo 6 and 4) was accompanied by a decrease of the peak INa-Ca to 70 % and less than 10 %, respectively. The peak INa-Ca was also inhibited to about 45 % of its initial value by increasing pHo to 10. The largest INa-Ca was found at pHo approximately 7.6. 4. Simultaneous measurements of [Ca2+]i and INa-Ca during partial proton block of the Na+-Ca2+ exchanger revealed that the exchange current was more inhibited by acidic pHo than the rate of Ca2+ transport. This observation is consistent with a change in the electrogenicity of the Na+-Ca2+ exchange cycle after protonation of the transporter. 5. We conclude that both extracellular alkalinization and acidification affect the Na+-Ca2+ exchanger during changes of pHo that may be present under pathophysiological conditions. During both extreme acidification or alkalinization the Na+-Ca2+ exchanger is strongly inhibited, suggesting that extracellular protons may interact with the Na+-Ca2+ exchanger at multiple sites. In addition, the electrogenicity and stoichiometry of the Na+-Ca2+ exchange may be modified by extracellular protons. PMID- 10699081 TI - Inhibition of sarcoplasmic reticulum function by polyunsaturated fatty acids in intact, isolated myocytes from rat ventricular muscle. AB - 1. We have studied the effects of two polyunsaturated fatty acids (PUFAs), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) on spontaneous and electrically stimulated contractions in single, isolated ventricular myocytes from rat hearts. 2. The frequency of spontaneous waves of calcium release and contraction (induced by elevation of the bathing calcium concentration) is reduced in the presence of EPA. At the same time the resting level of intracellular calcium falls, the resting cell length increases and the amplitude of shortening decreases. All these effects are reversed on removal of EPA. 3. Imaging of the waves of calcium release shows that the amplitude and the rate of propagation of the wave is increased in EPA. Consistent with the increased amplitude, integration of the caffeine-induced Na+-Ca2+ exchange current (a measure of the sarcoplasmic reticulum (SR) calcium content) is increased by both EPA and DHA. 4. EPA has a maintained negative inotropic effect on voltage clamped myocytes. This seems to be entirely due to inhibition of the L-type calcium current. Smaller depolarising pulses in control conditions that elicit the same calcium current as in EPA also activate the same level of contraction. This is in spite of the increased SR calcium content in EPA. 5. It is concluded that PUFAs have two effects on the SR; they reduce the availability of calcium for uptake and they inhibit the release mechanism. Both of these effects should lower the frequency of spontaneous waves of calcium release. As spontaneous release of calcium can initiate arrhythmias, some of the anti-arrhythmic action of PUFAs must be exerted at the level of the SR. PMID- 10699082 TI - Functional expression and regulation of the hyperpolarization activated non selective cation current in embryonic stem cell-derived cardiomyocytes. AB - 1. The biophysical and pharmacological characteristics of the hyperpolarization activated non- selective cation current (If) were recorded using whole-cell voltage clamp in embryonic stem (ES) cell-derived cardiomyocytes at different stages of development. 2. The cation current was detected in a large percentage (65 %) of early stage (EDS, differentiated for 7 + 3-4 days) cells at a current density of 11.4 +/- 0.6 pA pF-1 (n = 47). In late stage (LDS, differentiated for 7 + 9-12 days) cells the percentage of cells expressing If decreased (45 %), but If densities (15.5 +/- 0.9 pA pF-1, n = 20) were increased. 3. The muscarinic agonist carbachol (CCh, 1 microM) depressed basal If in EDS cells by 45.7 +/- 6.5 %, n = 5) and was without effect in LDS cardiomyocytes (n = 4). The beta adrenoceptor agonist isoprenaline (ISO, 1 microM) stimulated If in LDS cells by 33 +/- 5.2 % (n = 6) but not in EDS cells (n = 5). 4. Cell infusion with the catalytic subunit of the cAMP-dependent protein kinase (PKA, 7 microM) stimulated If in EDS cells by 37.0 +/- 2.9 %, (n = 4), but subsequent superfusion of 8-bromo cAMP (200 microM) was without effect. Intracellular perfusion of LDS cardiomyocytes with the highly selective peptide inhibitor of PKA (PKI, 20 microM) completely inhibited the stimulation of the L-type Ca2+ current (ICa,L) as well as of If by ISO (1 microM). 5. Extracellular superfusion with phosphodiesterase (PDE) inhibitors - IBMX, a non-selective antagonist, Erythro-9 (2-hydoxy-3-nonyl)adenine (EHNA), a PDE2 antagonist and rolipram, a PDE4 antagonist - resulted in stimulation of ICa,L and If in EDS cells. By contrast, milrinone and cilostamide, two PDE3 antagonists, stimulated ICa,L, but not If. 6. The present work demonstrates that If is functionally expressed during early cardiomyogenesis. Similar to ICa,L, If is regulated during embryonic development by phosphorylation via PKA. In contrast to ICa,L, If is not regulated by PDE3 suggesting different localization of these ion channels with respect to PDE3. PMID- 10699084 TI - P2X purinoceptor-induced sensitization of ferret vagal mechanoreceptors in oesophageal inflammation. AB - 1. Using an in vitro single unit recording technique we studied the changes in mechanical and chemical sensitivity of vagal afferent fibres in acute oesophagitis, with particular attention to inflammatory products such as purines. 2. Histologically verified oesophagitis was induced by oesophageal perfusion of 1 mg ml-1 pepsin in 150 mM HCl in anaesthetized ferrets for 30 min on two consecutive days. Controls were infused with 154 mM NaCl. 3. The number of action potentials evoked in oesophageal mucosal afferents by mucosal stroking with calibrated von Frey hairs (10-1000 mg) was stimulus dependent. In oesophagitis responsiveness was reduced across the range of stimuli compared with controls. 4. Topical application of the P2X purinoceptor agonist alphabeta-methylene ATP had no direct excitatory effect on afferents. In oesophagitis, but not in controls, there was a significant increase in responses to stroking with von Frey hairs during superfusion with alphabeta-methylene ATP (1 microM). 5. Mucosal afferents responded directly to one or more chemical stimuli: 26 % (5/19 afferents) responded in controls, and 47 % (7/15 afferents) in oesophagitis. There were no differences in responsiveness to bradykinin (1 microM), prostaglandin E2 (100 microM), 5-hydroxytryptamine (100 microM), capsaicin (1 mM) or hydrochloric acid (150 mM) between control and oesophagitis groups. 6. We conclude that a sensitizing effect of a P2X purinoceptor agonist on mechanosensory function is induced in oesophagitis. This effect is offset by a decrease in basal mechanosensitivity. PMID- 10699083 TI - Guinea-pig sympathetic neurons express varying proportions of two distinct P2X receptors. AB - 1. Characterization of P2X receptors on neurons of guinea-pig superior cervical ganglion (SCG) has been carried out using a whole-cell voltage-clamp technique. 2. Application of ATP and alpha,beta-methylene ATP (alphabeta-MeATP) produced fast activating inward currents, which desensitized slowly. The maximum response to alphabeta-MeATP was 36 +/- 23 % (range 0.1-100 %) of that evoked by ATP in the same cell. 3. Co-application of alphabeta-MeATP (300 microM) with ATP (300 microM) produced a response that was 97 +/- 1 % of that given by ATP alone. Following desensitization with alphabeta-MeATP, the decrease in response to ATP was equal to the absolute reduction in response to alphabeta-MeATP in the same cell. 4. The concentration-response curve for alphabeta-MeATP had an EC50 of 42 microM and a Hill coefficient of 1.17. For cells where the ratio of alphabeta MeATP/ATP currents at 100 microM was < 0.1, the ATP concentration-response curve had an EC50 of 56 microM and a Hill coefficient of 1.95. However, in cells where the ratio was > 0.7, the curve had an EC50 of 60 microM and a Hill coefficient of 0.97. 5. The response to 100 microM alphabeta-MeATP was inhibited by 2' (or 3')-O trinitrophenyl-ATP (TNP-ATP) with an IC50 of 70 nM. However, on cells where the ratio of alphabeta-MeATP/ATP currents was < 0.1, ATP was inhibited by TNP-ATP with an IC50 of 522 nM. 6. Immunohistochemical staining with antibodies raised against rat P2X2 and P2X3 epitopes suggested that both subunits were expressed by guinea-pig SCG neurons. 7. We conclude that varying proportions of two distinct P2X receptors coexist on the cell bodies of individual guinea-pig SCG neurons, which may correspond to homomeric P2X2 and heteromeric P2X2/3 receptors. PMID- 10699085 TI - Rapid compensatory changes in GABA receptor efficacy in rat vestibular neurones after unilateral labyrinthectomy. AB - 1. The inhibitory effects of the GABAA agonist muscimol and the GABAB agonist baclofen on tonically active medial vestibular nucleus (MVN) neurones were recorded in slices of the rat dorsal brainstem in vitro, to determine whether any changes occurred in the functional efficacy of GABAergic inhibition in these cells during the initial rapid stage of 'vestibular compensation', the behavioural recovery that takes place after unilateral labyrinthectomy (UL). These experiments were carried out in preparations where the midline was cut, severing all commissural connections between the two vestibular nuclei. 2. Slices of the MVN were prepared from normal animals and animals that had been unilaterally labyrinthectomised 4 h earlier. The mean in vitro discharge rate of MVN neurones in the rostral region of the ipsi-lesional nucleus after UL was significantly higher than that in control slices, confirming our earlier reports of an increase in intrinsic excitability of these cells in the early stage of vestibular compensation. The in vitro discharge rates of caudal ipsi-lesional MVN cells, and rostral and caudal contra-lesional MVN cells, were not different from controls. 3. Muscimol and baclofen caused reversible, dose-related inhibition of the tonic discharge rate of MVN cells in control slices. In slices prepared from UL animals, MVN cells in the rostral region of the ipsi-lesional nucleus showed a marked downregulation of their response to both muscimol and baclofen, seen as a rightward shift and a decrease in slope of the dose-response relationships for the two agonists. In the contra-lesional nucleus, there was a small but significant upregulation of the responsiveness of both rostral and caudal MVN cells to baclofen, and a marked upregulation of the responsiveness of caudal MVN cells to muscimol. 4. In slices from animals that had undergone bilateral labyrinthectomy 4 h earlier, the downregulation of the functional efficacy of GABA receptors in the rostral MVN cells did not occur. The changes in GABA receptor efficacy after UL are therefore not due to the vestibular de afferentation itself, but are instead due to the imbalance in excitability of the vestibular nuclei of the lesioned and intact sides, and the enhanced commissural inhibition of the ipsi-lesional MVN cells that follows UL. 5. The downregulation of GABA receptor efficacy in the ipsi-lesional MVN neurones is functionally compensatory, in that their response to commissural and cerebellar inhibitory drive will be significantly reduced after UL. Their intrinsic membrane conductances, and their remaining excitatory synaptic inputs, will consequently be more effective in causing depolarisation and the restoration of resting activity. Simultaneously the upregulation of GABAergic efficacy in the contra lesional MVN will tend to reduce the hyperactivity on the contralateral side. These adaptive changes therefore represent a plausible cellular mechanism for the recovery of resting discharge in the ipsi-lesional MVN neurones, and the 're balancing' of the excitability of the vestibular neurones of the lesioned and intact sides, as occurs after UL in vivo. 6. We propose that the adaptive regulation of the functional efficacy of GABA receptors in the MVN neurones may be an important cellular mechanism for the 'homeostasis of bilateral excitability' of the vestibular nuclei of the two sides. PMID- 10699086 TI - Paired-pulse modulation at individual GABAergic synapses in rat hippocampus. AB - 1. Unitary inhibitory postsynaptic currents (uIPSCs) were recorded in synaptically coupled pairs of CA1 hippocampal interneurons and pyramidal neurons in rat brain slices by using dual patch-clamp techniques. Paired-pulse modulation of uIPSCs at individual GABAergic synapses was tested. 2. GABAergic synapses could be divided into two subgroups, high and low failure, depending on their failure rate. 3. The external Ca2+ levels modulate the failure rate of uIPSCs. In 0.51 mM Ca2+, low-failure pairs had a high-failure characteristic, whereas high failure pairs had a low-failure characteristic in 8 mM Ca2+. The results suggest that uIPSC failures result from the Ca2+-dependent release mechanism rather than axon propagation failures. 4. Paired-pulse facilitation (PPF) occurred in high failure pairs when the interspike interval was 20 ms. Paired-pulse depression (PPD) was not predominant in high-failure pairs. 5. Potency of uIPSCs, the average amplitude of non-failure events, was enhanced by PPF, suggesting that multiple synapses connect each pair. Differing numbers of activated synapses contributed to the variable amplitude of uIPSCs from a given pair. 6. PPD occurred in low-failure pairs at the tested range of interspike intervals (20-200 ms). The uIPSC2 after a large uIPSC1 was smaller than the uIPSC2 after a small uIPSC1, suggesting that PPD is use dependent and due to a decrease in the quantal content (m) after the first release. 7. In 8 mM Ca2+, PPD occurred in high failure pairs and was larger in low-failure pairs, suggesting that the occurrence of PPF or PPD depends on the baseline release probability. 8. The GABAB receptor antagonist CGP 55845A (5 microM) decreased the baseline release probability of inhibitory synapses and attenuated PPD indirectly, rather than by blocking presynaptic GABAB autoreceptors. PMID- 10699088 TI - Coherent rhythmic discharges in sympathetic nerves supplying thermoregulatory circulations in the rat. AB - 1. In anaesthetised rats, activity recorded from sympathetic postganglionic neurones innervating the tail circulation has characteristic rhythmicity (0.4-1.2 Hz). At the population level this rhythmicity can be seen as a peak (T-peak) in autospectra of sympathetic activity recorded from ventral collector nerves (VCNs). 2. Here we investigated whether nerves supplying thermoregulatory circulations share common rhythmic discharges at T-peak frequency. Activity was recorded from nerve pairs consisting of left ventral collector nerve (LVCN) and one of the following: right ventral collector nerve (RVCN), left dorsal collector nerve (DCN), left saphenous nerve (SN) or left renal nerve (RN). 3. During central apnoea, T-peak frequencies in RVCN autospectra were similar to those of simultaneously recorded LVCN and these activities were coherent. Similar observations were made for nerve pairs involving LVCN-DCN and LVCN-SN. In contrast, autospectra of RN activity did not contain T-peaks. 4. In comparison to the peaks in autospectra of RN activity, when the frequency of rhythmic phrenic nerve activity was manipulated T-peaks in VCN, DCN and SN autospectra did not show obligatory 1:1 locking. 5. We conclude that T-peaks are a robust feature of autospectra of sympathetic discharges supplying thermoregulatory circulation but not those influencing the kidney. The high coherence demonstrated between the T peak discharges is consistent with the view that common/coupled oscillators located within the CNS influence cutaneous vasoconstrictor sympathetic activity. PMID- 10699087 TI - ATP as a mediator of mammalian central CO2 chemoreception. AB - 1. A role for P2 purinoceptors in the chemosensory response of respiratory neurones localised in the ventrolateral medulla to changes in arterial CO2 levels was investigated in the anaesthetised rat. Extracellular recordings were made from different classes of respiratory neurone and the effects of P2 receptor blockade on CO2-evoked changes in activity investigated. 2. Increasing inspired CO2 excited 85 % of inspiratory neurones in the pre-Botzinger complex. In all cases, CO2-evoked excitation was blocked by ionophoretic application of the P2 receptor antagonists suramin (0.02 M) and pyridoxal-phosphate-6-azophenyl-2',4' disulphonic acid (PPADS; 100 microM), but not the adenosine receptor antagonist 8 phenyltheophylline (8-PT; 100 microM). Suramin and PPADS often reduced ongoing activity, and blocked the excitatory effects of ATP. Inspiratory neurones were also excited by the P2X receptor agonist alphabeta-methyleneATP, suggesting a specific role for P2X receptors. 3. Sixty-six per cent of pre-inspiratory neurones were also excited by CO2. This effect was reduced or abolished by prior application of P2 receptor antagonists. Although post-inspiratory and expiratory neurones were excited by increasing levels of CO2, and also by ionophoretically applied ATP, the CO2-evoked effects were unaffected by P2 receptor blockade. 4. We suggest that ATP, possibly acting via P2X purinoceptors localised within the ventral respiratory group, is involved in central chemoreception. Specifically, these distinctive CO2-P2X-mediated actions were observed only in inspiratory neurones (incrementing inspiratory neurones and pre-inspiratory neurones), which appear to have purinoceptors with pH sensitivity that can account for the actions of CO2 in modifying ventilatory activity. PMID- 10699089 TI - Fast (3 Hz and 10 Hz) and slow (respiratory) rhythms in cervical sympathetic nerve and unit discharges of the cat. AB - 1. In seven decerebrate cats, recordings were taken from the preganglionic cervical sympathetic (CSy) nerves and from 74 individual CSy fibres. Correlation and spectral analyses showed that nerve and fibre discharges had several types of rhythm that were coherent (correlated) between population and unit activity: respiratory, '3 Hz' (2-6 Hz, usually cardiac related), and '10 Hz' (7-13 Hz). 2. Almost all units (73/74) had respiratory modulation of their discharge, either phasic (firing during only one phase) or tonic (firing during both the inspiratory (I) and expiratory (E) phases). The most common pattern consisted of tonic I-modulated firing. When the vagi were intact, lung afferent input during I greatly reduced CSy unit and nerve discharge, as evaluated by the no-inflation test. 3. The incidence of unit-nerve coherent fast rhythms (3 Hz or 10 Hz ranges) depended on unit discharge pattern: they were present in an appreciable fraction (30/58 or 52 %) of tonic units, but in only a small fraction (2/15 or 13 %) of phasic units. 4. When baroreceptor innervation (aortic depressor amd carotid sinus nerves) was intact, rhythms correlated to the cardiac cycle frequency were found in 20/34 (59 %) of units. The cardiac origin of these rhythms was confirmed by residual autospectral and partial coherence analysis and by their absence after baroreceptor denervation. 4. The 10 Hz coherent rhythm was found in 7/34 units when baroreceptor innervation was intact, where it co-existed with the cardiac-locked rhythm; after barodenervation it was found in 9/50 neurones. Where both rhythms were present, the 10 Hz component was sometimes synchronized in a 3:1 ratio to the 3 Hz (cardiac-related) frequency component. 5. The tonic and phasic CSy units seem to form distinct populations, as indicated by the differential responses to cardiac-related afferent inputs when baroreceptor innervation is intact. The high incidence of cardiac-related correlation found among tonic units suggests that they are involved in vasomotor regulation. The high incidence of respiratory modulation of discharge suggests that the CSy units may be involved in regulation of the nasal vasculature and consequent ventilation related control of nasal airway resistance. PMID- 10699090 TI - Habituation and desensitization of the Hering-Breuer reflex in rat. AB - 1. Many processes in mammalian and invertebrate central nervous systems exhibit habituation and/or sensitization of their responses to repetitive stimuli. Here, we studied the adaptive behaviours of the respiratory pattern generator in rat on repetitive vagal-afferent stimulation and compared these behaviours obtained in vivo with the reported effects of such stimuli on synaptic transmission in the corresponding signal pathway in vitro. 2. Sustained (1 min) electrical pulsed stimulation of the vagus nerve elicited the classic Hering-Breuer (HB) reflex slowing of the respiratory rhythm followed by a bi-exponential recovery, and a post-stimulus rebound (PR). The recovery from the HB reflex satisfied the classic criteria of habituation. 3. The fast component of the recovery and the PR were abolished by systemic administration of an NMDA receptor antagonist or electrolytic lesioning of the pontine Kolliker-Fuse nucleus. The characteristics of the fast recovery and PR suggest a vagally induced desensitization of the NMDA receptor-dependent pontine input to the respiratory pattern generator. 4. The slow component of recovery persist after both experimental interventions and accounted for the habituation to the vagal input. The characteristics of the slow recovery in vivo were reminiscent of the reported synaptic accommodation in vitro in the medullary region where vagal afferents terminate. 5. The habituation of vagal input and desensitization of pontine input act in concert to offset the HB reflex. Such simultaneous habituation-desensitization in parallel neural pathways with differing sensitivities to NMDA receptor activation represent a hitherto unknown pairing of dual non-associative learning processes in the mammalian brain. PMID- 10699091 TI - Lack of a role for cyclic nucleotide gated cation channels in lung liquid absorption in fetal sheep. AB - 1. Late gestation fetal sheep were chronically catheterised in utero to allow measurement of the rate of production of lung liquid (Jv) from 132-143 days gestation (term, 147 days), and to test the hypothesis that cyclic nucleotide gated cation channels mediate a component of fetal lung liquid absorption. 2. In eight experiments, 0.5 microg min-1 adrenaline caused a significant (P < 0.005) reduction in Jv from +18. 12 +/- 3.52 to -10.27 +/- 5.26 ml h-1. Dichlorobenzamil (a blocker of cyclic nucleotide gated cation channels) at 1.5 x 10-5 M did not significantly inhibit the adrenaline-induced lung liquid absorption (Jv dichlorobenzamil, -5.77 +/- 2.78 ml h-1; P > 0.1) when the data were grouped, but did exert a significant gestational effect (r = 0. 90, P < 0.01). Subsequent addition of 10-4 M amiloride (a blocker of epithelial sodium channels) abolished the adrenaline-induced absorption of lung liquid (mean Jv amiloride, +6.45 +/- 1.59 ml h-1; P < 0.01 relative to Jv adrenaline and P < 0.005 relative to Jv dichlorobenzamil). 3. In seven experiments, 0.5 microg min-1 adrenaline caused a significant (P < 0.0005) reduction in Jv from +18.95 +/- 2. 98 to -10.08 +/- 3.75 ml h-1. Amiloride (10-4 M) inhibited the adrenaline response (Jv amiloride, +5.46 +/- 1.09 ml h-1; P < 0.005). However, subsequent addition of 1.5 x 10-5 M dichlorobenzamil had no additive effect to that of amiloride (Jv dichlorobenzamil, +4.58 +/- 0.93 ml h-1; P > 0.1). 4. In six experiments, the cGMP analogue 8-Br-cGMP at 10-4 M caused a significant (P < 0.05) reduction in Jv from +15.20 +/- 2.81 to +11.63 +/- 1.71 ml h-1. Amiloride (10-4 M) did not block the effect of 8-Br-cGMP (Jv amiloride, +14.00 +/- 2.49 ml h-1; not significantly different from 8-Br-cGMP). Subsequent addition of 1.5 x 10-5 M dichlorobenzamil also did not block the effect of 8-Br-cGMP (Jv dichlorobenzamil, +11.37 +/- 1.22 ml h-1; not significantly different from either Jv amiloride or Jv 8-Br-cGMP). 5. We conclude that, in fetal sheep, neither adrenaline nor cGMP stimulate lung liquid absorption by actions on cyclic nucleotide gated cation channels, and that the effect of cGMP on fetal lung liquid secretion is minor and does not involve epithelial sodium channels. The effect of dichlorobenzamil, when given before amiloride, was probably due to an action on amiloride sensitive epithelial sodium channels. PMID- 10699092 TI - Short latency inhibition of human hand motor cortex by somatosensory input from the hand. AB - 1. EMG responses evoked in hand muscles by transcranial stimulation over the motor cortex were conditioned by a single motor threshold electrical stimulus to the median nerve at the wrist in a total of ten healthy subjects and in five patients who had electrodes implanted chronically into the cervical epidural space. 2. The median nerve stimulus suppressed responses evoked by transcranial magnetic stimulation (TMS) in relaxed or active muscle. The minimum interval between the stimuli at which this occurred was 19 ms. A similar effect was seen if electrical stimulation was applied to the digital nerves of the first two fingers. 3. Median or digital nerve stimulation could suppress the responses evoked in active muscle by transcranial electrical stimulation over the motor cortex, but the effect was much less than with magnetic stimulation. 4. During contraction without TMS, both types of conditioning stimuli evoked a cutaneomuscular reflex that began with a short period of inhibition. This started about 5 ms after the inhibition of responses evoked by TMS. 5. Recordings in the patients showed that median nerve stimulation reduced the size and number of descending corticospinal volleys evoked by magnetic stimulation. 6. We conclude that mixed or cutaneous input from the hand can suppress the excitability of the motor cortex at short latency. This suppression may contribute to the initial inhibition of the cutaneomuscular reflex. Reduced spinal excitability in this period could account for the mild inhibition of responses to electrical brain stimulation. PMID- 10699093 TI - There is no simple temporal relationship between the initiation of rapid reactive hand movements and the phase of an enhanced physiological tremor in man. AB - 1. A postural hand tremor of enhanced size was induced in eleven subjects. There was rhythmic activity in the posturally active extensor muscles synchronised to the tremor oscillation, which implied an oscillatory modulation of motor neurones. 2. The subjects executed single rapid wrist flexion movements in response to a flash of light. The light flash was presented at an instant when the wrist was spontaneously moving in the flexion direction, extension direction or at random. The time taken to generate a movement was not significantly different or more consistent in any of the conditions. 3. Inspection of individual and averaged acceleration and EMG records strongly suggests that the movement is made at a time that is independent of the tremor cycle at the time of stimulus presentation or at the moment of movement initiation. 4. These observations suggest that the central or spinal mechanism generating tremor does not gate the central mechanism that produces voluntary movement. PMID- 10699094 TI - The effect of fatigue on multifinger co-ordination in force production tasks in humans. AB - 1. This study investigated the effects of fatigue, induced by production of maximal isometric force for 60 s with four fingers, upon indices of multifinger co-ordination. 2. Measurements of individual finger forces were performed during single- and multifinger maximal force production (maximal voluntary contraction, MVC) for two sites of force application, the middle of the distal or the middle of the proximal phalanxes. Two fatiguing exercises were used, involving force production at the distal phalanxes and at the proximal phalanxes. Fourteen subjects were tested. 3. The total force in four-finger tasks dropped by about 43 % when it was produced at the site involved in the fatiguing exercise. During force production at the other site, MVC dropped by 23 %. During single-finger MVC tests, force drop with fatigue was similar across all four fingers (about -25 % of their corresponding MVCs). 4. Force production by one finger was accompanied by involuntary force production by other fingers (enslaving). Enslaving remained unchanged by fatigue when measured during force generation at the site involved in the fatiguing exercise, but increased during force production at the other site. 5. The total MVC of four fingers acting in parallel was smaller than the sum of the MVCs of these fingers in single-finger tasks (force deficit). The force deficit increased with fatigue. Force-sharing patterns during four-finger tasks showed only minor changes under fatigue. 6. These results indicate that the effects of fatigue were not limited to changes in the force-generating capabilities of the muscles. In particular, fatigue could lead to a reorganisation at a neural level that defines commands to individual fingers. PMID- 10699095 TI - Asthma in life context: Video Intervention/Prevention Assessment (VIA) AB - OBJECTIVE: Video Intervention/Prevention Assessment (VIA) was developed to determine whether medical information gathering might be augmented by video diaries created by patients to show clinicians the realities of managing chronic disease in the contexts of their lives. DESIGN: Children and adolescents who met National Heart, Lung, and Blood Institute criteria for moderate or severe asthma were enrolled from an urban pediatric hospital and an inner-city health center. Comprehensive, asthma-specific medical histories were obtained from study participants in standard clinical interviews. Participants were trained to use video camcorders and recorded visual narratives of how they lived with and managed their asthma over a 4- to 8-week period. These visual narratives were screened by a trained observer, who completed the initial comprehensive medical history based solely on viewing the video. Information from participants' medical history interviews was compared with observations of their visual narratives. RESULTS: Twenty young people 8 to 25 years old completed the VIA Asthma study. Important variations were found between participants' medical history interviews and their visual narratives. All 20 participants reported specific environmental triggers for their asthma; 19 had 1 or more of these triggers documented on video in their daily living environments (video illustrations online, available at: ). Exposures to known triggers ranged from 25% (noxious fumes) to 91% (mold). Exposure to tobacco smoke that was denied in the interview was revealed on video in 63%. The 18 participants who revealed medication use in their visual narratives were assessed for adherence: 33% exceeded prescribed doses, 28% discontinued medications without consulting a clinician, and 72% used ineffective inhaler technique. CONCLUSIONS: VIA visual narratives extended a comprehensive, standard of care medical history, yielding a more complete and accurate understanding of exacerbating environmental exposures and inappropriate medication usage of children and adolescents with asthma. VIA is an effective tool for revealing the physical and psychosocial environments in which young people live with disease. Patient-created video can enrich our understanding of the illness experiences of children and adolescents. VIA has the potential to enhance clinical data gathering, guide the development of more effective and sensitive management strategies, and educate clinicians about the realities of the young person living with illness. PMID- 10699096 TI - Evaluation of a pediatric hospitalist service: impact on length of stay and hospital charges. AB - OBJECTIVES: Inpatient medical services supervised by pediatric hospitalist physicians are a new development in academic medical centers in the United States. In a large pediatric teaching hospital, we compared length of stay, readmission rates, and hospital charges for children admitted to medical services with and without a hospitalist system of care. DESIGN: This retrospective observational study compared a baseline year of a traditional ward service (TS) with a subsequent year of a new hospitalist system of care called the Generalist Inpatient Service (GIS). Data were obtained from the hospital's clinical, demographic, and financial databases and from selected record review. All hospitalizations were at least 24 hours long and did not involve a stay in an intensive care unit. RESULTS: The average length of stay was longer for the 627 TS hospitalizations than for the 813 GIS hospitalizations (2.7 +/- 2.0 vs 2.4 +/- 1.7 days). Total hospital charges were significantly lower on the GIS ($3002 +/- $2160 vs $2720 +/- $1933) because of lower room and respiratory therapy charges. Three readmissions to the TS and 8 to the GIS occurred within 24 hours of hospital discharge and were, therefore, considered potentially preventable by a longer initial hospital stay. CONCLUSIONS: In a large pediatric teaching hospital, a system of inpatient care provided by hospitalists can reduce length of stay. This model has the potential to control hospital charges in a period of increasing health care costs. PMID- 10699097 TI - Outcome of children in the indomethacin intraventricular hemorrhage prevention trial. AB - BACKGROUND: For preterm infants, intraventricular hemorrhage (IVH) may be associated with adverse neurodevelopmental outcome. We have demonstrated that early low-dose indomethacin treatment is associated with a decrease in both the incidence and severity of IVH in very low birth weight preterm infants. In addition, we hypothesized that the early administration of low-dose indomethacin would not be associated with an increase in the incidence of neurodevelopmental handicap at 4.5 years of age in our study children. METHODS: To test this hypothesis, we provided neurodevelopmental follow-up for the 384 very low birth weight survivors of the Multicenter Randomized Indomethacin IVH Prevention Trial. Three hundred thirty-seven children (88%) were evaluated at 54 months' corrected age, and underwent neurodevelopmental examinations, including the Wechsler Preschool and Primary Scale of Intelligence-Revised (WPPSI-R), the Peabody Picture Vocabulary Test-Revised (PPVT-R), and standard neurologic examinations. RESULTS: Of the 337 study children, 170 had been randomized to early low-dose indomethacin therapy and 167 children had received placebo. Twelve (7%) of the 165 indomethacin children and 11 (7%) of the 158 placebo children who underwent neurologic examinations were found to have cerebral palsy. For the 233 English monolingual children for whom cognitive outcome data follow, the mean gestational age was significantly younger for the children who received indomethacin than for those who received placebo. In addition, although there were no differences in the WPPSI-R or the PPVT-R scores between the 2 groups, analysis of the WPPSI-R full-scale IQ by function range demonstrated significantly less mental retardation among those children randomized to early low-dose indomethacin (for the indomethacin study children, 9% had an IQ <70, 12% had an IQ of 70-80, and 79% had an IQ >80, compared with the placebo group, for whom 17% had an IQ <70, 18% had an IQ of 70-80, and 65% had an IQ >80). Indomethacin children also experienced significantly less difficulty with vocabulary skills as assessed by the PPVT-R when compared with placebo children. CONCLUSIONS: These data suggest that, for preterm neonates, the early administration of low-dose indomethacin therapy is not associated with adverse neurodevelopmental function at 54 months' corrected age. PMID- 10699098 TI - Predicting first-year relapses in children with nephrotic syndrome. AB - OBJECTIVE: More than half of the children diagnosed with nephrotic syndrome will have relapses. These can be infrequent relapses (IRs: <2 in 6 months or <3 in a year) or frequent relapses (FRs: >2 in 6 months or >3 in a year). Patients who relapse while on alternate day steroids or within 1 month of discontinuation of steroid therapy are considered steroid-dependent (SD; J Pediatr. 1982; 101:514 518). Patients with an IR course have a better long-term prognosis, and many of them have minimal-change disease without mesangial hypercellularity or sclerosis. The purpose of our study was to identify factors at initial presentation that could predict the relapse pattern in the first year after diagnosis, without taking into consideration the histopathology found on renal biopsy. DESIGN: We analyzed the medical records of children who were seen by us before March 1997 and followed for at least 1 year. Variables selected in the study were age, sex, race, presence or absence of hematuria, and days to remission (defined as protein free) at the initial presentation, because they could relate to the pattern of relapses (ie, IR, FR, and SD). RESULTS: Of 70 patients, 14 were excluded because of insufficient data. There were 38 males (67.9%) and 18 females (32.1%), giving a male:female ratio of 1.8:1. Median age at presentation was 3.25 years (range: 1.5-13), and 76.9% were white, 8.9% black, 7.1% Hispanic, and 7.1% other. Of all the patients, 23 were IR (41.1%), 9 were FR (16.1%), and 24 were SD (42. 9%). Median days to remission were 10 (range: 2-60), on Prednisone 60 mg/M(2) daily. Hematuria was present initially in 26 patients (46. 4%), and absent in 30 (53.6%). Age, sex, race, and hematuria, as independent variables, were not predictors of relapses in the first year. However, using a stratified analysis based on the presence or absence of hematuria, we found that if the remission occurred within the first week of therapy, the patients without hematuria were more likely to be IR. The sensitivity and specificity of this finding were 67% and 89%, respectively, with a positive predictive value of 94%. CONCLUSION: We conclude that of all the presenting features, the rapidity of initial response to steroid therapy combined with the presence of hematuria, could predict future relapses and should be well documented. PMID- 10699099 TI - Pediatricians' attitudes, beliefs, and practices regarding clinical practice guidelines: a national survey. AB - BACKGROUND: Clinical practice guidelines are increasingly being used for a wide variety of medical conditions, but not enough is known about physicians' attitudes and beliefs about guidelines, how often and under what circumstances they are used, and factors associated with their acceptance. OBJECTIVE: To determine practice guideline attitudes, beliefs, practices, and factors associated with use among a representative national sample of general pediatricians. STUDY DESIGN: Cross-sectional mail survey. SUBJECTS: Random sample of general pediatrician members of the American Academy of Pediatrics residing in all 50 states and Puerto Rico. SURVEY INSTRUMENT: Twenty-four multiple-choice, Likert scale, yes-no, and open-ended questions about pediatric clinical practice guidelines. RESULTS: From 1088 respondents, 461 specialists were excluded; the remaining 627 general pediatricians were mostly male (61%), white (81%), and in group practice (62%) in a suburban location (48%). Practice guidelines are used by 35% of pediatricians, in part by 44%, and not at all by 21%. Over 100 different practice guidelines are used, most commonly for asthma (77%), hyperbilirubinemia (27%), and otitis media (19%). Common reasons for use of practice guidelines include standardization of care (17%) and helpfulness (10%). Commonly cited problems with practice guidelines include failure to allow for clinical judgment (54%), use in litigation (16%), and limitation of autonomy (5%). In multivariate analysis, the odds of practice guideline use were greater among pediatricians in health maintenance organization practices (odds ratio [OR]: 9.1; 95% confidence interval [CI]: 1.2-68.0) and those who were nonwhite (OR: 2.3; 95% CI: 1.1-4.8), but lower in those with more weekly patient visits (OR:.7; 95% CI:.5-.9). Features most likely to lead to practice guideline use include simplicity (16%), feasibility (12%), and evidence of improved outcomes (10%). Most pediatricians agree that practice guidelines improve outcomes (89%), are motivated by a desire to improve quality (94%), and should not be used in litigation (82%) or disciplinary actions (77%), nor be motivated by a desire to reduce costs (73%). CONCLUSIONS: Most general pediatricians use practice guidelines, but no specific guidelines, except those for asthma, are used by >27% of pediatricians. The results of this study suggest that practice guidelines are most likely to be followed if they are simple, flexible, rigorously tested, not used punitively, and are motivated by desires to improve quality, not reduce costs. PMID- 10699100 TI - Reevaluation of outpatients with Streptococcus pneumoniae bacteremia. AB - BACKGROUND: The reevaluation process for outpatients recalled for Streptococcus pneumoniae bacteremia has not been standardized. Children who return ill or with new serious focal infections require admission and parenteral antibiotic therapy. Limited data exist to guide the follow-up management of those patients identified as having occult pneumococcal bacteremia. OBJECTIVES: Characterize the outcomes of outpatients with pneumococcal bacteremia based on their evaluation at follow up. For patients who are well-appearing without serious focal infection, propose a management scheme for reevaluation. METHODS: Retrospective review of outpatients with pneumococcal bacteremia. Patients with immunocompromise, those identified with focal bacterial infection at the initial visit, or those admitted at the initial visit were excluded. Data were collected from the initial visit (when blood culture drawn) and follow-up visit with regard to clinical parameters, laboratory data, diagnoses, and any antibiotic treatment. Decision tree analysis was used to generate a model to predict children at high risk for persistent bacteremia (PB). RESULTS: A total of 548 episodes of pneumococcal bacteremia were studied. Seventy-three children received no antibiotic, 239 oral antibiotic, and 236 parenteral antibiotic at the initial visit. Median age, temperature, and white blood cell (WBC) count were 13.5 months, 40.0 degrees C, and 20 400/mm(3). Forty-one patients had PB or new focal infections (15 with PB alone, 4 had focal infection and PB). Eight patients had meningitis at follow-up. Ninety-two percent returned because of notification of the positive blood culture result. A repeat blood culture was obtained in 92%, 23% had a lumbar puncture, 33% had a chest radiograph, and 12% were admitted. PB was associated with the antibiotic treatment group, elevation of temperature, and WBC count at follow-up. A simple management scheme using 2 sequential decision nodes of antibiotic treatment (none vs any) and then temperature at follow-up (>38.8 degrees C) would have predicted 16/19 patients with PB (sensitivity =.84 and specificity =.86). CONCLUSIONS: All patients with pneumococcal bacteremia need prompt reevaluation. For well-appearing patients without new focal infection, the utility of diagnostic testing (specifically repeat blood cultures) and the need for admission may be determined by the use of antibiotics at the initial evaluation and the presence of fever at follow-up. The majority of patients can be managed as outpatients entirely. Patients who did not receive antibiotics at the initial evaluation and those treated with oral antibiotics but remain febrile are at the highest risk for persistent bacteremia. PMID- 10699101 TI - Magnetic resonance imaging of intestinal necrosis in preterm infants. AB - BACKGROUND AND OBJECTIVE: Noninvasive diagnosis of intestinal necrosis is important in planning surgery in preterm infants with necrotizing enterocolitis (NEC). We aimed to assess the potential of magnetic resonance imaging (MRI) for the diagnosis of intestinal necrosis. STUDY PARTICIPANTS AND METHODS: Abdominal MRI scans were performed in a group of preterm infants with suspected NEC and compared with surgical findings and to MRI results in a group of control infants. In addition, MRI was performed in 2 preterm infants with suspected NEC who did not require surgery. RESULTS: Six infants with a median birth weight of 1220 g (range, 760-1770 g) and median gestational age at birth of 30 weeks (range, 28-34 weeks) were studied at a median postnatal age of 10 days (range, 4-19 days). Four infants had a bubble-like appearance in part of the intestinal wall, intramural gas, and an abnormal fluid level within bowel lumen. At surgery, NEC was found in 5 infants and sigmoid volvulus in 1. The site of the bubble-like appearance corresponded to the site of intestinal necrosis at surgery. Four control infants with a median birth weight of 1500 g (range, 730-2130 g) and a median gestational age of 31 weeks (range, 26-36 weeks) had abdominal MRI at a median postnatal age of 8 days (range, 4-70 days). None of the above findings were seen in any control infant. The bubble-like appearance was not seen in the 2 infants with suspected NEC who did not require surgery. CONCLUSION: Abdominal MRI allows the noninvasive diagnosis of bowel necrosis. This may aid the timing of surgical intervention in preterm infants with a clinical diagnosis of NEC.gangrene, ischemia, MRI, necrotizing enterocolitis. PMID- 10699102 TI - Psychoeducational outcome in children with early-treated congenital hypothyroidism. AB - OBJECTIVES: To describe the psychoeducational characteristics of children with congenital hypothyroidism (CH) identified through newborn screening and to study changes over time. METHOD: Examined were 83 children with early-treated CH, who were long-time participants in a prospective study of outcome after newborn screening, and 120 control children who were classmates (n = 80) or siblings (n = 42). Children were tested during the third (53 children with CH and 46 control children) or the sixth (51 children with CH and 76 control children) grades at school with 21 children with CH being seen in both grades. Test instruments included multiple measures of achievement and cognitive abilities as well as behavior rating scales completed by parents and teachers. RESULTS: CH was associated with a slightly increased risk of learning disabilities in grade 3 but not grade 6. Third grade CH children scored lower than control children on tests of reading comprehension and arithmetic but did not differ on word recognition, writing, or spelling. Sixth grade CH children performed similar to controls on basic achievement tests but were reported to be doing poorer in several subject areas. For children with CH in grade 3, delayed skeletal maturity at diagnosis was associated with poorer word recognition ability and a longer period for normalizing thyroid hormone in infancy was correlated with weaker skill in learning sound-symbol correspondences. CONCLUSION: Early-treated CH is associated with mild delays in several basic achievement areas (reading comprehension and arithmetic) at the third grade level, with catch up by the sixth grade. However, as other findings indicate cognitive problems do persist into adolescence in memory, attention, and visuospatial processing areas, the implications of these deficits for other educational accomplishments needs additional follow up.congenital hypothyroidism, thyroid hormone, newborn screening, achievement, behavior, attention. PMID- 10699103 TI - Rapid detection of microorganisms in blood cultures of newborn infants utilizing an automated blood culture system. AB - BACKGROUND: Neonatal sepsis is a low incidence, high-risk disease with many sepsis work-ups performed to detect a single case. Seventy-two hours of antibiotic therapy have been traditionally recommended pending negative culture results. Improved culture media and new technology integrated into blood culture systems could shorten incubation time required to detect positive culture results. This would then change the length of antibiotic therapy in the management of the newborn infant with suspected sepsis. In addition, previous data supporting the 72-hour recommendation were retrospectively acquired, utilized nonautomated systems, and reported in an era with a different population of microorganisms cultured in special care nurseries. OBJECTIVE: Evaluate the time of incubation to detect positive blood cultures from newborn infants with suspected sepsis using a computer-assisted, automated blood culture system, ESP (Trek Diagnostic Systems, Inc, Westlake, OH). DESIGN: Prospective, observational study. PATIENTS AND SETTING: All positive blood culture results that were obtained from term and preterm newborn infants born from November 1993 through June 1997 at a publicly funded hospital with over 6000 live births per year. METHODS: As positive blood culture results were identified, data were prospectively obtained from the patient's medical record. The computer algorithm in the automated blood culture system determined the time to positivity. Time to positivity was determined for blood cultures obtained before the initiation antimicrobial therapy and compared with those cultures obtained after beginning therapy. Time to positivity was also evaluated for clinically important Gram positive and Gram-negative bacteria and yeast. RESULTS: Four hundred fifty-five positive blood culture results were obtained from 222 patients. Gram-positive organisms accounted for 80% (366/455) of the positive culture results, Gram negative organisms accounted for 11% (48/455), and yeast for 9% (41/455). Virtually all cultures growing clinically significant Gram-positive and Gram negative organisms were positive by 24 to 36 hours of incubation. Cultures growing Staphylococcus epidermidis were virtually all positive after 36 to 48 hours of incubation. Of cultures growing yeast, 88% (36/41) were positive by 48 hours of incubation. There was no difference in time to positivity in pretherapy or posttherapy obtained positive blood cultures. Prenatally administered antibiotics did not affect time to positivity in positive cultures drawn on the first day of life. In a selected group of microorganisms that are the frequent cause of bacteremia in term infants, 97% and 99% of cultures were positive by 24 to 36 hours of incubation when only pretherapy cultures are evaluated. CONCLUSIONS: The ESP blood culture system identified 77%, 89% and 94% of all microorganisms at 24, 36, and 48 hours of incubation in aerobic cultures obtained from both term and preterm infants. Introduction of antimicrobial therapy did not affect time to positivity. Reducing duration of antibiotic therapy to 24 to 36 hours should be considered in term, asymptomatic newborn infants undergoing evaluation for suspected sepsis for maternal indications. Confirmation of similar rapidity of detection using other blood culture systems should be undertaken. PMID- 10699104 TI - Systemic to pulmonary collaterals in very low birth weight infants: color doppler detection of systemic to pulmonary connections during neonatal and early infancy period. AB - OBJECTIVE: Angiographic visualization of systemic to pulmonary collaterals (SPC) has been documented in premature infants needing prolonged ventilatory support. Noninvasive identification of such communications in premature infants was reported recently. The purpose of this study was to describe: 1) incidence, 2) clinical findings and implications, and 3) short-term follow-up of SPC diagnosed by echocardiography in very low birth weight (VLBW) infants admitted to the neonatal intensive care unit. METHODS: From December 1, 1994 to August 31, 1996, 196 infants with birth weight <1500 g were admitted to the neonatal intensive care unit; 133 of them received serial echocardiographic evaluations at 1 to 2 days, at 2 weeks, and at 1, 2, and 3 months of life. Follow-up echocardiograms were scheduled at 6 months and 1 year of age for patients with SPC persisting at 3 months of age. RESULTS: SPC were demonstrated in 88 patients (66%) at 1 to 90 days of life (mean 28 days). In most cases, the SPC originated at the distal aortic arch or the proximal descending aorta. Ten patients (11%) were treated for congestive heart failure. The symptoms improved and anticongestive therapy was discontinued in 9. One patient with persistent congestive heart failure underwent therapeutic cardiac catheterization and 1 prominent SPC was embolized. CONCLUSIONS: The incidence of SPC in VLBW infants is much higher than previously reported. We postulate that SPC are bronchopulmonary communications that enlarge and/or proliferate in response to a given stimulus. These communications are associated with increased time on positive pressure ventilation and length of stay in the hospital. SPC may lead to pulmonary edema and should be searched for in VLBW infants with a more complicated course. Echocardiographic examination with color Doppler performed in premature infants to evaluate left to right shunts should include careful search for systemic to pulmonary collaterals.echocardiography, systemic to pulmonary collaterals, aortopulmonary collaterals, prematurity, pulmonary edema. PMID- 10699105 TI - Predischarge bilirubin screening in glucose-6-phosphate dehydrogenase-deficient neonates. AB - OBJECTIVE: To assess the validity of predischarge serum bilirubin values in determining or predicting hyperbilirubinemia in glucose-6-phosphate dehydrogenase (G-6-PD)-deficient neonates, and to facilitate appropriate discharge planning. METHODS: Serum total bilirubin values were determined between 44 and 72 hours of life in a cohort of term, healthy neonates at high-risk for G-6-PD deficiency but with no other risk factors for hyperbilirubinemia. Percentile-based bilirubin nomograms were constructed for G-6-PD-deficient infants and normal infants according to age at sampling. The incidence of hyperbilirubinemia (serum bilirubin value > or =256 micromol/L [15 mg/dL]) for each group was determined according to the percentiles for that group. RESULTS: In both G-6-PD-deficient neonates (n = 108) and control neonates (n = 215) with serum bilirubin values <50th percentile for age, the incidence of hyperbilirubinemia was low in the G-6 PD-deficient neonates, with no measurable incidence in the controls. The incidence of hyperbilirubinemia became clinically consequential, and significantly higher in the G-6-PD-deficient groups, when the percentiles were > or =50: for those in the 50% to 74% range the incidence was moderate (23%) for the G-6-PD-deficient and small (7%) for the control infants (relative risk, 3.29; 95% confidence interval, 1.01-10.67). Among those infants > or =75th percentile, 82% of the G-6-PD-deficient infants, compared with 25% of the control infants, were either already hyperbilirubinemic at the time of screening or subsequently developed hyperbilirubinemia (relative risk, 3.23; 95% confidence interval, 1.99 5.24). CONCLUSIONS: Timed, predischarge serum bilirubin screening can be used to identify G-6-PD-deficient neonates at low, intermediate, or high-risk of developing severe neonatal hyperbilirubinemia, and thus offer a selective approach to the discharge and follow-up surveillance of these infants. PMID- 10699106 TI - Population-based estimates of surfactant protein B deficiency. AB - OBJECTIVE: Surfactant protein B deficiency is a lethal cause of respiratory distress in infancy that results most commonly from a homozygous frameshift mutation (121ins2). Using independent clinical ascertainment and molecular methods in different populations, we sought to determine allele frequency. STUDY DESIGN: Using clinical characteristics of the phenotype of affected infants, we screened the Missouri linked birth-death database (n = 1 052 544) to ascertain potentially affected infants. We used molecular amplification and restriction enzyme digestion of DNA samples from a metropolitan New York birth cohort (n = 6599) to estimate allele frequency. RESULTS: The point estimate and 95% confidence interval of the 121ins2 allele frequency in the Missouri cohort are 1/1000 individuals (.03-5.6/1000) and in the New York cohort are.15/1000 (. 08 .25/1000). These estimates are not statistically different. CONCLUSIONS: The close approximation of these independent estimates suggests accurate gene frequency (approximately one 121ins2 mutation per 1000-3000 individuals) despite its rare occurrence and that this mutation does not account for the majority of full-term infants with lethal respiratory distress. PMID- 10699107 TI - Early dexamethasone-attempting to prevent chronic lung disease. AB - BACKGROUND: We previously demonstrated improved survival and early outcomes in a pilot trial of 2 doses of intravenous dexamethasone for infants with surfactant treated respiratory distress syndrome. (1) A multicenter, randomized, double blind trial was undertaken to confirm these results. METHODS: Infants <30 weeks' gestation were eligible if they had respiratory distress syndrome, required mechanical ventilation at 12 to 18 hours of age, and had received at least 1 dose of exogenous surfactant. Infants were excluded if sepsis or pneumonia was suspected or if congenital heart disease or chromosomal abnormalities were present. A total of 384 infants were enrolled-189 randomized to dexamethasone (.5mg/kg birth weight at 12-18 hours of age and a second dose 12 hours later) and 195 to an equal volume of saline placebo. RESULTS: No differences were found in the dexamethasone versus placebo groups, respectively, regarding the primary outcomes of survival (79% vs 83%), survival without oxygen at 36 weeks' corrected gestational age (CGA; both 59%), and survival without oxygen at 36 weeks' CGA and without late glucocorticoid therapy (46% vs 44%). No significant differences between the groups in estimates from Kaplan-Meier survival analyses were found for median days on oxygen (50 vs 56 days), ventilation (20 vs 27 days), days to regain birth weight (15.5 vs 14 days), or length of stay (LOS; 88 vs 89 days). Infants given early dexamethasone were less likely to receive later glucocorticoid therapy for bronchopulmonary dysplasia during their hospitalization (27% vs 35%). No clinically significant side effects were noted in the dexamethasone group, although there were transient elevations in blood glucose and blood pressure followed by a return to baseline by study day 10. Among infants who died (40 vs 33), there were no differences in the median days on oxygen, ventilation, nor LOS. However, in survivors (149 vs 162), the following were observed: median days on oxygen 37 versus 45 days, ventilation 14 versus 19 days, and LOS 79 versus 81 days, for the dexamethasone versus placebo groups, respectively. CONCLUSIONS: This dose of early intravenous dexamethasone did not reduce the requirement for oxygen at 36 weeks' CGA and survival was not improved. However, early dexamethasone reduced the use of later prolonged dexamethasone therapy, and among survivors, reduced the median days on oxygen and ventilation. We conclude that this course of early dexamethasone probably represents a near minimum dose for instituting a prophylactic regimen against bronchopulmonary dysplasia. PMID- 10699108 TI - Evaluation of infants with subdural hematoma who lack external evidence of abuse. AB - OBJECTIVE: Advances in radiologic technique have increased the recognition of subdural hematoma. No study to date has addressed the role of child protective investigation into the cause and management of subdural hematoma in children who lack other indicators of abuse. METHODS: Medical records, radiology studies, and social service notes for all infants and children referred for child abuse investigation who had any form of intracranial hemorrhage were reviewed. The study covered the 12 months of 1997. All referrals were to the Suncoast Child Protection Team (St Petersburg, FL). RESULTS: There were 19 investigations because of subdural hematoma. Eight children had retinal hemorrhage as well as other major findings of trauma, such as bruises and/or fractures; all 8 were victims of child abuse. Two infants had tiny subdurals adjacent to accidental linear skull fractures. Nine infants were investigated for the possibility of abuse that had no findings of trauma apart from the subdural hematoma. These 9 cases form the basis for this study. The age range was 11 days to 15 months. Inflicted cerebral trauma was the medical diagnosis in 8 of the 9 cases; 1 case had a final diagnosis of possible inflicted injury in a high-risk setting. CONCLUSIONS: Infants with subdural hematoma but no other findings of abuse present a difficult challenge to child protection workers. Investigation by a medically oriented team can uncover the circumstances of the trauma in most instances and can usefully direct protective efforts. The high incidence of severe sequelae in infants with inflicted cerebral trauma warrants a vigorous approach. PMID- 10699109 TI - Training of pediatricians in care of physical disabilities in children with special health needs: results of a two-state survey of practicing pediatricians and national resident training programs. AB - OBJECTIVE: One goal of the American Academy of Pediatrics' Future of Pediatric Education II Project is to establish guidelines in training physicians to care for children with special health care needs (CWSN). Assessment of current practices in prescribing therapies and devices is necessary to meet this goal. Although much has been written about CWSN, there is a paucity of literature describing pediatricians' preparedness in prescribing such therapies and devices to children with physical disabilities. In an effort to assess physician preparedness, we surveyed pediatric residents nationwide and practicing pediatricians from 2 states, 1 urban and 1 rural. METHODS. A questionnaire aimed at identifying areas of concern regarding preparedness of physicians in practice and in training was prepared and mailed to prospective participants in Ohio and Mississippi. After follow-up mailings to nonresponders, approximately 59% responded. Summary statistics were reported as proportions with 95% confidence intervals. RESULTS: Among those polled, >70% reported no training in prescribing certain durable medical equipment and over 50% reported no training in prescribing certain therapies. In addition, at least 20% reported no training in treating some of the more common childhood physical disabilities. Nearly three fourths of the respondents indicated that they did not believe that they were adequately prepared to take an active role in prescribing therapies and durable medical equipment. Fewer respondents believed that they should be the sole providers of these therapies and durable medical equipment. CONCLUSIONS: The results of the survey indicate a lack of specific training and physician confidence in prescribing therapies and devices to CWSN, establishing the necessity of expanding training programs to better ensure quality health care for special needs children. Although additional ongoing research is necessary to fully evaluate the preparedness of physicians in caring for CWSN, this survey does help to identify areas of physician training that require improvement to provide quality health care for CWSN. PMID- 10699110 TI - High-risk periods for childhood injury among siblings. AB - OBJECTIVE: To determine whether the risk of unintentional injury requiring emergency department (ED) or inpatient care in children is transiently increased over a 90-day period after injury to a sibling. DESIGN: Retrospective cohort. SETTING: King County, Washington. Participants. A total of 41 242 children 0 to 15 years of age continuously enrolled in Medicaid and living in King County during the period October 1, 1992 through September 30, 1993 (27 450 child years). OUTCOME MEASURES: The outcome was an unintentional injury treated in the ED or inpatient setting. Incidence rates and hazard ratios were calculated for children whose sibling had been injured in the previous 90 days, compared with children without such exposure. Multivariate analysis was used to adjust for age, gender, race, sibling group size, and noninjury ED use. RESULTS: . There were 4921 injuries treated only in the ED and 82 hospital admissions. The incidence of ED treated injury was 305 per 1000 child-years among children whose sibling had been injured in the previous 90 days and 174 per 1000 child-years among children without this exposure (relative risk: 1.75; 95% confidence interval: 1.56-1.95). The incidence of injury-related hospitalization was 1.7 per 1000 child-years among children whose sibling had been injured in the previous 90 days, compared with 3.0 per 1000 child-years among children without this exposure (relative risk:.57; 95% confidence interval:.07-2.12). Injury risk peaked in the period 4 to 10 days after a sibling's injury and returned toward, but did not attain, baseline risk over the subsequent 21/2 months. The magnitude of this effect depended on the child's age; the relative risk of injury was higher among older children. CONCLUSIONS: Injuries treated in the ED or inpatient setting appear to cluster within sibling groups over brief periods of time. Shared social or environmental exposures may contribute to this clustering and may be amenable to targeted, time-limited prevention interventions. PMID- 10699111 TI - Parental perspectives of the health status and health-related quality of life of teen-aged children who were extremely low birth weight and term controls. AB - OBJECTIVES: To compare the health status and health-related quality of life of teen-aged children who were extremely low birth weight (ELBW) with matched controls from the perspective of their parents. STUDY DESIGN: Geographically defined cohort; longitudinal follow-up; cross-sectional interviews. PARTICIPANTS: parents of 149/169 (88%) ELBW children between 12 and 16 years of age (including 41 children with neurosensory impairments) and 126/145 (87%) parents of term controls. Health status of the teenagers was classified according to the 6 attributes of the Health Utilities Index Mark 2, based on information obtained during parent interviews. Parents were asked to imagine themselves living in their own child's health state and 4 preselected hypothetical health states when providing directly measured standard gamble utility scores. RESULTS: Parents of ELBW children reported a higher frequency and more complex functional limitations than parents of controls for their own children's health status. Also, the mean utilities were lower (ELBW =.91 vs controls =. 97) and the variability in their scores was greater. There were no differences in the valuation of the hypothetical health states provided by parents of ELBW and control children. CONCLUSIONS: ELBW children were reported to have a greater burden of disability than were control children based on parental descriptions. Nonetheless, parents of ELBW children, on average, rated the health-related quality of life of their children fairly high. Thus, differences in reported functional status are not necessarily associated with lower utility scores. PMID- 10699112 TI - Medical education about end-of-life care in the pediatric setting: principles, challenges, and opportunities. AB - OBJECTIVE: To identify the opportunities for and barriers to medical education about end-of-life (EOL) care in the pediatric setting. METHODS: A working group of pediatric specialists and ethicists was convened at the National Consensus Conference on Medical Education for Care Near the End-of-Life sponsored by the Open Society Institute's Project Death in America and the Robert Wood Johnson Foundation. The charge to the working group was to consider the unique aspects of death in childhood, identify critical educational issues and effective instructional strategies, and recommend institutional changes needed to facilitate teaching about EOL care for children. CONCLUSIONS: Although providing EOL care can be challenging, the cognitive and psychologic skills needed can be taught effectively through well-planned and focused learning experiences. The ultimate goals of such instruction are to provide more humane care to very sick children, enhance bereavement outcomes for their survivors, and develop more confident clinicians. Six specific principles regarding EOL care in the pediatric setting emerged as essential curricular elements that should be taught to all medical care providers to ensure competent patient-centered care. 1) Cognitively and developmentally appropriate communication is most effective. 2) Sharing information with patients helps avoid feelings of isolation and abandonment. 3) The needs of the patient are served when the ethical principles of self determination and best interests are central to the decision-making process. 4) Minimization of physical and emotional pain and other symptoms requires prompt recognition, careful assessment, and comprehensive treatment. 5) Developing partnerships with families supports them in their caregiving efforts. 6) The personal and professional challenges faced by providers of EOL care deserve to be addressed. These principles actually transcend patient age and can be used to inform medical education about the care of any terminally ill patient. Similarly, these principles of effective communication, ethical decision-making, and attention to the quality of life of patients, families, and providers apply to the care of all children regardless of diagnosis and prognosis. With this in mind, teaching about EOL care does not require a new and separate curriculum, but rather taking better advantage of the many teachable moments provided by caring for a dying patient. PMID- 10699113 TI - Beliefs about diagnosing asthma in young children. AB - OBJECTIVES: To determine what factors primary care pediatricians believe are important in establishing the initial diagnosis of childhood asthma and to identify variations in physicians' beliefs concerning this clinical decision. STUDY DESIGN: Massachusetts American Academy of Pediatrics Fellows were surveyed about their beliefs concerning the importance of 20 clinical factors associated with establishing the initial diagnosis of asthma. RESULTS: Most clinicians considered recurrent wheeze (96%), symptomatic improvement with a bronchodilator (90%), recurrent cough (89%), exclusion of alternative diagnoses (87%), and suggestive peak flow findings (82%) as important in diagnosing asthma. However, there was substantial heterogeneity among clinicians as to which combinations of factors they each considered relevant; for example, only 60% identified all 5 of the above factors to be necessary or important. Further, <50% identified presence of any of the 20 factors as necessary. Although national guidelines cite objective assessment of pulmonary function as essential, spirometry and peak expiratory flow testing were identified as necessary by only 8% and 10%, respectively. Two factors believed to contribute to establishing the asthma diagnosis contradicted the National Asthma Education and Prevention Program guidelines and expert opinion (age >2 years and absence of fever during episodes) and these beliefs were more likely held by those clinicians in practice for >5 years. CONCLUSIONS: The majority of pediatricians believe several common clinical factors establish a diagnosis of childhood asthma, but disagree over what combinations of these factors are important. Some misconceptions persist despite wide dissemination of clinical practice guidelines. We believe that future asthma guidelines will need to organize diagnostic criteria in an easily understood format, like a decision tree, to facilitate early recognition of asthma in young children. PMID- 10699114 TI - Informed proxy consent: communication between pediatric surgeons and surrogates about surgery. AB - OBJECTIVE: Informed consent for surgical procedures requires that the procedures are explained and that the patient understands the procedures and risks and agrees to undergo them. Proxy consent occurs when an individual is provided with the legal right to make decisions on behalf of another. This study was conducted to determine how surgeons communicate information to obtain an informed proxy consent, and to investigate how that information is received and processed by surrogates responsible for providing such consent. STUDY DESIGN: Twenty English speaking parents or legal guardians and 5 surgeons in an urban pediatric hospital were interviewed before, and 2 to 4 weeks after, the surgical procedure. In addition, the interview between the surgeon and surrogate, when consent was obtained, was audiotaped and subsequently analyzed. Semistructured interviews were used to elicit the motivations and influences on the surrogates to consent to the procedure. The same methodology was used to elicit the corresponding impressions of the surgeons. The data were analyzed using descriptive statistics and crosstabulations. RESULTS: Demographic data did not influence the results. Although there was concordance between the surrogate's understanding of the procedure and the surgeon's impression of this understanding, only 3 of 17 surrogates could recall any specifics of the explained procedure. Contrary to the stated belief of surgeons, surrogates consulted with a variety of others, including medical and paramedical professionals, family members, and spiritual leaders. CONCLUSIONS: Communication plays an important role within the surrogate surgeon dyad. Psychologic variables such as expectations, and the perception of both the surrogates and the surgeons, influence the amount of information that is proffered and the manner in which it is received. Improved communication may be achieved by use of visual aids, discussion of anesthesia and the postoperative course, recognition of the circumstances around the discussion, such as timing and location of the discussion, and personalization of the discussion. PMID- 10699115 TI - Childhood encephalopathies and myopathies: a prospective study in a defined population to assess the frequency of mitochondrial disorders. AB - OBJECTIVES: To assess the frequency of mitochondrial abnormalities in muscle histology, defects in respiratory chain enzyme activities, and mutations in mitochondrial DNA (mtDNA) in children with unexplained psychomotor retardation in the population of Northern Finland. BACKGROUND: The frequency of mitochondrial diseases among patients with childhood encephalopathies and myopathies is not known. Frequencies are difficult to estimate because the clinical presentation of these disorders is variable. METHODS: A total of 116 consecutive patients with undefined encephalopathies and myopathies were enrolled during a 7-year period in a hospital serving as the only neurologic unit for a pediatric population of 97 609 and as the only tertiary level neurologic unit for a pediatric population of 48 873. Biochemical and morphologic investigations were performed on muscle biopsy material, including oximetric and spectrophotometric analyses of oxidative phosphorylation, histochemistry, electron microscopy, and molecular analysis of mtDNA. RESULTS: Ultrastructural changes in the mitochondria were the most common finding in the muscle biopsies (71%). Ragged-red fibers were found in 4 cases. An oxidative phosphorylation defect was found in 26 children (28%), complex I (n = 15) and complex IV (n = 13) defects being the most common. Fifteen percent of patients (n = 17/116) with unexplained encephalomyopathy or myopathy had a probable mitochondrial disease. Common pathogenic mutations were found in the mtDNA of only 1 patient (.9%). CONCLUSIONS: The common known mutations in mtDNA are rarely causes of childhood encephalomyopathies, which is in contrast to the considerable frequency of the common MELAS mutation observed among adults in the same geographical area. Biochemically and morphologically verified mitochondrial disorders were nevertheless common among the children, making the analysis of a muscle biopsy very important for clinical diagnostic purposes. PMID- 10699116 TI - An analysis of clinical outcomes using color doppler testicular ultrasound for testicular torsion. AB - OBJECTIVES: To delineate the clinical outcomes of color Doppler ultrasound (US) in the equivocal torsion patient. METHODS: From 1992 to 1997, 130 patients (<23 years old) from 2 institutions underwent US imaging using a 7.5-mHz linear transducer to evaluate an acute scrotum equivocal, or of low suspicion, for torsion. The US reports and hospital charts of these patients were retrospectively reviewed. RESULTS: After clinical and radiologic evaluation, torsion was excluded in 110 patients without surgical exploration. In 3 patients, intermittent testicular torsion was diagnosed and in 17 patients, emergent exploration was performed for US diagnosis of testicular torsion. Twenty-five patients (22.7%) were subsequently lost to follow-up. Follow-up of 85 patients with US negative for torsion (mean length of follow-up = 466.9 days) revealed no testicular atrophy in 83. Two patients underwent delayed orchiectomy/contralateral orchiopexy for missed testicular torsion. Of 17 patients with US positive for torsion, 9 underwent orchiectomy for a necrotic torsed testis, 7 viable torsed testes were found, and 1 torsed appendix testis was found. Therefore, color Doppler US for the equivocal acute scrotum yielded a 1% false-positive rate, sensitivity of 88.9%, and specificity of 98.8%. CONCLUSION: When faced with ruling out testicular torsion, it is necessary to integrate the multiple pieces of patient data, knowing that each piece of data may have inaccuracies. With this in mind, this analysis of outcomes verifies that color Doppler US is an excellent adjunctive study in the clinically real situation in which the clinical evaluation is equivocal or low suspicion. PMID- 10699117 TI - Use of the national institutes of health criteria for diagnosis of neurofibromatosis 1 in children. AB - OBJECTIVE: The National Institutes of Health (NIH) Diagnostic Criteria for neurofibromatosis 1 (NF1) are very useful clinically, but some individuals who are later shown to have NF1 cannot be diagnosed in early childhood using these criteria. The aim of this study is to determine the value of the NIH Diagnostic Criteria for NF1 in early childhood, to determine the age at which diagnosis can confidently be made, and to clarify the age at onset of the cardinal clinical features used in the NIH Diagnostic Criteria. METHODS: We studied 1893 NF1 patients under 21 years old from the National Neurofibromatosis Foundation International Database to determine the age at which the features included in the NIH Diagnostic Criteria appear. RESULTS: Approximately 46% of sporadic NF1 cases fail to meet the NIH Diagnostic Criteria by 1 year of age. Nearly all (97%; 95% confidence interval: 94-98) NF1 patients meet the criteria for diagnosis by 8 years old, and all do so by 20 years old. The usual order of appearance of the clinical features listed as NIH criteria is cafe-au-lait macules, axillary freckling, Lisch nodules, and neurofibromas. Symptomatic optic glioma is usually diagnosed by 3 years old, and characteristic osseous lesions are usually apparent within the first year of life. CONCLUSION: The diagnosis of NF1 cannot always be made in young children using the NIH Diagnostic Criteria. Modification of these criteria may be necessary for children under 8 years old. PMID- 10699118 TI - Snowmobile injuries and deaths in children: a review of national injury data and state legislation. AB - BACKGROUND: Snowmobiling is a popular family sport, with annual expenditures over $9 billion. The size and speed of snowmobiles make them potentially dangerous to children. Pediatric snowmobile-related trauma has not been studied in the United States. METHODS: We analyzed 291 pediatric snowmobile- related injuries and 75 deaths reported to the Consumer Product Safety Commission from 1990 to 1998. We reviewed snowmobile legislation in the states that reported at least 1 death to the Consumer Product Safety Commission during this time period. RESULTS: The most common sites of injury were the extremities (48.8%) and the head, neck, and face (28.2%). Head and neck injuries were the predominant cause of death (66.7%). The most common diagnosis was contusion/abrasion (30.9%), followed by laceration (22%), fracture (20.3%), and strain/sprain (14.4%). Nonfatal injuries most often involved ejection from the snowmobile (26.1%), but striking a stationary object was the most common mechanism in fatal crashes. The review of state legislation revealed that few age restrictions or helmet laws exist. Children as young as 8 years old may legally operate a snowmobile in some states. Often, restrictions do not apply to snowmobile use on private property, where 43% of pediatric snowmobile-related injuries occurred. CONCLUSIONS: Head, neck, and face injuries are common nonfatal injuries and are the most common cause of death. State legislation often lacks age restrictions on private property, and laws requiring helmet use are rare. Legislators have not addressed the dangers of pediatric snowmobile-related injuries. Helmet laws and age restrictions similar to those enacted for motorcycle riders are necessary and appropriate. PMID- 10699119 TI - New policy on circumcision--cause for concern. PMID- 10699120 TI - How can we respond effectively to juvenile crime? PMID- 10699121 TI - Cholesterol screening in children and adolescents. PMID- 10699122 TI - Child health policymaking. PMID- 10699123 TI - The first video-based study in Pediatrics. PMID- 10699124 TI - Circumcision debate. Task Force on Circumcision, 1999-2000. PMID- 10699125 TI - Health appraisal guidelines for day camps and resident camps. American Academy of Pediatrics. Committee on School Health. AB - The American Academy of Pediatrics recommends that specific guidelines be established for pre-camp health appraisals of young people in day and resident camps. Camp guidelines also should include reference to health maintenance, storage and administration of medication, and emergency medical services. Although camps have diverse environments, there are general guidelines that apply to all situations, and specific recommendations are appropriate under special conditions. PMID- 10699126 TI - Access to pediatric emergency medical care. American Academy of Pediatrics. Committee on Pediatric Emergency Medicine. AB - Hundreds of thousands of pediatric patients require some level of emergency care annually, and significant barriers limit access to appropriate services for large numbers of children. The American Academy of Pediatrics has a strong commitment to identify barriers to access to emergency care, work to surmount these obstacles, and encourage through education increased levels of emergency care available to all children. It is also crucial to involve and incorporate the child's medical home into emergency care, both during acute presentation when the medical home is identified and by assisting in locating a medical home for follow up when none previously exists. PMID- 10699127 TI - Changing concepts of sudden infant death syndrome: implications for infant sleeping environment and sleep position. American Academy of Pediatrics. Task Force on Infant Sleep Position and Sudden Infant Death Syndrome. AB - The American Academy of Pediatrics has recommended since 1992 that infants be placed to sleep on their backs to reduce the risk of sudden infant death syndrome (SIDS). Since that time, the frequency of prone sleeping has decreased from >70% to approximately 20% of US infants, and the SIDS rate has decreased by >40%. However, SIDS remains the highest cause of infant death beyond the neonatal period, and there are still several potentially modifiable risk factors. Although some of these factors have been known for many years (eg, maternal smoking), the importance of other hazards, such as soft bedding and covered airways, has been demonstrated only recently. The present statement is intended to review the evidence about prone sleeping and other risk factors and to make recommendations about strategies that may be effective for further reducing the risk of SIDS. This statement is intended to consolidate and supplant previous statements made by this Task Force. PMID- 10699128 TI - Safety in youth ice hockey: the effects of body checking. American Academy of Pediatrics. Committee on Sports Medicine and Fitness. AB - Ice hockey is a sport enjoyed by many young people. The occurrence of injury can offset what may otherwise be a positive experience. A high proportion of injuries in hockey appear to result from intentional body contact or the practice of checking. The American Academy of Pediatrics recommends limiting checking in hockey players 15 years of age and younger as a means to reduce injuries. Strategies such as the fair play concept can also help decrease injuries that result from penalties or unnecessary contact. PMID- 10699129 TI - Injuries in youth soccer: a subject review. American Academy of Pediatrics. Committee on Sports Medicine and Fitness. AB - The current literature on injuries in youth soccer, known as football worldwide, has been reviewed to assess the frequency, type, and causes of injuries in this sport. The information in this review serves as a basis for encouraging safe participation in soccer for children and adolescents. PMID- 10699130 TI - Chemical-biological terrorism and its impact on children: a subject review. American Academy of Pediatrics. Committee on Environmental Health and Committee on Infectious Diseases. AB - There is an increasing threat that chemical and biological weapons will be used on a civilian population in an act of domestic terrorism. Casualties among adults and children could be significant in such an event. Federal, state, and local authorities have begun extensive planning to meet a chemical-biological incident by developing methods of rapid identification of potential agents and protocols for management of victims without injury to health care personnel. Because children would be disproportionately affected by a chemical or biological weapons release, pediatricians must assist in planning for a domestic chemical-biological incident. Government agencies should seek input from pediatricians and pediatric subspecialists to ensure that the situations created by multiple pediatric casualties after a chemical-biological incident are considered. This statement reviews key aspects of chemical-biological agents, the consequences of their use, the potential impact of a chemical-biological attack on children, and issues to consider in disaster planning and management for pediatric patients. PMID- 10699131 TI - Type 2 diabetes in children and adolescents. American Diabetes Association. PMID- 10699132 TI - Does mothering a doll change teens' thoughts about pregnancy? AB - OBJECTIVE: To determine the effect of age on the efficacy of the computerized, infant simulator doll Baby Think It Over (BTIO) for increasing middle school girls' knowledge about the responsibilities of parenthood and discouraging plans for teen childbearing. We hypothesized: 1) 8th grade students would be less apt than 6th grade students to equate BTIO care with mothering because they would rationalize that their infant would be easier to care for than BTIO; and 2) BTIO would be a more effective teen pregnancy prevention program with 6th grade students than with 8th grade students. METHODS: Nulliparous 6th (n = 68) and 8th (n = 41) grade girls attending an urban middle school in a predominantly lower socioeconomic, Hispanic, neighborhood were asked to care for BTIO for 3 days and 2 nights. Responses to a self-administered questionnaire were used to assess the girls' understanding of the responsibilities and difficulties associated with parenting, their feelings about the similarity of BTIO care and real infant care, and their childbearing intentions before and after caring for BTIO. RESULTS: Only 32 (29%) of the 109 girls thought that real infant care would be like BTIO care. Although 8th grade students were less apt than 6th grade students to equate BTIO care with real infant care (17% vs 37%), 6th grade students were more likely than 8th grade students to endorse statements suggesting that real infant care would be easier than BTIO care (37% vs 24%). Multivariate analyses revealed that this was largely because 6th grade students found BTIO care more difficult than did 8th grade students. Also, regardless of age or grade, the more difficult a girl found it to care for BTIO than anticipated, the more likely she was to endorse statements indicating that it would be easier to care for her own infant than it had been for her to care for BTIO. Little learning about the difficulties of parenting took place during the study. On average, the 6th grade students did not find BTIO care more difficult than anticipated and the 8th grade students actually found it easier than anticipated. Finally, caring for BTIO had no affect on the intent of students to become teen parents; 13 (12%) of the 109 students wanted to be teen parents before they cared for BTIO and 16 (15%) wanted to be teen parents after they cared for the doll. CONCLUSION: The results of this study demonstrate that the propensity of people this age for rationalizing their own immunity to the nocuous aspects of potentially desirable situations (the personal fable of omnipotence) allows those who perceive parenthood to be attractive to overlook the negative aspects of any parenting experience they have. PMID- 10699133 TI - Clinical validation of a polymerase chain reaction assay for the diagnosis of pertussis by comparison with serology, culture, and symptoms during a large pertussis vaccine efficacy trial. AB - OBJECTIVE: To assess the diagnostic sensitivity and specificity of a Bordetella pertussis polymerase chain reaction (PCR) assay using nasopharyngeal (NP) specimens from subjects with cough illnesses participating in a large pertussis vaccine efficacy trial. DESIGN: From 1991 to 1994, we conducted a large pertussis vaccine efficacy trial in Germany to determine the efficacy of the Lederle/Takeda acellular pertussis component diphtheria-tetanus toxoids in comparison with the Lederle whole-cell component diphtheria-tetanus toxoids vaccine. In the final year of the follow-up period of this trial, a second NP specimen for PCR, in addition to a culture specimen and blood for specific serology (enzyme-linked immunosorbent assay), was collected by use of a Dacron swab in subjects or family members with cough illnesses >/=7 days duration or in subjects with exposure to a cough illness in a household member to establish a diagnosis of B pertussis infection. Oligonucleotide primers (pTp1 and pTp2) that amplify a 191-bp-sized DNA fragment from the pertussis toxin operon, which is specific for B pertussis, were used. The PCR-amplified products were visualized by dot blot analysis followed by hybridization with a digoxigenin labeled probe and rated as 1+, 2+, or 3+ in comparison with positive controls representing approximately 1 to 10, 11 to 50, and >50 B pertussis organisms, respectively. In the present analysis, we compare PCR findings with those of serology, culture, positive household contact, and clinical characteristics of cough illnesses. RESULTS: Of 392 subjects with NP specimens obtained for PCR, 376 also had NP specimens collected for culture and 282 had serum specimens. PCR and culture were positive in 86 (22%) and 23 (6%) subjects, respectively. Of the positive PCR specimens, 40 were rated 3+, 32 were rated 2+, and 14 were rated 1+; 3+ positive specimens were more prevalent among DT recipients compared with pertussis vaccine recipients. Illnesses in subjects with 3+ positive PCR results were more typical of pertussis than were those in subjects with 2+ and 1+ positive results with a mean duration of cough of 48 days versus 43 and 42 days, respectively; presence of paroxysms, whoop or vomiting in 38% versus 17% and 10%, respectively; and a clinical diagnosis of definite or probable pertussis by the investigators of 26% versus 7% and 4%, respectively. Using serologic evidence of infection as the standard, sensitivity of PCR was 61%, and specificity was 88%. For 3+ positive PCR results, the respective values were 42% and 97%. CONCLUSION: Our findings demonstrate that PCR is more sensitive than conventional culture for the diagnosis of pertussis. They also demonstrate a high specificity of PCR when serology with or without other confirmative criteria (culture and household contact) is used as the reference. Analysis of semiquantitative PCR results revealed that subjects with a 3+ PCR more frequently experienced typical illness compared with patients with 1+ or 2+ PCR. Although specific serologic study remains a necessity in pertussis research its modification for diagnosis in the clinical setting results in low sensitivity and specificity. Therefore, because PCR is more sensitive than culture and is easy to perform, it is a useful addition in the clinical setting. PMID- 10699134 TI - Sports injuries: An important cause of morbidity in urban youth. District of Columbia Child/Adolescent Injury Research Network. AB - INTRODUCTION: Sports injuries account for substantial morbidity and medical cost. To direct intervention, a population-based study of the causes and types of sports injuries was undertaken. METHOD: An injury surveillance system was established at all trauma center hospitals that treat residents 10 to 19 years old in the District of Columbia and the Chief Medical Examiner's Office. Medical record abstractions were completed for those seen in an emergency department, admitted to the hospital, or who died from injury June 1996 through June 1998. FINDINGS: Seventeen percent (n = 2563) of all injuries occurred while participating in 1 of 6 sports (baseball/softball, basketball, biking, football, skating, and soccer) resulting in an event-based injury rate of 25.0 per 1000 adolescents or 25.0/1000 population year. Rates were higher in males for all sports. The most common mechanisms were falls (E880-888) and being struck by or against objects (E916-918). Hospitalization was required in 2% of visits and there were no deaths. Of those requiring hospitalization, 51% involved other persons, 12% were equipment-related, and 8% involved poor field/surface conditions. Of all baseball injuries, 55% involved ball or bat impact often of the head. Basketball injuries included several injuries from striking against the basketball pole or rim or being struck by a falling pole or backboard. Biking injuries requiring admission included 2 straddle injuries onto the bike center bar and collision with motor vehicles. Of all football injuries, 48 (7%) involved being struck by an opponent's helmet and 63 (9%) involved inappropriate field conditions including falls on or against concrete, glass, or fixed objects. In soccer there were 4 goal post injuries and a large proportion of intracranial injuries. There were 51 probable or clear assaults during sports and an additional 30 to 41 injuries from baseball bat assaults. CONCLUSIONS: Many sports including noncontact sports involved injuries of the head suggesting the need for improved head protection. Injuries involving collisions with others and assaults point to the need for supervision and enforcement of safety rules. The 16% of sports injury visits and 20% of hospitalizations related to equipment and environmental factors suggest that at least this proportion of injury may be amenable to preventive strategies. Design change may be warranted for prevention of equipment-related injuries. The many injuries involving inappropriate sports settings suggest the need for and use of available and safe locations for sports. PMID- 10699135 TI - Assessing the impact of pediatric-based development services on infants, families, and clinicians: challenges to evaluating the Health Steps Program. AB - BACKGROUND: Begun in 1996, the Healthy Steps for Young Children Program (HS) is a new model of pediatric practice that incorporates child development specialists and enhanced developmental services for families of young children. HS is for all families, not just those at high-risk. It is expected to strengthen parents' knowledge, attitudes, and behaviors in ways that promote child health and development, and in turn, to lead to improved child outcomes, such as improved language development, increased utilization of well child care, and decreased problem behaviors, hospitalizations, and injuries. The HS evaluation is designed to assess whether HS is successful in achieving the desired outcomes, measure the program's costs, and determine the relation of the program's costs to its outcomes. OBJECTIVE: This article is the first report of the HS evaluation. It describes the evaluation design and characteristics of the HS sites and sample for the evaluation. METHODS: The evaluation is following a cohort of children from birth to age 3 at 15 evaluation sites across the country. The sites represent a range of organizational practice settings that include group practices, hospital-based clinics, and health maintenance organization pediatric clinics. The evaluation design relies on 2 comparison strategies. At 6 randomization design sites, 400 children were randomized to the intervention or control group. At 9 quasi-experimental design sites, a comparison location with a similar organizational setting and patient profile has been selected and up to 200 children are being followed at each of these sites. At each site, 2 developmental specialists (or their full-time equivalents) work as a team with 4 to 8 pediatricians and pediatric nurse practitioners. The specialist conducts office visits (jointly or sequentially with the pediatric clinician) and home visits, assesses children's developmental progress, provides referrals and follow up to resources in the community, organizes and conducts parent discussion groups, coordinates early reading activities, and maintains a telephone information line for questions about child development and behavior. The evaluation relies on many data sources including self-administered provider surveys, key informant interviews, forms completed by parents at office visits, telephone interviews with parents, medical record reviews, data from each site on program costs and health services use, and an ongoing log of family contacts maintained by each developmental specialist. Analyses for this article are based on enrollment data for the Healthy Steps sample and national data on 1997 US live births. The chi2 goodness-of-fit test was used to evaluate whether the distribution of selected demographic variables, insurance, and infant's birth weight for the Healthy Steps sample was similar to the distributions for US births in 1997. In addition, comparisons were made between intervention and comparison families at the randomization and quasi-experimental evaluation sites. The chi2 test of independence was used to evaluate differences in variables across groups. RESULTS: Throughout a 26-month period, 5565 children enrolled in the evaluation, 2963 (53.2%) children in the intervention group and 2602 (46.8%) in the comparison group. More than 10% of mothers in the Healthy Steps sample are teenagers; 18% have 11 years of education or less; 27% have completed college; 18% are black or African-American; slightly >20% are of Hispanic origin; 36% are single; and close to one-third used Medicaid for their prenatal care. Approximately 7% of infants were low birth weight. When compared with national birth data for the United States as a whole, the Healthy Steps sample seems similarly diverse. However, with the exception of maternal age, the distribution of variables was significantly different from the distribution for US births. There are no differences between intervention and comparison families at randomization sites on any of PMID- 10699136 TI - Mercury intoxication and arterial hypertension: report of two patients and review of the literature. AB - Two children in the same household with symptomatic arterial hypertension simulating pheochromocytoma were found to be intoxicated with elemental mercury. The first child was a 4-year-old boy who presented with new-onset seizures, rash, and painful extremities, who was found to have a blood pressure of 171/123 mm Hg. An extensive investigation ensued. Elevated catecholamines were demonstrated in plasma and urine; studies did not confirm pheochromocytoma. Mercury levels were elevated. These findings prompted an evaluation of the family. A foster sister had similar findings of rash and hypertension. Both had been exposed to elemental mercury in the home. The family was temporarily relocated and chelation therapy was started. A Medline search for mercury intoxication with hypertension found 6 reports of patients ranging from 11 months to 17 years old. All patients showed symptoms of acrodynia. Because of the clinical presentation and the finding of elevated catecholamines, most of the patients were first studied for possible pheochromocytoma. Subsequently, elevated levels of mercury were found. Three children had contact with elemental mercury from a broken thermometer, 2 had played with metallic mercury and 1 had poorly protected occupational exposure. All responded to chelation therapy. Severe systemic arterial hypertension in infants and children is usually secondary to an underlying disease process. The most frequent causes of hypertension in this group include renal parenchymal disease, obstructive uropathy, and chronic pyelonephritis associated with reflux and renal artery stenosis. Less frequent causes include adrenal tumors, pheochromocytomas, neurofibromas, and a number of familial forms of hypertension. Other causes include therapeutic and recreational drugs, notably sympathomimetics and cocaine, and rarely, heavy metals. In children with severe hypertension and elevated catecholamines, the physician should consider mercury intoxication as well as pheochromocytoma. The health hazards of heavy metals need to be reinforced to the medical profession and the general public. PMID- 10699137 TI - Lymphocytic choriomeningitis virus: reemerging central nervous system pathogen. AB - Lymphocytic choriomeningitis virus (LCMV), a human zoonosis caused by a rodent borne arenavirus, has been associated with both postnatal and intrauterine human disease. Infection in man is acquired after inhalation, ingestion, or direct contact with virus found in the urine, feces, and saliva of infected mice, hamsters, and guinea pigs. Congenital LCMV infection is a significant, often unrecognized cause of chorioretinitis, hydrocephalus, microcephaly or macrocephaly, and mental retardation. Acquired LCMV infection, asymptomatic in approximately one third of individuals, is productive of central nervous system manifestations in one half of the remaining cases. Aseptic meningitis or meningoencephalitis are the predominant syndromes, although transverse myelitis, a Guillain-Barre-type syndrome, as well as transient and permanent acquired hydrocephalus have also been reported. Fatalities are rare. We report a patient with meningoencephalitis attributable to LCMV and discuss the spectrum of central nervous system disease, newer diagnostic modalities, and preventive strategies. lymphocytic choriomeningitis virus, aseptic meningitis, meningoencephalitis, zoonosis, hydrocephalus, arenavirus. PMID- 10699138 TI - The highly protective effect of newborn circumcision against invasive penile cancer. AB - OBJECTIVE: We determined the relation between newborn circumcision and both invasive penile cancer (IPC) and carcinoma in situ (CIS) among adult male members of a large health maintenance organization. SUBJECTS AND METHODS: Circumcision status was ascertained by a combination of pathology reports, medical record review, and questionnaires for 213 adult male members of a large prepaid health plan who were diagnosed with IPC or CIS. RESULTS: Of 89 men with IPC whose circumcision status was known, 2 (2.3%) had been circumcised as newborns, and 87 were not circumcised. Of 118 men with CIS whose circumcision status was known, 16 (15.7%) had been circumcised as newborns. CONCLUSIONS: Our results confirm the highly protective effect of newborn circumcision against IPC and the less protective effect against CIS. PMID- 10699139 TI - Mortality and time to death in very low birth weight infants: California, 1987 and 1993. AB - BACKGROUND: Recent advances in perinatal technology have dramatically increased the survival of very low birth weight (VLBW) infants (<1500 g). The possibility that these advances may also prolong the time to death and increase pain and suffering has been of concern, but there have been no population-based evaluations of this issue. METHODS: Infant, neonatal, and postneonatal mortality rates and time to death for infants 500 to 749 g, 750 to 999 g, 1000 to 1499 g, and all VLBW infants born during 1987 were compared with those outcomes for infants born in 1993 using statewide California linked birth/death cohort files. To assess the effects of improved survival and changes in time until death, we calculated the total days of life preceding an infant death per 1000 live born infants (TDD). RESULTS: VLBW infants comprised.96% of California's live births in 1987 and.92% of those in 1993. Between 1987 and 1993, VLBW infant mortality rate decreased 28.4% (from 290.7 to 208.3 per 1000 live born VLBW infants), VLBW neonatal mortality rate decreased 30. 3% (from 244.5 to 170.4), and VLBW postneonatal mortality rate decreased 25.3% (from 61.2 to 45.7 per 1000 VLBW alive at 28 days; P <.05 for each rate). Infant mortality rates decreased by 18.8% (718. 1 to 583.0 per 1000) for infants 500 to 749 g, 43.3% (375.1 to 202. 6) for infants 750 to 999 g, and 40.1% (127.9 to 76.7) for infants 1000 to 1449 g (P <.05 for each group). Neonatal mortality and postneonatal mortality rates also decreased in all 3 VLBW subgroups. These reductions in mortality rates were not accompanied by a significant difference in the distribution of times to death or a significant increase in the average time to death for all VLBW infants (22.0 vs 23.6 days) or for those with birth weights of 500 to 749 g (12.7 vs 71.5 days). Reduced mortality in larger infants was accompanied by an increase in the average time to death, from 24. 3 to 32.5 days in infants 750 to 999 g and from 32.3 to 47.0 days in infants 1000 to 1449 g. TDD decreased from 6410 to 4908 days for all VLBW infants. TDD was also reduced 26.4% (2401 days), 24.3% (2115 days), and 22.5% (1043 days) for the 3 VLBW birth weight groups. CONCLUSIONS: Both mortality rate and timing of death are important when assessing the impact of advances in perinatal technology. Although the average time to death was significantly increased in VLBW infants weighing >750 g, between 1987 and 1993, advances in perinatal technology dramatically decreased VLBW mortality. In the State of California in 1993, this resulted in 452 fewer VLBW deaths and 8233 fewer days preceding a VLBW death than expected. PMID- 10699140 TI - Clinical safety of iron-fortified formulas. AB - BACKGROUND: Iron-fortified formulas are recommended throughout infancy and are frequently used beyond, yet safety aspects have been inadequately studied. Iron could theoretically increase pro-oxidant stress, with potential adverse effects, including infection risk, and some clinicians suspect that iron-fortified formulas induce gastrointestinal disturbance. OBJECTIVE: A planned component of a large intervention trial has been to test the hypothesis that infants receiving iron-fortified formula do not have a higher incidence of infections (primary outcome) or gastrointestinal problems (secondary outcome) than infants on low iron-formulas or cow's milk. Methods. Children (n = 493) 9 months old receiving cow's milk were recruited in 3 UK centers and randomized to: 1) cow's milk as before, 2) formula containing.9 mg/L of iron, or 3) an otherwise identical formula but containing 12 mg/L of iron. Children were followed at 3 monthly intervals and the episodes of infections, diarrhea and constipation, and general morbidity to 18 months old were recorded. Hematologic indices of iron status were determined at 18 months old. RESULTS: Serum ferritin concentrations were increased in infants receiving iron-fortified formula but there were no intergroup differences in incidence of infection, gastrointestinal problems, or in general morbidity or weight gain. CONCLUSIONS: We were unable to identify adverse health effects in older infants and toddlers consuming a high iron containing formula (12 mg/L) even when used in populations with a low incidence of iron deficiency. PMID- 10699141 TI - A syndrome involving intrauterine growth retardation, microcephaly, cerebellar hypoplasia, B lymphocyte deficiency, and progressive pancytopenia. AB - We report a new complex syndrome involving profound failure to thrive with severe intrauterine growth retardation, cerebellar abnormalities, microcephaly, a complete lack of B lymphocyte development, and secondary, progressive marrow aplasia. B cell differentiation was found to be blocked at the pro-B cell stage. Although not strictly proven, a genetic origin is likely, according to similar cases reported in the literature. Three candidate genes, PAX5, encoding B cell specific activator protein, a factor involved in B cell lineage commitment, stromal cell-derived factor 1, and CXCR4, encoding a chemokine and its receptor, respectively, were thought to be responsible for this disease, given the similarity between the phenotype of the corresponding knock-out mice and the clinical features of the patient. However, the genomic DNA sequences of these 3 genes were normal, and normal amounts of stromal cell-derived factor 1 and CXCR4 were present. These data strongly suggest that another molecule is involved in early B cell differentiation, hematopoiesis, and cerebellar development in humans. PMID- 10699142 TI - Adverse effects of fetal cocaine exposure on neonatal auditory information processing. AB - BACKGROUND: Studies with animals have shown that in utero exposure to cocaine interferes with fetal brain development by disrupting the processes of neuronal proliferation, differentiation, and migration, often leading to subsequent neurobehavioral deficits. However, studies with humans have produced inconsistent findings. Although neurobehavioral abnormalities have been observed among cocaine exposed infants in several studies and in some cases dose-response effects have been found, the specific neurobehaviors affected vary from one study to the next. Researchers studying the effects of fetal cocaine-exposure are faced with many difficult challenges. For example, women who use cocaine typically use other substances in addition to cocaine, many of the methods available for identifying cocaine-exposed neonates are not reliable, and the available methods for assessing cocaine-exposed newborns may not be sufficiently sensitive to detect the subtle effects of cocaine on the developing central nervous system. Despite these difficulties, there is a growing body of research that suggests that fetal cocaine exposure is associated with subsequent language deficits among children exposed in utero. However, it is virtually impossible to disentangle the effects of the impoverished environments in which these children are often raised from the effect, if any, of fetal cocaine exposure. To determine the effects of fetal cocaine exposure independent of postnatal environmental effects, cocaine-exposed neonates would ideally be tested within the first few weeks of birth, and to identify early risks for subsequent language delay, well-researched auditory information processing measures could be used. OBJECTIVE: The purpose of the present study was to assess the effects of fetal cocaine exposure on neonatal auditory information processing ability. To overcome limitations of some previous studies on the neuroteratogenic effects of cocaine, such as unreliable subject identification techniques, inadequate control over confounding variables, and questionable measures of central nervous system integrity, a valid measure of auditory information processing was used in a rigorous, case-control design. METHOD: Newborn information processing was assessed using habituation and recovery of head-turning toward an auditory stimulus across the 3 phases of the procedure: familiarization, novelty, and dishabituation. During the familiarization phase, the infant orients and habituates to a repeated word; during the novelty phase, the infant recovers head-turning to a novel word and subsequently habituates to this word; and during the dishabituation phase the infant displays renewed head-turning to the return of the original stimulus. Testing takes approximately 20 minutes. This procedure has been shown previously to discriminate among infants at high-, moderate-, and low-risk for subsequent developmental delay. Twenty-five cocaine-exposed and 25 nonexposed control neonates, identified by meconium analysis, urine analysis, and/or maternal self report, were tested on the auditory information processing procedure. The majority of infants were tested within the first few days of birth. Cocaine exposed and control neonates were matched on birth weight, gestational age, Apgar scores, age at testing, and socioeconomic status as reflected by household income. Mothers were matched on age, weight gain, cigarette smoking, and alcohol consumption. RESULTS: Fetal cocaine exposure was associated with impaired auditory information processing. Both cocaine-exposed and nonexposed control neonates oriented to the familiarization stimulus, but cocaine-exposed neonates displayed impaired habituation. Moreover, cocaine-exposed neonates did not recover or habituate to the novel stimulus or dishabituate to the return of the familiarization stimulus. (ABSTRACT TRUNCATED) PMID- 10699143 TI - Infants with a thumb-in-fist posture. AB - OBJECTIVE: In early infancy the infant's thumb is not infrequently enclosed within the palm, ie, thumb-in-fist (TIF). This posture has received scant attention in the neurodevelopmental literature. Its prevalence, resolution, and clinical associations were investigated in this study. METHODOLOGY: Two hundred sequentially born, apparently healthy full-term newborn infants comprised the cohort. The whole study group was followed up until the disappearance of the TIF occurred. In the first 150 of the cohort, additional data on development and the neurological status were obtained at 12 months of life. RESULTS: In 125 infants (62.5%) of the total cohort, a TIF was noted. The mean age of disappearance was 1.5 months, and no TIF persisted after 7 months old. No relationship was noted between the TIF resolution and abnormal neurological signs or gross or fine motor development. The only association noted between age of resolution of the TIF and the neurodevelopmental status was a delay in language attainment at the 12-month screening. CONCLUSIONS: The TIF posture in infancy was noted in 65% of our cohort, and it had resolved in all infants by 7 months old. Therefore, a TIF posture after this age should alert the clinician to the possibility of possible neurological dysfunction. An unanticipated association of a delay in the 12-month language milestone attainment was noted in those infants with later resolution of the TIF posture. No data on language development in this group were obtained after 12 months old; therefore, the clinical significance of this finding is not yet elucidated. PMID- 10699144 TI - Prolonged recovery and delayed side effects of sedation for diagnostic imaging studies in children. AB - OBJECTIVE: Although sedation-related adverse events in children in the hospital setting have been extensively reported, limited data are available regarding adverse events after discharge home. Despite nationally recommended discharge criteria, in busy outpatient settings, children may be sent home into the care of their parents after a brief recovery from sedation, placing them at risk for adverse events in an unmonitored setting. Previous studies have not addressed issues such as requirement for escalation of care after discharge (ie, emergency department visits or hospitalization), or parental satisfaction with their child's sedation experience. This study was undertaken to evaluate the recovery and delayed adverse events after discharge of children who received sedation for magnetic resonance imaging or computed tomography. METHODS: With approval from the institutional review board and written informed consent from a parent, children (<18 years old) sedated for magnetic resonance imaging or computerized tomography were studied. Sedative drugs were ordered at the discretion of the radiologist responsible for the procedure in accordance with institutional sedation guidelines and in consideration of the child's health status. Pediatric nurses in the diagnostic areas administered the sedative agent(s) and monitored children according to preestablished institutional guidelines. Demographics, sedative(s) administered, and adverse events including hypoxemia (decrease in SpO(2) by >/=10% of baseline) and sedation events such as inadequate, failed, or excessive sedation, were documented on the institutional quality assurance tool. Children were discharged from the hospital when they met the following preestablished discharge criteria: return to baseline vital signs, level of consciousness close to baseline, and the ability to maintain a patent airway. The following day, parents were telephoned and questioned regarding the child's alertness, side effects, and whether medical follow-up had been sought. Parents also rated their overall satisfaction with the sedation experience. RESULTS: Three hundred seventy six children comprised the sample. Eighty nine percent of children received chloral hydrate (CH; 64 +/- 13 mg/kg), and 11% midazolam (.15 +/-.13 mg/kg) as the primary sedative. There was an 8% incidence of failed sedation, and a 1.6% incidence of hypoxemia during the procedure. Three children required prolonged monitoring in the postanesthesia care unit before discharge; 1 child attributable to an allergic reaction, a second attributable to wheezing and oxygen desaturation, and the third attributable to prolonged sedation from CH and midazolam. These children were discharged home from the postanesthesia care unit without additional sequelae. Side effects after discharge included: motor imbalance (31%), gastrointestinal effects (23%), agitation (19%), and restlessness (14%). Agitation and restlessness lasted greater than 6 hours in more than one third of children who experienced these effects. CH was more commonly associated with imbalance compared with midazolam, and restlessness and prolonged imbalance were associated with younger age. Medical advice was sought after discharge for 15 (4%) children, 3 of whom required a visit to the emergency department for excessive or prolonged sedation. Each of these children had received CH as a sole sedative in recommended doses (61-77 mg/kg). In 1 of these cases, the procedure had been aborted because of inadequate sedation in the hospital, yet the child became difficult to arouse at home. Only 48% of children returned to baseline activity and behavior within 8 hours of the procedure; however, 89% were back to baseline status within 24 hours. Notably, 5% of all children did not return to baseline activity until the second day after the procedure. Although not statistically significant, infants <12 months old experienced delayed recovery (ie, >/=24 hours) more frequently compared with older c PMID- 10699145 TI - Evaluation of children's health insurance: from New York State's CHild Health Plus to SCHIP. AB - BACKGROUND: The legislation and funding of the State Children's Health Insurance Program (SCHIP) in 1997 resulted in the largest public investment in child health care in 30 years. The program was designed to provide health insurance for the estimated 11 million uninsured children in the United States. In 1991 New York State implemented a state-funded program-Child Health Plus (CHPlus)-intended to provide health insurance for uninsured children who were ineligible for Medicaid. The program became one of the prototypes for SCHIP: This study was designed to measure the association between CHPlus and access to care, utilization of care, quality of care, and health care costs to understand the potential impact of one type of prototype SCHIP program. METHODS: The study took place in the 6-county region of upstate New York around and including the city of Rochester. A before and-during design was used to compare children's health care for the year before they enrolled in CHPlus versus the first year during enrollment in CHPlus. The study included 1828 children (ages 0-6.99 years at enrollment) who enrolled between November 1, 1991 and August 1, 1993. A substudy involved 187 children 2 to 12.99 years old who had asthma. Data collection involved: 1) interviews of parents to obtain information about demographics, sources of health care, experience and satisfaction with CHPlus, and perceived impact of CHPlus; 2) medical chart reviews at all primary care offices, emergency departments, and health department clinics in the 6-county region to measure utilization of health services; 3) claims analysis to assess costs of care during CHPlus and to impute costs before CHPlus; and 4) analyses of existing datasets including the Current Population Survey, National Health Interview Survey, and statewide hospitalization datasets to anchor the study in relation to the statewide CHPlus population and to assess secular trends in child health care. Logistic regression and Poisson regression were used to compare the means of dependent measures with and without CHPlus coverage, while controlling for age, prior insurance type, and gap in insurance coverage before CHPlus. RESULTS: ENROLLMENT: Only one third of CHPlus-eligible children throughout New York State had enrolled in the program by 1993. Lower enrollment rates occurred among Hispanic and black children than among white children, and among children from lowest income levels. PROFILE OF CHPlus ENROLLEES: Most enrollees were either previously uninsured, had Medicaid but were no longer eligible, or had parents who either lost a job and related private insurance coverage or could no longer afford commercial or private insurance. Most families heard about CHPlus from a friend, physician, or insurer. Television, radio, and newspaper advertisements were not major sources of information. Nearly all families had at least 1 employed parent. Two thirds of the children resided in 2-parent households. Parents reported that most children were in excellent or good health and only a few were in poor health. The enrolled population was thus a relatively low-risk, generally healthy group of children in low-income, working families. ACCESS AND UTILIZATION OF HEALTH CARE: Utilization of primary care increased dramatically after enrollment in CHPlus, compared with before CHPlus. Visits to primary care medical homes for preventive, acute, and chronic care increased markedly. Visits to medical homes also increased for children with asthma. There was, however, no significant association between enrollment in CHPlus and changes in utilization of emergency departments, specialty services, or inpatient care. QUALITY OF CARE: CHPlus was associated with improvements in many measures involving quality of primary care, including preventive visits, immunization rates, use of the medical home for health care, compliance with preventive guidelines, and parent-reported health status of the child. (ABSTRACT TRUNCATED) PMID- 10699146 TI - Evolution of a children's health insurance program: lessons from New York State's Child Health Plus. AB - The State Children's Health Insurance Program (SCHIP) was passed by Congress in 1997. It provides almost $40 billion in federal block grant funding through the year 2007 for states to expand health insurance for children. States have the option of expanding their Medicaid programs, creating separate insurance programs, or developing combination plans using both Medicaid and the private insurance option. New York State's child health insurance plan, known by its marketing name Child Health Plus, was created by the New York Legislature in 1990. New York's program, along with similar ones from several other states, served as models for the federal legislation, especially for state health insurance plans offered through private insurers. New York's program provides useful data for successful implementation of SCHIP. PMID- 10699147 TI - Evaluation of New York State's Child Health Plus: methods. AB - BACKGROUND: The State Children's Health Insurance Program (SCHIP) is the largest public investment in child health care in 30 years, targeting 11 million uninsured children, yet little is known about the impact of health insurance on uninsured children. In 1991 New York State implemented Child Health Plus (CHPlus), a health insurance program that was a prototype for SCHIP. A study was designed to measure the association between CHPlus and access to care, utilization of services, and quality of care. METHODS: The setting was a 6-county region in upstate New York (population 1 million) around and including the city of Rochester. A before-and-during design was used to compare children's health care for the year before they enrolled in CHPlus versus the first year during CHPlus, for 1828 children (ages 0-6.99 years at enrollment) who enrolled between November 1, 1991 and August 1, 1993. An additional study involved 187 children 2 to 12.99 years old who had asthma. Parents were interviewed to assess demographic characteristics, sources of health care, experience with CHPlus, and impact of CHPlus on their children's quality of care and health status. Medical charts were reviewed to measure utilization and quality of care, for 1730 children 0 to 6.99 years and 169 children who had asthma. Charts were reviewed at all primary care offices and at the 12 emergency departments and 6 public health department clinics in the region. CHPlus claims files were analyzed to determine costs during CHPlus and to impute costs before CHPlus from utilization data. ANALYSES: Logistic regression and Poisson regression were used to compare the means of dependent measures with and without CHPlus coverage, while controlling for age, prior insurance type, and gap in insurance coverage before CHPlus. CONCLUSIONS: This study developed and implemented methods to evaluate the association between enrollment in a health insurance program and children's health care. These methods may also be useful for evaluations of SCHIP. PMID- 10699148 TI - A profile of the population enrolled in New York State's Child Health Plus. AB - BACKGROUND: The recently enacted State Children's Health Insurance Program (SCHIP), designed to provide affordable health insurance for uninsured children, was modeled in part on New York State's Child Health Plus (CHPlus), which was implemented in 1991. All SCHIP programs involve voluntary enrollment of eligible children. Little is known about characteristics of children who enroll in these programs. OBJECTIVES: To provide a profile of children enrolled in CHPlus between 1993 and 1994 in the 6-county upstate New York study area, and to estimate the participation rate in CHPlus. Methods. A parent interview was conducted to obtain information about children, 0 to 6.9 years old, who enrolled in CHPlus in the study area. Two school-based surveys and the Current Population Survey were used to estimate health insurance coverage. Enrollment data from New York State's Department of Health, together with estimates of the uninsured, were used to estimate participation rates in CHPlus. RESULTS: Most children enrolled in CHPlus in the study area were white. Although 17% of all children in the study area who were <13 years old and living in families with incomes below 160% of the federal poverty level were black, only 9% of CHPlus-enrolled children were black. Twenty one percent of enrolled children were uninsured during the entire year before enrollment and 61% of children had a gap in coverage lasting >1 month. Children were generally healthy; only 4% had fair or poor health. Eighty-eight percent of parents of enrolled children had completed high school or a higher level of education. Parents reported that loss of a job was the main reason for loss of prior health insurance for their child. Most families learned about CHPlus from a friend (30%) or from their doctor (26%). The uninsured rate among children in the study area was approximately 4.1%. By 1993, the participation rate in CHPlus was about 36%. CONCLUSION: Blacks were underrepresented in CHPlus. Because the underlying uninsured rate was relatively low and parental education and family income were relatively high, the effects of CHPlus observed in this evaluation may be conservative in comparison to the potential effects of CHPlus for other populations of children. Participation rates during the early years of the program were modest. PMID- 10699149 TI - Evaluation of New York State's Child Health Plus: access, utilization, quality of health care, and health status. AB - BACKGROUND: The recently enacted State Children's Health Insurance Program (SCHIP) is modeled after New York State's Child Health Plus (CHPlus) program. Since 1991, CHPlus has provided health insurance to children 0 to 13 years old whose annual family income was below 222% of the federal poverty level and who were ineligible for Medicaid or did not have equivalent health insurance coverage. CHPlus covered the costs for ambulatory, emergency, and specialty care, and prescriptions, but not inpatient services. OBJECTIVES: To assess the change associated with CHPlus regarding 1) access to health care; 2) utilization of ambulatory, inpatient, and emergency services; 3) quality of health care; and 4) health status. SETTING: Six western New York State counties (including the city of Rochester). SUBJECTS: Children (0-6.99 years old) enrolled for at least 9 consecutive months in CHPlus. METHODS: The design was a before-and-after study, comparing individual-level outcomes for the 12 months immediately before CHPlus enrollment and the 12 months immediately after enrollment in CHPlus. Parent telephone interviews and medical chart reviews conducted 12 months after enrollment to gather information. Subjects' primary care charts were located by using interview information; emergency department (ED) charts were identified by searching patient records at all 12 EDs serving children in the study; and health department charts were identified by searching patient records at the 6 county health department clinics. Logistic regression and Poisson regression were used to compare the means of dependent measures with and without CHPlus coverage, while controlling for age, prior insurance type, and gap in insurance coverage before CHPlus. RESULTS: Complete data were obtained for 1730 children. Coverage by CHPlus was associated with a significant improvement in access to care as measured by the proportion of children reported as having a usual source of care (preventive care: +1.9% improvement during CHPlus and sick care: +2. 7%). CHPlus was associated, among children 1 to 5 years old, with a significant increase in utilization of preventive care (+.23 visits/child/year) and sick care (+.91 visits/child/year) but no measurable change in utilization of specialty, emergency, or inpatient care. CHPlus was also associated, among children 1 to 5 years old, with significantly higher immunization rates (up-to-date for immunizations: 76% vs 71%), and screening rates for anemia (+11% increased proportion screened/year), lead (+9%), vision (+11%), and hearing (+7%). For 25% of the children, a parent reported that their child's health was improved as a result of having CHPlus. CONCLUSION: After enrollment in CHPlus, access to and utilization of primary care increased, continuity of care improved, and many quality of care measures were improved while utilization of emergency and specialty care did not change. Many parents reported improved health status of their child as a result of enrollment in CHPlus. Implication. This evaluation suggests that SCHIP programs are likely to improve access to, quality of, and participation in primary care significantly and may not be associated with significant changes in specialty or emergency care. PMID- 10699150 TI - Evaluation of New York State's Child Health Plus: children who have asthma. AB - BACKGROUND: Little is known about the impact of providing health insurance to uninsured children who have asthma or other chronic diseases. OBJECTIVES: To evaluate the association between health insurance and the utilization of health care and the quality of care among children who have asthma. DESIGN: Before-and during study of children for a 1-year period before and a 1-year period immediately after enrollment in a state-funded health insurance plan. INTERVENTION: In 1991 New York State implemented Child Health Plus (CHPlus), a health insurance program providing ambulatory and ED (ED), but not hospitalization coverage for children 0 to 12.99 years old whose family incomes were below 222% of the federal poverty level and who were not enrolled in Medicaid. SUBJECTS: A total of 187 children (2-12.99 years old) who had asthma and enrolled in CHPlus between November 1, 1991 and August 1, 1993. MAIN OUTCOME MEASURES: Rates of primary care visits (preventive, acute, asthma-specific), ED visits, hospitalizations, number of specialists seen, and quality of care measures (parent reports of the effect of CHPlus on quality of asthma care, and rates of recommended asthma therapies). The effect of CHPlus was assessed by comparing outcome measures for each child for the year before versus the year after CHPlus enrollment, controlling for age, insurance coverage before CHPlus, and asthma severity. DATA ASCERTAINMENT: Parent telephone interviews and medical chart reviews at primary care offices, EDs, and public health clinics. MAIN RESULTS: Visit rates to primary care providers were significantly higher during CHPlus compared with before CHPlus for chronic illness care (.995 visits before CHPlus vs 1.34 visits per year during CHPlus), follow-up visits (.86 visits vs 1.32 visits per year), total visits (5.69 visits vs 7.11 visits per year), and for acute asthma exacerbations (.61 visits vs 0.84 visits per year). There were no significant associations between CHPlus coverage and ED visits or hospitalizations, although specialty utilization increased (30% vs 40%; P =.02). According to parents, CHPlus reduced asthma severity for 55% of children (no change in severity for 44% and worsening severity for 1%). Similarly, CHPlus was reported to have improved overall health status for 45% of children (no change in 53% and worse in 1%), primarily attributable to coverage for office visits and asthma medications. CHPlus was associated with more asthma tune-up visits (48% before CHPlus vs 63% during CHPlus). There was no statistically significant effect of CHPlus on several other quality of care measures such as follow-up after acute exacerbations, receipt of influenza vaccination, or use of bronchodilators or antiinflammatory medications. CONCLUSIONS: Health insurance for uninsured children who have asthma helped overcome financial barriers that prevented children from receiving care for acute asthma exacerbations and for chronic asthma care. Health insurance was associated with increased utilization of primary care for asthma and improved parent perception of quality of care and asthma severity, but not with some quality indicators. Although more intensive interventions beyond health insurance are needed to optimize quality of asthma care, health insurance coverage substantially improves the health care for children who have asthma. PMID- 10699151 TI - Evaluating Child Health Plus in upstate New York: how much does providing health insurance to uninsured children increase health care costs? AB - BACKGROUND: In response to the increase in the number of American children without health insurance, new federal and state programs have been established to expand health insurance coverage for children. However, the presence of insurance reduces the price of care for families participating in these programs and stimulates the use of medical services, which leads to an increase in health care costs. In this article, we identified the additional expenditures associated with the provision of health insurance to previously uninsured children. METHODS: We estimated the expenditures on additional services using data from a study of children living in the Rochester, New York, area who were enrolled in the New York State Child Health Plus (CHPlus) program. CHPlus was designed specifically for low-income children without health insurance who were not eligible for Medicaid. The study sample consisted of 1910 children under the age of 6 who were initially enrolled in CHPlus between November 1, 1991 and August 1, 1993 and who had been enrolled for at least 9 continuous months. We used medical chart reviews to determine the level of primary care utilization, parent interviews for demographic information, as well as specialty care utilization, and we used claims data submitted to CHPlus for the year after enrollment to calculate health care expenditures. Using this information, we estimated a multivariate regression model to compute the average change in expenditures associated with a unit of utilization for a cross-section of service types while controlling for other factors that independently influenced total outpatient expenditures. RESULTS: Expenditures for outpatient services were closely related to primary care utilization-more utilization tended to increase expenditures. Age and the presence of a chronic condition both affected expenditures. Children with chronic conditions and infants tended to have more visits, but these visits were, on average, less expensive. Applying the average change in expenditures to the change in utilization that resulted from the presence of insurance, we estimated that the total increase in expenditures associated with CHPlus was $71.85 per child in the year after enrollment, or a 23% increase in expenditures. The cost increase was almost entirely associated with the provision of primary care. Almost three-quarters of the increase in outpatient expenditures was associated with increased acute and well-child care visits. CONCLUSIONS: CHPlus was associated with a modest increase in expenditures, mostly from additional outpatient utilization. Because the additional primary care provided to young children often has substantial long-term benefits, the relatively modest expenditure increases associated with the provision of insurance may be viewed as an investment in the future. PMID- 10699152 TI - Genetically engineered insulin analogs: diabetes in the new millenium. AB - Tight glucose control is essential to minimize complications in diabetic patients. However, the pharmacokinetic characteristics of the currently available rapid-, intermediate-, and long-acting preparations of human insulin make it almost impossible to achieve sustained normoglycemia. Until recently, improvements in insulin formulations were seriously limited as advances were only achieved in insulin purity, species, and characteristics of the retarding agent. The availability of molecular genetic techniques opened new windows to create insulin analogs by changing the structure of the native protein and to improve the therapeutic properties. The first clinically available insulin analog, Lispro, confirmed the hopes by showing that improved glycemic control can be achieved without an increase in hypoglycemic events. This requires, however, optimal basal insulin replacement, either by multiple daily injections of neutral protein Hagedorn (NPH) insulin or by insulin pump. Evidence suggests that short acting insulin analogs would be better matched by a true basal insulin than by the erratically absorbed and rather short-acting NPH insulin. Therefore, future availability of long-acting analogs raises the hope to realize the true potential benefits of the currently available short-acting analog, Lispro, and of those still awaiting approval. The introduction of new short-acting and the first truly long-acting analogs, the development of analogs with increased stability, less variability and perhaps selective action will help to develop more individualized treatment strategies targeted to specific patient characteristics and to achieve further improvements in glycemic control. PMID- 10699153 TI - Molecular and cellular mechanisms of angiotensin II-mediated cardiovascular and renal diseases. AB - A growing body of evidence supports the notion that angiotensin II (Ang II), the central product of the renin-angiotensin system, may play a central role not only in the etiology of hypertension but also in the pathophysiology of cardiovascular and renal diseases in humans. In this review, we focus on the role of Ang II in cardiovascular and renal diseases at the molecular and cellular levels and discuss up-to-date evidence concerning the in vitro and in vivo actions of Ang II and the pharmacological effects of angiotensin receptor antagonists in comparison with angiotensin-converting enzyme inhibitors. Ang II, via AT(1) receptor, directly causes cellular phenotypic changes and cell growth, regulates the gene expression of various bioactive substances (vasoactive hormones, growth factors, extracellular matrix components, cytokines, etc.), and activates multiple intracellular signaling cascades (mitogen-activated protein kinase cascades, tyrosine kinases, various transcription factors, etc.) in cardiac myocytes and fibroblasts, vascular endothelial and smooth muscle cells, and renal mesangial cells. These actions are supposed to participate in the pathophysiology of cardiac hypertrophy and remodeling, heart failure, vascular thickening, atherosclerosis, and glomerulosclerosis. Furthermore, in vivo recent evidence suggest that the activation of mitogen-activated protein kinases and activator protein-1 by Ang II may play the key role in cardiovascular and renal diseases. However, there are still unresolved questions and controversies on the mechanism of Ang II-mediated cardiovascular and renal diseases. PMID- 10699154 TI - Transgenic approach to the study of body weight regulation. AB - Energy homeostasis is accomplished through a highly integrated and redundant neurohumoral system. Recently, novel molecular mediators and regulatory pathways for feeding and body weight regulation have been identified in the brain and the periphery. Because of the multitude and complexity of disturbances in energy intake, expenditure, and partitioning that are associated with obesity, it has been difficult to determine which abnormalities are causative versus less important phenomena that are consequences of the altered neuroendocrine and metabolic milieu. Transgenic technology has provided new opportunities to modify the complex body weight-regulating system and to assess the relative importance of the individual components. Observations of mutant mice have shed new light on the understanding of energy homeostasis equation. Once created, transgenic animal models may be useful in assessing the efficacy or determining the mode of action of potential new therapeutic agents. However, the interpretation of targeted mutation is sometimes not straightforward in unraveling the physiology because of the redundancy and compensation of the regulatory machinery, as well as the inherent problems of manipulation of the gene. Modifying the synthesis of a particular gene at all sites and developmental stages may be a relatively crude way of investigating its functions. Advanced gene-targeting strategies aimed at specific alterations (on and off) of a gene product at desired tissues and times could lead to a better understanding of the system. PMID- 10699155 TI - Role of high-affinity receptors and membrane transporters in nonsynaptic communication and drug action in the central nervous system. AB - Neurochemical and morphological evidence has shown that some neurotransmitters or substances may be released from both synaptic and nonsynaptic sites for diffusion to target cells more distant than those observed in regular synaptic transmission. There are functional interactions between neurons without synaptic contacts, and matches between release sites and localization of receptors sensitive to the chemical signal are exceptions rather than the rule in the central nervous system. This also indicates that besides cabled information signaling (through synapses), there is a "wireless" nonsynaptic interaction between axon terminals. This would be a form of communication transitional between discrete classical neurotransmission (in Sherrington's synapse) and the relatively nonspecific neuroendocrine secretion. Recent findings indicate that in addition to monoamines (norepinephrine, dopamine, serotonin), other transmitters, such as acetylcholine and nitric oxide (NO), may also be involved in these nonsynaptic interactions. It has been shown that NO, an ideal mediator of nonsynaptic communication, can influence the function of uptake carrier systems, which may be an important factor in the regulation of extracellular concentration of different transmitters. This review will focus on the role of nonsynaptic receptors and transporters in presynaptic modulation of chemical transmission in the central nervous system. The nonsynaptic interaction between neurons mediated via receptors and transports of high affinity not localized in synapses has the potential to be an important contributor to the properties and function of neuronal networks. In addition, it will be suggested for the first time that the receptors and transporters expressed nonsynaptically and being of high affinity are the target of drugs taken by the patient. PMID- 10699156 TI - C1-Esterase inhibitor: an anti-inflammatory agent and its potential use in the treatment of diseases other than hereditary angioedema. AB - C1-esterase inhibitor (C1-Inh) therapy was introduced in clinical medicine about 25 years ago as a replacement therapy for patients with hereditary angioedema caused by a deficiency of C1-Inh. There is now accumulating evidence, obtained from studies in animals and observations in patients, that administration of C1 Inh may have a beneficial effect as well in other clinical conditions such as sepsis, cytokine-induced vascular leak syndrome, acute myocardial infarction, or other diseases. Activation of the complement system, the contact activation system, and the coagulation system has been observed in these diseases. A typical feature of the contact and complement system is that on activation they give rise to vasoactive peptides such as bradykinin or the anaphylatoxins, which in part explains the proinflammatory effects of either system. C1-Inh, belonging to the superfamily of serine proteinase inhibitors (serpins), is a major inhibitor of the classical complement pathway, the contact activation system, and the intrinsic pathway of coagulation, respectively. It is, therefore, endowed with anti-inflammatory properties. However, inactivation of C1-Inh occurs locally in inflamed tissues by proteolytic enzymes (e.g., elastase) released from activated neutrophils or bacteria thereby leading to increased local activation of the various host defense systems. Here we will give an overview on the biochemistry and biology of C1-Inh. We will discuss studies addressing therapeutic administration of C1-Inh in experimental and clinical conditions. Finally, we will provide an explanation for the therapeutic benefit of C1-Inh in so many different diseases. PMID- 10699157 TI - Molecular interactions with mercury in the kidney. AB - Mercury is unique among the heavy metals in that it can exist in several physical and chemical forms, including elemental mercury, which is a liquid at room temperature. All forms of mercury have toxic effects in a number of organs, especially in the kidneys. Within the kidney, the pars recta of the proximal tubule is the most vulnerable segment of the nephron to the toxic effects of mercury. The biological and toxicological activity of mercurous and mercuric ions in the kidney can be defined largely by the molecular interactions that occur at critical nucleophilic sites in and around target cells. Because of the high bonding affinity between mercury and sulfur, there is particular interest in the interactions that occur between mercuric ions and the thiol group(s) of proteins, peptides and amino acids. Molecular interactions with sulfhydryl groups in molecules of albumin, metallothionein, glutathione, and cysteine have been implicated in mechanisms involved in the proximal tubular uptake, accumulation, transport, and toxicity of mercuric ions. In addition, the susceptibility of target cells in the kidneys to the injurious effects of mercury is modified by a number of intracellular and extracellular factors relating to several thiol containing molecules. These very factors are the theoretical basis for most of the currently employed therapeutic strategies. This review provides an update on the current body of knowledge regarding the molecular interactions that occur between mercury and various thiol-containing molecules with respect to the mechanisms involved in the renal cellular uptake, accumulation, elimination, and toxicity of mercury. PMID- 10699158 TI - International union of pharmacology. XXII. Nomenclature for chemokine receptors. AB - Chemokine receptors comprise a large family of seven transmembrane domain G protein-coupled receptors differentially expressed in diverse cell types. Biological activities have been most clearly defined in leukocytes, where chemokines coordinate development, differentiation, anatomic distribution, trafficking, and effector functions and thereby regulate innate and adaptive immune responses. Pharmacological analysis of chemokine receptors is at an early stage of development. Disease indications have been established in human immunodeficiency virus/acquired immune deficiency syndrome and in Plasmodium vivax malaria, due to exploitation of CCR5 and Duffy, respectively, by the pathogen for cell entry. Additional indications are emerging among inflammatory and immunologically mediated diseases, but selection of targets in this area still remains somewhat speculative. Small molecule antagonists with nanomolar affinity have been reported for 7 of the 18 known chemokine receptors but have not yet been studied in clinical trials. Virally encoded chemokine receptors, as well as chemokine agonists and antagonists, and chemokine scavengers have been identified in medically important poxviruses and herpesviruses, again underscoring the importance of the chemokine system in microbial pathogenesis and possibly identifying specific strategies for modulating chemokine action therapeutically. The purpose of this review is to update current concepts of the biology and pharmacology of the chemokine system, to summarize key information about each chemokine receptor, and to describe a widely accepted receptor nomenclature system, ratified by the International Union of Pharmacology, that is facilitating clear communication in this area. PMID- 10699159 TI - Improved graft survival after renal transplantation in the United States, 1988 to 1996. AB - BACKGROUND: The introduction of cyclosporine has resulted in improvement in the short-term outcome of renal transplantation, but its effect on the long-term survival of kidney transplants is not known. METHODS: We analyzed the influence of demographic characteristics (age, sex, and race), transplant-related variables (living or cadaveric donor, panel-reactive antibody titer, extent of HLA matching, and cold-ischemia time), and post-transplantation variables (presence or absence of acute rejection, delayed graft function, and therapy with mycophenolate mofetil and tacrolimus) on graft survival for all 93,934 renal transplantations performed in the United States between 1988 and 1996. A regression analysis adjusted for these variables was used to estimate the risk of graft failure within the first year and more than one year after transplantation. RESULTS: From 1988 to 1996, the one-year survival rate for grafts from living donors increased from 88.8 to 93.9 percent, and the rate for cadaveric grafts increased from 75.7 to 87.7 percent. The half-life for grafts from living donors increased steadily from 12.7 to 21.6 years, and that for cadaveric grafts increased from 7.9 to 13.8 years. After censoring of data for patients who died with functioning grafts, the half-life for grafts from living donors increased from 16.9 years to 35.9 years, and that for cadaveric grafts increased from 11.0 years to 19.5 years. The average yearly reduction in the relative hazard of graft failure after one year was 4.2 percent for all recipients (P<0.001), 0.4 percent for those who had acute rejection (P=0.57), and 6.3 percent for those who did not have acute rejection (P<0.001). CONCLUSIONS: Since 1988, there has been a substantial increase in short-term and long-term survival of kidney grafts from both living and cadaveric donors. PMID- 10699160 TI - Prevention of rejection in cardiac transplantation by blockade of the interleukin 2 receptor with a monoclonal antibody. AB - BACKGROUND: Alloantigen-activated T cells express the high-affinity interleukin-2 receptor. Specific blockade of this receptor with the human IgG1 monoclonal antibody daclizumab may prevent rejection of allografts after cardiac transplantation without inducing global immunosuppression. METHODS: We randomly assigned 55 nonsensitized patients undergoing a first cardiac transplantation to receive either induction therapy with daclizumab (1.0 mg per kilogram of body weight), given intravenously within 24 hours after cardiac-transplantation surgery and every two weeks thereafter, for a total of five doses, or generalized immunosuppressive therapy. Concomitant immunosuppression was achieved in both groups with cyclosporine, mycophenolate mofetil, and prednisone. The primary end points were the incidence and severity of acute rejection, and the length of time to a first episode of biopsy-confirmed rejection. RESULTS: Of the 55 patients in the study, 28 were randomly assigned to receive daclizumab and 27 served as the control group. During induction therapy, the mean frequency of acute rejection episodes (defined as a histologic grade of 2 or higher according to the classification of the International Society of Heart and Lung Transplants) was 0.64 per patient in the control group and 0.19 per patient in the daclizumab group (P=0.02). Acute rejection developed in 17 of 27 patients in the control group (63 percent), as compared with 5 of 28 patients in the daclizumab group (18 percent; relative risk, 2.8; 95 percent confidence interval, 1.1 to 7.4; P=0.04). Throughout follow-up, there were nine patients with episodes of acute rejection of histologic grade 3 in the control group, as compared with two in the daclizumab group (P= 0.03), and the time to a first episode of rejection was significantly longer in the daclizumab group (P=0.04). There were no adverse reactions to daclizumab and no significant differences between the groups in the incidence of infection or cancer during follow-up. CONCLUSIONS: Induction therapy with daclizumab safely reduces the frequency and severity of cardiac-allograft rejection during the induction period. PMID- 10699161 TI - Cultivation of the bacillus of Whipple's disease. AB - BACKGROUND: Whipple's disease is a systemic bacterial infection, but to date no isolate of the bacterium has been established in subculture, and no strain of this bacterium has been available for study. METHODS: Using specimens from the aortic [corrected] valve of a patient with endocarditis due to Whipple's disease, we isolated and propagated a bacterium by inoculation in a human fibroblast cell line (HEL) with the use of a shell-vial assay. We tested serum samples from our patient, other patients with Whipple's disease, and control subjects for the presence of antibodies to this bacterium. RESULTS: The bacterium of Whipple's disease was grown successfully in HEL cells, and we established subcultures of the isolate. Indirect immunofluorescence assays showed that the patient's serum reacted specifically against the bacterium. Seven of 9 serum samples from patients with Whipple's disease had IgM antibody titers of 1:50 or more, as compared with 3 of 40 samples from the control subjects (P<0.001). Polyclonal antibodies against the bacterium were generated by inoculation of the microorganism into mice and were used to detect bacteria in the excised cardiac tissue from our patient on immunohistochemical analysis. The 16S ribosomal RNA gene of the cultured bacterium was identical to the sequence for Tropheryma whippelii identified previously in tissue samples from patients with Whipple's disease. The strain we have grown is available in the French National Collection. CONCLUSIONS: We cultivated the bacterium of Whipple's disease, detected specific antibodies in tissue from the source patient, and generated specific antibodies in mice to be used in the immunodetection of the microorganism in tissues. The development of a serologic test for Whipple's disease may now be possible. PMID- 10699162 TI - Early expression of angiogenesis factors in acute myocardial ischemia and infarction. AB - BACKGROUND: When the myocardium is deprived of blood, a process of ischemia, infarction, and myocardial remodeling is initiated. Hypoxia-inducible factor 1 (HIF-1) is a transcriptional activator of vascular endothelial growth factor (VEGF) and is critical for initiating early cellular responses to hypoxia. We investigated the temporal and spatial patterns of expression of the alpha subunit of HIF-1 (HIF-1alpha) and VEGF in specimens of human heart tissue to elucidate the early molecular responses to myocardial hypoxia. METHODS: Ventricular-biopsy specimens from 37 patients undergoing coronary bypass surgery were collected. The specimens were examined by microscopy for evidence of ischemia, evolving infarction, or a normal histologic appearance. The specimens were also analyzed with the reverse-transcriptase polymerase chain reaction for HIF-1alpha and VEGF messenger RNA (mRNA) expression and by immunohistochemical analysis for the location of the HIF-1alpha and VEGF proteins. RESULTS: HIF-1alpha mRNA was detected in myocardial specimens with pathological evidence of acute ischemia (onset, <48 hours before surgery) or early infarction (onset, <24 hours before surgery). In contrast, VEGF transcripts were seen in specimens with evidence of acute ischemia or evolving infarction (onset, 24 to 120 hours before surgery). Patients with normal ventricles or evidence of infarction in the distant past had no detectable levels of either VEGF mRNA or HIF-1alpha mRNA. HIF-1alpha immunoreactivity was detected in the nuclei of myocytes and endothelial cells, whereas VEGF immunoreactivity was found in the cytoplasm of endothelial cells lining capillaries and arterioles. CONCLUSIONS: An increase in the level of HIF 1alpha is an early response to myocardial ischemia or infarction. This response defines, at a molecular level, one of the first adaptations of human myocardium to a deprivation of blood. HIF-1alpha is a useful temporal marker of acutely jeopardized myocardium. PMID- 10699163 TI - Images in clinical medicine. Varicella gangrenosa. PMID- 10699164 TI - Neurologic complications of the reactivation of varicella-zoster virus. PMID- 10699165 TI - Improving the success of organ transplantation. PMID- 10699166 TI - Whipple's disease--past, present, and future. PMID- 10699167 TI - A nuclear receptor to prevent colon cancer. PMID- 10699168 TI - Death and the research imperative. PMID- 10699169 TI - The np 3243 MELAS mutation: damned if you aminoacylate, damned if you don't. AB - The np 3243 MELAS mtDNA mutation in tRNA(leu(UUR))has been variously proposed as a loss-of-function or as a gain-of-function mutation, based on apparently contradictory studies in cultured cell lines. A new report describing the molecular effects of the mutation in vivo now mirrors this variability. This should prompt a more systematic re-investigation of cells carrying the mutation, in order to separate primary from secondary and pathogenic from compensatory effects, all of which may contribute to disease phenotype. Nuclear genetic and developmental background, mitochondrial haplotype, and epigenetic effects may all influence the pathological outcome. Defects in both base-modification and aminoacylation of the mutant tRNA could play critical roles. PMID- 10699170 TI - Decreased aminoacylation of mutant tRNAs in MELAS but not in MERRF patients. AB - Mutations in human mitochondrial tRNA genes are associated with a number of multisystemic disorders. Using an assay that combines tRNA oxidation and circularization we have determined the relative amounts and states of aminoacylation of mutant and wild-type tRNAs in tissue samples from patients with MELAS syndrome (mito- chondrial myopathy, encephalopathy, lactic acidosis, stroke like episodes) and MERRF syndrome (myoclonus epilepsy with ragged red fibers), respectively. In most, but not all, biopsies from MELAS patients carrying the A3243G substitution in the mitochondrial tRNA(Leu(UUR))gene, the mutant tRNA is under-represented among processed and/or aminoacylated tRNAs. In contrast, in biopsies from MERRF patients harboring the A8344G substitution in the tRNA(Lys)gene neither the relative abundance nor the aminoacylation of the mutated tRNA is affected. Thus, whereas the A3243G mutation may contribute to the pathogenesis of MELAS by reducing the amount of aminoacylated tRNA(Leu), the A8344G mutation does not affect tRNA(Lys)function in the same way. PMID- 10699171 TI - Functional analysis of ARHGAP6, a novel GTPase-activating protein for RhoA. AB - Microphthalmia with linear skin defects (MLS) is an X-linked dominant, male lethal syndrome characterized by microphthalmia, aplastic skin and agenesis of the corpus callosum, and is caused by the deletion of a 500 kb critical region in Xp22.3. Our laboratory isolated a novel rho GTPase-activating protein (rhoGAP) gene named ARHGAP6 from the MLS region. ARHGAP6 contains 14 exons encoding a 974 amino acid protein with three putative SH3-binding domains. Because exons 2-14 are deleted in all MLS patients, we hypothesized that ARHGAP6 may be responsible for some of the phenotypic features of MLS. We pursued two approaches to study the function of ARHGAP6 and its role in the pathogenesis of MLS: gene targeting of the rhoGAP domain in mouse embryonic stem cells and in vitro expression studies. Surprisingly, loss of the rhoGAP function of Arhgap6 does not cause any detectable phenotypic or behavioral abnormalities in the mutant mice. Transfected mammalian cells expressing ARHGAP6 lose their actin stress fibers, retract from the growth surface and extend thin, branching processes resembling filopodia. The ARHGAP6 protein co-localizes with actin filaments through an N-terminal domain and recruits F-actin into the growing processes. Mutation of a conserved arginine residue in the rhoGAP domain prevents the loss of stress fibers but has little effect on process outgrowth. These results suggest that ARHGAP6 has two independent functions: one as a GAP with specificity for RhoA and the other as a cytoskeletal protein that promotes actin remodeling. PMID- 10699172 TI - Chromosome 22-specific low copy repeats and the 22q11.2 deletion syndrome: genomic organization and deletion endpoint analysis. AB - The 22q11.2 deletion syndrome, which includes DiGeorge and velocardiofacial syndromes (DGS/VCFS), is the most common microdeletion syndrome. The majority of deleted patients share a common 3 Mb hemizygous deletion of 22q11.2. The remaining patients include those who have smaller deletions that are nested within the 3 Mb typically deleted region (TDR) and a few with rare deletions that have no overlap with the TDR. The identification of chromosome 22-specific duplicated sequences or low copy repeats (LCRs) near the end-points of the 3 Mb TDR has led to the hypothesis that they mediate deletions of 22q11.2. The entire 3 Mb TDR has been sequenced, permitting detailed investigation of the LCRs and their involvement in the 22q11.2 deletions. Sequence analysis has identified four LCRs within the 3 Mb TDR. Although the LCRs differ in content and organization of shared modules, those modules that are common between them share 97-98% sequence identity with one another. By fluorescence in situ hybridization (FISH) analysis, the end-points of four variant 22q11.2 deletions appear to localize to the LCRs. Pulsed-field gel electrophoresis and Southern hybridization have been used to identify rearranged junction fragments from three variant deletions. Analysis of junction fragments by PCR and sequencing of the PCR products implicate the LCRs directly in the formation of 22q11.2 deletions. The evolutionary origin of the duplications on chromosome 22 has been assessed by FISH analysis of non-human primates. Multiple signals in Old World monkeys suggest that the duplication events may have occurred at least 20-25 million years ago. PMID- 10699173 TI - Long glutamine tracts cause nuclear localization of a novel form of huntingtin in medium spiny striatal neurons in HdhQ92 and HdhQ111 knock-in mice. AB - Huntington's disease (HD) is caused by an expanded N-terminal glutamine tract that endows huntingtin with a striatal-selective structural property ultimately toxic to medium spiny neurons. In precise genetic models of juvenile HD, HdhQ92 and HdhQ111 knock-in mice, long polyglutamine segments change huntingtin's physical properties, producing HD-like in vivo correlates in the striatum, including nuclear localization of a version of the full-length protein predominant in medium spiny neurons, and subsequent formation of N-terminal inclusions and insoluble aggregate. These changes show glutamine length dependence and dominant inheritance with recruitment of wild-type protein, critical features of the altered HD property that strongly implicate them in the HD disease process and that suggest alternative pathogenic scenarios: the effect of the glutamine tract may act by altering interaction with a critical cellular constituent or by depleting a form of huntingtin essential to medium spiny striatal neurons. PMID- 10699174 TI - Genome-wide variation in recombination in female meiosis: a risk factor for non disjunction of chromosome 21. AB - Altered recombination patterns along non-disjoined chromosomes is the first molecular correlate identified for non-disjunction in humans. To understand better the factors related to this correlate, we have asked to what extent is recombination altered in an egg with a disomic chromosome: are patterns limited to the non-disjoined chromosome or do they extend to the entire cell? More specifically, we asked whether there is reduced recombination in the total genome of an egg with a non-disjoined chromosome 21 and no detectable recombination. We chose this subclass of non-disjoined chromosomes to enrich potentially for extremes in recombination. We found a statistically significant cell-wide reduction in the mean recombination rate in these eggs with non-disjoined chromosomes 21; no specific chromosomes were driving this effect. Most importantly, we found that this reduction was consistent with normal variation in recombination observed among eggs. Thus, given that recombination is a multifactorial trait, these data suggest that when the number of genome-wide recombination events is less than some threshold, specific chromosomes may be at an increased risk for non-disjunction. Further studies are required to confirm these results, to determine the importance of genetic and environmental factors that regulate recombination and to determine their impact on non-disjunction. PMID- 10699175 TI - A functional analysis of a natural variant of intercellular adhesion molecule-1 (ICAM-1Kilifi). AB - Intercellular adhesion molecule-1 (ICAM-1) is involved in a range of interactions both within the host and between the host and a number of pathogens. Recently we described a mutation within the coding region of the first N-terminal immunoglobulin-like domain of ICAM-1, present at high frequency within African populations, which increased the risk of cerebral malaria. To understand the mechanism by which such a polymorphism might be maintained despite counter selection by malaria, we have carried out functional assays using both forms of ICAM-1 as soluble Fc chimeric fusion proteins. ICAM-1Kilifi has reduced avidity for LFA-1 compared with ICAM-1ref and binding to soluble fibrinogen was completely abolished with the Kilifi variant. In Plasmodium falciparum adhesion assays, ITO4-A4u binding to ICAM-1Kilifi was reduced compared with binding to the reference form. These results allow for the possibility of balanced selection between the reference and Kilifi forms of ICAM-1 through modulation of inflammatory responses and indicate the existence of differences within ICAM-1 binding P. falciparum isolates which may be relevant to pathogenesis. PMID- 10699176 TI - Mapping of X-linked progressive retinal atrophy (XLPRA), the canine homolog of retinitis pigmentosa 3 (RP3). AB - X-linked progressive retinal atrophy (XLPRA) in the Siberian husky dog is a naturally occurring X-linked retinopathy closely resembling X-linked retinitis pigmentosa (XLRP) in humans. In affected males, initial degeneration of rods is followed by cone degeneration and complete retinal atrophy; carrier females have random patches of rod degeneration consistent with random X chromosome inactivation. By typing the XLPRA pedigree with five intragenic markers [dystrophin, retinitis pigmentosa GTPase regulator ( RPGR ), tissue inhibitor of metalloproteinases 1, androgen receptor and factor IX], we established a linkage map of the canine X chromosome, and confirmed that the order of these five genes is identical to that on the human X. XLPRA was tightly linked to an intragenic RPGR polymorphism (LOD 11.7, zero recombination), thus confirming locus homology with RP3. We cloned the full-length canine RPGR cDNA and three additional splice variants. No disease-causing mutation was found in the RPGR-coding sequence of the four splice variants characterized, a finding similar to approximately 80% of human XLRP patients whose disease maps to the RP3 locus. In addition, there were no significant differences in the proportional expression of each splice variant in normal and pre-degenerate XLPRA-affected retina. Expression of all RPGR splice variants increased later in the disease, when retinas were undergoing active degeneration. The results provide further evidence of cross-species retention of a complex splicing pattern in the 3' portion of RPGR, the functional significance of which is unknown. In addition, the possibility of another disease locus in the RP3 region is supported. PMID- 10699177 TI - Cell cycle-dependent phosphorylation of the ATRX protein correlates with changes in nuclear matrix and chromatin association. AB - Mutations in the ATRX gene are associated with an X-linked mental retardation (XLMR) syndrome most often accompanied by alpha-thalassaemia (ATR-X syndrome). The ATRX gene encodes a predicted protein of 280 kDa featuring a PHD zinc finger motif and an ATPase/helicase domain of the SWI/SNF type; the vast majority of mutations in the ATRX gene fall within these two motifs. Although these domains are suggestive of a role for ATRX in transcriptional regulation by affecting chromatin structure and/or function, the precise cellular role of the ATRX protein remains undefined. Using indirect immunofluorescence and biochemical fractionation, we demonstrate that the ATRX protein has a punctate nuclear staining pattern and that it is tightly associated with the nuclear matrix at interphase. At the onset of M phase, the ATRX protein was associated mainly with condensed chromatin. The association of the ATRX protein with chromosomes at mitosis is concomitant with phosphorylation of the protein and its association with heterochromatin protein 1alpha (HP1alpha). The phosphorylation-dependent changes in localization between the nuclear matrix and condensed chromatin are consistent with a dual role for ATRX, possibly involving gene regulation at interphase and chromosomal segregation at mitosis. PMID- 10699178 TI - Genetic variants of IL-13 signalling and human asthma and atopy. AB - Asthma and atopy show epidemiological association and are biologically linked by T-helper type 2 (T(h)2) cytokine-driven inflammatory mechanisms. IL-4 operates through the IL-4 receptor (IL-4R, a heterodimer of IL-4Ralpha and either gammac or IL-13Ralpha1) and IL-13 operates through IL-13R (a heterodimer of IL-4Ralpha and IL-13Ralpha1) to promote IgE synthesis and IgE-based mucosal inflammation which typify atopy. Recent animal model data suggest that IL-13 is a central cytokine in promoting asthma, through the stimulation of bronchial epithelial mucus secretion and smooth muscle hyper-reactivity. We investigated the role of common genetic variants of IL-13 and IL-13Ralpha1 in human asthma, considering IgE levels. A novel variant of human IL-13, Gln110Arg, on chromosome 5q31, associated with asthma rather than IgE levels in case-control populations from Britain and Japan [peak odds ratio (OR) = 2.31, 95% CI 1.33-4.00]; the variant also predicted asthma and higher serum IL-13 levels in a general, Japanese paediatric population. Immunohistochemistry demonstrated that both subunits of IL 13R are prominently expressed in bronchial epithelium and smooth muscle from asthmatic subjects. Detailed molecular modelling analyses indicate that residue 110 of IL-13, the site of the charge-modifying variants Arg and Gln, is important in the internal constitution of the ligand and crucial in ligand-receptor interaction. A non-coding variant of IL-13Ralpha1, A1398G, on chromosome Xq13, associated primarily with high IgE levels (OR = 3. 38 in males, 1.10 in females) rather than asthma. Thus, certain variants of IL-13 signalling are likely to be important promoters of human asthma; detailed functional analysis of their actions is needed. PMID- 10699179 TI - Developmentally distinct effects on human epsilon-, gamma- and delta-globin levels caused by the absence or altered position of the human beta-globin gene in YAC transgenic mice. AB - The human beta-globin locus has been an important model system in the study of developmentally regulated transcription in multigene chromosomal domains. In this study, primer extension and sensitive real-time RT-PCR assays were used to quantify the effects of beta-globin sequence modifications on epsilon-, gamma- and delta-globin levels in transgenic mice. E11.5 primitive erythroid cells showed a surprisingly large increase in epsilon-globin in the absence of the beta globin gene (beta- locus), which is weakly expressed at that stage of development. E17.5 fetal liver and adult erythroid cells, in which beta-globin expression approaches its maximum, showed an unexpectedly small, statistically insignificant stimulation of gamma- and delta-globin levels in the absence of beta-globin sequence. Analysis of erythroid colonies produced by in vitro differentiation of embryonic stem cells indicated that the absence of the human beta-globin gene had no effect on gamma-globin expression. These results suggest that competitive influences need not be linked directly to transcription level or distance from the locus control region (LCR), and that the large increases in gamma-globin levels seen in some human deletional beta-thalassemias and hereditary persistence of fetal hemoglobin conditions are most likely to be due to effects other than loss of beta-globin competition. In transgenic mice with beta-globin sequences inserted between epsilon and the LCR in a beta- locus (betaup), the expression of epsilon-, gamma- and delta-globins suggested that stage-specific sensitivity to loss of LCR activity may be a more important parameter than position relative to the LCR. The relationship of these measurements of transgenic globin expression to a possible binary model of globin LCR action and to mimicry from red blood cell loss due to transgenic globin imbalances are discussed. PMID- 10699180 TI - A genetic risk factor for mouse neural tube defects: defining the embryonic basis. AB - Genetic polymorphisms are thought to play an important role in determining susceptibility to neural tube defects (NTDs), for example between different ethnic groups, but the embryonic manifestation of these polymorphic genetic influences is unclear. We have used a mouse model to test experimentally whether polymorphic variations in the pattern of cranial neural tube closure can influence susceptibility to NTDs. The site at which cranial neural tube closure begins (so-called closure 2) is polymorphic between inbred mice. Strains with a caudal location of closure 2 (e.g. DBA/2) are relatively resistant to NTDs, whereas strains with a rostrally positioned closure 2 (e.g. NZW) exhibit increased susceptibility to NTDs. We tested experimentally whether altering the position of closure 2 can affect susceptibility to cranial NTDs, by back- crossing the splotch ( Sp (2H) ) mutant gene onto the DBA/2 background. As a control, Sp (2H) was transferred onto the NZW background, which resembles splotch mice in its closure pattern. Approximately 80% of Sp (2H) homozygotes develop NTDs, both cranial (exencephaly) and spinal (spina bifida). After transfer to the DBA/2 background, the frequency of cranial NTDs was reduced significantly in Sp (2H) homozygotes, confirming a protective effect of caudal closure 2. In contrast, Sp (2H) homozygotes on the NZW background had a persistently high frequency of cranial NTDs. The frequency of spina bifida was not altered in either backcross, emphasizing the specificity of this genetic effect for cranial neurulation. These findings demonstrate that variation in the pattern of cranial neural tube closure is a genetically determined factor influencing susceptibility to cranial NTDs. PMID- 10699181 TI - A new gene involved in DNA double-strand break repair and V(D)J recombination is located on human chromosome 10p. AB - V(D)J recombination, accountable for the diversity of T cell receptor- and immunoglobulin-encoding genes, is initiated by a lymphoid-specific DNA double strand break. The general DNA repair machinery is responsible for the resolution of this break. Any defect in one of the known components of the DNA repair/V(D)J recombination machinery (Ku70, Ku80, DNA-PKcs, XRCC4 and DNA ligase IV) leads to abortion of the V(D)J rearrangement process, early block in both T and B cell maturation, and ultimately to severe combined immune deficiency (SCID) in several animal models. A human SCID condition is also characterized by an absence of mature T and B lymphocytes, and is associated with an increase in sensitivity to DNA-damaging agents (RS-SCID). None of the above-mentioned genes are defective in these patients, arguing for the likelihood of the existence of yet another unknown component of the V(D)J recombination/DNA repair apparatus. Athabascan speaking (SCIDA) Navajo and Apache Native Americans have a very high incidence of T(-)B(-)SCID. The SCIDA locus is highly linked with markers on chromosome 10p, although the exact molecular defect has not been recognized in these patients. We show here that cells with the SCIDA defect are impaired in the DNA repair phase of V(D)J recombination similarly to RS-SCID, precisely an absence of V(D)J coding joint formation. Moreover, genotyping analysis in several RS-SCID families corroborates a linkage of the RS-SCID locus to the SCIDA region on chromosome 10p. These results demonstrate the presence of a new essential DNA repair/V(D)J recombination gene in this region, the mutation of which causes RS-SCID in humans. PMID- 10699182 TI - An imprinted locus associated with transient neonatal diabetes mellitus. AB - Recently, we reported the localization of a gene for transient neonatal diabetes mellitus (TNDM), a rare form of childhood diabetes, to an approximately 5.4 Mb region of chromosome 6q24. We have also shown that TNDM is associated with both paternal uniparental disomy (UPD) of chromosome 6 and paternal duplications of the critical region. The sequencing of P1-derived artificial chromosome clones from within the region of interest has allowed us to further localize the gene and to investigate the methylation status of the region. The gene is now known to reside in a 300-400 kb region of 6q24 which contains several CpG islands. At one island we have demonstrated differential DNA methylation between patients with paternal UPD of chromosome 6 and normal controls. In addition, two patients with TNDM, in whom neither paternal UPD of chromosome 6 nor duplication of 6q24 have been found, show a DNA methylation pattern identical to that of patients with paternal UPD of chromosome 6. Control individuals show a hemizygous methylation pattern. These results show that TNDM can be associated with a methylation change and identify a novel methylation imprint on chromosome 6 associated with TNDM. PMID- 10699183 TI - Whole-genome methylation scan in ICF syndrome: hypomethylation of non-satellite DNA repeats D4Z4 and NBL2. AB - The ICF (immunodeficiency, centromeric instability and facial abnormalities) syndrome is a rare recessive disease characterized by immunodeficiency, extraordinary instability of certain heterochromatin regions and mutations in the gene encoding DNA methyltransferase 3B. In this syndrome, chromosomes 1 and 16 are demethylated in their centromere-adjacent (juxtacentromeric) heterochromatin, the same regions that are highly unstable in mitogen-treated ICF lymphocytes and B cell lines. We investigated the methylation abnormalities in CpG islands of B cell lines from four ICF patients and their unaffected parents. Genomic DNA digested with a CpG methylation-sensitive restriction enzyme was subjected to two dimensional gel electrophoresis. Most of the restriction fragments were identical in the digests from the patients and controls, indicating that the methylation abnormality in ICF is restricted to a small portion of the genome. However, ICF DNA digests prominently displayed multicopy fragments absent in controls. We cloned and sequenced several of the affected DNA fragments and found that the non satellite repeats D4Z4 and NBL2 were strongly hypomethylated in all four patients, as compared with their unaffected parents. The high degree of methylation of D4Z4 that we observed in normal cells may be related to the postulated role of this DNA repeat in position effect variegation in facio- scapulohumeral muscular dystrophy and might also pertain to abnormal gene expression in ICF. In addition, our finding of consistent hypomethylation and overexpression of NBL2 repeats in ICF samples suggests derangement of methylation regulated expression of this sequence in the ICF syndrome. PMID- 10699184 TI - Constitutive and regulated modes of splicing produce six major myotonic dystrophy protein kinase (DMPK) isoforms with distinct properties. AB - Myotonic dystrophy (DM) is the most prevalent inherited neuromuscular disease in adults. The genetic defect is a CTG triplet repeat expansion in the 3' untranslated region of the myotonic dystrophy protein kinase ( DMPK ) gene, consisting of 15 exons. Using a transgenic DMPK-overexpressor mouse model, we demonstrate here that the endogenous mouse DMPK gene and the human DMPK transgene produce six major alternatively spliced mRNAs which have almost identical cell type-dependent distribution frequencies and expression patterns. Use of a cryptic 5' splice site in exon 8, which results in absence or presence of 15 nucleotides specifying a VSGGG peptide motif, and/or use of a cryptic 3' splice site in exon 14, which leads to a frameshift in the mRNA reading frame, occur as independent stochastic events in all tissues examined. In contrast, the excision of exons 13/14 that causes a frameshift and creates a C-terminally truncated protein is clearly cell type dependent and occurs predominantly in smooth muscle. We generated all six full-length mouse cDNAs that result from combinations of these three major splicing events and show that their transfection into cells in culture leads to production of four different approximately 74 kDa full-length (heart-, skeletal muscle- or brain-specific) and two C-terminally truncated approximately 68 kDa (smooth muscle-specific) isoforms. Information on DMPK mRNA and protein isoform expression patterns will be useful for recognizing differential effects of (CTG)(n)expansion in DM manifestation. PMID- 10699185 TI - Expression of the human CFTR gene from episomal oriP-EBNA1-YACs in mouse cells. AB - Plasmids carrying the origin of plasmid replication ( oriP ) and expressing the EBNA-1 protein from the Epstein-Barr virus replicate and segregate in human cells and are thus potentially useful vectors for gene therapy. As very large circular molecules, up to 660 kb in size, can be maintained episomally using this system, it is possible to include intact human genes with all their long-range controlling elements which might give high levels of tissue-specific and controlled gene expression. We have shown previously that a 320 kb yeast artificial chromosome (YAC) carrying the intact human CFTR gene can complement the Cambridge null cystic fibrosis mice as a transgene. We have now modified this YAC to a circular molecule carrying both oriP and the EBNA-1 gene. We show that this oriP-EBNA1-YAC can be stably maintained as unrearranged episomes in mouse LA 9 cells, which do not express endogenous cftr, and in mouse CMT-93 cells, which do express endogenous cftr. The human CFTR gene is expressed in some of the cell lines, but the level of expression is very variable between cell lines and is not related to the copy number of the elements. PMID- 10699186 TI - Beta-globin YAC transgenes exhibit uniform expression levels but position effect variegation in mice. AB - Expression of a construct integrated at different genomic locations often varies because of position effects that have been subcategorized as stable (decreased level of expression) and variegating (decreased proportion of expressing cells). It is well established that locus control regions (LCRs) generally overcome position effects in transgenes. However, whether stable and variegated position effects are equally overcome by an intact LCR has not been determined. We report that single-copy yeast artificial chromosome transgenes containing an unmodified human beta -globin locus were not subject to detectable stable position effects but did undergo mild to severe variegating position effects at three of the four non-centromeric integration sites tested. We also find that, at a given integration site, the distance and the orientation of the LCR relative to the regulated gene contributes to the likelihood of variegating position effects, and can affect the magnitude of its transcriptional enhancement. DNase I hypersensitive site (HSS) formation varies with the proportion of expressing cells, not the level of gene expression, suggesting that silencing of the transgene is associated with a lack of HSS formation in the LCR region. We conclude that transcriptional enhancement and variegating position effects are caused by fundamentally different but inter-dependent mechanisms. PMID- 10699187 TI - Spectrum of SPG4 mutations in autosomal dominant spastic paraplegia. AB - Autosomal dominant hereditary spastic paraplegia (AD-HSP) is a group of genetically heterogeneous neurodegenerative disorders characterized by pro- gressive spasticity of the lower limbs. Five AD-HSP loci have been mapped to chromosomes 14q, 2p, 15q, 8q and 12q. The SPG4 locus at 2p21-p22 has been shown to account for approximately 40% of all AD-HSP families. SPG4 encoding spastin, a putative nuclear AAA protein, has recently been identified. Here, sequence analysis of the 17 exons of SPG4 in 87 unrelated AD-HSP patients has resulted in the detection of 34 novel mutations. These SPG4 mutations are scattered along the coding region of the gene and include all types of DNA modification including missense (28%), nonsense (15%) and splice site point (26.5%) mutations as well as deletions (23%) and insertions (7.5%). The clinical analysis of the 238 mutation carriers revealed a high proportion of both asymptomatic carriers (14/238) and patients unaware of symptoms (45/238), and permitted the redefinition of this frequent form of AD-HSP. PMID- 10699188 TI - Mutation of the receptor tyrosine kinase gene Mertk in the retinal dystrophic RCS rat. AB - Vertebrate photoreceptor cells are the basic sensory apparatus of the retina, capable of converting the energy of absorbed photons into neuronal signals. The proximal portions of mammalian photoreceptor outer segments are synthesized daily by cell bodies, and outer segment tips are shed with a circadian rhythm, resulting in a complete turnover of outer segments about every 9 days. The shed outer segments are phagocytosed by adjacent retinal pigment epithelial (RPE) cells, and metabolites are recycled to photoreceptors. The Royal College of Surgeons (RCS) rat is a widely studied, classic model of recessively inherited retinal degeneration in which the RPE fails to phagocytose shed outer segments, and photoreceptor cells subsequently die. We have used a positional cloning approach to study the rdy (retinal dystrophy) locus of the RCS rat. Within a 0.3 cM genetic inclusion interval, we have discovered a small deletion of RCS DNA that disrupts the gene encoding the receptor tyrosine kinase Mertk. The deletion includes the splice acceptor site upstream of the second coding exon of Mertk and results in a shortened transcript that lacks this exon. The aberrant transcript joins the first and third coding exons, leading to a frameshift and a translation termination signal 20 codons after the AUG. The concordance of these and other data indicate that Mertk is probably the gene for rdy. Our results provide genetic evidence for an essential role of a receptor tyrosine kinase in a specialized form of phagocytosis and suggest a molecular model for ingestion of outer segments by RPE cells. PMID- 10699189 TI - Transduction of 3'-flanking sequences is common in L1 retrotransposition. AB - Active LINE-1 (L1) elements possess the ability to transduce non-L1 DNA flanking their 3' ends to new genomic locations. Occasionally, the 3' end processing machinery may bypass the L1 polyadenylation signal and instead utilize a second downstream polyadenylation site. To determine the frequency of L1-mediated transduction in the human genome, we selected 66 previously uncharacterized L1 sequences from the GenBank database. Fifteen (23%) of these L1s had transposed flanking DNA with an average transduction length of 207 nucleotides. Since there are approximately 400 000 L1 elements, we estimate that insertion of transduced sequences alone may have enlarged the diploid human genome as much as 19 Mb or 0.6%. We also examined 24 full-length mouse L1s and found two long transduced sequences. Thus, L1 retrotransposition in vivo commonly transduces sequence flanking the 3' end of the element. PMID- 10699190 TI - Maternal and fetal serum nitric oxide (NO) concentrations in normal pregnancy, pre-eclampsia and eclampsia. AB - OBJECTIVES: To measure the maternal and fetal serum concentrations of total nitrites and nitrates (as an index of nitric oxide production) in normal pregnancy, pre-eclampsia and eclampsia. DESIGN: Three groups of women were studied cross-sectionally: late pregnant women with pre-eclampsia and eclampsia (n=31); normal late pregnant women (n=32); and age-matched healthy non-pregnant women (n=21). Venous blood samples were collected from all women and both maternal and umbilical venous samples were collected from pregnant women. METHODS: Blood samples were assayed for nitric oxide (NO) production by Greiss reaction which measures the combined oxidation products of NO (total nitrites and nitrates). RESULTS: There was a significant increase in serum total nitrites and nitrates concentrations in normal pregnant women than in the serum of age-matched normal non-pregnant women (P<0.0001). Significantly higher total nitrites and nitrates levels were found in the maternal sera of the pre-eclamptic and eclamptic women compared with those of normal pregnant women (P<0.0001). Also, fetal blood levels of total nitrites and nitrates were significantly increased in pre-eclampsia and eclampsia compared with those of normal pregnancy (P<0.0001). CONCLUSIONS: (1) Serum nitric oxide (NO) production is increased in normal pregnancy than in the normal non-pregnancy. (2) Maternal and fetal serum NO levels are increased significantly in pre-eclampsia and eclampsia, which possibly represents a compensatory/protective mechanism to maintain blood flow and limit platelets aggregation in the fetal-maternal circulations. (3) The increase in NO production is directly related to the severity of pre-eclampsia; this would be of diagnostic significance for the prediction of the severity of this syndrome. PMID- 10699191 TI - Biochemical screening for Down syndrome: patients' perception of risk. AB - OBJECTIVES: To determine the utility of the triple test in routine clinical practice and in addition to the document, the acceptability of a cut-off of 1:250 for invasive testing. DESIGN: Retrospective analysis of data from screening and invasive testing for Down syndrome over a 5-year period in Hull Maternity Hospital. Computer-based records were accessed and individual data drawn from case notes were analyzed. RESULTS: 14827 (78%) of all patients opted for the triple test. A positive result (1:250 or greater) was found in 586 (4%). Fifteen percent of this group refused further testing with amniocentesis. 0.08% requested amniocentesis despite a negative triple test result. Of the screened pregnancies the triple test and selective invasive testing identified nine out of 15 (60%) of Down syndrome cases. CONCLUSION: Sixty percent of Down syndrome pregnancies were identified with a 4% invasive testing rate. Fifteen percent of women who had a positive test did not agree with the cut-off of 1:250 and therefore declined invasive testing. Invasive procedure complication rates do not equate with patients' perception of Down syndrome. PMID- 10699192 TI - Declining fetal growth standards in Enugu, Nigeria. AB - OBJECTIVE: To compare the birth-weights of babies born before and after the introduction of the Structural Adjustment Program in Nigeria. METHODS: A retrospective analysis of the birth-weights of all singleton live births at the University of Nigeria Teaching Hospital Enugu, Eastern Nigeria, in 1984 and 1998. RESULTS: The babies were on average 183 g lighter in 1998 than in 1984. The mean birth-weight for each gestational age, parity and fetal sex was significantly lower in 1998 than in 1984. The incidences of low birth-weight, intrauterine growth retardation and preterm delivery were significantly higher in 1998 than in 1984. The above findings were associated with a significant reduction in the mean gestational age, parity and utilization of the hospital's maternity services, probably consequences of an adverse socio-economic environment. CONCLUSION: Reversing the declining growth standards of Nigerian fetuses would entail improving the socio-economic status of Nigerian women as well as the quality of maternity care. PMID- 10699193 TI - Prenatal detection of a high-risk group for intrauterine growth restriction based on sonographic fetal biometry. AB - OBJECTIVE: This study was performed to evaluate the significance of sonographic fetal biometry in predicting low birth weight. METHOD: Five hundred and sixty eight single-term pregnancies were analyzed. They were stratified into seven subgroups by birth weight deviation (BWD). Among the 568 pregnancies, 115 were revealed to be small-for-gestational-age (SGA) (birth weight less than mean -1.5 S. D.). When IUGR was suspected by routine sonographic fetal biometry, 'IUGR work up' was carried out. The diagnostic performance of our screening method for the detection of SGA pregnancies in the general population was calculated. RESULT: The sensitivity, specificity, positive predictive value, negative predictive value and odds ratio of our screening method for the detection of SGA pregnancies in the general population were 73.0, 96.6, 83.2, 98.0% and 131.0, respectively. CONCLUSION: These data suggest that sonographic biometry is useful for the prenatal detection of high-risk cases of fetal growth restriction. PMID- 10699194 TI - Intrafascial abdominal hysterectomy: outcomes and complications of 867 operations. AB - OBJECTIVE: To prospectively evaluate the results and complications of the surgical technique of intrafascial abdominal hysterectomy. METHODS: From March 1993 to February 1998, 867 women at four institutions from the Department of Valle del Cauca, Colombia, underwent intrafascial abdominal hysterectomy. Information on sociodemographic and clinical characteristics before hysterectomy, indications for hysterectomy, surgical outcomes and intra- and post-operative complications were collected. Patients were evaluated at 1, 3, and 12 months post operatively and annually thereafter. RESULTS: The follow-up period ranged from 6 to 63 months (median=45 months). The mean blood loss was 286+/-112 ml. Operative time averaged 71+/-11 min. The overall operative site infection rate was 4%. Intra- and post-operative hemorrhage occurred in 0.2 and 1.0% of patients, respectively. The transfusion rate was 0.7%. The incidences of ureteral, bladder, and bowel injury were 0.1, 0.4 and 0.0%, respectively. To date, none of the patients followed up between 1 and 5 years have had evidence of vaginal vault prolapse. CONCLUSIONS: Intrafascial abdominal hysterectomy is a safe technique with a low rate of complications. PMID- 10699195 TI - Use of the abdominal wall fat index determined ultrasonographically for assessing the risk of post-operative pulmonary embolism. AB - OBJECTIVE: To assess the usefulness of the abdominal wall fat index (AFI) for predicting pulmonary embolism (PE) after gynecologic surgery. METHOD: The subjects were 115 female patients who underwent laparotomy for gynecologic disease. They were divided into low-dose heparin therapy (n=28) and control, without heparin (n=87) groups. The AFI ratio of the maximum preperitoneal fat thickness to the minimum subcutaneous fat thickness was determined using ultrasonography. RESULT: Post-operative PE occurred in four control patients. If the cut-off value of the AFI for predicting PE development was set at more than or equal to 0.85, the sensitivity was 1.00 and the specificity was 0.60 (P=0.03). If we restricted the criteria, and set the cut-off value of the AFI to more than or equal to 0.85 in patients with hypertension [systolic blood pressure (BP)>/=140 mmHg, diastolic BP>/=90 mmHg or patients were taking antihypertensive medication], the sensitivity and specificity were 0. 75 and 0.92, respectively (P=0.001). CONCLUSION: Visceral fat obesity is a risk factor for PE after gynecologic surgery, and the AFI is useful for predicting PE and for whom post operative low-dose heparin therapy may be indicated. PMID- 10699196 TI - A 10-year follow-up study of contraceptive Norplant implants. AB - OBJECTIVES: To evaluate the long-term efficacy and health problems associated with Norplant implants and re-implants after 5 full years of first implants. METHOD: From 1984 to 1988, 1657 women accepted first implants of Type I and Type II of Norplant, and 394 first acceptors had a re-implant, at a clinic for study. The clinical records and annual follow-up data of acceptors were analyzed. The life-table technique was the main method used. RESULTS: The continuation rates were very high. The cumulative pregnancy rates at 1, 3 and 5 full years of use were 0.0%, 0.1% and 0.7%, respectively. Users with heavier body weight had higher pregnancy rates. The rate of menstrual disturbances peaked at 73% after 3 months and consistently decreased to 20% at 5 years of use. Rates of menstrual disturbances associated with re-implants were much lower. CONCLUSION: Norplant is extremely effective and safe for long-term use. PMID- 10699197 TI - Intensive care for critically ill obstetric patients. PMID- 10699198 TI - A comparison of phentolamine and magnesium sulfate therapy in pre-eclampsia. PMID- 10699199 TI - Successful pregnancy following antenatal closure of uterine wall defect. PMID- 10699200 TI - Primary ovarian abscess. PMID- 10699201 TI - Intravesical migration of copper-T. PMID- 10699202 TI - Safety of three formulations of nonoxynol-9 containing vaginal spermicides. N-9 Formulation Preferences Study Group Committee. PMID- 10699203 TI - Life-threatening ovarian hyperstimulation syndrome. PMID- 10699204 TI - Postmenopausal leimyomatosis peritonealis disseminata. PMID- 10699205 TI - Preoperative rectosigmoidoscopy in gynecologic cancer patients. PMID- 10699206 TI - Citations from the literature. This is a selection of abstracts taken from the literature in the field of gynecology and obstetrics which the Journal's editors feel may be of interest to our readers(1) PMID- 10699207 TI - Interpreting large-scale experiments on effects of trawling on benthic fauna: an empirical test of the potential effects of spatial confounding in experiments without replicated control and trawled areas. AB - Disturbances due to trawling and dredging is a serious threat to assemblages of benthic marine animals. We tested hypotheses about effects of trawling on benthic assemblages in a manipulative field experiment, using gear and intensities relevant to future management of trawling in a Swedish fjord. Three trawled and three control sites were sampled at several times before and after trawling was initiated. This paper describes how conclusions about effects of trawling might differ between experiments involving replicate sites and experiments using only one trawled and one control site, as in several recent studies. Analyses of selected taxa showed that abundances of many species changed differently among control sites. Differences in temporal change between pairs of single trawled and control sites were also frequent. Neither the quantitative nor the qualitative nature of differences between treatments could, however, be coherently interpreted among the different combinations of trawled and control sites. This is consistent with results obtained from analyses using all sites, which showed no consistent effects of trawling on any of these taxa. These results provide empirical evidence that spatial confounding may cause serious problems to formal interpretation of experiments, which use only one control and one trawled area. Such potential problems can best be solved by ensuring that the study incorporates more than one control site. PMID- 10699208 TI - Relationship between specific dynamic action and protein deposition in calanoid copepods. AB - The link between specific dynamic action (SDA) and protein deposition was investigated in copepodites stage V of two calanoid copepod species, the neritic Acartia tonsa and the oceanic Calanus finmarchicus. This was done by measuring respiration before, during, and after a specific feeding period and measuring the incorporation of carbon into proteins. These were also measured on individuals incubated with cycloheximide, an antibiotic that inhibits protein synthesis. The cycloheximide treatment significantly diminished the magnitude of SDA in both A. tonsa and C. finmarchicus, and inhibited carbon incorporation into protein in both species. This provides evidence that the rate at which protein deposition takes place greatly affects the magnitude of SDA. The specific respiration rates of both starving and feeding copepods were generally higher in A. tonsa than in C. finmarchicus. This influenced SDA, the magnitude of SDA normalised to an 8 h feeding period being threefold higher in A. tonsa (78.7+/-25.7 nlO(2) ugC(-1)) than in C. finmarchicus (27.5+/-11.6 nlO(2) ugC(-1)). This difference may arise due to differences in energy allocation in the organisms of the copepodite V stage of the two species. In this stage C. finmarchicus deposits large quantities of storage lipids, predominately wax esters, whereas A. tonsa deposits proteins during somatic growth. PMID- 10699209 TI - The effect of feeding or starvation on resource allocation to body components during the reproductive cycle of the sea urchin Sphaerechinus granularis (Lamarck). AB - To determine the effects of feeding or starvation on resource allocation to body components during the reproductive cycle of Sphaerechinus granularis, sea urchins were placed in laboratory tanks and either fed ad libitum or starved during two different periods of their biological cycle, i.e. the mature stage and the recovery stage. The urchin growth was monitored over the whole experimental period, the gonad, gut, lantern indices and organic matter levels of different organs were determined at the end of the experiment. During the mature stage sea urchins in good nutritional conditions did not increase in size, but allotted energy to gonad production and stored reserves in body wall. Limiting food stopped the gonadal growth without complete regression. During the recovery period food allowed somatic growth, i.e. test growth and the storage of reserves in gonad somatic cells. This somatic production did not occur under food-limited conditions and the resources allotted for survival and maintenance were taken from different body components. PMID- 10699210 TI - Energy balance and cold adaptation in the octopus Pareledone charcoti. AB - A complete energy balance equation is calculated for the Antarctic octopus Pareledone charcoti at 0 degrees C. Energy used in respiration, growth, and excretion of nitrogenous and faecal waste, was recorded along with the total consumption of energy through food, for three specimens of P. charcoti (live weights: 73, 51 and 29 g). Growth rates were very slow for cephalopods, with a mean daily increase in body weight of only 0.11%. Assimilation efficiencies were high, between 95.4 and 97.0%, which is consistent with previous work on octopods. The respiration rate in P. charcoti was low, with a mean of 2.45 mg O(2) h(-1) for a standard animal of 150 g wet mass at 0 degrees C. In the North Sea octopus Eledone cirrhosa, respiration rates of 9.79 mg O(2) h(-1) at 11.5 degrees C and 4.47 mg O(2) h(-1) at 4.5 degrees C for a standard animal of 150 g wet mass were recorded. Respiration rates between P. charcoti and E. cirrhosa were compared using a combined Q(10) value between P. charcoti at 0 degrees C and E. cirrhosa at 4.5 degrees C. This suggests that P. charcoti are respiring at a level predicted by E. cirrhosa rates at 4.5 and 11.5 degrees C extrapolated to 0 degrees C along the curve Q(10)=3, with no evidence of metabolic compensation for low temperature. PMID- 10699211 TI - Precision of different methods used for estimating the abundance of the nitrogen fixing marine cyanobacterium, Trichodesmium Ehrenberg. AB - Variances involved in estimating the abundance of the nitrogen-fixing marine cyanobacterium, Trichodesmium Ehrenberg, were evaluated by repeated sampling using bottle casts and plankton net tows at two stations in the southern East China Sea. The variance among individual samples and the variance arising from subsampling processes were separated by the method of analysis of variance, and the coefficient of variation (C.V.) of an abundance estimate based on a single subsample was calculated. For bottle-collected samples, the major source of variation came from taking subsamples from a water bottle. The C.V. of a single subsample estimate ranged from 57% to 184%. For net-collected samples, variance in abundance estimation was mainly caused by distinctive net tows, and when distributing materials in the receiving bucket into smaller containers. The C.V.s of single subsample estimates were 34% and 40%, respectively. Trichodesmium abundance estimated with bottle- and net-collected samples were further compared using data obtained from 17 stations in the East China Sea. Although a general distribution pattern was supported by both methods, the correlation coefficient between them was 0.461, not significantly different from 0. PMID- 10699212 TI - Environmental controls on growth of the massive coral Porites. AB - Annual density banding provided growth characteristics for 245 similar-sized, massive colonies of Porites from similar locations on 29 reefs from across the length and breadth of the Great Barrier Reef (GBR), Australia. Values obtained were density, extension rate, and calcification rate. Tissue thickness, the depth to which skeletons were occupied by tissue at the time of collection, was also measured. Extension rate, calcification rate, and tissue thickness were significantly greater at the top of colonies than at the sides. Extension rate and calcification rate decreased from north to south along the GBR (latitudinal range of approximately 9 degrees ) and were significantly and directly related to annual average sea surface temperature (SST; range approximately 25-27 degrees C). For each 1 degrees C rise in SST, average annual calcification increased by 0.39 g cm(-2) year(-1) and average annual extension increased by 3.1 mm year(-1) (c.f. average values of 1.63 g cm(-2) year(-1) and 12.9 mm year(-1), respectively). Density was inversely correlated with extension rate and increased with distance offshore. Data for massive Porites colonies from the GBR were extended though 20 degrees of latitude and an average annual SST range of 23-29 degrees C using published data for the Hawaiian Archipelago (Grigg, R.W., 1981. Coral reef development at high latitudes in Hawaii. Proc. 4th Int. Coral Reef Symp., Manila, Vol. 1, pp. 687-693; Grigg, R.W., 1997. Paleoceanography of coral reefs in the Hawaiian-Emperor Chain - revisited. Coral Reefs 16, S33-S38) and Phuket, Thailand (Scoffin. T.P., Tudhope. A.W., Brown. B.E., Chansang. H., Cheeney. R.F., 1992. Patterns and possible environmental controls of skeletogenesis of Porites lutea, South Thailand. Coral Reefs 11, 1-11). The response of calcification rate to temperature remained linear. Variation in annual average SST accounted for 84% of the variance. For each 1 degrees C rise in SST, average annual calcification increased by 0.33 g cm(-2) year(-1) and average annual extension increased by 3.1 mm year(-1) (c.f. average values of 1.50 g cm(-2) year(-1) and 11.6 mm year(-1), respectively). The sensitivity of calcification rate in Porites to SST, combined with observed 20th Century increases in SSTs, suggests that calcification rates may have already significantly increased along the GBR in response to global climate change. PMID- 10699213 TI - Evidence of seasonal variation in bioluminescence of Amphipholis squamata (Ophiuroidea, Echinodermata): effects of environmental factors. AB - The bioluminescence of Amphipholis squamata was assessed from freshly collected individuals for 16 successive months, and from individuals maintained in the laboratory under various experimental conditions of salinity, temperature and photoperiodic regime. Field investigations showed that bioluminescence intensity and kinetics varied seasonally, with the light produced being brighter and faster in winter and summer. The seasonal variation was not correlated with changes of ambient salinity. However, it was correlated with changes in temperature, the luminescence being brighter and faster in coldest and warmest seasons, and with the changes of photoperiod, the luminescence being brighter and faster in seasons with shortest and longest day length. Laboratory investigations also demonstrated that luminescence was not affected by salinity conditions. Conversely, luminescence was affected by temperature, the light production being brighter and faster in warmer conditions (in agreement with field observations) and dimmer and slower in colder conditions (in disagreement with field observations). Light production was also affected by photoperiod since experimental changes of natural light:dark regime caused the bioluminescence to decrease. Considering that photoperiod guides the biology of A. squamata and that reproduction takes place during coldest months in the species, an endogenous factor of neurophysiological nature linked to the ophiuroid reproductive cycle is proposed to induce the luminescence to peak in winter. This was confirmed by the fact that seasonal variation of luminescence was different between adult and juveniles, the latter showing no winter peak of luminescence. It is suggested that the luminescence normally associated with defense could also be part of an intraspecific visual signal related to individuals aggregating for reproduction during winter. PMID- 10699214 TI - Induction of heat-shock (stress) protein gene expression by selected natural and anthropogenic disturbances in the octocoral Dendronephthya klunzingeri. AB - Previously it was found that the expression of selected heat-shock proteins is upregulated in corals after exposure to elevated temperature. We published that HSPs are suitable markers in sponges to monitor the degree of environmental stress on these animals. In the present study the heat-shock proteins (HSPs) with a molecular weight of 90 kDa have been selected to prove their potential usefulness as biomarkers under controlled laboratory conditions and in the field. The studies have been performed with the octocoral Dendronephthya klunzingeri4.5 fold higher steady-state level of the respective mRNA. Also animals taken from stressed locations in the field showed an increased expression. The amount of HSP90 protein in D. klunzingeri was found to be strongly increased under thermal stress, or exposure to polychlorinated biphenyl (congener 118), but not after treatment with cadmium. Field studies revealed that samples taken from a nonstressed area have a low level of HSP90, but those collected from locations at which the corals are under physical stress (sedimentation through landfilling) show a high expression of HSP90. It is concluded that the chaperone HSP90 might become a suitable biomarker to monitor environmental stress on corals. PMID- 10699215 TI - Alteration of photoresponses involved in diel vertical migration of a crab larva by fish mucus and degradation products of mucopolysaccharides. AB - Photoresponses involved in the descent phase of nocturnal diel vertical migration (DVM) of larvae of the crab Rhithropanopeus harrisii were measured in a laboratory system that mimicked the underwater angular light distribution. The test hypothesis was that kairomones from fish that activate zooplankton photoresponses involved in DVM are derived from polysaccharides from the external mucus of fishes. Studies considered fish mucus from the mummichog (Fundulus heteroclitus) and disaccharides (originating from chondroitin sulfate A and heparin polysaccharides) that are likely constituents of fish mucus. R. harrisii larvae descend at sunrise with an isolume and remain near the isolume during the day. Since depth maintenance near the isolume depends upon a negative phototaxis, the lowest light intensity (threshold) that induces this response was used to quantify the effects of the test chemicals. It was predicted that exposure to fish kairomones would lower the photoresponse threshold, thereby resulting in larvae remaining deeper in the water column where light for visual predation was reduced. The photoresponse threshold declined as the concentration of fish mucus increased. Disaccharides originating from chondroitin sulfate A and heparin also decreased the photoresponse threshold as compared to responses in aged, filtered seawater. Collectively, the results support the hypothesis and indicate that disaccharide degradation products of predator mucus containing sulfated and acetylated amines can serve as kairomones. PMID- 10699216 TI - The feeding behaviour and competition for carrion between two sympatric scavengers on a sandy shore in Hong Kong: the gastropod, Nassarius festivus (Powys) and the hermit crab, Diogenes edwardsii (De Haan). AB - Field and laboratory experiments compared the feeding behaviours of two marine scavengers, the gastropod, Nassarius festivus and the hermit crab, Diogenes edwardsii, sympatric on the lower intertidal shore at Starfish Bay, Hong Kong. Field trials demonstrated that they both preferred bivalve and fish to other carrion. N. festivus arrived at bait in greater numbers and more quickly than D. edwardsii. Numbers of the former species attracted to fish bait (1 cm(3)) peaked at a mean of 41 after 30 min, whereas the latter peaked at a mean of only 2.3 after 55 min. Laboratory investigations revealed that N. festivus80 cm, feed quickly (x=13.20 min) and, after 14 days starvation, had a high consumption rate (0.124 mg wet weight or 0.034 mg dry weight of food individual(-1) min(-1), or 1.4% and 1.3% of its wet and dry body weights min(-1), respectively). In contrast, D. edwardsii119.75 min and a lower consumption rate (0.003 mg wet weight and 0.001 mg dry weight of food individual(-1) min(-1), or 0.1% of its wet and dry body weights min(-1), respectively). Manipulative experiments provided evidence for interspecific competition for carrion between the two species. A morphological advantage, i.e. an extendible proboscis allowing feeding at a distance, and chemoreceptors that permit long distance food detection, combined with numerical superiority on the shore, are mechanisms by which N. festivus outcompetes D. edwardsii and dominates feeding clusters. Interactive behaviour studies revealed the occurrence of interspecific interactions between the two species and intraspecific interactions among D. edwardsii but not N. festivus. The number of interspecific interactions fell when the numbers of N. festivus increased and those of D. edwardsii decreased, and vice versa. The aggressive D. edwardsii did not significantly affect the feeding behaviour of N. festivus. When the number of D. edwardsii increased and the number of N. festivus decreased, the number of intraspecific interactions in the former increased. The feeding time of N. festivus, however, decreased with increasing numbers of conspecifics and a decreasing number of D. edwardsii. This suggests the possibility of intraspecific competition in N. festivus. PMID- 10699217 TI - The effects of symbiotic crabs on the pumping activity and growth rates of Chaetopterus variopedatus. AB - This study investigates how the presence of symbiotic crabs (Pinnixa chaetopterana or Polyonyx gibbesi) in the tubes of the polychaete Chaetopterus variopedatus affects the worms' pumping activity and growth rates under laboratory and field conditions. In the field, worms whose tubes are inhabited by Pinnixa beat their fan segments significantly more frequently than do worms hosting Polyonyx, but other measures of pumping activity do not differ according to symbiont species. In the lab, worms tend to move water through their tubes at higher rates when crabs are present. In 7-month laboratory experiments, growth rates of worms hosting either species of crab did not differ from growth rates of worms without crab symbionts. Although worms hosting Polyonyx are, on average, significantly larger than worms hosting Pinnixa, this appears to be due to competition between the crab species for hosts and not due to differential effects on host growth. Unlike the crabs in this study, pea crab species inhabiting bivalves are known to have strong deleterious effects on host growth and reproduction, suggesting that the evolution of virulence in symbiotic interactions is dependent upon specific ecological context. PMID- 10699218 TI - The effects of sea urchin grazing and drift algal blooms on a subtropical seagrass bed community. AB - Subtropical seagrass beds can be subject to relatively high levels of direct herbivory and large blooms of drift algae, both of which can have important effects on the floral and faunal components of the community. Caging experiments were used to investigate these factors in a Thalassia testudinum bed in Biscayne Bay, Florida. Abundance of sea urchins, Lytechinus variegatus, and drift algae was manipulated within the cages. Naturally occurring levels of urchin grazing do not appear to affect the T. testudinum population. With experimentally increased urchin densities in the winter, seagrass shoot density and aboveground biomass decreased significantly. Similar effects were not detected in the summer, indicating that the impact of grazing on T. testudinum is lessened during this time of year. Shoot density was more vulnerable to grazing than aboveground biomass. This may be a result of grazing-induced increases in seagrass productivity, in which the remaining shoots produce more or longer leaves. In the winter, drift algal blooms form large mats that cover the seagrass canopy. Under the normal grazing regime these algal blooms do not have significant negative effects on the seagrass. With increased grazing pressure, however, there is a synergistic effect of grazing and drift algae on seagrass shoot density. At intermediate urchin density (10 per m(-2)), cages without algae did not undergo significant decreases in shoot density, while those with algae did. At the high density of urchins, the number of seagrass shoots in cages both with and without algae decreased, but the effect was more pronounced for cages with algae. Invertebrate abundance at the field site was low relative to other seagrass beds. There were no discernible effects, either positive or negative, of urchin and algae manipulations on the sampled invertebrate community. PMID- 10699219 TI - Large scale population structure and gene flow in the planktonic developing periwinkle, Littorina striata, in Macaronesia (Mollusca: Gastropoda). AB - Allozymes were used to investigate the genetic structure of 42 populations of the planktonic developing, Macaronesian periwinkle Littorina striata, throughout its entire geographic range (Azores, Madeira, Canary Islands and Cape Verde Islands). This periwinkle is presumed to have a high dispersal and gene flow potential, because it has a planktonic development. It is therefore expected to show little population genetic differentiation. Indeed, based on Wright's hierarchical F statistics, no significant genetic differentiation could be detected among populations, at any of the specified hierarchical levels (i.e. population, island, and archipelago). Nevertheless, the Cape Verde Islands seemed genetically more diverse (highest mean number of alleles per locus). The number of loci revealing a significant genetic heterogeneity increased with increasing distance between populations, while private alleles based gene flow (Nm) estimates also revealed a tendency towards a geographical pattern. The distribution of rare and private alleles, might account for these observations which suggested some geographical effect. Because of the low frequency at which these alleles occur, their influence on the genetic population structure is negligible, and not picked up by F-statistics. PMID- 10699220 TI - Eye function of Mysidacea (Crustacea) in the northern Baltic Sea. AB - Eye spectral sensitivity, [S(lambda)], was measured in seven northern Baltic mysid species using an electroretinogram technique. Their S(lambda) curves were compared with the spectral distribution of underwater light at their normal habitats. In the littoral species Neomysis integer, Praunus flexuosus and Praunus inermis, the S(lambda) maxima, [S(lambda)(max)], were in the wavelength-bands of 525-535, 505-515 and 520-530 nm respectively. The neoimmigrant species Hemimysis anomala had a S(lambda)(max) around 500 nm and high sensitivity at 393 nm, possibly indicating UV-sensitivity. S(lambda) of the pelagic species Mysis mixta and Mysis relicta sp. II was at about 505-520 nm. M. relicta sp. I from Pojoviken Bay and fresh water humic Lake Paajarvi had S(lambda)(max) at approximately 550 nm and 570 nm respectively. This is in accordance with a similar long-wavelength shift in light transmittance of the respective waters. The eyes of the latter population were also damaged by strong light. The pontocaspian neoimmigrant H. anomala is clearly adapted to waters transmitting more blue light. PMID- 10699221 TI - Elemental composition of Mysis mixta (Crustacea, Mysidacea) and energy costs of reproduction and embryogenesis under laboratory conditions. AB - Mysis mixta were reared under laboratory conditions (temperature: 9-10 degrees C; salinity: 7 per thousand, ad libitum food). Dry weight, ash, total carbon and nitrogen content of mysids (muscle tissue, eggs, and embryos of different developmental stages) have been analyzed. We found significant variations in ash content and elemental composition during growth and maturation for both sexes. The proportion of carbon in abdominal muscle decreased gradually from juveniles with body weight of 3-4 mg (42.9%) to males and gravid females ( approximately 40.0%). The nitrogen content was relatively constant (11.4% in average) with significant differences only between juveniles (11.3%) and mature females (11.6%). In embryos, carbon and nitrogen content were highest in early stages (58.6 and 14.3%, respectively). By the end of the marsupial development, carbon had decreased to 51.4% and nitrogen to 12.6%. The C:N ratio reflected the change in somatic carbon content, and the ratio decreased 6.2% from juveniles to gravid females, indicating lipids to be an energy source during maturation and reproduction. The weight-specific female investment in reproduction increases with body size. In gravid females, intersegmental growth during brooding period was observed, while males appear to store energy only for copulation and die after mating. Ontogenetic variation in body composition has implications for elemental budgets of M. mixta, its value as prey for fish and in modeling energy and nutrient cycling. PMID- 10699222 TI - Experimental evidence of predation by juvenile flounder, Platichthys flesus, on a shallow water meiobenthic community. AB - In the northern Baltic Sea juvenile flounder, Platichthys flesus, occur in high abundances on shallow sandy bottoms in late summer and autumn. They feed mainly on meiofauna and the ontogenetic switch to macrofaunal sized food occurs at a larger size here than in other areas, exemplifying the high relative importance of meiofauna. Consequently, juvenile P. flesus in the Baltic feed for a longer period on meiofauna, and could thus be expected to have a stronger predation effect on the meiofaunal assemblages. In this study the predation effects of juvenile P. flesus on meiofaunal abundances and community structure were studied using microcosms that were sampled repeatedly over a 3-week period. Significant differences between treatment and control were found for the total number of taxa, for abundances of harpacticoids, copepod nauplii and ostracods. The nematode community was not affected, but one genus, Axonolaimus, was negatively affected by predation. The predation affected meiofaunal community structure as the major taxon diversity was significantly reduced. The results suggest that the meiofauna on shallow sandy bottoms may be structured by juvenile P. flesus, and the combined predation pressure of juvenile flounder and other epibenthic predators in the area might be considerable. Microcosms are effective in testing natural predation, especially on meiofaunal communities, but field experiments should be conducted to account for the physical characteristics of the area studied. PMID- 10699223 TI - Sleep loss and daytime sleepiness in the general adult population of Japan. AB - There are few epidemiological studies on sleep loss and daytime sleepiness in the general adult population of Japan. A total of 4000 adult people, aged 20 and over, were randomly drawn from five areas of Japan, and 3030 individuals were interviewed and completed a questionnaire including information about sleep duration and sleep problems. Overall, 29% slept less than 6 h at night, 23% reported having insufficient sleep, and 6% took sleep enhancing medications. The prevalence rates were 21% for symptoms of insomnia and 15% for excessive daytime sleepiness. Symptoms of insomnia were more prevalent in the elderly, whereas young people were more likely to report short sleep duration, subjective insufficient sleep and excessive daytime sleepiness. A multiple logistic regression model revealed that excessive daytime sleepiness had significant associations with young people, short sleep duration, insomnia symptoms, subjective insufficient sleep and sleep enhancing medication use. Short sleep duration was the strongest predictor of excessive daytime sleepiness. The findings indicate that sleep loss and excessive daytime sleepiness in the Japanese adult population are common, and comparable to those reported in Western countries. Excessive daytime sleepiness in the general adult population seems more likely to be attributed to short sleep duration. PMID- 10699224 TI - Current antipsychotic dose correlates to mononuclear cell counts in the cerebrospinal fluid of psychotic patients. AB - Elevated cerebrospinal fluid (CSF) angiotensin I-converting enzyme (ACE) levels have been evidenced in patients with schizophrenia who have been treated with antipsychotics. In order to explore a possible mononuclear cell origin of CSF ACE, the authors determined CSF ACE and CSF mononuclear cell counts from 25 acutely psychotic patients, who had been drug-free for at least 4 months but started on conventional antipsychotic medication within a few days before sampling. No correlations were found between CSF to serum ACE ratio and CSF mononuclear cell counts. However, CSF total mononuclear cell count, CSF lymphocyte count, and CSF mononuclear phagocyte count evidenced significant positive correlations with current dose of antipsychotic medication expressed as chlorpromazine equivalents. The authors conclude that no indication of a relationship between mononuclear cells and CSF ACE activity was found. Surprisingly, a relationship between chlorpromazine dose and CSF mononuclear cell counts was found, which may indicate drug-related changes in cell-mediated immunity. This finding needs replication and further corroboration in well designed studies. PMID- 10699225 TI - Learning and memory impairment in cocaine-dependent and comorbid schizophrenic patients. AB - Impairments in verbal learning and memory functioning have been found to be cardinal features among individuals with schizophrenia as well as among non schizophrenic cocaine abusers. Cognitive deficits in these areas, moreover, have been associated with poor treatment response and short-term outcome. Little is known, however, about the acute effects of cocaine abuse on schizophrenic patients' learning and memory functioning. Consequently, a potentially reversible and treatable source of cognitive impairment has been virtually ignored. The present study examined the extent of verbal learning and memory impairment in a group of cocaine-dependent schizophrenic patients (n=42) and a group of non schizophrenic cocaine-dependent patients (n=21) within 72 h of the last cocaine use using the California Verbal Learning Test (CVLT). Schizophrenic patients (n=34) without any substance-use disorders were also tested in an identical time frame and served as a comparison group. Results revealed that all groups demonstrated significant learning and memory impairment relative to CVLT published age and gender corrected norms. Both cocaine-dependent and non substance abusing schizophrenic groups presented a very similar pattern of impaired learning and recall performance across all CVLT task domains. Comorbid patients, in contrast, presented with marked deficits in their ability to learn and recall verbal information relative to either schizophrenic or cocaine-only groups. Moreover, the cocaine-abusing schizophrenic patients showed significant forgetfulness of the information that they did acquire during delayed recall conditions. The performance deficits exhibited by cocaine-abusing schizophrenic patients differed not only in relative severity of impairment, but also qualitatively in their increased rates of forgetfulness of acquired information. These results are interpreted in terms of the neurobiological substrates of learning and memory and the neurobiological impact of cocaine on schizophrenic patients' cognition during the early phase of inpatient hospitalization. These results suggest that comorbid patients should be targeted for specialized remediation efforts at the beginning phases of inpatient treatment. PMID- 10699226 TI - Influence of haloperidol on the relationship of frontal lobe function to psychomotor poverty and disorganization syndromes. AB - The aim of the study was to examine effects of haloperidol on the relationships between neuropsychological measures of frontal lobe functioning and the schizophrenia syndromes of psychomotor poverty and disorganization. Twenty-one participants with schizophrenia were initially evaluated when clinically stable and chronically treated with haloperidol, and 19 were evaluated again after a 3 week haloperidol-free period. Participants were evaluated with the Trail Making Test, the Wisconsin Card Sorting Test, the Purdue Pegboard, and psychiatric rating scales at each evaluation. There were significant correlations between schizophrenia syndromes and the tests sensitive to frontal lobe function when participants were medicated but not when drug-free. No significant changes in symptom severity or motor function occurred from the medication to the medication free evaluation. The results indicate that haloperidol mediates the relationship between tests sensitive to frontal lobe function and the schizophrenia syndromes of psychomotor poverty and disorganization. This mediation effect was not attributable to changes in overall symptom severity or motor function. PMID- 10699227 TI - Smooth pursuit eye tracking and visual fixation in psychosis-prone individuals. AB - Subjects identified by Perceptual Aberration-Magical Ideation (Per-Mag) scores (n=97), Social Anhedonia (SocAnh) scores (n=45), and Physical Anhedonia (PhysAnh) scores (n=31) as well as normal controls (n=94), underwent psychophysiological and clinical assessment. This is the first published investigation of pursuit system functioning in three groups of questionnaire-identified at-risk individuals. Pursuit during a simple non-monitor tracking task was measured using root-mean-square error (RMSE) scores and pursuit gain scores. Fixation performance was measured in terms of number of saccades away from the central fixation point. The at-risk subjects were more likely to display aberrant smooth pursuit tracking than controls, though there were no significant differences between the at-risk subjects endorsing items relevant to positive-symptom schizotypy and those endorsing items pertaining to negative-symptom schizotypy. The groups did not differ significantly in their visual fixation performance. Participants were also evaluated for the presence of Axis I symptomatology and psychotic-like experiences. Neither the experimental subjects nor the control subjects displayed a significant association between ocular motor performance and psychotic-like experiences. These findings are consistent with prior evidence that pursuit tracking is a trait characteristic, independent of clinical status. PMID- 10699228 TI - Cognitive factors and stress-induced changes in catecholamine biochemistry. AB - The purpose of the present research was to determine whether dysfunctional attitudes, a cognitive attribute, predicted changes in catecholamine biochemistry. A cognitive task was used to induce stress in female subjects (n=21), and levels of plasma norepinephrine (NE) and homovanillic acid (HVA) were measured at three time points: at baseline (T1); immediately after stress exposure (T2); and 40 min later (T3). Dysfunctional attitudes were significantly and positively related to levels of plasma NE at T3, controlling for baseline levels. Dysfunctional attitudes were not significantly related to plasma HVA levels at any time point. Our findings provide initial support for the idea that dysfunctional attitudes, an attribute shown to play an important role in some forms of unipolar depression, predict stress-induced alterations in noradrenergic output. PMID- 10699229 TI - Which temperamental characteristics are associated with substance use in subjects with psychotic and mood disorders? AB - The aim of this study was to assess the associations between substance use disorders and temperamental characteristics in subjects with non-affective psychotic disorders or mood disorders. Consecutively hospitalized patients were interviewed with a structured diagnostic interview to define DSM-IV diagnoses, including those of substance use. Temperamental characteristics were measured using the Sensation-Seeking Scale (SSS), the Barratt Impulsivity Scale (BIS) and the Physical Anhedonia Scale. Inpatients (n=103) with non-affective psychotic disorders (n=45) or mood disorders (n=58) were included. Among these patients, 25.2% presented with a lifetime (LT) history of alcohol abuse/dependence and 23.3% presented with a LT history of cannabis abuse/dependence. A LT history of alcohol misuse was independently associated with higher scores at the 'experience seeking' and 'disinhibition' subscales of the SSS. A LT history of cannabis misuse was independently associated with higher scores on the 'disinhibition' subscale of the SSS and on the 'non-planning activity' subscale of the BIS. These results suggest that sensation-seeking and impulsivity are temperamental characteristics that may favor substance use in patients with psychotic or mood disorders, independently from categorical diagnoses. PMID- 10699230 TI - Analysis of catechol-O-methyltransferase and 5-hydroxytryptamine transporter polymorphisms in patients at risk for suicide. AB - Genotype frequencies of functional polymorphisms in the genes encoding the serotonin transporter (5-HTT) and the enzyme catechol-O-methyltransferase (COMT) were not different in 51 suicidal inpatients compared to 51 control subjects. Within the patient group, increased hopelessness and suicide ideation were associated with homozygosity of the 5-HTT high promotor activity allele. PMID- 10699231 TI - Polymorphism in the promoter region of the alpha(2A)-adrenergic receptor gene and panic disorders. AB - alpha(2)-Adrenergic receptors have been thought to play a crucial role in the etiology or treatment of panic disorders. Polymorphism(s) in the promoter region of the alpha(2) receptor may affect gene expression and be associated with panic disorders. We studied the polymorphism of the alpha(2A) receptor gene at position -1291 reported by Lario et al. (Lario, S., Calls, J., Cases, A., Oriola, J., Torras, A., Rivera, F., 1997. Short repeat on DNA marker at candidate locus. MspI identifies a biallelic polymorphism in the promoter region of the alpha(2A) adrenergic receptor gene. Clinical Genetics 51, 129-130.) in 114 healthy control subjects and 55 patients with panic disorders. There was no statistically significant difference between controls and patients in either genotype or allele frequency. Our results suggest there is no association between this polymorphism in the promoter region of the alpha(2A) receptor gene and panic disorders. PMID- 10699233 TI - Congenital laryngeal webs: surgical management and clinical embryology. AB - Laryngeal webs are uncommon congenital anomalies. The formation of a laryngeal web represents anomalous embryologic development of the larynx. The extent of airway involvement varies which ultimately affects surgical management. A series of five congenital laryngeal webs each with subglottic involvement is reported. One patient also had a ventral laryngeal cleft. All patients ultimately required open laryngeal reconstruction, either laryngotracheal reconstruction (LTR) or thyrotomy (laryngofissure) and silastic keel, to correct the defect and all were decannulated. Findings at surgery correlate with recent descriptions of embryonic laryngeal development though the actual mechanism by which webs develop remains unknown. The findings suggest that congenital glottic webs require accurate endoscopic diagnosis and open airway reconstruction for definitive treatment. PMID- 10699232 TI - Magnesium oxide augmentation of verapamil maintenance therapy in mania. AB - The authors compared the antimanic effects of a verapamil-magnesium oxide (V-M) combination with a verapamil-placebo combination (V-P) in patients pretreated with verapamil. BPRS scores and serum magnesium levels were compared. The V-M combination was found to be significantly more effective than V-P in reducing manic symptoms (P=0.015). Serum magnesium levels were significantly higher in the V-M group (P<0.04). These data suggest that magnesium may increase antimanic efficacy of verapamil by mechanisms which may operate at the intracellular level. The magnesium-verapamil combination may have clinical application as an adjunct to verapamil in the maintenance therapy of mania. PMID- 10699234 TI - Airway foreign bodies in childhood. AB - OBJECTIVE: To define clinical spectrum of airway foreign body aspiration in children and to evaluate the outcome and complications. METHODS: A total of 53 patients (27 girls, 26 boys) with a mean age of 30.0+/-32.7 months, who aspirated foreign bodies were treated with bronchoscopy were divided into two groups with respect to the time they were diagnosed as early (Group 1, n=22, 1 on overnight polygraphy (OSAS-children), while 32 were considered primary snorers (PS), (OAHI<1). No statistically significant difference in nasalance scores was found between the OSAS- and PS children. Both groups of children had somewhat higher mean nasalance values both for oral and nasal passage sentences than the normative values for Finnish speech. In general, the most habitual snorers had lower nasalance scores than the less frequently snoring children (P=0.05). Earlier adenoidectomy or palatine tonsillar size did not have a significant influence on the nasalance. Adenotonsillectomy did not affect the nasalance scores of the nine children operated on during the follow-up period. CONCLUSIONS: According to the present study, nasalance measurements cannot be used to predict the incidence of OSAS among snoring children. PMID- 10699241 TI - Vallecular acinic cell carcinoma in a 9-year-old girl: report of an unusual case. AB - An unusual case of acinic cell tumour of the vallecula is presented. Acinic cell carcinoma occurs usually in the major salivary glands. Minor salivary gland location is unusual and vallecular origin exceptional. This peculiar histologic tumour should now be classified as an low grade carcinoma and adequate treatment has to be initiated. The patient, a 9-year-old girl, had undergone a suprahyoid access for total tumor removal with a bilateral neck exploration. Postsurgical radiotherapy has to be done in case with perineural invasion, invaded margins, node invasion or high grade tumor. The clinical and histopathological findings are discussed in the light of the literature. PMID- 10699242 TI - Otoacoustic emissions and auditory brainstem responses after neonatal hyperbilirubinemia. AB - Severe hyperbilirubinemia often results in hearing loss. Behavioral audiometry, auditory-evoked brainstem responses (ABRs) and otoacoustic emissions (OAEs) were performed in three such patients in an attempt to localize the pathophysiology of this hearing loss. Behavioral audiometric findings in these patients (all male, 4, 15 and 25 years old) ranged from severe in the 4-year-old, moderate in the 15 year-old and slight in the 25-year-old. Where obtained, ABR wave V thresholds were elevated or ABR were absent. However, absolute and inter-wave latency measurements were not indicative of brainstem pathology. OAEs (transient and distortion product) could only be obtained in the high- or low-frequency ranges in these patients. Our findings suggest that at least some lesions producing hearing loss in severe hyperbilirubinemia are in the cochlea, especially at the outer hair cells. Finally, we found that only moderately elevated serum bilirubin levels (<20 mg/dl) may contribute to the development of sensorineural hearing loss. PMID- 10699243 TI - Tracheobronchial foreign body management in an acutely ill neonate. AB - Pediatric foreign body management has become refined in recent years, both from a diagnostic and therapeutic standpoint. History, physical examination, and radiographic evaluation performed in a timely manner can lead to safe and successful foreign body retrieval. Advancement in videoendoscopic instrumentation and anesthetic techniques enable the airway surgeon to achieve simultaneous airway stabilization and foreign body removal. In the emergency setting, the surgeon may not have access to such instrumentation. We present a case of an acutely ill, extremely low birthweight infant who incurred foreign body aspiration. Basic tools of expediency and control of the airway remind us once again that technique must precede technology. PMID- 10699244 TI - Ludwig's angina in the pediatric population: report of a case and review of the literature. AB - Ludwig's angina is a rapidly progressing cellulitis involving the submandibular neck space. It is characterized by brawny induration of the submental region and floor of mouth, bearing the potential for rapid airway obstruction. Airway management, antibiotics, and judicious surgical intervention are the mainstays of successful therapy. We present a case of Ludwig's angina in a 5-year-old child and offer a meta-analysis of pediatric cases of Ludwig's angina described in the literature over the past 30 years. The presentation, etiology, management, and potential complications of this disorder in the pediatric population are discussed. PMID- 10699245 TI - Large intraosseous arteriovenous malformation of the maxilla - a case report with review of literature. AB - Large intraosseous arteriovenous malformations (AVM) of the maxilla are rare lesions, which are probably hamartomas. We report a case of an 8-year-old child who presented with exsanguinating hemorrhage after an attempted dental biopsy. The management of dental intraosseous AVMs includes transarterial embolization and direct intralesional injection of liquid acrylic (NBCA). This approach avoids mutilating surgery and its sequelae in children. We present this case for its rarity and the intralesional use of acrylic glue in its management. PMID- 10699246 TI - Solitary mastocytoma in an infant - case report with review of literature. AB - Solitary mastocytoma in infants is an uncommon disease which is characterized by mast cell hyperplasia and release of mast cell mediators. The most common presentation is pruritus. The treatment of solitary mastocytoma is symptomatic. Cutaneous mastocytoma tend to resolve by adulthood. PMID- 10699247 TI - Complications of surgical closure of tracheo-cutaneous fistula in pediatric patients - two case reports. AB - Tracheocutaneous fistula is seen frequently in decannulated children and respiratory complications associated with primary surgical closure can be potentially fatal. Cough is a precipitating factor for an air leak and we report two cases in which this occurred. A tracheotomy was performed on a 5-month-old girl for mechanical ventilation. Decannulation was successful at the first attempt. One year later, she presented with a persistent tracheo-cutaneous fistula. After surgical closure without drainage, she developed subcutaneous emphysema during a coughing episode. Sutures were removed. A 9-month-old boy presented with oxygen-dependence after lung disease and a tracheotomy was performed for respiratory support. Decannulation was successful at the first attempt 6 months later. He developed a pneumomediastinum after surgical closure of a tracheo-cutaneous fistula. Sutures were removed but replacement of a tracheotomy tube was required. In both cases the wounds were allowed to heal by secondary intention. PMID- 10699248 TI - Periorbital cellulitis secondary to ethmoiditis in a 5-week-old child. AB - Periorbital cellulitis is a condition primarily affecting young children. We present a 5-week-old boy who developed periorbital cellulitis and had a CT scan which identified acute ethmoiditis as the source of the sepsis. His clinical course is outlined, and the relevant literature is discussed. We believe this patient is the youngest case of periorbital cellulitis due to confirmed ethmoiditis reported. This emphasises the possibility of an underlying sinusitis in patients with periorbital cellulitis, even in this very young age group. PMID- 10699249 TI - A newborn's nose from the 6th millennium BC in Catal Huyuk. PMID- 10699250 TI - The selectable marker human dihydrofolate reductase enables sequential genetic manipulation of the Plasmodium berghei genome. AB - Genetic transformation of malaria parasites has been limited by the number of selectable markers available. For the rodent malaria parasite, Plasmodium berghei, only a single selection marker has been at hand, utilising the dihydrofolate reductase-thymidylate synthase gene from either P. berghei or Toxoplasma gondii to confer resistance to the anti-malarial drug pyrimethamine. Here we report the use of the human dihydrofolate reductase (hDHFR) gene as a new selectable marker, which confers resistance to the antifolate inhibitor WR99210 upon both pyrimethamine sensitive and resistant isolates of P. berghei. Transfection with circular constructs containing the hDHFR gene resulted in the generation of highly resistant parasites containing multiple copies of episomally maintained plasmids. These parasites showed around a 1000-fold increase in resistance to WR99210 compared to the parental parasites. We were also able to generate and select transgenic parasites harbouring only a single copy of hDHFR targeted into their genome, despite the fact that these parasites showed only a fivefold increase in resistance to WR99210 compared to the parental parasites. Importantly, and for the first time with malaria parasites, the hDHFR gene could be used in conjunction with the existing pyrimethamine selectable markers. This was demonstrated by reintroducing the circumsporozoite (CS) gene into transgenic CS-knockout mutant parasites that contained the P. berghei DHFR-TS selectable marker. The development of hDHFR as a second selectable marker will greatly expand the use of transformation technology in Plasmodium, enabling more extensive genetic manipulation and thus facilitating more comprehensive studies on the biology of the malaria parasite. PMID- 10699251 TI - Expression and characterisation of Plasmodium falciparum acidic basic repeat antigen expressed in Escherichia coli. AB - The acidic basic repeat antigen (ABRA) of Plasmodium falciparum has been localised on the merozoite surface and in the parasitophorous vacuole. It is one of the antigens enriched in the clusters of merozoites formed with growth inhibitory immune serum and possesses chymotrypsin-like activity. Chymostatin, an inhibitor of chymotrypsin, inhibits malaria invasion as well as autoproteolysis of ABRA. Based on these characteristics of ABRA, it seems important for invasion and should be investigated as a target for vaccine and drug design. For the functional characterisation of this protein, the full-length mature ABRA protein and its fragments with/without the putative protease active site were cloned, expressed and purified from Escherichia coli. The polyclonal serum raised against recombinant ABRA fragment recognised a parasite protein with a mobility of 101 kDa in an immunoblot assay and showed immunofluorescence activity with a schizont rich preparation of P. falciparum. Using a partially purified fragment containing the putative active site and fluorogenic and chromogenic substrates, we established that the protease activity of ABRA resides in the N-terminal portion of the protein and the highly charged C-terminal part of the protein is not required for this activity. The protease activity of ABRA was inhibited with serine protease inhibitors like chymostatin and phenyl methyl sulfonyl fluoride (PMSF) whereas leupeptin was not able to inhibit this enzyme activity. These results clearly indicated that ABRA is a protease with chymotrypsin-like specificity. This is the first report describing the expression and characterisation of recombinant ABRA protein. PMID- 10699252 TI - Isolation and characterization of Leishmania mutants defective in glycosomal protein import. AB - Kinetoplastid parasites contain a unique microbody organelle called the glycosome. Several important metabolic pathways found in the cytoplasm of higher eukaryotes are compartmentalized within the glycosome in these pathogens. This fundamental difference between the host and parasite has led to consideration of the glycosome as a potential chemotherapeutic target. The genetic basis of glycosome biogenesis is therefore of great interest. This report describes the isolation of multiple Leishmania mutant cell lines defective in glycosomal protein import, and the detailed characterization of three such lines. The mutants examined partially mislocalize a subset of glycosomal proteins to the cytosol yet retain wild-type numbers of glycosomes. One of the mutants has a mutation in the previously identified LdPEX2 (GIM1) gene. The other two mutants are demonstrated to contain cell-specific lesions in one or more genes distinct from PEX2. The identification of multiple genetically distinct mutants with defects in glycosome import provides an important genetic tool to facilitate the identification of genes involved in glycosome biogenesis. PMID- 10699253 TI - Genetic variability within the species Leishmania aethiopica does not correlate with clinical variations of cutaneous leishmaniasis. AB - Leishmania aethiopica infections in man result in a spectrum of diseases from LCL to DCL. These clinical manifestations have been attributed to genetic differences within the host or the parasites. In this study two different PCR-based methods were used to elucidate genetic variation within the species L. aethiopica. Inter- and intra-specific variations were detected in the ITS of the ribosomal operon in different strains and species of Leishmania, using a PCR-RFLP approach, and by a PCR fingerprinting technique that used single non-specific primers to amplify polymorphic regions of the genomic DNA. Both methods revealed genetic heterogeneity among ten L. aethiopica isolates examined. Unrooted distance trees separated the ten strains into two different genetic groups. This subdivision was correlated to the geographical origin of the isolates rather than to the clinical manifestation of the disease. PMID- 10699254 TI - Import of a constitutively expressed protein into mitochondria from procyclic and bloodstream forms of Trypanosoma brucei. AB - Trypanosoma brucei developmentally regulates mitochondrial function during its life cycle. Numerous nuclear encoded mitochondrial proteins undergo posttranslational regulation in a developmental fashion, but exactly how that regulation is achieved is unclear. We are interested in mitochondrial import as a potential regulatory step for nuclear encoded mitochondrial proteins. Previously, an in vitro import system was developed for the procyclic lifestage. We report here the development of an in vitro import system for bloodstream trypanosomes using a crude mitochondrial preparation. NADH dehydrogenase subunit K (NdhK) is a nuclear encoded mitochondrial protein that is constitutively expressed in bloodstream and procyclic trypanosomes. We examined the import of NdhK into procylic and bloodstream mitochondria in vitro. In both lifestages import of NdhK requires a membrane potential across the inner mitochondrial membrane, mitochondrial matrix ATP, and is time dependent. The precursor protein is processed by a matrix associated metalloprotease in a single cleavage step to mature protein. PMID- 10699255 TI - Cloning of the immunodominant 17-kDa antigen from Cryptosporidium parvum. AB - Infection with Cryptosporidium parvum causes a self-limiting diarrheal illness in immunocompetent humans and is associated with the development of a serum IgG antibody response dominated by the 27-kDa and 17-kDa parasite surface antigens. Antibodies against the 27-kDa and 17-kDa antigens may serve as useful markers for past infection in population-based studies of the risk factors associated with Cryptosporidium infection. A recombinant form of the 17-kDa antigen would be useful both in epidemiologic studies and in studies of the role of the humoral response in immunity. We have partially purified and sequenced the immunodominant 17-kDa surface antigen from sporozoites, and we have cloned a 975 bp open reading frame from C. parvum that includes all of the 17-kDa antigen peptide sequences. We show immunologic identity between a recombinant form of the protein and the native 17-kDa antigen. We conclude that the carboxy-terminal fragment of the cloned protein is the authentic 17-kDa antigen. PMID- 10699256 TI - High-throughput sequence typing of T-cell epitope polymorphisms in Plasmodium falciparum circumsporozoite protein. AB - We report a method for typing polymorphisms at the T-cell epitopes within the Th2R and Th3R regions of the Plasmodium falciparum circumsporozoite protein (CSP). This method combines the use of PCR and sequence specific oligonucleotide probes (PCR-SSOP), and allows the identification of single nucleotide polymorphisms in these epitope regions. PCR-SSOP is a robust and a high throughput sequence typing technique which has the same specificity and fidelity as direct sequencing. This method has been developed specifically for the assessment of the protective efficacy of RTS,S/SBAS2 vaccine against the 3D7 strain of P. falciparum (RTS,S/SBAS2 vaccine contains a part of the 3D7 CSP protein) in a phase IIb trial in Gambia which has been completed recently. PCR SSOP could be used to determine the allelic frequencies of other parasite antigens and their geographical distribution. PMID- 10699257 TI - Characterization of cyclic AMP phosphodiesterases in Leishmania mexicana and purification of a soluble form. AB - The cyclic AMP phosphodiesterase (PDE) activity in Leishmania mexicana is mainly located (>95%) in the soluble fraction of the cell. The intact parasite, as well as plasma membranes, showed PDE activity, probably indicating that at least part of the activity in the particulate fraction resides on the parasite cell surface, with its catalytic domain facing the extracellular moiety. For the first time, a highly specific cAMP phosphodiesterase (PDE) was purified from the soluble fraction to apparent homogeneity after a single step 2239-fold purification using pseudo-affinity chromatography on Cibacron Blue 3GA agarose. The enzyme was identified as a 61-kDa protein on SDS-PAGE, with a K(m) of 277 microM at 30 degrees C (optimum temperature). The native enzyme protein showed an apparent molecular size of approximately 200000 estimated by molecular sieve chromatography on Sephacryl S-300. Further characterization of the PDE activity present in the soluble fraction shows that the enzyme requires Mg(2+) for maximal activity. Furthermore, no activity was detected when assayed at pHs below 6.0, but above this value it increased dramatically, reaching the optimum at pH 7.2. On the basis of the K(m) and PDE activity in presence of specific drugs or modulators such as rolipram, OPC-3911, cGMP, IBMX, zaprinast, theophylline, caffeine and Ca(2+)/calmodulin, this enzyme does not seem to conform to any of the ten previously described Class I PDE families but to the PDE class II (or non mammalian PDEs) similar to the those found in Candida albicans, Dictyostelium discoideum, Saccharomyces cerevisiae or Vibrio fischeri. PMID- 10699258 TI - Plasmodium sporozoites invade cells with targeted deletions in the LDL receptor related protein. PMID- 10699259 TI - Two microsatellite loci from Brugia malayi show polymorphisms among isolates from Indonesia and Malaysia. PMID- 10699260 TI - GARP is highly conserved among Trypanosoma congolense Savannah, Forest and Kilifi subgroups. PMID- 10699261 TI - Failure to detect DNA in hydrogenosomes of Trichomonas vaginalis by nick translation and immunomicroscopy. PMID- 10699262 TI - Targeting of proteins to the nuclei of Giardia lamblia. PMID- 10699263 TI - Geographical structure and sequence evolution as inferred from the Plasmodium falciparum S-antigen locus. PMID- 10699264 TI - Meeting future challenges in topical ocular drug delivery: development of an air interfaced primary culture of rabbit conjunctival epithelial cells on a permeable support for drug transport studies. AB - The purpose of this study was to develop and characterize a functional air interfaced primary culture of rabbit conjunctival epithelial cells grown on a permeable support for drug transport studies. Conjunctival epithelial cells from the pigmented rabbit were isolated, seeded at 1.2 x 10(6) cells cm(-2) on permeable Transwell filters, and cultured at the air interface using a modified PC-1 medium. Conjunctival epithelial cell layers showed a transepithelial resistance of 1.1+/-0.1 kOmega cm(2), a potential difference of 17.0+/-0.5 mV, and an equivalent short-circuit current (I(eq)) of 16.1+/-0.4 microA cm(-2). The I(eq) was reduced by 35% using 0.01 mM bumetanide, 66% using 0.1 mM ouabain, 46% using 2 mM barium chloride (all three in the basolateral fluid), and 63% using 0.3 mM NPAA in the apical fluid, consistent with active Cl(-)-secretion across the conjunctival epithelial barrier. Amiloride-sensitive Na(+) channels were absent. The permeability of the cell layers to polar solutes decreased with increased solute size, and the calculated equivalent pore size was about 8.0 nm. The Papp of beta-blockers varied with lipophilicity in a sigmoidal fashion. Uridine transport showed temperature sensitivity and directionality, favoring transport in the apical-to-basolateral direction. Apical L-carnosine uptake was reduced by 46% in the absence of an inwardly directed proton gradient, and lowering the temperature to 4 degrees C abolished direction-dependent L-carnosine uptake. Furthermore, uptake was inhibited by 73% using apical 10 mM glycyl sarcosine (a dipeptide transporter substrate) and by 60% using 1 mM L valacyclovir (a dipeptide prodrug). In conclusion, a functional air-interfaced primary culture of rabbit conjunctival epithelial cell layers was established. This air-interfaced primary culture model may be useful for studying passive and active transport processes for ion and solute translocation in the mammalian conjunctival epithelial barrier in a defined experimental setting. PMID- 10699265 TI - Cellular delivery of functional peptides to block cytokine gene expression. AB - Advances in molecular and cellular biology have identified the cellular mediators that regulate many disease processes and have facilitated the development of new therapeutic agents that control these events. However, the size, complexity, and cellular inaccessibility of these therapeutic agents make their cellular delivery difficult. Here, we describe an efficient cellular delivery system that exploits the membrane-translocating ability of signal peptides to import functional peptides into cells. Molecular conjugates consisting of the signal import peptide (IP) and nuclear localization sequence (NLS) of the transcription factor NF kappaB were synthesized. Electrophoretic mobility shift and enzyme-linked immunosorbent assays were used to assess the inhibitory effects of these synthetic peptides on agonist-induced NF-kappaB nuclear translocation and transcriptional activation. Our results indicated that the peptides were effective in inhibiting both the nuclear translocation and transcriptional activation of NF-kappaB. However, their effects required the presence of the IP moiety for efficient cellular entry of the NLS. Structural analysis of IP showed that the hydrophobic domain, and to a lesser extent the N-terminal domain, was responsible for the membrane translocating activity of IP. PMID- 10699266 TI - Lectin-mediated drug delivery: the second generation of bioadhesives. AB - This paper reviews some recent developments in the area of bioadhesive drug delivery systems. The area of bioadhesion in drug delivery had started some 20 years ago by using so-called mucoadhesive polymers. Many of these polymers were already used as excipients in pharmaceutical formulations. This has facilitated the development of the first bioadhesive drug products, which are now commercially available. A major disadvantage of the hitherto known mucoadhesives, however, is their non-specificity with respect to the substrate. In particular for gastro-intestinal applications, this may cause some premature inactivation and moreover limits the duration of mucoadhesive bonds to the relatively fast mucus turnover. Nevertheless, for some mucoadhesive polymers other interesting functionalities were discovered, such as their ability to modulate epithelial permeability and to inhibit proteolytic enzymes. In contrast to the mucoadhesive polymers, lectins and some other adhesion molecules specifically recognize receptor-like structures of the cell membrane and therefore bind directly to the epithelial cells themselves ("cytoadhesion") rather than to the mucus gel layer. Furthermore, when bioadhesion is receptor-mediated, it is not only restricted to mere binding, but may subsequently trigger the active transport of large molecules or nanoscalic drug carrier systems by vesicular transport processes (endo-/transcytosis). Rather than only acting as a platform for controlled release systems, the concept of lectin-mediated bioadhesion therefore bears the potential for the controlled delivery of macromolecular biopharmaceuticals at relevant biological barriers, such as the epithelia of the intestinal or respiratory tract. PMID- 10699267 TI - Targeted drug delivery for brain cancer treatment. AB - The blood brain barrier (BBB) and the systemic toxicity of conventional chemotherapy present obstacles to the success of future blood-borne drug therapies of brain tumors. The work with polymer-encapsulated cancer drugs suggests an alternative and more focused treatment approach. Our experimental strategy integrates direct intracerebral drug delivery, sustained drug release from liposomes or polymer implants, and increased targeting of the drug either by chemically modifying the drug or by using tumor-specific carriers. This review will present some of the recent work on targeted drug delivery for brain cancer treatment. PMID- 10699268 TI - Drug solubilization in lung surfactant. AB - The relative affinity of glucocorticosteroids for lung surfactant was determined for the purpose of identifying chemopreventive agents with a high therapeutic index for lung cancer. The aqueous solubility and the extent of solubilization in Survanta, a native extract of bovine lung, of budesonide, triamcinolone acetonide, dexamethasone, and flunisolide were determined as a function of temperature by a dialysis technique. The aqueous solubilites at 37 degrees C were 19.6, 35.8, 104 and 120 microg/ml for the above listed compounds, respectively. The temperature dependence of the solubilities was modest consistent with the hydrophobic properties of the steroids. The amount of drug in solution was significantly enhanced in the presence of Survanta with solubilization ratios of 0. 019, 0.023, 0.014, and 0.02 microg drug dissolved per microg of Survanta phospholipid, respectively. In addition, the extent of solubilization also generally increased with temperature, although the phase transition of the surfactant lipid appeared to complicate the functional relation between temperature and solubilization. These results show that there is enhanced solubilization of glucocortosteroids by lung surfactant which is secreted by the cancer susceptible type II cells of the lung. PMID- 10699269 TI - Oral vaccine delivery. AB - Oral vaccination of animals and man, to provide effective mucosal and/or systemic immunity, is largely ineffective. This is due mainly to the very small quantity of antigen that survives degradation in the intestine and that crosses the intestinal wall. Over the past decade or so, a number of proteins have been identified that are effective at eliciting mucosal and systemic immune responses following oral administration. Uptake of these molecules by the gastro-intestinal tract (GIT) epithelium is dependent upon specific binding to the GIT epithelial cells. The identity of these molecules is discussed, as well as their possible application as 'carriers' for co-transporting haptens, proteins and nanoparticles across the GIT epithelium. PMID- 10699270 TI - Development of predictive pharmacokinetic simulation models for drug discovery. AB - As discovery chemistry produces increased numbers of potential drug compounds, the use of ADME (absorption, distribution, metabolism, and excretion) properties is becoming increasingly important in the drug selection and promotion process. A computer simulation model has been developed and validated to predict ADME outcomes, such as rate of absorption, extent of absorption, etc. using a limited number of in vitro data inputs. The oral bioavailability of ganciclovir in dogs and humans was simulated using a physiologically based model that utilized many biopharmaceutically relevant parameters, such as the concentration of ganciclovir in the duodenum, jejunum, ileum, and colon at various dose levels and solubility values. The simulations were run and compared to dog and human in vivo data. The simulation results demonstrated that the low bioavailability of ganciclovir is limited by compound solubility rather than permeability due to partitioning as previously speculated. This technology provides a breakthrough in in silico prediction of absorption and with its continued development and improvement, will aid drug discovery and development scientists to produce better pharmaceutical products. PMID- 10699271 TI - Molecular aspects of muco- and bioadhesion: tethered structures and site-specific surfaces. AB - Mucoadhesive controlled-release devices can improve the effectiveness of a drug by maintaining the drug concentration between the effective and toxic levels, inhibiting the dilution of the drug in the body fluids, and allowing targeting and localization of a drug at a specific site. Acrylic-based hydrogels have been used extensively as mucoadhesive systems. They are well suited for bioadhesion due to their flexibility and nonabrasive characteristics in the partially swollen state, which reduce damage-causing attrition to the tissues in contact. Crosslinked polymeric devices may be rendered adhesive to the mucosa. For example, adhesive capabilities of these hydrogels can be improved by tethering of long flexible chains to their surfaces. Tethering of long poly(ethylene glycol) (PEG) chains on poly(acrylic acid) hydrogels and their copolymers can be achieved by grafting reactions, or by copolymerization in the presence of several PEG containing acrylates. The ensuing hydrogels exhibit mucoadhesive properties due to enhanced anchoring of the chains with the mucosa. Theoretical calculations can lead to optimization of the tethered structure. Experimental results indicate that the chain interpenetration is a strong function of the PEG molecular weight, the polymer swelling ratio and the mucosa composition. PMID- 10699272 TI - Synthesis and characterization of superporous hydrogel composites. AB - Recently, we synthesized superporous hydrogels which swell fast with high swelling ratios for development of gastric retention devices. Due to their superabsorbent nature, superporous hydrogels are too mechanically weak for gastric retention application. The mechanical strength of superporous hydrogels was substantially increased by making superporous hydrogel composites. The composite materials used were hydrophilic particulate materials commonly used as disintegrants in pharmaceutical tablets. In this study, Ac-Di-Sol was used as a model composite material. Addition of Ac-Di-Sol resulted in significant improvement in the properties of superporous hydrogels. The dried superporous hydrogels maintained interconnected channels even after drying in the air. Thus, the swelling kinetics and the swelling ratio were not affected by air drying, which normally would have resulted in partial or total collapse of the interconnected pores. The presence of Ac-Di-Sol also increased the mechanical strength substantially. Scanning electron microscopic examination showed that the composite material increased the physical crosslinking density which provided high mechanical strength and prevented polymer chains from collapsing during air drying. The superporous hydrogel composites possess properties suitable for gastric retention. PMID- 10699273 TI - Interactions of functionalized dextran-coated liposomes with vascular smooth muscle cells. AB - Synthetic polymers are commonly used in the medical field as implants, polymeric drugs, or drug delivery systems. Among them, bioactive sulfated polysaccharides such as chemically modified dextrans are described to exhibit various properties including the inhibition of smooth muscle cell (SMC) growth. SMCs are key cellular components involved in the physiopathology of the vascular walls especially in atherosclerosis or after vascular surgeries. Interestingly, binding sites on vascular SMCs were already observed for an antiproliferative functionalized dextran (FDx). In this context, we hypothesized that this bioactive polymer could be used as a targeting moiety on the surface of drug delivery systems. In this work, liposomes constituted of phosphatidylcholine, phosphatidylethanolamine and cholesterol (70/10/20 mol.%) were prepared and coated with FDx hydrophobized by a cholesterol anchor (CholFDx) which penetrates the lipid bilayer during the liposome formation. The liposome interactions with SMCs were then followed using radiolabeled liposomes and fluorolabeled liposomes. Results of radioactivity on SMCs indicated higher interactions with CholFDx coated liposomes as compared to uncoated liposomes. The fluorescence of cells incubated with fluorolabeled CholFDx-coated liposomes also evidenced the liposome binding on SMC membranes. These data demonstrated that liposomes coated with FDx interacted with vascular SMCs. Consequently, the coating with such bioactive polymers appears promising for the design of new drug delivery systems for the targeting of vascular cells. PMID- 10699274 TI - Inner core segment design for drug delivery control of thermo-responsive polymeric micelles. AB - Modification of the thermo-responsive behavior of polymeric micelles for specific drug delivery functions was investigated using combinations of micellar inner cores and outer shell polymer chemistries. Polymeric micelles comprised of AB block copolymers of PIPAAm (poly(N-isopropylacrylamide)) with either PBMA (poly(butyl methacrylate)) or PSt (polystyrene) were employed. PIPAAm-PBMA and PIPAAm-PSt block copolymers formed a core-shell micellar structure after dialysis of the block copolymer solutions in organic solvents against water at 20 degrees C. The hydrophobic drug, adriamycin, (ADR) was loaded into the inner core of the polymeric micelles by dialysis. The polymers showed reversible intermicellar dispersion/aggregation in response to temperature cycles through an outer polymer shell lower critical solution temperature (LCST for PIPAAm=32.5 degrees C), observed by DLS (dynamic light scattering) and transmittance measurements. Upon heating above the LCST, PIPAAm-PBMA micelles exhibited an abrupt increase in micropolarity and an abrupt decrease in microrigidity sensed by pyrene and 1, 3 bis(1-pyrenyl)propane (PC(3)P), respectively. In contrast, PIPAAm-PSt micelles maintained constant values with lower micropolarity and higher microrigidity than those of PIPAAm-PBMA micelles over the temperature range 20 to 40 degrees C. From these results, structural deformations produced by outer shell polymer structural change with temperature cycles through the LCST are proposed for the PBMA core possessing a lower T(g) (ca. 20 degrees C) than the outer shell PIPAAm LCST. The PSt core with a much higher T(g) (ca. 100 degrees C) than the outer shell LCST retained its structure, regardless of outer shell changes. PIPAAm-PBMA micelles released ADR only when heated above the LCST, while PIPAAm-PSt micelles did not. Cell cultures treated with PIPAAm-PBMA micelles loaded with ADR showed high in vitro cytotoxicity when heated above the LCST, while PIPAAm-PSt micelles loaded with ADR expressed very low in vitro cytotoxicity irrespective of temperature change through the LCST. The nature of hydrophobic segments comprising the micelle inner core offers an important control point for thermo-responsive drug release and the drug activity of the thermo-responsive polymeric micelle. PMID- 10699275 TI - Polymer-bound camptothecin: initial biodistribution and antitumour activity studies. AB - Camptothecin (CPT) is a potent, antitumour drug acting mainly through inhibition of topoisomerase I during the S-phase of the cell cycle. Despite its impressive antitumour activity, clinical development was halted for unpredictable toxic events. Two soluble N-(2-hydroxypropyl) methacrylamide (HPMA) copolymers were synthesised to contain CPT (5 wt.% and 10 wt.%). CPT was covalently linked at its alpha-hydroxyl group to the polymers through a Gly-Phe-Leu-Gly- spacer. In-vitro, CPT-conjugates were fairly resistant to hydrolysis in plasma as in buffer at neutral pH (0.2-0. 4% free CPT/h), while elastase and cysteine-proteases were able to release the active drug. Plasma levels in mice after intravenous administration of CPT-conjugates confirmed the modest hydrolysis in plasma. Plasma levels were approximately 5-fold lower than those observed at the highest tolerated dose of CPT administered in classical vehicles. Biodistribution in HT29 human colon carcinoma bearing mice was carried out after i.v. injection of [3H]CPT-conjugate and free [3H]CPT. Radioactivity uptake in tumour was evident only after [3H]CPT-conjugate treatment. Repeated intravenous administration of CPT-conjugates to HT29-bearing mice gave more than 90% tumour inhibition, some complete tumour regressions and no toxic deaths. The improved pharmacological profile on HT29 human colon carcinoma xenografts of the first poly(HPMA)-CPT conjugates might be ascribed to their prolonged intra-tumour retention and sustained release of the active drug. PMID- 10699276 TI - Water-soluble dendritic unimolecular micelles: their potential as drug delivery agents. AB - To explore the potential of dendritic unimolecular micelles in drug delivery systems, dendritic unimolecular micelles with a hydrophobic core surrounded by a hydrophilic shell were prepared by coupling dendritic hypercores with poly(ethylene glycol) [PEG] mesylate. The monomer selected to build the dendritic cores was 4, 4-bis(4'-hydroxyphenyl) pentanol as this large monomer unit provides flexibility to the dendritic structure while contributing to the "container" capacity of the overall structure. Four generations of dendritic hypercores with six, 12, 24, and 48 phenolic end groups were prepared. Subsequent coupling reactions with PEG mesylate afforded four generations of dendritic unimolecular micelles. The micelles were characterized by Matrix Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry (MALDI-TOF MS), 1H-NMR and Gel Permeation Chromatography (GPC). The "container" property of micelles was demonstrated by solubilizing pyrene in aqueous solution. Entrapment of the model drug indomethacin in the dendritic micelles was achieved at 11 wt.% loading level, and preliminary in vitro release tests showed that sustained release characteristics were achieved. PMID- 10699277 TI - Dendrimers: relationship between structure and biocompatibility in vitro, and preliminary studies on the biodistribution of 125I-labelled polyamidoamine dendrimers in vivo. AB - Dendrimers are highly branched macromolecules of low polydispersity that provide many exciting opportunities for design of novel drug-carriers, gene delivery systems and imaging agents. They hold promise in tissue targeting applications, controlled drug release and moreover, their interesting nanoscopic architecture might allow easier passage across biological barriers by transcytosis. However, from the vast array of structures currently emerging from synthetic chemistry it is essential to design molecules that have real potential for in vivo biological use. Here, polyamidoamine (PAMAM, Starburst), poly(propyleneimine) with either diaminobutane or diaminoethane as core, and poly(ethylene oxide) (PEO) grafted carbosilane (CSi-PEO) dendrimers were used to study systematically the effect of dendrimer generation and surface functionality on biological properties in vitro. Generally, dendrimers bearing -NH(2) termini displayed concentration- and in the case of PAMAM dendrimers generation-dependent haemolysis, and changes in red cell morphology were observed after 1 h even at low concentrations (10 microg/ml). At concentrations below 1 mg/ml CSi-PEO dendrimers and those dendrimers with carboxylate (COONa) terminal groups were neither haemolytic nor cytotoxic towards a panel of cell lines in vitro. In general, cationic dendrimers were cytotoxic (72 h incubation), displaying IC(50) values=50-300 microg/ml dependent on dendrimer-type, cell-type and generation. Preliminary studies with polyether dendrimers prepared by the convergent route showed that dendrimers with carboxylate and malonate surfaces were not haemolytic at 1 h, but after 24 h, unlike anionic PAMAM dendrimers they were lytic. Cationic 125I-labelled PAMAM dendrimers (gen 3 and 4) administered intravenously (i.v.) to Wistar rats ( approximately 10 microg/ml) were cleared rapidly from the circulation (<2% recovered dose in blood at 1 h). Anionic PAMAM dendrimers (gen 2.5, 3.5 and 5.5) showed longer circulation times ( approximately 20-40% recovered dose in blood at 1 h) with generation-dependent clearance rates; lower generations circulated longer. For both anionic and cationic species blood levels at 1 h correlated with the extent of liver capture observed (30-90% recovered dose at 1 h). 125I Labelled PAMAM dendrimers injected intraperitoneally were transferred to the bloodstream within an hour and their subsequent biodistribution mirrored that seen following i.v. injection. Inherent toxicity would suggest it unlikely that higher generation cationic dendrimers will be suitable for parenteral administration, especially if they are to be used at a high dose. In addition it is clear that dendrimer structure must also be carefully tailored to avoid rapid hepatic uptake if targeting elsewhere (e.g. tumour targeting) is a primary objective. PMID- 10699278 TI - DNA delivery systems based on complexes of DNA with synthetic polycations and their copolymers. AB - Block and graft copolymers of N-(2-hydroxypropyl)methacrylamide (HPMA) with 2 (trimethylammonio)ethyl methacrylate were synthesised and used for preparation of polyelectrolyte complexes with calf thymus DNA intended for targeted delivery of genes in vivo. In this study the effects of the speed of component mixing, total concentration of polymers, ionic strength of solvents, copolymer structure and content of HPMA in the copolymers on parameters of the polyelectrolyte complexes was investigated. Static and dynamic light scattering methods were used as a main tool for characterising these complexes. The presence of HPMA units in the polycation had no significant effect on its ability to form complexes with DNA, but did affect molecular parameters and aggregation (precipitation) of the complexes. The size of the complexes increases whereas their molecular weight decreases with increasing content of HPMA units. The density of the complexes decreases with increasing HPMA content independently of the copolymer structure. In order to prepare stable DNA complexes containing single DNA molecule, the following rules should be observed: (1) copolymers should have a content of HPMA units higher than 40%; (2) the DNA concentrations in solutions should be kept below 4 x 10(-5) g/ml and (3) both components should be mixed together in deionized water. The stability of the complexes against precipitation in 0.15 M NaCl and the resistance of the complexed DNA to the action of nucleases was also studied. Whereas DNA complexes of all copolymers showed very good nuclease stability, the presence of a sufficiently high content of HPMA is necessary for their good stability in 0.15 M NaCl. The investigation of the stability and the interaction of DNA complexes in aqueous solutions of serum albumin and dilute human blood serum revealed adsorption of biomacromolecules on DNA complexes accompanied by significant changes in the zeta-potential which finally resulted in formation of a "protein layer" and in undesirable precipitation of DNA complexes. In in vitro transfection experiments, the transfection efficiency of DNA complexes with copolymers was always higher than that of the cationic homopolymer slightly increasing with increasing content of HPMA in the copolymers but being about 10-100-times lower than the complexes DNA-poly(L-lysine. In the cytoplasmic injections, it was observed that DNA complexes produced greater gene expression than a direct microinjection of free DNA. The block copolymer complexes were also found to be more efficient than the corresponding simple polycation complexes. In the nuclear microinjection, precisely the opposite behaviour was observed. PMID- 10699279 TI - Making microencapsulation work: conformal coating, immobilization gels and in vivo performance. AB - Microencapsulation of cells as a means of insulin or other protein delivery (for example, for gene therapy) has not yet realized its potential. Three aspects of this problem are illustrated with reference to the use of poly(hydroxyethyl methacrylate-co-methyl methacrylate) (HEMA-MMA). Conformal coating was used to coat cell aggregates with a very thin layer of a water-insoluble HEMA-MMA membrane that conforms to the shape of the aggregate, and minimizes the polymer's contribution to the total transplant volume. Cell aggregates were coated at a liquid-liquid interface of a discontinuous density gradient composed of both aqueous and organic liquids. Aggregates of HepG2 cells were coated and remained viable. Immobilization matrices were co-encapsulated in order to control cell phenotype. Ultralow gelling temperature agarose promoted the proliferation of HEK293 cells, while the viability of transfected C2C12 cells was improved in microcapsules that contained Matrigel. Rat or human hepatoma cells in HEMA-MMA microcapsules lost viability within a week after implantation into an omental pouch in Wistar rats. The loss of viability was attributed to the tissue reaction, although it is not clear if the cells lost their viability in vivo leading to the aggressive tissue reaction or if the latter caused the cells to starve or otherwise die. On the other hand, intraperitoneal implantation of microcapsules containing L929 cells in 'syngeneic' C3H mice in a high-strength agarose gel resulted in maintenance of viability of approximately 50% of the encapsulated cells. While progress is being made on several fronts, this type of tissue engineering construct is still several years away from routine use in humans. PMID- 10699280 TI - Poly-L-glutamic acid derivatives as vectors for gene therapy. AB - This paper describes the synthesis and evaluation of biodegradable derivatives of poly-L-glutamic acid as suitable vectors for gene therapy. When mixed with DNA the new polymers self assemble and form polyelectrolyte complexes. The formation of the complexes and determination of their stability towards disruption by serum albumin was monitored by Ethidium bromide (EtBr) fluorescence spectroscopy. All polymers were able to form complexes and their size, determined by photon correlation spectroscopy, was between 84.5+/-2 nm and 96. 7+/-1.6 nm, depending on the type of polymer and the charge ratio. All complexes were stable towards serum albumin. The results from the biodegradability tests, using tritosomes, show that the polymers are biodegradable and the rate of degradation is influenced by the number of charged groups in the side chains. Haemolysis and red blood cell (RBC) agglutination were assessed and compared to poly(L-lysine) (pLL) and polyethyleneimine (pEI). RBC agglutination was monitored with optical microscopy. Results show that the new polymers are less toxic than pLL and pEI. Preliminary transfection studies show that the polymers are suitable vectors for gene delivery. PMID- 10699281 TI - Molecular engineering of proteins and polymers for targeting and intracellular delivery of therapeutics. AB - There are many protein and DNA based therapeutics under development in the biotechnology and pharmaceutical industries. Key delivery challenges remain before many of these biomolecular therapeutics reach the clinic. Two important barriers are the effective targeting of drugs to specific tissues and cells and the subsequent intracellular delivery to appropriate cellular compartments. In this review, we summarize protein engineering work aimed at improving the stability and refolding efficiency of antibody fragments used in targeting, and at constructing new streptavidin variants which may offer improved performance in pre-targeting delivery strategies. In addition, we review recent work with pH responsive polymers that mimic the membrane disruptive properties of viruses and toxins. These polymers could serve as alternatives to fusogenic peptides in gene therapy formulations and to enhance the intracellular delivery of protein therapeutics that function in the cytoplasm. PMID- 10699282 TI - Nanoparticulate delivery system of a tyrphostin for the treatment of restenosis. AB - Restenosis, the principal complication of percutaneous transluminal coronary angioplasty is responsible for the 35-40% long-term failure rate following coronary revascularization. The neointimal formation, a morphological substrate of restenosis, is dependent on smooth muscle cells (SMC) proliferation and migration. Signal transduction through the platelet-derived growth factor (PDGF)/PDGF receptors system is involved in the process of post-angioplasty restenosis. The unsuccessful attempts to control restenosis by systemic pharmacological interventions have prompted many researchers to look for more promising therapeutic approaches such as local drug delivery. Tyrphostins are low molecular weight inhibitors of protein tyrosine kinases. We assessed the release kinetics and in vivo effects of nanoparticles containing PDGF-Receptor beta (PDGFRbeta) tyrphostin inhibitor, AG-1295. AG-1295-loaded poly(DL-lactide) (PLA) nanoparticles were prepared by spontaneous emulsification/solvent displacement technique. In vitro release rate and the impact of drug/polymer ratio on the nanoparticle size were determined. The degree of tyrosine phosphorylation was assessed by Western blot with phosphotyrosine-specific antibody in rat SMC extracts. Several bands characteristic of PDGF BB-stimulated SMC disappeared or weakened following tyrphostin treatment. Local intraluminal delivery of AG-1295 loaded PLA nanoparticles to the injured rat carotid artery had no effect on proliferative activity in medial and neointimal compartments of angioplastisized arteries, indicating a primary antimigration effect of AG-1295 on medial SMC. PMID- 10699283 TI - Virtual screening of intestinal drug permeability. AB - Lead compounds generated in high throughput drug discovery programmes often have unfavorable biopharmaceutical properties, resulting in a low success rate of such drug candidates in clinical development. Drug companies and researchers would thus like to have methods of predicting biopharmaceutical properties accurately. The intestinal permeability to a lead compound is one such property which is particularly important. Therefore, access to methods to accurately predict biopharmaceutical properties, such as the intestinal permeability of a large series of compounds, is of particular importance. This review deals with new theoretical methods used to predict intestinal drug permeability. There are several possible transport routes across the intestine, but theoretical methods generally deal with only one of them, the passive transcellular route. Therefore, this review will also discuss the relative importance of passive and active drug transport and efflux routes using recent data generated in cell cultures, animal models and human subjects. PMID- 10699284 TI - A coumarin-based prodrug strategy to improve the oral absorption of RGD peptidomimetics. AB - In recent years, major progress has been made in the design and synthesis of fibrinogen antagonists, which are peptidomimetic Arg-Gly-Asp (RGD) analogs. These RGD analogs are very promising antiplatelet agents. However, the clinical development of orally active RGD analogs has been hindered by the low oral bioavailability of many such RGD analogs. Aimed at enhancing their oral bioavailability, we have synthesized several coumarin-based cyclic prodrugs of RGD analogs, which have the two most polar functional groups, a carboxyl and an amino group, masked as an ester and an amide, respectively. As expected, these cyclic prodrugs have higher membrane interaction potentials as estimated by determining their partitioning between aqueous buffer and an immobilized artificial membrane than the corresponding RGD analogs. Consequently, these cyclic prodrugs are 5-6-fold more able to permeate monolayers of Caco-2 cells, an in vitro cell culture model of the intestinal mucosa barrier. Preliminary studies using dog also indicate the promising potential of using this coumarin-based prodrug strategy to improve the oral bioavailability of such RGD analogs. PMID- 10699285 TI - Oral uptake and translocation of a polylysine dendrimer with a lipid surface. AB - A series of lipidic peptide dendrimers based on lysine with 16 surface alkyl (C(12)) chains has been synthesised in our laboratories. One of the series, a fourth generation dendrimer with a diameter of 2.5 nm was chosen to study its absorption after oral administration to female Sprague-Dawley rats (180 g, 9 weeks old). It was synthesised as the tritiated derivative (all acetyl portions) and had a molecular weight of 6300 and log P (octanol/water) of 1.24. First a single oral dose 14 mg/kg was administered by gavage. Maximum levels of dendrimer observed were 15% in the small intestine, 5% in the large intestine and 3% in the blood at 6 h after administration, while 1.5% reached the liver, 0.1% the spleen and 0. 5% the kidneys. In a parallel study with a higher dose of 28 mg/kg, approximately 1% was absorbed via Peyer's patches of the small intestine at 3 h. The maximum uptake by small intestine enterocytes was 4% of the dose after 3 h. After 12 h, 0.3 and 4% dendrimer was measured respectively in Peyer's patches and enterocytes of the large intestine. When calculated on the basis of target tissue weight, the total percentage of the dose absorbed through Peyer's patches was greater than through normal enterocytes in the small intestine after 3 and 24 h, but the opposite was true in the large intestine. These levels of uptake and translocation are lower than those exhibited by polystyrene particles in the range from 50 to 3000 nm. This might suggest that there is an optimum size for nanoparticulate uptake by the gut. PMID- 10699286 TI - Nanosphere based oral insulin delivery. AB - Zinc insulin is successfully encapsulated in various polyester and polyanhydride nanosphere formulations using Phase Inversion Nanoencapsulation (PIN). The encapsulated insulin maintains its biological activity and is released from the nanospheres over a span of approximately 6 h. A specific formulation, 1.6% zinc insulin in poly(lactide-co-glycolide) (PLGA) with fumaric anhydride oligimer and iron oxide additives has been shown to be active orally. This formulation is shown to have 11.4% of the efficacy of intraperitoneally delivered zinc insulin and is able to control plasma glucose levels when faced with a simultaneously administered glucose challenge. A number of properties of this formulation, including size, release kinetics, bioadhesiveness and ability to traverse the gastrointestinal epithelium, are likely to contribute to its oral efficacy. PMID- 10699287 TI - Tumor vascular permeability and the EPR effect in macromolecular therapeutics: a review. AB - Most solid tumors possess unique pathophysiological characteristics that are not observed in normal tissues or organs, such as extensive angiogenesis and hence hypervasculature, defective vascular architecture, impaired lymphatic drainage/recovery system, and greatly increased production of a number of permeability mediators. The phenomenon now known as the enhanced permeability and retention (EPR) effect for lipid and macromolecular agents has been observed to be universal in solid tumors. Primarily, enhanced vascular permeability will sustain an adequate supply of nutrients and oxygen for rapid tumor growth. The EPR effect also provides a great opportunity for more selective targeting of lipid- or polymer-conjugated anticancer drugs, such as SMANCS and PK-1, to the tumor. In the present review, the basic characteristics of the EPR effect, particularly the factors involved, are described, as well as its modulation for improving delivery of macromolecular drugs to the tumor. Tumor-specific vascular physiology is also described. PMID- 10699288 TI - Development of microspheres for neurological disorders: from basics to clinical applications. AB - Drug delivery to the central nervous system remains a challenging area of investigation for both basic and clinical neuroscientists. Numerous drugs are generally excluded from blood to brain transfer due to the negligible permeability of the brain capillary endothelial wall, which makes up the blood brain barrier in vivo. For several years, we have explored the potential applications of the microencapsulation of therapeutic agents to provide local controlled drug release in the central nervous system. Due to their size, these microparticles can be easily implanted by stereotaxy in discreet, precise and functional areas of the brain without damaging the surrounding tissue. This type of implantation avoids the inconvenient insertion of large implants by open surgery and can be repeated if necessary. We have established the compatibility of poly(lactide-co-glycolide) microspheres with brain tissues. Presently, the most developed applications concern Neurology and Neuro-oncology, with local delivery of neurotrophic factors and antimitotic drugs into neurodegenerative lesions and brain tumours, respectively. The drugs that had been encapsulated by our group included nerve growth factor (NGF), 5-fluorouracil (5-FU), idoxuridine and BCNU. Preclinical studies have been performed with each drug. Studies with NGF are reported as an example. A phase I/II clinical trial has been carried out in patients with newly diagnosed glioblastomas to assess the potentialities of 5 FU-loaded microspheres when intracranially implanted. PMID- 10699289 TI - Epithelial application of Pseudomonas aeruginosa exotoxin A results in a selective targeting to cells in the liver, spleen and lymph node. AB - Pseudomonas aeruginosa exotoxin A (PE) is a 67-kDa protein expressed under the selective pressure of a low iron environment. Previous studies using non-toxic PE chimeras containing a viral surface antigen, the V3 loop of MN gp120 from human immunodeficiency virus type 1 (HIV-1), resulted in not only an effective mucosal immunization but also a striking systemic immune response following epithelial application. Presently, we have examined the possibility that such a strong dual immune response was generated by the efficient targeting of critical cells of the immune system. Mice were dosed with 10 microg of toxic PE or a non-toxic mutant of PE (ntPE) by intratracheal instillation. Examination of lung, liver and spleen tissues isolated 4, 8 and 12 h following intratracheal instillation with PE demonstrated specific cell damage in these tissues which was not observed in mice dosed with ntPE. Based upon the location and characteristics of observed responses, the cells targeted by PE appear to be involved in the antigen presentation arm of the immune response. Since ntPE chimeras with inserted peptide antigen epitopes from a wide variety of pathogens are easy to prepare and administer, these results support this approach for mucosal immunization. PMID- 10699290 TI - New clinical applications of transdermal testosterone delivery in men and women. AB - This paper reviews recent progress in the development and clinical application of testosterone transdermal delivery systems designed for physiological replacement therapy in men and women. The biopharmaceutic goal of physiologic replacement therapy is to produce serum levels and circadian patterns of testosterone and its active metabolites that mimic the normal physiology of testosterone in the particular target population. For the treatment of adult hypogonadal men, the nightly 24 h application of the Androderm testosterone transdermal system (5 mg per day) achieves this goal - as demonstrated in a series of clinical pharmacokinetic studies. For the treatment of adolescent males, physiologic replacement can be approximated by modifying the dose and duration of Androderm application so as to mimic the patterns of nocturnal testosterone secretion observed during puberty. With the objective of providing physiological replacement for women with diminished testosterone production, an experimental testosterone matrix transdermal system (TMTDS) has been developed and is currently undergoing clinical evaluation. In parallel with the development of testosterone transdermal systems, physicians have been investigating a number of conditions, in both males and females, where testosterone production is diminished and replacement therapy may be beneficial. Three of these new clinical applications will be illustrated - the use of Androderm for the treatment of adolescent males with beta-thalassemia, the use of Androderm for the treatment of HIV+ men, and the use of the TMTDS for the treatment of HIV+ women. From the biopharmaceutic and clinical perspectives, the development of testosterone transdermal systems represents an important achievement in controlled drug delivery. PMID- 10699291 TI - Preface PMID- 10699292 TI - The effect of egg albumin on the crystalline properties of carbamazepine in sustained release hydrophilic matrix tablets and in aqueous solutions. AB - The influence of egg albumin (EA) on the crystal habit properties of carbamazepine (CBZ) in aqueous solutions, solid-state, and in sustained release matrix tablets was investigated using differential scanning calorimetry (DSC), hot-stage microscopy (HSM), X-ray powder diffraction (XRD), scanning electron microscopy (SEM) and contact angle goniometer (CAG). The results suggest that in solid-state mixtures, EA affected the polymorphic transitions of CBZ from the beta to the alpha form. In hydrated matrix tablets and aqueous solutions, EA influenced the conversion rate of CBZ to the dihydrate form depending on EA concentration. It was found that increasing EA concentration enhanced CBZ dihydrate aggregation, an effect that leads to the formation of crystals with high mechanical strength and decrease of CBZ solubility. Possible mechanisms, which explain crystal growth and aggregation, as well as alteration of CBZ polymorphic transitions in the solid-state, gel layer, and in aqueous solution were suggested. In the gel layer of hydrated tablets the kinetics of CBZ transformation to the dihydrate form, crystal growth and aggregation were influenced by various processes: matrix hydration, erosion mechanism and the formation of metastable conditions, which favor aggregation and growth. The release kinetics of CBZ from the matrix highly correlated with the crystalline and morphological changes occurring in the matrix. PMID- 10699293 TI - Poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide)/poly(epsilon caprolactone) (PCL) amphiphilic block copolymeric nanospheres. II. Thermo responsive drug release behaviors. AB - Amphiphilic block copolymers composed of relatively hydrophilic PEO-PPO-PEO block copolymer (Pluronic) and poly (epsilon-caprolactone) with hydrophobic character were synthesized by ring-opening polymerization of epsilon-caprolactone in the presence of PEO-PPO-PEO block copolymer using stannous octoate as a catalyst. Pluronic/PCL block copolymeric nanospheres with core-shell structure were prepared by dialysis method. They showed the average diameter of 116-196 nm depending on the type of copolymer. All the nanosphere samples exhibited a narrow size distribution. The critical micelle concentrations of Pluronic/PCL amphiphilic block copolymers determined by fluorescence spectroscopy were lower than that of the common low molecular weight surfactant. Their core-shell structure was confirmed by 1H NMR spectroscopy. Pluronic/PCL block copolymeric nanospheres exhibited the reversible change of size depending on the temperature. Release behaviors of indomethacin from Pluronic/PCL block copolymeric nanospheres also showed temperature dependence and a sustained release pattern. In addition, cytotoxicity test using an MTT assay method revealed that these indomethacin loaded Pluronic/PCL nanospheres could remarkably reduce the cell damage compared with the unloaded free indomethacin. PMID- 10699294 TI - A vehicle for photodynamic therapy of skin cancer: influence of dimethylsulphoxide on 5-aminolevulinic acid in vitro cutaneous permeation and in vivo protoporphyrin IX accumulation determined by confocal microscopy. AB - Topical application of 5-aminolevulinic acid (5-ALA) followed by light irradiation is a new concept of photodynamic therapy (PDT) of skin cancers. 5-ALA is a prodrug that can be converted by the heme biosynthetic pathway into protoporphyrin IX, an effective photosensitizer. In the present work we propose the enhancement of 5-ALA-induced protoporphyrin IX accumulation by dimethylsulphoxide (DMSO) and ethylenediamine-tetraacetic acid disodium salt (EDTA). The presence of 20% DMSO (w/w) in oil-in-water emulsions increased the in vitro permeation of 5-ALA through hairless mouse skin. In vivo studies demonstrated a significant increase in the amount of protoporphyrin IX extracted from healthy hairless mouse skin after 3 h treatment with an oil-in-water emulsion containing 10% 5-ALA (w/w), 3% EDTA (w/w) and 20% DMSO (w/w). By confocal scanning laser microscopy imaging, an observed increase in red fluorescence, at 476 nm excitation and emission detected longer than 590 nm, in skin that had received this treatment, was attributed to protoporphyrin IX accumulation. Although no effect of EDTA on short-term protoporphyrin IX accumulation in skin was detected, this chelator could protect 5-ALA from decomposition during prolonged topical administration. The results obtained indicate that association of 5-ALA, EDTA and 20% DMSO may enhance the delivery of 5-ALA to the skin in the topical PDT. PMID- 10699295 TI - Development of a poly(ortho ester) prototype with a latent acid in the polymer backbone for 5-fluorouracil delivery. AB - A study has been carried out to determine whether the latest family of poly(ortho esters) can be converted into a practical delivery system. This polymer differs from the previously described polymers in that it incorporates a short segment of a latent acid in the polymer backbone. The following issues were specifically addressed: (a) can the erosion and drug release be reproducibly controlled to yield the desired drug release kinetics and erosion rates? (b) Is the polymer stable during radiation sterilization, on storage and on fabrication? (c) Can the polymer be prepared reproducibly at the desired molecular weights and molecular weight distribution? (d) Is the polymer safe for its intended application and does the in vivo erosion proceed to completion? (e) Can the polymer be easily fabricated into desired configurations? Studies have shown that if the synthesis is carefully controlled, the desired molecular weights can be reproducibly prepared, that the polymer is reasonably stable after irradiation at 24 kGy and during storage at room temperature under anhydrous conditions, and that it can be safely thermally fabricated at temperatures in the neighborhood of 120 degrees C. When polymer devices were implanted intraperitoneally in rats the polymer eroded to completion without any overt toxicity as determined by the measured parameters. PMID- 10699296 TI - Controlled release of beta-estradiol from PLAGA microparticles: the effect of organic phase solvent on encapsulation and release. AB - To determine the effect of the organic solvent used during microparticle preparation on the in vitro release of beta-estradiol, a number of formulations were evaluated in terms of size, shape and drug delivery performance. Biodegradable microparticles of poly(lactide-co-glycolide) were prepared containing beta-estradiol that utilized dichloromethane, ethyl acetate or a mixture of dichloromethane and methanol as the organic phase solvent during the particle preparation. The drug delivery behavior from the microparticles was studied and comparisons were made of their physical properties for different formulations. The varying solubilities of beta-estradiol and poly(lactide-co glycolide) in the solvents studied resulted in biodegradable microparticles with very different physical characteristics. Microparticles prepared from solid suspensions of beta-estradiol using dichloromethane as the organic phase solvent were similar in appearance to microparticles prepared without drug. Microparticles prepared from dichloromethane/methanol solutions appeared transparent to translucent depending on the initial amount of drug used in the formulation. Microparticles prepared using ethyl acetate appeared to have the most homogeneous encapsulation of beta-estradiol, appearing as solid white spheres regardless of initial drug content. Studies showed that microparticles prepared from either ethyl acetate or a mixture of dichloromethane and methanol gave a more constant release profile of beta-estradiol than particles prepared using dichloromethane alone. For all formulations, an initial burst of release increased with increasing drug loading, regardless of the organic solvent used. PMID- 10699297 TI - Development of fibrin derivatives for controlled release of heparin-binding growth factors. AB - The goal of this work was to develop a growth factor delivery system for use in wound healing that would provide localized release of heparin-binding growth factors in a biomimetic manner, such that release occurs primarily in response to cell-associated enzymatic activity during healing. A key element of the drug delivery system was a bi-domain peptide with an N-terminal transglutaminase substrate and a C-terminal heparin-binding domain, based on antithrombin III. The bi-domain peptide was covalently cross-linked to fibrin matrices during coagulation by the transglutaminase activity of factor XIIIa and served to immobilize heparin electrostatically to the matrix, which in turn immobilized the heparin-binding growth factor and slowed its passive release from the matrix. Basic fibroblast growth factor (bFGF) was considered as an example of a heparin binding growth factor, and cell culture experimentation was performed in the context of peripheral nerve regeneration. A mathematical model was developed to determine the conditions where passive release of bFGF would be slow, such that active release could dominate. These conditions were tested in an assay of neurite extension from dorsal root ganglia to determine the ability of the delivery system to release bioactive growth factor in response to cell-mediated processes. The results demonstrated that bFGF, immobilized within fibrin containing a 500-fold molar excess of immobilized heparin relative to bFGF, enhanced neurite extension by up to about 100% relative to unmodified fibrin. A variety of control experiments demonstrate that all components of the release system are necessary and that the bi-domain peptide must be covalently bound to the fibrin matrix. The results thus suggest that these matrices could serve as therapeutic materials to enhance peripheral nerve regeneration through nerve guide tubes and may have more general usefulness in tissue engineering. PMID- 10699298 TI - Ethosomes - novel vesicular carriers for enhanced delivery: characterization and skin penetration properties. AB - This work describes a novel carrier for enhanced skin delivery, the ethosomal system, which is composed of phospholipid, ethanol and water. Ethosomal systems were much more efficient at delivering a fluorescent probe to the skin in terms of quantity and depth, than either liposomes or hydroalcoholic solution. The ethosomal system dramatically enhanced the skin permeation of minoxidil in vitro compared with either ethanolic or hydroethanolic solution or phospholipid ethanolic micellar solution of minoxidil. In addition, the transdermal delivery of testosterone from an ethosomal patch was greater both in vitro and in vivo than from commercially available patches. Skin permeation of ethosomal components, ethanol and phospholipid, was demonstrated in diffusion-cell experiments. Ethosomal systems composed of soy phosphatidylcholine 2%, ethanol 30% and water were shown by electron microscopy to contain multilamellar vesicles. 31P-NMR studies confirmed the bilayer configuration of the lipids. Calorimetry and fluorescence measurements suggested that the vesicular bilayers are flexible, having a relatively low T(m) and fluorescence anisotropy compared with liposomes obtained in the absence of ethanol. Dynamic light scattering measurements indicated that ethanol imparted a negative charge to the vesicles. The average vesicle size, as measured by dynamic light scattering, was modulated by altering the ethosome composition. Experiments using fluorescent probes and ultracentrifugation showed that the ethosomes had a high entrapment capacity for molecules of various lyophilicities. PMID- 10699299 TI - Modified drug release from inert matrix tablets prepared from formulations of identical composition but different organisations. AB - This paper develops for the first time a concept to modify the release rate of a fixed formulation by changing only the organisation of the mix used to prepare the tablets (ordered mixing). To estimate the influence of the organisation of binary mixes, several mixes of ethylcellulose and niflumic acid of the same composition but different organisation were compacted. The tablet surfaces were examined by energy dispersive X-ray microanalysis before the release experiments. Finally, the cross-sections of the remaining matrix were examined by scanning electronic microscopy. Excipient-excipient and excipient-drug interactions are the major factors influencing the drug release rate from the tablets. In the case of interacting materials, the initial release behaviour depends on the tablet surface presented to the dissolution media. The dissolution properties of the tablets are governed by the percolating material. When the inert excipient is percolating, the release rate increases linearly with the excipient/drug size ratio, whereas when the drug is the only material percolating through the system, its release rate is independent of the size ratio. When both materials are percolating through the system, the release rate is independent of the component particle sizes. PMID- 10699300 TI - The preparation and evaluation of poly(epsilon-caprolactone) microparticles containing both a lipophilic and a hydrophilic drug. AB - An original dosage form for oral delivery based on the encapsulation of both, lipophilic and hydrophilic drugs, in poly(epsilon-caprolactone) (PCL) microparticles prepared either by the oil-in-water (o/w) or the water-in-oil-in water (w/o/w) solvent evaporation method was developed. Microparticles were characterized in terms of morphology, size, encapsulation efficiency and drug release. The physical state of the drugs and the polymer was determined by scanning electron microscopy (SEM), X-ray powder diffractometry, and differential scanning calorimetry (DSC). Nifedipine (calcium antagonist) and propranolol HCl (beta-blocker), used for the treatment of hypertension, were chosen as lipophilic and hydrophilic drugs. The microparticles were spherical with diameters in the range of 191-351 microm by the o/w-method, and in the range of 302-477 microm by the w/o/w-method. The encapsulation efficiency (EE) was 91% for nifedipine and 37% for propranolol HCl with the o/w-method, and 83% for nifedipine and 57% for propranolol HCl with the w/o/w-method. DSC and X-ray diffraction studies showed that PCL maintained its semi-crystalline structure, while the drugs were either dispersed or dissolved in the polymer. In vitro release studies revealed a controlled release of nifedipine and propranolol HCl from microparticles prepared by the o/w-method; a burst release of propranolol HCl was observed from microparticles prepared by the w/o/w-method. In conclusion, microparticles containing both a hydrophilic and a lipophilic drug were successfully prepared. PMID- 10699301 TI - Introduction and overview of noninvasive drug monitoring. PMID- 10699302 TI - Physics and instrumentation for imaging in-vivo drug distribution. AB - Several imaging methods are currently available to measure drugs noninvasively. Of these, two techniques are today central to such measurements: nuclear imaging and magnetic resonance imaging/spectroscopy (MRI and MRS). While other methods, such as optical techniques, are rapidly gaining in interest, they have not yet attained the degree of development that makes them effective in measuring drugs in living systems, except in a small number of examples. The following introduction provides some basic elements of the potential and the limitations of both nuclear imaging and MRI/MRS techniques, methods that will be used in the studies described in the articles in this issue. However, and for those desiring to gain a better understanding of both methods, the reader is advised to consult much more extensive reviews and books describing such methods. A suggested list of books and articles on Nuclear Imaging and MRI/MRS is given. PMID- 10699303 TI - In vivo monitoring of drugs using radiotracer techniques. AB - There is an increasing realization of the role of non-invasive monitoring of drug pharmacology. In this review, we discuss the role of positron emission tomography in such monitoring of tumour and normal tissue drug pharmacokinetics as well as assessment of tumour response, drug-receptor interactions and mechanisms of drug action and resistance. These studies represent a multidisciplinary research effort involving radiochemists, imaging scientists, clinicians, pharmacologists and mathematical modellers. This review evaluates achievements in the field from assessment of commonly used therapeutic agents such as 5-fluorouracil to target specific molecules such as markers for gene expression. It is envisaged that application of this technology will facilitate rational drug design and rapid translation of new ideas to the bedside. PMID- 10699304 TI - Nuclear imaging methods for non-invasive drug monitoring. AB - Functional imaging techniques provide complimentary information to that provided by structural studies such as MRI and CT. Functional imaging is based upon known parameters such as physiology, metabolism, biochemistry, pharmacology, and any other biological process. As such, this methodology plays a major role in understanding the basic mechanisms of a multitude of disorders, accurate diagnosis of certain diseases, and developing effective treatment for serious illnesses such as cancer and central nervous system maladies. Although this type of imaging can be performed with various modalities, nuclear procedures have played the leading role in this discipline. Advances made in labeling various radionuclides to biologically important compounds, and development of sophisticated instruments have substantially contributed to the growth of the field of functional imaging. The introduction of positron emission topography (PET), which is based on imaging of compounds labeled with elements such as carbon, nitrogen, and fluorine, has added a major dimension to the evolution of the discipline. This review deals with a brief introduction to the methodologies utilized with radiolabeled tracers and then deals with specific applications of this technology. These applications include assessment of blood flow and metabolism, receptor imaging, elucidating the pathophysiologic process, evaluating role of labeled therapeutic agents, and the potential of these techniques in the development of novel biologic therapies. Functional imaging with radiolabeled tracers will play an increasingly important role in modern medicine, and its impact will be substantial in the management of patients with various disorders. PMID- 10699305 TI - 19F-MRS studies of fluorinated drugs in humans. AB - The use of 19F-NMR as a noninvasive probe to measure directly the pharmacokinetics of drugs at their target (effector) site(s) is illustrated in this article by human studies with 5-fluorouracil (5-FU). This drug, and several of its metabolites, have been measured in vivo in animals and in patients using standard clinical MRI systems. Using a pharmacokinetic imaging approach the parameter that can be measured most readily is the tumoral t(1/2) of 5-FU. Patients whose tumoral t(1/2) of 5-FU is equal to/greater than 20 min are designated as "trappers", and those whose tumoral t(1/2) of 5-FU is less are nontrappers. Trapping of 5-FU in tumors is a necessary, albeit not a sufficient condition, for response. Problems associated with the technical aspects of these measurements have been discussed, as well as how modulators and other agents will affect the tumoral t(1/2) of 5-FU. The rationale for the biological processes underlying the fate of 5-FU in humans has been illustrated with the use of a 12 compartment model, where several of the steps have been discussed and the consequences of their inhibition/stimulation related to the noninvasive studies that can be performed with modulators of the action of 5-FU. These 19F-NMR studies have now been extended to other fluoropyrimidines, some of which are prodrugs of 5-FU, and others where the fluorine atoms are on the ribose ring. These studies also reveal information that has both scientific and clinical significance. The studies presented here illustrate some of the potential and some of the usefulness of 19F-MRS in patient management and in drug development. It is a technique that has proven itself. PMID- 10699306 TI - Monitoring pharmacokinetics of anticancer drugs: non-invasive investigation using magnetic resonance spectroscopy. AB - Magnetic resonance spectroscopy (MRS) offers the unique advantage of detecting, identifying and quantifying chemicals deep within the living body in a totally non-invasive manner. In studies on pharmacology and toxicology of anticancer drugs, MRS and the closely related technique magnetic resonance imaging (MRI) have many uses. MRS in particular, despite its low sensitivity, offers unique insights into pharmacokinetics (the changing concentration of the drug at its site of action) which can be monitored, and metabolism (both activation and detoxification) can be detected in real time. This review considers some recent work on (19)F, (31)P, (1)H and (13)C MRS of anticancer drugs. Future possibilities for (13)C MRS and (1)H MRS studies of drugs and their metabolites are considered in detail. PMID- 10699307 TI - Evolution from empirical dynamic contrast-enhanced magnetic resonance imaging to pharmacokinetic MRI. AB - For chemotherapy to be effective against cancers which grow as solid tumors, agents must reach all tumor cells in effective quantities. Although many clinical trials include studies of the pharmacokinetics of the agents in body fluids such as blood or cerebrospinal fluid (CSF), there is presently no widely applicable way to determine access of chemotherapeutic agents to all regions of a solid tumor in an individual patient. This review discusses a relatively new methodology in MR imaging - dynamic contrast-enhanced imaging for exploring tumor microcirculation and drug access by imaging the uptake, or leakage, of contrast agent into tumor interstitial (extracellular and extravascular) space. The aims and methods of dynamic contrast-enhanced MRI evaluations to measure contrast uptake are distinguished from dynamic contrast-enhanced MRI to measure blood volume or flow, by MR imaging of the first-pass effects of a contrast bolus. Measures of contrast uptake by dynamic MRI have demonstrated a convincing ability to aid in diagnosing the presence of viable tumor and to measure response for a range of human tumors. This body of clinical results will be summarized. While questions remain to be answered about how to extract non-invasive pharmacokinetic measures of drug access from these novel dynamic imaging methods, we are optimistic that these methods can provide important new clinical measures that reflect the range of biological variation within and between naturally-occurring solid tumors. PMID- 10699308 TI - Noninvasive measurements for studying the tumoral pharmacokinetics of platinum anticancer drugs in solid tumors. AB - An effective methodology to determine the amount of cisplatin or carboplatin at the solid tumor site in a noninvasive manner may enable clinicians to design drug regimens based on an individual's in situ pharmacokinetics. Such noninvasive methods may allow optimization of an individual's drug exposure at the target site, as well as provide a screening measure to determine individual efficacy based on exposure to these platinated drugs. 195mPt appears to be the radionuclide of platinum most suitable for radiolabeling cisplatin or carboplatin, and an analysis is presented of the methods available for preparing such radiolabeled drugs. The use of this methodology is illustrated in detail in studies in animals, as well as some preliminary studies in humans. The animals used were Sprague Dawley rats bearing the Walker 256 carcinoma, and drug biodistribution was studied following administration of cisplatin or carboplatin radiolabeled with 195mPt. This radionuclide permitted noninvasive imaging of the drug and its metabolites at the tumor site and at selected organs. The results obtained show an ability to estimate the amount of platinated drug species in the tumor environment using a noninvasive methodology. Various compartmental models were tested, some of which could be validated experimentally. This noninvasive method is able to provide individual estimates of the active component of the drug at the target site, and is therefore a method that can be implemented in human studies. PMID- 10699309 TI - Recent advances in cellular, sub-cellular and molecular targeting. PMID- 10699310 TI - Tuftsin-bearing liposomes in treatment of macrophage-based infections. AB - The use of liposomes as drug carriers in treatment of various diseases has been explored extensively for more than 20 years. 'Conventional' liposomes, when administered in vivo by a variety of routes, rapidly accumulate in the mononuclear phagocyte system (MPS). The inherent tendency of the liposomes to concentrate in MPS can be exploited in enhancing the non-specific host defence against infections by entrapping in them the macrophage modulators, and as carriers of antibiotics in treatment of intracellular infections that reside in MPS. This must further be enhanced by grafting on the liposome surface the ligands, e.g. tuftsin, that not only binds specifically to the MPS cells but also enhances their natural killer activity. Keeping this in view, we designed and developed tuftsin-bearing liposomes as drug carriers for the treatment of macrophage-based infections and outline these studies in this overview. PMID- 10699311 TI - Targeted drug delivery via the folate receptor. AB - The folate receptor is a highly selective tumor marker overexpressed in greater than 90% of ovarian carcinomas. Two general strategies have been developed for the targeted delivery of drugs to folate receptor-positive tumor cells: by coupling to a monoclonal antibody against the receptor and by coupling to a high affinity ligand, folic acid. First, antibodies against the folate receptor, including their fragments and derivatives, have been evaluated for tumor imaging and immunotherapy clinically and have shown significant targeting efficacy in ovarian cancer patients. Folic acid, a high affinity ligand of the folate receptor, retains its receptor binding properties when derivatized via its gamma carboxyl. Folate conjugation, therefore, presents an alternative method of targeting the folate receptor. This second strategy has been successfully applied in vitro for the receptor-specific delivery of protein toxins, anti-T-cell receptor antibodies, interleukin-2, chemotherapy agents, gamma-emitting radiopharmaceuticals, magnetic resonance imaging contrast agents, liposomal drug carriers, and gene transfer vectors. Low molecular weight radiopharmaceuticals based on folate conjugates showed much more favorable pharmacokinetic properties than radiolabeled antibodies and greater tumor selectivity in folate receptor positive animal tumor models. The small size, convenient availability, simple conjugation chemistry, and presumed lack of immunogenicity of folic acid make it an ideal ligand for targeted delivery to tumors. PMID- 10699312 TI - Receptor-mediated targeting of toxins to intraerythrocytic parasite Plasmodium falciparum. surolia@jncasr.ac.in. AB - The increasing prevalence of drug-resistant Plasmodium falciparum malaria and the absence of effective vaccines or of vector control measures makes the development of new antimalarial drugs and other approaches for treating malaria, an urgent priority. The development of immunotoxins for targeted cytotoxic effects to kill the parasite is an attractive alternative therapeutic concept. The cytocidal effect of such hybrid molecules is highly specific and requires only minute doses. Cell surface receptor-directed targeting of toxins (hybrid toxins or immunotoxins) to human malaria parasite could eventually be developed as an effective therapy for malaria. Hybrid toxins may provide means of controlling this dreadful disease and counter morbidity as well as mortality. Our results suggests that hybrid toxins are potent and efficacious in killing the parasite and that these agents should be examined in an appropriate in vivo model of malaria. PMID- 10699313 TI - Delivery of lipids and liposomal proteins to the cytoplasm and Golgi of antigen presenting cells. mangala.rao@na.amedd.army.mil. AB - Liposomes have the well-known ability to channel protein and peptide antigens into the MHC class II pathway of phagocytic antigen-presenting cells (APCs) and thereby enhance the induction of antibodies and antigen-specific T cell proliferative responses. Liposomes also serve as an efficient delivery system for entry of exogenous protein and peptide antigens into the MHC class I pathway and thus are very efficient inducers of cytotoxic T cell responses. Soluble antigens that are rendered particulate by encapsulation in liposomes are localized both in vacuoles and in the cytoplasm of bone marrow-derived macrophages. Utilizing fluorophore-labeled proteins encapsulated in liposomes we have addressed the question of how liposomal antigens enter the MHC class I pathway. After phagocytosis of the liposomes, the fluorescent liposomal protein and liposomal lipids enter the cytoplasm where they are processed by the proteasome complex. The processed liposomal protein is then transported via the TAP complex into the endoplasmic reticulum and the Golgi complex. Both the liposomal lipids and the liposomal proteins appear to follow the same intracellular route and they are processed as a protein-lipid unit. In the absence of a protein antigen (empty liposomes), there is no organelle-specific localization of the liposomal lipids. In contrast, when a protein is encapsulated in these liposomes, the distribution of the liposomal lipids is dramatically affected and the liposomal lipids localize to the trans-Golgi area. Localization of the protein in the trans-Golgi area requires liposomal lipids. Similarly, for the localization of liposomal lipids in the trans-Golgi area, there is an obligatory requirement for protein. Therefore, the intracellular trafficking patterns of liposomal lipids and liposomal protein are reciprocally regulated. Presence of both liposomal lipids and liposomal protein in the trans-Golgi therefore facilitates the entry of liposomal antigens into the MHC class I pathway. It is also possible that liposomal lipids are presented to T cells via the recently described CD1 pathway for lipid antigens. Because liposome-formulated vaccines have the potential to stimulate antibody as well as cellular immune responses to protein and lipid components, this approach could prove to be extremely useful in designing vaccine strategies. PMID- 10699314 TI - Lysosome membrane permeability: implications for drug delivery. AB - The membrane of the lysosome contains substrate-specific porters for a wide range of metabolites. Their physiological role is in promoting the efflux of the products of intralysosomal catabolism. With few exceptions, the specificity of these porters makes them unlikely candidates for the translocation of xenobiotics across the lysosome membrane. Where efflux from the lysosome is possible, it is likely to be accomplished by passive diffusion. Experimental studies on passive diffusion across the lysosome membrane have shown that its characteristics are similar to those of other biological membranes. Ease of permeation decreases with increasing hydrophilicity. Macromolecules and some highly hydrophilic molecules as small as sucrose are effectively non-permeant. The notional hydrogen-bonding capacity of molecules (an inverse correlate of oil:water partition coefficient) has been found a good predictor of permeance. Predictions of ease of permeation across lysosome membranes is of value when drug delivery strategies are contemplated that involve a drug-conjugate reaching the lysosome compartment and drug release there by the lysosomal enzymes. These strategies will be unsuccessful if the drug is unable to leave the lysosome and reach the cellular sites where its pharmacological action is required. PMID- 10699315 TI - Endosomes, lysosomes: their implication in gene transfer. AB - Plasmid DNA, naked or bound to a non-viral vector, is taken up by endocytosis. As a result, it has to travel through the intracellular endocytic pathway involving endosomes and lysosomes. However, some DNA molecules must escape these organelles to reach the nucleus where transcription takes place. In this paper, we consider different factors that could affect the trafficking of plasmid DNA and influence transfection efficiency. PMID- 10699316 TI - Bacterial pore-forming hemolysins and their use in the cytosolic delivery of macromolecules. AB - Advances in our understanding of fundamental cell biological processes have facilitated an expansion of therapeutic approaches to altering cellular physiology and phenotype. As many of these methods involve macromolecular agents that act on targets within the nucleus or cytoplasm, achieving their full potential ultimately requires the efficient delivery of these agents across the cell membrane barrier into the cytosol. Various strategies have been employed to enhance cytosolic delivery. These include either directly penetrating the plasma membrane, or avoiding degradation within the hydrolytic environment of the endosomal/lysosomal pathway after endocytic uptake. Some of the more promising methods in this regard have exploited the mechanisms utilized by certain viruses and bacteria for escaping into their host cell's cytosol. In this review, we will discuss some of these methods with an emphasis on the use of pore-forming proteins from bacteria. Particular attention will be drawn to the pH-sensitive endosomolytic bacterial hemolysins, such as listeriolysin O, and the potentiol for their use in cytosolic drug delivery systems. PMID- 10699317 TI - Gene-transfer systems for human endothelial cells. stewart.martin@nottingham.ac.uk. AB - By virtue of its location and importance in a number of pathophysiological processes the endothelium represents an attractive target tissue for gene transfer and gene-therapy strategies. Although it is important to maximise gene transfer to endothelial cells in such strategies primary human endothelial cells have proven to be rather intransigent to a variety of transfection techniques both in vitro and in vivo. We report on the variety of techniques in current use, revealing their strengths and weaknesses, indicate the steps that should ideally be taken to optimise expression and discuss the usefulness and future directions for viral mediated transduction. PMID- 10699318 TI - Drug delivery to mitochondria: the key to mitochondrial medicine. AB - The major function of mitochondria in human cells is to provide ATP by oxidative phosphorylation. However, mitochondria have many other roles including the modulation of intracellular calcium concentration and the regulation of apoptotic cell death. Furthermore, the mitochondrial respiratory chain is a major source of damaging free radicals. Consequently, mitochondrial dysfunction contributes to a number of human diseases, ranging from neurodegenerative diseases and ischaemia reperfusion injury to obesity and diabetes. In addition, mutations to nuclear or mitochondrial DNA cause a number of human diseases. Therefore, strategies to prevent mitochondrial damage or to manipulate mitochondrial function in clinically useful ways may provide new therapies for a range of human disorders. Here we outline why mitochondria are a potentially important target for drug delivery and discuss how to deliver bioactive molecules selectively to mitochondria within cells. PMID- 10699319 TI - Advantage of gene gun-mediated over intramuscular inoculation of plasmid DNA vaccine in reproducible induction of specific immune responses. AB - Utilizing a plasmid DNA encoding a single cytotoxic T lymphocyte (CTL) epitope and that encoding ovalbumin (OVA), we compared the reproducibility in the induction of immune responses by gene gun and intramuscular immunization. As compared to intramuscular inoculation, gene gun DNA immunization appeared to bring about highly reproducible and reliable results in the induction of specific CTL and IFN-gamma production to the CTL epitope and production of anti-OVA IgG. The results obtained by intramuscular inoculation vary significantly. Our data shown here strongly suggest that gene gun immunization of skin is a much more reliable method for DNA vaccination to induce effective immune responses in an animal model. PMID- 10699320 TI - Efficient extracellular production of recombinant Escherichia coli heat-labile enterotoxin B subunit by using the expression/secretion system of Bacillus brevis and its mucosal immunoadjuvanticity. AB - A gene encoding the mature Escherichia coli heat-labile enterotoxin B subunit (LTB) was introduced in a vector pNU212 and expressed at high levels in Bacillus brevis HPD31. The maximum amount of recombinant LTB (rLTB) secreted into the modified 5PY medium containing erythromycin was about 350 mg l(-1) when cultivated at 30 degrees C for 8 days. The rLTB purified directly from the culture supernatant by using D-galactose immobilized agarose was identical to the native LTB with respect to the molecular weight determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and the amino terminal amino acid sequence. Western blot analysis with antiserum to cholera toxin B subunit (CTB) indicated that rLTB had cross-reactivity to native CTB and its GM1 binding ability was almost the same as that of the CTB. The rLTB predominantly showed the pentameric form when non-boiled samples were applied to SDS-PAGE. When rLTB was administered intranasally to mice with diphtheria toxoid (D(T)), it resulted in the substantial stimulation of D(T)-specific serum IgG antibody, and in the induction of moderate levels of D(T)-specific mucosal IgA antibody responses in the nasal cavities and in the lung, suggesting that purified rLTB acts as a promising immunoadjuvant on mucosal immunizations. PMID- 10699321 TI - Single inoculation of replication-defective adenovirus-vectored vaccines at birth in piglets with maternal antibodies induces high level of antibodies and protection against pseudorabies. AB - In neonates, one limitation of vaccination is its inhibition by maternal antibodies. We show that piglets vaccinated intramuscularly once at birth with recombinant replication-defective adenoviruses developed comparable neutralizing antibody response against pseudorabies virus, independently of the presence or absence of maternal antibodies, and were partially protected against challenge 16 weeks later. PMID- 10699322 TI - Immunization of healthy adults with a single recombinant pneumococcal surface protein A (PspA) variant stimulates broadly cross-reactive antibodies to heterologous PspA molecules. AB - Pneumococcal surface protein A (PspA) is a highly variable protein found on all strains of pneumococci. To be successful, a PspA-based vaccine for S. pneumoniae must induce antibodies that are broadly cross-reactive. To address whether cross reactive antibodies could be induced in man, we evaluated serum from adults immunized with recombinant clade 2 PspA from strain Rx1. Immunization with 5-125 microg rPspA lead to a significant increase in circulating anti-PspA antibodies, as well as antibodies reactive to heterologous rPspA molecules. Increased binding of post-immune sera to 37 pneumococcal strains expressing a variety of PspA and capsule types was observed, versus pre-immune sera. The extent of cross-clade reactivity of human anti-rPspA followed roughly the amount of sequence homology to the non-clade 2 antigens. It is hypothesized that priming of humans by natural exposure to S. pneumoniae contributes to the breadth of the cross-reactivity of antibody to PspA. PMID- 10699323 TI - CpG DNA induces stronger immune responses with less toxicity than other adjuvants. AB - The ability to augment protective immune responses with minimal side effects is quintessential for a good adjuvant. This study has compared various adjuvants that are used in animal research (Freund's complete and incomplete adjuvants, Titermax Gold), are licensed for human use (alum), or are in clinical testing for humans (monophosphoryl lipid, CpG DNA), for their ability to augment humoral responses to a model antigen (hepatitis B surface antigen) and for the degree of damage they caused in the injected muscle. According to the data, the adjuvant combination CpG DNA+alum had the greatest potential to augment immune responses with minimal side effects at the injection site. Evaluation of antibody isotypes indicated Th2 responses (no IgG2a) with all adjuvants except monophosphoryl lipid and CpG DNA, which gave mixed Th1/Th2 responses (IgG1 and IgG2a). Strong Th1 responses (predominantly IgG2a) were obtained with combinations of CpG DNA with other adjuvants. PMID- 10699324 TI - Evaluation of the live attenuated cpts 248/404 RSV vaccine in combination with a subunit RSV vaccine (PFP-2) in healthy young and older adults. AB - The safety and immunogenicity of the live attenuated cold-passaged, temperature sensitive (cpts) 248/404 respiratory syncytial virus (RSV) A2 and the RSV A2 purified F glycoprotein (PFP-2) vaccine candidates were evaluated in a placebo controlled trial in 60 healthy young adults and 60 healthy elderly subjects using simultaneous and sequential (cpts 248/404 followed by PFP-2) vaccination schedules. Both vaccines were well tolerated. The cpts 248/404 vaccine was moderately infectious in both young and old volunteers, but was highly restricted in replication in those who were infected. After both vaccines, RSV neutralizing antibody (neut Ab) titers increased fourfold in 22% of young subjects and in 16% of elderly subjects. Of those with low levels of RSV neut Ab (titer <9), 10/12 (83% of) young subjects and six/eight (75% of) elderly subjects had a >/=four fold rise in neut Ab titer. Young and elderly subjects immunized simultaneously had similar serum IgG and IgA postimmunization titers to RSV F (IgG, 16.4 vs 16.2, IgA 11.6 vs 12. 5, respectively) as did those who were immunized sequentially (IgG 17.4 vs 17.0, IgA 13.0 vs 13.5). In both age groups, sequential immunization elicited higher postimmunization RSV F IgG and IgA titers than simultaneous immunization. Further studies that combine the PFP-2 subunit vaccine with a less attenuated RSV vaccine should be performed. PMID- 10699325 TI - Distribution of immunoglobulin G (IgG) and A (IgA) subclasses following Q fever vaccination with soluble phase I Coxiella burnetii extract. AB - High levels of IgG1, IgG3 and IgA2 antibodies have been observed in patients with Q fever following Coxiella burnetii infection. This IgG subclass distribution is more typical of viral and autoimmune diseases than of bacterial infections. It seemed, therefore, of interest to carry out a prospective study of the distribution of immunoglobulin subclasses after vaccination with phase I C. burnetii tricloroacetic soluble extracts to detect possible differences with respect to natural infection. The antibody response found in vaccinees was mainly restricted to the IgG1, IgG2 and IgA1 subclasses. These findings confirm differences in isotype distribution when compared to those of patients with acute or chronic Coxiella infections and opens an area of interest with respect to the role of IgA subclasses. PMID- 10699326 TI - Therapeutic vaccination against HSV-2: influence of vaccine formulation on immune responses and protection in mice. AB - Therapeutic immunisation may represent a means of influencing viral infections that persist in the host by modulating the nature or level of host immunity. To assess the influence of the form of the antigenic stimulus on immunity to type-2 herpes simplex virus (HSV-2), mice pre-infected with sublethal doses of HSV-2 were immunised with various HSV-2 vaccine formulations prior to challenge infection with heterologous HSV-1. Measurements of interleukin-2 (IL-2), interleukin-4 (IL-4) and interferon-gamma (IFN-gamma) levels in mouse spleen cell cultures restimulated in vitro with HSV-2 antigens showed that, depending on the form of HSV-2 antigen preparation used in this therapeutic context, changes in the levels of these cytokines could be effected. Measurement of HSV-specific antibody by serological tests support the contention that immunisation of HSV-2 infected mice can either enhance the existing Th1-like immune response elicited following HSV-2 infection, or modulate this response towards a more Th2-like profile, and this is dependent on the form of the antigenic stimulus. The degree of protection against subsequent lethal, heterologous HSV-1 challenge infection varied according to the nature of the infection and the immunisation history of the animals. PMID- 10699327 TI - Microparticles in MF59, a potent adjuvant combination for a recombinant protein vaccine against HIV-1. AB - Novel adjuvant formulations involving PLG microparticles with entrapped recombinant protein antigens (env gp120 and p24 gag) from human immunodeficiency virus type-1 (HIV-1), dispersed in the emulsion adjuvant MF59 were evaluated as potential HIV-1 vaccine candidates in mice and baboons. In mice, the adjuvant combination induced significantly enhanced antibody responses in comparison to either adjuvant used alone. In addition, the polylactide co-glycolide polymer (PLG) microparticles and MF59 combination induced CTL activity against HIV-1 p24 gag. In baboons, the adjuvant combination induced significantly enhanced antibody titers after a single dose of gp120, but the responses were comparable to gp120 in MF59 alone after boosting. Both MF59+gp120 alone and PLG/gp120 in MF59 induced neutralizing antibodies against a T cell line-adapted (TCLA) strain and a primary isolate of HIV-1. In contrast to the observations with gp120, immunization in baboons with PLG/p24 in MF59 induced significantly enhanced antibody responses after boosting, in comparison to immunization with MF59 alone + p24. PMID- 10699328 TI - Reversible protection of disulfide bonds followed by oxidative folding render recombinant hCGbeta highly immunogenic. AB - Active immunization of women against human chorionic gonadotropin (hCG) has been considered as a promising option for contraception. However, prototype hCG vaccines based on natural sources of antigen are expected to be costlier for use by common people. In the present report, a functionally active, cost-effective antigen of bacterial origin has been described. Sulfonation of thiol groups of the protein, anion-exchange purification, refolding with concomitant formation of disulfide bonds in the presence of cysteamine-cystamine redox buffer, and slow removal of denaturant resulted in 95% homogeneous, monomeric form of the antigen. The recombinant processed antigen [CGbeta(p)] obtained this way was highly immunopotent. Cellular DNA and endotoxin contaminants were appreciably low in the final product. The immunogenic response was drastically reduced with the unprocessed antigen. This finding envisages better prospect of a cost-effective hCG vaccine for birth control. PMID- 10699329 TI - Purification and characterization of Streptococcus pneumoniae palmitoylated pneumococcal surface adhesin A expressed in Escherichia coli. AB - All Streptococcus pneumoniae isolates tested to date express a species-common lipoprotein designated as pneumococcal surface adhesin A (PsaA). This protein is cell-associated, hydrophobic, immunogenic, and genetically conserved. It is currently under investigation as a potential component in third-generation pneumococcal vaccine formulations. To overcome the problem of low-level expression of native hydrophobic PsaA in S. pneumoniae, and also of the recombinant PsaA (rPsaA) in Escherichia coli, we generated a stable E. coli construct expressing functional palmitoylated rPsaA ( approximately 10 mg/l of fermentation culture) using Borrelia burgdorferi outer surface protein A (OspA, a hydrophobic lipoprotein) signal peptide. By Western blot analysis, the chimeric rPsaA ( approximately 34 kDa) was detected in the cell lysate using anti-PsaA antibodies. It was partially purified by extracting the cell pellet with PBS/Triton X(R)-114 buffers, followed by anion exchange filter chromatography. A trypsin digestion profile of rPsaA closely resembled that of the native protein, as revealed by SDS-PAGE/silver staining. Lipidation of rPsaA was confirmed by labeling recombinant E. coli cells with [(3)H] palmitic acid and analyzing the labeled E. coli cells by Western blotting coupled with autoradiography. Further, analysis of purified rPsaA by mass spectrometry (MALDI-TOF) revealed a heterogenous spectrum with a major peak (M+H)(+1) of mass 33,384 Da (theoretical mass of palmitoylated rPsaA=33,361 Da). Purified rPsaA was immunogenic in CBA/NCAHN-XID female mice following intranasal immunization with or without adjuvant, as determined by measurement of anti-PsaA serum IgG levels. These anti PsaA antibodies reacted with both native and rPsaA polypeptides. Our data strongly suggest that E. coli-expressed rPsaA is palmitoylated and closely resembles the native protein in structure and immunogenicity. It was also observed to elicit measurable protection against nasopharyngeal carriage with S. pneumoniae. PMID- 10699330 TI - IFN-gamma, IL-4, IL-10 and IL-12 gene expression in BCG-Leishmania vaccination of Trypanosoma cruzi-infected mice. AB - We have previously shown that vaccination of BALB/c mice with a combination of BCG plus killed Leishmania promastigotes, applied by the i.p. route 10 and 3 days before Trypanosoma cruzi inoculation, prolonged their survival and decreased their parasitaemia. In the present study we show that the BCG-Leishmania vaccine induced higher levels of circulating IFN-gamma in acute and chronic infection of mice [on day 25 and 40 post-infection (p.i.) respectively], in comparison to unvaccinated animals (PBS-treated). Though the IFN-gamma mRNA content of spleen cells of vaccinated and infected mice (on day 25 p.i.) was similar to that of unvaccinated animals, the BCG-Leishmania vaccine enhanced significantly the production of IFN-gamma by spleen cells stimulated with T. cruzi antigens. This effect was observed to a lower extent in BCG- and Leishmania-treated mice. The BCG-Leishmania vaccine reduced the expression of the IL-10 mRNA of splenocytes as soon as day 12 p.i., before the peak parasitaemia. Such this effect was not observed in BCG- or Leishmania-treated animals. On day 25 p.i., the BCG plus Leishmania- or BCG-treatment of mice abolished the capacity of spleen cells to produce IL-10 in response to T. cruzi antigens. The levels of mIL-4 RNA and protein production were not modified in any group of mice. T. cruzi infection in BCG-Leishmania-vaccined mice stimulated an early and high production of IL-12 transcripts in spleen cells during the acute phase of the infection, that was prolonged during the chronic phase of infection. This effect was weaker or absent in BCG- and Leishmania-treated animals, respectively. These results indicate that the BCG-Leishmania vaccine stimulates the production of IL-12 and IFN-gamma, but inhibits that of IL-10 and is without effect on IL-4 when mice are infected with T. cruzi. This highlights the key role of endogenously produced IFN-gamma, IL-10 and IL-12 in the control of T. cruzi acute and chronic infection in mice and the favorable modulation of their balance by a vaccination combining BCG and Leishmania. PMID- 10699331 TI - Case-control study of allergic reactions to Japanese encephalitis vaccine. AB - Japanese encephalitis (JE) vaccine is widely used in Asia for childhood immunizations, but the vaccine is also used for travellers to Asia from other parts of the world. In Denmark, more than 400,000 doses have been distributed from Statens Serum Institut since 1982. In 1989, the first allergic mucocutaneous reactions after JE vaccination were registered in Denmark and, although the number of reactions have decreased since 1992, reactions are still observed. No explanation of these reactions have been found. The present case-control study, including 49 travellers with allergic reactions and 148 travellers without similar reactions after JE vaccination was performed in order to clarify any possible risk factors. About one third of the adverse reactions to the vaccine could be attributed to an allergic predisposition in the vaccinees. The main risk factors were young age, female gender and previous allergic skin reactions or hayfever. The study also indicated that cases more often reacted to nickel and more often had severe edema after mosquito or other insect bites. Hormone intake was more often spontaneously reported by females in the case group. Accordingly, information on any history of allergy in young adults should be given before JE vaccination, the vaccination should be carried out more than a week before departure and antihistamine treatment should be available if a reaction occurs. PMID- 10699332 TI - DNA immunization utilizing a herpes simplex virus type 2 myogenic DNA vaccine protects mice from mortality and prevents genital herpes. AB - A gene transfer vector for DNA immunization was developed in which the promoter was derived from the murine muscle creatine kinase (MCK) gene; a gene expressed only in differentiated skeletal muscle. In vitro, we observed high-level, but unrestricted, gene expression from the cytomegalovirus (CMV) promoter unlike expression from the MCK promoter which was weak but restricted to myofibers. A myogenic DNA vaccine (MDV) that encoded the glycoprotein D gene from herpes simplex virus type-2 (HSV-2) was used to DNA immunize mice. MDV immunization resulted in virus specific immunity that protected HSV-2 infected mice from mortality and prevented the development of genital herpes. Therefore, we conclude that high-level gene expression or the use of a strong transcription unit was not a prerequisite for an efficacious DNA vaccine and the use of a nonviral tissue specific promoter could suffice. PMID- 10699333 TI - Effects of IL-12 on immune responses induced by transcutaneous immunization with antigens formulated in a novel lipid-based biphasic delivery system. AB - The development of non-invasive methods for the delivery of proteins through the permeability barriers, such as the intact skin, will greatly facilitate the administration of human and veterinary vaccines. In the present study we used recombinant Pasteurella haemolytica leukotoxin (Lkt) and hen egg lysozyme (HEL) as model antigens to investigate the ability of transdermal administration of vaccine antigens to induce humoral and cellular responses in mice and to assess the immunomodulatory effects of IL-12 on these antigen-specific immune responses. Mice were immunized by the transdermal route with Lkt or HEL formulated in a novel lipid-based biphasic delivery system (BPDS). Transdermal delivery of Lkt or HEL induced strong polarized Th2 responses characterized by enhancement of antigen-specific IgG1 antibody subclass and predominant induction of antigen specific IL-4 over IFN-gamma in spleen and draining lymph nodes cells. Animals immunized by topical application of formulations containing antigen and IL-12 developed significantly lower antibody titres without significant changes in IL-4 or IFN-gamma secreting cells (SC) in the draining lymph nodes or spleen cells. Our results indicated that application of antigens formulated in BPDS induced antigen-specific immune responses. Furthermore, incorporation of IL-12 to the vaccine formulation influences the induction of antibody responses induced by transdermal immunization. We demonstrated the feasibility of using this technology for the development of non-invasive methods of vaccine administration. PMID- 10699334 TI - Vaccination of mice with DNA encoding a large fragment of botulinum neurotoxin serotype A. AB - The potential utility of using DNA vaccination to protect mice from the microbial neurotoxin, botulinum toxin type A, was evaluated. A synthetically derived gene encoding a carboxyl-terminal 50 kDa fragment of the toxin was placed in two sites in the DNA inoculation vehicle pCMVint-BL (Vical), one predicted to lead to MHC I processing (pJT-1 construct) and the other to direct MHC II processing (pJT-2 construct). Mice were then inoculated at 3 week intervals with these two constructs and with the vehicle alone and evaluated for protection from botulinum toxin by i.p. challenges with various toxin doses. Protection was observed at about week 10-11 from toxin doses of 25-100 LD(50). Only animals inoculated with pJT-2 exhibited protection. In dose-response experiments, 50 micrograms of DNA was the minimal dose required to elicit a protective response against serotype A, while protection against serotypes B or E was not obtained. With standard ELISA testing, a relationship was observed between the level of protection and the level of ELISA reactive antibody. Our results support the concept that DNA vaccination is a viable methodology to use in cases where protection from toxins is the goal. PMID- 10699335 TI - Immunogenicity of Ty-VLP bearing a CD8(+) T cell epitope of the CS protein of P. yoelii: enhanced memory response by boosting with recombinant vaccinia virus. AB - We characterized the immunogenicity of the hybrid Ty-virus-like carrying the CD8(+) T cell epitope (SYVPSAEQI) of the circumsporozoite (CS) protein of Plasmodium yoelii (TyCS-VLP), a rodent malaria parasite. Balb/c mice were immunized with hybrid TyCS-VLP, and their CS-specific CD8(+) T cell response was quantitatively evaluated with the ELISPOT assay, based on the enumeration of epitope specific gamma-interferon secreting CD8(+) T cell. A single immunization with the TyCS-VLP by a variety of routes and doses indicated that the maximal response occurred in mice, which were immunized with 50 micrograms of these particles, administered via intramuscular. Combined immunization of mice with this TyCS-VLP followed by recombinant vaccinia virus expressing the entire P. yoelii CS protein (VacPyCS) or irradiated sporozoites, induced high levels of IFN gamma-producing cells. The immunization regime, priming with TyCS-VLP and boosting with VacPyCS generated a potent protective immune response, which strongly inhibited P. yoelii liver stages development and protected 62% of the mice against a subsequent live P. yoelii sporozoite challenge. PMID- 10699336 TI - Preparation of pneumococcal capsular polysaccharide-protein conjugate vaccines utilizing new fragmentation and conjugation technologies. AB - There is a global urgent need for a new efficient and inexpensive vaccine to combat pneumococcal disease, which should also be affordable in developing countries. In view of this need a simple low-cost technique to prepare such a vaccine was developed. The preparation of serotype 14 and 23F pneumococcal capsular polysaccharide (PnPS)-protein conjugates to be included in a forthcoming multivalent PnPS conjugate vaccine is described. Commercial lots of PnPSs produced according to Good Manufacturing Practice from Streptococcus pneumoniae serotype 14 (PS14) and 23F (PS23F) were partially depolymerized by sonication or irradiation in an electron beam accelerator. The PnPS fragments were conjugated to tetanus toxoid (TT) using a recently developed conjugation chemistry. The application of these new simple, efficient and inexpensive fragmentation and conjugation technologies allowed the synthesis of several PnPS-protein conjugates containing PnPS fragments of preselected sizes and differing in the degree of substitution. The PS14TT and PS23FTT conjugate vaccine candidates were characterized chemically and their immunogenicity was evaluated in rabbits and mice. All PnPS conjugate vaccines, unlike the corresponding plain polysaccharides, produced high IgG titres in both animal species. The PS14TT conjugates tended to be more immunogenic than the PS23FTT conjugates. The immune response to the PS14TT conjugates, but not to the PS23FTT conjugates, was related to the size of the conjugated polysaccharide hapten. Both types of conjugates elicited strong booster effects upon secondary immunizations, resulting in high IgG1, IgG2a and IgG2b titres. PMID- 10699337 TI - D-LysAsnProTyr tetrapeptide: a novel B-cell stimulant and stabilized bursin mimetic. AB - The tetrapeptide I (D-lysine-L-asparaginyl-L-prolyl-L-tyrosine or D LysAsnProTyr), and analogue sequences, were synthesized and evaluated for the ability to stimulate immune cell subsets. These sequences were selected based on their perceived ability to readily adopt a beta-turn structure. In vitro immunological assays revealed a robust stimulation of mitogen activated B-cell proliferation and a modest to significant stimulation of cytotoxic T lymphocytes (CTLs). Further, this in vitro stimulation of B-cells was accompanied by an in vivo expansion of B-cells in C57BL/6 mice, as demonstrated by immunophenotyping experiments. Interestingly, a conformational analysis of the low energy conformers of I and the endogenous B-cell stimulant bursin (LysHisGlyNH2) shows that these molecules can be superimposed. However, I displayed significantly enhanced physiological stability. For a number of reasons, I may be a particularly useful vaccine adjuvant. PMID- 10699338 TI - Safety, tolerability and humoral immune responses after intramuscular administration of a malaria DNA vaccine to healthy adult volunteers. AB - DNA-based vaccines are considered to be potentially revolutionary due to their ease of production, low cost, long shelf life, lack of requirement for a cold chain and ability to induce good T-cell responses. Twenty healthy adult volunteers were enrolled in a Phase I safety and tolerability clinical study of a DNA vaccine encoding a malaria antigen. Volunteers received 3 intramuscular injections of one of four different dosages (20, 100, 500 and 2500 microg) of the Plasmodium falciparum circumsporozoite protein (PfCSP) plasmid DNA at monthly intervals and were followed for up to twelve months. Local reactogenicity and systemic symptoms were few and mild. There were no severe or serious adverse events, clinically significant biochemical or hematologic changes, or detectable anti-dsDNA antibodies. Despite induction of excellent CTL responses, intramuscular DNA vaccination via needle injection failed to induce detectable antigen-specific antibodies in any of the volunteers. PMID- 10699339 TI - Estimation of the efficacy of live, attenuated influenza vaccine from a two-year, multi-center vaccine trial: implications for influenza epidemic control. AB - The authors provide an analysis of data from a two-year (1996-1998), multicenter (ten US cities), double-blinded, placebo-controlled influenza vaccine trial in children. The vaccine was the trivalent cold-adapted influenza vaccine. Estimates are made of the vaccine efficacy for susceptibility to culture-confirmed influenza (VE(S)) while taking inter-center variability in the risk of infection into account. Our overall estimate of VE(S) against influenza is 0.92 (95% confidence interval (CI) 0.89-0.94). In addition, for the second year, although the vaccine contained antigen for A/Wuhan-like (H3N2), the estimated VE(S) for epidemic variant A/Sydney-like (H3N2) was 0.89 (95% CI 0.81-0.94). Thus, the vaccine showed a high degree of protection against a variant not closely matched to the vaccine antigen. With regard to natural immunity, an influenza A infection in the first year reduces the estimated risk of an influenza A infection in the second year by a factor of 0.88 (95% CI 0.21-0.98). When comparing year 1 to year 2, there is a negative correlation of -0.50 in the center-specific attack rates in the placebo groups. This is consistent with the theory that natural immunity provides overall community protection to children. The authors argue that mass vaccination of 70% of the children with the cold-adapted influenza vaccine could provide substantial protection to the community at large. PMID- 10699340 TI - Inactivated meningococci and pertussis bacteria are immunogenic and act as mucosal adjuvants for a nasal inactivated influenza virus vaccine. AB - Whole killed meningococci (Nm) and pertussis bacteria (Bp) were tested for mucosal immunogenicity and as mucosal adjuvants for an inactivated influenza virus vaccine given intranasally to unanaesthetized mice. Virus was given alone, or simply mixed with one of the bacterial preparations, in four doses at weekly intervals. The virus alone induced low but significant increases of influenza specific IgG antibodies in serum measured by ELISA, whereas IgA responses in serum and saliva were insignificant compared to non-immunized controls. With Bp or Nm admixed, serum IgG and IgA and salivary IgA responses to the influenza virus were substantially augmented (P<0.005). However, this adjuvant effect of the bacterial preparations was not significant for responses in the intestine as measured by antibodies in faeces. Antibody responses to Bp itself, but not to Nm, were inhibited by the admixture of the virus vaccine. Moreover, the pertussis preparation induced salivary antibodies which cross-reacted with Nm. Whole-cell bacteria with inherent strong mucosal immunogenicity may also possess mucosal adjuvanticity for admixed particulate antigens which are weakly immunogenic by the nasal route. PMID- 10699341 TI - CpG DNA overcomes hyporesponsiveness to hepatitis B vaccine in orangutans. AB - Oligonucleotides containing immunostimulatory CpG motifs (CpG ODN) have been shown to be potent Th1-type adjuvants for augmenting antigen-specific responses in mice against hepatitis B surface antigen (HBsAg). The hepatitis B virus (HBV) infects only humans and great apes and appears to exist among wild chimpanzees and orangutans. An outbreak of HBV among orangutans being rehabilitated for re introduction to the jungle caused the death of several animals. A prophylactic vaccination program revealed that orangutans are quite hypo-responsive to a current commercial vaccine compared to results obtained previously in humans and chimpanzees. Addition of CpG ODN to hepatitis B vaccine greatly increased the seroconversion rate and the titers of antibody against HBsAg (anti-HBs). This is the first demonstration of CpG DNA in a great ape and the results have important implications for the vaccination of humans against HBV and other diseases. PMID- 10699342 TI - Effect of vaccination with 3 recombinant asexual-stage malaria antigens on initial growth rates of Plasmodium falciparum in non-immune volunteers. AB - A placebo controlled, randomised, double blind trial was conducted in human volunteers to test a mixture of three recombinant Plasmodium falciparum blood stage antigens for its ability to reduce the initial growth rates of parasites. The vaccine contained recombinant MSP2 (3D7 allele), a portion of MSP1 (190LCS.T3) and part of the RESA antigen (C terminal 771 amino acids) in the Montanide ISA 720 adjuvant (SEPPIC). Twelve volunteers received two doses of the vaccine, 6 weeks apart. The five participants in the placebo group received an equivalent volume of the adjuvant emulsion using the same schedule. Antibody responses were low, as has been reported in earlier studies with this combination, while T cell responses were stronger. All the volunteers were challenged with approximately 140 ring infected red cells of the 3D7 cloned line, 4 weeks after the second dose. Parasitaemia was determined once daily from day 4 using a sensitive and quantitative PCR assay. All the volunteers were infected and were treated on day 8, before any developed symptoms. There was no significant difference in initial parasite growth rates between the verum and placebo groups, nor was there any significant correlation between parasite growth rates and any of the measured immunological responses. These results suggest that the formulation tested in this trial did not generate immune responses that were strong enough to reduce parasite growth in naive volunteers. PMID- 10699343 TI - Protection of pigs against classical swine fever with DNA-delivered gp55. AB - Classical swine fever virus causes significant mortality and morbidity in commercial piggeries in many countries in Europe and Asia. The protective antigen, gp55, is highly conformation-dependent and thus killed virus or bacterially produced proteins are not protective. This report demonstrates that DNA vaccination with the gene encoding gp55 can provide protective immunity with inoculation of two doses of 25 microg DNA or a single shot of 200 microg. Furthermore, the DNA can be delivered intramuscularly or by a simple spring loaded needleless inoculator. In addition it is shown that inoculation of the DNA at a single site conveys the same level of immunity as division of the dose between two sites. PMID- 10699344 TI - Hepatitis A vaccine administration: comparison between jet-injector and needle injection. AB - INTRODUCTION: Type A hepatitis virus (HAV) is a serious health problem throughout the world and can be spread via fecal-oral contact. Both immune globulin and an HAV vaccine provide protection, but the vaccine gives complete protection. Efficacy of methods of vaccination in relation to the formation of anti-HAV antibodies is unclear; thus, this study seeks to determine if significant differences exist between the syringe as compared to the jet injection technique. The purpose of this study was to compare in a randomized trial Biojet jet injection system to a needle-syringe method. To determine if a significant difference between these two methods in seroconversion rates or geometric mean titers of anti HAV antibody occurs at day 15, 30, and 210 days after vaccination. METHOD: Anti-HAV IgG(-) adult hospital employees were randomized to receive 1440 EL.U of hepatitis a vaccine (HAVRIX(R)) in 2 doses by either needle or jet injector (Biojector(R)) system at month 0 and 6. HAV seroconversion titer results were measured by the Boehringer-Mannheim method. RESULTS/DISCUSSION: A higher proportion of persons who received HAV vaccine via the Biojector(R) seroconverted with anti-HAV level >/=20 mIU at day 15, 30, and month 7 when compared with a needle injection.Side-effect profiles reported by participants in both methods were below those identified in current published and insert information, but the Biojector(R) had greater local reactivity in all categories when compared to the needle method. PMID- 10699345 TI - Effectiveness of intranasal immunization with HIV-gp160 and an HIV-1 env CTL epitope peptide (E7) in combination with the mucosal adjuvant LT(R192G). AB - LT(R192G) is a novel mucosal adjuvant that induces protective immunity when co administered with certain whole inactivated bacteria or viruses or with subunits of relevant virulence determinants from these pathogens. LT(R192G) stimulates antigen-specific humoral and cellular immune responses, both systemically and in mucosal compartments, and is safe and nontoxic at adjuvant effective doses. Intranasal (IN) immunization of mice with LT(R192G) in conjunction with oligomeric HIV-1 gp160 elevates antigen-specific systemic and mucosal IgG and IgA production and Th1- and Th2-type cytokine responses. Isotype characterization of induced IgG reveals that gp160 alone fails to stimulate IgG2a responses in the absence of adjuvant. Both IgG1 and IgG2a are induced by immunization in the presence of LT(R192G). Additionally, intranasal immunization with a 15-amino acid peptide corresponding to an HIV-1 Env CTL determinant and LT(R192G) induces systemic, peptide-specific CTL activity and Th1 and Th2 cytokine responses that are absent when the adjuvant is excluded from the immunizations. These studies show that LT(R192G) quantitatively and qualitatively enhances cellular and humoral HIV-specific immune responses and that this adjuvant may offer significant advantages toward vaccine development against HIV. PMID- 10699348 TI - "Eliciting HIV-1 envelope-specific antibodies with mixed vaccinia virus recombinants" by samantha D. Rencher, timothy D. Lockey, ranga V. Srinivas, randall J. Owens and julia L. Hurwitz. Vaccine 15(3) pp. 265-272 (1997) PMID- 10699346 TI - Immunization of burn-patients with a Pseudomonas aeruginosa outer membrane protein vaccine elicits antibodies with protective efficacy. AB - The aim of this study was to determine whether the antibodies raised in burn patients by active immunization with a Pseudomonas aeruginosa OMPs vaccine have a protective efficacy against infection with P. aeruginosa. The binding patterns with P. aeruginosa OMPs of immunized burn patient sera were similar to the sera of immunized healthy humans as determined by immunoblot and immunoprecipitation analyses. The sera pooled from immunized burn patients after three immunizations showed a significantly higher opsonophagocytic-killing activity than the corresponding pre-immune sera, while the sera from unimmunized patients collected at the same day did not. Passive immunization of mice with post-immune sera of burn patients significantly enhanced the survival rate upon a lethal challenge with P. aeruginosa compared to the pre-immune sera, indicating the protective ability of the antibodies induced in burn patients by immunization. These results suggest that anti-P. aeruginosa OMPs antibodies elicited in burn patients by active immunization are protective against infection with P. aeruginosa, and provide a rational for further development of the vaccine for prevention against P. aeruginosa infection in burn patients. PMID- 10699347 TI - Quantification of the number of cytotoxic T cells specific for an immunodominant HCV-specific CTL epitope primed by DNA immunization. AB - Priming of strong cellular immune responses to hepatitis C (HCV) is thought to be important for eradication of infection. Although productive infection of HCV occurs only reproducibly in humans and chimpanzees, definition of HCV-specific T cell epitopes in mice is necessary to screen efficiently HCV vaccine strategies for their ability to prime cellular immune responses. Out of seven strains of mice screened for immunodominant CTL epitopes against HCV-1a Core, E2, NS5a and NS5b, only one epitope (p214K9) in only one mouse strain was identified. Enumeration of p214K9-specific CD8+ cells by flow cytometry revealed that the number of epitope specific CTL primed by 'naked' DNA immunization was lower than that reported during viral infection. The p214K9 epitope described here, combined with analysis of CTL responses by flow cytometry, should be instrumental in ranking various HCV vaccine strategies for their ability to prime CTL responses. PMID- 10699349 TI - "An economic evaluation of universal pertussis vaccination in Italy" by ph. Beutels, P. Bonanni, G. Tormans, F. Canale, P. Cuneo crovari. Vaccine 17(19) pp. 2400-2409 (1999) PMID- 10699350 TI - Brain potentials in affective picture processing: covariation with autonomic arousal and affective report. AB - Emotionally arousing picture stimuli evoked scalp-recorded event-related potentials. A late, slow positive voltage change was observed, which was significantly larger for affective than neutral stimuli. This positive shift began 200-300 ms after picture onset, reached its maximum amplitude approximately 1 s after picture onset, and was sustained for most of a 6-s picture presentation period. The positive increase was not related to local probability of content type, but was accentuated for pictures that prompted increased autonomic responses and reports of greater affective arousal (e.g. erotic or violent content). These results suggest that the late positive wave indicates a selective processing of emotional stimuli, reflecting the activation of motivational systems in the brain. PMID- 10699351 TI - Cardiovascular and secretory immunoglobulin A reactions to humorous, exciting, and didactic film presentations. AB - Secretory immunoglobulin A (sIgA) in saliva and cardiovascular activity were measured at rest and in response to three film extracts varying in affective content. Subjective ratings of film impact confirmed a priori assumptions; the humorous film was rated as funnier than the other two films, the didactic film as more boring than the other two films, and the exciting film as more exciting and more stressful than the other two films. The films elicited distinct patterns of cardiovascular autonomic activity. The exciting film provoked changes characteristic of beta-adrenergic activation: increased systolic blood pressure (SBP); heart rate (HR); cardiac output (CO); and shortened pre-ejection period (PEP). The didactic film had little impact on cardiovascular activity. While an increase in total peripheral resistance (TPR) occurred, the humorous film was largely notable for a reduction in beta-adrenergic drive, as evidenced by reduced CO and a lengthening of PEP. In contrast to previous research reporting a rise in sIgA particular to humorous exposures, the sIgA secretion rate, although enhanced by exposure to the films, did not vary with film content. PMID- 10699352 TI - Secretory immunoglobulin A and cardiovascular activity during mental arithmetic: effects of task difficulty and task order. AB - Secretory immunoglobulin A (sIgA) in saliva and cardiovascular activity were measured at rest and during mental arithmetic. Task difficulty was manipulated by presenting easy, hard, and impossible versions of the mental arithmetic task in counterbalanced order, while task novelty was operationalised as order of presentation (i.e. first, second, third). Mental arithmetic elicited significant increases in sIgA concentration and sIgA secretion rate, as well as significant cardiovascular effects. Performance decreased and rated difficulty increased with increasing task difficulty. However, sIgA and cardiovascular activity, with the exception of diastolic blood pressure, were insensitive to variations in task difficulty. In contrast, sIgA concentration and a broad range of cardiovascular variables were influenced by task novelty, with more pronounced activity characterising the task version presented first, irrespective of its level of difficulty. Task novelty would seem to be a more important determinant of sIgA and cardiovascular activity than task difficulty. PMID- 10699353 TI - Respiratory resistance during emotional stimulation: evidence for a nonspecific effect of experienced arousal? AB - We investigated the effects of phasic emotional stimuli on total respiratory resistance (TRR) in 16 nonasthmatic students. Six series of happy, neutral, and depressing affective pictures and self-referent Velten statements were presented. Each stimulus was presented for 12 s and subsequently imagined for 12 s. TRR was measured by forced oscillations throughout the stimulus series, together with ventilation, cardiac activity (including respiratory sinus arrhythmia), and facial EMGs (corrugator supercilii, orbicularis oculi, and masseter). In addition, self-reports of mood, pleasure and arousal were obtained. TRR was increased during happy and depressing stimuli compared to neutral stimuli, with stronger effects for the inspiratory component of TRR. Ventilatory parameters did not explain the changes observed in TRR. Discrimination of affective categories by facial EMG was weak. Although EMG masseter activity did not account for this result, an influence of the respiration measurement procedure on facial EMG cannot be ruled out. The TRR results are in accordance with clinical reports of asthmatic symptom aggravation due to positive or negative emotional arousal. PMID- 10699354 TI - The exhaustive additivity displayed by nitrous oxide has implications for cognitive-energetical theory. AB - The cognitive-energetical approach, which relies on the discrete stage model of additive factors logic, asserts that basal energetical mechanisms such as arousal act via particular information processing stages. The anaesthetic gas nitrous oxide (N(2)O) produces additivity at four of the five perceptual and central stages, but its effect on the remaining stage, feature extraction, is unknown. We investigated this stage using 12 subjects who performed a visual oddball experiment in which two levels of stimulus quality, three levels of breathing mixture (air, 25% and N(2)O, 35%) and three levels of stimulus probability were combined factorially. Reaction time (RT) and P300 were collected simultaneously. The RT results showed additivity between N(2)O, stimulus quality and probability. P300 latency also showed additivity between N(2)O and stimulus quality. Since the discrete stage model cannot easily account for the exhaustive additivity displayed by N(2)O on perceptual and central stages, we performed a continuous cascade model simulation to determine whether it is better able to account for this phenomenon. We found that exhaustive additivity could be reproduced by adding a time delay to the activation rate of the first stage, which we interpreted as evidence that N(2)O causes slowing prior to stage processing. To account for these results, we propose a two-tiered energetical model in which a lower GABAergic reticular system (influenced by N(2)O) modulates the activity of upper 'arousal-like' multidimensional ascending thalamocortical systems. The applicability of this model to drugs such as the barbiturates, the benzodiazepines and ethanol, as well as the aging process, is discussed. PMID- 10699355 TI - Induction of apoptosis by a novel copper-based anticancer compound, casiopeina II, in L1210 murine leukaemia and CH1 human ovarian carcinoma cells. AB - The activity of casiopeina II [Cu(1,4-dimethyl-1, 10 phenanthroline)(glycine)NO(3)], a novel anticancer agent, was tested in two cell lines, L1210 murine leukaemia, CH1 human ovarian carcinoma, cisplatin-resistant and sensitive. Exposure of the cells to a range of concentrations of casiopeina II indicates that this copper complex kills cells by apoptosis and necrosis. Condensed chromatin and nuclear fragmentation were observed after exposure to casiopeina II. The caspase inhibitor Z-Val-Ala-DL-Asp-fluoromethylketone (Z-VAD FMK) almost completely inhibited apoptosis induced by cisplatin; however, casiopeina II-induced apoptosis was inhibited only by 50-70%. These data are consistent with caspase activation (measured by Z-Asp-Glu-Val-Asp-7-amino-4 trifluoromethylcoumarin; Z-DEVD-AFC) by casiopeina II and cisplatin and confirm that caspases are activated in the apoptotic cell death induced by casiopeina II. DNA fragmentation was observed in L1210 cells, but not in CH1 cells. No difference in susceptibility to induction of apoptosis by casiopeina II was found between sensitive and cisplatin resistant cells. In this work we show that the novel copper-based antineoplastic agent casiopeina II is highly active against murine and human cancer cell lines, including cell lines resistant to cisplatin. PMID- 10699356 TI - Inhibition of NF-kappaB-DNA binding by mercuric ion: utility of the non-thiol reductant, tris(2-carboxyethyl)phosphine hydrochloride (TCEP), on detection of impaired NF-kappaB-DNA binding by thiol-directed agents. AB - Mercuric ion (Hg(2+)), a potent thiol inhibitor, prevents expression of nuclear factor kappaB (NF-kappaB) by mercaptide bond formation with a critical cysteine moiety (cys(62)) on the p50 subunit required for DNA binding. NF-kappaB-DNA binding is typically measured in reaction mixtures in which dithiothreitol (DTT) or other thiol reductants are used to maintain cys(62) in the reduced state. However, the presence of thiol reductants prevents accurate assessment of the Hg(2+) concentration required to prevent NF-kappaB-DNA binding because of competitive mercaptide bond formation. In the present studies we evaluated the efficacy of tris(2-carboxyethyl)phosphine-HCl (TCEP), a non-thiol reducing agent which does not bind Hg(2+), on NF-kappaB-DNA binding in vitro, using recombinant p50 protein and a (32)P-labelled kappaB oligonucleotide. We also measured the minimal Hg(2+) concentration required to prevent this interaction in the presence of either reagent. DTT promoted NF-kappaB-DNA binding in concentrations from 0.25 to 2.6mM in binding reactions. However, in the presence of 0.25mM DTT, inhibition of NF-kappaB binding was seen only at Hg(2+) concentrations greater than 100 microM and results were highly variable. In contrast, TCEP promoted NF-kappaB-DNA binding in a dose-related manner in concentrations from 0.25 to 6mM. In the presence of even 6mM TCEP, Hg(2+) prevented NF-kappaB-DNA binding at concentrations as low as 20 microM in binding reactions. Similar findings were observed with regard to the thiol alkylating agent N-ethylmaleimide (NEM). These findings demonstrate the utility of TCEP as reductant in nuclear binding reaction assays involving the interaction of thiol constituents. They also demonstrate inhibition of NF-kappaB-DNA binding at Hg(2+) concentrations comparable to those known to initiate toxicity and apoptotic cell death in vivo. PMID- 10699357 TI - Effect of PSC 833, verapamil and amiodarone on adriamycin toxicity in cultured rat cardiomyocytes. AB - Primary cultures of heart myocytes from neonatal rats were used as an in vitro cardiac cell system to study the effects of the p-170kDa glycoprotein (Pgp) blockers PSC 833 [(3'-keto-Bmt1)-(Val2)-cyclosporine], verapamil and amiodarone on adriamycin cardiotoxicity. Immunostaining revealed the presence of Pgp in the cardiomyocytes. Adriamycin induced a concentration-dependent increase in creatine kinase (CK) leakage, a parameter indicating cell death. None of the Pgp blockers was toxic up to 10 microM, but amiodarone markedly increased CK leakage at 25 microM. 1 microM of the Pgp blockers did not increase adriamycin induced CK leakage, whereas 10 microM of the Pgp blockers significantly augmented adriamycin induced CK leakage. In parallel, cytoplasmic vacuolization and plasma membrane disruptions were observed. Frequency of contraction of cardiomyocytes, as determined by digital image analysis, was concentration-dependently decreased by adriamycin. 1 microM PSC 833 had no additional effect on contractility, only 10 microM PSC 833 enhanced the impairment of contractility induced by adriamycin. Amiodarone and verapamil alone and in combination with adriamycin already at concentrations of 1 microM completely blocked contractility of cardiomyocytes. The results suggest that the increased toxicity of adriamycin in the presence of amiodarone, verapamil and PSC 833 is mediated by an effective blockage of the Pgp efflux pump. The results further indicate that the combination of adriamycin and PSC 833 might be better tolerated with regard to cardiac side-effects, than the combination of adriamycin and verapamil or amiodarone. PMID- 10699359 TI - The immunomodulatory effect(s) of lead and cadmium on the cells of immune system in vitro. AB - A number of studies documented that the heavy metals are not only toxic for the organisms but they may modulate immune responses. The immunomodulatory activity was proved in several in vivo and in vitro model systems. In the current study, immunomodulatory activities of lead and cadmium are presented. The viability of both lymphocytes and macrophages was affected by heavy metals in a dose- and time dependent manner. In the case of lead, the depression of N-oxide production closely correlated with increased blast transformation of spleen cells induced by concanavalin A (ConA). On the contrary, cadmium suppressed the production of N oxides but stimulated significantly the proliferation of spleen cells. The production of cytokines by lymphocytes and macrophages was dependent on the in vitro model used. Generally, the treatment of macrophages with lead results in disregulation of the production of proinflammatory cytokines [tumour necrosis factor alpha (TNF-alpha), interleukin 1alpha (IL-1alpha) and interleukin 6 (IL 6)] and preferential production of Th1 type of cytokines (IFN-gamma and IL-2). Cadmium seemed to trigger the Th2 cytokine regulatory pathway [interleukin 4 (IL 4), interleukin 10 (IL-10)]. The results suggest the metal-induced changes in immunoregulatory mechanism of host with potentially severe clinical consequences. PMID- 10699360 TI - Gap junction intercellular communication and cytotoxicity in normal human cells after exposure to smoke condensates from cigarettes that burn or primarily heat tobacco. AB - Heating tobacco, rather than burning it, reduces tobacco combustion and pyrolysis products. This study tested the hypothesis that the simplified smoke chemistry of a cigarette which primarily heats tobacco (TOB-HT) significantly reduces the potential to alter the structure or function of cellular plasma membranes relative to low "tar" 1R4F and ultra low "tar" lR5F Kentucky reference cigarettes which burn tobacco. Gap junction intercellular communication (GJIC) and lactate dehydrogenase release (LDH) were used to quantify functional and structural changes to the plasma membrane, respectively. Cigarette smoke condensate (CSC) from the mainstream smoke of TOB-HT, lR4F and 1R5F cigarettes were compared in the GJIC and LDH release assays following a 1-hr exposure in vitro. Human bronchial/tracheal epithelial cells, coronary artery endothelial cells, coronary artery smooth muscle cells, foreskin keratinocytes and the WB-344 rat liver epithelial cell line were studied. TOB-HT did not inhibit GJIC in any of the human cell types tested (P0.05) at concentrations where 1R4F and lR5F did inhibit GJIC (P<0.05). TOB-HT did not elevate LDH release (P0.05) when tested at concentrations where lR4F and lR5F did elevate LDH release (P<0.05). Our results suggest that CSC from TOB-HT cigarettes is less damaging to the structure or function of the cellular plasma membranes of a variety of human cell lines than CSC from 1R4F and 1R5F tobacco burning reference cigarettes. PMID- 10699361 TI - Inhibition of dihydrofolate reductase and cell growth activity by the phenanthroindolizidine alkaloids pergularinine and tylophorinidine: the in vitro cytotoxicity of these plant alkaloids and their potential as antimicrobial and anticancer agents. AB - The phenanthroindolizidine plant alkaloids pergularinine (PGL) and tylophorinidine (TPD) isolated from the Indian medicinal herb Pergularia pallida have been evaluated for their biological activity and assessed for the first time employing dihydrofolate reductase (DHFR) (5,6,7,8-THF: NADP(+) oxidoreductase, EC 1.5.1.3) as the probe in the present investigations. The enzyme is a key target in cancer chemotherapy and has been purified from Lactobacillus leichmannii. Cytotoxicity studies showed that both PGL and TPD are potently toxic and inhibited the growth of L. leichmannii cells (IC(50)=45 and 40 microM, respectively). Both the alkaloids significantly inhibited DHFR activity (IC(50)=40 and 32 microM for PGL and TPD, respectively). Alkaloid concentrations greater than 75-95 microM resulted in a complete loss of DHFR activity. Our results are suggestive of the alkaloids as potential antimicrobial and antitumour compounds. Alkaloid binding to DHFR is slow and reversible. Inhibition kinetics revealed K(i) values of 9x10(-6) M and 7x10(-6) M for PGL and TPD, respectively for the enzyme and inhibition in both the cases was a simple linear 'non competitive' type. PMID- 10699362 TI - The acute toxicity of surfactants on fish cells, Daphnia magna and fish-a comparative study. AB - There is a need to replace acute toxicity tests on fish (LC(50)) with more cost effective assays. The main objective of this study was to explore whether gill epithelial cells, hepatocytes and Daphnia magna could be used to predict acute toxicity of surfactants on fish. The acute toxicity of 10 synthetic surfactants (anionic, cationic, nonionic and zwitterionic) and two resin acids were determined on hepatocytes and gill epithelial cells from rainbow trout (Oncorhynchus mykiss), on Daphnia magna and on fish. Cell viability was measured with the fluorescent viability probe calcein-AM, immobilization was determined for Daphnia and 24-hr LC(50) for rainbow trout. The EC(50) values for the cellular tests were clearly higher than the corresponding values for Daphnia and fish, indicating that the cellular tests with the endpoint used are less sensitive than whole organisms. A combination of the EC(50) values for Daphnia and freshly isolated gill epithelial cells in suspension showed, however, a good correlation with acute toxicity on fish (r(2)=0.91 and slope=1.09). The combination seems to be a promising in vitro alternative to acute toxicity tests on fish (LC(50)), but a more exhaustive comparison, including a broad spectrum of chemicals should be made before the predictive value of the combined in vitro test can be evaluated. PMID- 10699363 TI - Stabilization of cytochrome P4502E1 protein by ethanol in primary hamster hepatocyte cultures. AB - We report the effect of ethanol on monooxygenase activities in primary hamster hepatocyte cultures maintained on collagen-coated dishes. The addition of 50mM ethanol to cell cultures both from control and ethanol pretreated animals almost completely maintained, at least for 72hr, the P4502E1-dependent aniline hydroxylase (AnH) activity and the 2E1 immunodetectable apoprotein content at the levels of the corresponding 4-hr plated hepatocytes. On the contrary, other P450 dependent monooxygenase activities, as assayed by testosterone hydroxylases, kept decreasing falling-after 72hr of culture-to the levels of the 4-hr plated hepatocytes. In both cases, in the absence of ethanol, a rapid decline of AnH activities and 2E1 apoprotein contents were also observed, attaining undetectable levels at 72hr. The hybridizable 2E1 mRNA also rapidly declined in both cultures, but such decline was not significantly altered by the presence of 50mM ethanol in the culture medium. Furthermore, we show that P4502E1 in the liver possesses a rapid degradation phase with a half-life of about 6hr. Thus, in the hamster, P4502E1 appears regulated at post-translational level, as in rat, probably by a protein stabilization mechanism. PMID- 10699364 TI - A QSAR model for the eye irritation of cationic surfactants. AB - A QSAR model for the eye irritation of cationic surfactants has been constructed using a dataset consisting of the maximum average scores (MAS-accordance to Draize) for 29 in vivo rabbit eye irritation tests on 19 different cationic surfactants. The parameters used were logP (log [octanol/water partition coefficient]) and molecular volume (to model the partition of the surfactants into the membranes of the eye), logCMC (log critical micelle concentration-a measure of the reactivity of the surfactants with the eye) together with surfactant concentration. The model was constructed using neural network analysis. MAS showed strongly positive, non-linear correlations with surfactant concentration and logCMC and a strongly negative, non-linear correlation with logP. The Pearson correlation between the actual and predicted values of MAS was 0.838 showing that around 70% (r(2)=0.702) of the variance in the dataset is explained by the model. This value is consistent with levels of biological variability reported historically for the Draize rabbit eye test. The relationship provides a potentially useful prediction model for the eye irritation potential of new or untested cationic surfactants with physicochemical properties lying within the parameter space of the model. PMID- 10699365 TI - Release of hydrogen peroxide by rat type II pneumocytes in the prolonged culture. AB - Type II pneumocytes (T II pneumocytes) produce hydrogen peroxide (H(2)O(2)), which may be potentially dangerous for the lung. These cells in culture differentiate to type I-like pneumocytes and it may reflect the differentiation which follows the injury of alveolar epithelium. This work was undertaken to estimate the H(2)O(2) release by T II pneumocytes, freshly isolated and cultured up to 8 days. The light and electron microscopy evaluation confirmed the differentiation of T II pneumocytes to type I-like cells. The release of H(2)O(2), estimated spectrofluorimetrically as homovanillic acid oxidation product obtained in the presence of horseradish peroxidase, was significantly higher at day 4 (0.63+/-0. 68nmol/mg protein/min, P2 S.D.s above the normal mean. These hyperuricosuric autistic individuals may comprise approx. 20% of the autistic population. In order to determine the metabolic basis for urate overexcretion in these patients, de novo purine synthesis was measured in the cultured skin fibroblasts of these patients by quantification of the radiolabeled purine compounds produced by incubation with radiolabeled sodium formate. For comparison, de novo purine synthesis in normal controls, in normouricosuric autistic patients, and cells from patients with other disorders in which excessive uric acid excretion is seen was also measured. These experiments showed that de novo purine synthesis is increased approx. 4-fold in the hyperuricosuric autistic patients. This increase was less than that found in other hyperuricosuric disorders. No unusual radiolabeled compounds (such as adenylosuccinate) were detected in these experiments, and no gross deficiencies of radiolabeled nucleotides were seen. However, the ratio of adenine to guanine nucleotides produced by de novo synthesis was found to be lower in the cells of the hyperuricosuric autistic patients than in the normal controls or the cells from patients with other disorders. These results indicate that the hyperuricosuric subclass of autistic patients have increased de novo purine synthesis, and that the increase is approximately that expected for the degree of urate overexcretion when compared to other hyperuricosuric disorders. No particular enzyme defect was suggested by either gross deficiency of a radiolabeled compound or the appearance of an unusual radiolabeled compound, and no potentially neurotoxic metabolites were seen. Although an enzyme defect responsible for the accelerated purine synthesis was not identified, the abnormal ratio of adenine to guanine nucleotides suggests a defect in purine nucleotide interconversion. PMID- 10699371 TI - Alterations of Na(+) homeostasis in hepatocyte reoxygenation injury. AB - Reperfusion injury represents an important cause of primary graft non-function during liver transplantation. However, the mechanism responsible for cellular damage during reoxygenation has not yet been completely understood. We have investigated whether changes in intracellular Na(+) distribution might contribute to cause hepatocyte damage during reoxygenation buffer after 24 h of cold storage. Hepatocyte reoxygenation resulted in a rapid increase in cellular Na(+) content that was associated with cytotoxicity. Na(+) accumulation and hepatocyte death were prevented by the omission of Na(+) from the incubation medium, but not by the addition of antioxidants. Blocking Na(+)/H(+) exchanger and Na(+)/HCO(3)( ) co-transporter by, respectively, 5-(N,N-dimethyl)-amiloride or omitting HCO(3)( ) from the reoxygenation medium significantly decreased Na(+) overload and cytotoxicity. Stimulation of ATP re-synthesis by the addition of fructose also lowered Na(+) accumulation and cell death during reoxygenation. A significant protection against Na(+)-mediated reoxygenation injury was evident in hepatocytes maintained in an acidic buffer (pH 6.5) or in the presence of glycine. The cytoprotective action of glycine or of the acidic buffer was reverted by promoting Na(+) influx with the Na(+)/H(+) ionophore monensin. Altogether, these results suggest that Na(+) accumulation during the early phases of reoxygenation might contribute to liver graft reperfusion injury. PMID- 10699372 TI - Q-band EPR investigations of neuromelanin in control and Parkinson's disease patients. AB - New insights into the understanding of the changes induced in the iron domain of neuromelanin (NM) upon development of Parkinson's disease (PD) have been gained by electron paramagnetic spectroscopy (EPR). The results of this study are compared with a previously reported variable temperature analysis of X-band EPR spectra of a NM specimen obtained from control brain tissues. The availability of high sensitivity instruments operating in the Q-band (34.4 GHz) allows us to deal with the low amounts of NM available from PD brains. The organization of iron in NM is in the form of polynuclear superparamagnetic/antiferromagnetic aggregates, but the lack of one or more signals in the EPR spectra of NM from PD suggests that the development of the pathology causes NM to decrease its ability to bind iron. Furthermore, the detection of the Mn(II) signal in the Q-band spectra is exploited as an additional internal probe to assess minor structural differences in iron domains of PD and control NM specimens. PMID- 10699373 TI - Measurements of the dielectric properties of peripheral blood mononuclear cells and trophoblast cells using AC electrokinetic techniques. AB - The separation of trophoblast cells from the maternal circulation could provide a valuable diagnostic tool for prenatal diagnosis of genetic abnormalities. This has been attempted using antibody methods, but due to non-specificity of the antibodies, maternal cell contamination remains a problem. We have investigated the potential of dielectrophoretic separation methods as a means of isolating trophoblast cells from mixed peripheral blood mononuclear cells. To determine the potential of this method the dielectric properties of trophoblast cells and mixed peripheral blood mononuclear cells were measured using dielectrophoretic crossover and single cell electrorotation methods. Both dielectrophoretic crossover data and electrorotation data gave an average specific membrane capacitance of the peripheral blood mononuclear cells of 11.5 mF m(-2). Trophoblast cells prepared using three different methods had a higher average specific membrane capacitance in the range 13-18 mF m(-2). The differences in capacitance between the cell types could be exploited as the basis of an AC electrokinetic-based system for the separation of trophoblast cells from peripheral blood mononuclear cells. PMID- 10699374 TI - The mouse N-acetylgalactosamine-6-sulfate sulfatase (Galns) gene: cDNA isolation, genomic characterization, chromosomal assignment and analysis of the 5'-flanking region. AB - Deficiency of lysosomal enzyme N-acetylgalactosamine-6-sulfate sulfatase (GALNS) leads to mucopolysaccharidosis IV A (MPS IV A), for which there is no definitive treatment so far. Although a number of mutations of the GALNS gene of MPS IV A patients have been described, pathogenesis of the disorder still remains elusive. In order to facilitate in vivo studies using model animals for MPS IV A, we isolated and performed molecular characterization of the mouse homolog of human GALNS. The 2.3-kb cDNA contains a 1560-bp open reading frame encoding 520 amino acid residues. The coding region has 84% similarity to the human GALNS cDNA at amino acid level. The mouse Galns gene was mapped by interspecific backcross analysis to the distal region of chromosome 8 where it co-segregates with Aprt. Northern blot analysis showed a wide expression of a single-copy gene, being higher especially in liver and kidney. The Galns gene was isolated from S129vJ genomic library and its genomic organization was characterized. The mouse Galns gene was about 50-kb long and organized into 14 exons and 13 introns. All intron exon splice junctions conformed to the GT/AG consensus sequence except exon 8/intron 8 junction. Primer extension shows multiple transcription initiation sites between -44 and -75 although major transcription initiation site was observed at -90 bp from the ATG codon. The 5'-flanking region lacks canonical TATA and CAAT box sequences, but is G+C rich with 10 GC boxes (potential Sp1 binding sites), characteristic of a housekeeping gene promoter. PMID- 10699375 TI - Succinylpurinemic autism: increased sensitivity of defective adenylosuccinate lyase towards 4-hydroxy-2-nonenal. AB - We studied the effect of trans-4-hydroxy-2-nonenal on the wild-type human adenylosuccinate lyase and on the enzyme from a patient compound-heterozygous for two missense mutations (P75A/D397Y; McKusick 103050.0003/103050.0004). Both the enzymes were inhibited by 10-50 microM trans-4-hydroxy-2-nonenal in a concentration-dependent manner by means of a mixed-type co-operative mechanism. A significantly stronger inhibition was noticed in the presence of the defective enzyme. Nonanal and trans-2,3-nonenal inhibited the enzymes to a less extent and at about 10-times higher concentrations. Hydroxylamine reversed the inhibition by trans-4-hydroxy-2-nonenal, trans-2,3-nonenal or nonanal in the case of the wild type enzyme, but it was ineffective to reverse the inhibition by trans-4-hydroxy 2-nonenal on the defective enzyme. Dithiothreitol slightly decreased the inhibition exerted by trans-4-hydroxy-2-nonenal on both the wild-type and the defective adenylosuccinate lyase, while it did not produce practically any change in the presence of trans-2,3-nonenal or nonanal. PMID- 10699377 TI - Corrigendum to: 'Molecular basis of carbohydrate-deficient glycoprotein syndromes type I with normal phosphomannomutase activity'. PMID- 10699376 TI - Iron overload in paediatrics undergoing cardiopulmonary bypass. AB - Pathological changes in iron status are known to occur during bypass and will be superimposed upon physiological abnormalities in iron distribution, characteristic of the neonatal period. We have sought to define the severity of iron overload in these patients. Plasma samples from 65 paediatric patients undergoing cardiopulmonary bypass (CPB) were analysed for non-haem iron, total iron binding capacity, transferrin and bleomycin-detectable iron. Patients were divided into four age groups for analysis. Within each age group, patients who were in iron overload at any time point were statistically compared to those who were not. The most significant changes in iron chemistry were seen in the plasma of neonates, with 25% in a state of plasma iron overload. 18.5% of infants and 14.3% of children at 1-5 years were also in iron overload at some time point during CPB. No children over 5 years, however, went into iron overload. Increased iron saturation of transferrin eliminates its ability to bind reactive forms of iron and to act as an antioxidant. When transferrin is fully saturated with iron, reactive forms of iron are present in the plasma which can stimulate iron-driven oxidative reactions. Our data suggest that paediatric patients are at greater risk of iron overload during CPB, and that some form of iron chelation therapy may be advantageous to decrease oxidative stress. PMID- 10699378 TI - A new method for preparing biodegradable microparticles and entrapment of hydrocortisone in DL-PLG microparticles using supercritical fluids. AB - An improved process for the production of polymeric microparticles, based on solution-enhanced dispersion by supercritical fluids (SEDS) using a combination of supercritical N(2) and CO(2), was evaluated. The biodegradable polymers, poly(D,L-lactide-co-glycolide): copolymer composition 50:50 (DL-PLG), poly(L lactide) (L-PLA), poly(D,L-lactide) (DL-PLA) and polycaprolactone, were used for preparation of microparticles by a modified SEDS process. Solutions of the polymers in organic solvents were dispersed and the solvent was extracted with supercritical CO(2) and N(2). The morphology, the size distributions and degree of hydrocortisone entrapment were determined. The combination of supercritical N(2) and CO(2) led to a more efficient dispersion of the polymer solutions than CO(2) alone. This resulted in a reduction of particle size of the microparticles produced from all of the amorphous polymers. Discrete spherical microparticles with a mean volumetric diameter of less than 10 microm were produced from DL-PLG, DL-PLA and L-PLA. Hydrocortisone was successfully entrapped within the DL-PLG microparticles. The modified SEDS process improved the dispersion of amorphous polymer solutions resulting in formation of small spherical microparticles. The SEDS process can be used for incorporation of drugs into the DL-PLG microparticles. PMID- 10699379 TI - Stereoselective pharmacokinetics of propafenone and its major metabolites in healthy Chinese volunteers. AB - The stereoselective pharmacokinetics of propafenone (PPF) and its active metabolite 5-hydroxypropafenone (5-OHP) as well as their glucuronide and sulfate conjugates have been investigated, in order to clarify the relationship between metabolism and stereoselective disposition of PPF in humans. After oral administration of 300 mg racemic PPF hydrochloride to 10 healthy Chinese subjects, the areas under the plasma concentration-time curves (AUCs) for (S)-PPF were significantly higher (S/R ratio, 1.50+/-0.17) and the apparent oral clearance significantly lower (S/R ratio, 0.68+/-0.07) than those parameters for (R)-PPF. In contrast, the AUCs of PPF glucuronide (PPF-G) were lower for (S)-PPF G than for the (R)-enantiomer (S/R ratio, 0.83+/-0.12). The partial clearance of (S)-PPF by glucuronidation pathway was lower than that of (R)-PPF and the enantiomeric ratio was 0.62+/-0.04. The t(max) values of PPF glucuronide diastereoisomers showed no statistically significant differences between each other, but were much shorter than the corresponding values of the parent drug, implying that glucuronidation may be the 'first-choice' pathway in presystemic metabolism of PPF. Glucuronidation of 5-OHP favored the (S)-enantiomer, whereas the sulfation showed a large preference for the (R)-enantiomer. After beta glucuronidase hydrolysis, no significant differences were observed in AUCs between 5-OHP enantiomers (including unconjugated and conjugated 5-OHP). The results suggest that the significant difference in disposition between PPF enantiomers may be, at least in part attributed to stereoselective metabolism in the glucuronidation pathway. PMID- 10699380 TI - Caffeine and nicotinamide enhances the aqueous solubility of the antimalarial agent halofantrine. AB - The aqueous solubility of the antimalarial agent halofantrine in phosphate buffers pH 5.9 and 7.0 (ionic strength 0.08) is increased by the addition of caffeine and nicotinamide. Solubility is increased to a greater extent in the presence of caffeine (12.5-125 mM) than nicotinamide (125 mM -2.0 M). The greatest increase in solubility was observed at pH 5.9 where the basal solubility of halofantrine rose from 0.91 to 435 microM when 125 mM caffeine was added. Phase solubility studies support the formation of a 1:1 complex between caffeine and halofantrine which is characterised by a K(1:1) constant of 2.75x10(3) M(-1) (pH 5.9). A less stable 1:1 complex is formed at pH 7.0 (K(1:1)=6.37x10(3) M( 1)). Differential scanning calorimetry of solid mixtures of caffeine and halofantrine showed the absence of the endotherms of the two drugs and the appearance of a distinct endotherm (with a smaller enthalpy) characteristic of the complex. An analysis of the 1H-NMR spectra of mixtures of caffeine and halofantrine revealed perturbations in the chemical shifts of the methyl group and proton at positions 4 and 8 of caffeine, and a change in splitting pattern of the H(9) proton of the phenanthrene ring in halofantrine. PMID- 10699381 TI - Carbonic anhydrase activators. Part 24. High affinity isozymes I, II and IV activators, derivatives of 4-(4-chlorophenylsulfonylureido-amino acyl)ethyl-1H imidazole. AB - N-1-Tritylsulfenyl histamine was synthesized by reaction of histamine (Hst) with tetrabromophthalic anhydride followed by protection of its imidazole moiety with tritylsulfenyl chloride. After hydrazinolysis, it afforded a key intermediate which was derivatized at the aminoethyl group in order to obtain new types of activators of the zinc enzyme carbonic anhydrase (CA, EC 4.2.1.1). Reaction of the key intermediate with 4-chlorophenylsulfonylureido amino acids (cpu-AA) in the presence of carbodiimides afforded, after deprotection of the imidazolic nitrogen atom, a series of compounds with the general formula cpu-AA-Hst (cpu, 4 ClC(6)H(4)SO(2)NHCO). Some structurally related dipeptide derivatives with the general formula cpu-AA1-AA2-Hst (AA, AA1 and AA2 represent amino acyl moieties) were also prepared by a strategy similar to that applied for the amino acyl compounds mentioned above. The new derivatives proved to be efficient activators of three CA isozymes. Best activity was detected against hCA I and bCA IV, for which some of the new compounds showed affinities in the 1-10 nM range (h, human; b, bovine isozymes). hCA II, on the other hand, was less prone to activation by the new derivatives, which possessed affinities around 20-50 nM for this isozyme. This new class of CA activators might lead to the development of drugs/diagnostic agents for CA deficiency syndrome, a genetic disease of bone, brain and kidneys. PMID- 10699382 TI - Development of pectin matrix tablets for colonic delivery of model drug ropivacaine. AB - The objective of this work was to develop pectin-based matrix tablets for colonic delivery of the model drug ropivacaine, with the future perspective of radiolabelling the system by neutron activation technique for a gamma scintigraphic study. The aim was to investigate some formulation factors that could reduce the release of the drug in the simulated gastric and intestinal fluids, increase the release in the simulated cecal fluid (with pectinolytic enzymes) and improve the poor compactibility of pectins. For dissolution studies, the flow-through apparatus with sequential dissolution liquids simulating the mouth-to-colon conditions was used. The effect of two pectin types, the incorporation of ethylcellulose as a dry matrix-additive and water or ethanol as granulation liquids were investigated in a study designed as a D-optimal mixture. Amidated pectin (Am.P) produced harder tablets than the calcium salt of pectin (Ca.P) and was more susceptible to enzymatic degradation. Addition of ethylcellulose increased the tablet strength and the dissolution rate. Furthermore, directly compressed Am.P tablets were produced by addition of coarse or micronised qualities of ethylcellulose. The latter improved the crushing strength markedly imposing a marginal release-reducing effect. Coating this formulation with Eudragit((R)) L 100 reduced the release in the simulated upper GI conditions without interference with the subsequent enzymatic activity. PMID- 10699384 TI - Protease inhibitors. Part 7. Inhibition of Clostridium histolyticum collagenase with sulfonylated derivatives of L-valine hydroxamate. AB - Sulfonylated L-valine hydroxamate derivatives were obtained by reaction of alkyl/arylsulfonyl halides with the title amino acid, followed by treatment with benzyl chloride, and conversion of the COOH moiety to the CONHOH group. Other derivatives were obtained by reaction of N-benzyl-L-valine with arylisocyanates, arylsulfonylisocyanates or benzoylisothiocyanate, followed by the similar conversion of the COOH into the CONHOH moiety, with hydroxylamine in the presence of carbodiimides. The obtained compounds were assayed as inhibitors of the Clostridium histolyticum collagenase, ChC (EC 3.4.24.3), a zinc enzyme which degrades triple helical collagen. The hydroxamate derivatives were generally 100 500 times more active than the corresponding carboxylates. In the series of synthesized derivatives, substitution patterns leading to best ChC inhibitors were those involving perfluoroalkylsulfonyl- and substituted-arylsulfonyl moieties, such as pentafluorophenylsulfonyl; 3- and 4-protected aminophenylsulfonyl-; 3- and 4-carboxyphenylsulfonyl-; 3 trifluoromethylphenylsulfonyl; or 1- and 2-naphthyl among others. Similarly to the matrix metalloproteinase hydroxamate inhibitors, ChC inhibitors of the type reported here must incorporate hydrophobic moieties at the P(2') and P(3') subsites, in order to achieve tight binding to the enzyme. Such compounds might lead to drugs useful in the treatment corneal bacterial keratitis. PMID- 10699383 TI - Evaluation of strength-enhancing factors of a ductile binder in direct compression of sodium bicarbonate and calcium carbonate powders. AB - This study evaluated the effectiveness of a ductile binder in direct compression of sodium bicarbonate and calcium carbonate powders. Properties associated with both the binder and the compound were studied. The addition of binder materials, such as polyethylene glycols (PEGs) of differing molecular weights or microcrystalline cellulose, generally resulted in an increase in the axial tensile strength of the corresponding compacts. The increase in tablet strength was generally greater with the PEGs than with microcrystalline cellulose. The results indicate that the improvement in tablet strength caused by the binder is dependent on properties of both the binder and the compound. By utilising different methods it was established that the fracture during tablet strength testing mainly occurred around the compound particles. As a consequence of this, it appears that the ability of the binder to fill the voids between the compound particles is a determinative factor for increasing tablet strength. The binder appeared to have less effect when added to compounds that fragmented during compaction. Characteristics of the binder resulting in the greatest decrease in porosity, and thus the greatest increase in the tensile strength of the compound, included a high degree of plastic deformation with a limited elastic component and a small particle size. Obviously, the amount of binder added to the mixture also affected the results. PMID- 10699385 TI - Near IR spectroscopy to quantify the silica content and difference between silicified microcrystalline cellulose and physical mixtures of microcrystalline cellulose and silica. AB - Silicified microcrystalline cellulose (SMCC) has been shown to have advantages over conventional microcrystalline cellulose (MCC). These advantages are (i) improved tablet strength compared to that achieved with MCC, (ii) the retention of compressibility after wet granulation, whereas MCC produces weaker tablets after wet granulation, and (iii) superior flow properties than MCC. In this study near IR spectroscopy has been used to study MCC, SMCC (with different loadings of colloidal silicon dioxide, CSD) and physical mixtures of MCC and CSD. It was found that even though SMCC and MCC were very similar, there was a region of the near IR spectra (second derivative peak at 2194 nm) where a distinctive response was seen for SMCC. The size of the peak was proportional to the CSD content for the co-processed SMCC samples. The peak was not present to the same extent for physical mixtures. A combination of near IR and a test for total silica content would make it possible to discern whether microcrystalline cellulose samples were SMCC material or simple physical mixtures. PMID- 10699386 TI - How dopamine was recognised as a neurotransmitter: a personal view. AB - Evidence of the neurotransmitter role of dopamine has come from research employing a variety of modalities. These have made use of the techniques of pharmacology, toxicology, clinical pathology, clinical chemistry and experimental neurology. This article deals more specifically with the contributions of the author and his collaborators in establishing the function of dopamine as the chemical mediator of the nigrostriatal tract PMID- 10699387 TI - Parkinson's disease subtypes: clinical classification and ventricular cerebrospinal fluid analysis. AB - The current study presents preliminary data regarding the development and validation of a rating system designed to classify PD patients into clinical subgroups. Using portions of the Unified Parkinson's Disease Rating Scale, a ratio value was derived, yielding three patient subtypes: a tremor-dominant group (T), an akinetic-rigid group (AR), and a mixed group (MX). Validation of the schema was conducted by grouping PD surgical candidates into specific disease subtypes and evaluating differences in neurotransmitter profiles among disease subtypes and non-PD neurological controls. High pressure liquid chromatography analysis of ventricular cerebrospinal fluid indicated 5-hydroxyindoleacetic acid was significantly lower in the AR and MX groups compared to non-PD controls; whereas, glycine was significantly higher in the AR group compared to the T, MX, and control groups. The results suggest that an operational approach can be utilized to differentiate between PD subtypes with distinct neurochemical profiles. PMID- 10699388 TI - A quantitative surface electromyogram analysis for diagnosis and therapy control in Parkinson's disease. AB - Computer analysis of EMG data on tonic and phasic activities of mm. biceps and triceps brahii was performed to evaluate objectively Parkinson's disease (PD) symptoms and to quantify levodopa therapy effects. Fifteen patients were evaluated in the OFF and eleven in the ON states. Ten healthy controls were also studied. The following EMG parameters were examined: average and maximal amplitudes at rest, occurrence of burst muscle discharges (BMD) with a frequency of 4-7Hz, phasic activation coefficients (PhAC) of the voluntarily contracting flexors and reflex agonist/antagonist muscle involvement under voluntary movement or tonic strain. Statistically significant correlations of resting EMG amplitudes and PhACs with the part III UPDRS motor scores were found. However, the level of antagonist muscle involvement correlated specifically with the part II UPDRS and dyskinesia (disability) scores. Treatment with levodopa produced a clear positive effect on resting amplitudes, PhAC values and BMD occurrence. But in some cases levodopa caused an enhancement of agonist and antagonist muscle involvement, which may be an objective indicator of the risk for developing drug-induced dyskinesia in PD patients. PMID- 10699389 TI - Torque variations during repeated passive isokinetic movements of the knee in subjects with Parkinson's disease and healthy control subjects. AB - The purpose of this study was to quantify response variations during isokinetic passive movements of the knee in subjects with Parkinson's disease. Parkinsonian patients demonstrated a greater decrease of resistive torque compared to healthy control subjects, particularly in tests at higher velocities and during knee flexion movements. Responses were influenced by electromyographic activity in stretched and shortened muscle groups and also by mechanical factors. The results indicate that repetition of movements needs to be taken into account when measuring hypertonia in parkinsonian subjects. PMID- 10699390 TI - Middle latency auditory evoked potentials in patients with parkinsonism. AB - The aim of this study was to assess the middle latency auditory evoked potential (MLAEP) findings in idiopathic Parkinson's disease (IPD) and in patients who are regarded as having atypical parkinsonian disorders (AP) and to determine whether MLAEPs could contribute to the differential diagnosis of IPD and AP.MLAEPs were evaluated in 19 control subjects and in a total of 35 patients with parkinsonism, of which 27 had IPD and 8 had AP. Among IPD patients, P1 was absent in 1 nondemented patient with young-onset Parkinson's disease (YOPD) (5%) and in 2 of 7 demented (28.5%) IPD patients. In the AP group, 3 of the 7 (42.8%) nondemented patients and the one patient with dementia showed the absence of P1. The absence of P1 was found to be significantly higher in AP patients than IPD patients (p=0.0335).In conclusion, MLAEPs were found to be normal in nondemented IPD patients with only a few exceptions. The absence of P1 in nondemented patients with parkinsonian symptoms may bring the diagnosis of IPD into question. The absence of P1 could be detected in AP patients at least as often as in demented IPD patients. Thus, the measurement of MLAEPs may be a clinically useful adjunct to the clinical examination of patients with parkinsonism. PMID- 10699391 TI - Clinical and electromyographic characteristics of tremor in patients with depression. AB - The aim of this investigation was to establish the clinical and electromyographic pattern of tremor in patients with depression.Twenty-eight patients with depression and tremor and 30 patients with tremor and generalized anxiety disorder were investigated. Tremor was scored clinically by the Webster Tremor Scale. Electromyographic examination of tremor activity from antagonistic hand muscles was performed.Our results revealed in both groups of patients a postural and kinetic tremor with characteristics of an enhanced physiological tremor. Tremor involved only both upper limbs and no other body parts in all patients.In conclusion tremor in depression and generalized anxiety disorder is an enhanced physiological tremor. PMID- 10699392 TI - Hyperkinesia contralateral to acute hemiplegia: relevance of previous frontal lesions. AB - Two patients with a hyperkinetic syndrome contralateral to acute hemiplegia are presented. One patient showed a right-sided hemiplegia associated with abnormal movements of the left upper limb. In the other patient hemiplegia was localised on the left side while abnormal movements involved the right lower limb. Brain imaging showed acute infarctions (respectively left middle cerebral artery area and right lacunar infarctions) associated with a pre-existing contralateral infarction involving anterior non-primary motor areas. Hyperkinetic syndromes in stroke are mainly related to acute lesion of the contralateral subcortical areas. In our patients, the acute lesions were located ipsilateral to the hyperkinetic body part while the pre-existing lesions were located contralateral to the hyperkinetic side. We speculated that these pre-existing lesions might play a role in the pathophysiology of this clinical syndrome. PMID- 10699393 TI - Worsening of Parkinson's disease by citalopram. AB - More than 50% of the patients with Parkinson's disease (PD) may experience mood disturbances. Serotonin-selective reuptake inhibitors (SSRI) are very active in the management of depression. Citalopram is a new SSRI increasingly used in the treatment of depression. The question of a negative impact of SSRI on the motor function of patients with PD is an important clinical issue. A number of such observations have published with various SSRI, but none, up-to-now with citalopram. We report the case of a patient with PD who experienced a worsening in the motor status soon after the addition of citalopram to her antiparkinsonian drug regime. This single case report suggests that this potential adverse event is a class related side effect. PMID- 10699394 TI - Hemiparkinsonian syndrome due to a cerebral tumor infiltrating the substantia nigra. AB - Parkinsonism due to a cerebral tumor is rare and usually caused by an indirect effect of supratentorial tumors on nigrostriatal pathways rather than by direct involvement of the substantia nigra. Only three previous cases of midbrain infiltration causing Parkinsonism have been reported. We report the case of a young woman with a left tremo-akineto-rigid parkinsonian syndrome caused by a tumor of the brainstem infiltrating the right substantia nigra. PMID- 10699395 TI - Ictal SPECT in paroxysmal non-kinesigenic dyskinesia. Case report and review of the literature. AB - Purpose: Paroxysmal non-kinesigenic dyskinesia (PNKD) should be included in the list of differential diagnosis in patients with refractory epilepsy. Although the pathophysiological mechanisms that underlie this disorder remain controversial, it is now accepted that the basal ganglia are the anatomical substrate responsible for it.Material and methods: We report a 16-year-old mentally retarded male with PNKD admitted for video-EEG monitoring and ictal SPECT, which showed hyperperfusion on the right caudate and thalamus.Conclusion: This case supports more evidence for the involvement of the caudate nucleus and thalamus in the mechanisms responsible for the production of PNKD. PMID- 10699396 TI - The clinical use of brainstem reflexes and hand-muscle reflexes. AB - Brainstem reflexes and hand-muscle reflexes can be elicited and recorded with routine EMG equipment. Not all these reflexes are useful in clinical neurology. But those that are - the subject of this review - exhibit distinct patterns of abnormality that have clinical diagnostic and localizing value in various diseases, including cranial neuropathies, focal lesions within the cervical cord, brainstem, and brain, movement disorders, and pain. PMID- 10699397 TI - Tracking the development of the N1 from age 3 to adulthood: an examination of speech and non-speech stimuli. AB - OBJECTIVES: To examine developmental changes in the N1a, N1b and N1c evoked by a tone and a speech consonant (/da/). METHODS: Subjects (n=70 for tones; n=69 for /da/) were grouped into 2 year intervals (age 3-16) and adults. They listened to a tone (2 kHz; 36 ms; 77 dB SL; ISI=600 ms; n=346) or a speech consonant /da/ (female voice recording; 7 ms VOT; 212 ms; 72 dB SL; ISI=600 ms; n=349) while watching a Disney((R)) screensaver. EEG was recorded from 26 electrodes referenced to Cz. An averaged reference was computed off-line. Amplitude and latency data were analyzed with repeated-measures ANOVAs for agexelectrode, and agexN1 component, respectively. RESULTS: Left hemisphere N1a was mature before age 3 whereas the right hemisphere N1a matured around 7-8 years. The vertex N1b showed a parietal distribution which shifted anteriorly with age. The N1c showed age- and stimulus-related changes. The N1c measured over the left hemisphere matured earlier than the N1c over the right hemisphere. The N1c to /da/ matured earlier than that to tones. CONCLUSIONS: Auditory processing undergoes steady and subtle developmental changes. These changes follow different maturational patterns depending on the type of stimuli. The evidence suggests earlier development of the left hemisphere and earlier development of the generators underlying speech processing. PMID- 10699398 TI - Differential temporal coding of the vibratory sense in the hand and foot in man. AB - OBJECTIVE: We studied the difference in the temporal tuning function of the vibratory senses between the hand and foot in man by using steady-state somatosensory evoked potentials (S-SEPs) to vibratory stimulation. METHODS: Vibratory stimuli were applied to the palm and sole, and the S-SEPs were then recorded in 8 normal subjects. A total of 200 responses were recorded from 4 electrodes including the ipsi-and contralateral somatosensory areas of the hand and foot. The amplitude of the first harmonic component (1F) was obtained by a Fourier analysis. The effect of modulation frequency (17-30 Hz) on the 1F at a stimulus intensity of 0.05 N was studied. RESULTS: The amplitudes of the S-SEPs were the greatest in the contralateral hand and foot areas. The mean 1F amplitudes of the palm S-SEPs as a function of the modulation frequency showed a narrow tuning curve with a peak near 21 Hz while those of the sole demonstrated a broad tuning curve. CONCLUSION: Our results suggest the differential temporal coding of the vibratory sense in the hand and foot areas of the somatosensory cortex in man. This is probably caused by the different characteristics in the receptors situated in the hand and foot. PMID- 10699399 TI - Dermatome versus homunculus; detailed topography of the primary somatosensory cortex following trunk stimulation. AB - OBJECTIVE: Identification of a detailed topography of the receptive area for each of the thoracic dermatomes in humans using somatosensory evoked magnetic fields (SEF). METHODS: We analyzed the location of the equivalent current dipole (ECD) of SEF following electrical stimulation of the skin at Th4, Th6, Th8, Th10 and Th12 dermatomes in 14 normal subjects. RESULTS: Three deflections, M18, M25 and M40, were obtained within 60 ms of stimulation of Th6, Th8 and Th10 dermatomes. No consistent deflection could be identified following Th4 and Th12 dermatomal stimulation, probably due to a poor signal-to-noise ratio and difficulty in fixing the stimulation electrodes. M18 was absent or small in amplitude. The latency of M25 ranged from short to long in the order Th6, Th8 and Th10 (P<0.05). ECDs of all components for each site stimulation were located in the truncal area of the primary somatosensory cortex. Although the locations of the ECDs tend to be arranged from lateral to medial in the sequence Th6, Th8 and Th10, the difference was not significant. CONCLUSION: The representation area of the trunk is small, and the receptive areas for the stimulation of Th6, Th8 and Th10 dermatomes are considered to be very close or to overlap. PMID- 10699400 TI - A new method to increase nociception specificity of the human blink reflex. AB - OBJECTIVE: The medullary R2 response of the blink reflex can be elicited by innocuous and noxious stimuli. The purpose of this study was to elicit a nociception specific R2 response with a new surface electrode. METHODS: In 10 healthy subjects the blink reflex was elicited using a standard (10-15 mA) and a new concentric surface electrode type (0.6-1.6 mA) which produces a pin-prick like pain. RESULTS: After topical local anaesthesia with lignocaine/prilocaine R1 was unchanged, R2 was attenuated by 12% after standard stimulation but was almost abolished (-91%) with the new electrode type. CONCLUSION: Stimulation with low stimulus intensities but electrode-dependent high current density allows preferential depolarization of superficial nociceptive A-delta fibres. This new method is less traumatic than others and is useful in the study of trigeminal nociception. PMID- 10699401 TI - Topographical characteristics of slow wave activities during the transition from wakefulness to sleep. AB - OBJECTIVES: This study examined the topographical characteristics of slow wave(delta and theta) activities during the transition from wakefulness to sleep, using topographic mapping of EEG power and coherence. METHODS: Sonography of nocturnal sleep was recorded on 10 male subjects. Each recording, from 'lights off' to 5 min after the appearance of the first sleep spindles, was analyzed. Typical EEG patterns during the transition from wakefulness to sleep were classified into 9 stages (EEG stages). RESULTS AND CONCLUSIONS: Topographic maps of coherence in delta-and theta-band activities demonstrated that the synchronous component at the anterior-central areas of the scalp appeared corresponding with increasing power. The populations of high coherence pairs among total pairs were computed for each band and each EEG stage to examine the regional differences of EEG. The populations of the delta-band activity increased clearly from EEG stage 6 in the anterior-central areas. The populations of the theta-band activity increased clearly from EEG stage 7 in the anterior-central areas. These results suggest that the dominant synchronous component of slow wave activities during the transition from wakefulness to sleep increased as a function of EEG stages in the anterior-central areas and increased clearly after the appearance of vertex sharp waves. PMID- 10699402 TI - Sleep onset REM period appearance rate is affected by REM propensity in circadian rhythm in normal nocturnal sleep. AB - OBJECTIVE: REM latency is usually 60-120 min, but under certain conditions, its latency may be less than 25 min, in which it is known as sleep onset REM period (SOREMP). In order to identify the factors responsible for the appearance of SOREMP, we used the nocturnal sleep interruption method to investigate whether REM propensity in normal nocturnal sleep, (i.e. circadian variations in REM sleep related to body temperature rhythm), could affect the rate of SOREMP appearance (SOREMP%). MATERIALS AND METHODS: After two adaptation and one baseline nights, we interrupted the nocturnal sleep, either in the second or the fourth cycle (early and late conditions, respectively) of 16 subjects and compared SOREMP% at the second sleep onset between these conditions by chi(2) test. Rectal temperature was measured. RESULTS: SOREMP% was found to be 58.1% in the early condition and 87.5% in the late condition - a significant difference. Body temperature dropped at the second sleep onset in both conditions but the drops did not differ significantly. CONCLUSION: We concluded that SOREMP% was affected by circadian variations in REM propensity but SOREMP% and body temperature drop did not show a linear relationship. Further studies to discriminate the influence of circadian rhythm factors and slow wave sleep on SOREMP% are called for. PMID- 10699403 TI - Sleep stage-related changes in sympathetic sudomotor and vasomotor skin responses in man. AB - OBJECTIVES: The aim of the study was to evaluate the characteristics of the spontaneous and evoked sympathetic skin responses (SSR) during sleep and wakefulness in comparison with the skin vasomotor responses (SVR). METHODS: Five healthy subjects underwent a night of videopolysomnographic recording. Spontaneous SSR were recorded via surface electrodes placed on the dorsal and ventral aspect of the hand while SVR were evaluated by means of an infrared photoelectric transducer placed on the index finger. SSR and SVR were evoked via electrical stimuli applied to the left supraorbital nerve. RESULTS: Spontaneous SSR frequency was highest during stage 4 of NREM sleep and lowest during REM phases. On the contrary, spontaneous SVR frequency reached its lowest value during stage 4 and its highest value during stage 2 of NREM sleep, remaining at levels above waking values during REM. SSR could be elicited by stimuli inducing arousal during light sleep but it was absent during deep NREM and REM sleep. SVR could be evoked throughout NREM and REM sleep. CONCLUSIONS: Spontaneous SSR and SVR act differently during physiological modifications of vigilance. Evoked SSR is strictly dependent upon the state of vigilance, whereas evoked SVR shows no modifications during the different stages of the wake-sleep cycle. PMID- 10699404 TI - EOG correction: comparing different calibration methods, and determining the number of epochs required in a calibration average. AB - OBJECTIVE: A form of EOG correction called the 'aligned-artifact average' (AAA) method has been advanced by the authors. In contrast to many previous methods, this used a correction coefficient (B), based on an average of eye movements rather than raw data, to remove eye movement related contamination from the EEG. The first part of this study was aimed at determining whether a variation of this procedure that is more easily implemented would produce a similar correction. The second part was designed to determine the number of epochs needed in an average to correct adequately. METHODS: Subjects performed a series of eye movement tasks whilst EEG and EOG data were recorded. Data were manipulated according to either the AAA or an alternate new ERP method (NERP) and the resultant Bs were compared in part A. In part B, averages were created from varying numbers of epochs, and the resultant r(2)-values were compared. RESULTS: The AAA and NERP methods produced the same Bs, and averages with at least 40 epochs were required for adequate B estimation for both VEOG and HEOG. CONCLUSION: There is no difference between the AAA and NERP methods and thus it is acceptable to use the more easily implemented NERP method for EOG correction. It is recommended that when applying this procedure, at least 40 epochs should be used to make up the averages from which to calculate correction coefficients. PMID- 10699405 TI - EOG correction of blinks with saccade coefficients: a test and revision of the aligned-artefact average solution. AB - OBJECTIVE: The 'aligned-artefact average' (AAA) procedure was advanced by the authors as a technique suitable for removing eye movement-related artefacts from the EEG. It was proposed that this method would correct both blink and non-blink artefact from the EEG, using the same set of correction coefficients (Bs). However, recent evidence suggests that this simplification is not always accurate. Thus, we test here a revision of the AAA, including an appropriate allowance for the radial EOG (REOG) component, that does allow the use of the same Bs for the correction of blink and non-blink artefact. METHODS: Blink (and saccade) ERP data from 15 subjects were corrected using the AAA method, with Bs calculated from the same blink (and saccade) data set (referent waveforms), or a different set of blink (and saccade) data, or using the new revised AAA procedure (RAAA). RESULTS: AAA Bs calculated from saccades corrected blinks poorly (and vice versa). However, the RAAA Bs corrected blink ERPs better than blink-derived Bs, and saccade ERPs better than saccade-derived Bs. It was also found that irrespective of correction type, inclusion of REOG improved correction. CONCLUSION: EOG correction is more accurate when the radial channel is included, but inclusion of REOG (and/or HEOG) is not sufficient to resolve the discrepancy between blink and saccade correction. Using the RAAA procedure, both blink and non-blink data can be corrected using the same set of Bs. PMID- 10699406 TI - Is ESES/CSWS a strictly age-related disorder? AB - OBJECTIVES: We report on a case of ESES/CSWS observed in a patient of 21 years and still persisting at the age of 25. Cases of ESES/CSWS have never been previously described in adult patients. ESES/CSWS is considered to be related to the degree of maturation of the central nervous system, and therefore strictly age-related. METHODS: Our case of ESES/CSWS was observed in a 2 1 year old woman referred for cognitive and behavioral disorders. She had previously had epileptic fits, but was seizure free at that time. The patient underwent a full-night polygraphic recording , which showed typical ESES/CSWS pattern, with a Spike-and Wave Index >8 5%. Polysomnography was repeated 9 times in a 4 year follow-up, during which the ESES/CSWS condition persisted, despite the pharmacological treatments. The patient also underwent cerebral magnetic resonance imaging and fludeoxyglucose F 18 positron emission tomography (PET). RESULTS: The PET study revealed reduced metabolic activity within the lower gyrus of the right parietal lobe, but no significant difference between subcortical structures and cortical mantle was seen. MRI scans were normal. CONCLUSIONS: This observation suggests that ESES/CSWS might not always be an age-related condition. Sleep EEG recordings should always be performed in patients with behavioral disorders and a history of epileptic fits. PMID- 10699407 TI - Test-retest reliability of cognitive EEG. AB - OBJECTIVE: Task-related EEG is sensitive to changes in cognitive state produced by increased task difficulty and by transient impairment. If task-related EEG has high test-retest reliability, it could be used as part of a clinical test to assess changes in cognitive function. The aim of this study was to determine the reliability of the EEG recorded during the performance of a working memory (WM) task and a psychomotor vigilance task (PVT). METHODS: EEG was recorded while subjects rested quietly and while they performed the tasks. Within session (test retest interval of approximately 1 h) and between session (test-retest interval of approximately 7 days) reliability was calculated for four EEG components: frontal midline theta at Fz, posterior theta at Pz, and slow and fast alpha at Pz. RESULTS: Task-related EEG was highly reliable within and between sessions (r0.9 for all components in WM task, and r0.8 for all components in the PVT). Resting EEG also showed high reliability, although the magnitude of the correlation was somewhat smaller than that of the task-related EEG (r0.7 for all 4 components). CONCLUSIONS: These results suggest that under appropriate conditions, task-related EEG has sufficient retest reliability for use in assessing clinical changes in cognitive status. PMID- 10699408 TI - The effects of body posture on resting electroencephalographic activity in sleep deprived subjects. AB - OBJECTIVE: This study examined the effects of posture on the resting electroencephalographic (EEG) activity of sleep-deprived volunteers. METHODS: EEG data were collected under two conditions at 13 separate time points. Testing was performed while subjects remained in a normal seated position and then repeated while subjects stood upright. RESULTS: Results indicated that delta and theta activity progressively increased as a function of sleep loss, and that standing upright attenuated this effect. CONCLUSIONS: These results suggest that an upright posture increases EEG arousal and may be useful for counteracting fatigue in sleep-deprived individuals. PMID- 10699409 TI - Negative myoclonus in Creutzfeldt-Jakob disease. AB - OBJECTIVE: To describe electrophysiological findings in a patient with Creutzfeldt-Jakob disease (CJD) showing negative myoclonus. METHODS AND RESULTS: We studied this CJD patient electrophysiologically, in comparison with two patients with cortical reflex positive myoclonus due to benign adult familial myoclonic epilepsy (BAFME). Spontaneous negative myoclonus was associated with periodic synchronous discharges (PSDs) on the electroencephalogram, but negative myoclonus could also be induced by electrical stimulation of the median nerve in the CJD patient. This patient showed giant somatosensory evoked potentials (SEPs) and enhanced C reflexes, and the duration of the induced EMG silences was found to be significantly correlated with the amplitude of cortical SEPs. The duration of silent periods (SPs) produced by magnetic stimulation of the motor cortex was extremely long. The study of recovery function of SEPs suggested that the excitability of the somatosensory cortex was decreased during a long post stimulus period. These findings were clearly different from those of patients with BAFME. CONCLUSIONS: This CJD patient had two types of negative myoclonus; one was associated with PSDs and the other was cortical reflex negative myoclonus. The long-lasting decrease in excitability of the sensorimotor cortices after stimulation could be related to the occurrence of both types of negative myoclonus. PMID- 10699410 TI - Automatic spike detection via an artificial neural network using raw EEG data: effects of data preparation and implications in the limitations of online recognition. AB - OBJECTIVE: Automatic detection of epileptic EEG spikes via an artificial neural network has been reported to be feasible using raw EEG data as input. This study re-investigated its suitability by further exploring the effects of data preparation on classification performance testing. METHODS: Six hundred EEG files (300 spikes and 300 non-spikes) taken from 20 patients were included in this study. Raw EEG data were sent to the neural network using the architecture reported to give best performance (30 input-layer and 6 hidden-layer neurons). RESULTS: Significantly larger weighting of the 10th input-layer neuron was found after training with prepared raw EEG data. The classification process was thus dominated by the peak location. Subsequent analysis showed that online spike detection with an erroneously trained network yielded an area less than 0.5 under the receiver-operating-characteristic curve, and hence performed inferiorly to random assignments. Networks trained and tested using the same unprepared EEG data achieved no better than about 87% true classification rate at equal sensitivity and specificity. CONCLUSIONS: The high true classification rate reported previously is believed to be an artifact arising from erroneous data preparation and off-line validation. Spike detection using raw EEG data as input is unlikely to be feasible under current computer technology. PMID- 10699411 TI - Motor evoked potentials of subjects over 70 years of age with and without recurrent falls. AB - Motor evoked potentials (MEPs) of 83 elderly (79+/-4 years) subjects, 43 with recurrent falls and 40 without, and of 31 healthy young (42+/-9 years) subjects were measured from thenar (and hypothenar) and tibialis anterior muscles. Forty four of the aged subjects without overt neurological diseases were used as controls. Absolute latencies from the cortex to the target muscles as well as the latency differences from the cortex to the level of the fifth lumbar vertebra (LV) were longer in the aged than in the young, but the latency difference from the cortex to the brachial plexus was shorter. The cortical, brachial plexus and lumbar (LV) latencies were all dependent on height as well as age. The latency differences from the cortex to the plexus or LV were not height-dependent but were age-dependent. The thenar MAXMEP/CMAP ratio was significantly higher in hands with thenar atrophy (in 30% of the aged subjects) than without; thenar atrophy thus excludes the use of this parameter in about one-third of the aged subjects. There were no significant differences in the MEP latencies or amplitudes of the recurrent fallers and the non-fallers. Subjects having more frequent falls, however, tended to have lower amplitudes of MEPs in the lower extremities. PMID- 10699412 TI - Antidromic corticospinal tract potential of the brain. AB - OBJECTIVE: To describe a novel potential component (antidromic corticospinal tract potential, ACSP) of the brain after translaminar spinal stimulation of a relaxed patient during scoliosis surgery. To study the origin of this component and to compare its source to known sources of the somatosensory evoked potentials (SEPs). METHODS: We studied 17 consecutive patients during posterior scoliosis surgery. SEPs and ACSPs were elicited by translaminar spinal stimulation at the Th 2 and L 1 levels. ACSPs and SEPs were recorded on the scalp midline. Neurogenic motor evoked potentials (NMEPs) were recorded on the popliteal spaces. Preoperative tibial SEPs were also recorded. RESULTS: ACSP was distinctly separated from the corresponding spinally evoked cortical SEP that showed longer latency than the ACSP. ACSPs decreased and disappeared when stimulation was moved to the caudal direction in the conus region while SEP persisted. In addition, the hemispheric origin of ACSP was confirmed with multichannel midline recordings of the scalp and neck. Thus there was no confusion to the response of nucleus gracilis, corresponding the P 31 response of the tibial nerve SEP. CONCLUSIONS: The origin of ACSP seemed to be in the rostral part of the corticospinal tract. ACSP diminished in the conus region when stimulation was moved caudally and it disappeared when the stimulus was given to the root level. This proves that ACSP is not a response of the somatosensory tract, instead ACSP represents antidromic response of the pyramidal tract. ACSP can be used in monitoring of the motor tracts during scoliosis surgery together with NMEPs. PMID- 10699413 TI - Rhythmic cortical and muscle discharges induced by fatigue in corticobasal degeneration. AB - We describe a patient presenting clinical features of corticobasal degeneration (CBD), including reflex myoclonus in the left upper limb. This patient complained of a marked worsening of involuntary movements in the left upper limb after exercise. We analysed the electrophysiological characteristics of myoclonus in the basal state and after a fatiguing exercise in the left upper limb. In the basal condition, single trials recording EEG showed a cortical complex occurring 20 ms after stimulation of the left median nerve. Surface EMG recordings of the left first dorsal interosseous (FDI) revealed an isolated biphasic C1 response 49 ms after stimulation. After exercise, single trials recording EEG following shocks to the left median nerve showed rhythmic complexes with a duration of approximately 80 ms. EEG complexes were made of a series of 3 bursts, with intervals between bursts tending to cluster at approximately 22 ms. These rhythmic complexes were associated with repetitive activity in the left FDI. We conclude that rhythmic cortical and muscle discharges can be induced by fatigue in CBD. PMID- 10699414 TI - Magnetoelectric brain stimulation in the assessment of brain physiology and pathophysiology. AB - OBJECTIVE: To review findings from transcranial magnetic stimulation (TMS) induced motor evoked potentials in normal subjects, in various neurological diseases and with pharmacologic manipulation. METHODS: MEDLINE was searched to identify pertinent articles and articles referenced therein were also reviewed. RESULTS: TMS is a safe and non-invasive technique which has been used widely in the study of corticospinal and corticocortical connectivity as well as in the assessment of basal ganglia disorders, diffuse diseases, and neuropharmacology. CONCLUSIONS: TMS motor measures have utility in examination of brain structure and function within and beyond the corticospinal tract. These measures have both research and clinical applications. PMID- 10699415 TI - Peripheral motor and sensory nerve conduction studies in normal infants and children. AB - OBJECTIVE: There are few data on electrophysiological data of motor and sensory fibres during nerve maturation. The aim of this study is to investigate the evolution of nerve conduction in the upper and lower limbs during the first years of life. METHODS: The study comprised 92 normal infants and children aged from 1 week to 6 years. Using surface electrodes, the investigation included the following data: (1) motor conduction velocity (MCV), corrected distal motor latency (DML) to a standard distance, and F-waves of the median, ulnar, peroneal and tibial nerves; (2) sensory conduction velocity (SCV) of the median and tibial nerves; and (3) amplitude and morphology of the muscle and sensory action potentials. RESULTS: Maximal MCV and SCV in the neonatal period was about half of adults; there was a steep conduction increase during the first year of life, adult values being reached around age 4. In the neonatal period corrected DML was greater than in adults with a further decrease during the first year. F-wave latencies also decreased during the first year with increase at the end of the study. CONCLUSIONS: This study corroborates the fact that 'maturation' of MCV and SCV occurs during the first 5 years of life, especially in the former. Evolution of DML is accounted for using correction. F-wave latency changes are explained both by an increase in MCV, and extremity growth. PMID- 10699416 TI - Face-elicited ERPs and affective attitude: brain electric microstate and tomography analyses. AB - OBJECTIVES: Although behavioral studies have demonstrated that normative affective traits modulate the processing of facial and emotionally charged stimuli, direct electrophysiological evidence for this modulation is still lacking. METHODS: Event-related potential (ERP) data associated with personal, traitlike approach- or withdrawal-related attitude (assessed post-recording and 14 months later) were investigated in 18 subjects during task-free (i.e. unrequested, spontaneous) emotional evaluation of faces. Temporal and spatial aspects of 27 channel ERP were analyzed with microstate analysis and low resolution electromagnetic tomography (LORETA), a new method to compute 3 dimensional cortical current density implemented in the Talairach brain atlas. RESULTS: Microstate analysis showed group differences 132-196 and 196-272 ms poststimulus, with right-shifted electric gravity centers for subjects with negative affective attitude. During these (over subjects reliably identifiable) personality-modulated, face-elicited microstates, LORETA revealed activation of bilateral occipito-temporal regions, reportedly associated with facial configuration extraction processes. Negative compared to positive affective attitude showed higher activity right temporal; positive compared to negative attitude showed higher activity left temporo-parieto-occipital. CONCLUSIONS: These temporal and spatial aspects suggest that the subject groups differed in brain activity at early, automatic, stimulus-related face processing steps when structural face encoding (configuration extraction) occurs. In sum, the brain functional microstates associated with affect-related personality features modulate brain mechanisms during face processing already at early information processing stages. PMID- 10699417 TI - High-density mapping in an N400 paradigm: evidence for bilateral temporal lobe generators. AB - OBJECTIVE: The aim of this study was to obtain a more detailed description of N400 scalp topography than has previously been reported. METHODS: High-density (128 channel) visual event-related potentials were measured in an N400 paradigm using semantically incongruous sentence endings. RESULTS: The stimuli elicited an N400 with a centroparietal scalp distribution. In addition, P400s with similar timing and functional characteristics were observed at non-standard recording locations inferior to the temporal lobes. CONCLUSIONS: The data are consistent with intracranial evidence for bilateral activation of anterior medial temporal lobe structures. These structures are oriented such that the positive regions of their scalp fields lie largely outside of the area sampled by standard electrode montages. P400s at other non-standard scalp locations, including infraorbital and infraoccipital sites, may reflect volume conduction from the same generators, or activation of non-temporal lobe generators. PMID- 10699418 TI - The individual replicability of mismatch negativity at short and long inter stimulus intervals. AB - OBJECTIVES: The individual replicability of the mismatch negativity (MMN) event related brain potential (ERP) was studied at two different inter-stimulus intervals (ISIs), to establish its potential value for routine clinical evaluation of sound discrimination and auditory sensory memory. METHODS: Ten healthy young subjects were presented sequences of 3 stimulus trains, in two recording sessions approximately 1 month apart. The stimuli in the trains were delivered at an ISI of 300 ms, whereas the inter-train intervals (ITIs) were 0.4 s and 4.0 s in different blocks. ERPs were averaged to standard (75 ms) and deviant (25 ms) tones started equiprobably the stimulus trains. RESULTS: Significant Pearson product-moment correlations coefficients were found between sessions at all scalp locations for the short ITI, when the MMN was quantified as the mean amplitude in the 100-200 ms latency window around its peak. However, none of the correlations reached significance for the longer ITI. CONCLUSIONS: MMN appears to be a reliable measure for single-case assessment and follow-ups when obtained at short ISIs and quantified as an integrated window of neuroelectric activation over a temporal span. PMID- 10699419 TI - Common basal extrastriate areas for the semantic processing of words and pictures. AB - OBJECTIVE: The recognition potential (RP) is an electrophysiological brain response which is sensitive to the semantic processing of meaningful stimuli. In this study we attempt to elucidate the topography and neural origin of the RP evoked by pictures and to compare it with the RP evoked by words. METHODS: Words, pictures, Chinese characters and control stimuli were presented to 20 subjects following the rapid stream stimulation procedure. The activity was recorded using 60 cephalic electrodes. RESULTS: We found a RP displaying its maximal amplitude at the left inferior parieto-occipital electrode (PO7) for words and at the right homologue electrode (PO8) for pictures and Chinese characters. Both the amplitude and the latency of the RP were larger in the case of words. A profile analysis indicated that the neural generators of the RP were common regardless of the type of stimulus, and a dipole analysis placed them about the lingual gyrus. CONCLUSIONS: Words and pictures share the same neural generators for the RP despite of subtle differences in lateralization. This is interpreted as an index of a multimodal semantic processing in basal extrastriate areas. PMID- 10699420 TI - Thyrotropin-receptor antibodies in thyroid diseases: advances in detection techniques and clinical applications. PMID- 10699421 TI - Markers of inflammation as predictors in cardiovascular disease. AB - Recent research suggests that inflammation in the coronary arteries may be intimately involved in the development of atherosclerosis and its associated acute coronary syndromes. Experimental and clinical studies based on the biochemical markers of inflammation and vascular perturbation in plasma as well as in atherosclerotic tissue have provided substantial evidence of ongoing inflammation in coronary heart disease. These studies have suggested a potential role for biochemical markers of inflammation in the prediction of risk for the development of coronary heart disease as well as in the prediction of adverse cardiac-related outcomes in patients with known coronary syndromes. Markers of inflammation appear to offer risk prediction information that is independent of, and possibly complementary to, traditional risk factors for the development of cardiovascular disease. There are numerous potential markers for these inflammatory processes and their interdependence remains somewhat unclear. Clinical studies have investigated some, but not all of these markers in a critical and comparative fashion. Therefore, further well-designed prospective studies should elucidate the role of markers of inflammation in the prediction, diagnosis, and management of cardiovascular disease. PMID- 10699422 TI - Free radicals antioxidant enzymes and lipid peroxidation in different types of leukemias. AB - Free radicals are highly reactive species that have been implicated in the pathogenesis of many diseases. Reactive oxygen species can initiate lipid peroxidation and DNA damage leading to mutagenesis, carcinogenesis and cell death, if the antioxidant system is impaired. This study was undertaken to examine the prevalence of oxidative stress and the role of antioxidant defence in untreated leukemia patients. The generation of superoxide anion and hydrogen peroxide by leukocytes, plasma malondialdehyde levels, red cell copper zinc superoxide dismutase (Cu-Zn SOD) and glutathione peroxidase (GSH-PX) activities were determined in 30 patients with different types of leukemias prior to therapy. The superoxide anion generation by polymorphonuclear leukocytes was found to be significantly increased in leukemia patients especially those with acute lymphocytic and nonlymphocytic leukemias, while the hydrogen peroxide levels were comparable to the control values. Plasma lipid peroxidation products in untreated leukemia patients were in the normal range. Red cell Cu-Zn SOD and GSH-PX activities were significantly increased and showed no correlation with the hemoglobin content. Although superoxide generation was high, lipid peroxide levels were normal in these patients. This might be due to the increased activities of the antioxidant enzymes (SOD, GSH-PX) which counteract lipid peroxidation. Increased free radical generation, especially superoxide anion in leukemia patients and increased antioxidant defence enzymes, which is an adaptive protective response, are indicative of mild oxidative stress. There were no significant differences for the parameters cited above between different types of leukemias, suggesting that the changes are not specific to the type of leukemia. PMID- 10699423 TI - Marked decrease in plasma apolipoprotein A-I and high density lipoprotein cholesterol in a case with Werner syndrome. AB - The patient was a 39-year-old Japanese male with a body height of 160 cm and weight of 48 kg who was diagnosed as Werner syndrome of homozygote for mutation 4. His plasma total cholesterol (TC), triglycerides (TGs), high density lipoprotein-cholesterol (HDL-C) and apolipoprotein A-I (apo A-I) levels were 7.2, 2.1, 1 mmol/l and 128 mg/dl, respectively. During the clinical course of treatment of this patient, his plasma levels of HDL-C and apo A-I declined drastically to levels of as low as 0.2 mmol/l and 10 mg/dl, respectively, with concurrent reciprocal increase in plasma TG levels. Plasma HDL-C, apo A-I and TG levels gradually returned to original values. Lipoprotein lipase activity and mass in post-heparin plasma were markedly low when the apo A-I and HDL-C levels decreased to 10 mg/dl and 0.21 mmol/l, respectively, and these values improved when the apo A-I and HDL-C levels returned to more normal values of 106 mg/dl and 0.94 mmol/l, respectively. The result of direct sequence of the exon 3 and 4, and the promoter region of the apo A-I gene of the patient revealed no single nucleotide changes. These results suggest that in the present patient, impaired hydrolysis of TGs in TG-rich lipoproteins, is due at least in part to a decreased LPL enzyme level, reduced the formation of nascent HDL, resulting in unusually low plasma levels of HDL-C and apo A-I. PMID- 10699424 TI - Pvu II intron 15 polymorphism at the LDL receptor gene is associated with differences in serum lipid concentrations in subjects with low and high risk for coronary artery disease from Brazil. AB - Coronary artery disease (CAD) has a high prevalence in the Brazilian population. Nevertheless, studies of genetic risk factors for CAD in this country have not been sufficiently conducted. We used the Pvu II polymorphism (intron 15) at the low-density lipoprotein receptor (LDLR) gene to study the effect of variation at this locus in determining plasma lipid concentrations in 128 white subjects presenting a lipid profile suggesting high risk for CAD (HRG) and 100 white normolipidemic individuals (controls, CG). The Pvu II polymorphism was detected by PCR-RFLP. The P1P1 genotype for Pvu II polymorphism (homozygous for absence of restriction site) was greater in HRG individuals than in CG subjects (57% vs. 38%, P<0.05). Moreover, the P1P1 genotype was strongly associated with high concentrations of total cholesterol (P=0.0001), triglycerides (P=0. 0295), LDL-C (P=0.0001), and VLDL-C concentrations (P=0.0280) and lower HDL-C concentrations (P=0.0051) in HRG subjects. Similarly, the CG individuals with P1P1 genotype presented high concentrations of total cholesterol and LDL-C compared to other genotypes (P=0. 0001). This study demonstrates the influence of Pvu II polymorphism of the LDLR on serum lipid concentrations of individuals with low and high risk for CAD from Brazil. PMID- 10699425 TI - Assay of a seric human hexapeptide (HWESAS) using a monoclonal antibody and ELISA. AB - Human serum contains low-molecular-weight growth factors potentiating some in vitro biological effects of IGF-I and IGF-II and recently two peptides were mainly identified: HWESAS and WGHE. In order to determine seric HWESAS concentration, a specific monoclonal antibody against HWESAS was prepared. Its specificity was studied by inhibition tests: this antibody cross-reacts with Y HWESAS, Cys-HWESAS. It does not react with HWESAS when its COOH is blocked, or with HWE, WGHE and tryptophan or with C3f (SSKITHRIHWESASLLR) which is a fragment of human complement containing HWESAS motif. Its affinity was measured by non competitive enzyme immunoassay (3.89+/-2.44.10(8) M(-1)). Then, this antibody was used in enzyme-linked immunosorbent assay (ELISA) and the preliminary assays were performed to detect HWESAS in serum. In contrast to healthy subjects, patients with chronic renal failure exhibited undetectable concentration of hexapeptide while after successful renal transplantation values increased to reach levels found in healthy subjects and varying according to post-operative evolution. These data are a strong hint that the kidney plays an important role in the production of this hexapeptide and underly the clinical interest of HWESAS detection in renal pathology. PMID- 10699426 TI - Cell adhesion molecules - can they be used to predict coronary artery disease in patients with familial hypercholesterolaemia? AB - Adhesion of leukocytes to endothelial cells via cell adhesion molecules (CAMS) is thought to be pivotal in the initiation of atherosclerosis. As patients with familial hypercholesterolaemia (FH) are known to develop severe, premature coronary artery disease (CAD), we investigated the usefulness of soluble forms of CAMS namely vascular cellular adhesion molecule-1 (VCAM), intercellular cell adhesion molecule-1 (ICAM) and E-selectin as predictive markers of the presence and severity of atherosclerosis in this patient group. Twenty heterozygous FH patients without CAD; 24 heterozygous FH patients with CAD; 17 homozygous FH patients without documented CAD; nine homozygous FH patients with overt CAD; and 50 healthy controls were studied. Carotid artery intima media thickness (IMT) was also measured in the homozygous patients. Levels of the adhesion molecules VCAM, ICAM and E-selectin were not significantly elevated in homozygous FH patients and heterozygous FH patients, both with and without CAD, compared to the normal control subjects. In addition the range of results was so wide and the overlap of values with normal controls so great, that the use of an individual level of either VCAM, ICAM or E-selectin was not predictive of either the presence or degree of atherosclerosis in the FH subjects. PMID- 10699427 TI - A new method for the determination of sulphide in gastrointestinal contents and whole blood by microdistillation and ion chromatography. AB - Hydrogen sulphide is produced in the human large intestine by the bacterial reduction of dietary inorganic sulphate and sulphite and by fermentation of sulphur amino acids. Sulphide may damage the colonic epithelium and has been implicated in the pathogenesis of ulcerative colitis. The accurate measurement of sulphide in biological samples, particularly in gut contents is difficult due to the volatile nature of the compound, and the viscosity and turbidity of the samples. Here we describe a method for the determination of sulphide in gut contents and whole blood which overcomes these problems. Initially, samples are treated with zinc acetate to trap sulphide. A microdistillation pretreatment is then used, which releases sulphide from its stable, stored state, coupled to ion chromatography with electrochemical detection. The limit of detection of the method was determined as 2.5 micromol/l, which enabled sulphide levels in gut contents and whole blood samples obtained from humans to be accurately determined. A preliminary investigation in healthy human subjects showed blood sulphide ranged from 10 to 100 micromol/l. Whole blood sulphide did not change significantly when increasing amounts of protein from meat were fed. However, faecal sulphide did show a significant increase from 164 to 754 nmol/g in four subjects fed diets which contained 60 and 420 g meat. PMID- 10699428 TI - Total iron-binding capacity and serum transferrin determination under the influence of several clinical conditions. AB - In this study TIBC and serum-transferrin concentrations were determined by immunochemical turbidimetry, immunochemical nephelometry and radial immunodiffusion under normal and pathological clinical conditions. A total of 246 (123 male/123 female) patients were included [iron deficiency: 60 (18/42), iron overload: 56 (39/17), chronic inflammation: 47 (23/24), undefined diseases: 35 (16/19), healthy volunteers 48 (27/21)]. The data show that determination of TIBC from conversion of transferrin values using a constant factor results in significantly higher values compared to conversion with a function of first degree. For clinical practice the influence of different diseases is negligible. This study indicates that it is not possible to develop a universal algorithm for the conversion of transferrin values into TIBC. PMID- 10699430 TI - A microtiter plate assay for superoxide dismutase using a water-soluble tetrazolium salt (WST-1). AB - A simple and reproducible microtiter plate assay for measuring superoxide dismutase (SOD) activity is described. Water-soluble tetrazolium, the sodium salt of 4-[3-(4iodophenyl)-2-(4-nitrophenyl)-2H-5-tetrazolio]-1,3-be nzene disulfonate, was used as a detector of superoxide radical generated by xanthine oxidase and hypoxanthine, in the presence of a range of concentrations of superoxide dismutase. A major advantage of the assay is that one reaction mixture is prepared and aliquotted into wells, avoiding pipetting errors and variable xanthine oxidase activity between samples. Inclusion of standardized SOD solution in each run enables inter-assay comparability without requiring a constant superoxide generation rate under all occasions. The assay is applicable for chloroform-ethanol red cell extracts as well as tissue homogenates without high speed centrifugation. Fifty percent inhibition of formazan formation was achieved at 2.4+/-0.1 ng of Cu, ZnSOD per well with the coefficient of variation 4.2%. PMID- 10699429 TI - The acute coronary syndrome diagnosis and prognostic evaluation by troponin I is influenced by the test system affinity to different troponin complexes. AB - It was suggested recently that cardiac troponins are released as T-I-C complexes and then further degraded to T and I-C. It is not known whether the various affinity to the T-I-C and I-C complex of different troponin I test systems influence the diagnostic and prognostic value of the test results in clinical practice. We studied 162 patients (61.3 S.D. 11.1 years) with suspected acute myocardial infarction (AMI) in a single center study. AMI was confirmed in 109 patients. Blood samples were taken at admission, after 1, 2, 4, 8, 12 and 24 h. Troponin I (TnI) was measured using the OPUS plus (TnI-O, cut-off 1.6 microg/l) and the Stratus II (TnI-S, cut-off 1.5 microg/l) analyzers. TnI-O has high affinity to the binary (I-C) and TnI-S to the ternary (T-I-C) troponin complex. A 6-month follow-up with respect to death and recurrent AMI was performed. The sensitivity (SE) and specificity (SP) for AMI diagnosis were 82.6 and 86.8% for TnI-S; 75.2 and 92.5% for TnI-O 0-2 h after admission. The ROC analysis showed a slightly better curve for TnI-S at 4 h (P<0.05). Logistic regression analysis shows prediction of 6 months outcome by 0-24 h serial TnI-S measurements (odds ratio 5.21, P=0.0356), and serial TnI-O measurements (odds ratio 4.92, P=0.0186). High affinity to the ternary troponin complex enhances the diagnostic but not the prognostic value of a test system. Indeed, the resulting differences are small but underline the need for standardization of biochemical markers. PMID- 10699431 TI - Non-cardiac nucleic acid composition and protein synthesis rates in hypertension: studies on the spontaneously hypertensive rat (SHR) model. AB - Various studies have shown the involvement of extracardiac tissues in hypertension, including the hepato-intestinal tract, musculo-skeletal system, skin, and the kidney. It was our hypothesis that these perturbations in non cardiac tissues would also include alterations in protein metabolism. Thus, the reported differences in soleus contractile protein composition may be related to changes in muscle protein synthesis or reduced protein synthetic efficiencies. The aim of the present study was to characterise tissue composition of nucleic acids and rates of protein synthesis in non-cardiac tissues, such as liver, skeletal muscle (i.e., the Type I fibre-predominant soleus and Type II fibre predominant plantaris), kidney, bone (tibia), skin and the gastrointestinal tract in a genetic model of hypertension (i.e., spontaneously hypertensive rats (SHRs), 15 weeks old) compared to their genetic aged-matched counterparts, i.e., normotensive Wistar-Kyoto (WKY) controls. Rates of protein synthesis were measured in vivo after injection with a flooding dose of L-[4-(3)H]phenylalanine. The results showed changed tissue wet weights (g per organ) for plantaris (+10%, P<0.05), liver (+25%, P<0.01), brain (-9%, P<0.01), jejunum (+39%, P<0.001) and tibia (+17%, P<0.001) in SHRs compared to WKY controls. Protein content (g or mg per organ) was increased in the liver (+32%, P<0. 01) and tibia (+37%, P<0.05). RNA contents (mg per organ) were increased in plantaris (+17%, P<0.01), liver (+22%, P<0.01) and jejunum (+11%, P<0.05). DNA (mg per organ) was increased in plantaris (+16%, P<0.025) and jejunum (+12%, P<0.025). The protein synthetic capacities (i.e., C(s), mg RNA/g protein) were higher in soleus (+41%, P<0.01) and plantaris (+6%, P<0.05) muscles of SHRs compared to WKYs, whereas values were lower in liver (-11%, P<0.01) and kidney (-6%, P<0.01) of SHRs compared to WKYs. The fractional rate of protein synthesis (i.e., k(s), the percentage of the protein pool renewed each day) was not significantly different for any of the tissues, though the rate of protein synthesis per unit RNA (i.e., k(RNA), mg protein/day per mg RNA) was reduced in the soleus (-24%, P<0.05) and the synthesis rate per unit DNA, i.e., k(DNA) (mg protein/day per mg DNA) was increased in the tibia (+31%, P<0.025). This is the first report of significant differences between indices of protein metabolism in extracardiac tissues in hypertension, which may reflect endocrine factors and/or the systemic influence of hypertension per se. PMID- 10699432 TI - Rapid analysis of cation constituents of urine by capillary electrophoresis. AB - Capillary electrophoresis was applied for the simultaneous determination of K(+), NH(4)(+), Na(+), Ca(2+) and Mg(2+) cations in urine. The separations were carried out in electrolyte containing 7. 5 mmol/l copper(II) acetate, 15 mmol/l ethylenediamine and 2 mmol/l triethanolamine at pH 8.0, with indirect UV detection at 230 nm. All cations were well separated in less than 4 min. The results obtained by this method were comparable with ion chromatography. The proposed system is simple, rapid and does not require any preliminary treatment of the urine samples except diluting. PMID- 10699433 TI - Determination of trace elements and calcium in bone of the human iliac crest by atomic absorption spectrometry. AB - A rapid and reliable analytical method for the determination of trace elements in human bone by atomic absorption spectrometry is reported. Calcium was determined to estimate the homogeneity of samples. Human bone from the iliac crest was obtained at autopsy of adult subjects. Before analysis samples were decomposed by microwave digestion and acid digestion in a Parr bomb. Zinc, rubidium, strontium, calcium and iron were determined by flame atomic absorption spectrometry (FAAS) and aluminium, copper and lead by electrothermal atomic absorption spectrometry (ETAAS) at optimum measurement conditions. The results for the two digestion procedures agreed for zinc, rubidium and calcium within +/-5%, for copper within +/-7% and for strontium, iron, aluminium and lead within +/-10%. The repeatability of measurement (R.S.D.) for determination of calcium and trace elements after microwave digestion and acid digestion in a Parr bomb was tested in one representative autopsy bone sample by six parallel determinations. It was found to be better than +/-5% either for microwave digested samples or samples digested in a Parr bomb, for all elements determined by FAAS and ETAAS techniques. The accuracy of the applied digestion procedures was checked by analysis of trace elements in NIST SRM 1486 Bone Meal reference material. Good agreement of the results with certified values was obtained for both digestion procedures. The microwave procedure developed for digestion of small amounts of sample was applied in trace elements analysis of bone biopsy samples from dialysis patients. PMID- 10699434 TI - Limitations on the quantitative determination of telomerase activity by the electrophoretic and ELISA based TRAP assays. AB - Telomerase is a promising new tumor marker and can be detected using the TRAP (Telomeric Repeat Amplification Protocol) method. To address factors affecting its quantitative determination, we evaluated two commercial TRAP assays, an electrophoretic and an ELISA assay formats, using cultured cells and human tumor samples. We found that both TRAP assays had a limited linearity from 250 to 5000 tumor cells, with a similar intra-assay variation. The quantification of TRAP products was affected by high cell number in sample, the presence of non-tumor cells, and interfering substances in patient specimens. Because both assays have different limitations, determination of telomerase by a combined use of the two may provide more accurate information on the telomerase activity in a specimen. Extracts of specimens should also be tested at several concentrations to insure that the result is not being falsely decreased by an inhibitor. The quantitative results for telomerase activity by the TRAP assays, however, should be interpreted cautiously. PMID- 10699435 TI - Quantitative assessment of apolipoprotein E genotypes by image analysis of PCR RFLP fragments. AB - Apolipoprotein E (APOE) genotyping usually involves polymerase chain reaction (PCR) and assessment of restriction fragment length polymorphism (RFLP) by gel electrophoresis. We made determination of HhaI restriction endonuclease digestive patterns more objective and improved diagnostic accuracy with a quantitative approach using sensitive DNA stain (SYBR Green) and image analysis of gel patterns. For distinguishing true and partially-digested restriction fragments, band ratios were calculated for the staining intensity of gel patterns from 116 sample runs of 63 human blood specimens. Each of these specimens was independently genotyped for APOE by at least two (and most of them by three) different PCR-RFLP methods. Based on the distribution of band ratios, decision levels were established and used for developing a program for computer-aided interpretation of APOE genotypes (Microsoft Excel software). Appropriateness of the decision levels for band ratios was validated by APOE genotyping of additional 61 specimens. The approach described here is applicable to a variety of other molecular diagnostic techniques that are based on PCR-RFLP or sequence specific signal amplifications. PMID- 10699436 TI - Differential modulation by native and glycated low density lipoproteins of peripheral blood mononuclear cells proliferation induced by phytohemagglutinin in insulin-dependent diabetes mellitus patients. PMID- 10699437 TI - Intracerebral transplantation of bone marrow with BDNF after MCAo in rat. AB - We tested the hypothesis that a composite graft of fresh bone marrow (BM) along with brain-derived neurotrophic factor (BDNF), transplanted into the ischemic boundary zone (IBZ) of rat brain, facilitates BM cells to survive and differentiate, and improves functional recovery after middle cerebral artery occlusion (MCAo). The fresh BM was harvested from adult rats injected with bromodeoxyuridine (BrdU) as a tracer. Rats (n=37) were subjected to 2h of MCAo, received grafts at 24h and were sacrificed at 7days after MCAo. Test groups consisted of: (1) control - MCAo alone (n=9); (2) injection of phosphate buffered saline (n=4); (3) transplantation of BM (n=8); (4) injection of BDNF (n=7); and (5) transplantation of BM with BDNF (n=9) into the IBZ. Immunohistochemistry was used to identify cells derived from the BM stem cells. Behavioral tests (rotarod motor test; adhesive-removal somatosensory test) were performed before and 7days after MCAo. The data indicate that intracerebral grafting of a combination of BM with BDNF enhances differentiation of BM cells and significantly improves motor recovery of rotarod (P<0.05) and adhesive-removal (P<0.05) tests. We anticipate that BM along with neurotrophic factors may provide a powerful autoplastic therapy for human neurological injury and degenerative disorders. PMID- 10699438 TI - Shc-binding site in the TrkB receptor is not required for hippocampal long-term potentiation. AB - Recent evidence shows that neurotrophins are not only involved in neuronal survival and differentiation but also in modulating synaptic strength in the developing and adult nervous system. To understand how neurotrophins induce changes in synaptic strength, we have investigated signaling pathways downstream of the TrkB receptor, which binds brain-derived neurotrophic factor (BDNF) or NT 4/5. To test whether the Shc-site activated signaling pathway, which has been shown to be important for neuronal survival in vivo, also plays a role in processes like long-term potentiation (LTP), we have generated a mouse strain carrying a mutation in the Shc-binding site of the TrkB receptor. In hippocampal slices from these mice we investigated whether basal synaptic transmission, early LTP (E-LTP) or late-LTP (L-LTP) were affected by this mutation. We found that homo- and heterozygous mutant mice show no difference in the induction-rate or magnitude of E-LTP and L-LTP induced by theta-burst or tetanus stimulation, suggesting that the Shc-binding site in the TrkB receptor and its downstream activated signaling cascade is not involved in hippocampal synaptic plasticity. PMID- 10699439 TI - Glutamate transport blockade has a differential effect on AMPA and NMDA receptor mediated synaptic transmission in the developing barrel cortex. AB - High affinity glutamate transport plays an important role in maintaining a low extracellular glutamate concentration in the CNS. Excitotoxicity due to a loss of glutamate transporter function has been implicated in disease processes such as stroke and amyotrophic lateral sclerosis (ALS). We studied the effects of glutamate transport inhibitors on thalamocortical synapses at developing (postnatal day 3-8) layer IV neurons in the barrel cortex using the thalamocortical slice preparation and whole-cell recordings. Inhibition of glutamate transport by D,L-threo-beta-hydroxyaspartate (THA), a combination of THA and dihydrokainate (DHK), or by L-trans-pyrrolidine-2,4-dicarboxylate (tPDC), caused a reversible blockade of AMPA and kainate receptor-mediated dual component excitatory postsynaptic currents (AMPA/KA EPSCs). This effect was not blocked by cyclothiazide (CTZ) indicating that is was not due to desensitisation of AMPARs. Under conditions in which NMDA receptors were unblocked the transport inhibitors caused the massive activation of NMDA receptors leading to the rapid loss of recordings. Previous studies using these transport inhibitors on brain slices from older animals reported no or only modest effects on synaptic transmission. Therefore the data in the present study suggest that neurons in the developing neocortex are particularly sensitive to glutamate transporter function. Furthermore the effects of transport inhibition are dependent upon whether neurons are sufficiently depolarised to relieve the voltage-dependent block of NMDA receptors. PMID- 10699440 TI - Role of sensory deficits in motor impairments after injury to primary motor cortex. AB - After a focal ischemic lesion in the hand representation of the primary motor cortex in squirrel monkeys, manual skill was mildly and transiently impaired on a reach-and-retrieval task. Performance was significantly poorer during weeks 1 and 3 post-lesion, but was normal by week 4. An unusual behavioral event was also observed after the lesion. Monkeys reached for pellets, but visually inspected the hand for the presence of the pellets, even when none were actually retrieved. This behavior, possibly indicative of a sensory deficit, was rarely observed prior to the lesion, but was observed at significantly higher levels during week one post-lesion. These results suggest that the primary motor cortex plays a significant role in somatosensory processing during the execution of motor tasks. Motor deficits heretofore identified as purely motor, may be at least partially due to a sensory deficit, or sensory-motor disconnection. PMID- 10699442 TI - Recovery from early cortical damage in rats, VIII. Earlier may be worse: behavioural dysfunction and abnormal cerebral morphogenesis following perinatal frontal cortical lesions in the rat. AB - The size of cortical removal was varied in rats that were given medial frontal lesions on postnatal day 2. In adulthood, the animals were trained on the Morris water task and Whishaw reaching task following which the brains were harvested and dendritic arborization and spine density was examined in the layer III pyramidal cells in Zilles' area Par1. There was a small relationship between lesion size and behavioral outcome as smaller lesions produced somewhat smaller deficits. In contrast, both small and large lesions produced large reductions in brain weight, dendritic arborization, and spine density. The cortex of newborn rats appears to be especially vulnerable to even restricted injury. This contrasts to the effects of similar injury a week later when animals show extensive functional recovery and anatomical compensation. PMID- 10699441 TI - Reactive astrocytic responses to denervation in the motor cortex of adult rats are sensitive to manipulations of behavioral experience. AB - Recent research has suggested that mild denervation of the neocortex of adult rats may facilitate neuronal growth in response to behavioral changes. Astrocytes react to denervation, produce growth-promoting factors and are a potential mediator of this denervation-facilitated growth. The present study assessed whether astrocytic reactions to denervation vary dependent upon post-injury behavioral experience. Denervation of the transcallosal afferents to the motor cortex was induced via partial transections of the corpus callosum. Transected- or sham-operated rats were then either forced to use the opposite forelimb (via limb-restricting vests) or permitted to use both forelimbs normally for 8 days. In the motor cortex, the surface density of glial fibrillary acidic protein (GFAP)-immunoreactive (IR) astrocytic processes and the density of basic fibroblast growth factor (FGF-2)-IR glial cells was significantly increased as a result of transections alone and as a result of forced forelimb-use alone in comparison to controls. The combination of transections and forced-use significantly enhanced GFAP-IR in comparison to all other groups, but did not further enhance FGF-2-IR. These findings are consistent with behavior and denervation having interactive influences on astrocytic reactivity in the motor cortex. These results also raise the possibility that astrocyte-mediated support of neural restructuring after brain injury might be enhanced with appropriate post-injury behavioral manipulations. PMID- 10699443 TI - Physiological consequences of morphologically detectable synaptic plasticity: potential uses for examining recovery following damage. AB - A growing literature indicates that brain structure is modified in various ways with experience. In this paper we briefly survey evidence that the brain retains the capacity to modify its organization in response to demands, including demands resulting from learning, throughout the lifetime. We attempt to address whether these experience-induced changes are accompanied by physiological changes that indicate a functional reorganization of the brain. The kinds of morphological changes that have been observed following brain injury appear to be very similar to those seen after learning. The similarity suggests that many of the basic mechanisms of synaptic change in the brain may be utilized for both functions. This suggests that we can take advantage of some of the methods used to test the changes in physiology with behavioral manipulations to examine the damaged brain. We advocate utilizing electrophysiological techniques to measure functional recovery from brain injury as these may be useful in evaluating both spontaneous recovery from damage and the therapeutic benefits of training, or other therapies. PMID- 10699444 TI - CNS plasticity and assessment of forelimb sensorimotor outcome in unilateral rat models of stroke, cortical ablation, parkinsonism and spinal cord injury. AB - We have reviewed a battery of useful tests for evaluating sensorimotor function and plasticity acutely and chronically in unilateral rat models of central nervous system injury. These tests include forelimb use for weight shifting during vertical exploration in a cylindrical enclosure, an adhesive removal test of sensory function, and forelimb placing. These tests monitor recovery of sensorimotor function independent of the extent of test experience. Data are presented for four models, including permanent focal ischemia, focal injury to the forelimb area of sensorimotor cortex, dopaminergic neurodegeneration of the nigrostriatal system, and cervical spinal cord injury. The effect of the dendrite growth promoting factor, Osteogenic Protein-1 (OP-1) on outcome following permanent middle cerebral artery (MCA) occlusion was used as an example to illustrate how the tests can be applied preclinically. OP-1 showed a beneficial effect on limb use asymmetry in the cylinder test. PMID- 10699445 TI - Loss of the innate cortical engram for action patterns used in skilled reaching and the development of behavioral compensation following motor cortex lesions in the rat. AB - Damage to the motor cortex of the rat (Rattus norvegicus) impairs skilled movements used in reaching for food with the contralateral forepaw. Nevertheless, there is substantial recovery in success over a two-week postsurgical period. The profile of behavioral recovery is believed to reflect the eventual normalization of behavior, but this idea has not been explicitly examined. The present experiments examined postsurgical reaching success and reaching movements as a function of (1) lesion type, (2) lesion size, (3) lesion location, (4) depletion of forebrain noradrenaline, and (4) presurgical and postsurgical experience. The results show that at least two separate processes contribute to recovery in postsurgical performance. The early postsurgical period was characterized by extreme difficulties in making reaching movements. The experiments suggest that this initial impairment was due to the loss of the innate cortical engram that supports the action patterns used for skilled movements. Subsequent recovery in reaching success was not due to the reacquisition of normal movements, but was due rather to the use of compensatory movements. The results are discussed in relation to the idea that true recovery from motor cortex injury will require that damaged neurons and their connections be rescued or replaced. PMID- 10699446 TI - Functional assessments in mice and rats after focal stroke. AB - This paper presents a comprehensive assessment of sensorimotor deficits in the mouse after focal ischaemia induced by occlusion of the middle cerebral artery. Twenty four hours after induction of middle cerebral artery occlusion, mice showed deficits in a range of sensory and motor tasks as assessed by the SHIRPA protocol. In addition they exhibited a decrease in rotarod performance and locomotor activity. Some behaviours, such as locomotor activity, were also impaired in sham operated animals compared to normal controls, although these impairments were not as marked as those exhibited by the ischaemic mice. This is the first comprehensive analysis of the short term effects of permanent focal ischaemia in mice. In a second series of experiments in the rat, rates of recovery over time were examined. Simple (neurological grades, rotarod) and complex (sticky label test) tasks were examined in rats after middle cerebral artery occlusion up to 7 days post-ischaemia. Ischaemic rats had a profound deficit in contralateral performance on the sticky label task with no evidence of recovery. A less marked deficit was also observed in ipsilateral performance of this task. These deficits were still present 7 days after ischaemia. Ischaemic rats also exhibited a deficit on rotarod performance but this had recovered 7 days post-ischaemia. Thus different sensorimotor tasks have different rates of recovery after focal cerebral ischaemia in the rat. Further characterisation of these tasks will enhance their utility meaningful preclinical means of assessing functional recovery of the administration of potential neuroprotective and regenerative therapies. PMID- 10699447 TI - Effects of subdural haematoma on sensorimotor functioning and spatial learning in rats. AB - Twenty per cent of all strokes are haemorrhagic in character and are associated with severe disturbances in sensorimotor behaviour and cognition. Although spontaneous recovery of pre-stroke functioning occurs in some cases, the process is demanding, slow, and often incomplete. A first step in the preclinical testing of new putative, neuroprotective and recovery-supporting therapeutics is to validate animal models of brain injury. In a series of four experiments we evaluated the behavioural impairments and the time course of recovery of functional deficits in rats with an experimentally induced subdural haematoma. We found that unilateral subdural haematoma resulted in dysfunction in both simple reflexive (experiment 1) and skilled sensorimotor behaviour (experiment 2). Reflexive behaviour did not recover, or recovered only marginally, and neither did the deficits in skilled forepaw use. Bilateral subdural haematoma impaired the learning and memory performance of adult (experiment 3) and old rats (experiment 4) in the Morris water escape task. Considering the diversity of the deficits found in our experiments, we conclude that different models are needed to cover the broad range of deficits seen in stroke patients. PMID- 10699448 TI - Defining post-stroke recovery: implications for design and interpretation of drug trials. AB - Measurement of stroke recovery is complex because definition of successful recovery is highly variable across measures and cut-off points for defining successful outcomes vary. The purpose of this paper is to describe patterns of recovery in stroke patients of varying severity when different measures are used and when different cut-off points are selected. 459 individuals enrolled in a prospective cohort study were assessed within 14 days post stroke and re evaluated at 1, 3, and 6 months. Recovery was assessed using the NIH Stroke Scale, the Fugl-Meyer Assessment of Motor Recovery, the Barthel Index of Activities of Daily Living, the Physical Function Index of the SF-36, and the Modified Rankin Outcome Scale. Subjects also defined their preference (utility) for their current health state with a time-trade off question. We compared patterns of recovery using the different measures and varying the cut-off points for defining successful recovery. The percentage of patients who are believed to have recovered depends on how recovery is defined. If recovery is defined at the disability level (Barthel > 90), the majority 57.3% of stroke survivors experience a full recovery. Fewer individuals are considered to be fully recovered if impairments are measured (NIH 90, 36.8%. Less than 25% of stroke survivors are considered recovered if recovery is defined relative to reported prior function in higher levels of physical activity. Shifting the definition of recovery on the modified Rankin scale from /=20 Gy). The time from the initial RT to re-treatment ranged from 8 to 136 months (median: 23 months). All patients were treated with external RT and conformal radiotherapy was used in 35 patients after 1993. Fifteen received radiosurgery as a boost treatment. The RT dose at the nasopharyngeal tumor area ranged from 20 to 67.2 Gy (median 50 Gy). Eighty-two patients received one to eight courses of cisplatin-based chemotherapy in addition to RT. RESULTS: The 1-, 3- and 5-year survival was 54.9, 22. 1 and 12.4%, respectively. Patients whose tumor relapsed later than 2 years after the first treatment had a better survival than those with earlier relapse (3-year survival: 30.1 vs. 10.8%; P=0.015), but the difference became insignificant in patients who received >/=50 Gy. Patients without evidence of intracranial invasion or cranial nerve palsy had better survival than those with such lesions (3-year survival: 30.9 vs. 3.7%; P=0.006). A re-treatment dose >/=50 Gy yielded better survival (3-year survival: 22.8 vs. 18.5%; P=0.003). Addition use of radiosurgery may improve survival. The use of chemotherapy did not improve survival. Conformal radiotherapy resulted in significantly fewer severe complications than conventional RT. CONCLUSIONS: A repeat course of RT for locally recurrent NPC successfully prolongs survival in a significant number of patients. Intracranial invasion and/or cranial nerve palsy and re-treatment dose affect the prognosis, with a dose of >/=50 Gy significantly improving survival. Radiosurgery boost may also improve survival. Our preliminary data indicates that conformal radiotherapy may decrease the severity of radiation-induced complications. However; longer follow-up and larger sample size is necessary to document the findings. PMID- 10699477 TI - Hypofractionation in glioblastoma multiforme. AB - PURPOSE: To compare conventional fractionation with hypofractionation in patients with a glioblastoma multiforme. Endpoints of the analysis are overall survival and palliative effect. MATERIALS AND METHODS: From 1988 to 1998, 155 patients with pathologically confirmed glioblastoma multiforme were prospectively analysed. Patients without irradiation and patients receiving an interstitial boost were excluded from this analysis. Three different radiation schemes were used in subsequent periods; 33x2, 8x5 and 4x7 Gy. In the last 5 years a scheme of 4x7 Gy conformal irradiation was given to poor prognosis patients. The more favourable group received the conventionally fractionated scheme up to 66 Gy. RESULTS: Median survival was 7, 5.6 and 6.6 months for the 33x2, 8x5 and 4x7 Gy, respectively. In general, patients in the hypofractionation group had far worse prognostic factors compared with patients treated with the conventional scheme. The period of neurological improvement or stabilisation was similar between the 4x7 and 33x2 Gy group. CONCLUSION: An extreme hypofractionation scheme of 4x7 Gy conformal irradiation in poor prognostic glioblastoma patients is well tolerated, convenient for the patient and provides equal palliation without negative effects on survival compared with conventional fractionation. PMID- 10699478 TI - Computer simulation of cytotoxic and vascular effects of radiosurgery in solid and necrotic brain metastases. AB - PURPOSE: Solid and necrotic brain tumors respond to radiosurgery, although necrotic lesions often contain a significant proportion of hypoxic cells which cannot become reoxygenated during the short overall treatment time of single dose application. In addition to the direct cytotoxic action, delayed vascular occlusion followed by ischemic tumor cell death could contribute to the effect of radiosurgery. MATERIALS AND METHODS: In order to determine the impact of the two possible effects on tumor response, a 3-dimensional computer simulation was developed and fitted to response data obtained from 90 patients who were treated by LINAC radiosurgery for 1-3 brain metastases with median marginal doses of 20 Gy. Complete response rates were as follows: small, solid lesions (diameter 0.4-1 cm), 52% (12/23); large solid lesions (1.1-5.2 cm), 28% (17/60); large necrotic lesions, 12% (6/50). The 3-dimensional computer model simulated the growth of small solid and large, solid or necrotic tumors situated in a vascularized stroma. Oxygen supply, tumor cell division (cell cycle time 5 days), neovascularization, tumor cell kill by single dose irradiation (linear-quadratic model, alpha/beta=10 Gy, oxygen enhancement ratio 3.0) and time-dependent vascular occlusion (alpha/beta=3 Gy) were modeled by Monte-Carlo simulation techniques. RESULTS: In the presence of neovascularization, solid tumors with a hypoxic fraction of 1-2% developed. Without neoangiogenesis, central necrosis occurred, and tumors had a hypoxic fraction of 20-25%. Assuming a pure cytotoxic effect of radiosurgery, neither the dose-response relationship for the solid lesions of different size nor that for the large lesions with solid or necrotic appearance could be reproduced for any given level of radiosensitivity. This was only possible by introducing a vascular effect that led to the occlusion of >/=99% of the vessels at the border of the target volume within 1 year after irradiation. In the presence of the vascular effect, the apparent radiosensitivity of the tumor cells was increased by 50-100%. Calculations of the dose-equivalent for the vascular effect show that it contributes 19-33% of the overall effect of single dose radiosurgery. CONCLUSION: This simulation study suggests that the therapeutic effect of single radiosurgery in malignant brain tumors cannot be understood without the consideration of vascular effects. The computer model might serve as a basis for exploring new treatment modalities that modify both cytotoxic and vascular effects of radiosurgery. PMID- 10699479 TI - Immediate toxicity during fractionated total body irradiation as conditioning for bone marrow transplantation. AB - BACKGROUND: Total body irradiation followed by bone marrow transplantation is well established as a part of the conditioning regimen in high dose therapy. The immediate tolerance of fractionated total body irradiation (FTBI) was investigated prospectively. METHODS: From January 1995 to December 1998 162 patients received a FTBI, 6x2 Gy on 3 consecutive days, lung dose 10 Gy, for allogeneic (n=112) or autologous (n=50) bone marrow transplantation. High dose chemotherapy (mostly Cyclophosphamide) was administered after the FTBI. A standardized supportive therapy was administered. The immediate toxicity of FTBI was evaluated prospectively prior to each radiation fraction using a defined questionnaire. RESULTS: Main symptoms distressing the patient during irradiation period were gastrointestinal symptoms like nausea and emesis. The prevalence of nausea per fraction increased to 26.1% after the 4th fraction, with a significant higher prevalence in children younger than 10 years at 1st and 2nd fractions. 42.6 and 22. 8%, respectively, of all patients complained of nausea and episodes of emesis, during FTBI. Mild xerostomia and parotiditis were observed in 29.9 and 7.1% of all patients. Further gastrointestinal side effects during FTBI were loss of appetite in 16.0%, indisposition in 25.3%, mild oesophagitis in 3.7% and diarrhoea in 3. 7% of the patients. During FTBI 41.4% of the patients developed a temporary skin irritation (mild erythema). Pruritus was registered in 3.7% of the patients. Headache was observed in 14.8% and Fatigue syndrome in 49.2% of women and 28.3% of men (P<0.005). CONCLUSION: FTBI is a well tolerated therapeutic regimen in high dose therapy. The 162 patients investigated revealed no severe immediate side effects. PMID- 10699481 TI - Mathematical modeling of chronical hypoxia in tumors considering potential doubling time and hypoxic cell lifetime. AB - PURPOSE: To model mathematically how potential doubling time and hypoxic cell lifetime affect the extent of chronical hypoxia in tumor tissue segments. Three capillary geometries were modeled under idealized steady state conditions. MATERIALS AND METHODS: The capillary geometries are: tissue surrounding an axial capillary, tissue enclosed by a cylindrical capillary network, and tissue enclosed by a spherical capillary network. The tissue segments are modeled as three-compartment systems, where well nourished cells proliferate near the vasculature and, in so doing, displace 'older' cells into a quiescent compartment and, ultimately into a hypoxic region. The extent of the hypoxic zone is the distance traversed by cells during their hypoxic lifetime before becoming necrotic. The steady state situation, where the necrotic cell loss equals the cell gain caused by cell proliferation was investigated. RESULTS: The hypoxic fraction, HF, was found to be inversely proportional to the potential doubling time of the tumor segment, T(pot), and proportional to the hypoxic cell lifetime, T(hypox). The extent of the oxygenated zone depends only on the capillary geometry, the capillary radius, the intracapillary oxygen tension, and the tissue respiration rate. The extent of the hypoxic zone in addition depends on T(pot) and T(hypox). CONCLUSIONS: Mathematical modeling of idealized steady state conditions shows that the ratio of hypoxic cell lifetime and potential doubling time, T(hypox)/T(pot), determines the hypoxic fraction, HF, in tumor segments. The extents of the oxygenated and the hypoxic zones can be predicted from the models. PMID- 10699480 TI - Definition and validation of a reference target volume in early stage node positive cervical carcinoma, based on lymphangiograms and CT-scans. AB - PURPOSE: To establish a reference planning target volume for postoperative radiotherapy in stage Ib and IIa N+ cervical carcinoma, based on 47 lymphangiograms and 15 CT-scans. METHODS: Radiation oncologists (n=17) from all radiotherapy institutes in The Netherlands were asked to define the clinical target volume (CTV) and planning target volume (PTV), and to delineate (on simulation films) the radiotherapy treatment portals following a radical hysterectomy with lymph node dissection for an early stage cervical carcinoma with positive iliac lymph nodes. A reference PTV was defined by using 47 normal lymphangiograms and CT-data of the pelvis from 15 patients who underwent surgery for cervical carcinoma. The simulation films were digitized and evaluated for adequacy in covering the PTV, previously individually determined by the radiation oncologists. Subsequently, the simulation films were also evaluated for adequacy in covering the reference PTV. RESULTS: Large variations were observed in the portals used and in treatment techniques. From the digitized films, it appeared that in 50% of the cases the defined PTV was not covered adequately. Furthermore, 71% of the treatment plans would not cover the lateral borders of the reference PTV sufficiently. CONCLUSIONS: There appears to be no consensus on the target volumes to be irradiated in postoperative radiotherapy of early stage cervical carcinoma. When a PTV defined on the basis of lymphangiograms and CT-data is taken as a reference, 71% of the treatment plans would not cover this PTV adequately. These findings indicate the need for a consensus in the design of standardized treatment volumes. PMID- 10699482 TI - Contractile properties of human renal cell carcinoma recruited arteries and their response to nicotinamide. AB - BACKGROUND AND PURPOSE: The manipulation of tumour blood supply and thus oxygenation is a potentially important strategy for improving the treatment of solid tumours by radiation. Increased knowledge about the characteristics that distinguish the tumour vasculature from its normal counterparts may enable tumour blood flow to be more selectively modified. Nicotinamide (NA) causes relaxation of preconstricted normal and tumour-supply arteries in rats. It has also been shown to affect microregional blood flow in human tumours. Direct effects of NA on human tumour supply arteries have not previously been reported. This paper describes our evaluation of the effects of NA on two parameters: 'spontaneous', oscillatory contractile activity and agonist (phenylephrine)-induced constriction in the arteries supplying human renal cell carcinomas. MATERIALS AND METHODS: Isolated renal cell carcinoma feeder vessels were perfused in an organ bath with the alpha(1)-adrenoceptor agonist phenylephrine (PE). When the arteries had reached a plateau of constriction, nicotinamide (8.2 mM) was added to the perfusate and changes in perfusion pressure were measured. RESULTS: PE (10 microM) induced a sustained constriction in the majority of the renal cell carcinoma feeder vessels examined, demonstrating that they retain contractile characteristics, at least in response to this alpha(1)-adrenoceptor agonist. In combination with NA (8.2 mM) the constriction was significantly attenuated in half of the preparations. In addition, seven arteries exhibited spontaneous contractile activity which was significantly attenuated by NA in six of them. CONCLUSIONS: NA can significantly attenuate both 'spontaneous' and agonist induced constrictions in tumour-recruited human arteries, though not all arteries are sensitive. PMID- 10699483 TI - Active steroidogenesis in the normal rat skin. AB - Using the radiolabeled precursors of adrenal steroids (14)C-11 deoxycorticosterone (DOC) and (14)C-progesterone ((14)C-PROG) we demonstrate that rat skin can synthesize a number of steroids. TLC separation of labeled metabolites show that among the (14)C-steroid products, two co-migrate with corticosterone (B) and 11-dehydrocorticosterone (A) standards. Thus, normal rodent skin possesses steroidogenic activity that can be shown using progesterone or DOC as primary substrates. PMID- 10699484 TI - Differential effects of gemcitabine on ribonucleotide pools of twenty-one solid tumour and leukaemia cell lines. AB - To gain a more detailed insight into the metabolism of 2', 2'-difluoro-2' deoxycytidine (dFdC, gemcitabine, Gemzar) and its effect on normal ribonucleotide (NTP) metabolism in relation to sensitivity, we studied the accumulation of dFdCTP and the changes in NTP pools after dFdC exposure in a panel of 21 solid tumour and leukaemia cell lines. Both sensitivity to dFdC and accumulation of dFdCTP were clearly cell line-dependent: in this panel of cell lines, the head and neck cancer (HNSCC) cell line 22B appeared to be the most sensitive, whereas the small cell lung cancer (SCLC) cell lines were the least sensitive to dFdC. The human leukaemia cell line CCRF-CEM accumulated the highest concentration of dFdCTP, whereas the non-SCLC cell lines accumulated the least. Not only the amount of dFdCTP accumulation was clearly related to the sensitivity for dFdC (R= 0.61), but also the intrinsic CTP/UTP ratio (R=0.97). NTP pools were affected considerably by dFdC treatment: in seven cell lines dFdC resulted in a 1.7-fold depletion of CTP pools, in two cell lines CTP pools were unaffected, but in 12 cell lines CTP pools increased about 2-fold. Furthermore, a 1.6-1.9-fold rise in ATP, UTP and GTP pools was shown in 20, 19 and 20 out of 21 cell lines, respectively. Only the UTP levels after treatment with dFdC were clearly related to the amount of dFdCTP accumulating in the cell (R=0.64 (P<0.01)), but not to the sensitivity to dFdC treatment. In conclusion, we demonstrate that besides the accumulation of dFdCTP, the CTP/UTP ratio was clearly related to the sensitivity to dFdC. Furthermore, the UTP levels and the CTP/UTP ratio after treatment were related to dFdCTP accumulation. Therefore, both the CTP and UTP pools appear to play an important role in the sensitivity to dFdC. PMID- 10699485 TI - N-terminally modified glucagon-like peptide-1(7-36) amide exhibits resistance to enzymatic degradation while maintaining its antihyperglycaemic activity in vivo. AB - Glucagon-like peptide-1(7-36)amide (tGLP-1) is inactivated by dipeptidyl peptidase (DPP) IV by removal of the NH(2)-terminal dipeptide His(7)-Ala(8). We examined the degradation of NH(2)-terminally modified His(7)99% of His(7) glucitol tGLP-1 remained intact at 12 h. His(7)-glucitol tGLP-1 was similarly resistant to plasma degradation in vitro. His(7)-glucitol tGLP-1 showed greater resistance to degradation in vivo (92% intact) compared to tGLP-1 (27% intact) 10 min after i.p. administration to Wistar rats. Glucose homeostasis was examined following i.p. injection of both peptides (12 nmol/kg) together with glucose (18 mmol/kg). Plasma glucose concentrations were significantly reduced and insulin concentrations elevated following peptides administration compared with glucose alone. The area under the curve (AUC) for glucose for controls (AUC 691+/-35 mM/min) was significantly lower after administration of tGLP-1 and His(7) glucitol tGLP-1 (36 and 49% less; AUC 440+/-40 and 353+/-31 mM/min, respectively; P<0.01). This was associated with a significantly higher AUC for insulin (98-99% greater; AUC 834+/-46 and 838+/-39 ng/ml/min, respectively; P<0.01) after tGLP-1 and His(7)-glucitol tGLP-1 administration compared to controls (421+/-30 ng/ml/min). In conclusion, His(7)-glucitol tGLP-1 resists plasma DPP IV degradation while retaining potent antihyperglycaemic and insulin-releasing activities in vivo. PMID- 10699486 TI - 1H-NMR and raman studies on perforating trauma-induced cataract formation in a mouse lens. AB - In order to provide new insight into the molecular mechanism of perforating trauma-induced cataract formation in an 8-week-old ddY mouse lens, we performed an in situ investigation into changes in the water-protein and/or protein-protein interactions by using 500 MHz (1)H-NMR spectroscopy, and into structural alterations in lens proteins by using Raman spectroscopy. Cross-relaxation times of water protons in the perforated opaque lens were considerably shorter than those in the intact transparent lens, whereas there was no significant difference in water content, suggesting a drastic change in water-protein and protein protein interactions in the perforated lens. In addition, there was no significant difference in the intensity ratios of several key Raman bands between intact and perforated lenses, indicating that no significant local and overall conformational changes in lens protein itself occur in the perforated lens. The present (1)H-NMR and Raman results lead us to the conclusion that changes leading to lens opacification in the perforating trauma-induced cataract appear to involve the rapid formation of immobile large lens protein aggregates without formation of intra- and intermolecular disulfide linkages, and rapid increase in a fraction of bound water associated with large protein aggregates. PMID- 10699487 TI - Expression of glypican-1, syndecan-1 and syndecan-4 mRNAs protein kinase C regulated in rat immature Sertoli cells by semi-quantitative RT-PCR analysis. AB - In seminiferous tubules, Sertoli cells provide structural and nutritional support for the developing germinal cells. Cell to cell signalization and cell adhesion require proteoglycans expressed at the cell membrane. A preliminary biochemical and structural approach indicated that cell surface proteoglycans are mostly heparan sulfate (HSPG) in immature rat Sertoli cells. The present study focused on the qualitative and quantitative expression of three membrane HSPG, syndecan 1, syndecan-4 and glypican-1 in Sertoli cells of 20-day-old rat. A semi quantitative multiplex RT-PCR strategy was developed to appreciate the effect of PKC activation on the mRNA expression of the three HSPG. Our data show that the syndecan-1 and glypican-1 mRNA expression is increased by the phorbol myristate acetate (PMA) suggesting a regulation of their expression by the phosphatidyl inositol pathway, as previously hypothesized (Fagen et al., Biochim. Biophys. Acta, 1472 (1999) 250-261). In addition, a physiological effector of the PKC as ATP gave similar effects. Thus, this over-expression could be related with paracrine factors secreted by germ cells. PMID- 10699488 TI - Serine and glycine transport in fetal ovine hepatocytes. AB - The role of hepatic serine and glycine transport in the regulation of the biosynthesis of serine by the fetal liver has not been studied. The goal of this study was to characterize serine and glycine transport and utilization at physiologic concentrations in primary cultures of fetal ovine hepatocytes. Primary culture of hepatocytes from mid gestation ( approximately 90 days) and term ( approximately 135 days) fetal sheep were studied after overnight serum free culture. At both gestational ages, the initial rate for sodium dependent 300 microM serine transport (1697+/-131 pmoles/min/mg protein at mid, 1765+/-544 at term) was fourfold greater than sodium dependent 300 microM glycine transport (309+/-54 at mid, 579+/-252 at term). At physiologic concentrations (300 microM), 69+/-7% of serine and 49+/-8% of glycine transport was sodium dependent. At term, sodium dependent serine transport has a V(max) of 1751+/-348 pmoles/min/mg protein and a K(m) of 159+/-111 microM. Sodium independent serine transport has a V(max) of 904+/-185 and a K(m) of 416+/-188 microM. Sodium dependent glycine transport has a V(max) of 410+/-69 and a K(m) of 2290+/-895 microM while sodium independent glycine transport exhibits non-saturable kinetics. Glycine (300 microM) sodium dependent transport was not inhibited by methyl-AIB while sodium dependent 300 microM serine transport was inhibited (31%). n-Ethylmaleimide inhibited sodium dependent serine and glycine transport by 36+/-9% and 37+/-2% respectively in term hepatocytes. Cysteine inhibited sodium dependent serine transport by 37%. Sodium independent glycine transport at 300 microM was higher in low glucose (1.1 mM) medium (881+/-76 pmoles/min/mg protein) compared to high glucose (5.5 mM) medium (510+/-60 P=0.004). There were no significant differences in serine or glycine incorporation into RNA, DNA, glycogen or lipid and protein. The predominance of serine transport over glycine at physiologic concentrations suggests that inward cellular amino acid transport of serine and glycine is not likely to be a regulatory mechanism that would favor serine biosynthesis in fetal ovine hepatocytes. PMID- 10699489 TI - Effects of beta-carotene on antioxidant enzyme activity, intracellular reactive oxygen and membrane integrity within post confluent Caco-2 intestinal cells. AB - As encountered with a plethora of other natural products, the antioxidant activity of beta-carotene has been proposed as one of the mechanisms by which diets rich in this pro-vitamin A active carotenoid apparently afford chemoprevention. Here, we report the ability of beta-carotene to alter endogenous reactive oxygen levels and antioxidant defences within non-stressed 'differentiated' monolayers of an intestinal epithelial cell line (Caco-2) and to subsequently effect resistance to general oxidative insult. The differentiated monolayers efficiently absorbed beta-carotene. Between 3 and 8 days post confluence, cultures exhibited a progressive increase in antioxidant enzyme activity and a corresponding reduction to intracellular ROS levels. The profile for antioxidant enzyme activity was unaffected by sustained daily supplementation with beta-carotene. However, after two daily treatments with 50 microM beta carotene intracellular ROS levels were significantly reduced and there was a trend towards reduced intracellular ROS within monolayers subject to five daily treatments with 0.5 and 5 microM beta-carotene. Prolonged supplementation with 0.1 and 0.5 microM beta-carotene or short supplementation periods with 5 and 50 microM beta-carotene did not alter susceptibility to H(2)O(2). However, cultures treated daily between 3 and 8 days post confluence with 5 or 50 microM beta carotene exhibited enhanced LDH release, increased non-adherence and enhanced Trypan blue staining when challenged with 10 mM H(2)O(2). In the absence of H(2)O(2), the beta-carotene treatments were not overtly toxic to the monolayers. These results indicate that beta-carotene does not enhance antioxidant defences within Caco-2 monolayers. The enhancement of H(2)O(2) toxicity by persistent, high doses of beta-carotene may contribute to the failure of this carotenoid to protect high risk individuals from certain degenerative conditions. PMID- 10699490 TI - Biochemical characterization of two crotamine isoforms isolated by a single step RP-HPLC from Crotalus durissus terrificus (South American rattlesnake) venom and their action on insulin secretion by pancreatic islets. AB - Crotamine, a neurotoxin present in the venom of the South American rattlesnake Crotalus durrisus terrificus exists as several polymorphic variants, as demonstrated by recombinant DNA technology (Smith and Schmidt, Toxicon 28 (1990) 575-585). We have isolated native crotamine by chromatography on Sephadex G75, and have purified two crotamine isoforms (F2 and F3) by a single step of RP-HPLC. Native crotamine and RP-HPLC fractions F2 and F3 produced skeletal muscle spasms and spastic paralysis in mice. At low glucose concentrations (2.8-5.6 mmol/l), none of the crotamines altered the insulin secretion by rat isolated islets. In the presence of 16.7 mmol glucose/l, F2 (5 microg/ml), but not F3, increased insulin secretion two-fold, whereas native crotamine (1.5, 5 and 16.5 microg/ml) potentiated the secretion dose-dependently. The increase in insulin secretion induced by F2 fraction (5 microg/ml) was similar to that obtained with 16.5 microg of native crotamine/ml. These results indicate that the mode of action of the F2 and F3 isoforms in beta-cells is different from that in muscle cells. This difference may be related to the binding affinity of each isoform for the Na(+) channels located in the beta-cell membrane. Crotamine isoforms may be valuable tools for studying the involvement of Na(+) channels in the mechanism of insulin secretion. PMID- 10699491 TI - Molecular cloning, tissue distribution and sequence analysis of complete glucokinase cDNAs from gilthead seabream (Sparus aurata), rainbow trout (Oncorhynchus mykiss) and common carp (Cyprinus carpio). AB - The enzyme glucokinase (GK) (EC 2.7.1.1) plays an important role in the control of glucose homeostasis. Qualitative and/or quantitative variations in GK enzyme have been postulated by previous studies to explain why dietary carbohydrate utilisation is lower in gilthead seabream (Sparus aurata) and rainbow trout (Oncorhynchus mykiss) than in common carp (Cyprinus carpio). In this study, we report the isolation and characterisation of a full-length cDNA coding for GK in these teleosts. Amino acid sequences derived from these cDNA clones are highly similar to other vertebrate GKs. These findings, including a detailed phylogenetic analysis, reveal that GK gene highly homologous to mammalian GK exists in these fish species with similar tissue specific expression (mainly liver). PMID- 10699492 TI - In vitro recycling of alpha-D-ribose 1-phosphate for the salvage of purine bases. AB - In this paper, we extend our previous observation on the mobilization of the ribose moiety from a purine nucleoside to a pyrimidine base, with subsequent pyrimidine nucleotides formation (Cappiello et al., Biochim. Biophys. Acta 1425 (1998) 273-281). The data show that, at least in vitro, also the reverse process is possible. In rat brain extracts, the activated ribose, stemming from uridine as ribose 1-phosphate, can be used to salvage adenine and hypoxanthine to their respective nucleotides. Since the salvage of purine bases is a 5-phosphoribosyl 1 pyrophosphate-dependent process, catalyzed by adenine phosphoribosyltransferase and hypoxanthine guanine phosphoribosyltransferase, our results imply that Rib-1P must be transformed into 5-phosphoribosyl 1-pyrophosphate, via the successive action of phosphopentomutase and 5-phosphoribosyl 1-pyrophosphate synthetase; and,in fact, no adenosine could be found as an intermediate when rat brain extracts were incubated with adenine, Rib-1P and ATP, showing that adenine salvage does not imply adenine ribosylation, followed by adenosine phosphorylation. Taken together with our previous results on the Rib-1P-dependent salvage of pyrimidine nucleotides, our results give a clear picture of the in vitro Rib-1P recycling, for both purine and pyrimidine salvage. PMID- 10699493 TI - Deficiencies of glycolytic enzymes as a possible cause of hemolytic anemia. AB - The critical minimum values of Na,K-ATPase and glycolytic enzyme activities at which the erythrocyte viability is lost were calculated using the mathematical model of the erythrocyte, which included all reactions of glycolysis, adenylate metabolism, ionic balance, and osmotic regulation of erythrocyte volume. The criterion for cell death was an increase in its volume to the level at which it is sequestrated from the circulation or is lysed. In hemolytic anemia associated with hexokinase or pyruvate kinase deficiency, activities of these enzymes measured in patient erythrocytes appeared to be close to the calculated critical values. By contrast, in hemolytic anemia associated with phosphofructokinase, glucosephosphate isomerase, triosephosphate isomerase, or phosphoglycerate kinase deficiency, activities of these enzymes measured in patient erythrocytes were significantly greater than the calculated critical values. In this case, if the deficient enzyme were stable, i.e. its activity in the cell were low, but constant in time, the deficiency observed would not account for the erythrocyte destruction observed and the development of hemolytic anemia. It was shown, however, that in phosphofructokinase, glucosephosphate isomerase, triosephosphate isomerase, or phosphoglycerate kinase deficiency, hemolytic anemia can arise because of the instability of these enzymes in time. PMID- 10699494 TI - Tissue-specific expression of c-series gangliosides in the extraneural system. AB - c-Series gangliosides in extraneural tissues from young and adult rats were examined using thin-layer chromatographic (TLC) immunostaining with a specific monoclonal antibody A2B5. The composition of c-series gangliosides significantly differed among tissues. In adult rats, while liver tissue contained GT1c, GQ1c, and GP1c, renal tissue had GT3 as the major c-series ganglioside with GT2 in a lesser amount. Pancreatic tissue expressed c-series gangliosides that consisted of GT3, GT2, GQ1c, and GP1c. In other tissues including adrenal, thyroid, and eye lens, GT3 constituted the main c-series ganglioside species. While total ganglioside contents of extraneural tissues were much lower than that of brain tissue, the proportions of c-series gangliosides to total gangliosides were higher in many extraneural tissues. Interestingly, eye lens had the highest GT3 content among rat tissues examined. The compositions and concentrations of c series gangliosides in liver and kidney significantly differed between 5-day-old and 7-week-old rats, suggesting the development-dependent expression of c-series gangliosides in these tissues. These results suggest that the expression of c series gangliosides in extraneural tissues is regulated in a tissue-specific manner. PMID- 10699495 TI - Effects of 6-formylpterin, a xanthine oxidase inhibitor and a superoxide scavenger, on production of nitric oxide in RAW 264.7 macrophages. AB - As well as superoxide generated from neutrophils, nitric oxide (NO) produced by inducible nitric oxide synthase (iNOS) in macrophages plays an important role in inflammation. We previously showed that 6-formylpterin, a xanthine oxidase inhibitor, has a superoxide scavenging activity. In the present study, to elucidate other pharmacological activities of 6-formylpterin, we investigated the effects of 6-formylpterin on production of nitric oxide (NO) in the murine macrophage cell line RAW 264.7 stimulated by lipopolysaccharide (LPS) and interferon-gamma (INF-gamma). 6-Formylpterin suppressed the expression of iNOS, and it also inhibited the catalytic activity of iNOS, which collectively resulted in the inhibition of NO production in the stimulated macrophages. However, 6 formylpterin did not scavenge the released NO from an NO donor, S-nitroso-N acetylpenicillamine (SNAP). These results indicate that 6-formylpterin inhibits pathological NO generation from macrophages during inflammation, but that it does not disturb the physiological action of NO released from other sources. PMID- 10699496 TI - Purification and characterization of cAMP dependent protein kinase from Microsporum gypseum. AB - A cyclic AMP dependent protein kinase (PKA), its regulatory (R) and catalytic (C) subunits were purified to homogeneity from soluble extract of Microsporum gypseum. Purified enzyme showed a final specific activity of 277.9 nmol phosphate transferred min(-1) mg protein(-1) with kemptide as substrate. The enzyme preparation showed two bands with molecular masses of 76 kDa and 45 kDa on sodium dodecyl polyacrylamide gel electrophoresis. The 76 kDa subunit was found to be the regulatory (R) subunit of PKA holoenzyme as determined by its immunoreactivity and the isoelectric point of this subunit was 3.98. The 45 kDa subunit was found to be the catalytic (C) subunit by its immunoreactivity and phosphotransferase activity. Gel filtration using Sepharose CL-6B revealed the molecular mass of PKA holoenzyme to be 240 kDa, compatible with its tetrameric structure, consisting of two regulatory subunits (76 kDa) and two catalytic subunits (45 kDa). The specificity of enzyme towards protein acceptors in decreasing order of phosphorylation was found to be kemptide, casein, syntide and histone IIs. Purified enzyme had apparent K(m) values of 71 microM and 25 microM for ATP and kemptide, respectively. Phosphorylation was strongly inhibited by mammalian PKA inhibitor (PKI) but not by inhibitors of other protein kinases. The PKA showed maximum activity at pH 7.0 and enzyme activity was inhibited in the presence of N-ethylmaleimide (NEM) which shows the involvement of sulfhydryl groups for the activity of PKA. PKA phosphorylated a number of endogenous proteins suggesting the multifunctional role of cAMP dependent protein kinase in M. gypseum. Further work is under progress to identify the natural substrates of this enzyme through which it may regulate the enzymes involved in phospholipid metabolism. PMID- 10699497 TI - Helicobacter pylori-antigen-binding fragments expressed on the filamentous M13 phage prevent bacterial growth. AB - Colonization of the human stomach by Helicobacter pylori is associated with the development of gastritis, duodenal ulcer, mucosa-associated lymphoid tissue (MALT) lymphoma, and gastric cancer. H. pylori-antigen-binding single-chain variable fragments (ScFv) were derived from murine hybridomas producing monoclonal antibodies and expressed as a g3p-fusion protein on a filamentous M13 phage. The recombinant ScFv-phage reacted specifically with a 30-kDa monomeric protein of a H. pylori surface antigen preparation and by means of immunofluorescence microscopy the phage was shown to bind to both the spiral and coccoid forms of the bacterium. In vitro, the recombinant phage exhibited a bacteriocidal effect and inhibited specifically the growth of all the six strains of H. pylori tested. When H. pylori was pretreated with the phage 10 min before oral inoculation of mice, the colonization of the mouse stomachs by the bacterium was significantly reduced (P<0.01). The results suggest that genetic engineering may be used to generate therapy-effective phages. PMID- 10699498 TI - Pasteurella multocida: the elusive determinants of virulence and immunity. PMID- 10699499 TI - The molecular biology of pasteurella multocida. AB - Pasteurella multocida is an important veterinary and opportunistic human pathogen. The species is diverse and complex with respect to antigenic variation, host predeliction and pathogenesis. Certain serological types are the aetiologic agents of severe pasteurellosis, such as fowl cholera in domestic and wild birds, bovine haemorrhagic septicaemia and porcine atrophic rhinitis. The recent application of molecular methods such as the polymerase chain reaction, restriction endonuclease analysis, ribotyping, pulsed-field gel electrophoresis, gene cloning, characterisation and recombinant protein expression, mutagenesis, plasmid and bacteriophage analysis and genomic mapping, have greatly increased our understanding of P. multocida and has provided researchers with a number of molecular tools to study pathogenesis and epidemiology at a molecular level. PMID- 10699500 TI - The immunogenicity and pathogenicity of Pasteurella multocida isolated from poultry in Indonesia. AB - Of 13 field isolates of Pasteurella from chickens and ducks in Indonesia, 10 were confirmed as P. multocida subspecies multocida, one as P. multocida subspecies gallicida and one as P. multocida subspecies septica. Nine were capsular Type A four were Serotype 1, one was Serotype 4, one was Serotype 11, one was Serotypes 4,12, and the remaining six were untypable. Five isolates were pathogenic for mice and two were pathogenic for chickens. Both a trivalent vaccine which included local field isolates and an imported commercial vaccine, were efficacious in layer chickens against challenge with virulent reference and local field strains. Though not statistically significant, the protection provided by the trivalent vaccine against virulent field isolate challenge was slightly better and could provide an improvement over the currently used imported vaccine although further field trials are required. A bacterin vaccine produced from a Serotype 1 field isolate grown in the allantoic sac of embryonated chicken eggs provided chickens with good cross protection against heterologous serotype challenge. PMID- 10699501 TI - Characteristics of a haemolytic extract from avian Pasteurella multocida. AB - In experimental fowl cholera, the intramuscular inoculation of Pasteurella multocida induces tissue damage that implies proteolytic or cytolytic activity of the bacteria. Such activity could not be demonstrated by conventional in vitro tests. The treatment of P. multocida strain VP21 with Tween-80 yielded an extract that lysed washed chicken red cells. Extracts were active to a maximum titre of 64. Haemolytic activity of the extract was neither affected by boiling nor by extremes of pH, indicating the active component was not a simple protein. Treatment with trypsin had no effect, but it was inactivated by Proteinase K. Yields were highest from bacteria grown in dextrose starch- or casein sucrose yeast broths; were similar if cultured in air or anaerobically, but were reduced if the bacteria were grown in 5% CO(2). Haemolytic activity was eliminated on exposure to serum or serum albumen. The extract from strain VP21 haemolysed red cells from the chicken, rabbit, sheep, horse, bovine and human, with the highest titres observed on chicken cells. Six other avian strains and seven out of 10 strains of P. multocida from other species yielded an extract which haemolysed chicken red cells. The elaboration of this cytotoxic substance in vivo and its role in pathogenesis remains to be determined. PMID- 10699502 TI - The demonstration of Pasteurella multocida in the alimentary tract of chickens after experimental oral infection. AB - A selective medium containing polymyxin B, crystal violet, thallous acetate, bacitracin and cycloheximide in 10% sheep blood dextrose starch agar, and a modified Pasteurella multocida-specific polymerase chain reaction (PCR) assay were developed for the respective isolation and detection of P. multocida from chicken alimentary tract. The selective medium and the PCR assay were highly sensitive, detecting 100 cfu from colon contents. These techniques were used to follow the localisation of an orally administered virulent P. multocida in chickens. Pasteurellae could be isolated from the crop of some birds up to 30 h, occasionally from other sites after 28 h. It was concluded that the crop was a likely site for colonisation and that infection was most likely to occur through the mucosa of the jejunum or ileum. PMID- 10699503 TI - The virulence and protective efficacy for chickens of pasteurella multocida administered by different routes. AB - The relative virulence for chickens of five strains of Pasteurella multocida was evaluated. Twenty groups, each of ten chickens, were inoculated with a standard dose of 10(5) of each of five strains by the intramuscular (I.m.), intravenous (I.v.), intratracheal (I.tr.) or conjunctival (Co) routes. The highest mortality occurred in the groups dosed I.m. and I.v., followed by I.tr. inoculation. The relative virulence of each strain did not change when inoculated by the different routes. The most virulent strain, VP161, caused 100% mortality by all except the Co route. The least virulent strain, VP17, caused a single mortality by the I.v. route, but gave a high level of protection to birds inoculated by both the I.m. and I.v. routes, when challenged by intramuscular injection with (VP161). There was no protection against I.m. challenge in the birds inoculated by the I.tr. or Co routes. Serum antibody levels measured by ELISA correlated with the level of protection against virulent challenge for groups inoculated I.m. or I.v., but not I.tr. Western blots of pooled sera from each group did not show any specific antigen recognition that might explain the observed differences in protection. Inoculation with strain VP17, (both I.m. and I.tr.) also gave a high level of protection to birds challenged with strain VP161 by intratracheal instillation. PMID- 10699504 TI - PCR detection and analysis of Pasteurella multocida from the tonsils of slaughtered pigs in Vietnam. AB - A total of 36 tonsil swab samples were collected from healthy swine prior to slaughter at the abattoirs in Can tho and Tien giang provinces of Southern Vietnam. The presence of Pasteurella multocida in these samples was detected by the combination of direct cultivation and isolation, mouse inoculation and the polymerase chain reaction (PM-PCR). P. multocida was detected in 16 samples by PCR, with 17 strains ultimately isolated. All samples were negative for serogroup B by HSB-PCR and conventional serotyping, with isolates identified as A:3, D:1 or D:3. In addition, all samples were determined to be negative for the P. multocida toxin (PMT). Characterisation of isolated P. multocida by REP-PCR and biotyping revealed nine distinct REP profiles and seven biotypes among the 17 isolates. Some correlation was seen with P. multocida isolated from a previous Australian outbreak of acute swine pasteurellosis, and those isolated from fowl cholera outbreaks in Vietnamese poultry. PMID- 10699505 TI - The type 4 fimbrial subunit gene of pasteurella multocida. AB - Colonisation of host tissue by Gram- negative bacteria is facilitated by various adhesins, one of which is type 4 fimbriae (pili). These structures have been associated with pathogenesis in several bacterial species, and have been shown to mediate colonisation of epithelial surfaces. Recently, type 4 fimbriae were identified and characterised from P. multocida strains A, B and D. The type 4 fimbrial subunit protein (PtfA) was identified as an 18-kDa protein which was isolated from whole membrane fractions. We report here the isolation and characterisation of the gene (ptfA) encoding the PtfA protein from P. multocida VP161 (serotype A:1). Part of the gene was cloned on a 2-kb genomic DNA fragment. The complete ptfA gene was obtained using inverse PCR. The gene and its flanking regions were characterised, and the deduced PtfA amino acid sequence was compared to type 4 subunit protein sequences from other bacterial species. The ptfA gene was amplified and sequenced from several P. multocida strains. Comparison of these sequences revealed variation within the type 4 subunit gene of P. multocida. PMID- 10699506 TI - Overexpression and immunogenicity of the Oma87 outer membrane protein of Pasteurella multocida. AB - The outer membrane protein of Oma87 from Pasteurella multocida A:1 has significant similarity to the D15 protective antigen of Haemophilus influenzae (Ruffolo and Adler, 1996). Four fragments of Oma87 from a P. multocida serotype D strain were cloned into a pGEX expression vector and transformed into E. coli JM105. Western blot analysis revealed that convalescent chicken sera reacted with only GST-F1 fusion protein which contained amino acids 18 through to 130 of Oma87 fused to the GST protein. Vaccination with the GST-F1 protein failed to protect chickens against challenge with a virulent P. multocida serotype A. PMID- 10699507 TI - Molecular characterisation of avian Pasteurella multocida isolates from Australia and Vietnam by REP-PCR and PFGE. AB - A total of 95 isolates of Pasteurella multocida were analysed by pulsed field gel electrophoresis (PFGE) using the enzyme ApaI, including 73 avian isolates from Australia and 22 from Vietnam. The majority of field isolates were capsular Type A, with the predominant somatic serovars of 1, 3, 4 and 3,4. Twenty-one distinct profiles were evident among the Australian isolates, with only 3 profiles observed among the 22 P. multocida strains isolated from Vietnam. Within the Australian isolates, related and unrelated outbreaks could be identified by PFGE. These results correlated well with previously published studies, with greater discrimination shown by PFGE. Repetitive extragenic palindromic sequence PCR (REP PCR) analysis of representative isolates from PFGE classifications yielded 21 profiles, with most of the subgroups in accordance with PFGE analysis. While REP PCR was shown to be less discriminating than PFGE, the epidemiological relatedness of strains compared favourably between the techniques. Thus, the ease and rapidity of REP-PCR while maintaining a high level of differentiation, supports the use of REP-PCR as a competent alternative to the more labour intensive PFGE system for strain identification and epidemiological studies of avian P. multocida. PMID- 10699508 TI - The molecular epidemiology of four outbreaks of porcine pasteurellosis. AB - Biochemical profiles, restriction endonuclease analysis (REA) and ribotyping were used to investigate a total of 38 Pasteurella multocida isolates from four separate outbreaks of pasteurellosis in Australian piggeries. Six isolates were obtained from Outbreak 1, 16 from Outbreak 2 and eight each from outbreaks 3 and 4. Outbreaks 1 and 2 were cases of pneumonic pasteurellosis while outbreaks 3 and 4 involved systemic pasteurellosis. Biochemical characterisation established that a number of different types of P. multocida were present in outbreaks 1 and 3 while outbreaks 2 and 4 were associated with a single type of P. multocida. Outbreaks 1 and 3 yielded isolates of P. multocida that belonged to the subspecies multocida and gallicida, with the subspecies multocida isolates being identified as biovar 3 (6 in total) or 12 (1 in total) and the subspecies gallicida isolates (7 in total) being identified as biovar 8. All 24 isolates from outbreaks 2 and 4 belonged to the subspecies multocida and were all biovar 3. REA and ribotyping showed that, in outbreaks 1 and 3, there were three different types of P. multocida in each outbreak with no common strains between the outbreaks. The molecular methods showed that only a single strain of P. multocida was associated with outbreaks 2 and 4, although the outbreaks were associated with strains that differed in REA profiles but shared a ribotype profile. This study has shown that both, systemic and pneumonic pasteurellosis can be associated with either a single strain or multiple strains of P. multocida. The results also indicate that the molecular typing methods of REA and ribotyping are superior to biochemical characterisation for epidemiological investigation of porcine pasteurellosis. PMID- 10699509 TI - Genetic organisation of the capsule biosynthetic locus of Pasteurella multocida M1404 (B:2). AB - Capsules from a range of bacterial species have been shown to be major virulence determinants and capsule has been implicated in virulence in Pasteurella multocida. Moreover, capsular serogroup appears to be related to disease predilection. Haemorrhagic septicaemia strains belong to serogroup B and E, fowl cholera strains to serogroup A and atrophic rhinitis strains to serogroup D. The entire capsule biosynthetic locus of P. multocida A:1 has been cloned and its nucleotide sequence determined (Chung et al., 1998. FEMS Microbiol. Lett. 166, 289-296); however, nothing is known of the P. multocida B:2 capsule locus. In this work we have determined the nucleotide sequence and genetic organisation of the P. multocida M1404 (B:2) capsule locus. By analogy with the cap loci of other bacteria, the nucleotide sequence can be divided into three functional regions. Regions 1 and 3 comprise six genes involved in transport of the polysaccharide capsule to the cell surface. The deduced products of these genes show high similarity to proteins involved in capsule export in other bacteria. Region 2 comprises nine genes which are likely involved in biosynthesis of the polysaccharide capsule. The deduced products of three of these genes (bcbA, bcbB and bcbC) show significant similarity to proteins known to be involved in polysaccharide biosynthesis while the other six show no similarity to known proteins. However, their organisation indicates they are co-transcribed with bcbA, bcbB, bcbC and the Region 1 capsule export genes, suggesting strongly that they are also involved in capsule biosynthesis. PMID- 10699510 TI - Cloning and characterisation of the Pasteurella multocida ahpA gene responsible for a haemolytic phenotype in Escherichia coli. AB - Haemolysins are membrane-damaging agents which have been described as bacterial virulence factors due to their ability to lyse erythrocytes and other host cells, and therefore inducing a greater inflammatory response (Elliott et al., 1998). Pasteurella multocida was found to be haemolytic under anaerobic conditions. In this study, we cloned and characterised a P. multocida gene, designated ahpA, which conferred a haemolytic phenotype on Escherichia coli when incubated under anaerobic conditions. A deletion was introduced into the ahpA open reading frame which abolished the haemolytic phenotype. The clone containing ahpA showed erythrocyte specificity, causing haemolysis of bovine and equine erythrocytes, and demonstrated weak haemolysis on ovine erythrocytes. Upon further investigation, AhpA was found to affect the expression of the E. coli K-12 latent haemolysin, SheA, under anaerobic conditions. PMID- 10699511 TI - Survival of avian strains of Pasteurella multocida in chicken serum. AB - The ability of bacteria to survive in serum is considered a likely virulence determinant in diseases where the infective bacteria become septicaemic. Optimal conditions were established to test the survival of Pasteurella multocida in chicken serum. Serum was used at 90%, the inoculum was 10(3)-10(4)cfu in phosphate buffered saline pH 7.4. Survival was measured after incubation for 2-4 h; if survival was <50% the strain was considered serum susceptible. Susceptible strains were either killed or their growth was inhibited. Some resistant strains not only survived but grew rapidly in unheated serum. Thirty-five strains, all originally isolated from clinical fowl cholera, were tested; eight were susceptible, of which three were killed and five inhibited, and the remainder (27) were resistant. Ten serum-resistant P. multocida serogroup A strains were grown in hyaluronidase to remove the capsule and survival in chicken serum was re tested. Three strains became susceptible, while seven strains remained resistant. Three serum susceptible strains were then tested in the presence of cytidine monophosphate-N-acetylneuraminic acid (CMP-NANA). This substance is present in the human serum, and is known to mask the effect of complement on Neisseria gonorrhoeae rendering susceptible strains resistant. Two of the three serum susceptible strains became resistant in the presence of CMP-NANA. Serum susceptibility/resistance was more complex than that of Escherichia coli, and the role of resistance to avian complement in the pathogenesis of fowl cholera remains to be determined. PMID- 10699512 TI - Phagocytic uptake and killing of virulent and avirulent strains of Pasteurella multocida of capsular serotype A by chicken macrophages. AB - The susceptibility of two highly virulent (VP161 and VP138) and two less virulent (VP17 and VP21) strains of Pasteurella multocida to phagocytic uptake and killing by chicken macrophages was compared using in vitro phagocytosis and bactericidal assays. When compared with VP17 and VP21, particularly after they were preopsonised with specific immune serum, VP161 and VP138 were more resistant to phagocytosis by chicken macrophages. The uptake of these bacteria increased following the removal of the bacterial capsules with hyaluronidase. All strains preopsonised with specific immune serum were killed to some extent by chicken macrophages. However, the percentages of killing for VP17 and VP21 were higher than those of VP161 and VP138. When the capsules of VP161 and VP138 were removed, the susceptibility of the bacteria to bactericidal activity of chicken macrophages increased. It can be concluded that the virulent strains of P. multocida were more resistant to phagocytosis and phagocytic killing by chicken macrophages compared with the less virulent strains. The hyaluronic acid capsule was considered to be important in the resistance, but might not be the only factor contributing to the resistance since the less virulent strains of P. multocida also possess capsules. PMID- 10699513 TI - Transdermal drug delivery system exposure outcomes. AB - Transdermal drug delivery systems are increasingly popular, yet few data exist regarding medical outcomes after exposures. Using data collected through a Regional Poison Information System, this retrospective study identified 61 cases of transdermal drug delivery system exposures reported over a recent 5-year period. Exposure routes included dermal (48 patients), oral (10 patients), combined oral and dermal (one patient), parenteral use of gel residue (one patient), and combined oral and parenteral (one patient). Forty-four exposures (72%) were managed by home telephone consultation only. Eleven of 17 patients (18%) evaluated in health care facilities were admitted, including eight (13%) to intensive care units. Hospital admission correlated statistically with clonidine and fentanyl exposures, oral exposures, and drug abuse. Clonidine exposure also correlated statistically with intensive care admission. One fatality was recorded, and all other patients recovered uneventfully. Transdermal drug delivery system exposures are infrequently reported to our regional poison information center but are associated with a significant hospital use and admission rate. PMID- 10699514 TI - Endotracheal lidocaine administration via an esophageal combitube. AB - The purpose of this study was to test the hypothesis that lidocaine is systemically absorbed after administration via a Combitube placed in the esophagus, and that therapeutically significant plasma lidocaine concentrations can be attained using this route with standard endotracheal doses (2.0 mg/kg). During general anesthesia, 27 elective surgical patients received 2.0 mg/kg lidocaine (diluted as necessary with 0.9% saline to a minimum total volume of 10 mL) via a Combitube (study group, n = 13) or an endotracheal tube (control group, n = 14). Venous blood samples were drawn for 3 h after lidocaine administration and plasma concentrations determined by gas chromatography using a nitrogen phosphorus detector (NPD). Overall, average lidocaine concentrations were maximal after 5 min, reaching 0.8+/-0.7 and 1.7+/-0.7 microg/mL in the Combitube and endotracheal tube groups, respectively. Individual patient peak concentrations averaged 1.0+/-0.7 and 2.2+/-1.1 microg/mL in the same two groups, 19+/-16 and 10+/-15 min after lidocaine administration, respectively. No patients reported chest discomfort or dyspnea upon awakening, and no other side effects were noted. In support of the hypothesis, administration of lidocaine via an esophageal Combitube results in systemic drug uptake; however, at conventional endotracheal doses, plasma concentrations are subtherapeutic. It remains to be determined whether higher doses of lidocaine administered via an esophageal Combitube will result in therapeutic plasma concentrations. PMID- 10699515 TI - Cardiac output measurement with an esophageal doppler in critically ill Emergency Department patients. AB - Cardiac output (CO) is a principal determinant of perfusion in many critically ill patients. The objectives of this study were to determine whether physicians' estimates of CO, or cardiac index (CI), are accurate compared with CO/CI measured by esophageal doppler, and to estimate the physician time necessary for Emergency Department (ED) CO/CI measurement. We prospectively evaluated a convenience sample of critically ill, adult ED patients. Based on all available clinical information, residents and emergency medicine attendings estimated patients' CI as being high, normal, or low. A blinded investigator measured CO/CI using an esophageal doppler probe. Times to achieve optimal doppler signal were recorded. Agreement between physician CI estimates and measured CI values was assessed using the weighted kappa statistic. Thirty-three patients were evaluated. There was no agreement beyond chance between physicians' estimates of CI and measured CI. The mean time for optimal doppler signal was 5.7+/-4.3 min. Physicians' estimates of CI were inaccurate compared with measured CI. Esophageal doppler measurement of CO/CI appears to be practical from a physician time standpoint. PMID- 10699516 TI - Base deficit level indicating major injury is increased with ethanol. AB - Analyses were performed to determine whether ethanol increases base deficit, independent of major injury, in blunt trauma patients from two Level I trauma centers. In 2140 Baltimore patients, base deficit was significantly higher in ethanol-positive patients (blood level > or =0.01%), independent of blood pressure (BP), Injury Severity Score (ISS), and blood loss. In 139 Youngstown, Ohio, patients, base deficit was significantly higher in ethanol-positive patients, independent of ISS and RBC units given. In 1796 awake Baltimore patients, major injury was defined as an ISS >10, presence of blood loss, or need for RBC transfusion. A base deficit < or =-4.1 for ethanol-positive and < or = 1.1 for ethanol-negative patients had higher rates of major injury (odds ratio 3.2 and 2.1, respectively) and abdominal trauma (odds ratio 3.6 and 3.2, respectively). In blunt trauma patients, base deficit is increased with ethanol, independent of major injury. A base deficit of < or =-4.1 for ethanol-positive and < or =-1.1 for ethanol-negative awake patients may be an early warning for occult injury and suggest the need for an abdominal computed tomography (CT) scan or ultrasound. PMID- 10699517 TI - "Medical clearance" of psychiatric patients without medical complaints in the Emergency Department. AB - This study was conducted to evaluate the benefit of comprehensive "medical clearance" (history, physical examination, vital signs, laboratory, radiography) in patients presenting to the Emergency Department (ED) with isolated psychiatric complaints. All patients 16 years and older who presented with a psychiatric complaint and required a psychiatric evaluation before discharge from the ED were included in the study. Data, obtained in a 5-month consecutive, retrospective chart review, included patient age, sex, initial complaint, past medical and psychiatric history, initial vital sign measurement, physical examination findings, laboratory analysis (electrolytes, complete blood count, toxicology screen), chest X-ray study results, and final disposition. The number of patients who could have been referred to a psychiatric unit after a history, physical examination, and stable vital signs, without additional laboratory or radiographic studies, was determined. There were 212 patients who met the inclusion criteria, and all their charts were available for review. Eighty patients (38%) presented with isolated psychiatric complaints coupled with a documented past psychiatric history. All received a comprehensive "medical clearance" in the ED followed by a psychiatric consultation. None of the patients had positive screening laboratory or radiographic results. All were either dispositioned home or to the psychiatric ED. The remaining 132 patients (62%) presented to the ED with medically based chief complaints or past medical history requiring further evaluation in the ED before discharge. The initial complaints of these patients correlated directly with the need for laboratory and radiographic "medical clearance" in the ED. Patients with a primary psychiatric complaint coupled with a documented past psychiatric history, negative physical findings, and stable vital signs who deny current medical problems may be referred to psychiatric services without the use of ancillary testing in the ED. PMID- 10699518 TI - Viral brachial neuritis in emergency medicine. AB - Brachial plexus neuritis is a rare neurologic disease that may be overlooked in emergency medicine because other conditions are much more common. We report a case of brachial plexus neuropathy due to cytomegalovirus infection. The diagnosis was based on history, clinical findings, laboratory tests, and electromyography. Early diagnosis and adequate treatment is important to avoid unnecessary investigation, prevent complications (especially adhesive capsulitis of the shoulder), and reassure the patient. PMID- 10699519 TI - Electric injury, part III: cardiac monitoring indications, the pregnant patient, and lightning. AB - A number of publications have presented recommendations for prolonged cardiac monitoring after electric injury. These recommendations are combined in this article to make criteria for monitoring that agree with the recommendations of most studies. It is generally agreed that cardiac monitoring is not needed unless there is an abnormal electrocardiogram or other suggestion of acute problems. Electric injury in pregnancy is also discussed, with recommendations for fetal monitoring after electric and mechanical trauma. Reports of fetal death after minor electric injury to the mother are described. Finally, selected aspects of lightning injury are described, including neurologic and vascular effects. This is the final article of a 3-part series on electric injury. PMID- 10699520 TI - Fatal tetanus in a drug abuser with "protective" antitetanus antibodies. AB - Tetanus is a rare disease in the United States. From 1995-1997, the average annual incidence of tetanus was 0.15/1,000,000 population. Injecting-drug users, particularly those who use heroin, are among the highest risk population for acquiring tetanus. We present a case of an injecting-drug user who was seen in the emergency department with worsening diffuse midthoracic back pain and spasms. He subsequently developed acute respiratory failure and central nervous system hypoxic injury. Serum obtained before administration of tetanus immune globulin showed a tetanus antibody titer greater than 16 times the level considered protective. Because of limited human data on the minimum protective level of neutralizing antibody, as well as reports of tetanus among individuals with "protective" antibody titers, the diagnosis of tetanus should not be excluded solely on the basis of antitetanus titers. PMID- 10699521 TI - Defecation-induced bronchospasm. AB - Acute asthma exacerbations are common. Patients with asthma experience symptoms in response to a wide variety of stimuli, and identifying the precipitating cause may be useful in guiding treatment and preventing future attacks. A case of asthma exacerbation occurring during multiple defecations is reported. Abnormal parasympathetic tone has been implicated in the pathogenesis of certain types of asthma, and defecation can be associated with increased parasympathetic tone. This patient's pattern of defecation-related asthma exacerbations responded to prophylactic anticholinergic medication. PMID- 10699522 TI - Thyrotoxic periodic paralysis and intravenous propranolol in the emergency setting. AB - A 27-year-old male of Malaysian descent presented to the Emergency Department (ED) with rapidly progressive flaccid paralysis that quickly compromised his respiratory effort. The patient was found to have a serum potassium of 1.9 meq/L, and was diagnosed as having an acute paralytic episode secondary to thyrotoxic periodic paralysis. The paralytic attack was aborted with a combination of potassium replacement and parenteral propranolol in large doses. We report the use of a rarely described, yet possibly more effective, therapy for an acute attack of thyrotoxic periodic paralysis. PMID- 10699523 TI - Trauma and thyroid-induced tachycardia. AB - Excluding a source of hemorrhage after blunt trauma in a patient who presents with a sustained tachycardia can be challenging. This report is of two trauma patients presenting with undiagnosed thyrotoxicosis. Trauma-triggered thyrotoxicosis is rarely reported in the literature. The confusing presentation, the laboratory analysis, and the response to therapeutic intervention are reviewed. PMID- 10699524 TI - Delayed presentation of spinal stab wound: case report and review of the literature. AB - Stab wounds to the spinal cord are relatively uncommon in North America, but even rarer is the presentation of such an injury in a delayed fashion. We report a case of a 31-year-old male who presented with neurologic deficit 4 weeks after a stab wound injury to the spine. Because of worsening neurologic deficit, the retained knife fragment was operatively removed, and the patient had an uneventful recovery. The management of such an injury is discussed, with a review of the literature. PMID- 10699525 TI - Atlanto-occipital dislocation: an unusual lethal airbag injury. AB - Airbag-induced injury fatality is increasing in frequency. We present the case of a 6-year-old passenger who sustained a fatal atlanto-occipital dislocation associated with airbag deployment in a low-speed motor vehicle crash. The current literature regarding airbag fatalities and methods to ameliorate airbag-induced injury are reviewed. PMID- 10699526 TI - Medical outcome of cocaine bodystuffers. AB - To describe the clinical course of cocaine "bodystuffers" presenting to regional emergency departments, a descriptive retrospective analysis was performed on all cases of cocaine bodystuffers received by a metropolitan poison control center and associated toxicology service from January 1993 to May 1994. We identified 46 cases of patients classified as bodystuffers. Of these, 34 patients (74%) remained asymptomatic. Eight patients (18%) had mild symptoms including hypertension and tachycardia that resolved with no treatment beyond decontamination or benzodiazepines (one patient). Two patients (4%) had moderate symptoms including agitation and fever that resolved with no treatment beyond decontamination or benzodiazepines (one patient). Two patients (4%) had severe symptoms including seizure and cardiac dysrhythmia. Both died. Radiographs of the abdomen were negative for foreign body in all 23 examinations performed. Mild cocaine intoxication is common in cocaine bodystuffers. Severe intoxication can occur, resulting in death. Abdominal radiographs are not of value for stuffers ingesting cellophane-wrapped packets. More experience is needed to determine the length of intensive care monitoring that these patients require. PMID- 10699527 TI - Intentional iron overdose in pregnancy--management and outcome. AB - The objectives of our study were to 1) determine if peak maternal serum iron level or toxicity stage after intentional overdose is associated with adverse maternal-fetal outcome, and 2) describe the use of deferoxamine antidote therapy in obstetric patients. A computer search of the English language literature from 1966-1998 used the key words iron toxicity, iron poisoning, deferoxamine, and pregnancy to identify peer-reviewed papers reporting intentional iron overdoses in pregnancy. Two investigators independently extracted data from articles and their references including stage of toxicity (0 = asymptomatic, 1 = gastrointestinal symptoms, 2 = metabolic disturbance, 3 = organ failure), with differences resolved by consensus. Statistical analysis used the Student t-test, Fisher exact test, or ANOVA, as appropriate. Fourteen publications were identified, describing 61 cases of obstetric iron overdose, including one recent case at our institution. Compared with women who had lower peak levels, women with peak serum iron levels > or =400 mcg/dL were more frequently symptomatic (12/13 vs. 5/10, respectively, p = 0.05). Peak iron level > or =400 mcg/dL was not associated with increased risk of spontaneous abortion, preterm delivery, congenital anomalies, or maternal death. However, patients with stage 3 toxicity were more likely to spontaneously abort (1/3 vs. 1/56, respectively), deliver preterm (2/3 vs. 6/56, respectively), or experience maternal death (3/3 vs. 0/56, respectively). The proportions of patients treated with deferoxamine and total dosages of deferoxamine were similar by peak iron level (> or =400 vs. <400 mcg/dL) and toxicity stage (0-3). Peak iron levels > or =400 mcg/dL are associated with symptomatic iron overdose. Stage 3 toxicity is associated with spontaneous abortion, preterm delivery, and maternal death. PMID- 10699528 TI - Anthrax threats: a report of two incidents from Salt Lake City. AB - The threat of anthrax as an agent of bioterrorism in the U.S. is very real, with 47 incidents of possible exposure involving 5664 persons documented by the Federal Bureau of Investigation over a 14-month period in 1998 and 1999. The highly visible and potentially devastating effects of these threats require a well-coordinated and well-organized Emergency Medical Services (EMS) and Emergency Department (ED) response to minimize panic and reduce the potential spread of an active and deadly biologic agent. This requires planning and education before the event. We describe the events of two anthrax threats in a major metropolitan area. The appropriate EMS and ED response to these threats is outlined. PMID- 10699529 TI - Tibiofibular syndesmosis and ossification. Case report: sequelae of ankle sprain in an adolescent football player. AB - Heterotopic ossification development within the interosseous membrane of the ankle is an uncommon occurrence after routine ankle sprains. We present a case of a high school football player who sustained a syndesmosis ankle sprain. After 4 weeks, he continued to have pain, swelling, and range of motion restriction despite being treated with cryotherapy, NSAIDs, supportive taping, and progressive rehabilitation. The radiographs revealed a heterotopic ossification within the interosseous membrane of the distal extremity. The patient was initially treated conservatively and went on to have surgical excision with an excellent result. Symptomatic patients will require definitive surgery even without frank synostosis. PMID- 10699530 TI - A global inventory of hospitals using powder-free gloves: a search for principled medical leadership. AB - Scientific experimental and clinical studies have demonstrated that cornstarch on surgical and examination gloves promotes disease by acting as a reactive foreign body in tissue and serving as a vector for latex allergy. Consequently, hospitals have selected an innovative glove selection program utilizing only powder-free gloves. Healthcare workers in emergency medical systems are now wearing powder free, latex-free gloves to care for the growing number of patients sensitized to latex. A global Internet search has now identified 70 hospitals in the United States and three hospitals in Europe that use only powder-free gloves. PMID- 10699531 TI - Blunt trauma with a medical twist. PMID- 10699532 TI - Orbital cellulitis with abscess formation caused by frontal sinusitis. PMID- 10699533 TI - Iatrogenic venous air embolism. PMID- 10699534 TI - Spontaneous esophageal rupture. PMID- 10699535 TI - Comprehensive assessment of patients in palliative care: a descriptive study utilizing the INTERMED. AB - Documentation in palliative care is often restricted to medical and sociodemographic information, and the assessment of physical and psychological symptoms or the quality of life. In order to overcome the lack of comprehensive information, we have evaluated the utility of the INTERMED-a biopsychosocial assessment method to document integrated information of patients' needs-in 82 consecutive patients for whom a palliative care consultation was requested. Results confirm the biopsychosocial heterogeneity of the sample, and the importance of integrated information to clinical, scientific, educational, and health care policy agendas. The INTERMED could become a useful method to tailor interdisciplinary interventions based on comprehensive patient needs assessment. PMID- 10699536 TI - Fatigue and psychiatric morbidity among Hodgkin's disease survivors. AB - Fatigue is prevalent among cancer patients, including Hodgkin's disease survivors (HDS). Fatigue is poorly understood, and the clinical management is consequently difficult. This cross-sectional study examined how fatigue related to psychiatric morbidity among 457 HDS (aged 19-74 years, 56% males) treated during the period 1971-1991. The subjects were mailed a questionnaire including the Fatigue Questionnaire, the Hospital Anxiety and Depression Scale, and measures of previous psychiatric problems. Fatigue correlated moderately with anxiety and depression (r = 0.44 and 0.41 respectively). Twenty-six percent of the HDS had substantial fatigue for 6 months or longer (=cases). They had higher levels of anxiety (mean 7.3, 95% CI 6.4-8.1) and depression (mean 4.5, 95% CI 3.8-5.2) than the non-cases (anxiety: mean 4.3, 95% CI 3.9-4.7; depression: mean 2.1, 95% CI 1.8-2.5). Past psychiatric problems were not reported more commonly among the fatigue cases than among the non-cases. A multiple logistic regression analysis identified age (OR 1.04, 95% CI 1.02-1.06), anxiety (OR 1.2, 95% CI 1.2-1.3), and no self-reported psychiatric symptoms during treatment (OR 2.3, 95% CI 1.3-4.2) as predictors of fatigue caseness. One-half of the fatigue cases among HDS have psychological distress that might respond to treatment. Chronic fatigue among HDS is not predicted by previous psychiatric problems. PMID- 10699537 TI - Impaired neuropsychological performance in chronic nonmalignant pain patients receiving long-term oral opioid therapy. AB - The study investigated neuropsychological performance in chronic nonmalignant pain patients receiving long-term oral opioid therapy. Forty patients treated solely with regular and stable doses of an oral opioid were compared with 40 healthy volunteers. The patients received daily opioid doses of 15-300 mg of oral morphine (median: 60 mg) or equianalgesic doses of other opioids. The neuropsychological tests consisted of continuous reaction time (CRT), which measured vigilance/attention; finger tapping test (FTT), which measured psychomotor speed; and paced auditory serial addition task (PASAT), which measured working memory. Three months after the study had been carried out, 14 of the controls were retested in order to determine the reliability of the three tests. The patients performed statistically significantly poorer than the controls in all the tests. Significantly positive correlations were found between the PASAT and pain visual analogue scales (VAS). In the retesting of 14 controls, it was found that the tests showed high reliability. Vigilance/attention, psychomotor speed, and working memory were significantly impaired in chronic nonmalignant pain patients. The present study cannot determine which factors influenced the test results, but pain itself seemed to have an arousal effect on working memory. PMID- 10699538 TI - Knowledge and use of sublingual nitroglycerin and cardiac-related quality of life in patients with chronic stable angina. AB - Relationships among sublingual nitroglycerin (SLN) knowledge and use, general aspects of angina self-management, and quality of life were investigated in 95 patients with angina (age 63 +/-11 years). Quality of life was measured using a reliable and valid questionnaire, the Seattle Angina Questionnaire. Older age (p = 0. 04), male gender (p = 0.0001), more recent diagnosis with coronary artery disease (p = 0.02), and distant recall of SLN instruction (p = 0.02) predicted poorer SLN knowledge (R(2) = 0.26, p < 0.001). Male gender (p = 0.001) predicted more difficulty with SLN use (R(2) = 0.15, p = 0.005). A "bad" experience with SLN was associated with poorer quality of life (r = -22, p = 0.047). Sixty-five percent lacked knowledge about using SLN to prevent symptoms and 32.6% took SLN for symptoms other than chest pain. Findings support the need for more frequent reinforcement of patient education, especially in the areas of preventive use of SLN and side effect management. PMID- 10699539 TI - Management of cancer treatment-related diarrhea. Issues and therapeutic strategies. AB - The cancer treatment-related diarrhea caused by acute graft-versus-host disease (GVHD) and chemotherapeutic agents, particularly fluoropyrimidines and irinotecan, significantly affects patient morbidity and mortality. The mechanisms causing cancer treatment-related diarrhea are not fully understood, but histopathologic evidence points to a multifactorial process that causes an absorptive and secretory imbalance in the small bowel. Cancer treatment-related diarrhea could be life-threatening, yet assessment and treatment are not currently standardized. Several clinicians participated in a closed roundtable meeting to review the mechanisms of chemotherapy-induced diarrhea (CID) and GVHD induced diarrhea, management issues in cancer treatment-induced diarrhea, and pharmacologic approaches to treatment. The meeting produced a proposal for new treatment guidelines and an algorithm, which include the use of octreotide for the management of CID- and GVHD-induced diarrhea. The development of diarrhea assessment guidelines that expand on the current National Cancer Institute criteria and allow for better patient management was also proposed. PMID- 10699540 TI - How useful is docusate in patients at risk for constipation? A systematic review of the evidence in the chronically ill. AB - The effectiveness of docusate for constipation has not been studied in the terminally ill. Controversy also exists concerning its effectiveness in the chronically ill. Because chronically ill patients and terminally ill patients have several risk factors for constipation in common, we undertook a systematic review of prospective controlled trials of oral docusate in the chronically ill to clarify the utility of this drug in populations with advanced disease. The data sources were Medline 1966-April 1997, CINAHL 1982-April 1997, Current Contents August 1996-April 1997, Cochrane Library, a hand search of Index Medicus 1940-1966, three palliative care journals, references in relevant articles and texts, and direct contact with experts. Prospective controlled trials evaluating oral docusate in humans with chronic illness and identifiable risk factors for, or preexisting, constipation were selected. Only materials abstracted in English or French were considered. Information was collected by two independent reviewers and included patient demographic data, study design, dose of docusate, outcomes of stool consistency, stool frequency, need for other laxatives, and assessment of methodologic and reporting quality. Of nine identified studies, four were eligible. These incorporated three different designs and sample sizes that ranged from 15 to 74. Quality assessment scores were low (range 0.46-0.52 with a perfect score being 1.0). Three studies were flawed in blinding of treatment allocation and the use of co-interventions. All studies showed a small trend toward increased stool frequency on docusate. Because of significant clinical heterogeneity in the identified studies, pooled data analysis was not feasible. At present, the use of docusate for constipation in palliative care is based on inadequate experimental evidence. Randomized controlled trials with chronically ill patients and patients with advanced disease are needed to determine its role in prevention and treatment of constipation. PMID- 10699541 TI - Symptoms in adults with lung cancer. A systematic research review. AB - Health care providers play a key role in providing adequate symptom management and promoting quality of life during chronic illness. Several studies have noted that adults with lung cancer experience more symptom distress than patients with other types of cancer. Therefore, symptom management in this group of patients is particularly important. An understanding of the research conducted in this area is important for further knowledge development and for potentially improving symptom management. This paper presents a systematic review of empirical studies examining symptoms in adults with lung cancer. The results of this systematic review revealed that although major strides have been made in understanding symptoms associated with lung cancer, further progress is needed to decrease the morbidity associated with uncontrolled symptoms. Theoretical, conceptual, and/or methodological issues identified through this review must be addressed in future research. In particular, the researcher should provide information about the theoretical or empirical framework guiding the study, give an explicit definition about the dimensions of the symptom experience being studied, report refusal rates and attrition, and use instruments that are reliable and valid. PMID- 10699543 TI - Commentary. The role of neuromuscular blockade in palliative medicine PMID- 10699542 TI - Case presentation: Undertreatment of pain: a risk associated with neuromuscular blockade in the intensive care unit. PMID- 10699544 TI - Commentary: Professionalism and pain management. PMID- 10699545 TI - Origins of an epidemic: the methodological and political emergence of rapid assessment. AB - The recent emergence of rapid assessment as a public health tool in the drug and alcohol field has been a relatively ahistorical process. The lack of such an explicit historical biography is understandable - the findings and impact of rapid assessment have rarely made their way into mainstream journals. However, its continued absence presents the manifest danger of an inward-looking field. This paper describes the development of rapid assessment during the past two decades, with a detailed focus on the emergence of rapid assessment in the drug and alcohol field. This focuses on the central role played by international agencies during the 1980s and 1990s, and the development of rapid methodologies by the World Health Organization to prevent epidemics of HIV among injecting drug users, and the use of rapid methodologies by the United Nations International Drug Control Programme to instruct drug policy reform. The paper also describes key events in other fields including: the emergence of the first formal rapid methodologies in the late 1970s, and the production of the first formal guidelines on conducting rapid assessment during the mid-1980s. The paper concludes by highlighting common challenges that rapid methodologies have faced throughout their history. PMID- 10699546 TI - Rapid assessment and response to injecting drug use in Madras, south India. AB - HIV infection among injecting drug users (IDUs) is preventable, and in order to develop appropriate interventions, an assessment was carried out at Madras, South India using the Rapid Assessment and Response Guide on Injecting Drug Use developed by WHO. Data were collected with multiple methods from multiple sources using the principles of triangulation and induction. A total of 100 IDUs were interviewed. These interviews were complemented by focus groups and observations. A community advisory board ensured community ownership and participation. Findings showed that heroin, buprenorphine, diazepam and avil were the drugs most commonly injected. The use of pharmaceutical preparations as a 'cocktail' was also prevalent. Drug injectors interviewed were males, and most (81%) were from low-income groups living in slums. Direct (69%) as well as indirect sharing (94%) was common. Such unhygienic injecting practices, and the lack of access to sterile water, contribute to the high incidence of adverse health consequences. Compared with the buprenorphine injectors, heroin injectors were more likely to share injecting equipment (P=0.0022), inject more frequently (P=0.0013), have more drug using network members (P=0.0104), frequent 'shooting' locations (P=0.002), use the dealer's place to inject (P=0.0317), and face threats of arrest (P=0.0023). Many buprenorphine injectors managed their life without serious crises, and seemed to adopt a 'natural' harm reduction response. Sexual risk behaviour was prevalent among opioid users, and a history of commercial sex was associated with daily alcohol use (P=0.0221). The assessment led to an action plan which was presented and endorsed in an advocacy meeting by key stake-holders and decision-makers. The critical importance of implementing quality, accessible, community-oriented, and effective HIV interventions with the capacity to reach the majority of IDUs is discussed. Public health responses to injecting drug use must target changes among individuals at-risk, as well as in the community and risk environment. PMID- 10699547 TI - A multi-centre rapid assessment of injecting drug use in India. AB - In 1998, a series of five rapid situation assessments (RSA) of injecting drug use were undertaken by The Society for Service to Urban Poverty (SHARAN) covering the major Metropolitan cities of Mumbai, Chennai, Calcutta, Delhi and Imphal. The RSA determined the extent and patterns of injecting drug use (IDU), the available responses, current and planned interventions, and drug users' perceptions of injecting and sexual-related risk behaviour. The RSA was necessary as there are a lack of data on IDU in India. This has resulted in the denial of injecting drug use except for the north-eastern states by official sources, thereby affecting the inputs for IDU-related interventions. The draft assessment reports were disseminated though city workshops, held between April 1998 and January 1999. Local NGOs involved in drug treatment and HIV related interventions, government officials, and the relevant State AIDS Cells were invited to the workshops in order to contribute to final city assessment reports, so as to promote ownership and to enhance coverage. While the data obtained from the RSA were largely as anticipated, the outcome of the dissemination workshops was phenomenal. PMID- 10699548 TI - Rapid assessment of drug use and HIV vulnerability in south-east and east Asia. AB - In an 8-week period in late 1997, an assessment of the situation of drug use and HIV vulnerability in east and south-east Asia was carried out for the United Nations Joint Programme on AIDS (UNAIDS). It served to assist UNAIDS' Asia Pacific team in setting priorities for action at a regional level, it having been realised that epidemics of HIV among injecting drug users (IDUs) were playing an important role in the development of the AIDS epidemic in Asia at both country and regional levels. Though essentially a desk exercise, contact with the extensive membership of the Asian Harm Reduction Network allowed a deeper and more efficient investigation than would otherwise have been possible, with access to key informants and 'grey' literature. The assessment found a situation of massive epidemics of HIV among IDUs either occurring, or about to occur, in most Asian countries; and parlous or non-existent public health responses to these problems in a context void of policy. As well as providing evidence to guide UNAIDS, and useful for advocacy by a wide range of people, the process of the situation assessment also generated interest and in some cases activity on the part of many individuals and institutions throughout the region. It is hoped that this will be the beginning of an ongoing monitoring of the situation. PMID- 10699549 TI - 'First steps': using rapid assessment and response methods to develop research, intervention and advocacy capacity for addressing drug use in Rosario City, Argentina. AB - During the last decade, injecting drug use (IDU) has increased in Rosario City, Argentina, and more than half of all reported HIV cases are thought to be related to the sharing of injection equipment. Despite this, valid and systematic data on the extent and nature of injecting drug use are rare, and only a limited number of HIV prevention interventions currently operate in the city. In response, the Universidad Nacional de Rosario have used the Spanish translation of the Rapid Assessment and Response Guide on Injecting Drug Use (IDU-RAR) developed by the World Health Organization (WHO. The Rapid Assessment and Response Guide on Injecting Drug Use (draft for field testing). Geneva: WHO, 1998) as a first step in producing more detailed research data, identifying urgently needed interventions, and designing a long-term harm reduction strategy for Rosario. This paper describes this process and the outcomes of the assessment where key findings indicate that: 65% of those sampled were HIV positive; 70% shared injecting equipment; the majority were not in contact with drug treatment services; 13% were under the age of 20; and 62% of those injecting drug users with HIV had neither medical care or treatment for their condition. PMID- 10699550 TI - Rapidly responding to injecting drug use and HIV in Brazil: a field-report from Sao Vicente, Sao Paulo State. AB - As part of Phase II of the World Health Organisation (WHO) Injecting Drug Use Multi-City Study, Rapid Assessment and Response (RAR) methodology was used in Sao Vicente City (Sao Paulo State, Brazil) to analyse the situation regarding injecting drug use and HIV infection. Over a period of 8 weeks, analysis of existing information, focus groups (four) and depth interviews (45) were conducted, with qualitative data analysis taking a further 12 weeks. This work was undertaken by a team of six researchers (working mostly part-time on the project), and the research process was overseen by an Advisory Community Group. Key topics addressed included: the general drug use situation; HIV/AIDS and drugs; and harm reduction. Through the use of qualitative analysis it was found that there was: a negative moral status associated with drug use and drug users; prejudices against this group; and a lack of information about health services and HIV prevention programmes within Sao Vicente city. In regard to risk behavior, injecting drug users reported awareness of risks and methods of prevention. However, safer sex was considered difficult by all participants. Interventions were proposed by the research team to minimize problems and gaps in provision, and some of these have already occurred. A review of existing quantitative data found that some key institutions do not collect systematic data on drug use. Conclusions of the study include the need for interventions and the progress on Harm Reduction proposals and AIDS prevention among IDUs. PMID- 10699551 TI - Injecting drug use in Romania: a field-report based on an initial assessment. AB - There are indications of increased injecting drug use in Romania associated with the increased cross-border traffic of heroin. At the same time, there are indications of HIV transmission associated with drug injecting in countries close by or neighbouring Romania. In the absence of data on patterns of injecting drug use, we undertook a preliminary assessment in four major Romanian cities (Bucharest, Iasi, Constanta and Timisoara) to describe the extent and nature of drug injecting and its associated adverse health consequences. The assessment is the first of its kind on injecting drug use in Romania. We found that little information exists on injecting drug use. In addition, we found recruiting IDUs into the assessment difficult. We note that future assessments need to find ways of recruiting IDUs directly in the community. We also note the need for future assessments to concentrate in greater detail on the potential for intervention developments in prevention, health promotion, treatment and policy. PMID- 10699552 TI - Processes and outcomes of training on rapid assessment and response methods on injecting drug use and related HIV infection in the Russian Federation. AB - In September 1997, Medecins Sans Frontieres-Holland (MSF-H) began a project to provide training and support for HIV/AIDS prevention among injecting drug users (IDUs) in the Russian Federation, focusing on the use of the World Health Organization Rapid Assessment and Response Guide on Injecting Drug Use, and the European Peer Support Manual. As part of the training, participants are asked to carry out a rapid situation assessment (RSA) in their city or region as a major step towards designing and implementing an effective program to prevent HIV transmission among IDUs. This paper focuses on the first four training cycles of the programme (from January 1998 to January 1999), in which 89 health professionals and others from 32 Russian cities took part. A total of 28 rapid situation assessments were completed or almost completed by participants during these four cycles. The paper provides an overview of the methods used and major problems faced by participants undertaking these assessments, as well as summarising the 14 harm reduction programmes which resulted. PMID- 10699553 TI - From kidnapping to corruption: some trials and tribulations in the implementation of rapid assessment studies. AB - In between the description of the methodology and the discussion of assessment results, lies a crucial area, namely that of implementation. If assessment methods cannot be implemented, there will be no assessment report, and no recommendations for intervention or policy development. Implementation is a critical, yet neglected, area of drug policy delivery. This paper describes some of the problems associated with the implementation of rapid assessment studies commissioned by the United Nations International Drug Control Programme (UNDCP) over the last 5 years. Drawing on the author's experience of commissioning rapid assessments on the extent and nature of drug use in a variety of international settings and political contexts, the paper draws eight main lessons for rapid assessments commissioned within the context of international development programmes. These are: (1) the need to get high level political support; (2) the need to employ competent social scientists to run the assessment; (3) the need to define exactly the object of the assessment; (4) the necessity of spelling out the methodology and the time scale; (5) the careful establishment of the financial framework; (6) the importance of cultivating contacts; (7) the necessity of actually getting a report; and (8) the danger of accepting an unseen or unread consultant's report as the basis for programme implementation. The paper emphasises the critical importance of planning prior to implementation and flexibility during it. PMID- 10699554 TI - The empirical and methodological comparative value of the rapid assessment of drug use patterns. AB - This paper examines the comparative value of rapid assessment in the field of illicit drug use. Three aims of rapid assessment are highlighted: one, to collect quality data to inform policy and practice; two, to encourage a multi-method research approach; and three, to promote and support greater local involvement and ownership of the project itself. A number of manuals and guidelines have been produced for rapid assessment. A key issue to address, therefore, is the extent to which a structured and standardised approach can provide sufficient flexibility to take into account the political, social and cultural differences in the respective countries in which assessment takes place. Using the empirical experience derived from four rapid assessment projects, a four-phase strategy is outlined: developing infrastructure and formative evaluation; research training and mapping exercises; data collection; and report writing and dissemination. In place of the current reactive approach, we need more strategic thinking at the macro-level. To enable comparison between projects we should move towards basic methodological standardisation, but this should be accomplished without stifling the sociological imagination, which is a crucial ingredient to successful rapid assessment. PMID- 10699555 TI - Muscle activation in the contralateral passive shoulder during isometric shoulder abduction in patients with unilateral shoulder pain. AB - Studies have shown an increased muscle activation at the opposite passive side during unilateral contractions. The purpose of the present study was to examine the influence of pain on muscle activation in the passive shoulder during unilateral shoulder abduction. Ten patients with unilateral rotator tendinosis of the shoulder and nine healthy controls performed unilateral maximal voluntary contractions (MVC) and sustained submaximal contractions with and without subacromial injections of local anaesthetics of the afflicted shoulder. Muscle activation was recorded by electromyography (EMG) from the trapezius, deltoid, infraspinatus and supraspinatus muscles in both shoulders. During MVCs, the EMG amplitude from muscles of the passive afflicted side was not different in patients and controls, and was not influenced by pain alterations. In contrast, the EMG amplitude from the muscles of the passive unafflicted side was lower in the patients and increased after pain reduction. During the sustained submaximal contraction the EMG amplitude increased gradually in the passive shoulder to 15 30% of the EMG amplitude observed during MVC. This response was not influenced by differences in pain. We conclude that muscle activation of the passive shoulder was closely related to the activation of the contracting muscles and thus related to central motor drive, and not directly influenced by changes in pain. PMID- 10699556 TI - The comparison of trunk muscles EMG activation between subjects with and without chronic low back pain during flexion-extension and lateral bending tasks. AB - The purpose of the study was to compare the electromyographic (EMG) activity of the trunk muscles between normal subjects and chronic low back pain (CLBP) patients during standardized trunk movements. Thirty-three male subjects (18 normals, 15 suffering from non specific CLBP) aged between 35 and 45 yr participated. A biomechanical analysis involving the recording of EMG signals from 12 trunk muscles, the kinematics of trunk segments and the computation of L5/S1 moments was performed. The subjects performed flexion-extension and lateral bending (left and right) tasks (three complete cycles) with and without a 12 kg load. Between group comparisons were performed on the full cycle average pattern of all biomechanical variables for each task. The reliability of EMG variables was evaluated for 10 subjects (5 normals and 5 CLBP) who performed the tasks on three different days. The reliability of EMG amplitude values was generally excellent for agonist muscles but poor to moderate for antagonists. The EMG amplitude analysis revealed significant differences between groups for some muscles (left lumbar and thoracic erector spinae). The abnormal (asymmetric) EMG patterns detected among CLBP patients were not explained by postural asymmetries. PMID- 10699557 TI - Muscle activities used by young and old adults when stepping to regain balance during a forward fall. AB - The current study was undertaken to determine if age-related differences in muscle activities might relate to older adults being significantly less able than young adults to recover balance during a forward fall. Fourteen young and twelve older healthy males were released from forward leans of various magnitudes and asked to regain standing balance by taking a single forward step. Myoelectric signals were recorded from 12 lower extremity muscles and processed to compare the muscle activation patterns of young and older adults. Young adults successfully recovered from significantly larger leans than older adults using a single step (32.2 degrees vs. 23.5 degrees ). Muscular latency times, the time between release and activity onset, ranged from 73 to 114 ms with no significant age-related differences in the shortest muscular latency times. The overall response muscular activation patterns were similar for young and older adults. However older adults were slower to deactivate three stance leg muscles and also demonstrated delays in activating the step leg hip flexors and knee extensors prior to and during the swing phase. In the forward fall paradigm studied, age differences in balance recovery performance do not seem due to slowness in response onset but may relate to differences in muscle activation timing during the stepping movement. PMID- 10699558 TI - Sensitivity of trapezius electromyography to differences between work tasks - influence of gap definition and normalisation methods. AB - Surface electromyography (EMG) has been used extensively to estimate muscular load in studies of work related musculoskeletal disorders, especially for the trapezius muscle. The occurrences of periods of EMG silence (gaps), the time below a predetermined threshold level (muscular rest) and various percentiles of the amplitude distribution (APDF) are commonly used summary measures. However, the effects of the criteria used to calculate these measures (e.g., gap duration, threshold level, normalisation method) on the sensitivity of these measures to accurately differentiate work loads is not well known. Bilateral trapezius EMG was recorded, for a full workday, for 58 subjects following both maximal (MVE) and submaximal (RVE) reference contractions. Gap frequency, muscular rest, and percentiles were derived for eight fundamental work tasks. The calculations were performed using different gap duration criteria, threshold levels and normalisation methods.A gap duration of less than 1/2 s, and threshold level approximately 0.3% MVE for gap frequency, and approximately 0.5% MVE for muscular rest, were the criteria that optimised sensitivity to task differences. Minimal sensitivity to tasks and a high sensitivity to individuals was obtained using gap frequency with a threshold level of approximately 1% MVE. Normalisation to RVE, rather than MVE, improved sensitivity to differences between tasks, and reduced undesirable variability. Muscular rest was more sensitive to task differences than APDF percentiles. PMID- 10699559 TI - Muscle pre- and coactivity during downward stepping are associated with leg stiffness in aging. AB - We have previously reported that elderly compared to young women executed downward stepping with substantially greater leg stiffness. Because antagonist muscle coactivity increases joint stiffness we hypothesized that increased leg stiffness in aging is associated with increased muscle coactivity. We also explored the possibility that the magnitude of the preparatory muscle activity preceding impact also differed between young and old subjects. Young (n=11, 20. 8 yr) and old (n=12, 69 yr) women performed downward stepping from a platform set at 20% body height. The leg was modeled as a simple mass-spring system. From video and ground reaction force data leg stiffness was computed as the ratio of force under the foot and the linear shortening of the limb. EMG activity of the vastus lateralis, biceps femoris, gastrocnemius lateralis, and tibialis anterior were recorded with a telemetric system. Elders compared to young subjects had 64% greater leg stiffness during downward stepping. Muscle activity over a 200-ms period preceding touch down was 136% greater in elderly than in young subjects. Biceps femoris and tibialis anterior coactivity during ground contact was 120% greater in the elders. Muscle pre- and coactivity, respectively, accounted for about 50% of the variance in leg stiffness. In conclusion, elderly people elevate muscle pre- and coactivity during downward stepping to stiffen the leg in compensation for impaired neuromotor functions. PMID- 10699560 TI - Test of a new technique for the diagnosis of carpal tunnel syndrome. AB - Several new techniques for carpal tunnel syndrome diagnosis have been developed in the last few years. This work tests a technique that compares the distal motor latency of the median nerve to the second lumbrical muscle (2L) with the distal motor latency of the ulnar nerve to the interossei muscle (INT). Results from 40 normal hands give the superior limit of the normal difference (2L-INT) as 0. 26 ms (&xmacr;+3 SD). In 55 hands with different levels of carpal tunnel syndrome, this new technique was more sensitive and accurate than the conventional test which uses the distal motor latency of the median nerve to the abductor pollicis brevis muscle (APB), especially in the less severe cases. With the absence of the compound muscle action potentials of the APB muscle caused by severe thenar atrophy, it is much easier to obtain the potential from the 2L muscle. We concluded that this is a sensitive, simple, rapid, and non-invasive new technique, and therefore, it should be incorporated as part of the routine ENMG procedures for carpal tunnel syndrome diagnosis. PMID- 10699561 TI - Differential angle-dependent modulation of the long-latency stretch reflex responses in elbow flexion synergists. AB - We determined the effect of elbow joint angle on the short-(M1) and long-latency stretch reflex (M2 and M3) responses of the elbow flexion synergists, the brachioradialis (BR), and the biceps brachii (BB), during weak isometric elbow flexion tasks. The elbow joint angle was 35,75 and 115 degrees (full-extension angle was 0 degrees ), and the muscle contraction level was 0,3 and 6% of maximum voluntary contraction (MVC) of the BR. In BR, the M1, M2 and M3 responses were significantly greater at 75 and 115 degrees than at 35 degrees. On the other hand, in BB, the M2 response was significantly greater at 35 degrees than at 75 and 115 degrees, while the M1 and M3 responses were not significantly different among the elbow joint angles. These results indicated that the stretch reflex responses of BR might be dependent on the changes of muscle length in stretch stimulus, while the M2 response of BB might not be dependent on the actual stimulus intensity. Therefore, we concluded that the M2 of BB might be modulated selectively by a higher reflex center in accordance with relationships of the mechanical advantages between synergistic muscles. PMID- 10699562 TI - Signal separation of background EEG and spike by using morphological filter. AB - A signal separation method for extracting background electroencephalogram (EEG) from EEG containing spikes was proposed. Morphological filters were designed for extracting spike waveforms, and then the background EEG was obtained by subtracting the detected spike waveforms from the EEG with spike. The proposed method was evaluated by using simulated EEG data, which consisted of a summation of EEG without spike and model waveform of typical spike. The background EEG separated by the method was processed by the automatic background EEG interpretation. PMID- 10699563 TI - Optimal control of FES-assisted standing up in paraplegia using genetic algorithms. AB - A practical system for Functional Electrical Stimulation (FES) assisted standing up in paraplegia should involve only a minimum of manual set up and tuning. An improved tuning method, using a genetic algorithm (GA) is proposed and demonstrated using computer simulation. Specifically, the GA adjusts the parameters of fuzzy logic (FL) and gain-scheduling proportional integral derivative (GS-PID) controllers that electrically stimulate the hip and knee musculature during the sit-stand maneuver. These new GA designed controllers were found to be effective in coordinating volitional and FES control according to formulated criteria. The latter was based on the deviations from a desired trajectory of the knee and hip joints and the magnitude of the voluntary upper body forces. The magnitude of the average arm forces were slightly higher when compared with the open-loop maximal stimulation of the hip and knee musculature; however, the terminal knee velocities were significantly reduced to less than 10 degrees /s. For practical implementation, the number of trials required to optimize the FL and GS-PID controllers can be reduced by a proposed pre-training procedure using a computer model scaled to the individual. The GA designed controllers remain near optimal provided the model-subject mismatch is small. PMID- 10699564 TI - A new sensor for monitoring chest wall motion during high-frequency oscillatory ventilation. AB - The recently developed technique of fibre optic respiratory plethysmography (FORP) has been modified to monitor the rapid, small amplitude movements of the chest wall during high-frequency oscillatory ventilation (HFOV). The FORP sensor is an expandable belt encircling the chest, in which is housed a fibre optic loop that alters its radius of curvature as a function of chest perimeter. These curvature changes cause variations in macrobending losses of light transmitted through the fibre, which are proportional to the chest perimeter. Dynamic measurement of transmitted light intensity can hence be used to monitor chest wall motion (CWM). For application to HFOV, the design of the FORP belt was altered to increase sensitivity and the materials were chosen to maximise macrobending effects induced by the CWM. FORP was tested in four piglets ventilated with HFOV, both in the normal and surfactant-deficient lung. Measurement of CWM was possible over the full range of tidal volumes and ventilation frequencies used during HFOV. In all cases, the measured frequency of the CWM fell within 3% of the applied ventilation frequency. In addition, the technique was sufficiently sensitive to detect changes in the amplitude of CWM in response to changes in applied tidal volume. It is anticipated that application of this new non-invasive measurement device will lead to an increased understanding of the dynamics of chest and abdominal wall motion during HFOV. PMID- 10699565 TI - A three-dimensional kinematic model of the human long finger and the muscles that actuate it. AB - A kinematic model of the human long finger and the six muscles that actuate it is presented. The model transforms finger pose into estimates of muscle excursions and fingertip location. The effects of abduction/adduction about the metacarpo phalangeal joint are accounted for, as are the effects of flexion of the three finger joints. A set of parameters are provided which approximate kinematics of the segments and muscles of a cadaver finger over the range of finger poses humans normally assume. PMID- 10699566 TI - Mechanical characterization in shear of human femoral cancellous bone: torsion and shear tests. AB - In order to investigate and compare the mechanical behaviour of human cancellous bone during different shear loading modes, two tests were performed to characterise human femoral cancellous bone in shear: a torsion test until failure and a shear test using a sharpened stainless steel tube. Paired cylindrical samples were core drilled from 12 human femoral heads, symmetrically with respect to the coronal plane and along the primary trabecular direction. The distal part of the sample was assigned to a torsion test and the shear test was performed on the proximal part along two perpendicular anatomical directions. Apparent densities and tissue densities were measured on both torsion and shear specimens. The mean torsion properties were shear modulus G, 289 (183) MPa, ultimate stress tau(torsion), 6.1 (2.7) MPa, ultimate strain gamma(ultimate), 4.6 (1.3)%, yield stress tau(yield), 4.3 (1.9) MPa and yield strain gamma(yield), 1.8 (0.3)%. Strong correlation was obtained between G and tau(torsion) (r'=0.853, p<0.001). These torsion properties were correlated with apparent density of torsion specimens showing, respectively: r'=0.713, p=0.005 and r'=0.671, p=0. 008. Properties from the shear test were invariable with regard to the two tested directions then isotropic ultimate shear stress and isotropic elementary shear stress, which represent the mean values of the two tested directions were, respectively, tau(shear), 10.0 (4. 5) MPa and tau(elem), 18.8 (6.1) MPa. Both shear stresses were correlated with apparent density of shear specimens: tau(shear), r'=0.564, p=0.045 and tau(elem), r'=0.636, p=0.024. Apparent densities for shear specimens were superior than for torsion specimens (p=0.06) and the comparison was the opposite for tissue densities (p=0.028), showing strong density gradients of cancellous bone in the femoral head. These torsion and shear tests which permit the evaluation of cancellous bone behavior under two different types of shear loading, may be performed on different human sites and the measured shear properties may be compared to structural properties of cancellous bone. PMID- 10699567 TI - An in vitro investigation of the dependence on sample thickness of the speed of sound along the specimen. AB - To measure the speed of sound (SOS), most quantitative ultrasound (QUS) devices use the transmission mode, whereby two transducers are placed on opposite sides of the sample. This mode is limited to a few specific skeletal sites because of the varying configuration of bone geometry and varying amounts of overlying soft tissue at most other sites. The aim of this study was to address the dependence of SOS measured along the sample on the thickness and composition of the bone sample. Bovine samples from mid-femur and trochanter, and perspex phantoms were used. We prepared the perspex samples in the shapes of blocks and cylinders to investigate the effect of wall thickness on SOS. The thickness of the blocks was decreased in decrements of 1 mm; a 22 mm diameter hole was drilled through the cylindrical samples and the hole size was gradually increased. The second configuration was also used with the bovine samples. For each experimental set-up five SOS measurements were acquired, with the probe aligned along the sample and a mean value computed. All measurements were taken with castor oil as the coupling agent, and in the cylindrical cases, the oil was used to fill the tube. The measurement precision determined as the root mean square coefficient of variation (RMSCV) was determined to be 0.14% and 0.65% for perspex and bovine samples respectively. The measured SOS on the perspex phantom (2760+/-4 m/s) was within the published values for bulk velocity. It was observed that for both perspex and bovine samples the SOS was independent of sample wall thickness greater than the wavelength (2.2 mm, 2.7 mm and 3.5 mm for perspex, trochanter and mid-femur respectively). The SOS decreased with sample wall thickness smaller than the wavelength in concordance with theoretical predictions. The SOS values obtained for bovine samples reflected either totally cortical (mid-femur) or a composite of cortical and cancellous bone (trochanter). PMID- 10699568 TI - An instrument for the multiparameter assessment of speech. AB - This paper describes the development of SNORS+, a clinical, user-friendly instrument for measurement of the articulators during speech. The design criteria for the instrument were based upon a wide-ranging review of current practice and available techniques. SNORS+ allows objective assessment of the function and co ordination of key articulators. Appropriate targeting of therapy is therefore possible. Visual feedback is provided, for therapy, and an objective measurement of outcome is easily obtained. Preliminary results are presented. These suggest that the instrument will prove extremely useful in the assessment and management of many speech disorders. PMID- 10699569 TI - Paraquat induces different pulmonary biochemical responses in Wistar rats and Swiss mice. AB - The paper presents results showing differential response to paraquat toxicity in Wistar rats and Swiss strain of mice. Paraquat-induced pulmonary biochemical responses in the two animal species were studied at different time point after giving a single intraperitoneal injection of the respective LD(10) doses of the herbicide paraquat to rats and mice. Paraquat induced different biochemical responses including different protective responses in the two animal species. As a protective response, NADPH-specific quinone reductase is induced in rats, while catalase is induced in mice. It is implied that an early induction of catalase in mice as opposed to rats may account for the resistance of Swiss mice to paraquat toxicity. Xanthine oxidase, which was induced in rats, remains unaffected in mice indicating that the enzyme contributes to paraquat toxicity only in Wistar rats. Time-course studies were also conducted to compare the differential responses of antioxidant enzymes and lipid peroxidation between the two species. The results of the study led us to suggest that the manifestation of paraquat toxicity involve distinct differences in early pulmonary biochemical responses in Wistar rats and Swiss mice. PMID- 10699570 TI - Response of Drosophila melanogaster to selection for P450-mediated resistance to isoquinoline alkaloids. AB - Several species of columnar cacti in the Sonoran Desert contain isoquinoline alkaloids that are toxic to all but the resident drosophilids that feed and breed in necrotic stems. Cytochrome P450 enzymes are known to be involved in the metabolic detoxification of these alkaloids by the desert Drosophila and are consequently responsible for their ability to utilize these substrates. D. melanogaster is not normally exposed to these xenobiotic compounds and cannot live in necrotic cactus tissue. However, a previous study found evidence of a phenobarbital-inducible P450 in adults of this species that is capable of metabolizing cactus alkaloids. The current investigation sought to determine whether D. melanogaster responds to selection for alkaloid resistance. Significant increases in larval viability and adult longevity as well as shorter larvae-to-adult development times were observed after 16 generations of selection on medium containing isoquinoline alkaloids. The selected lines that exhibited a positive response can now be used to assay for changes in gene regulation as a possible mechanism of their response. This information will contribute to the understanding of evolution of P450-mediated resistance in insects. PMID- 10699571 TI - The induction of hepatic microsomal UDP-glucuronosyltransferase by the methylsulfonyl metabolites of polychlorinated biphenyl congeners in rats. AB - The effects of nine methylsulfonyl (MeSO(2)) metabolites of tetra-, penta- and hexachlorinated biphenyls (tetra-, penta- and hexaCBs; 20 micromol/kg once daily for 4 days) on the hepatic microsomal UDP-glucuronosyltransferase (UDP-GT) were investigated in male Sprague-Dawley rats. Each of the seven 3-MeSO(2)-PCBs, 3 MeSO(2)-2, 2',4',5-tetraCB (3-MeSO(2)-CB49), 3-MeSO(2)-2,3',4',5-tetraCB (3 MeSO(2)-CB70), 3-MeSO(2)-2,2',3',4',5-pentaCB (3-MeSO(2)-CB87), 3-MeSO(2) 2,2',4',5,5'-pentaCB (3-MeSO(2)-CB101), 3-MeSO(2)-2,2',3', 4',5,6-hexaCB (3 MeSO(2)-CB132), 3-MeSO(2)-2,2',3',4',5,5'-hexaCB (3-MeSO(2)-CB141), 3-MeSO(2) 2,2',4',5,5',6-hexaCB (3-MeSO(2)-CB149) and 4-MeSO(2)-2,2',4',5,5'-pentaCB (4 MeSO(2)-CB101) increased the activities of UDP-GT toward chloramphenicol, 4 nitrophenol and 4-methylumbelliferone. 4-MeSO(2)-2,2',4',5,5',6-hexaCB (4-MeSO(2) CB149) increased the activity of UDP-GT toward chloramphenicol (UGT2B1) but not toward 4-nitrophenol (UGT1A6) and 4-methylumbelliferone (UGT1A6). The activity of UDP-GT toward thyroxine (T(4)) significantly increased after the administration of each of the seven 3-MeSO(2)-PCBs and 4-MeSO(2)-CB101. Significant correlation was found between the activity of UDP-GT toward T(4) and serum total T(4) concentration after the administration of each of the MeSO(2) derivatives except 4-MeSO(2)-CB149. In conclusion, seven 3-MeSO(2)-PCBs and 4-MeSO(2)-CB101 induce both UGT2B1 and UGT1A6, and 4-MeSO(2)-CB149 induces UGT 2B1. The results from the present study indicate that increase in the hepatic T(4) glucuronidation after the administration of the seven 3-MeSO(2)-PCBs and 4-MeSO(2)-CB101 possibly because of the induction of both UGT1A1 and UGT1A6 caused the reduction of serum T(4) levels. PMID- 10699572 TI - Selective fatty acid release from intracellular phospholipids caused by PCBs in rat renal tubular cell cultures. AB - The purpose of this study was to explore the influence of different polychlorinated biphenyls (PCBs) upon the release of oleic and palmitic acid from the intracellular lipids, which were previously labeled with [3H]oleic or [3H]palmitic acid, respectively. Studies have been realized with Aroclor 1248 (a commercial PCB mixture with 48% chlorine by weight), and two pure PCB congeners: 3,3',4, 4'-tetrachlorobiphenyl (a non-ortho-substituted planar congener) and 2,2',4,4',5,5'-hexachlorobiphenyl (a di-ortho-substituted nonplanar congener). The treatment of cells with Aroclor 1248 increased [3H]oleic acid release in a concentration-dependent manner. Our results showed that only the di-ortho substituted congener which prefers a nonplanar configuration stimulated the release of [3H]oleic acid from the intracellular phospholipids to the culture medium, while the exposure of cell cultures to the chosen non-ortho-substituted coplanar congener did not alter the release of [3H]oleic acid to the culture medium. Finally, none of the PCBs studied could increase the release of [3H]palmitic acid from the intracellular stores significantly. The possibility that these differential alterations in the fatty acid release affect cell function during PCB exposure should therefore be postulated. PMID- 10699573 TI - Kinetics of DNA alkylation, depurination and hydrolysis of anti diol epoxide of benzo(a)pyrene and the effect of cadmium on DNA alkylation. AB - Anti benzo[a]pyrene diol epoxide (BPDE) alkylates guanines of DNA at N7 in the major groove and at the exocyclic amino group in the minor groove. In this report we investigated the rates of BPDE hydrolysis, DNA alkylation and subsequent depurination of BPDE-adducted pBR322 DNA fragment using polyacrylamide gel electrophoresis. Preincubation studies showed that it hydrolyzed completely in triethanolamine buffer in <2 min. The depurination kinetics showed that a fraction of the N7 alkylated guanine depurinated rapidly; however a significant amount of N7 guanine alkylation remained stable to spontaneous depurination over a 4-h period. Similar results were obtained for the hydrolysis and alkylation rates of syn isomer but it required nearly 500 times more concentration to induce similar levels of N7 guanine alkylation. Cadmium ion strongly inhibited the N7 guanine alkylation of both isomers. But the minor groove alkylation was not affected as demonstrated by postlabeling assay which confirmed the presence of heat-and cadmium-stable minor groove adducts in BPDE-treated calf thymus DNA. Based on these and our earlier findings, we propose a mechanism for the synergistic effect of cadmium in chemically induced carcinogenesis. PMID- 10699574 TI - Cytokine vaccination: neutralising IL-1alpha autoantibodies induced by immunisation with homologous IL-1alpha. AB - High-affinity IgG autoantibodies (aAb) to IL-1alpha are among the most frequently found aAb to cytokines in humans. To establish an animal model with aAb to IL 1alpha, we immunised mice with recombinant murine IL-1alpha. Unprimed and Bacille Calmette-Guerin (BCG)-primed BALB/cA mice were vaccinated with IL-1alpha coupled to purified protein derivative of tuberculin (PPD). Both unprimed and primed animals developed IgG aAb to IL-1alpha. These aAb persisted at high levels more than 100 days after vaccination and did not cross-react with murine IL-1beta. The induced anti-IL-1alpha aAb inhibited binding of IL-1alpha to the murine T-cell line NOB-1 by simple competition and neutralised IL-1alpha, but not IL-1beta induced IL-6 in vivo. The aAb did not induce visible discomfort in the animals. In conclusion, long-lasting and high levels of neutralising and specific IgG aAb to IL-1alpha can be induced in mice by vaccination with recombinant murine IL 1alpha conjugated to PPD. Studies of the effects of IL-1alpha aAb in such animals may help clarify the importance of naturally occurring IL-1alpha aAb in humans and permit the evaluation of future therapies with cytokine aAb in patients with immunoinflammatory diseases and cytokine-dependent tumours. PMID- 10699575 TI - Rapid and sensitive immunomagnetic-electrochemiluminescent detection of staphyloccocal enterotoxin B. AB - Sensitive, rapid and reproducible detection of staphyloccocal enterotoxin B (SEB) in a range of different biological matrices was achieved using the ORIGEN((R)) Immunoassay System (Igen, Inc). The homologous immunoassay format consisted of a double antibody sandwich in which a biotinylated capture antibody, pre-bound to streptavidin-coated paramagnetic beads, was used to bind antigen from test samples. A detector antibody, labeled with ruthenium (II) tris-bipyridal chelate, was added and, when bound to the bead immunocomplex, generated light in the presence of an excess of tripropylamine. The light was detected and measured by the ORIGEN analyzer. The sensitivity of this assay was 1 pg of enterotoxin per ml of serum, urine, tissue, or buffer and was highly reproducible. Concentration curves generated from SEB standards produced consistently wide linear ranges (0.1 100 ng/ml), making quantitation possible with only two dilutions of sample (undiluted and 1:1000). The assay used 50 microl of sample per test and required a 30 min incubation period in addition to a 1 min per tube reading time (50 tubes maximum). This assay was significantly better in terms of sensitivity, linear range, and assay time than the standard microplate enzyme-linked immunosorbent assay and should permit early SEB detection in clinical samples, food, and environmental samples. PMID- 10699576 TI - Time-resolved fluorometric assay for leukocyte adhesion using a fluorescence enhancing ligand. AB - A new 96-well microtiter plate, time-resolved fluorometric assay was developed to measure leukocyte adhesion in vitro. The assay is based on loading leukocytes with a fluorescence enhancing ligand 2,2':6', 2"-terpyridine-6,6"-dicarboxylic acid (TDA), which in its acetoxymethyl ester form readily diffuses through the cell membrane. After hydrolysis by nonspecific intracellular esterases, the impermeable TDA accumulates inside the cells. When the TDA-labeled adherent leukocytes are lysed, the ligand is released and reacts with europium present in the lysis solution to produce a highly fluorescent and stable chelate. The fluorescence signal can be measured by time-resolved fluorometry and correlates directly with the number of adherent cells. In this study, we have optimized both the TDA-labeling and adhesion assay conditions in isolated human neutrophils. Furthermore, we have compared the assay with a traditional microscopic counting method. This time-resolved fluorometric assay provides a rapid, reproducible and convenient method for the routine analysis of leukocyte adhesion. PMID- 10699578 TI - Survival-promoting activity of IL-7 on IL-2-dependent cytotoxic T lymphocyte clones: resultant induction of G1 arrest. AB - Interleukin-7 (IL-7) is essential for both T cell and B cell development. Recent studies have suggested that IL-7 also functions as a survival-promoting factor for resting and activated T cells. In this study we examined the effects of IL-7 on survival and cytotoxicity of tumor-specific CD8(+) cytotoxic T lymphocyte (CTL) clones established and maintained either with IL-2 alone or with a combination of IL-2 and IL-7. While the CTL clones cultured in IL-2 alone died around day 10, the CTL clones cultured in the presence of IL-2 and IL-7 survived for more than 4 weeks after seeding. The long-term survival of the latter was correlated with the presence of IL-7 in the medium. In addition, IL-7 alone prolonged survival of other IL-2-dependent CTL clones after the removal of IL-2. IL-7 maintained the CTLs in G1 arrest after a slight proliferation during the initial phase during which low-level but sustained DNA synthesis was observed. However, there was no direct correlation between DNA synthesis and enhancement of long-term survival by IL-7 as demonstrated by the inhibiting proliferation of the CTL clones with the protein kinase inhibitor genistein. During long-term survival in the presence of IL-7, the cytotoxic activities of the CTL clones decreased gradually to background levels although they were restored soon after the next passage. These results suggested that IL-7 had the ability to set machinery in motion against apoptosis in the IL-2-dependent CTL clones. Such an effect of IL-7 might play a role in vivo in the process leading activated T cells to the resting, that is, memory state. PMID- 10699577 TI - Chemical agents and enzymes used for the extraction of gut lymphocytes influence flow cytometric detection of T cell surface markers. AB - Chemical agents (DTT, EDTA) and enzymes (collagenase, DNAse) are often used for the generation of a single cell suspension from tissue samples. In this study, flow cytometry was used to examine the effect of chemical agents and enzymes on the expression of cell membrane markers of T lymphocytes from tonsils and peripheral blood. Expression of CD4, CD8, CD25, CD38, L-selectin, CD44, alphaEbeta7 and alpha4beta7 were studied. Incubation of lymphocytes with DTT and EDTA resulted in a decrease of CD38, alphaEbeta7 and alpha4beta7 expression. Incubation with collagenase A and DNAse resulted in a decrease of CD25, L selectin, alphaEbeta7 and alpha4beta7. The results of this study indicate that a careful interpretation is necessary for phenotypic descriptions of lymphocyte populations obtained by enzymatic isolation techniques. PMID- 10699579 TI - Facile generation and use of immunogenic polypeptide fusions to a sparingly soluble non-antigenic protein carrier. AB - Many researchers attempt to prepare antipeptide antibodies by immunizing animals with preparations of fusion proteins or conjugates between the target peptide and a larger protein (such as GST or KLH). Often, the immune response to the larger protein dominates. We have engineered a protein to be sparingly soluble in aqueous solution and nonantigenic, and show that fusions of this sparingly soluble non-antigenic protein (SSNAP) to target peptide sequences can be purified easily to a point suitable for immunizations. When animals are immunized with such fusion proteins, the majority of the immune response is to the target peptide. In all three cases tested, the peptide-specific immune response generated using the SSNAP carrier was greater than that obtained with peptides chemically linked to BSA or KLH, or expressed as fusion proteins to GST. The SSNAP carrier induced a very early IgG response with all classes of IgG well represented in the specific antibody response. All of the SSNAP fusion peptide derived antibodies were capable of recognizing the full-length target protein in both ELISA and Western analysis. Based on the superior performance of the SSNAP antigens, these studies suggest this novel strategy will have broad utility for the generation of peptide antibodies. PMID- 10699580 TI - Development of a continuous IL-7-dependent murine pre-B cell line PB-1 suitable for the biological characterisation and assay of human IL-7. AB - Human interleukin-7 (IL-7) is a cytokine that appears to be critical for early T- and B-cell development and although IL-7 is currently under investigation as a therapeutic agent in a variety of hematolymphopoietic disorders, there have been few instances of the detection or investigation of this cytokine using a biological assay. This has been due, in the main, to the lack of a widely available, stable, easy to maintain and use, IL-7 responsive cell line. We have developed a pre-B-cell line, PB-1, from murine bone marrow, that is dependent on IL-7 for growth and has been maintained continually for up to 1 year without loss of responsiveness. The cells survive freezing and reviving, having been stored for periods of up to 4 years. The IL-7 bioassay is reproducible and sensitive, able to reliably detect 50 pg/ml IL-7. The assay is completely unresponsive to any other stimulatory cytokines tested and is not affected by a wide variety of inhibitory cytokines, with the exception of high levels of interferon alpha. The assay can be made completely specific for human IL-7 by including specific neutralizing antibodies for IL-7 and has been shown to be suitable for the estimation of IL-7 in both plasma and serum samples. PMID- 10699581 TI - Protein transfer of the costimulatory molecule, B7-2 (CD86), into tumor membrane liposomes as a novel cell-free vaccine. AB - Several approaches have been taken to enhance the immunogenicity of tumors. Genetically-modified tumors expressing various cytokines, major histocompatibility complex (MHC) molecules, or costimulatory molecules such as B7 1 (CD80) or B7-2 (CD86) can induce tumor-specific immune responses. In the present study, an alternative approach was explored based on direct protein transfer of purified recombinant B7-2 into tumor cell membranes. B7-2 was purified from recombinant baculovirus infected insect cells. Although differentially glycosolyated, the recombinant B7-2 retained the function to costimulate T-cell proliferation. Purified B7-2 was readily incorporated into tumor membranes using a detergent dialysis technique to form unilameller liposomes. The immunogenicity of tumor membrane proteoliposomes was significantly increased by incorporation of B7-2. These findings suggest an alternative method for the introduction of immunostimulatory molecules into tumor membranes to create novel tumor vaccines. PMID- 10699582 TI - An improved method for the immunological detection of mineral bound protein using hydrofluoric acid and direct capture. AB - Immunological detection of proteins adsorbed to mineral and ceramic surfaces has proved not only difficult but controversial. Unlike the immunological detection of proteins associated with carbonate or phosphate minerals (e.g. shells and bones) proteins adsorbed to siliceous minerals cannot readily be removed by dissolution of the mineral phase. We have previously examined alternative extraction methodologies which claim to bring the protein into solution, but found none of these to be effective. Here we report a novel strategy for immuno detection of proteins adsorbed to siliceous minerals, the Digestion and Capture Immunoassay (DACIA). The method involves the use of cold, concentrated (4M) hydrofluoric acid (HF) with the simultaneous capture of liberated protein onto a solid phase. The combination of low temperatures and surface stabilisation enables us to detect epitopes from even partially degraded proteins. The method may have a wide application in forensic, archaeological, soil and earth sciences. PMID- 10699583 TI - Production of monoclonal antibodies to porcine interleukin-18 and their use for immunoaffinity purification of recombinant porcine interleukin-18. AB - We have recently reported the cloning and expression of porcine interleukin-18 (IL-18). In this study, we describe the production of anti-porcine IL-18 monoclonal antibodies (mAb) and their use in the purification of a large amount of recombinant porcine IL-18 by immunoaffinity column chromatography. Five monoclonal antibodies (2-2-B, 2-5-B, 2-13-C, 3-1-C and 5-3-B) were established and characterized. Three (2-2-B, 3-1-C and 5-3-B) of them were of IgG1 subclass, and the other two were IgMs. Epitope analysis of the three IgG1 mAbs showed that they recognized the same epitope. All five mAbs demonstrated reactivity with baculovirus generated porcine IL-18 by immunoblot analysis. Biologically active porcine IL-18 was obtained by immunoaffinity chromatography using anti-porcine IL 18 mAb at more than 85% purity from culture supernatants of Trichoplusia ni (Tn5) derived cells infected with recombinant baculovirus containing the coding sequence of porcine mature IL-18. These results suggest that the anti-porcine IL 18 mAbs established in this study are useful for one-step purification of porcine mature IL-18 as well as the detection of porcine IL-18 by immunoblotting. PMID- 10699584 TI - Characterisation and specificity of two single-chain Fv antibodies directed to the protein tyrosine kinase Syk. AB - In order to obtain single chain Fv fragments (scFv) specific for the protein tyrosine kinase Syk, we screened a human synthetic phage-display library. Two glutathione S-transferase (GST):Syk fusion proteins containing both SH2 domains of Syk were used to perform three rounds of selection of the library. Among the scFv fragments resulting from the third round of selection, the ones specific for the GST portion of the fusion proteins were eliminated by performing enzyme linked immunosorbent assay tests on GST:Syk versus GST coated plates, and the monoclonal scFv fragments binding only to the GST:Syk coated plates with high affinities were further analysed. We report here the in vitro characterisation of G4G11 and G6G2 anti-Syk scFvs. G4G11 shows the best performance in immunoprecipitation and immunofluorescence experiments, and G6G2 is able to detect Syk in immunoprecipitation, immunofluorescence and on Western blots. Both scFvs are also able to detect the phosphorylated form of Syk, and neither of them binds to Zap-70, the other member of the Syk family of protein tyrosine kinases. PMID- 10699585 TI - Guinea pig C3 specific rabbit single chain Fv antibodies from bone marrow, spleen and blood derived phage libraries. AB - We constructed combinatorial immunoglobulin libraries from the whole rabbit antibody repertoire of bone marrow, spleen and peripheral blood of a rabbit immunized with guinea pig complement protein C3. By means of the phage display technology we selected guinea pig C3 specific single chain Fv (scFv) antibodies from each of the libraries. None of the scFv antibodies cross reacted with guinea pig C3a, human C3 or rat C3. The frequency of bone marrow derived C3 positive clones was much higher as compared to blood or spleen derived clones. Additionally bone marrow and spleen derived clones show higher diversity than clones, obtained from blood, as determined by fingerprint analysis with the restriction enzyme AluI. Dissociation rate constants for all scFvs were similar, indicating that the source of the scFvs had no influence on affinities. The antibody fragments were used to analyze complement activation during xenotransplantation. Several blood or bone marrow derived scFvs bound to C3 located on rat liver endothelium after hyperacute rejection of a heterotopically transplanted rat liver into guinea pig. These data demonstrate that monoclonal rabbit scFvs can be easily generated from recombinant phage display libraries, constructed from spleen, blood or bone marrow. The selected guinea pig C3 specific scFvs appear to be useful to detect complement activation during xenotransplantation in guinea pigs. PMID- 10699586 TI - Sheep monoclonal antibody fragments generated using a phage display system. AB - Monoclonal sheep antibodies have great potential for biomedical, veterinary and agricultural purpose. Although conventional sheep monoclonal antibodies can be generated by a modified hybridoma technology, the procedures are not routine. Here, we describe a method to generate recombinant sheep antibody fragments from immunised animals using a modified phage display system. Total RNA from pooled spleens of sheep immunised with the model antigens human serum albumin and conalbumin were used to amplify immunoglobulin V gene repertoires and an efficient two-step cloning method was employed to rapidly construct a phage display single-chain Fv (scFv) library. A total of 14 different scFvs were isolated and characterised. Sequence analysis indicated typical ovine immunoglobulin characteristics. Thirteen Vlambda and 11 VH genes were identified that could be grouped into the sheep Vlambda families I, II, VI and a single VH family. Soluble monomeric scFvs, produced in the periplasm of Escherichia coli, were subjected to affinity measurement via surface plasmon resonance analysis and affinities typical of the secondary immune response were observed. The method described here should be of value for the study of sheep immunology as well as for biorecognition in general. PMID- 10699587 TI - Characterizing the functionality of recombinant T-cell receptors in vitro: a pMHC tetramer based approach. AB - The very low affinity of the T-cell receptor (TCR) for the peptide-major histocompatibility complex (pMHC) has made it very challenging to design assays for testing the functionality of these molecules on small scales, which in turn has severely hampered the progress in developing expression and refolding methodologies for the TCR. We have now developed an ELISA assay for detecting pMHC binding to functional recombinant TCRs. It uses tetramers of biotinylated pMHCs bound to a neutravidin-horseradish peroxidase conjugate and detects the presence of functional TCR, bound in a productive orientation to an immobilized anti-Cbeta antibody. Specificity can be stringently demonstrated by inhibition with monomeric pMHCs. The assay is very sensitive and specific, and requires only very small amounts of protein. It has allowed us to study the unstable recombinant TCR P14, which we expressed and refolded from Escherichia coli. The TCR P14 is directed against the most abundant epitope of LCMV. We have confirmed the specificity of the interaction by BIAcore, and were able to determine the dissociation constant of the interaction of the P14 TCR and of the gp33-pMHC as 6 microM. This affinity ranks it among the tighter ones of TCR-pMHC interactions, and unusually low affinity thus does not seem to be the cause of the modest protective power of these T-cells, compared to others elicited in the anti-LCMV response. This strategy of multimerizing one partner and immobilizing the other in both a native form and productive orientation should be generally useful for characterizing the weak interactions of cell-surface molecules. PMID- 10699588 TI - "Affinity maturation" of ligands for HCV-specific serum antibodies. AB - We have previously screened a phage-displayed random peptide library using sera from patients and identified ligands binding to antibodies specifically associated with the hepatitis C virus infection. The ability of these peptides to detect HCV-specific antibodies was improved through an in vitro procedure which mimics the natural process of antibody affinity maturation operating in secondary immune response. Libraries were generated by mutating the sequence of the original peptide through a protocol that efficiently introduced substitution, insertion and deletion mutations on a single or population of clones. Screening these libraries isolated mutants that displayed increased specific reactivity with a broader range of sera from HCV-infected patients. Several variants of the original peptide were identified which discriminate between the various components of the specific polyclonal response. This methodology to select artificial ligands from RPL using sera and to enhance their diagnostic properties by affinity maturation makes the development of a diagnostic assay to detect disease-associated antibodies feasible, without requiring the natural antigen. PMID- 10699589 TI - Relationships between structure and molting hormonal activity of tebufenozide, methoxyfenozide, and their analogs in cultured integument system of Chilo suppressalis Walker. AB - The molting hormonal activity of methoxyfenozide (RH-2485), tebufenozide (RH 5992), five analogs with various alkyl groups, and 18 acyl analogs was measured by using cultured integument of rice stem borers, Chilo suppressalis Walker. The hormonal activity of methoxyfenozide was remarkably high (EC(50) = 1.1 x 10(-9) M), being equivalent to that of tebufenozide (RH-5992). The hormonal activity of several tebufenozide analogs with varying alkyl groups such as CH(3), n-C(3)H(7), i-C(3)H(7), n-C(4)H(9) and n-C(5)H(11) at the para-position of the benzene ring furthest from the tert-butyl group was lower than that of tebufenozide (alkyl group is C(2)H(5)). The activity decreased to varying degrees as a result of replacement of the 3,5-dimethylphenyl moiety of tebufenozide with either a phenyl, naphthyl, or cyclohexyl group. Both 1- and 2-naphthyl derivatives were very active (EC(50) = 4.3 x 10(-8) M and 3.2 x 10(-8) M, respectively) without any significant difference between them. The activity of the 1-cyclohexenyl analog (EC(50) = 1.0 x 10(-7) M) was about 40x that of the corresponding 3 cyclohexenyl analog (EC(50) = 4.4 x 10(-6) M), but 1/100 that of tebufenozide. The activity varied parabolically with respect to the molecular hydrophobicity, and decreased with longer acyl moieties. PMID- 10699590 TI - The effect of 7-oxo-DHEA acetate on memory in young and old C57BL/6 mice. AB - 7-Oxo-dehydroepiandrosterone, which can be formed from dehydroepiandrosterone (DHEA) by several mammalian tissues, is more effective than its parent steroid as an inducer of thermogenic enzymes when administered to rats. Using the Morris water maze procedure, we tested DHEA and its 7-oxo-derivative for their ability to reverse the memory abolition induced by scopolamine in young C57BL/6 mice, and for their effect on memory in old mice. A single dose of 7-oxo-DHEA-acetate at 24 mg/kg b.w. completely reversed the impairment caused by 1 mg of scopolamine per kg b.w. (P < 0.001). DHEA (20 mg/kg) was also effective (P < 0.01). In old mice given the same single doses followed by feeding 0.05% of the respective steroid in the diet, memory of the water maze training was retained through a four week test period in mice receiving 7-oxo-DHEA-acetate (P < 0.05) but not in the control or DHEA-treated groups. When old mice were not tested until five weeks after being trained 7-oxo-DHEA exerted a slight, but statistically insignificant, improvement in memory retention. The possible effect of 7-oxo-DHEA in human memory problems deserves investigation. PMID- 10699591 TI - New marine steroids, yonarasterols, isolated from the okinawan soft coral, Clavularia viridis. AB - Six new marine steroids, yonarasterols A through F, were isolated from the Okinawan soft coral, Clavularia viridis. Their structures were determined based on the results of spectroscopic analysis. PMID- 10699592 TI - Ecdysteroids from the caribbean sponge Iotrochota birotulata. AB - The sterol composition of the Caribbean sponge Iotrochota birotulata was investigated. Structure of a new ecdysteroid 2beta,3beta,14alpha, 20beta tetrahydroxy-22alpha-(2-hydroxyacetiloxy)-5b eta-colest-7-en-6- one (1) was assigned on the basis of spectroscopic and chemical evidence and molecular mechanics calculations. Isolation of the widespread ecdysteroids 2-5 is also reported. PMID- 10699593 TI - A convenient synthesis of 5beta-cholestan-26-oic and 5beta-cholestan-26,27-dioic acids. AB - A new method for the preparation of 5beta-cholestan-26-oic acids 7 and their analogs is described. The key steps in the synthesis are: iodination of bis- and tris-formyloxy-5beta-cholan-24-ols 3; nucleophilic substitution of iodides 4 with diethyl sodiomalonate; complete alkaline hydrolysis of esters 5; and subsequent decarboxylation of geminal diacids 6 in DMSO. PMID- 10699594 TI - Guinea pig 11beta-hydroxysteroid dehydrogenase type 1: primary structure and catalytic properties. AB - The 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) enzyme is responsible for the interconversion of glucocorticoids and their inactive metabolites, and thus modulates the intracellular level of bioactive glucocorticoids. The present study was designed to clone and characterize 11beta HSD1 in the guinea pig, a laboratory animal known for resistance to glucocorticoids. The cDNA encoding guinea pig 11beta-HSD1 was cloned by a modified 3'-RACE (rapid amplification of cDNA ends) protocol using the hepatic RNA as template. The cloned cDNA encodes a protein of 300 amino acids that shares 71 to 74% sequence identity with other known mammalian 11beta-HSD1 proteins. Sequence comparison analysis revealed that the deduced guinea pig 11beta-HSD1 was longer, by eight amino acids at the C terminus, than those of other mammals. Moreover, one of the two absolutely conserved consensus sites for N-glycosylation was absent. To examine the functional significance of these structural changes, we also characterized 11beta-HSD1 activity in the hepatic microsomes. Although the guinea pig hepatic enzyme was NADP(H)-dependent and reversible, it displayed equal affinity for cortisol and cortisone (apparent K(m) for both substrates was 3 microM). This is in marked contrast to 11beta-HSD1 in other mammals whose affinity for cortisone is approximately 10 times higher than that for cortisol (apparent K(m) of 0.3 vs. 3.0 microM). The apparent lower affinity of the guinea pig enzyme for cortisone would suggest that the intracellular bioformation of cortisol from circulating cortisone may be less efficient in this species. Northern blot analysis and RT-PCR revealed that the mRNA for 11beta-HSD1 was widely expressed in the adult guinea pig but at low amounts. In conclusion, the present study has identified distinct features in the deduced primary structure and catalytic function of 11beta-HSD1 in the guinea pig. Thus, the guinea pig provides a useful model in which the structural determinants of catalytic function of 11beta-HSD1 may be studied. PMID- 10699595 TI - Synthesis of 11beta-(4-dimethylaminophenyl)-17beta-hydroxy-17alpha- (3-methyl-1 butynyl)-4, 9-estradien-3-one and 11beta-(4-acetophenyl)- 17beta-hydroxy-17alpha (3-methyl-1-butynyl)-4, 9-estradien-3-one: two new analogs of mifepristone (RU 486). AB - From the structure activity relationship, two new analogs, 2 and 3, of the potent progesterone antagonist mifepristone 1 have been designed. The syntheses of these two analogs have been achieved in eleven steps through modified synthetic sequences and improved procedures starting from (+)-estrone. In comparison with mifepristone 1, the relative binding affinities of compound 2 for the progesterone receptor was found to be more, whereas that of compound 3 was less. PMID- 10699596 TI - The size and/or configuration of the cycloalkane D' ring in pentacyclic progesterone derivatives are crucial for their high-affinity binding to a protein in addition to progesterone receptor in rat uterine cytosol. AB - [(3)H]labeled progesterone and a number of its 16alpha, 17alpha-cycloalkano derivatives with an additional three to six-membered D' ring were investigated for mutual competition and equilibrium binding to proteins from rat uterine cytosol. The interaction of all studied [(3)H]ligands with proteins was characterized by comparable affinity (K(d) in nM region) and apparent homogeneity in terms of affinity. At the same time, the concentrations of binding sites for ligands bearing 16alpha,17alpha cyclopentano, cyclohexano, or cyclohexeno substituents were several-fold higher than those for progesterone or 16alpha, 17alpha-cyclopropanoprogesterone. In mutual competition experiments, when [(3)H]progesterone or [(3)H]16alpha, 17alpha-cyclopropanoprogesterone were used, the curves of 'bound radioactivity-log of competitor concentration' for all compounds studied were parallel and corresponded to a model of 'one protein-two ligands.' However, when [(3)H]ligands with bulky 16alpha, 17alpha-substituents (with the possible exception of cyclohexene derivative) were used, competitive curves for various ligands had different appearances and fell into two groups. Parallel curves for derivatives with 5 or 6 carbons in D' ring described by a model of 'one protein-two ligands' formed the 1st group. The 2nd group comprised curves for progesterone or 16alpha, 17alpha-cyclopropanoprogesterone that had lower slopes and could be described by a model of 'two proteins-two ligands.' Taken together, the results suggest the presence in rat uterine cytosol, of a protein in addition to progesterone receptor capable of discriminating between ligands with no or small 16alpha, 17alpha-cycloalkano substituents and ligands with more bulky substituents. PMID- 10699597 TI - The case for external beam treatment of early-stage prostate cancer. PMID- 10699598 TI - Brachytherapy. PMID- 10699599 TI - Management of patients with rising prostate-specific antigen after radical prostatectomy. PMID- 10699600 TI - G(1)/S cell cycle proteins as markers of aggressive prostate carcinoma. PMID- 10699601 TI - Clinical states in prostate cancer: toward a dynamic model of disease progression. PMID- 10699602 TI - Parenteral estrogen versus total androgen ablation in the treatment of advanced prostate carcinoma: effects on overall survival and cardiovascular mortality. The Scandinavian Prostatic Cancer Group (SPCG)-5 Trial Study. AB - OBJECTIVES: To compare the effect on overall survival of total androgen ablation (TAA) with that of parenteral estrogen and to pay special attention to cardiovascular mortality. TAA (orchiectomy or a luteinizing hormone-releasing hormone analogue combined with an antiandrogen) has been proposed as superior to other endocrine treatments for patients with prostate carcinoma. Recently, the use of parenteral estrogen has been suggested to reduce or even negate the well known cardiovascular side effects of oral estrogens. METHODS: Nine hundred fifteen patients were randomized to intramuscular injections of 240 mg polyestradiol phosphate (PEP) every second week for the first 8 weeks (5 doses) followed by a maintenance dose of 240 mg every month (n = 458) or to bilateral orchiectomy or triptorelin 3.75 mg every month combined with the antiandrogen flutamide 250 mg three times daily. The choice between orchiectomy and triptorelin was at the discretion of the clinician and patient. Patients were stratified according to performance status, presence of cardiovascular disease, and alkaline phosphatase level. An observer totally unaware of the treatment given classified all deceased patients. RESULTS: At a median follow-up of 18.5 months, no signs of a difference in overall survival were found between TAA and PEP (P <0.001). Of 458 patients, 266 (58.1%) had died in the PEP group compared with 269 (58.9%) of 457 patients in the TAA group. Within the TAA group, no difference in overall survival existed between patients who had undergone orchiectomy or who were given triptorelin. Furthermore, no differences in cardiovascular mortality were found (3.5% in the PEP group and 3.1% in the TAA group). CONCLUSIONS: The current parenteral estrogen regimen seems to be of comparable efficacy and cardiovascular safety as TAA in terms of overall survival. PEP has by far the lowest drug cost and also the lowest cumulative direct costs and thus has the highest cost-effectiveness. We suggest that parenteral estrogen be included as a therapeutic option in the endocrine management of prostate carcinoma. PMID- 10699603 TI - Editorial comment PMID- 10699604 TI - Reply by the authors PMID- 10699605 TI - Indications for surgical intensive care unit admission of postoperative urologic patients. AB - OBJECTIVES: To analyze the practice of surgical intensive care unit (SICU) admission of postoperative urologic patients and to define objective criteria to predict active treatment requirements and length of stay in the SICU. METHODS: The records of 90 consecutive patients admitted to the SICU postoperatively in the 12-month period from January 1996 to December 1996 were retrospectively reviewed. The Acute Physiology and Chronic Health Evaluation II (APACHE II) score was calculated from patient parameters acquired within the first 12 hours. The correlation of outcome variables to the length of stay and the requirements for active treatment in the SICU were analyzed and used to develop a risk stratification model. This algorithm was subsequently validated on a population of 46 patients who underwent radical cystectomy the following year. RESULTS: Only the preoperative American Society of Anesthesia class, the event of an intraoperative complication, and the APACHE II score were statistically significant (P <0.05) predictors of length of stay and active treatment. The patients were subsequently categorized into high and low-risk groups, which were found to have mean SICU stays of 39.9 +/- 3.92 hours and 20.2 +/- 0.45 hours, respectively (P = 0. 001), and an active SICU-specific treatment rate of 58.0% and 14.3%, respectively (P = 0.001). These results were confirmed in the validation population. CONCLUSIONS: Postoperative risk stratification may be helpful in predicting SICU requirements in the immediate postoperative period and in identifying patients at lower or higher risk of an adverse outcome. PMID- 10699606 TI - Anesthetic aspects of laparoscopic and open adrenalectomy for pheochromocytoma. AB - OBJECTIVES: To compare the anesthetic aspects and intraoperative hemodynamic data and immediate postoperative outcomes in patients whose pheochromocytoma resection was performed either laparoscopically or by traditional open surgery. METHODS: Fourteen consecutive patients who underwent laparoscopic procedures (a single surgeon) were compared with 20 patients who underwent open surgery. The patients' records were reviewed for demographic information, preoperative medical history and therapy, intraoperative hemodynamic data, fluid balance, and immediate postoperative course. RESULTS: No differences between the highest intraoperative blood pressures and number of hypertensive episodes between the two groups were found. However, in laparoscopic patients, the intraoperative hypotension was less severe (mean lowest blood pressure 98/57 mm Hg versus 88/50 mm Hg, P = 0.05), and the hypotensive episodes were less frequent (median 0 versus 2, P = 0.005) and required fewer interventions with vasopressors (P = 0.02). Extreme high and extreme low heart rates did not differ between the two groups. The estimated blood loss was lower in the laparoscopic group (P = 0.0001), but the total intraoperative fluid requirement and operative times were similar in the two groups. Patients in the laparoscopic group resumed walking earlier (median 1.5 versus 4 days, P = 0.002) and resumed oral food intake sooner (median 1 versus 3.5 days, P = 0.0001). The median duration of hospitalization in patients who underwent laparoscopic and open adrenalectomy was 3 and 7.5 days, respectively (P = 0.001). CONCLUSIONS: Intraoperative hemodynamic values during laparoscopic adrenalectomy for pheochromocytoma were comparable to those of traditional open surgery, but the patients who underwent the laparoscopic procedure had a faster postoperative recovery. PMID- 10699607 TI - Renal colic in pregnant women: role of renal resistive index. AB - OBJECTIVES: To investigate the value of the renal resistive index (RI) in the identification of acute renal obstruction in pregnant women. METHODS: The study included 22 pregnant women with acute unilateral ureteral obstruction due to a stone disease (group A), 71 normotensive pregnant patients without loin pain (group B), and 20 nonpregnant women of child-bearing age with both kidneys normal (group C). All patients underwent Doppler ultrasound (DUS) with determination of the RI and the difference between the RI of the corresponding and contralateral kidney (DeltaRI). The RI and DeltaRI was considered positive for obstruction with a value of 0.70 or greater and 0.04 or greater, respectively. Ureteral obstruction was confirmed by several clinical, radiologic, and endoscopic findings. The sensitivity, specificity, and overall accuracy of RI and DeltaRI for the diagnosis of acute unilateral ureteral obstruction were calculated. RESULTS: In group A, kidneys with ureteral obstruction (n = 22) had a mean RI of 0.69 +/- 0.03; the contralateral normal kidneys (n = 22) had a mean RI of 0.63 +/ 0.03, a significant difference (P <0.0001). The mean RI of all kidneys in group B (n = 142) and all kidneys in group C (n = 40) was 0.64 +/- 0.05 and 0.62 +/- 0.04, respectively; the difference was not statistically significant. A comparison between the mean RI of the normal kidneys of group A and all the kidneys of groups B and C revealed no significant difference. The mean RI of the obstructed kidneys in group A was significantly higher than the mean RI of all the kidneys in groups B and C. Similarly, the mean DeltaRI of group A was significantly higher than the mean DeltaRI of groups B and C (0.06 +/- 0.01 versus 0.006 +/- 0.003 versus 0.006 +/- 0.004, respectively). The RI was sensitive in 45%, specific in 91%, and accurate in 87%. The corresponding values for DeltaRI were 95%, 100%, and 99%. CONCLUSIONS: The DeltaRI is a sensitive and specific test that can replace intravenous urography in the diagnosis of acute unilateral ureteral obstruction in pregnant women. PMID- 10699608 TI - Accuracy of diagnosis by guided biopsy of renal mass lesions classified indeterminate by imaging studies. AB - OBJECTIVES: To define the accuracy, safety, and impact of percutaneous biopsies of indeterminate mass lesions as an additional diagnostic tool. The vast majority of renal mass lesions are routinely diagnosed by radiographic features alone. However, with the increased use of computed tomography scanning and ultrasound, many smaller renal masses, which are "indeterminate" (refractory to categorization on the basis of imaging alone), are now being discovered. METHODS: We retrospectively reviewed 583 patients (364 male and 219 female) with indeterminate renal mass lesions diagnosed by imaging studies that were further investigated by percutaneous biopsy. Patients were followed up for at least 5 years if the biopsy result demonstrated a benign lesion, or they underwent surgical exploration if the biopsy result demonstrated a malignancy. Biopsy or aspiration material was assessed by histopathologic and cytologic evaluation and, when appropriate, with biochemistry, Gram stain, culture, and sensitivity. The biopsy site was localized by computed tomography, ultrasound, or fluoroscopy. RESULTS: Five hundred eighty-three patients with indeterminate renal mass lesions (representing 7.2% of all renal masses diagnosed from 1967 through 1996) were diagnosed by imaging studies complemented by guided biopsy. Sixty-six patients were lost to follow-up, leaving 517 patients who were analyzed. In 393 cases (76%), the imaging-guided biopsy provided a definitive diagnosis. The incidence of false diagnoses was 1.2% (7 biopsies). In 124 of the cases (21%), imaging guided biopsy was unable to determine the etiology of the lesion with acceptable confidence; of these, 21 biopsies did not provide enough material to establish the diagnosis (16.9%). CONCLUSIONS: Overall, percutaneous biopsy of the kidney has proved to be a safe and accurate diagnostic procedure, with impact on the management of cystic or solid renal lesions. PMID- 10699609 TI - Randomized, double-blind study of electrical stimulation for urinary incontinence due to detrusor overactivity. AB - OBJECTIVES: To evaluate the usefulness of electrical stimulation for urinary incontinence due to detrusor overactivity in a randomized, double-blind manner. METHODS: Sixty-eight patients (29 men, 39 women, 70.0 +/- 11.2 years) were studied. Detrusor overactivity was urodynamically defined as involuntary detrusor contractions of more than 15 cm H(2)O during the filling phase. Ten-hertz square waves of 1-ms pulse duration were used. A vaginal electrode was used in the women and an anal or surface electrode in the men. The stimulation was given for 15 minutes twice daily for 4 weeks. The efficacy was evaluated on the basis of a frequency/volume chart and urodynamic study before and after treatment. RESULTS: Thirty-two patients in the active group and 28 in the sham group completed the study. The patient impressions were very good or good in 59% and 39% of the active and the sham group, respectively (P = 0.0354). On the cystometrogram, the bladder capacity at the first desire to void and the maximum desire to void increased significantly (P = 0.0104 and P = 0.0046, respectively) in the active group, but not in the sham group. Seven patients in the active group and 1 patient in the sham group were cured (P = 0.0324); 26 patients (81.3%) in the active group and 9 (32.1%) in the sham group improved (P = 0.0001). Of 17 patients in the active group, 13 remained cured or improved for an average of 8.4 months after completion of the 4-week treatment; in the sham group, 3 of 6 patients were cured or improved for an average of 4.7 months after completion of the 4-week treatment. CONCLUSIONS: Electrical stimulation was useful in treating urinary incontinence due to detrusor overactivity. PMID- 10699610 TI - Combined intravesical and oral oxybutynin chloride in adult patients with spinal cord injury. AB - OBJECTIVES: Detrusor hyperreflexia with elevated storage pressures presents a major risk factor for renal damage in patients with neurogenic lower urinary tract dysfunction. If standard anticholinergic treatment is unsuccessful, surgical treatment must be considered. We evaluated the effects of intravesical oxybutynin treatment on detrusor hyperreflexia in patients in whom standard oral treatment had failed. METHODS: Twenty-five patients (mean age 36. 7 years) with storage pressures greater than 40 cm H(2)O despite standard anticholinergic treatment received intravesical (15 mg three times daily) and oral oxybutynin chloride treatment. The follow-up evaluations included urodynamic testing, renal ultrasound, urine examination (urinalysis and urine culture), and evaluation of side effects. RESULTS: The mean follow-up was 6 months. Intravesical treatment led to an increase in bladder storage volume from 349 to 420 mL. The mean maximum storage pressure was significantly reduced from 54 to 26.5 cm H(2)O. Detrusor storage pressures returned to values less than 40 cm H(2)O in 21 of 25 patients. Dysreflexia was treated successfully in 3 of 5 patients. No patient developed renal damage. No severe side effects or drug-related discontinuation of treatment were observed. CONCLUSIONS: Intravesical oxybutynin therapy seems to be a safe and effective treatment option for detrusor hyperreflexia in adults and avoids surgical treatment in most patients. Long-term observations concerning side effects, acceptance, and efficacy are needed. PMID- 10699611 TI - Presence of carcinoma in situ and high 2C-deviation index are the best predictors of invasive transitional cell carcinoma of the bladder in patients with high-risk Quanticyt. AB - OBJECTIVES: Karyometric analysis (Quanticyt) has proved of value as a cytologic marker for bladder cancer. This study was conducted to identify diagnostic and prognostic factors in a high-risk Quanticyt population to predict the prognosis of transitional cell carcinoma (TCC) of the bladder. METHODS: Quanticyt is a karyometric system for quantitative bladder wash cytologic findings based on two nuclear features: the 2c-deviation index (2cDI) and the mean of nuclear shape. Samples are scored as low, intermediate, or high risk. Before 1995, 109 patients with high-risk quantitative bladder wash cytologic findings were identified at our clinic. Four patients with previous invasive tumors were excluded. RESULTS: Histologically proven malignancy was found in 54 of 105 patients at first high risk quantitative bladder wash cytologic findings. Invasive TCC was found in 16 patients, and another 10 patients had progression during a median follow-up of 3.7 years. In univariate analysis, the presence of carcinoma in situ (CIS), highest tumor grade, 2cDI, and highest tumor stage were significant predictors of progression. The presence of CIS proved to be the only predictor of progression in the multivariate analysis. A 2cDI of 2.00 c(2) or higher was a significant predictor of CIS, invasive TCC, and progression. At follow-up analysis after negative cystoscopy, 2cDI showed a tendency toward predicting progression. CONCLUSIONS: These data confirm earlier findings that CIS is an important marker of progression. 2cDI as assessed by quantitative cytology is a practical tool to predict CIS, invasive TCC, and subsequent progression. A 2cDI of 2. 00 c(2) can be used to further stratify high-risk quantitative bladder wash cytologic findings. PMID- 10699612 TI - Gender differences in stage distribution of bladder cancer. AB - OBJECTIVES: To compare stage distribution between men and women with bladder cancer at first presentation. METHODS: The population-based Netherlands Cancer Registry was used to investigate stage differences of newly diagnosed bladder cancer, including upper urinary tract tumors in female and male patients. RESULTS: The stage distribution at first presentation for both bladder cancer and upper urinary tract tumors was slightly worse in female patients with transitional cell carcinoma than in male patients. The stage differences were more clear in non-transitional cell carcinoma (squamous cell carcinoma, adenocarcinoma, and sarcoma) of the bladder, with female patients presenting with higher stages. Because of the large numbers, these gender differences in stage distribution were statistically significant in both TCC (P <0.0001) and non-TCC (P <0.0001). CONCLUSIONS: Treating physicians should be aware that female patients are more frequently diagnosed with higher stages at the first presentation for bladder cancer and upper urinary tract tumors. PMID- 10699614 TI - Prognostic value of nuclear morphometry on needle biopsy from patients with prostate cancer: is volume-weighted mean nuclear volume superior to other morphometric parameters? AB - OBJECTIVES: To compare the prognostic value of stereologically estimated volume weighted mean nuclear volume (MNV) with other nuclear morphometric parameters using pretreatment needle-biopsy specimens of prostate cancer. METHODS: The MNV, mean nuclear area, form factor, and coefficients of variation for nuclear area (VNA) and form factor were measured on pretreatment needle biopsy specimens from 66 patients with prostate cancer (clinical Stage B, n = 9; Stage C, n = 14; and Stage D, n = 43), all of whom underwent androgen deprivation therapy. The prognostic value of those morphometric parameters, as well as Gleason score and clinical stage, was examined in terms of cause-specific patient survival using univariate and multivariate analysis (Cox proportional hazard model). RESULTS: Univariate analysis of the nuclear morphometric parameters revealed that MNV, mean nuclear area, VNA, coefficient of variation for form factor, and clinical stage were significant prognostic factors for cause-specific patient survival. However, when the patients with Stage D disease were selectively analyzed for survival, only the VNA was a significant prognostic parameter. Furthermore, the multivariate analysis, including the morphometric parameters, clinical stage, and Gleason score revealed that only VNA and clinical stage were independent variables. CONCLUSIONS: The present comparative study could not demonstrate any prognostic superiority of MNV over other nuclear morphometric parameters in patients with prostate cancer. PMID- 10699613 TI - Percentage of free PSA in black versus white men for detection and staging of prostate cancer: a prospective multicenter clinical trial. AB - OBJECTIVES: In predominately white populations, measurement of the percentage of free prostate-specific antigen (%fPSA) has been shown to enhance the specificity of total PSA testing for prostate cancer while maintaining high sensitivity and to aid in prostate cancer staging. This study evaluated whether the %fPSA cutoff that maintained a 95% sensitivity in a white population yielded the same sensitivity and specificity in a black population and whether %fPSA was useful in predicting postoperative pathologic features in blacks. METHODS: We evaluated 647 white and 79 black men, prospectively enrolled at prostate cancer screening and surgical referral centers. Subjects were 50 to 75 years old with digital rectal examination findings that were not suspicious for prostate cancer and total PSA values between 4.0 and 10.0 ng/mL. All had undergone needle biopsy of the prostate. Hybritech's Tandem total and free PSA assays were used. RESULTS: Ninety five percent sensitivity was attained with a %fPSA cutoff of 25% in both races. Use of this cutoff could have avoided unnecessary biopsies in 20% of white and 17% of black subjects (P = 0.69). In receiver operating characteristic (ROC) curve analysis, the area under the curve (AUC) for %fPSA was significantly higher than for total PSA in both blacks (0.76 versus 0.56, P <0.01) and whites (0.70 versus 0.54, P <0.001). In both races, higher %fPSA values indicated a lower risk of cancer and also predicted favorable pathologic features in radical prostatectomy specimens. CONCLUSIONS: A 25% fPSA cutoff detected 95% of cancers and reduced unnecessary biopsies in both races. Higher %fPSA values were associated with favorable postoperative histopathologic findings in both races. PMID- 10699615 TI - Extent of extracapsular extension in localized prostate cancer. AB - OBJECTIVES: To measure the radial extent of extracapsular penetration by tumor cells, thereby providing estimates of the margins needed around target volumes. New radiotherapeutic techniques, like brachytherapy and conformal radiotherapy, irradiate small volumes and reduce the dose to periprostatic tissues. Even in the early stages of localized prostate cancer, extracapsular extension (ECE) is commonly seen. METHODS: Two hundred sixty-five consecutive radical prostatectomy specimens were analyzed for the presence of ECE. ECE was found in 92 of all cases (35%); measurements were performed in 79 of the 92 cases. A total of 98 ECE sites were evaluated in the 79 cases. The distance of tumor outside the capsule was measured in millimeters. Extension less than 0.1 mm was considered as "focal". RESULTS: The site of ECE was posterolateral in 53% of cases, lateral in 24%, posterior in 13%, and at the base in 10%. The median amount of ECE at all sites was 1. 1 mm (mean 1.7). However, the range was wide; the minimum measurable extent was 0.1 mm and the maximum 10.0 mm. The extent was within 3.8 mm for 90% of all cases. By stratifying cases with favorable and unfavorable tumors, the 90th percentiles of ECE were as follows: 3.3 mm for favorable tumors (clinical Stage T1-2, initial prostate-specific antigen 10 ng/mL or less, and biopsy Gleason score 6 or less) and 3.9 mm for unfavorable tumors (clinical Stage T3, initial prostate-specific antigen greater than 10 ng/mL, or biopsy Gleason score 7 or greater). CONCLUSIONS: Most of the ECE was at posterolateral sites. The extent of disease outside the prostate was within 4 mm in 90% of cases. Since ECE was observed in 30% to 60% of all patients with clinical Stage T1-2 prostate cancer, only 3% to 7% of all such cases would have disease extent exceeding 4 mm. The present study provides useful estimates of the amount of ECE. These estimates could be potentially used in planning the target volumes for treatment of prostate cancer with either conformal radiotherapy or brachytherapy. PMID- 10699616 TI - Clinical characteristics in black and white men with prostate cancer in an equal access medical center. AB - OBJECTIVES: To determine whether black men with newly diagnosed prostate cancer in an equal access health care center are more likely to present with metastatic disease, more poorly differentiated tumors, higher serum prostate-specific antigen (PSA) levels, and/or at younger ages compared with white men. METHODS: A retrospective survey was conducted that identified black and white men with newly diagnosed prostate cancer at the Los Angeles Regional Veterans Affairs Clinics between 1991 and 1997. Patient data were analyzed for racial differences in age at diagnosis, clinical stage, PSA level, and Gleason score of the prostate biopsy specimens. RESULTS: A total of 477 evaluable patients (230 black, 247 white) with newly diagnosed prostate cancer were identified. No significant differences in the average age (66.9 +/- 7.3 versus 67.9 +/- 7.5) or clinical stage at diagnosis were found between black and white men. Among black men, 87% presented with clinically localized disease (T1-2, Nx, M0) compared with 88% of white men. Only 6% of black men presented with distant disease (Tx, Nx, M1) compared with 4% of white men. Black men had higher median PSA levels than white men (14. 2 versus 9.4 ng/mL, P = 0.0001). Black men also had slightly higher average Gleason scores (6.2 versus 5.9, P = 0.025). CONCLUSIONS: This is the first study to show a low and equal percentage of black and white men presenting with metastatic prostate cancer. In this equal access center, no differences were found in patient age or clinical stage of prostate cancer between black and white men at the time of diagnosis. However, black men presented with higher serum PSA values and slightly higher Gleason scores. PMID- 10699617 TI - Does prolonged combined androgen blockade have survival benefits over short-term combined androgen blockade therapy? AB - OBJECTIVES: To explore whether less than 120 days of an antiandrogen plus a luteinizing hormone-releasing hormone agonist resulted in a different survival outcome than 120 days or more of combined treatment in patients with Stage D2 prostate cancer. METHODS: Survival data were available from a previously published controlled trial that had evaluated the efficacy and tolerability of two antiandrogens, bicalutamide and flutamide, each combined with a monthly depot preparation of leuprolide or goserelin, in 813 patients with Stage D2 prostate cancer. Cox's proportional hazards regression model assessed the relative effects of the length of combined androgen blockade (CAB) therapy on survival. This analysis was repeated in the subset of patients who lived at least 2 years beyond the date of randomization. Data were obtained at a median follow-up of 160 weeks. RESULTS: A survival benefit was demonstrated for patients receiving prolonged CAB therapy, with a hazard ratio of 0.275 (95% confidence interval 0.213 to 0.355, P = 0.0001) in favor of patients who received 120 days or more of CAB therapy (median survival 1035 days versus 302 days for less than 120 days of therapy). This result was confirmed in the patients who lived at least 2 years, in whom the median survival time was increased by 35%. The hazard ratio for 120 days or more of CAB therapy versus less than 120 days was 0.415 (95% confidence interval 0.246 to 0.702, P = 0.001). CONCLUSIONS: The results of the present exploratory analysis suggest that prolonged (120 days or more) antiandrogen treatment as part of CAB therapy may result in a better survival outcome. PMID- 10699618 TI - Editorial comment PMID- 10699619 TI - Reply by the authors PMID- 10699620 TI - Endocrine status in elderly men with lower urinary tract symptoms: correlation of age, hormonal status, and lower urinary tract function. The Prostate Study Group of the Austrian Society of Urology. AB - OBJECTIVES: To correlate endocrine parameters in elderly men with lower urinary tract symptoms (LUTS) to patient age and clinical parameters such as prostate volume, prostate-specific antigen (PSA) levels, and uroflowmetry and to compare the clinical and endocrinologic parameters in men with or without hypogonadism. METHODS: Men (40 years old or older) with untreated LUTS as defined by an International Prostate Symptom Score (IPSS) of 7 or greater due to benign prostatic hyperplasia were included in this study and underwent the following investigations: IPSS, free uroflow study, postvoid residual volume, transrectal ultrasound for assessment of prostate volume, serum PSA determination, and an endocrine study, including testosterone, human luteinizing hormone, human follicle-stimulating hormone, prolactin, dehydroepiandrostendione-sulphate (DHEA S), and prolactin. RESULTS: Three hundred twelve men (mean age 62.8 +/- 10.6 years, range 40 to 91) were analyzed. The serum levels of estradiol (correlation coefficient [r] = 0.19), human luteinizing hormone (r = 0.32), human follicle stimulating hormone (r = 0.19), and DHEA-S (r = -0.39) correlated (P <0.05) with age; no such correlation was seen for testosterone (r = 0.04; P0.05) or prolactin (r = 0.09; P0.05). Estradiol (but not testosterone) correlated (r = 0.17, P = 0.01) with prostate volume. The peak flow rate and PSA did not correlate with any endocrinologic parameter. Hypogonadism (serum testosterone less than 3.0 ng/mL) was detected in 22.1% of patients and had no impact on clinical (IPSS, peak flow rate, prostate volume, and PSA level) or endocrine (human luteinizing hormone, human follicle-stimulating hormone, estradiol, prolactin, and DHEA-S) parameters. CONCLUSIONS: A number of age-related endocrine changes are seen in elderly men with LUTS. Hypogonadism is seen in approximately one fifth of elderly men with LUTS, but in our study it had no impact on symptom status, PSA level, prostate volume, uroflowmetry, or endocrine parameters. PMID- 10699621 TI - Diagnosing and treating chronic prostatitis: do urologists use the four-glass test? AB - OBJECTIVES: To examine the diagnosis and treatment of chronic prostatitis, we conducted a national mail survey of practicing urologists in 1998. METHODS: Probability samples were drawn from the American Medical Association Registry of Physicians. RESULTS: Five hundred four urologists responded (response rate 64%). Urologists reported seeing a median of 30 patients (interquartile range 11 to 60) newly diagnosed with chronic prostatitis in the previous 12 months. Eighty percent of urologists responded that they "rarely" (47%) or "never" (33%) performed the Meares-Stamey four-glass diagnostic test. Only 4% answered "almost always." Forty percent of urologists responded that they treat "all" their patients with antibiotics and 42% more responded that they treat "more than half" with antibiotics. Physicians who routinely performed the four-glass test did not differ in antibiotic use from those who used the test less often; however, they were more likely to use treatment other than antibiotics. For example, alpha blockers were used in one half or more of the patients by only 35% of physicians who never do the four-glass test compared with 42% who rarely do the test and 57% who do the test more often (P = 0.002). Results were similar for treatment with natural remedies. CONCLUSIONS: Urologists frequently diagnose chronic prostatitis but rarely perform the four-glass diagnostic test. Use of the four-glass test does not appear to affect urologists' antibiotic treatment patterns. Although bacterial prostatitis is thought to be rare, antibiotic use in the population of men with prostatitis is not. The four-glass test and empiric antibiotics are practices in the diagnosis and treatment of prostatitis that deserve scrutiny. PMID- 10699622 TI - Assessment of a new transurethral balloon dilation catheter in the treatment of urethral stricture disease. AB - OBJECTIVES: To assess a newly designed balloon dilation catheter for the treatment of urethral stricture disease. The dilating capability of the catheter, the tolerability and safety of the procedure, and its short-term efficacy were evaluated. METHODS: Fifty-one patients with urethral strictures underwent dilation with the UrethraMax or a coude tip balloon dilation catheter. Efficacy parameters included measurement of the American Urological Association symptom score and maximum urinary flow rate 3, 6, and 12 months after treatment. The adequacy of dilation and the degree of mucosal trauma and hematuria were assessed endoscopically, and patient pain was measured using a visual analog scale. RESULTS: Forty-three patients (84.3%) were successfully dilated, achieving a urethral caliber of 20F or greater. Dilation resulted in statistically significant improvements in both the mean American Urological Association symptom score and mean maximum urinary flow rate at 3 and 6 months. Mucosal trauma was mild in all but 4 cases, and no patient developed significant hematuria. The mean visual analog pain score was 3.9 (range 0.1 to 9.4). CONCLUSIONS: Balloon dilation is a safe, well-tolerated, office-based procedure that theoretically offers several advantages over sequential rigid dilation and internal urethrotomy. It is associated with minimal complications, and its short-term efficacy is acceptable. We regard this as the dilation procedure of choice and first-line therapy for most strictures. PMID- 10699623 TI - Efficacy and tolerability of tolterodine in children with detrusor hyperreflexia. AB - OBJECTIVES: To investigate the urodynamic effects and tolerability of the new antimuscarinic drug tolterodine in children with detrusor hyperreflexia. METHODS: Twenty-two children (12 boys and 10 girls; age range 3 months to 15 years, mean age 5.7 years) with detrusor hyperreflexia resulting in maximum detrusor pressures exceeding 40 cm H(2)O during filling cystotonometry were enrolled to receive tolterodine tartrate (a total of 0.1 mg/kg orally daily, divided into two doses) either as a first-line therapy (n = 12, group 1) or replacing oxybutynin chloride therapy (n = 10, group 2). Within 3 months, all patients underwent urodynamic re-evaluation during ongoing tolterodine treatment. RESULTS: In group 1, the mean maximum bladder capacity increased from 120.2 to 173.0 mL (+44%), the mean detrusor compliance increased from 8.7 to 13.5 mL/cm H(2)O (+55%), and the mean maximum detrusor pressures decreased from 70.1 to 37.9 cm H(2)O (-46%); the differences were significant (P < 0.001). In group 2, no differences in the urodynamic effects of oxybutynin versus tolterodine were noted. Only 1 patient experienced a transient and moderately adverse effect with tolterodine. CONCLUSIONS: Although based on a limited number of subjects, these data indicate that in pediatric patients with detrusor hyperreflexia, tolterodine may be better tolerated than and equally effective as the standard drug oxybutynin chloride. PMID- 10699624 TI - La Vega slit procedure for the treatment of phimosis. AB - OBJECTIVES: The surgical treatment of phimosis is usually circumcision. In countries in which circumcision is not widely practiced, this approach results in a phallus that is cosmetically unacceptable. We applied a ventral slit procedure to boys with severe phimosis and achieved outstanding results. METHODS: All patients were selected during a 1-week medical mission to La Vega in the Dominican Republic during April 1997. Eight patients presented with severe phimosis. The patient age ranged from 3 to 7 years (mean 4.4). All patients were cleared by the team pediatrician before undergoing the procedure. RESULTS: Eight patients underwent the procedure without complications. The operative time was less than 10 minutes in all instances. All had excellent postoperative cosmesis, were able to retract their foreskins, and voided without difficulty. A follow-up mission to La Vega in March 1998 yielded no complications involving this group of patients. CONCLUSIONS: Unlike circumcision and the dorsal slit procedure, this approach yields a phallus that on initial appearance is indistinguishable from an uncircumcised phallus. The procedure is easily performed and should be considered in the treatment of phimosis whenever foreskin preservation is desired. PMID- 10699625 TI - Transvaginal bone anchored sling. AB - INTRODUCTION: To describe a new technique for the treatment of stress urinary incontinence by the transvaginal creation of a sling anchored to the pubic bone. This technique is minimally invasive and easy to learn, with minimal morbidity and complications. TECHNICAL CONSIDERATIONS: Miniature bone screws with No. 1 polypropylene sutures attached to them and a battery-operated screw inserter are used for the fixation of a biocompatible fabric sling to the pubic bone. The procedure is performed transvaginally with no abdominal or suprapubic incisions. One screw is inserted on each side of the urethra into the pubic bone below the bladder neck. A tunnel is made submucosally between these two holes just below the bladder neck, and the sling is passed through it. Using the sutures on each side of the urethra, the sling edges are tied and pulled toward the pubic bone. The openings made in the vaginal mucosa are closed with absorbable sutures. CONCLUSIONS: This sling procedure is minimally invasive, safe, and effective. Further experience and longer follow-up are necessary to establish its role in the treatment of women with stress urinary incontinence. PMID- 10699626 TI - Continence-preserving anatomic radical retropubic prostatectomy. AB - INTRODUCTION: Urinary continence, especially in regard to the time required to regain urinary control after radical prostatectomy, remains a significant complication of the procedure. TECHNICAL CONSIDERATIONS: The "no touch" or "avoidance" surgical principles that are the basis for continence-preserving radical prostatectomy primarily focus on the preservation of the following components of the external striated urethral sphincteric complex: the entire circumferential musculature of the rhabdosphincter, the fascial investments (the pubourethral ligaments anterolaterally and median fibrous raphe posteriorly), and the innervation of both the rhabdosphincter by way of the intrapelvic branch of the pudendal nerve (somatic) and the mucosal and smooth muscle components by way of the urethral branch of the inferior hypogastric plexus (autonomic). CONCLUSIONS: The clinical impact of preserving the external striated urethral sphincter, its innervation, and its fascial attachments by performing a continence-preserving anatomic retropubic prostatectomy is a shorter time to achieve urinary continence. PMID- 10699633 TI - Laser-assisted demucosalized gastrocystoplasty with autoaugmentation in a canine model. AB - OBJECTIVES: Laser-assisted autoaugmentation gastrocystoplasty has been performed successfully. Experiments were performed to determine the optimal laser for tissue welding during demucosalized autoaugmentation gastrocystoplasty using both a 1.9-microm diode and a 1.32-microm neodymium:yttrium-aluminum-garnet (Nd:YAG) laser with and without thermal control. METHODS: Autoaugmentation gastrocystoplasty was performed on 18 female mongrel dogs. Anastomoses were performed by either suture or laser welding with a 50% human albumin solution. A 1.9-microm diode laser was compared with a 1.32-microm Nd:YAG laser with and without thermal control. In vivo canine bladder capacity measurements were performed both before gastrocystoplasty and at euthanasia. The animals were studied on days 4 and 14. Samples of the anastomotic area from each group were taken to measure tensile strength. Histologic samples were assessed for tissue damage. RESULTS: There was a significant increase in bladder volume in the 4-day group compared with pregastrocystoplasty values. Both the 1.9-microm diode laser and suture control dogs with the 14-day repairs had significantly more tensile strength than their 4-day counterparts. In contrast, no statistical difference was found between the 4 and 14-day 1.32-microm Nd:YAG groups. The suture control group had evidence of minor tissue devitalization at the anastomosis at both 4 and 14 days. The 1.9 and 1.32-microm laser groups both had evidence of tissue devitalization at 4 and 14 days. The 1.32-microm laser group had primarily severe tissue injury. The laser groups at 14 days demonstrated an inflammatory reaction that was localized to the albumin. CONCLUSIONS: Demucosalized gastrocystoplasty with autoaugmentation can be safely and successfully performed with a 1.9-microm diode laser without significant differences in tensile strength when compared with suture controls. The 1.32-microm Nd:YAG laser can also be successfully used; however, the long-term results appear to be inferior to the 1.9-microm diode laser. PMID- 10699634 TI - Thoracoscopic transdiaphragmatic nephrectomy: feasibility study. AB - OBJECTIVES: Large-sized upper pole renal or adrenal tumors are often excised by the open thoracoabdominal approach. As an adjunct to existing transperitoneal and retroperitoneal laparoscopic approaches, we explore a novel minimally invasive technique, the thoracoscopic transdiaphragmatic approach, for performing nephrectomy. METHODS: Thoracoscopic transdiaphragmatic nephrectomy was performed bilaterally in 4 farm pigs (8 kidneys) using three ports placed intercostally. RESULTS: The mean surgical time was 69.3 minutes on the left side and 74.3 minutes on the right. Blood loss was 18.7 mL. The mean size of the diaphragmatic incision was 7.2 cm. Adequate retraction of the spleen and liver was feasible during left and right-sided nephrectomy, respectively. Excellent and expeditious access to the renal hilum was routinely obtained. In 5 of 8 procedures, the diaphragmatic incision was located peripherally along the posterior margin; difficulty in suture repair of the diaphragmatic incision was noted in each instance because of the thin diaphragm in this location. During porcine left nephrectomy with ipsilateral lung collapse (n = 4), arterial blood gases and end tidal carbon dioxide remained normal. CONCLUSIONS: Thoracoscopic transdiaphragmatic nephrectomy is feasible. This technique provides excellent and unique visualization of the renal vessels and the upper pole of the kidney and adrenal gland. When indicated, the thoracoscopic transdiaphragmatic approach, used in combination with current laparoscopic techniques, has the potential to provide the minimally invasive counterpart of the thoracoabdominal surgical approach in select patients with upper pole renal or adrenal pathologic findings. PMID- 10699635 TI - Expression of the human erythrocyte glucose transporter in transitional cell carcinoma of the bladder. AB - OBJECTIVES: It has previously been shown that glucose uptake and use is more prevalent in carcinomas than in normal cells and tissues. We hypothesized that human erythrocyte glucose transporter (Glut-1) expression is increased in bladder transitional cell carcinoma (TCC) and that the grade of expression might correlate with the degree of malignancy. METHODS: Immunostaining of Glut-1 protein was studied in normal bladder (5 cases), benign papilloma (10 cases), superficial tumor (48 cases), and invasive tumor (31 cases) tissue. The immunoreactivity grading system used was as follows: absence of immunoreactivity in tumor cell = 0; less than 10% of the tumor cells immunoreactive = 1+; 10% to 50% of the tumor cells immunoreactive = 2+; and greater than 50% of the tumor cells immunoreactive = 3+. RESULTS: Immunostaining of Glut-1 protein was not expressed in the normal bladder or benign papilloma samples, but it was expressed in 63.0% (46 of 73) of the TCC samples. In the pattern of expression of Glut-1 protein, superficial TCC was stained focally, but invasive TCC was stained diffusely in the tumor nests. The grade of Glut-1 protein expression increased more significantly in the invasive TCC than in the superficial TCC samples (P = 0.002) and more significantly in the high nuclear grade than in the low nuclear grade samples (P = 0.007). In the superficial TCC samples, the bladder tumor recurrence rate did not significantly correlate with Glut-1 protein expression (P = 0.40). CONCLUSIONS: Our results suggest that the Glut-1 protein is not expressed in normal bladder mucosa and benign lesions, that Glut-1 protein expression is strongly associated with neoplastic progression in bladder TCC, and that Glut-1 expression does not correlate with the recurrence rate in superficial bladder TCC. PMID- 10699636 TI - Effects of testosterone on pelvic autonomic pathways: progress and pitfalls. AB - Testosterone has potent effects on reproductive behavior, many of which are due to actions on brain nuclei and spinal motoneurons controlling perineal muscles. The autonomic circuits involved in penile erection, ejaculation and emission, have been less commonly considered as targets for circulating androgens. This review demonstrates that many components of pelvic autonomic reflex pathways, including preganglionic neurons, autonomic ganglion cells and primary afferent neurons, are likely to be influenced by testosterone. The steroid appears to play an important role in maintaining neuronal morphology, transmitter synthesis and receptor expression throughout adulthood. Surprisingly, the effects of testosterone are not limited to neurons involved in reproductive reflexes. The challenge is now to determine the range of neuronal features influenced by androgens, and the mechanisms by which these occur. Studies of androgen receptor location indicate that in many autonomic neurons gene expression may be directly influenced by androgens, but a mismatch between receptor distribution and androgen action shows that in some cells other mechanisms must exist. It is also possible that androgens are metabolised to estrogens by some peripheral neurons. Irrespective of the mechanism, it is time to acknowledge that testosterone is an important "maintenance factor" for autonomic neurons. PMID- 10699637 TI - Different cardiovascular neuron groups in the ventral reticular formation of the rostral medulla in rabbits: single neurone studies. AB - To examine whether the cardiovascular neurons of the ventral medulla consist of functionally different kinds of neurons, single neuronal activity of the ventral medulla, activity of the renal sympathetic nerves (RSNA), blood flow of the ear (EarBF) and arterial pressure (AP) were recorded in urethane-anesthetized, vagotomized and immobilized rabbits during electrical stimulation of the aortic nerve (AN, baroreceptor afferent fibers) and electrical stimulation of the dorsomedial hypothalamus (DMH) that reduced EarBF but less affected on AP and RSNA. The dorsolateral funiculus of the second cervical cord was stimulated to evoke antidromic spikes of medullary neurons. Two kinds of reticulo-spinal neurons were identified. Activities of one kind of neurons were facilitated by stimulation of DMH (latency 48.6+/-27.6 ms, n=11) but they did not respond to stimulation of the AN. Therefore, it was presumed that these neurons controlled vasomotion of the ear through the vasoconstrictor neurons in the spinal cord but did not participate in regulation of systemic AP. Activities of the other neurons were inhibited by stimulation of the AN (latency 47.8+/-8 4 ms, n=16) but they did not respond to the DMH stimulation. These neurons were identical to those reported previously as the RVLM neurons, and they contributed to regulate systemic AP but might not participate in control of cutaneous vascular movement. The former neurons were located medially to the latter in the reticular formation of the rostral ventral medulla. These results provided evidence at the single neuronal level that the cardiovascular neurons in the ventral medulla were consisted of functionally different sympatho-excitatory neurons and they were located at the different sites in the rostral ventral medulla. PMID- 10699638 TI - Involvement of two different mechanisms in trigeminal ganglion-evoked vasodilatation in the cat lower lip: role of experimental conditions. AB - The present study was designed to examine the vasodilator mechanisms elicited by electrical stimulation of trigeminal ganglion (TG) in cat lower lip of the cats. When vago-sympathectomized cats were fixed into a stereotaxic frame by means of ear-bars, etc., the lip blood flow (LBF) increase evoked by lingual nerve (LN) stimulation (parasympathetic reflex response) was almost abolished in 15 out of 34 animals, but unaffected in the other 19. With the animal in the stereotaxic frame, electrical stimulation at sites within the TG evoked an LBF increase whether or not the LN stimulation-induced reflex response was intact. However, hexamethonium abolished the TG stimulation-induced LBF increase in animals whose brainstem parasympathetic reflex was intact, but reduced it by only 50% in animals whose reflex was impaired. This difference was seen in all experiments in which the electrode site was within the TG proper, regardless of its exact position. Although the underlying mechanism is unclear, these data suggest that when the TG is stimulated the LBF increase is entirely mediated via the parasympathetic reflex mechanism in animals whose brainstem reflex is intact, and that an antidromic vasodilatation occurs only in animals whose brainstem parasympathetic reflex is impaired. PMID- 10699639 TI - Electro-acupuncture stimulation to a hindpaw and a hind leg produces different reflex responses in sympathoadrenal medullary function in anesthetized rats. AB - The effects of electro-acupuncture stimulation (EAS) of two different areas of a hindlimb with different stimulus intensities on sympathoadrenal medullary functions were examined in anesthetized artificially ventilated rats. Two needles of 160 microm diameter and about 5 mm apart were inserted about 5 mm deep into a hindpaw (Chungyang, S42) or a hind leg (Tsusanli, S36) and current of various intensities passed to excite various afferent nerve fiber groups at a repetition rate of 20 Hz and pulse duration of 0.5 ms for 30-60 s. Fiber groups of afferent nerves stimulated in a hindlimb were monitored by recording evoked action potentials from the afferents innervating the areas stimulated. The sympathoadrenal medullary functions were monitored by recording adrenal sympathetic efferent nerve activity and secretion rates of catecholamines from the adrenal medulla. EAS of a hindpaw at a stimulus strength sufficient to excite the group III and IV somatic afferent fibers produced reflex increases in both adrenal sympathetic efferent nerve activity and the secretion rate of catecholamines. EAS of a hind leg at a stimulus strength sufficient to excite the group III and IV afferent fibers produced reflex responses of either increases or decreases in sympathoadrenal medullary functions. All responses of adrenal sympathetic efferent nerve activity were lost after cutting the afferent nerves ipsilateral to the stimulated areas, indicating that the responses are the reflexes whose afferents nerve pathway is composed of hindlimb somatic nerves. It is concluded that electro-acupuncture stimulation of a hindpaw causes an excitatory reflex, while that of a hind leg causes either excitatory or inhibitory reflex of sympathoadrenal medullary functions, even if both group III and IV somatic afferent fibers are stimulated. PMID- 10699640 TI - Differential activation of nitric oxide synthase through muscarinic acetylcholine receptors in rat salivary glands. AB - Muscarinic receptors play an important role in secretory and vasodilator responses in rat salivary glands. Nitric oxide synthase (NOS) appears to be one of the multiple effectors coupled to muscarinic receptors in both submandibular and sublingual glands although some differences have been found depending on the gland studied. First, submandibular glands had a lower basal activity of nitric oxide synthase than sublingual glands and the concentration-response curve for carbachol was bell-shaped in the former but not in sublingual glands. Second, cGMP levels displayed a similar profile to that observed for NOS activity in both glands. Third, protein kinase C also coupled to muscarinic receptor activation in the glands might have a regulatory effect on nitric oxide production since its activity was higher in basal conditions in submandibular than sublingual glands and it also increased in the presence of the agonist at a concentration that inhibited NOS activity in submandibular glands. The effects appear to be partly related to the expression of a minor population of M(1) receptors in submandibular glands absent in sublingual as determined in binding and signaling experiments with the muscarinic receptor antagonist pirenzepine. PMID- 10699641 TI - Enhanced negative chronotropy by inhibitory receptors in transgenic heart overexpressing beta(2)-adrenoceptors. AB - Transgenic (TG) mice overexpressing beta(2)-adrenoceptors (AR) in the heart have enhanced beta-adrenergic activity. Since the degree of beta-adrenergic activation influences the negative chronotropic control of heart rate (HR), we studied the inhibitory effect of cholinergic and purinergic stimulation on HR in TG and wild type (WT) control mice. Bradycardia in response to vagal nerve stimulation and administration of acetylcholine or adenosine was studied in anesthetised animals and perfused hearts. Basal HR was significantly higher in TG than WT mice (P<0.01). Electrical stimulation of vagal nerves (1-32 Hz) induced a Hz-dependent reduction in HR and the response was more pronounced in TG than WT groups (P<0.01). In perfused hearts, HR reduction by acetylcholine (ACh) was more pronounced with EC(50) 110-fold lower in TG than WT hearts. Adenosine-induced bradycardia, which was abolished by a P(1) antagonist, was more pronounced in TG hearts. After pre-treatment with pertussis toxin (PT, 100 microg/kg), bradycardia by vagal nerve stimulation or ACh remained unchanged in WT, but markedly inhibited in TG hearts (both P<0.01). Conversely, inhibiting guanylyl cyclase with LY83583 (30 microM) or nitric oxide synthase with L-NMMA (100 microM) attenuated HR reduction by vagal nerve stimulation in WT but not in TG hearts. Immunobloting assay showed similar G(ialpha2) abundance in TG and WT hearts. Thus, cardiac overexpression of beta(2)AR with high beta-adrenergic activity leads to hypersensitivity of inhibitory receptors controlling HR due to increase in activity of PT-sensitive G-proteins. PMID- 10699642 TI - Connections of Barrington's nucleus to the sympathetic nervous system in rats. AB - Barrington's nucleus (BN) has been considered a pontine center related exclusively to the control of pelvic parasympathetic activity. The present study demonstrates an anatomical linkage between BN and autonomic outflow to visceral targets innervated exclusively by the sympathetic division of the autonomic nervous system. Temporal analysis of infection after injection of pseudorabies virus (PRV), a retrograde transynaptic tracer, into two sympathetically innervated organs, the spleen and the kidney, revealed the presence of infected neurons in BN at early post-inoculation survival intervals. Immunohistochemical localization of PRV after spleen injections showed that a small subpopulation of BN neurons became labeled in a time frame coincident with the appearance of infected neurons in other brain regions known to project to sympathetic preganglionic neurons (SPNs) in the thoracic spinal cord; a larger number of infected neurons appeared in BN at intermediate intervals after PRV injections into the spleen or kidney. Coinjection of the retrograde tracer Fluoro-Gold i.p. and PRV into the spleen demonstrated that parasympathetic preganglionic neurons in the caudal medulla or lumbo-sacral spinal cord were not infected, indicating that infected BN neurons were not infected via a parasympathetic route. Thus, BN neurons become infected after PRV injections into the spleen or kidney either directly through BN projections to SPNs, or secondarily via BN projections to infected pre-preganglionic neurons. These results demonstrate an anatomical linkage, either direct or indirect, between BN and sympathetic activity. Because BN receives numerous inputs from diverse brain regions, the relation of BN with both branches of the autonomic nervous system suggests that this nucleus might play a role in the integration of supraspinal inputs relevant to the central coordination of sympathetic and parasympathetic activity. PMID- 10699643 TI - Trigeminally induced cardiovascular reflex responses in spinalized rats. AB - The effects on cardiovascular functions of noxious stimulation to the orofacial areas innervated by trigeminal afferent nerves were analyzed in urethane anesthetized, spinal cord-intact rats and in rats acutely spinalized at the second cervical level. In the spinal cord-intact rats, pinching of the upper lip produced increases in both heart rate (HR) and mean arterial pressure (MAP). Both responses were considered to be due to activation of sympathetic efferent nerves to the cardiovascular organs. Both responses were attenuated but did not disappear after spinalization at the C2 level. In spinalized rats, sympathetic preganglionic neurons emerging from the thoracolumbar spinal cord could not receive any neural influences from the brain. The HR response in the spinal rats was abolished after either bilateral vagotomy or intravenous injection of a peripherally acting muscarinic cholinergic receptor antagonist, methylatropine. This suggests that the increase in HR was elicited via vagal cholinergic efferent fibers, probably by decreasing tonic activity of vagus nerves to the heart. In spinal rats, neither vagotomy nor cholinergic blockade affected the increase in MAP, but i.v. injection of the vasopressin V1 receptor antagonist, OPC-21268, abolished the response of MAP. This suggests that the response of MAP was due to peripheral vasoconstriction elicited by vasopressin secreted from the posterior pituitary lobe. The present study demonstrated that, in rats acutely spinalized at the C2 level, noxious stimulation of orofacial areas innervated by the trigeminal nerve could produce reflex increases both in HR, by decreasing cholinergic vagal nerve activity to the heart, and blood pressure, by secreting vasopressin from the pituitary gland, even though sympathetic efferent innervation to the cardiovascular organs could not be directly affected by trigeminal afferent nerve excitation. PMID- 10699644 TI - Distinct localization and target specificity of galanin-immunoreactive sympathetic preganglionic neurons of a teleost, the filefish Stephanolepis cirrhifer. AB - Immunoreactivity for galanin was examined in the sympathetic preganglionic neurons in the spinal cord, adrenal glands, sympathetic ganglia, and some sensory ganglia of the filefish Stephanolepis cirrhifer. Galanin-immunoreactive neurons were found only in the rostral part, but not in the caudal part of the central autonomic nucleus (a column of sympathetic preganglionic neurons of teleosts). Many galanin-immunoreactive nerve terminals were found in contact with neurons in the celiac ganglia and the cranial sympathetic ganglia on both sides of the body. Most neurons encircled by galanin-immunoreactive nerve fibers were negative for tyrosine hydroxylase. Galanin-immunoreactive nerve fibers were very sparse in the spinal sympathetic paravertebral ganglia. No galanin-immunoreactive nerve fibers were found in the adrenal glands. No sensory neurons of the trigeminal, vagal, or spinal dorsal root ganglia were positive for galanin-immunoreactivity. These results suggest that galanin-immunoreactive sympathetic preganglionic neurons have distinct segmental localization and might project specifically to a population of non-adrenergic sympathetic postganglionic neurons in the celiac and cranial sympathetic ganglia. PMID- 10699645 TI - Observer variations in short period spectral analysis of heart rate variability. AB - 25% of the corresponding mean LF/HF ratio. The smallest interobserver variations were found for the 'fixed frequency' method. The data showed that it is advantageous to select the 3-min ECG periods but not to select the frequency regions. Selection of the latter led to an increase in interobserver variation. The results of this study give a realistic impression of the intrasubject and interobserver variation to be expected when measuring the LF/HF ratio. This variation is considerable. PMID- 10699646 TI - Graded vascular autonomic control versus discontinuous cardiac control during gradual upright tilt. AB - Indexes of heart rate variability (HRV) and the slope of cardiac baroreflex are extensively used for non invasive assessment of circulatory autonomic control in pathophysiology. We performed this study (1) to assess the sensitivity of these indexes towards small graded postural stimulations and (2) to delineate the informations provided about the settings of both vascular tone and cardiac activity. Twenty healthy subjects were randomly tilted for eight minutes at each of the six angles: -10 degrees, 0 degrees (supine), 10 degrees, 30 degrees, 45 degrees, and 60 degrees. Instant RR-interval and finger blood pressure (BP) were continuously recorded, and venous blood was collected at the end of each 8 min position for catecholamines determination. Group average heart rate, noradrenaline and diastolic BP (DBP) increased linearly with head-up tilt angle from 10 degrees. Systolic BP (SBB) ranked only two distinct series -10 degrees, 0 degrees, 10 degrees versus 30 degrees, 45 degrees, 60 degrees, as did the number of spontaneous baroreflex (SBR) sequences. The spectral power of the low frequency (LF) and high-frequency (HF) of RR variability and the ratio LF/HF changed rather abruptly from either 30 degrees or 45 degrees, depending on each individual. Both HF/tot i.e. the ratio of HF to total spectral RR variability and the slope of SBR decreased markedly from 10 degrees to 30 degrees and less but more gradually from 30 degrees to 60 degrees. Thus, our observations argue for gradual adjustments of vascular tone as reflected by highly consistent changes in plasma noradrenaline and diastolic arterial pressure, contrasting with a main discontinuous autonomic setting of cardiac activity as reflected by changes in the harmonic components of spectral RR variability and in the slope of cardiac baroreflex. The pattern of changes in systolic arterial pressure attested the discontinuous cardiac autonomic control rather than the gradual setting of arterial tone. We submit that these different patterns of autonomic adjustments should be considered when assessing pathophysiological states. PMID- 10699647 TI - Effects of three days of dry immersion on muscle sympathetic nerve activity and arterial blood pressure in humans. AB - The present study was performed to determine how sympathetic function is altered by simulated microgravity, dry immersion for 3 days, and to elucidate the mechanism of post-spaceflight orthostatic intolerance in humans. Six healthy men aged 21-36 years old participated in the study. Before and after the dry immersion, subjects performed head-up tilt (HUT) test to 30 degrees and 60 degrees (5 min each) with recordings of muscle sympathetic nerve activity (MSNA, by microneurography), electrocardiogram, and arterial blood pressure (Finapres). Resting MSNA was increased after dry immersion from 23.7+/-3.2 to 40.9+/-3.0 bursts/min (p<0.005) without significant changes in resting heart rate (HR). MSNA responsiveness to orthostasis showed no significant difference but HR response was significantly augmented after dry immersion (p<0. 005). A significant diastolic blood pressure fall at 5th min of 60 degrees HUT was observed in five orthostatic tolerant subjects despite enough MSNA discharge after dry immersion. A subject suffered from presyncope at 2 min after 60 degrees HUT. He showed gradual blood pressure fall 10 s after 60 degrees HUT with initially well maintained MSNA response and then with a gradually attenuated MSNA, followed by a sudden MSNA withdrawal and abrupt blood pressure drop. In conclusion, dry immersion increased MSNA without changing MSNA response to orthostasis, and resting HR, while increasing the HR response to orthostasis. Analyses of MSNA and blood pressure changes in orthostatic tolerant subjects and a subject with presyncope suggested that not only insufficient vasoconstriction to sympathetic stimuli, but also a central mechanism to induce a sympathetic withdrawal might play a role in the development of orthostatic intolerance after microgravity exposure. PMID- 10699648 TI - Cellular expression of the neurokinin 1 receptor in the human antrum. AB - The localization of the neurokinin 1 receptor in rat and guinea pig gastrointestinal tract has been extensively studied but not in human tissues. The present study used antibodies to characterize the cellular expression of neurokinin 1 receptors in human antrum. Cryostat sections (40-80 microm) were immunostained for the neurokinin 1 receptor double labeled with substance P, von Willebrand's factor, c-kit, fibronectin, S-100, serotonin, gastrin and somatostatin. Neurokinin 1 receptor-immunoreactivity was observed on neurons within the myenteric and submucosal plexuses surrounded by substance P immunoreactive fibers and on von Willebrand's factor-immunoreactive endothelial cells lining blood vessels throughout the antral wall. c-Kit-immunoreactive interstitial cells of Cajal and gastrin cells were co-stained by the monoclonal neurokinin 1 receptor antibody. Finally, there was no evidence for the presence of the neurokinin 1 receptor on fibroblasts, Schwann, somatostatin, serotonin or smooth muscle cells. This study clearly demonstrates an expanded cellular expression of the neurokinin 1 receptor in the human antrum. PMID- 10699649 TI - Improvement of urge- and mixed-type incontinence after acupuncture treatment among elderly women - a pilot study. AB - The aim of this study was to investigate if sensory stimulation in the form of manual acupuncture could influence urge- or mixed-type incontinence among elderly women who were not satisfactorily relieved by standard pharmacological and non pharmacological treatments given at a specialised incontinence unit. The study is an open clinical follow-up study. The study included 15 elderly women who were treated with manual acupuncture 12 times. Both subjective scorings and objective measurements in the form of leakage in grams (48 h Inco-test) were used. Evaluations were performed at discharge and 1 and 3 months thereafter. Almost all outcome measurements were significantly improved even at follow-up 3 months after the last treatment. Global scorings showed that 12 of the 15 women considered themselves improved even at the follow-up 3 month after treatments were completed. The possible mechanisms of action are discussed, as is the way to perform more studies in this field. PMID- 10699650 TI - The human nucleus of the solitary tract: visceral pathways revealed with an "in vitro" postmortem tracing method. AB - Visceral relay neurons in the nucleus of the solitary tract (NTS) regulate behavior and autonomic reflex functions. NTS projections have been extensively characterized in animal studies but not in humans. For the first time, NTS fiber trajectories in the human medulla oblongata were revealed with an "in vitro" postmortem tracing method. Local intramedullary pathways were labeled by direct pressure injections of free horseradish peroxidase centered on the medial subnucleus at a level adjacent to true obex. Labeled elements were resolved by peroxidase histochemistry as a dark brown intracellular reaction product. A prominent transtegmental system of axons emerged from the NTS injection sites and entered the intermediate reticular zone, a region corresponding to an autonomic reflex center in other mammals. A medial system of axons arched across the dorsomedial reticular formation toward the dorsal medullary raphe and projected ventrally toward the nucleus gigantocellularis. A small lateral fiber trajectory coursed towards the dorsomedial zone of spinal trigeminal nucleus caudalis. Presumptive terminals appeared as dustings of fine punctate processes within the NTS, dorsomotor nucleus and reticular formation. NTS projections in humans resemble those identified in other mammals including primates. Axonal tracing studies predict that visceral impulses in humans may transmit over evolutionarily conserved pathways involved in autonomic feedback control and stress adaptation. PMID- 10699651 TI - Are alpha-blockers involved in lower urinary tract dysfunction in multiple system atrophy? A comparison of prazosin and moxisylyte. AB - Lower urinary tract dysfunction is a major cause of morbidity in patients with multiple system atrophy (MSA). alpha1-Adrenergic receptors are present in the proximal urethra where impaired relaxation may be responsible for voiding difficulty and a large amount of residual urine. An open study was designed to evaluate whether the blockade of these receptors by prazosin (a nonselective alpha1 blocker) and moxisylyte (an alpha1A-selective blocker) would improve bladder emptying in patients with MSA. Post-micturition residual volumes and clinical symptoms of 49 patients with MSA were evaluated at trial entry and after 4 weeks (prazosin; n=21 and moxisylyte; n=28). The respective means for the prazosin and moxisylyte groups were 38.1% and 35.2% reductions in residual urine volume (P<0.05), and there was lessening of urinary symptoms. Side effects due to orthostatic hypotension were seen in 23.8% of the prazosin group but in only 10.7% of the moxisylyte group. These effects were common in patients with postural hypotension of more than -30 mmHg at trial entry (P<0.05). Modulation of alpha1-receptors may function in the management of lower urinary tract dysfunction in MSA. PMID- 10699652 TI - Effects of acute and chronic maprotiline administration on inhibitory avoidance in male mice. AB - The effects of acute and chronic administration of maprotiline (5, 10 or 20 mg/kg, intraperitoneally) were assessed on inhibitory avoidance in male mice. Acute administration of maprotiline before training did not effect training phase latencies, but impaired performance (i.e. produced shorter latencies) in the test at doses of 5 and 20 mg/kg. When given after training, the drug did not modify test latencies at any of the doses used. Chronic administration for 21 days (interrupted 24 h before training) also shortened latencies in the test but not in training. An experiment on the acute effects of maprotiline on analgesia (determination of flinch and jump thresholds for increasing electric foot shock levels), at the doses stated, was carried out on naive animals. No analgesic effect of the drug was found. Taken together, the results indicate that acute maprotiline produces anterograde amnesia, and tolerance does not appear after 21 days of treatment. PMID- 10699653 TI - Quinpirole- and amphetamine-induced hyperdipsia: influence of fluid palatability and behavioral cost. AB - Daily administration of moderate doses of amphetamine or of the dopaminergic D2 agonist quinpirole is associated with the development of excessive, non regulatory drinking. Here we compared the influence of manipulating fluid palatability and behavioral cost on the development of this drinking augmentation. Experiment 1 was based on the phenomenon of contrafreeloading (CFL): animals work for a resource even though the same resource is freely available. The effects of 15 daily injections of amphetamine (1.0 and 1.7 mg/kg i.p. ) or quinpirole (0.1 and 0.56 mg/kg i.p.) were evaluated in mildly water deprived rats. For the first 6 days the rats obtained water by lever pressing (FR3) only; over the following 9 days water was also freely available (CFL). Initially, 0.56 mg/kg quinpirole reduced lever pressing for water. A complete recover of responding was then obtained, and was followed by a progressive increment in the amount water obtained by lever pressing during the CFL phase (from 10 to 50%). Amphetamine did not affect percent CFL, but at the highest dose (1.7 mg/kg) reduced total water intake during the last 3 days of treatment. In experiment 2 the rats had free access to two bottles, one of which contained tap water, and the other contained either an ethanol (6%) or a sucrose (5%) solution. After habituation to this regimen, the rats received 10 daily i.p. injections of vehicle, amphetamine (1.0 or 3 mg/kg), or quinpirole (0.1 or 0.56 mg/kg). Quinpirole 0.56 mg/kg enhanced daily fluid intake under both sucrose and ethanol conditions, but selectively reduced ethanol preference. The higher amphetamine dose reduced fluid intake and sucrose preference. In conclusion, chronic exposure to a dopaminergic D2 agonist, but not to amphetamine, produced an increment of drinking that was resistant to manipulation of either palatability or the behavioral cost of the fluid. PMID- 10699654 TI - Prior electrical stimulation of the inferior colliculus sensitizes rats to the stress of the elevated plus-maze test. AB - Besides its primary function in the transmission of acoustic signals, the inferior colliculus (IC) is involved in conveying auditory information of aversive nature to higher cortical structures. In the field of anxiety research, one widely used animal model is the electrical stimulation of a given structure supposed to be involved in the neural circuitry underlying emotional behavior. Indeed, electrical stimulation of the inferior colliculus produces fear-like responses. Moreover, prior exposure to stressful events sensitizes rodents' responsivity to fearful stimuli when they are subsequently tested in the elevated plus-maze. Based on these evidence it seems to be important to know whether animals stimulated in the inferior colliculus would show a heightened behavioral responsivity to subsequent stressful events. To this end, we examined the temporal course of the effects of the electrical stimulation of this midbrain region (5, 10 and 15 min afterward) on the conventional and ethological measures of the behavior of rats exposed to the elevated plus-maze test. Prior electrical stimulation of the inferior colliculus produced an 'anxiogenic' profile in the elevated plus-maze, i.e. a significant reduction in the number of entries and time spent into the open arms. Still, previous electrical stimulation of the inferior colliculus caused a significant decrease in rearing, scanning, peeping out, head dipping and end-arm activity while increased immobility. All these changes occurred after 5 min of inferior colliculus electrical stimulation. Therefore, stimulation of the inferior colliculus causes a heightened responsivity to anxiogenic stimuli inherent to the elevated plus-maze test. These findings bring additional support to the proposed role of this midbrain structure in the elaboration of adaptive responses to stressful situations. PMID- 10699655 TI - Evaluation of the spontaneously hypertensive rat as a model of attention deficit hyperactivity disorder: acquisition and performance of the DRL-60s test. AB - CD. However, the pattern of responding at DRL-60s suggested poor schedule control for the WKY rats. Therefore, the performance of SHR in the DRL test does not appear to represent a valid model of ADHD. Further, our findings with the WKY rat suggest that this strain is a poor behavioural control for the SHR. PMID- 10699656 TI - Impaired acquisition of a Morris water maze task following selective destruction of cerebellar purkinje cells with OX7-saporin. AB - Spatial learning in the Morris water maze task is believed to be dependent on an intact hippocampal system. However, evidence from human studies and animal experiments suggests a potential cerebellar involvement in spatial processing, place learning, and other types of 'higher-order' cognition. In order to investigate this possibility, intraventricular injections (ICV) of the anti neuronal immunotoxin OX7-saporin were used to selectively destroy cerebellar Purkinje cells, without affecting other brain areas believed to be critically involved in spatial learning and memory. Bilateral ICV injections of 2 microg OX7 saporin (4 microg total) in adult male rats produced substantial loss of Purkinje cells (56%) throughout the cerebellum without affecting hippocampal morphology or biochemical indices of cholinergic, serotonergic, or catecholaminergic function in the hippocampus, frontal cortex, or striatum. ICV OX7-saporin significantly impaired acquisition and performance of the standard Morris water maze task (though the impairment was less severe than reported in earlier studies that used alternate lesion methods or mutant mice species), but did not alter performance on the cued version of the task, or locomotor activity. In addition, lesioned animals spent significantly less time in the target quadrant on probe trial days 4 and 7 and the average distance to target scores (ADT) were significantly greater than controls on those days. Swim speed was not affected. Based on the specificity of the behavioral and neurobiological alterations, these data support the hypothesis that the cerebellum is involved in spatial processing and place learning. PMID- 10699657 TI - Skilled forelimb reaching for pasta guided by tactile input in the rat as measured by accuracy, spatial adjustments, and force. AB - Rats are capable of reaching for food with a single forelimb, but since they locate the target of their reach using olfaction, it is unclear how they adjust their limb movement to compensate for errors. Although it is thought that their reaching movement is ballistic and can only be adjusted by trial and error, whether they can use haptic cues to aid in locating and identifying a target has not been examined. The present study addressed this question by allowing rats to reach through a slot for rigidly held pieces of uncooked pasta of varying thickness, which could be oriented vertically or horizontally from different points around the slot and which were attached to a force transducer. The tasks required that animals not only adjust their reach and grasp to the target's location but also identify the target based on its texture. Acquisition curves were made of head orientation, limb transport trajectories, number of attempts per success, paw orientation, breaking direction and force of the grasp. A haptic discrimination test used pasta and similar sized metal rods with different tactile properties as discriminanda. The results indicated that whereas postural orientation and limb transport trajectory were not modified as a function of target orientation, paw orientation and grasp force did vary as a function of the sensory qualities of the target object, and the rats could make a haptic discriminative choice of a target object. The results show that the rat is capable of adjusting paw movements using haptic information, suggesting that somatosensory features of sensorimotor control of limb and paw movements in carnivores and primates are shared by rodents. This commonality points to a conservation of motor control in mammals, explains some of the idiosyncratic features of rat reaching behavior, and confirms that rodents provide a good model for investigating sensorimotor functions. PMID- 10699658 TI - Similar effects of amphetamine and methylphenidate on the performance of complex operant tasks in rats. AB - Methylphenidate and D-amphetamine are central nervous system stimulants that have been suggested to share certain behavioral and neurochemical effects. The current study was undertaken to determine whether methylphenidate and D-amphetamine have similar effects on the performance of a battery of complex operant tasks in rats. Thus, the effects of amphetamine (0.1-6.0 mg/kg, i.p.) and methylphenidate (1.12 18.0 mg/kg, i.p) on the performance of rats in three complex food-reinforced operant tasks were examined. The tasks (and the brain functions they are intended to model) included: (1) conditioned position responding (auditory/visual/position discrimination); (2) incremental repeated acquisition (learning); and (3) temporal response differentiation (time estimation). In addition, each of these tasks was paired with a progressive ratio task to assess drug effects on the rats' motivation to lever press for the food reinforcers used. Consistent with their effects in other behavioral paradigms, methylphenidate and D-amphetamine produced very similar patterns of disruption of the four tasks. Drug-induced changes in the endpoints of the progressive ratio task generally paralleled changes in the other three tasks, suggesting a major role for appetitive motivation in the effects of these agents. Several effects of these agents seen in the current study are consistent with their effects in children with attention deficit-hyperactivity disorder. These data further validate the use of this battery of operant tasks for the characterization of pharmacological agents, and suggest that findings using these tasks may be predictive of what is seen in humans. PMID- 10699659 TI - A comparison of the effects of hippocampal or prefrontal cortical lesions on three versions of delayed non-matching-to-sample based on positional or spatial cues. AB - We trained rats to perform one of three versions of delayed non-matching-to sample (DNMS): DNMS between two retractable levers in an enclosed operant chamber; varying-choice DNMS between two arms selected at random on every trial in an uncovered eight-arm radial arm maze; or recurring-choice DNMS between the same two arms on every trial in a covered radial maze (N=33/task). Rats with medial prefrontal cortical lesions showed delay-independent impairments on the retractable lever and recurring-choice tasks, but performed varying-choice DNMS normally. Rats with hippocampal lesions exhibited delay-independent impairments of the retractable lever task and delay-dependent impairments of both radial maze tasks. When rats trained initially to perform recurring choice DNMS were switched to varying choice DNMS, the impairments of both the prefrontal and hippocampal groups were reduced, although hippocampal animals remained significantly impaired. When rats trained initially to perform varying choice DNMS were switched to recurring choice DNMS, the impairment of the hippocampal group was exacerbated while the prefrontal group remained unimpaired. Thus training the prefrontal group to perform the varying choice task first seemed to protect from impairment when these rats were subsequently trained to perform recurring choice DNMS. This protection provides evidence against the possibility that factors related to proactive interference or to temporal discrimination can account for the effects of prefrontal lesions on delayed conditional discriminations involving two response alternatives in fixed locations. PMID- 10699660 TI - Ameliorative effect of tacrine on spatial memory deficit in chronic two-vessel occluded rats is reversible and mediated by muscarinic M1 receptor stimulation. AB - Our previous study demonstrated that permanent two-vessel occlusion (2VO)-induced working memory deficit was improved by daily administration of tacrine, a cholinesterase inhibitor. In this study, we investigated the mechanism underlying the effects of tacrine in 2VO rats using the eight-arm radial maze task. Daily administration of tacrine (0.1 or 0.3 mg/kg i.p.) started 5 weeks after the 2VO operation significantly improved the maze performance. In the delay-interposition task, a significant impairment of maze performance was observed in the tacrine (0.3 mg/kg, i.p.)-treated rats at a delay of 90 min but not delays of 5 or 30 min. Sham-operated rats were not affected by delay. After leaving animals with no further treatment for 4 weeks, the tacrine-pretreated 2VO rats showed significantly impaired performance compared to the sham-operated control animals. However, the performance of the tacrine-pretreated 2VO rats was significantly improved by restarting the daily administration of tacrine (0.3 mg/kg, i.p.). The effect of tacrine was reversed by the muscarinic antagonist scopolamine and the selective M1 antagonist pirenzepine. Moreover, a microdialysis study revealed that tacrine (1 or 3 mg/kg, i.p.) increased the extracellular acetylcholine (ACh) level for a period of over 3 h in the cerebral cortex of 2VO rats. These findings suggest that the ameliorative effect of tacrine on the spatial memory deficit in 2VO rats is reversible and may be mediated by stimulating the muscarinic M1 receptor via elevation of the extracellular ACh level in the brain. PMID- 10699661 TI - Reversal learning deficit in a spatial task but not in a cued one after telencephalic ablation in goldfish. AB - The fish telencephalon seems to be involved in spatial learning and memory in a similar manner to the hippocampus of the land vertebrates. For instance, telencephalon ablated goldfish are impaired in the post-operative retention of a 'spatial constancy' task, which requires the use of mapping strategies, but not in a directly cued task in which responses are based in a guidance strategy. In this regard, previous experiments showed that intact goldfish trained in the spatial constancy task presented considerable behavioral flexibility, as they showed fast reversal learning, that is, they required less training compared with animals trained in the directly cued task and made a lower number of errors to master the reversal than in acquisition. The purpose of the present work was to investigate if the goldfish telencephalon is involved in the faster reversal learning of the animals trained in the spatial constancy task. Goldfish with bilateral telencephalic ablation, sham operated or intact, were trained in the spatial constancy task or in the directly cued task. Telencephalic ablation selectively impaired reversal learning in the animals trained in the spatial constancy procedure. Ablated animals in this procedure reversed more slowly than control animals. By contrast, telencephalic ablation did not produce any significant deficit during reversal in the animals trained in the directly cued task. These results provide additional evidence that the fish telencephalon, as the land vertebrate hippocampus, plays a crucial role in the use of flexible spatial representations. PMID- 10699662 TI - The effects of local application of D2 selective dopaminergic drugs into the medial prefrontal cortex of rats in a delayed spatial choice task. AB - The study examined the effects of modulation of dopamine D2 receptors-mediated neurotransmission in the rat's prefrontal cortex (PFC) on storage and executive components of working memory. Rats were trained on delayed (delay interval, 3 s) and non-delayed choice in a U-maze. The prominence of proactive interference was evaluated by sorting errors in a current trial on the basis of animal reactions in a preceding trial. The erroneous runs to the same arm of the maze as in the previous trial were identified as the repetitions (RE) and the erroneous runs to the other arm in comparison with the previous trial were classified as alternations (AE). The bilateral microinfusion of D2 agonists PPHT (0.004 microg, 0.04 microg, 0.4 microg/1 microl) into medial wall of the PFC produced a dose dependent increase in the error rate of the delayed-response task and did not influence non-delayed choice. In delay condition PPHT enhanced the perseverative tendencies (the rate of RE was significantly higher than the rate of AE), in non delayed choice the erroneous performance was mainly represented by AE. In contrast, the infusion of D2-receptor antagonist sulpiride (0.03 microg, 0.3 microg, 3 microg/1 microl) increased the accuracy of delayed choice and changed the mode of intertrial dependence-rats made significantly more AE than RE. The results are discussed in terms of the involvement of D2 receptor dependent transmission of the PFC in different cognitive processes related to the delayed performance in U-maze (within-trial short-term storage of information versus dynamic control of between-trials working memory processing). PMID- 10699663 TI - Functional role of rat prelimbic-infralimbic cortices in spatial memory: evidence for their involvement in attention and behavioural flexibility. AB - The involvement of the medial prefrontal cortex (mPFC), and more particularly the prelimbic and infralimbic cortices (PL-IL area), in spatial memory remains controversial. The present study investigates the effects of neurotoxic lesions restricted to the PL-IL area of the mPFC in rats trained in two different spatial tasks. In experiment 1, PL-IL lesioned rats showed normal acquisition of a delayed non-matching to position task. They were also able to plan their responses for a prospective strategy but were transiently disrupted when the initial delay was extended. In experiment 2, rats were trained to locate one baited box among 13 identical boxes distributed on a circular arena. Lesioned rats performed normally when trained from a single start position but were severely disrupted when four start positions were used. A probe trial showed this deficit was not due to failure to learn the goal location. The addition of a proximal cue signalling the goal box helped lesioned rats to directly open the goal box, but did not compensate for greater distances that they travelled to reach it. Results from both experiments indicate that the PL-IL area is directly involved neither in allocentric spatial representations nor prospective memory and is not specifically involved in working memory. This area seems more likely to be involved in both attentional processes and behavioural flexibility that may be important for processing information for working memory as well as for spatial memory. PMID- 10699664 TI - Telencephalic nuclei control late but not early nestling calls in the budgerigar. AB - Bilateral lesions targeting the central nucleus of the anterior archistriatum (AAc) were placed in nestling budgerigars (Melopsittacus undulatus) aged 5, 9, 13, 22, 26, and 33 days post-hatch in order to evaluate the role of the telencephalon in producing nestling vocalizations in this species. In budgerigars, AAc is the final common pathway from telencephalic vocal control nuclei to brainstem respiratory and syringeal motorneuron pools. The results show that lesions destroying AAc bilaterally in addition to surrounding archistriatum and neostriatum do not alter the production of early simple patterned foodbegging calls but do prevent both the normal transition at 3-4 weeks post-hatch to more complex begging calls as well as the emergence of individually-distinctive contact calls around the time of fledging. These vocal results are strikingly similar to those obtained in previous studies in which early deafening of nestlings (Heaton and Brauth, 1999) and early lesioning of auditory areas in the anterior telencephalon (Hall WS, Brauth SE, Heaton JT. Comparison of the effects of lesions in nucleus basalis and field 'L' on vocal control learning and performance in the budgerigar (M. undulatus), Brain Behav. Evol., 1994;44:133 148) did not affect call production until 3-4 weeks post-hatch. These data combined support the idea that neither auditory feedback nor telencephalic sensorimotor circuits are necessary for the production of nestling calls before 3 weeks post-hatch. PMID- 10699665 TI - Social memory is impaired in neonatally ibotenic acid lesioned rats. AB - Postnatal lesion with ibotenic acid in the ventral hippocampus produces several behavioural effects that resemble the symptoms of schizophrenia. In the present study, we tested neonatally lesioned 1-year-old Sprague-Dawley rats for their social memory. It was found that social memory is worsened in lesioned rats. Subchronic treatment with haloperidol (0.025 mg/kg body weight) partly ameliorated this impairment. It was suggested that social memory might be a useful paradigm to test clinically used and potential antipsychotic drugs for their effects on learning and memory processes. PMID- 10699667 TI - Competitive PCR-DGGE analysis of bacterial mixtures: an internal standard and an appraisal of template enumeration accuracy. AB - Analysis of polymerase chain reaction (PCR) amplified 16S rDNA fragments from environmental samples by denaturing gradients of chemicals or heat [denaturing gradient gel electrophoresis (DGGE) and thermal gradient gel electrophoresis (TGGE)] within polyacrylamide gels is a popular tool in microbial ecology. Difficulties in acceptance of the technique and interpretation of the results remain, due to its qualitative nature. In this study we have addressed this problem by the construction and evaluation of a quantitative standard for incorporation into test DNA samples. The standard was based on a naturally occurring 16S rRNA gene carried by the X-endosymbiont of the psyllid Anomoneura mori, a gamma-proteobacterium. This sequence is the most AT-rich 16S rDNA gene recovered from any cultured organism or environmental sample described to date, and a specifically amplified rDNA fragment denatured under exceptionally low stringency denaturing conditions. The native sequence was modified to incorporate perfect matches to the PCR primers used. The efficiency of amplification of this standard in comparison to a range of 16S rDNA sequences and the errors involved in enumerating template molecules under a range of PCR conditions are demonstrated and quantified. Tests indicated that highly accurate counts of released target molecules from a range of bacterial cells could be achieved in both laboratory mixtures and compost. PMID- 10699668 TI - Widefield deconvolution epifluorescence microscopy combined with fluorescence in situ hybridization reveals the spatial arrangement of bacteria in sponge tissue. AB - Widefield deconvolution epifluorescence microscopy (WDEM) combined with fluorescence in situ hybridization (FISH) was performed to identify and characterize single bacterial cells within sections of the mediterranean sponge Chondrosia reniformis. Sponges were embedded in paraffin wax or plastic prior to the preparation of thin sections, in situ hybridization and microscopy. Serial digital images generated by widefield epifluorescence microscopy were visualized using an exhaustive photon reassignment deconvolution algorithm and three dimensional rendering software. Computer processing of series of images taken at different focal planes with the deconvolution technique provided deblurred three dimensional images with high optical resolution on a submicron scale. Results from the deconvolution enhanced widefield microscopy were compared with conventional epifluorescent microscopical images. By the application of the deconvolution algorithm on digital image data obtained with widefield epifluorescence microscopy after FISH, the occurrence and spatial arrangement of Desulfovibrionaceae closely associated with micropores of Chondrosia reniformis could be visualized. PMID- 10699669 TI - A comparison of enumeration methods for culturable Pseudomonas fluorescens cells marked with green fluorescent protein. AB - The detection of bacteria in environmental samples using genetic markers is valuable in microbial ecology. The green fluorescent protein (GFP) reporter gene was studied under nutrient starvation conditions at 4 degrees C, 23 degrees C and 30 degrees C in Pseudomonas fluorescens R2fG1 cells tagged with a red-shifted gfp. Fluorescence intensity was not significantly different in cells maintained in a buffer for at least 48 days at all the tested temperatures. gfp-Tagged R2fG1 cells were introduced into bulk soil microcosms and soil microcosms with wheat seedlings. GFP-marked cells were enumerated immediately after inoculation into soil and again in soil and root samples after 10 days. Counts of culturable colonies were obtained from drop plates using 5-microl aliquots of serial dilutions viewed with an epifluorescent microscope. Traditional spread plates (using 100-microl aliquots) and the most-probable-number (MPN) method using a spectrofluorometer were also used to enumerate the GFP-marked Pseudomonas cells in soil, rhizosphere and rhizoplane samples. Microcolonies were visualized on root surfaces under the epifluorescent microscope after immobilizing in agar and incubation for 24 h. Counts from traditional spread plates were significantly higher (P<0.05) than the population estimates of the MPN method for all treatments at any sampling time. Counts using the drop plate method, however, were not significantly different (P<0.05) except in one treatment, and provided similar estimates in half the time of spread plates and at an estimated third of the cost. PMID- 10699670 TI - Evaluation of mini-VIDAS rapid test for detection of Listeria monocytogenes from production lines of fresh to cold-smoked fish. AB - This study was conducted to evaluate the efficacy of the mini-VIDAS Listeria monocytogenes (LMO) system (BioMerieux Vitek, Inc., Missouri, USA) for detection of L. monocytogenes in environmental and fish samples from three Portuguese cold smoking plants and from their fresh fish suppliers. Mini-VIDAS-LMO is a fully automated system that uses fluorescent ELFA (Enzyme Linked Fluorescent Assay) technology for detection of Listeria monocytogenes antigens in food. It can be a rapid screening method alternative to time consuming classical isolation and identification. Two hundred and ninety five samples were tested in mini-VIDAS-LMO and in parallel by the ISO 11290-1 traditional protocol. The mini-VIDAS-LMO detected 8 of the 11 confirmed positive samples and presented 11 false positive results. The specificity of the mini-VIDAS-LMO found in this experiment was 0.96 and the sensitivity 0.73. PMID- 10699671 TI - Critical factors influencing the recovery and integrity of rRNA extracted from environmental samples: use of an optimized protocol to measure depth-related biomass distribution in freshwater sediments. AB - A protocol was developed for the efficient recovery of intact, high molecular weight rRNA from different environmental matrices. Critical variables were identified in sample processing that influenced yield and integrity of recovered nucleic acid. Most notably, the order of addition and the buffer to sample volume ratio profoundly influenced the efficiency of nucleic acid recovery from sediment material when utilizing a guanidine thiocyanate-beta-mercaptoethaol extraction buffer. Addition of one sample volume to five buffer volumes contributed to an order of magnitude increase in recovery relative to reverse order of addition (buffer addition to sample). An optimized extraction protocol was used to evaluate rRNA yield by seeding samples with whole cells and radiolabeled nucleic acid. Recovery of intact rRNA was confirmed by polyacrylamide gel electrophoresis, which was also used to provide another estimate of quantity. This optimized protocol was used to measure depth-related changes in biomass distribution in Lake Michigan deep-water sediments. This revealed a biomodal biomass distribution; a maximum near the water/sediment interface and a secondary peak associated with the oxic/suboxic boundary. A significant portion of the community at the oxic/suboxic boundary was composed of non-methanogenic Archaea. PMID- 10699672 TI - An indirect hemagglutination antibody test to detect antibodies to Borrelia burgdorferi in patients with Lyme disease. AB - An indirect hemagglutination antibody (IHA) test was evaluated for its ability to detect borrelial antibodies in serum samples from patients with Lyme disease. The key test reagent developed for this antibody detection system was tannic acid treated and glutaraldehyde-fixed sheep red blood cells (SRBC) containing Borrelia burgdorferi (Bb) antigens attached to the outer surface of the SRBC. In order to establish suitable cut-off titers, initial specificity and sensitivity measurements were made using sera from 100 anonymous healthy volunteers and 30 additional pre-determined samples known to be non-reactive or reactive for Lyme disease or syphilis. These results were compared with those obtained using a commercially available ELISA. At titers >/=64, the IHA test had a combined 98% specificity and 100% sensitivity for these 130 serum samples, 30 of which were known positives or negatives, whereas the ELISA was less specific (93%) and much less sensitive (80%). Subsequent testing was performed on sera from 65 patients with the erythema migrans (EM) rash and 20 patients with early disseminated (cardiac/neurologic) symptoms or with Lyme arthritis. At initial presentation, 46 48% of the EM patients had IHA reactivity, with titers >/=128, while 42% were positive in the ELISA. Follow-up testing performed on these EM patients, 8-12 days after receiving antibiotic treatment, revealed that Bb antibodies were detected best by the IHA test (83-86% reactive) relative to the ELISA (81% reactive). Bb antibodies were readily detectable on all of the serum samples from the early disseminated and late stage Lyme disease cases in both assay systems. Based on these results and because of its technical and interpretive simplicity, the IHA test should be considered as a useful and convenient alternative for the serological analysis of Bb infections. PMID- 10699673 TI - A modified microtiter-plate test for quantification of staphylococcal biofilm formation. AB - The tube test and the microtiter-plate test are the most frequently used techniques for quantifying biofilm formation, an important indicator for the pathogenicity of staphylococci. The purpose of the present study was to develop a modified microtiter-plate technique for quantification of biofilm formation. This technique involves fixing the bacterial film with methanol, staining with crystal violet, releasing the bound dye with 33% glacial acetic acid, and measuring the optical density (OD) of the solution at 570 nm by using an enzyme immunosorbent assay reader. Biofilm formation of 30 Staphylococcus strains was estimated by the tube test, the standard microtiter-plate test and the modified microtiter-plate test. The modified microtiter-plate test, as a quantitative assay, is superior to the tube test in terms of objectivity and accuracy. It is also superior to the standard microtiter-plate test because it enables indirect measuring of bacteria attached both to the bottom and to the walls of the wells, while in the standard test only the dye bound to the bacteria adhered to the bottom of the wells is spectrophotometrically registered. Highly significant differences between OD values obtained by the standard microtiter-plate test and those obtained by the modified test suggest that large number of bacteria were attached to the walls of the wells. Therefore, the modification of the standard microtiter-plate test by introduction of an additional step of decolorization by acetic acid seems to be a useful improvement of the technique. PMID- 10699674 TI - The measurement of toluene dioxygenase activity in biofilm culture of Pseudomonas putida F1. AB - Toluene dioxygenase (Tod) enzyme activity can be measured by the conversion of indole to indigo. Indigo is measured spectrophotometrically at 600 nm. However, this method is inadequate to measure the whole-cell enzyme activity when interference by suspended biomass is present. Indoxyl is a highly fluorescent intermediate in the conversion of indole to indigo by Tod. A fluorescence-based assay was developed and applied to monitor Tod activity in whole cells of Pseudomonas putida F1 biofilm from a continuously operated biofilter. Suspended growth studies with pure cultures indicated that indoxyl, as measured by fluorescence, correlated with indigo production (r(2)=0.89) as measured by spectrophotometry. Whole-cell enzyme activity was followed during growth on a minimal medium containing toluene. The maximum normalized whole cell enzyme activity of 19+/-1.5x10(-4) mg indigo (mg protein)(-1) min(-1) was reached during early stationary phase. P. putida F1 cells from a biofilm grown on vapor phase toluene had a normalized whole-cell enzyme activity of 5.0+/-0.2x10(-4) mg indigo (mg protein)(-1) min(-1). The half-life of whole-cell enzyme activity was estimated to be between 5.5 and 8 h in both suspended and biofilm growth conditions. PMID- 10699675 TI - Screening method for detecting staphylococci with reduced susceptibility to teicoplanin. AB - The incidence of infections caused by staphylococci with decreased susceptibility to teicoplanin (MIC>/=8 microg/ml) is increasing, but the disk diffusion test has difficulty detecting this low level of resistance. In addition, detection is complicated because of the heterogeneous phenotypes for teicoplanin. In this study, we evaluated an agar screening method to detect staphylococci with decreased susceptibility to teicoplanin or heterogeneous resistance. First, to investigate the inoculum density and teicoplanin concentration of screening agar, we used 10(5) and 10(6) CFU/ml and Mueller-Hinton agars supplemented with 6 and 8 microg of teicoplanin/ml to test 39 genetically distinct staphylococcal strains (15 strains with teicoplanin MICs>/=8 microg/ml and 24 strains with teicoplanin MICs/=8 microg/ml or showing heteroresistance could be detected. These findings indicate that the method can be used as a reliable screening method for detecting staphylococci with reduced susceptibility to teicoplanin. PMID- 10699676 TI - Health insurance markets and income inequality: findings from an international health policy survey. AB - OBJECTIVE: To assess disparities in access to health care, financial burden of medical bills and perceived quality of care between those with above average incomes and those with below average incomes in five nations and to examine the relationship inequities in care experiences to health insurance coverage. DESIGN: Cross-sectional analysis of a random survey of adults in 1998. SUBJECTS: 5059 adults ages 18 and over in five English-speaking countries: Australia, Britain, Canada, New Zealand and the United States (approximately 1000 per country). MAIN OUTCOME MEASURES: Failure to receive needed care, difficulty getting care, waiting for elective surgery, problems paying medical bills, failure to fill prescriptions due to cost, perceived quality of medical care received and of most recent doctor visit. RESULTS: There were two to three-fold differences between those with above and below average incomes on measures of access to care in the US, Australia and New Zealand. In Britain and Canada indicators of access of to care were similar for the two income groups. Problems paying medical bills were most prevalent in the US, yet significant differences by income also existed in Australia, Canada and New Zealand. Those with below average incomes were more likely to have not filled a prescription due to cost in Australia, Canada, New Zealand and the US, with gaps by income most severe in the US. Ratings of quality of doctor visit were significantly different for the two income groups in the US, but not other countries. CONCLUSIONS: The analysis finds striking differences among countries in the relative equity of health care experiences. In general, care experiences are more unequal in three countries such as the US, Australia and New Zealand where systems have relatively greater reliance on private health insurance and markets. Greater inequality in care experiences is also associated with more divided public opinion regarding the need for system reform and the direction of recent policy changes. In Canada and Britain where care experiences are more equal of the health system are similar across income groups. Reliance on private insurance and patient user fees appears to lead to more divided views of the overall health system as well as inequity in access to care. PMID- 10699677 TI - Health outcome measures used in cost-effectiveness studies: a review of original articles published between 1986 and 1996. AB - Theoretically, the preferred type of health economic evaluation is the cost benefit approach in which costs as well as benefits are measured in monetary units. This type of analysis is rarely found in practice, however, where cost effectiveness analysis (CEA), cost-utility analysis (CUA) and other forms of economic evaluations are instead favored. The use of quality adjusted life-years (QALYs) or life-years gained, if applicable, is generally recommended in CUA/CEA because these measures will make possible broad comparisons with other studies as well as with norms regarding society's willingness-to-pay for health benefits. The purpose of this paper is to study the choice of health outcome measures and the extent to which results from CUA and CEA are discussed from such a willingness-to-pay perspective. Based on the analysis of a sample of 455 studies included in the Health Economic Evaluations Database (HEED), it is concluded that major differences exist in the choice of health outcome measures across disease categories. There is no evidence that QALYs or life-years gained have become more common over the years and CEAs using intermediary outcome measures are as common as those using life-years gained. Furthermore, studies using QALYs or life-years gained often lack a relevant discussion of society's willingness-to-pay per QALY or life-years gained. PMID- 10699678 TI - The Swiss database on quality projects in medical institutions. AB - Since the early 1990s it is important for every medical institution to report on activities in the field of quality improvement and quality assessment because there is a certain pressure from the market and from health insurance laws in various countries. Nevertheless, researchers as well as clinicians or administrators are rarely informed on ongoing projects. To register projects in quality research, an Internet-based information system was established to register projects on quality of medical institutions. Among others, hospitals, private doctors' offices, medical specialty societies and cost-payers are regarded as institutions in this effort. An interactive database provides information on the institution performing a project and on what is being/has been performed in a certain place during a certain period of time. At present medical institutions are invited to report on their projects, but this initiative can only succeed if it provides information from as many different institutions as possible: for data skills and help your colleagues! Place your knowledge, activities and information at disposal to the public and profit from your colleagues. PMID- 10699679 TI - An update on Spain's health care system: is it time for managed competition? AB - Using national data and the most recent OECD figures, we provide an updated assessment of the Spanish health care system and its reforms. We compare figures from Spain with other major industrialized nations and find that the Spanish system appears macro-economically efficient and equitable. However, like many other countries in Europe and elsewhere, the Spanish health care system confronts continued pressures to provide high-quality universal care in the face of ever increasing costs and competing uses for financial resources. These pressures have prompted the enactment of several reforms, which are reviewed. We draw from the American experience with managed care and managed competition to illustrate possible paths for further reform. PMID- 10699680 TI - Development of a knowledge-base for automatic monitoring of renal function of intensive care patients over time. AB - Renal dysfunction is a major problem in the management of critically ill patients. Monitoring of renal parameters over time is a prerequisite for detection of any significant deterioration of kidney function. Thus, we developed a knowledge-base for the dynamic monitoring of renal function of critically ill patients. A database with renal parameters of 750 intensive care patients was analyzed for distribution of parameters within predefined intervals of the creatinine clearance. Additionally, a subgroup of 11 patients with (quite) normal renal function over 11 days was selected and the daily variability of renal parameters was analyzed. An interdisciplinary expert team selected a set of nine clinically relevant renal parameters and formulated, on the basis of the data analysis and the parameter set, eight definitions of renal function, which represent four levels of renal performance. These definitions were arranged into an hierarchical structure, considering only clinically relevant changes of renal function. A change from one functional state to another inside of 2 days indicates a relevant alteration of renal function. Monitoring of time courses can additionally be performed by statistical analysis of the daily variability of parameters and comparison with their 'normal' variability. Moreover, rules were established for the plausibility check of results and interpretations of single parameters and parameter sets formulated. PMID- 10699681 TI - Application of neural networks and sensitivity analysis to improved prediction of trauma survival. AB - The performance of trauma departments is widely audited by applying predictive models that assess probability of survival, and examining the rate of unexpected survivals and deaths. Although the TRISS methodology, a logistic regression modelling technique, is still the de facto standard, it is known that neural network models perform better. A key issue when applying neural network models is the selection of input variables. This paper proposes a novel form of sensitivity analysis, which is simpler to apply than existing techniques, and can be used for both numeric and nominal input variables. The technique is applied to the audit survival problem, and used to analyse the TRISS variables. The conclusions discuss the implications for the design of further improved scoring schemes and predictive models. PMID- 10699682 TI - The biometrical comparison of cardiac imaging methods. AB - OBJECTIVES: Biometrical comparison procedures for cardiac imaging methods with continuous outcome are reviewed mainly concentrating on assessment and design adequate comparison of accuracy and precision. Univariate graphical and numerical representation of corresponding deviations is outlined to derive a 'check list' of minimum information necessary to compare the measurement methods. DATA: The methods reviewed here are illustrated by the comparison of standard 2DE bidimensional cardial volumetry versus assessment using TDE colour imaging in 28 normal probands. SOURCES: The paired t-test and the corresponding confidence interval approach are used to assess deviations in location of two imaging methods; the test procedures of Maloney and Rastogi Hahn and Nelson and Grubbs are surveyed as proposals for the comparison of precisions in paired data. The Krippendorff coefficient and the Bradley/Blackwood test are illustrated as surrogate measures for method concordance. CONCLUSIONS: Since these methods can be performed by simple modification of standard options available in most statistics software packages, this review intends to enable cardiologists to choose appropriate methods for statistical data analysis and representation on their own. PMID- 10699683 TI - Computer assisted determination of ATP in environmental and food samples by bioluminescent assay: comparison of algorithms. AB - When the bioluminescent assay (BLA) for the determination of adenosine triphosphate (ATP) in environmental and food samples is performed by conventional photometers, a computing procedure is necessary to exclude the mixing time of reagents from the global time of the reaction because it produces a poor analytical precision [M. Mecozzi, A. Amici, G. Visco, Fres. J. Anal. Chem. 357 (1997) 747-751]. In this paper three automated procedures for correcting this interference were tested and compared with illustrate their advantages and drawbacks. The three techniques were the robust exponential regression and the outliers detection method (RER), the smoothing technique of Savitzky-Golay (SSG) and the smoothing technique of Kalman Filter (SKF). RER and SSG were found to be more effective than SKF in minimizing the effect of noise produced by the mixing of reagents. In addition, the application of computing procedures such as RER and SSG allow to improve the detection limit in the BLA performed by conventional photometers. PMID- 10699684 TI - An easily customized, random allocation system using the minimization method for multi-institutional clinical trials. AB - In a randomized clinical trial, random allocation of patients to treatment groups should be done to balance in the distribution of prognostic factors. Random allocation in a multi-institutional randomized clinical trial is conducted by a coordinating center, independent of the medical institution the attending doctor uses for his/her practice. This study provides a sophisticated system for doing an exact random allocation of patients to treatment groups. The minimization method proposed by Pocock was applied to this system to balance the distribution of prognostic factors between two treatment groups, even when the number of registered patients is relatively small (S.J. Pocock, Allocation of patients to treatment in clinical trial, Biometrics 35 (1979) 183-197). Furthermore, Zelen's method is used to balance the number of patients allocated to the two groups within each institution (M. Zelen, The randomization and stratification of patients to clinical trials, J. Chron. Dis. 27 (1974) 365-375.). This system was created by the 'PERL&RSQUO; language for writing common gateway interface (CGI) script, and can therefore, be easily extended to include data entry function by attending doctors as well as the random allocation function. This system is being used effectively in thirteen multi-institutional randomized clinical trials for stomach, colon-rectum and breast cancers in Japan. PMID- 10699685 TI - A software tool to acquire, synchronise and playback multimedia data: an application in kinesiology. AB - Assessing the physical condition of the human body frequently requires evaluating different tests performed by different devices. This information has to be analysed simultaneously to derive conclusions, and so a temporal relationship must be established between the data sources. In this paper, synchronisation of the sources of information is achieved by synchronising the clocks of computers connected in a Local Area Network. A clock synchronisation protocol is used and a global time is stamped in each information data flow. During analysis, the time stamp is used to playback the data in a synchronised way. The proposed system is valid for any medical application in which data synchronisation is needed. However, it has initially been used in a distributed medical environment for analysing electrical muscular activity and lumbar movement. The dual aim is to detect injuries and plan training sessions for athletes. It was specifically used to analyse data generated by: electromyographs; lumbar equipment; and images generated by video cameras. The simple operation of the application and the precision of the synchronisation protocol enabled several interesting medical conclusions to be made. PMID- 10699686 TI - TEODOLINDA. A communication architecture for hospital information systems. AB - This paper is focused on a system for the release and distribution of messages and services among hospital units, which extends hospital information systems features in the field of communication and supports hospital organisation to fulfil healthcare commitments. PMID- 10699687 TI - Concurrent cisplatin/etoposide chemotherapy plus twice daily thoracic radiotherapy in limited stage small cell lung cancer: a phase II study. AB - Thirty-one previously untreated patients with limited stage small-cell lung cancer (LSCLC) were included in a prospective study, to investigate the feasability and the efficacy of a combined modality treatment using concurrent hyperfractionated chest irradiation and cisplatin (P) plus etoposide (E) chemotherapy. All patients received intravenously P=75 mg/m(2) at day 1, plus E=120 mg/m(2) days 1-3, at 3-week intervals for six cycles. Irradiated patients received 45 Gy in two daily fractions, 5 days a week, from week 4 to week 6. During week 5, prophylactic cranial irradiation was initiated, in one daily fraction of 2.5 Gy for a total dose of 25 Gy. Twenty-nine patients were evaluable for response. Twenty-two (76%) achieved a complete response, five (17%) had a partial response. Five patients are currently alive. The overall response rate was 93% (CI 95% (83.7-100)). The median survival time was 14 months and the 2 year survival rate was 25%. Main toxicities were grade 3-4 esophagitis in half of the patients and myelosuppression. The results are not as optimistic as other studies using a similar regimen. PMID- 10699688 TI - A multicenter, randomized, phase III study of docetaxel plus best supportive care versus best supportive care in chemotherapy-naive patients with metastatic or non resectable localized non-small cell lung cancer (NSCLC). AB - This was an open-label randomized Phase III study of 207 patients with either unresectable or metastatic non-small cell lung cancer (NSCLC) who were treated with docetaxel plus best supportive care (BSC) or best supportive care alone. Patients in the chemotherapy arm of the study received docetaxel 100 mg/m(2) as a 1 h intravenous infusion every 21 days until they showed evidence of progressive disease, or estimated maximum benefit obtained or unacceptable side effects. Patients who received docetaxel were pretreated with oral dexamethasone. Patients in the BSC arm should not receive chemotherapy or anticancer therapy except for palliative radiotherapy. Overall survival obtained in the docetaxel arm was significantly longer than in the BSC arm (P=0.026). Two-year survival in the docetaxel arm was 12%, whereas none of the BSC patients survived after 20 months. The response rate was 13.1% (95% CI, 7.5-18.8%). There was a significantly longer time to progression in the docetaxel versus the BSC arm (P<0.001), and statistically significant improvement of clinical symptoms with docetaxel compared to BSC. The quality-of-life descriptors were in favor of docetaxel, and the difference was significant for pain, dyspnea and emotional functioning. The safety profile of docetaxel for this study was similar to that already reported in this patient population. PMID- 10699689 TI - Significance of serum pro-gastrin-releasing peptide as a predictor of relapse of small cell lung cancer: comparative evaluation with neuron-specific enolase and carcinoembryonic antigen. AB - Neuron-specific enolase (NSE) and carcinoembryonic antigen (CEA) have been reported to be useful markers for staging, monitoring treatment, and predicting relapse in patients with small cell lung cancer (SCLC). Recently, pro-gastrin releasing peptide (Pro-GRP) became available as a sensitive, specific, and reliable tumor marker for patients with SCLC. The aim of this study is to determine the most useful tumor marker to detect the relapse of SCLC. Furthermore, we analyzed the relationship between tumor markers at relapse and survival from relapse or response to salvage chemotherapy. Medical records were reviewed to obtain serum levels of Pro-GRP, NSE, and CEA before and after the initial chemotherapy, and at relapse. Consecutive 66 patients with SCLC, with an objective response and confirmed relapse treated at the National Cancer Center Hospital East, were analyzed in this study. The percentages of patients whose tumor marker level were elevated before treatment, decreased after the treatment, and increased again at relapse were 67% (95% CI, 55-78) for Pro-GRP, 20% (10-29) for NSE, and 38% (26-50) for CEA. Multivariate analysis indicated that poor performance status before initial treatment and elevated serum levels of lactate dehydrogenase at relapse were poor prognostic factors for patients with recurrent SCLC (P<0.005). None of the serum levels of Pro-GRP, NSE, and CEA at relapse was a significant prognostic factor and associated with an objective response to salvage chemotherapy. The present study demonstrated that serum levels of Pro-GRP reflect the disease course of patients with SCLC most accurately. PMID- 10699690 TI - Combined Nd-YAG laser/HDR brachytherapy versus Nd-YAG laser only in malignant central airway involvement: a prospective randomized study. AB - BACKGROUND: Laser debulking and prosthetic stents are useful modalities in the palliative treatment of initial inoperable or recurrent lung cancer. Recently, endobrochial brachytherapy was introduced to extend the duration of palliation and reduce the number of endoscopic treatments. This trial compares Nd-YAG laser alone and associated to high dose rated (HDR)-brachytherapy. PATIENTS AND METHODS: From 1995 to 1998, 29 consecutive patients, with non-small cell lung cancer (NSCLC) and central airway involvement, were randomized in two groups: group 1 (15 patients) received Nd-YAG laser only; group 2 (14 patients) underwent a combined Nd-YAG laser/ HDR brachytherapy treatment. RESULTS: There was no mortality or morbidity related to the treatment. The period free from symptoms was 2.8 months for group 1 and increased to 8.5 months in group 2 (P<0.05). The disease's progression free period grew from 2.2 months of group 1 to 7.5 months of group 2 (P<0.05) and the number of further endoscopic treatment reduced from 15 to 3 (P<0.05). CONCLUSION: The results confirm the potential of brachytherapy to prolong relief from symptoms, lessen disease progression and reduce costs of treatment. A detailed analysis is presented of both groups. PMID- 10699691 TI - Evasion mechanisms to tumor necrosis factor alpha (TNF-alpha) of small cell lung carcinoma and non-small cell lung carcinoma cell lines: comparison with the erythroleukaemia K-562 cell line. AB - The tumour necrosis factor alpha (TNF-alpha) is produced by mononuclear phagocytes as a defence mechanism against malignant cells. However, these cells can evade destruction by TNF-alpha. The present study evaluates in three lung cancer cell lines (small cell carcinoma NCI-H69, adenocarcinoma A-427, squamous carcinoma SK-MES-1) and one erythroleukaemia (K-562) cell line the following evasion mechanisms: (1) inhibition of TNF-alpha production, in indirect and direct co-cultures with monocytes; (2) the expression of type I and type II receptors for TNF-alpha (TNFRI and TNFRII) by tumour cell lines, using indirect immunofluorescence and flow cytometry; (3) the sensitivity of tumour cell lines to the toxic action of recombinant human TNF-alpha (rhTNF-alpha). With the exception of cell line NCI-H69, the other tumour cell lines liberated soluble factors that inhibited TNF-alpha production in monocytes. This effect occurred even after membrane contact with the A-427 and SK-MES-1 cell lines. Erythroleukaemia K-562 cells expressed both types of receptors for TNF-alpha, whereas the NCI-H69 cells expressed only TNFRI, and the A-427 and SK-MES-1 cells expressed no receptors. Lines NCI-H69, A-427 and K-562 were insensitive to the cytotoxic action of rhTNF-alpha. In conclusion, different lung cancer cell lines may evade destruction by TNF-alpha by various mechanisms that range from blocking TNF-alpha production by monocytes to blocking the cytotoxic action of this molecule. For selecting the most effective immunotherapy, knowledge of the evasion mechanisms would be useful. PMID- 10699692 TI - Detection of bone marrow metastases of small cell lung cancer with magnetic resonance imaging: early diagnosis before destruction of osseous structure and implications for staging. AB - Small cell lung cancer (SCLC) frequently metastasizes to bone and bone marrow. Skeletal scintigraphy and bone marrow cytology or biopsy, are incorporated into the staging procedures to examine these organs. However, skeletal scintigraphy is not highly specific to metastases, and only one or two bone marrow sites can be examined by cytology or biopsy. We have already reported that magnetic resonance imaging (MRI) could improve the sensitivity in detecting bone marrow metastases. The result of the bone marrow MRI was an independent prognostic factor of SCLC patients [9]. In the present study, we analyzed the results of skeletal scintigraphy and bone marrow aspiration with special reference to the results of MRI examination. We also analyzed the relationship between bone marrow lesions and bone lesions. For this purpose, we visualized bone marrow metastases with MRI and determined their anatomical locations and sizes. Approximately half of bone marrow lesions stayed in bone marrow during follow-up period ranging from 57 to 154 days, whereas about half of them were accompanied by hot spots in follow-up skeletal scintigraphy, which indicates the destruction of osseous structure. Additionally, 87.5% of osteolytic changes that newly appeared in skeletal scintigraphy were preceded by adjacent bone marrow lesions. All new lesions that appeared in follow-up skeletal scintigraphy within 3 months after the initial presentation had the preceding bone marrow lesions. These results mean that almost all lesions in skeletal scintigraphy derived from bone marrow metastases. Furthermore, appreciable volume of cancer cells is present in bone marrow before osteolytic changes appear in skeletal scintigraphy. PMID- 10699693 TI - Bone metastasis after a resection of stage I and II primary lung cancer. AB - In the present study, we reviewed the patients who developed bone metastases after a surgical resection of primary lung cancer and evaluated their clinicopathological features. From 1992 to 1995, 177 patients with stage I and II primary lung cancer underwent a surgical resection at the Kitakyushu Municipal Medical Center. Bone metastases were detected in 14 patients (7.9%) by follow-up examinations including bone scintigraphy (scan). Bone metastasis was one of the most frequent extra-thoracic recurrent forms. Patients with adenocarcinoma tended to develop bone metastases more frequently than those with squamous cell carcinoma. In the preoperative bone scans, an abnormal uptake was observed in 76 patients (42.9%), and 10 (13.1%) of them were found to develop bone metastases in the follow-up studies. A microscopic examination of the primary tumor demonstrated close correlation between intratumoral and peritumoral lymphatic vessel invasion and postoperative development of bone metastases. A bone scan is a very useful and indispensable procedure for diagnosing bone metastases. However, this scan may also show false positive finding in a number of benign conditions. Therefore, a surgical resection should be considered as the first line treatment for patients with positive findings in the bone scan when the diagnosis of bone metastasis can not be confirmed based on both their symptoms and other clinical examinations. PMID- 10699694 TI - Long term survival of a patient with malignant pleural mesothelioma as a late complication of radiotherapy for Hodgkin's disease treated with 90yttrium silicate. AB - A case of malignant pleural mesothelioma (PM) 24 years after thoracic radiotherapy for Hodgkin's disease is presented. As primary treatment and to relieve symptoms of dyspnea secondary to pleural effusion a thoracic drain was installed, followed by intracavitary radiation therapy with 90yttrium-silicate. Minor complaints of fever and a dry cough as a side-effect of this treatment were effectively treated with prednisone during 2 weeks. The patient remains in a good clinical condition now 6 years after diagnosis. Considering the few therapeutic options the use of 90yttrium-silicate intrapleural installation could be propagated as a safe and effective antitumour treatment for a selected group of patients with malignant PM. PMID- 10699695 TI - Incidence, duration and survival of ventricular fibrillation in out-of-hospital cardiac arrest patients in sweden. AB - The chance of survival from ventricular fibrillation (VF) is up to ten times higher than those with other cardiac arrest rhythms. To calculate the effect of out-of-hospital resuscitation organisations on survival, it is necessary to know the percentage of cardiac arrest patients initially in VF and the relationship between delay time to defibrillation and survival. AIM: To study the incidence of VF at the time of cardiac arrest and on first ECG, the duration of VF and the relation between time to defibrillation and survival. METHOD: The Swedish Cardiac Arrest Registry has collected standardised reports on out-of-hospital cardiac arrests from ambulance organisations in Sweden, serving 60% of the Swedish population. RESULTS: In 14065 cases of out-of-hospital cardiac arrest collected between 1990 and 1995, resuscitation was attempted in 10966 cases. INCIDENCE: The first ECG showed VF in 43% of all patients. The incidence of VF at the time of cardiac arrest was estimated to be 60-70% in all patients and 80-85% in the cases with probable heart disease. DURATION: The estimated disappearance rate of VF was slow. Thirty minutes after collapse approximately 40% of the patients were in VF. SURVIVAL: Overall survival to 1 month was only 1.6% for patients with non shockable rhythms and 9.5% for patients found in VF. With increasing time to defibrillation, the survival rate fell rapidly from approximately 50% with a minimal delay to 5% at 15 min. CONCLUSIONS: This study suggests a high initial incidence of VF among out-of-hospital cardiac arrest patients and a slow rate of transformation into a non-shockable rhythm. The survival rate with very short delay times to defibrillation was approximately 50%, but decreased rapidly as the delay increased. PMID- 10699696 TI - Possibilities of implementing dispatcher-assisted cardiopulmonary resuscitation in the community. An evaluation of 99 consecutive out-of-hospital cardiac arrests. AB - AIM: By evaluating tape recordings of true cardiac arrest calls, to judge the dispatchers ability to (a) identify cases as suspected cardiac arrest (CA), (b) give the case the right priority, (c) identify CA cases suitable for dispatcher assisted, telephone-guided cardiopulmonary resuscitation (T-CPR) and (d) accomplish T-CPR. METHODS: Evaluation of 99 tape recordings of consecutive cases that had been admitted to the two city hospitals in Goteborg after out-of hospital CA. RESULTS: In 70% of the interviews, the dispatcher demonstrated impeccable behaviour with short, distinct questions, quickly resulting in a decision on how to handle the case. In 30%, serious criticism could be voiced as the dispatcher displayed very stressful behaviour, or omitted to ask important questions such as whether the patient was conscious and breathing. In 21%, the interviews indicated a clear opportunity to perform T-CPR. In another 10%, there was a possibility of performing T-CPR. Only in 8% was T-CPR actually accomplished. CONCLUSIONS: (1) In the majority of the interviews, the quality was very high, while in one-third, serious criticism could be voiced. (2) In our study, only one-third (95% confidence interval, 22-41) of CA cases were suitable for T-CPR, and T-CPR was performed in only 8% of the 99 cases. (3) To optimise the dispatcher ability to identify suspected CA and initiate T-CPR, both medical knowledge and practical training are needed, preferably with protocols for pre arrival instructions. PMID- 10699697 TI - Attitudes of trained Swedish lay rescuers toward CPR performance in an emergency. A survey of 1012 recently trained CPR rescuers. AB - 59 years old. Only 1% had attended the course because of their own or a relative's cardiac disease. Ninety-four per cent believed there was a minor to major risk of serious disease transmission while performing CPR. When predicting their willingness to perform CPR in six scenarios, 17% would not start CPR on a young drug addict, 7% would not perform CPR on an unkempt man, while 97% were sure about starting CPR on a relative and 91% on a known person. In four of six scenarios, respondents from rural areas were significantly more positive than respondents from metropolitan areas about starting CPR. In conclusion, readiness to perform CPR on a known person is high among trained CPR rescuers, while hesitation about performing CPR on a stranger is evident. Respondents from rural areas are more frequently positive about starting CPR than those from metropolitan areas. PMID- 10699698 TI - Smaller tidal volumes with room-air are not sufficient to ensure adequate oxygenation during bag-valve-mask ventilation. AB - The European Resuscitation Council has recommended decreasing tidal volume during basic life support ventilation from 800 to 1200 ml, as recommended by the American Heart Association, to 500 ml in order to minimise stomach inflation. However, if oxygen is not available at the scene of an emergency, and small tidal volumes are given during basic life support ventilation with a paediatric self inflatable bag and room-air (21% oxygen), insufficient oxygenation and/or inadequate ventilation may result. When apnoea occurred after induction of anaesthesia, 40 patients were randomly allocated to room-air ventilation with either an adult (maximum volume, 1500 ml) or paediatric (maximum volume, 700 ml) self-inflatable bag for 5 min before intubation. When using an adult (n=20) versus paediatric (n=20) self-inflatable bag, mean +/-SEM tidal volumes and tidal volumes per kilogram were significantly (P<0.0001) larger (719+/-22 vs. 455+/-23 ml and 10.5+/-0.4 vs. 6.2+/-0.4 ml kg(-1), respectively). Compared with an adult self-inflatable bag, bag-valve-mask ventilation with room-air using a paediatric self-inflatable bag resulted in significantly (P<0.01) lower paO(2) values (73+/ 4 vs. 87+/-4 mmHg), but comparable carbon dioxide elimination (40+/-2 vs. 37+/-1 mmHg; NS). In conclusion, our results indicate that smaller tidal volumes of approximately 6 ml kg(-1) ( approximately 500 ml) given with a paediatric self inflatable bag and room-air maintain adequate carbon dioxide elimination, but do not result in sufficient oxygenation during bag-valve-mask ventilation. Thus, if small (6 ml kg(-1)) tidal volumes are being used during bag-valve-mask ventilation, additional oxygen is necessary. Accordingly, when additional oxygen during bag-valve-mask ventilation is not available, only large tidal volumes of approximately 11 ml kg(-1) were able to maintain both sufficient oxygenation and carbon dioxide elimination. PMID- 10699699 TI - Finger position for chest compressions in cardiac arrest in infants. AB - OBJECTIVE: To determine whether the recommended method of locating finger position for chest compression in infant cardiac arrest can cause pressure on the abdomen or xiphisternum. DESIGN: The length from the inter-nipple line to the xiphisternum was calculated in 30 infants. These lengths were compared with the finger position achieved by 30 adults, using the recommended method, on templates of infant chests. RESULTS: The mean infant lower sternal length was 2.3 cm (95% CI 1.6). The mean distance covered by the adults fingers was 4.4 cm (95% CI 0.9). CONCLUSION: If any infant in this study had chest compressions performed by any of the adults, using the recommended method, pressure would be exerted on the xiphisternum or abdomen. We suggest changing the method of locating finger position, to one using sternal anatomy. PMID- 10699700 TI - Adenosine by aortic flush fails to augment the brain preservation effect of mild hypothermia during exsanguination cardiac arrest in dogs - an exploratory study. AB - Most trauma cases with rapid exsanguination to cardiac arrest (CA) in the field, as well as many cases of normovolemic sudden cardiac death are 'unresuscitable' by standard cardiopulmonary-cerebral resuscitation (CPCR). We are presenting a dog model for exploring pharmacological strategies for the rapid induction by aortic arch flush of suspended animation (SA), i.e. preservation of cerebral viability for 15 min or longer. This can be extended by profound hypothermic circulatory arrest of at least 60 min, induced and reversed with (portable) cardiopulmonary bypass (CPB). SA is meant to buy time for transport and repair during pulselessness, to be followed by delayed resuscitation to survival without brain damage. This model with exsanguination over 5 min to CA of 15-min no-flow, is to evaluate rapid SA induction by aortic flush of normal saline solution (NSS) at room temperature (24 degrees C) at 2-min no-flow. This previously achieved normal functional recovery, but with histologic brain damage. We hypothesized that the addition of adenosine would achieve recovery with no histologic damage, because adenosine delays energy failure and helps repair brain injury. This dog model included reversal of 15-min no-flow with closed-chest CPB, controlled ventilation to 20 h, and intensive care to 72 h. Outcome was evaluated by overall performance, neurologic deficit, and brain histologic damage. At 2 min of CA, 500 ml of NSS at 24 degrees C was flushed (over 1 min) into the brain and heart via an aortic balloon catheter. Controls (n=5) received no drug. The adenosine group (n=5) received 2-chloro-adenosine (long acting adenosine analogue), 30 mg in the flush solution, and, after reperfusion, adenosine i.v. over 12 h (210 microg/kg per min for 3 h, 140 microg/kg per min for 9 h). The 24 degrees C flush reduced tympanic membrane temperature (T(ty)) within 2 min of CA from 37.5 to approximately 36.0 degrees C in both groups. At 72 h, final overall performance category (OPC) 1 (normal) was achieved by all ten dogs of the two groups. Final neurologic deficit scores (NDS; 0-10% normal, 100% brain death) were 5+/-3% in the control group versus 6+/-5% in the adenosine group (NS). Total brain histologic damage scores (HDS) at 72 h were 74+/-9 (64-80) in the control group versus 68+/-19 (40-88) in the adenosine group (NS). In both groups, ischemic neurons were as prevalent in the basal ganglia and neocortex as in the cerebellum and hippocampus. The mild hypothermic aortic flush protocol is feasible in dogs. The adenosine strategy used does not abolish the mild histologic brain damage. PMID- 10699701 TI - Pentoxifylline fails to improve organ dysfunction and survival when used in the resuscitation of a porcine model of haemorrhage and abdominal sepsis. AB - Pentoxifylline is a phosphodiesterase inhibitor, known to suppress tumour necrosis factor-alpha production and improve cardiopulmonary parameters and survival in animal models of sepsis. Using a porcine model of abdominal trauma resulting from the combined insults of haemorrhage and infection, a randomised placebo-controlled trial was conducted of pentoxifylline (20 mg/kg bolus followed by 20 mg/kg infusion over 1 h) administered in addition to a colloid resuscitation regimen. Female Large White pigs (45-60 kg) were bled 40% of their blood volume and peritonitis was induced using E. coli (O18: K1: H7) in an autoclaved faecal suspension. Animals were resuscitated with either colloid alone (n=5) or colloid plus pentoxifylline (n=5). Pentoxifylline attenuated increases in mean arterial and pulmonary artery pressures and reduced both systemic and pulmonary vascular resistance. It worsened the lactic acidosis associated with 'septic shock' and failed to reduce serum TNF-alpha levels. Pentoxifylline, in the high doses used in this study, does not have a role as an adjunct to resuscitation in this clinically relevant model of trauma. PMID- 10699702 TI - A case report of difficult ventilation with the Combitube - valve-like upper airway obstruction confirmed by fibreoptic visualisation. AB - This case report describes difficulty with ventilation because of valve-like upper airway obstruction by aryepiglottic folds after uncomplicated insertion of a Combitube in a 30-year-old female patient. After correct (oesophageal) placement increased ventilation pressures occurred and a fibreoptic device was used to investigate the cause. Valve-like obstruction was discovered and subsequently observed during controlled ventilation. After removal of the Combitube and mask ventilation no valve mechanism was seen. This effect appeared to be due to an increased air stream caused by the obstruction of seven out of eight Combitube perforations. PMID- 10699703 TI - The use of some ingredients for microemulsion preparation containing retinol and its esters. AB - A study of microemulsions with retinol and its esters. Physical properties of w/o and o/w microemulsions containing Tween 60, Tween 80, Epicurone 135 (soy bean lecithin) as surfactants, n-butanol, triacetin, propylene glycol as cosurfactants were examined. The drug-containing systems were characterised in regard to their ophthalmic parameters. Physiologically well-tolerated and physically stable multiple-components were developed. The concentrations of surfactants and cosurfactants which are necessary to form stable systems were evaluated. The values of the following parameters--refractive index, viscosity, pH value, osmotic tension, obtained in the study, proved suitable for the purpose and the preparations were physiologically tolerated, the use of microemulsions as potential drug delivery systems for ocular administration has been discussed. The influence of retinol and its esters on the physical parameters the preparation was investigated. Microemulsion stored at 20 degrees C up to 6 months showed no significant physical changes. PMID- 10699704 TI - Hydration of lipid films with an aqueous solution of Quil A: a simple method for the preparation of immune-stimulating complexes. AB - Immune-stimulating complexes (ISCOMs) are stable colloidal complexes of the adjuvant Quil A, cholesterol and phospholipid, which are effective carriers for subunit vaccines. The techniques currently available for the preparation of ISCOMs from the constituent components are rather complex and are based on either centrifugation or dialysis. This note reports a new simple procedure for the preparation of ISCOM matrices based on hydration of a cholesterol/phospholipid film with an aqueous solution of Quil A. It is demonstrated that ISCOM matrices do not form in the absence of phospholipid when prepared by this method. Further, the ratio by weight of phospholipid to either cholesterol or Quil A must be greater than that required for preparation by either dialysis or centrifugation. Photon correlation spectroscopy, negative stain transmission electron microscopy and centrifugation through a sucrose gradient demonstrate that ISCOM matrices can be prepared from cholesterol/lipid films by hydration with an aqueous solution of Quil A when the ratio of phospholipid:cholesterol:Quil A by weight is 6:1:4, respectively. Lower ratios of phospholipid:cholesterol reduce the efficiency of ISCOM formation while higher ratios produce systems containing a mixture of ISCOMs together with liposomes. PMID- 10699705 TI - Effects of alcohols and diols on the phase behaviour of quaternary systems. AB - The aim of the current study was to investigate the effect of different co surfactants on the phase behaviour of the pseudoternary system water:ethyl oleate:nonionic surfactant blend (sorbitan monolaurate/polyoxyethylene 20 sorbitan mono-oleate). Four aliphatic alcohols (1-propanol, 1-butanol, 1-hexanol and 1-octanol) and four 1, 2-alkanediols (1,2-propanediol, 1,2-pentanediol, 1,2 hexanediol and 1,2-octanediol) were used. The co-surfactant-free system forms two different colloidal structures, a water-in-oil microemulsion (w/o ME) and lamellar liquid crystals (LC) and two coarse dispersions, water-in-oil (w/o EM) and oil-in-water (o/w EM) emulsions. Microemulsion region area (%ME), liquid crystalline region area (%LC), amount of amphiphile blend required to produce a balanced microemulsion (%AMPH) and amount of water solubilised (%W) were used as assessment criteria to evaluate the co-surfactants. Seven calculated physico chemical descriptors were used to represent the different co-surfactants. 1 butanol, 1,2-hexanediol and 1, 2-octanediol produced balanced MEs capable of solubilising a high percentage of both oil and water. A similarity was observed between the descriptors attributed to 1-butanol and 1,2-hexanediol. The requirements of a co-surfactant molecule to produce a balanced microemulsion were: HLB value 7.0-8.0, a carbon backbone of 4-6 atoms, percentage carbon of 60 65%, percentage oxygen of 20-30%, logP value 0.2-0.9 and log 1/S (S: aqueous solubility) close to zero. PMID- 10699706 TI - Preparation of avidin-labelled gelatin nanoparticles as carriers for biotinylated peptide nucleic acid (PNA). AB - The possibility of preparing uniform nanoparticles consisting of proteins such as gelatin followed by covalent linkage of avidin was investigated. Gelatin nanoparticles were prepared by two step desolvation. Functional groups at the surface of the particulate system were quantified with site-specific reagents. The surface of the nanoparticles was thiolated and avidin was covalently attached to the nanoparticles via a bifunctional spacer at high levels. Biotinylated peptide nucleic acid (PNA) was effectively complexed by the avidin-conjugated nanoparticles. Avidin-conjugated protein nanoparticles should prove as potential carrier system for biotinylated drug derivatives in antisense therapy. PMID- 10699707 TI - Bench scale manufacture of multilamellar liposomes using a newly developed multistage pressure filtration device. AB - Liposomes are belonging to the modern kinds of drugs. They are facilitating the secure and well-targeted transport of substances inside the organism, and are gaining increasing importance in the pharmacological treatment of tumors and in gene-therapeutic strategies. Multilamellar liposomes have advantages to one-layer liposomes: the multiplicity of coats increases the effect of a reservoir and makes extremely prolonged releases of drugs possible. This study describes the aseptical manufacturing of different kinds of multilamellar liposomes. It has been shown that it is possible to produce liposome-suspensions with different shares of multilamellar liposomes in the scale of litres under aseptical circumstances using a newly constructed multi-stage-pressure-filtration-device. PMID- 10699708 TI - Nanosuspensions of poorly soluble drugs--reproducibility of small scale production. AB - The major problem of many newly developed pharmaceutical drugs is their poor solubility in water and simultaneously in organic media. To solve these problems formulation as nanosuspensions is an attractive alternative. During the drug development process screening for an optimal formulation by homogenisation is essential. Time and cost effective production in an initial phase of R&D can be conducted on lab scale by using the Micron Lab 40 in its discontinuous version. In this report reproducibility of small scale production parameters (particle size, size distribution, content of microparticles) was exemplary studied for the drug RMKP22. PMID- 10699709 TI - Nanosuspensions as a new approach for the formulation for the poorly soluble drug tarazepide. AB - Poorly soluble drugs are often a challenging problem in drug formulation, especially when the drug is not soluble in either aqueous media or organic solvents. Attempts to overcome the solubility problem are, e.g. solubilisation with mixed micelles or forming a complex using cyclodextrines, but these approaches are of limited success. Another problem with new high potential drug is that these drugs often show bioavailability problems. One tried to improve the in vivo performance of poorly soluble drugs by reducing the particles size of the drug thus leading to an increased surface area and an increased dissolution velocity (Muller et al., 1994, 1999). Some of these problems occurred with tarazepide and therefore it was tried to create a formulation with this drug as nanosuspension which is suitable for intravenous administration. PMID- 10699710 TI - Solid lipid nanoparticles as drug carriers for topical glucocorticoids. AB - Recent investigations both in vitro and in human subjects proved the benefit/risk ratio of prednicarbate (PC) to exceed those of halogenated topical glucocorticoids about 2-fold. To obtain a further highly desired increase by drug targeting to viable epidermis, PC was incorporated into solid lipid nanoparticles (SLN). Keratinocyte and fibroblast monolayer cultures, reconstructed epidermis and excised human skin served to evaluate SLN toxicity and PC absorption. Well tolerated preparations (e.g. cellular viability 94.5% following 18 h incubation of reconstructed epidermis) were obtained. PC penetration into human skin increased by 30% as compared to PC cream, permeation of reconstructed epidermis increased even 3-fold. The present study shows the great potential of SLN to improve drug absorption by the skin. PMID- 10699711 TI - Heavy metal contamination of nanosuspensions produced by high-pressure homogenisation. AB - High pressure homogenisation is a method for the production of nanosuspensions. In this process crystalline drug particles are pressed with high pressure through a narrow homogenisation gap. Due to the conditions in the gap it seems possible that metal erosion can occur. In this study the heavy metal (Fe) contamination of nanosuspensions produced by high pressure homogenisation was determined. Therefore nanosuspensions were analysed by atom absorption spectroscopy concerning their load of iron which is chosen as reference metal. The results show that the erosion of metal is below 1 ppm and will not cause any toxicological problems. PMID- 10699712 TI - Microencapsulation of peptides and proteins. AB - Microcapsules were prepared by using a double-emulsion technique. A new production method called 'induced phase separation method' was applied to encapsulate peptides and proteins. To find the optimal adjuvants a matrix was set up combining the appropriate organic solvents and the suitable surfactants. The polymer was chosen with regard to the required release period. The aqueous drug solution was intensively mixed with the organic polymer solution. An aqueous surfactant solution was slowly added to the O/W emulsion. The obtained W/O/W emulsion is stirred under partial vacuum conditions until the organic solvent was removed. After removing the solvent from the W/O/W emulsion the microcapsules were washed and lyophilized. The morphology of the microparticles (spheres, sponges, capsules, surplus polymer) was checked by microscopy, particle size distributions were measured by laser diffraction. PMID- 10699713 TI - Influences of process parameters on nanoparticle preparation performed by a double emulsion pressure homogenization technique. AB - The preparation of nanoparticles (NP) as an improved colloidal carrier system for proteins was investigated. Bovine serum albumin (BSA) was used as model drug. Owing to the high solubility of the protein in water, the double emulsion technique has been chosen as one of the most appropriate method. In order to both reaching submicron size as well as increasing the grade of monodispersity compared to previous preparation techniques, a microfluidizer as homogenization device was used. All experiments were performed using two biodegradable polymers, poly[D,L-lactic-co-glycolic acid] 50/50 (PLGA) and poly[epsilon-caprolactone] (PCL). The homogenization procedure has been optimized with regard to particle size and monodispersity by studying the influence of the homogenization time as well as the amount of polymer and surfactant in the external aqueous phase. The drug loading has been improved by varying the concentration of the protein in the inner aqueous phase. By increasing the protein concentration in the inner aqueous phase the polydispersity was slightly higher, while the particle size was not influenced significantly. The BSA encapsulation efficiency decreased with higher protein concentration in the inner aqueous phase. All release profiles were characterized by a initial burst effect, a higher release rate was obtained after 4 weeks for PLGA NP (60%) compared with PCL NP (47%). PMID- 10699714 TI - Influence of high pressure homogenisation equipment on nanodispersions characteristics. AB - In this study a comparison of the influence of the homogenising equipment supplied by different manufacturers on the quality of the lipid nanodispersions is given. An Avestin EmulsiFlex-B3 (B3) and APV Micron Lab 40 (LAB 40) were used for high pressure homogenisation. Particle size and particle size distribution were chosen as quality parameters. The influence of different process parameters was evaluated. The two homogenisers were compared in their quality of nanoparticles-production by hot and cold homogenisation technique and in processing nanoemulsions. Working with the B3 appeared as useful for preformulation studies and processing of expensive or rare drugs and excipients. This first scaling up within laboratory scale is evaluated and the problems and remarkable aspects working with the B3 are pointed out. PMID- 10699715 TI - Direct detection of dissolution of 14C-labeled compounds into an oil phase by the fat scintillation proximity method. AB - Traditional analysis of the dissolution of lipophilic compounds from aqueous phase into oil is often hampered by the necessity to separate the receiver oil compartment from the aqueous phase. In order to avoid possible artefacts associated with additional separation methods, a procedure was developed to selectively detect the entry of a compound into the oil phase of a oil/water dispersion. When a combination of a primary and secondary scintillator was predissolved in the oil, and solid 14C-tetrahydrolipstatin was added, increasing signals from the same container were measured upon prolonged incubation. The data are consistent with the hypothesis that 14C-THL that has dissolved in the oil phase is essentially responsible for the measured signal. The obtained dissolution profiles of 14C-THL into oil match with parallel experiments using classical procedures. PMID- 10699716 TI - Controlled release of solid-reversed-micellar-solution (SRMS) suppositories containing metoclopramide-HCl. AB - The investigated drug delivery system is a solid-reversed-micellar-solution (SRMS). The composition of this solution is 70% Witepsol W35 and 30% (w/w) lecithin. 1% (w/w) metoclopramide-HCl (MCP) was solubilized in the SRMS. After melting and on contact with water or any physicological aqueous media the SRMS exhibits an application induced transformation into a semisolid system of liquid crystalline microstructure. The structure of the liquid crystal has been identified by polarized light microscopy as a lamellar mesophase. Due to a low coefficient of diffusion in this mesophase a controlled release of the drug may be possible. The release profiles of the in vitro experiments have shown zero order kinetics and a sustained release of the SRMS-suppositories (SRMS-supp.) in comparison with commercial suppositories (Gastrosil-supp.). To examine bioavailability an in vivo study with rabbits was carried out. Five SRMS-supp. (10 mg MCP) and five Gastrosil-supp. (10 mg MCP) were tested in a parallel-group study. These experiments have shown a five times longer mean residence time (parameter of sustained release) in comparison with Gastrosil-supp. In vitro and in vivo studies have shown that rectal application of SRMS-supp. provides an appropriate route for controlled release of MCP via application induced transformation into liquid crystals. PMID- 10699717 TI - Desolvation process and surface characteristics of HSA-nanoparticles. AB - The objective of the present study was to characterise and optimise the desolvation process of human serum albumin (HSA) for the preparation of nanoparticles. Following the desolvation of the protein, the resulting nanoparticles were stabilised by the addition of varying amounts of glutaraldehyde. The particle size and the number of available amino groups on the surface of the nanoparticles were determined. The results indicated that the particle size depended mainly on the amount of desolvating agent added, but not on the amount of cross-linker. Increasing volumes of glutaraldehyde reduced the number of amino groups on the surface of HSA nanoparticles. PMID- 10699718 TI - Crystallographic investigation of cetylpalmitate solid lipid nanoparticles. AB - Solid lipid nanoparticles (SLN) as alternative intravenous colloidal drug carriers were produced by high pressure homogenisation of the melted lipid cetylpalmitate. The crystallographic properties of the used cetylpalmitate SLN were characterised by small angle X-ray scattering (SAXS) and X-ray diffraction (XRD). It was found that the SLN are available exclusively in a crystalline form. Different cetylpalmitate formulations showed all the same patterns and a uniform crystal lattice was obtained. A partial indexing of the signals of the cetylpalmitate was carried out and the unit cell of the cetylpalmitate was estimated by the Miller indices. A preferred orientation in 001-direction was observed. This can be explained by an impressive lamellar lattice structure of the cetylpalmitate. The results were compared with the crystal data of cetylpalmitate known from literature. There was no correlation with the monoclinic structure known so far. This could indicate that the SLN consist of crystallites of another modification of the cetylpalmitate. PMID- 10699719 TI - Solubilization of timolol maleate in reversed micellar systems. Measurement of particle size using SAXS and PCS. AB - Small angle X-ray scattering (SAXS) and photon correlation spectroscopy (PCS) are two methods to measure particle sizes in the order of 10 nm magnitude, which can be used to characterize reversed micellar systems, in this case reverse micelles consisting of lecithin and isopropyl myristate (IPM). In this study these micelles are loaded with different concentrations of the amphiphilic anti glaucoma drug timolol maleate (TM). The PCS results are consistent with those yielded by SAXS, showing a decrease of particle size with higher TM concentration. In addition, SAXS is capable to give information about the particle shape. This kind of evaluation yields an ellipsoidal shape for micelles with low drug loads, which transform into nearly spherical micelles at higher drug concentrations. PMID- 10699720 TI - Influence of different parameters on reconstitution of lyophilized SLN. AB - Drug-loaded solid lipid nanoparticles (SLN) suitable for parenteral administration were freeze-dried. The lipid matrix Imwitor 900 (concentration, 2.5%) was stabilized with Lipoid E 80 and sodium glycocholate. The influence of different parameters of lyophilization like the protective effect of cryoprotectants, freezing velocity, and thermal treatment was investigated. The results of this study demonstrate that, by optimizing critical process parameters, i.v.-injectable SLN-dispersions can be freeze-dried, preserving their small particle size. PMID- 10699721 TI - Degradation of phosphatidylcholine in liposomes containing carboplatin in dependence on composition and storage conditions. AB - In this study, the hydrolytic degradation of phosphatidylcholine in aqueous liposome-dispersions and the stability of the anti cancer drug carboplatin enclosed in the liposomes were investigated in dependence on liposome composition and storage conditions. Cholesterol containing liposomes show a high stability of the phosphatidylcholine and the encapsulated carboplatin during six months storage in refrigerator. The hydrolytic degradation of phosphatidylcholine is strongly increased by addition of the antioxidant ascorbyl palmitate, but despite the partial hydrolysis the advantages of the lipid membrane are retained-- no degradation of the drug and no changes in the particle size were detected during six months storage in refrigerator, in contrast to storage at room temperature. PMID- 10699722 TI - Comparison of wax and glyceride solid lipid nanoparticles (SLN). AB - The present study compares solid lipid nanoparticles (SLN) formulated with either wax or glyceride bulk material. While most published data deal with glyceride SLN, little knowledge is reported on wax carriers. The two types were compared with respect to drug encapsulation efficacy, particle size distribution after production and storage, and crystal packing. The inclusion of retinol as a model drug was investigated. Retinol is chemically unstable in water and rather stable in lipid phases. Thus, rapid degradation of retinol indicates rapid drug expulsion from the carrier. Good stability indicates an effective drug encapsulation in the lipid phase of the nanoparticles. Particle size distribution was measured by laser diffractometry. Subcell packing and assignment of polymorphic forms was investigated by WAXS measurements. Glyceride SLN showed good drug encapsulation, while physical stability was poor. In contrast, wax SLN possessed good physical stability but lacked sufficient drug encapsulation in the solidified state. These differences were attributed in part to different crystal packing. Less ordered crystal lattices favour successful drug inclusion, as in the case of glyceryl monosterate and glyceryl behenate SLN. The highly ordered crystal packing of wax SLN comprised of beeswax or cetyl palmitate, for instance, leads to drug expulsion, but also to superior physical stability. PMID- 10699723 TI - Visualization and quantification of polymer distribution in microcapsules by confocal laser scanning microscopy (CLSM). AB - Confocal laser scanning microscopy (CLSM) was employed in order to characterize microcapsules. Microcapsules were prepared by complex coacervation: gelatin and arabic gum were labelled with fluorescent markers. In the capsule wall a homogeneous distribution for both gelatin and arabic gum throughout the capsule wall was depicted. By the use of CLSM and a computational image analysis the quantification of the labelled polymer in the wall material was possible. Adding fluorescently labelled casein as a macromolecular model compound to the coacervation process, a gradiental distribution in the wall material was observed with highest concentration of casein at the oil-wall interface. The influence of casein concentration on its deposition behaviour in the capsule wall was imaged successfully and thereafter quantified by computational image analysis. PMID- 10699724 TI - Investigation on the viscoelastic properties of lipid based colloidal drug carriers. AB - The rheological behaviour of solid lipid nanoparticle dispersions (SLN) prepared by high pressure homogenization was investigated using a Haake RS-100 rheometer. Four preparations differing in their lipid content and macroscopic consistency were tested by continuous shear rheometry and oscillatory testing. Rheological data from continuous shear measurement reveal plastic flow for systems with low lipid content as well as for systems with high lipid content. By using oscillatory testing more detailed information concerning the structure could be achieved. Rheological measurements of 40% lipid dispersions show viscoelastic properties comparable to the data from standard dermal preparations. Therefore high concentrated lipid dispersions might constitute a promising vehicle for topical administration. PMID- 10699725 TI - Interactions of nanoparticles with body proteins--improvement of 2D-PAGE-analysis by internal standard. AB - Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) is the method of choice to investigate protein adsorption of blood proteins (opsonization) onto nanoparticular drug carriers. In general the reproducibility of the obtained adsorption patterns is satisfying. However, direct comparison between the amounts of single protein spots from gels obtained in different runs is difficult, because 2D-PAGE is a multistep procedure. A possible solution of the problem is to establish a protein as internal standard. Therefore, selected proteins (Bio rad) were under investigation. Due to its molecular weight and isoelectric point, soybean trypsin inhibitor (TI) does not interfere with plasma components. Therefore, a method was established to use TI as an internal standard protein to improve comparability between the 2D-PAGE gels obtained in different analytical runs. PMID- 10699726 TI - Preserved solid lipid nanoparticles (SLN) at low concentrations do cause neither direct nor indirect cytotoxic effects in peritoneal macrophages. AB - In order to investigate the interaction of preserved solid lipid nanoparticles (SLN) with murine peritoneal macrophages (Mpsi), cytotoxicity and proinflammatory effects of two different solid lipid nanoparticles (SLN) preparations consisting of either compritol (CO) or cetyl palmitate (CP) preserved with thiomersal were analyzed. Concentration-dependent cytotoxic effects were observed using the 3 (4,5-dimethylthiazol-2-yl)-2, 5-diphenyl-tetrazolium bromide (MTT) assay. Secretion of interleukin-6 by Mpsi following incubation with CO and CP SLN did not differ from secretion by untreated cells; proinflammatory cytokines interleukin-12 and tumor-necrosis-factor-alpha as further indicators of immunomodulatory effects were not detectable. These findings paralleled our previous findings that unpreserved CO and CP SLN did not induce immunomodulatory effects but cytotoxicity at higher concentrations. There were no synergistic cytotoxic effects of preservative and SLN. Thus, preservation of SLN using thiomersal does not appear to cause increased cytotoxicity and immunomodulatory effects following incubation with Mpsi. PMID- 10699727 TI - Influence of spin probe structure on its distribution in SLN dispersions. AB - Solid lipid nanoparticles (SLN) are drug carrier system composed of biodegradable substances, which are solid at room temperature. The physico-chemical properties and structure of the incorporated compounds can affect their partitioning in SLN dispersions. In this work the influence of lipophilicity and structure of different SP on its location in SLN were studied. By electron paramagnetic resonance (EPR) measurements it was found that lipophilic SP distribute between a solid glyceride core and a soft phospholipid layer, with the more polar part (piperidine ring or methylcarboxylic groups) oriented toward the water-lipid interface. The majority of SP is located in the phospholipid layer, but the portion in the solid lipid core increases with SP lipophilicity. The hydrophilic Tempol does not incorporate into SLN. PMID- 10699728 TI - Nanoparticles with decreasing surface hydrophobicities: influence on plasma protein adsorption. AB - The rapid uptake of i.v. injected nanoparticles by cells of the mononuclear phagocytic system (MPS) is a major obstacle for a long blood circulation time and a drug targeting to sites other than the MPS. The adsorption of proteins on the particles surface after i.v. administration depends on their surface characteristics and is regarded as key factor for the in vivo organ distribution. The objective of this study is to investigate changes in the plasma protein adsorption patterns in the course of surface hydrophobicity variation. Latex particles with decreasing surface hydrophobicity were synthesized as model colloidal carriers. Physicochemical characterization had been performed and considerable differences in the protein adsorption patterns on the particles could be detected by using two-dimensional polyacrylamide gel electrophoresis (2 D PAGE). Correlations between physicochemical characteristics and the protein adsorption patterns have been found and are discussed. PMID- 10699729 TI - Nuclear gene targeting using negatively charged liposomes. AB - Oligonucleotides are a very useful tool to control gene activity. Oligos work by complementary base-pairing with target sequences either in the nucleus or in the cytosol (Zelphati, O., Szoka, F.C., Jr., 1996. Liposomes as a carrier for intracellular delivery of antisense oligonucleotides: a real or magic bullet? J. Contr. Rel. 41, 99-119). In a new approach using chimeric oligonucleotides (Yoon, K., Cole Strauss, A., Kmiec, E.B., 1996. Targeted gene correction of episomal DNA in mammalian cells mediated by a chimeric RNA-DNA oligonucleotide. Proc. Natl. Acad. Sci. USA 93, 2071-2076) conversion of single base mutations with help of intranuclear repair mechanisms maybe an advantageous method to cure genetic diseases which are based on single point mutations. These chimeric oligonucleotides are constructed in a way that they form an intramolecular double strand of DNA and modified RNA-bases. We used a fluorescent labelled pure 68-mer DNA-analogue of a chimeric oligonucleotides to follow the intracellular fate of these kind of genetic material. The oligos were complexed with protamine sulfate and coated with three different liposomal formulations. The AVE-3 formulation shows enhanced properties compared to a classical neutral and negatively charged formulation. Nuclear localisation of oligos could only be observed with the AVE-3 formulation. Furthermore only the negatively charged liposome formulations interact with the protamine-complexed oligonucleotides. PMID- 10699730 TI - Nanosuspensions for the formulation of aphidicolin to improve drug targeting effects against leishmania infected macrophages. AB - A series of labdans and their derivatives have been identified as novel potential antileishmanial drugs using an in vitro test system against extracellular promastigotes and intracellular amastigotes of Leishmania donovani in murine macrophages (Kayser, O., Kiderlen, A.F., 1998. In vitro activity of leishmanicidal labdanes and related compounds. Proceedings of the Ninth International Congress of Parasitology, Monduzi Editore, Bologna, 925-929). Of these compounds, aphidicolin, a tetradecanhydro-3,9-dihydroxy-4,11b-dimethyl-8, 11a-methano-11aH-cyclo-hepta[a]naphthalin-4,9-dimethanol+ ++ (Fig. 1), was shown to be highly active at concentrations in the microgram range (EC(50) = 0.16 microg/ml). To improve drug targeting effects aphidicolin was formulated as nanosuspension and retested for its enhanced activity (EC(50) = 0.003 microg/ml). PMID- 10699731 TI - Silica nanoparticles modified with aminosilanes as carriers for plasmid DNA. AB - We synthesised silica nanoparticles (SiNP) with covalently linked cationic surface modifications and demonstrated their ability to electrostatically bind, condense and protect plasmid DNA. These particles might be utilised as DNA carriers for gene delivery. All nanoparticles were sized between 10 and 100 nm and displayed surface charge potentials from +7 to +31 mV at pH 7.4. They were produced by modification of commercially available (IPAST) or in-house synthesised silica particles with either N-(2-aminoethyl)-3 aminopropyltrimethoxysilane or N-(6-aminohexyl)-3-aminopropyltrimethoxysilane. All particles formed complexes with pCMVbeta plasmid DNA as evidenced by ratio dependent retardation of DNA in the agarose gel and co-sedimentation of soluble DNA with nanoparticles. High salt and alkaline pH did inhibit complex formation. Absorption onto the particles also decreased the hydrodynamic dimensions of plasmid DNA as shown by photon correlation spectroscopy. Complexes formed in water at a w/w ratio of Si26H:DNA (pCMVbeta) of 300 were smallest with a mean hydrodynamic diameter of 83 nm. For effective condensation a w/w ratio of Si26H:DNA of 30 was sufficient. Further, the absorbed DNA was protected from enzymatic degradation by DNase I. PMID- 10699732 TI - Imaging of head and neck tumors--methods: CT, spiral-CT, multislice-spiral-CT. AB - Spiral-CT is standard for imaging neck tumors. In correspondence with other groups we routinely use spiral-CT with thin slices (3 mm), a pitch of 1.3-1.5 and an overlapping reconstruction increment (2-3 mm). In patients with dental fillings a short additional spiral parallel to the corpus of the mandible reduces artifacts behind the dental arches and improves the diagnostic value of CT. For the assessment of the base of the skull, the orbital floor, the palate and paranasal sinuses an additional examination in the coronal plane is helpful. Secondary coronal reconstructions of axial scans are helpful in the evaluation of the crossing of the midline by small tumors of the tongue base or palate. For an optimal vascular or tissue contrast a sufficient volume of contrast medium and a start delay greater than 70-80 s are necessary. In our opinion the best results can be achieved with a volume of 150 ml, a flow of 2.5 ml/s and a start delay of 80 s. Dynamic enhanced CT is only necessary in some special cases. There is clear indication for dynamic enhanced CT where a glomus tumor is suspected. Additional functional CT imaging during i-phonation and/or Valsalva's maneuver are of great importance to prove vocal cords mobility. Therefore, imaging during i-phonation is an elemental part of every thorough examination of the hypopharynx and larynx region. Multislice-spiral-CT allows almost isotropic imaging of the head and neck region and improves the assessment of tumor spread and lymph node metastases in arbitrary oblique planes. Thin structures (the base of the skull, the orbital floor, the hard palate) as well as the floor of the mouth can be evaluated sufficiently with multiplanar reformations. Usually, additional coronal scanning is not necessary with multislice-spiral-CT. Multislice-spiral-CT is especially advantageous in defining the critical relationships of tumor and lymph node metastases and for functional imaging of the hypopharynx and larynx not only in the transverse plane but also in the coronal plane. PMID- 10699733 TI - Medical volume exploration: gaining insights virtually. AB - Since modern imaging modalities deliver huge amounts of data, which cannot be assessed easily, the visualization techniques are utilized to emphasize the structures of interest. To compare them, the different visualization techniques (maximum intensity projection, multiplanar reformations, shaded surface display and volume rendering) are regressed to a common ground whereby their strengths and weaknesses can be revealed. Additionally, medical image analysis can detect anatomical objects in volumetric data sets and provides their descriptions for further use. Usually, segmentation plays a crucial roll in that process. There are many segmentation methods which can be categorized in boundary-based and content-based ones. The extraction of anatomical objects also allows their quantification. Image analysis and visualization do not squeeze more information out of a data volume, but they provide different ways to look at it. As in real life, this alone may enlarge the insight. PMID- 10699734 TI - 2D and 3D visualisation of head and neck tumours from spiral-CT data. AB - PURPOSE: This paper intends to demonstrate the possibilities of two- and three dimensional visualisation methods from spiral-CT data sets in the head and neck region and demonstrates their value based on various studies. MATERIAL AND METHODS: The scanner was a Somatom PLUS 4 (Siemens, Forchheim/Germany). The patients were scanned using a slice thickness of 3 mm and a table feed of 5 mm (pitch 1,6). The images were reconstructed with an increment of 1 mm. Contrast agent (150 ml) was applied intravenously with a flow of 2.5 ml/s and a start delay of 80 s. In one study the start delay was 20 s in order to visualise the carotid arteries (extracranial aneurysm). Volumetric data sets were postprocessed with 'Vitrea' and 'VoxelView' (Vital Images) volume rendering software on a Silicon Graphics O2 workstation (virtual laryngoscopy). Multiplanar reformation and colour-coded 3D-reconstruction were done on a Prominence workstation (Siemens/Forchheim/Germany). RESULTS: In every region of the head and neck MPRs are useful as additional planes (with the exception of the hypopharynx and the larynx), SSDs are always useful if there is extensive bony destruction (skull, spine, skeleton larynx). Colour-coded three-dimensional reformations may be done for extensive tumours and before multi-specialty surgery. Perspective volume rendering is already in use for virtual endoscopy of the paranasal sinuses and the virtual laryngoscopy. In temporal bone evaluation, perspective volume rendering is a new and promising method of the future to assess the inner ear. SUMMARY: Two- and three-dimensional displays are used to visualise pathological findings in their topographic relation to anatomical leading structures. Thus, the radiologist can point out to the clinician the pathological findings by some essential images without having to demonstrate all axial slices. PMID- 10699735 TI - Methods: MRT. AB - MRI has become the imaging method of choice in special regions of the head and neck (e.g. nasopharynx, oropharynx, oral cavity, floor of the mouth). Superconducting MR-equipment with field strengths of 1.0-1.5 T are appropriate for the evaluation of the head and neck region. Signal acquisition is optimal with circular polarized head coils or with specially designed surface coils; the body coil is insufficient.When imaging tumors we need T1 contrast, T2 contrast and contrast medium information (enhancement information). For the T1 contrast T1 spin-echo is and remains the best sequence. For T2-contast T2 turbo-spin-echo with fat suppression has replaced the T2 spin-echo sequences because it is faster and shows good contrast between tumor and saturated fat tissue. Fat saturated T1 turbo-spin-echo enables best tissue contrast after Gd-DTPA application. PMID- 10699736 TI - Paranasal sinuses and nasopharynx CT and MRI. AB - Neoplastic disease of the nose, paranasal sinuses, the nasopharynx and the parapharyngeal space requires thorough assessment of location and extent in order to plan appropriate treatment. CT allows the deep soft tissue planes to be evaluated and provides a complement to the physical examination. It is especially helpful in regions involving thin bony structures (paranasal sinuses, orbita); here CT performs better than MRI. MRI possesses many advantages over other imaging modalities caused by its excellent tissue contrast. In evaluating regions involving predominantly soft tissue structures (ec nasopharynx and parapharyngeal space) MRI is superior to CT. The possibility to obtain strictly consecutive volume data sets with spiral CT or 3D MRI offer excellent perspectives to visualize the data via 2D or 3D postprocessing. Because head and neck tumors reside in a complex area, having a 3D model of the anatomical features may assist in the delineation of pathology. Data sets may be transferred directly into computer systems and thus be used in computer assisted surgery. PMID- 10699737 TI - Oropharynx, oral cavity, floor of the mouth: CT and MRI. AB - Pretherapeutic staging of tumors of the oropharynx, the oral cavity and the floor of the mouth is important and should be thorough and exact to ensure appropriate therapy. Particularly important is the assessment of infiltration of deeper compartments and the topographic relationship of tumor to vascular structures (lingual artery and vein, hypoglossal nerve), or the presence of spread of the tumor across the midline. As spread of tumor may occur to a large degree underneath normal appearing mucosa, clinical assessment of the true tumor extent is difficult. In the last 20 years computed tomography (CT) has proved its value as a supplementary non-invasive method and established its role in modern diagnostic evaluation. Magnetic resonance imaging (MRI) is an non-invasive scanning method that offers excellent tissue contrast. Ultrasonography (US) is of secondary importance, but provides useful guidance due to its wide availability and its easy use. This paper aims to depict the possibilities of modern CT and MRI to provide 'one-stop-shopping' information to the clinician as a basis for the right therapeutic approach and correct estimation of the individual patient's prognosis. A clear problem oriented imaging strategy with standardized diagnostic criteria will lead to a cost effective evaluation. PMID- 10699738 TI - Neoplastic invasion of laryngeal cartilage: radiologic diagnosis and therapeutic implications. AB - Cross-sectional imaging plays an indispensable complementary role to endoscopy in the pretherapeutic workup of laryngeal and hypopharyngeal cancer. Both computed tomography and magnetic resonance imaging are suitable for the detection of neoplastic cartilage invasion. Although MRI, due to its high negative predictive value, is now generally considered to be the most suitable imaging method for pretherapeutic evaluation of cartilage invasion CT continues to be commonly performed in many centers for practical reasons. Recent studies have shown that CT may yield acceptable sensitivity for neoplastic invasion of laryngeal cartilage if the diagnostic criteria are selected and combined appropriately. False positive results are inevitable with both CT and MRI because reactive inflammation may lead to overestimation of neoplastic cartilage invasion. PMID- 10699739 TI - Lymph node metastases: CT and MRI. AB - Imaging is playing a major role in the assessment of cervical lymphadenopathy. In head and neck malignancies, imaging can be helpful for staging, and sometimes in differentiating different types of metastases, such as squamous cell carcinomas, non-hodgkins disease and thyroid carcinomas. This article on imaging of cervical lymph node metastases will describe both radiological and clinical aspects. Computed tomography (CT) and magnetic resonance (MR) are widely used for primary tumor and nodal imaging. However, very seldom these modalities have clinical consequences for the management of the neck, such as a wait-and-see policy if no nodes are depicted. This is caused by the limited accuracy of both modalities caused by the fallibility of radiologic criteria for metastases. Ultrasound (US) is hampered by similar morphologic criteria, and only US-guided fine needle aspiration cytology (FNAC) can offer additional cytologic criteria which are more reliable. PMID- 10699740 TI - Head and neck tumors: imaging recurrent tumor and post-therapeutic changes with CT and MRI. AB - OBJECTIVE: To evaluate criteria for detection of tumor recurrence and post treatment changes in patients with head and neck malignancies in computed tomography (CT) and magnetic resonance imaging (MRI). METHODS AND MATERIALS: Thirty-nine patients with head and neck carcinoma receiving radiochemotherapy were examined before, during and after therapy with MRI. Changes in signal intensity were correlated to histology or clinical course. Three hundred and thirty-one patients with head and neck malignancies were examined with CT after therapy. CT diagnoses were correlated with histology or clinical course. RESULTS: Main criteria for recurrent/residual tumor in MRI was infiltrative mass with high signal intensity in T2-weighted images and enhancement after Gd-DTPA in T1 weighted images. Radiation-induced changes led to false positive diagnosis in 46% in the interval up to 3 months after therapy and in 58% in the interval 3-6 months after therapy. The combination of a circumscribed, infiltrative mass with contrast enhancement in CT had a sensitivity of 86% and a specificity of 80%. CONCLUSION: CT could accurately demonstrate postoperative changes and tumor recurrence. MRI had advantages in differentiation of tumor and scar, but edema after radiation therapy can spoil diagnosis. PMID- 10699741 TI - Gallium-desferrioxamine protects the cat retina against injury after ischemia and reperfusion. AB - This study sought to determine whether gallium-desferrioxamine (Ga/DFO) can curb free radical formation and mitigate biochemical and electrophysiological parameters of injury in the cat retina subjected to ischemia followed by reperfusion. For the biochemical studies, cat eyes were subjected to 90 min of retinal ischemia followed by 5 min of reperfusion, and enucleation of one eye of each cat was used to measure retinal reperfusion injury. Before enucleation of fellow eyes, 2.5 mg/kg Ga/DFO was injected intravenously 5 min before reperfusion. The flux of hydroxyl radicals, as measured directly by conversion of salicylate to 2,3- and 2,5-dihydroxybenzoic acid (2,3- and 2,5-DHBA), was significantly lower in Ga/DFO-treated eyes. The mean normalized level of 2,3-DHBA (considered a specific marker of hydroxyl radicals) was 3.5 times higher in untreated eyes. Ga/DFO caused a significant reduction, by 2.56-fold, in lipid peroxidation, as reflected by levels of malondialdehyde. Ascorbic acid, a natural antioxidant present in the retina, is severely depleted in untreated eyes. In contrast, in Ga/DFO-treated eyes, levels were 10 times higher than the control. Energy charge was 2.38 times higher in treated eyes. Levels of purine catabolites (hypoxanthine, xanthine, and uric acid) that reflect excessive metabolism of purine nucleotides were approximately twice higher in untreated retinas. Electroretionographic studies, performed on a different subset of animals, substantiated the biochemical results. In Ga/DFO-treated eyes the amplitude of the mixed cone-rod response b-wave (as compared with fellow nonischemic eyes) fully recovered within 24 h after ischemia (b-wave ratio 1.04 +/- 0.09, [mean +/- SEM]) whereas ischemic/reperfused and nontreated eyes recovered to only 0.33 +/- 0. 05. The results show that severe biochemical and functional retinal injury occurs in cat eyes subjected to ischemia and reperfusion. These severe changes were significantly reduced by a single administration of Ga/DFO just before reperfusion. We hypothesize that the protection afforded by Ga/DFO is due to a combined effect of "Push-Pull" mechanisms interfering with transition metal dependent and free radical-mediated injurious processes. PMID- 10699742 TI - The effect of copper supplementation on red blood cell oxidizability and plasma antioxidants in middle-aged healthy volunteers. AB - A multicenter European study (FoodCue) was undertaken to provide data on the significance of increased dietary copper as a pro-oxidant or antioxidant in vivo. The present work describes the effect of Cu supplementation on (2,2'-azo-bis(2 amidinopropane) hydrochloride (AAPH)-induced red blood cell oxidation in middle aged people. Double-blinded copper supplementation was achieved in 26 healthy volunteers (50-70 years) with pills containing 3 mg CuSO(4), 3 mg Cu glycine chelate (CuG) and 6 mg CuG. Each 6 week supplementation period was preceded and followed by 6 weeks of washout (WO) on placebo. The results show significant increases in time necessary to achieve 50% hemolysis (LT(50)) after 3CuSO(4) and 6CuG compared with values after WO periods. Cu supplementation did not increase the levels of (Cu,Zn)SOD activity in red blood cells. Resistance to hemolysis was significantly and positively correlated (r =.30, p <.01) with alpha- and beta carotene content in the plasma. Together, these data suggest that intake of copper as high as 7 mg/d has no pro-oxidant activity and may rather result in protection of red blood cells against oxidation. The decreased oxidizability of red blood cells did not result from increased (Cu,Zn)SOD activity and may occur through other mechanisms such as changes in membrane antioxidant content. PMID- 10699743 TI - Cyto- and genotoxic effects of novel aromatic nitroxide radicals in vitro. AB - Because of the increasing interest in the use of nitroxide radicals as antioxidants and probes for various applications in biological systems, the question of their toxicity is of paramount importance. Cytotoxicity and mutagenicity studies have been extensively performed with the commercially available aliphatic nitroxides, and the general outcome is that these compounds are nonmutagenic and relatively noncytotoxic. In this study, the cytotoxicity and genotoxicity of a new class of aromatic nitroxides that we have synthesized (i.e., indolinonic and quinolinic nitroxides), whose antioxidant activity has been established in both chemical and biological systems, were evaluated and compared with those of two commercial nitroxides and with that of butylated hydroxytoluene (BHT). The mutagenicity assay was performed using Salmonella typhimurium tester strains TA98, TA100, and TA102, chosen on the basis of their ability to detect various types of mutations and their sensitivity to oxidative damage. None of the compounds tested were found to be mutagenic. The colony forming assay (CFA) using Chinese hamster ovary (CHO) AS52 cells was employed for determining the cytotoxicity of the test compounds. On comparing the effective dose that inhibits the CFA by 50% (IC(50)), most of the compounds tested on an equal molar concentration basis were less toxic than BHT. Therefore, the overall results obtained correlate well with the data reported in the literature on the toxicity of aliphatic nitroxides and lend support to the possible use of these compounds as therapeutic antioxidants. PMID- 10699744 TI - Activity of the antimutagenic enzyme 8-oxo-2'-deoxyguanosine 5'-triphosphate pyrophosphohydrolase (8-oxo-dGTPase) in cultured chinese hamster ovary cells: effects of cell cycle, proliferation rate, and population density. AB - Mammalian 8-oxo-2'-deoxyguanosine 5'-triphosphate pyrophosphohydrolases (8-oxo dGTPases), such as MTH1, are believed to play the same antimutagenic role as their bacterial homologues, like MutT. Both decompose promutagenic 8-oxo-dGTP, a product of active oxygen's attack on dGTP. It is not known how 8-oxo-dGTPase expression and function are regulated. Therefore, we investigated the effect of cell population density, proliferation rate, and cell cycle phase on 8-oxo dGTPase specific activity in cultured Chinese hamster ovary K1-BH4 (CHO) cells. With increasing cell population density (from 30 to 95% confluence), the activity of 8-oxo-dGTPase per milligram protein decreased by 33% (p =.007 by ANOVA) while cells shifted by 9% into the G(0)/G(1) phase, with a 5% drop in cells in S phase. Importantly, inhibition of the cells' proliferation rate by calf serum deprivation caused a more dramatic 23% shift toward the G(0)/G(1) phase and a 25% drop in S phase, but had no effect on 8-oxo-dGTPase activity. Likewise, no differences in the enzyme activity were observed within cell populations of different cell cycle phases separated by centrifugal elutriation. Thus, the present results exclude cell cycle-dependent regulation of 8-oxo-dGTPase activity in CHO cells or its simple dependence on proliferation rate. The observed decrease of 8-oxo-dGTPase activity with increasing cell population density might be related to augmentation of cell-to-cell contact. PMID- 10699745 TI - Aromatic hydroxylation in PBN spin trapping by hydroxyl radicals and cytochrome P 450. AB - Phenyl N-tert-butylnitrone (PBN) is widely used as a spin trapping agent, but is not useful detecting hydroxyl radicals because the resulting spin adduct is unstable. However, hydroxyl radicals could attack the phenyl ring to form stable phenolic products with no electron paramagnetic resonance signal, and this possibility was investigated in the present studies. When PBN was added to a Fenton reaction system composed of 25 mM H(2)O(2) and 0.1 mM FeSO(4), 4 hydroxyPBN was the primary product detected, and benzoic acid was a minor product. When the Fe(2+) concentration was increased to 1.0 mM, 4-hydroxyPBN concentrations increased dramatically, and smaller amounts of benzoic acid and 2 hydroxyPBN were also formed. Although PBN is extensively metabolized after administration to animals, its metabolites have not been identified. When PBN was incubated with rat liver microsomes and a reduced nicotinamide adenine dinculeotide phosphate (NADPH)-generating system, 4-hydroxyPBN was the only metabolite detected. When PBN was given to rats, both free and conjugated 4 hydroxyPBN were readily detected in liver extracts, bile, urine, and plasma. Because 4-hydroxyPBN is the major metabolite of PBN and circulates in body fluids, it may contribute to the pharmacological properties of PBN. But 4 hydroxyPBN formation cannot be used to demonstrate hydroxyl radical formation in vivo because of its enzymatic formation. PMID- 10699746 TI - Increased lipoprotein oxidation in Alzheimer's disease. AB - Oxidation has been proposed to be an important factor in the pathogenesis of Alzheimer's disease (AD) and amyloid beta is considered to induce oxidation. In biological fluids, including cerebrospinal fluid (CSF), amyloid beta is found complexed to lipoproteins. On the basis of these observations, we investigated the potential role of lipoprotein oxidation in the pathology of AD. Lipoprotein oxidizability was measured in vitro in CSF and plasma from 29 AD patients and found to be significantly increased in comparison to 29 nondemented controls. The levels of the hydrophilic antioxidant ascorbate were significantly lower in CSF and plasma from AD patients. In plasma, alpha-carotene was significantly lower in AD patients compared to controls while alpha-tocopherol levels were indistinguishable between patients and controls. In CSF, a nonsignificant trend to lower alpha-tocopherol levels among AD patients was found. Polyunsaturated fatty acids, the lipid substrate for oxidation, were significantly lower in the CSF of AD patients. Our findings suggest that (i) lipoprotein oxidation may be important in the development of AD and (ii) the in vitro measurement of lipid peroxidation in CSF might become a useful additional marker for diagnosis of AD. PMID- 10699747 TI - Induction of replicative senescence biomarkers by sublethal oxidative stresses in normal human fibroblast. AB - We tested the long-term effects of sublethal oxidative stresses on replicative senescence. WI-38 human diploid fibroblasts (HDFs) at early cumulative population doublings (CPDs) were exposed to five stresses with 30 microM tert butylhydroperoxide (t-BHP). After at least 2 d of recovery, the cells developed biomarkers of replicative senescence: loss of replicative potential, increase in senescence-associated beta-galactosidase activity, overexpression of p21(Waf 1/SDI-1/Cip1), and inability to hyperphosphorylate pRb. The level of mRNAs overexpressed in senescent WI-38 or IMR-90 HDFs increased after five stresses with 30 microM t-BHP or a single stress under 450 microM H(2)O(2). These corresponding genes include fibronectin, osteonectin, alpha1(I)-procollagen, apolipoprotein J, SM22, SS9, and GTP-alpha binding protein. The common 4977 bp mitochondrial DNA deletion was detected in WI-38 HDFs at late CPDs and at early CPDs after t-BHP stresses. In conclusion, sublethal oxidative stresses lead HDFs to a state close to replicative senescence. PMID- 10699748 TI - Different roles for nitrogen monoxide and peroxynitrite in lipid peroxidation induced by activated neutrophils. AB - We studied the roles of nitrogen monoxide (NO&z.rad;) and peroxynitrite produced by the polymorphonuclear leukocytes (PMNs) isolated from an inflammatory exudate. PMNs were incubated either in a medium with a submicromolar concentration of iron or in a diethylenetriaminepenta-acetic acid (DTPA)-containing medium, and stimulated with phorbol 12-myristate 13-acetate (PMA) to generate free radicals. In both conditions superoxide anion (O(2)(*)(-)), NO&z.rad; and peroxynitrite were produced. In the presence of arachidonic acid, malondialdehyde (MDA) was generated. This MDA was generated in one of two way; the peroxynitrite iron independent mechanism (40%) and the Fenton reaction, caused by free iron (60%). We also observed that the addition of L-arginine was followed by a 42% reduction in MDA, which can be explained by the antioxidant effect of NO&z.rad;. These results indicate that lipid peroxidation can occur in the absence of iron, through a peroxynitrite-mediated mechanism, and that NO&z.rad; may act as an antioxidant when it is produced in large amounts. PMID- 10699749 TI - Altered eicosanoid biosynthesis in selenium-deficient endothelial cells. AB - Selenium (Se) is an integral part of the Se-dependent glutathione peroxidase (Se GSH-Px) catalytic domain. By modulating the cellular levels of fatty acid hydroperoxides, Se-GSH-Px can influence key enzymes of arachidonic acid cascade, in this case cyclooxygenase (COX) and lipoxygenase (LOX). To investigate this phenomenon, the effects of cellular Se status on the enzymatic oxidation of arachidonic acid were investigated in bovine mammary endothelial cells (BMEC), which were cultured in either Se-deficient (-Se) or Se-adequate (+Se) media. When stimulated with calcium ionophore A23187, BMEC produced eicosanoids of both COX and LOX pathways. Compared with the Se-adequate cells, the production of prostaglandin I(2) (PGI(2)), prostaglandin F(2) (PGF(2alpha)), and prostaglandin E(2) (PGE(2)) was significantly decreased in Se-deficient cells, whereas the production of thromboxane A(2) (TXA(2)) was markedly increased in the -Se BMEC cultures. Although the enzymatic oxidation of arachidonic acid by the LOX pathway was found to be relatively less than by the COX pathway, the BMEC cultured in -Se media produced significantly more 15-hydroperoxyeicosatetraenoic acid (15-HPETE) than the +Se cells produced. Based on these results, we postulate that cellular Se status plays an important regulatory role in the enzymatic oxidation of arachidonic acid by the COX and LOX pathways. The altered eicosanoid biosynthesis, especially the overproduction of 15-HPETE, in -Se BMEC may be one of the underlying biochemical phenomena responsible for vascular dysfunction during Se deficiency. PMID- 10699750 TI - Ionizing radiation potentiates the induction of nitric oxide synthase by interferon-gamma (Ifn-gamma) or Ifn-gamma and lipopolysaccharide in bnl cl.2 murine embryonic liver cells: role of hydrogen peroxide. AB - The effects of ionizing irradiation on the nitric oxide (NO) production in murine embryonic liver cell line, BNL CL.2 cells, were investigated. Various doses (5-40 Gy) of radiation made BNL CL.2 cells responsive to interferon-gamma alone for the production of NO in a dose-dependent manner. Small amounts of lipopolysaccharide (LPS) or tumor necrosis factor-alpha (TNF-alpha) synergized with IFN-gamma in the production of NO from irradiated BNL CL.2 cells, even though LPS or TNF-alpha alone did not induce NO production from the same cells. Immunoblots showed parallel induction of inducible nitric oxide synthase (iNOS). NO production in irradiated BNL CL.2 cells by IFN-gamma or IFN-gamma plus LPS was decreased by the addition of catalase, suggesting that H(2)O(2) produced by ionizing irradiation primed the cells to trigger NO production in response to IFN-gamma or IFN-gamma plus LPS. Furthermore, the treatment of nongamma-irradiated BNL CL.2 cells with H(2)O(2) made the cells responsive to IFN-gamma or IFN-gamma plus LPS for the production of NO. This study shows that ionizing irradiation has the ability to induce iNOS gene expression in responsive to IFN-gamma via the formation of H(2)O(2) in BNL CL.2 murine embryonic liver cells. PMID- 10699751 TI - Antioxidant and oxidative status in tissues of manganese superoxide dismutase transgenic mice. AB - Manganese superoxide dismutase (Mn-SOD) plays an important role in attenuating free radical-induced oxidative damage. The purpose of this research was to determine if increased expression of Mn-SOD gene alters intracellular redox status. Twelve week old male B6C3 mice, engineered to express human Mn-SOD in multiple organs, and their nontransgenic littermates were assessed for oxidative stress and antioxidant status in heart, brain, lung, skeletal muscle, liver, and kidney. Relative to their nontransgenic littermates, transgenic mice had significantly (p <.01) higher activity of Mn-SOD in heart, skeletal muscle, lung, and brain. Copper, zinc (Cu,Zn)-SOD activity was significantly higher in kidney, whereas catalase activity was lower in brain and liver. The activities of selenium (Se)-GSH peroxidase and non-Se-GSH peroxidase, and levels of vitamin E, ascorbic acid and GSH were not significantly different in any tissues measured between Mn-SOD transgenic mice and their nontransgenic controls. The levels of malondialdehyde were significantly lower in the muscle and heart of Mn-SOD mice, and conjugated dienes and protein carbonyls were not altered in any tissues measured. The results obtained showed that expression of human SOD gene did not systematical alter antioxidant systems or adversely affect the redox state of the transgenic mice. The results also suggest that expression of human SOD gene confers protection against peroxidative damage to membrane lipids. PMID- 10699752 TI - 2-ethoxycarbonyl-2-methyl-3,4-dihydro-2H-pyrrole-1-oxide: evaluation of the spin trapping properties. AB - The 2-ethoxycarbonyl-2-methyl-3,4-dihydro-2H-pyrrole-l-oxide (EMPO), an easily prepared pyrroline-N-oxide has been tested as a free radical scavenger. Spin adducts of superoxide, hydroxyl radical, and other free radicals were characterized in phosphate buffer at pH 7.0 and 5.6. At pH 7 in phosphate buffer, the EMPO/O(2)(-*) spin adduct was estimated to be about five times more persistent than its DMPO analogue. Furthermore, its decay does not produce the EMPO/HO&z.rad; adduct. PMID- 10699753 TI - Dynamic state of S-nitrosothiols in human plasma and whole blood. AB - In the vasculature, nitrosothiols derived from the nitric oxide (NO)-mediated S nitrosation of thiols play an important role in the transport, storage, and metabolism of NO. The present study was designed to examine the reactions that promote the decomposition, formation, and distribution of extracellular nitrosothiols in the circulation. The disappearance of these species in plasma and whole blood was examined using a high-performance liquid chromatography method to separate low- and high-molecular weight nitrosothiols. We found that incubation of S-nitrosocysteine (CySNO) or S-nitrosoglutathione (GSNO) with human plasma resulted in a rapid decomposition of these nitrosothiols such that <10% of the initial concentration was recovered after 10-15 min. Neither metal chelators (DTPA, neocuproine), nor zinc chloride (glutathione peroxidase inhibitor), acivicin (gamma-glutamyl transpeptidase inhibitor), or allopurinol (xanthine oxidase inhibitor) inhibited the decomposition of GSNO. With both CySNO and GSNO virtually all NO was recovered as S-nitrosoalbumin (AlbSNO), suggesting the involvement of a direct transnitrosation reaction. Electrophilic attack of the albumin-associated thiols by reactive nitrogen oxides formed from the interaction of NO with O(2) was ruled out because one would have expected 50% yield of AlbSNO. Similar results were obtained in whole blood. The amount of S nitrosohemoglobin recovered in the presence of 10 microM GSNO or CySNO was less than 100 nM taking into consideration the detection limit of the assay used. Our results suggest that serum albumin may act as a sink for low-molecular-weight nitrosothiols and as a modulator of NO(+) transfer between the vascular wall and intraerythrocytic hemoglobin. PMID- 10699754 TI - Decreased thioredoxin and increased thioredoxin reductase levels in Alzheimer's disease brain. AB - Increasing evidence supports the role of reactive oxygen species (ROS) in the pathogenesis of Alzheimer's disease (AD). Both in vivo and in vitro studies demonstrate that thioredoxin (Trx) and thioredoxin reductase (TR), the enzyme responsible for reduction of oxidized Trx, have protective roles against cytotoxicity mediated by the generation of ROS. The present study measured levels of Trx protein and activities of TR in the brain in AD compared with control subjects, and evaluated the possible protective role of TR and Trx against amyloid beta-peptide (Abeta) toxicity in neuronal cultures. Analysis of Trx protein levels in 10 AD and 10 control subjects demonstrated a general decrease in all AD brain regions studied, with statistically significant decreases in the amygdala (p <.05), hippocampus/parahippocampal gyrus (p <.05), and marginally significant (p <.10) depletions in the superior and middle temporal gryi. Thioredoxin reductase activity levels were increased in all AD brain regions studied with statistically significant increases occurring in AD amygdala (p =.01) and cerebellum (p =.007). To investigate the protective effects of Trx and TR against Abeta-induced toxicity, primary hippocampal cultures were treated with Trx or TR in combination with toxic doses of Abeta. Treatment of cultures with Trx led to a statistically significant concentration-dependent enhancement in cell survival against Abeta-mediated toxicity as did treatment with TR. Together, these data suggest that, although TR is protective against Abeta-mediated toxicity, the increase observed in AD brain offers no protection due to the significant decrease in Trx levels. This decrease in the antioxidant Trx-TR system may contribute to the increased oxidative stress and subsequent neurodegeneration observed in the brain in AD. PMID- 10699755 TI - Vitamin C prevents the acute atherogenic effects of passive smoking. AB - During passive smoking the body is attacked by an excess of free radicals inducing oxidative stress. In nonsmoking subjects even a short period of passive smoking breaks down serum antioxidant defense (TRAP) and accelerates lipid peroxidation leading to accumulation of their low-density lipoprotein (LDL) cholesterol in cultured human macrophages. We now studied whether these acute proatherogenic effects of secondhand smoke could be prevented by an effective free radical scavenger, vitamin C. Blood samples were collected from nonsmoking subjects (n = 10) as they were consecutively exposed to normal air or cigarette smoke during four separate days. During the last 2 d, a single dose of vitamin C (3 g) was given, which doubled its plasma concentration. Vitamin C did not influence the plasma antioxidant defense or the resistance of LDL to oxidation in normal air, but prevented the smoke-induced decrease in plasma TRAP (p <.001), the decrease in the resistance of LDL to oxidation (p <.05), and the accelerated formation of serum thiobarbituric acid reactive substances (TBARS) (p <.05) otherwise observed 1.5 h after the beginning of passive smoking. Vitamin C protected nonsmoking subjects against the harmful effects of free radicals during exposure to secondhand smoke. PMID- 10699756 TI - Liposome-delivered superoxide dismutase prevents nitric oxide-dependent motor neuron death induced by trophic factor withdrawal. AB - Inhibition of nitric oxide synthesis prevents rat embryonic motor neurons from undergoing apoptosis when initially cultured without brain-derived neurotrophic factor. Using an improved cell culture medium, we found that the partial withdrawal of trophic support even weeks after motor neurons had differentiated into a mature phenotype still induced apoptosis through a process dependent upon nitric oxide. However, nitric oxide itself was not directly toxic to motor neurons. To investigate whether intracellular superoxide contributed to nitric oxide-dependent apoptosis, we developed a novel method using pH-sensitive liposomes to deliver Cu, Zn superoxide dismutase intracellularly into motor neurons. Intracellular superoxide dismutase prevented motor neuron apoptosis from trophic factor withdrawal, whereas empty liposomes, inactivated superoxide dismutase in liposomes or extracellular superoxide dismutase did not. Neither hydrogen peroxide nor nitrite added separately or in combination affected motor neuron survival. Our results suggest that a partial reduction in trophic support induced motor neuron apoptosis by a process requiring the endogenous production of both nitric oxide and superoxide, irrespective of the extent of motor neuron maturation in culture. PMID- 10699757 TI - Using MT(-/-) mice to study metallothionein and oxidative stress. AB - Mice with null mutations for metallothionein genes MT-1 and MT-2 were used to study the role that metallothionein plays in protecting cellular targets in vivo from oxidative stress. Wild-type (MT(+/+)) and MT-null (MT(-/-)) mice were treated with either saline or zinc and exposed to two types of oxidative stress: gamma-irradiation or 2-nitropropane. There was no alteration in the antioxidant defense system (superoxide dismutase, catalase, or glutathione peroxidase and glutathione levels) to compensate for the lack of the metallothionein in the MT( /-) mice. The amount of oxidative damage to liver DNA, lipids, and proteins were similar for the MT(-/-) and MT(+/+) mice even though the levels of metallothionein in the livers of the saline- or zinc-pretreated MT(+/+) mice were 5- to 100-fold greater than found in the MT(-/-) mice. To determine if metallothionein can protect mice from the lethal effects of ionizing radiation, the mean survivals of MT(-/-) and MT(+/+) mice exposed to whole body gamma irradiation were measured and found to be similar. However, the mean survival increased significantly after zinc pretreatment for both the MT(-/-) and MT(+/+) mice. These results demonstrate that tissue levels of metallothionein do not protect mice in vivo against oxidative stress. PMID- 10699758 TI - Oxidative stress and gene regulation. AB - Reactive oxygen species are produced by all aerobic cells and are widely believed to play a pivotal role in aging as well as a number of degenerative diseases. The consequences of the generation of oxidants in cells does not appear to be limited to promotion of deleterious effects. Alterations in oxidative metabolism have long been known to occur during differentiation and development. Experimental perturbations in cellular redox state have been shown to exert a strong impact on these processes. The discovery of specific genes and pathways affected by oxidants led to the hypothesis that reactive oxygen species serve as subcellular messengers in gene regulatory and signal transduction pathways. Additionally, antioxidants can activate numerous genes and pathways. The burgeoning growth in the number of pathways shown to be dependent on oxidation or antioxidation has accelerated during the last decade. In the discussion presented here, we provide a tabular summary of many of the redox effects on gene expression and signaling pathways that are currently known to exist. PMID- 10699759 TI - Common pathways for ultraviolet skin carcinogenesis in the repair and replication defective groups of xeroderma pigmentosum. AB - The human disease xeroderma pigmentosum (XP) involves DNA repair and replication deficiencies that predispose homozygous individuals to a 1000-fold increase in nonmelanoma and melanoma skin cancers. Two major forms of XP are known with different biochemical defects: one form lacks nucleotide excision repair (NER); the other lacks the capacity to replicate damaged DNA. Since the clinical symptoms of both kinds of patients are almost the same, the different cellular defects must be reconciled with common clinical outcomes. An additional question among the NER defective patients is how to reconcile widely different skin and central nervous system symptoms with mutations in the same biochemical pathway. XP involves seven genes of the NER system (XPA through G). The XPA gene codes for a protein that is central to NER and binds to a variety of UV light and chemical damage to DNA. It also acts as a nucleation center for other repair proteins to attach and carry out excision and replacement synthesis. Mutations in XPA that are within the DNA binding site produce more severe CNS disorders, than mutations in the C-terminal region of the protein that interacts with the TFIIH complex. In contrast, mutations in two members of the TFIIH complex, the XPB and XPD genes are generally very severe with both skin and CNS disorders. Missense mutations within the helicase regions of these genes are associated with DNA repair deficiencies and XPD; mutations elsewhere in these genes are correlated with symptoms of XP and Cockayne syndrome and trichothiodystrophy. This raises the question whether the CNS disorders of XPA, XPB, and XPD patients are similar, or whether a careful clinical evaluation might reveal different mechanisms of development. The XP variant lacks the capacity to replicate damaged DNA due to mutations in hRad30, a damage-specific polymerase eta. The phenotype of XP variant cells becomes unstable and the cells become much more UV-sensitive when they are transformed by methods that inactivate p53. On a p53 negative background, the induction of recombination between sister chromatids occurs much more extensively than in normal cells, and we have evidence that DNA double strand breaks which trigger an apoptotic pathway involving caspase-3 are involved. The pathway for UV carcinogenesis may be the same for all XP patients if the ultimate cause of genomic instability is an increase in replication of damaged DNA by the error-prone polymerase zeta. The presence of unrepaired damage in the NER defective groups of XP would present more substrate for the error prone system leading to increased mutation rates. The absence of pol eta would require cells to use the error-prone pol zeta pathway, also increasing mutation rates from UV damage. A common pathway for increased mutagenesis therefore underlies both forms of XP. PMID- 10699760 TI - Copper, zinc-superoxide dismutase protects from ultraviolet B-induced apoptosis of SV40-transformed human keratinocytes: the protection is associated with the increased levels of antioxidant enzymes. AB - It has been reported that cellular oxidative stress induces apoptosis. Ultraviolet radiation that generates reactive oxygen intermediates (ROIs) also induces apoptosis. Superoxide dismutase (SOD) is among the most active scavengers of ROIs, providing defense against the cellular oxidative stress. Mammalian cells express two isozymes of SOD, copper, zinc-SOD (Cu, Zn-SOD) and manganese-SOD (Mn SOD). Using SV40-transformed human keratinocytes (SVHK cells), we investigated the role of SODs in the ultraviolet B (UVB) irradiation-induced apoptosis. UVB irradiation decreased transiently Cu, Zn- and Mn-SOD activities and their protein levels, with subsequent recovery to the basal levels by 24 h. The UVB-induced decrease in SOD activity was dose-dependent and the maximal effect was obtained at 75 mJ/cm(2). The decrease in Cu, Zn-SOD was more marked than that in Mn-SOD. The cell death assay, annexin-V/propidium iodide flow cytometry, and DNA fragmentation analysis revealed that UVB irradiation induces apoptosis in SVHK cells. The UVB-induced apoptosis was suppressed by the treatment of antioxidants, catalase, glutathione, and alpha-tochopherol. The stable transfection of Cu, Zn SOD expression vectors into SVHK cells was accompanied by the increased activities of antioxidant enzymes, catalase, and glutathione reductase, as well as glutathione and the cells were shown to be more resistant to UVB-induced apoptosis. In contrast, the transfection of Mn-SOD affected neither activities of antioxidant enzymes nor the UVB-induced apoptosis. The transfection of Cu, Zn-SOD antisense oligomers but not sense oligomers into SVHK or Cu, Zn-SOD cDNA transfected SVHK (C2) cells significantly decreased the antioxidant enzyme activities and increased the UVB-induced apoptosis. On the other hand, the transfection of Mn-SOD antisense oligomers did not affect the UVB-induced apoptosis. These results suggest that the transfection of Cu, Zn-SOD expression vector, which is accompanied by the increased level of antioxidant enzymes, suppresses the UVB-induced apoptosis of SVHK cells. PMID- 10699761 TI - A preliminary report of the treatment of blue nevus with dermal injection of riboflavin and exposure to near-ultraviolet/visible radiation (ribophototherapy). AB - Dye lasers are useful for treating pigmented skin lesions, but their equipment is expensive and bulky. A simple and cheap phototherapy would be acceptable to dermatologists for treating pigmented skin lesions such as nevus of Ota. We investigated as a pilot study whether dermal injection of riboflavin and exposure to near-ultraviolet/visible radiation (ribophototherapy) decreases the dermal pigment of blue nevi which are recalcitrant to laser therapy. The therapeutic efficacy was assessed by comparison of the amount of dermal pigment in hematoxylin-eosin specimens taken before and after treatment. Pigmentation of the nevus became faint to the depth of 1 mm with little noticeable epidermal change after 21 treatments. At the deeper dermis somewhere between 3 and 4 mm from the epidermis, ballooning degeneration of the dermal cells was observed in hematoxylin-eosin specimens. Ribophototherapy is hopeful for treating pigmented skin lesions. PMID- 10699762 TI - Suppressive effect of zinc ion on iNOS expression induced by interferon-gamma or tumor necrosis factor-alpha in murine keratinocytes. AB - Zinc, an essential metal, is a critical component of zinc binding proteins such as zinc fingers, zinc enzymes and metallothioneins. Recently, evidence for its anti-inflammatory property in skin has been accumulating, as shown in the treatment of acne, alopecia and zinc deficiency. In cutaneous inflammations, a large amount of nitric oxide (NO) is produced through induction of inducible nitric oxide synthase (iNOS) under the influence of proinflammatory cytokines, resulting in tissue damages in skin, as clarified in other organs. Therefore, we asked if the effect of zinc on NO production and/or on iNOS expression in keratinocytes may explain the anti-inflammatory property of zinc in skin. Accordingly, we sought to determine in this study whether zinc ion may have effect on IFN-gamma or TNF-alpha induced NO production and iNOS expression in cultured murine keratinocytes. Ten microM of zinc ion remarkably suppressed cytokine-induced NO production in keratinocytes. Furthermore, zinc ion also suppressed cytokine-induced iNOS expression in the protein level as well as in the messenger RNA level. These results suggest the possibility that the suppressive effect of zinc ion on cytokine-induced NO production in keratinocytes may be in part implicated in the anti-inflammatory property of zinc in some of skin disorders. PMID- 10699763 TI - Precursor of rat epidermal cathepsin L: purification and immunohistochemical localization. AB - Immunoblot study using anti-rat cathepsin L antibody revealed almost only a precursor present in a crude extract of homogenized lower epidermis of rat skin, while the precursor and mature form were detected in the upper epidermis. To elucidate mechanisms of synthesis of cathepsin L, we have purified a precursor of cathepsin L from rat epidermis and investigated its localization in the skin. The precursor was purified after separation from the mature form in the final purification step of active fraction for N-benzyloxycarbonyl-L-phenylalanyl-L arginine-7-amido-4-methylcoumari n. The precursor showed a single protein band with Mr 39 kDa on SDS/polyacrylamide gel electrophoresis and was immunoreacted with the anti-rat cathepsin L antibody. Two types of NH(2)-terminal sequences obtained were identical to the amino acid residues from -96 to -86 and those from -93 to -87 deduced from cDNA of the precursor of cathepsin L. The precursor was processed to mature form of the enzyme and the enzyme activity remarkably increased after 48-h incubation of the whole epidermis in 1 M acetate buffer (pH 3. 5) at 20 degrees C. In histological sections of the skin, a thin and diffuse staining pattern was present in the spinous layer and a dense and linear staining in the granular layer of the epidermis. In contrast, rat liver showed more mature form than the precursor by immunohistological findings. These results suggest that cathepsin L may have some roles in the terminal differentiation. PMID- 10699764 TI - In vivo mapping of the vascular changes in skin burns of anaesthetised mice by fibre optic confocal imaging (FOCI). AB - Burns (3 mm in diameter, 50 degrees C, 20 s duration) were induced on the skin of anaesthetised hairless mice. Anaesthesia was maintained throughout all experiments. Subsurface changes in the microvasculature at the burn site were imaged confocally following i. v. injection of fluorescently labelled (FITC) dextran. Blood cells moving through dermal blood vessels were seen and recorded on video tape. Multiple adjacent 2-D confocal images of the burn site and surrounding areas were assembled and enabled microscopic vascular imaging of the whole burn area (including zones of coagulation, stasis and hyperaemia) and the surrounding normal vessels. This mapping of the burn area by fibre optic confocal imaging (FOCI) in vivo demonstrated good congruence with vascular casts (Microfil MV-120, Flow tech, USA) made at 4 h post burn. This study demonstrates the usefulness of FOCI for in vivo vascular imaging in burns. PMID- 10699765 TI - Cutaneous blood flow and adrenoceptor response increase in segmental-type vitiligo lesions. AB - It has been proposed that two types of vitiligo exist from the physiological and clinical points of view. Nonsegmental-type vitiligo is associated with autoimmune diseases while segmental-type vitiligo results from the dysfunction of sympathetic nerves in the affected area. Using laser Doppler flowmetry and iontophoresis for cutaneous microcirculatory assessments, we evaluated these two types of vitiligo in regard to their physiological changes. Ten patients with facial stable stage segmental-type vitiligo and ten stable nonsegmental-type vitiligo patients were selected for this study. Our results revealed that a nearly threefold increase in cutaneous blood flow was noticed in segmental-type vitiligo as compared to contralateral normal skin. In contrast, a 1.4-1.5 times difference was found among nonsegmental-type vitiligo, lesion side clinically normal skin and contralateral normal skin. There was a significant increase in cutaneous alpha- and beta-adrenoceptor response in segmental-type vitiligo lesions. However, no change in plasma catecholamines or adrenoceptor densities on blood cells was noticed. Our findings suggest that a dysfunction of the sympathetic nerves exists in the affected skin and plays a role in the pathogenesis of segmental-type vitiligo. PMID- 10699766 TI - Characterization of acyclovir susceptibility and genetic stability of varicella zoster viruses isolated during acyclovir therapy. AB - We have characterized the susceptibility and genetic stability of varicella zoster viruses (VZV) isolated from skin lesions of three patients with herpes zoster and six patients with varicella treated with conventional short-term acyclovir (ACV) administration. The susceptibilities to ACV of the serial isolates from the patients were examined, and there was no significant difference in the susceptibility to ACV among the isolates before and during the ACV treatment, indicating that conventional short-term ACV treatment did not generate ACV-resistant VZV infections. Polymerase chain reaction (PCR) analyses of these as well as seven thymidine kinase-deficient VZV strains derived from in vitro ACV treatment were carried out to examine their genomic stability. Five regions containing tandem direct reiterations (R1-R5) were amplified by PCR and compared, and the region containing the Pst I-site was also examined. PCR analyses demonstrated that the R1, R5 and the Pst I-sites were stable and useful in epidemiological studies even after ACV treatment. The R2, R3 and R4 sites were far less stable in these experimental conditions. In this paper we discuss the results of the PCR analyses with regard to the dynamics of VZV population in patients with VZV infection treated with conventional short-term ACV administration. PMID- 10699767 TI - A performance, safety and cost comparison of reusable and disposable endoscopic biopsy forceps: a prospective, randomized trial. AB - BACKGROUND: Many gastroenterologists believe that disposable forceps are more expensive than reusable forceps. It has been shown, however, that cross contamination and spread of infection are possible with reusable forceps. We conducted a prospective, randomized study to evaluate the performance, safety and cost of reusable versus disposable biopsy forceps. METHODS: Endoscopists were randomly assigned reusable or disposable biopsy forceps during upper and lower endoscopy. Forceps were evaluated for ease of passage through the endoscope, ease of opening and closing, adequacy of sample, and overall evaluation following the endoscopy using an ordinal scale. The cost per biopsy session was calculated using the following formula: (Acquisition cost + Reprocessing costs)/Number of biopsy sessions. RESULTS: Disposable forceps received a predominantly excellent rating versus a predominantly good rating for reusable forceps. Disposable forceps were also found to be more cost-effective than reusable forceps with an average savings of $5. 94 per biopsy session. Examination of reusable forceps revealed residual patient debris despite "adequate" cleansing. CONCLUSIONS: Disposable forceps outperformed reusable forceps and were found to be more cost effective. Residual patient debris on reusable forceps may pose a risk of cross contamination and the spread of infection. PMID- 10699768 TI - Disposable versus reusable biopsy forceps: a prospective cost evaluation. AB - BACKGROUND: There is growing advocacy for the use of disposable medical accessories to reduce the risks of infection transmission. Their purchase costs can, however, be considered as prohibitive in an endoscopy unit operating under a cost-containment program. We therefore compared the costs of reusable and disposable biopsy forceps. METHODS: From October 1995 to September 1997, biopsies were obtained in 7740 sessions. The evaluation of costs included purchase prices, repairs, cleaning (chemicals, equipment, technician time) and autoclaving costs in a centralized sterilization unit. For comparison, the lowest purchase price for disposable biopsy forceps was $26.90 in 1997. RESULTS: A mean of 12 new reusable forceps were purchased every year for a total purchase price of $5460. A total of 315 biopsy sessions were performed per forceps (mean time life of 3 years per forceps, including 3 repairs). Yearly repair cost was $3308, equipment $1002, chemicals $3250, central sterilization $8333, and technician salary $4373. Total cost was $25,726 and cost per biopsy session was $6.65. CONCLUSIONS: Total purchase and reprocessing costs for reusable biopsy forceps were 25% of those of disposable devices. The use of disposable biopsy forceps would have led to a yearly extra cost of $78,377 in the operation of our endoscopy unit. PMID- 10699769 TI - A cost and performance evaluation of disposable and reusable biopsy forceps in GI endoscopy. AB - BACKGROUND: Biopsy forceps are widely used in gastrointestinal endoscopy, and yet few data exist on the usage and costs associated with disposable versus reusable forceps. METHODS: We prospectively measured the costs and operational performance of disposable and reusable forceps in 200 biopsy sessions; 100 sessions were performed using disposable forceps and 100 sessions were performed using reusable forceps. Total cost per use of the reusable forceps, including acquisition costs plus the costs of reprocessing per established guidelines, was determined. At the end of the study, the reusable forceps were disassembled to determine the cause of mechanical failures. RESULTS: The total cost per use of the disposable forceps was $38. For the reusable forceps, the acquisition cost per forceps was $415 and the total reprocessing cost was $16.56 +/- 0.07 per forceps. For 10, 15 and 20 uses, reusable forceps costs were $58.06, $44.23, and $37.31, respectively. Reusable forceps malfunction at 11 to 15 uses was 5%; at 16 to 20 uses was 25%; and at 21 to 25 uses reached 80% (p < 0.001). Dismantling of the reusable forceps at the end of the study demonstrated coiled sheath kinking, rust in the forceps closure mechanism, bent spikes, and biomaterial contamination. CONCLUSIONS: Up to 15 to 20 uses, disposable and reusable forceps costs were similar. If reusable forceps are used more than 20 times, then they are less expensive. However, in this range of uses, reusable biopsy forceps performance diminishes. With disposable biopsy forceps costing less than $40, cost differences between reusable and disposable forceps are minimal. PMID- 10699770 TI - Nitrous oxide for colonoscopy: a randomized controlled study. AB - BACKGROUND: Intravenous sedation/analgesia for colonoscopy is accompanied with certain risks and postprocedure drowsiness. We sought to determine whether inhaled nitrous oxide (Entonox: 50% nitrous oxide, 50% oxygen) provides adequate analgesia for colonoscopy and the impact of this agent on recovery. METHODS: All patients undergoing outpatient colonoscopy were considered for the study (n = 248) except those with previous colonic resection. Data for patients unsuitable for randomization (n = 58) and those who declined to participate (n = 88) were also analyzed. RESULTS: One hundred two patients were randomized to receive inhaled Entonox alone (n = 56) or intravenous midazolam and meperidine (n = 46). Forty-nine (88%) patients randomized to Entonox underwent complete colonoscopy without conversion to intravenous medications. Entonox patients reported more pain (p < 0.0001), tolerated colonoscopy less well (p < 0.0001), were less satisfied (p = 0.01), and less willing to undergo colonoscopy again under the same circumstances (p = 0.04). Of patients receiving intravenous medication, 91% found colonoscopy less unpleasant and 9% as unpleasant as anticipated; this compares with 52% and 21% Entonox patients, respectively, and an additional 27% Entonox patients who found colonoscopy more unpleasant than anticipated. Recovery was faster among Entonox patients (median 30 versus 60 minutes, p < 0.0001). CONCLUSION: Entonox is less effective than midazolam with meperidine for colonoscopy but is acceptable in many patients and allows faster recovery. PMID- 10699771 TI - Patient-controlled analgesia for conscious sedation during colonoscopy: a randomized controlled study. AB - BACKGROUND: The aim of this study was to assess whether patient-controlled anesthesia (PCA) can improve patient tolerance for colonoscopy. We compared baseline sedation and analgesia with baseline sedation and PCA. METHODS: Fifty six consecutive patients were alternatively allocated to 1 of 2 groups: either to control group (n = 28) to receive standard sedation (meperidine and midazolam as baseline and additional doses of meperidine administered by the anesthesiologist) or to a PCA group (n = 28) to receive the same baseline premedication but additional analgesia with meperidine being self-administered. Cardiopulmonary parameters were recorded and tolerance for the examination was evaluated by a numeric rating scale, 0 meaning "no pain" and 10 meaning "maximal pain." RESULTS: Patients' mean pain score (on a scale of 0 to 10) was 4.85 +/- 3.74 for the PCA group and 5.30 +/- 3.53 (not significant) for the control group. Physicians' assessment of patient tolerance registered a lower numeric rating score than patients' assessment. The duration of the procedure was slightly longer in the PCA group. None of the patients experienced a decline in oxygen saturation below 90%; a decrease in expiratory carbon dioxide during the examination was noted in both groups of patients, particularly during the first minutes of the examination. Mean additional sedation per patient in the PCA group was slightly higher, but not significantly different. CONCLUSIONS: Our results suggest that patient-controlled analgesia during colonoscopy is as effective as standard sedation with respect to patient tolerance and safety of the examination. PMID- 10699772 TI - Pharmacoeconomic evaluation of flumazenil for routine outpatient EGD. AB - BACKGROUND: Flumazenil is a benzodiazepine antagonist indicated for reversal of the sedative effects of benzodiazepines. Previous studies suggest that flumazenil may shorten recovery time after endoscopy, but there are few data on actual recovery room times and charges. METHODS: Fifty patients undergoing routine upper endoscopy were sedated with midazolam alone in the usual titrated manner. Patients were randomized in a double-blind fashion to receive either flumazenil or saline immediately after procedure. Assessments of responsiveness, speech, facial expression, and ptosis (Observer's Assessment of Alertness/Sedation [OAA/S] scale) were made before procedure, immediately after procedure and every 15 minutes thereafter. The patient was discharged from the recovery room when vital signs and OAA/S scale reached preprocedure levels. Recovery room times and charges were recorded. RESULTS: The flumazenil group demonstrated shorter recovery room times and recovery room charges than the placebo group (p < 0.001). The difference in recovery room charges was not statistically different when flumazenil charges were included (p = 0.09). CONCLUSIONS: The routine use of flumazenil after midazolam sedation for upper endoscopy significantly shortened recovery time and charges but did not statistically reduce overall charges. PMID- 10699773 TI - A randomized, controlled trial of transcutaneous carbon dioxide monitoring during ERCP. AB - BACKGROUND: Pulse oximetry, used to monitor oxygen saturation during endoscopy, does not directly measure hypoventilation. Study goals were to determine whether transcutaneous carbon dioxide (PtcCO(2)) monitoring during endoscopic retrograde cholangiopancreatography (ERCP) prevents severe hypoventilation and to assess the accuracy of clinical observation and pulse oximetry in detecting hypoventilation. METHODS: All patients received intensive clinical and electronic monitoring including pulse oximetry. Supplemental oxygen was administered for pulse oximetry < 90%. Patients were randomized to a treatment arm (group 1) where PtcCO(2) monitoring guided sedation or a control arm (group 2) where PtcCO(2) was recorded but unavailable for guiding sedation. RESULTS: Group 1 had significantly fewer episodes of severe carbon dioxide retention (rise in PtcCO(2) >/=40 mm Hg above baseline) than group 2 (0 of 199 versus 5 of 196, respectively, p = 0.03), as well a shorter mean duration of procedure discomfort (8.3% of procedure duration rated as "uncomfortable" versus 11.5%, p = 0.04). Correlations between clinical observation and objective measures of ventilation were poor: level of sedation versus PtcCO(2) (R = 0.3) or pulse oximetry (R = 0.06); slowest respiratory rate versus PtcCO(2) (R = 0.4) or pulse oximetry (R = -0.4). PtcCO(2) rises of greater than 20 mm Hg occurred without oxygen desaturation in 10.7% of patients receiving supplemental oxygen. CONCLUSIONS: Carbon dioxide retention during ERCP is not reliably detected by clinical observation or by pulse oximetry in patients receiving supplemental oxygen. The addition of PtcCO(2) monitoring prevents severe carbon dioxide retention more effectively than intensive clinical monitoring and pulse oximetry alone. The clinical relevancy of this observation needs to be determined in an appropriately designed outcome study. PMID- 10699774 TI - In vitro evaluation of antibiotic prophylaxis in the prevention of biliary stent blockage. AB - BACKGROUND: Bacterial adherence and biofilm formation are important factors in the blockage of biliary stents. Clinical studies with oral antibiotic prophylaxis to prevent stent blockage have produced conflicting results. The aim of this study was to evaluate the in vitro effect of single antibiotic (ciprofloxacin, ceftazidime, or ampicillin) treatment on adherence of Escherichia coli and Enterococcus to plastic stents. METHODS: Selected clinical isolates of E coli and Enterococcus were perfused through a modified Robbins device containing segments of polyethylene stents. The stents were removed daily and the number of bacteria attached was measured. The effect of antibiotic treatment on bacterial adherence was tested by the perfusion of individual antibiotics into separate modified Robbins devices using a side-arm adaptor and the results were compared with saline controls. RESULTS: Compared with the saline controls, ciprofloxacin and ceftazidime caused a 10- to 100-fold reduction in the number of E coli attached to the stents, whereas ampicillin had no effect on adherence of E coli. Ampicillin caused a 5- to 10-fold reduction in Enterococcus adherence but there was no change with ceftazidime. Sustained reduction in E coli adherence was observed with prolonged ciprofloxacin perfusion. CONCLUSION: Timely treatment with appropriate antibiotics reduced bacterial adherence in vitro and may be potentially beneficial in the prevention of stent blockage. PMID- 10699775 TI - Small bowel polyps in Peutz-Jeghers syndrome: management by combined push enteroscopy and intraoperative enteroscopy. AB - BACKGROUND: Polyps occur throughout the GI tract in Peutz-Jeghers syndrome; the major problem in the management of the syndrome lies in the small bowel. METHODS: From January 1979 to January 1998, seven patients with Peutz-Jeghers syndrome underwent surveillance. Between 1979 and 1992 they were managed with upper and lower endoscopy every 2 to 3 years and surgery when intestinal obstruction occurred. From 1993 they also underwent enteroclysis and, on the basis of radiologic findings, push enteroscopy and/or intraoperative enteroscopy. Push enteroscopy was then performed every 2 years in all patients. RESULTS: During the first period, 5 of 7 patients underwent emergency small bowel resection (2 operated twice). The patients were divided into 2 groups based on enteroclysis findings; the first comprised 4 patients with multiple polyps throughout the small bowel, and the second included 3 patients with polyps only in the proximal small bowel. Three of the 4 patients with diffuse polyposis underwent intraoperative enteroscopy during which on average 16 polyps per patient were removed (range 10 to 25 polyps; mean diameter 16 mm, range 3 to 50 mm). The remaining patient with diffuse polyposis had a single 25 mm polyp in the terminal ileum removed by retrograde ileoscopy; the more proximal polyps were removed by push enteroscopy. The patients with diffuse polyposis remained asymptomatic during follow-up (mean 50 months, range 47 to 57 months) and also underwent periodic push enteroscopy (mean 2.25 enteroscopies per patient, range 2 to 3) at which a mean of 8.5 polyps per patient (range 4 to 13 polyps) were removed (mean diameter 7.2 mm, range 3 to 15 mm). The 3 patients of the second group underwent periodic push enteroscopy alone (mean 3 per patient) during which a mean of 11.7 polyps per patient were removed (range 7 to 15 polyps: mean diameter 10.9 mm, range 3 to 40 mm). Enteroclysis was not repeated in these patients, who remained asymptomatic during follow-up (mean 47 months, range 46 to 48 months). CONCLUSIONS: More effective clearance of small bowel polyps via enteroscopy will help reduce the need for emergency surgery with extensive intestinal resection in patients with Peutz-Jeghers syndrome. PMID- 10699776 TI - Dilation of malignant esophageal stenosis to allow EUS guided fine-needle aspiration: safety and effect on patient management. AB - BACKGROUND: Endoscopic ultrasonography (EUS) with fine-needle aspiration identifies patients with esophageal cancer who are unlikely to be cured by surgery. In approximately 30% of patients the staging procedure cannot be completed without dilation of an obstructing tumor. METHODS: All EUS examinations for esophageal cancer requiring dilation from July 1995 to December 1998 were included. Yield was defined as newly diagnosed metastatic (celiac lymph nodes) or locally invasive disease that could not have been detected without dilation. RESULTS: EUS was performed in 132 patients. Forty-two (32%) required 44 dilations. No complications occurred. Of the 42 patients with obstruction, 18 (43%) had celiac adenopathy of which 7 had malignant cells confirmed histologically, 3 had benign adenopathy, and 8 did not undergo fine-needle aspiration due to T4 stage disease (5) or intervening vessels (3). Two patients were upstaged after successful dilation from T2 N1 Mx to T4 N1 Mx and from T3 Nx Mx to T3 N1 M1. Overall, dilation allowed detection of advanced disease in 8 of 42 (19%) patients. Dilation to 11 to 12.8 mm was insufficient (36% success rate) to complete EUS compared with dilation to 14 to 16 mm (87%, p < 0.01). CONCLUSION: Dilation of obstructing esophageal tumors allows identification of a large number of patients with advanced stage malignancy. Dilation to 14 to 16 mm is sufficient for complete staging in almost all patients. PMID- 10699777 TI - Usefulness of a pediatric colonoscope for colonoscopy in adults. AB - BACKGROUND: There are few published data on how different types of colonoscopes affect success in reaching the cecum and patient comfort. We examined the feasibility of using a pediatric colonoscope for routine colonoscopy in adults and investigated whether there were subgroups of patients in whom use of this instrument was preferable. METHODS: One-hundred fifty adults undergoing outpatient colonoscopy were randomized to colonoscopy with a standard colonoscope (Olympus CF-100L) or with a pediatric colonoscope (Olympus PCF-100). All procedures were performed by a faculty endoscopist and timed by an independent observer. After examinations, the endoscopist graded procedure difficulty and patients were given a questionnaire that assessed their experience. RESULTS: The adult (n = 77) and pediatric (n = 73) colonoscope groups were comparable in all outcomes measured, including success in reaching the cecum (91% vs. 93%, p = 0.61), mean time to reach the cecum (11.4 vs. 9.7 min, p = 0.07), mean total procedure time (21.8 vs. 21.9 min, p = 0.95), mean meperidine dose (55 vs. 52 mg, p = 0.17); median midazolam dose (2.0 mg in both groups, p = 0.10), the endoscopists' perception of procedure difficulty, and patient comfort scales. Of the 7 patients in whom colonoscopy with the adult colonoscope was unsuccessful, the cecum was reached in 4 by switching to a pediatric colonoscope (all women, 3 of whom had prior hysterectomy). In the 5 patients in whom colonoscopy with the pediatric colonoscope was unsuccessful, the cecum was reached in 1 by switching to an adult colonoscope. Including the cases in which the cecum was reached by switching to the alternative colonoscope, the overall frequency of cecal intubation was 143 of 150 (95%). Subgroup analysis disclosed no difference between the 2 groups in outcomes when gender, presence of diverticulosis, and patient size were considered. Colonoscopy with the pediatric colonoscope was more successful than with the adult instrument in reaching the cecum in women with prior hysterectomy (11 of 12 [92%] vs. 15 of 21 [71%]); however, the numbers in each group were relatively small and the difference was not significant (p = 0.22). CONCLUSIONS: The pediatric colonoscope is suitable for routine colonoscopy in adults. It is also useful in patients in whom colonoscopy with the adult colonoscope is unsuccessful in reaching the cecum (particularly in women). Additional study is needed to see if the pediatric colonoscope is actually superior to the adult colonoscopy for routine colonoscopy in women with prior hysterectomy. PMID- 10699778 TI - The prevalence, anatomic distribution, and diagnosis of colonic causes of chronic diarrhea. AB - BACKGROUND: The prevalence of chronic diarrhea from a colonic disease and the optimal method of its diagnosis have not been ascertained. METHODS: Eight hundred nine patients with chronic non-bloody diarrhea unassociated with human immunodeficiency virus (HIV) infection underwent colonoscopy with biopsy specimen taken from throughout the colon and, if reached, the terminal ileum. The prevalence and anatomic distribution of ileocolonic histopathology and whether flexible sigmoidoscopy or colonoscopy represents the safest and most cost effective test for diagnosis were determined. RESULTS: 122 of 809 patients (15%) had colonic histopathology (microscopic colitis in 80 patients, Crohn's disease in 23, melanosis coli in 8, ulcerative colitis in 5, other forms of colitis in 5, and nodular lymphoid hyperplasia in 1). A correct assessment of colonic histology (normal or abnormal) could have been made from biopsies of the distal colon in 99.7% of patients. CONCLUSION: In a referral setting, colonic histopathology occurs in 15% of patients with chronic diarrhea without HIV infection. According to this prevalence and the nearly universal diffuse anatomic distribution of colonic disease in these patients, a diagnostic investigation for chronic colonic diarrhea using a 60 cm flexible sigmoidoscope is highly efficient and cost effective. PMID- 10699779 TI - Hematoma of the esophagus. PMID- 10699780 TI - Pharyngeal stromal tumor. PMID- 10699781 TI - Compact parakeratosis of esophageal mucosa. PMID- 10699782 TI - Extramedullary hematopoiesis in juvenile polyposis coli. PMID- 10699783 TI - Obliteration of esophageal varices using EUS-guided sclerotherapy with color Doppler. AB - BACKGROUND: The current standard treatment of bleeding esophageal varices is band ligation. Although endoscopic sclerotherapy has largely been supplanted by band ligation, there are still clinical situations in which injection methods are useful. Endoscopic ultrasound (EUS) may allow for a more complete evaluation of esophageal varices and perforating veins and may allow for more effective delivery of sclerosant. Our aim was to evaluate the use of color Doppler EUS guided sclerotherapy for the obliteration of esophageal varices. METHODS: Five patients with esophageal varices (Child's A = 1, B = 2, C = 2) underwent dynamic EUS-guided sclerotherapy with color flow Doppler. EUS sclerotherapy was performed using Varijet (2.5 mm catheter) injector needles and sodium morrhuate directed at the perforating vessels until flow was completely impeded (2 to 4 mL per injection site). Data collected included (1) sessions to obliteration, (2) episodes of recurrent bleeding, (3) complications, and (4) mortality. RESULTS: Patients undergoing EUS-sclerotherapy required 2.2 sessions to achieve obliteration of varices. No patient had a recurrence of bleeding and no deaths occurred. One patient developed an esophageal stricture that responded to balloon dilation. CONCLUSIONS: Dynamic EUS-guided sclerotherapy with color flow Doppler may be safely and effectively used for the treatment of esophageal varices. It allows for effective delivery of sclerosant with favorable outcomes. Prospective, multicenter, randomized trials are warranted. PMID- 10699784 TI - Endoscopic bilateral metal stent placement for malignant hilar stenoses: identification of optimal technique. AB - BACKGROUND: The aim of this study was to identify factors that facilitate bilateral insertion of metal stents in malignant hilar stenoses, for which plastic stents often result in incomplete drainage and subsequent cholangitis. METHODS: Between January 1994 and April 1998, we collected 45 cases of advanced (Bismuth stage II or higher) hilar malignant stenoses. The insertion technique was progressively modified and the success rate in the early period (1994 to 1995) was compared with that of a later period (1996) and the most recent period (1997 to 1998). RESULTS: Overall success rate was 73.3% (33 of 45). The success rates for the three periods were 50%, 67%, and 88% (p = 0.008), respectively. Cholangitis occurred in 3 of the patients with unilateral stents compared with 1 with bilateral stents. CONCLUSION: We have described a technique for endoscopic insertion of bilateral metallic stents for malignant hilar stenoses that results in high (>88%) and reproducible success rates. PMID- 10699786 TI - Long-term outcome of non-surgical strictureplasty using metallic stents for intestinal strictures in Crohn's disease. PMID- 10699785 TI - Endoscopic mucosal resection using a partial transparent hood for lesions located tangentially to the endoscope. AB - BACKGROUND: Numerous methods have been developed to resect early-stage gastric and esophageal cancers, but it is difficult to resect lesions viewed tangentially with the endoscope. METHODS: We have designed and developed an original method of endoscopic mucosal resection using a partial transparent hood to treat difficult cases in which the lesions are located tangentially to the endoscope. The hood was attached on the right side of the endoscope and, after insertion into the stomach or the esophagus, was lightly pressed on the orad side of the lesion. Then the lesion was resected using grasping forceps and electrosurgical current snare. RESULTS: The average diameter of specimens was 26 +/- 8 mm in gastric lesions and 20 +/- 3 mm in esophageal lesions, both 6 mm larger than those obtained by previous methods. CONCLUSION: This device and technique were extremely useful for mucosal resection of lesions located tangentially to the endoscope. PMID- 10699787 TI - Self-expanding covered esophageal ultraflex stent for palliation of malignant colorectal anastomotic obstruction complicated by multiple fistulas. PMID- 10699788 TI - Small cell lung carcinoma diagnosed on EGD. PMID- 10699789 TI - Severe gastric mucosa injury after percutaneous pure ethanol injection therapy for hepatocellular carcinoma. PMID- 10699790 TI - Primary rectal teratoma: EUS features and review of the literature. PMID- 10699791 TI - Intussusception in an adult after colonoscopy. PMID- 10699792 TI - Streptococcus milleri liver abscesses: an unusual complication after colonoscopic removal of an impacted fish bone. PMID- 10699793 TI - Intestinal Behcet's disease associated with generalized myositis. PMID- 10699794 TI - Pancreatic squamous carcinoma mimicking a bleeding duodenal ulcer. PMID- 10699795 TI - Reusable versus disposable forceps: the dilemma of cost and safety. PMID- 10699797 TI - Teaching basic endoscopy. PMID- 10699796 TI - Conscious sedation: is there a need for improvement? PMID- 10699798 TI - Women and transnasal endoscopy. PMID- 10699799 TI - Glutaraldehyde-based formulations. PMID- 10699800 TI - Colonoscopy: antispasmodics not only for premedication, but also during endoscope withdrawal? PMID- 10699801 TI - Peppercorn's disease. PMID- 10699802 TI - Postsurgical follow-up of children with tympanostomy tubes: results of the American Academy of Otolaryngology-Head and Neck Surgery Pediatric Otolaryngology Committee National Survey. AB - Postsurgical follow-up of children with tympanostomy tubes is becoming a contentious issue in this era of managed care. Primary care providers believe themselves to be capable of evaluating these children. Otolaryngologists, on the other hand, have more specialized equipment available to them (suction apparatus, otomicroscopes, audiology devices, etc) for treating suppurative infections and monitoring the tympanic membrane for structural changes. In addition, the otolaryngologist is placed in an uncomfortable legal and ethical position if access to the patient with a tube-related complication is denied by the primary care provider. Attempts to develop an American Academy of Otolaryngology-Head and Neck Surgery (AAO-HNS) policy statement have been hampered by a lack of data on the incidence and severity of tube-related complications and the role that otolaryngologists can play in reducing these sequelae. A survey designed by the AAO-HNS Pediatric Otolaryngology Committee was distributed to 1000 board certified otolaryngologists and all members of the American Society of Pediatric Otolaryngologists and the American Academy of Pediatrics-Otolaryngology Section regarding current practice patterns and practitioners' experiences with tympanostomy tube complications. Specific information regarding complications that could have been avoided with earlier otolaryngology referral was also obtained. The results of the survey and its implications for AAO-HNS policy are presented. PMID- 10699803 TI - Vestibular dysfunction in Gulf War syndrome. AB - METHODS: Vestibular complaints of Gulf War veterans were characterized by a nested case-control study of 23 veterans with 3 different Gulf War syndromes and 20 matched control subjects. All subjects completed a standardized symptom questionnaire and underwent standard audiovestibular tests administered by audiologists blinded to group identities. RESULTS: The prevalence of reported dizzy spells was higher in veterans with Gulf War syndromes 1 (100%), 2 (85%), and 3 (100%) than in controls (25%, P < 0.0001). Dizzy spells were more frequent, lasted longer, and involved a wider variety of accompanying symptoms in veterans with syndrome 2 than in those with syndromes 1 and 3. Audiovestibular testing showed greater interocular asymmetry of nystagmic velocity on sinusoidal harmonic acceleration in syndromes 1 (P = 0.015) and 2 (P = 0.002), greater asymmetry of saccadic velocity in syndrome 2 (P = 0.4), diminished nystagmic velocity after caloric stimulation bilaterally in syndrome 3 (P = 0.02 to 0.04), more subjects with pathologic nystagmus (P = 0. 09), and greater interside asymmetry of wave I to III interpeak latency on auditory brain stem response in syndromes 1 (P = 0.005) and 2 (P = 0.07). Asymmetry of gain on sinusoidal harmonic acceleration and pathologic nystagmus were most strongly associated with symptoms of paroxysmal vertigo (P = 0.002 and 0.07, respectively); asymmetry of saccadic velocity, with the severity of vertigo (P = 0.004); and abnormal caloric response, with chronic dysequilibrium (P = 0.006). CONCLUSIONS: The findings are compatible with a subtle neurologic injury from organophosphate-induced delayed neurotoxicity. PMID- 10699804 TI - Tympanostomy tubes and otic suspensions: do they reach the middle ear space? AB - The treatment of patients with tympanostomy tubes (TTs) and otorrhea with medicated otic suspensions is well known, but confirmation of penetration into the middle ear is difficult. To address this question, we created an in vitro model of the human head and ear and then tested it with 5 different types of liquid exposure: tap water, soapy water, polymyxin B sulfate (Cortisporin), tobramycin and dexamethasone (TobraDex), and ciprofloxacin (Cipro) suspensions. A positive test result corresponded to liquids entering the middle ear through the TT. No positive test result was elicited with tap water (0/20), but soapy water did enter the middle ear (10/40) and was statistically significant (P = 0.0112). Without the use of slight tragal pressure, Cortisporin, TobraDex, and Cipro drops did not consistently pass through the TT (0/20, 1/25, 1/25). By placing the drops with the addition of tragal pressure, a statistically significant difference was obtained for each solution (20/20, 20/20, and 20/20, respectively [P < 0.0001]). We conclude that with a clean external auditory canal, patent TT, and no middle ear fluid, medicated otic suspensions enter the middle ear only when combined with slight tragal pressure. PMID- 10699805 TI - Cost-effective management of benign positional vertigo using canalith repositioning. AB - OBJECTIVE: The misdiagnosis and inappropriate treatment of benign positional vertigo have resulted in significant costs to the medical system. In the current medical-economic climate, there is an increased emphasis on cost control. Recent studies have shown that the canalith repositioning procedure (CRP) is effective; the next step is to show the impact of CRP in cost-effective management of benign positional vertigo. METHODS: Forty-six of 100 patients who underwent CRP for benign positional vertigo responded to a survey regarding the financial impact of their disease. They were asked to subjectively estimate the sum of all disease related expenses. Objective substantiation of this number was estimated by tabulating physician data, laboratory data, and failed treatment costs. RESULTS: The subjective figure totaled $2684.74 per individual. Summation of the tangible objective figures yielded $2009.63 per patient, corroborating the subjective figure. CONCLUSIONS: Because CRP is a relatively simple procedure that can obviate many wasted expenses in most patients, we believe that it is very cost effective and should be incorporated into routine practice. PMID- 10699806 TI - Endolymphatic mastoid shunt: a reevaluation of efficacy. AB - OBJECTIVES: The main goal of this paper was to statistically reevaluate the efficacy of the endolymphatic shunt procedure for Meniere's disease. METHODS: Thomsen et al (Arch Otolaryngol 1981;107:271-7) reported on the placebo effect in surgery for Meniere's disease in a controlled double-blind study. Thirty patients with typical Meniere's disease in whom medical treatment failed participated in the study. A placebo-controlled blinded surgical study has not since been replicated. We performed a retrospective statistical analysis using data extracted from the published report and reanalyzed it using both the original and new statistical measures and techniques. RESULTS: The original conclusions drawn by Thomsen et al differed considerably from ours in 5 key areas, including postoperative vertigo, nausea and vomiting, tinnitus, and combined score. CONCLUSIONS: This analysis strongly supports the effectiveness of the endolymphatic shunt in the management of Meniere's disease and refutes the placebo effect previously proposed. PMID- 10699807 TI - Preapplication of mitomycin C for enhanced patency of myringotomy. AB - OBJECTIVES: Ventilation tubes are the mainstay of surgical treatment for eustachian tube dysfunction and have been used successfully for many years. Certain disadvantages of ventilation tubes, however, have prompted research into alternative techniques including laser myringotomy. We investigated the use of KTP laser myringotomy in conjunction with topical mitomycin C to delay healing and prolong the patency of the myringotomy. METHODS: Twenty myringotomies were created in 10 Sprague-Dawley rats. A solution of mitomycin C was applied to the intact tympanic membrane for 15 minutes. The solution was then suctioned free, and a myringotomy was created with a KTP laser. Fifty-three rats with saline application serving as controls from a previous study were used to allow statistical assessment. RESULTS: The myringotomies remained open for a median of 9.5 weeks. Control myringotomies, which received saline solution instead of mitomycin C, healed within a median of 1.5 weeks. The difference was statistically significant at P < 0.0001. No complications were noted. CONCLUSION: Topically administered mitomycin C before laser myringotomy is effective in prolonging the patency of laser myringotomies in rats. The patency rate is similar to that achieved in experiments in which topical mitomycin C is placed into the myringotomy site created by the laser. PMID- 10699808 TI - Conductive hearing loss associated with pressure equalization tubes. AB - Pressure equalization (PE) tube placement traditionally has been used to lessen conductive hearing loss with chronic middle ear effusion. It is assumed that the small diameter of the tube should not interfere with the conduction of sound. In this article we present 5 patients in whom placement of a PE tube resulted in significantly worse conductive hearing. Occlusion of the PE tube with cigarette paper or Gelfoam improved hearing, as documented with audiometry. The average conductive hearing losses attributable to the ventilation tube for this series of patients were 22, 17, 15, 13, 4, and 10 dB at the frequencies of 250, 500, 1000, 2000, 4000, and 8000 Hz, respectively. This amount of change attributable to PE tubes in this small selection of patients is much greater than would be commonly appreciated. We conclude that the opening in the tympanic membrane provided by PE tubes can potentially result in a significant conductive hearing loss. Discussion includes those conditions in which reduced hearing may be more likely to occur. PMID- 10699810 TI - Octreotide scintigraphy for the detection of paragangliomas. AB - Paragangliomas are neuroendocrine tumors located primarily in the head and neck region, but they can also occur at other sites. Confirming the preoperative diagnosis and detecting synchronous tumors may be difficult in some patients. Octreotide is a somatostatin analog that, when coupled to a radioisotope, produces a scintigraphic image of tumors expressing somatostatin type 2 receptors. Paragangliomas, like many neuroendocrine tumors, have been found to have a high density of somatostatin type 2 receptors on the cell surface. This study compared the results of preoperative octreotide scintigraphy with the histopathology in 21 patients who underwent surgery for presumed head and neck paragangliomas. Octreotide scan findings were positive in 16 patients with paragangliomas and negative in 3 patients with other pathology. One false positive and 1 false-negative result were obtained. Thus, this test had an accuracy of 90%, a sensitivity of 94%, and a specificity of 75%. Previously unidentified synchronous tumors were identified in 5 patients. On the basis of this series of patients, octreotide scintigraphy appears to be a reliable test to detect paragangliomas and may be quite helpful in preoperative planning. PMID- 10699809 TI - Tall cell variant: an aggressive form of papillary thyroid carcinoma. AB - Twenty-four cases of the tall cell variant (TCV), a subset of papillary thyroid carcinoma, were identified in a group of 624 patients with thyroid cancer. All pathology specimens were reviewed, and each patient's carcinoma was categorized according to characteristics on presentation, local recurrence, distant metastases, follow-up, and tumor-related mortality. The TCV group was compared with a historical control group (Mazzaferri and Jhiang: 1355 patients). The TCV group had a statistically higher percentage of stage 3 and 4 carcinoma, extrathyroidal invasion, and tumor size less than 1.5 cm than the control group. There was no statistical relationship between age greater than 50 years and stage in the TCV group. No relationship could be found between TCV histology and recurrence or mortality. These findings, combined with those of studies that link stage on presentation to poor outcomes, have led to our conclusion that TCV is an aggressive malignancy warranting appropriate treatment and close follow-up. PMID- 10699811 TI - Potential role of growth factors and extracellular matrix in wound healing after laryngotracheal reconstruction. AB - Laryngotracheal reconstruction (LTR) has been used for more than 20 years to treat infants and children with subglottic stenosis. Results after pediatric LTR have been satisfactory; however, approximately 10% of children have recurrent airway narrowing after LTR. The purpose of our study was to determine whether a correlation existed between specific growth factors and extracellular matrix in patients with adequate wound healing capability as compared with patients with poor wound healing capability. Histologic sections from 27 patients who underwent LTR were cut, and immunohistochemical staining was performed for transforming growth factor-beta, platelet-derived growth factor, fibronectin, tenascin, transforming growth factor-alpha, and vascular endothelial growth factor. Results showed that patients with adequate wound healing capability had a positive correlation with vasculature fibronectin, vasculature tenascin, and stromal fibronectin. Patients with poor wound healing capability had a positive correlation with stromal vascular endothelial growth factor. PMID- 10699812 TI - Approaches to the sellar and parasellar region: a retrospective comparison of the endonasal-transsphenoidal and sublabial-transsphenoidal approaches. AB - Tumors of the hypophysis are often managed surgically by the neurosurgeon and the otolaryngologist. Three widely used anterior routes to the sella are the endonasal (transcolumellar) transsphenoidal, sublabial transsphenoidal, and transethmoidal approaches. We reviewed the charts of 60 patients who underwent surgery, 42 transcolumellar and 18 sublabial, for sellar and parasellar adenomas and compared the two transsphenoidal approaches. None of the patients in our study underwent the transethmoidal approach. Furthermore, 26 of the patients underwent an extensive interview to assess postoperative progress. Clinically, neither approach had any significant complications, and none of the patients in either group reported significant postoperative morbidity. On the basis of these results, we believe there is minimal difference in patient subjective reports and objective morbidity when comparing the sublabial and transcolumellar approaches. PMID- 10699813 TI - Endoscopically guided sinonasal cultures: a direct comparison with maxillary sinus aspirate cultures. AB - Sinusitis is a common medical problem that can at times be challenging to treat. Although most cases respond to empiric therapy, success is not achieved universally. If empiric therapy fails, it is important to identify the causative bacterial pathogen. Antral puncture is the traditional diagnostic method to recover and identify pathogens in sinusitis; however, it remains a painful, invasive test with potential complications. In contrast, rigid sinonasal endoscopy permits recovery of mucopus emanating from the sinus ostia with little pain and few possible complications. Endoscopy also affords important visual information that can confirm or refute a historical/clinical diagnosis of sinusitis. Although previous studies have shown poor correlation between nasal cavity swab cultures and maxillary sinus aspiration cultures, few investigations have compared endoscopically guided middle meatal cultures with cultures obtained from maxillary sinus aspiration. Thirteen patients with maxillary sinusitis in one or both sinuses underwent endoscopically guided culture of the middle meatus and maxillary sinus puncture with aspiration and culture (16 total study samples). Results from the microbiologic analysis were compared. Endoscopically guided middle meatal cultures accurately identified the predominant bacterial pathogen and correlated with the cultures from maxillary sinus aspiration in more than 90% of infections. These preliminary results suggest that endoscopically guided sinonasal cultures hold promise as a viable alternative to maxillary sinus aspiration. Endoscopically guided cultures appear to be an effective, noninvasive diagnostic tool for otolaryngologists managing sinusitis. PMID- 10699814 TI - Circulating neutrophil CD18 receptor expression and ischemic flap neutrophil infiltration in a guinea pig model. AB - This article demonstrates a correlation between circulating neutrophil CD18 expression, neutrophil infiltration, and varying periods of ischemia induced in guinea pig island skin flaps. Fifty adult female Hartley guinea pigs were equally separated into a control group, a sham group, and ischemic groups of 2, 4, and 10 hours. All, except those in the control group, had single guinea pig island flank skin flaps raised. Systemic neutrophil surface receptor (CD18) expression was analyzed with monoclonal antibodies, and flap skin biopsy specimens were analyzed for neutrophil infiltration. The results show that neutrophil counts and receptor detection increase as flap ischemia increases. However, a trend toward declining receptor expression was observed in the 10-hour ischemic group. In conclusion, systemic neutrophil adhesion receptor upregulation is correlated with cutaneous flap neutrophil infiltration and ischemia-reperfusion injury in a guinea pig model. A trend toward declining receptor expression with advanced ischemia was observed. PMID- 10699815 TI - Expression of Ki-67, tumor suppressor proteins, growth factor, and growth factor receptor in juvenile respiratory papillomatosis: Ki-67 and p53 as predictors of aggressive disease. AB - The enhanced proliferation of epithelial cells is a typical feature of respiratory papilloma. The mechanism or mechanisms leading to abnormal epithelial proliferation remain unclear. Overexpression of growth factors and their receptors and inactivation of tumor-suppressor proteins are known to cause cell transformation and proliferation. The objectives of this study were to evaluate the expression of these factors in juvenile respiratory papillomas with correlation to cellular proliferation activity, and to determine whether such expression is associated with the clinical course of the disease. The expression of transforming growth factor-alpha, epidermal growth factor receptor, p53 protein, retinoblastoma proteins and Ki-67 was quantified by immunohistochemistry in paraffin-embedded biopsy specimens taken at the initial surgical excision from children in whom respiratory papillomatosis was diagnosed. Clinical information regarding the number of disease sites, tracheobronchial spread, malignant transformation, and frequency of recurrences was reviewed. Thirty-five specimens were suitable for immunohistochemical evaluation. Ki-67 expression was significantly higher in patients with multiple sites of disease and frequent recurrences. High p53 expression was significantly associated with malignant transformation. We concluded that Ki-67 and p53 expression may be predictive of the clinical course in children with respiratory papillomatosis. PMID- 10699816 TI - Midline radiofrequency tissue reduction of the palate for bothersome snoring and sleep-disordered breathing: A clinical trial. AB - This study is a prospective, nonrandomized clinical trial initiated to assess the safety and efficacy of radiofrequency tissue reduction of the palate for the treatment of bothersome snoring and sleep-disordered breathing. Twelve healthy volunteers with socially disruptive snoring underwent a baseline polysomnogram along with a battery of visual analog scales (VASs) to measure daytime sleepiness, snoring level, pain, and disturbances of speech and swallowing. After radiofrequency tissue reduction of the palate, they were re-evaluated with a mean follow-up after the final procedure of 15.7 +/- 5.1 (mean +/- SD) weeks. As rated by the bed partner, a significant reduction in the level of snoring occurred in all 12 patients, with a mean pretreatment snoring level of 8.3 +/- 2.1 to a mean posttreatment snoring level of 2.1 +/- 1.4. (Student t test, P < 0.0001) These patients required an average of 2.3 treatment sessions each. Nine of 12 had a reduction in snoring from a bothersome level (VAS range 5-10) to a nonbothersome level (VAS range 0-3). Daytime sleepiness as measured by the Epworth Sleepiness Scale (0-24) decreased from 10.8 +/- 4.4 to 8.3 +/- 4.1 (P = 0.011). Posttreatment pain was considered absent or minimal in 11 of 12 patients and was managed with acetaminophen. No significant adverse events or complications were reported. PMID- 10699817 TI - Pharyngeal suspension suture with repose bone screw for obstructive sleep apnea. AB - OBJECTIVE: Multilevel surgery for obstructive sleep apnea syndrome (OSA) may improve success. This study's goal is to prospectively evaluate the feasibility and short-term subjective effectiveness of a new tongue-suspension technique. METHODS: A multicenter nonrandomized open enrollment trial used the Repose device to treat tongue obstruction in 39 snoring and OSA patients. Outcomes include 1- and 2-month subjective reports of general health, snoring, and sleep. RESULTS: Twenty-three patients completed 1 month and 19 completed 2 months of follow-up. In OSA patients, activity level, energy/fatigue, and sleepiness improved. Two month outcomes were less (activity level, energy/fatigue, and sleepiness). Fewer changes were observed in snorers than in OSA patients. There were 6 complications (18%), including sialadenitis (4), gastrointestinal bleeding (1), and dehydration (1) after the procedure. CONCLUSION: A pharyngeal suspension suture changes subjective outcomes. Improvement is incomplete. The procedure is nonexcisional, but significant complications may occur. Further evaluation is required to demonstrate effectiveness. PMID- 10699818 TI - Comparison of postoperative pain between laser-assisted uvulopalatoplasty, uvulopalatopharyngoplasty, and radiofrequency volumetric tissue reduction of the palate. AB - OBJECTIVES: This study compares the posttreatment discomfort between laser assisted uvulopalatoplasty (LAUP), uvulopalatopharyngoplasty (UPPP), and radiofrequency volumetric tissue reduction (RFVTR) of the palate through the use of visual analog pain scales and a quantitative assessment of the analgesic medication taken. METHODS: In one group, LAUP (n = 10) or UPPP (n = 9) was used to treat patients' snoring or sleep-disordered breathing (SDB), and the other group underwent RFVTR (n = 22). RESULTS: The mean numbers of days with pain after RFVTR, LAUP, and UPPP were 2.6, 13.8, and 14.3 days, respectively. Narcotic analgesics were required in the RFVTR, LAUP, and UPPP groups in 9%, 100%, and 100% of the subjects, respectively. The mean number of these days requiring narcotic pain medications for RFVTR, LAUP, and UPPP was 0.2, 11.8, and 12.4 days, whereas the total narcotic equivalent was 0.3, 7.4 and 29.6 days, respectively. CONCLUSION: RFVTR of the soft palate produced less posttreatment pain than LAUP or UPPP. LAUP and UPPP appeared to show little difference in the severity or duration of posttreatment discomfort. PMID- 10699819 TI - Radiofrequency volumetric reduction of the palate: An extended follow-up study. AB - OBJECTIVE: The goal was to evaluate the effect of radiofrequency (RF) of the palate on speech, swallowing, taste, sleep, and snoring 12 to 18 months after treatment. METHODS: Twenty-two patients were evaluated by clinical examination, questionnaires, and visual analog scales. The patients with relapse of snoring were offered further RF treatment. RESULTS: After a mean follow-up of 14 months, no adverse effect was reported. Subjective snoring scores relapsed by 29% overall. Nine patients (41%) noted relapse of snoring from 2.1 +/- 1. 1 to 5.7 +/ 2.7 (P < 0.001). Eight of the patients underwent further RF treatment with a reduction of snoring from 5.8 +/- 2.9 to 3.3 +/- 3.1 (P = 0.01). CONCLUSION: The success of RF volumetric reduction of the palate diminishes with time, as with other surgical procedures of the palate. However, the minimal invasiveness of the RF provided a high patient acceptance for retreatment, and relapse of snoring can be improved. PMID- 10699820 TI - Surgery and obstructive sleep apnea: long-term clinical outcomes. AB - OBJECTIVE: Outcome data on the surgical treatment of obstructive sleep apnea are, in general, based on short-term follow-up (<6-9 months). This examination was undertaken to assess long-term results. METHODS: Forty patients who underwent soft tissue and skeletal surgery were the subjects of this review. Methods of evaluation included polysomnographic variables (respiratory disturbance index [RDI], low oxyhemoglobin desaturation [LSAT]), body mass index, quality-of-life assessments, roentgenographic analysis, and complications. Statistical analysis used the SAS 6.12 system. RESULTS: Thirty-six of 40 patients (90%) showed long term clinical success. The mean preoperative RDI, nasal continuous positive airway pressure RDI, and long-term RDI were 71.2 +/- 27.0, 7.6 +/- 5.2 and 7.6 +/ 5.1, respectively. The mean preoperative LSAT, nasal continuous positive airway pressure LSAT, and long-term LSAT were 67. 5% +/- 14.8%, 87.1% +/- 3.2%, and 86.3% +/- 3.9%, respectively. The mean follow-up was 50.7 +/- 31.9 months. The patients showed a statistically significant long-term weight gain (P = 0.0002) compared with their 6-month postoperative level (body mass index 31. 4 +/- 6.7 vs 32.2 +/- 6.3). There was a positive correlation with the amount of skeletal advancement and clinical outcome. CONCLUSION: Comprehensive evaluation and surgical treatment can result in successful long-term clinical outcome. PMID- 10699821 TI - Xenon-enhanced computed tomography quantifies normal maxillary sinus ventilation. AB - OBJECTIVE: The purpose of this study was to determine the applicability, safety, and normal parameters of a xenon-enhanced CT technique to quantify maxillary sinus ventilation. PATIENTS AND METHODS: Nine healthy subjects inhaled a xenon oxygen-air mixture through their noses while repeated CT scans were performed through the same section of their sinuses. Images were obtained every 1 to 3 minutes and analyzed to measure the density of the gas in the maxillary sinus as a function of time. RESULTS: Individual nasal cavity time constants ranged from 0.5 to 18 minutes. Studies performed after decongestion showed poorer sinus ventilation. CONCLUSIONS: The xenon-CT washin/washout technique is safe, effective, and gives representative data. PMID- 10699823 TI - Management of chyle fistulization in association with neck dissection. AB - Chylous fistula after neck dissection is a relatively rare but potentially lethal complication. Sequelae range from severe fluid, electrolyte, and protein loss to fistula formation, skin-flap necrosis, and carotid blowout. A thorough knowledge of the anatomy is essential to avoid injury to the thoracic duct or right lymph duct. After surgery, drainage of large amounts of fluid, particularly if milky, may alert the surgeon to the danger of chylous leakage. Certain diagnosis, however, is not so easy. Once the diagnosis is made, the management has to address the immediate and late effects of the loss of chyle into an operative site. This article seeks to examine these factors through review of the literature and personal experience with the problem. Total parenteral nutrition allows for control of the fluid and protein loss while avoiding flow of chyle, and in most cases it results in resolution. In those cases that do not resolve, fibrin glue with some type of mesh and muscle flaps usually succeed in closure. PMID- 10699822 TI - Effects of intracochlear factors on spiral ganglion cells and auditory brain stem response after long-term electrical stimulation in deafened kittens. AB - Using an animal model, we have studied the response of the auditory brain stem to cochlear implantation and the effect of intracochlear factors on this response. Neonatally, pharmacologically deafened cats (100 to more than 180 days old) were implanted with a 4-electrode array in both cochleas. Then, the left cochlea of each cat was electrically stimulated for total periods of up to 1000 hours. After a terminal (14)C-2-deoxyglucose (2DG) experiment, the fraction of the right inferior colliculus with a significant accumulation of 2DG label was calculated. Using 3-dimensional computer-aided reconstruction, we examined the cochleas of these animals for spiral ganglion cell (SGC) survival and intracochlear factors such as electrode positions, degeneration of the organ of Corti, and the degree of fibrosis of the scala tympani. The distribution of each parameter was calculated along the organ of Corti from the basal end. There was a positive correlation between SGC survival and the level of fibrosis in the scala tympani, and a negative correlation between SGC survival and the degree of organ of Corti degeneration. Finally, there was a negative correlation between the 2DG-labeled inferior colliculus volume fraction and the degree of fibrosis, particularly in the 1-mm region nearest the pair of electrodes, and presumably in the basal turn. PMID- 10699824 TI - Treatment of benign paroxysmal positional vertigo: no need for postmaneuver restrictions. AB - The liberatory maneuver of Semont is an effective physical treatment for benign paroxysmal positional vertigo. It works because it causes otoconia to move out the posterior canal. The effectiveness of the maneuver is thought to be indicated by the appearance of a liberatory nystagmus. After the maneuver, patients are usually instructed to keep their heads erect for several days and not to lie on the pathologic side for about a week. Here we investigated the prognostic value of liberatory nystagmus and whether restrictions are necessary after treatment. Fifty-six patients with posterior canal benign paroxysmal positional vertigo underwent the Semont maneuver and were checked after 20 minutes, 24 hours, and 1 week. The patients were told that they could sleep or move as they pleased, without any particular precautions. We found that liberatory nystagmus had a high prognostic value and that it was not necessary for patients to avoid certain positions or movements after treatment. PMID- 10699825 TI - Laser-assisted outpatient septoplasty: results in 703 patients. AB - Laser-assisted outpatient septoplasty is a new technique devised to minimize and simplify surgery under local anesthesia. It takes 5 minutes and has a specific clinical application in chronic nasal obstruction because of moderate anterior septal deviation in adults. It is less invasive than traditional septoplasty and has less morbidity, lower medical costs, and faster return to full activity. Seven hundred three patients underwent this operation from August 1995 to June 1998, with a patient evaluation performed before and after surgery. The evaluation was first performed by means of a direct interview, with a clinical examination and acoustic rhinometry, and then by means of a telephone interview, with strictly standardized questioning. Our results show a surgical success rate of 90.8% on the nasal obstruction but also an improvement on nasal discharge, sneezing, recurrent headaches, and chronic rhinosinusitis. PMID- 10699826 TI - Clinical experiences of removing foreign bodies in the airway and esophagus with a rigid endoscope: a series of 3217 cases from 1970 to 1996. AB - This study examined 11,333 rigid endoscopy procedures performed in the Department of Otolaryngology, National Taiwan University Hospital, during a 27-year period from 1970 to 1996. Among these cases, 3217 were performed to remove foreign bodies from the airway (459 cases, 14.3%) and esophagus (2758 cases, 85.7%). Retrospective analysis of these data revealed that peanuts (217 cases) and animal bones (1184 cases) were the most frequent foreign bodies encountered in the airway and esophagus, respectively. The successful rate of removal of these foreign bodies was 99.9% (3213/3217). The complication rate was only 0.2% (8/3217), and the mortality rate was less than 0.1% (2/3217). On the basis of these results, we conclude that foreign bodies in the airway and esophagus can be removed safely under direct visualization through rigid endoscopy with relatively few complications. A significant finding in this study is the declining trend in the number of cases in recent years. Despite the decline in the number of procedures, endoscopic removal of foreign bodies remains as a vital skill of the aerodigestive tract surgeon. PMID- 10699827 TI - Pleomorphic adenoma of the parotid gland metastasizing to the cervical lymph node. PMID- 10699828 TI - Bilateral sensorineural hearing loss and spastic paraparesis in Lyme disease. PMID- 10699829 TI - Isolated frontal sinus aspergillosis. PMID- 10699830 TI - Lipoma of the tongue. PMID- 10699831 TI - Trigeminal herpes zoster/chocolate-vanilla tongue. PMID- 10699832 TI - Metastatic renal cell carcinoma to the nose. PMID- 10699833 TI - Explaining the Weber test. PMID- 10699834 TI - Explaining the weber test PMID- 10699835 TI - Subcranial approach to tumors of the anterior cranial base. PMID- 10699836 TI - Subcranial approach to tumors of the anterior cranial base PMID- 10699837 TI - Food pollution. PMID- 10699838 TI - Food pollution PMID- 10699839 TI - Editorial PMID- 10699840 TI - A molecular simulation picture of DNA hydration around A- and B-DNA. AB - Recent results from molecular dynamics (MD) simulations on hydration of DNA with respect to conformation are reviewed and compared with experimental data. MD simulations of explicit solvent around DNA can now give a detailed model of DNA that not only matches well with the experimental data but provides additional insight beyond current experimental limitations. Such simulation results are analyzed with a focus on differential hydration properties between A- and B-DNA and between C/G and A/T base pairs. The extent of hydration is determined from the number of waters in the primary shell and compared to experimental numbers from different measurements. High-resolution hydration patterns around the whole DNA are shown and correlated with the conformations. The role of ions associating with DNA is discussed with respect to changes in the hydration structure correlating with DNA conformation. PMID- 10699841 TI - Water and monovalent ions in the minor groove of B-DNA oligonucleotides as seen by NMR. AB - During the past 8 years, two complementary nmr techniques-magnetic relaxation dispersion and nuclear Overhauser effect spectroscopy-have been applied extensively to the study of water and monovalent ions in the minor groove of B DNA oligonucleotides in solution. In this review, the possibilities and limitations of the two methods are outlined, with emphasis on the interpretational steps whereby molecular-level information is extracted from the primary data. The results on sequence-dependent hydration and ion-DNA interactions obtained so far by these methods is summarized and critically assessed. The nmr results are also compared with structural data from x-ray crystallography. PMID- 10699842 TI - X-ray crystallographic analysis of the hydration of A- and B-form DNA at atomic resolution. AB - We have determined single crystal structures of an A-DNA decamer and a B-DNA dodecamer at 0.83 and 0.95 A, respectively. The resolution of the former is the highest reported thus far for any right-handed nucleic acid duplex and the quality of the diffraction data allowed determination of the structure with direct methods. The structures reveal unprecedented details of DNA fine structure and hydration; in particular, we have reexamined the overall hydration of A- and B-form DNA, the distribution of water around phosphate groups, and features of the water structure that may underlie the B to A transition. PMID- 10699843 TI - Effect of hydrostatic pressure on nucleic acids. AB - In comparison to other biomolecules, the effect of hydrostatic pressure on the conformational stability of DNA and RNA has received scant attention. However, the increasing interest in the hydration of biological molecules has resulted in a concomitant increase in the number of investigations of the effect of pressure upon the structure of nucleic acids. In this review, studies concerning the effect of pressure on DNA and RNA oligomers and polymers are presented. The greatest amount of research has been directed at studying the effect of pressure on the stability of the double helix. In general, under most conditions, the helical form of DNA or RNA is stabilized by pressure. The extent of stabilization is small relative to the effect of pressure on other biomolecular systems such as lipid membranes or protein quaternary structure. The absence of a larger pressure effect arises, in part because the state of ionization does not change as a function of the helical state. Initial experiments have also appeared on the effect of pressure upon helix-formation kinetics, B-Z and A-Z equilibria, and DNA topology. Fourier-transform ir spectroscopy of DNA polymers under high pressure has yielded data that showing that pressure does not induce large-scale structural changes. PMID- 10699844 TI - Volumetric properties of nucleic acids. AB - Volumetric studies can yield useful new information on a myriad of intra- and intermolecular interactions that stabilize nucleic acid structures. In particular, appropriately designed volumetric measurements can characterize the conformation-dependent hydration properties of nucleic acids as a function of solution conditions, including temperature, pressure, ionic strength, pH, and cosolvent concentration. We have started to accumulate a substantial database on volumetric properties of DNA and RNA, as well as on related low molecular weight model compounds. This database already has provided unique insights into the molecular origins of various nucleic acid recognition processes, including helix to-coil and helix-to-helix conformational transitions, as well as drug-DNA interactions. In this article, we review recent progress in volumetric investigations of nucleic acids, emphasizing how these data can be used to gain insight into intra-and intermolecular interactions, including hydration properties. Throughout this review, we underscore the importance of volume and compressibility data for characterizing the hydration properties of nucleic acids and their constituents. We also describe how such volumetric data can be interpreted at the molecular level to yield a better understanding of the role that hydration can play in modulating the stability and recognition of nucleic acids. PMID- 10699845 TI - Fermentation process kinetics. Reprinted from Journal of Biochemical Microbiological Technology and Engineering VOl. 1, No. 4 Pages 413-29 (1959). AB - Information on fermentation process kinetics is potentially valuable for the improvement of batch process performance; it is essential for continuous process design. An empirical examination of rate patterns in various fermentations discloses three basic types: (1) 'growth associated' products arising directly from the energy metabolism of carbohydrates supplied, (2) indirect products of carbohydrate metabolism and (3) products apparently unrelated to carbohydrate oxidation. Effects of operating variables on the primary kinetic processes, growth, sugar utilization and antibiotic formation, in the penicillin process, illustrate the special nature of this type. PMID- 10699846 TI - A kinetic study of the lactic acid fermentation. Batch process at controlled pH. Reprinted from Journal of Biochemical and Microbiological Technology Engineering Vol. I, No. 4. Pages 393-412 (1959). AB - Kinetic data are needed to develop basic understanding of fermentation processes and to permit rational design of continuous fermentation processes. The kinetics of the fermentation of glucose to lactic acid have been studied at six constant pH levels between 4. 5 and 6.0 by measuring the instantaneous rates of bacterial growth and of lactic acid formation throughout each fermentation. It was found that the instantaneous rate of acid formation dP/dt, should be related to the instantaneous rate of bacterial growth dN/dt, and to the bacterial density N, throughout a fermentation at a given pH, by the expression when the constants alpha and beta are determined by the pH of the fermentation. PMID- 10699847 TI - Steam sterilizable probes for dissolved oxygen measurement. Reprinted from Biotechnology and Bioengineering, Vol. VI, Issue 4, Pages 457-468 (1964). AB - Steam-sterilizable membrane probes for monitoring the dissolved oxygen level in fermentors, or the oxygen content of gas streams, are described. The probes have a silver cathode, a lead anode, and an acetate buffer as an electrolyte. The membrane is Teflon. The current output of the probes in the absence of oxygen is negligible. PMID- 10699848 TI - The cultivation of animal cells at controlled dissolved oxygen partial pressure. Reprinted from Biotechnology and Bioengineering Vol. X, Issue 6, Pages 801-814 (1968). AB - A 3-liter culture vessel has been developed for the growth of animal cells in suspension at controlled pH and dissolved oxygen partial pressure (pO(2)). The culture technique allows metabolically produced CO(2) to be measured; provision can be made to control the dissolved CO(2) partial pressure. In cultures containing a low serum concentration, gas sparging to control pO(2) was found to cause cell damage. This could be prevented by increasing the serum concentration to 10%, or by adding 0.02% of the surface-active polymer Pluronic F68. The growth of mouse LS cells in batch culture without pO(2) control was found to be limited by the availability of oxygen. Maximum viable cell populations were obtained when dissolved pO(2) was controlled at values within the range 40-100 mm Hg. PMID- 10699849 TI - Kinetics of product inhibition in alcohol fermentation. Reprinted from Biotechnology and Bioengineering, Vol. X, Issue 6, Pages 845-864 (1968). AB - The inhibitory effect of ethanol concentration p in a medium on the specific rates of growth mu and ethanol production nu of a specific strain of baker's yeast was studied in a chemostat, where except for ethanol as the product, only the concentration of glucose S was controlled to limit the metabolic activity of the yeast. This was designed to supplement the previous findings from the batch experiment, in which ethanol was added artificially and no substrate components were limiting the metabolism of the same yeast, that mu = mu(0)e(-k(1)p) and nu = nu(0)e(-k(1)p), where k(1) and k(2) are empirical constants and subscript the 0 denotes respective values at p = 0. The effects of p on the values of mu and nu were confirmed by the Line-weaver-Burk plot to belong to noncompetitive inhibition. The formulas here for mu and nu as affected by p, if extrapolated to the case of no limiting substrates, were in good agreement in respective forms with those derived previously from the batch experiment, though the values of corresponding coefficients in these formulas were different. The differential equations for mu and nu as functions of both p and S and, in addition for the rate of glucose consumption as correlated by the yield factors either with the cell growth rate or the rate of ethanol production, were solved properly with a digital computer. A kinetic pattern calculated so far was discussed with reference to the data obtained in the batch experiment and those relevant to actual "sake" brewing. PMID- 10699850 TI - Measurement of heat evolution and correlation with oxygen consumption during microbial growth. Reprinted from Biotechnology and Bioengineering, Vol. XI, Issue 3, Pages 269-281 (1968). AB - A procedure for measuring the rate of heat production from a fermentation has been developed. The method is based on measuring the rate of temperature rise of the fermentation broth resulting from metabolism, when the temperature controller is turned off. The heat accumulation measured in this manner is then corrected for heat losses and gains. A sensitive thermistor is used to follow the temperature rise with time. This procedure is shown to be as accurate as previous methods but much simpler in execution. Using this technique, the rate of heat production during metabolism was found to correlate with the rate of oxygen consumption. Experiments were performed using bacteria (E. coli and B. subtilis), a yeast (C. intermedia), and a mold (A. niger). The substrates investigated included glucose, molasses, and soy bean meal. The proportionality constant for the correlation is independent of the growth rate, slightly dependent on the substrate, and possibly dependent on the type of organism growth. This correlation has considerable potential for predicting heat evolution from the metabolism of microorganisms on simple or complex substrates and providing quantitative parameters necessary for heat removal calculations. PMID- 10699851 TI - Adriamycin, 14-hydroxydaunomycin, a new antitumor antibiotic from S. peucetius var. caesius. Reprinted from Biotechnology and Bioengineering, Vol. XI, Issue 6, Pages 1101-1110 (1969). AB - Streptomyces peucetius var. caesius, obtained from S. peucetius, the daunomycin producing microorganism, by mutagenic treatment, differs from the parent culture by the color of the vegetative and aerial mycelia and by its antibiotic producing ability. S. peucetius var. caesius accumulates adriamycin in submerged and aerated culture on a medium containing glucose, brewer's yeast, and inorganic salts both in shake flasks and in stirred fermenters. Isolation of the product is performed by solvent extraction, chromatography on buffered cellulose columns, and crystallization as the hydrochloride. The new antitumor agent, adriamycin, is the 14-hydroxy derivative of daunomycin. PMID- 10699852 TI - The enzymatic transformation of water-insoluble reactants in nonaqueous solvents. Conversion of cholesterol to cholest-4-ene-3-one by a Nocardia sp. Reprinted from Biotechnology and Bioengineering, Vol. XVII, Pages 815-826 (1975). AB - The rapid conversion of cholesterol to cholestenone by Nocardia in the presence of high proportions of water-immiscible solvent has been demonstrated. At high agitator speeds, the reaction rate was not limited by the rates of transfer of oxygen or cholesterol to the microorganisms. Using 100 g of thawed cells in 200 ml of carbon tetrachloride containing 16% (w/v) cholesterol, at 20 degrees C cholestenone was formed at 7 g/hr. Cells could be separated easily from the organic solvent and reused. After 7 runs (69 hr) the reaction rate had fallen only to half the value for the first run. PMID- 10699853 TI - Formulation of structured growth models. Reprinted from Biotechnology and Bioengineering, Vol. XVIII, No. 10, Pages 1481-1486 (1976). PMID- 10699854 TI - The immobilization of microbial cells, subcellular organelles, and enzymes in calcium alginate gels. Reprinted from Biotechnology and Bioengineering, Vol. XIX, No. 3, Pages 387-397 (1977). AB - Saccharomyces cerevisiae cells, Kluyveromyces marxianus cells, inulase, glucose oxidase, chloroplasts, and mitochondria were immobilized in calcium alginate gels. Ethanol production from glucose solutions by an immobilized preparation of S. cerevisiae was demonstrated over a total of twenty-three days, and the half life of such a preparation was shown to be about ten days. Immobilized K. marxianus, inulase, and glucose oxidase preparations were used to demonstrate the porosity and retraining properties of calcium alginate gels. Calcium alginate immobilized chloroplasts were shown to perform the Hill reaction. Some experiments with immobilized mitochondria are reported. PMID- 10699855 TI - A new approach to preparative enzymatic synthesis. Reprinted from Biotechnology and Bioengineering, Vol. XIX, No. 9, Pages 1351-1361. AB - A new approach to preparative organic synthesis in aqueous-organic systems is suggested. It is based on the idea that the enzymatic process is carried out in a biphasic system "water-water-immiscible organic solvent." Thereby the enzyme is localized in the aqueous phase-this eliminates the traditional problem of stabilizing the enzymes against inactivation by a nonaqueous solvent. Hence, in contrast to the commonly used combinations "water-water-miscible organic solvent," in the suggested system the content of water may be infinitely low. This allows one to dramatically shift the equilibrium of the reactions forming water as a reaction product (synthesis of esters and amides, polymerization of amino acids, sugars and nucleotides, dehydration reactions, etc.) toward the products. The fact that the system consists of two phases provides another very important sources for an equilibrium shift, i.e., free energies of the transfer of a reagent from one phase to the other. Equations are derived describing the dependence of the equilibrium constant in a biphasic system on the ratio of the volumes of the aqueous and nonaqueous phases and the partition coefficients of the reagents between the phases. The approach has been experimentally verified with the synthesis of N-acetyl-L-tryptophan ethyl ester from the respective alcohol and acid. Porous glass was impregnated with aqueous buffer solution of chymotrypsin and suspended in chloroform containing N-acetyl-L-tryptophan and ethanol. In water (no organic phase) the yield of the ester is about 0.01%, whereas in this biphasic system it is practically 100%. The idea is applicable to a great number of preparative enzymatic reactions. PMID- 10699856 TI - Application of mass and energy balance regularities in fermentation. Reprinted from Biotechnology and Bioengineering, Vol. XX, No. 10, Pages 1595-1621 (1978). AB - Material and energy balances for fermentation processes are developed based on the facts that the heat of reaction per electron transferred to oxygen for a wide variety of organic molecules, the number of available electrons per carbon atom in biomass, and the weight fraction carbon in biomass are relatively constant. Mass-energy balance equations are developed which relate the biomass energetic yield coefficient to sets of variables which may be determined experimentally. Organic substrate consumption, biomass production, oxygen consumption, carbon dioxide production, heat evolution, and nitrogen consumption are considered as measured variables. Application of the balances using direct and indirect methods of yield coefficient estimation is illustrated using experimental results from the literature. Product formation is included in the balance equations and the effect of product formation on biomass yield estimates is examined. Application of mass-energy balances in the optimal operation of continuous single-cell protein production facilities is examined, and the variation of optimal operating conditions with changes in yield are illustrated for methanol as organic substrate. PMID- 10699857 TI - Technological problems in cultivation of plant cells at high density. Reprinted from Biotechnology and Bioengineering, Vol. XXII, No. 6, Pages 1203-1218 (1981). AB - Using Cudrania tricuspidata cells as model plant cells which have high sensitivity to hydrodynamic stress, technological problems in the cultivation of the plant cells at high density were investigated. Using "shake" flasks on a reciprocal shaker and Erlenmeyer flasks on a rotary shaker and with a high supply of oxygen in order to obtain high cell densities in shaken cultures, particle breakdown and damage to the largest cell aggregate group (above 1981 microm in diameter) occurred and normal cell growth became impeded. The mass-transfer coefficient (K) for a model solid-liquid system (beta-naphthol particles and water) in place of a system of plant cells and a liquid medium was proposed as an intensity index of hydrodynamic stress effects on plant cells in suspension cultures under various conditions in the bioreactor systems. Normal cell growth was obtained under culture conditions for K values less than about 4.4 x 10(-3) cm/sec. The characteristics of various bioreactors used until now were investigated by considering the three main technological factors (capacity of oxygen supply, intensity of hydrodynamic stress effects on plant cells, and intensity of culture broth mixing and air-bubble dispersion). The most suitable bioreactor for culturing plant cells at high density was a jar fermentor with a modified paddle-type impeller (J-M). The yield of cell mass in the 10-liter J-M (working volume 5 liter) was about 30 g dry weight per liter of medium. PMID- 10699858 TI - Continuous enzymatic transformation in an enzyme membrane reactor with simultaneous NAD(H) regeneration. Reprinted from Biotechnology and Bioengineering, Vol. XXIII, No. 12, Pages 2789-2802 (1981). AB - Multienzyme reaction systems with simultaneous coenzyme regeneration have been investigated in a continuously operated membrane reactor at bench scale. NAD(H) covalently bound to polyethylene glycol with a molecular weight of 10(4) [PEG 10,000-NAD(H)] was used as coenzyme. It could be retained in the membrane reactor together with the enzymes. L-leucine dehydrogenase (LEUDH) was used as catalyst for the reductive amination of alpha-ketoisocaproate (2-oxo-4-methylpentanoic acid) to L-leucine. Formate dehydrogenase (FDH) was used for the regeneration of NADH. Kinetic experiments were carried out to obtain data which could be used in a kinetic model in order to predict the performance of an enzyme membrane reactor for the continuous production of L-leucine. The kinetic constants V(max) and k(m) of the enzymes are all in the same range regardless of whether native NAD(H) or PEG-10,000-NAD(H) is used as coenzyme. L-leucine was produced continuously out of alpha-ketoisocaproate for 48 days; a maximal conversion of 99.7% was reached. The space-time yield was 324 mmol/L day (or 42.5 g/L day). PMID- 10699859 TI - Analysis of growth rate effects on productivity of recombinant Escherichia coli populations using molecular mechanism models. Reprinted from Biotechnology and Bioengineering, Vol. 26, Issue 1, Pages 66-73 (1984). AB - The influence of growth rate on Escherichia coli plasmid content and expression of a cloned-gene product has been described by a mathematical model based upon the molecular mechanism of lambdadv plasmid replication and known relationships between growth rate and transcription and translation activities of the host cell. The model simulates correctly decreases in plasmid content with increasing growth rate as observed experimentally for pBR322, NR1, R1, and Col E1 plasmids. A maximum with respect to growth rate in intracellular product accumulation is indicated by the model, as is a transient overshoot in product concentration following a shift from smaller to larger growth rate. Available data, although very limited, show the same trends. These results, obtained without parameter or kinetic form adjustments or manipulation, clearly illustrate the advantages of kinetic descriptions of recombinant systems based upon the pertinent molecular mechanisms. PMID- 10699860 TI - Equations and calculations for fermentations of butyric acid bacteria. Reprinted from Biotechnology and Bioengineering, Vol. XXVI, Pp 174-187 (1984). AB - A stoichiometric equation has been derived which describes the interrelations among the various products and biomass in fermentations of butyric acid bacteria. The derivation of the equation is based on an assumed ATP yield, two biological regularities, and the biochemistry of product formation of the fermentations. The equation obeys the constraints imposed on growth and product formation by thermodynamics and the biochemical topology. The validity of the equation is tested using a variety of fermentation data from the literature. The uses, improvements, limitations, and extensions of the equation are also discussed in detail. For example, the fermentation equation is used to calculate the maximal possible yields of the main fermentation products. PMID- 10699861 TI - Computer model for glucose-limited growth of a single cell of Escherichia coli B/r-A. Reprinted from Biotechnology and Bioengineering, Vol. 26, Issue 3, Pp 203 216 (1984). AB - A computer model is described which is capable of predicting changes in cell composition, cell size, cell shape, and the timing of chromosome synthesis in response to changes in external glucose limitation. The model is constructed primarily from information on unrestricted growth in glucose minimal medium. The ability of the model to make reasonable quantitative predictions under glucose limitation is a test of the plausibility of the basic biochemical mechanisms included in the model. Such a model should be of use in differentiating among competing hypotheses for biological mechanisms and in suggesting as yet unobserved phenomena. The last two points are illustrated with the testing of a mechanism for the control of the initiation of DNA synthesis and predictions on cell-width variations during the division cycle. PMID- 10699862 TI - Hydrodynamic effects on animal cells grown in microcarrier cultures. Reprinted from Biotechnology and Bioengineering, Vol. 29, Issue 1, Pages 130-141 (1987). AB - Hydrodynamic phenomena in microcarrier cultures are investigated with regard to the development of improved reactor designs for large-scale operations. New concepts and theoretical models that describe new data as well as previously published data are presented. PMID- 10699863 TI - A kinetic analysis of hybridoma growth and metabolism in batch and continuous suspension culture: effect of nutrient concentration, dilution rate, and pH. Reprinted from Biotechnology and Bioengineering, Vol. 32, Pp 947-965 (1988). AB - Hybridomas are finding increased use for the production of a wide variety of monoclonal antibodies. Understanding the roles of physiological and environmental factors on the growth and metabolism of mammalian cells is a prerequisite for the development of rational scale-up procedures. An SP2/0-derived mouse hybridoma has been employed in the present work as a model system for hybridoma suspension culture. In preliminary shake flask studies to determine the effect of glucose and glutaminE, it was found that the specific growth rate, the glucose and glutamine metabolic quotients, and the cumulative specific antibody production rate were independent of glucose concentration over the range commonly employed in cell cultures. Only the specific rate of glutamine uptake was found to depend on glutamine concentration. The cells were grown in continuous culture at constant pH and oxygen concentration at a variety of dilution rates. Specific substrate consumption rates and product formation rates were determined from the steady state concentrations. The specific glucose uptake rate deviated from the maintenance energy model(1) at low specific growth rates, probably due to changes in the metabolic pathways of the cells. Antibody production was not growth associated; and higher specific antibody production rates were obtained at lower specific growth rates. The effect of pH on the metabolic quotients was also determined. An optimum in viable cell concentration was obtained between pH 7.1 and 7.4. The viable cell number and viability decreased dramatically at pH 6.8. At pH 7.7 the viable cell concentration initially decreased, but then recovered to values typical of pH 7.1-7.4. Higher specific nutrient consumption rates were found at the extreme pH values; however, glucose consumption was inhibited at low pH. The pH history also influenced the behavior at a given pH. Higher antibody metabolic quotients were obtained at the extreme pH values. Together with the effect of specific growth rate, this suggests higher antibody production under environmental or nutritional stress. PMID- 10699865 TI - Looking good at forty. PMID- 10699864 TI - Metabolic flux distributions in Corynebacterium glutamicum during growth and lysine overproduction. Reprinted from Biotechnology and Bioengineering, Vol. 41, Pp 633-646 (1993). AB - The two main contributions of this article are the solidification of Corynebacterium glutamicum biochemistry guided by bioreaction network analysis, and the determination of basal metabolic flux distributions during growth and lysine synthesis. Employed methodology makes use of stoichiometrically based mass balances to determine flux distributions in the C. glutamicum metabolic network. Presented are a brief description of the methodology, a thorough literature review of glutamic acid bacteria biochemistry, and specific results obtained through a combination of fermentation studies and analysis-directed intracellular assays. The latter include the findings of the lack of activity of glyoxylate shunt, and that phosphoenolpyruvate carboxylase (PPC) is the only anaplerotic reaction expressed in C. glutamicum cultivated on glucose minimal media. Network simplifications afforded by the above findings facilitated the determination of metabolic flux distributions under a variety of culture conditions and led to the following conclusions. Both the pentose phosphate pathway and PPC support significant fluxes during growth and lysine overproduction, and that flux partitioning at the glucosa-6-phosphate branch point does not appear to limit lysine synthesis. PMID- 10699866 TI - Construction of multicomponent catalytic films based on avidin-biotin technology for the electroenzymatic oxidation of molecular hydrogen. AB - Two methods based on the avidin-biotin technology were developed for the multimonolayer immobilization of Desulfovibrio gigas hydrogenase on glassy carbon or gold electrodes. In both methods the molecular structure of the modified interface was the result of a step-by-step process. The first method alternates monolayers of avidin and biotinylated hydrogenase, the mediator (methyl viologen) being free to diffuse in the structure. In the second method, the avidin monolayers were used to immobilize both the biotinylated enzyme and a long-chain biotinylated viologen derivative. The viologen head of this hydrophilic arm shuttles the electrons between the electrode and the enzyme. The modified electrodes were evaluated for the electroenzymatic oxidation of molecular hydrogen, which has interest for the development of enzymatic fuel cells. The parameters that affect the current density of mediated oxidation of H(2) at the modified electrodes was studied. The second structure, which has given typical catalytic currents of 25 microA per cm(2) for 10 monolayers, was found clearly less efficient than the first structure (500 microA per cm(2) for 10 monolayers). In both methods the catalytic currents increased linearly with the number of monolayers of hydrogenase immobilized, which indicates that the multilayer structures are spatially ordered. PMID- 10699867 TI - Characterization study of the sporulation kinetics of Bacillus thuringiensis. AB - A wild-type and an rDNA strain of Bacillus thuringiensis were cultured in a net draft-tube modified 20-L airlift bioreactor. A comparison of the sporulation patterns suggests that the early sporulation strain has a lower final spore count. Results from off-gas analysis suggests that the CO(2) profile could be an alternative indication to spore counts for the examination of fermentation performance or even the mortality in bioassay of the cultivation product. The difference in mortality tests exhibited by the microorganism was attributed to different patterns of sporulation as well as different levels of gene control inside the cell itself. The sporulation kinetics of B. thuringiensis was simulated by a simple modified Hill equation, where the initial glucose concentration could affect the timing of the onset of sporulation. The equation matches well with the experimental sporulation data for B. thuringiensis in both wild-type and rDNA strains. PMID- 10699868 TI - Analysis and prediction of the effect of uncertain boundary values in modeling a metabolic pathway. AB - The integration of large quantities of biological information into mathematical models of cell metabolism provides a way for quantitatively evaluating the effect of parameter changes on simultaneous, coupled, and, often, counteracting processes. From a practical point of view, the validity of the model's predictions would critically depend on its quality. Among others, one of the critical steps that may compromise this quality is to decide which are the boundaries of the model. That is, we must decide which metabolites are assumed to be constants, and which fluxes are considered to be the inputs and outputs of the system. In this article, we analyze the effect of the experimental uncertainty on these variables on the system's characterization. Using a previously defined model of glucose fermentation in Saccharomyces cerevisiae, we characterize the effect of the uncertainty on some key variables commonly considered to be constants in many models of glucose metabolism, i.e., the intracellular pH and the pool of nucleotides. Without considering if this variability corresponds to a possible true physiological phenomenon, the goal of this article is to illustrate how this uncertainty may result in an important variability in the systemic responses predicted by the model. To characterize this variability, we analyze the utility and limitations of computing the sensitivities of logarithmic-gains (control coefficients) to the boundary parameters. With the exception of some special cases, our analysis shows that these sensitivities are good indicators of the dependence of the model systemic behavior on the parameters of interest. PMID- 10699869 TI - Influence of bcl-2 on cell death during the cultivation of a Chinese hamster ovary cell line expressing a chimeric antibody. AB - The influence of Bcl-2 expression on the robustness of a CHO cell line (22H11) developed for the industrial production of a chimeric antibody was evaluated. Western blot analysis following transfection with the expression vector unexpectedly revealed upregulation of endogenous Bcl-2 expression in the control (Neo) cell line in response to exposure to the selection drug G418. This indicated that geneticin may function by inducing apoptosis in cells not carrying the control plasmid or expressing very low levels of survival genes. Thus, exposure to the drug enriched the culture for a population of cells which expressed enhanced levels of endogenous Bcl-2. In batch cultures, ectopic bcl-2 expression resulted in a 75% increase in maximum viable cell density over control cultures. Moreover, the rate of decrease in viability in the Bcl-2 cultures was significantly lower than that in the control cultures. After 18 days, the Bcl-2 viability was around 90%, compared to 20% in the control cultures. Evaluation of the mechanism of cell death revealed very few cells with classical apoptotic morphology. Around 10% were clearly necrotic, but the majority of dead cells were seen as chromatin free but otherwise relatively intact structures. Because of the relatively low rate of cell death in both cell lines, few cells were observed in the transitional, easily identifiable early stages of apoptosis. However, DNA gel electrophoresis revealed a clear ladder-pattern, but only in the control cultures, thus confirming high levels of apoptotic death. Antibody concentrations during both sets of cultures were very similar, both during the growth and death phases, with a maximum titer of around 40 microgram/ml. Analysis of Bcl-2 expression by flow cytometry revealed that the cultures contained two populations of cells: a large population which expressed high levels of Bcl-2 and a relatively smaller low-expressing population. During the course of the batch, the smaller, low-expressing population declined in frequency, suggesting that these cells were more sensitive to cell death. In addition, the mean level of Bcl-2 expression in the overexpressing population also declined significantly, presumably reflecting the exhaustion of precursors for protein synthesis following nutrient depletion. Importantly, when cells were taken from day 40 of the significantly extended Bcl-2 batch cultures, they immediately proliferated, confirming that they had retained their replicative potential. Cultivation of the cells in basal medium lacking (individually) serum, all amino acids, glutamate/asparagine, and, finally, glucose, resulted in relatively lower viable cell numbers and viability in the control cell line compared to the Bcl-2 cell line. Exposure of cells to ammonia toxicity also revealed the relative robustness of the bcl-2 transfected cells. When growth was arrested by treatment with 4 mM thymidine, Bcl-2 overexpressing cells exhibit a viability of over 80% after 5 days in culture, compared to only 40% in the control cell line. However, under growth-arrested conditions, there was no major difference in antibody titer between the two cell lines. PMID- 10699870 TI - Efficient bioconversion of ethanol to acetaldehyde using a novel mutant strain of the methylotrophic yeast Hansenula polymorpha. AB - We report the isolation of mutant strains of the methylotrophic yeast Hansenula polymorpha that are able to efficiently oxidize ethanol to acetaldehyde in an intact cell system. The oxidation reaction is catalyzed by alcohol oxidase (AOX), a key enzyme in the methanol metabolic pathway that is typically present only in H. polymorpha cells growing on methanol. At least three mutations were introduced in the strains. Two of the mutations resulted in high levels of AOX in glucose grown cells of the yeast. The third mutation introduced a defect in the cell's normal ability to degrade AOX in response to ethanol, and thus stabilizing the enzyme in the presence of this substrate. Using these strains, conditions for bioconversion of ethanol to acetaldehyde were examined. In addition to pH and buffer concentration, we found that the yield of acetaldehyde was improved by the addition of the proteinase inhibitor phenylmethylsulfonyl fluoride (PMSF) and by permeabilization of the cells with digitonin. Under optimal shake-flask conditions using one of the H. polymorpha mutant strains, conversion of ethanol to acetaldehyde was nearly quantitative. PMID- 10699871 TI - Suitability of immobilized metal affinity chromatography for protein purification from canola. AB - This work demonstrates that proper selection of a metal ion and chelating ligand enables recovery of a his(6)-tagged protein from canola (Brassica napus) extracts by immobilized metal affinity chromatography (IMAC). When using Co(2+) with iminodiacetate (IDA) as the chelating ligand, beta-glucuronidase-his(6) (GUSH6) can be purified from canola protein extract with almost homogeneous purity in a single chromatographic step. The discrimination with which metal ions bound native canola proteins followed the order Cu(2+) < Ni(2+) < Zn(2+) < Co(2+) in regard to elimination of proteins coeluted with the fusion protein. IDA- and nitrilotriacetate (NTA)-immobilized metal ions showed different binding patterns, whose cause is attributed to a more rigid binding orientation of the his(6) in forming a tridentate with Me(2+)-IDA than in forming a bidentate with Me(2+)-NTA. The more flexible binding allows for multisite interactions over the protein. PMID- 10699872 TI - Cultivation of immortalized human hepatocytes HepZ on macroporous CultiSpher G microcarriers. AB - Cultivation of the new immortalized hepatocyte cell line HepZ was performed with a 1:1 mixture of DMEM and Ham's F12 media containing 5% FCS. The cells were grown in their 40th passage in 100 mL and 1 L volumes in spinner flasks and in a bioreactor, respectively. For the production of adherently growing HepZ cells macroporous CultiSpher G gelatin microcarriers were used in various concentrations from 1 to 3 g/L. The cells were seeded in a density of 2 x 10(5) cells/mL when using a microcarrier concentration of 1 g/L and 5 x 10(5) cells/mL at a microcarrier concentration of 3 g/L. After 7 days of cultivation a maximum cell concentration of 4.5 x 10(6) cells/mL was obtained in the spinner culture using a microcarrier concentration of 1 g/L. With bubble-free aeration and daily medium exchange from day 7, 7.1 x 10(6) cells/mL were achieved in the bioreactor using a microcarrier concentration of 3 g/L. The cells exhibited a maximum specific growth rate of 0.84 per day in the spinner system and 1.0 per day in the bioreactor, respectively. During the growth phase the lactate dehydrogenase (LDH) activity rose slightly up to values of 200 U/L. At the end of cultivation the macroporous carriers were completely filled with cells exhibiting a spherical morphology whereas the hepatocytes on the outer surface were flat-shaped. Concerning their metabolic activity the cells predominantly consumed glutamine and glucose. During the growth phase lactate was produced up to 19.3 mM in the spinner culture and up to 9.1 mM in the bioreactor. Maximal oxygen consumption was 1950 nmol/(10(6) cells. day). HepZ cells resisted a 4-day long chilling period at 9.5 degrees C. The cytochrome P450 system was challenged with a pulse of 7 microgram/mL lidocaine at a cell density of 4.5 x 10(6) cells/mL. Five ng/mL monoethylglycinexylidide (MEGX) was generated within 1 day without phenobarbital induction compared to 26 ng/mL after a preceded three day induction period with 50 microgram/mL of phenobarbital indicating hepatic potency. Thus, the new immortalized HepZ cell line, exhibiting primary metabolic functions and appropriate for a mass cell cultivation, suggests its application for a bioartificial liver support system. PMID- 10699873 TI - Kinetics of cassava starch hydrolysis with Termamyl(R) enzyme. AB - Kinetic properties of Termamyl(R) 120L were investigated with respect to starch hydrolysis in a batch reactor at substrate concentrations of 50-270 g. dm(-3) and enzyme concentrations of 0. 17-1 cm(3). dm(-3) (i.e., cm(3) of Novo Nordisk enzymatic solution per dm(3) of raw cassava starch suspension). A general kinetic expression giving product concentration as a function of time was first developed; an equation relating the reaction rate of each product to the sum of the concentrations of oligosaccharides with a higher degree of polymerisation was then derived. The empirical model satisfactorily fits experimental data for oligosaccharides with a degree of polymerization ranging from 1 to 7. PMID- 10699874 TI - Enzymatic resolution of (S)-(+)-naproxen in a trapped aqueous-organic solvent biphase continuous reactor. AB - A trapped aqueous-organic biphase system for the continuous production of (S)-(+) 2-(6-methoxy-2-naphthyl) propionic acid (Naproxen) has been developed. The process consists of a stereoselective hydrolysis of the racemic Naproxen methyl ester by Candida rugosa lipase in a trapped aqueous-organic biphase system. The reaction has been carried out in a laboratory-scale continuous-flow stirred tank reactor (CSTR). The staring material has been supplied in and remaining substrate recovered by organic phase. YWG-C(6)H(5), a poorly polar synthetic support, has been employed to immobilize the lipase and to restrict the aqueous phase. Lipase immobilized on YWG-C(6)H(5) containing aqueous phase has been added into the CSTR to catalyze the hydrolysis. A dialysis membrane tube containing a continuous flow closed-loop buffer has been applied in the CSTR for the extraction of product and recruiting of the aqueous part consumed. Various reaction conditions have been studied. The activity of immobilized enzyme was effected by the polarity of support, the substrate concentration, logP value of organic phase and the product inhibition. At steady-state operating conditions, an initial conversion of 35% has been obtained. The CSTR was allowed to operate continuously for 60 days at 30 degrees C with a 30% loss of activity. The hydrolysis reaction yielded (S)-(+) Naproxen with >90% enantiomeric excess and overall conversion of 30%. PMID- 10699875 TI - Lipase and esterase-catalyzed acylation of hetero-substituted nitrogen nucleophiles in water and organic solvents. AB - The lipase- and esterase-catalyzed acylations of hydroxylamine and hydrazine derivatives with octanoic acid and ethyl octanoate are described. The influence of solvent and nucleophile on the initial reaction rate was investigated for a number of free and immobilized enzymes. Initial rates were highest in water, but the overall productivity was optimal in dioxane. Octanoic acid (250 g/L) was converted for 93% into the hydroxamic acid in 36 h with only 1% (w/w) Candida antarctica lipase B (Novozym 435) in dioxane at 40 degrees C. This translates to a catalyst productivity of 68.5 g. g(-1). day(-1) and a space time yield of 149 g. L(-1). day(-1), unprecedented figures in the direct reaction of an acid with a nitrogen nucleophile in an organic solvent. PMID- 10699876 TI - A forced-flow membrane reactor for transfructosylation using ceramic membrane. AB - A forced-flow membrane reactor system for transfructosylation was investigated using several ceramic membranes having different pore sizes. beta Fructofuranosidase from Aspergillus niger ATCC 20611 was immobilized chemically to the inner surface of a ceramic membrane activated by a silane-coupling reagent. Sucrose solution was forced through the ceramic membrane by crossflow filtration while transfructosylation took place. The saccharide composition of the product, which was a mixture of fructooligosaccharides (FOS), was a function of the permeate flux, which was easily controlled by pressure. Using 0.2 micrometer pore size of symmetric ceramic membrane, the volumetric productivity obtained was 3.87 kg m(-3) s(-1), which was 560 times higher than that in a reported batch system, with a short residence time of 11 s. The half-life of the immobilized enzyme in the membrane was estimated to be 35 days by a long-term operation. PMID- 10699877 TI - Reversible enzyme immobilization via a very strong and nondistorting ionic adsorption on support-polyethylenimine composites. AB - New tailor-made anionic exchange resins have been prepared, based on films of large polyethylenimine polymers (e.g., MW 25,000) completely coating, via covalent immobilization, the surface of different porous supports (agarose, silica, polymeric resins). Most proteins contained in crude extracts from different sources have been very strongly adsorbed on them. Ionic exchange properties of such composites strongly depend on the size of polyethylenimine polymers as well as on the exact conditions of the covalent coating of the solids with the polymer. On the contrary, similar coating protocols yield similar matrices by using different porous supports as starting material. For example, 77% of all proteins contained in crude extracts from Escherichia coli were adsorbed, at low ionic strength, on the best matrices, and less than 15% of the adsorbed proteins were eluted from the support in the presence of 0.3 M NaCl. Under these conditions, 100% of the adsorbed proteins were eluted from conventional DEAE supports. Such polyethylenimine-support composites were also very suitable to perform very strong and nondistorting reversible immobilization of industrial enzymes. For example, lipase from Candida rugosa (CRL), beta galactosidase from Aspergillus oryzae and D-amino acid oxidase (DAAO) from Rhodotorula gracilis, were adsorbed on such matrices in a few minutes at pH 7.0 and 4 degrees C. Immobilized enzymes preserved 100% of catalytic activity and remained fully immobilized in 0.2 M NaCl. In addition to that, CRL and DAAO were highly stabilized upon immobilization. Stabilization of DAAO, a dimeric enzyme, seems to be due to the involvement of both enzyme subunits in the ionic adsorption. PMID- 10699878 TI - Three-dimensional engineered heart tissue from neonatal rat cardiac myocytes. AB - A technique is presented that allows neonatal rat cardiac myocytes to form spontaneously and coherently beating 3-dimensional engineered heart tissue (EHT) in vitro, either as a plane biconcaval matrix anchored at both sides on Velcro coated silicone tubes or as a ring. Contractile activity was monitored in standard organ baths or continuously in a CO(2) incubator for up to 18 days (=26 days after casting). Long-term measurements showed an increase in force between days 8 and 18 after casting and stable forces thereafter. At day 10, the twitch amplitude (TA) of electrically paced EHTs (average length x width x thickness, 11 x 6 x 0.4 mm) was 0.51 mN at length of maximal force development (L(max)) and a maximally effective calcium concentration. EHTs showed typical features of neonatal rat heart: a positive force-length and a negative force-frequency relation, high sensitivity to calcium (EC(50) 0.24 mM), modest positive inotropic (increase in TA by 46%) and pronounced positive lusitropic effect of isoprenaline (decrease in twitch duration by 21%). Both effects of isoprenaline were sensitive to the muscarinic receptor agonist carbachol in a pertussis toxin-sensitive manner. Adenovirus-mediated gene transfer of beta-galactosidase into EHTs reached 100% efficiency. In summary, EHTs retain many of the physiological characteristics of rat cardiac tissue and allow efficient gene transfer with subsequent force measurement. PMID- 10699879 TI - Development and characterization of an oxygen-dependent inducible promoter system, the modified nar promoter in a mutant Escherichia coli. AB - A nar promoter system (a modified nar promoter in a mutant host Escherichia coli (pMW618/W3110narL(-))), which is maximally induced under microaerobic conditions, was developed and characterized through batch and fed-batch culture to see whether the modified nar promoter can be used as an oxygen-dependent inducible promoter in the absence of nitrate ion. The modified nar promoter (pMW618) derived by mutations at -10 and -35 regions of the wild-type nar promoter does not require nitrate ion for the full induction, while a mutant host E. coli, W3110narL(-), does not express nitrate-dependent regulatory protein, NARL, from the host chromosome. In this study, it was found from fed-batch culture that the specific beta-galactosidase activity expressed from the lacZ gene fused to the modified nar promoter in the absence of nitrate ion was maximal when E. coli was grown under aerobic conditions (dissolved oxygen (DO) at 80%) to absorbance at 600 nm (OD(600)) of 35, and then the modified nar promoter was induced by lowering DO to 1-2% with alternating microaerobic and aerobic conditions. The maximal specific beta-galactosidase activity became 58,000 Miller at OD(600) of 160 with an induction ratio of 20. On the basis of these results, we conclude that the modified nar promoter system (pMW618/W3110narL(-)), requiring only reduction of DO for the full induction, provides a convenient and effective high level expression system under conditions of fed-batch culture. PMID- 10699880 TI - Rapid in vivo evolution of a beta-lactamase using phagemids. AB - RNA viruses are capable of undergoing extremely rapid evolution due to their high rates of reproduction, small genome size, and a high frequency of spontaneous mutagenesis. Here we demonstrate that a virus-like, evolutionary state can be created by propagating a phagemid population in a hypermutator strain of Escherichia coli in the presence of a helper phage. This enables one to subject individual phagemid-encoded genes to rapid in vivo evolution. We applied this approach to TEM-1 beta-lactamase which confers resistance to 0.05 mg/L of the antibiotic cefotaxime. After 3 weeks of in vivo evolution we were able to isolate a double mutant, E104K/G238S, of the enzyme which confers a 500-fold increased level of resistance to cefotaxime compared to the starting enzyme. In two independent experiments we obtained a triple mutant, E104K/G238S/T263M, which confers a 1000-fold increase in resistance compared to the wild type enzyme. The same three mutations have been previously observed in TEM-4 beta-lactamase which was discovered in a highly cefotaxime-resistant clinical isolate. The probability of randomly obtaining a beta-lactamase carrying three identical point mutations is less than 10(-10). This indicates that phagemid evolution can rapidly reproduce evolution occurring in nature. PMID- 10699881 TI - Discrepancies between East and West. PMID- 10699882 TI - Pathologic analysis of sentinel lymph nodes in breast carcinoma. PMID- 10699883 TI - Despite limited data, coexistent lobular carcinoma in situ should not be a contraindication to breast conservation for women with invasive breast carcinoma. PMID- 10699884 TI - Despite limited data, coexistent lobular carcinoma in situ should not be a contraindication to breast conservation for women with invasive breast carcinoma: reply PMID- 10699885 TI - Salivary flow and its relation with oral symptoms in terminally ill patients. AB - BACKGROUND: Patients with terminal malignant disease commonly report hyposalivation or xerostomia. This leads to "dry mouth," fungal infection, and mucosal abnormalities. To the authors' knowledge oral symptomatology and findings have not been correlated previously with accurate salivary flow measurements. METHODS: Measurement of stimulated parotid salivary flow rate and clinical recording of oral symptoms within 24 hours from the time of hospital admission were obtained in 48 terminally ill cancer patients. Subjective reporting of symptoms by patients, parotid salivary flow rate, clinical recording of dental status, presence of candidiasis, angular cheilitis, and dryness of the floor of the mouth were obtained. RESULTS: A clinical diagnosis of oral candidiasis was made tentatively in 94% of patients, and 50% of the patients were found to have angular cheilitis. Thirty-one of 45 evaluable patients (68%) reported a sensation of oral dryness. Sixteen of the 48 patients (33%) had no saliva at the floor of the mouth. Analysis of individual salivary flow rates was stratified into 3 levels of secretion: 0, < 0.2, and > or= 0.2 mL/minute. Symptoms were found to correlate with salivary flow rates. CONCLUSIONS: In the current study, symptoms were found to be most severe in the patients with xerostomia followed by those patients with hyposalivation. Treatment should be directed individually to each group of patients using either salivary substitutes or stimulants. The rate of incidence of oral pathologic findings may be higher than formerly recognized. PMID- 10699886 TI - A phase I/II study of external beam radiation, brachytherapy, and concurrent chemotherapy for patients with localized carcinoma of the esophagus (Radiation Therapy Oncology Group Study 9207): final report. AB - BACKGROUND: A multiinstitutional, prospective study of the Radiation Therapy Oncology Group (RTOG) was designed to determine the feasibility and toxicity of chemotherapy, external beam radiation, and esophageal brachytherapy (EB) in a potentially curable group of patients with adenocarcinoma or squamous cell carcinoma of the esophagus. A preliminary analysis indicated a 17% 1-year actuarial risk of treatment-related fistulas. A final analysis of this study was considered important to determine the median survival time, local control, and late toxicity associated with this treatment regimen. METHODS: Planned treatment was 50 grays (Gy) of external beam radiation (25 fractions given over 5 weeks) followed 2 weeks later by EB (either high-dose-rate 5 Gy during Weeks 8, 9, and 10, for a total of 15 Gy, or low-dose-rate 20 Gy during Week 8). Chemotherapy was given during Weeks 1, 5, 8, and 11, with cisplatin 75 mg/m(2) and 5-fluorouracil 1000 mg/m(2)/24 hours in a 96-hour infusion. RESULTS: Of the 49 eligible patients, 45 (92%) had squamous histology and 4 (6%) had adenocarcinoma. Forty seven patients (96%) completed external beam radiation plus at least 2 courses of chemotherapy, whereas 34 patients (69%) were able to complete external beam radiation, EB, and at least 2 courses of chemotherapy. The estimated survival rate at 12 months was 49%, with an estimated median survival of 11 months. Life threatening toxicity or treatment-related death occurred in 12 (24%) and 5 (10%) cases, respectively. Treatment-related esophageal fistulas occurred in 6 cases (12% overall, 14% of patients starting EB) at 0.5-6.2 months from the first day of brachytherapy, leading to death in 3 cases. CONCLUSIONS: In this study, severe toxicity, including treatment-related fistulas, occurred within 7 months of brachytherapy. Based on the 12% incidence of fistulas, the authors continue to urge caution in employing EB, particularly when used in conjunction with chemotherapy. PMID- 10699887 TI - Differences in diagnostic criteria for esophageal squamous cell carcinoma between Japanese and Western pathologists. AB - BACKGROUND: Large discrepancies have been found between Western and Japanese pathologists in the diagnosis of adenoma/dysplasia versus carcinoma for gastric and colorectal glandular lesions. It is important to determine whether similar differences exist in the diagnosis of esophageal squamous lesions. METHODS: Eleven expert gastrointestinal pathologists from Japan, North America, and Europe individually reviewed a set of microscopic slides containing 21 sections of biopsies and corresponding endoscopic mucosal resection specimens from Japanese patients with superficial esophageal squamous neoplastic lesions. The pathologists indicated the pathologic findings on which they based each diagnosis. RESULTS: Invasion was the most important diagnostic criterion of carcinoma for the Western pathologists whereas nuclear and structural features were more important for the Japanese pathologists. For two sections showing low grade dysplasia according to most Western pathologists, the Japanese pathologists diagnosed suspected carcinoma in one case and definite carcinoma in the other. For nine sections with high grade dysplasia according to the Western pathologists, the Japanese pathologists diagnosed suspected carcinoma in two cases and definite carcinoma in seven cases. For six sections with suspected carcinoma according to most Western pathologists, the Japanese pathologists diagnosed suspected carcinoma in one case and definite carcinoma in five cases. Four sections showed definite carcinoma according to both the Western and Japanese pathologists. Thus, there was agreement among the Western and Japanese pathologists for only 5 of the 21 sections (kappa value, 0.04). However, when high grade dysplasia, noninvasive carcinoma, and suspected carcinoma were grouped together, the agreement was excellent (19 of the 21 sections; kappa value, 0.75). CONCLUSIONS: In Japan, esophageal squamous cell carcinoma is diagnosed mainly based on nuclear criteria, even in cases judged to be noninvasive low grade dysplasia in the West. This difference in diagnostic practice may contribute to the relatively high incidence rate and good prognosis of superficial esophageal carcinoma in Japan. To improve the comparability of research data, the authors recommend that high grade dysplasia, noninvasive carcinoma, and suspected carcinoma be grouped together into one category of "noninvasive high grade neoplasia." [See editorial on pages 969-70, this issue.] PMID- 10699888 TI - Preoperative treatment with tegafur suppositories enhances apoptosis and reduces the intratumoral microvessel density of human colorectal carcinoma. AB - BACKGROUND: This study examined the effect of tegafur, a depot of 5-fluorouracil, in human colorectal carcinomas in terms of apoptosis, cell proliferation, and expression of p53 gene and angiogenesis-related molecules. METHODS: A total of 32 patients with colorectal carcinoma were divided into 2 groups; 20 patients received tegafur suppositories (TS) at 1 g/day for 14 days before surgery, and 12 patients did not receive any chemotherapy. Surgically removed specimens were examined immunohistochemically for Ki-67, CD34, p53, p21, Bax, vascular endothelial growth factor (VEGF), and thymidine phosphorylase (dThdPase). Apoptotic tumor cells were visualized by the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-digoxigenin nick-end labeling (TUNEL) procedure. RESULTS: The mean percentage of apoptotic index (AI) was 6.9 +/- 1.2 in the 20 TS treated tumors and 4.4 +/- 1.0 in the 12 nontreated tumors (P < 0.001). In contrast, the mean percentage of Ki-67 labeling index (KI) became significantly lower in the former group (P < 0.05). The frequency of p21 expression was significantly higher in the TS-treated group than in the nontreated group (P < 0.05), whereas no difference was detected in p53 and Bax expression between the two groups. The mean intratumoral microvessel density was 47.8 +/- 19.8 in the TS treated tumors and 66.8 +/- 16.5 in the nontreated tumors (P < 0.01). The frequency of dThdPase expression, but not of VEGF expression, became significantly lower with the TS treatment. p53 expression did not correlate with AI, KI, IMV density, or the expression of VEGF, p21, or Bax, except for dThdPase, which was significantly higher in the 18 p53 positive tumors (P < 0.05). CONCLUSIONS: Preoperative TS treatment enhances apoptosis and suppresses angiogenesis of colorectal carcinomas in a p53-independent manner. PMID- 10699889 TI - Effect of viral status on recurrence after liver resection for patients with hepatitis B virus-related hepatocellular carcinoma. AB - BACKGROUND: Risk factors for recurrence after resection of hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC) require more precise definition. METHODS: Forty patients who underwent liver resection for HBV-related HCC were studied. Their clinical findings, laboratory data (including viral status), pathologic findings, and operative methods were evaluated for recurrence risk in univariate and multivariate analyses. RESULTS: The HB envelope (HBe) antigen, wild-type HBV, intrahepatic metastases, elevated serum activities of aspartate aminotransferase and alanine aminotransferase, and moderately or severely active hepatitis were more likely to be found in patients with a high viral load than in patients with a low viral load. Precore mutant-type HBV was more likely to be found in patients with a low viral load than in patients with a high viral load. The platelet count was significantly lower in the patients with a high viral load. A high viral load, the presence of wild-type HBV, the absence of anti-HBe, the absence of precore mutant-type HBV, Child score B, a low platelet count, and a positive surgical margin were risk factors for recurrence in univariate analysis. A nonanatomic resection tended to be a risk factor. A high viral load and positive surgical margin were independent risk factors for recurrence. CONCLUSIONS: The measurement of viral load and detection of anti-HBe, wild-type HBV, and precore mutant-type HBV are useful for estimating a patient's prognosis after resection of HBV-related HCC. PMID- 10699890 TI - Lack of frameshift mutations at coding mononucleotide repeats in hepatocellular carcinoma in Japanese patients. AB - BACKGROUND: Microsatellite instability occurs frequently in hereditary nonpolyposis colorectal carcinoma, in sporadic gastrointestinal carcinoma, and in other tumors. In these tumors, slippage-related frameshift mutations have been detected at coding mononucleotide repeats in genes such as those for transforming growth factor-beta receptor type II (TGFbetaRII), mannose 6-phosphate/insulinlike growth factor II receptor (M6P/IGFIIR), hMSH3, hMSH6, and Bcl-2-associated X protein (BAX). Because these genes regulate cell growth or repair DNA mismatches, loss of their function is thought to promote tumor development. The authors screened for these frameshift mutations and investigated the incidence of microsatellite instability (MI) in hepatocellular carcinoma (HCC) in Japan. METHODS: Fifty HCC samples were analyzed in this study. The authors used polymerase chain reactions to screen for frameshift mutation at the TGFbetaRII (A)(10) tract, the M6P/IGFIIR (G)(8) tract, the hMSH3 (A)(8) tract, the hMSH6 (C)(8) tract, and the BAX (G)(8) tract. For MI analysis, matched tumor and nontumor liver DNA were investigated with respect to 10 microsatellite loci. RESULTS: No frameshift mutation was detected in any case, and only 4% of these cancers exhibited MI in comparisons between tumor and nontumor liver specimens. CONCLUSIONS: This study suggests that frameshift mutation at coding mononucleotide repeats within TGFbetaRII, M6P/IGFIIR, hMSH3, hMSH6, and BAX genes did not seem to be involved in hepatocarcinogenesis in the Japanese population studied. PMID- 10699891 TI - p53 mutation pattern in hepatocellular carcinoma in workers exposed to vinyl chloride. AB - BACKGROUND: Vinyl chloride (VC) is a known animal and human carcinogen that is associated with liver angiosarcoma and most likely also with hepatocellular carcinoma (HCC) in humans. METHODS: The authors examined the presence of p53 gene mutations in 18 HCC specimens from patients with known exposure to VC (median, 8883 parts per million-years; median duration, 245 months). In all cases, other risk factors for the development of HCC (hepatitis B virus and hepatitis C virus infections, alcohol consumption, and metabolic or autoimmune disorders) were excluded. Three patients had concomitant cirrhosis. The p53 gene was examined by direct sequencing of exons 5-9. RESULTS: Mutations of the p53 gene were found in 11 of 18 HCCs examined. The point mutations detected were comprised of five transversions and five transitions. Five of 11 mutations (codons 175, 245, 248, 273, and 282) occurred at CpG sites. Histopathologic liver alterations (mild sinusoidal dilatation, [portal] fibrosis, and centrilobular siderosis) in tumor surrounding nonneoplastic liver confirmed exposure to VC. CONCLUSIONS: The results of the current study indicated a relation between VC exposure and the development of HCC. The mutation pattern of p53 with a nearly equal rate of incidence of transitions and transversions and a high rate of incidence of mutations at CpG sites may reflect endogenous mechanisms (e.g., deamination of 5 methylcytosine) rather than exogenous carcinogens. PMID- 10699892 TI - Molecular detection of metastatic pancreatic carcinoma cells using a multimarker reverse transcriptase-polymerase chain reaction assay. AB - BACKGROUND: The diagnosis of pancreatic carcinoma is often associated with a poor prognosis, because most patients already have advanced disease. A highly sensitive assay to detect the progression of pancreatic carcinoma would be of significant clinical utility. The authors developed multiple tumor mRNA markers for reverse transcriptase-polymerase chain reaction (RT-PCR) to detect metastatic tumor cells in the blood and tissue of patients with American Joint Committee on Cancer (AJCC) Stage II/III or IV pancreatic carcinoma. METHODS: An RT-PCR plus Southern blot assay was used to detect mRNA of tumor markers in blood and tissues. mRNA expression of the tumor progression markers MET (hepatocyte growth factor receptor gene c-met), GalNAc-T (beta1,4- N-acetyl-galactosaminyl transferase), and beta-hCG (beta-human chorionic gonadotropin) was evaluated in 9 pancreatic carcinoma cell lines, 13 tumor biopsy specimens, 5 nonmalignant pancreatic tissue specimens, and blood from 33 pancreatic carcinoma patients and 32 healthy donors. RESULTS: The detection limit of the assay was 1 rhog, 10 rhog, and 10 rhog for MET, GalNAc-T, and beta-hCG mRNA expression, respectively. The pancreatic carcinoma cell lines expressed all three mRNA markers. Of blood specimens from 17 patients with AJCC Stage IV pancreatic carcinoma, 82%, 65%, and 76% were MET, GalNAc-T, and beta-hCG mRNA positive, respectively. Of blood specimens from 16 patients with AJCC Stage II/III disease, 88% were positive for at least 1 mRNA marker. CONCLUSIONS: A multiple molecular marker assay was developed to detect cancer cells in blood and tissue from patients with different stages of pancreatic carcinoma. The detection of cancer cells in the blood may be used as a marker of pancreatic tumor progression and may be useful in monitoring response to therapy. PMID- 10699893 TI - Phase I trial of docetaxel and vinorelbine in patients with advanced nonsmall cell lung carcinoma. AB - BACKGROUND: With preclinical evidence of synergy, this dose-finding trial examining the combination of docetaxel and vinorelbine given with prophylactic filgrastim for the treatment of patients with nonsmall cell lung carcinoma was undertaken. METHODS: Twenty-seven patients with advanced nonsmall cell lung carcinoma received vinorelbine as an intravenous push immediately followed by docetaxel as a 1-hour intravenous infusion once every 2 weeks at 1 of 7 different dose levels. Vinorelbine was escalated from 15 mg/m(2) (Level I) to 45 mg/m(2) (Level VII) and docetaxel was increased from 50 mg/m(2) (Level I) to 60 mg/m(2) (Level VII). Prophylactic corticosteroids and filgrastim were employed prospectively. RESULTS: After completion of dose Level VII, accrual was terminated because Phase II dose intensity of both agents had been reached and further escalation was believed to be unsafe. At dose Level VII, one episode of first-cycle febrile neutropenia and a death after three treatment cycles due to Haemophilus influenzae sepsis (Grade 5 toxicity according to the Common Toxicity Criteria of the National Cancer Institute) without neutropenia were noted. In all, 209 treatment cycles were administered and febrile neutropenia was observed in only 4 of these treatments (1.9%). Bacteremia occurred in three patients (four episodes) in the absence of neutropenia. Symptomatic onycholysis was observed in three patients. Clinically significant peripheral neuropathy and fluid retention were rare. Confirmed partial responses were noted in 10 patients for a response rate of 37% (95% confidence interval, 20-57%). CONCLUSIONS: Docetaxel at a dose of 60 mg/m(2) and vinorelbine at a dose of 45 mg/m(2), both given every 2 weeks, can be combined safely to achieve Phase II dose intensity of both agents. An ongoing Phase II trial will define the activity of this treatment combination. PMID- 10699894 TI - Results of modern therapy for patients with mediastinal nonseminomatous germ cell tumors. AB - BACKGROUND: The aim of this study was to determine the effects of independent prognostic variables, such as prechemotherapy tumor markers, the extent of disease at diagnosis, the tumor markers postchemotherapy (PC), and the pathology of the PC residual mass on the overall survival of patients with primary mediastinal nonseminomatous germ cell tumors (PMNSGCT). METHODS: The authors undertook a retrospective review of 39 patients with PMNSGCT between 1983 and 1997 who received their initial chemotherapy at Indiana University and 36 additional patients referred for PC resection. All patients received chemotherapy based on the combination of cisplatin and etoposide. The median follow-up was 22 months (range, 12-144 months). RESULTS: The prechemotherapy tumor markers did not affect overall survival. Extent of disease (mediastinal only vs. visceral metastases) was an important prognostic factor for survival in univariate analysis (P = 0.042). Sixty-two of 75 patients underwent PC resection of residual disease. Fifteen of the 62 patients achieved a CR with chemotherapy alone, as the PC resection revealed only necrosis. Fourteen of these 15 patients continuously had no evidence of disease (NED). Forty-seven of the 62 patients had NED with chemotherapy and PC resection, including 31 with teratoma and 16 with carcinoma. However, 11 of 31 with teratoma and 11 of 16 with carcinoma subsequently relapsed. Although 18 patients had elevated tumor markers at the time of PC resection, 6 of 18 had only necrosis and 4 had teratoma. The PC tumor markers did not affect survival. The pathology of the resected specimen was the most significant predictor of survival in multivariate analysis (P < 0.001). CONCLUSIONS: Twenty-eight of 39 patients (71.8%) with PMNSGCT treated at Indiana University achieved NED status, but only 16 (41%) continuously had NED. Twenty of 36 (55.5%) referred for resection continuously had NED. Disease confined to the mediastinum and necrosis in the PC specimen were important prognostic factors for survival. PMID- 10699895 TI - ATM gene deletion in patients with adult acute lymphoblastic leukemia. AB - BACKGROUND: Loss of heterozygosity (LOH) at the ATM gene (mutated in ataxia telangiectasia [AT] patients) and ATM protein deficiency occur in 14% and 34%, respectively, of patients with chronic lymphocytic leukemia (CLL). ATM protein deficiency also is associated with aggressive disease and worse patient survival. Considering the aberrations in the ATM gene in CLL and the high rate of incidence of lymphoid neoplasias in AT patients, the authors investigated its incidence rate and significance in patients with adult acute lymphoblastic leukemia (ALL). METHODS: Samples from 36 adults with ALL were analyzed for LOH and homozygous deletion (HD) using a panel of three microsatellite markers located at the ATM gene (D11S2179), the MLL gene (D11S1356), and the BCL1 gene (D11S987) loci. These markers are located within the 11q13-q23 locus. RESULTS: Of the 36 informative cases, 10 (28%) showed deletions (7 LOH and 3 HDs) at the D11S2179 marker. In two patients, the deletions were extended to the MLL gene locus. These deletions were submicroscopic because only 3% (1 of 36) of patients showed abnormalities involving 11q23 using cytogenetic studies. The authors also estimated the levels of ATM protein in 15 ALL patients and 12 healthy volunteers by radioimmunoassay. The ATM protein levels in cases with LOH at the ATM gene were between 15-50% of those from normal bone marrow. In contrast to CLL patients, patients with LOH or HD at the ATM gene locus showed better survival compared with patients without ATM gene deletions (P = 0.003). CONCLUSIONS: LOH of the ATM gene and protein deficiency are common in adult ALL, are not demonstrated at the cytogenetic level, and are associated with a favorable prognosis. The authors speculate that ATM deficiency may increase the sensitivity of leukemic blasts to the chemotherapy used during induction and after disease remission in patients with adult ALL. The relatively high frequency of deletion of the D11S2179 marker compared with the D11S1356 marker suggests that ATM is the target gene of the deletion at the 11q23 locus, and that such deletions may play a role in the pathogenesis of ALL. PMID- 10699896 TI - Analysis of prognosis and disease progression after local recurrence of melanoma. AB - BACKGROUND: Local recurrence of melanoma is associated with a grave prognosis. However, the characteristics and the mode of disease progression for patients with local recurrence have not been adequately addressed in the literature. METHODS: A retrospective analysis of patients treated at a single institution revealed a subset of patients (n = 648) with local recurrence of melanoma as a first event. Patient characteristics, histologic determinants, and disease free interval were variables used to identify prognostic factors. RESULTS: In this group of patients, male gender (P = 0. 0163), increasing age (P = 0.0001), head and neck primaries (P = 0. 0001), thicker Breslow depths (P = 0.0022), deeper Clark levels (P = 0.0010), and ulceration of the primary tumor (P = 0.0348) suggested a shorter time until local recurrence. Breslow depth (P = 0.0004), Clark level (P = 0.0043), and ulceration (P = 0.0001) still factored into the survival prognosis after recurrence. Truncal primaries (P = 0.0005) and shorter disease free intervals (P = 0.0098) were also associated with poorer outcomes after recurrence. Of the 648 patients, 124 showed no progression, 196 developed another local recurrence, 178 developed in-transit/lymph node metastases, and 150 had systemic recurrences. Survival was only 33.6% for patients with further metastases, compared with 77.4% for patients with no progression of disease after a median follow-up of 38.9 months. CONCLUSIONS: There was a 48.5% mortality rate at 5 years of follow-up after local recurrence. Long term survival (> 10 years) was estimated to be 34.9%. The patterns of failure after local recurrence suggest that patients may benefit from aggressive locoregional therapy. PMID- 10699897 TI - The relation between the presence and extent of lobular carcinoma in situ and the risk of local recurrence for patients with infiltrating carcinoma of the breast treated with conservative surgery and radiation therapy. AB - BACKGROUND: When found in an otherwise benign biopsy, lobular carcinoma in situ (LCIS) has been associated with an increased risk of development of a subsequent invasive breast carcinoma. However, the association between LCIS and the risk of subsequent local recurrence in patients with infiltrating carcinoma treated with conservative surgery and radiation therapy has received relatively little attention. METHODS: Between 1968 and 1986, 1625 patients with clinical Stage I-II invasive breast carcinoma were treated at the Joint Center for Radiation Therapy at Harvard Medical School with breast-conserving surgery (CS) and radiation therapy (RT) to a total dose to the primary site of > or =60 grays. Analysis was limited to 1181 patients with infiltrating ductal carcinoma, infiltrating lobular carcinoma, or infiltrating carcinoma with mixed ductal and lobular features who, on review of their histologic slides, had sufficient normal tissue adjacent to the tumor to evaluate for the presence of LCIS and also had a minimum potential follow-up time of 8 years. The median follow-up time was 161 months. RESULTS: One hundred thirty-seven patients (12%) had LCIS either within the tumor or in the macroscopically normal adjacent tissue. The 8-year crude risk of recurrence was not significantly increased for patients with LCIS associated with invasive ductal, invasive lobular, or mixed ductal and lobular carcinoma. Among the 119 patients with associated LCIS adjacent to the tumor, the 8-year rate of local recurrence was 13%, compared with 12% for the 1062 patients without associated LCIS. For the 70 patients with moderate or marked LCIS adjacent to the tumor, the 8-year rate of local recurrence was 13%. The extent of LCIS did not affect the risk of recurrence. The risks of contralateral disease and of distant failure were similarly not affected by the presence or extent of LCIS. CONCLUSIONS: Breast-conserving therapy involving limited surgery and radiation therapy is an appropriate method of treating patients with invasive breast carcinoma with or without associated LCIS. Neither the presence nor the extent of LCIS should influence management decisions regarding patients with invasive breast carcinoma. [See editorial counterpoint and reply to counterpoint on pages 978-81 and 982-3, this issue.] PMID- 10699898 TI - Cutaneous fibrosis induced by docetaxel: a case report. AB - BACKGROUND: Docetaxel is a taxoid antineoplastic agent approved for use in the treatment of metastatic breast carcinoma. The current study reports an unusual case of generalized cutaneous fibrosis in a 39-year-old white female after treatment with 18 cycles of docetaxel for metastatic breast carcinoma. METHODS: Cutaneous fibrosis represents a rare and unique reaction associated with the cyclic use of docetaxel. The reaction is manifested by a distinct sequence of events involving pronounced edema followed by the rapid development of cutaneous fibrosis in dependent areas. RESULTS: Cessation of therapy results in dramatic reversal of the fibrotic process. CONCLUSIONS: This case report further substantiates the belief that docetaxel represents one of a very limited number of agents that appear capable of giving rise to scleroderma-like features. PMID- 10699899 TI - Pamidronate prevents skeletal complications and is effective palliative treatment in women with breast carcinoma and osteolytic bone metastases: long term follow up of two randomized, placebo-controlled trials. AB - BACKGROUND: Pamidronate therapy previously has been shown to reduce skeletal complications effectively for up to 12 months in breast carcinoma patients with bone metastases. The current study data provide further follow-up results regarding the effects of long term (up to 24 months) pamidronate treatment in women with breast carcinoma and osteolytic metastases. METHODS: Follow-up results from two prospective, multicenter, randomized, double-blind, placebo-controlled intervention trials conducted at academic and community oncology centers were combined to provide a large data set with which to evaluate the long term efficacy and safety of pamidronate therapy. Seven hundred fifty-four women with Stage IV breast carcinoma and osteolytic metastases were randomized to the 2 treatment arms of the trial. Three patients were excluded from the intent-to treat population for the analysis. A total of 751 evaluable patients were randomized to receive either a 90-mg intravenous pamidronate infusion (367 patients) or a placebo infusion (384 patients) every 3-4 weeks. The primary outcome measures were skeletal morbidity rate (events/year), proportion of patients developing a skeletal complication, and time to first skeletal complication. RESULTS: Of the 367 women receiving pamidronate, 115 (31.3%) completed the trial and 81 (22.1%) discontinued the study due to adverse events. Of the 384 women who received placebo, 100 (26.0%) completed the study and 76 (19.8%) discontinued the study due to adverse events. The skeletal morbidity rate was 2.4 in the pamidronate group and 3.7 in the placebo group (P < 0.001). In the pamidronate group, 186 of the 367 patients (51%) had skeletal complications compared with 246 of the 384 patients in the placebo group (64%) (P < 0.001). The median time to first skeletal complication was 12.7 months in the pamidronate group and 7 months in the placebo group (P < 0.001). Six patients treated with pamidronate discontinued treatment due to drug-related adverse events. Pain and analgesic scores were significantly worse in the placebo group compared with those patients in the pamidronate group. CONCLUSIONS: In the current study, monthly infusions of 90 mg of pamidronate as a supplement to antineoplastic therapy were found to be well tolerated and superior to antineoplastic therapy alone in preventing skeletal complications and palliating symptoms for at least 24 months in breast carcinoma patients with osteolytic bone metastases. PMID- 10699900 TI - Mutant p53 protein overexpression in women with ipsilateral breast tumor recurrence following lumpectomy and radiation therapy. AB - BACKGROUND: The p53 tumor suppressor gene encodes a nuclear phosphoprotein that is thought to be important to cell cycle regulation and DNA repair and that also may regulate induction of apoptosis by ionizing radiation. Somatic p53 gene mutations occur in 30-50% of breast carcinomas and are associated with poor prognosis. Mutations in the p53 gene result in prolonged stability of the protein that can be detected by immunohistochemical techniques. In a matched case-control study of breast carcinoma patients with ipsilateral breast tumor recurrence (IBTR) following lumpectomy and radiation therapy, the authors investigated the frequency and prognostic significance of somatic p53 mutations as well as the clinical characteristics of patients with these mutations. METHODS: Between 1973 and 1995, there were 121 breast carcinoma patients with IBTR following lumpectomy and radiation therapy, and the authors identified 47 patients in whom the paraffin embedded tissue blocks from the primary breast tumors were available for further molecular analysis. Forty-seven control breast carcinoma patients from the breast carcinoma data base were individually matched to the index cases who did not have IBTR for age, treatment date, follow-up, histology, margin status, radiation dose, and adjuvant treatment. Immunohistochemistry using a monoclonal antibody to mutant p53 protein was used to determine mutant p53 protein overexpression in breast tumors and appropriately scored. RESULTS: A total of 12 of 47 tumor specimens (26%) from index patients with breast tumor relapses demonstrated mutant p53 protein overexpression, whereas only 4 of 47 specimens from controls (9%) demonstrated high mutant p53 immunoreactivity (P = 0.02). The authors found that 9 of 23 patients (39%) with early breast tumor recurrences (recurrences within 4 years of diagnosis) had overexpression of mutant p53 protein, whereas only 1 of 23 control cases (4%) had high mutant p53 protein immunoreactivity (P = 0.003). In contrast, index cases from patients with late breast tumor relapses (more than 4 years after diagnosis), which are more likely to represent de novo breast tumors, and control cases from the breast carcinoma data base without IBTR had similar levels of mutant p53 protein overexpression (P = not significant). The 10-year distant disease free survival for patients with mutant p53 protein was 48%, compared with 67% for breast carcinoma patients without detection of mutant p53 protein (P = 0. 08). The authors found that 13 of 14 primary breast tumors (93%) with mutant p53 protein overexpression were estrogen receptor negative (P = 0.01) and 11 of 14 (79%) were progesterone receptor negative (P = not significant). CONCLUSIONS: In a matched case-control study, overexpression of mutant p53 protein has prognostic significance with respect to IBTR following lumpectomy and radiation therapy. Breast tumors with p53 mutations are generally estrogen receptor negative and are associated with compromised distant disease free survival. PMID- 10699901 TI - Pathologic analysis of sentinel and nonsentinel lymph nodes in breast carcinoma: a multicenter study. AB - BACKGROUND: Axillary lymph node status is a powerful prognostic factor in breast carcinoma; however, complications after axillary lymph node dissection are common. Sentinel lymph node biopsy is an alternative staging procedure. The sentinel lymph node postulate is that tumor cells migrating from the primary tumor colonize one or a few lymph nodes before colonizing subsequent lymph nodes. To validate this hypothesis, the distribution of occult and nonoccult metastases in sentinel and nonsentinel lymph nodes was evaluated. METHODS: Original pathology material was reviewed from 431 patients enrolled on a multicenter validation study of sentinel lymph node biopsy in breast carcinoma patients. Paraffin embedded tissue blocks of sentinel and nonsentinel lymph nodes were obtained for 214 lymph node negative patients. Additional sections from 100 and 200 microm deeper into the paraffin block were examined for the presence of occult metastatic carcinoma. Both routine and cytokeratin immunohistochemical stains were employed. RESULTS: Metastases were identified in 15.9% of sentinel lymph nodes and 4.2% of nonsentinel lymph nodes (odds ratio [OR] 4.3[ P < 0.001]; 95% confidence interval [95% CI], 3.5-5.4). Occult metastases were identified in 4. 09% of sentinel lymph nodes and 0.35% of nonsentinel lymph nodes (OR 12.3 [P < 0.001]; 95% CI, 5.6-28.6). The overall case conversion rate was 10.3%. All the occult metastases identified were < or = 1 mm in greatest individual dimension. The likelihood (OR) of metastases in nonsentinel lymph nodes was 13.4 times higher for sentinel lymph node positive than for sentinel lymph node negative patients (P < 0. 001; 95% CI, 6.7-28.1). CONCLUSIONS: The distribution of occult and nonoccult metastases in axillary lymph nodes validates the sentinel lymph node hypothesis. In addition, pathology review of cases confirmed the authors' previously reported finding that the sentinel lymph nodes are predictive of the final axillary lymph node status. Occult metastatic disease is more likely to be identified in sentinel lymph nodes, allowing future studies to focus attention on one or a few sentinel lymph nodes. However, the relation between occult metastatic disease in sentinel lymph nodes, disease free survival, and overall survival must be evaluated prior to endorsing the intensive analysis of sentinel lymph nodes in routine practice. [See editorial on pages 971-7, this issue.] PMID- 10699902 TI - Expression of MN/CA9 protein in Papanicolaou smears containing atypical glandular cells of undetermined significance is a diagnostic biomarker of cervical dysplasia and neoplasia. AB - BACKGROUND: Despite the enormous impact that Papanicolaou (Pap) smear screening has had on the incidence of cervical carcinoma in developed countries, there is still an unacceptably high frequency of occurrence of this cancer. In part, this is due to human error associated with cytologic diagnoses of Pap smears. Also, the use of new sampling devices, such as the cytobrush, has increased the complexity of diagnosing benign and neoplastic cervical cytology. This is particularly apparent in the diagnosis of atypical glandular cells of undetermined significance (AGUS). Approximately 40% of AGUS diagnoses have a corresponding significant lesion at biopsy follow-up, and 60% do not. There is clearly a need for an adjunct to cytologic diagnosis that can readily identify AGUS smears that are diagnostic of significant lesions. The authors have identified the MN/CA9 antigen as a strong candidate for an adjunct biomarker. METHODS: A total of 245 Pap smears of all AGUS diagnostic categories with histologic confirmation were studied. The median age of the patients was 39 years. The Bethesda system classification (AGUS-favor reactive, AGUS-not otherwise specified, and AGUS-favor neoplastic) was used. All of the Pap smears were decolorized and immunostained with monoclonal antibody to MN/CA9 antigen by the immunoperoxidase technique. The results of MN/CA9 immunoreactivity were correlated with the histologic data in a semiblinded fashion. RESULTS: The follow up biopsies showed that a high percentage (70%) of patients had low and high grade cervical intraepithelial neoplasia lesions, respectively (CIN I and CIN II or III). Clinically significant lesions-adenocarcinoma in situ/carcinoma (AIS/CA) and CIN II or III-were found in 50% of the cases. Among these, 11% were AIS/CA. In the three subcategories of AGUS diagnosis, the AGUS-not otherwise specified showed the broadest range of lesions in the follow-up biopsies. Three patterns of MN/CA9 immunoreactivity were observed in the Pap smears: 1) atypical cells, 2) normal endocervical cells only, and 3) all cells negative. All Pap smears that were MN/CA9 positive were histologically confirmed to be clinically significant lesions or CIN I; in addition, there were a very small number (n = 12) of cases of atypia. None of the benign lesions showed MN/CA9 expression in the corresponding Pap smears. Furthermore, the pattern of atypical cell immunostaining identified all cases with significant lesions (AIS/CA and CIN II or III) in the cervices. Conversely, the majority of CIN I cases (82%) and all cases of atypia showed positive immunostaining restricted to normal endocervical cells only. CONCLUSIONS: There is a clear association between MN/CA9 immunostaining of atypical cells and the presence of significant lesions in the cervix. Similarly, there is a clear association between lack of expression of MN/CA9 and the absence of cervical lesions. However, the screen does not allow discrimination between CIN I and atypia. The authors also found that, based on the combined patterns of morphology and immunostaining, they are able to discriminate between AIS and CIN II or III in AGUS Pap smear diagnoses. Thus, expression of the MN/CA9 antigen is indeed a discriminator of significant lesions in AGUS Pap smear diagnoses. PMID- 10699903 TI - Primary treatment choices for men with clinically localized prostate carcinoma detected by screening. AB - BACKGROUND: Increasingly, prostate carcinoma is diagnosed through screening. However, little is known regarding factors that influence a patient's decision concerning the treatment choices presented to him. METHODS: Subjects were prostate carcinoma patients detected through the Washington University PSA Prostate Cancer Screening Program between September 1989 and June 1998. The sources of data were the prostate specific antigen (PSA) screening database and follow-up questionnaire. RESULTS: Among 1809 study subjects, 79.2% chose radical prostatectomy (RP), 12.4% chose radiation therapy, and 8.4% chose watchful waiting (WW) as their decision regarding primary treatment. In bivariate analyses, education, income, age, indication for prostate biopsy, comorbidity score, serum PSA level, clinical stage, and pretreatment urinary and sexual function were associated significantly with treatment choice, but race, marital status, and Gleason grade were not. In a multivariate analysis, age, race, clinical stage, PSA level, and pretreatment urinary and sexual function were found to be associated significantly with treatment choice. For every 5-year decrease in age, the odds for choosing RP versus WW increased by 276%; for every 1-ng increase in PSA, the odds for choosing RP versus WW increased by 12%. Non African-American patients were greater than four times more likely to select RP versus WW. Patients with T2 tumors and those with normal pretreatment urinary function were three times more likely and twice more likely to choose RP versus WW, respectively. CONCLUSIONS: In the current study, RP was the most widely used treatment in patients with screen-detected prostate carcinoma. Age, race, PSA level, clinical stage, and pretreatment urinary and sexual function were significant factors influencing treatment selection. PMID- 10699904 TI - Expression of thymidine phosphorylase as an indicator of poor prognosis for patients with transitional cell carcinoma of the bladder. AB - BACKGROUND: Thymidine phosphorylase (TP), which is identical to platelet-derived endothelial cell growth factor (PD-ECGF), stimulates chemotaxis of endothelial cells and is involved in the angiogenesis of human solid tumors. METHODS: The activity and expression of TP were examined in human transitional cell carcinomas (TCCs) of the bladder, and their association with clinicopathologic findings was determined. The activity of the enzyme in 37 TCCs and 12 adjacent nonneoplastic tissues was measured spectrophotometrically. The expression of TP was also examined by immunoblotting. Immunohistochemical analysis was performed on 108 TCCs. RESULTS: TP activity in the carcinomas was higher than that in adjacent normal tissues (P = 0.002). TP activity in Grade 3 tumors or those classified as pT2-4 was higher than in Grade 1 and 2 tumors (P = 0.017) or those classified as pT1 (P = 0.007). The level of expression of TP detected by immunoblotting correlated well with TP activity. Immunohistochemical analyses showed that 62 of 108 cases (57.4%) were TP positive. There was a significant correlation between TP expression and histologic grade, infiltration pattern, local invasion, and lymph node metastasis. TP expression as a prognostic variable was studied using the Cox proportional hazards model. TP overexpression was an independent prognostic factor, as were lymph node metastasis and local invasion. CONCLUSIONS: These findings suggest that TP activity and its level of expression influence the progression of TCC and the prognoses of patients with this disease. PMID- 10699905 TI - Medullary thyroid carcinoma: clinical characteristics, treatment, prognostic factors, and a comparison of staging systems. AB - BACKGROUND: The clinical courses of patients with medullary thyroid carcinoma (MTC) vary, and a number of prognostic factors have been studied, but the significance of some of these factors remains controversial. METHODS: The study group consisted of 104 patients with MTC or C-cell hyperplasia managed at the hospitals of the University of California, San Francisco, between January 1960 and December 1998. Patients were classified as having sporadic MTC, familial non multiple endocrine neoplasia (MEN) MTC, MEN 2A, or MEN 2B. The TNM, European Organization for Research and Treatment of Cancer (EORTC), National Thyroid Cancer Treatment Cooperative Study (NTCTCS), and Surveillance, Epidemiology, and End Results (SEER) extent-of-disease stages were determined for each patient. The predictive values of these staging or prognostic scoring systems were compared by calculating the proportion of variance explained (PVE) for each system. RESULTS: Fifty-six percent of the patients had sporadic MTC, 22% had familial MTC, 15% had MEN 2A, and 7% had MEN 2B. The overall average age at diagnosis was 38 years, and patients with sporadic MTC presented at an older age (P < 0.05). Thirty-two percent of the patients with hereditary MTC were diagnosed by screening (genetic and/or biochemical). These patients had a lower incidence of cervical lymph node metastasis (P < 0.05) and 94.7% were cured at last follow-up (P < 0.0001) compared with patients not screened. Patients with sporadic MTC who had systemic symptoms (diarrhea, bone pain, or flushing) had widely metastatic MTC and 33.3% of those patients died within 5 years. Overall, 49.4% of the patients were cured, 12.3% had recurrent MTC, and 38.3% had persistent MTC. The mean follow-up time was 8.6 years (median, 5.0 years) with 10.7% (n=11) and 13.5% (n=14) cause specific mortality at 5 and 10 years, respectively. Patients with persistent or recurrent MTC who died of MTC lived for an average of 3.6 years (ranging from 1 month to 23.7 years). Patients who had total or subtotal thyroidectomy were less likely to have persistent or recurrent MTC (P < 0.05), and patients who had total thyroidectomy with cervical lymph node clearance required fewer reoperations for persistent or recurrent MTC (P < 0.05) than patients who underwent lesser procedures. In univariate analysis, age, gender, clinical presentation, TNM stage, sporadic/hereditary MTC, distant metastasis, and extent of thyroidectomy were significant prognostic factors. Only age and stage, however, remained independent prognostic factors in multivariate analysis. The TNM, EORTC, NTCTCS, and SEER staging systems were all accurate predictors of survival, but the EORTC prognostic scoring system had the highest PVE in this cohort. CONCLUSIONS: Screening for MTC and early treatment (total thyroidectomy with central neck lymph node clearance) had nearly a 100% cure rate. Patients with postoperative hypercalcitoninemia without clinical or radiologic evidence of residual tumor after apparently curative surgery may enjoy long term survival but have occult MTC. Only patient age at presentation and TNM stage were independent predictors of survival. The EORTC criteria, which included the greatest number of significant prognostic factors in our cohort, had the highest predictive value. PMID- 10699906 TI - Primary thyroid teratomas: a clinicopathologic study of 30 cases. AB - BACKGROUND: Primary thyroid teratomas are rare thyroid gland neoplasms of germ cell derivation that display features of trilineage differentiation. METHODS: The histologic and immunophenotypic features of 30 cases of thyroid teratomas were reviewed, patient follow-up was obtained, and the results were analyzed statistically. RESULTS: The patients included 15 females and 15 males ages newborn-56 years (mean, 12.4 years). All patients presented clinically with a mass in the thyroid, ranging in size from 2.0-13 cm in greatest dimension (mean, 6.0 cm). Histologically, the tumors usually were well circumscribed, although occasionally infiltrative into the thyroid parenchyma. Derivatives of all three germ layers (ectoderm, mesoderm, and endoderm) were present in varying degrees of maturity. The tumors were divided into benign (n = 7 tumors), immature (n = 14 tumors), and malignant (n = 9 tumors) as determined by an increasing percentage of tumor volume comprised of primitive mesenchymal or neural-type tissue. All the microscopically malignant tumors occurred in the adult population. Surgical excision was performed in 28 patients, followed by adjuvant therapy in 5 patients. Follow-up was obtained in 26 patients; 8 patients had died from or with tumor (5 neonates with immature histology and 3 adults with malignant histology; mean, 0.6 years) and 18 patients were alive with no evidence of disease at a mean of 16.9 years of follow-up. CONCLUSIONS: Thyroid teratomas are rare neoplasms that can be divided into three types depending on the presence and proportion of the immature component. The outcome is dependent largely on the age of the patient, the size of the tumor at the time of initial presentation, and the presence and proportion of immaturity. Surgical excision is the treatment of choice, with adjuvant therapy reserved for the malignant cases. PMID- 10699907 TI - Phase II evaluation of cisplatin and etoposide followed by mitotane at disease progression in patients with locally advanced or metastatic adrenocortical carcinoma: a Southwest Oncology Group Study. AB - BACKGROUND: A previous Southwest Oncology Group study demonstrated a 30% response rate with the combination of cisplatin and mitotane in the treatment of patients with metastatic adrenocortical carcinoma. Several case reports suggested that the combination of etoposide and cisplatin may be an active regimen in this disease. Because of these reports of potential activity, the authors conducted a Phase II trial evaluating the combination of etoposide and cisplatin. Due to the lack of data regarding the objective response rates to mitotane, the authors planned to evaluate the response rate to mitotane after disease progression on etoposide and cisplatin in patients with no prior mitotane therapy. METHODS: Patients with advanced, unresectable, or metastatic adrenocortical carcinoma with objectively measurable disease or biochemical abnormalities received cisplatin, 50 mg/m(2), intravenously on Days 1 and 2, and etoposide, 100 mg/m(2), on Days 1, 2, and 3. Cycles were repeated every 21 days. At the time of disease progression, patients who had not previously received mitotane received 1000 mg orally 4 times a day along with cortisone acetate and fludrocortisone acetate. RESULTS: Of the 47 patients entered onto the study, 45 were eligible. Nine patients had received mitotane previously and 36 had not. Objective responses were noted in 11% of patients (5 of 45 patients) (95% confidence interval, 3.7-24%). The median survival was 10 months. The most common toxic effects were hematologic, gastrointestinal, and neurologic. Only 16 patients with no prior mitotane therapy went on to receive mitotane at the time of disease progression. An objective response was noted in 13% of patients (2 of 16 patients). The most common toxic effects were edema and gastrointestinal effects. CONCLUSIONS: The current study demonstrates that the combination of cisplatin and etoposide has minimal activity in advanced and metastatic adrenocortical carcinoma and other treatment strategies are warranted. PMID- 10699908 TI - Alternating drug pairs with or without periodic reinduction in children with acute lymphoblastic leukemia in second bone marrow remission: a Pediatric Oncology Group Study. AB - BACKGROUND: Children with acute lymphoblastic leukemia (ALL) who experience hematologic recurrence while receiving chemotherapy or within 6 months after its cessation have a low cure rate. In this study (Pediatric Oncology Group Protocol 8303) two methods were examined for improving the outcome in these children. METHODS: After remission induction with prednisone, vincristine, daunorubicin, and asparaginase (PVDA) and consolidation chemotherapy with teniposide and cytarabine, patients received weekly continuation chemotherapy with rotating pairs of drugs, comprised of teniposide and cytarabine and vincristine and cyclophosphamide. In addition, they were randomized to receive or not receive repeated reinduction with PVDA. Patients with matched sibling donors were allowed to receive allogeneic bone marrow transplantation (BMT) instead of continued chemotherapy. RESULTS: Of 297 evaluable patients 258 (87%) achieved second complete hematologic remission. However, only 23 of these patients remained continuously free of leukemia > or =7 years after chemotherapy or BMT. Neither PVDA pulses nor BMT appeared to influence outcome at a statistically significant level. CONCLUSIONS: The results of the current study confirm prior reports of the low cure rate of children with ALL who experience hematologic recurrence during initial therapy or shortly after its cessation. New approaches are needed to prevent and retreat hematologic recurrence in pediatric ALL patients. PMID- 10699909 TI - Symptom and quality of life survey of medical oncology patients at a veterans affairs medical center: a role for symptom assessment. AB - BACKGROUND: The current study was conducted to assess symptom prevalence and symptom intensity and their relation to quality of life in medical oncology patients at a Veterans Affairs medical center. METHODS: Consecutive inpatients and outpatients were asked to complete the Functional Assessment Cancer Therapy (FACT-G), Memorial Symptom Assessment Scale (MSAS), and the Brief Pain Inventory. Symptoms then were analyzed by their relation to Karnofsky performance status (KPS) and quality of life. RESULTS: Two hundred forty patients participated. The median number of symptoms was 8 per patient (range, 0-30 symptoms). The 5 most prevalent symptoms were lack of energy (62%), pain (59%), dry mouth (54%), shortness of breath (50%), and difficulty sleeping (45%). Patients with moderate intensity pain had a median number of 11 symptoms and patients with moderate intensity lack of energy had a median number of 13 symptoms. The number of intense symptoms increased as the KPS decreased (P < 0.001). Patients with moderately intense pain or fatigue also were more likely to experience nausea, dyspnea, and lack of appetite. The number of symptoms rated as present on the MSAS was found to correlate significantly with the FACT-G Sum Quality of Life score. CONCLUSIONS: Intense symptoms were highly prevalent in this population. The presence of pain, lack of energy, or poor performance status should lead to comprehensive symptom assessment. Patients free of disease nevertheless still may experience intense symptoms. The number of symptoms present may be a helpful guide to quality of life. Routine comprehensive symptom assessment may identify a significant fraction of patients who urgently require intensive symptom palliation. PMID- 10699911 TI - Author reply PMID- 10699910 TI - Pretreatment staging by endoscopic ultrasonography does not predict complete response to neoadjuvant chemoradiation in patients with esophageal carcinoma. PMID- 10699912 TI - Gerald P. Murphy, M.D., D.Sc.: In memoriam PMID- 10699913 TI - Occurrence of primary cancers in association with multiple myeloma and Kaposi's sarcoma in the United States, 1973-1995. AB - The causes of multiple myeloma (MM) are obscure, but a laboratory association was recently reported between MM and human herpesvirus 8 (HHV-8), the probable etiologic agent of Kaposi's sarcoma (KS). Although there has been some additional laboratory corroboration, most laboratory studies have found no association between MM and HHV-8. We looked for indirect evidence of an HHV-8/MM association by evaluating whether MM is associated with KS in the United States. Cancer incidence and survival data were obtained from the Surveillance, Epidemiology, and End Results (SEER) program for the years 1973-1995. Strength of association was assessed for a number of cancer pairs using standardized incidence ratios (SIRs) (observed/expected double cancers). KS was strongly associated (SIR > 15) with non-Hodgkin's lymphoma and anal cancer, was modestly associated (2.5 < SIR < 5.5) with MM, Hodgkin's disease, and testicular cancer and was not significantly associated with 6 other cancers. Besides being associated with KS, MM was weakly associated (1.7 < SIR < 2.3) with Hodgkin's disease and testicular cancer. The SIRs for 7 other cancers paired with MM were all less than 1.6. Factors that might be responsible for the KS/MM association include MM-related immune dysfunction, HIV and HHV-8, but the role of these factors cannot be directly assessed through the SEER database. Although we cannot rule out the possibility that HHV-8 is linked to a small proportion of MM cases, the modest KS/MM association is evidence that the vast majority of MM cases are not likely to be associated with HHV-8. PMID- 10699914 TI - Low levels of urokinase plasminogen activator components in basal cell carcinoma of the skin. AB - Basal cell carcinoma of the skin (BCC) is the most common cancer worldwide. Unlike most other human malignancies, BCCs rarely metastasise. In this investigation, we show that the serine protease urokinase plasminogen activator (u-PA), which is causally involved in metastasis, is expressed at lower levels in BCCs compared to other skin cancers, such as squamous-cell carcinomas (SCCs) or malignant melanomas. Similarly, the u-PA receptor as well as the inhibitor PAI-1 were present at lower levels in BCCs relative to both SCCs and melanomas. In contrast to u-PA, tissue-plasminogen activator, which is not thought to be involved in metastasis, was present at similar levels in the different types of skin lesion investigated. We conclude that the failure of BCCs to metastasise may at least be partially related to low expression of components of the u-PA system. PMID- 10699915 TI - Expression of MAGE-antigens in normal tissues and cancer. AB - The human MAGE gene family encodes products that can be recognized by autologous cytotoxic T cells. Because MAGE genes are silent in most normal tissues except testis but are activated in a variety of neoplastic lesions, MAGE antigens represent ideal targets for immunotherapy. Current knowledge of MAGE gene expression is based primarily on mRNA typing and relatively little is known about MAGE protein expression. Monoclonal antibody (MAb) 57B, originally thought to be specific for MAGE-3, but now known to be reactive with other MAGE components, was used in the present study to analyze MAGE expression in a panel of normal and malignant tissues. In tests with a wide range of normal tissues, only spermatogenic cells of testis were reactive with 57B. In tumor tissues, significant immunoreactivity was observed in malignant melanomas and carcinomas of the lung, head and neck as well as urinary bladder. No 57B reactivity was seen with colorectal, prostatic or renal cell carcinomas. Lipo- and myosarcomas, as well as malignant fibrous histiocytoma (MFH), were negative, but synovial sarcomas showed intense immunoreactivity. A subset of seminomas was also strongly reactive with 57B. Tumor specimens showed great variability in the number of tumor cells showing 57B reactivity, with some tumors showing only small isolated clusters of positive cells to other tumors with uniform staining throughout the tumor. PMID- 10699916 TI - Stable amino-acid sequence of the mannose-6-phosphate/insulin-like growth-factor II receptor in ovarian carcinomas with loss of heterozygosity and in breast cancer cell lines. AB - The mannose-6-phosphate/insulin-like growth factor 2 receptor (Man-6-P/IGFII receptor) is involved in lysosomal enzyme sorting, IGFII degradation and pro TGFbeta activation. Genetic alterations in hepatocarcinomas and a few breast cancers suggest that this receptor behaves as a tumor suppressor. Moreover, hypersecretion and Man-6-P-independent targeting of cathepsins in breast and ovarian carcinomas also suggest alterations in this receptor. We studied the Man 6-P/IGFII receptor gene in 8 ovarian carcinomas, and 4 breast- and ovarian-cancer cell lines. The results confirmed a frequent loss of heterozygosity (LOH) in the 6q27-qter region in 5 out of 8 ovarian carcinomas. We used 23 overlapping RT-PCR fragments to sequence the whole coding region of the Man-6-P/IGFII receptor. The 2491 amino-acid sequence of this receptor was perfectly conserved in 9 out of 10 of our samples, including MCF7 and MDA-MB231 cells and 5 ovarian carcinomas with LOH. This allowed us to rectify the 2 previously published sequences which differed in several bases, and to propose a consensus amino-acid sequence. The only amino-acid change (Thr --> Ala) was in BG1 ovarian-cancer cells, and was due to an A-to-G substitution on one allele at nucleotide 2561. We found no bi allelic alterations in the 9 ovarian carcinomas, but 3 silent nucleotide substitutions leading to a lower cordon usage in 2 ovarian carcinomas with LOH. No mutation of the Man-6-P/IGFII receptor coding sequence was found in breast cancer cell lines to explain the cathepsin-D hypersecretion and Man-6-P independent trafficking described. We propose that, in breast and ovarian cancers, the frequent loss of one allele, associated with over-expression of some of its ligands, might be sufficient to saturate the receptor protein, displace the ligands to other sites, and consequently facilitate tumor progression. PMID- 10699917 TI - Detection of BRCA1 and BRCA2 mutations in breast cancer families by a comprehensive two-stage screening procedure. AB - We have developed a 2-stage protocol for BRCA1 and BRCA2 mutation screening from blood spot paper. Stage 1 screening was aimed to analyze patients at highest risk for the most common disease-associated sequence variants listed in the BIC database. Accordingly, stage1 testing implied detection of 18 disease- associated BRCA1 and 9 BRCA2 mutations by adapting the 5' nuclease assay to heterozygote screening. For stage 2 screening, we applied the conformation sensitive gel electrophoresis (CSGE) method by adapting this technique to automated heteroduplex analysis of BRCA1 and BRCA2 using fragment scanning on an ABI 377 sequencing device. Of the 120 patients with a family history of breast and ovarian cancer who took part in this study so far, 45 entered stage 1 testing. Disease-associated mutations were detected in 6 patients by stage 1 testing (13%). For these patients, the final result was available within 10 days. Mutation 300T-->G was found in 2 patients. One patient with mutation 3036delACAA in BRCA2 reported only 1 sister with a multifocal bilateral breast cancer. New disease-associated mutations were detected in 2 of the 114 patients who entered the stage 2 test (1.7%). Of particular interest was 1 patient who was diagnosed with a medullary breast carcinoma at age 39 and who had no family history of breast cancer. We conclude that pre-screening by 5' nuclease assay for the mutations most frequently seen in a given population represents a relatively effective first line of analysis. Subsequent detailed analysis by fluorescence conformation sensitive gel electrophoresis (F-CSGE) and fragment sequencing is a sensitive alternative to full nucleotide sequencing. PMID- 10699918 TI - Microsatellite analysis provides evidence of neoplastic transformation in long segment, but not in short-segment, Barrett's oesophagus. AB - It has been suggested that the high prevalence of short segments of specialised intestinal metaplasia (SIM) at the gastro-oesophageal junction is associated with the rising incidence of oesophageal adenocarcinoma. Our aims were to document the prevalence of short segments of SIM at the gastro-oesophageal junction in patients attending for routine endoscopy and to determine if there was molecular evidence of neoplastic transformation in those with SIM. Patients (n = 101) were recruited from randomly selected upper gastro-intestinal endoscopy lists. Biopsy specimens were taken at the squamo-columnar junction to assess the prevalence of SIM. Frozen sections were assessed for molecular evidence of neoplastic transformation using microsatellite analysis. Squamo-columnar biopsies were suitable for analysis in 95 patients, of whom 20 (21%) had oesophagitis and 2 (2%) had Barrett's oesophagus (>3 cm of endoscopically apparent columnar-lined oesophagus). Twenty patients had SIM at the gastro-oesophageal junction, including 2 with Barrett's oesophagus and 18 with short segments of SIM, one of whom had an associated intramucosal adenocarcinoma detected incidentally by histology. Three of the 20 cases with SIM exhibited novel microsatellite alleles, 2 with Barrett's oesophagus and 1 with short segment SIM and an associated adenocarcinoma. The 18 patients with short segments of SIM at the gastro oesophageal junction were significantly older than those without SIM. PMID- 10699919 TI - Human papillomavirus infection and invasive cervical cancer in Paraguay. AB - HPV types 16 and 18 have been categorized as human carcinogens based on their strong associations with cervical cancer in previous case-control studies. Recent IARC studies in the Philippines, Thailand and Morocco show strong associations between invasive cervical cancer and less common HPV types, including HPV 31, 33, 45, 51, 52 and 58. We present results of a further IARC case-control study conducted in Asuncion, Paraguay, to examine the association between specific HPV types and invasive cervical cancer as well as risk factors other than HPV. One hundred thirteen incident histologically confirmed invasive cervical cancer cases and 91 age-matched hospital controls were recruited. A standardized questionnaire was administered to investigate known and suspected risk factors for cervical cancer. For HPV status determination, cervical biopsy specimens from case subjects and exfoliated cervical cells from control subjects were obtained. HPV DNA was ascertained using a GP5+/6+ PCR-based assay capable of detecting more than 33 HPV types. Overall HPV prevalence was 97% in the cervical cancer cases and 20% in the control subjects. As a single infection, HPV 16 was the predominant type with a prevalence of 48% among case subjects and 5.5% among control subjects. Significant associations with the risk of cervical cancer were detected as follows: any HPV type (OR = 114; 95% CI: 36-361); HPV 16 (OR = 910); HPV 18 (infinite OR); HPV 31 (OR = 110); HPV 33 (OR = 261); HPV 45 (OR = 129); and HPV 58 (OR = 36). In the multivariate model, risk factors other than HPV significantly associated with cervical cancer risk were a higher number of lifetime sexual partners, lower educational status and never having had a Pap smear. Strong associations were found between invasive cervical cancer and specific HPV types 16, 18, 31, 33, 45 and 58. PMID- 10699920 TI - Clonal heterogeneity in sporadic melanomas as revealed by loss-of-heterozygosity analysis. AB - The major obstacle preventing effective treatment of melanoma is the biological heterogeneity of tumor cells. This study was performed to determine clonal genetic heterogeneity within primary melanoma and the evolution of these heterogeneous sub-clones during disease progression. DNA samples were obtained from 44 morphologically distinct areas identified within 10 primary tumors and from 15 metastases in the same patients. Loss of heterozygosity (LOH) analyses were performed using 17 microsatellite markers that mapped to chromosomes 6q, 9p, 10q and 18q, the most frequently deleted in melanoma. Of 10 primary tumors, 8 were revealed to have intratumoral genetic heterogeneity in terms of LOH of the 4 chromosome arms examined, 7 containing at least 2 different sub-clones harboring LOH of different chromosome areas, while the remaining one tumor showed prominent intratumoral genetic heterogeneity consisting of at least 6 genetically distinct sub-clones. LOH of 6q was detected only in a sub-set of multiple microdissected samples in most of the primary tumors, but was most frequently detected in metastases, suggesting that loss of this chromosome arm occurred late and played an important part in metastatic progression. Comparison of LOH between sub-clones within primary tumors and within metastases showed the divergence of metastatic clones from dominant populations within the primary tumor in 5 patients, whereas in the remaining three patients parent sub-clones were not identified, or constituted only a minor sub-population within the primary tumors. These results, showing considerable genetic heterogeneity in sporadic melanoma, have profound implications for the choice of future therapeutic strategies. PMID- 10699921 TI - Tobacco smoking, alcohol consumption and their interaction in the causation of hepatocellular carcinoma. AB - During a 4-year period from January 1995 to December 1998, blood samples and questionnaire data were obtained from 333 incident cases of hepatocellular carcinoma (HCC), as well as from 360 controls who were hospitalized for eye, ear, nose, throat or orthopedic conditions in Athens, Greece. Coded sera were tested for hepatitis B surface antigen (HBsAg) and antibodies to hepatitis C virus (anti HCV) by third-generation enzyme immunoassays, and information on smoking habits and beverage consumption was obtained. We found a significant dose-response, positive association between smoking and HCC risk [>/= 2 packs per day, odds ratio (OR)=2.5]. This association was stronger in individuals without chronic infection with either HBV or HCV (>/= 2 packs per day, OR=2.8). Consumption of alcoholic beverages above a threshold of 40 glasses per week increased the risk of HCC (OR=1.9). We also found evidence of a strong, statistically significant and apparently super-multiplicative effect of heavy smoking and heavy drinking in the development of HCC (OR for both exposures=9.6). This interaction was particularly evident among individuals without either HBsAg or anti-HCV (OR for both exposures=10.9). Coffee intake was not positively associated with HCC risk, but the reverse could not be excluded for the subgroup of chronically infected individuals. In conclusion, tobacco smoking and heavy alcohol consumption are associated with increased risk of HCC, especially when these 2 exposures occur together. PMID- 10699922 TI - Evidence against a role of general protein kinase C downregulation in skin tumor promotion. AB - Using isoenzyme-specific antibodies, we have performed an immunoblot analysis of the PKC isoenzyme pattern during the course of TPA-induced tumor promotion in the epidermis of NMRI mice. The TPA-sensitive PKC isoforms alpha, delta, straightepsilon, eta, nu (and TPA-insensitive PKCzeta), but not PKCbeta and gamma, were found to be expressed in both normal and neoplastic epidermis. The immune signals of all TPA-sensitive PKC isoforms were moderately and reversibly attenuated upon a single TPA treatment. Using different antibodies against PKCeta and PKCmu, this apparent downregulation could mainly be attributed to epitope changes of these enzymes, whereas for the other PKC species no such conclusion could be drawn. Except for PKCstraightepsilon, no substantial long-term attenuation of the immune signals of the other PKC isoforms occurred upon chronic phorbol ester treatment (i.e., 14 applications of 5 nmol TPA each over 7 weeks), which led to tumor development in initiated mouse skin. Specific PKC activity (related to tissue weight) was 40-50% lower in TPA-treated as compared with control epidermis whereby no clearcut difference was found between single and chronic TPA treatment. Benign and malignant skin tumors generated according to the initiation-promotion protocol did not exhibit consistent alterations in the immune pattern of the PKC isoenzymes with the exception of a decrease of PKCstraightepsilon and an increase of PKCmicro signal in carcinomas. Our data indicate that, in contrast with earlier assumptions, no general long-lasting PKC downregulation plays a critical role in skin tumor promotion. PMID- 10699923 TI - Vaccination with a recombinant vaccinia vaccine containing the B7-1 co stimulatory molecule causes no significant toxicity and enhances T cell-mediated cytotoxicity. AB - B7-1 is a co-stimulatory molecule that provides a second signal for T-cell activation. Several studies have demonstrated that vaccination with a vector containing genes encoding B7-1 and an antigen appears to be efficacious at promoting immune responsiveness to the antigen. To evaluate the safety of such a protocol and determine the effect of the B7-1 vector on immune responsiveness, female C57BL/6 mice were administered Wyeth wild-type vaccinia virus (V-WT) or V WT containing the gene for B7-1 (rV-B7-1) as a single s. c. injection or 3 monthly s.c. injections. Immunologic parameters were evaluated in half of the mice and general toxicity in the other half. Immunologic end points included determination of splenic lymphocyte phenotypes, mitogen-induced T- and B-cell proliferation, T-cell proliferation in response to alloantigens, cell-mediated cytotoxicity (CMC), natural killer cell activity and serum anti-nuclear antibody (ANA) titers. No significant signs of general toxicity were noted. The primary immunologic effect was an increase in the ability of spleen cells to lyse allogeneic targets and to proliferate in response to allogeneic stimulation. Numbers of splenic CD8(+) cells were also increased. These effects were more pronounced after 3 vaccinations than after a single vaccination. Minimal differences in ANA were observed between mice immunized with V-WT and rV-B7-1. In addition, no serum antibodies against B7-1 were detected in any mice. The data suggest that vaccination with rV-B7-1 augments CMC with minimal toxicity. PMID- 10699924 TI - Malignant transformation and EGFR activation of immortalized mouse liver epithelial cells caused by HBV enhancer-X from a human hepatocellular carcinoma. AB - We have previously observed that all human hepatocellular carcinomas (HCCs) from HBV carriers examined had the integrated X region. In this study, HBV DNA was isolated from an integration site in one HCC that had a single, very small integrated viral DNA including the X region, but it had no expression of X gene as poly(A)RNA. It was found that HBV DNA was present between alphoid repetitive sequences, and it included Enhancer and X regions, encompassing the adr sequence from 910 to 1811. Nucleotides for 8 amino acids at the 3' end, a stop codon of X gene and a poly(A) signal downstream of X gene were lost by integration, and nucleotides for 7 amino acids and a stop codon were substituted by a connected alphoid sequence. When this cloned HBV DNA was transfected with an expression vector to an immortalized mouse liver epithelial cell line, MLE-10, malignant transformation occurred. Transformants having expressed poly(A)RNA of the X gene showed anchorage-independent growth in soft agar and tumor formation in the subcutis of nude mice. The mRNA level of EGFR was found to be remarkably enhanced in X-transformed cells, in contrast with the absence of this mRNA in parental and ras-transformed MLE-10. Our data provide evidence that the Enhancer-X region alone is the key contributor to the malignant change of pre-malignant liver cells in HBV carriers through activation of some specific genes, such as EGFR. PMID- 10699925 TI - CD44s expression in human colon carcinomas influences growth of liver metastases. AB - CD44 is a family of cell-surface adhesion molecules which exist in several isoforms arising from mRNA alternative. Malignant transformation of colonic mucosa is associated with alterations in CD44 expression, which result in up regulation of high-molecular-weight CD44 isoforms and down-regulation of CD44s. We have demonstrated that stable transfection of CD44s into colon-carcinoma cell lines reduces their tumorigenicity. To understand the influence of CD44s expression on the metastatic potential of human colon carcinomas, we measured the ability of several different CD44s-transfected colon carcinomas to establish experimental liver metastases following splenic inoculation in mice. We observed that introduction of CD44s into 2 different human colon carcinoma cell lines, HT29 and KM12C6, resulted in reduced growth of liver metastases by as much as 75%. To explore the relationship between hyaluronate adhesion and metastasis, we transfected HT29 cells with cDNA encoding a mutant CD44s that does not bind to hyaluronate. HT29 transfectants expressing this mutant CD44s demonstrate an 84% reduction in growth of liver metastases, despite minimal binding to hyaluronate by the mutant CD44s. In concert, these results indicate that CD44s down regulation, which occurs with malignant transformation of colonic mucosa, is associated with enhanced growth of experimental liver metastases. Consequently, the functional consequences of CD44s down-regulation in colon carcinomas may be just as significant as the consequences of up-regulation of other CD44 isoforms. PMID- 10699926 TI - Late apoptotic effects of taxanes on K562 erythroleukemia cells: apoptosis is delayed upstream of caspase-3 activation. AB - The efficacy of taxanes on human leukemia cells is the object of intensive in vitro investigation concerning the influence of cell-type-specific characteristics on cytotoxic response to drugs. The present study dissects the response to taxanes of HL60 acute myelomonocytic leukemia and of K562 chronic myelogenous leukemia, in parallel over a 72-hr time-span. The kinetics of cytotoxicity following pulsed and continuous exposure to either taxol or taxotere showed a delayed response of K562 cells independently of dose and type of exposure. In K562 cells, apoptosis became evident at 48 hr and prominent at 72 hr of treatment. These events were mirrored by delayed kinetics of caspase-3 activation. Comparable microtubule targeting was demonstrated in HL60 and in K562 cell lines, as bcl-2 and raf-1 were phosphorylated following treatment with taxanes. These observations indicate that early activation processes were responsible for apoptosis, but that the delay was determined by other factors. In addition, cell-free-system experiments excluded the presence of excess nuclear and/or cytoplasmic inhibitory factors and demonstrated that K562 cells possess a fully competent caspase system which can be readily activated. Processing of caspase-3 pro-enzyme was in fact increased by addition of cytochrome c. These results extend to taxol and taxotere the notion that drug-induced apoptosis is delayed upstream of caspase-3 activation in K562 cells, that such kinetics is independent of drug concentration and exposure time, and that it is linked to intrinsic cellular characteristics mapping between bcl-2 phosphorylation and cytochrome c release. PMID- 10699927 TI - A truncated cytoplasmic topoisomerase IIalpha in a drug-resistant lung cancer cell line is encoded by a TOP2A allele with a partial deletion of exon 34. AB - To study the problem of acquired resistance to widely used anti-cancer drugs that target the 170 kDa topoisomerase IIalpha (topo IIalpha), a drug-resistant human small-cell lung cancer cell line, H209/VP, was selected in VP-16. H209/VP cells express reduced levels of the 170 kDa topo IIalpha that is localized normally in the nucleus and also express lower levels of a 160 kDa topo IIalpha-related protein that is located predominantly in the cytoplasm. Band depletion immunoblotting experiments suggest that the H209/VP nuclear 170 kDa topo IIalpha is able to form ternary complexes with DNA and VP-16 in intact cells, but the ability of the cytoplasmic 160 kDa protein to do so is greatly diminished. Sequence analysis of the 3; end of the H209/VP mutant topo IIalpha mRNA and the TOP2A gene indicates that the mRNA is missing 200 nt that corresponds to exon 34 because the partial loss of the minimal 3; splice-acceptor sequence at the beginning of exon 34 results in splicing of exon 33 to exon 35. The protein predicted to be encoded by this mutant mRNA does not contain the COOH-terminal 109 amino acids of the wild-type enzyme that we have demonstrated contain a strongly functional nuclear localization signal sequence. Consequently, our data explain both the size and the cytoplasmic localization of the H209/VP mutant topo IIalpha. The mutant TOP2A allele in H209/VP cells differs from those in previously characterized cell lines with cytoplasmic topo IIalpha and extends the number of types of resistance-associated deletions in this region to 4. These findings indicate that this region of the TOP2A gene may be a hot spot for mutations. PMID- 10699928 TI - A new human tumor-associated antigen (TLP) is naturally expressed in rat DHD-K12 colorectal tumor cells. AB - Renewed interest in cancer immunotherapy has been raised by the availability of a variety of tumor-associated antigens and animal models. We have recently described the presence of a new antigen, TLP, in sera and cancer tissue from lung and colorectal cancer patients. In order to develop an experimental model suitable for preclinical studies on cancer vaccines, we investigated the presence of TLP antigen in vitro, in the DHD-K12 cell line and in vivo, in metastases induced in syngeneic BDIX rats by DHD-K12 cell injection. TLP was not detected in any tissue of healthy rats nor in normal tissues of tumor-bearing rats. This is in agreement with our previous studies, in which we had demonstrated that TLP is expressed in human colorectal cancer and adenomas but not in normal colonic mucosa. Our results indicate TLP as a possible human tumor-specific antigen naturally expressed in DHD-K12 tumor syngeneic to immunocompetent BDIX rats. PMID- 10699929 TI - Galectin-3 overexpression protects from apoptosis by improving cell adhesion properties. AB - Galectin-3 is a carbohydrate-binding protein endowed with affinity for beta galactosides. It plays a role in cell-cell and cell-matrix interactions. Furthermore, it has been hypothesized to be involved in tumor progression and metastasis. To address the role of galectin-3 in the invasive and metastatic processes, we stably overexpressed galectin-3 in human breast carcinoma cell lines, and we evaluated the influence of elevated galectin-3 expression on several cell features, including cellular homotypic and heterotypic interactions and cell survival. No differences in various parameters related with cell growth features and proliferation were detected. By contrast, we found that galectin-3 overexpressing cells, with respect to low galectin-3 expressing cells, exerted: (1) a significantly enhanced adhesion to laminin, fibronectin and vitronectin exerted both directly or via increased expression of specific integrins, e.g., alpha-4 and beta-7; (2) a remodeling of those cytoskeletal elements associated with cell spreading, i.e., microfilaments; (3) an enhanced survival upon exposure to different apoptotic stimuli, such as cytokine and radiation. Collectively, our results indicate that overexpression of galectin-3 may play a role in tumor cell invasion and metastasis by specifically influencing cell adhesion to the extracellular matrix. This may confer selective survival advantage and resistance to the particular homeless-induced apoptosis called anoikia. PMID- 10699930 TI - FGF-2 isoforms of 18 and 22.5 kDa differentially modulate t-PA and PAI-1 expressions on the pancreatic carcinoma cells AR4-2J: consequences on cell spreading and invasion. AB - Pancreatic tumors overexpress FGF-2 and t-PA, but the implication of the growth factor in t-PA synthesis and t-PA-dependent tumor invasion remains unknown. FGF-2 is present in different isoforms: The 18 kDa FGF-2 is secreted, while the 22.5 kDa one is nuclearized and exerts intracrine regulations bypassing cell-surface FGF receptors. Rat pancreatic carcinoma AR4-2J cells producing either the 18 or the 22.5 kDa FGF-2 after transfection with FGF-2 cDNAs have been used to analyze the role of FGF-2 in t-PA expression and t-PA-related cell spreading. The 22.5 kDa FGF-2 reduced t-PA and PAI-1 synthesis 2-fold. Addition of recombinant 18 kDa FGF-2 (rFGF-2) to cell cultures resulted in increased t-PA and decreased PAI-1 expression. By contrast, rFGF-2 did not significantly modify t-PA synthesis in cells producing the 22.5 kDa FGF-2. Cell spreading was t-PA-dependent. Furthermore, cells producing the 22.5 kDa FGF-2 migrated less than control cells and cells producing the 18 kDa FGF-2. Overall, our data show that secretory FGF-2 is involved in t-PA synthesis by pancreatic cancer cells and facilitates cell spreading. The 22.5 kDa FGF-2 exerts opposite effects by decreasing t-PA expression in basal conditions and during rFGF-2 stimulation. Since the expression of the 22.5 kDa FGF-2 is under specific controls, its up-regulation might have the potential to reduce spreading of pancreatic cancer cells. PMID- 10699931 TI - Nk4, a new HGF/SF variant, is an antagonist to the influence of HGF/SF on the motility and invasion of colon cancer cells. AB - Hepatocyte Growth Factor/Scatter Factor (HGF/SF) is a heterodimeric molecule that plays a key role in the regulation of migration, invasion and angiogenesis in cancer, via activation of its receptor, c-met. HGF/SF is composed of an alpha chain, containing the N-terminal hairpin domain and 4 kringle domains, plus the serine protease-like beta-chain. We have examined here the properties of NK4, an HGF variant containing the N-terminal hairpin plus the 4 kringle domains, on tumour cell proliferation, dissociation and invasion using human colorectal cancer cells (HT115). The expression of HGF/SF and its receptor was also examined by RT-PCR and Western blotting. Analysis revealed NK4 to be an HGF/SF antagonist that, at a wide range of concentrations, did not exert any biological effects of its own. HT115 cells were shown to express the HGF/SF receptor mRNA and protein. HGF/SF-induced receptor tyrosine phosphorylation was suppressed, in a dose dependent manner, upon addition of NK4, whereas the addition of NK4 alone caused no phosphorylation. Tumour cell motility was induced by HGF and inhibited by NK4. Furthermore, HGF/SF induced the invasion of cells through Matrigel basement membrane components, and again this induced invasion was suppressed by NK4. Our results show that the ability of HGF/SF to stimulate tumour cell motility and invasion, properties required for metastatic spread, can be inhibited by NK4. Thus, NK4 may have an important role in the control of cancer metastasis. PMID- 10699932 TI - Catalytic activity of an in vivo tumor targeted anti-CEA scFv::carboxypeptidase G2 fusion protein. AB - Antibody-directed enzyme prodrug therapy (ADEPT) targets an enzyme selectively to a tumor where it converts a relatively non-toxic prodrug to a potent cytotoxic drug. Previous clinical work using antibody-enzyme chemical conjugates has been limited by the moderate efficiency of tumor targeting of these molecules. To address this a recombinant fusion protein composed of MFE-23, an anti carcinoembryonic antigen (CEA) single chain Fv (scFv) antibody, fused to the amino-terminus of the enzyme carboxypeptidase G2 (CPG2) has been constructed to achieve ADEPT in CEA-producing tumors. MFE-23::CPG2 fusion protein was overexpressed in Escherichia coli and purified using CEA affinity chromatography. Efficacy of MFE-23::CPG2 delivery to tumors in vivo was assessed by measuring catalytic activity after intravenous injection of purified MFE-23::CPG2 into nude mice bearing CEA-positive LS174T human colon adenocarcinoma xenografts. Recombinant MFE-23::CPG2 cleared rapidly from circulation and catalytic activity in extracted tissues showed tumor to plasma ratios of 1.5:1 (6 hr), 10:1 (24 hr), 19:1 (48 hr) and 12:1 (72 hr). (125)I-MFE-23::CPG2 was retained in kidney, liver and spleen but MFE-23::CPG2 catalytic activity was not, resulting in excellent tumor to normal tissue enzyme ratios 48 hr after injection. These were 371:1 (tumor to liver), 450:1 (tumor to lung), 562:1 (tumor to kidney), 1,477:1 (tumor to colon) and 1,618:1 (tumor to spleen). Favorable tumor : normal tissue ratios occurred at early time points when there was still 21% (24 hr) and 9.5% (48 hr) of the injected activity present per gram of tumor tissue. The high tumor concentrations and selective tumor retention of active enzyme delivered by MFE 23::CPG2 establish that this recombinant fusion protein has potential to give improved clinical efficiency for ADEPT. PMID- 10699933 TI - Over-expression of erbB-2/neu is paralleled by inhibition of mouse-mammary epithelial-cell differentiation and developmental apoptosis. AB - The erbB-2/neu oncogene is frequently over-expressed in many different tumors in humans, including those of breast and ovary. The oncogene encodes a receptor tyrosine kinase closely related to the epidermal-growth-factor receptor. We studied effects on differentiation and cell death of erbB-2/neu during mammary gland development in transgenic mice expressing an activated, oncogenic rat erbB 2/neu gene controlled by the mammary-gland-specific promoter from mouse-mammary tumor virus (MMTV-LTR). Transgenic animals develop mammary cancer after repeated pregnancies and lactation. We present evidence that over-expression of erbB-2/neu in these mice is restricted to tumor cells. Tumor cells fail to differentiate and express milk proteins such as beta-casein and whey acidic protein (WAP) during lactation. Epithelial-cell apoptosis during normal involution is characterized by non-random DNA degradation into oligonucleosomal fragments. Tumor cells were mostly refractory to this developmentally controlled programmed cell death. Distinct areas within tumors, however, showed spontaneous cell death as measured by in situ TUNEL staining that co-localized with caspase-3-like activity. Our results indicate that the control of developmental cell death during involution is disturbed in erbB-2/neu-induced tumors although cell death and caspase activation can take place. PMID- 10699934 TI - The effect of 2-methoxyoestrone-3-O-sulphamate on the growth of breast cancer cells and induced mammary tumours. AB - 2-Methoxyoestrogens are emerging as a new class of drug that can inhibit tumour growth and angiogenesis. As sulphamoylation of oestrogens enhances their potency and bioavailability we have synthesized 2-methoxyoestrone-3-O-sulphamate (2 MeOEMATE) and compared its ability to inhibit the proliferation of breast cancer cells with that of 2-methoxyoestrone (2-MeOE1). 2-MeOEMATE (1 microM) inhibited the growth of oestrogen receptor positive MCF-7 breast cancer cells by 52% whereas 2-MeOE1 had little effect at this concentration. 2-MeOEMATE also inhibited the growth of oestrogen receptor negative MDA-MB-231 breast cancer cells. Exposure of cells to 2-MeOEMATE caused them to round up and become detached suggesting that this compound may induce cells to undergo apoptosis. Cell cycle analysis revealed that 2-MeOEMATE caused cells to arrest in the G(2)/M phase with the increase in G(2)/M arrested cells being detectable by 12 hr. Exposure of MCF-7 cells to 2 L-MeOEMATE for 24 hr followed by culture in drug free medium for 24 hr did not reverse the arrest of cells in the G(2)/M phase. TUNEL analysis confirmed that 2-MeOEMATE induced apoptosis in a significant proportion of treated MCF-7 cells. In an in vivo study, employing nitrosomethylurea-induced mammary tumours in intact rats, 2-MeOE1 (20mg/kg/d, p.o. for 11 days) had little effect on tumour growth. In contrast, the same dose of 2-MeOEMATE resulted in the almost complete regression of 2/3 tumours over an 11-day period. We conclude that 2-MeOEMATE should have considerable therapeutic potential for the treatment of breast tumours. PMID- 10699935 TI - Sensitivity to DNA cross-linking chemotherapeutic agents in mismatch repair defective cells in vitro and in xenografts. AB - Together with tolerance to killing induced by methylating agents, loss of mismatch repair (MMR) has previously been found to be associated with hypersensitivity to the DNAcross-linking agent 1-(2-chloroethyl)-3-cyclohexyl nitrosourea(CCNU) in several human tumor cell lines (Aquilina et al., 1998). Here, we have investigated whether MMR might act as an efficient repair pathway and provide protection against the clastogenicity induced by CCNU and whether the hypersensitivity of MMR-defective cells is extended to other cross-linking agents. An increase in cell killing and in the frequency of micronuclei was observed after CCNU exposure in 2 hPMS2-defective clones (clones 6 and 7) compared with the parental HeLa cells. Introduction of a wild-type copy of chromosome 7 in clone 7 led to re-expression of the hPMS2 protein and brought survival and chromosomal damage upon CCNU exposure to parental levels. Our data indicate that MMR protects against the clastogenic damage induced by this drug. The hPMS2-defective HeLa cells were also hypersensitive to killing by mitomycin C. Mitomycin C sensitivity was confirmed in an hMLH1-defective clone derived from Raji cells and in msh2-defective mouse embryo fibroblasts derived from knock-out mice. hPMS2-defective and parental HeLa cells were transplanted into nude mice, and the animals were treated with mitomycin C. While parental cell growth rate was unaffected, the growth of MMR-defective tumor was significantly reduced. Our results indicate that the in vitro hypersensitivity to mitomycin C conferred by loss of MMR is paralleled in vivo and may have implications for the chemotherapy of MMR-defective tumors. PMID- 10699936 TI - Lack of expression for the suppressor PML in human small cell lung carcinoma. AB - The promyelocytic leukemia (PML) gene, which encodes a transformation and growth suppressor, was first identified at the chromosomal translocation break point t(15;17) in acute promyelocytic leukemia (PML). To determine if the PML gene might be involved in other neoplasias such as lung cancer, PML expression was analyzed by immunohistochemical staining and in situ hybridization. Considerable PML protein expression in the PML-oncogenic domain (POD) structure was found in adenocarcinomas (ADC) and squamous cell carcinomas (SCC) of the lung, but was almost completely absent in all the small cell lung carcinomas (SCLC) examined. In situ hybridization showed that both mRNA and DNA of PML were present in SCLC and in normal lung, suggesting that the decreased protein expression was due to either a defect in translation or protein instability, rather than the consequence of decreased transcription or gene deletion. Double staining showed that PML expression was inversely correlated with the proliferation marker Ki-67 and positively correlated with levels of apoptotic cells in these tumors. To determine if the precursor cells of SCLC, the neuroendocrine-producing cells, express PML, double labeling was performed with PML and chromogranin A, a bio marker for neuroendocrine cells. Neuroendocrine cells from normal tissues were found to be PML positive, indicating that the lack of PML protein in SCLC is associated with the tumorigenic phenotype and is not the result of cell-lineage specificity. Thus, the decreased PML expression may play an important role in SCLC development. PMID- 10699937 TI - Sequence analysis of the mismatch repair gene hMSH6 in the germline of patients with familial and sporadic colorectal cancer. AB - To evaluate the involvement of hMSH6 in colorectal cancer, the complete coding sequence and flanking intron regions of the gene were analyzed by DNA sequencing in 10 patients fulfilling Bethesda Guidelines for colorectal tumors and 10 patients with sporadic colorectal carcinoma. In addition, 10 mono- and 10 dinucleotide repeat markers were analyzed for microsatellite instability. A protein-truncating T insertion at codon 218 was identified in the index person of a hereditary non-polyposis colorectal cancer (HNPCC)-like kindred and was accompanied by a somatic T deletion in the tumor. The tumor of this patient was positive for mono- but negative for dinucleotide repeat instability and lacked allelic losses at loci frequently affected in colorectal carcinomas. A novel amino acid change, F340S, was found in a patient with sporadic colon and breast cancer and leukemia but was not detected in 246 chromosomes from healthy anonymous blood donors. In addition, we describe 2 silent and 15 intronic sequence variants not previously reported. Although the frequency is low, we present further evidence for hMSH6 germline mutations that predispose patients to HNPCC-like phenotypes and suggest that mono- and dinucleotide repeat instability testing may be useful for distinguishing between individuals harboring an hMSH2 or hMLH1 mutation and a mutation of the hMSH6 gene. PMID- 10699938 TI - Microsatellite alterations in urinary sediments from patients with cystitis and bladder cancer. AB - Microsatellite instability (MIN) and loss of heterozygosity (LOH) in bladder cancer have been suggested for diagnosis and follow-up of bladder cancer based on urinary sediments, reflecting tumor alterations. We have examined 6 microsatellites in urine sediments from 11 patients with transitional-cell carcinomas (TCC) and in 31 patients with benign prostatic hyperplasia (BPH), 22 of whom had cystitis. In the TCC patients, tumor tissue was available for comparison with urine. Microsatellites were amplified by PCR and compared with leukocyte DNA from the same individual in silver-stained gels. Altered mobility of bands, new bands and loss of bands were scored. We found MIN and LOH at relatively high frequency in markers from chromosomes 8 and 14 in urine from patients with TCC, but also in BPH patients who had cystitis. Even control patients with BPH without cystitis showed some instability and some losses. Novel bands in urine occurred significantly more often among TCC patients than among BPH patients with or without cystitis (p < 0.001). Band shifts in urine appeared to be more associated with BPH plus cystitis than with TCC. The alterations we found in urine from patients with bladder cancer did not always reflect those found in their tumors, the occurrence of novel bands being significantly higher (p < 0.008) in tumor tissue than in corresponding urine. In conclusion, microsatellite alterations in urine are indicators not only of malignancy but also of inflammatory conditions. PMID- 10699939 TI - Hybrid cell vaccination for cancer immune therapy: first clinical trial with metastatic melanoma. AB - Hybrid cell vaccination is a new cancer immune therapy approach that aims at recruiting T cell help for the induction of tumour specific cytolytic immunity. The vaccines are generated by fusion of the patients' tumour cells with allogeneic MHC class II bearing cells to combine the tumour's antigenicity with the immunogenicity of allogeneic MHC molecules. Safety and anti-tumour activity of this treatment were assessed in a clinical trial that has yielded one complete and one partial remission, and 5 cases of stable disease among 16 patients with advanced stage metastatic melanoma. As evidenced by histology, the vaccination induced T cell relocation into tumour nodules. Stable disease could be maintained by repeated booster injections for more than 24 months in some patients. The side effects were minor. Occasional occurrences of vitiligo spots after vaccination were indicative of a restricted therapy induced auto-immune reactivity. The results suggest that hybrid cell vaccination is a safe cancer immune therapy potentially effective for induction of acute anti-tumour response as well as long term maintenance. PMID- 10699940 TI - Site-specific risk of cutaneous malignant melanoma and pattern of sun exposure in New Zealand. AB - The site-specific relationship between melanoma and sun exposure was investigated in the high-risk region of New Zealand. Age and latitude of residence were used as biological and geographical proxy measures of exposure for the 16,117 newly incident cases and 3,150 death records reported between 1968 and 1993. Age standardized rates were the highest for the trunk in males and for the lower limbs in females, but once body surface area was accounted for, highest rates were found on fully exposed sites, particularly the ears in men. Melanomas occurred at a substantially younger age on intermittently exposed sites than chronically exposed ones (difference of about 13 and 27 years for men and women, respectively, between the trunk and the face). Age and latitude were found to influence melanoma rates in a sex- and site-specific fashion. For heavily exposed body areas, incidence rates increased more modestly with age before age 50 than after. In contrast, sharp increases in risk occurred from early age for episodically exposed sites with a reversal of trend observed among the elderly. For males, the magnitude of the latitude gradient was about 65% (incidence) and 50% (mortality) greater for body areas most intermittently exposed compared with those with a least intermittent pattern of exposure. The latitude gradient was steeper for males than females and for incidence than mortality, regardless of the pattern of site exposure. Sex- and age-specific differences in risk were largely explained by the varying patterns of exposure. These results confirm that intermittent exposure is probably more effective than continuous exposure in producing an early onset of melanoma. Reducing the episodes of acute exposure remains a paramount aspect to melanoma prevention strategies. PMID- 10699941 TI - Putative chromosomal deletions on 9P, 9Q and 22Q occur preferentially in malignant gastrointestinal stromal tumors. AB - To characterize the type of genetic alterations in gastrointestinal stromal tumors (GISTs), we performed a comprehensive allelotype study of 14 GISTs (2 benign, 7 borderline and 5 malignant) by polymerase-chain-reaction and loss-of heterozygosity (PCR-LOH) analysis using 102 microsatellite markers, and compared the results with comparative-genomic-hybridization (CGH) analysis. Among the 38 evaluated chromosomal arms, 16 (42.1%) showed LOH in at least one patient. Most frequent LOH was observed at chromosome 14p and 14q (9/14, 64%) and this was demonstrated in all types of GISTs (50% in benign, 71% in borderline and 80% in malignant). Additional chromosomal deletions were found in several chromosomal arms. Among them, deletions on chromosomal arms of 22q (3/14, 21.4%), 9p (2/14, 14.3%) and 9q (2/14, 14.3%) were the most frequent, and were detected only in malignant GISTs both by PCR-LOH and by CGH analysis. Additionally, 2 malignant GISTs with LOH on 9p showed homozygous deletions in the restricted area of 9p by multiplex PCR-LOH analysis. Thus, several putative chromosomal changes were preferentially present in malignant GISTs but rare in benign and borderline GISTs. These findings suggest that accumulated chromosomal changes may contribute to the progression and/or malignant transformation of GISTs. PMID- 10699942 TI - Non-melanoma skin cancer may be a marker of poor prognosis in patients with non Hodgkin's lymphoma. AB - According to recent results, patients with non-melanoma skin cancers are at increased risk of developing non-Hodgkin's lymphoma (NHL). The prognostic significance of this association is unknown. Two cohorts of patients with a first diagnosis of non-melanoma skin cancer and a subsequent diagnosis of either NHL (n = 170) or colon cancer (n = 435) were established using national cancer registry data in Denmark. Two other cohorts of patients in whom NHL (n = 600) or colon cancer (n = 1,541) was the patients' first known malignancy served as comparison groups. Mortality rates were compared using Cox's regression analysis. Among patients younger than 80 years at NHL diagnosis, a history of non-melanoma skin cancer was associated with significantly increased mortality [relative risk (RR) = 1.54; 95% confidence interval: 1.19-1.99]. This association was present in both men (RR = 1.38; 1.02-1.86) and women (RR = 2.15; 1.31-3.54) and was similar after both major subtypes of non-melanoma skin cancer. Overall, antedating non-melanoma skin cancer had no prognostic significance for colon cancer patients (RR = 1.00; 0.84-1.18). Whatever the underlying mechanism, our observation has potential clinical implications. If substantiated in other settings, NHL patients with prior non-melanoma skin cancer may constitute a subgroup of lymphoma patients in need of particular therapeutic attention. PMID- 10699943 TI - Gamma linolenic acid with tamoxifen as primary therapy in breast cancer. AB - Gamma linolenic acid (GLA) has been proposed as a valuable new cancer therapy having selective anti-tumour properties with negligible systemic toxicity. Proposed mechanisms of action include modulation of steroid hormone receptors. We have investigated the effects of GLA with primary hormone therapy in an endocrine sensitive cancer. Thirty-eight breast cancer patients (20 elderly Stage I-II, 14 locally advanced, 4 metastatic) took 8 capsules of oral GLA/day (total = 2.8 g) in addition to tamoxifen 20 mg od (T+GLA). Quality and duration of response were compared with matched controls receiving tamoxifen 20 mg od alone (n = 47). Serial tumour biopsies were taken to assess changes in oestrogen receptor (ER) and bcl-2 expression during treatment. GLA was well tolerated with no major side effects. T+GLA cases achieved a significantly faster clinical response (objective response vs. static disease) than tamoxifen controls, evident by 6 weeks on treatment (p = 0.010). There was significant reduction in ER expression in both treatment arms with T+GLA objective responders sustaining greater ER fall than tamoxifen counterparts (6-week biopsy p = 0.026; 6-month biopsy p = 0.019). We propose GLA as a useful adjunct to primary tamoxifen in endocrine-sensitive breast cancer. The effects of GLA on ER function and the apparent enhancement of tamoxifen-induced ER down-regulation by GLA require further investigation. PMID- 10699944 TI - Photodynamic therapy of vulvar intraepithelial neoplasia using 5-aminolevulinic acid. AB - Photodynamic (PDT) therapy is a relatively new technique with unique properties that make it attractive for the local treatment of superficial epithelial disorders. The objective of this study was to investigate the clinical response of PDT with the photosensitizing agent 5-aminolevulinic acid (5-ALA) in patients with vulvar intraepithelial neoplasia (VIN) grades 1 to 3. Twenty-five patients with 111 lesions of VIN 1-3 were topically sensitized with 10 ml of a 20% solution of 5-ALA and treated with 57 cycles of laser light at 635 nm (100 J/cm(2)). Seventy (64%) of the 111 VIN lesions regressed after various PDT cycles. A complete response was achieved in 13 patients (52%) with 27 lesions. All patients with VIN 1 and mono- and bifocal VIN 2-3 showed complete clearance. However, a complete response could be achieved in only 4 (27%) of 15 women with multifocal VIN 2-3, whereas a partial response was noted in 9 of these patients with a total of 70 lesions, out of which 44 (63%) lesions disappeared. No response was seen in 2 patients with multifocal VIN 3. Histological assessment of the fluorescence-directed biopsies revealed that increased pigmentation and hyperkeratosis of the lesions were associated with low response rates. PDT using 5-ALA represents an alternative treatment modality for VIN which is easy to perform and has the advantage of minimal tissue destruction, low side effects and excellent cosmetic results. However, multifocal VIN disease with pigmented and hyperkeratinic lesions remains difficult to treat. PMID- 10699945 TI - Identification of two distinct regions of allelic imbalance on chromosome 18Q in metastatic prostate cancer. AB - Like most cancers, prostate cancer (CaP) is believed to be the result of the accumulation of genetic alterations within cells. Previous studies have implicated numerous chromosomal regions with elevated rates of allelic imbalance (AI), using mostly primary CaPs with an unknown disease outcome. These regions of AI are proposed sites for tumor suppressor genes. One of the regions previously implicated as coding for at least one tumor suppressor gene is the long arm of chromosome 18 (18q). To confirm this observation, as well as to narrow the critical region for this putative tumor suppressor, we analyzed 32 metastatic CaP specimens for AI on chromosome 18q. Thirty-one of these 32 specimens (96.8%) exhibited AI at one or more loci on chromosome 18q. Our analysis using 17 polymorphic markers revealed statistically significant AI on chromosome 18q at 3 markers, D18S35, D18S64 and D18S461. Using these markers as a guide, we have been able to identify 2 distinct minimum regions of AI on 18q. The first region is between the genetic markers D18S1119 and D18S64. The second region lies more distal on the long arm of the chromosome and is between the genetic markers D18S848 and D18S58. To determine if 18q loss is a late event in the progression of CaP, we also examined prostatic intraepithelial neoplasia (PIN) and primary prostate tumors from 17 patients for AI with a subset of 18q markers. We found significantly higher AI in the metastatic samples. Our results are consistent with 18q losses occurring late in CaP progression. PMID- 10699946 TI - Induction of antibodies against GD3 ganglioside in melanoma patients by vaccination with GD3-lactone-KLH conjugate plus immunological adjuvant QS-21. AB - The gangliosides GD3, GD2 and GM2 are expressed on the cell surface of malignant melanomas, GD3 being the most abundant. We have shown that immunization of melanoma patients with GM2 adherent to Bacillus Calmette-Guerin (GM2/BCG) induced an IgM antibody response. Vaccines containing GM2-keyhole limpet hemocyanin (KLH) conjugate and the immunological adjuvant QS-21 induced a higher titer IgM response and consistent IgG antibodies. Patients with antibodies against GM2 survived longer than patients without antibody. On the other hand, our previous trials with GD3/BCG, GD3 derivatives including GD3-lactone (GD3-L)/BCG failed to induce antibodies against GD3. In our continuing efforts to induce antibody against GD3, we have immunized groups of 6 melanoma patients with GD3-KLH or GD3 L-KLH conjugates containing 30 microg of ganglioside plus 100 microg of QS-21 at 0, 1, 2, 3, 7 and 19 weeks. Prior to vaccination, no serological reactivity against GD3 or GD3-L was detected. After immunization, IgM and IgG antibodies were detected against both GD3 and GD3-L in the GD3-L group exclusively. The GD3 L-KLH vaccine induced IgM titers against GD3-L of 1:40-1/1,280 in all patients and IgG titers of 1/160-1/1,280 in 4 patients. These antibodies also strongly cross-reacted with GD3. ELISA reactivity was confirmed by immune thin-layer chromatography on GD3 and melanoma extracts. Sera obtained from 4 of these 6 patients showed cell surface reactivity by FACS and from 2 showed strong cell surface reactivity by immune adherence (IA) assay and complement lysis against the GD3 positive cell line SK-Mel-28. PMID- 10699947 TI - Co-culture with human fibroblasts increases the radiosensitivity of MCF-7 mammary carcinoma cells in collagen gels. AB - The growth and differentiation of normal and neoplastic epithelial cells may be regulated by the presence of adjacent normal tissues and cells, particularly stromal fibroblasts. However, the influence of normal fibroblast-tumor cell interactions on the response of malignant epithelial cells to radiation has not been adequately investigated nor has the possible role played by a 3-D environment in such modulation. We addressed this question by embedding MCF-7 mammary carcinoma cells into a collagen lattice, alone or mixed with HSF human dermal fibroblasts, and kept the gels anchored to the plastic surface or suspended in the culture medium. Some gels served as controls and others were irradiated with 6 MV photons fractionated into 3 daily doses totaling 5 or 10 Gy. After 2 or 7 days from the last treatment (7 or 12 days in culture, respectively), gels were processed in 1 of 2 ways: overall cell survival was determined by the MTT assay, while the survival of MCF-7 cells was selectively detected by a clonogenicity assay. Under these experimental conditions, we found that, in the presence of HSF fibroblasts, the growth of MCF-7 cells was restrained and radiosensitivity increased compared with MCF-7 cells cultured alone. For example, while the average number of MCF-7 foci/gel recovered from control gels with MCF-7 cells alone was 2,460 on day 7 and 3, 290 on day 12 of culture, it was 4 to 5 times lower (p < 0.001) in control gels with mixed MCF-7 and HSF cells. Radiation affected severely the survival of MCF-7 cells in all experimental groups but not sufficiently to mask the differences. For example, following exposure to the low dose of 5 Gy, the average number of MCF-7 foci/gel recovered from MCF-7-containing gels was 590 on day 7 and 329 on day 12 of culture, whereas numbers from the gels containing mixed MCF-7 and HSF cells were only 218 and 73, respectively (p < 0. 003 in both cases). HSF fibroblasts did not grow in our system, but they contracted strongly anchored and floating gels. PMID- 10699948 TI - Intracellular triggering of Fas, independently of FasL, as a new mechanism of antitumor ether lipid-induced apoptosis. AB - Antitumor ether lipid 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine (ET 18-OCH(3); edelfosine) induces apoptosis in cancer cells, sparing normal cells. We have found that the apoptotic action of ET-18-OCH(3) required drug uptake and Fas in the target cell. Failure to accomplish one of these requirements prevents cell killing by the ether lipid. In human lymphoid leukemic cells, ET-18-OCH(3) does not promote Fas or FasL expression and ET-18-OCH(3)-induced apoptosis is not inhibited by pre-incubation with an anti-Fas blocking antibody that abrogates cell killing mediated by Fas/FasL interactions. ET-18-OCH(3)-resistant normal human Fas-positive fibroblasts do not incorporate ET-18-OCH(3), but undergo apoptosis upon ET-18-OCH(3) microinjection. Murine fibroblasts L929 and L929-Fas, stably transfected with human Fas cDNA, do not incorporate ET-18-OCH(3) and are resistant to its action when added exogenously. Microinjection of ET-18-OCH(3) induces apoptosis in L929-Fas cells, but not in wild-type L929 cells. Confocal laser scanning microscopy shows that ET-18-OCH(3) induces Fas clustering and capping during triggering of ET-18-OCH(3)-induced apoptosis. Microinjection induced apoptosis and Fas clustering are specific for the molecular structure of ET-18-OCH(3). Our data indicate that ET-18-OCH(3) induces apoptosis via Fas after the ether lipid is inside the cell, and this Fas activation is independent of the interaction of Fas with its natural ligand FasL. This explains the selective action of ET-18-OCH(3) on tumors since only cancer cells incorporate sufficient amounts of the drug. PMID- 10699949 TI - Resistance to methotrexate in SKOV-3 cell lines after chronic exposure to carbamazepine is associated with a decreased expression of folate receptor. AB - The detrimental effect of chronic administration of carbamazepine (CBZ) on serum and erythrocyte folates of patients has been extensively analyzed. However, at present, no data have been reported on the effect of CBZ on the intracellular content and activity of antimetabolite cytotoxic agents that can be used together with CBZ in the treatment of cancer patients. To investigate this issue, we chronically exposed in vitro the human ovarian cancer cell line SKOV-3 (grown under physiological folate concentrations) to CBZ, thus selecting SKOV-CBZ clones (SKOV-CBZ-50-2, SKOV-CBZ-50-5 and SKOV-CBZ-100-2) able to grow in the continuous presence of the antiepileptic drug. All of the SKOV-CBZ clones were more resistant to methotrexate (MTX) than the parental cells. MTX resistance index, as determined by the ratio between MTX concentrations inhibiting cell growth by 50% (MTX IC(50)) in SKOV-CBZ clones and in the parental cells, ranged between 3- and 5-fold. This resistance was related to a reduced intracellular content of MTX. No alteration activity of the cellular enzymes directly involved in MTX cytotoxicity (dihydrofolate reductase, thymidylate synthase [TS] and folylpolyglutamate synthetase) was observed. SKOV-CBZ clones were cross-resistant to the TS inhibitors tomudex and CB3717, but not to the TS inhibitor 5-fluoro-deoxy uridine and other antineoplastic drugs (cisplatin, doxorubicin, vincristine and taxol), whose cellular uptake is derived from transmembrane transport mechanisms different from folate receptor alpha (FRalpha) or reduced folate carrier (RFC). FRalpha mRNA and protein levels were significantly lower in SKOV-CBZ clones than in the parental cells. The reduced level of FRalpha in SKOV-CBZ clones was associated with a decreased binding capacity of folic acid. No variation of mRNA RFC expression in the SKOV-CBZ clones as compared to the parental cells was observed. Thus, after chronic exposure to CBZ, SKOV-CBZ clones develop resistance towards MTX due to defective MTX uptake. PMID- 10699950 TI - Apigenin inhibits endothelial-cell proliferation in G(2)/M phase whereas it stimulates smooth-muscle cells by inhibiting P21 and P27 expression. AB - Apigenin is a plant flavonoid that is thought to play a role in the prevention of carcinogenesis. However, its mechanism of action has not yet been elucidated. Because of the importance of angiogenesis in tumor growth, we investigated the effect of apigenin on endothelial and smooth-muscle cells in an in vitro model. Apigenin markedly inhibited the proliferation, and, to a lesser degree, the migration of endothelial cells, and capillary formation in vitro, independently of its inhibition of hyaluronidase activity. In contrast, it strongly stimulated vascular smooth-muscle-cell proliferation. The molecular mechanisms of apigenin activity were analyzed in these 2 types of cells. Our results show that apigenin inhibits endothelial-cell proliferation by blocking the cells in the G(2)/M phase as a result of the accumulation of the hyperphosphorylated form of the retinoblastoma protein. Apigenin stimulation of smooth-muscle cells was attributed to the reduced expression of 2 cyclin-dependent kinase inhibitors, p21 and p27, which negatively regulate the G(1)-phase cyclin-dependent kinase. PMID- 10699951 TI - High frequency of allele-specific down-regulation of HLA class I expression in uveal melanoma cell lines. AB - Uveal melanoma is the most common primary intra-ocular tumor in adults and has a high mortality rate due to liver metastases, for which no effective treatment is available. To investigate whether immunotherapy might be feasible in uveal melanoma, the HLA class I surface expression of 6 uveal melanoma cell lines was analyzed by flow cytometry using a broad panel of allele-specific monoclonal antibodies. To up-regulate HLA expression, cells were also cultured with IFN alpha or -gamma. In general, expression of HLA-A alleles was high (except for cell line EOM-3) and could be further up-regulated by both IFN-alpha and -gamma. In cell line EOM-3, IFN-gamma treatment resulted in significant HLA-A expression while IFN-alpha treatment did not. Expression of HLA-B alleles was low or even negative. Variable effects were observed after IFN treatment. In 3 cell lines, expression of some HLA-B alleles could not be induced by IFN-alpha or -gamma: HLA B44 in cell line 92-1, HLA-B15 in cell line OCM-1 and HLA-B5 in cell line MEL 202. The other B alleles of these cell lines showed enhanced expression levels upon IFN stimulation. In OMM-1 cells, IFN-alpha and -gamma increased the expression of HLA-A but did not induce expression of the 2 B alleles, indicating an HLA-B locus-specific loss. We thus found a high frequency of allele-specific and locus-specific down-regulation of HLA expression in uveal melanoma cell lines. Some of these defects were not restored by IFN-alpha or -gamma treatment. The lack of HLA expression may explain why uveal melanoma cells escape immune surveillance by cytotoxic T cells and complicate the development of immunotherapy in uveal melanoma. PMID- 10699952 TI - UCN-01 selectively enhances mitomycin C cytotoxicity in p53 defective cells which is mediated through S and/or G(2) checkpoint abrogation. AB - We have previously reported that UCN-01 (7-hydroxystaurosporine), a protein kinase inhibitor that is under clinical trials as an anti-cancer agent in the USA and Japan, enhanced the anti-tumor activity of mitomycin C (MMC) in vitro and in vivo. Subsequent studies from other laboratories revealed that UCN-01 could selectively enhance cytotoxicity of DNA damaging agents in p53 defective cells and that this was mediated by abrogation of S and /or G(2) arrest by UCN-01. In this study, we report that UCN-01 selectively enhances the cytotoxicity of MMC in human p53 mutant cell lines. In contrast, UCN-01 showed little, if any, effect on MMC cytotoxicity in human p53 wild-type cell lines. Terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-nick end-labeling (TUNEL) assay revealed that the combination of MMC with UCN-01 increased DNA breaks consistent with apoptosis in p53 defective A431 epidermoid carcinoma cells. In p53 wild-type MCF-7 breast carcinoma cells, the cyclin-dependent kinase inhibitor protein p21/WAF1 was markedly induced after the treatment with MMC alone, although this response was significantly delayed from the time of MMC treatment. Detailed cell-cycle studies revealed that UCN-01 abrogated S and G(2) phase accumulation induced by MMC in p53 defective cells and to a lesser extent in p53 wild-type cell lines. The abrogation of arrest in p53 wild-type cells was observed prior to significant induction of the p53 response. Since MMC was less effective against p53 defective cell lines than against p53 wild-type cell lines and UCN-01 selectively enhanced MMC cytotoxicity in p53 defective cell lines, UCN-01 may provide a new modality of MMC-based cancer chemotherapy, particularly in p53 defective cancer patients. PMID- 10699953 TI - Resistance to taxanes is induced by c-erbB-2 overexpression in human MCF-10A mammary epithelial cells and is blocked by combined treatment with an antisense oligonucleotide targeting type I protein kinase A. AB - We have tested the sensitivity of human MCF-10A mammary epithelial cells and of their transformed derivatives overexpressing an activated c-Ha-ras gene (MCF-10A Ha-ras cells), the c-erbB-2 gene (MCF-10A c-erbB-2 cells) or both genes (MCF-10A HE cells) to different cytotoxic drugs. As compared with parental MCF-10A cells, the transformed cells exhibited an increased sensitivity to topoisomerase I- and topoisomerase II-inhibitors, and to platinum-derivatives with a 2- to 10-fold reduction in IC(50) values. A remarkable difference in sensitivity was observed following treatment with taxanes. While MCF-10A Ha-ras cells showed an increased sensitivity, MCF-10A c-erbB-2 and MCF-10A HE cells exhibited a relative resistance to taxol and taxotere, with an approximately 3.5- to 6.5-fold higher IC(50) as compared with MCF-10A cells suggesting that c-erbB-2 overexpression has a dominant effect compared with an activated c-Ha-ras gene. The type I cAMP dependent protein kinase (PKAI) is overexpressed in cancer cells. Inhibition of PKAI by antisense oligonucleotides targeting its RIalpha regulatory subunit results in cancer cell growth inhibition. To evaluate the effect of blocking PKAI on MCF-10A cell sensitivity to taxanes, we treated these cells with taxol or taxotere in combination with a PKAI antisense oligonucleotide. Treatment with this agent, but not with a control scramble sequence, was able to overcome the effect of c-erbB-2 overexpression on MCF-10A cell sensitivity to taxol and taxotere, with a 20- to 40-fold shift in the IC(50) values for the 2 drugs. PMID- 10699954 TI - Use of gastrin-releasing peptide promoter for specific expression of thymidine kinase gene in small-cell lung carcinoma cells. AB - For specific transduction of herpes simplex virus thymidine kinase (HSV-tk) into human small-cell lung carcinoma (SCLC) cells, we explored the 5'-flanking region (-1.1 kb) of the gastrin-releasing peptide (GRP) gene as a lung cancer-specific promoter. RT-PCR analysis demonstrated expression of GRP mRNA in the SBC5 human SCLC cell line but not in the RERF human SCLC cell line, the A549 human lung adenocarcinoma cell line or the HeLa human uterine cervix epithelioid carcinoma cell line. A reporting vector containing the GRP promoter (pGL2-GRP) exhibited higher luciferase activity in SBC5 than in the other 3 cell lines. After transfecting an expression vector containing the GRP promoter-bound HSV-tk gene (pGRP-TK) into the cells, we measured their sensitivity to ganciclovir (GCV). In SBC5, pGRP-tk-transfected cells became about 100 times more sensitive to GCV than parental cells in vitro. In nude mice, tumors of pGRP-tk-transfected SBC5 regressed completely after i.p. administration of GCV. GRP promoter might be a good tool for tumor-specific transduction of suicide genes in GRP-expressing SCLC cells. PMID- 10699955 TI - Defective Jak-STAT signal transduction pathway in melanoma cells resistant to growth inhibition by interferon-alpha. AB - Advanced malignant melanoma is an aggressive malignancy with poor prognosis. Current therapeutic strategies have a modest success rate. The most promising treatment consists of a combination of chemotherapy with interferon-alpha, but complete response rates remain less than 15%. Interferon-alpha is also effective in adjuvant therapy for non-advanced melanoma treated surgically. The molecular mechanisms leading to loss of growth restraints and gain of growth-promoting functions during carcinogenesis of malignant melanoma are not understood in detail. Here, we studied 9 human melanoma cell lines with regard to growth inhibition by interferon-alpha and defects in intracellular signal transduction through the Jak-STAT pathway. In 3 cell lines, we found a complete loss of growth restraint by interferon-alpha. In all of them, different components of the Jak STAT pathway were defective. Since signal transduction through the Jak-STAT pathway is necessary for antiviral and antiproliferative effects of interferons, we conclude that defects in this pathway may be one of the mechanisms that lead to cancer progression through loss of growth-restraining functions. Moreover, our results indicate that a subgroup of melanomas could be completely resistant to interferon-alpha and should therefore not be treated with this cytokine. PMID- 10699956 TI - CT10: a new cancer-testis (CT) antigen homologous to CT7 and the MAGE family, identified by representational-difference analysis. AB - Assays relying on humoral or T-cell-based recognition of tumor antigens to identify potential targets for immunotherapy have led to the discovery of a significant number of immunogenic gene products, including cancer-testis (CT) antigens predominantly expressed in cancer cells and male germ cells. The search for cancer-specific antigens has been extended via the technique of representational-difference analysis and SK-MEL-37, a melanoma cell line expressing a broad range of CT antigens. Using this approach, we have isolated CT antigen genes, genes over-expressed in cancer, e. g., PRAME and KOC, and genes encoding neuro-ectodermal markers. The identified CT antigen genes include the previously defined MAGE-A6, MAGE-A4a, MAGE-A10, CT7/MAGE-C1, as well as a novel gene designated CT10, which shows strong homology to CT7/MAGE-C1 both at cDNA and at genomic levels. Chromosome mapping localized CT10 to Xq27, in close proximity to CT7/MAGE-C1 and MAGE-A genes. CT10 mRNA is expressed in testis and in 20 to 30% of various human cancers. A serological survey identified 2 melanoma patients with anti-CT10 antibody, demonstrating the immunogenicity of CT10 in humans. PMID- 10699957 TI - Metabolic effects of photodynamically induced apoptosis in an erythroleukemic cell line. A (31)P NMR spectroscopic study of Victoria-Blue-BO-sensitized TF-1 cells. AB - Victoria Blue BO (VB BO) is a new and promising photosensitizer currently being evaluated for photodynamic therapy (PDT). Its photochemical processes are mediated by oxygen radicals, but do not involve singlet oxygen. We used (31)P NMR spectroscopy of VB-BO sensitized TF-1 leukemic cells to gain further insight into the biochemical mechanisms underlying PDT-induced cell death. Sham-treatment experiments were performed to evaluate the effects of this photosensitizer in the absence of light irradiation. Significant metabolic differences were detected for TF-1 cells incubated with VB BO but not exposed to light, as compared with native cells (controls). These changes include reductions in phosphocreatine, UDP-hexose and phosphodiester levels (as percentage of total phosphate) and slightly reduced intracellular pH. Complete phosphocreatine depletion, significant acidification and concomitant inorganic-phosphate accumulation were observed for TF-1 cells irradiated after incubation with VB BO. Moreover, significant changes in phospholipid metabolites, i.e., accumulation of cytidine 5'-diphosphate choline and a decrease in phosphodiester levels, were observed for PDT-treated vs. sham treated cells. Perturbations of phospholipid metabolism may be involved in programmed cell death, and the detection of a characteristic DNA ladder pattern by gel electrophoresis confirmed the existence of apoptosis in PDT-treated TF-1 cells. PMID- 10699958 TI - Cylindrical cell carcinomas of the paranasal sinuses do not show p53 alterations but loss of heterozygosity at 3p and 17p. PMID- 10699959 TI - Presence of simian virus 40 sequences in malignant pleural, peritoneal and noninvasive mesotheliomas. PMID- 10699961 TI - Regulation of the cdk inhibitor p27 and its deregulation in cancer. AB - p27 is a cell cycle inhibitor whose cellular abundance increases in response to many antimitogenic stimuli. In this review, we summarize the current knowledge on p27 function and its regulation by synthesis and by ubiquitin-mediated degradation. Importantly, p27 degradation is enhanced in many aggressive human tumors. The frequency with which this is observed suggests that loss of p27 may confer a growth advantage to these cancers. From a practical point of view, immunodetection of p27 in tumors may prove to be useful in the assessment of prognosis and may ultimately influence the therapy of this disease. PMID- 10699962 TI - Multiple functions of p27(Kip1) and its alterations in tumor cells: a review. AB - Cyclin-dependent kinases (CDKs), together with cyclins, their regulatory subunits, govern cell-cycle progression in eukaryotic cells. p27(Kip1) is a member of a family of CDK inhibitors (CDIs) that bind to cyclin/CDK complexes and arrest cell division. There is considerable evidence that p27(Kip1) plays an important role in multiple fundamental cellular processes, including cell proliferation, cell differentiation, and apoptosis. Moreover, p27(Kip1) is a putative tumor-suppressor gene that appears to play a critical role in the pathogenesis of several human malignancies and its reduced expression has been shown to correlate with poor prognosis in cancer patients. This study reviews current information on the functions of p27(Kip1), its abnormalities found in human tumors, and the possible clinical implications of these findings with respect to the management of cancer patients. PMID- 10699960 TI - Role of insulin-like growth factors and their binding proteins in growth control and carcinogenesis. AB - Interest in the role of the insulin-like growth factor (IGF) axis in growth control and carcinogenesis has recently been increased by the finding of elevated serum insulin-like growth factor I (IGF-I) levels in association with three of the most prevalent cancers in the United States: prostate cancer, colorectal cancer, and lung cancer. IGFs serve as endocrine, autocrine, and paracrine stimulators of mitogenesis, survival, and cellular transformation. These actions are mediated through the type 1 IGF-receptor (IGF-1R), a tyrosine kinase that resembles the insulin receptor. The availability of free IGF for interaction with the IGF-1R is modulated by the insulin-like growth factor-binding proteins (IGFBPs). IGFBPs, especially IGFBP-3, also have IGF-independent effects on cell growth. IGF-independent growth inhibition by IGFBP-3 is believed to occur through IGFBP-3-specific cell surface association proteins or receptors and involves nuclear translocation. IGFBP-3-mediated apoptosis is controlled by numerous cell cycle regulators in both normal and disease processes. IGFBP activity is also regulated by IGFBP proteases, which affect the relative affinities of IGFBPs, IGFs and IGF-1R. Perturbations in each level of the IGF axis have been implicated in cancer formation and progression in various cell types. PMID- 10699963 TI - Homocyst(e)ine induces calcium second messenger in vascular smooth muscle cells. AB - Homocysteine found in the plasma of patients with coronary heart disease, induces vascular smooth muscle cell (VSMC) proliferation and increases deposition of extracellular matrix (ECM) components. Yet, the mechanism by which homocysteine mediates this effect and its role in vascular disease is largely unknown. We hypothesized that homocysteine induces ECM production via intracellular calcium release in VSMC. To test this hypothesis, aortic VSMC from Sprague-Dawley rats were isolated and characterized by positive labeling for vascular smooth muscle alpha-actin. Early passage cells (p2-3) were grown in monolayer on coverslips. Calcium transients were quantified with fura2/AM spectrofluorometry. Homocysteine induced intracellular calcium [Ca(2+)](i) transients with an EC(50) of 60 +/- 5 nM. The EC(50) for glutathione and cysteine were 10 and 100-fold lower, respectively. Depleting extracellular calcium did not alter the homocysteine effect on intracellular calcium; however, thapsigargin pretreatment, which depletes intracellular Ca(2+) stores, abolished the homocysteine effect, demonstrating its dependence on intracellular Ca(2+) stores. Extracellular sodium depletion significantly (P < 0.05) increased [Ca(2+)](i) also suggesting a possible role of sodium-calcium exchange in the process. To begin to elucidate the intracellular pathways by which homocysteine might act, VSMC were pretreated with specific inhibitors and stimulators prior to homocysteine stimulation. Staurosporine and phorbol myrisate acetate (PMA), potent simulators of protein kinase C, augmented the release of Ca(2+) by homocysteine. Interestingly, pretreatment with the nitric oxide synthase inhibitor N-nitro-L-arginine methyl ester (L-NAME) greatly exacerbated the sensitivity of VSMC to homocysteine. In contrast, pretreatment with either the phospholipase A(2) activator neomycin, the antioxidant and hepatic hydroxymethyl glutaryl coenzyme A (HMG CoA) reductase inhibitor, pravastatin, the tyrosine kinase inhibitor genestein, or the calcium channel blocker, felodipine completely inhibited the homocysteine-induced Ca(2+) signal in VSMC. This suggests the role of multiple signaling pathways in the homocysteine effect on VSMC Ca(2+). Effects of homocysteine on collagen production, as ascertained by immunoblot analysis, correlated with its effect in intracellular calcium. Regardless of the signaling pathways involved, homocysteine, by virtue of its role on VSMC proliferation and ECM deposition, has the potential to affect vascular reactivity. To determine the effect of homocysteine on the ability of VSMC to react to potent agonist such as angiotensin II, VSMC were pretreated with homocysteine and exposed to a range of angiotensin II concentrations which normally have no effect on intracellular Ca(2+). After homocysteine pretreatment, VSMC were extremely responsive to angiotensin II at concentrations well below the physiologic range. These data taken together suggested that an initial effect of homocysteine is to induce release of intracellular Ca(2+) in VSMC and may induce vascular reactivity. The transient in Ca(2+) correlates with the effect on ECM associated with homocysteine. PMID- 10699965 TI - Influence of the cellular redox state on NF-kappaB-regulated gene expression. AB - The redox state has been shown to regulate a variety of biochemical functions including cellular proliferation. Previous studies from our laboratory and others have shown that the binding of many transcription factors to their cognate DNA sequences is sensitive to the redox environment. Therefore, it is likely that redox status serves as an additional regulatory control for the activity of transcription factors and that this may mediate the redox regulation of proliferation. To assess this possibility, the influence of altering the redox state on NF-kappaB-regulated gene expression was studied. A more-reducing environment favored higher levels of expression of gro, an endogenous gene associated with proliferation, when the redox levels were changed either naturally by altering culture density or chemically by treatment with modulators of glutathione synthesis. Furthermore, nuclear runoff studies showed that a more reducing redox increased transcription of gro. In order to ascertain the singular effect of the redox state on the activity of NF-kappaB, expression of a secreted alkaline phosphatase (SEAP) reporter gene solely under the control of an NF kappaB response element was measured under varying redox conditions. Changes in the redox state modulated the expression of this reporter system. Taken together, these results suggest the involvement of a redox mechanism regulating signaling events operating through the control of gene expression by transcription factors. PMID- 10699964 TI - Estradiol-17beta-BSA stimulates Ca(2+) uptake through nongenomic pathways in primary rabbit kidney proximal tubule cells: involvement of cAMP and PKC. AB - The effect of estradiol-17beta-BSA (E(2)-BSA) on Ca(2+) uptake and its related signal pathways were examined in the primary cultured rabbit kidney proximal tubule cells. E(2)-BSA (10(-9) M) significantly stimulated Ca(2+) uptake from 2 h by 13% and at 8 h by 35% as compared to control, respectively. This stimulatory effect of E(2)-BSA was not inhibited by tamoxifen (10(-8) M, an intracellular estrogen receptor antagonist), actinomycin D (10(-7) M, a transcription inhibitor), and cycloheximide (4 x 10(-5) M, a protein synthesis inhibitor). However, E(2)-BSA-induced stimulation of Ca(2+) uptake was blocked by methoxyverapamil (10(-6) M, an L-type calcium channel blocker) and 5-(N-ethyl-N isopropyl)-amiloride (10(-5) M, a Na(+)/H(+) antiporter blocker). These results suggest that E(2)-BSA stimulates Ca(2+) uptake through nongenomic pathways. Thus, we investigated which signal pathways were related to E(2)-BSA-induced stimulation of Ca(2+) uptake. 8-Br-cAMP (10(-6) M) alone increased Ca(2+) uptake by 22% compared to control. When E(2)-BSA combined with 8-Br-cAMP, Ca(2+) uptake was not significantly stimulated compared to E(2)-BSA. SQ 22536 (10(-6) M, an adenylate cyclase inhibitor) and myristoylated protein kinase A inhibitor amide 14-22 (10(-6) M, a protein kinase A inhibitor) blocked E(2)-BSA-induced stimulation of Ca(2+) uptake and E(2)-BSA also increased cAMP generation by 26% of that of control. In addition, TPA (0.02 ng/ml, an artificial PKC promoter) stimulated the Ca(2+) uptake by 14%, and the cotreatment of TPA and E(2)-BSA did not significantly stimulate Ca(2+) uptake compared to E(2)-BSA. E(2)-BSA-induced stimulation of Ca(2+) uptake was blocked by U 73122 (10(-6) M, a phospholipase C inhibitor) or bisindolylmaleimide I (10(-6) M, a protein kinase C inhibitor). Indeed, E(2)-BSA stimulated PKC activity by 26%. In conclusion, E(2)-BSA (10(-9) M) stimulated Ca(2+) uptake by nongenomic action, which is mediated by cAMP and PKC pathways. PMID- 10699966 TI - CO(2) inhibits specific inward rectifier K(+) channels by decreases in intra- and extracellular pH. AB - Hypercapnia has been shown to affect cellular excitability by modulating K(+) channels. To understand the mechanisms for this modulation, four cloned K(+) channels were studied by expressing them in Xenopus oocytes. Exposures of the oocytes to CO(2) for 4-6 min produced reversible and concentration-dependent inhibitions of Kir1.1 and Kir2.3 currents, but had no effect on Kir2.1 and Kir6.1 currents. Intra- and extracellular pH (pH(i), pH(o)) dropped during CO(2) exposures. The inhibition of Kir2.3 currents was mediated by reductions in both intra- and extracellular pH, whereas the suppression of Kir1.1 resulted from intracellular acidification. In cell-free excised inside-out patches with cytosolic-soluble factors washed out, a decrease in pH(i) produced a fast and reversible inhibition of macroscopic Kir2.3 currents. The degree of this inhibition was similar to that produced by hypercapnia when compared at the same pH(i) level. Exposure of cytosolic surface of patch membranes to a perfusate bubbled with 15% CO(2) without changing pH failed to inhibit the Kir2.3 currents. These results therefore indicate that (1) hypercapnia inhibits specific K(+) channels, (2) these inhibitions are caused by intra- and extracellular protons rather than molecular CO(2), and (3) these effects are independent of cytosol soluble factors. PMID- 10699967 TI - System A neutral amino acid transporter regulation by interleukin-1beta in human osteoarthritic synovial cells: evidence for involvement of prostaglandin E(2) as a second messenger. AB - We studied the long-terms effects of interleukin-1beta (IL-1beta; 3 to 6 h) on alpha-(methylamino) isobutyric acid (MeAIB), a nonmetabolizable amino acid transported by system A. We found that IL-1beta induced a large decrease in MeAIB uptake by human osteoarthritic synovial cells and a concomitant increase in prostaglandin E(2) (PGE(2)) synthesis. Therefore, we investigated whether PGE(2) acts as a mediator for the long-term action of IL-1beta. We found that exogenous PGE(2) inhibited MeAIB uptake, and that AH6809, a PGE(2) receptor antagonist, inhibited IL-1beta-mediated MeAIB uptake. To identify the enzymes involved in the IL-1beta-mediated synthesis of PGE(2) that inhibits MeAIB uptake, we studied the expression of secreted (s) and cytosolic (c) phospholipase A(2) (PLA(2)). Because both were expressed, we selected a broad spectrum of inhibitors to determine which of the two PLA(2)s was involved. We used AACOCF3, a cPLA(2) inhibitor, and dithiothreitol (DTT) and bromophenacyl bromide (BPB), which are sPLA(2) inhibitors. Our results suggest that the PLA(2) involved in the IL-1beta-mediated synthesis of PGE(2) was sPLA(2). We also showed the expression of cyclooxygenase (COX)-2 and its partial involvement using a potent selective COX-2 inhibitor, L 745337. These findings provide insight into the mechanisms underlying the IL 1beta-mediated regulation of transport system A. The Il-1beta-induced inhibition of MeAIB uptake in human osteoarthritic synovial cells thus seems to be essentially mediated by PGE(2) production via the activation of sPLA(2) and the partial activation of COX-2. PMID- 10699968 TI - Human three-dimensional fibroblast cultures express angiogenic activity. AB - Human neonatal fibroblasts were cultured on a lactate-glycollate copolymer scaffold for 12-16 days to form a three-dimensional dermal equivalent tissue. The cellular content of vascular endothelial growth factor (VEGF) mRNA in these three dimensional cultures was 22-fold greater than that observed in the same fibroblasts grown as monolayers. No induction was shown by hepatocyte growth factor (HGF) or angiopoietin 1 indicating that the effect was specific to VEGF. The predominant VEGF splice variant, detected by RT-PCR corresponded to the 121 amino acid form, with less of the 165 amino acid form. The cell-associated forms (189 and 206 amino acids) comprised less than 1% of the total VEGF mRNA. VEGF and HGF proteins, determined by ELISA, were secreted in physiologically significant amounts, 0.5-4 ng per 24 h/10(6) cells. Conditioned medium from the three dimensional cultures stimulated proliferation of endothelial cells in a dose dependent manner and induced cellular expression of integrin alpha(v)beta(3). Conditioned medium from the same dermal fibroblasts cultured in monolayer showed little angiogenic activity in any of these assays. Using the chorioallantoic membrane (CAM) angiogenesis assay, the cultures stimulated blood vessel production 2.8-fold over scaffold alone. VEGF-neutralizing antibody reduced the vessel development in the CAM to the level in the scaffold control. Anti-HGF antibody had no significant effect. In conclusion, three-dimensional cultures of dermal equivalent tissue express angiogenic activity to a greater extent than monolayer cultures, some of which can be assigned to VEGF. PMID- 10699970 TI - Adaptive responses of human monocytes infected by bordetella pertussis: the role of adenylate cyclase hemolysin. AB - The activation/adaptive responses of human monocytes exposed to Bordetella pertussis parental or mutant strains were evaluated and correlated to the expression of two bacterial toxins: adenylate cyclase-hemolysin and pertussis toxin. The marked rise in intracellular cyclic adenosine monophosphate (cAMP) observed in monocytes infected by B. pertussis parental strain, inversely correlated with (1) the production of tumor necrosis factor alpha; (2) the release of superoxide anion; and (3) the expression of the 72-kDa heat shock/stress protein, Hsp70. Experiments performed with mutants deficient in adenylate cyclase-hemolysin or with purified bacterial toxins confirmed the key role of adenylate cyclase-hemolysin in the control of monocytes' response to infection by B. pertussis. This bacterial strategy primarily involves evasion from antimicrobial defenses and, eventually, the sacrifice of the host cell. PMID- 10699969 TI - Protein tyrosine kinase inhibitors increase cytosolic calcium and inhibit actin organization as resorbing activity in rat osteoclasts. AB - Although there is evidence that protein tyrosine kinase inhibitors (PTKIs) suppress bone resorption activity, the mechanism of action of these compounds on osteoclastic bone resorption remains obscure. In the present study, we investigated the effect of PTKIs on cytosolic Ca(2+) concentration ([Ca(2+)](i)) and on the cytoskeleton in rat osteoclasts. The PTKIs, genistein and herbimycin A, reversibly elevated [Ca(2+)](i) measured by fura-2 microfluorimetry. The PTKI induced increase was abolished by omission of extracellular Ca(2+), but was not attenuated by depletion of Ca(2+) stores. The PTKI-induced increase was inhibited by addition of La(3+) and Ni(2+), but not abolished by dihydropyridine (DHP) Ca(2+) channel blockers. Genistin, an inactive analogue of genistein, had no effect on [Ca(2+)](i). In the cytoskeleton assay, genistein rapidly disrupted the actin ring formation that serves as a marker for the resorbing state of osteoclasts. Disruption of the actin ring formation was also diminished in Ca(2+) free extracellular solution. These results suggest that PTKIs in rat osteoclasts elevate [Ca(2+)](i) via activation of a DHP-insensitive, nonspecific Ca(2+) entry pathway and disrupt the formation of actin rings, resulting in suppression of bone resorption activity. The regulation of this Ca(2+)-influx by PTKIs is likely to contribute to inhibition of bone resorption by these compounds. PMID- 10699971 TI - Role of small heat shock protein HSP25 in radioresistance and glutathione-redox cycle. AB - Expression of heat shock proteins (HSPs) has been shown to protect mammalian cells exposed to a variety of stress stimuli. Among various HSPs, small HSPs from diverse species were shown to protect cells against oxidative stress. Here, we show that the overexpression of the mouse small hsp gene, hsp25, provides protection against ionizing radiation. Our results demonstrate that the radiation survival of the L929 cells stably transfected with hsp25 was enhanced compared with that of the parental or vector transfected control, L25#1 cells. Our results also demonstrate that the radiation-induced apoptosis was reduced in HSP25 overexpressors. A detailed analysis of glutathione composition of those clones that overexpressed HSP25 revealed the increases of the glutathione pool, which primarily resulted from the increase of reduced glutathione. Our data suggest that higher content of GSH in HSP25 overexpressors was because of a faster reduction of oxidized glutathione (GSSG) to GSH rather than an increased de novo synthesis of GSH. The activities of glutathione reductase (GRd) and glutathione peroxidase (GPx) were greater in HSP25 overexpressors but the activity of gamma glutamylcysteine synthetase was similar between the transfectants and the control cells. Consistent with our view, a steady state ratio of the GSH/GSSG was greater in the transfectants in comparison with the control L25#1 cells. A difference in the relative ratio became more significant after exposure to the ionizing radiation. To our knowledge, this study provides the first experimental evidence in support of the hypothesis that small HSP plays a key role in radioresistance by modulating the metabolism of glutathione. Based on the results obtained from the current investigation, we propose that HSP25 helps facilitate the glutathione redox cycle and therefore, enhances glutathione utilization and maintains the cellular glutathione pool in favor of the reduced states. PMID- 10699972 TI - Decreased accumulation of [14C]carboplatin in human cisplatin-resistant cells results from reduced energy-dependent uptake. AB - We have isolated cisplatin-resistant human liver carcinoma (7404-CP20) cells with reduced accumulation of cisplatin and other drugs (methotrexate, arsenate, and arsenite) to which these cells are cross-resistant. To determine whether the reduction of drug accumulation in cisplatin-resistant cells results from impaired uptake or from active efflux, [(14)C]carboplatin was used for kinetic analysis of drug uptake and efflux. We demonstrate here that the uptake of [(14)C]carboplatin in 7404 parental cells is time, temperature, and energy dependent, and that the rate of uptake is reduced in 7404-CP20 cells. Efflux of [(14)C]carboplatin in cisplatin-resistant cells was comparable to efflux in the parental cisplatin sensitive cells. There was little effect of temperature (between 37 degrees C and 4 degrees C) on efflux in cisplatin-resistant cells. Immunoblotting with specific antibodies directed to MRP1 and MRP2 (cMOAT) also showed that expression of these two ABC transporter genes was considerably reduced in 7404-CP20 cells and another cisplatin-resistant cell line KB-CP20, in contradistinction to previous studies suggesting that MRP might be responsible for cisplatin efflux. To rule out a generalized defect in uptake of small molecules, fluorescence-activated cell sorter (FACS) analysis of rhodamine 123 uptake showed that there was no difference between cisplatin-sensitive and -resistant cells. The presence of a pleiotropic defect in uptake of [(14)C]carboplatin, [(3)H]methotrexate, [(73)As]arsenate, and [(73)As]arsenite in cisplatin-resistant cells, in association with reduced expression of related cell surface proteins as demonstrated in our previous work, suggests a novel mechanism for acquisition of resistance to cisplatin associated with reduced activity of many different specific uptake systems. PMID- 10699973 TI - N- and E-cadherin mediate early human calvaria osteoblast differentiation promoted by bone morphogenetic protein-2. AB - Bone morphogenetic protein-2 (BMP-2) stimulates the differentiation of osteoblastic cells. However, the mechanisms involved in this effect are not well characterized. In this study, we determined the role of the cell-cell adhesion molecules N-cadherin and E-cadherin in the promotion of osteoblast differentiation by BMP-2 in immortalized human neonatal calvaria (IHNC) cells. In cells cultured in aggregates, recombinant human BMP-2 (rhBMP-2) increased messenger RNA levels for alkaline phosphatase (ALP), the osteoblast specific transcription factor Osf2/Cbfa1 and osteocalcin, and enhanced in vitro osteogenesis in long-term culture. RT-PCR, immunocytochemical, and Western blot analyses showed that IHNC cells express E-cadherin, N-cadherin, and neural cell adhesion molecule (N-CAM) mRNA and protein. Treatment with rhBMP-2 induced a rapid and transient increase in N-cadherin and E-cadherin but not N-CAM, mRNA, and protein levels. Incubation with the RNA polymerase II inhibitor 5, 6-dichloro 1-beta-D-ribofuranosyl benzimidazole prevented the upregulation of N- and E cadherins induced by rhBMP-2, suggesting that transcription is necessary for this effect. N- and E-cadherins were functional because rhBMP-2 increased cell-cell adhesion in a cell aggregation assay, and this effect was largely blocked by N cadherin- and E-cadherin-neutralizing antibodies. In addition, N- and E-cadherin antibodies decreased the basal ALP activity and completely suppressed the rhBMP-2 induced increase in ALP activity and mRNA levels. Furthermore, anti-N-cadherin or anti-E-cadherin antibodies markedly decreased Osf2/Cbfa1 mRNA levels and abolished the rhBMP-2-induced increased Osf2/Cbfa1 expression, and reduced the increased osteocalcin mRNA levels induced by rhBMP-2. We conclude that rhBMP-2 rapidly and transiently increases N- and E-cadherin expression, and this effect mediates the rhBMP-2-induced early promotion of cell-cell adhesion and osteoblast marker gene expression in human calvaria cells. PMID- 10699974 TI - Intestinal cell kinase (ICK) localizes to the crypt region and requires a dual phosphorylation site found in map kinases. AB - Identification of key regulatory kinases in the intestinal epithelium are useful to understand the molecular mechanisms that underlie proliferation and differentiation in cells found in this compartment. We used the polymerase chain reaction (PCR) to amplify the catalytic kinase domain of serine-threonine kinases by employing degenerate primers and then screened an intestinal crypt cDNA library to clone and sequence the open reading frame of a novel serine-threonine kinase. This was then further characterized by Northern blot analysis and RNA in situ hybridization. This kinase, designated intestinal cell kinase, harbors a dual phosphorylation site found in mitogen-activating protein (MAP) kinases that is important for kinase activity. PMID- 10699975 TI - In memoriam: Takis S. Papas, Ph.D. (1935-1999). PMID- 10699976 TI - Erratum: nairn LM, lindsay GS, woster PM, and wallace HM (2000): cytotoxicity of novel unsymmetrically substituted inhibitors of polyamine biosynthesis in human cancer cells. J. Cell physiol 182:209-213 AB - In Figure 3 on page 212 of the above cited article, the four separate parts of the figure legend should have been referred to individually. The figure and legend appear corrected below. PMID- 10699978 TI - Pathfinding, target recognition, and synapse formation of single regenerating fibers in the adult grasshopper Schistocerca gregaria. AB - After lesion of the peripheral tympanal nerve of the adult locust (Schistocerca gregaria), sensory axons regenerate into their original target areas. We examined the individual behavior of single regenerating auditory afferents during pathway and target selection by intracellularly recording and labeling them at different times postlesion. During axotomy, spontaneous activity is not increased in either the distal or proximal part of the cells. Stimulus response properties of lesioned cells with or without regenerating axons are not influenced. Surprisingly, only 55% of sensory neurons regenerate through the lesion site and often give rise to more than one axonal fiber. Within the central nervous system, 70% of regenerated axons consistently follow an incorrect pathway to reach the correct target region. Often, one of two processes formed by a cell chooses the correct pathway, and the other the incorrect one. In the target region, regenerated axons reconstitute somatotopically ordered projections and form synapses that resemble those of intact fibers in number and structure. The regeneration process does not induce a detectable expression of antigens that are known to be expressed during neural development in these neurons. Our study clearly demonstrates that precise synaptic regeneration is possible in adult animals within a completely differentiated central nervous system, although pathfinding and formation of arborizations are disturbed in a particular and probably system-related manner. The results strongly suggest that accurate pathfinding is unlikely to be a decisive factor in target area recognition and synaptogenesis. PMID- 10699977 TI - Retinoic acid regulates the morphological development of sympathetic neurons. AB - Interactions between all-trans-retinoic acid (RA) and bone morphogenetic proteins (BMPs) affect the expression of neurotrophin receptors in sympathetic neurons (Kobayashi et al., 1998). In this study, we examined the possibility that similar interactions might regulate the morphological development of these neurons. Under control conditions, embryonic rat sympathetic neurons formed axons but not dendrites; cells exposed to RA had a similar appearance. Profuse dendritic growth was observed upon exposure to BMP-7, and this was reduced by approximately 70% by RA. This inhibitory effect of RA was mediated primarily by retinoic acid receptors (RARs) and it exhibited substantial specificity because it was not associated with changes in either axonal elongation or cell survival. Moreover, mRNAs for enzymes required for synthesis of RA were expressed in the sympathetic neurons and retinoid activity was released from superior cervical ganglia. These observations suggest that retinoids may function as endogenous morphogens and regulate neural cell shape and polarity in developing sympathetic ganglia. PMID- 10699979 TI - Stability and variability of synapses in the adult molluskan CNS. AB - Synaptic transmission was examined between identified neurons in the central nervous system (CNS) of the freshwater mollusk, Lymnaea stagnalis. Four identified neurons were used: Right Pedal Dorsal one (RPeD1; a dopaminergic respiratory interneuron), Visceral Dorsal two and three (VD2/3), and Visceral Dorsal four (VD4; a cardiorespiratory interneuron). Neuron RPeD1 synapses onto both VD2/3 and VD4, while VD4 makes a reciprocal synapse onto RPeD1. When compared from animal to animal, the connections were variable in sign. Previously, we demonstrated that, in a given animal, the RPeD1 --> VD4 synapse could be either inhibitory, biphasic, or undetectable. The present study now expands this concept of variability by showing that the RPeD1 --> VD2/3 synapse was either excitatory or undetectable from animal to animal, while the synapse from VD4 to RPeD1 was observed as inhibitory, biphasic, depolarizing, excitatory, or undetectable. Next, we used 1-day organ culture to determine if the variability observed between animals is a product of ongoing change to the sign of these identified synapses and whether or not the extent of change could be influenced by the culture conditions. Changes to the sign of transmission occurred within minutes and, more commonly, after 24-h organ culture. All three synapses were investigated before and after 1-day organ culture, in either defined medium (DM) or brain-conditioned medium (CM). Regardless of culture conditions, the RPeD1 --> VD2/3 synapse showed no change of sign, i.e., it was relatively stable. However, the synapses between RPeD1 and VD4 did change sign, and when cultured in CM, the VD4 --> RPeD1 synapse changed significantly more than in DM. These data indicate that variability of some synapses reflects changes at these synapses. This is the first report that specific synapses in an adult CNS can change sign, and that the sign of transmission can be modulated by environmental conditions. PMID- 10699980 TI - Convergence of load and movement information onto leg motoneurons in insects. AB - The interaction of two feedback loops was investigated: one regulating cuticular stress in the stick insect's leg and the other controlling leg posture. Exclusive stimulation of either of the two relevant sense organs, the load-sensitive trochantero-femoral campaniform sensilla (CS) or the position-/movement-sensitive ventral coxal hairplate (cxHPv), elicits resistance reflex responses in the retractor and the protractor coxae motoneuron pools. Concurrent application of both stimulus modalities reveals that the strength of the postural feedback response is dependent on sign and amplitude of the load feedback response and vice versa. This superposition of the two reflex responses appears to be non linear. The results indicate that the CS information is underlying a force control function in this six-legged animal. It is hypothesized that the force control of each single leg could help to optimize the force distribution of the six-legged system, even - due to the mechanical coupling - without explicit neuronal pathways. On the level of the single leg control it was studied whether the different information provided by the two feedback transducers converge on the level of retractor coxae motoneurons or whether this information is fully preprocessed at the level of premotor interneurons. It is shown here that the hairplate afferents make direct, excitatory connections with the retractor motoneurons. Studies of the motoneurons' membrane conductances during exclusive CS stimulation reveal that both, excitatory as well as inhibitory synaptic drive is delivered onto the retractor motoneurons. Thus, the motoneuronal membrane is shown to be an important stage for the sensor fusion of the two modalities. PMID- 10699981 TI - Spinal motor axons and neural crest cells use different molecular guides for segmental migration through the rostral half-somite. AB - The peripheral nervous system in vertebrates is composed of repeating metameric units of spinal nerves. During development, factors differentially expressed in a rostrocaudal pattern in the somites confine the movement of spinal motor axons and neural crest cells to the rostral half of the somitic sclerotome. The expression patterns of transmembrane ephrin-B ligands and interacting EphB receptors suggest that these proteins are likely candidates for coordinating the segmentation of spinal motor axons and neural crest cells. In vitro, ephrin-B1 has indeed been shown to repel axons extending from the rodent neural tube (Wang & Anderson, 1997). In avians, blocking interactions between EphB3 expressed by neural crest cells and ephrin-B1 localized to the caudal half of the somite in vivo resulted in loss of the rostrocaudal patterning of trunk neural crest migration (Krull et al., 1997). The role of ephrin-B1 in patterning spinal motor axon outgrowth in avian embryos was investigated. Ephrin-B1 protein was found to be expressed in the caudal half-sclerotome and in the dermomyotome at the appropriate time to interact with the EphB2 receptor expressed on spinal motor axons. Treatment of avian embryo explants with soluble ephrin-B1, however, did not perturb the segmental outgrowth of spinal motor axons through the rostral half-somite. In contrast, under the same treatment conditions with soluble ephrin B1, neural crest cells migrated aberrantly through both rostral and caudal somite halves. These results indicate that the interaction between ephrin-B1 and EphB2 is not required for patterning spinal motor axon segmentation. Even though spinal motor axons traverse the same somitic pathway as neural crest cells, different molecular guidance mechanisms appear to influence their movement. PMID- 10699982 TI - The ultrastructural interactions of identified pre- and postsynaptic cells during synaptic target recognition in Drosophila embryos. AB - During the development of neural networks, what sets synaptogenic interactions apart from nonsynaptogenic interactions is not well understood at the subcellular level. Using a combination of intracellular dye injection and electron microscopy, we show that a specific motoneuron (RP3) and its synaptic partners (muscles 6 and 7), both often bearing microprocesses, develop intimate membrane contact sites characterized by junctional structures, prior to their initiating synaptogenesis in Drosophila embryos. Other motoneuron growth cones that extend alongside the RP3 growth cone to innervate surrounding muscles do not form such contacts with muscles 6 and 7. We also examined how specific target recognition molecules affect the development of these ultrastructural associations between synaptic partner cells. When Fasciclin III (Fas3), a "positive" target recognition molecule for RP3, is ectopically expressed in neighboring muscles, the RP3 growth cone ectopically develops membrane contact sites with Fas3 misexpressing muscles with which it would not normally associate. In contrast, when Toll, a "negative" target recognition molecule normally expressed by a subset of muscles that surrounds muscles 6 and 7, is misexpressed on muscles 6 and 7, the RP3 growth cone fails to exhibit its normal close contact with these muscles. We propose that the formation of close membrane associations and junctional structures can be regulated under the influence of synaptic target recognition molecules and signifies the beginning of subcellular events during synaptic target recognition. PMID- 10699983 TI - Nerve growth factor collaborates with myocyte-derived factors to promote development of presynaptic sites in cultured sympathetic neurons. AB - Nerve growth factor (NGF) acutely modulates synaptic transmission between sympathetic neurons and their cardiac myocyte targets. NGF also has developmental effects in establishing the level of synaptic transmission between sympathetic neurons and myocytes in culture, although little is known about the mechanisms by which NGF influences this synaptic connectivity. Here we report that NGF acts in conjunction with factors produced by cardiac myocytes to promote neuronal contact with the target and the extension of synaptic vesicle-containing growth cones. In conjunction with previously published results showing that NGF has long-term effects on synaptic transmission between sympathetic neurons and myocytes, this work suggests that NGF acts to promote sympathetic neurotransmission by increasing the number of sympathetic fibers establishing target contact. Further, we found that developmental changes in cardiac myocytes led to an increase in the density of synaptic vesicle-containing variocosities along sympathetic fibers, a process regulated by NGF. Thus, as myocytes mature they produce factors that promote the formation of sympathetic presynaptic structures. These results argue that multiple target interactions regulate the extent of synapse formation between sympathetic neurons and cardiac cells and suggest that NGF promotes presynaptic development by increasing neuronal contact with myocyte-derived cell surface or matrix-associated factors. PMID- 10699984 TI - Downregulation of LAR tyrosine phosphatase prevents apoptosis and augments NGF induced neurite outgrowth. AB - The identity of the protein tyrosine phosphatases (PTPs) regulating cell death and responses to neurotrophins during neural development remain unknown. To determine if the leukocyte common antigen-related (LAR) PTP regulates these processes, PC12 cells were made LAR-deficient via stable transfection with an LAR antisense transgene. LAR-deficient cells demonstrated a stable novel phenotype, including a two-fold increase in nerve growth factor- but not fibroblast growth factor-induced neurite outgrowth. Upon serum-deprivation, LAR-deficient cells exhibited a two- to three-fold decrease in cell death. The findings that an endogenous PTP promotes cell death and counter-regulates neurotrophin actions introduce a major new receptor gene family to neurotrophic processes and suggest novel strategies for preventing cell death and augmenting neurotrophin function. PMID- 10699985 TI - Lesions of the anterior forebrain song control pathway in female canaries affect song perception in an operant task PMID- 10699986 TI - Lack of molecular markers to predict malignant potential of oral precancer. AB - Studies on the natural history of oral squamous cell carcinoma suggest a clinically detectable phase which precedes the development of cancer. Among the precancerous states, leukoplakia is the most commonly encountered clinical lesion. Although epithelial dysplasia is the most important predictive factor, non-dysplastic lesions in the oral cavity may transform with time. Early reports describing p53 overexpression in oral leukoplakia has resulted in many authors misinterpreting this result by concluding that p53 is an early marker of oncogenesis. Using sequential biopsies, few reports have critically examined the clinical significance of p53 as a risk marker. These studies suggest that p53 protein stabilization, or indeed the spectrum of mutations of p53, has no major relationship to the biological characteristics and outcome of oral leukoplakia. When used as a single marker, p53 is unsuitable for the prediction of tumour development in high-risk subjects and no other maker has yet emerged as useful. Despite the impact of molecular diagnostics on the diagnosis of tumours, assessing a patient's risk for development of cancer of the oral cavity remains limited to 'H&E' pathology. PMID- 10699987 TI - Human DNA contamination of mortuaries: does it matter? AB - With the continuing development of extremely sensitive, automated systems for the detection of human DNA from a number of cellular sources, the problem of sample contamination from scenes of crime, cadavers, and the mortuary environment has become a potentially serious issue, with implications for all involved in forensic investigations. A recent survey of 20 mortuaries identified quantifiable human DNA on mortuary work surfaces and instruments which, when amplified, produced in some cases three or more profiles from single site samples. Possible sources of DNA contamination in the mortuary are discussed, along with implications related to its presence and its avoidance during the sampling process. These observations may not be confined to forensic practice. PMID- 10699988 TI - Molecular interactions in the Vogelstein model of colorectal carcinoma. AB - For about a decade, the model proposed by Fearon and Vogelstein has been the paradigm of the genetic alterations involved in the development of colorectal carcinoma. During this time, much information has become available on the function of the key genes in this model, as well as on their interactions. This review examines the impact of this new knowledge on the Vogelstein model. It is concluded that the model as such still stands and with a few modifications could even be strengthened in that, contrary to the original proposal, the order of genetic events seems to be essential. Crucial molecular events include derangement of the Wnt- and defects in the transforming growth factor beta (TGFbeta)-signalling pathways, which exert a synergistic effect on the cell cycle. Finally, with loss of p53 function, several checks and balances are disrupted, which paves the way to gross chromosomal aberrations and aneuploidy. PMID- 10699989 TI - p53 gene mutations in sequential oral epithelial dysplasias and squamous cell carcinomas. AB - Previous studies of oral cancer have suggested that alterations of the p53 tumour suppressor gene occur early in the precancerous stage of development. However, these observations have been based on cross-sectional assessment of abnormal p53 protein staining by immunohistochemistry and may not necessarily reflect gene changes. The purpose of this longitudinal study was to examine the changes in the p53 gene in progressive, sequential epithelial dysplasias and carcinomas from the oral cavity. The study analysed 24 formalin-fixed, paraffin-embedded tissue biopsies from ten patients with two or more temporally distinct lesions from the same site in the oral cavity with the diagnosis of hyperkeratosis, epithelial dysplasia, carcinoma in situ or squamous cell carcinoma. Exons 5-8 of the p53 gene were amplified from genomic DNA using intronic primers and directly sequenced using fluorescent-labelled primers. Standard immunohistochemistry with the DO7 monoclonal antibody was used to detect mutant and wild-type p53 protein. Mutations of the p53 gene were identified in 9 of 24 samples. Eight were missense mutations and one occurred at a splice site. In six patients, mutations of the p53 gene occurred late after the transformation of epithelial dysplasia to carcinoma. In two patients with progressive dysplasia, but who had yet to develop invasive carcinoma, p53 missense mutations occurred at the carcinoma in situ stage in one case and in a moderate dysplasia in the other. There was an inconsistent relationship between gene mutations and the level of p53 protein staining by immunohistochemistry. It is concluded that during oral carcinogenesis, p53 gene mutations seem to occur relatively late and are associated with transformation to the invasive phenotype. PMID- 10699990 TI - Ki-ras activation in vitro affects G1 and G2M cell-cycle transit times and apoptosis. AB - Mutant ras genes occur frequently in human neoplasia and, in particular, in pancreatic, colorectal, and lung adenocarcinomas. Recent evidence suggests that G ->T and G-->C transversions of the Ki-ras gene in codon 12 may lead to biological effects in vitro and in vivo that may be associated with an abnormal cell cycle and increased tumour aggressiveness. The role of Ki-ras activation (a G-->C transversion in codon 12, arginine for glycine) in the cell cycle and apoptosis was investigated using control and permanently transfected NIH3T3 mouse fibroblasts. Flow cytometry was used to evaluate the G1-, S- and G2M-phase transit times, the potential doubling time, the growth fraction, and the cell loss factor during asynchronous exponential growth. Apoptosis was induced in both cell lines by absence of growth factors for an extended period of time (72 h) and quantitatively evaluated using the TUNEL method coupled with flow cytometry. It was found that codon 12 G-->C Ki-ras transfected cells compared with controls, had a significant prolongation of G1 by about 50%, a reduction of the G2M transit time by 30%, and a decrease of the cell loss factor by about 90%. Apoptotic cells were about 10% in control and less than 0.5% in Ki-ras transfected cells after 72 h starvation-confluency. These data suggest that codon 12 G-->C Ki-ras activation in mouse NIH3T3 fibroblasts is associated with deregulation of checkpoint controls in the G1 and G2M phases of the cell cycle and inhibition of apoptosis. It appears plausible that these cell mechanisms are related to a proliferative advantage and that they may also be important in the progression of human tumours characterized by specific Ki-ras mutations. PMID- 10699991 TI - Macrophage infiltration is associated with VEGF and EGFR expression in breast cancer. AB - Angiogenesis is esential for tumour growth and metastasis. Vascular endothelial growth factor (VEGF) is a potent endothelial cell mitogen and is an important component of the angiogenic stimulus in a range of human neoplasias. In addition to its mitogenic activities, VEGF has also been found to stimulate migration in macrophages via the flt-1 VEGF receptor. It has previously been shown that increased focal tumour macrophage infiltration is associated with increased angiogenesis and worsened relapse-free and overall survival in breast cancer. Macrophages are able to stimulate angiogenesis by their production of a range of factors including VEGF, tumour necrosis factor-alpha (TNF-alpha), and thymidine phosphorylase (TP). Thus, in breast cancer, VEGF could have a dual role in the regulation of angiogenesis, by direct mitogenic stimulation of endothelial cells, and also indirectly by attracting macrophages into avascular tumours. The purpose of this study was to localize VEGF protein in a series of 96 consecutive primary breast carcinomas and to determine its relationship to focal macrophage infiltration (macrophage index). These two variables were also compared with the pathological features of the tumours, as well as oestrogen receptor (ER), epidermal growth factor receptor (EGFR), microvessel density, macrophage index, and survival. An inverse relationship (p=0.0006) was noted between VEGF and EGFR, with high VEGF expression correlating with low EGFR levels. In the EGFR-negative group of cases (n=56), positive associations were observed between VEGF expression and macrophage index (p=0.005), ER (p=0.05), p53 (p=0. 006), tumour grade (p=0.02), and tumour necrosis (p=0.03). Macrophage counts were higher in EGFR-positive tumours (p=0.0006) and no associations were found between VEGF expression and increased microvessel density. These results show that in breast cancers there are two types of macrophage infiltrates, one associated with the presence of EGFR and low VEGF expression in tumours and the other with high VEGF expression in EGFR-negative tumours. VEGF expression may be an important factor in the recruitment of tumour-associated macrophages into breast carcinomas and may thus have an additional, indirect, pathway of angiogenic stimulation in this type of tumour. PMID- 10699992 TI - Gastric carcinoma exhibits distinct types of cell differentiation: an immunohistochemical study of trefoil peptides (TFF1 and TFF2) and mucins (MUC1, MUC2, MUC5AC, and MUC6). AB - The expression of trefoil peptides (TFF1 and TFF2) and mucins (MUC1, MUC2, MUC5AC, and MUC6) has previously been described in gastric polyps. In the present study, the expression profile of these trefoil peptides and mucins was characterized in 96 gastric carcinomas, in an attempt to further the understanding of the histogenesis and cell differentiation of gastric carcinoma. Taking together the co-expression of trefoil peptides and mucins, three phenotypes were defined: complete gastric, incomplete gastric, and non-gastric phenotype. Gastric differentiation (complete and incomplete) was observed in 30 out of 33 (90.9%) diffuse carcinomas and in 38 out of 53 (71.7%) intestinal carcinomas. Non-gastric differentiation was observed in only three (9.1%) diffuse carcinomas and in 15 (28.3%) intestinal carcinomas. The phenotypes observed in intestinal carcinomas were similar to those previously observed in adenomatous polyps, whereas most diffuse carcinomas mimicked the phenotype of hyperplastic polyps. The percentage of cases displaying a non-gastric phenotype was higher, though not significantly, in tumours that had invaded the gastric wall than in T1 tumours, regardless of histotype. It is concluded that gastric-type differentiation is retained in the majority of gastric carcinomas, being more prominent in diffuse than in intestinal carcinomas, and in early than in advanced carcinomas. PMID- 10699993 TI - Multiple 'serrated adenocarcinomas' of the colon with a cell lineage common to metaplastic polyp and serrated adenoma. Case report of a new subtype of colonic adenocarcinoma with gastric differentiation. AB - A 70-year-old woman underwent right hemicolectomy and six carcinomas were recognized in the resected colon. These carcinomas were considered to be of a cell lineage common to serrated adenoma (SA) and hyperplastic (metaplastic) polyp (H/MP), because of the occurrence of multiple SAs and H/MPs around the carcinomas, as well as the co-existence of SA and H/MP areas within the carcinomas. These carcinomas had the following common histological and immunohistochemical features: a serrated structure resembling SA; a lace-like structure; infiltrative growth within the muscularis propria, with dedifferentiation at the invasive front; and immunohistochemical expression of pS2 and human gastric mucin. Based on these features, a new subtype of carcinoma is proposed, with a cell lineage common to SA and H/MP. It would also seem that p53 is involved in the serrated adenoma-carcinoma sequence. PMID- 10699994 TI - Staining patterns of p53 immunohistochemistry and their biological significance in colorectal cancer. AB - Immunohistochemistry (IHC) is a cheap and rapid method to detect p53 inactivation but the results are often discordant with gene mutation analysis. This study aimed to investigate whether there is a difference in the immunohistochemical staining patterns of p53-positive cells on comparing tumours with inactivating gene mutations with those without. Tissues of 142 colorectal cancers were investigated for p53 inactivation simultaneously by IHC and gene analysis using SSCP of exons 4-9 and sequencing. In addition, tumours were investigated immunohistochemically for the expression of mdm-2 protein, known to be transcriptionally transactivated by the wild-type (wt) p53 gene. p53-positive cells of tumours without detectable p53 gene mutations were microdissected using a PALM laser microscope system and subjected to p53 sequence analysis. Among the 142 cases of colorectal cancer (male/female=88/54; mean age 66a+/-11 years, range 24-90 years), 74% (n=105) of tumours were positive by p53 IHC and mutations in the p53 gene were found in 51% (73 patients). In 16% (12 patients) with mutations in the p53 gene, IHC for p53 was negative. In tumours with mutations in the p53 gene and positive p53 IHC, staining of all nuclei of the tumour was more frequently (57/61, 93%) found than in tumours without p53 gene mutations, where staining of scattered single cells was predominantly seen (29/44, 66%; p<0.0001). mdm-2 positivity (n=33) showed only staining of scattered single cells, predominantly (24/33, 82%; p<0.0001) in tumours without gene mutations. Single cell microdissection followed by mutation analysis of scattered p53-positive cells revealed no gene mutations. A scattered positive immunohistochemical reactivity of p53 in colorectal cancer cells might therefore represent a functionally active non-mutated p53 gene and should not be considered as a marker of gene mutation and inactivation. PMID- 10699995 TI - Allelic loss at the D9S171 locus on chromosome 9p13 is associated with progression of papillary renal cell carcinoma. AB - Papillary renal cell carcinomas (RCCs) have characteristic clinical and morphological features that separate them from the more common clear cell RCCs. The details of the molecular changes in papillary RCC progression are not well understood. In this study, four highly polymorphic microsatellite markers [D9S970 (9p12-9p13), D9S171 (9p13), D9S1748 (9p21) and D9S156 (9p21)] were used to determine the frequency and prognostic significance of 9p deletions in 37 papillary RCCs. Allelic deletions were detected in eight cases (22%). The highest rate of loss of heterozygosity (LOH) was observed in 6 of 29 informative patients (21%) at the D9S171 locus on 9p13. Only two patients displayed allelic loss at D9S1748, which resides in close proximity to p16(INK4). Two of 24 informative papillary RCCs (8%) showed LOH for D9S970. LOH at D9S171 (9p13) was associated with short patient survival (p=0.008), independently of tumour grade and stage. These data suggest a tumour suppressor gene centromeric to 9p21 that may contribute to papillary RCC progression. PMID- 10699996 TI - Endometrial precancer diagnosis by histopathology, clonal analysis, and computerized morphometry. AB - Management of endometrial precancers is compromised by longstanding debate over the natural history of endometrial hyperplasias and inconsistencies in their diagnosis. The recent demonstration that some hyperplasias, like cancers, are phenotypically monoclonal is useful in recognizing biological precancers. A clonal analysis has been undertaken of a series of 93 endometrial tissues and their morphology has been evaluated by subjective diagnostic classification and computerized morphometric analysis. A pathologist's diagnosis of atypical endometrial hyperplasia was highly associated with monoclonal growth. Both microsatellite-stable and microsatellite-unstable precancers were classified as atypical hyperplasias, indicating overlapping morphologies for these two groups. Diagnosis of non-atypical endometrial hyperplasias was not reproducible and identified a group of lesions equally likely to be monoclonal as polyclonal. Computerized morphometry resolved these lesions into monoclonal and polyclonal subgroups with a high degree of accuracy and reproducibility. The predictive value of morphometry was dominated by that fraction of the sample which consisted of stroma (volume percentage stroma). This can be measured manually and used to predict monoclonality when below the threshold value of 55%. This study shows that morphometric analysis reproducibly and precisely identifies monoclonal endometrial precancers from histological sections. It may serve, furthermore, to classify accurately lesions judged by pathologists as indeterminate (non-atypical hyperplasias). The material from this study (available at www.endometrium.org from March 1, 2000) and precisely defined architectural diagnostic criteria provide new tools for diagnostic standardization of endometrial precancers. PMID- 10699998 TI - Glomerular prolapse as precursor of one type of segmental sclerosing lesions. AB - A distinctive segmental glomerular abnormality is confined to the region of the tubular opening. The hypothesis was that this followed prolapse of the tuft into the tubule. Analysis was made of 39 renal biopsy specimens with acute postinfective glomerulonephritis, later material from ten cases, four specimens from three women with pre-eclampsia, and 21 control specimens, with morphometry of glomeruli and immunohistological examination for immunoproteins and monocytes/macrophages. Prolapse was found in 14 specimens with acute postinfective glomerulonephritis, associated in eight with adhesion to Bowman's capsule and local alterations in the tuft, which together constitute early tip changes. Another three had early tip changes only and eight others had thin adhesions between the tuft and capsule next to the tubular opening. Later material confirmed this order of development and showed another late change, with sclerosed and hyaline material in the tuft and adhesion at the tubular origin. Findings in pre-eclampsia were comparable. Glomeruli were significantly larger in acute postinfective glomerulonephritis than in controls and were shown by others to be larger in pre-eclampsia than in normal pregnancy. Immunohistology showed IgM and a few foamy monocytes/macrophages in early tip changes but not in prolapsed loops. Glomerular prolapse appears to be a temporary consequence of acute enlargement of the tuft, probably causes mechanical damage to epithelial cells, and is a precursor of permanent structural changes near the tubular origin. This gives a unifying hypothesis to explain how these changes can be seen in acute postinfective glomerulonephritis, pre-eclampsia, and many other human and experimental renal disorders. PMID- 10699997 TI - Oxytocin receptors in human adenocarcinomas of the endometrium: presence and biological significance. AB - Oxytocin receptors (OTRs) are expressed in endometrial cells and oxytocin (OT) participates in endometrial functions. In cancers derived from other OT target tissues, such as breast and neural tissues, the expression of OTRs and the antiproliferative effect of OT on cancer cells has been previously observed. This study was therefore designed to search for OTR expression and the OT effect in endometrial carcinomas. To demonstrate the presence and the location of OTRs and OTR mRNA immunocytochemical, reverse transcriptase-polymerase chain reaction (RT PCR) and in situ hybridization (ISH) procedures were employed in a series of human adenocarcinomas of the endometrium. Using an anti-OTR monoclonal antibody (IF3), OTRs were demonstrated in the large majority of endometrial carcinomas (82%), with a pattern of positivity varying from diffuse to focal, according to tumour differentiation. The OTR gene was demonstrated in 78% of the cases by RT PCR and its presence was confirmed in selected cases by ISH. Moreover, in a human endometrial carcinoma cell line (COLO 684) OTR was demonstrated by immunofluorescence and RT-PCR and it was observed that OT treatment (10(-11)-10( 7) M) significantly inhibited cell proliferation. Neither toxic effects nor apoptosis were induced by OT treatment. The addition of an inhibitor of protein kinase A (PKA) to the culture medium abolished the antiproliferative effect of OT, suggesting that cAMP via PKA could be the intracellular mediator of the OT effect, as previously observed in breast and neural tumours. In conclusion, this study presents evidence of OTR expression in human endometrial carcinomas and of an OT antiproliferative effect on human endometrial cancer cells in vitro. It is further suggested that OT and OTR may be involved in the regulation of endometrial cells, not only in physiological conditions but also in a neoplastic context. PMID- 10699999 TI - Stenotic glomerulotubular necks in radiation nephropathy. AB - Fibrosis of the renal interstitium is well correlated with kidney function and a greater extent of fibrosis predicts renal failure. Recent work has shown striking fibrotic constrictions at the glomerulotubular neck in porcine radiation nephropathy and in Wegener's granulomatosis in man. The present studies were designed to identify stenotic necks in a third species and to evaluate the effect of captopril treatment. Experimental radiation nephropathy was established with 17 Gy total body irradiation of barrier-maintained rats. Kidneys were obtained at 99 and 203 days for histology, using perfusion fixation. There was renal injury, with a rise in BUN, as expected, which was attenuated by captopril treatment. There was stenotic neck formation at 99 and 203 days. Captopril did not influence the absolute fraction of necks that were stenotic but it did prevent the evolution of glomeruli with necks to atubular glomeruli. It is concluded that stenosis of the glomerulotubular neck is a general phenomenon of any scarring kidney disease; that stenotic necks are probably an intermediate step in the evolution towards atubular glomeruli; and that timely use of captopril may prevent the progression of a stenotic neck towards an atubular glomerulus. PMID- 10700000 TI - Inhibition of tumour necrosis factor alpha does not prevent experimental paracetamol-induced hepatic necrosis. AB - Paracetamol-induced hepatic necrosis is the most common form of toxic liver injury experienced in clinical practice in the UK and USA. Recently, reports have described prevention of hepatic necrosis, induced by other hepato-toxins, by inhibiting tumour necrosis factor alpha (TNFalpha). The aim of the present study was to determine the role of TNFalpha in paracetamol-induced hepatic necrosis. Six-week-old CBA/J female mice were given 300 mg/kg paracetamol by intraperitoneal (IP) injection after an 8-h fast. Hepatic expression of TNFalpha was measured by enzyme-linked immunoassay (ELISA) and reverse transcriptase polymerase chain reaction (RT-PCR). Serum TNFalpha was measured by ELISA. One hour prior to paracetamol injection, mice were also given blocking anti-TNFalpha antibodies, soluble TNFalpha receptor, interleukin 10 (IL-10), and dexamethasone. Hepatic injury was measured by serum aspartate aminotransferase and histological assessment on haematoxylin and eosin (H&E)-stained liver sections. There was a significant increase in serum TNFalpha at 6 h (control 0.002+/-0.002 ng/ml, n=7; paracetamol-treated 0.022+/-0.007 ng/ml, n=5, p<0.05), but hepatic TNFalpha expression did not change up to 24 h following paracetamol injection. Histologically severe centrilobular hepatic necrosis was noted at 3 h and progressed for 24 h after paracetamol poisoning. Death rate, serum aspartate aminotransferase, and hepatic histology were not significantly different between the groups treated with blocking anti-TNFalpha antibodies, soluble TNFalpha receptor, IL-10, and dexamethasone, compared with controls. In conclusion, there is no evidence to suggest that modulation of TNFalpha expression affects hepatic injury following experimental paracetamol poisoning; anti-TNFalpha therapies are therefore unlikely to be effective in the corresponding clinical situation. PMID- 10700001 TI - Role of spleen macrophages in the clearance of scrapie agent early in pathogenesis. AB - The involvement of spleen macrophages in the early stages of scrapie pathogenesis was studied by applying the 'macrophage-suicide technique' to scrapie-infected mice. This method comprises critically the intravenous administration to mice of dichloromethylene disphosphonate encapsulated into liposomes. Depletion of spleen macrophages before scrapie infection induced an increased amount of scrapie inoculum in the spleen, consequently leading to accelerated scrapie agent replication in the early phase of pathogenesis, as followed by PrPres accumulation, a specific hallmark of scrapie. The same effect was observed when spleen macrophages were depleted just before the beginning of scrapie agent replication. These findings suggest that macrophages may partly control scrapie infection in peripheral tissues by sequestration of the scrapie inoculum and may thus impair early scrapie agent replication in the spleen. In addition to macrophages, most follicular dendritic cells and B lymphocytes, which are thought to support scrapie agent replication, were also transiently depleted by dichloromethylene disphosphonate administration. This suggests that a compensatory mechanism is sufficient to ensure the persistence of infection in these early stages of pathogenesis. PMID- 10700002 TI - Cultured human fibroblasts in agarose gel as a multi-functional control for immunohistochemistry. Standardization Of Ki67 (MIB1) assessment in routinely processed urinary bladder carcinoma tissue. AB - Immunohistochemistry (IHC) in clinical practice is hampered by lack of standardization and by subjectivity in interpretation and quantitation. This study aimed to develop a control system for IHC in routinely fixed and histoprocessed tissues. Such a system should be easy to handle in clinical practice and should reflect variations in fixation time, section thickness, section storage conditions, and staining protocols. In addition, in image analysis quantitation of immunostained tissues, when using classifiers computed on IHC-control images, the control system should be very stable. Cultured human fibroblasts were suspended in agarose, transferred into a length of tubing and stored at 4 degrees C. Three pieces of the cellgel control were separately fixed, histoprocessed, and paraffin-embedded as external controls. One piece was prepared together with each of 18 bladder carcinoma biopsies as internal controls. Slides with sections from the biopsy and all types of cellgel controls were stored at different temperatures and then stained using three different IHC protocols. The fibroblasts were homogeneously distributed in the agarose gel. Variation in section thickness did not influence immunostaining as evaluated by the MIB1 labelling index (MIB1 LI). The external controls decreased notably in MIB1 LI with increased fixation time. This was not seen in the 18 internal controls that were each fixed with a fresh biopsy. However, section storage and immunostaining conditions influenced the MIB1 expression equally in all control types and to a similar degree to the biopsies. Furthermore, colour-based image analysis quantitation of MIB1 LI in biopsies proved stable and independent of the control type used to compute the classifier. PMID- 10700003 TI - Analysis of the amplification refractory mutation allele-specific polymerase chain reaction system for sensitive and specific detection of p53 mutations in DNA. AB - The sensitivity of the amplification refractory mutation allele-specific polymerase chain reaction system (ARMAS-PCR) to detect known p53 mutations was determined using DNA extracted from two human tumour cell lines collected by cytobrush, as a model for its use in exfoliative cytology. Using DNA extracted from SW480 and CEM cell lines diluted with normal human fibroblasts, a nested ARMAS-PCR was more sensitive than a non-nested version and could detect one mutated cell amongst 100 000 normal cells. When compared with PCR-single stranded conformational polymorphism, nested ARMAS-PCR was 10 000 times more sensitive for detecting mutant p53 in extracted DNA. Primer design proved to be influential on the sensitivity and specificity of the assay; increased specificity was achieved by the use of deliberate mismatches upstream from the 3' end of mutation-specific primers. ARMAS-PCR was confirmed to be specific for the mutation that each primer was designed to detect. Nested ARMAS-PCR offered a rapid and sensitive method of analysis of cells with predetermined p53 mutations and has the potential to be applied to the study of the molecular progression of cancer, including diagnosis and detection of residual disease. It could also be extended to the in situ detection of aberrant cells. PMID- 10700005 TI - Authors' reply PMID- 10700004 TI - Re. Review article entitled 'The neoplastic pathogenesis of solitary and multiple osteochondromas'. PMID- 10700006 TI - Seeing is believing: non-invasive, quantitative and repetitive imaging of reporter gene expression in living animals, using positron emission tomography. AB - The ability to monitor reporter gene expression in living animals and in patients will permit longitudinal examinations both of somatically transferred DNA in experimental animals and patients and of transgenic constructs expressed in experimental animals. If investigators can non-invasively monitor the organ and tissue specificity, the magnitude and the duration of gene expression from somatically transferred DNA and from transgenes, conceptually new experimental paradigms will be possible. If clinicians can non-invasively monitor the location, extent and duration of somatically transferred genes, they will be better able to determine the correlations between expression of therapeutic genes and clinical outcomes. We have developed two reporter gene systems for in vivo reporter gene imaging in which the protein products of the reporter genes sequester positron-emitting reporter probes. The "PET reporter gene" dependent sequestration of the "PET reporter probes" is subsequently measured in living animals by Positron Emission Tomography (PET). We describe here the principles of PET reporter gene/PET reporter probe in vivo imaging, the development of two imaging systems, and the validation of their ability to non-invasively, quantitatively and repetitively image reporter gene expression in murine viral gene transfer and transgenic models. PMID- 10700007 TI - Involvement of OSP/claudin-11 in oligodendrocyte membrane interactions: role in biology and disease. AB - Oligodendrocyte-specific protein (OSP/claudin-11) is a four transmembrane protein concentrated in central nervous system myelin. Recent evidence has emerged suggesting that OSP/claudin-11 is involved in membrane interactions at tight junctions and with the extracellular matrix. OSP/claudin-11 seems to modulate proliferation and migration of oligodendrocytes presumably through these interactions. Furthermore, evidence is presented implicating OSP/claudin-11 as an autoantigen in the development of autoimmune demyelinating disease. PMID- 10700008 TI - Treatment of experimental autoimmune encephalomyelitis with antisense oligonucleotides against the low affinity neurotrophin receptor. AB - Upregulated expression of the low-affinity neurotrophin receptor (p75) in the central nervous system (CNS) during experimental autoimmune encephalomyelitis (EAE) has recently been demonstrated. To investigate whether p75 plays a role in disease pathogenesis, we adopted a gene therapy approach, utilizing antisense oligonucleotides to downregulate p75 expression during EAE. Phosphorothioate antisense oligonucleotides (AS), nonsense oligonucleotides (NS) or phosphate buffered saline (PBS) were injected daily for 18 days after immunization of SJL/J (H-2s)-mice with myelin proteolipid protein (PLP) peptide 139-151. In the AS group, there was a statistically significant reduction in both the mean maximal disease score (1.85 in the AS, 2.94 in the NS and 2.75 in the PBS-groups, respectively, P < 0.025) and in the cumulative disease incidence ( approximately 60% in the AS group and approximately 90% in the control groups). Histological and immunohistochemical analysis showed reduced inflammation and demyelination, as well as reduced p75 expression at the blood-brain barrier (BBB) in the AS treated mice in comparison with both control groups. There was no difference, however, in p75 expression on neural cells within the CNS between the three groups of mice. We conclude that p75 could play a proactive role in the pathogenesis of EAE and may exert its effect at the level of the BBB. PMID- 10700009 TI - Expression of galectin-1 mRNA correlates with the malignant potential of human gliomas and expression of antisense galectin-1 inhibits the growth of 9 glioma cells. AB - Although its precise function has not yet been established, galectin-1 seems to play a role in tumor progression. In this study, we investigated galectin-1 mRNA expression in human glioma specimens and glioma cell lines. Northern blot analysis showed higher galectin-1 mRNA levels in glioma tissues. The 0.7-kb galectin-1 mRNA transcript was detected, and the expression level correlated with the malignant state, from low-grade astrocytoma to glioblastoma. In several human glioma specimens, immunohistochemical examination with antiserum against a synthetic peptide corresponding to the predicted C-terminal sequence of the protein showed high levels of galectin-1 expression. To clarify the correlation between the expression of galectin-1 and the malignancy of gliomas, we examined whether expression of antisense galectin-1 would suppress tumor growth in rat 9L cells that express high levels of galectin-1. The cells were transfected with a plasmid DNA that produces antisense galectin-1 mRNA under the control of the metallothionein promoter, and stable clones expressing low levels of galectin-1 protein in comparison with control clones were isolated. Cells with low levels of galectin-1 displayed dramatic phenotypic changes in their morphology and growth properties compared with vector-transfected control 9L cells. Our data suggest that decreased expression of galectin-1 may arrest the growth of rat 9L cells. PMID- 10700010 TI - Intracellular pH changes during oligodendrocyte differentiation in primary culture. AB - We have studied the characteristics of pH(i) regulation at different stages of rat oligodendrocyte differentiation in primary culture. pH(i) was measured at 37 degrees C using the pH-sensitive fluorescent probe BCECF. In immature oligodendrocyte progenitor (OLP), three distinct ionic mechanisms were involved in pH(i) regulation: (i) a sodium-independent Cl(-)/HCO(-)(3) exchanger, (ii) a Na(+)/H(+) exchanger and (iii) a voltage-dependent Na(+)-HCO(-)(3) cotransporter. The two latter mechanisms were also detected in more differentiated pro oligodendrocytes and in mature oligodendrocytes whereas the Cl(-)/HCO(-)(3) exchanger was not active in these two later stages of differentiation. The presence of this Cl(-)/HCO(-)(3) exchanger (that acts as a chronic acidifying mechanism) only in immature OLP maintains in these cells a steady-state pH(i) value significantly lower than values measured in more differentiated cells. The possible involvement of this pH(i) change in triggering cell differentiation is discussed. PMID- 10700011 TI - Activation of dopamine D(3) receptors induces c-fos expression in primary cultures of rat striatal neurons. AB - A modulation of the expression of immediate-early genes (IEGs) such as c-fos is likely involved in the long-term influence of dopaminergic ligands on the activity of basal ganglia neurons. The roles of individual dopamine receptor types in this regard remain unclear, and the present study employed primary cultures of rat striatal neurons to examine the potential modulation of c-fos expression by D(3) receptors. Neurons were treated with the preferential D(3) receptor agonists, (+/-)-7-OH-DPAT and PD 128,907, which showed, respectively, 16 fold and 56-fold selectivity for recombinant rat D(3) vs. D(2) receptors (K(i) values, rD(2)/rD(3) = 59.5/3.7 nM and 251/4.5 nM, respectively). At concentrations of 3 and 30 nM, respectively, (+/-)-7-OH-DPAT and PD 128,907 significantly increased the expression of c-fos mRNA. The action of (+/-)-7-OH DPAT was expressed stereospecifically; its (+)-isomer (K(i) values, D(3)/D(2) = 1.6/56.7 elicited a 26% +/- 7.6% increase in c-fos expression whereas its (-) isomer (K(i) values, D(3)/D(2) = 59/1,060 nM) was ineffective. Further, stimulation of c-fos mRNA expression by PD 128,907 (20 nM) was markedly and significantly (P < 0.05) attenuated (-91.8% +/- 5.3%) by 30 nM of the selective D(3) receptor antagonist, (+)-S 14297 (K(i) values, D(3)/D(2) = 11/401 nM). In contrast, the action of PD 128,907 was significantly less affected (-24.5% +/- 7.8%) by 30 nM of its less active distomer, (-)-S 17777 (K(i) values, D(3)/D(2) = 294/3,191 nM). An examination of the concentration-response relationship revealed that (+/-)-7-OH-DPAT and PD 128,907 both produced bell-shaped dose-response curves for c-fos induction. The sequential activation of D(2) receptors-which inhibit striatal c-fos expression (Simpson and Morris [1995] Neuroscience 68:97 106)-by higher concentrations of (+/-)-7-OH-DPAT and PD 128,907 is presumably involved in the inflexion at higher doses. In conclusion, the present data demonstrate that activation of D(3) receptors results in a selective induction of c-fos mRNA expression in cultured striatal neurons. These data show that neuronal D(3) receptors control the expression of IEGs and suggest that D(3) receptors may mediate long-term adapative changes in the activity of neurons in the basal ganglia. PMID- 10700012 TI - Pre- and postsynaptic localization of RC3/neurogranin in the adult rat spinal cord: an immunohistochemical study. AB - RC3 (neurogranin; BICKS) is a neuron-specific calmodulin-binding protein kinase C substrate. Thus far, immunohistochemical studies on the localization of RC3 revealed its presence in all neuronal phenotypes, which were restricted to specific areas in the neostriatum, the neocortex, and the hippocampus. RC3 was mostly found in cell bodies and dendrites, with some infrequent presence in axonal profiles, i.e. in the internal capsule. Until now, RC3 expression was reported to be absent in the adult rat spinal cord. RC3 might, however, act as an intermediate of protein kinase C-mediated signaling pathways during synaptic development and plasticity. We hypothesized a role for this 78-amino-acid protein in dendritic plasticity occurring after spinal cord injury. To our surprise, an immunohistological analysis of the uninjured adult rat spinal cord revealed the presence of RC3-positive cell bodies and dendrites in specific regions in the gray matter. Interestingly, axon-containing structures, such as the dorsal and ventral corticospinal tract, were also found to be RC3-positive. This axonal labeling was confirmed by preembedding electron microscopy. In conclusion, we demonstrate here that RC3 is present in the adult rat spinal cord in pre- and postsynaptic structures. PMID- 10700013 TI - Modulation of stimulus-secretion coupling in porcine adrenal chromaffin cells by receptor-mediated increases in protein kinase C activity. AB - Catecholamine (CAT) secretion by adrenal chromaffin cells is primarily triggered by nicotinic receptor-dependent increases in cytosolic Ca(2+). The principal aim of the present study was to determine whether pituitary adenylate cyclase activating peptide (PACAP), which is coreleased with acetylcholine from the splanchnic nerve, can modulate nicotinic receptor-dependent Ca(2+) signaling and catecholamine secretion in porcine adrenal medullary chromaffin (PAMC) cells. Activation of protein kinase C (PKC) with phorbol myristate acetate (PMA) dose- and time-dependently inhibited nicotine (NIC)-induced Ca(2+) transients. At 100 nM PMA, peak Ca(2+) levels were reduced by 27% +/- 2% (P < 0.05) and 41% +/- 3% (P < 0. 05) after 10 and 20 min exposure, respectively. The inhibitory effects of PMA were significantly reduced by preincubation with the PKC inhibitor staurosporine. KCl-induced Ca(2+) transients were also reduced by 20 min PMA treatment (Delta -27% +/- 4%; P < 0.05), suggesting that PKC affects voltage gated Ca(2+) channel activity. Pretreatment with PACAP also resulted in both time and concentration-dependent suppression of Ca(2+) transients. After 20 min exposure to 1 microM PACAP, NIC- and KCl-induced transients were reduced by 36% +/- 5% (P < 0.05) and 51% +/- 6% (P < 0.05), respectively. These effects could also be prevented by staurosporine pretreatment. NIC-induced CAT secretion was significantly reduced by pretreatment with both PMA (Delta -56% +/- 2%; P < 0.05) and PACAP (Delta-53% +/- 7%; P < 0.05). This suppressive effect on secretion could be prevented by pretreatment with staurosporine. These data suggest that, in addition to having direct stimulatory effects on catecholamine synthesis and secretion, PACAP can also negatively modulate nicotinic receptor-dependent Ca(2+) signaling and secretion in PAMC cells. PMID- 10700014 TI - Expression of the RNA-binding protein TIAR is increased in neurons after ischemic cerebral injury. AB - T-cell restricted intracellular antigen-related protein (TIAR) is an RNA recognition motif-type RNA-binding protein that has been implicated in the apoptotic death of T-lymphocytes and retinal pigment epithelial cells. Western blots prepared with a monoclonal antibody against TIAR showed expression in normal rat hippocampus, and induction by 15 min of global cerebral ischemia. This increased expression was evident at 8 hr after ischemia and maximal at 24 hr, whereas expression at 72 hr was reduced below basal levels. Expression of TIAR protein was also increased in parietal cortex 6 and 24 hr after 90 min of focal cerebral ischemia induced by middle cerebral artery (MCA) occlusion, as well as in cultured cortical neurons and astroglia after exposure to hypoxia in vitro. Immunocytochemistry showed that increased expression of TIAR occurred mainly in the CA1 sector of hippocampus 24 hr after global ischemia, and in cortical and striatal neurons 24 hr after 20 or 90 min of focal ischemia. Double-labeling studies showed that TIAR protein expression was co-localized with DNA damage in neuronal cells. The findings suggest that TIAR may be involved in neuronal cell death after cerebral ischemic injury. PMID- 10700015 TI - Early acute necrosis, delayed apoptosis and cytoskeletal breakdown in cultured cerebellar granule neurons exposed to methylmercury. AB - Cerebellar granule cells (CGCs) are a sensitive target for methylmercury (MeHg) neurotoxicity. In vitro exposure of primary cultures of rat CGCs to MeHg resulted in a time- and concentration-dependent cell death. Within 1 hr exposure, MeHg at 5-10 microM caused impairment of mitochondrial activity, de-energization of mitochondria and plasma membrane lysis, resulting in necrotic cell death. Lower MeHg concentrations (0.5-1 microM) did not compromise cell viability, mitochondrial membrane potential and function at early time points. Later, however, the cells progressively underwent apoptosis and 100% cell death was reached by 18 hr treatment. Neuronal network fragmentation and microtubule depolymerization were detected as early as within 1.5 hr of MeHg (1 microM) exposure, long before the occurrence of nuclear condensation (6-9 hr). Neurite damage worsened with longer exposure time and proceeded to the complete dissolution of microtubules and neuronal processes (18 hr). Microtubule stabilization by taxol did not prevent MeHg-induced delayed apoptosis. Similarly ineffective were the caspase inhibitors z-VAD-fluoromethylketone and z-DEVD chloromethylketone, the L-type calcium channel inhibitor nifedipine, the calcium chelator EGTA and BAPTA, and the NMDA receptor antagonist MK-801. On the other hand, insulin-like growth factor-I partially rescued CGCs from MeHg-triggered apoptosis. Altogether these results provide evidence that the intensity of MeHg insult is decisive in the time of onset and the mode of neuronal death that follows, i.e., necrosis vs. apoptosis, and suggest that cytoskeletal breakdown and deprivation of neurotrophic support play a role in MeHg delayed toxicity. PMID- 10700016 TI - Low-affinity kainate receptor agonists induce insult-dependent apoptosis and necrosis in cultured murine cortical neurons. AB - Overstimulation of ionotropic glutamate receptors leads to excitotoxic neuronal death, which has been implicated in the neurodegeneration of neurological diseases. The present study examined the role of putative low-affinity kainate receptor subtype (GluR5-7) agonists in excitotoxicity in cultured murine cortical neurons. The concentration-dependent decrease in cell viability induced by the agonists kainate (1-1,000 microM) and (RS)-2-amino-3-(hydroxy-5-tert butylisoxazol-4-yl) propanoic acid (ATPA; 1-1,000 microM) was only attenuated by 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX; 10 microM) and 1-(4-aminophenyl)-4 methyl-7,8-methylenedioxy-5H-2,3-benzodiazepine (GYKI 52466; 20 microM). (S)-5 iodowillardiine (1-1,000 microM)-induced toxicity was attenuated by CNQX (20 microM), GYKI 52466 (20 microM) and MK-801 (10 microM); however, (2S, 4R)-4 methylglutamate (1-120 microM)-induced toxicity was not attenuated by the antagonists. None of the agonists possessed selective actions at GluR5-7. Morphological observations (phase-contrast and fluorescence microscopy) revealed that the agonists induced two distinct patterns of neuronal injury. After 24 hr of treatment, low concentrations of agonists (1-30 microM) produced cellular shrinkage and nuclear granulation consistent with slow, apoptotic-like neuronal death. Pyknotic labeling with the DNA binding dye Sytox green confirmed these apoptotic characteristics, which significantly decreased with increasing concentrations. After 4 hr, increasing concentrations of agonists (100-1,000 microM) induced cellular swelling, with subsequent extracellular debris; labeling with propidium iodide revealed isolated nuclei consistent with the increased involvement of rapid necrosis. Thus, all putative GluR5-7 agonists produced excitotoxicity across a necrotic-apoptotic continuum in murine cortical neuron cultures. PMID- 10700017 TI - Inhibitors of NO-synthase and donors of NO modulate kainic acid-induced damage in the rat hippocampus. AB - The effects of nitric oxide synthase (NOS) inhibitors, N(omega)-nitro-L-arginine and 7-nitroindazole, and the NOS substrate L-arginine on kainic acid (KA)-induced microglial reactivity and stress response were studied in the hippocampus 7 and 1 days after KA, respectively. Density of peripheral-type benzodiazepine receptors was measured as an index of microglial reactivity. Histological damage in hippocampus was evaluated at 7 days by neuronal counting. KA increased the maximal number of binding sites (B(max)) versus controls. Administration of either 7-nitroindazole (25 mg/kg) or N(omega)-nitro-L-arginine (20 and 50 mg/kg) 24 hr before KA, further increased B(max). This later effect was abolished by L arginine (1 g/kg), which given 24 hr before KA decreased B(max) to control values. Also, KA-induced HSP72 stress response was attenuated by pre-treatment with L-arginine. Histological evaluation showed reduced cell numbers in the pyramidal cell layer of the hippocampus in groups receiving KA, either alone or in combination with 7-nitroindazole. Administration of L-arginine before KA attenuated neuronal loss in CA3 but not CA1. A clear protective effect was observed, however, in CA1 and CA3, in rats receiving both L-arginine plus 7 nitroindazole before KA. The results show that the combination of a NO substrate with a NOS inhibitor reduces the neurotoxic effects of KA in the rat hippocampus. This study suggests that extremely fine regulation of NO levels in the different neural cell types can modulate excitotoxicity. PMID- 10700018 TI - Deficits in operant behavior and alteration of CA1, CA3 hippocampal dendritic arborization due to subicular lesions. AB - The deficits in operant behavior and the alterations in dendritic arborizations of Cornu Ammonis 1 and Cornu Ammonis 3 (CA1 and CA3) hippocampal areas were investigated in subicular lesioned rats. The subjects were female Wistar rats aged 120 days, and were divided into four groups: one serving as age-matched untrained control, a second group received training and sham lesioning, a third group were only trained, and the fourth group were first trained and then subjected to subicular lesions. The rats were food-deprived 24 hours prior to operant behavior training sessions. Two training sessions for operant behavior with continuous reinforcement of 10 minutes duration per day were done during the shaping session, following which rats were allowed 10 minutes of operant food reward for 10 days. On the eleventh day, only the operant behavior and sham operated rats were used for subicular lesion and sham surgery, respectively. After 72 hours of surgical recovery, operant behavioral testing was performed daily as before for a further period of 10 days. Later, all groups of rats were killed and the hippocampus was processed for rapid Golgi staining. Our results suggest that subicular lesions produce a significant reduction in operant learning. Further, the Golgi studies revealed a reduction in dendritic branching points and intersections of apical and basal CA1, CA3 neurons in lesioned rats. PMID- 10700019 TI - Functional properties, developmental regulation, and chromosomal localization of murine connexin36, a gap-junctional protein expressed preferentially in retina and brain. AB - Retinal neurons of virtually every type are coupled by gap-junctional channels whose pharmacological and gating properties have been studied extensively. We have begun to identify the molecular composition and functional properties of the connexins that form these 'electrical synapses,' and have cloned several that constitute a new subclass (gamma) of the connexin family expressed predominantly in retina and brain. In this paper, we present a series of experiments characterizing connexin36 (Cx36), a member of the gamma subclass that was cloned from a mouse retinal cDNA library. Cx36 has been localized to mouse chromosome 2, in a region syntenic to human chromosome 5, and immunocytochemistry showed strong labeling in the ganglion cell and inner nuclear layers of the mouse retina. Comparison of the developmental time course of Cx36 expression in mouse retina with the genesis of the various classes of retinal cells suggests that the expression of Cx36 occurs primarily after cellular differentiation is complete. Because photic stimulation can affect the gap-junctional coupling between retinal neurons, we determined whether lighting conditions might influence the steady state levels of Cx36 transcript in the mouse retina. Steady-state levels of Cx36 transcript were significantly higher in animals reared under typical cyclic-light conditions; exposure either to constant darkness or to continuous illumination reduced the steady-state level of mRNA approximately 40%. Injection of Cx36 cRNA into pairs of Xenopus oocytes induced intercellular conductances that were relatively insensitive to transjunctional voltage, a property shared with other members of the gamma subclass of connexins. Like skate Cx35, mouse Cx36 was unable to form heterotypic gap-junctional channels when paired with two other rodent connexins. In addition, mouse Cx36 failed to form voltage-activated hemichannels, whereas both skate and perch Cx35 displayed quinine-sensitive hemichannel activity. The conservation of intercellular channel gating contrasts with the failure of Cx36 to make hemichannels, suggesting that the voltage-gating mechanisms of hemichannels may be distinct from those of intact intercellular channels. PMID- 10700020 TI - Integrin-type signaling has a distinct influence on NMDA-induced cytoskeletal disassembly. AB - Adhesion responses triggered by integrin-class matrix receptors have been implicated in the synaptic reorganization events necessary for certain types of neuronal plasticity. Hippocampal slice cultures were used to test whether the related structural transformations elicited by NMDA receptor stimulation are regulated by integrin-type signals. Infusing the slices with NMDA for a short period induced the expected disassembly of the cytoskeletal network, measured with antibodies that selectively recognize spectrin cleavage sites targeted by the protease calpain. Marked levels of the 150-kDa breakdown product (BDP) were produced, whereas concentrations of the parent spectrin were not changed. Interestingly, the calpain cleavage events were attenuated by 60% when integrin type signaling was disrupted with the antagonist Gly-Arg-Gly-Asp-Ser-Pro (GRGDSP). This effect was RGDS-dependent, was largely evident in synapse-dense dendritic areas, particularly in subfield CA1, and was abolished when the NMDA exposure period was >5 min. These findings suggest that only those cytoskeletal alterations associated with brief synaptic activity are regulated by intact contact zones. AMPA-type glutamate receptors also were tested because, like spectrin, they are targets for calpain. Brief NMDA treatment caused a 15% loss of AMPA receptor GluR1 carboxytermini and this modification was augmented to 32% in the presence of GRGDSP. Thus, although blockage of matrix recognition signals decreased spectrin's susceptibility to disassembly, it increased the susceptibility of AMPA receptors to proteolysis. These data indicate that integrin-type signaling complexes are appropriately positioned to govern cytoskeletal reconfiguration while stabilizing the structural nature of AMPA receptors. PMID- 10700021 TI - Prenatal cocaine exposure increases serotonergic inhibition of electrically evoked acetylcholine release from rat striatal slices at adulthood. AB - This study tests the hypothesis that prenatal cocaine (pCOC) exposure (20 mg/kg, bidaily from embryonic days 15-21) modifies 5-HT(3) receptor regulation of electrically-evoked [(3)H]acetylcholine (ACh) overflow from adult male and female (proestrus, diestrus) rat striatal slices. Also, the influence of endogenous dopamine (DA) on serotonin (5-HT) regulation of ACh overflow was determined by assessing the effects alpha-methyl-para-tyrosine (AMPT) pretreatment or sulpiride. Phenylbiguanide (PBG, 5-HT(3) agonist) superfusion dose-dependently inhibited ACh overflow in all groups except the diestrus pCOC group in which there was an enhanced sensitivity to PBG. PBG (10, 30, and 60 microM) produced greater effects in the pCOC male than in the prenatal saline (pSAL) group. The pCOC male group also exhibited greater sensitivity to PBG (30 and 60 microM) than the pCOC proestrus group. PBG inhibition of ACh overflow was comparable in the pSAL male and female (proestrus) groups. PBG inhibition of ACh overflow was greater in the pCOC diestrus group than in the pCOC proestrus (10, 30, and 60 microM), the pSAL diestrus (10 and 30 microM), and the pCOC male (10 microM) conditions. In slices from untreated rats superfused with 30 microM PBG, AMPT pretreatment (68% DA loss) reduced inhibition of ACh overflow, and 1 microM sulpiride increased ACh overflow. ICS205-930 (5-HT(3) antagonist) reduced effectiveness of PBG indicating 5-HT(3) receptor specificity for PBG. In summary, pCOC exposure enhances modulatory effects of 5-HT (via 5-HT(3) receptors) on striatal ACh release in male and females rats and the inhibitory actions of 5 HT(3) receptors are mediated by DA. PMID- 10700022 TI - Dual ultrastructural localization of mu-opiate receptors and substance p in the dorsal horn. AB - Opiates active at the mu-opiate receptor (MOR) produce antinociception, in part, through actions involving substance P (SP), a peptide present in both unmyelinated primary afferents and interneurons within the dorsal horn. We examined potential functional sites for interactions between SP and MOR by using dual electron microscopic immunocytochemical localization of antisera against SP and a sequence-specific antipeptide antibody against MOR in rat cervical spinal dorsal horn. The distribution was compared with that of the functionally analogous dorsal horn of the trigeminal nucleus caudalis. Many of the SP immunoreactive terminals in the dorsal horn contacted dendrites that contain MOR (53% in trigeminal; 70% in cervical spinal cord). Conversely, within the cervical spinal dorsal horn 79% of the MOR-labeled dendrites that received any afferent input were contacted by at least one SP-containing axon or terminal. Although SP immunoreactive dendrites were rare, many of these (48%) contained MOR, suggesting that the activity of SP-containing spinal interneurons may be regulated by MOR ligands. A few SP-labeled terminals also contained MOR (12% in trigeminal; 6% in cervical spinal cord). These data support the idea that MOR ligands produce antinociception primarily through modulation of postsynaptic second-order nociceptive neurons in the dorsal horns of spinal cord and spinal trigeminal nuclei, some of which contain SP. They also suggest, however, that in each region, MOR agonists can act presynaptically to control the release of SP and/or glutamate from afferent terminals. The post- and presynaptic MOR sites are likely to account for the potency of MOR agonists as analgesics. PMID- 10700023 TI - Dual control of dorsal raphe serotonergic neurons by GABA(B) receptors. Electrophysiological and microdialysis studies. AB - We assessed the role of GABA(B) receptors in the control of serotonergic (5-HT) neurons of the dorsal raphe nucleus (DRN) by using microdialysis in vivo and intra- and extracellular recording in vitro in the rat. The GABA(B) agonist R(+)baclofen (but not the inactive S(-)enantiomer) enhanced the 5-HT output in the DRN (4. 7-fold at 15 mg/kg s.c.) and, to a much lesser extent, striatum of unanesthetized rats. Phaclofen (2 mg/kg s.c.) antagonized the effects of 6 mg/kg R(+)baclofen in dorsal striatum. Using dual-probe microdialysis, R(+)baclofen (0.1-100 microM) applied in the DRN enhanced the local 5-HT output (4.5-fold at 100 microM) but decreased that in striatum at 100 microM. At concentrations higher than 100 microM there was a moderate decrement in the elevation of 5-HT in the DRN. In midbrain slices, bath R(+)baclofen exerted a biphasic effect on DRN 5 HT neurons. Consistent with a reduced striatal 5-HT release when infused in the DRN, R(+)baclofen (0.1-30 microM) induced an outward current in 5-HT neurons (IC(50) = 1.4 microM). Lower R(+)baclofen concentrations (0.01-1 microM) preferentially reduced GABAergic inhibitory postsynaptic currents induced by N methyl-D-aspartate (20 microM) in 5-HT neurons (IC(50) = 72 nM). Using extracellular recordings, R(+)baclofen (300 nM) enhanced the ability of NMDA to induce firing in a subpopulation of serotonergic neurons. These results are consistent with a preferential activation by a low concentration of R(+)baclofen of presynaptic GABA(B) receptors on GABAergic afferents that could disinhibit 5 HT neurons and increase 5-HT release. PMID- 10700024 TI - Mapping of serotonin 5-HT(4) receptor mRNA and ligand binding sites in the post mortem human brain. AB - The anatomical localization of 5-HT(4) receptor mRNA and 5-HT(4) receptor protein was examined in sections of post-mortem human brain by in situ hybridization histochemistry and radioligand receptor autoradiography. In the in situ hybridization study, the highest levels of 5-HT(4) receptor mRNA were found in caudate nucleus, putamen, nucleus accumbens, and in the hippocampal formation. No 5-HT(4) receptor mRNA was detected in globus pallidus and substantia nigra. For receptor autoradiography, two new and highly selective radioligands were compared: [(3)H]prucalopride, which preferentially labels the G-protein coupled fraction of receptors, and [(3)H]R116712, which labels the entire receptor population at subnanomolar concentrations. [(3)H]Prucalopride and [(3)H]R116712 binding was performed on human brain hemisphere sections. The highest densities for both radioligands were found in the basal ganglia (caudate nucleus, putamen, nucleus accumbens, globus pallidus, substantia nigra). Moderate to low densities were detected in the hippocampal formation and in the cortical mantle. Mismatches between 5-HT(4) receptor mRNA and binding sites in the globus pallidus and the substantia nigra suggested that the binding sites may be localized on axonal projections originating from the striatum. To compare densities of binding sites, concentration binding curves with [(3)H]prucalopride, [(3)H]R116712 and [(3)H]GR113808 were performed on membranes from homogenates of several human brain regions. Comparison of B(max)-values obtained with [(3)H]prucalopride and [(3)H]R116712 indicated that the G-protein coupled fraction of 5-HT(4) receptors in the substantia nigra was exceptionally high (54%) in comparison with percentages (16-27%) found in the frontal cortex, the striatum and the hippocampus. Such a high percentage (40%) of [(3)H]prucalopride vs. [(3)H]R116712 binding was also observed in the substantia nigra in the receptor autoradiography experiments. The [(3)H]prucalopride binding was GppNHp-sensitive, whereas [(3)H]R116712 and [(3)H]GR113808 was not. These data indicate that in the substantia nigra 5-HT(4) receptors are more strongly coupled to their signal transduction pathway than in other brain regions. PMID- 10700025 TI - Dopamine innervation of monkey entorhinal cortex: postsynaptic targets of tyrosine hydroxylase-immunoreactive terminals. AB - Dopamine (DA) has been demonstrated to play an important role in regulating cortical activity in both neocortical and periallocortical regions. However, marked differences between these two types of cortices in the laminar pattern of DA axons, the types and distribution of DA receptors, and the postnatal development of the DA innervation suggest that DA may have region-specific effects. Such regional specialization may also include the types of cortical cells apposed to DA terminals. In neocortical regions, such as the prefrontal and motor cortices, the majority of structures apposed to DA terminals appear to be the dendritic spines and shafts of pyramidal cells, and a minority are dendrites immunoreactive for gamma-amino butyric acid (GABA). However, the identity of the neural elements apposed to DA terminals in the entorhinal cortex, a periallocortical region, is unknown. In this study, we used immunocytochemical techniques and antibodies against tyrosine hydroxylase (TH) and GABA, visualized with preembedding immunoperoxidase and immunogold-silver labels, respectively, to examine DA terminals and their targets with electron microscopy. In the superficial layers of the monkey entorhinal cortex, TH-immunoreactive (IR) terminals varied greatly in size and formed thin, symmetric synapses. The majority of dendritic structures apposed to these TH-terminals were not GABA-IR, and included both dendritic shafts (64%) and spines (14%). A minority (22%) of the apposed dendrites were GABA-IR. A similar distribution of targets was observed for the subset of TH-IR terminals with identifiable synaptic specializations. In addition, the proportions of GABA-labeled and unlabeled dendrites apposed to TH terminals did not differ from those previously reported for monkey prefrontal cortex. These findings indicate that DA terminals provide direct input to both excitatory and inhibitory cells in the monkey entorhinal cortex and suggest that the effects of DA are mediated through a set of targets that are common to both neo- and periallocortex. PMID- 10700026 TI - Detection of the effects of dopamine receptor supersensitivity using pharmacological MRI and correlations with PET. AB - Receptor supersensitivity is an important concept for understanding neurotransmitter and receptor dynamics. Traditionally, detection of receptor supersensitivity has been performed using autoradiography or positron emission tomography (PET). We show that use of magnetic resonance imaging (MRI) not only enables one to detect dopaminergic supersensitivity, but that the hemodynamic time course reflective of this fact is different in different brain regions. In rats unilaterally lesioned with intranigral 6-hydroxydopamine, apomorphine injections lead to a large increase in hemodynamic response (cerebral blood volume, CBV) in the striato-thalamo-cortico circuit on the lesioned side but had little effect on the intact side. Amphetamine injections lead to increases in hemodynamic responses on the intact side and little on the lesioned side in the same animals. The time course for the increase in CBV after either amphetamine or apomorphine administration was longer in striatum and thalamus than in frontal cortex. (11)C-PET studies of ligands which bind to the dopamine transporter (2 beta-carbomethoxy-3-beta-(4-fluorophenyl)tropane 1, 5-naphthalnendisulfonate, WIN 35, 428 or CFT) and D2 receptors (raclopride) confirm that there is a loss of presynaptic dopamine terminals as well as upregulation of D2 receptors in striatum in these same animals. Pharmacologic MRI should become a sensitive tool to measure functional supersensitivity in humans, providing a complementary picture to that generated using PET studies of direct receptor binding. PMID- 10700027 TI - Enhanced cerebral uptake of receptor ligands by modulation of P-glycoprotein function in the blood-brain barrier. AB - Low cerebral uptake of some therapeutic drugs can be enhanced by modulation of P glycoprotein (P-gp), an ATP-driven drug efflux pump at the blood-brain barrier (BBB). We investigated the possibility of increasing cerebral uptake of the beta adrenergic ligands S-1'-[(18)F]-fluorocarazolol (FCAR) and [(11)C]-carazolol (CAR) in P-gp knockout mice (mdr1a (-/-)) and by modulation of P-gp with cyclosporin A (CsA) in rats. Specific and nonspecific binding of FCAR in the rat brain were doubled by CsA, while target/nontarget ratios and clearance from plasma (area under curve (AUC)) were not affected. Cerebral uptake of CAR in rats was much lower than FCAR and nonspecific. CsA increased this uptake 5-6-fold, not only due to P-gp modulation in the BBB but also to a 2-fold higher plasma AUC. In the CNS of mdr1a (-/-) mice, uptake of FCAR and CAR was, respectively, 2-fold and 3-fold higher than in mdr1a (+/+) mice. These results indicate that the cerebral uptake of beta-adrenoceptor ligands can be increased by administration of P-gp modulators such as CsA without affecting regional distribution in the brain. P-gp modulation could improve the count statistics in PET studies of the CNS. PMID- 10700029 TI - Preface PMID- 10700028 TI - 3,4-Methylenedioxymetamphetamine (ecstasy) induces c-fos-like protein and mRNA in rat organotypic dorsal striatal slices. AB - 3,4-Methylenedioxymetamphetamine (MDMA, "ecstasy") is an increasingly abused drug, which has significant effects on the dopamine system in the striatum. The isolated single organotypic slice model allows investigation of the effects of drugs of abuse on the expression of transcription factors in the striatum without dopaminergic and glutamatergic interactions. In this study the effects of MDMA on the expression of c-fos mRNA by in situ hybridization as well as the c-fos-like protein by immunohistochemistry in isolated dorsal striatum was investigated. It was shown that 100 microM MDMA induced c-fos mRNA expression 30 min after treatment. Expression of c-fos-like protein was transiently detected 3 h afterwards. The c-fos expression was inhibited by MK 801 and metoclopramide, indicating the involvement of dopaminergic D2 receptors and glutamatergic NMDA receptors. The dopaminergic D1 receptor antagonist SCH 23390 did not affect c-fos expression. We conclude that MDMA treatment leads to the induction of c-fos expression in isolated rat striatal slices. This effect is independent of extrinsic neuronal circuitry and seems to be associated with direct interactions between MDMA and the dopamine/glutamate receptor system. PMID- 10700030 TI - Characterization of fungal spores by laser desorption/ionization time-of-flight mass spectrometry. AB - A considerable volume of research has now been completed on the application of matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) to the analysis of bacteria; however, to date no definitive studies have been made using this technique on fungi. Preliminary studies on the application of the MALDI-MS methodology, previously developed for the analysis of bacteria, to the analysis of intact fungal spores are described here. MALDI-MS and electrospray mass spectrometry enable the high molecular weight analysis of proteins, glycoproteins, oligosaccharides and oligonucleotides. Using MALDI-MS with bacteria has demonstrated the ability to produce 'fingerprints' of the intact cells with the ions observed being associated with the proteinaceous components of the cell wall. This paper reports the adaptation of this technique to the direct analysis of fungal cells. The high percentage of carbohydrate in the fungal cell wall indicates that the ions observed in the mass spectrometric experiments may be of carbohydrate origin. Penicillium spp., Scytalidium dimidiatum and Trichophyton rubrum have been studied in this preliminary investigation and all show individually distinctive spectra which would appear to provide a profile of the cellular material with discrete peaks being observed over the mass range 2 to 13 kDa. The spectra obtained are reproducible within the method used but, as shown in our previous studies on bacteria, washing may selectively release components from the fungal cell wall. PMID- 10700031 TI - Energy-dependent electrospray ionisation mass spectrometry: applications in transition metal carbonyl chemistry AB - Maps of electrospray mass spectrometry data, plotted as cone voltage versus mass to-charge ratio, provide a clear and compact method of visualising the accompanying fragmentation processes, in this case applied to the sequential removal of ligands from transition-metal carbonyl complexes. The technique is described as energy-dependent electrospray ionisation mass spectrometry (EDESI MS). Copyright 2000 John Wiley & Sons, Ltd. PMID- 10700032 TI - Capillary zone electrophoresis/mass spectrometry of 5-aminolaevulinic acid and porphobilinogen. AB - A capillary zone electrophoresis/electrospray ionisation mass spectrometry (CZE/ESI-MS) method has been developed for the separation and detection of 5 aminolaevulinic acid (ALA) and porphobilinogen (PBG). Capillaries were 70 cm long with an inner diameter of 75 micrometer and outer diameter of 375 micrometer. The buffer used was aqueous ammonium acetate (50mM, pH 5.2) with a co-axial 'make-up' flow of methanol/aqueous 0.1% formic acid (1:1 v/v) at a flowrate of 6 microL/min. A voltage of 20 kV was used for CZE and an ESI voltage of 3.5 kV. Full scan data was acquired over the range m/z 100-500 in positive ion mode, from which selected ion electropherograms were extracted; at m/z 132 for the protonated molecular ion of ALA and m/z 210 for the methylenepyrrolenine fragment ion of PBG. The protonated molecular ion of PBG, m/z 227, was found to be too facile to monitor, easily losing ammonia in the electrospray source and better sensitivity was achieved by monitoring the resulting fragment ion. The detection limits were circa 100 attomoles of ALA and 10 attomoles of PBG at a signal-to noise ratio (S/N) better than 10, providing sufficient sensitivity for clinical use and offering advantages over existing techniques. PMID- 10700033 TI - Identification of organically associated trace elements in wood and coal by inductively coupled plasma mass spectrometry. AB - 1-Methyl-2-pyrrolidinone (NMP) was used to extract samples of wood (forest residue) and coal; the extracts were analysed by inductively coupled plasma mass spectrometry (ICP-MS) using two different sample preparation methods, in order to identify trace elements associated with the organic part of the samples. A sample of fly ash was similarly extracted and analysed in order to assess the behaviour of the mineral matter contained within the wood and coal samples. 32% of the biomass was extracted at the higher temperature and 12% at room temperature while only 12% of the coal was extracted at the higher temperature and 3% at room temperature. Less than 2% of the ash dissolved at the higher temperature. Size exclusion chromatograms of the extracts indicated the presence of significant amounts of large molecular mass materials (>1000 mu) in the biomass and coal extracts but not in the ash extract. Trace element analyses were carried out using ICP-MS on the acid digests prepared by 'wet ashing' and microwave extraction. Sixteen elements (As, Ba, Be, Cd, Co, Cr, Cu, Ga, Mn, Mo, Ni, Pb, Sb, Se, V and Zn) were quantified, in the samples before extraction, in the extracts and in the residues. Concentrations of trace elements in the original biomass sample were lower than in the coal sample while the concentrations in the ash sample were the highest. The major trace elements in the NMP extracts were Ba, Cu, Mn and Zn from the forest residue; Ba, Cu, Mn, Pb and Zn from the coal; Cu and Zn from the ash. These elements are believed to be associated with the organic extracts from the forest residue and coal, and also from the ash. Be and Sb were not quantified in the extracts because they were present at too low concentrations; up to 40% of Mn was extracted from the biomass sample at 202 degrees C, while Se was totally extracted from the ash sample. For the forest residue, approximately 7% (at room temperature) and 45% (at 202 degrees C) of the total trace elements studied were in the extract; for the coal, approximately 8% (at room temperature) and 23% (at 202 degrees C) were in the extract. For the ash, only 1.4% of the trace elements were extracted at 202 degrees C, comprising 25% of Cd but less than 1% of Pb. PMID- 10700034 TI - Electron impact and chemical ionisation mass spectrometry in the characterisation of organometallic silicon, germanium, tin and lead compounds AB - The electron impact and chemical ionisation mass spectra of a number of organometallic derivatives of silicon, germanium, tin and lead, incorporating inter alia 2-bis(trimethylsilyl)methylpyridyl or cyclopentadienyl ligands have been recorded. Representative spectra are presented and illustrate the utility of these mass spectrometric techniques in the characterisation of such compounds, particularly so in light of the highly informative nature arising from their fragmentation patterns and the numerous isotopes of the nuclei involved. Copyright 2000 John Wiley & Sons, Ltd. PMID- 10700035 TI - Determination of trace elements by inductively coupled plasma mass spectrometry of biomass and fuel oil reference materials using milligram sample sizes. AB - Most of the analytical techniques used to quantify elements associated with solid samples suffer from high detection limits and cannot be used for trace elements in biomass samples, particularly when only 20 mg are available for analysis. Inductively coupled plasma mass spectrometry (ICP-MS) can achieve detection limits of parts-per-trillion with liquid sample introduction by solution nebulisation. This technique was therefore tested with two standard biomass reference materials: oriental tobacco leaves and cabbage leaves. Two preparations successfully used on coal standards were used to digest the solid samples: a total digestion method (wet ashing digestion) and a partial leaching (microwave extraction). The concentrations of up to seventeen elements (As, Ba, Be, Cd, Co, Cr, Cu, Ga, Mn, Mo, Ni, Pb, Sb, Se, Sn, V and Zn) were measured after the two preparations. The accuracy and sensitivity of the measurements improved when the dilution factor decreased from 5000 to 1000 and to 500. Since the proportion of mineral matter in biomass samples is small (5%), the microwave digestion extracted elements that are generally not completely extracted from coal samples (e.g. Sb). However, some trace element concentrations were below the limit of quantification after microwave extraction, even with a reduced dilution factor (As, Se and Mo) and could not be quantified. A fuel oil was also digested. The trace element concentrations were very low (between 28 and 0.1 microgram g(-1)) but acceptable results were obtained by applying a dilution factor of 100. Only six elements in the fuel oil (As, Ba, Co, Ni, Se and V) had certified or indicated values. Factors affecting the accuracy and sensitivity of the analyses are discussed. The reproducibility of analysis of the tobacco leaf standard was checked over a period of nine months by both digestion methods. The wet ashing method gave acceptable reproducibility for Ba, Cd, Co, Cu, Ga, Mn, Mo, Ni, Pb, V and Zn but poor precision for Cr, Se and Sn and showed evidence of residual chloride interference for As. The microwave extraction gave good reproducibility for As, Ba, Cd, Co, Cr, Cu, Mo, Ni and Zn but poor precision for Se and low recoveries for Ga, Mn, Sn and V. In spite of the small quantities of material analysed, it proved possible to determine the trace elements at levels down to 0.1 microgram g(-1) in the reference materials. PMID- 10700036 TI - Quantitative in vitro kinase reaction as a guide for phosphoprotein analysis by mass spectrometry. AB - A simple calculation using the radioactive decay of (32)P incorporated into a protein during in vitro kinase reactions is described that allows the overall stoichiometry of phosphorylation for the substrate protein or peptide to be calculated. Prior to using techniques such as diagnostic ion scanning to identify the molecular weight of an unknown phosphopeptide in a complex mixture followed by tandem mass spectrometry (MS/MS) to locate the phosphorylated residue within the phosphopeptide, such calculations are predictive of the chances for successful characterization by these methods. An example of estimating the stoichiometry of peptide phosphorylation will be presented along with calculations that predict when adequate phosphopeptide is present in any given spot on the thin-layer chromatography (TLC) plates used for two-dimensional phosphopeptide (2DPP) mapping to allow extraction and complete characterization by MS/MS. PMID- 10700037 TI - A library of atmospheric pressure ionization daughter ion mass spectra based on wideband excitation in an ion trap mass spectrometer. AB - A searchable library of MS/MS spectra obtained using a quadrupole ion trap mass spectrometer and electrospray or atmospheric pressure chemical ionization is presented. The application of wideband excitation (activation) and normalized collision energy leads to highly reproducible mass spectra which are searched using the NIST algorithm. Flow injection and LC/MS/MS applications of this powerful technique in the biomedical (diastereoisomeric steroids, morphine glucuronides, isovalerylcarnitine) and environmental (pirimicarb and desmethyl pirimicarb) areas are described. PMID- 10700038 TI - Chromatographic aspects in high throughput liquid chromatography/mass spectrometry. AB - Important parameters to consider when developing a liquid chromatography/mass spectrometry (LC/MS) method are buffer type and concentration, and column geometry. In the work presented here the choice of buffer for the analysis of basic compounds using a polar embedded phase (HyPURITYtrade mark ADVANCE) is illustrated for the analysis of tricyclic antidepressants. Method transfer from a 4.6 mm i.d. column to a 2.1 mm i.d. column is demonstrated for the analysis of triazines and anabolic steroids and their metabolites, with no change in selectivity and with added speed of analysis. Analysis of eight beta-blockers is achieved in 65 seconds by using a short 30 x 4.6 mm C18 column. PMID- 10700039 TI - Laser photo-induced dissociation using tandem time-of-flight mass spectrometry AB - A novel tandem time-of-flight (TOF) mass spectrometer has been developed for studying the photo-induced dissociation of large molecules and elemental clusters. It consists of a linear first stage TOF analyser for primary mass separation and precursor ion selection, and a second orthogonal reflecting field TOF analyser for product ion analysis. The instrument is equipped with a large volume throughput molecular beam source chamber allowing the production of jet cooled molecules and molecular clusters, as well as elemental clusters, using either a pulsed laser vaporisation source (LVS) or a pulsed are cluster ion source (PACIS). A second differentially pumped chamber can be used with effusive sources, or for infrared laser desorption of large molecules, followed by laser ionisation. These primary ions can then be irradiated with a second, high energy laser to induce photodissociation. Detailed information about the fragmentation mechanisms can be deduced from the product ion mass spectra. Preliminary results on the photo-induced dissociation (PID) of the molecule ion of aniline at 266 nm are presented. In this case the molecule ions were generated via two-photon laser ionisation at 266 nm using an effusive source. Results for the collision-induced dissociation (CID) of the aniline molecule ion, using a commercial mass spectrometer equipped with an atmospheric pressure electrospray ionisation interface, are also presented. Copyright 2000 John Wiley & Sons, Ltd. PMID- 10700040 TI - Generation and detection of PMID- 10700041 TI - Introduction to histological studies of gastric secretion PMID- 10700042 TI - Innervation of the gastric mucosa. AB - A plethora of neuronal messengers ("classical" transmitters, gaseous messengers, amino acid transmitters, and neuropeptides) are capable of mediating or modulating gastric functions. Accordingly, the stomach is richly innervated. Gastric nerves are either intrinsic to the gastric wall, i.e., they have their cell bodies in the intramural ganglia and thus belong to the enteric nervous system, or they reach the stomach from outside, originating in the brainstem, in sympathetic ganglia, or in sensory ganglia. Topographically, the nerve fibers in the stomach reach all layers from the most superficial portions of the gastric glands to the outer smooth muscle layer. This wide distribution implies that virtually all different cell types may be reached by neuronal messengers. Within the gastric mucosa endocrine and paracrine cells (e.g., gastrin cells, ECL cells, somatostatin cells), exocrine cells (parietal cells, chief cells, mucous cells), smooth muscle cells, and stromal cells are regulated by neuronal messengers. The sensory innervation, responding to capsaicin, plays an important role in mucosal protection, and in ulcer healing. Presumably also other nerves are involved and a plasticity in the neuropeptide expression has been demonstrated at the margin of gastric ulcers. Taken together, available data indicate a complex interplay between hormones, paracrine messengers and neuronal messengers, growth factors and cytokines in the regulation of gastric mucosal activities such as secretion, local blood flow, growth, and restitution after damage. PMID- 10700043 TI - Classification of gastric endocrine cells at the light and electron microscopical levels. AB - This review discusses the current concepts for the classification of gastric endocrine cells subdivided according to the type of mucosa in which they are located. In the oxyntic mucosa, the most important cell type is the ECL cell, involved in the synthesis and secretion of histamine. Proteins involved in many aspects of the biology of ECL cells including the response to the gastrin stimulus, membrane transport and docking, prevention of apoptosis, calcium homeostasis, autocrine activity, and maintenance of the differentiated cell phenotype have been localized to this cell type. Other cells of the oxyntic mucosa include: the D and EC cells producing somatostatin and serotonin, respectively, delivered through long cell processes; the X (or A-like) cells, possibly producing endothelin; and the D(1) and P cells of unknown function and possibly representing morphological variants of other cell types. In the antral mucosa, the three important cell types are represented by: the gastrin-producing G cells; the somatostatin-producing D cells, which are anatomically and functionally associated with G cells; and the serotonin-producing EC cells, which are located at the bottom of antral glands. PMID- 10700044 TI - Developmental biology of gastrin and somatostatin cells in the antropyloric mucosa of the stomach. AB - Gastrin is a hormone regulating gastric acid secretion and the growth of the gastrointestinal epithelium. It is expressed by endocrine tumors and by adenocarcinomas of the gastroenteropancreatic region and may represent an autocrine tumor growth factor. Gastrin is also implicated in the genesis of peptic ulcer disease both in conjunction with H. pylori infections and with gastrin-producing tumors. The secretion and expression of gastrin are under the paracrine control of somatostatin, produced by D cells situated in close contact with gastrin-producing G cells. D cells also contain neuronal nitric oxide synthase and appear to regulate apoptosis of G cells by paracrine release of nitric oxide. Both G and D cells are derived from a common multihormonal precursor cell present in the regenerative (isthmus) region of the gastric units. The precursor cells have been suggested to undergo asymmetrical divisions resulting in gastrin- and somatostatin-producing daughter cells that remain in paracrine contact during their migration into the glands. The precursor cells also give rise to the third main antropyloric endocrine cell type; the serotonin producing EC cell. The maturation of all of these cell types is regulated by a number of transcription factors containing homeobox motifs (Pdx-1, Pax 4 and 6, Isl-1, Nkx6.1). Many of these also regulate the development of the central nervous system and the pancreas. The use of different combinations of these factors for regulating the expression of different hormones may explain the phenomenon of abberant hormone expression during development and carcinogenesis and the occurrence of multihormonal cells. PMID- 10700045 TI - Morphological studies on the translocation of tubulovesicular system toward the intracellular canaliculus during stimulation of the gastric parietal cell. AB - The gastric parietal has two characteristic membrane systems. One is the intracellular canaliculus, which is specialized networks of enfolded luminal membrane channels lined with numerous microvilli. The other structures common to all parietal cells are the tubulovesicles or the tubulovesicular membranes, a system of tubules and vesicles. The tubulovesicular compartment is drastically depleted during maximal gastric acid secretion and this is coincident with an increase in the canalicular cell surface membrane. A plausible explanation for this redistribution is the fusion and transfer of tubulovesicular membranes to the plasma membrane. However, for many years there was no convincing evidence of connections between these two membrane systems. The mechanism of the transformation of tubulovesicular membrane into the plasma membrane without demonstrable connections has been an enigma to electron microscopists. Using a recently developed fixation technique for parietal cells [Sugai et al. (1995) Acta Anat Nippon 74:S101], we have investigated the organization of the cytoplasmic membrane systems in the rat resting and tetragastrin stimulated stomachs by ultra-high-resolution scanning electron microscopy (SEM). Gastric mucosae were microwave-fixed in a cacodylate buffer, (334 milliosmoles/kgH(2)O (mOsm)), to which 1.0% glutaraldehyde and 0.5% formaldehyde were added. Specimens examined by TEM of thin sections revealed the cytoplasm packed with tubular membranes similar to images detected by rapid-freeze/freeze-substitution fixation. To render the cytoplasmic membranes visible by SEM, fixed mucosae were treated by the aldehyde-osmium-DMSO-osmium maceration procedure. With much of the cell matrix and filaments removed, SEM revealed numerous 30-60-nm tubules, which formed a meshwork with small cisternae. Vesicles or isolated tubules were not found in adequately macerated parietal cells. The cytoplasmic surface of the intracellular canaliculus was smooth except for round openings representing the bases of macerated microvilli. In favorable sites, connections of the tubular membranes to the canaliculi were clearly visible. Stereo pair views were particularly useful to demonstrate these continuities. Connections between these two membrane compartments suggest the probability of rapid membrane transposition. In this article, the form and distribution of membrane systems of parietal cells in the resting state and after tetragastrin stimulation will be presented and discussed. Special emphasis is made to demonstrate connections between the tubulovesicular system and the intracellular canaliculus. PMID- 10700046 TI - Establishment of culture systems of human gastric epithelium for the study of pepsinogen and gastric lipase synthesis and secretion. AB - A main purpose of gastric secretion pertains to the digestion of dietary proteins and involves the release of pepsinogens by the fundic and antral mucosa. Over the last decade, data on human gastric physiology has expanded to equally include a significant role in fat digestion. Characteristics of human gastric lipase (HGL) such as optimum acid pH, resistance to proteolysis and non requirement of bile salts or cofactors, are advantageous in gastric lipolysis. Furthermore, the importance of HGL increases in the context of perinatal physiology and pathological situations where secretion of HGL could compensate, to some extent the depressed pancreatic activities. It is therefore important to understand the regulatory mechanisms involved in the synthesis and secretion of human gastric digestive enzymes. The establishment of an organ culture technique as well as a novel primary culture system of human gastric epithelium permitted us to demonstrate that Pg5 and HGL are colocalized in human chief cells and both digestive enzymes are efficiently synthesized and secreted in explants and primary cultures. Pepsin activity rises at the cellular level while its secretion remains constant. In contrast, cellular lipase activity drastically diminishes while being preferentially secreted. This nonparallelism supports the concept that Pg5 and HGL are differently regulated in culture. Furthermore, EGF downregulates HGL expression at the mRNA level via the p42/44(MAPK) pathway without affecting Pg5. Future studies should be designed to fully understand the cellular and molecular mechanisms involved in regulating HGL activity in normal and pathological conditions. PMID- 10700047 TI - Role of apoptosis in gastric epithelial turnover. AB - Gastric epithelial turnover is a dynamic process. It is characterized by continuous cell proliferation, which is counterbalanced by cell loss. The biological principle that mediates the homeostasis of epithelium is programmed cell death, or apoptosis. Currently, several subtypes of apoptosis are distinguished, which are mediated by different mechanisms. Various subtypes of apoptosis also occur in the gastric epithelium under various conditions. In the normal stomach, apoptosis due to cell isolation (anoikis) mediates the physiological epithelial turnover. Albeit rarely seen in routine histology, approximately 2% of epithelial cells in the normal stomach are apoptotic. In Helicobacter pylori-induced gastritis, apoptosis and epithelial proliferation are moderately increased, with approximately 8% apoptotic epithelial cells. In gastritis, factors such as CD95 ligand or tumor necrosis factor (TNF) alpha act as death factors. They bind to specific receptors, CD95 and TNF-R, which are induced either by other cytokines, such as interferon gamma, or by Helicobacter pylori itself. In addition to CD95, H.pylorican also induce upregulation of CD95 ligand expression. Taken together, the upregulated expression of CD95, and the presence of CD95L in the close proximity to apoptotic gastric epithelial cells suggest a functional role of the CD95-CD95L system in the induction of apoptosis in H.pylori-gastritis. The role of other pathways to apoptosis is currently under study. Apart from being a biological phenomenon, apoptosis in the stomach may also have direct clinical consequences. An extreme example is given in gastric graft-vs.-host disease when epithelial denudement occurs. PMID- 10700048 TI - Outcome of patients with acute myocardial infarction who are ineligible for primary angioplasty trials. AB - We determined acute outcome in 148 consecutive patients with ST segment elevation myocardial infarction undergoing angioplasty including 72 patients (48.7%) considered ineligible for primary angioplasty trials. Overall, in-hospital mortality for acute infarct angioplasty was 12%, with fivefold higher mortality in the trial-ineligible group (21% vs. 4%, P = 0.003). Thus, primary angioplasty trials continue to exclude nearly 50% of acute infarction patients and reported mortality rates of primary angioplasty trials are likely to be significantly lower than the unselected in-hospital mortality rates. Cathet. Cardiovasc. Intervent. 49:237-243, 2000. PMID- 10700049 TI - The power of (un)natural selection. PMID- 10700050 TI - Procedural costs of digital vs. analog archiving of diagnostic cardiac catheterizations. AB - The use of digital technology in the cardiac catheterization laboratory is expanding at a rapid pace. The cost-effectiveness of this new technology is yet to be proven. The aims of this study were to determine the direct cost differences of digital versus analog media (CDs) for the storage of diagnostic cardiac catheterizations and to explore the factors influencing these differences. Procedural costs of all diagnostic angiograms (n = 109), from three physicians, performed in an analog catheterization laboratory (room A) and a digital catheterization laboratory (room C) were compared during a 9-month period. The mean procedural cost was higher in room A than in room C ($1,102 vs. $1,087, P < 0.001). This cost difference was eliminated when recording media costs were excluded from analysis ($1,079 vs. $1,080, P = 0.931). Therefore, we conclude there is a procedural cost savings in a cardiac catheterization room that uses digital CDs versus cineangiogram film as the archival media. Cathet. Cardiovasc. Intervent. 49:246-250, 2000. PMID- 10700051 TI - The cost of doing business in a digital age. PMID- 10700052 TI - Response of the radial artery to three vasodilatory agents. AB - We examined the response of the radial artery to vasoactive agents (isosorbide dinitrate, ISDN, 1 mg, 3 mg, and 5 mg; verapamil, 1 mg, 3 mg, and 5 mg; and lidocaine, 10 mg, 30 mg, and 50 mg) in 100 consecutive patients admitted for elective coronary angiography. The drug solutions were directly injected into the radial artery from the puncture site. As a result, 5 mg of ISDN increased the diameter of the radial artery by 31% and 28.8% at the proximal and distal sites, respectively. Similarly, 5 mg of verapamil increased it by 9% and 10.8% at the proximal and distal site, respectively. But 10 mg of lidocaine decreased it by 15.6% and -12.1% at the proximal and distal site, respectively. At the doses utilized, ISDN was the most potent vasodilator for the radial artery and lidocaine caused paradoxical vasoconstriction. PMID- 10700053 TI - The radial artery approach: we have come such a long way for the long way. PMID- 10700054 TI - Percutaneous transluminal coronary angioplasty of chronic total occlusions. Determinants of primary success and long-term clinical outcome. AB - This study was conducted to assess the determinants of the procedural success and long-term clinical benefits of percutaneous transluminal balloon angioplasty (PTCA) of chronic total occlusion (CTO) in recent years. Two hundred and twenty six consecutive patients who underwent PTCA of CTO were divided into two groups according to the procedural success (n = 134) or failure (n = 92). Both groups were analyzed in terms of the initial success, predictors of procedural failure, and clinical outcome. The procedural success rate was noted to have improved to more than 70% since 1995. A multiple logistic regression analysis revealed that the presence of calcification, the length of the occlusion and the presence of multivessel disease were independent predictors of procedural failure. Cardiac death and the need for coronary surgery were significantly less frequent in patients with procedural success than in those with procedural failure. In properly selected cases, the success rate of PTCA of CTO is acceptable. Long-term clinical benefit is suggested by the high rate of freedom from coronary surgery and the low cardiac death rate in the patients who underwent successful revascularization. PMID- 10700055 TI - Advances and ongoing problems in the management of chronic total occlusion. PMID- 10700056 TI - Stent placement for ostial left anterior descending coronary artery stenosis: acute and long-term (2-year) results. AB - This study was performed to assess the acute and long-term results of elective stenting for the treatment of ostial left anterior descending coronary artery (LAD) stenosis. One hundred and eleven consecutive patients with ostial LAD stenting were included for this study. Follow-up angiography was performed at 6 months and clinical evaluation at regular intervals after stenting. Procedural success rate was 97.3%. Four patients developed non-Q myocardial infarction and one patient underwent emergency bypass surgery due to a large dissection after stenting. Angiographic restenosis rate was 26.1% (18/69), and target lesion revascularization rate 11.7%. The final luminal diameter after stenting was the only predictor of angiographic restenosis. Clinical follow-up was obtained in all patients at 21.5 +/- 16.0 months. Two patients died during the follow-up. Event free survival rate was 84.6 +/- 3.8%. In conclusions, stenting with or without debulking atherectomy may be considered as an acceptable therapeutic option for the treatment of ostial LAD stenosis. PMID- 10700057 TI - The illusive ostial and proximal LAD. PMID- 10700058 TI - Stenting of bifurcation lesions: classification, treatments, and results. AB - Percutaneous transluminal balloon coronary angioplasty (PTCA) of coronary bifurcations is associated with a low success rate, high rate of complications, and high incidence of target vessel revascularization (TVR). The strategy of systematic coronary stenting in bifurcation lesions involving a side branch >/= 2.2 mm in diameter was prospectively evaluated in a single-center observational study during a 35-month inclusion period. All patients meeting these criteria were consecutively included. Bifurcation lesions and treatment were predefined in the study. The study included 366 patients (12.1% of PTCA) with 373 bifurcation lesions, mean age 63.7 +/- 11.6 years, 79.2% male, 46.7% with unstable angina, and 8.3% acute MI. The left anterior descending/diagonal bifurcation was involved in 55.2% of cases, circumflex/marginal 22. 2%, PDA/PLA 10.4%, left main bifurcation in 6.8%, and others 5.4%. The main branch (2.78 +/- 0.42 mm reference diameter) was stented in 96.3% of cases and the side branch (2.44 +/- 0.43 mm) in 63.2% (the two branches were stented in 59.5% of cases). Procedural success was obtained in 96.3% in both branches and 99.4% in the main branch. At1-month follow up, The major cardiac event rate (MACE) was 4.8% (death 1.1%, emergency CABG 0.6%, Q-wave MI 0.9%, acute or subacute closure 1.4%, repeat PTCA 1.1%, and non-Q wave MI 2.3%). At 7-month follow-up, the total MACCE rate was 21.6%, including a TVR rate of 17.2%. Analysis of the 7-month outcome according to two study periods (period I, 1 January 1996 to 31 August 1997, 182 patients; period II, 1 September 1997 to 30 June 1998, 127 patients) showed that the TVR rate decreased from 20.6% to 13.8% (P = 0.04) and the MACE rate from 29.2% to 17.1% (P < 0.01) in period I and II, respectively. This was associated by univariate analysis with an increasing use of tubular stents deployed in the main branch (94.2% vs. 59.1%, P < 0.001) and kissing balloon inflation after coronary stenting (75.4% vs. 18.1%, P < 0.001). Bifurcation lesions are frequent. Procedural success of coronary stenting is high with a low rate of in-hospital MACE. TVR rate at follow-up is relatively low. In-hospital and follow-up results are influenced not only by the learning curve but also by the use of tubular stents in the main branch and final kissing balloon inflation. PMID- 10700059 TI - Ten-years clinical follow-up following successful percutaneous transvenous mitral commissurotomy: single-center experience. AB - The purpose of this study is to report the long-term follow-up outcome of patients undergoing percutaneous transvenous mitral commissurotomy (PTMC). The follow-up of 68 of 82 (83%) consecutive patients undergoing successful PTMC (mitral valve area of more than 1.5 cm(2) without major complications) in 1987 using the Inoue balloon was analyzed. The mean age at the time of PTMC was 52 +/- 11 years and 81% were female patients. The mean follow-up interval was 98 +/- 37 months (6 to 123). Actuarial survival rate was 98%, 97%, and 86% at 1, 5, and 10 years, respectively; the event-free (death, mitral valve replacement, and repeat PTMC) survival rate was 90%, 85%, and 66% at 1, 5, and 10 years, respectively. According to the echocardiographic findings, patients could be divided into three groups: pliable valve, semipliable valve, and rigid valve. Multivariable analysis identified echocardiographic subgrouping as the major significant predictor of any event: the event-free survival rate being 70% in group 1, 66% in group 2, and 20% in group 3 (P < 0.05). Echocardiographic follow-up was available in 49 of 68 patients (72%); the mitral valve area changed from 1.4 +/- 0.5 before to 2.1 +/- 0.4 immediately post-PTMC, and 1.8 +/- 0.4 cm(2) 10 years after the procedure. The long-term follow-up outcome following successful PTMC was favorable and seems to support it as a viable alternative to surgical commissurotomy in selected patients. Patients with rigid valves should be selected very carefully. PMID- 10700060 TI - Follow-up after percutaneous balloon mitral commissurotomy: hard data or first looks? PMID- 10700061 TI - Evaluation of left ventricular ejection fraction as a measure of pump performance in patients with chronic mitral regurgitation. AB - Left ventricular (LV) ejection fraction may not adequately detect a reduction in LV systolic performance resulting from chronic mitral regurgitation (MR), due to ventricular unloading into the low-impedance left atrium. To determine whether LV ejection fraction sufficiently gauges myocardial function in MR, nine patients were studied using micromanometer-measured LV pressures and biplane cineventriculography before and 1 year after mitral valve surgery. Six control patients were also studied. LV ejection fraction was normal in MR patients, despite an increase in LV end-systolic volume index. LV end-systolic pressure volume and stress-volume ratios in MR patients were lower than in controls (P < 0.05 and P < 0.01), suggesting that LV systolic performance fell. One year after mitral valve surgery, LV ejection fraction decreased (P < 0.05) even though LV end-systolic volume index (P < 0.05), pressure-volume (P < 0.05), and stress volume ratios (P < 0.01) all improved. Thus, LV ejection fraction inadequately reflected LV systolic function in MR patients before and after mitral valve surgery. PMID- 10700062 TI - A technique to prevent newly implanted stent displacement during subsequent catheter and sheath manipulation. AB - We describe a novel technique to prevent the displacement or migration of a newly implanted stent as a consequence of any subsequent catheter and sheath manipulation during the same catheterization procedure. The technique involves reinflation of the dilation balloon within the stent immediately after implant followed by advancing the long delivery sheath carefully over the balloon as the balloon is slowly deflated within the stent. The technique was used successfully in 78 stents in 30 patients without stent dislodgment or migration. PMID- 10700063 TI - Transcatheter closure of large PDA using 0.052" gianturco coils: controlled delivery using a bioptome catheter through a 4 French sheath. AB - For over 30 years, a number of devices have been used for transcatheter vascular occlusion procedures, with varying degrees of success. The most prevalent shortcoming with transcatheter occlusion devices is the lack of device control during implant. In patients with congenital heart defects, transcatheter occlusion is complicated by the wide range of vessel sizes, and various anatomical defects. Gianturco embolization coils are very effective and inexpensive occlusion devices. We report a new procedure for transcatheter vascular occlusion using readily available products (Gianturco coil, bioptome, long sheath) that provides complete control of the coil during implant and excellent results. PMID- 10700064 TI - Transcatheter coil occlusion of the small patent ductus arteriosus (<4 mm): improved results with a "multiple coil-no residual shunt" strategy. AB - We report our experience with transcatheter occlusion of the small PDA using Gianturco coils comparing a single coil strategy to a "multiple coil-no residual shunt strategy". Fifteen patients (Group I) had a single coil only placed irrespective of residual shunting and 20 (Group II) were treated using the no residual shunt strategy. Age, minimal PDA diameter, PDA length and PDA types were similar between groups. Closure rates in Group I patients were 60%, 80% and 87% at <1 month, 6 months and 1 year, respectively. In Group II, the <1 month and 6 month closure rates were 100%. The costs and hospital charges for coil closure were comparable to a concurrent surgical group; the total charges (hospital plus physician) were less for Group I, but similar between Group II and the surgical group. The complication rate for coil closure was significantly less than surgical closure. From these data, transcatheter closure with multiple coils can achieve the same closure rate as surgery at similar hospital charges with fewer complications. PMID- 10700065 TI - Balloon positioning difficulties during nonsurgical septal reduction therapy in a patient with hypertrophic obstructive cardiomyopathy. AB - Dual chamber (DDD) pacing and catheter-based nonsurgical septal reduction therapy (NSRT) with ethanol are evaluated for treatment of patients with hypertrophic cardiomyopathy. This report describes a patient with hypertrophic cardiomyopathy and left ventricular outflow tract obstruction who had failed to respond to DDD pacing but showed benefit from subsequent NSRT. Procedural difficulties during NSRT due to massive septal hypertrophy are presented. PMID- 10700066 TI - A case of multiple spontaneous coronary artery dissections. AB - Spontaneous coronary artery dissections are relatively uncommon. Many of the previously reported cases have involved a dissection in a single coronary artery of a peripartum or postpartum female. We describe an unusual case of multiple spontaneous coronary dissections in a middle-aged male. PMID- 10700067 TI - Percutaneous stenting of right pulmonary artery stenosis in fibrosing mediastinitis. AB - Pulmonary artery stenosis is an uncommon complication of fibrosing mediastinitis. Previous medical and surgical therapies have provided limited clinical efficacy without objective evidence of clinical improvement. With the advantages of limited invasiveness and absent need for prolonged drug therapy, percutaneous stent deployment to relieve pulmonary artery obstruction represents a novel treatment for this rare disorder. PMID- 10700068 TI - Combination of aneurysm and myocardial bridging at the same site of a coronary artery in a patient with obstructive hypertrophic cardiomyopathy. AB - This case report describes the rare finding of myocardial bridging and a coronary aneurysm in the same coronary artery segment of a 57-year-old patient with obstructive hypertrophic cardiomyopathy. At the site of the aneurysm in the proximal LAD, the myocardial bridging resulted in an almost normal vessel diameter during systole with an aneurysmatic expansion of the vessel during diastole. This accidental finding does not necessarily require special therapy, since the underlying coronary aneurysm is small, with a low risk of thrombus formation or rupture, but it is big enough to prevent a coronary obstruction due to the myocardial bridging. PMID- 10700069 TI - Congenital bilateral pulmonary venous stenosis in an adult:: diagnosis by Echo Doppler. AB - A 22-year-old man with life-long exertional fatigue and dyspnea was diagnosed as having bilateral congenital pulmonary venous stenosis by echocardiography with color Doppler examination. Fibrous membranes overlying the entrances of the veins to left atrium were the cause of obstruction and were easily resected. PMID- 10700070 TI - Air embolism in the right coronary artery occurring during the left coronary angioplasty using the guiding catheter with a side hole. AB - Coronary air embolism is one of the inadvertent complications of coronary angioplasty. We report two rare cases of complicating air embolism in the right coronary artery occurring during control left coronary angiography using a guiding catheter with a side hole, just prior to a coronary intervention procedure for a left coronary artery lesion. The air seemed to be injected into the right coronary artery through the side hole. When we use an angiographic or guiding catheter with a side hole, we should be aware that an air embolism can occur in the contralateral coronary artery and should carefully and repeatedly perform aspiration of the catheter. PMID- 10700071 TI - Transjugular approach to concurrent mitral-aortic and mitral-tricuspid balloon valvuloplasty. AB - A transjugular approach was successfully used for concurrent mitral-aortic and mitral-tricuspid valvuloplasty in one patient each. This approach simplifies antegrade transvenous aortic valve dilatation in rheumatic aortic stenosis. Advantages obtained by transjugular tricuspid valvuloplasty are easy crossing of the tricuspid valve and stable balloon position, co-axial with the tricuspid orifice. PMID- 10700072 TI - Percutaneous transseptal balloon mitral valvuloplasty: transfemoral vs. transjugular approaches. PMID- 10700073 TI - Late thrombosis following intracoronary brachytherapy. AB - Vascular brachytherapy has been the subject of an extensive ongoing investigation into the safety and efficacy of this technique for preventing restenosis. Preclinical studies have demonstrated reduction of the neointimal proliferation and the late vascular constriction with radiation therapy. However, radiation is also known to delay the healing process and may contribute to a new phenomenon of late thrombosis. We report on two cases of patients with in-stent restenosis who underwent intervention followed by intracoronary vascular radiation therapy (utilizing beta and gamma radiation) and presented with acute onset of unstable angina. Angiographic study demonstrated late thrombosis, which were treated successfully with the Angiojet thrombectomy device. PMID- 10700074 TI - Mid-term clinical and angiographic follow-up outcome after placement of a new balloon expandable stent in native coronary arteries. AB - The widely disparate characteristics that exist among the different stent designs currently available for clinical use may impact on their acute and late angiographic and clinical results. The BeStent (Medtronic Instent, MN) is a relatively new stainless steel, laser-cut, serpentine stent design with only very limited data regarding its performance. In this report, we examined the results of 74 consecutive patients (54 men, 20 women; mean age, 58 years) treated with 76 BeStents in 75 native coronary arteries with a mean reference size of 2.8 mm. Successful stenting without 30-day major adverse cardiac complications was achieved in 97.3% of procedures, resulting in a significant improvement in diameter stenosis from 85% to 2% (P = 0.0001). Six-month angiographic restudy in 88% of patients revealed a per-lesion in-stent restenosis rate of 27%. At a mean follow-up period of 9.3 months, there were no deaths or myocardial infarctions. In summary, the present study demonstrates that the BeStent has an excellent performance profile, is associated with a low risk of stent thrombosis, and yields an acceptable restenosis rate despite the inclusion of a high proportion of patients with diabetes (41%) and small vessels (< 3.0 mm in diameter; 77%). PMID- 10700075 TI - Infectious complications related to the use of the angio-seal hemostatic puncture closure device. PMID- 10700076 TI - Infectious complications related to the use of the angio-seal hemostatic puncture closure device PMID- 10700078 TI - Reply to the letter to the editor by gilchrist PMID- 10700077 TI - Transradial technical tips. PMID- 10700079 TI - "Inadvertent stenting of left main coronary artery complicated by laterin-stent restenosis". PMID- 10700080 TI - Coronary artery fistula closure in an infant. PMID- 10700081 TI - Radiation exposure to children during transcatheter occlusion of the patent ductus arteriosus. PMID- 10700082 TI - Changes in plasma catecholamine and corticosterone levels and gene expression of key enzymes of catecholamine biosynthesis in partially hepatectomized rats. AB - OBJECTIVE: To reexamine the possible role of catecholamines and corticosterone in the early period of liver regeneration after partial hepatectomy (PH)in conscious cannulated rats under carefully controlled conditions which would allow to obtain reliable information about sympathetic-adrenomedullary function after PH in the rat in vivo. METHODS: Plasma levels catecholamines (epinephrine - EPI, norepinephrine - NE) were estimated by radioenzymatic assay and these of corticosterone by competitive protein binding assay. The total RNA was isolated from the adrenals and tyrosine hydroxylase (TH) mRNA expression was estimated by hybridisation with cDNA after Northern blot. The level of immunoreactive protein was measured by using a monoclonal antibody to rat TH, visualized by Western light chemiluminescent detection system and analyzed by densitometry. The level of TH in adrenals was estimated with the aid of 3H-tyrosine and TH cofactor DL-6 methyl-5,6,7,8-tetrahydropterine and the formed radioactive water was measured by scintillation spectometry. RESULTS: The plasma level of norepinephrine (NE), epinephrine (EPI) and corticosterone rapidly increased 20 min. after PH or sham operation (laparotomy). Although the increase of plasma NE was about the same after both PH and laparotomy, that of EPI and corticosterone in PH rats was significantly higher as compared to the laparotomy. One hour after the surgery plasma NE levels in both groups decreased to the basal value and remained still unchanged 4 and 24 h later. At the interval of 4h the plasma level of EPI was higher than in laparotomized controls, but after 24 h the EPI levels returned to basal values. Adrenal tyrosine hydroxylase (TH) mRNA level was significantly elevated in both PH and laparotomized rats, however 24 h after the surgery they returned to the baseline. Adrenal TH immunoprotein levels and TH activity were significantly elevated in both groups 4 h after the surgery, while 24 h later they returned to the baseline in laparotomized rats bur remained elevated in PH rats. Adrenal phenylethanolamine N-methyl-transferase (PNMT) mRNA levels were increased 4 h after both the PH and laparotomy and declined within 24 h. CONCLUSIONS: The first peak of catecholamine and corticosterone levels might result from unspecific stressor associated with the surgery. These levels could be accompanied with the mechanism of the rat liver regeneration. Prolonged elevations of EPI found after PH seems to be specific for liver regeneration indicating that the rise in the adrenal TH mRNA appears to be translated into immunoreactive protein which further leads to the elevation of TH activity. These results contrast markedly with previous studies indicating that the regeneration is modulated predominantly by norepinephrine. PMID- 10700083 TI - Triiodothyronine stimulates 3beta-hydroxysteroid dehydrogenase activity in the porcine corpus luteum. AB - OBJECTIVE: To study the mechanism of thyroid hormone action on the activity of 3beta-hydroxysteroid dehydrogenase in the porcine corpus luteum. METHODS: Pig ovaries were obtained from slaughterhouse animals. Luteal cells were isolated from mid-developing (5-7 days after ovulation) corpora lutea and incubated for 24 h with or without triiodothyronine. Trilostane, an inhibitor od 3beta-HSD that blocks the conversion of pregnenolone to progesterone, was added to the medium in doses of 0,0.1, 1, 10 and 100 micromol. Each treatment was performed in triplicate and each culture system was set up in triplicate. Progesterone concentrations in culture media were determined by radioimmunoassays. RESULTS: Trilostane in a dose of 100 microM significantly decreased the basal progesterone secretion from luteal cells by 26% (P<0.05). However, such secretion was increased by triiodothyronine (T3) in a dose of 10(-9) M. In addition, in T3 treated cells dose dependent inhibitory effect of trilostane on progesterone secretion was observed. Control cultures grown in control medium revealed a relatively weak 3beta-HSD activity which, however, markedly increased after the addition of T3 to the culture medium. Trilostane remarkably decreased 3beta-HSD activity in T3-stimulated cells. CONCLUSION: It was found that T3 acts on luteal cell steroidogenesis via the activation of 3beta-hydrosysteroid dehydrogenase in these cells. PMID- 10700084 TI - Role of substance P in central control of ovulation in female rats. AB - OBJECTIVE: To investigate the influence of substance P on gonadotropin release from the pituitary as evaluated according to the final effect, i.e. the ovulation. METHODS: Stainless steel tube was implanted into the 3rd cerebral ventricle and substance P (SP), C-terminal hexapeptide of Substance P (SP6-11), Gn-RH and adrenaline were infused on the 3rd day of the cycle (proestrus). The oviducts were then isolated on the day of estrus and the ova were counted. RESULTS: SP or SP6-11 inhibited the ovulation which was not prevented by intracerebroventricular (i.c.v.) capsaicin pretreatment. The i.c. v. administration of noradrenaline on the 7-8th day of pseudopregnancy induced the ovulation which was prevented by the injection of SP immediately before noradrenaline. CONCLUSIONS: It is suggested that noradrenergic neuros which control the ovulation by influencing the release of Gn-RH into pituitary portal vessels are affected by SP-ergic neurons. PMID- 10700085 TI - Effect of octreotide acetate on thyrotropin-secreting adenoma: report of two cases and review of the literature. AB - OBJECTIVE: To present two cases with thyrotropin-secreting adenoma and the effectiveness of octreotide acetate treatment on their tumor size as well as on thyroid stimulating hormone (TSH) and thyroid hormone levels. CASE REPORTS: The first case presented with tremor, palpitations and sweating as suggestive of hyperthyroidism, but the other one presented with predominantly headache, while the other symptoms such as palpitation and nervousness were less prominent and he also did not have any thyroid enlargement at physical examination. Thyroid hormone levels in both cases were increased. However, TSH levels were not suppressed thus indicating an inappropriate secretion of TSH. Moreover, TSH levels did not change after T3 and TRH administration, which also contributed to the assumption of an inappropriate TSH secretion. One case had no increase in the TSH alpha subunit level, while this was increased in the other one. Both magnetic resonance imaging and somatostatin receptor scintigraphy revealed that there was a microadenoma (the first case; 6 x 7 mm in diameter)and a macroadenoma (the second case; 14 x 18 mm in diameter). Both patients were placed on a therapy with somatostatin analog octreotide (Sandostatin, Sandoz). Octreotide was initially given at a dose of 300 microg daily and then increased gradually up to 600 microg per day. There was some decrease in the levels of TSH and thyroid hormones at first. However, such decreases did not persist with ongoing therapy for 6 months. In addition, there was no change in the tumor size with this therapy at the end. CONCLUSIONS: We conclude that the treatment by somatostatin analogue octreotide may not be an effective means of reducing the pituitary tumor size, though it may be used to reduce TSH and thyroid hormones temporarily. PMID- 10700086 TI - Localisation of a 40kDa protein in rat steroid producing cells identified by a monoclonal antibody. AB - OBJECTIVE: To localize (by the light and electron microscopy) and partially characterize the antigen recognized by the Mab4E6 in rat ovaries. METHODS: Monoclonal antibody (4E6) against a rat ovarian granulosa cell antigen was prepared and identified the 40kDa protein specific for rat steroid producing cells. The localization of this antigen was studied by light and electron microscopic immunocytochemistry. RESULTS: The immunocytochemical observation suggested that the recognized antigen was localized in granulosa and thecal cells in all stages of follicular development. The intensity of immunostaining was found to depend on the developmental stage. In granulosa and thecal stage (health follicle) Mab 4E6 binding molecule was localized on the membranes of rough endoplasmic reticulum (RER) and on the surface of lipid droplets in close association with the RER. In atretic follicles we established that the final destination of the visualized antigen is in structures which we refer as the autophagic vacuoles in close contact with the steroidogenic organelles. In addition, we observed Mab 4E6 binding molecule in the cytoplasm of luteal cells. Leydig cells and adrenocortical cells. CONCLUSIONS: The results indicate that the 40kDa antigen may be common to all of rat steroidogenic organs. Our results suggest that the 40kDa protein may be associated with the processes governing steroidogenesis and/or follicular development. PMID- 10700087 TI - Localisation of atrial natriuretic factor (ANF) in rat testis after Leydig cell destruction: evidence for a potential role in regulating gonadal function. AB - OBJECTIVE: Since the atrial natriuretic factor (ANF) is synthesized in various peripheral tissues, where it acts in an autocrine or paracrine fashion, the aim was to gain new insight into ANF expression and function in rat testis after Leydig cell destruction (LCD). METHODS: Leydig cell destruction was performed by the treatment with ethane dimethane sulphonate (EDS). RESULTS: ANF was expressed after Leydig cell destruction and total elimination (24 h and 7 days after EDS treatment) and also after Leydig cell regeneration (21 and 45 days after EDS treatment). ANF staining in the interstitial compartment was observed in apoptotic Leydig cells 24 h after treatment. In seminiferous epithelium ANF labeling was detected in Sertoli and germ cell cytoplasm with a more prominent labeling in spermatids. The degenerating germ cells were totally labeled. CONCLUSIONS: The demonstration of ANF staining in seminiferous epithelium after Leydig cells elimination and androgen deprivation suggests that Leydig cells are not the sole source of ANF in rat testis and that the seminiferous epithelium may be a new site in which ANF may be synthesized. The result indicates that ANF plays a role in regulating gonadal function. PMID- 10700088 TI - Swapping science for consensus in Montreal. PMID- 10700089 TI - Synchronizing expectations. PMID- 10700090 TI - Sex, lies, and herbicides. PMID- 10700091 TI - GM crops and equivocal environmental benefits. PMID- 10700092 TI - Epigenomics. PMID- 10700094 TI - IBM joins SNP consortium PMID- 10700093 TI - Enzymes by post--restriction enzyme stability. PMID- 10700095 TI - British biotech: still waiting PMID- 10700097 TI - GMO roundup PMID- 10700096 TI - Athersys all the RAGE PMID- 10700099 TI - GMO fudge at BIA bash PMID- 10700098 TI - USDA names biotech panel PMID- 10700102 TI - Japan okays stem cells PMID- 10700101 TI - Functional map completed PMID- 10700100 TI - Support for agbiotech PMID- 10700103 TI - Boost for dutch biotech PMID- 10700104 TI - Cartoon PMID- 10700105 TI - Human genetics research in iceland to diversify PMID- 10700106 TI - GM health food PMID- 10700108 TI - Hot plants PMID- 10700109 TI - Spinal cord rejuvenation PMID- 10700107 TI - Research collaborations PMID- 10700111 TI - Solid-state DNA computer PMID- 10700110 TI - Illuminating cancer models PMID- 10700112 TI - Drug delivery coalesces PMID- 10700113 TI - Tracking phosphorylation in live cells PMID- 10700114 TI - Enantiomer selectivity inverted PMID- 10700116 TI - Chemical genetics and tissue engineering PMID- 10700115 TI - Cryopreservation by vitrification PMID- 10700118 TI - Transplastomics produces somatotropin PMID- 10700117 TI - Delivering PNAs to the nucleus PMID- 10700119 TI - Metabolic engineering PMID- 10700120 TI - Hemophilia bypassed PMID- 10700121 TI - Technical reports PMID- 10700123 TI - Review PMID- 10700122 TI - Testing Bt refuge strategies PMID- 10700124 TI - A view of the inside PMID- 10700125 TI - Biosafety rules get thumbs up. PMID- 10700126 TI - EU GMO applications continue to rot. PMID- 10700127 TI - Inquiry into gene therapy widens. PMID- 10700128 TI - Biotechnology venture capital booms in Japan. PMID- 10700129 TI - Geron issued UK Dolly patents. PMID- 10700130 TI - Blood products part of FDA xenotransplant plan. PMID- 10700131 TI - Overview of FY 2001 federal budget request. PMID- 10700132 TI - Combinatorial [correction of Combinational] chemistry gives cell biology some muscle. PMID- 10700133 TI - Novel fluorescent approaches for studying cell signaling in single cells. PMID- 10700134 TI - A chamber of hope for hemophilia. PMID- 10700135 TI - Testing Bt refuge strategies in the field. PMID- 10700136 TI - From genome to cellular phenotype--a role for metabolic flux analysis? PMID- 10700137 TI - Biotechnology development a stroll in the park? PMID- 10700138 TI - Ireland embraces biotechnology. PMID- 10700139 TI - Portugal opens its borders to biotechnology. PMID- 10700140 TI - Genomics forges ahead in East Asia. PMID- 10700141 TI - Biotechnology in the Bay. PMID- 10700142 TI - Structural genomics and its importance for gene function analysis. AB - Structural genomics projects aim to solve the experimental structures of all possible protein folds. Such projects entail a conceptual shift from traditional structural biology in which structural information is obtained on known proteins to one in which the structure of a protein is determined first and the function assigned only later. Whereas the goal of converting protein structure into function can be accomplished by traditional sequence motif-based approaches, recent studies have shown that assignment of a protein's biochemical function can also be achieved by scanning its structure for a match to the geometry and chemical identity of a known active site. Importantly, this approach can use low resolution structures provided by contemporary structure prediction methods. When applied to genomes, structural information (either experimental or predicted) is likely to play an important role in high-throughput function assignment. PMID- 10700143 TI - In vivo bypass of hemophilia A coagulation defect by factor XIIa implant. AB - Hemophilia A and B coagulation defects, which are caused by deficiencies of Factor VIII and Factor IX, respectively, can be bypassed by administration of recombinant Factor VIIa. However, the short half-life of recombinant Factor VIIa in vivo negates its routine clinical use. We report here an in vivo method for the continuous generation of Factor VIIa. The method depends on the implantation of a porous chamber that contains Factor Xa or XIIa, and continuously generates Factor VIIa bypass activity from the subject's own Factor VII, which enters the chamber by diffusion. Once inside, the Factor VII is cleaved to Factor VIIa by the immobilized Factor Xa or XIIa. The newly created Factor VIIa diffuses out of the chamber and back into the circulation, where it can bypass the deficient Factors VIII or IX, and enable coagulation to occur. In vitro, this method generates sufficient Factor VIIa to substantially correct Factor VIII-deficient plasma when assessed by the classical aPTT coagulation assay. In vivo, a Factor XIIa peritoneal implant generates bypass activity for up to one month when tested in rhesus monkeys. Implantation of such a chamber in a patient with hemophilia A or B could eventually provide a viable alternative to replacement therapies using exogenous coagulation factors. PMID- 10700144 TI - Vitreous cryopreservation maintains the function of vascular grafts. AB - Avoidance of ice formation during cooling can be achieved by vitrification, which is defined as solidification in an amorphous glassy state that obviates ice nucleation and growth. We show that a vitrification approach to storing vascular tissue results in markedly improved tissue function compared with a standard method involving freezing. The maximum contractions achieved in vitrified vessels were >80% of fresh matched controls with similar drug sensitivities, whereas frozen vessels exhibited maximal contractions below 30% of controls and concomitant decreases in drug sensitivity. In vivo studies of vitrified vessel segments in an autologous transplant model showed no adverse effects of vitreous cryopreservation compared with fresh tissue grafts. PMID- 10700145 TI - Effects in live cells of a c-myc anti-gene PNA linked to a nuclear localization signal. AB - Peptide nucleic acids (PNA) are synthetic homologs of nucleic acids in which the phosphate-sugar polynucleotide backbone is replaced by a flexible polyamide. In this study, a PNA construct was employed as an anti-gene agent in intact cells in culture. The cell lines studied were derived from Burkitt's lymphomas (BL) that presented a translocated and hyperexpressed c-myc oncogene. A 17-mer anti-myc PNA, complementary to a unique sequence located at the beginning of the second exon of the oncogene, and was covalently linked at its N terminus to a nuclear localization signal (NLS) (PNA-myc(wt)-NLS). When BL cells were exposed to PNA myc(wt)-NLS, the anti-gene construct was localized predominantly in the cell nuclei and a rapid consequent downregulation of c-myc expression occurred. Under these conditions, both completion of a productive cell cycle and apoptosis were inhibited. PMID- 10700146 TI - Myoseverin, a microtubule-binding molecule with novel cellular effects. AB - A new microtubule-binding molecule, myoseverin, was identified from a library of 2,6,9-trisubstituted purines in a morphological differentiation screen. Myoseverin induces the reversible fission of multinucleated myotubes into mononucleated fragments. Myotube fission promotes DNA synthesis and cell proliferation after removal of the compound and transfer of the cells to fresh growth medium. Transcriptional profiling and biochemical analysis indicate that myoseverin alone does not reverse the biochemical differentiation process. Instead, myoseverin affects the expression of a variety of growth factor, immunomodulatory, extracellular matrix-remodeling, and stress response genes, consistent with the activation of pathways involved in wound healing and tissue regeneration. PMID- 10700147 TI - Measurement of kinase activation in single mammalian cells. AB - We demonstrate a new method for the simultaneous measurement of the activation of key regulatory enzymes within single cells. To illustrate the capabilities of the technique, the activation of protein kinase C (PKC), protein kinase A (PKA), calcium-calmodulin activated kinase II (CamKII), and cdc2 protein kinase (cdc2K) was measured in response to both pharmacological or physiological stimuli. This assay strategy should be applicable to a broad range of intracellular enzymes, including phosphatases, proteases, nucleases, and other kinases. PMID- 10700148 TI - A fluorescent indicator for visualizing cAMP-induced phosphorylation in vivo. AB - We have developed a method for visualizing phosphorylation of proteins in living cells using a novel fluorescent indicator composed of two green fluorescent protein (GFP) variants joined by the kinase-inducible domain (KID) of the transcription factor cyclic adenosine monophosphate (cAMP)-responsive element binding protein (CREB). Phosphorylation of KID by the cAMP-dependent protein kinase A (PKA) decreased the fluorescence resonance energy transfer (FRET) among the flanking GFPs. By transfecting COS-7 cells with an expression vector encoding this indicator protein (termed ART for cAMP-responsive tracer), we were able to visualize activation dynamics of PKA in living cells. PMID- 10700149 TI - Inverting enantioselectivity by directed evolution of hydantoinase for improved production of L-methionine. AB - Using directed evolution, we have improved the hydantoinase process for production of L-methionine (L-met) in Escherichia coli. This was accomplished by inverting the enantioselectivity and increasing the total activity of a key enzyme in a whole-cell catalyst. The selectivity of all known hydantoinases for D 5-(2-methylthioethyl)hydantoin (D-MTEH) over the L-enantiomer leads to the accumulation of intermediates and reduced productivity for the L-amino acid. We used random mutagenesis, saturation mutagenesis, and screening to convert the D selective hydantoinase from Arthrobacter sp. DSM 9771 into an L-selective enzyme and increased its total activity fivefold. Whole E. coli cells expressing the evolved L-hydantoinase, an L-N-carbamoylase, and a hydantoin racemase produced 91 mM L-met from 100 mM D,L-MTEH in less than 2 h. The improved hydantoinase increased productivity fivefold for >90% conversion of the substrate. The accumulation of the unwanted intermediate D-carbamoyl-methionine was reduced fourfold compared to cells with the wild-type pathway. Highly D-selective hydantoinase mutants were also discovered. Enantioselective enzymes rapidly optimized by directed evolution and introduced into multienzyme pathways may lead to improved whole-cell catalysts for efficient production of chiral compounds. PMID- 10700150 TI - In vivo visualization of gene expression using magnetic resonance imaging. AB - High-resolution in vivo imaging of gene expression is not possible in opaque animals by existing techniques. Here we present a new approach for obtaining such images by magnetic resonance imaging (MRI) using an MRI contrast agent that can indicate reporter gene expression in living animals. We have prepared MRI contrast agents in which the access of water to the first coordination sphere of a chelated paramagnetic ion is blocked with a substrate that can be removed by enzymatic cleavage. Following cleavage, the paramagnetic ion can interact directly with water protons to increase the MR signal. Here, we report an agent where galactopyranose is the blocking group. This group renders the MRI contrast agent sensitive to expression of the commonly used marker gene, beta galactosidase. To cellular resolution, regions of higher intensity in the MR image correlate with regions expressing marker enzyme. These results offer the promise of in vivo mapping of gene expression in transgenic animals and validate a general approach for constructing a family of MRI contrast agents that respond to biological activity. PMID- 10700151 TI - A general definition of metabolic pathways useful for systematic organization and analysis of complex metabolic networks. AB - A set of linear pathways often does not capture the full range of behaviors of a metabolic network. The concept of 'elementary flux modes' provides a mathematical tool to define and comprehensively describe all metabolic routes that are both stoichiometrically and thermodynamically feasible for a group of enzymes. We have used this concept to analyze the interplay between the pentose phosphate pathway (PPP) and glycolysis. The set of elementary modes for this system involves conventional glycolysis, a futile cycle, all the modes of PPP function described in biochemistry textbooks, and additional modes that are a priori equally entitled to pathway status. Applications include maximizing product yield in amino acid and antibiotic synthesis, reconstruction and consistency checks of metabolism from genome data, analysis of enzyme deficiencies, and drug target identification in metabolic networks. PMID- 10700152 TI - High-yield production of a human therapeutic protein in tobacco chloroplasts. AB - Transgenic plants have become attractive systems for production of human therapeutic proteins because of the reduced risk of mammalian viral contaminants, the ability to do large scale-up at low cost, and the low maintenance requirements. Here we report a feasibility study for production of a human therapeutic protein through transplastomic transformation technology, which has the additional advantage of increased biological containment by apparent elimination of the transmission of transgenes through pollen. We show that chloroplasts can express a secretory protein, human somatotropin, in a soluble, biologically active, disulfide-bonded form. High concentrations of recombinant protein accumulation are observed (>7% total soluble protein), more than 300-fold higher than a similar gene expressed using a nuclear transgenic approach. The plastid-expressed somatotropin is nearly devoid of complex post-translational modifications, effectively increasing the amount of usable recombinant protein. We also describe approaches to obtain a somatotropin with a non-methionine N terminus, similar to the native human protein. The results indicate that chloroplasts are a highly efficient vehicle for the potential production of pharmaceutical proteins in plants. PMID- 10700153 TI - Field tests on managing resistance to Bt-engineered plants. AB - Several important crops have been engineered to express toxins of Bacillus thuringiensis (Bt) for insect control. In 1999, US farmers planted nearly 8 million hectares (nearly 20 million acres) of transgenic Bt crops approved by the EPA. Bt-transgenic plants can greatly reduce the use of broader spectrum insecticides, but insect resistance may hinder this technology. Present resistance management strategies rely on a "refuge" composed of non-Bt plants to conserve susceptible alleles. We have used Bt-transgenic broccoli plants and the diamondback moth as a model system to examine resistance management strategies. The higher number of larvae on refuge plants in our field tests indicate that a "separate refuge" will be more effective at conserving susceptible larvae than a "mixed refuge" and would thereby reduce the number of homozygous resistant (RR) offspring. Our field tests also examined the strategy of spraying the refuge to prevent economic loss to the crop while maintaining susceptible alleles in the population. Results indicate that great care must be taken to ensure that refuges, particularly those sprayed with efficacious insecticides, produce adequate numbers of susceptible alleles. Each insect/Bt crop system may have unique management requirements because of the biology of the insect, but our studies validate the need for a refuge. As we learn more about how to refine our present resistance management strategies, it is important to also develop the next generation of technology and implementation strategies. PMID- 10700154 TI - Artificial chromosomes for antibiotic-producing actinomycetes. AB - Bacteria belonging to the order Actinomycetales produce most microbial metabolites thus far described, several of which have found applications in medicine and agriculture. However, most strains were discovered by their ability to produce a given molecule and are, therefore, poorly characterized physiologically and genetically. Thus, methodologies for genetic manipulation of actinomycetes are not available and efficient tools have been developed for just a few strains. This constitutes a serious limitation to applying molecular genetics approaches to strain development and structural manipulation of microbial metabolites. To overcome this hurdle, we have developed bacterial artificial chromosomes (BAC) that can be shuttled among Escherichia coli, where they replicate autonomously, and a suitable Streptomyces host, where they integrate site-specifically into the chromosome. The existence of gene clusters and of genetically amenable host strains, such as Streptomyces coelicolor or Streptomyces lividans, makes this a sensible approach. We report here that 100 kb segments of actinomycete DNA can be cloned into these vectors and introduced into genetically accessible S. lividans, where they are stably maintained in integrated form in its chromosome. PMID- 10700155 TI - Custom fluorescent-nucleotide synthesis as an alternative method for nucleic acid labeling. AB - The variety of potentially useful dyes or haptenes available for fluorescent nucleic acid hybridization assays is far greater than what can be obtained from commercial sources. Since this diversity could be useful in many laboratory applications, we have developed a simple and inexpensive procedure for preparing nonpurified labeled nucleotides, for use in common nucleic acid labeling reactions, such as PCR and nick translation. The modified nucleotides were synthesized by coupling allylamine-dUTP to the succinimidyl-ester derivatives of the fluorescent dyes or haptenes such as biotin or digoxigenin, which require fluorescently labeled proteins for detection. This method allows custom preparation of most common fluorescent nucleotides and rapid testing of new ones, while reducing the cost of procedures such as multiplex fluorescent in situ hybridization (M-FISH) by 100-200 fold. PMID- 10700156 TI - Analyzing the USPTO's revised utility guidelines. United States Patent and Trademark Office. PMID- 10700157 TI - Recent patents in cloning PMID- 10700158 TI - Blood, genes, and profits PMID- 10700159 TI - People PMID- 10700160 TI - New products PMID- 10700161 TI - Trials and tribulations. PMID- 10700162 TI - Ellis-van Creveld syndrome and the Amish. PMID- 10700163 TI - Making the most of microarray data. PMID- 10700164 TI - Axin and hepatocellular carcinomas. PMID- 10700165 TI - Cancer cells, chemotherapy and gene clusters. PMID- 10700166 TI - TOUCHINGbase. Genomes galore. PMID- 10700167 TI - Are (CTG)n expansions at the SCA8 locus rare polymorphisms? PMID- 10700168 TI - Large, expanded repeats in SCA8 are not confined to patients with cerebellar ataxia. PMID- 10700169 TI - Reply- PMID- 10700171 TI - Capital errors PMID- 10700170 TI - Basal cell carcinomas in mice overexpressing Gli2 in skin. PMID- 10700172 TI - Fate of the lineage PMID- 10700173 TI - Analysing complex genetic traits with chromosome substitution strains. AB - Many valuable animal models of human disease are known and new models are continually being generated in existing inbred strains,. Some disease models are simple mendelian traits, but most have a polygenic basis. The current approach to identifying quantitative trait loci (QTLs) that underlie such traits is to localize them in crosses, construct congenic strains carrying individual QTLs, and finally map and clone the genes. This process is time-consuming and expensive, requiring the genotyping of large crosses and many generations of breeding. Here we describe a different approach in which a panel of chromosome substitution strains (CSSs) is used for QTL mapping. Each of these strains has a single chromosome from the donor strain substituting for the corresponding chromosome in the host strain. We discuss the construction, applications and advantages of CSSs compared with conventional crosses for detecting and analysing QTLs, including those that have weak phenotypic effects. PMID- 10700174 TI - Systematic variation in gene expression patterns in human cancer cell lines. AB - We used cDNA microarrays to explore the variation in expression of approximately 8,000 unique genes among the 60 cell lines used in the National Cancer Institute's screen for anti-cancer drugs. Classification of the cell lines based solely on the observed patterns of gene expression revealed a correspondence to the ostensible origins of the tumours from which the cell lines were derived. The consistent relationship between the gene expression patterns and the tissue of origin allowed us to recognize outliers whose previous classification appeared incorrect. Specific features of the gene expression patterns appeared to be related to physiological properties of the cell lines, such as their doubling time in culture, drug metabolism or the interferon response. Comparison of gene expression patterns in the cell lines to those observed in normal breast tissue or in breast tumour specimens revealed features of the expression patterns in the tumours that had recognizable counterparts in specific cell lines, reflecting the tumour, stromal and inflammatory components of the tumour tissue. These results provided a novel molecular characterization of this important group of human cell lines and their relationships to tumours in vivo. PMID- 10700175 TI - A gene expression database for the molecular pharmacology of cancer. AB - We used cDNA microarrays to assess gene expression profiles in 60 human cancer cell lines used in a drug discovery screen by the National Cancer Institute. Using these data, we linked bioinformatics and chemoinformatics by correlating gene expression and drug activity patterns in the NCI60 lines. Clustering the cell lines on the basis of gene expression yielded relationships very different from those obtained by clustering the cell lines on the basis of their response to drugs. Gene-drug relationships for the clinical agents 5-fluorouracil and L asparaginase exemplify how variations in the transcript levels of particular genes relate to mechanisms of drug sensitivity and resistance. This is the first study to integrate large databases on gene expression and molecular pharmacology. PMID- 10700176 TI - AXIN1 mutations in hepatocellular carcinomas, and growth suppression in cancer cells by virus-mediated transfer of AXIN1. AB - The Wnt signaling pathway is essential for development and organogenesis. Wnt signaling stabilizes beta-catenin, which accumulates in the cytoplasm, binds to 1 cell factor (TCF; also known as lymphocyte enhancer-binding factor, LEF) and then upregulates downstream genes. Mutations in CTNNB1 (encoding beta-catenin) or APC (adenomatous polyposis coli) have been reported in human neoplasms including colon cancers and hepatocellular carcinomas (HCCs). Because HCC5 tend to show accumulation of beta-catenin more often than mutations in CTNNB1, we looked for mutations in AXIN1, encoding a key factor for Wnt signaling, in 6 HCC cell lines and 100 primary HCC5. Among the 4 cell lines and 87 HCC5 in which we did not detect CTNNB1 mutations, we identified AXIN1 mutations in 3 cell lines and 6 mutations in 5 of the primary HCCs. In cell lines containing mutations in either gene, we observed increased DNA binding of TCF associated with beta-catenin in nuclei. Adenovirus mediated gene transfer of wild-type AXINI induced apoptosis in hepatocellular and colorectal cancer cells that had accumulated beta-catenin as a consequence of either APC, CTNNB1 or AXIN1 mutation, suggesting that axin may be an effective therapeutic molecule for suppressing growth of hepatocellular and colorectal cancers. PMID- 10700177 TI - Mutations in ACTN4, encoding alpha-actinin-4, cause familial focal segmental glomerulosclerosis. AB - Focal and segmental glomerulosclerosis (FSGS) is a common, non-specific renal lesion. Although it is often secondary to other disorders, including HIV infection, obesity, hypertension and diabetes, FSGS also appears as an isolated, idiopathic condition. FSGS is characterized by increased urinary protein excretion and decreasing kidney function. Often, renal insufficiency in affected patients progresses to end-stage renal failure, a highly morbid state requiring either dialysis therapy or kidney transplantation. Here we present evidence implicating mutations in the gene encoding alpha-actinin-4 (ACTN4; ref. 2), an actin-filament crosslinking protein, as the cause of disease in three families with an autosomal dominant form of FSGS. In vitro, mutant alpha-actinin-4 binds filamentous actin (F-actin) more strongly than does wild-type alpha-actinin-4. Regulation of the actin cytoskeleton of glomerular podocytes may be altered in this group of patients. Our results have implications for understanding the role of the cytoskeleton in the pathophysiology of kidney disease and may lead to a better understanding of the genetic basis of susceptibility to kidney damage. PMID- 10700178 TI - Evidence for gene transfer and expression of factor IX in haemophilia B patients treated with an AAV vector. AB - Pre-clinical studies in mice and haemophilic dogs have shown that introduction of an adeno-associated viral (AAV) vector encoding blood coagulation factor IX (FIX) into skeletal muscle results in sustained expression of F.IX at levels sufficient to correct the haemophilic phenotype. On the basis of these data and additional pre-clinical studies demonstrating an absence of vector-related toxicity, we initiated a clinical study of intramuscular injection of an AAV vector expressing human F.IX in adults with severe haemophilia B. The study has a dose-escalation design, and all patients have now been enrolled in the initial dose cohort (2 x 10(11) vg/kg). Assessment in the first three patients of safety and gene transfer and expression show no evidence of germline transmission of vector sequences or formation of inhibitory antibodies against F.IX. We found that the vector sequences are present in muscle by PCR and Southern-blot analyses of muscle biopsies and we demonstrated expression of F.IX by immunohistochemistry. We observed modest changes in clinical endpoints including circulating levels of F.IX and frequency of FIX protein infusion. The evidence of gene expression at low doses of vector suggests that dose calculations based on animal data may have overestimated the amount of vector required to achieve therapeutic levels in humans, and that the approach offers the possibility of converting severe haemophilia B to a milder form of the disease. PMID- 10700179 TI - Regulation of left-right patterning in mice by growth/differentiation factor-1. AB - The transforming growth factor-beta (TGF-beta) superfamily encompasses a large group of structurally related polypeptides that are capable of regulating cell growth and differentiation in a wide range of embryonic and adult tissues. Growth/differentiation factor-1 (Gdf-1, encoded by Gdf1) is a TGF-beta family member of unknown function that was originally isolated from an early mouse embryo cDNA library and is expressed specifically in the nervous systemin late stage embryos and adult mice. Here we show that at early stages of mouse development, Gdfl is expressed initially throughout the embryo proper and then most prominently in the primitive node, ventral neural tube, and intermediate and lateral plate mesoderm. To examine its biological function, we generated a mouse line carrying a targeted mutation in Gdf1. Gdf1-/- mice exhibited a spectrum of defects related to left-right axis formation, including visceral situs inversus, right pulmonary isomerism and a range of cardiac anomalies. In most Gdf1-/- embryos, the expression of Ebaf (formerly lefty-1) in the left side of the floor plate and Leftb (formerly lefty-2), nodal and Pitx2 in the left lateral plate mesoderm was absent, suggesting that Gdf1 acts upstream of these genes either directly or indirectly to activate their expression. Our findings suggest that Gdf1 acts early in the pathway of gene activation that leads to the establishment of left-right asymmetry. PMID- 10700180 TI - Familial dyserythropoietic anaemia and thrombocytopenia due to an inherited mutation in GATA1. AB - Haematopoietic development is regulated by nuclear protein complexes that coordinate lineage-specific patterns of gene expression. Targeted mutagenesis in embryonic stem cells and mice has revealed roles for the X-linked gene Gata1 in erythrocyte and megakaryocyte differentiation. GATA-1 is the founding member of a family of DNA-binding proteins that recognize the motif WGATAR through a conserved multifunctional domain consisting of two C4-type zinc fingers. Here we describe a family with X-linked dyserythropoietic anaemia and thrombocytopenia due to a substitution of methionine for valine at amino acid 205 of GATA-1. This highly conserved valine is necessary for interaction of the amino-terminal zinc finger of GATA-1 with its essential cofactor, FOG-1 (for friend of GATA-1; refs 9 12). We show that the V205M mutation abrogates the interaction between Gata-1 and Fog-1, inhibiting the ability of Gata-1 to rescue erythroid differentiation in an erythroid cell line deficient for Gata-1 (G1E). Our findings underscore the importance of FOG-1:Gata-1 associations in both megakaryocyte and erythroid development, and suggest that other X-linked anaemias or thrombocytopenias may be caused by defects in GATA1. PMID- 10700181 TI - Ror2, encoding a receptor-like tyrosine kinase, is required for cartilage and growth plate development. AB - Receptor tyrosine kinases often have critical roles in particular cell lineages by initiating signalling cascades in those lineages. Examples include the neural specific TRK receptors, the VEGF and angiopoietin endothelial-specific receptors, and the muscle-specific MUSK receptor. Many lineage-restricted receptor tyrosine kinases were initially identified as 'orphans' homologous to known receptors, and only subsequently used to identify their unknown growth factors. Some receptor tyrosine-kinase-like orphans still lack identified ligands as well as biological roles. Here we characterize one such orphan, encoded by Ror2 (ref. 12). We report that disruption of mouse Ror2 leads to profound skeletal abnormalities, with essentially all endochondrally derived bones foreshortened or misshapen, albeit to differing degrees. Further, we find that Ror2 is selectively expressed in the chondrocytes of all developing cartilage anlagen, where it essential during initial growth and patterning, as well as subsequently in the proliferating chondrocytes of mature growth plates, where it is required for normal expansion. Thus, Ror2 encodes a receptor-like tyrosine kinase that is selectively expressed in, and particularly important for, the chondrocyte lineage. PMID- 10700182 TI - Dominant mutations in ROR2, encoding an orphan receptor tyrosine kinase, cause brachydactyly type B. AB - Inherited limb malformations provide a valuable resource for the identification of genes involved in limb development. Brachydactyly type B (BDB), an autosomal dominant disorder, is the most severe of the brachydactylies and characterized by terminal deficiency of the fingers and toes. In the typical form of BDB, the thumbs and big toes are spared, sometimes with broadening or partial duplication. The BDB1 locus was previously mapped to chromosome 9q22 within an interval of 7.5 cM (refs 9,10). Here we describe mutations in ROR2, which encodes the orphan receptor tyrosine kinase ROR2 (ref. 11), in three unrelated families with BDB1. We identified distinct heterozygous mutations (2 nonsense, 1 frameshift) within a 7-amino-acid segment of the 943-amino-acid protein, all of which predict truncation of the intracellular portion of the protein immediately after the tyrosine kinase domain. The localized nature of these mutations suggests that they confer a specific gain of function. We obtained further evidence for this by demonstrating that two patients heterozygous for 9q22 deletions including ROR2 do not exhibit BDB. Expression of the mouse mouse orthologue, Ror2, early in limb development indicates that BDB arises as a primary defect of skeletal patterning. PMID- 10700183 TI - The gamete fusion process is defective in eggs of Cd9-deficient mice. AB - The cell-surface molecule Cd9, a member of the transmembrane-4 superfamily, interacts with the integrin family and other membrane proteins. and is postulated to participate in cell migration and adhesion. Expression of Cd9 enhances membrane fusion between muscle cells and promotes viral infection in some cells. Fertilization also involves membrane fusion, between gametes. In mammals, the sperm binds to microvilli on the egg surface, and sperm-egg membrane fusion first occurs around the equatorial region of the sperm head12. The fused membrane is then disrupted, and the sperm nucleus as well as the cytoplasm is incorporated into the egg. Cd9 is expressed on the plasma membrane of the mouse egg, and an anti-Cd9 monoclonal antibody inhibits sperm-egg surface interactions. We generated Cd9 mice and found that homozygous mutant females were infertile. Sperm egg binding was normal, but sperm-egg fusion was almost entirely inhibited in eggs from Cd9 females. Intracellular Ca2 oscillations, which signal fertilization, were absent in almost all mutant eggs; in rare cases, a response occurred after a long time period. In normal animals, Cd9 molecules were expressed on the egg microvilli and became densely concentrated at the sperm attachment site. Thus, our results show that Cd9 is important in the gamete fusion process at fertilization. PMID- 10700184 TI - Mutations in a new gene in Ellis-van Creveld syndrome and Weyers acrodental dysostosis. AB - Ellis-van Creveld syndrome (EvC, MIM 225500) is an autosomal recessive skeletal dysplasia characterized by short limbs, short ribs, postaxial polydactyly and dysplastic nails and teeth. Congenital cardiac defects, most commonly a defect of primary atrial septation producing a common atrium, occur in 60% of affected individuals. The disease was mapped to chromosome 4p16 in nine Amish subpedigrees and single pedigrees from Mexico, Ecuador and Brazil. Weyers acrodental dysostosis (MIM 193530), an autosomal dominant disorder with a similar but milder phenotype, has been mapped in a single pedigree to an area including the EvC critical region. We have identified a new gene (EVC), encoding a 992-amino-acid protein, that is mutated in individuals with EvC. We identified a splice-donor change in an Amish pedigree and six truncating mutations and a single amino acid deletion in seven pedigrees. The heterozygous carriers of these mutations did not manifest features of EvC. We found two heterozygous missense mutations associated with a phenotype, one in a man with Weyers acrodental dysostosis and another in a father and his daughter, who both have the heart defect characteristic of EvC and polydactyly, but not short stature. We suggest that EvC and Weyers acrodental dysostosis are allelic conditions. PMID- 10700185 TI - Rnx deficiency results in congenital central hypoventilation. AB - The genes Tlx1 (Hox11), Enx (Hox11L2, Tlx-2) and Rnx (Hox11L2, Tlx-3) constitute a family of orphan homeobox genes. In situ hybridization has revealed considerable overlap in their expression within the nervous system, but Rnx is singularly expressed in the developing dorsal and ventral region of the medulla oblongata. Tlx1-deficient and Enx-deficient mice display phenotypes in tissues where the mutated gene is singularly expressed, resulting in asplenogenesis and hyperganglionic megacolon, respectively. To determine the developmental role of Rnx, we disrupted the locus in mouse embryonic stem (ES) cells. Rnx deficient mice developed to term, but all died within 24 hours after birth from a central respiratory failure. The electromyographic activity of intercostal muscles coupled with the C4 ventral root activity assessed in a medulla-spinal cord preparation revealed a high respiratory rate with short inspiratory duration and frequent apnea. Furthermore, a coordinate pattern existed between the abnormal activity of inspiratory neurons in the ventrolateral medulla and C4 motorneuron output, indicating a central respiratory defect in Rnx mice. Thus, Rnx is critical for the development of the ventral medullary respiratory centre and its deficiency results in a syndrome resembling congenital central hypoventilation. PMID- 10700186 TI - The gene MAPK8IP1, encoding islet-brain-1, is a candidate for type 2 diabetes. AB - Type 2 diabetes is a polygenic and genetically heterogeneous disease . The age of onset of the disease is usually late and environmental factors may be required to induce the complete diabetic phenotype. Susceptibility genes for diabetes have not yet been identified. Islet-brain-1 (IB1, encoded by MAPK8IP1), a novel DNA binding transactivator of the glucose transporter GLUT2 (encoded by SLC2A2), is the homologue of the c-Jun amino-terminal kinase-interacting protein-1 (JIP-1; refs 2-5). We evaluated the role of IBi in beta-cells by expression of a MAPK8IP1 antisense RNA in a stable insulinoma beta-cell line. A 38% decrease in IB1 protein content resulted in a 49% and a 41% reduction in SLC2A2 and INS (encoding insulin) mRNA expression, respectively. In addition, we detected MAPK8IP1 transcripts and IBi protein in human pancreatic islets. These data establish MAPK8IP1 as a candidate gene for human diabetes. Sibpair analyses performed on i49 multiplex French families with type 2 diabetes excluded MAPK8IP1 as a major diabetogenic locus. We did, however, identify in one family a missense mutation located in the coding region of MAPK8IP1 (559N) that segregated with diabetes. In vitro, this mutation was associated with an inability of IB1 to prevent apoptosis induced by MAPK/ERK kinase kinase 1 (MEKK1) and a reduced ability to counteract the inhibitory action of the activated c-JUN amino-terminal kinase (JNK) pathway on INS transcriptional activity. Identification of this novel non-maturity onset diabetes of the young (MODY) form of diabetes demonstrates that IB1 is a key regulator of 3-cell function. PMID- 10700187 TI - Mice mutant for Egfr and Shp2 have defective cardiac semilunar valvulogenesis. AB - Atrioventricular and semilunar valve abnormalities are common birth defects, but how cardiac valvulogenesis is directed remains largely unknown. During studies of genetic interaction between Egfr, encoding the epidermal growth factor receptor, and Ptpn11, encoding the protein-tyrosine-phosphatase Shp2, we discovered that Egfr is required for semilunar, but not atrioventricular, valve development. Although unnoticed in earlier studies, mice homozygous for the hypomorphic Egfr allele waved-2 (Egfrwa2/wa2) exhibit semilunar valve enlargement resulting from over-abundant mesenchymal cells. Egfr-/- mice (CD1 background) have similar defects. The penetrance and severity of the defects in Egfrwa2/wa2 mice are enhanced by heterozygosity for a targeted mutation of exon 2 of Ptpn11 (ref. 3). Compound (Egfrwa2/wa2:Ptpn11+/-) mutant mice also show premature lethality. Electrocardiography, echocardiography and haemodynamic analyses showed that affected mice develop aortic stenosis and regurgitation. Our results identify the Egfr and Shp2 as components of a growth-factor signalling pathway required specifically for semilunar valvulogenesis, support the hypothesis that Shp2 is required for Egfr signalling in vivo, and provide an animal model for aortic valve disease. PMID- 10700188 TI - Mutations truncating the EP300 acetylase in human cancers. AB - The EP300 protein is a histone acetyltransferase that regulates transcription via chromatin remodelling and is important in the processes of cell proliferation and differentiation. EP300 acetylation of TP53 in response to DNA damage regulates its DNA-binding and transcription functions. A role for EP300 in cancer has been implied by the fact that it is targeted by viral oncoproteins, it is fused to MLL in Leukaemia and two missense sequence alterations in EP300 were identified in epithelial malignancies. Nevertheless, direct demonstration of the role of EP300 in tumorigenesis by inactivating mutations in human cancers has been lacking. Here we describe EP300 mutations, which predict a truncated protein, in 6(3%) of 193 epithelial cancers analysed. Of these six mutations, two were in primary tumours (a colorectal cancer and a breast cancer) and four were in cancer cell lines (colorectal, breast and pancreatic). In addition, we identified a somatic in-frame insertion in a primary breast cancer and missense alterations in a primary colorectal cancer and two cell lines (breast and pancreatic). Inactivation of the second allele was demonstrated in five of six cases with truncating mutations and in two other cases. Our data show that EP300 is mutated in epithelial cancers and provide the first evidence that it behaves as a classical tumour-suppressor gene. PMID- 10700189 TI - Mice deficient in Abl are osteoporotic and have defects in osteoblast maturation. AB - The c-Abl protein is a non-receptor tyrosine kinase involved in many aspects of mammalian development. c-Abl kinase is widely expressed, but high levels are found in hyaline cartilage in the adult, bone tissue in newborn mice, and osteoblasts and associated neovasculature at sites of endochondrial ossification in the fetus. Mice homozygous for mutations in the gene encoding c-Abl (AIM) display increased perinatal mortality, reduced fertility, foreshortened crania and defects in the maturation of B cells in bone marrow. Here we demonstrate that Abl-/- mice are also osteoporotic. The long bones of mutant mice contain thinner cortical bone and reduced trabecular bone volume. The osteoporotic phenotype is not due to accelerated bone turnover--both the number and activity of osteoclasts are similar to those of control littermates--but rather to dysfunctional osteoblasts. In addition, the rate of mineral apposition in the mutant animals is reduced. Osteoblasts from both stromal and calvarial explants showed delayed maturation in vitro as measured by expression of alkaline phosphatase (ALP), induction of mRNA encoding osteocalcin and mineral deposition. PMID- 10700190 TI - Mekk3 is essential for early embryonic cardiovascular development. AB - The early development of blood vessels consists of two phases, vasculogenesis and angiogenesis, which involve distinct and also overlapping molecular regulators, but the intracellular signal transduction pathways involved in these processes have not been well defined. We disrupted Map3k3 (also known as Mekk3), which encodes Mekk3, a member of the Mekk/Ste11 family, in mice. Map3k3-/- embryos died at approximately embryonic day (E) 11, displaying disruption of blood vessel development and the structural integrity of the yolk sac. Angiogenesis was blocked at approximately E9.5 in mutant embryos. Map3k3 disruption did not alter the expression of the genes encoding Vegf-1, angiopoietin or their receptors. The development of embryonic, but not maternal, blood vessels in the placentas of Map3k3-/- embryos was impaired, revealing an intrinsic defect in Map3k3-/- endothelial cells. Moreover, Mekk3 activated myocyte-specific enhancer factor 2C (Mef2c), a transcription factor crucial for early embryonic cardiovascular development through the p38 mitogen-activated protein kinase (Mapk) cascade. We conclude that Mekk3 is necessary for blood vessel development and may be a possible target for drugs that control angiogenesis. PMID- 10700191 TI - Genotype-based screen for ENU-induced mutations in mouse embryonic stem cells. AB - The ability to generate mutations is a prerequisite to functional genetic analysis. Despite a long history of using mice as a model system for genetic analysis, the scientific community has not generated a comprehensive collection of multiple alleles for most mouse genes. The chemical mutagen of choice for mouse has been N-ethyl-N-nitrosourea (ENU), an alkylating agent that mainly causes base substitutions in DNA, and therefore allows for recovery of complete and partial loss-, as well as gain-, of-function alleles . Specific locus tests designed to detect recessive mutations showed that ENU is the most efficient mutagen in mouse with an approximate mutation rate of 1 in 1,000 gametes. In fact, several genome-wide and region-specific screens based on phenotypes have been carried out. The anticipation of the completion of the human and mouse genome projects, however, now emphasizes genotype-driven genetics--from sequence to mutants. To take advantage of the mutagenicity of ENU and its ability to create allelic series of mutations, we have developed a complementary approach to generating mutations using mouse embryonic stem (ES) cells. We show that a high mutation frequency can be achieved and that modulating DNA-repair activities can enhance this frequency. The treated cells retain germline competency, thereby rendering this approach applicable for efficient generation of an allelic series of mutations pivotal to a fine-tuned dissection of biological pathways. PMID- 10700193 TI - Spring back-or forward? AB - While it's sometimes hard to remember whether the clock goes forwards or backwards, you'll save time in PCR quantitation, telomere length analysis and large-scale mRNA processing with the latest arrivals on the biomedical research scene. PMID- 10700192 TI - Mouse mutants from chemically mutagenized embryonic stem cells. AB - The drive to characterize functions of human genes on a global scale has stimulated interest in large-scale generation of mouse mutants. Conventional germ cell mutagenesis with N-ethyl-N-nitrosourea (ENU) is compromised by an inability to monitor mutation efficiency, strain and interlocus variation in mutation induction, and extensive husbandry requirements. To overcome these obstacles and develop new methods for generating mouse mutants, we devised protocols to generate germline chimaeric mice from embryonic stem (ES) cells heavily mutagenized with ethylmethanesulphonate (EMS). Germline chimaeras were derived from cultures that underwent a mutation rate of up to 1 in 1,200 at the Hprt locus (encoding hypoxanthine guanine phosphoribosyl transferase). The spectrum of mutations induced by EMS and the frameshift mutagen ICR191 was consistent with that observed in other mammalian cells. Chimaeras derived from ES cells treated with EMS transmitted mutations affecting several processes, including limb development, hair growth, hearing and gametogenesis. This technology affords several advantages over traditional mutagenesis, including the ability to conduct shortened breeding schemes and to screen for mutant phenotypes directly in ES cells or their differentiated derivatives. PMID- 10700194 TI - Stem cells-why wait? PMID- 10700195 TI - Origins of HTLV-1 in South America. PMID- 10700197 TI - Reply to Origins of HTLV-1 in South America. PMID- 10700196 TI - Origins of HTLV-1 in South America. PMID- 10700198 TI - Reply to Malaria vaccine research-setting the record straight. PMID- 10700199 TI - Malaria vaccine research--setting the record straight. PMID- 10700200 TI - Gene therapy investigations proliferate. PMID- 10700201 TI - Fate of 'Montagnier Center' to be decided. PMID- 10700202 TI - Kourilsky to reform the Pasteur Institute. PMID- 10700203 TI - University stem cells for sale. PMID- 10700204 TI - Distribution of mental health funding questioned. PMID- 10700205 TI - Another attempt at creating a global vaccine initiative. PMID- 10700206 TI - Spanish oncologists square-up. PMID- 10700207 TI - NIH announces plans for 2001 budget. PMID- 10700208 TI - Japan to permit stem cell research. PMID- 10700209 TI - Can GURP help indebted medical research centers? PMID- 10700210 TI - Alternative medicine chief sets research agenda. PMID- 10700211 TI - Canada expands networked research--up to a limit. PMID- 10700213 TI - Anthrax: The Investigation of a Deadly Outbreak. PMID- 10700212 TI - Malaria transmission-blocking vaccines--how can their development be supported? PMID- 10700214 TI - The Paradox of Sleep. PMID- 10700215 TI - Hormonal Chaos: The Scientific and Social Origins of the Environmental Endocrine Hypothesis. PMID- 10700216 TI - Neuronal progenitors-learning from the hippocampus. PMID- 10700217 TI - New islets from old. PMID- 10700218 TI - Smallpox, polio and now a cancer vaccine? PMID- 10700219 TI - Sane genetics for schizophrenia. PMID- 10700220 TI - Pot of gold for glioma therapy. PMID- 10700221 TI - Visualizing transcription. PMID- 10700222 TI - Blocking bacterial enterotoxins. PMID- 10700223 TI - 'New variant' Creutzfeldt-Jakob disease and bovine spongiform encephalopathy. PMID- 10700224 TI - Positive selection for autoimmunity. PMID- 10700225 TI - Both a 'magic bullet' and good aim are required to link public health interests and health care needs in HIV infection. PMID- 10700226 TI - Research News. PMID- 10700227 TI - A new biological agent for treatment of Shiga toxigenic Escherichia coli infections and dysentery in humans. AB - Gastrointestinal disease caused by Shiga toxin-producing bacteria (such as Escherichia coli O157:H7 and Shigella dysenteriae) is often complicated by life threatening toxin-induced systemic sequelae, including hemolytic-uremic syndrome. Such infections can now be diagnosed very early in the course of the disease, but at present no effective therapeutic intervention is possible. Here, we constructed a recombinant bacterium that displayed a Shiga toxin receptor mimic on its surface, and it adsorbed and neutralized Shiga toxins with very high efficiency. Moreover, oral administration of the recombinant bacterium completely protected mice from challenge with an otherwise 100%-fatal dose of Shiga toxigenic E. coli. Thus, the bacterium shows great promise as a 'probiotic' treatment for Shiga toxigenic E. coli infections and dysentery. PMID- 10700228 TI - In vitro neurogenesis by progenitor cells isolated from the adult human hippocampus. AB - Neurogenesis persists in the adult mammalian hippocampus. To identify and isolate neuronal progenitor cells of the adult human hippocampus, we transfected ventricular zone-free dissociates of surgically-excised dentate gyrus with DNA encoding humanized green fluorescent protein (hGFP), placed under the control of either the nestin enhancer (E/nestin) or the Talpha1 tubulin promoter (P/Talpha1), two regulatory regions that direct transcription in neural progenitor cells. The resultant P/Talpha1:hGFP+ and E/nestin:enhanced (E)GFP+ cells expressed betaIII-tubulin or microtubule-associated protein-2; many incorporated bromodeoxyuridine, indicating their genesis in vitro. Using fluorescence-activated cell sorting, the E/nestin:EGFP+ and P/Talpha1:hGFP+ cells were isolated to near purity, and matured antigenically and physiologically as neurons. Thus, the adult human hippocampus contains mitotically competent neuronal progenitors that can be selectively extracted. The isolation of these cells may provide a cellular substrate for re-populating the damaged or degenerated adult hippocampus. PMID- 10700229 TI - Reversal of insulin-dependent diabetes using islets generated in vitro from pancreatic stem cells. AB - Ductal structures of the adult pancreas contain stem cells that differentiate into islets of Langerhans. Here, we grew pancreatic ductal epithelial cells isolated from prediabetic adult non-obese diabetic mice in long-term cultures, where they were induced to produce functioning islets containing alpha, beta and delta cells. These in vitro-generated islets showed temporal changes in mRNA transcripts for islet cell-associated differentiation markers, responded in vitro to glucose challenge, and reversed insulin-dependent diabetes after being implanted into diabetic non-obese diabetic mice. The ability to control growth and differentiation of islet stem cells provides an abundant islet source for beta-cell reconstitution in type I diabetes. PMID- 10700231 TI - p27kip1 functions as an anergy factor inhibiting interleukin 2 transcription and clonal expansion of alloreactive human and mouse helper T lymphocytes. AB - Although recent in vitro studies have begun to decipher the molecular events that characterize the anergic state, their in vivo biologic relevance and potential clinical importance remain unclear. Here, using anergic human T-cell clones and tolerant alloreactive mouse T cells that do not induce graft-versus-host disease, we show that p27kip1 cyclin-dependent kinase inhibitor is an essential regulator responsible for the blockade of clonal expansion of anergic T cells in vitro and in vivo. Moreover, in anergic cells, p27kip1 associates with the c-Jun co activator JAB1, resulting in defective transactivation of AP-1 and interleukin 2 transcription. Therefore, pharmacological agents that upregulate the expression of or prevent the degradation of p27kip1 during antigen recognition should be part of new therapeutic strategies to induce antigen-specific T-cell unresponsiveness. PMID- 10700230 TI - Modulation of T-cell-mediated immunity in tumor and graft-versus-host disease models through the LIGHT co-stimulatory pathway. AB - LIGHT was recently described as a member of the tumor necrosis factor (TNF) 'superfamily'. We have isolated a mouse homolog of human LIGHT and investigated its immunoregulatory functions in vitro and in vivo. LIGHT has potent, CD28 independent co-stimulatory activity leading to T-cell growth and secretion of gamma interferon and granulocyte-macrophage colony-stimulating factor. Gene transfer of LIGHT induced an antigen-specific cytolytic T-cell response and therapeutic immunity against established mouse P815 tumor. In contrast, blockade of LIGHT by administration of soluble receptor or antibody led to decreased cell mediated immunity and ameliorated graft-versus-host disease. Our studies identify a previously unknown T-cell co-stimulatory pathway as a potential therapeutic target. PMID- 10700232 TI - Disruption of positive selection of thymocytes causes autoimmunity. AB - To differentiate into T cells, immature thymocytes must engage, through their antigen-specific T-cell receptor, peptides derived from self proteins presented by cortical epithelial cells in the thymus, a process called positive selection. Despite this requirement for self-recognition during development, mature T cells do not normally show autoreactivity. Mice injected in the thymus with procainamide-hydroxylamine, a metabolite of procainamide, develop autoimmune features resembling drug-induced lupus. Here, we show that when thymocytes undergo positive selection in the presence of procainamide-hydroxylamine, they fail to establish unresponsiveness to low affinity selecting self antigens, resulting in systemic autoimmunity. PMID- 10700233 TI - Suppression of tumor growth and metastasis in Mgat5-deficient mice. AB - Golgi beta1,6N-acetylglucosaminyltransferase V (MGAT5) is required in the biosynthesis of beta1,6GlcNAc-branched N-linked glycans attached to cell surface and secreted glycoproteins. Amounts of MGAT5 glycan products are commonly increased in malignancies, and correlate with disease progression. To study the functions of these N-glycans in development and disease, we generated mice deficient in Mgat5 by targeted gene mutation. These Mgat5-/- mice lacked Mgat5 products and appeared normal, but differed in their responses to certain extrinsic conditions. Mammary tumor growth and metastases induced by the polyomavirus middle T oncogene was considerably less in Mgat5-/- mice than in transgenic littermates expressing Mgat5. Furthermore, Mgat5 glycan products stimulated membrane ruffling and phosphatidylinositol 3 kinase-protein kinase B activation, fueling a positive feedback loop that amplified oncogene signaling and tumor growth in vivo. Our results indicate that inhibitors of MGAT5 might be useful in the treatment of malignancies by targeting their dependency on focal adhesion signaling for growth and metastasis. PMID- 10700234 TI - Anti-tumoral action of cannabinoids: involvement of sustained ceramide accumulation and extracellular signal-regulated kinase activation. AB - Delta9-Tetrahydrocannabinol, the main active component of marijuana, induces apoptosis of transformed neural cells in culture. Here, we show that intratumoral administration of Delta9-tetrahydrocannabinol and the synthetic cannabinoid agonist WIN-55,212-2 induced a considerable regression of malignant gliomas in Wistar rats and in mice deficient in recombination activating gene 2. Cannabinoid treatment did not produce any substantial neurotoxic effect in the conditions used. Experiments with two subclones of C6 glioma cells in culture showed that cannabinoids signal apoptosis by a pathway involving cannabinoid receptors, sustained ceramide accumulation and Raf1/extracellular signal-regulated kinase activation. These results may provide the basis for a new therapeutic approach for the treatment of malignant gliomas. PMID- 10700235 TI - Hepatocyte transplantation in a model of toxin-induced liver disease: variable therapeutic effect during replacement of damaged parenchyma by donor cells. AB - To provide long-term therapy in patients with severe toxin-induced hepatic parenchymal damage, donor hepatocytes would need to replicate and replace a large portion of the damaged parenchyma. Using a mouse model developed to reproduce this type of hepatic injury, we found that hepatocyte transplantation only slightly improved survival after transplantation despite the fact that many non survivors showed moderate liver repopulation by donor cells. Perhaps accounting for this outcome, donor parenchyma in non-survivors did not have typical lobular organization. These results indicate that the re-creation of functional parenchyma by transplanted hepatocytes requires time, during which donor cells proliferate and then establish normal parenchymal architecture. PMID- 10700236 TI - Sustained survival of human hepatocytes in mice: A model for in vivo infection with human hepatitis B and hepatitis delta viruses. AB - Persistence of hepatocytes transplanted into the same or related species has been established. The long-term engraftment of human hepatocytes into rodents would be useful for the study of human viral hepatitis, where it might allow the species, technical and size limitations of the current animal models to be overcome. Although transgenic mice expressing the hepatitis B virus (HBV) genome produce infectious virus in their serum, the viral life cycle is not complete, in that the early stages of viral binding and entry into hepatocytes and production of an episomal transcriptional DNA template do not occur. As for hepatitis delta virus (HDV), another cause of liver disease, no effective therapy exists to eradicate infection, and it remains resistant even to recent regimens that have considerably changed the treatment of HBV (ref. 13). Here, we demonstrate long term engraftment of primary human hepatocytes transplanted in a matrix under the kidney capsule of mice with administration of an agonistic antibody against c Met. These mice were susceptible to HBV infection and completion of the viral life cycle. In addition, we demonstrate super-infection of the HBV-infected mice with HDV. Our results describe a new xenotransplant model that allows study of multiple aspects of human hepatitis viral infections, and may enhance studies of human liver diseases. PMID- 10700237 TI - Regression of human metastatic renal cell carcinoma after vaccination with tumor cell-dendritic cell hybrids. AB - Reports of spontaneous regressions of metastases and the demonstration of tumor reactive cytotoxic T lymphocytes indicate the importance of the host's immune system in controlling the devastating course of metastatic renal cell carcinoma. Recent research indicates that immunization with hybrids of tumor and antigen presenting cells results in protective immunity and rejection of established tumors in various rodent models. Here, we present a hybrid cell vaccination study of 17 patients. Using electrofusion techniques, we generated hybrids of autologous tumor and allogeneic dendritic cells that presented antigens expressed by the tumor in concert with the co-stimulating capabilities of dendritic cells. After vaccination, and with a mean follow-up time of 13 months, four patients completely rejected all metastatic tumor lesions, one presented a 'mixed response', and two had a tumor mass reduction of greater 50%. We also demonstrate induction of HLA-A2-restricted cytotoxic T cells reactive with the Muc1 tumor associated antigen and recruitment of CD8+ lymphocytes into tumor challenge sites. Our data indicate that hybrid cell vaccination is a safe and effective therapy for renal cell carcinoma and may provide a broadly applicable strategy for other malignancies with unknown antigens. PMID- 10700238 TI - In vivo antigen challenge in celiac disease identifies a single transglutaminase modified peptide as the dominant A-gliadin T-cell epitope. AB - Celiac disease (CD) is an increasingly diagnosed enteropathy (prevalence, 1:200 1:300) that is induced by dietary exposure to wheat gliadins (as well as related proteins in rye and barley) and is strongly associated with HLA-DQ2 (alpha1*0501, beta1*0201), which is present in over 90% of CD patients. Because a variety of gliadin peptides have been identified as epitopes for gliadin-specific T-cell clones and as bioactive sequences in feeding studies and in ex vivo CD intestinal biopsy challenge, it has been unclear whether a 'dominant' T-cell epitope is associated with CD. Here, we used fresh peripheral blood lymphocytes from individual subjects undergoing short-term antigen challenge and tissue transglutaminase-treated, overlapping synthetic peptides spanning A-gliadin to demonstrate a transient, disease-specific, DQ2-restricted, CD4 T-cell response to a single dominant epitope. Optimal gamma interferon release in an ELISPOT assay was elicited by a 17-amino-acid peptide corresponding to the partially deamidated peptide of A-gliadin amino acids 57-73 (Q65E). Consistent with earlier reports indicating that host tissue transglutaminase modification of gliadin enhances gliadin-specific CD T-cell responses, tissue transglutaminase specifically deamidated Q65 in the peptide of A-gliadin amino acids 56-75. Discovery of this dominant epitope may allow development of antigen-specific immunotherapy for CD. PMID- 10700239 TI - Mutations in the tyrosine phosphatase CD45 gene in a child with severe combined immunodeficiency disease. AB - The hematopoietic-specific transmembrane protein tyrosine phosphatase CD45 functions to regulate Src kinases required for T- and B-cell antigen receptor signal transduction. So far, there have been no reports to our knowledge of a human deficiency in a tyrosine-specific phosphatase. Here, we identified a male patient with a deficiency in CD45 due to a large deletion at one allele and a point mutation at the other. The point mutation resulted in the alteration of intervening sequence 13 donor splice site. The patient presented at 2 months of age with severe combined immunodeficiency disease. The population of peripheral blood T lymphocytes was greatly diminished and unresponsive to mitogen stimulation. Despite normal B-lymphocyte numbers, serum immunoglobulin levels decreased with age. Thus, CD45 deficiency in humans results in T- and B lymphocyte dysfunction. PMID- 10700240 TI - Transdermal monitoring of glucose and other analytes using ultrasound. PMID- 10700242 TI - ON THE MARKET. PMID- 10700241 TI - In vivo magnetic resonance imaging of transgene expression. PMID- 10700244 TI - Mysterianism lite. PMID- 10700245 TI - Predicting perception from population codes. PMID- 10700246 TI - ChIPping away at potassium channel regulation. PMID- 10700247 TI - Toying with memory in the hippocampus. PMID- 10700248 TI - Attention - brains at work! PMID- 10700249 TI - Signaling dendritic growth in vivo PMID- 10700250 TI - Three-dimensional spatial selectivity of hippocampal neurons during space flight. PMID- 10700251 TI - NMDA receptor-mediated control of protein synthesis at developing synapses. AB - We demonstrate a rapid and complex effect of N-methyl-d-aspartate receptor (NMDAR) activation on synaptic protein synthesis in the superior colliculi of young rats. Within minutes of receptor activation, translation of alpha Ca2+/calmodulin dependent kinase II (alphaCamK II) was increased, whereas total protein synthesis was reduced. NMDAR activation also increased phosphorylation of eukaryotic elongation factor 2 (eEF2), a process known to inhibit protein translation by reducing peptide chain elongation. Low doses of cycloheximide, which reduce elongation rate independently of eEF2 phosphorylation, decreased overall protein synthesis but increased alphaCaMK II synthesis. These observations suggest that regulation of peptide elongation via eEF2 phosphorylation can link NMDAR activation to local increases in the synthesis of specific proteins during activity-dependent synaptic change. PMID- 10700252 TI - Rho GTPases regulate distinct aspects of dendritic arbor growth in Xenopus central neurons in vivo. AB - The development and structural plasticity of dendritic arbors are governed by several factors, including synaptic activity, neurotrophins and other growth regulating molecules. The signal transduction pathways leading to dendritic structural changes are unknown, but likely include cytoskeleton regulatory components. To test whether GTPases regulate dendritic arbor development, we collected time-lapse images of single optic tectal neurons in albino Xenopus tadpoles expressing dominant negative or constitutively active forms of Rac, Cdc42 or RhoA. Analysis of images collected at two-hour intervals over eight hours indicated that enhanced Rac activity selectively increased branch additions and retractions, as did Cdc42 to a lesser extent. Activation of endogenous RhoA decreased branch extension without affecting branch additions and retractions, whereas dominant-negative RhoA increased branch extension. Finally, we provide data suggesting that RhoA mediates the promotion of normal dendritic arbor development by NMDA receptor activation. PMID- 10700253 TI - Anatomical and physiological evidence for D1 and D2 dopamine receptor colocalization in neostriatal neurons. AB - Despite the importance of dopamine signaling, it remains unknown if the two major subclasses of dopamine receptors exist on the same or distinct populations of neurons. Here we used confocal microscopy to demonstrate that virtually all striatal neurons, both in vitro and in vivo, contained dopamine receptors of both classes. We also provide functional evidence for such colocalization: in essentially all neurons examined, fenoldopam, an agonist of the D1 subclass of receptors, inhibited both the Na+/K+ pump and tetrodotoxin (TTX)-sensitive sodium channels, and quinpirole, an agonist of the D2 subclass of receptors, activated TTX-sensitive sodium channels. Thus D1 and D2 classes of ligands may functionally interact in virtually all dopamine-responsive neurons within the basal ganglia. PMID- 10700254 TI - Growth cone and dendrite dynamics in zebrafish embryos: early events in synaptogenesis imaged in vivo. AB - We used time-lapse fluorescence microscopy to observe the growth of Mauthner cell axons and their postsynaptic targets, the primary motor neurons, in spinal cords of developing zebrafish embryos. Upon reaching successive motor neurons, the Mauthner growth cone paused briefly before continuing along its path. Varicosities formed at regular intervals and were preferentially associated with the target regions of the primary motor neurons. In addition, the postsynaptic motor neurons showed highly dynamic filopodia, which transiently interacted with both the growth cone and the axon. Both Mauthner cell and motor neurons were highly active, each showing motility sufficient to initiate synaptogenesis. PMID- 10700255 TI - Enrichment induces structural changes and recovery from nonspatial memory deficits in CA1 NMDAR1-knockout mice. AB - We produced CA1-specific NMDA receptor 1 subunit-knockout (CA1-KO) mice to determine the NMDA receptor dependence of nonspatial memory formation and of experience-induced structural plasticity in the CA1 region. CA1-KO mice were profoundly impaired in object recognition, olfactory discrimination and contextual fear memories. Surprisingly, these deficits could be rescued by enriching experience. Using stereological electron microscopy, we found that enrichment induced an increase of the synapse density in the CA1 region in knockouts as well as control littermates. Therefore, our data indicate that CA1 NMDA receptor activity is critical in hippocampus-dependent nonspatial memory, but is not essential for experience-induced synaptic structural changes. PMID- 10700256 TI - Muscles express motor patterns of non-innervating neural networks by filtering broad-band input. AB - We describe three slow muscles that responded to low-frequency modulation of a high-frequency neuronal input and, consequently, could express the motor patterns of neural networks whose neurons did not directly innervate the muscles. Two of these muscles responded to different frequency components present in the same input, and as a result each muscle expressed the motor pattern of a different, non-innervating, neural network. In an analogous manner, the distinct dynamics of the multiple intracellular processes that most cells possess may allow each process to respond to, and hence differentiate among, specific frequency ranges present in broad-band input. PMID- 10700257 TI - Microsaccadic eye movements and firing of single cells in the striate cortex of macaque monkeys. AB - When viewing a stationary object, we unconsciously make small, involuntary eye movements or 'microsaccades'. If displacements of the retinal image are prevented, the image quickly fades from perception. To understand how microsaccades sustain perception, we studied their relationship to the firing of cells in primary visual cortex (V1). We tracked eye movements and recorded from V1 cells as macaque monkeys fixated. When an optimally oriented line was centered over a cell's receptive field, activity increased after microsaccades. Moreover, microsaccades were better correlated with bursts of spikes than with either single spikes or instantaneous firing rate. These findings may help explain maintenance of perception during normal visual fixation. PMID- 10700258 TI - Lack of cortical contrast gain control in human photosensitive epilepsy. AB - Television and video games may be powerful triggers for visually induced epileptic seizures. To better understand the triggering elements of visual stimuli and cortical mechanisms of hyperexcitability, we examined eleven patients with idiopathic photosensitive epilepsy by recording visually evoked potentials (VEPs) in response to temporally modulated patterns of different contrast. For stimuli of low-medium, but not high, temporal frequency, the contrast dependence of VEP amplitude and latency is remarkably abnormal for luminance contrast (black white), but not so for chromatic contrast (equiluminant red-green) stimuli. We conclude that cortical mechanisms of contrast gain control for pattern stimuli of relatively low temporal frequency and high luminance contrast are lacking or severely impaired in photosensitive subjects. PMID- 10700259 TI - Learning to find a shape. AB - We studied the transition of stimuli from novel to familiar in visual search and in the guidance of attention to a particular object. Ability to identify an object improved dramatically over several days of training. The learning was specific for the object's position in the visual field, orientation and configuration. Improvement was initially localized to one or two positions near the fixation spot and then expanded radially to include the full area of the stimulus array. Characteristics of this learning process may reflect a shift in the cortical representation of complex features toward earlier stages in the visual pathway. PMID- 10700260 TI - Seeing multiple directions of motion-physiology and psychophysics. AB - Dot patterns sliding transparently across one another are normally perceived as independently moving surfaces. Recordings from direction-selective neurons in area MT of the macaque suggested that this perceptual segregation did not depend on the presence of two peaks in the population activity. Rather, the visual system seemed to use overall shape of the population response to determine the number and directions of motion components. This approach explained a number of perceptual phenomena, including susceptibility of the motion system to direction metamers, motion patterns combining three or five directions incorrectly perceived by subjects as comprising only two directions. Our findings offer insights into the coding of multi-valued sensory signals and provide constraints for biologically based computational models. PMID- 10700261 TI - A multimodal cortical network for the detection of changes in the sensory environment. AB - Sensory stimuli undergoing sudden changes draw attention and preferentially enter our awareness. We used event-related functional magnetic-resonance imaging (fMRI) to identify brain regions responsive to changes in visual, auditory and tactile stimuli. Unimodally responsive areas included visual, auditory and somatosensory association cortex. Multimodally responsive areas comprised a right-lateralized network including the temporoparietal junction, inferior frontal gyrus, insula and left cingulate and supplementary motor areas. These results reveal a distributed, multimodal network for involuntary attention to events in the sensory environment. This network contains areas thought to underlie the P300 event-related potential and closely corresponds to the set of cortical regions damaged in patients with hemineglect syndromes. PMID- 10700262 TI - The neural mechanisms of top-down attentional control. AB - Selective visual attention involves dynamic interplay between attentional control systems and sensory brain structures. We used event-related functional magnetic resonance imaging (fMRI) during a cued spatial-attention task to dissociate brain activity related to attentional control from that related to selective processing of target stimuli. Distinct networks were engaged by attention-directing cues versus subsequent targets. Superior frontal, inferior parietal and superior temporal cortex were selectively activated by cues, indicating that these structures are part of a network for voluntary attentional control. This control biased activity in multiple visual cortical areas, resulting in selective sensory processing of relevant visual targets. PMID- 10700263 TI - Voluntary orienting is dissociated from target detection in human posterior parietal cortex. AB - Human ability to attend to visual stimuli based on their spatial locations requires the parietal cortex. One hypothesis maintains that parietal cortex controls the voluntary orienting of attention toward a location of interest. Another hypothesis emphasizes its role in reorienting attention toward visual targets appearing at unattended locations. Here, using event-related functional magnetic resonance (ER-fMRI), we show that distinct parietal regions mediated these different attentional processes. Cortical activation occurred primarily in the intraparietal sulcus when a location was attended before visual-target presentation, but in the right temporoparietal junction when the target was detected, particularly at an unattended location. PMID- 10700264 TI - Plans for special focus issues. PMID- 10700265 TI - Nitrous oxide reductase from CuA to CuZ. PMID- 10700266 TI - New wine from old barrels. PMID- 10700267 TI - Class (I) will come to order--not. PMID- 10700268 TI - Passing the baton in base excision repair. PMID- 10700269 TI - A surprising function for SRP RNA? PMID- 10700270 TI - One motif--many different reactions. PMID- 10700271 TI - Relieving repression. PMID- 10700272 TI - Protein recognition by NMR. PMID- 10700273 TI - Antibiotics from soil bacteria. PMID- 10700274 TI - Picture story. Seeing double in living cells. PMID- 10700275 TI - A novel type of catalytic copper cluster in nitrous oxide reductase. AB - Nitrous oxide (N20) is a greenhouse gas, the third most significant contributor to global warming. As a key process for N20 elimination from the biosphere, N20 reductases catalyze the two-electron reduction of N20 to N2. These 2 x 65 kDa copper enzymes are thought to contain a CuA electron entry site, similar to that of cytochrome c oxidase, and a CuZ catalytic center. The copper anomalous signal was used to solve the crystal structure of N20 reductase from Pseudomonas nautica by multiwavelength anomalous dispersion, to a resolution of 2.4 A. The structure reveals that the CuZ center belongs to a new type of metal cluster, in which four copper ions are liganded by seven histidine residues. N20 binds to this center via a single copper ion. The remaining copper ions might act as an electron reservoir, assuring a fast electron transfer and avoiding the formation of dead end products. PMID- 10700276 TI - Crystal structure of the protein disulfide bond isomerase, DsbC, from Escherichia coli. AB - DsbC is one of five Escherichia coli proteins required for disulfide bond formation and is thought to function as a disulfide bond isomerase during oxidative protein folding in the periplasm. DsbC is a 2 x 23 kDa homodimer and has both protein disulfide isomerase and chaperone activity. We report the 1.9 A resolution crystal structure of oxidized DsbC where both Cys-X-X-Cys active sites form disulfide bonds. The molecule consists of separate thioredoxin-like domains joined via hinged linker helices to an N-terminal dimerization domain. The hinges allow relative movement of the active sites, and a broad uncharged cleft between them may be involved in peptide binding and DsbC foldase activities. PMID- 10700277 TI - Solution structure of the smallest cofactor-active fragment of thrombomodulin. AB - A glycosylated fragment of thrombomodulin containing two epidermal growth factor like domains (TMEGF45) was analyzed by NMR. The 4th-domains structure of this two domain fragment is similar to that of the individual domain previously determined. The 5th-domain, which has uncrossed disulfide bonds, is not as well determined in the two-domain fragment than the individual domain previously solved. The flexibility of the 5th-domain is consistent with low heteronuclear NOEs. In the individual 5th-domain, Met 388 was disordered, and key thrombin binding residues formed a hydrophobic core. By contrast, in TMEGF45, Met 388 is in the 5th-domain core, positioned by Phe 376 from the 4th-domain. As a result, key thrombin binding residues that were in the core of the individual domain are expelled. Upon thrombin binding, chemical shifts of two residues in the 4th domain, the three interdomain linker residues, and nearly all of the 5th-domain are perturbed. Thus, TMEGF45 binds thrombin by an induced fit mechanism involving a flexible 5th-domain. PMID- 10700278 TI - Substrate-induced exposure of an energy-coupling motif of a membrane transporter. AB - BtuB is an outer membrane protein responsible for the uptake of vitamin B12 by Escherichia coli. It belongs to a family of bacterial transport proteins that derive energy for transport by coupling to the trans-periplasmic energy-coupling protein TonB. Using site-directed spin labeling and EPR we investigated the structure and substrate-induced changes in the TonB box, a highly conserved region in all TonB dependent transporters that may couple to TonB. In the absence of substrate, the line widths and collision parameters from EPR are consistent with this domain existing in a structured helical conformation that contacts the barrel of the transporter. Addition of substrate converts this segment into an extended structure that is highly dynamic, disordered and probably extended into the periplasm. This structural change demonstrates that the TonB box cycles between sequestered and accessible states in a substrate-dependent fashion. In a transport defective mutant of BtuB, this conformational cycle is disrupted and the TonB box appears to be extended even in the absence of substrate. These data suggest that the TonB box extends into the periplasm and interacts with TonB only in PMID- 10700279 TI - A closer view of the conformation of the Lac repressor bound to operator. AB - Crystal structures of the Lac repressor, with and without isopropyithiogalactoside (IPTG), and the repressor bound to operator have provided a model for how the binding of the inducer reduces the affinity of the repressor for the operator. However, because of the low resolution of the operator-bound structure (4.8 A), the model for the allosteric transition was presented in terms of structural elements rather than in terms of side chain interactions. Here we have constructed a dimeric Lac repressor and determined its structure at 2.6 A resolution in complex with a symmetric operator and the anti inducer orthonitrophenylfucoside (ONPF). The structure enables the induced (IPTG bound) and repressed (operator-bound) conformations of the repressor to be compared in atomic detail. An extensive network of interactions between the DNA binding and core domains of the repressor suggests a possible mechanism for the allosteric transition. PMID- 10700280 TI - Structural basis of gene regulation by the tetracycline inducible Tet repressor operator system. AB - The tetracycline repressor (TetR) regulates the most abundant resistance mechanism against the antibiotic tetracycline in grain-negative bacteria. The TetR protein and its mutants are commonly used as control elements to regulate gene expression in higher eukaryotes. We present the crystal structure of the TetR homodimer in complex with its palindromic DNA operator at 2.5 A resolution. Comparison to the structure of TetR in complex with the inducer tetracycline-Mg2+ allows the mechanism of induction to be deduced. Inducer binding in the repressor core initiates conformational changes starting with C-terminal unwinding and shifting of the short helix a6 in each monomer. This forces a pendulum-like motion of helix a4, which increases the separation of the attached DNA binding domains by 3 A, abolishing the affinity of TetR for its operator DNA. PMID- 10700281 TI - A novel NMR method for determining the interfaces of large protein-protein complexes. AB - Identification of the interfaces of large (Mr > 50,000) protein-protein complexes in solution by high resolution NMR has typically been achieved using experiments involving chemical shift perturbation and/or hydrogen-deuterium exchange of the main chain amide groups of the proteins. Interfaces identified using these techniques, however, are not always identical to those revealed using X-ray crystallography. In order to identify the contact residues in a large protein protein complex more accurately, we developed a novel NMR method that uses cross saturation phenomena in combination with TROSY detection in an optimally deuterium labeled system. PMID- 10700282 TI - Structure of the Mad2 spindle assembly checkpoint protein and its interaction with Cdc20. AB - The checkpoint protein Mad2 inhibits the activity of the anaphase promoting complex by sequestering Cdc20 until all chromosomes are aligned at the metaphase plate. We report the solution structure of human Mad2 and its interaction with Cdc20. Mad2 possesses a novel three-layered alpha/beta fold with three alpha helices packed between two beta-sheets. Using deletion mutants we identified the minimal Mad2-binding region of human Cdc20 as a 40-residue segment immediately N terminal to the WD40 repeats. Mutagenesis and NMR titration experiments show that a C-terminal flexible region of Mad2 is required for binding to Cdc20. Mad2 and Cdc20 form a tight 1:1 heterodimeric complex in which the C-terminal segment of Mad2 becomes folded. These results provide the first structural insight into mechanisms of the spindle assembly checkpoint. PMID- 10700283 TI - Novel fold and capsid-binding properties of the lambda-phage display platform protein gpD. AB - The crystal structure of gpD, the capsid-stabilizing protein of bacteriophage lambda, was solved at 1.1 A resolution. Data were obtained from twinned crystals in space group P21 and refined with anisotropic temperature factors to an R factor of 0.098 (Rfree = 0. 132). GpD (109 residues) has a novel fold with an unusually low content of regular secondary structure. Noncrystallographic trimers with substantial intersubunit interfaces were observed. The C-termini are well ordered and located on one side of the trimer, relatively far from its three-fold axis. The N-termini are disordered up to Ser 15, which is close to the three-fold axis and on the same side as the C-termini. A density map of the icosahedral viral capsid at 15 A resolution, obtained by cryo-electron microscopy and image reconstruction, reveals gpD trimers, seemingly indistinguishable from the ones seen in the crystals, at all three-fold sites. The map further reveals that the side of the trimer that binds to the capsid is the side on which both termini reside. Despite this orientation of the gpD trimer, fusion proteins connected by linker peptides to either terminus bind to the capsid, allowing protein and peptide display. PMID- 10700284 TI - Crystal structures of homoserine dehydrogenase suggest a novel catalytic mechanism for oxidoreductases. AB - The structure of the antifungal drug target homoserine dehydrogenase (HSD) was determined from Saccharomyces cerevisiae in apo and holo forms, and as a ternary complex with bound products, by X-ray diffraction. The three forms show that the enzyme is a dimer, with each monomer composed of three regions, the nucleotide binding region, the dimerization region and the catalytic region. The dimerization and catalytic regions have novel folds, whereas the fold of the nucleotide-binding region is a variation on the Rossmann fold. The novel folds impose a novel composition and arrangement of active site residues when compared to all other currently known oxidoreductases. This observation, in conjunction with site-directed mutagenesis of active site residues and steady-state kinetic measurements, suggest that HSD exhibits a new variation on dehydrogenase chemistry. PMID- 10700285 TI - Site-site communication in the EF-hand Ca2+-binding protein calbindin D9k. AB - The cooperative binding of Ca2+ ions is an essential functional property of the EF-hand family of Ca2+-binding proteins. To understand how these proteins function, it is essential to characterize intermediate binding states in addition to the apo- and holo-proteins. The three-dimensional solution structure and fast time scale internal motional dynamics of the backbone have been determined for the half-saturated state of the N56A mutant of calbindin D9k with Ca2+ bound only in the N-terminal site. The extent of conformational reorganization and a loss of flexibility in the C-terminal EF-hand upon binding of an ion in the N-terminal EF hand provide clear evidence of the importance of site-site interactions in this family of proteins, and demonstrates the strength of long-range effects in the cooperative EF-hand Ca2+-binding domain. PMID- 10700295 TI - Sampling kinases in single cells PMID- 10700286 TI - Structure of a closed form of human malic enzyme and implications for catalytic mechanism. AB - Malic enzymes are widely distributed in nature and have many biological functions. The crystal structure of human mitochondrial NAD(P)+-dependent malic enzyme in a quaternary complex with NAD+, Mn++ and oxalate has been determined at 2.2 A resolution. The structures of the quaternary complex with NAD+, Mg++, tartronate or ketomalonate have been determined at 2.6 A resolution. The structures show the enzyme in a closed form in these complexes and reveal the binding modes of the cation and the inhibitors. The divalent cation is coordinated in an octahedral fashion by six ligating oxygens, two from the substrate/inhibitor, three from Glu 255, Asp 256 and Asp 279 of the enzyme, and one from a water molecule. The structural information has significant implications for the catalytic mechanism of malic enzymes and identifies Tyr 112 and Lys 183 as possible catalytic residues. Changes in tetramer organization of the enzyme are also observed in these complexes, which might be relevant for its cooperative behavior and allosteric control. PMID- 10700296 TI - Systemic local-anaesthetic-type drugs in chronic pain: a systematic review. AB - Basic research indicates that systemic local-anaesthetic-type drugs that block sodium channels are effective in pain due to nerve damage. These drugs were first used as analgesics in the 1950s and they are still commonly used to try to relieve neuropathic pain and incident pain caused by cancer. As they are potentially toxic, these drugs should not be used without proven effectiveness. For these reasons, a systematic review of randomized controlled trials of systemically administered local-anaesthetic-type drugs in chronic pain was performed. Main outcomes were pain relief or pain intensity difference and adverse effects. Twenty-one reports were found, and four publications were excluded. In the remaining 17 studies (450 patients), 10 used intravenous lignocaine, two used intranasal lignocaine, four used oral mexiletine and one used oral tocainide. The best documented effective dose of intravenous lignocaine was 5 mg/kg, and when infused over 30 min it was well tolerated. Intravenous lignocaine was effective in all four studies in non-cancer-related neuropathic pain. In migraine, lignocaine produced an inconsistent effect. Lignocaine was without effect in all three studies in cancer-related pain. Oral mexiletine showed some efficacy in all three studies in pain due to peripheral nerve damage, but lacked effect in the only study in central pain. Only minor dose-related adverse effects were reported in the 85 patients given mexiletine 225-750 mg. Local-anaesthetic-type drugs are effective in pain due to nerve damage, but there is little or no evidence to support their use in cancer-related pain. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700297 TI - Different direct pathways of locus coeruleus to medial prefrontal cortex and centrolateral thalamic nucleus: electrical stimulation effects on the evoked responses to nociceptive peripheral stimulation. AB - Projections from the locus coeruleus (LC) to the centrolateral thalamus (Cl) and the medial prefrontal cortex (PfCx) were studied using orthodromic and antidromic stimulation techniques. The LC is a major noradrenergic source in the central nervous system, and its descending projections provide an important source of pain suppression at spinal level. Previously, the author has described a cortico thalamic loop involved in pain modulation. The present paper reports on a study of the participation of LC as part of an ascending pain-control system acting on the cortico-thalamic loop.Rats were anaesthetized with halothane, and single unit recordings were made in LC using glass micropipettes. Stainless steel electrodes were placed in cortex and thalamus for electrical stimulation.Stimulation in PfCx or Cl produces antidromic responses in neurons in LC. The latencies, conduction velocity and location of neurons in LC projecting to PfCx or Cl structures are described. Separate projections to both structures have significantly different conducting velocities, arriving earlier at Cl (mean conduction velocities 0.27 and standard deviation +/-0.06 m/s) and then at PfCx (mean conduction velocities 0.20+/- 0.04 m/s). The presence of orthodromic responses suggests reciprocal connections. The paper also describes the suppression of spontaneous and nociceptive-evoked activity in the PfCx and Cl following electrical stimulation in LC.It is proposed that the LC innervation could be associated with an ascending noradrenergic system acting upon a Cl-PfCx pain-modulation mechanism. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700298 TI - Effects of an opioid antagonist on pain intensity and withdrawal reflexes during induction of hypnotic analgesia in high- and low-hypnotizable volunteers. AB - The aim of this investigation was to study the effect of suggestions of hypnotic analgesia on spinal pain transmission and processing. Pain intensity and amplitude of nociceptive withdrawal reflexes to electrical stimuli were measured in 10 high- and 10 low-hypnotizable subjects during two sessions taking place at least 24 h apart under five conditions of: (1) pre-hypnosis; (2) neutral hypnotic relaxation; (3) suggestions of hypnotic analgesia; (4) suggestions of hypnotic analgesia after injections of either naloxone (1 ml, 1 mg/ml) or saline (1 ml) under double-blinded conditions; and (5) post-hypnosis. The conditions of naloxone or saline were allocated at random to either Day 1 or Day 2 in a double blinded fashion. Results showed significant reductions of pain intensity during hypnotic analgesia, and a significant reduction in nociceptive reflexes during hypnotic analgesia on Day 1 in the highly hypnotizable group. No differences were found for low-hypnotizable subjects. The results support previous findings that pain intensity as well as the nociceptive reflex can be modulated by suggestions of hypnotic analgesia. While no effect of naloxone on pain intensity was found during hypnotic analgesia, naloxone significantly reversed the suppressive effect of suggestions of hypnotic analgesia on reflexes in high-hypnotizable subjects. Subsequent analysis showed that the effect of naloxone was associated with the intensity of the stimulus needed to elicit a reflex, and was unrelated to hypnotic susceptibility when controlling for stimulus intensity. These results suggest that the effect of naloxone was related to the greater stimulus intensities needed to elicit a reflex in the high-hypnotizable group, rather than to hypnosis or hypnotic susceptibility in itself. It is unclear why greater stimulus intensities were needed in high-hypnotizable subjects and further studies are needed. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700299 TI - Systemic morphine in the prevention of allodynia in the rat spinal nerve ligation model of neuropathic pain. AB - Peripheral nerve injury may lead to neuropathic pain that has been considered unresponsive to opioids. In animal models of neuropathic pain, there are previous data of both increased and decreased effect of opioids, but only limited information of the long-term effects of opioid treatment on the development of the symptoms of neuropathy. The possibility of preventing the development of signs of neuropathy with either a single pre-injury injection or chronic postinjury administration of morphine was studied in rats with unilateral peripheral neuropathy due to tight ligation of the L5 and L6 spinal nerves. These rats developed both mechanical and cold allodynia, but no thermal hyperalgesia. Neither subcutaneous morphine given as single injection (10 mg/kg) before the nerve injury nor chronic administration starting immediately after surgery using slow-release pellets (one, two or five pellets containing 75 mg of morphine, resulting in a total dose of 75, 150 or 375 mg) prevented the development of either mechanical or cold allodynia. No autotomy, signs of distress, altered social behaviour or morphine withdrawal was seen in any of the rats. The fact that neuropathic pain-like symptoms were not attenuated by any of the treatments studied could indicate that neither premedication nor postoperative pain management with systemic morphine is effective in preventing postoperative neuropathic pain. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700300 TI - Pain, coping and psychological well-being in late life. AB - In the present article, the relationships between pain, coping, functional capacity and psychological well-being are examined in a population of older patients (>/=60 years; n=202) treated for a variety of somatic complaints in a university-affiliated hospital. Results indicate that moderate to extreme pain is common in older patients and often occurs in several body regions simultaneously. Extreme pain in one or more body regions is associated with reduced daily functional capacity, lower values for life satisfaction and self-evaluated competence, and more negative attitudes towards the present and future. Results of a hierarchical cluster analysis reveal interindividual differences in coping approaches among older patients suffering from extreme pain; they also emphasize the importance of cognitive strategies and life-review coping. Relevance for clinical practice with older pain patients is discussed.In the present article, the relationships between pain, coping, functional capacity and psychological well-being are examined in a population of older patients (>/=60 years; n=202) treated for a variety of somatic complaints in a university-affiliated hospital. Results indicate that moderate to extreme pain is common in older patients and often occurs in several body regions simultaneously. Extreme pain in one or more body regions is associated with reduced daily functional capacity, lower values for life satisfaction and self-evaluated competence, and more negative attitudes towards the present and future. Results of a hierarchical cluster analysis reveal interindividual differences in coping approaches among older patients suffering from extreme pain; they also emphasize the importance of cognitive strategies and life-review coping. Relevance for clinical practice with older pain patients is discussed. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700301 TI - Transdermal fentanyl for pain control in adults with chronic cancer pain. AB - The transdermal therapeutic system (TTS) for fentanyl is a drug-delivery system for use in patients with chronic pain who require an opioid analgesic. A multicentre, randomized, double-blind, placebo-controlled study was performed to evaluate the efficacy and safety of TTS-fentanyl as an analgesic for chronic cancer pain. One hundred and thirty-eight patients entered a 15-day dose titration period, followed by a 9-day double-blind period (95 patients) with TTS fentanyl or placebo. Fifty-five patients entered a follow-up period of indefinite duration. For the majority of patients, TTS-fentanyl 50-75 ug/h provided effective analgesia. Due to an unexpectedly high placebo response, it was not possible to show fentanyl to be statistically superior to placebo at the 5% significance level. Nine patients treated with fentanyl and 13 treated with placebo were withdrawn from the study during the double-blind therapy because of insufficient efficacy (not significant), while 66% of fentanyl-treated patients experienced effective pain control compared with 48% of placebo-treated patients (p=0.071). During the course of the double-blind therapy, the mean dose of rescue morphine increased slightly more in the placebo group than in the fentanyl group. At the end of the double-blind phase, the investigators rated trial medication as being 'good' or 'excellent' in 30 patients in the fentanyl group and 23 in the placebo group. TTS-fentanyl appeared to be well tolerated, with a low incidence of constipation, somnolence and nausea. Due to an unexpectedly high placebo response it was not possible to demonstrate fentanyl to be statistically superior to placebo. This may reflect the practical difficulties of performing clinical trials in cancer patients with great inter-individual variability. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700302 TI - Influence of formalin concentration on the antinociceptive effects of anti inflammatory drugs in the formalin test in rats: separate mechanisms underlying the nociceptive effects of low- and high-concentration formalin. AB - The present study has assessed the relationship between formalin-induced nociception and formalin-induced inflammation by comparing the dose-related effects of anti-inflammatory treatments on both nociceptive scores and plasma extravasation in the rat hind paw in response to high and low concentrations of formalin. The degree of plasma extravasation produced by 1% formalin did not differ significantly from that produced by the same volume of saline, and was not significantly affected by either of the anti-inflammatory agents. The 5% formalin injection produced significant plasma extravasation that was dose-dependently reduced by both dexamethasone and ibuprofen. The early-phase nociceptive responses to either 1 or 5% formalin were not affected significantly by either of the anti-inflammatory agents. In contrast, the late-phase nociceptive responses to 5%, but not 1%, formalin were dose-dependently reduced by both dexamethasone and ibuprofen. The present study suggests that there is a positive correlation between the nociceptive and inflammatory effects of formalin in the rat hind paw. However, only a high concentration of formalin, which produces significant plasma extravasation, is capable of demonstrating the antinociceptive effects of anti inflammatory agents, and the effects are restricted to the late phase of the formalin test. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700303 TI - Where does it hurt? Describing the body locations of chronic pain. AB - Chronic pain patients have complex problems. Due to this, much research effort has been expended on the classification of pain patients and the classification of pain problems. A mainstay of most pain classification systems is the use of the physical location of the pain. Yet describing the location of the patient's pain is not straightforward. Many patients have pain at multiple sites and thus simple statements such as 'patients with low back pain' have considerable ambiguity. Does the statement refer to patients with pain just in the lower back or those with low back pain who may also have pain elsewhere (e.g. pain down the leg)? If patients do have pain elsewhere, at what other body sites and what are the implications of this? This paper presents data on the body location of pain for a large sample of 5279 patients seen with chronic pain in Scotland and the north of England. It shows that one-third of patients have pain in multiple locations, and that using a single body site to classify patients leads to groups with large overlaps. Thus, 38% of patients reported pain in the 'lower back/spine' and 34% reported pain in the 'buttock, leg, foot' - but there was considerable overlap between these groups. Nineteen percent of patients reported pain in both of these body areas, and one-third of these patients also had pain in at least one further body area. Furthermore, a systematic look at patients with diverse physical pain locations but a single site in common shows large demographic differences. Common pain groupings help to reduce the confusion; 13 pain site descriptions were able to account for 82% of all patients. The remaining 18% of patients had pain in a combination of body sites which they shared with fewer than 1% of other patients. Thus, pain patients are widely heterogeneous and complex. Patients report pain in more complex patterns than can easily be captured by a single body-site code. Further, large demographic differences between patients with different painful sites, even when they have at least one pain site in common, suggests that grouping patients based on a single site descriptor may be inappropriate. These findings have important implications for chronic pain description and classification. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700304 TI - Long-term intrathecal (i.t.) infusion of bupivacaine relieved intractable pain and spasticity in a patient with multiple sclerosis. AB - There is no reliable method to relieve both 'refractory' pain and spasticity in patients with multiple sclerosis (MS). This paper reports on the long-term use of continuous intrathecal bupivacaine infusion in such a patient. The patient under study was a 56-year-old woman affected for 18 years by MS, unsuccessfully treated with analgesics, baclofen, opioids, peripheral neurolysis (obturator nerves, lumbar plexus) and six intrathecal neurolyses of the L4-S3 nerve roots, each time with 1.5 ml of 50% phenol in glycerol. Intrathecal baclofen was not considered (MS with bulbar location and neurogenic pains). An intrathecal catheter was inserted via the L3-L4 interspace and its tip was placed at the height of the T12 L1 intervertebral disc. An intrathecal infusion of 0.5% bupivacaine at a rate of 3 ml (=15 mg)/day was started. The infusion rate was gradually increased from 20 mg on the first day to 95 mg/day after 68 days. The pain intensity decreased from a mean visual analogue score (VAS(mean)) of 7 before treatment to 1 (on a 0-10 scale) during the intrathecal treatment. The patient became free from pain and spasticity. No side-effects or complications were recorded. The treatment was given for 712 days, at which point the patient died (unrelated to the treatment). Intrathecal infusion of bupivacaine relieved 'refractory' spasticity and pain in a MS patient in whom administration of intrathecal baclofen was contraindicated and neurodestructive procedures had been inefficient. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700305 TI - Cerebral blood-flow changes evoked by two levels of painful heat stimulation: a positron emission tomography study in humans. AB - Positron emission tomography (PET) and accumulation of H(2)(15)O as a marker of neuronal activity were used to create maps of cerebral blood-flow changes evoked by painful heat stimulation in 10 subjects. Two levels of painful tonic and phasic heat stimuli were applied with use of a newly developed contact heat thermode on the volar surface of the dominant (right) arm. The subjects participated in two separate PET sessions. Maps reflecting low and high levels of painful tonic heat were obtained in the first session, and low and high levels of painful phasic heat in the second session. The subjects scored their peak pain intensity and unpleasantness on 10-cm visual analogue scales. For each subject, PET images were aligned to nuclear magnetic resonance (NMR) images and remapped into the standardized co-ordinate system of Talairach. After normalization of the PET volumes, subtraction images were formed voxel-by-voxel and converted to a t statistic volume. The perceived pain intensity and unpleasantness were identical with painful tonic and phasic heat stimulation. Directed searches revealed significant blood-flow increases in the contralateral primary sensorimotor cortex (MI/SI), SII, insular cortex and cingulate cortex when the low tonic heat map was subtracted from the high. A similar, but not identical, pain-processing network was observed for the maps representing the subtraction of low and high phasic heat. In this subtraction, the blood-flow increases in MSI/SI did not reach statistical significance, and significant blood flow decreases were found in the contralateral middle temporal gyrus. Finally, the location of the activation site in the cingulate cortex was different from that observed during tonic heat pain. This study has provided more evidence for the existence of a common pain processing network engaged during the perception of different levels of toxic and phasic heat pain. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700306 TI - Pain management of arteriovenous fistula cannulation in haemodialysis children: efficacy of EMLA anaesthetic cream. AB - The aim of the present study was to evaluate the efficacy of EMLA cream (containing a eutectic mixture of local anaesthetics) in controlling pain due to arteriovenous fistula cannulation in teenagers undergoing chronic haemodialysis. The study was conducted in two phases, one prospective, the other a blind randomized trial, at the Paediatric Haemodialysis service of the Paediatrics Department of Padua University, Italy. It involved six teenagers, aged 12-18 years. Pain was measured using the visual analogue scale, indirect evaluation by nurses and a four-category verbal rating scale. Results showed that: (1) the visual analogue scale calls for an adequate training period; and (2) the EMLA cream might be effective in controlling cannulation-related pain but emotional factors, such as uncontrolled fear and stress, can interfere with the global efficacy of the analgesic approach. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700307 TI - Morphine use and pharmacokinetics in patients with chest pain due to suspected or definite acute myocardial infarction. AB - The characteristics of chest pain due to suspected acute myocardial infarction and morphine use during the first 3 hospital days are described in a population of 2988 consecutive patients admitted to hospital. The duration of pain was usually less than 24 h (mean 20.9+/-0.55 h), and only 24.8% of patients experienced chest pain of longer duration. The majority of patients had only one attack of pain, but 34.4% experienced four or more attacks during hospitalization. A mean morphine dose of 6.7+/-0.2 mg was administered over the 3 hospitalization days, but surprisingly 52.4% of all patients required no morphine analgesia at all. Independent predictors of an increased morphine consumption were initial degree of suspicion of acute myocardial infarction, ST changes on admission ECG, male sex, a history of angina pectoris and a history of congestive heart failure. In a separate pharmacokinetic/pharmacodynamic study in 10 patients, plasma concentrations of morphine and its major metabolites, morphine-3 glucuronide (M3G) and morphine-6-glucuronide (M6G), were measured after intravenous administration of morphine. In this patient group, terminal half-life of unchanged morphine ranged from 0.77 to 3.22 h. M3G and M6G plasma concentrations increased gradually up to 60-90 min after the intravenous morphine injection. Initial pain intensity by numerical rating scale was 6.6+/-0.6 (arbitrary units), and after morphine administration, there was a rapid and significant decrease in pain intensity. After 20 min, pain relief was 69+/-11% and remained at this level during the following 8 h observation period. It is concluded that the need for morphine administration in patients with suspected or definite acute myocardial infarction, differs among subgroups of patients and, in particular, higher doses are needed in those with a strong suspicion of myocardial infarction at arrival. When intravenous morphine is given, it attains full effect 20 min after injection. Furthermore, the active morphine metabolites M3G and M6G appear rapidly in the circulation, which could influence the analgesic response to morphine treatment. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700308 TI - Imipramine does not affect argon-laser-induced pin-prick pain thresholds and laser-evoked cerebral potentials. AB - The tricyclic antidepressant imipramine has shown analgesic effect in human clinical and experimental pain studies. The aim of the present study was to test the effect of imipramine on a pure short-term nociceptive stimulus with pin-prick pain quality. In a randomized, placebo-controlled, double-blind, crossover study, the hypoalgesic effect of a single oral dose of 100 mg imipramine was investigated in 10 healthy volunteers. Test procedures performed before and 2, 4, 6, 8, 10, 12 and 14 h after medication included determination of warmth and pin prick pain thresholds to high-energy argon laser light stimulation on the hand, as well as laser-evoked cerebral potentials to suprathreshold stimulation. Both the warmth and the pin-prick pain thresholds (p=0.49 and 0.85) and the root mean square of the laser-evoked potentials (p=0.89) were unaltered by imipramine. It is concluded that a single oral dose of 100 mg imipramine has no effect on pin prick pain. This study demonstrates the important fact that a drug may show clear analgesic effect in some experimental pain models while it is without effect in other models; e.g. imipramine is known to affect pain tolerance and summation thresholds. Pre-clinical tests of potentially analgesic drugs should therefore be based on different pain-stimulation modalities. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700309 TI - Effects of staphylococcus toxoid vaccine on pain and fatigue in patients with fibromyalgia/chronic fatigue syndrome. AB - Positive results of pilot studies of the effect of staphylococcus toxoid vaccine in patients with fibromyalgia and chronic fatigue syndrome were the incitement to the present, placebo-controlled study. It included 28 patients who fulfilled the criteria for both fibromyalgia and chronic fatigue syndrome. The effect of vaccination with a staphylococcus toxoid was compared with the effect of injections of sterile water. Psychometric assessment was made using 15 items from the comprehensive psychopathological rating scale (CPRS), Zung's self-rating depression scale and clinical global impressions (CGI). The visual analogue scale (VAS) was used to measure pain levels, and a hand-held electronic pressure algometer was used to measure pressure pain thresholds. Significant improvement was seen in seven of the 15 CPRS items in the vaccine group when pretreatment values were compared to post-treatment values. In CPRS <>, there were significant intergroup differences, and in CPRS <> intergroup differences bordered on significance. There was no significant improvement in CPRS items in the placebo group. Clinical global impressions showed significant improvement in the vaccine-treated group, and VAS did so in both groups. In a follow-up study of 23 patients, the vaccine treatment was continued for 2-6 years. Fifty percent were rehabilitated successfully and resumed half-time or full-time work. The results of this study support the authors>> hypothesis that treatment with staphylococcus toxoid may be a fruitful strategy in patients with fibromyalgia and chronic fatigue syndrome. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700310 TI - Immunoisolating encapsulation of intrathecally implanted bovine chromaffin cells prolongs their survival and produces anti-allodynic effect in spinally injured rats. AB - We have previously reported that intrathecal (i.t.) implantation of bovine chromaffin cells has an anti-allodynic effect in a rat model of mechanical and cold allodynia-like neuropathic pain after spinal cord injury. The technique of encapsulation of the cells by a semipermeable membrane has been developed recently. The present study was undertaken to investigate the effects of encapsulated bovine chromaffin cells on the allodynia-like pain in the same model. Capsules with bovine chromaffin cells or control capsules were implanted in the spinal subarachnoidal space in rats. Their response in behavioural tests were recorded for 2 months. At termination, the capsules were explanted and examined morphologically with tyrosine hydroxylase immunohistochemistry. The mechanical allodynia was totally abolished from week 2 after implantation of the cells and throughout the 8-week test period. The abnormal cold response was also attenuated in about half of the animals. The threshold to acute nociceptive stimulation was not affected. Eight weeks after implantation, 60-80% of the encapsulated chromaffin cells were still tyrosine hydroxylase positive. No effects were observed with control capsules. The results indicate that spinal implantation of encapsulated xenogeneic chromaffin cells may be useful in treating some refractory painful states associated with spinal cord injury. Immunoisolation of chromaffin cells by a semipermeable membrane may inhibit immunorejection, prolong the survival of the cells and enhance their anti allodynic effect. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700311 TI - Inhibitory effects of glutamate-induced activation of thalamic nucleus submedius are mediated by ventrolateral orbital cortex and periaqueductal gray in rats. AB - This study found that in lightly-anesthetized rats a unilateral micro-injection of glutamate (200 mm, 0.5 ul) into the thalamic nucleus submedius (Sm) markedly depressed the radiant heat-evoked tail flick (TF) reflex. After injection, the mean TFL increased 25.6+/-6.5% (n=24) of the baseline at 5 min, up to a peak value (48.4+/-7.2%) at 20 min, and recovered to the baseline level at 60 min. This inhibitory effect was dose-related and repeatable over a time interval of 1.0-1.5 h in the same animal. Furthermore, micro-injections of gamma-aminobutyric acid (GABA) (100 mm) into the ipsilateral ventrolateral orbital cortex (VLO) (0.7 ul), or bilaterally into the lateral or ventrolateral parts of the periaqueductal gray (PAG) (0.5 ul on each side), eliminated the Sm-evoked inhibition. After GABA was injected into VLO or PAG, the Sm applications of glutamate failed to produce any significant changes in TFL, with the TFL changes being similar to the saline control (p>0.05). These results confirmed our previous findings that electrical stimulation of Sm depressed the rat TF reflex and that this inhibitory effect was blocked by electrolytic lesion of the VLO or PAG. Therefore, the present study provides further support for the hypothesis that Sm plays an important role in modulation of nociception, and that its effects are mediated by the VLO-PAG pathway, leading to activation of the brainstem descending inhibitory system and depression of the nociceptive inputs at the spinal cord level. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700312 TI - Ectopic mechanosensitivity in injured sensory axons arises from the site of spontaneous electrogenesis. AB - Injured sensory axons trapped in a neuroma or freely regenerating in the distal nerve stump, frequently display ectopic mechanosensitivity, spontaneous impulse discharge or both. This abnormal neural activity is thought to contribute to spontaneous and movement-evoked neuropathic paraesthesias, dysaesthesias and pain, as well as to allodynia and hyperalgesia. The present paper examines the relationship between mechanosensitivity and spontaneous discharge in three distinct sciatic nerve injury models in the rat: nerve transection (neuroma), nerve crush and chronic nerve constriction injury (CCI). Impulse pattern analysis was used to determine that the sites of mechanosensitivity and of spontaneous electrogenesis are either identical or very close to one another. This suggests that mechanosensitivity and spontaneous firing are aspects of a single underlying pathophysiological process. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700314 TI - Psychosomatic pain. PMID- 10700313 TI - Upside assay sensitivity in a dental pain model. AB - The extent of surgical trauma was related to postoperative pain intensity in a previous study. However, more extensive surgical procedures with higher baseline pain intensity did not appear to influence the ability to document the additive analgesic effect of codeine when given with paracetamol, partly due to large interindividual variation in baseline pain intensity. The aim of the present study was to attempt to improve upside assay sensitivity in this dental pain model by: (1) selecting patients with high baseline pain intensity; and (2) closer supervision of outpatients>> drug intake and compliance with protocol. Only patients with baseline pain >/=50 on a 100 mm visual analogue scale after wisdom tooth surgery were included. Twenty patients were given paracetamol 1000 mg with or without codeine 60 mg in repeated doses in a randomized and double blind manner. Intake of the first dose of test medication and its effects were closely supervised, while the two following doses were taken at 3-h intervals after the patient had left the clinic. Pain intensity was measured with the visual analogue scale for 8 h. More pain relief was revealed when codeine 60 mg was added to paracetamol 1000 mg on the following measures of effect: change of pain intensity with time (p<0.05, Mann-Whitney), sum of pain intensities (p=0.019), pain intensity difference (p> (p<0.01) and <> (p<0.05) less often and <> more often (p<0.01) than patients with work-related muscular pain in coping with stressful situations in general. No differences were revealed in pain-related coping between the groups. T-anxiety was positively correlated to pain-related <> (p<0.001) and negatively to abilities to control and reduce pain (p<0.05 andp<0.01, respectively). The correlation between general and pain specific coping was weak to moderate. In conclusion, patients with fibromyalgia scored significantly higher on trait-anxiety and seem to interpret stressful situations as more threatening than patients with work-related muscular pain. Anxiety seems to be of central importance for coping with chronic pain. Anxiety prone patients with fibromyalgia might benefit from psychological support in the process of coping with pain. Copyright 1998 The British Infection Society. All rights reserved. PMID- 10700327 TI - The NMDA (N-methyl-D-aspartate) receptor antagonist memantine in the treatment of postherpetic neuralgia: a double-blind, placebo-controlled study. AB - A double-blind, randomized, placebo-controlled trial was conducted to study the analgesic efficacy of the NMDA (N-methyl-D-aspartate) receptor antagonist memantine (1-amino-3,5-dimethyladamantane hydrochloride) in relieving postherpetic neuralgia (PHN). Memantine (or an identical-looking placebon=12/group) was administered at a dose of 10 mg/day for one week, and 20 mg/day for an additional 4 weeks. All patients were required to record their pain level twice daily during the entire study period, with the use of a 0-10 numerical pain scale (NPS). The McGill Pain Questionnaire (MPQ), spontaneous pain, and a series of mechanical and thermal stimuli-induced pain were measured with the use of a 0-100 visual analogue scale (VAS), on six office visits. Quantitative thermal testing (QTT) and routine blood tests were performed at the beginning and at the end of the study. Although reduction in spontaneous pain, mechanical and cold allodynia, mechanical hyperalgesia, and <> like pain were found in both groups, there were no significant differences between memantine and the placebo on any of the outcome measures. No changes were found in either group in MPQ scores or in quantitative thermal thresholds. Although three patients were withdrawn from the memantine group and only one from the control group, no differences in incidence of adverse effects between the two groups were found. Study results show that memantine is ineffective in reducing spontaneous and evoked pain in patients with PHN. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700328 TI - The importance of stimulus configuration for temporal summation of first and second pain to repeated heat stimuli. AB - Temporal summation of pain is suggested to be an important factor during various clinical conditions. Controversies exist as to whether temporal summation exists for Adeltafibre-mediated first pain. The aim of the present human experimental study was to investigate the importance of stimulus configuration (intensity, inter-pulse interval, location) for temporal summation of radiant (laser)- and contact-heat-induced pain. Consecutive stimuli were applied to the same or to adjacent skin locations. Both stimulation techniques evoked rapid temperature changes, which is an important parameter for recruitment of specific cutaneous nociceptors (Adelta and C fibres). Psychophysical thresholds and intensity ratings were used to assess the pain evoked by repeated stimuli applied to the hairy skin of nine volunteers. Inter-pulse interval (IPI) and stimulus intensity were important and interrelated parameters for temporal summation of pain. An increase in IPI resulted in a decreased summation, whereas increased stimulus intensity resulted in increased summation. Brief heat pulses evoked both first and second pain, and summation of the different pain qualities was investigated. Taking the latency from stimulation to perception into consideration, we were able to differentiate and find summation of first (Adeltafibre-mediated) and second pain (C fibre-mediated). Summation of first pain was more pronounced for high (38 degrees C) than for low (30-32 degrees C) baseline temperature. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700329 TI - Interactions in the antinociceptive effect of tramadol in mice: an isobolographic analysis. AB - Tramadol is a widely-used analgesic for pre- and post-operative pain which has a different pharmacological profile to that of classical opioids, since it does not induce respiratory depression, constipation, sedation, tolerance or dependence. However, tramadol frequently produces nausea and vomiting as side-effects. In the present study, the interactions between tramadol and several adrenergic and serotonergic compounds with antinociceptive activity were studied by isobolographic analysis. Antinociceptive activity was evaluated using the acetic acid writhing test in mice. Dose-response curves for the antinociceptive effect of tramadol, prazosin, clonidine, xylamine, clomipramine and cyproheptadine were obtained, and ED(50)s were calculated for isobolographic analysis, which was performed by administration of fixed-ratios of tramadol with each of these drugs, given both systemically and intrathecally. The isobolograms of all combinations tested, either systemically or intrathecally showed superadditivity. The synergies observed with these combinations suggest a complex modulation of the descending noradrenergic and serotonergic systems that exert inhibitory influences on the transmission of nociceptive information, probably in addition to effects on receptors in the primary neurons of the spinal cord. The co administration of analgesic drugs that produce superadditive effects constitutes a significant new avenue for the treatment of pain, since a similar level of antinociception can be obtained with considerable reductions in the dose of each analgesic. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700330 TI - Scoring the mouse formalin test: validation study. AB - The formalin test is a well-established model for assessing nociceptive processes and analgesic drug effects. Previous studies have provided statistical validation of optimal procedures for conducting and scoring the rat formalin test. In the mouse model, formalin concentration has been subjected to validation studies. The present research extended previous work by subjecting two additional parameters - the behaviors that should be scored and the optimal interval for second-phase formalin response - to empirical validation. Five behavioral (formalin-induced favoring, lifting, and biting/licking, rearing and locomotion) and two physiological measures (paw weight and paw thickness increases reflective of formalin-induced inflammation) were examined under four formalin concentrations (Exp. 1: 0.5, 1.0, 5.0, and 10.0%/25ul formalin) or under four morphine doses (Exp. 2: 0.0, 1.0, 5.0, and 10.0 mg/kg, s.c. with 5% formalin concentration). Multiple regression analyses defined the optimal second-phase formalin response in outbred, Swiss-Webster mice as 15-35 min post formalin injection, and revealed that the second-phase response is best characterized by the cumulative time spent biting/licking the injected paw. Finally, paw physiological measures provided convergent evidence of nociceptive and antinociceptive processes in the mouse formalin test. Copyright 1998European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700331 TI - The contribution of spinal neuronal changes to development of prolonged, tonic nociceptive responses of the cat induced by subcutaneous bee venom injection. AB - To elucidate neurophysiological mechanisms of persistent pain induced by tissue injury, the present study was designed to investigate the effects of s.c. bee venom injection on responses of the dorsal horn nociceptive neurons and those of behavior in anesthetized and awake cats, respectively. A parallel comparative study was also performed to compare the effects of s.c. bee venom and formalin injections on neuronal responses by using an extracellular single-unit recording technique. The present results showed that s.c. bee venom injection into the peripheral cutaneous receptive field resulted in a protracted, tonic monophase of increase in spike responses of wide-dynamic-range (WDR) neurons for more than 1 h, while injection of the same volume of vehicle did not have such an effect. The mean number of spikes during the 60-min period after bee venom was 6.74+/-2.58 spikes/s (n=10), which showed a significant increase in firing rate over the background activity (2.23+/-0.96 spikes/s). Behavioral observations showed that s.c. bee venom injection into the dorsum of a hind paw also produced a prolonged, tonic single phase of response indicative of pain, suggesting that central neuronal changes may contribute to development of bee venom-induced prolonged, tonic pain in cats. The increased neuronal firing induced by s.c. bee venom could be suppressed by a single dose of i.v. morphine and resumed by naloxone. Blockade of the sciatic nerve with lidocaine resulted in a complete suppression of the bee venom-induced neuronal firing, suggesting that the central neuronal changes following s.c. bee venom are peripherally-dependent. Comparative studies showed that the duration and frequency of the bee venom-induced neuronal responses were comparable to those induced by s.c. formalin; however, responses of WDR neurons to mechanical stimuli applied to the injection site of the two chemical agents were quite different. Bee venom produced a significant enhancement of mechanical responses of WDR neurons, while, on the contrary, formalin produced a desensitization of sensory receptors in the injection site, suggesting that the two tonic pain models may have different underlying mechanisms. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700332 TI - Functional anatomy of hypnotic analgesia: a PET study of patients with fibromyalgia. AB - Hypnosis is a powerful tool in pain therapy. Attempting to elucidate cerebral mechanisms behind hypnotic analgesia, we measured regional cerebral blood flow with positron emission tomography in patients with fibromyalgia, during hypnotically-induced analgesia and resting wakefulness. The patients experienced less pain during hypnosis than at rest. The cerebral blood-flow was bilaterally increased in the orbitofrontal and subcallosial cingulate cortices, the right thalamus, and the left inferior parietal cortex, and was decreased bilaterally in the cingulate cortex. The observed blood-flow pattern supports notions of a multifactorial nature of hypnotic analgesia, with an interplay between cortical and subcortical brain dynamics. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700333 TI - An investigation of stress and pain perception during manual therapy in asymptomatic subjects. AB - Stress perceived by the patient during the application of a manual therapy treatment technique may account for the reported findings of sympathoexcitation and hypoalgesia immediately after its application. This study investigated whether there was a difference in the level of perceived stress and pain before, during and after the application of a treatment condition (treatment, placebo or control technique), or whether the difference related to the time course of subject involvement in the study.Twenty-four asymptomatic subjects participated in a double-blind, placebo-controlled, within-subjects study in which stress was measured with a stress rating scale and a stress visual analogue scale. Pain experienced by the subject during the application of the treatment technique was measured with a visual analogue scale and modified McGill pain questionnaire. There was no effect of the treatment condition on perceived stress, but there was a reduction in stress levels over the time course of involvement in the experiment, as demonstrated by pre-application levels of stress. Pain was not produced by the treatment technique.This study demonstrated that stress and pain were not features of the lateral glide manipulation of the cervical spine in asymptomatic subjects. If the dorsal periaqueductal grey region is responsible for the initial effects of manual therapy, as has been previously suggested, then stress and pain do not appear to play a role in activating this system during the application of a manual therapy treatment technique in asymptomatic subjects. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700334 TI - Elevated plasma levels of neuropeptide Y in female fibromyalgia patients. AB - Neuropeptide Y(NPY) co-exists with norepinephrine in the sympathetic nervous system, and NPY may represent the sympathetic-neuronal output. Fibromyalgia syndrome (FMS) patients have perturbations in the hypothalmic-pituitary-adrenal (HPA) axis and in the sympathetic stress axis as well. As opioid peptides, monoamines and sex steroids are integrated in the regulation of stress, it is interesting to further explore the role of NPY in FMS patients, as they show many symptoms that are related to perturbations of those systems.In this study, plasma NPY levels were assessed in subgroups of FMS patients: cyclic (regular menstrual cycles), non-cyclic (post-menopausal), depressed and non-depressed patients. In order to examine whether pain and other symptoms seen in FMS patients are correlated to the NPY levels, the patients were also registering 15 different symptoms daily during 28 days. Sex and age-matched healthy controls were recruited for comparisons. Non-parametric tests were used for the statistical analyses.The results showed that the NPY levels were significantly elevated in the patients compared to the controls. In the luteal phase of the cyclic patients, the levels of the peptide were higher than in the corresponding controls. For the non-cyclic patients, there was a positive correlation between physical symptoms and NPY levels, however, pain per se did not reach the significant level of correlation. The non-depressed patients had the same levels of NPY as the depressed FMS patients, who also had a positive correlation between anxiety and NPY levels.These results suggest that FMS patients have an altered activity in the NPY system, most likely due to prolonged and/or repeated stress, and that the hormonal state and time of the menstrual cycle also may be of importance in the complex pathophysiologic mechanism behind the development of FMS. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700335 TI - Quantification and immunocytochemical characteristics of trigeminal ganglion neurons projecting to the cornea: effect of corneal wounding. AB - The number and immunocytochemical characteristics of trigeminal ganglion neurons providing sensory innervation to the cornea were studied in the mouse. Corneal neurons were retrogradely labelled with fluorogold placed on the cornea after removal of the epithelium with n-heptanol. Corneal neurons were counted, sized and characterized immunocytochemically with antisera against substance P (SP), calcitonin gene-related peptide (CGRP), calbindin, calretinin, and with a monoclonal antibody (RT97) against neurofilament proteins. A total of 258 corneal neurons were counted, most of them located in the ophthalmic division of the trigeminal ganglion. They represent only a small fraction (1.3%) of the population of trigeminal ganglion neurons. More than 70% of corneal neurons were classified as 'small dark' according to their cell body area and the absence of immunoreactivity to RT97. A low percentage of corneal neurons, usually large in size, contained calcium binding proteins. Fifty-eight percent of the corneal neurons were immunoreactive to CGRP, and 20% to SP. Corneal wounding with NaOH, which affects stromal nerve trunk, did not modify the total number of corneal neurons or their neuropeptide content. However, this increased the total number of calbindin-positive and decreased the RT97-positive neurons. Thus, unlike in other nociceptive neurons, peripheral axotomy did not modify the SP/CGRP content of corneal neurons.Trigeminal ganglion neurons projecting to the cornea are similar in size and neuropeptide content to nociceptive neurons of other territories. Their number is high in relation to the corneal surface, thus confirming that the cornea has a large nociceptive representation in the trigeminal ganglion. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700336 TI - Pain and mechanical injury of human skin following needle insertions. AB - Needle insertion is a very common invasive procedure, but little has been done to study how pain is related to the mechanical properties of the needle insertion, and how pain from the procedure may be minimized. The aim of the present study was to investigate the pain evoked by standardized needle insertion into the human skin. Needle insertions were performed with continuous registration of the mechanical force on the needle. Forty consecutive needle insertions to the depth of 8 mm with 27G and 30G needles, 2 and 19 mm/s velocities, and at 45 and 90 degree angles to the skin surface were performed in a randomized and blinded design in 30 healthy volunteers. The occurrence, intensity and quality of pain, as well as the maximum mechanical needle penetration force and the total mechanical workload were registered. The mechanical parameters of needle insertions were significantly related to needle size and velocity of insertion. Occurrence of pain and bleeding were significantly related to the needle size. High needle insertion velocity was significantly related to a higher rate of sharp and pricking pain, and low insertion velocity to a significantly higher rate of dull and burning pain. The present study provides quantitative data on pain and mechanical parameters of standardized needle insertions through normal human skin. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700337 TI - PET study on central processing of pain in trigeminal neuropathy. AB - Recent functional brain imaging studies with positron emission tomography (PET), in painful peripheral mononeuropathy and nitroglycerin-provoked cluster headache attacks, suggest a preference of the right hemisphere, especially the anterior cingulate cortex (ACC) and the medial prefrontal cortex (MPFC), in attributing emotional valence and attention to the pain suffering. We have investigated the central processing of painful trigeminal neuropathy (PTN) in patients treated with electric extradural precentral gyrus stimulation (PCGS). Increased regional cerebral blood flow (rCBF) was detected in the right caudal ACC [Brodmann area (BA) 24] and anterior limbic thalamus, while a decreased activity was observed in the right MPFC (BA 9/32) during the habitual-pain state, in comparison with the pain-alleviated state regardless of the inflicted side of PTN. The involvement of BA 9/32 and the anterior limbic thalamus spatially extended to the left hemisphere, but the local maxima and a significant negative correlation between the rCBF changes in the two structures were found only in the right hemisphere. The activation of the caudal BA24 further supports the theory that ACC is crucial for the suffering in chronic pain. Our study not only verifies the preferential role of the right hemisphere in the appreciation of pain suffering, but further supports that sustained chronic pain, being devoid of the motivational component of an escape response, targets the right hemisphere, particularly the BA24 of the ACC. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700338 TI - Recent and forgotten aspects of visceral pain. AB - Progress in the field of visceral pain research has been particularly rapid in recent years. Some aspects of the symptom that had previously been neglected for some time have now received a great deal of attention in both clinical and experimental studies. This regards, in particular, phenomena of hyperalgesia: (a) of visceral structures, because of local inflammatory/sensitizing processes (visceral hyperalgesia); (b) of areas of referred pain from viscera (referred somatic hyperalgesia from viscera); and (c) of a visceral structure, because of an algogenic process of another visceral domain (viscero-visceral hyperalgesia). Clinical studies in patients have led to characterisation of subjective and objective symptoms of these phenomena. A number of studies in human volunteers (employing experimental procedures to stimulate and measure pain reactivity in both visceral structures and somatic areas of referral) have further increased the knowledge about modalities of generation of the various forms of hyperalgesia.Animal experiments have improved understanding of pathophysiological mechanisms, mostly those underlying the referred hyperalgesia, with a number of findings supporting the notion of central changes at the basis of the phenomenon. An important aspect of laboratory experiments in recent years has been the setting up of animal models of visceral pain conditions closely mimicking a number of clinical pain states in patients. As a result, the outcome of experimental studies (electrophysiological, pharmacological, etc.) appears more directly applicable to the interpretation of the clinical reality.Finally, in the context of laboratory studies, a novel trend of investigation is represented by genetic experiments, particularly those employing 'knock-out' mice. These experiments, by generating animals lacking specific genes responsible for the production of various receptors implicated in pain transmission, have further contributed to the understanding of the generation of visceral pain symptoms. Although studies in this field are in their early stage, they seem particularly promising for a better understanding of the pathophysiology of visceral pain, and thus the establishment of more satisfying therapies in the future. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700339 TI - Experimental human muscle pain induced by intramuscular injections of bradykinin, serotonin, and substance P. AB - After intramuscular (m. tibialis anterior) injection of three different algogenic substances, the pain intensity was continuously scored on a visual analogue scale (VAS) in eight volunteers. The subject drew the distribution of the local and referred pain areas on a map. Four times within the first hour after injection, the pressure pain-thresholds (PPTs) and supra pressure-pain thresholds were assessed at the injection point, 2 cm distal from the injection site, at the arm, and at the contralateral leg. Measurements were done before and after injection of 0.5 ml of the algogenic substance [bradykinin (BKN), serotonin (5-HT), substance P (SP)], and isotonic saline as control. Cutaneous sensitivity to mechanical stimuli was assessed with a Von Frey hair at the same location as PPT determinations.The pain intensity (VAS-peak) after BKN (2, 4, and 10 nmol) and 5 HT (2, 4, and 20 nmol) was significantly higher (p< 0.05) than after SP (0.2, 0.4, and 0.8 nmol) and isotonic saline. The VAS-peak after infusions of hypertonic saline was significantly higher (p< 0.05) compared with VAS-peaks after all other substances. A significantly larger (p< 0.05) local pain area was found after BKN compared with isotonic saline. After injections of hypertonic saline, the offsets of evoked pain were significantly longer (p< 0.05) and the local and referred pain areas were significantly larger (p< 0.05) compared with all other substances. There was no dose-response relation between the pain intensity and the different doses of BKN, 5-HT, and SP. PPTs and skin sensitivity were not affected by any of the injections.We conclude that under the present experimental conditions, BKN and 5-HT can produce low levels of muscle pain after intramuscular injection. In the used concentrations, however, BKN, 5-HT, and SP did not generate cutaneous or muscular hyperalgesia. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700340 TI - Cerebral responses to pain in patients suffering acute post-dental extraction pain measured by positron emission tomography (PET). AB - Previous studies with normal volunteers have demonstrated distributed cortical responses to experimental heat pain within a network of structures. The network includes the insula, anterior cingulate, prefrontal, inferior parietal and somatosensory cortices. Patients suffering from chronic nociceptive pain following rheumatoid arthritis (RA) have shown damped central responses to experimental heat pain applied to the back of the right hand. In this study of patients with acute, left-sided, post-molar-extraction (surgical) pain, we assessed the cortical responses to experimental heat pain, applied to the back of the right hand, using positron emission tomography (PET), and compared the responses with a previously reported control group and the RA group. In response to the experimental heat pain, the surgical group indicated significantly increased regional cerebral blood flow in the prefrontal cortex [Brodman's area (BA) 44] ipsilateral to the heat stimulus. Contralateral increases were detected in the putamen and transverse temporal gyrus (BA 40/41/42) with bilateral increases in the insular cortex. Compared to the control and RA group, there were significantly reduced responses in the anterior cingulate (BA 24), pre-frontal medial, and orbito-frontal (BA 9/10/32/47) cortices. These results suggest that relatively discrete regions of the cerebral cortex are responsible for acute nociceptive processing during an acute inflammatory episode. The reduced frontal and anterior cingulate responses to the experimental heat pain (applied to the right hand) during acute inflammatory pain (left jaw) illustrates cortical modulation of nociceptive processing that may be related to non-somatotopic, bilateral, nociceptive inputs to these areas. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700341 TI - Effects of sciatic nerve injuries on delta -opioid receptor and substance P immunoreactivities in the superficial dorsal horn of the rat. AB - The aim of the present study was to examine the effects of transection combined with tight ligation, and crush of the sciatic nerve on delta -opioid receptor and substance P immunoreactivities in the superficial spinal dorsal horn at different time points after injury. Both the delta -opioid receptor and substance P are primarily localised to primary afferent fibres and terminals. Seven days following transection and ligation, a slight decrease in both delta -opioid receptor and substance P levels was seen in laminae I and II. The maximal reduction appeared to take place around 4 weeks. Restoration of immunoreactivity was observed by 32 weeks, and by 1 year the levels were almost back to normal. Regarding crush injury, the reduction in both delta -opioid receptor and substance P immunoreactivities were less pronounced and recovery was faster than after transection injury. Already by 16 weeks, the levels were almost back to normal.These results show that peripheral nerve injuries dramatically reduce the levels of delta -opioid receptor and substance P immunoreactivities in the superficial dorsal horn after short survivals and demonstrate recovery after long survivals. Whether the marked reduction of delta -opioid receptors in the dorsal horn is involved in the decreased ability of opioid analgesics to alleviate neuropathic pain remains to be studied. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700342 TI - Time course of UVA- and UVB-induced inflammation and hyperalgesia in human skin. AB - Dose-dependency and time course of hyperalgesia and erythema following UVA (16.8 and 36 J/cm(2)) and UVB (one and three times the minimum erythema threshold) irradiation was investigated in 10 healthy human subjects. Skin patches (1.5 cm in diameter) on the ventral side of the upper leg were irradiated with UVA or UVB light. Hyperaemia (Laser Doppler flowmetry, infrared thermography), thermal hyperalgesia to radiant heat stimuli, and mechanical hyperalgesia to controlled impact stimuli were tested at 1, 6, 12, 24, 48 and 96 h after irradiation. Dose dependent delayed hyperaemia and hyperalgesia was induced only by UVB irradiation. UVB-induced increase in blood flow peaked at 12 h after irradiation and normalized by 96 h. Although superficial blood flow, as measured by Laser Doppler flowmetry, increased up to eight-fold, no significant increase of skin temperature was detected using infrared thermography. Development of mechanical and thermal hyperalgesia was delayed and reached a plateau between 24 and 48 h. In contrast to UVB, UVA irradiation of up to 36 J/cm(2), sufficient to produce intense tanning of the skin, did not induce significant hyperalgesia or delayed hyperaemia. It is concluded that UVB- but not UVA-irradiation is a suitable experimental model of subacute thermal and mechanical hyperalgesia. The different time courses of erythema and hyperalgesia indicate that inflammatory mediators responsible for vasodilatation are not identical with those inducing hyperalgesia. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700343 TI - Cyclophosphamide cystitis in mice: behavioural characterisation and correlation with bladder inflammation. AB - The generation of transgenic 'knock-out' mice which lack genes relevant to pain is becoming increasing common. However, only one visceral pain model, the writhing test, is available in mice. The aim of this study was to adapt cyclophosphamide cystitis, a model of inflammatory visceral pain described in rats, for use in mice, and to characterise its behavioural effects. The toxic metabolites of systemically-administered cyclophosphamide are excreted in the urine, and induce bladder inflammation. We compared the effects of cyclophosphamide (100 and 300 mg/kg i.p., 4 h survival period) and vehicle (saline) in male mice on spontaneous behaviour (4 h continuous video-tape, and a 5-min Open Field test after 4 h). Involvement of the urinary bladder and other abdominal tissues was assessed by macroscopic examination and measurement of Evan's Blue plasma extravasation. Cyclophosphamide (300 mg/kg) produced significant changes in behaviour, including 22 +/- 6 min of 'crises' of visceral pain-related behaviour and a 53% reduction in activity, and also induced haemorrhage and substantial plasma extravasation in the bladder, but no change in other abdominal tissues. We conclude that cyclophosphamide cystitis has many advantages as a model of sub-acute, inflammatory visceral pain in mice. It does not require surgery or intubation, and we have found it to produce consistent, reproducible and quantifiable behavioural changes, which are significantly correlated with the degree of bladder inflammation in the absence of inflammation of other abdominal tissues. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700344 TI - Pressure pain thresholds asymmetry in left- and right-handers: Associations with behavioural measures of cerebral laterality. AB - Pressure pain threshold (PPT) asymmetry of the left and right third digits was assessed in 12 right-handed and 12 left-handed subjects using an automatised pressure algometer. A clear PPT asymmetry was found in right-handed participants, while left-handed participants revealed no PPT asymmetry. The PPT asymmetry of right-handed participants was due to a reduced PPT or increased pain sensitivity at the left hand. Behavioural laterality tests revealed a right ear or left hemisphere advantage for the processing of verbal material (consonant-vocal syllables) and a left visual field or right hemisphere advantage for the processing of emotional faces in all participants. PPT asymmetry was not associated with cerebral laterality assessed with these tests. We conclude that PPT asymmetry is associated with handedness, but neither PPT asymmetry nor handedness are closely associated with measures of cerebral laterality. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700345 TI - Pain in ambulatory HIV-infected patients with and without intravenous drug use. AB - The prevalence of pain in 211 HIV-infected patients with and without intravenous drug use was assessed and the prognostic information inherent in pain reporting was evaluated, using a questionnaire on pain and HIV-related symptoms combined with data on disease classification, route of HIV transmission, CD4+ lymphocyte counts in blood (CD4) and mortality rates at 15 months after completing the questionnaire. The pain prevalence was significantly higher among intravenous drug users (IDUs) compared with non-IDUs [76/89 (85%) vs 87/122 (71%);p<0.05], especially among the patients classified as asymptomatic [43/53 (81%) vs 35/59 (59%);p = 0.01]. No significant difference was found among AIDS patients. In non IDUs, a strong correlation was found between HIV disease stages according to the Centers for Disease Control classification (CDC) and pain prevalence (CDC A: 59%vs B: 74%vs C: 96%, p<0.001), and between the number of concurrent pain sites and both the CD4 levels (no pains: CD4 0.26 x 10(9)/l vs 1-2 pain sites: CD4 0.22 vs>2 pain sites: CD4 0.09;r = 0.35, p<0.001), and the mortality rate [no pains: 2/35 (6%) vs 1-2 pain sites: 8/45 (18%) vs> 2 pain sites: 12/42 (29%), p<0.01]. In IDUs, no such correlations were found. Our data demonstrates differences in the development, prevalence and prognostic value of pain among HIV-infected patients, with and without intravenous drug use, clearly indicating the need to differentiate risk groups in pain related studies. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700346 TI - The development of the DEGR(R): A scale to assess pain in young children with cancer. AB - The Gustave Roussy Child Pain Scale (Douleur Enfant Gustave Roussy, DEGR(R)Scale) is a scale for grading prolonged pain in children aged 2-6 years with cancer. The scale comprised six behaviours specific to pain items, five psychomotor inertia items, and four anxiety items, with a total score ranging from 0 to 60. This work was designed to confirm the scale structure and to study its construct validity and convergent validity.Our work was composed of two parts. In the first part of the study, 152 children with progressive cancer were scored by two nurses using the DEGR(R)scale, in a cross-sectional design. And in the second part, 53 of these 152 children were video-recorded. The tapes were assessed both by a panel of four pain specialists using a 0 to 7 Likert scale and by a nurse using DEGR(R)scale.As for the 152 children, the mean of the total scores derived from the DEGR(R)is 20.2 (SD = 6.2). Both the degree of agreement between the nurses (the weighted kappa coefficient) and the internal consistency of the scale (Cronbach alpha coefficient = 0.90) were high, providing evidence of good reliability. Multivariate factor analyses showed a first factor of intensity of pain (51% of the total variance) and a second factor (14% of the total variance) which distinguishes the psychomotor inertia items from the items concerning voluntary expression of pain. Also, the results showed that psychomotor inertia items contribute to both factors and that it is an important sign of prolonged pain. Construct validity was strengthened by the absence of correlation between DEGR(R)scores and variables not related to pain, including fever, neutropenia and anaemia (indicative of poor medical condition) and the absence of parents' visits (indicative of psychological distress).Concerning the 53 video-recorded children, the nurses' DEGR(R)ratings were strongly correlated with the specialists panel scores indicating a fairly good case for convergent validity. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700347 TI - Differential effects of morphine on pain and temperature perception in human volunteers. AB - Electrophysiological and behavioral evidence suggests that morphine may have a differential effect on nociceptive and thermal pathways. In this study, we explored the perceptual consequences of these differential actions by examining the effect of a low morphine dose (0.08 mg/kg) on pain and temperature sensations arising from cutaneous thermal stimuli. In a double-blind placebo-controled study, we compared the perceived temperature intensity and perceived pain intensity and unpleasantness of noxious and innocuous heat and cold applied to the face of human subjects, with and without low doses of systemic morphine. The results showed that morphine modified pain-related sensations. In contrast, perceived thermal intensity of both noxious and innocuous heat or cold stimuli was unchanged by low-dose morphine administration. These findings suggest that low doses of morphine have a differential effect on pain and temperature sensations arising from the same stimulus, and thus that these sensations could be subserved by different neuronal populations. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700348 TI - Patient-controlled extradural analgesia after caesarean section: a comparison between tramadol, sufentanil and a mixture of both. AB - In a double-blind randomised study into post-operative pain relief by extradural PCA, 66 Caesarean section patients were divided in to three groups to receive either sufentanil (2 u g/ml), tramadol (10 mg/ml) or a mixture of both. After a loading dose of 10 ml, patients were allowed to ask for additional boluses of 2.5 ml, respecting a lock-out time of 10 min and a 1-h limit of 10 ml. Every 6 h, VAS pain scores, consumption of drugs, number of demands and side-effects were registered.At 6 h, pain was significantly less in the combination group compared to patients receiving tramadol alone. The 24-h dose requirements for sufentanil and tramadol when used alone were 123.5+/-10.3 u g and 652+/-42 mg, respectively. Combining both drugs decreased the consumption and number of demands for tramadol only (22%, p<0.05 vs 18% for sufentanil, NS). In the tramadol groups, more failures and significantly more side-effects, mainly nausea and vomiting, were noticed.It may be concluded that the extradural use of tramadol is less beneficial than previously reported. Due to disturbing side-effects, relatively high dose requirements (even after the addition of a lipophilic opioid) and somewhat inferior analgesic quality, its extradural administration for postoperative pain relief cannot be recommended. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700349 TI - Assessing surgical patients' expectations and subsequent perceptions of pain in the context of exploring the effects of preparatory information: raising issues of gender and status. AB - The present study explored both patients' expectations and experiences of pain in the context of utilizing an established research procedure, which previously reported a significant effect of instructional information on post-surgical recovery. No research to date has systematically attempted to identify pre operative expectations of post-operative pain, nor has it compared pre- and post operative ratings. A further variable scrutinized during the study was patient gender.Thirty-two males and 31 females undergoing two types of elective surgery were randomly allocated into experimental and control conditions by a third party. Patients in the experimental group received additional instruction information pre-operatively from the anaesthetist and the control group received the routine pre-operative visit. Using a numerical pain scale, all participants provided ratings of expected pain levels and subsequently rated their pain following surgery.Results obtained were contrary to expectations with respect to the effects observed by the established study as instructional information had no significant effect on recovery variables. However, the study did suggest that pre and post-operative pain ratings were broadly equivalent. A significant gender difference was also found with male patients expecting less pain than they perceived and an opposite pattern for females. Measuring both pain expectations and perceptions is considered a useful technique to inform pain management and is worthy of further investigation. The inconsistent effect of instructional information and the gendered rating patterns are considered in relation to gender and status differentials between health providers and have important implications for future research. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700350 TI - Subjective future as a mediating factor in the relation between pain, pain related distress and depression. AB - The coincidence of chronic pain, psychological distress and depression has been well documented in several studies. However, there is still debate about the type of causality linking these factors and whether psychological distress and depression precede or are a consequence of pain. This study contributes to this debate through an analysis of the latent structure behind these complex concepts. To test the hypothesis that subjective future (i.e. how the pain patient perceives the future) has an impact on pain, data were analysed from 660 chronic pain patients who were tested with The Multidimensional Pain Inventory (MPI), The Symptom Distress Checklist (SCL-90) and a Future Scale, which was constructed from items of the Sense of coherence-scale. By use of path analysis and structural equation modeling (S.E.M.) four latent constructs were tested: Pain, Interference, Distress and Subjective future. The results indicated that Subjective future has a strong impact on Distress, is a mediating variable, which contributes to conceptually explaining and practically nullifying the relationship between Pain and Distress, and finally that Pain is a concept, that changes with increasing duration. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700351 TI - Connective tissue massage in the treatment of fibromyalgia. AB - The aim of this study was to investigate the effect of connective tissue massage in the treatment of individuals with fibromyalgia. The results of this random study of 48 individuals diagnosed with fibromyalgia (23 in the treatment group and 25 in the reference group) show that a series of 15 treatments with connective tissue massage conveys a pain relieving effect of 37%, reduces depression and the use of analgesics, and positively effects quality of life. The treatment effects appeared gradually during the 10-week treatment period. Three months after the treatment period about 30% of the pain relieving effect was gone, and 6 months after the treatment period pain was back to about 90% of the basic value. As long as there is a lack of effective medical treatment for individuals with fibromyalgia, they ought to be offered treatments with connective tissue massage. However, further studies are needed in the mechanisms behind the treatment effects. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700352 TI - Massage-is it really a reliable method of treatment? PMID- 10700353 TI - Pain and inflammatory hyperalgesia induced by intradermal injections of human platelets and leukocytes. AB - Acute and prolonged inflammatory processes, induced by intradermal injections of autologous platelet preparations, leukocytes, platelet-leukocyte mixtures and vehicle solutions, were monitored in 20 human subjects for up to 48 h. Psychophysical methods were employed to assess the time course of pain response, and the development of hyperalgesia. Time course of the axon reflex erythema was analysed by computer based videography. Injections of functionally intact platelets [4.4 x 10(7)+/- 0.6 x 10(7)in 50 ul (mean +/- SEM)] caused distinct pain sensations [median pain rating 7.25 (25-75%: 6.5-7.5) on a scale form 0-10]. Both platelets injected through a microfilter (0.2 um) and heat-inactivated platelets induced similar initial pain response; however, pain duration was significantly shorter than for intact platelets. Control injections of leukocytes (5.4 x 10(6)+/- 3.7 x 10(6)in 50 ul) and of vehicle solutions only caused minute pain sensations.Although platelet concentration in the mixture was only 50%, platelet/leukocyte mixtures produced axon reflex erythema that were significantly larger than those seen with naive platelets alone (11.1 +/- 2.3 vs 8.3 +/- 0.8 cm(2)). Intact platelets and heat-inactivated platelets both induced mechanical hyperalgesia and skin indurations at the injection site. Both reactions peaked between 6 and 24 h after the injection and disappeared within 48 h. Neither hyperalgesia, nor the development of indurations was noted at the injection sites of filtered platelets and vehicle solutions. Leukocyte injections induced hyperalgesia and indurations only in samples with a platelet contamination of more than 50 000 cells/ul. We conclude that platelets induce acute pain responses due to a soluble factor, whereas the development of hyperalgesia and the formation of indurations depends on a membrane-bound factor. Moreover, platelets and leukocytes act synergistically in nociceptor activation. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700354 TI - A psychometric evaluation of the Swedish version of the Multidimensional Pain Inventory (MPI-S): a gender differentiated evaluation. AB - A need to consider possible gender differences in pain research has been recognized by researchers during the last decades. As part of a psychometric evaluation of the Swedish version of the Multidimensional Pain Inventory (MPI-S), we performed gender-differentiated analyses of the internal consistency, validity and sensitivity to change of the MPI-S in a sample of 235 individuals (129 females, 106 males) suffering from long-term non-specific pain from the lower back and/or neck region. The construct validation and sensitivity analyses were performed by using validated self-report measures and direct observational assessment techniques as external constructs. For sections 1 and 2 of the MPI-S, the results support the internal consistency (alpha coefficients ranged from 0.74 to 0.85 for females and 0.62 to 0.89 for males) and construct validity across gender. The General Activity (GA) scale of section 3 of the MPI-S displayed acceptable internal consistency across gender (alpha = 0.79 for females, 0.80 for males) but not a satisfactory construct validity. Furthermore, the results yielded some support for the sensitivity to change of the Pain Severity (PS), Interference (I), Life Control (LC) and Affective Distress (AD) scales (from section 1) across gender. Unfortunately, the GA scale did not display a satisfactory sensitivity either for females or males. Altogether, the results showed a similar pattern across gender, although some divergences were detected, such as the substantially weaker negative correlation between perceived supportive behaviour from significant others and punishing responses for males compared to females. In conclusion, we recommend the use of sections 1 and 2 of the MPI-S as a psychometrically evaluated and comprehensive instrument in the assessment of individuals suffering from chronic non-specific low back pain or neck pain. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700355 TI - A survey of postoperative pain treatment in children of 3-14 years. AB - There is a lack of information concerning the characteristics of pediatric postoperative pain in Southern European countries. The aim of this study was to document how postoperative pain in children was managed routinely at Spanish surgical wards.The study was carried out in three hospitals on the first postoperative day. Children were divided in four groups according to their age (years): Group I (3-5), II (6-8), III (9-11) and IV (12-14). The parameters evaluated were: analgesia characteristics (type of prescription, drug used and route of administration, prescribed dose and whether the drug was or was not administered, need of non-prescribed analgesics) and the postoperative pain intensity. The results were analysed using descriptive statistics. U-Mann Whitney, chi(2), ANOVA, Kruskall-Wallis and Student's t -test were also used.A total of 348 children ranging from 3 to 14 years were studied. The average age (+/- SD) was 8.2 +/- 3.3 and the majority were male (74%). Urologic surgery was the most frequent type of operation, with age (p<0.05) and hospital differences (p<0.001). The majority of the patients (52%) were prescribed an analgesic, but only 26% of them had an analgesia order at fixed dosage intervals. Differences among the hospitals were observed (p<0.001). The most commonly used analgesics were metamizol, propyphenazone, paracetamol and codeine. Differences in choice of drug in relation to age and hospital were significant (p< 0.001). Rectal was the preferred route of drug administration. Patient's age was unrelated with the prescribed analgesic dose. An average of 68% of prescriptions were given and half of the patients without scheduled analgesia needed to have analgesics administered. Around 20% of patients had high pain scores.Few paediatric patients are given analgesics at fixed dose intervals to treat postoperative pain. Pain relief therapy for children differs notably to that of adults, in respect to the drugs prescribed and the administered route. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700356 TI - Capillary blood sampling: relation between lancet diameter, lancing pain and blood volume. AB - The relation between pain of capillary blood sampling and lancet diameter was studied in 52 healthy subjects. The tips of six fingers were pricked with a Softclix(R) II lancing device using lancets with identical facet geometry but needle diameters of 0.3, 0.4 and 0.8 mm. Two penetration depths (0.9 and 1.2 mm) were applied. Pain intensity and blood volume were recorded.At a puncture depth of 0.9 mm, pain did not differ between the three lancets. Pain increased with penetration depth, and at 1.2 mm the thicker lancets were somewhat more painful than the thinner ones. Blood volumes increased with lancet diameter and penetration depth. If the pain for punctures providing just enough blood for a glucose test (>/=10 ul) was compared, there was no difference between the lancets.It can be concluded that lancet diameter is of minor importance for the patient. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700357 TI - Electric stimulation of the cingulum bundle precipitates onset of autotomy induced by inflammation in rat. AB - The purpose of this work was to test if electric stimulation of the cingulum bundle in animals subjected to a hindpaw inflammatory process precipitates the onset and enhances autotomy behaviour. Wistar rats were implanted with bipolar parallel electrodes in the boundary of the cingulum bundle. The inflammatory process was induced in all subjects by injection of carrageenan. The groups were: A, sham; B, implanted and stimulated 10 min daily for 7 days; C, implanted and stimulated 2 h daily, for 7 days. Both groups were injected with CAR 2 days after ending the stimulation period; and D, implanted and stimulated 10 min daily for 5 days, the first stimulation being simultaneous to CAR injection. Results show that 100% of the subjects in stimulated groups presented autotomy as compared with 66% in the sham group. A significant shortening of the onset and increased rates in autotomy were observed in experimental groups (B, C and D) as compared to the sham group. We did not find differences between groups B and C, but there was an increment of autotomy in group D when compared with both B and C groups. We conclude that it is possible to facilitate the onset and to increase the intensity of the autotomy triggered by the inflammatory process with cingulum bundle electrical stimulation. The results also suggest that a fundamental condition to the development of the autotomy in this model is the presence of the noxious inflammatory process. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700358 TI - A comparison of the Faces Pain Scale and the Facial Affective Scale for children's estimates of the intensity and unpleasantness of needle pain during blood sampling. AB - To what degree can facial expression scales help children differentiate between the sensory and emotional aspects of the pain experience? This study examined the relationship between children's ratings on the Faces Pain Scale (an intensity measure), the Facial Affective Scale (an affective measure), and a paired mechanical visual analogue (MVAS) method for measuring the intensity and unpleasantness of pain. It was predicted that ratings on the Faces Pain Scale should correlate best with the MVAS measure of pain intensity rather than unpleasantness. Likewise, ratings on the Facial Affective Scale should correlate best with the MVAS measure of pain unpleasantness (assumed to reflect an emotional dimension) rather than intensity. Eighty children scheduled for blood sampling were selected in two age groups: 4 to 6, and 7 to 10 years. Children rated needle pain using each pain scale. As hypothesized, ratings on the Faces Pain Scale correlated more highly with the MVAS ratings for intensity (r =0.77) than for unpleasantness (r =0.52). A smaller reverse finding was confirmed for the Facial Affective Scale which correlated more highly with the MVAS for unpleasantness (r =0.64) than for intensity (r =0.51). Factor analysis indicated that 'pain dimension' (intensity vs affect) was a relatively weak factor as compared with shared instrument variance (two MVAS vs two face scales). No systematic age effects were observed. In conclusion, the Faces Pain Scale and the Facial Affective Scale may partly measure different aspects of the pain experience in children, although it remains to be determined to what degree the obtained differences are clinically meaningful. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700359 TI - Conditioned stress-induced analgesia in humans. AB - The purpose of this experiment was the demonstration of conditioned stress induced analgesia in humans. Following a baseline day (day 1), two groups of 10 subjects were each exposed to green light as the conditioned stimulus (CS) and mental arithmetic plus noise as the unconditioned stimulus (US) for 5 days (days 2-6), a third group of 10 subjects was only exposed to the US. On the test day (day 7), pain threshold and pain tolerance were tested, while groups 1 and 3 received the green light CS and group 2 received no CS. Group 1 showed a significantly higher pain threshold and higher pain tolerance on the test day, whereas the two control groups did not have altered pain perception. Heart rate data confirmed successful stress induction. Heart rate and blood pressure were unaltered on the test day, thus making cardiovascular mediation of the conditioning effect unlikely. Conditioned stress analgesia might be useful in the understanding of chronic pain-syndromes. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700360 TI - Impaired selective attention in chronic pain patients. AB - Several recent studies have suggested that cognitive complaints among chronic pain patients could result from an interference between ongoing pain and mental tasks, as they share common and limited attentional resources. This study was intended to further explore that presumed relationship between chronic pain and attentional disorders. For this purpose, a more sensitive version of the conventional colour-word Stroop task, with four subtasks of increasing difficulty, was used in a group of 33 consecutive patients with chronic non malignant pain and a group of 20 healthy subjects as controls. This task assesses specifically selective attention. Since the Stroop task is sensitive to dysthymic states, levels of anxiety and depression were also assessed in chronic pain patients.As expected, the increase of response times was positively related to the difficulty of the subtasks. However, only the patients with chronic pain of high intensity presented a significant increase in the response times to each subtask as compared to controls. Response accuracy was not affected. Patients with high pain had higher scores on trait-anxiety than those with low-intensity pain. However, ANCOVA showed that trait-anxiety scores x pain groups interactions were not significant and thus that trait-anxiety was not useful for predicting task performance.These results point to a disturbance of selective attention as a function of pain intensity in chronic pain patients. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700361 TI - The lifetime occurrence of Herpes zoster and prevalence of post-herpetic neuralgia: A retrospective survey in an elderly population. AB - One-thousand-and-seventy-one randomly chosen elderly persons (537 women, 534 men; median age 80) were recruited from the Institute of Human Aging (Dept of Psychiatry, University of Liverpool). Almost a quarter (23.8%; equal numbers of both sexes) had had shingles (HZ), at a median age of 60 (for both sexes); 39 subjects (3.6% of all respondents, 15% of those who had had shingles), two thirds of whom were female, developed post-herpetic neuralgia (PHN), defined as pain persisting for more than 3 months; they acquired HZ at a median age of 70. In 22 of them, pain had resolved by the time they were questioned, but in 17 it was ongoing (from less than 12 to 504 months). Two new independent risk factors for PHN were identified: (1) female gender; and (2) living alone at the time of HZ acquisition (p = 0.009). In addition to confirming the well-known factor of: (3) age at shingles acquisition (up to the early 90s); and (4) scarring, presumed to be a consequence of rash severity, was significantly commoner in subjects whose HZ was followed by PHN.Extrapolating the prevalence figures to the whole UK population, of whom 9.28 million were over 64 in 1992, it can be conservatively estimated that at any one time, some 200 000 people in the UK have PHN. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700362 TI - Principles of economic evaluation for interventions of chronic musculoskeletal pain. AB - Economic evaluation is attracting increasing attention to inform policy makers, insurers and other payers of the value of existing and new treatment modalities. Hence, it is desirable to assess not only the medical but also the economic consequences the new treatments produce. The available literature on economic evaluation revealed an urgent need for sound economic evaluation studies in the field of chronic musculoskeletal pain. Due to the generally weak methodology, the intended purpose of economic evaluation to help set funding priorities has often been bypassed. Although in general therapists have no direct responsibility for allocating scarce resources in the field of musculoskeletal pain, they are confronted with the results of these decisions in their everyday work. A clear understanding of the main principles of economic evaluation studies might therefore be advantageous. This paper addresses important methodological issues in economic evaluation research, such as the techniques for economic evaluation studies and the analytic perspective. In addition, the paper pays attention to the inclusion of costs and outcomes in economic evaluation research, sensitivity analysis, discounting, incremental analysis and ratios, and collecting of data.Further emphasis is placed on the transparent reporting of methods and study results. A clear reporting may help therapists and other researchers interpret the results of published studies and apply them to their own studies, and it may help decision makers generalize results from one setting to another. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700363 TI - Behavioural effects of LTP-inducing sciatic nerve stimulation in the rat. AB - A short-lasting tetanic sciatic nerve stimulation that previously has been shown to be nociceptive only during the stimulation, induces long-term potentiation (LTP) of nociceptive evoked responses in wide dynamic range neurons in the dorsal horn of rats. The LTP may contribute to the process of central sensitization. We have shown that the tetanic conditioning stimulation with muscular contractions induces LTP of both Abeta- and C-fibre evoked responses. However, the same stimulation during muscular paralysis induces LTP only of C-fibre evoked responses. In the present study, we investigated the effects of this conditioning stimulation with or without muscular paralysis in behavioural tests in rats. Conditioning stimulation with muscular contractions caused a significant reduction of weight borne on the stimulated side, suggesting muscular soreness and peripheral sensitization. Conditioning stimulation during neuromuscular paralysis, which only has given LTP of C-fibre evoked responses in intact animals, caused no change in the weight borne on the stimulated side, suggesting less or even absence of allodynia. However, in these animals the response temperature in the hot plate test was increased both on the stimulated and on the contralateral side compared to sham-operated rats. In view of our recent results indicating that a descending inhibition reduces the expression of LTP in dorsal horn cells, and the suggestion by others that long-term descending inhibition may override a segmental facilitation, it is suggested that an increased long-lasting endogenous nociceptive inhibition is induced after LTP-inducing stimulation. This is an interesting parallel to stimulation-induced analgesia in humans. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700364 TI - Opposite effects of spinal cord stimulation in different phases of carrageenan induced hyperalgesia. AB - A possibly beneficial effect of spinal cord stimulation (SCS) on acute and chronic nociceptive pain is still a controversial issue. In the present experimental study, SCS was applied, with parameters similar to those used clinically, to awake but restrained rats submitted to intraplantar injection of carrageenan (CAR). The paw circumference and the threshold of paw withdrawal and/or vocalization to pressure, applied by a pair of strain gauge calibrated flat forceps to the paw, were determined before and after CAR injection. As controls, one group of rats was treated by CAR injection only; a second group was subjected to SCS after CAR injection; a third group was pretreated with SCS for 3 days before the injection; a fourth group subjected to SCS only. In the acute phase, 3 h after the injection, SCS enhanced the CAR-induced hyperalgesia and oedema. Conversely, in the subacute phase at day 3-5 after the injection, SCS suppressed the hyperalgesia which was, however, less marked than in the acute phase. However, in that phase, the oedema was still increased by SCS. Pretreatment by SCS influenced neither the natural time course of hyperalgesia nor that of oedema. Circumstantial evidence suggests that the oedema augmentation by SCS in the acute phase may be the result of enhanced vasodilatation with fluid extravasation, probably associated with peripheral release of several substances, such as CGRP. The concomitant increase of the hyperalgesia may reflect an enhanced release of substance P and 5-HT, both peripherally and in the dorsal horn. The attenuation of hyperalgesia in the subacute phase may be due to an inhibitory effect of SCS on A-fibre-mediated wide-dynamic-range neuronal hyperactivity, presumably involving GABAergic mechanisms. However, a direct suppressive effect on sensitized nociceptive-specific second-order neurons cannot be excluded. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700365 TI - Thermal and tactile perception thresholds in acute herpes zoster. AB - This study was conducted to determine somatosensory perception thresholds in 97 immunocompetent patients with herpes zoster (HZ), and to evaluate their associations with the development of post-herpetic neuralgia (PHN). Warm, cold and heat pain thresholds were tested by Thermotest (SOMEDIC) and tactile thresholds by Semmes-Weinstein monofilaments. To establish reference values, 103 healthy subjects underwent somatosensory testing, from which values were calculated for both genders for four age groups (<60, 60-69, 70-79 and >/=80years) in five dermatomal levels (VI, C3, T3, T10 and S1). Patients with HZ underwent quantitative somatosensory testing within the affected dermatome, its mirror image dermatome and an adjacent dermatome bilaterally. The follow-up visits with somatosensory testing took place at 2 weeks, 6 weeks, 3 months and 6 months. When evaluated as means of the results, warm and cold thresholds were significantly elevated in the affected dermatome from the initial visit until 3 and 6 months, respectively. By contrast, heat pain thresholds were lowered at the initial visit but normalized by 2 weeks, and tactile thresholds remained unchanged. These threshold changes were associated neither with further development of PHN nor each other. It is concluded that measurement of somatosensory perception thresholds in early stages of HZ shows evidence of impaired neural function but is not helpful in predicting which patient will go on to develop PHN. Copyright 1999 European Federation of Chapters of the International Association for the Study of Pain. PMID- 10700366 TI - Tutorial II: Neuromodulation of Pain-Abstracts from a consensus meeting in Brussels, 16-18 January 1998. PMID- 10700367 TI - Adolescent participation in the U.K. national lottery games. AB - This study investigated illegal participation in the two U.K. National Lottery games (on-line game and scratchcards) by children under the age of 16 years. The sample consisted of 256 children aged between 13 and 15 years from four mixed-sex comprehensive schools/colleges, which serve city, town, rural and coastal catchment areas in the county of Devon, U.K. Pupils completed a questionnaire in a controlled environment at their respective schools/colleges. The findings indicated that 56% of the sample had participated in the National Lottery on-line game and 54% in the National Lottery Instants scratchcards. Regression analysis revealed that the best predictors of participation in the on-line game were income, household participation, whether the TV show was watched and whether a retailer had ever refused to sell a child a lottery ticket. The same variables (minus watching of the TV show) were also the best predictors of buying scratchcards. PMID- 10700368 TI - Deliberate self-poisoning in adolescence: why does a brief family intervention work in some cases and not others? AB - In a randomized trial of a brief family intervention with adolescents who had deliberately poisoned themselves, we have previously reported that, within the group of patients who did not have major depression, the family intervention was significantly superior to routine care in reducing suicidal thinking. The present paper examined whether efficacy was related to changes in family functioning or other possible mediating variables. Potential mediators included family functioning, hopelessness, depression, adolescent problem-solving and compliance with treatment. The efficacy of the family intervention in reducing suicidal ideation within the non-depressed sub-group was probably not mediated by changes in these variables. The implications of this finding are discussed. PMID- 10700369 TI - The effects of an anti-bullying intervention programme on peers' attitudes and behaviour. AB - This study aimed to evaluate the effect of an anti-bullying intervention programme on peers' attitudes towards bullying and their attempts to solve bully/victim conflicts. An experimental pre-test/post-test design with a control group was used. For secondary school students, positive outcomes were observed at post-test 1 on attitudes and on actual rates of intervention. However, the effects had disappeared at post-test 2. For primary school students, significant differences were found at post-test 2, showing a smaller decline in seeking teacher's help and in heightening students' support for victims. Students' competence to solve bully/victim problems in relation to their general pro-social development is discussed. PMID- 10700370 TI - Impact of attitudes and suicidal ideation on adolescents' intentions to seek professional psychological help. AB - Few adolescents who experience significant psychological distress seek professional psychological help, a finding particularly pertinent in New Zealand which has one of the highest youth suicide rates in the world. In the present study, 221 New Zealand high school students completed a questionnaire which examined the relationship between a variety of approach and avoidance factors associated with professional psychological help-seeking. Suicidal ideation, attitudes, psychological distress, treatment fears, gender and prior help-seeking were significant predictors accounting for approximately 23% of students', self rated help-seeking intentions. Contrary to expectations, higher levels of suicidal ideation led to lower levels of help-seeking intentions for suicidal thoughts. This findings is discussed in relation to the process of help-negation which has been identified in clinical suicidal samples. The implications of these findings for interventions that increase appropriate professional psychological help-seeking in adolescents are also discussed. PMID- 10700371 TI - Deliberate self-harm in adolescents in Oxford, 1985-1995. AB - Deliberate self-harm (DSH) has been one of the major health problems of adolescents in the U.K. for many years. Any changes in rates of DSH or the demographic characteristics of the patient population are likely to have important implications for clinical services and for future suicidal behaviour. Following a decline in rates in the late 1970s and mid 1980s, there were signs in the late 1980s that rates were beginning to increase again. We have used data collected by the Oxford Monitoring System for Attempted Suicide on the basis of patients presenting to the general hospital in Oxford to review trends in DSH in under 20-year-olds between 1985 and 1995. There was a substantial increase in the numbers of teenage DSH patients during the 11-year study period, with an increase between 1985-1986 and 1994-1995 of 27.7% in males, 28.3% in females, and 28.1% overall. There were no demographic changes within the catchment area to explain a change of this size. As rates of repetition of DSH also increased in both sexes during the study period the overall number of episodes of DSH rose even more between 1985-1986 and 1994-1995 (+56.9% in males, +46.3% in females, and +49.4% overall). As in previous studies the majority of adolescents had interpersonal problems and/or difficulties with studying or employment. Self-poisoning with paracetamol and paracetamol compounds became increasingly common such that by 1995 these were used in almost two-thirds of overdoses. The recent increase in DSH in adolescents has important implications for general hospital and adolescent psychiatric services. The greater frequency of repetition of DSH may herald increased future suicide rates. The case for restricting the amount of paracetamol available is overwhelming. Evaluative trials of specific interventions following adolescent DSH are urgently required. PMID- 10700372 TI - Effects of peer victimization in schools and perceived social support on adolescent well-being. AB - It has been suggested that the mental health of schoolchildren can be undermined by repeated bullying at school and further exacerbated by having inadequate social support. To evaluate this claim, the General Health Questionnaire (GHQ) was administered anonymously to 845 adolescent schoolchildren attending coeducational secondary schools in South Australia, together with measures of the extent to which each reported being bullied at school and the social support available to them. Multiple regression analyses indicated that for both sexes frequent peer victimization and low social support contributed significantly and independently to relatively poor mental health. PMID- 10700373 TI - An empirical study of adolescent dating aggression in the U.K. AB - The present study provides one of the first empirical investigations of adolescent dating aggression (ADA) in Britain. The survey found almost half of sampled boys, and more than half of sampled girls, experienced psychological, physical and/or sexual aggression. The study found no significant association between religious affiliation, household composition, age, social class or the use of alcohol and ADA. The study also combined quantitative and qualitative methods to investigate the "symmetry of violence" theory, concluding that when the meaning and context of aggression are considered, male physical and sexual aggression is a significant problem in adolescent heterosexual relationships. PMID- 10700374 TI - Adolescent males' experience of the counselling process. AB - This study examines 11 adolescent males' self-reports of their experiences of 23 counselling sessions to identify what they found helpful and unhelpful during key moments in the therapeutic process. The findings suggest that the experiences of the adolescent males in this study are similar in many ways to the to the reported experiences of adults in counselling. In particular, the experience of emotional support and relief appears to be highly significant for adolescent males, who give significantly lesser importance to cognitive task factors. PMID- 10700375 TI - Barriers to ego identity status formation: a contextual qualification of Marcia's identity status paradigm. AB - While many psychological, sociological and educational researchers acknowledge that ego identity formation is a socially embedded process, others have found that identity research often focuses, almost exclusively, on internal psychological development. The concept of "barriers" provides a means by which to describe external influences associated with adolescent and young adult ego identity exploration and commitment processes which affect and possibly limit individual developmental options. Based on Erikson's assumption that ego identity formation involves both personal growth and communal change, the barriers qualification expands upon Marica's identity status paradigm to more accurately reflect socio-cultural variables which may have impact upon individual internal psychological function. PMID- 10700376 TI - The internal consistency of the child abuse potential inventory with adolescent mothers. AB - In this study, 105 adolescents completed the Child Abuse Potential Inventory (CAP) at an average of 2 months postpartum. The purpose was to assess the reliability (internal consistency) of the CAP with adolescent mothers. The second purpose was to begin to establish a line of inquiry that examines the value of using the CAP with this population. The results showed that the alpha reliabilities were low for the CAP abuse scale (0.65) and low to moderate for its six subscales (range 0.59-0.74). The conclusion was drawn that further research is needed to understand the psychometric properties of the CAP with adolescent mothers. PMID- 10700377 TI - Purification and characterization of Candida albicans 20S proteasome: identification of four proteasomal subunits. AB - The 20S proteasome from yeast cells of Candida albicans was purified by successive chromatographic steps to apparent homogeneity, as judged by nondenaturing and denaturing polyacrylamide gel electrophoresis. Its molecular mass was estimated to be 640 kDa by gel filtration. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate gave at least 10 bands in the range 20-32 kDa. Two-dimensional electrophoresis revealed the presence of at least 14 polypeptides. By electron microscopy after negative staining, the proteasome preparation appeared as typical symmetrical barrel-shaped particles. The enzyme cleaved the peptidyl-arylamide bonds in the model synthetic substrates Cbz-G-G-L-p-nitroanilide, Cbz-G-G-R-beta-naphthylamide, and Cbz-L-L-E-beta naphthylamide (chymotrypsin-like, trypsin-like, and peptidylglutamyl-peptide hydrolyzing activities). The differential sensitivity of these activities to aldehyde peptides and sodium dodecyl sulfate supported the multicatalytic nature of this enzyme. Three proteasomal subunits were identified as alpha6/Pre5, alpha3/Y13, and alpha5/Pup2 by internal sequencing of tryptic fragments. Their sequences perfectly matched the corresponding deduced amino acid sequences of the C. albicans genes. A fourth subunit was identified as alpha7/Prs1 by immunorecognition with a monoclonal antibody specific for C8, the human proteasome subunit homologue. Treatment of the intact isolated 20S proteasome with acid phosphatase and Western blot analysis of the separated components indicated that the alpha7/Prs1 subunit is obtained as a multiply phosphorylated protein. PMID- 10700378 TI - Role of protein-solvent interactions in refolding: effects of cosolvent additives on the renaturation of porcine pancreatic elastase at various pHs. AB - The effects of cosolvent additives on the refolding of porcine pancreatic elastase were studied by comparing the enzymatic activity and the conformation of the enzyme renatured at various pHs with those of the native elastase under the same cosolvent and pH conditions. A lag period was observed before reaching the steady state of the hydrolysis of an amide substrate, and the lag period measured with the refolding enzyme was longer than that measured with the native elastase. Depending on the cosolvent studied (acetonitrile, dimethylsulfoxide, glycerol, methanol) there was or was not a dramatic increase in the duration of the lag period measured with the refolding enzyme, but not in the case of the native elastase. These results and additional kinetic data on inactivation of the enzyme demonstrated that dimethylsulfoxide, glycerol, and methanol enhance the stability of the intermediates able to refold into the native form, contrary to acetonitrile. In neither the case of the native enzyme nor that of the renatured enzyme, did the cosolvents modify the pK(app) of ionization of the amino acids that control enzymatic activity, indicating that they did not penetrate the core of the refolded elastase. Conversely, they shifted toward a more alkaline pH the structural transition of the native elastase, and the amplitude of the shift was comparable to that observed in bulk water with elastase whose Ser 195 has been acylated, suggesting that cosolvents stabilized the structure of the folded molecule by increasing its packing. PMID- 10700379 TI - Effect of limited proteolysis on phospholipase C-gamma1 kinetics. AB - Phospholipase C-gamma1 is a tightly regulated, multidomain protein that generates the second messengers inositol 1,4,5-trisphosphate and diacylglycerol. Kinetic analysis reveals that phospholipase C-gamma1 displays apparent allosteric behavior. A previous study determined that proteolytic cleavage of the SH domain region of phospholipase C-gamma1 yields an activated form of the enzyme (A. W. Fernald, G. A. Jones, and G. Carpenter Biochem. J. 302, 508, 1994). In this study, we show that activation of phospholipase C-gamma1 by proteolysis decreases both the cooperativity and the half-maximal value of the enzyme for substrate. Kinetic analysis revealed that the mole fraction of phosphatidylinositol 4,5 bisphosphate (PIP(2)) that resulted in half-maximal PIP(2) hydrolysis (S(0.5)) was lower for proteolyzed than uncleaved phospholipase C-gamma1 (0.08 mole fraction vs 0.18 mole fraction of PIP(2)). The cooperativity index was lower for proteolyzed than full-length phospholipase C-gamma1 (n = 2.5 vs n = 4). Kinetic analysis also revealed that the estimated dissociation constant was lower for phospholipase C-gamma1 that had been subjected to proteolysis (0.1 mM vs 1.0 mM PIP(2) for cleaved vs uncleaved phospholipase C-gamma1, respectively). It was previously hypothesized that activation of phospholipase C-gamma1 requires a conformational change that results in increased accessibility of substrate to the active site and that the SH domain of the enzyme is involved in the activation event. These experiments support the hypothesis that a portion of the protein covers the active site, allosterically inhibiting the enzyme, and that the removal of this "lid" domain activates the enzyme. PMID- 10700380 TI - Spontaneous porphyria of the Long-evans cinnamon rat: an animal model of Wilson's disease. AB - To confirm and extend our previous microspectrophotometric observations of 30 week-old male Long-Evans Cinnamon (LEC) rats, an animal model of human Wilson's disease, we analyzed the porphyrin patterns of the organs, urine, and plasma of LEC rats. Abnormal accumulation of porphyrins, especially highly carboxylated porphyrins (uro- and heptaporphyrin), in the kidneys and liver was seen in male and female LEC rats aged 30 weeks and also in 10-week-old rats, before the onset of spontaneous hepatic dysfunction. Accumulation of copper and iron in the kidneys was not observed in the 10-week-old rats. Massive accumulation of porphyrins was observed only in the kidneys of the 30-week-old male LEC rat, indicating that this symptom is related to sex and age. Renal accumulation of porphyrins was reflected in the rate of urinary porphyrin excretion. Hepatic accumulation of porphyrins appeared to be independent of sex and age. These results indicate that neither renal nor hepatic porphyrin accumulation is the result of renal deposition of metals or of spontaneous hepatic dysfunction and that porphyrinuria in the LEC rat is closely related to the renal accumulation of porphyrins. In contrast to these organs, a reduction in the porphyrin levels was observed in the brain of the LEC rat. This was independent of sex and age. The present work stresses the existence of an abnormal heme metabolism in the LEC rat, and thus, the necessity to study the heme metabolism in human Wilson's disease is strongly suggested. PMID- 10700381 TI - Importance of cysteine residues for the stability and catalytic activity of human pancreatic beta cell glucokinase. AB - The low-affinity glucose phosphorylating enzyme glucokinase has the function of a physiological glucose sensor in pancreatic beta cells and in liver. In contrast to the high-affinity hexokinase types I-III glucokinase shows extraordinary sensitivity toward SH group oxidizing compounds. To characterize the function of sulfhydryl groups cysteine residues in the vicinity of the sugar binding site (Cys 213, Cys 220, Cys 230, Cys 233, and Cys 252) as well as cysteine residues a distance from the active site (Cys 364, Cys 371, and Cys 382), they were replaced in human beta cell glucokinase by serine through site-directed mutagenesis. Controlled proteolysis of wild-type glucokinase by proteinase K revealed that the SH group oxidizing agent alloxan can induce the formation of multiple intramolecular disulfide bridges corresponding to a double-band pattern of glucokinase protein in nonreducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The formation of intramolecular disulfide bridges altered the mobility of the protein. None of the cysteine mutations could prevent the formation of the 49-kDa glucokinase conformation after alloxan treatment. The cysteine mutants Cys 233, Cys 252, and Cys 382 showed nearly complete loss of catalytic activity, whereas the V(max) values of the Cys 213, Cys 220, Cys 364, and Cys 371 mutants were decreased by 30-60%. Only the Cys 230 mutant showed kinetic characteristics comparable to those of wild-type glucokinase. The sensitivity of the Cys 213, Cys 230, Cys 364, and Cys 371 mutants toward alloxan induced inhibition of enzyme activity was up to 10-fold lower compared with wild type glucokinase. d-Glucose and dithiotreitol provided protection against alloxan induced inhibition of wild-type glucokinase and all catalytically active cysteine mutants. Conclusively our data demonstrate the functional significance of the cysteine residues of beta cell glucokinase for both structural instability of the enzyme and catalytic function. Knowledge of sensitive cysteine targets may help to develop strategies that improve glucokinase enzyme function under conditions of oxidative stress. PMID- 10700382 TI - Terpenoid secondary metabolism in Arabidopsis thaliana: cDNA cloning, characterization, and functional expression of a myrcene/(E)-beta-ocimene synthase. AB - The Arabidopsis genome project has recently reported sequences with similarity to members of the terpene synthase (TPS) gene family of higher plants. Surprisingly, several Arabidopsis terpene synthase-like sequences (AtTPS) share the most identity with TPS genes that participate in secondary metabolism in terpenoid accumulating plant species. Expression of a putative Arabidopsis terpene synthase gene, designated AtTPS03, was demonstrated by amplification of a 392-bp cDNA fragment using primers designed to conserved regions of plant terpene synthases. Using the AtTPS03 fragment as a hybridization probe, a second AtTPS cDNA, designated AtTPS10, was isolated from a jasmonate-induced cDNA library. The partial AtTPS10 cDNA clone contained an open reading frame of 1665 bp encoding a protein of 555 amino acids. Functional expression of AtTPS10 in Escherichia coli yielded an active monoterpene synthase enzyme, which converted geranyl diphosphate (C(10)) into the acyclic monoterpenes beta-myrcene and (E)-beta ocimene and small amounts of cyclic monoterpenes. Based on sequence relatedness, AtTPS10 was classified as a member of the TPSb subfamily of angiosperm monoterpene synthases. Sequence comparison of AtTPS10 with previously cloned monoterpene synthases suggests independent events of functional specialization of terpene synthases during the evolution of terpenoid secondary metabolism in gymnosperms and angiosperms. Functional characterization of the AtTPS10 gene was prompted by the availability of Arabidopsis genome sequences. Although Arabidoposis has not been reported to form terpenoid secondary metabolites, the unexpected expression of TPS genes belonging to the TPSb subfamily in this species strongly suggests that terpenoid secondary metabolism is active in the model system Arabidopsis. PMID- 10700383 TI - Lectin histochemical examination of rabbit bladder glycoproteins and characterization of a mucin isolated from the bladder mucosa. AB - The glycocalyx of the mucosal surface of urinary bladder acts as an effective barrier against invasion by pathogenic microorganisms and injury from toxic substances in the urine. Defects in these bladder mucosal components could thus be important factors in the development of diseases such as interstitial cystitis and lower urinary tract infections. However, information on the nature of glycoconjugates of mammalian bladder mucosa is very limited. In this study, the glycoconjugates of rabbit bladder were examined histochemically using biotinylated lectins with specificities for a variety of carbohydrate moieties. Three [Artocarpus integrifolia (Jacalin), Datura stramonium (DSL), and Maackia amurensis II (MAL-II)] of the lectins bound predominantly to the luminal cell layer, with decreased binding to the basal layers of the epithelium. In contrast, Ricinus communis I and Sambucus nigra lectins did not bind to the cells in the epithelium but strongly interacted with the subepithelial layers, especially the lamina propria. The intensity of the staining by Jacalin and MAL-II was significantly reduced by prior treatment of the bladder sections with O sialoglycoprotein endopeptidase, indicating that the ligands of these lectins are primarily mucin glycoproteins. In parallel biochemical studies, a high-molecular weight glycoprotein with characteristics typical of epithelial mucins was purified from the mucosa of rabbit bladder explant cultures metabolically labeled with [(3)H]glucosamine. Quantitative analysis of the sialic acid, uronic acid, and hexosamine contents of delipidated rabbit bladder mucosa revealed a larger proportion of sialoglycoproteins compared with glycosaminoglycans. Taken together, the results of histochemical and biochemical analyses indicate that glycoproteins rather than glycosaminoglycans are the major components of the bladder epithelium, and that the former include a mucin. PMID- 10700384 TI - Molecular cloning and functional expression of contortrostatin, a homodimeric disintegrin from southern copperhead snake venom. AB - Contortrostatin is a unique dimeric disintegrin isolated from southern copperhead snake venom. Through antagonism of integrins alphaIIbbeta3, alpha5beta1, alphavbeta3, and alphavbeta5, contortrostatin inhibits platelet aggregation and disrupts cancer cell adhesion and invasion. We cloned cDNA from a library made from the venom gland cells of Agkistrodon contortrix contortrix using polymerase chain reaction. We found that the contortrostatin gene is part of a precursor composed of proprotein, metalloproteinase, and disintegrin domains. The precursor cDNA is 2027 bp with a 1449-bp open reading frame. The disintegrin domain is 195 bp encoding 65 amino acids. Like other members of the disintegrin family, each subunit of contortrostatin has an RGD site, and the cysteine alignment is conserved. The disintegrin domain of the cDNA has been expressed in a eukaryotic expression system as a homodimeric fusion protein with an immunoglobulin. The recombinant protein is recognized by an antiserum against native contortrostatin in Western blot. Both the native and recombinant proteins bind to integrins alphavbeta3 and alphavbeta5. Like native contortrostatin, the recombinant fusion protein inhibits platelet aggregation, blocks cancer cell adhesion to fibronectin and vitronectin, and prevents invasion of cancer cells through a Matrigel barrier. The success of functional expression not only validates the cDNA cloning of this disintegrin, but also provides adequate material for functional studies of contortrostatin. PMID- 10700385 TI - Phospholipase A(2) with platelet aggregation inhibitor activity from Austrelaps superbus venom: protein purification and cDNA cloning. AB - Four phospholipase A(2) (PLA(2)) enzymes (Superbins a, b, c, and d) with varying platelet aggregation inhibitor activities have been purified from Austrelaps superbus by a combination of gel filtration, ion-exchange, and reversed-phase high-pressure liquid chromatography. Purity and homogeneity of the superbins have been confirmed by high-performance capillary zone electrophoresis and mass spectrometry. The electron spray ionization mass spectrometry data showed that their molecular masses range from 13,140 to 13,236 Da. Each of the proteins has been found to be basic and exhibit varying degrees of PLA(2) activity. They also displayed different platelet aggregation inhibitory activities. Superbin a was found to possess the most potent inhibitory activity with an IC(50) of 9.0 nM, whereas Superbin d was found to be least effective with an IC(50) of 3.0 microM. Superbins b and c were moderately effective with IC(50) values of 0.05 and 0.5 microM, respectively. The amino-terminal sequencing confirmed the identity of these superbins. cDNA cloning resulted in the identification of 17 more PLA(2) isoforms in A. superbus venom. It has also provided complete information on the precursor PLA(2). The precursor PLA(2) contained a 27-amino-acid signal peptide and 117- to 125-amino-acid PLA(2) (molecular mass ranging from 13,000 to 14,000 Da). Two of these PLA(2) enzymes resembled more closely (87%) Superbin a in structure. Two unique PLA(2) enzymes containing an extra pancreatic loop also have been identified among the isoforms. PMID- 10700386 TI - Ectonucleotide diphosphohydrolase activities in Entamoeba histolytica. AB - In this work, we describe the ability of living cells of Entamoeba histolytica to hydrolyze extracellular ATP. In these intact parasites, whose viability was determined by motility and by the eosin method, ATP hydrolysis was low in the absence of any divalent metal (78 nmol P(i)/h/10(5) cells). Interestingly, in the presence of 5 mM MgCl(2) an ecto-ATPase activity of 300 nmol P(i)/h/10(5) cells was observed. The addition of MgCl(2) to the extracellular medium increased the ecto-ATPase activity in a dose-dependent manner. At 5 mM ATP, half-maximal stimulation of ATP hydrolysis was obtained with 1.23 mM MgCl(2). Both activities were linear with cell density and with time for at least 1 h. The ecto-ATPase activity was also stimulated by MnCl(2) and CaCl(2) but not by SrCl(2), ZnCl(2), or FeCl(3). In fact, FeCl(3) inhibited both Mg(2+)-dependent and Mg(2+) independent ecto-ATPase activities. The Mg(2+)-independent ATPase activity was unaffected by pH in the range between 6.4 and 8. 4, in which the cells were viable. However, the Mg(2+)-dependent ATPase activity was enhanced concomitantly with the increase in pH. In order to discard the possibility that the ATP hydrolysis observed was due to phosphatase or 5'-nucleotidase activities, several inhibitors for these enzymes were tested. Sodium orthovanadate, sodium fluoride, levamizole, and ammonium molybdate had no effect on the ATPase activities. In the absence of Mg(2+) (basal activity), the apparent K(m) for ATP(4-) was 0.053 +/- 0.008 mM, whereas at saturating MgCl(2) concentrations, the corresponding apparent K(m) for Mg-ATP(2-) for Mg(2+)-dependent ecto-ATPase activity (difference between total and basal ecto-ATPase activity) was 0.503 mM +/- 0.062. Both ecto-ATPase activities were highly specific for ATP and were also able to hydrolyze ADP less efficiently. To identify the observed hydrolytic activities as those of an ecto-ATPase, we used suramin, a competitive antagonist of P(2) purinoreceptors and an inhibitor of some ecto-ATPases, as well as the impermeant agent 4'-4'-diisothiocyanostylbenzene-2'-2'-disulfonic acid. These two reagents inhibited the Mg(2+)-independent and the Mg(2+)-dependent ATPase activities to different extents, and the inhibition by both agents was prevented by ATP. A comparison among the ecto-ATPase activities of three amoeba species showed that the noninvasive E. histolytica and the free-living E. moshkovskii were less efficient than the pathogenic E. histolytica in hydrolyzing ATP. As E. histolytica is known to have a galactose-specific lectin on its surface, which is related to the pathogenesis of amebiasis, galactose was tested for an effect on ecto-ATPase activities. It stimulated the Mg(2+)-dependent ecto-ATPase but not the Mg(2+)-independent ATPase activity. PMID- 10700387 TI - In vivo processing of nonanchored Yapsin 1 (Yap3p). AB - A C-terminally truncated form of yapsin 1 (yeast aspartic protease 3) was overexpressed in yeast and its processing through the secretory pathway was followed by pulse-labeling and immunoprecipitation studies. In the soluble cell extract, three forms of yapsin 1-87, 74, and 18 kDa-were found. Identification of these forms of yapsin 1 using different antisera suggests that the 87-kDa form is pro-yapsin 1, which is processed into two subunits, alpha (18 kDa) and beta (74 kDa), by cleavage at a loop region not found in traditional aspartic proteases. By use of a temperature-sensitive mutant strain, sec18, the generation of the two subunits was found to occur in the endoplasmic reticulum. An active site-mutated yapsin 1 was not processed into the two subunits, suggesting that this process occurs in an autocatalytic manner. PMID- 10700388 TI - alpha(1,3)fucosyltransferases expressed by the gain-of-function Chinese hamster ovary glycosylation mutants LEC12, LEC29, and LEC30. AB - Gain-of-function glycosylation mutants provide access to glycosylation pathways, glycosylation genes, and mechanisms that regulate expression of a glycotype. Previous studies have shown that the gain-of-function Chinese hamster ovary (CHO) mutants LEC12, LEC29, and LEC30 express an N-ethylmaleimide-resistant alpha(1, 3)fucosyltransferase (alpha(1,3)Fuc-T) activity that is not detected in CHO cells and that generates the Lewis(X) but not the sialyl-Lewis(X) determinant. The three mutants differ, however, in lectin resistance properties, expression of fucosylated antigens, and in vitro alpha(1,3)Fuc-T activities. In this paper we show that each mutant expresses Fuc-TIX, but only LEC30 cells express Fuc-TIV. Using genomic PCR and reverse-transcriptase (RT)-PCR strategies, we isolated coding portions of the CHO Fut4 and Fut9 genes. Each gene is present in a single copy in the CHO and mutant genomes. The Fut4 gene is expressed only in LEC30 cells, while all three mutants express the Fut9 gene. Interestingly, the fucosylation phenotypes of LEC12 and LEC29 cells do not correlate with the relative abundance of their Fut9 gene transcripts (LEC29 >> LEC12). Compared to LEC29 cells, LEC12 cells have an approximately 40-fold higher in vitro alpha(1,3)Fuc-T activity and bind the VIM-2 monoclonal antibody, whereas LEC29 cells do not bind VIM-2. Mixing experiments did not detect Fuc-TIX inhibitory activity in LEC29 cell extracts, and CHO cells expressing a transfected Fut9 gene behaved like LEC12 cells. Therefore, it seems that LEC29 cells may not translate their more abundant Fut9 gene transcripts efficiently or may not synthesize appropriate acceptors for internal alpha(1,3)fucosylation. Alternatively, LEC12 cells may possess, in addition to Fuc-TIX, a novel alpha(1,3)Fuc-T activity. PMID- 10700389 TI - Reserpine attenuates interstitial adenosine-mediated activation of ecto-5' nucleotidase in rat hearts in vivo. AB - We examined whether reserpine-induced norepinephrine (NE) depletion attenuated the products of adenosine in rat heart. A flexibly mounted microdialysis technique was used to measure the concentration of interstitial adenosine and to assess the activity of ecto-5'-nucleotidase in rat hearts in situ. The microdialysis probe was implanted in the left ventricular myocardium of anesthetized rats and perfused with Tyrode solution containing adenosine 5' monophosphate (AMP) at rate of 1.0 microliter/min. The baseline level of dialysate adenosine was 0.51 +/- 0.09 microM. The introduction of AMP (100 microM) through the probe increased markedly the dialysate adenosine to 8.95 +/- 0.86 microM, and this increase was inhibited by ecto-5'-nucleotidase inhibitor, alpha, beta-methyleneadenosine 5'-diphosphate (AOPCP, 100 microM), to 0.66 +/- 0.38 microM. Thus, the level of dialysate adenosine is a measure of the ecto-5' nucleotidase activity in the tissue in situ. AMP concentration for the half maximal effect of adenosine release (EC(50)) was 107.3 microM. The maximum attainable concentration of dialysate adenosine (E(max)) by AMP was 21.1 microM. However, the EC(50) and E(max) values with reserpinized animals were 106.9 and 7.1 microM, respectively. Electrical stimulation of the left stellate ganglion increased significantly dialysate adenosine concentration, from the control level of 8.66 +/- 0.96 microM to 12.38 +/- 1.11 microM. After stimulation, dialysate adenosine returned to near the prestimulation level. When corresponding experiments were performed with reserpinized animals, the effect of electrical stimulation was abolished. Tyramine (endogenous catecholamine trigger) increased the adenosine concentration in a concentration-dependent manner. However, the elevation of adenosine concentration with reserpinized animals was not observed. These results suggest that reserpine attenuates NE-induced adenosine via stimulation of alpha(1)-adrenoceptor and protein kinase C mediated activation of ecto-5'-nucleotidase in rat heart. PMID- 10700390 TI - Effects of Al(3+) and related metals on membrane phase state and hydration: correlation with lipid oxidation. AB - The aim of the present study was to further understand how changes in membrane organization can lead to higher rates of lipid oxidation. We previously demonstrated that Al(3+), Sc(3+), Ga(3+), Be(2+), Y(3+), and La(3+) promote lipid packing and lateral phase separation. Using the probe Laurdan, we evaluated in liposomes if the higher rigidity of the membrane caused by Al(3+) can alter membrane phase state and/or hydration, and the relation of this effect to Al(3+) stimulated lipid oxidation. In liposomes of dimyristoyl phosphatidylcholine and dimyristoyl phosphatidylserine, Al(3+) (10-100 microM) induced phase coexistence and displacement of T(m). In contrast, in liposomes of brain phosphatidylcholine and brain phosphatidylserine, Al(3+) (10-200 microM) did not affect membrane phase state but increased Laurdan generalized polarization (GP = -0. 04 and 0.09 in the absence and presence of 200 microM Al(3+), respectively). Sc(3+), Ga(3+), Be(2+), Y(3+), and La(3+) also increased GP values, with an effect equivalent to a decrease in membrane temperature between 10 and 20 degrees C. GP values in the presence of the cations were significantly correlated (r(2) = 0.98, P < 0.001) with their capacity to stimulate Fe(2+)-initiated lipid oxidation. Metal-promoted membrane dehydration did not correlate with ability to enhance lipid oxidation, indicating that dehydration of the phospholipid polar headgroup is not a mechanism involved in cation-mediated enhancement of Fe(2+)-initiated lipid oxidation. Results indicate that changes in membrane phospholipid phase state favoring the displacement to gel state can facilitate the propagation of lipid oxidation. PMID- 10700391 TI - The existence of a lysosomal redox chain and the role of ubiquinone. AB - Several studies concerning the distribution of ubiquinone (UQ) in the cell report a preferential accumulation of this biogenic quinone in mitochondria, plasma membranes, Golgi vesicles, and lysosomes. Except for mitochondria, no recent comprehensive experimental evidence exists on the particular function of UQ in these subcellular organelles. The aim of a recent study was to elucidate whether UQ is an active part of an electron-transfer system in lysosomes. In the present work, a lysosomal fraction was prepared from a light mitochondrial fraction of rat liver by isopycnic centrifugation. The purity of our preparation was verified by estimation of the respective marker enzymes. Analysis of lysosomes for putative redox carriers and redox processes in lysosomes was carried out by optical spectroscopy, HPLC, oxymetry, and ESR techniques. UQ was detected in an amount of 2.2 nmol/mg of protein in lysosomes. Furthermore, a b-type cytochrome and a flavin-adenine dinucleotide (FAD) were identified as other potential electron carriers. Since NADH was reported to serve as a substrate of UQ redox chains in plasma membranes, we also tested this reductant in lysosomes. Our experiments demonstrate a NADH-dependent reduction of UQ by two subsequent one electron-transfer steps giving rise to the presence of ubisemiquinone and an increase of the ubiquinol pool in lysosomes. Lysosomal NADH oxidation was accompanied by an approximately equimolar oxygen consumption, suggesting that O(2) acts as a terminal acceptor of this redox chain. DMPO/(*)OH spin adducts were detected by ESR in NADH-supplemented lysosomes, suggesting a univalent reduction of oxygen. The kinetic analysis of redox changes in lysosomes revealed that electron carriers operate in the sequence NADH > FAD > cytochrome b > ubiquinone > oxygen. By using the basic spin label TEMPAMINE, we showed that the NADH-related redox chain in lysosomes supports proton accumulation in lysosomes. In contrast to the hypothesis that UQ in lysosomes is simply a waste product of autophagy in the cell, we demonstrated that this lipophilic electron carrier is a native constituent of a lysosomal electron transport chain, which promotes proton translocation across the lysosomal membrane. PMID- 10700392 TI - Inhibition of bacterial peptide deformylase by biaryl acid analogs. AB - Peptide deformylase is an essential eubacterial metalloenzyme involved in the maturation of proteins by cleaving the N-formyl group from N-blocked methionine polypeptides. Biaryl acid analogs containing tetrazole, acyl sulfonamide, or carboxylate pharmacophores were found to be potent inhibitors of recombinant Escherichia coli peptide deformylase. Two of these compounds, a biphenyl tetrazole, compound 1, and a biphenyl acyl sulfonamide, compound 4, were competitive inhibitors with K(i) values of 1.2 and 6.0 microM, respectively. By analogy to the binding of related compounds to other metalloenzymes such as Bacteroides fragilis metallo-beta-lactamase CcrA and human carbonic anhydrase, a mechanism of inhibition is proposed for these peptide deformylase inhibitors where the acidic moieties form direct ionic interactions with the active site metal cation. PMID- 10700393 TI - Disruption of interkringle disulfide bond of plasminogen kringle 1-3 changes the lysine binding capability of kringle 2, but not its antiangiogenic activity. AB - Kringle 1-3 of human plasminogen is a potent inhibitor of endothelial cell proliferation. To understand a possible role for the unique cystine bridge between kringle 2 and kringle 3, we disrupted the interkringle disulfide bond by mutating Cys(169) and Cys(297) to serine residues. The yield of the mutant during the refolding process was decreased significantly. Anti-endothelial cell proliferative activity of the mutant was similar to that of the wild type. There was no significant difference in in vivo antiangiogenic activity between the wild type and the mutant in chorioallantoic membrane assay. However, in the mutant, the weak lysine binding capability of kringle 2 was not detected and its mobility in nonreducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis is different from that of the wild type. These results support the notion that the overall antiangiogenic function of angiostatin is mediated by individual kringles, and suggest that the lysine binding capability of kringle 2 is likely not important for the antiangiogenic activity of kringle 1-3. PMID- 10700394 TI - Purification of recombinant flavanone 3beta-hydroxylase from petunia hybrida and assignment of the primary site of proteolytic degradation. AB - Flavanone 3beta-hydroxylase catalyzes the Fe(II)/oxoglutarate-dependent hydroxylation of (2S)-flavanones to (2R,3R)-dihydroflavonols in the course of flavonol/anthocyanin or catechin biosynthesis. The enzyme from Petunia hybrida consists of a 41,655-Da polypeptide that is prone to rapid proteolysis in crude plant extracts as well as on expression in Escherichia coli, and commercial protease inhibitors were inefficient in stopping the degradation. To pinpoint the primary site of proteolysis and to improve the activity yields, two revised schemes of purification were developed for the recombinant polypeptides. Applying a four-step protocol based on extraction and ion-exchange chromatography at pH 7.5, the primary, catalytically inactive proteolytic enzyme fragment (1.1 mg) was isolated and shown to cross-react on Western blotting as one homogeneous band of about 38 kDa. Mass spectrometric analysis assigned a mass of 37,820 +/- 100 Da to this fragment, and partial sequencing revealed an unblocked amino terminus identical to that of the native 3beta-hydroxylase. Thus, the native enzyme had been degraded by proteolysis of a small carboxy-terminal portion, and the primary site of cleavage must be assigned most likely to the Glu 337-Leu 338 bond, accounting for a loss of about 3800 Da. Alternatively, the enzyme degradation was greatly reduced when the extraction of recombinant bacteria was carried out with phosphate buffer at pH 5.5 followed by size exlusion and anion-exchange chromatography. This rapid, two-step purification resulted in a homogeneous 3beta hydroxylase of high specific acitivity (about 32 mkat/kg) at roughly 5% yield, and the procedure is a major breakthrough in mechanistic investigations of this class of labile dioxygenases. PMID- 10700395 TI - Effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin on the expression of luteinizing hormone receptors during cell differentiation in cultured granulosa cells. AB - Dioxin (2,3,7,8-tetrachlorodibenzo-p-dioxin, TCDD) is a common environmental pollutant causing public concern. By use of a cell culture system derived from rat granulosa cells that provides unique advantages for studying the molecular mechanisms underlying the action of TCDD, the influence of TCDD on luteinizing hormone receptor (LHR) induction was examined. Treatment with follicle stimulating hormone (FSH) produced, as expected, a substantial increase in specific LHR expression; concurrent treatment with TCDD (10 pM) resulted in a significant decrease in LHR after 24 h. Cotreatment with 30 ng/ml FSH and increasing doses of TCDD inhibited the levels of FSH-induced LHR mRNA in a dose dependent manner, and 1 pM TCDD inhibited FSH-induced LHR significantly after 48 h. The rate of LHR mRNA gene transcription, assessed by nuclear run-on transcription assay, was found to decrease after addition of TCDD. The decay curves for the 5.4-kb LHR mRNA transcript showed a significant decrease after addition of TCDD. PMID- 10700396 TI - Lithium's effects on rat liver glucose metabolism in vivo. AB - Oral administration of lithium carbonate to fed-healthy rats strongly decreased liver glycogen content, despite the simultaneous activation of glycogen synthase and the inactivation of glycogen phosphorylase. The effect seemed to be related to a decrease in glucose 6-phosphate concentration and to a decrease in glucokinase activity. Moreover, in these animals lithium markedly decreased liver fructose 2,6-bisphosphate, which could be a consequence of the fall in glucose 6 phosphate and of the inactivation of 6-phosphofructo-2-kinase. Liver pyruvate kinase activity and blood insulin also decreased after lithium administration. Lower doses of lithium carbonate had less intense effects. Lithium administration to starved-healthy and fed-streptozotocin-diabetic rats caused a slight increase in blood insulin, which was simultaneous with increases in liver glycogen, glucose 6-phosphate, and fructose 2, 6-phosphate. Glucokinase, 6-phosphofructo-2 kinase, and pyruvate kinase activities also increased after lithium administration in starved-healthy and fed-diabetic rats. Lithium treatment activated glycogen synthase and inactivated glycogen phosphorylase in a manner similar to that observed in fed-healthy rats. Glycemia was not modified in any group of animals. These results indicate that lithium acts on liver glycogen metabolism in vivo in at least two different ways: one related to changes in insulinemia, and the other related to the direct action of lithium on the activity of some key enzymes of liver glucose metabolism. PMID- 10700397 TI - Functional expression of an Arabidopsis cDNA clone encoding a flavonol 3'-O methyltransferase and characterization of the gene product. AB - We report that the cDNA clone (Accession No. U70424), previously isolated from Arabidopsis thaliana as encoding a caffeic acid/5-hydroxyferulic acid O methyltransferase (OMT) (1), has now been overexpressed in Escherichia coli BL21 and its recombinant protein identified as a novel flavonol 3'-OMT. It is, therefore, renamed AtOMT1. This cDNA clone has previously been identified on the basis of its 88% amino acid sequence similarity and 80% identity to the aspen bispecific lignin OMT (2), the type member of the group involved in lignin biosynthesis. Our data indicate that this novel OMT uses the flavonol quercetin as the preferred substrate, but neither of the hydroxycinnamic acids, caffeic or 5-hydroxyferulic, to any significant extent. This indicates that the high sequence similarity/identity of AtOMT1 to that of the aspen lignin OMT (2) is not sufficient to assign the function of this gene product. PMID- 10700398 TI - Contact-Angle Hysteresis Caused by a Random Distribution of Weak Heterogeneities on a Solid Surface. AB - A model according to which contact-angle hysteresis arises as the result of a random distribution of irregularities on the solid surface is investigated on the basis of probability theory. An estimate is obtained of the mathematical expectation of the number of stable equilibria when the effective angle between the liquid-gas surface and the solid surface with which the liquid is in contact deviates from the value, say theta(0), which would obtain if the solid surface were uniform, i.e., free from irregularities. It is found that when the effective contact angle deviates from theta(0) by less than a critical value, then the expected number of stable equilibria increases exponentially with the length of the contact line; therefore such a contact angle can occur under static conditions. But if the deviation of the contact angle from theta(0) exceeds the critical value, then the expected number of stable equilibria decreases exponentially with the length of the contact line, so a stable equilibrium is not possible for a macroscopic length of the contact line. The method is applicable only if the random deviations of the spreading power (defined as the solid-gas surface tension minus the sum of the liquid-gas and liquid-solid surface tensions) from its average are sufficiently small. It is found that the critical deviation of the contact angle from theta(0) is, apart from a slowly varying logarithmic factor, proportional to H(2)rho(s), where H is a measure of the amplitude of the surface irregularities and rho(s) is the surface density (i.e., number per unit area) of the irregularities. This qualitative feature agrees with the results previously obtained by several other authors, and, moreover, there is a surprisingly close agreement of the proportionality factor with the results of some earlier work in which the method of statistical analysis was much less elaborate than here. The effect of the logarithmic factor is to make the critical deviation of the contact angle increase more slowly than the first power of H(2)rho(s), and this is also in qualitative agreement with some earlier work. Copyright 2000 Academic Press. PMID- 10700399 TI - Formation of Layered Single- and Double-Metal Hydroxide Precipitates at the Mineral/Water Interface: A Multiple-Scattering XAFS Analysis. AB - Spectroscopic and microscopic studies have shown that Ni and Co sorption by clay minerals may proceed via formation of surface precipitates. Several studies employing X-ray absorption fine structure (XAFS) spectroscopy suggested the formation of turbostratic, alpha-type metal hydroxides, of layered double hydroxides (LDH) with Al-for-metal substitution, and of 1:1 or 2:1 phyllosilicates. Distinction of these phases is difficult because they have low crystallinity and/or a small mass compared to the sorbents, and because they have similar metal-metal distances in their hydroxide layers/sheets. Distinction of these phases is crucial, however, because they have substantially differing solubilities. In this paper we show that an XAFS beat pattern at about 8 A(-1) can be used as a fingerprint to unequivocally distinguish LDH from the alpha-type hydroxides and phyllosilicates. Full multiple-scattering simulations and experimental spectra of model compounds indicate that the beat pattern is due to focused multiple scattering at Me/Al ratios between 1 and 4 (Me=Ni, Co). By applying the fingerprint method to new and to already published XAFS data on Ni and Co surface precipitates, we found that LDH preferentially forms in the presence of the Al-containing sorbents pyrophyllite, illite, kaolinite, gibbsite, and alumina above pH 7.0. However, alpha-type metal hydroxides form in the presence of the Al-free sorbents talc, silica, and rutile, and in the presence of the Al-containing clay minerals montmorillonite and vermiculite. We believe that the high permanent charge of these latter minerals prevents or retards the release of Al. When Al is available, the formation of LDH seems to be thermodynamically and/or kinetically favored over the formation of alpha-type hydroxides. Copyright 2000 Academic Press. PMID- 10700400 TI - Precipitation of Zinc Sulfide Particles from Homogeneous Solutions. AB - Zinc sulfide particles were homogeneously precipitated by thermal decomposition of thioacetamide in acidic aqueous solutions in a one-step process. The influence of the operating conditions (initial concentration of zinc ion and TAA) on the nucleation time and number concentration of the generated particles was investigated. The experimental results show that the model of homogeneous nucleation previously developed and successfully tested for silver particle generation by a chemical reduction method can also be applied to the formation of zinc sulfide particles by homogeneous precipitation. Furthermore, in the particle formation method in which the nucleation time t* can be measured, the particle number concentration n* can be predicted by the simple relation n*=1/(4pir*Dt*) (r* is the critical nucleus radius, and D the monomer diffusion coefficient). Thus the particle number concentration can be easily predicted even if the rate expression and the critical supersaturation concentration are unknown. Copyright 2000 Academic Press. PMID- 10700401 TI - Micellar Behavior of n-Alkyl Sulfates in Binary Mixed Systems. AB - The micellar behavior of the binary mixed systems of sodium n-hexylsulfate with sodium n-decyl-, n-dodecyl-, and n-tetradecylsulfate has been studied. The critical micelle concentration of the mixtures was quantitatively estimated by conductance methods. The micellar composition in the micelles was determined by the Motomura model and the mutual interactions were estimated from Holland and Rubingh's theory. The surfactant mixtures were found to be nonideal. The influence of the alkyl chain length in these parameters was studied. Copyright 2000 Academic Press. PMID- 10700402 TI - Unusual Interfacial Phase Behavior of Two Nonmiscible Liquids in a Cylindrical Test Tube: Equilibrium Shapes and Stability of Axisymmetric Liquid Bridges under Gravity. AB - In this paper the unusual interfacial phase behavior of two nonmiscible fluids contained in a cylindrical glass test tube is reported. Water, which is the lighter phase, takes up the upper part of the tube, whereas the denser compound (a hydrofluorocarbon) is in the bottom. However, below some critical volume of water, the denser phase emerges at the air surface, by forming an axisymmetric liquid bridge through the aqueous phase. Above the critical condition, the formation of the bridge, the evolution of the shape of this bridge, and its final breakdown can be visually inspected after shaking the tube. The minority liquid (water) is dispersed in the majority phase (HCFC) as an unstable dispersion of droplets. Droplets rise to the air surface under the action of the buoyant force, and coalesce on the glass wall: this leads to the formation of a bridge (made from the dispersion in the middle of a hollow axisymmetric water drop), whose height increases and thickness decreases during the coalescence process, until it breaks down. Using a free energy analysis, we state the exact variational problem via its Euler-Lagrange equation. However, since this nonlinear differential equation cannot be solved analytically, a simplified "mean-field" approach is developed, which provides a comprehensive insight into the physical origin of these capillary bridges and their stability under gravity. Copyright 2000 Academic Press. PMID- 10700403 TI - Cubic-Phase-Based Concentrated Emulsions. AB - The effect of different types of added oil on the formation of a discontinuous micellar-type cubic phase was investigated in water-polyoxyethylene dodecyl ether (C(12)EO(25)) systems by phase study and small-angle X-ray scattering. The thermal stability of the cubic phase increases upon addition of oil, especially short-chain hydrocarbons. However, in the heptane system, the maximum melting temperature of the cubic phase is lower than that for decane due to the formation of a different liquid crystal phase. The effect of polyols on C(12)EO(25) cubic phases was also investigated. It was found that the thermal stability of the cubic phase decreases with polyol concentration. The destabilizing effect becomes large as the polyol molecule penetrates further into the surfactant palisade layer. Although the solubilization of oil in the cubic phase is very low, a large amount of excess oil can be incorporated and a transparent cubic-phase-based concentrated emulsion is formed. The transparency is attributed to the very small difference in the refractive indices between the cubic and excess-oil phases. Copyright 2000 Academic Press. PMID- 10700404 TI - Surface Heterogeneity of Passively Oxidized Silicon Carbide Particles: Hydrophobic-Hydrophilic Partition. AB - The surface of silicon carbide (SiC) particles previously subjected to passive oxidation has been characterized using various techniques such as adsorption of positively and negatively charged surfactants in aqueous solution, immersion microcalorimetry in three probe liquids, and flow microcalorimetry in organic media. Adsorption data show that around one quarter of the surface appears negatively charged and thus hydrophilic, while three quarters appear uncharged and hydrophobic. This is attributed to dissociation of silanols groups. Immersion calorimetry in liquids having well-defined polar and nonpolar components of surface energy shows that the Lifshitz-van der Waals component of SiC is very high and that the acid and basic components are weak. The experimental results appear to be consistent with both computations of surface energy using Lifshitz theory and experimental data previously obtained with other minerals. The three indexes are discussed and it is argued that they represent different terms of the solid surface energy. Copyright 2000 Academic Press. PMID- 10700405 TI - Oil/Alkanethiol Layers for the Study of Emulsified Protein Conformation by Surface Plasmon Resonance Using Monoclonal Antibodies. AB - A method combining surface plasmon resonance and epitope mapping was developed to study the protein conformation at the oil/water interface of an emulsion. The conformation of beta-lactoglobulin stabilizing dodecane/water and miglyol/water interfaces was investigated using five anti-beta-lactoglobulin monoclonal antibodies. The developed method allows us to specifically recognize the emulsified beta-lactoglobulin at the surface of a sensor chip with good repeatability; i.e., standard deviations range between 0.7 and 3.6%. Considering that the monoclonal antibodies, recognizing conformational epitopes, still bind to beta-lactoglobulin at oil/water interfaces, it is concluded that the protein retains a globular conformation. It is shown that the inhibition-binding values of two pairs of Mabs are different for beta-lactoglobulin stabilizing dodecane/water and miglyol/water interfaces. This indicates that the conformations of emulsified beta-lactoglobulin are slightly different according to the nature of the oil phase. Copyright 2000 Academic Press. PMID- 10700406 TI - Electroosmotic Flow of a General Electrolyte Solution through a Fibrous Medium. AB - The electroosmotic flow of a general electrolyte solution through a fibrous medium is modeled theoretically taking the effect of double-layer polarization into account. The result obtained is applicable to an arbitrary level of electrical potential. We show that if the effect of double-layer polarization is neglected using the linearized Poisson-Boltzmann equation will underestimate electroosmotic velocity. The deviation becomes inappreciable, however, if kappaa is either very large or very small, kappa and a being, respectively, the reciprocal Debye length and the radius of a fiber. If the surface potential is high, the variation of electroosmotic velocity as a function of kappaa may exhibit a local maximum and a local minimum, and the larger the porosity of the fibrous medium the lower the level of surface potential for the local extremals to occur. If kappaa is small, the effect of surface potential on the electroosmotic velocity is more significant than that of double-layer polarization, and the reverse is true if kappaa is large. Copyright 2000 Academic Press. PMID- 10700407 TI - Adhesion of Colloidal ZnO Particles on ZnS-Type Phosphor Surfaces. AB - Chemical and physical aspects of the adhesion of colloidal ZnO particles (d(50)=81 nm) on the surface of ZnS-type phosphors have been studied. Here, the green-emitting phosphor ZnS:Cu,Al,Cu (d(50)=5.0 um) applied in TV screens was chosen as model compound. The ZnS material was pretreated in various ways (H(2)O, HCl, H(2)O(2)) and reacted thereafter with a suspension containing colloidal ZnO particles. Analytical investigations (SEM, ESCA) have shown that the adhesion of colloidal ZnO particles is strongly affected by the degree of hydrolysis of the ZnS surface. Electroacoustic investigations (ESA) prove that both types of surfaces, hydrolyzed ZnS as well as colloidal ZnO, are positively charged. Even so, adhesion of ZnO particles is encouraged very much under these conditions, indicating that secondary attractive forces (electrostatic interaction, chemical bonding) determine the amount of colloidal ZnO adhered on a ZnS-type phosphor. Copyright 2000 Academic Press. PMID- 10700408 TI - Interglycolipid Membrane Interactions: pH-Dependent Aggregation of Nonionic Synthetic Glycolipid Vesicles. AB - Large unilamellar vesicles composed of a nonionic synthetic glycolipid, 1,3-di-O phytanyl-2-O-(beta-D-maltotriosyl)glycerol exhibited a pH-dependent aggregation disaggregation process; vesicle aggregation occurred in a lower pH region and vesicle disaggregation occurred in a higher pH region. This process was almost reversible and the aggregation threshold pH increased as NaCl concentration increased. Electrophoretic mobility measurements revealed that the glycolipid vesicles are negatively charged in the range pH 1.6-13. The change in zeta potentials as functions of pH and NaCl concentration could be well described by the Gouy-Chapman expression of the surface charges with an assumption that the interfacial charges arise from the "adsorption" of OH(-) at the vesicle-water interface and the dissociation of hydroxyl groups of the sugar headgroup in a higher pH regime (>pH 10). The pH-dependent aggregation process was reasonably well described by the classical DLVO theory. Thus, the double-layer repulsive forces appear to be a major factor in stabilizing the glycolipid vesicle suspension. Copyright 2000 Academic Press. PMID- 10700409 TI - Polyelectrolyte-Induced Aggregation of Microcrystalline Cellulose: Reversibility and Shear Effects. AB - The polyelectrolyte-induced aggregation of microcrystalline cellulose (MCC) was studied by focused beam reflectance measurement (FBRM) to determine the reversibility of MCC aggregation under high-shear conditions. A correlation was established between the mean chord length output of FBRM probing a high-shear zone with the mean particle size (laser diffraction) of an aliquot extracted from the low-shear bulk mixing zone. Flocs formed by addition of a cationic polyelectrolyte were ruptured by shear forces of mixing and did not reaggregate at low mixing intensities. Flocs formed by addition of both polyelectrolyte and colloidal silica sols were found to reaggregate at low shear quite reversibly following high-shear degradation. The Kolmolgoroff microscale, eta, was determined using a three-compartment mixing model for the FBRM experiments, and the minimum aggregate adhesion forces were calculated to be approximately 3 nN under the experimental mixing conditions. Shear-dependent FBRM studies are also used to estimate the radial dependence of particle adhesion forces within an aggregate. AFM-based surface force measurements between model anionic surfaces (mica and glass beads) showed more reversible adhesion forces in the presence of colloidal silica than with cationic polyelectrolyte only. A descriptive model of the interfaces giving rise to the observed MCC aggregation and adhesion behavior is proposed. Copyright 2000 Academic Press. PMID- 10700410 TI - A New Interpretation of Contact Angle Variations in View of a Recent Analysis of Immersion Calorimetry. AB - The values of the contact angles of different liquids on the same solid are classically interpreted by theories assuming that the vapor adsorption is negligible on solids presenting contact angles. This implies that the solid may be defined by a certain tension, called "critical tension", which is not clearly thermodynamically defined. These assumptions are in disagreement with modern theories of vapor adsorption science. In this paper, it is shown that it is possible to understand the contact angle values using a rigorous derivation of the Young equation, which links the contact angle, the liquid-vapor surface tension of the drop, and the surface tension of the solid in equilibrium with its own vapor. Numerical approximations allow the contact angle variation to be predicted, without use of ad hoc definitions. It is then possible to link vapor adsorption results and contact angle experiments. It is also possible to deduce the surface tension of solids simply and to compute contact angles on powders without using the Washburn method. Copyright 2000 Academic Press. PMID- 10700411 TI - Adsorbed Layers of Ferritin at Solid and Fluid Interfaces Studied by Atomic Force Microscopy. AB - The adsorption of the iron storage protein ferritin was studied by liquid tapping mode atomic force microscopy in order to obtain molecular resolution in the adsorbed layer within the aqueous environment in which the adsorption was carried out. The surface coverage and the structure of the adsorbed layer were investigated as functions of ionic strength and pH on two different charged surfaces, namely chemically modified glass slides and mixed surfactant films at the air-water interface, which were transferred to graphite substrates after adsorption. Surface coverage trends with both ionic strength and pH indicate the dominance of electrostatic effects, with the balance shifting between intermolecular repulsion and protein-surface attraction. The resulting behavior is more complex than that seen for larger colloidal particles, which appear to follow a modified random sequential adsorption model monotonically. The structure of the adsorbed layers at the solid surfaces is random, but some indication of long-range order is apparent at fluid interfaces, presumably due to the higher protein mobility at the fluid interface. Copyright 2000 Academic Press. PMID- 10700412 TI - Determination of Interparticle Forces by Colloidal Particle Scattering: A Simulation Study. AB - Colloidal particle scattering is a recently developed method for the determination of forces between micron-sized particles, which has given promising results. Inversion of the experimental scattering results to retrieve the interaction force requires the development of reliable computer code to match theory and experiment. We review the method used, discuss some operational details, and present some validating results for shear and sedimentation fields that compare favorably with the literature. We have used the simulation to compare particle scattering in a simple shear field for a range of exponential surface forces and discuss the sensitivity of these results to changes in potential parameters. We also discuss the sensitivity in relation to sources of error and their magnitude. In particular, we have incorporated a Brownian dynamics algorithm and compared the level of thermal noise with a simple theoretical formula. We introduce a straightforward means of representing the data and find that order of magnitude changes in the parameters evoke changes in the scattering pattern roughly equivalent to the Brownian noise level for a typical experiment with Pe=120. The results demonstrate the ability of the method, in principle, to distinguish between interaction potentials of different ranges and energy parameters. Copyright 2000 Academic Press. PMID- 10700413 TI - Zeta Potential of Nanobubbles Generated by Ultrasonication in Aqueous Alkyl Polyglycoside Solutions. AB - A simple and convenient method to measure microelectrophoretic mobilities was proposed to determine the zeta potential of nanobubbles generated by ultrasonication. Bubbles in pure water solutions and in aqueous solutions of alkyl polyglycoside (AG) with different alkyl chain lengths and degrees of polymerization in the head group were sonicated with a palladium-coated electrode designed specially by the manufacturer. The zeta potentials of bubbles with ordinary cationic and ionic surfactants are consistent with others' previous results. The average size of the bubbles generated by sonication is in the range of 300 to 500 nm. The zeta potentials of bubbles in both pure water and AG solutions at all pH values are negative. As the chain length of AG increases, zeta potentials significantly decrease at high pH. For nonionic AG, a possible charging mechanism based on known mechanisms is suggested to explain the negative charge, known to be unusual. Even with a very high concentration of H(+) ions in solution the bubbles are charged negatively because the interface is covered with slightly acidic alcohol groups of AGs. At high pH, the less polar the surfactant, the more negative the charge, since nonpolar surfactant molecules induce the adsorption of OH(-) ions, rather than H(+) ions that prefer hydration by water molecules. Copyright 2000 Academic Press. PMID- 10700414 TI - Electroosmotic Phenomena in Fractures. AB - Electroosmotic phenomena in fractures have been investigated in the linear limit for various double-layer thicknesses. The effects of the geometrical parameters were systematically studied for deterministic sinusoidal and random self-affine fractures. The numerical results show a definite influence of the surface amplitude on electroosmotic processes. For self-affine fractures, the roughness or Hurst exponent has a much larger incidence than the correlation parameter between the two surfaces. All the electroosmotic coupling coefficients can be gathered in a single relationship which depends on a characteristic length scale Lambda, applicable to every configuration. Such a length was already found useful for porous media, but the relationship is different. Copyright 2000 Academic Press. PMID- 10700415 TI - Influence of Gelation Temperature and Catalysts on the Mesoporous Structure of Resorcinol-Formaldehyde Aerogels. AB - Resorcinol-formaldehyde (RF) hydrogels were synthesized by sol-gel polycondensation of resorcinol with formaldehyde using a few kinds of catalysts at a gelation temperature of 238 or 332 K. RF aerogels were prepared by supercritical drying with carbon dioxide. The aerogels were characterized by nitrogen adsorption. The experimental results suggest that the gelation temperature does not greatly influence the porous properties and that sodium hydroxide and sodium hydrogencarbonate can be used as catalysts in the preparation of the aerogels. Copyright 2000 Academic Press. PMID- 10700416 TI - Radiotracer Study of the Specific Adsorption of Anions on Metal Oxides in Acid Media: An Experimental Approach. AB - The specific adsorption of anions (HSO(4)(-), H(2)PO(4)(-), and Cl(-)) was studied at Fe(2)O(3), ZnO, and CuO surfaces by the radiotracer technique in strongly acidic medium (1 M HClO(4)). A significant specific adsorption of HSO(4)(-) and H(2)PO(4)(-) ions was found in all cases studied while no measurable adsorption of Cl(-) ions was observed. In the cases of ZnO and CuO, the specific adsorption takes place over the course of continuous dissolution of the oxide. Copyright 2000 Academic Press. PMID- 10700417 TI - Synthesis of Temperature-Sensitive Poly(N-isopropylacrylamide) Hydrogel with Improved Surface Property. AB - A new PNIPA hydrogel was synthesized by carrying out the polymerization in the gelated corn starch aqueous solution. This PNIPA hydrogel has an improved surface property and does not form the disadvantageous bubbles during the shrinking process. This change is due to the hydrogen bonds between the corn starch and the hydrophilic side groups of the PNIPA chains, which let the starch act as the long graft-like chains of the PNIPA hydrogel. During the reswelling process, this PNIPA hydrogel exhibits a sigmoidal swelling pattern. This hydrogel with improved surface property may be very useful for the potential applications of the temperature-sensitive hydrogel. Copyright 2000 Academic Press. PMID- 10700418 TI - Determination of Bulk and Surface Diffusion Coefficients from Experimental Data for Thin Liquid Film Drainage. AB - This note presents a method for the determination of the surface diffusion coefficient and surface diffusion flux. The theoretical considerations are based on the Onsager linear theory for the definition of the surface diffusion flux and on the Einstein theorem for the definition of the surface diffusion parameter. In this interpretation the surface diffusion coefficient differs from the one commonly defined in the literature. It does not depend on the surfactant concentration and it is a function only of the type of surfactant and the liquid/liquid interface. The theoretical calculations indicate that the effect of the surface diffusion on the film drainage is stronger than that predicted by previous theoretical studies. The experimental data for thin liquid film drainage in the case of low surfactant concentration in the continuous phase could be used for the calculation of the bulk and surface diffusion coefficients. In the present study we utilized the experimental data for the drainage of nitrobenzene films stabilized by different concentrations of dodecanol. Copyright 2000 Academic Press. PMID- 10700419 TI - Human papillomavirus testing for triage of women with cytologic evidence of low grade squamous intraepithelial lesions: baseline data from a randomized trial. The Atypical Squamous Cells of Undetermined Significance/Low-Grade Squamous Intraepithelial Lesions Triage Study (ALTS) Group. AB - BACKGROUND AND OBJECTIVE: Human papillomavirus (HPV) infections appear to be central to the development of cervical cancer. This study addresses the question of whether testing women who have low-grade squamous intraepithelial lesions (LSILs) of the uterine cervix for HPV DNA is useful as a triage strategy. METHODS: Four clinical centers in different areas of the United States participated in a randomized clinical trial of the use of HPV DNA testing in women with cytologic evidence of atypical squamous cells of undetermined significance (ASCUS) or LSIL. The study sample in this article consists only of women who had LSIL at enrollment. Within 6 months of an LSIL diagnosis (based on a Pap smear read by a community-based cytopathologist), women who were 18 years of age or older completed a standardized questionnaire and underwent a pelvic examination that included collection of cervical specimens for HPV DNA testing by Hybrid Capture II (HCII)(R) assay. RESULTS: Among the 642 women referred with LSIL who had analyzable test results, the mean chronologic age and age at first coitus were similar among the four clinical centers, despite the centers' ethnic and geographic diversity. Overall, HPV DNA was detected in cervical samples from 532 (82.9%) of the 642 women (95% confidence interval = 79.7%-85.7%). This high frequency of HPV positivity was confirmed by polymerase chain reaction (PCR) assays in a subset of 210 paired specimens tested by HCII and PCR (81.4% were positive by both methods). CONCLUSION: Because a very high percentage of women with an LSIL diagnosis from Pap smears are positive for HPV DNA by HCII testing, there is limited potential for this assay to direct decisions about the clinical management of women with LSIL. The role of HPV testing in the management of women with ASCUS is still under study. PMID- 10700420 TI - Systemic vasculitis: epidemiology, classification and environmental factors. PMID- 10700421 TI - A visible and palpable cause of backache. PMID- 10700422 TI - Fragile without fractures. PMID- 10700423 TI - Lupus-molecular and cellular pathogenesis PMID- 10700424 TI - Immunogenetic differences between patients with familial and non-familial rheumatoid arthritis. AB - OBJECTIVE: To search for possible immunogenetic differencies between the patients with familial and non-familial rheumatoid arthritis (RA). METHODS: The study compared 129 familial RA patients with 217 non-familial patients for the frequencies of HLA-DR antigens including DR4 subtypes, DR4-DQB1*0301 and DR4 DQB1*0302 haplotypes and HLA-B27 antigen as well as the age of disease onset and existence of rheumatoid factor or joint erosions. RESULTS: Two major differences between familial and non-familial groups were found: firstly, familial RA patients had increased frequency of HLA-DR4 as compared with the non-familial RA group (68.2 v. 54.8%; p = 0.019). Secondly, the mean age at onset of RA was significantly lower in the familial than in the sporadic RA patients (42.0 v. 46.5 years; p = 0.0020) and the difference still remained when the DR4 positive and negative subgroups were compared separately. CONCLUSION: These results confirm the more prominent association with HLA-DR4 in familial than in the non familial cases and suggest that accumulation of HLA risk genes may, at least partly, explain the familial occurrence of the disease. Other susceptibility genes may also be concentrated in multiplex case families as suggested by an earlier age at the onset of RA in both HLA-DR4 positive and negative familial patients. PMID- 10700425 TI - Sonography for hip joint effusion in adults with hip pain. AB - OBJECTIVE: To study the prevalence of ultrasonic hip joint effusion and its relation with clinical, radiological and laboratory (ESR) findings in adults with hip pain. METHODS: Patients (n = 224) aged 50 years or older with hip pain, referred by the general practitioner for radiological investigation, underwent a standardised examination. The distance between the ventral capsule and the femoral neck, an increase in which represents joint effusion, was measured sonographically. Joint effusion was defined in three different ways: "effusion" according to Koski's definition, "major effusion", and "asymmetrical effusion" based on only individual side differences. RESULTS: "Effusion" was present in 80 (38%), "major effusion" in 20 (9%), and "asymmetrical effusion" in 47 (22%) patients. Pain in the groin or medial thigh, pain aggravated by lying on the side, decreased extension/internal rotation/abduction/flexion, painful external rotation, and pain on palpation in the groin showed a significant relation (adjusted for age and radiological osteoarthritis of the hip) with ultrasonic hip joint effusion. "Major effusion" showed a significant relation with an increased ESR. When patients with bilateral pain and increased ESR were excluded, a side difference in the range of motion of extension of the hip was shown to be a good predictor for "asymmetrical effusion" (positive predictive value: 71%, negative predictive value: 80%). CONCLUSION: This study showed a relatively high prevalence of ultrasonic joint effusion in adults with hip pain in general practice. Furthermore the results indicate a relation between joint effusion and clinical signs. PMID- 10700426 TI - Mast cells, extracellular matrix components, TGFbeta isoforms and TGFbeta receptor expression in labial salivary glands in systemic sclerosis. AB - OBJECTIVE: To determine whether there was altered elaboration of non-collagenous extracellular matrix proteins or expression of TGFbeta isoforms and their receptors in salivary glands of patients with systemic sclerosis (SSc) and Raynaud's phenomenon (RP). Because of the possible role of mast cells in the early stages of SSc their presence was also investigated. METHODS: Sections of normal labial salivary glands (n=10) and glands from patients with SSc (n = 13) and RP (n = 5) were stained immunohistochemically and using acid toluidine blue. RESULTS: SSc glands contained more mast cells than control tissues (p<0.005) and similar numbers to those found in RP specimens. There were no differences in the pattern or amount of non-collagenous matrix proteins detected. Tenascin and elastin were predominantly found surrounding ducts whereas fibronectin had a more general distribution. TGFbeta isoforms and receptors were expressed by glandular epithelium, fibroblasts, vascular endothelium and inflammatory cells. Cell counts showed no differences in expression of TGFbeta1 or TGFbeta receptors between groups. However, the percentage of TGFbeta2 positive fibroblasts was significantly higher in SSc glands compared with controls (p<0.004). RP glands showed an intermediate level of expression. By contrast, a lower percentage of RP fibrolasts expressed TGFbeta3 compared with controls with SSc glands showing an intermediate level of expression. CONCLUSIONS: These results show that (a) there are no changes in glandular expression of tenascin, elastin and fibronectin in SSc and RP and (b) both conditions are associated with an increased salivary gland mast cell population and changes in expression of TGFbeta2 and beta3 isoforms by glandular fibroblasts. PMID- 10700427 TI - Increased expression of CD40 ligand (CD154) on CD4+ T cells as a marker of disease activity in rheumatoid arthritis. AB - OBJECTIVES: The interaction between the activation induced surface glycoprotein CD40L (ligand) (CD154) on CD4+ T cells and its receptor CD40, which is expressed on various cell types, plays a crucial part in numerous cell mediated and humoral immune reactions that may be of pathogenetic importance in rheumatoid arthritis (RA). To further evaluate the pathogenetic role of CD40L in RA, expression of CD40L and various other T cell activation antigens as well as costimulatory molecules was investigated on CD4+ T cells in RA by flow cytometry. METHODS: Two colour flow cytometry was used to determine the percentage of CD4+ T cells expressing CD40L, CD69, CD25, HLA-DR, CD39, CD27 and CD28 in peripheral blood (PB) of 62 RA patients in comparison to 20 healthy controls (HC). Disease activity was assessed by clinical, laboratory and radiological examination. Status of clinical remission of RA was evaluated according to the ACR preliminary criteria for complete clinical remission of RA. RESULTS: CD40L was expressed on > 10% of CD4+ T cells in 29% of RA patients thus defining a CD40L(high+) patient group. Disease activity as estimated by C reactive protein, rheumatoid factor and status of clinical remission of disease (p = 0.049) was higher in this subgroup than in the RA CD40L(low+) group. Expression of CD69, CD25, and HLA-DR was significantly increased in both RA patient groups in comparison with HC. However, the percentage of CD39+ CD4+ T cells was increased only in the RA CD40L(high+) subgroup (versus HC p = 0.019, versus RA CD40L(low+) p = 0.044). Furthermore, expression of CD40L and CD39 on CD4+ T cells correlated positively as estimated by Spearman rank correlation (p<0.001). The percentage of CD4+ T cells lacking the costimulatory molecules CD27 (p = 0.002) and CD28 (p = 0.026) was increased in RA CD40L(low+) patients in comparison with HC. CONCLUSIONS: These data suggest that increased expression of CD40L on CD4+ T cells in RA indicates prolonged and increased activation of CD4+ T lymphocytes and is associated with active disease and possibly an unfavourable prognosis. Whether this phenotypically defined RA CD40L(high+) subgroup will preferentially respond to an anti-CD40L antibody treatment remains to be elucidated. PMID- 10700428 TI - Blockade of endogenous interleukin 12 results in suppression of murine streptococcal cell wall arthritis by enhancement of interleukin 10 and interleukin 1Ra. AB - OBJECTIVE: The goal of this study was to investigate the role of endogenous interleukin 12 (IL12) in acute murine streptococcal cell wall (SCW) arthritis. METHODS: C57black/6 mice were injected intraperitoneally with rat anti-murine IL12 (C17.8), shortly before induction of arthritis by intra-articular injection of 25 microg SCW fragments into the right knee joint. Joint swelling and chondrocyte synthetic function was analysed several days after induction of SCW arthritis. Local cytokine profile was determined, protein by using ELISA and mRNA by RT-PCR technology. To confirm the findings at later time points, tissue chamber model of inflammation was used. Histology was performed to examine cell influx and cartilage damage. RESULTS: Suppression of joint swelling was noted at days 2 and 4, whereas no suppressive effect of anti-IL12 was found at day 1. Severe inhibition of chondrocyte proteoglycan synthesis was seen at day 1 in both arthritic control and anti-IL12 treated mice. However, chondrocyte function was restored at day 4 of arthritis in the anti-IL12 injected animals, but not in the arthritic controls. Moreover, cell influx in synovial tissue and joint cavity was reduced by anti-IL12 treatment. Neutralisation of IL12 reduced the local levels of IL1beta, IL12 and interferon gamma, when examined shortly after induction of SCW arthritis, whereas tumour necrosis factor alpha levels were not affected. In contrast, IL10 and IL1Ra protein and mRNA levels were strongly up regulated in synovial tissues after IL12 blockade. Enhancement of IL10 and IL1Ra by anti-IL12 was confirmed in a tissue chamber model with SCW induced inflammation. CONCLUSIONS: This study indicates that IL12 is a pro-inflammatory cytokine during onset of acute SCW arthritis. Balances of proinflammatory and anti-inflammatory cytokines were strongly improved by anti-IL12 treatment. PMID- 10700429 TI - Prevalence of Raynaud's phenomenon in a healthy Greek population. AB - OBJECTIVE: Raynaud's phenomenon (RP) is comprised of repeated episodes of colour changes of the skin of digits on cold exposure or emotional stress. The prevalence of RP in the general population is variable fluctuating between 4% 15%, among surveys. The aim of this study was to estimate the prevalence of RP in a healthy working Greek population and to investigate the possible association of RP with various demographic, social and other factors. METHODS: A total of 756 employees of the University Hospital of Ioannina was included in the study. They belong to the administrative (120 subjects), nursing and technical (a representative sample of 418 and 218 subjects, respectively) personnel. Five hundred subjects (111 men and 389 women) responded in a face to face interview based on a specially conformed questionnaire. The study began in November 1997 and was completed in March 1998. RESULTS: Twenty six subjects with RP (1 man and 25 women) were found. Their mean (SD) age was 32.73 (5.77) years. The prevalence of RP was 5.2% (0.9% in men and 6.4% in women). The sex ratio, male/female, was 1/7.1. An association between RP and migraine was found. However, there were no significant correlations of RP with smoking, alcohol and coffee consumption, dietary habits, occupational history and drug exposure. No social or other demographic parameters associated to RP frequency were found. CONCLUSIONS: The prevalence of RP (5.2%) in the population studied is relatively low compared with previous studies. RP focuses on the fourth decade of life and affects mainly women. There was no evidence of any correlation of RP with social, environmental or personal parameters while an association of RP with migraine was found. Geographical or genetic factors, or both, may be responsible for these results. PMID- 10700430 TI - Subclinical gut inflammation in spondyloarthropathy patients is associated with upregulation of the E-cadherin/catenin complex. AB - OBJECTIVE: Previously an upregulation of E-cadherin and its associated molecules alpha-catenin, beta-catenin and plakoglobin has been demonstrated in clinically overt inflammatory bowel disease (IBD). The aim of this study was to investigate the expression of the E-cadherin/catenin complex in subclinically inflamed bowel mucosa from spondyloarthropathy (SpA) patients. METHODS: Ileal and colonic biopsy specimens from 19 SpA patients with subclinical inflammatory gut lesions and from seven controls were stained with monoclonal antibodies against E-cadherin, beta catenin and plakoglobin and a polyclonal antibody against alpha-catenin. E cadherin mRNA was detected using a riboprobe. Inflammation was histologically classified into acute, chronic active and chronic quiescent forms. RESULTS: In acute and chronic active bowel inflammation of SpA patients, upregulation of the E-cadherin/catenin glycoprotein complex could be observed. Chronic lesions in a quiescent state did not show such an upregulation. Furthermore, chronic inflammation was associated with an increase in E-cadherin mRNA. CONCLUSIONS: As some of the SpA patients with subclinical gut inflammation develop IBD, upregulation of the E-cadherin/catenin complex in inflamed bowel mucosa from SpA patients may point to early cellular changes in the development of IBD. However, at present it cannot be excluded that increased E-cadherin/catenin complex expression is a bystander phenomenon of active inflammation. PMID- 10700431 TI - Anaemia in systemic lupus erythematosus: aetiological profile and the role of erythropoietin. AB - OBJECTIVE: To study the prevalence of different causes of anaemia in patients with systemic lupus erythematosus (SLE) and their associations with immunological and clinical parameters and to evaluate the contribution of erythropoietin (Epo) and anti-erythropoietin (anti-Epo) autoantibodies to the development of SLE anaemia. METHODS: 132 SLE patients with anaemia (defined as haemoglobin of 12 g/dl or less for women and 13.5 g/dl or less for men) from among a total of 345 consecutive SLE patients were prospectively enrolled into the study. Standard haematological and immunological tests were performed and serum Epo and anti-Epo antibodies were assayed. RESULTS: The identified causes were anaemia of chronic disease (ACD) n=49 (37.1%), iron deficiency anaemia (IDA) n = 47 (35.6%), autoimmune haemolytic anaemia (AHA) n = 19 (14.4%) and other causes n = 17 (12.9%). There was significant heterogeneity in the severity of anaemia between the four groups (p<0.01) with AHA cases being on average more severe. The proportion of patients with anticardiolipin antibodies, low complement levels and anti-dsDNA differed significantly among the four groups; these markers were particularly common in patients with AHA, and uncommon in patients with IDA. Twenty one of 100 tested patients had anti-Epo antibodies. Such antibodies were seen practically only in patients with ACD (odds ratio 3.1, p = 0.041) and in patients with high lupus activity (ECLAM) scores (odds ratio 1.27 per point, p = 0.055). Epo response was inadequate in 42.4% and 41.2% of patients with ACD and AHA, respectively. CONCLUSIONS: Anaemia in SLE usually takes the form of ACD and IDA, however autoimmune haemolysis is not uncommon. SLE patients with different causes of anaemia differ in regard to several immunological parameters. Epo response is blunted in anaemic SLE patients, particularly those with ACD and AHA. PMID- 10700432 TI - Predictors of functional status in patients with early rheumatoid arthritis. AB - OBJECTIVE: To find disease parameters that can predict the functional capacity of patients with early rheumatoid arthritis (RA) at the first visit to the rheumatologist and one year after entry. METHODS: Patients referred to the outpatients clinic between 1995 and 1996, with a symptom duration of less than three years and fulfilling the American Rheumatism Association 1987 revised criteria for RA within one year after entry were included. Assessments of the duration of morning stiffness, the Disease Activity Score (DAS: a composite score based on erythrocyte sedimentation rate (ESR), number of painful and swollen joints and patient global assessment), pain (Visual Analogue Scale), the Arthritis Impact Measurement Scale (AIMS) and the Health Assessment Questionnaire (HAQ) were performed every three months. Possible predictors of the HAQ at entry and after one year were analysed by logistic regression. RESULTS: 133 patients were included in the study. The median duration of complaints was three months (range 0-35) and the median HAQ score at entry was 1.12 (range 0-3). There was no correlation between duration of complaints and the HAQ at entry (r = 0.01). An HAQ score under the 50th percentile at entry could be predicted correctly for 74% of the patients by entry DAS and C reactive protein concentration, and at one year could be predicted correctly for 73% of the patients by entry HAQ and pain score. CONCLUSION: Disease activity is strongly correlated with a lower functional capacity at entry, whereas disease duration is not. The functional status at entry is a good predictor for functional status at one year. Severity rather than duration of arthritis prompts referral in this cohort. PMID- 10700433 TI - Diastolic function abnormalities in rheumatoid arthritis. Evaluation By echo Doppler transmitral flow and pulmonary venous flow: relation with duration of disease. AB - OBJECTIVE: The aim of this study was to evaluate left ventricular filling in patients with rheumatoid arthritis (RA), analysing transmitral flow and pulmonary venous flow, with special regard to age and disease duration. METHODS: 32 patients affected by RA according to ARA criteria were selected, without evidence of cardiac disease, and compared with matched control subjects. All patients and the control group were submitted to M-mode, two dimensional, Doppler and colour Doppler (continuous and pulsed wave) echocardiography. The following diastolic parameters were evaluated: transmitralic flow (E/A ratio), pulmonary venous flow (S/D ratio), a-Pw, IVRT and DT. RESULTS: In RA patients left ventricular filling abnormalities were found characterised by a reduced E/A ratio (mean (SD) 1.16 (0.31) v. controls 1.37 (0.32); p = 0.02) and an increased S/D ratio (1.43 (0.40) v. controls 1.22 (0.29); p = 0.017). In the group of patients a relation was found between E/A ratio and disease duration (r= 0.40, p = 0.01 Spearman rank correlation). CONCLUSIONS: At present, it is concluded that RA patients, in absence of clinical evidence of heart disease, show diastolic dysfunction characterised by impaired E/A and S/D ratio. The relation between transmitral flow alteration and disease duration suggests a sub-clinical myocardial involvement. PMID- 10700434 TI - Development of additional autoimmune diseases in a population of patients with systemic lupus erythematosus. AB - OBJECTIVE: In view of the recognised clustering of autoimmune diseases (AID), the chronology of development of other autoimmune diseases in systemic lupus erythematosus (SLE) patients was considered. METHODS: A retrospective review of a well documented population of 215 patients with SLE was undertaken. The duration of follow up ranged from 2 to 18 years. RESULTS: Of these 215 patients, 65 (30%) had at least one other AID-including 51 (24%) having one other AID, 12 (6%) having two and two (1%) having three other AID in addition to their SLE. Twelve different autoimmune diseases were identified. The majority of patients developed a further AID after SLE had been diagnosed (62%) reflecting the relatively early age of onset of SLE. There was no significant difference in the age of onset of rheumatoid arthritis, Sjogren's syndrome and hypothyroidism that had developed before SLE compared with those who developed these diseases after SLE. However, those who developed autoimmune thrombocytopenia (AITP) before SLE were significantly younger than those who developed AITP after SLE (16.7 v. 38.7 years respectively, p<0.05). CONCLUSIONS: Physicians caring for SLE patients should remain alert to the possible development of a second AID during follow up. Further well matched case-control studies are required to define the exact relation between SLE and other AID. PMID- 10700435 TI - Importance of synovial fluid aspiration when injecting intra-articular corticosteroids. AB - OBJECTIVE: The aim of this prospective study was to find if a complete synovial fluid aspiration before injecting intra-articular corticosteroids influences the treatment result. METHODS: The study was performed in 147 patients with rheumatoid arthritis (RA). One hundred and ninety one knees with synovitis were randomised to arthrocentesis (n=95) or no arthrocentesis (n = 96) before 20 mg triamcinolone hexacetonide was injected. The duration of effect was followed up for a period of six months. All patients were instructed to contact the rheumatology department if signs and symptoms from the treated knee recurred. If arthritis could be confirmed by a clinical examination a relapse was noted. RESULTS: There was a significant reduction of relapse in the arthrocentesis group (p = 0.001). CONCLUSION: The study shows that aspiration of synovial fluid can reduce the risk for arthritis relapse when treating RA patients with intra articular corticosteroids. It is concluded that arthrocentesis shall be included in the intra-articular corticosteroid injection procedure. PMID- 10700436 TI - Autologous stem cell transplantation in a case of treatment resistant central nervous system lupus. AB - This case report describes a young woman with systemic lupus erythematosus starting at 16 years of age and giving rise to severe neurological complications including bilateral opticus neuritis and transverse myelitis. Despite heavy immunosuppression her condition steadily aggravated. At this point it was decided to perform autologous stem cell transplantation. Haematopoietic stem cells were mobilised with cyclophosphamide and granulocyte colony stimulating factor. Enrichment of CD34(+)cells was followed by depletion of peripheral T and B cells. The post-transplantation course was uneventful, and all the neurological deficits improved promptly during the 15 months of follow up. This is the first description of successful autologous stem cell transplantation in a case of life threatening central nervous system lupus. PMID- 10700437 TI - How could a genetic variant of the p22(phox) component of NAD(P)H oxidases contribute to the progression of coronary atherosclerosis? PMID- 10700438 TI - Anchors Aweigh!: ion channels, cytoskeletal proteins, and cellular excitability. PMID- 10700439 TI - Fibrillating myocardium : rabbit warren or beehive? PMID- 10700440 TI - Toward antiapoptosis as a new treatment modality. PMID- 10700442 TI - Myosin heavy chain isoform expression in the failing and nonfailing human heart. AB - In the heart, the relative proportions of the 2 forms of the motor protein myosin heavy chain (MyHC) have been shown to be affected by a wide variety of pathological and physiological stimuli. Hearts that express the faster MyHC motor protein, alpha, produce more power than those expressing the slower MyHC motor protein, beta, leading to the hypothesis that MyHC isoforms play a major role in the determination of cardiac contractility. We showed previously that a significant amount of alphaMyHC mRNA is expressed in nonfailing human ventricular myocardium and that alphaMyHC mRNA expression is decreased 15-fold in end-stage failing left ventricles. In the present study, we determined the MyHC protein isoform content of human heart samples of known MyHC mRNA composition. We demonstrate that alphaMyHC protein was easily detectable in 12 nonfailing hearts. alphaMyHC protein represented 7.2+/-3.2% of total MyHC protein (compared with approximately 35% of the MyHC mRNA), suggesting that translational regulation may be operative; in contrast, there was effectively no detectable alphaMyHC protein in the left ventricles of 10 end-stage failing human hearts. PMID- 10700441 TI - Selective upregulation of cardiac endothelin system in patients with ischemic but not idiopathic dilated cardiomyopathy: endothelin-1 system in the human failing heart. AB - Only scarce information is available on the activity and modifications of the cardiac endothelin (ET)-1 system in heart failure due to ischemic (ICM) or idiopathic dilated (DCM) cardiomyopathy. The activity of the ET-1 system was investigated by measuring cardiac ET-1 and big ET-1 formation and quantifying cardiac mRNA for prepro-ET-1 (ppET-1), ET-converting enzyme-1, and ET(A) and ET(B) receptors both in myocardium and in isolated myocytes using Northern blot, reverse transcription-polymerase chain reaction, and in situ hybridization in 22 patients with DCM and 20 with ICM who underwent cardiac transplantation and in 7 potential heart transplant donors (nonfailing hearts). Notwithstanding a similar increase of plasma ET-1 in the 2 groups, cardiac ET formation, mRNA levels for ppET-1, and ET(A) and ET(B) receptors were higher on both the myocardium and isolated myocytes from ICM than on those from DCM hearts (P<0.001 for all). ppET 1 and ET-converting enzyme-1 mRNAs were expressed on myocytes and endothelial and interstitial cells in ICM, whereas in DCM and nonfailing hearts they were mainly expressed on nonmyocyte cells. In both ICM and DCM, the ET(A) mRNA signal was expressed on both myocytes and nonmyocyte cells, whereas ET(B) mRNA was almost exclusively localized on nonmyocyte cells. ET(A)- and ET(B)-specific receptor binding was increased on both myocytes and cardiac membranes, showing a positive correlation with left ventricular ejection fraction in ICM (r=0.78 and 0.70) but not in DCM patients. The present results show that human ventricular myocytes express all of the components of the ET-1 system, which is selectively upregulated in ICM patients and appears to be functionally important in the maintenance of cardiac function. PMID- 10700443 TI - A variant of p22(phox), involved in generation of reactive oxygen species in the vessel wall, is associated with progression of coronary atherosclerosis. AB - A series of pro-oxidant and antioxidant enzymes, such as the NADPH oxidase system, maintain the redox state in the vessel wall. A major component of NADPH oxidase is p22(phox), which is implicated in atherosclerosis. We prospectively studied the association of the histidine (H)(72)-->tyrosine (Y) mutation in p22(phox) with the severity and progression/regression of coronary artery disease (CAD), plasma lipid levels, clinical events, and response to treatment with fluvastatin in a well-characterized population. Genotypes were determined by polymerase chain reaction and restriction digestion with RsaI enzyme in 368 subjects in the Lipoprotein and Coronary Atherosclerosis Study (LCAS). Fasting plasma lipids and quantitative coronary angiograms were obtained at baseline and 2.5 years after randomization to fluvastatin or placebo. Subjects with CC genotype (n=157) were identified by the presence of 396-bp and 113-bp products on gel electrophoresis. Those with TT (n=39) were identified by the presence of 316 bp, 113-bp, and 80-bp products, and those with CT (n=172) by the presence of 396 bp, 316-bp, 113-bp, and 80-bp products. Baseline and final plasma levels of lipids and the baseline severity of CAD were not significantly different among the genotypes. In the placebo group, subjects with the mutation had a 3- to 5 fold greater loss in mean minimum lumen diameter (MLD) (TT: -0.15+/-0.15; CT: 0.17+/-0.26; and CC: -0.03+/-0.22 mm; P=0. 006) and lesion-specific MLD (TT: 0.15+/-0.06; CT: -0.18+/-0.03; and CC: -0.06+/-0.03 mm; P=0.038) than those without. Progression was also more (TT: 8/17 [47%]; CT: 35/73 [48%]; and CC: 17/62 [27%]) and regression less (TT: 0/17 [0%]; CT: 1/73 [1%]; and CC: 11/72 [18%]) common in those with the mutation (P=0.002). The C(242)T mutation in p22(phox), involved in maintaining the redox state in the vessel wall, is associated with progression of coronary atherosclerosis in the LCAS population. PMID- 10700444 TI - Enhanced dispersion of repolarization and refractoriness in transgenic mouse hearts promotes reentrant ventricular tachycardia. AB - The heterogeneous distribution of ion channels in ventricular muscle gives rise to spatial variations in action potential (AP) duration (APD) and contributes to the repolarization sequence in healthy hearts. It has been proposed that enhanced dispersion of repolarization may underlie arrhythmias in diseases with markedly different causes. We engineered dominant negative transgenic mice that have prolonged QT intervals and arrhythmias due to the loss of a slowly inactivating K(+) current. Optical techniques are now applied to map APs and investigate the mechanisms underlying these arrhythmias. Hearts from transgenic and control mice were isolated, perfused, stained with di-4-ANEPPS, and paced at multiple sites to optically map APs, activation, and repolarization sequences at baseline and during arrhythmias. Transgenic hearts exhibited a 2-fold prolongation of APD, less shortening (8% versus 40%) of APDs with decreasing cycle length, altered restitution kinetics, and greater gradients of refractoriness from apex to base compared with control hearts. A premature impulse applied at the apex of transgenic hearts produced sustained reentrant ventricular tachycardia (n=14 of 15 hearts) that did not occur with stimulation at the base (n=8) or at any location in control hearts (n=12). In transgenic hearts, premature impulses initiated reentry by encountering functional lines of conduction block caused by enhanced dispersion of refractoriness. Reentrant VT had stable (>30 minutes) alternating long/short APDs associated with long/short cycle lengths and T wave alternans. Thus, optical mapping of genetically engineered mice may help elucidate some electrophysiological mechanisms that underlie arrhythmias and sudden death in human cardiac disorders. PMID- 10700445 TI - Distribution of excitation frequencies on the epicardial and endocardial surfaces of fibrillating ventricular wall of the sheep heart. AB - Tissue heterogeneities may play an important role in the mechanism of ventricular tachycardia (VT) and fibrillation (VF) and can lead to a complex spatial distribution of excitation frequencies. Here we used optical mapping and Fourier analysis to determine the distribution of excitation frequencies in >20 000 sites of fibrillating ventricular tissue. Our objective was to use such a distribution as a tool to quantify the degree of organization during VF. Fourteen episodes of VT/VF were induced via rapid pacing in 9 isolated, coronary perfused, and superfused sheep ventricular slabs (3x3 cm(2)). A dual-camera video-imaging system was used for simultaneous optical recordings from the entire epi- and endocardial surfaces. The local frequencies of excitation were determined at each pixel and displayed as dominant frequency (DF) maps. A typical DF map consisted of several (8.2+/-3.6) discrete areas (domains) with a uniform DF within each domain. The DFs in adjacent domains were often in 1:2, 3:4, or 4:5 ratios, which was shown to be a result of an intermittent Wenckebach-like conduction block at the domain boundaries. The domain patterns were relatively stable and could persist from several seconds to several minutes. The complexity in the organization of the domains, the number of domains, and the dispersion of frequencies increased with the rate of the arrhythmia. Domain patterns on the epicardial and endocardial surfaces were not correlated. Sustained epicardial or endocardial reentry was observed in only 3 episodes. Observed frequency patterns during VT/VF suggest that the underlying mechanism may be a sustained intramural reentrant source interacting with tissue heterogeneities. PMID- 10700446 TI - Kallistatin stimulates vascular smooth muscle cell proliferation and migration in vitro and neointima formation in balloon-injured rat artery. AB - Kallistatin, a serine proteinase inhibitor (serpin), is expressed in the endothelial and smooth muscle cells of blood vessels. The potential function of kallistatin in vascular biology was investigated by studying its role in the proliferation and migration of cultured primary aortic vascular smooth muscle cells (VSMCs) in vitro and in neointima formation in rat artery after balloon angioplasty in vivo. Exogenous kallistatin induced a >2-fold increase of VSMC proliferation and cell growth as measured by [(3)H]thymidine incorporation and cell counts and a 2.3-fold increase of cell migration in modified Boyden chambers. In balloon-injured vessels, endogenous kallistatin mRNA and protein levels increased up to 10-fold as determined by competitive polymerase chain reaction and by ELISA. Intense staining of kallistatin mRNA was identified in the proliferating VSMCs of balloon-injured arteries during cell migration from media to neointima by in situ hybridization histochemistry and immunohistochemistry. We observed an induction of kallistatin expression by platelet-derived growth factor (PDGF) and upregulation of p42/44 mitogen-activated protein kinase (MAPK) activity by kallistatin in cultured VSMCs. Conversely, adenovirus-mediated transfer of kallistatin antisense cDNA into cultured VSMCs inhibited PDGF-induced p42/44 MAPK activity and cell proliferation. Furthermore, local delivery of adenovirus carrying kallistatin antisense cDNA significantly downregulated kallistatin mRNA levels and attenuated neointima formation in balloon-injured rat arteries in vivo. These results indicate that kallistatin may play an important role in mediating PDGF-induced MAPK pathway on VSMC proliferation and in neointima formation after balloon angioplasty. PMID- 10700447 TI - Lateral zone of cell-cell adhesion as the major fluid shear stress-related signal transduction site. AB - It has been proposed previously that actin filaments and cell adhesion sites are involved in mechanosignal transduction. In this study, we present certain morphological evidence that supports this hypothesis. The 3D disposition of actin filaments and phosphotyrosine-containing proteins in endothelial cells in situ was analyzed by using confocal microscopy and image reconstruction techniques. Surgical coarctations were made in guinea pig aortas, and the same 3D studies were conducted on such areas 1 week later. Stress fibers (SFs) were present at both basal and apical regions of endothelial cells regardless of coarctation, and several phosphotyrosine-containing proteins were associated with SF ends. Apical SFs had one end attached to the apical cell membrane and the other attached to either the basal membrane or the lateral cell border. Within the coarctation area, the actin filament-containing and vinculin-containing structures became prominent, especially at the apical and the lateral regions. Substantially higher levels of anti-phosphotyrosine and anti-Src staining were detected in the constricted area, particularly at the cell-cell apposition, whereas the anti focal adhesion kinase, anti-CT10-related kinase, anti-platelet endothelial cell adhesion molecule-l, anti-vinculin, and phalloidin staining intensities increased only slightly after coarctation. We propose that apical SFs directly transmit the mechanical force of flow from the cell apex to the lateral and/or basal SF anchoring sites and that the SF ends associated with signaling molecules are sites of signal transduction. Our results support the idea that the cell apposition area is the major fluid shear stress-dependent mechanosignal transduction site in endothelial cells. PMID- 10700448 TI - Reduced neointima hyperplasia of vein bypass grafts in intercellular adhesion molecule-1-deficient mice. AB - Recently, we established a new mouse model of vein graft arteriosclerosis through the grafting of vena cava to carotid arteries. In many respects, the morphological features of this murine vascular graft model resemble those of human venous bypass graft disease. With this model, we studied the role of intercellular adhesion molecule-1 (ICAM-1) in the development of vein graft arteriosclerosis in ICAM-1-deficient mice. Neointimal hyperplasia of vein grafts in ICAM-1 -/- mice was reduced 30% to 50% compared with that of wild-type control animals. Immmunofluorescent analysis revealed that increased ICAM-1 expression was observed on the endothelium and smooth muscle cells (SMCs) of the grafted veins in wild-type, but not ICAM-1 -/-, mice. MAC-1 (CD11b/18)-positive cells that adhered to the surface of vein grafts in ICAM-1 -/- mice were significantly less as identified with en face immunofluorescence, and these positive cells were more abundant in the intimal lesions of vein grafts in wild-type mice. Furthermore, aortic SMCs cultivated from wild-type mice exhibited high ICAM-1 expression in response to tumor necrosis factor-alpha. When tumor necrosis factor alpha-stimulated SMCs were incubated with mouse spleen leukocytes, the number of cells that adhered to ICAM-1 -/- SMCs was significantly lower than the number that adhered to ICAM-1 +/+ SMCs, which was markedly blocked through pretreatment of leukocytes with the anti-MAC-1 antibody. Taken together, our findings demonstrate that ICAM-1 is critical in the development of venous bypass graft arteriosclerosis, which provides essential information for therapeutic intervention for vein graft disease in patients undergoing bypass surgery. PMID- 10700449 TI - Abnormal cardiac Na(+) channel properties and QT heart rate adaptation in neonatal ankyrin(B) knockout mice. AB - The cytoskeleton of the cardiomyocyte has been shown to modulate ion channel function. Cytoskeletal disruption in vitro alters Na(+) channel kinetics, producing a late Na(+) current that can prolong repolarization. This study describes the properties of the cardiac Na(+) channel and cardiac repolarization in neonatal mice lacking ankyrin(B), a cytoskeletal "adaptor" protein. Using whole-cell voltage clamp techniques, I(Na) density was lower in ankyrin(B)(-/-) ventricular myocytes than in wild-type (WT) myocytes (-307+/-26 versus -444+/-39 pA/pF, P<0.01). Ankyrin(B)(-/-) myocytes exhibited a hyperpolarizing shift in activation and inactivation kinetics compared with WT. Slower recovery from inactivation contributed to the negative shift in steady-state inactivation in ankyrin(B)(-/-). Single Na(+) channel mean open time was longer in ankyrin(B)(-/ ) versus WT at test potentials (V(t)) of -40 mV (1.0+/-0.1 versus 0. 61+/-0.04 ms, P<0.05) and -50 mV (0.8+/-0.1 versus 0.39+/-0.05 ms, P<0.05). Ankyrin(B)(-/-) exhibited late single-channel openings at V(t) -40 and -50 mV, which were not seen in WT. Late I(Na) contributed to longer action potential durations measured at 90% repolarization (APD(90)) at 1 Hz stimulation in ankyrin(B)(-/-) compared with WT (354+/-26 versus 274+/-22 ms, P<0.05). From ECG recordings of neonatal mice, heart rates were slower in ankyrin(B)(-/-) than in WT (380+/-14 versus 434+/-13 bpm, P<0.01). Although the QT interval was similar in ankyrin(B)(-/-) and WT at physiological heart rates, QT-interval prolongation in response to heart rate deceleration was greater in ankyrin(B)(-/-). In conclusion, Na(+) channels in ankyrin(B)(-/-) display reduced I(Na) density and abnormal kinetics at the whole-cell and single-channel level that contribute to prolonged APD(90) and abnormal QT-rate adaptation. PMID- 10700450 TI - Ca(2+) transients and Ca(2+) waves in purkinje cells : role in action potential initiation. AB - Purkinje cells contain sarcoplasmic reticulum (SR) directly under the surface membrane, are devoid of t-tubuli, and are packed with myofibrils surrounded by central SR. Several studies have reported that electrical excitation induces a biphasic Ca(2+) transient in Purkinje fiber bundles. We determined the nature of the biphasic Ca(2+) transient in aggregates of Purkinje cells. Aggregates (n=12) were dispersed from the subendocardial Purkinje fiber network of normal canine left ventricle, loaded with Fluo-3/AM, and studied in normal Tyrode's solution (24 degrees C). Membrane action potentials were recorded with fine-tipped microelectrodes, and spatial and temporal changes in [Ca(2+)](i) were obtained from fluorescent images with an epifluorescent microscope (x20; Nikon). Electrical stimulation elicited an action potential as well as a sudden increase in fluorescence (L(0)) compared with resting levels. This was followed by a further increase in fluorescence (L(1)) along the edges of the cells. Fluorescence then progressed toward the Purkinje cell core (velocity of propagation 180 to 313 microm/s). In 62% of the aggregates, initial fluorescent changes of L(0) were followed by focally arising Ca(2+) waves (L(2)), which propagated at 158+/-14 microm/s (n=13). Spontaneous Ca(2+) waves (L(2)*) propagated like L(2) (164+/-10 microm/s) occurred between stimuli and caused slow membrane depolarization; 28% of L(2)* elicited action potentials. Both spontaneous Ca(2+) wave propagation and resulting membrane depolarization were thapsigargin sensitive. Early afterdepolarizations were not accompanied by Ca(2+) waves. Action potentials in Purkinje aggregates induced a rapid rise of Ca(2+) through I(CaL) and release from a subsarcolemmal compartment (L(0)). Ca(2+) release during L(0) either induced further Ca(2+) release, which propagated toward the cell core (L(1)), or initiated Ca(2+) release from small regions and caused L(2) Ca(2+) waves, which propagated throughout the aggregate. Spontaneous Ca(2+) waves (L(2)*) induce action potentials. PMID- 10700451 TI - alpha(v)beta(3) integrin induces tyrosine phosphorylation-dependent Ca(2+) influx in pulmonary endothelial cells. AB - The endothelial alpha(v)beta(3) integrin occurs luminally, where its ligation by soluble agents may induce inflammatory signaling. We tested this hypothesis in bovine pulmonary artery endothelial cell monolayers with the use of vitronectin and cross-linking antibodies to ligate and aggregate the integrin. We quantified the endothelial cytosolic Ca(2+) concentration ([Ca(2+)](i)) according to the Fura 2 ratio imaging method in single cells of confluent monolayers. At baseline, endothelial [Ca(2+)](i) levels remained steady at 86 nmol/L for >20 minutes. Cross-linking of the alpha(v)beta(3) integrin through the sequential exposure of monolayers to anti-alpha(v)beta(3) monoclonal antibody LM609 and secondary IgG resulted in a [Ca(2+)](i) increase of 100% above baseline. This increase commenced in <0.5 minute, peaked in <2 minutes, and decayed to baseline in approximately 5 minutes. Similar responses occurred after the addition of vitronectin (400 microg/mL). In contrast, external Ca(2+) depletion blunted the cross-linking-induced [Ca(2+)](i) increase by 60%, a response that was completely inhibited when the monolayers were also pretreated with thapsigargin. Thus, the [Ca(2+)](i) increase was attributable in part to the release of Ca(2+) from endosomal stores but mostly to Ca(2+) influx across the plasma membrane. Induced aggregation of the alpha(v)beta(3) integrin enhanced tyrosine phosphorylation of phospholipase C-gamma1 and increased the accumulation of inositol-1, 4,5 trisphosphate. Genistein, a broad-spectrum tyrosine kinase inhibitor, abrogated both of these effects, as well as the alpha(v)beta(3)-induced [Ca(2+)](i) increases. We conclude that aggregation of the endothelial alpha(v)beta(3) integrin induces a rapid tyrosine phosphorylation-dependent increase in [Ca(2+)](i). This response may subserve the inflammatory role of alpha(v)beta(3) integrin in blood vessels. PMID- 10700452 TI - Coronary microvascular endothelial cell redox state in left ventricular hypertrophy : the role of angiotensin II. AB - Left ventricular hypertrophy (LVH) is associated with elevated plasma angiotensin II (Ang II) levels and endothelial dysfunction. The relationship between Ang II and endothelial dysfunction remains unknown, however, but it may involve an alteration in endothelial cell redox state. We therefore investigated the effect of Ang II on NADH/NADPH oxidase-mediated superoxide anion (O(2)(-)) production by cultured guinea pig coronary microvascular endothelial cells (CMVEs) and CMVEs freshly isolated from a guinea pig, pressure-overload model of LVH. Lucigenin chemiluminescence was used to measure O(2)(-) production in the particulate fraction of CMVE lysates. In cultured cells, incubation with Ang II (0.1 nmol/L to 1 micromol/L for 18 hours) resulted in significant (P<0.01) increases in both NADH- and NADPH-dependent O(2)(-) production, with a peak effect at 1 nmol/L. The latter was significantly (P<0.01) inhibited by the AT(1) receptor antagonist losartan (1 micromol/L for 18 hours). In contrast, the O(2)(-) response to Ang II (0.1 nmol/L to 1 micromol/L for 18 hours) was largely unaffected by concomitant exposure to the AT(2) antagonist PD 123319 (1 micromol/L). In freshly isolated CMVEs from nonoperated animals, NADH- and NADPH-dependent O(2)(-) production was not different from that in sham-operated animals but was significantly (P<0.05) elevated in the aortic-banded animals. Plasma Ang II levels were significantly (P<0.001) elevated in the aortic-banded (1.25+/-0.12 microg/L, n=12) compared with sham-operated animals (0.63+/-0.06 microg/L, n=12). These data suggest that the endothelial dysfunction associated with LVH may be due, at least in part, to the Ang II-induced upregulation of NADH/NADPH oxidase-dependent O(2)(-) production. PMID- 10700453 TI - Chimera analysis of troponin I domains that influence Ca(2+)-activated myofilament tension in adult cardiac myocytes. AB - The goal of this study was to investigate isoform-specific functional domains of the inhibitory troponin subunit, troponin I (TnI), as it functions within the intact myofilaments of adult cardiac myocytes. Adenovirus-mediated gene transfer was used to deliver and express a TnI chimera composed of the amino terminus of cardiac TnI (cTnI) and the carboxy terminus of slow skeletal TnI (ssTnI) in adult rat cardiac myocytes. The TnI chimera, designated N-card/slow-C TnI, was expressed and incorporated into myofilaments after gene transfer, without detectable changes in contractile protein stoichiometry or sarcomere architecture. Interestingly, force at submaximal Ca(2+) levels was markedly elevated in single permeabilized myocytes expressing the N-card/slow-C TnI chimera relative to force generated in adult myocytes expressing ssTnI or cTnI. Based on these results, a hierarchy of myofilament Ca(2+) sensitivity is emerging by use of TnI chimera analysis, with the order of sensitivity being N-card/slow-C TnI>>ssTnI>>cTnI. These results also strongly suggest that independent isoform specific domains in both the amino and carboxy portions of TnI influence myofilament Ca(2+) sensitivity. In additional studies carried out under pathophysiological ionic conditions (pH 6.2), the dramatic acidosis-induced decrease in myofilament Ca(2+) sensitivity observed in myocytes expressing cTnI was blunted in myocytes expressing N-card/slow-C TnI in a manner similar to that in ssTnI-expressing myocytes. These results demonstrate that there is a pH sensitive domain residing in the carboxy-terminal portion of TnI. The dissection of isoform-specific functional domains under physiological and acidic pH conditions demonstrates the utility of TnI chimeras for analysis of TnI function and provides important insights into the overall function of TnI within the intact myofilament of adult cardiac myocytes. PMID- 10700455 TI - UltraRapid communications : A novel anionic inward rectifier in native cardiac myocytes PMID- 10700454 TI - Identification of cis elements in the cardiac troponin T gene conferring specific expression in cardiac muscle of transgenic mice. AB - To investigate the underlying mechanism regulating cardiac gene expression, transgenic mice carrying the rat cardiac troponin T proximal promoter (-497 bp from the transcriptional start site) fused to a LacZ or chloramphenicol acetyltransferase (CAT) reporter gene were analyzed. The LacZ expression pattern throughout development was very similar to that of the endogenous cardiac troponin T gene. Within this promoter, a high degree of sequence homology was found at 2 sites, modules D (-335 to -289 bp) and F (-249 to -209 bp). Both regions contain at least a TCTG(G/C) direct repeat and an A/T-rich site, whereas only the F module has a muscle enhancer factor 2 (MEF2)-like motif. No significant decrease in CAT transgene expression was observed when only the MEF2 core sequence was mutated. However, when the MEF2 core sequence and its flanking TCTGG site were mutated (Mut5), CAT transgene expression was significantly decreased in the heart, and ectopic expression of the transgene was also observed. When mutations were introduced into this promoter to destroy all upstream TCTG(G/C) direct repeats in the D module (MutD), CAT expression remained cardiac specific, but the expression level was dramatically decreased. Relaxation of cardiac-specific transgene expression became even more severe in transgenic mice carrying double mutations (Mut[D+5]). In addition, CAT activity in the heart was nearly abolished. These results suggest that D and F modules have an additive function in determining the level of expression in the heart and only the F module confers cardiac-specific expression. PMID- 10700456 TI - A novel anionic inward rectifier in native cardiac myocytes. AB - Although the cationic inward rectifiers (Kir and hyperpolarization-activated I(f) channels) have been well characterized in cardiac myocytes, the expression and physiological role of anionic inward rectifiers in heart are unknown. In the present study, we report the functional and molecular identification of a novel chloride (Cl(-)) inward rectifier (Cl.ir) in mammalian heart. Under conditions in which cationic inward rectifier channels were blocked, membrane hyperpolarization (-40 to -140 mV) activated an inwardly rectifying whole-cell current in mouse atrial and ventricular myocytes. Under isotonic conditions, the current activated slowly with a biexponential time course (time constants averaging 179.7+/-23.4 [mean+/-SEM] and 2073.6+/-287.6 ms at -120 mV). Hypotonic cell swelling accelerated the activation and increased the current amplitude whereas hypertonic cell shrinkage inhibited the current. The inwardly rectifying current was carried by Cl(-) (I(Cl.ir)) and had an anion permeability sequence of Cl(-)>I( )>>aspartate. I(Cl.ir) was blocked by 9-anthracene-carboxylic acid and cadmium but not by stilbene disulfonates and tamoxifen. A similar I(Cl.ir) was also observed in guinea pig cardiac myocytes. The properties of I(Cl.ir) are consistent with currents generated by expression of ClC-2 Cl(-) channels. Reverse transcription polymerase chain reaction and Northern blot analysis confirmed transcriptional expression of ClC-2 in both atrial and ventricular tissues and isolated myocytes of mouse and guinea pig hearts. These results indicate that a novel I(Cl.ir) is present in mammalian heart and support a potentially important role of ClC-2 channels in the regulation of cardiac electrical activity and cell volume under physiological and pathological conditions. PMID- 10700457 TI - Asynchronous Ca(2+) waves in intact venous smooth muscle. AB - The rabbit inferior vena cava (IVC) is a large-capacitance vessel that displays typical contractile dose-response curves for caffeine and phenylephrine (PE). Using confocal microscopy on the endothelium-denuded IVC, we undertook experiments to correlate these whole-tissue contractile dose-response curves with changes in subcellular [Ca(2+)](i) signals in the in situ vascular smooth muscle cells (VSMCs). We observed that both caffeine and PE initially elicited Ca(2+) waves in individual VSMCs. The [Ca(2+)](i) in cells challenged with caffeine subsequently returned to baseline whereas the [Ca(2+)](i) in cells challenged with PE exhibited repetitive asynchronous Ca(2+) waves. These [Ca(2+)](i) oscillations were related to Ca(2+) release from the sarcoplasmic reticulum as they were inhibited by ryanodine and caffeine. The lack of synchronicity of the [Ca(2+)](i) oscillations between VSMCs can explain the observed tonic contraction at the whole-tissue level. The nature of these Ca(2+) waves was further characterized. For caffeine, the amplitude was all-or-none in nature, with individual cells differing in sensitivity, leading to their recruitment at different concentrations of the agonist. This concentration dependency of recruitment appears to form the basis for the concentration dependency of caffeine-induced contraction. Furthermore, the speed of the Ca(2+) waves correlated positively with the concentration of caffeine. In the case of PE, we observed the same characteristics with respect to wave speed, amplitude, and recruitment. Increasing concentrations of PE also enhance the frequency of the [Ca(2+)](i) oscillations. We therefore conclude that PE stimulates whole-tissue contractility through differential recruitment of VSMCs and enhancement of the frequency of asynchronous [Ca(2+)](i) oscillations once the cells are recruited. PMID- 10700458 TI - A novel PCR-based technique using expressed sequence tags and gene homology for murine genetic mapping: localization of the complement genes. AB - The complement system is a cascade of serum proteins and receptors which forms a vital arm of innate immunity and enhances the adaptive immune response. This work establishes the chromosomal localization of four key genes of the murine complement system. Mapping was performed using a novel and rapid PCR restriction length polymorphism method which was developed to exploit the murine expressed sequence tag (EST) database. This technique circumvents the laborious cDNA or genomic cloning steps of other mapping methods by relying on EST data and the prediction of exon-intron boundaries. This method can be easily applied to the genes of other systems, ranging from the interests of the individual researcher to large-scale gene localization projects. Here the complement system, probably one of the most well-characterized areas of immunology, was used as a model system. It was shown that the C3a receptor C1r and C1s genes form an unexpected complement gene cluster towards the telomeric end of chromosome 6. The second mannose binding lectin-associated serine protease gene was mapped to the telomeric end of chromosome 4, which is distinct from other complement-activating serine proteases. These results provide new insights into the evolution of this group of proteins. PMID- 10700459 TI - Roles of alpha(4) integrins/VCAM-1 and LFA-1/ICAM-1 in the binding and transendothelial migration of T lymphocytes and T lymphoblasts across high endothelial venules. AB - Several cell adhesion molecules that mediate the binding of lymphocytes to high endothelial venules (HEV) from flowing blood have been identified but the regulation of lymphocyte migration across the HEV wall into the lymph node (LN) is far from understood. In this study we have used an in vitro model of lymphocyte migration across HEV, and analysed the roles of two integrins in the binding and transendothelial migration of T lymphocytes and T lymphoblasts. The adhesion of T lymphocytes to high endothelial cells (HEC) cultured from rat LN HEV differed from that of T lymphoblasts since the percentage of T lymphoblasts that adhered and transmigrated was higher and was not increased by IFN-gamma pretreatment of HEC. Antibodies to alpha(4) integrins, VCAM-1 or LFA-1 maximally inhibited T lymphocyte adhesion by 40-50%, whereas antibodies to ICAM-1 were less effective (<20% inhibition). The effects of alpha(4) integrin and LFA-1 antibodies were additive, giving >90% inhibition. T lymphocytes which adhered in the presence of LFA-1 antibody showed reduced levels of transmigration and, in the presence of alpha(4) integrin antibody, slightly increased transmigration. Antibodies to alpha(4) integrins, VCAM-1, LFA-1 or ICAM-1 had little effect on T lymphoblast adhesion (maxima of 10-30% inhibition) and T lymphoblasts transmigrated normally in the presence of either alpha(4) integrin or LFA-1 antibodies. However, the effects of alpha(4) integrin and LFA-1 antibodies on T lymphoblast adhesion were synergistic, giving >90% inhibition of adhesion. These results suggest that the majority of T lymphoblasts use either alpha(4) integrins or LFA-1 to bind and transmigrate HEV, and the roles of these integrins on activated T cells are overlapping and redundant. In contrast, either integrin supports half-maximal binding of unactivated T lymphocytes to the surface of HEV and LFA-1 makes a larger contribution than alpha(4) integrins to transendothelial migration. PMID- 10700460 TI - Selective induction of p38 mitogen-activated protein kinase activity following A6H co-stimulation in primary human CD4(+) T cells. AB - We have recently described the novel A6H antigen expressed on human peripheral blood T cells and on renal cell carcinoma cells. Cross-linking of the A6H antigen results in co-stimulation of human CD4(+) T cells, characterized by induction of the transcription factor activator protein-1 (AP-1), proliferation and prominent IFN-gamma production, but low levels of IL-2. The proximal signaling events associated with A6H ligation include protein tyrosine kinase phosphorylation and association of p56 Lck, ZAP-70 and the TCR zeta chain. In this study we show that A6H co-stimulation selectively induced activation of the p38 mitogen-activated protein kinase (MAPK) pathway, whereas no significant c-Jun N-terminal kinases (JNK) activity was observed. In contrast, CD28 co-stimulation resulted in both p38 and JNK MAPK activities. Human CD4(+) T cells co-stimulated with A6H up regulated AP-1 binding proteins reactive with a proximal AP-1 binding site in the human IFN-gamma promoter and a consensus AP-1 binding site. Moreover, preincubation of the T cells with the specific p38 MAPK inhibitor SB203580 resulted in decreased AP-1 binding following A6H or CD28 co-stimulation. This suggests that the p38 MAPK pathway is required for induction of full AP-1 binding activity in human CD4(+) T cells co-stimulated with A6H or CD28. Blocking the p38 MAPK pathway by SB203580 completely inhibited IFN-gamma production from A6H co stimulated T cells and radically reduced IFN-gamma production from T cells co stimulated with anti-CD28. In contrast, no significant inhibition of IL-2 production was seen after blocking of the p38 MAPK in either A6H or CD28 co stimulated T cells. Since the p38 MAPK recently has been shown to be critically involved in regulation of IFN-gamma production from T(h)1 cells, we propose that A6H co-stimulation induces a specific pathway, mediated via p38 and AP-1 activation, for induction of a T(h)1 profile in human CD4(+) T cells. PMID- 10700461 TI - TCR v(beta) repertoire restriction and lack of CDR3 conservation implicate TCR superantigen interactions in promoting the clonal evolution of murine thymic lymphomas. AB - Thymic lymphoma development is a multistage process in which genetic and epigenetic events cooperate in the emergence of a malignant clone. The notion that signaling via TCR-ligand interactions plays a role in promoting the expansion of developing neoplastic clones is a matter of debate. To investigate this issue, we determined the TCR V(beta) repertoire of thymic lymphomas induced in AKR/J mice by either endogenous retroviruses or the carcinogen, N-methyl-N nitrosourea (MNU). Both spontaneous and MNU-induced lymphomas displayed restricted V(beta) repertoires. However, whereas V(beta)6, V(beta)8 and V(beta)9 were expressed by a greater than expected frequency of MNU-induced lymphomas, V(beta)8, V(beta)7, V(beta)13 and V(beta)14 were over-represented on spontaneous lymphomas. The dissimilar TCR V(beta) profiles indicate that different endogenous ligands promote neoplastic clonal expansion in untreated and MNU-treated mice. Although the nature of these ligands is not clear, the lack of conservation in TCR beta chain CDR3 regions among lymphomas that express the same V(beta) segment suggests that endogenous superantigens (SAG), as opposed to conventional peptide ligands, are likely to be involved in the selection process. The biased representation of lymphomas expressing V(beta)6-, V(beta)7- and V(beta)9 containing TCRs that recognize endogenous SAG is consistent with this hypothesis. The finding that Bcl-2 is expressed at high levels in spontaneous and MNU-induced lymphomas suggests that preneoplastic thymocytes may be resistant to SAG-induced clonal deletion. A working model is presented in which preneoplastic clones expressing TCRs that recognize endogenous SAG are selectively expanded as a consequence of sustained TCR-mediated signaling. PMID- 10700462 TI - V(beta)8(+) T cells protect from demyelinating disease in a viral model of multiple sclerosis. AB - Previous studies illustrated the influence of T cell subsets on susceptibility or resistance to demyelination in the Theiler's murine encephalomyelitis virus (TMEV) model of multiple sclerosis. Genetic segregation analysis showed a correlation with disease phenotype in this model with particular V(beta) genes. In this study we investigated the contribution of specific V(beta) TCR to the pathogenesis of virus-induced demyelinating disease. Spectratype analysis of cells infiltrating the CNS early in infection demonstrated an over-representation of V(beta)8(+) T cells in mice expressing a susceptible H-2 haplotype. We infected transgenic mice expressing the V(beta)8.2 TCR directed against a non TMEV antigen and found an increase in demyelinating disease in mice of either susceptible or resistant background compared with littermate controls. In addition, depletion studies with an anti-V(beta)8-specific antibody in both susceptible (B10.Q) and resistant (C57BL/6) mice resulted in increased demyelination. TCR analysis of VP2-specific cytotoxic T cell clones from mice with a resistant genotype identified only the V(beta)8.1 TCR, suggesting that limited T cell diversity is critical to TMEV clearance. Together, these results support a protective role for V(beta)8(+) T cells in virus-induced demyelinating disease. PMID- 10700463 TI - A repetitive sequence of Epstein-Barr virus nuclear antigen 6 comprises overlapping T cell epitopes which induce HLA-DR-restricted CD4(+) T lymphocytes. AB - Most human adults carry the Epstein-Barr virus (EBV) and develop immunological memory against the structural and the virus-encoded cellular proteins. The EBV nuclear antigen 6 (EBNA6) elicits cytotoxic T cell responses and it also maintains a persistent antibody response. The majority of sera from EBV seropositive individuals reacts with a synthetic peptide, p63, comprising 21 amino acids of a repetitive region of EBNA6. CD4(+) T lymphocytes, with specificity for p63, could be recalled from the T cell repertoire of EBV carriers that expressed certain HLA-DR allotypes which were identified as good binders of p63 by an in vitro flow cytometric assay. Analysis of the HLA-DR/p63 interaction by molecular mechanics calculations indicated the presence of multiple overlapping epitopes which were predicted to bind in a HLA-DRB1 allo- and subtype specific manner. Specific activation of p63-selected long-term CD4(+) T cell cultures resulted in a proliferative response, in the production of IL-2 and in the secretion of high levels of tumor necrosis factor as measured by bioassays. Proliferation and cytokine production of p63-specific T cells could be induced by p63-loaded HLA-DR-matched antigen-presenting cells and by B cells co-expressing relevant HLA-DR molecules and EBNA6. Our results show that peptides of an EBNA6 repeat region induce CD4(+) T cells which can react with EBNA6-carrying cells in many individuals. We suggest that these T(h) cells may be important in conditioning dendritic cells for initiation potent virus-specific immune responses, provide help for EBV-specific B cells, drive IgG isotype switch and support the sustained effector function of memory cytotoxic T lymphocytes. PMID- 10700464 TI - Synthetic oligodeoxynucleotide containing CpG motif induces an anti polysaccharide type 1-like immune response after immunization of mice with Haemophilus influenzae type b conjugate vaccine. AB - Synthetic oligodeoxynucleotides containing CpG motifs [immunostimulatory sequences (ISS)] have been described as potent adjuvants of type 1 immune responses when co-administered with protein or peptide vaccines. To investigate their role in the immune response to polysaccharides (CHO), different preparations of anti-Haemophilus influenzae type b (Hib) conjugate vaccine were administered to mice. The unconjugated CHO did not induce the synthesis of specific antibodies even in the presence of ISS. On the other hand, anti-CHO specific antibodies significantly increased in the presence of ISS, when tetanus (TT) or diphtheria [cross-reacting material (CRM)] toxoid-conjugated CHO were used to immunize mice. The adjuvant effect was also observed for the immune response against the carrier protein (TT and CRM). ISS insured an early and long lasting specific IgG production. The effects of ISS on the anti-CHO immune response could be attributed to the amplification of the T help provided by the carrier. The analysis of anti-CHO IgG subclasses showed a significant increase of IgG2a and IgG3 in the presence of ISS. ISS caused a rapid release of IL-12 and IFN-gamma in sera from treated mice. This data provide a first evidence for the ability of ISS to induce an anti-CHO type 1-like immune response and demonstrate that ISS have the potential to increase host antibody response against both the CHO and the protein component of a conjugated vaccine. PMID- 10700465 TI - Mapping the B cell superantigen binding site for HIV-1 gp120 on a V(H)3 Ig. AB - The emerging class of B cell superantigens includes HIV-1 gp120, which binds to many members of the V(H)3 Ig gene family. The present study addresses the structural features of V(H)3 antibodies conferring gp120 binding activity using a panel of recombinant full-length and Fab Ig proteins. Binding activity was fully conferred by the Fab portion of the Ig molecule. The V(H) region was the major determinant of binding; diverse light chains were permissive for gp120 binding. A series of recombinant V(H)3-V(H)1 chimeric molecules was created to analyze the contribution of different subregions of V(H)3 to gp120 binding. Hypervariable loop 1 (H1) substitution alone caused a 10-fold reduction in binding activity. The framework subregions (FR1, FR2 and FR3) and H2 also influenced binding, since substitutions of various combinations of these subregions conferred 10- to 100 fold binding reductions. We conclude that gp120 binding occurs through a non conventional interaction involving multiple discontinuously arrayed residues spanning the V(H), and including roles in gp120 contact and favorable conformation of the V(H). PMID- 10700466 TI - Identification of CD19(-)B220(+)c-Kit(+)Flt3/Flk-2(+)cells as early B lymphoid precursors before pre-B-I cells in juvenile mouse bone marrow. AB - The combined analysis of the expression of receptor tyrosine kinases c-Kit and Flt3/Flk-2 and of the human CD25 gene expressed as a transgene under the regulation of the mouse lambda5 promoter in the bone marrow of 1-week-old mice allows us to identify three stages of B lymphocyte development before the CD19(+)c-Kit(+) pre-B-I cells. Single-cell PCR analysis of the rearrangement status of the Ig heavy chain alleles allows us to order these early stages of B cell development as follows: (i) B220(+)CD19(-)c-Kit(lo)Flt3/Flk-2(hi)lambda5(-), (ii) B220(+)CD19(-)c-Kit(lo)Flt3/Flk-2(hi)lambda5(+) and (iii) B220(+)CD19(+)c Kit(lo)Flt3/Flk-2(lo)lambda5(+) before B220(+)CD19(+)c-Kit(lo)Flt3/Flk-2( )lambda5(+) pre-B-I cells. All these progenitors are clonable on stromal cells in the presence of IL-7 and can differentiate to CD19(+)c-Kit(-) B-lineage cells. A combination of stem cell factor, Flt3 ligand and IL-7 was also able to support the proliferation and differentiation of the progenitors in a suspension culture. Furthermore, the analyses indicate that the onset of D(H)J(H) rearrangements precedes the expression of the lambda5 gene. These progenitor populations were characteristic of juvenile mice and could not be detected in the bone marrow of adult mice. Hence the expression pattern, and probably the function, of the receptor tyrosine kinases in early B cell differentiation appears to be different in juvenile and adult mice. PMID- 10700467 TI - Inducible differentiation and apoptosis of the pre-B cell receptor-positive pre-B cell line. AB - The function of the pre-B cell receptor (pre-BCR) during B cell differentiation is not precisely defined. To investigate the pre-BCR receptor activity, we have established pre-BCR-positive pre-B cell lines that are able to differentiate into immature B cells in vitro. Antibody cross-linking of the pre-BCR induced apoptosis and differentiation accompanied with tyrosine phosphorylation. A specific tyrosine-phosphorylated 43 kDa protein (p43) was found down-stream of the pre-BCR. The results demonstrated the receptor function of pre-BCR, which indicates that a ligand-like molecule or a cross-linking structure on the cell surface is possibly present. PMID- 10700468 TI - Functional heterogeneity among bone marrow-derived dendritic cells conditioned by T(h)1- and T(h)2-biasing cytokines for the generation of allogeneic cytotoxic T lymphocytes. AB - Three distinct bone marrow (BM)-derived dendritic cells (BMDC) were expanded from BALB/c BM cells by culture with (i) granulocyte macrophage colony stimulating factor (GM-CSF) plus IL-3, (ii) GM-CSF, IL-3 plus T(h)1-biasing cytokines (IL-12 and IFN-gamma) or (iii) GM-CSF, IL-3 plus T(h)2-biasing cytokines (IL-4). All of these cells expressed the DC-specific marker CD11c, and were designated as BMDC0, BMDC1 and BMDC2 cells respectively. BMDC1 cells exhibited superior T cell stimulating activity in allogeneic mixed lymphocyte culture (MLC), while BMDC2 showed inferior stimulating activity. Specifically, BMDC1, as compared with BMDC2, induced a higher frequency of IFN-gamma-producing CD8(+) T cells in MLC. Moreover, BMDC1, but not BMDC2, were strong inducers of H-2(d)-specific cytotoxic T lymphocytes (CTL) in MLC. BMDC0 always showed intermediate stimulatory activity; however, when BMDC0 were cultured with IFN-gamma, they differentiated into BMDC1-like stimulator cells concomitant with the up-regulation of both MHC antigens and co-stimulatory molecules. In contrast, BMDC2 were refractory to differentiation into superior stimulator cells by treatment with IFN-gamma, although this treatment enhanced MHC expression. These findings indicate that T(h)1- and T(h)2-biasing cytokines, in addition to their effect on T(h) cell differentiation, may play a critical role in the functional skewing of DC. These findings have important implications for the development of DC-based immunotherapies. PMID- 10700469 TI - Tolerance and autoimmunity to a gastritogenic peptide in TCR transgenic mice. AB - The catalytic alpha and glycoprotein beta subunits of the gastric H/K ATPase are major molecular targets in human and mouse autoimmune gastritis. We have previously shown that the H/K ATPase beta subunit is required for the initiation of mouse gastritis and identified a gastritogenic H/K ATPase beta subunit peptide (H/Kbeta253-277). Here we report the generation of MHC class II-restricted TCR transgenic mice using V(alpha)9 and V(beta)8.3 TCR chains with specificity for the gastritogenic H/Kbeta253-277 peptide. We found an 8-fold reduction in CD4(+) T cells in the thymus of the transgenic mice. Despite the reduction in intrathymic CD4(+) T cells, V(beta)8. 3-expressing T cells comprised the majority (>90%) of peripheral spleen and lymph node T cells. These peripheral T cells retained their capacity to proliferate in vitro to the H/Kbeta253-277 peptide. Using the responsive T cells, we have restricted the gastritogenic T cell epitope to H/Kbeta261-274. Despite the capacity of the peripheral T cells to proliferate in vitro to the peptide, the majority ( approximately 80%, 13 of 16) of transgenic mice remained free of gastritis while a minority (20%, three of 16) spontaneously developed an invasive and destructive gastritis. Our results confirm that H/Kbeta261-274 is a gastritogenic peptide. The data also suggest that CD4 T cell tolerance to the gastritogenic peptide in the transgenic mice is maintained by a combination of intrathymic and peripheral tolerance mechanisms. PMID- 10700470 TI - The origin of anti-nuclear antibodies in bcl-2 transgenic mice. AB - bcl-2 transgenic mice develop anti-double-stranded (ds) DNA antibodies similar to those present in systemic lupus erythematosus. To begin to understand where a breakdown in the regulation of autoreactive lymphocytes is occurring, we have used a bcl-2 transgene (Tg) in conjunction with an Ig Tg that allows us to identify and track anti-dsDNA B cells. Previously, we have shown that anti-dsDNA B cells are actively tolerized in BALB/c mice as manifested by their developmental arrest, follicular exclusion, increased in vivo turnover rate and lack of their antibody in the serum. The bcl-2 Tg mice increased the lifespan of anti-dsDNA B cells, but did not alter the other features of tolerance, indicating that the anergy of the anti-dsDNA B cells is independent of their reduced lifespan. Furthermore, these data suggest that the serum anti-dsDNA antibodies in bcl-2 transgenic mice are not due to a breakdown in the induction or maintenance of B cell anergy; rather they may originate from B cells that have transited through a germinal center. PMID- 10700471 TI - Comparison of Fas- versus perforin-mediated pathways of cytotoxicity in TCR- and Thy-1-activated murine T cells. AB - T cell-mediated cytotoxicity can be triggered by cross-linking of TCR or Thy-1 surface proteins. While the TCR-triggered signaling initiates both perforin- and Fas ligand (FasL)-Fas-mediated mechanisms of cytotoxicity, it was not clear which mechanism was utilized by Thy-1-triggered signals and which pathway of cytotoxicity was triggered at low levels of antigen expression. It is shown that glycophosphatidylinositol-linked surface glycoprotein Thy-1 preferentially activates FasL-Fas- but not perforin-mediated cytotoxicity. This is explained by the lesser intensity of Thy-1-mediated signaling in T cells. The data suggest that Thy-1-triggered Fas-mediated cytotoxicity is completely dependent on cross talk between Thy-1 and TCR signals since mutations in TCR-CD3 complex molecules or inhibition of tyrosine kinases or calcineurin abolished or strongly inhibited Thy-1-triggered FasL-Fas-mediated cytotoxicity. Lower concentrations of antigenic peptide or levels of cross-linking with anti-TCR-CD3 mAb are required to trigger Fas-mediated than perforin-mediated cytotoxicity by different cytotoxic T lymphocyte (CTL) lines and clones, and it is shown that cross-linking of Thy-1 is much less efficient in triggering accumulation of second messengers (intracellular Ca(2+)) than cross-linking of TCR on CTL. Taken together, these data reflect the possibility of differential activation of FasL and/or perforin pathways of cytotoxicity depending on the nature of activating stimuli and surface receptor. PMID- 10700472 TI - Minimal peptide length requirements for CD4(+) T cell clones--implications for molecular mimicry and T cell survival. AB - CD4(+) T lymphocytes usually recognize peptides of 12-16 amino acids in the context of HLA class II molecules. We have recently used synthetic peptide combinatorial libraries to dissect in detail antigen recognition by autoreactive CD4(+) T cell clones (TCC). The results of these studies demonstrated that antigen recognition by T cells is highly degenerate and that many cross-reactive ligands can be defined, some of which much more potent than the selecting autoantigen. Based on these observations, we examined the response of a myelin basic protein-specific HLA class II-restricted CD4(+) TCC to truncation variants of optimal ligands. Surprisingly, pentapeptides, tetrapeptides and even tripeptides derived from different segments of the optimal ligands were recognized by the TCC, and some were even more potent than the selecting autoantigen. In addition, these peptides enhanced the survival of the TCC at low concentration. The relevance of this finding was supported by the generation of pentapeptide-specific CD4(+) TCC from peripheral blood lymphocytes. These observations not only change existing views on the length requirements for activation of CD4(+) HLA class II-restricted T cells, but also extend our knowledge about the flexibility of TCR recognition and the potential for cross reactivity in the immune system. PMID- 10700473 TI - Affiliation to mature B cell repertoire and positive selection can be separated in two distinct processes. AB - Using an 'oligoclonal' model, we have previously shown that mice transgenic for a mu chain (H3) and deficient for kappa chain expression display a mature B cell repertoire largely dominated by the H3/lambda1 pair, while the four H3/lambda available combinations can be observed in the immature B cell compartment. This led us to propose the existence of a positive selection process. To test this hypothesis, we have introduced the SJL lambda locus coding for a defective lambda1 chain (lambda1(s)) that creates a dysfunctional Ig receptor complex during B cell differentiation. Our results show that the lambda1(s) defect impairs the development of mature B cells when the H3-mu transgene insert is present in the hemizygous state. This suggests that the Gly --> Val substitution present in the C(lambda)1(s) chain at position 155 is sufficient to abrogate the selection of the H3/lambda1 pair. Unexpectedly, when the H3-mu transgene array is present in a homozygous state in lambda1(s) mice but not in 'wild-type' lambda1 mice (lambda1(+)), a significant number of mature B cells expressing all H3/lambda combinations can be developed. These results indicate that the overriding H3/lambda1 dominance observed in lambda1(+) mice is due to a positive selection process and not to a negative selection of other H3/lambda combinations. They also show that the export of B cells to the periphery can be controlled by the expression of the mu chain. PMID- 10700474 TI - B cell development and activation defects resulting in xid-like immunodeficiency in BLNK/SLP-65-deficient mice. AB - Engagement of the B cell receptor (BCR) leads to the activation of tyrosine kinases and other signaling molecules that ultimately determine the type and magnitude of the B lymphocyte's cellular response. The adaptor protein BLNK/SLP 65 plays a pivotal role in BCR signal transduction by coupling Syk activation to downstream elements such as Grb2, phospholipase C-gamma, Vav and Nck. We have generated BLNK(-/-) mice to determine the physiological role of this protein in B cell development and activation. BLNK(-/-) mice exhibit an incomplete block in B cell development with a severe inhibition of pro-B to pre-B cell differentiation. BLNK(-/-) sIgM(+) cells can develop, seed the peripheral lymphoid tissues and accumulate in numbers overtime. However, these mutant B cells failed to mature and are non-responsive to BCR cross-linking in terms of proliferation and up regulation of activation markers such as CD69 and CD86 (B7-2). In addition, the CD5(+) subset of B cells is absent. The immune response to T cell-independent antigen but not T cell-dependent antigen is also impaired. Overall, the phenotype of BLNK(-/-) mice bears a striking resemblance to that of xid mice which is the murine model of human XLA that has a mutation in Bruton's tyrosine kinase. This raises the interesting possibility that mutation in BLNK/SLP-65 may be responsible for certain human immunodeficiencies. PMID- 10700475 TI - Xenotransplantation: postponed by a millennium? PMID- 10700476 TI - A practical guide to continuous population-based data collection (PACE): a process facilitating uniformity of care and research into practice. PMID- 10700477 TI - Lipoprotein(a): from ancestral benefit to modern pathogen? AB - We review current concepts regarding the genetic, structural and metabolic features of lipoprotein(a), a major inherited cardiovascular pathogen. Although lipoprotein(a) is almost completely confined to a subset of primates, the hedgehog produces a lipoprotein(a)-like complex, which appears to have evolved independently from that of humans. The physiological role of lipoprotein(a) in humans is still unclear, and individuals with low or null concentrations of plasma lipoprotein(a) manifest no deficiency syndrome or disease. The integration of recent discoveries about the structure and metabolism of this unique lipoprotein particle has allowed the formulation of some hypotheses concerning the evolutionary advantages of synthesizing lipoprotein(a)-like particles. PMID- 10700478 TI - Diabetes and the Mediterranean diet: a beneficial effect of oleic acid on insulin sensitivity, adipocyte glucose transport and endothelium-dependent vasoreactivity. AB - Abnormalities in endothelial function may be associated with increased cardiovascular risk in diabetic patients. We examined the effect of an oleic-acid rich diet on insulin resistance and endothelium-dependent vasoreactivity in type 2 diabetes. Eleven type 2 diabetic patients were changed from their usual linoleic-acid-rich diet and treated for 2 months with an oleic-acid-rich diet. Insulin-mediated glucose transport was measured in isolated adipocytes. Fatty acid composition of the adipocyte membranes was determined by gas-liquid chromatography and flow-mediated endothelium-dependent and -independent vasodilatation were measured in the superficial femoral artery at the end of each dietary period. There was a significant increase in oleic acid and a decrease in linoleic acid on the oleic-acid-rich diet (p<0.0001). Diabetic control was not different between the diets, but there was a small but significant decrease in fasting glucose/insulin on the oleic-acid-rich diet. Insulin-stimulated (1 ng/ml) glucose transport was significantly greater on the oleic- acid-rich diet (0.56+/ 0.17 vs. 0.29+/-0.14 nmol/10(5) cells/3 min, p<0.0001). Endothelium-dependent flow-mediated vasodilatation (FMD) was significantly greater on the oleic-acid rich diet (3.90+/-0.97% vs. 6.12+/-1.36% p<0.0001). There was a significant correlation between adipocyte membrane oleic/linoleic acid and insulin-mediated glucose transport (p<0.001) but no relationship between insulin-stimulated glucose transport and change in endothelium-dependent FMD. There was a significant positive correlation between adipocyte membrane oleic/linoleic acid and endothelium-dependent FMD (r=0.61, p<0.001). Change from polyunsaturated to monounsaturated diet in type 2 diabetes reduced insulin resistance and restored endothelium-dependent vasodilatation, suggesting an explanation for the anti atherogenic benefits of a Mediterranean-type diet. PMID- 10700479 TI - Still hungry in hospital: identifying malnutrition in acute hospital admissions. AB - We assessed the prevalence, methods for recognition and clinical management of malnutrition in acute admissions in a large academic inner-city hospital. Of a total of 337 patients, it was possible to measure both height and weight in 219 patients (65% of admissions). As an alternative for bed-bound patients, mid-upper arm circumference was not very reliable in predicting BMI (sensitivity 98%; specificity 65%), and waist circumference even less so. Of these, 13% were malnourished (body mass index BMI <18.5 kg/m(2) or BMI 18.5-20 kg/m(2) with reported weight loss >3 kg in the last 3 months). Six patients (31% of those with BMI <18.5 kg/m(2)) and one with BMI 18.5-20 kg/m(2) were recognized as suffering from malnutrition and referred to the dietitian. Review of case records could not establish if the diagnosis was missed in the remainder, or if a conscious decision was taken not to manage malnutrition actively. Malnutrition in acute hospital admissions goes apparently unrecognized and unmanaged in 70% of cases. Since there are serious consequences, and effective simple treatment is readily available, increased awareness is required, with routine assessment of nutritional status in all patients. PMID- 10700480 TI - Enlarged vestibular aqueduct: a radiological marker of pendred syndrome, and mutation of the PDS gene. AB - Although the textbook view of Pendred syndrome is that of an autosomal recessive condition characterized by deafness and goitre, it is increasingly clear that not all such patients present this classical clinical picture. Malformations of the inner ear, specifically enlargement of the vestibular aqueduct, are common in Pendred syndrome and mutations in the PDS (Pendred Syndrome) gene have been recorded in patients presenting with deafness and vestibular aqueduct dilatation only, without other features of Pendred syndrome. Since this is the most common radiological malformation of the cochlea in deaf patients, we investigated what proportion of such cases were due to mutation of the PDS gene. We assessed 57 patients referred with radiological evidence of vestibular aqueduct enlargement, by history, clinical examination, perchlorate discharge test and molecular analysis of the PDS locus. Forty-one patients (72%) had unequivocal evidence of Pendred syndrome. The finding of a single heterozygous mutation at the PDS gene in a further eight was strongly suggestive of a critical role for pendrin, the protein product of the PDS gene, in the generation of enlarged vestibular aqueducts in at least 86% (49/57 cases) of patients with this radiological malformation. Securing the diagnosis of Pendred syndrome may be difficult, especially in the single case. Goitre is an inconstant finding, and the perchlorate discharge test, although helpful, is of diagnostic value only if abnormal. Enlargement of the vestibular aqueduct should be considered as the most likely presentation of Pendred syndrome and should prompt specific investigation of that diagnostic possibility. Pendred syndrome might henceforth be recharacterized as deafness with enlargement of the vestibular aqueduct, which is sometimes associated with goitre. PMID- 10700481 TI - Use of oral corticosteroids in the United Kingdom. AB - Administration of oral corticosteroids is associated with the development of osteoporosis and an increased risk of fractures. However, the size of the treated sub-population who would benefit from preventive therapy remains uncertain. The objective of this study was to investigate the usage pattern of oral corticosteroids in a large sample representative of the general population in England and Wales. Information was obtained from the General Practice Research Database (GPRD) which contains medical records of general practitioners. Oral corticosteroid users were patients aged 18 years or older who received one or more prescriptions for oral corticosteroids. Over 1.6 million oral corticosteroid prescriptions were issued to the cohort of 244 235 oral corticosteroid users. At any point in time, oral corticosteroids were being used by 0.9% of the total adult GPRD population. The highest use (2.5%) was by people between 70 and 79 years of age. Respiratory disease was the most frequently recorded indication for oral corticosteroid treatment (40%). Patients with arthropathies were most likely to use long-term, continuous treatment, and patients with chronic obstructive pulmonary disease least likely (19.3% and 6.1%, respectively, used oral corticosteroids for more than 2 years). The overall use of bone-active medication (oestrogens, bisphosphonates, vitamin D, and calcitonin) during oral corticosteroid treatment was low (between 4.0% and 5.5%). The current population in the UK at risk of developing corticosteroid-induced fractures might be as large as 350 000. Identification of these patients will be important for implementing preventive strategies in a cost-effective manner. PMID- 10700483 TI - Confidence limits and the limits of confidence. PMID- 10700482 TI - HIV, hepatitis C and risk behaviour in a Canadian medium-security federal penitentiary. Queen's University HIV Prison Study Group. AB - In a voluntary anonymous HIV and hepatitis C serology screen in a Canadian male medium security federal penitentiary, 68% of 520 prisoners volunteered a blood sample and 99% of those giving a blood sample completed a risk behaviour questionnaire which was linked numerically to the blood sample. Compared to previous screenings for HIV (4 years earlier), and hepatitis C (3 years earlier) in the same institution, HIV seroprevalence had risen from 1% to 2% and hepatitis C seroprevalence from 28% to 33%. The overwhelming risk association for hepatitis C was with drug use outside prison, although there was a small group of men who had only ever injected drugs inside prison, over half of whom had been infected with hepatitis C. The proportion of prisoners who had injected drugs in prison rose from 12% in 1995 to 24% in 1998. The proportion of surveyed individuals sharing injection equipment at some time in prison was 19%, and while HIV rates in the prison are currently low, HIV prevalence amongst Canadian street i.v. drug users is rising rapidly, underlining the need for urgent preventative measures in prisons. PMID- 10700485 TI - Antiphospholipid syndrome and renal artery stenosis. PMID- 10700484 TI - Thrombolysis for acute pulmonary embolism in Chinese patients. PMID- 10700486 TI - Racial variation in initial stroke severity. AB - BACKGROUND AND PURPOSE: Blacks experience greater morbidity and mortality from stroke than do whites. The degree to which this is due to the severity of the initial stroke is not known. The objective of this study is to determine whether there is a racial difference in initial stroke severity. METHODS: A secondary analysis of a prospective cohort of 984 veterans (29.7% black) admitted to any of 9 geographically diverse Veterans Administration Hospitals for acute stroke between April 1995 and March 1997 was performed. Initial stroke severity was ascertained by using the modified Canadian Neurological Scale (CNS) applied retrospectively to medical record data. Stroke severity, unadjusted and adjusted for covariates, was compared between black and white patients. RESULTS: Blacks had greater initial stroke severity than did whites (mean CNS score 7.96 versus 8.32, respectively; P=0.039), with a 0.5-point difference on the scale corresponding to a single-level decrement in either speech or strength of half of an extremity. This difference persisted with adjustment for other important predictors of stroke severity (P=0. 035). However, there was no significant racial difference in severity when CNS scores were collapsed into a priori clinically relevant categories. CONCLUSIONS: Compared with whites, blacks show greater severity of stroke at hospital admission. It remains uncertain whether the relatively small but significant difference at presentation fully explains the striking racial differences in morbidity and mortality from stroke. PMID- 10700487 TI - Effects of stress reduction on carotid atherosclerosis in hypertensive African Americans. AB - BACKGROUND AND PURPOSE: African Americans suffer disproportionately higher cardiovascular disease mortality rates than do whites. Psychosocial stress influences the development and progression of atherosclerosis. Carotid intima media thickness (IMT) is a valid surrogate measure for coronary atherosclerosis, is a predictor of coronary outcomes and stroke, and is associated with psychosocial stress factors. Stress reduction with the Transcendental Meditation (TM) program decreases coronary heart disease risk factors and cardiovascular mortality in African Americans. B-mode ultrasound is useful for the noninvasive evaluation of carotid atherosclerosis. METHODS: This randomized controlled clinical trial evaluated the effects of the TM program on carotid IMT in hypertensive African American men and women, aged >20 years, over a 6- to 9-month period. From the initially enrolled 138 volunteers, 60 subjects completed pretest and posttest carotid IMT data. The assigned interventions were either the TM program or a health education group. By use of B-mode ultrasound, mean maximum IMT from 6 carotid segments was used to determine pretest and posttest IMT values. Regression analysis and ANCOVA were performed. RESULTS: Age and pretest IMT were found to be predictors of posttest IMT values and were used as covariates. The TM group showed a significant decrease of -0.098 mm (95% CI -0. 198 to 0.003 mm) compared with an increase of 0.054 mm (95% CI -0.05 to 0.158 mm) in the control group (P=0.038, 2-tailed). CONCLUSIONS: Stress reduction with the TM program is associated with reduced carotid atherosclerosis compared with health education in hypertensive African Americans. Further research with this stress-reduction technique is warranted to confirm these preliminary findings. PMID- 10700488 TI - Sex differences in the relationship of risk factors to subclinical carotid atherosclerosis measured 15 years later : the Tromso study. AB - BACKGROUND AND PURPOSE: Ultrasound measurement of carotid artery intima-media thickness (IMT) is regarded as a valid index of atherosclerosis. Determinants of IMT in cross-sectional studies have been established, but the long-term relationship between cardiovascular risk factors and subclinical atherosclerosis has not been investigated thoroughly. METHODS: We included in the study 3128 middle-aged men and women in Tromso, Norway, who in 1980 attended the baseline examination with measurements of cardiovascular risk factors and who underwent carotid ultrasonography after 15 years of follow-up. RESULTS: Age, blood pressure, total cholesterol, HDL cholesterol, and body mass index were independent long-term predictors of IMT in both men and women. Triglyceride levels were associated with an increase in IMT in women only, while physical activity and smoking were predictors of IMT in men only. However, smoking was associated with increased risk of having atherosclerotic plaque in both men and women. There were no differences in the strength of risk factor effects on IMT in the common carotid artery and the carotid bifurcation. CONCLUSIONS: The present study indicates that established cardiovascular risk factors are independent predictors of subclinical atherosclerosis measured after 15 years of follow-up. However, there may be significant sex differences in the relationship between triglycerides, smoking, and physical activity and the risk of atherosclerosis. PMID- 10700489 TI - Management patterns and costs of acute ischemic stroke : an international study. For the Stroke Economic Analysis Group. AB - BACKGROUND AND PURPOSE: [corrected] With the ever-increasing pressure on healthcare budgets, we witness a growing demand for evidence of the economic implications of care across many therapeutic areas. Stroke is no exception. METHODS: Detailed information on healthcare use was collected in conjunction with two 12-week international trials designed primarily to assess the safety and efficacy of a new potential neuroprotective agent. The information was gathered prospectively by means of a customized resource use instrument that included both acute and long-term inpatient management as well as community care. In this report, the results pertaining to the 1341 acute ischemic stroke patients are described. RESULTS: More than 70% of the mean cost ($13 668) was explained by the initial hospitalization, which averaged 24 days. The total cost and its components varied according to patient age, the presence of comorbidities, and several indicators of disease severity. Pronounced country differences could be observed in the management of this fairly homogeneous patient group. CONCLUSIONS: This study provides a comprehensive picture of the healthcare services used for the treatment and rehabilitation of stroke victims, presented with respect to various patient and disease characteristics. It is expected that researchers evaluating the cost-efficiency of specific stroke treatments will benefit from the detailed information presented in this report. PMID- 10700490 TI - Increased platelet sensitivity to collagen in individuals resistant to low-dose aspirin. AB - BACKGROUND AND PURPOSE: The purpose of this study was to assess individual differences in the pharmacological effects of acetylsalicylic acid (ASA) on bleeding time as measured by in vitro platelet aggregation and to examine the consistency of responses over time. METHODS: We measured template IIR bleeding time and platelet aggregation in 8 healthy male volunteers before and 2 hours after ingestion of 324 mg of ASA. An individual was considered a nonresponder if his post-ASA bleeding time was not 2 SDs above his baseline bleeding time, where SD was estimated from the baseline bleeding times of the 8 volunteers. The same experiment was done after a 30-month interval. RESULTS: Five volunteers were identified as ASA responders, and 3 were identified as nonresponders. Bleeding time before and after ingestion of ASA was 408+/-121 seconds (mean+/-SD) and 720+/-225 seconds, respectively, in ASA responders and 330+/-30 seconds and 330+/ 52 seconds, respectively, in ASA nonresponders. The mean ED(50) for collagen induced platelet aggregation, that is, the mean concentration of collagen that caused a response at 50% of maximum, was 0.91 microg/mL (95% CI, 0.73 to 1. 14) in ASA responders and 0.48 microg/mL (95% CI, 0.38 to 0.60) in nonresponders. When optimum concentrations of collagen, ie, concentrations that yielded 90% maximum aggregation, were used as stimuli, the mean IC(50) for ASA, that is, the mean concentration that yielded 50% inhibition, was 322.5 micromol/L (95% CI, 264.8 to 392.6) in ASA responders and 336.1 micromol/L (95% CI, 261.0 to 432. 8) in nonresponders. The variability in individual responsiveness in the second experiment remained consistent with that in the first experiment. CONCLUSIONS: ASA resistance may be caused by an increased sensitivity of platelets to collagen. A platelet aggregation study specific for collagen dose response may be useful for strict selection of ASA responders for low-dose ASA therapy and for identifying ASA nonresponders for high-dose ASA therapy. PMID- 10700491 TI - Decreased perihematomal edema in thrombolysis-related intracerebral hemorrhage compared with spontaneous intracerebral hemorrhage. AB - BACKGROUND AND PURPOSE: Intracerebral hemorrhage (ICH) is a highly morbid disease process. Perihematomal edema is reported to contribute to clinical deterioration and death. Recent experimental observations indicate that clotting of the intrahematomal blood is the essential prerequisite for hyperacute perihematomal edema formation rather than blood-brain barrier disruption. METHODS: We compared a series of patients with spontaneous ICH (SICH) to a series of patients with thrombolysis-related ICH (TICH). All patients were imaged within 3 hours of clinical onset. We reviewed relevant neuroimaging features, emphasizing and quantifying perihematomal edema. We then analyzed clinical and radiological differences between the 2 ICH types and determined whether these factors were associated with perihematomal edema. RESULTS: TICHs contained visible perihematomal edema less than half as often as SICHs (31% versus 69%, P<0.001) and had both lower absolute edema volumes (0 cc [25th, 75th percentiles: 0, 6] versus 6 cc [0, 13], P<0.0001) and relative edema volumes (0.16 [0.10, 0.33] versus 0.55 [0.40, 0.83], P<0.0001). Compared with SICHs, TICHs were 3 times larger in volume (median [25th, 75th percentiles] volume 69 cc [30, 106] versus 21 cc [8, 45], P<0.0001), 4 times more frequently lobar in location (62% versus 15%, P<0.001), 80 times more frequently contained blood-fluid level(s) (86% versus 1%, P<0.001), and were more frequently multifocal (22% versus 0%, P<0.001). CONCLUSIONS: The striking qualitative and quantitative lack of perihematomal edema observed in the thrombolysis-related ICHs compared with the SICHs provides the first substantial, although indirect, human evidence that intrahematomal blood clotting is a plausible pathogenetic factor in hyperacute perihematomal edema formation. PMID- 10700492 TI - Abciximab in acute ischemic stroke. A randomized, double-blind, placebo controlled, dose-escalation study. AB - BACKGROUND AND PURPOSE: Abciximab is a potent parenterally administered platelet glycoprotein IIb/IIIa antagonist. Because this agent has been shown to improve outcomes in coronary artery disease, there is interest to evaluate whether it could improve cerebral perfusion and outcomes after ischemic stroke. This study was designed to evaluate the safety of abciximab in acute ischemic stroke and to obtain pilot efficacy data. METHODS: We conducted a randomized, double-blind, placebo-controlled, dose-escalation trial. Seventy-four eligible and consenting patients presenting within 24 hours after ischemic stroke onset at 38 study sites were randomly allocated to receive either an escalating dose of abciximab (54 patients) or placebo (20 patients) in a ratio of 3:1. We studied 4 escalating doses of abciximab. Patients underwent a scheduled follow-up head CT scan 24 to 36 hours after the completion of study agent administration to monitor for bleeding complications and were evaluated through 3 months. RESULTS: There were no cases of major intracranial hemorrhage. Asymptomatic parenchymal hemorrhages were detected on post-study agent CT in 4 of 54 abciximab patients (7%) and in 1 of 20 placebo patients (5%). Six additional abciximab patients had asymptomatic hemorrhagic lesions detected by unscheduled brain imaging during their follow-up period. Nine of 11 patients with asymptomatic hemorrhage had a baseline National Institutes of Health Stroke Scale score >14. At 3 months, there was a trend toward a higher rate of minimal residual disability (Barthel Index > or =95 or modified Rankin scale < or =1) among abciximab patients compared with those who received placebo. CONCLUSIONS: Abciximab appears to be safe when administered up to 24 hours after stroke onset, and it might improve functional outcome. PMID- 10700493 TI - High rate of complete recanalization and dramatic clinical recovery during tPA infusion when continuously monitored with 2-MHz transcranial doppler monitoring. AB - BACKGROUND AND PURPOSE: Clot dissolution with tissue plasminogen activator (tPA) can lead to early clinical recovery after stroke. Transcranial Doppler (TCD) with low MHz frequency can determine arterial occlusion and monitor recanalization and may potentiate thrombolysis. METHODS: Stroke patients receiving intravenous tPA were monitored during infusion with portable TCD (Multigon 500M; DWL MultiDop-T) and headframe (Marc series; Spencer Technologies). Residual flow signals were obtained from the clot location identified by TCD. National Institutes of Health Stroke Scale (NIHSS) scores were obtained before and after tPA infusion. RESULTS: Forty patients were studied (mean age 70+/-16 years, baseline NIHSS score 18.6+/ 6.2, tPA bolus at 132+/-54 minutes from symptom onset). TCD monitoring started at 125+/-52 minutes and continued for the duration of tPA infusion. The middle cerebral artery was occluded in 30 patients, the internal carotid artery was occluded in 11 patients, the basilar artery was occluded in 3 patients, and occlusions were multiple in 7 patients; 4 patients had no windows; and 1 patient had a normal TCD. Recanalization on TCD was found at 45+/-20 minutes after tPA bolus: recanalization was complete in 12 (30%) and partial in 16 (40%) patients. Dramatic recovery during tPA infusion (total NIHSS score <3) occurred in 8 (20%) of all patients (baseline NIHSS range 6 to 22; all 8 had complete recanalization). Lack of improvement or worsening was associated with no recanalization, late recanalization, or reocclusion on TCD (C=0.811, P< or =0.01). Improvement by > or =10 NIHSS points or complete recovery was found in 30% of all patients at the end of tPA infusion and in 40% at 24 hours. Improvement by > or =4 NIHSS points was found in 62.5% of patients at 24 hours. CONCLUSIONS: Dramatic recovery during tPA therapy occurred in 20% of all patients when infusion was continuously monitored with TCD. Recovery was associated with recanalization on TCD, whereas no early improvement indicated persistent occlusion or reocclusion. At 24 hours, 40% of all patients improved by > or =10 NIHSS points or recovered completely. Ultrasonic energy transmission by TCD monitoring may expose more clot surface to tPA and facilitate thrombolysis and deserves a controlled trial as a way to potentiate the effect of tPA therapy. PMID- 10700494 TI - Interrelation between plaque surface morphology and degree of stenosis on carotid angiograms and the risk of ischemic stroke in patients with symptomatic carotid stenosis. On behalf of the European Carotid Surgery Trialists' Collaborative Group. AB - BACKGROUND AND PURPOSE: The risk of ischemic stroke distal to an atherothrombotic carotid stenosis increases with the degree of stenosis. The main mechanism of stroke is thought to be embolism from fissured or ruptured plaque, but there are few published data on the relationship between plaque morphology and severity of stenosis and their independent effects on the risk of ischemic stroke. We sought to determine the interrelation between plaque surface morphology, degree of carotid stenosis, and the risk of ipsilateral ischemic stroke. METHODS: Severity of stenosis and plaque surface morphology were assessed on angiograms of the symptomatic carotid artery in 3007 patients in the European Carotid Surgery Trial and were related to baseline clinical characteristics, pathological characteristics of plaques examined at endarterectomy, and the risks of carotid territory ipsilateral ischemic stroke and other vascular events on follow-up. RESULTS: The early risk of ipsilateral ischemic stroke on medical treatment was closely related to the degree of carotid stenosis. However, the initial degree of carotid stenosis was not predictive of strokes occurring >2 years after randomization. Angiographic plaque surface irregularity and plaque surface thrombus at endarterectomy increased in frequency as the degree of stenosis increased (both P<0.0001). However, the degree of stenosis was still predictive of the 2-year risk of stroke on medical treatment after correction for plaque surface irregularity. Angiographic plaque surface irregularity was an independent predictor of ipsilateral ischemic stroke on medical treatment at all degrees of stenosis (hazard ratio=1.80; 95% CI, 1. 14 to 2.83; P=0.01). This relationship was maintained when the analysis was confined to strokes occurring >2 years after randomization (hazard ratio=2.75; 95% CI, 1.30 to 5.80; P=0.01). Neither the degree of stenosis nor plaque surface irregularity was predictive of the "background" stroke risk after endarterectomy or the risk of nonstroke vascular events. CONCLUSIONS: Angiographic plaque surface irregularity is associated with an increased risk of ipsilateral ischemic stroke on medical treatment at all degrees of stenosis. The increase in stroke risk with degree of stenosis is partly accounted for by the parallel increase in plaque surface irregularity and thrombus formation, but the degree of narrowing of the vessel lumen is still an independent predictor of ischemic stroke within 2 years of presentation. PMID- 10700495 TI - Low risk of ischemic stroke in patients with reduced internal carotid artery lumen diameter distal to severe symptomatic carotid stenosis: cerebral protection due to low poststenotic flow? On behalf of the European Carotid Surgery Trialists' Collaborative Group. AB - BACKGROUND AND PURPOSE: Patients with recently symptomatic severe carotid stenosis have a high risk of ischemic stroke on medical treatment. The main mechanism of stroke appears to be plaque surface thrombus formation and distal embolism. It is unclear to what extent reduction in blood flow across the stenosis, and the consequent reduction in cerebral perfusion pressure, is also important. Angiographic indices of reduced cerebral perfusion may identify patients at a particularly high risk of stroke who require urgent endarterectomy. The most direct angiographic correlate of poststenotic perfusion pressure is the degree of narrowing of the distal internal carotid artery (ICA) lumen. We sought to develop criteria for the definition of poststenotic narrowing of the ICA and to determine the effect of this and other angiographic characteristics likely to be associated with reduced cerebral perfusion on the risk of ipsilateral ischemic stroke in patients with recently symptomatic carotid stenosis. METHODS: We studied the carotid angiograms of 3007 patients in the European Carotid Surgery Trial. Poststenotic narrowing of the ICA was defined with use of the ratio of the lumen diameter of the ICA to that of the common carotid artery (CCA). The normal range of the ICA/CCA ratio was defined in 2966 symptomatic or contralateral carotid arteries with 0% to 49% stenosis. Arteries with 70% to 99% symptomatic stenosis and an ICA/CCA ratio below this range were categorized as narrowed. We related the presence of narrowing and other angiographic characteristics to the risk of ipsilateral ischemic stroke on medical treatment. RESULTS: An assessment of the ICA/CCA ratio had good interobserver reproducibility. Poststenotic narrowing of the ICA was defined as an ICA/CCA ratio of <0.42. The 5-year risk of ipsilateral carotid territory ischemic stroke on medical treatment was 8% in patients with 70% to 99% stenosis and narrowing of the ICA versus 25% in patients without narrowing (log rank test, P=0.02). This difference remained after correction for other clinical and angiographic variables (hazard ratio 0.40, 95% CI 0.17 to 0.94, P=0. 03). The other angiographic characteristics did not predict stroke. CONCLUSIONS: Poststenotic narrowing of the ICA was associated with a low risk of stroke on medical treatment. This suggests that low flow alone is not usually sufficient to cause ischemic stroke distal to symptomatic carotid stenosis. Poststenotic narrowing may be protective because blood flow distal to the stenosis is insufficient to carry emboli to the brain. PMID- 10700496 TI - Internal borderzone infarction: a marker for severe stenosis in patients with symptomatic internal carotid artery disease. For the North American Symptomatic Carotid Endarterectomy (NASCET) Group. AB - BACKGROUND AND PURPOSE: Among subcortical infarctions, internal borderzone infarcts (IBI) are considered to be separate entities from perforating artery infarcts (PAI). The purpose of the present study is to examine the relationship between the presence of IBI and the degree of angiographically defined internal carotid artery (ICA) stenosis in symptomatic patients. METHODS: A review of 1253 brain CTs from patients recruited by the North American Symptomatic Carotid Endarterectomy Trial was performed, using templates for the identification of subcortical and cortical vascular territories. RESULTS: A total of 413 patients had visible ischemic lesions on the side ipsilateral to their symptomatic ICA. Of these, 138 had PAI, 108 had IBI, 122 had cortical infarcts, and 45 had a combination of different lesions. Mean (+/-SD) lesion diameter was larger for IBI (11.0+/-5.9 mm) than for PAI (7.1+/-4.7 mm) (P<0.001 for comparing 2 means). IBI was associated with higher degrees of ICA stenosis (P<0. 001). Sixty-three percent of the patients with IBI had severe (70% to 99%) ICA stenosis compared with 42% of patients with PAI; 18% of the IBI patients had stenosis of 90% or more compared with 8% of the patients with PAI. Multiple logistic regression did not identify any patient characteristics as confounders. CONCLUSIONS: Among subcortical infarctions, IBI are associated with higher degrees of ICA stenosis in symptomatic patients. Differentiating between internal borderzone and perforating artery infarcts is important, because each may arise from different mechanisms, namely, carotid disease and small-vessel disease, respectively. PMID- 10700497 TI - Functional and neuroanatomic correlations in poststroke depression: the Sunnybrook Stroke Study. AB - BACKGROUND AND PURPOSE: The purpose of our study was to determine the functional and neuroanatomic correlates of poststroke depressive symptoms. METHODS: Patients with consecutive admissions to a regional stroke center for new-onset unilateral hemispheric stroke who met World Health Organization and National Institute of Neurological and Communicative Disorders and Stroke criteria were eligible for inclusion in a longitudinal study. Acutely, patients underwent CT scanning, and at 3 months and 1 year after stroke, depressive symptoms were assessed by using both the Montgomery-Asberg Depression Rating Scale and the Zung Self-Rating Depression Scale. The Functional Independence Measure (FIM) served as an indication of functional outcome and was obtained at 1 month, 3 months, and 1 year after stroke, along with other demographic information. The Talairach and Tournoux stereotactic atlas was used for the primary determination of CT lesion localization. Lesion proximity to the anterior frontal pole was also measured. RESULTS: Eighty-one patients participated in the longitudinal study. Stepwise linear regression analyses generated a highly significant model (F(3,76)=9.8, R(2)=28%, P<0.0005), with lower 1-month total FIM scores, living at home, and damage to the inferior frontal region predicting higher depression scores at 3 months. Similarly, lower 3-month total FIM scores correlated with higher 3-month depression scores, and lower 1-year total FIM scores correlated with higher 1 year depression scores. CONCLUSIONS: Functional measures correlated with poststroke depression across time and, together with neuroanatomic measures, predicted depressive symptoms longitudinally. Although inferior frontal lesion location, irrespective of side, appeared to play a role as a risk factor in this study, the degree of functional dependence after stroke imparted the greatest risk. PMID- 10700498 TI - Neurobehavioral outcome prediction after cardiac surgery: role of neurobiochemical markers of damage to neuronal and glial brain tissue. AB - BACKGROUND AND PURPOSE: The goal of the present study was to investigate the predictive value of neurobiochemical markers of brain damage (protein S-100B and neuron-specific enolase [NSE]) with respect to the short- and long-term neuropsychological outcomes after cardiac surgery with cardiopulmonary bypass (CPB). METHODS: We investigated 74 patients who underwent elective CABG or valve replacement surgery and who showed no severe neurological deficits after surgery. Patients were investigated with a standardized neurological examination and a comprehensive neuropsychological and neuropsychiatric assessment 1 to 2 days before surgery, 3 and 8 days after surgery, and 6 months later. Serial venous blood samples were taken preoperatively and 1, 6, 20, and 30 hours after skin closure. Protein S-100B and NSE were analyzed with immunoluminometric assays. RESULTS: Patients with severe postoperative neuropsychological disorders showed a significantly higher and longer release of neurobiochemical markers of brain damage. Patients who presented with a delirium according to DSM-III-R criteria 3 days after surgery had significantly higher postoperative S-100B serum concentrations. Multivariate analysis (based on postoperative NSE and S-100B concentrations and age of patients, type of operation, length of cross-clamp and perfusion time, and intraoperative and postoperative oxygenation) identified NSE and S-100B concentrations 6 to 30 hours after skin closure as the only variables that contributed significantly to a predictive model of the neuropsychological outcome. NSE, but not S-100B, release was significantly higher in patients undergoing valve replacement surgery. CONCLUSIONS: Postoperative serum concentrations and kinetics of S-100B and NSE have a high predictive value with respect to the early neuropsychological and neuropsychiatric outcome after cardiac surgery. The analysis of NSE and S-100B release might allow insight into the underlying pathophysiology of brain dysfunction, thus providing a valuable tool to monitor and evaluate measures to improve cardiac surgery with CPB. PMID- 10700499 TI - Decreased severity of brain infarct can in part explain the decreasing case fatality rate of stroke. AB - BACKGROUND AND PURPOSE: Case fatality rates for stroke has declined in most Western industrialized countries during recent decades. One possible explanation for this is a decrease in the severity of stroke symptoms. We therefore sought evidence for a change in stroke severity and its relationship with case fatality rates. METHODS: We compared the severity of symptoms among first-ever stroke patients in 2 population-based prospective stroke registers maintained during 1972 to 1973 and 1989 to 1991 in Finland. Patients who were evaluated by study assistants or the investigator during the first week after the onset of symptoms were included in the study, and their severity of symptoms was assessed with the use of comparable scales modified from the Scandinavian Stroke Scale. RESULTS: A total of 244 and 594 patients were registered, and a portion of them (155 [63.5%] and 360 [60.6%]) were included in the analyses in the registers for Espoo Kauniainen from 1972 to 1973 and for 4 separate districts in Finland from 1989 to 1991, respectively. The death rates during the first week among those who were not included did not differ between the registers. The severity of symptoms decreased significantly between the registers in both patients with brain infarct or intracerebral hemorrhage but not in those with subarachnoid hemorrhage. The severity of symptoms was an independent factor of case fatality at 1 month. CONCLUSIONS: The severity of symptoms of brain infarcts has decreased and can in part explain the decreased case fatality rate of stroke in Finland. However, the change in patients with intracerebral hemorrhage may be overestimated due to undiagnosed intracerebral hemorrhages in the first register resulting from the lack of brain CT. PMID- 10700500 TI - Evolution of cortical activation during recovery from corticospinal tract infarction. AB - BACKGROUND AND PURPOSE: Recovery from hemiparesis due to corticospinal tract infarction is well documented, but the mechanism of recovery is unknown. Functional MRI (fMRI) provides a means of identifying focal brain activity related to movement of a paretic hand. Although prior studies have suggested that supplementary motor regions in the ipsilesional and contralesional hemisphere play a role in recovery, little is known about the time course of cortical activation in these regions as recovery proceeds. METHODS: Eight patients with first-ever corticospinal tract lacunes causing hemiparesis had serial fMRIs within the first few days after stroke and at 3 to 6 months. Six healthy subjects were used as controls. Statistically significant voxels during a finger-thumb opposition task were identified with an automated image processing program. An index of ipsilateral versus contralateral activity was used to compare relative contributions of the 2 hemispheres to motor function in the acute and chronic phases after stroke. RESULTS: Controls showed expected activation in the contralateral sensorimotor cortex (SMC), premotor, and supplementary motor areas. Stroke patients differed from control patients in showing greater activation in the ipsilateral SMC, ipsilateral posterior parietal, and bilateral prefrontal regions. Compared with the nonparetic hand, the ratio of contralateral to ipsilateral SMC activity during movement of the paretic hand increased significantly over time as the paretic hand regained function. CONCLUSIONS: The evolution of activation in the SMC from early contralesional activity to late ipsilesional activity suggests that a dynamic bihemispheric reorganization of motor networks occurs during recovery from hemiparesis. PMID- 10700501 TI - Objective measurement of functional upper-extremity movement using accelerometer recordings transformed with a threshold filter. AB - BACKGROUND AND PURPOSE: The consensus is that the most important outcome for rehabilitation is functional activity in the life situation. Constraint-Induced Movement Therapy, a new treatment that transfers in-clinic gains to the life situation, demands objective measurement of real-world movement. However, direct, objective, and accurate measures of arm use in the real world are not available. Previous attempts to use accelerometry to measure extremity movement have failed because of unacceptable variability. This problem has been addressed here by use of a threshold filter. METHODS: Nine stroke patients and 1 healthy individual wearing accelerometers were videotaped while they carried out their usual activities at home or in the clinic; the duration of their arm, torso, and ambulatory movements was judged by 2 observation teams. In addition, 11 college students performed 5 standardized activities of daily living for varying durations in the laboratory. The accelerometer data were transformed; the raw value recorded for a given epoch was set to a constant if it exceeded a low threshold. RESULTS: The threshold-filtered recordings measured the duration of movement accurately and with very little variability. Correlations between the threshold-filtered recordings and the observer ratings of the duration of arm, torso, and ambulatory movements were 0.93, 0.93 and 0.99, respectively; the corresponding correlations for the raw values were -0.17, 0.34, and 0.85. CONCLUSIONS: These results present initial evidence for the validity of threshold filtered accelerometer recordings for objectively measuring the amount of real world upper-extremity movement as an index of treatment outcome for rehabilitation patients. PMID- 10700502 TI - A pilot study of somatotopic mapping after cortical infarct. AB - BACKGROUND AND PURPOSE: Animal studies have described remodeling of sensory and motor representational maps after cortical infarct. These changes may contribute to return of function after stroke. METHODS: Functional MRI was used to compare sensory and motor maps obtained in 35 normal control subjects with results from 2 patients with good recovery 6 months after a cortical stroke. RESULTS: During finger tapping in controls, precentral gyrus activation exceeded or matched postcentral gyrus activation in 40 of 42 cases. Patient 1 had a small infarct limited to precentral gyrus. Finger tapping activated only postcentral gyrus, a pattern not seen in any control subject. During tactile stimulation of a finger or hand in controls, postcentral gyrus activation exceeded or matched precentral gyrus activation in 11 of 14 cases. Patient 2 had a small infarct limited to postcentral gyrus and superior parietal lobule. Tactile stimulation of the finger activated only precentral gyrus, a pattern not seen in any control. In both patients, activation during pectoralis contraction was medial to the site activated during finger tapping. CONCLUSIONS: Results during finger tapping (patient 1) and finger stimulation (patient 2) may reflect amplification of a preserved component of normal sensorimotor function, a shift in the cortical site of finger representation, or both. Cortical map reorganization along the infarct rim may be an important contributor to recovery of motor and sensory function after stroke. Functional MRI is useful for assessing motor and sensory representational maps. PMID- 10700503 TI - Relating MRI changes to motor deficit after ischemic stroke by segmentation of functional motor pathways. AB - BACKGROUND AND PURPOSE: Infarct size on T2-weighted MRI correlates only modestly with outcome, particularly for small strokes. This may be largely because of differences in the locations of infarcts and consequently in the functional pathways that are damaged. To test this hypothesis quantitatively, we developed a "mask" of the corticospinal pathway to determine whether the extent of stroke intersection with the pathway would be more closely related to clinical motor deficit and axonal injury in the descending motor pathways than total stroke lesion volume. METHODS: Eighteen patients were studied > or =1 month after first ischemic stroke that caused a motor deficit by use of brain T2-weighted imaging, MR spectroscopic (MRS) measurements of the neuronal marker compound N-acetyl aspartate in the posterior limb of the internal capsule, and motor impairment and disability measures. A corticospinal mask based on neuroanatomic landmarks was generated from a subset of the MRI data. The maximum proportion of the cross sectional area of this mask occupied by stroke was determined for each patient after all brain images were transformed into a common stereotaxic brain space. RESULTS: There was a significant linear relationship between the maximum proportional cross-sectional area of the corticospinal mask occupied by stroke and motor deficit (r(2)=0.82, P<0.001), whereas the relationship between the total stroke volume and motor deficit was better described by a cubic curve (r(2)=0.76, P<0.001). Inspection of the data plots showed that the total stroke volume discriminated poorly between smaller strokes with regard to the extent of associated motor deficit, whereas the maximum proportion of the mask cross sectional area occupied by stroke appeared to be a more discriminatory marker of motor deficit and also N-acetyl aspartate reduction. CONCLUSIONS: Segmentation of functional motor pathways on MRI allows estimation of the extent of damage specifically to that pathway by the stroke lesion. The extent of stroke intersection with the motor pathways was more linearly related to the magnitude of motor deficit than total lesion volume and appeared to be a better discriminator between small strokes with regard to motor deficit. This emphasizes the importance of the anatomic relationship of the infarct to local structures in determining functional impairment. Prospective studies are necessary to assess whether this approach would allow improved early estimation of prognosis after stroke. PMID- 10700504 TI - Magnetic resonance perfusion imaging in acute ischemic stroke using continuous arterial spin labeling. AB - BACKGROUND AND PURPOSE: Continuous arterial spin-labeled perfusion MRI (CASL-PI) uses electromagnetically labeled arterial blood water as a diffusible tracer to noninvasively measure cerebral blood flow (CBF). We hypothesized that CASL-PI could detect perfusion deficits and perfusion/diffusion mismatches and predict outcome in acute ischemic stroke. METHODS: We studied 15 patients with acute ischemic stroke within 24 hours of symptom onset. With the use of a 6-minute imaging protocol, CASL-PI was measured at 1.5 T in 8-mm contiguous supratentorial slices with a 3.75-mm in-plane resolution. Diffusion-weighted images were also obtained. Visual inspection for perfusion deficits, perfusion/diffusion mismatches, and effects of delayed arterial transit was performed. CBF in predetermined vascular territories was quantified by transformation into Talairach space. Regional CBF values were correlated with National Institutes of Health Stroke Scale (NIHSS) score on admission and Rankin Scale (RS) score at 30 days. RESULTS: Interpretable CASL-PI images were obtained in all patients. Perfusion deficits were consistent with symptoms and/or diffusion-weighted imaging abnormalities. Eleven patients had hypoperfusion, 3 had normal perfusion, and 1 had relative hyperperfusion. Perfusion/diffusion mismatches were present in 8 patients. Delayed arterial transit effect was present in 7 patients; serial imaging in 2 of them showed that the delayed arterial transit area did not succumb to infarction. CBF in the affected hemisphere correlated with NIHSS and RS scores (P=0.037 and P=0.003, Spearman rank correlation). The interhemispheric percent difference in middle cerebral artery CBF correlated with NIHSS and RS scores (P=0.007 and P=0.0002, respectively). CONCLUSIONS: CASL-PI provides rapid noninvasive multislice imaging in acute ischemic stroke. It depicts perfusion deficits and perfusion/diffusion mismatches and quantifies regional CBF. CASL-PI CBF asymmetries correlate with severity and outcome. Delayed arterial transit effects may indicate collateral flow. PMID- 10700505 TI - Significance of acute multiple brain infarction on diffusion-weighted imaging. AB - BACKGROUND AND PURPOSE: Diffusion-weighted imaging (DWI) is superior to conventional MRI in identification of small new ischemic lesions and discrimination of recent infarcts from old ones. Thus, this technique is useful in the detection of acute multiple brain infarcts (AMBI). We sought to determine the frequency and the topographical and etiologic patterns of AMBI detected on DWI. METHODS: We studied 329 consecutive ischemic stroke patients who underwent DWI and MRI/MR angiography within 4 days of stroke onset. AMBI was defined as noncontiguous high signal intensities on DWI in >1 vascular territory. Stroke mechanism was determined according to the criteria of the Trial of Org 10172 in Acute Stroke Treatment (TOAST). RESULTS: We detected AMBI in 95 patients (28.9%). AMBI in anterior circulation was found in 62 cases: in 1 hemisphere in 42 (group A) and in bilateral hemispheres in 20 (group B). Twenty-two patients had AMBI in the posterior circulation (group C) and 11 in both anterior and posterior circulations (group D). The most frequent cause of stroke was large-artery atherosclerosis in groups A (33/42), B (9/20), and C (15/22) (P=0.02) and cardioembolism in group D (6/11) (P=0.02). Elevated fibrinogen or hematocrit was significantly associated with group B (P=0.01). In 9 patients in groups B and D, anatomic variations of anterior or posterior cerebral arteries or patent posterior communicating artery contributed to AMBI. CONCLUSIONS: Different topographical patterns of AMBI are associated with different vascular pathologies and stroke mechanisms. Hemorheologic abnormality or vascular anatomic variations may be contributing factors in the pathogenesis of AMBI in bilateral cerebral hemispheres or in both anterior and posterior circulations. PMID- 10700506 TI - MRI pontine hyperintensity after supratentorial ischemic stroke relates to poor clinical outcome. AB - BACKGROUND AND PURPOSE: MRI studies in patients with atherosclerosis often reveal ill-defined hyperintensity in the pons on T2-weighted images. This pontine hyperintensity (PHI) does not fulfill the criteria of a brain infarct, and its clinical relevance is not established. We examined the frequency, as well as the radiological and clinical correlates, of PHI in poststroke patients. METHODS: Three hundred nineteen patients were studied 3 months after supratentorial ischemic stroke with the use of 1.0-T MRI. Brain infarcts, atrophy, white matter hyperintensities, and PHI were registered. The clinical outcome was assessed 3 and 15 months after the stroke. RESULTS: Of the patients, 152 (47.6%) had PHI. The risk factors for stroke did not differ in patients without or with PHI. PHI was related to a higher frequency (P=0.002) and larger volume (P<0.001) of supratentorial brain infarcts, to parietal (P=0.020) and temporal (P=0.002) atrophy, to central atrophy (P< or =0.040), and to white matter hyperintensity grade (P<0.001). Brain infarcts that affected the corpus striatum (putamen, caudate, and pallidum) (P< or =0. 011) or pyramidal tract (P<0.001) were more frequent in patients with PHI. The 3- and 15-month outcomes were worse in patients with PHI (P< or =0.004). The total volume of brain infarcts (OR 1.22), mean atrophy (OR 3.59), and PHI (OR 3.76) were independent correlates of a poor 15-month outcome. CONCLUSIONS: PHI after supratentorial ischemic stroke deserves attention because it relates to poor clinical outcome. PMID- 10700507 TI - Carotid and transcranial color-coded duplex sonography in different types of carotid-cavernous fistula. AB - BACKGROUND AND PURPOSE: Patients with carotid-cavernous fistula (CCF) may undergo direct or indirect shunting. Ultrasonography has value that is complementary to angiography in the assessment and follow-up of these patients. The aim of this study was to characterize findings provided by carotid duplex sonography (CDS) and transcranial color-coded duplex sonography (TCCD) in patients with different types of CCF. METHODS: CDS and TCCD were independently performed by technologists and neurologists. Digital subtraction or MR angiography was interpreted by a neuroradiologist. Ultrasonographic studies were categorized into 4 types: I, direct shunting only; II, direct shunting with a carotid aneurysm; III, indirect shunting only; and IV, mixed (direct and indirect) shunting. In addition to carotid and intracranial flow velocities, volume, and pulsatility, other direct and indirect ultrasound signs of shunting were evaluated. The direct sign of CCF was a mosaic flash detected by TCCD. Alteration of hemodynamic parameters on CDS and demonstration of draining veins with the use of TCCD were considered indirect signs. RESULTS: Fifteen patients (8 men, 7 women) were included in the study. According to angiographic results, patients in ultrasonographic classification types I (n=7) and II (n=3) corresponded to type A of Barrow's classification. Patients with type III (n=8) were Barrow's type C. Type IV (n=1) had a combination of Barrow's types A and C. On ultrasound, both direct and indirect signs were seen in types I, II, and IV CCF. The presence of a 2-colored oval mass divided by a zone of separation without turbulence differentiated type I from type II CCF. All patients with type III CCF had indirect signs, and only 1 patient had direct signs on TCCD. Abnormal TCCD findings were most commonly seen through the transorbital window (100%), followed by the transtemporal window (63%) and transforaminal window (40%). CONCLUSIONS: If only indirect ultrasonographic signs of CCF are present, TCCD can be used to predict an indirect CCF type on the basis of the origin of the fistula. With direct communication between carotid artery and cavernous sinus, both direct and indirect ultrasonographic signs can be found. The combination of CDS/TCCD may provide a noninvasive and reliable way to classify patients with CCF. PMID- 10700508 TI - Longer duration of cardiopulmonary bypass is associated with greater numbers of cerebral microemboli. AB - BACKGROUND AND PURPOSE: Many patients who undergo cardiac surgery assisted with cardiopulmonary bypass (CPB) experience cerebral injury, and microemboli are thought to play a role. Because an increased duration of CPB is associated with an increased risk of subsequent cerebral dysfunction, we investigated whether cerebral microemboli were also more numerous with a longer duration of CPB. METHODS: Brain specimens were obtained from 36 patients who died within 3 weeks after CPB. Specimens were embedded in celloidin, sectioned 100 microm thick, and stained for endogenous alkaline phosphatase, which outlines arterioles and capillaries. In such preparations, emboli can be seen as swellings in the vessels. Cerebral microemboli were counted in equal areas and scored as small, medium, or large to estimate the embolic load (volume of emboli). RESULTS: With increasing survival time after CPB, the embolic load declined (P<0.0001). (Lipid emboli are known to pump slowly through the brain.) Also with increasing time after CPB, the percentage of large and medium emboli became lower (P=0.0034). This decline is consistent with the concept that the emboli break into smaller globules as they pass through the capillary network. A longer duration of CPB was associated with increased embolic load (P=0. 0026). For each 1-hour increase in the duration of CPB, the embolic load increased by 90.5%. CONCLUSIONS: Thousands of microemboli were found in the brains of patients soon after CPB, and an increasing duration of CPB was associated with an increasing embolic load. PMID- 10700509 TI - Relationship between pattern of intracranial artery abnormalities on transcranial doppler and Oxfordshire Community Stroke Project clinical classification of ischemic stroke. AB - BACKGROUND AND PURPOSE: The Oxfordshire Community Stroke Project (OCSP) devised a simple classification for acute stroke based on clinical features only, which is of value in predicting prognosis. We investigated whether the pattern of intracranial vascular abnormalities is related to the clinical syndrome. METHODS: Patients with acute ischemic stroke were classified by a stroke physician as having total or partial anterior circulation infarct (TACI or PACI, respectively), lacunar infarct (LACI), or posterior circulation infarct (POCI). Color-coded power transcranial Doppler was done whenever possible. Intracranial arterial velocities were compared in the 4 subtypes of ischemic stroke after adjustment for age and time to transcranial Doppler. RESULTS: Middle cerebral artery velocity was abnormal (hyperemia, reduced velocity, occlusion, or focal stenosis) in 38 of 69 TACIs (55%), 50 of 171 PACIs (29%), and 20 of 236 LACIs or POCIs (8%) (P<0.001). Velocity in the A1 segment of the anterior cerebral artery was reversed in 12 of 69 TACIs (17%), 20 of 171 PACIs (12%), and 8 of 236 LACIs or POCIs (3%) (P<0.001). Basilar artery velocity was abnormal in 8 of 121 POCIs (7%) compared with 5 of 355 (1%) of the other subtypes (P=0.005). Vertebral artery velocity was abnormal (reduced velocity, occlusion, stenosis) in 20 of 121 POCIs (17%) compared with 20 of 355 others (6%) (P=0.01). CONCLUSIONS: Intracranial arterial abnormalities were related to OCSP clinical subtype. Therefore, it is possible to stratify patients according to OCSP classification in trials of new treatments in which treatment effectiveness may depend on the underlying pattern of arterial pathology and before any arterial imaging is available. PMID- 10700510 TI - Plasma endothelin-1 levels neither increase nor correlate with neurological scores, stroke risk factors, or outcome in patients with ischemic stroke. AB - BACKGROUND AND PURPOSE: Endothelins (ETs) are potent vasoconstrictors and may play a role in the pathophysiology of several diseases. Limited and controversial data exist on their role in human ischemic stroke. We planned a prospective, observational, and longitudinal clinical study to test whether ET-1 levels increase in various phases of ischemic stroke and whether the ET-1 levels correlate with neurological scores, stroke etiology, stroke risk factors, or final outcome. METHODS: We measured plasma ET-1 levels with a sandwich-enzyme immunoassay method in 101 consecutive patients with ischemic stroke on admission and 1 week, 1 month, and 3 months after stroke and in 101 sex- and age-matched control subjects. At each sampling, the patients underwent a complete neurological evaluation. All stroke risk factors were recorded, an array of laboratory tests were performed, and the subtype of ischemic stroke was determined. The patients were contacted 3 years later for prognostic determination. RESULTS: ET-1 levels in patients (2.4+/-1.3 pg/mL on admission, 2.2+/-1.4 pg/mL at 1 week, 2.1+/-1.4 pg/mL at 1 month, and 2.1+/-1.2 pg/mL at 3 months) were not different from those of the control subjects (2.2+/-0.9 pg/mL) at any time point. No correlation was found between the ET-1 levels and stroke etiology, stroke risk factors, stroke recurrence risk, age, sex, or neurological scores, except that ET-1 levels correlated with the use of warfarin and with body mass index. CONCLUSIONS: Plasma ET-1 levels were normal in patients with ischemic stroke. Our findings cannot exclude a role of ETs in the pathophysiology of ischemic stroke because plasma levels might not accurately reflect intracerebral concentrations, but they also do not support the occurrence of a major plasma ET 1 level increase at any phase of stroke. Our patient population is the largest ever reported in whom ET-1 levels were measured, but it consisted of mild and moderately ill patients with stroke due to the study design, of which the aim was long-term observation, which excludes severely ill patients. PMID- 10700511 TI - Diffusion-weighted MRI and proton MR spectroscopic imaging in the study of secondary neuronal injury after intracerebral hemorrhage. AB - BACKGROUND AND PURPOSE: Cerebral ischemia has been proposed as contributing mechanism to secondary neuronal injury after intracerebral hemorrhage (ICH). Possible tools for investigating this hypothesis are diffusion-weighted (DWI) and proton magnetic resonance spectroscopic imaging ((1)H-MRSI). However, magnetic field inhomogeneity induced by paramagnetic blood products may prohibit the application of such techniques on perihematoma tissue. We report on the feasibility of DWI and (1)H-MRSI in the study of human ICH and present preliminary data on their contribution to understanding perihematoma tissue functional and metabolic profiles. METHODS: Patients with acute supratentorial ICH were prospectively evaluated using DWI and (1)H-MRSI. Obscuration of perihematoma tissue with both sequences was assessed. Obtainable apparent diffusion coefficient (Dav) and lactate spectra in perihematoma brain tissue were recorded and analyzed. RESULTS: Nine patients with mean age of 63.4 (36 to 87) years were enrolled. Mean time from symptom onset to initial MRI was 3.4 (1 to 9) days; mean hematoma volume was 35.4 (5 to 80) cm(3). Perihematoma diffusion values were attainable in 9 of 9 patients, and (1)H-MRSI measures were obtainable in 5 of 9 cases. Dav in perihematoma regions was 172.5 (120.0 to 302.5)x10(-5) mm(2)/s and 87.6 (76.5 to 102.1)x10(-5) mm(2)/s in contralateral corresponding regions of interest (P=0.002). One patient showed an additional area of reduced Dav with normal T(2) intensity, which suggests ischemia. (1)H-MRSI revealed lactate surrounding the hematoma in 2 patients. CONCLUSIONS: DWI and (1)H-MRSI can be used in the study of ICH patients. Our preliminary data are inconsistent with ischemia as the primary mechanism for perihematoma tissue injury. Further investigation with advanced MRI techniques will give a clearer understanding of the role that ischemia plays in tissue injury after ICH. PMID- 10700512 TI - Renal artery lesions in patients with moyamoya disease: angiographic findings. AB - BACKGROUND AND PURPOSE: Renal artery lesions in moyamoya disease have been described sporadically in several case reports. The purpose of this study is to evaluate the angiographic findings of renal artery lesions in moyamoya disease and to determine the prevalence of renal artery lesions in patients with moyamoya disease. METHODS: Eighty-six consecutive patients with idiopathic moyamoya disease were prospectively examined with both cerebral angiography and abdominal aortography. The findings of abdominal aortography were reviewed for the presence and appearance of renal artery lesions and compared with the clinical data and cerebral angiographic findings. RESULTS: Of 86 patients with idiopathic moyamoya disease, 7 patients (8%) were found to have renal artery lesions. Six patients (7%) had stenosis in the renal artery, and 1 patient (1%) had a small saccular aneurysm in the renal artery. Two patients (2%) with a marked renal artery stenosis presented with renovascular hypertension, which resulted in an intraventricular hemorrhage in 1 patient. Furthermore, the renal artery stenosis in the 2 patients with renovascular hypertension was successfully treated with percutaneous transluminal angioplasty. There was no significant correlation between the presence of renal artery lesions and cerebral angiographic findings. CONCLUSIONS: Seven (8%) of 86 patients with moyamoya disease showed renal artery lesions, including 6 stenoses (7%) and 1 aneurysm (1%). Renal artery lesions are a clinically relevant systemic manifestation in patients with moyamoya disease. PMID- 10700513 TI - Stroke in estrogen receptor-alpha-deficient mice. AB - BACKGROUND AND PURPOSE: Recent evidence suggests that endogenous estrogens or hormone replacement therapy can ameliorate brain damage from experimental stroke. Protective mechanisms involve enhanced cerebral vasodilation during ischemic stress as well as direct preservation of neuronal viability. We hypothesized that if the intracellular estrogen receptor subtype-alpha (ERalpha) is important to estrogen's signaling in the ischemic brain, then ERalpha-deficient (knockout) (ERalphaKO) female mice would sustain exaggerated cerebral infarction damage after middle cerebral artery occlusion. METHODS: The histopathology of cresyl violet-stained tissues was evaluated after reversible middle cerebral artery occlusion (2 hours, followed by 22 hours of reperfusion) in ERalphaKO transgenic and wild-type (WT) mice (C57BL/6J background strain). End-ischemic cerebral blood flow mapping was obtained from additional female murine cohorts by using [(14)C]iodoantipyrine autoradiography. RESULTS: Total hemispheric tissue damage was not altered by ERalpha deficiency in female mice: 51.9+/-10.6 mm(3) in ERalphaKO versus 60.5+/-5.0 mm(3) in WT. Striatal infarction was equivalent, 12.2+/-1.7 mm(3) in ERalphaKO and 13.4+/-1.0 mm(3) in WT mice, but cortical infarction was paradoxically smaller relative to that of the WT (20.7+/-4.5 mm(3) in ERalphaKO versus 30.6+/-4.1 mm(3) in WT). Intraocclusion blood flow to the parietal cortex was higher in ERalphaKO than in WT mice, likely accounting for the reduced infarction in this anatomic area. There were no differences in stroke outcomes by region or genotype in male animals. CONCLUSIONS: Loss of ERalpha does not enhance tissue damage in the female animal, suggesting that estrogen inhibits brain injury by mechanisms that do not depend on activation of this receptor subtype. PMID- 10700514 TI - Estradiol exerts neuroprotective effects when administered after ischemic insult. AB - BACKGROUND AND PURPOSE: 17beta-Estradiol (E2) has been reported to exert neuroprotective effects when administered before an ischemic insult. This study was designed to determine whether E2 treatment after ischemia exerts the same effects and, if so, how long this therapeutic window remains open, and whether the effects are related to changes in cerebral blood flow (CBF). METHODS: Female Sprague-Dawley rats were subjected to permanent middle cerebral artery occlusion (MCAO). In protocol 1, E2 was administered (100 microg/kg IV followed immediately by subcutaneous implantation of crystalline E2 in a silicone elastomer tube) to ovariectomized females (OVX+E2) at 0.5 (n=8), 1 (n=6), 2 (n=7), 3 (n=6), or 4 (n=9) hours after MCAO. Intact (INT; n=6) and ovariectomized females (OVX; n=12) were subjected to MCAO and received vehicle instead of E2. Two days after MCAO the animals were killed, and ischemic lesion volume was determined by 2,3,5 triphenyltetrazolium chloride staining. In protocol 2, CBF was monitored before and at 1, 24, and 48 hours in a group of animals receiving E2 or vehicle 0.5 hour after ischemia induction (INT, n=6; OVX, n=8; OVX+E2, n=6). RESULTS: Lesion volume was 20.9+/-2.2% and 21.8+/-1.2% in the INT and OVX groups, respectively. E2 was found to decrease lesion volume significantly when administered within 3 hours after MCAO. The lesion volumes were 6.3+/-0.5%, 10.3+/-2.1%, 11.8+/-1.8%, 13.5+/-1.6%, and 17.9+/-2.8% when E2 was administered at 0.5, 1, 2, 3, or 4 hours after MCAO, respectively. CBF decreased to 43.1+/-2.2% and 25.4+/-1.0% in the INT and OVX animals, respectively, at 5 minutes after MCAO. In comparison to OVX rats, CBF was not different at 1 hour after E2 administration but was increased significantly in the OVX+E2 group 1 and 2 days after E2 administration. CONCLUSIONS: E2 exerts neuroprotective effects when administered after ischemia, with a therapeutic window in a permanent focal cerebral ischemia model of approximately 3 hours. This effect of estradiol was associated with no immediate change in blood flow but with a delayed increase in CBF. PMID- 10700515 TI - Electromechanical alterations in the cerebrovasculature of stroke-prone rats. AB - BACKGROUND AND PURPOSE: Cerebrovascular pressure-dependent constriction (PDC) is associated with smooth muscle (SM) depolarization and Ca(2+) influx through voltage-gated channels. We studied the alterations in electromechanical contraction in the middle cerebral arteries (MCAs) of stroke-prone Wistar-Kyoto spontaneously hypertensive rats (SHRsp) in relation to the stroke-related loss of PDC. METHODS: Constriction to pressure, elevated [K(+)](o) and/or [Ca(2+)](o), and SM membrane potentials (E(m)) were measured in isolated pressurized MCAs of SHRsp and stroke-resistant SHR. RESULTS: MCAs of SHRsp exhibited an age-related decrease in PDC before hemorrhagic stroke and a loss of PDC after stroke. At 100 mm Hg, the MCAs of poststroke SHRsp maintained partial constriction that was not altered with pressure but was inhibited by nifedipine (1 micromol/L). The MCAs of poststroke SHRsp constricted to vasopressin (0.17 micromol/L) but not to elevated [K(+)](o). When pressure was reduced from 100 to 0 mm Hg, the MCAs from young prestroke SHRsp exhibited SM hyperpolarization (-38 to -46 mV), whereas those of poststroke SHRsp maintained a constant, depolarized E(m) (-34 mV). Alterations in E(m) with varying [K(+)](o) suggested that there was a decrease in SM K(+) conductance in the MCAs of poststroke SHRsp. CONCLUSIONS: The observation that the MCAs of poststroke SHRsp depolarize but do not constrict to elevated [K(+)](o) suggests the presence of dysfunctional voltage-gated Ca(2+) channels. The inability to alter E(m) with pressure or to constrict to depolarization could partially contribute to the loss of PDC in the MCAs of poststroke SHRsp. PMID- 10700516 TI - Impaired endothelial function in transgenic mice expressing both human renin and human angiotensinogen. AB - BACKGROUND AND PURPOSE: Chronic hypertension is a risk factor for carotid vascular disease and stroke. Mechanisms that account for alterations in carotid and cerebral vascular function during hypertension are poorly defined and based almost exclusively on studies in the spontaneously hypertensive rat, a model in which hypertension has an unknown etiology and in which the genetic background is dissimilar to the most commonly used normotensive control, the Wistar-Kyoto rat. METHODS: In this study we examined vascular function in a defined model of hypertension, double transgenic mice that overexpress both human renin (R+) and human angiotensinogen (A+). We studied vessels in vitro from R+/A+ mice as well as nontransgenic (R-/A-) and single transgenic (R-/A+ or R+/A-) littermate controls. RESULTS: After submaximal precontraction with U46619 or prostaglandin F(2alpha), acetylcholine, which produces relaxation mediated by endothelial nitric oxide synthase, produced marked relaxation of carotid arteries in control mice but was impaired in R+/A+ mice. For example, 1 micromol/L acetylcholine relaxed the carotid artery by 79+/-4% versus 44+/-7% (P<0.01) in control and R+/A+ mice, respectively. Impaired responses to acetylcholine in R+/A+ mice could be restored toward normal with indomethacin (10 micromol/L). In contrast, relaxation of the carotid artery in response to nitroprusside and papaverine was similar in R+/A+ mice and control mice. CONCLUSIONS: These findings indicate that acetylcholine-induced relaxation of carotid artery is impaired selectively in mice made hypertensive by expression of human renin and human angiotensinogen. The mechanism of this impairment may involve production of a cyclooxygenase derived contracting factor. PMID- 10700517 TI - Is pharmacological neuroprotection dependent on reduced glutamate release? AB - BACKGROUND AND PURPOSE: The aim of this study was to determinate the possible role of the ionotropic glutamate receptor in the expression of irreversible electrophysiological changes induced by in vitro ischemia and to test whether the neuroprotective action of various neurotransmitter agonists and drugs of clinical interest is related to a presynaptic inhibitory action at glutamatergic synapses. METHODS: Intracellular and extracellular recordings have been performed in a rat corticostriatal slice preparation. Different pharmacological compounds have been tested on corticostriatal glutamatergic transmission in control conditions and in an in vitro model of ischemia (oxygen and glucose deprivation). RESULTS: In vitro ischemia lasting 10 minutes produced an irreversible loss of the field potential recorded from striatal slices after cortical stimulation. Preincubation of the slices with 3 micromol/L 6-cyano-7-nitroquinoxaline-2,3-dione (an alpha-amino-3 hydroxy-5-methylisoxazole-4-propionic acid [AMPA] receptor antagonist) allowed a significant recovery of the field potential amplitude (P<0.05, n=6), whereas incubation with 30 micromol/L aminophosphonovaleric acid (an N-methyl-D-aspartate receptor antagonist) did not produce a significant recovery after 10 minutes of ischemia (P>0.05, n=7). Bath application of 3 mmol/L glutamate for 5 minutes produced a complete but reversible inhibition of the field potential amplitude. When a similar application was coupled with a brief period of ischemia (5 minutes), which produced, per se, only a transient inhibition of the field potential, it caused an irreversible loss of this parameter. We also tested the possible neuroprotective effect of neurotransmitter agonists reducing the release of glutamate from corticostriatal terminals. Agonists acting on purinergic (adenosine), muscarinic (oxotremorine), and metabotropic glutamate receptors (L serine o-phosphate [L-SOP]) significantly (P<0.001, n=8 for each agonist) reduced glutamatergic synaptic potentials, with each showing different potencies. The EC(50) was 26.4 micromol/L for adenosine, 0. 08 micromol/L for oxotremorine, and 0.89 micromol/L for L-SOP. Concentrations of these agonists producing the maximal inhibition of the synaptic potential were tested on the ischemia-induced irreversible loss of field potential. Adenosine (P<0.05, n=9) and oxotremorine (P<0.05, n=8) showed significant neuroprotective action, whereas L-SOP was ineffective (P>0.05, n=10). Similarly, putative neuroprotective drugs significantly (P<0.001, n=10 for each drug) reduced the amplitude of corticostriatal potential, with different EC(50) values (phenytoin, 33.5 micromol/L; gabapentin, 96.8 micromol/L; lamotrigine, 26.7 micromol/L; riluzole, 6 micromol/L; and sipatrigine, 2 micromol/L). Concentration of these drugs producing maximal inhibition of the amplitude of corticostriatal potentials showed a differential neuroprotective action on the ischemic electrical damage. Phenytoin (P<0.05, n=10), lamotrigine (P<0.05, n=10), riluzole (P<0.05, n=9), and sipatrigine (P<0.001, n=10) produced a significant neuroprotection, whereas gabapentin (P>0.05, n=11) was ineffective. The neuroprotective action of transmitter agonists and clinical drugs was not related to their ability in decreasing glutamate release, as detected by changes in the paired-pulse facilitation protocol. CONCLUSIONS: Ionotropic glutamate receptors, and particularly AMPA-like receptors, play a role in the irreversible loss of field potential amplitude induced by ischemia in the striatum. Drugs acting by reducing glutamatergic corticostriatal transmission may show a neuroprotective effect. However, their efficacy does not seem to be directly related to their capability to decrease glutamate release from corticostriatal terminals. We suggest that additional modulatory actions on voltage-dependent conductances and on ischemia induced ion distribution at the postsynaptic site may also exert a crucial role. PMID- 10700518 TI - The symptomatic carotid plaque. AB - BACKGROUND: The natural histories of equally severe symptomatic and asymptomatic carotid stenoses are very different, which suggests dichotomy in plaque behavior. The vascular biology of the symptomatic carotid plaque is presented in this review. SUMMARY OF REVIEW: Histology studies comparing asymptomatic and symptomatic plaques were identified from MEDLINE. Reports in which stenosis severity was not stated or not similar for symptomatic and asymptomatic patients were excluded. In vitro studies and reports from the coronary circulation were reviewed with regard to the vascular biology of the plaque. Histology studies comparing carotid plaques removed from symptomatic and asymptomatic patients reveal characteristic features of unstable plaques: surface ulceration and plaque rupture (48% of symptomatic compared with 31% of asymptomatic, P<0.001), thinning of the fibrous cap, and infiltration of the cap by greater numbers of macrophages and T cells. In vitro studies suggest that macrophages and T cells release cytokines and proteinase, which stimulate breakdown of cap collagen and smooth muscle cell apoptosis and thereby promote plaque rupture. CONCLUSIONS: Infiltration of inflammatory cells to the surface of carotid plaques may be a critical step in promoting plaque rupture and resultant embolization or carotid occlusion. Further understanding of cell recruitment and behavior in carotid atherosclerosis may allow better detection of unstable plaques and therapeutic methods of plaque stabilization. PMID- 10700519 TI - Stiffness of carotid artery wall material and blood pressure in humans: application to antihypertensive therapy and stroke prevention. AB - BACKGROUND AND PURPOSE: Because epidemiological studies show that increased pulse pressure and carotid wall-material stiffness are predictors of cardiovascular mortality independent of age, atherosclerosis, and conventional risk factors, the relationships between carotid wall stiffness and blood pressure are important to the optimization of cardiovascular prevention. SUMMARY OF REVIEW: In middle-aged hypertensive patients, mean and pulse pressures are increased, and systolic and diastolic pressures are increased to the same degree as mean pressure. Carotid hypertrophy is associated with normal wall stress, but no increased stiffness of wall material has been reported. With age, the normal wall stress is associated with a larger diameter and a stiffer material of carotid but not peripheral arteries. The stiffer wall involves calcifications, large amounts of collagen, and fragmentation and rupture of elastic tissue, which results in increased pulse wave velocity and alterations of amplitude and timing of wave reflections and thus causes a disproportionate increase in systolic and pulse pressure. During this period, acutely administered nitrates in elderly subjects are able to reduce selectively systolic and pulse pressures without altering diastolic and mean blood pressure and composition of the carotid wall. CONCLUSIONS: New therapeutic approaches acting mainly on the wall of large arteries are needed to treat hypertension in elderly patients and prevent stroke and myocardial infarction. These drugs could either selectively lower pulse pressure through changes in wave reflections (as nitrates do) or decrease arterial wall stiffness through modification of the composition of material (such as compounds that act on collagen cross-linking). PMID- 10700520 TI - A case of cerebral hemorrhage early after carotid stenting. PMID- 10700521 TI - Fatal cerebral hemorrhage after carotid stenting. PMID- 10700522 TI - Antithrombin therapy for intracerebral hemorrhage. PMID- 10700523 TI - Cerebral dynamics of autoregulation and hypoperfusion. PMID- 10700524 TI - Delayed ischemic hyperintensity of T1-weighted MRI. PMID- 10700525 TI - Extracranial cervical artery dissection. PMID- 10700526 TI - Abstracts of literature PMID- 10700527 TI - Artificial nutrition in pancreatic disease: what lessons have we learned from the literature? AB - Acute pancreatitis is a disease process that begins with an initial injury to the pancreatic acinar cell due to the erroneous premature activation and intracellular release of digestive enzymes. The local injury is amplified through the induction of a systemic inflammatory response, mediated by the generation and release of cytokines and an aggressive inflammatory cell recruitment. Failure to maintain gut integrity may exacerbate the stress response and the systemic inflammatory reaction associated with this process, worsening the overall clinical severity of the pancreatitis and contributing further to complications of organ failure and nosocomial infection. Emphasis in the clinical nutritional management of these patients has shifted from efforts to minimize stimulation of the gland, to attaining enteral access, starting tube feeds low in the gastrointestinal tract, and monitoring tolerance. While clinical guidelines help identify those patients with acute pancreatitis at greatest need for aggressive nutritional support, the proper timing to initiate feeding, the optimal composition of the enteral formula, and whether or not enteral feeding is better than no nutritional therapy is still not clear from the current literature. PMID- 10700528 TI - Fish oil emulsions: what benefits can they bring? PMID- 10700529 TI - Enteral tube feeding in the community: survey of adult patients discharged from a Dublin hospital. AB - BACKGROUND AND AIMS: No previous study has examined the state of patients on enteral tube feeding in the community in the Republic of Ireland. METHODS: Fifty adult patients discharged from a Dublin hospital on enteral tube feeding were assessed retrospectively. RESULTS: Sixty-six per cent of the sample were over 65 years of age. Patients required enteral tube feeding as a consequence of swallowing difficulties caused by stroke (46%) or cancer of the head and neck (24%). Most patients were on full nutritional support and, in total, had spent over 49 years tube feeding in the community. Geriatric stroke patients were found to have poor functional ability and nutritional assessment proved difficult to carry out on many of these patients. Problems encountered with feeding included blocked tubes (30%), infected stoma sites (16%), and logistical problems regarding feed and equipment. Nutritional follow-up was not routine in patients with poor mobility, and 55% of patients on long-term tube feeding had not been reviewed by a dietitian in over 1 year. Patients had little faith in their general practitioner's knowledge of enteral feeding. CONCLUSIONS: While patients and families appear to cope remarkably well with tube feeding in the community, more support is necessary to ensure appropriate feeding and to monitor the nutritional status of these patients. PMID- 10700530 TI - Quality of life in long-term home enteral nutrition patients. AB - BACKGROUND AND AIMS: Few data are available on the quality of life of home enteral nutrition (HEN) patients. This study was designed to assess both the quality of life of long-term HEN patients and the evolution of quality of life after initiation of HEN. METHODS: Quality of life-related parameters were analysed in 38 patients (24M, 14F) aged 56 +/- 5 years who had been on HEN for more than 2 months (mean 25 +/- 5 months). Patients or close relatives were asked to answer a subjective assessment questionnaire, and patients with normal consciousness (n+ 24) answered the self-administered SF-36 and EuroQol questionnaires. RESULTS: Since the initiation of HEN, patients had spent 1.9 +/- 0.5% of the time in the hospital, in 54% of cases because of HEN-related complications. Analysis of the generic questionnaires revealed poorer quality of life parameters in comparison to a general population, although better results were sometimes observed in younger patients (under 45 years), patients without cancer, and patients with more than one care-giver. Nevertheless, the patients' subjective assessment of the changes in their quality of life since beginning HEN was generally good, with most patients reporting improved or stable mental and physical well-being. CONCLUSIONS: Quality of life is poor in HEN patients, but subgroups of patients who score better in some quality of life dimensions can be identified. Most patients describe an improvement in their quality of life following the initiation of HEN that needs to be confirmed by a prospective study. PMID- 10700531 TI - Effects of glucocorticoids on hepatic sensitivity to insulin and glucagon in man. AB - AIMS: This study was undertaken to determine the effects of a short-term dexamethasone treatment on hepatic sensitivities to insulin and glucagon. METHODS: Eleven healthy subjects were studied during one or several of four protocols. In all protocols, somatostatin was infused continuously to inhibit pancreatic hormone secretion. In protocol 1, basal insulin was infused over 300 min while glucagon was infused at a rate of 0.5 mg/kg(-1)/min(-1)during 180 min, then at a rate of 1.5 ng/kg(-1)/min(-1)during 150 min. In protocol 2, the same experiment was performed after a 2 day treatment with 8 mg/day dexamethasone. In protocol 3, the two-step glucagon infusion was performed during insulin infusion at a rate aimed to reproduce the hyperinsulinemia observed during protocol 2. In protocol 4, continuous basal insulin and low glucagon (0.5 mg/kg(-1)/min(-1)) were infused over 330 min. RESULTS: In protocol 1, plasma glucose rose transiently by 2.0 +/- 0.3 mmol/l when the glucagon rate was increased and glucose production increased by 1.4 +/- 0.5 micromol/kg(-1)/min(-1). In protocol 2, the insulin infusion rate (1.85 +/- 0.36 nmol/kg(-1)/min(-1)) required to maintain glycemia was 3.3-fold higher than during protocol 1. Glucagon-induced stimulation of glycemia (by 1.47 +/- 0.5 mmol/l) and endogenous glucose production (by 0.8 +/- 0.3 micromol/kg(-1)/min(-1)) were blunted, but not abolished. In protocol 3, endogenous glucose production was suppressed by 75% by hyperinsulinemia and was not stimulated when the glucagon infusion rate was increased. In protocol 4, endogenous glucose production did not change significantly with time. CONCLUSION: These results indicate that high dose glucocorticoids induce a marked hepatic insulin resistance. Stimulation of glucose production by hyperglucagonemia was maintained in spite of hyperinsulinemia which can be attributed to either hepatic insulin resistance and/or increased hepatic glucagon sensitivity. PMID- 10700532 TI - Fever and sepsis during neutropenia are associated with expansion of extracellular and loss of intracellular water. AB - BACKGROUND AND AIMS: Shifts from intracellular to extracellular water are features of a catabolic reaction to sepsis. Bedside assessment of fluid shifts was carried out in neutropenic patients at high risk of systemic infection. METHODS: Multifrequency bioelectrical impedance analysis was performed in 41 patients with leukemia or high-malignant lymphoma and chemotherapy-induced neutropenia. RESULTS: Hydration was stable during afebrile periods except for transient intra- and extracellular dehydration after chemotherapy. The risk of over-hydration and dehydration increased 3-fold during fever. Over-hydration was more severe when occurring during fever. Extracellular water was highly variable and tended to increase, and intracellular water was slowly depleted. During sepsis, these alterations were enhanced. Changes in hydration status did not predict subsequent progression to sepsis because it developed more slowly than other symptoms of infection. CONCLUSIONS: Extracellular over-hydration and intracellular dehydration are observed in febrile infection in neutropenia, similar to severe sepsis. If the technical limits of bioelectrical impedance are taken into account, this method may be useful for non-invasive monitoring of these features of metabolic stress. PMID- 10700533 TI - Antioxidant and immune status in active Crohn's disease. A possible relationship. AB - BACKGROUND AND AIMS: As reactive oxygen has been demonstrated to participate in immune genes transcription, the aim of this study was to examine the relationship between systemic concentrations of several antioxidants and markers of inflammatory and immune activation in patients with Crohn's disease (CD). METHODS: In 26 CD patients and 15 controls we compared plasma selenium and zinc concentrations, erythrocyte glutathione peroxidase (GSHPx) and superoxide dismutase activities, as well as erythrocyte sedimentation rate (ESR), C-reactive protein, tumor necrosis factor-alpha, interleukin-6, blood neopterin and soluble receptors of interleukin-2 (sIL-2R), and examined the link between these parameters. RESULTS: Selenium concentration and GSHPx activity were decreased in CD patients (54.5 +/- 3.2 vs 79 ± 2.2 microg/l, P<< 0.05; 28 +/- 1.6 vs 38 +/- 2.6 IU/g Hb, P<< 0.05) and positively correlated to each other's (r= 0.59, P<< 0.01). TNF-alpha was significantly increased in patients (18 +/- 2.6 vs 5 +/- 0.6 pg/ml;P<< 0.001), negatively correlated to GSHPx activity (r= -0.56, P<< 0.05) and selenium concentration (r= -0.72, P<< 0.001), and positively to neopterin and sIL-2R concentrations. Selenium showed negative correlation with sIL-2R (r= -0.83, P<< 0.0001) and ESR. CONCLUSIONS: In CD patients low selenium concentration may participate in reduced GSHPx activity facilitating inflammatory and immune activation. In these patients, selenium monitoring and, if needed, supplementation may be of therapeutical interest. PMID- 10700534 TI - In vivo effects of olive oil-based lipid emulsion on lymphocyte activation in rats. AB - Numerous studies suggest that immune function may be compromised by lipid emulsions rich in polyunsaturated fatty acids, especially linoleic acid. In our study, we compared the effect of a new olive oil-based lipid emulsion (ClinOleic(R)) containing 18% linoleic acid, and an emulsion based on soybean oil (Ivelip(R); 52% linoleic acid) on lymphocyte functions. Weaning Wistar rats (n= 24) were fed for 4 weeks on an oral diet that contained 12% of total energy as lipids from soybean oil. Then they received, during 6 days, a total parenteral nutrition (260 kcal/kg/d) in which 12% of total energy was brought by one of the two lipid emulsions. The fatty acid profile of spleen lymphocyte phospholipids reflected lipid intakes, with a higher content of oleic acid in ClinOleic(R) group and linoleic acid in Ivelip(R) group. A greater proportion of cells expressed the interleukin-2 receptor a-chain (CD25) after administration of ClinOleic(R) when compared to Ivelip(R) (55.43 +/- 3.47 vs 45.48 +/- 3.26%, P<< 0.05). Moreover, the CD25 expression was positively correlated with oleic acid content of spleen lymphocyte phospholipids (r= 0.500, P<< 0.018). These results show that ClinOleic(R) is able to induce, in vivo, a greater proportion of cells expressing CD25, and suggest that oleic acid could have a role in the observed effects. PMID- 10700535 TI - Food intake of a typical Brazilian diet among hospitalized malnourished patients. AB - AIM: To verify whether malnourished inpatients receiving a typical Brazilian diet meet their food requirements. METHODS: Thirty-five consecutive surgical and medical hospitalized adults, able to feed themselves, received rice and beans based diets for 3 consecutive days. All served food was weighed before and after the meals. Nutrient intake was determined and results compared to American Recommended Dietary Allowances (RDA). Malnutrition was defined by the presence of at least one of these criteria: body mass index &lE 18.5 kg/m(2); height creatinine index << 70%; or albumin level << 3. 5 g/dl. RESULTS: Malnourished and non-malnourished patients were paired in relation to age, gender, diagnoses and clinical parameters. Despite showing distinct anthropometric parameters and laboratory data, malnourished patients ingested enough quantities of food and met or exceeded RDA for energy and other nutrients. CONCLUSIONS: Clinically-stable malnourished inpatients, supplied with rice- and beans based diets have adequate energy and nutrient intake, the same occurring for non-malnourished ones. PMID- 10700536 TI - Influence of surgical stress and parenteral nutrition on serum leptin concentration. AB - Experimental studies suggest that leptin may be an important metabolic signal for energy regulation. AIM: To assess whether surgical stress produces changes in serum leptin concentration and to investigate and compare the effect of total parenteral nutrition and hypocaloric parenteral nutrition on serum leptin levels. PATIENTS AND METHODS: Twenty-two surgical patients (11 male and 11 female) in need of parenteral nutrition were recruited. Parenteral nutrition was always initiated 24 h after surgical procedure. Group I (n=15) received total parenteral nutrition, while Group II (n=7) were treated with hypocaloric parenteral nutrition. Serum leptin concentration was determined before surgical procedure (day -1), after surgery and before parenteral nutrition was started (day +1), and after 5 days of treatment with parenteral nutrition (day +6). RESULTS: A tendency to increase serum leptin levels was observed after surgical procedure (6.0+/-1.9 vs 9.9+/-2.7 ng/ml;P= 0.07). After starting parenteral nutrition no significant changes on serum leptin concentrations were found in both groups, but a trend to raise serum leptin was observed in Group I (6.2+/-1.7 vs 8.3+/-2.7 ng/ml) whereas a trend to decrease serum leptin was detected in Group II (4.6+/-2.5 vs 1.6+/-0.5 ng/ml). On day +6 an increase of serum leptin and insulin levels was observed in Group I in comparison with Group II (8.3+/-2.7 vs 1.6+/-0.5 ng/ml;P<< 0.05 and 58+/-41 vs 12+/-15 microU/l;P<< 0.05 respectively). Finally, a positive correlation at day +6 between insulin and serum leptin levels was observed (r= 0.66;P<< 0.01). CONCLUSIONS: a) Surgical stress is associated to an increase of serum leptin concentrations; b) Total and hypocaloric parenteral nutrition produces quite different effects on serum leptin levels that could be related to distinct insulin response. PMID- 10700537 TI - Severe hypothyroidism in patients dependent on prolonged thyroxine infusion through a jejunostomy. AB - BACKGROUND AND AIMS: Enteral absorption of thyroxine (T4) is variable; the duodenum and jejunum appear to be the most important sites of absorption. Our objective is to demonstrate that T4 infused via a standard jejunostomy may occasionally be poorly absorbed. METHODS: Two patients underwent esophagolaryngeal resection for carcinoma of the cervical esophagus. The procedure was accompanied by complete removal of the thyroid and parathyroid glands. A neck fistula at the gastropharyngeal anastomosis led to a restriction of oral intake; daily requirements of T4 and nutrients were given via the jejunostomy. T4 plasma levels deteriorated and thyroid-stimulating hormone (TSH) levels increased and in the third postoperative week, T4 (300 microg) was administered via a nasogastric tube. RESULTS: Although given a high dose (300 microg) of T4, both patients developed severe hypothyroidism. Infusion of T4 through the nasogastric tube precipitated the normalization of T4 and TSH plasma levels. Both patients (cases 1 and 2) resumed oral intake during the fifth and sixth postoperative weeks respectively. CONCLUSION: T4 malabsorption may occur in patients dependent on prolonged T4 infusion via a standard jejunostomy. PMID- 10700538 TI - Very severe spinal muscular atrophy (SMA type 0): an expanding clinical phenotype. AB - The classical form of severe spinal muscular atrophy (SMA type 1; Werdnig Hoffmann disease) has a very consistent clinical phenotype that is well recognized by paediatricians. It usually presents at birth or within the first few months of life. There is general hypotonia, with axial and limb weakness; the legs are affected more than the arms and proximal muscles more than distal, leaving residual spontaneous activity in the feet and in the forearms and hands. Facial muscles are spared so that the infant usually has a bright normal expression. The intercostal muscles are always affected, whereas the diaphragm is spared, allowing adequate spontaneous respiratory activity until the infants are precipitated into respiratory failure by an incidental respiratory infection, or aspiration. With rare exception they die by 2 years of age with a median around 7 months and with about 80% of the children dying by the time they are 1 year old. There is a consistent homozygous deletion in exons 7 and 8 of the telomeric copy of the survival motor neuron (SMN) gene. In the current issue of the journal, MacLeod and her colleagues have documented five cases of more severe spinal muscular atrophy, with a history of diminished fetal movements in utero and presenting at birth with asphyxia and severe weakness. PMID- 10700539 TI - The limb-girdle muscular dystrophies: diagnostic guidelines. AB - At least 11 different disorders can be recognized to be genetically distinct within the group of muscle diseases known as the limb-girdle muscular dystrophies. Direct gene or protein based tests are available to confirm the diagnosis in one autosomal dominant and six autosomal recessive forms. In these disorders, therefore, a definition based on molecular pathology is becoming possible. Clinical studies in the genetically defined subgroups may also help to determine phenotypic correlates for the various diseases. An integrated approach to diagnosis in this group, based on clinical observations supported by the result of genetic and protein studies, is likely to provide the optimum level of information. PMID- 10700540 TI - Subependymal nodular heterotopia in patients with encephalocele. AB - Only incidental mention has been made to date of the combined occurrence of subependymal heterotopia and posterior encephalocele. We evaluated the presence of disseminated nodular subependymal heterotopia in two series of patients with posterior encephalocele. The first series consisted of all six patients who were treated in our hospital for encephalocele during the last 11 years and who underwent magnetic resonance imaging (MRI). In three, subependymal nodular heterotopia was found by MRI. The second series consisted of eight autopsy cases with encephalocele, representing all cases of encephalocele that came to autopsy during a 10-year period on whom full microscopic examination could be performed. Nodular heterotopia was found in four. The combined occurrence of these two rare conditions may not be accidental. PMID- 10700541 TI - Prenatal onset spinal muscular atrophy. AB - Five patients with severe spinal muscular atrophy (SMA) type I, all of whom presented with reduced fetal movements in utero, severe weakness at birth, and short survival time were assessed to attempt to determine whether their phenotype could be explained by their genotype. The diagnosis was confirmed by clinical, electrophysiological and histopathological features. Polymerase chain reaction assays were used to define the molecular diagnosis. A gene-dosage assay was used to assess the quantity of centromeric survival motor neuron gene (SMNc) present. In all cases the telomeric survival motor neuron gene (SMNt) was absent. The SMNc gene was present but in reduced copy number compared with a control group of children with less severe type I SMA, so may be important in determining severity. In the differential diagnosis of reduced fetal movements, SMA should be considered. The clinical classification may in future be clarified by molecular genetic findings. PMID- 10700542 TI - Home and hospital treatment of acute seizures in children with nasal midazolam. AB - Rectal diazepam is widely used in the treatment of acute seizures in children but has some disadvantages. Nasal/sublingual midazolam administration has been recently investigated for this purpose but never at home or in a general paediatric hospital. The aim of this open study was to determine the efficacy, the tolerance and the applicability of nasal midazolam during acute seizures in children both in hospital and at home. We included known epileptic children for treatment at home and all children with acute seizures in the hospital. In all, 26 children were enrolled, 11 at home and 17 in the hospital (including two treated in both locations); only one had simple febrile seizure. They had a total of 125 seizures; 122 seizures (98%) stopped within 10 minutes (average 3.6 minutes). Two patients in the hospital did not respond and in three, seizures recurred within 3 hours. None had serious adverse effects. Parents had no difficulties administering the drug at home. Most of those who were using rectal diazepam found that nasal midazolam was easier to use and that postictal recovery was faster. Among 15 children who received the drug under electroencephalogram monitoring (six without clinical seizures), the paroxysmal activity disappeared in ten and decreased in three. Nasal midazolam is efficient in the treatment of acute seizures. It appears to be safe and most useful outside the hospital in severe epilepsies, particularly in older children because it is easy for parents to use. These data should be confirmed in a larger sample of children. Its usefulness in febrile convulsions also remains to be evaluated. PMID- 10700543 TI - The maturation of auditory cortical evoked responses between (preterm) birth and 14 years of age. AB - In this study we report on the maturation of the auditory cortical evoked response (ACR) waveform between (preterm) birth and 14 years of age. From the results it can be concluded that the ACR waveform morphology shows substantial age-dependent changes until the age of 14 years. Two transitional periods could be recognized. The first between 36 and 41 weeks conceptional age; the second between 4 and 6 years of age. The adult waveform complex is achieved between 14 and 16 years of age. Further research is needed to determine whether these transitional periods in the maturation of ACRs correspond with important functional changes of the central auditory system. PMID- 10700544 TI - Van der Knaap's vacuolating leukoencephalopathy: two additional cases. AB - We present two new cases with infantile-onset megalencephaly and a characteristic magnetic resonance imaging (MRI) pattern including severe white-matter abnormalities and subcortical cysts. In one of the patients MRI at the early age of 9 months showed pronounced white matter swelling. In another patient the swelling of white matter was less pronounced at 12 years of age. PMID- 10700545 TI - The limb-girdle muscular dystrophies [directory]. PMID- 10700546 TI - Brain MRI detects fictional character. PMID- 10700547 TI - Distribution of delta opioid receptor immunoreactivity in the hamster suprachiasmatic nucleus and intergeniculate leaflet. AB - The hamster suprachiasmatic nucleus (SCN) is innervated by a dense plexus of enkephalin-containing axons originating from cells in the intergeniculate leaflet (IGL) of the thalamus. However, the distribution of opioid receptors within the hamster SCN has not been reported. Opioid receptors consist of three primary subtypes: mu, delta and kappa opioid receptors. Enkephalins have the highest affinity for delta opioid receptors. Therefore, in the present study, we examined the distribution of delta opioid receptor immunoreactivity in the hamster SCN and the IGL of the thalamus. Coronal sections of the hamster hypothalamus inclusive of the SCN or thalamic regions containing the IGL were prepared at specific times of the day and labeled with anti-delta opioid receptor polyclonal antisera using standard immunohistochemical techniques. delta opioid receptors were heavily distributed within rostral-caudal regions of the SCN, with the densest labeling located in the ventral and medial regions of the mid-SCN. Similar patterns of labeling were observed for tissue prepared during mid-day or mid-night times. In contrast, delta opioid receptor immunoreactivity only sparsely labeled cells in the IGL. Cellular staining in all regions appeared as dark punctate labeling surrounding cells, indicative of terminal boutons. Therefore, it is suggested that delta opioid receptors are located presynaptically on axon terminals within the hamster SCN and IGL. These results suggest that delta opioid receptors may play a role in modulating circadian rhythms generated within the SCN, possibly by regulating transmitter release within the nucleus. PMID- 10700548 TI - Effects of sleep deprivation on sleep and sleep EEG in three mouse strains: empirical data and simulations. AB - Gene targeted mice can be used as models to investigate the mechanisms underlying sleep regulation. Three commonly used background strains for gene targeting (129/Ola, 129/SvJ and C57BL/6J) were subjected to 4-h and 6-h sleep deprivation (SD), and their sleep and sleep EEG were continuously recorded. The two-process model of sleep regulation has predicted the time course of slow-wave activity (SWA) in nonREM sleep after several sleep-wake manipulations in humans and the rat [3] [9]. We tested the capacity of the model to predict SWA in nonREM sleep on the basis of the temporal organization of sleep in mice. The strains differed in the amount and distribution of sleep and the time course of SWA. After spontaneous waking episodes of 10-30 min as well as after SD, SWA was invariably increased. Simulations of the time course of SWA were successful for 129/SvJ and C57BL/6J, but were not satisfactory for 129/Ola. Since the time constants are assumed to reflect the dynamics of the physiological processes involved in sleep regulation, the results provide a basis for the use of gene targeted mice to investigate the underlying mechanisms. PMID- 10700549 TI - Oxidative stress, mitochondrial permeability transition and activation of caspases in calcium ionophore A23187-induced death of cultured striatal neurons. AB - Disruption of intracellular calcium homeostasis is thought to play a role in neurodegenerative disorders such as Huntington's disease (HD). To study different aspects of putative pathogenic mechanisms in HD, we aimed to establish an in vitro model of calcium-induced toxicity in striatal neurons. The calcium ionophore A23187 induced a concentration- and time-dependent cell death in cultures of embryonic striatal neurons, causing both apoptosis and necrosis. Cell death was significantly reduced by the cell-permeant antioxidant manganese(III)tetrakis(4-benzoic acid) porphyrin (MnTBAP). Cyclosporin A and its analogue N-MeVal-4-cyclosporin also reduced the incidence of cell death, suggesting the participation of mitochondrial permeability transition in this process. Furthermore, addition of either of two types of caspase inhibitors, Ac YVAD-CHO (acetyl-Tyr-Val-Ala-Asp-aldehyde) and Ac-DEVD-CHO (acetyl-Asp-Glu-Val Asp-aldehyde), to the striatal cells blocked A23187-induced striatal cell death in a concentration-dependent manner. These results suggest that oxidative stress, opening of the mitochondrial permeability transition pore and activation of caspases are important steps in A23187-induced cell death. PMID- 10700550 TI - Brainstem and hypothalamic areas involved in respiratory chemoreflexes: a Fos study in adult rats. AB - The adaptation to hypoxia and hypercapnia requires the activation of several anatomical structures along the neuraxis. In this study, using Fos immunoreactivity, we sought to map neuronal populations involved in chemoreflex networks activated during the responses to moderate hypoxia (O(2) 11%), and hypercapnia (CO(2) 5%) in the brainstem and the hypothalamus of the rat. In the medulla, hypoxia elicited marked and significant staining in the nucleus of the solitary tract (NTS), and in parapyramidal neurons located near the ventral surface, whereas hypercapnia evoked significantly c-fos only near the ventral surface in paraolivar neurons. In contrast, within pontine and suprapontine structures, both hypoxia and hypercapnia evoked similarly Fos immunoreactivity in the lateral parabrachialis area, the central grey, the caudal hypothalamus (dorsomedial and posterior hypothalamic nuclei), and in a ventro-lateral hypothalamic area, extending from the rostral limit of the mammillary nuclei to the retrochiasmatic area. More rostrally, labelling was observed in the paraventricular nucleus of the hypothalamus in response to hypercapnia, and in the supraoptic nucleus in response to hypoxia. These results support the hypothesis that chemoreflexes pathways are not only restricted to medulla and pons but also involved mesencephalic and hypothalamic regions. The parabrachialis area and the central grey may be key relays between caudal and ventral hypothalamic neurons, and medullary neurons involved in the response to hypoxia and hypercapnia. PMID- 10700551 TI - Spatial and temporal patterns of transneuronal labeling in CNS neurons after injection of pseudorabies virus into the sciatic nerve of adult rats. AB - The distribution of labeled neurons in the brain and spinal cord was studied after injecting the Bartha strain of pseudorabies virus (PRV) into the sciatic nerve to provide a baseline for studying neural circuitry after spinal cord injury (SCI) and regeneration. Following a single injection of viral particles into the left sciatic nerve, PRV labeling was found in the spinal cord at 2 days post-injection (p.i.). Increasing complexity in viral labeling from the spinal cord to supraspinal regions became apparent with increasing survival time. In brain regions, several neuronal groups that regulate sympathetic outflow, such as the rostroventrolateral medulla, the lateral paragigantocellular nuclei, and the A5 cells, were densely labeled. However, relatively sparse labeling was noticed in the lateral vestibular nuclei, the red nucleus and the motor cortex whose spinal projections regulate somatic motor function, although those areas were abundantly labeled with Fast blue (FB) in a double-labeling experiment in which FB was co-injected into the lumbar cord. The pattern of viral labeling became more complex beyond 5 days p.i. when increased numbers of cell groups were labeled with PRV but not FB. In addition, some infected neurons started to lyse, as evidenced by a decrease in viral labeling at 7 days p.i. Thus, the 5th day post-viral injection would appear to be an appropriate survival time to obtain maximal labeling with acceptable specificity. We suggest that transneuronal labeling using PRV should be appropriate for studying multi-neural circuitry after SCI and regeneration. PMID- 10700553 TI - Acquisition, extinction, and reinstatement of Pavlovian fear conditioning: the roles of the NMDA receptor and nitric oxide. AB - The acquisition and extinction of Pavlovian conditioned fear have been shown to be mediated by the N-methyl-D-aspartate (NMDA) glutamate receptor. This study found that the NMDA antagonist (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclo hepten-5,10-imine maleate (MK-801) blocked the reinstatement of Pavlovian conditioned fear in rats. The role of nitric oxide (NO) in the acquisition and extinction of Pavlovian fear conditioning was also examined. L-NAME, an NO synthase inhibitor, failed to block the acquisition or extinction of Pavlovian fear conditioning. The results are discussed in the context of hierarchical associations and the array of NMDA and calcium mediated mechanisms of synaptic strengthening. PMID- 10700552 TI - Cannabinoid CB(1) receptors colocalize with tyrosine hydroxylase in cultured fetal mesencephalic neurons and their activation increases the levels of this enzyme. AB - The incubation of cultured fetal mesencephalic neurons with Delta(9) tetrahydrocannabinol (Delta(9)-THC) increased the activity of tyrosine hydroxylase (TH) and this increase was reversed by SR141716A, a specific antagonist for cannabinoid CB(1) receptors. In the present work, we extended these earlier observations by addressing two objectives. First, we characterized at a molecular level the presence of CB(1) receptors in cultured fetal mesencephalic neurons using two strategies: (i) analyzing the presence of CB(1) receptor gene transcripts by Northern blot, and (ii) measuring [3H]WIN-55,212-2 binding in membrane fractions obtained from these cells, as well as evaluating the potential increase in [35S]-guanylyl-5'-O-(gamma-thio)-triphosphate ([35S]GTPgammaS) binding caused by the activation of these receptors with WIN 55,212-2, a synthetic agonist. Northern blot analyses demonstrated the presence of small, but measurable levels of CB(1) receptor mRNA in cultured fetal mesencephalic neurons. The presence of these transcripts was accompanied by the presence of receptor binding protein, as revealed by a small, but specific, [3H]WIN-55, 212-2 binding in membrane fractions obtained from these cells. These CB(1) receptors are coupled to GTP-binding proteins, as the incubation of membrane fractions obtained from these cells with WIN-55,212-2 slightly, but significantly increased [35S]GTPgammaS binding. This fact indicated the existence, not only of receptor binding, but also of a functional receptor transduction pathway. As a second objective, we examined the potential colocalization of CB(1) receptors and TH in these cells by double-labelling immunocytochemistry. We also determined by Western blotting whether the previously observed Delta(9)-THC-induced increase in TH activity was accompanied by increased TH protein levels. Cultured fetal mesencephalic neurons exhibit diverse cell phenotypes, with CB(1) receptors localized only on TH-containing neurons. In addition, we found that the incubation of fetal mesencephalic neurons with medium containing Delta(9)-THC increased TH protein levels, in concordance with the previously reported increase in TH activity. Collectively, our results support the notion that CB(1) receptors are present in cultured fetal mesencephalic TH-containing neurons, despite their absence in the corresponding neurons in the adult brain. Thus, it is likely that the effects of cannabinoids on TH activity are direct. All this data strengthen the view that cannabinoid receptors are atypically located during brain development and that they might play an important role during this process, in particular on the phenotypical expression of TH-containing neurons. PMID- 10700554 TI - Diphosphorylation and involvement of extracellular signal-regulated kinases (ERK1/2) in glutamate-induced apoptotic-like death in cultured rat cortical neurons. AB - Glutamate-induced excitotoxicity, with certain characteristics of apoptosis, has been implicated in a variety of neuronal degenerative disorders. In some physiological cases, extracellular signal-regulated kinases (ERK1/2) are activated by stimulation of glutamate receptors. In the present study, the activation (diphosphorylation) and role of ERK1/2 in glutamate-induced apoptotic like death in cultured cortical neurons were investigated. Protein levels and activation (diphosphorylation) levels of ERK1/2 were examined by Western immunoblot, probed with anti-ERK1/2 and anti-active (diphosphorylated) ERK1/2 antibodies, respectively. Apoptotic-like death was determined by DAPI staining. Before a remarkable increase of apoptotic-like cell death was observed at 9-18 h after 15 min exposure to 50 microM glutamate, diphosphorylation levels of ERK1/2 were rapidly increased, peaked at 5-15 min of the exposure, and reverted to sham control level 3 h after the exposure, while the protein levels of ERK1/2 were unaffected. The glutamate concentration effective for inducing apoptotic-like cell death was correlated with that for inducing ERK1/2 diphosphorylation. Both ERK1/2 diphosphorylation and the apoptotic-like cell death were largely prevented by MK-801, a specific NMDA receptor (a subtype receptor of glutamate) antagonist, or the elimination of extracellular Ca(2+) with EGTA. PD98059, a specific inhibitor of ERK1/2 kinase, completely inhibited ERK1/2 diphosphorylation and partially inhibited the apoptotic-like cell death. These results suggest that largely via NMDA receptor-mediated influx of extracellular Ca(2+), ERK1/2 were rapidly and transiently activated and were involved in glutamate-induced apoptotic-like death in cultured rat cortical neurons. PMID- 10700555 TI - Dephosphorylation-induced decrease of anti-apoptotic function of Bcl-2 in neuronally differentiated P19 cells following ischemic insults. AB - It is known that Bcl-2 has a protective effect against neuronal ischemia. Some reports speculate anti-apoptotic function of Bcl-2 depends not on the expression level but on the phosphorylation state. We found induction of apoptosis and CPP32 activation by energy impairment (3-nitropropionic acid (3-NP)-treatment or glucose-deprivation) in the neuronally differentiated P19 cells. Time course study of cell viability following ischemic insults showed that the number of viable cells decreased along with the increase in the amount of dephosphorylated Bcl-2 without obvious quantitative alteration of the protein. Then, we generated differentiated P19 cells overexpressing wild-type Bcl-2 (P19/wt. Bcl-2) or phosphorylation-negative Bcl-2 mutant (P19/mut.Bcl-2), in which alanine was substituted for serine 70. When the cell viability was examined within 24 h, P19/mut.Bcl-2 was more vulnerable to energy impairment as compared with P19/wt.Bcl-2. In addition, overexpression of wild-type Bcl-2 inhibited DNA laddering and CPP32 activation induced by the insults, while that of mutant Bcl-2 did not. These findings suggest that the phosphorylation state, as well as the expression level, of Bcl-2 plays an important role to modulate its protective effect against ischemic insults. PMID- 10700556 TI - Developmental neurotoxicity of chlorpyrifos in vivo and in vitro: effects on nuclear transcription factors involved in cell replication and differentiation. AB - Chlorpyrifos is a widely used organophosphate insecticide that is a suspected developmental neurotoxin. Although chlorpyrifos exerts some effects through cholinesterase inhibition, recent studies suggest additional, direct actions on developing cells. We assessed the effects of chlorpyrifos on nuclear transcription factors involved in cell replication and differentiation using in vitro and in vivo models. HeLa nuclear protein extracts were incubated with the labeled consensus oligonucleotides for AP-1 and Sp1 transcription factors in the presence and absence of chlorpyrifos. In concentrations previously shown to affect cell development, chlorpyrifos reduced AP-1, but not Sp1 DNA-binding activity. Next, chlorpyrifos was incubated with PC12 cells either during cell replication or after initiation of differentiation with NGF. Chlorpyrifos evoked stage-specific interference with the expression of the transcription factors: Sp1 was reduced in replicating and differentiating cells, whereas AP-1 was affected only during differentiation. Finally, neonatal rats were given apparently subtoxic doses of chlorpyrifos either on postnatal days 1-4 or 11-14 and the effects were evaluated in the forebrain (an early-developing, cholinergic target region) and cerebellum (late-developing region, poor in cholinergic innervation). Again, chlorpyrifos evoked stage-specific changes in transcription factor expression and binding activity, with greater effects on Sp1 during active neurogenesis, and effects on AP-1 during differentiation. The changes were present in both forebrain and cerebellum and were gender-specific. These results indicate that chlorpyrifos interferes with brain development, in part by multiple alterations in the activity of transcription factors involved in the basic machinery of cell replication and differentiation. Noncholinergic actions of chlorpyrifos that are unique to brain development reinforce the need to examine endpoints other than cholinesterase inhibition. PMID- 10700557 TI - Different patterns of respiratory and cardiovascular responses elicited by chemical stimulation of dorsal medulla in the rat. AB - Respiratory and cardiovascular responses to microinjections (10 nl) of L glutamate (10 mM) into the dorsal medulla were studied in spontaneously breathing urethane-anesthetized, adult male Wistar rats. A total of 10 patterns of respiratory and cardiovascular responses were observed: (1) hypotension alone; (2) hypotension and bradycardia; (3) hypotension and apnea; (4) hypotension, bradycardia, and apnea; (5) apnea alone; (6) hypotension and fast and shallow breathing; (7) hypotension, bradycardia, and fast and shallow breathing; (8) fast and shallow breathing alone; (9) sighs; and (10) increase in BP and HR accompanied with fast and shallow breathing. The sites from which a combination of hypotension, bradycardia, and apnea was elicited, occupied a region in the medial subnucleus of nucleus tractus solitarius (nTS), the reticular formation just ventral to it, and the dorsal motor nucleus of vagus. The sites from which hypotension alone or a combination of hypotension and apnea were elicited occupied the margins of the medial subnucleus of nTS. The sites from which apnea alone was elicited were located in the ventrolateral part of nTS and the reticular formation just ventral to it. In the commissural subnucleus of nTS, the responses comparable to those elicited by peripheral chemoreceptor stimulation (i.e., increase in BP, HR, and respiratory rate) were located in a midline region just caudal to the calamus scriptorius, the sites from which sighs were elicited were located slightly lateral and deeper, the sites from which fast and shallow breathing were elicited were located in the dorsal portion, slightly lateral to the midline. These results are expected to prove useful in studies in which microinjection technique is used to identify transmitters/receptors involved in mediating respiratory and cardiovascular reflex responses. PMID- 10700558 TI - Effects of prolactin on expression of Fos-related antigens in tyrosine hydroxylase-immunoreactive neurons in subdivisions of the arcuate nucleus. AB - Dual immunohistochemistry was employed to determine the effects of prolactin on expression of Fos and its related antigens (FRA) in tuberoinfundibular dopamine (TIDA) neurons located in the dorsomedial (DM) and ventrolateral (VL) subdivisions of the arcuate nucleus (ARC) in the male rat. Systemic administration of the DA receptor antagonist haloperidol caused a sustained (up to 12 h) increase in plasma prolactin concentrations that was accompanied by a transient increase (at 3 h) in the percentage of tyrosine hydroxylase (TH) immunoreactive (IR) neurons containing FRA-IR nuclei in the DM-ARC. In contrast, haloperidol caused a prolonged (1. 5 to 12 h) decrease in the percentage of TH-IR neurons with FRA-IR nuclei in the VL-ARC. Haloperidol had no effect, however, on the overall number of TH-IR neurons in either of these regions. Co-administration of prolactin antisera (PRL-AB) blocked haloperidol-induced increases in both plasma prolactin concentrations and the percentage of TH-IR neurons expressing FRA in the DM-ARC, but had no effect on haloperidol-induced inhibition of FRA expression in TH-IR neurons in the VL-ARC. Intracerebroventricular (i.c.v.) administration of prolactin also increased the percentage of TH-IR neurons containing FRA-IR nuclei in the DM-ARC, but this effect was of longer duration (up to 6 h) than that of haloperidol in all but the most caudal portion of the DM ARC. In the VL-ARC, prolactin caused a transient increase (at 1.5 h) in the percentage of TH-IR containing FRA-IR nuclei. These results demonstrate that prolactin regulates immediate early gene expression in TIDA neurons in male rats, and reveal that there are temporal differences in the responsiveness of discrete subpopulations of these neurons to prolactin. Prolactin causes a short-lived increase in FRA expression in TIDA neurons in the VL-ARC which is followed by a more prolonged activation of FRA expression in TIDA neurons in the DM-ARC. PMID- 10700559 TI - Functional brain areas used for the lifting of objects using a precision grip: a PET study. AB - Positron emission tomography (PET) was performed in 10 normal volunteers to investigate regional cortical and subcortical activation induced by the lifting of an object repetitively using a precision grip between the index finger and thumb. Data were obtained for three object weights (4, 200 and 600 g) and a resting condition. Grip and lift forces on a similar object and the activity of selected muscles in the hand, arm and shoulder were also recorded in separate lifting trials. A comparison between all movement conditions and the resting condition revealed significant activation of the primary motor (M1), primary sensory (S1), dorso-caudal premotor (PM), caudal supplementary motor (SMA) and cingulate motor (CMA) cortices contralateral to the hand used. On the ipsilateral side, activation of the M1, caudal SMA and inferior parietal cortex (BA 40) was also found. In the subcortical areas, the bilateral hemispheres and right vermis of the cerebellum, left basal ganglia and thalamus were activated. Behavioral adaptation to a heavier object weight was revealed in a nearly proportional increase of both grip and lift forces, prolonged force application period and a higher level of hand and arm muscle activities. An increase in the rCBF associated with these changes was noted in several cortical and subcortical areas. However, consistent object weight-dependent activation was observed only in the M1/S1 contralateral to the hand used. PMID- 10700560 TI - Neuron-specific localisation of the TR3 death receptor in Alzheimer's disease. AB - Death receptors are associated with the homeostatic and pathologic induction of cell death. TR3 is a recently characterised member of the death receptor family that is expressed in the adult brain. In order to establish the role of TR3 in acute CNS disease and chronic neurodegeneration, we analysed brain regions from Alzheimer's disease (AD), stroke and neurotrauma patients, using a novel anti peptide antibody generated to an exposed epitope in the extracellular domain of the receptor. We show a statistically significant increase in TR3 protein levels in AD brain samples but not in stroke, neurotrauma or control samples. The increase observed for TR3 was specific to neurons in regions associated with AD pathology. This is the first report describing the neuron-specific regulation of a death receptor in chronic disease and may indicate that a TR3 receptor-mediated signalling pathway is involved in AD-associated neuronal loss. PMID- 10700561 TI - Brain-derived neurotrophic factor requirement for activity-dependent maturation of glutamatergic synapse in developing mouse somatosensory cortex. AB - The maturation of cortical circuitry critically depends on experience. Recently, a model of silent synapse has been proposed as a mechanism of activity-mediated transition of immature synapse to mature synapse. It is not clear, however, how activity could regulate this transition. Here, we show the evidence that endogenous brain-derived neurotrophic factor (BDNF) is required for the maturation of glutamatergic synapse in developing mouse somatosensory cortex. Field potential recordings of thalamocortical glutamatergic synaptic activity with brain slices from the BDNF mutant mice showed that AMPA receptor responses are low, but NMDA receptor responses remain high in layer 4, thus, the relative contribution of AMPA receptor response is significantly lower compared to the age matched wild-type mouse. Furthermore, optical images of development of thalamocortical connectivity with a voltage-sensitive dye showed that NMDA receptor-dominant synapse is established first in layer 4 and layer 5/6 then AMPA receptor response appears later in concomitant with reduction of NMDA receptor response in layer 4 and that the maturation of the silent synapse is impaired in the BDNF mutant mice. In layer 5/6, NMDA receptor response was suppressed without upregulation of AMPA receptor response. This process also required BDNF function. Interestingly, whisker-trimming of the wild-type mouse from just after birth showed quite similar results with the homozygous mutant of their whiskers left intact. Therefore, we would propose that BDNF is a critical mediator for the maturation of glutamatergic synapse in developing mouse somatosensory cortex. PMID- 10700562 TI - CB1 cannabinoid receptor expression in brain regions associated with zebra finch song control. AB - Cannabinoids have been used for millennia through various preparations of Cannabis sativa. Despite this long history of use, the physiological significance of cannabinoid signaling in the vertebrate CNS is not well understood. High CB1 cannabinoid receptor densities in mammalian telencephalon and the results of behavioral studies suggest that cannabinoids play a role in cognitive function, learning, and memory. Since a network of discrete brain regions in zebra finch telencephalon controls song learning, we hypothesized that cannabinoid signaling may be relevant to songbird vocal development and behavior. Radioligand binding experiments using the cannabinoid agonist [3H]CP-55940 allowed identification of a dense population of high-affinity cannabinoid binding sites in zebra finch neuronal membranes. Northern blotting and RT-PCR experiments demonstrated expression of a predominant zebra finch CB1 mRNA of approximately 5.5 kb. Expression of this CB1 mRNA appears to change over the course of vocal development within the caudal telencephalon. As zebra finch caudal telencephalon contains the higher vocal center (HVC) and the robust nucleus of the archistriatum (RA), regions involved in song learning and production, we further investigated CB1 expression in these areas using in situ hybridization. In situ hybridization revealed that CB1 mRNA is expressed at high levels within both HVC and RA. Overall, these data demonstrate the presence of CB1 signaling systems within songbird telencephalon, notably within regions known to be involved in song learning and production. High-level CB1 expression in song regions suggests a potential role for cannabinoid signaling in zebra finch vocal development. PMID- 10700563 TI - Age-related impairment of coupling mechanism between neuronal activation and functional cerebral blood flow response was restored by cholinesterase inhibition: PET study with microdialysis in the awake monkey brain. AB - The effects of three cholinesterase inhibitors (physostigmine, E2020, and Tacrine), all of which are to be cognitive enhancers, on the functional regional cerebral blood flow (rCBF) response were studied in young (5.9+/-1.8 years old) and aged (18.0+/-3.3 years old) monkeys under awake conditions using high resolution positron emission tomography (PET). Under control condition, vibrotactile stimulation elicited increases in the rCBF response in the contralateral somatosensory cortices of both young and aged monkeys, but the degree of increase in rCBF response was significantly lower in aged (115.8%) than that in young monkeys (139.9%). Regional cerebral metabolic rate of glucose (rCMRglc) response to the stimulation, measured using [18F]-2-fluoro-2-deoxy Dphysostigmine) were consistent with the data obtained by microdialysis. In contrast, the cognitive enhancers did not alter rCBF response to stimulation in young monkeys. The present results demonstrated that the functional change in rCBF response to the stimulation was induced during the aging process by impairment of the coupling mechanism between the neuronal activation and rCBF response. Furthermore, the observation that cognitive enhancers partly restored the functional rCBF response suggested that the coupling mechanism might be regulated via cholinergic neuronal transmission. PMID- 10700564 TI - Comparison of the novel drug Ensaculin with MK-801 on the reduction of hydroxyl radical production in rat striatum after local application of glutamate. AB - Ensaculin interacts with various neurotransmitter systems (e.g., dopaminergic, serotoninergic, glutamatergic) and was originally designed for the treatment of dementia. In the present study Ensaculin was tested for its possible reduction of glutamate-induced hydroxyl free radical formation in vivo. The microdialysis experiment was carried out in non-anaesthetized Wistar rats, which were implanted with a microdialysis probe into the striatum. Salicylate (10 nmol/2 microl/min) was incorporated into the perfusion fluid to measure indirectly hydroxyl radicals indicated by 2,3-dihydroxybenzoic acid (DHBA) formation. After baseline recording, glutamate (100 or 500 nmol/2 microl/min) was perfused through the microdialysis probe (CMA 12, 4 mm, flow rate 2 microl/min). Ensaculin (0.1, 1 and 10 mg/kg), MK-801 (1 mg/kg) or saline was injected i.p. 20 min after the onset of glutamate perfusion (500 nmol/2 microl/min). Glutamate (100 nmol/2 microl/min) and (500 nmol/2 microl/min) perfusion produced a 2.6- and 17-fold increase of 2,3 DHBA, respectively. Treatment with Ensaculin (1 and 10 mg/kg i.p. ) significantly antagonized the formation of 2,3-DHBA, to values of 60.5% and 56.7% of control levels, respectively. In comparison, MK-801 attenuated 2,3-DHBA levels, to values of 65.8% compared to control values. Ensaculin may be useful in the treatment of neurodegenerative disorders associated with elevated hydroxyl free radicals and excitotoxicity. PMID- 10700565 TI - Free radicals are involved in the damage to protein synthesis after anoxia/aglycemia and NMDA exposure. AB - Neuronal protein synthesis is inhibited in CA1 pyramidal neurons for many hours after ischemia, hypoxia or hypoglycemia. This inhibition precedes cell death, is a hallmark characteristic of necrotic damage and may play a key role in the death of vulnerable neurons after these insults. The sequence of events leading to this inhibition remains to be fully elucidated. The protein synthesis failure after 7.5 min anoxia/aglycemia in the rat hippocampal slice can be prevented by blocking N-methyl-D-aspartate receptors in a reduced calcium environment during the insult. In this study, we demonstrate that N-methyl-D-aspartate exposure directly causes a dose-dependent, receptor-mediated and prolonged protein synthesis inhibition in CA1 pyramidal neurons. The free radical scavenger Vitamin E significantly attenuates this damage due to low concentrations of N-methyl-D aspartate (10 microM). Free radical generation by xanthine/xanthine oxidase (XOD) can directly damage protein synthesis in neurons of the slice. Vitamin E, ascorbic acid and N-acetylcysteine can each prevent the damage due to anoxia/aglycemia and to higher concentrations of N-methyl-D-aspartate (50 microM), provided calcium levels are reduced concomitantly. These findings indicate that both free radicals and calcium play a role in the sequence of events leading to protein synthesis failure after energetic stress like anoxia/aglycemia. They further suggest that the mechanism by which N-methyl-D aspartate receptor activation damages protein synthesis involves free radical generation. PMID- 10700566 TI - The effect of delta-9-tetrahydrocannabinol on forebrain ischemia in rat. AB - The purpose of the study was to evaluate the effect of delta-9 tetrahydrocannabinol (THC), the major psychoactive constituent of marijuana, on ischemic neuronal injury. A 12-min ischemic insult was induced by a reduction in systolic blood pressure to a mean of 50 mm Hg, followed by bilateral carotid artery occlusion at a middle ear temperature of 37.5 degrees C. THC at either a low (0.1 mg/kg; n=8) or high (10 mg/kg; n=8) dose was injected i.p. every 12 h for 7 days prior to ischemia. Non-treated ischemic (n=8) animals formed the control group. The animals were sacrificed 3 weeks post-ischemia for quantitative histopathology. THC at either dose did not significantly reduce ischemic neuronal damage in the hippocampus. The high dose THC-treated group showed significantly less neocortical injury, compared to either the control or low-dose THC groups (p<0.05). The striatum was markedly protected by both low and high dose THC (p<0.001). This regionally specific protection implies that either the hippocampus undergoes suprathreshold ischemic injury or that mechanisms of ischemic injury vary in different brain regions. PMID- 10700567 TI - The effect of ketamine isomers on both mice behavioral responses and c-Fos expression in the posterior cingulate and retrosplenial cortices. AB - Ketamine, a non-competitive NMDA receptor antagonist, is a racemic mixture. S(+) ketamine is presumed to be more potent as an anesthetic than R(-) ketamine, and causes less postanesthetic stimulation of locomotor activity than R(-) ketamine in animals at equihypnotic doses. In the present study, we investigated the effect of S(+), R(-), and racemic ketamines on mice behavioral responses and c Fos expression in the posterior cingulate and retrosplenial cortices (PC/RS), which are suggested to be the brain regions responsible for NMDA-receptor antagonist-induced psychotomimetic activity. Ataxia and head weaving and c-Fos expression in the PC/RS were significantly more induced by both S(+) and racemic ketamines than by R(-) ketamine at the same dose. S(+) ketamine induced significantly more potent ataxia than racemic ketamine at the same dose. Ketamine induced c-Fos expression in the PC/RS correlated well with the intensity of behavioral responses. These results imply that R(-) ketamine is weaker than both S(+) and racemic ketamines in a psychotomimetic effect. Also, S(+) ketamine is more potent than racemic ketamine in a psychotomimetic effect and possibly in an anesthetic effect. They also indicate that PC/RS is at least one of the specific brain regions responsible for ketamine-induced behavioral responses in animals and a psychotomimetic activity in humans. PMID- 10700568 TI - Distribution of tau protein kinase I/glycogen synthase kinase-3beta, phosphatases 2A and 2B, and phosphorylated tau in the developing rat brain. AB - When trying to elucidate the role played by tau protein kinase I/glycogen synthase kinase-3beta (TPKI/GSK-3beta) in tau phosphorylation, it is important to consider the balance that exists between the various kinases and phosphatases that are involved in vivo. We studied developmental changes in the expressions of TPKI/GSK-3beta and phosphatases 2A and 2B in rat brains using immunoblot analysis. The expression of the kinase peaked postnatally at days 8-11 and returned then to low level after 5 weeks. Phosphatase 2A showed a similar pattern, increasing postnatally until day 14 and decreasing thereafter. On the other hand, phosphatase 2B was undetectable at the juvenile stage, but later its presence increased rapidly to peak at 5 weeks after birth, after which it was maintained at high levels throughout the adult stage. Immunohistochemical studies using the PAP method revealed details of the distribution of TPKI/GSK-3beta. At postnatal days 3-21 both gray and white matter were immunoreactive. Later, after 5 weeks, the immunoreactivity became more restricted to the gray matter. The staining of tau phosphorylated at Ser 199, Ser 396, and Ser 413 followed mostly the pattern of the kinase distribution throughout all stages of development. These data, therefore, confirm that TPKI/GSK-3beta is expressed primarily in neurons and especially in neurites until postnatal day 21, whereafter the distribution is concentrated mostly in the cell soma and the proximal neurite region. PMID- 10700569 TI - Brain blood flow during hypercapnia in fish: no role of nitric oxide. AB - Very little is known about the regulation of cerebral blood flow (CBF) in lower vertebrates, especially fish. In mammals, hypercapnia causes cerebral vasodilation and increased CBF through mechanisms that involve the production of nitric oxide (NO). We have used epi-illumination microscopy in vivo to observe effects of hypercapnia on venular erythrocyte velocity, used as an index of CBF velocity, in rainbow trout (Oncorhynchus mykiss) and crucian carp (Carassius carassius). Rainbow trout exposed to a pCO(2) of 7.5 mmHg displayed a small increase of CBF velocity in two out of five fishes, while dorsal aortic blood pressure (P(DA)) did not change. Exposing trout to a pCO(2) of 22.5 mmHg, resulted in an 80% increase in CBF velocity and a 21% increase in P(DA). Trout exposed to a pCO(2) of 75 mmHg showed an additional increase in blood pressure, while no further increase was seen in CBF velocity compared to a pCO(2) of 22. 5 mmHg. By contrast, no change in CBF velocity was seen in crucian carp, even at a pCO(2) of 75 mmHg. None of the circulatory changes seen in the trout could be blocked by superfusing the brain surface with the NO synthase blocker N(G)-nitro L-arginine. The results point at striking species differences in the responses of CBF and P(DA) to hypercapnia in fish, and that the hypercapnia induced increase in CBF velocity seen in rainbow trout is independent of NO production. PMID- 10700570 TI - Central changes in nociceptin dynorphin B and Met-enkephalin-Arg-Phe in different models of nociception. AB - The newly identified neuropeptide nociceptin/orphanin FQ (NOC) was measured in different rat brain areas related to the descending anti-nociceptive pathways and compared to two opioid peptides, dynorphin B (DYN B) and Met-enkephalinArgPhe (MEAP). Two experimental models of chronic nociception, one neurogenic and one inflammatory, used in this study, reveal how different pathological conditions may influence these endogenous systems. Nerve injury is induced by ligation of the sciatic nerve and inflammation by a carrageenan injection in the gluteal muscle, 2 weeks prior to decapitation. Selected brain areas were dissected out and frozen. NOC-, DYN B- and MEAP-like immunoreactivity (LI) is determined by radioimmunoassay. Nerve injury increased the NOC-LI levels in the cortex cinguli, DYN B-LI levels in the dorsal and the ventral part of the spinal cord, whereas a decrease in the MEAP-LI levels is seen in the dorsal part of the periaqueductal grey (PAG). After inflammation, the NOC-LI levels increased in cortex cinguli, hypothalamus and in the dorsal spinal cord, whereas DYN B-LI levels increased in the dorsal part of the PAG. A general increase in MEAP-LI levels is found after inflammation in all analyzed brain areas except in hippocampus. In conclusion, increased levels of NOC-LI were found in cortex cinguli in both treatment groups and in hypothalamus and spinal cord following carrageenan treatment. The changes in the NOC-LI concentrations were not parallelled by changes in DYN B-LI and MEAP LI, suggesting that NOC and opioid peptides elicit different reactions in the systems of nociception/antinociception. PMID- 10700571 TI - Localization and age-related changes of nitric oxide- and ANP-mediated cyclic-GMP synthesis in rat cervical spinal cord: an immunocytochemical study. AB - An immunocytochemical technique was used to study the localization and developmental aspects of cyclic GMP (cGMP)-synthesizing structures in the cervical spinal cord of 2-week and 3-month-old Lewis rats in response to the nitric oxide (NO) donor sodium nitroprusside (SNP) and/or atrial natriuretic peptide (ANP). By using cell-specific markers, the cell structures involved were investigated. To visualize cGMP, a combined technique of low- and high-power magnification, using a confocal laser scanning microscope was used. NOS-mediated cGMP synthesis was observed in the cervical spinal cord in laminae I, II and III in 14-day-old rats, which activity was mainly absent at the age of 3 months. The involvement of NO in the NMDA-mediated increase in cGMP immunostaining (cGMP-IS) was demonstrated by the absence of cGMP-IS in slices incubated in the presence of NMDA together with the NOS inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME). This NO-mediated effect of NMDA on cGMP-IS was completely absent in the 3-month old rats. ANP-mediated cGMP synthesis resulted in an increase in cGMP in laminae I and II, which was generally similar at both ages. Astrocytes in both white and gray matter were found to be cGMP-IS in the basal, NO- and ANP-stimulated conditions. Using confocal laser microscopy, NO-mediated cGMP synthesis was observed in large cholinergic terminals nearby motor neurons in the ventral horn. An extensive colocalization between NO-stimulated cGMP synthesis and parvalbumin positive (GABAergic) neurons and fibers was observed in all laminae. In the ANP stimulated condition, a colocalization with parvalbumin structures was found in laminae II and III. No NO- or ANP-mediated cGMP synthesis was found in fibers immunopositive for the presynaptic glutamate transporter, serotonin, or tyrosine hydroxylase. PMID- 10700572 TI - Ionic changes accompanying astrocytic intercellular calcium waves triggered by mechanical cell damaging stimulation. AB - Mechanically poking or damaging a single cell within a confluent astrocyte culture produces the so-called intercellular calcium (Ca(2+)) waves, that is, cell-to-cell propagating changes of intracellular free Ca(2+). We were interested whether intercellular Ca(2+) waves are also associated with changes in other intra- or extracellular ions. To that purpose, we investigated spatiotemporal changes of intracellular Ca(2+) (Ca(i)2+), sodium (Na(i)+) and protons (H(i)+) in primary cultures of rat cortical astrocytes using microfluorescence imaging with fura-2, SBFI and BCECF, respectively; changes of extracellular potassium (K(e)+) were monitored with K(+)-sensitive microelectrodes. Mechanical damage to a single cell by stimulation with a piezo-electrically driven micropipette initiated intercellular Ca(2+) waves that propagated to about 160 microm away from the stimulation point. Na(i)(+) increases could be detected in cells located 2-3 cell diameters from the stimulated cell, acidification was observed 1-2 cell diameters away and Ke(+) increases were measured up to 75 microm away. Kinetic analysis suggests that the Na(i)(+) and H(i)(+) changes occur after, and thus secondary to the Ca(i)(2+) changes. In contrast, K(e)(+) changes occurred very fast, even before the Ca(i)(2+) changes, but their propagation speed was too fast to implicate them as a trigger of Ca(i)(2+) changes. As Na(i)(+) is an important regulator of glycolysis in astrocytes, we hypothesize that astrocytic Na(i)(+) changes in cells located remotely from a damaged cell might be a signal that activates glycolysis thereby producing more lactate that is transferred to the neurons and increases their energy potential to survive the inflicted damage. PMID- 10700573 TI - Sodium valproate inhibits production of TNF-alpha and IL-6 and activation of NF kappaB. AB - Sodium valproate (VPA) is frequently used to treat epilepsy and convulsive disorders. Several reports have indicated that anti-epileptic drugs (AED) affect the immune system, but the mechanism has not been clear. We examined whether the commonly used AEDs, diazepam (DZP), carbamazepine (CBZ), phenobarbital (PB), phenytoin (PHT), and VPA, can inhibit activation of the nuclear transcription factor kappa B (NF-kappaB), in human monocytic leukemia cells (THP-1) and in human glioma cells (A-172). NF-kappaB is essential to the expression of the kappa light chain of immunoglobulin and proinflammatory cytokines. Electrophoretic mobility shift assays (EMSA) of nuclear extracts demonstrated that VPA inhibits NF-kappaB activation induced by lipopolysaccharide (LPS), but the other AEDs do not. Western blot analysis revealed that this inhibition is not linked to preservation of expression of IkappaBalpha protein. Chloramphenicol acetyltransferase (CAT) assay indicated that NF-kappaB-dependent reporter gene expression is suppressed in glioma cells pretreated with VPA. VPA significantly inhibited LPS-induced production of TNF-alpha and IL-6 by THP-1 cells, whereas other AEDs did not. The findings are consistent with the idea that VPA suppresses TNF-alpha and IL-6 production via inhibition of NF-kappaB activation. Our results suggest that VPA can modulate immune responses in vitro. These findings raise the possibility that such modulation might occur with clinical use of VPA. PMID- 10700574 TI - Frequency-dependent effects of glutamate antagonists on the vestibulo-ocular reflex of the cat. AB - In the central nervous system, sensory and motor signals at different frequencies are transmitted most effectively by neural elements that have different dynamic characteristics. Dynamic differences may be due, in part, to the dynamics of neurotransmitter receptors. For example, N-methyl-D-aspartate (NMDA) receptors are thought to be a component of the "neural integrator" of the vestibulo-ocular reflex (VOR), which generates a signal proportional to eye position. We measured the effects of blockade of NMDA and AMPA/kainate receptors on the gain and phase of the VOR at frequencies between 0.1 and 8 Hz in alert cats. The competitive NMDA antagonist, APV, and the non-competitive antagonists, MK-801 and ketamine, all caused a pronounced reduction in VOR gain. Gain was more strongly attenuated at low frequencies (0.1-1 Hz) than at higher frequencies (2-8 Hz). The phase lead of the eye with respect to the head was increased up to 30 degrees. In contrast, the reduction in gain associated with drowsiness or surgical anesthesia was not frequency-dependent. Blockade of AMPA/kainate receptors by the competitive antagonists, CNQX and NBQX, reduced the gain of the VOR at all frequencies tested. We evaluated our results using a control systems model. Our data are consistent with participation of NMDA receptors in neural integration, but suggest that NMDA receptors also participate in transmission by other components of the VOR pathway, and that neural integration also employs other receptors. One possibility is that between 0.1 and 10 Hz, higher-frequency signals are transmitted primarily by AMPA/kainate receptors, and lower frequencies by NMDA receptors. This arrangement would provide a biological substrate for selective motor learning within a small frequency range. PMID- 10700575 TI - Glial growth factor 2 induces proliferation and structural changes in ensheathing cells. AB - Ensheathing cells were isolated from neonatal rat olfactory bulbs and cultured in the presence of glial growth factor 2 (GGF2). Proliferation assay showed that at concentrations of up to 60 ng/ml GGF2, ensheathing cells underwent a modest increase in proliferation rate. This stimulation was not maintained at high doses of GGF2 at 100 ng/ml or more. Chemotaxis chambers and scanning electron microscopy were used to determine whether GGF2 was a chemoattractant for ensheathing cells. Although the results showed no chemotactic response to GGF2, ensheathing cells demonstrated structural changes when cultured in the presence of 20 ng/ml GGF2. Ultrastructural observations revealed that GGF2 promoted increased deposition of extracellular matrix on the cell membrane, more cytoskeletal elements in the processes and as a possible consequence, contributed to a more rigid support. Ensheathing cells cultured in the absence of GGF2 often extended thinner and curved processes. Reverse transcription-polymerase chain reaction confirmed the presence of GGF2 transcripts in ensheathing cells, suggesting that ensheathing cells themselves are a source of GGF2. PMID- 10700576 TI - DNA fragmentation in the CA2 sector of gerbil hippocampus following transient forebrain ischemia. AB - It has been reported that following transient forebrain ischemia in the gerbil, "delayed neuronal death" and "reactive change" occur in hippocampal CA1 and CA2 sectors, respectively. In the present study, using the gerbil transient forebrain ischemia model, we examined brain sections after various recirculation periods and demonstrated, employing the in situ nick-end labeling (TUNEL) method, a nuclear DNA fragmentation in the damaged CA2 neurons. PMID- 10700577 TI - An ovarian steroid hormone regimen that increases hypothalamic oxytocin expression alters [3H] muscimol binding in the hypothalamic supraoptic nucleus of the female rat. AB - Administration of sequential estradiol (E(2)) and progesterone (P) for 2 weeks followed by withdrawal of P 48 h prior to sacrifice will increase oxytocin (OT) messenger ribonucleic acid (mRNA) levels in the paraventricular and supraoptic nuclei (PVN and SON) of the ovariectomized rat. Progesterone is known to mediate certain of its effects via binding to the gamma aminobutyric acid A (GABA(A)) receptor. E(2) and P are known to modulate the specific binding of the GABA(A) receptor agonist, muscimol, in certain brain regions. In the present study ovariectomized rats received empty or steroid-filled Silastic capsules for 2 weeks according to one of the following schedules: E(2) only (E(2) group) vs. sequential E(2) and P in which P was either removed 48 h prior to killing (E(2)/P group) or sustained until sacrifice (E(2)/P+ group). [3H]muscimol binding was measured in several brain regions of the animals. The steroid sequence that is known to increase SON OT mRNA (E(2)/P-) selectively decreased [3H]muscimol binding in the SON of ovariectomized rats. The results suggest that changes in GABA(A) receptor binding may, in part, play a role in the regulation of steroid induced increases in hypothalamic OT expression. PMID- 10700578 TI - Anorexigenic cocaine- and amphetamine-regulated transcript peptide intensifies fear reactions in rats. AB - An increasing number of appetite-regulating peptides are being discovered. The list of regulators inhibiting food intake is considerably longer than that of appetite stimulators. In many cases, the peptides inhibiting food intake facilitate fear reactions, whereas the majority of the agents reducing anxiety responses stimulate appetite. Cocaine- and amphetamine-regulated transcript (CART) cDNA was isolated from hypothalamic libraries and CART was reported to inhibit food intake and to mediate the anorectic effects of leptin. Here, we show that the active core fragment of CART (CART(89-103), 0.04-5.0 nmol) injected into lateral cerebral ventricle not only inhibits food intake, but also causes a dose dependent increase in anxiety-like reactions in elevated plus-maze test. Intracerebroventricular administration of CART(82-103) (0.04-5.0 nmol) did not inhibit water intake and did not affect spontaneous locomotor activity in the open field test ruling out unspecific effects of the peptide. Our results suggest that CART could be an endogenous factor in the brain mediating the effects of stress on appetite. PMID- 10700579 TI - Gene mapping of SEZ group genes and determination of pentylenetetrazol susceptible quantitative trait loci in the mouse chromosome. AB - Gene mapping of the newly discovered SEZ genes (seizure-related genes) in the mouse was performed by linkage analysis. SEZ6 was on chromosome 11, SEZ12 on chromosome 16, SEZ15 on chromosome 3 and SEZ17 (PTZ17) on chromosome 18. The mouse chromosomal locus related to high susceptibility to pentylenetetrazol (PTZ) was also determined by linkage analysis using the recombinant inbred mouse, BXD (C57BLxDBA). A significant level of PTZ susceptibility was found on chromosome 2. Chromosomal loci of the newly discovered SEZ genes were not coincident with the significant chromosomal loci to PTZ susceptibility. Since epilepsy is assumed to be a disease syndrome which is probably manifested by abnormal expression of multifocal genes, determination of the role of each chromosomal locus in the provocation of seizure activity is important. PMID- 10700580 TI - Plectin immunopositivity appears in the astrocytes in the white matter but not in the gray matter after stab wounds. AB - Plectin is associated with intermediate filaments, such as glial fibrillary acidic protein (GFAP) and vimentin, which have an important role in the glial reactions after lesions. The present study investigates the plectin immunoreactivity following stab wounds in the cortex and the underlying white matter of adult rats. Without lesion, plectin immunoreactive astrocytes were not found in the cortex and only scarcely in the white matter. Following lesion, a number of astrocytes became immunopositive in the white matter, while no immunoreactivity was found in the overlying cortex. GFAP and vimentin showed intense immunopositivity in both areas. Therefore, the participation of the plectin seems to distinguish the glial reactions developed in the white or gray matters. PMID- 10700581 TI - Effect of melatonin on cocaine-induced behavioral sensitization. AB - Melatonin, a pineal hormone and a potent free radical scavenger with neuroprotective actions, has been reported to act as an inhibitor of nitric oxide synthase (NOS). We have earlier shown that inhibitors of NOS (N(omega)-nitro-L arginine methyl ester [L-NAME], 7-nitroindazole [7-NI]) block cocaine-induced behavioral sensitization. In the present study, the effects of melatonin on cocaine behavior were studied. A single injection of melatonin markedly augmented cocaine-induced locomotor activity. Rats injected daily with melatonin prior to cocaine injections failed to elicit cocaine sensitization. These behavioral effects of melatonin do not completely mimic those of other NOS inhibitors, suggesting that the effects of melatonin on cocaine behavior are mediated by both NOS-dependent as well as NOS-independent mechanisms. PMID- 10700582 TI - Increased granule cell neurogenesis in the adult dentate gyrus following mossy fiber stimulation sufficient to induce long-term potentiation. AB - Neurons are continually added at a low rate to the granule cell layer of the dentate gyrus during adulthood in rats. The functional significance of this unusual feature is not completely understood, although recent studies suggest continued granule cell neurogenesis is essential for normal learning and memory. We report here that, in the adult rat, stimulation of the granule cell mossy fibers sufficient to induce long-term potentiation (LTP) increases the number of newly formed granule cells in the dentate gyrus, indicating that granule cell neurogenesis is regulated by efferent activity and, possibly, the induction of LTP. PMID- 10700583 TI - Region-specific localization of glutamine synthetase immunoreactivity in the mouse olfactory bulb: implications for neuron-glia interaction in bulbar synaptic plasticity. AB - Glutamine synthetase (GS) critically regulates the metabolism of glutamate and gamma-amino butyric acid (GABA), which mediate synaptic plasticity in the olfactory bulb. In this study, GS immunolocalization in the mouse olfactory bulb was examined. The main and accessory subdivisions of the olfactory bulb possess GS-positive cells and processes in the plexiform-, the mitral- and the granule cell layers. GS has been demonstrated to show a predominantly astrocytic localization; its presence in the cell layers implicated in glutamatergic and GABAergic function therefore suggests that bulbar synaptic plasticity in mice may be regulated by astroglia and, together with other lines of evidence, point to the possibility of a functional astroglia-neuron system in the mouse olfactory bulb. PMID- 10700584 TI - Brain-derived mast cells could mediate histamine-induced inhibition of food intake in neonatal chicks. AB - In the present study, the effect of intracerebroventricular (i.c.v.) administration of histamine on food intake of neonatal chicks was examined over 2 h. Histamine (100, 200 or 400 nmol, respectively) was injected in the lateral ventricle of 2-day-old chicks, and cumulative food intakes were measured. i.c.v. injection of histamine significantly inhibited food intake in a dose-dependent manner. In addition, compound 48/80, which causes degranulation of mast cells and release of histamine, or thioperamide, which is an antagonist of the histamine H3 autoreceptor and increases histamine release from histaminergic nerve terminals, was injected i.c.v. to clarify whether mast cell- or neuron-derived histamine in the central nervous system of chicks is essential to the feeding inhibition. Central administration of compound 48/80 inhibited food intake with a dose dependent manner, but thioperamide had no effect on feeding. An inhibitor of mast cell degranulation, sodium cromoglycate, somewhat attenuated food intake inhibited by compound 48/80. These results suggest that brain-derived mast cells could be a major source of histamine in the inhibition of food intake of neonatal chicks. PMID- 10700585 TI - Sustained facilitatory action of FK960 on hippocampal neurotransmission. AB - The present study investigated the effect of N-(4-acetyl-1-piperazinyl)-p fluorobenzamide monohydrate (FK960), a piperazine derivative developed as antidementia drug, on hippocampal neurotransmission. FK960 increased the amplitude of population spikes (PSs) from the granular cell layer in rat hippocampal slices in a bell-shaped dose-dependent manner at concentrations ranged from 10 nM to 10 microM. A similar potentiation was found in the intact mouse hippocampus, the action being still evident 120 min after injection; a maximal effect was obtained with 3 mg/kg intraperitoneal injection of FK960 at concentrations ranged from 0.3 to 5 mg/kg. The facilitatory action induced by FK960 did not occlude the potentiation induced by tetanic stimulation. The results of the present study indicate that FK960 induces a long-lasting facilitation of hippocampal neurotransmission, but by a mechanism independent of the tetanic long-term potentiation. This may account for the memory enhancing action of FK960. PMID- 10700586 TI - Stimulus pattern related plasticity of synapses between cones and horizontal cells in carp retina. AB - Stimulus pattern related synaptic plasticity in the luminosity-type horizontal cell (LHC) of isolated carp retina was investigated. The major findings were: (1) repetitive red flashes progressively strengthened the synaptic connection between red-cone and LHC, whereas weakened that between green-cone and LHC; (2) repetitive green flashes remarkably depressed the LHC's red response, but caused little changes in the cell's green response. A competitive depression between different cone signals is suggested. PMID- 10700587 TI - Quantitative differences between kinetic properties of Na(+) currents in postganglionic sympathetic neurones projecting to muscular and cutaneous effectors. AB - The activity of muscular and cutaneous sympathetic neurones has been shown to be differentially regulated. The differences may partially stem from the different ionic channel expression and current kinetics in these neurones, particularly that of Na(+) channels, which play a critical role in action potential generation and modulation of neuronal excitability. The whole cell patch-clamp technique was used to compare the kinetic properties of Na(+) currents in two groups of sympathetic neurones identified by the fluorescent tracer Fast Blue: putative muscular sympathetic neurones (PMSN) and putative cutaneous sympathetic neurones (PSSN). The tracer was injected into the muscular part of the diaphragm (to mark PMSN) and into the skin of the ear (to mark PSSN). Both kinds of neurones expressed fast activating, fast inactivating, voltage dependent and TTX sensitive Na(+) currents. However, the electrical characteristics of the cells were markedly different: (1) The capacitance of PMSN (21.7 pF) was larger than PSSN (12.7 pF). Maximum current in PMSN (3.1 nA) was also larger than in PSSN (2.0 nA). Calculated current density was smaller in PMSN (148.0 pA/pF) than in PSSN (181.1 pA/pF). Slope conductance was larger in PMSN compared to PSSN (102.7 nS and 73.6 nS respectively). (2) V(1/2) of activation for PMSN (-20.9 mV) was more negative than the potential recorded for PSSN (-16.7 mV); the slope factors were not different. (3) V(1/2) for inactivation was more negative for PMSN than for PSSN (-66.3 vs. -60.8 mV); again, the slope factors for inactivation were not different. (4) The rate of recovery from inactivation could be described by the sum of two exponential functions. In PMSN the fast and slow recovery exponential factors tau(f) and tau(s) were 12.6 (66%) and 83.9 (34%) ms, while in PSSN they were shorter and equalled 8.2 (62%) and 41.9 (38%) ms, respectively. We conclude that the Na(+) currents of PMSN and PSSN have different kinetic properties. PMID- 10700588 TI - Dynorphin B and spinal analgesia: induction of antinociception by the cannabinoids CP55,940, Delta(9)-THC and anandamide. AB - The endogenous opioid dynorphin B was evaluated for its role in cannabinoid induced antinociception. Previous work in our laboratory has shown that the synthetic, bicyclic cannabinoid, CP55,940, induces the release of dynorphin B whilst the naturally occurring cannabinoid, Delta(9)-tetrahydrocannabinol (Delta(9)-THC), releases dynorphin A. The dynorphins contribute in part to the antinociceptive effects of both cannabinoids at the level of the spinal cord. The present study compares dynorphin B released from perfused rat spinal cord in response to acute administration of anandamide (AEA), Delta(9)-THC and CP55,940 at two time points, 10 min and 30 min post administration, and attempts to correlate such release with antinociceptive effects of the drugs. Dynorphin B was collected from spinal perfusates of rats pretreated with Delta(9)-THC, CP55,940 or AEA. The supernatant was lyophilized and the concentrations of dynorphin B were measured via radioimmunoassay. At a peak time of antinociception (10 min), CP55,940 and Delta(9)-THC induced significant two-fold increases in the release of dynorphin B. AEA did not significantly release dynorphin B. Upon a 30-min pretreatment with the drugs, no significant dynorphin B release was observed, although antinociceptive effects persisted for CP55,940 and Delta(9)-THC. Previous work indicates that Delta(9)-THC releases dynorphin A while AEA releases no dynorphin A. This study confirms that although all three test drugs produced significant antinociception at 10 min, the endocannabinoid, AEA, does not induce antinociception via dynorphin release. Thus, our data indicate a distinct mechanism which underlies AEA-induced antinociception. PMID- 10700589 TI - Dopamine toxicity in neuroblastoma cells: role of glutathione depletion by L-BSO and apoptosis. AB - Dopamine (DA), while an essential neurotransmitter, is also a known neurotoxin that potentially plays an etiologic role in several neurodegenerative diseases. DA metabolism and oxidation readily produce reactive oxygen species (ROS) and DA can also be oxidized to a reactive quinone via spontaneous, enzyme-catalyzed or metal-enhanced reactions. A number of these reactions are cytotoxic, yet the precise mechanisms by which DA leads to cell death remain unknown. In this study, the neuroblastoma cell line, SK-N-SH, was utilized to examine DA toxicity under varying oxidant states. Cells pretreated with the glutathione (GSH)-depleting compound, L-buthionine sulfoximine (L-BSO), exhibited enhanced sensitivity to DA compared to controls (non-GSH-depleted cells). Furthermore, in cells pretreated with L-BSO, the addition of ascorbate (250 microM) afforded significant protection against DA-induced toxicity, while pyruvate (500 microM) had no protective effect. To further characterize the possibility that DA is associated with oxidative stress, additional studies were carried out with manganese (30 microM) as a pro-oxidant. Manganese and DA (200 microM), although not cytotoxic when individually administered to SK-N-SH cells, had a synergistic action on cytotoxicity. Finally, morphological and molecular markers of programmed cell death (apoptosis) were observed in cells treated with DA and L-BSO. These markers included membrane blebbing and internucleosomal DNA fragmentation. These results suggest that DA toxicity is tightly linked to intracellular oxidant/antioxidant levels, and that environmental factors, such as excessive Mn exposure, may modulate cellular sensitivity to DA. PMID- 10700590 TI - LPS-induced Fos expression in oxytocin and vasopressin neurons of the rat hypothalamus. AB - The aim of this study was to examine the involvement of the hypothalamic oxytocin (OXT) and vasopressin (AVP) neurons in acute phase reaction using quantitative dual-labeled immunostaining with Fos and either OXT and AVP in several hypothalamic regions. Administration of low dose (5 microg/kg) and high dose (125 microg/kg) of LPS induced intense nuclear Fos immunoreactivity in many OXT and AVP neurons in all the observed hypothalamic regions. The percentage of Fos positive nuclei in OXT magnocellular neurons was higher than that of AVP magnocellular neurons in the supraoptic nucleus (SON), the magnocellular neurons in the paraventricular nucleus (magPVN), rostral SON (rSON), and nucleus circularis (NC), whose axons terminate at the posterior pituitary for peripheral release. The percentage of Fos-positive nuclei in AVP parvocellular neurons in the paraventricular nucleus (parPVN) was higher than that of OXT parvocellular neurons, whose axons terminate within the brain for central release. Moreover, the percentage of Fos-positive nuclei in AVP magnocellular neurons of the SON and rSON was significantly higher than that of the magPVN and NC when animals were given LPS via intraperitoneal (i.p.)-injection. This regional heterogeneity was not observed in OXT magnocellular neurons of i.p.-injected rats or in either OXT or AVP magnocellular neurons of intravenous (i.v. )-injected rats. The present data suggest that LPS-induced peripheral release of AVP and OXT is due to the activation of the magnocellular neurons in the SON, magPVN, NC, and rSON, and the central release of those hormones is in part derived from the activation of parvocellular neurons in the PVN. It is also suggested that the activation of AVP magnocellular neurons is heterogeneous among the four hypothalamic regions, but that of OXT magnocellular neurons is homogenous among these brain regions in response to LPS administration. PMID- 10700591 TI - A novel action of the newly described prolactin-releasing peptides: cardiovascular regulation. AB - The physiological relevance of the recently described prolactin-releasing peptides (PrRPs) has yet to be established. Here, we demonstrate the low potency of the PrRPs (minimum effective dose: 100 nM), compared to that observed for thyrotropin-releasing hormone (TRH, minimum effective dose: 1.0 nM), to stimulate prolactin (PRL) release from cultured pituitary cells harvested from lactating female rats. Anatomic studies question the role of these peptides in neuroendocrine control of lactotroph function. Instead, peptide and peptide receptor mapping studies suggest potential actions in hypothalamus and brainstem unrelated to the control of anterior pituitary hormone secretion. Intracerebroventricular (i.c.v. ) administration of both PrRP-20 and PrRP-31 (0.4 and 4.0 nmol) resulted in significantly increased mean arterial blood pressure in conscious, unrestrained rats [peak elevations vs. baseline: PrRP-20, 10% and 16%, low and high dose peptide; PrRP-31, 7% and 10%; compared to the response to 0.1 nmol angiotensin II (A II), 15-17%]. Similar doses of peptide did not significantly alter water drinking in response to overnight fluid deprivation, or thirst or salt appetite in response to an isotonic hypovolemic challenge. Thus, the effect on blood pressure appeared relatively specific. We suggest that these peptides, identified originally as ligands for a receptor found in abundance in pituitary gland, play a broader role in brain function and that the ability of them to stimulate PRL release may not represent their primary biologic function. PMID- 10700592 TI - Neurotoxic relationship between dopamine and iron in the striatal dopaminergic nerve terminals. AB - The neurotoxic effect of dopamine (DA) and iron(III) on DAergic terminals in striatum has been studied by intracerebral microdialysis technique. Twenty-four hours after surgery (day 1), DA and/or iron(III) with and without DA reuptake inhibitor, nomifensine, were perfused for 1 h. Forty-eight hours after surgery (day 2), MPP(+) 1 mM was perfused for 15 min and the output of DA was measured, its amount being directly proportional to the remaining striatal DAergic terminals, supported by tyrosine hydroxylase immunohistochemistry technique. Perfusion of exogenous DA, as well as iron(III) 10 and 100 microM, did not produce any neurotoxic effect. However, perfusion of iron(III) (333 and 1000 microM) produced a concentration-dependent toxic effect. Co-perfusion of iron(III) at non-toxic concentration (100 microM) with DA (15 microM) produced a toxic effect. Elevation of the endogenous extracellular levels of DA by inhibiting its uptake with nomifensine increased the neurotoxic effect of iron(III) in a dose-dependent manner. The use of tetrodotoxin after elevation of DA with nomifensine partially prevented the neurotoxic effect of its co-perfusion with iron(III) (100 microM). These results suggest that DAergic system could be synergistically damaged by DA and iron(III). Thus, alterations in the clearance of DA from extracellular space along with an increase of iron may have significant consequences for DAergic system toxicity. PMID- 10700593 TI - Cyclic AMP and protein kinase A rhythmicity in the mammalian suprachiasmatic nuclei. AB - The levels of cyclic AMP and protein kinase A, as well as the activity of this enzyme, were measured in the hamster suprachiasmatic nuclei at different time points throughout the daily or circadian cycle. Significant diurnal variations for levels of AMPc and the catalytic subunit of protein kinase A and the activity of this enzyme were found. All of these parameters tended to increase throughout the nocturnal phase, reaching higher values at the end of the night and the beginning of the day and minimal values around the time of lights off. This rhythmicity appears to be under exogenous control, since constant darkness abolished fluctuations throughout the circadian cycle. In vitro incubation in the presence of melatonin during the day significantly decreased cyclic AMP levels and basal protein kinase A activity in the SCN, while neither neuropeptide Y nor light pulses affected these parameters. These results suggest a significant diurnal regulation of the cyclic AMP-dependent system in the hamster circadian clock. PMID- 10700594 TI - Developmental and pathological expression of peroxisomal enzymes: their relationship of D-bifunctional protein deficiency and Zellweger syndrome. AB - We present the developmental changes of peroxisomal enzymes, catalase, L bifunctional protein (L-BF) and D-bifunctional protein (D-BF), in the normal brains, and patients with D-BF deficiency, a new peroxisomal disease. D-BF immunoreactivity was observed in controls as early as 13 gestational weeks (GW) and increased with maturation. The adult pattern with fine granule staining of somata and dendrites became apparent in adolescence. L-BF appeared at 20 GW in the cerebral cortex and Purkinje cells and positive glia appeared early in the white matter at 17 GW, and then increased with age. Catalase-positive neurons were identified in the same manner as L-BF, D-BF deficiency in both fetus and infant showed markedly diminished enzyme immunoreactivity. Patients demonstrate reduced D-BF expression. Zellweger syndrome shows decreased expression for the three proteins. This study shows that the peroxisomal enzymes may be closely related to neuronal maturation and gliogenesis in human brain and to disturbance of neuronal migration as seen in Zellweger syndrome significant. D-BF deficiency may exhibit a range of symptoms during the neonatal and early infantile periods some of which may be similar to Zellweger syndrome. PMID- 10700595 TI - Intraperitoneal and intraamygdala N(6)-cyclohexyladenosine suppress hippocampal kindled seizures in rats. AB - Effects of intraperitoneal and intraamygdala N(6)-cyclohexyladenosine (CHA), a selective adenosine A(1) receptor agonist, and 1,3-dimethyl-8-cyclopentylxanthine (CPT), a selective adenosine A(1) receptor antagonist, were examined in fully hippocampal kindled rats. Intraperitoneal administration of CHA (0. 25, 0.5 and 1 mg/kg) decreased hippocampal secondary afterdischarge duration (SAD) and amygdala afterdischarge duration (ADD). Only the 1 mg/kg dose induced a significant increase in latency to stage 4. Intraperitoneal administration of CPT (0.25, 0.5 and 1 mg/kg) induced a significant increase in stage 5 duration, hippocampal SAD and ADD. Pretreatment of animals with CPT (1 mg/kg), antagonized effects of CHA on seizure parameters. Intraamygdala microinfusion (1 microl over 2 min) of CHA (5 nM-1 mM) significantly reduced hippocampal SAD and amygdala ADD. These effects were antagonized by intraamygdala CPT (1 microM). Results obtained suggest that in hippocampal kindled rats, amygdala may be regarded as a relay point for AD propagation specially in recruit activity of the hippocampus. PMID- 10700596 TI - Neutrophil elastase inhibition reduces cerebral ischemic damage in the middle cerebral artery occlusion. AB - It has been reported that activated neutrophils are involved in the development of cerebral damage induced by ischemia. Activated neutrophils release a lot of mediators including toxic oxygen metabolites, elastase and cytokines which damage brain tissue. Therefore, we investigated roles of neutrophil elastase in the development of cerebral damage using an elastase inhibitor, ONO-5046. The rat middle cerebral artery (MCA) was occluded by a thrombus induced by photochemical reaction between green light and the photosensitizer dye, Rose Bengal. Photochemical reaction causes endothelial injury followed by formation of a platelet and fibrin-rich thrombus at the site of the irradiation. Photochemical reaction is routinely used in our laboratory to produce arterial occlusion in experimental animals. Twenty-four hours after the MCA occlusion, the size of cerebral damage was measured by histochemical technique. Water content in the brain was measured and neuronal deficits were examined 24 h after the MCA occlusion. ONO-5046 was administered at various doses as continuous infusion for 24 h, starting just after the MCA occlusion or from 3 h after. ONO-5046 at doses of 10 and 30 mg/kg/h significantly (p<0.05 and p<0.01, respectively) reduced the size of cerebral damage and water content (p<0.05, p<0.01, respectively) in different eight rats. Further, ONO-5046 at a dose of 30 mg/kg/h significantly (p=0.01) improved neuronal deficits. ONO-5046 which was administered starting from 3 h after the MCA occlusion, also reduced the size of cerebral damage. Neutropenia by anti-neutrophil antibody injection significantly (p<0. 01) reduced the size of cerebral damage. Elastase released from activated neutrophils may play a key role in the development of cerebral damage. PMID- 10700597 TI - Glutamate stimulates ascorbate transport by astrocytes. AB - The concentrations of glutamate and ascorbate in brain extracellular fluid increase following seizure activity, trauma and ischemia. Extracellular ascorbate concentration also rises following intracerebral glutamate injection. We hypothesized that glutamate triggers the release of ascorbate from astrocytes. We observed in primary cultures of rat cerebral astrocytes that glutamate increased ascorbate efflux significantly within 30 min. The half-maximal effective concentration of glutamate was 180+/-30 microM. Glutamate-stimulated efflux of ascorbate was attenuated by hypertonic media. 4,4'-diisothiocyanatostilbene-2,2' disulfonic acid inhibited both Na(+)-dependent glutamate uptake and ascorbate efflux. Two other inhibitors of volume-sensitive organic anion channels (1, 9 dideoxyforskolin and 5-nitro-2-(3-phenylpropylamino) benzoic acid) did not slow glutamate uptake but prevented stimulation of ascorbate efflux. Glutamate also stimulated the uptake of ascorbate by ascorbate-depleted astrocytes. In contrast, glutamate uptake was not affected by intracellular ascorbate, thus ruling out a putative glutamate-ascorbate heteroexchange mechanism. These results are consistent with activation by glutamate of ascorbate-permeant channels in astrocytes. PMID- 10700598 TI - Low plasma epinephrine in elderly female subjects of dementia of Alzheimer type. AB - One of the robust features of brain pathologies of dementia of Alzheimer type (DAT) is the impairment of the hippocampus, especially the cholinergic system. Several animal studies have suggested that the cholinergic system in the hippocampus is involved in the control of the plasma level of catecholamines and glucose. The stimulation of the hippocampal cholinergic system has resulted in the elevation of plasma catecholamines and glucose in rats. In the present study, we measured the plasma level of epinephrine, norepinephrine, dopamine, glucose, and insulin during a fasting state in the morning in hospitalized DAT (n=66), vascular dementia (VD) (n=28), or non-demented (ND) (n=21) females (mean age DAT=82. 49+/-4.98, VD=82.86+/-5.86, ND=82.95+/-7.77, respectively). Statistical analysis showed that the plasma level of epinephrine during a fasting state in DAT subjects was significantly lower than that of ND subjects; however, in VD subjects the level of epinephrine was not different from that of ND subjects. Other values did not differ significantly among the groups. PMID- 10700599 TI - Dose-response study of caffeine effects on cerebral functional activity with a specific focus on dependence. AB - Caffeine is a behavioral stimulant consumed on a worldwide basis. The question of whether caffeine is addictive has been debated for over a decade. Caffeine acts as a mild positive reinforcer but is not consistently self-administered in humans or animals. With [14C]2-deoxyglucose autoradiography, we studied the effects of increasing doses of caffeine on cerebral glucose utilization in rats. At 1 mg/kg, caffeine activated the caudate nucleus mediating locomotion, and the raphe nuclei and locus coeruleus involved with mood and sleep. After 2.5 and 5 mg/kg caffeine, metabolic activation spread to other components of the nigrostriatal dopaminergic system, the thalamus, ventral tegmental area and amygdala. The functional activation of the shell of the nucleus accumbens, an area involved in addiction and reward, was only induced by the highest dose of caffeine, 10 mg/kg. At this dose, the activation of the shell of the nucleus accumbens occurred together with that of the core of the nucleus accumbens and of most other brain regions. These data correlate well with the known sensitivity of locomotion, mood and sleep to low doses of caffeine. They also show that low doses of caffeine which reflect the usual human level of consumption fail to activate reward circuits in the brain and thus provide functional evidence of the very low addictive potential of caffeine. PMID- 10700600 TI - Cortistatin modulates memory processes in rats. AB - Cortistatin (CST) is a recently described neuropeptide with high structural homology with somatostatin. Its mRNA is restricted to gamma amino butyric acid (GABA)-containing cells in the cerebral cortex and hippocampus. CST modulates the electrophysiology of the hippocampus and cerebral cortex of rats; hence, it may be modulating mnemonic processes. In this study, we have evaluated the effect of CST and somatostatin (SS) on short- and long-term memory (STM and LTM, respectively), as well as on the extinction of the behavior by using the footshock passive avoidance behavioral test. In addition, we tested the ability of both neuropeptides to affect the generation of cAMP in hippocampal neurons in culture. Results showed that the administration of either CST or SS into the hippocampal CA1 deteriorates memory consolidation in a dose-response fashion and facilitates the extinction of the learned behavior. CST was more potent than SS. Likewise, CST increases cAMP while SS decreases it. These results strongly support a modulatory role for CST in memory processes. PMID- 10700601 TI - T-588, a novel neuroprotective agent, delays progression of neuromuscular dysfunction in wobbler mouse motoneuron disease. AB - R(-)-1-(benzo[b]thiophen-5-yl)-2-[2-(N,N-diethylamino) ethoxy]ethanol hydrochloride (T-588) enhances acetylcholine release from the frontal cortex and hippocampus in rats, and can ameliorate cognitive dysfunction in various amnesia models of rodents. T-588 protects rat cerebellar granule cells from glutamate neurotoxicity in culture. This agent also inhibits facilitation in the crayfish neuromuscular junction and mammalian cerebellum. Clinical trials of T-588 are underway in patients with Alzheimer's disease. We attempted to determine whether T-588 treatment ameliorates neuromuscular dysfunction in the wobbler mouse, an animal model of motoneuron disease (MND). After the initial diagnosis of MND at the age of 3-4 weeks, wobbler mice were orally administered T-588 (3, 10, 30 mg/kg) or vehicle daily for 4 weeks in a blinded fashion. We compared symptomatic, pathological and biochemical changes among the groups. In comparison with vehicle, T-588 administration potentiated grip strength, attenuated forelimb contracture and increased the weight of the biceps muscles. T-588-treated mice had retarded denervation muscle atrophy and elevated activities of choline acetyltransferase (ChAT) or lactate dehydrogenase in the biceps muscles. T-588 treatment also enhanced ChAT activities and promoted formation of cyclic adenosine monophosphate in the cervical cord. Pharmacokinetic study also showed that T-588 was transported efficiently into the cerebrum and spinal cord following oral administration. Thus, T-588 treatment delayed the progression of wobbler murine MND. Our findings suggest that this agent has therapeutic potential in human motor neuropathy or MND. PMID- 10700602 TI - Comparative study of fluoxetine, sibutramine, sertraline and dexfenfluramine on the morphology of serotonergic nerve terminals using serotonin immunohistochemistry. AB - We compared the effects of treatment with high doses of fluoxetine, sibutramine, sertraline, and dexfenfluramine for 4 days on brain serotonergic nerve terminals in rats. Methylenedioxymethamphetamine (MDMA) and 5,7-dihydroxytryptamine (5,7 DHT) were used as positive controls because both compounds deplete brain serotonin. Food intake and body weight changes were also monitored and yoked, pair-fed animals were used to control for possible changes in morphology due to nutritional deficits. Fluoxetine, sibutramine, sertraline and dexfenfluramine all produced a significant reduction in body weight. Fluoxetine, sibutramine and sertraline treatment resulted in no depletion of brain serotonin but produced morphological abnormalities in the serotonergic immunoreactive nerve network. In contrast, dexfenfluramine and MDMA depleted brain serotonin and produced morphological changes in the serotonin nerve network. These results indicate that even though fluoxetine, sibutramine and sertraline do not deplete brain serotonin, they do produce morphological changes in several brain regions (as identified by serotonin immunohistochemistry). Dexfenfluramine and MDMA, on the other hand, markedly deplete brain serotonin and also produce morphological changes. Collectively, these results lend support to the concept that all compounds acting on brain serotonin systems, whether capable of producing serotonin depletion or not, could produce similar effects on the morphology of cerebral serotonin systems. PMID- 10700603 TI - SYM-2081 a kainate receptor antagonist reduces allodynia and hyperalgesia in a freeze injury model of neuropathic pain. AB - Cold-freeze injury at -4 degrees C to the rat sciatic nerve produces mechanical allodynia and thermal hyperalgesia [M.A. Kleive, P.S. Jungbluth, J.A. Uhlenkamp, K.C. Kajander, Cold injury to rat sciatic nerve induces thermal hyperalgesia or analgesia, 8th World Congress on Pain, Vancouver, BC, Canada, August 1996 (Abstract).]. The NMDA receptor, an excitatory amino acid (EAA) receptor, appears to be involved in the development of allodynia and hyperalgesia following nerve injury. The role, if any, of the kainate receptor, another EAA receptor, remains unknown. In the current study, we evaluated whether (2S,4R)-4-methylglutamic acid (SYM-2081), a recently developed kainate receptor antagonist, attenuates increased responsiveness following cold injury to the sciatic nerve. During baseline testing, Sprague-Dawley rats were evaluated for frequency of withdrawal from von Frey filaments and latency of withdrawal from a radiant thermal source. Animals were then anesthetized, the left sciatic nerve was exposed, and the nerve was cooled to -4 degrees C for 15 min (n=24). For control rats (n=24), all procedures were identical except that the nerve was maintained at 37 degrees C. Testing resumed on the third day following surgery. On the fifth post-operative day, SYM-2081 (150 or 100 mg/kg), fentanyl citrate (0. 04 mg/kg) or vehicle was injected intraperitoneally. Injury to the rat sciatic nerve induced a significant increase in withdrawal frequency and a significant decrease in withdrawal latency (ANOVA, p<0.05). SYM-2081 and fentanyl significantly reduced these responses (p<0.05). These results suggest that kainate and opioid receptors are involved in the mechanical allodynia and thermal hyperalgesia that develop following cold injury to the sciatic nerve. PMID- 10700604 TI - S-100beta protects cultured neurons against glutamate- and staurosporine-induced damage and is involved in the antiapoptotic action of the 5 HT(1A)-receptor agonist, Bay x 3702. AB - The serotonin (5-HT)(1A) receptor agonists have already been shown to protect cultured neurons from excitotoxic as well as from apoptotic damage [B. Ahlemeyer, J. Krieglstein, Stimulation of 5-HT(1A) receptors inhibits apoptosis induced by serum deprivation in cultured neurons from chick embryo, Brain Res. 777 (1997) 179-186. ; B. Ahlemeyer, A. Glaser, C. Schaper, I. Semkova, J. Krieglstein, The 5 HT(1A) receptor agonist, Bay x 3702, inhibited apoptosis induced by serum deprivation in cultured neurons, Eur. J. Pharmacol. 370 (1999) 211-216.; J.H.M. Prehn, M. Welsch, C. Backhauss, J. Nuglisch, F. Ausmeier, C. Karkoutly, J. Krieglstein, Effects of serotonergic drugs in experimental brain ischemia: evidence for a protective role of serotonin in cerebral ischemia, Brain Res. 630 (1993) 110-120.; I. Semkova, P. Wolz, J. Krieglstein, Neuroprotective effect of 5 HT(1A) receptor agonist, Bay x 3702, demonstrated in vitro and in vivo, Eur. J. Pharmacol. 359 (1998) 251-260.; B. Suchanek, H. Struppeck, T. Fahrig, The 5 HT(1A) receptor agonist, Bay x 3702, prevents staurosporine-induced apoptosis, Eur. J. Pharmacol. 355 (1998) 95-101.] and to increase the release of the neurotrophic protein, S-100beta [P.M. Whitaker-Azmitia, R. Murphy, E.C. Azmitia, Stimulation of astroglial 5-HT(1A) receptors releases the serotonergic growth factor, protein S-100, and alters astroglial morphology, Brain Res. 497 (1989) 80 86. ; P.M. Whitaker-Azmitia, R. Murphy, E.C. Azmitia, S-100 protein is released from astroglial cells by stimulation of 5-HT(1A) receptors, Brain Res. 528 (1990) 155-158.]. In this study, we tried to find out whether S-100beta can protect cultured neurons from glutamate- and staurosporine-induced damage and whether the neuroprotective activity of the highly selective 5-HT(1A) receptor agonist, Bay x 3702, is mediated by an induction of S-100beta. Extracellularly added S-100beta (1-10 ng/ml) reduced staurosporine-induced damage in pure neuronal cultures from chick embryo telencephalon as well as in mixed neuronal/glial cultures from neonatal rat hippocampus. In addition, S-100beta (1 ng/ml) reduced neuronal death induced by exposure to glutamate (0.25 mM, 30 min) in mixed neuronal/glial cultures from neonatal rat hippocampus. In cultured rat cortical astrocytes, a 24 h-treatment with Bay x 3702 (1 nM) increased the S-100beta content in the culture medium from 2.2+/-0.3 (controls) to 6.2+/-0.7 ng/ml. In the adult rat, a 4 h infusion of 4 microg/kg Bay x 3702 (i.v.) was found to increase the S-100beta content in the striatum 6 h after the beginning of the infusion to 153+/-37 microg/g compared with 60+/-20 microg/g in vehicle-treated rats. Bay x 3702 had no effect on the S-100beta content in the rat hippocampus. Finally, we tried to block the protective effect of Bay x 3702 against staurosporine-induced damage in mixed neuronal/glial cultures from rat neonatal hippocampus by anti-S-100beta antibodies. We found only a partial blockade, although the antibodies fully blocked the antiapoptotic effect of S-100beta itself demonstrating that the antibody was effective in blocking neuroprotection by S-100beta. Thus, we conclude that S-100beta was able to protect cultured neurons against glutamate- and staurosporine-induced damage. Furthermore, S-100beta mediated partially the protective effect of the 5-HT(1A) receptor agonist, Bay x 3702, against staurosporine-induced apoptosis in mixed neuronal/glial cultures from neonatal rat hippocampus. PMID- 10700605 TI - Effects of naloxone on the serum luteinizing hormone level and the number of Fos positive gonadotropin-releasing hormone neurons in immature female rats. AB - To examine developmental changes in the number of gonadotropin-releasing hormone (GnRH) neurons activated by an opioid receptor antagonist in female rats, blood sampling and double-labeled immunocytochemistry for Fos and GnRH were performed after the injection of naloxone (NAL) in immature (postnatal d16 and d30) and mature female rats. Three age groups of rats were perfused with 4% paraformaldehyde-PB 90 min after the subcutaneous injection of NAL (2.5 mg/kg) or saline. All tissue incubation and staining for double-labeled immunocytochemistry were simultaneously performed. Although no significant developmental change was observed in the total number of GnRH neurons (p0.05), NAL-induced increases in serum luteinizing hormone (LH) concentrations were much greater in the d16 group than those in the d30 and mature groups (p<0.01). Conversely, Fos-positive GnRH neurons were rarely observed in d16, and some Fos-positive GnRH neurons were observed in the d30 group (p<0.05 vs. saline) and the mature group (p<0.01 vs. saline). These results suggest that opiatergic inhibitory system on GnRH neuron in immature female rats is different from that in mature female rats. PMID- 10700606 TI - Distribution of mapacalcine receptors in the central nervous system of rat using the [125I]-labeled mapacalcine derivative. AB - Mapacalcine is a dimeric protein of Mr 19041 extracted from the marine sponge Cliona vastifica. Electrophysiological and pharmacological approaches have demonstrated that mapacalcine was blocking a calcium channel different from N-, L , P-, T- or Q-type calcium channels on mouse intestinal smooth muscle. Recently a [125I]-labeled derivative of mapacalcine has been synthesized and characterized as a tool usable as a probe to investigate mapacalcine receptors. On rat brain membranes, it binds to its receptor with a K(d)=0.35 nM and a maximal binding capacity of 706 fmol/mg protein. We use here [125I]-mapacalcine to study the mapping of its receptors in the rat brain. Data obtained show a practically homogeneous labeling of the brain. Our experiments suggest that mapacalcine receptors are present on neuronal and glial cells. Interestingly, choroid plexus demonstrates a high density of mapacalcine receptors. These data would suggest that mapacalcine sensitive calcium channels could be involved in the control of calcium homeostasis of the cerebrospinal fluid. PMID- 10700607 TI - Magnetic field desensitizes 5-HT(1B) receptor in brain: pharmacological and functional studies. AB - It was previously suggested that exposure to magnetic fields (MFs) could generate dysfunction of the CNS. The physiological manifestations described lead us to postulate that these symptoms might be related to a dysfunction of the serotonergic system and particularly of the 5-HT(1B) receptors. Accordingly, MFs could modify the conformation of these receptors altering their functional activities. In rat brain membrane preparations, we showed that the affinity constant of 5-HT for 5-HT(1B) receptors was modified under exposure to MFs since K(d) varied from 4.7+/-0.5 to 12+/-3 nM in control and exposed (2.5 mT) membranes, respectively. This effect was intensity-dependent (the sigmoidal dose response curve was characterized by an EI(50) of 662+/-69 microT and a maximal increase of 321+/-13% of the control K(d)), reversible, temperature-dependent and specific to the 5-HT(1B) receptors. Similar results have also been obtained with the human 5-HT(1B) receptors. In parallel assays, the functional activity of 5 HT(1B) receptors was investigated. The capacity of a 5-HT(1B) agonist to inhibit the cAMP production was reduced by 37% (53.7+/-3.5% to 33.7+/-4.1%) following exposure to MFs and the cellular activity of the receptors (inhibition of the synaptosomal release of 5-HT) also was markedly reduced (66.5+/-3.2% to 28.5+/ 4.2%). These results clearly show that in in vitro assays, MF specifically interacts with 5-HT(1B) receptors, inducing structural changes of the protein that result in a functional desensitization of the receptors. Thus, in vivo, exposure to MFs may lead to physiological changes, particularly in the field of mood disorders where the 5-HT system is strongly involved. PMID- 10700608 TI - The core-shell dichotomy of nucleus accumbens in the rhesus monkey as revealed by double-immunofluorescence and morphology of cholinergic interneurons. AB - Double-immunolabelling experiments for the combinations, calretinin (CR) calbindin, CR-tyrosine hydroxylase (TH) and calbindin-TH, were performed in rhesus monkeys to compare the chemical organization of the nucleus accumbens (ACC) in primates and rodents. Additionally, the soma sizes and numbers of primary dendrites of cholinergic neurons in the subregions of ACC were compared with those of caudate-putamen. Our findings subserve the shell-core concept also in the primate ACC, as like in the rat, CR immunoreactivity (-ir) due to intense neuropil labelling is very strong in the shell of rhesus monkey, but poor in the core. The staining intensity of this marker decreases in dorsoventral direction. An almost complementary pattern was noted in sections of the monkey ACC immunostained for both calbindin and TH. The cholinergic interneurons of the nucleus caudatus-putamen are clearly distinguished from those of the ACC and insula Calleja magna by their much bigger soma sizes and higher numbers of primary dendrites. Cholinergic neurons of the shell were found to be slightly, but significantly, larger than those of the core that also subserves subdivision of the primate ACC into shell and core. A low proportion of tyrosine-hydroxylase immunostained cells, already previously described below the rostral ACC, co expressed CR but not calbindin. A CR-immunoreactive neuronal population, intermingled with these cells, extends as a stripe medially to the ACC along the septal part of corpus callosum into the lateral septal area. The presumed origin of CR-immunoreactive fibres in the shell of ACC is discussed. PMID- 10700609 TI - 6-Hydroxydopamine induced apoptosis of dopaminergic cells in the rat substantia nigra. AB - The pathologic hallmark of Parkinson's disease is the dopaminergic cell death in the substantia nigra (SN). The cause of the cell death is, however, unknown. Even the question on whether the cells die by apoptosis or necrosis has not been answered with certainty. In 6-Hydroxydopamine induced Parkinsonian rats, the present study observed apoptotic nuclei from 1 day to 14 days after lesioning, using the TdT(terminal deoxynucleotidyl transferase)-mediated dUTP-biotin nick end labeling method. Tyrosine hydroxylase immunohistochemistry and haematoxylin staining further revealed that these apoptotic cells are dopaminergic cells in the substantia nigra. The results suggest that dopaminergic cells in SN undergo apoptosis in the rat model of Parkinson's disease. PMID- 10700610 TI - The development of terminal Schwann cells associated with periodontal Ruffini endings in the rat incisor ligament. AB - The postnatal development of the terminal Schwann cell, an analogue of the lamellar cell in cutaneous sensory receptors, was examined by histochemistry for non-specific cholinesterase and immunohistochemistry for S-100 protein in the periodontal Ruffini endings of the rat incisor. Double immunohistochemistry for S 100 protein and protein gene product 9.5 (PGP 9.5) was also performed to examine the relationship between terminal Schwann cells and axons. Histochemistry for non specific cholinesterase was able to demonstrate the age-related development of the terminal Schwann cells; the morphology and distribution of the developing terminal Schwann cells became almost identical to those in adults during postnatal days 15-18. Axons showing PGP 9.5-like immunoreactivity elongated and expanded after arrangement of terminal Schwann cells in the alveolus-related part. This suggests that the terminal Schwann cell is important in the development and maturation of the periodontal Ruffini endings. PMID- 10700611 TI - Blood-brain barrier formation of grafted human umbilical vein endothelial cells in athymic mouse brain. AB - Human umbilical vein endothelial cells (HUVECs) were transplanted in athymic mouse brain and neovascularization of grafted endothelial cells was studied. HUVECs were transfected by a reporter gene pEGFPE-N1 in vitro and grafted stereotactically in unilateral striatum of adult nude mice. Histological studies in 4 weeks revealed that grafted HUVECs newly formed microvessels in brain, which were migrated and fused with host vessels. Intravenous injection of Evans blue before sacrificing animals resulted in no extravasation of dye, indicating that a blood-brain barrier (BBB) was formed by the grafted HUVECs. Immunohistochemistry demonstrated that host astrocytes extended glial feet on the grafted endothelial cells and a part of the newly formed vessels was positive with glucose transporter-1. These results indicate that endothelial cells from an ectopic origin have the potential to form a BBB after grafting in the central nervous system. PMID- 10700612 TI - Tactile stimulation activates dopamine release in the lateral septum. AB - Little is known about the functional properties of the dopamine innervation of the lateral septum. In this study, the feasibility of using microdialysis to assess action-potential mediated release of dopamine in the lateral septum was established. A mild stressor, in the form of handling, significantly increased septal dopamine levels, implicating a role for dopamine in sensory-related processing associated with the septal complex. PMID- 10700613 TI - Influence of the ventral hippocampal formation on plasma vasopressin, hypothalamic-pituitary-adrenal axis, and behavioral responses to novel acoustic stress. AB - The ventral hippocampal formation (vHF) seems to constrain diverse responses to psychological stimuli, and disruption of this function may underlie severe neuropsychiatric diseases. In particular, the ventral subiculum inhibits hypothalamic-pituitary-adrenal axis (HPA) activity following psychological, but not systemic, stressors. Despite the difficulty in interpreting such HPA responses, they have been relied upon to further characterize vHF function, because increased HPA axis activity is implicated in neuropsychiatric disturbances, and reliance on behavioral and cognitive data is even more problematic. Plasma arginine vasopressin (pAVP), which is inhibited by psychological stimuli and is also implicated in diverse neuropsychiatric diseases, provides a less ambiguous measure of CNS function. To test if its inhibition by psychological stress is also mediated by the vHF, we conducted two studies. In the first, pAVP and behavioral responses to novel acoustic stress were assessed in rats with bilateral excitotoxic lesions of the ventral subiculum and the ventral hippocampus. The subiculum lesions blocked the fall in pAVP and enhanced escape behaviors, whereas the hippocampal lesions produced responses intermediate to those in the subiculum-lesioned and control rats. In the second study, the pAVP response was similarly blocked by small lesions restricted to those vHF subfields which project to the neuroendocrine hypothalamus, compared to the response in animals with lesions in other vHF subfields. These results indicate that discrete projections from the vHF inhibit the pAVP response to psychological stimuli, and suggest that pAVP may provide a reliable probe of vHF activity. PMID- 10700614 TI - Synaptic deficit in the temporal cortex of partial trisomy 16 (Ts65Dn) mice. AB - Down syndrome results from triplication of human chromosome 21. The distal end of mouse chromosome 16 shares a large region of genetic homology with the Down syndrome 'critical region' of human chromosome 21. Therefore, a partially trisomic mouse (Ts65Dn) that possesses a triplication of the distal region of chromosome 16 has been developed as a putative model for Down syndrome. Ts65Dn mice display learning and memory deficits. However, despite the importance of preserved synaptic integrity for learning and memory, the ultrastructure of neural connectivity has not yet been studied in Ts65Dn mice. Therefore, the density and apposition zone length of synapses in the temporal cortex of aged Ts65Dn mice (n=4) were compared with those in diploid controls (n=4), using quantitative electron microscopy. There were significantly less (30%) asymmetric synapses in the temporal cortex of Ts65Dn mice than in controls (t=-5.067; p=0.023). However, there was no significant difference between the mean density of symmetric synapses in Ts65Dn mice and control mice. In addition, the mean synaptic apposition lengths of both asymmetric (15%; t=9.812, p<0.0001) and symmetric (11%; t=5. 582; p<0.0001) synapses were significantly larger in Ts65Dn mice than in controls. These results suggest that excitatory synapses are preferentially affected in Ts65Dn mice and that there is an attempt to compensate for the deficit of asymmetric synapses by increasing the contact zone area of existing synapses. The results may also reveal the morphological basis for the learning and memory deficits observed in Ts65Dn mice and have a bearing on the cognitive deficits in Down syndrome in old age. PMID- 10700615 TI - Glial and neuronal localization of ionotropic glutamate receptor subunit immunoreactivities in the median eminence of female rats: GluR2/3 and GluR6/7 colocalize with vimentin, not with glial fibrillary acidic protein (GFAP). AB - Female rat median eminence was immunostained with anti-NR1, GluR1, GluR2/3, GluR6/7, or KA2. GluR2/3- and GluR6/7-immunoreactivities were detected in cells lining the basal portion of the third ventricle. To identify these cells as tanycytes, the median eminence was dual-immunostained with glutamate receptors and glial cytoskeletal marker proteins, such as vimentin or glial fibrillary acidic protein (GFAP). Both GluR2/3 and GluR6/7 were shown to colocalize with vimentin, not with GFAP. These results suggest the potential role for tanycytes in conducting glutamate signaling. PMID- 10700616 TI - Regulation of retinal neurite growth by alterations in MAPK/ERK kinase (MEK) activity. AB - Activation of the extracellular-signal regulated kinase (ERK) cascade may be involved in the promotion of neurite outgrowth by a variety of stimuli. For example, we have previously shown that laminin (LN) and N-cadherin activate ERK2 in chick retinal neurons, and that pharmacological inhibition of MAPK/ERK kinase (MEK), the major upstream ERK2 activator, severely impairs neurite growth induced by these proteins. We have therefore hypothesized that ERK activation through MEK is required for optimal induction of neurite growth by these proteins. Here we show that expression of mutant MEK in transfected retinal neurons alters neuronal responses to LN in a manner consistent with this hypothesis. Neurons expressing a constitutively active MEK construct extended longer neurites on LN than controls, while neurons transfected with a dominant negative construct extended shorter neurites. Further, experiments in which transfected neurons were replated onto polylysine substrates suggest that activation of MEK is sufficient for neurite promotion on a non-inducing substrate, and neurons replated onto LN confirm the pharmacological data that inhibition of MEK activation inhibits LN-induced neurite growth. We conclude that ERK activation plays a direct role in the promotion of neurite outgrowth from retinal neurons by LN. PMID- 10700617 TI - Cloning and expression of the alpha9 nicotinic acetylcholine receptor subunit in cochlear hair cells of the chick. AB - Hair cells of the vertebrate inner ear are subject to efferent control by the release of acetylcholine (ACh) from brainstem neurons. While ACh ultimately causes the hair cell to hyperpolarize through the activation of small conductance Ca(2+)-activated K(+) channels, the initial effect is to open a ligand-gated cation channel that briefly depolarizes the hair cell. The hair cell's ligand gated cation channel has unusual pharmacology that is well matched to that of the nicotinic subunit alpha9 expressed in Xenopus oocytes. We used sequence-specific amplification to identify the ortholog of alpha9 in the chick's cochlea (basilar papilla). Chick alpha9 is 73% identical to rat alpha9 at the amino acid level. A second transcript was identified that differed by the loss of 132 base pairs coding for 44 amino acids near the putative ligand-binding site. RT-PCR on whole cochlear ducts suggested that this short variant is less abundant than the full length alpha9 mRNA. In situ hybridization revealed alpha9 mRNA in sensory hair cells of the chick cochlea. The pattern of expression was consistent with the efferent innervation pattern. The alpha9 label was strongest in short (outer) hair cells on which large calyciform efferent endings are found. Tall (inner) hair cells receiving little or no efferent innervation had substantially less label. The cochlear ganglion neurons were not labeled, consistent with the absence of axo-dendritic efferent innervation in birds. These findings suggest that alpha9 contributes to the ACh receptor of avian hair cells and supports the generality of this hypothesis among all vertebrates. PMID- 10700618 TI - Detection of feigned recognition memory impairment using the old/new effect of the event-related potential. AB - Twenty-four undergraduate university students with no known neurological disorders completed the Recognition Memory Test (Warrington, A., 1984. Recognition Memory Test manual. Windsor, Berkshire: NFER-Nelson.) while event related potentials (ERPs) were recorded. Twelve subjects were instructed to feign a recognition memory deficit (malingering group), while the remainder served as controls. The malingerers performed poorly on the test compared to the control group. The 'old/new effect', an ERP measure thought to reflect recognition memory processes, did not differ between the groups, indicating recognition of previously learned material in the malingering group despite poor test performance. The study also revealed a second, early, old/new effect, maximal at left frontal sites in the malingering relative to the control group, suggesting task-related processing differences between the two groups. These effects appear to be of potential value in the detection of malingering of cognitive impairment in the clinical situation. PMID- 10700619 TI - Auditory event-related potentials in poor readers. AB - Although poor readers (PR) are considered the major group among reading-disabled children, there are not event-related potentials (ERP) studies reported of PR on the subject. In this study, attentional and memory processes were studied in an auditory oddball task in PR and normal controls. ERP to auditory stimuli were recorded in 19 leads of the 10/20 system, using linked earlobes as references, in 20 normal children (10 female) and 20 PR (10 female) of the same age (10-12 years old). Two pure tones (1000 and 3000 Hz) were used in an oddball paradigm. No significant differences were observed in the amplitudes and latencies of N100 between the groups. However, N200 to frequent stimuli and P200 to both frequent and infrequent stimuli were of higher amplitude in poor readers than in normal children. There were no differences between groups in the latency and amplitude of P300. The results suggest that PR use more attentional resources in the components occurring before P300 to both frequent and infrequent stimuli than the normal children, and this finding is particularly marked for PR girls. PMID- 10700620 TI - Emotional responsivity during daily life: relationship to psychosocial functioning and ambulatory blood pressure. AB - Emotional responsivity refers to acute changes in affective states. This study examined the relationship of emotional responsivity during daily life with ambulatory blood pressure (ABP) and psychosocial functioning. Subjects were 162 employed men and women, aged 25-45 years. Subjects underwent 24-h ABP monitoring in which they completed a behavioral diary with each cuff inflation. On a separate day, subjects completed a psychometric test battery including measures of depression, trait anxiety, and social support. Emotional Responsivity, an index of negative emotional variability during waking hours, was operationalized as the standard deviation of each individual's negative emotions scores throughout the day. Individuals with high levels of emotional responsivity showed greater increases in ABP and heart rate (HR) associated with negative emotions. Emotionally responsive individuals also reported less satisfaction with social support and higher levels of perceived daily stress, trait anxiety, and depressive symptoms. These findings suggest that psychosocial traits that have been linked to cardiovascular disease may be associated with more marked cardiovascular activation occurring in response to negative emotions experienced throughout the day. PMID- 10700621 TI - Brainstem frequency-following response and simple motor reaction time. AB - Simple motor reaction times (RT) in humans show marked trial-to-trial variations. In the present study, a brief tone (400 Hz, 37.5 ms duration) that was the imperative stimulus in a RT paradigm evoked the brainstem frequency-following response (FFR). Horizontal and vertical montage FFRs were recorded to evaluate neural responses with putative origins in auditory nerve and central brainstem, respectively. The main question concerned the possible relationship between trial to-trial variations in RT speed and FFR response properties. The results showed a reliable pattern in which fast RT trials yielded larger amplitudes (relative to slow trials) in earlier milliseconds of the FFR, and slow RT trials yielded relatively larger amplitudes in later milliseconds of the response. These results support the conclusion that early processing in the auditory brainstem is not automatic and invariant. Rather, short-latency evoked potentials appear to reflect trial-to-trial variations related to events far removed from the first synapse of sensory coding, perhaps depending upon cortically mediated influences such as cognition or attention. PMID- 10700622 TI - Cardiac responses associated with affective processing of unpleasant film stimuli. AB - The autonomic basis of cardiac reactions to unpleasant film stimuli was investigated. Film clips depicting major surgery, threats of violence, and neutral material were presented to 46 subjects. Self-report measures of emotion were obtained, as well as heart rate, respiration rate, respiratory sinus arrhythmia, T-wave amplitude and skin conductance level. Resting vagal tone was estimated in a paced breathing task prior to film viewing. Spontaneous blink rate was also taken as a measure of visual engagement during film viewing. Coherent increases in sympathetic activation accompanied the film containing violent threats, whereas the surgery film yielded greater electrodermal activation, as well as heart rate deceleration and T-wave increase. These data support the hypothesis of differential autonomic response patterns to specifically unpleasant material. As compared with threat and neutral films, greater blink rate inhibition was observed during the surgery film. Individual differences in parasympathetic cardiac control measured at rest were able to discriminate cardiac response patterns during film viewing. PMID- 10700623 TI - Modulation of attentional inhibition by norepinephrine and cortisol after psychological stress. AB - Two of the most salient physiological responses to stress are increased norepinephrine (NE) and cortisol (CORT) activities. However, it is unclear how these neurochemical events affect cognition, especially attention. We examined the effects of mild psychological stress on selective attention, as assessed by the negative priming (NP) paradigm. Salivary measures of the stress hormone CORT and alpha-amylase (a correlate of NE) were assayed to probe the relationship between the stress response and attentional inhibition. Healthy subjects (N = 20) engaged in the attention task, which was then followed by 15 min of a stressful video game before a return to the attentional task. Baseline saliva samples were obtained before the experiment began, 1 min after the video-game stressor, and 20 min post-stress. Subjects showed a significant reduction in NP and a decrease in reaction time (RT) after the video game. Moreover, alpha-amylase levels increased significantly after the stressor, indicating the role of NE in the acute stress response. While CORT levels remained unchanged after stress, CORT correlated significantly with both NP scores and RT after the stressor. These results imply that mild psychological stress can significantly alter attentional processes. Given the increase in alpha-amylase and the correlation between attention and CORT after stress, it seems likely that attentional processes are under tight control by brain systems which mediate the fight-or-flight response. PMID- 10700624 TI - An early antecedent to modern random dot stereograms --'the secret stereoscopic writing' of Ramon y Cajal. AB - The use of computerized random dot stimuli in modern neuroscience was introduced by Julesz in the 1960s. This method made it possible to study exclusively cortical processing of binocular information by disparity-sensitive neurons, and it has attained widespread use among neuroscientists and psychologists. It is now largely forgotten that in the last century, the famous neuroanatomist Ramon y Cajal had worked on random dot stereograms as a means of encoding written information. A brief note was finally published in a Spanish journal on photography in 1901. We present a translation of this text and summarize the early ideas on random dot stereograms, and we also supply a brief historical account on stereoscopic perception. PMID- 10700625 TI - EEG activity during the performance of complex mental problems. AB - This study investigated differences in cognitive processes related to problem complexity. It was assumed that these differences would be reflected in respondents' EEG activity--spectral power and coherence. A second issue of the study was to compare differences between the lower (alpha(1) = 7.9-10.0 Hz), and upper alpha band (alpha(2) = 10.1-12.9 Hz). In the first experiment two well defined problems with two levels of complexity were used. Only minor differences in EEG power and coherence measures related to problem complexity were observed. In the second experiment divergent production problems resembling tasks on creativity tests were compared with dialectic problems calling for creative solutions. Differences in EEG power measures were mainly related to the form of problem presentation (figural/verbal). In contrast, coherence was related to the level of creativity needed to solve a problem. Noticeable increased intra- and interhemispheric cooperation between mainly the far distant brain regions was observed in the EEG activity of respondents while solving the dialectic problems. These results are explained by the more intense involvement of the long cortico cortical fiber system in creative thinking. Differences between the lower and upper alpha band were significant for the power and coherence measures. In Experiment 2, fewer differences were observed in power measures in the upper alpha band than in the lower alpha band. A reverse pattern was observed for the coherence measures. These results hint to a functional independence of the two alpha bands, however, they do not allow to draw firm conclusions about their functional meanings. The study showed that it is unlikely that individuals solve well- and ill-defined problems by employing similar cognitive strategies. PMID- 10700626 TI - An evaluation of mattresses and mats in two dairy units. AB - In order to investigate the relative merits of mats and mattresses in terms of cow comfort, production and performance, 29 cows were housed on ethylethene vinyl acetate (EVA) mats and 29 on mattresses of loose rubber crumb with a polypropylene cover, at each of two similar dairy units (SAC Auchincruive and Myerscough). Both mats and mattresses were newly installed at the start of the trial. The cows were housed in the autumn after calving. Milk yield was recorded daily. Cows were weighed and scored for body condition, locomotion, dirtiness and hock and knee injury at fortnightly intervals. Feed offered was recorded daily and refusals were weighed weekly. Monthly milk records of milk yield, milk composition and somatic cell count data were available for both herds. In addition, 24 h behavioural observations of 15 core cows in each group were made at weeks 0, 2, 4, 6, 8, 16, and 32 post-housing. There was no difference between cows on mats and mattresses in milk yield, composition or quality; in feed intake; in weight loss or body condition score; in severe hock or knee injury, or in the incidence of lameness. Cows on mattresses tended to have slightly higher total dirtiness scores than those on mats (7.06 vs. 6.95, P=0.074) and had dirtier udders (mattress, 7.50 vs. mat, 6.52, P<0.05). However, over the whole housing period, cows on mattresses spent longer feeding, ruminating and lying and a greater proportion of their lying time was spent ruminating. They spent less time standing doing nothing (idling) than cows on mats and less time idling in cubicles. Cows on mattresses appeared to adapt to housing more quickly than those on mats. Overall, neither mat nor mattress gave advantages in terms of production or performance, cows were slightly cleaner on mats but behavioural indices suggest that cow comfort was greater on mattresses. PMID- 10700627 TI - Behavioural response to humans and the productivity of commercial dairy cows. AB - This study examined the relationships between the attitude and the behaviour of the stockperson towards cows and the behavioural response to humans and the milk production of cows at 31 commercial dairy farms over one lactation. The attitude of the stockperson was measured on the basis of the stockperson's opinion of the cow's behaviour and how the stockperson evaluated his own behaviour. The behaviour of the stockperson was measured by recording the nature and frequency of the tactile interactions and some visual and auditory interactions directed towards the cows. The behavioural response of cows to humans was assessed by observing their approach behaviour to an unfamiliar experimenter in a standard test and production records were collected for the entire lactation at each farm. Correlation and regression analyses using farm averages were used to examine relationships between human and cow variables. Several cow behaviour variables, indicative of fear of humans, were moderately (P<0.05) to highly (P<0.01) correlated with milk yield and composition and regression analysis indicated that fear of humans accounted for 19% of the variation in milk yield between farms. The results suggest that at farms where milk yield was low, cows showed less approach to the experimenter in the standard fear test than at farms where milk yield was higher. A composite attitude score, based on the responses of stockpeople to questions about patting and talking to cows, ease of movement of cows and cows recognising unfamiliar stockpeople, was moderately (P<0.05) to highly (P<0.01) correlated with the behaviour of the stockperson. While a number of stockperson behaviour variables were correlated (P<0.05) with milk yield, the former variables were generally poorly correlated with cow behaviour. Therefore these preliminary findings provide evidence that, as seen in the pig industry, sequential relationships may exist between the attitude and behaviour of the stockperson and the behaviour and productivity of commercial dairy cows. Research is required to further examine these relationships because of the possible implications on cow productivity and welfare. PMID- 10700628 TI - A laser-based method for measuring thermal nociception of cattle. AB - We describe a method for measuring nociception in cattle using a CO(2) laser aimed at the caudal aspect of the metatarsi. In Experiment 1, infrared thermography showed that calves responded by lifting their legs when skin temperatures reached 45-55 degrees C. In Experiment 2a, the validity of the method was tested by comparing the response latencies of 14 calves to two power settings (2.25 W vs. 4.5 W) with each setting being applied six times. We found that both leg-lift latencies and tail-flick latencies were lower at the higher power setting, and the calves were more likely to respond by kicking than by simply moving the leg. The standard deviations between and within calves were smaller at the higher power setting, and the large within-calf variation means that at least three tests were required to obtain reliable measures that could differentiate between calves. In Experiment 2b, application of the laser at a range of power settings (2.0, 3.0, 4.0, 4.5, 5.0 and 5.5 W) on 16 calves showed that response latencies decreased as power increased up to 4.5 W, after which no further change occurred. In Experiment 3, the repeatability of the method was evaluated on nine measures with the high power setting (4.5 W). The coefficient of variation associated with repetition of the measures was 36%. In general, we found little change in response latencies with repeated use of the laser, except that responses on the second test tended to be shorter. Experiment 4 showed that ambient temperatures between 16 degrees C and 27 degrees C did not affect response latencies, but these were longer at temperatures of 7 degrees C. We suggest that the method is a useful way of measuring cattle's sensitivity to nociception as the animals need not be restrained and the distance to the animal need not be closely controlled. However, to obtain accurate, valid and reliable measures it is necessary to use a high power setting (4.5 W) and take at least three consecutive measures of the response latency. PMID- 10700629 TI - The behavioural, physiological and immunological responses of lambs from two rearing systems and two genotypes to exposure to humans. AB - The behavioural, physiological and immunological responses of lambs from two rearing systems and two genotypes to exposure to humans was assessed during and immediately after testing in an open-field arena. Ninety-six lambs of two genotypes (Scottish Blackface: BF and Texelx(Blue-faced LeicesterxScottish Blackface): T) were used. From birth to weaning one of two management regimes was applied: extensive (E), whereby animals were handled as little as possible or semi-intensive (I), in which lambs experienced a greater level of human exposure. Eight lambs from each of the four treatment groups received an antigenic challenge (Mycobacterium a. paratuberculosis) at 9 weeks of age to allow subsequent testing of immunological reactivity. At 1 and 3 weeks after weaning and 1 year later, lambs were tested in groups of four in a 4.5x4.5 m indoor arena, marked with gridlines at 0.75 m intervals. There were a number of occasions where testing revealed significant effects of genotype, management or their interaction, but in an approximately equal number of instances no significant effects of either genotype or management were observed. Genotype significantly influenced the number of squares occupied in the test arena over a 10-min period before the human entered (100.4 vs. 110.5; sed 2.70 for BF and T lambs, respectively, p<0.001). In relation to the number of new squares entered, there was a genotypexmanagement interaction: BFE lambs entered fewer squares than TE lambs but following semi-intensive management (I) BF lambs entered more squares than T lambs (p<0.05). When a human entered the arena after this 10-min period, while there was a gradual reduction in the number of animals which had not moved over the next 5 min, 66 animals had not moved within the allocated time. Also during this period, BF lambs stood facing the human for significantly longer than T lambs (p<0.05). At the time of arena testing, 12 lambs from each treatment group were fitted with heart-rate monitoring equipment. There were significant differences in heart rate in relation to period of testing, i.e. before (107.9) or after (112.3) the point at which the human entered the arena or when the lambs were walking in the presence of a moving human (126.3 b.p.m.; sed 2.15, p<0.001). When lambs were alone in the test arena, BF lambs had higher heart rates than T lambs (p<0.05). The heart rate of E lambs increased more than that of I lambs when the human entered the pen (9.4 vs. 0.3 b.p.m.; sed 3.95, respectively; p=0.05). Immediately following completion of the behavioural tests, blood samples were collected from subsets of lambs. Plasma cortisol concentrations of BF lambs were greater than those of T lambs (82.0 vs. 53.5 nmol/l; sed 10.18, p<0.01) but there was no effect of management. Blood samples collected from the lambs challenged with a novel antigen prior to weaning showed a genotype but not a management effect on both antibody and cell mediated immune responses, although there was a genotypexmanagement interaction. However, it should also be noted that there were no significant effects of either genotype or management on a number of the indices recorded: latency of lambs to move from the initial entry position in the absence or subsequent presence of a human; length of time one individual was separated from the other three; distance moved in a raceway before stopping; plasma beta-endorphin concentrations; heart rate in the presence of a human. Overall, these results suggest that although differences in responsiveness associated with specific genotypes of sheep can be detected in a test situation, the early life management regime may also have an effect. The results of this study caution against drawing conclusions between studies where different genotypes are employed. PMID- 10700630 TI - Cereal aversion in behaviourally resistant house mice in Birmingham, UK. AB - In 1986, house mice in a small defined area of inner Birmingham were reported as not taking a variety of rodenticides from bait containers, a phenomenon labelled 'behavioural resistance'. This study investigated behavioural resistance by comparing the food preferences of West Midlands behaviourally resistant (WMBR) mice with those of normal (BC) mice. Nine bait boxes each containing one of nine different foods (cheese, chicken, tuna fish, peanut butter, canary seed, Cat stars, wheat, PCD (MOD) pellets and Non-tox) were introduced to 12 WMBR and seven BC sites (Experiment 1). The experiment was repeated in the laboratory with six pens of WMBR and six of BC mice (Experiment 2), and to investigate whether the preferences had a genetic basis the offspring were similarly assayed (Experiment 3). In each experiment the consumption of each food was measured over 7 days and the droppings around the bait boxes were counted to assess mouse activity. Food neophobia was noted in some populations of BC mice. Tested in the wild and in the laboratory WMBR mice showed an aversion to foods containing cereals, as did their offspring. These results, with other lines of evidence, strongly suggest that cereal aversion in WMBR mice has a physiological/genetic basis. Since cereal aversion allows WMBR mice to survive cereal-based rodenticidal baits, we conclude that WMBR mice have genetically based behaviours that allow them to survive poisoning regimes that kill other strains. PMID- 10700631 TI - Neonatal handling of Amazon parrots alters the stress response and immune function. AB - The influence of neonatal handling on behavior and immune function was assessed in Orange-winged Amazon parrots (Amazona amazonica). Chicks (n=11) were gently handled daily from 25 days of age until 38 days post-fledging, while control chicks (n=9) were not handled. At 10 days post-fledging ( approximately 66 days of age), chicks were given tests to evaluate tameness (e.g., willingness to perch on an offered finger). They were then restrained for 10 min, either by being held while perching (handled group) or, because they would not perch, by being restrained in a towel (nonhandled group). Serum corticosterone levels were measured and immune status was assessed by: the delayed-type hypersensitivity (DTH) response to phytohemagglutinin-P (PH-P) injection; the humoral response to a killed Newcastle disease virus (NDV) challenge; and heterophil:lymphocyte ratio (H:L). Handled chicks were tamer by all measures of tameness. DTH was greater in nonhandled chicks (P or =13 days) with either a high initial dose of levothyroxine (> or =9.5 microg/kg/d) or a low initial dose (<9.5 microg/kg/d). With these criteria, 4 treatment groups were formed. The results of the Bayley test, performed at the age of 10 to 30 months and expressed as mental developmental index (MDI) and psychomotor developmental index (PDI), were related to socioeconomic status, treatment group, initial free thyroxine (FT(4)) concentration, and mean FT(4) concentration during the first 3 months of treatment (FT(4)-A) and the ensuing 9 months (FT(4)-B). RESULTS: Mean (+/- SD) MDI was 113 +/- 14, and mean PDI was 114 +/- 12. In the severe CH group, only the patients treated early with a high initial dose had normal MDI scores (124 +/- 16), whereas the scores of the other groups ranged from 97 to 103. In contrast, all patients in the mild CH group had normal scores (range, 122-125), except those in the group treated late with a low initial dose, whose score was 110 +/- 10. Forty-three percent of the variance in MDI and PDI scores was explained by treatment factors, such as the treatment group, initial FT(4) concentration, FT(4)-A, and FT(4)-B. CONCLUSIONS: Our data suggest that optimal treatment includes achievement of euthyroidism before the third week of life by initiation of therapy before 13 days with a levothyroxine dose above 9.5 microg/kg/d and maintenance of FT(4) concentrations in the upper normal range during the first year. Thus treated, patients with CH can achieve normal psychomotor development at 10 to 30 months, irrespective of the severity of the disease. PMID- 10700684 TI - A single dose of intramuscularly administered dexamethasone acetate is as effective as oral prednisone to treat asthma exacerbations in young children. AB - OBJECTIVE: To evaluate whether a single dose of intramuscularly administered dexamethasone acetate (IM Dex) was as safe and effective as a 5-day course of oral prednisone (PO Pred) in the treatment of young children with mild-moderate exacerbations of asthma. STUDY DESIGN: A prospective, randomized, investigator blinded study was done in a tertiary care medical center in children (6 months to 7 years of age) who required corticosteroids to treat mild-moderate asthma exacerbations as outpatients. Patients were randomized to receive either a single dose of IM Dex ( approximately 1.7 mg/kg) or PO Pred ( approximately 2 mg/kg/d for 5 days). Clinical asthma score, behavioral changes, albuterol use, and tolerance of the medication were recorded in a home diary for 7 days. Cortisol/creatinine ratios on first morning void urine samples were obtained on day 14. The primary outcome measures were changes in clinical asthma score through day 5 and tolerance of the medication. RESULTS: Fifteen patients in the IM Dex group (mean age 37 months) and 17 in the PO Pred group (mean age 36 months) completed the study. Clinical asthma score improved significantly in both groups during the first 5 days of therapy, and no significant difference was seen in the rate of improvement between the 2 groups. Three children refused more than 75% of their prednisone doses, and another 4 missed 30% to 50% of the doses despite their parents' best efforts. The intramuscular injection caused no complications, and approximately 70% of parents in both groups stated that they would choose IM Dex to treat their child's next asthma exacerbation. CONCLUSION: In this group of children a single intramuscular injection of dexamethasone acetate was as effective as a 5-day course of PO Pred for the management of mild moderate outpatient asthma exacerbations. PMID- 10700685 TI - A randomized controlled trial of a 3-year home exercise program in cystic fibrosis. AB - OBJECTIVES: To evaluate the effects of a 3-year home exercise program on pulmonary function and exercise tolerance in mildly to moderately impaired patients with cystic fibrosis (CF) and to assess whether regular aerobic exercise is a realistic treatment option. STUDY DESIGN: Seventy-two patients with CF (7-19 years) were randomly assigned to an exercise group (a minimum of 20 minutes of aerobic exercise, at a heart rate of approximately 150 beats/min, 3 times weekly) or a control group (usual physical activity participation). Pulmonary function, exercise tolerance, clinical status, hospitalizations, and compliance with therapy were monitored during scheduled visits to the hospital's CF clinic. RESULTS: Sixty-five patients were included in the analyses. The control group demonstrated a greater annual decline in percent of predicted forced vital capacity compared with the exercise group (mean slope +/- SD, -2.42 +/- 4.15 vs 0.25 +/- 2.81; P =.02), with a similar trend for forced expiratory volume in 1 second (-3.47 +/- 4.93 vs -1.46 +/- 3. 55; P =.07). Patients remained compliant with the exercise program over the study period. An improved sense of well-being was reported with exercise. CONCLUSIONS: Pulmonary function declined more slowly in the exercise group than in the control group, suggesting a benefit for patients with CF participating in regular aerobic exercise. Consistent compliance with the home exercise program and a self-reported positive attitude toward exercise provide further evidence of the feasibility and value of including an aerobic exercise program in the conventional treatment regimen of patients with CF. PMID- 10700686 TI - Longitudinal evaluation of cardiopulmonary performance during exercise after bone marrow transplantation in children. AB - OBJECTIVE: Abnormalities in cardiopulmonary performance during exercise have been reported in children after bone marrow transplantation (BMT). We sought to study changes in exercise performance over time in pediatric BMT survivors. STUDY DESIGN: We retrospectively reviewed the results of serial cardiopulmonary exercise tests performed by patients who had undergone BMT at our institution. Four measurements of cardiopulmonary function are reported: maximum cardiac index (MCI), maximal oxygen consumption (Max VO(2)), oxygen consumption at ventilatory threshold (VO(2) at VT), and maximum work (Max Work) performed. A linear mixed effects model was fitted to assess changes in these parameters over time. RESULTS: Thirty-three patients performed 96 cardiopulmonary exercise tests. MCI and VO(2) at VT were depressed at initial testing and did not change over time. Max VO(2) increased by 4% per year to 69% predicted, and Max Work increased to 77% predicted at 6 years after BMT. CONCLUSIONS: In spite of an impaired cardiovascular response to exercise as indicated by the persistently low MCI, aerobic and physical working capacity increase. Improved Max VO(2) suggests that oxygen extraction at the musculoskeletal level becomes more efficient with recovery from BMT. This may represent a compensatory response to an impaired ability to increase cardiac output. PMID- 10700687 TI - Obstructive sleep apnea in infants: relation to family history of sudden infant death syndrome, apparent life-threatening events, and obstructive sleep apnea. AB - OBJECTIVES: Familial aggregation of obstructive sleep apnea (OSA) has been shown to be associated with sudden infant death syndrome (SIDS) and apparent life threatening events (ALTE) in infants. We wanted to determine the incidence of OSA in infants with siblings with ALTE and SIDS referred to our sleep clinic and to ascertain whether OSA was more common in infants who have family histories of SIDS, ALTE, and OSA. STUDY DESIGN: We studied 125 infants (mean age, 11.5 +/- 0.6 weeks) who were separated into 2 groups on the basis of their family history; polysomnographic studies were performed on each infant. RESULTS: Twenty infants had a multiple family history of SIDS, ALTE, or OSA (group 1), whereas the other 105 infants (group 2) had only one case of SIDS or ALTE within the family and no known history of OSA. We found that 19 of 20 infants in group 1 had OSA, whereas only 31 of 105 infants in group 2 had OSA (chi-squared analysis, P <.05). The OSA recorded was more frequent in infants of group 1 than in those of group 2. Follow up studies in some infants with OSA demonstrated a progressive decrease in OSA, which resolved between 6 and 12 months of age. CONCLUSION: We conclude that infants of families with multiple histories of SIDS, ALTE, and OSA are more likely to have OSA than infants of families with only one case of SIDS or ALTE. PMID- 10700688 TI - Birth weight and altitude: a study in Peruvian communities. AB - We tested the hypothesis that at high altitude birth weight decreases once a critical barometric pressure (Pb) is reached. Birth weight data covering the 1 year period from November 1997 to October 1998 were collected in Peru from the data files of 15 community and mining centers between sea level and 4575 m altitude. These centers are scattered along the main road that joins Lima (on the Pacific shore) to Cerro de Pasco (4330 m) and surroundings. Above approximately 2000 m (ie, at Pb below approximately 590 mm Hg, inspired O(2) partial pressure of approximately 114 mm Hg) and up to approximately 4500 m altitude birth weight declined at an average of 65 g for every additional 500 m altitude (or 105 g for every additional 50 mm Hg drop in Pb). This pattern did not differ between sexes. Averages and modal distributions of the birth weight from 2 hospitals in Cerro de Pasco (4330 m) serving different social groups were similar. Body length at birth was similar at various altitudes, with the exception of the 2 highest locations above 4500 m, where it was slightly reduced. From these data, together with additional data collected in the North of Peru (Chacas, 3360 m) and with results from other ethnic groups previously published, we conclude that the drop in birth weight at altitude is (1) apparent once the critical Pb of approximately 590 mm Hg is reached, corresponding to an altitude of approximately 2000 m, (2) proportional to the increase in altitude between approximately 2000 m and 4500 m, and (3) independent from socioeconomic factors. PMID- 10700689 TI - Prophylactic indomethacin: factors determining permanent ductus arteriosus closure. AB - BACKGROUND: Permanent closure of the ductus arteriosus (DA) requires both effective muscular constriction to block luminal blood flow and anatomic remodeling to prevent later reopening. OBJECTIVE: We examined the role of prophylactic indomethacin in producing permanent DA closure and the mechanism by which this occurs. METHODS: We studied 2 separate approaches to managing a patent DA in 257 preterm infants (gestation 24 to 27 weeks): (1) prophylactic indomethacin (all infants treated during the first 15 hours after birth) or (2) symptomatic treatment (infants in this group received indomethacin only if clinical symptoms appeared; infants whose ductus closed spontaneously and never received indomethacin were included in this group). Echocardiography was performed 24 to 36 hours after the last dose of indomethacin was administered or by age 5 days if spontaneous closure occurred. Infants were monitored for the development of ductus reopening. RESULTS: The prophylactic treatment group had a greater degree of initial ductus constriction, a higher rate of permanent anatomic closure, and a decreased need for surgical ligation than did the symptomatic treatment group. The degree of initial ductus constriction was the most important factor determining the rate of ductus reopening. Post-treatment echocardiography proved to be the best test for predicting eventual reopening. CONCLUSION: Prophylactic indomethacin improved the rate of permanent ductus closure by increasing the degree of initial constriction. Prophylactic indomethacin did not affect the remodeling process, nor did it alter the inverse relationship between infant maturity and subsequent reopening. Even when managed with prophylactic indomethacin, the rate of ductus reopening remained unacceptably high in the most immature infants. PMID- 10700690 TI - Neurodevelopmental outcome at 3.5 years of age in children treated with extracorporeal life support: relationship to primary diagnosis. AB - OBJECTIVE: Recent studies suggest that for neonates treated with extracorporeal membrane oxygenation (ECMO), children with congenital diaphragmatic hernia (CDH) have poorer neurodevelopmental outcome than children with other diagnoses. We therefore analyzed the neurodevelopmental outcome at 3(1/2) years of age in 130 neonatal ECMO survivors with 6 different primary diagnoses. STUDY DESIGN: Children were assessed with the McCarthy Scales of Children's Abilities, Peabody Picture Vocabulary Test, Vineland Adaptive Behavior Scales, and a neurologic/physical examination; 12 factors related to infant characteristics and ECMO/hospital course including primary diagnosis were identified as independent variables. Dependent variables included test scores and 2 outcome categories: functional status (normal, risk, abnormal) and major neurologic sequelae (presence or absence). Statistical tools included chi-squared analysis, t test, analysis of variance, and discriminant and regression analysis. RESULTS: No significant differences were found between diagnostic groups in functional status or neurologic sequelae. Hospital days was the only variable consistently expressed in all analyses as having significant influence on the outcome measures. This was not a factor of the longer hospital days experienced by children with CDH. CONCLUSION: Neurodevelopmental outcome in neonatal ECMO is multifactorial. Although hospital days has the greatest association with outcome at age 3(1/2) years, these days likely reflect degree of illness and various complications that are independent of diagnostic group. Further study is required to determine which factors influencing the length of hospital stay may be the best predictor of long-term outcome. PMID- 10700691 TI - Resting energy expenditure and prediction equations in young children with failure to thrive. AB - OBJECTIVE: To compare predicted and measured resting energy expenditure (REE) in young children (birth to 3 years) with failure to thrive (FTT). METHODS: REE (kcal/d) was measured by indirect calorimetry and compared with predicted REE from 3 sex and age group equations: World Health Organization (WHO), Schofield weight-based (SCH-WT), and Schofield weight- and height-based (SCH-WT-HT). The clinical characteristics associated with inaccuracy of predicted REE were examined. RESULTS: Forty-five subjects (47% female) were evaluated. Their clinical characteristics (mean +/- SD) included age 1.2 +/- 0.7 years, length/height z score -2.1 +/- 1.3, weight z score -2.7 +/- 1.0, and measured REE 438 +/- 111 kcal/d. All prediction equations were within 10% accuracy <50% of the time. However, SCH-WT-HT did not significantly differ from measured REE (450 +/- 138 vs 438 +/- 111 kcal/d, P =.2) and was least likely to underestimate REE. Younger age and more severe growth failure (based on weight, length/height, or both) were associated with underestimation of REE by prediction equations. CONCLUSION: REE should be measured in young infants and children with moderate to severe FTT when knowledge of caloric needs is required for optimal clinical care. The SCH-WT-HT equation was least likely to underestimate REE and is therefore preferred when REE cannot be measured in this group of children. PMID- 10700692 TI - Pulmonary infiltrates with eosinophilia syndromes in children. AB - Pulmonary infiltrates with eosinophilia (PIE) are a group of heterogeneous disorders having the common findings of lung disease and eosinophilia in the peripheral blood, bronchoalveolar lavage fluid, or pulmonary interstitium. Eleven cases of PIE syndromes were identified through a retrospective and prospective chart review: drug-induced (2), acute eosinophilic pneumonia (3), infant pulmonary eosinophilia (2), parasite-induced (2), Churg-Strauss syndrome (1), and atypical chronic PIE (1). Patient demographics, clinical presentation, and disease severity varied considerably among groups. Therapeutic interventions included bronchodilators (10), oxygen (7), corticosteroids (9), and mechanical ventilation (3). A single patient with acute eosinophilic pneumonia died. Our experience suggests that PIE syndromes are rare in childhood and that clinical presentation can vary widely. Because of the potential for significant morbidity and mortality, aggressive diagnostic evaluations are warranted, particularly in children with respiratory failure of unknown etiology. PMID- 10700693 TI - "White coat hypertension" in adolescents: increased values of urinary cortisol and endothelin. AB - OBJECTIVE: To investigate whether "white coat hypertension" (WCH) in adolescents is an innocent phenomenon or is associated with early changes of the vascular system and/or increased stress response, reflected in the urinary endothelin and cortisol values, respectively. STUDY DESIGN: The study group included 36 subjects, 14 with WCH (8 males and 6 females) aged 12.9 +/- 3 years and 22 normotensive control subjects (12 males and 10 females) aged 13 +/- 3.5 years. WCH was defined as systolic and/or diastolic blood pressure (BP) > or =95th percentile for age, sex, and height and with reported normal BP measurements at home. Urinary endothelin (UET1), urinary free cortisol (UFC), and plasma renin levels were determined by radioimmunoassay; and urinary albumin levels were determined by nephelometry. For statistical analysis, the Mann Whitney U test, Spearman correlation coefficient, and multivariate analysis of variance/multivariate analysis of covariance were used, as applicable. RESULTS: The 24-hour values of UET1 and UFC were greater in male subjects with WCH than in male control subjects (P =.02), whereas no such difference was found in female subjects. The difference in UFC values in male subjects was accounted for by the day values. In subjects with WCH, and not in control subjects, a positive correlation of UET1 to UFC (r = 0.59, P =.027), diastolic BP (r = 0.55, P =.04), and mean BP (r = 0.65, P =.012) was detected. CONCLUSIONS: Our data indicate that WCH in adolescence may not be an innocent phenomenon and may represent a prelude to permanent idiopathic hypertension of adulthood. PMID- 10700694 TI - Childhood obesity in a population at high risk for type 2 diabetes. AB - OBJECTIVES: To determine the prevalence of obesity and investigate its association with fasting glucose and insulin among children and adolescents in a population at high risk for type 2 diabetes. DESIGN: A cross-sectional screening survey involving anthropometry and fasting serum levels of glucose and insulin. SETTING: A remote aboriginal (Ojibwa-Cree) community in northern Manitoba, Canada. PARTICIPANTS: All children aged 4 to 19 years in the community were invited to participate, with a response rate of 82% (n = 719). MAIN OUTCOME MEASURES: Obesity is defined as body mass index exceeding the 85th percentile of the National Center for Health Statistics reference data. The diagnosis of diabetes and impaired fasting glucose is based on the new criteria of the American Diabetes Association. RESULTS: There is a high prevalence of obesity, with 64% (female) and 60% (male) exceeding the 85th percentile and 40% (female) and 34% (male) exceeding the 95th percentile. Body mass index is a significant predictor of both glucose and insulin in both sexes, independent of age. Obese children are at increased risk of being classified as having diabetes or impaired fasting glucose (odds ratio 5.1, 95% CI 1.51, 17.0). CONCLUSIONS: The early onset of type 2 diabetes in childhood is increasingly observed in many populations. Childhood obesity is a strong risk factor. Early detection and intervention directed at obesity are potential strategies to avert the long-term consequences of type 2 diabetes. PMID- 10700695 TI - Effective therapy for severe Henoch-Schonlein purpura nephritis with prednisone and azathioprine: a clinical and histopathologic study. AB - OBJECTIVES: To validate a scoring system to assess the severity of renal lesions and to correlate histology with clinical findings. We also examined the efficacy of treatment with prednisone (1 to 2 mg/kg/d) and azathioprine (1 to 2 mg/kg/d) for severe Henoch-Schonlein purpura (HSP) nephritis. METHODS: Twenty patients were evaluated retrospectively. All underwent biopsy before treatment, and 13 underwent biopsy after therapy. We developed a scale based on glomerular, tubulointerstitial (TI), and vascular changes and assigned all specimens acuity, chronicity, and TI scores. The outcomes of 17 patients were compared with those of a historical control group. RESULTS: Chronicity score at initial biopsy increased with increasing delay between onset of renal involvement and first biopsy (rho = 0.55, P =.016) but did not progress after treatment was initiated. Both acuity (rho = 0.57,P =. 016) and TI (rho = 0.69, P =.003) scores correlated with clinical severity at first biopsy. The TI score correlated negatively with serum albumin (rho = -.60, P <.01). Significantly more patients in the study group than in the control group had a favorable outcome (15 [88%] of 17 vs 32 [54%] of 59, P =.011). CONCLUSIONS: Our scale reflects disease activity and highlights the importance of TI changes in severe HSP nephritis. Outcome comparisons indicate that early treatment with prednisone and azathioprine prevents progression of chronic changes and improves outcome. PMID- 10700696 TI - The impact of recombinant human growth hormone treatment during chronic renal insufficiency on renal transplant recipients. AB - OBJECTIVE: To evaluate post-transplant outcomes for patients treated with human growth hormone (rhGH) during the course of chronic renal insufficiency (CRI). STUDY DESIGN: Patients (the "cohort" group) were identified who had been enrolled in 2 controlled studies to determine the efficacy and safety of rhGH in growth retarded children with CRI and were subsequently enrolled in the North American Pediatric Renal Transplant Cooperative Study (NAPRTCS) and received a renal transplant. Patient survival, graft survival, time to first acute rejection episode, causes of graft failure, adverse events, and serial growth data from transplant to 60 months were evaluated. Data from the cohort group of 102 patients were compared with data from 4913 primary transplants from "other NAPRTCS" recipients (the "control" group). RESULTS: No significant difference was seen in patient survival or graft survival, incidence of acute rejection episode, or time to first rejection episode between the cohort and control groups. No specific adverse events were attributable to previous rhGH treatment. Only 2 patients had post-transplant lymphoproliferative disease in the cohort group, with no other malignancies reported. The mean height z scores in the cohort group at baseline and 60 months after transplant were -1.92 and -1.90, and the Deltaz score at 60 months was +0.20 compared with the control group (-1.88 and -2.10). CONCLUSIONS: Treatment of growth-retarded patients with CRI does not adversely affect graft function after renal transplantation. "Catch-down" growth does not occur after renal transplantation. PMID- 10700697 TI - Pediatric evidence-based medicine: past, present, and future. PMID- 10700698 TI - Overlap of dyskeratosis congenita with the Hoyeraal-Hreidarsson syndrome. AB - X-linked dyskeratosis congenita (DKC) is characterized by mucosal leukoplakia and ulcerations, skin abnormalities, nail dystrophy, and pancytopenia. Hoyeraal Hreidarsson syndrome (HHS) includes intrauterine growth retardation, microcephaly, mental retardation, cerebellar malformation, and pancytopenia. A patient with striking features of both HHS and DKC has a de novo mutation in the DKC1 gene, known to be responsible for DKC. HHS may be a severe form of DKC, in which affected individuals die before characteristic mucocutaneous features develop. PMID- 10700699 TI - Loss of hypoglycemia awareness in an adolescent with type 1 diabetes mellitus during treatment with fluoxetine hydrochloride. AB - A 17-year-old boy with type 1 diabetes mellitus developed new loss of hypoglycemia awareness while being treated with fluoxetine hydrochloride for depression. Hypoglycemia unawareness resolved after this medication was discontinued. PMID- 10700700 TI - Death caused by perioperative fasting and sedation in a child with unrecognized very long chain acyl-coenzyme A dehydrogenase deficiency. AB - An adopted 3(1/2)-year-old girl with no prior medical problems died after a routine dental procedure. More than 2 years later, acylcarnitine analysis of dried blood found on her bedding revealed she had very long chain acyl-coenzyme A dehydrogenase (VLCAD) deficiency. Perioperative oral fasting, without intravenous administration of glucose, may be detrimental to children with certain metabolic and endocrine disorders. Newborn screening by tandem mass spectrometry will detect disorders of fatty acid oxidation such as VLCAD and allow early and preventive treatment. PMID- 10700701 TI - Carbohydrate-deficient glycoprotein syndrome type 1a: a variant phenotype with borderline cognitive dysfunction, cerebellar hypoplasia, and coagulation disturbances. AB - An 8-year-old boy is described with borderline cognitive impairment, cerebellar hypoplasia, a stroke-like episode, and venous thrombosis of the left leg after a period of immobilization. The pattern of multiple abnormalities in blood coagulation suggested carbohydrate-deficient glycoprotein syndrome type 1a. Isoelectric focusing of serum transferrin was abnormal. The activity of phosphomannomutase in leukocytes and fibroblasts was decreased. Mutation analysis of the PMM2 gene revealed the R141H/E151G genotype. These results confirm the presence of carbohydrate-deficient glycoprotein syndrome type 1a without severe psychomotor retardation. PMID- 10700702 TI - Corrected QT interval (QTc) prolongation and syncope associated with pseudohypoparathyroidism and hypocalcemia. AB - An adolescent presented with exercise-associated syncope and electrocardiographic corrected QT interval (QTc) prolongation. Pseudohypoparathyroidism-induced hypocalcemia was diagnosed. The QTc (485 to 505 milliseconds) shortened during normalization of calcium levels, and syncope has not reoccurred. PMID- 10700703 TI - Hypoplastic or absent mandibular frenulum: a new predictive sign of infantile hypertrophic pyloric stenosis. AB - Among 25 patients with hypertrophic pyloric stenosis, a hypoplastic or absent mandibular frenulum was noted in 92%, compared with 1.6% of 319 control infants (P <.001). This previously unrecognized sign may prove helpful in identifying newborns at risk of developing the disorder. PMID- 10700704 TI - Survival and dominant transmission of "lethal" platyspondylic dwarfism of the "West coast" types. AB - Torrance, San Diego, and Luton types ("West coast" types) of neonatal platyspondylic short-limbed dwarfism are suspected to be caused by dominant mutations that are obligatorily lethal. We report on an affected mother, who passed the disease to her daughter, confirming dominant disease transmission. Survival of the mother indicates a wider phenotypic spectrum. PMID- 10700705 TI - Iniencephaly: an uncommon neural tube defect. PMID- 10700706 TI - Infantile liver hemangiomatosis: evidence for molecular heterogeneity. PMID- 10700707 TI - Why do steroids increase blood pressure in preterm infants? PMID- 10700709 TI - Reply PMID- 10700708 TI - Detection of periventricular leukomalacia. PMID- 10700710 TI - IgE cross-reactivity between human and cow's milk proteins in atopic breast-fed infants. PMID- 10700711 TI - Reply PMID- 10700712 TI - Unilateral fixed dilated pupil in an infant after inhalation of nebulized ipratropium bromide. PMID- 10700713 TI - The role of the GH-IGF-I axis in the regulation of myocardial growth: from experimental models to human evidence. PMID- 10700714 TI - Iron supplementation in goitrous, iron-deficient children improves their response to oral iodized oil. AB - OBJECTIVE: In developing countries, many children are at high risk for both goiter and iron-deficiency anemia. Because iron deficiency may impair thyroid metabolism, the aim of this study was to determine if iron supplementation improves the response to oral iodine in goitrous, iron-deficient anemic children. DESIGN: A trial of oral iodized oil followed by oral iron supplementation in an area of endemic goiter in the western Ivory Coast. METHODS: Goitrous, iodine deficient children (aged 6-12 years; n=109) were divided into two groups: Group 1 consisted of goitrous children who were not anemic; Group 2 consisted of goitrous children who were iron-deficient anemic. Both groups were given 200mg oral iodine as iodized oil. Thyroid gland volume using ultrasound, urinary iodine concentration (UI), serum thyroxine (T(4)) and whole blood TSH were measured at baseline, and at 1, 5, 10, 15 and 30 weeks post intervention. Beginning at 30 weeks, the anemic group was given 60mg oral iron as ferrous sulfate four times/week for 12 weeks. At 50 and 65 weeks after oral iodine (8 and 23 weeks after completing iron supplementation), UI, TSH, T(4) and thyroid volume were remeasured. RESULTS: The prevalence of goiter at 30 weeks after oral iodine in Groups 1 and 2 was 12% and 64% respectively. Mean percent change in thyroid volume compared with baseline at 30 weeks in Groups 1 and 2 was -45.1% and -21.8% respectively (P<0.001 between groups). After iron supplementation in Group 2, there was a further decrease in mean thyroid volume from baseline in the anemic children (-34.8% and -38.4% at 50 and 65 weeks) and goiter prevalence fell to 31% and 20% at 50 and 65 weeks. CONCLUSION: Iron supplementation may improve the efficacy of oral iodized oil in goitrous children with iron-deficiency anemia. PMID- 10700715 TI - Thyroid structure and size and two-year follow-up of solitary cold thyroid nodules in an unselected population with borderline iodine deficiency. AB - OBJECTIVE: Multinodular goitre has been found with a high prevalence in iodine deficient areas, but less frequently in iodine-replete areas; the iodine intake sufficient to prevent goitre has not been established, however. METHODS: We report data from an ultrasonic investigation of the thyroid glands of 2656 randomly selected subjects aged 41 to 71 years in an area with borderline iodine deficiency. RESULTS: Median iodine concentration in spot urine samples was 70microg/l. Multinodular thyroid structure was found in 23% of the population, increasing in women from 20 to 46% with increasing age, and in men from 7 to 23%. Solitary, scintigraphically cold, thyroid nodules >10mm were found in 2.4% of the population with the same prevalence in the different age and sex groups. Two years of follow-up of these cold nodules revealed no signs of malignancies. Median thyroid volume was 11.0ml. Thyroid enlargement (>18ml for women and >25ml for men) was found among 13. 1% of the women and 6.2% of the men, and the prevalence increased with age. The presence of thyroid nodules was related to positive anti-thyroperoxidase antibody (TPO Ab) titres, whereas thyroid enlargement was associated with iodine excretion <50microg/day. CONCLUSIONS: Thyroid enlargement was associated with low iodine excretion and median thyroid volume was slightly increased compared with iodine-replete areas. Multinodular thyroid structure was found with a high prevalence and was associated with TPO Ab >200kU/l. Cold thyroid nodules were moderately prevalent, with no cases of detected malignancies during 2 years of follow-up. PMID- 10700716 TI - The effect of thyrotoxicosis on adrenocortical reserve. AB - OBJECTIVE: Variations in thyroid function are known to be associated with changes in adrenocortical activity. Previous studies in animals have suggested that long standing hyperthyroidism may be associated with diminished adrenal functional reserve despite a continuing hyperactivity of the hypothalamic-pituitary-adrenal (HPA) axis. In humans, there has been no direct assessment of adrenal secretory reserve in clinical thyrotoxicosis. This study aimed to assess adrenocortical reserve in response to low-dose ACTH, following dexamethasone suppression, in patients with severe thyrotoxicosis. DESIGN AND METHODS: Ten patients (four men and six women, 30-45 years) with severe long-standing thyrotoxicosis due to Graves' disease (n=6) or toxic nodular goitre (n=4) were studied at diagnosis and again when in a stable euthyroid state following drug therapy for 8-12 months. All patients underwent ACTH stimulation tests at 0800h with ACTH(1-24) (Cortrosyn; 0.1microg/kg body weight, i.v.) following overnight suppression of the HPA axis with dexamethasone (1mg per os at 2300h). Serum cortisol was assayed at -15, 0, 15, 30, 60 and 90min after the administration of ACTH. RESULTS: The mean (+/-s.d.) peak and delta cortisol responses to ACTH (634.5+/-164nmol/l and 618+/- 196nmol/l respectively), as well as the net area under the response curve (36769+/-12188nmol/lx min) in the hyperthyroid patients were significantly lower compared with the values when the same patients were euthyroid (911+/-157nmol/l, 905+/-160nmol/l and 57652+/-10128nmol/lxmin respectively; P<0.005). Subnormal peak cortisol responses (<500nmol/l) were observed in two severely toxic patients. The findings were independent of the cause of thyrotoxicosis. CONCLUSION: In patients with severe thyrotoxicosis, cortisol secretion in response to low-dose ACTH stimulation, following dexamethasone suppression, is lower in the hyperthyroid than in the euthyroid state. It appears that thyrotoxicosis is associated with subtle impairment of adrenocortical reserve. PMID- 10700717 TI - The role of cytokines and cortisol in the non-thyroidal illness syndrome following acute myocardial infarction. AB - OBJECTIVE: A number of different hormone changes have been described during the acute myocardial infarction (AMI), including those of the non-thyroidal illness syndrome (NTIS). DESIGN AND METHODS: We assessed the alterations of serum thyroid hormones, cytokines and cortisol levels in 30 patients with a first episode of AMI 4, 24, 48h and 10 days (240h) after the onset of the chest pain and we investigated the possible relationship of these alterations with the severity of AMI. RESULTS: Fifteen patients had left ventricular ejection fraction (LVEF) 50% (group II). A transient decrease of total tri-iodothyronine (T(3)), more prominent in group I (P<0.05, t-test) with a concomitant rise of reverse T(3 )(rT(3)) occurred at 24h. Total thyroxine (T(4)), free T(4) (FT(4)) and free T(4) index did not change significantly, but tended to be higher in group I patients, whereas TSH significantly increased in group II at 48h. Interleukin-6 (IL-6) increased significantly at 24h only in group I and declined thereafter (24 vs 240h, P<0.001) and this temporal change of IL-6 was associated with similar changes of creatine phosphokinase and creatine kinase isoenzyme MB (CK-MB). Tumor necrosis factor-alpha and IL-1beta remained low in both groups. Cortisol was higher at 4h and in 12 patients was above the normal values. Negative correlation was found between LVEF and IL-6 (P<0. 001), whereas T(3), T(4) or cortisol levels were not correlated with the LVEF. CONCLUSIONS: Our data indicate that NTIS, in association with increase of IL-6, occurs in the early post-infarction period. In the NTIS following AMI the high level of IL-6 is the best predictor, among several parameters, of the severity of AMI as assessed by the LVEF and the rise of CK-MB. PMID- 10700718 TI - Diagnostic efficiency of serum IGF-I, IGF-binding protein-3 (IGFBP-3), IGF I/IGFBP-3 molar ratio and urinary GH measurements in the diagnosis of adult GH deficiency: importance of an appropriate reference population. AB - OBJECTIVE: To analyse the diagnostic role of serum IGF-I, IGF-binding protein-3 (IGFBP-3), IGF-I/IGFBP-3 molar ratio and urinary GH (uGH) excretion in adult GH deficiency (GHD). DESIGN: Twenty-seven adults (age range: 18-71 years) with severe GHD, defined by a peak GH response to an insulin tolerance test below 3microg/l in patients with at least one additional pituitary hypofunction. Reference values were established from a selected age- and body mass index matched population (154 healthy adults grouped in four age groups). METHODS: IGF I and IGFBP-3 were measured by RIA (Nichols) and results expressed as standard deviation (s.d.) scores from our reference population and assay normative data (s.d. score Nichols). uGH was measured by IRMA. RESULTS: Within the control group, IGF-I, IGFBP-3, IGF-I/IGFBP-3 ratio standardisation regarding our control population and IGF-I with respect to the assay normative data resulted in disappearance of age-related differences. However, IGFBP-3 s.d. score Nichols resulted in mean values between +1.4 and +2.5 s.d. score. Greatest diagnostic efficiency was for IGF-I standardised with respect to our controls (97.2%), followed by s.d. score IGFBP-3 (92.9%). s.d. score IGF/IGFBP-3 ratio and uGH showed poor diagnostic efficiency. Any combination of at least two abnormal parameters raised specificity to 100%. IGF-I standardised with respect to assay reference (s.d. score Nichols) showed similar diagnostic value (95.0%) whereas IGFBP-3 showed low sensitivity (33. 3%). Within the GHD patients, those with three or more additional deficiencies had lower s.d. score IGF-I than those with only two or one. CONCLUSION: We underline the importance of an appropriate reference population for correct interpretation of GH secretion markers. Considering our results, specificity obtained with two simultaneous abnormal parameters when referred to an adequate reference population may add valuable information to alternative GH stimulation tests to confirm adult GHD. PMID- 10700719 TI - The effect of GH replacement therapy on endothelial function and oxidative stress in adult growth hormone deficiency. AB - OBJECTIVES: Controversy persists with regard to the atherogenic risk associated with adult growth hormone deficiency (GHD). Endothelial dysfunction and enhanced oxidative stress are early features of atherogenesis. Therefore, we have studied the effect of three months of low dose GH replacement therapy (0.03IU/kg/day) on these parameters in GHD adults. SUBJECTS AND METHODS: Eight hypopituitary GHD adults (4 male, 4 female), who were receiving conventional hormone replacement therapy, were studied before and after 3 months of GH replacement (0.03IU/kg/day). All observations obtained were compared with similar measurements made in 8 matched control subjects. All study subjects were non smokers, normotensive and gave no personal or family history of premature vascular disease. Endothelial function was assessed using a specialised vessel wall tracking system to measure endothelium-dependent, flow-mediated, brachial artery dilatation (FMD). Measurements were repeated following glyceryl-trinitrate (GTN) (endothelium-independent dilatation). Oxidative stress was assessed by directly measuring lipid-derived free radicals in venous blood by electron paramagnetic resonance spectroscopy. Fasting lipids, insulin, plasma glucose and IGF-I were also measured at baseline and following GH replacement. RESULTS: FMD, expressed as a percentage change from resting base-line diameter, was significantly impaired in the pre-treatment GHD patients compared with controls (3.1+/-2.1% vs 6.1+/-0.9%, P<0. 001; means+/-s.d.) indicating endothelial dysfunction. Significant increase in FMD was noted following GH therapy (3.1+/ 2.1% vs 6. 5+/-1.9%, P<0.001). Free radicals (arbitrary units) were elevated in the pre-treatment GHD patients compared with controls (0.36+/-0.09 vs 0.11+/ 0.12, P<0.05) and fell significantly following GH therapy (0.23+/-0.03 vs 0.36+/ 0.09, P<0.05), although they remained elevated compared with controls. Fasting insulin was significantly higher (25.9+/-18.8 vs 13.9+/-6.7mu/l, P<0.05) and IGF I concentrations lower (10.8+/-4.7 vs 20.2+/-6.3nmol/l, P<0.05) in the pre treatment GHD subjects. After treatment there were no changes in insulin concentration, although IGF-I levels were normalised (10. 8+/-2.3 vs 23.6+/ 11.4nmol/l, P<0.05). CONCLUSIONS: Endothelial dysfunction and enhanced oxidative stress are features of adult GHD. This study suggests plausible mechanisms underlying any proatherogenic tendency in adult GHD and demonstrates improvement of these factors following GH replacement. PMID- 10700720 TI - Serum leptin and habitual fatty acid dietary intake in patients with type 1 diabetes mellitus. AB - OBJECTIVE: To study the contribution of a normal intake of nutrients to the variability of serum leptin concentrations in persons with type 1 diabetes mellitus. DESIGN: We studied the relation between serum leptin and nutrient intake in a cross-sectional study. METHODS: Serum leptin measured by radioimmunoassay, nutritional data determined by a self-administered 7-day nutritional questionnaire, and the fatty acid composition of the serum phospholipids (measured by thin layer chromatography and gas chromatography) were determined in 60 patients with type 1 diabetes mellitus. Correlation and regression analyses were performed between serum leptin and dietary fatty acids and serum phospholipid fatty acids. RESULTS: In the prediction models for the concentrations of serum leptin in men with type 1 diabetes mellitus, the dietary fatty acids displaced the anthropometric variables, and were independent of the serum testosterone concentrations. This fact remained when the prediction was made on the basis of indirect markers of the intake, such as the serum phospholipid fatty acids. In the women, the fatty acids from the diet or from the serum phospholipids also partly explained the variation in serum leptin, although not displacing the anthropometric variables. CONCLUSIONS: Our data suggest that, in non-experimental conditions, the concentrations of serum leptin in men with type 1 diabetes mellitus and, to a lesser extent, those in women with diabetes, may be influenced by the composition of the habitual diet, especially the type of dietary fat. PMID- 10700721 TI - Allopregnanolone concentrations and premenstrual syndrome. AB - OBJECTIVE: To evaluate basal allopregnanolone and progesterone in both phases of the menstrual cycle in women suffering from premenstrual syndrome (PMS) and their response to a GnRH test. DESIGN: We selected 56 women (28 patients with PMS and 28 controls) aged between 18 and 32 years. Blood samples were drawn in both follicular and phases. Twenty-eight women (14 patients with PMS and 14 controls) underwent a GnRH test in the luteal phase. METHODS: We evaluated allopregnanolone by RIA, using a specific antibody. Serum progesterone and oestradiol were determined using a commercially available RIA kit. RESULTS: Luteal phase allopregnanolone concentrations were significantly lower in patients with PMS than in controls. Progesterone concentrations were significantly lower in patients with PMS in both the follicular and the luteal phase. Serum oestradiol concentrations were in the normal range in both groups of women, although slightly greater in those with PMS. Allopregnanolone and progesterone responses to a GnRH test were significantly blunted in women with PMS. CONCLUSIONS: Diminished concentrations of allopregnanolone and progesterone, its precursor, and a blunted response to the GnRH test lead us to hypothesise that patients with PMS may suffer from an inadequate production of ovarian neuroactive steroids, especially in the luteal phase. This would lead to an impaired anxiolytic GABA(A) mediated response in stressful physiological and psychological conditions, and may in part explain various psychoneuroendocrine symptoms that arise during PMS. PMID- 10700722 TI - Mechanism for the development of ovarian cysts in patients with congenital lipoid adrenal hyperplasia. AB - OBJECTIVE: Although ovarian cysts commonly occur in patients with congenital lipoid adrenal hyperplasia (CLAH), the mechanism of development remains to be determined. To clarify the pathogenesis of the ovarian cysts, endocrinological examinations were performed in patients with CLAH. METHODS: The subjects were three Japanese CLAH patients. Basal body temperature, serum and urinary gonadotropin levels, serum and/or urinary ovarian hormones and mutations of the steroidogenic acute regulatory protein (StAR) gene were examined. RESULTS: The basal body temperature was not biphasic in any patient. Basal LH levels were high in all CLAH patients and markedly responded to LH-releasing hormone in two patients. Urinary gonadotropin analysis revealed repetitive LH surges in the menstrual cycles of the CLAH patients. No increase in the urinary pregnanediol suggested anovulation in all patients, and bilateral ovarian cysts were found in two of the subjects. Examination of the StAR gene revealed a frameshift mutation 840delA at codon 238, a nonsense mutation Q258X at codon 258, a homozygotic mutation at Q258X, and a compound heterozygotic mutation with 251insG and Q258X. CONCLUSIONS: We concluded that the development of ovarian cysts may be derived from continued anovulation in CLAH patients. Elevated LH levels may be explained by increased sensitivity of the anterior pituitary to circulating estrogen. PMID- 10700723 TI - Hypothalamic amenorrhea with normal body weight: ACTH, allopregnanolone and cortisol responses to corticotropin-releasing hormone test. AB - OBJECTIVE: Hypothalamic amenorrhea (HA) is a functional disorder caused by disturbances in gonadotropin-releasing hormone (GnRH) pulsatility. The mechanism by which stress alters GnRH release is not well known. Recently, the role of corticotropin-releasing hormone (CRH) and neurosteroids in the pathophysiology of HA has been considered. The aim of the present study was to explore further the role of the hypothalamic-pituitary-adrenal axis in HA. DESIGN: We included 8 patients (aged 23.16+/-1.72 years) suffering from hypothalamic stress-related amenorrhea with normal body weight and 8 age-matched healthy controls in the follicular phase of the menstrual cycle. METHODS: We measured basal serum levels of FSH, LH, and estradiol and evaluated ACTH, allopregnanolone and cortisol responses to CRH test in both HA patients and healthy women. RESULTS: Serum basal levels of FSH, LH, and estradiol as well as basal levels of allopregnanolone were significantly lower in HA patients than in controls (P<0.001) while basal ACTH and cortisol levels were significantly higher in amenorrheic patients with respect to controls (P<0.001). The response (area under the curve) of ACTH, allopregnanolone and cortisol to CRH was significantly lower in amenorrheic women compared with controls (P<0.001, P<0.05, P<0.05 respectively). CONCLUSIONS: In conclusion, women with HA, despite the high ACTH and cortisol levels and, therefore, hypothalamus-pituitary-adrenal axis hyperactivity, are characterized by low allopregnanolone basal levels, deriving from an impairment of both adrenal and ovarian synthesis. The blunted ACTH, allopregnanolone and cortisol responses to CRH indicate that, in hypothalamic amenorrhea, there is a reduced sensitivity and expression of CRH receptor. These results open new perspectives on the role of neurosteroids in the pathogenesis of hypothalamic amenorrhea. PMID- 10700724 TI - Association between the expression of E1A oncogene and increased sensitivity to growth inhibition induced by sustained levels of cAMP in rat thyroid cells. AB - OBJECTIVE: The aim of this study was to investigate: (i) whether a persistent increase of cAMP interferes with the proliferation of transformed thyroid cells, and (ii) whether the degree of malignancy is correlated with the sensitivity to a transient and/or sustained increase in intracellular cAMP levels. DESIGN AND METHODS: To address these questions we used thyroid cell lines transformed with E1A oncogene from adenoviruses 5 (PC E1A cell line) or 2 (PC HE4 cell line), or infected with the polyoma murine leukemia virus (PC PyMLV cell line) carrying the middle T gene of the polyoma virus, or, finally, expressing both E1A and PyMLV. These cell lines present various degrees of malignancy: PC EIA and PC HE4 cells are not tumorigenic; PC PyMLV cells induce non-invasive tumors after a long latency period; and PC EIA+PyMLV cells are highly tumorigenic. RESULTS AND CONCLUSIONS: Thyroid cell proliferation required the transient increase of intracellular cAMP levels, while persistent elevation of cAMP blocked the proliferation of normal thyroid PC Cl 3 cells and of PC Cl 3 cells transformed by a variety of different oncogenes. In addition, sustained levels of cAMP induced apoptosis in cells carrying the adenovirus EIA oncogene, but not in cells transformed with other oncogenes or in the wild-type PC Cl 3 cells. Furthermore, middle T gene of the polyoma virus seemed to afford protection only from apoptosis induced by cAMP when middle T is present in thyroid cells along with the E1A gene. PMID- 10700725 TI - Steroidogenic acute regulatory protein mRNA expression in adrenal tumours. AB - The rate limiting step in steroidogenesis is cholesterol transport through the outer to the inner mitochondrial membrane and the cytochrome P450 side chain cleavage (P450scc) complex. The protein factor responsible for this transport, and as such necessary for regulating the acute production of steroids, has been identified and named the steroidogenic acute regulatory protein (StAR). We investigated the expression of StAR in functional and non-functional adrenal neoplasms and compared the expression with that of P450scc. Poly A RNA was extracted from normal adrenal glands (NAG, n=5), aldosterone producing adenomas (APA, n=4), cortisol producing adenomas (CPA, n=5), adrenocortical carcinomas (ACC, n=6) and non-functional adenomas (NFA, n=3), electrophoresed through a 1% agarose gel, blotted and hybridised with a PCR-generated cDNA labelled with [(32)P]CTP. The blots were stripped and re-hybridised with a P450scc cDNA and a mouse beta-actin probe. Compared with P450scc, StAR mRNA expression showed little variability in the magnitude of expression and did not correlate with the endocrine profiles (NAG: StAR 100+/-16%, P450scc 100+/-14%; APA: StAR 80+/-3%, P450scc 94+/-13%; CPA: StAR 71+/-10%, P450scc 109+/-15%; NFA: StAR 64+/-9.5%, P450scc 18+/-5%; means+/-s.e.m.). ACC expressed low levels of both genes probably as a result of dedifferentiation (StAR 29+/-9%, P450scc 46+/-18%). Incubation of the NCI-h295 tumour cell line with 10nmol ACTH and 10micromol forskolin induced an increase in the abundance of StAR and P450scc mRNA, demonstrating gene regulation by the cAMP protein kinase A pathway. Furthermore, we incubated the NCI-h295 tumour cell line with the adrenostatic compounds, aminoglutethimide and metyrapone. We could not detect an effect on the expression of StAR mRNA, whereas the expression of P450scc mRNA was significantly reduced. We conclude that, in contrast to P450scc, StAR seems to be evenly expressed in adrenocortical adenomas. Therefore, the endocrine activity of a given tumour cannot be explained by the abundance of StAR expression. In ACC, both StAR and P450scc expression is low, explaining the relatively inefficient steroid production of these tumours. PMID- 10700726 TI - Induction of cellular immunity in a parathyroid carcinoma treated with tumor lysate-pulsed dendritic cells. AB - BACKGROUND: Cytotoxic T-lymphocyte-mediated tumor immunity against major histocompatibility antigen class II-negative tumors requires help from CD4(+) T cells. The major antigen presenting cells for CD4(+) cell activation are dendritic cells. Studies in mice and humans have demonstrated the potent capacity of these cells to induce specific antitumor immunity. OBJECTIVE: To control the growth of a metastasized parathyroid carcinoma, by immunizing a patient with tumor lysate and parathyroid hormone-pulsed dendritic cells. DESIGN AND METHODS: Mature dendritic cells were generated from peripheral blood monocytes in the presence of granulocyte/macrophage colony-stimulating factor, interleukin-4 and tumor necrosis factor alpha. Antigen-loaded dendritic cells were delivered by subcutaneous and intralymphatical injections. After five cycles, we added keyhole limpet hemocyanin (KLH) as a CD4(+) helper antigen. RESULTS: After 10 vaccinations, a specific cellular immune response to tumor lysate was observed. In vitro T-cell proliferation assays revealed a dose-dependent stimulation index of 1.8-5.7 compared with 0.9-1.1 before vaccination. In vivo immune response was demonstrated by positive delayed-type hypersensitivity toward tumor lysate. Intradermal injection of tumor lysate resulted in an erythema and induration, suggesting the efficient generation of tumor lysate-specific memory T-cells. CONCLUSIONS: These data indicate that dendritic cell vaccination can induce in vitro and in vivo responses in a highly malignant endocrine carcinoma. Regardless of the clinical outcome of our patient, this approach might be generally applicable to other advanced, radio- and chemotherapy-resistant endocrine malignancies, such as adrenal carcinomas and metastasized medullary and anaplastic thyroid carcinomas. PMID- 10700728 TI - Why teach anatomy? Anatomists respond. PMID- 10700727 TI - Effect of long-term ovariectomy and estrogen replacement on the expression of estrogen receptor gene in female rats. AB - OBJECTIVE: Estrogen exerts a wide variety of actions involving many target tissues. We studied the effects of long-term ovariectomy (OVX) and OVX with 17beta-estradiol treatment (OVXE2) on the level of estrogen receptor (ER) gene expression in target tissues of female rats. DESIGN: Three groups of Sprague Dawley female rats were utilized in this study: sham operated (SO), OVX and OVXE2. METHODS: SO and OVX were performed 2 weeks before starting the 17beta estradiol treatment. All groups were maintained on liquid diet for 12 weeks from the time of estradiol treatment. Total RNA was prepared from the tissues of the rats and relative quantitative reverse transcription PCR was utilized to compare the ER alpha-subtype (ERalpha) mRNA level in the three groups for each target tissue. RESULTS: Following long-term OVX, the levels of ERalpha expression showed a significant increase in the uterus, kidney and cerebral cortex and no significant change in the liver, cerebellum, brainstem, heart and thoracic and abdominal aorta compared with their SO levels. On the other hand, a 12-week treatment of OVX rats with 17beta-estradiol restored the previously upregulated ERalpha mRNA to near SO levels except for the liver where the 17beta-estradiol treatment resulted in a significant increase in the ERalpha mRNA level compared with that in SO rats. CONCLUSIONS: We conclude that the regulation of ERs by its ligand is tissue specific. PMID- 10700729 TI - Anatomy at EB2000 PMID- 10700730 TI - Anatomical waxwork modeling: the history of the Bologna anatomy museum. PMID- 10700731 TI - Emotional reactions of medical students to dissecting human bodies: a conceptual approach and its evaluation. PMID- 10700732 TI - Dissecting the role of molecular motors in the mitotic spindle. AB - Accurate segregation of genetic material during both mitosis and meiosis is essential for the viability of future cellular generations. Genetic material is packaged in the form of chromosomes during cell division, and chromosomes are segregated equally into two daughter cells by a dynamic, microtubule-based structure known as the spindle. Molecular motor proteins of the kinesin and dynein superfamilies are essential players in the functional microanatomy of cell division. They power various aspects of spindle assembly and function, including establishing spindle bipolarity, spindle pole organization, chromosome alignment and segregation, regulating microtubule dynamics, and cytokinesis. This review highlights the roles that various members of the kinesin and dynein motor superfamilies play during mitosis and meiosis. Understanding how microtubule motors function during cell division will unravel how the spindle precisely segregates chromosomes, and may offer insights into the molecular basis of disease states that arise from spindle malfunctions. For example, chromosome non disjunction during meiosis causes such disorders as Klinefelter, Turner, and Down Syndromes. Chromosome non-disjunction during mitosis is an important contributing mechanism for tumor progression. In addition, since motor proteins are essential for spindle assembly and function, they provide obvious targets for intervention into the cell division cycle, and compounds that specifically block motor functions during mitosis may prove to be valuable chemotherapeutic agents. Anat Rec (New Anat) 261:14-24, 2000. PMID- 10700733 TI - Comparative respiratory morphology: themes and principles in the design and construction of the gas exchangers. AB - Along the evolutionary continuum, a kaleidoscope of gas exchangers has evolved from the simple cell membrane of the primeval unicells. The most momentous events in this process were: the intensification of molecular oxygen in the biosphere and its appropriation into aerobic metabolism, the rise of multicellular organisms, the development of a circulatory system and carrier pigments in blood, the advocacy of air breathing, adoption of suctional breathing, and the shift to endothermy. To satisfy species-specific needs for oxygen, some constraints were overcome through transactions that obliged certain compromises and trade-offs. Optimal designs of the gas exchangers for particular phylogenetic levels of development, habitat, and lifestyle have developed only so far as to satisfy prescribed needs. The efficiency of the human lung, for example, falls well below those of certain taxa that are considered to be relatively "less advanced." Utilizing different resources and strategies, in fascinating processes of conformity, different groups of animals have developed similar respiratory structures. In most cases, the analogy reflects evolutionary convergence in response to corresponding selective pressures rather than common ancestry. Anat Rec (New Anat) 261:25-44, 2000. PMID- 10700734 TI - Haines DE. 1999. Synopsis Of the 1999 IFAA meeting in rome. Anat rec (New anat) 257:185-186 PMID- 10700735 TI - Forthcoming topics AB - Forthcoming Topics PMID- 10700736 TI - Optimal treatment allocation in comparative biomedical studies. AB - In clinical trials involving several treatment protocols, there are usually several objectives of different importance. A general method for finding an optimal design which reflects this practical need is discussed. In particular, we show that the graphical method of Cook and Wong can be applied when there are two objectives. Several illustrative examples are discussed, including multiple objective designs for comparing several treatment groups with a placebo group. PMID- 10700737 TI - Comparing diagnostic tests: a simple graphic using likelihood ratios. AB - The diagnostic abilities of two or more diagnostic tests are traditionally compared by their respective sensitivities and specificities, either separately or using a summary of them such as Youden's index. Several authors have argued that the likelihood ratios provide a more appropriate, if in practice a less intuitive, comparison. We present a simple graphic which incorporates all these measures and admits easily interpreted comparison of two or more diagnostic tests. We show, using likelihood ratios and this graphic, that a test can be superior to a competitor in terms of predictive values while having either sensitivity or specificity smaller. A decision theoretic basis for the interpretation of the graph is given by relating it to the tent graph of Hilden and Glasziou (Statistics in Medicine, 1996). Finally, a brief example comparing two serodiagnostic tests for Lyme disease is presented. Published in 2000 by John Wiley & Sons, Ltd. PMID- 10700738 TI - Accounting for unreported and missing intercourse in human fertility studies. AB - In prospective studies of human fertility that attempt to identify days of ovulation, couples record each day whether they had intercourse. Depending on the design of the study, couples either (I) mark the dates of intercourse on a chart or (II) mark 'yes' or 'no' for each day of the menstrual cycle. If protocol I is used, intercourse dates that couples fail to record are indistinguishable from dates of no intercourse. Consequently, estimates of day-specific fecundability are biased upwards. If protocol II is used, data from menstrual cycles with missing intercourse information must be discarded in order to fit current fertility models. We propose methods to account for unreported and missing intercourse under the assumption that the missingness mechanism is independent of time conditional on the unobservable true intercourse status. We use probit mixture models to allow for heterogeneity among couples, both in fecundability and in the missingness and non-reporting mechanisms. Markov chain Monte Carlo (MCMC) techniques are used for Bayesian estimation. The methods are generally applicable to the analysis of aggregated Bernoulli outcomes when there is uncertainty in whether a given trial, out of a series of trials, was completed. We illustrate the methods by application to two prospective fertility studies. Published in 2000 by John Wiley & Sons, Ltd. PMID- 10700739 TI - Parametric empirical Bayes estimates of disease prevalence using stratified samples from community populations. AB - Studies of chronic diseases in a community setting often employ stratified sample designs to enable the study to attain multiple research goals at a reasonable cost. One important goal is estimation of disease prevalence in the whole community and in important subgroups. Some adjustment for the sample design is necessary; if the design has many strata with very disparate sampling fractions, simply upweighting observed stratum prevalences may lead to unstable estimators. We propose a parametric empirical Bayes estimator in the spirit of the work of Efron and Morris, and we compare it to the direct upweighted estimator and a regression-smoothed estimator. Simulation studies in realistic settings suggest that the new estimator performs best, giving estimates with low bias and good precision under a variety of models. PMID- 10700740 TI - Estimating age-related trends in cross-sectional studies using S-distributions. AB - Growth trends in children are often based on cross-sectional studies, in which a sample of the population is investigated at one given point in time. Estimating age-related percentiles in such studies involves fitting data distributions, each of which is specific for one age group, and a subsequent smoothing of the percentile curves. The first requirement for this process is the selection of a distributional form that is expected to be consistent with the observed data. If a goodness-of-fit test reveals significant discrepancies between the data and the best-fitting member of this distributional form, an alternative distribution must be found. In practice, there is seldom an objective argument for selecting any particular distribution. Also, different distributions can yield very similar fits, so that any selection is somewhat arbitrary. Finally, the shapes of the observed distributions may change throughout the age range so drastically that no single traditional distribution can fit them all in a satisfactory manner. To overcome these difficulties in population studies, non-parametric smoothing techniques and normalizing transformations have been used to derive percentile curves. In this paper we present an alternative strategy in the form of a flexible parametric family of statistical distributions: the S-distribution. We suggest a method that guides the search for well-fitting S-distributions for groups of observed distributions. The method is first tested with simulated data sets and subsequently applied to actual weight distributions of girls of different ages. As far as the results can be tested, they are consistent with observations and with results from other methods. PMID- 10700741 TI - GEE analysis of negatively correlated binary responses: a caution. AB - The method of generalized estimating equations has become almost standard for analysing longitudinal and other correlated response data. However, we have found that if binary responses have less than binomial variation over clusters, and are modelled using exchangeable correlations, prevailing software implementations may give unreliable results. Bounding the negative correlation away from its theoretical minimum may not always be a satisfactory solution. In such instances, using the independence working correlation structure and robust SEs is a more trustworthy alternative. PMID- 10700743 TI - Retrospective estimation of the birth prevalence for delayed onset disorders: application to cystic fibrosis in Nova Scotia. AB - Maximum likelihood methods are used to estimate the birth prevalence, the age-at diagnosis distribution, and the number of infants born with CF in Nova Scotia over a 20 year period. The data were taken from a registry of CF cases, and, because of the delayed onset of symptoms, some CF affected individuals were not diagnosed by the end of the study period. A goodness-of-fit test for assessing whether the age-at-diagnosis distribution is stable over the 20 years is provided. These methods could be applied equally well to data on any genetic disorder with delayed onset. PMID- 10700742 TI - Interval estimation for Cohen's kappa as a measure of agreement. AB - Cohen's kappa statistic is a very well known measure of agreement between two raters with respect to a dichotomous outcome. Several expressions for its asymptotic variance have been derived and the normal approximation to its distribution has been used to construct confidence intervals. However, information on the accuracy of these normal-approximation confidence intervals is not comprehensive. Under the common correlation model for dichotomous data, we evaluate 95 per cent lower confidence bounds constructed using four asymptotic variance expressions. Exact computation, rather than simulation is employed. Specific conditions under which the use of asymptotic variance formulae is reasonable are determined. PMID- 10700744 TI - Tutorial in biostatistics. Meta-analysis: formulating, evaluating, combining, and reporting by S-L. T. Normand, Statistics in Medicine, 18, 321-359 (1999) PMID- 10700745 TI - Tutorial in biostatistics. Meta-analysis: formulating, evaluating, combining, and reporting by S-L. Normand, Statistics in Medicine, 18, 321-359 (1999) PMID- 10700746 TI - Surgery for intracerebral hemorrhage. PMID- 10700747 TI - SHO education in the UK: the contribution of a distance learning course (MRCS STEP). PMID- 10700748 TI - Evolving models of recertification in the USA. PMID- 10700749 TI - Specialist registrar training in surgical emergencies: concern for Calman training in the United Kingdom. PMID- 10700750 TI - Vascular surgery: the European perspective. AB - Isaac Newton, among others, observed that 'we see so far because we are standing upon the shoulders of giants'. In vascular surgery most of the giants have been European, and this is a heritage which we as Europeans can take pride in and build upon if we chose to do so. As in other areas of life, commitment is essential in order to influence the future. For vascular surgeons in Europe this means active participation in the European scientific societies for vascular surgery and in the UEMS. The main value of the EBSQ.VASC assessments to date has been to expose the uneven standards of training in vascular surgery within the European Union. Only if action follows to address these inequalities will the tactics of the European Board of Vascular Surgery be vindicated. PMID- 10700751 TI - Surgical training and the trainer: creating the ideal situation. PMID- 10700752 TI - A strategic approach to implementing clinical governance. PMID- 10700753 TI - Orthopaedic trainee operative supervision and experience: South-East Thames Regional orthopaedic operative database 1991-1998. PMID- 10700754 TI - Investigations requested by house officers on surgical patients. PMID- 10700755 TI - Where does research fit in? PMID- 10700756 TI - Teaching and research training should be integrated with clinical training. PMID- 10700757 TI - Advances in non-invasive imaging of intracranial vascular disease. AB - Intra-arterial catheter angiography has, in the past, been the mainstay for the investigation of intracranial vascular disease. It is, however, invasive, usually requires in-patients admission, and is associated with a rate of neurological complications between 1% and 3%. In recent years, magnetic resonance angiography (MRA) and CT angiography (CTA) have emerged as non-invasive alternatives for imaging blood vessels and have made a significant impact on neuroradiological investigations. It is the purpose of this article to explain the basic technical principles of these two methods and to give an overview of their current clinical applications. PMID- 10700758 TI - Laparoscopic repair of perforated peptic ulcers. The role of laparoscopy in generalised peritonitis. AB - This non-randomised concurrent cohort study conducted in two teaching hospital Departments of Surgery examined the assumption that the benefits of elective laparoscopic upper gastrointestinal surgery would apply to those with generalised peritonitis due to perforated peptic ulcers. It compared 20 consecutive laparoscopic repairs of perforated peptic ulcers with a concurrent group of 16 consecutive open repairs. There were no differences pre-operatively between the two groups. The mean duration of surgery was similar (P = 0.46). There were no differences in the rate of GI tract recovery, but opiate analgesia requirement in the laparoscopic group was significantly less (P < 0.0001). Intensive care was required in three patients in the laparoscopic group (two with renal failure) and two in the open (no renal failure). Two patients in the laparoscopic and one in the open group died. The median duration of stay was five days in the laparoscopic group and six in the open. This comparison shows that the patho physiological insult of laparoscopy in the setting of generalised peritonitis does not obviously increase the peri-operative risk of organ failure but objective benefits are small. PMID- 10700759 TI - Experience of combined endoscopic percutaneous stenting with ultrasound guidance for drainage of pancreatic pseudocycts. AB - The therapeutic options for treatment of pancreatic pseudocysts are numerous. We report our experience of combined endoscopic and ultrasound guided percutaneous stenting for pancreatic pseudocysts. Data were prospectively collected for 20 consecutive patients. All patients had undergone a standard technique of combined endoscopic and ultrasound guided percutaneous placement of double J stents, between a pancreatic pseudocyst and the stomach. Patients age ranged between 25 and 84 years. Thirteen of the pseudocysts were due to acute pancreatitis and 7 were due to chronic pancreatitis. The duration of the combined procedure was mean 50 min (range 30-95 min). The length of hospital stay was mean 5 days (range 2-77 days. Only two patients suffered postoperative complications; one was re-admitted 2 weeks following stenting with acute cholecystitis, the other suffering a perforated duodenal ulcer 3 weeks after stenting. There were two failures early in the series, both due to stent migration, these stents were of a small size, (4.7 French). Following this the stent size was increased to at least 7 French, no further failures occurred. There was no operative mortality for the series. Follow-up ranged between 6 months and 5 years. We conclude that a combined percutaneous and endoscopic cyst-gastrostomy stent is a safe and effective treatment for patients with suitably placed pseudocysts. PMID- 10700760 TI - A useful mnemonic for severity stratification in acute pancreatitis. PMID- 10700762 TI - Association between Streptococcus milleri and abscess formation after appendicitis. AB - Abscesses after appendicitis occur in some patients despite timely surgery and antibiotics. The Streptococcus milleri group of bacteria are commonly associated with gastrointestinal abscesses. This study investigated the relationship between S. milleri and abscess formation after appendicectomy a total of 301 patients (172 males, 129 females, median age 22 years) with appendicitis were identified retrospectively from the hospital PAS computer system who had an appendicectomy and peritoneal bacteriology swabs taken. All but one patient had prophylactic antibiotics. Patients were divided into three groups according to peritoneal bacteriology: group 1 (S. milleri +/- mixed faecal organisms, n = 61); group 2 (mixed faecal organisms, n = 126); and group 3 (sterile, n = 114). The chi squared and Student t-tests were used for statistical analysis. Thirteen (21%) of group 1 patients developed an intra-abdominal abscess compared with 4 (3%) in group 2 and 2 (1.7%) in group 3 (P < 0.0001). There was no difference in the prevalence of gangrenous or perforated appendicitis between groups 1 and 2 (56% versus 52%) but these worse forms of appendicitis were less common in group 3 (22%). Group 1 patients had a mean total hospital stay of 10 days versus 6 days for group 2 and 4 days for group 3 (P < 0.001). S. milleri was associated with a 7-fold increase in abscess formation after appendicectomy and a longer hospital stay. Antibiotic prophylaxis did not prevent this complication. PMID- 10700761 TI - John Hunter, Frederick Treves and intussusception. AB - Early this century, intussusception in childhood was usually fatal. John Hunter, one of the founding fathers of scientific surgery was amongst the first to accurately describe the clinico-pathological features of the condition and one of the great nineteenth century surgeons, Sir Frederick Treves, suggested a plan of management for intussusception which remains little changed up to the present day. PMID- 10700763 TI - Audit of topical glyceryl trinitrate for treatment of fissure-in-ano. AB - PURPOSE: To clarify the clinical role of topical glyceryl trinitrate (GTN) in the management of anal fissures. PATIENTS AND METHODS: Fifty-six consecutive patients with fissure-in-ano attending a colorectal clinic from April 1997 to May 1998 included 16 acute and 40 chronic anal fissures. Patients were instructed to apply 0.2% 0.5 g of GTN to the painful area of the anus. Patients were followed-up in the clinic at 4, 8 and 12 weeks, and by telephone interviews at a median follow up of 10 months. RESULTS: Ten of 16 acute fissures (63%) were healed by 4 weeks and 13 (81%) by 8 weeks. Thirteen of 40 chronic fissures (33%) were healed by 8 weeks and 20 (50%) by 12 weeks. Seventeen patients (30%) underwent lateral sphincterotomy and all healed. There were five recurrences within 3 months of treatment with GTN. Thirty-four (61%) suffered from headaches, eight being severe headaches. None of the patients developed incontinence with GTN or lateral sphincterotomy. CONCLUSIONS: Treatment of fissure-in-ano using GTN ointment was effective in up to 50% of patients with chronic anal fissure, and has the benefit of being repeatable if the fissure recurs. Patients should be aware that treatment is likely to take some months to be effective and is associated with significant side effects in up to 15% of patients. PMID- 10700764 TI - Are surgeons aware of the dangers of diathermy? AB - Surgical diathermy is an invaluable aid in modern surgery and most contemporary diathermy machines are considered safe. However, diathermy accidents still do occur and a diathermy unit can be potentially lethal if adequate care is not exercised in its use. PMID- 10700765 TI - Current attitudes to total hip replacement in the younger patient: results of a national survey. AB - A postal questionnaire was sent to all practicing consultant orthopaedic surgeons in the UK seeking information regarding their usual total hip replacement practice, the age at which they would define a patient as falling into the 'young hip group' and whether this might modify their practice. In particular, in the 'younger' age group, we were interested in the frequency of usage of uncemented implants, the choice of implant and the bearing surfaces. Of 1242 surgeons surveyed, we had a response from 935 who currently undertake total hip arthroplasty. Their responses confirm that approximately 60,645 total hip replacements are performed annually in the UK of which 9,376 are performed in the younger age group (mean age 57.5 years). As with our previous survey, the most popular prosthesis in the 'older' age group overall was the Charnley (51%) followed by the Exeter (15%). These implants also proved to be the most popular in the 'younger' age group (40% Charnley, 18% Exeter), with 75% of surgeons choosing a cemented stem, and 65% also opting to cement the socket. 23% of surgeons used hydroxy-apatite coated implants on both the femoral and acetabular sides of the joint. Stainless steel remained the most popular choice of femoral head bearing surface (42%) followed by chrome-cobalt (33%) and ceramic (25%). On the acetabular side, high density polyethylene predominated--accounting for 95%, with only 3% using chrome cobalt and 2% ceramic. There would appear to be a remarkably conservative attitude among British surgeons, the majority of whom prefer to stick with tried and tested cemented femoral implants when dealing with the younger patient. There are a small number of uncemented acetabulae and the hybrid configuration. Hydroxy-apatite coatings seem to be the most popular choice for the non-cemented prostheses. Ceramic femoral heads are used more frequently than the ceramic acetabular bearing, and equally metal/metal bearings remain infrequently used. PMID- 10700766 TI - Trochanteric non-union in revision total hip arthroplasty: does it matter? AB - The aims of this study were to assess whether trochanteric non-union is an important factor in revision total hip arthroplasty in terms of postoperative morbidity. We studied prospectively 97 consecutive patients undergoing revision total hip arthroplasty in the years 1992-1996. All operations were performed by one surgeon through a Charnley trans-trochanteric approach. The patients were followed-up over a period of 1-4 years and at 12 months postsurgery were assessed using a modified scoring system devised by D'Aubigne. Anatomical union of the greater trochanter was assessed by an anterior-posterior pelvic radiograph at 12 months to decide if the greater trochanter was united in the correct anatomical position. The trochanteric non-union rate was 18.5% (18 out of 97 patients). There was no significant difference between the patients in terms of pain, function and satisfaction scores at one year between those with trochanteric union and those without. This study suggests that trochanteric non-union post revision total hip arthroplasty is not a cause of increased morbidity. PMID- 10700767 TI - Use of cut resistant glove liner in revision hip surgery. PMID- 10700768 TI - A one-centre prospective audit of peri- and postoperative blood loss and transfusion practice in patients undergoing hip or knee replacement surgery. AB - We prospectively audited peri-operative blood loss and blood transfusion practice in 42 elderly patients (mean age, 71.8 years, 68% female) undergoing hip or knee surgery in an orthopaedic unit. Only in 57% of all operations was blood loss recorded. Compliance with the Maximum Surgical Blood Ordering Schedule (MSBOS) was variable, and Cross-matching to Transfusion (C/T) ratios were low. In 86% of operations, blood had been issued pre-operatively (average three units, range = 1 61 units). Of these patients, 75% subsequently received a transfusion. In 26% of all the operations, the transfusion, although confirmed by the blood transfusion laboratory records, had not been recorded in the medical or nursing notes. The average pre-operative Hb in the transfusion group was 123 g/l (range, 80-144 g/l) and 112 g/l postoperatively and after a transfusion (range, 75-133 g/l). This compared to the non-transfusion group's value of 124 g/l (range, 86-186 g/l) and 113 g/l (range, 77-147 g/l) postoperatively. The high blood issuing and transfusion rates raise the concern that transfusions are being given in response to habit or blood availability, and not medical indications. This would imply that some patients are exposed to unnecessary risks. Furthermore, inadequate documentation of the transfusion process opens the medical profession to criticism and medical, legal and ethical complications regarding patient care. Positive improvements suggested by regular medical audit may help address these problems. PMID- 10700769 TI - Watson Jones Lecture. The organisation of trauma services in the UK. AB - To provide a high level of orthopaedic trauma care, education and research, across the country, trauma services in the UK require modification. Good information is necessary prior to formulating ideas and proposals. Trauma care provision must be considered comprehensively at both the national and local levels. As a first step, it is important to know just how many acute hospitals there are in the country. It is also important to know about the distribution of surgical specialities and the number of consultant orthopaedic surgeons staffing those hospitals. PMID- 10700770 TI - The effect of a 'fast-track' unit on the performance of a cardiothoracic department. AB - OBJECTIVE: The objective of this study was to describe the impact of a 'fast track' unit, combined with a computerised system for information collection and analysis, on the clinical practice and finance of a cardiothoracic department over the first 12 month period of its application. METHODS: Within 12 months, starting December 1996, 642 major cardiothoracic cases were performed at the Cardiothoracic Department, St Mary's Hospital, London, after the establishment of a 3-bed 'fast-track' unit, which was supported by a computerised system for admission planning and a pre-admission clinic. The main outcome measures were operating numbers, financial income, patient recovery and operative mortality. RESULTS: The 'fast-track' unit resulted in an increase of the operating numbers (11.3% increase in major cardiac cases) and income (38%), as compared with the year before. Some 525 patients out of 642 (81.8%) were scheduled for the 'fast track' unit and 492 (93.7%) were successfully 'fast-tracked'. Coronary artery bypass grafting operations had the lowest 'fast-track' failure and mortality rates. Re-do operations and complex coronary procedures presented a high 'fast track' failure rate of approximately 20-25%. Low cardiac output, postoperative bleeding and respiratory problems were the most frequent causes for 'fast-track' failure. CONCLUSIONS: The development of a 'fast-track' unit, supported by a computerised system for information collection and analysis and a pre-admission clinic, has resulted in a substantial improvement of operating numbers and financial income, without adversely affecting the clinical results. This task demanded close collaboration between a dedicated list manager and a designated member of the medical team. Patient selection with appropriate 'fast-track,' criteria may improve further the efficiency of 'fast-track' units in the future. PMID- 10700771 TI - Screening of abdominal aortic aneurysm: a pragmatic approach. AB - In order to evaluate the feasibility of a selective screening programme for abdominal aortic aneurysm (AAA) within an urban setting and assess its impact on the expected increase in workload for the local hospital(s), a population based, prospective study was performed. A total of 4823 men aged 65 years were invited for ultrasound examination of the abdominal aorta between January 1993 and April 1997 as part of a general practice-based aneurysm screening programme covering two districts with a general hospital each. All examinations were carried out by senior radiographers using a portable B mode grey scale machine and a 3.5 MHz curvi-linear array probe. Patients with a maximum aortic diameter of over 3 cm were annually recalled, those with over 4 cm were referred to hospital for an out patient's appointment. Those with AAA greater than 5 cm were considered for surgery. Of those approached, 3497 (72.5%) took part in the study, 1206 (25%) did not attend and 120 (2.5%) were excluded by their general practitioners (GPs) on medical grounds. Of the men taking part, 3130 (89.5%) had an aortic diameter equal to or less than 2.5 cm, 196 (5.6%) between 2.6 and 3.0 cm, and 171 (4.9%) had aortic diameters greater than 3 cm--29 of whom had AAA greater than 5 cm with a mean diameter of 6.0 cm (range 5.1-9.0 cm). Of 127 men with an initial diameter of 3.1-4.0 cm (mean progression in size of 2.3 mm/year), 22 enlarged to > 4 cm and 3 to > 5 cm. Of 24 men with an initial diameter of 4.1-5.0 cm, 6 enlarged to > 5 cm. Some 69 (2%) patients were referred to hospital requiring a total of 125 consultations (1.8 consultations per patient); 21 underwent surgery and one died from rupture whilst awaiting surgery. Five patients refused their operation and two failed to attend the clinic (all > 5 cm) but remain well to date. No patient died following surgery. We conclude that, screening for AAA in men at age 65 years within an urban setting is feasible and well received by patients and GPs. Screening does not lead to a huge increase in terms of outpatient appointments and operations for AAA. PMID- 10700772 TI - Elective abdominal aortic aneurysm operations--the results of a single surgeon series of 243 consecutive operations from a district general hospital. AB - BACKGROUND: There are few data on the morbidity and mortality of planned elective surgery for infrarenal abdominal aortic aneurysm (AAA) as a single surgeon series. This audit is of a consecutive series of AAA operations performed by one surgeon in one district general hospital over a 13-year period. METHODS: 243 patients were operated on for AAA between 1985 and 1998. Data were collected on the majority of patients prospectively. A reliable method was devised to identify all patients. Any missing complication and mortality data were then collected retrospectively. RESULTS: 13 patients died as a result of their operation (5.3%). In patients over the age of 80 years (36), five patients died (14%) and in the 207 patients under the age of 80 years, eight died (3.8%). Cardiac deaths were the most frequent cause (38%); 82 patients had recorded complications (34%). The operative mortality rate has increased in later years, (2.2% to 7.1%), largely due to an increase in the very elderly accepted for operation (12% to 16%), and a possible increase in co-morbidity. CONCLUSIONS: An acceptable and comparable mortality rate can be achieved in a district general hospital. The complication rate is high indicating the need for very intense medical and nursing care for these patients postoperatively. There is a considerable variance in mortality rates with age and risk even in the practice of one surgeon, indicating a need to be very knowledgeable and cautious in interpreting postoperative mortality data. This is the largest single surgeon series to date in the UK. PMID- 10700773 TI - Deep vein thrombosis prophylaxis protocol--needs active enforcement. AB - Each hospital department tends to have its own DVT prophylaxis protocol generally based on the recommendations of the THRIFT consensus group. This is developed to help the junior medical staff to prescribe the appropriate prophylaxis according to risk assessment. However, adherence to the protocol tends to be haphazard unless actively enforced. This study is aimed at determining whether active enforcement of the protocol improves the uptake of prophylaxis. PMID- 10700774 TI - Ultrasound-guided unilateral neck exploration for sporadic primary hyperparathyroidism: is it worthwhile? PMID- 10700775 TI - Safety of pancreatic surgery in a small DGH. PMID- 10700776 TI - Lung protective ventilatory strategies--will these prevail in the next millennium? PMID- 10700777 TI - Why we need large randomized studies in anaesthesia. PMID- 10700778 TI - Effects of hyperoxia and hypocapnia on regional venous oxygen saturation in the primary visual cortex in conscious humans. AB - Hyperoxia can improve oxygen delivery in patients exposed to hypocapnia for neurosurgical procedures but this effect may be modified by regional differences in the degree of hypocapnic vasoconstriction. Using functional magnet resonance imaging (fMRI), we have investigated the influence of hyperoxia on blood flow and blood oxygenation in the primary visual cortex in hypocapnic volunteers. Consecutive fMRI measurements were performed in 10 awake, male volunteers during hypocapnia (mean PE'CO2 3.3 (SD 0.1) kPa) and normocapnia (PE'CO2 5.3 (0.1) kPa) at FIO2 values of 0.21 and 1.0, respectively. Hypocapnia significantly reduced the pixel count in the primary visual cortex (median 169 (quartiles 34-246) vs 21 (0-40) pixels at an FIO2 of 0.21). Additional hyperoxia had no influence on this reduction in pixel count (16 (0-28) pixels at FIO2 1.0 vs 21 (0-40) pixels at FIO2 0.21). Hyperoxia did not influence hypocapnic vasoconstriction in the primary visual cortex. These data suggest that in the primary visual cortex, administration of oxygen alone may not be sufficient to improve oxygen delivery under hypocapnic conditions. PMID- 10700779 TI - Effects of target-controlled infusion of propofol on the transient hyperaemic response and carbon dioxide reactivity in the middle cerebral artery. AB - The transient hyperaemic response (THR) of blood flow velocity in the middle cerebral artery (vmca), measured by transcranial Doppler ultrasonography (TCD), can be used to assess cerebral autoregulation. We have studied the effects of propofol administered by target-controlled infusion on vmca, THR and carbon dioxide reactivity. We studied 20 healthy adult patients undergoing elective surgery. A standardized anaesthetic comprising alfentanil 10 micrograms kg-1, propofol via a target-controlled infusor and vecuronium 0.1 mg kg-1 was used in both parts of the study. In the first part, THR tests were performed on 10 subjects while awake and then at an 'induction' target concentration of propofol (the target at which consciousness was lost, mean 6.7 (SD 1.1) micrograms ml-1). In the carbon dioxide study, reactivity was tested in 10 patients while awake and at the 'induction' target concentration of propofol by altering the end-tidal carbon dioxide partial pressure by 1 kPa either side of baseline. Propofol caused a significant decrease in vmca but indices of autoregulation, THR ratio and strength of autoregulation increased significantly. Propofol had no effect on carbon dioxide reactivity. These results suggest that propofol may have a beneficial effect on cerebral haemodynamics. PMID- 10700780 TI - Gram stain of bronchoalveolar lavage fluid in the early diagnosis of ventilator associated pneumonia. AB - To assess the usefulness of the Gram stain in the early diagnosis of ventilator associated pneumonia (VAP), we performed 146 protected specimen brushings (PSB) and bronchoalveolar lavages (BAL) in 118 patients suspected of having nosocomial pneumonia. Gram stain and counts of infected cells were performed in all samples from BAL fluid. A final diagnosis of pneumonia was established in 51 patients and there was no infection in 95 cases. A threshold of 2% of infected cells was used to distinguish between VAP and the group without VAP (sensitivity 86.3%, specificity 78.9%, positive predictive value 68.7% and negative predictive value 91.4%); there was good agreement with the final diagnosis (kappa statistic 0.616; concordance 81.5%). Regarding detection of bacteria using the Gram stain, we found a sensitivity of 90.2%, specificity 73.7%, positive predictive value 64.8% and negative predictive value 93.3%; there was moderate agreement with the final diagnosis (kappa statistic 0.586; concordance 79.4%). In the VAP group, we analysed the degree of qualitative agreement between Gram stain and PSB quantitative cultures: the correlation was complete in 51% (26 of 51 VAP), partial in 39.2% (20 of 51 VAP) and there was no correlation in 9.8% (five of 51 VAP). We conclude that the Gram stain is useful for rapid diagnosis of VAP but unreliable for early adaptation of empiric therapy. PMID- 10700781 TI - Perioperative myocardial ischaemia, heart rate and arrhythmia in patients undergoing thoracotomy: an observational study. AB - We have studied myocardial ischaemia, heart rate and arrhythmia in 82 patients undergoing elective thoracotomy. Myocardial ischaemia was detected using a microprocessor-based surveillance system programmed to record leads V2 and V5. Patients were monitored on the day before and for up to 72 h after surgery. The total monitoring time was 5158 h. The incidence of silent myocardial ischaemia before operation was 11% (nine of 82). This increased to 24% (20 of 82) after operation. Postoperative myocardial ischaemia was associated with preoperative myocardial ischaemia in six patients. Before operation, the mean duration of myocardial ischaemia was 0.31 min per hour of monitoring. After operation, this increased to 1.36 min per hour of monitoring (P < 0.05). For the whole population, mean heart rate before operation was 74 beat min-1 and increased to 84 beat min-1 after operation (P < 0.01). Patients with ischaemia had a mean heart rate of 92.8 beat min-1 after operation compared with those with no ischaemia whose mean heart rate was unchanged at 81.8 beat min-1 (P < 0.05). The incidence of atrial tachyarrhythmia increased from one patient before operation to 12 patients after operation (P < 0.01). Atrial tachyarrhythmia was not associated with postoperative myocardial ischaemia. Nine patients had an adverse operative outcome; two had non-fatal myocardial infarction and seven died. Postoperative myocardial ischaemia was associated with adverse outcomes (P < 0.05). PMID- 10700782 TI - Malignant hyperthermia causing Gly2435Arg mutation of the ryanodine receptor facilitates ryanodine-induced calcium release in myotubes. AB - We have investigated if cultivated muscle cells from malignant hyperthermia (MH) patients can be distinguished pharmacologically from controls. Muscle specimens from four individuals carrying the Gly2435Arg mutation of the skeletal muscle ryanodine receptor protein (RYR1) and from four controls were used to culture myotubes. Resting intracellular calcium concentration ([Ca2+]i) of MH myotubes was similar to controls. However, when ryanodine 0.5 mumol litre-1 was added, the kinetics of the increase in the calcium signals in MH and control cells were significantly different; the time for half maximum increase was mean 197 (SD 131) s for MH cells and 474 (61) s for controls (n = 80 cells each). On average, the area under the MH response curves was twice the control value. These results give rise to hopes that the phenotype of MH can be characterized using cultured human muscle and that a culture-based test for MH susceptibility may eventually be developed. PMID- 10700783 TI - Rapacuronium for modified rapid sequence induction in elective caesarean section: neuromuscular blocking effects and safety compared with succinylcholine, and placental transfer. AB - We have compared rapacuronium 2.5 mg kg-1 (n = 20) with succinylcholine 1.5 mg kg 1 (n = 22) in a multicentre, blinded, randomized study in full-term parturients undergoing elective Caesarean section under general anaesthesia. Thiopental 5 mg kg-1 was given i.v. followed by the neuromuscular blocking agent. Sixty seconds later intubation was performed. Intubating conditions, evaluated as excellent, good or poor, were good to excellent in 95% and 91% in the intent-to-treat patients after rapacuronium and succinylcholine, respectively (ns). Mean onset times at the adductor pollicis muscle for rapacuronium and succinylcholine were 80.4 (SEM 14.4) s and 63.9 (5.6) s (ns) while maximum block was 96 (1.9)% and 99 (0.4)%, respectively (ns). Rate of recovery was significantly longer after rapacuronium; times for return of T1 to 25% were 16.9 (1.5) min and 9.6 (1.1) min for rapacuronium and succinylcholine, respectively (P = 0.0004). Maternal side effects included more tachycardia and skin erythema with rapacuronium; no maternal mortality or morbidity, including bronchospasm, occurred in either group. There were no neonatal adverse effects in either group based on: Apgar scores at 1 and 5 min; times to sustained respiration; neuroadaptive capacity scores at 15 min, 2 h and 24 h; and umbilical venous and arterial blood-gas values and acid-base status. At delivery (17.7 (3.2) min), mean maternal plasma concentrations of rapacuronium were 9041.4 (1259.1) ng ml-1 and 506.4 (24.9) ng ml-1 for Org 9488 (the main metabolite). Corresponding values for umbilical venous plasma were 808.0 (92.1) ng ml-1 and 59.1 (6.5) ng ml-1, and for umbilical arterial plasma, 361.4 (56.4) ng ml-1 and 29.7 (4.6) ng ml-1, respectively. Umbilical venous to maternal venous ratios for rapacuronium and Org 9488 were 8.8% (1.3)% and 10.2 (1.7)%, respectively. PMID- 10700784 TI - Combined spinal-epidural analgesia in labour: comparison of two doses of intrathecal bupivacaine with fentanyl. AB - We have compared intrathecal bupivacaine 1.25 mg and fentanyl 25 micrograms (group A) with bupivacaine 2.5 mg and fentanyl 25 micrograms (group B), for combined spinal-epidural analgesia in 49 labouring parturients in a prospective, randomized, double-blind study. Onset and quality of analgesia were similar in both groups, with median visual analogue scale pain scores of 0 achieved in 5-10 min. Median duration of analgesia was longer in group B (median 120 (range 90 120) min) compared with group A (75 (75-105) min) (P = 0.013). Median upper sensory level was higher in group B compared with group A at 15 min (T6-7 vs T11, P = 0.003) and at 30 min (T6 vs T11-12; P = 0.001). Motor block was greater in group B: seven patients had a modified Bromage score > or = 1 compared with none in group A at 15 min (P = 0.017). Group B also had a greater decrease in arterial pressure. Patient-midwife satisfaction scores and other side effects were similar. We conclude that intrathecal bupivacaine 1.25 mg with fentanyl 25 micrograms provided analgesia of similar onset and quality compared with bupivacaine 2.5 mg and fentanyl 25 micrograms. Although the duration of analgesia was shorter, the incidences of motor block and hypotension were less with the smaller dose. PMID- 10700785 TI - Interscalene brachial plexus anaesthesia with 0.5%, 0.75% or 1% ropivacaine: a double-blind comparison with 2% mepivacaine. AB - We have compared interscalene brachial plexus block performed with ropivacaine or mepivacaine in 60 healthy patients undergoing elective shoulder surgery. Patients were allocated randomly to receive interscalene brachial plexus anaesthesia with 20 ml of 0.5% ropivacaine (n = 15), 0.75% ropivacaine (n = 15), 1% ropivacaine (n = 15) or 2% mepivacaine (n = 15). Readiness for surgery (loss of pinprick sensation from C4 to C7 and inability to elevate the limb from the bed) was achieved sooner with 1% ropivacaine (mean 10 (SD 5) min) than with 0.5% ropivacaine (22 (7) min) (P < 0.001) or 2% mepivacaine (18 (9) min) (P < 0.02). Postoperative analgesia was similar with the three ropivacaine concentrations (11.5 (5) h, 10.7 (2) h and 10 (2.4) h with 0.5%, 0.75% and 1% concentrations, respectively) and nearly two-fold longer compared with 2% mepivacaine (5.1 (2.7) h) (P < 0.001). PMID- 10700786 TI - Effect of oral and i.v. tenoxicam in postoperative pain after total knee replacement. AB - We have evaluated the effect of oral and i.v. tenoxicam on postoperative pain after unilateral total knee replacement in a double-blind, randomized, controlled study. Tenoxicam was administered to two groups of patients, either before (40 mg orally) or after (40 mg i.v.) surgery, then at 24 h after surgery (40 mg i.v.) and at the end of each day for 8 days (20 mg orally). A third group were given placebo at all times. All patients had access to PCA morphine for the first 48 h and then co-dydramol tablets for the duration of the study. We studied 101 patients, mean age 67 yr. There was no significant reduction in the requirement for PCA morphine for the duration of the study in either of the treatment groups, or for co-dydramol in the first 2 days, but tenoxicam significantly reduced the need for co-dydramol over the remaining 7 days. There were no significant differences in mobility between groups. There was a high incidence of adverse events reported, with a similar number in each of the three groups. PMID- 10700787 TI - Asymptomatic intranasal abnormalities influencing the choice of nostril for nasotracheal intubation. AB - We have studied the prevalence of intranasal abnormalities that may influence the choice of nostril for intubation, using the fibreoptic laryngoscope, in 60 oral surgery patients presenting for nasotracheal intubation under general anaesthesia, who had no symptoms or signs of nasal obstruction. Videotape recordings were made during each nasendoscopy and later analysed by an anaesthetist and an otolaryngologist. A total of 68% of patients had intranasal abnormalities (10% bilateral and 58% unilateral) which resulted in one nostril being more patent than the other and therefore considered more suitable for intubation. The most common abnormality was deviated nasal septum which occurred in 57% of the study group; 22% were minor deviations, 13% were major deviations and 22% were impactions. Other abnormalities were simple spurs, unilateral polyp and hypertrophy of the inferior turbinate. In view of the relatively high incidence of intranasal pathology revealed on endoscopic examination, anaesthetists should consider using the fibreoptic laryngoscope to select the best nostril when performing nasotracheal intubation. PMID- 10700788 TI - Improving the shape and compliance characteristics of a high-volume, low-pressure cuff improves tracheal seal. AB - A prototype design of a compliant latex, high-volume, low-pressure cuffed tracheal tube cuff (CHVLP) was compared with the Mallinckrodt Hi-Lo, Sheridan preformed and Portex Profile high-volume, low-pressure (HVLP) cuffed tracheal tubes for leakage of dye placed above the cuff in a benchtop mechanical ventilation model and in five isolated pig tracheas. There was no leakage in the ventilation model or in the pig tracheas with the prototype CHVLP. There was rapid leakage in the ventilation model and in all the pig tracheas for the Mallinckrodt Hi-Lo, the Sheridan preformed and the Portex Profile cuffs. This benchtop study suggests that improved HVLP cuff compliance characteristics may be beneficial in the prevention of leakage of fluid to the lungs known to occur with HVLP cuffs. PMID- 10700789 TI - Variables used to set PEEP in the lung lavage model are poorly related. AB - Setting an appropriate positive end-expiratory pressure (PEEP) value is determined by respiratory mechanics, gas exchange and oxygen transport. As these variables may be optimal at different PEEP values, a unique PEEP value may not exist which satisfies both the demands of minimizing mechanical stress and optimizing oxygen transport. In 15 surfactant-deficient piglets, PEEP was increased progressively. Arterial oxygenation and functional residual capacity (FRC) increased, while specific compliance of the respiratory system decreased. Static compliance increased up to a threshold value of PEEP of 8 cm H2O, after which it decreased. This threshold PEEP did not coincide with the lower inflection point of the inspiratory limb of the pressure-volume (PV) loop. Oxygen transport did not correlate with respiratory mechanics or FRC. In the lavage model, the lower inflection point of the PV curve may reflect opening pressure rather than the pressure required to keep the recruited lung open. Recruitment takes place together with a change in the elastic properties of the already open parts of the lung. No single PEEP level is optimal for both oxygen transport and reduction of mechanical stress. PMID- 10700790 TI - Different effects of dopamine and dopexamine on the isolated perfused rat kidney. AB - We have investigated the effect of dopamine and dopexamine on the isolated perfused rat kidney. After an equilibration period of 20 min and two control periods of 10 min, dopexamine 1.0, 2.5 or 4.0 micrograms kg-1 min-1 or dopamine 2.0 micrograms kg-1 min-1 were perfused for a further 40 min in random order. Renal blood flow, urine volume, glomerular filtration rate, absolute sodium excretion and fractional sodium reabsorption of the isolated perfused kidney were measured every 10 min during the experiment. Dopamine increased significantly urine production from mean 61.54 (SEM 4.7) to 117.2 (9.7) microliters min-1 g-1 and absolute sodium excretion from 0.4 (0.1) to 1.2 (0.1) mumol min-1 g-1, and decreased significantly fractional sodium reabsorption from 97.3 (0.5) to 90.7 (0.7)%. Renal blood flow and glomerular filtration rate were not altered. In contrast, dopexamine had no effect on the isolated kidney. These data suggest that the diuretic and natriuretic effects of dopexamine in humans may not result from a direct action on the kidney. PMID- 10700791 TI - Effect of propofol on reperfusion injury after regional ischaemia in the isolated rat heart. AB - Free oxygen radicals and intracellular calcium homeostasis play important roles in the development of myocardial reperfusion injury. Propofol is a radical scavenger with calcium channel blocking properties. We have investigated the effects of propofol on myocardial reperfusion injury. We used an isolated rat heart model where heart rate, ventricular volume and perfusion pressure were constant. The left anterior descending coronary artery (LAD) was occluded for 30 min and reperfused for 2 h. We studied an untreated control group, an Intralipid group (1 microliter ml-1) and a propofol group (Intralipid 1 microliter ml-1 and propofol 1 microgram ml-1) (n = 12 each). Drugs were infused for 20 min starting 5 min before reperfusion. We measured left ventricular developed pressure (LVDP), coronary flow and infarct size. LAD occlusion reduced mean LVDP from 129 (SEM 4) to 36 (3) mm Hg and mean coronary flow from 12.2 (0.3) to 5.2 (0.2) ml min-1. During reperfusion, LVDP recovered to 98 (4) mm Hg and coronary flow to 11.9 (0.4) ml min-1. Haemodynamic variables were similar in all groups. Propofol had no effect on infarct size compared with the Intralipid group (25.0 (3.7) vs 26.9 (3.3)% of the area at risk; P = 0.89). Infarct size in the Intralipid group tended to be smaller compared with the control group (34.8 (3.2)%; P = 0.19). We conclude that propofol, at a clinically relevant concentration, provided no protective effect against myocardial reperfusion injury in the rat heart in vitro. PMID- 10700792 TI - Effects of tramadol stereoisomers on norepinephrine efflux and uptake in the rat locus coeruleus measured by real time voltammetry. AB - Despite its structural similarity to codeine, tramadol is an unusual analgesic whose antinociceptive efficacy is not solely a result of opioid actions but also of its apparent capacity to block monoamine uptake. Tramadol is a mixture of stereoisomers. In this study, we have examined the actions of racemic, (+)- and ( )-tramadol, in addition to O-desmethyltramadol (the main human metabolite), on electrically evoked norepinephrine efflux and uptake in the locus coeruleus brain slice, measured by fast cyclic voltammetry. Racemic tramadol and its (+)- and (-) enantiomers (all at 5 mumol litre-1) significantly increased stimulated norepinephrine efflux (P < 0.01) by mean 66 (SEM 10)%, 57 (7)% and 64 (13)%, respectively. However, only (-)-tramadol blocked norepinephrine reuptake (P < 0.01), increasing the reuptake half-time to 499 (63)% of pre-drug values. The metabolite O-desmethyl tramadol was inactive at the concentration tested (5 mumol litre-1). In the case of (-)-tramadol, the effect on norepinephrine efflux was directly proportional to, but significantly smaller than, the effect on norepinephrine uptake (P < 0.01). This appeared to be a result of compensatory alpha 2A autoreceptor tone as the selective alpha 2A autoreceptor antagonist BRL 44408 (1 mumol litre-1) eliminated this difference when its own effects on norepinephrine reuptake were taken into account. The efficacy of (-)-tramadol on norepinephrine uptake, at clinically relevant concentrations, may contribute to its antinociceptive efficacy. PMID- 10700793 TI - Development of academic anaesthesia in the UK up to the end of 1998. PMID- 10700794 TI - Antiemetic prophylaxis in cardiac surgery: comparison of metoclopramide and ondansetron. AB - We have compared the effectiveness of ondansetron (115 patients) and metoclopramide (101 patients) for prevention of postoperative nausea and vomiting in patients undergoing cardiac surgery involving cardiopulmonary bypass. In a prospective, randomized, controlled, double-blind study, patients received oral ondansetron 16 mg or oral metoclopramide 10 mg, 1-2 h before surgery. Anaesthesia was not standardized. Assessments of the severity of nausea and occurrence of vomiting were made at intervals after extubation and until discharge from the intensive care, or for a total of 24 h. Compared with the metoclopramide group, the ondansetron group had a higher incidence of nausea (49.6% vs 33.7%; P < 0.05) and vomiting (42.6% vs 24.8%; P < 0.01). There was no difference between groups in the number of patients who accepted postoperative antiemetics (ondansetron 43.4% vs metoclopramide 32.6%) and there was no difference in the incidence of symptoms of moderate or severe nausea. PMID- 10700795 TI - Aerobic, anaerobic and combination estimates of cerebral hypoperfusion during and after cardiac surgery. AB - We studied 15 patients undergoing cardiac surgery involving hypothermic cardiopulmonary bypass (CPB). Cerebral arteriovenous difference in oxygen content (AVDO2) was significantly less during CPB and for up to 18 h after operation compared with pre-CPB values (P < 0.05). There were no significant changes in mean jugular bulb oxyhaemoglobin saturation (SjvO2), cerebral arteriovenous difference in lactate content or lactate-oxygen index (LOI). SjVO2 and arterial carbon dioxide tension (PaCO2) (P = 0.005) were positively correlated as were AVDO2 and haemoglobin concentration (P = 0.012). AVDO2 and PaCO2 (P = 0.007) were negatively correlated as were LOI and arterial oxyhaemoglobin saturation (P = 0.037). There were no significant correlations between mean arterial pressure and any of the variables. SjVO2 and AVDO2 may require correction for changes in PaCO2 and haemoglobin concentration before relating these variables to cerebral outcome. PMID- 10700796 TI - Supralaryngeal tubeless combined high-frequency jet ventilation for laser surgery of the larynx and trachea. AB - We have developed a new technique of combined high-frequency jet ventilation (HFJV), characterized by simultaneous application of a low-frequency (LF) and a high-frequency (HF) jet stream. Tubeless supralaryngeal jet ventilation was delivered via a modified Kleinsasser laryngoscope. We studied 44 adults undergoing 45 elective surgical procedures of the larynx and trachea using a carbon dioxide laser during HFJV. Applied inspiratory oxygen ratios ranged from 0.4 to 1.0. Mean driving pressures of the HF and LF jet streams were 1.5 bar and 1.8 bar in adults, respectively. Mean duration of HFJV was 41 (range 10-180) min. HFJV resulted in mean PaO2 and PaCO2 values of 16.6 (range 9.8-26.9) kPa and 5.7 (3.0-7.6) kPa, respectively. Tubeless supralaryngeal HFJV was safe and effective in maintaining gas exchange in the presence of laryngeal or tracheal stenoses, providing optimal visibility of anatomical structures, offering maximum space for surgical manipulation, and avoiding the use of combustible material inside the larynx or trachea. PMID- 10700797 TI - Influence of sex on cerebrospinal fluid density in adults. AB - The extent of sensory block during spinal anaesthesia is unpredictable and is influenced by many factors, mainly patient position, site of injection, baricity and the dose of drug injected. Among other factors, cerebrospinal fluid (CSF) density has been advocated to affect subarachnoid distribution of local anaesthetics. In this study, we have investigated the influence of patient characteristics such as sex, age, weight and height on variations in the density of CSF in more than 46 consecutive patients undergoing spinal anaesthesia. CSF 2 ml was obtained after spinal puncture and before injection of local anaesthetic. Mean CSF density measured at 37 degrees C was mean 1.00054 (SD 0.00017) g ml-1, with significantly lower CSF densities in women (1.00049 (0.00011) g ml-1) than in men (1.00058 (0.00011) g ml-1) (P = 0.024). In contrast, there was no correlation between age, weight or height, and CSF density. These results suggest that sex significantly influenced CSF density and may therefore modify subarachnoid distribution of local anaesthetics. PMID- 10700798 TI - Inhibitory effect of clonidine on ketamine-induced norepinephrine release from the medial prefrontal cortex in rats. AB - We have investigated the effect of clonidine on ketamine-induced norepinephrine release from the medial prefrontal cortex in rats using microdialysis. Twenty-one male Wistar rats weighing 200-300 g were allocated randomly to one of four groups: i.p. injection of ketamine 100 mg kg-1 with clonidine 0 (saline: group C0, n = 6), 3 (group C3, n = 5), 30 (group C30, n = 5) and 300 micrograms kg-1 (group C300, n = 5). As reported previously, ketamine increases norepinephrine release. In groups C0 and C3, marked increases in norepinephrine release were observed with maximum values of mean 483 (SEM 55)% and 412 (53)% compared with basal values, respectively. Although significant increases in norepinephrine release were also observed (276 (43)%) in group C30, they were significantly lower than those in groups C0 and C3 (P < 0.01 and P < 0.05, respectively). In group C300, there was a significant reduction in norepinephrine release (62 (13)%) compared with basal and the three other groups (P < 0.01). This inhibitory effect of clonidine on norepinephrine may be related to reduction in undesirable emergence reactions after ketamine anaesthesia. PMID- 10700799 TI - Effects of a nitric oxide scavenger, carboxy-PTIO, on isoflurane MAC and cerebellar nitric oxide synthase activity in rats. AB - Recent studies have indicated that nitric oxide may play a role in inhalation anaesthesia. Inhibition of nitric oxide synthase reduces the minimum alveolar concentration (MAC) for inhalation anaesthetics and decreases cerebellar nitric oxide synthase (NOS) activity in rats. In this study, we have explored further the role of nitric oxide in isoflurane anaesthesia by examining the effects of a nitric oxide scavenger, 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide (carboxy-PTIO 0.075-0.6 mg kg-1) on MAC values and cerebellar NOS activity in rats. Bolus injection of carboxy-PTIO at doses greater than 0.15 mg kg-1 reduced the MAC value of isoflurane (mean 1.36 (SEM 0.07)% at 0.15 mg kg-1, 1.39 (0.14)% at 0.3 mg kg-1 and 1.31 (0.06)% at 0.6 mg kg-1 vs control value of 1.61 (0.19)%. Administration of carboxy-PTIO 0.125 and 0.15 mg kg-1 resulted in increased cerebellar NOS activity during isoflurane anaesthesia (P < 0.05). These findings suggest that the level of nitric oxide may set a baseline from which isoflurane then acts. PMID- 10700800 TI - Perinatal management of a neonate with airway obstruction caused by rhabdomyosarcoma of the tongue. AB - Intra-oral masses in neonates can seriously compromise the airway, potentially causing hypoxia and death if not recognized and managed appropriately. We report a case in which an intra-oral mass was diagnosed on antenatal ultrasound scan. Preparation for delivery involved a multidisciplinary team approach, with a strategy for management at delivery. The child was delivered by elective Caesarean section and had a patent airway. A tracheostomy was performed immediately after delivery. The infant underwent a debulking procedure 3 weeks after birth. A histological diagnosis of embryonal rhabdomyosarcoma was made and a course of chemotherapy commenced. The child had a partial response to treatment with considerable shrinkage of the tongue mass. We discuss the management options in neonates with intra-oral masses to provide an adequate airway and maintain fetal oxygenation. The differential diagnosis of fetal oral masses is reviewed. PMID- 10700801 TI - Epidural anaesthesia for caesarean section in a patient with severe Takayasu's disease. AB - Takayasu's arteritis or disease is a rare, idiopathic, chronic inflammatory disease which causes narrowing, occlusion or aneurysms of blood vessels. It preferentially affects large arteries such as the aorta and its branches and hence its alternative names of pulseless disease, occlusive thromboaortopathy or aortic arch syndrome. Although most commonly found in oriental women, it occurs sporadically throughout the world. We present the case of an elderly primigravida with long-standing Takayasu's disease complicated by hospital and needle phobia who underwent a successful Caesarean section under epidural anaesthesia. Her management is discussed in the light of current opinion regarding pregnancy and Takayasu's disease. PMID- 10700802 TI - Percussion--a new way to diagnose a pneumothorax. AB - We describe a new clinical sign in a case series of three patients who developed pneumothoraces during mechanical ventilation in the intensive care unit. All three patients were in the supine position. Two patients had x-rays that were inconclusive before insertion of chest drains and the third had a pneumothorax diagnosed on clinical findings alone. On each occasion we were able to diagnose pneumothorax using sternal percussion and simultaneous auscultation. The method relies on percussion of the sternum while simultaneously ausculating the anterior (superior) chest on the side of the suspected pneumothorax. The stethoscope is then placed on the other side of the chest. The percussion sound on the affected side has an exaggerated, resonant and booming quality. The percussion note is exaggerated partly because a stethoscope is used and partly because, in the supine patient, air localizes upwards to the anterior thorax. PMID- 10700803 TI - Malposition of the epiglottis after tracheal intubation via the intubating laryngeal mask. AB - The intubating laryngeal mask has been reported to be a successful method of tracheal intubation although advancement of the tracheal tube via the laryngeal inlet into the trachea cannot be seen. Damage to the larynx or other tissues may occur during blind passage of a tracheal tube. We report a case in which the tracheal tube, advanced blindly, tucked the epiglottis into the laryngeal inlet, resulting in oedema of the epiglottis. This case illustrates the potential for airway obstruction after extubation when using the intubating laryngeal mask as a blind intubation guide. PMID- 10700804 TI - Portable ultrasound for central venous cannulation. PMID- 10700805 TI - Induction of anaesthesia in morbidly obese patients. PMID- 10700806 TI - Oxygen concentration distinguishes pulmonary from bowel gas leak. PMID- 10700807 TI - Transient ischaemic attack during deep cervical plexus block. PMID- 10700808 TI - The value of delta Down during haemorrhage. PMID- 10700809 TI - Subconjunctival block for cataract extraction and keratoplasty. PMID- 10700810 TI - Low-dose clonidine infusion during labour. PMID- 10700811 TI - Efficacy and safety of the superficial cervical plexus block for carotid endarterectomy. PMID- 10700812 TI - Videoendoscopic parathyroidectomy. PMID- 10700813 TI - Magnesium: physiology and pharmacology. PMID- 10700814 TI - Itching after use of starch solutions. PMID- 10700815 TI - Anatomy of the lumbar epidural region. PMID- 10700816 TI - Intracoronary artery radiation. PMID- 10700817 TI - Retreatment of cancer after radical radiotherapy. PMID- 10700818 TI - Renal duplication artefacts in ultrasound imaging. AB - A duplication artefact is sometimes encountered on ultrasound examination of the left kidney. It is caused by sound beam refraction between the spleen and adjacent fat. The purpose of this study was to analyse the frequency and types of such artefacts, and examine their physical background. The examiner searched for a left renal duplication artefact in 150 unselected abdominal ultrasound studies. A disturbed contour of the left kidney could be formed in 34 of 123 patients in whom the spleen was in the image without covering the kidney completely. An artefact was seen significantly more often when the lower pole of the spleen was rounded (30/87) than when it was wedge shaped (4/36) (p < 0.02). The artefacts could be categorized into complete upper pole duplication (n = 4), incomplete upper pole duplication (n = 14) and bayonet artefact (n = 16). The thickness of the renal cortex appeared reduced with upper pole duplication artefacts owing to a geometric image compression, which could also be seen in an in vitro experiment. Left renal duplication artefacts are not rare when the upper part of the kidney is examined by ultrasound through the spleen. PMID- 10700819 TI - MRI of arachnoid granulations within the dural sinuses using a FLAIR pulse sequence. AB - The purpose of the study was to assess the signal intensities of arachnoid granulations within the dural sinuses using the FLAIR sequence for differentiation of space-occupying lesions in and adjacent to the dural sinuses. We retrospectively reviewed MR images of the brain of 1118 consecutive subjects, ranging in age from 0 to 93 years (mean 57.2 years). Nodules within the dural sinuses with signal intensities similar to that of cerebrospinal fluid (CSF) on both T1 and T2 weighted images were defined as arachnoid granulations. The location, signal intensity on T1 weighted spin echo (SE), T2 weighted fast SE and FLAIR images, the impression on the inner table of the skull, and the size of the lesion were assessed. 112 subjects (10.0%), age range 4-89 years old (mean 58.9 years), were found to have 134 arachnoid granulations. The commonest location was the transverse sinus, with 115 granulations (85.8%). The prevalence of the granulations showed a peak in the sixth decade of age. All granulations were isointense relative to CSF on T2 weighted images and almost all lesions were isointense relative to CSF on T1 weighted images. On FLAIR images, 90.3% of the granulations were isointense relative to CSF and the other 9.7% granulations were slightly hyperintense compared with the CSF. 21 (15.7%) subjects showed impressions on the inner table; one case involved the outer table. In conclusion, arachnoid granulations were isointense or slightly hyperintense relative to CSF on FLAIR. FLAIR images are helpful in differentiating arachnoid granulations from other dural sinus lesions or skull lesions which have an intensity similar to that of CSF on T1 weighted and T2 weighted images. PMID- 10700820 TI - Imaging of recurrent esthesioneuroblastoma. AB - Esthesioneuroblastoma is an uncommon neoplasm arising from the olfactory epithelium and characterized by frequent local recurrences. The purpose of this study was to determine the role of CT and MRI in the diagnosis of recurrent esthesioneuroblastoma. A total of 14 histologically confirmed recurrent esthesioneuroblastomas referred to our institution between 1986 and 1998 was retrospectively reviewed. All patients underwent both CT and MRI. The tumour recurrences displayed a variety of imaging characteristics and aggressiveness. They were typically expansile and destructive in their growth patterns. Erosion of the cribriform plate and involvement of the anterior cranial fossa were common findings. The CT and MRI appearances of recurrent esthesioneuroblastoma do not differ significantly from tumours imaged at initial presentation. Patients should receive close follow-ups and CT/MRI examinations for several years beyond diagnosis, as early diagnosis of recurrent disease predicts survival. PMID- 10700821 TI - The effect of antihistamine, endothelin antagonist and corticosteroid prophylaxis on contrast media induced bronchospasm. AB - Bronchospasm is a well recognized adverse reaction to radiographic contrast media (RCM) and may occur more frequently in asthmatics and atopics. This study was designed to identify RCM which are most likely to cause bronchospasm and to investigate underlying mechanisms mediating this response. Guinea pigs (mean body weight 550 g, n = 46) were anaesthetized with Hypnorm (5 ml kg-1) and Hypnovel (2 ml kg-1) and tracheal, jugular and pleural cannulae introduced. Total airways resistance (Raw) was calculated from the slope of the pressure/flow relationship. The effects of RCM (diatrizoate 370 mgI ml-1, ioxaglate 320 mgI ml-1, iotrolan 300 mgI ml-1 and iopromide 300 mgI ml-1) at a dose of 4 ml kg-1 body weight or control solutions matched for volume, pH and osmolarity administered via the jugular vein on Raw were studied. The effects of pre-treatment (30 min before the administration of RCM) with antihistamine (Mepyramine (30 mg kg-1 i.p.)) or non selective endothelin receptor antagonist (SB209670 (1 mg kg-1 i.v.)) were investigated. The effectiveness of corticosteroids prophylaxis (prednisolone (20 mg kg-1 i.p.)) administered 18-24 h and 1 h pre-RCM was also assessed. Control animals received normal saline pre-treatment before RCM administration. Lungs were taken for histological examination 30-40 min post-administration of RCM. Only ioxaglate caused a significant (p < 0.05) increase in Raw (5.19 +/- 0.58 to 13.95 +/- 3.53 mmHg ml-1 min-1). Neither mannitol nor saline control solutions had any effect on Raw. Pre-treatment with Mepyramine, SB209670 or prednisolone caused no significant change in the ioxaglate induced increase in Raw. Histological examination of lung tissue from ioxaglate treated animals showed no important abnormalities. In summary, only the ionic dimer ioxaglate caused an increase in Raw. This effect was independent of osmolarity and could be the result of the chemical composition of the contrast agent. It was not an inflammatory response and could not be prevented by prophylactic treatment with antihistamine, endothelin antagonist or corticosteroids. The mechanisms responsible for the increase in Raw remain uncertain. PMID- 10700822 TI - Positron emission tomography in breast cancer: a clinicopathological correlation of results. AB - We conducted a retrospective study to evaluate the sensitivity, specificity and accuracy of positron emission tomography (PET) scans in 109 patients with primary recurrent or metastatic breast cancer. All patients had a PET scan, X-ray or CT scan of the chest, an ultrasound or CT scan of the liver and a bone scan. Mammography was available for 86 patients. Correlation between the PET scan result and histological findings were made. The sensitivity, specificity and accuracy of the PET scan were calculated for both the primary tumour (T) and lymph nodes (N). In patients with metastasis (M) the accuracy of the PET scan was compared with other imaging modalities. Histological results of the site in question were available in only 105 patients. Information for the primary tumour was available for 93 patients and for nodes in 74. The PET scan was accurate in 89.2% for (T), with 3.2% false positive and 7.6% false negative. For (N) the PET scan was accurate in 90.5% with 9.5% false negative. In the 86 patients who underwent both mammography and PET scanning, the PET scan was more accurate in 89.5% versus 72% (p = 0.0003). In the 19 patients with metastasis, the PET scan was in agreement with other imaging modalities in 100% of cases. PET scanning is the only non-invasive imaging procedure that will detect tumours in the breast, lymph nodes, lung, liver, bone and bone marrow with high sensitivity, specificity and accuracy. It is a valuable tool in the management of patients in all stages of breast cancer for diagnosis, staging and following treatment response. PMID- 10700824 TI - Radiation doses from CT in the Sultanate of Oman. AB - The computed tomography dose index (CTDI), dose-length product (DLP) and the effective dose were determined for a range of CT examinations in the Sultanate of Oman. There was a wide variation in CTDI. This shows that there is a variation in both scanner design and the exposure settings used by hospitals. There was also a wide variation in DLP and effective dose, suggesting that in some cases too many slices are taken. Therefore, standard protocols should be designed and adhered to in order that radiation doses may be reduced in the future. PMID- 10700823 TI - Gamma-ray scattering for mandibular bone density measurement. AB - There has been considerable interest in the measurement of bone density in the mandible, either to study bone resorption following tooth loss or to determine the relationship between mandibular and skeletal bone mineral density. Measurements mostly have been made on dental radiographs but more significant correlations between mandibular and skeletal bone density have been obtained when more sophisticated techniques have been used, such as quantitative CT and dual energy X-ray absorptiometry. The present work investigates the feasibility of using gamma-ray scattering measurements to estimate mandibular bone density. Using a phantom to simulate the jaw, an 241Am source and a hyperpure germanium detector it is shown that bone density may be measured with a precision of about 1%. PMID- 10700825 TI - Comparison of proton therapy and conformal X-ray therapy in non-small cell lung cancer (NSCLC). AB - This study compares the performance of one proton and four conformal X-ray planning techniques in treating non-small cell lung cancer (NSCLC). The treatment volumes for 13 NSCLC patients undergoing radical radiotherapy were planned using the five different techniques and dose-volume histograms (DVH) were used extensively in the comparative analysis. The minimum dose to the phase 2 target volume was escalated to 90 Gy, or until the point at which pre-set tolerance limits of spinal cord or lung were exceeded. The proton plan could treat nine of the 13 patients up to a dose of 90 Gy. Among the four X-ray techniques, performance varied enormously. One of them could not treat any of the patients, even to the conventional 60 Gy level, without failing to meet one or more of the criteria, whilst another one could treat 10 out of the 13 patients, although with this technique only four were permitted to have the dose escalated to 90 Gy. It was also found that two of the 13 patients could not be treated by any of the proton or X-ray plans to the conventional level, and were therefore considered unsuitable for radical radiotherapy. Various issues in conformal NSCLC radiotherapy including organ movement, tumour control, other possible organs at risk etc., are also discussed. PMID- 10700827 TI - Dynamic contrast enhanced magnetic resonance scanning as a predictor of response to accelerated radiotherapy for advanced head and neck cancer. AB - Tumour perfusion has been assessed in patients with advanced head and neck cancer using dynamic contrast enhanced MRI prior to and at completion of accelerated radiotherapy, and related to local tumour control. Sequential MRI scans, at 3 s intervals after intravenous injection of gadolinium using a dynamic scan sequence through a tumour region of interest (ROI), were performed in 13 patients with advanced head and neck cancer before and on completion of radiotherapy. The scans have been analysed in terms of maximum tumour enhancement (E), slope of the enhancement versus time curve and the time taken to reach maximum tumour enhancement (Tmax), and these parameters related to tumour outcome after radiotherapy. Local tumour control was related to the value of E on a post radiotherapy scan and the difference in Tmax between a pre- and post-radiotherapy scan. Durable local control was seen in those tumours with a post-radiotherapy value for E of less than 8 and a mean fall in Tmax of 27.3 s. These results imply that tumours with diminished tumour perfusion at the end of radiotherapy are those most sensitive to treatment and that those tumours which show greater tumour enhancement after accelerated radiotherapy are likely to fail locally. This may reflect the persistence of viable perfused tumour at completion of radiotherapy. PMID- 10700826 TI - An investigation into the use of polymer gel dosimetry in low dose rate brachytherapy. AB - An investigation has been carried out into the properties of the BANG polymer gel and its use in the dosimetry of low dose rate brachytherapy. It was discovered that the response of the gel was reproducible and linear to 10 Gy. The gel was found to be tissue equivalent with a response independent of energy to within experimental accuracy (standard error of measurement +/- 5%). The slope of the calibration curve was found to increase from 0.28 +/- 0.01 s-1 Gy-1 to 0.50 +/- 0.02 s-1 Gy-1 for an increase in monomer concentration from 6 to 9%. Absorbed dose distributions for a straight applicator containing 36 137Cs sources were measured using the gel and the results compared with measurements made with thermoluminescent dosemeters (TLDs) and calculated values. Good agreement was found for the relative measurements. The root mean square residual percentage errors were 3%, 1% and 4% for the gel and the two groups of TLDs, respectively. There were some significant differences in absolute values of absorbed dose in the gel, possibly owing to the effects of oxygen. Measurements of a complex gynaecological insert were also made and compared with isodose curves from a planning system (Helax TMS), and in areas unaffected by oxygen diffusion the isodose levels from 100 to 50% agreed to within less than 0.5 mm. PMID- 10700828 TI - Clinical implementation of a computer controlled milling machine for compensating filter production. AB - The procedures required for the clinical implementation of a computer controlled milling machine for producing compensators for breast radiotherapy are described. Moulds are cut in a rigid polymer foam block and filled with stainless-steel granulate. Quality assurance procedures are described for ensuring that the compensators produced are consistent and accurate. Relative and absolute dosimetric measurements are presented showing that the compensators are accurate to better than 1% and demonstrate the technique to be clinically acceptable. PMID- 10700829 TI - Determination of colony numbers in pig epidermis as an estimate for radiosensitivity. A rapid assay based on in vitro BrdU-labelling. AB - A rapid assay has been developed for the quantitation of colonies arising from surviving clonogenic cells in pig epidermis after irradiation. The number of surviving clonogenic cells per unit area was related to the epidermal in vivo response of moist desquamation. After irradiation with single doses, ranging from 20 to 36 Gy, skin biopsies were taken and incubated in dispase for enzymatic separation of the epidermis and dermis. Full thickness epidermal sheets were labelled with bromodeoxyuridine (BrdU) in vitro. Proliferating cells were visualized using standard immunohistochemical procedures. Cell groups containing > or = 16 cells were counted as colonies. These colonies were first seen on day 14/15 after irradiation. The number of colonies per cm2, as a function of skin surface dose, yielded a cell survival curve with a D0 (+/- SE) of 3.87 +/- 0.57 Gy. The ED50 for the epidermal in vivo reaction of moist desquamation corresponded with a colony density of 2.7 colonies per cm2. After higher doses, abundant smaller colonies of 4-8 BrdU-positive cells were seen and these were more radioresistant, as represented by higher D0 values. PMID- 10700830 TI - Repeated 131I treatment of a residual ovarian teratoma containing malignant thyroid tissue. AB - A 49-year-old woman with ovarian teratoma received 131I treatment three times for an unresectable mass containing malignant thyroid tissue after surgery. Repeated 131I treatment effectively reduced serum thyroglobulin (Tg) level and tumour uptake of 131I, despite absence of any change in size of the treated tumour. Treatment did not inhibit the increase of serum CA-125 and tumour 201Tl uptake, associated with progression of a radioresistant intratumoral hyper-perfused tissue component, detected by colour Doppler ultrasound. Serum CA-125 level and tumour 201Tl uptake were not significantly changed despite temporary increases in serum Tg level after each 131I treatment. These observations indicate the importance of diagnostic measures using combined functional imaging and tumour markers in managing this rare tumour. PMID- 10700831 TI - Conversion of multiple solid testicular teratoma metastases to fatty and cystic liver masses following chemotherapy: CT evidence of "maturation". AB - Testicular germ cell tumour metastases may undergo "retroconversion" to mature differentiated teratoma following chemotherapy or irradiation. We report a patient with testicular germ cell liver metastases in whom computed tomography (CT) scans following chemotherapy demonstrated a reduction in CT attenuation of the liver lesions to that of cystic and fatty density. This is believed to represent CT evidence of liver metastasis "retroconversion", which offers the potential for non-invasive monitoring of histological progression. PMID- 10700832 TI - Renal metastases from a squamous cell carcinoma of the larynx. AB - Laryngeal squamous cell carcinoma (SCC) tends to exhibit local spread with a low incidence of distal metastases. The majority of distal metastases are to the lungs and renal involvement is extremely rare. We present a case of laryngeal SCC with metastatic spread to the left kidney presenting as a large, mainly cystic mass. The radiological differentiation of renal metastases from primary renal tumours is discussed. PMID- 10700833 TI - Osteoid osteoma of a cervical vertebral body. AB - An osteoid osteoma in the anterior part of the body of the fourth cervical vertebra occurred in a 22-year-old female. The patient's main complaint was neck pain and occasional numbness of the extremities. The pain was relieved by analgesics. Plain radiography and a 99Tcm MDP bone scan showed a non-specific abnormality. CT suggested the pathological diagnosis with reasonable certainty. The unusual location of the lesion and the role of various diagnostic modalities are discussed and the literature reviewed. PMID- 10700834 TI - CT evaluation of gastric wall pathology. AB - The purpose of this study is to show the CT features of common and infrequent pathological lesions of the gastric wall. Although CT features are not often specific, familiarity with the most frequent pathological gastric findings on CT can assist in differential diagnosis. PMID- 10700835 TI - A case of hoarse voice. PMID- 10700836 TI - The most pressing issue. PMID- 10700837 TI - Primary care and public involvement: achieving a balanced partnership. PMID- 10700838 TI - Postmortem and perimortem caesarean section: what are the indications? PMID- 10700839 TI - Genetic and immunological therapy for cancer. PMID- 10700840 TI - Isolated corticotropin deficiency in chronic alcoholism. AB - Three patients who chronically abused alcohol were found to be hyponatraemic with normal plasma potassium. The first had been admitted with confusion and weight loss, the second with hypotension and sepsis, and the third with confusion and hypoglycaemia-induced seizures. All three patients had a subnormal cortisol response in the short synacthen test; however, the plasma cortisol after three days of tetracosactrin administration was greater than 550 nmol/L. Baseline corticotropin levels were less than 10 pg/mL in all three. No structural lesions of the hypothalamo-pituitary tract were found and there was no evidence of other endocrinopathies. Glucocorticoid replacement therapy led to the resolution of hyponatraemia and hypoglycaemia, where present, and to clinical improvement. The two surviving patients remained hypocortisolaemic in the long term, without recurrence of hyponatraemia or hypoglycaemia. The features of isolated corticotropin deficiency are easily confused with other effects of chronic alcohol abuse. In alcoholic patients with unexplained hyponatraemia, hypoglycaemia or haemodynamic instability, a short tetracosactrin test is advisable. PMID- 10700841 TI - Investigation and treatment of thyroglossal cysts in children. AB - Thyroglossal cysts are the commonest midline neck masses in children. To evaluate current practice questionnaires were sent to all ear, nose and throat (ENT) and paediatric surgeons in the UK and 72% responded. The commonest investigation requested was an ultrasound scan (54%) and the commonest operation was a variant of Sistrunk's procedure (78%). Paediatric surgeons did fewer investigations than ENT surgeons and tended to excise more of the thyroglossal tract. Review of the published work suggests that ultrasound scanning and Sistrunk's procedure are the best management policy. The scan can avoid inadvertent excision of an ectopic thyroid gland. Extensive thyroglossal tract excisions give lower recurrence rates. PMID- 10700842 TI - Osteoporosis services in secondary care: a UK survey. AB - A 1994 survey indicated that only 13 health authorities in the UK were purchasing access to dual X-ray absorptiometry (DXA), the most accurate measure of osteoporosis risk. By 1998 the number of centres (including private facilities providing DXA) was 161. All these were sent a questionnaire concerning their activities. 124 (77%) responded, and the survey found that DXA machines operate, on average, for only 3.6 days a week. Funding of and access to diagnostic services for osteoporosis varies greatly. There is clear scope for greater efficiency in the use of existing DXA machines and more equitable access to diagnostic services is required for effective management of osteoporosis. PMID- 10700843 TI - Hyperventilation due to mitochondrial myopathy. PMID- 10700844 TI - Cadaveric renal transplantation with simultaneous iliac bypass. PMID- 10700845 TI - Cerebellar syndrome complicating Mycoplasma pneumoniae pneumonia. PMID- 10700847 TI - The silicone gel breast implant controversy: the rise of expert panels and the fall of junk science. PMID- 10700846 TI - Pulsatile varicose veins. PMID- 10700848 TI - Age-related distortion. PMID- 10700849 TI - Medicine in the Millennium. PMID- 10700850 TI - Red jackets and red noses: alcohol and the British Napoleonic soldier. PMID- 10700851 TI - Professor Jeffray's grand tour. PMID- 10700852 TI - Do infections prevent asthma? PMID- 10700853 TI - Replacement of damaged neural cells. PMID- 10700854 TI - Placebo and placebo effects in medicine. PMID- 10700855 TI - Parkinsonism secondary to carbon monoxide poisoning. PMID- 10700856 TI - Management of acute bursitis. PMID- 10700857 TI - Unanswered questions about NICE. PMID- 10700858 TI - Autologous transplantation of in vivo purged PBSC in CML: comparison of FISH, cytogenetics, and PCR detection of Philadelphia chromosome in leukapheresis products. AB - To determine the effectiveness of different methods for the detection of tumor cell contamination of collected peripheral stem cells, we performed a study on 39 chronic myelogenous leukemia (CML) patients who were consecutively treated at our department. Analyses of tumor cell contamination by fluorescence in situ hybridization (FISH), conventional cytogenetics, and polymerase chain reaction (PCR) showed marked differences in the percentage of evaluable results: Quantitative analysis of tumor cell contamination was feasible in 60 of 105 (57%) samples evaluated with the use of conventional cytogenetic analysis and in 105 of 107 (98%) samples analyzed by FISH. PCR was evaluable in all 85 samples tested (100%). Both methods were shown to be adequate overall in determining the number of BCR-ABL positive cells, although cytogenetics tended to produce slightly higher percentages. Based on these results, we conclude that FISH performed on leukapheresis products is a rapid and reliable method for assessing the quality of these products and should be used for routine evaluation of tumor cell contamination of CML stem cell products. PMID- 10700859 TI - Molecular definition of a small amplification domain within 3q26 in tumors of cervix, ovary, and lung. AB - A common amplification target encompassing chromosome region 3q25 to q27 has been identified by comparative genomic hybridization analyses in tumors of the cervix, ovary, endometrium, lung, and head and neck. Because this segment spans at least 30 megabases, we undertook a molecular analysis of copy number to more precisely define the amplification domain. Our Southern blot and fluorescence in situ hybridization results with the use of 17 markers confirmed the presence of low level 3q amplification events in cervical, ovarian, and variant SCLC tumors. Most of the tumor types studied appeared to have similar, broad amplification domains centered within 3q26.2, suggesting that the same target is being affected in all. The ovarian carcinoma cell line NIH:OVCAR3 had a highly restricted amplification domain spanned by four overlapping YAC clones, suggesting a small target. The region of highest amplification included the gene for the RNA component of telomerase (hTR), supporting it as a potential target. Although the importance of low-level amplification is unknown, the consistent and reproducible nature of this event in a variety of carcinomas suggests that 3q26.2 harbors an oncogene whose low-level amplification has a significant influence on tumor biology. PMID- 10700860 TI - Translocation (1;11)(q23;p15), a novel simple variant of translocation (7;11)(p15;p15), in a patient with AML (M2) accompanied by non-Hodgkin lymphoma and gastric cancer. AB - We describe a case of an acute myelogenous leukemia (AML) associated with t(1;11) (q23;p15), which is a novel simple variant translocation of t(7;11)(p15;p15). The patient was a Japanese man who had a history of non-Hodgkin lymphoma (NHL) and received MACOP-B combination chemotherapy. Fifteen months after the completion of the treatment, the patient developed AML (M2), which was regarded as a therapy related leukemia. Cytogenetic study of bone marrow cells showed t(1;11). Although he achieved complete remission by combination chemotherapy, a relapse of NHL and gastric cancer were revealed in the course of the consolidation chemotherapy for AML. The NHL was considered a histological conversion from follicular lymphoma because lymphoma cells carried t(14;18) (q32;q21) and were strongly positive for BCL2 protein. Translocation (1;11), together with AML having t(7;11) or inv(11) involving 11p15, shows that 11p15 is a common acceptor site of these chromosome aberrations and suggests the significance of the NUP98 gene located in 11p15 in therapy-related leukemia. PMID- 10700861 TI - Analysis of MLL-derived transcripts in infant acute monocytic leukemia with a complex translocation (1;11;4)(q21;q23;p16). AB - It has been proposed, on the basis of cytogenetic studies and molecular analysis of MLL-derived transcripts in acute leukemia with 11q23 rearrangement, that only one fusion gene transcript present on the der(11) chromosome is critical for leukemogenesis. This view is challenged by a recent observation in a case of leukemia with a complex translocation that results in MLL being fused in-frame to two different partner genes. We investigated a case of infant monocytic leukemia with a complex translocation, (1;11;4)(q21;q23;p16). Molecular studies revealed MLL rearrangement by both fluorescence in situ hybridization and Southern blot analysis, and MLL/AF1q, but not the reciprocal message (i.e., AF1q/MLL), was amplified by polymerase chain reaction. Sequence analysis of MLL/AF1q revealed an in-frame fusion between MLL exon 6 and the breakpoint located six bases upstream of the ATG start site for AF1q. Our data suggest that only one form of MLL fusion gene is implicated in leukemogenesis in our case to t(1;11;4). PMID- 10700862 TI - Constitutional t(3;11)(p21;q23) in a family, including one member with lymphoma: establishment of novel cell lines with this translocation. AB - We describe a family with an inherited constitutional chromosome translocation (3;11) (p21;q23). Of three proven translocation carriers, one had duodenal malignant lymphoma (B-cell diffuse lymphoma, medium-sized cell type). The t(3;11)(p21;q23) was detected not only in hematopoietic cells including the patient's lymphoma cells, non-pathological bone marrow, and phytohemagglutinin stimulated peripheral blood, but also in fibroblasts of the skin. We have successfully established an Epstein-Barr virus-transformed B-cell line and a Herpesvirus saimiri-transformed T-cell line from the patient, and found that both cell lines also carried this translocation. The patient's asymptomatic mother and sister had the same chromosomal abnormality. Chromosomal abnormalities of the 11q23 band occur frequently in various hematopoietic malignant disorders, and 3q21 has been linked to the pathogenesis of several solid tumors including carcinomas of the kidney, lung, and breast. Although 11q23 is known to recombine with many different chromosomal segments, t(3;11)(p21;q23) has not been reported to our knowledge. Further assessment is warranted to clarify if this constitutional translocation predisposes to certain malignancies. Our cell lines carrying the novel chromosome translocation would be useful for the molecular analysis of the rearranged genes involving both 3p21 and 11q23. PMID- 10700863 TI - Loss of Y chromosome in bilharzial bladder cancer. AB - Bilharzial bladder cancer is the most common malignant neoplasm in Egypt, also occurring with a high incidence in other regions of the Middle East and East Africa. In a previous study, using centromere probes specific for chromosomes 3, 4, 7-11, 16, and 17, we demonstrated that monosomy of chromosome 9 (48.4%), and numerical aberrations of chromosome 17 (19.4%) were the most common observed imbalances. The present study extends the establishment of the baseline cytogenetic profile of this type of malignancy. Interphase cytogenetics by fluorescence in situ hybridization with the use of a panel of centromere associated DNA probes for chromosomes 1, 2, 5, 6, 12, 13/21, 14, 15, 18, 19, 20, X, and Y was performed on paraffin-embedded bladder specimens from 25 Egyptian patients affected with bilharzial bladder cancer. No numerical aberrations were detected in the 25 cases for chromosomes 1, 2, 5, 6, 12, 13/21, 14, 15, 18, 19, 20, and X. However, loss of chromosome Y was observed in 7 of the 17 male cases studied (41.2%). No significant correlation was observed between loss of the Y chromosome and any of the different clinicopathologic characteristics of these cases. These data suggest that loss of the Y chromosome is the second frequent event that can occur in bilharzial bladder cancer. A molecular genetic model of bilharzial bladder cancer is evolving. PMID- 10700864 TI - Structural analysis of PAX7 rearrangements in alveolar rhabdomyosarcoma. AB - In the pediatric cancer alveolar rhabdomyosarcoma, the common 2;13 and less frequent 1;13 translocations fuse PAX3 and PAX7, respectively, with FKHR to produce chimeric genes. To compare structural features of these rearrangements, we cloned and mapped a 64-kb genomic region containing PAX7 exons 5 through 8. With the use of Southern blot methodology, rearrangements of the 30-kb PAX7 intron 7 were detected in 9 of 9 PAX7-FKHR-positive cases. Similar to our t(2;13) studies, the t(1;13) breakpoints were randomly distributed within the seventh intron. In contrast with the > 90% frequency of reciprocal rearrangements in the t(2;13), reciprocal rearrangements involving the 3' PAX7 region were detected in only 4 of 9 cases. Furthermore, we detected PAX7-FKHR genomic amplification in 10 of 11 cases, in contrast with the < 5% frequency of PAX3-FKHR amplification. The differences in occurrence, reciprocity, and amplification between the PAX3-FKHR and PAX7-FKHR fusions indicate important differences in the mechanism of the two associated chromosomal translocation events. PMID- 10700866 TI - Fluorescence in situ hybridization confirmation of 5q deletions in patients with hematological malignancies. AB - Fluorescence in situ hybridization (FISH) using specific probes for the 5q31-32 region and a whole chromosomal painting (WCP) probe for chromosome 5 were used to corroborate the results of classical cytogenetic examinations performed on G banded chromosomes of 77 patients with hematological malignancies. Using classical cytogenetic methods, we suspected the presence of clones with a deletion 5q in 63 patients, and complex rearrangements with involvement of chromosome 5 in 14 other cases. Fluorescence in situ hybridization proved the occurrence of deletion 5q31 in 23 patients and ascertained translocations of part of the long arms of deleted chromosome 5 with missing region 5q31 in 12 patients. In 2 cases, the 5q31 region was translocated to other chromosomes as a part of complex rearrangements. The combination of classical cytogenetics and FISH with specific probes for the 5q31 band yielded cytogenetic results in 35 cases. Routine FISH detection of deleted regions was possible by commercially available cosmid probes for the 5q31 chromosomal band. The interpretation of small deletions and frequent involvement of the deleted chromosomes 5 in complex translocations were ascertained by WCP probes. PMID- 10700865 TI - Identification by comparative genomic hybridization of genetic changes involved in tumoral progression of a T-cell non-Hodgkin lymphoma. AB - Comparative genomic hybridization (CGH) was used to detect chromosomal imbalances in tumor DNA from two relapsed samples obtained in stages II and IV of a T-cell non-Hodgkin lymphoma in order to identify genetic mechanisms involved in tumor progression of this neoplasm. With conventional cytogenetic techniques (CCT), a complex hyperdiploid karyotype was obtained in stage IV. Using CGH analysis, a normal profile was observed in stage II, whereas gains of 6p11.2, 7q11.2, 7q21- >q32, 7q34, 10p13, Xp11.4, and loss of 4q33-->qter chromosomal regions were detected in stage IV. PMID- 10700867 TI - Detection of numerical chromosome anomalies in interphase cells of benign and malignant thyroid lesions using fluorescence in situ hybridization. AB - Benign and malignant thyroid tumors constitute a wide range of neoplasias showing recurrent chromosome abnormalities. In an attempt to characterize specific numerical chromosome abnormalities in thyroid tissues, we present here the findings from a study of archival samples depicted by 10 malignant tumors, 30 benign lesions, and 10 normal thyroid tissues. Fluorescence in situ hybridization was performed on noncultured samples using biotinylated centromere-specific probes for chromosomes 7, 10, and 17. Trisomy or tetrasomy 7 were present in 19 benign and in 7 malignant tumors. Trisomy 10 or 17 were observed in 18 adenomas or goiters and in 9 carcinomas, and monosomy 17 was seen in 2 carcinomas. Our findings suggest that such abnormalities are an in vivo phenomenon and may be important in the neoplastic proliferation of thyroid gland. PMID- 10700868 TI - Multiple myeloma: monoallelic deletions of the tumor suppressor genes TP53 and RB1 in long-term follow-up. AB - Recently, a working-model of a stepwise malignant transformation in the molecular pathogenesis of multiple myeloma (MM) was proposed, involving the tumor suppressor gene TP53 and retinoblastoma gene (RB1) as prominent components of cell cycle control. To further define the role of TP53 and RB1 in disease progression, we retrospectively analyzed by fluorescence in situ hybridization (FISH) cytological material from 16 patients who underwent sequential bone marrow biopsies during the course of their disease. For TP53, no deletions were detected at presentation or during follow-up. It is possible that the patients reported here represent a subset with relatively long survival, and therefore did not demonstrate the TP53 deletions that had been reported in patients with a very poor prognosis. For RB1, monoallelic deletion was demonstrated in nine patients. In each case, the deletion appeared already in the first biopsy analyzed. The presence of a deletion did not affect the rate of tumor progression or the length of follow-up, and thus prognosis. Monoallelic deletions of RB1 appear to be a frequent and early event in the pathogenesis of MM, without obvious relevance for disease progression. PMID- 10700869 TI - DNA replication errors are frequent in mucinous cystadenocarcinoma of the ovary. AB - To elucidate the genetic etiology of sporadic epithelial ovarian carcinoma, we examined replication errors (RER) in its two common types, serous cystadenocarcinoma (SAC) and mucinous cystadenocarcinoma (MAC). The subjects were 63 patients with sporadic epithelial ovarian carcinoma, consisting of 34 patients with SAC and 29 with MAC. The specimens were formalin-fixed and paraffin-embedded tissues from the ovary. The presence of RER was examined by polymerase chain reaction using 5 microsatellite markers. Of the 61 informative patients with ovarian carcinoma, RERs were observed at greater than or equal to 1 locus in 15 patients (25%). Four of the 32 SAC patients (13%) were RER-positive, and 11 of the 29 MAC patients (38%) were RER-positive, the incidence being significantly higher in the latter than in the former (P < 0.05). The incidence of RER was not related to the patients' age, stage, histologic grade, or prognosis. The MAC patients in stages I and II showed a relatively high incidence of RER (30%). These results suggest that RER are involved in the development and/or progression of MAC among sporadic epithelial ovarian carcinoma in its relatively early stage. PMID- 10700870 TI - A broad amplification pattern at 3q in squamous cell lung cancer--a fluorescence in situ hybridization study. AB - Frequent DNA copy number gain at 3q, with minimal overlapping area at 3q24-qter, has previously been reported in squamous cell carcinoma of the lung (SQCC), implicating the importance of genes at 3q in the tumorigenesis of SQCC. To further characterize the gain of DNA sequences at 3q, we performed interphase fluorescence in situ hybridization (FISH) analysis on 16 paraffin-embedded SQCC tumor samples that had previously been studied by comparative genomic hybridization (CGH). Eleven yeast artificial chromosome (YAC) probes located at 3q25-q27 and a chromosome 3-specific centromeric probe were used in the analysis. All SQCC tumors showed increase in DNA sequence copy number with 9-11 probes. In 5 tumors (31%) the number of centromeric signals varied from 3 to 5 and the YAC/centromeric signal ratio was 1.0, suggesting that the increase in DNA sequence copy number at 3q in these cases resulted from polysomy of chromosome 3. In 11 tumors (69%), the YAC/centromeric signal ratio varied between 1.5 and 4.7, indicating that the increase in DNA sequence copy number was due to intrachromosomal gain of DNA sequences at 3q. In each case, several YACs showed increased number of signals, demonstrating that the gained area was relatively large. Our findings therefore suggest that multiple genes located at 3q25-q27 are involved in the tumorigenesis of SQCC. PMID- 10700871 TI - Chromosome abnormalities in peripheral T-cell lymphoma. AB - Data on chromosomal abnormalities in T-cell lymphomas are very rare as compared with those reported in B-cell lymphomas. We performed a cytogenetic study in 71 untreated patients with peripheral T-cell lymphoma, classified according to the criteria of the REAL classification. Fifty-seven patients (80.3%) had abnormal clones, whereas 9 karyotypes (12.7%) showed only normal metaphases; 5 karyotypes (7%) could not be analyzed. Recurrent numerical chromosomal abnormalities comprised +3 (21%), +5 (15.7%), +7 (15.5%), +21 (14%), -13 (14%), +8 (12.2%), +19 (12.2%), -10 (10.5%), and -Y (9% of male patients). Chromosomes involved in structural rearrangements were chromosome 6 (31.5%), mainly due to 6q deletions (19.2%), 1q (22.8%), 7q (22.8%), 9p (19.4%), 9q (19.2%), 4q (19.2%), 3q (19.2%), 2p (17.5%), 1p (17.5%), and 14q (17%). Trisomies 3 and 5 mainly correlated with angioimmunoblastic T-cell lymphoma. Isochromosome 7q, associated with trisomy 8, was present in two cases of hepatosplenic gamma/delta T-cell lymphoma. Rearrangements involving the location of T-cell receptor genes were rarely observed (chromosome band 7q35 was rearranged only in three cases, 14q11 in two cases, and 7p15 in none). No correlation could be found between the cytogenetic findings and histologic subgroup or clinical outcome in these patients. Further studies are needed to understand the significance of these abnormalities in peripheral T-cell lymphoma, and to reach a better evaluation of histologic correlations, as many differences persist between the two major classification systems, KIEL and REAL. PMID- 10700872 TI - A new case of Turner syndrome associated with multiple myeloma. AB - A 62-year-old woman with phenotypic stigmata of Turner syndrome and a mosaic cytogenetic pattern, 45,X/45,XX, developed multiple myeloma. The affected cells had a number of karyotypic changes in addition to the loss of the X chromosome. PMID- 10700873 TI - Acute promyelocytic leukemia relapsing into FAB-M2 acute myeloid leukemia with trisomy 8. AB - Acute promyelocytic leukemia was diagnosed in a 48-year-old man; the karyotype was normal, whereas reverse transcriptase polymerase chain reaction (RT-PCR) analysis identified PML/RAR alpha chimeric transcripts of the bcr3 type. Rather unexpectedly, the patient did not respond to alltrans retinoic acid administration; he attained complete remission with conventional chemotherapy and became PML/RAR alpha negative. Two years later, while PML/RAR alpha negative on RT-PCR, he presented with thrombocytopenia. Bone marrow examination was compatible with myelodysplasia of the RAEB type; the karyotype was normal. Then, after 10 months, he developed overt acute myeloid leukemia with PML/RAR alpha negative, French-American-British M2 blasts; karyotypic analysis revealed mosaicism for trisomy 8. PMID- 10700874 TI - Prenatally diagnosed sacrococcygeal teratoma: a unique expression of trisomy 1q. AB - Partial duplication of the long arm of chromosome 1 has been observed in fetal intracranial teratomas. We sonographically diagnosed a 19-week fetus with sacrococcygeal teratoma, cerebral ventriculomegaly, and cerebellar hypoplasia. Chromosomal analysis of amniocytes showed an unbalanced translocation between chromosomes 1 and 15, resulting in trisomy 1q21-->1 qter. Duplication or over expression at more than one locus on the long arm of chromosome 1 may be required for the development of an extra-gonadal teratoma. PMID- 10700875 TI - [HIV: no virus eradication with HAART. Highly active antiretroviral therapy]. PMID- 10700876 TI - [Obstructive sleep apnea in older patients after the closure of a tracheostoma]. AB - BACKGROUND AND OBJECTIVE: The closure of tracheostomies may postoperatively induce an obstructive sleep apnea (OSA) even in patients who never complained about sleep related breathing disorders before they underwent tracheotomy. The present study was designed to evaluate, whether OSA appears after the tracheostomy has been airtightly blocked and if so, whether there are typically illnesses predictive for OSA after operative closure of tracheostomies. PATIENTS AND METHODS: Twelve consecutive patients (3 male; 9 female; mean age 66 years), who addressed our clinic for operative closure of their tracheostomies were enrolled in this study over a period of 18 months. On basis of patient's history and clinical findings the patients were divided into two groups. Group A included 7 patients with laryngeal pathologies and group B included 5 patients with normal larynges. All patients underwent a 12-channel polysomnography (PSG) with airtightly blocked tracheostomies preoperatively. RESULTS: In 5 patients a mild OSA and in one patient a severe OSA was diagnosed by PSG. The patients of group A with laryngeal pathologies were older, had a lower Body Mass Index (BMI) and a higher Apnea-Hypopnea Index than patients of group B (p < 0.05). CONCLUSION: Laryngeal pathologies in the elderly might facilitate the development of OSA after operative closure of tracheostomies. Based on the results postoperative reevaluation is recommended for all elderly patients with laryngeal abnormalities after operative closure of the tracheostomy. PMID- 10700877 TI - [Dilatation tracheotomy after Ciglia--its use in an internal-medicine intensive care unit]. AB - BACKGROUND AND OBJECTIVE: Since the first description of percutaneous dilatation tracheostomy (DTT), it has become an alternative method of equal value to surgical tracheostomy. This study collected the experience with DTT in a medical intensive care unit (ICU), with special reference to early and late complications and their management, outcome, and changes in ventilation parameters and blood gases. PATIENTS AND METHODS: Between March 1994 and April 1998, 74 DTTs were performed on 71 patients (52 men, 19 women; mean age 61.8 [30-80]) years. The admission or main diagnoses were cardiovascular disease in 34 patients, pulmonary disease in 21, the remainder having had a variety of conditions. RESULTS: The procedure caused complications in 21 procedures (28%): 10 cases of stomal bleeding (13.5% of total number of procedures), 2 of intratracheal bleeding (2.7%), 2 of severe tracheal injury (2.7%) and mediastinal emphysema in 1 (1.3%). None required intervention because of these complications. 38 patients were discharged from hospital. Cause of death in the other 33 was unrelated to the DTT. One patient developed tracheomalacia as a late complication. Ventilatory parameters and blood gases 12 hours post-DTT were the same as before the procedure. CONCLUSIONS: Ciaglia's method of dilatation tracheostomy is a safe procedure also in the context of a medical ICU, if the indications are correct and the procedure performed by experienced personnel. Compared with surgical tracheostomy it significantly reduces the burden on the patient as well as requiring fewer personnel and less equipment. PMID- 10700878 TI - [The Wiskott-Aldrich syndrome in adulthood]. AB - HISTORY AND ADMISSION FINDINGS: A 24-year-old man with thrombocytopenia was referred for surgical resection of a bleeding polyp of the sigmoid. Examination showed a small haematoma and petechiae on both lower legs. The patient reported that several male family members also had a thrombocytopenic bleeding tendency. INVESTIGATIONS: Laboratory tests revealed thrombocytopenia (4000 platelets/ml, with small platelets: mean platelet volume [MPV] 5.6 ml). Serum immunoglobulins were normal. A mutation in the Wiskott-Aldrich (W-A) protein gene (intron 7 + 5 G ->A) was demonstrated both in the patient and his 26-year-old brother. DIAGNOSIS, TREATMENT AND COURSE: The diagnosis of W-A syndrome was made and, with perioperative administration of platelets, the polyp was resected without complication. CONCLUSION: Most patients with the W-A syndrome die by the time they are aged 10 years, unless appropriate treatment is given. This patient and his brother had a mutation of the W-A protein gene that unusually was in an intron rather than in an exon. Structurally normal W-A proteins were still being formed. This may explain the mild course and late onset of the disease. PMID- 10700879 TI - [The diagnosis of hereditary colorectal carcinomas]. PMID- 10700880 TI - [The sympathetic nervous system and coronary heart disease]. PMID- 10700881 TI - [At the end only "muffled consternation"--the Great War in the medical press. 1914-1918]. PMID- 10700882 TI - [Ocular myasthenia gravis and autoimmune thyroiditis]. PMID- 10700883 TI - [Bacteriuria with an indwelling bladder catheter]. PMID- 10700884 TI - [On the tracks of humane medicine]. PMID- 10700885 TI - [Febrile neutropenia--guideline in Japan]. AB - Fever in neutropenic patients is mostly culture negative, with a rapid progress and high mortality. Due to the necessity of starting treatment early, guidelines for the treatment were first issued by IDSA (Infectious Disease Society of America). We have recently prepared a Japanese version of these guidelines. The criteria for febrile neutropenia are a fever above 38.0 degrees C (in mouth), granulocytes below 1,000. Patients must be checked by blood count, bacterial cultures of blood and urine, and chest X ray. Empiric treatment must be started early before the culture data become available. The first treatment is either single therapy of penem or beta-lactum, or the combination of one of these agents with aminoglucoside. Patients are evaluated after 72 hours. Treatment is modified to add or change to glycopeptide or antifungals. These guidelines still require study for confirmation and to determine the response rates of each treatment arm, as well as to find antibacterial drugs which are effective in culture negative patients with granulocytopenia. PMID- 10700886 TI - [Standard chemotherapy for gastrointestinal malignancies based on evidence]. AB - It has been said that there is no standard chemotherapy for gastrointestinal malignancies. However, standard guidelines are essential to increase the level of medical treatment, and the death rate from gastrointestinal malignancies is very high in Japan. FAMTX, standard therapy for gastric cancer abroad, cannot be standard in Japan due to its toxicities. A combination of 5-FU and cisplatin (FP) is most commonly used as the the first choice for chemotherapy, but it's regimens vary. For colon cancer, it is said that a combination of 5-FU and Leucovorin (LV) is standard, but CPT-11, made in Japan, is a promising agent. There is no recommended drug for advanced pancreatic cancer, so palliative care or no chemotherapy are also available alternatives. PMID- 10700887 TI - [Chemotherapy for head and neck cancer]. AB - Chemotherapy for head and neck cancer was initially used as a palliative treatment in advanced and/or recurrent disease. The overall response rate was about 30% but patient survival was sometimes short. It was also observed that complete responders had a significantly longer survival period than non responders. Cisplatin-containing regimens including cisplatin plus 5-fluorouracil appear to be the most efficacious for this disease. In a large number of randomized trials, organ function preservation studies have shown the possibility of laryngeal preservation for T2 and T3 laryngeal and hypopharyngeal cancer. A survival benefit has been shown clearly in advanced nasopharyngeal cancer. Another survival prolongation has been demonstrated in cases of locally unresectable cancer in the oral cavity, pharynx, nose and paranasal sinus. Thus, we conclude that neoadjuvant chemotherapy can be effective in cases of locally unresectable cancer in the oral cavity, pharynx, and nose and paranasal sinus. In advanced N stage nasopharyngeal cancer, neoadjuvant chemotherapy plus adjuvant chemotherapy may be indicated. Advanced T stage nasopharyngeal cancer is a good candidate for concurrent chemoradiotherapy. For the aim of laryngeal preservation, neoadjuvant and/or concurrent chemoradiotherapy can be indicated for T2 and T3 laryngeal and hypopharyngeal cancer. PMID- 10700888 TI - [Evidence-based medicine for urological cancer chemotherapy]. AB - We reviewed the treatment results of urological cancer chemotherapy from the standpoint of evidence based medicine. In the treatment of advanced transitional cell carcinoma of the urothelium, M-VAC (MTX + VBL + ADM + CDDP) is regarded as the standard regimen; however, durable event-free survival is rare. There is no level 1 evidence to date showing that the use of neoadjuvant or adjuvant cisplatin-based regimens will improve survival in cases of locally advanced bladder cancer. Immunotherapy with interferon or interleukin-2 produces a small survival advantage in patients with metastatic renal cell carcinoma. There is no evidence that adjuvant interferon-alpha administration will improve the survival in those with non-metastatic renal cell carcinoma. Systematized cisplatin-based treatment protocols have been established in patients with advanced testicular germ cell tumor by means of many randomized controlled trials. Several clinical trials are under way to prove the efficacy of high dose chemotherapy (with autologous stem-cell support) in patients with poor risk germ cell tumors. We do not yet have sufficient data to conclude whether maximal androgen blockade will prolong the survival in patients with metastatic prostate cancer, nor to conclude whether neoadjuvant androgen depletion treatment improve disease free survival of the patients after radical prostatectomy. PMID- 10700889 TI - [Evidence-based chemotherapy for patients with bone and soft part sarcoma]. AB - In the present paper, we review the evidence for chemotherapy in patients with bone and soft part sarcoma and discuss the contributions and improvements made by chemotherapy to the treatment of patients with bone and soft part sarcoma. In the osteosarcoma and Ewing's sarcoma family, neoadjuvant and adjuvant chemotherapy have improved the 5-year disease-free survival to 60%, and limb-salvage operations have improved this to 70-80% in cases of non-metastatic malignant bone tumor. Several trials were conducted in order to overcome rate relapses and metastatic bone sarcoma. With osteosarcoma, thoracotomy improved the survival of lung metastatic patients, but CDDP-ADM branch switched according to the neoadjuvant chemotherapy and failed to elevate the continuous disease-free survival of patients. Dose intensive use of cytotoxic drugs with G-CSF or autologous bone marrow transplantation and multidrug programs were conducted in preliminary studies and achieved favorable results in a high risk factors group for tumors of the Ewing's sarcoma family. Surgical techniques have brought improvements in the treatment of soft tissue sarcoma, but there has been no impact by chemotherapy. Ifosfamide and adriamycin combination is being evaluated in the treatment of local advanced and metastatic soft part sarcoma by local control rate or survival from relapse. PMID- 10700890 TI - [Evidence based chemotherapy for pediatric cancers]. AB - The curability of pediatric cancer has been improved to nearly seventy percent. This change has been achieved by refinements in treatment strategy and supportive care. More than seventy percent of patients with ALL can be cured by modern chemo radiotherapy with reduced late effects. The stratification of the patients by risk factor, introduction of CNS prophylaxis, shortening of the duration of chemotherapy and intensification of the chemotherapy with agents such as HD-MTX have contributed to this remarkable success. Burkitt's lymphoma is a tumor for which the curability has improved from almost zero to ninety percent. With Wilms' tumor, clinical trials have been used to optimally refine the treatment strategy. The NWTS first compared the efficacy of combined VCR and Act-D with the single use of each drug. The difference was significant. The results of the systematic trials were then used to improve the survival rate of patients with Wilms' tumor from twenty to ninety percent and shorten the duration of chemotherapy to six months. On the other hand, tumors remain with which less than half of patients can survive for long. Advanced neuroblastoma and AML are typical such tumors. With these diseases, refinements in the treatment based on evidence derived from clinical trials have been insufficient. Further intensification of the treatment or novel approaches to control tumor growth are warranted for these diseases. In this article, I would like to describe the "standard" therapy for each tumor and the evidence on which improvements in those strategies have been based. PMID- 10700891 TI - [Neoadjuvant chemotherapy for esophageal cancer by administration of nedaplatin alone]. AB - The possibility of safe implementation of neoadjuvant chemotherapy using nedaplatin (254-S) was investigated with the aim of improving the therapeutic results in advanced esophageal cancer patients. The subjects had usually undergone two courses of 254-S, 80 or 100 mg/m2, preoperatively, at intervals of 4 weeks. After chemotherapy, responses were evaluated, and resection was undertaken 4 weeks after the final administration. The subjects were 10 patients with untreated esophageal cancer. As a result of evaluation of responses of main lesions to chemotherapy, a partial response (PR) was observed in 3 patients, and a minor response 3, showing an efficacy rate of 30%. Pathological findings before treatment were determined by staining of a cell cycle marker (Ki-67). The 3 patients who were evaluated as PR (clinical efficacy) showed high rates of positivity for Ki-67 of 47.4%, 52.7%, and 86%, respectively. There were no serious complications and no death related to operation. The observations suggested that 254-S, which has little or no side effects including nephrotoxicity, can become the standard remedy for esophageal cancer instead of CDDP in future. PMID- 10700892 TI - [An early phase II trial combining cisplatin, carboplatin and vindesine in patients with inoperable non-small cell lung cancer]. AB - We conducted an early phase II trial of advanced non-small cell lung cancer (NSCLC) to evaluate the response efficacy of a combination of cisplatin (CDDP), carboplatin (CBDCA) and vindesine (VDS). The twenty-four patients in the study had had no previous treatment. CDDP (15 mg/m2), CBDCA (200 mg/m2) and VDS (3 mg/m2) were administered on Day 1, CDDP (15 mg/m2) was administered on Days 2-5, and VDS (3 mg/m2) was administered on Day 8. We observed 9 partial responses (PR), with a total response rate of 39%. The overall median survival was 72 weeks, and the 1-year survival rate was 57%. Major toxicities were hematologic; leukopenia of grades 3 and 4 occurred in 25% patients, and thrombocytopenia occurred in 21%. Therefore, the combination of CBDCA with CDDP and VDS chemotherapy was effective against inoperable NSCLC with tolerable toxicities and a favorable median survival time. PMID- 10700893 TI - [Preliminary clinical evaluation of low-dose CDDP and continuous 5-FU therapy for advanced gallbladder cancer]. AB - 5-fluorouracil (5-FU) has been widely used for the treatment of gastrointestinal cancers. Low-dose cisplatin (CDDP) and continuous venous infusion of 5-FU have recently shown additive or synergistic antitumor effects in experimental models. In this study, we evaluated the clinical effects of low-dose CDDP and 5-FU (low dose FP therapy) in patients with advanced gallbladder cancer. From December, 1993 to June, 1998, 13 patients with advanced gallbladder cancer were treated with low-dose FP therapy. Patients were eligible for this study if they had a bidimensionally measurable tumor. 5-FU (160 mg/m2/day) was continuously infused over 24 hours using an implantable port, and CDDP (3 mg/m2/day) was infused for one hour. The administration schedule consisted of 5-FU for 7 consecutive days and CDDP for 5 days followed by a 2-day rest, each for four weeks according to response and tolerance. Low-dose FP therapy was given to 12 patients (92.3%). The response rate was 66.7% and the median survival time was 151 days. The regimen was tolerable, with the most common toxicity being nausea (38.5%). There were no severe side effects except for one patient who suffered from grade 3 nausea. We conclude that low-dose FP therapy may be useful as a palliative chemotherapy for cases of advanced gallbladder cancer. PMID- 10700894 TI - [Clinical study of selective intra-arterial infusion chemotherapy using trans radial arterial approach in 4 cases of advanced breast cancer]. AB - We administered neoadjuvant chemotherapy by a selective intra-arterial infusion method using a trans-radial approach in patients with advanced breast cancer (stage III and stage IV). The trans-radial approach uses the arterial flow, based on the Seldinger technique. In this method, the radial artery is cannulated, and epirubucin is infused into the artery that carries blood from the subclavical artery to the breast. We have used this method in 4 cases thus far. Two of the patients received a single intra-arterial infusion of epirubicin. The other 2 patients were catheterized before they received chemotherapy by intra-arterial infusion. This technique decreased the pain or discomfort caused by the catheterizations during chemotherapy in all 4 cases. However, the currently available catheters are not always able to approach the artery flowing into the breast, thus the protocol will need to be refined. PMID- 10700895 TI - [Plasma concentrations of toremifene citrate and N-desmethyltoremifene in postmenopausal patients with breast cancer--comparison of 120 mg of toremifene citrate administered once a day and divided into 3 separate doses (t.i.d.)]. AB - To determine whether plasma concentrations of toremifene citrate after administration of 120 mg/day of toremifene citrate given in three separate dose (t.i.d.) were similar to those when toremifene citrate was administered in a single daily doses (40 mg x 3 tablets), we examined changes in plasma concentrations of toremifene citrate (TOR) and its metabolite, N desmethyltoremifene (TOR-1). In both the t.i.d. administration group and the single-dose administration group, plasma TOR and TOR-1 concentrations reached a constant state within 2 weeks after administration was started. Under the constant state, plasma TOR concentrations were 1,493.3 +/- 120.3 ng/ml in the t.i.d. administration group and 1,348 +/- 341.0 ng/ml in the single-dose administration group. Plasma TOR-1 concentrations were 2,378.3 +/- 186.5 ng/ml in the t.i.d. administration group and 2,144 +/- 475.3 ng/ml in the single-dose administration group. In both groups, plasma TOR-1 concentrations were 2 or more times higher than plasma TOR concentrations. These results show there were no differences in plasma concentrations between administration of 120 mg/day of toremifene citrate in divided daily doses (t.i.d.) and in a single daily dose. The two administration methods appear to produce clinically similar actions. PMID- 10700896 TI - [Substance isolated from the kelp rhizoid identified as L-tryptophan shows high inhibition of breast cancer]. AB - In general, the root of a seaweed is poorly developed as compared with its thallus and is called the rhizoid or holdfast. In Laminaria, belonging to Phaeophyceae, although the thallus is used for food, the rhizoid is considered an unuseful natural resource. We attempted to detect anti-breast cancer substances from that resource. As a result, a substance having a weak absorptivity to aluminium oxide and Sephadex G-25 was found. According to analysis of the FAB-MS spectra and 1H NMR spectra, the substance was identified as tryptophan, an amino acid. Finally, it was concluded by a chiral column-HPLC method that the tryptophan was the L-form. PMID- 10700897 TI - [Evaluation of paclitaxel and carboplatin in patients with endometrial cancer]. AB - The purpose of this study was to evaluate the combination of paclitaxel and carboplatin in patients with endometrial cancer who have high-risk histopathologic criteria with vessel permeation and low grade, advanced or recurrent disease. The combination of paclitaxel (180 mg/m2 over 3 hours) and carboplatin (dosed at an area under the curve of 5-6) was given intravenously every 3 weeks. Response and toxicity were evaluated according to the Japan Society of Clinical Oncology's response and adverse effect criteria. Eighteen patients were entered in this study and a total of 94 courses were administered. Eleven patients had evaluable lesions. Complete and partial responses were achieved in 5 (45.5%) and 3 (27.3%) patients, respectively. Grade 3 or 4 leukopenia and neutropenia occurred in 49.2% and 90.5% of the patients. G-CSF support was needed in 52.4% of the patients. Only one patient received a platelet transfusion. As a high response rate was obtained, this regimen is considered to be promising treatment for endometrial carcinoma. Prospective comparative study between this combination therapy and the conventional therapy for endometrial carcinoma is warranted. PMID- 10700898 TI - [Interim report on JFMTC Study no. 21 on the effectiveness of UFT as an adjuvant therapy for semi-advanced cancer of the stomach]. AB - This interim report, for findings as of May, 1998, covers data on 435 gastrectomized patients with semi-advanced stomach cancer collected from 144 institutions between November, 1993 and March, 1996. The active arm of the study involved CDDP i.p. administration of 70 mg/m2 at the time of resective surgery, followed by UFT oral administration for one year at 3-4 capsules daily. A randomized control involved no adjuvant therapy after CDDP i.p. administered as in the active arm. The results obtained indicated no significant difference between the groups in terms of 3 year survival or disease free survival rates. Reports appearing elsewhere have suggested that 3-4 capsules/day of UFT may be insufficient to reach the threshold of the effective tissue level, and that 6 capsules may be necessary to obtain the expected results. (JFMTC: Japanese Foundation of Multidisciplinary Treatment for Cancer). PMID- 10700899 TI - [A case of stage IVb (H2P0N4T4) gastric cancer successfully treated with neoadjuvant chemotherapy (PMFE therapy)]. AB - A 69-year-old man was examined at our hospital because of a sense of upper abdominal fullness. He was diagnosed as having stage IVb (H2P0N4T4) gastric cancer and treated with neoadjuvant chemotherapy. One course of the regimen consisted of 10 mg CDDP (day 1-5), 10 mg MMC (day 1), 250 mg 5-FU (day 1-20) and 50 mg ETP (day 6, 7). The patient underwent the regimen three times in succession. After the chemotherapy, his hepatic metastases showed necrotic changes and the swelling of the para-aortic lymph nodes disappeared on a CT scan. A histological examination revealed that the cancer cells had completely vanished both at the site of the hepatic tumor and the para-aortic lymph nodes. This combination chemotherapy, named PMFE therapy, is considered effective without serious side effects for gastric cancer in patients with non-curative factors. PMID- 10700900 TI - [A case of gastric cancer with liver metastasis responding to low-dose CDDP/5-FU combination chemotherapy]. AB - We reported a case of a 62-year-old female with gastric cancer accompanied by liver, Virchow and paraaortic lymph nodes, and bone metastasis (taken low-dose cisplatin (CDDP)/5-fluorouracil (5-FU) combination chemotherapy). CDDP (10 mg/body/day) was injected on 1-5 days i.v. and 5-FU (500 mg/body/day) was injected i.v. continuously on 1-7 days. This treatment cycle was repeated for 4 weeks. After 4 cycles, liver metastasis disappeared without severe side effects. Primary lesion and Virchow's lymph nodes metastasis were reduced. However, bone and paraaortic lymph node metastasis showed no response. It was considered that low-dose CDDP/5-FU combination chemotherapy was effective for liver and lymph nodes metastasis of gastric cancer in this case. PMID- 10700901 TI - [A case of advanced gastric cancer with hepatic metastasis successfully treated by combined chemotherapy with UFT and lentinan]. AB - We have experienced successful treatment of a hepatic metastasis of gastric cancer with UFT and lentinan. The patient was a 65-year-old male, who underwent distal gastrectomy for gastric cancer with hepatic and lymphatic metastases. After operation, administrations of UFT 300 mg/day and lentinan 2 mg/2 weeks were given, and the hepatic metastasis disappeared by 17 months. We performed a resection of the residual stomach and lymphatic metastasis at 52 months after operation. For over 5 years the patient has shown no evidence of a recurrence of the hepatic metastasis. This chemotherapy regimen was very effective and improved the patients quality of life. PMID- 10700902 TI - [A case of complete regression of liver metastases by treatment with Futraful supposition]. AB - The patient was a 57-year-old man abnormalities indicated in examinations by X ray and ultrasonography in February, 1991. X-ray and endoscopic examination revealed a Borrmann type 3 carcinoma in the posterior wall and lesser curvature of the upper body of the stomach. The liver was swollen to 3 fingerbreadths on the right mid-clavicular line. Multiple liver metastases were revealed by computed tomography (CT). Proximal gastrectomy was done. From March 24, 1991, a Futraful suppository (1,500 mg/day) was given daily. After 4 months, CT showed the reduction and partial disappearance of the low-density areas of the liver. After 2 years and 7 months, CT showed very small low-density areas, which completely disappeared by April, 1998. The patient has had a good quality of life. According to the General Rules for Gastric Cancer Study, the patient belongs to the class of complete response. PMID- 10700903 TI - [A case of diffusely infiltrating carcinoma of the sigmoid colon associated with lymphangitis carcinomatosa effectively treated with sequential MTX.5-FU and 5' DFUR]. AB - A 37-year-old man was diagnosed as having a diffusely infiltrating carcinoma of the sigmoid colon associated with lymphangitis carcinomatosa. Sequential methotrexate (MTX).5-fluorouracil (5-FU) therapy with oral administration of doxifluoridine (5'-DFUR) was started. After 9 cycles of the MTX.5-FU therapy (total dose: MTX = 900 mg/body, 5-FU = 7,500 mg/body), radiographic examinations showed a partial response in the primary and pulmonary lesion, and paraaortic lymph nodes. Histological evaluation of the resected specimen by Hartmann's operation showed a grade 2 effect in the primary lesion and metastatic lymph nodes. This chemotherapy was repeated postoperatively. The patient died of pulmonary disease, deteriorating rapidly 60 days postoperatively. Diffusely infiltrating carcinoma of the large-bowel is generally far advanced at the time of diagnosis. The results suggest that sequential MTX.5-FU therapy and oral administration of 5'-DFUR are worth performing in clinical trials for patients with diffusely infiltrating carcinoma of the large-bowel. PMID- 10700904 TI - [Complete remission in a case of sigmoid colon cancer with multiple liver metastases-treatment with arterial chemotherapy and percutaneous microwave coagulation therapy]. AB - A 59-year-old man was admitted to our hospital for advanced sigmoid colon carcinoma with synchronous multiple liver metastases. The patient received sigmoidectomy with regional lymph node dissection on June 8, 1998. We started intra-arterial combination chemotherapy on July 1, 1998. MMC (4 mg/body) was administered via rapid intra-arterial infusion on day 1. After MMC administration, 5-day intra-arterial continuous infusion of 5-FU at 500 mg/body/day was performed with oral administration of LV (30 mg/body/day). The treatment cycle was defined as every three weeks. The patient was treated with 4 courses of chemotherapy. From September 30, he received intra-arterial infusion of bolus MMC 4 mg/body, LV 6 mg/body and 5-FU 1,000 mg/body/4 hrs every two weeks with oral administration of Tegafur-uracil 400 mg/day. After 4 intra-arterial chemotherapy sessions, the metastatic liver tumors disappeared except for a focus in the right lobe. Therefore we decided to give the remnant liver metastasis percutaneous microwave coagulation therapy (PMCT). He obtained a complete remission in the liver metastases after two PMCT (70 W, 60 sec) sessions. Intra arterial chemotherapy is effective for unresectable metastatic liver tumors from colon cancer. If a patient shows a partial response on the metastatic tumors through the chemotherapy, one must consider other modalities such as PMCT. PMID- 10700905 TI - [A case of malignant fibrous histiocytoma of mesocolon successfully resected after combined chemotherapy with epirubicin, CDDP and vincristine]. AB - We experienced a case of MFH of the sigmoid mesocolon which was successfully resected after preoperative combined chemotherapy. A 66-year-old male underwent a laparotomy following a diagnosis of retroperitoneal tumor. The tumor had extensively invaded the surrounding tissue and an incisional biopsy was done. It was diagnosed as MFH histologically. Ten cycles of post-operative chemotherapy with doxorubicin were effective for a complete remission. However, the MFH reappeared at the same site after 2 years and 9 months. Three cycles of combined chemotherapy with epirubicin, CDDP and vincristine led to a regression of the tumor and no distant metastasis was found. The tumor was successfully resected with a negative surgical margin. It proved to be MFH of sigmoid mesocolon origin. The patient has been in good health for 7 years and 2 months after the second operation without further therapy. In cases of MFH such as the present in which the patient is sensitive to chemotherapy, neoadjuvant chemotherapy might be effective in allowing minimal surgery and offering a better quality of life. PMID- 10700906 TI - [Improved QOL with cancer chemotherapy in two patients with breast cancer suffering form carcinomatous pleurisy and carcinomatous peritonitis]. AB - One of the breast cancer patients introduced here suffered from recurrent carcinomatous pleurisy and the other from recurrent carcinomatous peritonitis. The patient with recurrent carcinomatous pleurisy was a 47-year-old female with stage IIIa breast cancer. She underwent a standard mastectomy and, following surgery, radiotherapy (50 Gy) and CAF therapy (30 mg of ADM, 1,800 mg of futraful and 100 mg of CPA, administered p.o.). Dyspnea occurred 4 years after surgery. Pleural exudate cytodiagnosis proved positive and the patient was diagnosed with carcinomatous peritonitis. Continuous thoracic cavity drainage was carried out, and 30 mg of ADM was injected into the thoracic cavity. CAF therapy was performed. The dyspnea and thoracic effusion disappeared. At present, after one year and 7 months, the patient is receiving outpatient treatment and remains under observation. The patient with recurrent carcinomatous pleurisy was a 43 year-old female. The breast cancer was detected in a diagnosis of metastasis to the axillary lymph nodes. An increased CA15-3 level and ascitic retention were observed postoperatively at 5 months. Following administration of 600 mg of UFT and 1,200 mg of MPA, the ascites decreased and improvement of the thickened peritoneum was noted. The CA15-3 level was also lowered. It is anticipated that chemotherapy for carcinomatous pleurisy and carcinomatous peritonitis will contribute to an improvement in patients' QOL. PMID- 10700907 TI - [A case of prostate cancer treated with combined androgen-blockade]. AB - A 62-year-old man was admitted to our hospital because of urinary retention. A digital rectal examination revealed an enlarged, elastic, and hard prostate with an irregular surface. Prostatic biopsy was done and the pathological diagnosis was moderately differentiated adenocarcinoma. He was diagnosed as stage D2 by bone scan and MRI. We gave him 250 mg of fosfestrol intravenously for 13 days. Afterward, he underwent orchiectomy castrate and was given flutamide for two years. His condition has improved and he has experienced no relapse. PMID- 10700908 TI - [Case report of a patient with invading bladder cancer and lymph node swelling which decreased following 5'-DFUR treatment]. AB - We report a 75-year-old man with invading bladder cancer and enlarged lymph nodes, which decreased following 5'-DFUR treatment. A postoperative abdominal CT showed lymph node swelling around the great vessels. The patient refused aggressive chemotherapy, so he was given a 5-FU derivative, 5'-DFUR, 800 mg/day orally. The enlarged lymph node had decreased in size by the 22nd day after the treatment, and the patient has maintained a near CR. PMID- 10700909 TI - [Combination of levofolinate calcium and 5-fluorouracil]. AB - It has been determined that in the combination of Leucovorin (LV) and 5 fluorouracil (5-FU), LV potentiates the cytotoxic effect of 5-FU based on biochemical modulation. Many data suggest that LV/5-FU is a very effective combination, and most clinicians worldwide now regard it as the standard therapy for colorectal cancer. In Japan, clinical examinations of this combination using Levofolinate calcium (I-LV) have shown its effectiveness, and I-LV/5-FU for gastric and colorectal cancer was authorized by the Ministry of Public Welfare in Oct. 1999. PMID- 10700910 TI - Studies on risk of leprosy relapses in China: relapses after treatment with dapsone monotherapy. AB - Based upon the data from the Chinese National System for Leprosy Surveillance, this paper reports on the relapses in 297,343 leprosy patients [multibacillary (MB) 106,518, paucibacillary (PB) 190,825] cured by dapsone monotherapy. A total of 11,055 (MB 8675, PB 2380) patients relapsed during an accumulated follow-up period of 4,229,050 patient-years (PY), giving an overall relapse rate of 3.72 per 100 cases or 2.61 per 1000 PY, i.e., 8.14% or 5.91 per 1000 PY over an average follow-up period of 13.8 +/- 8.4 years in MB patients and 1.25% or 0.86 per 1000 PY over an average period of 14.5 +/- 8.9 years in PB patients. For either the overall relapse rate per 100 cases or per 1000 PY, the differences between MB and PB patients were statistically significant, except during 36-40 years of follow up. For both MB and PB patients, the relapse rates showed consistently significant decreases year by year, particularly in PB patients whose relapse rate per 1000 PY was 1.21 in year 10 of follow up; whereas it remained more than 10 per 1000 PY in MB patients. In view of that, the overall relapse rates in MB and PB patients cured by dapsone monotherapy were acceptably low, and most of these patients have been followed up for more than a mean incubation period of observed dapsone relapse. Along with the further extension of follow up, the risk of relapse in dapsone-cured patients will not be expected to increase. This conclusion should be considered when planning policy for the management of patients released from dapsone monotherapy. PMID- 10700911 TI - Studies on risk of leprosy relapses in China: relapses after treatment with multidrug therapy. AB - Based upon the data from the Chinese National System for Leprosy Surveillance, this paper reports on the relapses in 47,276 leprosy patients cured by or released from WHO-recommended multidrug therapy (WHO/MDT). The overall relapse rate was 0.73/1000 patient-years (PY). There was a statistically significant difference in the relapse rates of WHO/MDT-MB (0.61/1000 PY) and WHO/MDT-PB (1.04/1000 PY) (chi 2 = 15.7, p < 0.01) patients. For multibacillary (MB) patients, the relapse rate in patients treated with fixed-duration MDT (0.56/1000 PY) was comparable with that in patients treated with MDT until skin-smear negativity (0.73/1000 PY) (chi 2 = 2.20, p > 0.05). Our present study suggests that fixed-duration MDT is a cost-effective regimen for the treatment of leprosy in China. The present results also show that relapse of leprosy is acceptably low and has not yet become a serious clinical or public health problem but, based upon the incubation of relapse in MDT patients, it is necessary to encourage annual follow up for at least 5 years for paucibacillary (PB) and 10 years for MB patients after being released from WHO/MDT. PMID- 10700912 TI - Hypopigmented face patches; their distribution and relevance to ocular complications in leprosy. AB - Eighty-two leprosy patients with hypopigmented patches over the face (cases) and an equal number of age-, sex-, and classification-matched leprosy patients without any hypopigmented patches over the face (controls) were examined for the distribution of hypopigmented facial patches, areas of anesthesia over the face, and eye complications. The hypopigmented patches did not follow any pattern and overlapped in the areas of sensation supplied by the three branches of the trigeminal nerve. Anesthesia over the face, evaluated by a Semmes-Weinstein monofilament which exerted a force of 0.05 grams, was present in 19.5% of the cases and 15.9% of the controls. Patients with hypopigmented facial patches were found to have more corneal hypoesthesia than patients who did not have hypopigmented facial patches. The risk of having impaired corneal sensation was three to four times higher in patients with hypopigmented facial patches. This feature can be used to identify decreased corneal sensation among leprosy patients under field conditions where direct estimation of corneal sensation is not advocated. PMID- 10700913 TI - Isolation, characterization, molecular cloning and amplification of a species specific M. leprae antigen. AB - A polyclonal serum sample from a lepromatous leprosy (LL) patient, which presented a specific recognition pattern for leprosin, was used to screen a Mycobacterium leprae genomic library constructed with DNA isolated from human lepromas. One clone, designated ML4-1, which expressed a specific antigenic determinant of M. leprae as part of a beta-galactosidase fusion protein, was isolated. The 1.932 bp M. leprae-derived genomic fragment was sequenced, and it had an incomplete open-reading frame shown to code for a 644 amino-acid polypeptide (72.3 kDa). Some partial nucleotide homology to the M. tuberculosis MTCY9C4 cosmid and the M. leprae B1913 cosmid were found. Southern blot assays using the 584 bp Eco RI-Bam HI fragment excised from the ML4-1 clone revealed that this sequence is present only in the M. leprae genome and not in the 24 different mycobacterial DNA tested. Two oligonucleotides based on the genomic sequence were also synthesized and used as amplifiers for a polymerase chain reaction (PCR) test, giving a positive signal exclusively in M. leprae DNA. Furthermore, 32 sequential synthetic peptides, 20 amino-acids long, spanning the entire protein corresponding to the hypothetical ML4-1 clone sequence, were synthesized and evaluated by ELISA. A peptide included in the 221-240 region was significantly recognized by either lepromatous leprosy or healthy tuberculosis contact patient sera. Thus, PCR amplification of this fragment, along with the recognition of its protein sequence by leprosy patient sera, could be a useful tool for a potential diagnostic method in the detection of M. leprae infection in the future. PMID- 10700914 TI - HLA linked with leprosy in southern China: HLA-linked resistance alleles to leprosy. AB - According to the World Health Organization recommended multidrug therapy (WHO/MDT), we have carried out this study to investigate the presence of HLA linked susceptibility or resistance to leprosy in a southern Chinese population. Sixty-nine leprosy patients and 112 healthy controls participated in the study. HLA-DR2 subtypes, HLA-B and MHC Class I chain-related A (MICA) alleles were typed at the DNA level using the polymerase chain reaction-single strand conformation polymorphism method. The frequencies of HLA-DR2-DRB1 alleles did not show any significant differences between the patient and the control groups, suggesting that the disease susceptibility was not associated with the DR2 subtypes in this southern Chinese population. On the other hand, in the multibacillary (MB) patients significantly decreased allele frequencies of HLA-B46 (0.040 in MB patients vs 0.129 in controls) and MICA-A5 (0.200 vs 0.380) were observed compared with the healthy controls. The calculated relative risk (RR) for B46 was 0.28; for MICA-A5, 0.52. In addition, on haplotype analysis the frequency of the HLA-B46/MICA-A5 haplotype was significantly decreased in the MB patients compared to controls (0.060 vs 0.233, RR = 0.22, p < 0.01). These results suggest that an HLA-linked disease-resistant gene to MB leprosy in southern China is in strong linkage disequilibrium with the HLA-B46/MICA-A5 haplotype. In other words, the resistant gene may be located near the HLA-B/MICA region and not in the HLA-DR locus. PMID- 10700915 TI - Histopathological activity in paucibacillary leprosy patients after ROM therapy. AB - Histopathological activity was assessed in the skin tissue of 13 skin-smear negative, borderline tuberculoid leprosy patients after administration of a single dose of ROM (rifampin 600 mg, ofloxacin 400 mg and minocycline 100 mg) therapy. Biopsies taken just before therapy showed Mycobacterium leprae to be present in eight cases. After 6 months, only three showed granulomatous lesions and others showed only resolving or inactive lesions. Acid-fast bacilli (AFB) persisted in the nerves of three cases. At the end of 12 months, granulomas persisted in 2 out of 13 (15%) patients. No bacilli, however, were detected in any of them at the end of 12 months. This study demonstrated that 12 months after a single dose of ROM granuloma cleared in 85% of the patients and AFB were absent in all of them. PMID- 10700916 TI - Serologic response to mycobacterial proteins in hansen's patients during multidrug treatment. AB - Humoral immune responses were studied in 24 leprosy patients treated with multidrug therapy (MDT) and 16 contacts. The patients were monitored for 2 to 3 years with repeated determination of IgG antibody levels directed to different mycobacterial proteins (Mycobacterium tuberculosis, Mt70; M. bovis, Mb65; M. leprae, Ml36, 28, 18, 10 kDa, and the complete protein M. leprae extract, MLSA). All recombinant antigens were used at 5 micrograms/ml concentration and the complete soluble M. leprae extract at 2 micrograms/ml. The results shown in this study reveal a clear decline in IgG antibodies directed toward mycobacterial proteins in the 12 multibacillary (MB) patients when they were submitted to MDT. Initially we found strong reactivity toward complete cytosolic protein and M. leprae membrane protein. The most reactive recombinant proteins in MB patients were Ml10, Ml36, Mt70 kDa and, finally, Ml18 kDa when compared to the paucibacillary (PB) group. After treatment was completed all lepromatous and borderline lepromatous patients showed low or undetectable levels as compared with their initial values before starting treatment. PMID- 10700917 TI - Determination of circulating IgG subclasses against lipoarabinomannan in the leprosy spectrum and reactions. AB - IgG subclasses against lipoarabinomannan of mycobacteria were analyzed in the sera of leprosy patients. Patients with active leprosy [tuberculoid and lepromatous, patients undergoing erythema nodosum leprosum (ENL) and reversal reactions] and inactive cases (tuberculoid and lepromatous who were cured after chemotherapy) were included in this study. Active lepromatous patients had higher levels of IgG subclasses, except IgG4, compared to active tuberculoid patients. Some of the inactive cases (lepromatous patients cured after chemotherapy) were positive for the IgG1, IgG2 and IgG3 subclasses. However, their levels are lower than active lepromatous cases. On the other hand, no difference in the subclass levels between the active and inactive tuberculoid groups could be observed. While a significant fall in the level of IgG3 in ENL was observed as compared to lepromatous leprosy without ENL, higher levels of IgG1 and IgG2 were found in patients with reversal reactions compared to their active counterparts without reactions. PMID- 10700918 TI - Cytodiagnosis of primary neuritic leprosy. AB - The diagnosis of primary neuritic leprosy (PNL) and its differentiation from other causes of peripheral neuropathy is difficult since acid-fast bacilli (AFB) smears and skin biopsy are negative from anesthetic areas. A biopsy of the involved nerve is the only conclusive method of diagnosis. Such a biopsy may not necessarily be free of complications when a large nerve is involved. However, fine needle aspiration has in this study proved to be a simple technique to demonstrate inflammation granulomas and AFB from these involved nerves in 18 of the 27 cases suspected to have PNL. The validity of the cytological classification into morphological subtypes may have to be supplemented by a large series of studies. PMID- 10700919 TI - Leprosy in hypertensive nude rats (SHR/NCrj-rnu). AB - Since more than a decade ago, we have attempted to develop spontaneously hypertensive rats carrying the nude gene that permits high multiplication of Mycobacterium leprae. A congenic strain carrying nude (rnu) and hypertensive genes was produced using SHR/NCrj females and F344/NJcl-rnu males. Cross intercross was carried out 12 times to establish the hypertensive nude rat congenic strain. As a result of the genetic monitoring test with NE12F2 generation rats, the genetic profile of the SHR/NCrj-rnu rats was the same as that of the SHR/NCrj rats except for the rnu gene. We have successfully developed a hypertensive congenic nude rat strain (SHR.F344Hfh11; SHR/NCrj-rnu). An increase in the blood pressure in nude rats was found to begin at a slightly delayed age when compared with their hairy litter mates. Both female and male rats showed the highest blood pressure at approximately 20 weeks of age--166 +/- 1.4 and 197 +/- 11 mm Hg in nude rats and 175 +/- 11 and 193 +/- 3.2 mm Hg in their hairy litter mates in female and male rats, respectively. In the present study, comparisons were made on the susceptibility to M. leprae in hypertensive SHR/NCrj-rnu and normotensive F344/NJcl-rnu rats. We have reconfirmed that hypertensive SHR/NCrj-rnu rats of the NE12F3 generation were highly susceptible to M. leprae. In the SHR/NCrj-rnu rats of both sexes excellent massive swelling due to multiplication of M. leprae was observed and, also, nodular lesions were produced in uninoculated fore feet and lips while those sites in the F344/NJcl rnu rats showed only a slight swelling of the inoculated feet with mild nodular lesions. Although mild lymphocyte proliferation was seen only in the M. leprae inoculated site with numerous bacilli and partial necrosis in the SHR/NCrj-rnu rats, at noninoculated sites, multiplication of M. leprae was only observed in the cells of the mononuclear phagocyte system. However, in F344/NJcl-rnu rats, lymphocyte proliferation with a few neutrophils was seen at the site of inoculated hind foot pads and everywhere at the site of multiplication of M. leprae. There was a wide difference in the susceptibility to M. leprae between the SHR/NCrj-rnu and the F344/NJcl-rnu rats. PMID- 10700920 TI - Sciatic nerve of normal and T200x5R Swiss white mice fails to support multiplication of intraneurally injected M. leprae. AB - In a preliminary study we have shown that freshly harvested Mycobacterium leprae, when injected into the sciatic nerve in normal and immunosuppressed (TR) mice, induce massive but localized epithelioid and macrophage granuloma, respectively, in 3-4 weeks. In order to determine the fate of M. leprae injected intraneurally into normal and TR mice, in the present study we measured sequentially the viability, fold increase and clearance, if any, using semi-quantitative methods. The average M. leprae yield per nerve assessed at regular intervals, beginning at 24 hr and including 72 hr, 1 week, 2, 3, 4, 12, 24 and 48 weeks, showed neither a significant increase nor a decrease in either the normal or the TR mice. The viability of M. leprae, assessed using the standard mouse foot pad method, showed a significant decrease as compared to baseline growth effective at 24 hr and remained static until approximately 4 weeks. A further decline and total loss of viability was noted by 12 months. The results show that injection of M. leprae via the intraneural route in both normal and TR mice failed to sustain the viability and failed to support the multiplication of the organisms. PMID- 10700921 TI - Do antibodies to phospholipid antigens play any role in murine leprosy? AB - In order to know whether antibodies to phospholipids and other host lipids play a role in the pathology of murine leprosy, we looked for the presence of antibodies to cardiolipin, cerebroside sulfatide, and to lipids extracted from normal murine spleen, liver and brain in the sera of mice bearing a 6-month infection with Mycobacterium lepraemurium. We also looked for the presence of antibodies to lipids isolated from M. lepraemurium. We found that all of the 16 animals examined contained high levels of antibodies to the mycobacterial lipids of intermediate polarity (mostly glycolipids) but none of them had antibodies to the other lipids tested, including those isolated from mouse liver, spleen and brain, bovine cardiolipin and sulfatide, nor any significant levels of antibodies to mycobacterial lipids of high or low polarity. The infected animals also had high levels of antibodies to antigens sonically extracted from the microorganism. Antibodies to the socially extracted antigens (mostly proteins) were mainly IgG, while antibodies to the lipid antigens were predominantly IgM. Despite the low but significant percentage (1%-3%) of infected animals developing bilateral paralysis of the rear limbs, autoimmunity (due to antibodies to phospholipids and other host lipids) does not seem to be a feature of murine leprosy. PMID- 10700922 TI - The paleoepidemiology of leprosy: an overview. PMID- 10700923 TI - Leprosy at the age of 141 years: a case report. PMID- 10700924 TI - Apoptosis in leprosy patients. PMID- 10700925 TI - Mitsuda-negative, resistant nine-banded armadillos and enhanced Mitsuda response to live M. leprae. PMID- 10700926 TI - Portraying a positive image of persons (previously) affected by leprosy. PMID- 10700927 TI - Prevalence rate of leprosy in Brazil. PMID- 10700928 TI - Trials of preventive therapy. PMID- 10700929 TI - Elimination of leprosy in the federated states of micronesia by intensive case finding, treatment with WHO/MDT and administration of chemoprophylaxis. PMID- 10700931 TI - Implementation of chemoprophylaxis in Chuuk State, Federated States of Micronesia. PMID- 10700930 TI - Implementation of chemoprophylaxis in Pohnpei State, Federated States of Micronesia. PMID- 10700932 TI - Monitoring the effects of preventive therapy in the Federated States of Micronesia. PMID- 10700933 TI - Population screening and mass chemoprophylaxis in Kiribati. PMID- 10700934 TI - Population screening and chemoprophylaxis for household contacts of leprosy patients in the Republic of the Marshall Islands. PMID- 10700935 TI - Vaccine trials in leprosy--Venezuela, Malawi and India. PMID- 10700936 TI - Preventive treatment of leprosy: needs, opportunities, and feasibility. PMID- 10700937 TI - Drugs and regimens for preventive therapy against tuberculosis, disseminated Mycobacterium avium complex infection and leprosy. PMID- 10700938 TI - The future of leprosy elimination. PMID- 10700939 TI - New biological tools for leprosy surveillance. PMID- 10700940 TI - Rationale for the preventive treatment of leprosy. PMID- 10700941 TI - Subclinical infection by Mycobacterium leprae. PMID- 10700942 TI - American Association of Dental Schools 77th annual session. Washington, DC, USA. April 1-5, 2000. Abstracts. PMID- 10700943 TI - Storage of tissues by vitrification. PMID- 10700944 TI - Development of hypothermic continuous perfusion preservation machine equipped with nonpulsatile pump and its clinical application. PMID- 10700945 TI - The suboptimal donor: reduction of ischemic injury in fatty livers by gaseous oxygen persufflation. PMID- 10700946 TI - Biochemical effects and cyclic-AMP second messenger signal upon venous oxygen persufflation of ischemically preserved livers. PMID- 10700947 TI - Autooxidation of glutathione in organ preservation solutions. PMID- 10700948 TI - Warm and cold ischemia result in different mechanisms of injury to the coronary vasculature during reperfusion of rat hearts. PMID- 10700949 TI - Effect of acute rejection on expression of fibrosis associated genes in renal transplant recipients. PMID- 10700950 TI - Microcirculatory perfusion pattern during harvest of livers from non-heart beating donors: beneficial effect of warm preflush with streptokinase. PMID- 10700951 TI - Heparin/phentolamine does not improve kidney perfusion with HTK solution after prolonged warm ischemia in a rat non-heart-beating donor model. PMID- 10700952 TI - Liver preservation of non-heart-beating donors is improved by additional portal venous organ perfusion. PMID- 10700953 TI - Free radicals and apoptosis of the endothelial cells. PMID- 10700954 TI - Kupffer cell activation in liver preservation: cold storage vs machine perfusion. PMID- 10700955 TI - Swine pancreas preservation with Celsior solution. PMID- 10700956 TI - Supplemental reduced glutathione during cold ischemia does not improve early renal allograft function. PMID- 10700957 TI - Comparison between University of Wisconsin and Celsior solution on morphology and viability of rat aorta after cold storage. PMID- 10700958 TI - Celsior solution and clinical liver transplantation. PMID- 10700959 TI - Impact of limited cold ischemia on renal function. PMID- 10700960 TI - Evaluation of ex vivo renal function following prolonged cold ischemia. PMID- 10700961 TI - Isolated rat heart mitochondria and whole rat heart as models for mitochondrial cold ischemia-reperfusion injury. PMID- 10700962 TI - Limitation of lipid peroxidation and renal medullary cell injury of the kidney after 48- and 72-hour cold storage in University of Wisconsin solution: effect of trimetazidine. PMID- 10700963 TI - Evaluation of renal medulla injury after cold preservation and transplantation: noninvasive determination of medullar damage by proton nuclear magnetic resonance spectroscopy of urine and plasma. PMID- 10700965 TI - Mitochondrial oxidative injury in rat fatty livers exposed to warm ischemia reperfusion. PMID- 10700966 TI - School education, a basis for positive attitudes toward organ donation. PMID- 10700964 TI - Tauroursodeoxycholate reduces ischemic damage in human allografts: a biochemical and ultrastructural study. PMID- 10700967 TI - Efficacy of double filtration plasmapheresis pretreatment in clinical ABO incompatible transplantation and experimental pig-to-baboon xenotransplantation. PMID- 10700968 TI - Suppression of porcine xenoantigen expression by dominant-negative effect of alpha-1,3-galactosyltransferase (alpha-1,3-GT) splicing variants. PMID- 10700969 TI - Sharing of split livers between centers is easily feasible. PMID- 10700970 TI - Characterization and influence of risk factors on initial liver function after transplantation. PMID- 10700971 TI - Effects of an educational segment concerning organ donation and transplantation. PMID- 10700972 TI - Predictors of the intention to donate organs: an empirical model. PMID- 10700973 TI - True organ donor potential: a retrospective single-center study. PMID- 10700974 TI - How local transplant coordinators might increase the number of potential donors: our experience in lower silesia. PMID- 10700975 TI - Procurement of all the donor corneas needed: how is it achieved? PMID- 10700976 TI - A modified "Spanish model" for organ donation in the southeast region of Sweden. PMID- 10700977 TI - Organ donors with adequately treated bacterial meningitis may be suitable for successful transplantation. PMID- 10700978 TI - Ethical implications in donor-recipient allocation. PMID- 10700979 TI - Do prior open heart procedures affect the outcome after heart transplantation? PMID- 10700980 TI - Current arrangement and activity of organ transplantation after new organ transplant legislation in Japan. PMID- 10700981 TI - Reasons for turning down an offer of a kidney for transplantation. PMID- 10700983 TI - Need for a systematic approach to organ recovery: technology development for the non-heart-beating donor. PMID- 10700982 TI - Compliance with kidney allocation criteria in the Czech Republic in 1997. PMID- 10700984 TI - Evaluation of the efficiency of organ procurement and transplantation program. PMID- 10700985 TI - Double lung transplantation in cystic fibrosis patients: perioperative hemodynamic-volumetric monitoring. Rome Lung Transplantation Group. PMID- 10700986 TI - Arterial reconstruction in hepatic and pancreatic allograft transplantation following multi-organ procurement. PMID- 10700987 TI - Potential of interventional ventilation in organ transplantation. PMID- 10700988 TI - Gentle in situ liver manipulation during organ harvest increases oxygen consumption in liver. PMID- 10700989 TI - Living-related versus living-unrelated kidney transplantation using tacrolimus initial immunosuppression. PMID- 10700991 TI - Evaluation of live kidney donors with CT renal angiography: an experience with 38 cases. PMID- 10700990 TI - A step-by-step approach to laparoscopic live donor nephrectomy. PMID- 10700992 TI - Donor selection in a living related renal transplant program--an analysis of donor exclusion. PMID- 10700993 TI - Donor registration campaign: ministry of Public Health involves personal request to 12.2 million Dutch citizens > or = 18 years. PMID- 10700994 TI - Cancer in transplant recipients. PMID- 10700995 TI - Delayed graft function incidence as predictive variable of survival of kidney grafts retrieved from elderly donors. PMID- 10700996 TI - Successful transplantation of an injured liver. PMID- 10700997 TI - Polycystic liver grafts for orthotopic liver transplantation. PMID- 10700998 TI - The impact of donor age on renal graft survival. PMID- 10700999 TI - High-efficiency kidney transplantation: concept, technique, results, and cost analysis. PMID- 10701000 TI - Non-heart-beating donor transplantation: machine perfusion preservation is the answer to prolonged cold ischemia time. PMID- 10701001 TI - Our vision on organ donation in developing countries. PMID- 10701002 TI - Sensitive detection of cytomegalovirus infection in transplant recipients using nucleic acid sequence-based amplification. PMID- 10701003 TI - Mycobacterial infection in renal transplant recipients. PMID- 10701004 TI - Early detection of cytomegalovirus in renal transplant recipients: comparison of PCR, NASBA, pp65 antigenemia, and viral culture. PMID- 10701005 TI - Renal tubular epithelial cells tolerate prolonged warm ischaemia in vitro. PMID- 10701006 TI - Distribution and predictive value of apoptosis in biopsies from non-heart-beating donor kidneys. PMID- 10701007 TI - Usefulness of high-risk renal graft conditioning: functional improvement of high risk grafts by addition of reagents to continuous hypothermic perfusion preservation solution. PMID- 10701008 TI - Correlation between troponin I serum level and acute cardiac allograft rejection: a preliminary report. PMID- 10701009 TI - Lecithin cholesterol acyltransferase activity following orthotopic liver transplantation. PMID- 10701010 TI - Pretransplant evaluation of renal viability by glutathione S-transferase in machine perfusate. PMID- 10701011 TI - A useful predictor in machine perfusion preservation for kidney transplantation from non-heart-beating donors. PMID- 10701012 TI - Prospective evaluation of renal function. PMID- 10701013 TI - Potential prognostic indicators of liver function. PMID- 10701014 TI - Viability testing in the non-heart-beating donor. PMID- 10701015 TI - Interstitial fibrosis in the cortex of donor kidneys: relationship to donor type and posttransplant function. PMID- 10701016 TI - Excellent long-term graft survival with kidneys from the uncontrolled non-heart beating donor. PMID- 10701017 TI - Combined intravascular and intraperitoneal cooling in the non-heart-beating donor improves kidney function following transplantation. PMID- 10701018 TI - Influence of delayed graft function in renal transplants from cadaveric or non heart-beating donors. PMID- 10701019 TI - Comparison of fibrosis-associated genes after renal transplantation from cadaveric and non-heart-beating donors. PMID- 10701020 TI - Weekly protocol renal transplant biopsies allow detection of sub-clinical acute rejection episodes in patients with delayed graft function. PMID- 10701021 TI - Heart preservation of non-heart-beating donors by in situ perfusion: comparison of artificial oxygen carrier perfluorocarbons with University of Wisconsin solution in a big animal model. PMID- 10701022 TI - Preliminary analysis of a randomized trial comparing microemulsion cyclosporine and tacrolimus for recipients of renal transplants from non-heart-beating donors. PMID- 10701023 TI - Modification of the treatment of gout in renal transplant recipients. PMID- 10701024 TI - Orthotopic liver transplantation with poor neurologic outcome in valproate associated liver failure: a need for critical risk-benefit appraisal in the use of valproate. PMID- 10701025 TI - Telemetric system for ambulatory lung function analysis in transplanted patients. PMID- 10701026 TI - HOE 077 reduces fibrotic overgrowth around the barium alginate microcapsules. PMID- 10701027 TI - Effect of fibrin glue coating on the formation of new cartilage. PMID- 10701028 TI - Time-dependent difference of the 12-hour trough levels of cyclosporine in renal transplant recipients with sandimmune capsule. PMID- 10701029 TI - Effect of low-density lipoprotein-apheresis on nephrotic syndrome due to membranous nephropathy in renal allograft: a case report. PMID- 10701030 TI - Organic insecticides. AB - Organic insecticide poisoning continues to be a major health problem not only in the developing communities but also in the Western population. The insecticides commonly used are the organophosphates, organocarbamates, organochlorides and pyrethroids. Patients with organic insecticide poisoning present with a spectrum of manifestations ranging from gastrointestinal symptoms of nausea, vomiting and diarrhoea to severe neurological manifestations of fasciculations, seizures and neuromuscular weakness and paralysis or cardiac manifestations of arrhythmias and conduction disturbances. A strong clinical suspicion is necessary to make an early diagnosis of insecticide poisoning. Treatment is primarily supportive and includes decontamination, protection of airways and cardiac and respiratory monitoring. Specific therapy for organophosphates and organocarbamates includes the use of anticholinergics. The use of oximes, especially high dose, is controversial and may be associated with a higher mortality rate. Low-dose oximes given early in the course of the illness may be beneficial. This paper reviews the literature on organic insecticide poisoning worldwide. PMID- 10701031 TI - Morphine-sparing effect of ketoprofen after abdominal surgery. AB - In a double-blind, placebo-controlled clinical trial (power of 80% to detect a 30% reduction in morphine consumption, P < 0.05), we have determined that the administration of two doses of intravenous ketoprofen 100 mg, one at the end of surgery and the second 12 hours postoperatively, was associated with a significant reduction in morphine consumption at eight (P = 0.028), 12 (P = 0.013) and 24 hours (P = 0.013) but not four hours (P = 0.065) postoperatively, as compared to placebo, when assessed by patient-controlled analgesia. There was no difference between the groups in pain scores or in the incidence of nausea and vomiting. One patient in the placebo group suffered from excessive sedation while one patient from the ketoprofen group suffered from transient oliguric renal failure. There were no other adverse effects. The results of this study show that ketoprofen does provide a morphine-sparing effect in the management of postoperative pain after abdominal surgery. PMID- 10701032 TI - Effect of timing of ondansetron administration on incidence of postoperative vomiting in paediatric strabismus surgery. AB - This prospective, randomized, double-blinded study evaluated the effect of the timing of ondansetron administration on its antiemetic efficacy in children undergoing elective strabismus surgery. One hundred and twenty children aged one to 15 years, ASA physical status 1 or 2, were randomly allocated to receive intravenous ondansetron 100 micrograms/kg either at induction (Group 1) or at the end of the surgery (Group 2). All patients had general anaesthesia induced and maintained with nitrous oxide and halothane, muscle relaxation with vecuronium, endotracheal intubation, reversal with neostigmine and glycopyrrolate, and pethidine 0.5 mg/kg analgesia. Episodes of nausea and vomiting were evaluated at 0 to 2, 2 to 6 and 6 to 24 hour intervals by a blinded observer. Demographic data, duration of anaesthesia, type of surgery, incidence of previous postoperative nausea or vomiting and motion sickness and number of patients who developed oculocardiac reflex requiring atropine treatment were similar in both groups. The incidence of emesis in the first 24 hours following surgery was similar in both groups (35% Group 1, 33.3% Group 2, P = 1.00). Severity of emesis (median number of emetic episodes, rescue antiemetic requirement and mean time to the onset of first episode of emesis) and mean time to discharge from the post anaesthesia care unit were also similar in the two groups. We conclude that the timing of ondansetron administration either before or after the surgical manipulation of extraocular muscles had similar antiemetic efficacy following strabismus surgery in children. PMID- 10701033 TI - The accuracy of in vivo P50 at high haemoglobin saturation. AB - The in vivo P50 (P50iv) provides a useful index of haemoglobin-oxygen affinity and is calculated according to software algorithms incorporated into commercial blood gas analysers. These algorithms are known to be inaccurate at high haemoglobin saturation (SpO2 > 97%) although just how inaccurate has not been documented. This study examines the arterial blood gas profiles of patients admitted to a busy secondary referral Intensive Care Unit and stratifies them according to haemoglobin saturation in order to quantify the accuracy and potential clinical utility of the Siggaard-Andersen algorithm (SAA) for assessing P50iv in blood with SpO2 > 90%. Sicker patients, as identified by plasma pH < 7.35 or [lactate] > 2.0 mmol/l, were substratified and the SAA assessed as before. In both groups, the results show not only that the SAA is completely unreliable above 97% saturation, a fact acknowledged by Siggaard-Andersen in 1984, but it is also inaccurate in the range 92% < or = SpO2 < or = 97%, thus rendering P50iv calculations suspect in 90% of the patients in each of the study groups. PMID- 10701034 TI - Sedation for vitreoretinal surgery: a comparison of anaesthetist-administered midazolam and patient-controlled sedation with propofol. AB - Local anaesthesia is increasingly being used for vitreoretinal surgery, but the optimal technique for sedation remains unclear. Anaesthetist-administered midazolam, which is often used, was compared in this study to patient-controlled sedation with propofol in 43 patients undergoing 50 vitreoretinal procedures. A variety of patient, anaesthetist and surgical endpoints were measured. There were no significant outcome differences between the two agents except that midazolam produced more amnesia for the local anaesthetic eye block. However, several outcomes and the observations in patients who experienced both agents showed a trend in favour of propofol for intraoperative sedation. We conclude that both approaches are safe and that patient-controlled sedation with propofol is at least as satisfactory as anaesthetist-administered midazolam. PMID- 10701035 TI - The use of orbital morphine for postoperative analgesia in pterygium surgery. AB - A prospective double-blind study compared the analgesic effectiveness of peribulbar lignocaine with peribulbar morphine and lignocaine for postoperative analgesia in pterygium surgery. Twenty patients were randomly divided to receive a peribulbar injection preoperatively of either 1% lignocaine 2 ml or 1% lignocaine 1.6 ml and 4 mg morphine. Effects on pain at injection and pain at 24 hours and 48 hours postoperatively were measured with a visual analog pain scale. Effects of the injections on sedation, pupil size and unwanted side-effects were also recorded. The groups were comparable for age, weight and surgical technique. There was a significantly lower pain score at 24 hours after operation in the morphine group (P = 0.035). There were no significant differences in sedation or side-effects between the groups. The physiological effects of morphine on the eye are reviewed. The study suggests that orbital morphine may be an effective and safe form of analgesia for corneal surgery and further investigation is warranted. PMID- 10701036 TI - The effect of previous administration of nizatidine on the neuromuscular effects of vecuronium and the effect of nizatidine on gastric secretion. AB - Nizatidine, a new H2-receptor antagonist, has been reported to inhibit acetylcholinesterase activity. This could lead to an interaction with neuromuscular blocking drugs. This study examined the effects of nizatidine on the actions of vecuronium. Oral nizatidine has been reported to be an effective protective agent against acid aspiration syndrome, and we reevaluated this effect. The control group (n = 10) received a placebo with water 50 ml and the nizatidine group (n = 10) received nizatidine 300 mg with water 50 ml two hours before arrival in the operating room. Gastric contents were aspirated and the volume and pH measured before induction of anaesthesia. Anaesthesia was induced in all patients with thiopentone 5 mg/kg and 1.5% isoflurane in 98.5% oxygen followed by vecuronium 0.1 mg/kg. Vecuronium onset time and duration time 25 (time from injection until recovery of 25% of vaseline twitch amplitude) were obtained using electromyography. There were no significant differences between the two groups in vecuronium onset time or duration time 25. Gastric fluid volume was greater and gastric pH was lower in the control group than in the nizatidine group. 70% of the control group and none of the nizatidine group (P < 0.005) had a gastric content pH < 2.5 or volume > 25 ml. PMID- 10701037 TI - Retroperitoneoscopic excision of phaeochromocytoma--haemodynamic events, complications and outcome. AB - Over a period of 15 months, 11 patients with phaeochromocytoma underwent retroperitoneoscopic excision of their tumours. Five patients had bilateral tumours. All patients underwent thorough preoperative evaluation and preparation with alpha- and beta-blockade. In the majority of the patients a hypertensive response was seen during generation of pneumoretroperitoneum. However, the period of tumour dissection and excision was devoid of large haemodynamic fluctuations. The average time taken was 3.5 to 4 hours per gland. Blood loss in successful laparoscopic excision averaged 240 ml (range 120 to 700 ml). In these patients satisfactory postoperative analgesia could be provided with intramuscular pethidine or intramuscular diclofenac sodium. In three patients the procedure had to be converted to open laparotomy due to haemorrhage. All three patients had preoperative radiological evidence of inferior vena cava and aortic involvement. Patient selection plays an important role in a successful outcome. PMID- 10701038 TI - Generic polymerase chain reaction followed by DNA sequencing as a means of diagnosing bacteraemia. AB - There is increasing use of polymerase chain reaction techniques to diagnose infection. We report the use of polymerase chain reaction using a generic section of the bacterial 16S rDNA gene--followed by nucleotide sequencing--to determine the species of the infecting bacteria. In the first case, the clinical and microbiological diagnosis of meningococcal septicaemia was in agreement with the results from polymerase chain reaction technique. In the second case, a Yersinia enterocolitica bacteremia was detected by the polymerase chain reaction technique, but missed with conventional blood culture techniques. PMID- 10701039 TI - Sevoflurane anaesthesia with the Komesaroff vaporizer inside the circle system. AB - This study assessed the safety of sevoflurane anaesthesia using two Komesaroff vaporizers inside the circle with both spontaneous and controlled ventilation. Sevoflurane concentrations were continuously monitored using a mass spectrometer and the anaesthetic depth was easily controlled. Involuntary movements occurred in eight patients and breath-holding occurred in five patients after inhalational induction. With continuous monitoring of sevoflurane concentrations, induction and maintenance of anaesthesia using a vaporizer inside the circle is safe during both controlled and spontaneous ventilation. However the high incidence of involuntary movements may limit the feasibility of induction in spontaneously breathing patients. PMID- 10701040 TI - Cricothyroidotomy and transtracheal high frequency jet ventilation for elective laryngeal surgery. An audit of 90 cases. AB - We carried out an audit of needle cricothyroidotomy and transtracheal ventilation used during anaesthesia for elective endolaryngeal surgery. The data on 90 consecutive procedures was collected over two years. Patients were anaesthetized using a total intravenous technique. An intravenous cannula or Tuohy needle was placed through the cricothyroid membrane and the patient was ventilated via the cannula using high frequency jet ventilation. Technical details of the procedure and any perioperative complications were recorded. There were 12 complications in total. Only three of these were clearly related to the cricothyroid puncture, i.e., one minor bleed and two cases of limited local surgical emphysema. All complications were minor and resolved without sequelae. PMID- 10701041 TI - The accuracy and reliability of an infusion pump (STC-3121; Terumo Inst., Japan) during hyperbaric oxygenation. AB - We evaluated the accuracy of delivery of one type of infusion pump at three nominal flow rates under two environmental pressures (measured in atmospheres absolute, ATA), one of which is used for hyperbaric oxygenation. Overall, we observed significant differences between the two different pressures (1 ATA and 2 ATA). Changes in the rate of delivery of between about -4% and +2% were observed over the time course of hyperbaric oxygenation, and the inter-group difference was significant in the phases in which pressure was changing. The effect produced by 2 ATA was statistically significant but small, and its influence is likely to be negligible during practical hyperbaric oxygenation therapy. Thus, it would seem that this type of pump would deliver reliably at the nominal rate during hyperbaric oxygenation at these levels. PMID- 10701042 TI - Postobstructive pulmonary oedema--a case series and review. AB - Six cases of post-extubation pulmonary oedema in otherwise healthy patients are reported. All were preceded by an episode of laryngospasm and followed a clinical course similar to that previously documented in cases of post-obstructive pulmonary oedema. Frank haemoptysis was a feature of five of the presentations. One patient was reintubated and ventilated, two were admitted to the intensive care unit for mask CPAP, one was managed with CPAP in the recovery ward and two with supplemental oxygen only. All cases resolved fully within 24 hours. Some evidence points to the syndrome being the result of airway bleeding rather than true pulmonary oedema. The literature suggests that it occurs more commonly than is generally thought, with a frequency of 0.05 to 0.1% of all anaesthetics, and is often unrecognised or misdiagnosed. Most cases occur in the early postoperative period, so anaesthetists are well placed to witness, investigate and manage this interesting condition. PMID- 10701043 TI - Postoperative fibrinolysis diagnosed by thrombelastography. AB - Thrombelastography is a useful method for the monitoring of bedside coagulation, especially for fibrinolysis. We report a case where thrombelastography facilitated early detection of fibrinolysis with significant clinical bleeding in a patient immediately following hip replacement surgery. The early diagnosis enabled institution of antifibrinolytic therapy and monitoring of the patient's response. It is likely to have led to less blood product transfusion and may possibly have prevented unnecessary surgical re-exploration. PMID- 10701044 TI - Congenital myasthenic syndrome: a rare, potentially treatable cause of respiratory failure in a "floppy" infant. AB - A four-month-old infant, thought to suffer from cerebral palsy, presented with respiratory failure on the background of a gradually deteriorating general level of function. Whilst being ventilated in intensive care he was noted to have severe muscle weakness. A disorder of the neuromuscular junction was suspected and he was subsequently demonstrated to have a congenital myasthenic syndrome. Anticholinesterase therapy produced a dramatic recovery. The congenital myasthenic syndromes and the diagnosis of a "floppy baby" are briefly reviewed. PMID- 10701045 TI - Streptoccocus pyogenes: a forgotten cause of severe community-acquired pneumonia. AB - We report a case of severe community-acquired pneumonia caused by Streptococcus pyogenes (Lancefield Group A streptoccocus) that was complicated by a streptococcal toxic shock syndrome. Although this micro-organism is an uncommon cause of community-acquired pneumonia, previously well individuals may be infected and the clinical course may be fulminant. A household contact was the likely point of infection. Invasive group A streptococcal disease continues to remain an important cause of morbidity and mortality in the community and therefore will continue to be encountered by intensive care physicians. Treatment of Group A streptococcal infection remains penicillin; however, clindamycin should be added in severe infection. PMID- 10701046 TI - Fixed, dilated pupils in the ICU: another recoverable cause. AB - A 41-year-old woman presented unconscious with fixed dilated pupils following a massive overdose of carbamazepine and an unknown quantity of venlafaxine prescribed for the management of bipolar affective disorder. Her course in the intensive care unit was marked by a number of complications related to the overdose including prolonged coma, seizures and cardiac arrest. The patient eventually recovered to leave hospital. PMID- 10701047 TI - Tuberous sclerosis presenting for laparotomy. AB - A 30-year-old female patient with tuberous sclerosis presented for anaesthesia and surgery for haemorrhagic renal angiomyolipoma. The anaesthetic management of this case was tailored to the prevention of seizures. Diagnostic features and possible complications of the disease are also described. PMID- 10701049 TI - Turning the epidural needle. PMID- 10701048 TI - Diabetes, blood glucose and preoperative evaluation. PMID- 10701050 TI - Tuohy needle deformation. PMID- 10701051 TI - Hyperkalaemia and massive transfusion. PMID- 10701052 TI - Cerebral blood flow and tourniquet release. PMID- 10701053 TI - Fatal epidural infusion. PMID- 10701054 TI - [Do children and adolescents need supplements during puberty of calcium and vitamin D?]. PMID- 10701055 TI - [From rheumatism nodosa in children to idiopathic juvenile arthritis or from Diamantberger to the Durban criteria]. PMID- 10701056 TI - [For what reasons is the neurologic status of very premature infants altered between 1 and 2 years in a follow-up study? The contribution of a Frache-Comte region study]. AB - BACKGROUND: The purpose of this population-based study was firstly to compare the neuro-developmental outcome at one and two years of very preterm infants, and secondly, to identify the risk factors for a misdiagnosis of cerebral impairment at the age of one year. POPULATION AND METHODS: The preterm cohort included 203 infants born between 25 and 32 weeks of gestational age in the region of Franche Comte (France) during a two-year period. The control group included 196 full-term infants born in the same maternity wards. Neuro-developmental assessments were performed by pediatricians or physicians, both at one and two years of age, on 94% (161/171) surviving preterms and 89% (173/195) full-terms. RESULTS: There is a fair correlation between the two neurological evaluations of the control group (170/173, 98% have the same classification at the age of one and two). There is a weak correlation (kappa = 0.37) between the two neurological evaluations of the preterm group. Sixteen preterms (10%) had been classified more abnormal at one year than they were at two years. The presence of a broncho-pulmonary dysplasia, linked to male sex and extreme prematurity, was statistically linked to this first kind of misclassification. Seventeen preterms (10%) had been considered more normal at one year than they were at two years. The presence of a diplegia, family precariousness and the examination at one year of age by a general practitioner were statistically linked to this second kind of misclassification. CONCLUSION: This prospective population-based study identifies structural situations (bronchopulmonary dysplasia linked to extreme prematurity) and environmental situations (family precariousness, examiner's qualifications) linked to a misclassification of the neurological status of one-year-old former preterm infants. PMID- 10701057 TI - [Congenital hernia of the diaphragm. A retrospective study of 123 cases recorded in the Neonatal Medicine Department, URHC in Lille between 1985 and 1996]. AB - BACKGROUND: During the last ten years, new therapeutic strategies have been used in order to improve the management of congenital diaphragmatic hernia (CDH). CDH is associated with pulmonary hypoplasia, abnormal pulmonary vascular reactivity and pulmonary immaturity. Between 1985 and 1990, mechanical hyperventilation and early surgery were provided systematically. Since 1991, the management of CDH in our institution has involved a preoperative stabilization with exogenous surfactant replacement, gentle ventilation, high-frequency oscillation, nitric oxide or extracorporeal membrane oxygenation. PURPOSE: To analyse the impact of the new therapeutic strategy on the survival and outcome of newborns with CDH. METHODS: Retrospective review of all infants with CDH admitted to our institution from 1985 through 1996. Mortality and morbidity were compared between period I (1985-1990) and period II (1991-1996). RESULTS: Between 1985 and 1996, 123 neonates were admitted to our Neonatal Department. Nine of them had another severe congenital malformation and were excluded from the study. Survival was 23% (12/52) in period I and 56% (35/62) in period II (p < 0.001). In period II, complications were more frequent among survivors in whom an extracorporeal membrane oxygenation was required (13 infants): bronchopulmonary dysplasia 77% (10/13), gastroesophageal reflux 61% (8/13), and hypotrophy 61% (8/13). CONCLUSION: These data demonstrate a significant improvement in survival in CDH since the implementation of new therapeutic modalities. Nevertheless, a significant morbidity exists among the infants who survive a severe respiratory failure. PMID- 10701058 TI - [The incidence of prescriptions without marketing product license in a neonatal intensive care unit]. AB - Most of the drugs prescribed in pediatric units have no product licence. The lack of clinical studies in children and appropriate drug formulations decrease their safety. The lack of a legal framework makes the prescriber insecure. Even if the debate is not recent, few studies have been carried out in this field. The aim of the present study was to evaluate the rate of prescriptions of unlicensed and off label drugs in a neonatal intensive care unit. PATIENTS AND METHODS: The present study was carried out in our neonatal intensive care unit, from January 12 to February 12, 1998. Forty babies aged 0 to 128 days were included (90% newborns), with a gestational age between 25 to 40 weeks (88% were premature, with a birth weight lower than 1000 g). RESULTS: Two hundred and fifty-seven prescriptions were administered with 55 different types of drugs during this period. Ten percent of the prescribed drugs had no product licence. Sixty-two percent were off-label for premature infants and 64% for newborns: 90% due to age, 9.3% due to dose and 0.7% to method of administration. No therapeutic alternatives to these prescriptions were found among the few available licensed drugs. CONCLUSION: The prescriptions of unlicensed and off-label drugs in neonatal intensive care units are daily and repeated events. The prescribers are usually not aware of the exact status of the drug and do not realize neither he importance of the problem nor the legal and potential consequences. The lack of pediatric clinical studies is to a large extent responsible for the absence of drug registration in pediatrics. The pharmaceutical industry has few incentives to develop the pediatric product licences. PMID- 10701060 TI - [Acute Salmonella typhi meningitis in a 25-day-old newborn infant complicated by obstruction of the sylvian artery]. AB - Acute Salmonella typhi meningitis is rare in neonates, mostly reported from developing countries with poor socioeconomic conditions. CASE REPORT: A male Caucasian newborn presented with acute Salmonella typhi meningitis at the age of 25 days. His parents had traveled across several African countries under rudimentary hygienic conditions a few months before his birth. Despite early and adapted antibiotic therapy (cefotaxime plus netilmycine), the child developed ischemia in the region of the left sylvian artery. CONCLUSION: Salmonella meningitis must be considered while dealing with a sick newborn whose mother has traveled across countries with endemic typhoid. PMID- 10701059 TI - [Prevention of vitamin D deficiency in adolescents and pre-adolescents. An interventional multicenter study on the biological effect of repeated doses of 100,000 IU of vitamin D3]. AB - Recent studies have shown a high prevalence of calcium and vitamin D deficiencies in adolescents. The aim of this present study was to follow the changes in calcium status and 25 hydroxyvitamin D (25[OH]D) and parathyroid hormone (iPTH) levels during winter in preadolescents and adolescents from four university hospitals in northern France. SUBJECTS AND METHODS: Two groups of teenagers and adolescents (range: 10-15 years) were followed from October 1996 to June 1997. They were given either 100,000 IU of vitamin D (treated group n = 33) or a placebo (control group n = 35) in October, January and April. Serum calcium, phosphate, 25(OH)D and iPTH levels were measured at inclusion and every three months thereafter. RESULTS: At inclusion, plasma or serum 25(OH)D levels were < or = 10 ng/mL in 16 subjects and < 6 ng/mL in six. In control children, no significant change in 25(OH)D occurred during the study, while plasma or serum iPTH levels increased to 34 +/- 11 pg/mL. In the treated groups, 25(OH)D levels remained > 20 ng/mL in every subject; no hypercalcemia was observed; and the mean plasma or serum iPTH level was 25 +/- 14 pg/mL at the end of the study. CONCLUSION: Teenagers presented with a high prevalence of biological vitamin D deficiency at the end of summer. The increase of iPTH during winter in the unsupplemented group suggests that this has secondary consequences on their calcium homeostasis unless they are supplemented with vitamin D. We advocate a sufficient calcium supply and a 100,000 IU vitamin D supplement given two or three times during winter to preadolescents and adolescents living in northern France. PMID- 10701061 TI - [Heterozygous protein C deficiency: apropos of 2 cases with cerebral venous thrombosis in the neonatal period]. AB - Thrombotic accidents in the newborn, particularly cerebrovascular accidents, are reported in case of abnormalities in the coagulation system and rarely in heterozygous protein C deficiency; a low protein C level could be either physiological or acquired. CASE REPORT: Two cases of heterozygous protein C deficiency are reported in neonates. Severe neurologic distress was associated with bloody cerebrospinal fluid, and hemorrhagic lesions due to cerebral sinovenous occlusion were visualised by cerebral imaging. The course was severe. One case was associated with renal thrombosis. Mutation in the 168 proline/leucine was detected by molecular biology in the neonates and their mothers. In one case a treatment with protein C had no beneficial effect. CONCLUSION: Cerebral sinus venous thrombosis has to be sought by magnetic resonance imaging in the case of neurologic distress with profound cerebral hemorrhage in the newborn. A low level of protein C has to be interpreted with caution. The diagnosis of a heterozygous deficiency status can only be made through molecular biology. The effect of treatment with protein C concentrate is questionable. PMID- 10701062 TI - [Effectiveness and dangers of interferon-alpha in the treatment of severe hemangiomas in infants]. AB - Alarming hemangiomas, due to their site or repercussions, require pharmacological treatment. Corticosteroid therapy is indicated by first intention. In the event of failure, interferon alpha is proposed. CASE REPORTS: Case 1. A five-week-old infant was admitted to hospital for an extensive hemangioma of the left side of the face and neck with necrosis of the upper lip and ear. Prednisolone (2 mg/kg/day) by intravenous route brought about no improvement. Interferon alpha 2a (3 MU/m2/day of Referon by subcutaneous injection) enabled regression of lesions from the sixth month of treatment. After 11 months of treatment, the hemangioma had all but disappeared and interferon therapy was stopped. Repair surgery was planned at 24 months of age. Case 2. A one-month-old infant suffered from a hemangioma of the right side of the face with orbital invasion and risk of amblyopia. Prednisone (2 mg/kg/day) by oral route was ineffective. Interferon alpha 2a enabled regression of the hemangioma and the eye opened from the third month of treatment. Interferon therapy was stopped after 14 months. Initial repair surgery intervention was possible at two years of age. Spastic paraplegia was diagnosed at 18 months of age. The brain and medullar magnetic resonance imaging was normal. No etiology could explain the neurological attack. The possible toxic effect of interferon alpha is discussed. CONCLUSION: Interferon alpha is an effective treatment for hemangiomas. It significantly reduces spontaneous regression time. The uncertainty of long-term effects in infants with hemangiomas incites its indication to be limited to alarming corticosteroid resistant forms and necessitates prolonged neurological surveillance. PMID- 10701063 TI - [Ureaplasma urealyticum respiratory infection in newborn infants]. AB - The neonatal respiratory infection by Ureaplasma urealyticum is rare, but it could represent a major risk for the newborn infants. CASE REPORTS: A term newborn infant presented an early respiratory distress with persistent pulmonary hypertension, requiring artificial ventilation and inhaled nitric oxide therapy. Tracheal aspirates were positive for Ureaplasma urealyticum, although his mother was not contamined. A preterm newborn infant (gestational age: 33 weeks) presented a severe respiratory distress, requiring mechanical ventilation. The tracheal aspirates we positive for Ureaplasma urealyticum, as well as his mother's cervico-vaginal swab. Both recovered thanks to antibiotics (intravenous macrolid during ten days). The outcome was favorable for both babies. CONCLUSION: Neonatal infection due to Ureaplasma is serious. The clinical diagnosis is difficult, recalling group B streptococcal infection. Clinical aggravation, despite antibiotics associated with negative bacteriological standard detection, leads one to evoke this diagnosis and perform specific bacteriological cultures. PMID- 10701064 TI - [Carbohydrate-deficient blood glycoprotein syndrome]. AB - Carbohydrate-deficient glycoprotein syndrome (CDGS) is a newly delineated group of inherited multisystemic disorders associated with abnormal glycosylation of a number of serum glycoproteins. Several types have been described on the basis of clinical presentation and biochemical changes of the glycosylation of serum transferrin and attributed to different enzymatic defects; their clinical presentations are fully different and a clinical heterogeneity is observed within a same type of CDGS. Patients with CDGS type la usually present with neurologic (hypotonia, strabismus and cerebellar hypoplasia) and cutaneous (inverted nipples, abnormal distribution of adipose tissue) abnormalities, together with multivisceral involvement (digestive, hepatic, cardiac, renal). However, neurologic and cutaneous symptoms may be absent, so that CDGS must be looked for in case of unexplained organ failure such as isolated liver insufficiency, cardiomyopathy, pericarditis, tubulopathy, nephrotic syndrome, vascular accident or retinitis pigmentosa. Patients with CDGS type Ib present with liver disease, enteropathy and hypoglycemia without neurologic involvement. These patients are successfully treated with oral mannose administration emphasizing the importance of making the diagnosis. Patients with CDGS type Ic present with mild psychomotor retardation and seizures. Patients with CDGS type II have psychomotor retardation association with severe gastrointestinal disorder, dysmorphic features and abnormal electroretinogram. Other types (III, IV) are less clearly defined and the clinical presentation includes convulsive encephalopathy. Biological abnormalities such as mild hepatic cytolysis, hematologic and hormonal abnormalities are consistently observed in CDGS type I, as well as renal hyperechogeneity, leading one to look for this syndrome when they are associated. Until now, only four enzymatic deficiencies have been identified (types Ia, Ib, Ic, II). PMID- 10701066 TI - [The role of Helicobacter pylori in abdominal pain in children]. AB - Helicobacter pylori (H. pylori) colonizes the human stomach, especially during childhood. H. pylori gastritis, in the absence of duodenal ulcer, does not appear to be associated with specific symptoms. After eradication of H. pylori infection, abdominal pain is improved only in children with duodenal ulcer. Children with H. pylori gastritis cannot be distinguished from uninfected children on the basis of initial symptoms. However, although not demonstrated, a relationship between H. pylori and recurrent abdominal pain might exist since some studies showed that H. pylori-infected children present more frequent pain related to meals or ulcer-like symptoms. These discrepancies could be explained by the fact that H. pylori is probably not a frequent cause of recurrent abdominal pain. The use of refined clinical characteristics of abdominal pain could be of help identifying a subgroup of patients with abdominal pain in whom H. pylori infection needs to be sought and treated. Recent pediatric consensus conferences recommend testing for H. pylori infection by endoscopy only those patients presenting symptoms suggestive of an organic origin. PMID- 10701065 TI - [What is new in pediatric neurology?]. AB - Some significant advances in the field of pediatric neurology are reviewed. For many constitutional disorders, concepts and diagnostic procedures have progressed from various genetic techniques or from protein labeling in situ. Many neurodegenerative disorders, some poorly-defined metabolic diseases, and several syndromes associating mental retardation with neurologic or extraneurologic malformations have been characterized. In addition, for many disorders viewed as 'poorly specific' (mental retardation, epilepsy, migraine), familial forms have permitted us to define the first genes involved. In 'acquired' disorders, new data come from clinical trials (antiepileptic, anti-inflammatory drugs) rather than definite conceptual advances. Finally, clinics and biology are no longer the only approaches to brain functions, and clinical neurophysiology could encounter a second wind thanks to the techniques of functional imaging, especially in the fields of developmental neuropsychology. PMID- 10701067 TI - [Management and prevention of imported Plasmodium falciparum malaria. The 12th Consensus Conference of Anti-infectious Therapy of the French-speaking Society of Infectious Diseases, 14 April 1999]. AB - A consensus conference on management and treatment of imported malaria due to Plasmodium falciparum was organized by the French-speaking society for infectious diseases (Societe de pathologie infectieuse de langue francaise) on April 14, 1999 at Saint-Mande, France. The short text of recommendations of this conference is presented. PMID- 10701068 TI - [Are virtual worlds a threat to the mental health of children and adolescents?]. AB - Video games and the Internet cause enthusiasm but also worry. Among the possible risks, addiction (dependency), isolation, retiring within oneself, and loss of reality, are often put forward. Available data show that serious problems remain exceptional and non-specific, and that these new technological supports do not create new pathologies. Excessive use and isolation have to be solved on an educational basis. Nevertheless, virtual reality, whose applications for the general public are still considered part of the future, needs particular attention. PMID- 10701069 TI - [On what is renal clearance based]. PMID- 10701070 TI - [What evaluation methods for the perinatal care systems?]. PMID- 10701071 TI - [Associated celiac disease and venous thrombosis]. PMID- 10701072 TI - [Acute intestinal intussusception and arterial hypertension]. PMID- 10701073 TI - [Post-vaccination encephalopathy]. PMID- 10701074 TI - [Sleep epilepsy]. PMID- 10701075 TI - Ultrasound interaction with large unilamellar vesicles at the phospholipid phase transition: perturbation by phospholipid side chain substitution with deuterium. AB - The ultrasonic absorption, alpha lambda, as a function of temperature and frequency was determined in large unilamellar vesicles (LUVs) in which specific phospholipid side chains were deuterated. Deuteration significantly altered the temperature and frequency dependence of alpha lambda. The frequency change was especially marked, with decreased frequency and broadening of the ultrasound relaxation, even with only minor changes in the phase transition temperature. Deuteration decreased the Tm and enthalpy of the lipid phase transition, as shown by differential scanning calorimetry, whereas electron spin resonance showed that at and above the lipid phase transition, no differences in the mobility as a function of temperature were observed. These results show that the observed increase in ultrasonic absorption in LUVs at the phospholipid phase transition arises from the interaction of ultrasound with the hydrophobic side chains, probably coupling with structural reorganization of small domains of molecules, a process which is maximized at the phase transition temperature. PMID- 10701076 TI - Interbilayer lipid mixing induced by the human immunodeficiency virus type-1 fusion peptide on large unilamellar vesicles: the nature of the nonlamellar intermediates. AB - A peptide corresponding to the 23 N-terminal amino acid residues of the human immunodeficiency virus type-1 (HIV-1) gp41 has the capacity to induce intervesicular lipid mixing in large unilamellar liposomes composed of dioleoylphosphatidylcholine (DOPC), dioleoylphosphatidylethanolamine (DOPE) and cholesterol (CHOL) (molar ratio, 1:1:1). Cryo-transmission electron microscopy (cryo-TEM) of diluted vesicles to which peptides has been externally added reveals a morphology that is compatible with the formation of nonlamellar lipidic aggregates during the time-course of lipid mixing. 31P-nuclear magnetic resonance and 1-(4-trimethylaminophenyl)-6-phenyl-1,3,5-hexatriene (TMADPH) steady-state anisotropy data at equilibrium indicate that the peptide is able to modulate the lipid polymorphism in pelletted membranes by: (i) promoting the thermotropic formation of inverted phases; and (ii) driving the lamellar-to-nonlamellar transition towards the formation of isotropic phases. Therefore, our combined morphological and spectroscopic data reveal the existence of a direct correlation between the ability of the externally added peptide to induce lipid-mixing in dilute liposome samples and its capacity to modulate lipid polymorphism in stacked bilayers. PMID- 10701077 TI - Synthesis of new fluidity-enhanced amphiphilic compounds for soluble protein two dimensional crystallization purpose. AB - The synthesis of new amphiphilic compounds is described. The structures are rationally designed for soluble protein two-dimensional (2D) crystallization purpose. Special attention is devoted to fluidity properties expected of resulting monolayers. A series of 13 compounds was prepared containing unsaturated, branched or fluorinated alkyl chains. Structures are either symmetrical or dissymmetrical and present a hydroxyl group as polar head, eventually complemented with two other 'secondary' hydrophilic functions. PMID- 10701078 TI - Self-assembly of soluble proteins on functionalized lipid layers: a tentative correlation between the fluidity properties of the lipid film and protein ordering. AB - New series of amphiphilic structures are designed to exhibit various fluidity properties when spread at the air-water interface. The influence of the molecular structure of these lipids on the process of two-dimensional (2D) crystallization of the B subunit of DNA gyrase, a soluble protein, is investigated in terms of size of the crystals produced, protein ordering, and crystallization kinetics. Whereas no difference is observed concerning the mean size of the protein 2D crystals obtained on the different lipid supports, the ultimate protein ordering observable by electron microscopy using the negative-staining technique is more regularly attained with some of these new lipids. The most interesting point results from large discrepancies in crystallization kinetics as highly-ordered protein 2D crystals form within 6-24 h depending on the lipid layer structure. Thus, these new lipids reveal of special interest when studying proteins that suffer from extended incubation time at 4 degrees C or higher temperature and lose their functionality. PMID- 10701079 TI - A differential scanning calorimetry study of the interaction of lasalocid antibiotic with phospholipid bilayers. AB - Interaction of lasalocid sodium salt (Las-Na) with dipalmitoylphosphatidylcholine (DPPC) as a membrane model was investigated by highly-sensitive differential scanning calorimetry (DSC). The insertion properties of the antibiotic were studied both in multilamellar suspensions and unilamellar vesicles, for Las Na/DPPC molar ratios (r) ranging from 0.005 to 0.1. The effect of the antibiotic on the lipid thermotropic behavior is concentration dependent and drastically changes at a critical r of 0.04 in both model membranes. Below this ratio, Las-Na molecules interact with DPPC bilayers without disrupting the global organization of the membrane. In the multilamellar systems only the transition cooperativity is affected whereas for the mixed vesicles, a decrease in the enthalpy change suggests a different mode of insertion. Above this ratio, implantation of the antibiotic give rise to lateral phase separation in multilamellar systems. These structural modifications have repercussions on the formation of mixed LAS-Na/DPPC vesicles which seems limited to an r value of 0.04. PMID- 10701080 TI - Antioxidative and prooxidative effects of coumarin derivatives on free radical initiated and photosensitized peroxidation of human low-density lipoprotein. AB - The antioxidative and/or prooxidative activity of 4-methylcoumanrin (MC), 7 hydroxy-4-methylcoumarin (HMC) and 7,8-dihydroxy-4-methylcoumarin (DHMC), respectively, in the peroxidation of human low-density lipoprotein (LDL) has been studied. The peroxidation was initiated either thermally by water-soluble initiator 2,2'-azobis(2-amidinopropane hydrochloride) (AAPH), or photochemically by a triplet sensitizer benzophenone (BP) or its water-soluble analogue disodium 3,3'-disulfobenzophenonate (DSBP). The reaction kinetics were monitored by the uptake of oxygen and the depletion of alpha-tocopherol (TOH) present in the native LDL. Kinetic analysis of the peroxidation process demonstrated that DHMC is a good antioxidant for both the AAPH-initiated and BP- and DSBP photosensitized peroxidation; HMC is a prooxidant for the AAPH-initiated and DSBP photosensitized peroxidation, but an antioxidant for the BP-sensitized peroxidation; MC is a prooxidant in all of these initiation conditions. The antioxidative action of the coumarin derivatives may include trapping the initiating radicals, trapping the propagating lipid peroxyl radicals, recycling alpha-tocopherol and/or deactivating the excited photosensitizer. PMID- 10701081 TI - Study of the glycosylation of apolipoprotein H. AB - Apolipoprotein H is a single chain polypeptide composed of 326 amino acids highly glycosylated. Its carbohydrate content is approximately 19% of the molecular weight. We show that it is rich in sialic acid linked alpha (2-6) to galactose or N-acetylgalactosamine. Sialic acid is not alpha (2-3) linked to galactose. Galactose is beta (1-4) linked to N-acetylglucosamine and beta (1-3) linked to N acetylgalactosamine. Carbohydrate O-linked chains (mainly sialic acid) are alpha (2-6) linked to galactose or N-acetylgalactosamine. Galactose is also organised in O-linked chains and beta (1-4) linked to N-acetylglucosamine and beta (1-3) linked to acetylgalactosamine. Concanavalin A lectin was used to isolate two groups of apolipoprotein H molecules bearing biantennary and truncated hybrids and high mannose and hybrid oligosaccharides. Apolipoprotein H fails to bind lysine-Sepharose. Our results thus show that it presents truncated hybrid or hybrid-type carbohydrate chains which bear few unmasked mannose residues as a terminal sugar. Biochemical analysis of carbohydrate structures conducted on single isoforms separated through IEF revealed that no specific carbohydrate complex is bound to a single isoform. PMID- 10701082 TI - Traditional therapeutic agents. PMID- 10701084 TI - Antimalarials: unapproved uses or indications. PMID- 10701083 TI - Antibiotics: unapproved uses or indications. PMID- 10701085 TI - Dapsone: unapproved uses or indications. PMID- 10701086 TI - Hormones: androgens, antiandrogens, anabolic steroids, estrogens--unapproved uses or indications. PMID- 10701087 TI - Retinoids: unapproved uses or indications. PMID- 10701088 TI - Phototherapy, photochemotherapy, and photodynamic therapy: unapproved uses or indications. PMID- 10701089 TI - Vitamins A, B, C, D, E, F, trace elements and heavy metals: unapproved uses or indications. PMID- 10701090 TI - Immunosuppressant and cytotoxic drugs: unapproved uses or indications. PMID- 10701091 TI - Unapproved dermatologic indications for H2 receptor antagonists, cromolyn sodium, and ketotifen. PMID- 10701092 TI - Antihelmintic and antiparasitic drugs: unapproved uses or indications. PMID- 10701093 TI - Arachidonate transforming and immunomodulating agents: unapproved uses or indications. PMID- 10701094 TI - Unapproved treatments or indications in dermatology: physical therapy including balneotherapy. PMID- 10701095 TI - Miscellaneous treatments: thalidomide, potassium iodide, levamisole, clofazimine, colchicine, and D-penicillamine. PMID- 10701096 TI - Effects of the butylated hydroxyanisole (BHA) and butylated hydroxytoluene (BHT) on the arylamines N-acetyltransferase activity in rat white blood cells. AB - Butylated hydroxyanisole (BHA) and butylated hydroxytoluene (BHT) were used to determine any effects on the N-acetyltransferase (NAT) activity in rat whole blood and white blood cells as measured by high performance liquid chromatography assay for the amounts of N-acetyl-2-aminofluorene (AAF) and 2-aminofluorene (AF). Two assay systems were performed, one with cellular cytosols, the other with intact white blood cells. The NAT activity in the whole blood and white blood cell cytosols was suppressed by BHA and BHT in a dose-dependent manner, i.e. the higher the concentrations of BHA and BHT, the higher the inhibition of NAT activity. Time-course experiments showed that NAT activity measured from the intact white blood cells was inhibited by BHA and BHT up to 24 h. The results suggest that BHA and BHT suppressed AF acetylation in rat blood with intact white blood cells. PMID- 10701097 TI - Effect of cisplatin on Allium cepa root meristem cells. AB - Allium cepa root growth was retarded by cisplatin treatment in a dose-dependent manner. A decrease in the mitotic index (MI) and an increase in the number of interphase cells was seen in cisplatin treated root tips. An increase in the frequency of abnormal mitoses and chromosomal aberrations was also observed in cisplatin treated groups which indicates its genotoxic effect on plant cells. The endogenous glutathione (GSH) level in the root tips decreased significantly after cisplatin treatment which may favour its increased interaction with cellular DNA thereby developing enhanced chromosomal aberrations and affecting cell divisions and root growth. It is suggested that the decrease in endogenous GSH may be related to the development of cisplatin-mediated genotoxic effects in plants. PMID- 10701098 TI - The acquisition of a second language. AB - It is claimed that if children can begin to acquire a second language at an early age they will find it easier to develop fluency, and will speak it without an accent. Age is a factor in acquiring one's mother tongue, and this also applies when learning a second language. One essential to developing such a skill is the ability to switch from one language to the other, as appropriate. Studies on the effects of age on this learning are reviewed. Techniques such as positron emission tomography can now be used to show which areas of the brain are involved in developing new skills, and much has been learnt in this way. Differences can be demonstrated between the cerebral function of the children who learn a second language at an early age and those who do this when they are older, and also between those who acquire a high degree of fluency and those who never do. If children speak a second language by hearing it in the environment in which they live, they are acquiring it as they do their mother tongue, but if they start at the age of 12 years they are learning it like any other subject they study. If the opportunity is present, surely it is better to acquire a second language than learn it. PMID- 10701099 TI - The spinocerebellar ataxias: molecular progress and newly recognized paediatric phenotypes. PMID- 10701100 TI - Clinical characteristics of children with cerebral white matter abnormalities. AB - The rapidly expanding use of magnetic resonance imaging (MRI) in children with neurological impairments of unknown aetiology has revealed a large number of children with abnormalities of the cerebral white matter, some with leukodystrophy-like white matter abnormalities on MRI, but non-progressive in clinical presentation and course. The aim of this study was to investigate the clinical and neuroradiological characteristics of 26 children with white matter abnormalities of unknown origin and to find diagnostic clues or indicators of progressive versus nonprogressive disease. The typical child with white matter abnormalities was characterized by onset of symptoms within the first year of life, most often presenting as general developmental delay and hypotonia. Later appearing signs were spasticity and ataxia and as a rule severe learning and motor disabilities. Serious ophthalmological signs were frequently seen. Perinatal adverse events were rare, infectious aetiologies not indicated but prenatal stigmata relatively common. The clinical course was progressive in 11 children and non-progressive in 15. Late onset presentation was associated with a progressive course whereas prenatal stigmata and asymmetrical white matter lesions only were found in children with a non-progressive disorder. The MRI showed three main patterns: a) a generalized increase of the T2 signal of the white matter in 12 children, b) a bilateral, symmetric but not generalized abnormality in nine and c) asymmetric, focal or multifocal pathology in five. Useful information as to clinical entities and course was obtained from the combined clinical and radiological assessment. A precise nosological diagnosis could be made in six cases. The study showed that white matter abnormalities in children constitute a heterogeneous group of rare and 'anonymous' conditions, motivating collaborative studies for further clarification of background and management. PMID- 10701101 TI - Novel splice site mutation of aspartoacylase gene in a Turkish patient with Canavan disease. AB - Canavan disease is a severe, progressive autosomal recessive neurodegenerative leukodystrophy. Canavan disease occurs more frequently among Ashkenazi Jewish individuals with two predominant mutations in the aspartoacylase (ASPA) gene. The disease is less frequent in non-Jewish individuals and the mutations randomly reside on the ASPA gene, with one mutation seen more frequently among patients of European extraction. In the present study we report a novel homozygous donor splice site mutation of intron 4 in a child with first-cousin parents of Turkish extraction. PMID- 10701102 TI - Topiramate for drug-resistant epilepsies. AB - Topiramate is a new anti-epileptic drug with proven efficacy against partial seizures in adults. A retrospective assessment of the use of topiramate in drug resistant childhood epilepsy was undertaken. Thirty-four children (median age of 10 years; range 2-18 years) were treated for a median of 9 months (range 6-18 months). The starting dose was 0.25-2.0 mg/kg/day increasing to a maximum of 13 mg/kg/day. Generalized seizures occurred in 27 patients, partial seizures in 15 and infantile spasms in two. Epilepsies were localization-related in 15 patients and generalized in 18. One patient had severe myoclonic epilepsy in infancy. Two patients had Lennox-Gastaut syndrome, five (two currently and three previously) had West syndrome and one had epilepsy with myoclonic absences. Twenty patients had a substantial (> 50%) reduction in seizure frequency; two of whom became seizure-free. Two-patients had an increase in seizures. Efficacy was seen against simple and complex partial seizures, generalized tonic-clonic seizures (primarily generalized), atonic and tonic seizures, myoclonic seizures and infantile spasms. There was no response in the one patient with myoclonic absence seizures. Adverse effects were reported in nine patients; appetite suppression occurred in five patients, behaviour disturbances in three, somnolence in two and poor concentration in one patient. Topiramate is efficacious in a wide spectrum of childhood epilepsies and is well tolerated. PMID- 10701103 TI - Deletions in the spinal muscular atrophy gene region in a newborn with neuropathy and extreme generalized muscular weakness. AB - A newborn presented with respiratory insufficiency requiring artificial ventilation, inability to swallow, lack of spontaneous movements including the facial muscles, and areflexia. Nerve conduction velocities were not recordable. Molecular analysis showed a homozygous deletion in the spinal muscular atrophy (SMN) gene region on chromosome 5q. Pathological and neuropathological examination revealed a normal number of anterior horn cells, hypomyelinated axons in peripheral nerves and some atrophy of skeletal muscle fibres in combination with sarcoplasmic glycogen accumulation. This observation illustrates that severe congenital neuropathy can result from deletions in the SMN gene. PMID- 10701104 TI - Interstitial 6q deletion with a Prader-Willi-like phenotype: a new case and review of the literature. AB - We report on an additional fourth case of Prader-Willi (PW)-like phenotype and an interstitial deletion of 6q. Despite sharing clinical characteristics, patients with a PW-like phenotype and a deletion of 6q, have features which distinguish them from Prader-Willi syndrome (PWS) patients. This case emphasizes the need to examine patients with suspected PWS, but who are negative for recognizable deletions of 15q11-q13 or uniparental maternal disomy of chromosome 15, for a deletion of 6q. PMID- 10701105 TI - Adult and paediatric movement disorders. PMID- 10701106 TI - Development of SCAR markers to the PVY resistance gene Ryadg based on a common feature of plant disease resistance genes. AB - Sequence-characterized amplified regions (SCARs) were developed, based on nucleotide differences within resistance gene-like fragments isolated from a potato plant carrying the Ryadg gene, which confers extreme resistance to potato Y potyvirus (PVY). It originates from Solanum tuberosum subsp. andigena, and a susceptible potato plant. SCARs were tested using 103 potato breeding lines and cultivars with diverse genetic backgrounds derived from Europe, North America, and Japan. Two markers showed high accuracy for detection of the Ryadg gene. The SCAR marker RYSC3 was generated only in genotypes carrying Ryadg. The SCAR marker RYSC4 was detected in all genotypes carrying Ryadg but also in four PVY susceptible genotypes. Neither marker was detected in genotypes carrying other Ry genes originating from different species than S. tuberosum subsp. andigena. Therefore, these SCAR markers should be powerful tools in marker-assisted selection for Ryadg in potato breeding programs, and should also be useful for cloning of the Ryadg gene. PMID- 10701107 TI - Tirant is a new member of the gypsy family of retrotransposons in Drosophila melanogaster. AB - In this paper, we propose a consensus sequence for a putative complete Tirant retrotransposon. Several defective copies, as well as relevant sequences available in databases have been analyzed. The putative complete Tirant element is 8533 bp long, and presents all the structural features of a retrovirus-like transposable element of the gypsy family. It contains three ORFs (open reading frames) that encode putative products resembling the retroviral Gag, Pol, and Env proteins. Southern blot analyses show that complete and defective Tirant elements are widespread in Drosophila melanogaster. The different hybridization patterns observed in several natural populations of this species suggest that Tirant is an active element. PMID- 10701108 TI - Use of microsatellite DNA markers to investigate the level of genetic diversity and population genetic structure of coconut (Cocos nucifera L.). AB - We have used eight pairs of simple sequence repeat (SSR) primers to analyse the genetic diversity in 130 individuals of coconut (Cocos nucifera L.) comprising 75 tall individuals and 55 dwarf individuals, representing 94 different coconut ecotypes throughout the world. A total of 51 alleles were detected, with an average of 6.4 alleles per locus. Fifty alleles were detected in tall coconuts (talls; mean alleles/locus 6.3) compared with only 26 (mean/locus 3.3) in dwarfs, and the average diversity value in talls (0.589) was also significantly higher than that in dwarfs (0.348). Using the eight SSRs we were able to uniquely discriminate 116 of the 130 individuals. A phenetic tree based on DAD (absolute distance) values clustered individuals into five groups, each mainly composed of either talls or dwarfs. These results provide evidence in support of previous hypotheses concerning the dissemination of coconut, as well as important new information for conservation and breeding purposes. PMID- 10701109 TI - Variability in rDNA loci in Iberian species of the genus Zabrus (Coleoptera: Carabidae) detected by fluorescence in situ hybridization. AB - Fluorescence in situ hybridization (FISH) with a PCR-amplified 18S ribosomal probe was used to map rDNA loci in 19 taxa of the ground beetle genus Zabrus (2n = 47-63) from the Iberian Peninsula. A quantitative and qualitative variation has been observed among related species, subspecies, populations, and even individuals. The number of rDNA-carrying chromosomes varies from 2 to 12, and the extent of the signal from small dots to entire arms. Changes altering the number of rDNA clusters seem to be uncoupled from the variation found in the chromosome number. Mechanisms that explain the numerical variation and spreading of rDNA clusters throughout the genome within the genus Zabrus are briefly discussed. No concordance between the pattern of rDNA sites and the phylogenetic relationships as based on morphological characters has been found. PMID- 10701110 TI - Efficiency of RFLP, RAPD, and AFLP markers for the construction of an intraspecific map of the tomato genome. AB - We have constructed a tomato genetic linkage map based on an intraspecific cross between two inbred lines of Lycopersicon esculentum and L. esculentum var. cerasiforme. The segregating population was composed of 153 recombinant inbred lines. This map is comprised of one morphological, 132 RFLP (restriction fragment length polymorphism, including 16 known-function genes), 33 RAPD (random amplified polymorphic DNA), and 211 AFLP (amplified fragment length polymorphism) loci. We compared the 3 types of markers for their polymorphism, segregation, and distribution over the genome. RFLP, RAPD, and AFLP methods revealed 8.7%, 15.8%, and 14.5% informative bands, respectively. This corresponded to polymorphism in 30% of RFLP probes, 32% of RAPD primers, and 100% of AFLP primer combinations. Less deviation from the 1:1 expected ratio was obtained with RFLP than with AFLP loci (8% and 18%, respectively). RAPD and AFLP markers were not randomly distributed over the genome. Most of them (60% and 80%, respectively) were grouped in clusters located around putative centromeric regions. This intraspecific map spans 965 cM with an average distance of 8.3 cM between markers (of the framework map). It was compared to other published interspecific maps of tomato. Despite the intraspecific origin of this map, it did not show any increase in length when compared to the high-density interspecific map of tomato. PMID- 10701111 TI - Microsatellite repeats are not randomly distributed within Norway spruce (Picea abies K.) expressed sequences. AB - A Norway spruce (Picea abies K.) cDNA library obtained from vegetative bud tissue was screened for the presence of (AG)n and (AC)n microsatellite repeats. Ten (AG)n and six (AC)n microsatellites were found, with an average length of 25.5 repeat units. Most of the microsatellites are simple perfect repeats. The microsatellite distribution within the clones is clearly non-random, with different classes of repeats lying in different positions relative to the coding region and in a highly conserved orientation. An estimate of the frequency of dinucleotide microsatellites in expressed regions was obtained, showing that SSRs (simple sequence repeats) are found in genes about 20 times less frequently than in random genomic clones, with (AG)n repeats more frequent than (AC)n repeats. Potential applications of these sequences as expressed region-based molecular markers are shown by developing six SSR markers for the detection of natural variation in Norway spruce populations and testing two of them for the identification of illegitimate progenies from a mapping population. PMID- 10701112 TI - Properties of sequence-tagged-site primer sets influencing repeatability. AB - The polymerase chain reaction (PCR) has become a standard procedure in plant genetics, and is the basis for many emerging genomics approaches to mapping and gene identification. One advantage of PCR is that sequence information for primer sets can be exchanged between laboratories, obviating the need for exchange and maintenance of biological materials. Repeatability of primer sets, whereby the same products are amplified in different laboratories using the same primer set, is important to successful exchange and utilization. We have developed several hundred sequence-tagged site (STS) primer sets for wheat and barley. The ability of the primer sets to generate reproducible amplifications in other laboratories has been variable. We wished to empirically determine the properties of the primer sets that most influenced repeatability. A total of 96 primer sets were tested with four genomic DNA samples on each of four thermocyclers. All major bands were repeatable across all four thermocylers for approximately 50% of the primer sets. Characteristics most often associated with differences in repeatability included primer GC content and 3'-end stability of the primers. The propensity for primer-dimer formation was not a factor in repeatability. Our results provide empirical direction for the development of repeatable primer sets. PMID- 10701113 TI - Quantitative trait loci for root-penetration ability and root thickness in rice: comparison of genetic backgrounds. AB - Drought is the major abiotic stress limiting rice (Oryza sativa) production and yield stability in rainfed lowland and upland ecosystems. Root systems play an important role in drought resistance. Incorporation of root selection criteria in drought resistance improvement is difficult due to lack of reliable and efficient screening techniques. Using a wax-petrolatum layer system simulated to compacted soil layers, root traits were evaluated in a doubled haploid (DH) population derived from the cross between 'IR64' and 'Azucena'. Twelve putative QTLs (quantitative trait loci) were detected by interval mapping comprising four QTLs for root-penetration ability, four QTLs for root thickness, two QTLs for penetrated root number, and two QTLs for total root number. These QTLs individually explained 8.4% to 16.4% of the phenotypic variation. No QTL was detected for maximum penetrated root length by interval mapping. One QTL located between RG104 and RG348 was found to influence both root-penetration ability and root thickness. QTLs for root-penetration ability and root thickness were compared across two populations, 'IR64'-'Azucena' and 'CO39'-'Moroberekan', and different testing conditions. The identified consistent QTLs could be used for marker-assisted selection for deep and thick roots with high root-penetration ability in rice. PMID- 10701114 TI - Developing expressed sequence tags (ESTs) from polymorphic transcript-derived fragments (TDFs) in cassava (Manihot esculenta Crantz). AB - We applied the cDNA-AFLP (amplified fragment length polymorphism) technique to mRNA from the parents of a cassava (Manihot esculenta) genetic mapping population, and obtained more than 500 transcript-derived fragments (TDFs) that were unique in either parent. A subset of 50 TDFs were cloned and sequenced. Sequence alignment of the expressed sequence tags (ESTs) revealed mostly genes of unknown function. Six of the TDFs were mapped on to the cassava genetic map. We also demonstrated by genetic mapping of the TDFs, as RFLP (restriction fragment length polymorphism) markers, that TDFs are more polymorphic than random cDNAs. Generation of ESTs as differentially expressed sequences, in time or between different varieties, is proposed as a way of developing ESTs around specific traits for the candidate locus approach to mapping complex traits. PMID- 10701115 TI - Chloroplast microsatellite analysis reveals the presence of population subdivision in Norway spruce (Picea abies K.). AB - Three chloroplast microsatellites (cpSSRs), previously sequence characterized and for which paternal inheritance was tested and confirmed, were used to assess their usefulness as informative markers for phylogeographic studies in Norway spruce (Picea abies K.) and to detect spatial genetic differentiation related to the possible recolonization processes in the postglacial period. Ninety-seven populations were included in the survey. Some 8, 7, and 6 different size variants for the three cpSSRs, respectively, were scored by analysing 1105 individuals. The above 21 variants combined into 41 different haplotypes. The distribution of some haplotypes showed a clear geographic structure and seems to be related to the existence of different refugia during the last glacial period. The analysis of chloroplast SSR variation detected the presence of two main gene pools (Sarmathic-Baltic and Alpine--Centre European) and a relatively low degree of differentiation (RST of about 10%), characteristic of tree species with large distribution and probably influenced by an intensive human impact on this species. Based on our data, we were not able to detect any evidence concerning the existence of additional gene pools (e.g., from Balkan and Carpathian glacial refugia), though we cannot exclude the existence of genetic discontinuity within the species' European range. A large proportion of population-specific haplotypes were scored in this species, thus indicating a possible usefulness of these markers for the identification of provenances, seed-lots, and autochthonous stands. PMID- 10701116 TI - The 5S rRNA gene diversity in Kengyilia rigidula (Keng and S.L. Chen) J.L. Yang, Yen, and Baum (Poaceae: Triticeae): possible contribution of the H genome to the origin of Kengyilia. AB - Fifty-three units of 5S rDNA sequences from five accessions of Kengyilia rigidula, a member of the tribe Triticeae that also includes wheat, barley, rye, and their wild relatives, have been amplified by the polymerase chain reaction (PCR), cloned, and sequenced. The genome of K. rigidula consists of three haplomes, St, P, and Y. An evaluation of the aligned sequences of the diverse 53 different 5S DNA units yielded three 5S-unit classes. One unit class, Long S1, was assignable to the St haplome, one unit class, the Long P1, was assignable to the P haplome, and a third unit class, Long H1, was assignable to the H haplome. The last was expected to be assignable to the Y haplome, based on previous knowledge. Evolutionary scenarios are put forward to explain this finding. Among those possibilities is that the number of copies of units assignable to the Y haplome is very small and difficult to detect. Short units, reported earlier in K. alatavica, were not found in K. rigidula. PMID- 10701117 TI - Expression and genome organization of resistance gene analogs in soybean. AB - Sequence analysis of cloned plant disease-resistance genes reveals a number of conserved domains. Researchers have used these domains to amplify analogous sequences, resistance gene analogs (RGAs), from soybean and other crops. Many of these RGAs map in close proximity to known resistance genes. While this technique is useful in identifying potential disease resistance loci, identifying the functional resistance gene from a cluster of homologs requires sequence information from outside of these conserved domains. To study RGA expression and to determine the extent of their similarity to other plant resistance genes, two soybean cDNA libraries (root and epicotyl) were screened by hybridization with RGA class-specific probes. cDNAs hybridizing to RGA probes were detected in each library. Two types of cDNAs were identified. One type was full-length and contained several disease-resistance gene (R-gene) signatures. The other type contained several deletions within these signatures. Sequence analyses of the cDNA clones placed them in the Toll-Interleukin-1 receptor, nucleotide binding domain, and leucine-rich repeat family of disease-resistance genes. Using clone specific primers from within the 3' end of the LRRs, we were able to map two cDNA clones (LM6 and MG13) to a BAC contig that is known to span a cluster of disease resistance genes. PMID- 10701118 TI - Combined AFLP and RFLP mapping in two hexaploid oat recombinant inbred populations. AB - A combined RFLP and AFLP map was constructed for hexaploid oat (Avena spp.). The segregation of AFLP markers was scored in two hexaploid oat recombinant inbred line (RIL) populations, the 'Kanota' x 'Ogle' RFLP population, and a population derived from 'Clintland64' and 'IL86-5698', barley yellow dwarf virus (BYDV) sensitive and BYDV-tolerant lines, respectively. More than 300 AFLP markers were scored in each population, of which 97 could be scored in both populations. AFLP markers were linked to RFLP markers in 32 of 36 'Kanota' x 'Ogle' RFLP linkage groups. The addition of the AFLP markers to the 'Kanota' x 'Ogle' RFLP data set combined markers from four pairs of linkage groups and increased the size of the map from 1402 cM to 2351 cM. Thirty linkage groups were observed in the 'Clintland64' x 'IL86-5698' population, two of which could be consolidated by comparing the maps from both populations. The AFLP and RFLP markers showed very similar distributions in the 'Kanota' x 'Ogle' population with a tendency of each type of marker to cluster with markers of the same type. The placement of a set of AFLP markers on the 'Kanota' x 'Ogle' linkage map will enrich the RFLP map and allow others to relate AFLP markers for agronomically important genes to the reference 'Kanota' x 'Ogle' linkage map. PMID- 10701119 TI - A binary vector-based large insert library for Brassica napus and identification of clones linked to a fertility restorer locus for Ogura cytoplasmic male sterility (CMS). AB - We constructed and characterized a large DNA insert library for Brassica napus that would facilitate genome-related research and map-based cloning efforts in Brassica species. This library, consisting of 92,160 clones arrayed in 384-well microtiter dishes, was based on a conventional plant transformation vector (binary vector), and was constructed using a single ligation with transformation efficiency of over 5000 recombinants per microliter of ligation mixture. Every clone in this library contains an insert in the size range of 30-190 kb, facilitating both chromosome walking and plant transformation. Screening this library with three DNA markers (C2, F10, and CabR) that are linked to a fertility restorer locus for Ogura cytoplasmic male sterility (CMS) identified at least 17 positive clones for each probe. Among the 17 positive clones identified by C2, nine are linked to the restorer locus. Marker F10 identified 21 clones, of which only two are linked to the restorer locus. None of 68 clones identified by CabR is linked to the restorer locus. A stability test using two clones identified by the C2 marker indicated that large DNA inserts are stable in this conventional vector in both Escherichia coli and Agrobacterium. PMID- 10701120 TI - Molecular phylogeny of mangroves. VI. Intraspecific genetic variation in mangrove species Excoecaria agallocha L. (Euphorbiaceae). AB - Genomic DNA from 84 individuals of Excoecaria agallocha from seven mangrove populations were analysed for random amplified polymorphic DNAs (RAPDs) using 16 random 10-mer primers. Polymorphism within populations varied from 20% to 31%. At the interpopulation level, 111/149 (74%) of RAPDs were polymorphic. Restriction fragment length polymorphism (RFLP) analysis of 21 individuals (3 individuals randomly selected from the 7 populations) using 30 probe-enzyme combinations revealed a high level of interpopulation polymorphism (62.2%) indicating interpopulation genetic divergence. The polymorphic RAPDs and RFLPs were pooled, and clustering was carried out based on mean similarity for individual populations. The dendrogram showed groupings of populations from the West and East Coasts of India into separate clusters, at 60% similarity level. Further, RAPD and RFLP analysis of male and female plants showed approximately the same level of variation in both sexes, and no sex-linked markers were found. These results demonstrate that considerable intrapopulation and interpopulation genetic variations exist in E. agallocha, and that lack of genetic variation is not the reason for the morphological uniformity observed across the range of the species. PMID- 10701122 TI - A CAPS marker to assist selection of tomato spotted wilt virus (TSWV) resistance in pepper. AB - The hypersensitive resistance to tomato spotted wilt virus (TSWV) in pepper is determined by a single dominant gene (resistant allele: Tsw) in several Capsicum chinense genotypes. In order to facilitate the selection for this resistance, four RAPD (among 250 10-mer primers tested) were found linked to the Tsw locus using the bulked segregant analysis and 153 F2 individuals. A close RAPD marker was converted into a codominant cleaved amplified polymorphic sequence (CAPS) using specific PCR primers and restriction enzymes. This CAPS marker is tightly linked to Tsw (0.9 +/- 0.6 cM) and is helpful for marker-assisted selection in a wide range of genetic intercrosses. PMID- 10701123 TI - Chromosomal evidence for sibling species of the malaria vector Anopheles cruzii. AB - An analysis of the ovarian polytene chromosomes of Anopheles cruzii from three localities in Southeast Brazil revealed the existence of two genetic entities within this morphologically uniform taxon. These cryptic species differed in the banding patterns of the X chromosome and 3L arm. A pattern of bands that cannot be explained by the fixation of any of the known inversions in chromosome X was revealed and named chromosomal form B to distinguish it from the standard pattern of this X chromosome, form A. Each chromosomal form is characterized by a different set of inversions. The lack of heterozygotes (A/B) for these X chromosome forms in populations where both forms coexist is evidence of absence or limited gene flow between the two groups. PMID- 10701121 TI - Gene content and organization of a 281-kbp contig from the genome of the extremely thermophilic archaeon, Sulfolobus solfataricus P2. AB - The sequence of a 281-kbp contig from the crenarchaeote Sulfolobus solfataricus P2 was determined and analysed. Notable features in this region include 29 ribosomal protein genes, 12 tRNA genes (four of which contain archaeal-type introns), operons encoding enzymes of histidine biosynthesis, pyrimidine biosynthesis, and arginine biosynthesis, an ATPase operon, numerous genes for enzymes of lipopolysaccharide biosynthesis, and six insertion sequences. The content and organization of this contig are compared with sequences from crenarchaeotes, euryarchaeotes, bacteria, and eukaryotes. PMID- 10701124 TI - Characterization of university-level introductory genetics courses in Canada. AB - We conducted survey research with the intent to characterize post-secondary introductory genetics (IG) education in Canada during the 1996-1997 academic year. At least a minimum data set was obtained from 47 institutions through responses to a mailed questionnaire and on-line resources. The total reported enrollment (TRE) for IG was 10,500. Over half of the TRE used one particular text. A core curriculum of topics was identified as those given more than 30 min of lecture time in at least half of reporting institutions. Slightly more than half of the TRE had laboratory exercises associated with their IG course. Laboratory exercises tended to emphasize classical transmission genetics with very few exercises in molecular genetics. For the determination of academic equivalency between institutions, particular attention should be given to the breadth and duration of the tutorial and (or) laboratory components. The majority of personnel teaching IG were trained in Canada within the previous 15 years. We suggest mechanisms by which the Genetics Society of Canada could work to promote genetical literacy. PMID- 10701125 TI - Microsatellite loci in the phytoparasitic nematode Globodera. AB - A Globodera pallida genomic library, population Guiclan (Pa2/3), was screened for TG and TC microsatellite motifs. Screening of 50,000 clones revealed 48 positive matches. After sequencing, primers were designed to amplify 14 microsatellite loci. The specificity of the loci was tested with DNA templates of other populations of G. pallida, and also on other species of Globodera. Appearance of amplification products on several of these DNA templates showed that the microsatellite flanking regions are relatively conserved between G. pallida populations as well as between Globodera species. Evidence for allele polymorphism between individuals was demonstrated by using nine loci primers, in G. pallida population Guiclan and from a population of a closely related species G. "mexicana". Some alleles appeared to be species specific. PMID- 10701126 TI - Meiotic mutants of Medicago sativa show altered levels of alpha- and beta tubulin. AB - We have analysed the level of accumulation of alpha- and beta-tubulin polypeptides in flowers collected from different meiotic mutants of alfalfa (Medicago sativa L.). The H33 mutant previously identified as a producer of male and female gametes with the somatic chromosome number (2n gametes) as a result of defective spindle orientation or, more rarely, abnormal cytokinesis, showed a higher level of alpha- and beta-tubulin compared to control diploid plants and approximately the same level as control tetraploid plants. A higher level of tubulin was likewise observed in diploid plants displaying abnormalities in spindle orientation and cytokinesis, which had gone through 3-4 cycles of phenotypic recurrent selection to increase 2n gamete production. A similar analysis was performed on another class of Medicago meiotic mutants characterized by production of 4n pollen (jumbo pollen, due to the absence of cytokinesis at the end of meiosis) and 2n eggs. Again, the level of alpha- and beta-tubulin was found to be higher in the mutants than in diploid controls. We conclude that meiotic defects, such as abnormal spindle orientation or cytokinesis leading to the formation of 2n gametes, determine an increased level of tubulin, the main constituent of plant microtubules (MTs). PMID- 10701127 TI - Sequential meiotic prophase development in the pubertal Indian pygmy field mouse: synaptic progression of the XY chromosomes, autosomal heterochromatin, and pericentric inversions. AB - Sequential meiotic prophase development has been followed in the pubertal male pygmy mouse Mus terricolor, with the objective to identify early meiotic prophase stages. The pygmy mouse differs from the common mouse by having large heterochromatic blocks in the X and Y chromosomes. These mice also show various chromosomal mutations; for example, fixed variations of autosomal short arms heterochromatin among different chromosomal species and pericentric inversion polymorphism. Identification of prophase stages was crucial to analyzing effects of heterozygosity for these chromosomal changes on the process of homologous synapsis. Here we describe identification of the prophase stages in M. terricolor, especially the pachytene substages, on the basis of morphology of the XY bivalent. Based on this substaging, we show delayed pairing of the heterochromatic short arms, which may be the reason for their lack of chiasmata. The identification of precise pachytene substages also reveals an early occurrence of "synaptic adjustment" in the pericentric inversion heterobivalents, a mechanism that would prevent chiasma formation in the inverted segment and thereby would abate adverse effects of such heterozygosity. The identification of pachytene substages would serve as the basis to analyze the nature of synaptic anomalies met in M. terricolor hybrids (which will be the basis of a subsequent paper). PMID- 10701128 TI - Identification and chromosomal location of a new tandemly repeated DNA in maize. AB - Two clones of a new family of tandemly repeated DNA sequences have been isolated from a maize random genomic DNA library. MR68 is 410 bp, representing a monomeric unit and MR77 is 1222 bp, containing three units. The copy number was estimated to be about 3000 per 1C maize genome. Its methylation pattern was also determined. Fluorescent in situ hybridization (FISH) indicates that the sequence is located on the subtelomeric region of the long arm of chromosomes 3 and 6, as well as on the satellite of chromosome 6. PMID- 10701129 TI - Isolation, characterization, and chromosomal location of the tRNA(Met) genes in Atlantic salmon (Salmo salar) and brown trout (Salmo trutta). AB - This work describes the isolation, characterization, and physical location of the methionine tRNA in the genome of Atlantic salmon (Salmo salar L.) and brown trout (Salmo trutta L.). An Atlantic salmon genomic library was screened using a tRNA(Met) probe from Xenopus laevis. Two cosmid clones containing the Atlantic salmon tRNA(Met) gene were isolated, subcloned and sequenced. The tRNA(Met) was mapped to metaphase chromosomes by fluorescence in situ hybridization (FISH). Chromosomal data indicated that the tDNA of methionine is tandemly repeated in a single locus in both species. Analysis of genomic DNA by Southern hybridization confirmed the tandem organization of this gene. PMID- 10701130 TI - Physical characterization of the homoeologous group 5 chromosomes of wheat in terms of rice linkage blocks, and physical mapping of some important genes. AB - The wheat homoeologous Group 5 chromosomes were characterized physically in terms of rice linkage blocks using a deletion mapping approach. All three chromosomes, 5A, 5B, and 5D, were shown to have a similar structure, apart from the 4A-5A translocation on the distal end of chromosome arm 5AL. The physical mapping of rice markers on the deletion lines revealed that the whole of rice chromosome 9 is syntenous to a large block, proximal to the centromere, on the long arm. Likewise, a small segment of the distal end of the long arm showed conserved synteny with the distal one-third end of the long arm of rice chromosome 3. In between those conserved regions, there is a region on the long arm of the Group 5 chromosomes which shows broken synteny. The proximal part of the short arms of the Group 5 chromosomes showed conserved synteny with a segment of the short arm of rice chromosome 11 and the distal ends showed conserved synteny with a segment of rice chromosome 12. The physical locations of flowering time genes (Vrn and earliness per se) and the gene for grain hardness (Ha) on the Group 5 chromosomes were determined. These results indicate that comparative mapping using the deletion mapping approach is useful in the study of genome relationships, the physical location of genes, and can determine the appropriate gene cloning strategy. PMID- 10701131 TI - Construction of a bacterial artificial chromosome (BAC) library for potato molecular cytogenetics research. AB - Lack of reliable techniques for chromosome identification is the major obstacle for cytogenetics research in plant species with large numbers of small chromosomes. To promote molecular cytogenetics research of potato (Solanum tuberosum, 2n = 4x = 48) we developed a bacterial artificial chromosome (BAC) library of a diploid potato species S. bulbocastanum. The library consists of 23,808 clones with an average insert size of 155 kb, and represents approximately 3.7 equivalents to the potato genome. The majority of the clones in the BAC library generated distinct signals on specific potato chromosomes using fluorescence in situ hybridization (FISH). The hybridization signals provide excellent cytological markers to tag individual potato chromosomes. We also demonstrated that the BAC clones can be mapped to specific positions on meiotic pachytene chromosomes. The excellent resolution of pachytene FISH can be used to construct a physical map of potato by mapping molecular marker-targeted BAC clones on pachytene chromosomes. PMID- 10701132 TI - Identification of rpaP1-5 and rpaP2-6 genes encoding two additional variants of the 60S acidic ribosomal proteins of Schizosaccharomyces pombe. AB - In the fission yeast, four genes (rpaP1-1, rpaP1-3, rpaP2-2, and rpaP2-4) encoding two variants of the RpaP1 and RpaP2 ribosomal proteins (rp) have been characterized. We have identified cDNA for additional variants called RpaP1.5 and RpaP2.6. Sequence comparison suggests that RpaP1.5 diverged before RpaP1.1 and RpaP1.3 and that RpaP2.6 is closer to RpaP2.2 than to RpaP2.4. The corresponding genes, rpaP1-5 and rpaP2-6, are transcribed coordinately with other rp genes. PMID- 10701133 TI - Microsatellite markers useful throughout the genus Dianthus. AB - Using repeats found in sequences from Dianthus species present in the EMBL database, primers for STMS (sequence-tagged microsatellite site) analysis were developed and tested. Five loci were polymorphic and amplified products of sufficient quality in nearly all of the 26 Dianthus species tested, except MS DINGSTA, which amplified in only one-third of the species. Loci MS-DINMADSBOX and MS-DCDIA30 produced allele series that were mostly two nucleotides (the repeat unit) apart. MS-DCAMCRBSY and MS-DINCARACC also amplified regular series of alleles, but more than two fragments per individual were detected in a number of species. Both loci code for a member of the ACC synthase gene family. The observation that the loci amplified across a wide range of Dianthus species may imply that the different species within the genus are relatively closely related. Alternatively, it may indicate that the regions selected for primer design (some of which are in coding regions) are well conserved. These microsatellites will be useful for the measurement of genetic diversity in natural populations of Dianthus species and the identification of carnation varieties. PMID- 10701134 TI - Larger genomes for molluskan land pioneers. AB - The terrestrial pulmonate mollusks were found to have the significantly larger genomes than the aquatic pulmonates. Being shown in the independent phylogenetic branch, this phenomenon suggests that the previously observed genome enlargement in the vertebrate land pioneers (amphibians and lungfishes) was not casual. As in the vertebrates, the larger molluskan genomes are also more GC-rich. PMID- 10701135 TI - Expressed retrotransposed 5S rRNA genes in the mouse and rat genomes. AB - The analyses of previously described 5S rRNA gene sequences show that some of the expressed 5S rRNA genes present in the mouse and rat genomes were derived from the retrotransposition of 5S rRNA transcripts. These analyses demonstrate that new 5S rRNA gene copies can originate by retrotransposition and that some of these retrotranscribed genes are expressed. PMID- 10701136 TI - Abnormalities of renal function in the elderly. AB - OBJECTIVE: To investigate the glomerular and proximal tubular renal function and the prevalence of urinary abnormalities in the elderly. DESIGN: Cross-sectional study. SETTING: General community in the city of Sao Paulo. PARTICIPANTS: A population-based sample of 200 elderly subjects was randomly selected. Of these, 81 subjects (45 females and 36 males; mean +/- SD age: 73.7 +/- 6 years) accepted to undergo laboratory examination and were included in the study. MAIN OUTCOME MEASURES: 24-h creatinine clearance (CCr), microalbuminuria, urinary retinol binding protein (urRBP), leukocyturia, hematuria and total proteinuria. RESULTS: CCr was lower than 80 ml/min/1.73 m2 in 68% of the subjects. The median (range) CCr was 65 ml/min/1.73 m2 (21-112) in males and 77 ml/min/1.73 m2 (27-107) in females (p = 0.14). No individual had serum creatinine greater than 1.5 mg/dl. urRBP determination was normal in 79 of 81 subjects. The prevalence of microalbuminuria (> 20 micrograms/ml) was 31% (n = 25, 19 men and 6 women). These individuals presented higher mean systolic blood pressure (147 +/- 20 vs. 135 +/- 22 mmHg, p = 0.02) and mean serum creatinine (1.13 +/- 0.20 vs. 0.96 +/- 0.20 mg/dl, p < 0.01) than those without microalbuminuria. The prevalence of leukocyturia (> 10,000/mm3), hematuria (> 10,000/mm3) and total proteinuria (> or = 0.3 mg/dl) was 19%, 28% and 5% in males and 33%, 27% and 4% in females. CONCLUSIONS: Glomerular dysfunction and urinary abnormalities are frequent features in the elderly, however, proximal tubular dysfunction is uncommon in this population. PMID- 10701137 TI - Peritoneal dialysis in older individuals. AB - Peritoneal dialysis is a viable alternative to hemodialysis for management of elderly patients requiring renal replacement therapy. Peritoneal dialysis confers several advantages over hemodialysis for the elderly--namely independence, home treatment and perhaps preservation of residual renal function. Although there are a few contraindications, these are minimal and can largely be overcome with attention to special training and the use of healthcare partners to perform the technique of peritoneal dialysis exchanges. PMID- 10701138 TI - Acute renal failure in the elderly. PMID- 10701139 TI - Diabetic nephropathy in patients with type II diabetes. PMID- 10701140 TI - Aging kidneys in an aging population: how does this impact nephrology and nephrologists? AB - The 'demographic imperative' of a progressively aging society will place unprecedented demands on the health care system in the 21st century. Although improved education, public health measures, personal lifestyles, and health care will result in a large proportion of those born surviving to old age in robust health and vitality, the sheer numbers of 'baby-boomers' who will become the elderly and the inevitable association between aging and the associated multiple, especially chronic diseases and physiological impairments of old age will require more efficient and more effective systems of health care to meet the needs of the aging population. Generalists, specialists, and medical and surgical subspecialists will play important roles in meeting these needs, often in the multidisciplinary mode. Geriatricians will directly provide but a small minority of the care, focusing upon education, research, and consultation and in delivering primary care to the frail elderly and especially in long term care. Collaboration with subspecialists will be frequent in all these domains. Nephrologists, who already practice multidisciplinary team care of frail, complicated, chronically ill patients with end-stage renal disease, have much to contribute as their patient population progressively grows in numbers and age. Hence geriatricians and nephrologists have much to learn from and contribute to each other in addressing the 'age wave' of the 21st century. PMID- 10701141 TI - Managed care, geriatrics, and nephrology. PMID- 10701142 TI - Hypertension: a risk factor for dementia. PMID- 10701143 TI - Well-adjusted children: an alternate view of children with inflammatory bowel disease and functional gastrointestinal complaints. AB - Previous studies have suggested impaired psychosocial adjustment in children and adolescents with inflammatory bowel disease (IBD). We examined 62 subjects referred to a regional Pediatric Gastroenterology Clinic with IBD or functional gastrointestinal (FGI) complaints. Characteristics of the clinic include a unified team approach, regularly scheduled appointments at 3-month intervals, proactive medical care emphasizing maintenance of full functioning, and close medical-surgical interaction (joint clinics). A research assistant administered a questionnaire regarding children's perceptions of their illness, as well as the Child Depression Inventory (CDI), the Piers-Harris (PH) test of self-concept, and the Child Behaviour Checklist (CBCL). The 36 children with IBD (25 Crohn's disease, 11 ulcerative colitis, mean age 13.3 +/- 3.0 years) were compared with 26 patients with FGI complaints (16 recurrent abdominal pain, 10 functional megacolon, mean age 11.4 +/- 2.8 years). The scores on the standardized tests were not clinically significant for either group. In comparison, however, children with IBD were less depressed and had fewer behaviour problems than the FGI group. Surprisingly, only 19% (7 of 36) of children with IBD described their illness as a problem to them, compared with 65% (17 of 26) of children with FGI symptoms. The latter children also considered themselves significantly sicker than did those with IBD. We conclude that normal psychosocial adjustment is possible in pediatric patients with IBD. We speculate that this group benefitted from the professional supports that our clinic specifically provides to patients with IBD. The FGI group may have suffered from a lack of such professional supports, as well as from the absence of a specific diagnosis. PMID- 10701144 TI - A simple classification of Crohn's disease: report of the Working Party for the World Congresses of Gastroenterology, Vienna 1998. AB - Crohn's disease is a heterogeneous entity. Previous attempts of classification have been based primarily on anatomic location and behavior of disease. However, no uniform definition of patient subgroups has yet achieved broad acceptance. The aim of this international Working Party was to develop a simple classification of Crohn's disease based on objective variables. Eight outcome-related variables relevant to Crohn's disease were identified and stepwise evaluated in 413 consecutive cases, a database survey, and by clinical considerations. Allocation of variables was conducted with well-defined Crohn's disease populations from Europe and North America. Cross-table analyses were performed by chi-square testing. Three variables were finally elected: Age at Diagnosis [below 40 years (A1), equal to or above 40 years (A2)], Location [terminal ileum (L1), colon (L2), ileocolon (L3), upper gastrointestinal (L4)], and Behavior [nonstricturing nonpenetrating (B1), stricturing (B2), penetrating (B3)]. The allocation of patients to these 24 subgroups proved feasible and resulted in specific disease clusters. Cross-table analyses revealed associations between Age at Diagnosis and Location, and between Behavior and Location (all p < 0.001). The Vienna classification of Crohn's disease provides distinct definitions to categorize Crohn's patients into 24 subgroups. Operational guidelines should be used for the characterization of patients in clinical trials as well as for correlation of particular phenotypes with putative biologic markers or environmental factors. PMID- 10701146 TI - The key role of macrophages in the immunopathogenesis of inflammatory bowel disease. AB - Macrophages are important in the host's immunological and inflammatory responses. There is a large population of these cells in the normal intestinal mucosa where they represent the major antigen presenting cell population capable of determining the type of T cell responses that develop to luminal antigens. Studies suggest that the normal intestinal macrophages cannot be easily induced to mediate acute inflammatory responses. In active inflammatory bowel disease there is an increase in the mucosal macrophage population, derived from circulating monocytes. These recruited macrophages are phenotypically different from the resident population of cells and play a major role in mediating the chronic mucosal inflammation seen in patients with ulcerative colitis and Crohn's disease. They secrete many cytokines that are important in the proinflammatory responses, such as interleukin (IL)-1, IL-6, IL-8, IL-12, IL-18, and tumor necrosis factor-alpha. They also release reactive metabolites of oxygen and nitrogen and proteases that degrade the extracellular matrix. Macrophages also appear to be important during resolution of inflammation and repair of the intestinal mucosa that occurs during disease remission. PMID- 10701145 TI - Efficacy of mycophenolate mofetil in patients failing 6-mercaptopurine or azathioprine therapy for Crohn's disease. AB - Mycophenolate mofetil (MMF) is a novel immunomodulator that may be effective in the treatment of chronic active and perianal Crohn's disease (CD). The aim of this study is to assess the efficacy of MMF in CD patients who failed or were intolerant of 6-mercaptopurine (6-MP) or azathioprine (AZA). Eleven CD patients were treated with MMF after a failed course of 6-MP/AZA, and their records reviewed retrospectively. Reasons for 6-MP/AZA intolerance or failure were recorded. Response to MMF was determined by calculation of the Harvey-Bradshaw index and ability to taper steroids. Adverse reactions to MMF were recorded. Eleven patients were identified who failed a previous trial of 6-MP/AZA and other immunomodulators and required immunomodulator therapy. Of 11 patients who started MMF, four had early adverse reactions within 8 weeks and stopped the medication. Of the remaining seven patients who took MMF for at least 8 weeks, one had a complete response, two had a partial response, and four had no response to the medication. In patients who failed 6-MP/AZA, MMF was of benefit in 3 of 11 patients with only one complete responder. This lower-than-expected response rate may indicate that patients who are resistant to 6-MP or AZA may also be resistant to MMF. PMID- 10701147 TI - Viruses and inflammatory bowel disease: is there evidence for a causal association? AB - There has been a resurgent interest in potential microbial etiologies of inflammatory bowel disease (IBD). Over the past decade there have been both epidemiological and tissue studies exploring the potential role of paramyxoviruses in IBD, particularly Crohn's disease. This article will review the evidence and hence plausibility of a causal association between these viruses and IBD. PMID- 10701148 TI - Is laparoscopic surgery for most, a few, or no patients with Crohn's disease? PMID- 10701149 TI - Laparoscopic surgery for Crohn's disease?--a conditional yes. PMID- 10701150 TI - How to do without steroids in inflammatory bowel disease. AB - This review covers the use of steroids in the treatment of both ulcerative colitis and Crohn's disease. It looks at controlled trials and uncontrolled trials as to the benefits of this agent in both inducing and maintaining remission. The review also stresses the high incidence of toxicity with prolonged use of steroids and the fact that controlled trials have clearly shown that steroids do not maintain remission in either disorder. Alternatives to initiating steroids in mild to moderately active ulcerative colitis and Crohn's disease are presented. The use of steroids in fistulizing versus nonfistulizing Crohn's is also covered. Finally, there is a review of data and discussion of the role of antibiotics, immunosuppressives, and combination therapy for both ulcerative colitis and Crohn's disease. The expectation is that the reader will consider alternatives to initiating and maintaining steroids for prolonged periods of time in the treatment of inflammatory bowel disease. PMID- 10701151 TI - IBD highlights from the 1999 American College of Gastroenterology meeting: infliximab and 6-MP. PMID- 10701152 TI - PSC in UC: is it or is it not a risk factor for the development of neoplasia? PMID- 10701153 TI - Infliximab for fistulas: a hole in one? PMID- 10701154 TI - Intestinal pathogen or IBD, same T-cell response. PMID- 10701155 TI - Markov Polo: finding the costs of Crohn's in an Olmstead County pool. PMID- 10701156 TI - The role of liquid diet in the management of small bowel Crohn's disease. PMID- 10701157 TI - Reader response: antitumor necrosis factor therapy. PMID- 10701158 TI - Aggrecan from start to finish. PMID- 10701159 TI - Effect of low-calcium hemodialysate on bone metabolism. AB - In the present study, we investigated the kinetics of bone metabolism by determining serum bone metabolic markers and quantifying bone mineral density by dual-energy X-ray absorptiometry to clarify the effect of long-term use of low calcium hemodialysate on bone metabolism. After changing the calcium concentration in the dialysate from 3.0 mEq/l to 2.5 mEq/l, serum intact parathyroid hormone level, serum highly sensitive parathyroid hormone level, and serum bone metabolic markers were determined in ten patients with chronic nondiabetic renal insufficiency during 1 year. The doses of an oral phosphate binder and activated vitamin D were carefully regulated to control serum ionized calcium levels and serum inorganic phosphorus levels. Bone mineral density was determined at the distal 1/3 and 1/6 of the radius on the nonshunt side. As a result, the required amount of oral phosphate binder was increased; however, there was no need to significantly increase the amount of activated vitamin D. Intact parathyroid hormone showed no significant variation, but the highly sensitive parathyroid hormone was significantly increased. There were no significant changes in any bone metabolic markers or in bone mineral density. From these study results, it was found that it was difficult to increase the dose of activated vitamin D even if low-calcium hemodialysate was used, and that during use of the low-calcium hemodialysate the serum level of parathyroid hormone tended to increase but led to neither acceleration of bone turnover nor a decrease in bone mineral density. PMID- 10701160 TI - Expression of bone morphogenetic proteins and rat distal-less homolog genes following rat femoral fracture. AB - Expression of the genes encoding bone morphogenetic proteins (BMPs), BMP type IA receptor (BMPR-1A), and rat distal-less homolog (rDlx) was studied in bone, callus, and the surrounding soft tissue following rat femoral closed fracture, using RT-PCR-based techniques. Before fracture, the genes encoding BMP-5, BMP-6, and BMPR-1A were found to be expressed in both bone and the surrounding soft tissue, whereas the BMP-2 gene was expressed only in bone and BMP-7 was not expressed in either tissue. Expression of these genes was unaffected by fracture. The gene encoding BMP-4 was also expressed in both bone and the surrounding soft tissue before fracture. Moreover, although unchanged in bone, 6 h after fracture BMP-4 expression was increased tenfold in the surrounding soft tissue. The increased BMP-4 expression was transient and returned to prefracture levels within 72 h. Expression of rDlx was also increased in bone after fracture, but at later times than were observed with BMP-4: elevated rDlx expression was detected after 48 h and persisted for 30 days or more. No expression of rDlx was observed in the surrounding soft tissue before or after fracture. These findings indicate that BMP-4 and rDlx are selectively expressed following femoral fracture in the rat, and also suggest that they are involved in the formation of the callus at an early point during the postfracture healing of bone. PMID- 10701161 TI - Prolonged intake of fermented soybean (natto) diets containing vitamin K2 (menaquinone-7) prevents bone loss in ovariectomized rats. AB - The effect of the prolonged intake of dietary vitamin K2 (menaquinone-7, MK-7) on bone loss in ovariectomized (OVX) rats was investigated. OVX rats were freely given experimental diets containing the fermented soybean (natto; including 9.4 micrograms MK-7/100 g diet) without or with supplemental MK-7 (containing 14.1 or 18.8 micrograms of MK-7 as total per 100 g diet) for 150 days. Feeding produced a significant elevation of MK-7 concentration in the serum of OVX rats. In this case, the femoral MK-4 content was significantly increased, but MK-7 was not detected in the femoral tissues, indicating degradation of MK-7. Serum gamma carboxylated osteocalcin concentration was significantly decreased by OVX. This decrease was significantly prevented by the feeding of the natto diets with supplemental MK-7 (18.8 micrograms/100 g diets). OVX caused a significant decrease in femoral dry weight, femoral calcium content, and mineral density. These decreases were significantly prevented by feeding with diets containing natto with MK-7 (total, 18.8 micrograms/100 g diets). This study demonstrates that the prolonged intake of natto dietary including MK-7 has a preventive effect on bone loss induced by OVX. Dietary MK-7 may be useful in the prevention of osteoporosis. PMID- 10701162 TI - Synergistic effect of genistein and zinc on bone components in the femoral metaphyseal tissues of female rats. AB - The effect of genistein and zinc on bone components in rats was investigated. Femoral-metaphyseal tissues obtained from female rats (4 weeks old) were cultured for 24 h in a medium containing either vehicle or genistein (10(-7)-10(-5) M) in the absence or presence of zinc sulfate (10(-6)-10(-4) M) in vitro. The presence of genistein (10(-6) and 10(-5) M) or zinc (10(-6) and 10(-5) M) caused a significant increase in alkaline phosphatase activity, deoxyribonucleic acid (DNA), and calcium content in the metaphyseal tissues. These increases were significantly enhanced by the combination of each compound. The synergistic effect on bone components was seen in the combination with genistein (10(-6) and 10(-5) M) plus zinc (10(-5) M). Such an effect was completely blocked by the presence of cycloheximide (10(-6) M), an inhibitor of protein synthesis. Moreover, the oral administration of genistein (100 and 300 micrograms/kg body weight) or zinc sulfate (1 and 5 mg Zn/kg) to rats for 3 days caused a significant elevation of alkaline phosphatase activity, DNA, and calcium content in the femoral-metaphyseal tissues. These increases were significantly enhanced by the combination of each compound. The synergistic effect was seen in the case of genistein (100 and 300 micrograms/kg) plus zinc (5 mg/kg). These results demonstrate that the anabolic effect of genistein on bone components is synergistically enhanced by zinc in vitro and in vivo. This study further supports the view that the combination of nutritional factors has a potent anabolic effect on bone metabolism. PMID- 10701163 TI - Epidemiology of cervical and trochanteric fractures of the proximal femur in 1994 in Tangshan, China. AB - The purpose of this study was to determine the incidence of cervical and trochanteric fractures of the proximal femur in 1994 in Tangshan City, China. There are many reports on hip fracture incidence in many countries, suggesting that there are many factors affecting hip fractures. We visited 15 hospitals with an orthopaedic department within Tangshan City, and reviewed the medical records and radiographs of all patients with hip fractures occurring between January 1 and December 31, 1994. The population of Tangshan in 1994 was determined to be 1,454,543 (746,015 males and 708,528 females). The population of those over 65 years of age was 88,490 (41,519 males and 46,971 females), representing 6.08% of the total population. This study detected 184 cervical and trochanteric fractures of the proximal femur in 1994 in Tangshan (127 men and 57 women). The overall incidence or rate of the combined number of cervical and trochanteric fractures was 25 fractures per 100,000 population per year for men and 12 for women. There were a total of 147 cervical fractures (80%) and 37 trochanteric fractures (20%). The incidence of the combined number of cervical and trochanteric fractures in patients over 70 years of age increased to 108 for men and 156 for women. The incidence of hip fractures increased with age in both sex groups, especially in women over 65. Severe trauma fractures happened more often in younger groups, and mainly occurred in men, which may be a result of the particular composition of the population in Tangshan, which is young and male dominated. In addition, because Tangshan is an industrial city, many of its citizens are involved in occupations requiring a high level of physical activity. PMID- 10701164 TI - Epidemiology of cervical and trochanteric fractures of the proximal femur in 1996 in Kaohsiung City, Taiwan. AB - The goal of this study was to determine the incidence of cervical and trochanteric fractures of the proximal femur in 1996 in Kaohsiung City, Taiwan. Kaohsiung City is the industrial and commercial center of southern Taiwan, with a population of 1,433,621 in 1996. The number of individuals over 65 years of age accounted for 6.2% of the total population. Data from the archives of reimbursement of the National Health Insurance program were used to investigate the incidence of fractures of the proximal femur. This study detected 580 cervical and trochanteric fractures (40.5 fractures per 100,000 population per year) in 261 males (35.8 fractures per 100,000 men per year) and 319 females (45.3 fractures per 100,000 women per year), with 420 (72%) of these fractures occurring in individuals over 65 years of age. The age-specific incidences of cervical and trochanteric fractures increased exponentially with age in both genders. The overall ratio of cervical to trochanteric fractures was 1:1.04. The mean ages of women with cervical or trochanteric fractures (71.6 and 74.0 years, respectively) were significantly higher than those of males (59.9 and 64.8 years, respectively; P < 0.01). The age-adjusted incidences of fractures of the proximal femur in Kaohsiung City were higher than in other Asian countries, but were lower than in Western countries such as the United States and Norway. The urban lifestyle and low daily calcium intake may be responsible for this increased incidence of proximal femoral fractures in Kaohsiung City. PMID- 10701165 TI - Fracture threshold in the Thai elderly and bone mineral density evaluation. AB - Fractures in the elderly as the result of minor trauma or normal physiological stress in daily activities usually occur in load-bearing body areas such as the lumbar spine and the neck or trochanter of the femur, causing high morbidity and mortality. Surveillance of high-risk Thai elderly with low bone mineral density by determining the cutoff point as the fracture threshold may guide us in the proper management for preventing these unpleasant events. Of 329 elderly with age range of 50-110 years, 63 with lumbar spine fracture and 55 with hip fracture were descriptively and studied prospectively during May 1997 to December 1998 at Pramongkutklao Hospital. Bone mineral density (BMD) was analyzed to determine the fracture threshold using a receiver-operating characteristic (ROC) curve and compared with the total BMD at the lumbar spine and proximal femur. The cutoff point of the lumbar BMD at 0.799 g/cm2 (78.30% sensitivity, 73.60% specificity, and 74.5% accuracy) yields the likelihood of lumbar spine or hip fracture. For the femoral BMD, the cutoff point at 0.649 g/cm2 (92.5% sensitivity, 73.2% specificity, and 73.05% accuracy) is also used to predict the likelihood of lumbar spine or hip fracture. Of the nonfracture group, 27.33% had a total BMD value below the fracture threshold. In conclusion, early prevention among the elderly to decrease the risks of fracture is very important. Also, the detection and fracture prevention for normal population who had BMD below the fracture threshold are interesting. The BMD measurements and the loss of bone mass in Thai elderly people and the other risks of fracture need further studies. PMID- 10701166 TI - The impact of AIDS on medical ethics. PMID- 10701167 TI - Ignorance is bliss? HIV and moral duties and legal duties to forewarn. AB - In 1997, a court in Cyprus jailed Pavlos Georgiou for fifteen months for knowingly infecting a British woman, Janet Pink, with HIV-1 through unprotected sexual intercourse. Pink met Georgiou in January 1994 whilst on holiday. She discovered that she had contracted the virus from him in October 1994 but continued the relationship until July 1996 when she developed AIDS. She returned to the UK for treatment and reported Georgiou to the Cypriot authorities. There have been a number of legal cases involving deliberate transmission of HIV, but most have involved forced exposure to infected bodily fluids for example, rape or biting, and have been dealt with using the existing legislation for rape or assault. While it is often difficult to prove responsibility for transmission in cases of forced exposure to HIV, it is even more contentious in cases like those of Janet Pink where an individual has consented to sex but claims that he/she was not forewarned of his/her partner's HIV-positive status. At present there is no specific criminal offence of having unprotected sexual intercourse without disclosing one's HIV-positive status but a prosecution could possibly be brought under any one of a number of existing offences. Perhaps a change of policy needs to be considered. The Home Office has issued a consultation document which outlines a proposal that will allow the criminalization of intentional transmission of diseases, like HIV, that are likely to cause serious harm. This revised legislation would cover all other potentially fatal diseases (including salmonella and legionnaire's disease, for instance) but seems primarily to be targeted at HIV transmission. Should transmission of HIV through consensual sex, without the HIV-positive status of the individual being disclosed, be an offence? This question, and that of whether there is a moral obligation to disclose a positive HIV status prior to having a sexual relationship is the subject of this paper. PMID- 10701168 TI - The ethics of anonymized HIV testing of pregnant women: a reappraisal. AB - Seroprevalence monitoring of HIV in pregnant women by anonymized unlinked testing has been widely adopted in the UK and other countries. The scientific rationale is to eliminate participation and selection bias. The ethical justification is that the public good outweighs any harm to individuals. The assumption has been that individuals have had their autonomy respected by the offer of informed consent. In the light of new scientific evidence, it is doubtful that the public good is best served by the continuation of anonymously testing women receiving antenatal care. It is submitted that it is no longer ethical for health professionals to refrain from informing pregnant women of the benefits of voluntary named testing, or to request their consent to anonymized testing. The legal and moral concept of duty of care is examined, and the abrogation of this duty through anonymization is explained. PMID- 10701169 TI - Infectious health care workers: should patients be told? AB - The risk of transmission of HIV or hepatitis B from infectious health care workers to patients is low. However, inadvertent exposure causes great concern amongst patients of an infected health care worker. The patients of a Scottish dentist diagnosed hepatitis B e antigen positive were informed by letter of their exposure. A sample of patients was sent a postal questionnaire. Most (56%) respondents reported feeling anxious on receiving the letter but almost all (93%) thought patients should always be informed following treatment by an infectious health care worker, although the risk was very small. We discuss clinical and ethical factors relating to informing patients following exposure to an infectious health care worker. We suggest that a balance should be struck between patients' wishes to know of risks to which they have been exposed, however small, and the professional view that when risks are negligible, patients need not be informed. PMID- 10701170 TI - Ethical considerations in international HIV vaccine trials: summary of a consultative process conducted by the Joint United Nations Programme on HIV/AIDS (UNAIDS). AB - Research that is initiated, designed or funded by sponsor agencies based in countries with relatively high social and economic development, and conducted in countries that are relatively less developed, gives rise to many important ethical challenges. Although clinical trials of HIV vaccines began ten years ago in the US and Europe, an increasing number of trials are now being conducted or planned in other countries, including several that are considered "developing" countries. Safeguarding the rights and welfare of individuals participating as research subjects in developing countries is a priority. In September, 1997, the Joint United Nations Programme on HIV/AIDS (UNAIDS) embarked on a process of international consultation; its purpose was further to define the important ethical issues and to formulate guidance that might facilitate the ethical design and conduct of HIV vaccine trials in international contexts. This paper summarises the major outcomes of the UNAIDS consultative process. PMID- 10701171 TI - Live attenuated vaccine trials in medically informed volunteers: a special case? AB - A group of activist clinicians have offered to volunteer for clinical trials of live attenuated HIV vaccines. This has provided an important conceptual challenge to medical ethics, and to work on the development of HIV vaccines. In exploring these issues, this article highlights how the HIV field has altered the content as well as the tone of ethical discourse. The balance of expertise and authority between research subjects and triallists is profoundly changed, raising questions about the limits of voluntarism and differing perspectives on risk-benefit analysis. Care is needed to ensure that the novelty of the situation does not confuse the central ethical and scientific issues. PMID- 10701172 TI - Narrowing the gap: access to HIV treatments in developing countries. A pharmaceutical company's perspective. AB - The advent of new antiretroviral medicines means that the effects of HIV can now be curbed, but only one in twenty infected people have so far benefited. For those living in developing countries, the new treatments are practically unattainable. Governments, UNAIDS and pharmaceutical companies recognise this only too well and have rethought established assumption in order to try and overcome the challenges posed by cost, inadequate health services and unreliable local supply of medicines. PMID- 10701173 TI - Ethics in the laboratory examination of patients. AB - Various value problems are connected with the clinical examination of patients. The purpose of this literature review is to clarify: 1) in which patient examinations ethical problems are generally found; 2) what kind of ethical problems are found in the different phases of the examination process, and 3) what kind of ethical problems are found in connection with the use of examination results. Genetic testing, autopsy, prenatal and HIV examinations were ethically the most problematic laboratory examinations. The most problematic phase in the laboratory examination process proved to be the pre-analytic phase. At present the results of laboratory examination are used more and more often for the prediction of diseases. The problems appear when the examination results are used for discrimination and stigmatization. Because of the lack of empirical ethical research, it is important to chart empirical knowledge about present value conflict situations involved in the laboratory examination process. PMID- 10701174 TI - Frontiers in care: a case of compulsory treatment in AIDS dementia. Case study and commentaries. AB - A patient with AIDS dementia was confronted and compulsorily prevented from flying out of the country before being admitted against his will to hospital. While finding this on balance justified in the circumstances the commentators raise moral questions about the levels of care in general practice and within the couple's own relationships. PMID- 10701175 TI - An AIDS lexicon. PMID- 10701176 TI - Comparative efficiencies of different non-toxic microalgal diets in detoxification of PSP-contaminated oysters (Crassostrea gigas Thunberg). AB - Experimental PSP contamination of adult Pacific oysters (Crassostrea gigas) by the toxic dinoflagellate Alexandrium minutum Halim (120 cells.mL-1 continuously maintained in each flume) was carried out in a recirculated seawater system to obtain toxin levels above the safety threshold. In these conditions, 150 to 300 micrograms STX.eq.100 g-1 of shellfish tissues were produced at 16 degrees C within 8 to 15 days, corresponding to field values observed along French coasts. Diets based on non-toxic flagellates or diatoms were then used to detoxify the contaminated oysters. Despite large individual variations in toxin levels at the end of the contamination period, detoxification times were of the same order of magnitude (3 to 4 days), reaching a toxin level equal to or less than the safety threshold. These variations were most likely related to marked individual variability in valve and/or clearance activities. No significant differences in detoxification rates were found when oysters were fed Isochrysis galbana, Tetraselmis suecica, Thalassiosira weissflogii, or Skeletonema costatum. The different biochemical compositions of each algal species appeared to have no significant effect on detoxification rates. GTX2/GTX3 were the dominant compounds found in shellfish tissues during depuration, whereas C toxins were quite low (< 2 micrograms STX.eq.100 g-1) and STX or NeoSTX undetectable. These results do not suggest any bioconversion of paralytic toxins but indicate good correlation between the toxin composition of Alexandrium and oyster tissues. PMID- 10701177 TI - Detection of nerve growth factor (NGF) in venoms from diverse source: isolation and characterization of NGF from the venom of honey bee (Apis melifera). AB - Pearce (1973) reported the absence of NGF in the venoms of bees, scorpions, spiders, and toads. Contrary to the negative findings in the past, results of this research prove the presence of NGF in bee and scorpion venoms. Venoms from various species of snake, bee, scorpion, and toad were screened by two methods: immunological test ELISA using antibodies versus mouse NGF and venom NGF and the biological test of neurite outgrowth, the characteristic of NGF on PC cells. The presence of NGF was detected in snake, bee, and scorpion venoms, but not in toad venom by these tests. NGF was isolated from bee venom by HPLC fractionation using ion exchange chromatography. The molecular weight of bee NGF was found to be 14.0 kDa resolving into a single band by PAGE. The biological activity of bee NGF on PC12 cells was found to be 1/10 of the venom NGF. PMID- 10701178 TI - Histological and histochemical alterations in the liver following intramuscular injection with a sublethal dose of the Egyptian cobra venom. AB - In the present study, the effects of intramuscular (i.m.) injection of a sublethal dose (0.015 microgram/gm b.wt.) of Naja haje venom were histologically and histochemically examined in the hepatic tissues of rabbits after 3, 6, and 12 hr. of envenomation. Three hours after venom injection, the hepatic cells showed a generalized cytoplasmic granulation and cellular swelling accompanied with narrowing of the sinusoidal spaces. Occurrence of inflammatory cells and hypertrophy of Kupffer cells were also noticed. After 6 hr. of envenomation, the hepatic tissues revealed severe cellular swelling, cytoplasmic deterioration, nuclear pyknosis, and appearance of numerous basophilic granules. The central veins were engorged with hemolyzed blood. Hepatic tissues investigated after 12 hr. of envenomation exhibited highly damaged hepatic cells to the extent that the individual cell cannot be identified and remnants of cell debris were seen intermixed together. Some hepatic cells were intensively swollen and their contents were dissoluted except for a few dusty cytoplasmic granules and pyknotic nuclei. The histochemical observations showed a time dependent depletion in polysaccharide, lipid, and protein contents in the hepatic cells of the envenomed groups. As for the nucleic acids, slight depletion of RNA together with no changes in DNA contents were observed by 3 hr. of envenomation. Nevertheless, severe degrees of RNA depletion and moderate contents of DNA were recorded in 6 hr. envenomed tissues. Highly obvious depletion of RNA and DNA were demonstrated by 12 hr. after venom injection. From the above results, it is obvious that cobra venom induces a hepatotoxic action reflected by alterations in the histological and histochemical patterns of the hepatic tissues. These alterations are initiated at early stages of envenomation and could indicate a disturbance in the functional activities of the liver during envenomation. PMID- 10701180 TI - Mediators and renal hemodynamics in Russell's viper envenomation. AB - The effects of Russell's viper venom on vasoactive mediators and renal hemodynamics were studied in five mongrel dogs. Intravenous administration of Russell's viper venom to the dogs caused a reduction of mean arterial pressure, renal blood flow, and glomerular filtration rate. The filtration fraction was decreased. This was accompanied by a rise in plasma norepinephrine, endothelin, 6 keto-PGF1 alpha, a stable metabolite of PGI2, and TXB2, a metabolite of TXA2. Plasma levels of epinephrine and dopamine showed no significant changes. The increase of plasma levels of both vasodilatator and vasoconstrictor were critical to systemic and renal hemodynamics. While vasodilatation predominated in the systemic circulation and resulted in hypotension, vasoconstriction played a major role in decreasing renal hemodynamics. Decreased renal blood flow and decreased glomerular filtration rate were the result of renal vasoconstriction and hypotension. PMID- 10701179 TI - Nature of the postsynaptic action of crotoxin at guinea-pig diaphragm end-plates. AB - Crotoxin is known to desensitize the nicotinic receptor of Torpedo marmorata and Electrophorus electricus electroplaques. The aim of the present study was to elucidate whether the postsynaptic effect of crotoxin at a mammalian muscle end plate is also caused by receptor desensitization or results from a curaremimetic action. For this purpose, we investigated the action of 4-aminopyridine (4-AP) on crotoxin-induced blockade of miniature end-plate potentials (m.e.p.p.s) and of the depolarization of end-plates produced by carbachol. The experiments were carried out in guinea-pig diaphragms bathed in Tyrode solution at 37 degrees C and gassed with 95% O2, 5% CO2. The potentials were measured with conventional techniques using glass microelectrodes. Even at low concentrations, crotoxin blocked the m.e.p.p.s and this blockade was antagonized by 4-AP. Neostigmine was without effect. 4-AP did not restore the m.e.p.p.s blocked by either d tubocurarine (dTc) or beta-bungarotoxin (beta-BTX). 4-AP also antagonized the crotoxin-induced blockade of the end-plate depolarization produced by carbachol. These results show that the postsynaptic effect of crotoxin at the guinea-pig muscle end-plate also results from nicotinic receptor desensitization. PMID- 10701181 TI - Botanical biocides. 4. Mosquitocidal activity of certain Thymus capitatus constituents. AB - Successive extraction of Thyme plant, Thymus capitatus (L.) Hoffm. and Link (Lamiaceae), by different solvents of increasing polarity, showed that potency was highly attributed to the non-polar fraction (e.g., petroleum ether) when tests were carried out against the larvae and adults of Culex pipiens (L). Of special concern to the mosquitocidal activity, the following fractions and isolates were recognized: the volatile oil, unsaponifiable portion, and certain compounds isolated from the unsaponifiable portion (e.g., Thymol, alpha-Amyrin, Carvacrol + beta-Caryophyllene). The volatile oil, Thymol, and the unsaponifiable portion proved high larvicidal potency against the tested insect (LC50 = 49.0, 58.0, and 100.0 ppm, respectively). Non-lethal concentrations of these substances synergized the toxicity of Malathion, while induced additive or antagonistic effects when mixed with Permethrin or Pirimiphos-methyl. The unsaponifiable portion and volatile oil showed the highest adulticidal potency (LC50 = 0.0070 and 0.0076 mg/cm2, respectively). The efficacy of the tested candidates as repellent agents was found in the following order: unsaponifiable portion > alpha Amyrin > Thymol > volatile oil > Carvacrol + beta-Caryophyllene. Thymol as well as volatile oil affected egg hatchability, causing Sterility Indices accounting for 0.70 and 0.74, respectively, while the unsaponifiable portion showed lower degree of sterility (0.81). The results obtained in this study may shed light on the importance of T. capitatus as a probable source of some biologically active agents for mosquito control in the future. PMID- 10701182 TI - Survey of paralytic toxins in shellfish in southern Taiwan between 1995 and 1997. AB - To establish the safety data of shellfish in southern Taiwan, a total of 3,074 specimens of 30 shellfish species were seasonally collected from August 1995 to March 1997. These samples were assayed for the presence of tetrodotoxin (TTX) and paralytic shellfish poisons (PSP) by the bioassay methods. It was found that some major shellfish including oyster, clam, ear shell, and purple clam were nontoxic, but four species, Babylonia formosae, Niotha clathrata, Natica lineata, and Natica vitellus, were toxic. The toxic percentages of these four species was 1% in B. formosae, 56% in N. clathrata, 37% in N. lineata, and 23% in N. vitellus. The toxic composition was TTX in B. formosae. In the other three shellfish types the toxic composition was TTX and PSP. PMID- 10701183 TI - Brain delivery of biotinylated NGF bounded to an avidin-transferrin conjugate. AB - Recent study showed that transferrin receptors were concentrated on the plasma membrane of brain endothelia cells and mediated transcytosis of transferrin (Tf) through the blood-brain barrier (BBB). This property allows the transferrin to act as the brain drug transporter vector. The present investigation examined the pharmacokinetic behavior of nerve growth factor (NGF), which was conjugated to transferrin by the avidin/biotin technology, especially its brain-uptake efficiency. The area under the plasma concentration curve and the mean residence time were not significantly different for either bio-NGF or bio-NGF/AV-Tf. At the first hour after single intravenous injection, the BBB permeability surface area product of bio-NGF/AV-Tf was 0.77 microliter/min/g; it was about 8-fold higher than that of bio-NGF, and equal to that of AV-OX26. The delivery of bio-NGF/AV-Tf to brain was 0.075% of injected dose per gram brain, and it was 5-fold higher than that of bio-NGF, and 2-fold higher than that of AV-OX26. In summary, these studies demonstrated that the use of Tf as brain drug delivery vector was as effective in transporting biotinylated therapeutics as OX26, and avoided the disadvantages of its antigenicity. PMID- 10701184 TI - Fumonisin B1 from the fungus Fusarium moniliforme causes contact toxicity in plants: evidence from studies with biosynthetically labeled toxin. AB - Fumonisin B1 (FB1) is the most abundant of a series of sphingosine analog mycotoxins produced by the fungus Fusarium moniliforme, a ubiquitous contaminant of stored corn (maize) worldwide. FB1 exhibits a variety of biological activities including phytotoxicity, which is of particular interest for its potential role as a virulence factor to facilitate invasion of plant tissues by the fungus. Droplets of FB1 solution applied to the leaf surface of jimsonweed, black nightshade, and susceptible tomatoes caused necrosis, growth inhibition, and death. With Arabidopsis thaliana grown on agar plates, an IC50 (concentration causing half maximal phytotoxicity) of less than 1 ppm was observed. [3H]FB1 was prepared by biosynthetic incorporation of commercially-available radiolabeled presumptive precursors into the toxin in rice medium solid cultures of F. moniliforme JW#1. The labeled toxin produced by incorporation of [9,10( 3)H]palmitate induced phytotoxic symptoms identical to unlabeled material, indicating it had full biological activity. The area of necrosis on treated leaves was similar in light and dark treated plants. Using liquid scintillation counting to quantify radioactivity in excised plant parts, over 95% of the [3H]FB1 radioactivity applied to leaves of light or dark-treated plants was recovered from the treated leaf. When [3H]FB1 was applied to a wound site on target plants, severe damage occurred at the site of FB1 application and in tissue above the site. These results indicate that FB1 applied to intact surfaces of target plants exhibits primarily contact activity. Translocation of FB1 is limited, occurring only when FB1 is applied to a wound site, and it results in damage to tissue above the point of application, indicating that FB1 is xylem mobile. PMID- 10701185 TI - What is your diagnosis? Axial osteosarcoma. PMID- 10701186 TI - Palmoplantar hyperkeratosis in Irish terriers: evidence of autosomal recessive inheritance. AB - An abnormal development of the epidermis of the footpad was observed in Irish terriers. At the age of six months, the affected animals developed smooth parchment-like footpads. The pad epidermis then hardened and grew lateral cone like protrusions of up to 5 mm in diameter. Fissures and cracks developed and these predisposed the animal to secondary infection. The repeated occurrence in subsequent generations led to the assumption of a hereditary form of hyperkeratosis. Evidence for an autosomal recessive mode of inheritance was derived from a retrospective analysis of the breeder's records. The clinical, histopathological and ultrastructural features of the disease are presented and the genetic transmission and its implications discussed. PMID- 10701187 TI - Influence of selective breeding on the prevalence of chorioretinal dysplasia and coloboma in the rough collie in Sweden. AB - A total of 8204 rough collies, representing 76 per cent of all collies registered by the Swedish Kennel Club between 1989 and 1997, were examined before 10 weeks of age for collie eye anomaly (CEA). All dogs were permanently identified and examination results were registered and computerised. The policy of breeders during the study period was to select against coloboma in breeding stock, but to allow breeding of chorioretinal dysplasia (CRD) affected animals. The prevalence of CRD increased significantly from 54.2 per cent to 68.1 per cent (P < 0.001) from 1989 to 1997, while the prevalence of coloboma did not (8.3 per cent to 8.5 per cent, P = 0.4). These results are not compatible with a simple, recessive, autosomal inheritance for the entire CEA complex. PMID- 10701188 TI - Capillary blood sampling from the ear of dogs and cats and use of portable meters to measure glucose concentration. AB - Two new methods for collection of capillary blood from the ear of dogs and cats for the measurement of blood glucose concentration using portable blood glucose meters (PBGMs) are described. The first method uses a lancing device after pre warming the ear, while the second employs a vacuum lancing device. Both methods generated blood drops of adequate size, although the latter method was faster and easier to perform. Accuracy of the two PBGMs was evaluated clinically and statistically. Although assessment of statistical accuracy revealed differences between the PBGMs and the reference method, all of the PBGM readings were within clinically acceptable ranges. Measurement of capillary blood glucose concentration is easy to perform, inexpensive and fast. It may be used by owners to determine blood glucose concentrations at home, and could serve as a new tool for monitoring diabetic dogs and cats. PMID- 10701189 TI - Multiple endocrine neoplasia type 1 in a crossbred dog. AB - Multiple endocrine neoplasia type 1 was diagnosed in a 12-year-old male crossbred dog. Relevant history included polyuria and polydipsia of four months' duration. Physical examination revealed abdominal enlargement, seborrhoea and polypnoea. Diagnostic tests indicated hypercalcaemia, elevated serum alkaline phosphatase and alanine aminotransferase, an exaggerated response to adrenocorticotropic stimulation of the adrenal gland, lack of cortisol suppression with a low dose dexamethasone suppression test and suppression of cortisol secretion with a high dose dexamethasone test. An enlarged right parathyroid gland was removed surgically and confirmed histopathologically to be a parathyroid adenoma. The pituitary-dependent hyperadrenocorticism was treated successfully with mitotane for 14 months before the patient was euthanased for an unrelated problem. PMID- 10701190 TI - Use of a mucosal advancement flap for the treatment of nasopharyngeal stenosis in a cat. AB - A three-and-a-half-year-old male neutered domestic short-haired cat was presented with a three-year history of stertor and intermittent open-mouth breathing. No airflow was detected from either nostril when checked using a cold slide test. Oropharyngeal swabs were positive for calicivirus, while skull radiographs were suggestive of a dorsal deviation of the soft palate. The diagnosis of nasopharyngeal stenosis was confirmed via cannulation of the nasal passages and direct examination of the oropharynx under general anaesthesia. A midline approach through the soft palate was used to excise the adhesions. The resulting defect was reconstructed by advancement of a mucosal flap elevated from the dorsal nasopharynx and laryngopharynx. The cat was free of clinical signs 28 months later. PMID- 10701191 TI - Radial carpal bone fractures in 15 dogs. AB - Between 1977 and 1992, 15 adult dogs were presented to the Universities of Bristol, Glasgow, Liverpool and London with fractures of the radial carpal bone which had occurred without a known episode of violent trauma. Eleven of the dogs were boxers and the other breeds affected were a Pharaoh hound, rottweiler, springer spaniel and a greyhound. The condition was bilateral in three cases. Eleven dogs were treated surgically by cast immobilisation (four cases), fragment removal (one case), lag screw fixation (three cases), pan-carpal arthrodesis (one case) and lag screw reduction followed by pan-carpal arthrodesis (two cases). The remaining four dogs were treated conservatively with rest and tactical use of non steroidal anti-inflammatory drugs. Follow-up periods ranged from two months to 11 years. Varying degrees of lameness, which tended to be exacerbated by exercise, persisted in all cases. The radial carpal bone appears to have at least three separate centres of ossification which eventually fuse; their planes of fusion correspond approximately to the two main fracture lines. These areas of fusion could be weak points within the radial carpal bone. PMID- 10701192 TI - Pyogranulomatous blepharitis in two dogs. AB - Two cases of pyogranulomatous blepharitis (inflammation of the lid margins) in the dalmatian are described. The diagnosis was confirmed on biopsy of the lid lesions. Bacteriology performed in one case was negative. Both cases responded to treatment: one responded well to a course of systemic steroids (prednisolone 1 mg/kg at a decreasing dose over three weeks), while the other, which was negative on culture, responded to a six-week course of cephalexin (30 mg/kg twice daily). This second dog also presented with a localised lymphadenopathy; the owner had suffered a similar reaction three years previously as a result of a penetrating injury by a pyracantha thorn. PMID- 10701194 TI - [Rise and fall of mobile preventive medicine]. PMID- 10701193 TI - [Tropical medicine, travel medicine and humanitarian medicine: is there a place for the 3 trainings in France?]. PMID- 10701195 TI - [Scuba diving in the tropics]. PMID- 10701196 TI - [Ketoconazole]. PMID- 10701197 TI - [Indonesia: the largest archipelago in the world]. PMID- 10701198 TI - [2020, the odyssey of tropical medicine: the emergence of non-transmissible diseases]. PMID- 10701200 TI - [Antibiotic therapy in developing countries]. PMID- 10701199 TI - [French Federation of Tropic Medicine and International Health Institute: a new tool for training and research]. PMID- 10701201 TI - [Preliminary evaluation of usable indicators during a control program for urinary bilharziosis in Niger]. AB - The performance of several indirect screening tests was evaluated during the start-up phase of a urinary schistosomiasis control program in Niger. Urine tests were carried out on a total of 354 children attending 3 primary schools on five consecutive days. Tests included filtration of 10 ml of urine, search for microscopic hematuria using reagent strips, and gross examination of urine. In addition a questionnaire was administered on the first day to identify signs of dysuria and hematuria. Repeat testing had a strong effect on the epidemiological profile of urinary schistosomiasis in the 3 schools. Although day-to-day counts varied greatly, egg excretion could be considered as high in all infected subjects. The screening sensitivity of urine filtration was low when the level of endemicity was moderate (up to 55 p. 100). Microscopic hematuria was common. However the sensitivity of this method was overestimated in comparison with urine filtration alone and use of reagent strips can be inconvenient. Using carefully defined diagnostic criteria, gross examination of urine was as effective as urine filtration and easier to perform. The value of the questionnaire for evaluation of morbidity was low despite relatively good performance of the diagnostic techniques. The children's responses concerning hematuria were not objective and questions concerning dysuria were poorly understood and time-consuming. In the next phase of study, these findings will be validated by ultrasound imaging. PMID- 10701202 TI - [Relationship between the intensity of Loa loa filariasis transmission and prevalence of infections]. AB - Filarial loiasis differs from other filariases in that most infected subjects are amicrofilaremic. This difference raises the notion of occult infection. The aim of this study was to evaluate the relationship between the intensity of transmission and incidence of infection. For this purpose we determined the incidence of loiasis both microscopically and by PCR in 201 subjects from three villages in the province of Haut Ogooue in Gabon. Intensity of transmission, expressed in ATP (annual transmission potential) in these villages was estimated to be 250 infecting larvae per individual per year (L3/man/yr) in Moyabi, 180 L3/man/yr in N'dokaye, and 43,000 L3/man/yr in Okoumbi. Although there was no significant difference between the three villages with regard to the incidence of microfilaremia (21 p. 100 and 22 p. 100), the incidence of occult infection, i.e., positive PCR in amicrofilaremic subjects, was 45 p. 100 in Moyabi, 79 p. 100 in N'dokaye and 80 p. 100 in Okoumbi. The overall incidence of loiasis was 57 p. 100 in Moyabi and 85 p. 100 in both N'dokaye and Okoumbi. These findings demonstrate that the incidence of loiasis is correlated with the intensity of transmission (p < 0.001), especially in children. Taking this information into account will improve control of Loa loa in endemic areas. PMID- 10701203 TI - [Diarrhea in urban agricultural workers in Nouakchott in Mauritania]. AB - Nearly 200 million people in the developing world are dependent or urban gardening for food and income. This practice has been accelerated by the droughts of recent decades which have forced more and more migrants into urban areas. Numerous potential health hazards have been attributed to urban gardening but the exact risks in Sahelian areas remain unclear. The purpose of this cross-sectional study was to evaluate the incidence of diarrhea at the Tel Zatar gardening site in urban Nouakchott, Mauritania. In addition, a case-control study was carried out to identify risk factors for diarrhea in function of gardeners' activity and living conditions. Statistical analysis was performed using univariate and logistical regression methods. The annual incidence of diarrhea ranged from 6.9 (IC95 p. 100 = 5.0-8.8) to 8.5 (IC95 p. 100 = 6.2-10.8) episodes per gardener and year. Multivariate analysis identified four significant risk factors. Two of these factors were unrelated to gardening, i.e., not having spent more than USD 3.50 the previous day (odds ratio (OR = 2.8, IC95 p. 100 = 1.01-7.81) and poor food hygiene (cooking outside (OR = 4.69, IC95 p. 100 = 1.06-20.83). The other two factors were regular consumption of raw vegetables (OR = 25.5, IC95 p. 100 = 2.0-32.0) and use of untreated well water (OR = 3.85, IC95 p. 100 = 1.08-14.29). Unprotected well water was the cause of 59.2 p. 100 of diarrheal episodes reported by gardeners at Tel Zatar. The results of this study confirm that vegetable production in urban gardens such as Tel Zatar is associated with health risks. Public health measures should address not only the garden sites but also domestic hygiene. PMID- 10701204 TI - [Acquired antibiotic resistance in Madagascar: first evaluation]. AB - The purpose of this study was to evaluate the incidence of acquired resistance to antibiotics in Madagascar. Testing was carried out on total of 1267 strains of medically significant bacteria isolated from specimens sent to the Pasteur Institute of Madagascar in Antananarivo between October 1997 and October 1998. Antibiograms were performed using the diffusion technique on gel media with antibiotic disks. Results were read according to the criteria of the Antibiogram Committee of the French Society of Microbiology. Preliminary findings documented a high incidence of resistance to widely available, low-price antibiotics including penicillin G and tetracycline for which 84 p. 100 and 65 p. 100 of Staphylococcus aureus respectively demonstrated resistance; tetracyclin to which 80 p. 100 of streptococcus were resistant; and ampicillin, cotrimoxazole, and phenicoles to which 60 p. 100, 60 p. 100 and 28 p. 100 of Escherichia coli respectively and 77 p. 100, 83 p. 100, and 71 p. 100 of Shigella sp. respectively were resistant. Second-line antibiotics including penicillin M, macrolides, nalidixic acid, and nitrofuranes were still relatively active, thus providing an effective alternative. Newly developed antibiotics such as fluoroquinolones and third-generation cephalosporines were highly effective but a few resistant strains were observed. Although not representative of Madagascar as a whole, the findings of this preliminary study indicate that acquired resistance must be taken into account in designing simplified decision charts for front-line laboratories, that appropriate information must be made available to health care workers, and that further testing is needed to monitor the evolution of antibiotic resistance. PMID- 10701205 TI - [Evidence of risk factors for condom breakage among French military personnel stationed overseas]. AB - The French Army Medical Service has been distributing free condoms to overseas personnel since 1989. An epidemiologic study conducted in Cambodia showed that the rate of condom failure during protected sex was 16.3 p. 100. In view this high failure rate, a study to evaluate safety guidelines for condom use was deemed necessary. This report describes the results of an inquiry designed to identify risk factors for condom breakage. The study population included 124 servicemen stationed overseas who consulted an army physician following condom failure. An anonymous questionnaire was completed by the physician with the informed consent of the patient. Data was analyzed by multiple correspondence analysis. Anogenital intercourse and alcohol abuse were the main risk factors for condom failure. Oral sex before penetration, carrying the condom in the pocket, and improper application by the sex partner were also risk factors for failure. The major finding of this study was that 64 of the 124 (51.6 p. 100) denied any sexual activity or improper handling that might have led to condom breakage. Although condoms are still indispensable for prevention sexually transmitted diseases, the results of this study show that they do not provide absolute protection. Health care officials should inform the public as to the risk factors for breakage of latex condoms. PMID- 10701206 TI - [Adults with sickle cell anemia in Senegal. Clinical study of 40 homozygote subjects]. AB - While the well-documented life expectancy of patients with homozygous sickle cell anemia (SS) is 40 years in industrialized countries, this question remains unanswered in black Africa. The purpose of this prospective study was to establish the clinical phenotype for Senegal. A severity score based on 12 clinical, laboratory, radiological, and prognostic findings was calculated and correlated with age and hemoglobin F level. A total of 40 SS homozygotes over 15 years of age (mean age: 25 years) were hospitalized between January 1996 and January 1998 at the Principal Hospital in Dakar. The most common events requiring hospitalization were vasoocclusive phenomena (n = 26) but the incidence of these complications declined significantly with age (p < 0.05). The mean hemoglobin level was 4.4 mmol/l and the mean hemoglobin F level was 6.2 p. 100. The incidence of visceral involvement was low (lithiasic vesicules in 17 cases, necrosis of the head of the femur in 7, and abnormal cardiac ultrasound findings in 10). Only one patient died during the study. No correlation was found between severity score and either age or hemoglobin F level. These findings confirm that the phenotype is less severe in Senegal. However they also show that organ damage is common by the time that patients reach adulthood and thus underline the need for prevention and education to improve survival of SS homozygotes in Senegal. PMID- 10701207 TI - [Diagnostic test to identify human Plasmodium species by the quantitative buffy coat test]. AB - The quantitative buffy coat system (QBC Test) was designed for rapid diagnosis of malaria by identifying the presence of hemoparasites. The main drawback of the technique is failure to identify the Plasmodium species. The purpose of this study was to attempt to remedy this problem by studying the distribution of the parasites at the bottom of the test tube. Indeed since the QBC Test is based on gradient centrifugation of blood components, the distribution of the parasites in the test tube depends on density. Blind QBC Tests were performed on specimens obtained from two different batches, i.e., one from France and the other from Burkina Faso. Distribution curves were obtained by counting the number of parasites in all microscopic fields in the five-millimeter test tube. Our findings showed differences in distribution curve depending on species. For Plasmodium falciparum, the number of parasites was nearly the same in all fields suggesting that the arrangement of the parasites in the QBC Test tube was linear. For Plasmodium vivax and Plasmodium ovale, the number of parasites was markedly lower near the cap of the tube suggesting that a non-linear arrangement with a decreasing number of parasites toward the top of the tube. In 97 p. 100 of cases, we were able to propose a differential diagnosis of Plasmodium falciparum versus Plasmodium vivax or Plasmodium ovale. However it was not possible to distinguish between Plasmodium vivax and Plasmodium ovale. In case of mixed infection it would be difficult to distinguish Plasmodium falciparum from the other species. The ability to identify Plasmodium species would add to the advantages of the rapid and sensitive QBC Test. PMID- 10701208 TI - [Chronic genital ulcerations and HIV infection: 29 cases]. AB - Genital ulcers are common manifestations of infectious disease. The incidence of genital ulcers featuring a chronic course has increased since the beginning of the AIDS epidemic. The purpose of this 18-month cross-sectional study was to determine the main infectious causes of chronic genital ulcers (CGU) and their correlation with HIV infection. A total of 29 patients with CGU defined as an ulcer showing no sign of healing after more than one month were studied. Mean age ranged from 24 to 54 years. The male-to-female sex ratio was 1:5. The etiology was herpes in 19 cases (65.5 p. 100), chancroid in 6 cases (20.6 p. 100), streptococcal infection in 2 cases (6.8 p. 100), Pseudomonas aeruginosa infection in 1 case (3.4 p. 100) and cutaneous amibiasis in 1 case (3.4 p. 100). Twenty-two patients (75.8 p. 100) presented HIV infection including 16 with HIV1 and 6 with HIV1 and HIV2. All patients with herpes were HIV-positive. Eighteen of these patients were in stage C3 of HIV infection. Genital herpes was the main etiology of UGC in patients with HIV infection (p < 0.001). Conversely chancroid was the main etiology in patients without HIV infection (p < 0.05). This finding suggests that herpetic CGU is highly suggestive of AIDS whereas chancroid CGU is not. Although syphilis is widespread in Africa, it was not a cause of CGU in this study. Search for herpes simplex virus or Haemophilus ducreyi in patients with CGU is an important criteria for presumptive diagnosis of AIDS in Africa. PMID- 10701209 TI - [Management of severe malaria in children in developing countries. A protocol for economic evaluation]. AB - This prospective one-year study was conducted as a preliminary phase to setting up a protocol for economic appraisal of management of severe malaria at Albert Royer Children's Hospital in Dakar, Senegal. Data was routinely collected using a standardized checklist. The four key indicators chosen for this study were nurse workload, adequacy of care (number of patients receiving adequate care), direct cost, and mortality rate. The mean daily care workload was estimated to be 27.2 minutes. This indicator assesses the relationship between supply and demand. Based on 5 criteria, care was considered as adequate in 54.5 p. 100 of patients. This indicator is helpful in judging the effectiveness of the therapeutic modalities used. The direct cost of treating severe malaria was estimated to be 45963 CFA francs. This indicator will be useful in establishing controls to reduce costs. The mortality rate was 12.2 p. 100. Comparison of this rate with previous years suggests little improvement in the outcome of malaria management at the institution. This indicator must be taken into account in the ongoing quality control program. Overall these findings should enable institutional decision-making to improve management of severe malaria based on objective measurable data. PMID- 10701210 TI - [Marine life envenomations: example in New Caledonia]. AB - Marine life in the waters of New Caledonia is extraordinarily rich. However some of the animals inhabiting this wonderland are dangerous including a number of venomous species. A retrospective study conducted at the Territorial Hospital in Noumea for the three-year period between 1995 and 1998 showed that nearly 200 people/year were victims of envenomation by marine animals. Findings also indicated that the incidence of envenomation was rising as the practice of marine activities by the local population and tourists increased. Venomous species can be classified into 4 categories according to the mechanism of envenomation, i.e., biting animals such as sea snakes, cephalopoda, and eels; stinging animals including not only fish such as scorpion fish (Pterois, stonefish), sting-rays, saltwater catfish, surgeon fish, and flatfish but also cones and crown of thorns (Acanthaster planci); animals with contact venoms such as cnidaria (jellyfish, corals, sea anemones, and men-of-war), glaucus, sea cucumbers (holothurioidae), and sponges; and animals with more than one envenomation apparatus such as sea urchins and sea worms which can bite and sting. Study focused on the characteristics of each species including biology, envenomation apparatus, and chemical composition and action of the venom; pharmacological and clinical aspects of envenomation; and management and prevention of accidents. PMID- 10701211 TI - [Malaria in the Americas]. AB - In 1996, malaria involving Plasmodium vivax, Plasmodium falciparum, and, to a lesser extent, Plasmodium malariae was endemic in 21 countries in the Americas. The Amazon river basin and bordering areas including the Guyanas were the most affected zones. Until the mid 1970s, endemic malaria appeared to be under control. However in the ensuing 15 year period, the situation deteriorated drastically. Although trends varied depending on location, aggregate indexes indicated a twofold increase with recrudescence in previously settled areas and emergence in newly populated zones. Since 1990, the situation has worsened further in some areas where increased incidences have been associated with a high levels of drug-resistant Plasmodium falciparum. However this species remains in minority except in the Guyanas where the highest annual incidences (100 to 500 cases per 1000) and the most drug-resistant Plasmodium have been reported. The causes underlying this deterioration are numerous and complex. In regions naturally prone to transmission of the disease, outbreaks have been intensified by unrestrained settlement. The resulting deforestation has created new breeding areas for Anopheles darlingi, the main vector of malaria in the Americas. Migration of poor populations to newly opened farming and mining areas has created highly exposed areas for malaria infection. Implementation of adequate medical care and prevention measures has been hindered by a lack of money and sociopolitical unrest. Climatic phenomenon related the El Nino have also been favorable to the return of malaria to the region. Except with regard to financial resources and political unrest, the same risk factors for malaria are present in French Guiana. PMID- 10701212 TI - [Cholera in Guinea: the 1994-1995 epidemic]. AB - Since the first outbreak in 1970, cholera epidemics have occurred regularly in Guinea. Until 1994, epidemics recurred every 8 years and were confined to the capital and coastal areas. The first cases in every epidemic were observed in coastal lagoons near the Sierra Leone border. In 1994, the disease demonstrated its migratory ability as the first cases were reported in towns located in far eastern inland areas. Spread of the disease from war-torn Sierra Leone and Liberia where epidemics have also been reported cannot be ruled out. Control measures have gradually been implemented to deal with these outbreaks and a treatment facility was built at the University of Conakry in 1994. Bacteriological studies including antibiotic susceptibility tests carried out at this center showed that the offending bacteria was Vibrio cholerae El Tor of the Ogawa group. Although this strain is relatively sensitive to all antibiotics, analysis of epidemiological data revealed high mortality rates at the beginning of every outbreak probably due to delays in organizing appropriate care. A major effort is now being made to improve the response time particularly in remote inland areas. PMID- 10701213 TI - [Hepatitis C virus: first pathogen found in blood donors in Burundi]. PMID- 10701214 TI - [Treatment effectiveness of chromomycoses by surgery-thiabendazole in Madagascar (Fort Dauphin)]. PMID- 10701215 TI - [Is primary gallbladder cancer rare in Congo-Brazzaville?]. PMID- 10701216 TI - [A case of autochtonous anguilluliasis]. PMID- 10701217 TI - [A case of autochotonous anguilluliasis]. PMID- 10701218 TI - [The persistence of Wuchereria bancrofti in Ouvea Island (New Caledonia)]. PMID- 10701219 TI - The scientific environment of the Tuskegee Study of Syphilis, 1920-1960. PMID- 10701220 TI - Writing about their science: American interest in Soviet psychiatry during the post-Stalin Cold War. PMID- 10701221 TI - Dietl's crisis: the rise and fall of medical eponyms. PMID- 10701222 TI - Falling back on the brain. PMID- 10701223 TI - Low level exposure to ionizing radiation: do ecological and evolutionary considerations imply phantom risks? PMID- 10701225 TI - Mental-dental interface: window to the psyche and soma. PMID- 10701224 TI - When are children worth it? An evolutionary reading of Euripides' Medea. PMID- 10701226 TI - Medi?ation: meditation and medication in a personal tale of clinical depression. PMID- 10701227 TI - On dying with personhood: socratic death. PMID- 10701228 TI - The performance of a lifetime: a metaphor for the phenotype. PMID- 10701229 TI - The symbiotic nature of animal research. PMID- 10701230 TI - [Inpatient rehabilitation in inpatients with chronic obstructive lung diseases (COPD): effect on physical capacity for work, psychological wellbeing and quality of life]. AB - Success of inpatient rehabilitation for patients with chronic obstructive pulmonary disease (COPD) was assessed in a prospective study of 39 patients (mean age = 71 years). Six months after hospital stay (mean duration 22.4 days), physical endurance as assessed by the 6-minute walk distance remained unchanged. Subjective health measures improved (SF36 21 vs 29%; p < 0.05), MRC-dyspnoea was reduced (2.61 vs 2.19; p < 0.05) and measures of global quality of life also improved (56.1 vs 67.5; p < 0.05). Meanwhile, anxiety symptoms were reduced (7.7 vs 6.0; p < 0.05); no changes in depression scores were observed (6.8 vs 6.2; p = 0.271). A novel visual method to assess the burden of suffering, PRISM (Pictorial Representation of Illness and Self Measure), was applied for the first time in COPD patients. PRISM scores improved significantly (6.3 vs 12.2; p < 0.001). In conclusion, inpatient pulmonary rehabilitation improved subjective physical health measures and reduced anxiety levels. PMID- 10701231 TI - [Herpes simplex virus type 1 and 2 in Switzerland]. AB - A worldwide increase in the incidence of genital herpes infections has been described in recent years. Transmission of the herpes simplex virus type 2 (HSV 2) by asymptomatic seropositive subjects is considered to be a relevant mode of infection. The seroprevalence of HSV-2 varies considerably between different populations. In Europe, data are scarce and the epidemiological situation in Switzerland is unknown. In 1997 we performed serological examinations in 151 adult volunteers (87% between 20 and 49 years of age) of a low-risk population from the region of Basel with no history of genital herpes or any other sexually transmitted disease. The overall seroprevalence of HSV-1 was 77% and an annual seroconversion rate of 4.6% (95% CI: 3.8-5.6) was estimated for both sexes. Of the 51 subjects with no symptoms of orolabial herpes, 25 (49%) proved to be HSV-1 seropositive. In contrast, of 91 patients with symptoms of orolabial herpes, 90 (97%) had serum antibodies against HSV-1. The seroprevalence of HSV-2 was 14.6% for women (n = 89) and 8.1% for men (n = 62). The annual seroconversion rate was estimated to be 0.61% (95% CI: 0.14-1.4) for women and 0.49% (95% CI: 0.09-1.4) for men for the period after 1985 (when "safer sex" and the use of condoms were promoted). Our results indicate the significance of herpes simplex virus type 2 infections in Switzerland. More detailed studies are needed to describe the epidemiology of HSV-2 infections more reliably, especially in view of progress in the development of vaccines against HSV infections. PMID- 10701232 TI - Haemorrhagic shock and encephalopathy syndrome: report of two cases with special reference to hypoglycaemia. AB - Haemorrhagic shock and encephalopathy syndrome (HSES) is a devastating disorder affecting infants. So far no cases have been reported in Switzerland. It is characterised by the abrupt onset of hyperpyrexia, shock, encephalopathy, diarrhoea, disseminated intravascular coagulation (DIC) and renal and hepatic failure in previously healthy infants. Severe hypoglycaemia has been repeatedly reported in association with HSES. However, the pathophysiology of the hypoglycaemia is not clear. We report on two infants (2 and 7 months old) with typical HSES, both of whom were presented with nonketotic hypoglycaemia. In the first case, plasma insulin was 23 pmol/l at the time of hypoglycaemia (0.1 mmol/l). In the second case, increased values for interleukin-6 (IL-6) (319 pg/ml) and IL-8 (1382 pg/ml) were found 24 hours after admission, whereas IL-1 and tumour necrosis factor-alpha (TNF-alpha) were not measurable. Alpha-1 antitrypsin was decreased (0.6 g/l). In hyperpyrexic, unconscious and shocked infants, HSES should be considered and hypoglycaemia should be specifically looked for. Hypoglycaemia is not caused by hyperinsulinism but may be secondary to the release of cytokines. PMID- 10701233 TI - [Eosinophilic fasciitis (Shulman syndrome)]. AB - We report on a 35-year-old female with eosinophilic fasciitis (Shulman's syndrome). The characteristic features of this disease are scleroderma-like skin indurations, predominantly on the extremities, with joint contractures and intermittent blood eosinophilia. Histologic findings include fibrosis of muscle fascia and eosinophilic infiltration. Systemic corticosteroid therapy usually results in remission of symptoms. In this case refractory to systemic corticosteroids, we report for the fist time a successful therapy using cyclophosphamide. PMID- 10701234 TI - [Pseudohyponatremia]. PMID- 10701235 TI - Cancer nursing: the second century. PMID- 10701236 TI - The influence of technology on cancer nursing. AB - OBJECTIVES: To summarize technologies that have been described in nursing literature (in the context of oncology) over the last 25 years, and to present projections of possible technology in the future of oncology nursing. DATA SOURCES: CINAHL (1982 to present), MEDLINE (1976 to 1984), and author identification of articles. CONCLUSIONS: Technology has greatly influenced the evolution of oncology nursing, particularly concerning treatment (chemotherapy, radiation therapy, biotherapy, marrow and blood transplantation), access devices, and genetic and information technologies. IMPLICATIONS FOR NURSING PRACTICE: Nurses must be prepared to deal with how technology affects their philosophical perspective of nursing and the challenges presented by technology. PMID- 10701237 TI - Shaping advanced nursing practice in the new millennium. AB - OBJECTIVES: To provide a review of converging themes and trends that are shaping advanced practice nursing roles in oncology nursing. DATA SOURCES: Review and research articles, text-books, and organization documents. CONCLUSIONS: The current managed care environment provides many opportunities and challenges for oncology advanced practice nurses. Advanced practice nurses have both clinical and organization competencies that enable them to mediate the clinical needs of patients and organization goals within the health care system. IMPLICATIONS FOR NURSING PRACTICE: Advanced practice nurses can help shape their roles and practice by active participation in the development of systems to support access to clinical and financial information for effective decision making, collaboration among disciplines, and incorporating evidence-based care in their clinical practices. PMID- 10701238 TI - Oncology nursing education: peril and opportunities in the new century. AB - OBJECTIVES: To review the historic evolution of oncology nursing education, the health care trends affecting educational programs, and the educational innovations required for the future. DATA SOURCES: Articles, textbooks, and professional documents. CONCLUSIONS: Oncology nursing education has evolved into a premier model for specialty nursing education. Oncology nursing certification at both the generalist and advanced levels is well recognized. Current trends, however, suggest that some forms of oncology nursing education are on the decline and new and innovative educational approaches need to be considered for the future. IMPLICATIONS FOR NURSING PRACTICE: The 21st century will bring new challenges and opportunities for oncology nurse educators to develop new and innovative approaches to prepare oncology nurses to be effective and valued health care providers. PMID- 10701239 TI - Research and oncology nursing practice. AB - OBJECTIVE: To trace the evolution of oncology nursing research and to discuss its contribution to oncology nursing practice and health care. DATA SOURCES: Articles, textbooks, organizational documents, conference proceedings, and personal communication. CONCLUSIONS: Placing research in perspective will help suggest where we should go as a nursing specialty to provide excellent cancer care to patients, families, communities, and populations. It will also suggest directions for research priorities and for conceptual, methodologic, and health policy activities. IMPLICATIONS FOR NURSING PRACTICE: To influence oncology care, it is essential that oncology nurse researchers be responsive to trends that are relevant to both the conduct and utilization of oncology nursing research. PMID- 10701240 TI - Perspectives on cancer patient education. AB - OBJECTIVES: To provide an overview on cancer patient education. DATA SOURCES: Literature related to demographic and health care trends, patient education, health literacy, and cancer patient information needs. CONCLUSIONS: Cancer patients need information related to diagnosis, treatment, side effects, self care needs, and effects on work and relationships. Planned patient teaching includes a variety of teaching strategies and written materials, and contributes to better patient outcomes. Inadequate health literacy and other barriers present challenges to providing effective patient education. IMPLICATIONS FOR NURSING PRACTICE: More research is warranted in developing assessment tools and effective health education techniques and in measuring outcomes and costs related to patient education. The education needs of cancer patients will not drastically change in the next century. How nurses meet those needs, however, will change as we apply new learning theories and technologies of teaching. PMID- 10701241 TI - New opportunities for nurses as patient advocates. AB - OBJECTIVES: To define the role of nurses as patient advocates and to explore new strategies for the future. DATA SOURCES: Review articles, research studies, education and communications materials, and personal experience with oncology professionals, patients, and family members. CONCLUSIONS: Cancer nurses' roles in patient advocacy have progressed and grown as the profession of oncology nursing has itself matured. As resources continue to diminish, nurses need to consider the power of their roles as change agents, coordinators, and directors as well as interventionists. IMPLICATIONS FOR NURSING PRACTICE: There are many needs for research in this area as well as new roles for nurses who care for patients. Nurses need to be aware of ongoing research in areas such as health communications and consider partnering with persons in these other disciplines to enhance productivity and to use their time most efficiently. PMID- 10701242 TI - The National Cancer Program. AB - OBJECTIVES: To provide an understanding of the history, progress, and future of the National Cancer Program. DATA SOURCES: Published articles, reports, book chapters, and the National Cancer Institute (NCI) web site. CONCLUSIONS: The NCI is the largest agency for cancer research. The cancer incidence and burden remains significant in spite of many advances. Oncology nurses can contribute to the prevention and cure of cancer through an enhanced understanding of the NCI's program. IMPLICATIONS FOR NURSING PRACTICE: The NCI provides many opportunities for oncology nurses. Nurses can conduct NCI-sponsored research trials, serve on NCI advisory boards, and participate in clinical research. Nurses can advise patients and the public of the many resources available to patients from the NCI and assist patients with informed decision making. PMID- 10701244 TI - 3rd Brazilian Symposium on Basic Research of HIV/AIDS. Comandatuba, Brahia, Brazil. 16-20 October 1999. Abstracts. PMID- 10701243 TI - Health policy and legislation: impact on cancer nursing and care. AB - OBJECTIVES: To review policy and legislative initiatives in which nursing has played roles and to provide insight into trends and issues that characterize the policy and political agendas in which oncology nurses can play advocacy roles in the new millennium. DATA SOURCES: Professional journals, books, newspapers, news magazines, and internet web sites. CONCLUSIONS: Oncology nurses can offer solutions to the dilemmas posed by fiscal realities, unique American values and expectations, and the complexities of cancer care in the United State's market based health care system. Oncology nurses must assume roles as health policy specialists. IMPLICATIONS FOR NURSING PRACTICE: For nurses to gain acceptance in health policy arenas, individual nurses and groups of nurses must become expert in assessing the environment, the interpretation of cues, and the development and implementation of realistic strategies targeting priority health policy and legislative issues. PMID- 10701245 TI - [LIV Congress of The Brazilian Society of Cardiology. Abstracts]. PMID- 10701246 TI - Fabrication of metallic microstructures using exposed, developed silver halide based photographic film AB - This paper demonstrates that the pattern of silver particles embedded in the gelatin matrix of exposed and developed silver halide-based photographic film can serve as a template in a broadly applicable method for the microfabrication of metallic microstructures. In this method, a CAD file is reproduced in the photographic film by exposure and developing. The resulting pattern of discontinuous silver grains is augmented and made electrically continuous by electroless deposition of silver, and the electrically continuous structure is then used as the cathode for electrochemical deposition of an additional layer of the same or different metal. The overall process can be completed within 2 h, starting from a CAD file, and can generate electrically continuous structures with the smallest dimension in the plane of the film of approximately 30 microns. Structures with aspect ratio of up to 5 can also be obtained by using the metallic structures as photomasks in photolithography using SU-8 photoresist on the top of the electroplated pattern and exposed from the bottom, followed by development and electroplating through the patterned photoresist. This method of fabrication uses readily available equipment and makes it possible to develop prototypes of a wide variety of metallic structures and devices. The resulting structures--either supported on the film backing or freed from it--are appropriate for use as passive, structural materials such as wire frames or meshes and can also be used in microfluidic, microanalytical, and microelectromechanical systems. PMID- 10701247 TI - Atmospheric pressure matrix-assisted laser desorption/ionization mass spectrometry. AB - A novel ionization source for biological mass spectrometry is described that combines atmospheric pressure (AP) ionization and matrix-assisted laser desorption/ionization (MALDI). The transfer of the ions from the atmospheric pressure ionization region to the high vacuum is pneumatically assisted (PA) by a stream of nitrogen, hence the acronym PA-AP MALDI. PA-AP MALDI is readily interchangeable with electrospray ionization on an orthogonal acceleration time of-flight (oaTOF) mass spectrometer. Sample preparation is identical to that for conventional vacuum MALDI and uses the same matrix compounds, such as alpha-cyano 4-hydroxycinnamic acid. The performance of this ion source on the oaTOF mass spectrometer is compared with that of conventional vacuum MALDI-TOF for the analysis of peptides. PA-AP MALDI can detect low femtomole amounts of peptides in mixtures with good signal-to-noise ratio and with less discrimination for the detection of individual peptides in a protein digest. Peptide ions produced by this method generally exhibit no metastable fragmentation, whereas an oligosaccharide ionized by PA-AP MALDI shows several structurally diagnostic fragment ions. Total sample consumption is higher for PA-AP MALDI than for vacuum MALDI, as the transfer of ions into the vacuum system is relatively inefficient. This ionization method is able to produce protonated molecular ions for small proteins such as insulin, but these tend to form clusters with the matrix material. Limitations of the oaTOF mass spectrometer for singly charged high-mass ions make it difficult to evaluate the ionization of larger proteins. PMID- 10701248 TI - An investigation of the concentration dependence and response to analyte mixtures of carbon black/insulating organic polymer composite vapor detectors. AB - The responses relative to an air background of carbon black/polymer composite vapor detectors have been determined as a function of the concentration of a homologous series of alcohols (n-CnH2n+1OH, 1 < or = n < or = 8), a homologous series of alkanes (n-CnH2n+2, 5 < or = n < or = 10 and n = 12, 14), and a set of diverse solvent vapors. In all cases, the steady-state relative differential resistance responses, delta R/Rb, of the carbon black/polymer composite vapor detectors were well-described by a linear relationship with respect to the analyte partial pressure, at least over the tested concentration range (P/P degree = 0.005-0.03, where P degree is the vapor pressure of the analyte). When two vapors in air were simultaneously presented to the detectors, the delta R/Rb response, relative to an air background, was the sum of the delta R/Rb values obtained when each analyte was exposed separately to the carbon black/polymer composite detectors under study. Similarly, when an analyte was exposed to the detectors on top of a background level of another analyte, the delta R/Rb values of the array of detectors were very close to those obtained when the test analyte was exposed to the detectors only in the presence of background air. The initial training requirements from the array response output data of such detectors are minimized because the delta R/Rb response pattern produced by the analyte of concern can be associated uniquely with that odor, under the conditions explored in this work. PMID- 10701249 TI - Electrochemical impedance spectroscopy and X-ray photoelectron spectroscopy study of the response mechanism of the chalcogenide glass membrane iron(III) ion selective electrode in saline media. AB - The response mechanism of the iron(III) chalcogenide glass membrane ion-selective electrode (ISE) in saline media has been studied using electrochemical impedance spectroscopy (EIS) and X-ray photoelectron spectroscopy (XPS). EIS equivalent circuits and XPS surface compositions for the FeIII ISE are consistent with the presence of two surface films probably comprising a outer surface layer (OSL) and an Fe-deficient modified surface layer (MSL), along with a low-frequency charge transfer impedance that is attributable to the reduction of Fe3+. In accordance with literature data for the conductivity of low-bearing iron(III) chalcogenide glasses, a high-impedance MSL is internally consistent with XPS data for an Fe deficient MSL. It is evident that the impedance of the MSL diminishes on exposure to solutions containing Fe3+, and this finding is consistent with the ion exchange of Fe3+ within the MSL. Likewise, the charge-transfer impedance also decreases at elevated levels of Fe3+, demonstrating that Fe3+ is a participant in the reversible charge-transfer reaction occurring at the electrolyte/electrode interface. The kinetics of charge transfer are facilitated by Fe chelating agents (e.g., citrate, salicylate, EDTA, etc.) due presumably to the complexation of the products of the charge transfer process (possibly Fe2+). It is shown unequivocally that the response of the FeIII ISE in saline buffers is independent of pH, demonstrating that the ISE is responding directly to Fe3+, not H+. A mechanism involving a combination of charge transfer and ion exchange of FeIII, at the electrode diffusion layer, has been proposed to explain the 30 mV/decade slope of the FeIII ISE. PMID- 10701250 TI - Electrode coatings based on chitosan scaffolds. AB - Thin films of a biopolymer chitosan (CHIT) were cast on glassy carbon electrodes, modified by grafting Lucifer Yellow VS dye (LYVS) onto chitosan chains, and cross linked with glutaric dialdehyde (GDI). The ion-transport and ion-exchange properties of such polymeric structures (CHIT, CHIT-LYVS, CHIT-LYVS-GDI) were studied using cyclic voltammetry, rotating disk electrode, and flow injection analysis. The results showed that the chitosan matrix supported a fast ion transport as demonstrated by aqueous-like values of the apparent diffusion coefficients of Ru(NH3)6(3)+ and dopamine in the films. Anionic LYVS dye introduced a permselectivity against anions (e.g., Fe(CN)6(4)-, ascorbate) into the CHIT-LYVS films. The cross-linking of such films with GDI further increased their permselectivity as well as their stability. A unique combination of high permselectivity and fast ion transport in the CHIT-LYVS-GDI films is discussed in terms of the mixed-transport mechanism involving both pore and membrane diffusion in a highly hydrated chitosan matrix. The results indicate that the chemically modified chitosan is an attractive new coating for the development of fast, selective, and reversible sensors. PMID- 10701251 TI - Voltammetry as a tool for monitoring micellar structural evolution? AB - Self-assembled systems such as micelles and liquid crystals are currently of interest as templates for the controlled formation of nanoscale structures. Knowledge of the mesophase structure, structural evolution, and interparticle interaction is of great importance in understanding the behavior of such systems especially for applications such as nanoreactors. Here, we compare the use of cyclic voltammetry, chronoamperometry, and the rotating disk electrode (RDE) for the determination of micellar hydrodynamic radii and show that only the steady state RDE yields values directly comparable with nonelectrochemical techniques. The RDE is applied for the determination of cetyltrimethylammonium chloride micellar structure and observing micellar structural evolution as well as evaluating the usual intermicellar interactions. The results clearly show (a) the collapse of the micellar shear plane toward the hard-sphere surface with increasing electrolyte concentration, (b) the electrolyte-dependent spherical expansion of the micellar hard-spheres due to increasing aggregation (N) number, (c) the structural transition from spherical to rodlike micelles, and (d) micellar elongation. As well as structural evolution, the evolutionary changes in interaction processes are also observed, i.e. the transition from Coulombic interactions to excluded volume interaction. This paper describes in detail the voltammetric measurement of these processes and explicates the necessary experimental conditions for successful observation of micellar structural evolution. PMID- 10701252 TI - Temperature-independent near-infrared analysis of lysozyme aqueous solutions. AB - Digital Fourier filtering is used to produce a temperature-insensitive univariate calibration model for measuring lysozyme in aqueous solutions. Absorbance spectra over the 5000-4000 cm-1 spectral range are collected for lysozyme standards maintained at 14 degrees C. These spectra are used to compute the calibration model while a set of spectra collected at temperatures ranging from 4 to 24 degrees C are used to validate the accuracy of this model. The root-mean-square error of prediction (RMSEP) is 0.279 mg/mL over a tested lysozyme concentration range of 0.036-51.6 mg/mL. The detection limit is 0.68 mg/mL. In addition, multivariate calibration models based on partial least-squares regression (PLS) are evaluated and compared to the results from the univariate model. PLS outperforms the univariate model by providing a RMSEP of 0.090 mg/mL. Analysis of variance showed that both calibration methods effectively eliminate the adverse affects created by variations in solution temperature. PMID- 10701253 TI - Immunosensing platforms using spontaneously adsorbed antibody fragments on gold. AB - This paper describes the construction and characterization of miniaturized antigenic immunosurfaces composed of spontaneously adsorbed Fab'-SH fragments on gold. Rabbit Fab'-SH fragments contain a free sulfhydryl group that forms a thiolate bond with a gold substrate as detailed by X-ray photoelectron spectroscopy. This approach creates surfaces of higher epitope density, a factor critical to the early detection of disease, than surfaces composed of adsorbed whole molecule IgG on gold. The viability and specificity of antigenic Fab'-SH immunosurfaces is demonstrated using atomic force microscopy and confocal fluorescence microscopy, and possible explanations for the larger epitope density are discussed. PMID- 10701254 TI - Detection of secretion from single pancreatic beta-cells using extracellular fluorogenic reactions and confocal fluorescence microscopy. AB - Confocal microscopy with Zinquin, a fluorogenic Zn(2+)-specific indicator, was used for spatially and temporally resolved measurement of Zn2+ efflux from single pancreatic beta-cells. When cells were incubated in buffer containing Zinquin, application of insulin secretagogues evoked an increase in fluorescence around the surface of the cell, indicative of detection of Zn2+ efflux from the cell. The fluorescence increases corresponded spatially and temporally with measurements of exocytosis obtained simultaneously by amperometry. When images were taken at 266-ms intervals, the detection limit for Zn2+ was approximately 0.5 microM. With this image frequency, it was possible to observe bursts of fluorescence which were interpreted as fluctuations of Zn2+ level due to exocytosis. The average intensity of these fluorescence bursts corresponded to a Zn2+ concentration of approximately 7 microM. Since insulin is co-stored with Zn2+ in secretory vesicles, it was concluded that the Zn2+ efflux corresponded to exocytosis of insulin/Zn(2+)-containing granules from the beta-cell. Exocytosis sites identified by this technique were frequently localized to one portion of the cell, indicative of active areas of release. PMID- 10701255 TI - Mitigation of Rayleigh and Raman spectral interferences in multiway calibration of excitation-emission matrix fluorescence spectra. AB - A weighted parallel factor analysis (W-PARAFAC) model is applied to excitation emission matrix (EEM) fluorescence spectra of carbamate pesticides to aid with calibration in the presence of Raman scattering. Traditional PARAFAC inefficiently models the Raman scattering, resulting in prediction and calibration errors when a significant background is present. Four different weighting strategies were investigated and compared with subtraction of the appropriate sample background. Using a binary weighting strategy produced superior results, compared with a continuous distribution of weights. Further choice of weighting strategies, which are optimized to include either maximum analyte signal or to exclude a maximum amount of background scattering, is dependent on the degree of overlap and relative signal intensity. PMID- 10701256 TI - Development of an 19F NMR method for the analysis of fluorinated acids in environmental water samples. AB - This investigation was carried out to evaluate 19F NMR as an analytical tool for the measurement of trifluoroacetic acid (TFA) and other fluorinated acids in the aquatic environment. A method based upon strong anionic exchange (SAX) chromatography was also optimized for the concentration of the fluoro acids prior to NMR analysis. Extraction of the analyte from the SAX column was carried out directly in the NMR solvent in the presence of the strong organic base, DBU. The method allowed the analysis of the acid without any prior cleanup steps being involved. Optimal NMR sensitivity based upon T1 relaxation times was investigated for seven fluorinated compounds in four different NMR solvents. The use of the relaxation agent chromium acetylacetonate, Cr(acac)3, within these solvent systems was also evaluated. Results show that the optimal NMR solvent differs for each fluorinated analyte. Cr(acac)3 was shown to have pronounced effects on the limits of detection of the analyte. Generally, the optimal sensitivity condition appears to be methanol-d4/2M DBU in the presence of 4 mg/mL of Cr-(acac)3. The method was validated through spike and recovery for five fluoro acids from environmentally relevant waters. Results are presented for the analysis of TFA in Toronto rainwater, which ranged from < 16 to 850 ng/L. The NMR results were confirmed by GC-MS selected-ion monitoring of the fluoroanalide derivative. PMID- 10701257 TI - Development of an amino acid sequence and D/L-configuration determination method of peptide with a new fluorescence Edman reagent, 7-methylthio-4-(2,1,3 benzoxadiazolyl) isothiocyanate. AB - On the basis of the relationship between the fluorescence characteristics of the benzofurazan compounds and the Hammett constants (sigma p), a new fluorescence Edman reagent, 7-methylthio-4-(2,1,3-benzoxadiazolyl) isothiocyanate (MTBD-NCS) was designed and synthesized. MTBD-thiohydantoin (TH)-amino acid derivatives produced by the Edman sequencing method gave fluorescence, whereas other degradation byproducts such as MTBD-thiocarbamoyl (TC)- or carbamoyl (CA)-amino acids did not fluoresce. MTBD-NCS was applicable as an Edman sequencing reagent to the simultaneous determination of both the sequence and D/L-configuration of amino acids in peptides. Boron trifluoride (BF3) and HC1/methanol were adopted as the cyclization/cleavage and conversion reagents to suppress the amino acid residue racemization. The MTBD-TH-amino acids were separated on a reversed-phase column for amino acid sequencing, and their enantiomers were resolved on two types of polysaccharide-based chiral stationary phases for D/L-configuration determination. The method was successfully applied to the sequence and D/L configuration determination of D-amino acid-containing peptide [D-Ala2] deltorphin II. PMID- 10701258 TI - Light-scattering studies of packed stationary phases for capillary electrochromatography AB - Multiply scattered light through turbid media, packed particles, or compressed powders will inherently have a significantly longer optical path length than that of light which is not scattered. The concept of using the multiply scattered light potentially generated in the packed stationary phase of a capillary electrochromatography (CEC) column for enhanced detection as a result of its increased optical path length was examined. Ultraviolet (UV) light at 365 nm or laser light at 635 nm was focused to a small spot onto the packed section of a 3 microns spherisorb ODS1 CEC column (100 microns i.d.). The light was transported inside the capillary, and an image of the multiply scattered light several millimeters along the capillary was collected using a charged-couple device detector. Even if the spot size was less than 100 microns in diameter, evidence of light scattering was observed at a detection spatial off-set distance of 1-2 mm from the illumination point. When the calcium channel blocking drug felodipine was flushed through the column, the light intensity value dropped (increase in absorbance) to a greater degree at a spatial off-set (1.5 mm) than at the illumination point. The greater absorbance values at the spatial off-set were examined experimentally when felodipine was eluted from the column in the CEC mode in 6 min using MeCN/50 mM TRIS (pH 8.0) (80:20, v/v) at an applied voltage of 300 V/cm and an injection time of 2 s at 10 kV. A factor of 8.5 increase in absorbance was observed at a spatial off-set of 1 mm compared to the value obtained at the illumination point. An efficiency value of approximately 234,000 plates m-1 was obtained for this higher felodipine peak. Higher noise values, however, were also observed with this increase in absorbance. Using a spectrophotometer or an open capillary to obtain reference values for optical length, it was possible to estimate the average optical path length of light traveled through the packed stationary phase when transmitted at a spatial off set. It was concluded that, although an increase in absorbance of 8.5 was observed at a spatial off-set, this most likely arises from the light being "redirected" and scattered in a straightforward fashion along the capillary. It was expected that if substantial multiple scattering did occur inside the packed stationary phase, a significantly larger absorbance increase would be attained. A number of proposals are thus given to explain the relatively low degree of multiple scattering in this stationary phase and suggestions offered on means to attain even higher absorbance increases at a spatial off-set. Additional potential applications are also discussed. PMID- 10701259 TI - Analysis of human skin emanations by gas chromatography/mass spectrometry. 2. Identification of volatile compounds that are candidate attractants for the yellow fever mosquito (Aedes aegypti). AB - Volatile compounds emanated from human skin were studied by gas chromatography/mass spectrometry (GC/MS). The purpose of this study was to identify compounds that may be human-produced kairomones which are used for host location by the mosquito, Aedes aegypti (L.). The procedure used to collect volatiles was chosen because of prior knowledge that attractive substances can be transferred from skin to glass by handling. Laboratory bioassays have shown that the residuum on the glass remains attractive to mosquitoes until the compounds of importance evaporate. The sampling and analytical procedures modeled the above cited process as closely as possible except that the evaporation of compounds from the glass surface was accomplished by thermal desorption from glass beads in a heated GC injection port. This made possible the solventless injection of volatiles onto the column. The compounds were cryofocused on the head of the column with liquid nitrogen prior to GC separation. A single stage of mass spectrometry on a triple quadrupole instrument was used for mass analysis. A combination of electron ionization and pulsed positive ion/negative ion chemical ionization modes on two different GC columns (one polar, one relatively nonpolar) was used to identify most of the 346 compound peaks detected by this technique. PMID- 10701260 TI - Automated identification of amino acid sequence variations in proteins by HPLC/microspray tandem mass spectrometry. AB - Amino acid sequence variations resulting from single-nucleotide polymorphisms (SNPs) were identified using a novel mass spectrometric method. This method obtains 99+% protein sequence coverage for human hemoglobin in a single LC microspray tandem mass spectrometry (microLC-MS/MS) experiment. Tandem mass spectrometry data was analyzed using a modified version of the computer program SEQUEST to identify the sequence variations. Conditions of sample preparation, chromatographic separation, and data collection were optimized to correctly identify amino acid changes in six variants of human hemoglobin (Hb C, Hb E, Hb D Los Angeles, Hb G-Philadelphia, Hb Hope, and Hb S). Hemoglobin proteins were isolated and purified, dehemed, (S)-carboxyami-domethylated, and then subjected to a combination proteolytic digestion to obtain a complex peptide mixture with multiple overlaps in sequence. Reversed-phase chromatographic separation of peptides was achieved on-line with MS utilizing a robust fritless microelectrospray interface. Tandem mass spectrometry was performed on an ion trap mass spectrometer using automated data-dependent MS/MS procedures. Tandem mass spectra were collected from the five most abundant ions in each scan using dynamic and isotopic exclusion to minimize redundancy. The spectra were analyzed by a version of the SEQUEST algorithm modified to identify amino acid substations resulting from SNPs. PMID- 10701262 TI - Laser-induced electron capture mass spectrometry AB - Two techniques are reported for detection of electrophorederivatized compounds by laser-induced electron capture time-of-flight mass spectrometry (LI-EC-TOF-MS). In both cases, a nitrogen laser is used to induce the electron capture. The analyte is deposited in a matrix consisting of a compound with a low ionization potential such as benzo[ghi]perylene in the first technique, where the electron for electron capture apparently comes from this matrix. In the second technique, the analyte is deposited on a silver surface in the absence of matrix. It seems that "monoenergetic" ions instantly desorb from the target surface in the latter case, since the peak width in the continuous extraction mode essentially matches the pulse width of the laser (4 ns). Ten picomoles of 3-O-(pentafluorobenzyl) alpha-estradiol were detected at a S/N > or = 50, where the spot size of the laser was approximately 0.25% of the sample spot. It is attractive that simple conditions can enable sensitive detection of electrophores on routine TOF-MS equipment. The technique can be anticipated to broaden the range of analytes in both polarity and size that can be detected by EC-MS relative to the range for GC/EC-MS. PMID- 10701261 TI - An LC-MS-MS method for the comprehensive analysis of cocaine and cocaine metabolites in meconium. AB - A sensitive, precise, and accurate liquid chromatography-mass spectrometry (LC-MS MS) method was developed to quantitate cocaine and cocaine metabolites, which were simultaneously extracted from suspected drug-positive meconium samples using solid-phase extraction. The ability to analyze cocaine and multiple cocaine metabolites in meconium makes this method a powerful tool for the study of cocaine exposure and metabolism in neonates. Of 22 samples, only 1 did not show the presence of cocaine or any metabolite of cocaine. The identified metabolites varied both qualitatively and quantitatively between samples. Ecgonine appears to hold the most promise as a diagnostic marker compound for neonatal cocaine exposure as this metabolite was present in 21 of 21 of the positive samples tested, and at a relatively high median concentration. However, a core group of eight metabolites (present in at least 20 of 21 positive samples) was identified that appears to possess the greatest utility for determining cocaine exposure. Finally, the use of this method for assessment of the magnitude of fetal cocaine exposure was demonstrated. PMID- 10701263 TI - Tuning of an electrospray ionization source for maximum peptide-ion transmission into a mass spectrometer. AB - We describe assembly and optimization of a continuous flow nanoelectrospray source for high-performance analysis on a routine basis. It is derived from an inJection adaptable Fine Ionization Source ("JaFIS"), previously shown to be durable and easy to use (Geromanos, S.; et al. Rapid Commun. Mass Spectrom. 1998, 12, 551-556) and now modified for maximum sensitivity. Proper design, manufacturing, and quality control of spray needles with specific orifice diameters, in combination with precisely controlled helium backpressure and applied voltage, enable stable flows at 1-2 nL/min. Needle positioning and ion spray potential are hereby exceedingly important, as shifts by 0.5 mm or 25 V, respectively, cause significant reduction in signal strength. In addition to prolonged analysis times, ultralow flows also yield higher sensitivity, the result of an improved "overall ion transfer efficiency" measured to be approximately 5% at 1.6 nL/min. Used in combination with a "microtip" (Erdjument Bromage, H.; et al. J. Chromatogr. A 1998, 826, 167-181), the optimized JaFIS implements infusion-style ESI-MS at sensitivities approaching capillary LC-MS. Spraying times in excess of 20 h allow for any number of tandem mass spectrometric analysis routines to be performed, and to average thousands of scans in every experiment, thereby further improving sensitivity. This was fully illustrated by extensive analysis of a 2-fmol peptide mixture, in a 2-microL volume, using a multimode MS approach. PMID- 10701264 TI - Collision-induced dissociation of ions within the orifice-skimmer region of an electrospray mass spectrometer AB - An equation was derived to describe the variation of the gas number density within the region between the orifice and the skimmer of an electrospray ionization mass spectrometer. The equation was used to develop a semi quantitative model to predict the value of orifice voltages that lead to ion fragmentation within this region. This model made it possible to predict the types of solvent adducts observed for analytes at various orifice voltages. In addition, it is shown that a small number of high-energy collisions is equally effective for collision-induced dissociation as compared to a large number of low energy collisions. Finally, this model is tested with different background electrolyte solutions and a different electrospray mass spectrometer. It is demonstrated that controlled fragmentation studies can be performed on single quadrupole mass spectrometers, and the proposed model gives a reasonable description of the fragmentation process in both spectrometers. PMID- 10701265 TI - Quantification and rapid metabolite identification in drug discovery using API time-of-flight LC/MS. AB - Liquid chromatography/mass spectrometry (LC/MS), utilizing a time-of-flight (TOF) mass analyzer, has been evaluated and applied to problems in bioanalysis for pharmacokinetics and drug metabolism. The data obtained by TOF MS differ from those obtained using quadrupole mass spectrometer instruments in that full-scan spectra can be routinely collected with greater sensitivity and speed. Both quantitative and qualitative information, including compound concentration in rat plasma and full-scan atmospheric pressure ionization mass spectra, are concurrently obtained. This approach has been used to characterize the disposition of several drug compounds that have been simultaneously dosed to rats in a cassette format. Quantitation limits in the 5-25 ng/mL range (approximately 20 nM) were obtained from nominal mass chromatograms (0.5 Da resolution). A reference lock mass was used to provide accurate mass measurement to reach third decimal place accuracy in the monoisotopic molecular weight. An improvement in quantitation limits was demonstrated after using accurate mass determinations. Several possible preliminary drug metabolites were confirmed or refuted, based on accurate mass. The trend of metabolite formation and clearance was qualitatively evaluated. PMID- 10701267 TI - Automated instrumentation for comprehensive isotachophoresis-capillary zone electrophoresis AB - An automated comprehensive isotachophoresis-capillary zone electrophoresis (ITP CZE) system is described. The sample is focused in the first capillary by ITP and injected repeatedly into the second smaller diameter capillary for more rapid CZE separation. Since only small portions of the concentrated zones are sequentially injected for CZE separation, overloading was not observed. Moreover, the sensitivity is enhanced because all of the concentrated zones are analyzed and the results are summed. A single detector (only for the CZE dimension) is required, and accurate timing for CZE injection is not necessary. The system was evaluated using a mixture of angiotensins. The effect of addition of leading electrolyte at the junction of the ITP and CZE capillaries before each CZE run on comprehensive ITP-CZE peak area was studied, and leading electrolyte volumes between 7 and 11 microL led to the best sensitivity. Under optimized conditions, a detection limit of approximately 5 nM could be achieved by injecting 10 microL of angiotensin solution. PMID- 10701266 TI - Selective detection of alkanolamine vapors by ion mobility spectrometry with ketone reagent gases. AB - The ion mobility (IMS) spectra of the alkanolamines, monoethanolamine (MEA), 3 amino-1-propanol (PRA), 4-amino-1-butanol (BUA), and 5-amino-1-pentanol (PEA) with acetone and 4-heptanone reagent gases have been measured using a hand-held spectrometer. Monomer and dimer peak patterns were observed for all the alkanolamines with acetone reagent gas. Drift times of monomer and dimer ion clusters for each alkanolamine increased linearly in order of size of alkyl group. Ammonia, Freon 22, and F76 diesel vapors, having similar or coincident mobilities, caused severe interference. Replacement of acetone with 4-heptanone reagent gas resulted in good separation by the altering drift times of product ions. The limit of detection was 0.005 ppm having a linear range of 0.005-0.7 ppm, and signal saturation occurred above 0.88 ppm. Detection was reversible, with a response time of 4 min and a slower recovery time of > 60 min, at vapor levels of 0.7 ppm and ambient nozzle and drift-region temperatures. In contrast to acetone chemistry, single-peak patterns were observed for the alkanolamines with the 4-heptanone reagent. Further, drift times unexpectedly remained stagnant with increasing alkyl-group size. From atmospheric pressure chemical ionization (APcI) tandem mass spectral identifications and collision induced studies, dynamic changes in product-ion equilibria in the IMS drift region compensated by differences in collision cross sections were suggested as the governing causes of the unusual mobility effect. PMID- 10701268 TI - Capillary electrophoresis of supercoiled DNA molecules: parameters governing the resolution of topoisomers and their separation from open forms. AB - We describe the separation of covalently closed and open circular DNA forms with capillary electrophoresis. This technique is expected to be applied in the research of novel anticancer molecules targeting the activity of topoisomerase I. The separation of a plasmid mixture containing fully supercoiled molecules, single topoisomers, and their relaxed and open circular forms was tested in an electric field of 200 V/cm using Tris/borate buffer with the addition of magnesium ions at low concentrations and various sieving polymers. The resulting separation is quite simple to achieve and is clearly comparable to that obtained in agarose gels run at low voltage, but with an improved resolution, a higher quantitativity, and a higher speed of analysis. We identified three main parameters that influence the separation: (I) Low concentrations of MgCl2 in the separation buffer are required for a good resolution of topoisomers. (II) Cellulose derivatives can be used as sieving polymers; in our hands, HPMC and HEC worked best. (III) High molecular mass forms of sieving polymers allow the best separations. PMID- 10701269 TI - Separation of nontarget compounds by DNA aptamers. AB - The ability of DNA aptamers to separate nontarget compounds is demonstrated. Two G-quarter forming aptamers, a 15-mer and a 20-mer, were covalently linked to fused silica capillary columns to serve as stationary-phase reagents in capillary electrochromatography. Separations of binary mixtures of amino acids (D-trp and D tyr), enantiomers (D-trp and L-trp), and polycyclic aromatic hydrocarbons were achieved. Aptamers offer several attractive features for stationary-phase reagents, including ease of synthesis and of attachment to surfaces and modification of their binding properties through minor changes in sequence. PMID- 10701270 TI - Cell separation by dielectrophoretic field-flow-fractionation. AB - Dielectrophoretic field-flow-fractionation (DEP-FFF) was applied to several clinically relevant cell separation problems, including the purging of human breast cancer cells from normal T-lymphocytes and from CD34+ hematopoietic stem cells, the separation of the major leukocyte subpopulations, and the enrichment of leukocytes from blood. Cell separations were achieved in a thin chamber equipped with a microfabricated, interdigitated electrode array on its bottom wall that was energized with AC electric signals. Cells were levitated by the balance between DEP and sedimentation forces to different equilibrium heights and were transported at differing velocities and thereby separated when a velocity profile was established in the chamber. This bulk-separation technique adds cell intrinsic dielectric properties to the catalog of physical characteristics that can be applied to cell discrimination. The separation process and performance can be controlled through electronic means. Cell labeling is unnecessary, and separated cells may be cultured and further analyzed. It can be scaled up for routine laboratory cell separation or implemented on a miniaturized scale. PMID- 10701271 TI - Simultaneous quantification of acetanilide herbicides and their oxanilic and sulfonic acid metabolites in natural waters. AB - This paper describes a procedure for simultaneous enrichment, separation, and quantification of acetanilide herbicides and their major ionic oxanilic acid (OXA) and ethanesulfonic acid (ESA) metabolites in groundwater and surface water using Carbopack B as a solid-phase extraction (SPE) material. The analytes adsorbed on Carbopack B were eluted selectively from the solid phase in three fractions containing the parent compounds (PCs), their OXA metabolites, and their ESA metabolites, respectively. The complete separation of the three compound classes allowed the analysis of the neutral PCs (acetochlor, alachlor, and metolachlor) and their methylated OXA metabolites by gas chromatography/mass spectrometry. The ESA compounds were analyzed by high-performance liquid chromatography with UV detection. The use of Carbopack B resulted in good recoveries of the polar metabolites even from large sample volumes (1 L). Absolute recoveries from spiked surface and groundwater samples ranged between 76 and 100% for the PCs, between 41 and 91% for the OXAs, and between 47 and 96% for the ESAs. The maximum standard deviation of the absolute recoveries was 12%. The method detection limits are between 1 and 8 ng/L for the PCs, between 1 and 7 ng/L for the OXAs, and between 10 and 90 ng/L for the ESAs. PMID- 10701272 TI - Pesticide residue analysis by off-line SPE and on-line reversed-phase LC-GC using the through-oven-transfer adsorption/desorption interface. AB - A new method to determine pesticide residue in water is presented. The described method includes using off-line solid-phase extraction (SPE) and on-line reversed phase liquid chromatography-gas chromatography (RPLC-GC). An interface, based on a modified programmed temperature vaporizer (PTV) injector, packed with a suitable trapping material, is used for on-line RPLC-GC. The changes made in the PTV injector affect the pneumatic system, sample introduction, and solvent elimination. The new interface is easily capable of automation. Methanol/wate (70/30) is used as the eluent in the LC preseparation step. The LC column flow during elution is different from the flow during the transfer step. The transferred volumes range from 500 to 1400 microL (volume of the fractions of interest). Solvent elimination is almost 100% before the sample reaches the GC column. The described system does not show any variation of the peak retention times. The detection limit for real samples ranges from 0.04 to 1.5 ng/L, using NP detection. PMID- 10701273 TI - Mechanism of DNA hydrodynamic separation in chromatography. AB - An alternative chromatographic procedure for the separation of large double stranded DNA molecules was discovered recently and called "slalom chromatography". This fractionation is based on a new hydrodynamic process that is determined by the progression of the mobile-phase flow through the interstitial spaces created between the highly packed particles inside the column. Here, the separation is treated as the result of a slowing down of the large double-stranded DNA fragments in relation to their size with the flow direction changing around the particles. A model, based on the concept derived from the reorientation time of macromolecules, was adequate to describe the hydrodynamic phenomenon. This model constitutes an attractive tool to enhance the expansion of this chromatographic procedure and provide valuable information on the dynamic behavior of biological polymers. PMID- 10701274 TI - DNA microsatellite analysis using ion-pair reversed-phase high-performance liquid chromatography. AB - Genotyping based on short tandem repeat (STR) regions is used in human identification and parentage testing, gene mapping studies, cancer diagnostics, and diagnosis of hereditary diseases. Analysis of STR systems using slab gel electrophoresis requires lengthy and labor-intensive procedures. Therefore, alternative methods such as capillary electrophoresis or ion-pair reversed-phase high-performance liquid chromatography (IPRP HPLC) have been used to analyze DNA. IPRP HPLC offers an attractive substitute to gel electrophoresis for STR analysis because of the reduced analysis time, and there is no need for the waste disposal associated with radioisotopic, enzyme-linked, or fluorescence detection systems. We evaluated the use of IPRP HPLC for the sizing and typing of STR alleles from the HUMTHO1 locus. The IPRP HPLC conditions (column temperature, flow rate, percent organic modifier per minute) were optimized for the separation of PCR products. Using the optimized separation conditions, the alleles of the HUMTHO1 system were sized in their native state (double standard) with the use of internal markers. The typing results correlated 100% to accepted methods of DNA typing. The analysis time for the HUMTHO1 locus was less than 14 min, and the alleles could be peak captured for further examination following such as sequencing. PMID- 10701276 TI - Instrumentation for chemical cytometry. AB - Capillary electrophoresis is ideally suited to chemical analysis of individual cells. Small mammalian somatic cells (approximately 15 microns in diameter) can be analyzed by injecting the intact cell into a capillary, lysing the cell, separating and detecting the cellular components, and reconditioning the capillary prior to the next injection. In this paper, we report on technical improvements to single-cell analysis. We designed an inexpensive multipurpose single-cell injector that facilitates the following: (i) monitoring of injection, (ii) reproducible pressure- or electrokinetic-driven injection of the cell, (iii) complete cell lysis by SDS within 30 s of injection, and (iv) pressure-driven capillary reconditioning. Furthermore, we report on the analysis of glycosylation and glycolysis in single human carcinoma cells (HT29 cell line). The reliability and quality of the analysis is confirmed by comparing electropherograms from single cells and those from purified cell extracts. PMID- 10701275 TI - Trace-level amino acid analysis by capillary liquid chromatography and application to in vivo microdialysis sampling with 10-s temporal resolution. AB - A sensitive method was developed to determine 16 amino acids, including all the neurotransmitter amino acids and neuromodulators, in physiological samples. Samples were derivatized with o-phthalaldehyde/tert-butyl thiol followed by two scavenging reactions that reduced the chemical background caused by excess derivatization reagent by approximately 90%. A total of 250 nL of the derivatized sample was injected and concentrated onto a 50-micron-inner diameter capillary column packed with 5-micron reversed-phase particles and separated using gradient elution. Analytes were detected amperometrically at a cylindrical 9-micron carbon fiber microelectrode. The combination of on-column concentration, scavenging reactions after derivatization, high sensitivity electrochemical detection, and protocols to minimize amine contamination allowed detection limits of 90-350 pM (20-80 amol) for all the amino acids tested. This method was used to analyze in vivo microdialysate samples from probes implanted in the striatum of anesthetized rats. Probes were perfused at 1.2 microL/min and fractions collected every 10 s. The 200-nL fractions were diluted to 2 microL to facilitate sample handling for off-line analysis. The suitability of this method for simultaneous monitoring of all the major amino acid neurotransmitters with 10-s temporal resolution under basal conditions, during potassium stimulation, and during selective uptake inhibition of gamma-aminobutyric acid is demonstrated. PMID- 10701277 TI - Development of a hollow waveguide sampler for detection of chlorinated aromatic compounds in soils. AB - In this paper, a Fourier transform infrared (FT-IR) spectroscopic method for detection of chlorinated aromatic compounds in soils was developed. The sensing device of this method was based on an infrared hollow waveguide, the inner surface of which was coated with a hydrophobic film. Vaporized chlorinated aromatic compounds from soils were trapped onto the hydrophobic film of the hollow waveguide sampler following detection by FT-IR spectrometry. The extraction process in this method was similar to the headspace solid-phase microextraction (HSSPME) in principle. Means of increasing the speed of transfer of the vaporized organic species to the sampler were also studied. Results indicated that, with a negative pressure on the end of the sampler, the speed of transfer increased significantly. Vapor pressures of the analytes were used as an indication to test the limitation of this method in the analysis of organic compounds in soils. Results showed that analytes with vapor pressures lower than 12 Torr could be detected quantitatively. The typical R-square of the regression on the concentration and IR signals was around 0.99 and the typical detection limits were in the range of hundreds of parts per billion. PMID- 10701278 TI - The effect of position and mattress on interface pressure. AB - The aim of this investigation was to determine which positions resulted in the lowest pressures to the skin of persons lying in bed. Pressures were recorded in 10 different lying positions on 2 mattresses in 62 healthy volunteers. The study revealed that the 30 degrees semi-Fowler position and the prone position resulted in the lowest interface pressures. The 30 degrees laterally inclined position had lower pressure readings than the 90 degrees side lying position; 90 degrees side lying position gives the highest pressure readings and thus should be avoided. A Tempur polyethylene-urethane mattress reduces interface pressures by 20 to 30% in comparison to a standard hospital mattress (12-cm-thick cold foam). PMID- 10701279 TI - Informational needs of surgical patients following discharge. AB - A decreased length of hospital stay for surgical patients dramatically reduces the time available for nurses to teach patients how to manage postdischarge self care. Nurses need guidelines to prioritize teaching content. This study explored the perceptions of information needed by 45 patients who were recently discharged following short-term surgical procedures. Patients identified information related to activity, wound care, complications, and pain management as highly important. Patient reports of information given and satisfaction with information were also examined. These findings can contribute to the development of teaching programs for patients who are discharged following short-term surgical procedures. PMID- 10701280 TI - A professional-patient partnership model of discharge planning with elders hospitalized with heart failure. AB - Despite efforts to improve the discharge planning process and subsequent outcomes, existing mechanisms fail to accurately identify elders' needs for follow-up care. Studies report rehospitalization rates ranging from 12 to 50%. The two aims of this study were to (1) examine the difference in outcomes for elders hospitalized with heart failure and caregivers who participated in a professional-patient partnership model of discharge planning compared to those who received the usual discharge planning and (2) examine differences in costs associated with hospital readmission and use of the emergency room following hospital discharge. A before-and-after nonequivalent control group design was used for this study. Data were collected from the control and the intervention cohorts before discharge and at 2 weeks and 2 months postdischarge. One hundred and fifty-eight patient-caregiver dyads completed both the predischarge and 2 weeks postdischarge interviews; 140 also completed a 2-month follow up. The average age of elders was 73.7 years; the average age of the caregivers was 58.5 years. The findings indicated that elders in the intervention cohort felt more prepared to manage care, reported more continuity of information about care management and services, felt they were in better health, and when readmitted spent fewer days in the hospital than the control cohort. Caregivers in the intervention cohort also reported receiving more information about care management and having a more positive reaction to caregiving 2 weeks postdischarge than the control cohort. PMID- 10701281 TI - Nursing documentation versus standardized assessment of cognitive status in hospitalized medical patients. AB - Although the literature discusses the importance of assessing cognitive status, little research has explored the concordance of nurses' documentation of cognitive status and standardized assessment. This study examined nurses documentation of cognitive status in 42 medically hospitalized individuals (mean age 51.9, SD = 10.1 years) using a variety of standardized measures. Although the chart review revealed no documentation of impaired cognitive status, impaired performance in 24 to 67% of the cognitive measures was identified. This study suggests nurses are missing cognitive impairment in hospitalized patients by limiting assessment to orientation. Use of a combination of several brief screening measures, such as the Clock Drawing Test and the standardized Mini Mental State Examination, would provide timely, effective, and inexpensive assessment of cognitive status. PMID- 10701282 TI - Acute confusion assessment instruments: clinical versus research usability. AB - Acute confusion (AC), also referred to as delirium (AC/delirium), is a common problem seen by health professionals who work in a variety of care settings. This is an evaluative report on the clinical usability of instruments to assess AC/delirium as a part of nursing practice. Specifically, five instruments [the Confusion Assessment Method (CAM), Delirium Rating Scale (DRS), Delirium Symptom Inventory (DSI), Mini-Mental State Examination (MMSE), and Neelon/Champagne (NEECHAM) Confusion Scale] are discussed. The work demonstrates how the cooperation of nurses in practice, education, and research can improve both patient and staff outcomes. PMID- 10701283 TI - Nurses' job satisfaction and organizational climate in a dynamic work environment. AB - The infrastructure and organization of hospitals are changing rapidly as a result of major transitions in health care. Downsizing in hospitals has caused employees to have to take on new tasks and, often, multiple tasks with a decrease in available resources and an increase in job complexity. Naturally, such organizational changes have a profound effect on the nature and duration of patient hospitalization and on the job responsibilities and roles of inpatient staff. In many hospitals, there is a perception of chaos, sometimes resulting in frustration among the nursing personnel. The purpose of this study was to describe the relationship between nurses' job satisfaction and organizational climate. PMID- 10701284 TI - Confusion and aggression in restrained elderly persons undergoing hip repair surgery. AB - The purpose of this pilot study was to observe and describe the behaviors of confusion and aggression in physically restrained elderly hospitalized persons following hip repair surgery. Five elderly persons who were 75-95 years of age and physically restrained following hip repair surgery comprised the sample of this descriptive study. Descriptive data including age, sex, medical diagnosis, current medications, serum electrolyte and arterial blood gas results, date/time of hip fracture, subsequent surgical repair, and restraint application were obtained from the medical record. Observations of behaviors associated with confusion and aggression were conducted at six separate observation times lasting approximately 20 to 30 minutes each. The observation periods were divided into two mornings, two afternoons, and two evening sessions. These behaviors were assessed by utilizing a combined observational tool developed by the researcher, and was created from the Clinical Assessment of Confusion--A, by Vermeersch, and Ryden Aggression Scale. After data analysis, three patterns of confusion emerged. These were the major pattern of confusion, the minor pattern of confusion, and pattern of confusion relative to time. All patients experienced low serum sodium levels immediately prior to the application of physical restraints. Based on the results of the pilot study, further investigation is needed. PMID- 10701285 TI - Application of Cohen and Garrison's respirator cartridge service life prediction model to 1,6-hexamethylene diisocyanate (HDI) monomer. PMID- 10701286 TI - Control of fire hazards in commercial drycleaning shops using petroleum-based solvents. National Institute for Occupational Safety and Health. PMID- 10701287 TI - Overexposure to methylene bisphenyl isocyanate (MDI) in a motor vehicle parts manufacturing facility. PMID- 10701288 TI - The use of participatory action research and ergonomics in the prevention of work related musculoskeletal disorders in the newspaper industry. AB - The newspaper industry is one of many in which employees are reported to be at risk for work-related musculoskeletal disorders of the upper extremities and low back. The purpose of this 18-month demonstration project was to assess the usefulness of a participatory ergonomics process as a strategy to reduce the risk factors associated with musculoskeletal disorders at a metropolitan newspaper company. The company involved had 455 employees and a daily circulation of 75,200. Employees from both office and production areas participated. The participatory action research approach utilized required investigators to work collaboratively with the study population. Using a five-step continuous improvement process, the ergonomics committee identified and evaluated jobs having ergonomic risk factors. This was followed by the development, implementation, and evaluations of interventions aimed at reducing risk factor exposure. The committee's productivity and participant feedback were used as measures of the committee's effectiveness. During the project period, interventions were implemented in 11 of 12 targeted departments. Participant ratings of effectiveness for different aspects of the ergonomics process were generally favorable. The mean and median cost for ergonomic interventions were $376 and $25, respectively. This project demonstrated that participatory action research could be used to develop and implement ergonomic solutions that reduce the risk factors associated with work-related musculoskeletal disorders. PMID- 10701289 TI - Personal sampling in parallel with open-face filter cassettes and IOM samplers for inhalable dust--implications for occupational exposure limits. AB - Parallel personal sampling was carried out with the open-face filter cassette and the IOM sampler for inhalable dust for nine types of organic dust. Parallel samples numbering 749 were obtained from 152 plants. Extremely large values and outliers were disregarded, and the remaining data for each type of dust were divided into subsets according to type of product or work task, and analyzed with the aid of linear regression. The coefficient of regression for each subset ranged between 0.2 and 0.7. Hypothetical occupational exposure limits (OELs) for inhalable dust were calculated based on the linear relation obtained between the dust concentrations measured with the open-face filter cassette and the IOM sampler. The fraction of person days with time-weighted average (TWA) concentrations exceeding the calculated hypothetical OELs for inhalable dust was obtained from the distribution of measured TWA inhalable dust concentrations. Based on the results of this study and the difference in sampling efficiency for large particles between the two samplers, it was concluded that the numerical value of the OEL for inhalable dust may be set at approximately twice the numerical value of the corresponding limit value for "total dust." Additional consideration of recently discovered health effects, and technical and economical factors may result in other numerical values of future OELs for inhalable dust. PMID- 10701290 TI - Comparison of the constituents of two jet engine lubricating oils and their volatile pyrolytic degradation products. AB - Leaking oil seals in jet engines, at locations prior to the compressor stage, can be a cause of smoke in the cabins of BAe-146 aircraft. Compressed combustion air is bled off to pressurize the cabin and to provide a source of fresh air. Bleed air is diverted from a location just prior to the combustion chamber at a temperature around 500 degrees C. To prevent oil breakdown products from entering the cabin air, catalytic converters have been used to clean the air. During an oil seal failure this device becomes overloaded and smoke is observed in the cabin. Some aircraft companies have removed the catalytic converters and claim an improvement in air quality. During an oil seal failure, however, the flight crew is potentially exposed to the thermal breakdown products of the engine oils. Because very little is known regarding the thermal breakdown products of jet engine lubrication oils, two commercially available oils were investigated under laboratory conditions at 525 degrees C to measure the release of CO, CO2,NO2, and HCN as well as volatiles which were analyzed using GC-Mass spectrometry in an attempt to see if the neurotoxic agents tricresyl phosphates (TCPs) and trimethyl propane phosphate (TMPP) would be present or formed. TMPP was not found in these experiments. Some CO2 was generated along with CO which reached levels in excess of 100 ppm. HCN and NO2 were not detected. GC compositions of the two bulk oils and their breakdown products were almost identical. The presence of TCPs was confirmed in the bulk oils and in the volatiles. Localized condensation in the ventilation ducts and filters in the air conditioning packs are likely the reason why the presence of TCPs has not been demonstrated in cabin air. It was recommended that this needed to be verified in aircraft. PMID- 10701291 TI - Improved estimation of dermal pesticide dose to agricultural workers upon reentry. AB - Agricultural workers reentering fields after pesticide application to engage in hand labor activities are subject to potentially significant dermal exposures to residues on foliage and in soil. Environmental Protection Agency (EPA) guidelines for assessment of post-application exposures were originally described in the 1984 Pesticide Assessment Guidelines Subdivision K which is currently undergoing revision. A successor document will eventually appear as Series 875, Group B Postapplication Exposure Monitoring Test Guidelines. Regulatory protocols found in these documents utilize dislodgeable foliar residues, foliage-to-human transfer coefficients, and duration of activity to estimate exposure. Dermal absorption factors are then used to estimate dose. However, the experiments from which absorption factors are derived typically involve constant or nearly constant exposures which are not consistent with assumed field exposure conditions. This can lead to inconsistent interpretation and questionable dose estimates. An AFL-CIO challenge to procedures used by EPA to estimate the dose of the fungicide captan [N-trichloromethylthio-4-cyclohexene-1,2-dicarboximide] to strawberry harvesters, which elicited a response from EPA, provides a useful opportunity for examination of the derivation and use of absorption factors. An improved, but still relatively simple, method for dermal dose estimation featuring explicit treatment of the time dependence of absorption has been developed. A benefit of the proposed method is capability for consideration of the effect of delay in post-shift washing on dose. PMID- 10701292 TI - Review of occupational standards and guidelines for hand-arm (segmental) vibration syndrome (HAVS). AB - This article reviews the health effects, treatment, prevention strategies, and international standards for hand-arm vibration syndrome (HAVS). It draws attention to the proposed International Standards Organization (ISO) 1998 revision requirement to base the assessment of vibration on the root-sum-of squares for all directions rather than the dominant direction; the European Economic Commission (EEC) directives with their threshold, action, and ceiling levels; and the intent of the EEC, United Kingdom, and Japanese jurisdictions to propose action levels to prevent the incidence of vibration-induced Raynaud's phenomenon (VWF) from exceeding the background level in their countries. Proposals for the revision of the American Conference of Governmental Industrial Hygienists (ACGIH) Threshold Limit Values (TLVs) are presented. PMID- 10701293 TI - Characterization of an aerosol chamber for human exposures to endotoxin. AB - The objective of this study was to develop and characterize an exposure chamber in which human subjects could be exposed to low dust concentrations carrying an endotoxin coating. An exposure chamber, dust dispersion method, and endotoxin characterization technique were developed for inhalation exposures. A 6.27 m3 exposure chamber was designed and constructed from cinder block, glass windows, and Plexiglas. Using an acetone adhesion process, Enterobacter agglomerans were adsorbed onto respirable cellulose particles to create the endotoxin aerosol. The size distribution of the endotoxin-treated particles was verified using light microscopy and cascade impactors. A dry powder dust generator was refined to consistently disperse small quantities of the aerosol into the chamber to maintain dust concentrations at approximately 250 micrograms/m3. Dust levels during the chamber exposures were monitored using a portable continuous aerosol monitor (PCAM). During initial exposure runs, PCAM monitoring stations were positioned at different locations within a 0.5-meter matrix to document mixing patterns. Total dust and cascade impactor samples were collected throughout each exposure period to characterize the chamber operating system and insure the mean airborne dust concentration fulfilled target levels. A one-factor analysis of variance at the 95 percent confidence interval illustrated that there was not a statistically significant difference in the mean dust concentration throughout the exposure runs compared to the individual runs. Together the consistency of the total dust filters, endotoxin concentrations, and aerosol-monitoring instrument were adequate to allow use of the chamber for experimental studies involving human volunteers. PMID- 10701294 TI - Occupational exposure and respiratory illness symptoms among textile industry workers in a developing country. AB - This study investigates the respiratory health profile of textile mill workers in Bangladesh, aiming to develop workers' awareness and public attention, and to ensure a proper implementation of health and safety measures. Forced vital capacity was measured by peak expiratory flow rate instrument among 210 subjects. The personal history, the occupational history, and the state of health were also determined using a questionnaire and checklists. The subjects who had a considerably low peak expiratory flow rate (< 290 liters/min), and had symptoms of chronic respiratory illness, underwent X-ray examination. A statistically significant low peak expiratory flow rate was identified among 52.9 percent of workers. Among them, 42.9 percent had symptoms of cough with or without phlegm; 5.7 percent had a history of chronic bronchitis and/or asthma, and 4.3 percent experienced chest tightness or breathlessness. This study showed a high degree (p < .001) of respiratory-related illness symptoms present among the workers in the blow/card rooms and the workers in the spinning section. Irrespective of variation of age as well as work pattern, non-smokers were less likely to be affected. Whether worker were occupationally exposed to other incidences was also investigated. The results of these investigations are presented and the findings discussed in light of other studies carried among similar occupational groups. PMID- 10701295 TI - Copper resistance & its correlation to multiple drug resistance in Salmonella typhi isolates from south Karnataka. AB - A study was conducted to ascertain the prevalence of copper resistant Salmonella typhi isolates in south Karnataka. Of the 186 strains studied, 26 (13.97%) were found to be copper resistant. Among the copper resistant strains 19 (73.08%) were found multi drug resistant. All copper resistant strains remained uniformly sensitive to ceftriaxone, ciprofloxacin and norfloxacin. Multiple drug resistance was exclusively associated with E1 phage types. PMID- 10701296 TI - Multi-drug resistant non-typhoidal Salmonella spp. associated with acute diarrhoeal disease. AB - The prevalence of different serotypes of non-typhoidal Salmonella spp. among patients suffering from acute diarrhoea admitted to the Infectious Diseases Hospital, Calcutta was investigated. The predominant serogroup was C and Salmonella infantis was the major serotype isolated followed by S. worthington, S. enteritidis, S. typhimurium, S. weltevereden and S. newport. All the Salmonella strains were isolated from adults. Multidrug resistance to various antimicrobial agents was observed in 37.5 per cent of the strains. All the strains were sensitive to ciprofloxacin, norfloxacin and gentamycin. PMID- 10701297 TI - Evaluation of various methods of susceptibility to ofloxacin in strains of Mycobacterium tuberculosis. AB - A comparison of three methods of susceptibility testing was undertaken on 30 susceptible and 25 resistant strains of Mycobacterium tuberculosis to determine an acceptable in vitro definition of resistance of ofloxacin. The strains were tested by the proportion method on Lowenstein Jensen (L-J) and 7H11 media and also by the BACTEC radiometric method. Using a criterion of 1 per cent or more growth at a concentration of 2 mg/1, there was a 100 per cent agreement with the conventional MIC method by the proportion tests on L-J as well as on 7H11 media. The BACTEC radiometric method, at the same concentration, yielded 98 per cent agreement. Thus, any of these methods could be used depending upon the infrastructure available. PMID- 10701298 TI - Anti-C1q antibody as a marker of disease activity in systemic lupus erythematosus. AB - The present study was conducted to examine the usefulness of anti-C1q antibody as a marker of disease activity in Indian patients with systemic lupus erythematosus (SLE). We standardized the assay for detection of IgG anti-C1q antibody using ELISA. The normal cut-off level was determined by testing 57 healthy, age and sex matched controls to be 53 units/m1 (mean +/- 2 SD). Patients with SEL (97 females and 13 males) were studied and the following parameters were obtained on all: SLE disease activity index (SLEDAI), anti-C1q, anti-ds DNA and C3. Correlations were tested between these parameters using Spearman's rank correlation coefficients. Anti-C1q was found positive in 66 (60%) patients while anti-ds DNA was found in 78 (71%). The positive predictive values of anti-C1q and anti-ds DNA for lupus nephritis were 59 and 61 per cent respectively. The titres of anti-C1q correlated positively with SLEDAI (P < 0.01) and anti-ds DNA (P < 0.01) and negatively with C3 levels (P < 0.001). No significant correlation was observed between anti-C1q positivity and any particular organ involvement. Similarly, no correlation was found between anti-C1q and proliferative lupus nephritis. Anti-C1q was found positive in 5 of 9 patients with moderate SLEDAI scores and negative for anti-ds DNA antibody. It is concluded that anti-C1q antibody can serve as a general marker for lupus activity, supplementing the currently used serum markers. PMID- 10701299 TI - Efficacy of non-breath-hold magnetic resonance cholangiography at midfield strength. AB - The efficacy of non-breath-hold magnetic resonance (MR) cholangiography at mid field strength (0.5 Tesla) was evaluated for delineating biliary anatomy and the cause and extent of biliary obstruction. We performed 65 MR cholangiograms on a mid-field 0.5 Tesla MR unit and correlated them with contrast cholangiography and/or surgery. MR cholangiography was found to be both sensitive and specific in the detection of biliary obstruction and in the definition of its cause (sensitivity 98%, specificity 100%, positive predictive value 100%, negative predictive value 85.7%, accuracy 98%). MR cholangiography accurately predicted the level of obstruction in 94 per cent of strictures. Normal caliber intra hepatic biliary radicles were visualised in only 6 per cent of the MR cholangiograms. In contrast, 94 per cent of dilated intrahepatic biliary radicles were demonstrated. The confluence, and right and left hepatic ducts were visualized in 98 per cent; the gall bladder in 65 per cent; the cystic duct in 45 per cent and the cystic duct insertion in 25 per cent. The extrahepatic bile duct was seen in 82.7 per cent. A normal caliber pancreatic duct was seen in 18 per cent while a dilated pancreatic duct was seen in 86 per cent. The pancreatico biliary junction was visualised in 7 per cent. Non-breath-hold MR cholangiography at midfield strength is a highly accurate method of evaluating the cause and level of biliary obstruction, comparable to high-field MR cholangiography. The spatial resolution however is inadequate for the detection of variations in biliary or pancreatic ductal anatomy when the ducts are of normal caliber. PMID- 10701300 TI - Oxidative stress in alcoholic liver disease. AB - We report on the prooxidant (lipid peroxides) and antioxidant levels (ascorbic acid, reduced glutathione, superoxide dismutate activity) in healthy individuals (30) and patients with cirrhosis (37; 22 alcoholic cirrhosis and 15 non alcoholic cirrhosis). A significant increase in plasma lipid peroxide (P < 0.05) and ascorbic acid (P < 0.01) and a significant decrease in reduced glutathione (P < 0.001) and superoxide dismutase activity (P < 0.05) in haemolysate was observed in cirrhosis patients compared to the control group. A significant decrease in reduced glutathione (P < 0.01) and superoxide dismutase (P < 0.05) activity was also observed when the alcoholic cirrhosis group was compared to non alcoholic group. A significant increase in aspartate transaminase (P < 0.05), gamma glutamyl transaminase (P < 0.01) and aspartate transaminase/alanine transaminase (P < 0.05) ratio was seen in alcoholic cirrhosis group. A significant positive correlation between gamma glutamyl transferase and lipid peroxides (r = 0.48, P < 0.05) was observed in alcoholic cirrhosis. PMID- 10701301 TI - Synergism analysis of biochemical systems. I. Conceptual framework. AB - The detection of synergisms--deviations from additive or linear behaviour--is often an important step in uncovering mechanisms of biochemical processes. Yet, a theoretical background for systemic analysis of synergisms in metabolic networks is lacking. Based on suitable mathematical models, such a theoretical approach should allow predicting synergisms and analysing what mechanistic features contribute to specific synergisms. This work presents a conceptual framework and formalism that fulfil these purposes. The synergism between perturbations of a pair of parameters is quantified as the difference between the response to the simultaneous perturbation of both parameters and the sum of the individual responses to the perturbations of each parameter. A generalisation measures deviations from multiplicative or power-law behaviour. These deviations were called log-synergisms, as in logarithmic coordinates they are quantified in the same way as the synergisms are in Cartesian coordinates. For small perturbations, synergisms and log-synergisms are approximately proportional to the second derivatives (in Cartesian and logarithmic coordinates, respectively) of the observable to the perturbed parameter(s). These derivatives, here called synergism or log-synergism coefficients, measure how steeply the responses diverge from linearity/additivity or power-law/multiplicativity. The formalism now presented allows evaluating (log-)synergism coefficients for systemic steady state responses, and relates these coefficients to intrinsic kinetic properties of the underlying processes. A robust homeostasis of metabolite concentrations requires that these have moderate systemic log- and relative-synergism coefficients. PMID- 10701302 TI - Synergism analysis of biochemical systems. II. Tensor formulation and treatment of stoichiometric constraints. AB - The previous paper outlined a conceptual and mathematical framework for synergism analysis of kinetic models. Though the formalism presented there is adequate for studying simple models, the analysis of large-scale models benefits from the more effective formulation achieved in this work. The present formulation is based on simple tensor operations and takes advantage of the analogy between the formalisms for synergism and log-synergism analysis presented before. Well-known relationships of first-order sensitivity analysis and new relationships for (log )synergism coefficients of various steady-state properties are cast in the new formal setting. The formalism is then extended to models that are subject to constraints between variables, fluxes and/or parameters. This treatment, which generalizes Reder's concept of link matrices, is applied to networks that include moiety conservation cycles [C. Reder, Metabolic control theory: a structural approach, J. Theor. Biol. 135 (1988) 175]. It is also used to take advantage of flux conservation at steady-state to simplify synergism analysis. Issues of numerical effectiveness are briefly discussed, and the theory illustrated with the study of synergistic behaviour in the metabolism of reactive oxygen species and of a scheme of dynamic channelling. PMID- 10701303 TI - A mathematical model of cancer treatment by immunotherapy. AB - In this paper, a detailed mathematical study of cancer immunotherapy will be presented. General principles of cancer immunotherapy and the model equations and hypotheses will be discussed. Mathematical analyses of the model equations with regard to dissipativity, boundedness of solutions, invariance of non-negativity, nature of equilibria, persistence, extinction and global stability will be analyzed. It will also be shown that bifurcations can occur, and criteria for total cure will also be derived. PMID- 10701304 TI - A model of HIV-1 pathogenesis that includes an intracellular delay. AB - Mathematical modeling combined with experimental measurements have yielded important insights into HIV-1 pathogenesis. For example, data from experiments in which HIV-infected patients are given potent antiretroviral drugs that perturb the infection process have been used to estimate kinetic parameters underlying HIV infection. Many of the models used to analyze data have assumed drug treatments to be completely efficacious and that upon infection a cell instantly begins producing virus. We consider a model that allows for less then perfect drug effects and which includes a delay in the initiation of virus production. We present detailed analysis of this delay differential equation model and compare the results to a model without delay. Our analysis shows that when drug efficacy is less than 100%, as may be the case in vivo, the predicted rate of decline in plasma virus concentration depends on three factors: the death rate of virus producing cells, the efficacy of therapy, and the length of the delay. Thus, previous estimates of infected cell loss rates can be improved upon by considering more realistic models of viral infection. PMID- 10701305 TI - [Chlamydia, atherosclerosis, asthma and MS]. PMID- 10701306 TI - [Vasopeptidase inhibitors. Omapatrilat in hypertension and heart failure]. PMID- 10701307 TI - [Neurotoxicity of anti-infective agents]. PMID- 10701308 TI - [Idiopathic peripheral facial paralysis. Therapeutic possibilities]. PMID- 10701309 TI - [Beverages]. PMID- 10701310 TI - [A century of aspirin. The history of the most successful drug of the last ry]. PMID- 10701311 TI - [Phytoestrogens: potential agents for prevention and treatment of breast cancer]. PMID- 10701312 TI - [Fixed solutions by means of melting extrusion--an integrated manufacturing concept]. PMID- 10701314 TI - [AIDS: new FDA approved agents]. PMID- 10701313 TI - [Bisphosphonates in review]. PMID- 10701315 TI - [In Process Citation] PMID- 10701316 TI - [Do angiotensin II receptor antagonists substitute angiotensin converting enzyme inhibitors in the treatment of high blood pressure?]. AB - Angiotensin converting enzyme inhibitors (ACEI) and angiotensin II receptor antagonists (AIIA) are both pharmacological groups that inhibit the actions of angiotensin II. ACEI prevent the formation of angiotensin II from angiotensin I, whereas A II A inhibit the final crucial step of angiotensin II binding with the AT1 receptor site. A similar antihypertensive efficacy has been described for both groups but A II A drugs have a better safety profile above all due to the absence of dry cough. Despite the fact that evidence with ACEI is more conclusive, A II A seems to achieve the same protective effects on the target organ damage in hypertensive patients. At present, ACEI are the drugs of choice in the treatment of patients with cardiac dysfunction and failure. The information of ongoing trials with A II A will be of great value in deciding the optimal treatment for hypertensive patients with different cardiovascular diseases. PMID- 10701317 TI - [Epidemiology of cerebrovascular disease]. PMID- 10701318 TI - [Incidence, mortality and risk factors for stroke in the Manresa Study: 28 years of follow-up]. AB - INTRODUCTION AND OBJECTIVES: The information concerning stroke mortality is limited in Spain, and the information on morbidity is even scarcer similarly to other countries. This is true also for the decrease of frequency observed in the last decades. The objective of this paper is to provide data in the incidence, mortality and cardiovascular risk factors associated to stroke in our surrounding through by the prolonged observation of a working population. MATERIAL AND METHODS: In the Manresa Study, which began in 1968, a cohort of 1,059 men, from 30 to 59 years old, was followed for 28 years. We recorded new cases of fatal and nonfatal stroke and the relationship between stroke incidence and risk factors of cardiovascular disease found in the initial examination. RESULTS: Incidence rate for stroke was 183 x 100,000 per year, 64% of the cases were registered after they turned 60 years of age. Mortality rate due to stroke was 88 x 100,000 per year, 91.6% of fatal cases were over 60 years old. Factors associated to the stroke morbimortality incidence were age, high blood pressure and overweight. In a bivariate regression model, stroke mortality was found significantly associated to the presence of atrial fibrillation, diabetes, hypercholesterolemia and tobacco smoking. CONCLUSIONS: Stroke frequency rates in the Manresa cohort are ranged at a medium level compared to data from other general population studies. The role of atrial fibrillation in the stroke morbimortality has been confirmed. The associated factors, age, high blood pressure and overweight, are similar role to that which was found in other research studies. The priorities in the cerebrovascular disease prevention in our surroundings are discussed. PMID- 10701319 TI - [Percutaneous closure of atrial septal defects: midterm results of a new therapeutic strategy]. AB - OBJECTIVE: To evaluate the midterm results of percutaneous closure of the atrial septal defect using two new devices. PATIENTS AND METHODS: Nine children (weight 19.7 +/- 7 kg, age 5.1 +/- 1.9 years) underwent percutaneous type II atrial septal defect closure through the antegrade pathway under general anaesthesia, and monitored by transesophageal echocardiography. The closing devices used were DAS-Angel Wings and Ampaltzer. RESULTS: The hemodynamic results were: mean diameter of the defects was 11.4 +/- 2 mm by TEE measurement and 12.3 +/- 2.6 mm using balloon occlusion reference. Mean pulmonary artery pressure was 12.7 +/- 2 mmHg and mean pulmonary vascular resistance 1.5 +/- 0.5 U/m2. A total of 13 devices were used: 9 Amplatzer and 4 DAS-Angel Wings. Four Amplatzer through the introducer were retrieved without complications. Two of which because of lack of sufficient stability in the atrial septum because they were too small inappropriate and the other two because of inappropriate expansion of distal disk of the device. Finally in all patients the device was a successfully deployed. The angiographic evaluation immediate post-procedure showed a minimal shunt in five patients that was no longer present by color Doppler echocardiography 24 hours later. The children were discharged 38 +/- 12 hours after the procedure and at a mean follow up of 9.6 +/- 2.2 months they remain asymptomatic without any clinical or technical problems. CONCLUSION: With the right selection of patients percutaneous closure of atrial septal defects can obtain a very high success rate without complications. PMID- 10701320 TI - [Results of coronary stenting in acute myocardial infarction]. AB - OBJECTIVE: To describe the angiographic results and the in-hospital clinical outcome of patients with an acute phase of myocardial infarction treated with coronary angioplasty and stent placement. METHODS: 268 patients with myocardial infarction were treated with angioplasty and coronary stenting within in our center 12 hours after the onset of symptoms from January in 1992 to March 1998. 366 stents were placed (1.4 +/- 0.7 per patient), 35% being Palmaz-Schatz, 26% Wiktor, 21% Multi-Link and 18% others. Stenting was elective in 171 patients (64%), and the majority of patients (91%) were treated with aspirin plus ticlopidine. RESULTS: A successful angiographic result was achieved in 258 patients (96%). Minimum lumen diameter was increased from 0.2 +/- 0.3 to 2.7 +/- 0.7 mm (p < 0.001), and stenosis decreased from 94 +/- 8% to 13 +/- 11% (p < 0.001). Mortality was 15.3% (3.2%, 24.4% and 67.7% in patients in Killip class I, II-III and IV, respectively). Nonfatal reinfarction and recurrent ischemia rates were 2.6% and 9%, respectively. Stent thrombosis occurred in 8 patients (3.0%), and new target vessel revascularization was needed in 12 (4.5%). CONCLUSIONS: Stent placement in acute myocardial infarction is associated with high angiographic success rate, as well as a good in-hospital outcome. Mortality is localized, especially in patients with cardiac failure at the beginning of the procedure. PMID- 10701321 TI - [Treatment of chronic stable angina: follow-up study with nifedipine gastrointestinal therapeutic system. SENIOR Study Group]. AB - OBJECTIVE: To gather information about efficacy and tolerability of nifedipine GITS in patients with stable angina, and its impact on the patient quality of life. PATIENTS AND METHODS: 1076 patients of both sexes (63.5 +/- 12.8 year old) with stable angina (classes I to III of the CCVS) and evidence of coronary disease (43.3% previous myocardial infarction) were included. The treatment with nifedipine GITS 30-60 mg/day (monotherapy or combination) lasted for 6 months. The study variables were: weekly rate of anginal attacks, short-acting nitrate consumption, changes in the antianginal drug treatment, tolerability, and changes in the questionnaire score concerning the quality of life. RESULTS: A decrease in the number of the anginal attacks and in the short-acting nitrates consumption by 80.7% and 83.3%, respectively (both, p = 0.001), was found. Furthermore, the proportion of patients experiencing anginal attacks the week before the assessment visit fell from 71.7% to 10.9% (p < 0.001). At the end of the study, a remarkable decrease in the use of other antianginal medications was seen. Side effects were reported by 10.9% of the patients, 2.7% of which were withdrawn from the study for this reason. A favourable change in the patient quality of life was also noted. CONCLUSION: In patients with stable angina, nifedipine GITS is an effective, safe and well tolerated drug that remarkably enhances the patient quality of life. PMID- 10701322 TI - [The effect of pre-infarction unstable angina on the size of myocardial necrosis]. AB - INTRODUCTION AND OBJECTIVES: Recent studies suggest that preinfarction angina (PA) might induce less myocardial necrosis. The objective of this study is to evaluate whether patients with PA have smaller myocardial infarctions. METHODS: Patients with acute myocardial infarction of less than 12 hours since the onset symptoms were included. PA was defined as unstable angina at rest during the 7 days before the infarction. Infarct size was assessed with the area under the curve of CK-MB levels in the 24 hours following the onset of the infarct. RESULTS: One hundred-seventy nine patients were included, 75 (41.9%) with PA. There were more men in the group with PA (89.3% vs 70.2%, p = 0.004) and a higher prevalence of ex-smokers (38.7% vs 19.2%, p = 0.006). We did not find significant differences in myocardial infarction size between both groups, but a statistically significant interaction between PA and pre-treatment with sulfonylurea drugs was noted (p = 0.050). CONCLUSIONS: Preinfarction angina does not seem to induce less myocardial necrosis in this study. There is a significant interaction between preinfarction angina and pre-treatment with sulfonylurea drugs. PMID- 10701323 TI - [Systematic review of the effectiveness and indications of antidigoxin antibodies in the treatment of digitalis intoxication]. AB - INTRODUCTION AND OBJECTIVES: Cardiac glucoside intoxication is a frequent medical problem given the following: the very narrow therapeutic range, its use in advanced aged patients, in patients with altered renal function, and because of interaction with other drugs. There are two types of digitalis intoxication: one that appears as a complication of the treatment with digitalis, and the other as a result of an accidental ingestion or in suicide attempt. The objective of this study was to review and assess the level of scientific evidence on the effectiveness and the indications of use of Fab fragments of antidigoxine antibodies. METHODS: A systematic bibliographic search in the following databases was made: MEDLINE, The Cochrane Library, The Iowa Drug Information Service, Embase, LMS/R&D Insight, and Indice Medico Espanol. The selected papers were classified according to their level of scientific evidence. RESULTS: Abstracts of 252 references were reviewed. In the reviewed bibliography no controlled, randomized trials were found. Most of the studies found are descriptions of case series or single cases that were treated with antidigoxin Fab fragments. These types of studies provide little or no scientific evidence to speak of. None of the treatment regimes with antidigoxin antibody Fab fragments so far proposed have proven to be valid in a controlled, randomized clinical trial. CONCLUSIONS: There is a very high level of concordance among the studies reviewed with regards to the efficacy and the indications for the use of Fab fragments in severe acute accidental digitalis intoxication and in suicide attempts. Regarding those intoxications that result in patients undergoing digitalis therapy, usual therapeutic approach is traditional treatment and the monitorization of the severity of the intoxication. PMID- 10701324 TI - [Effect of cyclooxygenase inhibition on the adaptation of coronary blood flow to tachycardia]. AB - BACKGROUND: The role of different endothelium-derived vasoactive substances in the regulation of coronary circulation during tachycardia is not well defined. In order to elucidate the contribution of prostacyclin to the adaptation of coronary blood flow to tachycardia, the effect of meclofenamate, a cyclooxygenase inhibitor on the coronary blood flow response to rapid atrial pacing was analyzed in a porcine model. METHODS: A group of seventeen pigs were instrumented for coronary blood flow, aortic pressure and atrial pacing. Heart rate was increased by 20 beats every 5 minutes. Coronary blood flow and aortic pressure were measured, and coronary resistance calculated, basally and at each pacing interval, before and after saline serum (n = 6), meclofenamate 5 mg/kg, i.v. (n = 7) or meclofenamate 35 mg/kg, i.v. (n = 4). RESULTS: Neither saline nor meclofenamate modified the normal increase of coronary blood flow provoked by rapid atrial pacing (163 +/- 28% increase before versus 172 +/- 29% after saline; 159 +/- 21% increase before versus 161 +/- 22% after meclofenamate low doses and 201 +/- 39% before vs 172 +/- 36 after meclofenamate high doses). There were no differences in the response of coronary vascular resistance to tachycardia before and after meclofenamate (44% reduction vs 40% respectively). CONCLUSION: Cyclooxygenase blockade does not modify the response of coronary circulation to rapid atrial pacing, suggesting that prostacyclin does not play a limiting role in the regulation of coronary blood flow during tachycardia in this model. PMID- 10701325 TI - [Spanish Society of Cardiology practice guidelines on arterial hypertension]. AB - High blood pressure is a well-known cardiovascular risk factor that is responsible for an elevated morbidity and mortality. However, although efficacious drugs for treatment and numerous and updated scientific training programs are available, the reality is that only a low percentage of patients are followed up in accordance with the rates which are presently considered normal. The purpose of these guidelines is to provide medical guidance for the prevention, detection and evaluation of hypertension, and to provide the best diagnosis and treatment. The factors involved in cardiovascular complications in the hypertensive patient are multiple. That is why this report places more emphasis in the individual cardiovascular risk stratification as part of the treatment strategy. The information obtained in the most recent studies published confirms the interest in achieving the greatest decrease in rates of blood pressure. This treatment to lower levels is especially useful in the high-risk subgroup. It maintains the necessity of nonpharmacological measures or lifestyle modifications in all patients with high blood pressure who either need or do not need drug therapy. All pharmacological groups may be used, but it is appropriate to choose the specific antihypertensive agent adapted to the clinical and individual situation with the use of low doses of drugs to initiate therapy and the use of appropriate drug combinations. PMID- 10701326 TI - [Spanish Society of Cardiology practice guidelines on ambulatory monitoring of electrocardiogram and blood pressure]. AB - In the present paper, a historical review and a clinical up-date are done on two procedures of great medical interest: Holter electrocardiography and ambulatory blood pressure monitoring. Technical and methodological characteristics of each procedure are carefully exposed, emphasizing each the lack of an international agreement in order to establish regulations that make all the equipment homogeneous and reliable in order to increase both accuracy and reliability in diagnosis. Based on published international scientific documents and the personal experience of the authors, guidelines for clinical applications, indications and limitations of each technique are analyzed in relation to capacities of the Spanish political and social public health system profile. New concepts and dynamics of developments such as; dynamic QT, RR variability or pulse wave velocity are exposed, in the frame of the present time and future for improving efficiency and clinical application. PMID- 10701327 TI - [Blood pressure variability and cardiovascular morbimortality]. AB - Blood pressure is a changing parameter that is influenced by intrinsic body rhythms, physical and emotional environmental factors that act on the individual, and the differences produced by activity and rest periods of every subject. Blood pressure variability seems to be related with target organ damage due to high blood pressure. Its specific effect on left ventricular mass is likely, although it is not absolutely confirmed. Testing blood pressure variability should be a routine exploration in patients with a target organ lesion. PMID- 10701329 TI - [Sudden death (V). Identification and treatment of patients with hypertrophic cardiomyopathy at risk of sudden death]. AB - During the last 20 years, the principal objective in hypertrophic cardiomyopathy research has been the refinement of algorithms for the identification and treatment of patients at risk of sudden death. Sudden death is an important problem in hypertrophic cardiomyopathy, with an incidence of 4-6% in referral populations and approximately 1% in non-referral centers and because it affects young and often asymptomatic patients. We now know that hypertrophic cardiomyopathy is not a single disease, but a group of diseases caused by mutations in genes encoding different sarcomeric proteins. The phenotypic expression depends on multiple modifying genetic and environmental factors. Even though genetic testing is not presently a practical approach in hypertrophic cardiomyopathy risk stratification, it is important to consider new genetic data in the prognostic evaluation of patients. In this paper, we review the published data on risk stratification in hypertrophic cardiomyopathy and we set forth our opinion with regard to the available therapeutic options and their indications in the prevention of sudden death. PMID- 10701328 TI - [Circadian alterations of the cardiovascular system]. AB - Circadian variations have been known for a long time for the influence they have on the physiological systems, including the cardiovascular system. The study of the mechanisms with circadian variation that change the function of the cardiovascular system and its diseases has increased greatly in recent years due to its clinical prominence. Through these studies, physiopathology, epidemiology and factors involved in cardiovascular diseases are more understandable. Thus, the incidence of cardiac events has been clearly associated with the morning hours, as well as the possible mechanisms involved in this variation during the daytime hours. The arterial blood pressure, plasma catecholamine levels and cortisol, platelet aggregation, and fibrinolytic system action are the most implicated mechanisms. From this knowledge, it is possible to design new therapeutic strategies that should consider the time of the day of higher risk for the onset of cardiovascular events. PMID- 10701330 TI - [Aberrant right subclavian artery]. PMID- 10701331 TI - [Absent pulmonary valve syndrome with ductal origin of the left pulmonary artery. Diagnosis only by 2-D echo doppler color flow mapping]. AB - A two-month old infant is described with the rare combination of absent pulmonary valve syndrome, ventricular septal defect, pulmonar "anular" stenosis and ductal origin of the left pulmonary artery. The diagnosis that was confirmed in the operating room was made by 2-D echocardiographic Doppler color flow mapping study without the support of cardiac catheterization. PMID- 10701332 TI - [Multiple changes of the morphology of ST segment in a patient with Brugada syndrome]. AB - The Brugada syndrome is characterized by in a electrocardiographic pattern of right bundle branch block and ST-segment elevation in the right precordial leads, absence of any structural heart disease and syncope episodes or sudden death. We report the case of a 50 year-old men with Brugada syndrome and manifold changes of the precordial morphology of ST segment. PMID- 10701333 TI - [Employment of St. Jude "silzone" valve in the surgical treatment of early prosthetic valve endocarditis: a preliminary case report and review of the literature]. AB - Prosthetic valve endocarditis remains as one of the most life-threatening complication of valve replacement surgery. Homografts are the valve of choice with a lower early risk of endocarditis than other valve substitutes, however they are not always available. Recently a new prosthesis has been introduced with a silver-coated sewing cuff (St. Jude Medical with Silzone coating). Silver is an antimicrobial agent that has been proven to reduce bacterial colonization. We present the case of a 48-year-old man with an early prosthetic valve endocarditis which affected an aortic stentless prosthesis. He was successfully treated with a silver-coated prosthesis. Indications for surgery and the use of this prosthesis as a valuable option in this disease entity are discussed. Although the present patient is an isolated case, the interest of this article is the encouraging result obtained with this new prosthesis for this serious complication. Moreover, the clinical experience is reduced with only a few reports in the literature. PMID- 10701334 TI - [Pulmonary artery primary sarcoma: diagnosis with transthoracic and transesophageal echocardiogram]. AB - Pulmonary artery sarcoma is a rare malignant disease and diagnosis before surgery or autopsy is difficult. We present a case of a pulmonary artery sarcoma diagnosed with transtoracic and transesophagic echocardiogram which was treated surgically. PMID- 10701335 TI - [Pericardial tamponade as the first manifestation of primary hypothyroidism]. PMID- 10701336 TI - Light scattering studies on supersaturated protein solutions. AB - The difficulties associated with protein crystallization is a major obstacle in modern structural biology. The increasing demand from the protein crystal structure community for quick and non-invasive experimental techniques as well as the rapid progress of modern optics and electronics have led during the past decade to a considerable expansion of the laser light scattering techniques. The latter are now very often employed to elucidate the aggregation kinetics of supersaturated protein solutions and the mechanism(s) underlying the early nucleation stages. The experimental verification of the nucleation processes, the prediction of the effective interaction potentials and the development of appropriate diagnostics schemes are discussed. PMID- 10701337 TI - Structural investigations of calcium and zinc binding in proteins. AB - Metal ions are used in a variety of ways in our cells to regulate, activate, and stabilise specific protein molecules. In this review, we describe some of the regulatory functions of calcium and zinc that have been examined using X-ray crystallographic structural studies of specific proteins. These studies indicate that very precise control of cellular activity can be achieved by exploiting the specific physio-chemical properties of different metal ions. PMID- 10701338 TI - Dissection of the class A scavenger receptor by peptide engineering. AB - The construction of portions of a protein provides information about its functional mechanisms. Peptide engineering is a recently developed technique to construct a small tertiary structure or a portion of a protein, and allows expansion to applications for bio- and physicochemistries. In this article, we focus on the class A scavenger receptor, which plays a key role in atherogenesis, and may be involved in other pathogenic processes. The receptor is mainly composed of simple tertiary structures, such as collagen and alpha-helical coiled coil structures, which have different functions. We constructed both the collagen like and alpha-helical coiled coil domains by peptide engineering, and analysed the structure-function relationships of the receptor. Understanding the mechanisms of their functions at the amino acid level should help us to mimic the functional domains and to create de novo designed proteins with new functions. PMID- 10701339 TI - D. J. du Plessis Lecture. Surgical research in a developing country. PMID- 10701340 TI - Acute acalculous cholecystitis--a clinical-pathological disease spectrum. AB - OBJECTIVE: To assess the influence of disease setting on clinical and pathological features of acute acalculous cholecystitis (AAC). DESIGN: Analysis of prospectively accumulated clinical data. Blinded histopathological review. LOCATION OF STUDY: Tygerberg Hospital, Western Cape. PATIENTS: Fifty-seven consecutive patients with AAC treated over a 9-year period. MAIN OUTCOME MEASURES: Clinical, ancillary and pathological features of AAC in each of 3 arbitrarily designated types. Type I (N = 24) occurred in patients hospitalised for trauma or critical illness. Patients with type II disease (N = 20) presented primarily with symptoms of acute cholecystitis. Type III AAC (N = 13) was associated with non-calculous gallbladder outflow obstruction. RESULTS: Type I AAC was associated with the highest mortality rate (45.8%), occurred predominantly in males (75%) and was diagnosed pre-operatively in 50% of patients. Acute ischaemic cholecystitis was the most frequent histological diagnosis (66.7%). Only 1 death (5%) was associated with type II AAC despite patients being older, and all but 2 patients (10%) having chronic underlying disease. Acute cholecystitis was diagnosed pre-operatively in 90% of patients. Thirteen patients (65%) were males. Acute-on-chronic cholecystitis was the most frequent histological diagnosis (50%), followed by acute ischaemic cholecystitis in 30%. Type III was associated with an intermediate mortality rate (23.1%) and was the type most seldom diagnosed pre-operatively (15.4%). Histological findings reflected the nature and duration of underlying obstructive pathology. CONCLUSION: The circumstances in which AAC occurs appear to be associated with distinct clinical-pathological variants of the disease. Their recognition could serve to enhance understanding of this challenging condition. PMID- 10701341 TI - Heterotopic pancreas--an unusual cause of cholecystitis. AB - A 47-year-old woman presented with signs and symptoms of acute cholecystitis. A routine cholecystectomy was performed. No gallstones were present. On histological examination of the gallbladder, a nodule impinging on the lumen was present in the region of the cystic duct. This nodule was composed of mature, uninflamed exocrine and endocrine pancreatic tissue. In the absence of gallstones, the heterotopic pancreas was the cause of obstruction, with subsequent cholecystitis. PMID- 10701342 TI - Inferior vena caval injury in the firearm era. AB - BACKGROUND: This study compared the outcome of intra-abdominal caval injuries in the current era of firearm injuries with the outcome during the previous era of stab wounds. METHODS: Patients with intra-abdominal vena caval injuries treated at King Edward VIII Hospital, Durban, from December 1990 to December 1995 were reviewed. This group was compared with a similar cohort reviewed a decade earlier. RESULTS: The historical group consisted of 28 patients and the current group of 26 patients. Modes of injury in the historical group were: stabs (15, 53%), firearm injuries (7, 33%), blunt trauma (4, 14%) and iatrogenic injuries (2, 7%). Modes of injury in the current group were: stabs (5, 19%), firearm injuries (17, 65%) and blunt trauma (4, 16%). Mortality rose from 35.7% in the historical group to 88% in the current series. This mortality figure included 5 patients who died later from complications of the associated injuries. CONCLUSIONS: Firearm injuries are more destructive than stab wounds. The increase in firearm injuries partly explains this higher mortality. However, the failure to apply current concepts of abbreviated laparotomy and damage control combined with excessive delays in transferring patients to theatre have contributed to this high mortality. PMID- 10701343 TI - [The formation and developmental outlook of medical rehabilitation in the Armed Forces of the Russian Federation]. AB - In medical service system of AF RF rehabilitation means combination of medical, military and professional, social and economic and pedagogical measures directed to recovery of health, fighting efficiency (ability to work) which were disturbed or lost by servicemen because of disease or trauma. In the article the main landmarks of rehabilitation development in Russian military medicine are pointed out, today's state of system on the whole and stages in particular is analyzed, perspectives of development are determined. The authors have noted considerable contribution made by Central Military Clinical Hospital N 6 to development of medical rehabilitation. Arsenal of modern rehabilitation and restorative measures is indicated. Methodological principles of rehabilitation conduction are shown. The main ways in further improvement of medical rehabilitation are development of its specialization, rise in economic and social efficiency of rehabilitation measures at the expense of significant unloading of hospital urgent beds and decrease in periods of patient return to military service who will be ready to perform their duties in whole volume. Introduction of modern methodological and organizational principles of medical rehabilitation into the practice of medical support of the Armed Forces' personnel will contribute to achievement of success in this area. PMID- 10701344 TI - [The methodological aspects of optimizing medical support for the troops from the position of functional systems theory]. PMID- 10701345 TI - [The characteristics of the medical evacuation of the wounded by special aviation transport from areas of armed conflicts]. PMID- 10701346 TI - [Food additives and old Russian grain bread for the army table]. PMID- 10701347 TI - [The methodological approaches to organizing the rehabilitation of servicemen with amputation defects of the extremities]. AB - 10-year experience in prosthetics of extremity stumps in conditions of the Central Military Clinical Hospital N 6 was analyzed. In the hospital during the period from 1989 to 1999 prosthetic appliance was made in 203 patients with amputating defects of extremities, 37 of them with pair stumps and 1 with lack of segments of all extremities. Most patients (80.5%) had stumps at the level of femur and crus. Basing on the analysis of modern scientific literature and experience gained in hospital the authors have formulated organizational and methodical states. Thanks to observance of the stated principles it was possible to optimize considerably the process of medico-psychological and social rehabilitation, to achieve good functional results in all patients treated in the hospital. Adequate use of experience in other military medical and prophylactic institutions will contribute to significant decrease in periods of return of servicemen to social useful labour and increase in efficiency of rehabilitation measures. PMID- 10701348 TI - [The medical rehabilitation of patients with non-insulin-dependent diabetes mellitus at the hospital stage]. PMID- 10701349 TI - [The restoration of professional vision in military specialists at a general rehabilitation center]. PMID- 10701350 TI - [The current diagnosis and treatment of erectile dysfunction]. PMID- 10701351 TI - [Bioacoustical psychocorrection in neurotic disorders]. PMID- 10701352 TI - [Current approaches to the medical rehabilitation of patients with a psychoneurological profile at the hospital stage]. PMID- 10701353 TI - [Traction therapy in the medical rehabilitation of servicemen with vertebrogenic pathology]. PMID- 10701354 TI - [The use of Lerivon in the therapy of mood disorders]. PMID- 10701355 TI - [The treatment of chronic dermatoses with glucocorticoids from the Belupo firm]. PMID- 10701356 TI - [The medico-psychological rehabilitation of participants in combat actions in a general hospital]. AB - There were investigated 453 servicemen--participants of fighting actions in Chechnya [correction of Chechenskaya] Republic in whom the incidence of psychologic stress reactions nearly twice exceeded disorders of other types of psychic dysfunction. 5 groups of combatants were selected and severity of psychic disorders depending on duration of participation in fighting actions was determined. Posttraumatic stress disorders were revealed in 14.5% of servicemen, no signs of psychologic desadaptation were detected in 16.6%, 16.9% had pathologic psychogenic reactions, 18.6% showed pathologic level of psychic disorders, and psychologic stress reactions were noted in 33.4%. The developed individual programs of medico-psychologic rehabilitation depending on type and severity of psychic disorders will allow to reduce the period of restorative treatment and significantly decrease servicemen discharge from the Armed Forces because of psychic disorders. PMID- 10701357 TI - [A system of unconscious verbal audio and visual suggestion in optimizing medical rehabilitation]. PMID- 10701358 TI - [A system for restoring the professional health of flight personnel]. PMID- 10701359 TI - [The psychoneurological characteristics of the initial manifestations of cerebral arteriosclerosis in sailors]. PMID- 10701360 TI - A phase II study of mitomycin C, etoposide, and cisplatin in advanced non-small cell lung cancer. AB - Standard chemotherapeutic regimens, such as cisplatin and etoposide, may improve quality of life and prolong survival in patients with incurable non-small cell lung cancer (NSCLC). This trial was designed to evaluate the activity and toxicity of a regimen combining three of the most active agents against advanced stage NSCLC: mitomycin C, etoposide, and cisplatin (MEP). Sixty-eight patients with stage IIIB (pleural effusion) or IV NSCLC received cisplatin 80 mg/m2 i.v. on day 1 and etoposide 80 mg/m2 i.v. on days 1, 2, and 3 every 3 weeks along with mitomycin C 10 mg/m2 i.v. on day 1 of the first and third cycles for a median of four cycles (range, 1-11). Median age was 59 years, and nine patients were enrolled after relapse from previously treated early-stage NSCLC. Eighty-eight percent of patients had stage IV disease, and 14 (21%) had brain metastases at diagnosis. Palliative radiotherapy was given to 10 patients (15%) before MEP and to 17 (25%) concurrent with MEP. The major toxicity of MEP was myelosuppression, with grade 3-4 neutropenia in 74% of patients. Sixteen patients (24%) had documented infections, and there were eight (12%) treatment-related deaths. Partial response was observed in 24 patients (35%) with a median duration of 4.4 months, (range 1.4-13 months). Median survival was 8.1 months (range, 1-34 months), and 1-year survival was 32%. The addition of mitomycin C to cisplatin and etoposide resulted in response and survival rates comparable with those achieved with standard regimens in patients with advanced NSCLC but was associated with substantial hematologic toxicity and unacceptable treatment related mortality. PMID- 10701361 TI - c-erbB-2 gene amplification in nasopharyngeal carcinoma. AB - Amplification of the c-erbB-2 gene has been associated with poor prognosis in different types of cancer. However, there are no data on the c-erbB-2 expression levels in nasopharyngeal cancer. In this study, amplification of the gene has been investigated in the tumor tissue of patients with nasopharyngeal cancer by competitive polymerase chain reaction c-erbB-2 amplification was observed in 43.3% of the patients. The increase in the gene copy number correlated with the T stage. No correlation was found with lymph node involvement, histologic grade, differentiation, presence of metastases, or age and sex. We conclude that c-erbB 2 amplification may contribute to the pathogenesis of nasopharyngeal cancer. Our report is the first study investigating the expression of the c-erbB-2 gene in nasopharyngeal cancer at the DNA level. PMID- 10701362 TI - Fatigue patterns observed in patients receiving chemotherapy and radiotherapy. AB - The purpose of this study was to describe the patterns of cancer-related fatigue (CRF) and vigor in patients receiving chemotherapy or radiation therapy. Five studies that measured fatigue and vigor with the Profile of Mood States were used to describe the pattern of CRF and vigor during and after both types of treatment. Repeated-measures ANOVA was used to determine differences over time in each study. Results demonstrate different patterns of CRF for patients receiving chemotherapy and radiation therapy. Chemotherapy-related CRF peaks in the days after chemotherapy, whereas radiation therapy-related CRF gradually accumulates over the course of treatment. The CRF associated with both forms of treatment gradually declines over time. The prevalence, intensity, and persistence of CRF during treatment and for months after treatment is complete make this symptom one that cannot be ignored. PMID- 10701363 TI - Enhancement of 5-fluorouracil anabolism by methotrexate and trimetrexate in two rat solid tumor models, Walker 256 carcinosarcoma and Novikoff hepatoma, as evaluated by 19F-magnetic resonance spectroscopy. AB - Although 5-fluorouracil (5-FU) is one of the most effective single agents in treating solid tumors, its low effectiveness as a single agent has led to development of a number of modulators intended to enhance its therapeutic effectiveness. Of these, methotrexate (MTX) and trimetrexate (TMTX) have been shown to have synergistic anticancer activity with 5-FU. The effect of these two drugs on the uptake and the intratumoral metabolism of 5-FU was studied in two rat tumor models using 19F-nuclear magnetic resonance spectroscopy: on excised samples of Walker 256 carcinosarcoma and noninvasively (in vivo) in Novikoff hepatoma. In the rats bearing the Walker 256 tumor, a 4-hr pretreatment with MTX showed the maximal increase in the rate of conversion from 5-FU to its fluorinated nucleotides/nucleosides. In the rats bearing the Novikoff hepatoma, both modulators increased the amounts of cyctotoxic anabolites of 5-FU, but at the doses administered, the cumulative amounts of 5-FU anabolites formed after MTX were significantly higher than those formed after TMTX or after saline control. On the other hand, the increase in the levels of the fluorinated nucleotides/nucleosides after TMTX peaked at a later time. The possible significance of these findings is that timing of administration of a modulator is important because it affects both transport and metabolism of 5-FU. The two modulators studied, both antifolates, act differently on transport and on metabolism: MTX affects both, whereas TMTX, at the level studied, appears to affect predominantly the metabolic process. In addition, significant differences exist between tumor models. These data suggest possible mechanisms and processes that should be studied further in humans, using these noninvasive pharmacokinetic imaging methods for monitoring 5-FU targeting and metabolism. PMID- 10701364 TI - A phase II study of 9-aminocamptothecin in patients with refractory breast cancer. AB - We evaluated 9-aminocamptothecin (9-AC) in patients with metastatic or locally recurrent breast cancer who were no longer responsive to standard therapy. Patients were treated with 9-AC with a 72-hr continuous infusion given at a dose of 45 micrograms/m2/hr every 2 weeks. Granulocyte colony-stimulating factor 5 micrograms/kg was given subcutaneously for 7-10 days after completion of the treatment. Eighteen patients were treated, with all patients assessable for toxicity and 15 patients assessable for response. There were two partial responses seen in the 15 patients lasting 3.5 and 5 months, respectively. The major toxicity seen was myelosuppression, with 12 patients having grade 3 or greater granulocytopenia with four episodes of significant infectious complications. In addition, significant thrombocytopenia was seen in 14 patients. The other complications commonly seen were nausea and vomiting and alopecia. 9-AC given as a 3-day continuous infusion has limited activity in previously treated metastatic and locally recurrent breast cancer. PMID- 10701365 TI - Role of taxanes in adjuvant therapy. PMID- 10701366 TI - ras-p21-induced cell transformation: unique signal transduction pathways and implications for the design of new chemotherapeutic agents. PMID- 10701367 TI - Psychosexual adjustment in adolescent cancer survivors. PMID- 10701368 TI - Trends in Medicaid and managed care. PMID- 10701369 TI - Radiation therapy options in the treatment of prostate cancer. PMID- 10701370 TI - Chromosome breaks and genomic instability. AB - Tumorigenesis is known to result from multiple genetic changes. Although endogenous and environmental insults can damage DNA, cellular mechanisms exist to repair various forms of damage or to kill those cells irreparably damaged. Hence, the accumulation of numerous genetic changes that would lead to cancer in normal cells is extremely rare. Nevertheless, disruption of a DNA repair pathway has the potential to expedite tumorigenesis by resulting in a cell that is hypermutable. Multiple pathways exist to repair the various forms of DNA damage that can cause mutagenesis. Recent studies have demonstrated a key role for homologous recombination in DNA repair, in particular in the repair chromosomal double strand breaks. This review summarizes those studies and discusses how disruption of homologous recombination pathways can create genetic instability. PMID- 10701371 TI - Small bioactive peptides and cell surface peptidases in androgen-independent prostate cancer. PMID- 10701372 TI - Cancer-related fatigue. PMID- 10701373 TI - Treatment of acute myeloid leukemia M3 in a patient with Crohn's disease. PMID- 10701374 TI - PubMed Central: signing on. PMID- 10701375 TI - Osler's unusual case. PMID- 10701376 TI - Violence in the health care workplace. PMID- 10701377 TI - Peerless accuracy (or not) PMID- 10701378 TI - High marks for the physical exam. PMID- 10701379 TI - High marks for the physical exam. PMID- 10701380 TI - High marks for the physical exam. PMID- 10701381 TI - High marks for the physical exam. PMID- 10701382 TI - Effect of preventive home visits by a nurse on the outcomes of frail elderly people in the community: a randomized controlled trial. AB - BACKGROUND: Timely recognition and prevention of health problems among elderly people have been shown to improve their health. In this randomized controlled trial the authors examined the impact of preventive home visits by a nurse compared with usual care on the outcomes of frail elderly people living in the community. METHODS: A screening questionnaire identified eligible participants (those aged 70 years or more at risk of sudden deterioration in health). Those randomly assigned to the visiting nurse group were assessed and followed up in their homes for 14 months. The primary outcome measure was the combined rate of deaths and admissions to an institution, and the secondary outcome measure the rate of health services utilization, during the 14 months; these rates were determined through a medical chart audit by a research nurse who was blind to group allocation. RESULTS: The questionnaire was mailed to 415 elderly people, of whom 369 (88.9%) responded. Of these, 198 (53.7%) were eligible, and 142 consented to participate and were randomly assigned to either the visiting nurse group (73) or the usual care group (69). The combined rate of deaths and admissions to an institution was 10.0% in the visiting nurse group and 5.8% in the usual care group (p = 0.52). The rate of health services utilization did not differ significantly between the 2 groups. Influenza and pneumonia vaccination rates were significantly higher in the visiting nurse group (90.1% and 81.9%) than in the usual care group (53.0% and 0%) (p < 0.001). INTERPRETATION: The trial failed to show any effect of a visiting nurse other than vastly improved vaccination coverage. PMID- 10701383 TI - Time required for approval of new drugs in Canada, Australia, Sweden, the United Kingdom and the United States in 1996-1998. AB - BACKGROUND: The timeliness with which national regulatory agencies approve new drugs for marketing affects health care professionals and patients. An unnecessarily long approval process delays access to new medications that may improve patients' health status. The author compared drug approval times in Canada, Australia, Sweden, the United Kingdom and the United States. METHODS: Application and approval dates of new chemical or biological substances (excluding diagnostic products, and new salts, esters, dosage forms and combinations of previously approved substances) approved for marketing in the 5 countries from January 1996 to December 1998 were requested from the relevant pharmaceutical companies. Data on new drug approvals during the study period were also obtained from the national drug regulatory agencies in Canada, Australia and Sweden and from publications of the US Food and Drug Administration. RESULTS: A total of 219 new drugs were identified as being approved in at least one of the countries during the study period: 23 (10.5%) in all 5 countries, 23 (10.5%) in 4, 27 (12.3%) in 3, 42 (19.2%) in 2, and 104 (47.5%) in 1 country. By individual nation, 97 drugs were identified as being approved in Canada, 94 in Australia, 107 in Sweden, 55 in the UK and 123 in the US. Approval times in Canada and Australia were similar (medians 518 and 526 days respectively), but both countries had significantly longer approval times than Sweden (median 371 days), the UK (median 308 days) and the US (median 369 days). This pattern was consistent across all 3 years and for the 23 new drugs approved in all 5 countries during the 3-year period. Median approval times in Canada were similar in all of the reviewing divisions of Health Canada's Therapeutic Product Program (539-574 days) except the Central Nervous System Division (428 days) and the Bureau of Biologics and Radiopharmaceuticals (698 days). INTERPRETATION: Median drug approval times during 1996-1998 decreased by varying amounts from the 1995 values in all 5 countries. However, the median approval time in Canada continues to be significantly longer than the times achieved in Sweden, the UK and the US, and it remains considerably longer than Canada's own target of 355 days for all new drugs. PMID- 10701384 TI - Health-related quality of life among final-year medical students. PMID- 10701385 TI - Evaluating innovation in the care of Canada's frail elderly population. PMID- 10701386 TI - X-rays and technology as metaphor. PMID- 10701387 TI - Jung at heart: assessing one's suitability for medical training. PMID- 10701388 TI - Medical journals are dead. Long live medical journals. PMID- 10701389 TI - Pharmaceutical policies in Canada: another example of federal-provincial discord. AB - Pharmaceutical policy in Canada is set at both the federal and provincial levels of government. The federal government is responsible for intellectual property rights of manufacturers (patents) and the initial approval and labelling of prescription drugs and for ensuring overall market competitiveness. The provincial government has responsibility and jurisdiction over the funding of all health care services, including pharmaceuticals. Various interactions between the pharmaceutical industry, the federal and provincial governments and consumers have shaped the current landscape for prescription drugs in Canada. One key failing of the system is that the federal government is almost completely insulated from the impact of its policies because, although it regulates drug prices, it does not buy any drugs. In contrast, provincial governments have no jurisdiction over market competitiveness or pricing, yet end up paying for most of the drug expenditures incurred. PMID- 10701390 TI - Plastic bread-bag clips in the gastrointestinal tract: report of 5 cases and review of the literature. AB - Plastic bread-bag clips have been identified as a cause of local perforation or obstruction at many sites in the gastrointestinal tract. This study is the largest case series yet reported, consisting of 3 cases presenting as small-bowel perforation, 1 case in which the clip was found incidentally in the small bowel at laparotomy during vascular surgery and 1 case in which the clip was found incidentally in the small bowel at autopsy. In all cases there was no radiographic evidence to suggest a foreign body in the gastrointestinal tract. People older than 60 years of age who have either partial or full dentures seem to be particularly at risk for the accidental ingestion of these devices. If accidentally ingested, plastic bread-bag clips represent a significant health hazard. As the population ages, small-bowel perforation secondary to ingestion of such clips may occur with increasing frequency. The authors recommend elimination or redesign of the clips, to prevent their being swallowed and becoming impacted in the small bowel or to allow them to be identified in the gastrointestinal tract by conventional radiography. PMID- 10701391 TI - Automated external defibrillation: is survival only a shock away? PMID- 10701392 TI - Continuous positive airway pressure for congestive heart failure. PMID- 10701393 TI - Continuous venovenous hemodiafiltration for renal failure and sepsis. PMID- 10701394 TI - Report card on renal transplantation. PMID- 10701395 TI - Latest ER crisis hit communities large and small. PMID- 10701396 TI - Military medical service no longer has MD at helm. PMID- 10701397 TI - American Academy of Allergy, Asthma, and Immunology 56th annual meeting. San Diego, California, USA. March 3-8, 2000. Abstracts. PMID- 10701398 TI - [Primary epiploic appendicitis: CT diagnosis for conservative treatment]. AB - OBJECTIVE: Primary epiploic appendicitis is an uncommon, self-limited disease with spontaneous resolution. Diagnosis is usually made at surgery as the disease is generally mistaken for acute appendicitis or sigmoid diverticulitis. CT allows a non-invasive diagnosis, thus avoiding unnecessary surgery. The condition can be managed conservatively with the use of analgesic drugs only, and clinical evolution is uneventful. The aim of this study is to report the CT features of this pathologic process before and after medical treatment by analgesics. PATIENTS AND METHOD: Six patients (4 men and 2 women aged from 23 to 70 year old mean; 29 year old) underwent abdominal CT scan because of acute abdominal pain located in the left lower quadrant (n = 3) and right lower quadrant (n = 3). No patient had fever. Laboratory findings were normal in all cases. Follow-up CT scans were obtained in 4 patients respectively at 2, 4, 8 and 80 weeks. RESULTS: CT scan showed in all cases a fatty mass located to the anterolateral wall of the colon, delineated by an hyper attenuating rim. Infiltration of the pericolic fat was noted in all cases. Follow up CT scans obtained in 4 patients showed that the inflammatory signs had cleared in all cases, the lesion had disappeared (n = 2), decreased in size (n = 1) or a residual paracolic node was observed (n = 1). CONCLUSION: Primary epiploic appendicitis is a rare disease. Knowledge of CT signs allowed the correct diagnosis and a conservative management. PMID- 10701399 TI - [Importance of allergologic consultation after a ++perianesthetic complication. Report on a year of activities at the Poitiers University Hospital Center]. AB - OBJECTIVE: Patients who present an accident during general anesthesia must be tested 4 to 8 weeks later in order to determine the exact cause and prevent any further acute accidents. We report the results of a prospective study based on a 1-year period of allergology consultations after peroperative accidents in the Poitiers University Hospital. PATIENTS AND METHODS: Previous history, surgical procedure, the nature and gravity, of the reaction, risk factors and tests results were recorded. All observations were transmitted to the Regional Pharmacovigilance Center. The level of causality of the administered drug was determined from these data which were integrated into the national computer bank of side effects. RESULTS: After 2,500 general anesthesia procedures, a total of 21 patients had been referred to the allergology consultation. Eighteen were positive for at least 1 test (skin tests or laboratory tests). Among the substances administered, causality was probable for 16, possible for 4 and doubtful for 1. CONCLUSION: Our findings demonstrate the importance of allergy tests and the need for close collaboration between anesthetists, allergologists and pharmacologists. PMID- 10701400 TI - [Right heart failure caused by thiamine deficiency (cardiac beriberi)]. AB - BACKGROUND: Vitamin B1 deficiency (beriberi) is very uncommon in France. It leads to high output cardiac failure totally different from the situation observed in alcoholic patients. We report a typical case of cardiac beriberi. CASE REPORT: The patient was referred for dyspnea with high output cardiac failure. Echocardiography evidenced severe pulmonary hypertension and high cardiac output. The more common causes of heart failure were ruled out. The dietary habits of the patient (suggested beriberi which was confirmed by the low serum thiamin and therapeutic test with vitamin B1. DISCUSSION: High output cardiac failure should suggest possible Shoshin beriberi, particularly in subjects with imported dietary habits living in a precarious socioeconomic situation. PMID- 10701401 TI - [Acute interstitial pneumopathy with fever caused by hydroxyurea]. AB - BACKGROUND: Hydroxyurea has few side effects excepting the known bone marrow toxicity. Fever with or without pneumonia has occasionally been reported. CASE REPORT: A patient given hydroxyurea for polycythemia suddenly developed severe interstitial pneumonia with fever and hypoxemia. All bacteriological tests were negative and an empirical antibiotic regimen was ineffective. Fever recurred after reintroducing hydroxyurea and definitive cure was achieved after its withdrawal. The clinical course was rapidly favorable without the need for corticosteroids. DISCUSSION: Fever, and in some cases interstitial pneumonia, in patients given hydroxyurea generally suggests an infection. However, 15 cases of pneumonia have been reported as caused by hydroxyurea in patients treated for a myeloproliferative syndrome. Delay to onset is 3 to 8 weeks after initiating treatment. The course is favorable after withdrawal, with or without corticosteroids. Fever may be the only sign of a drug reaction, resolving with withdrawal and recurring at re-challenge. The underlying mechanism remains unknown. Definitive cure can be achieved by discontinuing hydroxyurea, avoiding the need for further investigations. PMID- 10701402 TI - [Pneumococcal pelvioperitonitis revealing HIV seropositivity]. PMID- 10701403 TI - [Acute methadone poisoning]. PMID- 10701404 TI - [Immune deposit glomerulonephritis in an HIV-infected patient after discontinuance of antiretroviral treatment]. PMID- 10701405 TI - [Dangerous connection: alcohol and youth...]. PMID- 10701406 TI - [Medical ethics and therapeutic progress: the example of lung cancer. Hippocrates, help!]. PMID- 10701407 TI - [The 39th ICAAC (San Francisco) and the 7th European Conference on Clinical Aspects and Treatment of HIV-infection (Lisbon). HIV infection: antiretroviral agents in the development and trial stage]. PMID- 10701408 TI - [Natural latex allergy. Primary and secondary prevention in work environment]. AB - AT RISK GROUPS: The incidence of latex hypersensitivity of latex hypersensitivity has increased over the last decade. The main at-risk groups for developing latex allergy are: health care workers and employees working in latex industries, patients with atopic diathesis and subjects with repeated surgical procedures during childhood. SENSITIZATION: The use of cornstarch powder gloves can sensitize healthy subjects and exacerbate symptoms of allergic patients as the powder spreads the latex allergens into the environment. PRACTICAL ATTITUDE: We propose here some practical recommendations for prevention, both for the general population and for allergic subjects. PMID- 10701409 TI - [Management of diabetes during corticosteroid therapy]. AB - BACKGROUND: Corticosteroids are generally contraindicated in diabetic patients due to the risk of disrupting glucose control leading to acute decompensation. In some cases however, corticosteroid therapy can be beneficial if given early with a well-controlled regimen. Glucose disequilibrium after withdrawal can be anticipated with proper knowledge of the pharmacokinetics of the glucocorticoid used. FOR PATIENTS WITH TYPE I DIABETES: Ketose acidosis is a real risk in these patients. Insulin dose must be increased and the administration scheme optimized. FOR PATIENTS WITH TYPE II DIABETES: Whether oral drugs should be continued is a question of debate, excepting cases where the underlying disease might cause acute decompensation requiring insulin. Outside this situation, oral drugs can be continued at a higher dose if the fasting serum glucose is below 2 g/L. Finally, it is important to recognized steroid-induced diabetes in order to initiate proper antidiabetic measures. FOR ALL PATIENTS: The glucose curve is reproducible. Basically, the postprandial level rises, warranting repeated insulin injections. Rapid-release analogs and alpha-glucosidase inhibitors appear to be promising; biguanides affect insulin resistance. PMID- 10701410 TI - [Non-steroidal anti-inflammatory drugs with selectivity for cyclooxygenase-2 in Alzheimer's disease. Rationale and perspectives]. AB - POSSIBLE INFLAMMATORY MECHANISMS: Alzheimer's disease (AD) is a degenerative disease of the brain including possibly inflammatory mechanisms, as illustrated by the presence of activated microglial cells in the periphery of senile plaques and neurofibrillary tangles and the subsequent release of proinflammatory mediators with neurotoxic potency. RATIONALE FOR NSAID USE: Although not firmly demonstrated, the rationale for the prescription of non steroidal anti inflammatory drugs (NSAIDS) as neuroprotective agents in AD lies on epidemiological data having shown a reduced risk of developing AD in patients on long-term therapy with NSAIDs (non selective cyclo-oxygenase [COX] inhibitors). RATIONALE FOR THE USE OF SELECTIVE COX-2 INHIBITORS: The rationale for the prescription of selective COX-2 inhibitors as neuroprotective drugs in AD lies on: Epidemiological data having shown a reduced risk of developing AD in patients treated with anti-inflammatory doses of classical NSAIDs (inhibition of COX-1 and COX-2) but not with antithrombotic doses of aspirin (selective inhibition of COX 1), Cellular experiments, Demonstration of a better gastro-intestinal (GI) safety profile with selective COX-2 inhibitors than with classical NSAIDs in short-term studies, allowing a possible long-term use in AD. BEFORE PRESCRIBING: COX-2 may have an ambivalent functionality in the brain since the basal production of prostaglandins through COX-2 may participate in neuronal homeostasis whereas the expression of COX-2 is associated with brain development. Classical NSAIDs are ineffective in reducing the formation of senile plaque and neurofibrillary tangles in AD, which is consistent with an ability to reduce inflammation associated with activation of microglia but illustrates their failure to suppress the degenerative process. Prophylactic use of selective COX-2 NSAIDs can be considered on the basis of their good GI safety after 6 months of marketing in United States but need to be confirmed for a longer time. CURRENT TRIALS: Clinical studies focusing on both the prevention and the slowing down of early AD are under way with two recently launched selective COX-2 inhibitors, celecoxib and rofecoxib. PMID- 10701411 TI - [Cardiac anomalies in chronic renal failure]. AB - PRINCIPAL CARDIOVASCULAR COMPLICATIONS IN END STAGE RENAL DISEASE: Cardiovascular diseases are the leading causes of morbidity and mortality in end stage renal disease patients. Very often, complications observed are left ventricular hypertrophy and various forms of arterial degenerative lesions involving coronary arteries, less frequently pericarditis and calcifying valvulopathy are diagnosed. THE REASONS ARE COMPLEX: Risk factors can be either specific of uremia per se such as anemia, overhydration, fistula or the same as in the general population. Hemodynamic alterations including tensile stress or blood flow play a major role associated to various locally or generally generated substances whose role remains currently to be determined. THEIR TREATMENTS: Treatments of cardiovascular complications are not specific in this end stage renal disease population but are more often the treatment of the etiology: reduction of fistula blood flow, increase of hemoglobin, best control of weight gain between two hemodialysis sessions or blood pressure control. PMID- 10701412 TI - [A multitude of glandular-cystic gastric polyps]. PMID- 10701413 TI - Simultaneous spectrophotometric and volumetric determinations of amoxycillin, ampicillin and cloxacillin in drug formulations: reaction mechanism in the base catalysed hydrolysis followed by oxidation with iodate in dilute acid solution. AB - A method for the analysis of degraded products of amoxycillin, ampicillin and cloxacillin in drug formulations, obtained as a result of their base hydrolysis is described. Simultaneous spectrophotometric and volumetric determinations of the antibiotic is based on the neutralization of the degraded product by dilute hydrochloric acid to get a pH approximately 2 to be conducive for redox titration using potassium iodate as titrant. A red purple colour is developed in carbon tetrachloride at the end point. Spectrophotometry is done after separating the organic layer and measuring the absorbance of red-purple colour at lambda(max) 520 nm. The pathways of different degraded products and their oxidation mechanism is described on IR, TLC and UV spectroscopic studies. PMID- 10701414 TI - Fluorimetric determination of prenalterol hydrochloride in pharmaceuticals and biological fluids based on its oxidation reaction by hexacyanoferrate(III). AB - A rapid and sensitive fluorimetric method for the determination of prenalterol hydrochloride is presented, based on the oxidation of the analyte with potassium hexacyanoferrate(III) in a slightly alkaline medium (pH 9.23). The different experimental parameters were carefully studied and incorporated into the procedure. The oxidation product exhibits a blue fluorescence with its emission maximum at 427 nm, and excitation maximum at 314 nm. Fluorescence intensity is a linear function of prenalterol hydrochloride concentration over the range of 0.2 3.6 microg/ml(-1) in the solution finally measured. The method was successfully applied to the determination of prenalterol hydrochloride in pharmaceutical formulations and biological fluids. A proposal for the reaction pathway is suggested. PMID- 10701415 TI - Quantification of hyaluronan in pharmaceutical formulations using high performance capillary electrophoresis and the modified uronic acid carbazole reaction. AB - The amount of hyaluronan (HA) in pharmaceutical formulations was determined by high-performance capillary electrophoresis (HPCE) and the results were compared with the carbazole reaction established by Bitter and Muir (T. Bitter, H.M. Muir, Anal. Biochem. 4 (1962) 330-334), HA analysis was performed in less than 10 min by using an untreated fused silica capillary with bubble detection cell. The influence of several buffers and pH values was examined. Calibration curve shows good linearity from 0.01 to 5.0 mg/ml. The lower limit of detection by monitoring the absorbance at 195 nm was 10 microg/ml at a signal to noise ratio of 5. PMID- 10701416 TI - Indirect polarographic and cathodic stripping voltammetric determination of cefaclor as an alkaline degradation product. AB - Cefaclor is not reducible at a mercury electrode, but it can be determined polarographically and by cathodic stripping voltammetry as its initial alkaline degradation product which is obtained in high yield by hydrolysis of cefaclor in Britton-Robinson (B-R) buffer pH 10 at 50 degrees C for 30 min (reduction peak at pH 10, -0.70 V). Differential pulse polarographic calibration graphs are linear up to at least 1 x 10(-4) mol/l(-1). Recoveries of 93% of the cefaclor (n = 3) were obtained from urine spiked with 38.6 microg/ml(-1) using this polarographic method with 1 ml urine made up to 10 ml with pH 10 buffer. Using cathodic stripping voltammetry and accumulating at a hanging mercury drop electrode at - 0.2 V for 30 s, linear calibration graphs were obtained from 0.35 to 40 microg/ml(-1) cefaclor in B-R buffer pH 10. A relative standard deviation of 4.2% (eta = 5) was obtained, and the limit of detection was calculated to be 2.9 ng/ml(-1). Direct determination of cefaclor in human urine (1 ml of urine was made up to 10 ml with pH 10 buffer) spiked to 0.39 microg/ml(-1) was made (recovery 98.6%). PMID- 10701417 TI - Stereoselective determination of trihexyphenidyl in human serum by LC-ESI-MS. AB - The antiparkinsonian drug trihexyphenidyl (THP) is currently manufactured and administered as a racemate. However, stereochemistry can play significant role in the drug's pharmacokinetics, biotransformation, metabolism, interaction with cellular and tissue components and overall effect on human body. It is necessary to consider such a drug as a mixture of two compounds (drug enantiomers), with their own effect on the human body. The present paper describes a simple and sensitive LC-MS method for the stereoselective determination of THP in human serum. In this study, the sample was prepared by a solid-phase extraction (SPE) procedure. The enantiomer separation was done using native beta-cyclodextrin stationary phase LC column. The combination of ESI-MS detection and SPE showed excellent sensitivity and selectivity of the method. The limits of detection of <0.1 ng/ml can be easily achieved, which is 7,000 times lower than the detection limits achievable by a UV detection method. The method has at least a 3-order of magnitude linear dynamic range for both enantiomers (concentrations up to 1,323 ng/ml were tested). This is 24 times wider than the therapeutic range of THP (peak THP plasma concentration of 55 ng/ml was previously reported). The recoveries of THP enantiomers from the human serum were > 95%. PMID- 10701418 TI - A simple high-performance liquid chromatography assay for simultaneous determination of plasma norepinephrine, epinephrine, dopamine and 3,4 dihydroxyphenyl acetic acid. AB - A reversed-phase HPLC assay coupled with electrochemical detection for simultaneously measuring plasma levels of norepinephrine, epinephrine, dopamine, and 3,4-dihydroxyphenylacetic acid (DOPAC) was developed. Separation of the catecholamines and the internal standard isoproterenol was obtained by a mobile phase consisting of 7% methanol in 0.1 M citrate buffer containing 0.3 mM sodium ethylenediaminetetraacetic acid (EDTA), and 0.5 mM 1-octanesulfonic acid, operated under isocratic condition at a flow rate of 1.2 ml/min. The potential of the guard cell was set at +650 mV, the first electrode of the analytical cell at +100 mV and the second at + 350 mV. Using a signal-to-noise ratio of > 3, the minimum detection limit assessed by direct on column injection was < 10 pg for analyte. The assays were linear from basal concentrations to 400 ng/ml. The intra and inter-assay variations were < 10 and 15%, respectively. The assay has been applied successfully to measure plasma concentrations of these catecholamines in humans, rabbits and rats. PMID- 10701419 TI - Development of a chiral HPLC method to evaluate in vivo enantiomeric inversion of an unstable, polar radiosensitizer in plasma. AB - A chiral HPLC method to quantify in vivo enantiomeric inversion of prodrug CI 1010 (IR) or its drug IIR (PD 146923), a radiosensitizer, upon X-irradiation of dosed rats was developed. These polar enantiomers were separated only by using normal-phase chiral HPLC. A Chiralpak AS column provided the best separation. Isolation of analytes from plasma employed solid-phase extraction (SPE), and required conditions that were compatible with normal-phase HPLC. Options for SPE were restricted by the chemically reactive nature of both prodrug and drug, which produced analyte losses as high as 100%. Acceptable recoveries using SPE required evaluation of conditions for analyte chemical stability. The validated method gave a lower-limit of quantitation (LLOQ) of 200 ng/ml for each enantiomer extracted from 0.15 ml of plasma. The LLOQ of the inverted enantiomer could be detected in the presence of 10,000 ng/ml of the dosed enantiomer. Precision (RSD) ranged from 14.2 to 4.4%, and from 24.2 to 5.1% for IIS and IIR, respectively. Accuracy (RE) was +/- 13.1 and +/- 13.2%, respectively. Recoveries ranged from 44.3 to 71.4%, and from 40.7 to 67.9%, for IIS and IIR, respectively. PMID- 10701420 TI - Application of first-derivative UV-spectrophotometry, TLC-densitometry and liquid chromatography for the simultaneous determination of mebeverine hydrochloride and sulpiride. AB - Three methods are described for the simultaneous determination of mebeverine hydrochloride (MB) and sulpiride (SU) in combined pharmaceutical tablets. The first method depends on first-derivative ultraviolet spectrophotometry, with zero crossing measurement method. The first derivative amplitudes at 214.2 and 221.6 nm were selected for the assay of MB and SU, respectively. Calibration graphs follow Beer's law in the range of 10-30 and 2-8 microg/ml(-1), and the linearity was satisfactory (r = 0.9999), for MB and SU, respectively. The second method was based on the application of the thin layer chromatographic separation of both drugs followed by the densitometric measurements of their spot areas. After separation on silica gel GF254 plates, using ethanol: diethyl ether: triethylamine (70:30:1 v/v) as the mobile phase, the chromatographic zones corresponding to the spots of MB and SU were scanned at 262 and 240 nm, respectively. The calibration function was established in the ranges of 4-12 microg for MB and 2-8 microg for SU. The third method was an internal standard procedure based on high performance liquid chromatographic separation of the two drugs on a reversed-phase, Bondapak CN column. The detection was done at 243 nm using buclizine hydrochloride as internal standard. All chromatographic methods showed good linearity, precision and reproducibility. No spectral or chromatographic interference from the tablet excipients were found. The proposed methods were successfully applied to the assay of commercial tablets and content uniformity test. The procedures were rapid, simple and suitable for quality control application. PMID- 10701421 TI - Analytical and preparative supercritical fluid extraction of chamomile flowers and its comparison with conventional methods. AB - Supercritical fluid extraction (SFE) was compared with Soxhlet extraction, steam distillation and maceration for the isolation of the active components present in chamomile flowerheads. The obtained fractions were analysed by GC-MS and reversed phase HPLC. The yield of essential oil achieved by a 30-min extraction with pure CO2 at 90 atm and 40 degrees C was 4.4 times higher than that produced by steam distillation performed for 4 h. The recovery of the flavonoid apigenin obtained by supercritical CO2 after a 30-min extraction at 200 atm and 40 degrees C was 71.4% compared to Soxhlet extraction performed for 6 h and 124.6% compared to maceration performed for 3 days. However, the highly polar flavonoid apigenin-7 glucoside was not extracted by 100% CO2 (recovery values < 1.1%). Its extraction efficiency was markedly improved by the addition of the polar modifier methanol (5%, v/v) to the CO2 fluid, yet the obtained recoveries were unsatisfactory (14.6 19.5%). The SFE method was scaled-up for preparative applications using a pilot plant with three separation stages operating in series. Large-scale SFE was technically feasible with pure CO2 as the extracting fluid. However, the use of CO2 modified with organic solvents was not effective at the pilot-plant scale. PMID- 10701422 TI - Determination of remifentanil in human heparinised whole blood by tandem mass spectrometry with short-column separation. AB - For the use in pharmacokinetic studies, a fast and sensitive assay method was developed for the determination of remifentanil in human heparinised whole blood samples of 0.5 ml. The assay method is based on tandem mass spectrometry detection (LC-MS/MS). The limit of quantification is 0.1 ng/ml and linear up to 50 ng/ml. The precision, accuracy, recovery and applicability were found to be adequate for pharmacokinetic studies. PMID- 10701423 TI - Assay for the determination of low dosage form of formoterol dry syrup by capillary electrophoresis with head-column field-amplified sample stacking. AB - The development of a capillary zone electrophoresis method with head-column field amplified sample stacking injection for the determination of formoterol (FMTR) in a low dosage dry syrup form was described. To obtain the highest sensitivity, the sample solution was prepared by high content of organic solvent with the presence of a small amount of H+ (60-100 microM) and the capillary inlet end was dipped in water before electroinjection. This method was fully validated in terms of repeatability (RSDs for migration time, peak area of FMTR and peak area ratio between FMTR and I.S. at 1 microg/ml of FMTR was 0.76, 1.10 and 0.55% respectively), reproducibility (RSDs from different capillaries, analytes, days and instruments were 1.52%, 1.04%, 1.16% and 1.93% respectively), linearity (y = 0.827x - 0.085, r = 0.9993 (n = 6) over the range of 0.25-2.0 microg/ml), limits of quantitation, ruggedness and robustness. The method was applied to the determination of the drug in commercial dry syrup preparation (recovery was 100.9%, RSD = 1.5%, n = 5) and proved to be fast and reliable for the quantitation analysis of FMTR in the pharmaceutical form. PMID- 10701424 TI - Stability of meperidine in an implantable infusion pump using capillary gas chromatography-mass spectrometry and a deuterated internal standard. AB - A capillary gas chromatographic-mass spectrometric (GC MS) method is described for the analysis of meperidine using 3,3,5,5-[2H4]-meperidine as an internal standard. Chromatography was performed on a (5% phenyl) methylpolysiloxane column (30 m x 0.32 mm I.D., 0.25 microm film thickness) operated at 195 degrees C; helium carrier gas-50 cm/s(-1), tR = 2.3 min. Ionization was by electron impact (EI) and detection by selected ion monitoring of the molecular ions. The method provided high response linearity (mean r = 0.9982) and precision (< 6.5% C.V.). Application of this method to a pilot study of aqueous meperidine x HCl (10 mg/ml(-1)) stability in a surgically implantable infusion pump at 37 degrees C for 90 days revealed no demonstrable drug degradation. PMID- 10701425 TI - Fluorimetric determination of pipemidinic acid using the europium chelate. AB - The sensitized luminescence of europium ion in the complexes with pipemidinic acid is investigated. It was shown that in the result of intramolecular energy transfer from ligand to lanthanide ion the luminescence intensity of the latter increases by 10(10) times. The luminescence properties of the complex were studied and the high sensitive luminescence method for the determination of pipemidinic acid has been developed. PMID- 10701426 TI - Determination of melatonin in biological fluids in the presence of the melatonin agonist S 20098: comparison of immunological techniques and GC-MS methods. AB - Immunoassays were investigated for the determination of melatonin in biological samples in the presence of a naphthalenic structural analogue S 20098, which is currently under development as a melatonin agonist. The lack of specificity of commercially available antibodies in the presence of closely related molecules led us to develop an LC-RIA procedure with a quantification limit set at 15 pg/ml(-1). Because this technique was not sensitive enough and difficult to use on a routine basis, a more sensitive GC-MS technique was developed. This method involved automated solid-phase extraction (plasma) or liquid-liquid extraction (saliva), derivatization of the indolic moiety and GC separation with an automated switching device before MS detection. The method was validated over the range 1-100 pg/ml(-1), with a quantification limit set at 1 pg/ml(-1) in human plasma and saliva. Intra-assay and inter-assay precision and accuracy were within 16% for all concentrations investigated and each biological matrix. The stability of melatonin in plasma and saliva under various storage conditions was also determined. The specificity of the assay for the analysis of melatonin in the presence of S 20098 and its metabolises was demonstrated. The method was subsequently applied for the determination of endogenous melatonin concentrations in plasma and saliva samples from clinical studies performed with S 20098 to provide pharmacodynamic data. PMID- 10701427 TI - Development of flexible and efficient strategies for optimizing chromatographic separations. AB - The intensive research in chemometrics is resulting in continuous development of new concepts and optimization methods. The practical chromatographic optimization examples described in this paper highlight the importance of developing efficient and flexible optimization strategies, which are adapted to the (complex) separation problems encountered in the real chromatographic world. The availability of efficient and user-friendly software should contribute to a more systematic use of chemometrical approaches. Two primordial aspects are discussed in more details: (1) the selection of adequate optimization criteria and (2) the optimum robustness. PMID- 10701428 TI - An inductively coupled plasma method for determination of cyclophosphamide loaded to polymeric systems. AB - A new method for the determination of cyclophosphamide content of polyalkylcyanoacrylate nanoparticles was developed. The analyses were carried out by inductively coupled plasma atomic emission spectrometry (ICP-AES) by measuring the phosphorus content in the drug. The results obtained by this non-selective technique were compared with those given by high performance liquid chromatography (HPLC) a selective procedure that permits the detection of the cyclophosphamide molecule, and its degradation products. Sensitivity and reproducibility of both procedures were also determined. The ICP-AES method was demonstrated to be valid for sensitivity, precision, accuracy and specificity. In spite of ICP method is not a suitable procedure to analyze the degradation products of cyclophosphamide, the sensitivity of ICP is higher than chromatographic technique. Nevertheless, both procedures are appropriate for the determination of cyclophosphamide-loaded nanoparticles. PMID- 10701429 TI - Improved RPLC determination of acyclovir using hexylamine as silanol masking agent. AB - The aim of the present work is to improve the sensitivity in the RPLC determination of acyclovir [9-(2-hydroxy ethoxymethyl) guanine] (ACV) and guanine, the major impurity of the drug synthesis and one of the compounds found in the chemical degradation process of ACV. The method was applied to the quantification of drug in liposomal formulations. The most important problem for RPLC analysis of both compounds are their high pKa values, mainly guanine, and the interaction with reactive silanol groups in the stationary phase. In order to avoid these problems there are four basic strategies: (i) ionic pair reagents, (ii) deactivated silica columns, (iii) polymeric based columns and (iv) silanol masking agents. A validation protocol was followed to develop the analytical method, using a Spherisorb ODS (250 x 4.6 mm i.d.) analytical column, with a mobile phase of 95% aqueous phosphate buffer (pH 3.0) and 5% HPLC methanol pumped isocratically at 1.3 ml/min(-1), with ultraviolet detection at 254 nm. The results showed a high reproducibility in retention time value, with R.S.D. of 2.37% for ACV and 0.32% for guanine. The lowest concentration levels assayed, 0.15 microg/ml(-1) for guanine and 1 microg/ml(-1) for ACV, showed good R.S.D. in the quantification parameter (peak area) 11.0% (guanine) and 9.64% (ACV) PMID- 10701430 TI - Carbonyl metallo immuno assay: a new application for Fourier transform infrared spectroscopy. AB - We describe here the development of a new, non-isotopic immunological assay termed CMIA (carbonyl metallo immunoassay) that uses metal carbonyl complexes as tracers and Fourier transform infrared spectroscopy (FT-IR) as the detection method. This assay is based on the particular spectral features of these complexes, which show very strong absorption bands in the 1,800-2,200 cm(-1) spectral range where proteins and organic molecules do not absorb. In Section 1, the optimisation of the quantitative detection of these tracers is detailed. In Section 2, the implementation of mono-CMIA is described, including the CMIA assays of three antiepileptic drugs (carbamazepine, phenobarbital, phenytoin). Finally, extension to the simultaneous double- and triple-CMIA of these drugs is reported. PMID- 10701431 TI - Solid phase extraction of clenbuterol from plasma using immunoaffinity followed by HPLC. AB - An immuno-extraction column for clenbuterol has been prepared. Optimum conditions for the selective retention and elution of clenbuterol have been developed, based on a modification of our earlier work on morphine, chlortoluron and isoproturon. Clenbuterol could be retained on the immuno-column then eluted in one x one ml fraction using 50% methanol in phosphate buffered saline pH 2. On columns containing antisera (but not to clenbuterol) the clenbuterol was removed in the washing step. HPLC-UV determination gave clean traces. Day-to-day reproducibility was improved by precipitating the plasma proteins with acetonitrile. PMID- 10701432 TI - Gas chromatography-mass spectrometry assay method for the therapeutic drug monitoring of the antiepileptic drug tiagabine. AB - A gas chromatography-mass spectrometry assay method suitable for the therapeutic drug monitoring of the antiepileptic drug tiagabine is described. Tiagabine and its desmethylated analogue used as internal standard were first extracted from serum by liquid-liquid extraction using an ethyl ether-isobutanol 98:2 mixture. Tiagabine and the internal standard were then methylated in the organic phase in presence of methanol by means of a safe and stable diazomethane derivative. After evaporation, the reconstituted extracts were chromatographed on a crosslinked phenyl methyl siloxane capillary column and detected by mass fragmentometry at m/z = 156. No other antiepileptic drug possibly administrated in polytherapy and no metabolite were found to interfere in the assay. The limit of quantification was 5 ng/ml. The precision and the accuracy were found to be suitable for the therapeutic drug monitoring of tiagabine. PMID- 10701433 TI - An HPLC method for the determination of diastereomeric prodrug RS-79070-004 in human plasma. AB - Ganciclovir is an antiviral nucleoside analogue approved for treatment and prevention of cytomegalovirus infections in immunocompromised subjects. RS-79070 194, a diastereomeric monovalyl ester of ganciclovir (hydrochloride salt), is under evaluation as a prodrug to increase the bioavailability of ganciclovir. An HPLC method with column switching has been developed and validated for quantification of the corresponding free base RS-79070-004 in human plasma. In the method, proteinaceous material in 0.25 ml of plasma is precipitated by trichloroacetic acid. An aliquot of the supernatant is analyzed by HPLC, with automated column switching to remove late-eluting materials that might interfere with the analyte peaks in subsequent runs. Detection of RS-79070-004 is by UV lambda = 254 nm). The peak areas for each isomer are summed to generate a value for total RS-79070-004. The method has a validated range of 0.0400-4.00 microg/ml and a lower limit of quantification of 0.0400 microg/ml. All intra- and inter assay %CVs were < 7.5%, and all recoveries (accuracy) were within 6% of nominal values. No interference was observed by ganciclovir, caffeine, acetaminophen, or ibuprofen. Analyte stability in plasma and in the sample extracts is adequate for the specified collection, storage, and analysis conditions. The validated method has been successfully used to analyze clinical study samples. PMID- 10701434 TI - A rapid, sensitive HPLC method for the determination of ganciclovir in human plasma and serum. AB - A method for ganciclovir determination in human serum and plasma has been developed and validated. The method has a lower limit of quantification (LLOQ) adequate for sensitive pharmacokinetic studies ( < or = 0.05 microg/ml), has run times of < or = 15 min, and uses aliquot volumes adequate for pediatric studies (0.25 ml). In the method, proteinaceous material in serum or plasma is precipitated by trichloroacetic acid. An aliquot of the supernatant is analyzed by HPLC; automated column switching removes late-eluting materials that might interfere with the analyte peak in subsequent runs. Detection and quantification of ganciclovir is by fluorescence (lambda(ex) = 278 nm; lambda(em) = 380 nm). The method has a validated range of 0.0400-4.00 microg/ml and an LLOQ of 0.0400 microg/ml. All intra- and inter-assay % C.V. values were < 8%; all recoveries (accuracy) were within 7% of nominal values. No interference was observed by mycophenolic acid or its glucuronide metabolite, by AZT, salicylic acid, acetaminophen, ibuprofen, naproxen prednisone, acyclovir, or cyclosporine. Ganciclovir is very stable in the samples and the extract during storage and sample processing. Both serum and plasma methods have been validated for use and have been successfully used to analyze samples from clinical studies. PMID- 10701435 TI - Hyphenation of high performance liquid chromatography with nuclear magnetic resonance spectroscopy for the characterization of beta-carotene isomers employing a C30 stationary phase. AB - The hyphenation of HPLC together with NMR spectroscopy proves advantageous for the structure elucidation of oxidation- and UV-sensitive compounds such as beta carotene isomers. In the closed-loop HPLC-NMR system, degradation or isomerization of separated compounds is largely hindered. With the help of 3 microm C30 stationary phases a better separation efficiency towards the different beta-carotene cis/trans isomers could be obtained in comparison to a 5-microm material, resulting in sharper peaks and a better resolution of all compounds. This effect greatly facilitated the structure determination of the isomers by HPLC-NMR coupling. Due to the introduction of a superior stationary phase, the structure of seven cis-isomers of beta-carotene could thereby be determined employing the stopped-flow HPLC-1H-NMR mode. PMID- 10701436 TI - HPLC separation of 99mTc-L-cysteine acetyldiglycine diethylester and its monoester-monoacid and diacid derivatives. PMID- 10701437 TI - Quantitative determination of azathioprine in tablets by 1H NMR spectroscopy. PMID- 10701438 TI - Determination of lisinopril from pharmaceutical preparations by derivative UV spectrophotometry. PMID- 10701439 TI - Development of neurogenic placodes in Xenopus laevis. AB - The development of neurogenic placodes in Xenopus laevis from the time of neural fold closure to larval stages is described. Placodes were reconstructed from camera lucida drawings of serial sections, and the spatiotemporal pattern of placodal neurogenesis was analyzed using in situ hybridization for the genes X NGNR-1, XNeuroD, X-MyT1, and X-Delta-1, all of which have been implicated in the regulation of neurogenesis. Olfactory, profundal, and trigeminal placodes, a series of dorsolateral placodes (otic placode and five lateral line placodes), a series of epibranchial placodes, and two hypobranchial placodes were identified. Earlier claims that all placodes in anurans develop from a common primordium could not be confirmed. Profundal and trigeminal placodes, however, are partially fused, and all lateral line placodes arise from a common precursor. Epibranchial and hypobranchial placodes develop ventral to other placodes and dorsal and ventral to the pharyngeal pouches, respectively. Hypobranchial placodes give rise to neurons that become intimately associated with the developing heart. All neurogenic placodes strongly express the neuronal differentiation gene XNeuroD. The neuronal determination gene X-NGNR-1, however, is expressed strongly in only some placodes and not in dorsolateral placodes, indicating that neurogenesis in the latter relies on other determination genes. X-Delta-1 is expressed not only in the neurogenic parts of the placodes but also in the primordia of the lateral lines. This suggests that Delta-Notch-mediated lateral inhibition may be involved not only in placodal neurogenesis, but also in the patterning of lateral line neuromasts. PMID- 10701440 TI - PACAP and glutamate are co-stored in the retinohypothalamic tract. AB - The retinohypothalamic tract (RHT) relays photic information from the eyes to the suprachiasmatic nucleus (SCN). Activation of this pathway plays a role in adjusting circadian timing to the light/dark environment. Two transmitters, glutamate and pituitary adenylate cyclase activating polypeptide (PACAP) having phase shifting capacity during the night and day, respectively, are located in the RHT. Using double staining immunohistochemistry at the light and electron microscopic level, we showed that PACAP was co-stored with glutamate in a subset of retinal ganglion cells and in nerve terminals in the retino-recipient area of the SCN. These findings provide an anatomical basis for the recent demonstration of the interaction between these two transmitters on the SCN phase response at night. PMID- 10701441 TI - Relationship between neurokinin-1 receptor and substance P in the striatum: light and electron microscopic immunohistochemical study in the rat. AB - The synaptic relationship between substance P (SP) and its receptor, i.e., neurokinin-1 receptor (NK1R), was examined in the striatum of the rat by confocal laser-scanning microscopy and electron microscopy. For confocal laser-scanning microscopy, triple-immunofluorescence histochemistry was performed to label NK1R, SP, and vesicular acetylcholine transporter (a specific marker for cholinergic neurons). In electron microscopic double-immunolabeling study, immunoreactivity for NK1R was detected with the silver-intensified gold method, while immunoreactivity for SP was detected with peroxidase immunohistochemistry. Simultaneous immunolabeling of NK1R and SP revealed significant mismatch at the synaptic level: although some SP-immunopositive axon terminals were in synaptic contact with NK1R-immunopositive sites of plasma membrane, NK1R-immunoreactivity was observed at both synaptic and non-synaptic sites of plasma membrane. Thus, SP released from the sites remote from NK1Rs might diffuse in the extracellular fluid to act, as a paracrine neurotransmitter, on NK1Rs distant from its releasing site. SP neurotransmission in the striatum might occur not only synaptically but also extrasynaptically. The SP-NK1R system might constitute an association system within the striatum. PMID- 10701442 TI - Actin-associated protein synaptopodin in the rat hippocampal formation: localization in the spine neck and close association with the spine apparatus of principal neurons. AB - Dendritic spines are sites of synaptic plasticity in the brain and are capable of remodeling their shape and size. However, little is known about the cellular mechanisms that regulate spine morphology and motility. Synaptopodin is a recently described actin-associated protein found in renal podocytes and dendritic spines (Mundel et al. J Cell Biol. [1997] 139:193-204), which is believed to play a role in spine plasticity. The present study analyzed the distribution of synaptopodin in the hippocampal formation. In situ hybridization histochemistry revealed a high constitutive expression of synaptopodin mRNA in the principal cell layers. Light microscopic immunohistochemistry showed that the protein is distributed throughout the hippocampal formation in a region- and lamina-specific manner. Postembedding immunogold histochemistry demonstrated that synaptopodin is exclusively present in dendrites and spines, specifically in the spine neck in close association with the spine apparatus. Spines lacking a spine apparatus are not immunoreactive for synaptopodin. These data suggest that synaptopodin links the spine apparatus to actin and may thus be involved in the actin-based plasticity of spines. PMID- 10701443 TI - Maternal and mating-induced aggression is associated with elevated citrulline immunoreactivity in the paraventricular nucleus in prairie voles. AB - Lactating female rodents are fiercely aggressive against intruders when they are rearing and protecting pups. In monogamous prairie voles, Microtus ochrogaster, males are parental and exhibit a dramatic increase in aggression, termed mating induced aggression, in association with reproduction. In mice, the gas, nitric oxide (NO), inhibits male aggression, but may have an excitatory role in the production of maternal aggression. In this study, we combined aggressive behavioral testing of female and male prairie voles with immunohistochemistry for citrulline, a marker of NO synthesis, to examine NO synthesis indirectly during maternal and mating-induced aggression. A significant increase in the number of citrulline-positive cells was identified in the paraventricular nucleus (PVN) of the hypothalamus in aggressive lactating females compared with unstimulated lactating females. A significant increase in the number of citrulline-positive cells was also observed in the PVN of aggressive mated males compared with nonaggressive unmated males and unstimulated mated males. Both nonaggressive unmated males and unstimulated mated males show similar levels of citrulline immunoreactivity in the PVN. In other regions of the brain, no changes in the number of citrulline-positive cells were observed. These results suggest that NO is released specifically in the PVN during both maternal and mating-induced aggression in prairie voles. PMID- 10701444 TI - Retinal ganglion cells in the South American opossum (Didelphis aurita). AB - By using the Golgi technique, the authors investigated the morphology of ganglion cells in the retinas of South American opossums. In flat-mount preparations of the retinas, cell bodies, entire dendritic fields, and the stratification level of ganglion cells were studied. Fractal dimensions of dendritic trees, an objective quantitative measure of morphological complexity, were included as a morphological parameter of classification. Based on these characteristics, nineteen types of ganglion cells were described. A great number of opossum ganglion cell types had dendrites stratifying in both sublaminae of the inner plexiform layer (IPL) in five different ways (S1-S3 [G9], S1-S4 [G17 and G22], S2/S3 [G19], S2-S4 [G15, G16, G21 and G221, and S2-S5 [G61), and only two types (G8, and G10) showed narrow field dendritic trees ramifying in S4 only. Morphological types of opossum ganglion cells were compared to their counterparts in cat retina. The distribution pattern of large cell bodies on the ganglion cell layer was analyzed employing the Nissl staining method, immunocytochemistry for neurofilaments, and the reduced silver neurofibrillar staining method. The results showed a random pattern of distribution. PMID- 10701446 TI - Glutamate-like immunoreactivity marks compartments of the mushroom bodies in the brain of the cricket. AB - In the mushroom bodies of the brain of the cricket Gryllus bimaculatus, the distribution of glutamate-like immunoreactivity is shown by using several immunocytochemical staining protocols and confocal and conventional microscopy. Glutamate-like staining of intrinsic cells of mushroom bodies (Kenyon cells), their axons and projections, is demonstrated for the first time. Two types of Kenyon cells constituting distinct, separated populations within the perikaryal layer and in prominent neuropilar subcompartments exhibit strong (type III cells) or medium (type II cells) glutamate-like immunoreactivity, whereas the small neurons of a central population (type I cells) lack staining above background. Type III Kenyon cells display a strong immunoreactivity similarly found in some giant neurons and in identified antennal motorneurons by using glutamate as an excitatory transmitter, indicating that also distinct populations of the Kenyon cells use glutamate as a putative transmitter. The pattern of glutamate-like immunoreactivity in the mushroom bodies and in other parts of the brain is different from gamma-aminobutyric acid (GABA)-like immunoreactivity (investigated for comparison). GABA-like immunostaining is particularly prominent in the mushroom body calyces where Kenyon cells have their dendritic branchings. Differences in glutamate-like immunostaining in Kenyon cell subpopulations, together with differences in their arborization and axonal projection patterns, indicate a functional diversity of these neurons. PMID- 10701445 TI - Hearing loss and glutamate efflux in the perilymph following transient hindbrain ischemia in gerbils. AB - The mechanism underlying ischemia-induced hearing loss was studied in gerbils with transient hindbrain ischemia. Occlusion of the vertebral arteries caused an increase in the concentration of glutamate in the perilymph and elevated the compound action potential (CAP) threshold to 24.6 dB at 5 minutes. the CAP threshold subsequently recovered on reperfusion, gradually reaching 8.3 dB 120 minutes after reperfusion. Under electron microscopy, afferent dendrites of the cochlear nerve in contact with inner hair cells exhibited abnormal swelling 5 minutes after ischemia/reperfusion. These morphological changes were not observed in cochleas treated with an alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA)/kainate-type glutamate receptor antagonist, 6-7-dinitroquinoxaline 2,3-dione (DNQX), before hindbrain ischemia; an N-methyl-D-aspartate (NMDA)-type receptor antagonist, D-2-amino-5-phosphonopentanoate (D-AP5), was ineffective. Moreover, the histopathological alterations noted 5 minutes after reperfusion were spontaneously ameliorated 120 minutes after ischemia/reperfusion. These findings suggest that the ischemia-induced increase in extracellular glutamate concentration with subsequent activation of AMPA/kainate receptors is responsible for neurite degeneration and hearing loss in the early stages following transient hindbrain ischemia. PMID- 10701447 TI - Electrochemical modification of proteins. A review. PMID- 10701448 TI - Continuous production of N-(benzyloxycarbonyl)-L-glycyl-L-phenylalanine methyl ester utilizing extractive reaction in aqueous/organic biphasic medium. AB - N-(Benzyloxycarbonyl)-L-glycyl-L-phenylalanine methyl ester was continuously synthesized, enzymatically, utilizing an extractive reaction in an aqueous/organic biphasic system. The extremely high yield, ca. 100%, was obtained continuously in a water/butyl acetate biphasic medium. PMID- 10701449 TI - Investigation of expression of HOX 2C and HOX 4B homeobox genes in human colorectal cancer by using an RT-PCR method. AB - Investigation of expression of both HOX 2C and HOX 4B homeobox genes in the same patient with colorectal cancer was proposed by using an RT-PCR method. In order to conduct this investigation, PCR products of 445 bp of HOX 2C and 301 bp of HOX 4B were amplified in both tumor and normal samples of ten patients. Expressions of HOX 2C gene were observed in both tumor and normal samples of four patients and in only a tumor sample of one patient, while the expression was not observed in both tumor and normal samples of five patients. Expressions of HOX 4B gene were not observed in both tumor and normal samples of ten patients. In the present study, it was found that individuality seems to be important. The results of these two genes, observed in patients with colorectal cancer, should be taken into consideration for further researches. PMID- 10701450 TI - Novel process for enzymatic hydrolysis of proteins in an aqueous two-phase system for the production of peptide mixture. AB - A novel process for the production of peptide mixtures is proposed. Biologically active peptides were synthesized using a thermolysin-catalyzed hydrolysis of a corn protein (zein) in an aqueous two-phase system. The mixture of peptides which was selectively recovered from the dextran-rich bottom phase had a higher angiotensin-converting enzyme (ACE) inhibitory activity than native zein. PMID- 10701451 TI - Synthesis and analgesic activity of N-(N-acetyl-L-amino-acyl)-5 methoxytryptamines. AB - 5-Methoxytryptamine was acylated with N-acetyl-L-amino acids to give rise the corresponding N-(N-acetyl-L-amino acyl)-5-methoxytryptamines. The analgesic activity was evaluated by the tail flick test. Among the 6 compounds, the analgesic potency of N-(N-acetyl-tryptophanyl)-5-methoxytryptamine (5e) and N-(N acetyl-glycyl)-5-methoxytryptamine (5a) are much more potent than that of melatonin. PMID- 10701452 TI - Purification and characterization of carbonic anhydrase from bovine erythrocyte plasma membrane. AB - Carbonic anhydrase (CA) was purified from bovine erythrocyte plasma membrane and characterized in this study. For this purpose, the blood taken from young animals was hemolysed, the membrane fraction was separated, and this fraction was repeatedly washed. The enzyme (CA) was removed from the membrane with buffered TritonX-100 (1%); it could be purified at a factor of 22.8 by affinity chromatography. The CA obtained from erythrocyte membrane has an esterase activity as well as hydratase activity. The Vmax and Km of the enzyme for the substrate (p-nitrophenyl acetate) are 1.948x10(-3) mM/L x dak, and 3.596 mM, respectively. The purification degree of the enzyme was controlled by SDS-PAGE (3 10), which showed two distinct bands. It was determined that the enzyme had activity within the pH range of 4.5-9.5 and that the optimal pH was 7.5. The temperature at which it showed activity was 20-60 degrees C and optimal temperature was 37 degrees C. Molecular weight of CA was found to be 29844 and 61706 Dalton by gel filtration. On the other hand, sulfanilamide and acetazolamide affected the enzyme. PMID- 10701453 TI - Purification of the ADP-ribosylating enzyme from S. solfataricus by SDS polyacrylamide gel electrophoresis and electroelution. AB - The ADPribosylating enzyme from the thermophilic archaeon S. solfataricus was purified by a simple procedure which included preparative electrophoresis on a 0.1% SDS- polyacrylamide gel. The gel slice containing the enzymatic protein was cut out and the enzyme was solubilized by electroelution. The pure enzyme was obtained by chromatography of the electroeluted sample on a DNA-Sepharose column. The purified enzyme retained both its full activity and the structuring ability as a function of temperature increase. PMID- 10701454 TI - Preliminary study on the cleavage of fusion protein GST-CMIV with palladium(II) complex. AB - A novel method for post-treatment of gene-engineered proteins is reported. A coden of Cys-His unit is introduced into the N-terminal of cecropin CMIV by using PCR. The gene is expressed in E. coli fused with GST. After purification, the fusion protein is cleaved by [Pd(en)(H2O)2]2+ at the His-Arg bond and the cecropin CMIV with antibacterial activity is obtained. The preliminary results held some promise of success for application of the palladium(II) complex as cleavage agent for the production of peptide drugs from gene-engineering fusion proteins. PMID- 10701455 TI - Titanium dioxide ceramics control the differentiated phenotype of cardiac muscle cells in culture. AB - A new approach, the cultivation of heart muscle cells on biocompatible scaffolds made from titanium dioxide ceramics was established to provide a mechanism for in vitro engineering of a vital heart tissue. Terminally differentiated ventricular myocytes isolated from hearts of adult rats were kept in primary culture for long periods of time and used as an experimental model. The microenvironmental properties of titanium dioxide ceramics helped to maintain the tissue-like structural organisation of the cardiac cells in vitro. Coating of the cell substrata with fine-grained titanium dioxide ceramics imitating cell surface topography favoured the formation of focal adhesion complexes in the ventral plasma membrane of cardiomyocytes. It also promoted the cellular expression of vinculin, a protein that connects the ECM integrin receptors to the network of cytoplasmic filaments, which define cell shape. This topographical reinforcement of cell-material interactions led to stabilisation of the molecular linkage between the extracellular contacts and the intracellular cytoskeleton and thus assisted the preservation and maintenance of the heart muscle cell differentiated phenotype in long-term primary culture. The results of this work demonstrate a promising pathway for the regulation of cellular organisation in vitro by local geometric control. PMID- 10701456 TI - Phagocytosis of wear debris by osteoblasts affects differentiation and local factor production in a manner dependent on particle composition. AB - Wear debris is considered to be one of the main factors responsible for aseptic loosening of orthopaedic endoprostheses. Whereas the response of cells in the monocytic lineage to foreign materials has been extensively studied, little is known about cells at the bone formation site. In the present study, we examined the hypothesis that the response of osteoblasts to wear debris depends on the chemical composition of the particles. We produced particles from commercially pure titanium (cpTi), Ti-6Al-4V (Ti-A), and cobalt-chrome (CoCr) and obtained ultrahigh molecular weight polyethylene (UHMWPE; GUR 4150) particles from a commercial source. The equivalent circle diameters of the particles were comparable: 1.0 +/- 0.96 microm for UHMWPE; 0.84 +/- 0.12 microm for cpTi; 1.35 +/- 0.09 microm for Ti-A, and 1.21 +/- 0.16 microm for CoCr. Confluent primary human osteoblasts and MG63 osteoblast-like cells were incubated in the presence of particles for 24 h. Harvested cultures were examined by transmission electron microscopy to determine if the cells had phagocytosed the particles. Particles were found intracellularly, primarily in the cytosol, in both the primary osteoblasts and MG63 cells. The chemical composition of the particles inside the cells was confirmed by energy-dispersive X-ray analysis. Morphologically, both cell types had extensive ruffled cell membranes, less-developed endoplasmic reticulum, swollen mitochondria, and vacuolic inclusions compared with untreated cells. CpTi, Ti-A, and CoCr particles were also added to cultures of MG63 cells to assess their effect on proliferation (cell number) and differentiation (alkaline phosphatase activity), and PGE2 production. All three types of particles had effects on the cells. The effect on cell number was dependent on the chemical composition of the particles; Ti-A and CoCr caused a dose-dependent increase, while cpTi particles had a biphasic effect with a maximal increase in cell number observed at the 1:10 dilution. Alkaline phosphatase specific activity was also affected and cpTi was more inhibitory than Ti-A or CoCr. PGE2 production was increased by all particles, but the magnitude of the effect was particle dependent: CoCr > cpTi > Ti-A. This study demonstrates clearly that human osteoblast-like cells and MG63 cells can phagocytose small UHMWPE, CoCr, Ti-A, and cpTi particles. Phagocytosis of the particles is correlated with changes in morphology, and analysis of MG63 response shows that cell proliferation, differentiation, and prostanoid production are affected. This may have negative effects on bone formation adjacent to an orthopaedic implant and may initiate or contribute to the cellular events that cause aseptic loosening by inhibiting bone formation. The effects on alkaline phosphatase and PGE2 release are dependent on the chemical composition of the particles, suggesting that both the type and concentration of wear debris at an implant site may be important in determining clinical outcome. PMID- 10701457 TI - Influence of glass composition on the properties of glass polyalkenoate cements. Part III: influence of fluorite content. AB - The influence of fluorite content of the glass on the formation and properties of glass polyalkenoate cements was investigated. A series of glass powders based on 1.5SiO2 x 0.5P2O5 x Al2O3 x CaO x XCaF2 were synthesised. The glass transition temperature of the glass fell with increasing fluorite content. Setting and working times of the cement pastes decreased with increasing fluorite content of the glass. Compressive strength and un-notched fracture strength increased with increasing fluorite content of the glass. Fracture toughness and toughness of the cements were relatively insensitive to fluorite content. PMID- 10701458 TI - Tissue-engineered cartilage using serially passaged articular chondrocytes. Chondrocytes in alginate, combined in vivo with a synthetic (E210) or biologic biodegradable carrier (DBM). AB - In vitro multiplication of isolated autologous chondrocytes is required to obtain an adequate number of cells to generate neo-cartilage, but is known to induce cell-dedifferentiation. The aim of this study was to investigate whether multiplied chondrocytes can be used to generate neo-cartilage in vivo. Adult bovine articular chondrocytes, of various differentiation stages, were suspended in alginate at densities of 10 or 50 million/ml, either directly after isolation (P0) or after multiplication in monolayer for one (P1) or three passages (P3). Alginate with cells was seeded in demineralized bovine bone matrix (DBM) or a fleece of polylactic/polyglycolic acid (E210) and implanted in nude mice for 8 weeks. The newly formed tissue was evaluated by Alcian Blue and immunohistochemical staining for collagen type-II and type-I. Structural homogeneity of the tissue, composed of freshly isolated as well as serially passaged cells, was found to be enhanced by high-density seeding (50 million/ml) and the use of E210 as a carrier. The percentage of collagen type-II positive staining P3-cells was generally higher when E210 was used as a carrier. Furthermore, seeding P3-chondrocytes at the highest density (50 million/ml) enhanced collagen type-II expression. This study shows promising possibilities to generate structurally regular neo-cartilage using multiplied chondrocytes in alginate in combination with a fleece of polylactic/polyglycolic acid. PMID- 10701459 TI - Development of tailor-made collagen-glycosaminoglycan matrices: EDC/NHS crosslinking, and ultrastructural aspects. AB - The many biocharacteristics of glycosaminoglycans (GAGs) make them valuable molecules to be incorporated in collagenous biomaterials. To prepare tailor-made collagen-GAG matrices with a well-defined biodegradability and (bioavailable) GAG content, the crosslinking conditions have to be controlled. Additionally, the ultrastructural location of GAGs in engineered substrates should resemble that of the application site. Using chondroitin sulfate (CS) as a model GAG, these aspects were evaluated. The methodology was then applied for other GAGs. CS was covalently attached to collagen using 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS). A maximum of about 155 mg CS/g matrix could be immobilized. CS incorporation and bioavailability, as evaluated by interaction with specific antibodies and glycosidases, was dependent on the molar ratio EDC:carboxylic groups of CS. The denaturation temperature could be modulated from 61 to 85 degrees C. The general applicability of EDC/NHS for immobilizing GAGs was demonstrated with dermatan sulfate, heparin, and heparan sulfate. These matrices revealed comparable physico-chemical characteristics, biodegradabilities, and preserved bioavailable GAG moieties. At the ultrastructural level, GAGs appeared as discrete, electron-dense filaments, each filament representing a single GAG molecule. Distribution was independent of GAG type. They were observed throughout the matrix fibers and at the outer sites, and located, either parallel or orthogonally, at the periphery of individual collagen fibrils. Compositional and ultrastructural similarity between matrices and tissue structures like cartilage and basement membranes can be realized after attachment of specific GAG types. It is concluded that EDC/NHS is generally applicable for attachment of GAGs to collagen. Modulation of crosslinking conditions provides matrices with well-defined GAG contents, and biodegradabilities. Ultrastructural similarities between artificially engineered scaffolds and their possible application site may favor the use of specific collagen-GAG matrices in tissue engineering. PMID- 10701460 TI - Biodegradable three-dimensional networks of poly(dimethylamino ethyl methacrylate). Synthesis, characterization and in vitro studies of structural degradation and cytotoxicity. AB - In ophthalmology, there is a need for novel degradable biomaterials for e.g. controlled drug release in the vitreous body. These degradable materials should feature both excellent biocompatibility, and well-defined kinetics of degradation. In most cases, poly(D,L-lactic acid), or poly(lactic-co-glycolic acid) are used. These materials, however, suffer from some serious drawbacks, since the degradation kinetics are difficult to control, especially since the so called 'burst-degradation' occurs. Here, we describe a set of novel polymeric networks which largely consist of poly(dimethylamino ethyl methacrylate) (poly(DMAEMA)); these materials are crosslinked via a dimethacrylate molecule that contains two carbonate groups. This system is susceptible to hydrolytic scission. The degradation products do not exert a catalytic effect on the ongoing degradation reaction (i.e. there is no burst effect). We describe the synthesis of three of these materials, which differ merely with regard to the crosslinker content. These materials were characterized through DMTA, 1H NMR and FT-IR spectroscopy, and scanning electron microscopy. The reaction DMAEMA + 2 hydroxyethyl methacrylate (HEMA) was studied in detail, using 1H NMR spectroscopy, and these experiments revealed that the reaction of DMAEMA and HEMA produces a random (Bernouillian-type) copolymer. From this, we contend that the new materials have more or less uniform distribution of the crosslinks throughout their volume. Structural degradation of the three materials was studied in vitro, at pH 7.4, 9.1 and 12.0. It is found that the materials exhibit smooth hydrolysis, which can be controlled via the crosslink density and the pH, as was expected a priori. It should be noted that degradation of these materials produces non-hydrolysable, but water-soluble, oligo(DMAEMA) and poly(DMAEMA) molecules. We subsequently performed in vitro studies on the biocompatibility of these materials. The MTT cytotoxicity assay revealed that the materials were cytotoxic to chondrosarcoma cells. This is most probably due to local increase of the pH due to the basic character of the pending dimethylamino groups. Cytotoxicity remained virtually unchanged after extended washing with water. This indicates that the cytotoxicity is an intrinsic property of the material and was not caused by remnants of free monomer. Cytotoxicity was also seen in cell cultures (human fibroblasts isolated from donor corneas) which were grown in contact with the materials. It is concluded that the new materials have attractive degradation characteristics, but their cytotoxicity makes them unsuitable for applications in ophthalmology. PMID- 10701462 TI - Preparation and characterisation of monoclinic hydroxyapatite and its precipitated carbonate apatite intermediate. AB - Five 100 g batches of a carbonate apatite (the intermediate) were produced by heating an aqueous slurry of CaCO3 and CaHPO4 with an overall Ca/P mole ratio of 5/3 with vigorous stirring. Each intermediate produced by boiling off water was heated in vacuum at 1100 degrees C to remove carbonate, then steamed at 900 degrees C to ensure complete hydroxylation. Comparison of calculated and observed X-ray diffraction patterns showed final products containing 50-100 wt% monoclinic hydroxyapatite (remainder hexagonal). Rietveld refinements in P6(3)/m gave structures similar to several hydroxyapatite standards, including NIST SRM 2910, although there was no evidence from X-ray diffraction that the latter was in the monoclinic form. Refinements from standards and final products were slightly different from published single crystal data for Holly Springs hydroxyapatite. This is attributed to known impurities in mineral hydroxyapatite and indicates that parameters from the Rietveld refinements are closer to the true values for pure hydroxyapatite. Rietveld refinements for intermediates showed small, but significant differences from the final product, the largest being in O1x, O2x and O(H)z. All P-O bond lengths were shorter than in the final product, resulting in a 3.2% lower PO4 tetrahedron volume. The occupancies of P and Ca(2) were reduced. These differences are attributed to partial replacement of PO4(3) by CO3(2-) ions. PMID- 10701461 TI - Surface modification using silanated poly(ethylene glycol)s. AB - Surface-grafted poly(ethylene glycol) (PEG) molecules are known to prevent protein adsorption to the surface. The protein-repulsive property of PEG molecules are maximized by covalent grafting. We have synthesized silanated monomethoxy-PEG (m-PEG) for covalent grafting of PEG to surfaces with oxide layers. Two different trialkoxysilylated PEGs were synthesized and characterized. The first trialkoxysilylated PEG was prepared by direct coupling of m-PEG with 3 isocyanatopropyltriethoxysilane through a urethane bond (silanated PEG I). The other silanated PEG (silanated PEG II) containing a long hydrophobic domain between PEG and a silane domain was prepared by reacting m-PEG with 1,6 diisocyanatohexane and 10-undecen-1-ol in sequence before silylation with 3 mercaptopropyl trimethoxysilane. Silanated PEGs I and II were grafted onto glass, a model surface used in our study. The PEG-grafted glass surfaces were characterized by contact angle, X-ray photoelectron spectroscopy (XPS), and atomic force microscopy (AFM). Although contact angle did not change much as the bulk concentration of silanated PEG used for grafting increased from 0.1 to 20 mg/ml for both PEGs I and II, the surface atomic concentrations from XPS measurements showed successful PEG grafting. Surface PEG grafting increased concentration of surface carbon but decreased silicone concentration. The high resolution C1s spectra showed higher ether carbon with lower hydrocarbon compositions for the PEG-grafted surfaces compared to the control surface. AFM images showed that more PEG molecules were grafted onto the surface as the bulk concentration used for grafting was increased. AFM images of the dried surfaces showed that the surfaces were not completely covered by PEG molecules. After hydration, however, the surface appears to be covered completely probably due to the hydration of the grafted PEG chains. Glass surfaces modified with silanated PEGs reduced fibrinogen adsorption by more than 95% as compared with the control surface. Silanated PEGs provides a simple method for PEG grafting to the surface containing oxide layers. PMID- 10701463 TI - Microgrooved silicone subcutaneous implants in guinea pigs. AB - Cell-substratum interactions are of fundamental importance for the reaction of body tissues to surgically implanted foreign materials. In our study we investigated the influence of 2 microm wide microgrooves, with various depths (0.5-6 microm), on capsule formation around subcutaneous silicone implants, in an animal experiment. Silicone sheets with microtexture were glued around silicone tubes. These implants were placed subcutaneously in eight guinea pigs for 10 weeks. The implanted tubes were removed including all surrounding tissues, and processed for light microscopy and subsequent histomorphometrical evaluation. All removed implants were surrounded by a thin fibrous capsule, and it was observed that this capsule was separated from the implants by a thin, single layer of mono and multinucleated phagocytotic cells. In histomorphometry no significant differences were seen in relation to the reaction towards the various textures. We conclude that microtextures do not have an effect on the morphological characteristics of capsule formation around silicone implants in soft tissue. PMID- 10701464 TI - Treatment of tooth fracture by medium energy CO2 laser and DP-bioactive glass paste: compositional, structural, and phase changes of DP-bioglass paste after irradiation by CO2 laser. AB - Nowadays, fractured teeth are difficult to treat effectively. Currently, root fractures are usually treated by root amputation, hemisection or tooth extraction. If the fusion of tooth fracture by laser were possible, it would offer a different therapy to repair fracture teeth. We tried to use a developed DP-bioactive glass paste to fuse or bridge the tooth crack line by a medium energy continuous-wave CO2 laser. The study is divided into three parts: (1) The compositional and structure changes in tooth enamel and dentin after laser treatment; (2) The phase transformation and recrystallization of DP-bioactive paste during exposure to the CO2 laser; (3) The thermal interactions and bridge mechanism between DP-bioactive glass paste and enamel/dentin when they are subjected to CO2 laser. The present report will focus on the second part that will examine the changes of laser-exposed DP-bioactive glass paste by means of X ray diffractometer (XRD), Fourier transforming infrared spectroscopy (FTIR), differential thermal analysis/thermogravimetric analysis (DTA/TGA), and scanning electron microscopy (SEM). From the study, we could find that the temperature increase due to laser irradiation is greater than 900 degrees C and that the DP bioactive glass paste could be melted in a short period of time after irradiation. In the study, we successfully developed a DP-bioactive glass paste which could form a melting glass within seconds after exposure to a medium energy density continuous-wave CO2 laser. The paste will be used in the near future to bridge the enamel or dentin surface crack by the continuous-wave CO2 laser. PMID- 10701465 TI - Anxiety disorders: a conceptual history. PMID- 10701466 TI - IDS-C and IDS-sr: psychometric properties in depressed in-patients. AB - Sixty-eight depressed in-patients were assessed at admission (DO), and after 5 days (D5), ten days (D10) and 28 days (D28) of antidepressant treatment, with the Inventory for Depressive Symptomatology-Clinician (IDS-C) and the Inventory for Depressive Symptomatology-Self-Rated (IDS-SR) (Rush et al., 1986), the Montgomery and Asberg Depression Rating Scale (MADRS) (Montgomery and Asberg, 1979) and the depression factor of the Symptom Check List (SCL-90R) (Derogatis, 1977), in order to assess IDS-C and IDS-SR psychometric properties in depressed in-patients and to compare IDS-C to MADRS and IDS-SR to the SCL-90R depression factor. Most of the IDS-C and IDS-SR items were significantly correlated to the final score and the Cronbach alpha coefficients were high (0.75 for the IDS-C and 0.79 for the IDS-SR). Principal Component Analyses (PCA) showed three factors for both IDS-C and IDS-SR: 'depression', 'anxiety/arousal' and 'sleep/appetite'. These results suggest satisfactory internal consistency of IDS-C and IDS-SR. Concurrent validity of the IDS-C with the MADRS was high (r = 0.81), as well as concurrent validity of the IDS-SR with the SCL-90R depression factor (r = 0.84). Concerning sensitivity to change, the four scales were able to discriminate between different levels of severity of depression. Moreover, considering paired t-tests on score changes, IDS-C sensitivity to change may be higher than MADRS sensitivity to change, this phenomenon being related to the number of items and degrees but not to the item contents. Contrary to IDS-C and MADRS, IDS-SR and SCL 90R depression factor were not different in terms of sensitivity to change. Finally, psychometric properties of IDS-C and IDS-SR in depressed in-patients are satisfactory and close to those obtained in depressed out-patients. The high sensitivity to change of the IDS-C may be an advantage for this scale as compared to the MADRS, especially in antidepressant drug trials. PMID- 10701467 TI - Temperament in bipolar illness: impact on prognosis. AB - OBJECTIVE: The present study was designed to investigate the relations between temperament and outcome in bipolar illness. METHODS: Seventy-two patients presenting with bipolar type I disorder were recruited from consecutive admissions and evaluated when euthymic. The criteria developed by Akiskal and Mallya (Criteria for the 'soft' bipolar spectrum: treatment implications. Psychopharmacol. Bull. 1987;23:68-73) were used to assess both depressive (DT) and hyperthymic temperaments (HT) in a dimensional approach. RESULTS: Multiple regression analysis showed that a higher DT score or a lower HT score were significantly associated with a greater number of episodes. Furthermore, a higher DT score was strongly associated with a higher percentage of major depressive episodes. Conversely, a higher HT score was associated with a trend to manic rather than depressive episodes. Suicide attempts appeared more frequent in the history of patients presenting with higher DT scores. CONCLUSIONS: Our findings strengthen the hypothesis that temperament is one of the main variables accounting for some features in the clinical evolution of bipolar disorder such as polarity of episodes. Furthermore, these findings are consistent with the hypothesis of a trait-state continuum between personality and affective episodes. PMID- 10701468 TI - Efficacy and safety of tianeptine in the treatment of depressive disorders in comparison with fluoxetine. AB - BACKGROUND: Depression is treated by a great variety of antidepressant treatments. SSRIs (such as fluoxetine) are well known: it is, however, sure that further progress is needed and the search for antidepressants with other mechanisms of action (such as tianeptine) or different efficacy is still of interest. METHODS: A multinational study compared tianeptine with fluoxetine in 387 patients with Depressive Episode, or Recurrent Depressive Disorder, or Bipolar Affective Disorder (ICD-10), in a double-blind parallel group design. They were treated for six weeks. RESULTS: At inclusion, no significant difference between groups was shown. Final MADRS scores were 15.7 and 15.8 with tianeptine and fluoxetine, respectively (ITT population) (p = 0.944). MADRS responders were 58% and 56% with tianeptine and fluoxetine, respectively (p = 0.710). No statistical difference was observed for the other efficacy parameters. Thirty-six withdrawals occurred in each group, without any difference for the reasons of discontinuation. There was no major difference between groups for the other safety parameters. CONCLUSIONS: In this study, both tianeptine and fluoxetine exhibited a good efficacy and safety. PMID- 10701469 TI - Reduced tolerance and cardiovascular response to ischemic pain in minor depression. AB - BACKGROUND: A paradoxical association between a higher prevalence of clinical pain and a reduced sensitivity to brief experimental pain seems to exist during depression. METHODS: We assessed the responses to sustained ischemic pain produced by a maximal effort tourniquet procedure in 32 controls and 11 individuals with minor depression (Zung autoscale > or =50). Stethoscopic blood pressures and heart rates were monitored throughout the procedure. RESULTS: Measures of pain threshold, and measures of pain intensity and pain unpleasantness during the ischemic procedure were similar in depressed and control subjects. Yet, the overall numerical ratings of ischemic pain during the procedure was 28% higher and pain tolerance was 44% lower in depressed compared to control subjects. Clinical pain complaints were reported by 91% of depressed but only by 41% of control subjects (P = 0.01). Sustained ischemic pain induced significant elevations of systolic and mean arterial blood pressures in controls but not in depressed subjects. LIMITATIONS: The main limitation of the present study was the preponderance of females in the depressed group. Yet, we did not find significant gender differences in the sensory-affective and autonomic responses to ischemic pain in our sample. CONCLUSIONS: These findings suggest alterations in the sensory and autonomic nervous systems during minor depression. PMID- 10701470 TI - Gender differences in the short-term course of unipolar depression in a follow-up sample of depressed inpatients. AB - OBJECTIVE: This paper examined sex differences in the short-term course of depression and assessed the impact of possibly outcome-affecting factors, including sex-specific recall artefacts and demographic and clinical characteristics. METHODS: A cohort of 179 unipolar depressed inpatients was followed up 1 (T1) and 7 months (T2) after discharge. RESULTS: Residual depression at T1 was comparable in both sexes as was the rate of follow-up nonremissions in patients who had failed to remit from the index episode at T1. In contrast, female gender was a significant predictor of relapse. This sex difference was partly attributable to women who relapsed after T1 and were again in remission at T2. Potential sex-related recall artefacts were tested by contrasting the patients' retrospective assessment of their T1-depression status reported at T2 with their interviewer-rated depression status assessed at T1. Results suggest that the observed sex difference in relapses could neither be explained by memory artefacts nor by differences in demographic and clinical sample compositions. CONCLUSIONS: It is concluded that due to their higher risk for early relapses, particular efforts with regard to continuation treatment are required for women during the critical period of remission. PMID- 10701471 TI - Reduced adenylyl cyclase immunolabeling and activity in postmortem temporal cortex of depressed suicide victims. AB - BACKGROUND: Previous studies have found altered receptor/G protein-modulated adenylyl cyclase (AC) activity in subjects with mood disorders. METHODS: To investigate whether these effects are associated with altered levels of specific isoforms of AC, we measured AC isoform I, IV and V/VI immunoreactivities in postmortem temporal cortex from nine depressed suicide victims, nine subjects with bipolar disorder (BD) and 18 age-matched non-psychiatric controls. Basal, GTPgammaS- and forskolin-stimulated AC activities were measured in the temporal cortex from the nine depressed suicide victims and their controls. RESULTS: Western blotting revealed significant reductions in immunolabeling in AC type IV (-49%; p < 0.05) in depressed suicide subjects compared to age-matched controls, but no differences were found in AC type I or type V/VI. There were no statistically significant differences in AC type I, IV or V/VI immunoreactivities between BD and matched control subjects. Functionally, there was a significant reduction in forskolin-stimulated AC activity in depressed suicide subjects compared to controls, which may be, in part, related to higher basal AC activity in the former group. LIMITATIONS: Our sample size was small with diverse subject characteristics. CONCLUSIONS: These preliminary findings suggest altered levels and/or function in AC type IV may contribute to disturbances in the postreceptor cAMP signaling cascade in depression. PMID- 10701472 TI - Temperament and character inventory dimensions as a predictor of response to antidepressant treatment in major depression. AB - BACKGROUND: Cloninger's theory of personality, including 4 temperament dimensions and 3 character dimensions, is one of the most noteworthy theories in recent years. Several studies have explored temperament dimensions as a predictor of response to antidepressant treatments in major depression, but these have provided inconsistent results. The present study explored temperament as well as character dimensions, as measured by the Temperament and Character Inventory (TCI), as possible predictors of response to maprotiline, the most-widely prescribed antidepressant in Japan. METHODS: 86 consecutive patients with major depression underwent a 16-week open trial of maprotiline. They filled out the TCI at baseline, and were followed up at weeks 8 and 16 by using the Hamilton Rating Scale for Depression. RESULTS: Hierarchial logistic regression analyses demonstrated that response to maprotiline was significantly predicted by the cooperativeness score at the 8-week outcome assessment, and by the self directedness score at the 16-week outcome assessment, after controlling the possible effects of clinical variables on the response. There was no evidence that either temperament dimensions or their 2-way interactions significantly predicted the response. LIMITATIONS: Large replication studies with other antidepressants are needed for generalizing the results in this study. CONCLUSIONS: The results in this study regarding temperament dimensions seem consistent with findings in previous studies, which are, as a whole, inconsistent with each other. It is suggested that character dimensions (particularly cooperativeness and self-directedness), rather than temperament dimensions, may be important predictors of response to antidepressants. Antidepressants may differ in the personality configurations that predict optimal responses. PMID- 10701473 TI - Winter and summer outdoor light exposure in women with and without seasonal affective disorder. AB - BACKGROUND: The annual decrease of daylight duration initiates a depressive phase in patients with seasonal affective disorder (SAD), and light therapy treats it. How much bright light exposure in winter and summer these patients actually receive may help understand the pathogenetic factors initiating SAD. METHODS: During a week in winter and summer, women with and without SAD kept daily logs of the time spent outdoors, subjective sleep, and self-ratings of mood and alertness. RESULTS: Compared with the winter depressive state, mood, alertness, and sleep of SAD patients improved in summer to control values, but did not correlate with the amount of light exposure. In summer, patients with SAD spent more time outdoors than controls. LIMITATION: Light logs--in comparison with light monitor measurements--may overestimate light exposure outdoors. CONCLUSION: Women with SAD do not spend less time outdoors in winter than controls, but spend more time outdoors in summer. CLINICAL RELEVANCE: Patients with SAD show a high amplitude seasonal difference in outdoor light exposure. The susceptibility to winter depression may arise not from behaviourally-related lack of sufficient light exposure, but an increased vulnerability to the amount of light received. They may require more light than controls to remain euthymic (higher light exposure in summer, light therapy in winter). PMID- 10701474 TI - A double-blind, randomized, placebo-controlled trial of once-daily venlafaxine extended release (XR) and fluoxetine for the treatment of depression. AB - BACKGROUND: We compared the efficacy and tolerability of venlafaxine XR with that of fluoxetine in a multicenter, randomized, double-blind, placebo-controlled study in depressed outpatients. METHODS: Outpatients, 18 years and older, who met DSM-IV criteria for major depressive disorder were included (n = 301 randomized; 232 completed). Patients were randomly assigned to eight weeks of treatment with either venlafaxine XR 75-225 mg/day (n = 100), fluoxetine 20-60 mg/day (n = 103), or placebo (n = 98). The primary efficacy outcome measures were the final ratings on the Hamilton Rating Scale for Depression (HAM-D21) total score, HAM-D21 depressed mood item, Montgomery-Asberg Depression Rating Scale total score, and Clinical Global Impressions Scale. RESULTS: Withdrawal from the study due to adverse events occurred in 6% of the patients in the venlafaxine XR group and 9% of the patients in the fluoxetine group. Patients treated with venlafaxine XR, but only rarely those treated with fluoxetine, had statistically significant improvements in their depression ratings compared with placebo at the end of the study. The percentages of patients who achieved full remission of their depression (HAM-D21 total score < or = 7) at the end of treatment were 37%, 22% and 18% for the venlafaxine XR, fluoxetine and placebo groups, respectively. The differences in remission rates between venlafaxine XR and the other groups were statistically significant (p < 0.05). LIMITATIONS: The superior remission outcome observed with venlafaxine XR treatment needs to be replicated in additional studies. CONCLUSION: Venlafaxine XR is a well-tolerated and efficacious treatment for depression. The results of this study suggest that venlafaxine XR is as well tolerated as fluoxetine but may have some efficacy advantages over fluoxetine. PMID- 10701475 TI - Retrospective controlled study of inpatient ECT: does it prevent suicide? AB - BACKGROUND: This study examined the use of ECT among inpatients who committed suicide at a provincial psychiatric hospital. METHODS: A total of 45 psychiatric in-patients who committed suicide at a provincial psychiatric hospital were compared with a gender, age and admission diagnosis matched group of 45 hospitalized patients to examine the use of electroconvulsive therapy during the last 3 months of hospitalization. RESULTS: No difference in the utilization of ECT was found in the two groups. LIMITATIONS: Retrospective design and small sample size. CONCLUSIONS: We failed to demonstrate that ECT had prevented suicide in hospitalized patients. Future prospective studies with large sample size are needed to further examine this question. PMID- 10701476 TI - Lower serum zinc in major depression in relation to changes in serum acute phase proteins. AB - There is now some evidence that major depression is accompanied by activation of the inflammatory response system (IRS). Other signs of IRS activation, which have been reported in major depression are lowered serum zinc (Zn) and serum albumin (Alb) concentrations. In serum, Zn is closely bound to Alb. The aims of the present study were to replicate previous findings that major depression is accompanied by lowered serum Zn and Alb and to examine whether the decrease in serum Zn may be explained by that in serum Alb. The above variables were determined in 48 major depressed patients and in 15 age-sex-matched healthy volunteers. Serum Zn and Alb were significantly lower in major depressed patients than in normal volunteers. In healthy volunteers and major depressed patients, there were significant and positive correlations between serum Zn and Alb. We found that 53.8% of the variance in serum Zn could be explained by the combined effects of serum Alb and diagnostic classification. The results suggest that lower serum Zn in depression is in part explained by lowered serum Alb and by another depression-related mechanism. It is suggested that lower serum Zn in depression may be secondary to sequestration of metallothionein in the liver, which may be related to increased production of interleukin-6. PMID- 10701477 TI - Free thyroxine and thyroid-stimulating hormone levels in patients with seasonal affective disorder and matched controls. AB - Seasonal affective disorder (SAD) is characterized by recurrent episodes of depression in the fall and winter that alternate with nondepressed periods in the spring and summer. Because some symptoms of SAD, such as decreased energy and weight gain, also occur in hypothyroidism, it is possible that individuals with SAD have a subtle decrease in thyroid function. To test this hypothesis, we studied blood levels of free thyroxine (T4) and thyroid-stimulating hormone (TSH) in SAD patients and matched controls in the winter. We found that free T4 blood levels were slightly but significantly lower in patients than in healthy volunteers. The difference between TSH levels in SAD patients and controls was not statistically significant. Future research will be needed to determine whether the difference in thyroid function between SAD patients and controls is an epiphenomenon or is related to the biological mechanisms that cause symptoms of SAD. PMID- 10701478 TI - Sexual dysfunction before antidepressant therapy in major depression. AB - BACKGROUND: Decreased sexual interest and function both occur as a consequence of antidepressant medication use, and are especially associated with serotonin reuptake inhibitors (SRIs). However, few investigators have reported the base rate for disturbances in sexual desire, arousal and orgasm or ejaculation in patients with major depression (MD) prior to antidepressant treatment. The purpose of this report is to define the frequency of sexual dysfunction (SD) in 134 patients with MD and examine the relationship between SD and demographic, clinical and personality variables. METHOD: A consecutive series of 55 male and 79 female MD patients diagnosed by SCID-DSM IV assessment completed a series of psychometric measures including a Sexual Function Questionnaire, which asked about change in sexual interest and function as well as sexual activity during the preceding month. RESULTS: Only 50% of women and 75% of men reported sexual activity during the preceding month. Over 40% of men and 50% of women reported decreased sexual interest. Reduced levels of arousal were more common in both men and women (40-50%) than ejaculatory or orgasm difficulties (15-20%). In women, problems with arousal and orgasm correlated with higher neuroticism and lower extraversion. There was no relationship between SD and personality measures in men. While age at onset of depression and number of prior episodes showed a modest correlation with SD measures, there were no correlations with severity of depression or specific symptoms clusters. LIMITATIONS AND CONCLUSIONS: Although limited by a relatively small sample of drug free patients with MD, and by the absence of a non-depressed comparison sample, these results emphasize the importance of factors beyond specific drug effects in the assessment of antidepressant related sexual dysfunction. PMID- 10701479 TI - Timing of movements in depressed patients and healthy controls. AB - BACKGROUND: Psychomotor disturbances are fundamental psychopathological features of major depression and observable components of behaviour. Human behaviour is segmented into action units with duration of a few seconds due to central nervous motor processing. Timing may depend on cognitive and emotional functions which are affected in depression. Therefore, time structure of action units in depressed patients was compared to healthy controls. METHODS: Included were patients with major depression and melancholic features. Upper limb movements (total n = 566) of depressed patients and matched controls were evaluated using videotaped interviews and frame-by-frame analysis with a temporal resolution of 40 ms. RESULTS: Behaviour of depressed patients in interview sessions was organised in action units with a narrow time span of only a few seconds. Single, non-repetitive action units were significantly shorter (median = 1.20 s) and repetitive units longer (median = 4.92 s) in patients compared to controls (median = 2.08 and 2.96, respectively). LIMITATIONS: Behaviour in interview sessions might differ from activities of daily living. DISCUSSION: Altered temporal segmentation of movements appears to be an observable, measurable sign of melancholic depression and may allow further insights in pathophysiological dysfunctions of the disease. Clinical implications of these motor changes for differential diagnosis, course and treatment of depression are discussed and need further evaluation. PMID- 10701480 TI - Childhood trauma and depression in alcoholics: relationship to hostility. AB - OBJECTIVE: To examine for a relationship between childhood trauma and depression in alcoholics. METHODS: Euthymic depressed alcoholics (N = 23) were compared with never depressed alcoholics (N = 20) for their scores on the Childhood Trauma Questionnaire (CTQ). Subjects also completed the Hostility and Direction of Hostility Questionnaire (HDHQ). RESULTS: Euthymic depressed alcoholics had significantly higher scores on the CTQ for childhood emotional abuse, physical abuse, sexual abuse, and emotional neglect. They also had significantly higher hostility scores on the HDHQ. There were significant correlations between adult hostility scores and CTQ scores for childhood emotional neglect, physical neglect, sexual abuse and total childhood trauma. CONCLUSION: A history of childhood trauma was correlated with adult depression in male alcoholics: a hostile personality dimension might be a mediating variable. LIMITATION: Subjects were queried on their memories of childhood traumas. Prospective studies are needed. PMID- 10701481 TI - The potential role of haloperidol in the treatment of trichotillomania. AB - BACKGROUND: Trichotillomania is categorized as an impulse control disorder in DSM IV and is considered by some to be closely related to Obsessive Compulsive Disorder (OCD). We review the clinical phenomenology and pharmacological response of trichotillomania, and suggest that it may be more related to Tourette Syndrome than to OCD. Serotonin reuptake inhibitors (SRIs) are typically employed in the treatment of OCD, while neuroleptic medications such as haloperidol are typically used in the treatment of Tourette Syndrome. Evidence for the efficacy of treatment of trichotillomania with drugs typically used for OCD is equivocal. METHOD: Nine patients with trichotillomania were treated with haloperidol. Six patients unresponsive to SSRI medication had haloperidol added to their treatment. Three patients received only haloperidol. Response to treatment was assessed using descriptions of hair pulling, quantity of hair pulled, and severity of depilation at hair pulling sites. RESULTS: Eight of nine patients responded to haloperidol treatment, with seven experiencing complete or near complete cessation of hair pulling. LIMITATIONS: Inferences from the results of this study are limited by the lack of a control group, the small sample size, and the use of unstandardized ratings as measures of symptom severity. CONCLUSIONS: Results suggest that the addition of haloperidol to SSRIs or haloperidol alone may be effective in the treatment of trichotillomania. Results also encourage speculation about the relation between OCD, Tourette Syndrome, and trichotillomania. PMID- 10701482 TI - Open trial of fluoxetine in children and adolescents with dysthymic disorder or double depression. AB - BACKGROUND: Chronic depressions commonly present in youth and cause significant morbidity. No treatment studies in this age group are currently available. METHODS: 19 pediatric subjects with dysthymic disorder or 'double depression' were recruited. After four weeks of psychosocial treatment, subjects failing to improve began open treatment with fluoxetine (20 mg) for eight weeks. Subjects were then reassessed for treatment response. RESULTS: Fifteen subjects entered the medication phase, and eleven (73%) no longer met criteria for dysthymic disorder or Major Depression at final assessment. CONCLUSIONS: Fluoxetine shows promise as a safe and effective treatment for youth with chronic depressions. Controlled trials are indicated. LIMITATIONS: Open label design, no comparison treatment condition. CLINICAL RELEVANCE: As in adults, treatment with antidepressants may prove to be a useful intervention with children and adolescents with chronic forms of depression. PMID- 10701483 TI - A double-blind study comparing idazoxan and bupropion in bipolar depressed patients. AB - BACKGROUND: There is a small body of evidence indicating that idazoxan, a potent and selective alpha-2 antagonist, may be effective in treating bipolar depressive disorder. The purpose of this prospective controlled study is to compare idazoxan to bupropion, an antidepressant which has been suggested to have some advantages over other antidepressants in treating bipolar depressed patients. METHODS: Bipolar I depressed patients were randomly assigned in this 6-week double-blind out-patient study to receive either idazoxan, titrated to 240 mg/day and placebo bupropion, or bupropion, titrated to 450 mg/day and placebo idazoxan. These doses were achieved after 2 weeks. Depression severity was assessed with the Hamilton Depression Rating Scale and possible psychosis with the Brief Psychiatric Rating Scale. Side effects, heart rate, weight, and orthostatic blood pressure were also monitored. RESULTS: Fourteen patients completed this study (seven in each group). Both idazoxan and bupropion demonstrated significant improvement over time with reductions in Hamilton scores of 50%. LIMITATIONS: Limitations of this study include lack of a placebo group and small sample size. CONCLUSION: In light of our preliminary findings suggesting the usefulness of idazoxan in bipolar depression, larger more rigorous studies are indicated. PMID- 10701485 TI - Immunoisolation of cells and tissues for transplantation. PMID- 10701484 TI - Highlights of the cell transplantation meeting at Montreux, March 1999. PMID- 10701486 TI - Encapsulated CNTF-producing cells for Huntington's disease. PMID- 10701487 TI - Xenografting human T2 sympathetic ganglion from hyperhidrotic patients provides short-term restoration of catecholaminergic functions in hemiparkinsonian athymic rats. AB - Previous studies have suggested that allografting peripheral sympathetic ganglia, such as superior cervical ganglia, partially relieves clinical or behavioral deficits in parkinsonian patients and animals. However, removal of these ganglia can cause Homer's syndrome, which limits the utilization of this approach. Hyperhidrosis, a disease of excessive sweating, is commonly seen in young Orientals. Treatment of hyperhidrosis often involves surgical removal of the second thoracic sympathetic ganglia (T2G), which contain catecholaminergic neurons. The purpose of our study was to investigate behavioral responses and tyrosine hydroxylase (TH) immunoreactivity in hemiparkinsonian rats at different time points after transplantation of human T2G from hyperhidrotic patients. Athymic Fisher 344 rats were injected unilaterally with 6-hydroxydopamine into the medial forebrain bundle to destroy the nigrostriatal dopaminergic (DA) pathway. The effectiveness of lesions was tested by measuring methamphetamine (MA)-induced rotations. These unilaterally lesioned rats were later transplanted with T2G or T2 fiber tract (T2F) obtained from adult hyperhidrotic patients. Animals grafted with T2G showed a reduction in MA-induced rotation by 2 weeks; however, rotation returned to the pregrafting levels by 3 months. Animals receiving T2F grafts did not show any reduction of rotation over a 3-month period. Animals were later sacrificed for TH immunostaining at different time points. Tyrosine hydroxylase-positive [TH(+)] cell bodies and fibers were found in the lesioned striatum 2-4 weeks after T2G grafting, suggesting the survival of transplants. Two to 3 months after grafting, TH(+) fibers were still found in almost all the recipients. However, TH(+) cell bodies were found in only three of seven rats studied. Animals receiving T2F grafting did not show any TH immunoreactivity in the lesioned striatum over the 3-month period. These data indicate that T2G transplants from adult hyperhidrotic patients can survive and provide transient normalization of the motor behavior in the hemiparkinsonian athymic rats. Because of the short-term improvement in behavior after grafting, the use of T2G in human trials should be cautious at the present time. Further laboratory research is required. PMID- 10701488 TI - Fetal porcine dopaminergic cell survival in vitro and its relationship to embryonic age. AB - One of the critical factors in the survival of embryonic neural grafts is the age at which the population of donor neurons is harvested. This is especially the case for the developing dopaminergic neurons in the embryonic ventral mesencephalon, which are used for neural grafts in Parkinson's disease (PD). The donor age for optimal harvesting of these cells has been well characterized in the mouse, rat, and marmoset, and to a lesser extent in humans. However, the best donor age for porcine ventral mesencephalic tissue has not been ascertained, even though the use of this tissue for xenografts has been explored both experimentally and clinically. In this study the effect of donor age on dopaminergic cell survival was assessed in vitro, from a range of fetal pigs aged from E24 to E35. The number of tyrosine hydroxylase (TH)-positive cells per ventral mesencephalon was then calculated after 1 and 7 days in culture. E26-E27 embryos gave the highest yield of such cells at both survival time points, suggesting that this will be the optimal age for harvesting tissues whether for experimental or clinical nigral xenograft programs. PMID- 10701489 TI - Porcine embryonic brain cell cytotoxicity mediated by human natural killer cells. AB - Intracerebral transplantation of porcine embryonic dopamine-producing neurons has been suggested as a method to treat patients with Parkinson's disease. Even though the brain is an immunologically privileged site, neuronal xenografts are usually rejected within a few weeks. T cells are important for this process, but the exact cellular events leading to rejection are poorly characterized. Brain cells from ventral mesencephalon of 26-27-day-old pig embryos were used as target cells in flow cytometry-assessed cytotoxicity assays using non- and IL-2 activated CD3- CD16+ CD56+ human natural killer (NK) cells as effector cells. The ability of human NK cells to kill pig embryonic brain cells by antibody-dependent cellular cytotoxicity (ADCC) in the presence of nondepleted and anti-Gal alpha1,3Gal antibody-depleted human blood group AB serum (AB serum) was evaluated using the same assay. Both nondepleted and anti-Gal alpha1,3Gal antibody-depleted AB serum could mediate ADCC of pig embryonic VM cells when human NK cells were used as effector cells. Nonactivated NK cells did not show any direct cytotoxic effect on freshly isolated VM cells, whereas IL-2-activated NK cells killed approximately 50% of the VM cells at an effector-to-target ratio of 50:1 in a 4-h cytotoxicity assay. Activation of VM cells by TNF-alpha did not change their sensitivity to human NK cell cytotoxicity. Human NK cells may thus contribute to a cellular rejection of pig neuronal xenografts by ADCC, or following IL-2 activation, by a direct cytotoxic effect. PMID- 10701490 TI - Viability of periosteal tissue obtained postmortem. AB - Periosteal autografts have the potential to regenerate articular cartilage defects, but this potential is limited by the patient's age. Allograft transplantation from a young donor to an older recipient might bypass this limitation. The effect of the time delay, between death and harvesting of a periosteal graft, on the chondrogenic potential of periosteum is important not only for transplantation but also for studies dealing with tissues retrieved postmortem (i.e., including the periosteal explant model). The purpose of this study was to investigate the chondrogenic potential of periosteum obtained postmortem and a possible beneficial effect of hypothermia. Thirty NZ white rabbits (2 months old) were sacrificed and stored at room temperature or 4 degrees C for 0, 4, 6, 8, 12, 16, 18, or 24 h. Periosteal explants were then obtained and a standard cartilage yield assay performed by culturing them for 6 weeks using the periosteal organ culture model as previous published. TGF-beta1 (10 ng/ml) was added for the first 14 days of culture. Histochemical analysis and quantitative collagen typing were performed. In the explants from the animals kept for 4 h at room temperature growth and chondrogenesis were dramatically reduced. Little or no chondrogenesis was seen in explants from rabbits maintained at room temperature after 4-8 h (or more) postmortem. Cooling the rabbits to 4 degrees C partially prevented this loss of viability and continued to do so for 24 h. Even storage at 4 degrees C did not eliminate the decrease in chondrogenic potential, though it did permit partial preservation of chondrogenic potential. If periosteum is to be used for allograft transplantation, or if it is used for experimental study, its viability must be assured. This is best accomplished by harvesting it immediately postmortem. Preservation techniques, cryopreservation, or hypothermia might be useful in preserving periosteal chondrogenic potential. PMID- 10701491 TI - Subcutaneous transplantation of bovine and human adrenocortical cells in collagen gel in scid mice. AB - Adrenocortical cells of bovine origin and of adult and fetal human origin were transplanted subcutaneously (s.c.) in scid mice after being embedded in collagen gel. In this site the cells survived, became vascularized by invasion of host endothelial cells, and secreted steroids into the circulation. The animals' own adrenal glands were removed at the time of cell transplantation. Steroids secreted by the transplants replaced the essential functions of the animals' own adrenal glands. Adrenalectomized animals without transplanted cells died after several days, but most animals with transplanted bovine or adult human adrenocortical cells survived; fewer animals survived with transplanted fetal human adrenocortical cells. The histology of the tissues formed from transplanted cells resembled that of the normal adrenal cortex. A few proliferating cells were observed in tissue from bovine or adult human cells; there was a greater percentage of dividing cells in tissue derived from fetal cells. Subcutaneous transplantation of bovine or human primary adrenocortical cells in collagen provides a model for the study of the physiology, cell biology, and molecular biology of adrenocortical cells in a three-dimensional vascularized tissue structure in a host animal. PMID- 10701492 TI - Immunosuppression and rejection of cartilage formed by allogeneic chondrocytes in rats. AB - Rat syngeneic and allogeneic chondrocytes were transplanted intramuscularly or into defects prepared in articular cartilage (intracartilaginous transplants). Recipients of allogeneic transplants received cyclosporin A (CsA), cladribine (2 chlorodeoxyadenosine, 2-CdA), or both drugs in combination. Transplants were taken for examination after 5 weeks. Cartilage formed intramuscularly by syngeneic chondrocytes was ossified. Allogeneic cartilage was resorbed by infiltrating cells. CsA or 2-CdA partially suppressed, and both these agents in combination strongly suppressed, formation of infiltrations. Both syngeneic and allogeneic chondrocytes formed cartilage in joint surface defects but only allogeneic cartilage was attacked by infiltrating cells. CsA + 2-CdA treatment slightly decreased intensity of infiltrations but did not prevent cartilage resorption. Antichondrocyte response was studied by evaluation of spleen mononuclear cells (SMC) stimulation in mixed splenocyte-chondrocyte cultures and by detection of antichondrocyte cytotoxic antibodies. SMC stimulation index (SI) was calculated separately for syngeneic and allogeneic chondrocytes. Comparison of SMC SI for syngeneic and allogeneic chondrocytes indicated lack of stimulation of SMC from control or syngeneic transplant recipients and significant stimulation of SMC from recipients of allogeneic transplants. SMC from animals treated with CsA + 2-CdA were not stimulated. Additional experiments aiming at an explanation of the lack of stimulation of SMC from intact animals by syngeneic chondrocytes reported in this work and contrary to other findings disclosed that it was caused by the use of collagenase solution containing N alpha-p-tosyl-l lysine chloromethyl ketone for chondrocyte isolation. Spontaneous antichondrocyte cytotoxic antibody activity was found in intact rats raised only in sera from recipients of allogeneic intramuscular transplants without immunosuppression. Thus, strong immunosuppressive treatment of rats with allogeneic chondrocyte transplants was more effective in relation to the general immunological response than to the local reaction. PMID- 10701493 TI - Pretreatment with glucocorticoids enhances T-cell effector function: possible implication for immune rebound accompanying glucocorticoid withdrawal. AB - Glucocorticoids (GCs) exert their immunosuppressive/antiproliferative effects largely through inhibition of cytokine expression, and paradoxically upregulate the expression of (proinflammatory) cytokine receptors on select nonlymphoid cells. Clinically, withdrawal of GCs was frequently associated with worsening of the outcome of heightened immunity disorders, thereby implicating enhanced cytokine and cytokine receptor expression as a possible consequence of acute/short-term GCs withdrawal. In view of the significance of this complication of GC therapy, we addressed the effect of GC withdrawal on cytokine receptor expression and subsequent T-cell effector function, using the proliferation of human T cells as biological readout. To mimic GC withdrawal, T cells were treated with GCs or controls, stimulated, and incubated for 16-20 h at 37 degrees C, washed, and reactivated for a further 4-48 h. Surface marker expression was assessed by FACS analysis, and proliferation was determined by measuring the cellular uptake of tritiated thymidine. Dexamethasone (DEX) and prednisolone (PRED), in a concentration-dependent manner, inhibited T-cell proliferation induced by anti-CD28 Ab + PMA. However, pretreatment of T cells activated with mitogens, cross-linking antibodies, or PMA + ionomycin ("CD3-bypass" stimulation regimen), but not resting T cells, with DEX or PRED resulted in a marked increase in IL-IR, IL-2R alpha, and IL-6R expression, which was accompanied by a significant enhancement in T-cell proliferation. This effect of GCs was neither stimulus specific nor did it result from alteration in cell viability, and was paralleled by augmentation in cytokine (rIL-2) effects on DEX-pretreated and preactivated T cells. Taken together, our results underline the dual effects of GCs in regulating T-cell activation and cytokine expression. In essence, GCs directly inhibited T-cell proliferation by suppressing cytokine production, and, by enhancing cytokine receptor expression, pretreatment with GCs augmented T-cell proliferation. PMID- 10701494 TI - Xenogeneic transplantation of porcine hepatocytes into the CCl4 cirrhotic rat model. AB - Liver support using extracorporeal devices and hepatocyte transplantation has received renewed interest for the management of acute and chronic liver failure. The aim of this study was to determine whether xenogeneic porcine hepatocytes could integrate into the liver parenchyma of cirrhotic Lewis rats when administered by an intrasplenic route. Cirrhosis was induced by carbon tetrachloride (CCl4) inhalation and confirmed histologically. Freshly isolated porcine hepatocytes were infused directly into the splenic pulp at laparotomy over a 5-15-min interval. Using (111)In-labeled hepatocytes, the degree of localization of porcine hepatocytes to the spleen and liver was found to be greater than 60% in both control and cirrhotic rats. Integration of porcine hepatocytes into the rat liver parenchyma was determined by immunohistochemical staining for porcine albumin in rat liver sections. Further confirmation was provided by in situ hybridization using a porcine-specific probe that binds to a distinct repetitive element (PRE) in porcine DNA. Evidence of integrated porcine hepatocytes was seen for over 50 days in animals under cyclosporine immunosuppression. These data demonstrate the integration of xenogeneic porcine hepatocytes into the liver of the cirrhotic rat and their ability to produce porcine albumin for up to 50 days. PMID- 10701495 TI - Efficient gene transfer and expression in islets by an adenoviral vector that lacks all viral genes. AB - Although adenoviral vector-mediated gene transfer has significant potential for gene therapy, host immune responses to virally expressed proteins and small insert capacity may limit its clinical application. In order to overcome these disadvantages, a new adenoviral vector that lacks all viral genes has been developed. Using the green fluorescent (GFP) gene as a reporter gene, we investigated the efficiency of gene transfer by this all-viral-genes-deleted and minimal cis-element remaining adenoviral vector (miniAd-GFP) in islets in vitro and ex vivo, and compared it with the E1-deleted adenoviral vector (E1-GFP). One day after in vitro infection, GFP was expressed in both miniAd-GFP- and E1-GFP infected islets. The percentage of GFP-positive single cells was not significantly different between miniAd-GFP-infected islets and E1-GFP-infected islets. When these islets were transplanted into syngeneic diabetic mice, both miniAd-GFP- and E1-GFP-infected islet grafts reversed diabetes, and normal blood glucose levels were maintained for over 20 weeks posttransplantation. Mild lymphocyte infiltration was found in all E1-GFP-infected islet grafts at all time points. However, this was not seen in most miniAd-GFP-infected islet grafts. Our results indicate that gene transfer by an adenoviral vector that lacks all viral genes is as efficient as E1-deleted adenoviral vector-mediated gene transfer in islets. Furthermore, this adenoviral vector might be less immunogeneic than the E1-deleted adenoviral vector. PMID- 10701496 TI - Differentiation and expansion of beta cell mass in porcine neonatal pancreatic cell clusters transplanted into nude mice. AB - Neonatal porcine pancreas has considerable capacity for growth and differentiation, making it an attractive potential source of islet tissue for xenotransplantation. Pancreases from 1-3-day-old newborn pigs were digested with collagenase and cultured for 8 days. The resulting cellular aggregates are called porcine neonatal pancreatic cell clusters (NPCCs). The mean yield of NPCCs from a newborn pig was 28,200 +/- 1700 islet equivalents. Cytokeratin 7 (CK7) was used as a marker for the immunostaining of pancreatic duct cells. In neonatal pancreas, 18% of the insulin-positive cells co-stained for CK7, thus being protodifferentiated. NPCCs also contained protodifferentiated cells; insulin/PP and insulin/somatostatin co-stained cells were more common than insulin/glucagon cells. Between 1 and 8 days of culture, the DNA content of the NPCCs fell to 16% and the insulin content to 33% of the starting value, mainly due to the preferential loss of exocrine cells. Transplantation of 2000 or 4000 NPCCs into diabetic nude mice typically normalized glucose values in 10-20 weeks. Mice with successful grafts had lower fasting blood glucose levels than normal mice and accelerated glucose clearance after an i.p. glucose load. The starting NPCCs consisted of 17% insulin-staining cells, but the grafts of mice with reversed diabetes consisted of 94% beta cells, with some co-stained for CK7, indicating that the grafts still contained immature cells. The mass of insulin-producing cells rose from 0.22 +/- 0.08 mg 1 week after transplantation to 4.34 +/- 0.27 mg in mice sacrificed at 27-35 weeks. In summary, NPCCs contain mostly islet precursor cells, which when transplanted into nude mice undergo striking differentiation and beta cell expansion. PMID- 10701497 TI - Secretion from islets and single islet cells following cryopreservation. AB - The ability to cryopreserve pancreatic islets has allowed the development of low temperature banks that permit pooling of islets from multiple donors and allows time for sterility and viability testing. However, previous studies have shown that during cryopreservation and thawing there is a loss of islet mass and a reduction in islet function. The aim of this study was to measure and compare insulin secretion from cultured nonfrozen and frozen-thawed canine islets and beta-cells. Canine islets were isolated from mongrel dogs using intraductal collagenase distention, mechanical dissociation, and EuroFicoll purification. One group of purified islets was cultured overnight before dissociation into single cells and subsequent analysis. Remaining islets were cultured overnight (22 degrees C) and then cryopreserved in 2 M dimethyl sulfoxide (DMSO) solution using a slow stepwise addition protocol with slow cooling to -40 degrees C before storage in liquid nitrogen (-196 degrees C). Frozen islets were rapidly thawed (200 degrees C/min) and the DMSO removed using a sucrose dilution. From a series of seven consecutive canine islet isolations, islet recovery following postcryopreservation tissue culture was 81.5 +/- 4.8% compared to precryopreservation counts. In vitro islet function was equivalent between cultured nonfrozen and frozen-thawed islets with a calculated stimulation index of 10.4 +/- 1.5 (mean +/- SEM) for the frozen-thawed islets, compared with 12.4 +/- 1.2 for the cultured nonfrozen controls (p = ns, n = 7 paired experiments). Amperometric detection of secretion from single beta-cells in vitro has the sensitivity and temporal resolution to detect single exocytotic events and allows secretion to be monitored from single beta-cells in real time. Secretion from single beta-cells elicited by chemical stimulation was detected using a carbon fiber microelectrode. The frequency of exocytosis events was equivalent between the cultured nonfrozen and frozen-thawed beta-cells with an average of 7.0 +/- 1.32 events per stimulation for the cultured nonfrozen group compared with 6.0 +/ 1.45 events from the frozen then thawed preparations (minimum of 10 cells per run per paired experiment, p = ns) following stimulation with tolbutamide. The average amount of insulin released per individual exocytosis event was equivalent for the cultured nonfrozen and frozen-thawed islets. In addition, beta-cells responded to both tolbutamide and muscarinic stimulation following cryopreservation. It was determined that beta-cells recovered following cryopreservation are capable of secreting insulin at levels and frequencies comparable to those of cultured nonfrozen islet preparations. PMID- 10701498 TI - The effects of microencapsulation on pancreatic islet osmotically induced volumetric response. AB - Microencapsulation of pancreatic islets has been proposed as a means to prevent allograft rejection and to protect islets during cryopreservation. The aim of this study was to investigate: 1) the effects of the cryoprotectants (CPAs) dimethyl sulfoxide (DMSO) and ethylene glycol (EG) on the volume of Ca2+ alginate microcapsules, and 2) the effects of microencapsulation on the volumetric response of human and canine pancreatic islets during CPA equilibration. Stock sodium alginate with a high mannuronic acid content (HM) or a high guluronic acid content (HG) was used to generate empty capsules (mean diameter 200 microm) with an electrostatic generator. The capsules were held in place by a holding pipette system and videotaped during the addition of 2 or 3 M CPA at 22 degrees C. Islets (isolated from human cadaveric donors and mongrel dogs and then cultured overnight at 37 degrees C) were encapsulated in alginate (HM), loaded into a microperfusion chamber, and the change in islet volume was videotaped after exposure to the same CPAs and concentrations. These were compared to the volume responses of nonencapsulated islets. Images were analyzed using a computerized image analysis system and the data were analyzed using ANOVA. HG microcapsules showed a significant (p < 0.05) increase in volume following exposure to EG but not to DMSO. HM microcapsule volume did not change significantly following exposure to either EG or DMSO and was therefore chosen as the substrate for islet encapsulation. Free, nonencapsulated canine and human islets responded to the osmotic challenge of the 2 M DMSO by shrinking to 70.00 +/- 1.04% (mean +/- SEM) and 70.11 +/- 1.05%, and in 2 M EG to 72.89 +/- 1.93% and 69.33 +/- 1.38%, respectively, of the isotonic volume before returning to the original cell volume. Exposure to 3 M DMSO or EG resulted in a further dehydration to 65.89 +/- 0.91% and 67.67 +/- 1.91% for canine and 62.22 +/- 0.66.% or 65.89 +/- 1.30% for human islets. Minimum volumes were reached within 30-40 s after exposure to the cryoprotectant. Encapsulated human islets reached 86.88 +/- 1.47% of their original volume in 2 M and 80.33 +/- 0.89% in 3 M DMSO, and 87.33 +/- 1.86% in 2 M and 82.80 +/- 1.57% in 3 M EG. This volume change was significantly less (p < 0.01) than that observed in corresponding free islets. Encapsulated canine islets reached 83.67 +/- 2.13% of their original volume in 2 M and 78.22 +/- 0.95% in 3 M DMSO, and 85.44 +/- 1.92% in 2 M and 78.11 +/- 2.01% in 3 M EG. As with human islets, this was significantly different than free islets (p < 0.01). These minimal volumes were reached within 30-50 s. These results demonstrate that there are cryoprotectant and alginate-specific interactions and that microencapsulation modulates the degree of osmotically induced shrinkage of islets. The development or modification of existing cryopreservation protocols to improve postcryopreservation recovery or function must account for these factors. PMID- 10701499 TI - Lessons from in vitro perifusion of pancreatic islets isolated from 80 human pancreases. AB - We report the average insulin response to acute glucose measured by in vitro perifusion of pancreatic islets isolated from 80 consecutive human organs. Different perifusion parameters were considered [basal release, stimulation index (SI), time to peak, incremental area under the curve delta-AUC alpha)], and the correlation among them was determined. SI positively correlated with delta-AUC alpha (p < 0.001, r = 0.80) while negatively with time to peak (p < 0.05, r = 0.23). We also evaluated several variables of the isolation procedure that might affect responsiveness to glucose by human islets. Sex and age of pancreas donors, cold ischemia time, duration of the digestion, collagenase concentration, and lot characteristics (collagenase, trypsin, clostripain, and proteases activity), and final islet yield were considered. Multivariate regression analysis showed only an independent association between SI and the concentration of collagenase (p = 0.01). PMID- 10701501 TI - A quantitative approach to the dielectric properties of the skin. AB - The results of measurements using an open-ended coaxial probe of the audio/radiofrequency dielectric properties of human skin in vivo, either dry or moistened with physiological saline, are reported. Permittivity and conductivity dispersion curves were parametrized by using a newly reported dispersion function (Raicu V 1999 Dielectric properties of biological matter: model combining Debye type and 'universal' responses Phys. Rev. E 60 4677), and the results obtained are discussed in the light of the recent advances in this field. It is suggested that the coaxial probe reports on the properties of the superficial layer, the stratum corneum, when the skin surface is dry, whilst the signal from deeper skin layers becomes dominant after wetting the skin with conductive physiological saline. PMID- 10701500 TI - Prospects for in vivo Raman spectroscopy. AB - Raman spectroscopy is a potentially important clinical tool for real-time diagnosis of disease and in situ evaluation of living tissue. The purpose of this article is to review the biological and physical basis of Raman spectroscopy of tissue, to assess the current status of the field and to explore future directions. The principles of Raman spectroscopy and the molecular level information it provides are explained. An overview of the evolution of Raman spectroscopic techniques in biology and medicine, from early investigations using visible laser excitation to present-day technology based on near-infrared laser excitation and charge-coupled device array detection, is presented. State-of-the art Raman spectrometer systems for research laboratory and clinical settings are described. Modern methods of multivariate spectral analysis for extracting diagnostic, chemical and morphological information are reviewed. Several in-depth applications are presented to illustrate the methods of collecting, processing and analysing data, as well as the range of medical applications under study. Finally, the issues to be addressed in implementing Raman spectroscopy in various clinical applications, as well as some long-term directions for future study, are discussed. PMID- 10701502 TI - Electron fluence perturbation correction factors for solid state detectors irradiated in megavoltage electron beams. AB - The perturbation correction factor gamma(p) is defined as the deviation of the absorbed dose in the medium from that predicted by the Spencer-Attix extension of the Bragg-Gray cavity theory where the medium occupies exactly the same volume as the solid state cavity and the electron fluence energy spectrum in the cavity is identical in shape, but not necessarily in magnitude, to that in the medium. The value of gamma(p) has been examined for TL detectors irradiated in megavoltage electron beams (5-20 MeV) using the EGS4 Monte Carlo code. LiF and CaF2 solid state detectors simulated were standard size discs of thickness 1 mm and diameter 3.61 mm irradiated in a water phantom with their centres at d(max) or close to it. Values of gamma(p) for LiF ranged from 0.998 +/- 0.005 to 0.994 +/- 0.005 for electron beams with initial energies of 5 and 20 MeV respectively. For CaF2 the corresponding values were 0.956 +/- 0.006 to 0.989 +/- 0.006 for the same size cavities irradiated at the same depth. EGS4 Monte Carlo simulations demonstrate that the total electron fluence (primary electrons and delta-rays) in these solid state detector materials is significantly different from that in water for the same incident electron energy and depth of irradiation. Thus the Spencer-Attix assumption that the electron fluence energy spectrum in the cavity is identical in shape to that in the medium is violated. Differences in the total electron fluence give rise to electron fluence perturbation correction factors which were up to 5% less than unity for CaF2, indicating a strong violation in this case, but were generally less than 1% for LiF. It is the density of the cavity which perturbs the electron fluence, but it is actually the atomic number differences between the medium and cavity that are responsible for the large electron fluence perturbation correction factors for detectors irradiated close to d(max) because the atomic number affects the change in stopping power with energy. When correction is made for the difference between the electron fluence spectrum in the uniform water phantom and the solid state cavity, the Spencer-Attix cavity equation predicts the dose to water within 0.3% in both clinical and monoenergetic electron beams. Harder's formulation for computing the average mass collision stopping power of water to calcium fluoride, surprisingly, requires perturbation correction factors that are closer to unity than those determined using the Spencer-Attix integrals at depths close to d(max). PMID- 10701503 TI - Photon fluence perturbation correction factors for solid state detectors irradiated in kilovoltage photon beams. AB - Dose perturbation correction factors, gamma(p), for LiF, CaF2 and Li2B4O7 solid state detectors have been determined using the EGS4 Monte Carlo code. Each detector was simulated in the form of a disc of diameter 3.61 mm and thickness 1 mm irradiated in a clinical kilovoltage photon beam at a depth of 1 cm in a water phantom. The perturbation correction factor gamma(p) is defined as the deviation of the absorbed dose ratio from the average mass energy absorption coefficient ratio of water to the detector material, (mu(en)/rho)med,det, which is evaluated assuming that the photon fluence spectrum in the medium and in the detector material are identical. We define another mass energy absorption coefficient ratio, (kappa(en)/rho)med,det, which is evaluated using the actual photon fluence spectrum in the medium and detector for LiF and CaF2 rather than assuming they are identical. (kappa(en)/rho)med,det predicts the average absorbed dose ratio of the medium to the detector material within 0.3%. When the difference in atomic number between the cavity and the phantom material is large then their photon fluence spectra will differ substantially resulting in a difference between (kappa(en)/rho)med,det and (mu(en)/rho)med,det. The value of gamma(p) calculated using (mu(en)/rho)med,det is up to 27% greater than unity for a cavity of CaF2 in 50 kV x-rays. When the atomic number of the medium and detector are similar, their photon fluence spectra are similar, and the difference between (kappa(en)/rho)med,det and (mu(en)/rho)med,det is small. For instance their difference for LiF is less than 2%. The average mass energy absorption coefficient ratio, (mu(en)(E)/rho)w,LiF, evaluated using the mean or representative energy, E, is up to 8% different from (mu(en)/rho)w,LiF. For calcium fluoride the difference between (mu(en)/rho)w,CaF2 and (mu(en)(E)/rho)w,CaF2 is up to 42% in the energy range studied. PMID- 10701504 TI - Tumour control probability: a formulation applicable to any temporal protocol of dose delivery. AB - An analytic expression for the tumour control probability (TCP), valid for any temporal distribution of dose, is discussed. The TCP model, derived using the theory of birth-and-death stochastic processes, generalizes several results previously obtained. The TCP equation is [equation: see text] where S(t) is the survival probability at time t of the n clonogenic tumour cells initially present (at t = 0), and b and d are, respectively, the birth and death rates of these cells. Equivalently, b = 0.693/Tpot and d/b is the cell loss factor of the tumour. In this expression t refers to any time during or after the treatment; typically, one would take for t the end of the treatment period or the expected remaining life span of the patient. This model, which provides a comprehensive framework for predicting TCP, can be used predictively, or--when clinical data are available for one particular treatment modality (e.g. fractionated radiotherapy)--to obtain TCP-equivalent regimens for other modalities (e.g. low dose-rate treatments). PMID- 10701505 TI - Feasibility of penumbra compensating filters for conformal prostate radiotherapy. AB - Radiation therapy beams demonstrate a gradual dose fall off at the field edges, due to factors affecting the physical penumbra and transport of radiation. Adequate target coverage requires an increase in field size larger than the target volume itself for a uniform dose to be delivered to that target volume. A method is presented for the design and fabrication of penumbra compensating filters (PCFs) which essentially sharpen the penumbra on a field-by-field basis and are used in conjunction with custom shielding blocks. We have explored the feasibility of using PCFs to reduce the field margins required for our four-field conformal prostate treatments. The penumbra compensation is designed based on a profile measured along the direction perpendicular to the blocked field edge that shows the greatest 50% to 95% isodose distance for a typical conformal prostate patient. Rigid foam material is milled and filled with a low melting point alloy material to create a filter which provides dose compensation in the field periphery of the custom shielding block. The accuracy of our methodology has been established using film dosimetry. By employing PCFs, the reduction in the rectal margin ranges from approximately 4 mm in the posterior region to 13 mm in the superior-posterior region, as compared with the shielding blocks alone. The reduction in bladder margin ranges from approximately 4 mm in the superior anterior region to 10 mm in the superior region. Dose-volume histograms for an idealized cylindrical rectum indicate a substantial reduction in the volume treated to high doses. The calculated normal tissue complication probability values were 8.7% and 10.5% with and without PCFs included in the blocked fields respectively. The advantages of using PCFs, compared with multileaf collimator based techniques, are discussed. PMID- 10701506 TI - Optimum beam configurations in tomographic intensity modulated radiation therapy. AB - We review and extend the theory of tomographic dose reconstruction for intensity modulated radiotherapy (IMRT). We derive the basis for a saturation with beam number of dose conformation, and provide an analysis which ranks particular beam orientations in terms of the contribution to the delivered dose. Preferred beam directions are found which effectively reduce the number of beams necessary to achieve a given level of dose conformation. The analysis is a new application of the tomographic projection-slice theorem to the problem of beam orientation determination. The effects of the beam front filter and the positivity constraint arising from the tomographic approach are analysed, and modifications of the beam front filter for small beam numbers are suggested. The theory is applied to simple geometric shapes in two dimensions. A Gaussian ellipse, where analytical results are obtained, and simple hard-edged convex prescribed dose shapes are examined to illustrate beam selection based on the beam overlap metric. More complex concave prescribed dose shapes which contain a sensitive organ are also analysed and for low beam numbers are found to have preferred beam directions. PMID- 10701507 TI - 3D cone-beam CT reconstruction for circular trajectories. AB - 3D reconstruction from 2D projections obtained along a single circular source trajectory is most commonly done using an algorithm due to Feldkamp, Davis and Kress. In this paper we propose an alternative approach based on a cone-beam to parallel-beam rebinning step, a corresponding rebinning step into a rectangular virtual detector plane and a filtered backprojection. This approach yields an improved image quality reflected by a decreased low-intensity drop which is well known for 3D reconstruction from projection data obtained along circular trajectories. At the same time the computational complexity is lower than in Feldkamp's original approach. Based on this idea, a hybrid 3D cone-beam reconstruction method is formulated that enlarges the reconstruction volume in its dimension along the rotation axis of the cone-beam CT system. This enlargement is achieved by applying different reconstruction conditions for each voxel. An optimal ratio between the reconstructible and irradiated volume of the scanned object is achieved. PMID- 10701509 TI - Point kernels and superposition methods for scatter dose calculations in brachytherapy. AB - Point kernels have been generated and applied for calculation of scatter dose distributions around monoenergetic point sources for photon energies ranging from 28 to 662 keV. Three different approaches for dose calculations have been compared: a single-kernel superposition method, a single-kernel superposition method where the point kernels are approximated as isotropic and a novel 'successive-scattering' superposition method for improved modelling of the dose from multiply scattered photons. An extended version of the EGS4 Monte Carlo code was used for generating the kernels and for benchmarking the absorbed dose distributions calculated with the superposition methods. It is shown that dose calculation by superposition at and below 100 keV can be simplified by using isotropic point kernels. Compared to the assumption of full in-scattering made by algorithms currently in clinical use, the single-kernel superposition method improves dose calculations in a half-phantom consisting of air and water. Further improvements are obtained using the successive-scattering superposition method, which reduces the overestimates of dose close to the phantom surface usually associated with kernel superposition methods at brachytherapy photon energies. It is also shown that scatter dose point kernels can be parametrized to biexponential functions, making them suitable for use with an effective implementation of the collapsed cone superposition algorithm. PMID- 10701510 TI - Dose rate calculations around 192Ir brachytherapy sources using a Sievert integration model. AB - The classical Sievert integral method is a valuable tool for dose rate calculations around brachytherapy sources, combining simplicity with reasonable computational times. However, its accuracy in predicting dose rate anisotropy around 192Ir brachytherapy sources has been repeatedly put into question. In this work, we used a primary and scatter separation technique to improve an existing modification of the Sievert integral (Williamson's isotropic scatter model) that determines dose rate anisotropy around commercially available 192Ir brachytherapy sources. The proposed Sievert formalism provides increased accuracy while maintaining the simplicity and computational time efficiency of the Sievert integral method. To describe transmission within the materials encountered, the formalism makes use of narrow beam attenuation coefficients which can be directly and easily calculated from the initially emitted 192Ir spectrum. The other numerical parameters required for its implementation, once calculated with the aid of our home-made Monte Carlo simulation code, can be used for any 192Ir source design. Calculations of dose rate and anisotropy functions with the proposed Sievert expression, around commonly used 192Ir high dose rate sources and other 192Ir elongated source designs, are in good agreement with corresponding accurate Monte Carlo results which have been reported by our group and other authors. PMID- 10701511 TI - On the equivalence of rotational and concentric therapy. AB - The aim of this work is to analyse the equivalence of two classes of radiation therapy. One class of therapy is characteristic of Gamma Knife type irradiations and is defined by pencil beam concentric irradiation converging on multiple centres throughout the patient's body. The other class of treatment is characteristic of accelerator based, beam intensity modulated type irradiation defined by a rotation of wide beams around a single centre. We focus our attention on deriving formulae that relate treatments in these two classes and characterize conditions under which they are valid. PMID- 10701508 TI - Personal exposure distribution of solar erythemal ultraviolet radiation in tree shade over summer. AB - The personal radiant exposure distribution of solar erythemal UV in tree shade for an upright posture was measured, with measurements over the whole summer for a total of 17 trees. For each tree, the personal radiant exposure distribution was measured for both the morning and afternoon periods. The exposure ratios averaged over all the trees and over the morning and afternoon periods ranged from 0.16 to 0.49 for the different anatomical sites. A numerical model was employed to estimate the UV radiant exposure to humans in tree shade over the entire summer. The body sites with the higher exposure ratios in the tree shade were the vertex of the head, shoulders and forearms with radiant exposures over the summer of 1300 MED to the vertex of the head and 1100 MED to the shoulders and forearms. These radiant exposures in the shade are substantially higher than the ambient erythemal UV measured in full sun on a horizontal plane over a full summer at a more temperate northern hemisphere latitude. The average radiant exposures per day to each anatomical site for a complete day in the tree shade ranged from 4.6 to 14.6 MED. This research has provided new data that is essential to quantify human UV exposure during outdoor activities. PMID- 10701512 TI - Photon beam characterization and modelling for Monte Carlo treatment planning. AB - Photon beams of 4, 6 and 15 MV from Varian Clinac 2100C and 2300C/D accelerators were simulated using the EGS4/BEAM code system. The accelerators were modelled as a combination of component modules (CMs) consisting of a target, primary collimator, exit window, flattening filter, monitor chamber, secondary collimator, ring collimator, photon jaws and protection window. A full phase space file was scored directly above the upper photon jaws and analysed using beam data processing software, BEAMDP, to derive the beam characteristics, such as planar fluence, angular distribution, energy spectrum and the fractional contributions of each individual CM. A multiple-source model has been further developed to reconstruct the original phase space. Separate sources were created with accurate source intensity, energy, fluence and angular distributions for the target, primary collimator and flattening filter. Good agreement (within 2%) between the Monte Carlo calculations with the source model and those with the original phase space was achieved in the dose distributions for field sizes of 4 cm x 4 cm to 40 cm x 40 cm at source surface distances (SSDs) of 80-120 cm. The dose distributions in lung and bone heterogeneous phantoms have also been found to be in good agreement (within 2%) for 4, 6 and 15 MV photon beams for various field sizes between the Monte Carlo calculations with the source model and those with the original phase space. PMID- 10701513 TI - Hardware-sensitive optimization for intensity modulated radiotherapy. AB - The multileaf collimator (MLC) hardware constraints are usually neglected in the process of intensity-modulated beam optimization. Consequently, it is not always possible to deliver planned beam modulation using dynamic MLC. Beam optimization is significantly diminished if the results must be approximated due to limitations imposed by the delivery device. To overcome this problem, an inverse beam optimization method which incorporates the hardware constraints has been developed. The hardware constraints, including the leaf velocity, the dose rate and the minimum required gap between opposing and adjacent leaves, were considered. An iterative search for feasible modulation was conducted alternately in the dosimetric space and the MLC position-time space. The optimization algorithm was designed for a unidirectional leaf trajectory and a constant dose rate. A scheme to reduce tongue-and-groove underdosage during optimization was also implemented. Comparisons were made between the solutions produced by this method and conventional optimization disregarding the hardware restrictions. The beam profiles generated by the conventional method were modified to satisfy the hardware specifications. The results indicate that inclusion of MLC constraints during optimization can improve the degree of conformity that is deliverable. PMID- 10701514 TI - Viability of the EUD and TCP concepts as reliable dose indicators. AB - The concept of equivalent uniform dose (EUD) was introduced to provide a method of reporting radiotherapy dose distributions which takes account of the nonlinearity of tissue dose-response, whilst not attempting to make predictions of absolute outcome. The purpose of this investigation was to determine the level of sensitivity of EUD to model parameters for significant variations in dose distribution and consequently the reliability of the factor as a dose-indicator, and to compare EUD with the more familiar index, tumour control probability (TCP). EUD and TCP, derived from the linear-quadratic formalism, were investigated for a test tissue being irradiated non-uniformly. Variations in the parameters of the model (tissue cell characteristics, dose heterogeneity, fractionation parameters) indicated the sensitivity of EUD and TCP to them. For time independent factors--cell density, cell radiosensitivity, radiosensitivity heterogeneity (population averaged) and ratio alpha/beta--EUD was found to vary insignificantly in comparison with TCP, though this is a function of the actual form of the dose distribution under consideration. For fractionated treatments where the mean dose per fraction is varying (due to dosimetric/positioning errors for example), both EUD and TCP showed little variation with the degree of dose non-uniformity. For other time dependent factors, fractionation rate and cell repopulation times, TCP again showed significant variation relative to EUD. The relative insensitivity of EUD implies that this index will be useful for dose evaluation when parameters are not known with accuracy, for the intercomparison of dose control studies and as a radiobiologically based optimization objective. However, given confidence in model parameters, the sensitivity of TCP would make it a more reliable tool for indicating potentially successful and unsuccessful irradiation strategies. It is suggested that both parameters be used in conjunction, with EUD and TCP results viewed with an appreciation of the characteristics of each model. PMID- 10701515 TI - Correlation between CT numbers and tissue parameters needed for Monte Carlo simulations of clinical dose distributions. AB - We describe a new method to convert CT numbers into mass density and elemental weights of tissues required as input for dose calculations with Monte Carlo codes such as EGS4. As a first step, we calculate the CT numbers for 71 human tissues. To reduce the effort for the necessary fits of the CT numbers to mass density and elemental weights, we establish four sections on the CT number scale, each confined by selected tissues. Within each section, the mass density and elemental weights of the selected tissues are interpolated. For this purpose, functional relationships between the CT number and each of the tissue parameters, valid for media which are composed of only two components in varying proportions, are derived. Compared with conventional data fits, no loss of accuracy is accepted when using the interpolation functions. Assuming plausible values for the deviations of calculated and measured CT numbers, the mass density can be determined with an accuracy better than 0.04 g cm(-3). The weights of phosphorus and calcium can be determined with maximum uncertainties of 1 or 2.3 percentage points (pp) respectively. Similar values can be achieved for hydrogen (0.8 pp) and nitrogen (3 pp). For carbon and oxygen weights, errors up to 14 pp can occur. The influence of the elemental weights on the results of Monte Carlo dose calculations is investigated and discussed. PMID- 10701516 TI - Production of new thermoluminescent mini-dosimeters. AB - A method of producing CaSO4:Dy thermoluminescent mini-dosimeters was reported in 1986 by B W Wessels for determination of the in vivo absorbed dose in radioimmunotherapy, a field in which absorbed dose gradients are important. These dosimeters, which undergo dissolution when used in a liquid environment, showed a sensitivity loss of up to 30% after 4 days of immersion in our tests. Moreover, several studies have shown that biocompatibility problems can occur during in vivo studies in animals. This paper describes the production and testing of a new type of thermoluminescent mini-dosimeter obtained by microextrusion of a mixture of LiF:Mg,Cu,P polypropylene and plastic adjuvants. These dosimeters, in the form of long 400 microm diameter filaments, can be cut to the desired length. The production process allows an LiF:Mg,Cu,P load of up to 50%. Results obtained in external irradiation indicate that these new miniature LiF:Mg,Cu,P dosimeters have good sensitivity (about 1.6 times that of CaSO4:Dy mini-TLDs), homogeneous response within a production batch (mean +/-4%), response stability in water (0.7% of variation in sensitivity after 2 weeks of immersion) and stability in aqueous solutions at different pH. LiF:Mg,Cu,P mini-dosimeters appear to be highly promising for internal dosimetry, and evaluation is in progress in animals. PMID- 10701517 TI - Verification of dynamic multileaf collimation using an electronic portal imaging device. AB - High standards of treatment verification are necessary where complex new delivery techniques, such as intensity modulated radiation therapy using dynamic multileaf collimation, are being developed. This paper describes the use of a fluoroscopic electronic portal imaging device (EPID) to provide real-time qualitative verification of leaf position during delivery of a dynamic MLC prescription in addition to off-line quantitative verification. A custom-built circuit triggers the EPID to capture a series of snap-shot images at equally spaced dose points during a dynamic MLC prescription. Real-time verification is achieved by overlaying a template of expected leaf positions onto the images as they are acquired. Quantitative off-line verification is achieved using a maximum gradient edge detection algorithm to measure individual leaf positions for comparison with required leaf positions. Investigations have been undertaken to optimize image acquisition and assess the edge detection algorithm for variations in machine dose rate, leaf velocity and beam attenuation. On-line verification enables the operator to monitor the progress of a dynamic delivery and has been used for independent confirmation of accurate dynamic delivery during intensity modulated treatments. Off-line verification allows measurement of leaf position with a precision of 1 mm although image acquisition times must be less than or equal to 140 ms to ensure coincidence of the maximum gradient in the image with the 50% dose level. PMID- 10701518 TI - The measurement of proton stopping power using proton-cone-beam computed tomography. AB - A cone-beam computed tomography (CT) system utilizing a proton beam has been developed and tested. The cone beam is produced by scattering a 160 MeV proton beam with a modifier that results in a signal in the detector system, which decreases monotonically with depth in the medium. The detector system consists of a Gd2O2S:Tb intensifying screen viewed by a cooled CCD camera. The Feldkamp-Davis Kress cone-beam reconstruction algorithm is applied to the projection data to obtain the CT voxel data representing proton stopping power. The system described is capable of reconstructing data over a 16 x 16 x 16 cm3 volume into 512 x 512 x 512 voxels. A spatial and contrast resolution phantom was scanned to determine the performance of the system. Spatial resolution is significantly degraded by multiple Coulomb scattering effects. Comparison of the reconstructed proton CT values with x-ray CT derived proton stopping powers shows that there may be some advantage to obtaining stopping powers directly with proton CT. The system described suggests a possible practical method of obtaining this measurement in vivo. PMID- 10701519 TI - Physical properties of hydrated tissue determined by surface interferometry of laser-induced thermoelastic deformation. AB - Knee meniscus is a hydrated tissue; it is a fibrocartilage of the knee joint composed primarily of water. We present results of interferometric surface monitoring by which we measure physical properties of human knee meniscal cartilage. The physical response of biological tissue to a short laser pulse is primarily thermomechanical. When the pulse is shorter than characteristic times (thermal diffusion time and acoustic relaxation time) stresses build and propagate as acoustic waves in the tissue. The tissue responds to the laser induced stress by thermoelastic expansion. Solving the thermoelastic wave equation numerically predicts the correct laser-induced expansion. By comparing theory with experimental data, we can obtain the longitudinal speed of sound, the effective optical penetration depth and the Gruneisen coefficient. This study yields information about the laser tissue interaction and determines properties of the meniscus samples that could be used as diagnostic parameters. PMID- 10701520 TI - Modelling infrared temperature measurements: implications for laser irradiation and cryogen cooling studies. AB - The use of thermographic techniques has increased as infrared detector technology has evolved and improved. For laser-tissue interactions, thermal cameras have been used to monitor the thermal response of tissue to pulsed and continuous wave irradiation. It is important to note that the temperature indicated by the thermal camera may not be equal to the actual surface temperature. It is crucial to understand the limitations of using thermal cameras to measure temperature during laser irradiation of tissue. The goal of this study was to demonstrate the potential difference between measured and actual surface temperatures in a quantitative fashion using a ID finite difference model. Three ablation models and one cryogen spray cooling simulation were adapted from the literature, and predictions of radiometric temperature measurements were calculated. In general, (a) steep superficial temperature gradients, with a surface peak, resulted in an underestimation of the actual surface temperature, (b) steep superficial temperature gradients, with a subsurface peak, resulted in an overestimation, and (c) small gradients led to a relatively accurate temperature estimate. PMID- 10701521 TI - Source distribution in adjoint Monte Carlo calculation. PMID- 10701522 TI - Stereotactic pallidotomy performed without using microelectrode guidance in patients with Parkinson's disease: surgical technique and 2-year results. AB - OBJECT: Pallidotomy for the treatment of medically refractory Parkinson's disease (PD) has enjoyed renewed popularity. However, the optimal surgical technique, lesion location, and long-term effectiveness of pallidotomy remain subjects of debate. In this article the authors describe their surgical technique for performing pallidotomy without using microelectrode guidance, and the clinical and radiological results of this procedure. METHODS: Patients were evaluated preoperatively by using a battery of validated clinical rating scales and magnetic resonance (MR) imaging of the brain. Individuals with severe treatment refractory idiopathic PD who were believed to be good candidates for surgery underwent computerized tomography scanning- and MR imaging-guided stereotactic pallidotomy. Intraoperative macrostimulation was used to optimize lesion placement and to avoid injury to nearby structures. Lesion location and size were calculated from MR imaging sequences of the brain obtained within the first 24 hours after surgery and again 3 months later. Clinical examinations were conducted at 1.5, 3, 6, 12, and 24 months after surgery. Seventy-five patients (mean age 61 years, range 38-79 years) underwent unilateral pallidotomy. Significant improvements were observed in the "off' period scores for the activities of daily living portion of the Unified Parkinson's Disease Rating Scale (UPDRS), the UPDRS motor scores, total "on" time, levodopa-induced dyskinesias, and contralateral tremor. These improvements were maintained 24 months postoperatively. The mean lesion volume measured on the immediate postoperative MR image was 73 +/- 5.4 mm3. Radiological analysis suggests that initial lesion volume does not predict outcome. The only permanent major complication was a single visual field defect. CONCLUSIONS: Pallidotomy performed without using microelectrode guidance is a safe and effective treatment for selected patients with medically refractory PD. PMID- 10701523 TI - Cognitive outcome following pallidotomy: the influence of side of surgery and age of patient at disease onset. AB - OBJECT: The authors studied the neuropsychological correlates of stereotactically guided lesioning of the right and left posteroventral globus pallidus internus (GPi) in a prospective series of patients suffering from Parkinson's disease (PD). METHODS: Eighteen patients with PD who underwent stereotactically guided lesioning of the GPi (left side in 10 patients and right side in eight) completed neuropsychological evaluations before and after surgery. Patients served as their own controls. Multiple two-by-two repeated-measures analyses of variance were used to assess neuropsychological changes as a function of the side in which lesioning was performed (lesioning on the left side compared with that on the right) and surgery (presurgery compared with postsurgery). Relationships between cognitive variables and patient age at disease onset, age at surgery, and disease duration were examined using a linear regression model. The most striking findings were evident from results of a phonemic word fluency test in which patients in whom a left-sided pallidotomy had been performed achieved a mean performance score that was lower than the score of patients in whom a right-sided pallidotomy had been performed; this score declined even more as a result of surgery. Change in performance on the word fluency test across pre- and postoperative assessments was also related to patient age at onset of PD in those who had undergone left-sided pallidotomy, with patients of an older age at disease onset showing the greatest decline in performance. CONCLUSIONS: These preliminary findings indicate that the side on which surgery was performed and patient age at onset of PD are important in the prediction of postoperative cognitive outcome. The findings also indicate that stereotactically guided lesioning of the GPi presents a relatively mild cognitive risk. PMID- 10701525 TI - Long-term follow up of progesterone receptor status in benign meningioma: a prognostic indicator of recurrence? AB - OBJECT: A long-term prospective analysis of patients with benign meningioma was undertaken to determine whether progesterone receptor (PR) status of the excised tumor has any influence on recurrence. METHODS: Between 1983 and 1985, a total of 62 meningiomas in 53 patients (age range 19-79 years, mean age 55.6 years) were studied for clinical, histological, and pathological characteristics, including hormone receptor status and DNA features. Progesterone receptor status was quantified by cryostat section assay, and then factors affecting recurrence were analyzed. During 1997 all case records were reviewed to determine whether tumor had recurred in any patient, and PR status was correlated with tumor recurrence. Of the 62 tumors, 60 were benign, and of the benign tumors 29 (48%) were PR positive. Patients harboring 14 of the 60 benign tumors were lost to follow up. Of the 46 tumors included in the final analysis, 13 were recurrent (all within 5 years) and 33 were nonrecurrent. Of the 33 nonrecurrent tumors, 14 had not recurred 5 to 10 years postresection and 19 had not recurred after more than 10 years. Chi-square analysis of the results did not show an association between recurrence and patient's sex, extent of resection, histological subtype, or tumor site but did show an association between recurrence and PR negativity (p = 0.013). CONCLUSIONS: The results indicate that benign meningiomas that are PR positive are less likely to recur, a finding that has prognostic and therapeutic implications. PMID- 10701524 TI - Plasma endothelin concentrations after aneurysmal subarachnoid hemorrhage. AB - OBJECT: The pathogenesis of cerebral vasospasm and delayed ischemia after subarachnoid hemorrhage (SAH) seems to be complex. An important mediator of chronic vasospasm may be endothelin (ET), with its powerful and long-lasting vasoconstricting activity. In this study the author investigated the correlation between serial plasma concentrations of ET and ischemic symptoms, angiographically demonstrated evidence of vasospasm, and computerized tomography (CT) findings after aneurysmal SAH. METHODS: Endothelin-1 immunoreactivity in plasma was studied in 70 patients with aneurysmal SAH and in 25 healthy volunteers by using a double-antibody sandwich-enzyme immunoassay (immunometric) technique. On the whole, mean plasma ET concentrations in patients with SAH (mean +/- standard error of mean, 2.1 +/- 0.1 pg/ml) did not differ from those of healthy volunteers (1.9 +/- 0.2 pg/ml). Endothelin concentrations were significantly higher (p < 0.05) in patients who experienced delayed cerebral ischemia with fixed neurological deficits compared with those in other patients (post-SAH Days 0-5, 3.1 +/- 0.8 pg/ml compared with 2.1 +/- 0.2 pg/ml; post-SAH Days 6-14, 2.5 +/- 0.4 pg/ml compared with 1.9 +/- 0.2 pg/ml). Patients with angiographic evidence of severe vasospasm also had significantly (p < 0.05) elevated ET concentrations (post-SAH Days 0-5, 3.2 +/- 0.8 pg/ml; post-SAH Days 6 14, 2.7 +/- 0.5 pg/ml) as did those with a cerebral infarction larger than a lacuna on the follow-up CT scan (post-SAH Days 0-5, 3.1 +/- 0.8 pg/ml; post-SAH Days 6-14, 2.5 +/- 0.4 pg/ml) compared with other patients. Patients in whom angiography revealed diffuse moderate-to-severe vasospasm had significantly (p < 0.05) higher ET levels than other patients within 24 hours before or after angiography (2.6 +/- 0.3 compared with 1.9 +/- 0.2 pg/ml). In addition, patients with a history of hypertension or cigarette smoking experienced cerebral infarctions significantly more often than other patients, although angiography did not demonstrate severe or diffuse vasospasm more often in these patients than in others. CONCLUSIONS: Endothelin concentrations seem to correlate with delayed cerebral ischemia and vasospasm after SAH. The highest levels of ET are predictive of the symptoms of cerebral ischemia and vasospasm, and ET may also worsen ischemia in patients with a history of hypertension. Thus, ET may be an important causal or contributing factor to vasospasm, but its significance in the pathogenesis of vasospasm remains unknown. PMID- 10701526 TI - Prevention of subdural fluid collections following transcortical intraventricular and/or paraventricular procedures by using fibrin adhesive. AB - OBJECT: Subdural fluid collections following transcortical intraventricular and/or paraventricular neurosurgical procedures for tumors are common and can be difficult to treat. The authors prospectively studied the efficacy of a fibrin adhesive (Tisseel) in closing cortical and ependymal defects following intraventricular and/or paraventricular lesion resection and in preventing the development of subdural fluid collections. METHODS: Twenty-five patients who underwent 29 transcortical approaches for the resection of intraventricular and/or paraventricular lesions were studied. No patient developed a symptomatic subdural fluid collection and no new seizure or progression of a preexisting seizure disorder was encountered during a median follow-up time of 29 months (range 1-57 months). The incidence of preoperative hydrocephalus was 72% and four (22%) of these patients required postoperative shunt placement. CONCLUSIONS: The use of a fibrin adhesive to seal cortical and ependymal defects after transcortical procedures appears to prevent the development of subdural fluid collections. PMID- 10701527 TI - Neurosurgical implications of Carney complex. AB - OBJECT: The authors present their neurosurgical experience with Carney complex. Carney complex, characterized by spotty skin pigmentation, cardiac myxomas, primary pigmented nodular adrenocortical disease, pituitary tumors, and nerve sheath tumors (NSTs), is a recently described, rare, autosomal-dominant familial syndrome that is relatively unknown to neurosurgeons. Neurosurgery is required to treat pituitary adenomas and a rare NST, the psammomatous melanotic schwannoma (PMS), in patients with Carney complex. Cushing's syndrome, a common component of the complex, is caused by primary pigmented nodular adrenocortical disease and is not secondary to an adrenocorticotropic hormone-secreting pituitary adenoma. METHODS: The authors reviewed 14 cases of Carney complex, five from the literature and nine from their own experience. Of the 14 pituitary adenomas recognized in association with Carney complex, 12 developed growth hormone (GH) hypersecretion (producing gigantism in two patients and acromegaly in 10), and results of immunohistochemical studies in one of the other two were positive for GH. The association of PMSs with Carney complex was established in 1990. Of the reported tumors, 28% were associated with spinal nerve sheaths. The spinal tumors occurred in adults (mean age 32 years, range 18-49 years) who presented with pain and radiculopathy. These NSTs may be malignant (10%) and, as with the cardiac myxomas, are associated with significant rates of morbidity and mortality. CONCLUSIONS: Because of the surgical comorbidity associated with cardiac myxoma and/or Cushing's syndrome, recognition of Carney complex has important implications for perisurgical patient management and family screening. Study of the genetics of Carney complex and of the biological abnormalities associated with the tumors may provide insight into the general pathobiological abnormalities associated with the tumors may provide insight into the general pathobiological features of pituitary adenomas and NSTs. PMID- 10701528 TI - Paracrine delivery of IL-12 against intracranial 9L gliosarcoma in rats. AB - OBJECT: Interleukin-12 (IL- 12) has potential for the treatment of tumors because it can stimulate an antitumor immune response and possesses antiangiogenic properties. In the study reported here, the authors investigated the therapeutic role of locally delivered IL-12 in a malignant brain tumor model. METHODS: After genetically engineering 9L gliosarcoma cells to express IL-12 (9L-IL12 cells), the authors used these cells as a source of locally delivered cytokine. First, they investigated the behavior of these cells, which were implanted with the aid of stereotactic guidance into the rat brain, by using serial magnetic resonance imaging and histopathological examination. Second, they assessed the antitumor efficacy of proliferating, as well as nonproliferating (irradiated), 9L-IL12 cells by implanting these cells in animals challenged by wild-type 9L gliosarcoma (9Lwt) cells. The IL-12 expression in brain regions injected with 9L-IL12 was confirmed by reverse transcription-polymerase chain reaction. Last, the authors explored whether animals treated with 9L-IL12 cells developed an antitumor immunological memory by rechallenging the survivors with a second injection of 9Lwt cells. The authors demonstrated that local delivery of IL-12 into the rat brain by genetically engineered cells significantly prolongs survival time in animals challenged intracranially with a malignant glioma. CONCLUSIONS: These findings support continued efforts to refine local delivery systems of IL-12 in an attempt to bring this therapy to clinical trials. PMID- 10701529 TI - Locomotion and proliferation of glioblastoma cells in vitro: statistical evaluation of videomicroscopic observations. AB - OBJECT: The motility and doubling of human glioblastoma cells were investigated by means of statistical evaluation of large sets of data obtained using computer aided videomicroscopy. METHODS: Data were obtained on cells in four established glioblastoma cell lines and also on primary tumor cells cultured from fresh surgical samples. Growth rates and cell cycle times were measured in individual microscopic fields. The averages of cell cycle time and the duplication time for the recorded cell populations were 26.2 +/- 5.6 hours and 38 +/- 4 hours, respectively. With these parameters, no significant differences among the cell lines were revealed. Also, there was no correlation in the cell cycle time of a parent cell and its progeny in any of the cultures. Statistical analysis of cell locomotion revealed an exponential distribution of cell velocities and strong fluctuations in individual cell velocities across time. The average velocity values ranged from 4.2 to 27.9 micro/hour. In spite of the uniform histopathological classification of the four tumors, each cell line produced by these tumors displayed distinct velocity distribution profiles and characteristic average velocity values. A comparison of recently established primary cultures with cell lines that had propagated multiple times indicated that cells derived from different tumors sustain their characteristic locomotor activity after several passages. CONCLUSIONS: It can be inferred from the data that statistical evaluation of physical parameters of cell locomotion can provide additional tools for tumor diagnosis. PMID- 10701530 TI - Induction of tolerance against ischemia/reperfusion injury in the rat brain by preconditioning with the endotoxin analog diphosphoryl lipid A. AB - OBJECT: Inflammatory responses and oxygen free radicals have increasingly been implicated in the development of ischemic brain injury. In some cases, an attenuation of inflammation or free-radical injury can provide tissue protection. Diphosphoryl lipid A (DPL) is a detoxified derivative of a lipopolysaccharide (endotoxin) of Salmonella minnesota strain R595, which is capable of stimulating the immune system without eliciting direct toxic effects. In this study the authors examined the influence of preconditioning with DPL on ischemia/reperfusion injury in rats. METHODS: Sprague-Dawley rats were injected intravenously with either DPL or vehicle. Twenty-four hours later, some animals were tested for superoxide dismutase (SOD) activity. Others were subjected to a 3 hour period of focal cerebral ischemia and, after a reperfusion period of 24 hours, were killed. Infarction volume, SOD activity, and myeloperoxidase (MPO) activity were assayed in the postischemic animals. Pretreatment with DPL produced significant reductions in cerebral infarction and MPO activity in the ischemic penumbra. A significant enhancement of basal SOD activity was observed 24 hours after DPL treatment (that is, before ischemia), and a further enhancement of SOD activity was seen in the ischemic penumbra 24 hours after reperfusion. CONCLUSIONS: These data provide the first evidence of a neuroprotective effect of preconditioning with DPL in an in vivo model of cerebral ischemia. Although the precise mechanisms through which DPL exerts its neuroprotective influence remain to be established, an inhibition of the complex inflammatory response to ischemia and an enhancement of endogenous antioxidant activity are leading candidates. PMID- 10701531 TI - Altered diffusion and perfusion in hydrocephalic rat brain: a magnetic resonance imaging analysis. AB - OBJECT: It can be inferred from data published in the literature that brain compression occurs in the early stages of acute hydrocephalus and that drainage of extracellular waste products is impaired. The authors hypothesized that compression of the cortex would alter water distribution and retard the diffusion of fluid in the hydrocephalic brain. METHODS: Proton diffusion, blood perfusion, and T1 and T2 relaxation times were determined in adult rat brain by using magnetic resonance imaging prior to, and 1 and 8 days after induction of hydrocephalus by kaolin injection. Five anatomical regions of interest were studied. The striatum, dorsal cortex, and lateral cortex exhibited decreased T2 and apparent diffusion coefficient (ADC) values but no change in perfusion. Examination of white matter revealed an initial decrease in ADC followed by a significant increase. The T2 relaxation times increased and perfusion decreased progressively between 1 and 8 days after induction of hydrocephalus. CONCLUSIONS: Acute experimental hydrocephalus causes compression of gray matter, perhaps associated with reduction in total water, which impairs diffusion of water in the tissue. White matter compression and hypoperfusion precede the development of edema. These findings have importance for understanding the neurochemical changes that occur in hydrocephalic brains. PMID- 10701532 TI - Neuroprotective effects of citicoline on brain edema and blood-brain barrier breakdown after traumatic brain injury. AB - OBJECT: Cytidine 5'-diphosphocholine (CDPC), or citicoline, is a naturally occurring endogenous compound that has been reported to provide neuroprotective effects after experimental cerebral ischemia. However, in no study has such protection been shown after traumatic brain injury (TBI). In this study the authors examined the effect of CDPC on secondary injury factors, brain edema and blood-brain barrier (BBB) breakdown, after TBI. METHODS: After anesthesia had been induced in Sprague-Dawley rats by using 1.5% halothane, an experimental TBI was created using a controlled cortical impact (CCI) device with a velocity of 3 m/second, resulting in a 2-mm deformation. Four sham-operated control animals used for brain edema and BBB breakdown studies underwent the same surgical procedure, but received no injury. Brain edema was evaluated using the wet-dry method 24 hours postinjury, and BBB breakdown was evaluated by measuring Evans blue dye (EBD) extravasation with fluorescein 6 hours after TBI. The animals received intraperitoneal injections of CDPC (50, 100, or 400 mg/kg two times after TBI [eight-10 animals in each group]) or saline (eight animals) after TBI. Traumatic brain injury induced an increase in the percentage of water content and in EBD extravasation in the injured cortex and the ipsilateral hippocampus. No significant benefit from CDPC treatment was observed at a dose of 50 mg/kg. Cytidine 5'-diphosphocholine at a dose of 100 mg/kg attenuated EBD extravasation in both regions, although it reduced brain edema only in the injured cortex. In both regions, 400 mg/ kg of CDPC significantly decreased brain edema and BBB breakdown. CONCLUSIONS: This is the first report in which dose-dependent neuroprotective effects of CDPC have been demonstrated in the injured cortex as well as in the hippocampus, a brain region known to be vulnerable to injury, after experimental TBI. The results of this study suggest that CDPC is an effective neuroprotective agent on secondary injuries that appear following TBI. PMID- 10701533 TI - Pallidal stimulation for generalized dystonia. Report of three cases. AB - Pallidal stereotactic surgery is a well-accepted treatment alternative for Parkinson's disease. Another indication for this procedure is medically refractory dystonia, especially generalized dystonia with abnormal axial and extremity movements and postures. Improvement of dystonia after pallidotomy has been reported in several recent papers. In this report the authors describe three patients with generalized dystonia (two primary, one secondary) and their improvement after bilateral pallidal stimulation at follow-up times of between 6 and 18 months. PMID- 10701534 TI - Long-term follow-up study of chronic globus pallidus internus stimulation for posttraumatic hemidystonia. AB - The authors report the first case of chronic globus pallidus internus (GPi) stimulation for treatment of medically intractable hemidystonia for which long term follow-up data are available. The patient had developed left-sided low frequency tremor and hemidystonia after a severe head trauma sustained at 15 years of age. He experienced relief of the tremor but not of the hemidystonia after a thalamotomy was performed in the right hemisphere 3 years postinjury. When the patient was 24 years old, the authors performed a magnetic resonance guided stereotactic implantation of a monopolar electrode in the right-sided posteroventral GPi. Chronic deep brain stimulation resulted in remarkable improvement of dystonia-associated pain, phasic dystonic movements, and dystonic posture, which was accompanied by functional gain. Postoperative improvement was sustained after 4 years of follow up. Chronic GPi stimulation appears to be a valuable treatment option for posttraumatic dystonia. PMID- 10701535 TI - Primary meningeal melanocytoma of the pineal region. Case report. AB - A unique case of a meningeal melanocytoma located in the pineal region is presented. This 48-year-old man presented with a round pineal region tumor that caused triventricular hydrocephalus and exhibited slow clinical progression. During surgery a black encapsulated tumor was found and totally removed. On histopathological examination, the tumor proved to be a meningeal melanocytoma. In this report cell culture data are presented and the relevant literature is reviewed. The problems of histopathological diagnosis and management of patients with melanocytomas are also discussed. PMID- 10701536 TI - Congenital primary cerebral angiosarcoma. Case report. AB - Reports of angiosarcoma arising in the central nervous system are rare. The authors present the case of a 30-day-old infant with clinical manifestations of projectile vomiting and tense anterior fontanelle resulting from a left frontotemporal tumor. Total excision of this highly vascular, well-circumscribed tumor was performed without incident, and histopathological examination revealed a malignant angiosarcoma. Immunohistochemical reaction of the neoplastic cells was diffusely positive for endothelium-specific antigens including factor VIII related antigen, CD31, and CD34. The final diagnosis of congenital primary cerebral angiosarcoma was thus confirmed. The patient's postoperative course was uneventful, and he was discharged 2 weeks after the operation. He was in good condition with no sign of recurrence after 11 months; follow-up computerized tomography, magnetic resonance (MR) imaging, and abdominal ultrasonography studies demonstrated no tumor regrowth. The characteristic findings for this tumor on MR imaging, the immunohistochemical findings, and surgical outcome are discussed. PMID- 10701537 TI - Acute posterior fossa syndrome following lumbar drainage for treatment of suboccipital pseudomeningocele. Report of three cases. AB - The authors report on a series of patients who underwent lumbar drainage of cerebrospinal fluid (CSF) for treatment of posterior fossa pseudomeningoceles and who subsequently developed an acute posterior fossa syndrome. These patients were found to have similar radiological findings demonstrating acute mass effect secondary to movement of CSF from the pseudomeningocele into the cerebellar parenchyma. Discontinuation of lumbar drainage resulted in symptomatic and radiological improvement in all patients. From these cases the authors infer that not all pseudomeningoceles communicate directly with the subarachnoid space. A readily recognizable appearance on magnetic resonance imaging that is useful in diagnosing this reversible complication of treatment for posterior fossa pseudomeningocele is also illustrated. PMID- 10701538 TI - Intracranial hypotension without meningeal enhancement on magnetic resonance imaging. Case report. AB - Meningeal enhancement on magnetic resonance (MR) imaging is considered the hallmark radiological feature of intracranial hypotension. The authors report on a patient who exhibited progressively symptomatic intracranial hypotension due to a lumbar cerebrospinal fluid (CSF) leak, but in whom MR imaging demonstrated no pachymeningeal enhancement. This 24-year-old man presented with a 6-week history of progressive orthostatic headaches that were associated with photo- and phonophobia. Four weeks before the onset of the headaches, the patient had undergone a lumbar laminectomy. Brain MR images revealed subdural fluid collections and brain sagging; however, meningeal enhancement was not present. Myelography demonstrated a CSF leak at the site of the laminectomy. At surgery, a large dural tear was repaired. The patient recovered well from the surgery, with complete resolution of his headaches. The absence of meningeal enhancement on MR imaging does not exclude a diagnosis of symptomatic intracranial hypotension. PMID- 10701539 TI - Risk of intraoperative aneurysm clip slippage: a new experience with titanium clips. AB - Slippage of an aneurysm clip as a result of insufficient clip-closing force cannot be predicted, even when using force-testing devices. Descriptions of intraoperative clip slippages are rarely found in the literature. The authors summarize four unusual cases in which titanium aneurysm clips slipped by a scissorslike mechanism during surgery. They analyze the possible factors implicated in such a dangerous situation and discuss corrective choices. PMID- 10701540 TI - Patient presentation, angiographic features, and treatment of strangulation induced bilateral dissection of the cervical internal carotid artery. Report of three cases. AB - Domestic violence leading to strangulation by an abusive spouse can cause carotid artery dissection. This phenomenon is rare and has been described in only three previous instances. The authors present their management strategies in three additional cases. Three young women aged 24 to 43 years were victims of manual strangulation committed by their spouses 3 months to 1 year before presentation. Two of the patients suffered delayed cerebral infarctions before presentation and angiography demonstrated focal, mirror-image severe residual stenoses in the high cervical internal carotid artery (ICA), which were characteristic of a healed chronic dissection; there was no evidence of fibromuscular dysplasia. One of these patients underwent unilateral percutaneous angioplasty with stent placement, and the other underwent bilateral percutaneous angioplasty. Both patients have recovered from their strokes and remain clinically stable at 8 and 20 months posttreatment, respectively. The third patient presented with bilateral ischemic frontal watershed infarctions resulting from an occluded left ICA and a severely narrowed right ICA. Given the extent of the established infarctions, this case was managed with a long-term regimen of anticoagulation medications, and the patient remains neurologically impaired. These cases illustrate the susceptibility of the manually compressed ICA to traumatic injury as a result of domestic violence. They identify bilateral symmetrical ICA dissection as a consistent finding and the real danger of delayed stroke as a consequence of strangulation. Endovascular therapy in which percutaneous angioplasty and/or stent placement are used can be useful in treating residual focal stenoses to improve cerebral perfusion and to lower the risk of embolic or ischemic stroke. PMID- 10701541 TI - Interdural origin of the ophthalmic artery at the dural ring of the internal carotid artery. Report of two cases. AB - The authors report two cases in which the ophthalmic artery (OA) originated from the interdural portion of the internal carotid artery at the carotid dural ring and coursed within the dura. This configuration was observed during surgeries performed in 82 cases of juxta-dural ring aneurysms. In surgery for such an aneurysm, if the OA is not seen intradurally, an attempt should be made to find this kind of variation by using a Doppler flowmeter before sectioning the dural ring. PMID- 10701542 TI - Benign aqueductal cyst causing bilateral internuclear ophthalmoplegia after external ventricular drainage. Case report. AB - The introduction of magnetic resonance (MR) imaging to the field of neuroimaging has allowed detection of various lesions that cause aqueductal stenosis. The authors report the case of a 3-year-old boy in whom a benign ventricular cyst developed in the aqueduct. The patient became drowsy after having complained of headache and vomiting; MR imaging revealed mild triventricular dilation and a normal-sized fourth ventricle. Repeated MR imaging performed 1 week later revealed an aqueductal cyst that had markedly enlarged during the intervening period. An external ventricular drainage system was installed, but recovery of consciousness in the child was unsatisfactory and a new bilateral internuclear ophthalmoplegia developed. Fenestration of the cyst wall and placement of a ventriculocisternostomy in the third ventricle were performed simultaneously by using a flexible neuroendoscope. By 2 weeks postsurgery, the patient's neurological symptoms had completely resolved. This case illustrates that simple rerouting of ventricular cerebrospinal fluid (CSF) can aggravate the symptoms of this rare lesion by causing severe compression of periaqueductal structures by a cyst that maintains a high intracystic pressure. Endoscopic surgery was an excellent choice of treatment to achieve both cyst fenestration and normalization of intracranial CSF pressure by creating a ventriculocisternostomy. PMID- 10701543 TI - Neural transplantation cannula and microinjector system: experimental and clinical experience. Technical note. AB - The authors present a simple, reliable, and safe system for performing neural transplantation in the human brain. The device consists of a transplantation cannula and microinjector system that has been specifically designed to reduce implantation-related trauma and to maximize the number of graft deposits per injection. The system was evaluated first in an experimental rat model of Parkinson's disease (PD). Animals in which transplantation with this system had been performed showed excellent graft survival with minimal trauma to the brain. Following this experimental stage, the cannula and microinjector system were used in eight patients with PD enrolled in the Halifax Neural Transplantation Program who received bilateral putaminal transplants of fetal ventral mesencephalic tissue. A total of 16 transplantation operations and 64 trajectories were performed in the eight patients, and there were no intraoperative or perioperative complications. Magnetic resonance imaging studies obtained 24 hours after surgery revealed no evidence of tissue damage or hemorrhage. Transplant survival was confirmed by fluorodopa positron emission tomography scans obtained 6 and 12 months after surgery. As neural transplantation procedures for the treatment of neurological conditions evolve, the ability to deliver viable grafts safely will become critically important. The device presented here has proved to be of value in maximizing the number of graft deposits while minimizing implantation-related trauma to the host brain. PMID- 10701544 TI - Median nerve penetration by an anomalous tendon. Case illustration. PMID- 10701545 TI - Spontaneous resolution of a sphenoid sinus encephalocele. Case illustration. PMID- 10701546 TI - Pneumoangiogram in a patient with severe head injury. Case illustration. PMID- 10701547 TI - Giant interdural cavernous hemangioma at the convexity. Case illustration. PMID- 10701548 TI - Enlarging vertebrobasilar dolichoectasia with subarachnoid hemorrhage heralded by recurrent ischemia. Case illustration. PMID- 10701549 TI - Pneumocephalus causing pulsatile tinnitus. Case illustration. PMID- 10701550 TI - Solitary metastasis to the choroid plexus. Case illustration. PMID- 10701551 TI - Tirilazad and subarachnoid hemorrhage. PMID- 10701552 TI - Bilateral pallidotomy. PMID- 10701554 TI - Third ventriculostomy. PMID- 10701553 TI - Papaverine and vasospasm. PMID- 10701556 TI - Effect of temperature on fatty acid composition in each lipid fraction of Spirometra erinaceieuropaei plerocercoids. AB - The effects of temperature and host fatty acids on the fatty acid contents of Spirometra erinaceieuropaei plerocercoids were investigated to clarify their role in sparganosis. After 24 hr incubation at 18 C in host snake serum, omega6 series fatty acids, especially arachidonic acid in the phospholipid fraction of the plerocercoids, increased compared with those of plerocercoids incubated at 37 C. The changes in the ratio of polyunsaturated to saturated fatty acids in the phospholipid fraction of plerocercoids incubated in physiological saline for 6 hr at 10 C were almost the same as the changes at 37 C. The ratio of polyunsaturated to saturated fatty acids of the triglyceride fraction showed almost opposite change versus the phospholipid fraction. The percentage of arachidonic acid in the phospholipid fraction of plerocercoids increased during the first 3 hr of incubation and then decreased, regardless of temperature. At 37 C, the percentage of arachidonic acid in the free fatty acid fraction fell for the first 3 hr of incubation and was significantly elevated at the end of the 6-hr incubation. At 10 C, however, arachidonic acid in the free fatty acid fraction decreased for the first hour of incubation, increased at 3 hr of incubation, then decreased again. These results suggest that fatty acids of the plerocercoids are frequently exchanged between fractions. Plerocercoids can mobilize arachidonic acid to the free fatty acid fraction more quickly at lower temperature than at higher temperature. They may utilize mobilized arachidonic acid early in the infection stage to produce prostaglandins. Alternatively, they can incorporate arachidonic acid into the phospholipid fraction again when arachidonic acid is readily available in the environment. PMID- 10701555 TI - Characterization of acid phosphatase and phosphorylcholine hydrolase in adult Haemonchus contortus. AB - An acid phosphatase (AP) and a phosphorylcholine hydrolase (PCH) were detected in excretory-secretory (ESP) products from adult Haemonchus contortus. The AP had a pH optimum of 4.5 and was inhibited by tartaric acid and sodium fluoride, but not by o-phenanthroline. The AP hydrolyzed paranitrophenol (pnp)-phosphate and to a lesser extent pnp-phenyl-phosphonate but did not hydrolyze diester substrates. Purified AP consisted of heterodimers with relative molecular weight (Mr) of 41.9 and 48.7 kDa and had a native molecular weight of 98 kDa by size-exclusion chromatography (SEC). The PCH had a pH optimum of about 9.5 and was inhibited by EDTA and o-phenanthroline but not by the specific phospholipase inhibitor D609. The specific activity of PCH in the ESP was approximately 25-fold less than that of AP. PCH also hydrolyzed 5'-thymidine monophosphate-pnp at a rate about 40% lower than pnp-phosphorylcholine but did not hydrolyze 3'-thymidine monophosphate pnp. Partial purification of PCH suggests an Mr of 50.2 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and an Mr of 102 kDa by SEC. Both AP and PHC were secreted in vitro in a time-dependent manner and had their highest concentrations in the intestine. The results indicate that H. contortus adults secrete significant amounts of AP that might be a digestive enzyme. PCH is also an intestinal enzyme and is secreted in lesser amounts than AP. The PCH is probably not a phospholipase C but has some characteristics of a type I phosphodiesterase. PMID- 10701557 TI - Purification and characterization of a secretory serine proteinase of Acanthamoeba healyi isolated from GAE. AB - We purified and characterized a serine proteinase secreted by Acanthamoeba healyi to evaluate it as a possible virulence factor in the pathogenesis of granulomatous amoebic encephalitis (GAE). Ammonium sulfate precipitated culture supernatant of A. healyi OC-3A strain was purified by chromatography on CM Sepharose, Sephacryl-S200, and Q-2 anion-exchange columns. The purified 33-kDa enzyme had a pH optimum of 8.0 and a temperature optimum of 40 C. Phenylmethylsulfonylfluoride and diisopropyl fluorophosphate, serine proteinase inhibitors, diminished activity of the enzyme to near zero. In addition to types I and IV collagen and fibronectin, the main components of the extracellular matrix, other proteins such as fibrinogen, IgG, IgA, albumin, and hemoglobin were also degraded by the enzyme. The broad substrate specificity of this secreted serine proteinase suggests that it may play an important role in pathogenesis of GAE by A. healyi. PMID- 10701558 TI - Ultrastructural and lectin-histochemical differences between the scolex/strobila and bladder teguments of the Taenia taeniaeformis strobilocercus. AB - The strobilocercus stage of the cat tapeworm Taenia taeniaeformis is surrounded by a single syncytial sheet of cytoplasm called the tegument. The outer membrane of the tegument covers both the scolex/strobila (S/S) and the bladder portions of the strobilocercus, but only the S/S region is resistant to intestinal digestion. It has been suggested that the glycocalyx, the surface-exposed glycoconjugates of the outer membrane, may serve to insulate underlying surface membrane components from digestion. In this study, we used lectin binding to test the hypothesis that the glycocalyx of the S/S is different from that of the bladder and that this may serve as the resistance mechanism of the S/S to digestion. Biotin-labeled lectins and an avidin-glucose oxidase detection system were applied to whole strobilocerci and to 1-microm epon-araldite plastic-embedded sections. Lectins bound to either both regions of the strobilocerci, to the S/S regions only, or did not bind at all. The restriction of some glycoconjugates to the glycocalyx of the S/S region only is consistent with our hypothesis. PMID- 10701559 TI - Comparative development and merozoite production of two isolates of Sarcocystis neurona and Sarcocystis falcatula in cultured cells. AB - The development and merozoite production of Sarcocystis falcatula and 2 isolates (SN6 and SN2) of Sarcocystis neurona were studied in various cultured cell lines inoculated with culture-derived merozoites. All 3 parasites underwent multiple cycles of schizogony in VERO cells, bovine monocytes (M617 cells), and bovine pulmonary artery endothelial cells (CPA). Sarcocystis neurona strains SN6 and SN2 formed schizonts in rat myoblasts (L6) but not in quail myoblasts (QM7); S. falcatula formed schizonts in QM7 cells but not in L6 cells. Merozoites did not develop to sarcocysts in the myoblast cells lines. During a 47-day culture period in VERO cells, SN6 produced substantially more merozoites than did SN2 or S. falcatula. M617 cells produced substantially more merozoites of SN6 than did VERO or CPA cells. During a 17-day culture period of SN6, M617 cells produced mean totals of 4.7 x 10(8) merozoites, VERO cells produced 1.9 x 10(8) merozoites, and CPA cells produced 5.9 x 10(7) merozoites. At 4-12 days after inoculation of cultured cells with SN6, M617 cells cultured in the presence of 10% fetal bovine serum (FBS) produced a mean merozoite total of 5.1 x 10(8) compared to 3.6 x 10(8) for culture medium containing 1% FBS. PMID- 10701560 TI - Pharyngeal bot flies in Cervus elaphus in central Spain: prevalence and population dynamics. AB - The prevalence and intensity of infestations by bot flies Pharyngomyia picta and Cephenemyia auribarbis in red deer (Cervus elaphus) from Quintos de Mora (Toledo, Spain) were determined over a 1-yr period. Bots were present all year. No clear correlations were found between age or sex of the host and parasitization levels (prevalence and intensity). Considerable variation was found in prevalence and intensity, with larger values from December to March. Cephenemyia auribarbis was restricted from November to March, with maximum numbers of L-3 in February. Pharyngomyia picta showed a more complex profile with 2 peaks (March and August), indicating 2 generations per year. PMID- 10701561 TI - Molecular phylogenetic analysis of ixodid ticks based on the ribosomal DNA spacer, internal transcribed spacer 2, sequences. AB - An internal transcribed spacer (ITS2) sequence between the 5.8S and 28S rRNA genes was used to estimate the phyletic relationships among Ixodes spp. tick vectors of Lyme disease-causing Borrelia spirochetes. Analysis indicates that Borrelia burgdorferi sensu lato species associated with Lyme disease are found mainly in ticks of the Ixodes ricinus species complex. Other closely related tick species are not known to transmit the Borrelia-that cause Lyme disease in humans, but they appear to have a specific association with other closely related Borrelia species. There is a high degree of concordance in the phylogenetics of Borrelia taxa and the phylogenetic relationships among Ixodes ticks. PMID- 10701563 TI - A light microscopy study of the migration of Naegleria fowleri from the nasal submucosa to the central nervous system during the early stage of primary amebic meningoencephalitis in mice. AB - The migratory pathway of Naegleria fowleri from the nasal submucosa to the central nervous system (CNS) during the early stage of primary amebic meningoencephalitis (PAM) was investigated in mice. Twenty-one-day-old CD-1 mice were inoculated by intranasal instillation of 1 x 10(6) amebas. Animals were divided into 3 groups of 5 and, after being anesthetized, were killed at intervals of 24, 32, and 48 hr postinoculation by transcardial perfusion with formaldehyde, acetic acid, and methanol. The heads were decalcified, divided in the midsagittal plane, and the area of the cribriform plate removed and embedded in paraffin. Serial sections were cut at 8 microm and stained with a combination of celestin blue, Harris' hematoxylin, and acid fuchsin for light microscopy. Focal inflammation and amebas were observed in the submucosal nerve plexus, olfactory nerves penetrating the cribriform plate, and the olfactory bulb of the brain as early as 24 hr postinoculation. The time periods selected assured that the disease process would not obliterate soft tissue structures. Earlier studies used moribund mice in which the inflammation and the number of amebas were overwhelming. The present study provides convincing evidence that amebas gain initial access to the CNS through olfactory nerves within the cribriform plate during the early stages of PAM. PMID- 10701562 TI - Comparison of efficacy of American and African Amblyomma ticks as vectors of heartwater (Cowdria ruminantium) infection by molecular analyses and transmission trials. AB - The ability of Amblyomma americanum, Amblyomma cajennense, Amblyomma maculatum, and Amblyomma variegatum to acquire and transmit Cowdria ruminantium infection was investigated. Uninfected nymphs were fed on clinically reacting C. ruminantium-infected sheep and then analyzed for infection by specific DNA detection assays and by tick transmission trials. By polymerase chain reaction (PCR), the mean infection prevalence of A. maculatum ticks (50.7%) was similar to that of A. variegatum, Elevage strain (43.5%; P = 0.83) and Petit Bourg strain (45.9%; P = 0.26) ticks. Though Amblyomma hebraeum were not tested by PCR, by DNA probe their infection prevalence was 94%. In contrast, A. americanum and A. cajennense ticks demonstrated very low susceptibility to C. ruminantium, and the prevalence of infection by PCR was approximately 1%. The higher susceptibility of A. maculatum and A. variegatum to C. ruminantium correlated with superior vector efficiency, depicted by similar prepatent periods and severity of disease transmissions to sheep. Amblyomma americanum and A. cajennense failed to transmit infection, confirming that low susceptibility to C. ruminantium correlates with the poor vector status of these species. These results highlight the importance of A. maculatum as a potential vector that is likely to play a major role in the establishment and maintenance of heartwater, if the disease were to be introduced to the U.S.A., Central, and South America. PMID- 10701564 TI - Experimental infections of farmed eels with different Trypanosoma granulosum life cycle stages and investigation of pleomorphism. AB - Trypanosoma granulosum, a flagellate protozoon commonly found in the blood of the European eel Anguilla anguilla, was injected experimentally into uninfected eels purchased from a local farm. In order to investigate the infectivity of different stages in the life cycle, trypanosomes from various sources were used for inoculation. Infectivity was greatly reduced in in vitro culture stages inoculated at 20 C. Isolated bloodstream stages injected into groups of animals held at 12 and 20 C could be detected for over 70 days but did not appear to multiply. Naturally infected Hemiclepsis marginata, a piscivorous leech known to serve as vector, produced detectable, single-peak infections in eels held at 20 C. Infections were characterized by a prepatent stage and a phase of rising parasitemia. Peak infection intensities ranged between 1 and 7 x 10(4) trypanosomes/ml. Trypanosomes in the bloodstream of eels experimentally infected with leeches, divided at a very low rate during the early stages of infection. Small morphs present during the early phase of rising parasitemia were gradually replaced by larger trypanosomes. The overall length frequency distribution of trypanosomes was unimodal. PMID- 10701565 TI - Analysis of messages expressed by Echinostoma paraensei miracidia and sporocysts, obtained by random EST sequencing. AB - A lambdaZAP Express cDNA library was constructed with mRNA obtained from immature miracidia within eggs, hatched miracidia, and sporocysts of Echinostoma paraensei. This cDNA library was amplified and 213 expressed sequence tag (EST) sequences (averaging 466 nucleotides in length) were obtained. The mean percentage of unresolved bases within the EST sequences was 0.4%, ranging from 0 to 4.6%. The 213 ESTs represent 151 unique messages. BLAST (version 2.0.8) analysis disclosed that 64 unique E. paraensei messages (42.4%) had significant similarities (BLAST score < or =e-5), at deduced amino acid or nucleotide levels, with known sequences in the nonredundant GenBank databases or the dbEST database (NCBI). The remainder, 57.6% of the unique EST-encoded messages, scored nonsignificant hits. Most of the E. paraensei messages that could be assigned a cellular role based on sequence similarities were involved in gene/protein expression. Several ESTs scored highest similarities with sequences obtained from trematode species. A total of 22,560 nucleotides present in open reading frames from ESTs that aligned with known sequences was used to determine codon usage for E. paraensei. Analysis of a subset of eight ESTs that contained full-length open reading frames did not reveal a bias in codon usage. Also, EST sequences were found to contain 3' untranslated regions with an average length of 69.9 +/- 88.4 nucleotides (n = 46). The EST sequences were submitted to GenBank/dbEST, adding to the 51 available Echinostoma-derived sequences, to provide reference information for both phylogenetic analysis and study of general trematode biology. PMID- 10701566 TI - Taenia crassiceps cysticercosis: protective effect and immune response elicited by DNA immunization. AB - The nucleotide sequence of a protective recombinant antigen of Taenia crassiceps cysticerci present in all stages of Taenia solium (KETc7), cloned into pcDNA3 plasmid with the signal peptide sequence of the beta-glycan receptor (pTc-sp7), has been shown to be effective in protecting mice against experimental infection of T. crassiceps. To explore further the possibilities of this form of immunization and the immune response induced, mice were injected intramuscularly (i.m.) or intradermally (i.d.) with 3 doses of pTc-sp7. Similar levels of resistance were found using either i.m. or i.d. immunization. Spleen cells from i.d. and i.m. DNA immunized mice induced a specific T-cell response to T. crassiceps antigens and to a synthetic peptide from the immunogen itself (GK-1). Proliferated cells were especially enriched in CD8+ CD4- T-lymphocytes. A clear increase in the percentage of CD3+ cells that produce gamma-interferon and interleukin-2 was detected when measuring the intracellular cytokine production, an indication of the pTc-sp7 capacity to induce an effective cellular response. These results provide encouraging information on the use of KETc7 in the prevention of cysticercosis as well as a first insight into the characterization of the immune response induced by pTc-sp7 that hints to the relevance of cellular immunity in protection. PMID- 10701568 TI - Leishmania amazonensis infection does not inhibit systemic nitric oxide levels elicited by lipopolysaccharide in vivo. AB - Leishmaniasis is a parasitic disease that leads to chronic inflammation. Macrophages, depending on their activation state, are either hosts or killers of the parasites. Downregulation of nitric oxide (NO) synthesis by the parasite infecting the macrophages has been proposed to be an important evading mechanism based on in vitro studies. We confirmed inhibition of NO release by macrophages infected with Leishmania amazonensis in vitro. To examine the role of the parasite in regulating NO production in vivo, we monitored systemic NO levels elicited by challenging naive and L. amazonensis-infected BALB/c mice with lipopolysaccharide (LPS). Animals were challenged after 1, 2, 6, and 9 wk of infection. NO production was monitored by electron paramagnetic resonance spectroscopy as the levels of hemoglobin nitrosyl complexes (HbNO) present in the animal's blood. No significant differences in HbNO levels were observed between LPS-treated naive and inoculated mice at any time during infection. To control for increased macrophage numbers in infected mice, naive mice were injected with a macrophage cell line before LPS challenge; this treatment did not increase produced NO levels. The results argue against a major role for the parasite in downregulating NO production in vivo. PMID- 10701567 TI - Failure to identify alveolar echinococcosis in trappers from South Dakota in spite of high prevalence of Echinococcus multilocularis in wild canids. AB - Echinococcus multilocularis causes a rare but potentially lethal zoonotic disease in humans. This tapeworm has been known to be endemic in foxes (Vulpes vulpes) and coyotes (Canis latrans) within the northern United States since the 1960s. One purpose of this study was to provide recent data on the prevalence of E. multilocularis in foxes and coyotes from eastern South Dakota. In a survey conducted from 1987 to 1991 and involving 137 foxes and 9 coyotes from this area, 74.5% of the foxes and 4 of the coyotes were infected. To assess the possible prevalence of alveolar echinococcosis in a group at presumptive high risk, we also conducted a serological survey of members of the South Dakota Trappers Association in 1990 and 1991. Serum samples from 115 trappers were evaluated for the presence of E. multilocularis antibodies using enzyme-linked immunosorbent assay tests involving a purified antigen called Em2, a crude E. multilocularis antigen, and a recombinant E. multilocularis antigen called II/3-10. None of the trappers showed antibody evidence for the presence of E. multilocularis. Roughly half of the surveyed individuals had trapped more than 50 foxes during their life, and almost one-fourth had trapped more than 1,000 foxes. PMID- 10701569 TI - Spinitectus mexicanus n. sp. (Nematoda : Cystidicolidae) from the intestine of the freshwater fish Heterandria bimaculata in Mexico. AB - A new nematode, Spinitectus mexicanus n. sp., is described on the basis of the specimens recovered from the intestine of Heterandria bimaculata (Heckel) (Poeciliidae, Cyprinodontiformes) from 3 rivers of the Papaloapan River basin (type locality La Basura River), Los Tuxtlas, Veracruz State, Mexico. It differs from its congeners mainly in having the spination of the cuticle separated into 4 longitudinal sectors, each with posteriorly diminishing numbers of larger spines at the anterior part of body. It is the first species of Spinitectus described from a poeciliid fish and the second reported from freshwater fishes in Mexico. PMID- 10701570 TI - A phylogenetic hypothesis for species of the genus Taenia (Eucestoda : Taeniidae). AB - Cladistic analysis of a numerical data matrix describing 27 characters for species of Taenia resulted in 4 most parsimonious phylogenetic trees (174 steps; consistency index = 0.28; homoplasy index = 0.72; retention index = 0.48). Monophyly for Taenia is diagnosed by the metacestode that is either a cysticercus or a form derived from a bladder-like larva; no other unequivocal synapomorphies are evident. Tree structure provides no support for recognition of a diversity of tribes or genera within the Taeniinae: Fimbriotaeniini and Taeniini have no phylogenetic basis. Hydatigera, Fimbriotaenia, Fossor, Monordotaenia, Multiceps, Taeniarhynchus, Tetratirotaenia must be subsumed within Taenia as synonyms. Taenia saginata and Taenia asiatica are sister species and distantly related to Taenia solium. Cospeciation with respect to carnivorous definitive hosts and Taenia appears to be limited. Although felids are putative ancestral hosts, contemporary associations appear to have resulted from extensive host-switching among felids, canids, hyaenids, and others. In contrast, relationships with herbivorous intermediate hosts are indicative of more pervasive coevolution; rodents as intermediate hosts are postulated as ancestral for the Taeniidae, Taenia + Echinococcus. Patterns appear consistent with rapid shifts between phylogenetically unrelated carnivores but among those that historically exploited a common prey resource within communities in specific biogeographic regions. PMID- 10701571 TI - Strelkovimermis amphidis n. sp. from chironomid adults emerging from Lake Itasca and Long Lake, Minnesota. AB - Mermithid nematodes, Strelkovimermis amphidis n. sp., emerged from chironomid imagos from Lake Itasca in Minnesota in the fall of 1996, 1997 and from Long Lake in the fall of 1998. The species is distinguished from the other 11 members of the genus by the long cephalic papillae, absence of an excretory pore, pointed termini in both sexes, large amphids, body diameter decrease at the vulva, long vagina, and the absence of lateral genital papillae. Strelkovimermis amphidis n. sp. is the fifth member of this genus recorded from Lake Itasca. The presence of and nature of the bursal sleeve is suggested as a useful distinguishing characteristic. The ratios involving spicule axis length, diameter of the body at the genital pore, and the length of the tail are also discussed in distinguishing species of Strelkovimermis. An expanded key to the species of Strelkovimermis is included. PMID- 10701572 TI - Tetracapsula renicola n. sp. (Myxozoa : Saccosporidae); the PKX myxozoan--the cause of proliferative kidney disease of salmonid fishes. AB - Proliferative kidney disease (PKD) of salmonid fishes is caused by the extrasporogonic stage of an enigmatic myxozoan, referred to as PKX. Sporogenesis occurs in the renal tubules, resulting in monosporous pseudoplasmodia. The spores are ovoid with indistinguishable valves and measure 12 microm in length and 7 microm in width. Two spherical polar capsules (2 microm diameter) with 4 coils occur at the anterior end of the spore. Prominent capsulogenic cell nuclei posterior to the polar capsules are evident in histological sections stained with hematoxylin and eosin. Regardless of the true nature of the valves (indistinguishable or absent), this myxozoan is morphologically distinct from all other described members of the phylum Myxozoa. Comparisons of small subunit rDNA sequences of PKX with other myxozoans demonstrated that it branches from all other members of the myxosporeans from fish examined thus far, including representatives of the phenotypically most closely related genera, Sphaerospora and Parvicapsula. Recent reports, based on rDNA comparisons, indicate that the alternate stage of PKX occurs in bryozoans, and that PKX clusters in a clade with Tetracapsula bryozoides. Our analyses and those of others, along with phenotypic observations, indicate that salmonids are the primary myxosporean hosts for PKX, that the cryptic spores of PKX in salmonids are the fully formed myxospores as they occur in the fish host, and that PKX represents distinct species that we previously place in the genus Tetracapsula in the family Saccosporidae. The latter 2 taxa were described based on stages from bryozoans, and the myxosporean stage in fish of the type species, T. bryozoides, has not been identified (if it exists). Thus, more complete resolution of the life cycle of both PKX and T. bryozoides, as well as more genetic data, are required to determine the precise relationship of these organisms. PMID- 10701573 TI - Paraquimperia Africana n. sp (Nematoda : Quimperiidae), a new intestinal parasite of the eel Anguilla mossambica Peters, in South Africa. AB - A new seuratoid nematode of the family Quimperiidae, Paraquimperia africana n. sp., is described from the small intestine of the longfin eel, Anguilla mossambica Peters, from the Eastern Cape Province, South Africa. The new species is characterized mainly by the presence of a ventral sucker in mature males, short spicules (147-171 microm), the number and arrangement of caudal papillae, the postesophageal position of the excretory pore, and by the slender female tail. In this new species, a variability in the number (3-5 pairs) of subventral preanal papillae was observed. Paraquimperia africana is the first representative of the genus in Africa. In view of recent reports, Paraquimperia aditum (Mueller, 1934) is considered a junior synonym of Paraquimperia tenerrima (Linstow, 1878). Paraquimperia xenentodonia Gupta and Bakshi, 1984 is considered a species inquirenda. PMID- 10701574 TI - Three new Procamallanus (Spirocamallanus) species from freshwater fishes in Mexico. AB - The following 3 new species of Procamallanus (Spirocamallanus) are described from the intestines of freshwater fishes in Mexico, all belonging to the morphological group characterized by the presence of wide caudal alae, 3 pairs of subventral preanal papillae, and unequal spicules in the male: Procamallanus (Spirocamallanus) jaliscensis n. sp. (type host: Agonostomus monticola) and Procamallanus (Spirocamallanus) gobiomori n. sp. (hosts: Gobiomorus maculatus [type host], Gobiomorus polylepis and Eleotris picta) from 2 rivers in Jalisco State, western Mexico, and Procamallanus (Spirocamallanus) mexicanus n. sp. (type host: Cichlasoma geddesi) from Xalapa District, Veracruz State (Gulf of Mexico region), southeastern Mexico. Procamallanus jaliscensis is characterized by the length of the spicules (606-900 microm and 282-354 microm), number (15-16) of spiral ridges in the buccal capsule, and the digit-like protrusion with 1 terminal cuticular spike on the female tail; P. mexicanus by the length of the spicules (456-480 microm and 231-233 microm), number (10-12) of spiral ridges in the capsule, and the shape of the female tail (conical with a suddenly narrowed distal part, without any terminal spikes); and P. gobiomori by the length of spicules (318-348 microm and 156-192 microm), number (8-10) of spiral ridges and by the digit-like protrusion with 2 terminal cuticular spikes on the female tail. PMID- 10701576 TI - Trichinella murrelli n. sp: etiological agent of sylvatic trichinellosis in temperate areas of North America. AB - Trichinella T5, collected from sylvatic carnivores in North America, was identified previously as a different phenotype of Trichinella, with an uncertain taxonomic level due to the availability of only 2 isolates. Cross-breeding experiments carried out with single female and male larvae of 2 strains of Trichinella T5, with single female and male larvae of 2 strains of Trichinella spiralis, Trichinella nativa, Trichinella britovi, Trichinella pseudospiralis, Trichinella nelsoni, and Trichinella T6, showed a reproductive isolation of Trichinella T5. Viable offspring were obtained only when a female of Trichinella T5 was crossed with a male of T. britovi, but not vice versa. Furthermore, the analysis of biological, biochemical, and molecular data of 32 isolates collected from sylvatic animals in the Nearctic region and identified as Trichinella T5 permitted its reassessment at the species level. Trichinella murrelli n. sp. is characterized by the following: distribution in temperate areas of the Nearctic region; newborn larvae production in vitro of 29-36/72 hr; nurse cell development time between 24 and 70 days postinfection; reproductive capacity index in Swiss mice 1.2-9.5, in wild mice 29.5-159.8, in rats 0.7-2.4, and in pigs 0.03-0.0004; no resistance to freezing; ribosomal DNA fragments of 7.2 kb and/or 11.4 kb, plus 2.2 kb and 1.8 kb present after Dra I digested DNA when probed with total T. spiralis RNA; a specific amplicon of 179 bp after polymerase chain reaction (PCR) amplification with the primer set SB147G; a specific fragment of 1,600 bp after PCR amplification with the primer set Ts43CA and Hhb I digestion; long incubation period; and moderate to severe pathogenicity for humans. The new species is most similar to T. britovi, though it differs from T. britovi in the pattern of 2 allozymes, in the patterns of major ribosomal DNA and PCR-restriction fragment length polymorphism fragments, and in geographical distribution. PMID- 10701575 TI - Small subunit ribosomal DNA phylogeny of microsporidia with particular reference to genera that infect fish. AB - Molecular data have proved useful in the study of microsporidia phylogeny. Previous studies have shown that there are several important differences between phylogenies based on rRNA and morphological data. In the present study, small subunit (SSU) rDNA sequences were obtained from 7 different fish-infecting microsporidia from 4 different genera (Glugea Thelohan, 1891, Loma Morrison and Sprague, 1981, Pleistophora Gurley, 1893, and Spraguea Weissenberg, 1976). The lengths of the SSU rDNA genes in these species were between 1,332 and 1,343 base pairs. Phylogenetic analysis was performed using parsimony, maximum likelihood, and Kimura 2-parameter with neighbor joining. The analyses revealed that the microsporidia could be divided into 3 major groups. With the exception of Nucleospora salmonis Hedrick, Groff, and Baxa, 1991, all the microsporidia infecting fishes occurred in the same group. The analysis showed that Pleistophora mirandellae Vaney and Conte, 1901 and Pleistophora aguillarum Hoshina, 1951 are not species of Pleistophora. Furthermore, the analysis showed that Loma is not a member of Glugeidae Thelohan, 1892. PMID- 10701577 TI - Detection of Echinococcus granulosus coproantigens in Australian canids with natural or experimental infection. AB - Coproparasitological and purging methods for diagnosing canids infected with the intestinal helminth Echinococcus granulosus, an important zoonotic parasite, are unreliable. Detection of coproantigens in feces of infected dogs by enzyme-linked immunosorbent assay (ELISA) is suitable for detecting patent and prepatent infections with a high degree of sensitivity and specificity. In the present study, natural and experimental infections in domestic and wild Australian canids were investigated using a coproantigen capture ELISA. Experimental infection of dogs with E. granulosus was detected at between 14 and 22 days postinfection (PI), and optical density (OD) values remained high until termination of experiments 35 days PI. After chemotherapy, coproantigen levels in infected dogs dropped rapidly, becoming negative 2-4 days after treatment. In experimentally infected red foxes (Vulpes vulpes), the coproantigen excretion profile was different, with ELISA OD levels peaking 15-17 days PI, then falling to low or undetectable levels by 30 days PI. Coproantigens were detected in the feces of naturally infected Australian wild dogs (dingoes, dingo/domestic dog hybrids) with infection levels ranging between 2 worms and 42,600. Preliminary data on the stability of coproantigen in dog feces exposed to environmental conditions indicated that there was no change in antigenicity over 6 days. The results suggest the coproantigen ELISA could be successfully used to monitor E. granulosus prevalence rates in Australian domestic dogs, foxes, and wild dogs. PMID- 10701578 TI - Randomly amplified polymorphic DNA (RAPD) polymerase chain reaction assay for identification of Schistosoma mansoni strains sensitive or tolerant to anti schistosomal drugs. AB - The genetic differences between Schistosoma mansoni strains from different geographic areas that were reportedly resistant or sensitive to anti-schistosomal drugs were studied with randomly amplified polymorphic DNA (RAPD) and simple sequence repeat (SSR) polymerase chain reaction (PCR) assays. Of the 20 RAPD primers we chose, 19 showed the capacity to produce a medium to high level of amplification and 6 revealed difference PCR bands between drug-resistant and drug sensitive strains. One particular primer, 5'-CAGCGACAAG-3', showed 2 major difference bands between praziquantel (PZQ)-resistant and PZQ-sensitive strains from the endemic area of Egypt. These results demonstrate that defined sequence primers could be applied as a useful tool for differentiating drug-resistant and sensitive schistosome parasites in the field. PMID- 10701579 TI - Plagioporus sinitsini (Digenea : Opecoelidae): a one-host life cycle. AB - Adult Plagioporus sinitsini occur within daughter sporocysts voided with the feces of prosobranch snails Elimia symmetrica in Basin Creek, North Carolina. These worms produced eggs containing active miracidia while still in the snail. Adults in snails and adults in rosyside dace, Clinostomus funduloides, collected from the same stream were indistinguishable morphometrically. Adults in snails develop from cotylocercous cercariae sequestered in daughter sporocysts that pass through the metacercaria stage. These observations, and previous study in Michigan, suggest that the life cycle of P. sinitsini has 3 potential pathways, i.e., a 3-host life cycle involving molluscan, arthropod, and piscine hosts, a 2 host life cycle involving only molluscan and piscine hosts, and a 1-host life cycle involving only the snail host. The truncated life cycles do not appear to be the result of paedomorphosis. PMID- 10701580 TI - Arthropod and helminth parasites from the Plains Pocket Gopher, Geomys bursarius bursarius from the hosts' northern boundary range in Minnesota. AB - As part of a continuing and more general study of the diversity of parasites from subterranean mammals in central North America, individuals of the Plains Pocket Gopher, Geomys bursarius bursarius, were collected from 7 localities in northwestern Minnesota from September 1991 through October 1996. Arthropods collected included the fleas, Opisocrostis bruneri (4 of 124, 3.2%), Foxella ignota ignota (85 of 124, 68.5%); the chewing louse, Geomydoecus geomydis geomydis from 98 of 124 (79%), and larvae of the tick, Dermacentor variabilis (1 of 124, 0.8%). Nematodes found included Physaloptera limbata (2 of 118 gophers, 1.7%), Capillaria americana (4 of 118, 3.4%), and Ransomus rodentorum (31 of 118, 26.3%). Cestodes recovered included Anoplocephaloides infrequens (12 of 136 gophers, 8.8%), Anoplocephaloides variabilis (19 of 136, 14%), Andrya macrocephala (20 of 136, 14.7%), and Hymenolepis weldensis from 12 of 136, 8.8%. The acanthocephalan, Moniliformis clarki was found in 1 of 118 gophers (0.8%). No parasites were found in the cheek pouches, thoracic, or peritoneal cavities. PMID- 10701581 TI - The blood parasites of the spiny pocket mouse Liomys salvini (Thomas, 1893) from Costa Rica. AB - A survey of the blood parasites of 20 Liomys salvini revealed 3 types of parasites. Sixty percent of the mice harbored Trypanosoma zeledoni, 5% an unnamed species of Hepatozoon, and 20% of the mice were infected with a species of Haemobartonella. PMID- 10701582 TI - The susceptibility of Lymnaea fuscus to experimental infection with Fasciola hepatica. AB - Three experiments on the infection of Lymnaea fuscus with Fasciola hepatica were carried out to determine if successful infections and maturation of the parasite were dependent on the size of snails at miracidial exposure. The first experiment was performed using 1-4-mm-high snails from 2 populations of L. fuscus and 1 population of Lymnaea palustris. In these snails each subjected to a single bimiracidial exposure, the prevalence of F. hepatica infection at day 35 postexposure ranged from 20.3% to 46.2% in snails measuring 1 mm in height at exposure; it was lower in the 2-mm snails and was 0 in higher size classes. The second experiment was performed by subjecting 1- and 4-mm L. fuscus to 1, 2, and 3 bimiracidial exposures. The prevalence of F. hepatica infection at day 35 postexposure was maximum in the 1-mm snails exposed once to miracidia and decreased with increasing number of exposures. The results were negative in 4-mm snails. Cercarial shedding of F. hepatica was studied in the third experiment using 1- and 2-mm L. fuscus each subjected to a single bimiracidial exposure. The total number of cercariae released from these snails was less than 50. From these results, it can be concluded that L. fuscus showed a partial resistance to F. hepatica infection due to snail age. PMID- 10701584 TI - Determination of the activity of diclazuril against Sarcocystis neurona and Sarcocystis falcatula in cell cultures. AB - Diclazuril is a benzeneacetonitril anticoccidial that has excellent activity against the extraintestinal stages of Toxoplasma gondii and Neospora caninum. It also is highly active against intestinal coccidia of poultry. The present study examined the efficacy of diclazuril in inhibiting merozoite production of Sarcocystis neurona and Sarcocystis falcatula in bovine turbinate cell cultures. Diclazuril inhibited merozoite production by more than 80% in cultures of S. neurona or S. falcatula treated with 0.1 ng/ml diclazuril and greater than 95% inhibition of merozoite production was observed when infected cultures were treated with 1.0 ng/ml diclazuril. Diclazuril may have promise as a therapeutic agent in the treatment of S. neurona-induced equine protozoal myeloencephalitis in horses and S. falcatula infections in birds. PMID- 10701583 TI - Isolation of Sarcocystis speeri Dubey and Lindsay, 1999 parasite from the South American opossum (Didelphis albiventris) from Argentina. AB - Sarcocystis sporocysts from the intestines of 2 opossums (Didelphis albiventris) from Argentina were fed to gamma-interferon knockout (KO) and nude mice. Protozoal schizonts were seen in brain, liver, spleen, and adrenal glands of mice examined 33-64 days after feeding sporocysts. Sarcocysts were seen in skeletal muscles of KO mice 34-71 days after feeding sporocysts. Schizonts and sarcocysts were structurally similar to Sarcocystis speeri Dubey and Lindsay, 1999 seen in mice fed sporocysts from the North American opossum Didelphis virginiana from the United States. PMID- 10701585 TI - Trichobius joblingi, Aspidoptera falcata, and Megistopoda proxima (Diptera : Streblidae) parasitic on Carollia perspicallata and Sturnia lillium (Chiroptera : Phyllostomidae) in southeastern Brazil: sex ratios, seasonality, host site preference, and effect of parasitism on the host. AB - This note examines the effect of parasitism on host size, the preference of the parasite for a specific host body area, and the seasonal abundance for the 3 most abundant bat flies (i.e., Trichobius joblingi Wenzel, a parasite of the bat Carollia perspicillata [Linnaeus], and Aspidoptera falcata Wenzel and Megistopoda proxima [Seguy], parasites on Sturnira lilium [Geoffroy]). Trichobius joblingi and A. falcata are moderately dorsoventrally flattened and were collected on the wing membranes of their hosts, and M. proxima is moderately laterally compressed, has long, thin hind legs, and was collected in the body fur of the host. These 3 parasites also showed distinct seasonal patterns. There was a significant negative correlation between the simultaneous occurrence of A. falcata and M. proxima on the host. Parasitism by M. proxima was correlated with a significant weight loss in male S. lilium, which may reflect the large size, high activity, and constant feeding of this parasite, thereby causing a significant negative effect on the host. Sex ratios favoring male flies could be explained by the tendency of female flies to leave the host immediately before the bat leaves the shelter in search for food or immediately after bats are collected but could also be a consequence of higher mortality among females, especially gravid ones. Finally, collecting may have influenced the skewed sex ratio because male flies, being more active, were more evident to the collector. PMID- 10701587 TI - Evaluation of molecular techniques to biotype Giardia duodenalis collected during an outbreak. AB - Twenty-seven Giardia duodenalis cyst-positive specimens (human, animal, or drinking water) were obtained from a waterborne outbreak in a community in British Columbia, western Canada. Parasite isolates were characterized using molecular techniques at 4 different steps of organism retrieval. None of the drinking water samples (n = 20) infected gerbils and none was successfully amplified using polymerase chain reaction (PCR). We were able to genotype 4 of 7 (human and animal) isolates by amplification of DNA from original specimens at the triosephosphate isomerase (tpi) gene locus using PCR followed by restriction fragment length polymorphism (RFLP) analysis. Five of the original specimens inoculated into Mongolian gerbils (Meriones unguiculatus) were infective and genotyped at the tpi locus using parasite material collected from the gerbil (cysts and trophozoites). Pulsed field gel electrophoresis (PFGE) was used to biotype trophozoites collected from the gerbils as well as trophozoites from the 4 isolates that adapted to culture. Four of these 5 isolates displayed the same (designated outbreak) biotype at all parasite retrieval steps with all molecular techniques including the originally amplified isolates. PCR-RFLP identified an additional biotype group. The 4 isolates that adapted to in vitro culture were also characterized by isoenzyme electrophoresis (IE). Biotype groups identified in these axenized isolates were all the same with each molecular technique (PCR RFLP, PFGE, IE) tested. Results of this study demonstrate a need for more sensitive molecular methods to detect and characterize Giardia in original host and environmental samples. Results are also consistent with evidence of biotype changes that occur during the presently used process of isolate retrieval. PMID- 10701586 TI - Scabies and head-lice infestations in different environmental conditions of Lower Silesia, Poland. AB - A comparison of external parasitic infestations among inhabitants of Legnica, Walbrzych, and Wroclaw districts, in the Lower Silesia region of Poland showed a direct relationship between the high incidence of scabies and low standard ecological indices, as well as social economic setting of the communities. In the years 1990-1997, the highest mean incidences of scabies per 100,000 people (80 and 46) were noted, respectively, in the Legnica and Walbrzych districts, compared to only 7.9 in the Wroclaw district. Infestation was correlated with percentages of the population with higher education (4.8; 4.2, 10.1, respectively) and the number of patients per physician (795, 632, 288, respectively), and the percentages of degraded land/and land threatened by degradation (10/37, 5/16, 0.7/10, respectively), forest stands damaged by gases and particulates (99.4, 99.4, 58.8, respectively), and air pollution emission indices in the towns of Legnica and Walbrzych (30 and 21 tons/km2) and Wroclaw (16). Scabies infestation was highest in children and teenagers (0-19) and was gender-associated (in all age groups, women were more often infested than men). A decreasing rate of scabies infestation, especially from the mid-1990s, was noted for both scabies and pediculosis in Walbrzych district; in the 0-19-yr-old inhabitants, it varied from 0.75% in 1994 to 0.41% in 1996. PMID- 10701588 TI - A chlorodiazirine analog of pentamidine with anti-trypanosomal activity. AB - A chlorodiazirine derivative of pentamidine was synthesized and tested for anti trypanosomal activity using EATRO stock 164 trypanosomes in cell culture. Anti trypanosomal activity was measured as a decrease in [3H]hypoxanthine incorporation by the organisms. The derivative, 3,3'-[1,5-pentanediylbis(oxy-4,1 phenylene)]bis(3-chloro-3H-diazir ine), at a treatment level of 0.1 microM inhibited isotope incorporation by 40-50% compared to nontreated controls. At this concentration, pentamidine inhibited incorporation only 10-15%. The derivative is a nonionic molecule with much different solubility properties than the parent compound and should readily cross the blood-brain barrier. PMID- 10701589 TI - Variation between ASP-1 molecules from Ancylostoma caninum in China and the United States. AB - Hookworm infection continues to be a serious problem in rural areas of China. Rapid reinfection and high cost limit the effectiveness of deworming programs. Vaccination offers an attractive alternative to mass chemotherapy. However, variation in vaccine antigens from field hookworm populations could conceivably limit efficacy of a vaccine developed from laboratory strains. Reported here are initial experiments to ascertain levels of molecular variation in a promising vaccine antigen, ASP-1, from the dog hookworm Ancylostoma caninum. ASP-1 from a Chinese strain of A. caninum was isolated from a third-stage larval cDNA library and compared to ASP-1 from a U.S. strain. There was 97% and 98% similarity in the DNA and amino acid sequences, respectively. There were 42 polymorphic sites between the sequences, 30 of which were synonymous. The 12 nonsynonymous substitutions resulted in 10 changes in the deduced amino acid sequence. Five of the amino acid changes were in the N-terminal domain, whereas the C-terminal domain was more highly conserved, containing only 2 amino acid changes. The results suggest that the effect of molecular variation in antigens from geographically separated parasite populations should be considered during vaccine development. PMID- 10701591 TI - Acanthocephalus tumescens (Acanthocephala, Echinorhynchidae) in Galaxias maculatus (Pisces, Galaxiidae) of Lake Gutierrez, Patagonia, Argentina. AB - The seasonal distribution of Acanthocephalus tumescens (Acanthocephala : Echinorhynchidae) among Galaxias maculatus (Pisces : Galaxiidae) in Lake Gutierrez was studied from March 1994 to June 1996. Acanthocephalus tumescens always occurs in the intestine, has an overdispersed frequency distribution, a similar proportion of sexes, and females are larger than males. Mean intensity and prevalence are low and increase with host length. The pattern of the infection shows seasonality, with recruitment in winter and a reproductive period during spring-summer. PMID- 10701590 TI - Close genotypic relationship between Enterocytozoon bieneusi from humans and pigs and first detection in cattle. AB - The reservoirs and the routes of transmission of Enterocytozoon bieneusi are still unknown. In humans, it is the most commonly found microsporidial species. It has also been found repeatedly in pigs, too. The first detection of E. bieneusi in cattle is reported herein. Two distinct genotypes were characterized and compared with 4 other genotypes from humans, 6 from pigs, and 1 from a cat. From these 13 E. bieneusi genotypes known to date, 25 polymorphic sites could be identified in the internal transcribed spacer of the rRNA gene. The spectrum of polymorphisms within and between each of the 4 host species indicates a close relationship between E. bieneusi strains from humans and pigs, whereas those from cattle are more distantly related. The data suggest the absence of a transmission barrier between pigs and humans for this pathogen. PMID- 10701592 TI - Ectoparasitic insects of bats in British Columbia, Canada. AB - One species of parasitic bug (Hemiptera : Cimicidae), 3 species of fleas (Siphonaptera: Ischnopsyllidae), and 2 species of parasitic flies (Diptera : Nycteribiidae) were collected from 9 species of bats (Chiroptera : Vespertilionidae) in southern interior and northeastern British Columbia, Canada. Female bats that return daily to maternity roosts were more frequently infested with both cimicids and ischnopsyllids than were male bats. Some differences in ectoparasite infestation can be attributed to differences in roosting behavior of the host. New national records for 2 parasite species, and 8 new host records are established for Canada. PMID- 10701593 TI - The longevity of actinosporean spores from oligochaetes of Lake Sasajewun, Algonquin Park, Ontario, and their reaction to fish mucus. AB - The longevity of 7 forms of actinosporean spores and the reaction of 6 forms of actinosporeans to fish mucus were investigated. The maximum longevity of actinosporean spores kept at ambient laboratory temperatures was 14 days. Spore longevity ranged from 11 to 14 days among actinosporeans. The reaction of spores to fish mucus varied among the actinosporeans. Triactinomyxon F of Xiao and Desser, 1998 reacted only to the mucus of the common shiner Luxilus cornutus, and golden shiner Notemigonus crysoleucas, whereas the aurantiactinomyxon form of Xiao and Desser, 1998, and raabeia B of Xiao and Desser, 1998 reacted readily to mucus of all fish species tested. The differences in reaction to fish mucus among actinosporeans may indicate their different host range. These results indicate that actinosporean spores are short-lived and that actinosporeans respond to their hosts by chemodetection. PMID- 10701594 TI - Rules to live by: decision rules. PMID- 10701596 TI - Personal reflections on the growth of diagnostic imaging. PMID- 10701595 TI - Vascular malformations and hemangiomas: a practical approach in a multidisciplinary clinic. PMID- 10701597 TI - Must new radiographs be compared with all previous radiographs, or only with the most recently obtained radiographs? PMID- 10701598 TI - Continuous speech recognition in MR imaging reporting: advantages, disadvantages, and impact. AB - OBJECTIVE: Our objective was to describe our experience with a commercially available continuous speech recognition system, highlighting the advantages, disadvantages, and costs compared with those of conventional transcription for MR imaging reports. MATERIALS AND METHODS: Data from 5072 reports generated in our MR imaging section during a 9-month period after the implementation of a commercial continuous speech recognition system were compared with 4552 reports produced during the same period 1 year earlier. Information pertaining to the use of continuous speech recognition, report turnaround time, word recognition rate, report appearance, and equipment costs was collected. RESULTS: After its system installation, continuous speech recognition was used to dictate 81.8% of all reports. The mean report turnaround time decreased from 87.8 to 43.6 hr, and report availability at 24 hr increased from 10.5% to 62.5%. The system was found to have an average word recognition accuracy of 92.7% for spontaneous dictation. Mean report length declined from 95 to 60 words, with an increase in spacing errors from 0.3 to 8.0 per 1000 words and a decrease in spelling errors from 3.0 to 0.8 per 1000 words. Initial hardware and software costs were approximately $10,000, compared with a yearly cost of $12,000 for human transcription. CONCLUSION: Although the technology is still evolving and was evaluated in its earliest implementation stages, continuous speech recognition nonetheless markedly improved report turnaround time and proved cost-effective. PMID- 10701599 TI - Using gray-scale and color and power Doppler sonography to detect prostatic cancer. AB - OBJECTIVE: We performed a prospective study to assess gray-scale and color and power Doppler sonography for the detection of prostatic cancer and to determine the impact of operator experience. SUBJECTS AND METHODS: Four radiologists with prior experience using gray-scale and Doppler imaging and four urologists with prior experience limited to gray-scale imaging performed sextant biopsies on 251 patients. Each biopsy site was prospectively scored for gray-scale and Doppler abnormality. RESULTS: Cancer was detected in 211 biopsy sites from 85 patients. Overall agreement between sonographic findings and biopsy results as measured with the kappa statistic was minimally superior to chance (kappa = 0.12 for gray scale, kappa = 0.11 for color Doppler, kappa < or =0.09 for power Doppler). With respect to gray-scale diagnosis of cancer, the performance of radiologists (kappa = 0.12) and urologists (kappa = 0.13) was similar. With respect to power Doppler, the performance of radiologists (kappa = 0.09) was superior to that of urologists (kappa = -0.03, p<0.002). Among patients with at least one positive biopsy for cancer, foci of increased power Doppler flow detected by a radiologist were 4.7 times more likely to contain cancer than adjacent tissues without flow. CONCLUSION: Gray-scale and Doppler imaging did not reveal prostatic cancer with sufficient accuracy to avoid sextant biopsy. Power Doppler may be useful for targeted biopsies when the number of biopsy passes must be limited. There is benefit from increased operator experience with Doppler imaging, but there is no demonstrable benefit of power Doppler over conventional color Doppler sonography. PMID- 10701600 TI - Accuracy of normal-dose contrast-enhanced MR angiography in assessing renal artery stenosis and accessory renal arteries. AB - OBJECTIVE: The purpose of this study was to evaluate the accuracy of breath-hold contrast-enhanced MR angiography in the assessment of renal artery stenosis and accessory renal arteries using a standard dose of gadolinium. SUBJECTS AND METHODS: Thirty-eight patients suspected of having renal artery stenosis underwent MR angiography and intraarterial digital subtraction angiography, which was the method of reference. Three-dimensional gradient-echo MR subtraction angiography (TR/TE, 5.8/1.8 msec) was performed on a 1.5-T imager using a phased array body coil. Before imaging, a separate timing bolus sequence was used, administering 1.0 ml of contrast agent. Gadopentetate dimeglumine (15 ml) was injected using an MR power injector. Two observers, who were unaware of each other's interpretation and of MR findings, assessed digital subtraction angiography. Likewise, two other observers assessed MR angiography. RESULTS: Digital subtraction angiography depicted 75 main and 17 accessory renal arteries (n = 92). All main renal arteries and 13 accessory renal arteries were identified on MR angiography. Compared with digital subtraction angiography, MR imaging correctly classified 57 of 66 arteries without a hemodynamically significant stenosis (0-49%), 22 of 22 arteries as significantly stenotic (50-99%), and four of four occluded arteries; five stenoses were overestimated. There was one false positive finding of an accessory renal artery on MR angiography that was identified retrospectively on digital subtraction angiography. Interobserver agreement was high. Sensitivity and specificity for grading significant stenosis were 100% and 85%, respectively. CONCLUSION: Contrast-enhanced MR angiography, using +/-0.1 mmol/kg of gadolinium, is an accurate method in the assessment of renal artery stenosis and accessory renal arteries. PMID- 10701601 TI - MR imaging of the kidneys after laparoscopic cryoablation. AB - OBJECTIVE: We describe the MR imaging findings of patients who underwent laparoscopic renal lesion cryoablation. MATERIALS AND METHODS: Twenty-one patients (men, 11; women, 10; age range, 36-84 years; average age, 65.5 years; SD, 11.9) with 23 small renal masses (< or =4 cm) underwent laparoscopic renal lesion cryoablation. Twenty patients (22 masses) underwent follow-up MR imaging on the first day after surgery, 12 (13 masses) at 1 month, 16 (18 masses) at 3 months, 14 (15 masses) at 6 months, and 12 (12 masses) at 12 months. Three radiologists retrospectively reviewed MR images for the signal intensity, characteristics, and size of cryolesions. CT-guided needle biopsy was performed 6 months after cryoablation (18 patients) and no evidence of malignancy was discovered. RESULTS: Including all lesions at all times on T1-weighted images, cryolesion signal intensity was isointense to renal parenchyma (47/76, 61.8%) or isointense with hyper- or hypointense foci (7/76, 9.2%). On T2-weighted images, almost all lesions (72/76, 94.7%) were isointense or hypointense, and there was a hypointense rim between the cryolesion and renal parenchyma in 38.2% of lesions (29/76). A thin peripheral rim of enhancement was noted in 19.7% (14/74) of lesions. Cryolesions decreased in size an average of 61.5% (SD, 22.82; n = 12) at 1 month, 78.7% (SD, 13.5; n = 17) at 3 months, 83.5% (SD, 24.3; n = 15) at 6 months, and 94.2% (SD, 8.1; n = 11) at 1 year after cryoablation (one patient was not scanned 1 day after cryoablation and was not included in our calculations). CONCLUSION: After renal cryoablation, MR imaging revealed common signal characteristics such as low-signal-intensity rims on T2-weighted images, enhancement patterns such as thin peripheral rims, and interval size changes. PMID- 10701602 TI - Sonography of obstetric and gynecologic emergencies: Part I, Obstetric emergencies. PMID- 10701603 TI - Sonography of obstetric and gynecologic emergencies: Part II, Gynecologic emergencies. PMID- 10701604 TI - MR-based three-dimensional modeling of the normal pelvic floor in women: quantification of muscle mass. AB - OBJECTIVE: Our objective was to use a combination of axial MR source images and three-dimensional (3D) models to describe the anatomy of the normal pelvic floor in young nulliparous women and to measure the volume of the levator ani. SUBJECTS AND METHODS: Ten healthy nulliparous female volunteers (average age, 27 years) underwent T2-weighted MR imaging of the pelvis. Three-dimensional color-coded models of the pelvic bones and organs and the three major components of the levator ani--puborectalis, iliococcygeus, and coccygeus--were created. Source images were used to measure muscle width and signal intensity and to identify ligamentous structures. Using 3D models, we measured the volume of the levator ani, the angle of the levator plate, the posterior urethrovesical angle, and the distance of the bladder neck from the symphysis pubis and the pubococcygeal line. RESULTS: In all volunteers, the signal intensity of the puborectalis exceeded that of the obturator externus. The average volume of the levator ani was 46.6 ml, the average width of the levator hiatus was 41.7 mm, and the average posterior urethrovesical angle was 143.5 degrees. Vaginal shape in the volunteers followed no recognizable pattern. CONCLUSION: Muscle morphology, signal intensity, and volume is relatively uniform among healthy young women. PMID- 10701605 TI - Dynamic MR imaging of the pelvic floor in asymptomatic subjects. AB - OBJECTIVE: Dynamic MR imaging may be used as an alternative to dynamic cystoproctography for the evaluation of pelvic floor prolapse and configuration. MR criteria for normality are derived from proctographic studies because no large MR study of asymptomatic individuals has been performed. Our study aimed to define the normal range of dynamic pelvic MR appearances in a large group of asymptomatic individuals. SUBJECTS AND METHODS: Fifty healthy adult volunteers (25 men and 25 women; age range, 20-66 years; mean age, 34 years) were prospectively recruited and examined using dynamic MR imaging. All subjects were interviewed and established as healthy using a validated questionnaire. Axial, coronal, and sagittal MR imaging was performed at rest and during maximum pelvic strain using a static 1.0-T unit and a fast-field-echo sequence, providing 10 slices in 31 sec. Standardized measurements of pelvic configuration were taken, and rest and strain imaging were compared to determine the range of normal appearances. RESULTS: Three women developed a cystocele during maximum pelvic strain, two of whom also showed grade 1 uterocervical prolapse, which was also seen in another woman. Three men showed posterior pelvic floor descent in excess of 3 cm during straining. No rectocele, enterocele, rectal prolapse, or perineal hernia was seen in any subject. CONCLUSION: The normal range of pelvic organ descent in asymptomatic subjects seen on dynamic MR imaging included cystocele, uterocervical prolapse, and excessive anorectal junction descent. Although we encountered pelvic prolapse in seven volunteers, it was infrequent and low grade, suggesting that criteria for abnormality derived from proctography are generally applicable to MR imaging. PMID- 10701606 TI - Renal carcinoma presenting with flank pain: a potential drawback of unenhanced CT. PMID- 10701607 TI - Imaging findings in pancreatic lymphoma: differential aspects. AB - When the radiologist is faced with a well-circumscribed tumoral mass in the pancreas, knowing when to direct the patient toward nonsurgical biopsy instead of surgical biopsy and staging is critical. Lymphoma does not require surgical staging or a palliative Whipple's procedure before chemotherapy or radiation therapy. A better overall prognosis with nonsurgical treatment is additional impetus to search for secondary signs of primary pancreatic lymphoma. In patients with primary pancreatic lymphoma, no marked pancreatic ductal dilatation is present even with ductal invasion. Adenocarcinoma commonly dilates the more distal pancreatic duct when more proximal ductal invasion has taken place. Lymph node involvement below the level of the renal veins was another finding not seen with adenocarcinoma. Clinical and imaging findings are otherwise not specific in the differentiation of pancreatic lymphoma and pancreatic cancer, but a bulky homogeneous tumoral mass without alteration of Wirsung's duct or the peripancreatic vessels should suggest the diagnosis. In patients with diffuse infiltration of the pancreatic gland without clinical signs of pancreatitis, the radiologist should be alert to the possibility of pancreatic lymphoma. PMID- 10701609 TI - The ileosigmoid knot: CT findings. PMID- 10701608 TI - CT assessment of the inferior peripancreatic veins: clinical significance. AB - OBJECTIVE: The purpose of this study was to evaluate and clarify the clinical significance of CT scans of the inferior peripancreatic veins. MATERIALS AND METHODS: Forty-three patients with suspected pancreatic disease underwent three phase helical CT (collimation, 5 mm; reconstruction, 2.5 mm; scan delay, 30, 60, and 150 sec). The frequency of visualization on CT of the anterior and posterior inferior pancreaticoduodenal veins, inferior pancreaticoduodenal vein, and first jejunal trunk was assessed and correlated with angiographic and pathologic findings. RESULTS: The frequency of visualization of normal inferior peripancreatic veins in patients (n = 22) with a normal portomesenteric vein was 36% for the anteroinferior pancreaticoduodenal vein, 36% for the posteroinferior pancreaticoduodenal vein, 59% for the inferior pancreaticoduodenal vein, and 100% for the first jejunal trunk. The smaller inferior peripancreatic veins were frequently not visualized when normal. In patients (n = 13) with pancreatic carcinoma involving the portosuperior mesenteric vein, all of the inferior peripancreatic veins were dilated and easily recognizable. When the tumor did not involve the portosuperior mesenteric vein but did involve the anteroinferior pancreaticoduodenal, posteroinferior pancreaticoduodenal, and inferior pancreaticoduodenal veins (n = 8), some of the other peripancreatic veins (first jejunal trunk, anterior and posterior superior pancreaticoduodenal veins, and gastrocolic trunk) were dilated. Dilatation indicated tumor extension to the third portion of the duodenum. In patients (n = 7) with involvement of the inferior pancreaticoduodenal vein, the first jejunal trunk, or both without the involvement of the portosuperior mesenteric vein, dilatation of the other peripancreatic veins (anteroinferior pancreaticoduodenal vein, posteroinferior pancreaticoduodenal vein, anterosuperior pancreaticoduodenal vein, posterosuperior pancreaticoduodenal vein, and gastrocolic trunk) indicated tumor invasion of only the second portion of the extrapancreatic nerve plexus (n = 4) and tumor invasion of both the second portion of the extrapancreatic nerve and the mesenteric root (n = 3). CONCLUSION: Dilatation of peripancreatic veins with nonvisualization of inferior peripancreatic veins suggests tumor invasion of peripancreatic tissue. PMID- 10701610 TI - MR enteroclysis: evaluation of small-bowel obstruction in a patient with pseudomyxoma peritonei. PMID- 10701611 TI - CT and MR imaging of hepatic metastases. PMID- 10701612 TI - Perihepatic lymph nodes as a marker of antiviral response in patients with chronic hepatitis C infection. AB - OBJECTIVE: In patients with chronic hepatitis C, the sonographically determined total perihepatic lymph node volume reflects liver histology and viremia. The aim of this prospective study was to assess whether the response to antiviral therapy is reflected by changes in the total perihepatic lymph node volume. SUBJECTS AND METHODS: In 59 patients with chronic hepatitis C infection. the total perihepatic lymph node volume was assessed using sonography before the initiation of antiviral treatment, at the end of treatment, and at the end of a 6-month follow up period. Hepatitis C viremia was assessed by reverse transcription-polymerase chain reaction assay at the same time points. Liver biopsy was performed in all patients before therapy and in 40 of the 59 patients 6 months after therapy. RESULTS: At the end of follow-up, the total perihepatic lymph node volume was significantly smaller in the 15 patients with a sustained virologic response than in the 44 patients who failed to respond to treatment (0.5+/-0.3 ml versus 2.0+/ 1.2 ml; p<0.0001). In the group of sustained virologic responders, the decline of the perihepatic lymph node volume was associated with an improvement in liver histology. CONCLUSION: Total perihepatic lymph node volume changes according to the antiviral response and leads to progressive normalization of the perihepatic lymph node volume in sustained virologic responders. A decrease in the perihepatic lymph node volume is associated with an improvement in liver histology. PMID- 10701613 TI - CT of focal nodular hyperplasia of the liver. PMID- 10701614 TI - Cervical spine injury: a clinical decision rule to identify high-risk patients for helical CT screening. AB - OBJECTIVE: We aimed to validate the routine use of a clinical decision rule to direct diagnostic imaging of adult blunt trauma patients at high risk for cervical spine injury. MATERIALS AND METHODS: We previously developed and have since routinely used a prediction rule based on six clinical parameters to identify patients at greater than 5% risk of cervical spine injury to undergo screening helical CT of the cervical spine. During a 6-month period, 4285 screening imaging studies of the cervical spine were performed in adult blunt trauma patients. Six hundred one patients (398 males, 203 females; age range, 16 100 years; median age, 38 years) underwent helical CT, and the remainder underwent 3684 conventional radiographic examinations. Clinical and report data were extracted from the radiology department database, medical records, and the hospital trauma registry. Abnormal findings were independently confirmed by additional imaging studies, autopsy results, or clinical outcome. RESULTS: The true-positive cervical spine injury rates in helical CT- and conventional radiography-screened patients who presented directly to our trauma center were 40 (8.7%) of 462 and seven (0.2%) of 3684, respectively. The cervical spine injury rate in patients who were transferred from outside institutions to our trauma center and who underwent helical CT was 37 (26.6%) of 139. This figure included 20 patients already known to have cervical spine fracture. CONCLUSION: The clinical decision rule can distinguish patients at high and low risk of cervical spine injury, thus supporting its validity. PMID- 10701615 TI - MR imaging of cyclops lesions. AB - OBJECTIVE: Localized anterior fibrosis (cyclops lesion) is a known cause of extension loss of the knee after anterior cruciate ligament (ACL) reconstruction. We describe MR imaging as a noninvasive diagnostic tool to examine cyclops lesions. SUBJECTS AND METHODS: Thirty-three MR studies of 31 patients with residual persistent extension loss after ACL reconstruction using patellar tendon autograft were reviewed and compared with results of second arthroscopy. We used MR imaging to describe the ACL graft signal intensity and course, tibial and femoral tunnel placement. quantitative measurements of notch size and shape, and the presence or absence of cyclops lesions. When a cyclops lesion was revealed on MR imaging, the signal-intensity characteristics, location, and size were documented. Preoperative MR imaging findings were then correlated with findings at arthroscopy. RESULTS: The sensitivity, specificity, and accuracy of revealing a cyclops lesion on MR imaging were 85.0%, 84.6%, and 84.8%, respectively. We found no statistically significant differences in the size of intercondylar notches for patients with and patients without cyclops lesions. CONCLUSION: MR imaging was sensitive, specific, and accurate in revealing cyclops lesions in a subgroup of patients with extension loss after ACL reconstruction. PMID- 10701616 TI - Sacral stress fractures in long-distance runners. AB - OBJECTIVE: Sacral stress fractures in athletes are rare but important to recognize because the symptoms often mimic sciatica and can lead to delay in diagnosis and treatment. The radiographic findings are characteristic and can facilitate early diagnosis and lead to appropriate treatment. CONCLUSION: The clinical presentation of runners with sacral stress fractures can mimic disk disease. However, stress fractures in athletes, especially long-distance runners, must be treated differently. The imaging characteristics appear as linear abnormal signal intensity paralleling the sacroiliac joint on MR imaging and linear sclerosis with cortical disruption on CT. Imaging with bone scintigraphy shows increased uptake that parallels the sacroiliac joint. PMID- 10701617 TI - Analysis of diffusion changes in posttraumatic bone marrow using navigator corrected diffusion gradients. AB - OBJECTIVE: This study was undertaken to analyze diffusion characteristics of normal and posttraumatic bone marrow. MATERIALS AND METHODS: Fifty consecutive patients with knee pain underwent both conventional and diffusion-weighted MR imaging (b values, 0-980 sec/mm2). Diffusion maps derived from source data were analyzed on a workstation using region-of-interest techniques. Apparent diffusion values recorded in normal marrow were compared with values recorded in abnormal posttraumatic bone marrow (square centimeters per second). RESULTS: Normal bone marrow identified in 35 patients showed minimal diffusion, with a mean value of 0.15x10(-5) cm2/sec. Bone marrow in 15 patients sustaining direct traumatic injury (21 bone bruises) showed markedly increased diffusion, with a mean value of 0.8x10(-5) cm2/sec (range, 0.4-1.3 cm2/sec). CONCLUSION: Marrow injury after trauma with trabecular damage allows increased movement or diffusion of interstitial water relative to normal marrow. The magnitude of diffusion change appears to reflect the severity of marrow injury. PMID- 10701618 TI - Correlation between bone marrow edema and collapse of the femoral head in steroid induced osteonecrosis. AB - OBJECTIVE: The purpose of this study was to clarify whether bone marrow edema is detectable on initial MR imaging of steroid-induced osteonecrosis of the femoral head. SUBJECTS AND METHODS: Forty-eight hips with osteonecrosis were examined consecutively with MR imaging and radiography. In a previously reported screening program, osteonecrosis was diagnosed on MR imaging when subchondral bands of abnormal signals were present. In the screening program, the MR images of 200 hips of 100 patients receiving high-dose steroid therapy were examined prospectively. Subchondral bands were detected in 48 hips at a mean of 14 weeks after the initiation of steroid therapy. RESULTS: On follow-up MR imaging of 47 hips (one hip excluded) bone marrow edema was initially observed in 13 hips after the onset of hip pain. MR imaging of the remaining 34 hips did not reveal bone marrow edema and the patients were all asymptomatic. MR imaging of 31 of the 34 hips continued to show subchondral bands and MR imaging of the other three hips indicated that the subchondral bands had disappeared. When bone marrow edema was detectable, abnormal findings on radiography were slight but 11 (85%) of the 13 hips progressed to advanced osteonecrosis. Bone marrow edema was highly correlated with the subsequent collapse of the femoral head (p<0.0001). CONCLUSION: Bone marrow edema was not present on initial MR imaging of osteonecrosis. Bone marrow edema should be considered a marker for potential progression to advanced osteonecrosis, and careful examinations for osteonecrosis are necessary when bone marrow edema is seen. PMID- 10701619 TI - Efficacy of step-oblique mammography for confirmation and localization of densities seen on only one standard mammographic view. AB - OBJECTIVE: Step-oblique mammography is a technique used to determine with confidence whether a mammographic finding visible on multiple images on only one projection (but not elucidated using standard additional mammographic projections such as the roll view) represents a summation artifact or a true mass, and to precisely localize the true mass for further evaluation (if applicable). This paper describes the step-oblique technique and evaluates its efficacy. MATERIALS AND METHODS: Between January 1, 1993 and December 31, 1998, 69 consecutive women underwent step-oblique mammography for the evaluation of densities seen on multiple images in only one standard projection. Additional images were obtained at 15 degrees stepped increments in obliquity. If a one-projection-only finding was not seen on step-oblique images, the density was judged to represent a summation artifact, completing the examination. If a density was visualized and could be triangulated concordantly on step-oblique images ranging from the craniocaudal to the 90 degrees lateral projection, then it was judged to represent a real lesion. Such a lesion was further characterized (mass, neodensity, architectural distortion, focal asymmetric density) and was localized precisely in three dimensions, permitting imaging-guided tissue diagnosis, if appropriate. For all study patients, we also recorded BI-RADS (American College of Radiology Breast Imaging and Data Reporting System) assessment categories; pathology results for biopsied lesions; and mammographic follow-up, clinical follow-up, and linkage to regional tumor registry for nonbiopsied lesions for which at least 2 years had elapsed since step-oblique mammography. RESULTS: Step oblique mammography differentiated 50 real lesions from 19 summation artifacts. All 50 real lesions, although initially visible on only one standard projection, were successfully localized in three dimensions. Subsequent management resulted in the prompt detection and diagnosis of seven breast cancers and 21 benign lesions. None of the remaining findings managed by follow-up rather than biopsy have subsequently been found to be malignant. CONCLUSION: Step-oblique mammography is an effective means of evaluating the mammographic finding visible on multiple images on only one standard projection. PMID- 10701620 TI - Mammographic abnormalities caused by percutaneous stereotactic biopsy of histologically benign lesions evident on follow-up mammograms. AB - OBJECTIVE: The purpose of our study was to evaluate how often a mammographic abnormality thought to be produced by the biopsy procedure was evident on the initial follow-up mammogram after percutaneous biopsy of impalpable histologically benign lesions. We compared three stereotactic percutaneous biopsy methods. CONCLUSION: A mammographic density seen well only in the projection parallel to the biopsy needle tract was evident in 2% (5/226) of the lesions for which 11-gauge directional vacuum-assisted biopsy was used, 0% (0/96) of the lesions for which 14-gauge directional vacuum-assisted biopsy was used, and 0% (0/422) of the lesions for which 14-gauge automated large-core biopsy was used. No mammographic abnormalities assessed as BI-RADS categories 3, 4, or 5 (according to the American College of Radiology's Breast Imaging Reporting and Data System) and thought to be produced by the biopsy procedure were evident after any of the biopsy methods. PMID- 10701621 TI - Tubular adenomas of the breast: imaging findings with histologic correlation. AB - OBJECTIVE: The purpose of this study is to describe the imaging features of tubular adenomas, which are rare benign breast tumors usually found in women younger than 35 years old. CONCLUSION: In young women, tubular adenomas can look like noncalcified fibroadenomas on mammography and sonography. In older women, tubular adenomas may resemble malignant masses with microcalcifications. Awareness of these findings may help in assessing concordance between imaging and histologic findings after percutaneous core biopsy of these rare lesions. PMID- 10701622 TI - Evolution of peripheral lung adenocarcinomas: CT findings correlated with histology and tumor doubling time. AB - OBJECTIVE: This study was performed to evaluate the evolution of peripheral lung adenocarcinomas using CT findings and histologic classification related to tumor doubling time. MATERIALS AND METHODS: The subjects were 34 patients, each with an adenocarcinoma smaller than 3 cm. All patients underwent chest radiography and 10 of them had previously undergone CT more than 6 months before surgery. Tumor doubling time was estimated by examining sequential radiographs using the method originally described by Schwartz. Tumor growth was also observed by studying the changes on CT in the 10 patients who had previously undergone CT. The histologic classification (types A-F) was evaluated according to the criteria of Noguchi et al. RESULTS: Five (83%) of the six adenocarcinomas with tumor types A or B showed localized ground-glass opacity on high-resolution CT. All six tumors had a tumor doubling time of more than 1 year. Fifteen (71%) of the 21 tumors with type C showed partial ground-glass opacity mixed with localized solid attenuation on high-resolution CT. Ten (48%) of these 21 type C tumors had a tumor doubling time of more than 1 year. In types B and C, the solid component or the development of pleural indentation and vascular convergence increased during observation before surgery. All seven tumors with types D, E, and F showed mostly solid attenuation, and the tumor doubling time was less than 1 year in six (87%) of the seven tumors. CONCLUSION: Two main types of peripheral lung adenocarcinoma exist. The first type appears on CT as a localized ground-glass opacity with slow growth, and the other appears as a solid attenuation with rapid growth. PMID- 10701623 TI - Prognostic value of thoracic FDG PET imaging after treatment for non-small cell lung cancer. AB - OBJECTIVE: We determined the prognostic value of 18F-fluorodeoxyglucose (FDG) positron emission tomography (PET) for patients with treated lung cancer. MATERIALS AND METHODS: We examined patients who underwent FDG PET after first line treatment for non-small cell lung cancer. FDG PET results were correlated with survival rates to determine whether FDG PET findings were predictive of outcomes. RESULTS: After initial therapy, 113 patients with non-small cell lung cancer underwent FDG PET. One hundred patients had positive FDG PET results and a median survival of 12 months (95% confidence interval, 9.2-15.4). Thirteen patients had negative FDG PET results, and 11 (85%) of these patients are still living at a median follow-up of 34 months. The difference in survival for patients with positive and negative FDG PET results was statistically significant (p = 0.002). CONCLUSION: FDG PET has prognostic value and strongly correlates with survival rates of patients with treated lung cancer. Patients with positive FDG PET results have a significantly worse prognosis than patients with negative results. Additionally, FDG PET may be helpful in guiding therapeutic treatments. PMID- 10701624 TI - CT depiction of regional nodal stations for lung cancer staging. PMID- 10701625 TI - Using thoracic helical CT to assess iodine concentration in a small volume of nonionic contrast medium during vascular opacification: a prospective study. AB - OBJECTIVE: The goal of the study was to assess whether, using thoracic helical CT, diagnostic mediastinal and hilar vascular enhancement can be obtained with a small amount of nonionic contrast material (80 ml) injected at a relatively slow rate (2 ml/sec). SUBJECTS AND METHODS: One hundred twenty patients (60 in their fourth decade of life and 60 in their seventh decade of life) referred for contrast-enhanced thoracic CT for malignancies or infections prospectively entered the study. They were randomly assigned to be given one of three iodine concentrations of a nonionic contrast material: 250 mg/ml (1250), 300 mg/ml (1300), and 350 mg/ml (1350). Two radiologists independently graded perivenous artifacts and arterial enhancement of mediastinal and hilar vessels on a 4-point scale: 1, poor; 2, fair; 3, good; and 4, excellent. Measurements of arterial attenuation values (quantitative assessment) were obtained on the aorta and pulmonary artery. RESULTS: Mean scores were equal to or greater than 3 for all vessels only using 1350. The higher the iodine concentration was, the higher the mean score, but there was a statistically significant difference only between scores obtained with 1350 and those obtained with 1300 or 1250. Mean scores were higher for the patients in their seventh decade of life than those in their fourth decade; however, there was no statistically significant difference between scores of the two decade groups. We found a highly significant statistical relationship between scores and arterial attenuation values. CONCLUSION: During contrast-enhanced helical CT examinations for general thoracic evaluations, good opacification of central vascular structures is obtained with a low volume of high iodine concentration nonionic contrast medium. PMID- 10701626 TI - Talcosis associated with IV abuse of oral medications: CT findings. AB - OBJECTIVE: Our objective was to evaluate the CT appearance of talcosis associated with IV abuse of oral medications and to compare the findings of talcosis related to methylphenidate with those findings seen with other drugs. MATERIALS AND METHODS: The CT scans of 12 patients with talcosis (seven men, five women), 33-54 years old (mean age, 44 years), were analyzed retrospectively. Seven patients had abused methylphenidate; five patients had no history of abuse. The diagnosis of talcosis was made histologically in 11 patients and at funduscopy in one patient. CT was performed with 1- to 1.5-mm collimation (n = 10 patients) or 5- to 10-mm collimation (n = 2). RESULTS: The predominant abnormalities seen on CT consisted of a diffuse fine nodular pattern (n = 2), a combination of nodules and lower lobe panacinar emphysema (n = 3), and ground-glass attenuation (n = 2). Emphysema was the only abnormality seen in the remaining five patients (lower lobe panacinar, n = 4; upper lobe centrilobular, n = 1). No significant difference in the prevalence of nodules and ground-glass attenuation was seen between the methylphenidate and non-methylphenidate groups. Lower lobe panacinar emphysema was more common in methylphenidate abusers (six [86%] of seven patients) than in non-mnethylphenidate drug abusers (one [20%] of five, p<0.05, Fisher's exact test). CONCLUSION: The CT manifestations of talcosis consist of a fine micronodular pattern, ground-glass attenuation, and emphysema. A significantly increased prevalence of lower lobe panacinar emphysema is seen in IV drug addicts who abuse methylphenidate. PMID- 10701627 TI - Semiinvasive pulmonary aspergillosis: CT and pathologic findings in six patients. AB - OBJECTIVE: We describe the chest CT and pathologic findings of semiinvasive pulmonary aspergillosis in six patients. CONCLUSION: Semiinvasive pulmonary aspergillosis should be considered in the mildly immunocompromised patient with CT findings that reveal persistent parenchymal abnormalities. Patterns include consolidation and mass. PMID- 10701628 TI - Psyllium aspiration causing bronchiolitis: radiographic, high-resolution CT, and pathologic findings. PMID- 10701629 TI - Interscan variation in coronary artery calcium quantification in a large asymptomatic patient population. AB - OBJECTIVE: We evaluated interscan variation in coronary artery calcium scores in a large screening population as determined by electron beam CT. MATERIALS AND METHODS: One thousand patients (average age, 53 years; age range, 18-85 years) who were asymptomatic for coronary artery disease underwent two consecutive scans of the heart on an electron beam CT scanner. Scans were performed with ECG gating, breath-hold, 3-mm collimation, and 100-msec exposure. Two contiguous pixels with density values greater than 130 H were used as the minimum criterion for a calcific lesion. The calcium score was determined on a vessel-by-vessel basis for both scans of each patient. Interscan variation in calcium and vessels involved with calcification was evaluated on the basis of age, sex, and average calcium score. RESULTS: The percentage of difference between calcium scores in scans was 28.4% and 43.0% for women and men, respectively. For the individual epicardial arteries (left main, left anterior descending, circumflex, and right coronary), the percentage of difference for calcium scores was 20.2-24.2% for women and 30.5-44.9% for men. A difference between the two scans in at least one vessel of the total coronary arteries identified with calcium was noted in 31% of patients. CONCLUSION: Interscan variability in calcium scores may be important in the determination of risk stratification. Subjects with a nonzero calcium score may benefit from undergoing two scans at the time of initial imaging. PMID- 10701630 TI - Angiography of leaks after endovascular repair of infrarenal aortic aneurysms. AB - OBJECTIVE: We examined whether leaks that persist after stent grafting are associated with outflow arteries. SUBJECTS AND METHODS: Selective angiography was performed in 21 patients with persistent leaks after undergoing endovascular repair of infrarenal aneurysms of the abdominal aorta. Late leaks occurred in five patients whose prostheses were originally sealed. Before angiography, the size and position of leaks were determined with CT and color Doppler sonography. RESULTS: Superselective angiography was successful in 19 of 21 patients. In two patients, angiography was performed over the afferent artery supplying the leak. We found one outflow artery at the site of the leak in 10 patients (47%); two outflow arteries in five (23.8%); and as many as five outflow arteries in three (14%). Angiography overlooked outflow arteries in three patients (14%). The lumbar and inferior mesenteric, urethral, and testicular arteries were identified as outflow arteries. CONCLUSION: Other than feeder arteries, persistent leaks are associated with outflow vessels that contribute to the patency of leaks. PMID- 10701631 TI - Vertebral artery Doppler waveform changes indicating subclavian steal physiology. AB - OBJECTIVE: The goal of this study was to characterize and classify changes in antegrade vertebral artery waveforms that may represent the early stages of subclavian steal physiology. SUBJECTS AND METHODS: A prospective examination of waveforms from 1914 vertebral arteries produced a total of 40 that had a transient sharp decline in velocities at mid or late systole. In these patients, an ECG tracing was synchronized with the pulsed Doppler waveform, and reactive hyperemia was induced in the ipsilateral arm with a blood pressure cuff. The same protocol was performed in a control group of 52 patients with normal vertebral artery waveforms. Correlation between the waveforms and subclavian disease shown on angiography was made in 10 cases collected from the prospective study and in an additional 10 cases identified from a record search. RESULTS: Four prototypic waveforms were identified on the basis of the degree of flow deceleration in mid systole. Flow velocity at the nadir of the mid systolic notch was greater than that of the end diastole for type 1 waveforms, equal to the end diastole for type 2, at the baseline for type 3, and below the baseline for type 4. The blood pressure cuff maneuver induced a change to more abnormal waveforms in 36 of 40 patients but did not change the waveforms of the control group. The correlation between waveform type and subclavian disease was statistically significant (p = 0.03). CONCLUSION: Identifiable changes in the pulse contour of antegrade vertebral artery waveforms seem to represent the early stages of subclavian steal physiology. These changes can be organized into waveform types that indicate increasingly abnormal hemodynamics. PMID- 10701632 TI - Usefulness of CT angiography with volume rendering after carotid angioplasty and stenting. PMID- 10701633 TI - Innovative catheter fixation using a low-profile device. PMID- 10701634 TI - CT and MR imaging of generalized cystic lymphangiomatosis in pediatric patients. AB - OBJECTIVE: The aim of this study was to describe the spectrum of abnormalities seen in generalized cystic lymphangiomatosis as shown by CT and MR imaging and to correlate these findings to gross pathology. CONCLUSION: MR imaging and CT may substantially broaden visualization of the spectrum of abnormalities seen in generalized cystic lymphangiomatosis by revealing the complete extent of disease and, thus, may contribute to clinical management of the disease by preventing initial misdiagnosis. PMID- 10701635 TI - MR-guided biopsy using respiratory-triggered high-resolution T2-weighted sequences. PMID- 10701636 TI - Imaging-based nodal classification for evaluation of neck metastatic adenopathy. AB - OBJECTIVE: This study was undertaken to create an imaging-based classification for the lymph nodes of the neck that will be readily accepted by clinicians, result in consistent nodal classification, and be easily used by radiologists. SUBJECTS AND METHODS: Over an 18-month period, the necks of 50 patients with cervical lymphadenopathy were scanned with CT, MR imaging, or both. Imaging anatomic landmarks were sought that would create a nodal classification of these necks similar to the clinically based nodal classifications of the American Joint Committee on Cancer and the American Academy of Otolaryngology-Head and Neck Surgery. Each nodal level was defined to ensure consistent nodal classification and eliminate areas of confusion existing in the clinically based classifications. RESULTS: Necks were classified using the imaging-based classification and then compared with the classification of the same necks using the most common clinically based classifications. The imaging-based nodal classifications of the superficial nodes were the same as the clinically based classifications; however, the deep nodes of eight patients were found only by imaging. The anatomic precision and the level definition afforded by sectional imaging allowed the radiologists to use the imaging-based classification in a consistent manner. CONCLUSION: This imaging-based classification has been endorsed by clinicians who specialize in head and neck cancer. The boundaries of the nodal levels were easily discerned by radiologists and yielded consistent nodal classifications. The reproducibility of this classification will allow it to become an essential component of future classifications of metastatic neck disease. PMID- 10701637 TI - Epistaxis: vascular anatomy, origins, and endovascular treatment. AB - Embolization can play an important role in controlling epistaxis. However, one must be careful to avoid nontarget embolization via the dangerous anastomoses between the ECA branches, the carotid siphon, and ophthalmic arteries. PMID- 10701638 TI - Cerebral infarctions: evaluation with single-axis versus trace diffusion-weighted MR imaging. AB - OBJECTIVE: Our purpose was to determine the usefulness of single-axis diffusion weighted imaging versus trace diffusion-weighted imaging in the evaluation of cerebral infarctions. SUBJECTS AND METHODS: Twenty-six patients harboring 34 infarctions were examined using single-axis and trace diffusion-weighted imaging within 48 hr of the onset of symptoms. Two neuroradiologists who were not aware of the clinical findings reviewed all images obtained with both techniques and noted the following: type of infarction (small [<15 mm] versus territorial), location of infarction, presence of infarction (seen only on single-axis images, seen only on trace images, seen on both), lesion conspicuity (better on single axis images, better on trace images, or equal on both), and lesion size (larger on single-axis images, larger on trace images, or equal on both). Differences in opinion were resolved by consensus. RESULTS: Of the 18 small and 16 territorial infarctions, all were identified on both single-axis and trace imaging. Lesion conspicuity was judged to be slightly better on trace images for both types of infarctions. Lesion size was judged to be larger on single-axis images for territorial infarctions. CONCLUSION: Both single-axis and trace diffusion weighted imaging showed all small and territorial cerebral infarctions. Both types of infarctions were slightly larger on single-axis images but this did not affect correct interpretation in any case. The single-axis technique provided sufficient information for the diagnosis of cerebral infarction in our clinical settings. PMID- 10701639 TI - Comparison of patient age with MR imaging features of gangliogliomas. AB - OBJECTIVE: The purpose of this study was to compare MR imaging features of gangliogliomas in children less than 10 years old with those seen in patients at least 10 years old. MATERIALS AND METHODS: Our study population consisted of 15 female patients and 10 male patients with a mean age of 20 years. The early childhood group was composed of six children with a mean age of 5.5 years. The older group was composed of 19 patients with a mean age of 25.6 years. We assessed tumor volume, tumor location, percentage of tumor that was cystic, pattern of contrast enhancement, and degree of edema. RESULTS: The temporal lobe was the most common tumor location in both groups. Mean tumor volume in the early childhood group was 83 cm3, which was significantly larger than the mean tumor volume (9.78 cm3) for the older group (p = 0.001). Cystic tumors were more common in the early childhood group (83%) than in the older group (63%), and the average percentage of cysts in the cystic tumors was much higher in the early childhood group (67%) than in the older group (30%). Contrast enhancement was seen in five of six early childhood tumors and 13 of 16 tumors in older patients. Four of six tumors in the early childhood group and five of 19 tumors in the older patient group had associated edema. CONCLUSION: The mean tumor volume of gangliogliomas in the early childhood group was significantly larger than that of the older patient group. This finding may be indicative of differences in tumor growth patterns in the two groups, ability of the hemicranium to adjust to mass effect in childhood, or sampling error as a result of a relatively small sample size. PMID- 10701640 TI - MR findings in subacute combined degeneration of the spinal cord: a case of reversible cervical myelopathy. PMID- 10701641 TI - When doing a small-bowel series, what is considered a normal transit time for barium to reach the cecum? PMID- 10701642 TI - What is the natural history of posttraumatic osteochondritis dissecans of the talus? PMID- 10701643 TI - Professional courtesy: no thanks. PMID- 10701644 TI - Peritumoral sparing of fatty liver: another important instance of focal sparing caused by a hepatic tumor. PMID- 10701645 TI - Focal fibrosis of the breast in diabetes. PMID- 10701646 TI - Negative findings on helical CT in suspected pulmonary embolism. PMID- 10701647 TI - Notice of duplicate publication, AJR 1999;172:35-37. PMID- 10701648 TI - Dermoid cyst in the lumbosacral region: radiographic findings. PMID- 10701649 TI - Erdheim-Chester disease involving bilateral lower extremities: MR features. PMID- 10701650 TI - Acquired spontaneous intercostal hernia of the lung diagnosed on helical CT. PMID- 10701651 TI - Extrahepatic portal vein aneurysms. PMID- 10701652 TI - Langerhans' cell histiocytosis showing low-attenuation mediastinal mass and cystic lung disease. PMID- 10701653 TI - CT features of basidiobolomycosis with gastrointestinal and urinary involvement. PMID- 10701654 TI - Review of current literature. PMID- 10701655 TI - Sound localization with eccentric head position. AB - This study investigates the influence of head-to-trunk position on auditory localization in humans. Various methods of head pointing, of two-alternative forced choice, and hand pointing were employed. Head-pointing toward actual sound sources in darkness, by using only the subjective median plane of the head as a reference, resulted in systematic underestimations of target eccentricity. The deviations of the terminal head position from the target shifted with a mean slope of approximately 0.1 degrees per degree change in head position. A corresponding shift in the localization of virtual sound sources (presented via headphones during eccentric head positions) was demonstrated by requiring forced choice (left or right) responses with respect to the subjective median plane of the head. Head pointing toward remembered auditory targets in darkness resulted in undershoots similar to those found with actual targets. However, when a visual marker of the actual median plane of the head was additionally presented to the subject during these tasks (by a laser attached to the head that projected a spot onto a screen), sound localization was fairly accurate. Localization of eccentric auditory targets by using a swivel hand pointer also showed systematic errors similar to those found with head pointing in darkness when the head was simultaneously oriented toward the sound. When the head remained in alignment with the trunk, hand pointing resulted in overshooting responses. These results may be related to neural processes, presumably in the posterior parietal cortex, that transform auditory and visual spatial coordinates into a common, trunk centered, frame of reference. PMID- 10701656 TI - A tryptophan-free diet markedly reduces frontocortical 5-HT release, but fails to modify ethanol preference in alcohol-preferring (sP) and non-preferring (sNP) rats. AB - It has been hypothesised that rat lines genetically selected for their alcohol preference consume large amounts of ethanol because they have a low 5-HT content. Since brain tryptophan (TRP) availability controls the rate at which neurons synthesise and release serotonin (5-HT), we assessed whether the administration of a TRP-supplemented or TRP-free diet for 3 consecutive days influenced alcohol intake in alcohol-preferring and non-preferring sP and sNP rats, respectively. In the same animals extracellular 5-HT concentration was monitored by microdialysis in the frontal cortex. A TRP-free diet progressively and markedly decreased cortical extracellular 5-HT in sP and sNP rats during the treatment period with respect to a balanced diet. However, the TRP-free diet failed to modify alcohol consumption and preference in sP and sNP rats. The TRP-supplemented diet also failed to alter the intake of alcohol in either group of rats. Therefore, these results do not support a specific role of 5-HT transmission in ethanol intake and preference in sP and sNP rats. PMID- 10701657 TI - Anesthesia induced retrograde amnesia is ameliorated by ephrinA5-IgG in mice: EphA receptor tyrosine kinases are involved in mammalian memory. AB - EphA receptors and their ephrin-A ligands were previously thought to play a role only in embryonic development of the brain. Recently, however, these proteins were shown to be expressed in the adult mouse brain, primarily in the hippocampus, and were implicated in hippocampal synaptic plasticity and learning. What aspects of learning EphA receptors mediate have not been studied? Using the fear conditioning paradigm we demonstrate that EphA receptors play roles in memory. We show that post-training surgical anesthesia leads to robust context specific retrograde amnesia in mice, and post-anesthesia activation of EphA receptors induces a significant amelioration of this amnesia. As acquisition was left unaffected and performance factors were found unaltered, we suggest that the amelioration was due to changes in cognition leading to improved memory. Our data represent the first pieces of evidence for the involvement of EphA receptor tyrosine kinase receptors in mammalian memory, a finding that opens a new avenue into the functional analysis of the largest receptor tyrosine kinase subfamily in the brain. PMID- 10701658 TI - Impairment of spatial memory and changes in astroglial responsiveness following loss of molar teeth in aged SAMP8 mice. AB - In order to evaluate the mechanism(s) responsible for senile impairment of cognitive function as a result of reduced mastication, the effects of the loss of the molar teeth (molarless condition) on the hippocampal expression of glial fibrous acidic protein (GFAP) and on spatial memory in young adult and aged SAMP8 mice were studied using immunohistochemical and behavioral techniques. Aged molarless mice showed a significantly reduced learning ability in a water maze test compared with age-matched control mice, while there was no difference between control and molarless young adult mice. Immunohistochemical analysis showed that the molarless condition enhanced the age-dependent increase in the density and hypertrophy of GFAP-labeled astrocytes in the CA1 region of the hippocampus. These effects increased the longer the molarless condition persisted. When the extracellular K+ concentration ([K+]o) was increased from 4 to 40 mM for hippocampal slices in vitro, the mean increase in the membrane potential was about 57 mV for fine, delicate astrocytes, the most frequently observed type of GFAP-positive cell in the young adult mice, and about 44 mV for the hypertrophic astrocytes of aged mice. However, there was no significant difference in resting membrane potential between these cell types. The data suggest that an impairment of spatial memory and changes in astroglial responsiveness occur following the loss of molar teeth in aged SAMP8 mice. PMID- 10701659 TI - Further experiments on the relationship between hippocampus and orientation following phase-shift in homing pigeons. AB - Following a clock- or phase-shift of the light dark cycle, hippocampal lesioned pigeons (Columba livia) consistently display a larger deviation in vanishing bearings away from the homeward direction compared to intact birds; an effect never seen in unshifted birds. In Experiment 1, control and hippocampal lesioned pigeons oriented similarly after being held 1 week under artificial lighting in the absence of a phase-shift. Housing under artificial light by itself does not result in between group orientation differences. In Experiment 2, control and hippocampal lesioned pigeons oriented equally well under overcast conditions, indicating that both groups had a functional magnetic compass. The between group difference in orientation following phase-shift does not appear to be a consequence of control birds being able to use both the sun and earth's magnetic field for orientation and the hippocampal lesioned pigeons only being able to use the sun. In Experiment 3, lengthening the time held under 6-h clock-shift from 1 to 2 weeks had no effect on the magnitude of the difference in orientation, but fast shifting produced clearer effects than slow shifting. Taken together, the data suggest that hippocampal lesions alter how a pigeon responds to a rapidly changing light-dark cycle, particularly following a fast-shift manipulation, suggesting an as yet unspecified relationship between the avian hippocampus and the circadian rhythm(s) that regulate sun compass orientation. PMID- 10701660 TI - Lorazepam-induced modifications of saccadic and smooth-pursuit eye movements in humans: attentional and motor factors. AB - In a placebo-controlled, double-blind study, we measured the effects of low dose lorazepam on attentional and motor factors involved in saccadic and smooth pursuit eye movements. We manipulated the temporal interval between the extinction of the central fixation target and the appearance of a second eccentric target (gap/overlap step paradigm). The second target was either stationary (saccade trial) or moving in a direction opposite to the step (pursuit trial). Gap/overlap effects on the latency of saccadic and smooth pursuit eye movements were measured before and after oral intake of either lorazepam or placebo. Pharmacological effects on the dynamics and the accuracy of both types of eye movements were also investigated. In 14 healthy volunteers, we found that the temporal interval between fixation target offset and eccentric target onset modulates the latency of saccadic and smooth pursuit eye movements in a similar way. As compared to placebo, lorazepam significantly increased the latency of both types of eye movements, but did not modify the gap/overlap effect. Moreover, lorazepam significantly decreased the peak velocity of the first saccade towards the eccentric stationary target, as well as the gain of tracking towards the eccentric moving target. However, the overall accuracy of both behaviors was not significantly affected, indicating that systematic errors in foveating or tracking were detected and corrected by appropriate corrective or catch-up saccades, respectively. Results are discussed in terms of shared/different mechanisms for saccadic and pursuit systems in primates. PMID- 10701661 TI - The superior olivary complex is necessary for the full expression of the acoustic but not tactile startle response in rats. AB - The acoustic startle response (ASR) in rats is mediated by an oligosynaptic pathway from the cochlea via the brainstem to spinal and cranial motoneurons. The present study tested whether the superior olivary complex (SOC) plays a role in the mediation of the ASR. The SOC receives auditory information from the ventral cochlear nuclei and projects to the caudal pontine reticular nucleus (PnC), the sensorimotor interface of the ASR. Axon-sparing excitotoxic lesions of the SOC strongly reduced the ASR amplitude and slightly prolonged ASR onset and peak latencies. The integrity of PnC which is adjacent to the SOC was confirmed by testing the tactile startle response which was not affected by SOC lesions. We suggest that the SOC is necessary for a full expression of the ASR and discuss possible auditory input structures involved in the mediation of the ASR. PMID- 10701662 TI - Functional ablation of deep cerebellar nuclei temporarily impairs learned coordination of forepaw and tongue movements. AB - The role of the cerebellum in complex skilled movements was assessed by the use of functional ablation technique. Rats were trained to synchronize tongue and forepaw movements in a drinking box equipped with a retractable spout which was automatically withdrawn after every lick but could be returned by pressing and releasing a lever placed 4 cm below the spout. The animals learned to perform short presses synchronized with the lick cycle in such a way as to allow continuous drinking. The contribution of the neocerebellum to these lick associated instrumental movements was estimated by intracranial injection of 2 ng of tetrodotoxin into the dentate and lateral part of interposed nuclei. Bilateral blockade of the mainly neocerebellar output interfered with learned synchronization of licking and bar pressing, but did not suppress licking from a stationary spout and only decreased the licking frequency by 10%. It is concluded that the tongue-forepaw synchronization is disrupted by elimination of the neocerebellar output but for a much shorter time (< 9 h) than the tetrodotoxin induced inactivation of the lateral part of the caudate nucleus (72 h) reported earlier. The results confirm participation of cerebellar hemispheres in learned tongue-forepaw synchronization, but indicate at the same time that elimination of this link can be easily compensated. PMID- 10701663 TI - Previous maze experience required to increase open arms avoidance in rats submitted to the elevated plus-maze model of anxiety. AB - Studies have shown an increased open arm avoidance in rats re-exposed to the elevated plus-maze (EPM), which suggests a qualitative shift in emotional states from an unconditioned (Trial 1) to a learned (Trial 2) form of fear response, but a precise source of aversion has not been determined. Using rats submitted to the EPM or various EPM-derived configurations, this study was designed to investigate what previous maze experiences in Trial 1 are required to increase avoidance of open arms in EPM Trial 2. Results obtained from rats submitted to the EPM or EPM derived configurations confirmed the increased open arms avoidance in Trial 2. Rats confined to either open or enclosed arms failed to show the increased avoidance of open arms in Trial 2. The results are discussed in terms of the minimum prerequisite in Trial 1 to elicit an avoidance learning response to open arms in Trial 2, and also the implications of an acquired fear response in rats for the study of the biological basis of anxiety. PMID- 10701664 TI - Measuring fear and anxiety in the marmoset (Callithrix penicillata) with a novel predator confrontation model: effects of diazepam. AB - This report describes a new experimental method for measuring fear and anxiety in Cerrado marmosets (Callithrix penicillata). In order to test the sensitivity of the behavioral parameters to an anxiolytic substance, the effects of the benzodiazepine diazepam on the anxiety measures were examined. The strategy was to use a naturally occurring stimulus known to elicit anxiety and fear in this species. A taxidermized predator (the wild cat Felis tigrina) was chosen as the stimulus to induce anxiety-related behaviors on the basis of a preceding study in which various stimuli were systematically compared in their effectiviness to induce fear responses. The apparatus consisted of three parallel arms of equal dimensions, joining two perpendicular arms at each end, thus comprising a figure eight-like or five-arm continuous rectangular maze. The wild-cat was placed outside of one corner of the maze's outer parallel arms. Each subject was submitted to six treatments given in random order: three drug sessions (diazepam 1, 2 and 3 mg/kg, i.m.), saline, sham (injection control), and a control session, involving neither manipulation nor injection. Subjects were placed into the back of the chamber, out of sight of the 'predator', 20 min after a treatment and given free access to the maze for 30 min. The behavioral repertoire was recorded via videocamera. The following behaviors were considered to be possible indices of emotionality relevant to exposure to the predator in the paradigm used: scratching, scent marking, exploration, frequency and time spent in each of 13 defined sections of the maze. Administration of diazepam induced a significant reduction in scratching and an increase in the time spent in the vicinity of the 'predator', as well as in the frequency of exploratory behaviors, indicative of an anxiolytic effect. Gender did not influence the effect of treatment. These results suggest that this new ethologically-based test may be a useful method for studying anxiety and fear-induced avoidance in non-human primates and for pre clinical research on psychoactive drugs. PMID- 10701665 TI - Mother rats bar-press for pups: effects of lesions of the mpoa and limbic sites on maternal behavior and operant responding for pup-reinforcement. AB - This series of studies explored the operant response rates for pup-reinforcement of female Sprague Dawley rats that were either postpartum or cycling and sustained lesions of the medial preoptic area (mpoa), the lateral amygdala, the nucleus accumbens, or sham lesions. The last experiment tested the effects on operant responding of preventing direct access to pups in mpoa and sham-lesioned postpartum mothers. All animals were trained prior to mating on an FR-1 bar-press schedule to criterion (50 presses in 30 min) for a food (Froot Loops) reward in an operant chamber. At the end of pregnancy animals that were to be tested postpartum were provided in their home cages with six newborn foster pups; mother litter interactions were observed on the last 3 days of pregnancy and throughout the postpartum period. On each of these same days after a period of separation from pups, females were tested in the operant box for delivery of rat pups. With each bar-press response, a rat pup rather than a Fruit Loop was delivered down a gentle shoot into the hopper. Non-postpartum, but maternal, multiparous animals who were showing estrous cycles were tested using the same procedures. The first and second studies showed that animals (both postpartum and as cycling multiparous animals) with mpoa lesions exhibited a significant reduction in bar press rate for pup reinforcement in the operant box. In postpartum animals, amygdala lesions also produced a bar-press deficit, whereas nucleus accumbens lesions did not. All lesioned groups showed deficits in maternal responding in the home cage and deficits in retrieval in the operant box. These results indicate that systems associated with the mpoa mediate both the stereotypical maternal behaviors and pup-reinforcement. In contrast, the expression of home cage maternal behavior is dependent on the integrity of both the amygdala and nucleus accumbens, whereas operant responding need not be. These results indicate a dissociation of mechanisms mediating expression of the species-typical maternal behavior and pup-reinforcement. PMID- 10701666 TI - Ion exchange on resins with temperature-responsive selectivity III. Influence of complex formation stoichiometry on temperature dependence of resin selectivity. AB - The influence of temperature (293308 K 1:1 IDA:Ni2+ complex is dominating. The Cu2+-Ni2+ exchange equilibrium from sulfate medium is characterized by the formation of nickel complexes of both stoichiometries within the whole temperature range studied. The dependence of alpha on T in Zn2+-Co2+ exchange system has been shown to be weaker than that in the Cu2+-Ni2+ system. This result is in a good agreement with the predictions made in the first communication of this series. The results of thermostripping experiments carried out for Cu2+-Co2+ exchange have shown that the efficiency of the thermostripping process depends on both the interval of working temperatures (deltaT) and its position on the temperature scale. The efficiency of thermostripping rises with an absolute deltaT value and also increases following the shift of temperature interval to the lower temperature range. PMID- 10701667 TI - Studies of retention and stability of a horizontally polymerized bonded phase for reversed-phase liquid chromatography. AB - We have studied the novel horizontally polymerized mixed trichloropropyl trichlorooctadecyl silane bonded phase described by Wirth. These materials can be reproducibly prepared and give very high bonded phase density (>7.5 micromol m( 2)). They show significantly improved alkaline stability and chromatographic selectivity towards PAHs similar to conventional monomeric phases. Study of retention by the Linear Solvation Energy Relationship approach as well as measurement of dead volume and retention of methanol indicate that less mobile phase is sorbed by a horizontally polymerized phase than by conventional phases. Silanophilic interaction of amines are decidedly weaker on a silica modified by horizontal polymerization compared to a conventionally modified phase. In addition, this work provides additional support for the "partition-like" retention mechanism of bonded phase RPLC. PMID- 10701668 TI - Optimization of simulated moving bed plants with low efficient stationary phases: separation of fructose and glucose. AB - An optimization procedure for simulated moving bed (SMB) plants with low efficient stationary phases is presented. The new aspect is that the desorbent consumption can be cut by 70% by running the plant with lower internal liquid flows and a corresponding larger switch time while the productivity is kept constant. This concept was validated by the separation of fructose and glucose in water on a calcium resin with an eight-column SMB plant. The separation can be predicted well by a true moving bed (TMB) and a simulated moving bed simulation. Adsorption isotherms were determined up to 300 kg/m3 for glucose and 500 kg/m3 for fructose from 25 to 80 degrees C. Experimental SMB runs were performed over a wide range of feed concentrations (10-350 kg/m3) and temperatures (25-80 degrees C). The strong influence of the delay volume is pointed out. For an experimental run with high feed concentration a complete set of data is presented. To reduce biological growth separation at 80 degrees C is recommended. PMID- 10701669 TI - Chiral and electrokinetic separation of amino acids using polypyrrole-coated adsorbents. AB - An optically active and electroconductive polymeric adsorbent has been developed for the use in chromatographic resolution of nonderivatized amino acids. The chiral selectivity of the adsorbent is based upon ligand exchange of coordinated copper(II) complexes of D or L-amino acids and a molecular imprinting technique by modifying the resin surface with polypyrrole coating. Applying a potential difference of +/-1.5 V to the chiral and conductive column, racemic amino acids are separated according to their charge characteristics, and simultaneously resolved with respect to their optical isomerisms. A pH-controlled mixture of D,L lysine and D,L-aspartic acid is resolved displaying enantioselectivity values of 1.19 and 2.08, respectively, and a baseline separation of the two amino acids is accomplished by alternating the polarity of the electric field. The synthesized adsorbent also exhibits size exclusion factor discriminating amino acids with larger side chains. PMID- 10701670 TI - Enantiomeric and diastereomeric high-performance liquid chromatographic separation of cyclic beta-substituted alpha-amino acids on a teicoplanin chiral stationary phase. AB - High-performance liquid chromatographic (HPLC) separation of stereomeric cyclic beta-substituted or-quaternary alpha-amino acids was performed on a chiral stationary phase based on the glycopeptide antibiotic teicoplanin. The investigated amino acids are the 1-amino-2-methylcyclohexanecarboxylic acids, the 1-amino-2-hydroxycyclohexanecarboxylic acids, Ala, Cha, Phe and Tle. The effects of the mobile phase composition (type and content of organic modifier, pH) and of the temperature on the enantio- and diastereoselectivity were studied and the conditions were optimised to resolve the four stereomers of one amino acid in a single chromatographic run. The influence of the modifier concentration and the pH of the mobile phase reveal two enantiomeric and diastereomeric discrimination mechanisms based on different interactions with the stationary phase. For optimal separation of diastereomers the column has to be conditioned with an acidic eluent. PMID- 10701671 TI - Determination of ammonium in seawater by column-switching ion chromatography. AB - A system which combines column switching and concentration was developed. A binary eluent mechanism was developed to study the effect of high sample concentration matrix on retention time shifts of the trace analyte. Separation conditions were chosen according to this mechanism to reduce the retention time shift of ammonium in the presence of high concentration sodium ion: high concentration sulfuric acid (25 mmol/l) was used as the eluent, and a hydronium selective column of high capacity--a CS12 column, was employed. Since the retention time shift was reduced, the interval between onset and the complete elution of a high concentration ammonium standard (10 mg/l) was directly defined as the column-switching time window, which greatly simplified the procedure for determining the time window. Results showed that for ammonium below 1 mg/l, 90% ammonium was introduced and concentrated. Detection limits of 12.8 microg/l were obtained for ammonium with sodium at 1000 mg/l. PMID- 10701672 TI - Different elution modes and field programming in gravitational field-flow fractionation. 2. Experimental verification of the range of conditions for flow rate and carrier liquid density programming. AB - Gravitational field-flow fractionation utilises the Earth's gravitational field as an external force that causes the settlement of particles towards the channel accumulation wall. Hydrodynamic lift forces oppose this action by elevating of particles from the channel accumulation wall. Therefore there are several possibilities to modulate the resulting force field acting on particles in gravitational field-flow fractionation. Regarding the force field programming in gravitational field-flow fractionation, this work focused on two topics: changes of the difference between particle density and carrier liquid density in Brownian and focusing elution modes and influencing of lift forces achieved by changing the flow-rate in focusing elution mode. We have found and described the experimental conditions applicable to force field programming in the case of separations of silica gel particles by gravitational field-flow fractionation. It was shown that the effect of carrier liquid viscosity in the water-methanol system is implemented as an additional factor enhancing the desired effect of carrier liquid density. Some other forces influencing the retention behaviour of the model particles are discussed. PMID- 10701673 TI - Comparison of gas chromatography-pulsed flame photometric detection-mass spectrometry, automated mass spectral deconvolution and identification system and gas chromatography-tandem mass spectrometry as tools for trace level detection and identification. AB - The complexity of a matrix is in many cases the major limiting factor in the detection and identification of trace level analytes. In this work, the ability to detect and identify trace level of pesticides in complex matrices was studied and compared in three, relatively new methods: (a) GC-PFPD-MS where simultaneous PFPD (pulsed flame photometric detection) and MS analysis is performed. The PFPD indicates the exact chromatographic time of suspected peaks for their MS identification and provides elemental information; (b) automatic GC-MS data analysis using the AMDIS ("Automated Mass Spectral Deconvolution and Identification System") software by the National Institute of Standards and Technology; (c) GC-MS-MS analysis. A pesticide mixture (MX-5), containing diazinon, methyl parathion, ethyl parathion, methyl trithion and ethion was spiked, in descending levels from 1 ppm to 10 ppb, into soil and sage (spice) extracts and the detection level and identification quality were evaluated in each experiment. PFPD-MS and AMDIS exhibited similar performance, both superior to standard GC-MS, revealing and identifying compounds that did not exhibit an observable GC peak (either buried under the chromatographic background baseline or co-eluting with other interfering GC peaks). GC-MS-MS featured improved detection limits (lower by a factor of 6-8) compared to AMDIS and PFPD-MS. The GC PFPD-MS-MS combination was found useful in several cases, where no reconstructed ion chromatogram MS-MS peaks existed, but an MS-MS spectrum could still be extracted at the elution time indicated by PFPD. The level of identification and confirmation with MS-MS was inferior to that of the other two techniques. In comparison with the soil matrix, detection limits obtained with the loaded sage matrix were poorer by similar factors for all the techniques studied (factors of 5.8, >6.5 and 4.0 for AMDIS, PFPD-MS and MS-MS, respectively). Based on the above results, the paper discusses the trade-offs between detectivity and identification level with the compared three techniques as well as other more traditional techniques and approaches. PMID- 10701674 TI - Automated capillary gas chromatographic system to monitor ethylene emitted from biological materials. AB - An automated capillary gas chromatographic system to measure ethylene emitted from biological materials is presented. The system consists of an on-line sampling device, a thermodesorption preconcentration apparatus and a capillary gas chromatograph with a flame ionization detection system. The limit of detection achievable on the GC system alone is 5 pg ethylene. The use of the strong Carboxen 1000 adsorbent at a sampling temperature as low as -50 degrees C allows sampling of volumes up to a few liters. Ethylene concentrations at low ppt levels can be accurately and reproducibly determined. PMID- 10701675 TI - Purity testing of organometallic catalysts by packed capillary supercritical fluid chromatography. AB - A packed capillary column supercritical fluid chromatography system with flame ionization detection has been used for purity testing of candidates for homogeneous catalysis such as methyl tricarbonyl pentamethylcyclopentadienyl tungsten [Cp*W(CO)3Me], methyl tricarbonyl cyclopentadienyl tungsten [CpW(CO)3Me], tetramethyl pentamethylcyclopentadienyl iridium (Cp*IrMe4), trimethyl (1,4,7-trimethyl-1,4,7-triazocyclononane) rhodium (CnRhMe3), trimethylphosphine hydride dicarbonyl cyclopentadienyl molybdenum [eta5 CpMoH(CO)2PMe3] and triphenylphosphine hydride dicarbonyl cyclopentadienyl molybdenum [eta5-CpMoH(CO)2PPh3]. A mass limit of detection of 240 pg was found for eta5-CpMoH(CO)2PMe3 when using a 60-nl injection volume and pure CO2 as mobile phase on a 5 microm Kromasil C18 column. The stability of the catalysts in solution has been examined. After 24 h more than 70% of eta5-CpMoH(CO)2PMe3 and 50% of eta5-CpMoH(CO)2PPh3 had decomposed. Due to the instability of the compounds the purity testing had to take place rapidly after sample dissolution. PMID- 10701676 TI - High-performance thin-layer chromatographic determination of six major ginsenosides in Panax ginseng. AB - A densitometric determination of six major ginsenosides in Panax ginseng, separated by high-performance thin-layer chromatography (HPTLC), was optimized. Simple extraction and clean-up methods of the target constituents and the development of standardized conditions of chromatoplates with a quaternary solvents system allowed an efficient saponins recovery from the plant material and their selective separation. After exposure of the chromatograms to thionyl chloride vapors and further heating, stable reaction products of ginsenosides, which showed absorption maxima at lambda=275 nm as well as a fluorescence (lambda(excitation)=366 nm, lambda(emission)=400 nm), allowed the application of a sensitive and reproducible method for their simultaneous determination. The method was validated by spiking the ginseng extracts with pure standards. PMID- 10701677 TI - Calculation of electrophoretic mobilities in water-organic modifier mixtures in capillary electrophoresis. AB - In order to correlate/predict electrophoretic mobility data in the mixture of water+organic modifier four equations have been presented and examined. The experimental mobilities of five analytes were determined in a water-methanol mixture. These data have been used to assess the accuracy and predictability of the models. Also, some previously published mobility data in water-organic modifier mixtures has been employed for further evaluation of the models. The models produced accurate results and the means of percentage deviations were in the range of 0.66-1.30. PMID- 10701679 TI - Simultaneous stereoselective analysis of tramadol and its main phase I metabolites by on-line capillary zone electrophoresis-electrospray ionization mass spectrometry. AB - On-line combination of partial filling capillary electrophoresis and electrospray ionization mass spectrometry was demonstrated for the simultaneous enantioseparation of tramadol and its main phase I metabolites. The partial filling technique was efficient at avoiding MS contamination by the chiral selector. Different experimental factors were investigated, including the chiral selector nature and concentration, plug length as well as the separation temperature. The best enantioseparation of the investigated compounds was achieved with a coated polyvinyl alcohol capillary and a 40 mM ammonium acetate buffer, pH 4.0, adding sulfobutyl ether beta-cyclodextrin (2.5 mg/ml) as the chiral selector. The charged cyclodextrin not only allowed enantioseparation of tramadol and its metabolites, but also improved the selectivity of compounds with the same molecular mass. Finally, CE-electrospray ionisation-MS was successfully applied to the stereoselective analysis of tramadol and its main metabolites in plasma after a simple liquid-liquid extraction. PMID- 10701678 TI - Enantioseparation of atropine by capillary electrophoresis using sulfated beta cyclodextrin: application to a plant extract. AB - A capillary zone electrophoresis (CZE) method, with sulfated beta-CD as chiral selector, was optimized by means of an experimental design for the enantioseparation of atropine. In this study, a central composite design was used and the following factors were varied simultaneously: buffer concentration, buffer pH and sulfated beta-CD concentration. The resolutions between littorine and its positional isomer ((-)-hyoscyamine) and between atropine enantiomers, as well as the separation time and generated current were established as responses. A model was obtained for each response by linear multiple regression of a second degree mathematical expression. The most favorable conditions were determined by maximizing the resolution between atropine enantiomers and by setting the other responses at threshold values. Successful results were obtained with a 55 mM phosphate buffer at pH 7 in the presence of 2.9 mM sulfated-beta-CD at 20 degrees C and 20 kV. Under these optimized conditions, a baseline separation of littorine and atropine enantiomers was achieved in less than 5 min. Finally, the method allowed the enantiomeric separation of atropine in a pharmaceutical formulation and was also found to be suitable for the enantiomeric purity evaluation of (-) hyoscyamine in plant extracts, in relation with the extraction procedure. It was demonstrated that supercritical fluid extraction induced less racemization than classical liquid-solid extraction procedures. PMID- 10701680 TI - Towards a quantitative definition of perfection in chromatographic analyses. AB - The paper published by Ghaoui and Rothman [J. High Resolut. Chromatogr. 15 (1992) 36] and particularly its Fig. 5, is further considered here because it contains the germ of an idea of how to measure improvements to a chromatographic method, and how to define the goal of perfection in terms of "zero defects" as required by quality assurance schemes. From this, a new role emerges for signal averaging in capillary chromatography: a role to quantify and measure method improvements, and one which can be generally applied to measure improvements in instrument design too. PMID- 10701681 TI - Determination of critical micelle concentration and interactions between cephalosporins and charged surfactants. PMID- 10701682 TI - Expression and localization of the annexins II, V, and VI in myocardium from patients with end-stage heart failure. AB - Annexins II, V, and VI belong to a family of Ca(2+)-dependent phospholipid binding proteins that have been involved mainly in signal transduction, differentiation, membrane trafficking events, or binding to the extracellular matrix, or that might be effective as Ca(2+)-channels. They are abundant in the mammalian myocardium and might play a role in ventricular remodeling and altered calcium handling during heart failure. To test this hypothesis, we compared the expression and distribution of these annexins in nonfailing (n = 9) and failing human hearts with idiopathic dilated cardiomyopathy (n = 11). Northern blot and slot blot analysis were used to determine the annexin mRNA levels and Western blots were used to quantify the amounts of annexin proteins. Distribution of annexins was studied by immunohistofluorescence labeling and compared with that of a sarcolemmal marker (Na+/K(+)-ATPase) and of a myofibrillar protein (alpha actinin). We showed that nonfailing hearts contained a higher amount of annexin VI than of annexin V or II (13.5 +/- 1.8, 3.7 +/- 0.2, and 2.5 +/- 0.5 microg/mg protein, respectively). In failing hearts, there was a parallel increase in both mRNA and protein levels of annexin II (146% and 132%, p < 0.05, respectively) and annexin V (152%, p < 0.01, 147%, p < 0.005, respectively); the protein level of annexin VI was also increased (117%, p < 0.05), whereas the increase of its mRNA level was statistically insignificant. We observed a predominant localization of annexin II in interstitium, and of annexins V and VI in cardiomyocytes at the level of the sarcolemma, T-tubules, and intercalated disks in nonfailing hearts, whereas in failing hearts enlarged interstitium contained all three annexins. Furthermore, annexin V staining at the level of cardiomyocytes almost disappeared. In conclusion, we showed that heart failure is accompanied by marked overexpression of annexins II and V, as well as translocation of annexin V from cardiomyocytes to interstitial tissue. The data suggest that annexins may contribute to ventricular remodeling and annexin V to impaired Ca2+ handling in failing heart. PMID- 10701683 TI - FK506 augments glucocorticoid-mediated cyclooxygenase-2 down-regulation in human rheumatoid synovial fibroblasts. AB - Prostaglandins (PG) formed by cyclooxygenase (COX) enzymes are important mediators of inflammation in rheumatoid arthritis. The contribution of the inducible COX-2 to inflammation in the rheumatoid synovium is well documented. We examined the regulation of COX-2 mRNA and protein expression in response to both glucocorticoids (GC) and FK506 using rheumatoid synovial fibroblasts. Combined treatment of FK506 and a low concentration of dexamethasone (DEX) (10(-9) M) down regulated synovial COX-2 mRNA and protein expression. In contrast, neither FK506 nor DEX (10(-9) M) alone influenced COX-2 expression. Immunocytochemical studies showed that pretreatment with FK506 enhanced the nuclear translocation of the glucocorticoid receptor (GR) in synovial fibroblasts in the presence of low concentrations of DEX (10(-9) M). Transient transfection experiments showed that treatment of cells with FK506 enhanced the expression of glucocorticoid responsive gene reporter in the presence of DEX (10(-9) M). NF-kappaB is known to mediate the transcriptional activation of the COX-2 gene. Electrophoretic mobility shift assay demonstrated that DNA-binding activity of NF-KB was suppressed more profoundly by FK506 plus DEX (10(-9) M) treatment with those of DEX (10(-9)M) alone in IL-1beta-stimulated synovial cells. Our results indicated that FK506-induced potentiation of GR-mediated repression of synovial COX-2 gene transcription is the result of increased translocation of GR to the nucleus and subsequent repression of NF-kappaB transactivation. Our results also suggest that FK506 may exert anti-inflammatory effects in the rheumatoid synovium by potentiating GR-mediated signal transduction. PMID- 10701684 TI - Repetitive acute pancreatic injury in the mouse induces procollagen alpha1(I) expression colocalized to pancreatic stellate cells. AB - Pancreatic stellate cells may be a major source of extracellular matrix deposition during injury. This study was undertaken to establish whether pancreatic stellate cells are a source of Type I collagen in vivo and whether they continue to be a source of matrix production in the post-injury fibrotic pancreas. To induce pancreatic fibrogenesis, acute pancreatic injury was induced in mice three times weekly with supraphysiologic doses of cerulein. Animals were treated for 6 weeks and allowed to recover for an additional 6 weeks. Stellate cell activation and pancreatic collagen expression were measured by immunohistochemistry, whole tissue RNA analysis, and in situ hybridization. Histology and digital image analysis demonstrated the development of substantial pancreatic fibrosis after 6 weeks of treatment. During recovery, incomplete resolution of the fibrosis was found. Procollagen alpha1(I) mRNA increased more than 15-fold during treatment and continued to be 5-fold elevated during the post injury phase. In situ hybridization studies demonstrated that collagen gene expression was colocalized to activated pancreatic stellate cells. Collagen expression and fibrosis persisted in focal areas during recovery. These findings show that pancreatic stellate cells are the major source of collagen during repetitive injury in vivo. Additionally, focal areas of sustained pancreatic fibrogenesis persist after cessation of cerulein treatment, and these areas may contribute to sustained total organ collagen expression in the absence of ongoing injury. PMID- 10701685 TI - Biodistribution and feasibility of non-viral IGF-I gene transfers in thermally injured skin. AB - Gene therapy using cationic liposomes containing cDNA is a relatively new approach with great potential; however, little is known about the mechanisms of dermal gene transfer, its biodistribution, systemic transfection, and cellular uptake. This study identifies mechanisms, transfection rates, and biodistribution of liposomal gene transfers in the skin of thermally injured rats using cDNA gene constructs coding for insulin-like growth factor-I (IGF-I) and Lac Z. Male Sprague-Dawley rats (350 to 375 g) were given a 60% total body surface area full thickness scald burn that was followed by weekly subcutaneous injections of normal saline (control, n = 10), liposomes plus 0.2 microg Lac Z cDNA construct driven by a cytomegalovirus (CMV) promoter (vehicle, n = 10), or liposomes containing 2.2 microg cDNA coding for IGF-I plus 0.2 microg Lac Z cDNA construct driven by a CMV promoter (IGF-I cDNA, n = 10). Gene transfection was determined by histochemical and luminescent beta-galactosidase assays of blood, skin, liver, spleen, and kidney. Transcription of IGF-I cDNA to IGF-I mRNA was determined in skin cells by Northern blot analyses. Levels of IGF-I protein in blood, skin, liver, spleen, and kidney were measured by radioimmunoassay. The biological activity of the translated IGF-I was evaluated by the mitogenic activity in dermal cells and the rate of re-epithelization. Gene transfection was observed only in skin cells. The expression of IGF-I mRNA increased in skin cells of burned rats receiving liposomes containing the IGF-I cDNA construct compared with liposomes without the construct or normal saline. IGF-I protein levels in the skin of rats receiving the IGF-I cDNA was 176 +/- 4 ng/ml compared with 105 +/- 6 ng/ml for liposomes alone or 90 +/-3 ng/ml for saline (p < 0.05). The translated IGF-I protein was found biologically active in the skin by increasing skin cell proliferation and accelerating re-epithelization 33 days after thermal injury (p < 0.05). No systemic transfection could be detected. Skin cells transfected with liposomes encapsulating the IGF-I cDNA constructs increased the expression of IGF I mRNA transcript and the expression of a biologically active IGF-I protein. Liposomes containing the cDNA coding for IGF-I present an effective approach to gene therapy in the skin. PMID- 10701686 TI - Accelerated appearance of multiple B cell lymphoma types in NFS/N mice congenic for ecotropic murine leukemia viruses. AB - Spontaneous lymphomas occur at high frequency in NFS x V+ mice, strains congenic for ecotropic murine leukemia virus (MuLV) proviral genes and expressing virus at high titer. In the present study, a total of 703 NFS x V+ lymphomas were studied by histopathology, immunophenotypic analysis, immunoglobulin heavy chain or T cell receptor beta chain rearrangements, and somatic ecotropic MuLV integrations; 90% of the lymphomas tested were of B cell lineage. Low-grade tumors included small lymphocytic, follicular, and splenic marginal zone lymphomas, while high grade tumors comprised diffuse large-cell (centroblastic and immunoblastic types), splenic marginal zone, and lymphoblastic lymphomas. Comparison of mice of similar genetic background except for presence (NFS x V+) or absence (NFS x V-) of functional ecotropic MuLV genomes showed that NFS x V-clonal lymphomas developed at about one-half the rate of those occurring in NFS x V+ mice, and most were low-grade B cell lymphomas with extended latent periods. In NFS x V+ mice, clonal outgrowth, defined by Ig gene rearrangements, was associated with acquisition of somatic ecotropic proviral integrations, suggesting that, although generation of B cell clones can be virus independent, ecotropic virus may act to increase the rate of generation of clones and speed their evolution to lymphoma. The mechanism remains undefined, because only rare rearrangements were detected in several cellular loci previously associated with MuLV insertional mutagenesis. PMID- 10701687 TI - Apoptosis induced in vitro and in vivo during infection by Ebola and Marburg viruses. AB - Induction of apoptosis has been documented during infection with a number of different viruses. In this study, we used transmission electron microscopy (TEM) and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling to investigate the effects of Ebola and Marburg viruses on apoptosis of different cell populations during in vitro and in vivo infections. Tissues from 18 filovirus-infected nonhuman primates killed in extremis were evaluated. Apoptotic lymphocytes were seen in all tissues examined. Filoviral replication occurred in cells of the mononuclear phagocyte system and other well-documented cellular targets by TEM and immunohistochemistry, but there was no evidence of replication in lymphocytes. With the exception of intracytoplasmic viral inclusions, filovirus-infected cells were morphologically normal or necrotic, but did not exhibit ultrastructural changes characteristic of apoptosis. In lymph nodes, filoviral antigen was co-localized with apoptotic lymphocytes. Examination of cell populations in lymph nodes showed increased numbers of macrophages and concomitant depletion of CD8+ T cells and plasma cells in filovirus-infected animals. This depletion was particularly striking in animals infected with the Zaire subtype of Ebola virus. In addition, apoptosis was demonstrated in vitro in lymphocytes of filovirus-infected human peripheral blood mononuclear cells by TEM. These findings suggest that lymphopenia and lymphoid depletion associated with filoviral infections result from lymphocyte apoptosis induced by a number of factors that may include release of various chemical mediators from filovirus infected or activated cells, damage to the fibroblastic reticular cell conduit system, and possibly stimulation by a viral protein. PMID- 10701688 TI - Cardiac dysfunction occurs in the HIV-1 transgenic mouse treated with zidovudine. AB - Cardiomyopathy in AIDS is an increasingly important clinical problem. Mechanisms of AIDS cardiomyopathy were explored using AIDS transgenic mice that express replication-incompetent HIV-1 (NL4-3delta gag/pol). Transgenic and FVB/n mice (n = 3 to 6 per cohort) received water ad libitum with and without zidovudine (3' azido-2',3'-deoxythymidine; AZT; 0.7 mg/ml) for 21 or 35 days. After 21 days, echocardiographic studies were performed and abundance of mRNA for cardiac sarcoplasmic reticulum calcium ATPase (SERCA2), sodium calcium exchanger (NCX1), and atrial natriuretic factor were determined individually using Northern analysis of extracts of left ventricles. After 35 days, contractile function and relaxation were analyzed in isolated work-performing hearts. Histopathological and ultrastructural (transmission electron microscopy) changes were identified. After 21 days, molecular indicators of cardiac dysfunction were found. Depressed SERCA2 and increased atrial natriuretic factor mRNA abundance occurred in left ventricles from AZT-treated transgenic mice. NCX1 abundance was unchanged. Eccentric left ventricle hypertrophy was determined echocardiographically. After 35 days, cardiac dysfunction was worst in AZT-treated and AZT-untreated transgenic mice. Decreases in the first derivative of the maximal change in left ventricle systolic pressure with respect to time (+dP/dt) occurred in transgenic mice with and without AZT. Increased half-time of relaxation and ventricular relaxation (-dP/dt) occurred in AZT-treated and -untreated transgenic mice. Increased time to peak pressure was found only in AZT-treated transgenic mice. In AZT-treated FVB/n mice, -dP/dt was decreased. Ultrastructurally, mitochondrial destruction was most pronounced in AZT-treated transgenic mice, but also was found in AZT-treated FVB/n mice. Transgenic mice that express HIV-1 demonstrate cardiac dysfunction. AZT treatment of FVB/n mice causes mitochondrial ultrastructural alterations that are similar to those in other species. In transgenic mice, AZT treatment worsens molecular and ultrastructural features of cardiomyopathy. HIV-1 constructs and AZT each contribute to cardiac dysfunction in this murine model of AIDS cardiomyopathy. PMID- 10701689 TI - Methacarn fixation: a novel tool for analysis of gene expressions in paraffin embedded tissue specimens. AB - To establish a quantitative method for analysis of gene expressions in small areas of tissue after paraffin embedding, preliminary validation experiments with RT-PCR and Western blotting were performed using methacarn-fixed rodent tissues and a cultured PC12 cell line. A total RNA yield of 52 +/- 15 ng/mm2, sufficient for a quantitative RT-PCR of many genes, could be extracted from a deparaffinized 10-microm-thick rat-liver section by a simple, single-step extraction method. The low concentration of contaminating genomic DNA and the resolution of ribosomal RNAs in RNA gel proved the purity and integrity of the extracted RNA samples, allowing PCR amplification of a long mRNA sequence and mRNA species expressing low copy numbers. PCR amplification of mRNA-derived target gene fragments could be achieved by optimizing the amount of total RNA for reverse transcription and the number of subsequent PCR cycles for each gene. By this validation, organ- and sex-specific mRNA expression could be detected in methacarn-fixed paraffin embedded tissues without additional DNase treatment of RNA samples. RT-PCR analysis could also be performed with total RNA extracted from deparaffinized tissue dissected with a laser capture microdissection system. In addition, extraction of protein yielded 4.9 +/- 2.1 microg/mm2 from a 10-microm-thick rat liver section, allowing a quantitative expression analysis of protein by Western blotting. Thus, in addition to its advantages for immunohistochemistry, methacarn fixed paraffin-embedded tissue has benefits for analysis of both RNAs and proteins in the cells of histologically defined areas. PMID- 10701690 TI - Pseudomonas aeruginosa virulence factors delay airway epithelial wound repair by altering the actin cytoskeleton and inducing overactivation of epithelial matrix metalloproteinase-2. AB - To investigate the role of P. aeruginosa virulence factors in the repair of human airway epithelial cells (HAEC) in culture, we evaluated the effect of stationary phase supernatants from the wild-type strain PAO1 on cell migration, actin cytoskeleton distribution, epithelial integrity during and after repair of induced wounds, and the balance between matrix metalloproteinases (MMP) and their tissue inhibitors (TIMP). PAO1 supernatant altered wound repair by slowing the migration velocity in association with altered actin cytoskeleton polymerization in the lamellipodia of migrating airway epithelial cells and delaying or inhibiting the restoration of epithelial integrity after wound closure. PAO1 virulence factors overactivated two of the gelatinolytic enzymes, MMP-2 and MMP 9, produced by HAEC during repair. During HAEC repair in the presence of PAO1 virulence factors, enhanced MMP-2 activation was associated with decreased rates of its specific inhibitor TIMP-2, whereas enhanced MMP-9 activation was independent of changes of its specific inhibitor TIMP-1. These inhibitory effects were specific to P. aeruginosa elastase-producing strains (PAO1 and lipopolysaccharide-deficient AK43 strain); supernatants from P. aeruginosa strain elastase-deficient PDO240 and Escherichia coli strain DH5alpha had no inhibitory effect. To mimic the effects of P. aeruginosa, we further analyzed HAEC wound closure in the presence of increasing concentrations of activated MMP-9 or MMP-2. Whereas increasing concentrations of active MMP-9 accelerated repair, excess activated MMP-2 generated a lower migration velocity. All these data demonstrate that P. aeruginosa virulence factors, especially elastase, may impede airway epithelial wound closure by altering cell motility and causing an imbalance between pro- and activated forms of MMP-2. PMID- 10701691 TI - A comparison of the histopathology of premalignant and malignant mammary gland lesions induced in sexually immature rats with those occurring in the human. AB - The injection of sexually immature female rats with 1-methyl-1-nitrosourea results in a rapid induction of premalignant and malignant mammary gland lesions within 35 days of carcinogen administration. This model affords the opportunity for investigators to study the process of mammary carcinogenesis over a very short latency and to investigate early events in this process. We have recently published on various aspects of this system including the histology of the lesions induced, the time frame of their occurrence, and their dependence on ovarian hormones for their maintenance and growth. In this report we present evidence that many aspects of the histopathology of mammary lesions in this model system are similar to those occurring in humans. We also discuss aspects of the human disease, which are not recapitulated in this model. PMID- 10701692 TI - High frequency of allelic loss in dysplastic lichenoid lesions. AB - Oral lichen planus (OLP) is a common mucosal condition that is considered premalignant by some, whereas others argue that only lichenoid lesions with epithelial dysplasia are at risk of progressing into oral carcinoma. A recent study from this laboratory used microsatellite analysis to evaluate OLP for loss of heterozygosity (LOH) at loci on three chromosomal arms (3p, 9p, and 17p) (Am J Path 1997;Vol151:Page323-Page327). Loss on these arms is a common event in oral epithelial dysplasia and has been associated with risk of progression of oral leukoplakia to cancer. The data showed that, although dysplastic epithelium demonstrated a high frequency of LOH (40% for mild dysplasia), a significantly lower frequency of LOH was noted in OLP (6%), which is even lower than that in hyperplasia (14%). Such results do not support OLP as a lesion at risk for malignant transformation. As a second step of the research, we determined LOH frequencies in 61 dysplastic lichenoid lesions (mild 35; moderate 19; severe 7) using the same microsatellite markers and compared these results with data obtained from the first study and from 13 normal mucosal specimens. Dysplastic lichenoid lesions showed a high frequency of loss (54% for lichenoid lesions with mild dysplasia), but values did not differ significantly from those observed in dysplasia of similar degree without lichenoid appearance. None of the normal mucosa demonstrated LOH. Epithelial dysplasia is a sign of malignant risk, independent of lichenoid changes. Such results suggest that pathologists should search for dysplasia carefully in lesions that otherwise qualify as OLP and that caution should be used when discounting dysplasia as being merely a reactive condition in lichenoid lesions. PMID- 10701693 TI - Big prolactin 60 kDa is overexpressed in salivary glandular epithelial cells from patients with Sjogren's syndrome. AB - Characterization of endogenous synthesis of prolactin (PRL) proteins and their cellular localization in labial salivary glands of patients with Sjogren's syndrome (SS) were achieved. PRL, PRL-receptors (PRL-R), and S100A6 protein were detected by immunohistochemistry. In situ prolactin synthesis was investigated in controls and SS patients by ex vivo incubation of minor salivary glands biopsies and immunoprecipitation assay. Increased PRL-immunoreactivity was found in cytoplasmic acinar epithelial cells in SS patients compared with normal subjects. PRL-R was distributed only in ductal epithelial cells in which S100A6 protein (a PRL-R-associated protein) was also present. PRL, PRL-R, or S100A6 immunoreactivity was not detected in infiltrating mononuclear cells. Immunoprecipitation demonstrated that PRL synthesis occurred in minor salivary glands with increased synthesis of two distinct PRL-like proteins (one major band at 60 kDa and a minor at 16 kDa) in SS glands compared with normal glands. Expression of PRL gene was demonstrated in SS salivary glands using RT-PCR. A positive correlation was found between the presence of PRL-like proteins in acinar epithelial cells of SS patients and clinical extraglandular manifestations. The presence of anti-Ro and anti-La antibodies also positively correlated with a higher percentage of PRL in acinar epithelial cells. In conclusion, PRL-like proteins are synthetized and overexpressed in glandular epithelial cells of labial salivary glands from SS patients and correlate with the aggressiveness of the disease. PMID- 10701694 TI - Age and organ dependent spontaneous generation of nuclear 8-hydroxydeoxyguanosine in male Fischer 344 rats. AB - 8-Hydroxydeoxyguanosine (8-OHdG) is a major oxidative DNA adduct playing roles in senescence, carcinogenesis and various disease processes. High-performance liquid chromatography with an electrochemical detection (HPLC-ECD) method has been widely used to assess organ levels of 8-OHdG, and a recently introduced immunohistochemical approach has made it possible to clarify intra-organ localization. In the present study, these methods were employed to reveal age dependent changes in nuclear 8-OHdG within various tissues of male Fischer 344 rats between 18 fetal days and 104 weeks of age. 8-OHdG was detected in the nuclei of cerebellar small granule and small cortical cells, cerebral nerve cells, and choroid plexus epithelia of the brain and ependymal cells of the spinal cord; parenchymal cells in the anterior lobe of the pituitary and adrenal glands (mainly cortex); bronchial epithelium of the lung; intra-hepatic bile duct, pancreatic duct, glandular gastric and intestinal epithelial cells; renal tubular epithelial cells (mainly medulla); and spermatogonia and spermatocytes of the testis and seminal vesicle epithelia. The nuclear 8-OHdG levels were high (more than two lesions per 10(6) deoxyguanosines) from 7 days to 104 weeks of age in the brain, 3 to 6 weeks in the adrenal gland, 6 to 104 weeks in the lung, and 3 to 52 weeks in the testis. In the other organs, the nuclear 8-OHdG levels remained low throughout. These findings provide a basis for research dealing with oxidative stress by indicating organ-specific and age- but not aging-dependent changes in the localization of spontaneously generated nuclear 8-OHdG in intact rats. The immunohistochemical approach has advantages for assessing variation of 8-OHdG formation at the cellular level not accessible to the HPLC-ECD method. PMID- 10701695 TI - Human scavenger receptor B1 is involved in recognition of apoptotic thymocytes by thymic nurse cells. AB - Recognition and uptake of apoptotic cells by neighboring phagocytes is essential for the clearance of dying cells without accompanying inflammation or tissue damage. In the thymus, many apoptotic cells are generated in the process of negative selection, and both thymic macrophages (professional phagocytes) and nursing thymic epithelial cells (nursing TEC; nonprofessional phagocytes) recognize and ingest them. However the receptors responsible for this recognition and uptake have not been identified. In the present study, we have established a human nursing TEC line and examined the expression of several genes of the scavenger receptor family considered to be potential receptors for apoptotic cells. Human scavenger receptor-B1 (hSR-B1)/CLA-1, previously shown to recognize apoptotic cells, was strongly expressed in nursing TEC, whereas there was little or no expression of the other scavenger receptors tested: scavenger receptor class A, CD36, or CD68. Suppression of hSR-B1/CLA-1 expression using antisense oligonucleotides decreased the binding of apoptotic thymocytes to nursing TEC by more than 40%. These results indicate that hSR-B1/CLA-1 may play a major role in the clearance of apoptotic cells in the thymus, mediating the recognition and ingestion of apoptotic thymocytes by nursing TEC. PMID- 10701696 TI - Detection of MYCN amplification in neuroblastoma using competitive PCR quantitation. PMID- 10701697 TI - Identification of methylated cytosine from archival formalin-fixed paraffin embedded specimens. PMID- 10701698 TI - Pharmacokinetics of 9alpha-fluoromedroxyprogesterone acetate in rats: comparison with medroxyprogesterone acetate. AB - Medroxyprogesterone acetate (MPA) is widely used in endocrine therapy for breast cancer and other diseases. Recently, it has been demonstrated that 9alpha fluoromedroxyprogesterone acetate (FMPA) also has anti-tumour activity in chemical-induced rat mammary tumour and its activity is greater than that of MPA. In the present study, the physico-chemical properties of FMPA and MPA and their pharmacokinetics in female rats were investigated. Partition coefficients (log P) of FMPA and MPA were 3.1 and 3.8, respectively, while the solubilities of FMPA and MPA in phosphate buffer saline were 3.8 and 1.1 microg/mL, respectively. When the two agents were intravenously or orally administered into female rats, there was no significant difference between their plasma concentrations. However, unmetabolized drug excreted into urine accounted for 4.7 and 0.7% of the intravenous dose of FMPA and MPA, respectively. The free fraction of FMPA in rat plasma was approximately four times that of MPA. Assuming the well-stirred model, hepatic intrinsic clearances of FMPA and MPA were estimated to be 64 and 293 L/h per kg, respectively. In addition, the free fraction of FMPA in blood is estimated to be higher than that of MPA, which may explain the higher anti-tumour activity. PMID- 10701699 TI - Single dose pharmacokinetics and bioavailability of nevirapine in healthy volunteers. AB - The results of two randomized, single-dose, crossover bioavailability studies are presented which describe the pharmacokinetics and oral bioavailability of nevirapine, a novel nonnucleoside antiretroviral drug. In the first study 12 healthy male volunteers received nevirapine 15 mg via short-term i.v. infusion or orally as a 50 mg tablet or reference solution (50 mg/200 mL). Following the i.v. dose, nevirapine had a low systemic clearance (Mean +/- S.D., Cl = 1.4 +/- 0.3 L/h) and a prolonged elimination phase (t(1/2beta) = 52.8 +/- 14.8 h; MRT = 81.4 +/- 22.4 h). Nevirapine absolute bioavailability was 93 +/- 9% and 91 +/- 8% for the tablet and oral solution, respectively. In the second study, 24 healthy male volunteers were administered nevirapine as a 200 mg production-line tablet or oral reference solution (200 mg/200 mL). There was no significant difference in bioavailability between the tablet and reference solution. Overall, comparison of the pharmacokinetic parameters between the 50 and 200 mg doses indicates that nevirapine is well absorbed at clinically relevant doses. The absorption profiles using deconvolution revealed no evidence of differential enzyme induction between the two doses or routes of administration following a single dose. PMID- 10701700 TI - Plasma protein binding of celecoxib in mice, rat, rabbit, dog and human. AB - The plasma protein binding of celecoxib was determined for animals and humans using in vitro and ex vivo methods. Eight, healthy, human volunteers (three male, five female, 20-39 years) received celecoxib (600 mg) BID for 7 days, blood samples were collected and concentrations of bound and unbound celecoxib determined. The fraction of bound drug in the volunteers was constant (97.4 +/- 0.1%) at total celecoxib plasma concentrations ranging from 0.01 to 4.02 microg/mL. The ex vivo plasma protein binding of celecoxib in the animals was concentration-independent up to approximately 12, 8 and 10 microg/mL for mouse, rat and dog, respectively. The plasma protein binding of celecoxib after a single oral dose of 10 and 300 mg/kg to mice was 98.3 +/- 0.2%, of 1 and 400 mg/kg to rats was 98.3 +/- 0.2% and of 1 and 100 mg/kg to dogs was 98.5 +/- 0.1%. The percent binding of celecoxib to plasma proteins in vitro was slightly lower than those values determined ex vivo. The in vitro binding of celecoxib to plasma protein was constant over the concentrations of 0.1-10 microg/mL for all species, except rat. PMID- 10701701 TI - Percutaneous absorption of [3H]tretinoin and systemic exposure to mequinol after dermal application of 2% mequinol/0.01% [3H]tretinoin (Solage) solution in healthy volunteers. AB - Solage is a combination product composed of 2% mequinol (4-hydroxyanisole) and 0.01% tretinoin (all-trans-retinoic acid) in an ethanolic solution, which is being studied for its safety and efficacy as a topical treatment for disorders of skin hyperpigmentation. The purpose of this study was to evaluate the extent of percutaneous absorption of [3H]tretinoin and to estimate the systemic exposure to mequinol from this combination product when topically applied to the backs of healthy subjects. Eight subjects received bid topical applications of nonradiolabelled 2% mequinol/0.01% tretinoin solution on a 400 cm2 area of the back for 14 days. The subjects then received a single topical application of 2% mequinol/0.01% [3H]tretinoin solution. After 12 h, the radiolabelled dose was removed and bid treatment with nonradiolabelled 2% mequinol/0.01% tretinoin solution was continued for 7 days. Plasma, urine and faecal samples were analysed for total radioactivity and plasma was analysed for both mequinol and tretinoin by GC/MS procedure. Mean percutaneous absorption of [3H]tretinoin based on the cumulative recoveries of radioactivity in the urine and faeces was about 4.5% (median 2.18%). Tretinoin concentrations in plasma did not increase above endogenous levels. This was consistent with the concentrations of radioactivity in plasma, which showed an average Cmax of 91 pg-eq/mL (median 26 ng/mL). Average Cmax and AUC(0-12 h) values for mequinol were 10 ng/mL and 33 ng h/mL, respectively. Based on the results of this study, systemic toxicity from topical application of tretinoin in this formulation is unlikely, because percutaneous absorption of tretinoin is minimal and because endogenous levels of tretinoin are not increased following bid dosing with this combination formulation. The safety of mequinol in this combination formulation is supported by the low systemic exposures of the subjects in this study compared with the systemic exposures at the highest doses in the dermal toxicity studies in mice (16.6-fold) and rats (34.6-fold). PMID- 10701702 TI - Disposition and exposure of the fibrinogen receptor antagonist XV459 on alphaIIBbeta3 binding sites in the guinea pig. AB - The disposition of XV459, a potent, selective GP IIb/IIIa antagonist, has been examined following intravenous administration of XP280, the benzenesulphonate salt, and 3H-SA202, the trifluroacetic acid salt, to male guinea pigs. A liquid chromatography-mass spectrometry (LC-MS) method was developed and validated for XV459 quantitation in guinea pig plasma with an LLOQ of 0.1 ng/mL. Intravenous infusions (30 min) of XP280 at doses of 0.5 and 2.0 microg/kg were administered to guinea pigs which were sequentially sacrificed at 0.5, 1, 1.5, 4, 8, 12, 24, 48 and 72 h postinitiation of infusion. Maximum total (unbound and GP IIb/IIIa displaced) XV459 plasma concentration of approximately 3.5 microg/mL was obtained at the 2.0 microg/kg dose. Pooling individual concentration-time data yielded a systemic clearance of 1.42 mL/min/kg, Vss of 0.24 L/kg, and a terminal half-life of 2.8 h in the guinea pig at the 0.5 microg/kg dose. The 2.0 microg/kg dose yielded XV459 exposure that was less than proportional to the previous dose. Similar behaviour has been observed in human trials. Cumulative (up to 72 h) urinary and faecal recovery of total radioactivity was 66.4 and 11.2%, respectively. The time course of spleen, marrow and whole blood radioactivity profiles was similar, suggesting that XV459 was not preferentially sequestered on non-plasma GP IIb/IIIa binding sites. Tissue to blood ratios of 20.7 and 8.3 for the spleen and bone marrow, respectively, indicate that increased (relative to blood) exposure was evident for sites containing the GP IIb/IIIa receptor. In vitro studies confirmed the similarity of XV459 binding to both resting and activated platelets in the guinea pig and humans. Given the comparability of dissociation rate constants and IC50s based on in vitro platelet aggregation, human dosimetry estimates should assume similar partitioning of radiolabelled XV459 as in the guinea pig. These results suggest that the guinea pig may indeed be an appropriate animal model for pharmacokinetic and distribution studies with DMP754; in conjunction with recent pharmacological findings with GP IIb/IIIa antagonists, our results suggest that the guinea pig may be the rodent species of choice for preclinical studies with some other GP IIb/IIIa antagonists. PMID- 10701703 TI - The effects of mechanical forces on lung functions. AB - The lung is a dynamic organ that is subjected to mechanical forces throughout development and adult life. This review article addresses the types of mechanical forces in the lung and their effects on development and normal lung functions. The effects of mechanical forces on the various different cell types of the lung are discussed, as are the mechanisms underlying mechanotransduction. PMID- 10701705 TI - Role of nucleus isthmi in the ventilatory response to hypoxia of Bufo paracnemis. AB - Nucleus isthmi (NI) is a mesencephalic structure of the amphibian brain that has recently been reported to participate in CO2-ventilatory response. The present study was designed to test the hypothesis that NI is also involved in hypoxia induced hyperventilation and in the breathing pattern of the toad Bufo paracnemis. Pulmonary ventilation was directly measured by pneumotachography method in control, sham-operated and NI-lesioned toads exposed to normoxia and hypoxia (7 and 5% inspired O2). Under normoxic conditions, NI lesion caused no significant change in the ventilatory pattern or in the pulmonary ventilation. Hypoxia caused a significant (P < 0.05) increase in ventilation in control and sham-operated animals mainly due to an elevated VT. The hypoxia-induced hyperventilation was greater (P < 0.05) in the NI-lesioned toads, due to increases in both fR and VT. Such increased fR under hypoxia was due to a higher number of breaths per burst. The data indicate that NI plays no role under normoxic conditions but is involved in the ventilatory response to hypoxia, exercising an inhibitory modulation on pulmonary ventilation. PMID- 10701704 TI - Medroxyprogesterone acetate with acetazolamide stimulates breathing in cats. AB - Both medroxyprogesterone acetate (MPA) and acetazolamide (ACET) increase ventilation. Combined administration of these agents could result in an additional improvement of blood gases, for example in patients with chronic obstructive pulmonary diseases. The aim of this study in anaesthetized female (ovariohysterectomized, pre-treated with 17-beta-estradiol) cats was to compare the effects on the CO2 response curve of MPA alone (4 microg kg(-1), i.v.) with those after MPA followed by ACET (4 mg kg(-1) i.v.). We performed dynamic end tidal CO2 forcing and analysed the data with a two-compartment model comprising a fast peripheral and slow central compartment, characterized by CO2 sensitivities (Sp and Sc, respectively) and a single offset (the apnoeic threshold B). MPA reduced Sp from 0.22 +/- 0.09 (mean +/- S.D.) to 0.13 +/- 0.06 L min(-1) kPa(-1) (P < 0.01) and Sc from 1.01 +/- 0.38 to 0.88 +/- 0.32 L min(-1) kPa(-1) (P < 0.01). B decreased from 4.02 +/- 0.27 to 3.64 +/- 0.42 kPa (P < 0.01). Subsequent administration of ACET reduced Sp and Sc further to 0.09 +/- 0.06 and to 0.70 +/- 0.49 L min(-1) kPa(-1) (P < 0.01), respectively. The apnoeic threshold decreased further to 2.46 +/- 1.50 kPa (P < 0.01). Because both treatments reduced ventilatory CO2 sensitivity, we conclude that a simulating effect on ventilation is due to a decrease in the apnoeic threshold. Combined administration of MPA and ACET may lead to larger increases in ventilation than treatment with either drugs alone. PMID- 10701706 TI - Temporal evolution of pneumothorax: respiratory mechanical and histopathological study. AB - Respiratory mechanics, chest wall configuration, and lung morphometry were determined in rats before and at 30 (PTX.30) and 60 (PTX.60) min after pneumothorax induction (intrathoracic injection of 8 ml of room air; 50% collapse). Pneumothorax increased respiratory system and lung elastances and viscoelastic/inhomogeneous pressures in both groups, but respiratory system and lung resistive pressures increased only in PTX.60 group. Antero-posterior diameters at the third intercostal space and xiphoid levels, circumference at xiphoid level, and thoracic cephalo-caudal diameter increased significantly after pneumothorax induction independently of temporal evolution. In both groups lung collapse, hyperinflation, and interstitial and alveolar edema were present. Additionally, in PTX.60 group the central airways calibre diminished in relation to PTX.30. In conclusion pneumothorax yields changes in respiratory system and lung elastic and viscoelastic parameters, which are related to alveolar collapse and edema, respectively. Temporal evolution of pneumothorax also leads to changes in lung resistive pressure, probably because of airway narrowing. PMID- 10701707 TI - Airway cross section strongly influences alveolar plateau slope of capnograms for smaller tidal volumes. AB - We compared the predictions of the single path convection-diffusion model (SPM) and the well-mixed single acinus model (SAM) with the normalized slopes (NS) of experimentally measured volumetric capnograms in which VT was varied in three healthy spontaneously breathing adults. For values of VT greater than 15 ml/kg, the tidal volume penetrates deep into the acinar airways, mixing by molecular diffusion is rapid and the predictions of the SAM and SPM both agree with the experiment. The SPM however shows much better agreement with the experimental NS data than does the SAM for values of VT less than 10 ml/kg. The explanation for the departure of the SAM from the observed experimental data, at small VT, is that it represents the limiting case (of well-mixed alveolar gas) for the SPM, only at large VT, where the assumption of rapid mixing is most accurate. We conclude that in general, gas phase diffusivity and total acinar airway cross sectional area variation with cumulative volume into the lung are essential to realistically model airway gas exchange between VT and FRC and to obtain agreement with experimental data under the widest range of breathing conditions. PMID- 10701708 TI - Airflow limitation and control of end-expiratory lung volume during exercise. AB - To test the hypothesis that the presence of airflow limitation (AFL) influences the control of end-expiratory lung volume (EELV) during exercise, 11 subjects with normal lung function, performed submaximal exercise (SM) on a cycle ergometer, with and without AFL. AFL was achieved during exercise by increasing the density of the air via a hyperbaric chamber, compressed to a depth of 3 atm (3 ATA; with AFL). Five subjects achieved AFL during SM exercise at 3 ATA while the remaining six subjects did not achieve AFL. SM exercise was performed with the same apparatus in the hyperbaric chamber at sea level pressure with none of the subjects achieving AFL (SL; no-AFL). EELV (% of TLC, BTPS), was significantly larger during exercise at 3 ATA than during exercise at SL for the AFL group (SL = 44 +/- 6%; 3 ATA-AFL = 51 +/- 9%, P < 0.05; but, was not for the no-AFL group (SL = 46 +/- 6%; 3 ATA-no AFL = 46 +/- 7%). End inspiratory lung volume was significantly elevated during exercise at 3 ATA compared with SL in the AFL group (SL = 80 +/- 6%; 3 ATA-AFL = 86 +/- 6%; P = 0.01) but not in the no-AFL group (SL = 82 +/- 4%; 3 ATA-no AFL = 84 +/- 4%). Tidal volume and ventilation were not different for any condition. These data suggest that the occurrence of AFL influences the control of EELV. PMID- 10701709 TI - CO2 transport and excretion in rainbow trout (Oncorhynchus mykiss) during graded sustained exercise. AB - A quantitative analysis of CO2 transport and excretion was conducted in seawater acclimated rainbow trout (Oncorhynchus mykiss) swimming at different sustained swimming velocities. CO2 excretion increased linearly with cardiac output during exercise but arterial P(CO2) (Pa(CO2)) and total CO2 levels also increased indicating a diffusion limitation to CO2 excretion. The elevated Pa(CO2) was not accompanied by a decrease in pH, indicating that the acid-base compensation was rapid. Mixed-venous P(CO2) increased to a greater extent than Pa(CO2) resulting in a large increase in the venous arterial difference in P(CO2) (Pv(CO2) - Pa(CO2)). The Pv(CO2) - Pa(CO2) difference was used to calculate the proportion of total CO2 excreted comprised of dissolved CO2 which accounted for less than 1% of total CO2 excreted in fish swimming at 11 cm sec(-1) but increased to about 9% at the greatest swimming velocity indicating that the pattern of CO2 excretion changes during exercise. There was no effect of exercise on the proportion of CO2 excreted which was dependent upon HCO3-/Cl- exchange (54%) or that which was dependent upon the dehydration of HCO3- that resided within the red cell prior to gill blood entry (42%). The large proportion of total CO2 excreted that was dependent upon HCO3-/Cl- exchange is significant because this is thought to be the rate limiting step in CO2 excretion. PMID- 10701710 TI - The interaction between O2 and CO2 exchange in rainbow trout during graded sustained exercise. AB - A quantitative analysis of O2 and CO2 transport was conducted in resting and exercising rainbow trout, and these data were used to quantify the magnitude of coupling between O2 and CO2 exchange, in vivo. The release of Bohr protons during haemoglobin-oxygenation was non-linear over the Hb-O2 equilibrium curve used in trout subjected to different levels of sustained exercise. At low swimming speeds, when venous blood O2 content (CvO2) was high, there was a small acidosis as blood passed through the gills, indicating more protons were released during oxygenation of Hb than were consumed during HCO3- dehydration. At higher swimming speeds, when CvO2 was low, there was a significant alkalosis in arterial relative to venous blood, indicating that fewer protons were released upon oxygenation than HCO3- ions were dehydrated to CO2. Haldane coefficients (moles of protons released per mole of O2 which binds to Hb), calculated from steady state arterial and mixed-venous parameters, revealed that under resting conditions all blood CO2 removed from the blood during gill transit was stoichiometrically related to O2 uptake through the release of Bohr protons during Hb oxygenation. The magnitude of coupling between CO2 excretion and O2 uptake decreased from 100% to less than 40% at the maximal swimming velocity when the largest region of the Hb-O2 equilibrium curve was used for gas exchange. The non-linear release of Bohr protons over the range of Hb-O2 saturation in the blood reduces HCO3- dehydration at the gills during greater work loads elevating arterial P(CO2) levels, leading to an increase in HCO3- buffer capacity of the blood and tissues. PMID- 10701711 TI - Vitamin E supplementation and endurance exercise: are there benefits? AB - It has been widely noted that vitamin E shows numerous beneficial effects through and beyond its antioxidative properties; consequently, vitamin E is expected to prevent degenerative diseases. In the field of sports medicine, many studies dealing with vitamin E have been conducted originally from the point of view of its effects on physical performance. Although some earlier studies indicated that vitamin E supplementation could improve physical performance, defects in the study design or statistical analysis were pointed out at a later time. The majority of subsequent well controlled studies have reported no significant effect on physical performance from vitamin E supplementation. Recent studies suggest that endurance exercise may promote free radical generation in the body, and vitamin E may play an important role in preventing the free radical damage associated with endurance exercise. Although there is evidence of free radical involvement in exercise-induced muscle injury, vitamin E supplementation might not be expected to prevent muscle damage caused by exercise in humans without a vitamin E deficiency. Since it is still unclear whether exercise induces lipid peroxidation in the human body, the beneficial effect of vitamin E supplementation on exercise-induced lipid peroxidation has not yet been established. However, it is proposed that as a result of exercise vitamin E may be mobilised from store tissues and redistributed in the body to prevent oxidative damage. Therefore, we are convinced that vitamin E contributes to preventing exercise-induced lipid peroxidation. It has also been indicated that strenuous endurance exercise may enhance the production of oxidised low density lipoprotein (LDL), which plays a key role in the initiation and progression of atherosclerosis. It is also suggested that this enhanced production of oxidised LDL could be reduced if a higher vitamin E status is maintained. Supplementation with 100 to 200mg of vitamin E daily can be recommended for all endurance athletes to prevent exercise-induced oxidative damage and to reap the full health benefits of exercise. PMID- 10701712 TI - Analgesia following exercise: a review. AB - Over the past 20 years a number of studies have examined whether analgesia occurs following exercise. Exercise involving running and cycling have been examined most often in human research, with swimming examined most often in animal research. Pain thresholds and pain tolerances have been found to increase following exercise. In addition, the intensity of a given pain stimulus has been rated lower following exercise. There have been a number of different noxious stimuli used in the laboratory to produce pain, and it appears that analgesia following exercise is found more consistently for studies that used electrical or pressure stimuli to produce pain, and less consistently in studies that used temperature to produce pain. There is also limited research indicating that analgesia can occur following resistance exercise and isometric exercise. Currently, the mechanism(s) responsible for exercise-induced analgesia are poorly understood. Although involvement of the endogenous opioid system has received mixed support in human research, results from animal research seem to indicate that there are multiple analgesia systems, including opioid and non-opioid systems. It appears from animal research that properties of the exercise stressor are important in determining which analgesic system is activated during exercise. PMID- 10701713 TI - The effect of aerobic exercise training on the lipid-lipoprotein profile of children and adolescents. AB - Longitudinal paediatric population studies have provided evidence that the risk factor theory may be extended to children and adolescents. These studies could assist in identifying individuals at increased coronary risk. Numerous studies have focused on the effects of regular exercise on the paediatric lipoprotein profile, a recognised primary risk factor, with equivocal results. Cross sectional comparisons of dichotomised groups provide the strongest evidence of an exercise effect. 'Trained' or 'active' children and adolescents demonstrate 'favourable' levels of high density lipoprotein-cholesterol (HDL-C), triacylglycerol, total cholesterol (TC)/HDL-C and low density lipoprotein cholesterol (LDL-C)/HDL-C, whilst TC is generally unaffected. The evidence regarding LDL-C in these studies is equivocal. A possible self-selection bias means that a cause-effect relationship between exercise and the lipoprotein profile cannot be readily established from this design. Correlational studies are difficult to interpret because of differences in participant characteristics, methods employed to assess peak oxygen uptake and habitual physical activity (HPA), and the statistical techniques used to analyse multivariate data. Directly measured cardiorespiratory fitness does not appear to be related to lipoprotein profiles in the children and adolescents studied to date, although there are data to the contrary. The relationship with HPA is more difficult to decipher. The evidence suggests that a 'favourable' lipoprotein profile may be related to higher levels of HPA, although differences in assessment methods preclude a definitive answer. While few prospective studies exist, the majority of these longitudinal investigations suggest that imposed regular exercise has little, if any, influence on the lipoprotein levels of children and adolescents. However, most prospective studies have several serious methodological design weaknesses, including low sample size, inadequate exercise training volume and a lack of control individuals. Recent studies have suggested that increases in HDL-C and reductions in LDL-C may be possible with regular exercise. The identification of a dose-response relationship between exercise training and the lipoprotein profile during the paediatric years remains elusive. PMID- 10701714 TI - Evaluation of the ergogenic properties of ginseng: an update. AB - Ginseng has been used in the Orient for several thousand years as an 'adaptogenic' as well as a 'restorative' agent. It has been used to treat nervous disorders, anaemia, wakefulness, dyspnoea, forgetfulness and confusion, prolonged thirst, decreased libido, chronic fatigue, angina and nausea. Although the mechanisms underlying the alleged effects of ginseng remain to be elucidated, there is an extensive animal literature dealing with the effects of ginseng on the cardiovascular system, central nervous system, endocrine system, metabolism, and immune system. In our previous review dealing with the efficacy of ginseng, we concluded that while studies with animals show that ginseng, or its active components, may prolong survival to physical or chemical stress, there is generally a lack of controlled research demonstrating the ability of ginseng to improve or prolong performance in fatigued humans. In this review, we extend our earlier analysis on the potential efficacy of ginseng use in the enhancement of physical performance and modification of fatigue states. Our analysis reveals that published literature appearing since our earlier review has not resolved the equivocal nature of research evidence involving animals or humans. Also, the lack of unanimity in this research can be explained on the basis of various methodological problems such as inadequate sample size and lack of double-blind, control and placebo paradigms. In addition, the absence of acceptable approaches to the problem of 'sourcing', in concert with an absence of compliance data in human research, further complicates the interpretation of this research literature. Nevertheless, the use of ginseng continues to grow, and current sales are estimated to be over $US300 million annually. There is clearly a need for systematic research dealing with the efficacy of ginseng, and this research needs to take into account basic, fundamental design considerations if there is to be any hope of establishing whether or not ginseng possesses efficacy. PMID- 10701716 TI - Measuring physical activity in peripheral arterial disease: a comparison of two physical activity questionnaires with an accelerometer. AB - Peripheral arterial disease (PAD)-related exertional leg pain may limit physical activity, thereby contributing to mobility loss and increasing cardiovascular morbidity and mortality in men and women with PAD. The objectives of this study were: (1) to compare objectively measured physical activity levels between patients with and without PAD, (2) to assess the validity of two physical activity questionnaires in patients with PAD. Twenty PAD patients from a noninvasive vascular laboratory and 21 patients without PAD from a general medicine practice wore an accelerometer continuously for 7 days to measure physical activity objectively. After 7 days, participants completed the leisure time physical activity questionnaire (LTPAQ), derived from the Health Interview Survey, (continued on next page)and the Stanford 7-day physical activity recall questionnaire (PARQ). PAD participants had markedly lower physical activity levels than non-PAD participants as measured by accelerometer (803 kcal/week +/ 364 (range=284-2,000, median=708) vs 1,750 kcal/week +/-1,296 (range=882-6,586, median=1,278), p<0.001). For the LTPAQ, physical activity levels in PAD and non PAD participants were 609 kcal/week +/-576 (range=0-2,085, median=529) vs 832 kcal/week +/-784 (range=53-2,820, median= 623), p=0.128. For the PARQ, physical activity levels in PAD and non-PAD participants were 234 METS/week +/-21 (range=214-301, median=229) vs 238 METS/week +/- 11 (range=225-268, median=234), p=0.454, respectively. Pearson's correlation coefficient for the association between the accelerometer and the log-transformed LTPAQ measure was 0.419 (p=0.006). Pearson's correlation coefficient was 0.348 for the association between the accelerometer and the log-transformed PARQ measure of physical activity (p=0.026). In conclusion, PAD patients have significantly lower physical activity levels than non-PAD patients. Two commonly used physical activity questionnaires were less sensitive than objective measurement to the association between PAD and inactivity. PMID- 10701715 TI - Chronic Achilles tendinosis: recommendations for treatment and prevention. AB - Chronic Achilles tendinosis is a condition with an unknown aetiology and pathogenesis that is often, but not always, associated with pain during loading of the Achilles tendon. Histologically, there are no inflammatory cells, but increased amounts of interfibrillar glycosaminoglycans and changes in the collagen fibre structure and arrangement are seen. In situ microdialysis has confirmed the absence of inflammation. It is a condition that is most often seen among recreational male runners aged between 35 and 45 years, and it is most often considered to be associated with overuse. However, this condition is also seen in patients with a sedentary lifestyle. Chronic Achilles tendinosis is considered a troublesome injury to treat. Nonsurgical treatment most often includes a combination of rest, NSAIDs, correction of malalignments, and stretching and strengthening exercises, but there is sparse scientific evidence supporting the use of most proposed treatment regimens. It has been stated that, in general, nonsurgical treatment is not successful and surgical treatment is required in about 25% of patients. However, in a recent prospective study, treatment with heavy load eccentric calf muscle training showed very promising results and may possibly reduce the need for surgical treatment of tendinosis located in the midportion of the Achilles tendon. The short term results after surgical treatment are frequently very good, but in the few studies with long term follow-up there are signs of a possible deterioration with time. Calf muscle strength takes a long time to recover and, furthermore, a prolonged progressive calcaneal bone loss has been shown on the operated side up to 1 year after surgical treatment. PMID- 10701717 TI - Ventricular ectopic activity, left ventricular mass, hyperinsulinemia, and intracellular magnesium in normotensive patients with obesity. AB - Insulin resistance, a well-known feature of obesity, is associated with several pathological changes, which are potentially arrhythmogenic. Ventricular ectopic activity in normotensive obese patients has not been studied in detail. Therefore the authors designed a study to investigate potential relationships among ventricular ectopic activity, left ventricular mass, hyperinsulinemia, and intracellular magnesium concentration in obese patients. Thirty-two obese patients and 32 nonobese control subjects, who were referred to outpatient department because of ventricular ectopy, participated in the study. The groups were matched for age and gender. All had normal glucose tolerance. All subjects underwent a 75-g glucose tolerance test, and blood samples were obtained at 30, 60, and 120 minutes thereafter for determination of glucose and insulin concentrations. Echocardiography was performed and left ventricular mass index was calculated. The number of ventricular ectopic beats per hour (VEB/hour) was recorded by 24-hour ECG Holter monitoring. Plasma and erythrocyte magnesium concentrations were determined by atomic absorption spectrophotometer. Obese patients had higher body weight, body mass index, heart rate, and left ventricular mass index. Obese subjects had higher fasting insulin as well as insulin/glucose ratio and broader area under the curve of insulin (AUC-I) compared to nonobese subjects. Insulin sensitivity appeared to be lower in the obese group. Holter monitoring showed more VEB/hour in the obese group. Magnesium concentration in serum and in erythrocytes was lower in obese persons. In the obese group a positive correlation was found between left ventricular mass index and fasting insulin (r=0.345, p=0.027), insulin/glucose index (r=0.351, p=0.049), and AUC-I (r=0.405, p=0.011). The number of VEB/hour in obese patients was in positive correlation with age (r=0.681, p<0.001), left ventricular mass index (r=0.542, p=0.001), fasting insulin (r=0.380, p=0.016), and AUC-I (r=0.493, p=0.002) and in negative correlation with magnesium concentration in erythrocytes (r=-0.457, p=0.004). Multiple regression analysis showed that age and AUC-I are the only determinants of VEB/hour and together explained 56% of the variability in the obese subjects. It appears that in obese normotensive subjects, ventricular ectopic beats are related to age, insulin resistance, left ventricular mass index, and decreased intracellular magnesium content. PMID- 10701718 TI - Long-term follow-up of primary stenting with coil stent in acute myocardial infarction. AB - The authors evaluated clinical and angiographic outcomes for 1 year after primary stenting using coil stent for acute myocardial infarction. Twenty-eight patients underwent primary stenting with coil stent. Follow-up coronary arteriography at 3 months and 1 year was planned in all patients. Procedural success was achieved in 96%. There was no acute or subacute thrombosis. Minimal lumen diameter (MLD) was increased from 0.08 +/- 0.19 to 2.73 +/- 0.49 mm after stenting. MLD had decreased significantly for 3 months (MLD at 3 months: 2.03 +/- 0.86 mm, p = 0.001). On the other hand, MLD did not differ between 3-month; and 1-year follow up (MLD at 1 year: 2.26 +/- 0.73 mm, p = NS). Only one patient manifested reocclusion at 3-month follow-up. The cumulative restenosis rate and target lesion revascularization rate at 1-year follow-up were 25.9% (7/27) and 11.1% (3/27). Primary stenting using coil stent is safe and feasible in patients with acute myocardial infarction and may improve clinical outcome and decrease restenosis and target lesion revascularization rate. PMID- 10701719 TI - Effects of transvenous regional guanethidine block in the treatment of critical finger ischemia. AB - The objective of this study was to determine the effects of transvenous regional guanethidine block in the treatment of patients with critical finger ischemia. Twenty-seven patients (17 collagen vascular disease, four thromboangiitis obliterans, three embolism, three atherothrombosis) presenting with ischemic rest pain and/or ulcerations of the fingers received a single block with 5 mg guanethidine injected in 60 mL into the clinically more affected hand under 30 minutes of arterial arrest. Marked hyperemia was induced in the treated upper limb, increases (p < 0.01) in finger blood flow, finger skin temperature, and laser Doppler flux were higher and longer lasting than in forearm blood flow, persisting for a whole month. Effects in patients with ischemic finger ulcers were less pronounced than in those without, yet statistically significant increases of all evaluated parameters were observed in these patients too. No effects were seen in the contralateral untreated upper limb or in systemic blood pressure. Subjective symptoms (reduction of rest pain, numbness, vasospastic attacks) were improved in 25/27 (92.6%) patients, ischemic rest pain disappeared in 20/27 (74.1%), and complete healing of finger tip ulcerations within 1 month was achieved in 10/12 (83.3%) affected patients. No side effects were observed. This described method combines good clinical efficacy with lack of undesirable side effects and can be repeated easily. Therefore, this technique is recommended for broader clinical use. PMID- 10701720 TI - Influence of the ultrasound contrast agent Levovist on human nailfold capillary microcirculation. AB - Little is known about the behavior of ultrasound contrast microbubbles in human capillaries. The evaluation of circulatory effects of echo contrast media may bring valuable information for the interpretation of echo contrast phenomena in the human myocardium. In 12 healthy volunteers (aged 31 +/- 6.7 years; five women), nailfold capillaries were examined by means of TV microscopy. The authors investigated acral microcirculation at rest and after local cold application with and without saccharide-based microbubbles (10 mL Levovist 300 mg/mL IV). The mean blood flow velocity at rest was 1.18 +/- 0.18 mm/s (mean value +/-1 SD) and 1.11 +/- 0.11 mm/s (mean value +/- 1 SD) after the injection of Levovist (ns). One minute after local cold exposure a decrease of the blood flow velocity by 61% before and by 75% after intravenous Levovist was found. In both groups the cold induced decrease of blood flow velocity was statistically significant (p<0.01), whereas there was no significant difference in flow reaction between the two groups. No wall adhesion of blood cells or extravasation of contrast into the surrounding tissue was detected. After intravenous injection of a regular dose of saccharide-based microbubbles Levovist, no change of blood cell flow velocity and no wall adhesion or extravasation could be found at rest and after cold application in human nailfold capillaries. Since microcirculatory flow characteristics in the finger nailfold capillaries are not influenced by Levovist, it might be assumed also that myocardial blood flow behavior remains unchanged, so that this contrast agent may be used as a flow tracer for cardiac investigation. PMID- 10701721 TI - Morphologic and functional changes of left ventricle in dialyzed patients after treatment with recombinant human erythropoietin (r-HuEPO). AB - Dysfunction of the cardiovascular system is a common complication of chronic renal insufficiency. Many factors can cause left ventricular hypertrophy (LVH), and hypertension and anemia are among them. They play an important role in the pathogenesis of LVH as well as in the development of cardiac dysfunction. Echocardiography enables early detection of functional macrocirculatory changes as well as adequate measuring of cardiac structures and LV mass. Anemia of end stage chronic renal insufficiency (ESRD) is only one among its many complications and has complex pathogenesis; one of the primary factors causing anemia is insufficient production of erythropoietin, a leading factor in the production of erythropoiesis. Anemia correction with recombinant human erythropoietin (r-HuEPO) in ESRD has a positive effect on the cardiovascular system. In this study the authors examined the hemodynamic effect of erythropoietin in anemic patients undergoing hemodialysis and observed its positive effect on the cardiovascular system. Twenty-two patients were included in the study (13 men and 9 women) mean age x=39.5 years. All patients were dialyzed three times a week for 4 hours and were all (Abstract continued) treated, according to protocol, with r-HuEPO for 8 months. Left ventricular mass was measured by the Penn Convention formula. The authors noticed the effectiveness of this therapy through an increase of hemoglobin of 35% and of hematocrit of 34% and a direct effect on the cardiovascular system. Echocardiographic findings showed decrease of LV mass from 391 to 274 mg (30%). The correction of renal anemia with erythropoietin leads to structural microcirculatory changes and partial morphologic regression of preexistent LVH, which again leads to regression of cardiac dysfunctions and improved hemodynamic effect, physical capacity, and cardiopulmonary status, and ultimately better quality of life for dialyzed patients. PMID- 10701722 TI - Sustained classic clinical spectrum of thromboangiitis obliterans (Buerger's disease). AB - Thromboangiitis obliterans (TAO) has been reported to become less common in general population but more common in women, and in elderly patients. The authors looked at the clinical characteristics of TAO in Poland where there was no significant decrease of smoking and the extent of aging of the general population is less profound. They retrospectively reviewed the records of 377 patients with the diagnosis of TAO hospitalized in their institution from 1970 to 1995. If young smoking males demonstrated distal-extremity ischemia with no bruits audible over major arteries, upper limbs involvement, or superficial thrombophlebitis, the diagnosis of TAO was considered certain. When at least one of those criteria was missed, and in men older than 35 years, but in all females, typical arteriographic findings were required for the diagnosis of TAO. Connective-tissue disease, hyperlipidemia, diabetes, and hypercoagulable state were excluded. Three hundred forty-two men (91%), and 35 (9%) women had a mean age of 29.5 years at the onset of the disease (the oldest patient was 50 years old). The prevalence of TAO in southwest Poland is 8.1/100,000 and the incidence of the disease steadily declines; there was no increase of TAO in women. Three hundred thirty-seven (89%) experienced rest pain, 321 (85%) had ischemic necrosis, and 233 (62%) thrombophlebitis at some (continued on next page) time in the course of the disease. Raynaud's phenomenon occurred in only 39 patients (10%). Those patients who had quit smoking had a 50% decrease of the disease recurrences compared to their smoking period. Because the cause of declining incidence of TAO is obscure, the authors critically evaluated previously used explanations of this phenomenon. They did not confirm the observation of a change in the TAO clinical spectrum: occurrence in women did not increase, the aging of the TAO population was not observed. In Poland TAO is still a disease affecting the peripheral circulation of young smoking males with recurrent episodes of superficial thrombophlebitis and common involvement of the upper extremities; Raynaud's phenomenon is rather infrequent. Smoking cessation ameliorates the course of the disease but does not invariably stop further exacerbations. PMID- 10701723 TI - An unusual case of cortical blindness associated with aortography--a case report. AB - Transient cortical blindness is a rare but well-recognized benign complication of angiography that is due to neurotoxicity of the contrast agent. The blindness completely resolves in a few days. This report describes a patient suffering an unusual course of cortical blindness following aortography resulting in permanent, partial blindness. It was found that blindness was the consequence of bilateral occipital embolisms. This case emphasizes that cortical blindness associated with angiography does not always have a favorable outcome, since it might be, in rare cases, due to isolated occipital embolisms, which initially produce a clinical picture identical to that of the neurotoxic effect of contrast agents. PMID- 10701724 TI - Incomplete caval protection following suprarenal caval filter placement--a case report. AB - Inferior vena cava filters are increasingly utilized to prevent pulmonary emboli originating from lower extremity, pelvis, or caval thromboses. Unique indications exist for filter placement in the suprarenal portion of the vena cava. The largest reported experience in suprarenal position has involved the use of the Greenfield filter. Although unique in design, little differences have been described between the stainless steel and titanium version of this device. The authors present a case report of incomplete caval protection after successful placement of a stainless steel Greenfield filter in the suprarenal cava. Subtle differences between the titanium and the stainless steel Greenfield filter may exist and should be taken into consideration for placement in the suprarenal cava. Physiologic conditions supporting this premise are described. Further investigation between the two filter types appears justified. PMID- 10701725 TI - Deep venous thrombosis and pulmonary thromboembolism associated with a huge uterine myoma--a case report. AB - A 51-year-old woman with a large uterine myoma suffered from acute pulmonary thromboembolism. Venography revealed thrombosis in the right common iliac vein and almost complete obstruction of the left common iliac vein. The ascending lumbar vein showed collateral drainage. Treatment was initiated with continuous intravenous heparin sodium, and a Greenfield filter was inserted to prevent the extension of the pulmonary embolism during and after hysterectomy. After a total hysterectomy, venography revealed restoration of patency in the bilateral common iliac veins, and no flow was seen in the ascending lumbar vein. Thorough clinical examinations failed to identify any other prothrombotic conditions. These results suggest that a large uterine myoma compressed veins in the pelvis, and the resulting impaired blood flow caused deep venous thrombosis and pulmonary thromboembolism. PMID- 10701726 TI - Ruptured aneurysm of the sinus of Valsalva in association with persistent left superior vena cava--a case report. AB - A 58-year-old man presenting with chest pain and dyspnea was diagnosed by transesophageal echocardiography and cardiac catheterization to have the rare combination of ruptured aneurysm of noncoronary sinus of Valsalva into the right ventricle in association with persistent left superior vena cava. These defects were confirmed at cardiac surgery. This case shows the importance of complementary use of invasive and noninvasive methods together in the diagnosis of rare combinations of lesions. PMID- 10701727 TI - Internal jugular vein thrombosis, Lemierre's syndrome; oropharyngeal infection with antibiotic and anticoagulation therapy--a case report. AB - The authors present a case of Lemierre's syndrome that is an uncommon septic thrombophlebitis of the internal jugular vein. A 31-year-old man developed pharyngeal pain one month before hospital admission when he suffered from a severe headache and painful swelling of the left side of his neck. He was diagnosed with tonsillitis. Contrast-enhanced computed tomography and magnetic resonance imaging of the neck revealed the presence of an occlusive thrombosis of the left internal jugular vein and an inflamed mesopharynx. His symptoms and the jugular vein thrombus showed remarkable improvement after administration of antibiotic and anticoagulation therapy. No pulmonary embolism or other metastatic infection were observed. It was suggested that accurate diagnosis during early treatment is essential to obtain a successful prognosis for Lemierre's syndrome. PMID- 10701728 TI - Regarding an unusual case of bilateral coronary artery fistulas--a case report. PMID- 10701729 TI - Intraoperative radiation therapy in resected bile duct cancer. PMID- 10701730 TI - Granulocyte macrophage-colony stimulating factor (GM-CSF) and sucralfate in prevention of radiation-induced mucositis: a prospective randomized study. AB - PURPOSE: To compare subcutaneously given molgramostim (GM-CSF) and sucralfate mouth washings to sucralfate mouth washings in prevention of radiation-induced mucositis. METHODS AND MATERIALS: Forty head and neck cancer patients were randomly assigned to use either GM-CSF and sucralfate (n = 20) or sucralfate alone (n = 20) during radiotherapy. Sucralfate was used as 1.0 g mouth washing 6 times daily after the first 10 Gy of radiotherapy, and 150-300 microg GM-CSF was given subcutaneously. The grade of radiation mucositis and blood cell counts were monitored weekly. Salivary lactoferrin was measured as a surrogate marker for oral mucositis. RESULTS: We found no significant difference between the molgramostim and the control groups in the oral mucositis grade, oral pain, use of analgesic drugs, weight loss, or survival. The median maximum neutrophil counts (median, 9.2 x 10(9)/L vs. 5.9 x 10(9)/L, p = 0.0005), eosinophil counts (median, 1.3 x 10(9)/L vs. 0.2 x 10(9)/L, p = 0.0004), and salivary lactoferrin concentrations were higher in patients who received GM-CSF. The most common toxicities in the GM-CSF plus sucralfate group were skin reactions at the GM-CSF injection site (65%), fever (30%), bone pain (25%), and nausea (15%), whereas the toxicity of sucralfate given alone was minimal. CONCLUSION: We found no evidence indicating that subcutaneously given GM-CSF reduces the severity of radiation induced mucositis. PMID- 10701731 TI - Oral glutamine to alleviate radiation-induced oral mucositis: a pilot randomized trial. AB - PURPOSE: To evaluate the influence of oral glutamine on radiation-induced oral mucositis in the radiotherapy of head and neck cancer. METHODS AND MATERIALS: From July 1997 through June 1998, 17 patients with head and neck cancer receiving primary or adjuvant irradiation were randomized to either glutamine suspension (16 g in 240 ml normal saline) (n = 8) or placebo (normal saline) (n = 9) arm. Patients were instructed to swish the test solutions (30 ml) four times per day. All patients received half-mouth irradiation at least. Patients were treated 1.8 Gy per fraction daily, 5 days a week. We evaluated the grading of oral mucositis daily fraction at each day of treatment until 45 Gy/25 fractions. World Health Organization (WHO) step analgesic medication and body weight change were compared between the two arms. RESULTS: The duration of objective oral mucositis > or = Grade 1 (p = 0.0097), Grade 2 (p = 0.0232), and Grade 3 (p = 0.0168) was shorter in the glutamine arm. Mean maximum grade of objective oral mucositis was less severe in the glutamine arm (1.6 vs. 2.6) (p = 0.0058). Glutamine did not reduce the duration and severity of subjective oral mucositis except for duration > or = Grade 3 (p = 0.0386). In the analysis of mean maximum WHO step of analgesic medication, there was no statistical difference (p = 0.5374) between the two arms. Mean body weight change was also not significantly different (p = 0.8070). CONCLUSIONS: Oral glutamine may significantly reduce the duration and severity of objective oral mucositis during radiotherapy. It may shorten the duration of > or = Grade 3 subjective mucositis. PMID- 10701732 TI - Lymph node metastasis in maxillary sinus carcinoma. AB - PURPOSE: To evaluate the incidence and prognostic significance of lymph node metastasis in maxillary sinus carcinoma. METHODS AND MATERIALS: We reviewed the records of 97 patients treated for maxillary sinus carcinoma with radiotherapy at Stanford University and at the University of California, San Francisco between 1959 and 1996. Fifty-eight patients had squamous cell carcinoma (SCC), 4 had adenocarcinoma (ADE), 16 had undifferentiated carcinoma (UC), and 19 had adenoid cystic carcinoma (AC). Eight patients had T2, 36 had T3, and 53 had T4 tumors according to the 1997 AJCC staging system. Eleven patients had nodal involvement at diagnosis: 9 with SCC, 1 with UC, and 1 with AC. The most common sites of nodal involvement were ipsilateral level 1 and 2 lymph nodes. Thirty-six patients were treated with definitive radiotherapy alone, and 61 received a combination of surgical and radiation treatment. Thirty-six patients had neck irradiation, 25 of whom received elective neck irradiation (ENI) for N0 necks. The median follow-up for alive patients was 78 months. RESULTS: The median survival for all patients was 22 months (range: 2.4-356 months). The 5- and 10-year actuarial survivals were 34% and 31%, respectively. Ten patients relapsed in the neck, with a 5-year actuarial risk of nodal relapse of 12%. The 5-year risk of neck relapse was 14% for SCC, 25% for ADE, and 7% for both UC and ACC. The overall risk of nodal involvement at either diagnosis or on follow-up was 28% for SCC, 25% for ADE, 12% for UC, and 10% for AC. All patients with nodal involvement had T3-4, and none had T2 tumors. ENI effectively prevented nodal relapse in patients with SCC and N0 neck; the 5-year actuarial risk of nodal relapse was 20% for patients without ENI and 0% for those with elective neck therapy. There was no correlation between neck relapse and primary tumor control or tumor extension into areas containing a rich lymphatic network. The most common sites of nodal relapse were in the ipsilateral level 1-2 nodal regions (11/13). Patients with nodal relapse had a significantly higher risk of distant metastasis on both univariate (p = 0.02) and multivariate analysis (hazard ratio = 4.5, p = 0.006). The 5-year actuarial risk of distant relapse was 29% for patients with neck control versus 81% for patients with neck failure. There was also a trend for decreased survival with nodal relapse. The 5-year actuarial survival was 37% for patients with neck control and 0% for patients with neck relapse. CONCLUSION: The overall incidence of lymph node involvement at diagnosis in patients with maxillary sinus carcinoma was 9%. Following treatment, the 5-year risk of nodal relapse was 12%. SCC histology was associated with a high incidence of initial nodal involvement and nodal relapse. None of the patients presenting with SCC histology and N0 necks had nodal relapse after elective neck irradiation. Patients who had nodal relapse had a higher risk of distant metastasis and poorer survival. Therefore, our present policy is to consider elective neck irradiation in patients with T3-4 SCC of the maxillary sinus. PMID- 10701733 TI - Treatment of locally advanced adenoid cystic carcinoma of the head and neck with neutron radiotherapy. AB - PURPOSE: To examine the efficacy of fast neutron radiotherapy for the treatment of locally advanced and/or recurrent adenoid cystic carcinoma of the head and neck and to identify prognostic variables associated with local-regional control and survival. METHODS AND MATERIALS: One hundred fifty-nine patients with nonmetastatic, previously unirradiated, locally advanced, and/or recurrent adenoid cystic carcinoma (ACC) of the head and neck region were treated with fast neutron radiotherapy during the years 1985-1997. One hundred fifty-one patients had either unresectable disease, or gross residual disease (GRD) after an attempted surgical extirpation. Eight patients had microscopic residual disease and were analyzed separately. Sixty-two percent of patients had tumors arising in minor salivary glands, 29% in major salivary glands, and 9% in other sites such as the lacrimal glands, tracheal-bronchial tree, etc. Fifty-five percent of patients were treated for postsurgical recurrent disease and 13% of patients had lymph node involvement at the time of treatment. The median duration of follow-up was 32 months (range 3-142 months). Actuarial curves for survival, cause-specific survival, local-regional control, and the development of distant metastases are presented for times out to 11 years. RESULTS: The 5-year actuarial local-regional tumor control rate for the 151 patients with GRD was 57%; the 5-year actuarial overall survival rate was 72%; and the 5-year actuarial cause-specific survival rate was 77%. Variables associated with decreased local-regional control in the patients with GRD as determined by multivariate analysis included base of skull involvement (p < 0.01) and biopsy only versus an attempted surgical resection prior to treatment (p = 0.03). Patients without these negative factors had an actuarial local-regional control rate of 80% at 5 years. Patients with microscopic residual disease (n = 8) had a 5-year actuarial local-regional control rate of 100%. Base of skull involvement (p < 0.001), lymph node metastases at the time of treatment (p < 0.01), biopsy only prior to neutron radiotherapy (p = 0.03), and recurrent tumors (p = 0.04) were found to be associated with a diminished cause-specific survival as ascertained by multivariate analysis. Patients with base of skull involvement and positive lymph nodes at presentation had an increased rate of the development of distant metastases at 5 years, (p < 0.01 and p < 0.001, respectively). No statistical difference in outcome was observed between major and minor salivary gland sites. CONCLUSIONS: Fast neutron radiotherapy is an effective treatment for locally advanced ACC of the head and neck region with acceptable toxicity. Further improvements in local-regional control are not likely to impact survival until more effective systemic agents are developed to prevent and/or treat distant metastatic disease. PMID- 10701734 TI - Scrutiny of the ASTRO consensus definition of biochemical failure in irradiated prostate cancer patients demonstrates its usefulness and robustness. American Society for Therapeutic Radiology and Oncology. AB - PURPOSE: The goals of this study are: (1) to establish the robustness of the Fox Chase Cancer Center (FCCC) and the American Society for Therapeutic Radiology and Oncology (ASTRO) consensus definitions of failure by comparing biochemical estimates under various modifications of the censoring and failure time components to their respective unaltered definitions; (2) to isolate the source of variation between the two definitions of failure; and (3) to describe the hazard of failure over time for each definition. METHODS: Between May 1989 and May 1997, 670 men were treated at Fox Chase Cancer Center for localized prostate cancer using three-dimensional conformal radiation therapy (3DCRT). These men were stratified into three groups for analysis: 111 men treated with adjuvant hormones; 204 men treated with radiation therapy alone and presenting with more favorable prognosis tumor characteristics; 255 men treated with radiation therapy alone and presenting with less favorable prognosis tumor characteristics. For each group, biochemical failure was estimated and compared using the FCCC and ASTRO definitions of failure. The robustness of each definition was evaluated by comparing estimates under the definition as stated to those under various modifications of the censoring and failure components. Analyses were also performed while excluding slow-progressing patients. To isolate the source of variation between the two failure definitions, estimates were compared for patients with agreement in failure status. Estimates of biochemical failure, and thus hazard rates, were made using Kaplan-Meier methodology. RESULTS: ASTRO biochemical failure estimates were higher than the FCCC failure estimates in the first 5 years post-treatment. Beyond 5 years, ASTRO estimates level off, while the FCCC failure estimates continued to increase. These failure patterns were similar in all patient groups; however, patients treated with adjuvant hormones had a much higher risk of failure immediately following treatment under the ASTRO definition. Modifying the censoring pattern had little effect on failure estimates in any patient group, regardless of definition used. The exclusion of patients with slow prostate-specific antigen (PSA) doubling time did not result in biochemical estimates that differed significantly from those for all patients. The analysis of patients with agreement in failure status continued to demonstrate significant differences in estimates between the two definitions, and thus differences may be attributed to the specification of time to failure. For all patient groups, hazard rates were dependent upon failure definition: under the FCCC failure definition, patients were at constant risk of failure over the observation period; under the ASTRO failure definition, patients were at risk of failure during the first 4 years following treatment, and then at low risk of failure beyond 5 years. CONCLUSIONS: Both FCCC and ASTRO failure definitions were robust to modifications in censoring and the inclusion of patients with long doubling times. The ASTRO failure definition was robust to specifying the time to failure at first rise, as opposed to midway between nadir and first rise. Similarities in estimates for all patients versus patients with agreeing failure status suggest that differences in failure definition lie in the specification of time to failure. The ASTRO definition of failure is more appropriate because it does not impose an empirical failure marker but is based on the initiation of biochemical rise. The use of the ASTRO consensus definition demonstrated little risk of biochemical failure 4 years beyond treatment. The ASTRO failure definition should be adopted in all research involving biochemical failure analysis of men treated with radiation therapy. PMID- 10701735 TI - Higher than standard radiation doses (> or =72 Gy) with or without androgen deprivation in the treatment of localized prostate cancer. AB - PURPOSE: To study the effect on biochemical relapse-free survival (bRFS) and clinical disease-free survival of radiation doses delivered to the prostate and periprostatic tissues for localized prostate cancer. METHODS AND MATERIALS: A total of 1041 consecutive localized prostate cancer cases treated with external beam radiotherapy (RT) at our institution between 7/86 and 2/99 were reviewed. All cases had available pretreatment parameters including pretreatment prostate specific antigen (iPSA), biopsy Gleason score (bGS), and clinical T stage. The median age was 69 years. Twenty-three percent of cases (n = 238) were African American. The distribution by clinical T stage was as follows: T1 in 365 cases (35%), T2 in 562 cases (54%), and T3 in 114 cases (11%). The median iPSA level was 10.1 ng/ml (range: 0.4-692.9). The distribution by biopsy Gleason score (bGS) was as follows: < or =6 in 580 cases (56%) and > or =7 in 461 cases (44%). Androgen deprivation (AD) in the adjuvant or neoadjuvant setting was given in 303 cases (29%). The mean RT dose was 71.9 Gy (range: 57.6-78.0 Gy). The median RT dose was 70.2 Gy, with 458 cases (44%) receiving at least 72.0 Gy. The average dose in patients receiving <72 Gy was 68.3 Gy (median 68.4) versus 76.5 Gy (median 78.0) for patients receiving > or =72 Gy. The mean follow-up was 38 months (median 33 months). The number of follow-up prostate-specific antigen (PSA) levels available was 5998. RESULTS: The 5- and 8-year bRFS rates were 61% (95% CI 55-65%) and 58% (95% CI 51-65%), respectively. The 5-year bRFS rates for patients receiving radiation doses > or =72 Gy versus <72 Gy were 87% (95% CI 82 92%) and 55% (95% CI 49-60%), respectively. The 8-year bRFS rates for patients receiving radiation doses > or =72 Gy versus <72 Gy were 87% (95% CI 82-92%) and 51% (95% CI 44-58%), respectively (p < 0.001). A multivariate analysis of factors affecting bRFS was performed using the following parameters: age (continuous variable), race, T-stage (T1-T2 vs. T3), iPSA (continuous variable), bGS (< or =6 vs. > or =7), use of AD (yes vs. no), radiation technique (conformal versus standard), and radiation dose (continuous variable). T-stage (p < 0.001), iPSA (p < 0.001), bGS (p < 0.001), and RT dose (p < 0.001) were independent predictors of outcome. Age (p = 0.74), race (p = 0.96), radiation technique (p = 0.15), and use of AD (p = 0.31) were not. We observed 11% clinical failures (local, distant, or both) at 5 years and 15% at 8 years for the entire cohort. There was a statistically significant improvement with higher radiation doses (p = 0.032). The 5-year clinical relapse rates for patients receiving > or =72 Gy versus <72 Gy were 5% and 12%, respectively. The 8-year clinical relapse rates for patients receiving radiation doses > or =72 Gy versus <72 Gy were 5% and 17%, respectively (p = 0.026). CONCLUSION: Patients receiving radiation doses exceeding 72 Gy had significantly better bRFS and clinical disease-free survival rates. Although results need to be confirmed with longer follow-up, these preliminary results are extremely encouraging. If these results are confirmed by other institutions and by longer follow-up, RT doses exceeding 72 Gy should be considered as standard of care. PMID- 10701736 TI - Short-course intensity-modulated radiotherapy for localized prostate cancer with daily transabdominal ultrasound localization of the prostate gland. AB - PURPOSE: To present our initial observations on the clinical feasibility of the technique of short-course intensity-modulated radiotherapy (SCIM-RT) in the treatment of localized prostate cancer coupled with daily transabdominal ultrasound localization of the prostate. The proposed regimen consists of a hypofractionated course delivering 70.0 Gy in 28 fractions. METHODS AND MATERIALS: The treatment data of the first 51 patients treated with SCIM-RT at the Cleveland Clinic Foundation are presented in this report. The technique consisted of intensity-modulated radiotherapy using 5 static fields (anterior, 2 laterals, and 2 anterior obliques). Inverse plans were generated by the Corvus treatment-planning system. The treatment delivery was performed with a Varian Dynamic Multileaf Collimator. The target was the prostate only in patients with low-risk disease (stage T1-T2, pretreatment PSA < or =10, and biopsy Gleason < or =6). The target was the prostate and seminal vesicles in patients with high-risk disease (stage T3 or pretreatment PSA > 10 or biopsy Gleason > or =7). In the Corvus planning system, the margins for the planning target volume (PTV) were 4 mm posteriorly, 8 mm laterally, and 5 mm in all other directions. A total of 70.0 Gy (mean prostate dose approximately 75 Gy) was prescribed in all cases at 2.5 Gy per fraction to be delivered in 28 fractions over 5 1/2 weeks. Prior to treatment delivery, the patients were minimally immobilized on the treatment table, only using lasers and skin marks. The location of the prostate gland was verified daily with the BAT transabdominal ultrasound system and patient position adjustments were performed accordingly. Fifty-one patients completed therapy between October 1998 and May 1999. RESULTS: The dose was prescribed to an isodose line ranging from 82.0% to 90.0% (mean: 87.2%). The range of the individual prostate mean doses was 73.5 to 78.5 Gy (average: 75.3 Gy). The range of the maximum doses was 77.4 to 84.5 Gy (average: 80.2 Gy). The range of the minimum doses was 64.3 to 69.2 Gy (average: 67.5 Gy). The average time for the prostate position verification and alignment of the prostate using the BAT system was 5 minutes. The entire localization/alignment process was performed by the radiation therapists. The daily alignment images were automatically saved and reviewed by the radiation oncologist, a process similar to port film checks. The total treatment (beam-on) time was around 6 minutes using the 5 static intensity modulated fields. The mean and standard deviation (SD) of bladder volumes irradiated to 50, 60, and 70 Gy were as follows: 24 +/- 11 cc, 16 +/- 8 cc, and 8 +/- 6 cc. The mean and SD of rectal volumes irradiated to 50, 60, and 70 Gy were as follows: 22 +/- 11 cc, 15 +/- 8 cc, and 7 +/- 5 cc. The RTOG acute bladder toxicity scores were as follows: 0 in 3 (6%), 1 in 38 (74%), and 2 in 10 (20%). The RTOG acute rectal toxicity scores for SCIM-RT cases were as follows: 0 in 10 (20%), 1 in 33 (65%), and 2 in 8 (16%). No Grade 3 or 4 acute toxicities were observed. CONCLUSION: The delivery of our proposed hypofractionated-schedule SCIM RT in combination with daily target localization/alignment with the BAT transabdominal ultrasound system is clinically feasible. It is an alternative method of dose escalation in the treatment of localized prostate cancer. The proposed schedule would significantly increase convenience to patients due to the decrease in overall treatment time. Preliminary acute toxicity results are extremely encouraging. Long-term follow-up is needed to assess late complications and treatment efficacy. PMID- 10701737 TI - Benefits of adjuvant radiotherapy after radical resection of locally advanced main hepatic duct carcinoma. AB - PURPOSE: The objective of this study was to determine the benefits of adjuvant radiotherapy after radical resection of locally advanced main hepatic duct carcinoma (Klatskin tumor). METHODS AND MATERIALS: We conducted a retrospective review of 63 patients who underwent surgical resection of Stage IVA Klatskin tumor. Of the 63 patients, 47 had microscopic tumor residue (RT1). Twenty-eight of the 47 patients with RT1 were treated by adjuvant radiotherapy and the remaining 19 patients were treated exclusively by surgical resection. Seventeen of the 28 patients with RT1 were treated by both intraoperative radiotherapy (IORT) and postoperative radiotherapy (PORT); of the remaining 11 patients with RT1, 6 underwent resection and IORT, and 5 underwent resection and PORT. RESULTS: The major complication and 30-day operative death rates were significantly lower in the radiation group (9.5% and 0.0%, respectively) than in the resection alone group (28.5% and 9.5%, respectively). Of the eight 5-year survivors with RT1, 6 had adjuvant radiotherapy and the remaining 2 had resection alone. Adjuvant radiotherapy for patients with RT1 yielded significantly (p = 0.0141) higher 5 year survival rates (33.9%) than in the resection alone group (13.5 %). The best 5-year survival rate (39.2 %) was found in patients who underwent a combination of IORT and PORT after resection. The local-regional control rate was significantly higher in the adjuvant radiation group than in the resection alone group (79.2% vs. 31.2%). CONCLUSION: Our data clearly suggest the improved prognosis of patients with locally advanced Klatskin tumor by integrated adjuvant radiotherapy with IORT and PORT to complete gross tumor resection with acceptable treatment mortality and morbidity. PMID- 10701738 TI - Results of irradiation or chemoirradiation following resection of gastric adenocarcinoma. AB - PURPOSE: To evaluate the results of postoperative irradiation +/- chemotherapy for carcinoma of the stomach and gastroesophageal junction. METHODS AND MATERIALS: The records of 63 patients who underwent resection for stomach cancer were retrospectively reviewed. Twenty-five patients had complete resection with no residual disease but with high-risk factors for relapse. Twenty-eight had microscopic residual and 10 had gross residual disease. Doses of irradiation ranged from 39.6 to 59.4 Gy with a median dose of 50.4 Gy in 1.8 Gy fractions. Fifty-three of the 63 (84%) patients received 5-fluorouracil (5-FU)-based chemotherapy. RESULTS: The median duration of survival was 19.3 months for patients with no residual disease, 16.7 months for those with microscopic residual disease, and 9.2 months for those with gross residual disease (p = 0.01). The amount of residual disease also significantly impacted locoregional control (p = 0.04). Patients with linitis plastica did significantly worse in terms of survival, locoregional control, and distant control than those without linitis plastica. The use of 4 or more irradiation fields was associated with a significant decrease in the rate of Grade 4 or 5 toxicity when compared to the patients treated with 2 fields (p = 0.05). CONCLUSIONS: There was a significant association between survival and extent of residual disease after resection as well as the presence of linitis plastica. Distant failures are common and effective systemic therapy will be necessary to improve outcome. The toxicity of combined modality treatment appears to be reduced by using greater than 2 irradiation fields. PMID- 10701739 TI - Evaluating changes in stable chromosomal translocation frequency in patients receiving radioimmunotherapy. AB - PURPOSE: The lack of any consistent correlation between radioimmunotherapy (RIT) dose and observed hematologic toxicity has made it difficult to validate RIT radiation dose estimates to marrow. Stable chromosomal translocations (SCT) which result after radiation exposure may be a biologic parameter that more closely correlates with RIT radiation dose. Increases in the frequency of SCT are observed after radiation exposure and are highly correlated with absorbed radiation dose. SCT are cumulative after multiple radiation doses and conserved through an extended number of cell divisions. The purpose of this study was to evaluate whether increases in SCT frequency were detectable in peripheral lymphocytes after RIT and whether the magnitude of these increases correlated with estimated radiation dose to marrow and whole body. METHODS AND MATERIALS: Patients entered in a Phase I dose escalation therapy trial each received 1-3 intravenous cycles of the radiolabeled anti- carcinoembryonic antigen (CEA) monoclonal antibody, 90Y-chimeric T84.66. Five mCi of 111In-chimeric T84.66 was co-administered for imaging and biodistribution purposes. Blood samples were collected immediately prior to the start of therapy and 5-6 weeks after each therapy cycle. Peripheral lymphocytes were harvested after 72 hours of phytohemagglutinin stimulation and metaphase spreads prepared. Spreads were then stained by fluorescence in situ hybridization (FISH) using commercially available chromosome paint probes to chromosomes 3 and 4. Approximately 1000 spreads were evaluated for each chromosome sample. Red marrow radiation doses were estimated using the AAPM algorithm and blood clearance curves. RESULTS: Eighteen patients were studied, each receiving at least one cycle of therapy ranging from 5-22 mCi/m2. Three patients received 2 cycles and two patients received 3 cycles of therapy. Cumulative estimated marrow doses ranged from 9.2 to 310 cGy. Increases in SCT frequencies were observed after each cycle for both chromosomes 3 and 4 in 16 of 18 patients and in at least one chromosome for the remaining 2 patients. Cumulative increases in SCT frequencies ranged from 0.001 to 0.046 with no major differences observed between chromosomes 3 and 4. A linear correlation between cumulative marrow dose and increases in SCT frequencies was observed for chromosome 3 (R2 = 0.63) and chromosome 4 (R2 = 0.80). A linear correlation was also observed between increases in SCT frequency and whole body radiation dose or administered activity (R2 = 0.67-0.89). There was less correlation between observed decrease in wbc or platelet counts and marrow dose, whole body dose, or administered activity (R2 = 0.28-0.43). CONCLUSIONS: Increases in SCT frequency were detectable in peripheral lymphocytes after low dose-rate RIT irradiation. A linear correlation was observed between increases in SCT and marrow dose, whole body dose, and administered activity. This correlation provides one of the strongest radiation dose-response and activity-response relationships observed with RIT. The detection of SCT may therefore have application as an in situ integrating biodosimeter after RIT. This biologic parameter should prove useful in comparing effects on marrow for different therapeutic radionuclides and in comparing effects of RIT and external beam radiation doses on a cGy per cGy basis. As a result, this should allow for a more direct comparison between different methods of irradiation and in further refinement of radioimmunotherapy dose estimates and dosimetry methodology. PMID- 10701740 TI - Recovery of sperm production following radiation therapy for Hodgkin's disease after induction chemotherapy with mitoxantrone, vincristine, vinblastine, and prednisone (NOVP). AB - PURPOSE: The effect on human male fertility of radiotherapy following chemotherapy for the treatment of Hodgkin's disease (HD) is unknown. The impact of radiation therapy, given after mitoxantrone, vincristine, vinblastine, and prednisone (NOVP) chemotherapy, on sperm production is the focus of this study. PATIENTS: Serial semen analyses were performed on 34 patients with HD Stages I III before NOVP chemotherapy, after chemotherapy prior to radiation, and after radiation therapy. The most inferior radiation portals for patients were: mantle, 1 patient; paraaortic-spleen, 3 patients; upper abdomen, 24 patients; abdominal spade, 4 patients; and pelvic, 2 patients. Testicular radiation dose measurements were available for 20 of these patients. RESULTS: Before the start of radiation, 90% of patients were normospermic. The magnitude of the decline in sperm counts was related to the measured testicular dose and/or radiation fields employed. The minimum postradiotherapy counts, expressed as a fraction of pretreatment counts, for the various treatment groups are as follows: paraaortic-spleen, 20%; upper abdomen, testicular dose < 30 cGy, 4%; upper abdomen, testicular dose 30-39 cGy, 0.9%; abdominal spade, 0.02%; and pelvis, 0%. The time to nadir of sperm counts averaged 4.5 months. Recovery to normospermic levels occurred in 96% of patients, with most recovering to that level within 18 months. CONCLUSION: The effect of radiation following NOVP chemotherapy on sperm counts was no greater than would be expected with radiation therapy alone. In most patients, sperm counts recovered to levels compatible with normal fertility. PMID- 10701741 TI - Radiobiological considerations in the design of fractionation strategies for intensity-modulated radiation therapy of head and neck cancers. AB - PURPOSE: The dose distributions of intensity-modulated radiotherapy (IMRT) treatment plans can be shown to be significantly superior in terms of higher conformality if designed to simultaneously deliver high dose to the primary disease and lower dose to the subclinical disease or electively treated regions. We use the term "simultaneous integrated boost" (SIB) to define such a treatment. The purpose of this paper is to develop suitable fractionation strategies based on radiobiological principles for clinical trials and routine use of IMRT of head and neck (HN) cancers. The fractionation strategies are intended to allow escalation of tumor dose while adequately sparing normal tissues outside the target volume and considering the tolerances of normal tissues embedded within the primary target volume. METHODS AND MATERIALS: IMRT fractionation regimens are specified in terms of "normalized total dose" (NTD), i.e., the biologically equivalent dose given in 2 Gy/fx. A linear-quadratic isoeffect formula is applied to convert NTDs into "nominal" prescription doses. Nominal prescription doses for a high dose to the primary disease, an intermediate dose to regional microscopic disease, and lower dose to electively treated nodes are used for optimizing IMRT plans. The resulting nominal dose distributions are converted back into NTD distributions for the evaluation of treatment plans. Similar calculations for critical normal tissues are also performed. Methods developed were applied for the intercomparison of several HN treatment regimens, including conventional regimens used currently and in the past, as well as SIB strategies. This was accomplished by comparing the biologically equivalent NTD values for the gross tumor and regional disease, and bone, muscle, and mucosa embedded in the gross tumor volume. RESULTS: (1) A schematic HN example was used to demonstrate that dose distributions for SIB IMRT are more conformal compared to dose distributions when IMRT is divided into a large-field phase and a boost phase. Both were shown to be significantly superior compared to dose distributions obtained using conventional beams for the large-field phase followed by IMRT for the boost phase. (2) The relationship between NTD and nominal dose for HN tumors was found to be quite sensitive to the choice of tumor clonogen doubling time but relatively insensitive to other parameters. (3) For late effect normal tissues embedded in the tumor volume and assumed to receive the same dose as the tumor, the biologically equivalent NTD for the SIB IMRT may be significantly higher. (4) Normal tissues outside the target volume receive lower dose due to the higher conformality of the IMRT plans. The biologically equivalent NTDs are even lower due to the lower dose per fraction in the SIB strategy. CONCLUSIONS: IMRT dose distributions are most conformal when designed to be delivered as SIB. Using isoeffect radiobiological relationships and published HN data, fractionation strategies can be designed in which the nominal dose levels to the primary, regional disease and electively treated volumes are appropriately adjusted, each receiving different dose/fx. Normal tissues outside the treated volumes are at reduced risk in such strategies since they receive lower total dose as well as lower dose/fx. However, the late effect toxicities of tissues embedded within the primary target volume and assumed to receive the same dose as the primary may pose a problem. The efficacy and safety of the proposed fractionation strategies will need to be evaluated with careful clinical trials. PMID- 10701742 TI - Effect of incomplete repair on normal tissue complication probability in the spinal cord. AB - PURPOSE: To incorporate the effects of repair into a model for normal tissue complication probability (NTCP) in the spinal cord. METHODS AND MATERIALS: We used an existing model of NTCP for the spinal cord, based on a critical volume concept, into which we incorporated an incomplete repair (IR) scheme. Values for the repair half time were taken from existing experimental data. Repair corrections were expanded to account for the possibility of biphasic repair, namely the existence of long and short components of repair. RESULTS: We found that the model predicts complete repair to occur at approximately 15 hours, consistent with experimental data. The dependence of the model on the value of the dose per fraction was also studied. It was found that there is a sparing effect as the dose per fraction is decreased below 2 Gy. Surface plots of the NTCP as a function of both the interfraction interval (IFI) and the dose per fraction were generated. We investigated "iso-NTCP" curves, which may allow freedom in choice of treatment plans in terms of the optimal IFI and dose per fraction. As for biphasic repair, as the relative weights of the long and short components of repair were varied, the NTCP changed as well. The model showed little difference between mono- and bi-exponential repair in the time to complete repair, due to a dominance of the long component at long IFIs. CONCLUSIONS: Incorporating IR into NTCP modeling of the spinal cord is consistent with current experimental data. The concept of iso-NTCP curves is an approach which may be clinically useful. PMID- 10701743 TI - Early alteration in TGF-beta mRNA expression in irradiated rat liver. AB - PURPOSE: Radiation of the liver results in hepatic fibrosis as a late complication. TGF-beta has been implicated in the pathogenesis of fibrosis. The purpose of this study was to determine if there is early alteration in TGF-beta expression before hepatic fibrosis is evident. METHODS AND MATERIALS: Male Sprague-Dawley rats weighing 150-175 g were used. A partial volume of liver as large as a 2 cm x 1 cm rectangle was given a single dose of 25 Gy gamma radiation. Animals were sequentially sacrificed from day 0 to day 28. Appearance of hepatic fibrosis was tested by trichrome stain. Levels of mRNA expression of TGF-beta1 and TGF-beta3 were measured by Northern blot hybridization. Change in the level of mRNA expression was analyzed by densitometry. The expression of TGF betas was also analyzed in tissue with immunohistochemical staining. RESULTS: In trichrome-stained liver tissues obtained through 28 days after irradiation, there was no evidence of hepatic fibrosis. The expression of mRNAs of TGF-beta1 and TGF beta3 showed different features; The level of TGF-beta1 mRNA showed a gradual increase to the peak level of 3.6-fold at day 28, the last analyzed time. In contrast, TGF-beta3 mRNA showed an early peak of 4.8-fold at day 7 followed by a decrease to the lowest level of 1.6-fold at the last analyzed time. The expression of TGF-betas was also analyzed in tissue with immunohistochemical staining. At day 28 after radiation, increased positive staining for TGF-beta1 was observed around the central vein. Positive staining appeared mainly in nonhepatocytic cells. For TGF-beta3, the same pattern of positive staining was observed at day 7. CONCLUSION: The results of this study suggest that the alteration in mRNA expression of TGF-beta1 and TGF-beta3 occurs very early after radiation. The contrasting difference in the mRNA expression pattern of TGF-beta1 and TGF-beta3 suggests that interaction of the TGF-betas may be involved in fibrogenesis of irradiated liver, with TGF-beta1 as a positive regulator and TGF beta3 as a negative regulator. PMID- 10701744 TI - The new tubulin-inhibitor combretastatin A-4 enhances thermal damage in the BT4An rat glioma. AB - PURPOSE: To investigate the toxicity of combretastatin A-4 disodium phosphate (CA 4) and its vascular effects in the subcutaneous (s.c.) BT4An rat glioma, and additionally, to determine the tumor response of CA-4 combined with hyperthermia. METHODS AND MATERIALS: For assessment of drug toxicity, rats were given 50, 75, or 100 mg/kg CA-4 and followed by daily registration of weight and side effects. Interstitial tumor blood flow was determined by laser Doppler flowmetry in rats injected with 50 mg/kg CA-4. In the tumor response study we administered CA-4 50 mg/kg alone or combined with hyperthermia (waterbath 44 degrees C for 60 min) 0 or 3 h later. RESULTS: We found that CA-4, at a well-tolerated dose of 50 mg/kg, induced a considerable time-dependent decrease in the tumor blood flow. Tumor blood flow was reduced by 47-55% during the first 110 min after injecting CA-4, and thereafter remained decreased until the measurements were terminated. Administering CA-4 3 h before hyperthermia yielded the best tumor response and increased tumor growth time significantly compared with simultaneous administration of CA-4 and hyperthermia (p = 0.03). Interestingly, CA-4 alone did not influence tumor growth. CONCLUSION: CA-4 induces a gradual reduction in tumor blood flow which can be exploited to sensitize the BT4An tumor for hyperthermia. PMID- 10701745 TI - Alteration of sensitivity of intratumor quiescent and total cells to gamma-rays following thermal neutron irradiation with or without 10B-compound. AB - PURPOSE: Changes in the sensitivity of intratumor quiescent (Q) and total cells to gamma-rays following thermal neutron irradiation with or without 10B-compound were examined. METHODS AND MATERIALS: 5-Bromo-2'-deoxyuridine (BrdU) was injected to SCC VII tumor-bearing mice intraperitoneally 10 times to label all the proliferating (P) tumor cells. As priming irradiation, thermal neutrons alone or thermal neutrons with 10B-labeled sodium borocaptate (BSH) or dl-p boronophenylalanine (BPA) were administered. The tumor-bearing mice then received a series of gamma-ray radiation doses, 0 through 24 h after the priming irradiation. During this period, no BrdU was administered. Immediately after the second irradiation, the tumors were excised, minced, and trypsinized. Following incubation of tumor cells with cytokinesis blocker, the micronucleus (MN) frequency in cells without BrdU labeling (= Q cells at the time of priming irradiation) was determined using immunofluorescence staining for BrdU. The MN frequency in the total (P + Q) tumor cells was determined from the tumors that were not pretreated with BrdU before the priming irradiation. To determine the BrdU-labeled cell ratios in the tumors at the time of the second irradiation, each group also included mice that were continuously administered BrdU until just before the second irradiation using mini-osmotic pumps which had been implanted subcutaneously 5 days before the priming irradiation. RESULTS: In total cells, during the interval between the two irradiations, the tumor sensitivity to gamma rays relative to that immediately after priming irradiation decreased with the priming irradiation ranking in the following order: thermal neutrons only > thermal neutrons with BSH > thermal neutrons with BPA. In contrast, in Q cells, during that time the sensitivity increased in the following order: thermal neutrons only < thermal neutrons with BSH < thermal neutrons with BPA. The longer the interval between the two irradiations, the higher was the BrdU-labeled cell ratio at the second irradiation. The labeled cell ratio at the same time point after each priming irradiation increased in the following order: thermal neutrons only < thermal neutrons with BSH < thermal neutrons with BPA. CONCLUSION: These findings indicated that the use of 10B-compound, especially BPA, in thermal neutron irradiation causes the recruitment from the Q to P population. PMID- 10701746 TI - Intraarterial beta irradiation induces smooth muscle cell apoptosis and reduces medial cellularity in a hypercholesterolemic rabbit restenosis model. AB - PURPOSE: Ionizing radiation has been shown to be a powerful inhibitor of neointimal hyperplasia following arterial injury in several animal models of post percutaneous transluminal coronary angioplasty (post-PTCA) restenosis. This was previously shown to be associated with a reduction in smooth muscle cell (SMC) mitotic activity. This study evaluated the effect of intraarterial beta irradiation on the arterial wall SMC density and apoptosis. METHODS AND MATERIALS: Twenty-five carotid and 7 iliac arteries of hypercholesterolemic New Zealand white rabbits were injured using the Baumgartner technique. The impact of an 18 Gy beta radiation dose administered after balloon injury was studied and compared to a nonirradiated injured control group. The medial SMC density as well as the percentage of apoptotic cells were determined at 8 days, 21 days, and 6 weeks after injury using an automated computer-based software. Apoptotic cells were identified using in situ end-labeling of fragmented DNA. RESULTS: The values for medial apoptosis in control vs. irradiated arteries were: 0.014 +/- 0.023 vs. 0.23 +/- 0.28%, p = NS, at 8 days; 0.012 +/- 0.018 vs. 0.07 +/- 0.07%, p = 0.05, at 21 days; and 0 +/- 0 vs. 0.16 +/- 0.11%, p = 0.03, at 6 weeks. The overall incidence of medial apoptotic cells at all time points was 0.01 +/- 0.017 vs. 0.13 +/- 0.14% in controls and irradiated arteries respectively, p = 0.004. Medial SMC density was significantly decreased in irradiated arteries in comparison with controls (p < 0.01 at all time-points). CONCLUSIONS: Intraarterial beta irradiation stimulates medial SMC apoptosis in balloon-injured arteries. This, together with a decrease in SMC mitotic activity, contributes to a decrease in the arterial wall cellularity. PMID- 10701747 TI - Evaluation of compensation in breast radiotherapy: a planning study using multiple static fields. AB - PURPOSE: A method that uses electronic portal imaging to design intensity modulated beams for compensation in breast radiotherapy was implemented using multiple static fields in a planning study. We present the results of the study to verify the algorithm, and to assess improvements to the dosimetry. METHODS AND MATERIALS: Fourteen patients were imaged with computed tomography (CT) and on a treatment unit using an electronic portal imager. The portal imaging data were used to design intensity-modulated beams to give an ideal dose distribution in the breast. These beams were implemented as multiple static fields added to standard wedged tangential fields. Planning of these treatments was performed on a commercial treatment planning system (Target 2, IGE Medical Systems, Slough, U.K.) using the CT data for each patient. Dose-volume histogram (DVH) analysis of the plans with and without multileaf collimator (MLC) compensation was carried out. This work has been used as the basis for a randomized clinical trial investigating whether improvements in dosimetry are correlated with the reduction of long-term side effects from breast radiotherapy. RESULTS: The planning analysis showed a mean increase in target volume receiving 95-105% of prescribed dose of 7.5% (range -0.8% to 15.9%) when additional MLC compensation was applied. There was no change to the minimum dose for all 14 patient data sets. The change in the volume of breast tissue receiving over 105% of prescribed dose, when applying MLC compensation, was between -1.4% and 11.9%, with positive numbers indicating an improvement. These effects showed a correlation with breast size; the larger the breast the greater the amount of improvement. CONCLUSIONS: The method for designing compensation for breast treatments using an electronic portal imager has been verified using planning on CT data for 14 patients. An improvement was seen in planning when applying MLC compensation and this effect was greater the larger the breast size. PMID- 10701748 TI - Fractionated radiotherapy for metastatic bone pain: evidence-based medicine or...? PMID- 10701749 TI - CT of the entire chest for treatment planning of breast cancer to locate the internal mammary vessels is not necessary. PMID- 10701750 TI - RBE as a function of dose for effects on tissues and tumors assessed by the linear-quadratic model. PMID- 10701751 TI - Electroporation--a new possibility in boron neutron capture therapy? PMID- 10701752 TI - Somatotopic organization of cortical fields in the lateral sulcus of Homo sapiens: evidence for SII and PV. AB - The human somatosensory cortex in the Sylvian fissure was examined using functional magnetic resonance imaging to describe the number and internal organization of cortical fields present. Somatic stimuli were applied to the lips, face, hand, trunk, and foot of 18 human subjects. Activity patterns were transposed onto three-dimensional magnetic resonance images of the brain so that the location of activity associated with the different stimuli could be related to specific regions of the cortex. There were several consistent findings. First, there were three regions of activity in the lateral sulcus associated with stimulation of the contralateral body. The most consistent locus of activation was on the upper bank of the lateral sulcus, continuing onto the operculum. The other two areas, one rostral and one caudal to this large central area, were smaller and were activated less consistently. Second, when activity patterns in the large central area resulting from stimulation of all body parts were considered, this region appeared to contain two fields that corresponded in location and somatotopic organization to the second somatosensory area (SII) and the parietal ventral area (PV). Finally, patterns of activation within SII and PV were somewhat variable across subjects. Repeated within-subject stimulus presentation indicated that differences across subjects were not due to inconsistent stimulus presentation. Comparisons with other mammals suggest that some features of organization are found only in primates. It is hypothesized that these features may be associated with manual dexterity and coordination of the hands, a characteristic generally restricted to the primate lineage. PMID- 10701753 TI - Phenotypical characterization of the neurons expressing the D1 and D2 dopamine receptors in the monkey striatum. AB - The striatum is regulated by dopaminergic inputs from the substantia nigra. Several anatomical studies using in situ hybridization have demonstrated that in rodents, dopamine D1 and D2 receptors are segregated into distinct striatal efferent populations: dopamine D1 receptor into gamma-aminobutyric acid (GABA)/substance P striatonigral neurons, and dopamine D2 receptor into GABA/enkephalin striatopallidal neurons. The existence of such a segregation has not been investigated in primates. Therefore, to quantify the efferent striatal GABAergic neurons in the adult Cynomolgus monkey, we detected GAD67 mRNA expression while considering that only a minority of the GABAergic population is composed of interneurons. To characterize the peptidergic phenotype of the neurons expressing dopamine D1 or D2 receptors, we examined the mRNA coding for these receptors in the striatum at the cellular level using single- and double in situ hybridization with digoxigenin and 35S ribonucleotide probes. Double in situ hybridization demonstrated a high coexpression of dopamine D1 receptor and substance P mRNAs (91-99%) as well as dopamine D2 receptor and preproenkephalin A mRNAs (96-99%) in medium-sized neurons throughout the nucleus caudatus, putamen, and nucleus accumbens. Only a small subpopulation (2-5%) of the neurons that contained dopamine D1 receptor mRNA also expressed dopamine D2 receptor mRNA in all regions. Large-sized neurons known to be cholinergic expressed D2R mRNA. However, within the nucleus basalis of Meynert, the large cholinergic neurons expressed D2R mRNA, but the neurons producing enkephalin expressed neither D1R nor D2R mRNA. These results demonstrate that the striatal organizational pattern of D1 and D2 receptor segregation in distinct neuronal populations described in rodent also exists in primate. PMID- 10701754 TI - Endogenous dopaminergic regulation of horizontal cell coupling in the mammalian retina. AB - Horizontal cells in an isolated wholemount preparation of the mouse retina were injected with Lucifer yellow and neurobiotin to characterize both the pattern of gap junctional connectivity and its regulation by dopamine. The injected horizontal cells had a uniform morphology of a round cell body, a compact dendritic tree, and an axon, which could sometimes be traced to an expansive terminal system. The dendro-dendritic gap junctions between neighboring cells mediated both weak Lucifer yellow dye coupling and strong neurobiotin tracer coupling. The extent of the tracer coupling was decreased by either exogenous dopamine (100 microM) or cyclic adenosine monophosphate (cAMP) analogs and was significantly increased by the D1 antagonist SCH 23390 (10 microM). These results provide the first evidence in the mammalian retina that the gap junctions between horizontal cells are endogenously regulated by dopamine, which acts through D1 receptors to increase the intracellular cAMP. It has been proposed that the gap junctional coupling between horizontal cells is mediated by connexin 32 (Cx32), but the pattern and dopaminergic regulation of horizontal cell coupling were unaffected in Cx32-knockout mice, ruling out the possible involvement of Cx32. Every tracer-coupled horizontal cell showed calbindin immunoreactivity, and vice versa, providing strong evidence that the horizontal cells in the mouse retina comprise a single cell type. Like the axonless horizontal cells in other mammalian retinas, the axon-bearing horizontal cells in the mouse retina are coupled by gap junctions that are permeable to Lucifer yellow and dopamine sensitive, suggesting that the mouse horizontal cells have hybrid properties to compensate for the absence of axonless horizontal cells. PMID- 10701755 TI - Central lateral line pathways in a vocalizing fish. AB - The organization of the central lateral line pathways in the midshipman fish, Porichthys notatus, was identified following biotin injections into physiologically identified sites in the lateral line-recipient nucleus ventrolateralis in the midbrain. Retrogradely filled neurons are located primarily in nucleus medialis, the principal termination site of lateral line nerve afferents in the medulla, whereas terminal fields are mainly identified in isthmal (nucleus praeeminentialis) and diencephalic (posterior thalamic) nuclei. Compared to other teleosts, nucleus medialis has a distinctive cytoarchitecture in that most of its somata are confined to a dense cell plate adjacent to the fourth ventricle. Injections into nucleus ventrolateralis reveal a caudal (MEDc) and a rostral (MEDr) division of nucleus medialis which are separated by a dorsomedial division of the descending octaval nucleus. MEDc is further divisible into a caudal spherical and a more extensive rostral Purkinje-like cell division. MEDr includes a caudal division of Purkinje-like cells and a rostral division of round and fusiform-shaped cells that form a lateral band under the cerebellar crest. In addition to labeling terminals in nucleus ventrolateralis, biotin injections into MEDc and MEDr further distinguish intrinsic connectivity within nucleus medialis, and also label somata and terminals within other octavolateralis nuclei in the medulla. Injections into both nucleus ventrolateralis and nucleus medialis identify sites which may be processing information from both the auditory and lateral line systems, including the eighth nerve-recipient descending octaval nucleus, the acoustic division of the midbrain, and nucleus praeeminentialis which receives auditory input from the midbrain in midshipman. PMID- 10701756 TI - Development of the cholinergic, nitrergic, and GABAergic innervation of the cat dorsal lateral geniculate nucleus. AB - Cholinergic projections from the brainstem have been shown to be important modulators of visual activity in the dorsal lateral geniculate nucleus (dLGN) of the adult, but little is known about the role of these modulatory inputs during development. We examined the postnatal development of the cholinergic innervation of the dLGN by using an monoclonal antibody against choline acetyl transferase (ChAT). We also investigated the development of GABAergic interneurons in the dLGN by using an antibody against glutamic acid decarboxylase (GAD), and the developmental expression of brain nitric oxide synthase (BNOS) by using an antibody against this enzyme. We found that brainstem cells surrounding the brachium conjunctivum express ChAT at birth, although axons in the dLGN do not express ChAT until the end of the first postnatal week. Cholinergic synaptic contacts were observed as early as the second postnatal week. The number of axons stained with the ChAT antibody increased slowly during the subsequent weeks in the dLGN and reached adult levels by the eighth postnatal week. GABAergic interneurons were present at birth and reached their adult soma size by the third postnatal week. GABAergic fibers are dense at birth but change during development from a diffuse pattern to clustered arrangements that can be recognized as distinct rings of GAD staining by P35. Cellular expression of BNOS was seen within all dLGN laminae during development. The BNOS-stained cells are tentatively identified as interneurons because their soma sizes were similar to those of GAD-stained cells. Although cellular BNOS staining remained robust in the C1-3 laminae through adulthood, cellular expression of BNOS in the A laminae declined during the first five postnatal weeks and remains sparse in the adult. As cellular BNOS staining declined, there was a steady increase in BNOS-stained fibers, which paralleled the increase of ChAT-stained fibers that are known to colocalize BNOS in the adult. Our results emphasize the continued transformations of intrinsic as well as extrinsic innervation patterns that occur during the development, of the dLGN. PMID- 10701757 TI - Proline-specific dipeptidyl peptidase activity in the cockroach brain and intestine: partial characterization, distribution, and inactivation of tachykinin related peptides. AB - Proline-specific dipeptidyl peptidase (DPP IV) is an established enzyme known to degrade neuropeptides and peptide hormones in vertebrate tissues. DPP IV cleaves peptides at the Pro2 residue. Because several neuropeptides of the cockroach Leucophaea maderae, such as LemTRP-1 (APSGFLGVRamide), are potential substrates for this peptidase, we investigated the occurrence of proline-specific DPP activity in cockroach tissues. Partly purified DPP activity was characterized from the brain and midgut of L. maderae by using Gly-Pro-4-nitroanilide as a substrate. The highest activity was obtained from the membrane fraction of intestine; about 10 times less activity (per milligram protein) was obtained from brain membranes. A smaller amount of soluble DPP activity could also be identified in both tissues. Gel chromatography of the solubilized intestinal DPP activity revealed a molecular mass of about 75 kDa. The enzyme had a pH optimum of 8.5. Diprotin A (Ile-Pro-Ile) was an efficient competitive inhibitor of the cockroach DPP, whereas other known DPP inhibitors were found to be less potent. When incubated with human and cockroach DPP IV, the cleavage products of LemTRP-1 were AP and SGFLGVRamide (des-AP-LemTRP-1) as determined by mass spectrometry of high-performance liquid chromatography (HPLC)-purified peptide fragments. The AP fragment was biologically inactive and the des-AP fragment had a drastically reduced myostimulatory activity on the hindgut of L. maderae. The blowfly TRP callitachykinin-I (CavTK-I; APTAFYGVRamide) was cleaved in two steps to des-AP CavTK-I and desAPTA-CavTK-I, showing that cockroach DPP does not only liberate Xaa-Pro, but also Xaa-Ala dipeptides. The fragment desAPTA-CavTK-I was completely inactive on the cockroach hindgut. To compare, LemTRP-3 and CavTK-II, which lack a Pro2, were not cleaved by DPP IV. Enzyme histochemistry for DPP IV was performed on cryostat sections of brain and intestine with Gly-Pro-4-methoxy-2 naphthylamide as the substrate and Fast Blue B as the chromogen. Strong histochemical labeling was seen in specific neuropils of the brain such as the calyces of the mushroom bodies, the antennal glomeruli, and the central body. Also, the inner lining of the midgut (the peritrophic membrane) and the malpighian tubules were strongly labeled by reaction product. In both the brain and intestine, the enzyme-histochemical reaction was inhibited by diprotin A. PMID- 10701758 TI - Serotonergic connections to the ventral oral pontine reticular nucleus: implication in paradoxical sleep modulation. AB - Cholinergic microstimulation of the ventral part of the oral pontine reticular nucleus (vRPO) in cats generates and maintains paradoxical sleep. The implication of rostral raphe nuclei in modulating the sleep-wakefulness cycle has been based on their serotonergic projections to the pontine structures responsible for the induction of paradoxical sleep. However, serotonergic neurons have also been described in brainstem structures other than the raphe nuclei. The aim of the present work is to trace the origin of the serotonergic afferents to the vRPO and to the locus coeruleus alpha and perilocus coeruleu alpha nuclei, closely related with different paradoxical sleep events. Anterograde and retrograde horseradish peroxidase conjugated with wheat germ agglutinin tracer injections in these nuclei in cats were combined with serotonin antiserum immunohistochemistry. Our results demonstrate that reciprocal connections linking the rostral raphe nuclei to those oral pontine nuclei are scarce. The percentage of double-labeled neurons after injections in the vRPO averaged 18% in rostral raphe nuclei, while a level of 82% was estimated in mesopontine tegmentum structures other than the raphe nuclei. These results showed that the main source of serotonin to the vRPO, implicated in generation and maintenance of paradoxical sleep, arises from these mesopontine tegmentum structures. This indicates that the serotonin modulation of paradoxical sleep could be the result of activation in non-raphe mesopontine tegmentum structures. The existence of a complicated network in the vRPO, which maintains a balance between different neurotransmitters responsible for the generation and alternance of paradoxical sleep episodes, is discussed. PMID- 10701759 TI - Dynamic transformation of Bergmann glial fibers proceeds in correlation with dendritic outgrowth and synapse formation of cerebellar Purkinje cells. AB - Bergmann glia (BG) are unipolar cerebellar astrocytes, whose radial (or Bergmann) fibers associate with developing granule cells and mature Purkinje cells (PCs). In the present study, we investigated the morphodifferentiation of BG by immunohistochemistry for glutamate transporter GLAST and electron microscopy. GLAST was expressed widely in cerebellar radial glia/astrocytes during fetal and neonatal periods and became concentrated in BG postnatally. During the second postnatal week when PC dendrites grow actively, GLAST immunostaining revealed dynamic cytologic changes in Bergmann fibers in a deep-to-superficial gradient; Bergmann fibers traversing the external granular layer were stained as rod-like fibers, whereas in the molecular layer, the rod-like pattern was gradually replaced with a reticular meshwork. At postnatal day 10, the superficial rod-like domain was composed of glial fibrillary acidic protein (GFAP)-positive/GLAST positive straight fibers, forming cytoplasmic swellings and short filopodia. Along this domain, the tip of growing PC dendrites ascended vertically and entered the base of the external granular layer. The deeper reticular domain of Bergmann fibers was characterized by active expansion of GFAP-negative/GLAST positive lamellate processes, which surrounded PC synapses almost completely. Therefore, the transformation of Bergmann fibers proceeds in correlation with dendritic differentiation of PCs. The intimate PC-BG relationships during cerebellar development raise the possibility that a preexisting glial shaft could serve as a structural substrate that directs dendritic outgrowth toward the pial surface, whereas the successive formation of a reticular glial meshwork should lead to structural maturation of newly formed PC synapses. PMID- 10701761 TI - Secretion pattern of thyroid-stimulating hormone in dogs during euthyroidism and hypothyroidism. AB - In as many as one third of dogs with primary hypothyroidism a plasma thyrotropin (TSH) concentration within the reference range for euthyroid dogs is found. To determine whether this is due to fluctuations in the release of TSH, the plasma profiles of TSH were analyzed in 7 beagle bitches by collecting blood samples every 10 min for 6 hr, both before and after induction of primary hypothyroidism. After induction of primary hypothyroidism, a 37-fold increase in mean basal plasma TSH concentration and a 34-fold increase in mean area under the curve for TSH were found. Analysis by the Pulsar program demonstrated pulsatile secretion of TSH in the hypothyroid state, characterized by relatively low amplitude pulses (mean [+/-SEM]) amplitude 41 +/- 3% of basal plasma TSH level) and a mean pulse frequency of 2.0 +/- 0.5 pulses/6 hr. In the euthyroid state, significant TSH pulses were identified in only 2 dogs. The mean basal plasma TSH level correlated positively (r = 0.84) with the mean amplitude of the TSH pulses, and correlated negatively (r = -0.88) with the TSH pulse frequency. The results of this study demonstrate pulsatile secretion of TSH in dogs during hypothyroidism and only small fluctuations in plasma TSH concentrations during euthyroidism. The findings also suggest that the low TSH values occasionally found in dogs with spontaneous primary hypothyroidism may in some cases in part be the result of ultradian fluctuations. PMID- 10701760 TI - Selenium stimulates estradiol production in bovine granulosa cells: possible involvement of nitric oxide. AB - Reduction in fertility is well known to be possibly related to selenium deficiencies, even if target organ for selenium action is, at present, unclear. The present study was aimed to examine whether selenium directly influences granulosa cells. Bovine granulosa cells from different size follicles were used to investigate the effect of selenium (5 ng/ml), with or without bovine follicle stimulating hormone (bFSH) (100 ng/ml), on proliferation and steroidogenesis. In addition, we sought to determine if selenium modulates the production of nitric oxide, which is known to play an important role in ovarian activity. Our data demonstrate that selenium significantly (P < 0.001) stimulates the proliferation of the cells from small follicles; moreover, it further potentiates the stimulatory effect of the gonadotropin in the same cells. Furthermore, selenium significantly (P < 0.01) augments E2 output by cells from both kinds of follicles. bFSH increases E2 production (P < 0.01) by cells from large follicles, whereas it exerts a stimulatory (P < 0.01) effect only in the presence of selenium in the cells from the small ones. The production of nitric oxide is significantly increased (P < 0.001) by bFSH, but only in cells from small follicles. Selenium inhibits (P < 0.001) nitric oxide production in cells from both kinds of follicles and significantly decreases (P < 0.001) bFSH-induced nitric oxide production in cells from the small ones. We conclude that selenium acts on granulosa cells by modulating their proliferation and E2 synthesis; moreover, its effect could be mediated, at least in part, through an inhibition of nitric oxide. PMID- 10701762 TI - Effects of a simulated estrous cycle on sodium, volume, ACTH, and AVP in sheep. AB - The studies were designed to test for effects of acute increases in estradiol and progesterone, similar in magnitude and duration to those in the ovine estrous cycle, on adrenocorticotropic hormone (ACTH) and plasma vasopressin (AVP) under resting conditions and in response to hypotension. Ewes (7 per group) were studied as intact, ovariectomized, ovariectomized and treated with progesterone for 7-8 days, or subsequently treated with estradiol. During progesterone treatment plasma sodium and AVP were increased significantly. However, neither plasma volume nor blood pressure was altered. Plasma AVP responses to hypotension were not altered by either progesterone or estradiol treatment. The peak plasma ACTH response to hypotension was not altered by steroid treatment; however, the duration of the response was greater in progesterone-treated ewes than in intact ewes. The results indicate that changes in gonadal steroids similar to those in the ovine estrous cycle cause a small increase in plasma sodium that stimulates AVP, but do not alter regulation of blood pressure or volume or AVP or ACTH responses to hypotension. PMID- 10701763 TI - Increase in prolactin receptor (PRL-R) mRNA level in the mammary gland after hormonal induction of lactation in virgin ewes. AB - In order to examine the hormonal regulation of the prolactin-receptor (PRL-R) gene expression during mammary gland development, ewes were treated to induce lactation via an estrogen-progesterone-hydrocortisone and ovine growth hormone treatment. In situ hybridization analysis was used and revealed that sex steroids increased PRL-R mRNA levels in the mammary gland. Using RNase protection assay we showed that the estradiol + progesterone treatment increased both the levels of the long and the short forms of PRL-R mRNA. Addition of hydrocortisone increased the level of alphaS1-casein transcripts and the level of the ratio of the long to the short form of the PRL-R mRNA. This ratio can be further enhanced by addition of ovine growth hormone to the latter treatment. This suggests a role of hydrocortisone and ovine growth hormone in the alternative splicing that leads to the preferential expression of the long form of the PRL-R mRNA. In conclusion, the present experiments suggest that estrogen, progesterone and hydrocortisone are the major regulators of the PRL-R gene expression during pregnancy and prepare the mammary gland for its differentiation. PMID- 10701764 TI - Changes in concentrations of plasma immunoreactive follicle-stimulating hormone, luteinizing hormone, estradiol-17beta, testosterone, progesterone, and inhibin in heifers from birth to puberty. AB - This study was designed to clarify the characteristics of changes in plasma concentrations of reproductive hormones in heifers from birth to puberty. Weekly or daily hormonal changes were observed in 39 heifers. Daily changes in the concentration of follicle-stimulating hormone (FSH) demonstrated a consistent cycle of hormone changes over a 7- to 8-day period in heifers from approximately 10 days to 9 months old. Weekly changes in reproductive hormones showed that there were three brief periods in heifers between birth and puberty in which dramatic changes occur. The first period was the first week after birth, during which a reciprocal relationship between steroid hormones and gonadotropins was observed. At birth, the concentrations of steroid hormones were higher than those at any other age. These hormone levels rapidly decreased within the first week after birth. Gonadotropin levels, however, increased from birth to 1 week of age. The second period of major change was at approximately 4 weeks of age when there was an increase in the concentrations of luteinizing hormone (LH), estradiol 17beta, testosterone, and immunoreactive inhibin. The third period was the last 5 weeks before the first ovulation, when there was an increase in the concentrations of estradiol-17beta followed by an increase in (LH). These results suggest that regular hormone changes start from 10 days after birth and that the periods from birth to 4 weeks of age and the last 5 weeks before the first ovulation in heifers are important to the development of reproductive functions before puberty. PMID- 10701765 TI - Physiological response to acute endotoxemia in swine: effect of genotype on energy metabolites and leptin. AB - Certain high lean gain swine genotypes have greater sensitivity to pathogen and nonpathogen stressors evident by reduced productivity and increased mortality during disease stress or in suboptimal production environments. Saline (control) and an immunologic challenge (LPS; 25 microg lipopolysaccharide/kg BW) were administered to three genetic populations (each pig used as its own control): high lean (H), moderate lean terminal cross (MT), and moderate lean maternal cross (MM). LPS induced anorexia, and significantly increased body temperature and circulating TNF-alpha, cortisol, and NEFA in all genotypes (P < 0.0004). LPS reduced circulating glucose, insulin, and IGF-1 in all genotypes (P < 0.05). The LPS-induced hypoglycemia was significantly greater in MM versus MT and H pigs (P < 0.03). The hypoinsulinemia was significantly greater in MM versus H pigs (P < 0.02). MM pigs recovered from hypoinsulinemia slower than MT pigs (P < 0.03). Control insulin was higher in H versus MT pigs (P < 0.08), but relative to basal, the insulin response to LPS was similar. Plasma haptoglobin response to LPS was lower for MM versus MT and H pigs (P < 0.02), and tended to be lower in MT versus H pigs (P < 0.09). LPS treatment caused similar decreases in plasma IGF-1 concentrations among genotypes. Ten hours after LPS treatment, leptin mRNA abundance in adipose tissue was significantly reduced (relative to control) in MM and H pigs (P < 0.02) but not in MT pigs (P > 0.05). Physiological differences in leptin, a potent regulator of food intake and energy metabolism, may be important factors in the genetic variation in sensitivity to environmental stress. PMID- 10701766 TI - Review: brain aromatization and other factors affecting male reproductive behavior with emphasis on the sexual orientation of rams. PMID- 10701767 TI - Interactions of photoperiod, testosterone, and naloxone on GnRH and LH pulse parameters in the male sheep. AB - This study tested the hypothesis that the effects of the opiate antagonist naloxone on GnRH (and LH) secretion is affected by photoperiod length and testosterone (T) concentrations. The effect of infusing naloxone on GnRH and LH pulse patterns was determined in four groups of orchidectomized sheep: long day (LD) photoperiod treated with T, LD without T (LDC), short day photoperiod (SD) with T, SDC (n = 5-7/group). Hypophyseal-portal and jugular blood samples were collected at 10 min intervals for 4 h before and 4 h during naloxone infusion (1 mg/kg/h). Neither photoperiod nor T affected either mean GnRH or LH whereas naloxone (P < 0.01) increased both. LD photoperiod (P < 0.01), T (P < 0.01) and naloxone (P < 0.01) all increased LH pulse amplitude whereas only naloxone increased GnRH pulse amplitude (P < 0.01). There was an interaction (P < 0.01) between steroid and naloxone on LH, but not GnRH, pulse amplitude. Both LD photoperiod and T increased both LH and GnRH (P < 0.01) interpulse-interval (IPI). Naloxone decreased GnRH IPI (P < 0.01). The LH/GnRH pulse amplitude ratio was (P < 0.02) increased by T--likely a secondary response to the T-induced increase in IPI. These results are interpreted as showing that in the ram the endogenous opiate peptides regulate both GnRH pulse frequency and amplitude, but that their specific role is modulated by photoperiod and T. These results do not support the concept that the opiate peptides are the primary mediators of the negative feedback effects of T. PMID- 10701768 TI - Immunohistochemical analysis of estrogen receptors in feline mammary gland benign and malignant lesions: comparison with biochemical assay. AB - Estrogen receptors (ER) were determined by both the biochemical dextran-coated charcoal (DCC-ER) and the immunohistochemical Avidin biotin-peroxidase complex (IHC-ER) methods in proliferative mammary lesions collected from 37 cats: 20 malignant tumors without metastasis at first presentation, seven malignant tumors with lung and/or lymph node metastases and 10 benign tumors and dysplasias. Total number of samples analyzed by both methods was 44. The DCC-ER method was applied to frozen tissue samples and the IHC-ER method was applied to neutral buffered formalin-fixed, paraffin wax-embedded tissue samples by using the NCL-6F11 monoclonal antibody. Biochemically, 21 (47.7%) cases had equal or more than 5 fmol/mg of protein (standard positivity threshold). Immunohistochemically, 11 (25%) cases were scored positive, the percentage of positive nuclei being statistically linked to the intensity of immunostaining. Normal mammary gland tissue (13 cases) and/or dysplastic areas (5 cases) found in the surroundings of the main lesion were IHC-ER positive in 76.9% and 40% of the cases, respectively. Concordance between DCC-ER and IHC-DCC was 72.7% and the results of the DCC and the IHC-ER methods were significantly correlated (P < 0.05) by the chi2 test. Specificity (true negatives) and sensitivity (true positives) of the ICH-ER method were 95.6% and 47.6%, respectively. One out of eleven DCC-ER positive and IHC-ER negative discordant cases (9.09%) was a DCC-ER false positive, because the surrounding normal mammary gland tissue was IHC-ER positive. The remaining 10 cases had ER content values equal or lower than 23 fmol/mg of protein, a figure that could represent the sensitivity threshold of the immunohistochemical method employed. PMID- 10701769 TI - Oxidized-low density lipoprotein inhibits cyclic AMP production by porcine luteal cells. AB - Oxidized(OX)-low density lipoprotein (LDL) inhibits steroidogenesis by luteal cells (LC) from regressing porcine CL. The present study was designed to investigate the mechanism of inhibition by determining whether OX-LDL inhibits basal and agonist-stimulated cAMP production in regressing LC. Collagenase dispersed porcine LC (n = 7 animals, estrous cycle Day 12-15) were cultured (2.5 x 10(5) cells/0.5 ml) in serum-free DMEM/Hams F-12 in duplicate wells at 37 degrees C. Approximately 18 hr after plating, media were replaced and LC were immediately treated with human LDL (0, 25, or 100 microg/ml) or OX-LDL (25 or 100 microg/ml). LC were incubated for 2 hr before addition of isobutylmethylxanthine (IBMX) to inhibit phosphodiesterase activity, immediately followed by hCG (100 ng/ml), cholera toxin (CT; 0.1 microM), forskolin (FS; 50 microM), or no further treatment (controls). LC were incubated for an additional 90 min. After removal of culture media, cells were extracted with 0.1 N HCl. Cell extracts were assayed for cAMP by enzyme immunoassay (EIA). HCG, CT, and FS increased (P < 0.05) cAMP production approximately four-, 10-, and 25-fold, respectively, relative to controls. OX-LDL (25 and 100 microg/ml) inhibited (P < 0.05) cAMP production by unstimulated, hCG-, and CT-stimulated LC, but not that by FS-stimulated LC. The highest concentration of OX-LDL (100 microg/ml) reduced cAMP formation by 39.8 +/ 6.6%, 44.7 +/- 10.5%, and 67.7 +/- 4.5% in unstimulated, hCG-, and CT-stimulated LC, respectively. In contrast, unmodified LDL (25 and 100 microg/ml) did not alter cAMP production. We conclude that OX-LDL can interfere with the cAMP signaling pathway in regressing luteal cells by acting at sites proximal to adenylate cyclase activation. PMID- 10701770 TI - Effect of endotoxin challenge on hepatic 5'-deiodinase activity in cattle. AB - Thyroid status is compromised in a variety of acute and chronic infections and toxin-mediated disease states. Conversion of thyroxine (T4) into the metabolically active hormone, triiodothyronine (T3), is catalyzed by 5' deiodinase (5'D). Our objective was to determine the effect of endotoxin (LPS) challenge with and without L-arginine (Arg) infusion on hepatic activity of 5'D and plasma concentrations of T4 and T3. In a 2 x 2 factorial, beef heifers (275 310 kg b.wt.) were fed low (8% CP; 6.5 kg/d) or high (14% CP; 7.2 kg/d) isocaloric protein diets (1.96 Mcal/kg DM) for 10 d before LPS challenge. L Arginine in saline (0.5 g/kg b.wt.) or saline alone was infused i.v. throughout an 8 hr period starting 2 hr before bolus LPS injection (Escherichia coli, 055: B5; 0.2 microg/kg; i.v.). Blood samples were collected at -2, 0, 3, 6, 12, and 24 hr relative to LPS injection. Liver samples were obtained 20 hr before, and then 6 and 24 hr after LPS challenge using a biopsy needle. Plasma T4 and T3 concentrations were not affected by dietary CP or Arg. Compared with levels at 0 hr, LPS challenge decreased plasma T4 (P < 0.01) and T3 (P < 0.001), respectively, 8.4% and 28.9% at 6 hr and 19.7% and 31.3% at 24 hr. Consistent with these changes, the T3:T4 ratio was lower than that at 0 hr (P < 0.001) 22.0% at 6 hr and 13.5% at 24 hr. Hepatic 5'D activities 20 hr before LPS injection were 2.80 +/- 0.11 nmol I- x hr(-1) x mg protein(-1) and decreased 24 hr after LPS, respectively, 45.4% (P < 0.01) and 17.6% (P < 0.05) in saline- and Arg infused heifers. The results indicate that mild LPS challenge in cattle inhibits hepatic generation of T3 and decreases plasma concentrations of thyroid hormones. The data also suggest that the impact of LPS on 5'D activity in liver can be altered by Arg supplementation. PMID- 10701772 TI - Nutritional and somatotropin regulation of the mitogenic response of mammary cells to mammary tissue extracts. AB - Our objective was to investigate the mitogenic response of primary mammary epithelial cells to extracts of mammary parenchyma from 24 prepubertal Friesian heifers treated with placebo or growth hormone at either a low or a high feeding level. The mitogenic responses to mammary extracts were tested by using primary mammary epithelial organoids obtained from prepubertal heifers cultured for 4 to 5 d in collagen gels in serum-free medium supplemented to 5% concentration of the mammary extracts. Cell proliferation was determined using [methyl-3H]thymidine incorporation as a measure of DNA synthesis. High feeding level reduced DNA synthesis in response to mammary extracts. At low feeding level, growth hormone treatment decreased DNA synthesis in response to mammary extracts whereas, at high feeding level, growth hormone increased DNA synthesis in response to mammary extracts. These results suggest that locally produced growth factors are involved in the regulation of mammary development when mammary growth is modulated by feeding level and growth hormone treatment. PMID- 10701771 TI - Growth hormone treatment of breeding bulls used for artificial insemination improves fertilization rates. AB - To evaluate new therapeutical concepts for male subfertility, we tested the effects of exogenous recombinant bovine growth hormone (rbGH) on various endocrine and metabolic parameters both in blood and in seminal plasma of bulls. Sperm quality was assessed morphometrically and by monitoring the number of successful artificial inseminations (AIs) defined as non-return rates (NRR). Aliquots of 450 semen samples were used from each bull and each experimental period (4 wk before, 14 weeks during and 6 wk after treatment). Six out of ten sires (average age 8.4 years) were treated every two weeks with 640-mg depot formulated rbGH (Eli Lilly). Four bulls received vehicle only. Blood plasma bGH, IGF-I, insulin and glucose concentrations were increased with rbGH treatment. In seminal plasma there was no effect of rbGH treatment on fructose and citrate or on testosterone concentrations. With one exception, rbGH-treated bulls had greater IGFBP-3 concentrations in seminal plasma. Motility of spermatozoa after freezing and thawing was increased compared with pretreatment rates. Most interestingly, the number of successful AIs was increased by an average of 6.0% NRR when ejaculates from rbGH-treated bulls were used. PMID- 10701773 TI - Lack of collagen type specificity for lysyl hydroxylase isoforms. AB - Lysyl hydroxylase is the enzyme catalyzing the formation of hydroxylysyl residues in collagens. Large differences in the extent of hydroxylysyl residues are found among collagen types. Three lysyl hydroxylase isoenzymes (LH1, LH2, LH3) have recently been characterized from human and mouse tissues. Nothing is known about the distribution of these isoforms within cells or whether they exhibit collagen type specificity. We measured mRNA levels of the three isoforms, as well as the mRNAs of the main collagen types I, III, IV, and V and the alpha subunit of prolyl 4-hydroxylase, another enzyme involved in collagen biosynthesis, in different human cell lines. Large variations were found in mRNA expression of LH1 and LH2 but not LH3. Immunoblotting was utilized to confirm the results of Northern hybridization. The levels of mRNA of LH1, LH2, and the alpha subunit of prolyl 4-hydroxylase showed significant correlations with each other. The LH3 mRNA levels did not correlate with those of LH1, LH2, or the alpa subunit of prolyl 4-hydroxylase, clearly indicating a difference in the regulation of LH3. No correlation was observed between LH isoforms and individual collagen types, indicating a lack of collagen type specificity for lysyl hydroxylase isoforms. Our observations suggest that LH1, LH2, and the alpha subunit of prolyl 4 hydroxylase are coregulated together with total collagen synthesis but not with the specific collagen types and indicate that LH3 behaves differently from LH1 and LH2, implying a difference in their substrates. These observations set the basis for further studies to define the functions of lysyl hydroxylase isoforms. PMID- 10701774 TI - Transcription of human cathepsin B is mediated by Sp1 and Ets family factors in glioma. AB - Cathepsin B expression is increased at both the mRNA and protein levels in a wide variety of tumors. The mechanisms responsible for this regulation are not well elucidated. We have isolated a 2.2-kb cathepsin B genomic fragment that contains the 5'-flanking region of the cathepsin B gene. Using reporter gene analysis in human glioblastoma U87MG cells, we have mapped a 228-bp fragment (-172 to +56) having high promoter activity. This promoter region has a high G+C content; contains potential Spl, Ets, and USF binding motifs; and lacks canonical TATA and CAAT boxes immediately upstream of the major transcriptional initiation site. Cotransfection experiments demonstrated that Spl and Ets1 could trans-activate cathepsin B transcription, whereas Ets2 could not. Electrophoretic mobility shift assays and supershift assays revealed that three of the four putative Sp1 sites in this promoter region form a specific complex containing the Sp1 transcription factor. Mutating all four of the Spl binding sites individually markedly reduced the promoter activity of transfected reporter genes in U87 cells. Cotransfection of this cathepsin B promoter construct with Spl family expression vectors in Schneider's Drosophila line 2 (SL2) cells demonstrated that Spl and Sp3, but not Sp4, activated cathepsin B transcription. Taken together, these results suggest that Sp1, Sp3, and Ets1 are important factors in cathepsin B transcription. The regulation of cathepsin B transcription by Sp1- and Sp1-related factors is mediated through multiple GC boxes. PMID- 10701775 TI - Role for C-tail residues in delta opioid receptor downregulation. AB - The delta opioid receptor, a member of the G-protein-coupled receptor superfamily, was used as a model system to characterize opioid receptor downregulation. Metabolic labeling followed by immunoprecipitation resulted in the isolation of the epitope-tagged mouse delta opioid receptor as a approximately 60-kDa protein. Prolonged agonist treatment with 100 nM d-Ala2, d Leu5-enkephalin (DADLE) caused significant (approximately 60%) reduction in the level of receptor. The delta opioid receptor contains a number of phosphorylatable residues in the C tail. Point mutations of the majority of Ser/Thr sequences did not affect the level of downregulation, whereas mutation of Thr353 to Ala did. In order to test if phosphorylation at this site is involved in receptor downregulation, we generated a Thr353Glu mutant that would mimic the phosphorylated Thr at this site. This mutant exhibited a significantly higher extent of downregulation than the Thr353Ala mutant. In order to critically evaluate the requirement of Thr353 in receptor downregulation, we examined the downregulation of wildtype rat delta receptor (which does not contain Ala353) and an Ala353Thr point-mutant rat delta receptor. The wild-type receptor exhibited poor agonist-mediated downregulation, whereas Ala353Thr mutant exhibited increased downregulation. These results and results from additional studies with rat/mouse chimeric receptors support a role for phosphorylation of sites within the C tail in efficient downregulation of delta opioid receptors. PMID- 10701776 TI - Estrogen modulates HNF-3beta mRNA levels in the developing chick oviduct. AB - Steroid hormones are involved in many physiological processes, including tissue specific gene expression, homeostasis, and development. The chick oviduct represents an excellent system in which to study many of these events, as it is highly steroid responsive. Here, we report the cloning of chick HNF-3beta from an oviduct cDNA library and its expression pattern in adult tissues and in the developing oviduct in response to estrogen treatment. Overall, cHNF-3beta was expressed at high levels in the immature chick oviduct and lung and, to a lesser extent, in the liver, kidney, and muscle. This expression pattern is divergent from that of mammalian HNF-3beta, which is not expressed in kidney or muscle. Furthermore, several lengths of cHNF-3beta mRNA transcripts were detected that were expressed tissue specifically. Interestingly, cHNF-3beta mRNA levels were differentially influenced by estrogen as a result of a post-transcriptional effect on the cHNF-3beta message in some tissues. Finally, a role for cHNF-3beta is proposed in the estrogen-stimulated differentiation and development of the oviduct, as cHNF-3beta mRNA expression is induced in the early stages of oviduct development and declines as the animal becomes sexually mature. PMID- 10701777 TI - Expression of the hydrogen peroxide-generating enzyme fatty acyl CoA oxidase activates NF-kappaB. AB - Peroxisome proliferators are a class of hepatic carcinogens in rodents and have been proposed to act in part by increasing oxidative stress. Fatty acyl CoA oxidase (FAO), which is highly induced by peroxisome proliferators, is the hydrogen peroxide-generating enzyme of the peroxisomal beta-oxidation pathway. We previously showed that the treatment of rats and mice with the peroxisome proliferator ciprofibrate resulted in increased hepatic NF-kappaB activity and suggested that this effect may be secondary to the action of H2O-generating enzymes. To test this possibility directly, we have determined whether transient overexpression of FAO, in the absence of peroxisome proliferators, leads to NF kappaB activation. Here, we show that FAO overexpression in Cos-1 cells, in the presence of an H2O-generating substrate, can activate a NF-kappaB regulated reporter gene. Electrophoretic mobility shift assays further demonstrated that FAO expression increases nuclear NF-kappaB DNA binding activity in a dose dependent manner. The antioxidants vitamin E and catalase can inhibit this activation. These results indicate that FAO mediates, at least in part, peroxisome proliferator-induced NF-kappaB activation. PMID- 10701778 TI - Structure and expression of the mouse gene encoding the orphan nuclear receptor TEC. AB - Translocated in extraskeletal chondrosarcoma (TEC) is an orphan nuclear receptor involved in the control of cell proliferation and apoptosis and is expressed mainly in the mammalian central nervous system. To help understand the regulation of its expression, we have characterized the mouse genomic locus encoding TEC and analyzed its expression pattern in various tissues. The gene spans approximately 40 kb and contains 8 exons, of which the first two are noncoding. The promoter region does not contain any identifiable TATA box or CCAAT box elements; however, several binding sites for the transcription factors cyclic AMP-responsive element binding (CREB) protein and Spl are present. Two types of transcripts generated by alternative splicing were characterized by RT-PCR: one encodes the full-length receptor of 627 amino acids; the other encodes a truncated receptor of 429 amino acids lacking the entire carboxyl-terminal domain. Northern blots and RT-PCR analyses showed that mRNAs encoding both isoforms are expressed in all mouse tissues examined, with the highest levels being found in the brain. This expression pattern suggests that TEC may perform some basic housekeeping cellular function in addition to its role in cell proliferation. PMID- 10701779 TI - Molecular characterization of TgHBox4, a Drosophila Abd-B homolog found in the sea urchin Tripneustes gratilla. AB - We have isolated and sequenced a cDNA clone that, as judged by the sequence of the homeobox region, encodes a sea urchin homolog of the homeobox containing the gene Abdominal-B of Drosophila. The total length of the cDNA is 3634 nucleotides and includes an open reading frame, which encodes a protein that is 32,321 Da. The N-terminal region of the homeodomain includes consensus sequences found in some of TgHBox4's Abdominal-B relatives. A genomic clone representing the 5' part of the message was also isolated. This clone and a previously isolated clone were found to represent the full-length cDNA sequence. We have also raised antibodies against a bacterially expressed portion of the TgHBox4 protein and used them to determine the location of TgHBox4 proteins during development. The protein displays ubiquitous expression early in development but becomes more restricted, to posterior regions, late in embryogenesis. Thus, in contrast to its Abd-B homologs in bilateral metazoans, TgHBox4 is probably not involved in pattern formation but may have a posterior-defining role late in embryogenesis. PMID- 10701780 TI - How common are various causes of dizziness? A critical review. AB - BACKGROUND: Although dizziness is a common symptom in both primary care and referral practices, the relative frequency of various causes has not been well delineated. METHODS: A MEDLINE search identified 12 articles containing original data on the etiology of dizziness in consecutive patients. Study sites included primary care offices (n = 2), emergency room (n = 4), and referral clinics (n = 6). Each study's strength of design was graded using nine quality criteria. RESULTS: Dizziness was attributed to a peripheral vestibulopathy in 44% of patients, a central vestibulopathy in 11%, psychiatric causes in 16%, other conditions in 26%, and an unknown cause in 13%. Certain serious causes were relatively uncommon, including cerebrovascular disease (6%), cardiac arrhythmia (1.5%), and brain tumor (<1%). CONCLUSIONS: Dizziness is due to vestibular or psychiatric causes in more than 70% of cases. Since serious treatable causes appear uncommon, diagnostic testing can probably be reserved for a small subset of patients. PMID- 10701781 TI - The major histocompatibility complex and inflammation. AB - The major histocompatibility complex (MHC) is of major medical interest because of its contribution to transplant rejection and to variation among individuals in susceptibility to a variety of autoimmune disorders. In addition to its role in influencing the propensity for known autoimmune diseases, the MHC contains genes contributing to several other hereditary disorders that are not autoimmune in nature or in which the role of autoimmunity is uncertain. Recently, a cluster of genes encoding inflammation-related proteins were found, and our review focuses on these findings and their clinical relevance. PMID- 10701782 TI - Blunt thoracic aortic injuries: initial evaluation and management. AB - In at least one large study, the average time from arrival at the emergency department to arrival in the operating room was nearly 6 hours. That 30% of survivors will die in the same amount of time underscores the need for rapid diagnosis and treatment. In blunt thoracic aortic injury, beta-blockers have been shown to reduce the incidence of rupture, and their use is rarely contraindicated. A working knowledge of the mechanisms of injury likely to produce this lesion, commonly associated injuries, clinically relevant and easily recognizable chest film findings, and appropriate use of beta-blockade can have a significant impact on mortality. Any physician responsible for evaluation of trauma patients should be familiar with this information. PMID- 10701783 TI - Domestic violence in a university emergency department. AB - BACKGROUND: We attempted to determine the prevalence and demographics of domestic violence (DV) among male and female patients in a university emergency department (ED). METHODS: The validated Index of Spouse Abuse (ISA) was used. Patients aged 18 years or older seen during 28 randomly selected 4-hour shifts were eligible. RESULTS: Of the 97 participants in the study, 57 were female. One man and 3 women were victims of present physical DV, with 1 male and 2 female victims of present nonphysical abuse. Three of the 40 men and 22 of the 57 women had been victims of past physical violence. One man and 15 women had been victims of past nonphysical abuse. Alcohol use, suicidal ideation, family history, and psychiatric history were all strongly correlated with DV. CONCLUSIONS: The prevalence of DV past was significantly higher in the females. Present violence was more rare and less than that reported in other ED studies. PMID- 10701784 TI - Impact of a formal trauma program on a small rural hospital in Mississippi. AB - BACKGROUND: Since Mississippi is largely rural, most of the initial care given to trauma patients is at small community hospitals. This study examined the impact of the institution of a formal trauma program on trauma care at such a hospital. METHODS: All trauma cases at a single institution during 1998 were retrospectively analyzed. Cases were stratified into two groups, which depended on whether trauma care was given before or after institution of the trauma program. Various parameters were evaluated and compared. RESULTS: The disposition of trauma cases changed after institution of a formal protocol: both transfers to trauma centers and hospital admissions decreased, while the number of cases cleared in the emergency department (ED) increased. The percentage of cases inappropriately managed also decreased. CONCLUSIONS: Institution of a formal trauma program increased the efficiency of resource utilization and improved the level of care received by trauma patients. PMID- 10701785 TI - Drug use and HIV risk--related sex behaviors: a street outreach study of black adults. AB - BACKGROUND: Our street outreach project investigated the relationship between use of noninjecting drugs (alcohol, marijuana, cocaine) and human immunodeficiency virus (HIV) risk-related sex behaviors of black adults. The study focused on three HIV-related risks: multiple sex partners, unprotected sex, and drugs during sex. METHODS: Data for this study were collected in a street outreach community survey for a drug abuse and HIV intervention study in Birmingham, Ala. A total of 780 black men and women completed the survey. RESULTS: High-risk sex behaviors were far more prevalent among cocaine users than marijuana or alcohol users. A greater number of cocaine users reported having multiple sex partners, not using condoms, and using drugs during sex. Female cocaine users showed the same risk level for HIV infection as male cocaine users. CONCLUSIONS: Increased risk of HIV infection through sexual transmission is associated with use of noninjecting cocaine for both men and women. Condom use should be considered as a major component of HIV prevention programs. PMID- 10701786 TI - The cost of running American city hospitals: the Gorgas 1910 survey. AB - BACKGROUND: COL William C. Gorgas was appointed Chief Sanitary Officer of the Isthmian Canal Commission during construction of the Panama Canal (1904-1914). In 1910, Gorgas sought to determine the administrative and operating costs of major metropolitan hospitals in the United States and compare these with similar costs in the Canal Zone hospitals. METHODS: Gorgas sent a questionnaire to hospitals in Atlanta, Baltimore, Boston, Chicago, Cleveland, New York, Philadelphia, Pittsburgh, San Francisco, and Washington, DC. The information requested included number of beds, daily census, details about resident and nursing staff, salaries, length of stay, and hospital cost per patient per day. RESULTS: The survey results provide information about metropolitan hospitals in the United States at the turn of the century. Hospital costs varied from $.22 to $2.76 per patient per day. CONCLUSION: Gorgas concluded that the costs of operating hospitals in the Canal Zone compared favorably with those in the United States. PMID- 10701787 TI - Aortopulmonary fistula. AB - BACKGROUND: Aortopulmonary fistula is an uncommon but usually fatal condition if not treated surgically. The most frequent cause is erosion of a false aneurysm of the descending thoracic aorta into the left lung. METHODS: Review of charts of all patients who had had resection of a thoracic aortic aneurysm at the MidAmerica Heart Institute (1971 to 1997) revealed three cases in which the presentation was hemoptysis resulting from an aortopulmonary fistula. The clinical features and course of each patient are summarized in this report. RESULTS: The three patients with hemoptysis due to an aortopulmonary fistula had emergency surgical intervention with no major complication. CONCLUSIONS: Any patient who has an otherwise unexplained hemoptysis and a history of a previous thoracic aortic surgical procedure or is known to have a thoracic aortic aneurysm should have appropriate clinical evaluation to exclude the presence of an aortopulmonary fistula. If an aortopulmonary fistula cannot be excluded, emergency operation should be done. PMID- 10701788 TI - Comparing the toxicity of digoxin and digitoxin in a geriatric population: should an old drug be rediscovered? AB - BACKGROUND: Little information is available regarding toxicity rates of the two available forms of cardiac glycosides (digoxin, digitoxin) when used in elderly patients. METHODS: We retrospectively analyzed the charts of all patients more than 60 years of age who were chronically managed with a cardiac glycoside and were hospitalized during the period January 1995 through January 1998. Toxicity was defined as any clinical event that required either a reduction in dose of the drug or its discontinuance. RESULTS: Toxicity occurred among 7.6% of hospitalizations in which digitoxin was used, compared with 18.3% of hospitalizations in which digoxin was used. In multivariate analysis, the odds of toxicity adjusted for other clinical characteristics were three times greater for patients taking digoxin than for patients taking digitoxin. CONCLUSION: Hospitalized elderly patients taking digitoxin had a lower rate of toxicity than those taking digoxin. PMID- 10701789 TI - Accuracy of clinical evaluation in the determination of brain death. AB - BACKGROUND: The accuracy of the clinical diagnosis of brain death has never been established. METHODS: Seventy-one consecutive clinically brain dead patients were studied retrospectively. Inclusion criteria were complete cessation of brain function with profound coma of known cause, complete absence of brain stem reflexes, and apnea, all persisting for a least 6 hours. A formal apnea test with a documented Pco2 of > 60 mm Hg was required. All evaluations were done by experienced neurosurgery or neurology resident or staff physicians. The clinical diagnosis was compared with the results of radionuclide angiography and with the clinical course and final outcome. RESULTS: Seventy patients had no arterial blood flow on radionuclide angiography. One blood flow study was considered to have yielded a false-negative result. No patient recovered or survived. CONCLUSIONS: The clinical diagnosis of brain death is highly reliable when made by experienced examiners using established criteria. In this study, the accuracy was 100%. PMID- 10701791 TI - Acute mesenteric ischemia and malpractice claims. AB - BACKGROUND: Acute mesenteric ischemia can be a difficult diagnosis to make, but delay contributes directly to infarction, and this may provide a setting for malpractice claims. METHODS: We reviewed 180 consecutive malpractice claims submitted by attorneys for medical expert (ME) review during the 12 years ending in late 1998. Seven cases involved acute mesenteric ischemia. RESULTS: Alleged failure to make a timely diagnosis was the basis for 5 of these claims, failure to provide anticoagulant protection for 1, and failure to prevent nonocclusive ischemic infarction for 1. Six claims were closed after ME review and 1 claim involving late diagnosis was settled before trial. CONCLUSIONS: The risk of a malpractice claim is reduced by consideration of computed tomography (CT), angiography, and surgical consultation as soon as a patient is seen whose differential diagnosis includes acute mesenteric ischemia. PMID- 10701790 TI - Anserine bursitis in patients with osteoarthritis of the knee. AB - BACKGROUND: We sought to determine the frequency of anserine bursitis (AB) in Koreans with osteoarthritis (OA) of the knee and its relationship to age, sex, and radiographic severity of OA, and to evaluate its response to various forms of therapy. METHODS: In a retrospective study, we reviewed the charts of patients with OA of the knee and graded the radiographic severity by the Kellgren-Lawrence grading scheme. RESULTS: Of 62 patients, 29 had AB. No difference in age, sex, and radiographic severity was noticed between patients with AB and those without AB. Eleven of 12 patients who received a local injection of methylprednisolone plus lidocaine at the anserine bursa had relief, whereas 7 of 17 patients who received noninjection therapy for AB showed improvement. CONCLUSIONS: Anserine bursitis is commonly found in Koreans with OA of the knee, and its presence is unrelated to age, sex, and radiographic severity. Local injection at the anserine bursa is more effective than noninjection therapy. PMID- 10701792 TI - Use of technetium Tc 99m sestamibi scintigraphy for recurrent tertiary hyperparathyroidism from a parathyroid forearm graft. AB - Technetium Tc 99m sestamibi scintigraphy is a sensitive technique for localizing recurrent parathyroid disease in the neck or mediastinum. We report the case of a 60-year-old woman with recurrent tertiary hyperparathyroidism after total parathyroidectomy. Technetium Tc 99m sestamibi images of the neck and mediastinum were negative; however, images of the right arm revealed a hyperfunctioning parathyroid autotransplant. Partial resection of the autograft resulted in prompt resolution of the hyperparathyroidism. PMID- 10701793 TI - Typhlitis as a presenting manifestation of acute myelogenous leukemia. AB - We describe a case of typhlitis with onset before diagnosis of acute myelogenous leukemia or initiation of chemotherapy. Typhlitis is usually seen as a complication of chemotherapy for hematologic malignancies. It is being reported with increasing frequency in association with other disease states that produce profound neutropenia. PMID- 10701794 TI - Pancreatic pseudocyst masquerading as an incarcerated inguinal hernia. AB - We present a case in which a pancreatic pseudocyst dissected into the right inguinal region, thus masquerading as an incarcerated right inguinal hernia. Computed tomography (CT) confirmed the diagnosis of a pseudocyst, and the patient was treated successfully with percutaneous drainage. PMID- 10701795 TI - Concomitant acute sinusitis and acute lung rejection. AB - Two lung transplant recipients had concomitant acute sinusitis and acute lung rejection. Antibiotics and decongestants alleviated the sinusitis, but the symptoms of cough and dyspnea as well as spirometric defects necessitated treatment of acute lung rejection. In patients with clinical evidence of acute sinusitis after lung transplantation, concomitant acute lung rejection should be suspected if dyspnea or pulmonary dysfunction is also present. This appears to be the first report of concomitant acute sinusitis and acute lung rejection. PMID- 10701797 TI - Acquired segmental megacolon in an adult patient with cystic fibrosis. AB - Cystic fibrosis (CF) is characterized by symptoms related to pulmonary dysfunction and pancreatic insufficiency. Constipation, though a frequent complaint of patients with CF, receives less attention. We report a case of acquired segmental megacolon and constipation necessitating surgical colonic resection in an adult patient with CF. The differential diagnosis and possible causes of megacolon in this setting are discussed. PMID- 10701796 TI - Carcinosarcoma of the uterus associated with a nongestational choriocarcinoma. AB - Choriocarcinoma has been reported in association with endometrial carcinoma and as a metaplastic change in multiple carcinomas, including liver, urinary bladder, lung, and the gastrointestinal tract. We report choriocarcinoma in conjunction with a carcinosarcoma (also called malignant mullerian mixed tumor) in a 71-year old woman whose hysterectomy specimen revealed two polypoid lesions of the endometrium, one arising from the anterior endometrium and one arising from the posterior endometrium. Histologic examination revealed three histologic patterns. The anterior endometrial lesion showed a FIGO grade 2 endometrioid endometrial adenocarcinoma. The posterior endometrial lesion showed a carcinosarcoma composed of a high-grade adenocarcinoma and scant homologous stromal sarcoma. In addition, a choriocarcinoma was identified intermixed with the adenocarcinoma. The syncytiocytotrophoblasts and cytotrophoblasts stained strongly with 0 human chorionic gonadotropin (beta-hCG) and human placental lactogen (hPL). The patient's beta-hCG levels on postoperative days 14, 27, and 42 were 283, 32, and 7 mIU/mL, respectively. This unusual case suggests the importance of identifying the choriocarcinomatous component, since the serum beta-hCG can serve as a marker of tumor recurrence postoperatively. PMID- 10701798 TI - Polymicrobial cholangitis and liver abscess in a patient with the acquired immunodeficiency syndrome. AB - Cholangitis/cholangiopathy associated with the human immunodeficiency virus (HIV) infection is characterized by chronic abdominal pain, low-grade fever, cholestasis, and sometimes areas of focal or diffuse dilatation of the bile ducts that may be apparent on noninvasive imaging studies. Although the etiology of this biliary disease may be multifactorial, it appears to be the result of immunosuppression and/or secondary opportunistic infections rather than a direct cytopathic effect of HIV itself. Various opportunistic pathogens, including cytomegalovirus, Cryptosporidium, Campylobacter fetus, and Candida albicans, have been implicated as causes of HIV-associated cholangitis. We report an unusual case of polymicrobial cholangitis and liver abscess in a patient with HIV infection. PMID- 10701800 TI - Interstitial cystitis and the potential role of gabapentin. AB - Gabapentin, an antiepileptic agent, is a safe and versatile medication also used in the adjunctive treatment of painful disorders. These include neuropathic pain, such as postherpetic neuralgia, diabetic neuropathy, and the pain of reflex sympathetic dystrophy. Interstitial cystitis, a painful disease entity, shares many common features of these chronic pain states, and the use of gabapentin can assist in pain control. Gabapentin, as an adjunctive agent, may reduce use of cotherapeutics such as narcotics. Two patients with interstitial cystitis improved functional capacity within their activities of daily living and received adequate pain control with the addition of gabapentin to their medication regimen. PMID- 10701799 TI - Massive upper gastrointestinal hemorrhage due to cytomegalovirus infection in two patients with acquired immunodeficiency syndrome. AB - Massive upper gastrointestinal (GI) hemorrhage is a rare manifestation of GI cytomegalovirus (CMV) infection. A review of the English language literature yielded 21 well-documented cases of gastric ulcers due to CMV, and 7 of these 21 cases were complicated by significant GI bleeding. This report describes two cases of massive upper GI hemorrhage due to CMV infection in patients with acquired immunodeficiency syndrome (AIDS). PMID- 10701801 TI - Scoliosis associated with typical Mayer-Rokitansky-Kuster-Hauser syndrome. AB - Disorders that cause congenital scoliosis include Mayer-Rokitansky-Kuster-Hauser (MRKH) syndrome. We present the case of a 46-year-old karyotypical (XX) woman with mullerian agenesis (MRKH type A, typical form), a rudimentary bicornate uterus, a blind vaginal pouch, and adenocarcinoma of both ovaries with subsequent bilateral salpingo-oophorectomy. She also had scoliosis of the thoracic and lumbar spine, an association thus far seen only among patients with type B (atypical) MRKH. We describe typical and atypical forms of MRKH and emphasize how these various anomalies associated with mullerian agenesis have affected the classification of the syndrome. We also outline possible embryologic etiologies of mullerian agenesis. PMID- 10701802 TI - Acute compartment syndrome after rupture of the medial head of the gastrocnemius muscle. AB - Rupture of the gastrocnemius muscle is an uncommon injury, with most cases occurring in athletically active individuals. The presentation of a gastrocnemius rupture is the acute onset of calf pain and subsequent ecchymosis. Most of these injuries can be treated symptomatically with good results. We present an unusual case of gastrocnemius muscle tear complicated by acute compartment syndrome. Physicians need to be aware of this potentially devastating complication of gastrocnemius rupture. PMID- 10701803 TI - The tilted disc syndrome. PMID- 10701804 TI - Epithelial inclusions in association with mucin ball development in high-oxygen permeability hydrogel lenses. AB - Debris trapped behind contact lenses may be associated with adverse reactions in extended wear. Although streaks and clumps of cellular material have been reported after overnight wear of conventional materials, recent experience with several high-oxygen permeability (Dk), silicone-containing hydrogel lenses indicates that certain participants are prone to the development of a unique back surface debris. This debris forms as spherical, translucent entities and results in depressions within the ocular surface after lens removal. Little information is known about these spherical bodies, particularly with respect to their composition and development. In this report, we provide photographic evidence of this debris (sometimes termed "mucin balls" or "lipid plugs"), discuss its differential diagnosis and describe a case in which material seems to be embedded in the epithelium as a direct consequence of their presence. PMID- 10701805 TI - Contrast is enhanced by yellow lenses because of selective reduction of short wavelength light. AB - PURPOSE: Although many studies have shown a subjective preference for yellow lenses, there has been little success in determining the clinical nature of this benefit. METHOD: Contrast sensitivity, color vision, accommodative-convergence, and visual acuity were measured in a group of 20 young subjects along with subjective rating of their perception through clear control lenses (380-nm cut off), yellow lenses (450-nm cut-off), dark yellow lenses (511-nm cut-off), and orange lenses (527-nm cut-off). RESULTS: A systematic detriment to color vision was found to occur with increasing cut-off wavelength of the yellow lenses (p < 0.001) and this was significantly correlated to subjective ratings of color (r = 0.66) and brightness (r = -0.34). Perceived brightness significantly improved for the yellow (450-nm cut-off) lens only (p < 0.001). Although tinted lenses reduced contrast sensitivity to a white on black grating, there was a significant improvement in low to midrange spatial frequencies when measured using a white-on blue grating. CONCLUSIONS: The detriment in color vision caused by yellow-colored lenses enhances contrast when viewing bright objects against a blue-based background, such as the sky. Contrast of overlying objects is enhanced is due to the selective reduction of short-wavelength light by the yellow lenses. PMID- 10701806 TI - Stimulus motion increases measured visual field extent in children 3.5 to 30 months of age. AB - PURPOSE: To examine the influence of stimulus motion on measured visual field extent of 3.5- to 30-month-old children and adults. METHODS: Each subject was tested with LED-hybrid and LED-kinetic perimetry procedures, using a black double arc perimeter. Targets in both procedures were identical in size, color, luminance, contrast, and flicker rate. However, in the LED-hybrid procedure, peripheral targets were sequentially illuminated from more peripheral to more central locations, whereas in the LED-kinetic procedure, a peripheral target on a black wand was manually moved centrally along the perimeter arm. A subset of subjects was also tested with white sphere kinetic perimetry (WSKP). RESULTS: The LED-kinetic procedure produced larger measured visual field extent than the LED hybrid procedure in 3.5-, 11-, 17-, and 30-month-olds, but not in 7-month-olds or adults. Data from subjects tested with WSKP indicated that both stimulus motion and discrepancies in scoring methods contributed to the difference reported previously between visual field measurements obtained with WSKP vs. LED-hybrid perimetry. CONCLUSION: In infants and toddlers, measured visual field extent is larger for moving than for nonmoving targets. Further research is needed to determine whether the effect of motion is related to the visual system or to attentional factors. PMID- 10701807 TI - Retest variability of human infant contrast sensitivity: how many tests are sufficient? AB - Retest variability of a new infant contrast sensitivity (CS) card procedure was assessed by binocular measurement of a group of 20 6-month-olds twice within a 1 week period. Coefficient of reliability analyses showed that within-subject variability between tests was only slightly less than variation across subjects, which suggests that results from a single test are a poor predictor of an infant's "true" visual functioning. To determine how many tests are needed to estimate when infant CS stabilizes to within an acceptable (0.15 log unit) criterion, a second experiment was conducted in which a small group of subjects was tested repeatedly over a 2-week period. The results showed that averaging performance on 2 to 3 tests was required before an accurate estimate of the subject's performance could be obtained. Our results suggest that caution should be taken in the interpretation of a single measurement of infant visual functioning. PMID- 10701808 TI - The negative directional aftereffect associated with adaptation to the prismatic effects of spectacle lenses. AB - PURPOSE: To determine whether subjects would demonstrate a negative directional aftereffect in a sparse visual environment after being trained to point accurately through a spectacle lens. METHODS: Subjects were made myopic using a contact lens and then the myopia was corrected with a spectacle lens. Pointing behavior was used to assess directional localization. Training was carried out by showing subjects their pointing errors. RESULTS: Seven of 10 subjects demonstrated a negative directional aftereffect after spectacle lens adaptation. CONCLUSIONS: The presence of a negative directional aftereffect indicates that some patients who switch between spectacles and contact lenses can accurately localize objects only in one condition. Patients who do not demonstrate a negative directional aftereffect may be able to correctly localize objects with both spectacles and contact lenses. PMID- 10701809 TI - A method for using relay lenses with display monitors for vision testing at far and near. AB - Resolution limitations preclude the use of display monitors for near testing of acuity and contrast sensitivity. Relay lenses can form minified aerial images of the display at any given near viewing distance, but the image will differ in spatial frequency from the display. Equations are presented that can be used to specify the far- and near-viewing distances and the necessary focal length of a lens so that the display and its near aerial image have identical spatial frequencies when viewed by a subject at a fixed location. Modulation transfer function (MTF) calculations show that achromatic doublets will not degrade the resolution across a 300-mm wide display, thereby providing the versatility of display monitors for near vision testing. PMID- 10701811 TI - Angiotensin-converting enzyme gene I/D polymorphism and carotid artery disease in renovascular hypertension. AB - There is evidence linking the activation of the renin-angiotensin system (RAS) with target organ damage in renovascular hypertension (RVH). A genetic association of the DD genotype of the angiotensin-converting enzyme (ACE) gene with cardiovascular complications has been found in various clinical conditions. The aim of our study was to determine whether the insertion/deletion (I/D) polymorphism of the ACE gene is associated with the high prevalence of target organ damage reported in RVH. A total of 65 atherosclerotic patients (age 68.2 +/ 5.2 years) with RVH and 49 atherosclerotic patients (age 68.0 +/- 6.3 years) with essential hypertension (EH) were sequentially enrolled when attending the outpatient clinic for specialist assessment of their vascular disorder. Cardiac, renal, and vascular involvement were assessed in both groups and blood was taken for genetic analysis. Patients with RVH had a higher prevalence of left ventricular hypertrophy (LVH), carotid artery disease, and albuminuria than those with EH. In RVH, but not in EH, the DD genotype was significantly associated with severe arterial disease. In RVH, carotid disease (lumen narrowing >60%) was present in 62% of DD patients versus 25% of the other genotypes (OR = 4.90, 95% CI: 1.70-14.13). Such an association was also present in peripheral vascular disease: 72.4% in DD patients versus 41.6% in the other genotypes (OR = 3.67, 95% CI = 1.29-10.36). Logistic regression analysis showed that the DD genotype was the strongest predictor of risk of severe carotid disease. We conclude that, in atherosclerotic RVH, there is an association of the severity of vascular disease with the DD genotype of the ACE gene. PMID- 10701810 TI - Gene polymorphism of the renin-angiotensin system associates with risk for lacunar infarction. The Ohasama study. AB - The polymorphism of the angiotensin-converting enzyme gene is considered to be associated with increased risk for stroke, but there is a diversity in the results obtained. The genetic involvement of the renin-angiotensin system in stroke also remains unclear. To predict the genetic risk of lacunar infarction, we conducted an association study in an Ohasama population, which is the cohort in a rural region of northern Japan. A total of 134 subjects without major neurological, cardiovascular, or metabolic disorders were recruited. Using brain magnetic resonance imaging, the number of lacunae in each of four brain regions were calculated, and periventricular hyperintensity was classified into five grades. We used the following four candidate gene polymorphisms: angiotensin converting enzyme (ACE)/Insertion(I)-Deletion(D), angiotensinogen (AGT)/M235T, angiotensin II type 1 receptor (AT1)/ A1166C, type 2 receptor (AT2)/C3123A, to examine the association between polymorphisms and the severity of lacunar infarction. AGT/M235T was significantly associated with the number of lacunae in the brain stem, the basal ganglia (P < .05), and whole brain (P < .005) regions. The AT1 polymorphism was also significantly associated with the number of lacunae in the basal ganglia and whole brain regions (P < .05), and with periventricular hyperintensity grade (P < .005) in the younger population. However, ACE and AT2 polymorphisms failed to show an association with either the number of lacunae or the PVH grade. We concluded that AGT and AT1 polymorphisms are independent genetic risk factors for lacunar infarction. PMID- 10701812 TI - Analysis of promoter region polymorphism in the aldosterone synthase gene (CYP11B2) as a risk factor for myocardial infarction. AB - Several polymorphisms in genes of the reninangiotensin-aldosterone system have been found to have pleiotropic effects on cardiovascular disorders. Recently, a polymorphism (-344 C/T) in the promoter region of the aldosterone synthase gene (CYP11B2), which may influence plasma aldosterone levels, has been reported to strongly influence left ventricular diameters and mass in young adults and arterial stiffness in essential hypertensives. We investigated any association with risk of myocardial infarction (MI). CYP11B2 -344 polymorphism genotypes were determined by polymerase chain reaction (PCR) in 542 acute MI cases and 500 control subjects without history of coronary disease. All subjects were white and <75 years old. There was no significant difference in either genotype distributions (cases CC 17%, CT 52%, TT 31%; controls CC 22%, CT 47%, TT 31%, P = .10) or allele frequencies (cases C/T 0.43/0.57, controls C/T 0.46/0.54, P = .39) between cases and controls. The odds ratio (OR) for MI associated with the CC genotype was 0.75 (0.54-1.05), and remained insignificant when analysis was restricted to the 129 (24%) cases and 193 (37%) controls < 55 years of age (OR 0.68 [0.36-1.27], P = .20). In further analyses, there was no interaction of the polymorphism with other cardiovascular risk factors (smoking, hypertension, diabetes, body mass index, or cholesterol level) in determining MI risk, and the polymorphism did not influence the frequency of these risk factors in either cases or controls. In the case cohort, age at MI was not significantly different in subjects with the three genotypes (CC 61.2 +/- 9.8 years, CT 61.8 +/- 9.1 years, TT 62.2 +/- 9.0 years, P = .69). We conclude that the aldosterone synthase -344 promoter region polymorphism does not significantly influence the risk of MI either directly or via interaction with other risk factors. PMID- 10701813 TI - Human G-protein beta3 subunit variant is associated with serum potassium and total cholesterol levels but not with blood pressure. AB - The activity of a sodium-proton exchanger is enhanced in the patients with essential hypertension and regulated via G-protein, which is a signal transducer between receptors and intracellular effectors. A recent study has revealed that a novel variant (C825T) in exon 10 of the gene encoding the beta3 subunit of heterotrimetric G proteins (GNB3) is a genetic factor predisposing to hypertension in Caucasians. We examined the association between GNB3/ C825T and blood pressure, lipids, electrolytes, and other parameters in a Japanese population. Subjects (n = 352) were selected from the Ohasama Study, the population of which is regarded as from a rural community in Japan. To obtain precise clinical measurements, 24-h ambulatory blood pressure monitoring (ABPM), brain magnetic resonance imaging (MRI), and carotid ultrasonography (CUS) were conducted in this population. In addition, we recruited 762 subjects from outpatients at the Osaka University Medical School to carry out the association study between hypertension and GNB3. The GNB3 genotype distribution did not differ significantly between normotensives and hypertensives in either of the two studies. The T825 allele of GNB3 was not associated with the presence of hypertension, blood pressure level, the number of brain lacunae or carotid wall thickness. However, the serum potassium and total cholesterol levels were significantly higher in subjects with the T allele (P < .005). The T825 allele of GNB3 is associated with increased serum potassium and total cholesterol levels but not with blood pressure in a Japanese population. PMID- 10701814 TI - Exposure to cadmium and conventional and ambulatory blood pressures in a prospective population study. Public Health and Environmental Exposure to Cadmium Study Group. AB - This prospective population study investigated in a random sample of 692 subjects (age 20-83 years) how changing environmental exposure to cadmium influenced blood pressure (BP) and the incidence of hypertension. At baseline (1985 to 1989; participation rate, 78%) and follow-up (1991 to 1995; re-examination rate, 81%), blood pressure was measured by conventional sphygmomanometry (CBP; 15 readings in total) and, at follow-up, also by 24-h ambulatory blood pressure monitoring (ABP). Systolic/diastolic CBP at baseline averaged 128.4/77.3 mm Hg. At baseline, blood cadmium concentration (B-Cd) and urinary cadmium excretion (U-Cd) averaged (geometric means) 11.1 nmol/L and 10.2 nmol/24 h. Over 5.2 years (median follow up), B-Cd fell by 29.6% and U-Cd by 15.2%. B-Cd fell less in subjects living closer to three zinc smelters and in premenopausal women. During follow-up, systolic CBP decreased by 2.2 mm Hg in men and remained unchanged in women, and diastolic CBP increased by 1.8 mm Hg in both sexes. No relationship could be demonstrated between the secular trends in CBP and B-Cd or U-Cd or between B-Cd or U-Cd at baseline and the incidence of hypertension. In addition, in cross sectional analyses involving the average of all available CBP measurements in each participant or 24-h ABP at follow-up (mean, 119.1/71.4 mm Hg), blood pressure was not correlated with B-Cd or U-Cd. In conclusion, environmental exposure to cadmium was not associated with higher CBP or 24-h ABP or with increased risk for hypertension. The lesser fall in B-Cd in the residents living closer to the zinc smelters or in premenopausal women underscores the necessity to sanitize cadmium-polluted areas and to systematically reinforce the preventive measures to be adopted by exposed communities to reduce cadmium uptake. PMID- 10701815 TI - Racial differences in response to acute dosing with hydrochlorothiazide. AB - Blacks demonstrate a higher response rate to diuretic therapy for hypertension than do whites. This study examined the pharmacokinetic (PK), pharmacodynamic, and neurohumoral effects of hydrochlorothiazide (HCTZ) administration in a matched group of 9 black and 9 white hypertensive patients (mean +/- SD for black and white). After a 4-week washout period and 7-day control diet, subjects received a single dose of HCTZ (25 mg at 8 AM) with serial blood and urine collections for 36 hours. After HCTZ sodium excretion increased comparably in both groups (blacks: 122 +/- 42 pre to 265 +/- 49 mEq/24 hours post; whites: 117 +/- 29 pre to 255 beta 39 mEq/24 hrs post). Potassium excretion tended to be higher at baseline and was significantly higher following HCTZ in whites (blacks: 45 beta 20 pre to 66 beta 13 mEq at 24 hours post; blacks: 57 +/- 9 pre to 86 +/- 14 mEq at 24 hours post) with most of the post-dosing difference being observed in the hours 0 to 12 after HCTZ. There were no between group PK differences for urinary HCTZ. Aldosterone excretion followed a normal circadian pattern in the whites but did not show this pattern in the blacks. Aldosterone excretion (0 to 12 hours) was generally lower post-dosing in blacks. In conclusion, whereas the PK and single-dose natriuretic response for HCTZ were not racially distinct, potassium excretion was notably less in blacks. Aldosterone excretion was also lower in blacks and without its normal circadian pattern which may, in part, explain their altered potassium excretion pattern. PMID- 10701816 TI - Assessment of vascular aging and atherosclerosis in hypertensive subjects: second derivative of photoplethysmogram versus pulse wave velocity. AB - The pulse wave velocity (PWV) and the photoplethysmogram (PTG) are noninvasive methods for evaluating the pulse wave. The PWV has been associated with age and arterial hypertension, and an index of the second derivative of PTG (SDPTG) is correlated with age and other risk factors for atherosclerosis. The aim of this study was to compare SDPTG and PWV concerning the influencing factors of vascular compliance, including age and atherosclerosis, in a large hypertensive population. We studied consecutively 524 essential hypertensives, 140 with atherosclerotic alterations (AA), defined on the basis of clinical events including coronary heart disease, peripheral vascular disease, stroke, and abdominal aorta aneurysm. The PWV carotid-femoral was measured by a Complior device and the SDPTG was recorded by Fukuda FCP-3166. The augmentation index (AUI) of PTG was defined as the ratio of the late systolic peak to that of the early systolic peak in the pulse. The SDPTG consists of an a,b,c, and d wave in systole and an e wave in diastole; an SDPTG aging index (AI) was calculated as (b c-d-e)/a. The patients with AA presented a higher PWV (14.9 +/- 4 m/sec v 12.4 +/ 2 m/sec, P < .001), PTG AUI (0.322 +/- 0.16 v 0.252 +/-0.09, P < .001), and SDPTG AI (-0.093 +/- 0.03 v -0.271 +/- 0.018, P < .001). However, in patients 60 years of age, only PWV remained higher in those with AA, whereas in patients >60 yr, both PWV and SDPTG AI remained higher in those with AA. The PWV was independently influenced by age, systolic blood pressure, glucose, AA, and plasma creatinine, whereas the PTG AUI was influenced by age and systolic pressure and the SDPTG AI by age and AA. In a logistic regression model for the presence of AA, including age, plasma creatinine, smoking, and diastolic BP, PWV was a significant independent determinant of AA, whereas SDPTG-AI weakly entered into the model. This study provides evidence that the aortic PWV reflects better than the SDPTG the modifications of the arterial compliance related to age, blood pressure, and atherosclerosis. However, the SDPTG AI may be useful for evaluation of vascular aging in hypertensives. PMID- 10701817 TI - Endothelium-derived factors in microalbuminuric and nonmicroalbuminuric essential hypertensives. AB - Previous evidence has demonstrated a relationship between growth factors and cardiovascular diseases. This study was aimed at evaluating levels of some endothelium-derived growth factors, and their relationship with microalbuminuria (MAU), in essential hypertension. Ninety-nine mild-moderate essential hypertensives (EH) and 25 healthy controls were studied. All patients underwent 24-h blood pressure monitoring, serum endothelin-1 (ET-1), basic fibroblast growth factor (bFGF) and platelet-derived growth factor (PDGF), and 24-h MAU assays. Later, EH were divided into two subsets consisting of microalbuminurics (MAU >11 microg/min) and nonmicroalbuminurics (MAU <11 microg/min). In microalbuminuric EH, circulating ET-1, bFGF, and PDGF were significantly higher than in nonmicroalbuminurics (P < .0001, P < .0001, P < .005, respectively) or in controls. In the group of 99 EH, significant positive correlations of MAU with both ET-1 and bFGF (r = 0.35, P < .001, and r = 0.34, P < .001, respectively) were found. ET-1 and bFGF correlated significantly (r = 0.31, P < .002). Circulating bFGF also correlated significantly with MAU in the microalbuminuric EH subset (r = 0.49, P < .01). Our results show that in microalbuminuric EH circulating levels of certain growth factors are increased. In human essential hypertension these factors are linked with MAU, an early cardiovascular and renal damage marker. PMID- 10701818 TI - A clinic and community-based approach to hypertension control for an underserved minority population: design and methods. AB - This paper describes the design and methodology of the Community Hypertension Intervention Project (CHIP). CHIP is investigating the environmental and psychosocial factors related to treatment adherence and examining the effects of combining usual hypertension care with the effects of three interventions designed to improve patient compliance with treatment for high blood pressure in a high-risk, underserved minority population. Thirteen hundred and sixty-seven inner-city hypertension patients (75% black and 25% Hispanic) have agreed to participate in the 4-year longitudinal study. These participants were randomized to usual care or one of three intervention groups: individualized counseling sessions; home visits/discussion groups; or computerized appointment-tracking system. Participants are representative of the surrounding, predominantly low income minority community and are treated in a hospital-based clinic and in a private clinic in the community. About 65% have blood pressure levels considered to be out of control. It was concluded that structural changes at the clinic site, along with the targeted interventions, would improve patient satisfaction, increase treatment adherence, and improve blood pressure control. PMID- 10701819 TI - Electronic pill-boxes in the evaluation of antihypertensive treatment compliance: comparison of once daily versus twice daily regimen. AB - The objective was to compare the compliance of hypertensive patients treated with captopril twice daily or trandolapril once daily. After a 2-week placebo period, hypertensive patients (diastolic BP 95-115 mm Hg) were randomly allocated to trandolapril 2 mg once daily or to captopril 25 mg twice daily for 6 months. Trandolapril and captopril were packed in electronic pill-boxes equipped with a microprocessor that recorded date and time of each opening (MEMS). Patients' compliance was assessed both by standard pill-count and by electronic monitoring. Blood pressure was measured using a validated semi-automatic device at the end of the placebo period and of the treatment period. One hundred sixty-two patients entered the study. Compliance data were evaluable for 133 patients (62 in the captopril group and 71 in the trandolapril group). Treatment groups were comparable at baseline except for age (P = .046). Using electronic pill-box, overall compliance was 98.9% in the trandolapril group and 97.5% in the captopril group (P = .002). The percentage of missed doses was 2.6% in the trandolapril group and 3.3% in the captopril group (P = .06). The percentage of delayed doses was 1.8% in the trandolapril group and 11.7% in the captopril group (P = .0001). The percentage of correct dosing periods, ie, a period with only one correct recorded opening, was 94.0% in the trandolapril group and 78.1% in the captopril group (P = .0001). Results were unchanged when adjusted for age. At the end of the study, 41% of patients in the trandolapril group and 27% in the captopril group (NS) had their blood pressure normalized (systolic BP <140 and diastolic BP <90 mm Hg). In this 6-month study, the electronic pill-box allowed refined analysis of compliance of hypertensive patients. Patients' compliance with once daily trandolapril was higher than with twice daily captopril. The between-group difference is mainly explained by an increase in delayed doses in the twice daily group. PMID- 10701820 TI - Angiotensin II upregulates transforming growth factor-beta type I receptor on rat vascular smooth muscle cells. AB - Angiotensin II (Ang II) and transforming growth factor-beta (TGF-beta) modulate cell growth and metabolism. Our objective was to evaluate the effect of Ang II on the characteristics and expression of TGF-beta receptors on vascular smooth muscle cells (VSMC) from Wistar-Kyoto rats. The addition of TGF-beta1 elicited a biphasic response on DNA synthesis in cultured VSMC in the absence of Ang II, but TGF-beta1 did not stimulate DNA synthesis in the presence of Ang II. TGF-beta binding data showed that Ang II increased the specific binding of 125I-TGF-beta1 by enhancing the expression of lower affinity receptors and increasing the number of binding sites. Ang II alone did not stimulate DNA synthesis in these cultures. However, Ang II significantly stimulated DNA synthesis after the inhibition of endogenous TGF-beta with a neutralizing antibody. The DNA synthesis stimulated by phorbol ester milisterol (PMA) was not affected by the TGF-beta neutralizing antibody. Affinity labeling data revealed receptor-ligand complexes of 280, 85, and 70 kDa, corresponding to TGF-beta type III, II, and I receptors, respectively. Incubation of VSMC with Ang II but not with PMA markedly increased the expression of the TGF-beta type I receptor. Reverse transcription and polymerase chain reaction data also indicated that Ang II, but not PMA, significantly increased the expression of TGF-beta type I receptor mRNA. Results suggest that Ang II increases the binding of TGF-beta with upregulation of TGF beta type I receptor via a C-kinase-independent pathway. The enhanced expression of the TGF-beta type I receptor may counteract Ang II-promoted growth of VSMC. PMID- 10701821 TI - Effect of imidapril on myocardial remodeling in L-NAME-induced hypertensive rats is associated with gene expression of NOS and ACE mRNA. AB - Chronically administered N(omega)nitro-L-arginine methyl ester (L-NAME) produces vascular structural changes and fibrosis of the left ventricle (LV). However, very few studies have evaluated whether the beneficial effects of angiotensin converting enzyme (ACE) inhibitors on these myocardial remodelings are associated with local gene expression of nitric oxide synthase (NOS) and ACE mRNA in the LV. Effects of long term treatment with imidapril, an ACE inhibitor, on gene expression of endothelial-cell NOS (eNOS) and ACE mRNA in the LV and its relation to myocardial remodeling in L-NAME-induced hypertensive rats were evaluated. Fifteen male Sprague-Dawley rats were given L-NAME (60 mg/ kg/day) in drinking water for 6 weeks to induce hypertension, and then treated with imidapril (L-NAME I, n = 8, 1 mg/kg/day, subdepressor dose), or a vehicle (L-NAME-V, n = 7) for 4 weeks. Age-matched rats (C, n = 7) served as a control group. Blood pressure in L NAME-V and L-NAME-I was similar and significantly higher than that in C. The level of eNOS mRNA in the LV was significantly decreased in L-NAME-V compared with C, and was significantly increased in L-NAME-I compared with C and L-NAME-V. The ACE mRNA and type I collagen mRNA expression levels were significantly increased in L-NAME-V compared with C, and significantly suppressed in L-NAME-I compared with L-NAME-V. L-NAME-V demonstrated a significant increase in wall-to lumen ratio, perivascular fibrosis, and myocardial fibrosis. These changes in the microvasculature were improved significantly by imidapril. Myocardial remodeling in L-NAME-induced hypertensive rats was significantly ameliorated by a subdepressor dose of imidapril, which may be due to an increase in local eNOS mRNA expression and a decrease in angiotensin II in the LV. PMID- 10701822 TI - Relaxation of vascular smooth muscle by cicletanine in aged wistar aorta under stress conditions: importance of nitric oxide. AB - The vascular mechanism of action of cicletanine, an antihypertensive agent, was studied on isolated Wistar rat aortas (24-months-old) in presence and in absence of endothelium in two different stress conditions, normoxic and hypoxic, in presence of norepinephrine (NE). Under normoxic conditions, in presence of endothelium, cicletanine (10(-9)-10(-5)M) induced a concentration-dependent relaxation, whereas in absence of endothelium, cicletanine (10(-9)-10(-5)M) was ineffective although it relaxed the smooth muscle at higher concentrations (10( 4)M). At pharmacologic concentrations (below or equal 10(-5)M), relaxation induced by cicletanine, in presence of endothelium, was prevented by N(omega) nitro-L-arginine (L-NNA) (P <.005) and relaxation induced by the highest concentration (10(-4)M) was reversed by BaCl2 (P <.005). Under hypoxic conditions, in presence of NE and endothelium, the aorta displayed an increased developed tension that was significantly (P <.05) attenuated by cicletanine (10( 5)M) and insensitive to indomethacine (10(-7)M). When the two compounds were added together, the relaxation induced by cicletanine was significantly improved (P <.005). These results indicated that cicletanine, under stress conditions, relaxes vascular smooth muscle through an endothelium-dependent action mediated by the nitric oxide (NO) synthase pathway. We proposed that the observed vascular effects could be associated with the counter-regulation mechanisms linked to the antihypertensive action of cicletanine. PMID- 10701823 TI - Candesartan cilexetil is not associated with cough in hypertensive patients with enalapril-induced cough. Multicentre Cough Study Group. AB - The aim of this study was to evaluate the occurrence of dry cough during treatment with candesartan cilexetil, enalapril, or placebo in patients with hypertension and a history of angiotensin converting enzyme (ACE)-inhibitor related cough. Patients with confirmed cough during an enalapril (10 mg) challenge period, followed by no cough during a placebo dechallenge period were randomized to 8 weeks of double-blind treatment with candesartan cilexetil (8 mg) (n = 62), enalapril (10 mg) (n = 66), or placebo (n = 26). Incidence and severity of dry cough was evaluated by the symptom assessment questionnaire, frequency of dry cough by a visual analog scale, and the possible impact on quality of life by the minor symptom evaluation (MSE) profile. The percentage of patients with cough was significantly lower with candesartan cilexetil (35.5%) than with enalapril (68.2%, P < .001), and did not differ between candesartan cilexetil and placebo (26.9%, P > .20). Patients coughed less frequently and with less severe cough with candesartan cilexetil than with enalapril, and similarly with candesartan cilexetil and placebo. Changes in the MSE profile were minor, although candesartan cilexetil had better scores for contentment than placebo (P = .03), and also tended to be associated with better sleep than enalapril (P = .08). In hypertensive patients with ACE-inhibitor-induced cough, the incidence, frequency, and severity of dry cough was significantly lower with candesartan cilexetil than with enalapril, and no different from that found with placebo. PMID- 10701824 TI - Expression of nerve growth factor, brain-derived neurotrophic factor, and neurotrophin-3 in the somatosensory cortex of the mature rat: coexpression with high-affinity neurotrophin receptors. AB - Neurotrophins, including nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neurotrophin-3 (NT-3), are critical for the maintenance and plasticity of central nervous system (CNS) neurons. We tested the hypothesis that cortical neurons participate in redundant autocrine/paracrine systems. Three sets of studies determined the distribution of NGF-, BDNF-, and NT-3-expressing neurons, the frequency of neurons coexpressing NGF and BDNF, and the frequency of neurons expressing a neurotrophin and its associated high-affinity receptor. The distribution of NGF-, BDNF, and NT-3-immunoreactive neurons was identical. Neurotrophin-positive cells were parceled throughout the cortex, although the labeling frequency was not the same in all layers. More than 30% of the neurons in layers II/III, V, and VI were labeled, whereas only 5-10% of the neurons in layer IV was immunopositive for a neurotrophin. Some glia were also neurotrophin positive, particularly BDNF-positive glia. About 70% of the neurons in layers II/III and V coexpressed NGF and BDNF or coexpressed NGF and NT-3. Ligand receptor colabeling was also common among cortical neurons. For example, nearly 70% of the NGF-, BDNF-, and NT-3-positive neurons in layer V colabeled with their respective high-affinity receptors, i.e., trkA, trkB, and trkC, respectively. Thus, (a) neurons express multiple neurotrophins and (b) cortical neurons (e.g., layer V neurons) contain the components required for autocrine/paracrine and/or anterograde communication (e.g., neurons in layer II/III support layer V neurons). These systems mean that the cortex is capable of regulating itself autonomously. PMID- 10701825 TI - Differential synaptic localization of the glutamate transporter EAAC1 and glutamate receptor subunit GluR2 in the rat hippocampus. AB - EAAC1, a neuron-specific glutamate transporter, is likely to play an important role in the regulation of glutamate levels in the synaptic cleft. Ultrastructural studies have demonstrated that the glutamate receptor subunit proteins (e.g., GluR2) are frequently preferentially located at the postsynaptic density of asymmetric synapses. While the glutamate/glutamate receptor interaction is likely to be influenced by the activity and location of the transporter molecules, the spatial localization of the transporter molecules relative to the receptor molecules is not well delineated. Thus, we analyzed the cellular, ultrastructural, and synaptic distribution of EAAC1 in the context of the distribution of the AMPA receptor subunit GluR2 in the hippocampus. While GluR2 and EAAC1 are both present in hippocampal projection neurons, their intracellular distribution patterns differ. Both GluR2 and EAAC1 are present in the dendritic membranes and cytoplasm; however EAAC1 has a distinctive punctate distribution in the dendrite compared to the more diffuse labeling reflected by GluR2. Pre embedding ultrastructural studies also revealed cytoplasmic and membrane associated pools of EAAC1 within dendritic shafts and spines, as well as in a subset of axonal profiles and terminals. Postembedding double label immunogold localization demonstrated a similar intraneuronal distribution, but in addition showed that membrane-associated EAAC1 is not intermingled with GluR2 within the synaptic complex, but in contrast is primarily located perisynaptically, often immediately outside the synaptic specialization. In addition, there is a significant presynaptic pool of EAAC1, whereas GluR2 is essentially absent from the pre-synaptic profile. Thus, membrane-associated EAAC1 within the synaptic region is ideally situated to restrict the site of action of glutamate with respect to ionotropic receptors to the synaptic cleft, as well as regulate glutamate levels in the perisynaptic and presynaptic domains, the ultrastructural sites that have been associated with metabotropic receptor localization. PMID- 10701826 TI - Functional units of a compound nose: aesthetasc sensilla house similar populations of olfactory receptor neurons on the crustacean antennule. AB - The lateral flagellum of the antennule of the spiny lobster Panulirus argus houses more than 1,000 morphologically similar olfactory sensilla, called aesthetascs. By using a high-resolution activity labeling technique that depends on entry of agmatine into olfactory receptor neurons (ORNs) through cation channels during odor stimulation, we examined the distribution of different functional types of ORNs within and across mature aesthetascs. A significant number of ORNs in mature aesthetascs are labeled with agmatine during stimulation by single odorants, including adenosine-5'-monophosphate, ammonium chloride, cysteine, glycine, proline, and taurine. The percentage of ORNs per aesthetasc that was agmatine labeled during odor stimulation averaged 0.5-1.6% for single compounds and 4.6% for a 33-component mimic of oyster tissue. For most antennules and antennular regions studied, the percentage of agmatine-labeled ORNs by stimulation with single or complex odorants was statistically homogeneous across most or all aesthetascs. The extent of heterogeneity among mature aesthetascs was correlated with their age: extensive heterogeneity was observed only in the distal part of the flagellum containing the oldest aesthetascs and their ORNs. Thus, it appears that over most of the length of the aesthetasc-bearing region of the lateral flagellum, different and distinct functional types of aesthetascs do not exist. Rather, aesthetascs appear to be repetitive morphological and functional units in olfactory coding. However, because odor sensitivity of ORNs can change with the age of an aesthetasc, some development-related functional heterogeneity exists among aesthetascs. PMID- 10701827 TI - Development of peripheral hindlimb and central spinal cord innervation by subpopulations of dorsal root ganglion cells in the embryonic rat. AB - The development of rat peripheral hindlimb and lumbar dorsal horn innervation by different subpopulations of dorsal root ganglion cells was investigated from embryonic day (E)13 to birth by using immunostaining. Antibodies to protein gene product (PGP) 9.5, growth associated protein (GAP) 43, and peripherin were used as pan-neuronal markers, RT97 for A fibres; calcitonin gene-related peptide (CGRP), trkA, and the lectin IB4 for small A and C fibres. Size frequency analysis showed that RT97 is a selective marker for large A cells at E18. Although both A and C fibres enter the hindlimb at E13-14, A fibres are the first to innervate the skin and dominate over small fibre innervation until later fetal life. CGRP expression in sensory axons appears at E19 and in all regions simultaneously, suggesting expression in existing fibres. All sensory terminals grow transiently to the skin surface before retracting subepidermally at late embryonic stages. The development of peripheral and central innervation by the same subpopulations of sensory neurons was compared. The entry of A fibre terminals into the lumbar dorsal horn at E14 coincided with hindlimb skin innervation. In contrast, C fibres were not detected in the dorsal horn until E18, 4 days after peripheral innervation. CGRP expression appears in both spinal cord and hindlimb targets at E19. IB4 binding in the central terminals began at E18 but was never observed in embryonic peripheral axons. These results demonstrate that in fetal skin, A fibre innervation dominates over C fibres. In addition, alathough peripheral and central innervation by A fibres coincide, this is not true for C fibres, suggesting that central target factors may control C fibre terminal growth within the dorsal horn. PMID- 10701828 TI - Identification of signal substances in synapses made between primary afferents and their associated axon terminals in the rat trigeminal sensory nuclei. AB - The relationships between primary afferent terminals (PATs) and their associated presynaptic terminals in the rat trigeminal sensory nuclear complex (TSNC) were examined with special reference to amino acid transmitters glutamate (Glu) and gamma-aminobutyric acid (GABA). Primary afferent terminals anterogradely labeled from the trigeminal ganglion with the B subunit of cholera toxin conjugated to horseradish peroxidase (CTB-HRP) were sectioned for electron microscopy. Serial sections from the principal nucleus (Vp), dorsomedial parts of the oral and interpolar nuclei (Vdm), and lamina III/IV of caudal nucleus (Vc) were immunostained for Glu and GABA by using a postembedding immunogold technique. The tracer, CTB-HRP to the trigeminal ganglion, preferentially labeled myelinated primary afferents. Sections immunostained with Glu antiserum showed that most labeled PATs were enriched with immunoreactivity (IR) for Glu. The Glu-IR PATs contained clear, round, synaptic vesicles and formed asymmetric synaptic contacts with somata or dendrites. They were frequently postsynaptic to, unlabeled axon terminals filled with a mixture of clear, round, oval, and flattened vesicles (p endings), with symmetric synaptic junctions. The frequency of synapses onto somata or primary dendrites per Glu-IR PAT was higher in the Vdm than in either the Vp or Vc lamina III/IV. The frequency of contacts of the p-endings per Glu-IR PAT was higher in the Vp than in the Vdm and Vc lamina III/IV. Sections immunostained with GABA antiserum showed that most axon terminals presynaptic to PATs were enriched with GABA in the three nuclei. The GABA-IR axon terminals and their postsynaptic PATs had a similar ultrastructural character to p-endings and their postsynaptic Glu-IR PATs, respectively. The present study suggests that primary afferent neurons with large-caliber fibers use glutamate as a neurotransmitter and are subject to presynaptic modulation by GABAergic fibers. PMID- 10701829 TI - Distribution and developmental changes in GABA-like immunoreactive neurons in the central nervous system of pond snail, Lymnaea stagnalis. AB - We examined three-dimensionally the arrangement of gamma-aminobutyric acid (GABA) like immunoreactive neurons in the central nervous system (CNS) of the pond snail, Lymnaea stagnalis, by a combination of immunohistochemistry and confocal laser scanning microscopy on whole-mount preparations. GABA-like immunoreactivity was detected in all ganglia of the adult CNS. The following distribution of immunoreactive cell bodies was noted in the adult snail. Buccal ganglia: one cell body and five pairs of cell bodies, cerebral ganglia: one pair of cell bodies, pedal ganglia: two single cell bodies, two pairs of cell bodies, and three pairs of cell clusters, and pleural ganglia: one pair of cell bodies. In the asymmetrical parietal ganglia, three cell bodies were located in the left parietal ganglion; three cell bodies and three cell clusters were located in the right parietal ganglion. In the single visceral ganglion, a few scattered individual cell bodies and a cell cluster were GABA-like immunoreactive. Our results showed that the occurrence of GABA is widely spread in the CNS of adult L. stagnalis. GABA-like immunoreactivity in the CNS was not detected in the embryo but was observed after hatching, although the number of stained cells was less than in the adult, with the exception of those in the cerebral ganglia where their number decreased with maturation. Our results provide detailed maps of the central GABA-like immunoreactive neurons in juveniles, immatures, and adults of L. stagnalis. PMID- 10701830 TI - Pontomedullary distribution of 5-HT2A receptor-like protein in the rat. AB - Serotonin (5-HT) exerts excitatory effects in many brainstem regions involved in autonomic, somatic, motor, and sensory functions, and in control of vigilance. To determine the potential role of 5-HT2A receptors in these effects, we immunohistochemically mapped the distribution of 5-HT2A receptor-like protein in the rat pontomedullary brainstem. Areas containing the densest labeling included the trigeminal, facial, hypoglossal, dorsal vagal motor nuclei, medullary linear nucleus, and the inferior olive. In the nucleus ambiguus, labeled cells were located in the areas containing pharyngeal and laryngeal motoneurons. Intensely labeled cells were loosely scattered in the reticular formation adjacent to the raphe magnus and obscurus nuclei, in the gigantocellular region, in the caudal pedunculopontine and laterodorsal tegmental nuclei, dorsomedial pontine reticular formation, and nucleus subcoeruleus. In the nucleus prepositus hypoglossi, all vestibular, abducens, cuneate, and lateral reticular nuclei, labeled neurons commingled with unlabeled ones. Few labeled neurons were located in the rostral and caudal ventrolateral medulla and parvicellular reticular formation. In the nucleus of the solitary tract, two patches of diffuse labeling not associated with cellular profiles were present: one in the medial, and the other in the interstitial subnucleus. Similar diffuse labeling was present in the lateral parabrachial region and the lateral rim of the caudal spinal trigeminal sensory nucleus. No labeled cells were found in the locus coeruleus, dorsal raphe, superior olive, or area postrema. The distinct pontomedullary distribution of 5 HT2A receptors, combined with the known arousal-dependent activity of serotonergic neurons, show that these receptors may mediate post- and presynaptic effects in the motor, selected somatic and visceral sensory, oculo-vestibulo precerebellar, and sleep-related regions. PMID- 10701831 TI - Sexual dimorphism in the song system of the Carolina wren Thryothorus ludovicianus. AB - Sexual and interspecific differences in the size of passerine bird song repertoires are related to differences in the size of song-control regions (SCR) within the brain. Most species of Thryothorus wrens (family Certhiidae) are known to duet, and, in both sexes, song repertoire sizes are related to the size of the SCR. However, one member of this genus, the Carolina wren T. ludovicianus, is very sexually dimorphic in its singing behavior: Males develop large song repertoires, whereas females do not sing. In this study, Nissl staining was used to investigate whether the marked gender difference in the behavior of this species is related to sexual dimorphism of the SCR. Carolina wren males, as predicted, possess the largest premotor song nuclei within the genus; these nuclei could not be identified within Nissl-stained female tissue. The cellular bases for gender differences in SCR morphology also were examined: Males and females differed strongly in the size and density of neurons making up the regions in which SCRs exist in the male forebrain. Interspecific comparison provided no evidence for a decoupling of behavioral and neural evolution within this clade. Male Carolina wrens possess the largest song repertoires and SCRs within the genus, whereas females of this species represent the opposite behavioral and neural extremes of this songbird group. These results are consistent with the hypothesis that the size of the passerine song repertoire is limited by the amount of neural tissue devoted to singing. PMID- 10701832 TI - Expression of presynaptic proteins is closely correlated with the chronotopic pattern of axons in the retinotectal system of the chick. AB - Newly synthesized presynaptic integral membrane proteins in neurons are transported in precursor vesicles from the site of protein biosynthesis in the cell body by fast axonal flow to the presynaptic terminal. We followed the path that presynaptic proteins travel on the way to their central targets of the highly ordered primary visual pathway of the chick and analyzed the developmental changes in the expression of synaptic vesicle protein 2 (SV2), synaptotagmin, and syntaxin. Immunofluorescences revealed that: (1) the onset of protein expression in the retinal ganglion cells occurs in a central to peripheral developmental pattern from embryonic day 4 (E4) onward; (2) the proteins were found first in the inner and later in the outer plexiform layer of the retina; and (3) they were redistributed from the photoreceptor inner segments and cell bodies to the terminals in the outer plexiform layer. From E4 onward, immunopositive axons for SV2, synaptotagmin, and syntaxin were found in the optic nerve, disappearing after E9 for SV2 and synaptotagmin. The optic tract was stained for SV2 and synaptotagmin between E7 and E12, for syntaxin until the posthatching period. Finally, immunoreactivities for the investigated proteins were present at the surface of the tectum from E8 onward, when first retinal axons arrived there. The present study revealed that SV2 and synaptotagmin, but not syntaxin, are, expressed in a transient wave that follows the advancement of optic axons and the proteins towards the optic tectum. PMID- 10701833 TI - Expression of cholesteryl glucoside by heat shock in human fibroblasts. AB - We investigated the heat-induced alteration of glycolipids in human cultured cells, TIG-3 fibroblasts, to show the expression of steryl glucoside by heat shock. A glycolipid band was detected on a thin-layer chromatography plate in lipid extracts from TIG-3 cells exposed to high temperature (42 degrees C) for 15 and 30 minutes, while it was hardly detectable without heat shock. Both cholesterol and glucose were almost exclusively detected by gas liquid chromatography as degradation products of the lipid. The structure of the lipid molecule was elucidated by electrospray mass spectrometry to be a cholesteryl glucoside. This is the first report to show the occurrence of a steryl glucoside in mammalian cells, and this substance is considered to have a significant role in heat shock responses in mammalian cells. PMID- 10701834 TI - Enhanced protein denaturation in indomethacin-treated cells. AB - Indomethacin, a potent anti-inflammatory drug, activates the DNA-binding activity of human heat shock transcription factor 1 (HSF1), but this is insufficient to elevate heat shock gene expression. However, indomethacin pretreatment leads to a complete heat shock response at temperatures that are by themselves insufficient. Here, we showed that the heat-induced loss of enzymatic activity of a nuclear or a cytoplasmic luciferase expressed in murine cells was enhanced when cells had been pretreated with indomethacin. Additionally, in these cells the 70-kDa constitutive heat shock protein exhibited an enhanced aggregation in the presence of indomethacin. Similarly an increase in the aggregation of beta-galactosidase was observed. These data suggest that indomethacin at moderate temperatures accelerates the presence of denatured proteins in the cell, thus lowering the temperature threshold for a heat shock response. PMID- 10701835 TI - Preparation and characterization of polyclonal antibodies against human chaperonin 10. AB - Early pregnancy factor (EPF) has been identified as an extracellular homologue of chaperonin 10 (Cpn10), a heat shock protein that functions within the cell as a molecular chaperone. Here, we report the production of polyclonal antibodies directed against several different regions of the human Cpn10 molecule and their application to specific protein quantitation and localization techniques. These antibodies will be valuable tools in further studies to elucidate the mechanisms underlying the differential spatial and temporal localization of EPF and Cpn10 and in studies to elucidate structure and function. PMID- 10701836 TI - Cloning, sequencing, and transcriptional analysis of the dnaK heat shock operon of Listeria monocytogenes. AB - The complete dnaK operon of Listeria monocytogenes was isolated by chromosome walking using the previously cloned dnaK gene as a probe. Molecular analysis of the locus identified 6 genes in the order hrcA, grpE, dnaK, dnaJ, orf35, and orf29. Primer extension analysis revealed 3 transcription start sites-S1, S2, and S3-upstream of the hrcA, grpE, and dnaJ, respectively. The transcription from S1 was heat inducible. Analysis of the sequences revealed the consensus promoter sequences of gram-positive bacteria, P1 and P2 upstream of the hrcA and dnaJ, respectively. The hrcA gene and a regulatory sequence, designated CIRCE (controlling inverted repeat of chaperone expression), play a role in the regulation of expression of the dnaK locus in response to heat shock in several gram-positive bacteria. Their presence upstream of the dnaK locus in L. monocytogenes suggested a similar regulatory mechanism for the transcription initiated at the promoter, P1. Northern blot analysis led to the detection of 4 mRNA species of 4.9 kb, 3.6 kb, 3.6 kb, and 1.2 kb; the first 2 species were heat inducible. The current results indicate that 4 distinct transcripts directed by 3 promoters are involved in the expression of the dnaK operon of L. monocytogenes. PMID- 10701837 TI - The stress kit: a new method based on competitive reverse transcriptase polymerase chain reaction to quantify the expression of human alphaB-crystallin, Hsp27, and Hsp60. AB - We describe a reverse transcriptase-polymerase chain reaction method for the semiquantitative detection of mRNAs encoding the human heat shock proteins alphaB crystallin, Hsp27, and Hsp60. The method involves the coamplification of cellular mRNA-derived cDNA with a dilution series of a competitor fragment (internal standard), using 1 primer pair common to both templates. Internal standards were based on cellular-derived cDNA engineered to be slightly smaller to differentiate between the target and the standard on electrophoretic separation. Initial cDNA quantitations can be corrected for possible variations during cDNA synthesis by standardizing to the levels of beta-actin-encoding cDNA. We show that the coamplified templates accumulate in a parallel manner with the cellular-derived cDNA throughout both the exponential and the nonexponential phase of amplification. Furthermore, we illustrate the utility of this technique by quantifying increased expression of alphaB-crystallin, Hsp27, and Hsp60 mRNA in astroglioma cells on heat shock. PMID- 10701838 TI - Stress-induced, tissue-specific enrichment of hsp70 mRNA accumulation in Xenopus laevis embryos. AB - In this study, we have employed whole-mount, in situ hybridization to study the spatial pattern of hsc70 and hsp70 mRNA accumulation in normal and heat shocked embryos during Xenopus laevis development. Our findings revealed that hsc70 mRNA was constitutively present in a global fashion throughout the embryo and was not heat inducible. Accumulation of hsp70 mRNA, however, was detected only in heat shocked embryos. Furthermore, hsp70 mRNA accumulation was enriched in a tissue specific manner in X. laevis tailbud embryos within 15 minutes of a 33 degrees C heat shock. Abundant levels of heat shock-induced hsp70 mRNA were detected in the head region, including the lens placode, the cement gland, and in the somitic region and proctodeum. Preferential heat-induced accumulation of hsp70 mRNA was first detected at a heat shock temperature of 30 degrees C. Placement of embryos at 22 degrees C after a 1-hour, 33 degrees C heat shock resulted in decreased hsp70 mRNA with time, but the message persisted in selected tissues, including the lens placode and somites. Treatment of tailbud embryos with either sodium arsenite or zinc chloride induced a tissue-specific enrichment of hsp70 mRNA in the lens placode and somitic region. These studies reveal the complex nature of the heat shock response in different embryonic tissues and suggest the presence of regulatory mechanisms that lead to a stressor-induced, tissue-specific enrichment of hsp70 mRNA. PMID- 10701839 TI - Heat shock transcription factor activation and hsp72 accumulation in aged skeletal muscle. AB - Induction of the protective heat shock proteins (Hsps), and of Hsp72 in particular, has been reported to be decreased in certain tissues from aged animals. To determine if both fast and slow skeletal muscles from aged animals demonstrate an altered ability to induce and accumulate Hsp72, adult (age, 6 months) and aged (age, 20 months) Fischer 344 rats were subjected to heat stress. At selected times (0, 1, 3, and 24 hours) after a 10-minute, 41 degrees C heat stress, fast (white gastrocnemius [WG]) and slow (soleus) skeletal muscles were examined for either heat shock transcription factor (HSF) activation (trimerization and DNA-binding activity) or Hsp72 content using electrophoretic gel mobility shift assays and Western blotting, respectively. Immediately after heat stress, the level of HSF activation between aged and adult animals was similar for both muscles. HSF activation was undetectable at 1 and 3 hours after heat stress in all cases. Twenty-four hours after heat stress, Hsp72 content in the WG muscles from both aged and adult animals was significantly increased compared with unstressed, age-matched controls (P < 0.05). In contrast, perhaps because of their high constitutive Hsp72 levels, soleus muscles from both aged and adult animals did not demonstrate a significant increase in Hsp72 content after heat shock, but there was a trend toward increased levels. Hsp72 content in both the soleus and WG muscles demonstrated no significant differences between adult and aged animals in either the unstressed state (controls) or after heat shock. These results suggest that skeletal muscles from aged animals are capable of inducing the heat shock response and accumulating Hsp72. PMID- 10701840 TI - The Hsp90-specific inhibitor geldanamycin selectively disrupts kinase-mediated signaling events of T-lymphocyte activation. AB - The 90-kDa heat shock protein (Hsp90) is the most abundant molecular chaperone of eukaryotic cells. Its chaperone function in folding nascent proteins seems to be restricted to a subset of proteins including major components of signal transduction pathways (eg, nuclear hormone receptors, transcription factors, and protein kinases). Improper function of these proteins can be induced by selective disruption of their complexes with Hsp90 using the benzoquinonoid ansamycin geldanamycin. In this study, we demonstrate that geldanamycin treatment blocks interleukin (IL)-2 secretion, IL-2 receptor expression, and proliferation of stimulated T-lymphocytes. Moreover, geldanamycin decreases the amount and phosphorylation of Lck and Raf-1 kinases and prevents activation of the extracellular signal regulated kinase (ERK)-2 kinase. Geldanamycin also disrupts the T-cell receptor-mediated activation of nuclear factor of activated T-cells (NF-AT). Treatment with geldanamycin, however, does not affect the activation of lysophosphatide acyltransferase, which is a plasma membrane enzyme coupled to the T-cell receptor after T-cell stimulation. Through demonstrating the selective inhibition of kinase-related T-lymphocyte responses by geldanamycin, our results emphasize the substantial role of Hsp90-kinase complexes in T-cell activation. PMID- 10701842 TI - Defining the efficiency of fluorescence in situ hybridization on uncultured amniocytes on a retrospective cohort of 27407 prenatal diagnoses. AB - Rapid prenatal detection of selected numerical chromosomal abnormalities by using fluorescence in situ hybridization (FISH) on uncultured amniotic fluid samples was described six years ago. It allows a very rapid identification of selected aneuploidies. We have indexed the results of our 27407 fetal karyotypes obtained by conventional cytogenetics during the last five years, noting the type of chromosomal abnormality and the reasons for prenatal diagnosis. We have also indexed the chromosomal abnormality regarding the prognosis of the chromosomal aberations to evaluate the real impact of a non-diagnosis. Within the population of bad prognosis abnormalities, the percentage of abnormalities with bad prognosis detectable by FISH is 94.6% for advanced maternal age, 85.3% for ultrasonographic anomalies and 86.4% for positive maternal screening. The use of FISH alone on our cohort is not a suitable method to diagnose the chromosomal abnormalities. PMID- 10701841 TI - In vivo and in vitro interaction of DnaK and a chloroplast transit peptide. AB - Chloroplast transit peptides have been proposed to function as substrates for Hsp70 molecular chaperones. Many models of chloroplast protein import depict Hsp70s as the translocation motors that drive protein import into the organelle, but to our knowledge, no direct evidence has demonstrated that transit peptides function either in vivo or in vitro as substrates for the chaperone. In this report, we demonstrate that DnaK binds SStp (the full-length transit peptide for the precursor to the small subunit of Rubisco) in vivo when fused to either glutathione-S-transferase (GST) or to an His6-S-peptide tag (His-S) via an ATP dependent mechanism. Three independent biophysical and biochemical assays confirm the ability of DnaK and SStp to interact in vitro. The cochaperones, DnaJ and GrpE, were also associated with the DnaK/SStp complex. Therefore, both GST-SStp and His-S-SStp can be used as affinity-tagged substrates to study prokaryotic chaperone/transit peptide interactions as well as to provide a novel functional probe to study the dynamics of DnaK/DnaJ/GrpE interactions in vivo. The combination of these results provides the first experimental support for a transit peptide-dependent interaction between a chloroplast precursor and Hsp70. These results are discussed in light of a general mechanism for protein translocation into chloroplasts and mitochondria. PMID- 10701843 TI - Prenatal diagnosis and carrier detection for molybdenum cofactor deficiency type A in northern Israel using polymorphic DNA markers. AB - Molybdenum cofactor deficiency (MoCoD) is an autosomal recessive, fatal neurological disorder, characterized by the combined deficiency of sulphite oxidase, xanthine dehydrogenase and aldehyde oxidase. We have recently reported an excessive occurrence of this fatal disorder among segments of the Arab population in Northern Israel suggesting that the true incidence of MoCoD is probably underestimated in this highly inbred population. This lethal disease can be diagnosed prenatally by assay of sulphite oxidase activity in chorionic villus samples in pregnancies of couples who have had previously affected children (obligatory carriers). However, to date, there is no biochemical assay for carrier detection among the population at risk. Recently we demonstrated the linkage of a MoCoD gene to an 8-cM region on chromosome 6p21.3 in two consanguineous Israeli-Arab unrelated kindreds. The description of the MOCS1 gene that maps to the same region and which carries multiple mutations in MoCoD type A followed this finding. We describe here one additional kindred of Arab-Israeli origin, which is also linked to the MOCS1 locus, and demonstrate the feasibility of prenatal diagnosis and carrier detection using microsatellite markers in selected families when mutations are unknown. A complete correlation between the biochemical and DNA assays was found in a total of six samples (five chorionic villus and one amniocyte culture sample) obtained from the three MoCoD families. PMID- 10701844 TI - Serum inhibin A levels in pregnant women with systemic lupus erythematosus or antiphospholipid syndrome. AB - Maternal serum inhibin A levels are increased on average in pregnancies affected by Down syndrome (DS). However, some reports have found increased serum levels in women with pre-eclamptic toxaemia as well. In the current study, maternal serum inhibin A was retrospectively measured in a series of 32 serum samples from pregnant women previously diagnosed as having either systemic lupus erythematosus (SLE) or primary antiphospholipid syndrome (APS). For comparison, normal medians were calculated from 57 unaffected control pregnancies together with a total of 854 samples tested at 13-19 weeks of gestation as part of the routine antenatal DS screening. All results were expressed in multiples of the gestation specific normal medians (MoM). A cubic regression formula was fitted, weighting for the number of women tested at each gestation. The median MoM value in the 16 cases of SLE and the 16 cases of primary APS is 0.60 (95% confidence interval 0.40-0.91) and 0.88 (95% confidence interval 0.66-1.17), respectively. For primary APS this was not statistically significant, whereas the SLE patients had a highly statistically significant reduction of serum inhibin A (p<0.002, Wilcoxon Rank sum Test, 2 tailed). Six pregnancies in the SLE group had a complicated obstetric outcome, i.e. missed abortion, placental abruption, exacerbation of the underlying disease which necessitated delivery, and severe postpartum haemorrhage. In 85% of this subgroup, serum inhibin A levels were below the normal 10th centile. The current data suggest that serum inhibin A is decreased on average in SLE patients. Those preliminary results might have various obstetric implications such as antenatal DS screening of SLE patients, identification of pregnant women at risk of developing SLE, who have presented for routine DS screening and for monitoring SLE patients throughout their pregnancy. PMID- 10701845 TI - Prenatal detection of trisomy 21: combined experience of two British hospitals. AB - A retrospective study was performed to determine the detection rate of trisomy 21 in two British hospitals using a combination of: (1) second trimester serum screening with maternal age, alphaFP and hCG; (2) karyotyping for raised maternal age and high background risk of aneuploidy; and (3) second trimester fetal anomaly ultrasonography at 18-22 week gestation. 36-410 women with a median age of 27 years were studied. Trisomy 21 detected by the combination of methods in both hospitals was compared with the actual number of pregnancies affected by trisomy 21, to determine the detection rate. Serum screening as the backbone of the service detected 31/48 (65%) trisomy 21 affected pregnancies. Karyotyping for maternal age and previous aneuploidy detected eight trisomy 21 affected pregnancies, and second trimester ultrasound a further six, giving a total detection rate of 45/56 (80%). Thus, the detection rate of trisomy 21 in our population is 65% by serum screening alone. This is similar to demonstration projects, but the addition of second trimester ultrasonography and karyotyping for maternal age and prior risk, contributes further to improve the overall sensitivity to 80%. The invasive procedure rate was 4.8% of all women. PMID- 10701846 TI - Survey of attitudes of pregnant women towards Down syndrome screening. AB - This study aimed to examine whether pregnant women made informed decisions based on an accurate understanding of the antenatal screening process and to explore their attitude to screening and termination of a Down syndrome fetus. Women's aspirations were the keystone that informed the development of the first strategy for antenatal screening for congenital anomalies. Semi-structured interviews were carried out with a sample of pregnant women in South Wales in 1995. A total of 34 women aged less than 35 years, who were 20 weeks pregnant, were interviewed. These women were selected because the screening policy differed between hospitals for this age group. The majority of women were not aware that screening tests were voluntary: tests were presented as routine. About half of the sample were not well informed to make decisions. Only five out of a sampling frame of 101 women refused screening; they tended to be better educated and of higher social class. All women wanted to be given the choice whether to be screened. Seven out of 34 would not terminate an affected fetus. Staff communication skills, especially in delivering risk estimate, were criticized. The survey findings supported the view that women required an information package tailored to their individual needs. PMID- 10701847 TI - Fetal 'space-suit' hydrops in the first trimester: differentiating risk for chromosome abnormalities by delineating characteristics of nuchal translucency. AB - Detecting first trimester fetuses with pan-body hydrops, giving the appearance of a 'space-suit,' is associated with a marked increased risk for chromosome abnormalities. In 30 consecutive fetuses prospectively characterized by space suit hydrops, detected at or before 13.9 weeks' gestation, 26 (86.7%) were characterized by chromosome abnormalities. However, as opposed to the preponderance of autosome abnormalities among first-trimester fetuses with prominent nuchal translucencies, 15 of the 26 fetuses (57.7%) with abnormal complements were characterized by sex chromosome aneuploidies. Genetic counselling and consideration of invasive prenatal testing is warranted when space-suit hydrops is detected in the first trimester. PMID- 10701848 TI - Prenatal diagnosis in adenylosuccinate lyase deficiency. AB - Adenylosuccinate lyase deficiency, an autosomal recessive inborn error of purine synthesis, provokes accumulation in body fluids of succinylaminoimidazolecarboxamide riboside and succinyladenosine, the dephosphorylated derivatives of the two substrates of the enzyme. Most patients display severe psychomotor retardation, often accompanied by epilepsy and/or autistic features, although some are only mildly retarded. About 20 mutations are known. Prenatal diagnosis was performed twice on chorion villi of the mother of a previously diagnosed patient with a C5T mutation (exon 1) on the maternal allele, and a C1185A mutation (exon 11) on the paternal allele. Both suppress a Fnu4HI restriction site. In a first fetus, incubation of PCR products generated from genomic DNA of exon 1 with Fnu4HI yielded a 113 bp fragment from a control and the father's gene, and both a 113 bp and 170 bp fragment from the mother, affected sibling and fetus. Incubation of PCR products of exons 11-12 with Fnu4HI yielded a 550 bp fragment from a control and the mother's gene, and a 550 bp and 600 bp fragment from the father, affected sibling and fetus. Assay of adenylosuccinate lyase on the aborted fetal liver confirmed the enzyme deficiency. A second fetus displayed only the maternal mutation. PMID- 10701849 TI - Parental decisions following prenatal diagnosis of sex chromosome aneuploidy: a trend over time. AB - Research over the last 20 years has considerably changed the understanding of the natural history and prognosis for individuals with a diagnosis of sex chromosome aneuploidy (SCA). A cross-sectional retrospective analysis of factors influencing parental decisions following a prenatal diagnosis of SCA during the time period of 1971-97 was performed. The records of 169 fetuses with a prenatal karyotype of 45,X, 47,XXX, 47,XXY, and 47,XYY were reviewed. Mosaic karyotypes for SCA were also included. Information reviewed involved: parental decision, the type of SCA, the presence or absence of mosaicism, the presence or absence of a fetal anomaly diagnosed by ultrasound examination, indication for prenatal diagnosis, prenatal procedure performed, parental age, marital status, previous pregnancy history, family history, ethnicity, religion, education, and profession. A significant correlation was found between the decision to continue a pregnancy and the type of SCA and the presence of fetal abnormalities on ultrasound examination. In addition, this study examined differences in parental decisions over time for the years in question. A statistically significant trend was observed with a higher rate of pregnancy continuation in the more recent years. PMID- 10701850 TI - Cytogenetical diagnosis in paraffin-embedded fetoplacental tissue using comparative genomic hybridization. AB - Comparative genomic hybridization (CGH) is a FISH-related technique used to assess global chromosomal aberrations in a variety of human tumours. Recently CGH has been applied to cytogenetic analysis of fresh frozen fetoplacental tissues. Here we report the application of CGH to paraffin-embedded placental samples. Ten samples from paraffin-embedded blocks of 6 control placentas and fetoplacental tissue from 10 aneuploidies, and 2 unbalanced aberrations were evaluated. Balanced karyotype profiles were obtained from samples of healthy placentas and all samples from the same placenta appeared to have similar confidence intervals. CGH analysis of four cases of trisomy 21, three cases of trisomy 18, one case of trisomy 13, one case of trisomy 15 and one case of trisomy 7 all showed overrepresentation of the respective trisomic chromosome. The CGH profile was also in accordance with the karyotyping of a case with isochromosome 21. The CGH profile of a case with der (2)t(2;6)(q37.3;q22.2) revealed partial trisomy for chromosome 6 between q21 and q27. CGH may be a useful adjunct in prenatal genetic diagnosis when retrospective diagnosis is needed from archival samples. PMID- 10701851 TI - Differentiation between human fetal breathing patterns by investigation of breathing-related tracheal fluid flow velocity using Doppler sonography. AB - We report our results from the analysis of Doppler measurements of breathing related fluid flow velocity waveforms in the trachea in human fetuses. Our aim was to determine whether, using the proposed method, reproducible patterns can be recognized over the latter half of gestation. Breathing-related tracheal fluid flow velocity of 47 normal fetuses at 20-39 weeks' gestation were analysed. Colour Doppler was used to document 'streaming' of fluid in the trachea, followed by spectral Doppler to record flow velocity waveforms. More than 40 (median 94; range 42-725) continuous breathing cycles (inspiration+expiration) were obtained in each case. Although breathing-related fetal tracheal fluid flow waveforms were found to be highly variable, we were able to distinguish by visual analysis between a regular and an irregular pattern. Among the regular patterns, we further differentiate between a regular symmetric (sinusoidal type) and a regular asymmetric (deep inspiration with expiratory flow retardation) pattern. The regular pattern occurred consistently in all age groups studied and there were no significant (p<0.05) differences in the occurrence rate of the regular symmetric and asymmetric pattern. The incidence of the regular pattern increased significantly (p<0.05) from 11.74+/-3.38% (mean +/- SEM) at 24-27 weeks to 20.72+/-1.75% at 28-31 weeks of gestation and remained constant thereafter. This study shows that the proposed method can provide detailed information on breathing-related tracheal fluid flow velocity as early as 20 weeks of gestation. The information that a regular symmetric pattern was observed throughout the second half of gestation is important. Hence, a higher standardization of on going fetal breathing movements studies may be achieved by measuring breathing related tracheal fluid flow velocity waveform parameters only during this pattern. PMID- 10701852 TI - Prenatal diagnosis and outcome in sacrococcygeal teratomas: a review of cases between 1992 and 1998. AB - A review of sacrococcygeal teratomas diagnosed in the antenatal period in the West Midlands region over a six year interval is reported. The aim of the study was to assess the contribution of ultrasound scanning to the management of cases and to determine the outcome of prenatally diagnosed sacrococcygeal teratomas. A retrospective review of 10 cases was performed to obtain pregnancy details, ultrasound scan data and outcome information. Two fetuses were electively aborted. Perinatal mortality was 62.5% in the remaining cases with all stillbirths and neonatal deaths occurring in babies delivered preterm (at or before 34 weeks' gestation). Marked increase in tumour size (mainly vascular/solid) was observed in five of the fetuses, which was often associated with local compression effects and the development of hydrops. Eight out of 10 cases were delivered vaginally, one following aspiration of the large cystic tumour. Three of the four neonates surviving to surgery underwent successful resection of their benign tumours. As well as guiding prognosis, serial ultrasound scans may also allow the mode of delivery to be planned more effectively. The importance of a multidisciplinary team approach to these difficult cases is emphasized. PMID- 10701853 TI - Prenatal diagnosis of RAG-deficient Omenn syndrome. AB - Mutations in recombination activating genes (RAG) 1 and 2 have been found to cause Omenn syndrome (OS), a severe combined immunodeficiency (SCID) with a peculiar phenotype. Here we report the prenatal diagnosis performed in three OS patients. Mutations were detected in the probands as well as in their parents by genomic sequencing of the complete coding regions of both RAG 1 and RAG 2, which are contained in a single exon. All the three probands had RAG 1 mutations in both alleles, at least one of which was a missense substitution. Of the three fetuses tested, one had a wild type sequence on both alleles, while the other two had one mutated allele. None of the three patients were predicted to be affected and this was confirmed at birth. Detection of RAG genes mutations on fetal samples by direct sequencing is an easy and effective way to investigate fetuses from families affected with RAG-dependent SCID and OS families affected by RAG dependent SCID and OS. PMID- 10701854 TI - Gaucher disease: considerations in prenatal diagnosis. PMID- 10701855 TI - Rapid identification of a small dicentric supernumerary marker derived from chromosome 16 with a modified FISH technique on amniotic fluid. AB - Small supernumerary marker chromosomes are seldom found in prenatal diagnosis and the majority of them are difficult to identify. The only possibility to give a more precise prognosis is by establishing its origin. FISH is the best technique to identify the chromosomal origin, but in the majority of cases large amounts of chromosomal material are needed and this is time consuming. We have used a modification of the FISH technique that allows the hybridization of several probes on one slide. Using this method, we have identified the first de novo mosaic dicentric supernumerary marker derived from chromosome 16 (smaller than chromosome 21) in amniotic fluid. The gestation and the follow-up of the baby were normal. PMID- 10701856 TI - Rapid detection of expansions by PCR and non-radioactive hybridization: application for prenatal diagnosis of myotonic dystrophy. AB - The mutation specific for myotonic dystrophy (DM) is an unstable expanded CTG repeat located in the 3'-untranslated region of the myotonin protein kinase gene. Expansion of the CTG repeat shows a positive correlation with the severity of the disease and increases in successive generations of DM patients. Children with the congenital form of DM show the most severe phenotype and have large expansions, usually > 1000 repeats. For pregnant women with DM, prenatal diagnosis of DM may be offered. To reduce the time between chorionic villus sampling or amniocentesis and final results of DNA analysis in these cases, a fast and efficient method has been developed. This method combines direct PCR analyses for normal alleles with a nested PCR system followed by non-radioactive hybridization with a single stranded probe. PMID- 10701857 TI - Prenatal diagnosis of pyloric atresia-junctional epidermolysis bullosa syndrome in a fetus not known to be at risk. AB - Junctional epidermolysis bullosa with pyloric atresia (PA-JEB) is a highly lethal, inherited, autosomal recessive disease. Thus far, prenatal diagnosis of this syndrome was only realized on pregnancies at risk for recurrence. We report the case of a 26-year-old woman, first cousin to her husband, who had undergone amniocentesis for polyhydramnios. The karyotype was normal but the amniotic fluid contained acetylcholinesterase. A targeted scan at 25 weeks' gestation did not find spina bifida, but polyhydramnios with a dilated stomach, and several other anomalies: echogenic particles in the amniotic fluid, a thin skin which closely adhered to the nasal bones, narrow nostrils, abnormal ears, fisted hands, malposition of both first toes, and kidney malformation. Despite no previous case in the family, it was thought that sonographic findings were suggestive of the PA JEB syndrome. A fetal skin biopsy was carried out at 28 weeks' gestation. The ultrastructural examination of fetal skin displayed JEB. Genetic analysis detected a homozygous mutation in the gene encoding integrin alpha 6. Termination of pregnancy was carried out at 29 weeks' gestation. These results illustrate that in the case of a fetus not known to be at risk, diagnosis of PA-JEB can be achieved by ultrasound findings leading to fetal skin biopsy and ultrastructural examination of blistered epidermis. Some new sonographic signs should raise the possibility of significant cutaneous desquamation and blister formation in a fetus, especially when there is positive amniotic acetylcholinesterase coupled with elevated alpha-fetoprotein or suspected pyloric atresia. PMID- 10701858 TI - Prenatal diagnosis of mosaicism for a del(22)(q13). AB - A case of prenatally detected mosaicism for a del(22)(q13) is reported. CVS was performed because of abnormal fetal ultrasound findings: cystic 'tumour' in the fetal neck and the upper thoracic aperture. Karyotypes from chorionic villi were suspicious of an aberration concerning the long arm of one chromosome 22. FISH analysis demonstrated mosaicism for a distal 22q deletion in fetal fibroblasts. The deletion was postnatally confirmed by FISH with a chromosome-specific 22q probe. The 'tumour' on autopsy turned out to be cystic thymic tissue. Apart from this, no other obvious fetal anomalies were found. PMID- 10701859 TI - Early and mid-trimester amniocentesis. PMID- 10701860 TI - Maternal serum hyaluronic acid is ineffective for Down syndrome screening. PMID- 10701861 TI - Current awareness in prenatal diagnosis. PMID- 10701862 TI - Native extracellular matrix induces a well-organized bipolar outgrowth pattern with neurite extension and retraction in cultured neurons. AB - Cultured anterior pagoda (AP) neurons from the leech develop characteristic outgrowth patterns that depend on the molecular composition of the substrate. This article analyzes how native substrates from the central nervous system (CNS), such as the extracellular matrix (ECM) inside the capsules that enwrap the ganglia, determine the outgrowth patterns of AP neurons. When plated on the internal side of ganglion capsules, the remaining primary portion (stump) of AP neurons sprouted two main branches in opposite directions with bifurcations. This T-shaped pattern was distinctive for AP neurons and was different from the patterns of the same cell type plated on the external side of the capsule or on leech laminin extracts, in which they generated multiple neurites and branching points. AP neurons plated on tritonized CNS homogenates reproduced the outgrowth pattern displayed on ganglion capsules, in terms of the number of primary neurites, their length, their orientation, and the number of branch points. The development of the T-shaped outgrowth pattern of AP neurons on ganglion capsules and CNS homogenates started by the sprouting of one branch that later bifurcated, followed by a second branch in the opposite direction after a lag of several hours. Extension of the second branch and retraction of secondary neurites of the first were synchronous and contributed to refine the T-shaped pattern. These results suggest that during development or regeneration of the CNS, particular sets of ECM proteins have multiple effects regulating the number, direction, extension, and retraction of neurites. PMID- 10701864 TI - Connective tissue response to tubular implants for peripheral nerve regeneration: the role of myofibroblasts. AB - The presence of contractile cells, their organization around regenerating nerve trunks, and the hypothetical effect of these organized structures on the extent of regeneration across a tubulated 10-mm gap in the rat sciatic nerve were investigated. Collagen and silicone tubes were implanted both empty and filled with a collagen-glycosaminoglycan (GAG) matrix. Nerves were retrieved at 6, 30, and 60 weeks postoperatively and time-dependent values of the nerve trunk diameter along the tubulated length were recorded. The presence of myofibroblasts was identified immunohistochemically using a monoclonal antibody to alpha-smooth muscle actin. Myofibroblasts were circumferentially arranged around the perimeter of regenerated nerve trunks, forming a capsule which was about 10 times thicker in silicone tubes than in collagen tubes. The nerve trunk diameter that formed inside collagen tubes was twice as large as that inside silicone tubes. In contrast, the collagen-GAG matrix had a relatively small effect on capsule thickness or diameter of regenerate. It was hypothesized that the frequency of successful bridging by axons depends on the balance between two competitive forces: the axial forces generated by the outgrowth of axons and nonneuronal cells from the proximal stump and the constrictive, circumferential forces imposed by the contractile tissue capsule that promote closure of the wounded stumps and prevent axon elongation. Because the presence of the collagen-GAG matrix has enhanced greatly the recovery of normal function of regenerates in silicone tubes, it was hypothesized that it accelerated axonal elongation sufficiently before the hypothetical forces constricting the nerve trunk in silicone tubes became sufficiently large. The combined data suggest a new mechanism for peripheral nerve regeneration along a tubulated gap. PMID- 10701863 TI - Transneuronal labeling from the rat distal colon: anatomic evidence for regulation of distal colon function by a pontine corticotropin-releasing factor system. AB - Neural circuits that are positioned to regulate rat distal colon function were identified by immunohistochemical detection of pseudorabies virus (PRV) and corticotropin-releasing factor (CRF). The distribution of PRV-immunoreactive neurons was examined in spinal cord and brain at increasing times (72-118 hours) after distal colon injection. At 72-80 hours, PRV-labeling was confined to the spinal cord, in the parasympathetic preganglionic column in the lumbosacral spinal cord and in the intermediolateral column of the thoracic spinal cord. At longer survival times (88 hours), PRV-immunolabeled neurons in the lumbosacral spinal cord were also distributed in superficial layers of the dorsal horn, the dorsal commissure, and around the central canal. Trans-synaptic labeling was identified in the medullary raphe nuclei, parapyramidal region, A5, Barrington's nucleus, A7, and the dorsal cap of the paraventricular nucleus of the hypothalamus after longer survival times (88-91 hours). Substantial labeling of the locus coeruleus, periaqueductal gray and forebrain regions occurred at later survival times (> or = 96 hours). In dual-labeled sections, CRF terminal labeling surrounded PRV-labeled neurons in the parasympathetic preganglionic column of the lumbosacral spinal cord. Additionally, many neurons in Barrington's nucleus, but not other CRF-containing nuclei, were double labeled for CRF and PRV. These results, taken with previous studies, support a convergence in transneuronal labeling from different pelvic viscera that may be related to coordination of overall pelvic visceral functions. Importantly, they provide an anatomic substrate for an impact of CRF from Barrington's nucleus in normal and pathophysiological functions of the distal colon. PMID- 10701865 TI - Microglial cells in the retina of Carassius auratus: effects of optic nerve crush. AB - To study the morphology and distribution of the retinal microglial cells of the goldfish retina in normal conditions and after optic nerve crush, we have used the nucleoside diphosphatase (NDPase) technique, applied to whole-mounts or sections, for light and electron microscopy. In normal retinas, two populations of NDPase-positive cells were identified: compact cells associated with the retinal vessels on the vitreal surface of the retina and microglial cells in various retinal layers. The microglial cells had a bipolar or multipolar morphology. Bipolar cells were observed in the nerve fibre layer, and multipolar cells were visualised in the ganglion cell layer (GCL), inner plexiform layer (IPL), and outer plexiform layer. The highest densities of multipolar cells were observed in the IPL layer, where they adopted a regular mosaic-like arrangement in which the occasional spaces were occupied by cells of the GCL. After optic nerve crush, we observed an increase in the number of compact cells associated with the vessels and changes in NDPase activity, morphology, and distribution of the retinal microglial cells. These cells showed an increase in NDPase activity in all retinal layers from day 1 to day 15 after axotomy, and retraction of their processes from day 1 to day 7. In addition, the densities of microglial cells increased in the GCL between 2 and 15 days after axotomy, and decreased in the IPL by day 4 after axotomy. These microglial changes resemble those observed in other regenerating and nonregenerating neuronal systems and may reflect a general response of microglia directed to help the regeneration process. PMID- 10701866 TI - Dorsal medullary pathways subserving oromotor reflexes in the rat: implications for the central neural control of swallowing. AB - Retrograde and anterograde axonal transport techniques were used to investigate the organization of inputs from the dorsomedial medulla, a region known to elicit patterned swallowing reflexes following focal stimulation, to the fifth (MoV), seventh (VII), tenth (nucleus ambiguus, NA), and twelfth (XII) cranial nerve motor nuclei in the rat, those motor nuclei most directly involved in the control of deglutition. The results may be summarized as follows. 1) Dorsal medullary inputs to MoV, VII, and XII arise primarily from an extended region of the caudal reticular formation immediately ventral to the nucleus of the solitary tract (NTS), which we term the dorsal medullary reticular column (DMRC). Projections from the DMRC are largely bilateral and are distributed preferentially to the ventral subdivision of MoV, to the dorsal and intermediate subdivisions of VII, and to both the dorsal and the ventral subdivisions of XII. In addition, a subpopulation of large multipolar neurons embedded within the DMRC gives rise to a primarily crossed input to the dorsal subdivision of MoV. 2) Dorsal medullary inputs to the NA arise from the NTS, are largely uncrossed, and are organized such that the ventrolateral, intermediate, and interstitial subdivisions of the NTS project to the semicompact formation and to the rostral extension of the compact formation (which supplies the pharynx) and to the loose formation (larynx), whereas the central subdivision of the NTS provides input to the compact formation (esophagus). 3) Neither the NTS nor the DMRC gives rise to significant projections to the central subnucleus of the NTS. Together, these results provide evidence for discrete medullary pathways subserving sequential activation of swallowing reflexes. PMID- 10701867 TI - Innervation of the digit on the forepaw of the raccoon. AB - The innervation of the digits on the raccoon forepaw was examined by using immunochemistry for protein gene product 9.5, calcitonin-gene related peptide, substance P, neuropeptide-Y, tyrosine hydroxylase, and neurofilament protein. The larger-caliber axons in the ventral glabrous skin terminate as Pacinian corpuscles deep in the dermis, small corpuscles and Merkel endings around the base of dermal papillae, and Merkel endings on rete pegs in dermal papillae. Extensive fine-caliber innervation terminates in the epidermis and on the microvasculature. The innervation is more dense in the distal than in the proximal volar pads. Pacinian endings are also concentrated in the transverse crease separating the distal and proximal pads. In the dorsal hairy skin, hair follicles are well innervated with piloneural complexes. Merkel innervation is located under slight epidermal elevations and in some large Merkel rete pegs located at the apex of transverse skin folds just proximal to the claw. No cutaneous Ruffini corpuscles were found anywhere on the digit. The claw is affiliated with dense medial and lateral beds of Pacinian endings, bouquets of highly branched Ruffini-like endings at the transition from the distal phalanx and unmyelinated innervation in the skin around the perimeter. Encapsulated endings are located at the lateral edge of the articular surface of the distal phalanx. Extensive fine-caliber innervation is affiliated with sweat glands and with the vasculature and is especially dense at presumptive arteriovenous sphincters. Virtually all of the sweat gland and vascular innervation is peptidergic, whereas most of the unmyelinated epidermal innervation is nonpeptidergic. PMID- 10701868 TI - Afferents of cranial sensory ganglia pathfind to their target independent of the site of entry into the hindbrain. AB - In vertebrates, sensory neurons interconnect a variety of peripheral tissues and central targets, conveying sensory information from different types of sensory receptors to appropriate second-order neurons in the central nervous system (CNS). To explore the possibility that the different rhombomere environments where sensory neurons enter into the hindbrain affect the pathfinding capability of growth cones, we studied the development of the VIIIth ganglion afferent both in vivo and in vitro. We focused on the vestibular nerve because it is the only cranial nerve projecting to the cerebellum, allowing for ready identification from its pattern of projection. Embryonic rat brain was cut along the dorsal midline and, with the VIIIth and Vth ganglia still attached, flat mounted and visualized with antibodies specific for sensory ganglia. Axons reached the cerebellar primordium at embryonic day (E) 13, then splayed out towards the edges of the rhombic lip of rostral hindbrain. In vitro, the VIIIth ganglion showed development similar to that in vivo and innervated the cerebellum, an appropriate target, indicating that mechanisms for axon guidance and target recognition are preserved in vitro. When the VIIIth ganglion was transplanted to the position of the Vth ganglion, axons from the transplanted ganglion entered the cerebellar primordium with a trajectory characteristic of the VIIIth nerve. These results indicate that the central projection pattern of the VIIIth nerve is not affected by the environment of nerve entry into the brainstem, suggesting that axons of sensory cranial ganglion intrinsically possess the capacity to find their target correctly. PMID- 10701870 TI - Spatial training and high-frequency stimulation engage a common pathway to enhance glutamate release in the hippocampus. AB - We have measured depolarization-induced release of endogenous glutamate in synaptosomes prepared from the dentate gyrus after the induction of LTP by high frequency stimulation in anesthetized rats, and after training in the water maze. Both spatial training and LTP in untrained rats were accompanied by an increase in glutamate release from dentate synaptosomes. The enhancement of synaptosomal glutamate release induced by high-frequency stimulation was abolished in well trained rats, and was reduced in partially trained rats and in rats trained in a nonspatial task. However, the magnitude of LTP was similar in well-trained and untrained groups. These results indicate that spatial training activates a glutamate release pathway that converges with that activated in LTP, and demonstrate an unexpected dissociation between increased glutamate release and LTP. PMID- 10701869 TI - Concentrated expression of Ca2+/ calmodulin-dependent protein kinase II and protein kinase C in the mushroom bodies of the brain of the honeybee Apis mellifera L. AB - We have previously used the differential display method to identify a gene that is expressed preferentially in the mushroom bodies of worker honeybees and to show that it encodes a putative inositol 1,4,5-trisphosphate receptor (IP3R) homologue (Kamikouchi et al. [1998] Biochem. Biophys. Res. Commun. 242:181-186). In the present study, we examined whether the expression of some of the genes for proteins involved in the intracellular Ca2+ signal transduction is also concentrated in the mushroom bodies of the honeybee by isolating cDNA fragments that encode the Ca2+/calmodulin-dependent protein kinase II (CaMKII) and protein kinase C (PKC) homologues of the honeybee. In situ hybridization analysis revealed that the expression of these genes was also concentrated in the mushroom bodies of the honeybee brain: The CaMKII gene was expressed preferentially in the large-type Kenyon cells of the mushroom bodies, whereas that for PKC was expressed in both the large and small types of Kenyon cells. The expression of the genes for IP3R and CaMKII was concentrated in the mushroom bodies of the queen and drone as well as in those of the worker bee. Furthermore, the enzymatic activities of CaMKII and PKC were found to be higher in the mushroom bodies/central bodies than in the optic and antennal lobes of the worker bee brain. These results suggest that the function of the intracellular Ca2+ signal transduction is enhanced in Kenyon cells in comparison to other neuronal cell types in the honeybee brain. PMID- 10701871 TI - Long-term habituation to spatial novelty in blind cave fish (Astyanax hubbsi): role of the telencephalon and its subregions. AB - Blind cave fish, when released into a novel environment, show a typical exploratory behavior characterized by high swim speed along walls shortly after release. This behavior wanes during prolonged exposure and thus may reflect habituation to novelty. As the hippocampus of mammals, which plays a crucial role in spatial learning, is part of the telencephalon, the possible involvement of this brain structure of fish was investigated in exploratory behavior. Ablation of the whole telencephalon or bilateral removal of dorsal parts of the hemispheres reduced activity; in contrast, unilateral lesions of one hemisphere, bilateral lesions of dorsal and dorsoventral parts, and removal of olfactory bulbs increased activity. However, the time course of habituation in a novel environment remained unchanged, except for ablated animals in which there was virtually no habituation in swim speed. These data suggest that the telencephalon of fish may not participate in long-term habituation to spatial novelty but, rather, support the notion of the telencephalon being involved in generation of arousal. PMID- 10701872 TI - Protein synthesis-dependent memory and neuronal enhancement in Hermissenda are contingent on parameters of training and retention. AB - Following contiguous pairings of light and rotation, light alone elicits a conditioned contraction of Hermissenda's foot, indicative of an associative memory. After a 5-min retention interval, this conditioned response was evident following two or nine (but not one) conditioning trials but persisted for 90 min only after nine trials. In vivo incubation of animals in the protein synthesis inhibitor anisomycin (ANI; 1 microM) did not affect the conditioned response at the 5-min retention interval but significantly attenuated conditioned responding at the 90-min interval even following nine training trials. Deacetylanisomycin (DANI; 1 microM; an inactive form of anisomycin) had no effect on either 5- or 90 min retention. In a companion procedure, groups of isolated nervous systems were exposed to comparable light and rotation pairings, and the B photoreceptors (considered a site of storage for the associative memory) underwent electrophysiological analysis. An increase in neuronal excitability (indexed by depolarizing voltage responses to injected current) in the B photoreceptors paralleled the expression of conditioned responding in intact animals, that is, two training trials produced a short-term increase in excitability that dissipated within 45 min, whereas nine trials produced a persistent (at least 90 min) increase in excitability. In a fmal experiment, isolated nervous systems were exposed to nine training trials, and ANI or DANI was either present in the bathing medium before and during training or was introduced 5 min after training. Following training in ANI, a short-term (5- to 45-min) but not persistent (90 min) increase in excitability in the B photoreceptors was observed. ANI had no effect on either the short-term or persistent increase in excitability if the drug was applied 5 min after the last (ninth) training trial, and DANI had no effect on training-induced increases in excitability at any retention intervals. These results suggest that short-term retention in Hermissenda is protein synthesis independent but that new protein synthesis initiated during or shortly after the training event is necessary for even 90-min retention. Moreover, these results indicate that under some conditions, a critical threshold of training must be exceeded to initiate protein synthesis-dependent retention. PMID- 10701873 TI - Identification of specific mRNAs affected by treatments producing long-term facilitation in Aplysia. AB - Neural correlates of long-term sensitization of defensive withdrawal reflexes in Aplysia occur in sensory neurons in the pleural ganglia and can be mimicked by exposure of these neurons to serotonin (5-HT). Studies using inhibitors indicate that transcription is necessary for production of long-term facilitation by 5-HT. Several mRNAs that change in response to 5-HT have been identified, but the molecular events responsible for long-term facilitation have not yet been fully described. To detect additional changes in mRNAs, we investigated the effects of 5-HT (1.5 hr) on levels of mRNA in pleural-pedal ganglia using in vitro translation. Four mRNAs were affected by 5-HT, three of which were identified as calmodulin (CaM), phosphoglycerate kinase (PGK), and a novel gene product (protein 3). Using RNase protection assays, we found that 5-HT increased all three mRNAs in the pleural sensory neurons. CaM and protein 3 mRNAs were also increased in the sensory neurons by sensitization training. Furthermore, stimulation of peripheral nerves of pleural-pedal ganglia, an in vitro analog of sensitization training, increased the incorporation of labeled amino acids into CaM, PGK, and protein 3. These results indicate that increases in CaM, PGK, and protein 3 are part of the early response of sensory neurons to stimuli that produce long-term facilitation, and that CaM and protein 3 could have a role in the generation of long-term sensitization. PMID- 10701874 TI - Analysis of sequence-dependent interactions between transient calcium and transmitter stimuli in activating adenylyl cyclase in Aplysia: possible contribution to CS--US sequence requirement during conditioning. AB - An important recent insight in a number of neurobiological systems is that during learning, individual dually regulated proteins with associative properties function as critical sites of stimulus convergence. During conditioning in Aplysia, the Ca2+ /calmodulin-sensitive adenylyl cyclase (AC) in mechanosensory neurons serves as a molecular site of interaction between Ca2+ and serotonin [5 hydroxytryptamine (5-HT)]-two signals that represent the CS and US in these cells. Conditioning requires that the CS and US be paired within a narrow time window and in the appropriate sequence. AC shows an analogous sequence preference: It is more effectively activated when a pulse of Ca2+ precedes a pulse of 5-HT than when the 5-HT precedes Ca2+. One mechanism that contributes to this sequence preference is that Ca2+/calmodulin binding to AC accelerates the rate of AC activation by receptor-Gs. We have identified two additional properties of AC activation that would cause pairing with Ca2+ preceding 5-HT to be more effective than simultaneous pairing or pairing with the reciprocal sequence: (1) Activation of Aplysia AC by a Ca2+ pulse rose with a delay compared with activation by a 5-HT pulse. (2) A late pulse of Ca2+, which arrived after 5 HT, acted, via calmodulin, to accelerate the decay of AC activation by receptor Gs. Together, these activation properties of AC may contribute to the CS-US sequence requirement of classical conditioning. PMID- 10701875 TI - Perforant path activation modulates the induction of long-term potentiation of the schaffer collateral--hippocampal CA1 response: theoretical and experimental analyses. AB - In one computational model of hippocampal function, the entorhinal cortical input to CA1 is hypothesized to play a key role in the ability of CA1 to decode CA3 recodings. Here, we develop a modification of this CA1 decoder hypothesis that is applicable to several computational theories of hippocampal function, and then we electrophysiologically investigate one assumption of this new hypothesis. First, using biologically realistic estimates, we calculate that CA3-induced CA1 excitation is too high and that inhibition plausibly plays a role in this CA1 decoder model. Thus motivated, we turn to a physiological demonstration to substantiate the plausibility of the proposed mechanism. Using the rat hippocampal slice, we examine an interlaminar interaction between the distal perforant path input to hippocampal CA1 stratum moleculare and the more proximal Schaffer collateral input to stratum radiatum. Perforant path activation provides sufficient inhibition to block homosynaptic long-term potentiation elicited by a suitably strong stratum radiatum input. For this interlaminar interaction to be most effective, perforant path activation must both precede and follow Schaffer collateral activation. Perforant path-evoked inhibition in CA1 can thus serve as a viable mechanism in the learned decoder theory of hippocampal CA1. PMID- 10701876 TI - Behavioral modulation induced by food odor aversive conditioning and its influence on the olfactory responses of an oscillatory brain network in the slug Limax marginatus. AB - We compared behaviorally and physiologically the olfactory responses of slugs (Limax marginatus) that had been subjected to aversive, appetitive, or unpaired training with food odors (carrot or cucumber). In the aversive training, the slugs were exposed to the food odor as a conditioned stimulus (CS), and then quinidine sulfate solution as an unconditioned stimulus (UCS) was immediately applied to the lip of the slugs. This training caused a decrease in preference level for the CS. The unpaired training, in which the CS and the UCS were presented to the slugs with a 5-min interval, induced no change in the preference level for the CS. In the appetitive training, the slugs were allowed to eat the CS odor source without UCS application. When we used nonstarved slugs, it was found that the preference level for the CS increased upon the appetitive training. These results indicate that each training changed the preference for the odors in a characteristic manner. In the physiological experiments, we used brain-inferior tentacular nose preparations isolated from slugs and investigated the olfactory responses of the oscillations in the local field potential (LFP) of the procerebral (PC) lobe. We found that odor presentation induced various types of changes in the LFP oscillation frequency, although the rate of occurrence of the frequency modulation differed between odors used in the aversive and the unpaired training (aversive-conditioned and unpaired odors). The aversive conditioned odors induced a decrease in the oscillatory frequency. Unpaired odors did not change it. Moreover, odors used in the appetitive training (appetitive conditioned odors) induced an increase in the frequency. Thus, it was considered that those modulations of PC lobe oscillatory activity were independent of odor and reflected learned preference for odors. PMID- 10701878 TI - Optical recording analysis of olfactory response of the procerebral lobe in the slug brain. AB - We studied the oscillatory properties and the olfactory responses of the procerebral (PC) lobe of the cerebral ganglion in the terrestrial mollusc Limax marginatus. The PC lobe, a central olfactory organ in Limax, is a highly interconnected network of local interneurons that receives olfactory inputs from the inferior and superior tentacular noses. We used an optical recording technique with a voltage-sensitive dye to record the activity of the PC lobe from either the posterior or the dorsal surface. The recordings revealed that almost all PC interneurons showed spontaneous oscillatory activities that had been entrained with each other. Upon presentation to the nose of odors to which the slugs had been aversively conditioned, the basal level of the oscillation changed biphasically. In the early phase of the response, depolarization in the basal level of the oscillation occurred in one or more belt-shaped regions parallel to the dorsoventral axis. In the late phase of the response, hyperpolarization of basal potential level of the PC lobe oscillations occurred in a wider area. Such spatial and temporal modulation was not observed when the unpaired control odors were presented to the preparation, whereas the same preparations responded to the aversively conditioned stimuli. Thus, it was considered that the spatial and temporal response in the basal level of oscillation was specific to the aversively conditioned odors. Furthermore, the spatial pattern of the depolarization modulation in the early phase was repeatable in multiple trials performed using the same odor, although different odors produced different spatial patterns of the modulation. From these results, we conclude that in the PC lobe learned odors are represented as spatial and temporal activity patterns of oscillators that constitute a coherent network. PMID- 10701877 TI - Mapping of interneurons that contribute to food aversive conditioning in the slug brain. AB - To determine the distribution of neurons that contribute to memory formation induced by odor-taste associative conditioning in the slug's brain, we examined neuronal activity of the central nervous system of the slug Limax marginatus using a fluorescent activity marker [Lucifer yellow (LY)]. When LY was injected into the body cavity just after the conditioning, many of the procerebral (PC) interneurons were labeled. The PC lobe was considered to play important roles in the olfaction of the slug, because the olfactory afferent fibers from both the inferior and the superior tentacular noses innervate it. Such strong dye-uptake activity of PC interneurons was not observed when LY was injected just after unpaired control treatment. Thus, it was suggested that enhancement of dye-uptake activity upon conditioning was caused by the association of a conditioning stimulus (CS) with an unconditioned stimulus (UCS). The distribution patterns of PC interneurons that were labeled by LY after conditioning showed a characteristic feature: They usually formed a belt-shaped cluster parallel to the dorsoventral axis. This feature of the distribution was maintained when different odors were used as a CS. Furthermore, the number of the clusters reflected the number of CS odors but not the number of conditioning sessions. From these observations, we considered that enhancement of the neural activity involving dye uptake in each belt-shaped cluster contributed to formation of each odor memory. PMID- 10701879 TI - Two critical periods of protein and glycoprotein synthesis in memory consolidation for visual categorization learning in chicks. AB - A protein synthesis inhibitor, anisomycin (ANI), and an inhibitor of glycoprotein synthesis, 2-deoxygalactose (2-D-gal), were used to investigate memory consolidation following visual categorization training in 2-day-old chicks. ANI (0.6 micromole/chick) and 2-D-gal (40 micromoles/chick) were injected intracerebrally at different time intervals from 1 hr before to 23 hr after the training. Retention was tested 24 hr post-training. Both ANI and 2-D-gal injections revealed two periods of memory sensitivity to pharmacological intervention. ANI impaired retention when injected from 5 min before to 30 min after the training or from 4 hr to 5 hr post-training, thus demonstrating that consolidation of long-term memory in this task requires two periods of protein synthesis. 2-D-Gal first produced an amnesia when it was injected in the interval from 5 min before to 5 min after the training. Injections made between 5 min and 5 hr post-training were without effect on the retention. The second period of memory impairment by 2-D-gal started at 5 hr post-training and lasted until 21 hr after the training. Administration of 2-D-gal made 23 hr after the training did not influence retention in the test at either 24 hr or 26 hr. These results are consistent with the hypothesis that two waves of protein and glycoprotein synthesis are necessary for the formation of long-term memory. The prolonged duration of performance impairment by 2-D-gal in the present task might reflect an extended memory consolidation period for a categorization form of learning. PMID- 10701880 TI - Deficit in learning of a motor skill requiring strategy, but not of perceptuomotor recalibration, with aging. AB - We investigated the effect of aging on different aspects of motor skill learning using two computer-presented perceptuomotor tasks. The relationship between visual and proprioceptive feedback was transformed in the first task, which was open to the formation and use of strategies. This task was designed to lead to perceptuomotor adaptation that was then measured by performance on a very similar second task that was not open to the use of strategy task. Older participants showed impaired learning of the strategic task but not of the nonstrategic task. This is in line with the suggestion that the effect of aging on learning and memory may be to reduce working memory resources. PMID- 10701881 TI - A model of cerebellar metaplasticity. AB - The term "learning rule" in neural network theory usually refers to a rule for the plasticity of a given synapse, whereas metaplasticity involves a "metalearning algorithm" describing higher level control mechanisms for apportioning plasticity across a population of synapses. We propose here that the cerebellar cortex may use metaplasticity, and we demonstrate this by introducing the Cerebellar Adaptive Rate Learning (CARL) algorithm that concentrates learning on those Purkinje cell synapses whose adaptation is most relevant to learning an overall pattern. Our results show that this biologically plausible metalearning algorithm not only improves significantly the learning capability of the cerebellum but is very robust. Finally, we identify several putative neurochemicals that could be involved in a cascade of events leading to adaptive learning rates in Purkinje cell synapses. PMID- 10701882 TI - Backward inhibitory learning in honeybees: a behavioral analysis of reinforcement processing. AB - One class of theoretical accounts of associative learning suggests that reinforcers are processed according to learning rules that minimize the predictive error between the expected strength of future reinforcement and its actual strength. The omission of reinforcement in a situation where it is expected leads to inhibitory learning of stimuli indicative for such a violation of the prediction. There are, however, results indicating that inhibitory learning can also be induced by other mechanisms. Here, we present data from olfactory reward conditioning in honeybees that show that (1) one- and multiple trial backward conditioning results in conditioned inhibition (CI); (2) the inhibition is maximal for a 15-sec interval between US and CS; (3) there is a nonmonotonic dependency on the degree of CI from the US-CS interval during backward pairing; and (4) the prior association of context stimuli with reinforcement is not necessary for the development of CI. These results cannot be explained by models that only minimize a prediction error. Rather, they are consistent with models of associative learning that, in addition, assume that learning depends on the temporal overlap of a CS with two processes, a fast excitatory and a slow inhibitory one, both evoked by a reinforcer. The fmdings from this behavioral analysis of reinforcement processing are compared with the known properties of an individual, identified neuron involved in reinforcement processing in the bee brain, to further understand the mechanisms underlying predictive reward learning. PMID- 10701883 TI - Conservative treatment of childhood phimosis with topical conjugated equine estrogen ointment. AB - PURPOSE: To assess the application of topical conjugated equine estrogen for the treatment of boys with phimosis. METHODS: Fifteen boys with phimosis were included in the study. Conjugated equine estrogen (Premarin) 0.1% ointment was applied on the prepuce once daily. The treatment was continued until the prepuce was fully retractable. The patient was examined each second week up to a maximum treatment of 8 weeks. Retractability and the appearance of the foreskin were graded before and after treatment. RESULTS: Thirteen of 15 boys (87%) referred with phimosis were successfully treated with conjugated equine estrogen ointment. An adverse effect of gynecomastia was seen in one boy (7%). CONCLUSION: Conjugated equine estrogen ointment application for phimosis may be an alternative to surgery. PMID- 10701884 TI - Prognostic significance of tumor grade for renal cell carcinoma. AB - BACKGROUND: The natural history and prognosis of renal cell carcinoma cannot be predicted. Based on the Japanese classification system, the value of nuclear grade were assessed as a possible prognostic factor for renal cell carcinomas. METHODS: In this retrospective study of 116 patients with renal cell carcinoma, radical nephrectomy was performed. Survival rates were calculated using the Kaplan-Meier method and multivariate analysis was performed using Cox's proportional hazard model. RESULTS: Distribution by stage and grade in the population of renal cell carcinomas was as follows: pT1 in 13 cases (11.3%), pT2 in 65 cases (56.5%), pT3 in 36 cases (31.3%) and pT4 in one case (0.9%) and grade 1, 28 (24.1%), grade 2, 69 (59.5%) and grade 3, 16 (13.8%). Three cases could not be determined because of pre-operative embolization of the renal cell carcinomas. Nuclear grade was correlated with stage (P=0.0002), the presence of perirenal fat involvement (P=0.003) and metastases (P=0.007). A significant difference in survival was found between grades 1 and 3 (P=0.0001) and grades 2 and 3 (P=0.0001), respectively. Survival was significantly correlated with sex (P=0.0125), tumor size (P=0.0001), the presence of lymph node metastasis (P=0.0001), renal vein involvement (P=0.0001), perirenal fat involvement (P=0.002) or distant metastasis (P=0.0001). The multivariate analysis showed that the occurrence of tumor grade (P=0.0006) or distant metastasis were independent prognostic values. CONCLUSION: The observations lead us to conclude that the nuclear grade according to the Japanese classification system appears to be of reliable prognostic value for renal cell carcinomas. PMID- 10701885 TI - Endocrine cell distribution and expression of tissue-associated antigens in human female paraurethral duct: possible clue to the origin of urethral diverticular cancer. AB - BACKGROUND: To investigate (i) what determines the histologic differences seen among female urethral diverticular cancers and (ii) the possible embryologic origin of the female paraurethral duct, we performed a distribution analysis of endocrine cells and a comparative study of tissue-associated antigens in the female paraurethral duct. METHODS: Six human female urethras were obtained from surgical and autopsy cases including two cases of urethral diverticular cancer (columnar/mucinous type adenocarcinoma). The urethral and paraurethral epithelia were examined histologically and immunohistochemically. RESULTS: Immunoreactive endocrine cells predominated and carcinoembryonic antigen (CEA) was strongly expressed in the larger portion of the paraurethral duct close to the urethral lumen. Conversely, prostate-specific antigen and prostatic acid phosphatase were positive only in the smaller distal duct. In two cases of adenocarcinoma including endocrine cells, cancer cells were strongly positive for CEA. CONCLUSIONS: This study suggests that the proximal and distal parts of the paraurethral duct have different histologic characteristics and that the pathologic differences seen among female diverticular cancers may result from their cancer-genesis from different parts of the paraurethral duct. PMID- 10701886 TI - Telomerase activity in human renal cell carcinoma. AB - PURPOSE: Telomerase activity has been detected in a wide variety of human tumors. The present study evaluated telomerase activity in association with the acquisition of renal cell carcinoma (RCC). METHODS: Telomerase activity was examined in 30 RCC and the adjacent normal kidney tissue, obtained as surgical specimens. The activity was assayed by polymerase chain reaction-based telomeric repeat amplification protocol assay. RESULTS: Among the 30 RCC, 18 (60%) displayed telomerase activity, whereas none of the normal tissue samples exhibited it. Subdivision of the tumors according to telomerase activity did not reveal any obvious difference in distribution with regard to tumor size, stage, histocytological subtype, or DNA-ploidy. However, a statistically significant relationship was found between the frequency of telomerase-positive activity and both serum immunosuppressive acidic protein level in the patient and tumor grade (P<0.05). There was no significant difference in the recurrent-free survival and the disease-specific survival between patients with positive telomerase activity and patients with negative activity. CONCLUSION: The present results indicate that telomerase activity might be related to the progression of RCC and thus a marker of malignant potential. PMID- 10701887 TI - A case of incidentally detected Castleman's disease with retroperitoneal paravertebral localization. AB - Castleman's disease, or angiofollicular lymph node hyperplasia, is a fairly rare benign tumor of lymphoid origin. The expected localization is mediastinum and rarely retroperitoneum. An asymptomatic case is reported with multimodality imaging and postoperative findings. The lesion was detected incidentally on routine chest radiogram. Surgical exploration revealed a retroperitoneal mass and the mass was resected successfully through a thoracoabdominal flank incision. Castleman's disease should be included in the list of differential diagnosis of retroperitoneal masses which are mostly malignant tumors. PMID- 10701888 TI - Enterocolitis with pathogenic Escherichia coli infection in renal transplant recipients: case reports. AB - PURPOSE: We report three cases of enterocolitis associated with pathogenic Escherichia coli infection in renal transplant recipients. METHODS/RESULTS: Patients presented with abdominal pain and diarrhea at 1, 3 and 7 years after living-related renal transplantation. Pathogens of enterocolitis were identified by stool culture as verotoxin-2-producing O157:H7 E. coli, non-verotoxin producing E. coli 06 and 0125. All patients were basically treated with fluid replacement with additional fosfomycin administration in the patient with O157:H7 E. coli infection. Immunosuppressive drugs were kept at maintenance doses throughout the treatment. CONCLUSION: All patients recovered uneventfully within 10 days after the onset of enterocolitis without severe complications. PMID- 10701889 TI - Antiandrogens fail to block androstenedione-mediated mutated androgen receptor transactivation in human prostate cancer cells. PMID- 10701890 TI - Howard Henry Tooth (1856-1925). PMID- 10701891 TI - Diagnosis and management of normal-pressure hydrocephalus. AB - The syndrome of normal-pressure hydrocephalus (NPH) remains a diagnostic and therapeutic challenge, especially as many patients do not display the classical clinical and neuroimaging patterns of NPH, thus questioning the usefulness of a shunt. Gait impairment remains the cardinal symptom, while mental deterioration may be subtle and even unrecognized. NPH is rarely the cause of severe dementia, and substantial improvement in NPH-related mental deterioration is limited to 30 40% of shunted patients. Many ancillary investigations have been described that can increase the probability of selecting the appropriate candidates for a shunt. The reliability and reproducibility of these tests are limited. Unfortunately, the best predictive tests are technically complex and are used only in a few specialized centers. The best management is still to adhere to strict clinical and magnetic resonance imaging criteria and to rely on a positive - but not negative - CSF tap test and the occurrence of B-waves during at least 50% of the continuous intracranial pressure recording time, when this procedure is available. PMID- 10701892 TI - Mortality in epilepsy. AB - Both community-based studies and reports from more selected epilepsy populations consistently reveal persons with epilepsy to have a mortality rate two to three times that of the general population. This increased rate is most pronounced in patients with remote symptomatic epilepsy, although many studies also report a significant excess mortality also among those with idiopathic epilepsy. The highest standardized mortality ratios (SMR) are seen in young age groups, mainly due to the low expected mortality in children, and during the first 5-10 years after diagnosis. Many of these observations suggest that the higher mortality is partly related to the underlying disorder causing epilepsy rather than a direct consequence of the seizures. For example, mortality in cerebrovascular diseases is increased, with SMRs ranging from 1.8 to 5.3 in the various studies. Deaths due to neoplasms, and in particular brain tumors, is also increased among patients with epilepsy. Accidents, status epilepticus, and sudden unexpected death (SUD) are more directly seizure-related causes of death. The incidence of such deaths vary considerably depending on the population being studied. While rare among patients with new-onset epilepsy, seizure-related deaths may account for up to 40% of all deaths in patients with chronic epilepsy. SUD is probably the most frequent seizure-related cause of death among young adults with epilepsy, with an SMR more than 20 compared with the general population. Seizure induced autonomic cardiorespiratory effects have been suggested, but the mechanisms behind SUD are far from fully understood. It appears, however, that the risk of SUD is closely related to seizure frequency, being 40 times higher in patients who continue to have seizures than in those who are seizure-free. PMID- 10701894 TI - The contribution of fast-FLAIR MRI for lesion detection in the brain of patients with systemic autoimmune diseases. AB - Fast fluid-attenuated inversion recovery (fFLAIR) is more sensitive that conventional or fast spin echo T2-weighted magnetic resonance imaging (MRI) for detecting lesions in the brain of patients with ischemic, inflammatory, or demyelinating diseases of the CNS. We investigated whether the use of fFLAIR also increases the sensitivity of brain MRI assessment in patients with systemic autoimmune disorders. Turbo spin echo (TSE) dual-echo and fFLAIR scans of the brain were obtained from patients affected by systemic lupus erythematosus (SLE) with (NSLE, n = 9) and without clinical CNS involvement (n = 15), Behcet disease (n = 5), Wegener granulomatosis (n = 9), and antiphospholipid antibody syndrome (n = 6). Brain hyperintense lesions were counted and classified according to their size and their location by two observers by consensual agreement. The total lesion volume was measured using a semiautomated technique for lesion segmentation on both TSE and fFLAIR scans. The imaging modalities showed brain hyperintense lesions in all 9 SLE patients with CNS involvement, 5 of 15 SLE patients without CNS involvement, 5 of 9 patients with Wegener granulomatosis, 1 of 5 with Behcet disease, and 3 of 6 with antiphospholipid antibody syndrome. A total of 342 lesions were seen on both sequences; 88 were seen only on TSE and 54 only on fFLAIR scans. The average number of brain lesions per scan was higher on TSE than on fFLAIR, since significantly more discrete (P<0.002) and small (P = 0.004) lesions were seen on TSE than on fFLAIR. The median total lesion volume, however, was similar on TSE and fFLAIR. Our study indicates that the use of fFLAIR does not improve the sensitivity of fast dual-echo MRI for detecting brain abnormalities in patients with systemic autoimmune disorders. PMID- 10701893 TI - High-dose immunoglobulin therapy in sporadic inclusion body myositis: a double blind, placebo-controlled study. AB - Sporadic inclusion body myositis (s-IBM) is an acquired inflammatory muscle disease of unknown cause. In general, s-IBM presents with slowly progressive, asymmetric weakness, and atrophy of skeletal muscle. There is a mild transitory or nil responsiveness to standard immunosuppressive treatment. A controlled cross over study of 22 s-IBM patients over 3 months showed a partial improvement in those treated with high-dose intravenous immunoglobulin therapy (IVIG) versus placebo. The present study included 22 patients aged 32-75 years and with a mean duration of disease of 5.2+/-3.6 years. They were randomized by a double-blind, placebo-controlled, cross-over design to monthly infusions of 2 g/kg bodyweight IVIG or to placebo for 6 months each, followed by the alternative treatment. After 6 and 12 months the response to treatment was evaluated, using a modified Medical Research Council scale, Neuromuscular Symptom Score (NSS), the patient's own assessment of improvement, arm outstretched time, and electromyography. No serious side effects were seen, in particular no viral infection and no major cardiac or neurological complications. Overall there was no progression of the disease in 90% of patients, unlike that which might have been expected in untreated patients. A mild and significant improvement (11%) in clinical symptoms was found using NSS, but not with other test procedures. There was a trend to mild improvement in treated patients when using other tests. Individual responses to treatment was heterogeneous. The validity of this study may be reduced by mismatch of groups with regard to age at onset and variability in disease expression. The findings of this study largely confirm those of a previous IVIG trial. Treatment with IVIG may be mildly effective in s-IBM by preventing disease progression or inducing mild improvement. Long-term studies are needed to evaluate further the benefit of IVIG therapy in s-IBM. PMID- 10701895 TI - The use of magnetic resonance imaging in multiple sclerosis treatment trials: power calculations for annual lesion load measurement. AB - Phase III definitive treatment trials of new multiple sclerosis (MS) therapies now routinely incorporate an annual magnetic resonance imaging protocol, with change in T2-weighted brain lesion load providing an important outcome measure. To date the accepted strategy has been to perform a core imaging protocol on all patients in such studies. The aim of this study was to provide power calculations based on this MRI endpoint. Serial MRI data from 128 patients with either relapsing remitting (RR) or secondary progressive (SP) MS were used to calculate sample size requirements using a repeated measures analysis of variance design. We provide sample size calculations based on various follow-up intervals and effect sizes. Sample sizes for the SPMS cohort were substantially larger than for the RRMS group, reflecting the greater variance in lesion load changes between patients in the SPMS group. With a follow-up of 3 years, we estimate that only 12 and 33 patients per arm are needed to show stabilisation of MRI lesion load in the RRMS and SPMS groups, respectively. Our results suggest that ongoing phase III treatment trials are more than adequately powered to detect even subtle treatment effects, and indicate that incorporating measurements from longer follow-up durations increases power substantially. We conclude that an annual imaging protocol provides a robust and powerful tool for assessing effects on the radiological appearance of the disease process. PMID- 10701896 TI - Sleep-disordered breathing among patients with first-ever stroke. AB - Sleep-disordered breathing (SDB) in the form of obstructive sleep apnea is a possible risk factor for stroke. We carried out a cross-sectional survey out in a rehabilitation center among patients with first-ever stroke to further determine the incidence and types of SDB and its relationship to known risk factors for stroke. Full polysomnography was performed in 147 consecutive patients (95 men, 52 women, age 61+/-10 years) admitted to our neurological Rehabilitation Department 46+/-20 days after first-ever stroke. Subjective sleepiness (Epworth Sleepiness Scale), vascular risk factors, anthropometric data, and polysomnographic findings were compared between stroke patients with varying degrees of SDB. With a cutoff point for the respiratory disturbance index (RDI) of 5, 10, 15, or 20 the respective prevalence of SDB was 61%, 44%, 32%, and 22%. The type of SDB was generally obstructive, with dominant central apneas in only 6% of patients. Patients with an RDI of 20 or higher had less REM sleep, thicker necks, and a more central type of obesity. Even in patients with an RDI of 20 or higher subjective sleepiness, although higher than in those without SDB, was not a predominant symptom. Snoring and anthropometric data suggest that obstructive SDB may have existed prior to stroke. The prevalence of hypertension and coronary heart disease were higher among stroke patients with an RDI of 20 or higher than in those without SDB. We conclude that the prevalence of SDB among patients with stroke is high. Examination of stroke should include screening for SDB. PMID- 10701897 TI - Thalamic degeneration with negative prion protein immunostaining. AB - A 34-year-old woman presented with an insidious 5-year history of cognitive decline and apathy, associated with hypersomnia, ataxia, and dysarthria. Magnetic resonance imaging of the brain showed cortical and subcortical atrophy. At autopsy we found abnormalities in the subcortical grey matter and brainstem, with a relatively preserved cerebral cortex. The thalami showed symmetrical neuronal loss and astrocytosis, particularly severe in the dorsal medial nucleus, followed by the lateral nuclei group. Prion protein immunostaining was negative, and there was no spongiform change. No mutations were detected in the prion protein gene. PMID- 10701898 TI - The monoamine oxidase B gene GT repeat polymorphism and Parkinson's disease in a Chinese population. AB - Monoamine oxidase B (MAOB) metabolises dopamine and activates neurotoxins known to induce parkinsonism in humans and primates. Therefore the MAOB gene (MAOB; Xp15.21-4) is a candidate gene for Parkinson's disease (PD). Longer length dinucleotide repeat sequences in a highly polymorphic GT repeat region of intron 2 of this gene showed an association with PD in an Australian cohort. We repeated this allele-association study in a population of 176 Chinese PD patients (90 men, 86 women) and 203 agematched controls (99 men, 104 women). Genomic DNA was extracted from venous blood and the polymerase chain reaction was used to amplify the appropriate regions of the MAOB gene. The length of each (GT) repeat sequence was determined by 5% polyacrylamide denaturing gel electrophoresis. There was no significant difference in allele frequencies of the (GT) repeat allelic variation between patients and controls (chi2 = 2.48; df = 5, P<0.75). Therefore the longer length GT repeat alleles are not associated with PD in this Chinese population. Possible reasons for the discrepancy between Chinese and Australian populations include a different interaction between this genetic factor and environmental factors in the two populations and the possibility that the long length GT repeat alleles may represent a marker mutation, genetically linked to another susceptibility allele in whites but not in Chinese. Methodological differences in the ascertainment of cases and controls in this cohort could also explain the observed differences. Further study is required to determine whether the longer length GT repeat alleles are true susceptibility alleles in PD. PMID- 10701899 TI - Omeprazole-induced delirium. PMID- 10701900 TI - Hyalinosis cutis et mucosae: a case report. PMID- 10701901 TI - Myasthenia gravis associated with limited scleroderma (CREST syndrome) PMID- 10701902 TI - CNS involvement in primary Sjogren's syndrome: a case with a clue for the pathogenesis. PMID- 10701903 TI - Leber's hereditary optic neuropathy presenting as multiple sclerosis-like disease of the CNS. PMID- 10701904 TI - Peroneal nerve compression by fibrous histiocytoma. PMID- 10701905 TI - I. Acquisition of social knowledge is related to the prefrontal cortex. PMID- 10701906 TI - II. Regional distribution in the brain of amines associated with Parkinson's disease. PMID- 10701907 TI - Investigation of the chemical equivalence of the trypanocidal products, Samorin and Veridium. AB - A procedure for the evaluation of chemical equivalence of proprietary formulations of isometamidium is described. The method combines the analysis of the principal component (isometamidium), HPLC profiling of related substances and determination of the inorganic impurity, ammonium chloride, using a modification of the Berthelot (Indophenol) reaction. Application of these procedures to analyses of commercially available sachets from four different batches of Samorin and four different batches of Veridium has demonstrated that there are marked qualitative and quantitative differences between batches from these two sources. Whilst Samorin samples showed inter-batch consistency of composition, there was considerable inter-batch variation between the samples of Veridium. PMID- 10701908 TI - Amoxicillin sodium-potassium clavulanate: evaluation of gamma radiation induced effects by liquid chromatography on both the individual drugs and their combination. AB - The effects of gamma irradiation on the stability of potassium clavulanate, amoxicillin sodium and their combination were investigated. A decrease in purity and increase in degradation products up to 30 days after the irradiation were evaluated by reversed phase HPLC. The comparison between unirradiated and irradiated amoxicillin sodium, performed within 24 h following the irradiation process, showed no significant increase in the pre-existing impurities and no evidence of newly induced degradation products. On the contrary, an appreciable increase in the content of some impurities was evidenced 30 days after the irradiation. The chromatographic profile of irradiated potassium clavulanate showed the appearance of one unidentified new product and a slight increase of one pre-existing impurity. No further change in the impurity content was noted 30 days after the irradiation. The amoxicillin sodium-potassium clavulanate combination underwent the same kind of radiation induced degradation as the single compounds. PMID- 10701909 TI - Simultaneous assay of ephedrine hydrochloride, theophylline, papaverine hydrochloride and hydroxyzine hydrochloride in tablets using RP-LC. AB - The analytical problem was to control the quality of imported antiasthmatic tablets containing ephedrine hydrochloride, theophylline, papaverine hydrochloride and hydroxyzine hydrochloride. The aim of the analytical method for the assay was to separate, identify and quantify all compounds, at the same time. A gradient capable RP-LC system was used, using a commercially packed Nucleosil C18 column connected to a dual channel variable, programmable wavelength detector. The analysis was performed in the gradient program of increasing concentration of acetonitrile in water. The influence of pH of the mobile phase was established. The proposed method is reliable, reproducible, easy to perform and satisfies the aim. PMID- 10701910 TI - Evaluation of the H-point standard additions method (HPSAM) and the generalized H point standard additions method (GHPSAM) for the UV-analysis of two-component mixtures. AB - The H-point standard additions method (HPSAM) and two versions of the generalized H-point standard additions method (GHPSAM) are evaluated for the UV-analysis of two-component mixtures. Synthetic mixtures of anhydrous caffeine and phenazone as well as of atovaquone and proguanil hydrochloride were used. Furthermore, the method was applied to pharmaceutical formulations that contain these compounds as active drug substances. This paper shows both the difficulties that are related to the methods and the conditions by which acceptable results can be obtained. PMID- 10701911 TI - Enantiometric separation of verapamil and norverapamil using Chiral-AGP as the stationary phase. AB - Simultaneous enantiomeric separation of verapamil and its main metabolite norverapamil was achieved using Chiral-AGP as the stationary phase. The optimized chromatographic system was obtained using statistical experimental design with partial least squares as regression method. The three variables studied were buffer pH, content of acetonitrile and column temperature. A high buffer pH favors enantioselectivity as well as the selectivity between (S)-verapamil and (R)-norverapamil. The concentration of the organic modifier in the mobile phase was a compromise as a high content of acetonitrile decreased enantioselectivity but increased the selectivity mentioned above. Increased column temperature increased the separation between (S)-verapamil and (R)-norverapamil with only a slight decrease in enantioresolution. PMID- 10701912 TI - Urea determination using pH-enzyme electrode. AB - A pH-membrane electrode with n-tridodecylamine (TDDA) as the hydrogen-ion selective ionophore was used for the construction of a potentiometric biosensor for urea determination. The electrode was enzymatically modified by covalent binding of urease molecules directly to the surface of the potentiometric membrane. Incorporation of the urea biosensor into simple double-channel flow injection analysis (FIA) system allows reproducible urea determination in a millimolar range of concentration. The utility and limitations of the presented biosensor-FIA system for analysis of various real samples has been investigated. The system can be useful for some biomedical and pharmaceutical applications such as analyses of urine, posthaemodialysis fluid and extracts from pharmaceutical ointments containing urea. PMID- 10701913 TI - Narrowbore high performance liquid chromatography of berberine and palmatine in crude drugs and pharmaceuticals with ion-pair extraction using cobalt thiocyanate reagent. AB - For the simultaneous determination of berberine and palmatine from Phellodendri Cortex, Coptidis Rhizoma and pharmaceuticals, a new narrowbore HPLC method was developed with a simple and selective sample clean-up using cobalt thiocyanate reagent. Samples were sonicated with 2% hydrochloric acid for 30 min and protoberberine-type alkaloids in the resulting mixture were extracted with cobalt thiocyanate reagent and dichloroethane. The aliquot of dichloroethane layer was evaporated and the residue was dissolved for HPLC analysis. The recoveries of berberine and palmatine from Phellodendri Cortex, and Coptidis Rhizoma were better than 90%. Calibration curves for berberine and palmatine were linear over the concentration range of 0.1-50 microg/ml. Limits of quantitation for berberine and palmatine were 0.5 ng. This sample preparation process can be used for the identification of protoberberine-type alkaloids from crude drugs and oriental pharmaceutical preparations. PMID- 10701914 TI - Determination of captopril and its degradation products by capillary electrophoresis. AB - Captopril, an antihypertensive agent, and its degradation products have been quantified in pharmaceutical formulations by capillary zone electrophoresis (CZE). A method using cetyltrimethylammonium bromide (CTAB) added to a sodium phosphate buffer (pH 5.5; 100 mM) as running buffer and using N-acetyl-L-tyrosine as an internal standard has been developed and validated for the quantitative determination of captopril in tablets. The method is an indicator of compound stability and can also be applied to the purity control of the raw material and for the determination of the degradation products. For this purpose, salicylic acid is used as an internal standard. PMID- 10701915 TI - Chiral separation and quantitation of pentazocine enantiomers in pharmaceuticals by capillary zone electrophoresis using maltodextrins. AB - The chiral separation of pentazocine was achieved by capillary electrophoresis using oligosaccharides. Enantiomers were separated on 100 mM Tris/H3PO4 buffer (pH 2.5) with 5% maltodextrin as a chiral selector, and migration behavior was monitored at 200 nm. Under these conditions, (-)- and (+)-pentazocine and dextromethorphan (internal standard) migrated within 9 min, and the resolution of pentazocine enantiomers was 2.54. Linear calibration curves were obtained in the range 5-50 microg/ml(-1) for each enantiomer. The detection limit of pentazocine enantiomers was 29 pg, and the recoveries of(-)- and (+)-pentazocine were 98.9 (R.S.D., 3.4%) and 101.4% (R.S.D., 4.3%) with 10 microg/ml(-1), respectively. PMID- 10701916 TI - Analysis of diclofenac sodium and derivatives. AB - There are two reasons explaining why several researchers have carried out the in vitro release studies of diclofenac sodium (DFNa) using pH media of above 6.5. Firstly the pH dependence of solubility, and secondly the intramolecular cyclization suffered under acidic conditions which causes the salt to become inactivated. Nevertheless, many commercially available pharmaceutical dosage forms have no protective coat to avoid the inactivation in the gastric juices. A possible explanation may be found if reconstitution of the cyclated form takes place. It is therefore necessary to study the behaviour of diclofenac sodium when it is submitted to the action of different solutions in a wide pH range. To perform this study five analytical methods have been employed: UV-vis spectrophotometry, differential scanning calorimetry (DSC), infrared analysis (IR), X-ray diffractometry (DRX) and energy dispersive X-ray analysis (EDS). PMID- 10701917 TI - Use of ANN modelling in structure--retention relationships of diuretics in RP HPLC. AB - Structure retention relationship study, conducted by RP HPLC, was used to investigate physical chemical parameters related to the RP retention times of amiloride, hydrochlorothiazide and methyldopa in order to predict the separation of amiloride and methylclothiazide from Lometazid tablets. Retention data were obtained with an ODS column using a mobile phase methanol water (pH adjusted with phosphoric acid). Physical chemical properties were calculated directly from the molecular structure. Artificial neural networks (ANNs) were used to correlate chromatograms retention times with mobile phase composition and pH, and with physical chemical properties of amiloride, hydrochlorothiazide and methyldopa and to predict separation of amiloride and methylclothiazide from Lometazid tablets. Sensitivity analysis was performed to interpret the meaning of the descriptors included in the models. Results confirmed the dominant role of the polar modifier in such chromatographic systems. Within a series of solutes chromatographed under identical conditions, the retention parameters could be approximated by a non linear combination of logP, logD, pKa, surface tension, parachor, molar volume and to minor extend by polarisability, reetractivity index and density. This study has demonstrated that the use ANNs techniques can result in much more efficient use of experimental information. As HPLC is the most popular analytical technique, improvements in HPLC methods development can yield significant gains in the overall analytical effort. The ANNs extension presented could be the method of choice in some advanced research settings and serves as an indication of the broad potential of neural networks in chromatography analysis. PMID- 10701918 TI - The determination of methanesulphonic acid content of busulfan drug substance and busulfan (Myleran) tablets by ion chromatography. AB - A robust ion chromatographic procedure is described for the determination of the methanesulphonic acid content of busulfan drug substance and busulfan (Myleran) 2 mg tablets. The sample was dissolved in aqueous acetonitrile, butanesulphonic acid was added as an internal standard and the solution was injected onto an ion selective column, with ion suppression and conductivity detection. The method has been fully validated and is linear over the concentration range 0.295-14.73 microg of methanesulphonic acid/ml. The method has been demonstrated to be stability indicating. PMID- 10701919 TI - The influence of data pre-processing in the pattern recognition of excipients near-infrared spectra. AB - The effect of data pre-processing (no pre-processing, offset correction, de trending, standard normal variate transformation (SNV), SNV + de-trending, multiplicative scatter correction, first and second derivative transformation after smoothing) on the identification of ten pharmaceutical excipients is investigated. Four pattern recognition methods are tested in the study, namely the Mahalanobis distance method, the SIMCA residual variance method, the wavelength distance method and a method based on triangular potential functions. The performance of the 32 method combinations is evaluated on the basis of two NIR data sets. The first one, measured in 1994, is used to build the classification models, the second, measured from 1994-1997, is used to assess the quality of the models. The best approach for the given data sets is the wavelength distance method combined with de-trending, a simple baseline correction method. More general recommendations for pre-processing excipient NIR data and for choosing an appropriate classification method are given. PMID- 10701920 TI - Spectrophotometric determination of fluoxetine and sertraline using chloranil, 2, 3 dichloro-5, 6 dicyano benzoquinone and iodine. AB - Spectrophotometric procedures are presented for the determination of two commonly used antidepressant drugs, fluoxetine (I) and sertraline hydrochloride (II). The methods are based mainly on charge transfer complexation reaction of these drugs with either pi acceptors chloranil and 2, 3 dichloro-5, 6-dicyanoquinone (DDQ) or sigma acceptor iodine. The colored products are quantified spectrophotometrically at 550, 450 and 263 nm for fluoxetine and at 450, 455 and 290 nm for sertraline in chloranil, DDQ and iodine methods, respectively. The molar combining ratio and the optimum assay conditions were studied. The methods determine the cited drugs in concentration ranges of 8-640, 16-112 and 7.5-60 microg/ml with mean percentage recoveries of 99.83, 99.76 and 100.00% and R.S.D. of 1.24, 0.95 and 1.13% in fluoxetine and ranges of 16-160, 15-105 and 6-48 microg/ml with mean percentage recoveries of 100.39, 99.78 and 99.69% and R.S.D. of 1.02, 0.81 and 0.57% in sertraline for chloranil, DDQ and iodine methods, respectively. A more detailed investigation of the complex formed was made with respect to its composition, association constant K(AD)c, molar absorptivity xiAD(A) and free energy change deltaG. The proposed methods were applied successfully to the determination of the cited drugs either in pure or dosage forms with good accuracy and precision. The results were compared statistically with those given by the reported methods. PMID- 10701921 TI - LC method development for ibuprophen and validation in different pharmaceuticals. AB - An isocratic LC method for the simultaneous determination of ibuprophen (IBU) and its degradation product (4-isobutylacetophenone) has been developed and validated for different pharmaceuticals (sachets, tablets and gels). The chromatographic separation was achieved with phosphoric acid solution (pH 3.2)-acetonitrile (50:50, v/v) as mobile phase, a Hypersil C18 column and UV detection at 254 nm. In all cases, sample preparation was required before HPLC analysis. PMID- 10701922 TI - Determination of the oral platelet aggregation inhibitor Sibrafiban in rat, dog, and human plasma utilising HPLC-column switching combined with turbo ion spray single quadrupole mass spectrometry. AB - A sensitive and selective HPLC-column switching method with single quadrupole mass spectrometric detection was developed for the simultaneous determination of the oral platelet aggregation inhibitor Sibrafiban (double protected prodrug), its prodrug and the active metabolite in rat, dog, and human plasma. The three analytes together with their tri-deuterated internal standards were isolated from plasma by protein precipitation (0.5 M perchloric acid). The de-proteinated samples were injected onto a standard-bore trapping column (4.0 mm i.d., LC-ABZ) of an HPLC-column switching system. Polar plasma components were removed by flushing the trapping column with ammonium formate (pH 3.6; 5 mM). Enriched compounds (including the analytes of interest) were backflushed onto a narrow bore analytical column (2.1 mm i.d., Inertsil ODS-2) and separated by gradient elution (formic acid/ methanol). The whole effluent (200 microl/min) from the analytical column was passed to the turbo ion spray interface without splitting. Selected ion monitoring (SIM) was used for mass spectrometric detection. The limit of quantification for all three analytes was 1 ng/ml, using a 250-microl specimen of plasma. The mean precision and inaccuracy for the three analytes in all species were < 6 and < 5%, respectively. The practicability of the new analytical method was demonstrated by the analysis of about 500 rat and dog plasma and about 14,000 human plasma samples. The new method represents a successful example for the application of LC single MS with ionspray ionisation to the analysis of small molecule drugs in biological matrices from toxicokinetic studies and large clinical trials. PMID- 10701923 TI - Nonaqueous versus aqueous capillary electrophoresis for the dosage of N butylscopolamine in various pharmaceutical formulations. AB - A simple nonaqueous capillary electrophoresis method is described for the separation of several atropine and scopolamine related drugs. The analysis of these pharmaceutical compounds was performed in a methanol-acetonitrile (25/5, v/v) mixture containing 25 mM ammonium acetate and 1 M acetic acid. The robustness was proved using a full factorial design at two levels. The method was validated and successfully applied for the determination of N-butylscopolamine in different pharmaceutical preparations. Results were compared to those obtained by a capillary electrophoresis method based on aqueous media. PMID- 10701924 TI - Determination of nicorandil concentrations in human plasma using liquid chromatography. PMID- 10701925 TI - A method for simultaneous determination of five anticoagulant rodenticides in whole blood by high-performance liquid chromatography. PMID- 10701926 TI - Quantitative determination of ginsenosides from Panax ginseng roots and ginseng preparations by thin layer chromatography--densitometry. PMID- 10701927 TI - Detection and separation of artesunate and artelinic acid with capillary zone electrophoresis. PMID- 10701928 TI - Determination of chloroquine and desethylchloroquine in plasma and blood cells of Plasmodium vivax malaria cases using liquid chromatography. PMID- 10701929 TI - Reversed-phase liquid chromatographic determination of cryptotanshinone and its active metabolite in pig plasma and urine. PMID- 10701930 TI - The study of the voltammetric behaviour of flunarizine. PMID- 10701931 TI - Determination of cisapride in pharmaceutical dosage forms by reversed-phase liquid chromatography. PMID- 10701932 TI - Simplified reversed-phase LC method with spectrophotometric detection for estimation of sparfloxacin in human plasma. PMID- 10701933 TI - Atomic emission spectrometric determination of ephedrine, cinchonine, chlorpheniramine, atropine and diphenhydramine based on formation of ion associates with ammonium reineckate. AB - Ion-associate complexes of ephedrine HCl (I), cinchonine HCl (II), chlorpheniramine maleate (III), atropine sulphate (IV) and diphenhydramine HCl (V) with ammonium reineckate were precipitated and their solubilities were studied as a function of pH, ionic strength and temperature. Saturated solutions of each ion-associate under the optimum precipitation conditions were prepared and the Cr ion content in the supernatant was determined. The solubility products were thus elucidated at different temperatures. A new accurate and precise method using direct current plasma-atomic emission spectrometry for the determination of the investigated drugs in pure solutions and in pharmaceutical preparations is described. The drugs can determined by the present method in the ranges 1.6 52,2.64-85.8,3.12-101.4,5.52-180.4 and 2.72-75.85 microg/ml solutions of I, II, III, IV and V, respectively. PMID- 10701934 TI - Spectrofluorometric determination of alpha-aminocephalosporins in biological fluids and pharmaceutical preparations. AB - A selective and highly sensitive fluorometric method was developed for the determination of four alpha-aminocephalosporins, namely cefaclor, cefadroxil, cephalexin and cephradine. The method involves the reaction of the target compounds with fluorescamine at a specific pH, ranging from 7.8 to 8.4. The produced derivatives exhibit maximum fluorescence intensities at 472-478 nm after excitation at 370-372 nm. The method is highly specific because other alpha aminocephalosporins whose alpha amino group was blocked do not react similarly and hence do not interfere. At the specific pH range of the reaction where no degradation may occur with that medium the proposed method can be utilised as a stability-indicating assay. The different experimental parameters affecting the derivatisation reaction were carefully studied and incorporated into the procedure. Under the described conditions, the proposed method is linear over the concentration range of 0.05(-1) microg/ml(-1) for both cefaclor and cephalexin, and 0.05-0.65 and 0.025-0.5 microg/ml(-1) for cefadroxil and cepharadine, respectively and the coefficients of determination were greater than 0.999 (n = 3). The recoveries of the title compounds from spiked serum ranged from 88.6 to 89.7% and from spiked urine from 92.2 to 93.3% with a limit of quantitation (LOQ) of 25-50 ng/ml(-1) and limit of detection (LOD) of 5 ng/ml(-1) (S/N = 2) for all drugs. The mechanism of the fluorometric reaction is proposed and the advantages of the proposed method are discussed. PMID- 10701935 TI - Flow-injection extraction-spectrophotometric method for the determination of chlorhexidine in pharmaceutical preparations. AB - The spectrophotometric determination of trace amounts of chlorhexidine was carried out by liquid-liquid extraction using bromophenol blue with a flow system. The determination of chlorhexidine in the range of 1 x 10(-4) to 1 x 10( 5) M was possible with a sampling frequency of 40 samples per hour. The method was satisfactorily applied to the determination of chlorhexidine in pharmaceutical preparations. PMID- 10701936 TI - Liquid chromatographic analysis of dihydrostreptomycin sulfate. AB - The analysis of dihydrostreptomycin sulfate using a column packed with base deactivated reversed phase silica gel and ultraviolet detection at 205 nm is described. The mobile phase consists of an aqueous solution containing 4 g/l of sodium sulfate, 1.5 g/l of sodium octanesulfonate, 100 ml/l of acetonitrile and 50 ml/l of a 0.2-M phosphate buffer at pH 3.0. The method allows separation of streptidine, dihydrostreptomycin B, streptomycin, dihydrostreptomycin and deoxydihydrostreptomycin, as well as some other components which were not identified. The total time of analysis is 55 min. The effects of the different chromatographic parameters on the separation were also investigated. A number of commercial samples were analyzed using this method. PMID- 10701937 TI - A comparative study of the ratio spectra derivative spectrophotometry, Vierordt's method and high-performance liquid chromatography applied to the simultaneous analysis of caffeine and paracetamol in tablets. AB - Two spectrophotometric methods and high-performance liquid chromatography were proposed for the simultaneous analysis of caffeine and paracetamol in a tablet formulation. The ratio spectra derivative method is based on the use of the analytical signals obtained by measuring at 267.9 and 291.0 nm for caffeine and 237.0 and 251.8 nm for paracetamol in the first derivative of the ratio spectra. Calibration graphs were prepared in the range 4-40 microg/ml for caffeine and 8 48 microg/ml for paracetamol. In Vierordt's method, A1(1) (1%, 1cm) values of paracetamol and caffeine were determined at 242.9 and 273.0 nm in zero-order spectra. The matrix for A1(1) (1%, 1cm) values was written and the amounts of both drugs were calculated by means of the program 'Matlab' software. The results obtained by these spectrophotometric methods were compared with the results of HPLC method. PMID- 10701938 TI - Flow-injection spectrophotometric methods for the determination of tenoxicam. AB - Two sensitive and rapid flow-injection spectrophotometric methods are proposed for the determination of tenoxicam (TX). In the first method, a Fe(III)-tenoxicam complex is formed in a methanolic medium and the absorbance is measured at 540 nm, while the second method involves measurement of the absorbance at 355 nm of a solution containing the drug in hydrochloric acid medium. In both methods, the peak heights were proportional to tenoxicam concentration over the ranges 7.0-320 and 0.5-8.5 mg/l(-1), respectively. The methods have been applied to the routine determination of the drug in dosage forms. PMID- 10701939 TI - Quantification of amphetamine plasma concentrations by gas chromatography coupled to mass spectrometry. AB - We developed a fast and sensitive method for identification and quantification of plasma concentrations of amphetamine using gas chromatography with mass spectrometry detection (GC-MS). Amphetamine-d8 served as internal standard. The method involves a single extraction procedure and an easy treatment of the samples that allowed no losses during the evaporation process. Derivatisation of amphetamine with N-methyl-bis(trifluoroacetamide), a potent acylating agent, provides many advantages to the method compared with common derivatisation reactions usually used for amphetamines. The limits of detection and quantification following this method were 0.43 and 1.42 ng/ml, respectively. The assay has been successfully employed in the quantification of amphetamine in plasma samples from healthy volunteers at four different doses. PMID- 10701941 TI - Liquid extraction and ion-pair HPLC for determination of hydrophilic 3 hydroxypyridin-4-one iron chelators. AB - Hydrophilic 3-hydroxypyridin-4-ones (HPOs), such as 1-(2'-carboxyethyl)-2-methyl 3-hydroxypyridin-4-one (CP38), 1-(3'-hydroxypropyl)-2-methyl-3-hydroxypyridin-4 one (CP41) and 1-(2'-hydroxyethyl)-2-ethyl-3-hydroxypyridin-4-one (CP102), are orally active iron chelators and ester prodrugs of these molecules are currently under investigation. A liquid extraction method using acetonitrile and 2-propanol (80:20 v/v) under acidic and NaCl-saturated conditions has been developed in order to efficiently extract these HPOs from various matrices. The extracted HPOs were determined using a reversed phase polymer HPLC column (PLRP-S 100 A) and the gradient ion-pair mobile phase containing tetrabutylammonium chloride (5 mM) and EDTA (0.5 mM). The extraction recovery of these chelators in phosphate buffer, rat blood and liver homogenate varied from 85 to 94%. The coefficients of variation (C.V.) for within-day determination were in the range of 1.4-3.3% at 1 mM and 2.0-4.7% at 0.1 mM. High accuracy of determination was also achieved. PMID- 10701940 TI - An improved method for the determination of fluticasone propionate in human plasma. AB - Therapeutic monitoring of the potent, highly lipophilic glucocorticoid, fluticasone propionate (FP), was initially performed by a radioimmunoassay method. However an improved method with a lower limit of quantitation (LLOQ) of at least 25 pg per ml (pg/ml(-1)) was needed to measure the low levels of FP present in human plasma following inhalation administration of doses in the range 50-250 microg twice daily. A sensitive and specific liquid chromatographic, tandem mass spectrometric method (LC-MS/MS) with automated solid phase extraction (SPE) was developed and validated. Fluticasone propionate was extracted from plasma using Bond Elut C18 cartridges and analysed using reverse-phase chromatography with atmospheric pressure chemical ionisation followed by selective reaction monitoring. The method used a 13C-labelled internal standard and was validated over a concentration range of 25-500 pg/ml(-1). The method was shown to be specific, sensitive and reliable in the analysis of clinical samples. The main advantages of this method over the radioimmunoassay method previously used were improved sensitivity, specificity, ease of sample preparation and shortened analysis time. PMID- 10701942 TI - Monosaccharide composition analysis of pamiteplase by anion exchange chromatography with pulsed amperometric detection. AB - The monosaccharides (neutral and amino sugars) of palmiteplase (recombinant modified human tissue plasminogen activator) were analyzed by high performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD). Since the palmiteplase formulation contains sucrose, it was removed by reverse phase high-performance liquid chromatography (RP-HPLC) prior to analysis. Acid hydrolysis with TFA was performed at 100 degrees C for 4 h. Fucose, glucosamine, galactose and mannose were detected by HPAEC-PAD analysis after hydrolysis. The linearity range of HPAEC-PAD analysis was 20-200 pmol/ml(-1) (r > 0.999) and the RSD value for repeatability was less than 7%. The recovery of each monosaccharide spiked into samples was more than 90%. The monosaccharide composition of palmiteplase suggests that it has complex-type oligosaccharides lacking in high mannose-type oligosaccharides. PMID- 10701943 TI - Interaction of substituted phenoxazine chemosensitizers with bovine serum albumin. AB - The binding of 10-[3'-[N-bis(hydroxyethyl)amino]propyl]phenoxazine [BPP], 10-[3' [N-bis(hydroxyethyl)amino]propyl]-2-chlorophenoxazine [BPCP], 10-[3'-[N-bis (hydroxyethyl)amino]propyl]-2-trifluoromethylphenoxazin e [BPFP], 10-(3'-N pyrrolidino propyl)-2-chlorophenoxazine [PPCP] or 10-(3'-N-pyrrolidinopropyl)-2 trifluoromethylphenoxazine [PPFP] to bovine serum albumin (BSA) has been measured by gel filtration and equilibrium dialysis methods. The binding of these modulators to bovine serum albumin based on dialysis experiments has been characterized by the following parameters: percentage (beta) of bound drug, the association constant 'K1', the apparent binding constant 'k' and the free energy deltaFdegrees. The binding of phenoxazine derivatives to bovine serum albumin is correlated with their octanol-water partition coefficient, log10P. In addition, the displacing activity of hydroxyzine and acetylsalicylic acid on the binding of phenoxazines to albumin has been studied. The results of the displacing experiments showed that the phenoxazine benzene rings and the tertiary amines attached to the side chain of the phenoxazine moiety are bound to a hydrophobic area on the albumin molecule. PMID- 10701944 TI - Determination of eletriptan in plasma and saliva using automated sequential trace enrichment of dialysate and high-performance liquid chromatography. AB - The use of the system, automated sequential trace enrichment of dialysates (ASTED), to prepare plasma and saliva prior to high pressure liquid chromatography of eletriptan (HPLC) is described. Chromatographic identification of one metabolite, UK-135,800 was also established. Using this technique the procedure was observed to be specific and linear over the range 0.50-250 ng/ml. The intra-batch imprecision (C.V.) of the method ranged from 0.56 to 5.70% at plasma eletriptan concentrations from 5.00 to 200 ng/ml, and the corresponding inter-batch imprecision ranged from 1.44 to 6.36%. At these plasma analyte concentrations, the overall inaccuracy (% bias) of the procedure ranged from 5.00 to 1.50%. Similar performances were observed for the estimation of eletriptan in saliva using near identical assay conditions. The application of the assay to a pharmacokinetic investigation during a clinical study is presented. PMID- 10701945 TI - Voltammetric and spectrophotometric techniques for the determination of the antihypertensive drug Prazosin in urine and formulations. AB - A sensitive method was developed to determine Prazosin using a nafion modified carbon paste electrode (NMCPE). Prazosin was accumulated at a potential of 750 mV in Britton-Robinson buffer (pH 6.0) and then a negative sweep was made obtaining a cathodic peak close to 0 V. Cyclic voltammetric studies indicated that the process was quasi-reversible, and fundamentally controlled by adsorption. To obtain a good sensitivity, the instrumental and accumulation variables were studied using differential pulse voltammetry (DPV). Adsorptive voltammetric peak currents showed a linear response for Prazosin concentrations in the range between 4.0 x 10(-11) and 4.0 x 10(-8) M with two different slopes, and a detection limit (LOD) of 3.1 x 10(-11)M was obtained. The variation coefficient (CV) for a 8.0 x 10(-10) M solution (n = 10) was 4.08%. A spectrophotometric study of Prazosin was also carried out and two absorption bands were obtained at 246 and 329 nm (pH 1.8). The band at 329 nm was pH-dependent and its height and position changed with the pH values, so this allowed the pK'a determination (7.14 +/- 0.20) using different methods. The detection limit reached by means of UV spectrophotometry was 0.9 x 10(-7) M, and the variation coefficient for 1.5 x 10( 5) M Prazosin solutions was 1.14% (n = 10). Although the sensitivity of the UV spectrophotometric method was lower than that obtained using adsorptive stripping differential pulse voltammetry (AdS-DPV), it could be applied to the determination of Prazosin in Minipres tablets. The voltammetric method was used for the determination of the drug in human urine samples at trace levels with good recoveries. PMID- 10701946 TI - Voltammetric study and determination of buprenorphine in pharmaceuticals. AB - The oxidation of buprenorphine on a carbon paste electrode has been studied using voltammetric techniques under both semi-infinite linear diffusion and hydrodynamic conditions. By applying a simple electrode pretreatment a good reproducibility of the current signal is obtained (R.S.D. = 0.85%, n = 6 for a 1.0 x 10(-5) M buprenorphine concentration). The limit of detection was found to be 2.0 x 10(-7) M. The voltammetric method developed for the determination of buprenorphine in pharmaceutical preparations was examined for its applicability to liquid and solid preparations. PMID- 10701947 TI - Chemiluminescence analysis of menadione sodium bisulfite and analgin in pharmaceutical preparations and biological fluids. AB - A novel chemiluminescence (CL) flow system for two sulfite-containing drugs, namely, menadione sodium bisulfite (MSB) and analgin is described. It is based on the weak chemiluminescence induced by the oxidation of sulfite group in drugs with dissolved oxygen in the presence of acidic Rh6G. Tween 80 surfactant micelles showed a strong enhancement effect on this weak chemiluminescence. For MSB analysis, online conversion of MSB in alkaline medium into sodium bisulfite was necessary, whereas analgin could be determined directly. The proposed method allowed the measurement of 0.05-50 microg/ml(-1) MSB and 0.05-10 microg/ml(-1) analgin. The limits of detection (3sigma) were 0.01 microg/ml(-1) MSB and 0.003 microg/ml(-1) analgin. The method was applied satisfactorily to pharmaceutical preparations as well as biological fluids. PMID- 10701948 TI - Spectrophotometric estimation of D-penicillamine in bulk and dosage forms using 2,6-dichloroquinone-4-chlorimide (DCQ). AB - A simple colorimetric method for the determination of D-penicillamine in pure form and pharmaceutical formulations is described. The method is based on coupling between D-penicillamine and 2,6-dichloroquinone-4-chlorimide (DCQ) in dimethylsulphoxide. The optimum conditions for the reaction were investigated and incorporated into the procedure. The reaction forms a yellow 1:1 complex with maximum absorbance at 431 nm (epsilon = 3,700). Regression analysis of Beer's plot showed good correlation (r = 0.9998) and the calibration graph was rectilinear over the range 4-20 microg/ml(-1) with a detection limit of 0.15 microg/ml(-1) . The average recovery for the commercial capsules was 101.66% with an RSD of 1.57%. The results obtained were sufficiently accurate, reproducible and in accordance with those given by the official method. The method is specific for the intact drug, and can be adopted in the presence of some interfering substances. PMID- 10701949 TI - Rapid determination of theophylline in serum by selective extraction using a heated molecularly imprinted polymer micro-column with differential pulsed elution. AB - Molecular imprinting of theophylline in poly(methacrylic acid ethylene dimethacrylate) form binding sites with complementary size, shape and chemical functionalities to theophylline. This molecularly imprinted polymer (MIP) can be packed into a micro-column for selective solid phase extraction (SPE) of theophylline from 20 microl of sample solution. Its chemical inertness and thermal stability allow the use of various organic solvents and elevated column temperatures for effective binding of theophylline. Non-specific adsorption of interfering drugs on the MIP surface is eliminated by an intermediate wash with 20 microl of acetonitrile, prior to quantitative desorption of the bound theophylline by 20 microl of methanol for in-line UV spectrophotometric determination. In this differential pulsed elution (DPE) technique, both the column temperature and solvent flow rate can be optimized to enhance selectivity. Application of this micro-analytical method, molecularly imprinted solid phase extraction DPE (MISPE-DPE), is demonstrated for accurate determination of theophylline in human blood serum. The method is validated over a linear range from 2 microg/ml to at least 20 microg/ml. PMID- 10701950 TI - Analysis of low concentration sufentanil citrate/bupivacaine hydrochloride admixtures, using solid phase extraction followed by high-performance liquid chromatography. AB - A method has been developed for the quantitative determination of low concentrations of sufentanil citrate (1.0 microg/ml), in the presence of bupivacaine hydrochloride (0.125%), in the quality control of pharmaceutical preparations. The main problem in analysis of this combination is the low concentration of sufentanil citrate in the presence of relatively high concentrations of bupivacaine hydrochloride. This paper describes the validation of a HPLC method of sufentanil citrate in an admixture with bupivacaine hydrochloride using solid phase extraction (SPE). The optimized method shows good linearity, precision and accuracy. The limits of detection (0.09 microg/l) and quantification (0.29 microg/l) for sufentanil citrate are lower than the maximal accepted limits. This method is currently used in stability studies. PMID- 10701951 TI - Photodegradation studies on Atenolol by liquid chromatography. AB - The photostability of the beta-blocker drug Atenolol was evaluated at pH 9, 7.4 and 4.0. The drug was exposed to UVA-UVB radiations and the photoproducts were detected by reversed phase LC methods. The photodegradation was found to increase with the pH value decreasing. The major photodegradation product at pH 7.4 was identified as 2-(4-hydroxyphenyl)acetamide. The LC method developed for routine analyses (column: C-18 Alltima; mobile phase: TEA acetate (pH 4; 0.01 M) acetonitrile 96:4) was found to be suitable for the stability indicating determination of Atenolol in pharmaceutical dosage forms. PMID- 10701952 TI - A solid-phase assay for quantitative analysis of sulfated glycosaminoglycans at the nanogram level. Application to tissue samples. AB - A sensitive and accurate solid-phase methodology for the quantitative analysis of glycosaminoglycans is described. Chondroitin-4-sulfate (CSA) was labelled with biotin hydrazide after the reaction of its carboxyl groups with it in the presence of carbodiimide. Polystyrene plates modified with sequential reaction with glutaraldehyde (GH) and spermine to possess amino groups were used to immobilize electrostatically the biotin labelled CSA. Exogenously added sulfated glycosaminoglycans (GAGS) [variously sulfated chondroitin sulfates and heparan sulfate (HS)] were found to compete to this immobilization in a concentration dependent mode, within a concentration range from 10 up to 300 ng/ml. Glycosaminoglycan-derived oligosaccharides competed to a degree similar to that of intact molecules. Hyaluronan (HA) and keratan sulfate (KS) did not compete the immobilization. The procedure was applied for the rapid and reproducible determination of the sulfated glycosaminoglycans in proteinase digests of small tissue samples or cell cultures with high sensitivity and accuracy. PMID- 10701953 TI - Stability indicating spectrodensitometric determination of ranitidine hydrochloride using linear and non-linear regression. PMID- 10701954 TI - Determination of biotin levels in cerebrospinal fluid samples. PMID- 10701955 TI - Electrochemical study on the determination of tinidazole in tablets. AB - The electrochemical reduction of tinidazole has been carried out in aqueous solution in the pH range 1.8-11.3 by differential-pulse (DP) polarography. Tinidazole exhibits one or two reduction peaks depending on pH. In strongly acidic solution (pH < 4.5), one reduction peak was obtained and it was suitable for analytical purposes. A method for the determination of tinidazole by DP polarography in Britton-Robinson buffer of pH 3.0, which allows quantification over the range 0.03-7.30 microg/ml, was proposed. The method was successfully applied to the determination of tinidazole in tablets with mean recovery and relative standard deviation of 98.7 and 3%, respectively. Excipients did not interfere in the determination. PMID- 10701956 TI - A high-performance liquid chromatographic assay for sparfloxacin. AB - A specific and sensitive high-performance liquid chromatographic procedure was developed for the assay of sparfloxacin in raw material and tablets. It was also found that the excipients in the commercial tablet preparation did not interfere with the assay. The method validation yielded good results and included the range, linearity, precision, accuracy, specificity and recovery. This method can also be applied to stability studies. PMID- 10701957 TI - Determination of benzodiazepine in tablets studied by thermal desorption gas chromatography. AB - Thermal desorption gas chromatography (TDGC) was applied to the analysis of 13 kinds of tablets containing different benzodiazepines (BZDs). An aliquot of ground tablet sample (0.1-1 mg), weighed into a platinum sample cup, was placed in a furnace pyrolyzer where the sample was heated up to a suitable temperature (150-500 degrees C) so that BZD was desorbed from the sample powder. The desorbed components of BZDs were immediately transferred into a separation column by a helium carrier gas without using any trapping techniques. The desorbed components were identified by TDGC-mass spectrometry. Among various BZDs, fludiazepam, nimetazepam and mexazolam in tablet samples were determined by the present method. Thus, the results obtained were in good agreement with the specified values. Correlation coefficients of the calibration lines for the three BZDs' ranged from 0.997 to 0.999 for several micrograms to about 10 microg of the components. Relative standard deviations of this method were < 4.1% for 4 or 5 runs. PMID- 10701959 TI - Choosing the calibration model in assay validation. AB - Data transformations and weighting schemes are normally used to obtain the best fit of standard curves in bioanalysis and the calibration model is usually selected during prevalidation. In the present study, a comparison has been made between unweighted and weighted (1/x, 1/x2, and 1/square root of x) regression models with or without an intercept in achieving the best-fit for the standard curve of CDRI compound 81/470, a new anthelmintic agent, in cow milk. Validation samples in milk at the LLOQ, medium, and high concentrations were also analysed by each of the calibration models. An unweighted regression equation with an intercept overestimated the concentrations at the LLOQ. An unweighted equation without intercept and weighted equations with or without an intercept significantly minimized the bias at the LLOQ without distorting the results at higher concentrations. Hence, an unweighted equation for a straight line passing through the origin was found to be the best model for a standard curve of 81/470 in milk. Similar results were obtained for 81/470 and UMF-078 in serum and plasma, respectively. Bioanalysts should routinely test these models to obtain the best fit model for their calibration curves as part of their assay validation not during prevalidation. PMID- 10701958 TI - Spectrophotometric determination of fluoxetine hydrochloride in bulk and in pharmaceutical formulations. AB - Two new rapid, sensitive and economical spectrophotometric methods are described for the determination of fluoxetine hydrochloride in bulk and in pharmaceutical formulations. Both methods are based on the formation of a yellow ion-pair complex due to the action of methyl orange (MO) and thymol blue (TM) on fluoxetine in acidic (pH 4.0) and basic (pH 8.0) medium, respectively. Under optimised conditions they show an absorption maxima at 433 nm (MO) and 410 nm (TB), with molar absorptivities of 2.12 x 10(-4) and 4.207 x 10(-3) l mol(-1) cm( 1) and Sandell's Sensitivities of 1.64 x 10(-2) and 0.082 microg cm(-2) per 0.001 absorbance unit for MO and TB, respectively. The colour is stable for 5 min after extraction. In both cases Beer's Law is obeyed at 1-20 microg mol(-1) with MO and 4-24 microg mol(-1) with TB. The proposal method was successfully extended to pharmaceutical preparations capsules. The results obtained by both the agreement and E.P. (3rd edition) were in good agreement and statistical comparison by Student's t-test and variance ratio F-test showed no significant difference in the three methods. PMID- 10701960 TI - A stability-indicating method for the determination of melphalan and related impurity content by gradient HPLC. AB - A robust gradient high performance liquid chromatographic (HPLC) procedure is described for the simultaneous determination of melphalan content and related impurities in melphalan drug substance. The sample solution is prepared in methanol and injected. A linear gradient from 5 to 60% acetonitrile in water containing 0.05% v/v acetic acid, 0.01% v/v triethylamine, and 0.05% w/v ammonium acetate is applied over 20 min. The chromatographic conditions are capable of separating and quantifying all impurities found in routine production batches of melphalan at above 0.1% area/area. The method has been fully validated and is linear over the column loading range of 0-3 microg of melphalan. All related impurities occurring in routine batches at above 0.1% area/area have been identified, and structures assigned. The method has been applied to melphalan samples stored under stressed conditions, and shown to be stability-indicating. PMID- 10701961 TI - Development and validation of a new high-performance liquid chromatographic assay of centpropazine, a new antidepressant compound, in serum. AB - Centpropazine is a new anti-depressant compound developed by Central Drug Research Institute, Lucknow (India). We report here the development and validation of a new HPLC assay of a parent drug in the serum of humans, monkeys and rats for pharmacokinetic studies. Centpropazine was eluted on a C18 column with a mobile phase consisting acetonitrile phosphate buffer (60:40) pumped at 1.5 ml min(-1) flow rate and quantitated by UV detector at 270 nm. Considering the sample volume available from different species and to enhance the sensitivity of assay, three sample clean up methods requiring 0.05, 0.5 and 4 ml serum for a linear quantitation range of 312.5 ng ml(-1)-5 microg ml(-1), 0.04-2.5 microg ml( 1) and 2.5-80 ng ml(-1) respectively were developed. The lowest limit of quantitation of the method was 2.5 ng ml(-1) requiring 4 ml serum, 40 ng ml(-1) requiring 0.5 ml serum and 312 ng ml(-1) requiring 50 microl serum sample. All these methods were fully validated in human serum and extended to monkey and rat serum. The recovery of centpropazine at 5, 80, 625, 1280 and 2500 ng ml(-1) ranged between 92 and 105%. The within and between run variability in precision and accuracy were less than 10% and the drug in serum was stable up to three freeze-thaw cycles. Overall the method is simple, quick and robust for biopharmaceutical applications. The method was applied to analyse concentrations of centpropazine in rat serum after administering single 20 mg kg(-1) peroral and 5 mg kg(-1) i.v. dose. The chromatograms of treated rat serum exhibited three well resolved peaks of metabolites and one of them was identified as hydroxy metabolite of centpropazine. PMID- 10701962 TI - Macrolide and ketolide antibiotic separation by reversed phase high performance liquid chromatography. AB - Twenty different macrolide and ketolide antibiotics were analyzed by reversed phase high performance liquid chromatography on an ODS-2 cartridge column. Each of these compounds was uniquely separated and purified by varying the flow rate. Retention times of the individual drugs were proportional to the flow rate of the mobile phase. Recovery of antimicrobial activity for most of the drugs was greater than 90% based on a microbiological assay of material recovered from the column. Retention times were related to structural differences between these antimicrobial agents. PMID- 10701963 TI - Identification of oxidative degradates of the thrombin inhibitor, 3-(2 phenethylamino)-6-methyl-1-(2-amino-6-methyl-5-methyleneca rboxamidomethylpyridinyl)pyrazinone, using liquid chromatography/mass spectrometry and liquid chromatography/tandem mass spectrometry. AB - Liquid chromatography/mass spectrometry was used to identify reaction products from a solution of 3-(2-phenethylamino)-6-methyl-1-(2-amino-6-methyl-5 methyleneca rboxamidomethylpyridinyl)pyrazinone (L-375,378) and hydrogen peroxide, a system that generates high levels of the oxidative degradates which form in the tablets and intravenous (i.v.) solutions of L-375,378. Two major hydrogen peroxide reaction products of L-375,378 (m/z 407) with m/z values of 369 and 370 were separated and identified. Both compounds were products of ring opening with elimination of three carbon atoms from the center pyrazinone ring. The structural assignments for these two products were alpha-amidinoamide and alpha-diamide compounds, respectively. In addition, five products (m/z 423) with a molecular weight 16 Da greater than that for L-375,378 were separated. Further liquid chromatography/tandem mass spectrometry experiments indicated that three of these M + 16 products were phenolic derivatives of L-375,378. Among them, the para-hydroxy compound has been verified using an authentic standard. The other two phenolic compounds were believed to be the meta- and ortho-hydroxy derivatives of L-375,378. The fourth M + 16 product was derived from hydroxylation of the methyl group on the center pyrazinone ring. The fifth M + 16 product was derived from oxidation on the aminopyridine moiety, most likely N oxide of the pyridine ring. Other minor hydrogen peroxide reaction products were not studied in detail because they did not appear in tablets or i.v. formulations. PMID- 10701964 TI - Determination of cyanocobalamin, betamethasone, and diclofenac sodium in pharmaceutical formulations, by high performance liquid chromatography. AB - The aim of this work was to develop an analytical method for a simultaneous determination of cyanocobalamin (Vitamin B12), betamethasone, and diclofenac, present in pharmaceutical formulations, by high performance liquid chromatography, assuring rapidity, accuracy, precision, and selectivity. The working conditions were as follows: RP18 column of 125 mm x 4 mm ID and a particle size of 5 microm; mobile phase acetonitrile-water (40:60; v/v) (pH* 3.45) adjusted with acetic acidl flow gradient from 0.8 to 1.9 ml/min.; injection volume of 20 microl; temperature 34 degrees C and detection at 240 nm. The method was adequately validated, and linearity, accuracy, as well as the system, method and interday precision, for each active principle, were determined. PMID- 10701965 TI - On the use of liquid chromatography with radio- and ultraviolet absorbance detection coupled to mass spectrometry for improved sensitivity and selectivity in determination of specific radioactivity of radiopharmaceuticals. AB - Pneumatically assisted electrospray mass spectrometry was evaluated as a complementary detection technique to UV absorbance, for determination of specific radioactivity of tracer molecules to be used in positron emission tomography. Tracers labelled with radionuclides having short half-lives can be synthesised with high specific radioactivity. The UV absorbance detection that is commonly used for the determination does not always have the sensitivity required for those analyses. In comparison, mass spectrometry gave improved detection limits in all but one (nicotine) of the 12 compounds studied. The magnitude of this improvement was more than 100-fold for the compounds ketamine (2-methylamino-2-(2 chloro-phenyl)cyclohexanone), SCH-23390 ((R)-(+)-7-chloro-8-hydroxy-1-methyl-1 phenyl-2,3,4,5-tetra-hydro-1H-3-b enzazepine) and N-methyl-piperidylbenzilate. These improved detection limits, specificity, plus the added certainty of product identity provided by mass spectral data demonstrated the value of the mass spectrometer as a complementary detector in the determination of specific radioactivity. PMID- 10701966 TI - Sedimentation field-flow fractionation application to Toxoplasma gondii separation and purification. AB - Toxoplasmosis is a worldwide disease caused by Toxoplasma gondii, an intracellular protozoa of micronic size range (4-10 microm). Its classical purification processes are complex and often associated with low recovery. All investigation procedures concerning this parasite require its isolation and purification from at least the mouse ascitic fluid. For this purpose, a recently developed laboratory technology was used, i.e. sedimentation field-flow fractionation. This chromatographic-like separation technology was demonstrated to be particularly selective for isolation and separation of micron-sized biological particles. Sedimentation field-flow fractionation operated on the steric-hyperlayer mode was used to isolate the parasite from the remanent ascitic contaminants of different origins and from red blood cells. With this technology, 86% recovery with 97% viability was obtained in less than 30 min. PMID- 10701967 TI - In-house packing and testing of capillaries for capillary electrochromatography using simple equipment. AB - The feasibility of producing packed capillaries for capillary electrochromatography (CEC) is evaluated. Emphasis is put on the fact that only material was used that is already available in any LC/CE orientated laboratory. An experimental set-up is developed for filling the capillaries by use of an ordinary LC pump, and frits are sintered with a glowing resistance wire, fed by a d.c. power supply. Electrochromatography was carried out in an in-house built capillary electrophoresis apparatus, without pressurizing the vials. Under these conditions, the capillaries performed well, producing up to 190,000 plates per meter. Bubble formation did not appear, on condition that the mobile phase was thoroughly helium degassed. Even at ambient temperature, electrophoresis obeyed Ohm's law up to a voltage of 30 kV, proving that Joule heating was not a major concern. PMID- 10701968 TI - Stability of reconstituted solutions of ceftazidime for injections: an HPLC and CE approach. AB - The stability of aqueous reconstituted ceftazidime injection vials containing ceftazidime pentahydrate blended with anhydrous sodium carbonate was investigated in different storage conditions (4 degrees C and 10 degrees C for 7 days in a refrigerator, 20 and 30 degrees C for 24 h) with validated HPLC and (micellar) CE methods. Stability indicating data were obtained for ceftazidime and two degradation products: pyridine and the delta2-ceftazidime isomer. Other degradation products were also identified (the complementarity of the two used experimental procedures was useful in such exercise) and characterized by their UV spectra and retention times. Stability data (7 days at 4 degrees C in a refrigerator and 18 h at room temperature) resulted in agreements with the manufacturers prescription and point out the need of a strict temperature control of the refrigerator's compartment used to store the reconstituted solution. PMID- 10701969 TI - Stability indicating assays for the determination of piroxicam--comparison of methods. AB - Photodegradation of piroxicam, a 1,2-benzothiazine oxicam, is studied laying special emphasis on the investigation of the correlation between concentration of the sample solution and stability. A comparison of three different methods (HPTLC/densitometry, HPLC, CE) developed for the photostability testing of the title compound is presented. The stability indicating capability of the assays is proved using forced degradation by exposing a sample solution to artificial irradiation from a xenon source. The chromatograms and the electropherogram of the resulting solution show piroxicam well resolved from the degradation products. For quantitation external calibration is employed, all calibration curves being linear in the respective concentration range of interest. Piroxicam solutions of three different concentrations (2 mg ml(-1); 250 microg ml(-1); 40 microg ml(-1)) are subjected to simulated sunlight for 480 min. The stability is investigated by quantitation of piroxicam by the methods mentioned. The methods are compared in respect of performance and precision. Costs and time of analysis are regarded also. PMID- 10701970 TI - A simple HPLC method using a microbore column for the analysis of doxorubicin. AB - Doxorubicin is one of the most potent anti-tumor agents generally used in the treatment of bone cancer. A simple and sensitive HPLC method was developed and validated for the assay of doxorubicin. The method used a C18 Luna microbore column (50 x 1 mm) with a fluorescent detector (505 nm Ex. and 550 nm Em.). The mobile phase consisted of water-acetonitrile-acetic acid (80:19:1, v/v/v, pH 3.0) and the flow rate was 0.1 ml min(-1). Daunomycin was used as the internal standard. This isocratic system required a 10-min run-time, giving a detection limit of 0.02 ng (0.035 pmol per injection). Standard curves were linear over the concentration range of 0.01-0.1 microg ml(-1). Relative standard deviations (R.S.D.) for the within-day, day-to-day precision, and the accuracy measurement for the assay were less than 4.0, 3.2, and 4.1%, respectively. This HPLC method was used to study the in vitro release characteristics of doxorubicin from implantable drug delivery system. PMID- 10701971 TI - High sensitivity ELISA determination of taxol in various human biological fluids. AB - Taxol (paclitaxel)--the natural product isolated from Pacific yew (Taxus brevifolia)--is a novel agent with high activity in the treatment of patients with several malignant tumors including those resistant to other cytotoxic drugs. The therapeutic index of this promising anticancer drug could be further increased by the exploration of its pharmacokinetic pharmacodynamic relationship in cancer patients. Since taxol is highly protein bound, a very specific and highly sensitive analytical method is required in order to determine free, protein unbound and biologically active taxol species in human physiological fluids: plasma; plasma ultrafiltrate; and salivary fluids. In order to accomplish this, a new indirect competitive enzyme-linked immunosorbent assay (ELISA), for quantitating such a low bioactive taxol concentration level, has been developed in our laboratories. This method uses taxol competitive inhibition of mouse anti taxol antibodies binding to the solid phase coated antigen 7-succinyltaxol-bovine serum albumin. This indicates recognition of the active taxol in the solution phase, where a diluted horseradish peroxidase labeled goat anti-mouse enzyme conjugate is used. While employing this technique, after systematic optimization of the experimental conditions, we are able to detect the anticipated taxol in plasma ultrafiltrate and salivary fluids at the concentration level of subpicogram per milliliter. The working range of the assay is approximately five orders in magnitude, i.e. from pg ml(-1) to 100 ng ml(-1). The clinical part of this study verified the working range of the ELISA method using samples of physiological fluids from a cancer patient treated with 3 h intravenous (i.v.) infusion of this drug. Our results of taxol determination in plasma, plasma ultrafiltrate and saliva demonstrate the applicability of the newly developed ELISA method for further pharmacokinetic studies of free, biologically active taxol species in cancer patients. PMID- 10701972 TI - Spectrophotometric determination of certain cephalosporins through oxidation with cerium(IV) and 1-chlorobenzotriazole. PMID- 10701973 TI - Potentiometric determination of monofluorphosphate in dentifrice: a critical discussion and a proposal for new improved procedures. PMID- 10701974 TI - Chromatographic and UV-VIS and IR spectral methods for qualitative and quantitative analysis of some synthetic bromo-flavone derivatives. PMID- 10701975 TI - Cathodic stripping voltammetric behaviour of nitrofurazone and its determination in pharmaceutical dosage form, urine and serum by linear sweep voltammetry. PMID- 10701976 TI - Spectrofluorometric determination of diclofenac in tablets and ointments. PMID- 10701977 TI - Voltammetric determination of amoxicillin using a poly (N-vinyl imidazole) modified carbon paste electrode. PMID- 10701978 TI - Determination of Lansoprazole in pharmaceutical dosage forms by two different spectroscopic methods. AB - Two different ultraviolet (UV) spectroscopic methods were developed for determination of Lansoprazole in pharmaceutical dosage forms. The solutions of the standard and the sample were prepared in 0.1 M NaOH and phosphate buffer pH 6.6. Both UV spectrophotometric and derivative spectroscopic techniques were applied. Second-order derivative spectra were generated between 200 and 400 nm at N = 9, deltalambda = 31.5. The linear range for the UV spectrophotometric method was 3.0-25.0 microg ml(-1) and that for the derivative spectroscopic method was 0.5-25.0 microg ml(-1). The developed methods were applied to three different pharmaceutical preparations. The percentage recovery was 100.2%. PMID- 10701979 TI - Determination of roquefortine C in blue cheese using on-line column-switching liquid chromatography. AB - A method is described for the determination of roquefortine C in (blue) cheese. After liquid-liquid extraction with a mixture of hydrochloric acid and methanol, and filtration, an aliquot is analysed using column-switching reversed-phase liquid chromatography. The recovery of roquefortine C in Fetta cheese is about 85%, the calibration curve is linear from 10 to 2500 ng g(-1) (r2 = 0.998), and the detection limit is about 10 ng g(-1). In different batches of Danish Blue concentrations of 1000-2000 ng g(-1) of roquefortine C are found. As regards the stability of roquefortine C its half-life in diffuse daylight is ca. 50 min, while after irradiation with ultraviolet light, it is about 10 min. PMID- 10701980 TI - Spectrodensitometric determination of clorazepate dipotassium, primidone and chlorzoxazone each in presence of its degradation product. AB - The present work describes a quantitative thin layer procedure for estimating primidone, clorazepate dipotassium and chlorzoxazone in bulk powders and in dosage forms, each in the presence of its degradation product. The method consists of dissolving the drug in ethanol (for primidone), or methanol (for clorazepate dipotassium and chlorzoxazone) and then spotting this solution on a thin layer of silica gel G254. Quantitation is achieved by comparing the areas under the peaks obtained from scanning the thin layer chromatographic plates in a spectrodensitometer. PMID- 10701981 TI - Effects of cimetidine on the pharmacokinetics of proguanil in healthy subjects and in peptic ulcer patients. AB - The pharmacokinetics of orally administered proguanil and its metabolites were determined in six healthy volunteers and in six peptic ulcer patients, before and after a 3-day course of cimetidine (400 mg given two times daily for 2 days and 400 mg on the third day 1 h before proguanil). Cimetidine significantly increased Cmax (P < 0.05), AUCo-alpha (P < 0.005) and elimination half-life t 1/2b of proquanil in plasma of healthy subjects. In ulcer patients, cimetidine significantly increased, AUCo-alpha (P < 0.05), elimination half life (P < 0.005) and Cmax. Cimetidine significantly reduced (P < 0.05) Total body clearance in both healthy subjects and in peptic ulcer patients. The Cmax and AUCo-alpha of the active metabolite cycloguanil was significantly decreased (P < 0.05) in both the healthy subjects and in the peptic ulcer patients. The Cmax of the inactive metabolite, 4-CPB was significantly decreased in healthy subjects and AUCo-alpha significantly decreased in peptic ulcer patients. PMID- 10701982 TI - Liquid chromatography method for separation of clindamycin from related substances. AB - A reversed-phase liquid chromatography method has been developed for the separation of clindamycin from 7-epiclindamycin, clindamycin B, lincomycin, lincomycin B, 7-epilincomycin and other impurities of unknown identity. The method uses a Hypersil ODS, 5 microm, 250 x 4.6 mm i.d. column maintained at 45 degrees C. The mobile phase comprises acetonitrile phosphate buffer (1.35% v/v phosphoric acid, adjusted to pH 6.0 with ammonium hydroxide)-water (35:40:25, v/v) at a flow rate of 1.0 ml/min. UV detection is performed at 210 nm. The method was tested on several C-18 columns and showed good robustness. Robustness was further evaluated by performing a full-fraction factorial design experiment. The method showed good selectivity, linearity, and repeatability. It is also suitable for analysis of clindamycin formulations. PMID- 10701983 TI - Improved benzodiazepine radioreceptor assay using the MultiScreen Assay System. AB - In this article, an improved benzodiazepine radioreceptor assay is described, which allows substantial reduction in assay time. The filtration in this method was performed by using the MultiScreen Assay System. The latter consists of a 96 well plate with glass fibre filters sealed at the bottom, which allows both the incubation and the filtration of the specimen in the same plate. After the filtration, the filters were punched out for quantitation of the bound labeled ligand [3H]flunitrazepam. The results obtained with the MultiScreen Assay System did not differ significantly from the data obtained with the conventional filtration manifold (48S): The Ki's of lorazepam were 2.4 +/- 0.30 and 1.9 +/- 0.15 nM, respectively. In case a radioactive label is replaced by a fluorescent label, the bound labeled-ligand usually cannot be determined in the presence of the receptor material. Here, the bound labeled-ligand has to be dissociated after the filtration step. To dissociate the ligand-receptor complex, Tris- HCl buffer, containing 10 microM flumazenil, was added to the filters and the second filtrates were collected containing the previously bound fractions in the absence of receptor material. This approach showed the same Ki for lorazepam, 2.5 +/- 0.04 nM as without dissociation, when using the radio-labeled benzodiazepine [3H]flunitrazepam. PMID- 10701984 TI - Isolation and identification of cyclic imide and deamidation products in heat stressed pramlintide injection drug product. AB - This report summarizes the identification of six cyclic imide [Asu] and two deamidation products from a sample of pramlintide final drug product that had been stressed at 40 degrees C for 45 days. The pramlintide degradation products were isolated by cation exchange high-performance liquid chromatography (HPLC) followed by reversed-phase HPLC. The isolated components were characterized by mass spectrometry (MS), tandem MS (MS/MS) and when necessary, by enzymatic (thermolysin) digestion followed by liquid chromatography/mass spectrometry (LC/MS) and sequence analysis. The isolated products were identified as [Asu14] pramlintide, [Asu21]-pramlintide, [Asu22]-pramlintide, [Asu35]-pramlintide, [1 21]-succinimide-pramlintide, and [1-22]-succinimide-pramlintide. Also identified were [Asp35]-pramlintide, the deamidation product of pramlintide at Asn35, and [Tyr37-OH]-pramlintide, the deamidation product of the pramlintide amidated C terminal Tyr. Together these data support those presented earlier (C. Hekman et al., Isolation and identification of peptide degradation products of heat stressed pramlintide injection drug product. Pharm Res 1998;15:650-9) indicating that the primary mechanism of degradation for pramlintide in this pH 4.0 formulation is deamidation, with six of the eight possible deamidation sites observed to undergo deamidation. Gln-10 and Asn-31 are the only two residues subject to deamidation for which none is observed. The data indicate that the cyclic imide products account for approximately 20% of the total thermal degradation while the deamidation products account for 64%. The remaining degradation is due to peptide backbone hydrolysis. PMID- 10701985 TI - Characterization of a technique for rapid pharmacokinetic studies of multiple co eluting compounds by LC/MS/MS. AB - A method for rapid pharmacokinetic screening of multiple potential drug candidates has been developed. This technique, based on the ability of liquid chromatography coupled with tandem mass spectrometry (LC/MS/MS) to independently monitor multiple components, enables the quantification of substances which may or may not be chromatographically resolved. Our results indicate that the limit of quantitation and accuracy of this multiple-compound LC/MS/MRM quantitation method are comparable to a single-compound LC/MS/MRM quantitation method. No apparent ion suppression due to the existence of extraneous compounds in the analytical solution and biological matrix effect are observed in the range of the calibration curve. The issue of potential residual molecule cross-talk interference existing in the multiple-reaction monitoring mode has been discussed. This multiple-compound LC/MS/MRM quantitation method can be used for high throughput pharmacokinetic screening and to assay mixtures that have co eluting analytes or similar m/z of precursor/product ion pairs. PMID- 10701986 TI - Transfer from manual to automated sample preparation: a case study. AB - A manual sample preparation for a controlled release capsule formulation has been converted to an automated sample preparation. Each step of the manual sample preparation was evaluated as to its feasibility for automation in terms of precision, carryover, filter selection and other critical issues. Although most steps of the manual method were easily translated to the automated procedure, certain 'simple' details such as filter selection, sample storage, and the conversion from volumetric to gravimetric measurements needed closer investigation. PMID- 10701987 TI - High-performance liquid chromatographic analysis of new triazole antifungal agent SYN-2869 and its derivatives in plasma. AB - A simple reversed-phase high-performance liquid chromatography (HPLC) method with UV detection was developed and validated for the quantitation of SYN-2869, a novel triazole antifungal agent and its analogs in rat plasma. The method involved a simple precipitation of plasma protein with acetonitrile (1:10 ratio). The reconstituted sample after evaporation to dryness was injected onto a HPLC column. SYN-2869 and its analogs were separated from the matrix components on a symmetry C18 column using an aqueous mobile phase of acetonitrile and water with a flow rate of 1 ml min(-1). A step gradient of 40-80% acetonitrile eluted all four compounds. The run time was 30 min. The linear range was 0.5 10 microg ml( 1)(r2 > 0.999). The limit of quantitation was 0.5 microg ml(-1). The inter-day precision and accuracy for SYN-2869 standard concentration were from 1.9 to 8.5% and from 1.4 to +/- 4.40%, respectively. The precision and accuracy of intra-day quality control samples were from 4.6 to 5.2% and from 4.6 to 12%, respectively. PMID- 10701988 TI - Determination of LY295501 in human plasma by reverse phase HPLC with UV detection. AB - A high performance liquid chromatographic (HPLC) assay was developed and validated for the quantitative determination of LY295501 in human plasma. A structural analog, LY186641, was selected as the internal standard. The samples were processed by protein precipitation with acetonitrile followed by concentration of the supernatants and reconstitution. Chromatographic resolution of LY295501 from endogenous plasma components was accomplished with a Waters Novapak C18 HPLC column (3.9 x 150 mm, d(p) 4 mm). Detection was by absorbance at 260 nm. The linear dynamic range was from 5 to 400 microg ml(-1) of human plasma using a 0.25 ml aliquot. The inter-day precision (%RSD) and accuracy (%RE) in plasma ranged from 2.4 to 4.7, and -4.9 to 1.4, respectively. This assay is both simple and rapid, and has been used to successfully analyze over 1500 samples from human clinical trials. PMID- 10701989 TI - Determination of the nutraceutical, glucosamine hydrochloride, in raw materials, dosage forms and plasma using pre-column derivatization with ultraviolet HPLC. AB - A selective and specific high performance liquid chromatography method was developed to quantitate glucosamine hydrochloride in raw materials, dosage forms and plasma. Reverse phase chromatography using pre-column derivatization with phenylisothiocyanate, and ultraviolet detection (lambda = 254 nm) was used to quantify the eluate. The mobile phase consisted of MeOH/H2O/CH3COOH (10:89.6:0.04) and was pumped at a flow rate of 1.2 ml/min. The standard curves for glucosamine hydrochloride showed linearity (r > or = 0.99) over the selected concentration range from 6.65 to 16.63 microg/ml for raw materials and dosage forms. The precision of the dosage form assay, expressed as the % relative standard deviation (R.S.D.), was < 5% at all concentrations. The intra-day and inter-day accuracy, as indicated by the relative error (R.E.), ranged from - 2.54 to 2.70% for glucosamine hydrochloride. For the plasma assay, beagle dog plasma was used to prepare standard curves in the concentration range of 1.25-20 microg/ml. Precipitation of plasma proteins was accomplished with acetonitrile to separate interfering endogenous products from the compound of interest. The supernatant was derivatized using phenylisocyanate in phosphate buffer (pH = 8.3) and subsequently evaporated to dryness under a nitrogen stream at 42 degrees C. The residue was dissolved in 250 microl mobile phase and injected onto the chromatographic system. The assay was linear in concentration ranges of 1.25-20 microg/ml (r > or = 0.999). Intra- and inter-day precision was < or = 5.23 and 5.65%, respectively and the intra- and inter-day accuracy, indicated by R.E., ranged from - 8.6 to 10.35%. The method was found to be specific and with excellent linearity, accuracy and precision and is well suited for the quantitation of glucosamine hydrochloride in raw materials, dosage forms, and pharmacokinetic studies. PMID- 10701991 TI - Development of a mentoring program for new authors. PMID- 10701990 TI - Distinction among eight opiate drugs in urine by gas chromatography-mass spectrometry. AB - Opiates are commonly abused substances, and forensic urine drug-testing for them requires gas chromatographic-mass spectrometric (GC-MS) confirmation. There are also medical reasons to test urine for opiates, and confirmation procedures other than GC-MS are often used for medical drug-testing. A thin-layer chromatographic (TLC) method distinguishes morphine, acetylmorphine, hydromorphone, oxymorphone, codeine, dihydrocodeine, hydrocodone, and oxycodone in clinical specimens. In certain clinical circumstances, GC-MS confirmation is requested for opiates identified by TLC, but, to our knowledge, no previous report examines all of the above opiates in a single GC-MS procedure. We find that they can be distinguished by GC-MS analyses of trimethylsilyl (TMS) ether derivatives, and identities of 6 keto opiates can be further confirmed by GC-MS analysis of methoxime (MO)-TMS derivatives. Inclusion of deuterium-labeled internal standards permits identification of the opiates in urine at concentrations below the TLC cutoff level of 600 ng/ml, and the GC-MS assay is linear over a concentration range that spans that level. This GC-MS procedure has proved useful as a third-stage identification step in a medical drug-testing sequence involving prior immunoassay and TLC. PMID- 10701992 TI - Synthesis of glucose oxidase and catalase by Aspergillus niger in resting cell culture system. AB - The synthesis of glucose oxidase and catalase by Aspergillus niger was investigated using a resting cell culture system without growth being established. Calcium carbonate induced the synthesis of both enzymes and calcium chloride inhibited it. The optimal pH for the biosynthesis of glucose oxidase and catalase was 6.0 and 5.7, respectively. The effects of other bivalent cations, reductive compounds and metabolic products on enzyme synthesis were also tested. The biosynthesis of glucose oxidase and catalase was promoted by MnCO3, thioglycolic acid, pyroracemic acid and gluconic acid. PMID- 10701993 TI - Proceedings of the International Conference on Nuclear Analytical Methods in the Life Sciences. Beijing, China, 26-30 October 1998. PMID- 10701994 TI - Proceedings of the 1st Central European Oncology Congress. Opatija, Coratia, 1998. PMID- 10701995 TI - ["Human albumin--time to say good-bye?"]. PMID- 10701996 TI - In memoriam: Norman Dion Levine (1912-1999). PMID- 10701997 TI - The Fifth European Workshop on Virus Evolution and Molecular Epidemiology, Leuven, Belgium, 30 August to 4 September 1999. Evolving viruses and virologists. PMID- 10701998 TI - Proprotein and prohormone convertases: a family of subtilases generating diverse bioactive polypeptides. AB - Proproteins and prohormones are the fundamental units from which bioactive proteins and peptides as well as neuropeptides are derived by limited proteolysis within the secretory pathway. Precursors are usually cleaved at the general motif (K/R)--(X)n--(K/R)down arrow, where n=0, 2, 4 or 6 and X is any amino acid and usually is not a Cys. Seven mammalian precursor convertases (PCs) have been identified: PC1, PC2, furin, PC4, PC5, PACE4 and PC7. Each of these enzymes, either alone or in combination with others, is responsible for the tissue specific processing of multiple polypeptide precursors both in the brain and in periphery. This combinatorial mechanism generates a large diversity of bioactive molecules in an exquisitively regulated manner. The production of null mice allowed the assessment of the critical role of convertases in vivo. Thus, male PC4 (-/-) mice are infertile, furin (-/-) and PC1(-/-) mice are embryonic lethal, and PC2 (-/-) mice are mildly diabetic and runted. Interestingly, animals deficient in 7B2, a PC2-specific binding protein, exhibit a Cushing-like syndrome and die soon after birth. Recently, the first member of a new class of subtilisin -kexin-like convertases, called SKI-1, was identified. Its structure is closer to pyrolysin than to mammalian PCs and it exhibits a specificity for cleavage at the motif (R/K)--X--X--(L,T) down arrow as deduced from its ability to process sterol regulatory element binding proteins and pro-brain derived neurotrophic factor. Thus, while PCs are responsible for the processing of neuropeptides, adhesion molecules, receptors, growth factors, cell surface glycoprotein and enzymes, SKI 1 cleaves proproteins that are critical for the control of cholesterol and fatty acid metabolism and for neuronal protection and growth. PMID- 10701999 TI - Outcome in antenatally diagnosed renal pelvis dilatation. PMID- 10702000 TI - Unlicensed and off label drug use in neonates. PMID- 10702001 TI - Glycosaminoglycans in neonatal urine. PMID- 10702002 TI - Proceedings of an international workshop on biomechatronics. Enschede, The Netherlands, April 1999. PMID- 10702003 TI - Re: The Clinical Quality Improvement Network (CQIN) Investigators. Thromboembolic prophylaxis in 3,575 hospitalized patients with atrial fibrillation. 1998;14:695 702. PMID- 10702004 TI - [Anatolii L'vovych Mikhn'ov (on the 90th anniversary of his birth)]. PMID- 10702005 TI - Dentistry on stamps. PMID- 10702006 TI - Monitoring, surveillance and research needs. Public health planning priorities and policy options. PMID- 10702007 TI - Proceedings of the 5th International Conference on Equine Exercise Physiology. Utsunomiya, Japan, 20-25 September 1998. PMID- 10702008 TI - 5th National Rural Health Conference. 14 March 1999. Abstracts. PMID- 10702009 TI - Cigars. A hazard to health. PMID- 10702010 TI - Guess what! Perforating pilomatricoma resembling keratoacanthoma. PMID- 10702011 TI - Guess what! Malignant eccrine spiradenoma. PMID- 10702012 TI - Guess what! Malignant peripheral nerve sheath tunour in a patient with neurofibromatosis type I. PMID- 10702013 TI - Literature review. PMID- 10702014 TI - Message from the chairperson. National Kidney Foundation. PMID- 10702015 TI - British Society for Haematology Slide Session, Annual Scientific Meeting, Brighton, 1999. PMID- 10702016 TI - Reference values for the activated partial thromboplastin time in infants using a synthetic reagent. PMID- 10702017 TI - A case of idiopathic hypereosinophilic syndrome with hypersegmented and hypogranular eosinophils. PMID- 10702018 TI - Infection of Helicobacter pylori in gastric adaptation to continued administration of aspirin in humans. PMID- 10702019 TI - Evaluation of endoscopic variceal ligation in prophylactic therapy for bleeding of oesophageal varices: a prospective, controlled trial compared with endoscopic injection sclerotherapy. PMID- 10702020 TI - The role of surveillance endoscopic retrograde cholangiopancreatography in preventing episodic cholangitis in patients with recurrent common bile duct stones. PMID- 10702021 TI - Metal stents improve dysphagia, nutrition and survival in malignant oesophageal stenosis: a randomized controlled trial comparing modified Gianturco Z-stents with plastic Atkinson tubes. PMID- 10702022 TI - Basic techniques of ERCP. PMID- 10702023 TI - Endodontic case difficulty assessment: the team approach. PMID- 10702024 TI - Unified nomenclature for the winged helix/forkhead transcription factors. PMID- 10702025 TI - Sharing of an HLA-B27-restricted H-Y antigen between rat and mouse. PMID- 10702026 TI - [Alternative medicine and natural healing]. PMID- 10702027 TI - [Interventional cardiology: the problem of recurrent stenosis. 2nd European Congress of Pharmacology. Budapest, 2-7 July 1999]. PMID- 10702028 TI - Papers from the 15th International Conference on Kinins. Part II. Nara, Japan, 19 24 October 1998. PMID- 10702029 TI - Papers of the 15th International Conference on Kinins. Part III. Nara, Japan, 19 24 October 1998. PMID- 10702030 TI - Abdominal pressure in the critically ill: measurement and clinical relevance. PMID- 10702031 TI - High frequency oscillatory ventilation. PMID- 10702032 TI - Management of cerebral vasospasm in patients with aneurysmal subarachnoid hemorrhage. PMID- 10702033 TI - History of The Journal of Hand Surgery: 1976-1999. PMID- 10702034 TI - ESHHS. European Society for the History of the Human Sciences. Report on the Eighteenth Annual Conference, Florence, 3-7 September 1999. PMID- 10702035 TI - Gender, chronic infection and myocardial infarction. PMID- 10702036 TI - Metal complexes of taurine. The first reported solution equilibrium studies for complex formation by taurine at physiological pH; the copper(II)-glycylglycinate taurine and the copper(II)-glycylaspartate-taurine systems. AB - The first solution studies at physiological pH for the formation of metal complexes of taurine, +NH3CH2CH2S03-, one of the most abundant low molecular weight organic compounds in the animal kingdom, are reported. The complexes Cu(Gly-GlyH-1) (1) and [Cu(Gly-AspH-1)] (2) react with taurine to give the ternary complexes [Cu(Gly-GlyH-1)taurine]- (3) (log K=2.95+/-0.03, I=0.2M, T=25.0 degrees C) and [Cu(Gly-AspH-1)taurine]2- (4) (log K=2.68+/-0.02) in which taurine acts as an N-donor ligand, most likely monodentate, without involvement of the sulphonate group in coordination. The results of the pH-metric studies are confirmed by visible and EPR spectrophotometric studies. The taurine complexes are less stable than the analogous complexes of beta-alanine due to the decreased basicity of the amino group in the former ligand, and in the case of the Cu(Gly GlyH-1) complexes due to involvement of the carboxylate group of beta-alanine in axial coordination. PMID- 10702037 TI - Acute optic neuritis in Australia: a 13 year prospective study. PMID- 10702038 TI - Post-traumatic hydrocephalus: influence of craniectomy on the CSF circulation. PMID- 10702039 TI - Diencephalic amnesia and apraxia after left thalamic infarction. PMID- 10702040 TI - Transverse myelopathy in the antiphospholipid antibody syndrome: pinworm infestation as a trigger? PMID- 10702041 TI - Radiologically selective visual pathway involvement in adult onset cerebral adrenoleukodystrophy. PMID- 10702042 TI - Golf ball epilepsy. PMID- 10702043 TI - Sensory predominant neuropathy with GM, antibodies, conduction blocks, and orbital pseudotumour. PMID- 10702044 TI - Alterations of muscarinic acetylcholine receptor subtypes in diffuse Lewy body disease: relation to Alzheimer's disease. PMID- 10702045 TI - Mortality from Parkinson's disease. PMID- 10702046 TI - Anaphylactoid reaction to methylprenisolone. Is it surprising when pharmacological and immune effects of a drug differ? PMID- 10702047 TI - Literature alerts. PMID- 10702048 TI - Carter-Thomason uterine suspension and positioning by ligament investment and truncation. PMID- 10702049 TI - Carter-Thomason uterine suspension and positioning by ligament investment and truncation. PMID- 10702050 TI - Carter-Thomason uterine suspension and positioning by ligament investment and truncation. PMID- 10702052 TI - 2020 Extra: more NIH heads view the future. PMID- 10702051 TI - Economic impact of automated primary screening for cervical cancer. PMID- 10702053 TI - Joseph Decherd Guess, M.D. (1891-1981). PMID- 10702054 TI - International activities of the Doctors Mayo. PMID- 10702055 TI - The relation between somatic symptoms and depression. PMID- 10702056 TI - The relation between somatic symptoms and depression. PMID- 10702057 TI - Idiopathic eosinophilia. PMID- 10702058 TI - Idiopathic eosinophilia. PMID- 10702059 TI - Idiopathic eosinophilia. PMID- 10702060 TI - Multidrug-resistant Salmonella enterica serotype typhimurium DT104. PMID- 10702061 TI - Respiratory distress associated with zanamivir. PMID- 10702062 TI - Recovery of heart rate after exercise. PMID- 10702063 TI - Recovery of heart rate after exercise. PMID- 10702064 TI - Recovery of heart rate after exercise. PMID- 10702065 TI - Biliary tract cancers. PMID- 10702066 TI - Toll genes and responsiveness to bacterial endotoxins. PMID- 10702067 TI - Treatment of neurogenic incontinence with botulinum toxin A. PMID- 10702068 TI - Proteomics is getting easier in some ways. PMID- 10702069 TI - [XXI National Congress of the Italian Society of Angiology and Vascular Pathology. Bologna, Italy, November 28-December 1, 1999. Proceedings]. PMID- 10702070 TI - Infectious diseases diagnosis: current status and future trends. PMID- 10702071 TI - Index of suspicion. Case 4. Epiglottitis. PMID- 10702072 TI - Index of suspicion. Case 2. Carbon monoxide (CO) poisoning. PMID- 10702073 TI - Index of suspicion. Case 3. Congenital sucrase-isomaltase deficiency. PMID- 10702074 TI - Index of suspicion. Case 5. Esophageal atresia (EA) with tracheoesophageal fistula (TEF). PMID- 10702075 TI - Index of suspicion. Case 6. Acute rheumatic fever (ARF). PMID- 10702076 TI - Invited editorial: preimplantation genetic diagnosis. PMID- 10702077 TI - [Dental quality assurance]. PMID- 10702078 TI - [Anonymous: Saint Apollonia. The Bonadent Collection]. PMID- 10702079 TI - [Everything about intraoral video systems--part II. Advice on the purchase decision and on use]. PMID- 10702080 TI - [The emergency in dental practice: rare but dangerous]. PMID- 10702081 TI - [Citius, altius, fortius--but for whose benefit?]. PMID- 10702082 TI - [Quality standards: the GHZ takes a position. Helvetische Gesellschaft der Zahnarzte (Helvetian Society of Dentists)]. PMID- 10702083 TI - [Everything about intraoral video systems--part I. Advice on the purchase decision and on use]. PMID- 10702084 TI - [The juridical aspects of quality management (quality assurance) in dentistry]. PMID- 10702085 TI - Assessment of the cardiovascular system at the workplace. PMID- 10702086 TI - Cardiovascular evaluation of the worker and workplace: a practical guide for clinicians. AB - Unlike several other branches of medicine (e.g., pulmonology), primary cardiology has yet to fully develop a discipline of occupational cardiology. The authors outline an approach for including a focused occupational history in the CV work up and present a graded, risk-stratified algorithm for occupational cardiologic assessment. This work-up can help clinicians make specific recommendations concerning working conditions, as these impact upon the patient's CV status. PMID- 10702087 TI - Clinical issues: return to work and public safety. AB - Return to work after cardiac events is an especially delicate question for the clinician. The cardiologic caregiver must evaluate the full clinical picture. including symptoms and morphological and functional status, as well as address complex personal. psychological, social, economic, legal. and ethical issues. The importance of job characteristics is illustrated by the existing, albeit limited, longitudinal data showing that return to high-strain work is a significant independent predictor of mortality in young men post-myocardial infarction. PMID- 10702088 TI - Screening and management of the workplace for CVD risk. PMID- 10702089 TI - Costs of occupational circulatory disease. AB - This chapter analyzes the overt and hidden economic costs of work-related cardiovascular diseases Affected individuals and society at large, rather than employers, pay much of the costs. Economic interventions to motivate public health approaches to preventing CVD are explored. PMID- 10702090 TI - Legal and legislative issues. PMID- 10702091 TI - Workplace intervention studies. AB - The author reviews and discusses several intervention studies with implications for CVD or CV risk. The studies address chemical exposures. work schedules and working hours, and psychosocial factors at work. Effective strategies for prevention of CVD at the workplace are based on intervention research and integrate prevention at different levels. PMID- 10702092 TI - The workplace and cardiovascular health: conclusions and thoughts for a future agenda. AB - The evidence in this book provides convergent validation of a causal relationship between workplace stressors and CVD. Here, the editors explore new strategies for enhanced prevention and clinical management, work place interventions, and social policy to reduce the impact of CVD. These strategies acquire an urgent public health dimension, given the magnitude of the CVD epidemic and the current deterioration in conditions of working life. PMID- 10702093 TI - Silica dust and lung cancer. PMID- 10702094 TI - Re-estimated equations for 5th percentiles of lung function variables. PMID- 10702095 TI - [Guidelines for diagnosis and therapy of disorders in sexual differentiation. German Society of Urology]. PMID- 10702096 TI - [Dissertations defended in 1999]. PMID- 10702097 TI - [Lieutenant General of the Medical Service Evgenii Vladislavovich Gembitskii (on the 80th anniversary of his birth)]. PMID- 10702098 TI - [A researcher on the demographic consequences of wars (on the 75th anniversary of the birth of L. E. Poliakov)]. PMID- 10702099 TI - [Outstanding Russian surgeon Iu. K. Shimanovskii]. PMID- 10702100 TI - [The 80th anniversary of the Epidemiological Health Detachment of the Volga Military District]. PMID- 10702101 TI - [The 40th anniversary of the Rest Base for Submariners of the Northern Fleet]. PMID- 10702102 TI - Proceedings of the World Conference on Low-Calorie Sweeteners. Barcelona, Spain, April 25-28, 1999. PMID- 10702103 TI - 16th Meeting of the European Intestinal Transport Group. Bad Herrenalb, Germany, September 19-20, 1999. Abstracts. PMID- 10702104 TI - [Symposium of the German Section of the International Society for the Study of Hypertension in Pregnancy (ISSHP). Heidelberg, 22-23 May 1998. Proceedings]. PMID- 10702105 TI - Premedication with midazolam in pediatric anesthesia. PMID- 10702106 TI - Socioeconomic Status and Health in Industrial Nations: Social, Psychological, and Biological Pathways. Bethesda, Maryland, USA. May 10-12, 1999. Proceedings. PMID- 10702107 TI - A randomised controlled trial of specialist health visitor intervention for failure to thrive. PMID- 10702108 TI - Definitive diagnosis of nut allergy. PMID- 10702109 TI - Interhospital transfer of sick children: proposal for a unified approach. PMID- 10702110 TI - RSV prevention. PMID- 10702111 TI - Varicella: to vaccinate or not vaccinate? PMID- 10702112 TI - Androgen secreting adrenocortical carcinomas. PMID- 10702113 TI - Fatal chickenpox: negative electron microscopy of vesicular samples may be misleading. PMID- 10702114 TI - Effect of cellular interaction on glycolytic oscillations in yeast: a theoretical investigation. AB - On the basis of a detailed model of yeast glycolysis, the effect of intercellular dynamics is analysed theoretically. The model includes the main steps of anaerobic glycolysis, and the production of ethanol and glycerol. Transmembrane diffusion of acetaldehyde is included, since it has been hypothesized that this substance mediates the interaction. Depending on the kinetic parameter, the single-cell model shows both stationary and oscillatory behaviour. This agrees with experimental data with respect to metabolite concentrations and phase shifts. The inclusion of intercellular coupling leads to a variety of dynamical modes, such as synchronous oscillations, and different kinds of asynchronous behavior. These oscillations can co-exist, leading to bi- and tri-rhythmicity. The corresponding parameter regions have been identified by a bifurcation analysis. The oscillatory dynamics of synchronized cell populations are investigated by calculating the phase responses to acetaldehyde pulses. Simulations are performed with respect to the synchronization of two subpopulations that are oscillating out of phase before mixing. The effect of the various process on synchronization is characterized quantitatively. While continuous exchange of acetaldehyde might synchronize the oscillations for appropriate sets of parameter values, the calculated synchronization time is longer than that observed experimentally. It is concluded either that addition to the transmembrane exchange of acetaldehyde, other processes may contribute to intercellular coupling, or that intracellular regulator feedback plays a role in the acceleration of the synchronization. for appropriate sets of parameter values, the calculated synchronization time is longer than that observed experimentally. It is concluded either that addition to the transmembrane exchange of acetaldehyde, other processes may contribute to intercellular coupling, or that intracellular regulator feedback plays a role in the acceleration of the synchronization. PMID- 10702115 TI - Treatment of oral cancer. Biopsy under local anaesthetic is inadequate. PMID- 10702116 TI - Moving beyond journals. Physicists do it in large groups. PMID- 10702117 TI - Changing perceptions in osteoporosis. Replacing bone mineral density with bone turnover is not a solution. PMID- 10702118 TI - Caltech Faint Galaxy Redshift Survey. XI. The Merger Rate to Redshift 1 from Kinematic Pairs. AB - The rate of mass accumulation due to galaxy merging depends on the mass, density, and velocity distribution of galaxies in the near neighborhood of a host galaxy. The fractional luminosity in kinematic pairs combines all of these effects in a single estimator that is relatively insensitive to population evolution. Here we use a k-corrected and evolution-compensated volume-limited sample having an R band absolute magnitude of Mk,eR/=0.2M*) is 0.02+/-0.01&parl0;1+z&parr0;0.1+/-0.5M* Gyr-1. Present-day high-luminosity galaxies therefore have accreted approximately 0.15M* of their mass over the approximately 7 Gyr to redshift 1. Since merging is likely only weakly dependent on the host mass, the fractional effect, deltaM&solm0;M approximately 0.15M*&solm0;M, is dramatic for lower mass galaxies but is, on the average, effectively perturbative for galaxies above 1M*. PMID- 10702119 TI - Cosmic Shear from Galaxy Spins. AB - We discuss the origin of galactic angular momentum and the statistics of the present-day spin distribution. It is expected that the galaxy spin axes are correlated with the intermediate principal axis of the gravitational shear tensor. This allows one to reconstruct the shear field and thereby the full gravitational potential from the observed galaxy spin fields. We use the direction of the angular momentum vector without any information of its magnitude, which requires a measurement of the position angle and inclination on the sky of each disk galaxy. We present the maximum likelihood shear inversion procedure, which involves a constrained linear minimization. The theory is tested against numerical simulations. We find the correlation strength of nonlinear structures with the initial shear field and show that accurate large-scale density reconstructions are possible at the expected noise level. PMID- 10702120 TI - Stochastic Acceleration and Nonthermal Radiation in Clusters of Galaxies. AB - We calculate the distribution of electrons in clusters of galaxies, which results from thermalization processes in the presence of stochastic acceleration due to plasma waves. We show that the electron distribution can deviate from a Maxwell Boltzmann distribution because of the effect of the stochastic energy gain, provided that waves can be sustained against damping. The bremsstrahlung emission of the nonthermal tail of electrons can result in a flux of hard X-rays that is compatible with the ones recently detected in some clusters of galaxies. PMID- 10702121 TI - Superwind Model of Extended Lyalpha Emitters at High Redshift. AB - We propose a new model for the extended Lyalpha blobs found recently at high redshift (z approximately 3). The observational properties of these blobs are as follows: (1) the observed Lyalpha luminosities are approximately 1043 h-2 ergs s 1, (2) they appear elongated morphologically, (3) their sizes amount to approximately 100 kpc, (4) the observed line widths amount to approximately 1000 km s-1, and (5) they are not associated with strong radio continuum sources. All these observational properties seem to be explained in terms of galactic winds driven by successive supernova explosions shortly after the initial burst of massive star formation in the galactic centers. The observed number density of Lyalpha blobs ( approximately 3.4x10-5 h3 Mpc-3) may be explained if their present-day counterparts are elliptical galaxies with a luminosity above approximately 1L*. PMID- 10702122 TI - The Variability of Seyfert 1.8 and 1.9 Galaxies at 1.6 Microns. AB - We present a study of Seyfert 1.5-2.0 galaxies observed at two epochs with the Hubble Space Telescope (HST) at 1.6 um. We find that unresolved nuclear emission from nine of 14 nuclei varies at the level of 10%-40% on timescales of 0.7-14 months, depending upon the galaxy. A control sample of Seyfert galaxies lacking unresolved sources and galaxies lacking Seyfert nuclei show less than 3% instrumental variation in equivalent aperture measurements. This proves that the unresolved sources are nonstellar and associated with the central parsecs of active galactic nuclei. Unresolved sources in Seyfert 1.8 and 1.9 galaxies are not usually detected in HST optical surveys; however, high angular resolution infrared observations will provide a way to measure time delays in these galaxies. PMID- 10702123 TI - The Mid-Infrared Spectra of Normal Galaxies. AB - The mid-infrared spectra (2.5-5 and 5.7-11.6 um) obtained by ISOPHOT reveal the interstellar medium emission from galaxies powered by star formation to be strongly dominated by the aromatic features at 6.2, 7.7, 8.6, and 11.3 um. Additional emission appears in between the features, and an underlying continuum is clearly evident at 3-5 um. This continuum would contribute about a third of the luminosity in the 3-13 um range. The features together carry 5%-30% of the 40 120 um far-infrared (FIR) luminosity. The relative fluxes in individual features depend very weakly on galaxy parameters such as the far-infrared colors, direct evidence that the emitting particles are not in thermal equilibrium. The dip at 10 um is unlikely to result from silicate absorption since its shape is invariant among galaxies. The continuum component has a fnu~nu+0.65 shape between 3 and 5 um and carries 1%-4% of the FIR luminosity; its extrapolation to longer wavelengths falls well below the spectrum in the 6-12 um range. This continuum component is almost certainly of nonstellar origin and is probably due to fluctuating grains without aromatic features. The spectra reported here typify the integrated emission from the interstellar medium of the majority of star forming galaxies and could thus be used to obtain redshifts of highly extincted galaxies up to z=3 with SIRTF. PMID- 10702124 TI - Prompt Optical Observations of Gamma-Ray Bursts. AB - The Robotic Optical Transient Search Experiment (ROTSE) seeks to measure simultaneous and early afterglow optical emission from gamma-ray bursts (GRBs). A search for optical counterparts to six GRBs with localization errors of 1 deg2 or better produced no detections. The earliest limiting sensitivity is mROTSE>13.1 at 10.85 s (5 s exposure) after the gamma-ray rise, and the best limit is mROTSE>16.0 at 62 minutes (897 s exposure). These are the most stringent limits obtained for the GRB optical counterpart brightness in the first hour after the burst. Consideration of the gamma-ray fluence and peak flux for these bursts and for GRB 990123 indicates that there is not a strong positive correlation between optical flux and gamma-ray emission. PMID- 10702125 TI - Binary Black Hole Mergers from Planet-like Migrations. AB - If supermassive black holes (BHs) are generically present in galaxy centers, and if galaxies are built up through hierarchical merging, BH binaries are at least temporary features of most galactic bulges. Observations suggest, however, that binary BHs are rare, pointing toward a binary lifetime far shorter than the Hubble time. We show that, almost regardless of the detailed mechanism, all stellar dynamical processes are too slow in reducing the orbital separation once orbital velocities in the binary exceed the virial velocity of the system. We propose that a massive gas disk surrounding a BH binary can effect its merger rapidly, in a scenario analogous to the orbital decay of super-Jovian planets due to a proto-planetary disk. As in the case of planets, gas accretion onto the secondary (here a supermassive BH) is integrally connected with its inward migration. Such accretion would give rise to quasar activity. BH binary mergers could therefore be responsible for many or most quasars. PMID- 10702126 TI - Faint Infrared Flares from the Microquasar GRS 1915+105. AB - We present simultaneous infrared and X-ray observations of the Galactic microquasar GRS 1915+105 using the Palomar 5 m telescope and Rossi X-Ray Timing Explorer on 1998 July 10 UT. Over the course of 5 hr, we observed six faint infrared (IR) flares with peak amplitudes of approximately 0.3-0.6 mJy and durations of approximately 500-600 s. These flares are associated with X-ray soft dip/soft-flare cycles, as opposed to the brighter IR flares associated with X-ray hard-dip/soft-flare cycles seen in 1997 August by Eikenberry et al. Interestingly, the IR flares begin before the X-ray oscillations, implying an "outside-in" origin of the IR/X-ray cycle. We also show that the quasi-steady IR excess in 1997 August is due to the pileup of similar faint flares. We discuss the implications of this flaring behavior for understanding jet formation in microquasars. PMID- 10702127 TI - Magnitude Bias of Microlensed Sources toward the Large Magellanic Cloud. AB - There are lines of evidence suggesting that some of the observed microlensing events in the direction of the Large Magellanic Cloud (LMC) are caused by ordinary star lenses as opposed to dark MACHOs in the Galactic halo. Efficient lensing by ordinary stars generally requires the presence of one or more additional concentrations of stars along the line of sight to the LMC disk. If such a population behind the LMC disk exists, then the source stars (for lensing by LMC disk objects) will be drawn preferentially from the background population and will show systematic differences from LMC field stars. One such difference is that the (lensed) source stars will be farther away than the average LMC field stars, and this should be reflected in their apparent baseline magnitudes. We focus on red clump stars; these should appear in the color-magnitude diagram at a few tenths of a magnitude fainter than the field red clump. Suggestively, one of the two near-clump confirmed events, MACHO-LMC-1, is a few tenths of magnitude fainter than the clump. PMID- 10702128 TI - Discovery of High Proper-Motion Ancient White Dwarfs: Nearby Massive Compact Halo Objects? AB - We present the discovery and spectroscopic identification of two very high proper motion ancient white dwarf stars, found in a systematic proper-motion survey. Their kinematics and apparent magnitude clearly indicate that they are halo members, while their optical spectra are almost identical to the recently identified cool halo white dwarf WD 0346+246. Canonical stellar halo models predict a white dwarf volume density that is 2 orders of magnitude less than the rho approximately 7x10-4 M middle dot in circle pc-3 inferred from this survey. With the caveat that the sample size is very small, it appears that a significant fraction, approximately 10%, of the local dark matter halo is in the form of very old, cool, white dwarfs. PMID- 10702129 TI - Formation of Short-Period Binary Pulsars in Globular Clusters. AB - We present a new dynamical scenario for the formation of short-period binary millisecond pulsars in globular clusters. Our work is motivated by the recent observations of 20 radio pulsars in 47 Tuc. In a dense cluster such as 47 Tuc, most neutron stars acquire binary companions through exchange interactions with primordial binaries. The resulting systems have semimajor axes in the range approximately 0.1-1 AU and neutron star companion masses approximately 1-3 M middle dot in circle. For many of these systems, we find that when the companion evolves off the main sequence and fills its Roche lobe, the subsequent mass transfer is dynamically unstable. This leads to a common envelope phase and the formation of short-period neutron star-white dwarf binaries. For a significant fraction of these binaries, the decay of the orbit due to gravitational radiation will be followed by a period of stable mass transfer driven by a combination of gravitational radiation and tidal heating of the companion. The properties of the resulting short-period binaries match well those of observed binary pulsars in 47 Tuc. PMID- 10702130 TI - Detection of Planetary Transits of the Star HD 209458 in the Hipparcos Data Set. AB - A search of the Hipparcos satellite photometry data for the star HD 209458 reveals evidence for a planetary transit signature consistent with the planetary properties reported by Henry et al. and Charbonneau et al. and allows further refinement of the planet's orbital period. The long time baseline (about 2926 days or 830 periods) from the best Hipparcos transit-like event to the latest transit reported by Henry et al. for the night of 1999 November 15 (UT) allows for an orbital period determination of 3.524736 days with an uncertainty of 0.000045 days (3.9 s). The transit events observed by Charbonneau et al. fall at the interim times expected to within the errors of this newly derived period. A series of statistical tests was performed to assess the likelihood of these events occurring by chance. This was crucial given the ill-conditioned problem presented by the sparse sampling of the light curve and the non-Gaussian distribution of the points. Monte Carlo simulations using bootstrap methods with the actual Hipparcos HD 209458 data set indicate that the transit-like signals of the depth observed would only be produced by chance in 21 out of 1 million trials. The transit durations and depths obtained from the Hipparcos data are also consistent with those determined by Charbonneau et al. and Henry et al. within the limitations of the sampling intervals and photometric precision of the Hipparcos data. PMID- 10702132 TI - Equilibrium Chemistry in a Brown Dwarf's Atmosphere: Cesium in Gliese 229B. AB - The distribution of Cs in Gliese 229B's atmosphere reveals how equilibrium chemistry establishes the atmospheric composition. The rapid kinetics of cesium chemistry keeps the Cs abundance in thermochemical equilibrium and renders Cs a sensitive measure of chemical processes in brown dwarf atmospheres. Observations of Gliese 229B indicate a subsolar bulk abundance of Cs, the depletion of alkali metals in the upper atmosphere from condensation, and a partitioning of heavy elements different from that of the Sun. PMID- 10702131 TI - The Spectroscopic Orbit of the Planetary Companion Transiting HD 209458. AB - We report a spectroscopic orbit with period P=3.52433+/-0.00027 days for the planetary companion that transits the solar-type star HD 209458. For the metallicity, mass, and radius of the star, we derive [Fe/H&sqbr0;=0.00+/-0.02, M*=1.1+/-0.1 M middle dot in circle, and R*=1.2+/-0.1 R middle dot in circle. This is based on a new analysis of the iron lines in our HIRES template spectrum and also on the absolute magnitude, effective temperature, and color of the star, and it uses isochrones from four different sets of stellar evolution models. Using these values for the stellar parameters, we reanalyze the transit data and derive an orbital inclination of i=86&fdg;1+/-1&fdg;6. For the planet, we derive a mass of Mp=0.69+/-0.05 MJup, a radius of Rp=1.40+/-0.17 RJup, and a density of rho=0.31+/-0.07 g cm-3. PMID- 10702133 TI - Interstellar Scintillations of Polarization of Compact Sources. AB - We demonstrate that the measurement of fluctuations of polarization due to the Galactic interstellar scintillations may be used to study the structure of the radiation field at compact radio sources. We develop a mathematical formalism and demonstrate it on a simple analytical model in which the scale of the polarization variation through the source is comparable to the source size. The predicted amplitude of modulation of the polarized radiation flux is approximately 20%pismsc, where pis is the characteristic degree of polarization of radiation at the source and msc is the typical modulation index due to scattering, i.e., msc approximately 1 for diffractive scintillations and msc<1 for refractive scintillations. PMID- 10702134 TI - Global Simulations of Differentially Rotating Magnetized Disks: Formation of Low beta Filaments and Structured Coronae. AB - We present the results of three-dimensional global magnetohydrodynamic simulations of the Parker-shearing instability in a differentially rotating torus initially threaded by toroidal magnetic fields. An equilibrium model of a magnetized torus is adopted as an initial condition. When beta0=Pgas&solm0;Pmag approximately 1 at the initial state, magnetic flux buoyantly escapes from the disk and creates looplike structures similar to those in the solar corona. Inside the torus, the growth of nonaxisymmetric magnetorotational (or Balbus & Hawley) instability generates magnetic turbulence. Magnetic field lines are tangled on a small scale, but on a large scale they show low azimuthal wavenumber spiral structure. After several rotation periods, the system oscillates around a state with beta approximately 5. We found that magnetic pressure-dominated (beta<1) filaments are created in the torus. The volume filling factor of the region in which beta50% (> or =56-65% of energy) of their subsistence from animal foods, whereas only 14% of these societies derived >50% (> or =56-65% of energy) of their subsistence from gathered plant foods. This high reliance on animal-based foods coupled with the relatively low carbohydrate content of wild plant foods produces universally characteristic macronutrient consumption ratios in which protein is elevated (19-35% of energy) at the expense of carbohydrates (22-40% of energy). PMID- 10702161 TI - Overweight among children and adolescents in a Native Canadian community: prevalence and associated factors. AB - BACKGROUND: The prevalence of pediatric obesity in North America is increasing. Native American children are at especially high risk. OBJECTIVES: The objective was to evaluate the prevalence of pediatric overweight and associated behavioral factors in a Native Canadian community with high rates of adult obesity and type 2 diabetes mellitus. DESIGN: Height and weight were measured in 445 children and adolescents aged 2-19 y. Fitness level, television viewing, body image concepts, and dietary intake were assessed in 242 subjects aged 10-19 y. Overweight was defined as a body mass index > or =85th percentile value for age- and sex specific reference data from the third National Health and Nutrition Examination Survey (NHANES III). Multiple logistic regression was used to examine factors associated with overweight, with adjustment for age and sex. RESULTS: The overall prevalence of overweight in subjects aged 2-19 y was significantly higher than NHANES III reference data [boys: 27. 7% (95% CI: 21.8, 34.5); girls: 33.7% (95% CI: 27.9, 40.1)]. In the subset aged 10-19 y, > or =5 h television viewing/d was associated with a significantly higher risk of overweight than was < or =2 h/d [odds ratio (OR) = 2.52; 95% CI: 1.06, 5.98]. Subjects in the third and fourth quartiles of fitness had a substantially lower risk of overweight than did those in the first quartile [third quartile compared with first quartile: OR = 0.24 (95% CI: 0.09, 0.66); fourth quartile compared with first quartile: OR = 0.13 (95% CI: 0.03, 0. 48)]. Fiber consumption on the previous day was associated with a decreased risk of overweight (OR = 0.69; 95% CI: 0.47, 0.99 for each 0.77 g/MJ increase in fiber intake). CONCLUSIONS: Pediatric overweight is a harbinger of future diabetes risk and indicates a need for programs targeting primary prevention of obesity in children and adolescents. PMID- 10702162 TI - Dietary medium-chain triacylglycerol prevents the postprandial rise of plasma triacylglycerols but induces hypercholesterolemia in primary hypertriglyceridemic subjects. AB - BACKGROUND: Previous studies showed divergent results concerning the influence of medium-chain triacylglycerol (MCT) on lipoprotein metabolism. OBJECTIVE: The objective of this study was to compare the effects of MCT and corn oil on plasma lipids in primary hypertriglyceridemic patients. DESIGN: Ten subjects ate different proportions of corn oil and MCT for 12 wk. The subjects first ate a low fat diet for 2 wk and during the next 4 wk, corn oil was added as the sole source of fat. Thereafter, for 2-wk periods, the subjects were sequentially fed corn oil and MCT mixed in the following proportions: 3:1, 1:1, and 0:1. Fasting plasma total cholesterol, triacylglycerol, and HDL-cholesterol concentrations were measured at the end of each period. At the end of the 100%-corn oil and of the 100%-MCT periods, subjects were fed a test meal containing the respective oil (40 g fat/m(2) body surface area) and total cholesterol and triacylglycerols were measured at 2-h intervals over 8 h; fasting lipoprotein composition was also measured. RESULTS: Compared with corn oil, MCT was associated with a higher mean (+/-SD) fasting total cholesterol concentration (6.39 +/- 1.14 compared with 5.51 +/- 0.98 mmol/L, respectively; P < 0. 05); non-HDL-cholesterol concentrations were also higher with MCT (5. 36 +/- 1.11 mmol/L) than with corn oil (4.51 +/- 0.92 mmol/L; P < 0. 005). In response to the liquid test meal, plasma total cholesterol did not change with either diet but triacylglycerols increased with the 100%-corn oil diet. CONCLUSIONS: MCT prevents the risk of pancreatitis due to postprandial hypertriglyceridemia but has the inconvenience of raising total cholesterol concentrations in primary hypertriglyceridemic subjects. PMID- 10702163 TI - Changes in plasma lipids and other cardiovascular risk factors during 3 energy restricted diets differing in total fat and fatty acid composition. AB - BACKGROUND: The well-established relation between changes in dietary fatty acids and plasma lipids has been determined in energy-balance states. Whether this relation is altered in states of energy restriction and active weight loss is not clear. OBJECTIVE: The objective of this 12-wk study was to compare the time course of lipid changes and other cardiovascular risk factors in 3 energy restricted diets (all 6500 kJ) with different total fat and fatty acid compositions. DESIGN: Sixty-two subjects with a body mass index (in kg/m(2)) >24 were stratified into 1 of 3 parallel dietary intervention groups: 1) a very-low fat (VLF) diet (10% of energy from fat; 3% from saturated fat), 2) a high saturated-fat (HSF) diet (32% of energy from fat; 17% from saturated fat), and 3) a high-unsaturated-fat (HUF) diet (32% of energy from fat; 6% from saturated fat). RESULTS: After 12 wk, LDL cholesterol decreased by 0. 66 +/- 0.11 (mean +/- SEM) and 0.68 +/- 0.12 mmol/L ( approximately 20%) with the VLF and HUF diets, respectively, compared with a decrease of only 0.24 +/- 0.11 mmol/L (7%) with the HSF diet (P < 0.02 between groups). Diet affected the time course of changes in HDL cholesterol with both high-fat diets, resulting in smaller reductions in HDL cholesterol at weeks 1 (P = 0.0004) and 4 (P = 0.02); however, these differences were no longer apparent by 12 wk. Overall weight loss was 8.6 +/- 0.4 kg (9.7%) and waist circumference decreased by 7.3 +/- 5 cm (8%) for the combined groups, with no significant differences between diets. CONCLUSIONS: Significantly greater decreases in LDL cholesterol during active weight loss are achieved with diets low in saturated fatty acids. Changes in HDL cholesterol between diets appear dependent on both the fat content of the diet and the duration of energy restriction. PMID- 10702164 TI - Apolipoprotein B gene polymorphisms and serum lipids: meta-analysis of the role of genetic variation in responsiveness to diet. AB - BACKGROUND: The genetic variance determining plasma lipid and lipoprotein concentrations may modify individual responsiveness to alterations in dietary fat and cholesterol content. OBJECTIVE: The aim was to examine the role of apolipoprotein (apo) B DNA polymorphisms in responsiveness of plasma lipids and lipoproteins to diet. DESIGN: A controlled dietary intervention study was conducted in 44 healthy, middle-aged subjects with a 3-mo baseline, a 1-mo fat controlled, a 1-mo high-fat, and a 1-mo habitual diet period. We also conducted a meta-analysis of all published dietary trials, including our own. RESULTS: In our own dietary study, the apo B XbaI restriction-site polymorphism affected the responsiveness to diet of the plasma LDL-cholesterol concentration (P < 0.05, repeated-measures analysis of variance). Especially during the high-fat diet, homozygous absence of the XbaI restriction site (X(-)/X(-)) was associated with a greater increase in LDL cholesterol (44 +/- 5%) than was X(+)/X(+) (27 +/- 7%) or X(+)/X(-) (40 +/- 5%). The high-fat diet also induced a larger increase in plasma LDL cholesterol in subjects with the R(-)/R(-) genotype (homozygous absence of the EcoRI restriction site) (59 +/- 10%) than in those with the R(+)/R(-) (39 +/- 6%) or R(+)/R(+) (36 +/- 4%) genotype. The M(+)/M(+) genotype (homozygous presence of the MspI restriction site) was also more responsive (41 +/- 3% increase in LDL cholesterol) than the M(+)/M(-) genotype (27 +/- 10% increase). The meta-analysis supported the finding of the significant role of the EcoRI and MspI polymorphisms, but not that of the XbaI polymorphism. CONCLUSIONS: The present study indicated that the apo B EcoRI and MspI polymorphisms are associated with responsiveness to diet. PMID- 10702165 TI - Role of dietary factors in ethnic differences in early risk of cardiovascular disease and type 2 diabetes. AB - BACKGROUND: The disparity in the prevalence of cardiovascular disease and type 2 diabetes between African Americans and whites has been well established, and ethnic differences in several risk factors for these diseases are evident in childhood. OBJECTIVE: The current study explored whether dietary factors explain ethnic differences in serum lipids and insulin profiles in children, independent of body composition and social class background. DESIGN: The sample included 95 African American and white children (mean age: 10.0 y). Macronutrient and food group intakes were derived from three 24-h recalls. Cardiovascular disease and type 2 diabetes risk were determined on the basis of total cholesterol, triacylglycerol, insulin sensitivity (S(i)), and acute insulin response (AIR). Data were analyzed by using t tests, analysis of covariance, and multiple regression. RESULTS: African American children had lower triacylglycerol (P < 0.01), lower S(i) (P < 0.001), and higher AIR (P < 0.001) than whites. Intake of fruit and vegetables was significantly higher, and dairy intake lower, in African American than in white children after adjustment for social class and total energy intake. Several direct relations were observed between diet and insulin action: carbohydrate and fruit intakes were positively associated with S(i) (P = 0.02), and vegetable intake was negatively associated with AIR (P = 0.01). However, neither macronutrient nor food group intake accounted for the ethnic differences in triacylglycerol and AIR. CONCLUSIONS: The African American children in our sample showed a greater disease risk than did the white children, even after body composition, social class background, and dietary patterns were adjusted for. PMID- 10702166 TI - Skeletal muscle weakness is associated with wasting of extremity fat-free mass but not with airflow obstruction in patients with chronic obstructive pulmonary disease. AB - BACKGROUND: Skeletal muscle weakness is a prominent problem in many patients with chronic obstructive pulmonary disease (COPD). OBJECTIVE: The aim of the study was to determine the relation between skeletal muscle function, body composition, and lung function in COPD (emphysema and chronic bronchitis) patients and healthy volunteers. DESIGN: In 50 patients with chronic bronchitis, 49 patients with emphysema, and 28 healthy volunteers, skeletal muscle function was assessed by handgrip and linear isokinetic dynamometry. Whole-body and subregional fat-free mass (FFM) were assessed by dual-energy X-ray absorptiometry. RESULTS: Whole-body and extremity FFM were significantly lower in patients with emphysema (P < 0.001) and chronic bronchitis (P < 0.05) than in healthy volunteers, but trunk FFM was significantly lower only in patients with emphysema (P < 0.001). Extremity FFM was not significantly different between the COPD subtype groups, despite significantly lower values for whole-body and trunk FFM (P < 0.05) in patients with emphysema. Absolute skeletal muscle function (P < 0. 001) and muscle function per kilogram of whole-body FFM were significantly lower in both COPD subtype groups than in healthy volunteers (P < 0.05), but no significant difference was found between patients with chronic bronchitis and those with emphysema. Muscle function per kilogram of extremity FFM was not significantly different between the 3 groups and was not associated with forced expiratory volume in 1 s. CONCLUSION: Skeletal muscle weakness is associated with wasting of extremity FFM in COPD patients, independent of airflow obstruction and COPD subtype. PMID- 10702167 TI - Psychological measures of eating behavior and the accuracy of 3 common dietary assessment methods in healthy postmenopausal women. AB - BACKGROUND: Factors affecting the accuracy of reported energy intake (rEI) need to be identified. OBJECTIVE: Our objective was to investigate the association of psychological measures of eating behavior with the accuracy of rEI assessed by 7 d weighed intakes, a 24-h recall, and a food-frequency questionnaire. DESIGN: Subjects were 26 restrained eaters aged 60.3 +/- 0.6 y (mean +/- SEM) and weighing 63.8 +/- 1.7 kg and 34 unrestrained eaters aged 59.4 +/- 0.6 y and weighing 64.0 kg. rEI was assessed by using 3 dietary assessment methods and total energy expenditure (TEE) was determined by using doubly labeled water. Calculated EI (cEI) was determined as TEE corrected for the estimated change in body energy. Subjects completed the Eating Inventory. RESULTS: rEI values were significantly lower than TEE values for all 3 dietary assessment methods (P < 0.05); there was no significant relation between rEI and TEE by any method. There was no significant difference in 100 x rEI:TEE between restrained and unrestrained eaters by any of the dietary assessment methods. When combined data from the 3 methods were used, 100 x rEI:cEI was not significantly different from 100% in unrestrained eaters (99 +/- 6.8%) but was lower in restrained eaters (89.1 +/- 5.3%; P < 0.05). There was a positive relation between hunger and 100 x rEI:TEE (P < 0.05). CONCLUSIONS: Low hunger is associated with undereating relative to normal eating during measurement of dietary intake; high dietary restraint may be associated with a reduction in reporting of consumed foods. Dietary hunger and restraint assessed with use of the Eating Inventory may help to identify subjects likely to underreport dietary intake. PMID- 10702168 TI - Lack of efficacy of a food-frequency questionnaire in assessing dietary macronutrient intakes in subjects consuming diets of known composition. AB - BACKGROUND: We compared the validity of a semiquantitative food-frequency questionnaire in assessing intakes of macronutrients (absolute amounts and percentages of energy) by 19 subjects fed natural-food diets of known composition. In small subsets (n = 5 or 6), we also tested 3-d diet records. OBJECTIVE: The objective of this study was to investigate the efficacy of food frequency questionnaires and diet records in subjects fed natural-food diets of known composition. DESIGN: Each subject consumed 3 different diets for >/=6 wk and self-reported his or her food intake by using a food-frequency questionnaire and a diet record. The diets varied in their chemically analyzed contents of fat (15-35% energy), saturated fat (5-14%), monounsaturated fat (5-14.5%), polyunsaturated fat (2.5-10.5%), carbohydrate (49-68%), and cholesterol (108-348 mg/d). RESULTS: The food-frequency questionnaire significantly underestimated fat, saturated fat, monounsaturated fat, and protein intakes and significantly overestimated carbohydrate intake with the high-fat diet. The percentage of energy from fat was significantly underestimated for the high-fat diet and significantly overestimated for the very-low-fat diet. Estimates from the food frequency questionnaire differed significantly from actual intakes for fat (absolute and percentage), saturated fat (absolute and percentage), monounsaturated fat (absolute and percentage), and protein (percentage) in the high-fat diet and for polyunsaturated fat (absolute and percentage), saturated fat (percentage), fiber (absolute), and cholesterol (daily absolute; in mg/d) in the lower-fat diet. Estimates from the diet records better agreed with actual intakes than did estimates from the food-frequency questionnaire except for monounsaturated fat (absolute and percentage) in the high-fat diet and polyunsaturated fat (percentage) in the lower-fat diet and the very-low-fat diet. CONCLUSION: Our data indicated that the food-frequency questionnaire did not provide reliable estimates of absolute amounts of dietary fats or cholesterol. PMID- 10702169 TI - Energy expenditure at rest and during sleep in children with Prader-Willi syndrome is explained by body composition. AB - BACKGROUND: Obesity in Prader-Willi syndrome (PWS) seems to be related to a low basal metabolic rate (BMR). In addition, abnormal sleep patterns reported in PWS might affect sleeping metabolic rate (SMR). OBJECTIVE: Our objective was to assess BMR and SMR adjusted for fat-free mass in young PWS patients. DESIGN: Subjects were 17 PWS patients (10 females and 7 males aged 7.5-19.8 y) and 17 obese control subjects matched for sex and bone age. SMR was measured in a respiratory chamber, BMR with a ventilated-hood system, and body composition by deuterium dilution. RESULTS: BMR and SMR were significantly lower in the PWS group than in the control group (5.36 +/- 1.18 and 4.62 +/- 1.08 MJ/d compared with 6.38 +/- 1.55 and 5.60 +/- 1.52 MJ/d, respectively; P < 0.05). When fat-free mass was included in the analysis, multiple regression showed no differences in BMR and SMR between groups. When weight was included in the analysis instead of fat-free mass, SMR was lower in the PWS group. Fat-free mass was lower in the PWS group both as an absolute value and when adjusted for height. CONCLUSION: BMR and SMR are low in young patients with PWS because of a low fat-free mass. PMID- 10702170 TI - Threonine requirement of young men determined by indicator amino acid oxidation with use of L-[1-(13)C]phenylalanine. AB - BACKGROUND: Threonine is an indispensable amino acid with a complex degradative pathway. Use of the indicator amino acid oxidation technique should provide an estimate of the threonine requirement that is not affected by its metabolic pathway. OBJECTIVE: Our objective was to determine the requirement for threonine in men by using the indicator amino acid oxidation method and to provide statistical estimates of the population mean and 95% CIs of the threonine requirement. We hypothesized that the current World Health Organization estimate of the threonine requirement, 7 mg*kg(-)(1)*d(-)(1) (based on nitrogen balance studies), is too low. DESIGN: Six healthy men each received 6 different threonine intakes while consuming an energy-sufficient diet with 1.0 g L-amino acid mixture*kg(-)(1)*d(-)(1). The effect of graded alterations in dietary threonine intake on phenylalanine flux and oxidation was studied by using L-[1 (13)C]phenylalanine as the indicator amino acid. RESULTS: The results of two phase linear regression crossover analysis showed that the mean threonine requirement, based on indicator oxidation, was 19.0 mg*kg(-)(1)*d(-)(1) with an upper safe intake of 26.2 mg*kg(-)(1)*d(-)(1). CONCLUSIONS: This is the first application of the indicator amino acid oxidation technique in humans to study the requirement for an indispensable amino acid with a complex degradative pathway. We found that the upper safe intake for 95% of the population is almost 4-fold higher than the current World Health Organization estimate. PMID- 10702171 TI - Amino acid losses during hemodialysis with polyacrylonitrile membranes: effect of intradialytic amino acid supplementation on plasma amino acid concentrations and nutritional variables in nondiabetic patients. AB - BACKGROUND: Malnutrition is highly prevalent in hemodialysis patients. Amino acid (AA) losses during the dialysis procedure may be a contributing factor. OBJECTIVES: The objectives of this study were 1) to prospectively evaluate AA losses and their effect on plasma AA concentrations during dialysis with polyacrylonitrile at baseline and after administration of AAs by intradialysis and 2) to investigate the effects of intradialytic AA supplementation on nutritional status. DESIGN: Seventeen stable patients without diabetes who were receiving hemodialysis were studied. In the first phase, AA losses were evaluated over 2 wk in 10 patients randomly assigned to receive AA supplementation. AA losses were analyzed during the first week without supplementation and during the second week with AA administration. In the second phase, the patients' nutritional status was investigated after 3 mo of AA supplementation and was compared with those in 7 patients not receiving AAs. RESULTS: Mean +/- SD) AA losses during a 4-h dialysis session were 12 +/- 2 g; there was a significant decrease in plasma AA concentrations (386 +/- 298 micromol/L for essential and 902 +/- 735 micromol/L for nonessential AAs). After administration of AAs, the losses increased to 28 +/- 4 g. However, this procedure produced a positive net balance of AAs (10.6 +/- 5.6 g for total AAs), preventing a reduction in plasma concentrations. After 3 mo of AA administration, there was a significant increase in protein catabolic rate and serum albumin and transferrin. This improvement occurred without any change in the dialysis dose, ruling out the possibility that an increase in dialysis efficiency played a role. CONCLUSIONS: Intradialysis adequately provides AA supplements, prevents reductions in plasma AA concentrations, and favorably affects the nutritional status of patients receiving hemodialysis. PMID- 10702172 TI - Antioxidant intakes and smoking status: data from the continuing survey of food intakes by individuals 1994-1996. AB - BACKGROUND: Cigarette smoking is a major risk factor for several chronic oxidative diseases that can be ameliorated by antioxidants. OBJECTIVES: This study identified the typical dietary intakes and the major food group contributors of the antioxidants beta-carotene, vitamin C, and vitamin E by smoking status. DESIGN: The 1994-1996 Continuing Survey of Food Intakes by Individuals (CSFII) provided the current sample (n = 6749), who were categorized as non- (n = 3231), former (n = 1684), and current (n = 1834) smokers. In the CSFII, individuals' food intakes were estimated with two 24-h dietary recalls. Data were analyzed by using a chi-square test with a simultaneous Fisher's z test, analysis of variance with Scheffe's test, multivariate analysis of covariance, and analysis of covariance with Bonferroni adjustment for multiple comparisons. RESULTS: The sample consisted of 3707 men and 3042 women. Current smokers tended to be younger with less education and lower incomes than nonsmokers and former smokers. The average body mass index (in kg/m(2)) of current smokers was 25.8, the lowest of the 3 groups. Current smokers had the lowest dietary antioxidant intake. Fatty foods such as luncheon meats, condiments and salad dressings, and ground beef contributed more to the antioxidant intakes of current smokers than to those of the other 2 groups, whereas fruit and vegetables contributed less. Current smokers consumed the fewest numbers of servings of all nutrient-bearing groups in the food guide pyramid, except the meat group. CONCLUSION: Future interventions should target the clustering of cigarette smoking and other unhealthy lifestyle habits, eg, an imprudent diet. PMID- 10702173 TI - Changes in dietary zinc and copper affect zinc-status indicators of postmenopausal women, notably, extracellular superoxide dismutase and amyloid precursor proteins. AB - BACKGROUND: Zinc is an essential trace element for human health and well-being; however, methods currently available for the assessment of zinc status in humans are unsatisfactory. OBJECTIVE: The objective was to critically evaluate the use of various indicators of zinc status in humans in a controlled metabolic ward study. DESIGN: Indicators of zinc status were measured in 25 healthy postmenopausal women aged 64.9 +/- 6.7 y. After a 10-d equilibration period, volunteers consumed a diet with either a low (1 mg/d; n = 12) or a high (3 mg/d; n = 13) copper content based on a total energy content of 8.4 MJ. They received the same amount of copper throughout the study. Both groups were fed the basal diet (3 mg Zn/d) with no zinc supplement for one 90-d period, and the diet supplemented with 50 mg Zn/d for another 90-d period. RESULTS: Zinc supplementation significantly increased (P < 0.0001) extracellular but not erythrocyte superoxide dismutase activity. This increase was more apparent when subjects were fed the low-copper diet. Zinc supplementation in combination with the low-copper diet significantly decreased (P < 0.01) amyloid precursor protein expression in platelets. Other indicators of zinc status that were significantly elevated after zinc supplementation were as follows: plasma zinc and free thyroxine concentrations and mononuclear 5'-nucleotidase activity. CONCLUSION: The measurement of serum extracellular superoxide dismutase activity may be useful as a marker for the functional assessment of zinc status in humans. PMID- 10702174 TI - Iron and zinc supplementation improves indicators of vitamin A status of Mexican preschoolers. AB - BACKGROUND: The coexistence of multiple micronutrient deficiencies is a widespread public health problem in many regions of the world. Interactions between zinc deficiency and vitamin A metabolism have been reported but no longitudinal studies have evaluated the effect of iron deficiency on vitamin A. OBJECTIVE: The objective of this study was to investigate the effect of supplementation with iron, zinc, or both on vitamin A and its metabolically related proteins retinol binding protein (RBP) and transthyretin. DESIGN: The study was a longitudinal, double-blind, placebo-controlled trial in which 219 rural Mexican children aged 18-36 mo were randomly assigned to receive 20 mg Zn/d, 20 mg Fe/d, 20 mg Zn/d plus 20 mg Fe/d, or placebo. RESULTS: Six months after supplementation, plasma retinol increased in all supplemented groups. Compared with placebo, zinc supplementation was associated with significantly higher plasma retinol and transthyretin but the increase in RBP was not significant. Iron supplementation significantly increased plasma retinol, RBP, and transthyretin. Supplementation with zinc plus iron significantly increased plasma retinol but not RBP or transthyretin. Children deficient in zinc, iron, or vitamin A (as indicated by nutrient plasma concentration) at the beginning of the study had a significantly greater increase in retinol than did children with adequate nutrient status. CONCLUSIONS: Supplementation with zinc, iron, or both improved indicators of vitamin A status. The results of this study agree with previous observations of a metabolic interaction between zinc and vitamin A and suggest an interaction between iron and vitamin A metabolism. PMID- 10702175 TI - Carotenoids and carotenoids plus vitamin E protect against ultraviolet light induced erythema in humans. AB - BACKGROUND: Carotenoids and tocopherols, known to be efficient antioxidants and capable of scavenging reactive oxygen species generated during photooxidative stress, may protect the skin from ultraviolet light-induced erythema. beta Carotene is widely used as an oral sun protectant but studies on its protective effects are scarce. OBJECTIVE: The objective of this study was to investigate the protective effects of oral supplementation with carotenoids and a combination of carotenoids and vitamin E against the development of erythema in humans. DESIGN: A carotenoid supplement (25 mg total carotenoids/d) and a combination of the carotenoid supplement and vitamin E [335 mg (500 IU) RRR-alpha-tocopherol/d] were given for 12 wk to healthy volunteers. Erythema was induced by illumination with a blue-light solar simulator. Serum beta-carotene and alpha-tocopherol concentrations and skin carotenoid levels were assessed by HPLC and reflection photometry. RESULTS: Serum beta-carotene and alpha-tocopherol concentrations increased with supplementation. Erythema on dorsal skin (back) was significantly diminished (P < 0.01) after week 8, and erythema suppression was greater with the combination of carotenoids and vitamin E than with carotenoids alone. CONCLUSION: The antioxidants used in this study provided protection against erythema in humans and may be useful for diminishing sensitivity to ultraviolet light. PMID- 10702176 TI - Evaluation of serum retinol, the modified-relative-dose-response ratio, and breast-milk vitamin A as indicators of response to postpartum maternal vitamin A supplementation. AB - BACKGROUND: Conflicting results have been reported regarding the relative performance of serum retinol, the modified-relative-dose-response (MRDR) ratio, and breast-milk vitamin A concentrations in detecting changes in maternal vitamin A status. OBJECTIVE: We used receiver operating characteristic analyses and standardized differences to compare the ability of these indicators to detect a response to postpartum vitamin A supplementation in lactating Bangladeshi women. DESIGN: At 2 wk postpartum, women were randomly assigned to receive either a single dose of vitamin A [200000 IU (60000 retinol equivalents); n = 74] or placebo (n = 73). Data from maternal serum and breast milk collected 3 mo postpartum and from infant serum collected 6 mo postpartum were used to examine the ability of serum retinol, the MRDR ratio, and breast-milk vitamin A to discriminate between individuals in the supplemented and unsupplemented groups. Breast milk was collected by expressing the entire contents of one breast that had not been used to feed an infant for > or =2 h (full samples) or without controlling the time since the last breast-feeding episode (casual samples). RESULTS: Casual breast-milk samples performed better than full breast-milk samples in detecting a response to maternal supplementation. The MRDR ratio performed better than serum retinol in both the women and their infants. Overall, the most responsive indicator was the measurement of breast-milk vitamin A per gram of fat in casual breast-milk samples. CONCLUSIONS: Breast-milk vitamin A and the MRDR ratio are responsive indicators of vitamin A status, especially in women with mild vitamin A deficiency. PMID- 10702177 TI - Altered lipid profile, lipoprotein composition, and oxidant and antioxidant status in pediatric Crohn disease. AB - BACKGROUND: Growing evidence supports a role for peroxidation in the pathogenesis of Crohn disease (CD). The activation of inflammatory cells, the release of their mediators, and the excessive production of free radicals may affect circulating lipids. OBJECTIVE: We examined the lipid profile, lipoprotein composition, and oxidant-antioxidant status of children with CD. DESIGN: We studied 22 pediatric CD patients and 10 healthy control subjects. RESULTS: The proportion of saturated and monounsaturated fatty acids in plasma of CD patients was higher but that of polyunsaturated fatty acids was lower than in control subjects. This resulted in higher ratios in CD patients of palmitoleic acid to linoleic acid (P < 0. 05) and of eicosatrienoic acid to arachidonic acid (P < 0.04), 2 established indexes of essential fatty acid deficiency. Hypocholesterolemia was noted in CD patients as a result of lower LDL-cholesterol concentrations than in control subjects (P < 0.02). Plasma apolipoproteins B (P < 0.02) and A-I (P < 0.02) were also lower in CD patients, whereas plasma triacylglycerols were higher (P < 0.005). Lipoprotein composition was altered in CD patients, with relative triacylglycerol depletion and protein enrichment in VLDL. In contrast, intermediate-density lipoprotein of CD patients was characterized by an increased percentage of triacylglycerol and protein (P < 0.005) and a reduced proportion of phospholipids (P < 0. 01). Additional abnormalities were observed in the chemical distribution of HDL(2) and HDL(3) moieties. Lipid peroxidation was documented by higher plasma malondialdehyde concentrations in CD patients (P < 0.05), accompanied by lower retinol concentrations (P < 0.02). CONCLUSION: Disturbances in the lipid profile, in lipoprotein concentrations and composition, and in oxidant-antioxidant status occur in CD patients. PMID- 10702178 TI - Short-term growth and substrate use in very-low-birth-weight infants fed formulas with different energy contents. AB - BACKGROUND: Currently available preterm formulas with energy contents of 3350 kJ (800 kcal)/L promote weight and length gain at rates at or above intrauterine growth rates but disproportionately increase total body fat. OBJECTIVE: The objective of this study was to determine whether fat accretion in formula-fed, very-low-birth-weight (VLBW) infants could be decreased and net protein gain maintained by reducing energy intakes from 502 kJ (80 kcal)*kg(-)(1)*d(-)(1) [normal-energy (NE) formula] to 419 kJ (100 kcal)*kg(-)(1)*d(-)(1) [low-energy (LE) formula] while providing similar protein intakes (3.3 g*kg(-)(1)*d(-)(1)). DESIGN: The study was a randomized, controlled trial enrolling 20 appropriate-for gestational-age (AGA) and 16 small-for-gestational-age (SGA) VLBW infants (mean birth weight: 1.1 kg; mean gestational age: 31 wk); energy expenditure and nutrient balance were measured at 4 wk of age and anthropometric measurements were made when infants weighed 2 kg. RESULTS: The percentage of fat in newly formed tissue was significantly lower in AGA infants fed the LE formula (n = 9) than in those fed the NE formula (n = 10) (9% compared with 23%; analysis of variance, P = 0.001). Energy expenditure was higher in AGA infants fed the NE formula than in those fed the LE formula. Skinfold thickness was markedly lower in AGA infants fed the LE formula than in those fed the NE formula, resulting in a lower estimated percentage body fat (8.0 +/- 1.9% and 10.8 +/- 3.5%, respectively; P < 0.05). Three of 6 SGA infants fed the LE formula were excluded during the study because of poor weight gain. CONCLUSIONS: Body composition can easily be altered by changing the energy intakes of formula-fed VLBW infants. Energy intakes in these infants should be >419 kJ (100 kcal)*kg(-)(1)*d(-)(1). PMID- 10702179 TI - Randomized diet in the neonatal period and growth performance until 7.5-8 y of age in preterm children. AB - BACKGROUND: Preterm children are at high risk of poor growth performance. In 2 randomized trials, preterm infants fed preterm formula grew better in the neonatal period than those fed banked donor breast milk or standard term formula. OBJECTIVE: Our objective was to test the hypothesis that for preterm infants, the neonatal period is a critical one for programming growth performance and that early diet influences long-term growth. DESIGN: A total of 926 preterm infants were recruited into 2 parallel, randomized trials of neonatal diet. In trial 1, infants were fed either banked donor breast milk or preterm formula whereas in trial 2, infants were fed either standard term formula or preterm formula. Within each trial, the allocated milk was the sole diet for some infants (study A), whereas for others it was a supplement to maternal breast milk, given when not enough expressed breast milk was available (study B). We followed up 781 of 833 survivors (94%) to age 7.5-8 y. Trained assessors obtained anthropometric measurements according to a standard protocol. RESULTS: Despite significantly better neonatal growth performance in infants fed preterm formula (compared with either banked donor breast milk or standard formula), early diet had no influence on weight, height, head circumference, or skinfold thicknesses at 9 or 18 mo postterm or at age 7.5-8 y. CONCLUSIONS: These findings suggest that the preterm period is not a critical window for nutritional programming of growth, which contrasts with evidence from these trials showing that early diet influences later neurodevelopment. PMID- 10702180 TI - Paternal body fat is a longitudinal predictor of changes in body fat in premenarcheal girls. AB - BACKGROUND: Longitudinal studies in infants and children suggest that low total energy expenditure (EE) (TEE) and parental body composition are important predisposing factors to obesity. OBJECTIVE: The aim of this study was to examine potential predictors of changes in total or percentage body fat over 2.7 y in premenarcheal girls. DESIGN: We studied 47 normal-weight prepubertal girls aged 4.8-8.9 y in 3 visits. The girls' age, total and percentage body fat at baseline, sleep EE (SEE) and activity-related EE (AEE) adjusted for fat-free mass (FFM) and total body fat, mothers' and fathers' total and percentage body fat and FFM at baseline, and time to follow-up visits were measured; 24-h EE and SEE were measured by whole-room indirect calorimetry. AEE was calculated as TEE minus (SEE + 0.1 TEE), with the assumption that the thermic effect of food was 10% of TEE. The girls' body composition was measured at each visit and that of the parents was measured at the time of the girls' enrollment by using dual-energy X-ray absorptiometry. RESULTS: From baseline to the first (mean: 1.6 y) and the second (mean: 2.7 y) follow-up visits, the girls' mean (+/-SD) change in total fat adjusted for FFM was 1.2 +/- 2.7 and 3.3 +/- 4.0 kg, respectively, and the mean change in percentage body fat was -2.0 +/- 5.0% and -0. 8 +/- 5.9%, respectively. Fathers' total and percentage body fat were the main predictors of changes in the girls' total and percentage body fat. For the first follow-up visit, SEE, girls' age at baseline, and AEE were significant predictors of percentage body fat. CONCLUSION: Fathers' total and percentage body fat were predictors of changes in body fat of premenarcheal girls during a 2. 7-y period. PMID- 10702181 TI - Nutritional and metabolic effects of the endotoxin bacterial lipopolysaccharide in orally and parenterally fed rats. AB - BACKGROUND: Animals treated with tumor necrosis factor alpha (TNF-alpha) developed severe metabolic abnormalities despite receiving sufficient protein and energy by total parenteral nutrition (TPN). OBJECTIVE: We sought to investigate the nutritional and metabolic effects of bacterial lipopolysaccharide (LPS) in rats. DESIGN: Rats were randomly allocated to 5 groups: oral nutrition (ON control; n = 7), TPN control (n = 7), ON+LPS (n = 6), TPN+LPS (n = 9), and pair fed (PF) in relation to ON+LPS (n = 6). RESULTS: Body weight decreased significantly as diet consumption decreased in the ON+LPS and PF groups compared with the ON control group. Relative carcass weights were significantly lower in the TPN+LPS and ON+LPS groups than in their respective control groups. Diaphragm and extensor digitorum longus weights were significantly lower in the ON+LPS and PF rats, but not in the TPN+LPS rats, compared with their respective controls. Biochemical abnormalities and plasma corticosterone concentrations were greater in the TPN+LPS group than in the other groups. CONCLUSIONS: These data suggest that provision of sufficient protein and energy by TPN does not prevent general carcass wasting induced by LPS but may protect individual muscles. However, compared with an oral ad libitum diet, TPN providing sufficient protein and energy worsens the biochemical abnormalities induced by LPS. More rapid clearance of TNF-alpha and low corticosterone concentrations in weight-losing animals may help reduce the severity of the metabolic effects of LPS. PMID- 10702182 TI - Benefits and risks of antiobesity agents. PMID- 10702183 TI - Efficacy of antiobesity therapies. PMID- 10702184 TI - Blockade of pancreatic lipase. PMID- 10702185 TI - Orlistat and weight loss. PMID- 10702189 TI - Protein, fat, and ischemic heart disease. PMID- 10702190 TI - Animal protein and ischemic heart disease. PMID- 10702192 TI - Geriatric cachexia: a role for magnesium deficiency as well as for cytokines? PMID- 10702194 TI - Adolphe Quetelet. PMID- 10702195 TI - Digestive disease centers: it's not about the money. PMID- 10702196 TI - Tegaserod accelerates orocecal transit in patients with constipation-predominant irritable bowel syndrome. AB - BACKGROUND & AIMS: This study evaluated the effects of a partial 5 hydroxytryptamine (5-HT)(4) agonist, tegaserod, on gastric small bowel and colonic transit in constipation-predominant irritable bowel syndrome (IBS). METHODS: After a 1 week run-in period, 24 patients with constipation-predominant IBS were randomized to 1 week of tegaserod, 2 mg twice daily, or placebo treatment. Scintigraphic gastric emptying, small bowel transit, and colonic transit were determined before administration of study drug and after 1 week on the medication. Colonic transit was also measured using radiopaque markers and a single radiograph on day 5. RESULTS: Gastric emptying was unaltered by tegaserod. Proximal colonic filling at 6 hours, a measure of orocecal transit, was accelerated by tegaserod (70.4% +/- 1.3% [mean +/- SEM] vs. placebo, 46.4 +/- 1.9; P = 0.015). Proximal colonic emptying half-time and geometric center at 48 hours were also accelerated by tegaserod compared with baseline, but not compared with placebo. Mean colonic transit time was similar in both groups at baseline and after drug administration (tegaserod, 59.5 +/- 2.1 hours; placebo, 62.1 +/- 2.1 hours). CONCLUSIONS: Tegaserod accelerates orocecal transit, tends to accelerate colonic transit, and deserves further study in patients with constipation-predominant IBS. PMID- 10702197 TI - Topographic analysis of esophageal double-peaked waves. AB - BACKGROUND & AIMS: Esophageal double-peaked waves occur with increased frequency in patients with functional esophageal symptoms. This study was undertaken to further understand the mechanisms responsible for their production. METHODS: Topographic methods that consider temporal and spatial relationships of pressure data were used to examine 74 double-peaked waves detected in 18 subjects referred for manometric evaluation of unexplained symptoms. The studies were performed with a computerized data acquisition and analysis system designed for topographic plotting. RESULTS: The second peak appeared to represent muscle contraction that merged with an unusually strong pressure site in the third topographic segment and covered 6.3 +/- 1.6 cm (33.5% +/- 8.5% esophageal length) proximal to this site. In 50 swallows (67.6%), the peak itself progressed in a retrograde direction at 13.2 +/- 10.8 cm/s, suggesting cephalad extension of a strong distal motor event. Analysis of wave onsets and movement of the peristaltic trailing edge detected retrograde propagation in up to 33.8% of waves, antegrade propagation in 2.7%, and simultaneous contraction in the remainder. CONCLUSIONS: In symptomatic patients, the second peak in a double-peaked wave is typically a short, simultaneous, or retrograde pressure event in the region of and merging with the third topographic segment in the distal esophageal body. Topographic methods help explain the common association of these waveforms with other features of exaggerated contraction in the distal esophagus and suggest their relationship to inadequate inhibitory nerve function. PMID- 10702198 TI - Heme oxygenase activity in the internal anal sphincter: effects of nonadrenergic, noncholinergic nerve stimulation. AB - BACKGROUND & AIMS: To date, the exact role of carbon monoxide (CO) in the nonadrenergic, noncholinergic (NANC) relaxation is not known. This is partly related to the lack of an appropriate method to measure heme oxygenase (HO) activity in the gastrointestinal tissues. METHODS: HO activity of the opossum internal anal sphincter (IAS) smooth muscle was determined using a newly developed assay system that used radiolabeled hemin as a substrate. Enzyme activity of the IAS tissues was measured in the basal state, after electric field stimulation (EFS), ganglionic stimulant dimethyl diphenyl piperazinium iodide (DMPP), and neuropeptide vasoactive intestinal polypeptide (VIP). The presence and localization of HO was examined by Western blot analysis and immunocytochemistry. RESULTS: NANC nerve stimulation of the IAS smooth muscle by EFS (0.25-5 Hz), DMPP, and VIP caused a significant increase in the HO activity of the IAS. The increase in HO activity by EFS was inhibited by the HO inhibitor Tin protoporphyrin (1 x 10(-4) mol/L). Both HO-1 and HO-2 were present in the IAS tissue extracts, and both enzymes were localized in the neurons of the myenteric plexus. The method for HO activity determination used in the present study was found to be reliable and reproducible. CONCLUSIONS: The data suggest that the HO pathway may have a role in neurally mediated relaxation of the IAS. The exact site of involvement and the source of HO activity, however, remains to be determined. PMID- 10702199 TI - Cyclooxygenase 2 expression in Barrett's esophagus and adenocarcinoma: Ex vivo induction by bile salts and acid exposure. AB - BACKGROUND & AIMS: Barrett's esophagus (BE) results from chronic, severe gastroesophageal reflux and predisposes to esophageal adenocarcinoma. Cyclooxygenase (COX)-2 is involved in chronic inflammation and epithelial cell growth. We investigated COX-2 expression in BE and esophageal adenocarcinoma to explore a potential relation between COX-2 expression and metaplasia or carcinogenesis. METHODS: Endoscopic mucosal biopsy specimens of Barrett's intestinal metaplasia (n = 30), Barrett's dysplasia (n = 11), and esophageal adenocarcinoma (n = 5) were compared with normal esophagus (n = 46) and duodenum (n = 46) and analyzed by Western blotting and immunohistochemistry. RESULTS: Immunoblots revealed constitutive expression of COX-2 in normal esophagus and duodenum. COX-2 protein expression was significantly higher in patients with Barrett's metaplasia, dysplasia, and adenocarcinoma compared with normal squamous esophageal or columnar duodenal epithelia and was heterogenous in different regions of the BE surface. Immunohistochemistry revealed prominent staining in the glands of BE, dysplasia, and adenocarcinoma and faint staining in the basal layers of squamous esophagus and the surface of the duodenum. In response to pulses of acid or bile salts in an ex vivo organ culture system, COX-2 expression increased significantly in BE tissues, and this effect was attenuated by the selective COX-2 inhibitor NS-398. CONCLUSIONS: The results show COX-2 expression in normal esophagus, which increases significantly in BE and esophageal adenocarcinoma. COX-2 is regulated ex vivo by exposure to acid or bile salts. PMID- 10702200 TI - Integrin alpha6beta4 as a suppressor and a predictive marker for peritoneal dissemination in human gastric cancer. AB - BACKGROUND & AIMS: Because alterations of integrin expression in cancers contribute to cancer cell biology, we analyzed the association between the potential for peritoneal dissemination and integrin expression. METHODS: The dissemination potential of 10 human gastric cancer cell lines in mice with severe combined immunodeficiency (SCID) was compared with the expression of various integrins. The relationship between integrin expression and peritoneal dissemination was also investigated in surgically resected gastric cancer cases. RESULTS: The level of integrin beta4 subunit expression was inversely correlated with dissemination potential. Introduction of a full-length complementary DNA (cDNA) for beta4 subunit into cancer cells showing negligible beta4 subunit expression markedly suppressed peritoneal dissemination and inhibition of endogenous integrin alpha6beta4 by introduction of a cytoplasmic domain-deleted beta4 subunit cDNA into cells showing high expression of beta4 subunit promoted peritoneal dissemination. Apoptosis, which was histologically evident in peritoneal nodules of SCID mice, was induced in the cells with high beta4 subunit expression by attachment to laminin and stimulation with growth factors in vitro. An immunohistochemical study of specimens from 120 cases of primary gastric cancer showed that patients with beta4 subunit-positive tumors exhibited peritoneal dissemination only infrequently (P < 0.0001) and had a better outcome (P < 0.01). CONCLUSIONS: These results indicate that integrin alpha6beta4 is both a suppressor and a predictive marker for peritoneal dissemination in gastric cancer. PMID- 10702201 TI - Overexpression of protein kinase C-beta1 isoenzyme suppresses indomethacin induced apoptosis in gastric epithelial cells. AB - BACKGROUND & AIMS: We have previously reported that nonsteroidal anti inflammatory drugs (NSAIDs) could induce apoptosis of gastric epithelial cells both in vivo and in vitro. This study investigated the role of protein kinase C (PKC) isoforms in the regulation of NSAID-induced apoptosis. METHODS: Protein levels of 12 PKC isoforms in AGS cells, in the presence or absence of indomethacin, were determined by Western blot. The effect of PKC-beta1 overexpression by transfection with its complementary DNA (cDNA) on indomethacin induced apoptosis and apoptosis-related genes, including p53, p21(waf1/cip1), and c-myc, was further investigated. RESULTS: Treatment with indomethacin decreased the abundance of PKC-beta1 and increased that of PKC-beta2, eta, and epsilon, but did not alter the expression of PKC alpha, gamma, zeta, delta, iota, and micro. Overexpression of PKC-beta1 attenuated the apoptotic response of AGS cells to indomethacin, associated with overexpression of p21(waf1/cip1) in both messenger RNA and protein levels. Inhibition of PKC-beta1-mediated overexpression of p21(waf1/cip1) by its antisense cDNA partially reduced the antiapoptotic effect of PKC-beta1. CONCLUSIONS: Indomethacin-induced apoptosis in gastric cancer cells is partly mediated by differential regulation of PKC isoform expression. Enhanced expression of exogenous PKC-beta1 protects against indomethacin-induced apoptosis through up-regulation of p21(waf1/cip1). PMID- 10702202 TI - IL-1beta-induced apoptosis in rat gastric enterochromaffin-like cells is mediated by iNOS, NF-kappaB, and Bax protein. AB - BACKGROUND & AIMS: Enterochromaffin-like (ECL) cells are histamine-containing endocrine cells in the gastric mucosa. Previous studies have shown that the proinflammatory cytokine interleukin (IL)-1beta present during chronic gastritis inhibits histamine synthesis in ECL cells and leads to sustained functional impairment. This study investigated the effects of IL-1beta on ECL cell apoptosis and the related signal-transduction mechanisms. METHODS: ECL cells were isolated by pronase digestion and a combination of elutriation, gradient centrifugation, and 48-hour culture (purity >/=90%). Apoptosis was measured by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling reaction and cell death detection enzyme-linked immunosorbent assay. RESULTS: IL 1beta (100 pg/mL) increased the rate of programmed cell death 2-3 fold in ECL cells after 24 hours of incubation (total of 12%-14%). This effect was completely inhibited by the NF-kappaB inhibitor, proteasome inhibitor I, and the nitric oxide synthase inhibitor (iNOS) N(G)-monomethyl-L-arginine (10(-4) mol/L), but not by the caspase 3 inhibitor, Asp-Glu-Val-Asp-CHO. Western blot analysis, reverse-transcription polymerase chain reaction (PCR), and in situ PCR showed that IL-1beta induced gene expression (after 2-4 hours) and protein synthesis (6 18 hours) of the iNOS isoform in ECL cells. Bax protein expression was increased in response to IL-1beta. In contrast, bcl-2 gene expression was increased in response to basic fibroblast growth factor, which has been shown to counteract IL 1beta- induced apoptosis. CONCLUSIONS: These data suggest that IL-1beta induces programmed cell death in isolated rat ECL cells via activation of NF-kappaB, iNOS, and the Bax protein. PMID- 10702203 TI - Rapid activation of NF-kappaB and AP-1 and target gene expression in postischemic rat intestine. AB - BACKGROUND & AIMS: The molecular mechanisms underlying intestinal mucosal damage repair processes induced by ischemia-reperfusion (IR) remain unknown. We determined nuclear factor-kappaB (NF-kappaB) and activator protein 1 (AP-1) activities and the expression of potential target genes relevant to damage-repair events. METHODS: Rat jejunal segment was subjected to ischemia for 30 minutes followed by reperfusion for defined times. NF-kappaB and AP-1 activities; mucosal p105, p50, and inhibitor kappaB-alpha (IkappaB-alpha) levels; and c-fos, neurotensin, and ferritin H expression were determined by electrophoretic mobility shift assay and Western and Northern analyses, respectively. RESULTS: NF kappaB and AP-1 activities were significantly elevated from 1 to 12 hours after reperfusion. The activated NF-kappaB in the nuclear extract consisted of solely p50 homodimers. Activation of p50 was associated with a decrease of p105, generation of p50, and increased phosphorylation and degradation of IkappaB alpha. The activated AP-1 contained c-fos but not c-jun, fosB, and Fra-1. Reperfusion induced a transient elevation of c-fos, prolonged increase of neurotensin, and early reduction followed by recovery of ferritin H messenger RNA. CONCLUSIONS: The intestine shows organ-specific responses to IR, characterized by prolonged NF-kappaB and AP-1 activation involving NF-kappaB p50 dimers and excluding AP-1 c-jun protein. Degradation of the IkappaB-gamma component of p105 and partial reduction IkappaB-alpha selectively activate p50/p50 dimers. Temporal patterns of target gene expression reflect functional relevance to mucosal damage-repair processes after IR. PMID- 10702204 TI - Polarized transport of hydrophilic compounds across rat colonic mucosa from serosa to mucosa is temperature dependent. AB - BACKGROUND & AIMS: In both clinical and experimental studies, intestinal epithelial barrier function is routinely assessed by measuring mucosal permeability to various hydrophilic compounds. We performed experiments to determine whether permeation of several hydrophilic compounds across rat colonic mucosa is polarized. METHODS: Sheets of colonic mucosa, stripped of the underlying seromuscular coats, were mounted in Ussing chambers. RESULTS: The rates of permeation across colonic mucosa by numerous hydrophilic compounds (fluorescein isothiocyanate [FITC]-dextrans with molecular weights of 4000 [FD4] and 70,000 [FD70] daltons, fluorescein disulfonic acid [FS], lucifer yellow [LY], lactulose, and mannitol) were several times greater in the serosal-to-mucosal (S- >M) direction than in the opposite direction. Increased S-->M permeation by FD4, lactulose, and mannitol was evident at 37 degrees C, but not at 4 degrees C. Efflux of FD4 and FS in the S-->M direction was dose-dependently inhibited by verapamil, an inhibitor of the P-glycoprotein efflux system. Indomethacin, an anion transporter inhibitor, showed no effect on the S-->M permeation of FD4, FD70, FS, or LY. Adding an excess of unlabeled dextran (mol wt, 10,000 daltons) dose-dependently decreased the S-->M efflux of FD4, but not FS or LY. CONCLUSIONS: The transport across rat colonic mucosa of a number of hydrophilic substances, including some compounds that are commonly used to measure intestinal permeability in clinical practice, is greater in the S-->M than in the M-->S direction. S-->M transport of these hydrophilic solutes is temperature dependent, suggesting that the process is an active one. S-->M transport of FD4 may occur via a process that manifests some degree of substrate specificity for polysaccharides. PMID- 10702205 TI - The role of medial hypothalamic serotonin in the suppression of feeding in a rat model of colitis. AB - BACKGROUND & AIMS: Experimental colitis is associated with anorexia that is attenuated by treatment with an interleukin (IL)-1 receptor antagonist. Serotonin (5-hydroxytryptamine [5-HT]) is a potent inhibitor of feeding, and its release from the hypothalamus is stimulated by IL-1. We have tested the hypotheses that anorexia associated with experimental colitis results from increased activity of hypothalamic 5-HT neurons and that the increase in activity occurs secondary to an increase in availability of tryptophan, the precursor of 5-HT. METHODS: In vivo 5-HT release and regional hypothalamic 5-HT and tryptophan concentrations were measured in rats with 2,4,6,-trinitrobenzene sulfonic acid (TNBS)-induced colitis, healthy controls, and animals pair-fed to match the food intake of the colitic group. Food intake in the colitic group was assessed after depletion of brain 5-HT by p-chlorophenylalanine (PCPA). RESULTS: In the colitic group, release of 5-HT from the hypothalamic paraventricular nucleus (PVN) was 3-fold (P = 0.01) and 14-fold (P < 0.001) higher than in control and pair-fed groups, respectively. Concentrations of tryptophan were similar in each group in all hypothalamic regions. Food intake was significantly increased in the colitic group after PCPA treatment but was not restored to control values. CONCLUSIONS: In animals with TNBS-induced colitis, 5-HT release from the PVN is increased. The increase in food intake after depletion of brain 5-HT suggests that hypothalamic 5-HT contributes to anorexia but is not the only mediator. Increased 5-HT release in the colitic group was not driven by increased precursor availability. PMID- 10702206 TI - Hepatitis B genotypes correlate with clinical outcomes in patients with chronic hepatitis B. AB - BACKGROUND & AIMS: Six genotypes (A-F) of hepatitis B virus (HBV) have been identified; however, the genotype-related differences in the pathogenicity of HBV remain unknown. Therefore, we investigated the prevalence of HBV genotypes in Taiwan and the association between distinct genotypes and severity of liver disease in a cross-sectional study. METHODS: Using a molecular method, HBV genotypes were determined in 100 asymptomatic carriers and in 170 patients with histologically verified chronic liver disease and hepatocellular carcinoma (HCC). RESULTS: All genotypes except genotype E were identified in Taiwan, and genotypes B and C were predominant. Genotype C was prevalent in patients with cirrhosis and in those with HCC who were older than 50 years compared with age-matched asymptomatic carriers (60% vs. 23%, P < 0.001, and 41% vs. 15%, P = 0.005, respectively). Genotype B was significantly more common in patients with HCC aged less than 50 years compared with age-matched asymptomatic carriers (80% vs. 52%, P = 0.03). This predominance was more marked in younger patients with HCC (90% in those aged 4 N DNA content. These cells exhibited a 20-40% reduction in growth rate, which was rescued by plasmid-borne over-expression of BIR1 but not by its human counterpart, survivin. Deletion analysis revealed that the N-terminal domain of Bir1, containing the conserved baculovirus IAP repeat, was able to partially complement the cell growth defect caused by BIR1 deletion. Moreover, the full length and truncated forms of Bir1 accelerated cell division in wild-type cells. Finally, BIR1 heterozygous mutants exhibited grossly altered cell morphology with misshapen or abnormally long buds connected to an unusually large mother cell. These findings identify a novel function of IAP proteins in the pleiotropic control of cell division, in addition to their role in the suppression of apoptosis. PMID- 10702225 TI - NMR studies of Bacillus subtilis tRNA(Trp) hyperexpressed in Escherichia coli. Assignment of imino proton signals and determination of thermal stability. AB - 15N-Labeled Bacillus subtilis tRNA(Trp) wild type and a series of mutants were hyperexpressed in Escherichia coli and purified for NMR studies with the use of two-dimensional nuclear Overhauser effect spectroscopy (NOESY) and heteronuclear single quantum correlation (HSQC) and three-dimensional NOESY-HSQC techniques. These made possible chemical shift assignments of imino protons and determination of the thermal stability of the tRNA(Trp) molecules. Almost all of the imino protons in the helical regions and the tertiary base pairs were assigned, except three imino protons of the AU base pairs whose peaks were not clearly observed. Several base triplets found in the crystal structure of tRNA were observed in the present study as well. These studies also revealed two components of tRNA(Trp), which could not be separated by high pressure liquid chromatography, corresponding to s(4)U and U at position 8 of the tRNA(Trp), as indicated by two different sets of peaks for the TpsiC and D arms. The modification at position 8 altered the local conformation of the core region of the tRNA. Thermal unfolding experiments showed that the unfolding process is cooperative in the presence of a high concentration of magnesium ions and that the component corresponding to the s(4)U8 is more stable than the U8 component, thus providing evidence that the thiolation of U8 stabilizes the tertiary structure of tRNA. PMID- 10702226 TI - Molecular cloning and expression of mouse GD1alpha/GT1aalpha/GQ1balpha synthase (ST6GalNAc VI) gene. AB - A novel member of the mouse CMP-NeuAc:beta-N-acetylgalactosaminide alpha2,6 sialyltransferase (ST6GalNAc) subfamily, designated ST6GalNAc VI, was identified by BLAST analysis of expressed sequence tags. The sequence of the cDNA clone of ST6GalNAc VI encoded a type II membrane protein with 43 amino acids composing the cytoplasmic domain, 21 amino acids composing the transmembrane region, and 269 amino acids composing the catalytic domain. The predicted amino acid sequence showed homology to the previously cloned ST6GalNAc III, IV, and V, with common amino acid sequences in sialyl motif L and S among these four enzymes. A fusion protein with protein A and extracts from L cells transfected with ST6GalNAc VI in an expression vector showed enzyme activity of alpha2,6-sialyltransferase for GM1b, GT1b, and GD1a but not toward glycoproteins. Thin layer chromatography immunostaining revealed that the products were GD1alpha, GQ1balpha, and GT1aalpha. Northern blotting revealed that this gene was expressed in a wide range of mouse tissues such as colon, liver, heart, spleen, and brain. It is concluded that this enzyme is a novel sialyltransferase involved in the synthesis of alpha-series gangliosides in the nervous tissues and many other tissues. PMID- 10702227 TI - Identification of an interleukin-3-regulated aldoketo reductase gene in myeloid cells which may function in autocrine regulation of myelopoiesis. AB - The EML hematopoietic progenitor cell line is a model system for studying molecular events regulating myeloid commitment and terminal differentiation. We used representational difference analysis to identify genes that are expressed differentially during myeloid differentiation of EML cells. One gene (named mAKRa) encoded a novel member of the aldoketo reductase (AKR) superfamily of cytosolic NAD(P)(H)-dependent oxidoreductases. mAKRa mRNA was detected in murine hematopoietic tissues including bone marrow, spleen, and thymus. In myeloid cell lines, mAKRa was expressed at highest levels in cells representative of promyelocytes. mAKRa mRNA levels increased rapidly in response to interleukin-3 over the first 24 h of EML cell differentiation when the cells undergo lineage commitment and extensive proliferation. mAKRa mRNA levels decreased later in the differentiation process particularly when the EML cells were cultured with granulocyte/macrophage colony-stimulating factor and retinoic acid to induce terminal granulocytic maturation. mAKRa mRNA levels decreased during retinoic acid-induced terminal granulocytic differentiation of the MPRO promyelocyte cell line. AKRs act as molecular switches by catalyzing the interconversion or inactivation of bioactive molecules including steroids and prostaglandins. We propose that mAKRa may catalyze the production or catabolism of autocrine factors that promote the proliferation and/or lineage commitment of early myeloid progenitors. PMID- 10702228 TI - T cells activated by zwitterionic molecules prevent abscesses induced by pathogenic bacteria. AB - Immunologic paradigms classify bacterial polysaccharides as T cell-independent antigens. However, these models fail to explain how zwitterionic polysaccharides (Zps) confer protection against intraabdominal abscess formation in a T cell dependent manner. Here, we demonstrate that Zps elicit a potent CD4+ T cell response in vitro that requires available major histocompatibility complex class II molecules on antigen-presenting cells. Specific chemical modifications to Zps show that: 1) the activity is specific for carbohydrate structure, and 2) the proliferative response depends upon free amino and carboxyl groups on the repeating units of these polysaccharides. Peptides synthesized to mimic the zwitterionic charge motif associated with Zps also exhibited these biologic properties. Lysine-aspartic acid (KD) peptides with more than 15 repeating units stimulated CD4+ T cells in vitro and conferred protection against abscesses induced by bacteria such as Bacteroides fragilis and Staphylococcus aureus. Evidence for the biologic importance of T cell activation by these zwitterionic polymers was provided when human CD4+ T cells stimulated with these molecules in vitro and adoptively transferred to rats in vivo conferred protection against intraabdominal abscesses induced by viable bacterial challenge. These studies demonstrate that bacterial polysaccharides with a distinct charge motif activate T cells and that this activity confers immunity to a distinct pathologic response to bacterial infection. PMID- 10702230 TI - Mechanistic basis for catalytic activation of mitogen-activated protein kinase phosphatase 3 by extracellular signal-regulated kinase. AB - The dual specificity mitogen-activated protein kinase phosphatase MKP3 has been shown to down-regulate mitogenic signaling through dephosphorylation of extracellular signal-regulated kinase (ERK). Camps et al. (Camps, M., Nichols, A., Gillieron, C., Antonsson, B., Muda, M., Chabert, C., Boschert, U., and Arkinstall, S. (1998) Science 280, 1262-1265) had demonstrated that ERK binding to the noncatalytic amino-terminal domain of MKP3 can dramatically activate the phosphatase catalytic domain. The physical basis for this activation has not been established. Here, we provide detailed biochemical evidence that ERK activates MKP3 through the stabilization of the active phosphatase conformation, inducing closure of the catalytic "general acid" loop. In the closed conformation, this loop structure can participate efficiently in general acid/base catalysis, substrate binding, and transition-state stabilization. The pH activity profiles of ERK-activated MKP3 clearly indicated the involvement of general acid catalysis, a hallmark of protein-tyrosine phosphatase catalysis. In contrast, unactivated MKP3 did not display this enzymatic group as critical for the low activity form of the enzyme. Using a combination of Bronsted analyses, pre-steady state and steady-state kinetics, we have isolated all catalytic steps in the reaction and have quantified the specific rate enhancement. Through protonation of the leaving group and transition-state stabilization, activated MKP3 catalyzes formation of the phosphoenzyme intermediate approximately 100-fold faster than unactivated enzyme. In addition, ERK-activated MKP3 catalyzes intermediate hydrolysis 5-6-fold more efficiently and binds ligands up to 19-fold more tightly. Consistent with ERK stabilizing the active conformation of MKP3, the chemical chaperone dimethyl sulfoxide was able to mimic this activation. A general protein-tyrosine phosphatase regulatory mechanism involving the flexible general acid loop is discussed. PMID- 10702229 TI - N-t-butyl hydroxylamine, a hydrolysis product of alpha-phenyl-N-t-butyl nitrone, is more potent in delaying senescence in human lung fibroblasts. AB - Alpha-phenyl-N-t-butyl nitrone (PBN), a spin trap, scavenges hydroxyl radicals, protects tissues from oxidative injury, and delays senescence of both normal human lung fibroblasts (IMR90) and senescence-accelerated mice. N-t-butyl hydroxylamine and benzaldehyde are the breakdown products of PBN. N-t-Butyl hydroxylamine delays senescence of IMR90 cells at concentrations as low as 10 microM compared with 200 microM PBN to produce a similar effect, suggesting that N-t-butyl hydroxylamine is the active form of PBN. N-Benzyl hydroxylamine and N methyl hydroxylamine compounds unrelated to PBN were also effective in delaying senescence, suggesting the active functional group is the N-hydroxylamine. All the N-hydroxylamines tested significantly decreased the endogenous production of oxidants, as measured by the oxidation of 2', 7'-dichlorodihydrofluorescin and the increase in the GSH/GSSG ratio. The acceleration of senescence induced by hydrogen peroxide is reversed by the N-hydroxylamines. DNA damage, as determined by the level of apurinic/apyrimidinic sites, also decreased significantly following treatment with N-hydroxylamines. The N-hydroxylamines appear to be effective through mitochondria; they delay age-dependent changes in mitochondria as measured by accumulation of rhodamine-123, they prevent reduction of cytochrome C(FeIII) by superoxide radical, and they reverse an age-dependent decay of mitochondrial aconitase, suggesting they react with the superoxide radical. PMID- 10702231 TI - Store-operated calcium entry in vascular endothelial cells is inhibited by cGMP via a protein kinase G-dependent mechanism. AB - Store-operated Ca(2+) entry in vascular endothelial cells not only serves to refill the intracellular Ca(2+) stores, but also acts to stimulate the synthesis of nitric oxide, a key vasodilatory factor. In this study, we examined the role of cGMP in regulating the store-operated Ca(2+) entry in aortic endothelial cells. Cyclopiazonic acid (CPA) and thapsigargin, two selective inhibitors of endoplasmic reticulum Ca(2+)-ATPase, were used to induce store-operated Ca(2+) entry. 8-Bromo-cGMP, an activator of protein kinase G, inhibited the CPA- or thapsigargin-induced Ca(2+) entry in a concentration-dependent manner. An inhibitor of protein kinase G, KT5823 (1 microM) or H-8 (10 microM), abolished the inhibitory action of 8-bromo-cGMP and resumed Ca(2+) entry. Addition of S nitroso-N-acetylpenicillamine (a nitric oxide donor) or dipyridamole (a cGMP phosphodiesterase inhibitor) during CPA treatment elevated cellular cGMP levels, stimulated protein kinase G activity, and at the same time reduced Ca(2+) influx due to CPA. Patch clamp study confirmed the existence of a CPA-activated Ca(2+) permeable channel sensitive to cGMP inhibition. These results suggest that cGMP via a protein kinase G-dependent mechanism may play a key role in the regulation of the store-operated Ca(2+) entry in vascular endothelial cells. PMID- 10702232 TI - Inactivation of interferon regulatory factor-1 tumor suppressor protein by HPV E7 oncoprotein. Implication for the E7-mediated immune evasion mechanism in cervical carcinogenesis. AB - In studying biological roles of interferon regulatory factor (IRF)-1 tumor suppressor in cervical carcinogenesis, we found that HPV E7 is functionally associated with IRF-1. Binding assays indicate a physical interaction between IRF 1 and HPV E7 in vivo and in vitro. The carboxyl-terminal transactivation domain of IRF-1 was required for the interaction. Transient co-expression of E7 significantly inhibits the IRF-1-mediated activation of IFN-beta promoter in NIH 3T3 cells. Co-transfection of E7 mutants reveals that the pRb-binding portion of E7 is necessary for the E7-mediated inactivation of IRF-1. It was next determined whether histone deacetylase (HDAC) is involved in the inactivation mechanism as recently suggested, where the carboxyl-terminal zinc finger domain of E7 associates with NURD complex containing HDAC. When trichostatin A, an inhibitor of HDAC, was treated, the repressing activity of E7 was released in a dose dependent manner. Furthermore, the mutation of zinc finger abrogates such activity without effect on the interaction with IRF-1. These results suggest that HPV E7 interferes with the transactivation function of IRF-1 by recruiting HDAC to the promoter. The immune-promoting role of IRF-1 evokes the idea that our novel finding might be important for the elucidation of the E7-mediated immune evading mechanism that is frequently found in cervical cancer. PMID- 10702233 TI - Transcriptional regulation of the human CYP1B1 gene. Evidence for involvement of an aryl hydrocarbon receptor response element in constitutive expression. AB - The cytochrome P450 1B1 gene (CYP1B1) is expressed constitutively and is inducible by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in the human breast adenocarcinoma cell line MCF-7 but not in the human hepatoma cell line HepG2. Genomic DNA isolated from both cell lines was digested with the methylation sensitive restriction enzyme isoschizomers MspI and HpaII, and subjected to Southern analysis with a probe for the CYP1B1 promoter/enhancer region. Although differences were observed in methylation patterns for the CYP1B1 gene from MCF-7 and HepG2 cells, treatment with the demethylating agent 5-azacytidine (10 microM for 6 days) did not activate CYP1B1 mRNA expression in HepG2 cells. Furthermore, treatment with the histone deacetylase inhibitor trichostatin A (100 nM for 24 h) did not activate CYP1B1 mRNA expression in HepG2 cells. Comparative analysis of the constitutive expression of luciferase/1B1 reporter constructs containing a series of deletions in the 5' enhancer region indicated that in MCF-7 cells the region from -987 to -732 (relative to the transcription start site) was necessary for maximal levels of activity. Mutation of the aryl hydrocarbon receptor response elements (dioxin response elements) in this region showed that the dioxin response elements located at -833 is essential for constitutive gene expression in MCF-7 cells. In HepG2 cells, reporter gene activity was at least equal or greater than the activity observed in MCF-7 cells, which is in marked contrast to the expression of the native CYP1B1 gene. Taken together these findings indicate that the observed cell-specific differences in CYP1B1 constitutive expression are not mediated by DNA promoter/enhancer methylation, but are likely due to either 1) inaccessibility of the 5'-enhancer region in HepG2 cells to transcriptional activators due to a higher order chromatin structure that does not involve histone acetylation, or 2) the action of a repressor protein at cis-elements located outside of the -2296 to +25 region examined with the CYP1B1 reporter constructs. Furthermore, at least one of the dioxin response elements in the enhancer region is required for constitutive expression of CYP1B1. PMID- 10702234 TI - pH and calcium regulate the water permeability of aquaporin 0. AB - Aquaporins increase the water permeability in many cell types across many species. We investigated the effects of external pH and Ca(2+) on water permeability of Xenopus oocytes injected with aquaporin cRNA by measuring the rate of swelling in hypotonic solutions. Lowering pH to 6.5 increased the water permeability of aquaporin (AQP0) 3.4 +/- 0.4-fold. Diethylpyrocarbonate pretreatment increased water permeability 4.2 +/- 0.5-fold and abolished pH sensitivity, suggesting that the pH regulation is mediated by an external histidine. Lowering Ca(2+) increased water permeability 4.1 +/- 0. 4-fold. The effects of Ca(2+) and pH each required the presence of histidine 40, indicating a critical role of this amino acid in facilitating the modulation of water permeability. Clamping intracellular Ca(2+) at high or low values abolished sensitivity to external Ca(2+), suggesting that Ca(2+) acts at an internal site. Three different calmodulin inhibitors each increased AQP0 water permeability, suggesting that Ca(2+) may act through calmodulin. None of the above altered the water permeability induced by AQP1 or AQP4. Because the greatest change in AQP0 water permeability is in the normal pH range found in the lens (7.2-6.5), this paper provides evidence for regulation of an aquaporin by pH under physiological conditions. PMID- 10702235 TI - The study of guanosine 5'-diphosphate 3'-diphosphate-mediated transcription regulation in vitro using a coupled transcription-translation system. AB - The effects of the "alarmone" guanosine 5'-diphosphate 3'-diphosphate (ppGpp) on regulation of the Salmonella typhimurium histindine operon and the Escherichia coli tRNA(leu) operon were analyzed in vitro using a DNA-dependent transcription translation system, S-30. The expression of the hisG promoter is positively regulated by ppGpp, whereas that of the leuV promoter (of tRNA(1eu)) is negatively regulated by ppGpp. In an attempt to understand the global regulatory mechanism of ppGpp control, interrelationship between ppGpp-dependent activation and repression of gene expression was examined using these promoters as models. It has been traditionally supposed that the ppGpp-dependent regulation, at least for the activation, is by a passive mode of control: the activation of gene expression by ppGpp is a consequence of the repression of stable RNA gene expression in the condition of RNA polymerase limiting. To test this model, the ppGpp-dependent regulations of both an activable promoter (hisGp) and a repressible promoter (leuVp) were determined in vitro simultaneously using a mixed template setup. The rationale for this exercise was to see whether the ppGpp-dependent activation and repression are inversely correlated in the in vitro condition in which RNA polymerase is limiting. No correlation was observed. It was concluded that the ppGpp-dependent activation is independent of the repression. Moreover, it was proposed that ppGpp-dependent activation and repression are mediated by titratable factors, each of which operate independently. PMID- 10702236 TI - Increased endosomal sorting of ligand to recycling enhances potency of an interleukin-2 analog. AB - An interleukin-2 (IL-2) variant containing adjacent point mutations (L18M/L19S, termed 2D1) displaying binding affinity to the heterotrimeric IL-2 receptor similar to that of wild-type IL-2 (WT) had been previously found to surprisingly exhibit increased bioactivity in a peripheral blood lymphocyte proliferation assay. In order to provide an explanatory mechanism for this unexpected potency enhancement, we hypothesize that altered endocytic trafficking of the 2D1 variant might be responsible by increasing the number of ligand-receptor complexes. We demonstrate here that the internalization kinetics of 2D1 via the high affinity IL-2 receptor are equivalent to those of WT but that a significantly increased fraction of internalized 2D1 is sorted to recycling instead of to lysosomal degradation. We further find a reduced pH sensitivity of binding to IL-2 receptor alpha relative to IL-2 receptor beta compared with WT, which could be responsible for the altered sorting behavior of 2D1 in the acidic endosomal compartment. Accordingly, the 2D1 variant displays a half-life 36 h longer than that of IL-2 in T-lymphocyte culture at concentrations equal to the K(D) of the IL-2 receptor. The extended half-life of intact 2D1 provides enhanced mitogenesis as compared with IL-2. In addition, 2D1 stimulates natural killer cells to a lesser degree than IL-2 at equal concentrations. We conclude that this IL-2 variant provides increased mitogenic stimulation that could not be easily predicted from its cell surface receptor binding affinity while minimizing undesired stimulation of natural killer cells. This concept of altering trafficking dynamics may offer a generalizable approach to generating improvements in the pharmacological efficacy of therapeutic cytokines. PMID- 10702237 TI - Resonance Raman characterization of biotin sulfoxide reductase. Comparing oxomolybdenum enzymes in the ME(2)SO reductase family. AB - Resonance Raman spectroscopy has been used to define active site structures for oxidized Mo(VI) and reduced Mo(IV) forms of recombinant Rhodobacter sphaeroides biotin sulfoxide reductase expressed in Escherichia coli. On the basis of (18)O/(16)O labeling studies involving water and the alternative substrate dimethyl sulfoxide and the close correspondence to the resonance Raman spectra previously reported for dimethyl sulfoxide reductase (Garton, S. D., Hilton, J., Oku, H., Crouse, B. R., Rajagopalan, K. V., and Johnson, M. K. (1997) J. Am. Chem. Soc. 119, 12906-12916), vibrational modes associated with a terminal oxo ligand and the two molybdopterin dithiolene ligands have been assigned. The results indicate that the enzyme cycles between mono-oxo-Mo(VI) and des-oxo Mo(IV) forms with both molybdopterin dithiolene ligands remaining coordinated in both redox states. Direct evidence for an oxygen atom transfer mechanism is provided by (18)O/(16)O labeling studies, which show that the terminal oxo group at the molybdenum center is exchangeable with water during redox cycling and originates from the substrate in substrate-oxidized samples. Biotin sulfoxide reductase is not reduced by biotin or the nonphysiological products, dimethyl sulfide and trimethylamine. However, product-induced changes in the Mo=O stretching frequency provide direct evidence for a product-associated mono-oxo Mo(VI) catalytic intermediate. The results indicate that biotin sulfoxide reductase is thermodynamically tuned to catalyze the reductase reaction, and a detailed catalytic mechanism is proposed. PMID- 10702238 TI - Thymidine diphosphate-6-deoxy-L-lyxo-4-hexulose reductase synthesizing dTDP-6 deoxy-L-talose from Actinobacillus actinomycetemcomitans. AB - The serotype c-specific polysaccharide antigen of Actinobacillus actinomycetemcomitans NCTC 9710 contains an unusual sugar, 6-deoxy-L-talose, which has been identified as a constituent of cell wall components in some bacteria. Two genes coding for thymidine diphosphate (dTDP)-6-deoxy-L-lyxo-4 hexulose reductases were identified in the gene cluster required for biosynthesis of serotype c-specific polysaccharide. Both dTDP-6-deoxy-L-lyxo-4-hexulose reductases were overproduced and purified from Escherichia coli transformed with the plasmids containing these genes. The sugar nucleotides converted by both reductases were purified by reversed-phase high performance liquid chromatography and identified by (1)H nuclear magnetic resonance and gas-liquid chromatography. The results indicated that one of two reductases produced dTDP-6-deoxy-L-talose and the other produced dTDP-L-rhamnose (dTDP-6-deoxy-L-mannose). The amino acid sequence of the dTDP-6-deoxy-L-lyxo-4-hexulose reductase forming dTDP-6-deoxy-L talose shared only weak homology with that forming dTDP-L-rhamnose, despite the fact that these two enzymes catalyze the reduction of the same substrate and the products are determined by the stereospecificity of the reductase activity. Neither the gene for dTDP-6-deoxy-L-talose biosynthesis nor its corresponding protein product has been found in other bacteria; this biosynthetic pathway is identified here for the first time. PMID- 10702239 TI - Three-dimensional migration of neurites is mediated by adhesion site density and affinity. AB - Three-dimensional neurite outgrowth rates within fibrin matrices that contained variable amounts of RGD peptides were shown to depend on adhesion site density and affinity. Bi-domain peptides with a factor XIIIa substrate in one domain and a RGD sequence in the other domain were covalently incorporated into fibrin gels during coagulation through the action of the transglutaminase factor XIIIa, and the RGD-dependent effect on neurite outgrowth was quantified, employing chick dorsal root ganglia cultured two- and three-dimensionally within the modified fibrin. Two separate bi-domain peptides were synthesized, one with a lower binding affinity linear RGD domain and another with a higher binding affinity cyclic RGD domain. Both peptides were cross-linked into fibrin gels at concentrations up to 8.2 mol of peptide/mol of fibrinogen, and their effect on neurite outgrowth was measured. Both two- and three-dimensional neurite outgrowth demonstrated a bi-phasic dependence on RGD concentration for both the linear and cyclic peptide, with intermediate adhesion site densities yielding maximal neurite extension and higher densities inhibiting outgrowth. The adhesion site density that yielded maximal outgrowth depended strongly on adhesion site affinity in both two and three dimensions, with lower densities of the higher affinity ligand being required (0.8-1.7 mol/mol for the linear peptide versus 0.2 mol/mol for the cyclic peptide yielding maximum neurite outgrowth rates in three dimensional cultures). PMID- 10702240 TI - Cathepsin G activates protease-activated receptor-4 in human platelets. AB - Of the four known protease-activated receptors (PARs), PAR1 and PAR4 are expressed by human platelets and mediate thrombin signaling. Whether these receptors are redundant, interact, or play at least partially distinct roles is unknown. It is possible that PAR1 and/or PAR4 might confer responsiveness to proteases other than thrombin. The neutrophil granule protease, cathepsin G, is known to cause platelet secretion and aggregation. We now report that this action of cathepsin G is mediated by PAR4. Cathepsin G triggered calcium mobilization in PAR4-transfected fibroblasts, PAR4-expressing Xenopus oocytes, and washed human platelets. An antibody raised against the PAR4 thrombin cleavage site blocked platelet activation by cathepsin G but not other agonists. Desensitization with a PAR4 activating peptide had a similar effect. By contrast, inhibition of PAR1 function had no effect on platelet responses to cathepsin G. When neutrophils were present, the neutrophil agonist fMet-Leu-Phe triggered calcium signaling in Fura-2-loaded platelets. Strikingly, this neutrophil-dependent platelet activation was blocked by the PAR4 antibody. These data show that PAR4 mediates platelet responses to cathepsin G and support the hypothesis that cathepsin G might mediate neutrophil-platelet interactions at sites of vascular injury or inflammation. PMID- 10702241 TI - Molecular cloning and expression of three isoforms of the 100-kDa a subunit of the mouse vacuolar proton-translocating ATPase. AB - We have identified cDNAs encoding three isoforms (a1, a2, and a3) of the 100-kDa a subunit of the mouse vacuolar proton-translocating ATPase (V-ATPase). The predicted protein sequences of the three isoforms are 838, 856, and 834 amino acids, respectively, and they display approximately 50% identity between isoforms. Northern blot analysis demonstrated that all three isoforms are expressed in most tissues examined. However, the a1 isoform is expressed most heavily in brain and heart, a2 in liver and kidney, and a3 in liver, lung, heart, brain, spleen, and kidney. We also identified multiple alternatively spliced variants for each isoform. Reverse transcriptase-mediated polymerase chain reaction revealed that one splicing variant of the a1 isoform (a1-I) was expressed only in brain, whereas two other variants (a1-II and a1-III) were expressed in tissues other than brain. These alternatively spliced forms differ in the presence or absence of 6-7 amino acid residues near the amino and carboxyl termini of the proteins encoded. The a3 isoform is also encoded by three alternatively spliced variants, two of which are predicted to encode a protein that is truncated near the border of the amino- and carboxyl-terminal domains of the a subunit and therefore lacks the integral transmembrane-spanning helices thought to participate in proton translocation. Expression of each isoform (with the exception of a1-I) was detectable at all developmental stages investigated, with a1-I absent only in day 7 embryos. The results obtained suggest that isoforms of the 100-kDa a subunit may contribute to tissue-specific functions of the V-ATPase. PMID- 10702242 TI - Pairing of the nucleotide binding domains of the transporter associated with antigen processing. AB - The transporter associated with antigen processing (TAP) comprises two structurally related subunits, TAP1 and TAP2, that form stable complexes in endoplasmic reticulum (ER) membranes. TAP complexes function in the translocation of peptides from the cytosol into the ER lumen for presentation by major histocompatibility complex class I molecules. Each TAP subunit contains an N terminal membrane-spanning region with multiple membrane-spanning segments, and a C-terminal, cytosolic nucleotide binding region. To study the nature of the interactions occurring on the cytosolic face of TAP1/TAP2 complexes, we investigated quaternary associations mediated by two C-terminal fragments of human TAP1 (T1c, residues 452-748 and T1ctr, residues 472-748) and two C-terminal fragments of human TAP2 (T2c, residues 399-686 and T2ctr, residues 433-686). Each of these constructs contains the core nucleotide binding region as well as a long or short N-terminal extension. We show stable complex formation between T1c and T2c but not between T1ctr and T2ctr. The mechanistic implications of these results are discussed. We also show that each of the constructs except T1ctr interacts with wild type TAP1 and TAP2, indicating possibilities for homodimerization of TAP1 and TAP2, or of oligomerization of TAP1/TAP2 heterodimers on membranes. PMID- 10702243 TI - Probing the native structure of stathmin and its interaction domains with tubulin. Combined use of limited proteolysis, size exclusion chromatography, and mass spectrometry. AB - Stathmin is a cytosoluble phosphoprotein proposed to be a regulatory relay integrating diverse intracellular signaling pathway. Its interaction with tubulin modulates microtubule dynamics by destabilization of assembled microtubules or inhibition of their polymerization from free tubulin. The aim of this study was to probe the native structure of stathmin and to delineate its minimal region able to interact with tubulin. Limited proteolysis of stathmin revealed four structured domains within the native protein, corresponding to amino acid sequences 22-81 (I), 95-113 (II), 113-128 (III), and 128-149 (IV), which allows us to propose stathmin folding hypotheses. Furthermore, stathmin proteolytic fragments were mixed to interact with tubulin, and those that retained affinity for tubulin were isolated by size exclusion chromatography and identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The results indicate that, to interact with tubulin, a stathmin fragment must span a minimal core region from residues 42 to 126, which interestingly corresponds to the predicted alpha-helical "interaction region" of stathmin. In addition, an interacting stathmin fragment must include a short N- or C-terminal extension. The functional significance of these interaction constrains is further validated by tubulin polymerization inhibition assays with fragments designed on the basis of the tubulin binding results. The present results will help to optimize further stathmin structural studies and to develop molecular tools to target its interaction with tubulin. PMID- 10702244 TI - Casein kinase II phosphorylates lens connexin 45.6 and is involved in its degradation. AB - Connexin (Cx) 45.6, an avian counterpart of rodent Cx50, is phosphorylated in vivo, but the sites and function of the phosphorylation have not been elucidated. Our peptide mapping experiments showed that the Ser(363) site in the carboxyl (COOH) terminus of Cx45.6 was phosphorylated and that this site is within casein kinase (CK) II consensus sequence, although showing some similarity to CKI sequence. The peptide containing Ser(363) could be phosphorylated in vitro by CKII, but not by CKI. Furthermore, CKII phosphorylated Cx45.6 in embryonic lens membrane and the fusion protein containing the COOH terminus of Cx45.6. Two dimensional peptide mapping experiments showed that one of the Cx45.6 peptides phosphorylated in vivo migrated to the same spot as one of those phosphorylated by CKII in vitro. Furthermore, CKII activity could be detected in lens lysates. To assess the function of this phosphorylation event, exogenous wild type and mutant Cx45.6 (Ser(363) --> Ala) were expressed in lens primary cultures by retroviral infection. The mutant Cx45.6 was shown to be more stable having a longer half-life compared with wild type Cx45.6. Together, the evidence suggests that CKII is likely a kinase responsible for the Ser(363) phosphorylation, leading to the destablization and degradation of Cx45.6. The connexin degradation induced by phosphorylation has a broad functional significance in the regulation of gap junctions in vivo. PMID- 10702245 TI - Physical properties of the transmembrane signal molecule, sn-1-stearoyl 2 arachidonoylglycerol. Acyl chain segregation and its biochemical implications. AB - sn-1,2-diacylglycerol (DAG), a key intermediate in lipid metabolism, activates protein kinase C and is a fusogen. Phosphoinositides, the main sources of DAG in cell signaling, contain mostly stearoyl and arachidonoyl in the sn-1 and -2 positions, respectively. The polymorphic behavior of sn-1-stearoyl-2 arachidonoylglycerol (SAG) was studied by differential scanning calorimetry, x ray powder diffraction, and solid state magic angle spinning (MAS) (13)C NMR. Three alpha phases were found in the dry state. X-ray diffraction indicated that the acyl chains packed in a hexagonal array in the alpha phase, and the two sub alpha phases packed with pseudo-hexagonal symmetry. In the narrow angle range strong diffractions of approximately 31 and approximately 62 A were present. High power proton-decoupled MAS (13)C NMR of isotropic SAG gave 16 distinct resonances of the 20 arachidonoyl carbons and 5 distinct resonances of the 18 stearoyl carbons. Upon cooling, all resonances of stearoyl weakened and vanished in the sub-alpha(2) phase, whereas arachidonoyl carbons from 8/9 to 20 gave distinct resonances in the frozen phases. Remarkably, the omega-carbon of the two acyl chains had different chemical shifts in alpha, sub-alpha(1), and sub-alpha(2) phases. Large differences in spin lattice relaxation of the stearoyl and arachidonoyl methene and methyl groups were demonstrated by contact time (cross polarization) MAS (13)C NMR experiments in the solid phases alpha, sub-alpha(1), and sub-alpha(2). This shows that stearoyl and arachidonoyl in SAG have different environments in the solid states (alpha, sub-alpha(1), and sub-alpha(2) phases) and may segregate during cooling. The NMR and long spacing x-ray diffraction results suggest that SAG does not pack in a conventional double layer with the two acyls in a hairpin fashion. Our findings thus provide a physicochemical basis for DAG hexagonal phase domain separation within membrane bilayers. PMID- 10702246 TI - Chemokine receptors CXCR-1/2 activate mitogen-activated protein kinase via the epidermal growth factor receptor in ovarian cancer cells. AB - Ovarian cancer typically disseminates widely in the abdomen, a characteristic that limits curative therapy. The mechanisms that promote ovarian cancer cell migration are incompletely understood. We studied model SK-OV-3 ovarian cancer cells and observed robust expression of the alpha chemokine receptors CXCR-1 and CXCR-2. Interleukin-8 (IL-8) treatment caused shape changes in the cells, with membrane ruffling and formation/retraction of thin actin-like projections, as detected by time-lapse microscopy. Stimulation of the CXCR-1/2 receptors by human interleukin 8 (IL-8) rapidly activated the p44/42 mitogen-activated protein (extracellular signal-regulated kinase (Erk1/2)) kinase pathway. Treatment of SK OV-3 cells with the inhibitors genestein and herbimycin A indicated that tyrosine kinases were involved in the IL-8 activation of Erk1 and Erk2. Of note, IL-8 induced transient phosphorylation of the epidermal growth factor (EGF) receptor and its association with the adaptor molecules Shc and Grb2. This transactivation of the EGF receptor was dependent on intracellular Ca(2+) mobilization. Furthermore AG1478, a specific inhibitor of the EGF receptor kinase, blocked Erk1 and Erk2 activation. c-Src kinase was not involved in the IL-8-mediated phosphorylation of the EGF receptor, but was critical for Shc phosphorylation and downstream Erk1/2 kinase activation. These results suggest important "cross-talk" between chemokine and growth factor pathways that may link signals of cell migration and proliferation in ovarian cancer. PMID- 10702247 TI - Cloning of an alkaline ceramidase from Saccharomyces cerevisiae. An enzyme with reverse (CoA-independent) ceramide synthase activity. AB - Ceramide is not only a core intermediate of sphingolipids but also an important modulator of many cellular events including apoptosis, cell cycle arrest, senescence, differentiation, and stress responses. Its turnover may be tightly regulated. However, little is known about the regulation of its metabolism because most enzymes responsible for its synthesis and breakdown have yet to be cloned. Here we report the cloning and characterization of the yeast gene YPC1 (YBR183w) by screening Saccharomyces cerevisiae genes whose overexpression bestows resistance to fumonisin B1. We demonstrate that the yeast gene YPC1 encodes an alkaline ceramidase activity responsible for the breakdown of dihydroceramide and phytoceramide but not unsaturated ceramide. YPC1 ceramidase activity was confirmed by in vitro studies using an Escherichia coli expression system. Importantly, YPC1p also has reverse activity, catalyzing synthesis of phytoceramide from palmitic acid and phytosphingosine. This ceramide synthase activity is CoA-independent and is resistant to fumonisin B1, thus explaining why YPC1 was cloned as a fumonisin B1-resistant gene. PMID- 10702248 TI - The roles of specific template nucleosides in the formation of stable transcription complexes by Escherichia coli RNA polymerase. AB - We have examined the effects of removing individual template nucleosides on promoter escape by Escherichia coli RNA polymerase in vitro. The ability of DNA templates containing random single nucleoside gaps generated by hydroxyl radical treatment to support the production of stable ternary transcription complexes was analyzed. On two templates containing different promoter and initial transcribed regions, we found that removal of nucleosides on the template strand in the region from -13 to at least +8 relative to the transcription start site interfered with ternary complex formation. The downstream border of this region varied for the two templates, suggesting an effect of the specific nucleotide sequence on the stability of intermediates in the promoter escape process. On the nontemplate strand, removal of nucleosides in the vicinity of the -10 consensus promoter element interfered with escape, whereas removal of nucleosides in the vicinity of the transcription start site actually enhanced the yield of ternary complexes. On one template, removal of nucleosides in an A-tract containing region upstream of the promoter caused a significant decrease in promoter escape, consistent with previous suggestions that contacts between this region and the RNA polymerase play a role in promoter binding and/or initiation. PMID- 10702249 TI - The Hsp organizer protein hop enhances the rate of but is not essential for glucocorticoid receptor folding by the multiprotein Hsp90-based chaperone system. AB - A system consisting of five purified proteins: Hsp90, Hsp70, Hop, Hsp40, and p23, acts as a machinery for assembly of glucocorticoid receptor (GR).Hsp90 heterocomplexes. Hop binds independently to Hsp90 and to Hsp70 to form a Hsp90.Hop.Hsp70.Hsp40 complex that is sufficient to convert the GR to its steroid binding form, and this four-protein complex will form stable GR.Hsp90 heterocomplexes if p23 is added to the system (Dittmar, K. D., Banach, M., Galigniana, M. D., and Pratt, W. B. (1998) J. Biol. Chem. 273, 7358-7366). Hop has been considered essential for the formation of receptor.Hsp90 heterocomplexes and GR folding. Here we use Hsp90 and Hsp70 purified free of all traces of Hop and Hsp40 to show that Hop is not required for GR.Hsp90 heterocomplex assembly and activation of steroid binding activity. Rather, Hop enhances the rate of the process. We also show that Hsp40 is not essential for GR folding by the five protein system but enhances a process that occurs less effectively when it is not present. By carrying out assembly in the presence of radiolabeled steroid to bind to the GR as soon as it is converted to the steroid binding state, we show that the folding change is brought about by only two essential components, Hsp90 and Hsp70, and that Hop, Hsp40, and p23 act as nonessential co-chaperones. PMID- 10702250 TI - Cell surface monkey CD9 antigen is a coreceptor that increases diphtheria toxin sensitivity and diphtheria toxin receptor affinity. AB - Monkey (Mk) CD9 antigen has been shown previously to increase the diphtheria toxin (DT) sensitivity of cells when co-expressed with Mk proHB-EGF (DT receptor). We have elucidated here the mechanism whereby Mk CD9 influences Mk proHB-EGF and present evidence that Mk CD9 is a coreceptor for DT. We observed that Mk CD9 not only increased the DT sensitivity but also increased the DT receptor affinity of cells. Furthermore, the higher the Mk CD9/Mk proHB-EGF ratio, the higher the affinity. In contrast, mouse (Ms) CD9 did not increase the toxin sensitivity or receptor affinity of cells when co-expressed with Mk proHB EGF. Using Mk/Ms chimeric CD9 molecules, we determined that the second extracellular domain of Mk CD9 is responsible for both increased sensitivity and receptor affinity. This domain of Mk CD9 also interacts with Mk proHB-EGF in a yeast two-hybrid system. Our findings thus suggest that Mk CD9 has a direct physical interaction with Mk proHB-EGF to form a DT receptor complex and that this contact may change the conformation of the receptor to increase DT binding affinity and consequently increase toxin sensitivity. We thus propose that Mk CD9 is a coreceptor for DT. PMID- 10702251 TI - 4-hydroxyretinoic acid, a novel substrate for human liver microsomal UDP glucuronosyltransferase(s) and recombinant UGT2B7. AB - It is suggested that formation of more polar metabolites of all-trans-retinoic acid (atRA) via oxidative pathways limits its biological activity. In this report, we investigated the biotransformation of oxidized products of atRA via glucuronidation. For this purpose, we synthesized 4-hydroxy-RA (4-OH-RA) in radioactive and nonradioactive form, 4-hydroxy-retinyl acetate (4-OH-RAc), and 5,6-epoxy-RA, all of which are major products of atRA oxidation. Glucuronidation of these retinoids by human liver microsomes and human recombinant UDP glucuronosyltransferases (UGTs) was characterized and compared with the glucuronidation of atRA. The human liver microsomes glucuronidated 4-OH-RA and 4 OH-RAc with 6- and 3-fold higher activity than atRA, respectively. Analysis of the glucuronidation products showed that the hydroxyl-linked glucuronides of 4-OH RA and 4-OH-RAc were the major products, as opposed to the formation of the carboxyl-linked glucuronide with atRA, 4-oxo-RA, and 5,6-epoxy-RA. We have also determined that human recombinant UGT2B7 can glucuronidate atRA, 4-OH-RA, and 4 OH-RAc with activities similar to those found in human liver microsomes. We therefore postulate that this human isoenzyme, which is expressed in human liver, kidney, and intestine, plays a key role in the biological fate of atRA. We also propose that atRA induces its own oxidative metabolism via a cytochrome P450 (CYP26) and is further biotransformed into glucuronides via UGT-mediated pathways. PMID- 10702252 TI - Topoisomerase II from Chlorella virus PBCV-1. Characterization of the smallest known type II topoisomerase. AB - Type II topoisomerases, a family of enzymes that govern topological DNA interconversions, are essential to many cellular processes in eukaryotic organisms. Because no data are available about the functions of these enzymes in the replication of viruses that infect eukaryotic hosts, this led us to express and characterize the first topoisomerase II encoded by one of such viruses. Paramecium bursaria chlorella virus 1 (PBCV-1) infects certain chlorella-like green algae and encodes a 120-kDa protein with a similarity to type II topoisomerases. This protein was expressed in Saccharomyces cerevisiae and was highly active in relaxation of both negatively and positively supercoiled plasmid DNA, catenation of plasmid DNA, and decatenation of kinetoplast DNA networks. Its optimal activity was determined, and the omission of Mg(2+) or its replacement with other divalent cations abolished DNA relaxation. All activities of the recombinant enzyme were ATP dependent. Increasing salt concentrations shifted DNA relaxation from a normally processive mechanism to a distributive mode. Thus, even though the PBCV-1 enzyme is considerably smaller than other eukaryotic topoisomerase II enzymes (whose molecular masses are typically 160-180 kDa), it displays all the catalytic properties expected for a type II topoisomerase. PMID- 10702253 TI - Identification of alternative spliced variants of human hypoxia-inducible factor 1alpha. AB - Mammalian cells are able to sense oxygen and regulate a number of genes in response to hypoxia. The transcription factor Hypoxia Inducible Factor-1 (HIF-1) was identified as an important key component of the hypoxia signaling pathway. HIF-1 is a heterodimer composed of two members of the basic helix-loop-helix transcription factor superfamily containing a PAS (PER-ARNT-SIM) domain: HIF 1alpha and HIF-1beta/ARNT. During the cloning by reverse transcriptase-polymerase chain reaction of the human HIF-1alpha subunit, we isolated two cDNA clones which corresponded to alternative splicing of the HIF-1alpha gene. Polymerase chain reaction analysis and sequencing revealed that both clones possessed three additional base pairs between exons 1 and 2. Also, one of them lacked 127 base pairs corresponding to exon 14. We demonstrate that the mRNA of this truncated form is expressed in several human cells lines and human skin but apparently not in rodents. When transfected in HEK 293 cells, the corresponding 736 amino acid protein (HIF-1alpha(736)) is regulated by hypoxia in a similar manner as the full length HIF-1alpha (HIF-1alpha(FL)). In luciferase transfection assays, both recombinant proteins HIF-1alpha(736) and HIF-1alpha(FL) dimerize with HIF 1beta/ARNT and activate the VEGF promoter upon hypoxia. However, the shorter HIF 1alpha isoform is 3-fold less active than HIF-1alpha(FL), a result consistent with the lack of the C-terminal transactivation domain. As expected, this small isoform can compete with the endogenous and transfected full-length HIF-1alpha. Altogether, these results suggest that the HIF-1alpha(736) isoform modulates gene expression upon hypoxia. PMID- 10702254 TI - Reactions of BglI and other type II restriction endonucleases with discontinuous recognition sites. AB - Type II restriction enzymes generally recognize continuous sequences of 4-8 consecutive base pairs on DNA, but some recognize discontinuous sites where the specified sequence is interrupted by a defined length of nonspecific DNA. To date, a mechanism has been established for only one type II endonuclease with a discontinuous site, SfiI at GGCCNNNNNGGCC (where N is any base). In contrast to orthodox enzymes such as EcoRV, dimeric proteins that act at a single site, SfiI is a tetramer that interacts with two sites before cleaving DNA. BglI has a similar recognition sequence (GCCNNNNNGGC) to SfiI but a crystal structure like EcoRV. BglI and several other endonucleases with discontinuous sites were examined to see if they need two sites for their DNA cleavage reactions. The enzymes included some with sites containing lengthy segments of nonspecific DNA, such as XcmI (CCANNNNNNNNNTGG). In all cases, they acted at individual sites. Elongated recognition sites do not necessitate unusual reaction mechanisms. Other experiments on BglI showed that it bound to and cleaved DNA in the same manner as EcoRV, thus further delineating a distinct group of restriction enzymes with similar structures and a common reaction mechanism. PMID- 10702255 TI - Direct evidence for the control of mitochondrial respiration by mitochondrial creatine kinase in oxidative muscle cells in situ. AB - The efficiency of stimulation of mitochondrial respiration in permeabilized muscle cells by ADP produced at different intracellular sites, e.g. cytosolic or mitochondrial intermembrane space, was evaluated in wild-type and creatine kinase (CK)-deficient mice. To activate respiration by endogenous production of ADP in permeabilized cells, ATP was added either alone or together with creatine. In cardiac fibers, while ATP alone activated respiration to half of the maximal rate, creatine plus ATP increased the respiratory rate up to its maximum. To find out whether the stimulation by creatine is a consequence of extramitochondrial [ADP] increase, or whether it directly correlates with ADP generation by mitochondrial CK in the mitochondrial intermembrane space, an exogenous ADP-trap system was added to rephosphorylate all cytosolic ADP. Under these conditions, creatine plus ATP still increased the respiration rate by 2.5 times, compared with ATP alone, for the same extramitochondrial [ADP] of 14 microM. Moreover, this stimulatory effect of creatine, observed in wild-type cardiac fibers disappeared in mitochondrial CK deficient, but not in cytosolic CK-deficient muscle. It is concluded that respiration rates can be dissociated from cytosolic [ADP], and ADP generated by mitochondrial CK is an important regulator of oxidative phosphorylation. PMID- 10702256 TI - SPACRCAN, a novel human interphotoreceptor matrix hyaluronan-binding proteoglycan synthesized by photoreceptors and pinealocytes. AB - The interphotoreceptor matrix is a unique extracellular complex occupying the interface between photoreceptors and the retinal pigment epithelium in the fundus of the eye. Because of the putative supportive role in photoreceptor maintenance, it is likely that constituent molecules play key roles in photoreceptor function and may be targets for inherited retinal disease. In this study we identify and characterize SPACRCAN, a novel chondroitin proteoglycan in this matrix. SPACRCAN was cloned from a human retinal cDNA library and the gene localized to chromosome 3q11.2. Analysis of SPACRCAN mRNA and protein revealed that SPACRCAN is expressed exclusively by photoreceptors and pinealocytes. SPACRCAN synthesized by photoreceptors is localized to the interphotoreceptor matrix where it surrounds both rods and cones. The functional protein contains 1160 amino acids with a large central mucin domain, three consensus sites for glycosaminoglycan attachment, two epidermal growth factor-like repeats, a putative hyaluronan binding motif, and a potential transmembrane domain near the C-terminal. Lectin and Western blotting indicate an M(r) around 400,000 before and 230,000 after chondroitinase ABC digestion. Removal of N- and O-linked oligosaccharides reduces the M(r) to approximately 160,000, suggesting that approximately 60% of the mass of SPACRCAN is carbohydrate. Finally, we demonstrate that SPACRCAN binds hyaluronan and propose that associations between SPACRCAN and hyaluronan may be involved in organization of the insoluble interphotoreceptor matrix, particularly as SPACRCAN is the major proteoglycan present in this matrix. PMID- 10702257 TI - Effect of pH and monovalent cations on the formation of quinonoid intermediates of the tryptophan synthase alpha(2)beta(2) complex in solution and in the crystal. AB - Quinonoid intermediates play a key role in the catalytic mechanism of pyridoxal 5'-phosphate-dependent enzymes. Whereas the structures of other pyridoxal 5' phosphate-bound intermediates have been determined, the structure of a quinonoid species has not yet been reported. Here, we investigate factors controlling the accumulation and stability of quinonoids formed at the beta-active site of tryptophan synthase both in solution and the crystal. The quinonoids were obtained by reacting the alpha-aminoacrylate Schiff base with different nucleophiles, focusing mainly on the substrate analogs indoline and beta mercaptoethanol. In solution, both monovalent cations (Cs(+) or Na(+)) and alkaline pH increase the apparent affinity of indoline and favor accumulation of the indoline quinonoid. A similar pH dependence is observed when beta mercaptoethanol is used. As indoline and beta-mercaptoethanol exhibit very distinct ionization properties, this finding suggests that nucleophile binding and quinonoid stability are controlled by some ionizable protein residue(s). In the crystal, alkaline pH favors formation of the indoline quinonoid as in solution, but the effect of cations is markedly different. In the absence of monovalent metal ions the quinonoid species accumulates substantially, whereas in the presence of sodium ions the accumulation is modest, unless alpha-subunit ligands are also present. Alpha-subunit ligands not only favor the formation of the intermediate, but also reduce significantly its decay rate. These findings define experimental conditions suitable for the stabilization of the quinonoid species in the crystal, a critical prerequisite for the determination of the three-dimensional structure of this intermediate. PMID- 10702258 TI - Partial uncoupling of the mitochondrial membrane by a heterozygous null mutation in the gene encoding the gamma- or delta-subunit of the yeast mitochondrial ATPase. AB - Prior genetic studies indicated that the yeast mitochondrial ATP synthase can be assembled into enzyme complexes devoid of the gamma-, delta-, or epsilon-subunits (Lai-Zhang, J., Xiao, Y., and Mueller, D. M. (1999) EMBO J. 18, 58-64). These subunit-deficient complexes were postulated to uncouple the mitochondrial membrane thereby causing negative cellular phenotypes. This study provides biochemical and additional genetic data that support this hypothesis. The genetic data indicate that in a diploid cell, a heterozygous deletion mutation in the gene encoding the gamma- or delta-subunit of the ATPase is semidominant negative due to a decrease in the gene number from 2 to 1. However, the heterozygous atp2Delta mutation is epistatic to the heterozygous mutation in the gene encoding gamma or delta, suggesting that the semidominant negative effect is because of a gain of activity in the cells. Biochemical studies using mitochondria isolated from the yeast strains that are heterozygous for a mutation in gamma or delta indicate that the mitochondria are partially uncoupled. These results support the hypothesis that the negative phenotypes are caused by the formation of a gamma- or delta-less ATP synthase complex that is uncoupled. PMID- 10702259 TI - Loss of the effector function in a transducin-alpha mutant associated with Nougaret night blindness. AB - A missense mutation, G38D, was found in the rod transducin alpha subunit (Galpha(t)) in individuals with the Nougaret form of dominant stationary night blindness. To elucidate the mechanism of Nougaret night blindness, we have examined the key functional properties of the mutant transducin. Our data show that the G38D mutation does not alter the interaction between Galpha(t) and Gbetagamma(t) or activation of transducin by photoexcited rhodopsin (R*). The mutant Galpha(t) has only a modestly (approximately 2.5-fold) reduced k(cat) value for GTP hydrolysis. The GTPase activity of Galpha(t)G38D can be accelerated by photoreceptor regulator of G protein signaling, RGS9. Analysis of the Galpha(t)G38D interaction with cGMP phosphodiesterase revealed marked impairment of the mutant effector function. Galpha(t)G38D completely fails to bind the inhibitory PDE gamma subunit and activate the enzyme. Altogether, our results demonstrate a novel molecular mechanism in dominant stationary night blindness. In contrast to known forms of the disease caused by constitutive activation of the visual cascade, the Nougaret form has its origin in attenuated visual signaling due to loss of effector function by transducin G38D mutant. PMID- 10702260 TI - Building a thermostable membrane protein. AB - The poor stability of membrane proteins in detergent solution is one of the main technical barriers to their structural and functional characterization. Here we describe a solution to this problem for diacylglycerol kinase (DGK), an integral membrane protein from Escherichia coli. Twelve enhanced stability mutants of DGK were obtained using a simple screen. Four of the mutations were combined to create a quadruple mutant that had improved stability in a wide range of detergents. In n-octylglucoside, the wild-type DGK had a thermal inactivation half-life of 6 min at 55 degrees C, while the quadruple mutant displayed a half life of 35 min at 80 degrees C. In addition, the quadruple mutant had improved thermodynamic stability. Our approach should be applicable to other membrane proteins that can be conveniently assayed. PMID- 10702261 TI - Ca(2+), Sr(2+), and Ba(2+) identify distinct regulatory sites on adenylyl cyclase (AC) types VI and VIII and consolidate the apposition of capacitative cation entry channels and Ca(2+)-sensitive ACs. AB - Ca(2+)-sensitive adenylyl cyclases may act as early integrators of the two major second messenger-signaling pathways mediated by Ca(2+) and cAMP. Ca(2+) stimulation of adenylyl cyclase type I (ACI) and adenylyl cyclase type VIII (ACVIII) is mediated by calmodulin and the site on these adenylyl cyclases that interacts with calmodulin has been defined. By contrast, the mechanism whereby Ca(2+) inhibits adenylyl cyclase type V (ACV) and adenylyl cyclase type VI (ACVI) is unknown. In this study, Ca(2+), Sr(2+), and Ba(2+) were compared to probe the involvement of E-F hand-like domains in both Ca(2+) stimulation and inhibition of ACVIII and ACVI, respectively. HEK 293 cells transfected with ACVIII cDNA and C6 2B glioma cells (where the endogenous adenylyl cyclases is predominantly ACVI) were used to compare the effects of these three cations in in vitro and in vivo measurements. The in vitro data identified two Ca(2+) regulatory sites for both ACVIII and ACVI. Strikingly different potency series for these cations at mediating high affinity stimulation and inhibition of ACVIII and ACVI, respectively, effectively rule out the possibility that calmodulin or proteins utilizing similar Ca(2+)-binding motifs mediate inhibition of ACVI. On the other hand, the low affinity inhibition that is common to both ACVIII and ACVI showed virtually identical potency profiles for the IIa cation series, indicating a common site of action. Remarkably, whereas Sr(2+) was rather ineffective at regulating these cyclases (particularly ACVI) in vitro, adequate concentrations accumulated in the vicinity of these enzymes as a consequence of capacitative cation entry to partially regulate both of these activities in vivo. This latter finding consolidates earlier observations that Ca(2+)-sensitive adenylyl cyclases detect and respond to capacitative cation entry rather than global cytosolic cation concentrations. PMID- 10702262 TI - Reversible G(1) arrest induced by inhibition of the epidermal growth factor receptor tyrosine kinase requires up-regulation of p27(KIP1) independent of MAPK activity. AB - We have used quinazoline inhibitors of the epidermal growth factor receptor (EGFR) tyrosine kinase to study the link between EGFR signaling and G(1) to S traverse. Treatment of A431 and MDA-468 human tumor cells with 0.1-10 microM AG 1478 inhibited basal and ligand-stimulated EGFR phosphorylation without a decrease in receptor content, EGF-binding sites, or binding affinity. Incubation of A431 cells with 0.1-1 microM AG-1517 abrogated (125)I-EGF internalization. Both AG-1478 and AG-1517 markedly inhibited A431 and MDA-468 colony formation in soft agarose at concentrations between 0.01 and 1 microM. Daily injections of AG 1478 at 50 mg/kg delayed A431 tumor formation in athymic nude mice. A transient exposure of A431 cells to AG-1478 resulted in a dose-dependent up-regulation of the cyclin-dependent kinase inhibitor p27, down-regulation of cyclin D1 and of active MAPK, and hypophosphorylation of the retinoblastoma protein (Rb). These changes were temporally associated with recruitment of tumor cells in G(1) phase and a marked reduction of the proportion of cells in S phase. Upon removal of the kinase inhibitor, EGFR and Rb phosphorylation and the levels of cyclin D1 protein were quickly restored, but the cells did not reenter S phase until p27 protein levels were decreased. Phosphorothioate p27 oligonucleotides decreased p27 protein in A431 cells and abrogated the quinazoline-mediated G(1) arrest. Treatment of A431 cells with PD 098509, a synthetic inhibitor of MEK1, inhibited MAPK activity without inducing G(1) arrest or increasing the levels of p27. However, treatment with LY 294002, an inhibitor of phosphatidylinositol 3-kinase (PI3K), inhibited basal Akt activity, up-regulated p27, and recruited cells in G(1). These data suggest that p27 is required for the growth arrest that follows interruption of the EGFR kinase in receptor-overexpressing cells. In addition, the G(1) arrest and up-regulation of p27 resulting from EGFR blockade are not due to the interruption of MAPK, but to the interruption of constitutively active PI3K function. PMID- 10702263 TI - Phosphorylation of the membrane proximal region of tumor necrosis factor receptor CD120a (p55) at ERK consensus sites. AB - The interaction of tumor necrosis factor-alpha with its receptor CD120a (p55) initiates downstream signaling cascades that include the activation of the mitogen-activated protein kinase (MAPK), p42(mapk/erk2). The membrane proximal region of CD120a (p55) is Ser-, Thr-, and Pro-rich and contains four mitogen activated protein kinase consensus phosphorylation sites. In recent work, we showed that CD120a (p55) itself is a target of phosphorylation by p42(mapk/erk2), and after phosphorylation, the receptor is redistributed from the cell surface and Golgi complex to intracellular tubular structures associated with elements of the endoplasmic reticulum. The goal of this study was to define the specific amino acid residues that are phosphorylated. Deletional mutagenesis of the cytoplasmic domain of CD120a (p55) indicated that two sites located between residues 207-254 and 250-300 were phosphorylated predominantly on Thr and Ser residues, respectively. Site-directed mutagenesis of Ser and Thr residues contained within the extracellular signal-regulated kinase (ERK) consensus sequences indicated that the preferred residues were Thr-236 and Ser-270. Primary phosphorylation at these sites appeared to enable subsequent phosphorylation at Ser-240 and Ser-244, although the level of phosphorylation of these latter two sites was less than the preferred sites. Through the use of specific ligation of CD120a (p55) alone and mice deficient in CD120a (p55), CD120b (p75), or both receptors, CD120a (p55) was shown to be necessary and sufficient for the induction of kinase activity. These findings thus suggest that the phosphorylation of Thr-236 and Ser-270 within the membrane proximal region of CD120a (p55) are the preferred sites of phosphorylation by p42(mapk/erk2) and may set in motion phosphorylation at other sites. PMID- 10702264 TI - Evidence for a primary endocytic vesicle involved in synaptic vesicle biogenesis. AB - The regulated release of neurotransmitters at synapses is mediated by the fusion of neurotransmitter-filled synaptic vesicles with the plasma membrane. Continuous synaptic activity relies on the constant recycling of synaptic vesicle proteins into newly formed synaptic vesicles. At least two different mechanisms are presumed to mediate synaptic vesicle biogenesis at the synapse as follows: direct retrieval of synaptic vesicle proteins and lipids from the plasma membrane, and indirect passage of synaptic vesicle proteins through an endosomal intermediate. We have identified a vesicle population with the characteristics of a primary endocytic vesicle responsible for the recycling of synaptic vesicle proteins through the indirect pathway. We find that synaptic vesicle proteins colocalize in this vesicle with a variety of proteins known to recycle from the plasma membrane through the endocytic pathway, including three different glucose transporters, GLUT1, GLUT3, and GLUT4, and the transferrin receptor. These vesicles differ from "classical" synaptic vesicles in their size and their generic protein content, indicating that they do not discriminate between synaptic vesicle-specific proteins and other recycling proteins. We propose that these vesicles deliver synaptic vesicle proteins that have escaped internalization by the direct pathway to endosomes, where they are sorted from other recycling proteins and packaged into synaptic vesicles. PMID- 10702265 TI - The RcsAB box. Characterization of a new operator essential for the regulation of exopolysaccharide biosynthesis in enteric bacteria. AB - The interaction of the two transcriptional regulators RcsA and RcsB with a specific operator is a common mechanism in the activation of capsule biosynthesis in enteric bacteria. We describe RcsAB binding sites in the wza promoter of the operon for colanic acid biosynthesis in Escherichia coli K-12, in the galF promoter of the operon for K2 antigen biosynthesis in Klebsiella pneumoniae, and in the tviA (vipR) promoter of the operon for Vi antigen biosynthesis in Salmonella typhi. We further show the interaction of RcsAB with the rcsA promoters of various species, indicating that rcsA autoregulation also depends on the presence of both proteins. The compilation of all identified RcsAB binding sites revealed the conserved core sequence TaAGaatatTCctA, which we propose to be termed RcsAB box. The RcsAB box is also part of Bordetella pertussis BvgA binding sites and may represent a more distributed recognition motif within the LuxR superfamily of transcriptional regulators. The RcsAB box is essential for the induction of Rcs-regulated promoters. Site-specific mutations of conserved nucleotides in the RcsAB boxes of the E. coli wza and rcsA promoters resulted in an exopolysaccharide-negative phenotype and in the reduction of reporter gene expression. PMID- 10702266 TI - Dissecting G protein-coupled receptor signaling pathways with membrane-permeable blocking peptides. Endogenous 5-HT(2C) receptors in choroid plexus epithelial cells. AB - To determine the intracellular signaling mechanism of the 5-HT(2C) receptor endogenously expressed in choroid plexus epithelial cells, we implemented a strategy of targeted disruption of protein-protein interactions. This strategy entails the delivery of conjugated membrane-permeable peptides that disrupt domain interaction at specific steps in the signaling cascade. As proof of concept, two peptides targeted against receptor-G protein interaction domains were examined. Only G(q)CT, which targets the receptor-G(q) protein interacting domain, disrupted 5-HT(2C) receptor-mediated phosphatidylinositide hydrolysis. G(s)CT, targeting the receptor-G(s) protein, disrupted beta2 adrenergic receptor mediated activation of cAMP but not 5-HT(2C) receptor-mediated phosphatidylinositide hydrolysis. The peptide MPS-PLCbeta1M, mimicking the domain of phospholipase Cbeta1 (PLCbeta1) interacting with active Galpha(q), also blocked 5-HT(2C) receptor activation. In contrast, peptides PLCbeta2M and Phos that bind to and sequester free Gbetagamma subunits were ineffective at blocking 5-HT(2C) receptor-mediated phosphoinositol turnover. However, both peptides disrupted Gbetagamma-mediated alpha(2A) adrenergic receptor activation of mitogen activated protein kinase. These results provide the first direct demonstration that active Galpha(q) subunits mediate endogenous 5-HT(2C) receptor activation of PLCbeta and that Gbetagamma subunits released from Galpha(q) heterotrimeric proteins are not involved. Comparable results were obtained with metabotropic glutamate receptor 5 expressed in astrocytes. Thus, conjugated, membrane permeable peptides are effective tools for the dissection of intracellular signals. PMID- 10702267 TI - Essential role of a single arginine of photosystem I in stabilizing the electron transfer complex with ferredoxin. AB - PsaE is one of the photosystem I subunits involved in ferredoxin binding. The central role of arginine 39 of this 8-kDa peripheral polypeptide has been established by a series of mutations. The neutral substitution R39Q leads to a 250-fold increase of the dissociation constant K(d) of the photosystem I ferredoxin complex, as large as the increase induced by PsaE deletion. At pH 8.0, this K(d) value strongly depends on the charge of the residue substituting Arg 39: 0.22 microM for wild type, 1.5 microM for R39K, 56 microM for R39Q, and more than 100 microM for R39D. The consequences of arginine 39 substitution for the titratable histidine were analyzed as a function of pH. The K(d) value of R39H is increased 140 times at pH 8.0 but only 5 times at pH 5.8, which is assigned to the protonation of histidine at low pH. In the mutant R39Q, the association rate of ferredoxin was decreased 3-fold compared with wild type, whereas an 80-fold increase is calculated for the dissociation rate. We propose that a major contribution of PsaE is to provide a prominent positive charge at position 39 for controlling the electrostatic interaction and lifetime of the complex with ferredoxin. PMID- 10702268 TI - Multiple phosphorylation events regulate the activity of the mannitol transcriptional regulator MtlR of the Bacillus stearothermophilus phosphoenolpyruvate-dependent mannitol phosphotransferase system. AB - D-mannitol is taken up by Bacillus stearothermophilus and phosphorylated via a phosphoenolpyruvate-dependent phosphotransferase system (PTS). Transcription of the genes involved in mannitol uptake in this bacterium is regulated by the transcriptional regulator MtlR, a DNA-binding protein whose affinity for DNA is controlled by phosphorylation by the PTS proteins HPr and IICB(mtl). The mutational and biochemical studies presented in this report reveal that two domains of MtlR, PTS regulation domain (PRD)-I and PRD-II, are phosphorylated by HPr, whereas a third IIA-like domain is phosphorylated by IICB(mtl). An involvement of PRD-I and the IIA-like domain in a decrease in affinity of MtlR for DNA and of PRD-II in an increase in affinity is demonstrated by DNA footprint experiments using MtlR mutants. Since both PRD-I and PRD-II are phosphorylated by HPr, PRD-I needs to be dephosphorylated by IICB(mtl) and mannitol to obtain maximal affinity for DNA. This implies that a phosphoryl group can be transferred from HPr to IICB(mtl) via MtlR. Indeed, this transfer could be demonstrated by the phosphoenolpyruvate-dependent formation of [(3)H]mannitol phosphate in the absence of IIA(mtl). Phosphoryl transfer experiments using MtlR mutants revealed that PRD-I and PRD-II are dephosphorylated via the IIA-like domain. Complementation experiments using two mutants with no or low phosphoryl transfer activity showed that phosphoryl transfer between MtlR molecules is possible, indicating that MtlR-MtlR interactions take place. Phosphorylation of the same site by HPr and dephosphorylation by IICB(mtl) have not been described before; they could also play a role in other PRD-containing proteins. PMID- 10702269 TI - cAMP response element-binding protein-binding protein binds to human papillomavirus E2 protein and activates E2-dependent transcription. AB - cAMP response element-binding protein-binding protein (CBP) is a eucaryotic transcriptional co-activator that contains multiple protein-protein interaction domains for association with various transcription factors, components of the basal transcriptional apparatus, and other co-activator proteins. Here, we report that CBP is also a co-activator of the human papillomavirus (HPV) E2 protein, which is a sequence-specific transcription/replication factor. We provide biochemical, genetic, and functional evidence that CBP binds directly to HPV E2 in vivo and in vitro and activates E2-dependent transcription. Mutations in an amphipathic helix within HPV-18 E2 abolish its transcriptional activation properties and its ability to bind to CBP. Furthermore, the binding of CBP to E2 was shown to be necessary for E2-dependent transcription. Interestingly, the histone acetyltransferase activity of CBP plays a role in CBP activation of E2 dependent transcription. PMID- 10702270 TI - The alpha(4) integrin subunit Tyr(187) has a key role in alpha(4)beta(7) dependent cell adhesion. AB - The integrin alpha(4)beta(7) is the cell adhesion receptor for the mucosal vascular addressin MAdCAM-1, and this interaction is dominant in lymphocyte homing to Peyer's patch high endothelial venules, and plays key roles in lymphocyte recruitment at sites of inflammation. To identify alpha(4) subunit amino acids important for alpha(4)beta(7)/MAdCAM-1 interaction, we expressed mutant alpha(4) and wild type beta(7) chains in K562 cells and analyzed the effect of the mutations on cell adhesion to a soluble MAdCAM-1 (sMAdCAM-1-Ig). Transfectants expressing mutated alpha(4) at Tyr(187) displayed a substantial decrease in adhesion to this ligand, which was associated with a reduced alpha(4)beta(7)/sMAdCAM-1-Ig interaction, as determined by soluble binding assays. Addition of Mn(2+) to the adhesion assays did not restore the impaired adhesion. Mutations at alpha(4) Gln(152)Asp(153) also affected transfectant adhesion to sMAdCAM-1-Ig, but did not involve an alteration of alpha(4)beta(7)/MAdCAM-1 binding, and adhesion was restored by Mn(2+). Instead, mutations at alpha(4) Asn(123)Glu(124) did not affect this adhesion. Mutation of alpha(4) Tyr(187) abolished alpha(4)beta(7)-mediated cell adhesion to CS 1/fibronectin, an additional ligand for alpha(4)beta(7), while alpha(4) Gln(152)Asp(153) transfectant mutants showed a reduced adhesion. These results identify alpha(4) Tyr(187) as a key residue during receptor alpha(4)beta(7)/ligand interactions, indicating that it plays important roles in alpha(4)beta(7)-mediated leukocyte adhesion, and provide a potential target for therapeutic intervention in several inflammatory pathologies. PMID- 10702271 TI - Interleukin (IL)-7 induces rapid activation of Pyk2, which is bound to Janus kinase 1 and IL-7Ralpha. AB - Interleukin-7 (IL-7) receptor signaling begins with activation of the Janus tyrosine kinases Jak1 and Jak3, which are associated with the receptor complex. To identify potential targets of these kinases, we examined Pyk2 (a member of the focal adhesion kinase family) using an IL-7-dependent murine thymocyte line, D1. We demonstrate that stimulation of D1 (or normal pro-T) cells by IL-7 rapidly increased tyrosine phosphorylation and enzymatic activity of Pyk2, with kinetics slightly lagging that of Jak1 and Jak3 phosphorylation. Conversely, IL-7 withdrawal resulted in a marked decrease of Pyk2 phosphorylation. Pyk2 was found to be physically associated with Jak1 prior to IL-7 stimulation and to increase its association with IL-7Ralpha chain following IL-7 stimulation. Pyk2 appeared to be involved in cell survival, because antisense Pyk2 accelerated the cell death process. Activation of Pyk2 via the muscarinic and nicotinic receptors using carbachol or via intracellular Ca(2+) rise using ionomycin/phorbol myristate acetate promoted survival in the absence of IL-7. These data support a role for Pyk2 in coupling Jak signaling to the trophic response. PMID- 10702272 TI - Disruption of Raf-1/heat shock protein 90 complex and Raf signaling by dexamethasone in mast cells. AB - Antigen stimulation of mast cells via the IgE receptor, FcepsilonRI, results in the recruitment of the cytosolic tyrosine kinase, Syk, and the activation of various signaling cascades. One of these, the extracellular signal-regulated kinase (ERK2) cascade, is inhibited by low concentrations of the immunosuppressant drug, dexamethasone, probably at a step prior to the activation of Raf-1 (Rider, L. G., Hirasawa, N., Santini, F., and Beaven, M. A. (1996) J. Immunol. 157, 2374-2380). We now show that treatment of cultured RBL-2H3 mast cells with nanomolar concentrations of dexamethasone causes dissociation of the Raf-1.heat shock protein 90 (Hsp90) complex. Raf-1 bereft of this protein fails to associate with the membrane or Ras in antigen-stimulated cells. Upstream events such as the Syk-dependent phosphorylation of Shc, the engagement of Shc with the adapter protein, Grb2, and the activation of Ras itself are unaffected. Interestingly, the counterpart of Raf-1 in the c-Jun N-terminal kinase (JNK) cascade, MEKK-1 (mitogen-activated protein kinase/ERK kinase), is similarly associated with Hsp90, and this association as well as the activation of MEKK-1 are disrupted by dexamethasone treatment. Disruption of the ERK and JNK cascades at the level of Raf-1 and MEKK-1 could account for the inhibitory action of dexamethasone on the generation of inflammatory mediators in stimulated mast cells. PMID- 10702273 TI - Ionomycin, a carboxylic acid ionophore, transports Pb(2+) with high selectivity. AB - Studies utilizing phospholipid vesicle loaded with chelator/indicators for polyvalent cations show that ionomycin transports divalent cations with the selectivity sequence Pb(2+) > Cd(2+) > Zn(2+) > Mn(2+) > Ca(2+) > Cu(2+) > Co(2+) > Ni(2+) > Sr(2+). The selectivity of this ionophore for Pb(2+) is in contrast to that observed for A23178 and 4-BrA23187, which transport Pb(2+) at efficiencies that are intermediate between those of other cations. When the selectivity difference of ionomycin for Pb(2+) versus Ca(2+) was calculated from relative rates of transport, with either cation present individually and all other conditions held constant, a value of approximately 450 was obtained. This rose to approximately 3200 when both cations were present and transported simultaneously. 1 microM Pb(2+) inhibited the transport of 1 mM Ca(2+) by approximately 50%, whereas the rate of Pb(2+) transport approached a maximum at a concentration of 10 microM Pb(2+) when 1 mM Ca(2+) was also present. Plots of log rate versus log ionomycin or log Pb(2+) concentration indicated that the transporting species is of 1:1 stoichiometry, ionophore to Pb(2+), but that complexes containing an additional Pb(2+) may occur. The species transporting Pb(2+) may include H.IPb.OH, wherein ionomycin is ionized once and the presence of OH(-) maintains charge neutrality. Ionomycin retained a high efficiency for Pb(2+) transport in A20 B lymphoma cells loaded with Indo-1. Both Pb(2+) entry and efflux were observed. Ionomycin should be considered primarily as an ionophore for Pb(2+), rather than Ca(2+), of possible value for the investigation and treatment of Pb(2+) intoxication. PMID- 10702274 TI - Macromolecular inhibitors of HIV-1 protease. Characterization of designed heterodimers. AB - Defective variants of human immunodeficiency virus type 1 (HIV-1) protease (HIV PR) have been engineered to inhibit wild-type (wt) HIV PR activity. These variants were designed to promote the formation of heterodimers and to destabilize the formation of inactive variant homodimers of HIV-1 protease through substitutions at Asp-25, Ile-49, and Gly-50 (Babe, L. M., Rose, J., and Craik, C. S. (1995) Proc. Natl. Acad. Sci. U. S. A. 92, 10069-10073; McPhee, F., Good, A. C., Kuntz, I. D., and Craik, C. S. (1996) Proc. Natl. Acad. Sci. U. S. A. 93, 11477-11481). The mechanism of action of these dominant-negative inhibitors was established using recombinantly expressed defective monomers. The defective monomers were refolded in vitro in the presence of wt HIV PR and showed dose-dependent inhibition of proteolytic activity. This inhibition was shown to result from the formation of inactive heterodimers between defective and wt HIV PR monomers. Heterodimer formation was detected by (i) isolating refolded, inactive heterodimers using histidine-tagged defective monomers and (ii) isolating heterodimers from bacteria coexpressing both wt and defective variants of HIV PR. Single-chain variants of HIV PR, in which the C terminus of the wt HIV PR monomer was covalently tethered to the N terminus of the defective monomer, were also expressed and analyzed. Thermal denaturation of these single-chain heterodimers using differential scanning calorimetry revealed a 1.5-7.2 degrees C greater thermal stability than single-chain wt HIV PR. The thermodynamic trend shown by these three variants mirrors their relative inhibition in provirus transfection assays. These data support the model that the effects seen both in tissue culture and in vitro arise from an increase in stability conferred on these heterodimers by interface mutations and identifies heterodimer formation as their mechanism of inhibition. PMID- 10702275 TI - Lack of oxidative phosphorylation and low mitochondrial membrane potential decrease susceptibility to apoptosis and do not modulate the protective effect of Bcl-x(L) in osteosarcoma cells. AB - We explored the role of low mitochondrial membrane potential (DeltaPsim) and the lack of oxidative phosphorylation in apoptosis by assessing the susceptibility of osteosarcoma cell lines with and without mitochondrial DNA to staurosporine induced death. Our cells without mitochondrial DNA had low DeltaPsim and no functional oxidative phosphorylation. Contrary to our expectation, these cells were more resistant to staurosporine-induced death than were the parental cells. This reduced susceptibility was associated with decreased activation of caspase 3 but not with the mitochondrial permeability transition pore or cytochrome c release from the mitochondria. Apoptosis in both cell lines was associated with an increase in DeltaPsim. Bcl-x(L) could protect both cell types against caspase 3 activation and apoptosis by a mechanism that does not appear to be mediated by mitochondrial function or modulation of DeltaPsim. Nevertheless, we found that Bcl-x(L) expression can stimulate cell respiration in cells with mitochondrial DNA. Our results showed that the lack of functional oxidative phosphorylation and/or low mitochondrial membrane potential are associated with an antiapoptotic effect, possibly contributing to the development of some types of cancer. It also reinforces a model in which Bcl-x(L) can exert an antiapoptotic effect by stimulating oxidative phosphorylation and/or inhibiting caspase activation. PMID- 10702276 TI - The protein core of the proteoglycan perlecan binds specifically to fibroblast growth factor-7. AB - Perlecan is a multifaceted heparan sulfate proteoglycan that is expressed not only as an intrinsic constituent of basement membranes but also as a cell-surface and pericellular proteoglycan. Perlecan functions as a ligand reservoir for various growth factors that become stabilized against misfolding or proteolysis and acts as a co-receptor for basic fibroblast growth factor by augmenting high affinity binding and receptor activation. These biological properties are mediated by the heparan sulfate moiety. Rather little is known about the protein core's mediation of functions. We have recently discovered that fibroblast growth factor-7 (FGF7) binds to perlecan protein core and that exogenous perlecan efficiently reconstitutes FGF7 mitogenic activity in perlecan-deficient cells. In this report we examined the specific binding of FGF7 to various domains and subdomains of perlecan protein core. Using several experimental approaches including overlay protein assays, radioligand binding experiments, and the yeast two-hybrid system, we demonstrate that FGF7 binds specifically to the N-terminal half of domain III and to a lesser extent to domain V, with affinity constants in the range of 60 nM. Thus, perlecan protein core should be considered a novel biological ligand for FGF7, an interaction that could influence cancer growth and tissue remodeling. PMID- 10702277 TI - 2-(oxalylamino)-benzoic acid is a general, competitive inhibitor of protein tyrosine phosphatases. AB - Protein-tyrosine phosphatases (PTPs) are critically involved in regulation of signal transduction processes. Members of this class of enzymes are considered attractive therapeutic targets in several disease states, e.g. diabetes, cancer, and inflammation. However, most reported PTP inhibitors have been phosphorus containing compounds, tight binding inhibitors, and/or inhibitors that covalently modify the enzymes. We therefore embarked on identifying a general, reversible, competitive PTP inhibitor that could be used as a common scaffold for lead optimization for specific PTPs. We here report the identification of 2 (oxalylamino)-benzoic acid (OBA) as a classical competitive inhibitor of several PTPs. X-ray crystallography of PTP1B complexed with OBA and related non-phosphate low molecular weight derivatives reveals that the binding mode of these molecules to a large extent mimics that of the natural substrate including hydrogen bonding to the PTP signature motif. In addition, binding of OBA to the active site of PTP1B creates a unique arrangement involving Asp(181), Lys(120), and Tyr(46). PTP inhibitors are essential tools in elucidating the biological function of specific PTPs and they may eventually be developed into selective drug candidates. The unique enzyme kinetic features and the low molecular weight of OBA makes it an ideal starting point for further optimization. PMID- 10702278 TI - Binding of an antibody mimetic of the human low density lipoprotein receptor to apolipoprotein E is governed through electrostatic forces. Studies using site directed mutagenesis and molecular modeling. AB - Monoclonal antibody 2E8 is specific for an epitope that coincides with the binding site of the low density lipoprotein receptor (LDLR) on human apoE. Its reactivity with apoE variants resembles that of the LDLR: it binds well with apoE3 and poorly with apoE2. The heavy chain complementarity-determining region (CDRH) 2 of 2E8 shows homology to the ligand-binding domain of the LDLR. To define better the structural basis of the 2E8/apoE interaction and particularly the role of electrostatic interactions, we generated and characterized a panel of 2E8 variants. Replacement of acidic residues in the 2E8 CDRHs showed that Asp(52), Glu(53), and Asp(56) are essential for high-affinity binding. Although Asp(31) (CDRH1), Glu(58) (CDRH2), and Asp(97) (CDRH3) did not appear to be critical, the Asp(97) --> Ala variant acquired reactivity with apoE2. A Thr(57) - > Glu substitution increased affinity for both apoE3 and apoE2. The affinities of wild-type 2E8 and variants for apoE varied inversely with ionic strength, suggesting that electrostatic forces contribute to both antigen binding and isoform specificity. We propose a model of the 2E8.apoE immune complex that is based on the 2E8 and apoE crystal structures and that is consistent with the apoE binding properties of wild-type 2E8 and its variants. Given the similarity between the LDLR and 2E8 in terms of specificity, the LDLR/ligand interaction may also have an important electrostatic component. PMID- 10702279 TI - The yeast mitochondrial citrate transport protein. Probing the roles of cysteines, Arg(181), and Arg(189) in transporter function. AB - Utilizing site-directed mutagenesis in combination with chemical modification of mutated residues, we have studied the roles of cysteine and arginine residues in the mitochondrial citrate transport protein (CTP) from Saccharomyces cerevisiae. Our strategy consisted of the sequential replacement of each of the four endogenous cysteine residues with Ser or in the case of Cys(73) with Val. Wild type and mutated forms of the CTP were overexpressed in Escherichia coli, purified, and reconstituted in phospholipid vesicles. During the sequential replacement of each Cys, the effects of both hydrophilic and hydrophobic sulfhydryl reagents were examined. The data indicate that Cys(73) and Cys(256) are primarily responsible for inhibition of the wild-type CTP by hydrophilic sulfhydryl reagents. Experiments conducted with triple Cys replacement mutants (i.e. Cys(192) being the only remaining Cys) indicated that sulfhydryl reagents no longer inhibit but in fact stimulate CTP function 2-3-fold. Following the simultaneous replacement of all four endogenous Cys, the functional properties of the resulting Cys-less CTP were shown to be quite similar to those of the wild type protein. Finally, utilizing the Cys-less CTP as a template, the roles of Arg(181) and Arg(189), two positively charged residues located within transmembrane domain IV, in CTP function were examined. Replacement of either residue with a Cys abolishes function, whereas replacement with a Lys or a Cys that is subsequently covalently modified with (2-aminoethyl)methanethiosulfonate hydrobromide, a reagent that restores positive charge at this site, supports CTP function. The results clearly show that positive charge at these two positions is essential for CTP function, although the chemistry of the guanidinium residue is not. Finally, these studies: (i) definitely demonstrate that Cys residues do not play an important role in the mechanism of the CTP; (ii) prove the utility of the Cys-less CTP for studying structure/function relationships within this metabolically important protein; and (iii) have led to the hypothesis that the polar face of alpha-helical transmembrane domain IV, within which Arg(181), Arg(189), and Cys(192) are located, constitutes an essential portion of the citrate translocation pathway through the membrane. PMID- 10702280 TI - Role of megalin (gp330) in transcytosis of thyroglobulin by thyroid cells. A novel function in the control of thyroid hormone release. AB - When thyroglobulin (Tg) is endocytosed by thyrocytes and transported to lysosomes, thyroid hormones (T4 and T3) are released. However, some internalized Tg is transcytosed intact into the bloodstream, thereby avoiding proteolytic cleavage. Here we show that megalin (gp330), a Tg receptor on thyroid cells, plays a role in Tg transcytosis. Following incubation with exogenous rat Tg at 37 degrees C, Fisher rat thyroid (FRTL-5) cells, a differentiated thyroid cell line, released T3 into the medium. However, when cells were incubated with Tg plus either of two megalin competitors, T3 release was increased, suggesting that Tg internalized by megalin bypassed the lysosomal pathway, possibly with release of undegraded Tg from cells. To assess this possibility, we performed experiments in which FRTL-5 cells were incubated with either unlabeled or (125)I-labeled Tg at 37 degrees C to allow internalization, treated with heparin to remove cell surface-bound Tg, and further incubated at 37 degrees C to allow Tg release. Intact 330-kDa Tg was released into the medium, and the amount released was markedly reduced by megalin competitors. To investigate whether Tg release resulted from transcytosis, we studied FRTL-5 cells cultured as polarized layers with tight junctions on permeable filters in the upper chamber of dual chambered devices. Following the addition of Tg to the upper chamber and incubation at 37 degrees C, intact 330-kDa Tg was found in fluids collected from the lower chamber. The amount recovered was markedly reduced by megalin competitors, indicating that megalin mediates Tg transcytosis. We also studied Tg transcytosis in vivo, using a rat model of goiter induced by aminotriazole, in which increased release of thyrotropin induces massive colloid endocytosis. This was associated with increased megalin expression on thyrocytes and increased serum Tg levels, with reduced serum T3 levels, supporting the conclusion that megalin mediates Tg transcytosis. Tg transcytosis is a novel function of megalin, which usually transports ligands to lysosomes. Megalin-mediated transcytosis may regulate the extent of thyroid hormone release. PMID- 10702281 TI - Uncoupling ceramide glycosylation by transfection of glucosylceramide synthase antisense reverses adriamycin resistance. AB - Previous work from our laboratory demonstrated that increased competence to glycosylate ceramide conferred adriamycin resistance in MCF-7 breast cancer cells (Liu, Y. Y., Han, T. Y., Giuliano, A. E. , and M. C. Cabot. (1999) J. Biol. Chem. 274, 1140-1146). This was achieved by cellular transfection with glucosylceramide synthase (GCS), the enzyme that converts ceramide to glucosylceramide. With this, we hypothesized that a decrease in cellular ceramide glycosylation would result in heightened drug sensitivity and reverse adriamycin resistance. To down regulate ceramide glycosylation potential, we transfected adriamycin-resistant breast cancer cells (MCF-7-AdrR) with GCS antisense (asGCS), using a pcDNA 3.1/his A vector and developed a new cell line, MCF-7-AdrR/asGCS. Reverse transcription-polymerase chain reaction assay and Western blot analysis revealed marked decreases in both GCS mRNA and protein in MCF-7-AdrR/asGCS cells compared with the MCF-7-AdrR parental cells. MCF-7-AdrR/asGCS cells exhibited 30% less GCS activity by in vitro enzyme assay (19.7 +/- 1.1 versus 27.4 +/- 2.3 pmol GC/h/microg protein, p < 0.001) and were 28-fold more sensitive to adriamycin (EC(50), 0.44 +/- 0.01 versus 12.4 +/- 0.7 microM, p < 0. 0001). GCS antisense transfected cells were also 2.4-fold more sensitive to C(6)-ceramide compared with parental cells (EC(50) = 4. 0 +/- 0.03 versus 9.6 +/- 0.5 microM, p < 0.0005). Under adriamycin stress, GCS antisense transfected cells compared with parental cells displayed time- and dose-dependent increases in endogenous ceramide and dramatically higher levels of apoptotic effector, caspase-3. Western blotting showed that adriamycin sensitivity, introduced by asGCS gene transfection, was independent of P-glycoprotein and Bcl-2 expression. In summary, this work shows that transfection of GCS antisense tempers the expression of native GCS and restores cell sensitivity to adriamycin. Therefore, limiting the potential to glycosylate ceramide, which is an apoptotic signal in chemotherapy and radiotherapy, provides a promising approach to combat drug resistance. PMID- 10702282 TI - A role for phospholipase D in GLUT4 glucose transporter translocation. AB - Based on recent studies showing that phospholipase D (PLD)1 is associated with intracellular membranes and promotes membrane budding from the trans-Golgi, we tested its possible role in the membrane trafficking of GLUT4 glucose transporters. Using immunofluorescence confocal microscopy, expressed Myc epitope tagged PLD1 was found to associate with intracellular vesicular structures by a mechanism that requires its N-terminal pleckstrin homology domain. Partial co localization with expressed GLUT4 fused to green fluorescent protein in both 3T3 L1 adipocytes and Chinese hamster ovary cells was evident. Furthermore, microinjection of purified PLD into cultured adipocytes markedly potentiated the effect of a submaximal concentration of insulin to stimulate GLUT4 translocation to cell surface membranes. Insulin stimulated PLD activity in cells expressing high levels of insulin receptors but no such insulin effect was detected in 3T3 L1 adipocytes. Taken together, these results are consistent with the hypothesis that PLD1 associated with GLUT4-containing membranes acts in a constitutive manner to promote the mechanism of GLUT4 translocation by insulin. PMID- 10702283 TI - Inhibition of the hepatitis C virus NS3/4A protease. The crystal structures of two protease-inhibitor complexes. AB - The hepatitis C virus NS3 protein contains a serine protease domain with a chymotrypsin-like fold, which is a target for development of therapeutics. We report the crystal structures of this domain complexed with NS4A cofactor and with two potent, reversible covalent inhibitors spanning the P1-P4 residues. Both inhibitors bind in an extended backbone conformation, forming an anti-parallel beta-sheet with one enzyme beta-strand. The P1 residue contributes most to the binding energy, whereas P2-P4 side chains are partially solvent exposed. The structures do not show notable rearrangements of the active site upon inhibitor binding. These results are significant for the development of antivirals. PMID- 10702284 TI - Truncated estrogen receptor product-1 suppresses estrogen receptor transactivation by dimerization with estrogen receptors alpha and beta. AB - The estrogen receptor (ER) is a ligand-activated transcription factor that acts as a homodimer. Truncated estrogen receptor product-1 (TERP-1) is a pituitary specific, estrogen-induced, isoform of rat ERalpha that is transcribed from a unique start site and contains only the C-terminal region of the full-length receptor. TERP-1 does not affect transcription directly but suppresses ligand activated ERalpha and ERbeta activity. Because TERP-1 contains a dimerization domain and part of the coactivator binding pocket, we hypothesized that it modulates ER function by direct interactions with full-length ER or the steroid receptor coactivator, SRC-1. Localization studies demonstrate that TERP-1 is present in the cytoplasm and nucleus of transfected cells and colocalizes with nuclear ER. Protein binding studies show that TERP-1 forms heterodimers with both ERalpha and ERbeta and inhibits ERalpha binding to its cognate DNA response element. TERP-1 also binds SRC-1, and increasing levels of SRC-1 decrease the TERP-1-ERalpha interactions, in agreement with the rescue of TERP-1-suppressed ERalpha transcriptional activity by SRC-1. Mutational analysis of TERP-1 and ERalpha in the activation helix and the AF-2 dimerization helix indicates that TERP-1 acts predominantly through dimerization with ERalpha. Therefore, TERP-1 suppression of ER transcription occurs primarily by formation of inactive heterodimers and secondarily by competition for coactivators. PMID- 10702285 TI - The trimeric GTP-binding protein (G(q)/G(11)) alpha subunit is required for insulin-stimulated GLUT4 translocation in 3T3L1 adipocytes. AB - To investigate the potential role of trimeric GTP-binding proteins regulating GLUT4 translocation in adipocytes, wild type and constitutively active G(q) (G(q)/Q209L), G(i) (G(i)/Q205L), and G(s) (G(s)/Q227L) alpha subunit mutants were expressed in 3T3L1 adipocytes. Although expression of neither the wild type nor G(i)/Q205L and G(s)/Q227L alpha subunit mutants had any effect on the basal or insulin-stimulated translocation of a co-expressed GLUT4-enhanced green fluorescent protein (EGFP) fusion protein, expression of G(q)/Q209L resulted in GLUT4-EGFP translocation in the absence of insulin. In contrast, microinjection of an inhibitory G(q)/G(11) alpha subunit-specific antibody but not a G(i) or G(s) alpha subunit antibody prevented insulin-stimulated endogenous GLUT4 translocation. Consistent with a required role for GTP-bound G(q)/G(11), expression of the regulators of G protein signaling (RGS4 and RGS16) also attenuated insulin-stimulated GLUT4-EGFP translocation. To assess the relationship between G(q)/G(11) function with the phosphatidylinositol 3-kinase dependent pathway, expression of a dominant-interfering p85 regulatory subunit, as well as wortmannin treatment inhibited insulin-stimulated but not G(q)/Q209L stimulated GLUT4-EGFP translocation. Furthermore, G(q)/Q209L did not induce the in vivo accumulation of phosphatidylinositol-3,4,5-trisphosphate (PIP(3)), whereas expression of the RGS proteins did not prevent the insulin-stimulated accumulation of PIP(3). Together, these data demonstrate that insulin stimulation of GLUT4 translocation requires at least two independent signal transduction pathways, one mediated through the phosphatidylinositol 3-kinase and another through the trimeric GTP-binding proteins G(q) and/or G(11). PMID- 10702286 TI - Vear, a novel Golgi-associated protein with VHS and gamma-adaptin "ear" domains. AB - The molecular basis of the selectivity and the details of the vesicle formation in endocytic and secretory pathways are still poorly known and most probably involve as yet unidentified components. Here we describe the cloning, expression, and tissue and cell distribution of a novel protein of 67 kDa (called Vear) that bears homology to several endocytosis-associated proteins in that it has a VHS domain in its N terminus. It is also similar to gamma-adaptin, the heavy subunit of AP-1, in having in its C terminus a typical "ear" domain. In immunofluorescence microscopy, Vear was seen in the Golgi complex as judged by a typical distribution pattern, a distinct colocalization with the Golgi marker gamma-adaptin, and a sensitivity to treatment of cells with brefeldin A. In cell fractionation, Vear partitioned with the post-nuclear membrane fraction. In transfection experiments, hemagglutinin-tagged full-length Vear and truncated Vear lacking the VHS domain assembled on and caused compaction of the Golgi complex. Golgi association without compaction was seen with the ear domain of Vear, whereas the VHS domain alone showed a diffuse membrane- and vesicle associated distribution. The Golgi association and the bipartite structure along with the differential targeting of its domains suggest that Vear is involved in heterotypic vesicle/suborganelle interactions associated with the Golgi complex. Tissue-specific function of Vear is suggested by its high level of expression in kidney, muscle, and heart. PMID- 10702287 TI - Hepatitis C virus NS5A protein modulates transcription through a novel cellular transcription factor SRCAP. AB - Hepatitis C virus NS5A protein transcriptionally modulates cellular genes and promotes cell growth. NS5A is likely to exert its activity in concert with cellular factor(s). Using a yeast two-hybrid screen, we have demonstrated that NS5A interacts with the C-terminal end of a newly identified cellular transcription factor, SRCAP. The authenticity of this interaction was verified by a mammalian two-hybrid assay, in vitro pull-down experiment, and an in vivo coimmunoprecipitation assay in human hepatoma (HepG2) cells. An in vitro transient transfection assay demonstrated that SRCAP can efficiently activate transcription when recruited by the Gal4 DNA-binding domain to the promoter. However, down-regulation of p21 promoter activity by NS5A was enhanced following ectopic expression of SRCAP. Together these results suggest that the interaction of NS5A and SRCAP may be one of the mechanisms by which NS5A exerts its effect on cell growth regulation contributing to hepatitis C virus-mediated pathogenesis. PMID- 10702288 TI - Myeloid leukemia cell growth and differentiation are independent of mitogen activated protein kinase ERK1/2 activation. AB - The mitogen-activated protein kinase ERK1/2 pathway is essential in the control of cell proliferation and differentiation in most cellular systems. As such, it has been considered a potential target for antineoplastic therapy. For this purpose, we have examined the role of ERK activation in myeloid leukemia cell growth and differentiation. Using a representative set of myeloid leukemia cell lines, we show that cell proliferation was not accompanied by increases on ERK1/2 activation, and mitogenic stimulation did not enhance ERK activity. Moreover, abolition of ERK function by the inhibitor PD98059 or by a dominant inhibitory mutant ERK2 had no significant effects on proliferation. With the aid of various differentiation inducers, we found that within the same cell line, differentiation to a given lineage could occur with and without ERK1/2 activation, depending on the stimulus. Also, a differentiator could have the same effect in the presence or absence of ERK stimulation, depending on the cell line. ERK inhibition did not affect the differentiation elicited by stimuli whose effects were accompanied by ERK activation. Finally, constitutive ERK activity was also ineffective on proliferation and differentiation. Thus, our results indicate that ERK1/2 activation is not an essential requirement for leukemic cell growth and differentiation. PMID- 10702289 TI - Functional domain mapping and subcellular distribution of Dal82p in Saccharomyces cerevisiae. AB - Previous studies have shown that (i) Dal81p and Dal82p are required for allophanate-induced gene expression in Saccharomyces cerevisiae; (ii) the cis acting element mediating the induced transcriptional response to allophanate is a dodecanucleotide, UIS(ALL); and (iii) Dal82p binds specifically to UIS(ALL). Here we show that Dal82p is localized to the nucleus and parallels movement of the DNA through the cell cycle. Deletion analysis of DAL82 identified and localized three functional domains. Electrophoretic mobility shift assays identified a peptide (consisting of Dal82p amino acids 1-85) that is sufficient to bind a DNA fragment containing UIS(ALL). LexA-tethering experiments demonstrated that Dal82p is capable of mediating transcriptional activation. The activation domain consists of two parts: (i) an absolutely required core region (amino acids 66-99) and (ii) less well defined regions flanking residues 66-99 that are required for full wild type levels of activation. The Dal82p C terminus contains a predicted coiled-coil motif that down-regulates Dal82p-mediated transcriptional activation. PMID- 10702290 TI - GTP enhances the degradation of the photosystem II D1 protein irrespective of its conformational heterogeneity at the Q(B) site. AB - The light exposure history and/or binding of different herbicides at the Q(B) site may induce heterogeneity of photosystem II acceptor side conformation that affects D1 protein degradation under photoinhibitory conditions. GTP was recently found to stimulate the D1 protein degradation of photoinactivated photosystem II (Spetea, C. , Hundal, T., Lohmann, F., and Andersson, B. (1999) Proc. Natl. Acad. Sci. U. S. A. 96, 6547-6552). Here we report that GTP enhances the cleavage of the D1 protein D-E loop following exposure of thylakoid membranes to either high light, low light, or repetitive single turnover flashes but not to trypsin. GTP does not stimulate D1 protein degradation in the presence of herbicides known to affect the accessibility of the cleavage site to proteolysis. However, GTP stimulates degradation that can be induced even in darkness in some photosystem II conformers following binding of the PNO8 herbicide (Nakajima, Y., Yoshida, S., Inoue, Y., Yoneyama, K., and Ono, T. (1995) Biochim. Biophys. Acta 1230, 38-44). Both the PNO8- and the light-induced primary cleavage of the D1 protein occur in the grana membrane domains. The subsequent migration of photosytem II containing the D1 protein fragments to the stroma domains for secondary proteolysis is light activated. We conclude that the GTP effect is not confined to a specific photoinactivation pathway nor to the conformational state of the photosystem II acceptor side. Consequently, GTP does not interact with the site of D1 protein cleavage but rather enhances the activity of the endogenous proteolytic system. PMID- 10702291 TI - Cloning and characterization of a novel Kruppel-associated box family transcriptional repressor that interacts with the retinoblastoma gene product, RB. AB - The retinoblastoma gene product, RB, seems to function as a key tumor suppressor by repressing the expression of genes activated by members of the E2F family of transcription factors. In order to accomplish this, RB has been proposed to interact with a transcriptional repressor. However, no genuine transcriptional repressors have been identified by virtue of interaction with RB. By using the yeast two-hybrid system, we have identified a novel member of a known family of transcriptional repressors that contain zinc fingers of the Kruppel type and a portable transcriptional repressor motif known as the Kruppel-associated box (KRAB). The mouse and human forms of the novel RB-associated KRAB protein (RBaK) are widely expressed. The amino acid motif that links the KRAB domain and zinc fingers appears to be required for interaction with RB in vitro. Human RBaK ectopically expressed in fibroblasts is an 80-kDa protein that is localized to the nucleus. The expression of either RB or RBaK in 10T1/2 fibroblasts represses the activation of an E2F-dependent promoter and decreases DNA synthesis to a similar degree. However, a mutant form of RBaK that cannot interact with RB in vitro is unable to prevent DNA synthesis. We present a model in which RB physically interacts with the novel transcriptional repressor RBaK to repress E2F dependent genes and prevent DNA synthesis. PMID- 10702292 TI - Cell wall structure of a mutant of Mycobacterium smegmatis defective in the biosynthesis of mycolic acids. AB - A mutant strain of Mycobacterium smegmatis defective in the biosynthesis of mycolic acids was recently isolated (Liu, J., and Nikaido, H. (1999) Proc. Natl. Acad. Sci. U. S. A. 96, 4011-4016). This mutant failed to synthesize full-length mycolic acids and accumulated a series of long chain beta-hydroxymeromycolates. In this work, we provide a detailed characterization of the localization of meromycolates and of the cell wall structure of the mutant. Thin layer chromatography showed that the insoluble cell wall matrix remaining after extraction with chloroform/methanol and SDS still contained a large portion of the total meromycolates. Matrix-assisted laser desorption/ionization and electrospray ionization mass spectroscopy analysis of fragments arising from Smith degradation of the insoluble cell wall matrix revealed that the meromycolates were covalently attached to arabinogalactan at the 5-OH positions of the terminal arabinofuranosyl residues. The arabinogalactan appeared to be normal in the mutant strain, as analyzed by NMR. Analysis of organic phase lipids showed that the mutant cell wall contained some of the extractable lipids but lacked glycopeptidolipids and lipooligosaccharides. Differential scanning calorimetry of the mutant cell wall failed to show the large cooperative thermal transitions typical of intact mycobacterial cell walls. Transmission electron microscopy showed that the mutant cell wall had an abnormal ultrastructure (without the electron-transparent zone associated with the asymmetric mycolate lipid layer). Taken together, these results demonstrate the importance of mycolic acids for the structural and functional integrity of the mycobacterial cell wall. The lack of highly organized lipid domains in the mutant cell wall explains the drug-sensitive and temperature-sensitive phenotypes of the mutant. PMID- 10702293 TI - Oxalate decarboxylase from Collybia velutipes. Molecular cloning and its overexpression to confer resistance to fungal infection in transgenic tobacco and tomato. AB - Oxalic acid is present as nutritional stress in many crop plants like Amaranth and Lathyrus. Oxalic acid has also been found to be involved in the attacking mechanism of several phytopathogenic fungi. A full-length cDNA for oxalate decarboxylase, an oxalate-catabolizing enzyme, was isolated by using 5'-rapid amplification of cDNA ends-polymerase chain reaction of a partial cDNA as cloned earlier from our laboratory (Mehta, A., and Datta, A. (1991) J. Biol. Chem. 266, 23548-23553). By screening a genomic library from Collybia velutipes with this cDNA as a probe, a genomic clone has been isolated. Sequence analyses and comparison of the genomic sequence with the cDNA sequence revealed that the cDNA is interrupted with 17 small introns. The cDNA has been successfully expressed in cytosol and vacuole of transgenic tobacco and tomato plants. The transgenic plants show normal phenotype, and the transferred trait is stably inherited to the next generation. The recombinant enzyme is partially glycosylated and shows oxalate decarboxylase activity in vitro as well as in vivo. Transgenic tobacco and tomato plants expressing oxalate decarboxylase show remarkable resistance to phytopathogenic fungus Sclerotinia sclerotiorum that utilizes oxalic acid during infestation. The result presented in the paper represents a novel approach to develop transgenic plants resistant to fungal infection. PMID- 10702294 TI - Re-engineering of human urokinase provides a system for structure-based drug design at high resolution and reveals a novel structural subsite. AB - Inhibition of urokinase has been shown to slow tumor growth and metastasis. To utilize structure-based drug design, human urokinase was re-engineered to provide a more optimal crystal form. The redesigned protein consists of residues Ile(16) Lys(243) (in the chymotrypsin numbering system; for the urokinase numbering system it is Ile(159)-Lys(404)) and two point mutations, C122A and N145Q (C279A and N302Q). The protein yields crystals that diffract to ultra-high resolution at a synchrotron source. The native structure has been refined to 1.5 A resolution. This new crystal form contains an accessible active site that facilitates compound soaking, which was used to determine the co-crystal structures of urokinase in complex with the small molecule inhibitors amiloride, 4-iodo benzo(b)thiophene-2-carboxamidine and phenylguanidine at 2. 0-2.2 A resolution. All three inhibitors bind at the primary binding pocket of urokinase. The structures of amiloride and 4-iodo-benzo(b)thiophene-2-carboxamidine also reveal that each of their halogen atoms are bound at a novel structural subsite adjacent to the primary binding pocket. This site consists of residues Gly(218), Ser(146), and Cys(191)-Cys(220) and the side chain of Lys(143). This pocket could be utilized in future drug design efforts. Crystal structures of these three inhibitors in complex with urokinase reveal strategies for the design of more potent nonpeptidic urokinase inhibitors. PMID- 10702295 TI - Probing chemical and conformational differences in the resting and active conformers of platelet integrin alpha(IIb)beta(3). AB - Integrin alpha(IIb)beta(3) is the fibrinogen receptor that mediates platelet adhesion and aggregation. The ligand binding function of alpha(IIb)beta(3) is "activated" on the platelet surface by physiologic stimuli. Two forms of alpha(IIb)beta(3) can be purified from platelet lysates. These forms are facsimiles of the resting (Activation State-1 or AS-1) and the active (Activation State-2 or AS-2) conformations of the integrin found on the platelet surface. Here, the differences between purified AS-1 and AS-2 were examined to gain insight into the mechanism of activation. Four major findings are put forth. 1) The association rate (k(1)) between fibrinogen and the integrin is a key difference between AS-1 and AS-2. 2) Although the divalent ion Mn(2+) enhances the ligand binding function of AS-1, this ion is unable to convert AS-1 to AS-2. Therefore, its effect on integrin is unrelated to activation. 3) Peptide mass fingerprints indicate that the chemical structure of AS-1 and AS-2 are virtually identical, calling into question the idea that post-translational modifications are necessary for activation. 4) The two forms of alpha(IIb)beta(3) have significant conformational differences at three positions. These include the junction of the heavy and light chain of alpha(IIb), the divalent ion binding sites on alpha(IIb), and at a disulfide-bonded knot linking the amino terminus of beta(3) to the cysteine-rich domain. These observations indicate that integrin is activated by a series of specific conformational rearrangements in the ectodomain that increase the rate of ligand association. PMID- 10702296 TI - Human transaldolase-associated repetitive elements are transcribed by RNA polymerase III. AB - Repetitive elements flanked by exons 2 and 3 of the human transaldolase gene, thus termed transaldolase-associated repetitive elements, TARE, were identified in human DNA. Nonpolyadenylated TARE transcripts were detected by Northern blot analysis and cloned by reverse transcriptase-mediated polymerase chain reaction from human T lymphocytes. A dominant 1085-nucleotide long transcript, TARE-6, contained two adjacent Alu elements, a right monomer and a complete dimer, oriented opposite to the direction of transcription of the transaldolase gene. Reverse transcriptase-polymerase chain reaction and in vitro transcription analyses showed that transcription of TARE-6 proceeded in the orientation of the RNA pol III promoter of the Alu dimer and opposite to the orientation of the TAL H gene. TAREs lacking RNA polymerase III promoter showed no transcriptional activity. In vitro transcription of TARE-6 was resistant to 1 microg/ml alpha amanitin but sensitive to 100 microg/ml alpha-amanitin and tagetitoxin, suggesting involvement of RNA polymerase III. TAREs in both the transaldolase and HSAG-1 genomic loci were surrounded by TA target site duplications. Homologies between transaldolase and HSAG-1 break off internally at splice donor and acceptor sites. The results suggest RNA polymerase III-mediated transcription of TARE may be a source of repetitive elements, contributing to distinct genes and thus shaping the human genome. PMID- 10702297 TI - Identification of structural and functional domains in mixed lineage kinase dual leucine zipper-bearing kinase required for complex formation and stress-activated protein kinase activation. AB - Accumulating evidence suggests that mitogen-activated protein kinase signaling pathways form modular signaling complexes. Because the mixed lineage kinase dual leucine zipper-bearing kinase (DLK) is a large modular protein, structure function analysis was undertaken to examine the role of DLK domains in macromolecular complex formation. DLK mutants were used to demonstrate that a DLK leucine zipper-leucine zipper interaction is necessary for DLK dimerization and to show that DLK dimerization mediated by the leucine zipper domain is prerequisite for DLK activity and subsequent activation of stress-activated protein kinase (SAPK). Heterologous mixed lineage kinase family members can be co immunoprecipitated. However, the DLK leucine zipper domain interacted specifically only with the DLK leucine zipper domain; in contrast, DLK NH(2) terminal region was sufficient to co-immunoprecipitate leucine zipper kinase and DLK. DLK has been shown to associate with the putative scaffold protein JIP1. This association occurred through the DLK NH(2)-terminal region and occurred independently of DLK catalytic activity. Although the DLK NH(2)-terminal region associated directly with JIP-1, this region did not interact directly with either DLK or leucine zipper kinase. Therefore, DLK may interact with heterologous mixed lineage kinase proteins via intermediary proteins. The NH(2)-terminal region of overexpressed DLK was required for activation of SAPK. These results provide evidence that protein complex formation is required for signal transduction from DLK to SAPK. PMID- 10702298 TI - Cloning and characterization of the promoter region of the rat epidermal growth factor receptor gene and its transcriptional regulation by nerve growth factor in PC12 cells. AB - Our previous studies have shown that treatment of PC12 cells with nerve growth factor (NGF) causes a profound down-regulation of the epidermal growth factor receptor (EGFR) mRNA and protein. Further, the NGF-induced down-regulation of the EGFR is under transcriptional control. To elucidate the molecular mechanism of this down-regulation we have cloned a 2.7-kilobase sequence from the promoter region of the rat EGFR from a rat P1 library. Six transcriptional start sites were identified by 5'-rapid amplification of cDNA ends and primer extension. Sequence analysis showed a 62% overall homology with the human EGFR promoter region. To investigate its transcription, 1.1 kilobases of the 5'-flanking sequence were fused to a luciferase reporter gene. This sequence exhibited functional promoter activity in transient transfection experiments with PC12, C6, and CV-1 cells. Treatment of PC12 cells with NGF inhibited promoter activity. By transfection of promoter deletion constructs, a silencer element was found between nucleotides -260 and -181, and TCC repeat sequences appeared to be at least partially responsible for the down-regulation of the EGFR by NGF. Supportive evidence for the relevance of this sequence was obtained from gel mobility shift assays and by transfection of TCC mutation constructs. Our results demonstrate that TCC repeat sequences are required for the down-regulation of rat EGFR by NGF in PC12 cells and may lead to the identification of the NGF responsive transcription factors. PMID- 10702299 TI - Insulin inhibits the activation of transcription by a C-terminal fragment of the forkhead transcription factor FKHR. A mechanism for insulin inhibition of insulin like growth factor-binding protein-1 transcription. AB - The forkhead rhabdomyosarcoma transcription factor (FKHR) is a promising candidate to be the transcription factor that binds to the insulin response element of the insulin-like growth factor-binding protein-1 (IGFBP-1) promoter and mediates insulin inhibition of IGFBP-1 promoter activity. Cotransfection of mouse FKHR increased IGFBP-1 promoter activity 2-3-fold in H4IIE rat hepatoma cells; insulin inhibited FKHR-stimulated promoter activity approximately 70%. A C terminal fragment of mouse FKHR (residues 208-652) that contains the transcription activation domain fused to a Gal4 DNA binding domain potently stimulated Gal4 promoter activity. Insulin inhibited FKHR fragment-stimulated promoter activity by approximately 70%. Inhibition was abolished by coincubation with the phosphatidylinositol-3 kinase inhibitor, LY294002. The FKHR 208-652 fragment contains two consensus sites for phosphorylation by protein kinase B (PKB)/Akt, Ser-253 and Ser-316. Neither site is required for insulin inhibition of promoter activity stimulated by the FKHR fragment, and overexpression of Akt does not inhibit FKHR fragment-stimulated Gal4 promoter activity. These results suggest that insulin- and phosphatidylinositol-3 kinase-dependent phosphorylation of another site in the fragment by a kinase different from PKB/Akt inhibits transcription activation by the fragment. Phosphorylation of this site also may be involved in insulin inhibition of transcription activation by full-length FKHR, but only after phosphorylation of Ser-253 by PKB/Akt. PMID- 10702300 TI - Isolation of a somatic cell mutant resistant to the induction of apoptosis by oxidized low density lipoprotein. AB - Oxidized low density lipoprotein (oxLDL) induces apoptosis in macrophages, smooth muscle cells, and endothelial cells. To elucidate the molecular mechanism of oxLDL-induced cytotoxicity and determine its tissue specificity, we have used Chinese hamster ovary (CHO)-K1 cells expressing human CD36 (CHO/CD36). Expression of CD36 rendered these cells susceptible to killing by oxLDL. This cytotoxicity was due to the induction of apoptosis. Therefore, CD36 expression is the only requirement for oxLDL-induced apoptosis. Oxysterols apparently mediate the cytotoxicity of oxLDL in macrophage foam cells and endothelial cells. 25 Hydroxycholesterol, at concentrations higher than 1 microg/ml, killed CHO-K1 cells, by apoptosis, in medium supplemented with serum as a source of cholesterol. These effects were not seen in a 25-hydroxycholesterol-resistant CHO/CD36 mutant (OX(R)), which was otherwise capable of undergoing apoptosis in response to staurosporine. This mutant was also resistant to killing by oxLDL, suggesting that oxysterols are at least partially responsible for the toxic effects of oxLDL. Oxysterol-induced apoptosis did not involve regulation of sterol regulatory element-binding protein proteolysis or the cholesterol biosynthetic pathway. 25-Hydroxycholesterol stimulated calcium uptake by CHO-K1 cells within 2 min after addition. Treatment of CHO or THP-1 (macrophage) cells with the calcium channel blocker nifedipine prevented 25-hydroxycholesterol induction of apoptosis. OX(R) showed no enhanced calcium uptake in response to 25 hydroxycholesterol. PMID- 10702301 TI - Comparative expression of homologous proteins. A novel mode of transcriptional regulation by the coding sequence folding compatibility of chimeras. AB - Recombinant acetylcholinesterases (AChE) are produced at systematically different levels, depending on the enzyme species. To identify the cause of this difference, we designed expression vectors that differed only by the central region of the coding sequence, encoding Torpedo, rat, and Bungarus AChEs and two reciprocal rat/Bungarus and Bungarus/rat chimeras. We found that folding is a limiting factor in the case of Torpedo AChE and the chimeras, for which only a limited fraction of the synthesized polypeptides becomes active and is secreted. In contrast, the fact that rat AChE is less well produced than Bungarus AChE reflects the levels of their respective mRNAs, which seem to be controlled by their transcription rates. A similar difference was observed in the coding and noncoding orientations; it seems to depend on multiple cis-elements. Using CAT constructs, we found that a DNA fragment from the Bungarus AChE gene stimulates expression of the reporter protein, whereas a homologous fragment from the rat AChE gene had no influence. This stimulating effect appears different from that of classical enhancers, although its mechanism remains unknown. In any case, the present results demonstrate that the coding region contributes to control the level of gene expression. PMID- 10702302 TI - The N-terminal region of the human progesterone A-receptor. Structural analysis and the influence of the DNA binding domain. AB - The role of the N-terminal region in nuclear receptor function was addressed by a biochemical and biophysical analysis of the progesterone receptor A-isoform lacking only the hormone binding domain (NT-A). Sedimentation studies demonstrate that NT-A is quantitatively monomeric, with a highly asymmetric shape. Contrary to dogma, the N-terminal region is structured as demonstrated by limited proteolysis. However, N-terminal structure is strongly stabilized by the DNA binding domain, possibly explaining the lack of structure seen in isolated activation domains. Upon DNA binding, NT-A undergoes N-terminal mediated assembly, suggestive of DNA-induced allostery, and consistent with changes in protease accessibility of sites outside the DNA binding domain. Microsequencing reveals that protease-accessible regions are limited to previously identified phosphorylation motifs and to functional domain boundaries. PMID- 10702303 TI - Functions of amino acid residues in the active site of Escherichia coli pyrroloquinoline quinone-containing quinoprotein glucose dehydrogenase. AB - Several mutants of quinoprotein glucose dehydrogenase (GDH) in Escherichia coli, located around its cofactor pyrroloquinoline quinone (PQQ), were constructed by site-specific mutagenesis and characterized by enzymatic and kinetic analyses. Of these, critical mutants were further characterized after purification or by different amino acid substitutions. H262A mutant showed reduced affinities both for glucose and PQQ without significant effect on glucose oxidase activity, indicating that His-262 occurs very close to PQQ and glucose, but is not the electron acceptor from PQQH(2). W404A and W404F showed pronounced reductions of affinity for PQQ, and the latter rather than the former had equivalent glucose oxidase activity to the wild type, suggesting that Trp-404 may be a support for PQQ and important for the positioning of PQQ. D466N, D466E, and K493A showed very low glucose oxidase activities without influence on the affinity for PQQ. Judging from the enzyme activities of D466E and K493A, as well as their absorption spectra of PQQ during glucose oxidation, we conclude that Asp-466 initiates glucose oxidation reaction by abstraction of a proton from glucose and Lys-493 is involved in electron transfer from PQQH(2). PMID- 10702304 TI - A unique organization of the protein subunits of the DNA polymerase clamp loader in the archaeon Methanobacterium thermoautotrophicum deltaH. AB - Replication factor C (RFC, also called activator 1), in conjunction with proliferating cell nuclear antigen (PCNA), is responsible for processive DNA synthesis catalyzed by the eukaryotic replicative DNA polymerases delta and epsilon. Here we report the isolation and characterization of homologues of RFC and PCNA from the archaeon, Methanobacterium thermoautotrophicum DeltaH. In contrast to the five subunit RFC complex isolated from eukaryotic cells, the mthRFC contains only two subunits. The two genes encoding the RFC subunits called, mthRFC1 and mthRFC3, were cloned, and the proteins (54.4 and 36.8 kDa, respectively) were overexpressed in Escherichia coli and purified individually and as a complex. The gene encoding PCNA was also cloned, and the protein was purified after overexpression in E. coli. Based on sizing column elution and subunit composition, the mthRFC complex appears to be a hexamer consisting of two mthRFC1 protomers and four mthRFC3 protomers. Although mthRFC differs in organization from its eukaryotic counterpart, it was shown to be functionally similar to eukaryotic RFC in: (i) catalyzing DNA-dependent ATP hydrolysis; (ii) binding preferentially to DNA primer ends; (iii) loading mthPCNA onto singly nicked circular DNA; and (iv) supporting mthPolB-catalyzed PCNA-dependent DNA chain elongation. The importance and roles of RFC and PCNA in M. thermoautotrophicum DeltaH replication are discussed. PMID- 10702305 TI - p53 induces apoptosis by caspase activation through mitochondrial cytochrome c release. AB - The p53 tumor suppressor gene is critically involved in cell cycle regulation, DNA repair, and programmed cell death. Several lines of evidence suggest that p53 death signals lead to caspase activation; however, the mechanism of caspase activation by p53 still is unclear. Expressing wild type p53 by means of an adenoviral expression vector, we were able to induce apoptotic cell death, as characterized by morphological changes, phosphatidylserine externalization, and internucleosomal DNA fragmentation, in p53(null) Saos-2 cells. This cell death was accompanied by caspase activation as well as by cleavage of caspase substrates and was preceded by mitochondrial cytochrome c release. The addition of the broad-spectrum caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp fluoromethyl ketone (zVAD-fmk) directly after transduction almost completely prevented p53-induced apoptotic cell death but did not inhibit mitochondrial cytochrome c release. In contrast, N-acetylcysteine, even at high concentrations, could not prevent induction of programmed cell death by p53 expression. Cytosolic extracts from Saos-2 cells transduced with p53, but not from Saos-2 cells transduced with the empty adenoviral vector, contained a cytochrome c-releasing activity in vitro, which was still active in the presence of zVAD-fmk. When Bax was immunodepleted from the cytosolic extracts of p53-expressing cells before incubation with isolated mitochondria, the in vitro cytochrome c release was abolished. Thus, we could demonstrate in cells and in vitro that p53 activates the apoptotic machinery through induction of the release of cytochrome c from the mitochondrial intermembrane space. Furthermore, we provide in vitro evidence for the requirement of cytosolic Bax for this cytochrome c-releasing activity of p53 in Saos-2 cells. PMID- 10702306 TI - Regulation of c-myc transcription by interleukin-2 (IL-2). Identification of a novel IL-2 response element interacting with STAT-4. AB - Regulation of c-myc expression is known to occur at the level of transcription initiation. However, the participating promoter elements and their cognate binding proteins have not been fully characterized. c-myc transcription can be stimulated by a number of cytokines including interleukin-2 (IL-2). We have identified a novel IL-2-responsive element, located in the 5'-flanking region of the c-myc gene, between nucleotides -1406 and -1387 (relative to the P2 promoter). This element belongs to the family of interferon-gamma activation site like responsive elements and has the core sequence TTCCAATAA. We confirmed that IL-2-mediated signaling involves activation by phosphorylation of Jak2 tyrosine kinase and subsequently STAT4. The transcription factor STAT4 binds the TTCCAATAA motif within this responsive element and, therefore, is probably involved in enhancing c-myc transcription upon IL-2 stimulation. Our results propose participation of Jak2 and STAT4 in IL-2-induced up-regulation of c-myc. PMID- 10702307 TI - Epitope randomization redefines the functional role of glutamic acid 110 in interleukin-5 receptor activation. AB - Sequence randomization through functional phage display of single chain human interleukin (IL)-5 was used to investigate the limits of replaceability of the Glu(110) residues that form a part of the receptor-binding epitope. Mutational analysis revealed unexpected affinity for IL-5 receptor alpha chain with variants containing E110W or E110Y. Escherichia coli-expressed Glu(110) variants containing E110W in the otherwise sequence-intact N-terminal half, including a variant with an E110A replacement in the sequence-disabled C-terminal half, were shown by their CD spectra to be folded into secondary structures similar to that of single chain human IL-5 (scIL-5). Biosensor kinetics analysis revealed that (E110W/A5)scIL-5 and (E110W/A6)scIL-5 had receptor alpha chain binding affinities similar to that of (wt/A5)scIL-5. However, (E110W/A6)scIL-5 had a significantly reduced bioactivity in TF-1 cell proliferation compared with both (wt/A5)scIL-5 and (E110W/A5)scIL-5, and this activity reduction was disproportionately greater than the much smaller effect of Glu(110) mutation on receptor binding affinity. The marked and disproportionate decrease in TF-1 proliferation observed with (E110W/A6)scIL-5 suggests a role for Glu(110) in the biological activity mediated by the signal transducing receptor betac subunit of the IL-5 receptor. This is also consistent with the lack of stimulation of JAK2 phosphorylation by the (E110W/A6)scIL-5 mutant in recombinant 293T cells, as compared with the concentration-dependent stimulation seen for scIL-5. The results reveal the dispensability of charge in the Glu(110) locus of IL-5 for receptor alpha chain binding and, in contrast, its heretofore underappreciated importance for receptor activation. PMID- 10702308 TI - TAK1 mitogen-activated protein kinase kinase kinase is activated by autophosphorylation within its activation loop. AB - TAK1, a member of the mitogen-activated kinase kinase kinase family, is activated in vivo by various cytokines, including interleukin-1 (IL-1), or when ectopically expressed together with the TAK1-binding protein TAB1. However, this molecular mechanism of activation is not yet understood. We show here that endogenous TAK1 is constitutively associated with TAB1 and phosphorylated following IL-1 stimulation. Furthermore, TAK1 is constitutively phosphorylated when ectopically overexpressed with TAB1. In both cases, dephosphorylation of TAK1 renders it inactive, but it can be reactivated by preincubation with ATP. A mutant of TAK1 that lacks kinase activity is not phosphorylated either following IL-1 treatment or when coexpressed with TAB1, indicating that TAK1 phosphorylation is due to autophosphorylation. Furthermore, mutation to alanine of a conserved serine residue (Ser-192) in the activation loop between kinase domains VII and VIII abolishes both phosphorylation and activation of TAK1. These results suggest that IL-1 and ectopic expression of TAB1 both activate TAK1 via autophosphorylation of Ser-192. PMID- 10702309 TI - Natriuretic peptides inhibit G protein activation. Mediation through cross-talk between cyclic GMP-dependent protein kinase and regulators of G protein-signaling proteins. AB - Atrial natriuretic peptide (ANP) inhibits the proliferation of many cells, in part through interfering with signal transduction enacted by G protein-coupled growth factor receptors. Signaling interactions between ANP and the G protein coupled growth factor receptor ligand, endothelin-3 (ET-3), regulate astrocyte proliferation at a very proximal but undefined point. Here, we find that ANP inhibits the ability of ET-3 to activate Galpha(q) and Galpha(i) in these cells. ANP stimulated the translocation of endogenous regulators of G protein-signaling (RGS) proteins 3 and 4 from the cytosol to the cell membrane, and enhanced their association with Galpha(q) and Galpha(i). ANP effects were significantly blocked by HS-142-1, an inhibitor of guanylate cyclase activation, or by ET-3. KT5823, an inhibitor of cyclic GMP-dependent protein kinase (PKG) reversed the RGS translocation induced by ANP; conversely, expression of an active catalytic subunit of PKG-I, or 8-bromo-cyclic GMP stimulated RGS translocation. ANP caused the phosphorylation of both RGS proteins in a PKG-dependent fashion, and the expressed PKG (in the absence of ANP) also stimulated RGS phosphorylation. A novel cross-talk between PKG and RGS proteins is stimulated by ANP and leads to the increased translocation and association of RGS proteins with Galpha. The rapid inactivation of G proteins provides a mechanism by which ANP inhibits downstream signaling to the cell proliferation program. PMID- 10702310 TI - Characterization of a novel serine/threonine kinase associated with nuclear bodies. AB - A novel protein kinase, Mx-interacting protein kinase (PKM), has been identified in a yeast two-hybrid screen for interaction partners of human MxA, an interferon induced GTPase with antiviral activity against several RNA viruses. A highly conserved protein kinase domain is present in the N-terminal moiety of PKM, whereas an Mx interaction domain overlaps with C-terminal PEST sequences. PKM has a molecular weight of about 127,000 and exhibits high sequence homology to members of a recently described family of homeodomain-interacting protein kinases. Recombinant PKM has serine/threonine kinase activity that is abolished by a single amino acid substitution in the ATP binding domain (K221W). PKM catalyzes autophosphorylation and phosphorylation of various cellular and viral proteins. PKM is expressed constitutively and colocalizes with the interferon inducible Sp100 protein and murine Mx1 in discrete nuclear structures known as nuclear bodies. PMID- 10702311 TI - Cell surface display and intracellular trafficking of free glycosylphosphatidylinositols in mammalian cells. AB - In addition to serving as membrane anchors for cell surface proteins, glycosylphosphatidylinositols (GPIs) can be found abundantly as free glycolipids in mammalian cells. In this study we analyze the subcellular distribution and intracellular transport of metabolically radiolabeled GPIs in three different cell lines. We use a variety of membrane isolation techniques (subcellular fractionation, plasma membrane vesiculation to isolate pure plasma membrane fractions, and enveloped viruses to sample cellular membranes) to provide direct evidence that free GPIs are not confined to their site of synthesis, the endoplasmic reticulum, but can redistribute to populate other subcellular organelles. Over short labeling periods (2.5 h), radiolabeled GPIs were found at similar concentration in all subcellular fractions with the exception of a mitochondria-enriched fraction where GPI concentration was low. Pulse-chase experiments over extended chase periods showed that although the total amount of cellular radiolabeled GPIs decreased, the plasma membrane complement of labeled GPIs increased. GPIs at the plasma membrane were found to populate primarily the exoplasmic leaflet as detected using periodate oxidation of the cell surface. Transport of GPIs to the cell surface was inhibited by Brefeldin A and blocked at 15 degrees C, suggesting that GPIs are transported to the plasma membrane via a vesicular mechanism. The rate of transport of radiolabeled GPIs to the cell surface was found to be comparable with the rate of secretion of newly synthesized soluble proteins destined for the extracellular space. PMID- 10702312 TI - Molecular and biochemical characterization of rat gamma trimethylaminobutyraldehyde dehydrogenase and evidence for the involvement of human aldehyde dehydrogenase 9 in carnitine biosynthesis. AB - The penultimate step in carnitine biosynthesis is mediated by gamma trimethylaminobutyraldehyde dehydrogenase (EC 1.2.1.47), a cytosolic NAD(+) dependent aldehyde dehydrogenase that converts gamma-trimethylaminobutyraldehyde into gamma-butyrobetaine. This enzyme was purified from rat liver, and two internal peptide fragments were sequenced by Edman degradation. The peptide sequences were used to search the Expressed Sequence Tag data base, which led to the identification of a rat cDNA containing an open reading frame of 1485 base pairs encoding a polypeptide of 494 amino acids with a calculated molecular mass of 55 kDa. Expression of the coding sequence in Escherichia coli confirmed that the cDNA encodes gamma-trimethylaminobutyraldehyde dehydrogenase. The previously identified human aldehyde dehydrogenase 9 (EC 1.2.1.19) has 92% identity with rat trimethylaminobutyraldehyde dehydrogenase and has been reported to convert substrates that resemble gamma-trimethylaminobutyraldehyde. When aldehyde dehydrogenase 9 was expressed in E. coli, it exhibited high trimethylaminobutyraldehyde dehydrogenase activity. Furthermore, comparison of the enzymatic characteristics of the heterologously expressed human and rat dehydrogenases with those of purified rat liver trimethylaminobutyraldehyde dehydrogenase revealed that the three enzymes have highly similar substrate specificities. In addition, the highest V(max)/K(m) values were obtained with gamma-trimethylaminobutyraldehyde as substrate. This indicates that human aldehyde dehydrogenase 9 is the gamma-trimethylaminobutyraldehyde dehydrogenase, which functions in carnitine biosynthesis. PMID- 10702313 TI - The pyridinyl imidazole inhibitor SB203580 blocks phosphoinositide-dependent protein kinase activity, protein kinase B phosphorylation, and retinoblastoma hyperphosphorylation in interleukin-2-stimulated T cells independently of p38 mitogen-activated protein kinase. AB - Pyridinyl imidazole inhibitors, particularly SB203580, have been widely used to elucidate the roles of p38 mitogen-activated protein (MAP) kinase (p38/HOG/SAPKII) in a wide array of biological systems. Studies by this group and others have shown that SB203580 can have antiproliferative activity on cytokine activated lymphocytes. However, we recently reported that the antiproliferative effects of SB203580 were unrelated to p38 MAP kinase activity. This present study now shows that SB203580 can inhibit the key cell cycle event of retinoblastoma protein phosphorylation in interleukin-2-stimulated T cells. Studies on the proximal regulator of this event, the phosphatidylinositol 3-kinase/protein kinase B (PKB)(Akt/Rac) kinase pathway, showed that SB203580 blocked the phosphorylation and activation of PKB by inhibiting the PKB kinase, phosphoinositide-dependent protein kinase 1. The concentrations of SB203580 required to block PKB phosphorylation (IC(50) 3-5 microM) are only approximately 10-fold higher than those required to inhibit p38 MAP kinase (IC(50) 0.3-0.5 microM). These data define a new activity for this drug and would suggest that extreme caution should be taken when interpreting data where SB203580 has been used at concentrations above 1-2 microM. PMID- 10702314 TI - The differential time-course of extracellular-regulated kinase activity correlates with the macrophage response toward proliferation or activation. AB - Bone marrow-derived macrophages proliferate in response to specific growth factors, including macrophage colony-stimulating factor (M-CSF). When stimulated with activating factors, such as lipopolysaccharide (LPS), macrophages stop proliferating and produce proinflammatory cytokines. Although triggering opposed responses, both M-CSF and LPS induce the activation of extracellular-regulated kinases (ERKs) 1 and 2. However, the time-course of ERK activation is different; maximal activation by M-CSF and LPS occurred after 5 and 15 min of stimulation, respectively. Granulocyte/macrophage colony-stimulating factor, interleukin 3, and TPA, all of which induced macrophage proliferation, also induced ERK activity, which was maximal at 5 min poststimulation. The use of PD98059, which specifically blocks ERK 1 and 2 activation, demonstrated that ERK activity was necessary for macrophage proliferation in response to these factors. The treatment with phosphatidylcholine-specific phospholipase C (PC-PLC) inhibited macrophage proliferation, induced the expression of cytokines, and triggered a pattern of ERK activation equivalent to that induced by LPS. Moreover, PD98059 inhibited the expression of cytokines induced by LPS or PC-PLC, thus suggesting that ERK activity is also required for macrophage activation by these two agents. Activation of the JNK pathway did not discriminate between proliferative and activating stimuli. In conclusion, our results allow to correlate the differences in the time-course of ERK activity with the macrophagic response toward proliferation or activation. PMID- 10702315 TI - Modulation of beta-amyloid precursor protein processing by the low density lipoprotein receptor-related protein (LRP). Evidence that LRP contributes to the pathogenesis of Alzheimer's disease. AB - Beta-amyloid peptide (Abeta), which plays a central role in the pathogenesis of Alzheimer's disease, is derived from the transmembrane beta-amyloid precursor protein (APP) by proteolytic processing. Although mechanisms associated with Abeta generation are not fully understood, it is known that Abeta can be generated within endosomal compartments upon internalization of APP from the cell surface. The low density lipoprotein receptor-related protein (LRP) was previously shown to mediate the endocytosis of APP isoforms containing the Kunitz proteinase inhibitor domain (Kounnas, M. Z., Moir, R. D., Rebeck, G. W., Bush, A. I., Argraves, W. S., Tanzi, R. E., Hyman, B. T., and Strickland, D. K. (1995) Cell 82, 331-340; Knauer, M. F., Orlando, R. A., and Glabe, C. G. (1996) Brain Res. 740, 6-14). The objective of the current study was to test the hypothesis that LRP-mediated internalization of cell surface APP can modulate APP processing and thereby affect Abeta generation. Here, we show that long term culturing of cells in the presence of the LRP-antagonist RAP leads to increased cell surface levels of APP and a significant reduction in Abeta synthesis. Further, restoring LRP function in LRP-deficient cells results in a substantial increase in Abeta production. These findings demonstrate that LRP contributes to Abeta generation and suggest novel pharmacological approaches to reduce Abeta levels based on selective LRP blockade. PMID- 10702316 TI - FKBP12-rapamycin-associated protein (FRAP) autophosphorylates at serine 2481 under translationally repressive conditions. AB - The FKBP12-rapamycin associated protein (FRAP, also RAFT, mTOR) belongs to a family of phosphatidylinositol kinase-related kinases. These kinases mediate cellular responses to stresses such as DNA damage and nutrient deprivation in a variety of eukaryotes from yeast to humans. FRAP regulates G(1) cell cycle progression and translation initiation in part by controlling the phosphorylation states of a number of translational and cell cycle regulators. Although FRAP is known to be phosphorylated in vivo and to phosphorylate several proteins (including itself) in vitro, FRAP's phosphorylation sites and substrate specificity are unknown. We report here the identification of a FRAP autophosphorylation site. This site, Ser-2481, is located in a hydrophobic region near the conserved carboxyl-terminal FRAP tail. We demonstrate that the COOH terminal tail is required for FRAP kinase activity and for signaling to the translational regulator p70(s6k) (ribosomal subunit S6 kinase). Phosphorylation of wild-type but not kinase-inactive FRAP occurs at Ser-2481 in vivo, suggesting that Ser-2481 phosphorylation is a marker of FRAP autokinase activity in cells. FRAP autophosphorylation is blocked completely by wortmannin treatment but not by rapamycin treatment, amino acid deprivation, or serum withdrawal, treatments that lead to acute dephosphorylation of eIF4E-binding protein (4E-BP1) and p70(s6k). Ser-2481 phosphorylation increases slightly upon c-Akt/PKB activation and dramatically upon calyculin A treatment of T-cells. These results suggest that FRAP-responsive dephosphorylation of 4E-BP1 and p70(s6k) occurs through a mechanism other than inhibition of intrinsic FRAP kinase activity. PMID- 10702317 TI - Identification of AUF1 as a parathyroid hormone mRNA 3'-untranslated region binding protein that determines parathyroid hormone mRNA stability. AB - Parathyroid hormone (PTH) mRNA levels are post-transcriptionally increased by hypocalcemia and decreased by hypophosphatemia, and this is mediated by cytosolic proteins binding to the PTH mRNA 3'-untranslated region (UTR). The same proteins are also present in other tissues, such as brain, but only in the parathyroid is their binding regulated by calcium and phosphate. The function of the PTH mRNA 3' UTR-binding proteins was studied using an in vitro degradation assay. Competition for the parathyroid-binding proteins by excess unlabeled 3'-UTR destabilized the full-length PTH transcript in this assay, indicating that these proteins protect the RNA from RNase activity. The PTH RNA 3'-UTR-binding proteins were purified by RNA affinity chromatography of rat brain S-100 extracts. The eluate from the column was enriched in PTH RNA 3'-UTR binding activity. Addition of eluate to the in vitro degradation assay with parathyroid protein extracts stabilized the PTH transcript. A major band from the eluate at 50 kDa was sequenced and was identical to AU-rich binding protein (AUF1). Recombinant AUF1 bound the full length PTH mRNA and the 3'-UTR. Added recombinant AUF1 also stabilized the PTH transcript in the in vitro degradation assay. Our results show that AUF1 is a protein that binds to the PTH mRNA 3'-UTR and stabilizes the PTH transcript. PMID- 10702318 TI - A conserved mechanism for controlling the translation of beta-F1-ATPase mRNA between the fetal liver and cancer cells. AB - To characterize the mechanisms governing the biogenesis of mitochondria in cancer, we studied the mitochondrial phenotype and the mechanisms controlling the expression of the beta subunit of the mitochondrial H(+)-ATP synthase (beta-F1 ATPase) gene in the rat FAO and AS30D hepatomas. When compared with normal adult rat liver, the relative cellular content of the mitochondrial beta-F1-ATPase and glutamate dehydrogenase, as well as of mitochondrial DNA, was severely reduced in both cell lines. A paradoxical increase in the cellular abundance of beta-F1 ATPase mRNA was observed in cancer cells. Run-on transcription assays and the estimation of mRNA half-lives revealed that the increased abundance of beta-F1 ATPase mRNA results from the stabilization of the transcript in cancer. In vitro translation assays revealed a specific inhibition of the synthesis of the beta precursor when translation reactions were carried out in the presence of extracts derived from cancer cells. The inhibitory effect was recapitulated using an RNA chimera that contained the 3'-untranslated region of beta-F1-ATPase mRNA. Hepatoma extracts also contained an increased activity of the developmentally regulated translation-inhibitory proteins that bind the 3'-untranslated region of beta-F1-ATPase mRNA. The results indicate that the expression of this gene in hepatoma cells is controlled by the same mechanisms that regulate its expression in the liver during fetal development. PMID- 10702319 TI - Tonsils and adenoids. PMID- 10702321 TI - Earning CME credit-completing the PIR quiz PMID- 10702320 TI - Aortic and pulmonary stenosis. PMID- 10702322 TI - Pediatric foreign body aspiration. PMID- 10702323 TI - Index of suspicion. Case 1. Ataxia. PMID- 10702324 TI - Idiopathic thrombocytopenic purpura. PMID- 10702325 TI - Visual diagnosis: An infant who has fever and rash. Diagnosis: measles. PMID- 10702326 TI - Fluoride in drinking water and cancer mortality in Taiwan. AB - The possibility that cancer risk is associated with naturally fluoridated water in Taiwan is examined. The 1982-1991 age-adjusted mortality rates for cancer for 10 municipalities whose water supplies contained the highest naturally occurring fluoride concentrations in Taiwan were compared to those rates for 10 matched municipalities with unfluoridated water. The two groups had similar urbanization levels and sociodemographic characteristics. Our study does not support the suggestion that fluoridation of water supplies is associated with an increase in cancer mortality in Taiwan. PMID- 10702327 TI - Estimating cancer risk from outdoor concentrations of hazardous air pollutants in 1990. AB - A public health concern regarding hazardous air pollutants (HAPs) is their potential to cause cancer. It has been difficult to assess potential cancer risks from HAPs, due primarily to lack of ambient concentration data for the general population. The Environmental Protection Agency's Cumulative Exposure Project modeled 1990 outdoor concentrations of HAPs across the United States, which were combined with inhalation unit risk estimates to estimate the potential increase in excess cancer risk for individual carcinogenic HAPs. These were summed to provide an estimate of cancer risk from multiple HAPs. The analysis estimates a median excess cancer risk of 18 lifetime cancer cases per 100,000 people for all HAP concentrations. About 75% of estimated cancer risk was attributable to exposure to polycyclic organic matter, 1,3-butadiene, formaldehyde, benzene, and chromium. Consideration of some specific uncertainties, including underestimation of ambient concentrations, combining upper 95% confidence bound potency estimates, and changes to potency estimates, found that cancer risk may be underestimated by 15% or overestimated by 40-50%. Other unanalyzed uncertainties could make these under- or overestimates larger. This analysis used 1990 estimates of concentrations and can be used to track progress toward reducing cancer risk to the general population. PMID- 10702328 TI - Metals in albatross feathers from midway atoll: influence of species, age, and nest location. AB - Female birds sequester some heavy metals in their eggs, which are then transferred to the developing embryo. Semiprecocial birds such as albatrosses are fully covered with down at hatching, but are dependent on their parents for food for many weeks. At hatching, levels of metals in the chick's down represent exposure from the female via egg, while levels in fully formed feathers at fledgling, several months later, represent mainly exposure from food provided by their parents. In this paper we examine the concentrations of "metals" (heavy metals, mercury, lead, cadmium, chromium, manganese, tin; and metalloids, arsenic and selenium), in the down and contour (body) feathers of half-grown young albatrosses, and contour feathers of one of their parents. We collected feathers from Laysan Diomedea immutabilis and black-footed Diomedea nigripes albatrosses from Midway Atoll in the central Pacific Ocean. We test the null hypotheses that there is no difference in metal levels as a function of species, age, feather type, and location on the island. Using linear regression we found significant models accounting for the variation in the concentrations of mercury, lead, cadmium, selenium, chromium, and manganese (but not arsenic or tin) as a function of feather type (all metals), collection location (all metals but lead), species (selenium only), and interactions between these factors. Most metals (except mercury, arsenic, and tin) were significantly higher in down than in the contour feathers of either chicks or adults. Comparing the two species, black-footed albatross chicks had higher levels of most elements (except arsenic) in their feathers and/or down. Black-footed adults had significantly higher levels of mercury and selenium. We also collected down and feathers from Laysan albatross chicks whose nests were close to buildings, including buildings with flaking lead paint and those that had been lead-abated. Lead levels in the down and feathers of chicks close to nonabated buildings were 10 times higher than for chicks from other locations. Conversely, levels of cadmium and tin were lower near the buildings. Near lead-abated buildings, lead levels decreased as a function of distance, indicating residual contamination on the soil. Our results indicate that black-footed albatross adults and chicks generally have higher levels of heavy metals in their feathers than Laysans. Chicks of both species have higher levels in their down than in their contour feathers, indicating potentially higher exposure during the early chick phase. PMID- 10702329 TI - Urban CO exposure and its health effects on traffic policemen in Ankara. AB - Carbon monoxide (CO) is an important component of air pollution caused by traffic exhaust fumes. CO can cause chronic poisoning which shows its first symptoms as headaches, blurry vision, difficulty in concentration, and confusion. With the increasing number of vehicles in metropolitan areas of Turkey, the CO level has also increased in the city air as is the case in the capital city of Ankara, especially at certain locations. As far as the effects of CO on humans are concerned, traffic policemen are the population group under risk due to their inhalation of CO-rich air while on duty at the crowded cross-sections of the city. The traffic policemen on duty at these cross-sections are exposed to these high levels of CO for at least 6 h. This study was performed to investigate the traffic policemen (traffic organizers) who are exposed to high concentrations of CO at crowded cross-sections of Ankara City and to find out if chronic CO intoxication exits among this risk group. The CO levels in the ambient air at these cross-sections have also been compared to, and correlated with measurements of CO in the expired air of the target population. Additional factors like smoking, general health status, type of heating systems used at home, etc., have been taken into consideration by evaluating special questionnaires filled out by the policemen. A control group of clerk policemen, who were not engaged in street traffic activities was formed for comparative purposes. PMID- 10702330 TI - Lead exposure among small-scale battery recyclers, automobile radiator mechanics, and their children in Manila, the Philippines. AB - Blood lead (PbB) and hemoglobin levels (Hb) were determined in 40 battery repair/recycling shop workers, 16 radiator repair shop workers, and 20 children living in the immediate vicinity of these shops. Unexposed residents with similar socioeconomic status were also investigated. Mean PbB level was significantly higher for battery workers (54.23 microg/dL) when compared to radiator workers (20.04 microg/dL) and unexposed adults (12.56 microg/dL) (P<0.001). Among battery workers, 94% had PbB levels above the WHO permissible exposure limit of 40 microg/dL for males and 30 microg/dL for females. There was no demarcation between workplace and living quarters; therefore, workers' families were similarly exposed to hazards. Children living in the immediate vicinity of battery shops also had significantly higher mean PbB levels (49.88 microg/dL) compared to radiator shop children (11.84 microg/dL) and unexposed children (9.92 microg/dL). For workers with PbB > 40 microg/dL, 90% were anemic (Hb < 13 g/dL for males and <11.5 g/dL for females). Linear regression showed a correlation (r= 0.214; P=0.03) between Hb level and log(10)PbB. There was no significant relationship between anemia and blood lead in children (r=-0.146). We conclude that radiator repair activities appeared to increase the body burden of lead, although not up to a level significantly different from unexposed counterparts. Battery recycling/repair activities, however, significantly increased blood lead levels in workers and their children. PMID- 10702331 TI - Alterations in the properties and isoforms of sciatic nerve Na(+), K(+)-ATPase in methylcyclopentadienyl manganese tricarbonyl-treated mice. AB - The in vivo effect of methylcyclopentadienyl manganese tricarbonyl (MMT), an organic manganese-containing compound, on the mouse motor nerve was studied. The motor nerve conduction velocity was markedly decreased in MMT-treated mice. The Na(+),K(+)-ATPase activity of sciatic nerve isolated from MMT-treated mice was decreased; however, the sciatic nerve Na(+),K(+)-ATPase activity was not affected by the in vitro treatment of MMT. Moreover, [(3)H]ouabain binding of sciatic nerve isolated from MMT-treated mice was decreased. Using Western blot analysis, the amount of Na(+),K(+)-ATPase catalytic alpha1 subunit polypeptide in sciatic nerve of MMT-treated mice was also decreased. These results indicate that a causal relationship may exist between reduced nerve Na(+),K(+)-ATPase activity and motor nerve conduction velocity in MMT-treated mice and that a measurable decrease in alpha1 catalytic subunit isoform of Na(+),K(+)-ATPase may be necessary for the development of peripheral neuropathy by MMT. PMID- 10702332 TI - Involvement of oxidative stress in crystalline silica-induced cytotoxicity and genotoxicity in rat alveolar macrophages. AB - Alveolar macrophages (AMs) occupy a key position in silica-induced pulmonary fibrosis, although the mechanisms are yet to be elucidated. In the present study we examined the involvement of oxidative stress and reactive oxygen species formation in silica-induced cytotoxicity and genotoxicity in cultured rat AMs. A lucigenin-dependent chemiluminescence test was used to determine superoxide anion (O(-)(2)), and a 2',7'-dichlorofluorescin diacetate fluorescence test was employed to measure the hydrogen peroxide (H(2)O(2)) level. The cytotoxic and genotoxic effects caused by silica in AMs were examined by lactate dehydrogenase (LDH) leakage and single-cell gel electrophoresis (comet assay), respectively. The results showed that silica enhanced O(-)(2) and H(2)O(2) formation in AMs. There were clear dose- and time-dependent relationships in silica-induced cytotoxicity and genotoxicity. Furthermore, superoxide dismutase and catalase were able to reduce silica-induced LDH leakage and DNA damage, with concurrent significant inhibition on silica-induced oxidative stress in AMs. These findings provide convincing evidence that oxidative stress mediates the silica-induced cytotoxicity and genotoxicity. The understanding of such a mechanism may provide a scientific basis for the possible application of antioxidants in preventing the hazardous effects of silica. PMID- 10702333 TI - A comparison of sampling media for environmental viable fungi collected in a hospital environment. AB - Quantitative evaluation of fungal exposure is often conducted by analysis of the composition of microbes in air samples and calculation of the concentrations afterward. The collecting medium that favors the growth for most saprophytic fungi is considered to be the ideal choice in most circumstances. Currently, the culture medium most frequently adopted in environmental sampling for airborne fungi is MEA (malt extract agar) recommended by the ACGIH for its suitability for most fungal growth. DG18 (dichloran glycerol-18), developed in 1980, is suggested for growth at lower water activity (a(w)=0.95) specifically and is not as commonly used in general studies. This investigation collected airborne viable fungi using a single stage/N6 Andersen impactor with MEA and DG18 agar plates attached simultaneously to the same set of samplers. The sampling locations were at 17 sites within a central air-conditioned hospital. After incubation and morphological identification, concentrations of airborne fungi and bacteria were expressed as CFU/m(3) (colony forming units/m(3)). There are 405 DG18 plates and 378 plates available for statistical analysis. Results show that the airborne fungal concentrations, shown by geometric mean (GM), are higher from the DG18 plates than from the MEA plates. The total fungal concentrations is 68.6 vs 12.94 CFU/m(3), and for Aspergillus spp., the concentration is 1.58 vs 0.72 CFU/m(3); for Penicillium spp., 3.37 vs 0.71; and for yeast, 5.09 vs 0.49 CFU/m(3). In addition, the number of different genera present is greater on the DG18 plates than on the MEA plates, on average, 2.85 types vs 1.72. This study suggests that in a hospital environment with 24-h, central air conditioning, DG18 plates appear to be more effective in collecting more fungal colonies in terms of both quantity and types of genera. Such a finding is presumed to be attributed to the characteristic of DG18 in slowing colony growth so that the dominating genus will not over occupy the culture plate surface before the less competitive genus can fully develop. Future studies on related biological mechanisms are essential to conclude whether the above results sustain when sampling is conducted in other environments. PMID- 10702334 TI - Tissue uptake of bismuth from shotgun pellets. AB - Shotgun pellets containing bismuth have been suggested to be less environmentally toxic than those containing other metals. We sought to find if bismuth from shotgun pellets embedded within an animal enters the tissues of that animal. Five bismuth-containing shotgun pellets were placed intraperitoneally into adult mice. Four or 9 weeks later the tissue distribution of bismuth was examined histologically using silver lactate autometallography. Bismuth was seen in the nervous system of the mice, either in cells with processes outside the nervous system or in cells not protected by the blood-brain barrier. Bismuth was also seen in the kidney, liver, spleen, and lung. The amount of bismuth within tissues varied widely between animals at both time intervals. Bismuth from shotgun pellets enters the tissues of mice, with some mice taking up more bismuth than others. Some animals wounded with bismuth pellets are therefore likely to accumulate large amounts of potentially toxic bismuth in their tissues. PMID- 10702336 TI - Environmental research, section B PMID- 10702335 TI - Environmental and geographical factors contributing to watershed contamination with Cryptosporidium parvum oocysts. AB - Cryptosporidium parvum is a waterborne parasite which infects cattle and produces life-threatening zoonosis in people with impaired immune systems. Digital maps of 100-year floodplain boundaries, land use/cover, and livestock operations were used to select and characterize cattle farms in the floodplain area in Lancaster County, Pennsylvania, U.S.A. Over 21% of the cattle farms were located within 100 year floodplain boundaries. On average, a single farm comprised 12.8 ha of pasture (including buildings and farmyard) at risk of inundation. In all farms cattle had unlimited access to the creek. Manure samples collected from closed-in calf pens, cow/heifer yard runoff, and cattle paths through the creek were tested for C. parvum. On 64% of the farms (n=50) at least one sample was positive for C. parvum, and 44% of the farms had oocysts in all manure samples. Concentration varied from 90 to 371 oocysts/g and was significantly higher (P<0.02) in calf samples than in manure from cow and cow/heifer. PMID- 10702337 TI - Arsenic, dioxin, and the promotional step in cancer creation. AB - With an uncanny symmetry, both arsenic and dioxin act at the promotional step of cancer creation in a select but broad array of tissues: arsenic to promote initiated cancer cells and dioxin to promote blocking them. The symmetry is explored. PMID- 10702338 TI - Microbial resistance to metals in the environment. AB - Many microorganisms demonstrate resistance to metals in water, soil and industrial waste. Genes located on chromosomes, plasmids, or transposons encode specific resistance to a variety of metal ions. Some metals, such as cobalt, copper, nickel, serve as micronutrients and are used for redox processes, to stabilize molecules through electrostatic interactions, as components of various enzymes, and for regulation of osmotic pressure. Most metals are nonessential, have no nutrient value, and are potentially toxic to microorganisms. These toxic metals interact with essential cellular components through covalent and ionic bonding. At high levels, both essential and nonessential metals can damage cell membranes, alter enzyme specificity, disrupt cellular functions, and damage the structure of DNA. Microorganisms have adapted to the presence of both nutrient and nonessential metals by developing a variety of resistance mechanisms. Six metal resistance mechanisms exist: exclusion by permeability barrier, intra- and extra-cellular sequestration, active transport efflux pumps, enzymatic detoxification, and reduction in the sensitivity of cellular targets to metal ions. The understanding of how microorganisms resist metals can provide insight into strategies for their detoxification or removal from the environment. PMID- 10702339 TI - Anti-HIV activity of extracts and compounds from algae and cyanobacteria. AB - The human immunodeficiency virus (HIV) is the retrovirus that causes the acquired immune deficiency disease syndrome (AIDS). This review discusses the anti-HIV activity of extracts and compounds isolated from freshwater and marine algae, and cyanobacteria (formerly called "blue-green algae"). Compounds and extracts with anti-HIV activity are also active against other retroviruses such as herpes simplex virus (HSV), but the amount of antiviral activity varies with the compound and the virus. Most of the research has focused on sulfated homopolysaccharides and heteropolysaccharides. Sulfoglycolipids, carrageenans, fucoidan, sesquiterpene hydroquinones, and other classes of compounds with anti HIV activity that have been isolated from algae have received less attention. Most studies have used in vitro test systems, but a few in vivo studies have been carried out using compounds isolated from algae or analogs produced synthetically or isolated from other natural sources. Sulfated homopolysaccharides are more potent than sulfated heteropolysaccharides. The presence of the sulfate group is necessary for anti-HIV activity, and potency increases with the degree of sulfation. Studies using nonsulfated and sulfated homo- and heteropolysaccharides isolated from algae or other natural sources, or synthesized, have revealed the mechanisms of binding of drugs to the virion, and the mechanisms of viral binding to host cells. However, given the few classes of compounds investigated, most of the pharmacopeia of compounds in algae and cyanobacteria with antiretroviral activity is probably not known. PMID- 10702340 TI - Specific detection of membrane-toxic substances with a conductivity assay. AB - A conductivity assay that represents a new biotest able to detect the effects of membrane-toxic compounds, e.g., detergents, organic solvents, and radical formers, on various organisms was previously described and developed. The conductivity assay measures ion leakage from cells, tissues, or whole plant and animal organisms whose membrane systems have been damaged by membrane-toxic compounds. In this study the specificity of the conductivity assay for membrane toxic compounds was tested by comparing the electrolyte efflux from Elodea canadensis leaves during incubation with a well-known detergent (benzalkonium chloride) using different plant physiological and biochemical techniques (photochemical efficiency, plasmolysis capacity, NBT reduction, and electron microscopy of membranes of E. canadensis leaves). The comparison of the different methods proved that the electrolyte loss during benzalkonium chloride incubation determined in the conductivity assay is due to membrane impairment. The observed electrolyte loss correlated with a reduction of photochemical efficiency and a decrease in both plasmolysis and NBT reduction capacity. Furthermore, a disintegration of the plasmalemma could be seen in the electron micrographs. These results indicate that the measured electrolyte loss in the conductivity assay is a specific effect of membrane-toxic compounds. PMID- 10702341 TI - A simple and rapid evaluation of methemoglobin toxicity of 8-aminoquinolines and related compounds. AB - Methemoglobin, a toxic ferric form of hemoglobin, is continuously formed in normal erythrocytes, but during abnormal situations in situ, the level is enhanced. 8-Amino-quinolines and related compounds are causative agents for methemoglobin formation. Employing oxyhemoglobin, methemoglobin toxicity was about six times higher with primaquine compared to CDRI Compound 80/53 at 10(-9) M concentration. Methemoglobin reductase activity was also completely inhibited by primaquine, whereas 24% inhibition was noted in the case of 80/53 at the same concentrations. Mastomys, a rodent animal model, was found to be equally good for comparative evaluation of methemoglobin toxicity. Further, with the use of primaquine transdermal tape on the Mastomys model, a rise in methemoglobin occurred with increase in time. In conclusion, the study presents simple, economical, less time-consuming methods for the evaluation of methemoglobin toxicity, in vitro and in vivo, without employing the conventional Beagle dog model. PMID- 10702342 TI - Use of bioassays for assessment of water-extractable ecotoxic potential of soils. AB - The characterization of contaminated soils is based on heterogeneous strategies considering chemical analyses or bioassays. In the report Bioassays for soils, test methods which are at an advanced state of development and standardization are recommended by the German DECHEMA (German Society for Chemical Apparatus, Chemical Engineering and Biotechnology e.V.). Following this report six soil samples contaminated with different organic and inorganic substances are applied to bioassays using the following organisms: Scenedesmus subspicatus, Daphnia magna, Vibrio fischeri, and Pseudomonas putida. Additionally, they are chemically analyzed. The test results demonstrate that toxic contaminants are present and bioavailable in water elutriates from two soils, indicating the necessity of bioassays and chemical analyses to predict potential risks from contaminated soils. Furthermore it is demonstrated that the described qualitative approach for the assessment of test results is not sufficient to quantify the risk potential of contaminated soils. PMID- 10702343 TI - Organochlorine pesticides in wolves from Galicia. AB - Levels of seven organochlorine pesticides (heptachlorepoxide, dieldrin, endrin, p,p'-DDE, o-p'-DDT, p,p'-DDT, and methoxychlor) and DDE/DDT ratios were determined in spleen, liver, muscle, kidney and suprarenal from 12 wolves from three provinces of Galicia (eight male and four female). Analysis was carried out by GC-ECD. Heptachlorepoxide was in found only 25% of samples, while p,p'-DDE was the most dominant of the organochlorine compounds analyzed. DDE/DDT ratios higher than 1 were found in liver and muscle. PMID- 10702344 TI - Toxicity of sodium molybdate and sodium dichromate to Daphnia magna straus evaluated in acute, chronic, and acetylcholinesterase inhibition tests. AB - As a result of a widespread application in numerous industrial processes, chromium is a contaminant of many environmental systems. Chromium and their compounds are toxic to both invertebrates and vertebrates and, for this reason, there has been a search for suitable and less toxic alternatives. Molybdenum compounds have been studied as alternative to chromium compounds for some industrial applications. The toxicity of chromium is well known but the effects of molybdenum and molybdenum mining on natural populations and communities of freshwater invertebrates have not often been studied. However, chromium, and molybdenum (and their compounds) are included in the same list (List II) of European Union dangerous substances. In this study, the acute and chronic effects of sodium molybdate and sodium dichromate to Daphnia magna Straus were evaluated. Furthermore, in vitro and in vivo effects of these two metals on acetylcholinesterase (AChE) activity of D. magna Straus were investigated. LC(50) values determined at 48 h were 0.29 and 2847.5 mg L(-1) for chromium (as sodium dichromate) and molybdenum (as sodium molybdate), respectively. No significant in vitro effects of both metals on AChE were found. However, both toxicants inhibited AChE in vivo at concentrations under the respective 48-h LC(50) values. Both sodium dichromate and sodium molybdate inhibited the reproduction and growth of D. magna, but the concentrations inducing significant effects were different for the two chemicals. Sodium molybdate had significant lower toxicity to D. magna Straus than sodium dichromate. PMID- 10702345 TI - Cellular method for evaluation of noxiousness of inorganic pollutants in industrial wastes: calculation of a safety index for monitoring sludge discharge. AB - This article deals with a biological test of safety applicable to industrial wastes. The test is based on the measurement of the growth rate of cultured human cells exposed to waste samples with different dilutions. As a first approach, 15 chemicals in which discharge concentrations are submitted to sanitary regulations were tested one by one. For Zn, Cu, Ni, Cd, Ag, Co, Mg, sulfates, and fluorides, it was possible to detect concentrations that are below the allowed limit. For Hg, Al, As(V), Cr(III), Fe, and Pb, the concentrations that affect cell growth are higher than the allowed limit. Tests were also performed using actual samples (liquid effluent from a laundry and sludge from waste-water treatment plants). Results indicate that, in contrast to chemical analyses, the current biological test has the advantage of providing an indication of global toxicity, integrating all substances and factors that can be harmful to life processes. From the sludge data and the observed threshold of concentration that does not affect cell growth, a numeric safety index has been calculated which indicates the amount of sludge that could be dispersed, as a fertilizer, per hectare of agricultural soil. Such an index could be conveniently used for designing sewage sludge disposal strategies. PMID- 10702346 TI - Evaluation of nickel-zinc interactions by means of bioassays with amphibian embryos. AB - The nickel hazard was evaluated by means of a 7-day toxicity test with Bufo arenarum embryos. The LC(50) values for this metal from 24 to 168 h diminished from about 26 to 1.8 mg Ni(2+)/L, respectively, but from 96 h onward, the LC(50) varied very slightly. Although a noticeable difference among the LC(50) and LC(10) or LC(90) was observed at 24 h of exposure, these parameters tended to a similar value at 168 h of exposure while the confidence intervals of LC(50) overlapped all other confidence interval values. These results, plotted as toxicity profile curves, are useful for determining time and concentration thresholds for Ni. Nickel-zinc interactions on B. arenarum embryos were evaluated by means of simultaneous treatments with both cations (Ni: 5-35 mg Ni(2+)/L; Zn: 0.5-130 mg Zn(2+)/L). As a general pattern, low Zn concentrations (0.5 mg Zn(2+)/L) did not have a clear-cut effect on Ni toxicity, higher Zn concentrations (2-20 mg Zn(2+)/L) enhanced Ni toxicity, and concentrations of 30 mg Zn(2+)/L and higher had a beneficial effect in most cases. The metal interaction studies provide a scientific basis for the establishment of water quality criteria for wildlife protection purposes. PMID- 10702347 TI - Shake-flask test for determination of biodegradation rates of (14)C-labeled chemicals at low concentrations in surface water systems. AB - A simple shake-flask surface water biodegradability die away test with (14)C labeled chemicals added to microgram per liter concentrations (usually 1-100 microg/L) is described and evaluated. The aim was to provide information on biodegradation behavior and kinetic rates at environmental (low) concentrations in surface water systems. The basic principle of measurement was to determine evolved CO(2) indirectly from measurements of total organic activity in subsamples after stripping off their content of CO(2). Used with surface water alone the test simulates a pelagic environment and amended with sediments (0.1-1 dry weight/L) the test is intended to simulate a water environment with suspended solids (e.g., resuspended sediments). A protocol of the test used with the (14)C technique or with specific chemical analysis was recently developed by the International Organization for Standardization. Practical experience with the method is presented for a set of reference substances. These substances could be ranked in five groups of decreasing biodegradability: aniline>p-nitrophenol, 2, 4 dichlorophenoxyacetic acid>4-chloroaniline>maleic hydrazide, pentachlorophenol>atrazine. It was found that degradation rates and lag periods varied considerably among sampling sites and sometimes also among samples from the same site. No significant correlation could be established between degradation rates and microbial biomass estimates. Even small portions of added sediments greatly enhanced biodegradation of the absorbable compound pentachlorophenol, probably by providing sites for microbial attachment. Repeated tests indicated consistent degradation behavior for the readily degradable substances, whereas degradation sometimes stopped or failed with the more recalcitrant substances. A preadaptation step involving regular reinoculation with freshly collected surface water could, however, overcome the problems of false-negative results. PMID- 10702348 TI - Interaction of aluminum with exogenous and endogenous iron in the organism of rats. AB - The aim of these experiments was to find changes in free erythrocyte protoporphyrins (FEP) and in the concentration of endogenous iron in the blood, erythrocytes, serum, liver, kidneys, and spleen of rats, as well as in the dynamics of aluminum concentrations in the serum of rats after oral application of aluminum chloride (AlCl(3)) separately or with ferrum chloride (FeCl(2)), depending on the time and doses administered. The experiments were carried out on female Wistar rats which received (p.o.) 100 mg Al/kg separately or with iron (4 mg Fe/kg) daily for 35 days. The effects of aluminum administration were noticed after the second week. The experiments demonstrated that the increase in the level of free erythrocyte protoporphyrins in the blood is the most sensitive indicator of exposure to AlCl(3). A decrease in iron concentration in erythrocytes, blood, and spleen was also noticed. The response and the sequence of the investigated effects were recorded according to aluminum and iron concentration in the serum. Joint administration of iron and aluminum decreases concentration of aluminum in serum and prevents changes in the investigated indicators in rats exposed to aluminum chloride. PMID- 10702349 TI - Effects of dietary cadmium and its bioconcentration in tilapia Oreochromis niloticus. AB - The administration of cadmium, as food supplement, its bioaccumulation, and the effects on the development of tilapia Oreochromis niloticus were investigated. The average size and weight and its behavior compared with controls were investigated during the period January 31, 1997, until March 31, 1999. At intervals of 60 days the measurements of size and weight were performed, and the concentration of cadmium in feces, water, muscular tissue, and viscera were determined by atomic absorption spectroscopy. The initial average cadmium concentration in food was 5 mg small middle dotkg(-1) and only after 6 months a small effect on size and weight could be observed. With increases in cadmium concentration to 50 mg small middle dotkg(-1), beginning after the 7th month, and 100 mg small middle dotkg(-1) after the 16th month, a clear difference in size and weight and also in behavior could be observed. An LC(50) value of 40 mg small middle dotkg(-1) was observed after the 23rd month. PMID- 10702350 TI - Changes in the metabolic elimination profile of testosterone following exposure of the crustacean Daphnia magna to tributyltin. AB - The biocide tributyltin has been found to cause the development of pseudohermaphroditic conditions in some neogastropod species. These abnormalities of the reproductive system have adversely affected the fecundity of some field populations of gastropods, resulting in local population declines. Current evidence suggests that tributyltin elicits these effects by interfering with the biotransformation of testosterone to other steroid derivatives, resulting in an elevation in endogenous testosterone or some of its bioactive derivatives. The purpose of the present study was to determine whether tributyltin altered testosterone metabolism in daphnids (Daphnia magna), a species commonly used in ecotoxicology testing. Exposure of daphnids to 1.2 microg (tin)/L caused a general increase in the rate of elimination of oxido-reduced, hydroxylated, and glucose-conjugated derivatives of testosterone. However, tributyltin exposure had no significant effect on the rate of elimination of the glucose-conjugated forms of the various oxido-reduced and hydroxylated derivatives of testosterone. As a result, the percentage of the oxido-reduced and hydroxylated metabolites of testosterone eliminated as glucose conjugates decreased with increasing tributyltin exposure levels. These results demonstrate that tributyltin causes alterations in testosterone metabolism in daphnids that would result in an increase in the production of oxido-reduced derivatives. These products are preferentially retained in the tissues of daphnids and are variously androgenic in vertebrates. The increased production of oxido-reduced derivatives of testosterone may be mechanically responsible for the masculinizing effects of tributyltin in some species and suggests that daphnids may be a suitable surrogate for evaluating the potential of chemicals to elicit this form of toxicity. PMID- 10702351 TI - Pharmacokinetics of intravascularly administered (65)Zinc in channel catfish (Ictalurus punctatus). AB - Comparison was made of the pharmacokinetics of the radioisotope (65)Zinc ((65)Zn) in blood, plasma, and whole body of adult channel catfish (Ictalurus punctatus) following intravascular (iv) administration. A two-compartment model described the pharmacokinetics of (65)Zn in plasma and blood during the first 40 days following iv administration, but was unable to describe the long-term disposition of (65)Zn. Whole-body counting revealed that approximately half of the (65)Zn dose was sequestered in a slowly exchangeable pool with a half-life of 1.5 years. Greater than 99% of the circulating (65)Zn was bound to plasma proteins, whereas there was less than 1% binding to red blood cells. Synthesis of the results for channel catfish and existing data in other species indicates three phases in the pharmacokinetics of zinc. The first phase consists of initial distribution outside the vascular system to kidney, liver, and other organs (alpha phase in blood and plasma; t(1/2) of 4 to 5 h). The second phase involves distribution from organs to a slowly exchangeable zinc pool, likely consisting of bone (beta phase in blood and plasma; alpha phase in whole body; t(1/2) of 4 to 20 days). The third phase appears to involve a slow turnover of sequestered zinc (t(1/2) greater than 1 year). Blood sampling or short-term whole-body measurements will underestimate the persistence of zinc in fish, thus prolonged sampling and measurement of whole-body concentrations are necessary to characterize the pharmacokinetics of zinc. PMID- 10702352 TI - Monitoring Wadi El Raiyan lakes of the egyptian desert for inorganic pollutants by ion-selective electrodes, ion chromatography, and inductively coupled plasma spectroscopy. AB - Wadi El Raiyan is a great depression located southwest of Cairo in the western desert of Egypt, one of the most arid regions of the world. In 1973, Wadi El Raiyan was connected with the agricultural wastewater drainage system of the El Faiyum province to provide a reservoir for the wastewater that exceeded the capacity of Lake Qarun north of the province. Pollutants from agricultural waste including pesticides and fertilizers as well as other effluents of industrial activities and runoffs certainly will pass into the biotic elements of the ecosystem. This report presents the status of inorganic pollutants including anions, cations, and trace metals in the two lakes and the surrounding springs of Wadi El Raiyan using ion chromatography, ion-selective electrodes, and inductively coupled plasma emission spectroscopy. The report also includes the levels of selected metals in the vegetation community of the area. The result of this investigation revealed a great improvement in water quality of the Wadi El Raiyan lakes compared to 1988 report by Saleh et al. Mercury was not detected in any of the samples and the level of lead was significantly reduced. Cadmium levels were much higher than those seen earlier. The higher level of cadmium might be used as an indicator to track the contamination of water by human waste. Concentrations of common anions were not significantly different from those reported earlier. However, an increase in the level of cyanide was observed. Levels of heavy metals in vegetation around the lakes were also found to be lower than previously reported. PMID- 10702353 TI - Papers to Appear in Environmental Research Section A. PMID- 10702354 TI - Post-transcriptional regulation of endothelial cell plasminogen activator inhibitor-1 expression during R. rickettsii infection. AB - Intracellular infection of endothelial cells with Rickettsia rickettsii results in increased steady-state levels of plasminogen activator inhibitor-1 (PAI-1) mRNA. Control mechanisms governing such increased expression in response to this novel stimulus have not been defined. In this study, we compared the stability of PAI-1 mRNA in infected and uninfected endothelial cells (EC) and explored the requirement for de novo host cell protein synthesis in the infection-induced increase of steady-state levels. The half-life of PAI-1 mRNA, which is constitutively expressed in cultured EC, increased from 18 h in uninfected EC to greater than 30 h in EC infected for 24 h, a time point at which increases in steady-state PAI-1 mRNA levels are noted. There was no change in stability of gamma-actin due to infection. Nuclear run-on studies revealed no apparent increase in transcription rate at 4, 18 and 24 h. R. rickettsii -induced increase in PAI-1 mRNA was blocked by the eukaryotic protein synthesis inhibitor, cycloheximide, which suggests that this response requires de novo host cell protein synthesis. These results provide evidence that post-transcriptional control mechanisms are operative in the regulation of PAI-1 during R. rickettsii infection. PMID- 10702355 TI - Sequence and characterization of the glyceraldehyde-3-phosphate dehydrogenase of Mycobacterium avium: correlation with an epidermal growth factor binding protein. AB - Mycobacterium avium is a common pathogen in AIDS patients. The extracellular environment within the granuloma shown to support mycobacterial growth is in the caseous fluid. Previous work demonstrated that the presence of human epidermal growth factor (EGF), which is found in the tissue of chronic granulomous lesions, increases the growth rate of M. avium and Mycobacterium tuberculosis. Previously, a protein capable of binding recombinant human EGF (rEGF) in a western blot was identified with homology to glyceraldehyde-3-phosphate dehydrogenase (GAP) in both M. avium and M. tuberculosis but not Mycobacterium smegmatis. Surface GAPs have been identified in group A Streptococcus, enteropathogenic Escherichia coli, Candida albicans and Schistosoma mansoni. We have cloned the gap gene of M. avium. M. avium GAP has high homology with M. tuberculosis GAP. The protein was also expressed in M. smegmatis, conveying the ability to bind rEGF, but no growth increase was observed in 7H9 broth in the presence of rEGF up to 500 ng/ml. Only one copy of the GAP gene was identified in M. avium These results contribute to the understanding of M. avium pathogenesis by characterizing its interaction with a host protein present in the site of infection. PMID- 10702356 TI - Characterization of P1-deficient isogenic mutant of Haemophilus influenzae biogroup aegyptius associated with Brazilian purpuric fever. AB - Haemophilus influenzae biogroup aegyptius (formerly H. aegyptius) is the etiologic agent of Brazilian purpuric fever (BPF). A surface-exposed epitope on the outer membrane protein P1 is present on most strains of H. influenzae biogroup aegyptius associated with BPF but is absent in almost all non-disease associated strains. The role of the outer membrane protein P1 in the pathogenesis of this disease was evaluated by utilizing an isogenic P1-deficient mutant. We compared the ability of the wild type and P1 isogenic mutant to grow under various conditions. The P1-deficient strain grew at a similar rate to the wild type in both complex and chemically defined medium. The P1-deficient mutant also had a similar growth rate to the wild type under anaerobic conditions. Anaerobic growth, however, resulted in up-regulation of the P1 protein in the wild type strain. Three assays were used to examine the pathophysiologic role of the P1 protein in BPF: 1) serum resistance; 2) sustained bacteremia in the infant rat model; and 3) the human microvascular endothelial cell (HMEC) cytotoxicity assay. Both the mutant and wild-type strains were resistant to killing in 95% normal human serum. The P1-deficient strain was also as virulent as the wild type in both the infant rat model of bacteremia and in the HMEC-1 tissue culture model. These results demonstrate that serum resistance, sustained bacteremia in the infant rat, and cytotoxicity of HMEC cells occur in the absence of P1. The P1 protein is not essential for the pathogenic potential identified by these assays. However, these results demonstrate that an anaerobic environment is a potent physiologic regulator of P1 protein expression. The impact of anaerobiosis on protein expression and pathogenesis will require further investigations. PMID- 10702357 TI - Immune recognition of porin and lipopolysaccharide epitopes of Salmonella typhimurium in mice. AB - We investigated the antigenic specificity of the humoral immune response to infection by Salmonella typhimurium, by competitive inhibition enzyme-linked immunosorbent assay and Western immunoblots. A panel of eight murine monoclonal antibodies, raised to OmpC and OmpD porins and lipopolysaccharide (LPS)-O antigens, was used to define the specificity of the polyclonal immune response in mice. The monoclonal antibody panel recognized five distinct epitopes; these were localized to surface-exposed loops of OmpC and OmpD porin, to the "eye-let" forming loop L3 of OmpC/OmpD, and to LPS-O4 and O5 factors. The immune mouse serum raised to infections with S. typhimurium LT-2 strain WB600 (wild-type) competitively inhibited the binding of biotin-labelled monoclonal antibodies to the epitopes that they recognize, indicating that all five epitopes were targets of the host immune response to natural infection. However, only two epitopes, one within a surface-exposed loop of OmpC porin, and the other in the LPS-O4 factor, were immunodominant. Furthermore, the bacterial LPS core and O-antigen structure influenced the immune response to the porins. Surface epitopes of porins were dominant in the rough strain SH5014 (rfa), whereas the immune recognition of LPS epitopes was predominant in mice infected with the smooth, wild-type strain (WB600). Finally, the immune response to LPS epitopes O4 and O5 was more pronounced in mice immunized with heat-killed cells than those infected with live S. typhimurium. PMID- 10702358 TI - Applying in vivo expression technology (IVET) to the fungal pathogen Histoplasma capsulatum. AB - Understanding how pathogens survive within the host cell is of paramount importance in the development of vaccines and therapeutic agents. This task has been particularly daunting in the study of fungal pathogens due to the lack of easily manipulated genetic systems. In recent years several molecular genetic reporter systems have been developed to identify genes expressed during the infection process and potential virulence determinants. The development of one method in particular, in vivo expression technology (IVET), has led to the discovery of several genes from various bacterial pathogens necessary for survival during infection. The recent development of molecular genetic tools for Histoplasma capsulatum has enabled us to adapt the IVET technology for this pathogenic fungus utilizing the URA5 gene, which is essential for H. capsulatum survival in mice and in cultured macrophages, as a reporter of in vivo gene expression. We report the first successful application of IVET screening of a fungal pathogen for genes expressed exclusively during infection. PMID- 10702359 TI - Characterization of a putative virulence island in the chromosome of uropathogenic Escherichia coli possessing a gene encoding a uropathogenic specific protein. AB - This study was initiated to search for a homologue of the Vibrio cholerae zot gene in uropathogenic Escherichia coli (UPEC) using a specific DNA probe. The faint signal obtained at low stringency with some UPEC strains associated with prostatitis cases prompted us to examine UPEC strains by PCR using primers designed from the conserved regions of the proteins of the Zot group of putative NTPases containing the classical NTP binding motif. This led to the discovery of a DNA fragment in UPEC strains which hybridized with a probe designed from the PCR. Further analysis of this DNA fragment revealed an ORF which was designated as uropathogenic specific protein (Usp). The gene encoding Usp was 1038 bp long and codes for 346 amino acids with an appropriate SD sequence. Upstream and downstream analysis of usp revealed motifs of prokaryotic consensus promoters and three small ORFs with SDs and ribosome binding sites transcribed in the same direction of usp. The proximity of these set of genes in a specific area of the bacterial chromosome resembling a block of genes preferentially associated with UPEC coupled with the presence of a motif matching that of a Tn3 transposon family lead us to believe that this could be an hitherto unknown pathogenicity island. PMID- 10702360 TI - Mechanisms of cyclosporine A-induced apoptosis in rat hepatocyte primary cultures. AB - In rat hepatocytes and isolated liver mitochondrial fractions, Cyclosporine A (CsA) is often used as a specific inhibitor of mitochondrial Ca(2+) release and as a specific blocker of mitochondrial membrane potential and permeability transition (MPT), which are all processes involved in the inhibition of apoptosis. However, neither inhibition nor induction of apoptosis by CsA has yet been described in the rat hepatocyte primary culture during incubation for 4 and 20 h. It was the purpose of the present study to examine by means of morphological and biochemical criteria the effects of CsA on apoptosis and to characterize the underlying mechanisms. Rat hepatocytes were cultured for 4 or 20 h with CsA at concentrations of 0, 10, 25, and 50 microM. Chromatin condensation and fragmentation, DNA fragmentation (TUNEL), membrane phosphatidylserine distribution (Annexin V), caspase-1, -3, and -6 activity, mitochondrial membrane potential (Rhodamine 123), and cytochrome c release into the cytosol were investigated. Four hours after CsA treatment, chromatin condensation and fragmentation and the number of TUNEL- and Annexin V-positive cells increased dose-dependently without any observable enzyme leakage, which indicated the integrity of the outer cell membrane. After 20 h of CsA incubation apoptosis parameters were further increased and were accompanied by the increased activity of the cysteine protease, caspase-3 (CPP 32), and slightly increased caspase-6 (Mch 2), but not caspase-1 (ICE). The caspase-3 inhibitor, Ac-DEVD-CHO, inhibited caspase-3 activation and attenuated CsA-induced apoptosis and LDH leakage. The caspase-6 inhibitor, Ac-VEID-CHO, only marginally inhibited CsA-induced apoptosis. Decreased mitochondrial membrane potential and cytochrome c release went in parallel with ultrastructural mitochondrial changes and might be regarded as early events that trigger the apoptosis cascade. Transmission electron microscopy confirmed an increase in the number of necrotic cells after 20 h, but not after 4 h, compared with controls. PMID- 10702361 TI - Residual oil fly ash induces cytotoxicity and mucin secretion by guinea pig tracheal epithelial cells via an oxidant-mediated mechanism. AB - Inhalation of ambient air particulate matter (PM) is associated with pulmonary injury and inflammation. Using primary cultures of guinea pig tracheal epithelial (GPTE) cells as an in vitro model of airway epithelium, we examined effects of exposure to suspensions of six different emission and ambient air PM samples: residual oil fly ash (ROFA) from an electrical power plant; fly ash from a domestic oil burning furnace (DOFA); ambient air dust from St. Louis (STL), Ottawa (OT), and Washington, DC (WDC); and volcanic ash from the eruption of Mount Saint Helens (MSH) in 1980. Effects of these particulates on cell viability (assessed via LDH assay), secretion of mucin (measured by a monoclonal antibody based ELISA), and steady-state mRNA levels of the mucin gene MUC2 were determined. ROFA was the most toxic of the dusts tested, as it significantly increased LDH release following a 24-h incubation with 50 microg/cm(2) ROFA. ROFA also enhanced MUC2 mRNA after 4-h exposure, and mucin secretion after 8 h. ROFA induced mucin secretion and cytotoxicity were attenuated by the oxidant scavenger, dimethylthiourea (DMTU). ROFA exposure also depleted cells of glutathione (GSH). Relatedly, depletion of intracellular GSH by treatment of the cells with buthionine sulfoxamine (BSO) also provoked mucin secretion, as well as enhancing the secretory effect of ROFA when the two agents were added together. L NMA, the nitric oxide synthase (NOS) inhibitor, did not affect ROFA-induced mucin secretion. Of the soluble transition metals in ROFA (nickel, iron, vanadium), only vanadium individually, or combinations of the metals containing vanadium, provoked secretion. The results suggest ROFA enhances mucin secretion and generates toxicity in vitro to airway epithelium via a mechanism(s) involving generation of oxidant stress, perhaps related to depletion of cellular antioxidant capacity. Deleterious effects of inhalation of ROFA in the respiratory tract in vivo may relate to these cellular responses. Vanadium, a component of ROFA, may be important in generating these reactions. PMID- 10702362 TI - Cadmium-induced acute hepatic injury is exacerbated in human interleukin-8 transgenic mice. AB - It is reported repeatedly that severe hepatocellular necrosis along with infiltration of neutrophils occurs after acute cadmium exposure. Neutrophils, which migrate by the gradient of chemoattractants such as interleukin-8, are believed to play an important role in inflammation at the damaged sites. To investigate whether neutrophils aggravate or repair the liver injury induced by cadmium, we checked the hepatotoxic effects of cadmium on human interleukin-8 transgenic mice (hIL-8Tg), which overexpressed IL-8 and displayed an inability of neutrophil migration resulting from both the lack of chemotactic gradient and the downregulation of l-selectin on the surface of neutrophils. A significantly lower survival rate was observed in hIL-8Tg compared with wild-type mice after subcutaneous administration of cadmium. Evident liver injury characterized by abrupt increases in plasma GOT and GPT levels was found in hIL-8Tg at 18 h after cadmium administration. Histological examinations, including H & E staining and esterase staining, revealed the infiltration of numerous neutrophils into the damaged liver tissues in wild-type mice, and the inhibition of the neutrophil migration into the liver as well as enhanced hepatocellular necrosis in hIL-8Tg. Peripheral white blood cell and polymorphonuclear cell counts increased and reached their peaks at 12 h after cadmium administration in wild-type mice, whereas the increase in blood leukocyte counts was delayed in hIL-8Tg. There was no significant difference in the amounts of cadmium accumulated in liver and kidneys between wild-type mice and hIL-8Tg. In conclusion, an acute cadmium hepatotoxic effect was exacerbated in hIL-8Tg resulting from inhibited neutrophil migration, suggesting that migrated neutrophils can prevent aggravation of liver injury by acute cadmium administration. PMID- 10702363 TI - Effects of microsomal enzyme inducers on outer-ring deiodinase activity toward thyroid hormones in various rat tissues. AB - Microsomal enzyme inducers, such as phenobarbital (PB), pregnenolone-16alpha carbonitrile (PCN), 3-methylcholanthrene (3MC), and Aroclor 1254 (PCB) are more effective at reducing serum thyroxine (T(4)) than serum triiodothyronine (T(3)). It is possible that rats treated with PB and PCN maintain serum T(3) by increasing serum TSH, which stimulates the thyroid gland to synthesize more T(3). However, it is unclear how serum T(3) is maintained in rats treated with 3MC or PCB, because serum TSH is not increased in these rats. We hypothesized that increased conversion of T(4) to T(3), catalyzed by outer-ring deiodinases (ORD) type-I and -II, is the reason serum T(3) is maintained in rats treated with 3MC or PCB. Furthermore, 3MC and PCB do not increase serum TSH, whereas PB and PCN do, because type-II ORD activity in the pituitary of 3MC- and PCB-treated rats is increased greater than in rats treated with PB or PCN. To test these two hypotheses, male Sprague-Dawley rats were fed either a basal diet or a diet containing PB (300, 600, 1200, or 2400 ppm), PCN (200, 400, 800, or 1600 ppm), 3MC (50, 100, 200, or 400 ppm), or PCB (25, 50, 100, or 200 ppm) for 7 days. Type I ORD activity was measured in thyroid, kidney, and liver, whereas type-II ORD activity was measured in brown adipose tissue, pituitary, and brain. Type-I ORD activity in thyroid was not affected by PB, 3MC, or PCB treatments, and was slightly increased by PCN. Type-I ORD activity in kidney was not affected by PB, PCN, or 3MC treatments, and was reduced by PCB treatment. Type-I ORD activity in liver was reduced by PB, PCN, 3MC, and PCB treatments. Type-II ORD activity in brown adipose tissue was unaffected by any of the four treatments. Type-II ORD activity in pituitary was unaffected by PB or 3MC treatments, and was increased by PCN or PCB treatments. Type-II ORD activity in brain was unaffected by PB treatment, and was increased by PCN, 3MC, and PCB treatments. Overall, total ORD activity, calculated by summation of ORD activities in thyroid, kidney, liver, brown adipose tissue, pituitary, and brain, was reduced rather than increased by the four microsomal enzyme inducers. In conclusion, increased conversion of T(4) to T(3) is not the reason serum T(3) concentration is maintained in 3MC- or PCB treated rats. Furthermore, the reason serum TSH is not increased in 3MC- and PCB treated rats is the result of mechanisms other than increased type-II ORD activity in pituitary. PMID- 10702364 TI - Enhanced expression of pulmonary gamma-glutamylcysteine synthetase heavy subunit in rats exposed to cadmium aerosols. AB - This investigation sought to determine the effect of cadmium (Cd) aerosol exposure on the pulmonary expression of the heavy subunit (HS) of gamma glutamylcysteine synthetase (gamma-GCS), the rate-limiting enzyme in de novo synthesis of glutathione (GSH). Using Northern hybridization analysis, we demonstrated that CdO inhalation caused time- and dose-dependent increases in the steady-state levels of gamma-GCS-HS mRNA that were highly correlated with lung Cd burden. Observed increases in gamma-GCS-HS gene expression were maximal 2 h following a single aerosol exposure to Cd and appeared to be triggered by an oxidant stress, characterized by a decline in the reduced to oxidized glutathione ratio. Immunoblotting of proteins in lung extracts from treated and untreated animals produced a single protein band corresponding to a molecular weight of 73 kDa. Elevated levels of gamma-GCS-HS mRNA and gamma-GCS-HS protein in lungs of Cd exposed animals were also accompanied by higher gamma-GCS enzymatic activity and elevations in glutathione (GSH). Immunohistochemical and in situ hybridization studies were used to identify compartments in the lung where Cd-induced expression of gamma-GCS-HS was localized. The most prominent staining for gamma GCS-HS protein and gamma-GCS-HS mRNA was observed in the alveolar epithelium of Cd-exposed animals. Quantitative image analysis confirmed a good agreement between relative levels of protein and mRNA transcripts for gamma-GCS-HS. These observations suggest that resistance to Cd toxicity in the lung may reflect the ability of specific lung cells to upregulate gamma-GCS expression and increase de novo GSH synthesis as an adaptive response. PMID- 10702365 TI - Lead targets GRP78, a molecular chaperone, in C6 rat glioma cells. AB - Exposure to potentially neurotoxic levels of lead (Pb) occurs in about 9% of American children under 6 years of age. Astroglia in the brain serve as a Pb depot, sequestering Pb and preventing its contact with the more sensitive neurons. Astroglia have the capacity to adapt to Pb exposure, and as such are able to tolerate relatively high intracellular Pb accumulation. This tolerance mechanism has yet to be defined in biochemical terms. In the present study, we present evidence that glucose-regulated protein (GRP78), a molecular chaperone in the ER, participates directly or indirectly in the tolerance mechanism. Exposure of cultured C6 rat glioma cells, an astroglia-like cell line, to 1 microM Pb acetate for 1 week raised the intracellular levels of two proteins, one of which was identified by sequence analysis as GRP78. GRP78 accumulation started within 1 day and progressed with time of exposure. Studies in vitro showed that GRP78 bound tightly to affinity columns with Pb(2+) as the affinity ligand and bound weakly when either Zn(2+) or Ni(2+) replaced the Pb(2+). The reduced form of GSH and BSA did not compete with GRP78 to chelate Pb(2+). However, the heavy metal binding domain (HMB) of Menkes protein competed with GRP78 for chelating Pb(2+). The data provide evidence that GRP78 may be a component of the Pb tolerance mechanism through its direct interaction with Pb(2+). Its increased synthesis could be part of the adaptive response to Pb exposure. PMID- 10702366 TI - Chloride secretion in kidney distal epithelial cells (A6) evoked by cadmium. AB - The effect of Cd(2+) on chloride secretion was examined in A6 renal epithelia cells by chloride-sensitive fluorescence (SPQ probe) and by the short-circuit current (I(sc)) technique. Depleting the cells of Cl(-) suggests that the Cd(2+) activated I(sc) (DeltaI(sc(Cd))) is dependent on the presence of Cl(-) ions. Among the Cl(-)-channel inhibitors the fenemates, flufenamic acid (FFA) and niflumic acid (NFA), and 5-nitro-2-(3-phenylpropylamino)-benzoate (NPPB) significantly lowered DeltaI(sc(Cd)) compared with control level. In SPQ-loaded A6 cells, Cd(2+) evoked an increase in Cl(-) secretion ([DeltaCl(-)](Cd)), which significantly exceeded the basal Cl(-) transport and was blockable by FFA and NFA. The closely related metals, Zn(2+) or Ni(2+), were also able to activate Cl( ) secretion. Preexposure of Zn(2+) or Ni(2+) completely prevented [DeltaCl( )](Cd), suggesting that Zn(2+) and Ni(2+) probably use similar mechanisms. Like Cd(2+), thapsigargin (TG), an inhibitor of intracellular Ca(2+)-ATPase and the Ca(2+)-ionophore A23187, induced an increase in I(sc). Moreover, TG and Cd(2+) were able to neutralize the responses of the counterparts as also observed in I(sc) measurements, which indicates that Cd(2+) activates Cl(-) secretion in a Ca(2+)-dependent manner. Hence, this study supports the idea that basolateral Cd(2+) (possibly also Zn(2+) and Ni(2+)), probably through a Ca(2+)-sensing receptor, causes calcium mobilization that activates apical fenemate-sensitive chloride channels leading to chloride secretion in A6 cells. PMID- 10702368 TI - Chemical index for volume 163 PMID- 10702367 TI - Pharmacokinetics of inhaled manganese phosphate in male Sprague-Dawley rats following subacute (14-day) exposure. AB - Methylcyclopentadienyl manganese tricarbonyl (MMT) is used as a gasoline octane enhancer. Manganese phosphate is the primary respirable (PM(2.5)) MMT-combustion product emitted from the automobile tailpipe. The goal of this study was to determine the exposure-response relationship for inhaled manganese phosphate in adult male CD rats. Rats were exposed 6-h/day for either 5 days/week (10 exposures) or 7 days/week (14 exposures) to manganese phosphate at 0, 0.03, 0.3, or 3 mg Mn/m(3) (MMAD congruent with 1.5 micrometer). The following tissues collected at the end of the 2-week exposure: plasma, erythrocytes, olfactory bulb, striatum, cerebellum, lung, liver, femur, and skeletal muscle (n = 6 rats/exposure group) were analyzed for manganese content by neutron activation analysis. Intravenous (54)MnCl(2) tracer studies were also conducted following the 14th exposure (n = 6 rats/concentration), and whole-body gamma spectrometry was performed immediately after injection and at 1, 2, 4, 8, 12, and 16 weeks after (54)MnCl(2) administration. Increased manganese concentrations were observed in olfactory bulb, lung, femur, and skeletal muscle following exposure to 3 mg Mn/m(3) (10 or 14 exposures). Increased manganese concentrations were also observed in olfactory bulb, striatum, and lung following exposure to 0.3 mg Mn/m(3) (14 exposures only). Red blood cell and plasma manganese concentrations were increased only in rats exposed to 3 mg Mn/m(3) (10 exposures). Rats exposed to 3 mg Mn/m(3) also had an increased whole-body manganese clearance rate when compared to air-exposed control animals. Our results suggest that the rat olfactory bulb may accumulate more manganese than other brain regions following inhalation exposure. PMID- 10702369 TI - Selectively attending to auditory objects. AB - The ability to maintain a conversation with one person while at a noisy cocktail party has often been used to illustrate a general characteristic of auditory selective attention, namely that perceivers' attention is usually directed to a particular set of sounds and not to others. Part of the cocktail party problem involves parsing co-occurring speech sounds and simultaneously integrating these various speech tokens into meaningful units ("auditory scene analysis"). Here, we review auditory perception and selective attention studies in an attempt to determine the role of perceptual organization in selective attention. Results from several behavioral and electrophysiological studies indicate that the ability to focus attention selectively on a particular sound source depends on a preliminary analysis that partitions the auditory input into distinct perceptual objects. Most findings can be accounted for by an object-based hypothesis in which auditory attention is allocated to perceptual objects derived from the auditory scene according to perceptual grouping principles. PMID- 10702370 TI - Multiple facets of sialomucin complex/MUC4, a membrane mucin and erbb2 ligand, in tumors and tissues (Y2K update). AB - Sialomucin complex (SMC, MUC4) is a high Mr glycoprotein heterodimer, composed of mucin (ASGP-1) and transmembrane (ASGP-2) subunits. ASGP-2 contains two EGF-like domains and acts as an intramembrane ligand for the receptor tyrosine kinase ErbB2. Transfection studies with SMC DNAs showed that SMC expression could markedly reduce both cell-cell and cell-matrix interactions in vitro and increase the growth of primary tumors and the formation of metastatic foci of human A375 melanoma cells as xenotransplants in nude mice, possibly through the ability to suppress apoptosis. SMC is expressed in most vulnerable epithelia as a protective agent, which is found in both membrane and soluble forms at luminal surfaces and secreted into fluids such as milk and tears. SMC appears to be constitutively expressed by most accessible epithelia, notable exceptions being the mammary gland and uterine luminal epithelium, in which it is tightly regulated during pregnancy. Down-regulation at the luminal uterine surface appears necessary for blastocyst implantation. TGF-b is a potent repressor of SMC expression in the mammary gland and uterus, though by different mechanisms. These combined results suggest that SMC has multiple functions in epithelia and is tightly regulated in those tissues where its special functions are required. PMID- 10702371 TI - Pathogenesis and treatment of HIV-1 infection: recent developments (Y2K update). AB - Human immunodeficiency virus type 1 (HIV-1) is the etiologic agent of acquired immunodeficiency syndrome (AIDS). The pathogenesis of HIV-1-induced disease is complex and characterized by the interplay of both viral and host factors, which together determine the outcome of infection. An improved understanding of the pathogenic mechanisms of AIDS, combined with recent insights into the dynamics of viral infection may provide powerful new opportunities for therapeutic intervention against this virus. PMID- 10702372 TI - Neurophysiological mechanisms of auditory selective attention in humans. AB - This chapter reviews the main data on the physiological substrates of auditory selective attention and their contribution to theoretical models of cognitive psychology. While event-related potentials, magnetoencephalography, and more recently neuroimaging techniques have provided fundamental information on the neural correlates of attention in the central cortical system, measurements of the frequency-following responses in the brainstem and evoked otoacoustic emissions at the cochlea strongly suggest attentional phenomena at the auditory periphery. We propose an adaptive filtering mechanism for selective auditory attention that can be flexibly and dynamically tuned depending on the attentional demand. PMID- 10702373 TI - The development of auditory attention in children. AB - In this paper we review the development of four components of auditory attention: arousal, orienting, selective attention and sustained attention. We focus especially on the processes responsible for the selection of specific stimuli for further processing because these are essential for learning and development. Although much work still needs to be done, there is evidence of developmental change in some of the components of attention, especially early in infancy. Later developmental improvements seem to be primarily attributable to higher cognitive processes, such as motivation, strategy development and implementation, and voluntary direction and regulation of attention. PMID- 10702374 TI - Cellular and molecular basis of beta-amyloid precursor protein metabolism. AB - In molecular neurobiology, perhaps no molecule has been as thoroughly examined as Alzheimer's beta-amyloid precursor protein (beta-APP). In the years since the cDNA encoding beta-APP was cloned, the protein has been the subject of unparalleled scrutiny on all levels. From molecular genetics and cellular biology to neuroanatomy and epidemiology, no scientific discipline has been left unexplored - and with good reason. beta-amyloid (Abeta) is the main constituent of the amyloidogenic plaques which are a primary pathological hallmark of Alzheimer's disease, and bta-APP is the protein from which Abeta is cleaved and released. Unraveling the molecular events underlying Abeta generation has been, and remains, of paramount importance to scientists in our field. In this review we will trace the progress that has been made in understanding the molecular and cellular basis of beta-APP trafficking and processing, or alternatively stated, the molecular basis for Abeta generation. Imperative to a complete understanding of Abeta generation is the delineation of its subcellular localization and the identification of proteins that play either direct or accessory roles in Abeta generation. We will focus on the regulation of beta-APP cleavage through diverse signal transduction mechanisms and discuss possible points of therapeutic intercession in what has been postulated to be a seminal molecular step in the cascade of events terminating in the onset of dementia, loss of neurons, and eventual death from Alzheimer's disease. PMID- 10702375 TI - The control of mitosis. AB - A precise coordination of multiple cell cycle events is required to ensure proper mitosis. Chromosome cohesion must be maintained until all chromosomes are attached to opposite poles of the mitotic spindle and aligned at the metaphase plate. At the onset of anaphase, the activity of separins contributes to the release of cohesins from chromosomes, allowing for the segregation of bivalents to opposite spindle poles. Separin activity is blocked by binding to a class of proteins known as securins, whose turnover at the metaphase-to-anaphase transition is triggered by the Anaphase Promoting Complex or cyclosome. The mitotic spindle cell cycle checkpoint coordinates the timing of these events and acts as input mechanism for DNA damage/stress pathways. Failure of this precise network leads to genomic instability and/or cell death. PMID- 10702376 TI - Lipoperoxidation damage of spermatozoa polyunsaturated fatty acids (PUFA): scavenger mechanisms and possible scavenger therapies. AB - The lipid metabolism in sperm cells is important both as one of the main sources for energy production and for cell structure. The double leaflets of the membrane should be considered not simply as a passive lipid film, but as a very specialized structure. The complete maturation of the sperm cell membrane is attained after testicular lipid biosynthetic processes and after passage through the epididymis. A special composition of membrane phospholipids, rich in polyunsaturated fatty acids (PUFA), and the different composition of sperm and immature germ cell membrane are described and discussed. Testis germ cells as well as epididymal maturing spermatozoa are endowed with enzymatic and non enzymatic scavenger systems to prevent lipoperoxidative damage. Catalase, superoxide dismutase and GSH-dependent oxidoreductases are present in variable amounts in the different developmental stages. Phospholipid hydroperoxide GSH peroxidase (PHGPx) activity and alpha tochopherol of epididymal spermatozoa are considered in detail. Their distribution and roles in caput and cauda epididymal sperm cells are discussed. Seminal plasma also has a highly specialized scavenger system that defends the sperm membrane against lipoperoxidation and the degree of PUFA insaturation acts to achieve the same goal. Systemic predisposition and a number of pathologies can lead to an anti-oxidant/pro-oxidant disequilibrium. Scavengers, such as GSH, can be used to treat these cases as they can restore the physiological constitution of PUFA in the cell membrane. The results of GSH therapy are presented and discussed. PMID- 10702377 TI - Centripetal versus centrifugal bias in visual line bisection: focusing attention on two hypotheses. AB - A variety of stimulus factors have been shown to influence the degree of leftward displacement of perceived line midpoint (i.e., pseudoneglect), which typifies the performance of normal subjects in line bisection tasks [M.E. McCourt & G. Jewell: Neuropsychologia 37, 843-855 (1999); G. Jewell & M.E. McCourt: Neuropsychologia 38, 93-110 (2000)]. One such factor is the position of lines within the visual field, where two conflicting patterns of bisection error have been reported. Some authors report a centrifugal pattern of error, where perceived line midpoint shifts away from the vertical midline, regardless of line position, i.e., relatively leftward for leftward displaced lines and vice versa. Others have reported a centripetal pattern of bisection error, where perceived line midpoint is always displaced centrally, toward the vertical midline, regardless of line position. There is no satisfactory explanation for these discrepant findings. An experiment using a tachistoscopic forced-choice line bisection protocol is described which discloses that neurologically normal right-handed subjects (N=82) typically display a centrifugal pattern of bisection error when lines are azimuthally displaced over a relatively small range, whereas a centripetal pattern is observed when lines are displaced over a wider range. Results from ancillary control experiments, in which eye position was measured during testing, confirm that systematic differences in gaze direction do not occur as a function of line position, and thus cannot account for the different patterns of bisection error. We conclude that stimulus context significantly modulates the strategy with which observers deploy spatial attention. When line position is constant, or varies over a narrow range, observers hold attention steady and widen its aperture to accommodate the relevant range of spatial location. Centrifugal bisection error is thus produced by the asymmetric cueing effect of laterally displaced lines, according to the activation-orientation theory [M. Kinsbourne: Acta Psychologica 33, 193-201 (1970)]. When the range of line position exceeds the aperture of focal attention, we hypothesize that observers adopt a strategy in which attention is dynamically scanned in the direction of azimuthally displaced lines. The effects of attentional scanning on line bisection performance are quite robust. The centripetal scanning proposed to occur for widely displaced lines is consistent with the centripetal pattern of bisection error in this condition. PMID- 10702378 TI - Visual search: bottom-up or top-down? AB - The aim of the experiments in this paper was to explore the relationship between top-down and bottom-up processes in visual search. Employing behavioral techniques, we first consider the possible role of the magnocellular visual pathway in visual search, and find that visual search does not necessarily depend on processing by this visual sub-system. We next use functional imaging (positron emission tomography) to explore the effect of varying top-down strategy during visual search. Our findings indicate that the neural processes underlying visual search are distributed over an extensive network of brain regions, with varying roles for different parts of the network as the dynamics of top-down vs. bottom up influences shift. The conjunction of bottom-up processing with top-down attentional suppression of an irrelevant singleton could account for activity found in right primary visual cortex (V1). The conjunction of bottom-up processing with top-down attentional set could explain activity noted in the right superior temporal gyrus/insular cortex. The left lateral cerebellum appears to play a role in attention, either in signaling popout or in switching attention repeatedly between multiple visual attributes. Loci in left parietal cortex (parietal operculum/superior temporal gyrus, parieto-occipital fissure and precuneus) are implicated in attention-demanding search for a target shape. Returning to behavioral experiments, we find that, when multiple feature singletons compete for attention, interference between them is strongest for features closely related to the distinguishing target feature. This competition appears to be feature-level rather than object-level, and is characterized by a varying degree of specificity for different features. Task complexity modulates interference effects, even for abrupt visual onsets, which are often considered to capture attention involuntarily. Overall, our observations converge on the conclusion that visual search is extremely flexible and subject to considerable specificity of top-down control, although such specificity is clearly not absolute. PMID- 10702379 TI - Tyrosine kinase expression is increased in papillary thyroid carcinoma of children and young adults. AB - Tyrosine kinases (TKs) are important candidate genes for malignant transformation and at least 21 different TKs have been identified in the thyroid gland. We hypothesized that the collective activity of these TKs might be increased in thyroid carcinoma and have association with the clinical behavior of individual tumors. To test this, we determined TK expression by immunohistochemistry in 74 archival thyroid tissue blocks (48 papillary thyroid carcinoma, PTC; 9 follicular thyroid carcinoma, FTC; 17 benign thyroid diseases) from children and young adults. Mean TK expression was greater for PTC (2.1 +/- 0.11) than benign lesions (1.6 +/- 0.2, p = 0.027), and also tended to be greater in FTC (2.1 +/- 0.25, p = 0.12). Recurrence risk was three-fold greater for PTC with intense TK expression (4/15, 27%) than for PTC with minimal - moderate TK expression (3/33, 9.0%). However, this was not statistically significant (p = 0.10). In PTC, TK expression correlated with expression of the receptor for hepatocyte growth factor / scatter factor (cMET, r = 0.31, p = 0.044). In FTC, TK expression did not correlate with cMET, but tended to be greater in young patients (r = -0.59, p = 0.09). We conclude that TK expression is increased in PTC and possibly associated with an increased recurrence risk. PMID- 10702380 TI - Cell cycle control of pancreatic beta cell proliferation. AB - Diabetes mellitus ensues as a consequence of the body's inability to respond normally to high blood glucose levels. The onset of diabetes is due to several pathological changes, which are a reflection of either the inability of the pancreatic beta cells to secrete sufficient insulin to combat the hyperglycemia or a state of insulin resistance in target tissues. However, the significance of changes in beta cell mass and decreased beta cell proliferation or growth in progression of diabetes has been under-appreciated. Beta cells, like all other cells of our body are under the regulatory checks and balances enforced by changes in cell cycle progression. However, very little is known regarding the key components of the cell cycle machinery regulating cell cycle control of beta cells. Knowledge of key elements involved in cell cycle regulation of beta cells will go a long way in improving our understanding of the replication capacity and developmental biology of beta cells. This information is essential for us to design new approaches that can be used to correct beta cell deficiency in diabetes. This review focuses on the current knowledge of factors important for proliferation of beta cells and proposes a cell cycle model for regeneration of the beta cell population lost or reduced in diabetes. PMID- 10702381 TI - Vaccination against malaria: targets, strategies and potentiation of immunity to blood stage parasites. AB - Malaria is the world's major parasitic disease, for which effective control measures are urgently needed. One of the difficulties hindering successful vaccine design against Plasmodium is an incomplete knowledge of antigens eliciting protective immunity, the precise types of immune response for which to aim, and how these can be induced. A greater appreciation of the mechanisms of protective immunity, on the one hand, and of immunopathology, on the other, should provide critical clues to how manipulation of the immune system may best be achieved. This review discusses the current state of malaria vaccine development and research to understand the factors involved in the modulation of vaccine-potentiated immunity to the pathogenic blood-borne stages of the parasite. PMID- 10702382 TI - Theoretical and experimental approaches to design effective antisense oligonucleotides. AB - Among the large number of possible antisense oligonucleotides (asODN) against a given target nucleic acid, only a small number of species seems to give rise to satisfactorily strong inhibition of target gene expression in living cells. Therefore much attention is paid to strategies that help to successfully design effective asODN. Here, selected experimental approaches and theoretical concepts will be briefly described that have been developed to increase the probability of success in the use of asODN. Advantages and disadvantages of these strategies will be compared and the relatively new and controversially discussed concept of a theoretical and computer-supported design of effective asODN will be addressed. PMID- 10702383 TI - Genetic alterations in adult diffuse glioma: occurrence, significance, and prognostic implications. AB - Our understanding of diffuse glioma development and progression has expanded remarkably over the past decade. As the genetic alterations responsible for these tumors are identified, molecular models of glioma pathogenesis are emerging and hold great promise to explain the biologic mechanisms of these neoplasms. Although these models continue to evolve and remain highly simplified, some of the genetic alterations that they encompass appear to be prognostically useful. Among the astrocytic gliomas, age and tumor grade are the most powerful indicators of patient survival, however, a wide range of variability remains, particularly among the low- grade and anaplastic astrocytomas. Recent reports indicate that alterations of the PTEN tumor suppressor gene are independent predictors of overall survival for anaplastic astrocytoma patients, helping to distinguish the cases with behavior resembling their more malignant counterparts, the glioblastomas. Among the oligodendroglial tumors, alterations of the 1p and 19q chromosome arms have emerged as potentially powerful predictors of overall patient survival and in vivo chemotherapeutic response, while alterations of the p16/CDKN2A tumor suppressor gene suggest shorter overall survival. As our molecular models continue to improve, through functional analyses and the identification of additional genetic contributors, we will expand our capacity to more effectively prognose these patients and to design rational therapeutic strategies. PMID- 10702384 TI - Role of pRB dephosphorylation in cell cycle regulation. AB - pRB, the tumor suppressor product of the retinoblastoma susceptibility gene, is regarded as one of the key regulators of the cell cycle. This protein exerts its growth suppressive effect through its ability to bind and interact with a variety of cellular proteins. In turn, pRB binding and interacting ability is governed by its phosphorylation state. In recent years, this negative growth regulatory protein has captured a great deal of attention from investigators around the world due to its ability to modulate the activity of transcription regulatory proteins, enzymes which modify chromatin, and other cellular proteins which contribute to its complex role in mammalian cells. Hypophosphorylated pRB binds and sequesters transcription factors, most notably those of the E2F/DP family, inhibiting the transcription of genes required to traverse the G1 to S phase boundary. This cell cycle inhibitory function is abrogated when pRB undergoes phosphorylation mediated by cyclin/cdk complexes following cell stimulation by mitogens. Removal of these phosphates appears to be carried out by a multimeric complex of protein phosphatase type 1 (PP1) and noncatalytic regulatory subunits at the completion of mitosis. This dephosphorylation returns pRB to its active, growth suppressive state. While the mechanism of pRB phosphorylation has and continues to be extensively studied, dephosphorylation of pRB has received disproportionately less attention. The goal of this review is to revisit the role of pRB dephosphorylation in regulating the cell cycle. Emphasis will be placed on understanding the function and regulation of pRB during the cell cycle as well as our ever-expanding notions of pRB-PP1 interaction and the mechanism of pRB dephosphorylation at mitotic exit. PMID- 10702385 TI - Bacterial resistance to aminoglycosides and beta-lactams: the Tn1331 transposon paradigm. AB - Aminoglycosides (Ags) are a group of antibiotics that exert their bactericidal activity primarily by inhibition of protein synthesis. Aminoglycoside (Ag) molecules bind to the bacterial 30S ribosomal subunit rendering the ribosomes unavailable for translation, which results in cell death. Although these antibiotics are and have been very useful to treat a variety of bacterial infections, in recent years the number of Ag resistant and multiresistant isolates has seriously increased. Mechanisms of resistance to Ag include enzymatic inactivation by acetyltransferases, nucleotidyltransferases (adenylyltransferases), and phosphotransferases, ribosomal alterations, and reduced permeability. Of all Ags, amikacin (Ak) is the most resistant to the action of Ag-modifying enzymes. However, AAC(6')-I type enzymes (a group of 6'-N acetyltransferases) can utilize Ak as substrate and confer resistance to this antibiotic in addition to other Ags. The gene aac(6')-Ib was found in various bacterial species and various research groups performed mutagenesis studies on this or related enzymes. In one case, aac(6')-Ib was identified in a transposable element, Tn1331, included in pJHCMW1, a plasmid isolated from a clinical K. pneumoniae strain. Tn1331 includes genes encoding two Ag-modifying enzymes (aac(6')-Ib and ant(3")-Ia) and two beta-lactamases (blaTEM and blaOXA-9). Characterization of other functions of the pJHCMW1 plasmid showed the presence of an RNA-regulated replication origin and a functional oriT. Stability by multimer resolution is achieved by the Tn1331 resolvase. PMID- 10702386 TI - Human T cell lymphotropic virus type I genomic expression and impact on intracellular signaling pathways during neurodegenerative disease and leukemia. AB - HTLV-I has been identified as the etiologic agent of neoplasia within the human peripheral blood T lymphocyte population, and a progressive neurologic disorder based primarily within the central nervous system. We have examined the role of HTLV-I in these two distinctly different clinical syndromes by examining the life cycle of the virus, with emphasis on the regulation of viral gene expression within relevant target cell populations. In particular, we have examined the impact of specific viral gene products, particularly Tax, on cellular metabolic function. Tax is a highly promiscuous and pleiotropic viral oncoprotein, and is the most important factor contributing to the initial stages of viral-mediated transformation of T cells after HTLV-I infection. Tax, which weakly binds to Tax response element 1 (TRE-1) in the viral long terminal repeat (LTR), can dramatically trans-activate viral gene expression by interacting with cellular transcription factors, such as activated transcription factors and cyclic AMP response element binding proteins (ATF/CREB), CREB binding protein (CBP/p300), and factors involved with the basic transcription apparatus. At the same time, Tax alters cellular gene expression by directly or indirectly interacting with a variety of cellular transcription factors, cell cycle control elements, and cellular signal transduction molecules ultimately resulting in dysregulated cell proliferation. The mechanisms associated with HTLV-I infection, leading to tropical spastic paraparesis (TSP) are not as clearly resolved. Possible explanations of viral-induced neurologic disease range from central nervous system (CNS) damage caused by direct viral invasion of the CNS to bystander CNS damage caused by the immune response to HTLV-I infection. It is interesting to note that it is very rare for an HTLV-I infected individual to develop both adult T cell leukemia (ATL) and TSP in his/her life time, suggesting that the mechanisms governing development of these two diseases are mutually exclusive. PMID- 10702387 TI - Hereditary hemorrhagic telangiectasia: A model for blood vessel growth and enlargement. PMID- 10702388 TI - Differential viral protein expression in Kaposi's sarcoma-associated herpesvirus infected diseases: Kaposi's sarcoma, primary effusion lymphoma, and multicentric Castleman's disease. AB - Kaposi's sarcoma (KS)-associated herpesvirus (KSHV) is linked to KS, primary effusion lymphomas (PEL), and a subset of multicentric Castleman's disease (MCD). Transcript mapping studies using PEL cell lines have allowed preliminary classification of viral gene expression into constitutive (class I) and inducible (class II/III) categories. To determine whether viral gene expression differs in vivo, we examined tissue sections of KSHV-infected disorders, using specific antibodies against proteins that are representative of the different expression classes of KSHV genes. ORF73/LANA appears to be a surrogate marker for KSHV infection because it is constitutively expressed in vitro and in vivo in all KSHV infected cells. Expression of vIRF1, vIL6, and PF-8 proteins in the infected B cells of MCD lymph nodes reproduces the expression pattern observed in TPA stimulated KSHV-infected B-cell lines. In contrast, the protein expression of the inducible viral genes that we tested in KS and PEL biopsies is restricted to PF-8 and vIL6, respectively. The tightly restricted expression of KSHV proteins in vivo differs from the dysregulated expression of inducible KSHV genes in vitro and suggests that viral gene expression in KSHV-infected cell lines does not accurately reflect what occurs in diseased tissues. These differences may be related to either cell-specific or immune restriction of viral replication. PMID- 10702390 TI - Gene expression of osteoprotegerin ligand, osteoprotegerin, and receptor activator of NF-kappaB in giant cell tumor of bone: possible involvement in tumor cell-induced osteoclast-like cell formation. AB - Giant cell tumor of bone (GCT) is a rare primary osteolytic tumor of bone that is characterized by massive tissue destruction at the epiphysis of long bones. There is no evidence that tumor cells themselves are capable of bone destruction; instead, it appears that the tumor cells of GCT act by promoting osteoclastogenesis and, as a consequence, osteoclastic bone resorption. However, the mechanism by which this is achieved is not understood. Here we attempted to determine whether osteoprotegerin ligand (OPGL), the factor that is necessary and essential for osteoclastogenesis, is involved in tumor cell-recruited osteoclast like giant cell formation in GCT. Using fluorescence in situ hybridization, we sought to determine mRNA expression of OPGL, its receptor RANK, and its decoy receptor OPG in three major cell types of GCT. We demonstrated that OPG mRNA was expressed in all three cell types of GCT, OPGL transcripts were mainly detected in spindle-shaped stromal-like tumor cells, whereas RANK was expressed only in macrophage-like mononuclear cells and multinuclear osteoclast-like giant cells. By semiquantitative RT-PCR, we also showed that the level of OPGL mRNA in GCT is much higher than that in normal bone and osteogenic osteosarcoma. In contrast, a similar level of OPG transcripts was detected in these three kinds of tissues, and RANK mRNA was detectable only in GCT tissues. We have further examined the regulation of gene expression of OPGL and OPG in tumor cells in response to osteotropic hormones. Administration of 1,25(OH)(2)D(3) and dexamethasone resulted in maximum up-regulation of OPGL level and down-regulation of OPG level in cultured GCT stromal-like tumor cells and the mouse bone marrow-derived ST-2 stromal cell line. Furthermore, we have shown that tumor cells of GCT induce differentiation of RANK-expressing myeloid RAW(264.7) cells into osteoclast-like cells in the presence of 1,25(OH)(2)D(3) and dexamethasone. Our findings suggest that OPGL is involved in the tumor cell-induced osteoclast-like cell formation in GCT. The ratio of OPGL/OPG by tumor cells may contribute to the degree of osteoclastogenesis and bone resorption. PMID- 10702389 TI - Genetic alterations of the retinoblastoma-related gene RB2/p130 identify different pathogenetic mechanisms in and among Burkitt's lymphoma subtypes. AB - Alterations of cell cycle-associated genes probably contribute to the pathogenesis of Burkitt's Lymphoma (BL), in addition to c-myc translocation. Mutations disrupting the nuclear localization signal of the retinoblastoma related gene RB2/p130 have been documented recently in BL cell lines and primary tumors. Given the importance of the RB2/p130 gene in controlling cell growth, mutations of this gene may result in uncontrolled cell proliferation. We tested the expression and genomic organization of the RB2/p130 gene in relation to the proliferative features of a series of BL samples collected from the endemic and sporadic regions, regardless of whether the samples were acquired immune deficiency syndrome (AIDS)-related. The expression of the Rb2/p130, p107, and cell proliferation-related proteins (cyclin A and B) was determined by immunohistochemistry. The structures of exons 19 through 22 of the RB2/p130 gene, encoding for the B domain and C terminus, were analyzed by polymerase chain reaction (PCR) analysis and single-strand conformation polymorphism (SSCP) technique. The direct PCR products were sequenced to identify the actual mutations. Our results suggest that BL is composed of a mixture of molecular types with distinct genetic and phenotypic patterns, probably resulting from different pathogenetic mechanisms. In endemic BL, the RB2/p130 gene is mutated in most of the cases, and the protein is restricted to the cytoplasm. In AIDS related BL, high levels of nuclear expression of the wild-type pRb2/p130, p107, and cell proliferation-related proteins were detected. This finding is in line with the molecular mechanisms observed in virus-linked oncogenesis. Sporadic BLs were mainly characterized by the low nuclear values of the wild-type pRb2/p130 and, conversely, the high values of p107. The increased cell proliferation due to different alterations of cell growth control by Rb-related proteins may be the first step in lymphomagenesis, during which additional genetic changes, including missense mutations of c-myc, may subsequently occur. PMID- 10702391 TI - Activated leukocyte cell adhesion molecule/CD166, a marker of tumor progression in primary malignant melanoma of the skin. AB - Expression of activated leukocyte cell adhesion molecule (ALCAM)/CD166 correlates with the aggregation and metastatic capacity of human melanoma cell lines (Am J Pathol 1998, 152:805-813). Immunohistochemistry on a series of human melanocytic lesions reveals that ALCAM expression correlates with melanoma progression. Most nevi (34/38) and all thin melanomas studied (Clark levels I and II) did not express ALCAM. In contrast, immunoreactivity was detected in the invasive, vertical growth phase of 2 of the 13 Clark level III lesions tested. The fraction of positive lesions further increased in Clark level IV (13/19) and in Clark level V (4/4) lesions. ALCAM expression was exclusively detectable in the vertical growth phase of the primary tumor. In melanoma metastases, approximately half of the lesions tested (13/28) were ALCAM positive. According to the Breslow thickness, ALCAM expression was observed in less than 10% of the lesions that were thinner than 1.5 mm and in over 70% of the lesions that were thicker than 1.5 mm. Our results strongly suggest that ALCAM plays an important role in melanocytic tumor progression and depict it as a new molecular marker for neoplastic progression of primary human melanoma. PMID- 10702392 TI - Neurokinin-1 (NK-1) receptor is required in antigen-induced cystitis. AB - Interstitial cystitis (IC) is a debilitating disease that has been adversely affecting the quality of women's lives for many years. The trigger in IC is not entirely known, and a role for the sensory nerves in its pathogenesis has been suggested. In addition to inflammation, increased mast cell numbers in the detrusor muscle have been reported in a subset of IC patients. Experimentally, several lines of evidence support a central role for substance P and neurokinin-1 (NK-1) receptors in cystitis. The availability of mice genetically deficient in neurokinin-1 receptor (NK-1R(-/-)) allows us to directly evaluate the importance of substance P in cystitis. An unexpected finding of this investigation is that NK-1R(-/-) mice present increased numbers of mast cells in the bladder when compared with wild-type control mice. Despite the increase in mast cell numbers, no concomitant inflammation was observed. In addition, bladder instillation of wild-type mice with a sensitizing antigen induces activation of mast cells and an acute inflammatory response characterized by plasma extravasation, edema, and migration of neutrophils. Antigen-sensitized NK-1R(-/-) mice also exhibit bladder mast cell degranulation in response to antigen challenge. However, NK-1R(-/-) mice are protected from inflammation, failing to present bladder inflammatory cell infiltrate or edema in response to antigen challenge. This work presents the first evidence of participation of NK-1 receptors in cystitis and a mandatory participation of these receptors on the chain of events linking mast cell degranulation and inflammation. PMID- 10702393 TI - ATIC-ALK: A novel variant ALK gene fusion in anaplastic large cell lymphoma resulting from the recurrent cryptic chromosomal inversion, inv(2)(p23q35). AB - The subset of CD30-positive anaplastic large cell lymphomas (ALCL) with the NPM ALK gene fusion arising from the t(2;5)(p23;q35) forms a distinct clinical and prognostic entity. Recently, various cytogenetic, molecular, and protein studies have provided evidence for the existence of several types of variant ALK fusions in up to 20% of ALK+ ALCL, of which only one, a TPM3-ALK fusion resulting from a t(1;2)(q25;p23), has so far been cloned. A cryptic inv(2)(p23q35) has been described as another recurrent cytogenetic alteration involving ALK and an unidentified fusion partner in some ALCL. In a screen for variant ALK gene fusions, we identified two ALCL that were negative for NPM-ALK by reverse transcriptase-polymerase chain reaction, but were positive for cytoplasmic ALK with both polyclonal and monoclonal antibodies to the ALK tyrosine kinase domain, consistent with ALK deregulation by an alteration other than the t(2;5) Case 1 was a T-lineage nodal and cutaneous ALCL in a 52-year-old woman, and Case 2 was a T-lineage nodal ALCL in a 12-year-old girl. FISH analysis confirmed ALK rearrangement in both cases. An inverse polymerase chain reaction approach was then used to identify the ALK translocation partner in Case 1. We found an in frame fusion of ALK to ATIC, a gene previously mapped to 2q34-q35. We then confirmed by DNA polymerase chain reaction the localization of ATIC to yeast artificial chromosome (YAC) 914E7 previously reported to span the 2q35 break in the inv(2)(p23q35). FISH analysis in Case 1 confirmed rearrangement of YAC 914E7 and fusion to ALK. The ATIC-ALK fusion was confirmed in Case 1 and also identified in Case 2 by conventional reverse transcriptase-polymerase chain reaction using ATIC forward and ALK reverse primers. ATIC encodes an enzyme involved in purine biosynthesis which, like other fusion partners of ALK, is constitutively expressed and appears to contain a dimerization domain. ATIC-ALK fusion resulting from the inv(2)(p23q35) thus provides a third mechanism of ALK activation in ALK+ ALCL. PMID- 10702394 TI - KIT extracellular and kinase domain mutations in gastrointestinal stromal tumors. AB - Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal neoplasms arising in the gastrointestinal tract. GISTs express the KIT receptor tyrosine kinase, and many cases have activating mutations in the KIT juxtamembrane region. We now report an analysis of KIT cDNA and genomic sequences in eight GISTs that lack juxtamembrane region mutations. Six cases contained heterozygous exon 9 mutations in which six nucleotides, encoding Ala-Tyr, were duplicated. The other two cases contained homozygous exon 13 missense mutations, resulting in substitution of Glu for Lys(642), that were associated with constitutive KIT tyrosine phosphorylation. Sequence analysis of DNAs from nonneoplastic companion tissues revealed that both the exon 9 and exon 13 mutations were somatic. These are the first descriptions, in any tumor, of mutations in KIT exons encoding the C-terminal end of the extracellular domain and the first part of the split kinase domain. These findings indicate that KIT may be activated by mutations in at least three domains-extracellular, juxtamembrane, and kinase-in GISTs. PMID- 10702395 TI - Elevated abeta42 in skeletal muscle of Alzheimer disease patients suggests peripheral alterations of AbetaPP metabolism. AB - The levels of amyloid-beta40 (Abeta40) and Abeta42 peptides were quantified in temporalis muscles and brain of neuropathologically diagnosed Alzheimer disease (AD) and of nondemented individuals. This was achieved by using a novel analytical approach consisting of a combination of fast-performance liquid chromatographic (FPLC) size exclusion chromatography developed under denaturing conditions and europium immunoassay on the 4.0- to 4.5-kd fractions. In the temporalis muscles of the AD and nondemented control groups, the average values for Abeta42 were 15.7 ng/g and 10.2 ng/g (P = 0.010), and for Abeta40 they were 37.8 ng/g and 29.8 ng/g (P = 0.067), respectively. Multiple regression analyses of the AD and control combined populations indicated that 1) muscle Abeta40 and muscle Abeta42 levels were correlated with each other (P < 0.001), 2) muscle Abeta40 levels were positively correlated with age (P = 0. 036), and 3) muscle Abeta42 levels were positively correlated with Braak stage (P = 0.042). Other forms of the Abeta peptide were discovered by mass spectrometry, revealing the presence of Abeta starting at residues 1, 6, 7, 9, 10, and 11 and ending at residues 40, 42, 44, 45, and 46. It is possible that in AD the skeletal muscle may contribute to the elevated plasma pool of Abeta and thus indirectly to the amyloid deposits of the brain parenchyma and cerebral blood vessels. The increased levels of Abeta in the temporalis muscles of AD patients suggest that alterations in AbetaPP and Abeta metabolism may be manifested in peripheral tissues. PMID- 10702396 TI - API2-MALT1 chimeric transcripts involved in mucosa-associated lymphoid tissue type lymphoma predict heterogeneous products. AB - Recent progress in molecular analysis of low-grade B cell lymphoma has revealed that API2 at 11q21 and a novel gene, MALT1 at 18q21, are involved in t(11;18)(q21;q21), a characteristic chromosome aberration for mucosa-associated lymphoid tissue (MALT) type lymphoma. We describe here the establishment of a reverse transcription-polymerase chain reaction (RT-PCR) assay that we used to analyze 22 cases of MALT lymphoma. All five cases that were shown to possess t(11;18)(q21;q21) showed the specific amplification of API2-MALT1 chimeric transcripts. Of the remaining 17 cases for which cytogenetic data were not available, three cases demonstrated the presence of fusion transcripts, indicating that a significant percentage of MALT lymphoma cases of the present series appeared to possess t(11;18). A single fragment was observed in each of these cases, but the size varied from case to case. Sequencing analysis revealed that there are two breakpoints in API2 and three in MALT1, and that all of the fusion transcripts are in-frame. On the basis of these results, four kinds of chimeric proteins can be predicted for the present series. Thus, the RT-PCR assay used here should serve as an effective molecular tool for understanding molecular pathogenesis and the clinical significance of API2-MALT1 for MALT lymphomas. PMID- 10702397 TI - Telomere shortening is an in vivo marker of myocyte replication and aging. AB - To determine whether adult cardiac myocytes are capable of multiple divisions and whether this form of growth is restricted to a subpopulation of cells that retain this capacity with age, telomere lengths were measured in myocyte nuclei isolated from the left ventricle of fetal and neonatal Fischer 344 rats and rats at 4, 12, and 27 months after birth. Two independent methodologies were used for this analysis: laser scanning cytometer and confocal microscopy. In each case, fluorescence intensity of a peptide nucleic acid probe specific for telomeric sequence was evaluated. The two techniques yielded comparable results. Telomeric shortening increased with age in a subgroup of myocytes that constituted 16% of the entire cell population. In the remaining nondividing cells, progressive accumulation of a senescent associated nuclear protein, p16(INK4), was evidenced. In conclusion, a significant fraction of myocytes divides repeatedly from birth to senescence, counteracting the continuous death of cells in the aging mammalian rat heart. PMID- 10702398 TI - Overexpression of the hepatocyte growth factor (HGF) receptor (Met) and presence of a truncated and activated intracellular HGF receptor fragment in locally aggressive/malignant human musculoskeletal tumors. AB - Enhanced hepatocyte growth factor (HGF) receptor (Met) signaling has been suggested to play an important role in the development and progression of various epithelial and nonepithelial tumors. N-terminally truncated forms of the HGF receptor have been shown to be constitutively activated and tumorigenic in animal experiments. In the present study, 102 benign and malignant human musculoskeletal tumors were examined for expression of the HGF receptor by Western blotting and/or immunohistochemistry. A clear predominance of HGF receptor expression was seen in malignant as compared to benign tumors (Western blotting, P < 0.001; immunohistochemistry, P < 0.02). For the first time we show HGF receptor expression in the following four tumor types: dermatofibrosarcoma protuberans, clear cell sarcoma of tendons, malignant primitive neuroectodermal tumor, and benign fibrous histiocytoma. In three cases of sarcoma with high HGF receptor expression by Western blotting, we found indications of a short 85-kd N terminally truncated HGF receptor that was tyrosine phosphorylated and located in the cytoplasm. Although fragments of this length were seen in 18 of 65 tumors, most were not tyrosine-phosphorylated. Northern blotting revealed only the 7.5-kb full-length HGF receptor transcript, suggesting that the 85-kd fragment is generated by an alternative initiation of translation or by proteolytic cleavage. Southern blotting detected no amplification of the Hgfr/Met gene in the 35 tumors examined, in contrast to our recent report of Hgfr/Met gene amplification in 7, 12-dimethylbenz(a)anthracene (DMBA)-induced rat sarcomas. The present data suggest that the locally aggressive and malignant properties of human mesenchymal tumors maybe related, in part, to high levels of full-length HGF receptors, and in some cases to the occurrence of N-terminally truncated HGF receptors, activated independently of HGF. PMID- 10702399 TI - Papillary carcinoma of the thyroid: hepatocyte growth factor (HGF) stimulates tumor cells to release chemokines active in recruiting dendritic cells. AB - Tissue distribution of dendritic cells was investigated in eight cases of papillary carcinoma of the thyroid using immunohistochemistry. Most dendritic cells had an immature phenotype (CD1a++, CD11c+, CD40+, CD86-, HLA-DR-) and were located at the invasion edge of the tumor. This pattern of distribution was profoundly different from that of CD68+ macrophages, which were evenly distributed throughout the tumor. The ability of tumor cells to release chemotactic factors active on dendritic cells was investigated in primary cultures of the same cases of papillary carcinoma, and was compared to that of the corresponding normal thyroid cells obtained from the tumor-free contralateral lobe. Chemotactic activity of culture supernatants was tested against dendritic cells in a chemotaxis chamber. It was found that papillary carcinoma cells were active in releasing chemotactic activity, that hepatocyte growth factor (HGF; 100 ng/ml) or interleukin (IL)-1beta (10(3) U/ml) induced a fourfold increase in the amount of chemotactic activity released, and that normal thyroid cells obtained from the same patients were as effective as tumor cells. Characterization of chemokines at RNA level revealed that unstimulated cells contain large amounts of IL-8 and monocyte chemotactic protein (MCP)-1 RNAs, and that stimulation with HGF or IL-1beta induced RNAs for regulated upon activation normal T expressed and secreted (RANTES), macrophage inflammatory protein (MIP)-3alpha, interferon-gamma inducible protein 10 (IP-10), and, to a lesser extent, MIP-1alpha and MIP-1beta. The possibility that HGF/Met interaction has a biological role in vivo was investigated in serial sections of six tumors immunostained for CD1a+, Met protein, and HGF. It was found that all six tumors were intensely and diffusely positive for Met protein, that HGF staining was present in tumor cells of the advancing edge, and that HGF+/Met+ tumor cell nests were infiltrated by CD1a+ dendritic cells. The foregoing observations are consistent with the possibility that HGF stimulation of Met+ tumor cells is one of the molecular mechanisms involved in the recruitment of dendritic cells. PMID- 10702400 TI - Amplification and deletion of topoisomerase IIalpha associate with ErbB-2 amplification and affect sensitivity to topoisomerase II inhibitor doxorubicin in breast cancer. AB - Topoisomerase IIalpha (topoIIalpha) is a key enzyme in DNA replication and a molecular target for many anti-cancer drugs called topoII inhibitors. The topoIIalpha gene is located at chromosome band 17q12-q21, close to the ErbB-2 oncogene (HER-2/neu), which is the most commonly amplified oncogene in breast cancer. Because of the physical proximity to ErbB-2, copy number aberrations may also occur in the topoIIalpha gene. These topoIIalpha gene copy number aberrations may be related to the altered chemosensitivity to topoII inhibitors that breast cancers with ErbB-2 amplification are known to have. We used fluorescence in situ hybridization to study copy number aberrations of both topoIIalpha and ErbB-2 in nine breast cancer cell lines and in 97 clinical breast tumors, which were selected for the study according to their ErbB-2 status by Southern blotting. TopoIIalpha-protein expression was studied with Western blot and sensitivity to doxorubicin (a topoII inhibitor) with a 96-well clonogenic in vitro assay. Two of the five cell lines with ErbB-2 gene amplification (SK-BR-3 and UACC-812) showed amplification of topoIIalpha. In MDA-361 cells, ErbB-2 amplification (14 copies/cell) was associated with a physical deletion of topoIIalpha (four copies of chromosome 17 centromere and two copies of topoIIalpha). The topoIIalpha amplification in UACC-812 cells was associated with 5.9-fold-increased topoIIalpha protein expression and 2.5-fold-increased sensitivity to the topoII inhibitor, doxorubicin, whereas the deletion in MDA-361 leads to decreased protein expression (45% of control) and a 2.4-fold-increased chemoresistance in vitro. Of 57 ErbB-2-amplified primary breast carcinomas, 25 (44%) showed ErbB-2-topoIIalpha coamplification and 24 (42%) showed a physical deletion of the topoIIalpha gene. No topoIIalpha copy number aberrations were found in 40 primary tumors without ErbB-2 amplification. TopoIIalpha gene amplification and deletion are common in ErbB-2-amplified breast cancer and are associated with increased or decreased sensitivity to topoII inhibitors in vitro, respectively. These findings may explain the altered chemosensitivity to topoII inhibitors reported in ErbB-2-amplified breast cancers. PMID- 10702401 TI - Association of EWS-FLI1 type 1 fusion with lower proliferative rate in Ewing's sarcoma. AB - The Ewing's sarcoma (ES) family of tumors, including peripheral neuroectodermal tumor (PNET), is defined genetically by specific chromosomal translocations resulting in fusion of the EWS gene with a member of the ETS family of transcription factors, either FLI1 (90-95%) or ERG (5-10%). A second level of molecular genetic heterogeneity stems from the variation in the location of the translocation breakpoints, resulting in the inclusion of different combinations of exons from EWS and FLI1 (or ERG) in the fusion products. The most common type of EWS-FLI1 fusion transcript, type 1, is associated with a favorable prognosis and appears to encode a functionally weaker transactivator, compared to other fusion types. We sought to determine whether the observed covariation of structure, function, and clinical course correlates with tumor cell kinetic parameters such as proliferative rate and apoptosis, and with expression of the receptor for insulin-like growth factor I (IGF-1R). In a group of 86 ES/PNET with defined EWS-ETS fusions (45 EWS-FLI1 type 1, 27 EWS-FLI1 non-type 1, 14 EWS-ERG), we assessed proliferation rate by immunostaining for Ki-67 using MIB1 antibody (n = 85), apoptosis by TUNEL assay (n = 66), and IGF-1R expression by immunostaining with antibody 1H7 (n = 78). Ki-67 proliferative index was lower in tumors with EWS-FLI1 type 1 than those with non-type 1 EWS-FLI1, whether analyzed as a continuous (P = 0.049) or categorical (P = 0.047) variable. Logistic regression analysis suggests that this association was secondary to the association of type 1 EWS-FLI1 and lower IGF-1R expression (P = 0.04). Comparing EWS-FLI1 to EWS-ERG cases, Ki-67 proliferative index was higher in the latter (P = 0.01, Mann-Whitney test; P = 0.02, Fisher's exact test), but there was no significant difference in IGF-1R. TUNEL results showed no significant differences between groups. Our results suggest that clinical and functional differences between alternative forms of EWS-FLI1 are paralleled by differences in proliferative rate, possibly mediated by differential regulation of the IGF-1R pathway. PMID- 10702402 TI - A novel association between the human heat shock transcription factor 1 (HSF1) and prostate adenocarcinoma. AB - A search for differentially expressed genes in a pair of nonmetastatic (PC-3) versus metastatic variant (PC-3M) human prostate carcinoma cell lines led to identification of the human heat shock factor (HSF1) as an overexpressed gene product in PC-3M cells. Analysis of primary prostate cancer specimens indicated that HSF1 is generally up-regulated in most of the malignant prostate epithelial cells relative to the normal prostate cells. Among the known effectors of HSF1 action, constitutive levels of HSP70 and HSP90 are not significantly altered by the naturally elevated expression of HSF1 as in PC-3M cells or by transduced overexpression of HSF1 in PC-3 cells. The basal levels of HSP27 in both cases are, however, consistently increased by two- to threefold. With respect to response to heat shock, high basal concentration of HSP90 is not further enhanced in these cells, and HSP70 is up-regulated irrespective of HSF1 level. Heat shock, however, causes an increase in HSP27 when HSF1 is up-regulated, except when the expression of HSF1 is already too high. These results document for the first time that HSF1 is overexpressed in human prostate cancer cells, at least one consequence of which in the prostate cancer cell lines tested is stimulation of both basal and stress-induced expression of HSP27, an important factor in cell growth, differentiation, or apoptosis. PMID- 10702403 TI - Expression of nuclear beta-catenin and c-myc is correlated with tumor size but not with proliferative activity of colorectal adenomas. AB - Most colorectal cancers have loss-of-function mutations in the adenomatosis polyposis coli (APC) tumor suppressor gene. This leads to the accumulation of nuclear beta-catenin, which, together with the DNA-binding protein TCF-4, functions as a transcriptional activator. The recently defined target genes c myc, cyclin D1, and matrilysin are responsible for tumor proliferation or malignant progression and explain the oncogenic potential of nuclear beta catenin. To investigate its role in early colon carcinogenesis, we analyzed the expression of beta-catenin, its target gene c-myc, and the proliferative activity in 88 colorectal adenomas of varying size and grade of dysplasia. The results revealed i) the most significant correlation of nuclear beta-catenin and c-myc expression was not with the grade of dysplasia but with the size of the colon adenoma; ii) perfect correlation of nuclear beta-catenin and c-myc expression; iii) no significant correlation of adenoma size with the proliferative activity; and iv) no significant correlation of proliferative activity and the nuclear expression of beta-catenin and c-myc. These results imply that APC mutations have additional beta-catenin-independent functions; APC mutations alone are not sufficient for nuclear overexpression of beta-catenin; and nuclear beta-catenin has additional important functions for exceeding a threshold tumor size. PMID- 10702404 TI - DNA copy number changes in Schistosoma-associated and non-Schistosoma-associated bladder cancer. AB - DNA copy number changes were investigated in 69 samples of schistosoma-associated (SA) and non-schistosoma-associated (NSA) squamous cell carcinoma (SCC) and transitional cell carcinoma (TCC) of the bladder by comparative genomic hybridization (CGH). DNA copy number changes were detected in 47 tumors. SA tumors had more changes than NSA tumors (mean, 7 vs. 4), whereas the number of changes in SCC and TCC tumors was similar. SA tumors displayed more gains than losses (1.7:1), whereas NSA tumors showed an equal number of gains and losses. Changes that were observed at similar frequencies in SCC and TCC, irrespective of the schistosomal status, included gains and high-level amplifications at 1q, 8q, and 20q and losses in 9p and 13q. These changes may be involved in a common pathway for bladder tumor development and progression independent of schistosomal status or histological subtype. Losses in 3p and gains at 5p were seen only in SCC (P < 0.01) and losses in 5q were more frequent in SA-SCC than in other tumors (P < 0.05). However, changes that were more frequent in TCC than those in SCC included gains at 17q (P < 0.01) and losses in 4q (P < 0.05) and 6q (P < 0.01). Gains and high-level amplifications at 5p were seen only in SA-SCC (P < 0. 01), whereas gains and high-level amplifications with minimal common overlapping regions at 11q13 were more frequently seen both in SA-SCC and SA-TCC tumors (P < 0.01). In addition to the above mentioned alterations, several other changes were also seen at lower frequencies. The variations in the DNA copy number changes observed in TCC, SCC, SA, and NSA bladder carcinomas suggest that these tumors have different genetic pathways. PMID- 10702405 TI - Lack of endothelial nitric oxide synthase aggravates murine accelerated anti glomerular basement membrane glomerulonephritis. AB - Nitric oxide (NO) radicals generated by endothelial nitric oxide synthase (eNOS) are involved in the regulation of vascular tone. In addition, NO radicals derived from eNOS inhibit platelet aggregation and leukocyte adhesion to the endothelium and, thus, may have anti-inflammatory effects. To study the role of eNOS in renal inflammation, the development of accelerated anti-glomerular basement membrane (GBM) glomerulonephritis was examined in mice lacking a functional gene for eNOS and compared with wild-type (WT) C57BL/B6j mice. WT C57BL/6j mice (n = 12) and eNOS knockout (-/-) mice (n = 12) were immunized intraperitoneally with sheep IgG (0.2 mg in complete Freund's adjuvant). At day 6.5 after immunization, mice received a single i.v. injection of sheep anti-mouse GBM (1 mg in 200 microl PBS). Mice were sacrificed at day 1 and 10 after induction of the disease. All WT mice survived until day 10, whereas 1 eNOS-/- mouse died and 2 more became moribund, requiring sacrifice. At day 10, eNOS-/- mice had higher levels of blood urea nitrogen than WT mice (P < 0.02), although proteinuria was comparable. Immunofluorescence microscopy documented similar IgG deposition in both WT and eNOS-/- mice, but eNOS-/- mice had more extensive glomerular staining for fibrin at day 10 (P < 0.007). At day 10, light microscopy demonstrated that eNOS-/- mice had more severe glomerular thrombosis (P < 0.003) and influx of neutrophils (P < 0. 006), but similar degrees of overall glomerular endocapillary hypercellularity and crescent formation. In conclusion, accelerated anti-GBM glomerulonephritis is severely aggravated in eNOS-/- mice, especially with respect to glomerular capillary thrombosis and neutrophil infiltration. These results indicate that NO radicals generated by eNOS play a protective role during renal inflammation. PMID- 10702406 TI - Heparin-binding EGF-like growth factor is up-regulated in the obstructed kidney in a cell- and region-specific manner and acts to inhibit apoptosis. AB - The expression of certain growth factors in the epidermal growth factor (EGF) family is altered in response to renal injury. Recent studies have demonstrated that heparin binding EGF-like growth factor (HB-EGF) expression may be cytoprotective in response to apoptotic signals. The purpose of this study was to investigate the potential role of HB-EGF in the upper urinary tract following unilateral ureteral obstruction. We present evidence that: i) ureteral obstruction induced cell-specific but transient activation of HB-EGF gene expression; ii) HB-EGF expression in renal epithelial cells increased under conditions where mechanical deformation, such as that caused by hydronephrotic distension, induces apoptosis, but HB-EGF expression did not increase in renal pelvis smooth muscle cells under identical conditions; and iii) enforced expression of HB-EGF served to protect renal epithelial cells from stretch induced apoptosis. These results suggest a potential mechanism by which the kidney protects itself from apoptosis triggered by urinary tract obstruction. PMID- 10702407 TI - Galectin-3 induces endothelial cell morphogenesis and angiogenesis. AB - Increasing evidence suggests that carbohydrate-binding proteins play an essential role in tumor growth and metastasis. However, conflicting results on their function in the regulation of cell proliferation and differentiation during angiogenesis have been reported. We have examined the role of galectin-3 in the regulation of human umbilical vein endothelial cell proliferation, differentiation, migration, and neovascularization. Galectin-3, a carbohydrate binding protein, with specificity for type 1 and 11 ABH blood group epitopes and polylactosamine glycan containing cell surface glycoproteins, is the major nonintegrin cellular laminin-binding protein. Because galectin-3 expression was shown to be associated in some tumor systems with metastasis, we questioned whether it induces endothelial cell morphogenesis. Here we show that galectin-3 affects chemotaxis and morphology and stimulates capillary tube formation of HUV EC-C in vitro and angiogenesis in vivo. Endothelial cell morphogenesis is a carbohydrate-dependent process, as it is neutralized by specific sugars and antibodies. These findings demonstrate that endothelial cell surface carbohydrate recognition event(s) can induce a signaling cascade leading to the differentiation and angiogenesis of endothelial cells. PMID- 10702408 TI - Endoglin expression is reduced in normal vessels but still detectable in arteriovenous malformations of patients with hereditary hemorrhagic telangiectasia type 1. AB - Endoglin is predominantly expressed on endothelium and is mutated in hereditary hemorrhagic telangiectasia (HHT) type 1 (HHT1). We report the analysis of endoglin in tissues of a newborn (family 2), who died of a cerebral arteriovenous malformation (CAVM), and in a lung specimen surgically resected from a 78-year old patient (family 5), with a pulmonary AVM (PAVM). The clinically affected father of the newborn revealed a novel mutation that was absent in his parents and was identified as a duplication of exons 3 to 8, by quantitative multiplex polymerase chain reaction. The corresponding mutant protein (116-kd monomer) and the missense mutant protein (80-kd monomer) present in family 5 were detected only as transient intracellular species and were unreactive by Western blot analysis and immunostaining. Normal endoglin (90-kd monomer) was reduced by 50% on peripheral blood-activated monocytes of the HHT1 patients. When analyzed by immunostaining and densitometry, presumed normal blood vessels of the newborn lung and brain and vessels adjacent to the adult PAVM showed a 50% reduction in the endoglin/PECAM-1 ratio. A similar ratio was observed in the CAVM and PAVM, suggesting that all blood vessels of HHT1 patients express reduced endoglin in situ and that AVMs are not attributed to a focal loss of endoglin. PMID- 10702409 TI - Impaired elastogenesis in Hurler disease: dermatan sulfate accumulation linked to deficiency in elastin-binding protein and elastic fiber assembly. AB - Hurler disease resulting from a deficiency in alpha-L-iduronidase, which causes an accumulation of dermatan sulfate and heparan sulfate glycosaminoglycans, is characterized by connective tissue and skeletal deformations, cardiomyopathy, cardiac valve defects, and progressive coronary artery stenosis. In this report, we present evidence that accumulation of dermatan sulfate but not heparan sulfate moieties is linked to impaired elastic fiber assembly that, in turn, contributes substantially to the development of the clinical phenotype in Hurler disease. Our data suggest that dermatan sulfate-bearing moieties bind to and cause functional inactivation of the 67-kd elastin-binding protein, a molecular chaperone for tropoelastin, which normally facilitates its secretion and assembly into elastic fibers. We demonstrate that, in contrast to normal skin fibroblasts and cells from Sanfilippo disease, which accumulate heparan sulfate, Hurler fibroblasts show reduced expression of elastin-binding protein and do not assemble elastic fibers, despite an adequate synthesis of tropoelastin and sufficient production of a microfibrillar scaffold of elastic fibers. Because cultured Hurler fibroblasts proliferate more quickly than their normal counterparts and the addition of exogenous insoluble elastin reduces their proliferation, we suggest that cell contacts with insoluble elastin play an important role in controlling their proliferation. PMID- 10702410 TI - Semaphorin SEMA3F localization in malignant human lung and cell lines: A suggested role in cell adhesion and cell migration. AB - Semaphorins/collapsins are a family of secreted and membrane-associated proteins involved in nerve growth cone migration. However, some are expressed widely in adult tissues suggesting additional functions. SEMA3F/H.SemaIV was previously isolated from a 3p21.3 homozygous deletion region in human lung cancer. We studied SEMA3F cellular localization using our previously characterized anti SEMA3F antibody. In normal lung, SEMA3F was found in all epithelial cells at the cytoplasmic membrane and, to a lesser extent, in the cytoplasm. In lung tumors, the localization was predominantly cytoplasmic, and the levels were comparatively reduced. In non-small-cell lung carcinomas, low levels correlated with higher stage. In all tumors, an exclusive cytoplasmic localization of SEMA3F correlated with high levels of vascular endothelial growth factor and was related to the grade and aggressiveness. This suggests that vascular endothelial growth factor might compete with SEMA3F for binding to their common receptors, neuropilin-1 and -2 and might contribute to SEMA3F delocalization and deregulation in lung tumor. In parallel studies, SEMA3F distribution was examined in cell cultures by confocal microscopy. Marked staining was observed in pseudopods and in the leading edge or ruffling membranes of lamellipods or cellular protrusions in motile cells. SEMA3F was also observed at the interface of adjacent interacting cells suggesting a role in cell motility and cell adhesion. PMID- 10702411 TI - Expression of human apolipoprotein E4 in neurons causes hyperphosphorylation of protein tau in the brains of transgenic mice. AB - Epidemiological studies have established that the epsilon 4 allele of the ApoE gene (ApoE4) constitutes an important risk factor for Alzheimer's disease and might influence the outcome of central nervous system injury. The mechanism by which ApoE4 contributes to the development of neurodegeneration remains unknown. To test one hypothesis or mode of action of ApoE, we generated transgenic mice that overexpressed human ApoE4 in different cell types in the brain, using four distinct gene promoter constructs. Many transgenic mice expressing ApoE4 in neurons developed motor problems accompanied by muscle wasting, loss of body weight, and premature death. Overexpression of human ApoE4 in neurons resulted in hyperphosphorylation of the microtubule-associated protein tau. In three independent transgenic lines from two different promoter constructs, increased phosphorylation of protein tau was correlated with ApoE4 expression levels. Hyperphosphorylation of protein tau increased with age. In the hippocampus, astrogliosis and ubiquitin-positive inclusions were demonstrated. These findings demonstrate that expression of ApoE in neurons results in hyperphosphorylation of protein tau and suggests a role for ApoE in neuronal cytoskeletal stability and metabolism. PMID- 10702413 TI - Alterations of cell cycle regulators in localized synovial sarcoma: A multifactorial study with prognostic implications. AB - Genetic alterations of cell cycle regulators are thought to represent uncommon and possible secondary events in sarcomas characterized by recurrent chromosomal translocations. The present study investigates this hypothesis on synovial sarcoma (SS), assessing the frequency of expression and possible clinical implications of detecting alterations in critical cell cycle regulatory proteins. A homogeneous cohort of 49 patients with localized SS, restricted to the extremity and with available long-term follow-up information, was selected from our files. We focused our study on molecules involved in the G1 checkpoint and G1 S transition, including cyclins D1 and E, p21(WAF1), p27(Kip1), mdm2, p53, and Ki67. A cutoff point of 10% immunoreactive tumor cell nuclei was selected to define a positive phenotype for any given marker, except for Ki67. High Ki67 proliferative index was considered when >/=20% tumor cells displayed nuclear immunoreactivity. Biphasic SS were analyzed, taking into account separately the expression of these proteins in the spindle and glandular components. Disease specific survival was modeled using the Kaplan-Meier method with log rank test and Cox regression. The cohort of patients analyzed included 23 females and 26 males, and the histological type distribution was 35 monophasic and 14 biphasic SS. The median follow-up for survivors was 53 months, with a 5-year disease specific survival of 63% and a metastatic disease-free survival of 40%. The positive phenotypes identified for the different markers studied were as follows: cyclin D1, 59%; cyclin E, 29%; p21, 51%; p27, 69%; mdm2, 59%; p53, 16%; and Ki67, 59%. We observed that positive p53, cyclin E, and high Ki67 proliferative index were correlated with survival, but only Ki67 and p53 were independent variables for prognostication. The present study suggests that alterations of cell cycle regulators are more common events in SS than originally thought. p53 overexpression could be of use as a marker together with a high Ki67 proliferative index, in identifying a subset of SS patients with increased risk of tumor relapse. PMID- 10702412 TI - Hypoxia-induced vascular endothelial growth factor expression precedes neovascularization after cerebral ischemia. AB - We investigated the hypothesis that hypoxia induces angiogenesis and thereby may counteract the detrimental neurological effects associated with stroke. Forty eight to seventy-two hours after permanent middle cerebral artery occlusion we found a strong increase in the number of newly formed vessels at the border of the infarction. Using the hypoxia marker nitroimidazole EF5, we detected hypoxic cells in the ischemic border of the neocortex. Expression of vascular endothelial growth factor (VEGF), which is the main regulator of angiogenesis and is inducible by hypoxia, was strongly up-regulated in the ischemic border, at times between 6 and 24 hours after occlusion. In addition, both VEGF receptors (VEGFRs) were up-regulated at the border after 48 hours and later in the ischemic core. Finally, the two transcription factors, hypoxia-inducible factor-1 (HIF-1) and HIF-2, known to be involved in the regulation of VEGF and VEGFR gene expression, were increased in the ischemic border after 72 hours, suggesting a regulatory function for these factors. These results strongly suggest that the VEGF/VEGFR system, induced by hypoxia, leads to the growth of new vessels after cerebral ischemia. Exogenous support of this natural protective mechanism might lead to enhanced survival after stroke. PMID- 10702414 TI - Intestinal restitution: progression of actin cytoskeleton rearrangements and integrin function in a model of epithelial wound healing. AB - Superficial injury involving the mucosa of the gastrointestinal tract heals by a process termed restitution that involves epithelial sheet movement into the damaged area. The forces that drive epithelial sheet movement are only partially understood, although it is known to involve changes in the morphology of cells bordering the damage, such as the formation of large, flat, cytoplasmic extensions termed lamellae. We investigated the mechanism of epithelial sheet movement by following the response of the actin cytoskeleton and specific integrins (alpha6beta4, alpha6beta1, and alpha3beta1) to wounding. To model this event in vitro, monolayers of T84 cells, well-differentiated colon carcinoma cells, were damaged by aspiration and the ensuing response was analyzed by a combination of time-lapse video microscopy, fluorescence confocal microscopy and antibody inhibition assays. We show that wound healing begins with retraction of the monolayer. alpha6beta4 integrin is localized on the basal surface in structures referred to as type II hemidesmosomes that persist throughout this early stage. We hypothesize that these structures adhere to the substrate and function to retard retraction. Once retraction ceases, the wound is contracted initially by actin purse strings and then lamellae. Purse strings and lamellae produce a pulling force on surrounding cells, inducing them to flatten into the wound. In the case of lamellae, we detected actin suspension cables that appear to transduce this pulling force. As marginal cells produce lamellae, their basal type II hemidesmosomes disappear and the alpha6 integrins appear evenly distributed over lamellae surfaces. Antibodies directed against the alpha6 subunit inhibit lamellae formation, indicating that redistribution of the alpha6 integrins may contribute to the protrusion of these structures. Antibodies directed against the alpha3beta1 integrin also reduce the size and number of lamellae. This integrin's contribution to lamellae extension is most likely related to its localization at the leading edge of emerging protrusions. In summary, wounds in epithelial sheets initially retract, and then are contracted by first an actin purse string and then lamellae, both of which serve to pull the surrounding cells into the denuded area. The alpha6 integrins, particularly alpha6beta4, help contain retraction and both the alpha6 integrins and alpha3beta1 integrin contribute to lamellae formation. PMID- 10702415 TI - Prevention of hepatic apoptosis and embryonic lethality in RelA/TNFR-1 double knockout mice. AB - Mice deficient in the nuclear factor kappaB (NF-kappaB)-transactivating gene RelA (p65) die at embryonic days 14-15 with massive liver apoptosis. In the adult liver, activation of the NF-kappaB heterodimer RelA/p50 can cause hepatocyte proliferation, apoptosis, or the induction of acute-phase response genes. We examined, during wild-type fetal liver development, the expression of the Rel family member proteins, as well as other proteins known to be important for NF kappaB activation. We found these proteins and active NF-kappaB complexes in the developing liver from at least 2 days before the onset of lethality observed in RelA knockouts. This suggests that the timing of NF-kappaB activation is not related to the timing of lethality. We therefore hypothesized that, in the absence of RelA, embryos were sensitized to tumor necrosis factor (TNF) receptor 1 (TNFR-1)-mediated apoptosis. Thus, we generated mice that were deficient in both RelA and TNFR-1 to determine whether apoptotic signaling through TNFR-1 was responsible for the lethal phenotype. RelA/TNFR-1 double knockout mice survived embryonic development and were born with normal livers without evidence of increased hepatocyte apoptosis. These animals became runted shortly after birth and survived an average of 10 days, dying from acute hepatitis with an extensive hepatic infiltration of immature neutrophils. We conclude that neither RelA nor TNFR-1 is required for liver development and that RelA protects the embryonic liver from TNFR-1-mediated apoptotic signals. However, the absence of both TNFR-1 signaling and RelA activity in newborn mice makes these animals susceptible to endogenous hepatic infection. PMID- 10702416 TI - Evidence for growth hormone (GH) autoregulation in pituitary somatotrophs in GH antagonist-transgenic mice and GH receptor-deficient mice. AB - Growth hormone (GH) modulates the hypothalamic release of somatostatin and GH releasing hormone; however, there has been no evidence of GH autoregulation on the pituitary somatotroph. To determine the effects of GH on its own regulation, we examined the pituitaries of giant transgenic mice expressing a GH agonist (E117L), dwarf transgenic mice expressing a GH antagonist (G119K), and dwarf mice devoid of the GH receptor/binding protein (GHR/BP). In the E117L transgenic mice, the number and distribution of pituitary GH-immunoreactive cells were unchanged from nontransgenic littermate controls; an ultrastructural examination revealed typical, densely granulated somatotrophs. In contrast, the pituitaries of the G119K mice contained both moderately granulated somatotrophs and a sparsely granulated (SG) population with well-developed synthetic organelles and a distinct juxtanuclear globular GH-staining pattern. GHR/BP-deficient mice exhibited a marked reduction in the intensity of cytoplasmic GH immunoreactivity; however, prominent GH staining in the juxtanuclear Golgi was seen. GH immunoreactive cells were increased in number, and the reticulin network pattern was distorted; stains for proliferating cell nuclear antigen confirmed mild hyperplasia. Electron microscopy showed that the somatotrophs were hyperactive SG cells with prominent endoplasmic reticulum membranes, large Golgi complexes, and numerous mitochondria. These findings are consistent with synthetic and secretory hyperactivity in pituitary somatotrophs due to the reduced GH feedback regulation. The changes are most striking in animals that are devoid of GHR/BP and less marked in animals expressing a GH antagonist; both models had reduced insulin-like growth factor-I levels, but the more dramatic change in the GHR/BP animals can be explained by abrogated GH signaling. This represents the first evidence of direct GH feedback inhibition on pituitary somatotrophs, which may have implications for the use of GH analogs in different clinical settings. PMID- 10702417 TI - Somatic mutation of the 5' noncoding region of the BCL-6 gene is associated with intraclonal diversity and clonal selection in histological transformation of follicular lymphoma. AB - Follicular lymphoma (FL) is a B cell non-Hodgkin's lymphoma (NHL) that frequently displays a t(14;18) translocation. Clonal evolution and histological transformation of FL is frequently associated with the accumulation of secondary genetic alterations. It has been demonstrated that the BCL-6 gene can be altered by chromosomal rearrangements and by mutations clustering in its 5' noncoding region in a significant fraction of FL and diffuse large cell lymphoma (DLCL). To elucidate the role of the BCL-6 gene alterations in the histological transformation and clonal progression of FL, we analyzed serial biopsy specimens from 12 patients with FL. Two cases of FL showed no histological alteration in the second biopsy, and 10 cases of FL showed morphological transformation to DLCL in the second biopsy. Southern blot analysis was used to detect rearrangement of the BCL-6 gene, polymerase chain reaction-single strand conformation polymorphism and sequence analysis were performed for identification of mutations in the 5' noncoding region of the BCL-6 gene, and immunohistochemical analysis was applied to reveal the BCL-6 protein expression. No BCL-6 gene rearrangement was detected in any of the samples, but a total of 58 mutations were found in the 5' noncoding region of the BCL-6 gene in seven cases. In five cases, both the FL and the clonally related FL or DLCL, and in two cases only the DLCL samples were mutated. The mutations were identical in multiple biopsy specimens of FL that did not show morphological transformation. In six patients where FL cells underwent morphological transformation, considerable intraclonal sequence heterogeneity was observed, indicating an ongoing type of somatic mutation. Based on the pattern of shared and nonshared mutations, the genealogical relationship of neoplastic clones could be established. In all of these cases, the histological transformation of FL was associated with the emergence of a subpopulation marked by new sites of mutations in the BCL-6 5' noncoding sequences. In three of these six cases, the histological transformation is also associated with the reduced expression of the BCL-6 protein. These findings demonstrate that mutation of the 5' noncoding region of the BCL-6 gene developed in the clonal evolution of FL, and at different time points in the lymphoma evolution different clonotypes dominate. PMID- 10702418 TI - Bax is increased in the retina of diabetic subjects and is associated with pericyte apoptosis in vivo and in vitro. AB - Diabetes of even short duration accelerates the death of capillary cells and neurons in the inner retina by a process consistent with apoptosis. We examined whether the process is accompanied by changes in the expression of endogenous regulators of apoptosis. In postmortem retinas of 18 diabetic donors (age 67 +/- 6 years, diabetes duration 9 +/- 4 years) the levels of pro-apoptotic Bax were slightly, but significantly, increased when compared with levels in 20 age matched nondiabetic donors (P = 0.04). In both groups, Bax localized to vascular and neural cells of the inner retina. Neither pro-apoptotic Bcl-X(S), nor pro survival Bcl-X(L) appeared affected by diabetes. The levels of these molecules could not be accurately quantitated in lysates of retinal vessels because of variable degrees of glial contamination. However, studies in situ showed in several pericytes, the outer cells of retinal capillaries, intense Bax staining often in conjunction with DNA fragmentation. Bovine retinal pericytes exposed in vitro to high glucose levels for 5 weeks showed elevated levels of Bax (P = 0.03) and increased frequency of annexin V binding, indicative of early apoptosis. Hence, human diabetes selectively alters the expression of Bax in the retina and retinal vascular pericytes at the same time as it causes increased rates of apoptosis. The identical program induced by high glucose in vitro implicates hyperglycemia as a causative factor in vivo, and provides a model for establishing the role of Bax in the accelerated death of retinal cells induced by diabetes. PMID- 10702419 TI - Anti-inflammatory effects of mutant forms of secretory leukocyte protease inhibitor. AB - The secretory leukocyte protease inhibitor (SLPI) is found in a variety of secreted fluids in mammals and is a known inhibitor of serine proteases. Wild type (WT) SLPI has recently been shown to block nuclear factor kappaB (NF-kappaB) activation in rat lungs and to interfere with the ensuing inflammatory response and recruitment of neutrophils after an intrapulmonary deposition of IgG immune complexes. In this study, WT SLPI and SLPI mutants with various degrees of protease-inhibitory capacity (for trypsin, chymotrypsin, and elastase) were evaluated for their ability to suppress the lung-vascular leak, neutrophil accumulation, and NF-kappaB activation in the lung inflammatory model. The SLPI mutant with Gly(72) (replacing Leu(72) ) lost its ability to block in vivo activation of NF-kappaB, as well as its ability to suppress the lung vascular leak and neutrophil recruitment. The Phe(72) and Gly(20) mutants were as effective as the WT SLPI in suppressing NF-kappaB activation and neutrophil recruitment. The Lys(72) mutant had the most suppressive effects of the lung vascular leak and for neutrophil recruitment into the lung. The in vivo suppressive effects of SLPI mutants on lung vascular permeability, neutrophil recruitment, and NF-kappaB activation appear to be most closely related to their trypsin-inhibiting activity. These data suggest that the suppressive effects of SLPI on the intrapulmonary activation of NF-kappaB and neutrophil recruitment into the lung may be linked to their antiprotease activity, directed, perhaps, at the intracellular proteases. PMID- 10702420 TI - New roles for glial cell line-derived neurotrophic factor and neurturin: involvement in hair cycle control. AB - Glial cell line-derived neurotrophic factor (GDNF), neurturin (NTN), and their receptors, GDNF family receptor alpha-1 (GFRalpha-1) and GDNF family receptor alpha-2 (GFRalpha-2), are critically important for kidney and nervous system development. However, their role in skin biology, specifically in hair growth control, is as yet unknown. We have studied expression and function of GDNF, neurturin, GFRalpha-1, and GFRalpha-2 in murine skin during the cyclic transformation of the hair follicle (HF) from its resting state (telogen) to active growth (anagen) and then through regression (catagen) back to telogen. GDNF protein and GFRalpha-1 messenger RNA are prominently expressed in telogen skin, which lacks NTN and GFRalpha-2 transcripts. Early anagen development is accompanied by a significant decline in the skin content of GDNF protein and GFRalpha-1 transcripts. During the anagen-catagen transition, GDNF, GFRalpha-1, NTN, and GFRalpha-2 transcripts reach maximal levels. Compared with wild-type controls, GFRalpha-1 (+/-) and GFRalpha-2 (-/-) knockout mice show a significantly accelerated catagen development. Furthermore, GDNF or NTN administration significantly retards HF regression in organ-cultured mouse skin. This suggests important, previously unrecognized roles for GDNF/GFRalpha-1 and NTN/GFRalpha-2 signaling in skin biology, specifically in the control of apoptosis-driven HF involution, and raises the possibility that GFRalpha 1/GFRalpha-2 agonists/antagonists might become exploitable for the treatment of hair growth disorders that are related to abnormalities in catagen development. PMID- 10702421 TI - Reactive changes of retinal microglia during fatal murine cerebral malaria: effects of dexamethasone and experimental permeabilization of the blood-brain barrier. AB - Microglial activation and redistribution toward blood vessels are some of the earliest observable events occurring within the central nervous system (CNS) during fatal murine cerebral malaria (FMCM). To investigate stimuli that might modulate microglial reactivity during FMCM we have performed two experimental manipulations and observed microglial responses in retinal whole mounts. First, to determine whether increased blood-brain barrier (BBB) permeability in the absence of the malaria parasite initiates the microglial changes, BBB function was compromised experimentally by intracarotid injection of arabinose and retinae were examined 12, 24, or 36 hours later. Second, to determine whether the immune response against the malaria parasite modulates microglial reactivity, infected mice were treated with dexamethasone before day 4 postinoculation. This treatment regime ameliorates cerebral complications without affecting parasite growth. We observed that increased BBB permeability was sufficient to elicit thickening of microglial processes and redistribution of microglia toward the vasculature, characteristic of the early stages of FMCM. However, despite the presence of plasma constituents in the CNS for up to 36 hours, microglia with amoeboid and vacuolated morphology were not observed. Dexamethasone treatment inhibited the up regulation of alpha-D-galactose expression and reactive morphological changes in microglia during FMCM. These results suggest that disruption of the CNS milieu by entry of plasma constituents, or circulating malaria parasites in the absence of an immune response, by themselves are insufficient to induce the reactive microglial changes that are characteristic of FMCM. In addition, dexamethasone sensitive event(s), presumably associated with immune system activation, occurring within the first few days of malaria infection are essential for the development of reactive microglia and subsequent fatal neurological complications. PMID- 10702422 TI - Single-cell PCR analysis of T helper cells in human lymph node germinal centers. AB - The T helper cell population of human lymph node germinal centers (GCs) was analyzed for clonality and signs of antigen selection. Frozen sections of lymph node biopsies taken from three different individuals were used to micromanipulate single T cells from one particular GC for each of the specimens. T cell receptor (TCR) beta gene rearrangements were amplified from these single cells and directly sequenced. Although only unique rearrangements were amplified from T cells of GC2 and GC3, 11 of 28 potentially functional rearrangements amplified from GC1 originated from four different clones. In all three GCs, TCR gene rearrangements neither showed obvious biases in gene segment usage nor similarities in complementarity determining region 3 amino acid sequence. Thus, it appears that T lymphocytes in human GCs usually represent a diverse population of cells. Sequence analysis of V region genes did not provide evidence that in the human the process of somatic hypermutation acts on the TCRbeta loci. For one of the GCs (GC3), immunoglobulin heavy chain (IgH) gene rearrangements were amplified and sequenced from single micromanipulated GC B cells. The detection of clonal expansions accounting for more than half of the sampled B cells in addition to ongoing somatic hypermutation of Ig V region genes suggested that GC3 was a fully developed GC. PMID- 10702423 TI - Targeted disruption of the galectin-3 gene results in attenuated peritoneal inflammatory responses. AB - Galectin-3 is a member of a growing family of beta-galactoside-binding animal lectins. Previous studies have demonstrated a variety of biological activities for this protein in vitro, including activation of cells, modulation of cell adhesion, induction of pre-mRNA splicing, and regulation of apoptosis. To assist in fully elucidating the physiological and pathological functions of this protein, we have generated galectin-3-deficient (gal3(-/-)) mice by targeted interruption of the galectin-3 gene. Gal3(-/-) mice consistently developed fewer inflammatory cell infiltrations in the peritoneal cavities than the wild-type (gal3(+/+)) mice in response to thioglycollate broth treatment, mainly due to lower numbers of macrophages. Also, when compared to cells from gal3(+/+) mice, thioglycollate-elicited inflammatory cells from gal3(-/-) mice exhibited significantly lower levels of NF-kappaB response. In addition, dramatically different cell-spreading phenotypes were observed in cultured macrophages from the two genotypes. Whereas macrophages from gal3(+/+) mice exhibited well spread out morphology, those from gal3(-/-) mice were often spindle-shaped. Finally, we found that peritoneal macrophages from gal3(-/-) mice were more prone to undergo apoptosis than those from gal3(+/+) mice when treated with apoptotic stimuli, suggesting that expression of galectin-3 in inflammatory cells may lead to longer cell survival, thus prolonging inflammation. These results strongly support galectin-3 as a positive regulator of inflammatory responses in the peritoneal cavity. PMID- 10702424 TI - Surface antigen expression and complement susceptibility of differentiated neuroblastoma clones. AB - Human neuroblastoma cell lines typically consist of heterogenous subpopulations of cells that are morphologically and biochemically distinct. The cell types are characterized as neuroblastic (N-type), substrate-adherent Schwann-like (S-type), or intermediate (I). These cell types can undergo spontaneous or induced transdifferentiation in vitro. We investigated the complement sensitivity of different neuroblastoma cell lines and of matched sets of cloned N- and S-type neuroblastoma cell lines. Human neuroblastoma cell lines that consisted predominantly of a neuroblastic phenotype were shown to be significantly more susceptible to human complement-mediated lysis than cell lines of other cancer types. Complement sensitivity of neuroblastoma cell lines was correlated with low levels of CD59, decay-accelerating factor, and membrane cofactor protein expression. We found that cloned S-type neuroblastoma cells were much more resistant to complement-mediated lysis than cloned N-type cells. The increased complement resistance of S-type cells was shown to be due to increased expression of membrane-bound complement inhibitors. CD59 was the single most important protein in providing S-type cells with protection from complement lysis. S-type cells were also found to express lower levels of GD2, a target antigen for a complement activating monoclonal antibody currently in clinical trials for neuroblastoma immunotherapy. The ability of S-type cells to evade complement, and the ability of S-type cells to differentiate into the more tumorigenic N-type cells, may represent a mechanism of tumor survival and regrowth, a phenomenon often observed with this cancer. PMID- 10702425 TI - Molecular analysis of phyllodes tumors reveals distinct changes in the epithelial and stromal components. AB - Phyllodes tumors are fibroepithelial mammary lesions that tend to behave in a benign fashion but may undergo sarcomatous transformation. A study of clonality in these tumors has suggested that the epithelial component is polyclonal, but the stroma is monoclonal, and thus forms the neoplastic component of the lesion. In this study microsatellites on chromosome 1q and chromosome 3p were assessed for allelic imbalance (AI) in 47 phyllodes tumors; in all cases stroma and epithelium were analyzed separately. Ten of 42 (24%) phyllodes tumors showed AI at one or more markers on 3p, and 14 of 46 (30%) showed AI on chromosome 1. Five tumors had changes in both the epithelium and stroma. Eight tumors had changes only detectable in the stroma and eight, changes in the epithelium only. Three tumors exhibited low-level microsatellite instability in the epithelium but not in the stroma. The results show that AI on 3p and 1q does occur in phyllodes tumors and that it can occur in both the stroma and epithelium, sometimes as independent genetic events. These unexpected findings throw into doubt the classical view that phyllodes tumors are simply stromal neoplasms and raise questions about the nature of stromal and epithelial interactions in these tumors. PMID- 10702426 TI - Analysis of the effect of endogenous viral genes in the Smyth line chicken model for autoimmune vitiligo. AB - The Smyth line (SL) chicken, an animal model for autoimmune human vitiligo, is characterized by a spontaneous posthatch pigment loss, determined to be the result of an autoimmune phenomenon. Because endogenous virus (EV) genes have been reported to be associated with a number of autoimmune diseases of human and animal models, we designed this experiment to investigate the role of EV in the SL vitiligo by using the complete sequence of Rous-associated virus-2 as a probe for EV. An F(2) resource population was developed by the matings of SL and parental control (BL) chickens. Linkage disequilibrium between vitiligo and EV was apparent (16.2-kb SacI fragment, P 30 kg/m(2)) adolescents are comparable to those in similarly aged nonobese adolescents when dosing is calculated based on total body mass and not lean body mass. When a rapid sequence induction of anesthesia is considered in an obese adolescent, the dose of succinylcholine should be based on actual (not lean) body mass. PMID- 10702441 TI - Cardiovascular criteria for epidural test dosing in sevoflurane- and halothane anesthetized children. AB - This study was designed to determine the detectability of a simulated IV test dose in children during administration of general anesthesia by using heart rate (HR), systolic blood pressure (SBP), and T wave criterion. Forty-two children (0.5-8 yr old) received an IV injection containing epinephrine 0.5 microg/kg and another IV injection containing saline during either halothane or sevoflurane anesthesia administration at 1.0 minimum alveolar concentration in nitrous oxide. A positive test response was defined as a change in T wave amplitude >/=25%, SBP increase >/=15 mm Hg, and HR increase >/=10 bpm. By using the T wave, SBP, and HR criteria, a positive response rate to epinephrine was 100%, 95%, and 71%, respectively, during sevoflurane, and 90%, 71%, and 71%, respectively, during halothane anesthesia administration. These data suggest that the T wave criterion is superior to conventional hemodynamic criteria, and that sevoflurane attenuates T wave and SBP responses less than halothane; however, chronotropic responses are similar to halothane. IMPLICATIONS: We found a greater reliability of the T wave criterion over conventional hemodynamic criteria for detecting intravascular injection of a simulated epidural test dose. Sevoflurane may increase the likelihood of recognition of an accidental intravascular injection of epinephrine containing solutions in clinical practice compared with halothane. PMID- 10702442 TI - The cost effectiveness of anesthesia workforce models: a simulation approach using decision-analysis modeling. AB - The objective of this study was to evaluate the incremental cost effectiveness of anesthesia workforce staffing scenarios, as a function of skill mix, by using the technique of decision analysis. A decision tree model was constructed to compare the incremental cost effectiveness of alternative delivery systems for anesthesia care from the perspective of the payer. Five different staffing scenarios, ranging from physician-intensive to nurse-intensive, were modeled. In the nurse intensive model, low- and intermediate-risk patients were cared for by solo certified registered nurse anesthetists (CRNAs) and high-risk patients were cared for by physicians. In the physician-intensive model, physicians anesthetized all patients. In the first-, second-, and third-team models, all high-risk patients were cared for by physicians working alone, and all intermediate-risk patients were cared for using an anesthesia care team approach with a ratio of one physician to two CRNAs. The low-risk patients were managed by using an anesthesia care team approach with physician to CRNA ratios of 1:2, 1:4, and 1:8 in the first-, second-, and third-team models, respectively. The findings of this decision-analysis model suggest that physician-only anesthesia is not cost effective. However, the third-team model is cost effective when compared with the nurse-intensive model. IMPLICATIONS: An anesthesia care-team approach with a physician to certified registered nurse anesthetist (CRNA) ratio of 1:2 is the preferred staffing scenario for intermediate-risk patients. Although medical direction of CRNAs caring for low-risk patients is cost-effective, the small improvement in outcome resulting from increasing the physician to CRNA ratio from 1:8 to 1:4 may not be justified by the added cost. PMID- 10702443 TI - Comparing methods of clinical measurement: reporting standards for bland and altman analysis. AB - In this era of medical technology assessment and evidence-based medicine, evaluating new methods to measure physiologic variables is facilitated by standardization of reporting results. It has been proposed that assessing repeatability be followed by assessing agreement with an established technique. If the "limits of agreement" (mean bias +/- 2SD) are not clinically important, then one could use two measurements interchangeably. Generalizability to larger populations is facilitated by reporting confidence intervals. We identified 44 studies that compared methods of clinical measurement published during 1996 to 1998 in seven anesthesia journals. Although 42 of 44 (95.4%) used the limits of agreement methodology for analysis, several inadequacies and inconsistencies in reporting the results were noted. Limits of agreement were defined a priori in 7.1%, repeatability was evaluated in 21.4%, and relationship (pattern) between difference and average was evaluated in 7.1%. Only one of the articles reported confidence intervals. A computer macro for the Minitab statistical package (State College, PA) is described to facilitate reporting of Bland and Altman analysis with confidence intervals. We propose standardization of nomenclature in clinical measurement comparison studies. IMPLICATIONS: A literature review of anesthesia journals revealed several inadequacies and inconsistencies in statistical reports of results of comparison studies with regard to interchangeability of measurement methods. We encourage journal editors to evaluate submissions on this subject carefully to ensure that their readers can draw valid conclusions about the value of new technologies. PMID- 10702444 TI - The effects of morphine on blood-brain barrier disruption caused by intracarotid injection of hyperosmolar mannitol in rats. AB - This study was performed to evaluate whether morphine could alter the degree of disruption of the blood-brain barrier (BBB) caused by hyperosmolar mannitol. Under isoflurane anesthesia, rats in a control group were infused with 25% mannitol into the internal carotid artery before measuring the transfer coefficient (Ki) of (14)C-alpha-aminoisobutyric acid. Infusion of morphine 3 mg/kg in the small-dose morphine group and 10 mg/kg in the large-dose morphine group was completed, 10 min before administering mannitol. There were no statistical differences in systemic blood pressures between these three groups of animals. In the control group, the Ki of the ipsilateral cortex where mannitol was injected, increased to 4.6 times that of the contralateral cortex (19.5 +/- 8.5 vs 4.2 +/- 1.2 microL. g(-1). min(-1), P < 0.002). The Ki of the ipsilateral cortex of the small-dose morphine group was 13.5 +/- 7.6 microL. g(-1). min(-1). The Ki of the ipsilateral cortex of the large-dose morphine group was 9.2 +/- 4.5 microL. g(-1). min(-1) and was smaller than that of control animals (P < 0.05). There was no significant difference in the Ki of the contralateral cortex among the three groups. In conclusion, morphine attenuated BBB disruption induced by hyperosmolar solution without significant effects on systemic blood pressure. IMPLICATIONS: Our study suggests that morphine may be effective in reducing the blood-brain barrier disruption by hyperosmolar mannitol without significant effects on systemic blood pressure. PMID- 10702445 TI - The effects of remifentanil on cerebral capacity in awake volunteers. AB - Remifentanil, a short-acting potent mu-opioid agonist proposed for intraoperative analgesia but also for postoperative pain therapy, has not been investigated with regard to the effects of the drug on cerebral capacity in awake humans. We assessed cerebral capacity noninvasively by means of phase-contrast magnetic resonance imaging measurement of systolic cerebrospinal fluid peak velocity in the aqueduct of Sylvius before and during infusion of remifentanil (0.1 microg. kg(-1). min(-1) IV) in normocapnic humans. Remifentanil had no significant effect on systolic cerebrospinal fluid peak velocity as compared with baseline (mean +/- SD): baseline, -4.3 +/- 1.3 cm/s versus remifentanil (0.1 microg. kg(-1). min( 1)): -4.7 +/- 1.0 cm/s. Small-dose remifentanil (0.1 microg. kg(-1). min(-1)) did not influence cerebral capacity in healthy, awake volunteers free of intracranial pathology. IMPLICATIONS: Knowledge about the influence of remifentanil on cerebral capacity is crucial before routine use of the drug in neuroanesthesia. Thus, we assessed the influence of remifentanil on cerebral capacity noninvasively by means of phase-contrast magnetic resonance imaging measurement of systolic cerebrospinal fluid peak velocity in the aqueduct of Sylvius in humans. PMID- 10702446 TI - Investigating hypoxemia during apnea: validation of a set of physiological models. AB - The aim of our study was to validate the Nottingham Physiology Simulator (NPS) for examining pulmonary denitrogenation and apnea by reproducing the methodology and results of previous clinical studies. Only four studies provided sufficient detail in their description of their methodology to allow accurate reproduction by using the NPS or provided a sufficiently detailed description of their subjects to allow accurate modelling. The results of the NPS recreation of the studies were within 13% of the values found clinically in all cases and were within 2% in the majority of cases. The four studies included healthy and morbidly obese patients, conscious and anesthetized patients, and included examination of the effect of denitrogenation and apnea on plasma pH and on lung and arterial oxygen and carbon dioxide tensions at various lung volumes. IMPLICATIONS: We used mathematical, physiological models to recreate the methods and subjects of four clinical studies investigating oxygenation and low oxygen levels during cessation of breathing. Our aim was to validate the models, allowing theoretical investigations into this area. The blindly recreated results closely matched the clinical studies, validating the models. PMID- 10702447 TI - Factors determining the onset and course of hypoxemia during apnea: an investigation using physiological modelling. AB - We used the Nottingham Physiology Simulator to examine the onset and course of hypoxemia during apnea after pulmonary denitrogenation. The following factors, as possible determinants of the hypoxemia profile, were varied to examine their effect: functional residual capacity, oxygen consumption, respiratory quotient, hemoglobin concentration, ventilatory minute volume, duration of denitrogenation, pulmonary venous admixture, and state of the airway (closed versus open). Airway obstruction significantly reduced the time to 50% oxyhemoglobin saturation (8 vs 11 min). Provision of 100% oxygen rather than air to the open, apneic patient model greatly prolonged time to 50% oxyhemoglobin saturation (66 vs 11 min). Hemoglobin concentration, venous admixture, and respiratory quotient had small, insignificant effects on the time to desaturation. Reduced functional residual capacity, short duration of denitrogenation, hypoventilation, and increased oxygen consumption significantly shortened the time to 50% oxyhemoglobin saturation during apnea. IMPLICATIONS: Reduction in oxygen levels during cessation of breathing is dangerous and common in anesthetic practice. We used validated, mathematical, physiological models to reveal the impact of physiological factors on the deterioration of oxygen levels. This study could not be performed on patients and reveals important information. PMID- 10702448 TI - Cyclooxygenase inhibitors attenuate bradykinin-induced vasoconstriction in septic isolated rat lungs. AB - Cyclooxygenase (COX) products play an important role in modulating sepsis and subsequent endothelial injury. We hypothesized that COX inhibitors may attenuate endothelial dysfunction during sepsis, as measured by receptor-mediated bradykinin (BK)-induced vasoconstriction and/or receptor-independent hypoxic pulmonary vasoconstriction (HPV). Rats were administered intraperitoneally a nonselective COX inhibitor (indomethacin, 5 or 10 mg/kg) or a selective COX-2 inhibitor (NS-398, 4 or 8 mg/kg) 1 h before lipopolysaccharide (LPS, 15 mg/kg), or saline (control). Three hours later, the rats were anesthetized, the lungs were isolated, and pulmonary vasoreactivity was assessed with BK (0.3, 1.0, and 3.0 microg) and HPV (3% O(2)). Perfusion pressure was monitored as an index of vasoconstriction. To investigate what receptor-subtype is mediating BK responses, the BK(1)-receptor antagonist des-Arg(9)-[Leu(8)]-BK, the BK(2)-receptor antagonist HOE-140, or the thromboxane A(2)-receptor antagonist SQ 29548 (all at 1 microM) were added to the perfusate. BK-induced vasoconstriction was significantly increased in LPS lungs (1.4-5.2 mm Hg) compared with control (0.1 1.1 mm Hg). In LPS lungs, indomethacin 10 mg/kg significantly decreased BK vasoconstriction by 78% +/- 9%, whereas 5 mg/kg did not. NS-398, 4 mg/kg, significantly attenuated BK vasoconstriction at 0.3 microg (71% +/- 7%) and 1.0 microg (56% +/- 12%), whereas 8 mg/kg attenuated 0.3 microg BK (57% +/- 14%), compared with LPS lungs. HPV was increased in LPS lungs (21.5 +/- 2 mm Hg) compared with control lungs (9.8 +/- 0.6 mm Hg). Indomethacin 5 mg/kg increased HPV in LPS lungs; otherwise, HPV was not altered by COX inhibition. BK-induced vasoconstriction was prevented by BK(2), but not BK(1) or thromboxane A(2) receptor antagonism. This study suggests that nonselective COX inhibition, and possibly inhibition of the inducible isoform COX-2, may attenuate sepsis-induced, receptor-mediated vasoconstriction in rats. IMPLICATIONS: This study demonstrated that, in an isolated rat lung model, nonselective inhibition of the cyclooxygenase pathway, and possibly selective inhibition of the inducible cyclooxygenase-2 isoform, may attenuate sepsis-induced endothelial dysfunction. PMID- 10702449 TI - A comparison of epidural analgesia with 0.125% ropivacaine with fentanyl versus 0.125% bupivacaine with fentanyl during labor. AB - We previously found that the extent of an epidural motor block produced by 0.125% ropivacaine was clinically indistinguishable from 0.125% bupivacaine in laboring patients. By adding fentanyl to the 0. 125% ropivacaine and bupivacaine solutions in an attempt to reduce hourly local anesthetic requirements, we hypothesized that differences in motor block produced by the two drugs may become apparent. Fifty laboring women were randomized to receive either 0. 125% ropivacaine with fentanyl 2 microg/mL or an equivalent concentration of bupivacaine/fentanyl using patient-controlled epidural analgesia (PCEA) with settings of: 6-mL/hr basal rate, 5-mL bolus, 10-min lockout, 30-mL/h dose limit. Analgesia, local anesthetic use, motor block, patient satisfaction, and side effects were assessed until the time of delivery. No differences in verbal pain scores, local anesthetic use, patient satisfaction, or side effects between groups were observed; however, patients administered ropivacaine/fentanyl developed significantly less motor block than patients administered bupivacaine/fentanyl. Ropivacaine 0.125% with fentanyl 2 microg/mL produces similar labor analgesia with significantly less motor block than an equivalent concentration of bupivacaine/fentanyl. Whether this statistical reduction in motor block improves clinical outcome or is applicable to anesthesia practices which do not use the PCEA technique remains to be determined. IMPLICATIONS: By using a patient-controlled epidural analgesia technique, ropivacaine 0.125% with fentanyl 2 microg/mL produces similar analgesia with significantly less motor block than a similar concentration of bupivacaine with fentanyl during labor. Whether this statistical reduction in motor block improves clinical outcome or is applicable to anesthesia practices which do not use the patient-controlled epidural analgesia technique remains to be determined. PMID- 10702450 TI - The inhibition of epidural morphine-induced pruritus by epidural droperidol. AB - IV droperidol inhibits epidural morphine-induced pruritus, but this effect disappears when the dose is increased from 2.5 to 5.0 mg. This study was performed to determine whether epidural droperidol would have a similar effect. In this double-blinded study, we enrolled 140 patients undergoing Cesarean delivery under epidural anesthesia who were randomly allocated to four groups. Anesthesia consisted of 150 mg of 0.5% bupivacaine with 1:200,000 epinephrine, with 2 mg of morphine and 0.0, 1.25, 2.5, or 5.0 mg of droperidol (Groups 1 to 4). During the postoperative period, patients were assessed for pruritus (absent, mild, moderate, or severe) and other untoward symptoms. The chi(2) test was used to compare the incidence of the side effects. For the analysis of pruritus, we used the Mantel-Haenszel test for linear association. Droperidol induced a dose related reduction in the incidence of pruritus (P < 0.001). This reduction was independent of the incidence of somnolence, which increased with droperidol dose (P < 0.05 when the incidence of somnolence in Groups 1 and 4 was compared). We conclude that droperidol, in doses up to 5 mg epidurally, induces a dose-related reduction in the incidence of pruritus without inducing significant side effects. IMPLICATIONS: Epidural morphine is effective for pain control but yields some side effects, including pruritus, that can be severe. Studying patients undergoing Cesarean delivery, we found a dose-related reduction in the incidence of pruritus using epidural droperidol. PMID- 10702451 TI - A comparison of epidural levobupivacaine 0.75% with racemic bupivacaine for lower abdominal surgery. AB - Levobupivacaine, the S(-) isomer of bupivacaine, is less cardiotoxic than racemic bupivacaine. In this prospective, randomized, double-blinded study of epidural anesthesia, the onset, extent, and duration of sensory and motor block produced by 0.75% levobupivacaine (20 mL, 150 mg) was compared with that of 0.75% racemic bupivacaine in 56 patients undergoing elective lower abdominal surgery. The time to onset of adequate sensory block (T10 dermatome) was similar in both treatment groups (13.6 +/- 5.6 min for levobupivacaine and 14.0 +/- 9.9 min for bupivacaine), with an average peak block height of T5 reached at 24.3 +/- 9.4 and 26.5 +/- 13.2 min, respectively. Time to complete regression of sensory block was significantly longer with levobupivacaine (550.6 +/- 87.6 min) than bupivacaine (505.9 +/- 71.1 min) (P = 0.016). Abdominal muscle relaxation was adequate for the scheduled procedure in all patients, and there were no significant differences between the groups in rectus abdominis muscle scores (P = 0.386) and quality of muscle relaxation as determined by the surgeon and anesthesiologist (P = 0. 505 and 0.074, respectively). In conclusion, both 0.75% levobupivacaine and 0.75% bupivacaine produced effective epidural anesthesia and their effects were clinically indistinguishable. IMPLICATIONS: The results of this study indicate that the sensory and motor block produced by 0.75% levobupivacaine is equivalent to that of 0.75% racemic bupivacaine. Both local anesthetics are well tolerated and effective in producing epidural anesthesia for patients undergoing lower abdominal surgery. PMID- 10702452 TI - What concentration of sufentanil should be combined with ropivacaine 0.2% wt/vol for postoperative patient-controlled epidural analgesia? AB - In this randomized double-blinded study, we sought to determine an optimal dose combination of sufentanil with ropivacaine 0.2% wt/vol as postoperative epidural analgesics. One hundred twenty patients undergoing major abdominal surgery under general and thoracic epidural anesthesia (T9-11) were assigned to groups receiving patient-controlled epidural analgesia with ropivacaine 0.2% wt/vol (R), ropivacaine 0.2% wt/vol + sufentanil 0.5 microg/mL (R+S0.5), 0. 75 microg/mL (R+S0.75), 1.0 microg/mL (R+S1). A visual analog score of less than 40 was considered effective, and all side effects were recorded. In randomized subgroups (10 patients per group), plasma pharmacokinetic data were obtained for both epidural drugs. Four patients in Group R and two in Group R+S0.5 were excluded because of inadequate analgesia. The drug infusion rates (range of means: 5.4-5. 9 mL/h) were similar in all patients. Analgesia was superior for sufentanil 0.75 microg/mL with no further enhancement by the larger sufentanil concentration of 1 microg/mL. Sufentanil plasma levels were within the range of the minimal effective concentrations (highest in R+S1), and there was no covariation between plasma levels and pain relief. Free ropivacaine plasma concentrations remained stable for 96 h. No severe side effects were detected, although pruritus correlated with an increasing dose of sufentanil. We conclude that the combination of ropivacaine 0.2% wt/vol and 0.75 microg/mL sufentanil provided the best analgesia with the fewest side effects of the three combinations tested. IMPLICATIONS: Sufentanil is added to epidural infusions of ropivacaine 0.2% wt/vol to improve the effectiveness of postoperative pain management. Regarding the risk of side effects, however, it is still unclear what concentration of sufentanil should be added to the local anesthetic. For postoperative thoracic epidural analgesia after major abdominal surgery, the combination of ropivacaine 0.2% wt/vol and 0.75 microg/mL sufentanil resulted in an appropriate cost:benefit ratio between good analgesia and side effects. PMID- 10702453 TI - Spinal nerve function in five volunteers experiencing transient neurologic symptoms after lidocaine subarachnoid anesthesia. AB - The etiology of transient neurologic symptoms (TNS) after 5% lidocaine spinal anesthesia remains undetermined. Previous case reports have shown that patients acutely experiencing TNS have no abnormalities on neurologic examination or magnetic resonance imaging. The aim of our study was to determine whether volunteers with TNS would exhibit abnormalities in spinal nerve electrophysiology. Twelve volunteers with no history of back pain or neurologic disease underwent baseline electromyography (EMG), nerve conduction studies, and somatosensory-evoked potential (SSEP) testing. Then, the volunteers were administered 50 mg of 5% hyperbaric lidocaine spinal anesthesia and were placed in a low lithotomy position (legs on four pillows). The next day, all volunteers underwent follow-up EMG, nerve conduction, and SSEP testing and were questioned and examined for the presence of complications including TNS (defined as pain or dysthesia in one or both buttocks or legs occurring within 24 h of spinal anesthesia). Volunteers who had TNS underwent additional EMG testing 4-6 wk later. Five of the 12 volunteers reported TNS. No volunteer had an abnormal EMG, nerve conduction study, or SSEP at 24 h follow up, nor were there any changes in EMG studies at delayed testing in the five volunteers experiencing TNS. On statistical analysis, the right peroneal and the right tibial nerve differed significantly for all volunteers from pre- to postspinal testing. When comparing pre- and postspinal testing of the TNS and non-TNS volunteers, statistically significant changes occurred in the nerve conduction tests of the right peroneal and left tibial nerve. There was no difference in measurements of F response, H reflex latency, amplitude, or velocity for either leg. Multivariate analysis of variance showed no significant difference between TNS and non-TNS volunteers for the changes in the nine nerve conduction tests when considered together (P = 0.4). We conclude that acute TNS after lidocaine spinal anesthesia did not result in consistent abnormalities detectable by EMG, nerve conduction studies, or SSEP in five volunteers. IMPLICATIONS: Electrophysiologic testing in volunteers experiencing transient neurologic symptoms is not abnormal. PMID- 10702454 TI - The efficacy of intravenous 0.15 versus 0.25 mg/kg intraoperative morphine for immediate postoperative analgesia after remifentanil-based anesthesia for major surgery. AB - We evaluated the effect of perioperative administration of two doses of morphine for postoperative analgesia after remifentanil-based anesthesia. The prospective, randomized study included 245 patients from 33 centers. All patients were scheduled for abdominal or urological surgery lasting more than 1 h. General anesthesia used remifentanil as the perioperative opioid (1 microg/kg as a bolus then, 0.5 microg/kg as a continuous infusion). A morphine bolus of 0. 15 mg/kg (0.15-mg group) or 0.25 mg/kg (0.25-mg group) was administered 30 min before the end of surgery. In the postanesthesia care unit, pain scores for patients were evaluated by using behavioral pain scores of 1-3, verbal pain scores of 0-3, and visual analog scale scores of 0-10). Postoperative analgesia was obtained by a morphine titration (3 mg every 5 min). Demographic and surgery characteristics were similar in both groups. The delay for first demand of morphine was similar in the 0.15-mg and the 0.25-mg groups (26 [9-60] and 30 [10-60] min, respectively). The frequency of morphine titration was similar in both groups (75% and 66%, respectively). The amount of morphine used in the postanesthesia care unit was smaller in the 0.25-mg group (0.16 [0.0-1.25] vs 0.10 [0.0-0.56] mg/kg; P = 0.008). In the 0.25-mg group, the behavioral pain score was lower at 15 min, the verbal pain score was lower at 60 min (P < 0.001), and similar at 30 min. The visual analog scale pain score at 30 min and 60 min was similar in both groups. The incidence of minor side effects was similar in both groups. However, three cases of postoperative respiratory depression occurred in the 0.25-mg group compared with no cases in the 0.15-mg group. In conclusion, perioperative administration of morphine alone does not provide entirely adequate immediate postoperative pain control after remifentanil-based anesthesia in major surgery. IMPLICATIONS: The administration of 0.15 or 0.25 mg/kg perioperative morphine during remifentanil-based anesthesia for major surgery does not preclude additional morphine administration in the postanesthesia care unit. The larger dose of 0.25 mg/kg slightly improves postoperative analgesia; however, it may be responsible for postoperative respiratory depression. PMID- 10702455 TI - The analgesic efficacy of intravenous tenoxicam as an adjunct to patient controlled analgesia in total abdominal hysterectomy. AB - Nonsteroidal antiinflammatory drugs may reduce postoperative opioid consumption. We evaluated the analgesic efficacy of preoperatively administered tenoxicam in patients undergoing total abdominal hysterectomy. Patients were randomly assigned to receive IV either normal saline 4 mL (Group NS), tenoxicam 20 mg (Group T20), or tenoxicam 40 mg (Group T40) before the induction of anesthesia in a double blinded fashion. Patient-controlled analgesia with fentanyl was used to assess postoperative opioid requirements. Pain was evaluated by visual analog scale at 2, 4, 6, 8, and 24 h postoperatively. Intraoperative bleeding as assessed by the surgeon, incidence of nausea, and gastrointestinal symptoms were recorded. No statistically significant difference was identified between groups in fentanyl consumption or pain scores. The incidence of nausea was similar in all groups. Two patients in Group T20 and two in Group T40 exhibited mild gastrointestinal symptoms. Intraoperative oozing was noted in two patients in Group T40. We conclude that patients undergoing total abdominal hysterectomy and receiving fentanyl via patient-controlled analgesia postoperatively do not benefit from tenoxicam pretreatment. On the contrary, the drug may be associated with an increased incidence of side effects. IMPLICATIONS: The preoperative administration of 20 or 40 mg IV tenoxicam does not reduce fentanyl consumption via Patient-Controlled Analgesia, compared with placebo, after total abdominal hysterectomy. Additionally, tenoxicam may increase intraoperative bleeding and gastrointestinal side effects. PMID- 10702456 TI - The role of heme oxygenase in neuropathic and incisional pain. AB - Heme oxygenase (HO) catalyzes the formation of free iron, biliverdin, and the second messenger molecule carbon monoxide from heme. We document a role for HO in both neuropathic and incisional pain models. For our neuropathic model, the L5 and L6 nerve roots of rats were ligated unilaterally resulting in mechanical allodynia and thermal hyperalgesia in the ipsilateral hind paws. Both changes were dose-dependently reversed by systemic administration of the HO inhibitor tin protoporphyrin (Sn-P). Likewise, a 1-cm incision made in one hind paw resulted in mechanical allodynia and thermal hyperalgesia, again reversible by using Sn-P. The 50% effective doses for Sn-P ranged from 4.0 to 6.8 micromol/kg depending on the model and nociceptive stimulus. We also observed that the blood-brain barrier impermeable HO inhibitor zinc protoporphyrin had little analgesic activity in these models when injected systemically. Using an enzymatic assay, we observed increased HO activity in lumbar spinal cord tissue from either nerve root ligated or incised animals as compared with tissue from sham-operated animals. Taken together, we interpret our results to indicate that an increase in spinal cord HO activity at least partially underlies the allodynia and hyperalgesia seen in rat models of neuropathic and incisional pain. IMPLICATIONS: Central nervous system heme oxygenase likely plays a role in nociceptive signaling in both neuropathic and incisional models of pain. Therefore, inhibitors of heme oxygenase activity may be viable analgesics in these settings. PMID- 10702457 TI - The effects of sevoflurane on serum creatinine and blood urea nitrogen concentrations: a retrospective, twenty-two-center, comparative evaluation of renal function in adult surgical patients. AB - Despite mounting clinical evidence that supports its safety, the question of the potential adverse effects of sevoflurane on renal function continues to generate some controversy. This study retrospectively evaluated pooled renal laboratory data from 22 different clinical trials that compared sevoflurane with three widely used anesthetics. The trials examined postoperative changes in serum creatinine and blood urea nitrogen levels from a total of 3, 436 ASA physical status I-IV adult surgical patients administered either sevoflurane (n = 1941) or a control drug (isoflurane, enflurane, or propofol; n = 1495) as the maintenance anesthetic. The incidences of increased serum creatinine and blood urea nitrogen concentrations were similar among patients administered sevoflurane and those administered control drugs. Additionally, no trends specific to sevoflurane were observed with respect to postoperative serum creatinine concentration and fresh gas flow rate, concurrent treatment with nephrotoxic antibiotics, or type of carbon dioxide absorbent. IMPLICATIONS: Our data for changes in serum creatinine and blood urea nitrogen indicate that, for exposures of less than 4 minimum alveolar anesthetic concentration/h, sevoflurane is not associated with an increased risk of renal toxicity compared with other commonly used anesthetics. For clinical purposes, the pre- to postoperative changes in serum creatinine and blood urea nitrogen are appropriate measures of renal function in surgical patients. PMID- 10702458 TI - Reversal of rapacuronium block during propofol versus sevoflurane anesthesia. AB - We studied the antagonism of rapacuronium with edrophonium-atropine during propofol- or sevoflurane- based anesthesia in 60 healthy outpatients. After the induction of anesthesia with standardized doses of propofol and fentanyl, rapacuronium 1.5 mg/kg was administered to facilitate tracheal intubation. Patients were randomized to receive either a propofol infusion (100 microg. kg( 1). min(-1)) or sevoflurane (1.0%, end-tidal) in combination with nitrous oxide 66% for maintenance of anesthesia. Neuromuscular block was monitored by using electromyography at the wrist and reversed with edrophonium 1.0 mg/kg and atropine 0.015 mg/kg when the first twitch (T(1)) response of the train-of-four (TOF) stimulation recovered to 25% of the baseline value. The clinical duration of action (i.e., time to 25% T(1) recovery) was similar during both propofol (13.1 +/- 3.6 min) and sevoflu-rane (13.7 +/- 4.4 min) anesthesia. The time from 25% T(1) recovery to TOF ratio of 0.8 was also similar with propofol (3.4 +/- 2.1 min) and sevoflurane (5.9 +/- 8.7 min) (P > 0.05). Although none of the patients in the propofol group required more than 9 min to achieve a TOF ratio of 0. 8, two patients receiving sevoflurane required 31 min and 37 min. Adequate antagonism of rapacuronium block with edrophonium can be achieved within 10 min during propofol anesthesia. However, more prolonged recovery may occur in the presence of sevoflurane. IMPLICATIONS: We studied the reversal of rapacuronium induced block with edrophonium and found that the residual rapacuronium block can be readily antagonized during propofol-based anesthesia. However, reversal of rapacuronium appears to be less predictable during sevoflurane-based anesthesia. PMID- 10702459 TI - Age-related thermoregulatory differences in a warm operating room environment (approximately 26 degrees C). AB - Inadvertent hypothermia occurs frequently at typical ambient operating room (OR) temperatures, especially in elderly patients receiving general anesthesia. The aims of the current study were to 1) determine the incidence and magnitude of core hypothermia in an unusually warm OR environment, and 2) to assess age related differences in perioperative thermoregulatory responses under these circumstances. Forty patients receiving general anesthesia for orthopedic surgical procedures (20 younger patients, 20-40 yr old) and (20 older patients, 60-75 yr old) were enrolled. Mean ambient temperature in the ORs was 25.8 degrees +/- 0.2 degrees C. Core temperature, vasoconstriction, and shivering were compared in the younger and older age groups. Mean core temperature on admission to the postanesthesia care unit was not significantly different in the younger (36.7 degrees +/- 0.1 degrees C) and older (36.4 degrees +/- 0.1 degrees C) age groups. Only 10% of patients (n = 4, 1 younger, 3 older) were admitted with a core temperature <36.0 degrees C. Only 2% of patients (n = 1, older group) had a core temperature <35.5 degrees C. This very mild degree of hypothermia was associated with postoperative vasoconstriction in 80% of the younger and 55% of the older patients (P = 0.18). Postoperative shivering occurred in 40% of the younger patients and in 10% of the older patients (P = 0.06). In summary, an ambient OR temperature near 26 degrees C (79 degrees F) is effective in preventing core hypothermia during general anesthesia regardless of patient age. Even very mild postoperative hypothermia may initiate thermoregulatory responses. IMPLICATIONS: By increasing ambient temperature in the operating room to 26 degrees C (79 degrees F), the incidence of core hypothermia can be dramatically reduced in both younger and older patients. PMID- 10702460 TI - Sedative, amnestic, and analgesic properties of small-dose dexmedetomidine infusions. AB - This research determined the safety and efficacy of two small-dose infusions of dexmedetomidine by evaluating sedation, analgesia, cognition, and cardiorespiratory function. Seven healthy young volunteers provided informed consent and participated on three occasions with random assignment to drug or placebo. Heart rate, blood pressure, respiratory rate, ETCO(2), O(2) saturation, and processed electroencephalogram (bispectral analysis) were monitored. Baseline hemodynamic measurements were acquired, and psychometric tests were performed (visual analog scale for sedation; observer's assessment of alertness/sedation scale; digit symbol substitution test; and memory). The pain from a 1-min cold pressor test was quantified with a visual analog scale. After a 10-min initial dose of saline or 6 microg. kg(-1). h(-1) dexmedetomidine, volunteers received 50 min IV infusions of saline, or 0.2 or 0.6 microg. kg(-1). h(-1) dexmedetomidine. Measurements were repeated at the end of infusion and during recovery. The two dexmedetomidine infusions resulted in similar and significant sedation (30%-60%), impairment of memory (approximately 50%), and psychomotor performance (28%-41%). Hemodynamics, oxygen saturation, ETCO(2), and respiratory rate were well preserved throughout the infusion and recovery periods. Pain to the cold pressor test was reduced by 30% during dexmedetomidine infusion. Small-dose dexmedetomidine provided sedation, analgesia, and memory and cognitive impairment. These properties might prove useful in a postoperative or intensive care unit setting. IMPLICATIPNS: The alpha(2) agonist, dexmedetomidine, has sedation and analgesic properties. This study quantified these effects, as well as cardiorespiratory, memory and psychomotor effects, in healthy volunteers. Dexmedetomidine infusions resulted in reversible sedation, mild analgesia, and memory impairment without cardiorespiratory compromise. PMID- 10702461 TI - The visual analog scale allows effective measurement of preoperative anxiety and detection of patients' anesthetic concerns. AB - The advent of managed care, reduction of costs, and advances in medical technology place increasing demands on anesthesiologists. Preoperative anxiety may go unnoticed in an environment that stresses increased productivity. The present study compares different methods for measuring preoperative anxiety, identifies certain patient characteristics that predispose to high anxiety, and describes the quantity and quality of anxiety that patients experience preoperatively. Seven hundred thirty-four patients participated in the study. We assessed aspects of anxiety by means of visual analog scales (VAS) and the State Anxiety Score of the Spielberger State-Trait Anxiety Inventory (STAI). The mean STAI anxiety score was 39 +/- 1 (n = 486) and the mean VAS for fear of anesthesia was 29 +/- 1 (n = 539). Patients feared surgery significantly more than anesthesia (P < 0.001). The VAS measuring fear of anesthesia correlated well with the STAI score (r = 0.55; P < 0.01). Young patients, female patients, and patients with no previous anesthetic experience or a previous negative anesthetic experience had higher anxiety scores. Patients worried most about the waiting period preceding surgery and were least concerned about possible awareness intraoperatively. Factor analysis of various anxiety items showed three distinct dimensions of fear: 1) the fear of the unknown 2) the fear of feeling ill, and 3) the fear for one's life. Among these dimensions, fear of the unknown correlated highest with the anxiety measuring techniques STAI and VAS. The simple VAS proved to be a useful and valid measure of preoperative anxiety. IMPLICATIONS: The study of qualitative aspects of anxiety reveals three distinct dimensions of preoperative fear: fear of the unknown, fear of feeling ill, and fear for one's life. Groups of patients with a higher degree of preoperative anxiety and their specific anesthetic concerns can be identified using the visual analog scale. PMID- 10702462 TI - Is an infusion pump necessary to safely administer remifentanil? AB - We sought to determine if remifentanil could be administered as safely and effectively from an IV drip as from a calculator pump, because not all anesthesiologists have access to a calculator pump. Forty healthy adults undergoing outpatient knee arthroscopy were premedicated with midazolam, 2 mg. Total IV anesthesia was induced with propofol by bolus (2 mg/kg) and maintained by a continuous infusion of propofol and remifentanil. On a randomized, double blinded basis, they received, IV, either remifentanil (50 microg/mL) by syringe from an infusion pump or from a bag of saline containing remifentanil 20 microg/mL through a minidrip set. The remifentanil infusion syringe pump rate was 0.4 microg. kg(-1). min(-1) until skin incision and then 0.2 microg. kg(-1). min( 1), whereas that from the bag/minidrip set was set to approximate the delivery rate from the pump. Both a syringe pump and bag/minidrip set infusion were administered to each patient but only one contained remifentanil, that one being determined in a randomized, double-blinded manner. There were no differences in demographic data, time to recovery of open eyes, response to command, ability to speak (approximately 7 min), total dose and time of administration of propofol and remifentanil, the incidence of intraoperative hypotension and bradycardia, and postoperative shivering. We demonstrated that remifentanil can be administered as safely and effectively from a bag with a minidrip set as from a syringe in a calculator infusion pump, provided the anesthesiologist is paying attention to the drip rate from the bag. IMPLICATIONS: Because remifentanil is rapidly degraded in the body, it can be safely and effectively administered from a bag through a minidrip set. We showed that there was no difference with this less expensive method of administration than from the more precise method of a calculator infusion pump. PMID- 10702463 TI - A comparison of rabeprazole, lansoprazole, and ranitidine for improving preoperative gastric fluid property in adults undergoing elective surgery. AB - Acid aspiration syndrome at the induction of anesthesia is still a potentially life-threatening complication. Its severity is affected by both pH and volume of the gastric juice that is aspirated. We compared the effects of rabeprazole (a new proton pump inhibitor), lansoprazole, and ranitidine on gastric fluid properties in a prospective, randomized, double-blinded fashion in 180 adult patients undergoing elective surgery. Patients were divided into six groups (n = 30 in each) according to their premedication. Patients in each group received placebo-rabeprazole (PLA-RAB), rabeprazole-placebo (RAB-PLA), rabeprazole rabeprazole (RAB-RAB), lansoprazole-lansoprazole (LAN-LAN), placebo-ranitidine (PLA-RAN), or placebo-placebo (PLA-PLA) for the first-second medication. Each dose of the study drug was 20 mg for rabeprazole, 30 mg for lansoprazole, and 150 mg for ranitidine. The first medication was given orally at 9:00 PM on the day before surgery and the second at 5:30 AM on the day of surgery. Each patient fasted overnight and took the drug with 20 mL of water. After tracheal intubation, gastric fluid was aspirated via an orogastric tube, and the volume and pH of the aspirate was measured. Preoperative gastric fluid acidity and volume were improved by the study drugs in the following order: PLA-RAN (pH 5.3, volume 0.10 mL/kg), RAB-RAB, LAN-LAN, PLA-RAB, and RAB-PLA (pH 3.8, volume 0.22 mL/kg). The proportion of patients at risk of acid aspiration syndrome according to the traditional criteria (pH < 2.5 and volume > 0.4 mL/kg) was minimized in Groups RAB-RAB and PLA-RAN (0%). We concluded that a single morning dose of ranitidine rather than two doses (bedtime and morning) of rabeprazole was the most effective premedicant to control gastric fluid properties and to minimize the risk of aspiration pneumonitis. IMPLICATIONS: Acid aspiration syndrome at the induction of anesthesia is rare but still a potentially life-threatening complication. We compared rabeprazole, lansoprazole, and ranitidine for reduction of preoperative gastric fluid acidity and volume in elective surgery and found that a combination of bedtime and morning doses of rabeprazole, or a morning dose of ranitidine, similarly minimized the variables. In adult patients who are at risk of aspirating gastric contents, improvement of gastric fluid environment by rabeprazole can reasonably be anticipated to provide protection against pneumonitis should regurgitation and aspiration of gastric contents occur. PMID- 10702464 TI - Supraspinal, not spinal, alpha(2) adrenoceptors are involved in the anesthetic sparing and hemodynamic-stabilizing effects of systemic clonidine in rats. AB - Clonidine, an alpha(2) agonist, reduces the anesthetic requirement and attenuates harmful hemodynamic responses to noxious stimuli. We examined the responsible sites of action in the central nervous system for the minimum alveolar anesthetic concentration (MAC) and MAC blocking adrenergic response (MAC-BAR) reducing effects of systemically administered clonidine in halothane-anesthetized rats. The MAC for halothane was determined by the tail clamp method, and MAC-BAR was defined as the MAC which attenuated hemodynamic responses within 10% after the tail clamp. We examined the effect of IV clonidine in the presence of rauwolscine, an alpha(2) antagonist given through IV, intrathecal (IT), intracisternal (IC), or intracerebroventrical (ICV) routes. IV clonidine reduced MAC and MAC-BAR dose-dependently. IV and ICV rauwolscine antagonized the MAC reducing effect of clonidine, whereas IC and IT rauwolscine did not. In comparison, IV, ICV, and IC rauwolscine antagonized the MAC-BAR-reducing effect of clonidine; IT rauwolscine had no effect. Our data demonstrate that the alpha(2) adrenoceptors in the regions above mesencephalon and both the regions above mesencephalon and the lower brainstem are responsible for the MAC and MAC BAR-reducing effect of systemic clonidine in rats, respectively. However, the spinal alpha(2) adrenoceptors were not involved in these effects of clonidine. IMPLICATIONS: In the regions above mesencephalon, alpha(2) adrenoceptors were the most responsible for the minimum alveolar concentration-reducing effect and both the lower brainstem and regions above mesencephalon were involved in the minimum alveolar concentration blocking adrenergic response-reducing effect of clonidine. The spinal alpha(2) adrenoceptors did not significantly contribute to these effects of clonidine. PMID- 10702465 TI - Patch-clamp analysis of anesthetic interactions with recombinant SK2 subtype neuronal calcium-activated potassium channels. AB - Small conductance calcium-activated potassium channels (SK) mediate spike frequency adaptation and underlie the slow afterhyperpolarization in central neurons. We tested the actions of several anesthetics on the SK2 subtype of recombinant SK channels, cloned from rat brain and functionally expressed in a mammalian cell line. Butanol, ethanol, ketamine, lidocaine, and methohexital blocked recombinant SK2 channel currents, measured in the whole-cell patch clamp recording mode. The block was reversible, dose-dependent, and of variable efficacy. The inhaled anesthetics chloroform, desflurane, enflurane, halothane, isoflurane, and sevoflurane produced little or no block when applied at 1 minimum alveolar anesthetic concentration; varying degrees of modulation were observed at very large concentrations (10 minimum alveolar concentration). The extent of block by inhaled anesthetics did not appear to depend on concentration or membrane voltage. IMPLICATIONS: We describe differential effects of anesthetics on cloned small conductance calcium-activated potassium channels from brain that may play a role in generating the effects or side effects of anesthetics. PMID- 10702467 TI - The awareness of being observed changes the patient's psychological well-being in anesthesia. PMID- 10702466 TI - No implicit memory for stories played during isoflurane/alfentanil/nitrous oxide anesthesia: a reading speed measurement. AB - Implicit memory of intraoperatively presented stories was recently detected by using the reading speed paradigm during propofol-alfentanil-nitrous oxide anesthesia. Our main goal was to evaluate the reading speed test procedure under another anesthetic regimen, i.e., isoflurane combined with nitrous oxide and alfentanil-infusion. In both experiments, patients were premedicated with oral midazolam. In a previous experiment, patients postoperatively read "old" stories that had been presented during anesthesia quicker compared with "new," unpresented stories. The same study design and test material as in the previous experiment were used. One of two audio tapes with two short stories was played randomly to patients during lumbar disk surgery and to awake controls. Approximately 7 h later, a structured interview and the reading speed test were used to determine whether the participants had any explicit or implicit memories of the presented stories. The results of 30 patients and 30 controls were calculated. Whereas the control participants showed an intact explicit and implicit memory of the previously presented material, no such effect was found in the anesthetized patients. The present experiment shows that changing the main anesthetic in otherwise equal study conditions, i. e., propofol to isoflurane (end-expiratory 0.7%), implicit memory is abolished in anesthetized patients. IMPLICATIONS: We showed that implicit memory during general anesthesia can be abolished by changing the hypnotic anesthetic. Increased postoperative reading speed for stories presented during propofol-alfentanil-nitrous oxide anesthesia was shown in a previous experiment, but not in our study using isoflurane for balanced anesthesia. PMID- 10702468 TI - Suspected malfunction of the SAF-T-FILL(TM) valve assembly of the Suprane (Desflurane, USP) refill bottle. PMID- 10702469 TI - Lighted stylet tracheal intubation: a review. PMID- 10702470 TI - Difficult endotracheal intubation associated with torus mandibularis. PMID- 10702471 TI - Subcutaneous carbon dioxide insufflation does not cause hypercarbia during endoscopic thyroidectomy. PMID- 10702472 TI - Air embolism: a complication during transcervical resection of the endometrium. PMID- 10702473 TI - Inappropriate statements can lead to misleading conclusions. PMID- 10702474 TI - Preemptive analgesia with ketamine. PMID- 10702475 TI - Perineural concentration of lidocaine is more relevant to spinal neurotoxicity than the concentration administered. PMID- 10702476 TI - A simple technique to reduce preservative/excipient related neurotoxicity of intrathecal (spinal) drugs. PMID- 10702477 TI - Thoracic epidurals in coronary artery bypass surgery. PMID- 10702478 TI - The difficult neuraxial block: predictors or self-fulfilling prophecy? PMID- 10702479 TI - Is laparoscopic surgery associated with specific hemodynamic changes? PMID- 10702480 TI - Jugular bulb desaturations during propofol anesthesia in neurosurgical procedures. PMID- 10702481 TI - Increased safety in the administration of aprotinin: need for a test-dose. PMID- 10702482 TI - The lateral approach to the sciatic nerve. PMID- 10702484 TI - Monitoring specific T-cell responses to melanoma vaccines: ELISPOT, tetramers, and beyond. PMID- 10702483 TI - Immunoglobulin D: properties, measurement, and clinical relevance. PMID- 10702485 TI - Evaluation of the modified ELISPOT assay for gamma interferon production in cancer patients receiving antitumor vaccines. AB - Frequencies of vaccine-responsive T-lymphocyte precursors in peripheral blood mononuclear cells (PBMC) prior to and after administration of peptide-based vaccines in patients with cancer can be measured by limiting-dilution assays (LDA) or by ELISPOT assays. We have used a modified version of the ELISPOT assay to monitor changes in the frequency of gamma interferon (IFN-gamma)-producing T cells in a population of lymphocytes responding to a relevant peptide or a nonspecific stimulator, such as phorbol myristate acetate-ionomycin. Prior to its use for monitoring of patient samples, the assay was validated and found to be comparable to the LDA performed in parallel, using tumor-reactive cytolytic T lymphocyte (CTL) lines. The sensitivity of the ELISPOT assay was found to be 1/100,000 cells, with an interassay coefficient of variation of 15%, indicating that it could be reliably used for monitoring of changes in the frequency of IFN gamma-secreting responder cells in noncultured or cultured lymphocyte populations. To establish that the assay is able to detect the T-cell precursor cells responsive to the vaccine, we used CD8(+) T-cell populations positively selected from PBMC of HLA-A2(+) patients with metastatic melanoma, who were treated with dendritic cell-based vaccines containing gp100, MELAN-A/MART-1, tyrosinase, and influenza virus matrix peptides. The frequency of peptide specific responder T cells ranged from 0 to 1/2,600 before vaccination and increased by at least 1 log unit after vaccination in two patients, one of whom had a clinical response to the vaccine. However, no increases in the frequency of peptide-responsive T cells were observed in noncultured PBMC or PBMC cultured in the presence of the relevant peptides after the melanoma patients enrolled in another trial were treated with the intramuscular peptide vaccine plus MF59 adjuvant. Thus, while the ELISPOT assay was found to be readily applicable to assessments of frequencies of CTL precursors of established CTL lines and ex vivo amplified PBMC, its usefulness for monitoring of fresh PBMC in patients with cancer was limited. In many of these patients antitumor effector T cells are present at frequencies of lower than 1/100,000 in the peripheral circulation. Serial monitoring of such patients may require prior ex vivo amplification of specific precursor cells. PMID- 10702486 TI - Diagnosis of tuberculosis based on the two specific antigens ESAT-6 and CFP10. AB - Tests based on tuberculin purified protein derivative (PPD) cannot distinguish between tuberculosis infection, Mycobacterium bovis BCG vaccination, or exposure to environmental mycobacteria. The present study investigated the diagnostic potential of two Mycobacterium tuberculosis-specific antigens (ESAT-6 and CFP10) in experimental animals as well as during natural infection in humans and cattle. Both antigens were frequently recognized in vivo and in vitro based on the induction of delayed-type hypersensitivity responses and the ability to induce gamma interferon production by lymphocytes, respectively. The combination of ESAT 6 and CFP10 was found to be highly sensitive and specific for both in vivo and in vitro diagnosis. In humans, the combination had a high sensitivity (73%) and a much higher specificity (93%) than PPD (7%). PMID- 10702487 TI - Differential induction of complement fragment C5a and inflammatory cytokines during intramammary infections with Escherichia coli and Staphylococcus aureus. AB - The prompt recruitment of neutrophils to the site of infection is essential for the defense of the bovine mammary gland against invading pathogens and is determinant for the outcome of the infection. Escherichia coli is known to induce clinical mastitis, characterized by an intense neutrophil recruitment leading to the eradication of the bacteria, whereas Staphylococcus aureus induces subclinical mastitis accompanied by a moderate neutrophil recruitment and the establishment of chronic mastitis. To elicit the neutrophil recruitment into the udder, inflammatory mediators must be produced after recognition of the invading pathogen. To our knowledge, those mediators have never been studied during S. aureus mastitis, although understanding of the neutrophil recruitment mechanisms could allow a better understanding of the differences in the pathogeneses elicited by E. coli and S. aureus. Therefore, we studied, at several time points, the accumulation of neutrophils and the presence of the chemoattractant complement fragment C5a and of the cytokines interleukin-1beta (IL-1beta), tumor necrosis factor alpha, and IL-8 in milk after inoculation of E. coli or S. aureus in lactating bovine udders. The low levels of C5a and the absence of cytokines in milk from S. aureus-infected cows, compared to the high levels found in milk from E. coli-infected animals, mirror the differences in the severities of the two inflammatory reactions. The cytokine deficit in milk after S. aureus inoculation in the lactating bovine mammary gland could contribute to the establishment of chronic mastitis. This result could help in the design of preventive or curative strategies against chronic mastitis. PMID- 10702488 TI - Characterization of a predominant immunogenic outer membrane protein of Riemerella anatipestifer. AB - The ompA gene, encoding the 42-kDa major antigenic outer membrane protein OmpA of Riemerella anatipestifer, the etiololgical agent of septicemia anserum exsudativa, was cloned and expressed in Escherichia coli. Recombinant OmpA displayed a molecular mass similar to that predicted from the nucleotide sequence of the ompA gene but lower than that observed in total cell lysates of R. anatipestifer. The ompA gene showed a conserved C-terminal region comprising the OmpA-like domain and a variable N-terminal region. This structure is similar to those of the analogous outer membrane proteins of several gram-negative bacteria. However, OmpA of R. anatipestifer contains six EF-hand calcium-binding domains and two PEST regions, which distinguish it from other outer membrane proteins. The occurrence of these motifs in OmpA suggests a possible role in virulence for this protein. The ompA gene is present in the R. anatipestifer type strain and in all serotype reference strains. However, it exhibits some minor genetic heterogeneity among different serotypes, which seems not to affect the strong antigenic characteristics of the protein. OmpA is a conserved and strong antigenic determinant of R. anatipestifer and hence is suggested to be a valuable protein for the serodetection of R. anatipestifer infections, independent of their serotype. PMID- 10702489 TI - Anti-idiotypic antibodies in patients with different clinical forms of paracoccidioidomycosis. AB - Paracoccidioidomycosis (PCM) is the most prevalent systemic mycosis in Latin America. Patients with PCM show a wide spectrum of clinical and pathological manifestations depending on both host and pathogen factors. Two clinical forms of the disease are recognized: the acute or juvenile form and the chronic or adult form. The major antigenic component of the parasite is a glycoprotein of 43 kDa (gp43). All patient sera present antibodies against gp43 (anti-gp43) and, as demonstrated before by our group, spontaneous anti-idiotypic (anti-Id) antibodies (Ab2) can be detected in patient sera with high titers of anti-gp43. Since it has been postulated that anti-Id antibodies may have a modulating function, we decided to purify and characterize anti-Id antibodies in this system. The possible correlation of Ab2 titers with different clinical forms of disease was also verified. Results showed that purified human anti-Id antibodies (human Ab2) recognized specifically the idiotype of some murine monoclonal anti-gp43 (17c and 3e) but not others (40.d7, 27a, and 8a). Spontaneous anti-Id antibodies were found in all clinical forms of disease. The majority of patients (88%, n = 8) with the acute form of PCM had high titers of Ab2. However, among patients with the multifocal chronic form of the disease, only 29% (n = 14) had high titers of Ab2; 70% (n = 10) of patients with the unifocal chronic form had low titers of Ab2. A correlation between Ab2 titers and anti-gp43 titers was observed before and during antimycotic treatment. Our results suggest that titers of anti-Id antibodies correlate with the severity of PCM in humans. PMID- 10702490 TI - Down regulation of CD4 expression following isolation and culture of human monocytes. AB - The down regulation of CD4 by cultured monocytes has been observed by our group and by other investigators. Flow cytometric experiments were done to examine which factors might influence this phenomenon. The addition of lipopolysaccharide, granulocyte-macrophage colony-stimulating factor, macrophage colony-stimulating factor, or interleukin-10 to monocyte cultures failed to inhibit the decrease in monocyte CD4 expression routinely observed following overnight culture. The down regulation was an adherence-independent phenomenon and was not influenced by the type of anticoagulant into which the peripheral blood was collected or by the presence or absence of lymphocytes within the cultures. The avoidance of the use of Ficoll-Paque to isolate peripheral blood mononuclear cells did not prevent monocyte CD4 down regulation. Finally, by tagging monocyte CD4 with an anti-CD4 phycoerythrin-conjugated monoclonal antibody prior to culture, we were able to determine that the down regulation observed was the result of the internalization of the molecule. At this time, we conclude that the observed down regulation of monocyte CD4 is probably due to the differentiation of blood monocytes into tissue culture-derived macrophages rather than to some artifact of the isolation procedure. PMID- 10702492 TI - Lipid removal from human serum samples. AB - The efficacy of lipid removal from human serum samples obtained by using Cleanascite HC, a commercially available product, was compared to that obtained by the standard chloroform method. Separate samples of 21 frozen, banked human serum samples used in the preparation of samples for proficiency testing were treated with either Cleanascite HC or chloroform. The lipid content was measured before and after treatment. The total percentages of lipid removed ranged from 61 to 70% with Cleanascite HC and from 60 to 62% with chloroform. The advantage of Cleanascite HC over chloroform is based on the simplicity of the procedure with Cleanascite HC without the environmental concerns inherent in the use of chloroform. In 15 serum samples known to contain antibodies to treponemal and nontreponemal syphilis antigens, Cleanascite HC bound some immunoglobulin, but with only minimal loss of reactivity in the serologic tests for syphilis. Cleanascite HC is therefore an acceptable alternative to chloroform for lipid reduction in human serum samples. PMID- 10702491 TI - Immunoglobulin A (IgA) deficiency and alternative celiac disease-associated antibodies in sera submitted to a reference laboratory for endomysial IgA testing. AB - Immunoglobulin A (IgA) deficiency occurs more frequently in patients with celiac disease (CD) than in the general population and can lead to false-negative results in the best serologic test for CD, endomysial IgA (EMA). To evaluate the impact of IgA deficiency on serologic detection of CD in a reference laboratory setting, IgA levels were measured in 510 consecutive serum specimens submitted for testing for EMA; 510 consecutive serum specimens submitted for Helicobacter pylori IgG testing served as a gastrointestinal symptom control group. The frequency of IgA deficiency was significantly higher among the specimens submitted for testing for EMA (5.1%) than among the specimens from the symptom control group (1.4%). Three subsets of sera from the group of specimens submitted for testing for EMA were then tested by additional serologic assays for CD; these subsets were EMA-positive sera (n = 25), EMA-negative, IgA-deficient sera (n = 26), and control sera (from EMA-negative, IgA-nondeficient patients age matched to IgA-deficient patients; n = 26). The proportions of EMA-positive sera positive by other assays for CD were 92% for transglutaminase IgA (TG-IgA), 80% for gliadin IgA, 84% for gliadin IgG, 60% for endomysial IgG (EMG), and 32% for transglutaminase IgG (TG-IgG). Very low proportions (0 to 8%) of IgA-deficient sera and control sera were positive for TG-IgA, gliadin IgA, EMG, and TG-IgG. Eight of 26 (31%) IgA-deficient serum samples were positive for gliadin IgG, whereas 3 of 26 (12%) control serum samples were positive for gliadin IgG, but this difference was not statistically significant. Physicians supplied clinical data for 18 of 26 patients with IgA deficiency; only 4 patients had undergone small-bowel biopsy, and 0 of 4 patients showed villous atrophy. These findings show that IgA deficiency is found more frequently among sera submitted for testing for EMA in a reference laboratory setting, but there was no clear-cut serologic or clinical evidence of CD in EMA-negative, IgA-deficient patients. PMID- 10702493 TI - Expression of regeneration and tolerance factor correlates directly with human immunodeficiency virus infection and inversely with hepatitis C virus infection. AB - Hepatitis C virus (HCV) and human immunodeficiency virus (HIV) cause two of the most prevalent debilitating viral infections. HIV appears to induce a skewing toward a Th2 response, while in HCV infection a Th1 response appears to dominate. Regeneration and tolerance factor (RTF) may participate in driving or sustaining a Th2 cytokine response. The expression of RTF on CD3(+) T cells of HIV seropositive (HIV(+)) individuals is increased. The purpose of this study was to compare the expression of RTF during HIV infections with that during HCV infections. Three-color flow-cytometric analysis of peripheral blood collected from HIV(+) HCV-seropositive (HCV(+)), HIV- and HCV-seropositive (HIV(+) HCV(+)), and HIV- and HCV-seronegative (HIV(-) HCV(-)) individuals was performed. Levels of RTF expression on T-lymphocyte subsets from these groups were compared, as were levels of RTF expression on activated T cells expressing CD38 and HLA-DR, to determine the relationship of RTF expression to these infections. We demonstrated that the expression of RTF on surfaces of T cells from HIV(+) individuals is upregulated and that its expression on T cells from HCV(+) individuals is downregulated. A twofold increase in the mean channel fluorescence of RTF on CD3(+) T cells was seen in both HIV(+) and HIV(+) HCV(+) individuals compared to HIV(-) HCV(-) individuals. HCV(+) individuals had lower levels of RTF expression than HIV(-) HCV(-) individuals (P < 0.005 for CD4(+); P < 0.0005 for CD8(+)). In terms of percentages of T cells expressing RTF, the groups were ranked as follows: HIV(+) > HIV(+) HCV(+) > HIV(-) HCV(-) > HCV(+). The results indicate that RTF expression correlates with HIV-associated immune activation and may be associated with Th2-type responses. PMID- 10702494 TI - Sequential use of paraformaldehyde and methanol as optimal conditions for the direct quantification of ZEBRA and rta antigens by flow cytometry. AB - A technique was developed with flow cytometry to quantify the two immediate-early proteins ZEBRA and Rta, which are involved in the activation of Epstein-Barr virus replication. We evaluated four monoclonal antibodies on four cell lines (B95-8, RAJI, Namalwa, and P3HR1) with varying levels of expression of these replication-phase antigens. The Namalwa lymphoma cell line was used as a negative control. Four fixation-permeabilization procedures were compared. The preparation of cells with paraformaldehyde and methanol in sequence, and antigen detection with AZ125 and AR 5A9 monoclonal antibodies, were found to be the optimal conditions in these cell lines. Our procedure allowed ZEBRA antigen to be detected in 4.85% of peripheral blood mononuclear cells from a transplant recipient with a lymphoproliferative disease. PMID- 10702495 TI - Relationship between IS901 in the Mycobacterium avium complex strains isolated from birds, animals, humans, and the environment and virulence for poultry. AB - A total of 738 strains of Mycobacterium avium complex (MAC) were examined in biological experiments on poultry by use of PCR methods with primers for detection of the insertion sequence IS901. Serotype strains of MAC from all known 28 serotypes were examined. Further strains were isolated from human immunodeficiency virus (HIV)-negative and HIV-positive patients, 6 animal species, 17 bird species, and the environment. Of 165 strains virulent for poultry, characterized by generalized tuberculosis, 164 strains contained IS901, a result which is statistically highly significant (P, 0.01). The remaining 573 strains were nonvirulent; however, IS901 was present in 24 strains. From among 20 strains of serotypes 1, 2, and 3, IS901 was found in 15 strains, only 5 of which were virulent for poultry. The remaining 111 strains, of serotypes 4 to 28, were nonvirulent and did not incorporate IS901. None of the 152 strains isolated from humans was virulent for poultry, including 12 strains which were IS901 positive. PMID- 10702496 TI - Cytoskeletal alterations in lipopolysaccharide-induced bovine vascular endothelial cell injury and its prevention by sodium arsenite. AB - Morphological changes, especially cytoskeletal alterations, in lipopolysaccharide (LPS)-induced vascular endothelial cell injury were studied by using LPS susceptible bovine aortic endothelial cells (BAEC). BAEC in cultures with LPS showed cell rounding, shrinking, and intercellular gap formation. In those cells, LPS caused the disorganization of actin, tubulin, and vimentin. LPS also induced a reduction in the F-actin pool and an elevation in the G-actin pool. Cytoskeletal disorganization affected transendothelial permeability across the endothelial monolayer. Pretreatment of BAEC with sodium arsenite (SA) prevented alterations in LPS-induced BAEC injury. However, posttreatment with SA had no protective effect on them. SA upregulated the expression of heat shock protein in the presence of LPS. The role of SA in prevention of LPS-induced BAEC injury is discussed. PMID- 10702497 TI - Characterization of the priming effect by pituitary canine growth hormone on canine polymorphonuclear neutrophil granulocyte function. AB - In this report, we demonstrate that canine growth hormone (cGH) is capable of priming canine polymorphonuclear neutrophil granulocytes (PMN) in a manner resembling that of human PMN. The cGH influences important functions that are involved in the process of recruitment of PMN, i.e., shape change, chemotaxis, CD11b/CD18 expression, adhesion, and subsequent transendothelial migration. Also, intracellular O(2)(-) production was evaluated. We investigated the priming effect by incubating PMN with purified pituitary cGH at various concentrations (10 to 800 microg/liter). The capacity for shape change was significantly (P < 0.05) enhanced, whereas the chemotactic response under agarose was significantly (P < 0.05) reduced. The chemotactic migration in Boyden chambers (10-microm-thick polycarbonate filter; lower surface count technique) was significantly (P < 0.05) enhanced, presumably due to cGH-induced hyperadhesiveness to the lower surface of the filters. The adhesion in albumin-coated microtiter plates and adherence to canine pulmonary fibroblasts were significantly (P < 0.05) increased, and the increased adhesion resulted in a significant (P < 0.01) increase in transendothelial migration using canine jugular vein endothelial cells. The increase in adhesion was associated with a significant increase in CD11b/CD18 expression. Furthermore, intracellular O(2)(-) production was significantly enhanced in response to both phorbol myristate acetate (P < 0.01) and opsonized zymosan (P < 0.05). In the absence of a PMN-stimulating agent, cGH did not influence the effector functions investigated except for an increased expression of CD11b/CD18. PMID- 10702498 TI - Use of an attenuated leishmanial parasite as an immunoprophylactic and immunotherapeutic agent against murine visceral leishmaniasis. AB - The ability of the leishmanial parasite UR6 to act as an immunoprophylactic and immunotherapeutic agent against Leishmania donovani infection in BALB/c mice was investigated. Unlike the virulent L. donovani AG83 (MOHOM/IN/1983/AG83), UR6 given through intracardiac route failed to induce visceral infection, but when it was injected subcutaneously, UR6 induced a short-lived and localized self-healing skin lesion. Priming of peritoneal macrophages with UR6 in vitro induced superoxide (O(2)(-)) generation, whereas similar experiments with virulent AG83 inhibited O(2)(-) generation. It was observed that priming of mice with either live or sonicated UR6 in the absence of any adjuvant provided strong protection against subsequent virulent challenge. Further, UR6-primed infected mice not only displayed a strong antileishmanial delayed-type hypersensitivity (DTH) response but also showed an elevated level of the serum antileishmanial immunoglobulin G2a (IgG2a) isotype, whereas infected mice failed to mount any antileishmanial DTH response and showed an elevated level of IgG1. This indicates that UR6 priming and subsequent L. donovani infection allowed the expansion of Th1 cells. Our studies indicate that UR6 has potential to be used as an immunoprophylactic and immunotherapeutic agent against experimental visceral leishmaniasis. PMID- 10702499 TI - Anticardiolipin antibodies in patients with chronic hepatitis C virus infection: characterization in relation to antiphospholipid syndrome. AB - The antiphospholipid syndrome (APS) is usually defined by the association of clinical manifestations that comprise venous and/or arterial thrombosis, recurrent fetal losses, and thrombocytopenia, along with the presence of anticardiolipin (aCL) antibodies and/or lupus anticoagulant. Various infectious diseases can induce aCL; however, these antibodies are not usually associated with thrombotic events, as happens with autoimmune diseases, in which these antibodies need the presence of beta(2)-glycoprotein I. Levels of immunoglobulin G (IgG) and IgM aCL antibodies were determined by enzyme-linked immunosorbent assay for 243 patients with chronic hepatitis C virus (HCV) infection and 100 healthy controls. Clinical events of APS, the level of beta(2)-glycoprotein dependence of aCL, the presence of cryoglobulins and other autoantibodies, and cross-reactivity between purified aCL and HCV were evaluated. Positive results for aCL antibodies were found more frequently (3. 3%) for the patients with HCV infection than for healthy controls (0%). All positive aCL antibodies were beta(2)-glycoprotein I independent. No significant association was found between aCL antibodies and clinical manifestations of APS, neither was one found between the presence of other autoantibodies or cryoglobulins and that of aCL. Finally, no cross-reactivity between aCL antibodies and HCV antigens was observed. As previously reported, aCL antibodies seem to be an epiphenomenon, and they do not have clinical or laboratory significance in HCV patients. PMID- 10702500 TI - Cocaine causes increased type I interferon secretion by both L929 cells and murine macrophages. AB - Cocaine has been demonstrated to have a number of different effects on immune cell functions. We have reported alterations of cellular functions by macrophages (Mphi) exposed to cocaine in vitro, including the inhibition of mouse hepatitis virus replication. Here, we present evidence that cocaine stimulates the secretion of an antiviral product that is neutralized by anti-interferon (anti IFN). A dose-dependent increase in the secretion of IFN by both Mphi and L929 cells incubated with cocaine, with a concomitant decrease in virus replication, is also reported. The increase in IFN secretion was most pronounced when cells were cultured in the presence of the IFN inducer poly(I.C). The effect of cocaine on IFN production was found to be primarily at the transcript level in both Mphi and L929 cells. These findings further support our previous research demonstrating an antiviral activity of cocaine in vitro. The relevance of this activity to viral infections in general remains to be determined. PMID- 10702501 TI - Conservation of the 17-kilodalton antigen gene within the genus Bartonella. AB - The 17-kDa antigen of Bartonella henselae has previously been shown to elicit a strong humoral immune response in patients with cat scratch disease (CSD) and to be useful in screening human serum samples for CSD. In this study, PCR amplification of genes homologous to the 17-kDa antigen gene of B. henselae was performed using genomic DNAs from several species of Bartonella, including the currently recognized human pathogens. Amplicons of similar size were demonstrated using the following chromosomal DNA templates: B. henselae (two strains), B. quintana (two strains), B. elizabethae, B. clarridgeiae, B. vinsonii subsp. vinsonii, and B. vinsonii subsp. berkhoffii. No evidence of a B. bacilliformis homolog of the 17-kDa antigen gene was obtained using multiple primer pairs. DNA sequencing revealed open reading frames capable of coding for proteins with sizes similar to that of the 17-kDa antigen of B. henselae in all of the amplicons; however, extensive sequence divergence across the genus was noted. Cloning of the amplified products into pUC19 resulted in recombinants that directed synthesis of homologs of the 17-kDa protein. Immunoblot analysis using human sera from CSD cases demonstrated very little cross-reactivity among different species for this protein. In contrast, immunoblots using rabbit serum raised to the recombinant B. henselae antigen showed extensive cross-reactivity with the proteins of other Bartonella species. The data suggest that the use of the 17-kDa antigen as a serologic reagent may allow the development of more specific diagnostic assays. Furthermore, the nucleotide sequences from the various versions of the 17-kDa antigen gene should be useful for rapid identification of Bartonella at the species level. PMID- 10702502 TI - Improved repetitive-element PCR fingerprinting of Salmonella enterica with the use of extremely elevated annealing temperatures. AB - Modified thermal cycling conditions were explored in an effort to improve the reproducibility and resolving power of repetitive-element PCR (rep-PCR) fingerprinting. Assay performance was rigorously evaluated under standard and modified cycling conditions, using as a test set 12 strains putatively representing 12 serovars of Salmonella enterica. For all three fingerprint types (ERIC2, BOXA1R, and composite fingerprints), the use of extremely elevated annealing temperatures plus an initial "touchdown" cycling routine yielded significant improvements in day-to-day reproducibility and discriminating power despite the somewhat sparser appearance of the fingerprints. Modified cycling conditions markedly reduced the variability of fingerprints between cyclers, allowing fingerprints from different cyclers to be analyzed together without the degradation of assay performance that occurred with between-cycler analyses under standard cycling conditions. With modified cycling, composite fingerprints exhibited the lowest reproducibility but the highest net discriminating power of the three fingerprint types. rep-PCR fingerprints led to the discovery of a serotyping error involving one of the 12 test strains. These data demonstrate that modified cycling regimens that incorporate elevated annealing temperatures (with or without an initial touchdown routine) may markedly improve the performance of rep-PCR fingerprinting as a bacterial typing tool. PMID- 10702503 TI - Improved repetitive-element PCR fingerprinting for resolving pathogenic and nonpathogenic phylogenetic groups within Escherichia coli. AB - Repetitive-element PCR (rep-PCR) fingerprinting is a promising molecular typing tool for Escherichia coli, including for discriminating between pathogenic and nonpathogenic clones, but is plagued by irreproducibility. Using the ERIC2 and BOXA1R primers and 15 E. coli strains from the ECOR reference collection (three from each phylogenetic group, as defined by multilocus enzyme electrophoresis [MLEE], including virulence-associated group B2), we rigorously assessed the effect of extremely elevated annealing temperatures on rep-PCR's reproducibility, discriminating power, and ability to reveal MLEE-defined phylogenetic relationships. Modified cycling conditions significantly improved assay reproducibility and discriminating power, allowing fingerprints from different cyclers to be analyzed together with minimal loss of resolution. The correspondence of rep-PCR with MLEE with respect to tree structure and regression analysis of distances was substantially better with modified than with standard cycling conditions. Nonetheless, rep-PCR was only a fair surrogate for MLEE, and when fingerprints from different days were compared, it failed to distinguish between different clones within all-important phylogenetic group B2. These findings indicate that although the performance and phylogenetic fidelity of rep PCR fingerprinting can be improved substantially with modified assay conditions, even when so improved rep-PCR cannot fully substitute for MLEE as a phylogenetic typing method for pathogenic E. coli. PMID- 10702504 TI - Transmission of human T-cell lymphotropic virus type 1 tax to rabbits by tax-only positive human cells. AB - The human T-cell lymphrotropic virus type 1 (HTLV-1) is causally related to adult T-cell leukemia and lymphoma and the neurodegenerative diseases tropical spastic paraparesis and HTLV-1-associated myelopathy. In the United States the prevalence of infection has been estimated to range from 0.016 to 0.1% on the basis of serologic tests for antibodies to the viral structural proteins. Blood from donors positive for antibodies to HTLV-1 or HTLV-2 is not used for transfusion. However, patients with the cutaneous T-cell lymphoma mycosis fungoides (MF) are HTLV-1 and -2 seronegative yet harbor proviral sequences identical to those that encode the HTLV-1 transactivating and transforming gene product p40tax in their peripheral blood mononuclear cells (PBMCs), and they usually have antibodies to p40(tax). Moreover, a study of 250 randomly selected blood donors revealed that approximately 8% of these seronegative individuals also had HTLV-1 tax sequences and antibodies to p40(tax), while they lacked sequences and antibodies related to gag, pol, or env. Thus, it seemed important to determine whether the "tax-only" state can be transmitted by transfusion. To this end, PBMCs from HTLV-1 and -2 seronegative tax-only-positive MF patients or from healthy tax-only-positive blood donors were injected into adult rabbits, an established animal model for HTLV-1 infection. The PBMCs of all injected rabbits became tax sequence positive. These observations suggest that HTLV-1 tax can be transmitted by tax-only positive mononuclear cells. PMID- 10702505 TI - CD8(+) T-cell gamma interferon production specific for human immunodeficiency virus type 1 (HIV-1) in HIV-1-infected subjects. AB - The CD8(+)-T-cell response to human immunodeficiency virus type 1 (HIV-1) is considered to be important in host control of infection and prevention of AIDS. We have developed a single-cell enzyme immunoassay (enzyme-linked immunospot assay) specific for gamma interferon (IFN-gamma) production stimulated by either autologous B-lymphoblastoid cell lines (B-LCL) infected with vaccinia virus vectors expressing HIV-1 proteins or synthetic peptides representing known HIV-1 CD8(+) cytotoxic T-lymphocyte (CTL) epitopes. Single-cell IFN-gamma production stimulated by HIV-1 Gag-, Pol-, and Env-expressing B-LCL was a reliable measure of HIV-1-specific T-cell immunity in peripheral blood CD8(+) T cells from HIV-1 infected individuals. This method was more sensitive than stimulation of IFN gamma by direct infection of the cultures with HIV-1-vaccinia virus vectors. Comparable results were found for IFN-gamma production in CD8(+) T cells from HIV 1-negative, cytomegalovirus (CMV)-seropositive, healthy donors stimulated with B LCL expressing the CMV pp65 lower matrix protein. HIV-1 peptides were immunodominant for both CD8(+) single-cell IFN-gamma production and CTL precursor frequencies. The number of cells producing IFN-gamma decreased in individuals with late-stage HIV-1 infection and was temporally enhanced during combination antiretroviral therapy with two reverse transcriptase nucleoside inhibitors and a protease inhibitor. PMID- 10702506 TI - Development, characterization, and diagnostic applications of monoclonal antibodies against bovine rotavirus. AB - Hybridomas secreting monoclonal antibodies (MAbs) against the Nebraska calf diarrhea strain of bovine rotavirus (BRV) were characterized. Indirect fluorescent-antibody assay, immunodot assay, and immunoprecipitation were used to select hybridomas that produced anti-BRV MAbs. Seven of the MAbs were shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot assay to be reactive with the BRV outer capsid protein, VP7, which has a molecular mass of 37.5 kDa. None of the seven MAbs were reactive with canine rotavirus, bovine coronavirus, or uninfected Madin-Darby bovine kidney cells. Two clones, 8B4 (immunoglobulin G2a [IgG2a]) and 2B11 (IgG1), were found suitable for use in an antigen capture enzyme-linked immunosorbent assay for detecting BRV in bovine fecal samples. Both were subtype A specific (G6 subtype) but did not react with all isolates of BRV group A. PMID- 10702507 TI - Antibody response to lipopolysaccharide in patients colonized or infected with an endemic strain of Acinetobacter genomic species 13 sensu Tjernberg and Ursing. AB - The levels of antilipopolysaccharide (anti-LPS) antibodies in patients colonized with an endemic Acinetobacter strain were compared to those in patients with bloodstream infections. Seropositivity and seronegativity correlated with positive and negative blood cultures, respectively, indicating that determination of the level of anti-LPS antibodies is useful for diagnosing Acinetobacter infections. PMID- 10702508 TI - Phagocytic function of monocytes in children with human immunodeficiency virus type 1 infection. AB - We investigated the phagocytic function of monocytes in 7- to 10-year-old children horizontally infected with human immunodeficiency virus type 1 (HIV-1) in comparison to that in healthy sex- and age-matched controls. CR3-mediated phagocytosis was increased in patients with HIV-associated pulmonary tuberculosis, independently of CD4 counts and p24 antigenemia. PMID- 10702509 TI - Identification of different states of hepatitis B virus infection with a quantitative PCR assay. AB - The level of hepatitis B virus (HBV) DNA in serum reflects the replicative activity of HBV. To compare serum HBV DNA levels in different states of hepatitis B, 47 sera of patients with HBeAg-positive chronic hepatitis B, 4 sera of patients with HBeAg-negative chronic hepatitis B, 40 samples of patients after HBeAg seroconversion during alpha interferon treatment, 57 sera of inactive HBsAg carriers, and 42 sera of patients who had recovered from chronic hepatitis B more than 12 months prior to blood collection were checked for the presence of HBV DNA with the Amplicor HBV Monitor Test. In patients with HBeAg-positive chronic hepatitis B, the median of serum HBV DNA levels (8.3 x 10(8) copies/ml) was significantly higher than that for patients after HBeAg seroconversion (6.2 x 10(3) copies/ml) and than that for inactive HBsAg carriers (5.6 x 10(3) copies/ml). None of the patients who had recovered from hepatitis B had detectable HBV DNA in serum. Quantitative PCR proved to be a valuable tool for identification of different states of HBV infection. This technique was found to be a good method for determination of serum HBV DNA levels both for patients with HBeAg seroconversion and for inactive carriers who showed low viremia not detectable by conventional hybridization assays. PMID- 10702510 TI - Clustering of clinical strains of Helicobacter pylori analyzed by two-dimensional gel electrophoresis. AB - Strain variations of Helicobacter pylori have been tested by numerous methods and compared among different patient groups. The aim of this study was to investigate whether H. pylori expresses disease-specific proteins that can be detected by two dimensional polyacrylamide gel electrophoresis (2-D PAGE). H. pylori strains isolated from duodenal ulcer, gastric cancer, and gastritis patients were analyzed. Extensive variation in spot patterns was observed between the strains, but a dendrogram analysis revealed that some strains within each disease group clustered together. Eight proteins were sequenced and found in the H. pylori genome sequence. 2-D PAGE is a useful method for studies of protein expression and for highlighting the extensive strain variation that H. pylori exhibits. PMID- 10702511 TI - Measurement of T-lymphocyte responses in whole-blood cultures using newly synthesized DNA and ATP. AB - The proliferative response is most frequently determined by estimating the amount of [(3)H]thymidine incorporated into newly synthesized DNA. The [(3)H]thymidine procedure requires the use of radioisotopes as well as lengthy periods of incubation (>72 h). An alternative method of assessing T-lymphocyte activation in whole-blood cultures involves the measurement of the nucleotide ATP instead of [(3)H]thymidine incorporation. In addition, the Luminetics assay of T-cell activation measures specific T-lymphocyte subset responses through the use of paramagnetic particles coated with monoclonal antibodies against CD antigens. This assay permits rapid (24 h) analysis of lymphocyte subset activation responses to mitogens and recall antigens in small amounts of blood. PMID- 10702512 TI - Clinical application of a dot blot test for diagnosis of enteric fever due to Salmonella enterica serovar typhi in patients with typhoid fever from Colombia and Peru. AB - Clinical application of a dot blot test to detect immunoglobulin G (IgG) (88% sensitivity and specificity) and IgM (12.1% sensitivity and 97% specificity) against flagellar antigen from Salmonella enterica serovar Typhi was performed in Peruvian and Colombian patients with typhoid fever. This test can be used as a good predictor of serovar Typhi infection in regions lacking laboratory facilities and in field studies. PMID- 10702513 TI - Comparison of polymorphonuclear cells from healthy donors and differentiated HL 60 cells as phagocytes in an opsonophagocytic assay using antigen-coated fluorescent beads. AB - Polymorphonuclear cells (PMNs) from healthy donors and differentiated HL-60 cells were compared in an opsonophagocytic assay using fluorescent latex beads coated with Streptococcus pneumoniae polysaccharide conjugates. Serum-specific phagocytosis was efficiently mediated by both sources of cells, as measured by flow cytometry, but the mean number of beads ingested per cell was three- to fivefold higher when PMNs were used than when HL-60 cells were used. Nevertheless, differentiated HL-60 cells could be a convenient and standardized source of cells to evaluate the functionality of specific antibodies to vaccine candidates as a coating on fluorescent beads. PMID- 10702514 TI - Identification of potentially diagnostic Leishmania braziliensis antigens in human cutaneous leishmaniasis by immunoblot analysis. AB - The antibody response in patients with American cutaneous leishmaniasis was analyzed by immunoblotting with soluble and insoluble antigens of Leishmania braziliensis. The recognition of the 27- and/or 30-kDa soluble antigens was considered relevant for the diagnosis of cutaneous leishmaniasis. Immunoblotting was found to be significantly more sensitive and specific than indirect immunofluorescence and enzyme-linked immunosorbent assay. PMID- 10702515 TI - Description of a nonlethal herpes simplex virus type 1 glycoprotein D deletion mutant affecting a site frequently used for PCR. AB - We report a herpes simplex virus type 1 mutant which failed to amplify with a commonly used glycoprotein D primer set. The virus contained a nine-base deletion in the gene's 5' nontranslated region. The altered amplicon was clearly distinguishable on a 4% high-resolution agarose gel. PMID- 10702516 TI - Real-time quantification of human telomerase reverse transcriptase mRNA in tumors and healthy tissues. AB - BACKGROUND: Expression of the hTERT gene, which codes for the catalytic subunit of telomerase, is associated with malignancy. We recently developed a real-time reverse transcription-PCR assay, based on TaqMan technology, for accurate and reproducible determination of hTERT mRNA expression (Lab Investig 1999;79:911-2). This method may be of interest for molecular tumor diagnostics in tissues and corresponding body fluids, washings, or brushes. METHODS: In this study, we measured hTERT expression in a subset of healthy tissues and tumors to select those tumor types with the best potential for quantification of hTERT in corresponding body fluids. To demonstrate the use of the method in body fluids, we quantified hTERT expression in voided urine of patients with bladder cancer and controls. RESULTS: Real-time measurement of hTERT expression could discriminate between all healthy and malignant tissue samples from pancreas, lung, esophagus, and bladder, but not for colon tissues. Moreover, in five of nine (55%) urine samples, hTERT could be quantified. CONCLUSIONS: The present study demonstrates that accurate quantitative measurement of hTERT expression has high potential for discrimination between healthy and tumor cells in tissues and urine and supports future measurements in pancreatic fluid, bronchoalveolar lavage fluid, esophageal brushings, and urine or bladder washings. PMID- 10702517 TI - Plasma DNA as a prognostic marker in trauma patients. AB - BACKGROUND: Recently, much interest has developed in the potential use of plasma DNA as a diagnostic and monitoring tool. We hypothesized that plasma DNA is increased in patients with trauma and may be prognostic in such patients. METHODS: We studied 84 patients who had sustained an acute blunt traumatic injury. We measured plasma DNA by a real-time quantitative PCR assay for the beta globin gene. Blood samples were collected at a median time of 60 min following injury. Blood samples were also obtained from 27 control subjects. RESULTS: The median plasma DNA concentrations in the control, minor/moderate trauma (Injury Severity Score <16; n = 47), and major trauma (Injury Severity Score > or =16; n = 37) groups were 3154 kilogenome-equivalents/L, 13 818 kilogenome-equivalents/L, and 181 303 kilogenome-equivalents/L, respectively. Plasma DNA concentrations in patients with adverse outcomes, including acute lung injury, acute respiratory distress syndrome, and death, had 11. 6- to 12-fold higher plasma DNA concentrations than those who did not develop these complications. At a cutoff of 232 719 kilogenome-equivalents/L, the sensitivities of plasma DNA analysis for the prediction of acute lung injury, acute respiratory distress syndrome, and death were 100% (95% confidence interval, 100-100%), 100% (95% confidence interval, 100-100%), and 78% (95% confidence interval, 40-97%), respectively. The respective specificities were 81% (95% confidence interval, 71-89%), 80% (95% confidence interval, 70-88%), and 82% (95% confidence interval, 71-90%). CONCLUSIONS: Plasma DNA is increased after trauma and may be a potentially valuable prognostic marker for these patients. PMID- 10702518 TI - Development of conventional and real-time PCR assays for the rapid detection of group B streptococci. AB - BACKGROUND: Group B streptococci (GBS), or Streptococcus agalactiae, are the leading bacterial cause of meningitis and bacterial sepsis in newborns. Currently available rapid methods to detect GBS from clinical specimens are unsuitable for replacement of culture methods, mainly because of their lack of sensitivity. METHODS: We have developed a PCR-based assay for the rapid detection of GBS. The cfb gene encoding the Christie-Atkins-Munch-Petersen (CAMP) factor was selected as the genetic target for the assay. The PCR primers were initially tested by a conventional PCR method followed by gel electrophoresis. The assay was then adapted for use with the LightCycler(TM). For this purpose, two fluorogenic adjacent hybridization probes complementary to the GBS-specific amplicon were designed and tested. In addition, a rapid sample-processing protocol was evaluated by colony-forming unit counting and PCR. A total of 15 vaginal samples were tested by both standard culture method and the two PCR assays. RESULTS: The conventional PCR assay was specific because it amplified only GBS DNA among 125 bacterial and fungal species tested, and was able to detect all 162 GBS isolates from various geographical areas. This PCR assay allowed detection of as few as one genome copy of GBS. The real-time PCR assay was comparable to conventional PCR assay in terms of sensitivity and specificity, but it was more rapid, requiring only approximately 30 min for amplification and computer-based data analysis. The presence of vaginal specimens had no detrimental effect on the sensitivity of the PCR with the sample preparation protocol used. All four GBS positive samples identified by the standard culture method were detected by the two PCR assays. CONCLUSION: These assays provide promising tools for the rapid detection and identification of GBS. PMID- 10702519 TI - Development and evaluation of three immunofluorometric assays that measure different forms of osteocalcin in serum. AB - BACKGROUND: Circulating human osteocalcin (hOC) has been used as a marker of bone formation. Our aim was to validate three immunofluorometric assays (IFMAs), measuring different forms of hOC. METHODS: The two-site IFMAs were based on previously characterized monoclonal antibodies. Assay 2 recognized intact hOC, assays 4 and 9 measured the NH(2)-terminal mid-fragment and the intact hOC. In addition, assay 9 required hOC to be gamma-carboxylated. RESULTS: A 76-79% increase of serum immunoreactive hOC was found in the postmenopausal group compared with the premenopausal group with all IFMAs. With EDTA-plasma samples, the observed increases were lower (49-65%). The hOC concentration in the postmenopausal group receiving hormone replacement therapy was 42-44% lower than that in the postmenopausal control group in both serum and EDTA-plasma samples. The depressed carboxylation in warfarin-treated patients was accompanied by lower results in assay 9. The ratio of assay 9 to assay 4 totally discriminated the warfarin-treated patients from the controls. Assay 9 showed the smallest decreases in measured hOC after storage of serum or plasma for 4 weeks at 4 degrees C, followed by assay 4 and assay 2. Results from the last assay were <17% of their initial values after 4 weeks of storage. No diurnal variation was observed with assay 9 as opposed to the two other IFMAs. CONCLUSION: The three assays with their distinct specificity profiles (intact vs fragmented and carboxylated vs decarboxylated hOC) may provide valuable tools for investigating the significance of different hOC forms in various bone-related diseases. PMID- 10702520 TI - Correlation of antemortem serum creatine kinase, creatine kinase-MB, troponin I, and troponin T with cardiac pathology. AB - BACKGROUND: Spurious increases in serum troponins, especially troponin T, have been reported in patients with and without acute myocardial syndromes. METHODS: We studied 78 autopsied patients without clinical myocardial infarction (MI) and correlated histologic cardiac findings with antemortem serum creatine kinase (CK), its MB isoenzyme (CK-MB), cardiac troponin I (cTnI), and cardiac troponin T (cTnT). RESULTS: There was no significant myocardial pathology in 15 patients. Cardiac pathologies were in five groups: scarring from previous MI or patchy ventricular fibrosis (n = 9), recent MI (n = 27), healing MI (n = 7), degenerative myocyte changes consistent with congestive heart failure (CHF; n = 12), and other cardiac pathologies (n = 8). The median concentrations in the five groups were not significantly different for either CK or CK-MB. Compared with the no-pathology group, only the MI group was significantly different for cTnI, and the MI and other pathology groups were significantly different for cTnT. For patients with MI, 22%, 19%, 48%, and 65% had increased CK, CK-MB, cTnI, and cTnT, respectively; for CHF and other cardiac pathologies combined, the percentages were 28%, 17%, 22%, and 50%. For patients with increased cTnI, 72% and 28% had MI and other myocardial pathologies, respectively; patients with increased cTnT had 64% and 36%, respectively. Patients without myocardial pathology had no increases in CK-MB, cTnI, or cTnT. CONCLUSIONS: All patients with increased serum CK-MB, cTnI, and cTnT had significant cardiac histologic changes. The second-generation cTnT assay appears to be a more sensitive indicator of MI and other myocardial pathologies than the cTnI assay used in this study. PMID- 10702521 TI - Development of a new assay for complex I of the respiratory chain. AB - BACKGROUND: Measurement of complex I activity has been hampered by the large amounts of tissue required and the resulting turbidity of the assay solution, which makes spectrophotometric analysis difficult. We have developed a new assay for measuring the activity of complex I in isolated mitochondria that is also applicable to skeletal muscle homogenate in patients with suspected mitochondrial diseases. METHODS: The method was a radioenzymatic assay based on the preferential oxidation of the 4B hydrogen of NADH by complex I. We prepared tritiated isoforms of NADH for both the respective 4A-(3)H and 4B-(3)H positions. Enzyme in the form of purified mitochondria or homogenate was prepared from rat or human skeletal muscle and incubated with the respective radioisotopes. The product ((3)H(2)O) was collected after charcoal adsorption of unreacted NADH and taken as an indicator of NADH oxidation. Sensitivity to rotenone was used as a measure of complex I specific activity. RESULTS: The assay was linear with time and protein for isolated mitochondria and tissue homogenates from rats and humans. The V(max) and K(m) values obtained for 4B-NADH with isolated rat skeletal muscle mitochondria were 35 micromol/L and 90 micromol. min(-1). mg protein(-1), respectively. The assay was reproducible and usable for routine measurements in human skeletal muscle. The sensitivity was >10-fold higher than the sensitivities of spectrophotometric techniques. CONCLUSIONS: The results of our studies demonstrate the successful development of a new assay for complex I that is rapid, easy to perform, and that enables the processing of multiple samples at one time. PMID- 10702522 TI - Estimating the long-term effects of storage at -70 degrees C on cholesterol, triglyceride, and HDL-cholesterol measurements in stored sera. AB - We estimated the effects of long-term storage at -70 degrees C on serum total cholesterol, HDL-cholesterol, and triglycerides in specimens that had been stored for up to 7 years. These estimates were made using measurements in serial specimens collected from the placebo control group of the Air Force/Texas Coronary Atherosclerosis Prevention Study over a period of approximately 5 years. We compared the group means for pairs of serial specimens taken at 6- and 12 month intervals, assuming that (a) a negligible placebo effect occurred between the serial specimen pairs; (b) in the absence of storage effects, the variation in the group means would reflect only normal biological variation and would not materially affect the group means for the serial specimens; (c) any systematic changes in these group means would reflect storage-related changes; and (d) storage-related changes are cumulative, i.e., the overall changes for a given storage period are the sum of the changes during previous storage periods. We observed average decreases of 2.0% per year for total cholesterol over 7 years and 2.8% per year in triglycerides for the first 5 years. HDL-cholesterol decreased by 1.3% per year, but this change was not statistically significant. This approach may be useful for estimating storage-related changes for studies in specimens stored for a period of years and for which stability data may not be available. PMID- 10702523 TI - Determination of the acyl glucuronide metabolite of mycophenolic acid in human plasma by HPLC and Emit. AB - BACKGROUND: The acyl glucuronide (AcMPAG) of mycophenolic acid (MPA) has been found to possess pharmacologic and potentially proinflammatory activity in vitro. To establish its pharmacologic and toxicologic relevance in vivo, a reversed phase HPLC method was modified to simultaneously determine MPA, the phenolic MPA glucuronide (7-O-MPAG), and AcMPAG. In addition, cross-reactivity of AcMPAG in the Emit assay for MPA was investigated. METHODS: The procedure used simple sample preparation, separation with a Zorbax Eclipse-XDB-C8 column, and gradient elution. AcMPAG was quantified as 7-O-MPAG-equivalents. RESULTS: The assay was linear up to 50 mg/L for MPA, 250 mg/L for 7-O-MPAG, and 10 mg/L for AcMPAG (r >0.999). Detection limits were 0.01, 0.03, and 0.04 mg/L for MPA, 7-O-MPAG, and AcMPAG, respectively. The recoveries were 99-103% for MPA, 95-103% for 7-O-MPAG, and 104-107% for AcMPAG. The within-day imprecision was <5.0% for MPA (0.2-25 mg/L), <4.4% for 7-O-MPAG (10-250 mg/L), and < or =14% for AcMPAG (0.1-5 mg/L). The between-day imprecision was <6.2%, <4.5%, and < or =14% for MPA, 7-O-MPAG, and AcMPAG, respectively. When isolated from microsomes, purified AcMPAG (1-10 mg/L) revealed a concentration-dependent cross-reactivity in an Emit assay for the determination of MPA ranging from 135% to 185%. This is in accordance with the bias between HPLC and Emit calculated in 270 samples from kidney transplant recipients receiving mycophenolate mofetil therapy, which was greater (median, 151.2%) than the respective AcMPAG concentrations determined by HPLC. AcMPAG was found to undergo hydrolysis when samples were stored up to 24 h at room temperature or up to 30 days at 4 degrees C or -20 degrees C. Acidified samples (pH 2.5) were stable up to 30 days at -20 degrees C. CONCLUSIONS: The HPLC and Emit methods for AcMPAG described here may allow investigation of its relevance for the immunosuppression and side effects associated with mycophenolate mofetil therapy. PMID- 10702524 TI - Detection and measurement of urinary 2-hydroxyestradiol 17-sulfate, a potential placental antioxidant during pregnancy. AB - BACKGROUND: Preeclampsia is associated with a quantitative imbalance between lipid peroxide and an antioxidant coproduced in the placenta. To investigate our hypothesis that 2-hydroxyestradiol 17-sulfate (2-OH-ES) is the placental antioxidant during pregnancy, we developed an assay for 2-OH-ES in urine and studied samples from women with and without preeclampsia. METHODS: The detection and measurement of 2-OH-ES in the urine of pregnant women were performed by RIA using highly specific antiserum to 2-OH-ES. To confirm the reliability of the RIA method, the same samples were analyzed by HPLC using an electrochemical detector. RESULTS: Urinary 2-OH-ES values obtained by RIA showed a close relationship to those obtained by HPLC (y = 1.1x - 0.01; r = 0.96). The urinary 2-OH-ES concentrations during the first, second, and third trimesters were 2. 0 +/- 0.6 (mean +/- SE, n = 13), 5.3 +/- 1.3 (n = 21), and 15.3 +/- 2.0 microg/mg creatinine (n = 54), respectively, and <0.15 microg/mg creatinine (n = 10) at 2 24 h after delivery. The concentrations in preeclamptic women during the third trimester were significantly lower, 3.9 +/- 1.9 microg/mg creatinine (mean +/- SE, n = 12). CONCLUSIONS: RIA can be used to measure urinary 2-OH-ES during pregnancy. The increase in urinary 2-OH-ES during gestation, its decrease after delivery, and the lower values in preeclampsia are consistent with a role of 2-OH ES as a placental antioxidant. PMID- 10702525 TI - Leptin binding and binding capacity in serum. AB - BACKGROUND: Leptin, a hormone produced primarily by adipose tissue, is known to be present in serum as both monomeric (free) and higher molecular mass (bound) forms, but little is known about the nature of the bound forms or physiological variation in binding capacity. METHODS: A new method to quantify the free and bound forms was developed, based on HPLC separation and RIA quantification in chromatography fractions. Reanalysis of specimens after addition of exogenous leptin allowed direct determination of leptin-binding capacity and the degree of saturation of leptin-binding capacity. RESULTS: HPLC chromatography fractionated serum leptin into both the free form and as a broad peak of 59-130 kDa. Several experiments were conducted to validate the new method. The concentrations of bound leptin in serum were 0.45-3.94 micro/L, and they increased as total leptin (reflecting adiposity) increased in 24 lean and obese volunteers. Leptin was readily dissociated from the bound fraction by competition from exogenous leptin. Rechromatography of the bound fraction led to dissociation of leptin, which was promoted by warming the sera before chromatography. Leptin-binding capacity was 1.8-5.3 microg/L; binding capacity was nearly constant over a range of total leptin concentrations of 2-10 microg/L, and slowly increased at higher total leptin concentrations. Saturation of binding capacity was low (15%) at very low total leptin concentrations (<5 microg/L), but rose quickly to a plateau near 80% at higher total leptin concentrations. CONCLUSIONS: The new method facilitates measurement of free and bound fractions of serum leptin, and is the first method measuring leptin-binding capacity. These experiments demonstrate that the concentration of bound leptin and leptin-binding capacity vary physiologically, with binding/binding capacity increasing with adiposity. Except in very lean individuals, binding capacity is nearly completely saturated. PMID- 10702527 TI - Total protein determination in urine: elimination of a differential response between the coomassie blue and pyrogallol red protein dye-binding assays. AB - BACKGROUND: The total protein content of urine is a good index of renal function, but its determination is unreliable. Protein dye-binding assays are simple, but they characteristically lack a uniform response to different proteins. METHODS: We investigated a differential response of the Sigma Microprotein Coomassie Brilliant Blue (CBB) and Pyrogallol Red-molybdate (PRM) protein dye-binding assays to urine, using human albumin, albumin/globulin, or urinary protein as calibrator. RESULTS: The urine protein values (n = 60) obtained with the CBB assay were 110-13 500 mg/L (mean, 2390 mg/L) compared with 160-18 300 mg/L (mean, 3470 mg/L) obtained with the PRM assay (CBB:PRM protein concentration ratio, 0.46 0.88, mean, 0. 69 +/- 0.10). The differential response was highly reproducible as indicated by Sigma urine control Level 1 (within-day CBB:PRM ratio, 0.68 +/- 0.02; between-day CBB:PRM ratio, 0.67 +/- 0.04) and Sigma urine control Level 2 (within-day CBB:PRM ratio, 0.60 +/- 0.01; between-day CBB:PRM ratio, 0.59 +/- 0.02). The use of urinary protein as a calibrator (rather than human albumin) greatly improved the agreement between the assays when applied to urine (y(CBB) = 0. 972x(PRM) - 16 vs y(CBB) = 0.685x(PRM) + 17). In studies using urine controls, this calibrator also improved agreement between the CBB, PRM, trichloroacetic acid (TCA), and benzethonium chloride protein methods and, to a lesser extent, agreement with the TCA-Ponceau S method. CONCLUSION: The use of a urinary protein calibrator improves the agreement between different methods used to determine total protein in urine. PMID- 10702526 TI - Correlation between plasma total homocysteine and copper in patients with peripheral vascular disease. AB - BACKGROUND: Increased concentrations of both plasma total homocysteine and copper are separately associated with cardiovascular disease. Correlations between plasma total homocysteine, trace elements, and vitamins in patients with peripheral vascular disease have not been investigated. METHODS: The concentrations of trace elements in plasma were determined by the multielement analytical technique of total-reflection x-ray fluorescence spectrometry. Plasma total homocysteine was determined by HPLC. RESULTS: In the univariate and multivariate regression analyses, copper was positively correlated with plasma total homocysteine in all subjects (coefficient +/- SE, 0.347 +/- 0.113; P = 0.0026 and coefficient +/- SE, 0.422 +/- 0.108; P = 0.0002, respectively), and in patients with peripheral vascular disease (coefficient +/- SE, 0.370 +/- 0.150; P = 0.016; and coefficient +/- SE, 0.490 +/- 0.151; P = 0.0025, respectively). Correlation between copper and plasma total homocysteine was not detected in healthy control subjects. The concentration of calcium in plasma (67.5 vs 80. 8 microg/g) was significantly lower in the patients than in the control subjects (P = 0.02). When the patients were divided into groups, the patients with suprainguinal lesions had significantly higher copper concentrations (P = 0.04) and significantly lower selenium and calcium concentrations (P = 0.01 and 0.008, respectively) than the healthy subjects. Patients had higher concentrations of autoantibodies against oxidized LDL and concentrations of thiobarbituric acid reactive substance than the healthy subjects (P <0.0001 and P = 0.001, respectively). The concentrations of plasma total homocysteine and alpha tocopherol were significantly higher, and the concentrations of vitamin B(6) and beta-carotene were lower in the patients than the healthy subjects. CONCLUSION: Our findings suggest that the atherogenicity of homocysteine may be related to copper-dependent interactions. PMID- 10702528 TI - Cerebrospinal fluid protein concentrations in children: age-related values in patients without disorders of the central nervous system. AB - BACKGROUND: The published reference values for cerebrospinal fluid (CSF) total protein concentrations in children suffer from two major drawbacks: (a) the age related range often is too broad when applied to the steeply falling concentrations in early infancy; and (b) no values have been published for widely used dry chemistry methods. METHODS: We conducted a 2-year retrospective survey of CSF results obtained in a children's hospital with a dry chemistry-based method set up on the Vitros 700 analyzer. RESULTS: The data related to ambulatory children up to 16 years of age and term neonates with no clinical or biological signs of brain disease (n = 1074). Seven age groups with significantly different CSF protein values were identified, and their age-related percentiles (5th, 50th, and 95th) were determined. On the basis of the upper 95th percentile, from age 0 to 6 months the CSF protein concentrations fell rapidly from 1.08 to 0.40 g/L. A plateau (0.32 g/L) was reached from age 6 months to 10 years, followed by a slight increase (0.41 g/L) in the 10-16 years age range. CONCLUSIONS: These results imply that CSF total protein concentrations in the pediatric setting, particularly in infants, must always be interpreted with regard to narrow age related reference values to avoid false-positive results. PMID- 10702529 TI - Analysis of folate form distribution by affinity followed by reversed- phase chromatography with electrical detection. AB - BACKGROUND: Naturally occurring folates exist in multiple forms, differing in pteridine ring structure and number of glutamate residues. The ability to measure these folate coenzymes in tissues and cells gives important information about in vivo folate metabolism. METHODS: Folates were heat-extracted from biological samples. A two-column HPLC system with four-channel coulometric electrochemical detection was used for analysis. An affinity column was used first to purify folates from the extract. Purified folates were eluted from the affinity column onto a phenyl analytical column, utilizing a switching valve, and folate forms were separated using an acetonitrile gradient. RESULTS: Folate forms differing in pteridine ring structure and number of glutamate chain residues were identified by retention time and characteristic response across the channels of the detector. Folates were quantified by comparison to an external calibration mixture. Limits of detection for pentaglutamyl folates ranged from 0.21 pmol for tetrahydrofolate to 0.41 pmol for 5-methyltetrahydrofolate. CVs (n = 5) for peaks containing 9-67 pmol of folate were 0.6-6.4% (within day) and 5.2-8. 4% (between days). CVs (n = 5) for peaks containing 0.9-3.5 pmol folate were 5.7-16% (within day) and 8.4-13% (between days). CONCLUSIONS: This automated HPLC system allows the simultaneous determination of polyglutamyl forms of folates from biological samples, including tetrahydrofolate, 5-methyltetrahydrofolate, formylated folates, and pteroylglutamate. The low detection limits allow analysis of folate form distribution in human samples such as erythrocytes and lymphocytes. PMID- 10702530 TI - Lectin immunoassay for macrophage-activating factor (Gc-MAF) produced by deglycosylation of Gc-globulin: evidence for noninducible generation of Gc-MAF. PMID- 10702531 TI - Technical and clinical validation of an immunoradiometric assay for circulating parathyroid hormone-related protein. PMID- 10702532 TI - Intermethod discordant free thyroxine measurements in bone marrow-transplanted patients. PMID- 10702533 TI - Determination of plasma serotonin and 5-hydroxyindoleacetic acid in healthy subjects and cancer patients. PMID- 10702534 TI - Melting temperature assay for a UGT1A gene variant in Gilbert syndrome. PMID- 10702535 TI - Rapid beta-globin genotyping by multiplexing probe melting temperature and color. PMID- 10702536 TI - Procalcitonin is not a reliable marker for the assessment of severity in acute pancreatitis without infectious complications. PMID- 10702537 TI - Premetrological variation of thyrotropin, thyroxine (non-protein bound), and triiodothyronine concentrations in serum. PMID- 10702538 TI - Circadian variation in serum CrossLaps concentration is reduced in fasting individuals. PMID- 10702539 TI - Response to "Increased creatine kinase MB and cardiac troponin T with normal cardiac troponin I in metastatic alveolar rhabdomyosarcoma". PMID- 10702540 TI - Sensitivity of assays designed for the detection of disseminated epithelial tumor cells is influenced by cell separation methods. PMID- 10702542 TI - Compiled by david E. Bruns editor (dbruns@aacc.org) PMID- 10702541 TI - Is lipoprotein(a) cholesterol a significant indicator of cardiovascular risk? PMID- 10702543 TI - Management of intravascular catheter-related infections. PMID- 10702544 TI - In vitro antibacterial activity and mechanism of action of J-111,225, a novel 1beta-methylcarbapenem, against transferable IMP-1 metallo-beta-lactamase producers. AB - IMP-1 beta-lactamase, a class B zinc metallo-enzyme encoded by the transferable bla(IMP) gene, is known to confer high-level resistance to carbapenems as well as to penicillins and cephalosporins. J-111, 225 is a novel 1beta-methylcarbapenem with a structurally unique side chain comprising a trans-3,5-disubstituted pyrrolidinylthio moiety at the C2 position. It inhibited 17 Serratia marcescens and two Pseudomonas aeruginosa IMP-1-producing clinical isolates at a concentration of 32 mg/L (range 4-32 mg/L). It showed synergy with imipenem against IMP-1-producing S. marcescens BB5886 and P. aeruginosa GN17203 with minimal FIC indices of 0.38 and 0.5, respectively. J-111,225 was more resistant than imipenem to hydrolysis by class B metallo-beta-lactamases. In kinetic studies, J-111,225 inhibited the IMP-I enzyme with a K(i) of 0.18 microM when imipenem was used as a substrate. In contrast, J-111,225 was the substrate for hydrolysis by other class B beta-lactamases such as Bacteroides fragilis CcrA, Stenotrophomonas maltophilia L1 and Bacillus cereus type II enzyme with respective K(m) values of 11, 10 and 148 microM. The greater antibacterial activity of J-111,225 against IMP-1-producing bacteria may result from its unique interaction with the beta-lactamase. PMID- 10702545 TI - Diversity among high-level aminoglycoside-resistant enterococci. AB - A total of 55 Enterococcus faecalis and 21 Enterococcus faecium non-replicate isolates were obtained from routine clinical specimens, during a 1 year period, in a tertiary care hospital in Athens, Greece. The most common isolation site was the urinary tract (44% of E. faecalis and 33% of E. faecium isolates). No vancomycin resistance was detected. Ampicillin-resistant isolates did not produce beta-lactamase. High-level gentamicin resistance was detected in 22% and 0% of E. faecalis and E. faecium isolates, respectively. The corresponding figures for high-level streptomycin resistance were 40% and 33%. The aminoglycoside-modifying enzyme gene aac(6')+aph(2") was detected by PCR in 10 of 12 high-level gentamicin resistant E. faecalis isolates, and the ant(6)-I gene in all high-level streptomycin-resistant isolates of both species. DNA fingerprinting by PFGE grouped 31 of 55 E. faecalis isolates into 10 clusters, and 10 of 21 E. faecium isolates into two clusters, containing two to seven isolates each. Two E. faecalis PFGE types, comprising isolates expressing high-level aminoglycoside resistance, and not observed among non-high-level aminoglycoside-resistant strains, were disseminated in building A of the hospital. In contrast, high-level aminoglycoside resistance seemed to have been acquired nosocomially by a number of genotypically different E. faecium types. Molecular typing was therefore instrumental in understanding the differences in the mode of spread and acquisition of high-level aminoglycoside resistance among these two different enterococcal species. PMID- 10702546 TI - A comparison of antimicrobial resistance rates in Gram-positive pathogens isolated in the UK from October 1996 to January 1997 and October 1997 to January 1998. AB - Rates of resistance for two consecutive years for 28 centres (10 Teaching, nine Associate Teaching and nine District General hospitals) in the UK were compared. Combined rates of resistance for each of the hospital types of Staphylococcus aureus to methicillin revealed an increase in the rate of resistance in Teaching hospitals (12.5% year 1, 23.5% year 2), but, for Associate Teaching and District General hospitals rates fell (Associate Teaching 19.1% year 1, 11.9% year 2; District General 16.5% year 1 and 11.3% year 2). Using conventional methodology to determine MICs, no strain was considered to have reduced susceptibility to vancomycin. Among coagulase-negative staphylococci, increased resistance was observed for Staphylococcus epidermidis to rifampicin, for Staphylococcus haemolyticus to clindamycin, for Staphylococcus saprophyticus to penicillin and for Staphylococcus spp. to clindamycin, methicillin and rifampicin. For Streptococcus pneumoniae an upward trend in low-level resistance to penicillin was observed (18 of the 28 centres), however, for high-level resistance the trend was in the opposite direction (only four centres showed an increase). For Enterococcus faecalis there was a trend to a fall in levels of resistance, the only exception being an increase in high-level gentamicin resistance (10.5% year 1, 15.1% year 2, P = 0.0388). For Enterococcus faecium rates of resistance were not significantly different except for increases in resistance to nitrofurantoin and rifampicin. PMID- 10702547 TI - Activity and spectrum of 22 antimicrobial agents tested against urinary tract infection pathogens in hospitalized patients in Latin America: report from the second year of the SENTRY antimicrobial surveillance program (1998). AB - The potency and spectrum of various antimicrobial agents tested against 434 bacterial isolates causing urinary tract infection (UTI) in hospitalized patients in Latin America were evaluated. The genotypes of the extended-spectrum beta lactamase-producing and selected multi-resistant isolates were also evaluated by molecular typing techniques. Escherichia coli (60.4%) was the most common aetiological agent causing UTI, followed by Klebsiella spp. (11.2%) and Pseudomonas aeruginosa (8.3%). In contrast, Enterococcus spp. isolates caused only 2.3% of UTIs. Fewer than 50% of E. coli isolates were susceptible to broad spectrum penicillins. The resistance rates to ciprofloxacin and the new quinolones were also high among these isolates. The molecular characterization of ciprofloxacin-resistant E. coli showed that most of them have a double mutation in the gyrA gene associated with a single mutation in the parC gene. The Klebsiella pneumoniae isolates studied demonstrated high resistance rates to beta lactam drugs, including broad-spectrum cephalosporins. The carbapenems were the compounds with the highest susceptibility rate among these isolates (100.0% susceptible) followed by cefepime (91.7% susceptible). Meropenem, imipenem and cefepime were also the most active drugs against Enterobacter spp. Among P. aeruginosa isolates, meropenem (MIC(50), 2 mg/L) was the most active compound, followed by imipenem (MIC(50), 4 mg/L), cefepime (MIC(50), 8 mg/L) and ceftazidime (MIC(50), 16 mg/L). The results presented in this report confirm that bacterial resistance continues to be a great problem in Latin American medical institutions. PMID- 10702548 TI - In vitro susceptibilities of Rickettsia and Bartonella spp. to 14-hydroxy clarithromycin as determined by immunofluorescent antibody analysis of infected vero cell monolayers. AB - The in vitro susceptibilities of Rickettsia akari, Rickettsia conorii, Rickettsia prowazekii, Rickettsia rickettsii, Bartonella elizabethae, Bartonella henselae and Bartonella quintana to different concentrations of clarithromycin, 14-hydroxy clarithromycin (the primary metabolite of clarithromycin) and tetracycline in Vero cell cultures, were determined by enumeration of immunofluorescently-stained bacilli. The extent of antibiotic-induced inhibition of foci was recorded for each dilution of antibiotic and compared with an antibiotic-negative control. Based upon MIC data, clarithromycin alone is highly active against all three Bartonella spp., R. akari and R. prowazekii, while 14-hydroxy-clarithromycin is active against R. conorii, R. prowazekii and R. rickettsii. Further testing is warranted in animal models and human clinical trials, to examine the activity of both clarithromycin and its primary metabolite and to define further the role of clarithromycin in therapy, particularly of infections caused by obligate intracellular bacteria such as Rickettsia and Bartonella spp. PMID- 10702549 TI - Therapeutic effects of parenteral beta-lactam antibiotics on experimental otitis media caused by penicillin-resistant Streptococcus pneumoniae in guinea-pigs. AB - The therapeutic effects of parenteral beta-lactam antibiotics were evaluated in experimental acute otitis media caused by penicillin-resistant Streptococcus pneumoniae (PRSP) in guinea-pigs. Cefotaxime, ceftriaxone and piperacillin significantly reduced viable cell counts of PRSP in the middle ear at a dose of 50 mg/kg bd for 3 days (P < 0.01 compared with control). The therapeutic effects of cefotaxime, ceftriaxone and piperacillin were superior to those of cefotiam and ceftazidime. These therapeutic effects reflected both in vitro activity and pharmacokinetic properties of the drugs. PMID- 10702550 TI - Evaluation of combined ceftriaxone and dexamethasone therapy in experimental cephalosporin-resistant pneumococcal meningitis. AB - The treatment of meningitis caused by strains of Streptococcus pneumoniae with decreased susceptibility to third-generation cephalosporins is an increasingly frequent and difficult problem. In this study a rabbit model of meningitis was used to determine the efficacy of ceftriaxone at different dosages, and to establish the effect of the addition of dexamethasone to the chemotherapeutic regimen. Groups of eight rabbits were inoculated with 10(6) cfu/mL of a cephalosporin- resistant strain of S. pneumoniae (MIC of cefotaxime/ceftriaxone 2 mg/L). Eighteen hours after inoculation, ceftriaxone (50 or 100 mg/kg/day) with or without dexamethasone (0. 25 mg/kg/ day) was administered for a period of 48 h. The ceftriaxone dose of 50 mg/kg/day was not fully effective in this model (therapeutic failure rate 28%). With a dose of 100 mg/kg/day there were no therapeutic failures and all CSF cultures were below the level of detection at 48 h. CSF ceftriaxone concentrations, area under the time-concentration curve and time above the MIC were not significantly different with or without dexamethasone. However, concomitant use of dexamethasone resulted in higher CSF bacterial counts and a higher number of therapeutic failures (57% with the 50 mg/kg/day dose and 28% with the 100 mg/kg/day dose). Increasing doses of ceftriaxone might be an effective mode of therapy for meningitis caused by S. pneumoniae with MIC , T(ss) > MIC, f2.gif" BORDER="0"> and AUIC(ss) were calculated for each clinical case included in the study, from simulated plasma level curves corresponding to the dosage regimen administered. A univariate correlation analysis was performed considering efficacy (%) as the dependent variable and indices as the independent variables according to linear and non-linear pharmacokinetic-pharmacodynamic models (PK-PD models). The results prove that log transformation of the independent variable improves the data fitting to linear model. The four estimated indices show a log-linear relationship with outcome, T(ss) > MIC and AUIC(ss) being the parameters best correlated with percentage efficacy. The E(max) model with intrinsic response is an additional correlation strategy for T(ss) > MIC, leading to estimated values of E(max) and E(0) of 100.34 +/- 25.09% and 24.40 +/- 11.7%, respectively. The wide range of bacteria responsible for the infections considered, including Gram-positive pathogens such as staphylococci, might explain the good correlation between T(ss) > MIC and percentage efficacy found for ciprofloxacin in this study. PMID- 10702552 TI - High vancomycin dosage regimens required by intensive care unit patients cotreated with drugs to improve haemodynamics following cardiac surgical procedures. AB - The aim of this study was to evaluate retrospectively the importance of a Bayesian pharmacokinetic approach for predicting vancomycin concentrations to individualize its dosing regimen in 18 critically ill patients admitted to intensive care units following cardiothoracic surgery. The possible influence of some coadministered drugs with important haemodynamic effects (dopamine, dobutamine, frusemide) on vancomycin pharmacokinetics was assessed. Vancomycin serum concentrations were measured by fluorescence polarization immunoassay. Vancomycin dosage regimens predicted by the Bayesian method (D(a)) were compared retrospectively with Moellering's nomogram-based dosages (D(M)) to assess possible major differences in vancomycin dosing. D(a) values were similar to D(M) in 10 patients (D(a) approximately D(M) group) (20.52 +/- 8.40 mg/kg/day versus 18.81 +/- 7.24 mg/kg; P = 0.15), whereas much higher dosages were required in the other eight patients (D(a) >> D(M) group) (26.78 +/- 3.01 mg/kg/day versus 18.95 +/- 3.41 mg/kg/day; P < 0.0001) despite no major difference in attained vancomycin steady-state trough concentration (C(min ss)) (9.22 +/- 1. 33 mg/L versus 8.99 +/- 1.26 mg/L; = 0.75) or estimated creatinine clearance (1.23 +/- 0.49 mL/min/kg versus 1.21 +/- 0.24 mL/min/kg; P = 0.95) being found between the two groups. The ratio between D(a) and D(M) was significantly higher in the D(a) >> D(M) group than in the D(a) approximately D(M) group (1.44 +/- 0.18 versus 1.10 +/- 0. 21; P < 0.01). In four D(a) >> D(M) patients the withdrawal of cotreatment with haemodynamically active drugs was followed by a sudden substantial increase in the vancomycin C(min ss) (13.30 +/- 1. 13 mg/L versus 8.79 +/- 0.87 mg/L; P < 0.01), despite no major change in bodyweight or estimated creatinine clearance being observed. We postulate that these drugs with important haemodynamic effects may enhance vancomycin clearance by inducing an improvement in cardiac output and/or renal blood flow, and/or by interacting with the renal anion transport system, and thus by causing an increased glomerular filtration rate and renal tubular secretion. Given the wide simultaneous use of vancomycin and dopamine and/or dobutamine and/or frusemide in patients admitted to intensive care units, clinicians must be aware of possible subtherapeutic serum vancomycin concentrations when these drugs are coadministered. Therefore, therapeutic drug monitoring (TDM) for the pharmacokinetic optimization of vancomycin therapy is strongly recommended in these situations. PMID- 10702553 TI - Pharmacokinetics and burn eschar penetration of intravenous ciprofloxacin in patients with major thermal injuries. AB - Adequate penetration of antibiotics into burn tissue and maintenance of effective serum levels are essential for the treatment of patients sustaining major thermal injuries. The pharmacokinetics and burn eschar penetration of intravenous ciprofloxacin were determined in 12 critically ill patients with burn injuries. Mean age for the 12 patients was 45 +/- 17 (range 25-82 years), total body surface area burned (TBSAB) = 38 +/- 15% and Acute Physiology and Chronic Health Evaluation (APACHE) II score = 8 +/- 6. Patients received recommended doses of ciprofloxacin, 400 mg q12h iv, for three doses beginning 72 h post-burn. Serum concentrations were measured at t = 0, 0.25, 0.5, 0.75, 1.0, 1.25, 1.5, 2.0, 4.0 and 12.0 h after the first and third doses. Burn eschar biopsies were obtained after the third ciprofloxacin dose. Three of these 12 patients (25%) manifested later signs of clinical sepsis (TBSAB = 61 +/- 6% and APACHE II score = 11 +/- 3) and underwent a second infusion of three doses of intravenous ciprofloxacin, blood sampling and eschar biopsy. Serum and eschar concentrations were determined by high performance liquid chromatography. Serum ciprofloxacin concentrations were comparable to those of normal volunteers (C(max) = 4.0 +/- 1 mg/L and AUC = 11.4 +/- 2 mg.h/L) during the immediate post-burn period after dose 1 (C(max1) = 4.8 +/- 3 mg/L and AUC(0-12) = 12.5 +/- 7 mg. h/L) and dose 3 (C(max3) = 4.9 +/- 2 mg/L and AUC(24-36) = 17.5 +/- 11 mg.h/L). Mean burn eschar concentration during the 72 h post-burn was significantly lower than that found during clinical sepsis (18 +/- 17 compared with 41.3 +/- 54 microg/g; P < 0.05 by t test). Similar serum concentrations were achieved in patients with clinical sepsis (C(max1) = 4.2 +/- 0.2 mg/L and AUC(0-12) = 15.0 +/- 3 mg. h/L; C(max3) = 5.0 +/- 1 mg/L and AUC(24-36) = 22.8 +/- 9 mg.h/L). A positive correlation between burn eschar concentrations and C(max) (r = 0.71, r(2) = 0.51, P = 0.01) was found by linear regression analysis. A C(max)/MIC ratio > 10 (MIC = 0.5 mg/L) and an AUC/MIC ratio > 100 SIT(-1).h (serum inhibitory titre) (MIC = 0.125 mg/L) were achieved. High burn eschar concentrations and serum levels, similar to those found in normal volunteers, can be achieved after intravenous ciprofloxacin infusion in critically ill burns patients. PMID- 10702554 TI - Clinical pharmacokinetics of nelfinavir combined with efavirenz and stavudine during rescue treatment of heavily pretreated HIV-infected patients. AB - Nelfinavir is a novel protease inhibitor that exhibits good inhibitory activity against human immunodeficiency virus type 1 (HIV-1) and is currently used in combination with reverse transcriptase inhibitors for the management of HIV infection. In this study we analysed the pharmacokinetic profile of nelfinavir after multiple oral doses in 18 HIV-infected patients during a combination regimen of nelfinavir plus efavirenz and stavudine. Patients who received the study drug for >/=4 weeks were considered for pharmacokinetic evaluation. Blood samples were obtained at the following times: 0 (before nelfinavir administration), 1, 2, 3, 4, 6 and 8 h after administration. Nelfinavir plasma concentrations were analysed by a specific and validated HPLC assay with ultraviolet detection. Nelfinavir concentration-time data were analysed by compartmental and non-compartmental techniques and the pharmacokinetic parameters of nelfinavir were determined according to a one-compartment model. We found a high variability between individuals in nelfinavir plasma concentrations. The mean average drug plasma concentration was 2.22 +/- 1.25 mg/L and the mean AUC during the dosing interval was 17.7 +/- 10.0 mg*h/L. The mean nelfinavir trough plasma concentration was 1.58 +/- 1.0 mg/L. A good relationship was found between AUC(0-8h) and the plasma concentrations measured at 6 h, and the trough plasma concentrations made total body exposure for nelfinavir less predictable. Alternatively, a 2 h abbreviated AUC provides a good estimate of the full AUC(0 8h). Comparing the pharmacokinetic parameters obtained in our patients with those reported for patients receiving nelfinavir monotherapy or nelfinavir combined with nucleoside analogues, one observes substantial overlap with nelfinavir concentrations achieved without efavirenz. PMID- 10702555 TI - First identification of an SHV-12 extended-spectrum beta-lactamase in Klebsiella pneumoniae isolated in Italy. AB - A clinical isolate of Klebsiella pneumoniae highly resistant to third- and fourth generation cephalosporins, cephamycins and aminoglycosides, was isolated in 1991 from urine. Analysis of a crude extract showed the presence of three beta lactamases with isoelectric points of 6.6, 7.5 and 8.2. The enzyme with pI 8.2 was transferred by conjugation into Escherichia coli K-12 J53 and was responsible for the resistance to third-generation cephalosporins and monobactams, but not to other antibiotics. Kinetic studies of partially purified beta-lactamase from the transconjugant strain confirmed that the enzyme was able to hydrolyse ceftazidime, cefotaxime and aztreonam but not cephamycins. Analysis of the transconjugant showed the presence of two small non-conjugative plasmids of 14 and 6 kb. A polymerase chain reaction was performed using primers specific for the bla(SHV) gene and a fragment of the expected size (about 961 bp) was obtained with both the K. pneumoniae clinical isolate and the transconjugant. Nucleotide sequence analysis of the fragment showed that it encoded the enzyme SHV-12, derived from SHV-5 (with Gln-35 to Leu). This is the first report of an SHV-12 like enzyme isolated in Italy. PMID- 10702556 TI - Multiple mutations conferring ciprofloxacin resistance in Staphylococcus aureus demonstrate long-term stability in an antibiotic-free environment. AB - Two unrelated strains of Staphylococcus aureus, one with a single mutation in grlA, the other with multiple mutations in gyrA, gyrB, grlA, grlB, norA and the norA promoter region, encoding low-level and high-level ciprofloxacin resistance, respectively, were studied. The characterized mutations in these genes were conserved when both strains were passaged for at least 500 generations in an antibiotic-free environment. New, rapidly stabilized mutations and higher MICs were detected for strains passaged in sub-MIC ciprofloxacin concentrations. The seeming irreversibility of quinolone resistance may affect the long-term success of this drug class. PMID- 10702557 TI - In vitro activity of R-95867, the active metabolite of a new oral carbapenem, CS 834, against anaerobic bacteria. AB - The in vitro activity of R-95867, the active metabolite of a new oral carbapenem, CS-834, was compared with those of DU-6859a, cefditoren, ampicillin/sulbactam and clindamycin against a variety of anaerobic bacteria. R-95867 inhibited 90% of anaerobic strains at /=10 can be reached for 95%, 84.3%, 83.1% and 81.5% of isolates, respectively, for clinafloxacin, trovafloxacin, moxifloxacin and sparfloxacin (P < 0. 001 compared with levofloxacin and ciprofloxacin). In spite of the rare but serious adverse events associated with the new-generation quinolones, these agents may become very useful in the treatment of certain severe or life-threatening infectious conditions due to S. maltophilia, notably lower respiratory tract infections. PMID- 10702559 TI - In vitro activity of gemifloxacin (SB 265805; LB20304a) against human mycoplasmas. AB - The in vitro activity of gemifloxacin, a new broad-spectrum fluoroquinolone, was compared with those of ciprofloxacin, erythromycin, azithromycin and doxycycline against 29 human respiratory or urogenital tract mycoplasmas. Gemifloxacin was highly active against all of the mycoplasma and ureaplasma species tested (MIC range 0.001-0.25 mg/L) and was 5- to 100-fold more active than ciprofloxacin. Doxycycline was less active than gemifloxacin against the mycoplasmas (MIC range 0.01-1 mg/L) but had similar activity against Ureaplasma urealyticum (MIC ranges 0.025-0.25 mg/L and 0.1-0. 25 mg/L, respectively). The macrolides, particularly azithromycin, were more active than gemifloxacin against Mycoplasma pneumoniae (MIC range 0.001-0.0025 mg/L) and Mycoplasma genitalium (0.0005-0. 001 mg/L) isolates but were less active against Mycoplasma fermentans and U. urealyticum and inactive against Mycoplasma hominis. Gemifloxacin may therefore be useful in the treatment of respiratory, urogenital or systemic mycoplasma infections in humans. PMID- 10702560 TI - Antifungal activity of itraconazole compared with hydroxy-itraconazole in vitro. AB - Microbroth dilution tests in vitro showed the same IC(50) values for itraconazole and hydroxy-itraconazole, within a mode +/- one dilution range of experimental error, for 90% of 1481 isolates of pathogenic fungi, representing 48 genera. Some 10-15% of Candida glabrata and Trichophyton mentagrophytes isolates were more susceptible to itraconazole than hydroxy-itraconazole. Replicate tests with bioassay marker strains of Candida kefyr and Candida albicans showed no susceptibility variations outside the mode +/- one dilution range. We conclude that few fungi differ substantially in their susceptibility to itraconazole and hydroxy-itraconazole. PMID- 10702561 TI - Anticryptosporidial activity of ranalexin, lasalocid and azithromycin alone and in combination in cell lines. AB - The in vitro anticryptosporidial activities of ranalexin, lasalocid and azithromycin alone and in combination were investigated against four clinical isolates of Cryptosporidium parvum. Susceptibility was tested by inoculating the isolates on to cell monolayers and determining the parasite count after 48 h incubation at 37 degrees C. The culture medium was supplemented with Dulbecco's modified Eagle's medium containing serial dilutions of the above-mentioned compounds. Ranalexin showed moderate anticryptosporidial activity: at a concentration of 64 mg/L it reduced parasite counts by 33.8%. Azithromycin at a concentration of 8 mg/L gave inhibition comparable to that observed with the highest concentration of ranalexin. Lasalocid showed the highest activity, with a 70.3% reduction in parasite counts at 2 mg/L. The combination of ranalexin 64 mg/L and lasalocid 2 mg/L completely suppressed parasite growth without harming the monolayer. PMID- 10702562 TI - Therapeutic efficacy of J-111,225, a novel trans-3,5-disubstituted pyrrolidinylthio-1beta-methylcarbapenem, against experimental murine systemic infections. AB - In a murine model of systemic infection with methicillin-resistant Staphylococcus aureus (MRSA), J-111,225 showed an ED(50) value of 5. 83 mg/kg, which was comparable to vancomycin (ED(50) 4.84 mg/kg), whereas imipenem failed to cure infected mice (ED(50) >100 mg/kg). Against a mixed infection caused by MRSA and Pseudomonas aeruginosa, monotherapy with J-111,225 showed an ED(50) value of 7.23 mg/kg, whereas combined treatment with vancomycin plus imipenem (1:1) had an ED(50) of 20.86 mg/kg. J-111,225 showed good therapeutic efficacy against methicillin-susceptible S. aureus, penicillin-resistant Streptococcus pneumoniae, Escherichia coli, Klebsiella pneumoniae and P. aeruginosa. The unusually broad spectrum suggests that monotherapy with this novel carbapenem may be suitable for polymicrobial infections associated with MRSA. PMID- 10702563 TI - Once-daily versus multiple-daily gentamicin in empirical antibiotic therapy of febrile neutropenia following intensive chemotherapy. AB - The clinical efficacy and toxicity of once-daily compared with multiple-daily gentamicin dosing, in combination with azlocillin, were studied retrospectively in febrile neutropenic episodes following intensive chemotherapy. Fifty-two episodes were studied in 28 patients with acute myeloid leukaemia. Reasons for initiation of antibiotic therapy, dose, duration of treatment, organism isolation rates, response, cost comparison and toxicity were studied in the two treatment groups. The main indication for initiation of antibiotic therapy was neutropenic fever without a documented infection (80.8% of episodes). The response rate to once-daily gentamicin dosing and azlocillin was three times higher than to multiple-daily gentamicin dosing and azlocillin (P = 0.0112). The incidence of toxicity was low overall and was slightly but not significantly higher in the once-daily group. In this clinical context once-daily gentamicin at a dose of 7 mg/kg/day is more effective than a multiple-daily dosing regimen but may be more toxic. PMID- 10702564 TI - Use of a treatment protocol in the management of community-acquired lower respiratory tract infection. AB - The aim of the present study was to examine the impact of an antimicrobial prescribing protocol on clinical and economic outcome measures in hospitalized patients with community-acquired lower respiratory tract infection (LRTI). The study was performed as a prospective controlled clinical trial within the medical wards at Antrim Area Hospital, Northern Ireland. Data were collected on all hospitalized adult patients with a primary diagnosis of LRTI during the period December 1994 to February 1995 (normal hospital practice; control group; n = 112). After an LRTI management protocol (medical, microbiological and pharmacy staff) had been developed, all hospitalized adult patients with a primary diagnosis of LRTI over the period December 1995 to February 1996 formed the intervention group (treated according to the protocol; n = 115). The results showed a statistically significant impact of the protocol in terms of clinical and economic outcome measures. Patients treated using the algorithmic prescribing protocol had significant reductions in length of hospital stay (geometric mean 4.5 versus 9.2 days), iv drug administration (34.8% versus 61.6%), duration of iv therapy (geometric mean 2.1 versus 5.7 days) and treatment failures (7.8% versus 31.3%). Healthcare costs were also significantly reduced. The use of the protocol was a major factor in streamlining the prescribing of antimicrobial therapy for community-acquired LRTI and led to more cost-effective patient management. PMID- 10702565 TI - Non-hospital consumption of antibiotics in Spain: 1987-1997. AB - Spain has one of the highest incidences of bacterial resistance to antimicrobials, possibly linked to drug consumption patterns. Using Ministry of Health and Consumer Affairs records, data were obtained on non-hospital sales of antibiotics for the period 1987-1997, and equivalents calculated in weight of active drug ingredient and defined daily doses per 1000 population per day (DDD/1000/day). The number of packages sold declined from 75 million in 1987 to 55 million in 1997. None the less, there was a gradual yet steady rise in consumption in tonnage terms (249 to 275 tonnes). Furthermore, in terms of DDD/1000/day, consumption rose sharply until 1995 and then held steady at 21 DDD/1000/day, a level comparable to the mean for other developed countries. Penicillins were the group to register the highest consumption in Spain, followed in the latter years of the study-by macrolides, cephalosporins and quinolones. The marked rise in these latter three groups was noteworthy. Despite the decrease in the number of packages sold, antibiotic consumption in Spain has risen. This consumption pattern is different from that of other European countries and might serve to explain differences in the generation of resistance. PMID- 10702566 TI - Pneumococcal macrolide resistance-not a myth. PMID- 10702568 TI - Defining high-level gentamicin resistance in enterococci. PMID- 10702569 TI - Effect of the incubation atmosphere on the susceptibilities of Haemophilus influenzae isolates to macrolides and amoxycillin as determined by the Etest. PMID- 10702570 TI - Trends in the susceptibilities of Proteus mirabilis isolates to quinolones. PMID- 10702571 TI - Primary resistance to flucytosine among clinical isolates of Candida spp. PMID- 10702572 TI - Septicaemias caused by a strain of Staphylococcus haemolyticus exhibiting intermediate susceptibility to teicoplanin in multiple intensive care unit patients. PMID- 10702573 TI - Acute pancreatitis associated with severe lactic acidosis in human immunodeficiency virus-infected patients receiving triple therapy. PMID- 10702576 TI - Dietary factors that affect the bioavailability of carotenoids. AB - Carotenoids are thought to contribute to the beneficial effects of increased vegetable consumption. Various dietary factors have an effect on the bioavailability of carotenoids. The type of food matrix in which carotenoids are located is a major factor. The bioavailability of beta-carotene from vegetables in particular has been shown to be low (14% from mixed vegetables) compared with that of purified beta-carotene added to a simple matrix (e.g., salad dressing), whereas for lutein, the difference is much smaller (relative bioavailability of 67% from mixed vegetables). Processing, such as mechanical homogenization or heat treatment, has the potential to enhance the bioavailability of carotenoids from vegetables (from 18% to a sixfold increase). The amount of dietary fat required to ensure carotenoid absorption seems low (approximately 3-5 g per meal), although it depends on the physicochemical characteristics of the carotenoids ingested. Unabsorbable, fat-soluble compounds reduce carotenoid absorption, and interaction among carotenoids may also result in a reduced carotenoid bioavailability. Research into the functional benefits of carotenoids should consider the fact that the bioavailability of beta-carotene in particular is one order of magnitude higher when provided as a pure compound added to foods than when it is present naturally in foods. PMID- 10702577 TI - Acetic acid suppresses the increase in disaccharidase activity that occurs during culture of caco-2 cells. AB - To understand how blood glucose level is lowered by oral administration of vinegar, we examined effects of acetic acid on glucose transport and disaccharidase activity in Caco-2 cells. Cells were cultured for 15 d in a medium containing 5 mmol/L of acetic acid. This chronic treatment did not affect cell growth or viability, and furthermore, apoptotic cell death was not observed. Glucose transport, evaluated with a nonmetabolizable substrate, 3-O-methyl glucose, also was not affected. However, the increase of sucrase activity observed in control cells (no acetic acid) was significantly suppressed by acetic acid (P < 0.01). Acetic acid suppressed sucrase activity in concentration- and time-dependent manners. Similar treatments (5 mmol/L and 15 d) with other organic acids such as citric, succinic, L-maric, L-lactic, L-tartaric and itaconic acids, did not suppress the increase in sucrase activity. Acetic acid treatment (5 mmol/L and 15 d) significantly decreased the activities of disaccharidases (sucrase, maltase, trehalase and lactase) and angiotensin-I-converting enzyme, whereas the activities of other hydrolases (alkaline phosphatase, aminopeptidase N, dipeptidylpeptidase-IV and gamma-glutamyltranspeptidase) were not affected. To understand mechanisms underlying the suppression of disaccharidase activity by acetic acid, Northern and Western analyses of the sucrase-isomaltase complex were performed. Acetic acid did not affect the de novo synthesis of this complex at either the transcriptional or translational levels. The antihyperglycemic effect of acetic acid may be partially due to the suppression of disaccharidase activity. This suppression seems to occur during the post-translational processing. PMID- 10702578 TI - Food intake, energy balance and serum leptin concentrations in rats fed low protein diets. AB - Studies examining the effects of low-protein diets on food intake and body weight have shown varied results. Many researchers have found low dietary protein to increase food intake, while others have found no effect or even a decrease. In 63 male Sprague-Dawley rats, we examined several low levels of dietary protein (2%, 5%, 8%, 10%, 15% vs. 20% casein) to determine the dose-response relationships between low dietary protein and food intake, body composition, energy balance and serum leptin concentrations. Food intake, over the range of low dietary protein, showed a quasi bell-shaped response curve with peak intake occurring in rats fed 8-10% casein. Peak feeding occurred at or just below the estimated protein requirement of the rats (10-12.5% casein). Compared to the 20% casein controls, food intake was severely reduced in rats fed 2% casein, while it was greater in the other low-protein groups. The amount of body fat steadily increased between the 15% casein group and the 8% casein group, and sharply declined between the 5% casein group and 2% casein group. The change in body fat reflected both the change in food intake and altered energy partitioning. Serum leptin concentrations were greater in rats fed the 5 and 8% casein diets than in control rats fed 20% casein. Serum leptin concentrations were positively associated with body fat content (r(2) = 0.763, P < 0.001). Increased serum leptin concentrations in the presence of increased food intake is suggestive of a state of leptin resistance. This animal model may provide important insights into diet-induced obesity. PMID- 10702579 TI - Colonic cell proliferation and aberrant crypt foci formation are inhibited by dairy glycosphingolipids in 1, 2-dimethylhydrazine-treated CF1 mice. AB - Dietary sphingomyelin (SM) inhibits early stages of colon cancer (appearance of aberrant crypt foci, ACF) and decreases the proportion of adenocarcinomas vs. adenomas in 1,2-dimethylhydrazine (DMH)-treated CF1 mice. To elucidate the structural specificity of this inhibition, the effects of the other major sphingolipids in milk (glycosphingolipids) were determined. Glucosylceramide (GluCer), lactosylceramide (LacCer) and ganglioside G(D3) were fed individually to DMH-treated (six doses of 30 mg/kg body weight) female CF1 mice at 0.025 or 0.1 g/100 g of the diet for 4 wk. All reduced the number of ACF by > 40% (P < 0.001), which is comparable to the reduction by SM in earlier studies. Immunohistochemical analysis of the colons revealed that sphingolipid feeding also reduced proliferation, with the most profound effect (up to 80%; P < 0.001) in the upper half of the crypts. Since the bioactive backbones of the glycosphingolipids (i.e., ceramide and other metabolites) are the likely mediators of these effects, the susceptibility of these complex sphingolipids to digestion in the colon was examined by incubating 500 microgram of each sphingolipid with colonic segments from mice and analysis of substrate disappearance and product formation by tandem mass spectrometry. All of the sphingolipids (including SM) disappeared over time with a substantial portion appearing as ceramide. Partially hydrolyzed intermediates (such as GluCer from LacCer or G(D3)) were not detected, which suggests that the cleavage involves colonic (or microflora) endoglycosidases. In summary, consumption of dairy SM and glycosphingolipids suppresses colonic cell proliferation and ACF formation in DMH treated mice; hence, many categories of sphingolipids affect these key events in colon carcinogenesis. PMID- 10702580 TI - Albumin synthesis is diminished in men consuming a predominantly vegetarian diet. AB - Albumin synthesis was calculated in healthy male volunteers consuming diets differing in the relative contribution of protein from animal or vegetable sources. In one study (Study 1, n = 4) two isoenergetic and isonitrogenous diets were consumed for a period of 10 d each. One diet (diet A) was animal protein rich (74%), the other one (diet V) contained 67% of vegetable protein. Albumin synthesis rate was measured from L-[(2)H(5)]phenylalanine incorporation (43 mg/kg) at the end of each dietary period. Both albumin fractional synthesis rate (FSR) (5.7 +/- 0.6 vs. 6.7 +/- 0. 8%/d, P = 0.04) and absolute synthesis rate (ASR) (123 +/- 6 vs. 143 +/- 8 mg. kg(-1). d(-1), P = 0.05) were reduced after diet V. In a second study (Study 2, n = 8) a third dietary treatment was added (Diet VS). This was similar to diet V but supplemented with soy protein (18g/d). The results of study 2 confirmed that albumin synthesis was reduced after diet V (FSR: 5.9 +/- 0.3 vs. 6.7 +/- 0. 5%/d, P = 0.015; ASR: 126 +/- 7 vs. 146 +/- 9 mg. kg(-1). d(-1), P = 0.007), but it also showed that the drop could be prevented by adding supplemental protein to the predominantly vegetarian diet (Diet VS) (FSR: 6.4 +/- 0.3%/d, P = 0.08; ASR: 140 +/- 7 mg. kg(-1). d(-1), P = 0.03). Albumin synthesis appears to be modulated by changes in the proportion of animal vs. vegetable protein occurring in the diet. The mechanism might be related to differences in digestibility and consequently in net amino acid availability between diets. PMID- 10702581 TI - The bioavailability of beta-carotene in stir- or deep-fried vegetables in men determined by measuring the serum response to a single ingestion. AB - To evaluate the bioavailability of beta-carotene from plant foods, the serum beta carotene response to a single ingestion of various beta-carotene sources was determined in 10 healthy men. Tested beta-carotene sources included stir-fried shredded carrot, stir-fried water convolvulus leaves, deep-fried sweet potato ball, purified beta-carotene in a capsule (beadlets) and beadlets with beta carotene free oriental radish (beadlets + radish). The maximal change in serum beta-carotene concentration occurred at 24 or 32 h post ingestion. This response to beadlets was significantly higher than that to the other four tested beta carotene sources (P < 0.05). The maximal serum response to beadlets + radish was also significantly higher than that to the three food beta-carotene sources (P < 0.05). The maximal serum response to sweet potato was significantly higher than that to water convolvulus leaves (P < 0. 05). The bioavailability relative to beta-carotene beadlets was calculated by dividing the maximal change in serum concentration to each test meal of each subject by his own serum maximal change in response to beadlets. Accordingly, the bioavailability was 65% for beadlets + radish, 33% for carrots, 26% for water convolvulus leaves and 37% for sweet potatoes. Concurrent ingestion of oriental radish reduced the bioavailability of beadlets to two-thirds of its original value, which partially accounted for the difference between the bioavailability of beadlets and natural foods. The relative bioavailability of beta-carotene from stir-fried and deep-fried vegetables was about one-third to one-fourth that of the purified beta-carotene beadlets. These bioavailabilities are higher than previously reported values. PMID- 10702582 TI - Time course of and effect of dietary iron level on iron incorporation into erythrocytes by infants. AB - As a part of our effort to explore various aspects of ferrokinetics in infancy, the present study was designed to determine the timing of entry of an orally ingested iron isotope into circulating erythrocytes, and the effect of the level of dietary iron [0.3 mg/100 kcal (418.4 kJ) vs. 1.8 mg/100 kcal] after isotope administration on erythrocyte incorporation of the isotope. We administered the stable isotope, (58)Fe, orally to 56-d-old and 168-d-old infants. All infants were fed a low-iron formula (LF) before and until 5 h after isotope administration. Thereafter, half the infants were fed a formula high in iron (HF group) while the remaining infants continued to receive the LF (LF group) for an additional 28 d. The quantity of (58)Fe in circulating erythrocytes increased from 14 to 28 d after isotope administration was nearly constant from 28 through 84 d of age (plateau value) and decreased between 84 and 112 d. Erythrocyte incorporation of (58)Fe was greater by the 168-d-old infants than by the 56-d-old infants, presumably because of the lesser iron stores of the older infants. In the 56-d-old infants, erythrocyte incorporation of (58)Fe was greater by the LF than by the HF group, but this difference was not significant in the 168-d-old infants. Thus, at least in younger infants, the level of iron intake after administration of an iron isotope affects erythrocyte incorporation of the isotope. The fact that less isotope was present in erythrocytes 112 d than 84 d after administration indicates that the life span of erythrocytes of infants, even beyond the immediate newborn period, is less than the 120-d life span of erythrocytes in the adult. PMID- 10702583 TI - Growth faltering is prevented by breast-feeding in underprivileged infants from Mexico City. AB - This study was designed to test whether breast-feeding protects infants reared in unfavorable environments from growth-stunting by averting acute infections. The body weight and length, feeding mode and morbidity of 170 healthy infants were assessed at 15-d intervals from birth to 6 mo. Birth weight and length were not different between groups, but at 6 mo, breast-fed infants were heavier and tended to be taller (P = 0.1) than infants fed formula. Relative to NCHS values, infants had lower mean birth weights than a sample of American and European BF infants. At 6 mo, the weight of BF infants caught up to the weight of NCHS standards, while infants fed formula fell to around -1 NCHS-Z-score for weight and length. The cumulative 6-mo weight increments were negatively related to the number of episodes of diarrhea, and positively to duration of lactation (P = 0.03, R(2) = 0.17). The 6-mo length gain was negatively related to infections but not to duration of lactation (P = 0.004, R(2) = 0.19). Never-ill infants attained a better weight (P = 0.04) and length (P = 0.02) than infants who suffered one or more episodes of diarrhea. Weight and length gain of infants suffering at least one episode of diarrhea was positively related to breast-feeding and socioeconomic status. Weight increments of 15-d were positively related to breast feeding and negatively to the introduction of solids. In conclusion, breast feeding positively affected the growth performance of the recipient infants by averting infections and possibly by improving nutrient intake during infections. PMID- 10702584 TI - Vitamin B-6 inadequacy is prevalent in rural and urban Indonesian children. AB - The vitamin B-6 status of Indonesian children was evaluated by determining their dietary vitamin B-6 intakes, erythrocyte alanine aminotransferase activity coefficients and plasma pyridoxal phosphate (PLP) concentrations. Thirty-eight third-grade elementary school children (ages = 8-9 y) in rural and 39 in urban areas of Bogor, West Java, Indonesia, voluntarily served as subjects. The subjects included 39 male and 38 female students. The mean vitamin B-6 intake of the subjects was 0.57 mg/d. Fifty-five percentage of the children reported consuming <0.5 mg/d of vitamin B-6 (the 1998 Estimated Average Requirement for those 4-8 y). Erythrocyte alanine aminotransferase activity coefficients >/= 1.25 were observed in 30%, and plasma PLP concentrations 0.10) thereafter. Fractional rates of protein synthesis (K(s), %/d) based on SRA in plasma- or intracellular-free phenylalanine did not differ (P > 0.10) in all tissues except pancreas (P < 0.05). Diet affected K(s )in liver (P < 0.01) and colon (P < 0.05) but not in pancreas, duodenum, jejunum and cecum. Based on plasma-free phenylalanine SRA, liver K(s)were 85.4 +/- 11.0 vs. 60.5 +/- 5.2 (mean +/- SEM) in CC- and BCM-fed pigs, respectively; these values were 82.3 +/- 4.7 vs. 98.2 +/- 5.8 in the colon. The absolute amount of protein synthesis (g.d( 1)) was higher in the liver (P < 0.05) and pancreas (P < 0. 05) of the CC pigs compared to BCM pigs. No dietary effects were observed in all other organs (P > 0.10). The present results suggest that feeding growing pigs a BCM diet that induces high ENL does not affect PSR in the small intestine of growing pigs from which >50% of ENL originates. PMID- 10702587 TI - Nutritional status affects intestinal carotene cleavage activity and carotene conversion to vitamin A in rats. AB - Validation of an in vivo method we developed recently and its application to assess the role of dietary factors in carotene conversion were tested in rats. We compared the ratio of area under plasma vitamin A time-curves (AUC(0-12h)) obtained after a dose of beta-carotene to that after a dose of vitamin A, with the in vitro intestinal supernatant beta-carotene dioxygenase activity. In separate experiments, vitamin A (AD) and protein deficiencies (PD) were produced in male WNIN weanling rats. Corresponding food-restricted (AR and PR) and unrestricted rats (AA and PA) served as controls. Three rats in each of the AD, AR and AA groups received oral doses of 50-300 microgram beta-carotene or 25-150 microgram vitamin A and four rats in each of the PD, PR and PA groups received only 100 microg beta-carotene or vitamin A. The plasma vitamin A AUC(0-12h) with beta-carotene or vitamin A were significantly and positively correlated (r = 0.714-0.918, n = 9-12, P < 0.05) with the dose in AD, AR and AA groups. The AUC(0 12h) slope ratios in AD, AR and AA rats were 0.33, 0.20 and 0.26, respectively. The beta-carotene dioxygenase activity (pmol retinal. h(-1). mg protein(-1)) was significantly higher in the AD group (14.9 +/- 2.43) compared to both AR (6.7 +/- 0.62) and AA (6.3 +/- 1.37) groups and was parallel with in vivo conversion of beta-carotene to vitamin A. The AUC(0-12h) ratio was lower in PD rats (0.13) compared to PR (0.26) and PA (0.5) groups. Similarly, the in vitro enzyme activity (pmol retinal. h(-1). mg protein(-1)) in PD rats was significantly lower (3.6 +/- 1.30) compared to PR (13.7 +/- 0.92) and PA groups (13.8 +/- 1.6). Thus the results validate the methodology and confirm the role of nutritional factors in carotene conversion to vitamin A. PMID- 10702588 TI - Improved cholecalciferol nutrition in rats is noncalcemic, suppresses parathyroid hormone and increases responsiveness to 1, 25-dihydroxycholecalciferol. AB - We examined how cholecalciferol (vitamin D) nutrition affected serum 25 hydroxycholecalciferol (25(OH)D) and 1, 25-dihydroxycholecalciferol (1,25(OH)(2)D). Rats were fed conventional diet (vitamin D, 4.5 IU/g, or 7 nmol/d) or the same diet plus 18 nmol/d of extra vitamin D for 3 wk. The extra vitamin D resulted in greater serum 25(OH)D (51 +/- 3, vs. control of 21 +/- 2 nmol/L), and kidney mRNA for vitamin D receptor [VDR mRNA] (P = 0. 026) and lower serum 1,25(OH)(2)D (72 +/- 16 vs. control of 161 +/- 10 pmol/L, P = 0.001), and parathyroid hormone (PTH) (89 +/- 4 vs. control of 160 +/- 15 ng/L, P = 0.001). Kidney VDR mRNA relative to GAPDH mRNA correlated inversely with serum 1,25(OH)(2)D (r = -0.714, P = 0.006). There were no differences in serum calcium, phosphate, alkaline phosphatase, or weight gain. Experiment 2 compared groups supplemented with 0.2, 2 or 20 nmol/d of vitamin D orally, or 20 nmol/d dermally to see how vitamin D nutrition influenced the response of 1,25(OH)(2)D to changes in diet calcium. Vitamin D did not affect urinary calcium or pyridinoline excretion, serum calcium, phosphate, vitamin D binding protein or alkaline phosphatase. In groups given 20 nmol/d of vitamin D, renal mitochondrial 25(OH)D 1alpha-hydroxylase was lower (P < 0.01) and 25(OH)D-24-hydroxylase was higher (P < 0.05). Higher 25(OH)D concentration was related to proportionally lower 1,25(OH)(2)D at every calcium intake, indicating greater tissue sensitivity to 1, 25(OH)(2)D. We conclude suppression of 1,25(OH)(2)D and PTH, and higher renal VDR mRNA and 24-hydroxylase did not involve higher free 1,25(OH)(2)D concentration or a first pass effect at the gut. Thus, 25(OH)D or a metabolite other than 1,25(OH)(2)D is a physiological, transcriptionally and biochemically active, noncalcemic vitamin D metabolite. PMID- 10702589 TI - In vitro fermentation of swine ileal digesta containing oat bran dietary fiber by rat cecal inocula adapted to the test fiber increases propionate production but fermentation of wheat bran ileal digesta does not produce more butyrate. AB - This experiment evaluated three hypotheses: i) production of propionate is increased during fermentation of substrate containing oat bran (OB)(6); ii) production of butyrate is increased during fermentation of substrate containing wheat bran (WB) and iii) results of in vitro fermentations using physiological substrates and inocula agree with in vivo data. Ileal digesta collected from swine fed OB and WB were the substrates. Digesta was fermented for 0-96 h in an anaerobic in vitro system using inocula prepared from ceca of rats fed the same fiber sources. Carbohydrate and short-chain fatty acid (SCFA) contents in the fermentations were measured by gas chromatography. Fermentation of WB digesta did not produce more n-butyrate (P > 0.05) and was significantly slower (P < 0.05) than fermentation of OB digesta. OB digesta fermentation produced a significantly greater (P < 0.05) molar proportion of SCFA as propionate. Bacterial mass increased more and was maintained longer during fermentation of OB digesta than the WB digesta. Our results indicate that dilution of undigested WB fiber and not n-butyrate production is one mechanism by which WB may protect colonic mucosa; propionate production is increased during fermentation of beta-glucan in OB; and an in vitro system using physiological sources of inoculum and substrate containing WB and OB yields results that agree with in vivo findings in humans and rats. PMID- 10702590 TI - A high oat-bran intake does not impair zinc absorption in humans when added to a low-fiber animal protein-based diet. AB - Oat bran has a high phytate content and a low or inactivated phytase activity. A high intake of oat bran could therefore result in an impaired absorption of trace elements. The effect of a mean daily intake of 142 g of oat bran (102 g/10 MJ) on absorption of zinc was evaluated by the use of stable isotopes and fecal monitoring in 12 healthy subjects (6 males and 6 females). Each subject participated in two separate diet periods each of 21 d with identical low-fiber diets and with oat bran added in one of the periods. The oat bran was incorporated into bread and served at three daily main meals. The intake of zinc and phytate per 10 MJ was 138 micromol (9.0 mg) and 0.5 mmol, respectively, in the low-fiber period and 225 micromol (14.7 mg) and 4.0 mmol, respectively, in the oat bran period. Stable isotopes of zinc ((70)Zn) were added to the diets at d 7 of each period. The fractional absorptions (means +/- SD) of zinc from the low-fiber and oat bran diets were 0.48 +/- 0.11 and 0.40 +/- 0.15 (P = 0.07), respectively. The higher zinc content in the oat bran period resulted in a greater amount of zinc absorbed (64 +/- 19 micromol and 99 +/- 51 micromol, respectively, P = 0.009). Balance data suggest that the higher absorbed amount of zinc resulted in correspondingly higher intestinal endogenous excretion of zinc. In conclusion, the absorption of zinc was high and not affected by addition of oat bran. PMID- 10702591 TI - Gender and hormonal status affect the hypolipidemic mechanisms of dietary soluble fiber in guinea pigs. AB - The objective of this study was to assess the effects of gender on the secondary mechanisms by which dietary soluble fiber lowers plasma LDL cholesterol. For that purpose, male, female and ovariectomized (to mimic menopause) guinea pigs (8-10 per group) were allocated to two dietary treatments. Diets were identical in composition except for the fiber source: the control diet contained 10 g/100 of cellulose and 2.5 g/100 g of guar gum, while the soluble fiber (SF) diet contained 5 g/100 of psyllium, 5 g/100 of pectin and 2.5 g/100 g of guar gum. SF intake resulted in 44% lower plasma LDL cholesterol, 64% lower apo B and 22% lower plasma triacylglycerol (TAG) concentrations (P < 0.01) compared to guinea pigs fed the control diet. However, ovariectomized guinea pigs had higher plasma cholesterol, apo B and TAG concentrations (P < 0.01) compared to males and females, even those fed SF. Plasma HDL-cholesterol was higher in females than in males (P < 0.05). LDL size, as measured by LDL composition and fast protein liquid chromatography, was larger in females than males. Guinea pigs fed SF had smaller LDL than controls. LDL susceptibility to oxidation was 80% lower in male and females fed the SF diet (P < 0.001) than in controls, while there was no effect of diet in ovariectomized guinea pigs. Hepatic free cholesterol and TAG were lower, and activities of 3-hydroxy-3-methyl-glutaryl coenzyme A (HMG-CoA) reductase and cholesterol 7alpha-hydroxylase were higher in guinea pigs fed SF (P < 0.05) than in controls. These results indicate that gender plays an important role in the metabolic responses to dietary soluble fiber and that estrogen deprivation leads to a detrimental lipoprotein profile. PMID- 10702592 TI - Insulin stimulates phenylalanine uptake across the hind limb in fed lambs. AB - Five lambs ( approximately 6 mo of age and 30 kg), with an external iliac artery and vein catheterized and fed to maintain body weight, were used to examine effects of close arterial infusion of insulin and branched-chain amino acids (BCAA) on net phenylalanine (Phe) uptake across the hind limb. Treatments, administered randomly on five consecutive days to each lamb, were 400 min infusions of: i) saline (control); ii) insulin to double iliac artery concentration (low insulin); iii) as ii but to quadruple insulin concentration (high insulin); iv) 30 micromol/min leucine and 22.5 micromol/min each isoleucine and valine (BCAA) and v) co-infusion of ii and iv. Blood was sampled over the last 200 min from the iliac vein and right ventricle of the heart. High insulin caused a slight decrease (-13%, P < 0.05) in systemic glucose concentration, but did not alter systemic insulin concentration. Insulin, at both doses and in combination with BCAA, resulted in 9-fold greater net Phe uptake (P < 0.05) than the control, as did BCAA alone. Because BCAA alone increased net Phe uptake, these may have stimulatory effects directly or may enhance endogenous insulin. Maximum stimulation was achieved with low insulin because there was no increase in net Phe uptake with high insulin or from co-infusion with BCAA. Insulin, at low concentrations, may be important to growth in animals with marginal nutrition. PMID- 10702593 TI - Dietary sulfur amino acid requirement of juvenile yellow perch fed the maximum cystine replacement value for methionine. AB - We conducted three separate experiments designed to determine the dietary methionine requirement, ability of cyst(e)ine to spare methionine, and the total sulfur amino acid requirement (TSAA) of juvenile yellow perch when fed the maximal amount of cyst(e)ine. The purified basal diet used in each experiment contained 33.6 g of crude protein/100 g diet and 12.0 g of lipid/100 g diet. In the first experiment,;>L-methionine was added to eight diets providing methionine concentrations ranging from 0.37 to 1.77 g/100 g diet in gradations of 0.2 g/100 g diet. Diets were fed for 12 wk to juvenile yellow perch initially weighing 4.7 g/fish. Broken-line analyses of weight gain and feed efficiency data indicated that the dietary methionine requirement was 1.0 g/100 g diet (3.1 g TSAA/100 g dietary protein) and 1.1 g/100 g diet (3.4 g TSAA/100 g dietary protein), respectively. In the second experiment, various ratios of L-cyst(e)ine and L methionine were added to the basal diet and fed for 12 wk to determine the cyst(e)ine replacement value of yellow perch initially weighing 19.3 g/fish. Weight gain and feed efficiency (FE) data indicated that cyst(e)ine spared up to 51% of the methionine requirement. In the final experiment, graded levels of cyst(e)ine plus methionine in a ratio of 51:49 were added to the basal diet in gradations of 0.1 g/100 g diet (0.5 to 1.2 g TSAA/100 g diet) to determine the dietary total sulfur amino acid requirement. Diets were fed to satiation for 10 wk to fish initially weighing 8. 1 g. Broken-line analyses of weight gain, feed intake and FE data indicated that the dietary TSAA requirement was 0.85, 0.87 and 1.0 g of TSAA/100 g diet (2.5 to 3.0 g of TSAA/100 g of dietary protein), respectively. The majority of dietary TSAA requirements of fish are in the range of 2 to 4 g/100 g of dietary protein and are generally similar to those of both birds and swine, but lower than estimates for rodents. PMID- 10702594 TI - Hepatic ascorbic acid saturation is the most stringent response criterion for determining the vitamin C requirement of juvenile European sea bass (Dicentrarchus labrax). AB - Our main objective was to verify whether the dietary ascorbic acid (AA) requirement of juvenile European sea bass (Dicentrarchus labrax) varies as a function of different physiological needs. Practical diets with eight (0, 5, 10, 20, 40, 80, 160, 320 mg AA/kg diet) levels of ascorbic acid polyphosphate were fed to sea bass (mean weight: 0.7 g) for 15 wk. At the beginning and at the end of the feeding trial, tissues were sampled for vitamin C and hydroxyproline (HyPro) analysis. Dose-dependent responses of skin and whole body HyPro concentrations and hepatic AA concentration to dietary vitamin C levels were observed. Skin and whole body HyPro concentrations were low in sea bass fed AA deficient diet, 217 and 15 nmol/g tissue, respectively. HyPro levels increased with increasing dietary levels, reaching plateaus of 297 and 45 nmol/g tissue in the skin and whole body at dietary vitamin C levels of at least 5 and 31 mg AA/kg. Hepatic AA level increased with increasing dietary levels, reaching a plateau of 474 pmol/g tissue in juveniles fed at least 121 mg of AA/kg. We concluded that hepatic AA saturation is the most stringent response criterion for determination of the vitamin C requirement in juvenile European sea bass. PMID- 10702595 TI - Both iron deficiency and daily iron supplements increase lipid peroxidation in rats. AB - Numerous studies have shown that iron-loaded diets increase markers of lipid peroxidation in rats, but few have addressed the effects of oral iron supplements on these markers. We investigated the effects of daily and intermittent iron supplements on iron and vitamin E status, and lipid peroxidation. Iron supplements were administered in doses equivalent to those often given to pregnant women in the developing world. In Study 1, iron-deficient (D) and iron normal (N) rats were fed either 0 or 8000 microgram of supplemental iron daily for 21 d. In Study 2, D rats were fed either the same supplements daily or once every 3 d (8 supplements total). Lipid peroxidation was assessed by breath ethane and pentane and by malondialdehyde (MDA) (using GC-MS). In Study 1, daily supplemented N and D rats had liver nonheme iron concentrations that were 1.8- and 2.7-fold higher, respectively, than those in unsupplemented N rats. Breath ethane levels were also higher in supplemented rats (P < 0.05), but MDA (in plasma, liver, kidney) and liver vitamin E did not differ. Unexpectedly, severely D, anemic rats had significant elevations in the levels of breath ethane, liver MDA and kidney MDA. In Study 2, liver iron and breath ethane decreased progressively (P < 0.05) from 1 d to 3 d after the last iron dose in intermittently supplemented rats. We conclude that iron deficiency results in lipid peroxidation, but that its correction with daily iron supplements results in abnormal iron accumulation and increased lipid peroxidation in rats. These effects are mitigated by intermittent iron supplementation. PMID- 10702596 TI - Dietary vitamin A modulates the concentrations of RRR-alpha-tocopherol in plasma lipoproteins from calves fed milk replacer. AB - The practice of supplementing milk replacers fed to neonatal calves with high concentrations of vitamin A has raised concerns regarding the effect of excess vitamin A on the bioavailability of vitamin E. A 4 x 2 factorial experiment evaluated the effects of four dietary amounts of vitamin A [0, 1.78 [National Research Council (NRC)(6) requirement, control], 35.6 and 71.2 micromol daily as retinyl acetate] and two forms of vitamin E (RRR-alpha-tocopherol and RRR-alpha tocopheryl acetate, 155 micromol daily) on plasma RRR-alpha-tocopherol and RRR gamma-tocopherol and RRR-alpha-tocopherol associated with plasma lipoproteins (Lp) from milk replacer-fed Holstein calves from birth to 28 d of age. The VLDL, LDL, HDL and very high-density lipoprotein (VHDL) fractions were separated by ultracentrifugal flotation, and the amount of vitamin E associated with each fraction was determined by normal-phase HPLC. The amount and distribution of RRR alpha-tocopherol in Lp fractions were unaffected by the form of dietary vitamin E. Plasma and Lp RRR-alpha-tocopherol concentrations increased with age (P < 0.0001) and were maximal at 28 d of age. Concentrations of RRR-alpha-tocopherol associated with Lp were 25% (P < 0.01) to 39% (P < 0.0001) lower in calves fed 35.6 and 71.2 micromol of vitamin A daily than in control calves at 28 d of age. The RRR-gamma-tocopherol concentrations were unaffected by dietary vitamin A (P >/= 0.05). In conclusion, dietary vitamin A modulated the amount and distribution of RRR-alpha-tocopherol in the circulation of milk replacer-fed neonatal calves. Because of the essential antioxidant role of vitamin E, the health-related consequences associated with the depression of the LP RRR-alpha-tocopherol concentrations in calves fed vitamin A at 35.6 and 71.2 micromol need to be investigated. PMID- 10702597 TI - Intravenous infusion of hexamethonium and atropine but not propranolol diminishes apolipoprotein A-IV gene expression in rat ileum. AB - To clarify the role of neural factors in the regulation of apolipoprotein (apo) A IV expression in the small intestine, we investigated the effect of neural blockers on mRNA levels of apo A-IV in rat small intestine. Either ganglionic blocker (hexamethonium), cholinergic blocker (atropine) or beta-adrenergic blocker (propranolol) was infused intravenously to unrestrained conscious rats for 8 h, and then total RNA was isolated from the small intestine and analyzed using Northern hybridization. Apo A-IV mRNA levels in the ileum were significantly lower in hexamethonium- or atropine-infused rats than in saline- (control) or propranolol-infused rats. Immunoblot analysis showed no difference in plasma apo A-IV concentrations between hexamethonium- and saline-infused groups. The lower mRNA levels of apo A-IV in the ileum of hexamethonium-infused rats were observed even in bile-drained rats, indicating that the lower expression was not due to any changes in bile availability. The ileal apo A-IV mRNA levels were significantly higher in rats infused with lipid emulsion into the ileum than in rats infused with glucose-saline, and the concomitant infusion of intravenous hexamethonium did not affect the higher levels of apo A-IV mRNA. These results suggest that the basal expression of the ileal A-IV gene is at least partially regulated in a site-specific manner by cholinergic neurons. PMID- 10702599 TI - The canary in the cell: a sentinel role for beta-carotene. PMID- 10702598 TI - Olestra consumption is not associated with macular pigment optical density in a cross-sectional volunteer sample in Indianapolis. AB - The associations between the intake of the fat-substitute olestra and the concentrations of macular carotenoid pigments and serum lutein and zeaxanthin were investigated in a volunteer cross-sectional sample in Indianapolis. The study was conducted in January through March, 1998 after olestra-containing savory snacks had been sold in central Indiana for a year. Volunteers (n = 280) aged 18-50 y were recruited to make a single clinic visit during which macular pigment optical density (MPOD) was determined by psychophysical flicker photometry, serum was obtained for determination of lutein and zeaxanthin concentration, usual intake of olestra, carotenoids and nutrients were assessed by 1-y food frequency questionnaire, and health habits including smoking, physical characteristics such as eye color, demographics and medical history were determined by questionnaire. Intake of olestra at least one time per month for the past year was reported by 81:280 subjects and their mean, median and 90(th) percentile intakes were 1.09, 0.34 and 2.43 g olestra/d, respectively. Mean macular pigment optical density was not significantly different between olestra consumers and nonconsumers (0.213 +/- 0.014 vs. 0.211 +/- 0.010) nor was serum lutein and zeaxanthin concentration (0.361 +/- 0.017 vs. 0.375 +/- 0. 013 micromol/L) or intake (1242 +/- 103 mg/d vs. 1042 +/- 58 mg/d) in one-way or two way ANOVA. Olestra intake was not associated with MPOD or serum lutein and zeaxanthin before or after correction for significant covariates of MPOD. Thus, olestra intake over the past year in a cross-sectional volunteer sample in Indianapolis was not associated with MPOD. PMID- 10702600 TI - 3rd International Symposium on the Role of Soy in Preventing and Treating Chronic Disease. Washington DC, USA. October 31-November 3, 1999. Proceedings and abstracts. PMID- 10702601 TI - Absorption and metabolism of soy isoflavones-from food to dietary supplements and adults to infants. PMID- 10702602 TI - Beyond ERalpha and ERbeta: estrogen receptor binding is only part of the isoflavone story. PMID- 10702603 TI - Phytoestrogens and prostate disease. PMID- 10702604 TI - Hormonal effects of soy isoflavones: studies in premenopausal and postmenopausal women. PMID- 10702605 TI - Soy and cardiovascular disease: cholesterol lowering and beyond. PMID- 10702607 TI - Oral Presentation Abstracts. PMID- 10702606 TI - Soybean isoflavones as an alternative to traditional hormone replacement therapy: are we there yet? PMID- 10702608 TI - Poster Presentation Abstracts. PMID- 10702609 TI - Screening for genetic mutations. A review. AB - A point mutation of a nucleotide within a single gene can have a profound effect on a specific organ and/or the entire human body. DNA sequences associated with human diseases may differ from the corresponding normal sequences by single nucleotide mutations or by large alterations such as deletions, insertions, duplications, or translocations of DNA segments or entire chromosomes. As a result of the heterogeneity of DNA alterations and genetic mutations, various screening approaches are required to detect these alterations. However, methods which facilitate the detection of large mutations in the genome are typically insensitive to point mutations, whereas methods which detect point mutations are not appropriate to detect large alterations within the genome. Since there is no single perfect method to screen for unknown mutations, combinations of these methods may be necessary for accurate genetic diagnosis. The applications of polymerase chain reaction (PCR) technology to genomic screening have made rapid and accurate genetical diagnosis possible. Furthermore, recent developments in the technology of DNA microarrays have opened the way for high throughput sequence analysis by hybridization, which shows great potential in both molecular biology and medicine in the near future. PMID- 10702610 TI - High-throughput screening of genetic mutations/polymorphisms by capillary electrophoresis. AB - Genetic mutations/polymorphisms analyses play a great role in genetic and medical research, and clinical diagnosis. Most conventional methods for genetic assay are based on slab gel electrophoresis that is both labor-intensive and time consuming. Recently, capillary electrophoresis (CE) has been used for genetic analysis instead of conventional slab gel electrophoresis. This technique can be automated and is characterized by short analysis time, small sample and reagents requirements, and high separation efficiency. CE has been successfully applied for mutation detection involving human tumor suppressor genes, oncogenes and disease-causing genes, and has shown a great potential for genetic mutation/polymorphism screening of large numbers of clinical samples. In this article, an overview of the fundamental aspects of mutation/polymorphism assay methods in combination with CE is given and some key applications are summarized. PMID- 10702612 TI - Solid-phase synthesis and biological screening of N-alpha-mercaptoamide template based matrix metalloprotease inhibitors. AB - A series of N-alpha-mercaptoacetyl containing dipeptides have been prepared on solid-phase supports as putative matrix metalloprotease (MMP) inhibitors. Inhibitor design was based on a positional scanning approach of the amino acids present within a template molecule, previously shown to be an MMP inhibitor with good pharmacological characteristics. This study is the first step in a unique programme, designed to expand the repertoire of molecular templates which can be chosen as starting points for the development of more focused parallel and/or combinatorial libraries of MMP inhibitors as a means to accelerate the lead discovery process. This paper reports the success of such an approach in the development of agents with activity against a number of pathologically important MMPs. After screening of these positional scanning libraries, we have obtained important SAR information, in particular, pharmacophores with the ability to impart selectivity for particular MMP species. PMID- 10702611 TI - Measurement of cdk4 kinase activity using an affinity peptide-tagging technology. AB - Cyclin-dependent kinases such as Cdk4 are involved in the control of cell cycle progression, and misregulation of Cdk4 has been implicated in many types of cancers. In the present study, we report the development of a novel homogeneous assay using an affinity peptide-tagging technology for rapidly discovering Cdk4 inhibitors. The DNA sequence encoding a streptavidin recognition motif, or StrepTag (AWRHPQFGG), was cloned and expressed at the C-terminus of a fusion protein of a 152-amino acid hyperphosphorylation domain (Rb152) of the retinoblastoma protein (Rb) linked to GST at the N-terminus. This affinity peptide-tagged protein (GST-Rb152-StrepTag), which contains the two known phosphorylation sites of Rb, specifically phosphorylated by Cdk4 in vivo, was used as a substrate in the current in vitro kinase assay. After phosphorylation, scintillation proximity assay (SPA) scintillant beads coated with streptavidin were added. Radiolabeled GST-Rb152-StrepTag was brought in close proximity to the SPA scintillant beads through the interaction between StrepTag and streptavidin, resulting in the emission of light from beads. By applying the affinity peptide tagging technology, we have eliminated the separation and wash steps which are normally required in a radioactive filtration assay. Therefore, this homogeneous method is simple, robust, and highly amenable to high-throughput screening of Cdk4-specific inhibitors. Furthermore, the affinity peptide tagging technique reported here is a simple, generic method that can be applied to many recombinant proteins for the development of kinase and protein-protein interaction assays. PMID- 10702613 TI - A solution-phase combinatorial synthesis of selective dopamine D4 ligands. AB - Utilizing combinatorial synthesis and a preparative LC-MS automated chromatography system we have prepared and purified a library of 4-[2-(1,2,4 oxadiazolyl)]piperidines that were designed to be novel and selective dopamine D4 ligands. In one round of synthesis we identified N-4-chlorobenzyl-4-[2-(3-(2 thienyl)-1,2, 4-oxadiazolyl)]piperidine with a Kd of 5 nM for the human D4 receptor. PMID- 10702614 TI - Generation of a polyclonal fab phage display library to the human breast carcinoma cell line BT-20. AB - We have previously described a vector system for generating recombinant polyclonal antibody libraries. This system uses bidirectional phagemid and mammalian expression vectors to facilitate mass transfer of selected variable light and variable heavy (VL-VH) region gene pairs from the phagemid to the mammalian vector, to express polyclonal libraries of whole IgG antibodies. We report here the first stage of generating a polyclonal antibody library to the human breast carcinoma cell line BT-20, using this vector system. VL and VH region gene pairs were obtained from a mouse immunized with BT-20 cells, and cloned, in opposite transcriptional orientations, in the bidirectional phagemid vector, to produce an Fab phage display library. This library was selected by panning on BT-20 cells and shown to bind specifically to BT-20 cells. Such libraries, after suitable negative selection to eliminate major reactivities against normal tissue, could be transferred in mass to our bidirectional mammalian expression vector for production of libraries of chimeric antibodies with mouse V regions and human constant (C) regions. These polyclonal antibody libraries will mediate effector functions and are expected to be useful for breast cancer therapy, as well as diagnosis. PMID- 10702615 TI - Quantitative structure--activity relationship (QSAR) studies on non steroidal anti-inflammatory drugs (NSAIDs). AB - Different chemical structures have been found to possess different anti inflammatory activities. Inflammation is a normal and essential response to any noxious stimulus which threatens the host and may vary from a localized response to a more generalized one. In view of the complexity and multitude of biochemical factors involved in inflammatory events, few general correlations of chemical structures and physicochemical properties with biological activities would be expected. Nevertheless some general features seem to be commonly associated with a large number of active drugs. However, these main features are not sufficient, but they could reflect certain physicochemical requirements for in vivo efficacy. QSAR is a useful means for maximizing the potency of a new lead compound. In the lead optimization phase of the synthetic project various QSAR procedures with the aid of computer-technology have been proposed. Among them, the classical Hansch approach has been widely used leading to quite a few successful examples. In the QSAR approaches, the prescription to optimise the lead structure is inferred from mathematical equations correlating variations in the potency of a certain biological activity with physicochemical and structural descriptors among congeneric molecules. The QSAR procedures are based on physical organic concepts and involve calculational operations. In the last years, quantum-chemical descriptors have been used in QSAR studies, because of the large physical information content encoded in many of the descriptors. Several anti-inflammatory receptor site models have been proposed. Since inflammation is a complex phenomenon involving interrelationships between humoral and cellular reactions through a number of inflammatory mediators, there is not much evidence on QSAR studies. Several QSAR studies have been reported obtaining only partial results. It was found that substituents which contribute to the high lipophilicity, were favourable to the activity. Substituents of short length (H, CH3) have also a favourable effect. Satisfactory relationships between the in vivo activities and deprotonation energies, the HOMO energies and lipophilicities were found. PMID- 10702616 TI - Medicinal chemistry based on the sugar code: fundamentals of lectinology and experimental strategies with lectins as targets. AB - Theoretical calculations reveal that oligosaccharides are second to no other class of biochemical oligomery in terms of coding capacity. As integral part of cellular glycoconjugates they can serve as recognitive units for receptors (lectins). Having first been detected in plants, lectins are present ubiquitously. Remarkably for this field, they serve as bacterial and viral adhesins. Following a description of these branches of lectinology to illustrate history, current status and potential for medicinal chemistry, we document that lectins are involved in a wide variety of biochemical processes including intra- and intercellular glycoconjugate trafficking, initiation of signal transduction affecting e. g. growth regulation and cell adhesion in animals. It is thus justified to compare crucial carbohydrate epitopes with the postal code ensuring correct mail routing and delivery. In view of the functional relevance of lectins the design of high-affinity reagents to occupy their carbohydrate recognition domains offers the perspective for an attractive source of new drugs. Their applications can be supposed to encompass the use as cell-type-selective determinant for targeted drug delivery and as blocking devices in anti-adhesion therapy during infections and inflammatory disease. To master the task of devising custom-made glycans/glycomimetics for this purpose, the individual enthalpic and entropic contributions in the molecular rendezvous between the sugar receptor under scrutiny and its ligand in the presence of solvent molecules undergoing positional rearrangements need to be understood and rationally exploited. As remunerative means to this end, cleverly orchestrated deployment of a panel of methods is essential. Concerning the carbohydrate ligand, its topological parameters and flexibility are assessed by the combination of computer-assisted molecular-mechanics and molecular-dynamics calculations and NMR spectroscopic measurements. In the presence of the receptor, the latter technique will provide insights into conformational aspects of the bound ligand and into spatial vicinity of the ligand to distinct side chains of amino acids establishing the binding site in solution. Also in solution, the hydrogen-bonding pattern in the complex can be mapped with monodeoxy and monofluoro derivatives of the oligosaccharide. Together with X-ray crystallographic and microcalorimetric studies the limits of a feasible affinity enhancement can be systematically probed. With galactoside-binding lectins as instructive mo del, recent progress in this area of drug design will be documented, emphasizing the general applicability of the outlined interdisciplinary approach. PMID- 10702617 TI - Enaminones-versatile therapeutic pharmacophores. Further advances. AB - Enaminones, enamines of ss-dicarbonyl compounds, have been know for many years. In our initial account (Current Med. Chem. 1994, 1, 159-175), we reported on the anticonvulsant activity of a series of enaminones, notably methyl 4-[(p chlorophenyl)amino]-6-methyl-2-oxo-cyclohex-3-en- 1-oate, 9a (R=CH3, R1=4-Cl), which, in animal tests, compared favorably to phenytoin and carbamazepine. Since that time, further research in our laboratory and other laboratories have expanded the therapeutic potential of these compounds. In addition to new anticonvulsant derivatives, we have uncovered a novel brain transport mechanism for the enaminones and developed a preliminary regression model for further synthetic direction. These topics will each be presented and elaborated. PMID- 10702618 TI - Orally active peptidomimetic RGD analogs that are glycoprotein IIb/IIIa antagonists. AB - Peptidomimetic RGD (Arg-Gly-Asp) analogs, which bind to glycoprotein (GP) IIb/IIIa on the surface of activated platelets, have been shown to inhibit platelet aggregation. Consequently, such RGD analogs can be used for the treatment of unstable angina pectoris and myocardial infarction. However, the low oral bioavailability for this class of compounds has been hindering their clinical development. Although many factors affect the oral activity of a drug, the limited membrane permeability of RGD analogs due to charge and high polarity is thought to be a major factor leading to the low oral activity of such compounds. Another factor is the metabolic lability of some such RGD analogs in the presence of proteases and peptidases. During the last 5 years, major progress has been made in the development of orally active RGD analogs. To improve the metabolic stability of RGD analogs, N-alkylation and C-terminal modification methods have been used successfully. To improve the membrane permeability of RGD analogs, two major strategies have been used. The first one is the strategy of prodrug. Along this line, simple ester prodrugs, double prodrugs, triple prodrugs, and cyclic prodrugs have been prepared with improved membrane permeability and oral activity. The second approach used is the de novo design of centrally constrained RGD analogs with improved oral bioavailability while maintaining the desired potency against GP IIb/IIIa. The lessons learned from the modification of RGD analogs could also help the future design of other peptidomimetic drugs with improved oral bioavailability. PMID- 10702619 TI - Approaches to the design of effective HIV-1 protease inhibitors. AB - Recently, western countries have recorded a decrease in the death rate imputed to AIDS. This success has been largely attributed to the presence on the market of chemotherapies that inhibit the infectivity of the predominant causative agent, the HIV-1 virus, by targeting essential viral enzymes. One of these is the protease (HIV-1 PR) whose activity is a prerequisite for viral replication. Two main sites have been identified as potential targets for the inhibition of HIV-1 PR, the active site and the interface, the latter being largely responsible for the stabilization of the enzyme dimeric structure. The compounds that have reached clinical application so far target the active site of HIV-1 PR. These molecules act as transition state analogues and result from modifications of the peptidic scaffold into peptidomimetics. In order to improve their bioavailability, systematic biological screening and de novo design have been used to suggest new non-peptide inhibitors combining both antiviral potency and favorable pharmacokinetic properties. In parallel, compounds targeting other potential sites of inhibition have been tested. Peptides and peptidomimetics based on the terminal sequence of the enzyme, a site which is proposed to be less susceptible to mutations, have been shown to lead to HIV-1 PR inactivation. Cupric ion was described to bind a sequence on the protease surface, which includes cysteine and histidine residues, leading to the inhibition of the enzyme. In the future, these non-active site inhibitors could provide an alternative in anti-HIV drug combination strategies. PMID- 10702620 TI - Gossypol: prototype of inhibitors targeted to dinucleotide folds. AB - Gossypol, a disesquiterpene from cottonseed, exhibits multiple biological properties, including male antifertility activity and anticancer activity. Gossypol also inhibits the growth of numerous parasitic organisms and shows antiviral activity against a number of enveloped viruses, including the AIDS virus. Derivatives of gossypol, in which the aldehyde functional groups that contribute to toxicity have been modified, retain or even show enhanced biological activity. Ring substituted 2,3-dihydroxy-1-naphthoic acids, which are structural analogs of gossypol, share with gossypol the ability to complex with dehydrogenases at the dinucleotide fold (Rossmann fold) with selectivity, suggesting that gossypol may be considered the prototype of a new class of drugs targeted to dehydrogenases. Most of the biological activities of gossypol and related compounds may result from inhibition of dehydrogenases. PMID- 10702621 TI - Cardiovascular disease risk and hormone replacement therapy (HRT): a review based on randomised, controlled studies in postmenopausal women. AB - Epidemiological data suggest that the use of oestrogen replacement therapy (ERT) and combined oestrogen/progestagen replacement therapy (HRT) in healthy postmenopausal women is associated with a decreased risk of cardiovascular events. In sharp contrast, the HERS study, a secondary prevention trial in postmenopausal women with established coronary heart disease, did not show a favourable effect, with a trend towards an increased risk of cardiovascular disease in the first year of treatment. This paper provides an overview of randomised, controlled trials (RCTs) in postmenopausal women published in the literature and discusses possible explanations for the contrast between data from the epidemiological studies and the results of the HERS study. ERT and HRT are associated with: 1) an improved lipid profile; and 2) a decrease in homocysteine and endothelin levels. Data on factor VII and fibrinogen were not consistent. There were insufficient data on the effects on blood pressure, glucose metabolism, vasomotor regulation, arterial stiffness, thrombomodulin, adhesion molecules, and clotting and fibrinolysis, as well as on the effects of route of administration and the role of progestagens. Finally, endothelium-dependent vasodilatation appears to increase with ERT, but the effects of HRT are less clear This paucity of controlled data indicates that, although ERT and HRT improve surrogate measures of risk of atherothrombosis, adverse effects of ERT and HRT on biological mechanisms related to risk of atherothrombosis can by no means be excluded. PMID- 10702622 TI - Sex steroids and the endothelium. AB - Gonadal steroids clearly influence the course of atherosclerotic cardiovascular disease in women. This observation has suggested that these hormones have beneficial effects on the physiology of the vascular wall. Increased arterial vascular caliber after estrogen treatment, decreased lipid levels in subjects receiving hormone replacement therapy, and the markedly decreased extent of atherosclerotic plaque formation in young women as compared with young men support a cardioprotective effect of ovarian steroids. Generally, it appears that the effects of 17beta-estradiol are particularly beneficial, and the mechanism of action is targeted largely to the endothelial cell. This review describes the evidence for positive effects of estrogens on endothelial cell biology and considers potential mechanisms for estrogen actions on endothelial cell signal transduction. PMID- 10702623 TI - Rapid non-genomic vasodilator actions of oestrogens and sex steroids. AB - There is now convincing experimental evidence documenting acute and transient actions of steroid hormones in the vasculature. Steroids can rapidly activate signalling cascades within endothelial and smooth muscle cells that seem to bypass the classical, genomic receptor. Activation of these signalling cascades, involving alterations in intracellular Ca2+ and MAPK activity, leads to changes in membrane potential and/or Ca2+ fluxes through L-type channels. In addition to stimulating NO production acutely in endothelial cells, chronic exposure to 17beta-oestradiol may activate expression of eNOS via a genomic receptor(s). PMID- 10702624 TI - Steroids and the endometrium. AB - Steroids are commonly employed in current clinical practice. The benefits of steroids in hormone replacement therapy, contraception and prevention or treatment of breast cancer are limited by their side effects arising from disorders in endometrial function. These side effects are complex and enclose bleeding problems and endometrial proliferation during hormone replacement therapy and antioestrogen treatment or menstrual disturbances during oral contraception. Numerous reports have identified gene targets influenced by steroids and have implicated these products as contributors to endometrial physiology or pathology. The expression of estrogen and progesterone receptors is regulated by steroids. The new estrogen receptor (ER) subtype ERbeta with different functional characteristics from ERalpha was recently described in endometrium. In addition, there is now increasing evidence that the functionally distinct progesterone receptor (PR) isoforms A and B are differentially expressed in this tissue. The relative proportions of these steroid receptors and their interaction determine the expression of specific genes upon steroidal stimulation. Steroids induce endometrial expression of various growth and angiogenic factors. Dysregulations of this steroid modulated expression is believed to be involved in the pathogenesis of many endometrial diseases. Irregular bleeding induced by steroidal contraception, for example, is thought to involve aberrant endometrial vascular development and expression of angiogenic growth factors. The antioestrogen tamoxifen induces growth factors like vascular endothelial growth factor and adrenomedullin which may be key mediators of endometrial neoplastic effects. This review describes recent advances regarding the mechanism of action of steroids on endometrium. The expression of oestrogen and progesterone receptors as well as steroid hormone dependent growth factors and angiogenic modulators are going to be discussed. PMID- 10702625 TI - Estrogen receptors alpha and beta: two receptors of a kind? AB - Ever since the discovery of estradiol and the elucidation of its chemical structure, there has been a great deal of interest in its mechanism of action and its potential therapeutic value. It is now well established that estrogens have many different functions in many different cell-types. With respect to the potential use of estrogens as therapeutics, there is an interest in controlling reproductive function, bone metabolism, cardiovascular disease, as well as in the prevention of hot flushes, mood changes and Alzheimer s disease. For over a decade, it was believed that estrogens signal through a a single estrogen receptor, now referred to as ERalpha, which belongs to a family of ligand activated transcription factors. More recently, however, a second estrogen receptor ERbeta was identified. The current review describes similarities as well as differences between these two distinct estrogen receptors. Both ERalpha and ERbeta bind 17beta-estradiol with high affinity and they bind to classical estrogen response elements in a similar if not identical fashion. However, there are also major differences between ERalpha and ERbeta for instance with respect to their tissue distribution, the phenotype of the corresponding knock-out mice and their transcriptional activities. It is anticipated that a better understanding of these two receptors will eventually lead to more selective ways of modulating physiological processes which are influenced by estrogens. For this purpose, the development of ERalpha and ERbeta specific ligands, both agonists as well as antagonists, will be of great importance. PMID- 10702626 TI - Selective estrogen receptor modulators (SERMs): effects on multiple organ systems. AB - SERMs represent a structurally diverse group of compounds which interact with the estrogen receptor but which elicit agonist or antagonist activity depending on the organ system and physiological context. Evaluation of the actions of these compounds has led researchers to a fuller understanding not only of the antiestrogens but also of steroid signaling in general. Based on their evolving clinical profiles, SERMs have the potential to address long-term health maintenance needs of women in their non-reproductive years. PMID- 10702627 TI - Effects on haemostasis variables by second and third generation combined oral contraceptives: a review of directly comparative studies. AB - Previous reports and reviews indicate differences in effects of second and third generation combined oral contraceptives (COCs) on haemostasis variables. This review analyses directly comparative studies on such effects. From the literature, 17 longitudinal comparative studies with parallel groups were retrieved, containing data on comparisons between COCs containing levonorgestrel (second generation COCs) and COCs containing desogestrel, gestodene or norgestimate (third generation COCs) with 30-35 ug ethinylestradiol. Six or more comparisons were available only for fibrinogen, platelet count, antithrombin III, factor VII, factor VIII and factor X. The comparisons reveal a consistently larger increase in factor VII with the third generation COCs compared to the second generation COCs. The effects on factor VII do not coincide in these comparative studies with effects on factor X and prothrombin, rendering a specific sensitivity of the vitamin K-dependent mechanisms for progestogens unlikely. Fibrinogen effects tend to be different for the different progestogens, suggesting a progestogen-specific dependence. Trends in antithrombin III are towards more reduction for the third generation COCs, but the effects are very minor. The effects on factor V suggest a possible progestogen specificity, which may be relevant to explain the difference in APC-resistance between second and third generation of COCs. In general, direct comparisons of effects of different types of COCs on haemostatic variables are available for only a very few factors, which hampers the drawing of general conclusions with respect to haemostatic consequences. PMID- 10702628 TI - Structure-activity relationship of chemical penetration enhancers in transdermal drug delivery. AB - Transdermal drug delivery (TDD) is the administration of therapeutic agents through intact skin for systemic effect. TDD offers several advantages over the conventional dosage forms such as tablets, capsules and injections. Currently there are about eight drugs marketed as transdermal patches. Examples of such products include nitroglycerin (angina pectoris), clonidine (hypertension), scopolamine (motion sickness), nicotine (smoking cessation), fentanil (pain) and estradiol (estrogen deficiency). Since skin is an excellent barrier for drug transport, only potent drugs with appropriate physicochemical properties (low molecular weight, adequate solubility in aqueous and non-aqueous solvents, etc) are suitable candidates for transdermal delivery. Penetration enhancement technology is a challenging development that would increase significantly the number of drugs available for transdermal administration. The permeation of drugs through skin can be enhanced by physical methods such as iontophoresis (application of low level electric current) and phonophoresis (use of ultra sound energy) and by chemical penetration enhancers (CPE). In this review, we have discussed about the CPE which have been investigated for TDD. CPE are compounds that enhance the permeation of drugs across the skin. The CPE increase skin permeability by reversibly altering the physicochemical nature of the stratum corneum, the outer most layer of skin, to reduce its diffusional resistance. These compounds increase skin permeability also by increasing the partition coefficient of the drug into the skin and by increasing the thermodynamic activity of the drug in the vehicle. This review compiles the various CPE used for the enhancement of TDD, the mechanism of action of different chemical enhancers and the structure-activity relationship of selected and extensively studied enhancers such as fatty acids, fatty alcohols and terpenes. Based on the chemical structure of penetration enhancers (such as chain length, polarity, level of unsaturation and presence of some special groups such as ketones), the interaction between the stratum corneum and penetration enhancers may vary which will result in significant differences in penetration enhancement. Our review also discusses the various factors to be considered in the selection of an appropriate penetration enhancer for the development of transdermal delivery systems. PMID- 10702629 TI - New trends in thromboxane and prostacyclin modulators. AB - Thromboxane A2 (TXA2) and prostacyclin (PGI2) are two labile products formed from arachidonic acid by the way of cyclooxygenase. An overproduction of thromboxane A2 has been detected in a series of diseases whereby this prostanoid is assumed to contribute to the underlying pathomechanisms by its potent stimulation of platelet aggregation and smooth muscle contraction. This increased TXA2 biosynthesis is frequently accompanied by a stimulation of prostacyclin formation which is one of the most potent inhibitors of platelet aggregation and smooth muscle contraction. Therefore, TXA2 / prostaglandin endoperoxide H2 receptor antagonists, thromboxane synthase inhibitors and drugs which combine both activities have been developed with the aim to suppress the formation and/or the action of thromboxane A2. Since prostacyclin has been demonstrated to counterbalance the pathological effects of TXA2, several PGI2 agonists have also been developed. This review will highlight the evolution and some of the latest findings in the field of prostacyclin and thromboxane A2 modulators mainly those which are under clinical evaluation or marketed. PMID- 10702630 TI - Phosphinic acid compounds in biochemistry, biology and medicine. AB - This review summarizes our knowledge of biochemical, biological and medical applications and properties of phosphinic acid compounds. Phosphinic acid compounds (phosphinates) are derivatives of phosphinic acid H2P(O)(OH). The major attention of this article is focused on applications of phosphinates of a pseudopeptide character, however interesting examples of phosphinates of a non peptide nature are mentioned too. Phosphinic acid peptides (phosphinic pseudopeptides) are peptide isosteres where one peptide bond is substituted by the nonhydrolysable phosphinate moiety -P(O)(OH)-CH2- or -P(O)(OH)-. This substitution represents a very convenient mimic of a substrate in the transition state for at least two distinct classes of hydrolytic enzymes, Zn metalloproteinases and aspartic acid proteinases. In this review we discuss about thirty different protein targets for which the phosphinates have found applications as modulators of their functions in vitro and/or in vivo. These proteins are mainly proteinases, however other types of proteins such as transferases, synthetases, ligases or even receptors are also discussed. Genome sequencing projects have been identifying protein sequences faster than it is possible to discover their functions. The development of combinatorial chemistry in the past few years has boosted up the interest in the use of chemistry to address biological problems. We believe that phosphinates, especially in conjunction with combinatorial chemistry approaches, can represent an extremely versatile tool in the search for proteome and its function. PMID- 10702631 TI - Recent studies on natural products as anti-HIV agents. AB - The discovery of medicinal agents capable of specifically inhibiting human immunodeficiency virus (HIV) is urgently needed due to its globally widespread infection. Most of clinically useful anti-HIV agents are nucleosides but their use is limited due to their severe toxicity and emerging drug resistance. More than 50% of world marketed drugs have their origin of the nature. Natural products, of which structural diversity is so broad, are good sources for the effective discovery of anti-HIV agents with decreased toxicity. Over the past decade, substantial progress has been made in research on the natural products for the anti-HIV agents. New natural products that have potent anti-HIV activities with novel structures were reviewed in this article. These compounds, isolated mainly from medicinal plants, in this review have been classified as secondary metabolites such as terpenes, phenolics, and naturally scarce peptides and sugars. Especially, terpenes and phenol substances have gained much interest due to their significant anti-HIV activities along with their structural diversity. Recent studies also showed that several polysaccharides are effective inhibitors of HIV replication. Most of chemotherapeutic targets reviewed in this article are found to be HIV reverse transcriptase (RT). PMID- 10702632 TI - Discovery and development of GS 4104 (oseltamivir): an orally active influenza neuraminidase inhibitor. AB - Rational drug design utilizing available X-ray crystal structures of sialic acid analogues bound to the active site of influenza virus neuraminidase has led to the discovery of a series of potent carbocyclic influenza neuraminidase inhibitors. From this series, GS 4104 (oseltamivir, TAMIFLU ) has emerged as a promising antiviral for the treatment and prophylaxis of human influenza infection. This article will summarize the design, discovery, and development of oseltamivir as an oral therapeutic to treat influenza infection. PMID- 10702633 TI - The impact of immunosuppressive drugs on the analysis of T cell activation. AB - The discovery of the immunosuppressive properties of cyclosporin A (CSA) and its successful utilisation in organ transplantation was a milestone in clinics. CSA has revolutionised transplantation both in term of efficiency and quality-of-life of the patient. In addition, the analysis of the mode of action of CSA has been rewarding in the understanding the mechanisms leading to T lymphocytes activation. CSA binds to a family of cytosolic receptors, the cyclophilins, a highly conserved family of proteins. Once this complex is formed, a third protein, the calcineurin, is recruited. The calcineurin, a calcium-dependent phosphatase, loose its activity when complexed. Dephosphorylation of NFAT, a substrate of calcineurin is a mandatory step for its translocation to the nucleus where NFAT acts as a transactivator involved in the regulation of the genes encoding many cytokines. CSA preventing NFAT dephosphorylation blocks cytokine production this in turn allowing for a better engrafting. The resolution of the tertiary structure of CSA alone or complexed with cyclophilin and calcineurin has important implication in the modelling of new drugs devoid of its side effects. Indeed, the high incidence of cancer is one of the main problems linked to CSA utilisation. Recent data suggest that CSA may promote cancer inducing the transcription of the gene encoding transforming growth factor beta. Other molecules sharing with CSA its immunosuppressive activity were later described. Some of them, as FK506, have the some mode of action; others, as rapamycin, mycophenolate mofetil or leflunomide, act at different steps of T cell activation. PMID- 10702634 TI - Apoptosis at the interface of immunosuppressive and anticancer activities: the examples of two classes of chemical inducers, oxysterols and alkylating agents. AB - Recent progresses in the understanding of molecular and biochemical pathways involved in apoptotic cell death offer novel perspectives for therapeutic interventions, in particular in immunosuppressive and anti-cancer therapies. In this review, we examine some chemical, biological, and mechanistic aspects of two classes of apoptosis chemical inducers: oxysterols and alkylating agents. Oxysterols represent a vast family of oxygenated derivatives of sterols. Found in both animal and vegetal kingdoms, they can be considered as ultimate products of an oxidative stress, and are chemically inert. Some of them (7beta hydroxycholesterol, 25-hydroxycholesterol and 7, 25-dihydroxycholesterol) are cytotoxic at micromolar concentrations towards normal and tumor cells in culture, particularly lymphocytes, and reduce the growth of murine transplanted tumors. Thus, possible applications of oxysterols in medicine as immunosuppressants or as anticancer agents may be considered. Alkylating agents, on the other hand, have been widely used in cancer chemotherapy for decades. There toxicity results from their high chemical reactivity, causing lesions to macromolecules through covalent linkage. Some representative members of this class, mainly bifunctional derivatives which possess dichloroethyl groups, such as Chlormethine, Cyclophosphamide and Chlorabucil, express a pronounced cytotoxicity against lymphoid cells, and have therefore potent immunosuppressive properties. Because they triggers apoptosis via both common and distinct mechanisms, oxysterols and alkylating agents provide unique tools for exploring the initiation of this phenomenon in lymphoid cells, and may help design novel pharmacological approaches based on apoptotic modulation of these cells. PMID- 10702635 TI - Immunomodulation and anti-cancer activity of polysaccharide-protein complexes. AB - In the last three decades, numerous polysaccharides and polysaccharide-protein complexes have been isolated from mushrooms and used as a source of therapeutic agents. The most promising biopharmacological activities of these biopolymers are their immunomodulation and anti-cancer effects. They are mainly present as glucans with different types of glycosidic linkages such as (1-->3), (1-->6)-beta glucans and (1-->3)-alpha-glucans, and as true herteroglycans, while others mostly bind to protein residues as polysaccharide-protein complexes. Three antitumor mushroom polysaccharides, i.e. lentinan, schizophyllan and protein bound polysaccharide (PSK, Krestin), isolated respectively, from Lentinus edodes, Schizophyllum commune and Coriolus versicolor, have become large market items in Japan. Lentinan and schizophyllan are pure beta-glucans, whereas PSK is a protein bound beta-glucan. A polysaccharide peptide (PSP), isolated from a strain of Coriolus versicolor in China, has also been widely used as an anti-cancer and immunomodulatory agent. Although the mechansim of their antitumor action is still not completely clear, these polysaccharides and polysaccharide-protein complexes are suggested to enhance cell-mediated immune responses in vivo and in vitro and act as biological response modifiers. Potentiation of the host defense system may result in the activation of many kinds of immune cells that are vitally important for the maintenance of homeostasis. Polysaccharides or polysaccharide-protein complexes are considered as multi-cytokine inducers that are able to induce gene expression of vaious immunomodulatory cytokines and cytokine receptors. Some interesting studies focus on investigation of the relationship between their structure and antitumor activity, elucidation of their antitumor mechanism at the molecular level, and improvement of their various biological activities by chemical modifications. PMID- 10702636 TI - FK506, an immunosuppressant targeting calcineurin function. AB - The macrolactam natural product, FK506 (Tacrolimus), acts as a powerful and clinically useful immnosuppressant through disruption of signaling events mediated by the calcium-dependent serine/threonine protein phosphatase, calcineurin (CaN), in T lymphocytes. Its mechanism of action involves the formation of a molecular complex with the intracellular FK506-binding protein-12 (FKBP12), thereby acquiring the ability to interact with CaN and to interfere with its access to and dephosphorylation of various substrates. Among the CaN substrates whose activity is altered by FK506, the nuclear factors of activated T cells (NFAT), a family of transcription factors regulating lymphokine gene expression, have been shown to play a prominent role in FK506-induced immunosuppression. Over the past few years, additional members of the FKBP and NFAT families of proteins have been identified, providing further insights into the complexity of FK506 biological effects. Furthermore, it has become clear that, predominantly as a result of CaN inhibition, FK506 alters multiple biochemical processes in a variety of cells besides lymphocytes. This may account for the adverse side effects of the drug, including neurotoxicity and nephrotoxicity. Extensive medicinal chemistry efforts have been devoted to the generation of analogs of FK506 with the hope of identifying compounds with an improved therapeutic index, that could have broader therapeutic utility than the parent drug. These efforts yielded several compounds with unique biochemical attributes, showing evidence for a dissociation between immunosuppressive and toxic properties, which may pave the way towards designing safer FK506-related immunosuppressants. PMID- 10702637 TI - Targets of oncogenes and tumor suppressors: key for understanding basic mechanisms of carcinogenesis. AB - Changes in expression of protooncogenes and tumor suppressor genes play a key role in oncogenesis. Dysfunction of their protein products leads to abnormal regulation of signaling pathways, which control the cell cycle, apoptosis, genetic stability, cell differentiation, and morphogenetic reactions. Changes in these important physiological processes are obviously responsible both for initial steps of neoplastic cell transformation and for determination of subsequent tumor progression resulting in the development of malignant tumors. PMID- 10702638 TI - Function of the p53 gene: choice between life and death. AB - Gene p53 is a central component of a system that eliminates pathologically damaged cells from an organism. Multiple signal pathways monitor the state of a cell and when damage or a fault is found that could cause heritable changes, p53 protein is activated to either coordinate the repair process or induce cell suicide. Thus, the p53 gene acts as a supreme judge that decides the fate of cells and guarantees their social behavior. Loss of the p53 gene results in uncontrolled accumulation of genetic damage causing failure of control by the organism, malignant cell growth, and death of the organism. PMID- 10702639 TI - Suppression of p53: a new approach to overcome side effects of antitumor therapy. AB - The p53 protein is traditionally believed to be a tumor suppressor. Activation of p53-dependent apoptosis in response to damage to cell DNA provides for the elimination of possible tumor cell precursors. However, in some cases the activity of p53 can be dangerous for the organism. Thus, p53-dependent apoptosis induced in normal tissues during chemo- and radiotherapy can cause severe side effects of antitumor therapy and, therefore, limits its efficiency. This review analyzes experimental data on the role of p53 in the primary and late tissue response to DNA-damaging exposures. Comparison of normal and p53-deficient mice indicated that the apoptosis in radiosensitive tissues during the first hours after irradiation is really caused by the activity of p53 which, in turn, is determined by a high level of expression of mRNA of p53. We supposed that a temporary suppression of p53 can decrease the damage to sensitive tissues and accelerate their recovery after the antitumor radio- and chemotherapy. To test this hypothesis, we have isolated a chemical inhibitor of p53 and determined its activity in vitro and in vivo. This compound, called pifithrin-alpha, protects wild-type mice against lethal doses of radiation, has no effect on p53-deficient animals, and does not induce visible tumors. These results show that the suppression of p53 is a promising approach in the prevention of side effects of antitumor therapy. PMID- 10702640 TI - Kinases of the Src family: structure and functions. AB - Tyrosine kinases of the Src family are involved in different signal transduction pathways in cells. The corresponding genes participate in such vital processes as growth, differentiation, adhesion, transcription, etc. Specific structural changes confer oncogenic properties to the Src protein. In this review, we summarize the available data on the structure, substrates, regulation mechanisms, and role of nonreceptor tyrosine kinases by the example of the src gene product (as the prototype member of this family) and a number of related proteins. PMID- 10702641 TI - Phosphatidylinositol-3 kinase dependent pathways: the role in control of cell growth, survival, and malignant transformation. AB - Phosphatidylinositol-3 kinase (PI3K) is one of the most important regulatory proteins that is involved in different signaling pathways and controlling of key functions of the cell. The double-enzymatic activity of PI3K (lipid kinase and protein kinase) as well as the ability of this enzyme to activate a number of signal proteins including some oncoproteins determines its fundamental significance in regulation of cell functions such as growth and survival, aging, and malignant transformation. Among the main effectors of PI3K are the mitogen transducing signal proteins (protein kinase C, phosphoinositide-dependent kinases, small G-proteins, MAP (mitogen activated protein) kinases), which are activated either via their interaction with lipid products of PI3K or through PI3K-dependent phosphorylation of proteins. The anti-apoptotic effect of PI3K is realized by activation of proteins from another signaling pathway--protein kinase B (PKB) and/or PKB-dependent enzymes (GSK-3, ILK). PI3K plays a critical role in malignant transformation. PI3K itself possesses oncogenic activity and also forms complexes with some viral or cellular oncoproteins (src, ras, rac, alb, T antigen), whose transforming activities are realized only in presence of PI3K. The transforming effect of PI3K is supposed to occur on the basis of complex alterations in cellular signaling pathways: appearance of constitutively generated PI3K-dependent mitogen signal and activation of some protooncogenes (src, ras, rac, etc.), PI3K/PKB-pathway stimulation resulting in delay of apoptosis and increase of cell survival, and actin cytoskeleton reorganization. PMID- 10702642 TI - Virus-associated human tumors: cervical carcinomas and papilloma viruses. AB - The latest experimental data on the role of viruses in the origin of human tumors are discussed. This group of viruses consists of T-cell leukemia virus type 1 (HTLV 1), herpes viruses (HHV 8 and Epstein-Barr virus), hepatitis B virus, and human papilloma viruses. The most typical feature of this group of viruses is a very long latent period from the initial infection to the development of the disease that varies between 10 and 40 years. The mechanism of malignant cell conversion is specific for each viral type but is mainly associated with a disruption of functions of cellular genes participating in the control of cell division and proliferation. It can be a direct inactivation of tumor suppressor genes by their interaction with viral gene products (papilloma viruses), or a trans-activation of cellular genes modulating cell proliferation by viral gene products (hepatitis B virus and HTLV 1). Viruses play an initiative role and additional genetic changes in the genome of infected cells are necessary for complete expression of the oncogenic potential of the viral genes. Only these cells will give rise to a monoclonal cell population with uncontrolled proliferation. New approaches for the creation of vaccines against cancers associated with hepatitis B virus and papilloma viruses (hepatocellular carcinomas and cervical tumors, respectively) are in progress. These vaccines have been found to be effective in prevention of the disease in the experimental models and are now beginning to be used for human vaccination. PMID- 10702643 TI - Natural selection and early changes of phenotype of tumor cells in vivo: acquisition of new defense mechanisms. AB - This review summarizes results obtained in the author's and collaborating laboratories within the last decade and is designed to attract the attention of researchers to discrete biochemical mechanisms of protection acquired in vivo by cells of malignant tumors against effectors of innate antitumor immunity. Tumor progression in vivo is associated with the appearance and selection of tumor cells with new specific characteristics: a high level of H(2)O(2)-catabolizing (antioxidant) activity (H(2)O(2)CA) and the ability for immediate release of E2 type prostaglandin (PGES) on contact with natural killers, macrophages, and neutrophils; the expression of the [H(2)O(2)CA + PGES] phenotype provides the tumor cells with two mechanisms of local protection against effectors of innate and acquired antitumor immunity. This results in a 10-100-fold less effective rejection of tumor cells in immune and normal animals and corresponding increase of tumorigenicity. The in vitro transformation of normal fibroblasts, spontaneous or induced by oncogenes LTSV40, E1a,b, Ha-ras, myc, and also by p53(175) and bcl 2 does not result in the [H(2)O(2)CA + PGES] phenotype expression, but during subsequent in vivo growth of the above-mentioned transformants the selection of tumor cells of the [H(2)O(2)CA + PGES] phenotype is correlated with a 30-200-fold increase in their tumorigenicity (accompanied or not accompanied by spontaneous metastatic activity). Unlike the transformation induced by the above-mentioned oncogenes, the transformation of normal cells by the v-src gene results in the [H(2)O(2)CA + PGES] phenotype expression. The data presented confirm the determining role of the v-src gene in the expression of the [H(2)O(2)CA + PGES] phenotype. In various primary viral carcinogenesis (SV40, SA7(C8)) the natural selection of cells expressing the [H(2)O(2)CA + PGES] phenotype begins even within the latent period and can be completed by the appearance of primary tumors. PMID- 10702644 TI - Cellular mechanisms of multidrug resistance of tumor cells. AB - Multidrug resistance (MDR) is the protection of a tumor cell population against numerous drugs differing in chemical structure and mechanisms of influence on the cells. MDR is one of the major causes of failures of chemotherapy of human malignancies. Recent studies show that the molecular mechanisms of MDR are numerous. Cellular drug resistance is mediated by different mechanisms operating at different steps of the cytotoxic action of the drug from a decrease of drug accumulation in the cell to the abrogation of apoptosis induced by the chemical substance. Often several different mechanisms are switched on in the cells, but usually one major mechanism is operating. The most investigated mechanisms with known clinical significance are: a) activation of transmembrane proteins effluxing different chemical substances from the cells (P-glycoprotein is the most known efflux pump); b) activation of the enzymes of the glutathione detoxification system; c) alterations of the genes and the proteins involved into the control of apoptosis (especially p53 and Bcl-2). PMID- 10702645 TI - Differentiation mechanisms and malignancy. AB - This review considers the relationship between differentiation mechanisms and the genesis and maintenance of tumor phenotype. To a certain extent, carcinomas preserve differentiation markers of normal tissue, and hemoblastoses precisely reflect the direction and differentiation level of their precursor cells. Both tumor types retain the ability to differentiate. Mechanisms of T and B cell differentiation are reviewed considering the activation of protooncogenes by translocation to the region of tissue-specific genes including the immunoglobulin (Ig) and T cell receptor (TCR) genes. Apart from the classical oncogenes (MYC, PRAD, BCL-2), heterologous differentiation of trans-factors can be activated in a similar manner. Their activation at inappropriate time and place induces oncogenic transformation in a number of hemoblastoses. Chimeric genes and fused proteins are analyzed, including their genesis by specific translocation resulting in transformation and their role in differentiation and maintenance of the tumor phenotype. Induction of terminal differentiation in leukemia can have significant therapeutic effect. These hemoblastoses include hairy cell leukemia, promyelocytic leukemia, and in part chronic myeloid leukemia. Specific attention is given to the role of intercellular interactions in the control of tumor growth and maintenance of a differentiated state of the cells. It is suggested that alterations in these interactions during tumor progression simultaneously stimulate malignant growth and decrease differentiation level, thus inducing re expression of embryonic antigens in the tumors. PMID- 10702646 TI - Molecular mechanisms of alpha-fetoprotein gene expression. AB - Alpha-fetoprotein (AFP) is the main component of mammalian fetal serum. It is synthesized by visceral endoderm of the yolk sac and by fetal liver. Immediately after birth AFP level in blood decreases dramatically. AFP synthesis is reactivated in liver tumors and germinogeneous teratoblastomas, in a lesser degree after chemical and mechanical liver injuries followed by regeneration (i.e., acute viral hepatitis). AFP blood level change is an important marker for liver tumors that is widely used in clinical practice. Therefore, the study of the molecular and cellular mechanisms participating in regulation of the oncoembryonal protein AFP is an important task. On various experimental models it has been shown that the expression is regulated mainly on the transcriptional level, the AFP gene having a 7 kb regulatory region upstream. Within this region a tissue-specific promoter, three independent enhancers, and a silencer that is at least partially responsible for AFP gene expression decrease in adult liver have been defined. Some ubiquitous and some tissue-specific transcription factors, including hepatocyte nuclear factors (HNFs), which mediate the transcription of most of the liver-specific genes, have been shown to bind to the promoter. However, the mechanisms determining drastic changes of AFP synthesis level in the course of ontogenesis and carcinogenesis remain incompletely clarified. Also, little is known about negative regulators of AFP gene expression in cells of non-hepatic origin and in adult liver. PMID- 10702647 TI - Scintigraphic predictor of left ventricular size after acute myocardial infarction. AB - The aim of this study was to evaluate the relation between thallium-201 scintigraphic indices and left ventricular size after acute myocardial infarction. Forty-seven patients with acute myocardial infarction underwent rest redistribution thallium-201 scintigraphy at 2 weeks and left ventriculography at 4 weeks, after the onset of myocardial infarction. Percent (%) fixed defect, %redistribution and %reverse redistribution, calculated as a percentage of whole left ventricular area, were quantified with computer-generated unfolded map method of the myocardial radioactivity. Despite no significant difference in peak plasma creatine phosphokinase between the two groups, patients with anterior myocardial infarction (28 patients) had larger %fixed defect (p < 0.01), which was associated with higher end-diastolic pressure (p < 0.05) and larger end diastolic volume index (p < 0.01) than those with inferior myocardial infarction (19 patients). End-diastolic volume index was not related to %redistribution and %reverse redistribution, but there was a good relation between end-diastolic volume index and %fixed defect in anterior (r = 0.79, p < 0.001) and in inferior (r = 0.73, p < 0.001) myocardial infarction. However, left ventricular end diastolic volume index in anterior myocardial infarction was larger than that of inferior myocardial infarction at any given %fixed defect. Thus, site as well as size of fixed defect at 2 weeks after the onset of acute myocardial infarction was related to left ventricular end-diastolic volume at chronic phase. PMID- 10702648 TI - Obstructive sleep apnea as a risk marker in coronary artery disease. AB - STUDY OBJECTIVES: Obstructive sleep apnea (OSA) is associated with a range of cardiovascular sequelae and increased cardiovascular mortality. The aim of our study was to assess the prevalence of OSA in patients with symptomatic angina and angiographically verified coronary artery disease (CAD). In addition, we analyzed the association of OSA and other coronary risk factors with CAD and myocardial infarction. METHODS: Overnight non-laboratory-monitoring-system recordings for detection of OSA was performed in 223 male patients with angiographically verified CAD and in 66 male patients with exclusion of CAD. A logistic regression analysis was performed to assess associations between risk factors and CAD and myocardial infarction. RESULTS: CAD patients were found to have OSA in 30.5%, whereas OSA was found in control subjects in 19.7%. The mean apnea/hypopnea index (AHI) was significantly higher (p < 0.01) in CAD patients (9.9 +/- 11.8) than in control subjects (6.7 +/- 7.3). Body-mass-index (BMI) was significantly higher in patients with CAD and OSA than in patients with CAD without OSA (28. 1 vs. 26.7 kg/m(2); p < 0.001). No significant difference was found with regard to other risk factors and left ventricular ejection fraction (LVEF) between both groups. Hyperlipidemia (OR 2.3; CI 1. 3-3.9; p < 0.005) and OSA defined as AHI >/=20 (OR 2.0; CI 1.0-3.8, p < 0.05) were independently associated with myocardial infarction. CONCLUSIONS: There is a high prevalence of OSA among patients with angiographically proven CAD. OSA of moderate severity (AHI >/=20) is independently associated with myocardial infarction. Thus, in the care of patients with CAD, particular vigilance for OSA is important. PMID- 10702649 TI - Endothelium-dependent vasodilator response is augmented in peripheral resistance vessels of patients with vasospastic angina. AB - Although the mechanism underlying coronary conduit artery spasm in vasospastic angina (VAP) remains unknown, coronary endothelial dysfunction has been suggested as playing an important role. However, it remains unknown whether this endothelium-mediated abnormal vasomotion is uniformly evident irrespective of site of vascular bed or vessel size. Plethysmographic studies were carried out to measure changes in forearm resistance vessel blood flow (FBF) induced by acetylcholine (Ach), sodium nitroprusside, and 10-min occlusion-induced reactive hyperemia in 12 patients with VAP, 14 patients with atherosclerotic (>75% fixed stenosis) coronary artery disease (CAD), and 16 healthy controls. FBF responses induced by the endothelium-dependent vasodilator Ach were significantly augmented in patients with VAP compared to controls, whereas this type of FBF response in patients with CAD was significantly blunted (at the maximum dose: VAP, 24.1 +/- 3.0; controls, 17.2 +/- 1.9; CAD, 12.8 +/- 1.9 ml/min per 100 ml; p < 0.01). However, there were no significant differences in FBF responses during infusion of sodium nitroprusside among the three subject groups. Reactive hyperemic FBF which represents maximum vasodilatory capacity did not differ among the three. To assess the role of nitric oxide in the augmented endothelium-dependent response in VAP, Ach-induced FBF response was determined before and after administration of the nitric oxide synthase inhibitor N(G)-monomethyl-L-arginine in another VAP group (n = 7). After nitric oxide synthase inhibition, the peak FBF response induced by Ach decreased from 23.3 +/- 3.2 to 14.1 +/- 1.6 ml/min per 100 ml (p < 0.05). In conclusion, the above data suggest that endothelium-dependent vasomotor response to Ach in limb resistance vessels is augmented in patients with vasospastic angina. It seems unlikely that endothelial function is consistently depressed in any area of the vascular bed in this disorder. PMID- 10702650 TI - Outcome of patients with malignant hypertension and end-stage renal failure treated by long-term hemodialysis. AB - Malignant hypertension is associated with high mortality and morbidity usually caused by cardiovascular events. The course and prognosis of malignant hypertension patients treated with renal replacement therapy has not been thoroughly investigated. In the present work, we compared the clinical evolution and survival of 24 end-stage renal failure malignant hypertension patients with that of a group of individually matched renal failure patients admitted to the same dialysis center during a period of 21 years. Survival rates at 1, 5 and 8 years were 87, 82 and 50% for malignant hypertension patients and 87, 75 and 65% for controls, respectively (p = 0.766, NS). Nonfatal cardiovascular complications occurred in 2 individuals of each group. The most important cause of death in both groups was cardiovascular. The frequency of fatal cardiovascular events was similar in the two groups: 64% of deaths for malignant hypertension and 71% for controls (NS). In conclusion, previous malignant hypertension did not increase the risk of patients in long-term hemodialysis in our series. PMID- 10702651 TI - Beta-blockers after myocardial infarction: do benefits ever outweigh risks in asthma? AB - beta-Blockers are well documented to prolong life in patients after myocardial infarction (MI), yet patients who also have asthma are frequently and understandably denied this therapy. We reviewed the literature (via MEDLINE) for the past 35 years for beta-blocker-induced asthma, and reexamined potential beta blocker use in the context of NIH guidelines for asthma classification and management. Because beta-blockers can cause fatal or life-threatening asthma, their use should be avoided in moderate to severe persistent asthmatics. Benefits of low-dose beta(1)-blockers (e.g. atenolol 50 mg daily) may outweigh risks in some patients with mild intermittent or well-controlled mild persistent asthma. Further study is needed to verify that low doses of beta(1)-blockers are effective in prolonging life after MI, and that use specifically in mild intermittent or mild persistent asthma per NIH classification is safe. PMID- 10702652 TI - ACE inhibitors suppress ischemia-induced arrhythmias by reducing the spatial dispersion of ventricular repolarization. AB - The effect of angiotensin-converting enzyme (ACE) inhibitors on ischemia-induced spatial dispersion of ventricular repolarization was investigated in 12 pentobarbitone-anesthetized sheep. The obtuse marginal coronary artery was occluded in the pretreated animals (enalapril maleate, 0.4 mg/kg, i.v., n = 6) and controls (normal saline, i.v., n = 6). The activation-recovery intervals were determined from the unipolar ECGs acquired from the ischemic region. There was a significant increase in the pooled activation-recovery interval dispersion in both groups at 30 min of coronary occlusion (p < 0.01), however, the increase in the treatment group was smaller than that of the controls (15.9 +/- 9.7 vs. 43.6 +/- 19.9 ms, p < 0. 01). Ventricular ectopic beats were observed in the 6 controls and in only 1 pretreated animal. CONCLUSIONS: ACE inhibitors suppress the spatial dispersion of ventricular repolarization and this may be attributed to, at least in part, its antiarrhythmic effect. PMID- 10702653 TI - Effects of losartan titrated to Losartan/Hydrochlorothiazide and amlodipine on left ventricular mass in patients with mild-to-moderate hypertension. A double blind randomized controlled study. AB - To study the effects of the angiotensin II receptor antagonist Losartan and Amlodipine on left ventricular mass (LVM), we performed blood pressure measurements and transthoracic echocardiographies at baseline. After a 4-week placebo run-in period, 25 patients with mild-to-moderate essential hypertension were randomly allocated to active treatment with Losartan 50 mg titrated to Losartan 50 mg/hydrochlorothiazide (HCT) 12.5 mg (n = 11) or Amlodipine 5 mg titrated to 10 mg (n = 14) for 16 weeks. After treatment, blood pressure decreased significantly in both groups. LVM and LVM index (mean +/- SD/median) in the Losartan group at baseline were 311 +/- 101/288 g and 163 +/- 55/150 g/m(2) and decreased significantly to 252 +/- 25/255 g and 133 +/- 22/128 g/m(2) (p = 0.003 for LVM; p = 0. 01 for LVM index) after 16 weeks of active treatment. In the Amlodipine group LVM and LVM index decreased from 259 +/- 47/243 g and 136 +/ 25/ 131 g/m(2) to 240 +/- 42/234 g and 126 +/- 24/123 g/m(2) (n.s.). In conclusion, LVM decreased significantly as early as 16 weeks after initiation of antihypertensive treatment with the Angiotensin II antagonist Losartan. PMID- 10702654 TI - Pain recollection after chest pain of cardiac origin. AB - Memory for pain is an important research and clinical issue since patients ability to accurately recall pain plays a prominent role in medical practice. The purpose of this prospective study was to find out if patients, with an episode of chest pain due to suspected acute myocardial infarction could accurately retrieve the pain initially experienced at home and during the first day of hospitalization after 6 months. A total of 177 patients were included in this analysis. The patients rated their experience of pain on a numerical rating scale. The maximal pain at home was retrospectively assessed, thereafter pain assessments were made at several points of time after admission. After 6 months they were asked to recall the intensity of pain and once again rate it on the numerical rating scale. The results from the initial and 6-month registrations were compared. In general, patients rated their maximal intensity of chest pain as being higher at the 6-month recollection as compared with the assessments made during the initial hospitalization. In particular, in patients with a high level of emotional distress, there was a systematic overestimation of the pain intensity at recall. PMID- 10702655 TI - The prognostic implications of negative T waves in the leads with ST segment elevation on admission in acute myocardial infarction. AB - We assessed the prognostic significance of negative T waves on admission in leads with ST elevation in 2,853 patients with acute myocardial infarction treated with thrombolysis. Patients were classified into 2 groups based on the presence of negative (T-) or positive (T+) T waves in the leads with ST elevation on admission. T+ and T- waves on admission were detected in 2,601 (91%) and 252 (9%) patients, respectively. T- waves were observed in 6.7 and 9.6% of patients admitted 2 h after symptom onset. T- patients admitted 2 h after onset suffered higher mortality (20/196 patients; 10.2%) than T+ patients (100/1,836 patients; 5.4%; p = 0.01). Multivariate analysis of the data on patients treated >2 h after onset demonstrated T- waves to be associated with mortality (OR 1.86; 95% CI 1.07-3.25; p = 0.017). T- waves in leads with ST elevation upon admission are associated with adverse prognosis in patients presenting >2 h after symptom onset, whereas in patients presenting p21. PMID- 10702665 TI - Assignment of laminin alpha 2-chain gene (Lama2) to mouse chromosome 10A4-B1 by fluorescence in situ hybridization. PMID- 10702664 TI - cDNA cloning, characterization and chromosome mapping of the gene encoding human cartilage associated protein (CRTAP). AB - We have recently isolated and characterized cDNA clones coding for a novel developmentally regulated avian and mouse embryo protein, CASP for Cartilage Associated Protein. Here we describe the isolation and characterization of the gene coding for the human CASP. The comparison of the putative human and mouse protein sequences with the chick sequence revealed an overall high identity (89% and 51%, respectively). Homology search with known DNA and protein sequences showed that CASPs are related to two mammalian nuclear proteins. Here we demonstrate definitively that CASPs are distinct from these nuclear proteins. However, sequence comparison analyses suggest that all of these proteins belong to a new family. In all human tissues examined two CASP mRNA species were detected, whereas a single mRNA and three mRNAs were found in chick and mouse, respectively. The human CASP gene (CRTAP) was assigned to chromosome 3p22 by fluorescence in situ hybridization. PMID- 10702666 TI - Assignment of the SLC25A12 gene coding for the human calcium-binding mitochondrial solute carrier protein aralar to human chromosome 2q24. PMID- 10702667 TI - Genetic mapping of the mouse homologue of the human angiopoietin-1 gene (Agpt) to mouse chromosome 9E2 by in situ hybridization. PMID- 10702668 TI - Identification of the mouse beta'-COP Golgi component as a spermatocyte autoantigen in scleroderma and mapping of its gene Copb2 to mouse chromosome 9. AB - In an immunological screening of a mouse testicular cDNA library with a human CREST serum we isolated five overlapping cDNA clones encoding the mouse homolog of a Golgi coatomer complex protein (accession number AF043120), designated beta' COP in bovine and p102 in humans. We generated antibodies against this protein which specifically recognize the Golgi apparatus of mouse spermatocytes. FISH analyses assigned the beta'-COP gene Copb2 to mouse Chromosome 9, region E3-F1. Our results demonstrate that CREST sera can contain antibody components against Golgi proteins as well as against nuclear proteins. PMID- 10702670 TI - Assignment of the human PLC delta3 gene (PLCD3) to human chromosome band 17q21 by fluorescence in situ hybridization. PMID- 10702669 TI - VPREB3: cDNA characterization and expression in human and chromosome mapping in human and mouse. AB - The pre-B cell receptor (pre-BCR) regulates pre-B cell expansion and allelic exclusion at the immunoglobulin (Ig) heavy chain locus and mediates the selection of Ig heavy chain variable gene segments. During the early phase of pre-BCR assembly in the mouse, the membrane Ig mu heavy chain transiently associates with the VPREB3 protein in the endoplasmic reticulum. Here, we present the human VPREB3 cDNA sequence and its B cell-specific expression in hematopoietic cell lines. We have localized this gene to chromosome 22q11 close to IGLL genes in human and to chromosome 10C in mouse. PMID- 10702671 TI - Assignment of stra13 to the sub-telomeric region of mouse chromosome 6 by in situ hybridization. PMID- 10702673 TI - Assignment of amyloid-precursor-like protein 2 gene (APLP2) to 11q24 by fluorescent in situ hybridization. PMID- 10702672 TI - Assignment of the yeast APG5 human homologue APG5L to chromosome band 6q21 by fluorescence in situ hybridisation. PMID- 10702674 TI - Assignment of the rat Crebbp and Rxrip13 genes to chromosome bands 10q12-->q21 and 10q24 respectively by in situ hybridization. PMID- 10702675 TI - Assignment of ACVR2 and ACVR2B the human activin receptor type II and IIB genes to chromosome bands 2q22.2-->q23.3 and 3p22 and the human follistatin gene (FST) to chromosome 5q11.2 by FISH. PMID- 10702676 TI - Assignment of SH3BP5/Sh3bp5 encoding sab, an SH3 domain-binding protein which preferentially associates with Bruton's tyrosine kinase, to human chromosome 1q43 and mouse chromosome 14B by in situ hybridization. PMID- 10702677 TI - Protein tyrosine phosphatase receptor type C polypeptide (PTPRC) on human chromosome band 1q31-->q32 localizes with marker D1S413(1) on a 610-kb yeast artificial chromosome. PMID- 10702678 TI - Characterization of chromosomal abnormalities in prostate cancer cell lines by spectral karyotyping. AB - Human prostate cancer is characterized by multiple gross chromosome alterations involving several chromosome regions. However, the specific genes involved in the development of prostate tumors are still largely unknown. Here we have studied the chromosome composition of the three established prostate cancer cell lines, LNCaP, PC-3, and DU145, by spectral karyotyping (SKY). SKY analysis showed complex karyotypes for all three cell lines, with 87, 58/113, and 62 chromosomes, respectively. All cell lines were shown to carry structural alterations of chromosomes 1, 2, 4, 6, 10, 15, and 16; however, no recurrent breakpoints were detected. Compared to previously published findings on these cell lines using comparative genomic hybridization, SKY revealed several balanced translocations and pinpointed rearrangement breakpoints. The SKY analysis was validated by fluorescence in situ hybridization using chromosome-specific, as well as locus specific, probes. Identification of chromosome alterations in these cell lines by SKY may prove to be helpful in attempts to clone the genes involved in prostate cancer tumorigenesis. PMID- 10702679 TI - Cytogenetic assignment of 29 functional genes to chicken microchromosomes by FISH. AB - We assigned 29 functional genes to chicken microchromosomes by fluorescence in situ hybridization (FISH). Two linkage groups in the genetic linkage map of the East Lansing breed were identified in this study by localizing the genes AGRN and H2FA to microchromosomes. The frequency of the genes mapped on 30 pairs of microchromosomes, which account for roughly 30% of the whole chicken genome, was about 40% of the 73 genes randomly mapped in our laboratory. This result confirms the important role of microchromosomes for avian genome function and supports the likelihood of a high gene density on avian microchromosomes. PMID- 10702681 TI - Identification and chromosomal localization of murine ORC3, a new member of the mouse origin recognition complex. AB - A new member of the murine origin recognition complex (ORC) related to Saccharomyces cerevisiae ORC3 has been cloned. Transcription of ORC3 is not suppressed in mouse NIH3T3 fibroblasts made quiescent by serum starvation. The transcription level of the ORC3 gene is constantly high in all phases of the cell cycle. Murine ORC3 protein contains a putative nuclear localization signal and a non-basic helix-loop-helix motif. Both motifs are conserved in eukaryotes. A potential dimerization partner of ORC3p in the murine ORC complex is ORC1p which also contains an HLH motif. This HLH motif is also highly conserved in all eukaryotic ORC1 proteins. Comparison of murine ORC3p with other ORC3-related proteins shows high amino acid homology and motif conservation leading to the conclusion that ORC3p is part of the initiation machinery conserved in eukaryotes. The mouse ORC3 gene Orc3 was assigned to mouse chromosome 4A3 by fluorescence in situ hybridization (FISH) analysis. PMID- 10702680 TI - The integrin alpha10 subunit: expression pattern, partial gene structure, and chromosomal localization. AB - Herein we report the cloning of cDNAs and incompletely processed hnRNAs from endothelia and heart that encode the alpha10 subunit forming part of the novel collagen type II-binding integrin alpha10beta1 of chondrocytes. Analysis of hnRNA clones and reported expressed sequence tags revealed the positions of 17 putative intron-exon splice junctions shared with those of the p150,95 (ITGAX) gene. Human alpha10 transcripts of 5.4 and 1.8 kb were not restricted to chondrocytes but, instead, were widely expressed in a panel of 24 tissue types, where the highest expression was found in muscle and heart. The human alpha10 subunit gene (ITGA10) was localized to band q21 of chromosome 1. PMID- 10702682 TI - Assignment of apoptotic protease activating factor-1 gene (APAF1) to human chromosome band 12q23 by fluorescence in situ hybridization. PMID- 10702683 TI - Assignment of the human PLC delta4 gene (PLCD4) to human chromosome band 2q35 by fluorescence in situ hybridization. PMID- 10702684 TI - Assignment of the human translation termination factor 1 (ETF1) to 5q31.1 and of the proximal marker D5S1995 by radiation hybrid mapping. PMID- 10702685 TI - Assignment of the human melanoma cell adhesion molecule gene (MCAM) to chromosome 11 band q23.3 by radiation hybrid mapping. PMID- 10702686 TI - Reassignment of EST w23312 to human chromosome 2q35-->q37 by fluorescence in situ hybridization. PMID- 10702687 TI - Assignment of a member of the ribosomal protein S6 kinase family, RPS6KA5, to human chromosome 14q31-->q32.1 by radiation hybrid mapping. PMID- 10702688 TI - The human HERC3 gene maps to chromosome 4q21 by fluorescence in situ hybridization. PMID- 10702689 TI - Mapping of the CCXCR1, CX3CR1, CCBP2 and CCR9 genes to the CCR cluster within the 3p21.3 region of the human genome. AB - Human CC-chemokine receptor genes are known to be clustered. The detailed structure of this cluster was established by radiation hybrid mapping, and organization of BAC contigs by fluorescence hybridization on combed genomic DNA. A main cluster of six genes (CCR1, CCR3, CCRL2, CCR5, CCR2 and CCXCR1), covered by four BACs, was mapped to the 3p21.3 region of the human genome. Five other genes (CCR9, CCBP2, CX3CR1, CCR8 and CCR4) were found to be spread over a relatively large region between this main cluster and the 3p telomere. PMID- 10702690 TI - Assignment of the gene encoding inwardly rectifying potassium channel, subfamily J, member 3 (Kcnj3) to rat chromosome 3q32 by in situ hybridization and radiation hybrid mapping. PMID- 10702691 TI - Assignment of NEK6, a NIMA-related gene, to human chromosome 9q33. 3-->q34.11 by radiation hybrid mapping. PMID- 10702692 TI - Genetic mapping of the rat Lcn2 gene to chromosome 3. AB - The expression of rat 24p3, encoded by the Lcn2 gene, has been associated with rat mammary carcinomas initiated by the neu oncogene (Stoesz and Gould, 1995). In this study, we assign the Lcn2 gene to rat chromosome band 3q12 by genetic linkage analysis. PMID- 10702693 TI - Assignment of the thyrotropin-releasing hormone gene (TRH) to human chromosome 3q13.3-->q21 by in situ hybridization. PMID- 10702694 TI - Assignment of the inositol polyphosphate 4-phosphatase type I gene (INPP4A) to human chromosome band 2q11.2 by in situ hybridization. PMID- 10702695 TI - Micro- and macrochromosome paints generated by flow cytometry and microdissection: tools for mapping the chicken genome. AB - Despite the chicken being one of the most genetically mapped of all animals, its karyotype remains poorly defined. This is primarily due to microchromosomes that belie assignment by conventional methods. To address this problem, we have developed chromosome-specific paints using flow cytometry and microdissection. For the microchromosomes it was necessary to amplify and label DNA from single microdissected chromosomes. PMID- 10702696 TI - Generation of whole-chromosome painting probes specific to each chicken macrochromosome. AB - The realization of physical and genetic maps of the chicken genome is dependent on progress in cytogenetic knowledge of its karyotype. To help achieve this goal, we constructed amplified representative DNA samples of the chicken chromosomes 1, 2, 3, 4, 5, 6, 7, 8, Z, W and of the terminal heterochromatin part of Zq by chromosome microdissection and DOP-PCR amplification. These chromosome DNA samples, which represent about 75% of the chicken genome, were used to generate whole chromosome painting probes for FISH. The direct application of these chromosome specific probes is dual FISH localization and characterization of panels of chicken interspecific somatic hybrids. We discuss some aspects of the chicken genome and its repeated sequences. PMID- 10702697 TI - Association of genomic imbalances with resistance to therapeutic drugs in human melanoma cell lines. AB - The reason why human malignant melanomas respond poorly to chemotherapy is not known. In an attempt to identify genes responsible for such resistance or sensitivity to therapeutic drugs, we studied the parental human melanoma cell line MeWo, as well as eight drug-resistant sublines of MeWo. These have low and high levels of resistance to four chemotherapeutic drugs with different modes of action: Vindesine, cisplatin, fotemustine and etoposide. Comparative genomic hybridizations with genomic DNA from these cell lines as probes revealed a number of chromosome gains and losses which occurred upon selective pressure during development of the sublines. The MeWo subline with high resistance to the topoisomerase II inhibitor, etoposide, exhibited the highest number of acquired chromosome imbalances. Interestingly, the two lines with high resistance to cisplatin and fotemustine, respectively, shared three additional imbalances, loss of 9p, loss of distal 12p and gain on distal 15q. The importance of these coincident imbalances is discussed. PMID- 10702698 TI - Isolation and characterization of the novel gene, TU3A, in a commonly deleted region on 3p14.3-->p14.2 in renal cell carcinoma. AB - We previously identified a 700-kb region of common allelic loss on 3p14.3-->p14.2 in renal cell carcinoma (RCC). We further analyzed this region and constructed a sequence ready bacterial artificial chromosome (BAC) contig. This region was totally covered by six overlapping BAC clones and was roughly estimated to be 700 kb. Furthermore, we isolated a gene in this region that we termed TU3A. This gene encodes a protein consisting of 144 amino acids. Homology search did not show any significant similarities with known genes or proteins. Northern analysis with normal tissue identified a 3.0-kb transcript that was expressed ubiquitously. Although our mutation search using 37 primary RCCs as well as five RCC cell lines failed to detect any somatic alterations in the TU3A gene, two of five RCC cell lines had totally lost its expression. Considering the fact that we found no genetic alterations in TU3A, it is possible that some epigenetic alteration may have suppressed its expression. PMID- 10702699 TI - Effects of clopidogrel on platelet activation and coagulation of non anticoagulated rat blood under high shear stress. AB - Clopidogrel is a new thienopyridine derivative similar to ticlopidine, which inhibits adenosine diphosphate-induced platelet aggregation. The in vitro effects of clopidogrel on shear-induced platelet activation and coagulation were assessed after oral administration to rats, by subjecting non-anticoagulated blood to haemostatometry. Clopidogrel significantly inhibited shear-induced platelet activation and coagulation 2 h after administration at doses of 7.5 and 15 mg/kg. Both ticlopidine (200 mg/kg) and aspirin (200 mg/kg) inhibited shear-induced platelet activation, but not coagulation. The peak inhibition of plaetelet activation by clopidogrel occurred 2 h after oral administration, but significant inhibition persisted even after 24 h. These results suggest that clopidogrel could be a more potent antithrombotic agent than ticlopidine or aspirin, and also that ADP plays an important role in shear-induced platelet activation. PMID- 10702700 TI - Usefulness of screening for congenital or acquired hemostatic abnormalities in women with previous complicated pregnancies. AB - Activated protein C resistance (APCR) is a common cause of familial thrombophilia and venous thrombosis. The aim of the study was to investigate the prevalence of APCR associated with factor V Leiden mutation and its relevance in comparison to other risk factors for thromboembolic disorders in women with a history of previous complicated pregnancies (history of fetal loss in the second and third trimester n = 34, preeclampsia n = 46). The frequency of APCR was significantly higher in women with a history of fetal loss and preeclampsia (23.5 and 26.1%, respectively) compared with a control group (3.8%). The prevalence of antithrombin, protein C and protein S deficiencies and the presence of antiphospholipid antibodies were also investigated: the prevalence of at least one disorder was 41.2% in the group with previous fetal loss, 37.0% in the group with previous preeclampsia and 7.5% in the control group. PMID- 10702701 TI - Biochemical characterization of a thrombin inhibitor from the bloodsucking bug Dipetalogaster maximus. AB - From the bloodsucking bug Dipetalogaster maximus, a protein with anticoagulant activity was isolated and biochemically characterized. The isolated protein, named dipetalogastin, possesses an average molecular mass of 11.8 kD. Its N terminal sequence shows homology to rhodniin, a thrombin inhibitor isolated from the bug Rhodnius prolixus. The in vitro anticoagulant activity of dipetalogastin occurs via the inhibition of thrombin. The anticoagulant and thrombin inhibitory potency of dipetalogastin is comparable to that of recombinant hirudin. Its specific thrombin inhibitory activity is 9,300 antithrombin units/mg protein. Dipetalogastin forms only 1:1 molar complexes with thrombin. It is a tight binding inhibitor of thrombin possessing a dissociation constant of 125 fM. It does not inhibit factor Xa or alpha-chymotrypsin and only weakly inhibits trypsin. PMID- 10702702 TI - Genetic risk factors in acute coronary disease. AB - OBJECTIVE: We investigate whether each of the following: HPA-1, Factor V Leiden, prothrombin gene variant and the methylene tetrahydrofolate reductase gene (MTHFR) mutation, are risk factors for acute coronary disease in Portuguese patients. MATERIAL AND METHODS: 100 blood donors and 52 patients with an established diagnosis of myocardial infarction or unstable angina were evaluated for genetic risk factors, by determining HPA-1 genotype, Factor V Leiden, Prothrombin 20210 variant and MTHFR mutation. RESULTS: We found a prevalence of 2.0% for Factor V Leiden, 5.0% for the Prothrombin 20210 variant and 66% for the MTHFR mutation in blood donors. These values are similar to those found in the patients (1.9, 3.8 and 58%, respectively). We found that 28/100 controls had the PI(A2) polymorphism, a frequency statistically different from that in the patients (23/52). This difference was even more pronounced in patients less than 60 years old (27/96 vs. 13/24). CONCLUSION: Factor V Leiden, Prothrombin 20210 variant and MTHFR mutation do not seem to represent risk factors for acute coronary disease. However, the PI(A2) polymorphism could have a role in the pathogenesis of this disease. The presence of multiple genetic factors, more than single ones, could influence the development and outcome of myocardial infarction and unstable angina. Larger studies are needed in order to have a better insight into the pathophysiological mechanisms of this disease, along with its prevention and the development of new treatments. PMID- 10702703 TI - Hemorheological profile in chronic venous insufficiency after surgery. AB - In recent years, there have been rheological abnormalities reported in chronic venous insufficiency (CVI), mainly an increase of erythrocyte aggregability (EA), which probably take part in the pathophysiology of the disease. The aim of this study was to analyze the hemorheological profile after stripping in 45 patients suffering from CVI. Follow-up included laboratory tests on the 7th, 60th and 180th day after surgery. EA was assessed with a photometric aggregometer (MA1, Myrenne) in stasis and low shear (3 s(-1)). The results show an increase of EA on the 7th day after surgery (p<0.001). Two and 6 months later, EA values returned to those found prior to surgery. The plasma fibrinogen level changes in a way parallel to EA. The association between rheological disturbances and thrombogenesis is well known, so the hyperaggregability found supports the antithrombotic prophylaxis in the early postsurgical period. On the other hand, the hemorheological abnormalities persist after stripping, so postsurgical treatment to inhibit EA may be beneficial. PMID- 10702704 TI - Determination of the prevalence of fibrinogen Banks peninsula mutation (gamma280Tyr-->Cys) by PCR. AB - A mutation of Tyr-->Cys at position 280 of the gamma chain of fibrinogen was recently determined to be the cause for fibrinogen Banks peninsula. A novel polymerase chain reaction/restriction digestion assay for this mutation was developed and used for the screening of 300 apparently healthy European blood donors. None of these subjects carried the mutation. PMID- 10702705 TI - Acquired Bernard-Soulier syndrome: a case with necrotizing vasculitis and thrombosis. AB - We describe a patient with positive antinuclear antibodies, polyclonal gammopathy and high level of circulating immunocomplexes, resulting in vascular purpura. In addition, the patient had a slightly prolonged bleeding time and an isolated defect of ristocetin-induced platelet aggregation (RIPA) in platelet-rich plasma (PRP). The patient's plasma also inhibited RIPA in normal PRP and in normal platelet suspension. The activity and multimeric structure of plasmatic von Willebrand factor showed no alteration. We could demonstrate an autoantibody against platelet membrane glycoprotein (GP) Ib, using an ELISA-type assay. These data suggest an acquired Bernard-Soulier syndrome. We suggest that the patient had an immunocomplex-mediated leukocytoclastic vasculitis accompanied by production of antinuclear autoantibodies as well as the presence of an autoantibody against GPIb. The titer of the anti-GPIb antibody, however, was too low to induce significant platelet-type bleeding tendency, only laboratory alterations were found. Moreover, in a later stage of her disease, she developed a severe necrotizing vasculitis which was followed by a deep venous thrombosis. PMID- 10702706 TI - Protein-bound heparin/heparan sulfates in human adult and umbilical cord plasma. AB - We used thrombin times and a competitive radiometric assay to identify, quantitate and characterize endogenous heparin-like molecules in umbilical cord (n = 58) and normal adult (n = 25) plasma. Thrombin times for cord plasma (29.6+/ 3.6 s) were significantly longer (p< or = 0.0005) than those for adult plasma (18. 9+/-2.3 s), suggesting increased endogenous heparins. A radiometric assay based on the displacement of (125)I-heparin from protamine-Sepharose revealed that protease-digested plasma contained heparin/heparan sulfate, and plasma that was not digested with protease appeared not to contain heparin/heparan sulfate. More heparin/heparan sulfate was identified in cord than in adult plasma (p< or =0.05), but heparinase digestion produced significantly (p< or =0.001) reduced concentrations of heparin/heparan sulfate in only 39% of the samples. The lack of heparinase sensitivity in 61% of the protease-digested samples apparently was due to low molecular weight (LMW) heparins, for control heparin fragments of 5 kD that did not extend thrombin times were also less affected by heparinase, but the same LMW heparins were detected by radiometric assay. Despite normal thrombin times in all samples, the amounts of endogenous heparin/heparan sulfate identified in protease-digested samples by radiometric assay were of sufficient concentrations to produce inordinately prolonged thrombin times when compared with the same concentrations of unfractionated heparin. Collectively, these findings suggest the presence of a plasma reservoir of endogenous heparin/heparan sulfates in normal cord and adult plasma. These endogenous heparin/heparan sulfates are bound to plasma proteins, and an as yet undetermined proportion of these bound heparin/heparans are most likely LMW molecules. PMID- 10702707 TI - Time-related effects of cytomegalovirus infection on the development of chronic renal allograft rejection in a rat model. AB - Cytomegalovirus (CMV) infection is a risk factor for chronic allograft rejection. The histological findings of chronic renal allograft rejection include inflammation, vascular intimal thickening, glomerulosclerosis, tubular atrophy and fibrosis. We have developed a rat model of renal transplantation in which transplants, after an early inflammatory episode, end up with chronic rejection within 60 days. During the early phase of the process in this model, CMV increased and prolonged the inflammatory response, the expression of adhesion molecules, intercellular adhesion molecule-1 and vascular cell adhesion molecule 1 and their ligands, lymphocyte function antigen-1 and very late antigen-4 in the graft. Simultaneously, the production of various growth factors, such as transforming growth factor beta, platelet-derived growth factor and connective tissue growth factor was upregulated, which induce smooth muscle cell proliferation in the vascular wall and collagen synthesis by fibroblasts. Chronic rejection developed within 20 days in CMV-infected grafts. In summary, CMV infection accelerated and enhanced the early immune response, the induction of growth factors and collagen synthesis, and the development of chronic rejection in renal allografts. PMID- 10702708 TI - Overcoming the problem of cytomegalovirus infection after organ transplantation: calling for Heracles? AB - Although diagnosis of CMV infections and treatment of CMV disease with effetive antiviral drugs have become much easier, the persistent problem of CMV infection after solid-organ transplantation still requires solid knowledge of the pathophysiology of its clinical manifestations in order to minimize the impact of CMV infections in the future. The complex symptomatology of CMV infection after solid-organ transplantation is reviewed as well as some of the new theories attempting to explain the myriad of symptoms seen after transplantation. PMID- 10702710 TI - Human cytomegalovirus escape from immune detection. AB - Human cytomegalovirus (CMV) has devised numerous means of escaping immune surveillance. The CMV genome encodes at least 4 genes involved in downregulating surface expression of HLA class I molecules. In addition, it sequesters CC chemokines, induces Fc receptors, interferes with induction of HLA class II antigens, and can inhibit natural killer cell activity. CMV can efficiently block the presentation of immediate early antigens, the first viral proteins to be produced. Together, these mechanisms probably contribute to the ability of CMV to persist in its host and may play a role in the immunopathology of CMV disease. PMID- 10702709 TI - Clinical significance of cytomegalovirus-specific T helper responses and cytokine production in lung transplant recipients. AB - Cytomegalovirus (CMV) disease continues to be a major problem for lung transplant recipients. In CMV-seropositive individuals, we detected two types of CMV specific responses: a self-restricted response stimulated by soluble CMV antigen (sCMV-Ag) and a non-self-restricted response induced by CMV-infected cells (cCMV Ag). Lung transplant recipients who develop the CMV-specific self-restricted T helper response have a low risk of recurrent CMV disease. In contrast, during CMV disease, lung transplant recipients exhibit only the non-self-restricted T helper responses. We characterized the T cell activation and the kinetics of cytokine production of sorted CD4+ and CD8+ T cells from PBLs of CMV seropositive donors. The two types of CMV antigens induced cytokine production in both T cell subsets. We also performed competitive RT-PCR for Granzyme B (GB) in BAL cells of lung transplant recipients prior to, during and following CMV disease. CMV disease was associated with increase in GB gene expression when was accompanied by acute cellular rejection while it remained low in patients with CMV disease that did not have a complicated course. In summary, CMV-activated T cells within the allograft may produce inflammatory cytokines and effector molecules that may promote allograft rejection. PMID- 10702711 TI - Human cytomegalovirus reactivation in bone-marrow-derived granulocyte/monocyte progenitor cells and mature monocytes. AB - Monocyte/granulocyte progenitor cells of the bone marrow are a major site of human cytomegalovirus (HCMV) latency. The mechanisms of establishment and maintenance of HCMV latency are still unknown. Reactivation of the latent virus in bone-marrow-derived progenitor cells has been demonstrated in vitro and suggested to occur also in vivo. Clinical studies have shown that reactivation is a rather frequent event not only in immunosuppressed but also in nonimmunosuppressed patients and in healthy blood donors. At least three independent mechanisms of virus reactivation are discussed: systemic inflammation connected with strong tumor necrosis factor alpha release; application of cAMP elevating drugs, and highly stressful events associated with increased plasma catecholamine levels. PMID- 10702712 TI - Human cytomegalovirus latency and reactivation - a delicate balance between the virus and its host's immune system. AB - Human cytomegalovirus (HCMV) is a ubiquitous herpesvirus that still causes severe morbidity and mortality in immunocompromised individuals. During its evolution, the virus has developed sophisticated methods to evade immune recognition and to establish life-long persistence in its host. Today, we know that the virus establishes latency in myeloid lineage cells and that the virus is dependent on immune activation mechanisms to reactivate it from latency to produce a new viral progeny. During this process, a number of viral proteins are produced that interfere with different immune recognition pathways. The current knowledge of the delicate balance between the virus' continuous existence and its host's immune system will be summarized in this chapter. PMID- 10702713 TI - Measurement of anti-human cytomegalovirus T cell reactivity in transplant recipients and its potential clinical use: a mini-review. AB - By allowing direct determination of the frequencies of antigen-specific memory T cells in peripheral blood, novel techniques based on flow cytometry provide new diagnostic opportunities in various clinical settings, including organ transplantation. While the importance of the T cell compartment for the anti human cytomegalovirus (HCMV) immune response is undisputed, efficient monitoring of this response was previously impossible because the conventional methods for measuring CD4+ and CD8+ T cell responses are too time-consuming and cost intensive. We analyzed how the rapid induction of anti-HCMV CD4+ and CD8+ memory T cells by HCMV viral lysate or HCMV-derived peptides, respectively, followed by a flow-cytometric detection step, may be used to monitor HCMV-specific CD4+ and CD8+ memory T cells in solid-organ recipients. We also discuss a number of preconditions for integrating such testing into the clinical routine. PMID- 10702714 TI - Viral inhibition of interferon signal transduction. AB - The type I and II interferons (IFNs) are potent stimulators of antigen processing and presentation and are essential in antiviral immunity. IFNs upregulate the transcription of major histocompatibility complex (MHC) class I and II molecules, associated antigen-processing proteins, and induce the production of direct antiviral effector molecules such as 2',5'-oligoadenylate synthetase, double stranded-RNA-dependent protein kinase and Mx proteins. It is increasingly evident that viruses have evolved mechanisms to globally inhibit the actions of IFNs through disruption of their signal transduction pathways. Herein, we review the ability and novel mechanisms of several diverse viruses to inhibit IFN-induced JAK/STAT signal transduction. PMID- 10702715 TI - Murine cytomegalovirus homologues of cellular immunomodulatory genes. AB - The study of 'molecular mimicry' or 'genetic piracy', with respect to the utilisation of cellular genes captured and modified during the course of virus evolution, has been an area of increasing research with the expansion in virus genome sequencing. Examples of cellular immunomodulatory genes which have been captured from hosts have been identified in a number of viruses. This review concentrates upon studies of murine cytomegalovirus (MCMV), investigating the functions of viral genes homologous to G protein-coupled receptors, MHC class I and chemokines. The study of recombinant MCMV engineered with specific disruptions of these genes has revealed their significance during virus replication and dissemination within the host. In the case of the latter two classes of genes, evidence suggests they interfere with cellular immune responses, although the detailed mechanisms underlying this interference have yet to be delineated. PMID- 10702716 TI - Molecular mimicry by cytomegaloviruses. Function of cytomegalovirus-encoded homologues of G protein-coupled receptors, MHC class I heavy chains and chemokines. AB - Cytomegaloviruses (CMVs) are well known for their high prevalence rate within host populations as well as their ability to induce lifelong infections. To maintain a persistent and stable relationship with their host, CMVs have evolved various molecular mechanisms to both control host cell metabolism and evade immune surveillance. Among the viral gene products that are likely to be involved in these processes are homologues of cellular G protein-coupled receptors, MHC class I molecules and chemokines. The viral genes encoding these homologues have probably been pirated by the viruses during a long pathogen/host coevolution. In this report, we will discuss the possible functions of these homologues in the pathogenesis of CMV infections. PMID- 10702717 TI - Cytomegalovirus-induced transendothelial cell migration. a closer look at intercellular communication mechanisms. AB - A variety of cells such as leukocytes and tumor cells may adhere to endothelial cells and subsequently transmigrate into the solid tissue by involving specific intercellular molecular pathways. One important prerequisite for transendothelial migration is the loosening of endothelial cell-to-cell contact sites, which can be triggered by extravasating cells. Cytomegalovirus (CMV) has obviously evolved the ability not only to influence host cells floating in the blood stream to adhere to endothelial cells, but also to induce the formation of intercellular gaps within the endothelium, resulting in transendothelial migration. These features allow the virus to disseminate and evade the immune system. In coculture experiments with human endothelial monolayers and human CMV (HCMV)-infected neuroblastoma cells or leukocytes, changes in the integrity of the monolayer were observed and further analyzed on the molecular level. For example, HCMV may activate the integrin beta1alpha5 (VLA-5) that triggers adhesion to endothelial cells with subsequent focal disruption of endothelial cell-to-cell connections. It is hypothesized that a Ca(2+)-independent pathway following VLA-5 binding disconnects the cadherin-catenin-actin complex within the endothelial cells. The loss of cadherin function causes the loss of contact to the neighboring endothelial cells and thus could represent an important mechanism in HCMV-induced cellular transendothelial migration and disruption of the endothelial integrity. PMID- 10702719 TI - Human cytomegalovirus infection of immature dendritic cells and macrophages. AB - A central aspect of human cytomegalovirus (HCMV) pathogenesis is the interaction of the virus with different antigen-presenting cell (APC) types of the host. In principle, a number of various cell types have the potential of antigen presentation when MHC II expression is induced by appropriate stimuli. The most potent antigen presenters are monocytes/macrophages and dendritic cells (DCs), therefore called professional APCs. Interestingly, these cells seem to be targets of productive HCMV infection. The susceptibility of the monocyte/macrophage system has been analyzed intensively during the past decade. Investigation of the role of DCs during HCMV infection, however, has begun only recently. PMID- 10702718 TI - Altered expression of extracellular matrix in human-cytomegalovirus-infected cells and a human artery organ culture model to study its biological relevance. AB - The influence of human cytomegalovirus (HCMV) on the transcription of 11 selected, representative extracellular matrix genes was investigated in cell culture. Northern blot hybridization indicated the downregulation of all mRNAs investigated. Based on our results and the known repression of other extracellular matrix transcripts and the beta-actin transcription during HCMV infection, we suggest that one molecular mechanism contributing to the cytopathic effect may be the transcriptional downregulation of genes encoding proteins involved in cell structure and intercellular connection. To further study the biological relevance of this and other pathogenetic mechanisms, we established a human renal artery organ culture system and characterized this new infection model for HCMV. Our model is a new suitable system for the investigation of molecular as well as functional consequences of HCMV infection in a more physiological microenvironment. PMID- 10702720 TI - Diagnostic value of nucleic-acid-sequence- based amplification for the detection of cytomegalovirus infection in renal and liver transplant recipients. AB - To evaluate the diagnostic value of nucleic-acid-sequence-based amplification (NASBA) for the detection of cytomegalovirus (CMV) infection in transplant recipients, we compared immediate early 1 (IE1) and late pp67 mRNA detection by NASBA with the antigenemia assay, PCR and viral culture in 72 renal transplant (RTx) recipients and with antigenemia and serology in 25 liver transplant (LTx) recipients. Antigenemia, viral culture and pp67 NASBA were almost equivalent for the detection of CMV in RTx recipients. In LTx recipients, antigenemia detected more positive samples and more positive recipients compared to pp67 NASBA. In RTx recipients, PCR detected more positive samples and positive recipients compared to pp67 NASBA, antigenemia and viral culture. Also the first day of detection was slightly earlier for PCR. However, IE1 NASBA was the most sensitive test and detected 96% of all positive samples and positive transplant recipients. In addition, IE1 NASBA preceded PCR and all other positive results. This makes IE1 NASBA a very attractive screening test for the early detection of CMV infection. PMID- 10702721 TI - Towards standardization of the human cytomegalovirus antigenemia assay. AB - The Human Cytomegalovirus antigenemia (HCMV-Agemia) test has been accepted worldwide as a clinical tool in the diagnosis and management of HCMV-associated syndromes in immunocompromised patients. The many modifications proposed since the first description by our laboratory make standardisation of the HCMV-Agemia assay necessary to enable multicenter clinical trials. We report the initial work for standardization of the HCMV-Agemia assay. A standard protocol is proposed, the optimal distribution conditions are investigated and the results of the shipment of positive and negative test slides as well as of two sets of coded internal standard slides are discussed. The main conclusions are that standard slides can be distributed at room temperature and that the results of participating laboratories with the coded internal standard slides were strikingly similar in spite of differences in HCMV-Agemia protocols used by participating laboratories. PMID- 10702722 TI - New advances in the diagnosis of congenital cytomegalovirus infection. AB - With the advances in anticytomegalovirus (anti-CMV) serology, the new recombinant IgM tests seem likely to become the screening tests for pregnant women whose prepregnancy serological status for CMV is unknown. When a woman is found to be IgM-positive, further diagnostic evaluation focused on determining whether this is due to a primary infection should be carried out. Maternal primary infections that were difficult to determine until a few years ago unless documented by seroconversion can now be readily diagnosed from the presence of low-avidity anti CMV antibody which persists for approximately 20 weeks after primary infection. In primarily infected mothers prenatal diagnosis can be performed between 21 and 23 weeks of gestation, and the amniotic fluid (AF) represents the pathological material of choice to determine intrauterine virus transmission. In AF, the virus can be detected by culture and/or PCR. Both procedures differentiate uninfected from infected fetuses, but cannot predict fetal outcome. The determination of the viral load in AF carried out by quantitative PCR is more promising and could represent an important starting point for preemptive fetal therapy. PMID- 10702723 TI - Significance of qualitative polymerase chain reaction combined with quantitation of viral load in the diagnosis and follow-up of cytomegalovirus infection after solid-organ transplantation. AB - Quantitative PCR was evaluated in the monitoring of patients with ongoing posttransplantation cytomegalovirus (CMV) infection and antiviral therapy, compared to leukoDNAemia and serology. From January 1998 until May 1999, 61 patients were followed up weekly during 3 months after transplantation by a qualitative PCR. The quantitative PCR was performed on plasma samples from 21 selected patients, of whom 12 had a primary infection and 9 a reactivation or reinfection. Analysis of the viral load differences showed that the viral loads in patients with a primary infection were significantly higher than viral loads in patients with a reactivation (p < 0.01). Based on the results of our study, we can state that qualitative PCR is a good marker for initiating preemptive therapy. In addition, viral quantitation is clinically useful for accurate diagnosis of established CMV disease, and monitoring of antiviral therapy. PMID- 10702724 TI - Viral dynamics during active cytomegalovirus infection and pathology. AB - The central role that cytomegalovirus (CMV) load plays in its pathogenesis is being unravelled. In AIDS patients with active CMV replication, many months prior to the development of CMV disease, elevated CMV load in the blood and urine are significantly associated with an increased risk of disease progression. In addition, elevated load in blood is associated with an increased risk of death. Intervention with ganciclovir acts to rapidly inhibit CMV replication in vivo and has allowed estimates of the clearance/replication rate of CMV to be performed. These data indicate that CMV replicates dynamically in the human host with a doubling time of approximately 1 day. This knowledge has been used to determine the relative contribution of initial viral load and rate of change of viral load as predictors of CMV disease in organ transplant recipients. The data show that both these parameters have prognostic value in multivariate models and should allow the development of novel patient management strategies. PMID- 10702725 TI - Inhibition of cytomegalovirus in vitro and in vivo by the experimental immunosuppressive agent leflunomide. AB - Despite progress in antiviral chemotherapy, cytomegalovirus (CMV) remains a major cause of morbidity and mortality among pharmacologically immunosuppressed transplant recipients, frequently engaging the clinician in a struggle to balance graft preservation with control of CMV disease. Leflunomide, an inhibitor of protein kinase activity and pyrimidine synthesis, is an experimental immunosuppressive agent effective against acute and chronic rejection in animal models. Herein we summarize our recent studies demonstrating that leflunomide inhibits the production of multiple clinical CMV isolates (including multi-drug resistant virus) in both human fibroblasts and endothelial cells. In contrast to all other anti-CMV drugs currently in use, leflunomide does not inhibit viral DNA synthesis, but rather appears to interfere with virion assembly. Finally, preliminary studies in a rat model suggest that this agent reduces viral load in vivo. These findings imply that leflunomide, an effective immunosuppressive agent, shows potential to concurrently attenuate a major complication of immunosuppression, CMV disease, by a novel mechanism of antiviral activity. PMID- 10702726 TI - Proinflammatory potential of cytomegalovirus infection. specific inhibition of cytomegalovirus immediate-early expression in combination with antioxidants as a novel treatment strategy? AB - We observed the effects of antiviral therapy on CMV and/or oxidative-stress induced stimulation of proinflammatory molecules including interleukin-8 (IL-8), melanoma growth stimulatory activity-alpha (GRO-alpha) and intercellular adhesion molecule 1 (ICAM-1) using human foreskin fibroblasts. Ganciclovir, foscarnet or cidofovir completely suppressed virus replication, as demonstrated by CMV late (L) antigen production. These drugs did not influence CMV immediate-early (IE) antigen expression and had no effects on CMV-induced cellular changes in IL-8, GRO-alpha and ICAM-1 levels. Phosphorothioate oligonucleotide (ISIS 2922) suppressed both CMV IE and L antigen by 99%. ISIS 2922 completely suppressed CMV induced upregulation of both chemokines and ICAM-1. Induction of oxidative stress by H(2)O(2) upregulated IL-8 expression. Oxidative stress and CMV infection showed synergistic effects on IL-8 expression. ISIS 2922 only partially inhibited the upregulation of IL-8 in infected cells treated with H(2)O(2), whereas cotreatment with ISIS 2922 and antioxidants inhibited the upregulation almost completely. The results showed that inhibition of CMV IE expression alone or in combination with antioxidants is promising for the treatment of CMV disease. PMID- 10702727 TI - Skull base growth in craniosynostosis. AB - Although considerable scientific work has been published on the role of the skull base in craniosynostosis, the changes with age throughout childhood have not been fully outlined. The realisation that little attention has been paid to the posterior skull in craniosynostosis, resulted in renewed interest in skull base growth. The availability of computer-based image analysis provides a new accurate method of study in three dimensions. Using three-dimensional visualisation techniques, 34 points of the skull base were identified on CT scans of 50 children with craniosynostosis of various types, aged from 1 month to 5 years. Several distances and angles between the various landmarks were measured in an attempt to quantify the growth of skull fossae with age. Comparisons were made with normal controls. In children with craniosynostosis, the anterior fossa was overdeveloped in the males, whereas in the females remained underdeveloped throughout the first 2 years of life. The body of the sphenoid showed moderate underdevelopment in the first 2 years in both sexes, the effect being more prominent in the males. The middle fossae showed overdevelopment in both sexes in the first 2 years of life. The posterior fossa was underdeveloped in both sexes in the first 2 years of life, the effect being more prominent in the females. Craniosynostosis seems to affect both sexes to a similar degree, but there are regional differences in the growth pattern. Better understanding of the normal growth pattern of the skull base and the effect of craniosynostosis upon it may assist our approach to surgical treatment and in particular the role of anterior and posterior skull expansive surgery. PMID- 10702728 TI - Delayed repair of open depressed skull fracture. AB - INTRODUCTION: Elevation and repair of an open depressed skull fracture is often thought of as an emergency procedure. Common indications for emergent elevation of a depressed skull fracture have been dural tear, seizure, gross contamination or mass effect from bone or a sizable underlying intracerebral hematoma. As treatment of head injury moves towards management of cerebral perfusion pressure (CPP) rather than intracranial pressure (ICP), we sought a way to maximize CPP in the initial treatment of head-injured patients with depressed skull fractures that would eventually require surgery by delaying surgery, when possible, until after the initial period of elevated ICP. METHODS: Over a 12-month period, 7 patients (all male, ages 1-15 years) were admitted to our institution with the diagnosis of open depressed skull fracture without significant mass effect requiring urgent decompression. All had significant head trauma with altered mental status and a Glasgow Coma Score of 3-12. Patients were treated with antibiotic prophylaxis (nafcillin, ceftriaxone, metronidazole), seizure prophylaxis (phenytoin) and underwent CPP management in an intensive care unit setting as indicated by intracranial pressure monitoring or clinical assessment. Length of medical management of CPP ranged from 4 to 12 days. Upon stabilization of CPP, patients were operated for repair of their dural, bone and scalp injuries. RESULTS: All 7 patients treated in the above manner suffered no ill effects from their delayed surgery: there was no meningitis, no late seizures, and no cerebrospinal fluid leak. Complications attributable to delay were not present at follow-up ranging from 12 to 24 months. CONCLUSIONS: We have delayed surgery for repair of open depressed skull fractures in order to maximize medical management of CPP in the setting of acute trauma. Among other considerations, the risk of intraoperative hypotension occurring at a time of acutely raised ICP was avoided by this delay. We conclude that there is a role, in this specifically defined subset of head trauma patients, for delayed surgical repair of open depressed skull fractures. PMID- 10702729 TI - Melatonin as a free radical scavenger in experimental head trauma. AB - Head trauma causes two kinds of injury in the neural tissue. One is the primary injury which occurs at the time of impact. The other one is a secondary injury and is a progressive process. Free radicals are produced during oxidative reactions formed after trauma. They have been thought to be responsible in the mechanism of the secondary injury. Some studies have been conducted to demonstrate the role of free oxygen radicals in neuronal injury. The alterations in the free radical level during the early posttraumatic period and the effect of a free radical scavenger on these alterations have not been studied as a whole. We aimed to demonstrate the free oxygen radical level changes in the early posttraumatic period and the effect of melatonin, which is a potent free radical scavenger, on the early posttraumatic free radical level. A two-staged experimental head trauma study was designed. In stage one, posttraumatic free radical level changes were determined. In the second stage, the effect of melatonin on the free radical level changes in the posttraumatic period was studied. Two main groups of rats each divided into four subgroups were studied. Rats in one of the main groups underwent severe head trauma, and malondealdehyde (MDA) levels were measured in the contused cerebral tissue at different time points. Rats in the other main group also underwent the same type of trauma, and melatonin was injected intraperitoneally at different time points after trauma. The MDA level alteration in the tissue was determined after the injection of melatonin. The MDA level increased rapidly in the early posttraumatic period. But in time, it decreased in the groups with only trauma. In the melatonin-treated group, the MDA level decreased after the injection of melatonin, when injected in the early posttraumatic period, compared to the control and trauma groups. However, melatonin increased MDA to a higher level than in the groups with only trauma and the control group when injected later than 2 h after trauma. The MDA level increases in the very early posttraumatic period of cerebral trauma and decreases in time. Melatonin, which is the most potent endogenous free radical scavenger, when injected intraperitoneally to the cerebral traumatized rats in the very early posttraumatic period, causes a significant decrease in the MDA level. But, melatonin, when injected more than 2 h after trauma, increases the MDA level in experimental cerebral trauma in rats. PMID- 10702730 TI - Current patterns of inflicted head injury in children. AB - The purpose of this study was to examine the current patterns of head trauma associated with child abuse. We reviewed the records of all patients admitted to our medical center between 1995 and 1997 with a primary diagnosis of head trauma, and analyzed the clinical presentation, mechanism of injury, socioeconomic status and outcome for these patients. Head trauma was deliberately inflicted in 38/405 children (9%). There were 25 boys and 13 girls, with a median age of 5.5 months. Two thirds of the families lived in the inner city. Of the 99 children under the age of 2 years admitted for head trauma, the injury was inflicted in 32 (32%). Acute subdural hematoma was present in 22/32 (69%) of children with inflicted trauma, but in only 5/68 (7%) with accidental trauma. Retinal hemorrhages were present in 17/32 (53%) abused children, but in no cases of accidental trauma (0/68). Deliberately inflicted injury is a frequent cause of serious head trauma in young children. Head injury is a major cause of morbidity and mortality in the abused child. Child abuse cases correlated strongly with low socioeconomic status. Nonaccidental trauma must be considered strongly in children under 2 years of age who present with acute subdural hematoma in the absence of a history of a motor vehicle accident. PMID- 10702731 TI - Primary Ewing's sarcoma of the skull in children. Utility of molecular diagnostics, surgery and adjuvant therapies. AB - Ewing's sarcoma (ES) of the skull is rare. Herein, we present 2 cases of ES that involved the cranium in young children. In one case, the lesion originated in the petrous temporal bone; in the other, the frontal bone. Both children were acutely compromised neurologically by signs and symptoms of raised intracranial pressure. In both cases, radiographs revealed massive tumors affecting the skull. Neurosurgical resection of the tumor was undertaken in both instances, and the diagnosis of ES was confirmed by immunohistochemistry, cytogenetic analysis (translocation 11;22), spectral karyotyping and RT-PCR (demonstration of a EWS/FLI1 fusion transcript). Following aggressive surgical resection, both children received intensive chemotherapy. No child has received radiation therapy. One child is alive and well 8 years after diagnosis without any evidence of residual disease. The other is currently undergoing chemotherapy for her tumor. The principles involved in the management of children with cranial-based ES are discussed. These 2 cases serve to illustrate the fact that even children with massive ES tumors of the cranium may be salvaged with aggressive combination therapy. PMID- 10702732 TI - The endocrine spectrum of arachnoid cysts in childhood. AB - BACKGROUND: On clinical grounds, arachnoid cysts are usually associated with neurological dysfunction. There is little information concerning their involvement in endocrinological disorders. PATIENTS: The experience in 6 children (birth to 12 years) with hypothalamic-pituitary disturbances secondary to the presence of intracranial arachnoid cysts is reported and the literature is reviewed. RESULTS: Three of our children were diagnosed with isolated hormone abnormalities (2 children with precocious puberty and 1 child with growth hormone, GH deficiency). One child presented the unusual combination of GH deficiency and precocious puberty. The remaining 2 children developed panhypopituitarism associated with diabetes insipidus. CONCLUSION: Arachnoid cysts may cause a wide spectrum of endocrinological disorders. Periodical and complete follow-up of every patient is recommended. PMID- 10702733 TI - Infantile cerebral aneurysms with visual pathway compression. AB - Intracranial aneurysms are rare in infancy. The commonest presentation is intracranial hemorrhage, but signs of mass effect are more frequent than in adults. We report 2 infants with cerebral aneurysms, one presenting with macrocephaly and another with strabismus. Both had visual loss and optic disc pallor; MRI revealed a suprasellar mass and anterior visual pathway compression. In both cases, the preoperative diagnosis was craniopharyngioma. It is essential to recognize that, although exceedingly uncommon, cerebral aneurysms do occur in infants and have features that differ from those in adults. PMID- 10702734 TI - Approach via the floor of the fourth ventricle for hydatid cyst of the pons. AB - A very rare case of a pontine hydatid cyst is reported. It was diagnosed preoperatively on the basis of magnetic resonance imaging findings. The patient was operated on in the sitting position. Total excision of the cyst using the Dowling technique and gravity effect was performed through the floor of the fourth ventricle. The patient was discharged without extra neurological sequel. The significance of an accurate preoperative diagnosis, surgical approaches and technique in the management of this pathology is discussed. PMID- 10702735 TI - Posterior fossa craniotomy: an alternative to craniectomy. PMID- 10702736 TI - Value in microsurgery of the ultrapulse carbon dioxide laser for rapid skin deepithelialization. AB - Although laser technology continues to evolve, its role in microsurgery is limited to minor adjunctive applications including skin deepithelialization. In the past, continuous-wave carbon dioxide (CO(2)) lasers were the best for performing skin deepithelialization. The newer ultrapulse CO(2) lasers also show promise in this regard. They have become readily available and are very popular for aesthetic skin resurfacing. Their use for skin deepithelialization is an essential part of microsurgical procedures for free flap contouring, flap onlay at the recipient site, and revision of skin grafted free flap donor sites. Complications are essentially nonexistent and competency in using this modality is straightforward for any microsurgeon. PMID- 10702737 TI - Microvascular repair following a modified crush-avulsion injury in a rat model: effect of recombinant human tissue-type plasminogen activator on the patency rate. AB - The failure rate of replantations following a crush-avulsion type injury is high. This study has been designed to reproduce an effective standardized crush avulsion injury model to the femoral artery of the rat and evaluate the antithrombotic efficacy of systemic intravenous administration of recombinant human tissue-type plasminogen activator (rt-PA). The crush-avulsion injury was reproduced by using a bulldog clamp and two hemostats and followed by microvascular repair. The animals were divided into three groups of 20 rats each and received either normal saline, heparin 100 U/kg body weight, or rt-PA 3.5 mg/kg body weight intravenously. Patency tests were performed 20 min and 48 h after blood flow reestablishment. Results showed that this experimental crush avulsion injury model ensures low patency in the control group, whereas systemic rt-PA administration improves the patency rate statistically significantly compared to control and heparin groups at both 20 min and 48 h postrevascularization. PMID- 10702738 TI - Xenotransplantation model for vascularized musculoskeletal tissues in rodents. AB - The purpose of this study was to establish a model and to define the mechanism of rejection for the transplantation of vascularized musculoskeletal xenografts between C57BL/6j (B6) mice and Lewis rats. This was accomplished by using conventional skin xenografts to determine immunologic baseline data between these species and by performing musculoskeletal grafts from the B6 mice transplanted into Lewis rats. After the transplant, the xenografts were examined histologically and the recipients were assessed for immune reaction using in vitro assays to measure both cell-mediated and humoral responses. The results obtained from the skin xenografts showed activation of both cellular and humoral immunologic responses. All musculoskeletal xenografts were rejected between 3 and 4 postoperative days. Histologically, the grafts showed extensive vascular injury manifested by thrombosis and hemorrhage, suggesting an early humoral response. Anti-donor antibody production was detected in the recipient's sera soon after rejection of the xenogeneic tissue. The cell-mediated immune response, although detectable by the in vitro assays, was less pronounced than the humoral response and corroborated the histologic findings of mild lymphocyte infiltration in the rejected tissue. These results demonstrate that humoral rejection plays a predominant role in the rejection of vascularized musculoskeletal xenotransplants between concordant species. This mouse-to-rat vascularized xenograft model will be utilized for further studies on inducing tolerance to vascularized musculoskeletal xenografts. PMID- 10702739 TI - Tissue specificity in rat peripheral nerve regeneration through combined skeletal muscle and vein conduit grafts. AB - Diffusible factors from the distal stumps of transected peripheral nerves exert a neurotropic effect on regenerating nerves in vivo (specificity). This morphological study was designed to investigate the existence of tissue specificity in peripheral nerve fiber regeneration through a graft of vein filled with fresh skeletal muscle. This tubulization technique demonstrated experimental and clinical results similar to those obtained with traditional autologous nerve grafts. Specifically, we used Y-shaped grafts to assess the orientation pattern of regenerating axons in the distal stump tissue. Animal models were divided into four experimental groups. The proximal part of the Y-shaped conduit was sutured to a severed tibial nerve in all experiments. The two distal stumps were sutured to different targets: group A to two intact nerves (tibial and peroneal), group B to an intact nerve and an unvascularized tendon, group C to an intact nerve and a vascularized tendon, and group D to a nerve graft and an unvascularized tendon. Morphological evaluation by light and electron microscopy was conducted in the distal forks of the Y-shaped tube. Data showed that almost all regenerating nerve fibers spontaneously oriented towards the nerve tissue (attached or not to the peripheral innervation field), showing a good morphological pattern of regeneration in both the early and late phases of regeneration. When the distal choice was represented by a tendon (vascularized or not), very few nerve fibers were detected in the corresponding distal fork of the Y-shaped graft. These results show that, using the muscle-vein-combined grafting technique, regenerating axons are able to correctly grow and orientate within the basement membranes of the graft guided by the neurotropic lure of the distal nerve stump. PMID- 10702740 TI - Free flaps for reconstruction of the lower back and sacral area. AB - Free flap reconstruction of the lower back and sacrum is complicated by a paucity of recipient vessels and difficulties in postoperative care. From 1983 to 1997, six patients with intractable wounds of the lower back and sacral area were treated with free flaps. The flaps used were latissimus dorsi (three), combined latissimus dorsi and serratus anterior (one), and filleted leg tissue (two). The recipient vessels were the deep femoral vessels, the perforator vessels of the deep femoral system, the inferior epigastric vessels, and the superior gluteal and inferior gluteal vessels. The patients were observed in the intensive care unit for 1 week and kept in prone position for 4 weeks. All flaps survived and wounds healed primarily. For large or multiple defects of the lower back and sacrum, free tissue transfer is effective in achieving primary healing, particularly when local flaps are inadequate or have failed. PMID- 10702741 TI - Microcirculatory changes following reperfusion insult in diabetic rat skeletal muscles. AB - We investigated the microcirculatory changes of ischemia/reperfusion injury in the diabetic rat cremaster muscle as well as the therapeutic effect of insulin. Streptozotocin-induced diabetic rats were maintained hyperglycemic for up to 8 weeks or were treated with insulin in the diabetic period. The rat cremaster muscle was prepared as an island flap and subjected to 2-h clamp ischemia followed by 1-h reperfusion. In nonischemic conditions, effective concentrations for 50% response (EC50) of serial orders of arterioles to norepinephrine were higher in diabetic muscles. Ischemia/reperfusion insult significantly decreased the EC50 of arterioles in the normal group, but not in the diabetic group. Light microscopy showed that the diabetic cremasters had more collapsed capillaries and smooth muscle-disarranged arterioles. Insulin therapy showed significant improvement in the diabetes-caused reduction of perfused capillary density, but not in the contractility of the diabetic arterioles. These results indicate that diabetes mellitus may damage the skeletal muscle microvasculature irreversibly and make it less responsive to autonomic regulation. Insulin therapy can improve capillary perfusion, but not the microvascular reactivity of diabetic muscles. PMID- 10702742 TI - Assessment of muscle flap sensibility by evoked potentials in the rat. AB - This study investigated whether the sensory-to-motor reinervation of the muscle flap provides a better sensory recovery of an overlying skin graft. Fifty-four animals were studied in three groups of 18 rats each: group I (control): 1 cm of the gastrocnemius muscle motor nerve was excised and no repair was performed; group II (motor-to-motor repair): the motor nerve of the gastrocnemius flap was transected and repaired; group III (sensory-to-motor repair): the motor nerve of the gastrocnemius muscle and sural nerve were transected and their distal and proximal ends, respectively, were repaired. At follow-up periods of 6, 12, and 24 weeks, evaluation of hair growth, muscle atrophy, and sensory evoked potentials was performed. Somatosensory evoked potentials (SSEP) at 6 weeks in the sensory to-motor repair (group III) revealed a significant (P < 0. 05) increase (104.4% +/- 22.9) in the relative response of peak-to-peak potentials when compared with group I (46.6% +/- 19) and group II (51.8% +/- 14.0). Muscle flap stimulation was most prominent at 6 weeks in sensory-to-motor reinvervated flaps (group III 133.1% +/- 25.4; group I 84.9% +/- 20.2). In this study, sensory-to-motor nerve repair significantly improved the sensibility of skin flaps at 6 weeks. Denervated flaps presented with 3 months of sensory recovery delay. PMID- 10702743 TI - Model for avulsion injury in the rat brachial plexus using passive acceleration. AB - We have developed an experimental model for brachial plexus injuries in the rat that closely simulates the characteristics of human injury. This model produces avulsion injuries in a noninvasive manner. A prototype apparatus was designed that allowed a force to be transmitted to a restrained limb by passive acceleration. Reproducible results were obtained in 32 rats. A significant correlation was found between the test weight and the number of roots avulsed (r = 0.92; P < 0.05). The amount of force also correlated to the pattern of avulsion injury: a 230-g weight produced either C6 (54%), C7 (15%), or C6 and C7 (31%) avulsions; a 330-g weight produced C6 (18%), C7 (9%), or C6 and C7 (73%) avulsions; a 530-g weight produced C5 through C8 (75%) or C6 through T1 (25%) avulsions. This model of brachial plexus injury may be useful to further our understanding of the cellular response to this incapacitating injury and to develop therapeutic strategies with behavioral correlates. PMID- 10702744 TI - Do African Americans have lower energy expenditure than Caucasians? AB - OBJECTIVE: To review current studies that examine differences in energy expenditure between African Americans and Caucasians as possible modulators of attained differences in overweight status. DESIGN: Literature review of recent clinical and laboratory studies. METHODS: Studies chosen for review were those that examined directly resting metabolic rate (RMR), using indirect calorimetry, and total daily energy expenditure (TDEE) and physical activity energy expenditure (PAEE), using doubly labeled water. RESULTS: Ten of 15 studies reviewed reported a lower RMR in African Americans than in Caucasians. The differences in RMR between African Americans and Caucasians ranged from 81 to 274 kcal/day and could not be explained by differences in age, fat-free mass (FFM) or methodological concerns. Two of six studies of energy expenditure using doubly labeled water suggest that Black adults have a tendency for lower TDEE that can be accounted for primarily by a lower PAEE. CONCLUSIONS: If future studies indicate conclusively that African Americans do have lower RMR, TDEE and PAEE than Caucasians, then the disproportionally higher risk of obesity and associated metabolic disorders in Black adults may be preventable-especially in Black women. If these race differences are indeed a result of both physiological and behavioral factors, then interventions designed to reduce caloric intake and/or increase energy expenditure through lifestyle activity or structured exercise programs become especially important for African Americans and should be encouraged. International Journal of Obesity (2000)24, 4-13 PMID- 10702745 TI - Lifetime dietary change and its relation to increase in weight in Spanish women. AB - INTRODUCTION: Changes in dietary patterns and a decrease in physical activity have occurred in Western countries. These are factors in the variation in body composition observed in populations, characterized by a progressive accumulation of fat with age and a consequent increase in the risk of suffering from common chronic illnesses such as obesity, cardiovascular disease and cancer. OBJECTIVE: To investigate weight gain throughout the life-cycle and its relation to modifications in dietary patterns, analyzing the causes of these modifications and their implications for patterns of adult overweight and obesity. DESIGN: Cross-sectional sample of Spanish women from a socio-economically disadvantaged class. SUBJECTS: 1037 healthy perimenopausal women (age: 45-65 y). MEASUREMENTS: Juvenile body mass index (BMI), current BMI, food frequency questionnaire, retrospective food habits. RESULTS AND CONCLUSIONS: Of these women, 48.8% had changed their dietary habits during their lifetime. A change in diet due to migration or marriage occurred at approximately 20 years of age and was characterized by an increased frequency of consumption of foods rich in protein and complex carbohydrates, while a change due to illness occurred at around 50 years of age and was characterized by a decrease in the consumption of these types of food. The change in dietary behavior due to migration was associated with weight gain. Weight gain was also inversely associated with BMI during youth; women who in their youth had a BMI<18.5 kg/m2 gained an average of 21.4 kg, compared with those with a BMI>27 kg/m2 in their youth, who gained an average of 5.4 kg. International Journal of Obesity (2000)24, 14-19 PMID- 10702746 TI - Misreporting of total energy intake in older African Americans. AB - OBJECTIVES: (1) To examine misreporting of total energy intake in older African American men and women using the double-labeled water procedure; and (2) to identify significant physiological and demographic determinants of total energy intake misreporting in older African Americans. DESIGN: Cross-sectional study examining gender differences and determinants of misreporting of total energy intake in older African-American men and women. SUBJECTS: Sixty-four, older African-American men (n=28) and women (n=36); 52-84 y old; body mass index of 20.5-45.1 kg/m2. MEASUREMENTS: Misreporting of total energy intake (difference between reported intakes and measured energy expenditure by doubly labeled water procedure), peak VO2, resting metabolic rate (by indirect calorimetry), indices of body fat and fat distribution (by dual-energy X-ray absorptiometry and anthropometry), income, living arrangement and education (by interview). RESULTS: Older African-American men and women under-reported total energy intake to a modest degree and there were no gender differences in the magnitude of the misreporting. Peak VO2 (a determinant of daily energy requirements) and percentage intakes of fat and protein were significant correlates of misreporting of total energy intake in older African Americans. When these correlates were entered in a multiple regression model, only percentage dietary fat and protein intakes independently predicted misreporting of total energy intake. CONCLUSIONS: Cultural differences in attitudes regarding food and weight have significant effects on misreporting of total energy intake in older African Americans. In addition, individuals who misreported their total energy intake to a greater extent reported consuming less fat and more protein. Misreporting of total energy intake may occur less frequently in older African Americans as compared to other racial groups, since they may be less preoccupied with body size and image. International Journal of Obesity (2000)24, 20-26 PMID- 10702747 TI - Changes in plasma leptin and insulin action with resistive training in postmenopausal women. AB - OBJECTIVE: To determine the effects of 16 weeks of resistive training alone (RT) and with weight loss (RT+WL) on insulin action, plasma leptin concentrations and leptin's relationship to beta-cell sensitivity to glucose, resting metabolic rate (RMR), and plasma catecholamines in older women. SUBJECTS: Fifteen obese postmenopausal women aged 50-69 y. MEASUREMENTS: Body composition (by dual-energy X-ray absorptiometry), RMR (by indirect calorimetry), insulin action (by 2 h hyperglycemic clamps; 7.9 mmol/l above basal plasma glucose levels), plasma leptin and insulin (by RIA), and plasma catecholamines (by enzymatic methods). RESULTS: RT and RT+WL resulted in significant improvements in muscular strength (P<0.01) with no changes in maximal oxygen consumption. Body weight, fat mass and percent body fat did not change with RT, but decreased with RT+WL (P<0.001). Fat free mass and RMR increased after training when both groups were combined (P<0.05). The insulin response during the last 20 min of the 2 h hyperglycemic clamps decreased 16% after RT (P=0.05), 43% after RT+WL (P<0.05), and 29% in the entire group (P<0. 01) without any changes in glucose utilization. Plasma leptin levels did not change after RT, but decreased by 36% after RT+WL (P<0.05). Baseline leptin levels correlated with body weight (r=0.68, P<0.01), body fat mass (r=0.77, P<0.001), and RMR (kcal/d; (r=0.69, P<0.005), but not with baseline norepinephrine or epinephrine levels. Plasma leptin levels correlated with basal insulin (r=0.73, P<0.005), and approached significance with the 0-10 min and 100 120 min insulin response to hyperglycemia before training (both r=0.51, P=0.07). In the entire group, the change in insulin response from 100-120 min during the clamp correlated with the change in leptin levels (r=0.60, P<0.05), but this was not independent of changes in fat mass. CONCLUSIONS: Although changes in leptin levels were not related to changes in RMR or plasma catecholamines after RT with and without weight loss, the increase in insulin action after training and weight loss may be related to the decrease in leptin levels that were mediated by the loss of body fat in the obese, postmenopausal women. International Journal of Obesity (2000)24, 27-32 PMID- 10702748 TI - Mortality associated with body fat, fat-free mass and body mass index among 60 year-old swedish men-a 22-year follow-up. The study of men born in 1913. AB - OBJECTIVE: To describe differences in the 22 y mortality risk associated with body mass index (BMI), body fat or fat-free mass, in order to examine if the differential health consequences of fat and fat-free mass may be responsible for elevated mortality rates at both high and low BMI. DESIGN: Prospective cohort study, a 22 y follow-up. SETTING: General community. The study of men born in 1913, Gothenburg. SUBJECTS: 787 men aged 60 y. MAIN OUTCOME MEASURES: Number and time of total deaths from 1973 to 1995. RESULTS: The risk of dying was a linear function of percentage fat and fat-free mass, and increased from a relative risk of 1.00 in men belonging to the lowest fifth to 1.4 (95% confidence interval 1.11 1.99) in men in the highest fifth of percentage fat mass. For BMI the lowest risk was observed for men belonging to the middle fifth of BMI. When the relative risk was set at 1.00 for subjects belonging to the middle fifth of BMI the risk associated with the low BMI fifth was 1.3 (95% confidence interval 0.94-1.68) and that with the highest fifth was 1. 5 (95% confidence interval 1.09-1.96). Analyses including both body fat and fat-free mass showed that total mortality was a linear increasing function of high fat and low fat-free mass. CONCLUSION: The apparent U-shaped association between BMI and total mortality may be the result of compound risk functions from body fat and fat-free mass. International Journal of Obesity (2000)24, 33-37 PMID- 10702749 TI - Reproducibility, power and validity of visual analogue scales in assessment of appetite sensations in single test meal studies. AB - OBJECTIVE: To examine reproducibility and validity of visual analogue scales (VAS) for measurement of appetite sensations, with and without a diet standardization prior to the test days. DESIGN: On two different test days the subjects recorded their appetite sensations before breakfast and every 30 min during the 4.5 h postprandial period under exactly the same conditions. SUBJECTS: 55 healthy men (age 25.6+/-0.6 y, BMI 22.6+/-0.3 kg?m2). MEASUREMENTS: VAS were used to record hunger, satiety, fullness, prospective food consumption, desire to eat something fatty, salty, sweet or savoury, and palatability of the meals. Subsequently an ad libitum lunch was served and energy intake was recorded. Reproducibility was assessed by the coefficient of repeatability (CR) of fasting, mean 4.5 h and peak/nadir values. RESULTS: CRs (range 20-61 mm) were larger for fasting and peak/nadir values compared with mean 4.5 h values. No parameter seemed to be improved by diet standardization. Using a paired design and a study power of 0.8, a difference of 10 mm on fasting and 5 mm on mean 4.5 h ratings can be detected with 18 subjects. When using desires to eat specific types of food or an unpaired design, more subjects are needed due to considerable variation. The best correlations of validity were found between 4.5 h mean VAS of the appetite parameters and subsequent energy intake (r=+/-0.50-0.53, P<0.001). CONCLUSION: VAS scores are reliable for appetite research and do not seem to be influenced by prior diet standardization. However, consideration should be given to the specific parameters being measured, their sensitivity and study power. International Journal of Obesity (2000)24, 38-48 PMID- 10702750 TI - Determination of energy density of freely selected diets: methodological issues and implications. AB - BACKGROUND: There is increasing evidence that dietary energy density (ED, kJ/g) may be an important dietary characteristic, particularly in respect to control of energy intake; however, there are no agreed methods for deriving the ED of freely selected diets, and ED values may be markedly affected by the inclusion or exclusion of specific dietary items, particularly beverages. OBJECTIVE: To highlight the consequences of using six different methods of ED calculation, and their implications for characterizing differences between weight status groups and identifying associations of ED with macronutrient intakes. DESIGN: ED was calculated using six defined methods: (1) all food and beverages; (2) all food and energy beverages; (3) food, milk and alcohol; (4) food only; (5) all dry matter; (6) protein, carbohydrate and fat only, of varying exclusions of different beverages and water. For illustrative purposes, data from 41 lean (LE, body mass index (BMI) 20-25 kg/m2) and 34 obese (OB, BMI>/=30 kg/m2) adults who kept 4-day weighed dietary intake records are described. RESULTS: ED values (and coefficient of variation, CV) differed substantially by methods of calculation. OB reported significantly greater mean ED compared with LE by one method (all food, milk and alcohol, excluding other non-alcoholic beverages); however, the opposite was found using another method (dry weight). For most calculation methods, ED was negatively associated with percentage energy from carbohydrate for LE, in contrast to OB. All methods found positive correlations for ED and fat (g) among LE, but only one method found such a correlation among OB. Similarly, three methods produced positive correlations between ED and percentage energy fat amongst LE; however, this was only observed amongst OB with one method. CONCLUSIONS: Methods of calculating ED of freely selected diets must be carefully defined, and can markedly influence apparent relationships of ED with other dietary measures and subject characteristics. International Journal of Obesity (2000)24, 49-54 PMID- 10702751 TI - Daily energy expenditure in Mexican and USA Pima indians: low physical activity as a possible cause of obesity. AB - BACKGROUND: Obesity is caused by an imbalance between energy intake and energy expenditure. However, it is unknown whether increased physical activity protects susceptible populations against the development of obesity and type 2 diabetes. OBJECTIVE: To investigate the potential protective role of environment and physical activity against obesity by measuring total energy expenditure in Mexican and USA Pima Indians. METHODS: We compared the physical activity level of 40 (17 female and 23 male; 37+/-11 y, 66+/-13 kg) Mexican Pima Indians from a remote, mountainous area of Northwest Mexico, with 40 age-and-sex matched (17 female and 23 male; 37+/-12 y, 93+/-22 kg) Pima Indians from the Gila River Indian Community in Arizona, USA. We measured total energy expenditure (TEE) by doubly labeled water and calculated physical activity by different methods: physical activity level (PAL) as the ratio of TEE on resting metabolic rate (RMR), TEE adjusted for RMR by linear regression, activity energy expenditure adjusted for body weight (AEE), and activity questionnaire. RESULTS: Physical activity was higher in Mexican Pima Indians when compared with USA Pima Indians as assessed by PAL (1.97+/-0.34 vs 1.57+/-0.16, P<0.0001), TEE adjusted for RMR (3289+/-454 vs 2671+/-454 kcal/day, P<0.0001) and AEE adjusted for body weight (1243+/-415 vs 711+/-415 kcal/day, P<0. 0001). Questionnaires revealed more time spent on occupational activities among Mexican Pima compared with USA Pima (23.9+/-13.3 vs 12.6+/-13.9 h/week, P<0.001). CONCLUSION: These data support a significant role for physical activity in the prevention of obesity in genetically susceptible populations. International Journal of Obesity (2000)24, 55-59 PMID- 10702752 TI - Visceral fat loss evaluated by total body magnetic resonance imaging in obese women operated with laparascopic adjustable silicone gastric banding. AB - OBJECTIVE: To investigate the changes of visceral fat, as compared with total and subcutaneous adipose tissue (AT) in obese patients operated with laparascopic adjustable silicone gastric banding (LAP-BAND). SUBJECTS: Six premenopausal morbid obese (body mass index range: 41.4-44.2 kg/m2) women, aged 38-42 y, operated with LAP-BAND, evaluated before, 8 weeks after, and 24 weeks after surgery. MEASUREMENTS: Fat distribution was analysed by total body multi-slices MRI. Total AT, gluteo-femoral subcutaneous AT, abdominal subcutaneous AT, and abdominal visceral AT volumes were measured. FM was calculated from MRI determined total AT volume and AT density. RESULTS: A weight loss of 9.9+/-3.8 kg was observed in the first 8 weeks after LAP-BAND (0-8 weeks), and a further weight loss of 7.1+/-4.9 kg in the subsequent 16 weeks (8-24 weeks). Total AT showed a statistically significant reduction of 6.2+/-4.0 l in 0-8 weeks and a further significant reduction of 7.7+/-3.9 l in 8-24 weeks (P<0.01 from baseline). A similar trend was observed for both abdominal and gluteo-femoral subcutaneous AT. Visceral AT showed a statistically significant reduction of 1.0+/-0.9 l in the 0-8 weeks (P<0.05) and a further non-significant reduction of 0.6+/-0.7 l in 8-24 weeks (P<0.05 from baseline). In 0-8 weeks, the relative reduction of visceral AT was higher than the relative reduction of both total AT and gluteo-femoral subcutaneous AT. A highly significant correlation was observed between the reduction of total AT and the reduction of both abdominal and gluteo femoral subcutaneous AT. By contrast, in 0-8 weeks, the reduction of total AT and the reduction of visceral AT were not correlated. In a subsequent analysis, both observations collected in the first 8 weeks after LAP-BAND and observations collected in the last 16 weeks are simultaneously considered, leading to a total of 12 time periods (two time periods for each individual patient). In order to identify factors associated with preferential visceral fat reduction, we calculated for each of the 12 time periods the difference between the percentage changes of visceral AT and the percentage changes of total AT. The relationship between this difference and several other variables were investigated by simple correlation analysis. The only variables found to be associated were the initial visceral AT volume, the absolute level of weight loss (kg) per week of observation, and the relative level of weight loss (%) per week of observation. CONCLUSION: In the phase of rapid weight loss following LAP-BAND, a preferential mobilization of visceral fat, as compared with total and subcutaneous AT, can occur. However, this preferential visceral fat reduction occurs only in those patients presenting higher levels of visceral fat deposition at baseline and higher levels of weight loss. International Journal of Obesity (2000)24, 60-69 PMID- 10702753 TI - Subcutaneous adipose tissue expression of plasminogen activator inhibitor-1 gene during very low calorie diet in obese subjects. AB - OBJECTIVE: To determine whether changes in subcutaneous adipose tissue plasminogen activator inhibitor-1 (PAI-1) expression influence plasma PAI-1 level during weight loss in obese humans. DESIGN: Study of the variations of PAI-1 levels both in plasma and in subcutaneous abdominal adipose tissue in 15 volunteer non-diabetic obese subjects, body mass index (BMI) 40.4.+/-1.9 kg/m2, aged 48+/-3 y, before and after a 3 week very low calorie diet (VLCD) programme (3.9+/-0.1 MJ/day). MEASUREMENTS: Plasma and adipose tissue PAI-1 protein levels were measured by enzyme-linked immunosorbent assay and PAI-1 mRNA levels were quantified by quantitative RT-competitive PCR. RESULTS: VLCD induced weight loss (5.8+/-0.8 kg) and decreased plasma PAI-1 concentration (-26% (P<0. 01)). Surprisingly, PAI-1 mRNA and protein abundance in subcutaneous adipose tissue increased by 87% (P<0.05) and by 44% (P<0.01), respectively. CONCLUSION: These data indicate thus that changes in subcutaneous adipose tissue PAI-1 expression are not involved in the decrease of plasma PAI-1 levels during VLCD in obese subjects. International Journal of Obesity (2000)24, 70-74 PMID- 10702754 TI - Distribution of food intake as a risk factor for childhood obesity. AB - OBJECTIVE: The purpose of our study was to assess the relationship between nutrient intake, partitioning of food intake, parents' overweight and adiposity in a group of children. SUBJECTS: 530 7-11-year-old children: 278 males, 252 females. METHODS: Energy intake, nutrient intake and percentage distribution of the intake of energy among the different meals were assessed by means of diet history. Body composition was obtained by measuring skinfold thickness. RESULTS: We identified the relationship between the children's adiposity and their parents' body mass index (BMI) mother: r=0.12, P<0.01; father: r=0.13; P<0.01), carbohydrate (r=-0. 15, P<0.001) and fat intake (r=0.14, P<0.002), and the proportion of energy taken at dinner (r=0.1, P<0.05). A multiple regression analysis was run with a stepwise procedure using relative adiposity as the dependent variable and parents' BMI, dinner intake (percentage of energy intake), EI/BMR ratio (an index of energy intake validity), and sex (dummy variable) as independent variables. All the independent variables, except percentage of fat intake, were included in the final model. The equation was able to explain approximately 19% (R=0.44, P<0.001) of inter-individual fat mass percentage variability. CONCLUSIONS: Diet composition did not contribute to explain the children's adiposity when the parents' overweight (BMI) was taken into account. However, the percentage distribution of the intake of energy among the different meals, particularly at dinner, contributed to explain inter-individual variance of fatness in children of both sexes. International Journal of Obesity (2000)24, 75-80 PMID- 10702755 TI - Physical activity beliefs and behaviours among adults attempting weight control. AB - OBJECTIVE: To compare the frequency and duration of varying intensities of physical activity performed by adults trying to lose weight, avoid gaining weight and not actively trying to control their weight, and to compare these groups' beliefs about the physical activity they should perform. METHOD: Random postal survey of 2500 Victorian adults selected from the Australian electoral roll (response rate=42%). MEASURES: One-week physical activity recall (frequency and duration of walking, other moderate activity and vigorous activity), BMI (based on self-reported height and weight) and weight-control behaviour. RESULTS: At the time of the survey, 27% of respondents were actively trying to avoid gaining weight, 23. 9% trying to lose weight and 49.1% undertaking no weight control. Respondents spent a mean time of 4.0 (+/-7.1) h walking, 5.5 (+/-7. 9) h in moderate activity and 3.1(+/-5.9) h in vigorous activity during the week prior to the survey. Women trying to lose weight or avoid gaining weight engaged in vigorous activity more often than women not trying to control their weight. After adjusting for age, education and BMI, women trying to avoid gaining weight were 2.4 times more likely, and women trying to lose weight 2.5 times more likely, to have met current physical activity guidelines than women undertaking no weight control. On average, respondents believed they should spend 5.2 (+/-6.9) h walking, 6.5 (+/-8.2) h in moderate activity and 4.3 (+/-6.5) h in vigorous activity each week. Women trying to lose weight felt they should perform vigorous activity more often than other women. Weight-control behaviour was not associated with physical activity beliefs and behaviours of men. CONCLUSION: Walking is a common activity among adults attempting weight control. However, many men and women do not fully recognize the value of moderate-intensity physical activity. Future efforts should be directed at promoting the role of moderate-intensity activity in weight control, particularly activity that can be performed outside of planned activity sessions. International Journal of Obesity (2000)24, 81-87 PMID- 10702756 TI - Body mass index in 17-year-old Israeli males of different ethnic backgrounds; national or ethnic-specific references? AB - OBJECTIVE: To evaluate whether a single national reference is appropriate for assessing prevalence of overweight in heterogeneous populations, or whether ethnic-specific references are needed. DESIGN: A population-based study of Israeli Jewish males who underwent routine physical and clinical examinations prior to army recruitment served as the basis for the development of two types of references for body mass index (BMI): a national reference (NR) and an ethnic specific reference (ER). SUBJECTS: Consecutive cohorts of all 17-y-old Jewish male recruits (n=109, 570). MEASUREMENTS: Weight, height and blood pressure values were obtained. BMI was calculated, and the 85th percentile of BMI was used as a cut-off point for overweight, using both types of references. Prevalence of hypertension among recruits was used as a biomarker to support the reliability of the ER when discrepancy in classification between the two references was found. RESULTS: As compared to the NR, three ethnic groups had a BMI distribution shifted to the left (light sub-population) and five were shifted to the right (heavy sub-population). In the light sub-population, 7% of the inductees who were classified as having normal weight by the NR were considered overweight by the ER and had a hypertension rate similar to that of those defined as overweight by both references (3.1 per 1000). In the heavy sub-population the 4% of subjects who were overweight by NR and normal by ER had hypertension rates similar to those defined as normal weight by both references (2.7/1000), and significantly lower than that of those classified as overweight by both references (10.8/1000). CONCLUSION: In heterogeneous populations, ethnic references should be used to evaluate prevalence of overweight, rather than one national reference. International Journal of Obesity (2000)24, 88-92 PMID- 10702758 TI - Loss of total body potassium during rapid weight loss does not depend on the decrease of potassium concentration in muscles. Different methods to evaluate body composition during a low energy diet. AB - OBJECTIVE: The aim of the study was to elucidate whether combustion of skeletal muscle glycogen during a very low calorie diet (VLCD) was associated with decreased muscle potassium content. A comparison between different methods was also performed to evaluate body composition during a VLCD and a low calorie diet (LCD). DESIGN: Dietary treatment of obese women by VLCD and LCD. Measurements after 1 and 2 weeks of VLCD and 6 months of LCD. SUBJECTS: Fifteen perimenopausal obese women aged 46.5+/-1.3 y and 15 of 48.0+/-0.7 y of age. MEASUREMENTS: Skeletal muscle biopsies under local anaesthesia. Body composition measurements by means of deal-energy X-ray absorptiometry (DEXA), and measurements of total body potassium (40K) and total body nitrogen (TBN). Measurements of electrolytes and glycogen concentration in muscle samples. RESULTS: In the first study (1 week of VLCD) skeletal muscle glycogen decreased (P<0.01), but muscle potassium increased (P<0.01). Muscle sodium decreased (P<0.01), while muscle magnesium was unaltered. Body weight decreased by 2.9+/-0.5 kg and 40K decreased. Fat-free mass (FFM) calculated from 40K and DEXA decreased by 2.7 vs 1.9 kg (P<0.001). Body fat measured with DEXA decreased by 1.1 kg (P<0.01), but not body fat calculated from 40K. TBN decreased by 0.03+/-0.01 kg (P<0.05) and FFM calculated from TBN by 2.9+/-0.5 kg (P<0.002). In the second study, 6 months on the LCD resulted in 17.0+/-2.0 kg weight reduction and this was mainly due to reduced body fat, 14. 0+/-2.0 kg measured with DEXA and from 40K (P<0.001). The decrease in FFM was slight. CONCLUSION: One week of VLCD resulted in muscle glycogen depletion but increased muscle potassium content in spite of decreased total body potassium. FFM contributed to the main part of body weight loss during short periods of severe energy restriction, but remained unchanged during long-term dietary treatment. Body fat became mostly responsible for the body weight loss during long-term LCD. Calculations of changes of FFM from 40K and TBN seem to overestimate the FFM decrease associated with short-term VLCD. International Journal of Obesity (2000)24, 101-107 PMID- 10702757 TI - Association of sets of alleles of genes encoding beta3-adrenoreceptor, uncoupling protein 1 and lipoprotein lipase with increased risk of metabolic complications in obesity. AB - OBJECTIVE: To investigate the relationship between the polymorphisms of the beta3 AR (Trp64Arg), UCP1 (A-->G) and LPL (HindIII and PvuII) loci and the metabolic complications associated with obesity in a Turkish population. SUBJECTS: 271 unrelated individuals of Turkish origin including obese (body mass index, BMI>30 kg?m2) and lean (BMI< or =25 kg?m2) subjects. MEASUREMENTS: Anthropometric (weight, height and blood pressure) and metabolic measurements (plasma levels of glucose, cholesterol and triglycerides), and determination of beta3-AR, UCP1 and LPL genotypes by polymerase chain reaction followed by enzymatic digestion. RESULTS: The distributions of genotypes for each candidate gene (beta3-AR, UCP1 and LPL) were similar between the obese and the lean subjects. The Arg64 allele of the beta3-AR gene was absent from massively obese men. GG carriers of the A- >G variant of the UCP1 gene showed BMI-associated increases of cholesterol levels which were more marked than both AA (P=0.027) and AG (P=0.039) carriers. Obese P+ carriers of the LPL PvuII variant had significantly higher levels of glucose than non-carriers (P=0.011), whereas obese P+P+ carriers did not have significantly different levels of triglycerides than non-carriers (P=0.087). Moreover, carriers of both alleles (G&P+) had higher levels of glucose than non-carriers (P=0.048), but did not have significantly different levels of triglycerides than non carriers (P=0.125). However, the BMI-associated increase of triglycerides of P+&G carriers was significantly more marked than that of P+ carriers (P=0.0085). CONCLUSION: Our data support the idea that alleles of specific genes (UCP1, LPL and beta3-AR) might play a role in the development of certain metabolic complications of obesity and might have additive effects when combined with each other (as in the case of UCP1 and LPL). International Journal of Obesity (2000)24, 93-100 PMID- 10702759 TI - Body mass index and alternative indices of obesity in relation to height, triceps skinfold and subsequent mortality: the Busselton health study. AB - OBJECTIVES: The ideal index for leanness and obesity in epidemiological studies should correlate strongly with body weight and with a direct measure of fat while minimizing the influence of height. The preferred index is expected to show meaningful associations with subsequent mortality. Our aims were to compare weight/height, weight/height(2) (body mass index or BMI), and weight/height(3) as candidates for this index. DESIGN: We analysed cross-sectional data from surveys of 6948 adults (3334 men (mean age 43 y, mean BMI 24.8 kg/m2), and 3614 women (mean age 42 y, mean BMI 24.3 kg/m2)) in Busselton, Australia whose weight, height, triceps skinfold, and cardiovascular risk factors were measured from 1966 through to 1978. In these same subjects we studied the mortality risks of indices of obesity using Cox regression analysis for survival time from first survey to death, or to follow up at the end of December 1995, after adjustment for age. Subjects dying within 5 y of the baseline survey were excluded from the analysis to avoid the bias of concurrent illness. We also studied subgroups including never smokers, subjects with no heart disease, and subjects <60 years of age at first survey. RESULTS: In men, weight/height2 met the criteria for a satisfactory index in that there was a very strong correlation with triceps skinfold, and a negligible correlation with height. For women, weight/height was as good a measure as weight/height2, with both having strong correlations with triceps skinfold, and minimal correlations with height. Weight/height2 as a predictor of mortality in men of all ages showed the typical U-shaped associations that were similar and consistent and of variable statistical significance. The significances of the hazard ratio curves were the strongest for cardiovascular disease deaths (all men P=0.001; men without heart disease at baseline P<0.001; never smoking men P=0.007). In never smoking men there was a near linear positive relationship with all-cause mortality (P=0.018). In women weight/height2 showed no consistent associations with mortality. There was a shallow U-shaped relationship with all-cause mortality (P=0.087), also seen in never smoking women (P=0.075). In assessing 'ideal' weight for height in this population, a weight/height2 of 25 kg/m2 (range 22.5-27.5 kg/m2) is appropriate. Weight/height and mortality showed very similar patterns in men to weight/height2 with quite similar levels of statistical significance. In women much more pronounced U shaped curves were apparent in all groups and subgroups, with a significant all cause mortality trend for all women (P=0.029) and never smoking women (P=0.034). In assessing 'ideal' weight for height a weight/height of 42.5 kg/m (range 35-50 kg/m) appears appropriate for men and women. CONCLUSIONS: Weight/height2 is an appropriate index of leanness and obesity in males at all ages, whereas weight/height is at least as good an index for females. In mortality studies weight/height2 and weight/height predict mortality similarly in males, but weight/height is a better discriminant of mortality in females. International Journal of Obesity (2000)24, 108-115 PMID- 10702760 TI - A descriptive study of weight loss maintenance: 6 and 15 year follow-up of initially overweight adults. AB - OBJECTIVE: To describe factors associated with long-term maintenance of weight loss. DESIGN AND SUBJECTS: We identified initially overweight individuals (body mass index >27 kg/m2, n=911) from the nationwide Finnish Twin Cohort and studied those who lost at least 5% of their body weight between 1975 and 1981. Subjects who had maintained weight loss until 1990 (38 men, 17 women) were compared to both regainers (28 men, 26 women) and the other overweight subjects in the cohort. MEASUREMENTS: Self-report data on weight, height, health behaviours and perceived well-being; self-report and register-based data on health status and use of medication. RESULTS: Only 6% of all overweight individuals lost and maintained at least 5% weight loss. In men weight loss maintenance was associated with a low level of stress and health-promoting behaviours but also with medical problems. Failure to maintain weight loss seemed to be associated with stressful life and past high alcohol intake. In women weight loss maintenance was associated with low initial well-being and health-compromising behaviours that improved after weight loss. CONCLUSION: Long-term weight loss maintenance is rare. Predictors of weight loss maintenance are different between women and men. International Journal of Obesity (2000)24, 116-125 PMID- 10702761 TI - Extra cellular water and increase in capillary permeability to albumin in overweight women with swelling syndrome. AB - OBJECTIVE: To evaluate extracellular water (ECW) in the recumbent and the upright position, in overweight and lean women with swelling syndrome, and to correlate the excess in ECW with an increase in capillary filtration of albumin (CFA). PATIENTS: Fifty-one women with a swelling syndrome were investigated, 26 of whom were overweight. MEASUREMENTS: ECW was measured by the bioelectrical impedance method, in the recumbent position and again after a postural test which consisted of walking around for 30 min. CFA was studied by an isotopic test using 99m technetium-labelled albumin. RESULTS: ECW increased (>107% of the theoretical value) in 22 of the 26 overweight patients and 23 of the 25 lean patients. The CFA isotopic test was abnormal in half (11/22) of the overweight patients with increased ECW and in three of the four overweight patients with a normal ECW value. It was abnormal in 18 of the 23 lean patients with increased ECW and in the two lean patients with a normal ECW value. During the postural test, a significant (by> or =4%) increase in ECW occurred in a higher proportion of overweight patients tested (14/22) than among the lean women tested (0/5; P=0. 04). CONCLUSIONS: The swelling syndrome is indeed related to an increase in ECW in lean and overweight subjects and to a further increase in ECW after a postural test only in the overweight patients. It is also associated with microcirculatory disorders in most of the lean patients who complain of swelling and in only half of the overweight patients with the same complaints, which suggests that other factors (e.g. hormonal disorders) may be involved in the overweight patients. International Journal of Obesity (2000)24, 126-130 PMID- 10702762 TI - Interventions to prevent weight gain: a systematic review of psychological models and behaviour change methods. AB - OBJECTIVE: To identify and review published interventions aimed at the prevention of weight gain. DESIGN: A systematic review of published interventions aimed at the prevention of weight gain. METHODS: Search strategies-we searched eight databases, manually checked reference lists and contacted authors. Inclusion and exclusion criteria-studies of any design, in which participants were selected regardless of weight or age, were included. Interventions targeting a specific subgroup, multifactorial interventions, interventions aimed at weight loss, and those with an ambiguous aim were excluded. Data extraction-data were extracted on behaviours targeted for change, psychological model, behaviour change methods and modes of delivery, methodological quality, characteristics of participants, and outcomes related to body weight and self-reported diet and physical activity. Classification and validation-a taxonomy of behaviour change programmes was developed and used for classification of underlying model, behaviour change methods, and modes of delivery. The data extraction and subsequent classification were independently validated. RESULTS: Eleven publications were included, describing five distinct interventions in schools and four in the wider community. Where diet and physical activity were described, positive effects were usually obtained, but all were measured by self-report. Effects on weight were mixed but follow-up was generally short. Smaller effects on weight gain were found among low-income participants, students and smokers. Many participants in the community-based studies were overweight or obese. Study dropout was higher among thinner and lower-income subjects. CONCLUSION: Interventions to prevent weight gain exhibited various degrees of effectiveness. Definite statements about the elements of the interventions that were associated with increased effect size cannot be made as only one of the five studies that involved an RCT design reported a significant effect on weight. This intervention involved a correspondence programme and a mix of behaviour change methods including goal setting, self-monitoring and contingencies. Future interventions might be more effective if they were explicitly based on methods of behaviour change that have been shown to work in other contexts. Effective interventions would be more easily replicated if they were explicitly described. Effectiveness might be more precisely demonstrated if more objective measures of physical activity and diet were used, and if the follow-up was over a longer period. International Journal of Obesity (2000) 24, 131-143 PMID- 10702763 TI - A clinical trial of the use of sibutramine for the treatment of patients suffering essential obesity. AB - OBJECTIVE: To evaluate the safety and efficacy of Sibutramine 10 mg per os, once a day in obese patients over a period of 6 months. DESIGN: A monocenter, double blind, placebo-controlled, parallel, prospective clinical trial. SUBJECTS: 109 male and female obese patients (BMI>30 kg/m2) from 16 to 65 y entered the trial. MEASUREMENTS: Body weight, body mass index (BMI), waist and waist/hip ratio, medical history, assessment of hunger, satiety and diet compliance, standard laboratory assessments, blood pressure, heart rate and ECG. RESULTS: 40 out of 55 patients in the Sibutramine group and 44 out of 54 patients in the placebo group completed the trial. Using the method of last observation carried forward (LOCF), the weight loss in the Sibutramine group was 7.52 kg (95% confidence intervals (95% CI) 6.15; 8.9) and that in the placebo group was 3.56 kg (95% CI 2.41; 4.7). The BMI loss was 3.14 kg/m2 (95% CI 2.58; 3.69) in the Sibutramine group and 1.46 kg/m2 (95% CI 0.99; 1.93) in the placebo group. The waist reduction was 12. 51 cm (95% CI 9.25; 15.77) in the Sibutramine group and 3.26 cm (95% CI 1.38; 5.14) in the placebo group (P<0.05 by paired Student's t-test for all the intragroup comparisons). 32 Sibutramine patients had 45 adverse events, the most frequent adverse events in the Sibutramine group being dry mouth (n=19), increase in blood pressure (n=5), constipation (n=5) and tachycardia (n=5); 23 placebo patients had 29 adverse events, mainly increase in blood pressure (n=11) and dry mouth (n=10). Two Sibutramine patients withdrew from the trial due to adverse events. CONCLUSION: Sibutramine induces significant loss of body weight, BMI and waist, but does not significantly affect cardiovascular function. Sibutramine was well tolerated by most of the patients. PMID- 10702764 TI - Obesity morbidity and health care costs in France: an analysis of the 1991-1992 Medical Care Household Survey. AB - OBJECTIVE: To estimate the direct medical costs associated with obesity in France. DESIGN: Analysis of the French 1991-1992 National Household Survey database comprising a representative sample of 14, 670 individuals aged 18 y and over. A subgroup of subjects with a body mass index (BMI)>/=30 kg/m2 was compared with a control group of normal-weight individuals (BMI 18.5-25 kg/m2) matched on age, gender and education level. MEASUREMENTS: Self-reported weight and height used to calculate individual body mass index and health expenditures in a 3 month period, and morbidity as declared by respondents to the national household survey and verified on medical records. RESULTS: The direct cost attributable to obesity (BMI>/=30 kg/m2) was estimated to be in the range 4.2-8.7 billion French Francs (FF) in 1992 value, that is between 0.7 and 1.5% of total health expenditures. CONCLUSION: These results were of the same order of magnitude as similar estimates obtained by a top-down approach for the same year and setting. International Journal of Obesity (2000) 24, 151-155 PMID- 10702765 TI - Up-regulation of muscle UCP2 gene expression by a new beta3-adrenoceptor agonist, trecadrine, in obese (cafeteria) rodents, but down-regulation in lean animals. AB - OBJECTIVE: The anti-obesity properties of a new beta3-adrenergic agonist (Trecadrine) were examined in a diet-induced obesity model, including the effects on OB and uncoupling protein (UCP-1 and -2) gene expression. MEASUREMENTS: Control rats and cafeteria-fed rats were treated with placebo or Trecadrine for 35 days. Leptin and UCP (1 and 2) mRNA levels were determined by reverse transcription-polymerase chain reaction (RT-PCR) methodology in adipose tissue and gastrocnemius muscle. RESULTS: Animals fed a cafeteria diet increased body weight, fat content, white adipose tissue (WAT), brown adipose tissue (BAT) weights and oxygen consumption in relation to lean controls. A rise in plasma leptin, WAT OB gene expression as well as circulating free fatty acids levels was found in obese rats as compared with lean controls. Trecadrine administration to cafeteria-fed animals decreased fat content, WAT weight, circulating leptin and fatty acids concentrations, and WAT OB gene expression, reaching comparable values to lean controls, while WAT O2 consumption was increased in these animals. Also, an increase in BAT UCP1 mRNA levels was found through a two-way analysis of variance in control and obese animals after Trecadrine administration. Gastrocnemius muscle UCP2 gene expression was reduced in lean Trecadrine-treated and diet-induced obese animals as compared to controls, while an increase was found in cafeteria-fed animals after Trecadrine administration. A negative correlation between WAT O2 consumption and UCP2 expression was found in control animals, but not in the cafeteria-fed groups, suggesting a differential response to the beta3-adrenergic compound in lean and obese animals, which is in agreement with the reported statistical interactions between obesity and Trecadrine administration found for WAT O2 consumption and muscle UCP2 expression, as well as for plasma leptin and WAT leptin expression. CONCLUSION: The new beta3 adrenergic agonist, Trecadrine, decreases fat content and increases gastrocnemius muscle UCP2 gene expression in a diet-induced obesity model. This sheds additional light on the action mechanism of compounds with affinity for beta3 adrenoceptors and other potential anti-obesity agents. PMID- 10702766 TI - A cross-sectional survey of the opinions on weight loss treatments of adult obese patients attending a dietetic clinic. AB - OBJECTIVE: To investigate the views and opinions on weight loss treatments of adult obese patients attending a dietetic clinic. DESIGN: Cross-sectional survey. SUBJECTS: 161 adults attending dietetic outpatients clinics in Portsmouth for obesity with a body mass index of at least 30 kg/m2. MEASUREMENTS: Self administered questionnaire developed from a series of focus groups with obese adults. Key topics were previous attempts to lose weight, methods used, the role of physical activity and patients' views about treatment from health professionals. RESULTS: The preferences and usefulness of different methods to lose weight varied according to the number of attempts to lose weight, gender, age, body mass index and medical condition of the patient. Men were less likely to use special slimming products, attend slimming groups and swimming than women (odds ratios (95% confidence interval), 0.1 (0.03-0.6) for slimming groups other than Weight Watchers, 0.3 (0.1-0.80) for special slimming products and 0.3 (0.1 0.6) for swimming). Men were more likely to use physical activity (2.6 (1.1-6.2)) and in particular walking (3.7 (1.0-13.6)) and cycling (2.8 (1.0-7.6)) and were more likely to see the dietitian (3.8 (1.4-9.9)) than women. Those with more than 10 attempts to lose weight were more likely to see the dietitian (3.6(1.6-8.2)), use Weight Watchers (2.5 (1.1-5. 6)) and newspapers and magazines (4.4 (1.8 10.9)) than those with fewer attempts. The younger age group were more likely to use more vigorous forms of exercise (4.2 (1.6-11.2) for keep fit and 3.7 (1. 5 9.6) for cycling) than the older subjects. The most obese were more likely to have negative views on their treatment by health professionals (4.4 (1.9-9.8) 'chairs are never big enough' and 4.0 (1.8-8.8) 'I am regarded as a second class citizen') than those who were less obese. Those without a medical condition were more likely to exercise (2.8 (1.3-6.3)) and use books (4.8 (2.0-11.2)) than those with a medical condition. CONCLUSION: The views of obese people should be considered when planning services for the treatment of obesity and a variety of options should be available. International Journal of Obesity (2000) 24, 164-170 PMID- 10702767 TI - Body fat distribution in Alaskan Eskimos of the Bering Straits region: the Alaskan Siberia Project. AB - OBJECTIVE: To describe the body fat content and distribution of adult Alaska Natives of the Bering Straits Region. DESIGN: Cross-sectional screening in the spring of 1994. SUBJECTS: 454 non-pregnant native residents from four rural Alaskan villages. MEASUREMENTS: Height, weight, waist, hip and thigh circumference, bioelectrical impedance, sagittal abdominal diameter, and triceps, biceps, suprailiac, subscapular and thigh skinfolds. RESULTS: Mean height, weight and subscapular-to-triceps ratio were higher in men than women. The women had larger waist, hip and thigh circumferences, higher body fatness, as well as larger skinfolds than the men. There were no demonstrable differences between men and women in measures of body fat distribution. The proportions of women and men with high waist-to-hip ratio (>/=0.8 for women, >/=0.9 for men) for low (<25 kg/m2), medium (25-30 kg/m2) and high (>30 kg/m2) body mass index (BMI) groups were compared with a Canadian study of all races. 1 In the lowest BMI subgroup (<25 kg/m2) a much higher proportion of Eskimo women exhibited a high waist-to hip ratio (91%) than Eskimo men (42%) or Canadian women (29%) or men (51%). In the highest BMI subgroup (>30 kg/m2) Eskimo women were similar in proportion of high waist-to-hip ratio (99%) compared to Eskimo men (100%), but still demonstrated a much greater proportion of subjects with high waist-to-hip ratio than either Canadian men (90%) or women (76%). CONCLUSIONS: The large abdominal fat depots found in Eskimo women were similar to men, and may indicate that future increases in diabetes mellitus and other metabolic alterations can be anticipated. International Journal of Obesity (2000) 24, 171-179 PMID- 10702768 TI - Predictive value of abdominal obesity cut-off points for hypertension in blacks from west African and Caribbean island nations. AB - BACKGROUND: Waist circumferences (WC) >/=94 cm for men and >/=80 cm for women (action level I) and >/=102 cm for men and >/=88 cm for women (action level II) have been suggested as limits for health promotion purposes to alert the general public to the need for weight loss. In this analysis we examined the ability of the above cut-off points to correctly identify subjects with or without hypertension in Nigeria, Cameroon, Jamaica, St Lucia and Barbados. We also determined population- and gender-specific abdominal adiposity cut-off points for epidemiological identification of risk of hypertension. METHODS: Waist measurement was made at the narrowest part of the torso as seen from the front or at midpoint between the bottom of the rib cage and 2 cm above the top of the iliac crest. Sensitivity and specificity of the established WC cut-off points for hypertension were compared across sites. With receiver operating characteristics (ROC), population- and gender-specific cut-off points associated with risk of hypertension were determined over the entire range of WC values. RESULTS: Predictive abilities of the established WC cut-off points for hypertension were poor compared to the specific cut-off points estimated for each population. Different values of WC were associated with increased risk of hypertension in these populations. In men, WC cut-off points of 76, 81, 80, 83 and 87 cm provided the highest sensitivity for identifying hypertensives in Nigeria, Cameroon, Jamaica, St Lucia and Barbados, respectively. The analogous cut-off points in women were 72, 82, 85, 86 and 88 cm. CONCLUSIONS: The waist cut-off points from this study represent values for epidemiological identification of risk of hypertension. For the purpose of health promotion, the decision on what cut-off points to use must be made by considering other additional factors including overall impact on health due to intervention (e.g. weight reduction) and potential burden on health services if a low cut-off point is employed. There is a need to develop abdominal adiposity cut-off points associated with increased risks for cardiovascular diseases in different societies, especially for those populations where the distribution of obesity and associated risk factors tends to be very different from those of the technologically advanced nations. International Journal of Obesity (2000) 24, 180-186 PMID- 10702769 TI - Orlistat fails to alter postprandial plasma lipid excursions or plasma lipases in normal-weight male volunteers. AB - OBJECTIVES: After 10 d of orlistat administration (120 mg three times/day), the primary objective was to determine the drug's effect on postprandial plasma lipoprotein lipase (LPL) and hepatic triglyceride lipase (HTGL) activities on day 10 after an oral fat-load. The secondary objectives were to determine the effects of orlistat on 12 h postprandial measures of: (1) preheparin HTGL and LPL; and (2) serum triglycerides, very-low-density lipoprotein cholesterol, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol and free fatty acids. METHODS: Twenty-four normal-weight, healthy male volunteers were randomized to either 120 mg orlistat (n=12) or placebo (n=12) three times a day with meals for 10 d. Preheparin LPL and HTGL activities and LPL specific activity were measured in the fasted state on days 1, 5, and 10. On days 5 and 10 the study medication (orlistat or placebo) was taken at the beginning of a fat-rich breakfast and serum lipid and lipoprotein levels monitored for 12 h postprandially. On day 10, 15 min postheparin HTGL activity was measured 8 h after the fat-rich breakfast. RESULTS: No differences were found between groups in fasting levels of preheparin LPL or HTGL activity or in LPL-specific activity on days 1, 5 and 10. No difference was found between the two treatment groups in postheparin HTGL activity 8 h after the fat-rich breakfast. Also, no differences were found between the two groups in plasma triglycerides or lipoproteins. CONCLUSION: The results indicate that the oral administration of orlistat (120 mg t. i.d.) does not significantly alter plasma triglycerides or lipoproteins, and that the inhibitory effect of orlistat on lipases is limited to the gastrointestinal tract and is not manifested systemically. PMID- 10702770 TI - Impact of the Peroxisome Proliferator Activated Receptor gamma2 Pro12Ala polymorphism on adiposity, lipids and non-insulin-dependent diabetes mellitus. AB - OBJECTIVE: The Pro12Ala polymorphism of the Peroxisome Proliferator Activated Receptor gamma2 (PPARgamma2) gene has been inconsistently associated with body mass index variations and non-insulin-dependent diabetes mellitus (NIDDM). We investigated the impact of this polymorphism on obesity markers, lipid and glucose variables in a sample of French subjects and evaluated its possible role in the onset of NIDDM. DESIGN AND SUBJECTS: Within the framework of the WHO MONICA project, a population study composed of 1195 subjects aged 35-64 y was randomly sampled from the electoral rolls of the urban community of Lille, in northern France. Subjects receiving medical treatment for hypercholesterolemia, hypertension or diabetes mellitus were excluded for the analyses, to avoid any interferences between medical treatment and biological variables. This resulted in a sample size of 839 subjects (421 men/418 women, age=49.4+/-8.1 y, body mass index (BMI)=25.7+/-4.4 kg/m2). To evaluate the role of the Pro12Ala polymorphism in the onset of NIDDM, we evaluated its distribution in 170 Caucasian NIDDM subjects from a clinical series (117 men/53 women, age=62.3+/-9.0 y, BMI=30.1+/ 3.6 kg/m2). MEASUREMENTS: The PPARgamma2 Pro12Ala polymorphism genotyping was carried out with allele specific oligonucleotides hybridisation. Data were statistically analysed for association with various obesity markers (body weight (BW), BMI, waist-to-hip ratio (WHR), plasma leptin concentrations, lipid and glucose variables. RESULTS: In the WHO-MONICA population, the Ala allele frequency was 0.11. The presence of the Ala allele was significantly associated with higher body weight (P=0.002), BMI (P=0.02), height (P=0.02) and waist circumference (P=0.04). Increased plasma concentrations of total cholesterol (P=0.01), LDL-cholesterol (P=0.004) and apolipoprotein B (P=0.01) were also detected in Ala allele bearers. The distribution of the Pro12Ala polymorphism was similar in NIDDM subjects (Ala allele frequency: 0.10) and in the WHO-MONICA population subjects. CONCLUSION: Our results suggest that genetic variability of PPARgamma2 affects body weight control and lipid homeostasis in humans and do not support a significant role for the PPARgamma2 Pro12Ala polymorphism in the aetiology of NIDDM. International Journal of Obesity (2000) 24, 195-199 PMID- 10702771 TI - Validation of the BOD POD with hydrostatic weighing: influence of body clothing. AB - OBJECTIVE: Whole body air-displacement plethysmography (BOD POD), a new body composition technique, was validated against hydrodensitometry (UWW) in 67 women wearing a one-piece swimsuit (OP) who represent a wide range of body fatness and age. Additionally, the effect of trapped isothermic air in clothing while in the BOD POD was examined by comparing different clothing schemes (a one-piece swimsuit (OP), two-piece swimsuit (TP), a hospital gown (HG), and a hospital gown previously included in a volume calibration (GC)) in a subset of 25 women. DESIGN: Cross-sectional data analysis. SUBJECTS: 67 healthy Caucasian females. MEASUREMENTS: Body density g/cm3 (Db) by BOD POD and UWW. RESULTS: In 67 females UWW Db (1.030+/-0.020 g/cm3) was higher (P<0.01) than BOD POD Db (1. 028+/-0.020 g/cm3). This is a difference of 1.0% fat. The R2 was 0. 94, SEE was 0.005 g/cm3 and the regression between Db by UWW and BOD POB did not significantly deviate from the line of identity. In the subset group of 25 subjects, OP Db (1.040+/ 0.014 g/cm3) and TP Db (1.040+/-0.014 g/cm3) were significantly lower (P<0.01) than UWW Db (1.044+/-0.014 g/cm3) or a difference of 1.9% fat. The R2 was 0.86 and the SEE was 0.005 g/cm3 and the regression between Db by UWW and both OP and TP did not significantly deviate from the line of identity. HG Db (1.056+/-0.016 g/cm3) and GC Db (1.037+/-0.016 g/cm3) were significantly different (P<0.01) from UWW Db (1.044+/-0. 014 g/cm3). This difference in density translates to a difference of 5.5% and 3.2% fat respectively. The regression between Db by UWW and both HG and GC significantly deviated from the line of identity. CONCLUSION: This study supports the use of the BOD POD as a substitute for UWW. However, caution should be made in using the BOD POD if subjects are clothed in anything other than a tight fitting swimsuit. PMID- 10702772 TI - Melanocortin 3 receptor (MC3R) gene variants in extremely obese women. AB - OBJECTIVE: Following several reports of linkage of obesity related phenotypes to human chromosome 20q we sought to determine whether variations of the melanocortin 3 receptor (MC3R) gene are associated with obesity. DESIGN: We screened the MC3R gene coding region and approximately 2 kb of 5' and 3' flanking sequences for DNA variants in unrelated extremely obese women and average weight controls using polymerase chain reaction (PCR) single strand conformation polymorphism (SSCP) analysis and DNA sequencing. SUBJECTS: 124 unrelated extremely obese women (body mass index, (BMI)>/=40 kg/m2) and 85 average weight controls (BMI<27 kg/m2). MEASUREMENTS: Radiation hybrid (RH) mapping was performed to localize the MC3R gene. 5' and 3' flanking sequences of MC3R gene were cloned. PCR-SSCP and DNA sequencing were used to detect mutations in the MC3R gene coding region and flanking sequences. RESULTS: RH mapping localized the MC3R gene to 20q13, between markers D20S100 and D20S149. 1083 bp 5' and 653 bp 3' flanking region of the MC3R gene were cloned. A missense mutation (+241, codon 81 ATT/GTT, Ile-->Val) was found in the MC3R coding region. Four more variants were detected in the 5' flanking sequence: -201(C-->G), -239 (A-->G), -762(A-->T) and 769(T-->C). Compared with controls, no significant allele frequency differences were found. Racial differences were found for the +241, -201, -239 and -762 polymorphisms. CONCLUSIONS: Several sequence variants were found in the MC3R gene coding region and in 5' flanking sequences. However, none of the variants were associated with obesity phenotypes. The linkage of extreme human obesity on 20q13 is likely caused by genes other than MC3R. International Journal of Obesity (2000) 24, 206-210 PMID- 10702773 TI - Growth and overweight of Navajo youth: secular changes from 1955 to 1997. AB - OBJECTIVE: To examine the growth status, prevalence of risk of overweight and of overweight, and secular changes in growth status in Navajo youth from 1955 to 1997. SUBJECTS: 526 (256 males, 270 females) Navajo children 6-12 y of age. MEASUREMENTS: Stature and mass were measured and the body mass index (BMI) was calculated. ANALYSIS: All three variables were plotted relative to age- and sex specific US reference data and the prevalence rates for risk of overweight and of overweight were estimated using the BMI as the criterion. The cut-off for the risk of overweight was the age- and sex-specific 85th and 95th percentiles of NHANES I, while the cut-off for overweight was a BMI>/=95th percentiles. Age specific sex differences were compared using independent samples t-tests. Secular changes for body size were estimated by comparing age- and sex-specific means for stature, mass, and the BMI in the present study and two previous studies in 1955 and 1989. RESULTS: No statistically significant differences were observed between sexes within age groups. In both sexes, mean age-specific stature appeared to be relatively stable around the 50th percentile of US reference values. Mean age specific mass appeared to be relatively stable between the 50th and 90th percentiles of the reference values, while the mean BMI tended to fluctuate about the 85th percentile. Approximately 41% of the Navajo boys and girls 6-12 y of age had BMIs >/=85th percentiles of US reference data. Compared to corresponding data on Navajo youth in 1955 and 1989, the current sample was larger in mass and the BMI. The estimated rate of secular change in mass was about 1.5 kg/decade in younger boys and girls, and about 3 kg/decade in older boys and girls between 1955 and 1997. The estimated rate of secular change in the BMI was about 0.5-1.0 units/decade between 1955 and 1997, while that for stature was about 2 cm/decade between 1955 and 1997. CONCLUSIONS: The results are consistent with recent findings on the Navajo Health and Nutrition Survey that overweight is a serious public health concern across the lifespan in the Navajo, and that the problem begins in childhood. Furthermore, Navajo children appear to be heavier than about a decade ago. International Journal of Obesity (2000) 24, 211-218 PMID- 10702774 TI - Primary prevention of weight gain for women aged 25-34: the acceptability of treatment formats. AB - OBJECTIVE: To determine if treatment format will affect the willingness of women aged 25-34 to participate in a program for primary prevention of weight gain. DESIGN: 102 normal-weight women aged 25-34 were randomized to one of three treatment formats (group meetings, correspondence course, no-treatment control). Acceptability was evaluated by determining the proportion of women participating in their assigned format. Efficacy was assessed by determining mean weight changes at post-treatment (10 weeks) and 6-month follow-up, and the proportions of women who remained at baseline weights. RESULTS: Significantly fewer women chose to participate in a group format, compared to the correspondence course and no-treatment control (42%, 84% and 62%, respectively). However, the group format produced the largest short-term changes in weight (-1.9+/-1.8 kg, -1.1+/-2.1 kg and -0.2+/-1.3 kg, respectively). CONCLUSIONS: The format of prevention programs may influence the willingness of subjects to participate, as well as treatment outcome. Both format acceptability and efficacy should be considered in determining the overall effectiveness of a program. International Journal of Obesity (2000) 24, 219-225 PMID- 10702775 TI - Effect of menopausal status on body composition and abdominal fat distribution. AB - OBJECTIVE: Preliminary studies suggest that the menopause transition is associated with deleterious changes in body composition and abdominal fat distribution. Limitations of the methodology used in these studies, however, render their conclusions controversial. Thus, the present study used radiologic imaging techniques to examine the effect of menopausal status on body composition and abdominal fat distribution. DESIGN: Cross-sectional. SUBJECTS: Fifty-three healthy, middle-aged, premenopausal women (mean+/-SD; 47+/-3 y) and 28 early postmenopausal women (51+/-4 y). MEASUREMENTS: Total and regional body composition by dual energy X-ray absorptiometry and abdominal fat distribution by computed tomography. RESULTS: No differences in total body fat-free mass or appendicular skeletal muscle mass were noted between groups. In contrast, total body fat mass was 28% higher (23+/-7 vs 18+/-7 kg) and percentage fat 17% higher (35+/-6 vs 30+/-9%; both P<0.01) in postmenopausal women compared with premenopausal women. Postmenopausal women had a 49% greater intra-abdominal (88+/ 32 vs 59+/-32 cm2; P<0.01) and a 22% greater abdominal subcutaneous fat area (277+/-93 vs 227+/-108 cm2; P<0.05) compared to premenopausal women. The menopause-related difference in intra-abdominal fat persisted (P<0.05) after statistical adjustment for age and total body fat mass, whereas no difference in abdominal subcutaneous fat was noted. A similar pattern of differences in total and abdominal adiposity was noted in sub-samples of pre- and postmenopausal women matched for age or fat mass. CONCLUSION: Our data suggest that early postmenopausal status is associated with a preferential increase in intra abdominal fat that is independent of age and total body fat mass. International Journal of Obesity (2000) 24, 226-231 PMID- 10702776 TI - Body mass index (BMI) in Turner Syndrome before and during growth hormone (GH) therapy. AB - OBJECTIVE: To study whether body mass index (BMI) is different in girls with Turner syndrome (TS) compared to normal girls, and whether BMI in TS is affected by growth hormone (GH) treatment. DESIGN: A retrospective cross-sectional study. SUBJECTS: 2468 girls with TS enrolled in the National Cooperative Group Study (NCGS), a collaborative surveillance study for assessing GH-treated children. MEASUREMENTS: BMI and BMI standard deviation score (BMI SDS) at baseline and during GH treatment were computed from height and weight data. RESULTS: BMI in TS patients increases with age as expected. However, BMI SDS increased starting at about age 9 y. A similar pattern of increase in BMI SDS was observed after each year of GH treatment for up to 4 y, but GH treatment did not change the magnitude of increase. BMI and BMI SDS curves before and during GH treatment were essentially superimposable. CONCLUSION: These findings suggest that mechanisms specific for TS are responsible for the age-related increase in BMI SDS. This increase was unaffected by GH treatment. PMID- 10702777 TI - Short-term weight cycling in aging female rats increases rate of weight gain but not body fat content. AB - OBJECTIVE: To examine the effects of short-term repeated weight cycling (WC) above and below the baseline (BL) body weight (BW) on body weight regulation, feeding efficiency, and fat content in old female Wistar rats when dietary fat content was kept constant. DESIGN: Completely randomized. ANIMALS AND METHODS: Female Wistar rats, 11 months old at the beginning of the study, were randomly divided into six groups (12 per group) after a group of rats (BL) was sacrificed for baseline data collection: the high fat gain (HFG) group gained weight to 20% above the BL weight with a high fat diet (HF) and returned to BL level by food restriction (50% of ad-libitum amount) for five cycles; the high fat loss (HFL) group lost weight to 20% below the BL weight by food restriction (50% of ad libitum amount) and regained to BL level by ad-libitum feeding for four cycles; the high fat ad-libitum (HFA) and low fat ad-libitum (LFA) groups were fed HF and low fat (LF) diet, respectively, ad-libitum for the entire study; the high fat restricted (HFR group) and low fat restricted rats (LFR group) were fed the HF and LF diet, respectively, in restricted amounts to maintain BW at BL level. RESULTS: A trend of increased rates of weight gain and feeding efficiencies from the first to last cycles for both WC groups was observed, and significant increases was observed between cycles 4 and 5. The rate of weight gain and feeding efficiency of HFL was significantly higher than that of the HFG group for all cycles (P<0. 05). The rates of weight loss were significantly decreasing with each successive cycle for HFG, but were unchanged for HFL. Percentage of body fat was not modified permanently from BL to sacrifice for both HFG and HFL groups. The body fat of HFA was higher than that of the other groups (P<0.01), while the body fat of LFA was significantly higher than that of the LFR, BL and HFL groups (P<0.01), but was similar to that of the HFG and HFR groups. The body fat of WC groups and HFR were similar to each other. The percentage of internal fat (retroperitoneal+omental) were similar for the WC groups. The percentage of internal fat of the HFG, HFR and LFA groups were similar, but were significantly higher than that of the BL and LFR groups (P<0.05). The percentage of internal fat of HFA was significantly higher than that of the rest of the groups (P<0.01). CONCLUSION: Short-term WC did not affect body fat content in these animals, but since weight gain became easier and weight loss became more difficult for animals in the HFG group, repeated WC may promote obesity in these rats. PMID- 10702778 TI - Central obesity, depression and the hypothalamo-pituitary-adrenal axis in men and postmenopausal women. AB - OBJECTIVE: We examined the relationship of adiposity to pituitary-adrenal responses to corticotrophin-releasing hormone (CRH) in men and postmenopausal women, controlling for the influence of depression. DESIGN: Studies of CRH responses, cortisol metabolite levels and depression scores in relation to adiposity in men and postmenopausal women. SUBJECTS: Thirteen men: age (median, interquartile range) 62 y (52-63), body mass index (BMI) 29.0 kg/m2 (26.3-33.1), waist circumference (waist) 105 cm (97-111), waist:hip ratio (WHR) 1.03 (0.98 1.07), subscapular to triceps skinfold thickness ratio (STR) 2.0 (1.2-2.4), total body fat (TBF) 25.4 kg (19.8-28.8); and eight women: age 54 y (53-62), BMI 30 kg/m2 (23-41), waist 86 cm (79-117), WHR 0.94 (0.87-1.10), STR 1.0 (0.85-1.07), TBF 35.0 kg (18.7-48.8). MEASUREMENTS: A standard CRH test was conducted with additional basal samples taken for leptin and interleukin 6 (IL-6). Total urine cortisol metabolites (TCM) and the ratio of urinary cortisol:cortisone (Fm/Em) metabolites were measured. Depression scores were measured by the General Health Questionnaire (GHQ-30) and Hospital Anxiety and Depression Scale (HAD) questionnaire. All subjects completed an overnight dexamethasone suppression test. RESULTS: The basal to peak percentage increments (%inc.) in adrenocorticotrophic hormone (ACTH) and cortisol in men correlated directly with STR (ACTH %inc. r=0.70, P<0.01; cortisol %inc. r=0.55, P=0.05); this relationship was independent of depression scores. In women, the ACTH area under incremental curve (AUIC) correlated negatively with STR (r=-0.81, P<0.05). In men, but not in women, there was a significant correlation between GHQ-30 score and ACTH AUIC (r=0.62, P<0.05) and cortisol AUIC (r=0.72, P<0.01). Depression scores were consistently and directly related to indices of obesity and central obesity. There were no significant relationships in either sex between urinary TCM or Fm/Em ratio and BMI, waist, WHR, TBF, STR or CRH responses. The urinary Fm/Em ratio was higher in men than in women (median 0.74 vs 0.66, P<0.05). In men, but not in women, GHQ-30 scores correlated positively with urinary TCM (r=0.57, P=0.05) and HAD-depression scores were inversely related to the urine Fm/Em ratio (r=-0.65, P<0.05). All subjects suppressed normally with dexamethasone. CONCLUSIONS: Cortisol metabolite levels were increased in depression in men, but were not related to adiposity in either sex. We demonstrate that central obesity in men, but not postmenopausal women, is associated with an enhanced pituitary adrenal response to CRH and that this relationship is independent of depression score. International Journal of Obesity (2000) 24, 246-251 PMID- 10702779 TI - Green tea and thermogenesis: interactions between catechin-polyphenols, caffeine and sympathetic activity. AB - The thermogenic effect of tea is generally attributed to its caffeine content. We report here that a green tea extract stimulates brown adipose tissue thermogenesis to an extent which is much greater than can be attributed to its caffeine content per se, and that its thermogenic properties could reside primarily in an interaction between its high content in catechin-polyphenols and caffeine with sympathetically released noradrenaline (NA). Since catechin polyphenols are known to be capable of inhibiting catechol-O-methyl-transferase (the enzyme that degrades NA), and caffeine to inhibit trancellular phosphodiesterases (enzymes that break down NA-induced cAMP), it is proposed that the green tea extract, via its catechin-polyphenols and caffeine, is effective in stimulating thermogenesis by relieving inhibition at different control points along the NA-cAMP axis. Such a synergistic interaction between catechin polyphenols and caffeine to augment and prolong sympathetic stimulation of thermogenesis could be of value in assisting the management of obesity. International Journal of Obesity (2000) 24, 252-258 PMID- 10702780 TI - Leptin deficiency due to lipid apheresis: a possible reason for ravenous hunger and weight gain. AB - OBJECTIVE: To investigate how extracorporal cholesterol lowering therapy affects circulating leptin levels in patients with ravenous hunger after treatment and permanent weight gain. DESIGN: A case report. SUBJECT: 51 y old caucasian male patient with moderate chronic renal failure. MEASUREMENTS: Serum Leptin concentration (RIA, Linco Research Inc, St. Louis, MO, USA), total cholesterol, low density lipoprotein cholesterol, blood glucose levels, calorie intake by food records. RESULTS: During treatment total cholesterol was reduced by 50%. Serum Leptin levels showed a 42% reduction at the end of treatment, that by far exceeds the physiological diurnal variation. Calorie intake was significantly increased on days of treatment. CONCLUSION: We conclude that this artificial reduction in circulating leptin plays an important role in the pathogenesis of ravenous hunger and weight gain under extracorporal cholesterol lowering therapy in this case. PMID- 10702781 TI - Weight and ventilation. PMID- 10702782 TI - The epidemiology of mouth cancer: a review of global incidence. AB - Mouth cancer (143-145 ICD-9) is a major health problem in many parts of the world. While its incidence is relatively low in most western countries there are some important exceptions to this trend: on the Indian subcontinent and in other parts of Asia it remains one of the most common forms of cancer. This review article summarises the global incidence of mouth cancer using cancer maps. Data have been compiled from the latest edition of Cancer Incidence in Five Continents and recent studies from various locations around the world. Significant geographic variation is noted in the incidence of mouth cancer, with high rates reported for the Indian subcontinent and parts of Asia (male incidence rates in excess of 10 per 100,000 per annum). It is also noted that as with other forms of oral cancer, the majority of population-based data for mouth cancer comes from the Western world with a paucity of reliable data from the so-called developing countries. Mouth cancer remains a serious health problem in many parts of the world with many regions reporting increasing incidence rates particularly in males. Ongoing research into the aetiologic risk factors associated with this disease must remain a very high priority if the causes of mouth cancer are to be established and disease control protocols introduced widely. PMID- 10702783 TI - The epidemiology of tongue cancer: a review of global incidence. AB - The tongue (141 ICD-9) is the most common intraoral site for cancer in most countries, however its global epidemiology shows significant geographic variation. This review paper summarises the global incidence of cancer of the tongue using cancer maps and references to recent studies from various locations. Tongue cancer remains a serious health problem in many countries including India (male incidence rates up to 6.5 per 100,000 per annum) and parts of Europe (male incidence rates in France up to 8.0 per 100,000 per annum). It is noted that as with other forms of oral cancer the majority of population-based data for tongue cancer comes from the Western world with a paucity of reliable data from the so called developing countries. The tongue remains the most common intraoral site for oral cancer worldwide and in a number of countries it is a serious public health problem with significant morbidity and mortality. While the incidence of tongue cancer appears to be stable or falling in some regions of the world, in other areas it is rising, particularly among younger people. PMID- 10702784 TI - Pathology and clinical correlates in oral candidiasis and its variants: a review. AB - Although Candida albicans is well recognised as the major agent of oral candidiasis, it is not clear why several variants such as pseudomembranous (PC), erythematous (EC) and hyperplastic candidiasis (HC) manifest in different individuals, sometimes singly and on other occasions, in combination. The present review focuses on recent histopathologic and immunocytochemical studies as well as the pathogenic attributes of the yeast, in an attempt to address the following queries. (1) Do histopathologic studies of the different variants of candidiasis in immunocompetent and immunocompromised individuals help explain these varying manifestations? (2) Under what circumstances does oral candidiasis manifest as a pseudomembranous rather than an erythematous lesion or vice versa? (3) Are there differences in immunoreactivity in closely adjacent mucosae so that the variable presentation of such lesions reflect differences in the local mucosal immune system? Recent studies of PC, EC and HC offer some insights into the pathogenic mechanisms involved. Histopathologic and immunohistochemical finding in cases of PC and EC in HIV-infected patients and controls appear to be comparable, with a marked reduction or even an absence of CD4+ cells. The latter phenomenon is marked in PC compared with the EC, and explicable in terms of a breakdown of the local immune response in the former, and a hypersensitivity reaction against Candida antigens in the latter. Hyperplastic candidiasis on the other hand could be considered a superficial cellular reaction against the pathogen, which cannot entirely be eradicated by the systemic or local host immune response. The virulent attributes of the fungus, such as the production of extracellular proteinases, do significantly differ within and between species and thereby play a contributory role in the genesis of the clinical variants. Although the available data do give a tantalising glimpse of the contributory mechanisms for the aetiopathology of PC, EC and HC, further research is warranted to elucidate response of the host to this ubiquitous fungal pathogen. PMID- 10702785 TI - HIV topic update: oro-genital transmission of HIV. AB - Several viruses, including the human immunodeficiency virus (HIV), can be found in blood and many body fluids including saliva, and are transmissible sexually across genital and particularly anal mucosae. A persisting concern has been the question of transmission of HIV by oral sexual practices. This review discusses the evidence for oro-genital transmission of HIV, detailing the presence and infectivity of HIV in genital fluids and saliva, the case reports and epidemiology of oro-genital HIV transmission, and the evidence from animal studies. Oral intercourse is not risk-free. The evidence suggests that the risk of HIV transmission from oro-genital sexual practices is substantially lower than that from penile-vaginal or penile-anal intercourse, that exposure to saliva presents a considerably lower risk than exposure to semen, and that oral trauma and ulcerative conditions might increase the risk of HIV transmission. PMID- 10702786 TI - An epidemiological study of supernumerary primary teeth in Japanese children: a review of racial differences in the prevalence. AB - OBJECTIVE: To clarify the prevalence of supernumerary primary teeth in Japanese children, we evaluated this prevalence in a sample of 8122 children aged 3-6 years. Furthermore, we undertook a statistical comparison of the findings of the present study with those of previous investigations on the prevalence of supernumerary primary teeth in Japanese, Chinese and Caucasian children. SUBJECTS: The subjects used in the study consisted of 8122 children aged 3-6 years (4102 boys and 4020 girls). The children were examined in kindergartens. METHODS: The prevalence of supernumerary primary teeth was recorded by visual inspection. Statistical analysis was carried out using the chi 2 test. RESULTS: Four cases of supernumerary primary teeth were found among the children examined, and thus the prevalence was 0.05%. All of the supernumerary primary teeth were located in the maxillary lateral incisor area. CONCLUSIONS: The prevalence in a sample of 8122 Japanese children was 0.05% and that in a combined sample of 65,068 Japanese children was 0.06%. PMID- 10702787 TI - Isolation of fusobacteria from the oral cavities of malnourished Nigerian children living in agricultural and herding villages. AB - A previous study demonstrated the presence and possible involvement of Fusobacterium necrophorum in the pathogenesis of noma lesions of children living in agricultural and herding villages in northwestern Nigeria. In order to determine if F. necrophorum was part of the oral flora of malnourished children with no noma lesions, a study of the fusobacteria present in the oral cavities of 30 children, 2-6 years of age in Sokoto State, was undertaken. Swabs taken of the oral cavity were cultured on selective fusobacteria medium using conventional anaerobic microbiological techniques. F. nucleatum was recovered from each child and F. necrophorum was isolated from the oral cavity of only one child. The presence of F. nucleatum and the lack of F. necrophorum, except in one case, suggests that the latter is not normal flora in the children at risk for noma. F. necrophorum, a putative trigger organism for noma may gain a foothold only when certain staging conditions (i.e., lowered host resistance and/or oral lesion) are present. PMID- 10702788 TI - Occurrence of oral lesions in relation to clinical and immunological status among HIV-infected adult Tanzanians. AB - OBJECTIVE: To determine the association, if any, between the presence of oral lesions and clinical and immunological status of untreated HIV-infected adults in Tanzania. DESIGN: A cross-sectional study. SETTING: AIDS Clinical Trial Clinic (ATCC) at Muhimbili Medical Centre in Dar-es-Salaam, Tanzania. SUBJECTS: 192 HIV infected individuals not receiving treatment; 156 individuals confirmed to be HIV seronegative acted as a control group. METHODS: Examination of oral structures, determination of HIV serostatus, clinical status, and peripheral CD4+ T cell and total lymphocyte counts. MAIN OUTCOME MEASURE: Presence of oral lesions. RESULTS: Intra-oral lesions were seen among 7.7% of the HIV-seronegative, 10.4% of the HIV seropositive and 36.8% of the AIDS groups, respectively. Enlarged parotid glands were seen in 20% of the AIDS patients, 11.9% of the HIV-seropositives, and 5.1% of the HIV seronegatives. Enlargement of submandibular salivary glands was seen in 29.6% of the AIDS patients, 31.3% of the HIV-seropositives compared with 14.7% among the HIV-seronegatives. Multiple regression analysis was used to calculate adjusted odds ratio (OR) for presence of oral lesions. OR for an intra-oral lesion was 1.6 (95% CI = 0.5; 5.0) among the HIV-seropositives and 8.2 (95% CI = 3.5; 19.7) among the AIDS patients using the HIV-seronegatives as reference. OR for an intra-oral lesion was 0.9 (95% CI = 0.3; 2.9) in HIV-infected patients with peripheral CD4+ T cell count of between 200-500 cells mm-3 and 2.7 (95% CI = 0.9; 7.7) in patients with less than 200 cells mm-3. OR for an intra-oral lesion was 0.4 (95% CI = 0.2; 0.9) for patients with peripheral total lymphocyte counts of between 1000-2000 cells mm-3 and 0.9 (95 CI = 0.4; 2.0) for patients with less than 1000 cells mm-3. CONCLUSION: The association of oral lesions with the clinical stage of HIV infection and to a lesser extent peripheral CD4+ T cell count does suggest that these lesions could be used as additional markers of immunosuppression and AIDS. PMID- 10702789 TI - Tumor-associated glycoprotein 72 (TAG-72) expression in salivary gland neoplasia: an immunohistochemical study using the monoclonal antibody (MAb) CC49. AB - OBJECTIVES: The purpose of this study was to investigate immunohistochemically the expression of tumor-associated glycoprotein 72 (TAG-72) using the monoclonal antibody (MAb) CC49 in salivary gland neoplasia and normal salivary glands in an attempt to determine the potential usefulness of MAb CC49 in diagnostic and therapeutic applications. MATERIALS AND METHODS: Eighty-six specimens (21 benign tumors, 41 malignant, and 24 normal salivary glands), fixed in 10% formalin and embedded in paraffin, were retrieved from the files of the Department of Oral Medicine and Oral Pathology at the Dental School of Aristotle University, Thessaloniki, Greece, and were retrospectively studied with hematoxylin and eosin and with the streptavidin-biotin-complex method using the MAb CC49. RESULTS: Strong immunoreactivity for TAG-72 was observed in salivary duct carcinoma, adenocarcinoma, papillary cystadenocarcinoma, low-grade mucoepidermoid carcinoma, normal submandibular, sublingual, and minor salivary glands. Weak or no immunoreactivity was found in adenoid cystic carcinoma, basal cell adenocarcinoma, polymorphous low-grade adenocarcinoma, and normal parotid gland. CONCLUSIONS: Our results suggest the potential use of MAb CC49 in the differential diagnosis of some salivary gland neoplasms in which their histopathologic features overlap, and in the radiation immunolocalization and immunotherapy of malignant tumors that are localized in the parotid gland. PMID- 10702790 TI - The effect of sodium lauryl sulphate, triclosan and zinc on the permeability of normal oral mucosa. AB - OBJECTIVE: Sodium lauryl sulphate (SLS), an important component in many oral health products, is well established as a contact irritant in skin. Recent studies have suggested that it may also affect the structural integrity of oral mucosa. SLS is rarely used alone in dentifrices or mouthwashes and the aim of this study was to establish the effect of SLS both alone and in combination with Triclosan (TCN) and zinc (Zn) on the permeability barrier properties of normal human oral mucosa. METHOD: Ventral tongue mucosa was obtained from nine males and seven females within 60 h of death and stored frozen at -70 degrees C until use. The permeability of the tissue to tritiated water was measured after pretreatment for 15 min with SLS alone, SLS/TCN, SLS/Zn and a SLS/TCN/Zn mixture. Treatment with distilled water (DW) served as control. The histological appearance of the tissue before and after treatment was also examined by light microscopy. RESULTS: SLS treatment caused a significant increase in water permeability compared to control tissue (Kp = 11.7 +/- 1.00; 4.96 +/- 0.50 respectively; P < 0.005). Treatment with a SLS/TCN/Zn mixture, however, had no effect on the permeability to water (Kp = 5.5 +/- 0.56). Histological examination revealed that tissue exposed to SLS had a marked disruption of the epithelial surface whilst tissue treated with a SLS/TCN/Zn mixture was indistinguishable from controls. CONCLUSION: Although mucosa exposed to SLS alone showed an increase in permeability to water, the addition of TCN and Zn to SLS appeared to prevent this effect. As SLS is included in some dental products to solubilise compounds such as TCN, its presence may have no effect on the permeability barrier property of oral mucosa. PMID- 10702791 TI - Effects of 0.12% chlorhexidine gluconate on experimental gingivitis in non-human primates: clinical and microbiological alterations. AB - OBJECTIVE: This study examined the efficacy of 0.12% chlorhexidine gluconate (Peridex) to reduce gingival inflammation in the absence of mechanical hygiene and its effect on the oral microbial ecology in a non-human primate (NhP) model of gingivitis. DESIGN: Twelve NhP were stratified based on existing inflammation into two groups of six NhP per group. Oral hygiene was performed on both groups so as to reach a level of gingival health (BOP < or = 0.3) at the conclusion of the hygiene phase. One group received 30 ml of 0.12% chlorhexidine gluconate twice daily 7 days/week, and a second group received 30 ml of placebo (distilled water colored to match the active) using the same regimen for 10 weeks. MEASUREMENT OUTCOMES: Clinical parameters including plaque (PLI), pocket depth (PD), attachment level (AL), and bleeding on probing (BOP) were evaluated at 2 week intervals. Subgingival plaque samples were collected by paper point at 2 week intervals and cultured for predominant cultivable bacteria. RESULTS: By week 2, there was a difference in BOP between the groups, which reached statistical significance by week 4. This difference in BOP was maintained throughout the course of the study. Chlorhexidine gluconate (0.12%) had no significant effect on PLI, PD, or AL; although PD was greater in the placebo group after week 2 and throughout the study. Microbiologically, at week 4, the treatment group had a reduction in total bacterial counts, as well as Gram positive bacteria, and total black pigmented bacteria, compared to the placebo group. However, only the differences in Actinomyces spp. reached significance. Interestingly, when both groups received only one treatment/day on the weekends (i.e., day 6 and 7), an associated loss of statistically significant differences between the two groups was observed. Additional experiments dosing the non-human primates once daily, 5 days/week yielded no significant differences in clinical parameters, including bleeding, when compared with the placebo group. CONCLUSION: Non-human primates provided a model system of gingivitis for testing antimicrobial agent effects on the subgingival ecology and accompanying inflammatory responses. Chlorhexidine gluconate (0.12%), even in the absence of mechanical hygiene, was effective in inhibiting clinical signs of inflammation, associated with alterations in the subgingival microbial ecology, most notably Actinomyces spp. PMID- 10702792 TI - NF-kappa B is required for H-ras oncogene induced abnormal cell proliferation and tumorigenesis. AB - Oncogenic mutations in ras lead to constitutive activation of downstream signaling pathways that modulate the activities of transcription factors. In turn, these factors control the expression of a subset of genes responsible for neoplastic cell transformation. Recent studies suggest that transcription factor NF-kappa B contributes to cell transformation by inhibiting the cell death signal activated by oncogenic Ras. In this study, inhibition of NF-kappa B activity by forced expression of a super-repressor form of I kappa B alpha, the major inhibitor of NF-kappa B, markedly decreased the growth rate, saturation density and tumorigenicity of oncogenic H-Ras transformed rat embryo fibroblasts. Such clonally isolated cells overexpressing I kappa B alpha super-repressor not only were viable but also exhibited no sign of spontaneous apoptosis. Inhibition of NF kappa B in these cells was functionally demonstrated by both the loss of cytokine induced DNA binding activity and a profoundly increased sensitivity to cell death in response to TNF-alpha treatment. In contrast, inhibition of NF-kappa B activity in non-transformed fibroblasts had minimal effect on growth, but rendered the cells resistant to a subsequent transformation by H-ras oncogene. Similar results were also obtained with rat intestinal epithelial cells harboring an inducible ras oncogene. Taken together, these findings suggest that NF-kappa B activity is essential for abnormal cell proliferation and tumorigenicity activated by the ras oncogene and highlight an alternative functional role for NF kappa B in oncogenic Ras-mediated cell transformation that is distinct from its anti-apoptotic activity. Oncogene (2000) 19, 841 - 849. PMID- 10702793 TI - The adenovirus type 5 E1B-55K oncoprotein is a highly active shuttle protein and shuttling is independent of E4orf6, p53 and Mdm2. AB - The E1B-55K and E4orf6 oncoproteins of adenovirus type 5 are involved in the export of viral mRNAs. Previously, it was suggested that a complex composed of E1B-55K and E4orf6 serves as a nucleocytoplasmic transporter for viral mRNAs in which the E4orf6 protein directs both nuclear import and export. We now demonstrate that the E1B-55K protein itself shuttles efficiently in the absence of E4orf6. In addition, E1B-55K trafficking was independent of the defined shuttle proteins Mdm2 or p53, which interacts with E1B-55K. The identified N terminal E1B-55K leucine-rich nuclear-export signal (NES) conferred rapid nuclear export even in a heterologous system in contrast to the postulated E4orf6NES. Interestingly, although shuttling was blocked by inhibitors of the CRM1 mediated export pathway, E1B-55K inhibited neither the activity nor the trafficking of the retroviral shuttle proteins HIV-1 Rev and HTLV-1 Rex. In contrast, Rev or Rex blocked the nuclear export of E1B-55K, most likely by competing for essential export factors. Our results provide new insights into the regulation of the adenovirus mRNA export system and the processes of adenovirus mediated transformation. Oncogene (2000) 19, 850 - 857. PMID- 10702794 TI - The MAP-kinase ERK2 is a specific substrate of the protein tyrosine phosphatase HePTP. AB - HePTP is a tyrosine specific protein phosphatase that is strongly expressed in activated T-cells. It was recently demonstrated that in transfected T-cells HePTP impairs TCR-mediated activation of the MAP-kinase family members ERK2 and p38 and it was suggested that both ERK and p38 MAP-kinases are substrates of HePTP. The HePTP gene has been mapped to human chromosome 1q32.1. Abnormalities in this region are frequently found in various hematopoietic malignancies. HePTP is highly expressed in acute myeloid leukemia and its expression in fibroblasts resulted in transformation. To address a possible involvement of HePTP in hematopoietic malignancies we sought to identify HePTP substrate(s) in leukemic cells. Using substrate trapping mutants we have identified the MAP-kinase ERK2 as a specific target of HePTP in the myelogenous leukemia cell line K562. Tyrosine phosphorylated ERK2, but not ERK1, p38, or JNK1, efficiently bound to catalytically inactive HePTP mutants in which the active site cysteine (HePTP C/S) or the conserved aspartic acid residue (HePTP-D/A) had been exchanged for serine and alanine, respectively. Moreover, the interaction of ERK2 with HePTP trapping mutants was dependent on ERK2 tyrosine phosphorylation, indicating that HePTP is specifically targeted to activated ERK2. Using a deletion mutant of HePTP (HePTP-dLD), in which 14 amino acid residues within the N-terminus are missing, we show that regions outside the catalytic domain are also required for the interaction. Furthermore, overexpression of HePTP in K562 cells and fibroblasts interfered with PMA or growth factor induced MAP-kinase activation and HePTP efficiently dephosphorylated active ERK2 on the tyrosine residue in the activation loop in vitro. Together, these data identify ERK2 as a specific and direct target of HePTP and are consistent with a model in which HePTP negatively regulates ERK2 activity as part of a feedback mechanism. Oncogene (2000) 19, 858 869. PMID- 10702795 TI - HYAL1LUCA-1, a candidate tumor suppressor gene on chromosome 3p21.3, is inactivated in head and neck squamous cell carcinomas by aberrant splicing of pre mRNA. AB - The hyaluronidase first isolated from human plasma, Hyal-1, is expressed in many somatic tissues. The Hyal-1 gene, HYAL1, also known as LUCA-1, maps to chromosome 3p21.3 within a candidate tumor suppressor gene locus defined by homozygous deletions and by functional tumor suppressor activity. Hemizygosity in this region occurs in many malignancies, including squamous cell carcinomas of the head and neck. We have investigated whether cell lines derived from such malignancies expressed Hyal-1 activity, using normal human keratinocytes as controls. Hyal-1 enzyme activity and protein were absent or markedly reduced in six of seven carcinoma cell lines examined. Comparative genomic and fluorescence in situ hybridization identified chromosomal deletions of one allele of HYAL1 in six of seven cell lines. Initial RT - PCR analyses demonstrated marked discrepancies between levels of HYAL1 mRNA and protein. Despite repeated sequence analyses, no mutations were found. However, two species of transcripts were identified when primers were used that included the 5' untranslated region. The predominant mRNA species did not correlate with protein translation and contained a retained intron. A second spliced form lacking this intron was found only in cell lines that produced Hyal-1 protein. Inactivation of HYAL1 in these tumor lines is a result of incomplete splicing of its pre-mRNA that appears to be epigenetic in nature. Oncogene (2000) 19, 870 - 877. PMID- 10702796 TI - Molecular requirements for the effect of neuregulin on cell spreading, motility and colony organization. AB - Neuregulin can trigger morphogenetic signals in cells both in vivo and in culture through the activation of receptors from the ErbB family. We have ectopically expressed various ErbB-receptors in 32D myeloid cells lacking endogenous ErbB proteins, and in CHO cells, which express only ErbB-2. We show here that activation of ErbB-3/ErbB-2 heterodimeric receptors triggers PI3-kinase-dependent lamellipodia formation and spreading, while individual ErbB-receptor homodimers as well as ErbB-3/ErbB-1 heterodimers are much less effective. CHO cells expressing ErB-3/ErbB-2 together with N-cadherin, an adhesion receptor, form epithelioid colonies. Neuregulin activates cell motility leading to transition of these colonies into ring-shaped multicellular arrays, similar to those induced by neuregulin in epithelial cells of different types (Chausovsky et al., 1998). This process requires both PI3-kinase and MAP kinase kinase activity and depends on coordinated changes in the actin- and microtubule-based cytoskeleton. Transactivation of ErbB-2 is not sufficient for the activation of cell motility and ring formation, and the C-terminal domain of ErbB-3 bearing the docking sites for the p85 subunit of PI3-kinase is essential for these morphogenetic effects. Thus, ErbB-3 in conjunction with ErbB-2 mediates, via its C-terminal domain, cytoskeletal and adhesion alterations which activate cell spreading and motility, leading to the formation of complex structures such as multicellular rings. Oncogene (2000) 19, 878 - 888. PMID- 10702797 TI - Cooperative interaction between mutant p53 and des(1-3)IGF-I accelerates mammary tumorigenesis. AB - Mammary tumorigenesis was analysed in transgenic mice which overexpress des(1 3)hIGF-I (WAP-DES) and/or a mutant form of p53 (p53172R-H). Nonlactating, multiparous WAP-DES mice exhibited hyperplastic lesions termed mammary interepithelial neoplasia (MIN) which constitutively expressed WAP-DES. By 23 months of age, 53% of the WAP-DES mice developed mammary adenocarcinomas. A 75% reduction in both apoptosis and proliferation was observed in the normal mammary glands of WAP-DES mice. Mammary tumor incidence in WAP-DES/p53 bitransgenic mice was similar to that of WAP-DES and 2 - 3-fold greater than that of nontransgenic and p53172R-H females. Tumor latency, however, was reduced by 8 months in bitransgenic mice as compared to mice of the other three genotypes. Aneuploidy was frequently observed in tumors from bitransgenic and p53172R-H mice, but not from mice expressing only the WAP-DES transgene. Expression of IGFBP3 was elevated in tumors from WAP-DES, but not bitransgenic mice, indicating an alteration in the p53/IGF-I axis. These studies indicate that overexpression of des(1-3)hIGF-I increases the frequency of MIN and stochastic mammary tumors and that the appearance of tumors displaying genomic instability is accelerated by mutant p53172R-H. Oncogene (2000) 19, 889 - 898. PMID- 10702798 TI - Initiation of Apaf-1 translation by internal ribosome entry. AB - The apoptotic protease activating factor (Apaf-1) plays a central role in apoptosis: interaction of this protein with procaspase-9 leads to cleavage and activation of this initiator caspase. In common with other mRNAs whose protein products have a major regulatory function, the 5' untranslated region (UTR) of Apaf-1 is long, G-C rich and has the potential to form secondary structure. We have shown that the 5' UTR of Apaf-1 contains an internal ribosome entry segment, located in a 233 nucleotide region towards the 3' end of the leader, and that the translation initiation of this mRNA occurs only by internal ribosome entry. The Apaf-1 IRES is active in almost all human cell types tested, including Human cervical carcinoma (HeLa), Human liver carcinoma (HepG2), Human breast carcinoma (MCF7), Human embryonic kidney (HK293), African Green Monkey kidney (COS7) and Human lung (MRC5). The Apaf-1 IRES initiates translation as efficiently as the HRV IRES, but is less active than the c-myc IRES. We propose that the Apaf-1 IRES ensures that a constant cellular level of Apaf-1 protein is maintained even under conditions where cap-dependent translation is compromised. Oncogene (2000) 19, 899 - 905. PMID- 10702799 TI - The ETV6-NTRK3 gene fusion encodes a chimeric protein tyrosine kinase that transforms NIH3T3 cells. AB - The congenital fibrosarcoma t(12;15)(p13;q25) rearrangement splices the ETV6 (TEL) gene on chromosome 12p13 in frame with the NTRK3 (TRKC) neurotrophin-3 receptor gene on chromosome 15q25. Resultant ETV6-NTRK3 fusion transcripts encode the helix - loop - helix (HLH) dimerization domain of ETV6 fused to the protein tyrosine kinase (PTK) domain of NTRK3. We show here that ETV6-NTRK3 homodimerizes and is capable of forming heterodimers with wild-type ETV6. Moreover, ETV6-NTRK3 has PTK activity and is autophosphorylated on tyrosine residues. To determine if the fusion protein has transforming activity, NIH3T3 cells were infected with recombinant retroviral vectors carrying the full-length ETV6-NTRK3 cDNA. These cells exhibited a transformed phenotype, grew macroscopic colonies in soft agar, and formed tumors in severe combined immunodeficient (SCID) mice. We hypothesize that chimeric proteins mediate transformation by dysregulating NTRK3 signal transduction pathways via ligand-independent dimerization and PTK activation. To test this hypothesis, we expressed a series of ETV6-NTRK3 mutants in NIH3T3 cells and assessed their transformation activities. Deletion of the ETV6 HLH domain abolished dimer formation with either ETV6 or ETV6-NTRK3, and cells expressing this mutant protein were morphologically non-transformed and failed to grow in soft agar. An ATP-binding mutant failed to autophosphorylate and completely lacked transformation activity. Mutants of the three NTRK3 PTK activation-loop tyrosines had variable PTK activity but had limited to absent transformation activity. Of a series of signaling molecules well known to bind to wild-type NTRK3, only phospholipase-Cgamma (PLCgamma) associated with ETV6-NTRK3. However, a PTK active mutant unable to bind PLCgamma did not show defects in transformation activity. Our studies confirm that ETV6-NTRK3 is a transforming protein that requires both an intact dimerization domain and a functional PTK domain for transformation activity. Oncogene (2000) 19, 906 - 915. PMID- 10702800 TI - DNA damage-related gene expression as biomarkers to assess cellular response after gamma irradiation of a human lymphoblastoid cell line. AB - Since defects in molecular mechanisms controlling DNA repair, cell cycle checkpoint and apoptosis could modify cellular sensitivity to DNA damaging agents, we have conducted a multiparametric molecular analysis for better understanding the regulation pathways leading to cell survival or cell death after irradiation. Using a human lymphoblastoid cell line, we have analysed, following gamma irradiation (0.5, 1, 2, 4, 8, 16 and 32 Gy, at 0.5, 24, 48 and 72 h after treatment), the correlation between proliferation, cell cycle analysis, apoptosis and micronuclei frequency with the expression of TP53, WAF1, DNA LIGASE 1, PCNA, BAX, BLC-2, BAK, DAD1, ICH1-Long and -Short forms mRNAs. We have found that whereas TP53, BAK, ICH1-Short form, and DAD1 were expressed at constant levels, WAF1, PCNA, BAX were up-regulated, ICH1-Long form, DNA LIGASE 1, and BCL 2 were down-regulated. These modifications of expression were significantly correlated with doses, survival, proliferation, cell cycle delays, and apoptosis. A positive correlation of WAF1 and BAX, and a borderline negative correlation with BCL-2 expressions were observed with micronuclei frequency for doses ranging from 0.5 to 4 Gy. In conclusion, our data clearly demonstrate that gene expression profiling, which is easier and more rapid to conduct than the assessments of classical phenotypic responses, could be useful to improve knowledge concerning pathways involved in cellular response to irradiation, knowing that such biomarkers could constitute tools to assess radio sensitivity/radio-resistance. Oncogene (2000) 19, 916 - 923. PMID- 10702801 TI - Ras signaling through PI3K confers hormone-independent proliferation that is compatible with differentiation. AB - Hormones are specialized mitogens that stimulate proliferation in their differentiated target cells. Thyrotropin (TSH), the physiologic regulator of thyroid cells, stimulates cAMP-mediated proliferation and thyroid-specific gene expression. The mitogenic effects of TSH require Ras, therefore Ras activation should be compatible with the maintenance of thyroid differentiation. However, expression of activated Ras extinguishes the differentiated phenotype of thyroid cells. One explanation for this apparent paradox is the selective utilization of Ras effector pathways. We tested the hypothesis that Ras signaling through PI3K mediates the mitogenic effects of TSH in cells which retain their differentiated character. Expression of a Ras effector mutant (RasV12S35) that signals preferentially through Raf-1, although sufficient to confer TSH-independent proliferation, abolished hormone-regulated expression of thyroglobulin and the sodium/iodide symporter. In contrast, expression of a Ras mutant (RasV12C40) that binds selectively to PI3K conferred TSH-independent proliferation without marked effects on thyroid-specific gene expression. Unlike the inhibitory effects of TSH on the proliferation of RasV12S35-expressing cells, TSH enhanced RasV12C40 stimulated proliferation by further increasing the activity of p70s6k, an important mediator of the mitogenic effects of TSH and RasV12C40. These results demonstrate that channeling Ras-dependent signals to PI3K confers TSH with the ability to stimulate proliferation in differentiated cells. Oncogene (2000) 19, 924 - 932. PMID- 10702802 TI - Role of TRAF2/GCK in melanoma sensitivity to UV-induced apoptosis. AB - Radiation resistance is a hallmark of human melanoma, and yet mechanisms underlying this resistance are not well understood. We recently established the role of ATF2 in this process, suggesting that stress kinases, which contribute to regulation of ATF2 stability and activity, play an important role in the acquisition of such resistance. Here we demonstrate that changes in the expression and respective activities of TRAF2/GCK occur during melanoma development and regulate its sensitivity to UV-induced apoptosis. Comparing early and late-stage melanoma cells revealed low expression of TRAF2 and GCK in early stage melanoma, which coincided with poor resistance to UV-induced, TNF-mediated apoptosis; forced expression of GCK alone or in combination with TRAF2 efficiently increased JNK and NF-kappaB activities, which coincided with increased protection against apoptosis. Conversely, forced expression of the dominant negative form of TRAF2 or GCK in late-stage melanoma cells reduced NF kappaB activity and decreased Fas expression, resulting in a lower degree of UV induced, Fas-mediated cell death. Our results illustrate a mechanism in which protection from, or promotion of, UV-induced melanoma cell death depends on the nature of the apoptotic cascade (TNF or Fas) and on the availability of TRAF2/GCK, whose expression increases during melanoma progression. Oncogene (2000) 19, 933 - 942. PMID- 10702803 TI - DNA methylator and mismatch repair phenotypes are not mutually exclusive in colorectal cancer cell lines. AB - A potential link between DNA repair and de novo methylation of exogenous sequences in colorectal cancer cell lines suggested that cells deficient in mismatch repair (MMR-) had an increased ability to silence the introduced virus promoter by DNA methylation due to the presence of a methylator phenotype (MET+) (Lengauer et al., 1997a). We explored this relationship in more detail and found that although there was a clear difference in the abilities of MMR+ cells to express the viral promoter compared to their MMR- counterparts, this difference was not consistently explained by levels of methylation in the viral promoter. Furthermore, we were unable to distinguish differences between the levels of methylation of six endogenous known CpG islands or 100 random DNA fragments containing CCGG sites within the cells. No consistent differences between the abilities of the cells to methylate the CpG island in exon 2 of the p16 gene were observed after transient demethylation by 5-aza-2'-deoxycytidine nor in the levels of expression of three human methyltransferase enzymes. Our results do not therefore support the existence of mutually exclusive DNA methylation (MET) and DNA repair (MMR) phenotypes. Oncogene (2000) 19, 943 - 952. PMID- 10702804 TI - Dominant action of mutated erythropoietin receptors on differentiation in vitro and erythroleukemia development in vivo. AB - J2E cells produce rapid, fatal erythroleukemias in vivo but still respond to erythropoietin (epo) in vitro by differentiating, proliferating and remaining viable in the absence of serum. Mutant epo receptors were introduced into these cells to determine whether they could influence the different biological responses to epo in vitro and the development of erythroleukemias. Three mutant receptors were used as cytoplasmic truncation mutants Delta257 and Delta321 (above box 1 and below box 2 respectively), and the cytoplasmic point mutant W282R (defective for JAK2 activation). Strikingly, the Delta321 mutation produced a hyper-sensitive response in vitro to epo-induced differentiation and viability, but not to proliferation. In contrast with the Delta321 receptor, the Delta257 and W282R mutants inhibited all biological responses to epo due to impaired JAK2 phosphorylation. Significantly, erythroleukemias took almost twice as long to develop with cells containing the W282R mutation, indicating that JAK2 plays an important role in the emergence of these leukemias. These data demonstrate that mutant epo receptors dominantly altered responses of J2E cells to epo in culture and the development of erythroleukemias. Oncogene (2000) 19, 953 - 960. PMID- 10702805 TI - A role for E2F1 in Ras activation of p21(WAF1/CIP1) transcription. AB - We recently reported that E2F1 could transactivate the p21 promoter via cis acting elements between -119 to +16 bp of the p21 gene. Here we show that activated V12-H-Ras can induce the p21 promoter through the same region of the p21 promoter by a p53-independent mechanism in NIH3T3 cells. In contrast, activated Ras was not able to induce the p21 promoter in E2F1-/- fibroblasts, suggesting that E2F1 is required for induction of the p21 promoter by activated Ras. Cotransfection of increasing concentrations of dominant negative E2F1 alone, or with dominant negative DP1 into NIH3T3 cells suppressed induction of the p21 promoter by activated Ras. These data suggest that p53-independent induction of the p21 promoter by activated Ras is mediated at least in part by E2F1. Oncogene (2000) 19, 961 - 964. PMID- 10702806 TI - APC resistance, oral contraceptive therapy and deep vein thrombosis: settled and unsettled problems. PMID- 10702807 TI - Refining prognosis of acute myeloid leukemia patients. PMID- 10702808 TI - Co-existence of hereditary spherocytosis and a new red cell pyruvate kinase variant: PK mallorca. AB - BACKGROUND AND OBJECTIVE: A partial red blood cell (RBC) pyruvate-kinase (PK-R) deficiency was found in a patient with concomitant hereditary spherocytosis (HS) and chronic hemolytic anemia. Clinical, biological and molecular studies were performed in the patient, his parents and a brother, in order to characterize the specific PK-R gene mutation and the inheritance mechanism of the transmission of both red cell defects in this particular family. DESIGN AND METHODS: Conventional biological studies were used to identify the PK-LR gene mutation responsible for hereditary transmission of PK-R deficiency and HS. The family study was completed with genotypic and RBC membrane protein analyses in the patient and his family. RESULTS: Molecular study of the PK deficiency was performed in all the family members and demonstrated a heterozygous condition for the 1516 G->A (506Val->Ile) mutation at the PK-LR gene in both the patient and his mother. Since this mutation has not been reported previously, it is provisionally named PK "Mallorca". The study of RBC membrane proteins demonstrated the existence of partial band 3 and protein 4.2 deficiencies in the propositus and his father but not in the mother and brother, who were also studied. These results support the dominant mode of inheritance of HS and PK-LR gene in this family. INTERPRETATION AND CONCLUSIONS: HS and PK deficiency are not exceptional in Spain. The co existence of both RBC defects in the same patient, however, is very rare; only a few cases have been described to date. Our findings suggest that performing an elementary RBC enzyme survey in all patients with HS would help to determine the real frequency of this apparently rare association. PMID- 10702809 TI - Blood dyscrasias in clozapine-treated patients in Italy. AB - BACKGROUND AND OBJECTIVE: Clozapine is a dibenzodiazepine derivative that is more effective than standard neuroleptic drugs in refractory schizophrenic patients, but its introduction in some countries was delayed by its propensity to cause blood dyscrasias. However, over the last ten years, different reports have clearly demonstrated that agranulocytosis and neutropenia can be easily prevented by means of strict hematologic surveillance. This article reviews the results of the first five years of the Italian Clozapine Monitoring System (ICLOS). DESIGN AND METHODS: The hematologic parameters of 2,404 patients registered between 1995 and 1999 were collected in a central database, before the patients began clozapine-treatment, weekly for the first 18 weeks, and then monthly throughout the duration of therapy. On the basis of conventional criteria, different risk levels have been identified with total leukocyte <3. 0x10(9)/L and/or an absolute neutrophil count <1.5x10(9)/L leading to immediate discontinuation of the drug. RESULTS: The analysis shows that 0.9% of the patients developed neutropenia and 0.7% agranulocytosis, mainly during the first 18 weeks of clozapine treatment. Drug discontinuation led to the normalization of hematologic parameters in all cases, and the use of growth factors reduced the risk of infectious complications. Transient leukocytosis and eosinophilia were also observed but these did not have any serious clinical effects. INTERPRETATION AND CONCLUSIONS: The ICLOS study confirms that regular hematologic monitoring is highly effective in minimizing the incidence of clozapine-associated blood dyscrasias. The lower than initially expected rates of agranulocytosis and associated deaths are encouraging in view of the benefits of this drug in treatment-resistant schizophrenia and other neurologic disorders. PMID- 10702810 TI - Cellular redox state and its relationship to the inhibition of clonal cell growth and the induction of apoptosis during all-trans retinoic acid exposure in acute myeloblastic leukemia cells. AB - BACKGROUND AND OBJECTIVE: All-trans retinoic acid (ATRA) induces growth arrest and apoptosis in acute myeloblastic leukemia (AML) cells. Since cellular redox state regulates these events, we were interested in studying whether it has any role in the responsiveness of AML cells to ATRA. DESIGN AND METHODS: Two human AML cell lines, the ATRA-sensitive OU-AML-3, and the ATRA-resistant OU-AML-7, were used as models. Clonogenic cell culture assay, annexin V method, and measurement of mitochondrial membrane potential were used for the determination of cell growth and apoptosis. Peroxide formation was analyzed by flow cytometry, glutathione and g-glutamylcysteine synthetase (g-GCS) activity was determined spectrophotometrically, and the expression of manganese superoxide dismutase (MnSOD) by Western blotting. RESULTS: ATRA inhibited clonogenic cell growth and induced apoptosis particularly in OU-AML-3 cells. The OU-AML-7 cells had a higher basal level of glutathione and g-GCS activity than the OU-AML-3 cells. ATRA enhanced the generation of peroxides after 24h exposure, which was more prominent in the sensitive than the resistant cell line and was not preventable by N-acetyl L-cysteine. ATRA also increased the activity of g-GCS, which was associated with increased intracellular glutathione in the resistant cell line, while the glutathione level was maintained in the sensitive cell line. During ATRA exposure, MnSOD was induced in the sensitive cell line, but not until after 72 h. Buthionine sulfoximine significantly increased the inhibitory effect of ATRA on colony formation in both cell lines, but only marginally enhanced the effect of ATRA on the induction of apoptosis. INTERPRETATION AND CONCLUSIONS: The balance between oxidative and antioxidative actions of ATRA, as well as the basal redox state of the cells seem to have a definite influence on the responsiveness of AML cells to ATRA. PMID- 10702811 TI - Identification of a group of AML/MDS patients with a relatively favorable prognosis who have chromosome 5 and/or 7 abnormalities. AB - BACKGROUND AND OBJECTIVE: Patients with AML, RAEB-t, or RAEB and abnormalities involving chromosomes 5 and/or 7 (-5, -7) generally, but not always, have poorer prognoses than patients with a normal karyotype. Our objective was to see whether the occasional relatively favorable outcome in -5/-7 patients is a random event or, rather, reflects true heterogeneity in -5/-7. DESIGN AND METHODS: We examined 3 factors known to be prognostic in AML for their prognostic significance in 400 5/-7 patients treated at the M.D. Anderson Cancer Center from 1980-1998 for AML or MDS. The outcome of comparative interest was survival as assessed by log-rank test. RESULTS: There was evidence that outcome was better in -5/-7 patients with a simple (rather than complex) karyotype, with > 1 normal metaphase (rather than only metaphases containing -5/-7), and without an antecedent hematologic disorder. More importantly, the 10% of the patients with a simple karyotype, > 1 normal metaphase, and no antecedent hematologic disorder not only had a better outcome than the other -5/-7 patients but had essentially identical outcomes to the 669 AML/MDS patients with a normal karyotype treated at M.D. Anderson during the same period. INTERPRETATION AND CONCLUSIONS: The results indicate that the 5/-7 group should not a priori be regarded as having an unfavorable prognosis, and more generally suggest the need to refine prognosis within each of the cytogenetic subsets of AML. PMID- 10702812 TI - Chromosomal aberrations evaluated by CGH, FISH and GTG-banding in a case of AIDS related Burkitt's lymphoma. AB - BACKGROUND AND OBJECTIVE: We have previously reported on a complex chromosome rearrangement [der(17)] in a B-cell line, BRG A, established from an AIDS patient with Burkitt's lymphoma (BL). The aim of the present study was the definition of der(17) composition and the identification of complete or partial chromosome gains and losses in two cell clones (BRG A and BRG M) derived from this patient. DESIGN AND METHODS: We applied comparative genome hybridization (CGH) to detect the DNA misrepresentations in the genome of the two cell clones. Findings from CGH and banding analysis could then direct the choice of probes for chromosome painting experiments to elucidate der(17) composition. RESULTS: CGH analysis identified gains of chromosomes 1q, 7q, 12q, 13q, 15q, 17p, 20p,q and losses of chromosomes 3p and 5q in BRG A and gain of chromosome 1q and loss in chromosome 6q in BRG M. Some of the detected alterations had already been described in lymphomas, while others appeared to be new. The combination of these techniques allowed a precise definition of der(17), composed by translocated regions from chromosomes 12 and 15. INTERPRETATION AND CONCLUSIONS: We demonstrated CGH to be a powerful tool in the identification of recurrent chromosome aberrations in an AIDS-related BL and in ascertaining the origin of marker chromosomes. We were also able to identify a different pattern of aberrations and assess an independent sequence of events leading to the 1p gain in the two subclones. PMID- 10702813 TI - Late Epstein-Barr virus infection of a hepatosplenic gamma delta T-cell lymphoma arising in a kidney transplant recipient. AB - BACKGROUND AND OBJECTIVE: gd T-cell lymphomas are only exceptionally observed in transplanted patients. Aim of this study was the detailed characterization of one such case. DESIGN AND METHODS: The patient developed spontaneous splenic rupture six years after kidney transplantation. The splenic red pulp was infiltrated by medium-sized and large lymphoid cells with two or more nucleoli. At autopsy, similar lymphoid cells infiltrated the hepatic sinusoids. Histologic, immunologic and molecular studies were carried out. RESULTS: By immunohistochemistry, the atypical lymphoid cells were found to express CD3, CD45 and CD43, indicating their T-lineage origin. Approximately 99% of spleen mononuclear cells (MNC) were CD3(+), gammadelta TcR+, CD4-, CD8-, alphabeta TcR-. A clonal gammadelta TcR rearrangement (Vgamma1-Jgamma1.3/2.3-Cgamma2; Vdelta1-Ddelta2-Jdelta1) was detected. The final diagnosis was peripheral T-cell lymphoma, hepato-splenic gammadelta-type. EBV infection of spleen MNC was documented by molecular studies. However, in situ hybridization for EBER-1 (EBV-RNA) showed that only a minority of malignant lymphoid cells (5-7%) were EBV-infected. INTERPRETATION AND CONCLUSIONS: It is concluded that EBV infection was as a late event involving an already transformed gd T-cell clone. PMID- 10702814 TI - A variant of ProMACE-CytaBOM chemotherapy for non-Hodgkin's lymphoma with threefold higher drug dose size but identical cumulative dose intensity. A pilot study of the Italian lymphoma study group (GISL). AB - BACKGROUND AND OBJECTIVE: The positive results of high-dose chemotherapy followed by rescue with bone marrow progenitor cell transplantation are generally ascribed to the high dose size (DS) of the drugs given. However, a concomitant marked increase in dose intensity (DI) is always involved. With the aim of comparing the role of DS and DI in non-Hodgkin's lymphomas, a variant of Fisher's ProMACE CytaBOM regimen was designed in which the projected cumulative drug DIs remained the same as in the original schedule but the DSs were tripled. DESIGN AND METHODS: Dosages in mg/m(2), route and days of administration were the following: cyclophosphamide 1,950 i.v. on days 1, 64; methotrexate 360 i.v. days 15, 78; vincristine 1.4 iv days 15, 78, 43, 106; etoposide 360 i.v. days 29, 92; epirubicin 120 i.v. days 29, 92; bleomycin 15 i.v. days 43, 106; cytarabine 900 i.v. days 50, 113. Thirty-six outpatients with intermediate- and high-grade non Hodgkin's lymphomas entered the pilot study; 29 were untreated and 7 had relapse disease. Clinical stage was I in 1 patient, II in 7, III in 5 and IV in 23; 10 had B symptoms; the IPI score was 0-2 in 29 cases and > or =3 in the remaining 7. RESULTS: Of the 29 previously untreated patients, 16 achieved complete remission, 8 partial remission, 4 developed progressive disease and 1 was withdrawn early from the study because of acute viral hepatitis; subsequently 4 relapsed and 3 died (2 of disease progression, 1 of causes unrelated to the disease). In the pre treated group 3 patients obtained complete remission, 2 partial remission and in 1 patient the disease progressed; 3 of these pre-treated patients died (1 of progressive disease, 1 of a new relapse, 1 of myocardial infarction during therapy). With a 20-month median follow-up, the 30-month overall and relapse-free survival were 0.58 and 0.70, respectively. G-CSF was administered to all but 2 patients, with median delivery throughout the whole regimen of 8, 400 microg per patient. Actual cumulative DI was 0.82+/-0.11. Grade 3-4 hematologic toxicity consisted of anemia in 3 cases, of leukopenia in 8 and of thrombocytopenia in 2; the same grade of non-hematologic toxicity involved the liver in 2 cases, the heart in 1 (the above mentioned death), the digestive mucosa in 2 and the peripheral nerves in 1 patient. INTERPRETATION AND CONCLUSIONS: The iso-DI sequential variant of the ProMACE-CytaBOM regimen can be considered feasibile, relatively non-toxic, and can be given on an out-patient basis. Limited use of G CSF is required (about 3 vials after each drug administration). Thus, a randomized trial with the original ProMACE-CytaBOM regimen can be designed. PMID- 10702815 TI - High-dose therapy in multiple myeloma: effect of positive selection of CD34+ peripheral blood stem cells on hematologic engraftment and clinical outcome. AB - BACKGROUND AND OBJECTIVE: Positive selection of peripheral blood stem cells (PBSC) has been investigated in multiple myeloma (MM) with the aims of reducing plasma cell (PC) contamination of the leukaphereses and improving clinical outcome of autografted patients. DESIGN AND METHODS: In our center 39 untreated patients with stage II and III MM, younger than 65 years, started high-dose therapy consisting of 4 VAD cycles, collection of PBSC mobilized by 7 g/m(2) cyclophosphamide + G-CSF, and myeloablative treatment with 12 mg/kg busulfan plus 120 mg/m(2) melphalan. The leukaphereses from 23/39 patients (59%) were processed for positive selection of CD34(+) cells using an avidin-biotin immunoaffinity device. RESULTS: A reduction of PC contamination of as much as 2 log was found in the post-selection products by a flow-cytometric technique using the monoclonal antibody CD 138 alternatively coupled with CD38 and cytoplasmatic k or l light chains in separate samples. Hematologic reconstitution and clinical outcome of the 23 patients reinfused with selected CD34(+) cells (SEL group) were compared with those of the 16 patients reinfused with unselected cells (UNSEL group). No significant differences were observed between the 2 groups with regards to the median duration of neutropenia and thrombocytopenia, the hematologic support required, the incidence of febrile episodes and bacteremias. At a median follow up of 18 months (range 5-34) after ASCT, there were 7/23 (32%) continuous complete remissions (CR) in the SEL group and 4/16 (25%) in the UNSEL group; there were 10/23 (44%) continuous partial remissions (PR) and 5/16 (31%) in the SEL and UNSEL groups, respectively. Two patients in the UNSEL group and one patient in the SEL group died of progressive disease. INTERPRETATION AND CONCLUSIONS: Our data show that positive selection allows rapid engraftment of hematopoiesis and low morbidity. Although no significant difference was detected between the two groups in the frequency of CR and PR 3 and 18 months after ASCT, a longer follow-up is needed to evaluate definitively the effect of CD34(+) selection on the clinical outcome after ASCT. PMID- 10702816 TI - Comparison of two different time interval protocols for central venous catheter dressing in bone marrow transplant patients: results of a randomized, multicenter study. The Italian Nurse Bone Marrow Transplant Group (GITMO). AB - BACKGROUND AND OBJECTIVE: Care of central venous catheter (CVC) in patients undergoing bone marrow transplantation (BMT) raises significant problems related to the high risk of local infections due to the immunodeficient status, which in itself is a predisposing factor for systemic blood-stream infections. Although frequent changes of CVC dressing might theoretically reduce the incidence of infections, they are also accompanied by significant skin toxicity and patient discomfort. No study has yet addressed these points. The objective of this study was to compare two different time interval protocols for CVC dressing in order to assess the effects on local infections and toxicity. DESIGN AND METHODS: In a multicenter study, 399 bone marrow transplant (BMT) patients with a tunneled CVC (Group A, 230 pts) or a non-tunneled one (Group B, 169 pts) were randomly allocated to receive CVC dressing changes every 5 or 10 days, if belonging to Group A, or 2 or 5 days, if in Group B. Transparent, impermeable polyurethane dressings were used for all patients. The rate of local infections at the site of CVC insertion was assessed by microbiological assays every 10 days, while the severity of skin toxicity was measured according to the ECOG scale. RESULTS: Sixty-five per cent of enrolled patients were finally evaluable. Patients (in both Groups) receiving CVC dressing changes at longer intervals did not show a significant increase in the rate of local infections, while those who received dressing every 2 days had a significant increase in local skin toxicity. Longer intervals were accompanied by a reduction in costs. INTERPRETATION AND CONCLUSIONS: The results of this study demonstrate that the increase in time interval between CVC dressing changes in BMT patients did not raise the risk of local infections, while significantly reducing patient discomfort and costs. PMID- 10702817 TI - Thrombin facilitates primary platelet adhesion onto vascular surfaces in the absence of plasma adhesive proteins: studies under flow conditions. AB - BACKGROUND AND OBJECTIVE: The effect of local and circulating thrombin on platelet adhesion onto vascular surfaces was explored in the absence of plasma adhesive proteins using flow conditions. DESIGN AND METHODS: To study the local effects of thrombin, denuded rabbit aorta segments were incubated with thrombin concentrations of 0.001, 0.01 and 0.1 U/mL. To evaluate the effects of circulating thrombin, the same concentrations were added to perfusates consisting of washed platelets and washed red blood cells suspended in a human albumin solution (5%). In some experiments, purified von Willebrand's factor (vWF) (Haemate-P) was added to the perfusates (0. 8 U/mL of vWF, final concentration). A humanized chimeric antibody to the GPIIb-IIIa complex (Reopro) was used to determine the role of this glycoprotein on platelet adhesion under the conditions described. The effect of blocking GPIb was also assessed. Perfusions were carried out at 800 s(-1) for 10 min. The interaction of platelet with the vessel surface was morphometrically evaluated and expressed as percentage of surface coverage (%SC). Changes in the surface expression of the major platelet antigens were also analyzed by flow cytometry. RESULTS: Incubation of subendothelial surfaces with thrombin enhanced platelet deposition with respect to control levels (increases in SC of 64%, 79% and 86% with 0.001, 0.01 and 0.1 U/mL of thrombin, respectively). Low concentrations of thrombin (0.001 and 0.01 U/mL) incorporated in the perfusates resulted in a similar pro-adhesive effect (increases in SC of 64% and 71%, respectively) while the highest concentration (0.1 U/mL) failed to produce a pro-adhesive effect due to the augmented formation of platelet aggregates with subsequent thrombocytopenia (15+/-1 vs. 160+/-5x10(9) plt/L in the perfusates). Similar results were obtained when VWF was present in the perfusate. Reduction of platelet deposition by blockade of GPIIb-IIIa (to 5.3+/ 0.7%) was partially restored by thrombin. Blockade of GPIb prevented platelets from adhering even when thrombin was present (%SC of 2.0+/-0.8%). No significant changes in the distribution of platelet membrane glycoproteins during perfusion experiments were detected. INTERPRETATION AND CONCLUSIONS: Our results suggest that thrombin facilitates primary platelet adhesion onto vascular surfaces even in the absence of plasma adhesive proteins. This effect seems to be mainly dependent on the GPIb/vWF axis. PMID- 10702818 TI - Clotting alterations in primary systemic amyloidosis. AB - BACKGROUND AND OBJECTIVE: The bleeding manifestations frequently observed in patients with immunoglobulin light chain amyloidosis (AL) have been attributed to different pathogenetic factors: amyloid deposits in several organs and systems leading to failures of these latter, the affinity of amyloid for some clotting factors, and the presence of plasma components interfering with fibrin formation could all induce alterations of clotting tests. This investigation was aimed at defining the prevalence of clotting abnormalities and their clinical manifestations in patients with AL. DESIGN AND METHODS: Thirty-six consecutive patients with biopsy proven amyloidosis and documented monoclonal gammapathy were enrolled within one year. The following clotting tests were considered in the study: activated partial thromboplastin time (aPTT), prothrombin time (PT), thrombin time (TT), reptilase time (RT), Russell's viper venom time (RVTT), fibrinogen, factor X and alpha-2 antiplasmin. RESULTS: Hemorrhagic manifestations were mild to moderate in nine patients, but severe and untractable in one. The most frequent clotting anomaly was defective fibrinogen conversion to fibrin, as demonstrated by prolongation of both TT (85% of cases) and RT (90% of cases). Low levels of factor X activity were observed in about 1 out of 4 samples, while fibrinogen and alpha2 antiplasmin levels were distributed over a wide range of values. PT was prolonged in 8 and aPTT in 25 patients. The search for lupus anticoagulant was negative in samples showing a prolongation of aPTT and/or RVVT. INTERPRETATION AND CONCLUSIONS: The prolongation of TT and RT is not dependent on either the presence of a heparin-like substance in the plasma or on fibrinogen levels; furthermore, the prolongation of RVVT is not related to factor X level. The hypothesized presence in the plasma of an inhibitor of fibrin formation could also affect factor X activation by Russell viper venom. The prolongation of TT and RT represents a peculiar feature of amyloidosis. The variability in the behavior of the other clotting times and hemostatic factors studied is mirrored in the heterogeneity of the clinical features observed in this disease. PMID- 10702819 TI - The FXIII Val34Leu polymorphism in venous and arterial thromboembolism. AB - BACKGROUND AND OBJECTIVE: Several hereditary disorders affecting coagulation factors have been identified as prothrombotic risk factors. Recently, the common Val34Leu polymorphism of the A-chain factor XIII gene, associated with high factor XIII activity, has been identified as a protective genetic factor against occlusive arterial and venous diseases in British and Finnish populations. The aim of our study was to investigate the role of this polymorphism in arterial and venous thromboembolic disorders in a distinct population. DESIGN AND METHODS: We analyzed the prevalence of this polymorphism in three case/control studies of consecutive patients from the south of Spain diagnosed as having acute coronary syndromes (101), acute cerebrovascular events (104), and deep venous thrombosis (97). RESULTS: No significant differences were detected in the prevalence of genotypes or alleles between patients and controls. INTERPRETATION AND CONCLUSIONS: The Leu 34 allele does not play an important role in the development of thromboembolic episodes in the Spanish population. PMID- 10702820 TI - Eosinophils and C4 predict clinical failure of combination immunotherapy with very low dose subcutaneous interleukin-2 and interferon in renal cell carcinoma patients. AB - BACKGROUND AND OBJECTIVE: The clinical and immunologic activities of interleukin 2 (IL-2) in cancer patients have been extensively studied and described; however, in most of these studies, IL-2 was administered by intravenous bolus or continuous infusion, while the immunologic effects of IL-2 given by the subcutaneous (s.c.) route have not yet been well studied. DESIGN AND METHODS: The present study was aimed at evaluating the effects of IL-2, given at very low doses s.c. to patients with advanced renal cell carcinoma (RCC), on a number of immunologic parameters: number of total lymphocytes, number of CD4-, CD8-, CD25 positive cells, number of natural killer (NK) cells, titers of IL-2 soluble receptor (sIL-2R) and of C4, eosinophils, eosinophilic cationic protein (ECP) and eosinophilic protein X (EPX). Finally, a logistic regression model was performed to identify early immunologic parameters that correlate with a favorable or unfavorable treatment outcome. RESULTS: Independently from the mere report of the changes induced by immunotherapy, the analysis showed that, within the pre treatment model, a large eosinophil number predicts the failure of IL-2 treatment; in contrast, within the post-treatment model, high C4 serum titers and, again, a large number of circulating eosinophils predict immunotherapy failure. INTERPRETATION AND CONCLUSIONS: As far as concerns C4, its negative predictive value could be related to the fact that it is an indirect index of macrophage activation; thus, even though macrophages release substances with antitumor activity, they can also stimulate the release of sIL-2R, which may compete for exogenous IL-2. Some authors have postulated that macrophages may even stimulate tumor cell growth, or impair NK activity. Despite a great amount of uncertainty concerning the role of eosinophils, in our study, blood eosinophilia predicts a poor response to immunotherapy in patients with advanced RCC, thus supporting previous observations from our own group. PMID- 10702821 TI - Low intensity regimens with allogeneic hematopoietic stem cell transplantation as treatment of hematologic neoplasia. AB - Conventional myeloablative conditioning regimens for allografting rely on the use of toxic myeloablative and immunosuppressive therapies to achieve engraftment and control of hematologic neoplasias. Unfortunately, these regimens have resulted in substantial morbidity and mortality. Preclinical and pilot clinical studies have shown that conditioning regimens can be reduced in intensity (resulting in reduced morbidity and mortality) since stem cell allografts can create their own space in the host's bone marrow. Initial promising results with these attenuated conditioning regimens confirm that such an approach is feasible in patients with hematologic neoplasias and genetic diseases ineligible for conventional allografting because of age and/or organ toxicity. The combination of high-dose therapy/autografting followed by a low intensity conditioning regimen (Flu-Cy protocol) and donor mobilized hematopoietic stem cell infusion (mini allografting) may ultimately be useful in advanced resistant hematologic neoplasia. Finally, these initial promising results with attenuated conditioning regimens have been achieved in transplants with HLA-identical siblings. In the future the main goal will be to explore non-toxic conditioning regimens in the context of transplants from related MHC-mismatched or unrelated MHC-matched donors by increasing the patient's immunosuppression. PMID- 10702822 TI - Lymphangioma of the spleen in an elderly patient. AB - Splenic cystic lymphangioma is a very rare condition, and is classified among cystic proliferations of the spleen. It is considered to be the result of a developmental malformation of the lymphatic system and can involve the spleen alone or be a part of multiorgan disease. It is usually seen in children, often found incidentally. We describe a case of cystic lymphangioma of the spleen in an elderly woman putting emphasis on the rarity of the case in old age, and on the problems of differential diagnosis with the other cystic proliferations of the spleen, in particular hydatid disease, in the absence of histologic information. PMID- 10702823 TI - A case of phagocytic multiple myeloma. PMID- 10702824 TI - A Caucasian boy with Gilbert's syndrome heterozygous for the (TA)(8) allele. PMID- 10702825 TI - Molecular characterization of glucose-6-phosphate dehydrogenase deficiency in Turkey. PMID- 10702826 TI - Expression of transmembrane and soluble forms of CD44H in human myeloid cell lines and its regulation by hyaluronic acid. PMID- 10702827 TI - Isolated asymptomatic severe neutropenia as the presentation of myeloid/natural killer cell acute leukemia. PMID- 10702828 TI - Liposome encapsulated daunorubicin (daunoxome) for acute leukemia. PMID- 10702829 TI - Usefulness of the prognostic score for advanced Hodgkin's disease in patients with human immunodeficiency virus-associated Hodgkin's lymphoma. PMID- 10702830 TI - Mini-ICE regimen allows mobilization of peripheral blood progenitor cells in a patient with chronic myelogenous leukemia failing the ICE protocol. PMID- 10702831 TI - Lamivudine for the prevention of hepatitis B virus reactivation during autologous stem cell transplantation. A case report. PMID- 10702832 TI - Effectiveness of extracorporeal photochemotherapy in treating refractory chronic graft-versus-host disease. PMID- 10702833 TI - Delayed graft versus leukemia effect after allogeneic peripheral stem cell transplantation in a patient with chronic lymphocytic leukemia. PMID- 10702834 TI - Molecular biotyping methods for epidemiologic studies of candidemia in patients with acute leukemia. PMID- 10702835 TI - Disseminated toxoplasmosis after CD34+-selected autologous peripheral blood stem cell transplantation. PMID- 10702836 TI - Frequency of Gilbert's syndrome associated with UGTA1 (TA)(7) polymorphism in Southern Italy. PMID- 10702838 TI - Newer vaccines: Like Marie Antoinette said, "let the poor eat cake" - Reply. PMID- 10702837 TI - AFP surveillance Let it not be "Targetoma" - Reply. PMID- 10702839 TI - Diagnostic and therapeutic approaches for masses in the posterior mediastinum. AB - Between January 1980 and December 1997 twenty-one patients with a mass in the posterior mediastinum came under our observation. All of the patients underwent chest radiography, bronchoscopy, respiratory function tests, perfusional and ventilatory radionuclide scans, a computed tomography (CT) of the chest and blood gas analysis. In cases involving neurogenic tumours magnetic resonance imaging (MRI) was used. Ten patients underwent CT guided transthoracic needle biopsy. The excision was performed by means of a thoracotomy in 12 cases (57.1%) and by video assisted thoracoscopy surgery (VATS) in the other 9 (42.9%); no deaths were recorded. Eleven neurilemmomas, two bronchogenic cysts, two paragangliomas, two neuroepitheliomas, one neurogenic sarcoma and three esophageal duplications were found. The authors believe an accurate pre-operative assessment of the lesion can be obtained using CT and MRI. The video assisted thoracoscopy (VAT) is a useful method of diagnosis and treatment as it can be converted into VATS if the lesion is benign or cystic. Thoracotomy is necessary when the mass is malignant or when there is adhesion to or invasion of surrounding tissues. PMID- 10702840 TI - [Cleft palate and dysfunction of the eustachian tube]. AB - Secretory otitis media (SOM) is a frequent complication in infants with cleft palate. In cleft palate the muscles that open the Eustachian tube (tensor palatini and levator palatini) have abnormal connections thereby making the tube opening either difficult or impossible. This will lead to secretory otitis media in 95% of cases, since the middle ear will not be aerated. In this paper, 14 patients operated on for cleft palate during the first year of life were examined. SOM was treated only by medical therapy without the insertion of tympanostomy tubes. Post-operative follow-up ranged from 2 months to 5 years. In all patient SOM was still present at last follow-up with poor efficacy of medical therapy. Therefore, our therapeutic protocol includes myringotomy and insertion of tympanostomy tubes during the first general anesthesia for cleft palate treatment. PMID- 10702841 TI - [The treatment of pathologic calcification of shoulder tendons with E.D.T.A. bisodium salt by mesotherapy]. AB - The Authors treated at the Physiotherapy Service of the Clinic Orthopedic in Parma, 31 patients affected by shoulder's calcific tendonpathie++ were treated with E.D.T.A. bisodium salt they were painful and showed functional restriction besides they all shared a crystal's hydroxyapatite deposition. It was considered the pain by the Scott-Huskisson analogous--visual scale and the radiographic variation of calcification. It was noticed that with minimum pharmacological doses satisfying therapeutical results were achieved. That was not only as far as pain was concerned (disappeared in 29 patients (93.5%), but it was also obtained a significant reduction (4 patients 13%) or in some cases the disappearing of calcifications (25 patients 80%). PMID- 10702842 TI - Endometriosis of the abdominal wall. AB - Endometriosis is ectopic endometrial tissue that responds to hormonal stimulation and is found 8-15 per cent of all menstruating women. Endometrioma in/or close to a surgical scar is rare and occurs in 0.1 per cent of women who underwent cesarean section. When localized at the abdominal wall, the disease presents as a painful swelling resembling other lesions, such as hernias, post-operative ventral hernias, hematomas, granulomas, abscesses, and tumors. Endometriosis of the abdominal wall may not be considered in the differential diagnosis of masses detected in/or close cesarean scar. Three cases are reported here. All of them underwent surgery and the error of the pre-operative diagnosis was revealed by histology in two cases. Actually, only one case was suspected pre-operatively. PMID- 10702843 TI - [Repair of vesicovaginal fistula by the Martius technique]. AB - Vesico-vaginal fistula (VVF) is still an unpleasant complication of female genital system surgery. We report the case of a 57-years-old woman undergone to neoadjuvant radiotherapy and afterwards abdomino-perineal resection who developed after one months a VVF. PMID- 10702845 TI - Racism and mental health into the 21st century: perspectives and parameters. AB - Definitions and theories of racism are reviewed, and the influence of racism on the American mental health system is examined, with special attention to the effects on racial and ethnic minorities of the sociopolitical climate of the 1990s. The aims of this special section are outlined and an overview is presented of the articles, which define some of the key problems of racism and mental health, describe their scope and effects, and propose approaches to remediation as we move into the 21st century. PMID- 10702844 TI - [Ileal conduit: our experience]. AB - In the last 100 year, many progresses have been done in the ileal urinary diversion after cystectomy. Although the interest for the continent urinary diversion has increased enormously in the last ten years, the ileal conduit still represents the golden standard. We report our experience. PMID- 10702846 TI - Racism as a clinical syndrome. AB - This paper examines the clinical effects of racism on its targets and, in particular, on its agents, the individuals who, wittingly or not, partake of the culture of racial privilege. It proposes a paradigm shift in regard to the clinical study of racism, and presents a structural model of racism, analogous to addiction as a disease, which holds that racism has an etiology and a clinical taxonomy that lends itself to differential diagnosis and treatment of those who manifest symptoms. PMID- 10702847 TI - From evil to illness: medicalizing racism. AB - Arguments for treating racism as an illness or an addiction are critiqued, and it is suggested that such efforts constitute a step backward in the battle against racism and discrimination. Medicalization, rather than being a catalyst for social change, is a mode of social control. The assumptions underlying the disease model are examined, and a strategy is outlined for dealing with racism as a structural phenomenon broader and more complex than personal prejudice and individual pathology. PMID- 10702848 TI - Invisibility syndrome: a clinical model of the effects of racism on African American males. AB - Adaptive behavior and psychological well-being of African Americans can be affected by prejudice and discrimination. Encountering repeated racial slights can create "psychological invisibility." The invisibility syndrome is presented as a conceptual model for understanding the inner evaluative processes and adaptive behavior of African Americans in managing experiences of racism. PMID- 10702849 TI - A multidimensional conceptualization of racism-related stress: implications for the well-being of people of color. AB - A conceptualization of racism-related stress and its impact on well-being is offered that integrates existing theory and research on racism, multicultural mental health, and the stress process. The conceptualization is relevant to diverse racial/ethnic groups, considers the larger social and historical context, and incorporates attention to culture-based variables that may mediate the relationship between racism and well-being. Implications for intervention are discussed. PMID- 10702850 TI - Treatment delay among Asian-American patients with severe mental illness. AB - Length of treatment delay and cultural-familial correlates were studied in a group of 62 Asian-American patients with severe mental illness, and 40 of their relatives. Contrary to prior findings of long treatment delay among Asian Americans, this cohort reported relatively low levels of stigma and shame and relatively short delay between onset of psychiatric symptoms and inception of treatment. Higher levels of shame and stigma felt by the relatives were associated with patients' longer treatment delay. PMID- 10702851 TI - Training in cultural schemas: an antidote to unintentional racism in clinical practice. AB - Although recent attempts have been made to bolster multicultural training, problems remain in defining effective training and providing equitable service delivery to minority consumers. This article argues that cultural schema is a useful construct for helping clinicians identify, organize, interpret, and integrate cultural data into clinical practice. Further, it is proposed that training in the use of cultural schemas will serve to also reduce the prevalence of unintentional racism in the mental health field. PMID- 10702852 TI - The long and winding road: personal reflections of an anti-racism trainer. AB - This paper addresses the intense personal and professional preparation necessary for becoming an effective anti-racism trainer. The author draws upon diversity training models and theory, as well as personal experiences and reflections charted as a white man actively involved in this field over the past ten years. PMID- 10702853 TI - Early intervention programs for children with autism: conceptual frameworks for implementation. AB - Four diverse early intervention programs for children with autism--discrete trial training, LEAP, floor time, and TEACCH--are described. For each program, the concepts of learning, development, and autism are summarized, intervention procedures are outlined, and connections between theory and practice are illustrated. Research outcomes for each of the four programs are discussed. PMID- 10702854 TI - Relation of mothers' affective developmental history and parenting behavior: effects on infant medical risk. AB - Mothers of infants with varying degrees of medical risk were grouped according to their perception of acceptance or rejection in childhood. Those who recalled the highest degree of acceptance showed greater warmth and flexibility as parents, regardless of their infants' degree of medical risk. However, infant medical risk was an important moderator in relations between maternal perceptions of childhood rejection and parental behavior. PMID- 10702855 TI - Emotional availability: conceptualization and research findings. AB - The emotional availability construct (based on observations of parent-child interactions) was first reconceptualized for research in 1991 as a way to describe the quality of parent-child interactions. Since then, there has been considerable refinement of the construct. EA refers to several parental dimensions (sensitivity, structuring, non-intrusiveness, non-hostility) and two child dimensions (responsiveness to parent and involvement of parent). The EA empirical link with attachment and parent-child relationship are reviewed and avenues for future research are suggested. PMID- 10702857 TI - Differences among families coping with serious mental illness: a qualitative analysis. AB - Families of 180 people with serious mental illness, representing various socioeconomic and ethnic groups, were interviewed about their understanding of their family member's illness, coping with problems caused by the illness, sources of support, effects of medication and substance abuse, and dealing with mental health professionals. Several significant areas of concern emerged, and are explored with attention to differences based on gender, ethnic group, socioeconomic status, and role of the family member. PMID- 10702856 TI - Preschool children's exposure to violence: relation of behavior problems to parent and child reports. AB - A group of 155 parents and their preschool children attending Head Start reported on the children's exposure to community violence, level of distress symptoms, and behavioral problems. The behavioral correlates of exposure were found to differ according to exposure modality: internalizing problems were more likely in children who witnessed violence, and externalizing problems in those victimized by violence. Issues regarding self-reports by preschool children are highlighted, and clinical and research implications discussed. PMID- 10702858 TI - Impact of welfare reform on teenage parent recipients: an analysis of two cohorts. AB - To assess early effects of welfare reform, administrative data was used to compare prereform and postreform cohorts of teenage parents regarding the impact of reform on welfare enrollments, case closures, child maltreatment, and subsequent births. The relation of mandated living arrangements to outcomes was also examined. Cohort differences were observed in enrollments and reasons for closure, but not in maltreatment or birth rates. Living arrangements were found to be associated with case closure. PMID- 10702859 TI - An in vitro model to evaluate the effect of an organophosphoric agropesticide on cell proliferation in mouse seminiferous tubules. AB - Recently, there has been public concern about the toxic effects of organophosphoric pesticides (OP) upon human and animal populations. Since the seminiferous epithelium is an actively proliferating tissue, it was of interest to study germ cell proliferation in the isolated seminiferous tubules of mice that were cultured in the presence of Parathion or paraoxon, its metabolite. Eighteen 3-month-old CF-1 male mice were used. Paraoxon (PO) and Parathion (PT) were added to cultures of seminiferous tubules in the following groups: (1) Ham F 10 medium pH 7.4 (M) (control); groups 2 to 5, M + decreasing doses of either PO or PT (0.8; 0.4; 0.04; 0.004; 0.0004 mM). Each group consisted of six mice. Incubation of the tubules was carried out for 5 h at 35 degrees C, 5% CO2, 95% air. One hour before the end of incubation 5 microCi of 3H-thymidine was added to the cultures. DNA uptake was measured by scintillation counting. PO and PT at concentrations of 0.8 mM elicited a sharp decrease in testicular DNA synthesis. Recuperation at concentrations under 0.4 mM was different. With PO it was rapid, possibly due to the high detoxificative ability of the testis, which contains high quantities of the enzyme 'type A esterases' that hydrolyses PO. At concentrations lower than PO, PT has an inhibitory effect upon germ cell proliferation, which deserves further research. PMID- 10702860 TI - Chronic immobilization-induced stress increases plasma testosterone and delays testicular maturation in pubertal rats. AB - We investigated whether chronic stress, applied from prepuberty to early puberty, interferes with the spermatogenic and androgenic testicular functions. Male pubertal rats (40 days old) were immobilized 6 h per day for 15 days. Plasma concentrations of corticosterone, prolactin and testosterone were significantly augmented following immobilization, whereas plasma luteinizing hormone decreased and follicle-stimulating hormone was not altered. Acute immobilization (5 min) increased prolactin and testosterone levels in control rats but caused a significantly higher increase in these hormones when superimposed on chronic stress. A lower extent of testicular maturation was observed in pubertal rats immobilized from prepuberty. PMID- 10702861 TI - Clinical findings in congenital absence of the vasa deferentia. AB - In a clinical study, 105 patients with congenital bilateral absence of the vas deferens (CBAVD) and 18 with congenital unilateral absence of the vas deferens (CUAVD) were investigated. CUAVD was observed on the left side in 66%. Renal agenesis was more frequent in CUAVD (73.7%) than in CBAVD (11.8%). The leading signs of CBAVD are low pH level (average 6.5) and low volume of the ejaculate (average 0.95 ml). Testicular biopsies of 52 patients revealed normal spermatogenesis or hypospermatogenesis (33% in CBAVD; 45% in CUAVD). Genetic probing and counselling concerning cystic fibrosis are necessary if extracorporal micro-fertilization is considered. The absence of the vas deferens was often overlooked by the first investigator, the average time until correct diagnosis being 4.3 years. As artificial reproduction technology becomes more common, detection of vasal agenesis will certainly be made earlier and more frequently in the future. In order to assure compatibility of subsequent prospective studies about CBAVD and CUAVD, the following investigations are considered to be necessary: (i) semen analysis (pH, volume); (ii) renal ultrasonography or excretory urogram (screening for renal agenesis); (iii) genetic cystic fibrosis screening. PMID- 10702862 TI - Effect of varicocelectomy on seminal plasma transferrin values: a comparative clinical trial. AB - The possible effects of varicocele and of the varicocelectomy procedure on Sertoli cell function were investigated. Transferrin concentrations in seminal plasma in men with varicocele before and 3 months after the operation were evaluated. Concentrations were measured in 10 normozoospermic fertile men as a control group and 32 oligozoospermic men with varicocele. Also, sperm analysis before and 3 months after the operation was performed. The mean transferrin level in seminal plasma was 108.4 +/- 17.5 micrograms ml-1 in normoozoospermic men and 58.1 +/- 14.4 micrograms ml-1 in patients with varicocele before the operation (P < 0.0001). Mean sperm concentration, motility and normal morphology ratio showed significant improvement 3 months after the operation (P < 0.0001). Although the mean transferrin level increased slightly (to 60.8 +/- 16.2 micrograms ml-1; P = 0.2), there was a statistically significant correlation between the change in transferrin concentration and the change in sperm concentration after the operation (r = 0.56, P = 0.0008). These results showed that elevated transferrin secretion after the treatment seems to be associated with an increase in sperm concentration after varicocelectomy. The finding of improvements in seminal parameters after the operation but insignificant changes in seminal transferrin levels indicates that varicocelectomy results in a greater improvement in sperm quality than in Sertoli cell function. PMID- 10702863 TI - Glucocorticoid receptor distribution in rat testis during postnatal development and effects of dexamethasone on immature peritubular cells in vitro. AB - In this study, the occurrence of the glucocorticoid receptor in the rat testis during early stages of postnatal development and its potential functional significance were investigated. Quantitative analyses of immunohistochemically labelled paraffin sections revealed that the receptor was present during all stages of postnatal development in the nuclei of interstitial cells such as Leydig cells, macrophages and fibroblasts, and endothelial cells of blood vessels. The labelling index increased initially, with maximum levels reached within the second week of postnatal development, and decreased thereafter. Within the seminiferous tubules, the glucocorticoid receptor could be detected in the nuclei of germ cells as well as Sertoli cells, reaching the highest levels in 3 week-old rats, mainly due to immature germ cell staining. In contrast, approximately 50% of the peritubular cell nuclei were stained throughout postnatal development. In vitro experiments on immature and immortalized peritubular cells demonstrated a dose-dependent and significant decrease in proliferation and fibronectin secretion after administration of dexamethasone. The data of this study suggest that glucocorticoids have a consistently repressive effect on peritubular cells throughout postnatal development. In summary, labelling of germ cells, especially in immature rats, might indicate an inhibition of spermatogenesis by corticosteroids. PMID- 10702864 TI - Does acupuncture treatment affect sperm density in males with very low sperm count? A pilot study. AB - Classic therapies are usually ineffective in the treatment of patients with very poor sperm density. The aim of this study was to determine the effect of acupuncture on these males. Semen samples of 20 patients with a history of azoospermia were examined by light microscope (LM) and scanning electron microscope (SEM), with which a microsearch for spermatozoa was carried out. These examinations were performed before and 1 month after acupuncture treatment and revealed that the study group originally contained three severely oligoteratoasthenozoospermic (OTA), two pseudoazoospermic and 15 azoospermic patients. The control group was comprised of 20 untreated males who underwent two semen examinations within a period of 2-4 months and had initial andrological profiles similar to those of the experimental group. No changes in any of the parameters examined were observed in the control group. There was a marked but not significant improvement in the sperm counts of severely OTA males following acupuncture treatment (average = 0.7 +/- 1.1 x 10(6) spermatozoa per ejaculate before treatment vs. 4.3 +/- 3.2 x 10(6) spermatozoa per ejaculate after treatment). A definite increase in sperm count was detected in the ejaculates of 10 (67%) of the 15 azoospermic patients. Seven of these males exhibited post treatment spermatozoa that were detected even by LM. The sperm production of these seven males increased significantly, from 0 to an average of 1.5 +/- 2.4 x 10(6) spermatozoa per ejaculate (Z = -2.8, P < or = 0.01). Males with genital tract inflammation exhibited the most remarkable improvement in sperm density (on average from 0.3 +/- 0.6 x 10(6) spermatozoa per ejaculate to 3.3 +/- 3.2 x 10(6) spermatozoa per ejaculate; Z = -2.4, P < or = 0.02). Two pregnancies were achieved by the IVF-ICSI procedure. It is concluded that acupuncture may be a useful, nontraumatic treatment for males with very poor sperm density, especially those with a history of genital tract inflammation. PMID- 10702865 TI - Peritubular cell-Sertoli cell interactions: factors involved in PmodS activity. AB - The widespread occurrence of peritubular myoid cells in mammalian and other species suggests that they form an integral and functional component of the testis. Peritubular cells contribute to the contractile activity of testicular tubules and maintain mesenchymal-epithelial interactions with Sertoli cells both by cooperation in the deposition of extracellular matrix elements and by secretion of paracrine agonists. One of the most intriguing of these paracrine agonists is known as PModS (Peritubular factor that Modulates Sertoli cell function). The demonstration that, at least under some conditions, PModS production may be stimulated by androgens has led to the hypothesis that PModS may mediate part or all of the effects of androgens on Sertoli cells. The identity of PModS, however, remains elusive. Here we summarize data showing: (1) that production of PModS (-like factors) may not be limited to peritubular cells; (2) that the role of androgens in the control of PModS production remains controversial; (3) that other known mediators including IGF-I, bFGF, cytokines and heregulins mimic some or all of the effects of PModS; (4) that combinations of such growth factors have potent effects. It is concluded that, until PModS has been identified unambiguously, the hypothesis that it acts as an essential andromedin in the testis should be regarded with caution. PMID- 10702866 TI - Macrophage migration inhibitory factor (MIF) as a paracrine mediator in the interaction of testicular somatic cells. AB - Originally the macrophage migration inhibitory factor (MIF) was described as a classical T-cell cytokine. Recently, a much broader tissue distribution for MIF has been revealed. We demonstrated that MIF protein and mRNA are present in the Leydig cells of the normal adult rat testis. Addition of recombinant MIF to cultures of rat seminiferous tubules resulted in decreased secretion of inhibin, whereas follistatin and activin levels remained unchanged, suggesting a paracrine role for MIF in Sertoli cell regulation. Furthermore, MIF showed unique compensatory production in the rat testis. Depletion of the original MIF source, the Leydig cells, by the specific toxin EDS prompted MIF expression by the previously negative Sertoli cells. Leydig cell re-population of the interstitial tissue by precursor cells resulted in a switch back to production by Leydig cells. Therefore, testicular MIF appears to be under very tight paracrine control. MIF has thus been identified as a new mediator in the cross-talk between Leydig cells and the somatic cells of the seminiferous tubules of the rat testis. PMID- 10702867 TI - Inhibin and activin subunits and spermatogenesis. PMID- 10702868 TI - The interleukin-1 system in the testis. AB - The interleukin-1 (IL-1) family consists of two agonist proteins, IL-1 alpha and IL-1 beta, and one antagonist, IL-1 receptor antagonist (IL-1ra), which blocks the action of the agonists by binding and competing at the IL-1 receptor level. IL-1 beta and to a lesser extent IL-1 alpha were originally described as rapidly inducible proinflammatory cytokines released by activated macrophages. However, IL-1 alpha has been found to be constitutively produced by certain tissues, and noninflammatory functions have been proposed for this protein, although they have not yet been well elucidated. Consistent with this suggestion, we previously showed that the intact rat testis constitutively produces large amounts of IL-1 alpha at both the mRNA and protein levels. The expression of IL-1 alpha was found to be confined to Sertoli cells, with evidence of a developmental as well as a stage-dependent production pattern. In more recent studies, we have found indications that the testis can also initiate production of IL-1 beta upon stimulation with inflammatory inducers such as endotoxin. Further, we have detected constitutive testicular expression of IL-1ra, opening up the possibility that IL-1 action in the testis may be specifically regulated by paracrine mechanisms. Recent data have demonstrated that the testis can produce more than one isoform of IL-1 alpha with indications of both post-transcriptional and post translational modifications, resulting in at least three distinct bio- and immunoreactive IL-1 alpha proteins. We conclude that all three classical IL-1 ligands and novel IL-1 alpha isoforms are present in the testis and may serve as paracrine mediators under physiological or pathophysiological conditions. The function of this testicular IL-1 agonist-antagonist network is a current focus of investigation. PMID- 10702870 TI - Hepatocyte growth factor receptor expression in rat testis. PMID- 10702869 TI - Multiple roles of the messenger molecule cGMP in testicular function. AB - The messenger molecule cyclic guanosine monophosphate (cGMP) is produced by different isoforms of the enzyme guanylate cyclase (GC). Natriuretic peptides (ANP and CNP) bind to and activate particulate GCs, whereas NO and CO activate a soluble form of GC. The specific relevance of the cGMP system for reproductive functions has been recently demonstrated by the successful use of sildenafil (Viagra), an inhibitor of cGMP-specific phosphodiesterase type 5, for the treatment of erectile dysfunction. In the testis, cGMP signal transduction pathways are involved in a variety of local functions, based on autocrine or paracrine effects. In particular, cGMP has been suggested to influence motility in spermatozoa, development of testicular germ cells, relaxation of peritubular lamina propria cells, testosterone synthesis in Leydig cells and dilatation of testicular blood vessels. The physiological significance of cGMP accumulation in Scrtoli cells is not yet clear. Taken as a whole, the evidence suggests that cGMP mediated processes might influence both the potentia coeundi within the penis and the potentia generandi at various levels within the testis. PMID- 10702871 TI - ECE-1 is cyclically expressed in Sertoli cells and mediates the spatiotemporal control of tubular contractility. PMID- 10702872 TI - Paracrine mechanisms in the rat testis: developmental aspects. PMID- 10702873 TI - Transforming growth factor-alpha stimulates Sertoli cell proliferation in vitro. PMID- 10702874 TI - Constitutive and inducible production of proinflammatory cytokines by the rat testis. PMID- 10702875 TI - Synthesis and silica-based immobilization of monofunctionalized cyclomaltoheptaose derivatives for enantioselective HPLC. AB - Heptakis(6-O-tert-butyldimethylsilyl-2,3-di-O-methyl)cyclomaltohep taose (6-TBDMS 2,3-Me-beta-CD) and heptakis(2,3,6-tri-O-methyl)cyclomaltoheptaose (per-Me-beta CD) were monofunctionalized by introduction of a 5-cyanopentyl group attached to one of the O-2, O-3 or O-6 positions and subsequent reduction with lithium aluminum hydride to give the corresponding mono-O-(omega-aminohexyl) derivatives. Alternatively, after attachment of a 7-octenyl group and further epoxidation the corresponding mono-omega-epoxyoctyl derivatives of 6-TBDMS-2,3-Me-beta-CD were obtained. The mono-O-(omega-aminohexyl) derivatives were immobilized by reaction with glycidoxypropyl and 'aldehyde' silica, whereas aminopropyl silica was used for the immobilization of the monoepoxyoctyl derivatives. The immobilized cyclodextrin derivatives were partially evaluated as chiral stationary phases in high-performance liquid chromatography (HPLC) and micro-HPLC. PMID- 10702876 TI - Syntheses of amphiphilic glycosylamides from glycosyl azides without transient reduction to glycosylamines. AB - Protected glycosyl azides react with acyl chlorides in the presence of triphenylphosphine to afford glycosylamides in high yields, at room temperature. Starting from the beta-glycosyl azides, the reaction is highly stereoselective and occurs with retention of configuration, whereas the alpha-azido anomers display a lower stereoselectivity giving rise to alpha/beta mixtures of glycosylamides. The reaction was applied to several monosaccharidic azides and to lactosyl azide with various acyl chlorides; it was shown to be of general use for preparing 1,2-trans beta-glycosylamides. PMID- 10702877 TI - Hydrolytic activity of alpha-galactosidases against deoxy derivatives of p nitrophenyl alpha-D-galactopyranoside. AB - The four possible monodeoxy derivatives of p-nitrophenyl (PNP) alpha-D galactopyranoside were synthesized, and hydrolytic activities of the alpha galactosidase of green coffee bean, Mortierella vinacea and Aspergillus niger against them were elucidated. The 2- and 6-deoxy substrates were hydrolyzed by the enzymes from green coffee bean and M. vinacea, while they scarcely acted on the 3- and 4-deoxy compounds. On the other hand, A. niger alpha-galactosidase hydrolyzed only the 2-deoxy compound in these deoxy substrates, and the activity was very high. These results indicate that the presence of two hydroxyl groups (OH-3 and -4) is essential for the compounds to act as substrates for the enzymes of green coffee bean and M. vinacea, while the three hydroxyl groups (OH-3, -4, and -6) are necessary for the activity of the A. niger enzyme. The kinetic parameters (K(m) and Vmax) of the enzymes for the hydrolysis of PNP alpha-D galactopyranoside and its deoxy derivatives were obtained from kinetic studies. PMID- 10702878 TI - Synthesis and preliminary characterisation of charged derivatives and hydrogels from scleroglucan. AB - The synthesis of negatively and positively charged polyelectrolytes from scleroglucan is described. Polycarboxylates were synthesised through nucleophilic substitution with chloroacetic acid or through a selective 2,2,6,6-tetramethyl-l piperidinyloxy (TEMPO)-mediated oxidation of the primary alcohol groups. Amine groups were introduced through nucleophilic substitution with 2-chloroethylamine or 3-chloropropylamine. Reaction conditions were varied to obtain insight into the influence of variables on the degree of substitution. The conformational behaviour of the obtained polyelectrolytes was studied as a function of pH, temperature and solvent. For the products with a low degree of modification, evidence of an ordered conformation was found, whereas the polymers with a higher degree of modification behaved as random coils in solution. The negatively charged polymers were reticulated using the Ugi four-component condensation, obtaining negatively charged hydrogels. The positively charged polymers were reticulated using diethyl squarate (3,4-diethoxy-3-cyclobutene-1,2-dion, DES) to obtain positively charged hydrogels. PMID- 10702879 TI - The reduction of CrVI to CrIII by the alpha and beta anomers of D-glucose in dimethyl sulfoxide. A comparative kinetic and mechanistic study. AB - The reduction of CrVI by alpha-D-glucose and beta-D-glucose was studied in dimethyl sulfoxide in the presence of pyridinium p-toluensulfonate, a medium where mutarotation is slower than the redox reaction. The two anomers reduce CrVI by formation of an intermediate CrVI ester precursor of the slow redox step. The equilibrium constant for the formation of the intermediate chromic ester and the rate of the redox steps are different for each anomer. alpha-D-Glucose forms the CrVI-Glc ester with a higher equilibrium constant than beta-D-glucose, but the electron transfer within this complex is slower than for the beta anomer. The difference is attributed to the better chelating ability of the 1,2-cis-diolate moiety of the alpha anomer. The CrV species, generated in the reaction mixture, reacts with the two anomers at a rate comparable with that of CrVI. The EPR spectra show that the alpha anomer forms several linkage isomers of the five coordinate CrV bis-chelate, while beta-D-glucose affords a mixture of six coordinate CrV monochelate and five-coordinate CrV bis-chelate. The conversion of the CrV mono- to bis-chelate is discussed in terms of the ability of the 1,2-cis- versus 1,2-trans-diolate moieties of the glucose anomers to bind CrV. PMID- 10702880 TI - Phase separation in the mixture of schizophyllan and poly(ethylene oxide) in aqueous solution driven by a specific interaction between the glucose side chain and poly(ethylene oxide). AB - We found that the mixture of schizophyllan and poly(ethylene oxide) in aqueous solution underwent phase separation at around 3-4 degrees C, and this temperature was independent of both polymer concentration and the difference in poly(ethylene oxide) molecular weight (Mw 6000 and 70,000). The phase-separation took place at the same temperature at which the optical rotation changed. Since the optical rotation change is ascribed to the difference in the nature of hydrogen bonding between the schizophyllan side chain and water, the phase separation is also considered to be due to an interaction between poly(ethylene oxide) and schizophyllan. The phase-separation temperature increased on changing H2O to D2O in accordance with a change in the optical rotation, confirming the specific interaction essential for the phase separation. PMID- 10702881 TI - Topography of the 1:1 alpha-cyclodextrin-nitromethane inclusion complex. AB - Dissolution of alpha-cyclodextrin (alpha-CD) in 9:1 water-nitromethane smoothly generates the title compound, which crystallizes as the pentahydrate in the orthorhombic space group P2(1)2(1)2(1) with a = 9.452(4), b = 14.299(3), c = 37.380(10) A, and Z = 4. Its crystal structure analysis revealed the alpha-CD macrocycle in an unstrained conformation stabilized through a ring of O-2...O-3' hydrogen bonds between five of the six adjacent glucose residues. The nitromethane is located in the alpha-CD cavity in an orientation parallel to the plane of the macrocycle, and assumes two sites of equal population with the nitro group in excessive thermal motion; the guest is held by van der Waals contacts and C-H...O-type hydrogen bonds to the pyranose H-3 and H-5 protons. The packing of the macrocycles in the crystal lattice is of cage herringbone-type with an extensive intra- and intermolecular hydrogen bonding network. The ready formation of a nitromethane inclusion complex in aqueous nitromethane, and the subtleties of its molecular structure amply demonstrate the ease with which water is expelled from the alpha-CD cavity by a more hydrophobic co-solvent. PMID- 10702882 TI - Cilostazol pharmacokinetics after single and multiple oral doses in healthy males and patients with intermittent claudication resulting from peripheral arterial disease. AB - OBJECTIVE: To study the pharmacokinetics of cilostazol following single oral administration of 50 to 200 mg in healthy young males, and after repeated oral administration of 100 mg every 12 hours to patients with peripheral arterial disease (PAD). DESIGN: The healthy male single dose study was a single-centre, randomised sequence, open-label, incomplete block, 3-period, 4-treatment, crossover design. The patient study was a single-centre, multiple dose, open label study. STUDY PARTICIPANTS: 20 healthy nonsmoking male volunteers were enrolled and successfully completed the single dose study. 26 patients (21 males, 5 females) with intermittent claudication resulting from PAD were enrolled and completed the single/multiple dose study. MAIN OUTCOME MEASURES: Noncompartmental pharmacokinetic parameters, the area under the plasma concentration-time curve from zero to the time of last measurable plasma concentration, and maximum plasma concentration. RESULTS: Peak plasma concentrations of cilostazol occurred about 3 hours after drug administration and then declined biexponentially with concentrations detectable (> 20 micrograms/L) in the plasma for at least 36 hours postdose. The apparent elimination half-life of cilostazol (approximately 11 hours) was similar after a single dose or after multiple doses, with steady state being reached within 4 days. Cilostazol accumulated 1.7-fold following multiple dose administration. The apparent volume of distribution (Vz/F; 2.76 L/kg) suggested extensive distribution of cilostazol in the tissues. The oral clearance of cilostazol (CL/F; 0.18 L/h/kg) was much lower than liver blood flow, indicating a low extraction ratio drug, and hence low probability of a significant first-pass effect. None of the administered doses were recovered in the urine as unchanged cilostazol, suggesting that metabolism, rather than urinary excretion, is the major elimination route. Following single oral doses of 50 to 200 mg, the plasma concentrations of cilostazol and its metabolites increased less than proportionally to the dose. The pharmacokinetics of cilostazol in normal healthy volunteers are predictive of those in patients with PAD. Single oral doses of 50 to 200 mg cilostazol as well as 100 mg cilostazol every 12 hours were well tolerated. CONCLUSION: The plasma concentration of cilostazol and its metabolites increased less than proportionally with increasing doses. The relatively low plasma clearance and high volume of distribution of cilostazol suggest a low first-pass effect and extensive distribution. The pharmacokinetics of cilostazol in normal volunteers is predictive of that in patients with PAD. Cilostazol was well tolerated in healthy volunteers and patients with intermittent claudication resulting from PAD. PMID- 10702883 TI - Relative bioavailability and effects of a high fat meal on single dose cilostazol pharmacokinetics. AB - OBJECTIVES: The objectives of this research were to (1) assess the relative bioavailability following administration of a 100 mg cilostazol suspension versus 100 mg tablet; (2) assess dosage form equivalency (2 x 50 mg compared with 1 x 100 mg); (3) compare the relative bioavailability following a single 50 mg dose of cilostazol administered as an ethanolic solution versus a 50 mg tablet; and (4) determine the effects of high fat diet on the pharmacokinetics of cilostazol following a single dose of 100 mg cilostazol in the fed or fasted state. Results were compiled from 3 separate studies to address these objectives. DESIGN: All studies involved healthy adult males receiving single oral doses of cilostazol in the fed or fasted state. The fed state consisted of administering cilostazol after ingestion of a high fat meal. One study compared the relative bioavailability of 100 mg suspension and 2 x 50 mg tablet versus 100 mg tablet in a randomised crossover design. The study involving administration of a 50 mg cilostazol ethanolic solution was a single treatment study. The effects of food on the pharmacokinetics of cilostazol after administration of 100 mg cilostazol in the fed or fasted state as well as the pharmacokinetic profile following administration of a single 50 mg oral dose of cilostazol were assessed in a randomised crossover design. STUDY PARTICIPANTS: All participants were healthy nonsmoking males aged between 19 and 48 years whose bodyweight was within 15% of ideal bodyweight. MAIN OUTCOME MEASURES: Noncompartmental pharmacokinetic parameters were determined for each study participant. RESULTS: The area under the plasma concentration-time curve (AUC) parameters were within the 80 to 125% criterion for bioequivalence for the cilostazol and its primary metabolite, OPC 13015. The maximum observed plasma concentrations (Cmax) for these formulations were not equivalent and indicated that the absorption of cilostazol from a suspension is more rapid than from a tablet. The apparent terminal half-lives (t1/2z) of cilostazol and OPC-13015 were shorter after administration of the suspension compared with the tablet. Cmax and AUC following administration of a single 50 mg cilostazol tablet were approximately 80% of that from the same dose administered as an ethanolic solution. The t1/2z of cilostazol decreased from 15.5 hours after a tablet to 2.5 hours after an ethanolic solution. Upon coadministration with a high fat meal, the Cmax of cilostazol increased 90% and AUC infinity increased 25% (p < 0.05). The t1/2z decreased from 15.1 +/- 14.5 hours (mean +/- SD) in the fasted state to 5.4 +/- 2.0 hours in the fed state. Single oral doses of 50 and 100 mg cilostazol were well tolerated. CONCLUSIONS: The relative bioavailability of the 100 mg cilostazol tablet versus an oral 100 mg cilostazol suspension is 100%. The 2 x 50 mg and 1 x 100 mg tablets are considered to be bioequivalent. The absorption following administration of 50 mg cilostazol ethanolic solution is faster and appears to be greater than that after administration of the 50 mg tablet. Coadministration of food increases the rate and extent of cilostazol absorption. The oral pharmacokinetics of cilostazol and metabolites are absorption-rate limited. The significant differences in the t1/2z observed when comparing cilostazol tablet, suspension, and solution as well as the effects of food suggest 'flip-flop' pharmacokinetics. PMID- 10702884 TI - Effect of hepatic impairment on the pharmacokinetics of a single dose of cilostazol. AB - OBJECTIVE: The pharmacokinetic profiles of cilostazol and its metabolites following a single oral dose of cilostazol 100 mg were compared between individuals with impaired and normal liver function. DESIGN: The study was conducted as a single-centre, open-label, single dose pharmacokinetic and tolerability trial. STUDY PARTICIPANTS: 12 patients with impaired and compensated liver function were compared with 12 volunteers with normal liver function. Participants in each group were matched for gender, age and weight. Of the 12 patients with hepatic impairment examined in this study, 10 had mild impairment (Child-Pugh class A) and 2 had moderate impairment (Child-Pugh class B). MAIN OUTCOME MEASURES: Blood and urine were collected up to 144 hours after drug administration. Pharmacokinetics were determined by noncompartmental methods. RESULTS: Protein binding did not differ between the groups (95.2% healthy volunteers, 94.6% hepatically impaired patients). Mean +/- SD unbound oral clearance of cilostazol decreased by 8.6% because of hepatic impairment (3380 +/- 1400 ml/min in healthy volunteers, 3260 +/- 2030 ml/min in hepatically impaired patients). Total urinary excretion of metabolites was significantly higher in healthy volunteers (26 vs 17% of dose). Overall, the pharmacokinetics of cilostazol and its metabolites, OPC-13213 and OPC-13015, were not substantially different in those with mild and moderate hepatic disease compared with values in healthy volunteers. Except for terminal-phase disposition half-life and apparent terminal-phase volume of distribution for cilostazol, the ratios of geometric means of pharmacokinetic parameters for plasma cilostazol, OPC-13213 and OPC 13015 in those with hepatic impairment versus healthy volunteers were close to 100%. CONCLUSIONS: Based on the results of the pharmacokinetic analysis, dose adjustment in patients with mild hepatic impairment is not necessary. However, caution should be exercised when cilostazol is administered to patients with moderate or severe hepatic impairment. PMID- 10702885 TI - Effect of renal impairment on the pharmacokinetics of cilostazol and its metabolites. AB - OBJECTIVE: The pharmacokinetics of cilostazol were studied in patients with mild, moderate and severe renal impairment and in healthy volunteers after administration of 50 mg single and multiple doses of cilostazol. DESIGN: This was an open-label, single and multiple dose study administering 50 mg cilostazol every 12 hours to healthy volunteers and patients with varying degrees of renal impairment. PARTICIPANTS: 6 normal volunteers [creatinine clearance (CLCR) > or = 90 ml/min]; 6 patients with mild (CLCR 50 to 89 ml/min), 5 with moderate (CLCR 26 to 49 ml/min) and 6 with severe (CLCR 5 to 25 ml/min) renal impairment. OUTCOME MEASURES: Noncompartmental pharmacokinetic parameters were determined for each study participant. RESULTS: At steady state, in the severe renal disease group, cilostazol and OPC-13015 peak concentrations (Cmax) were 29 and 41% lower and the areas under the concentration-time curve over the dosage interval (AUC tau) 39 and 47% lower than in the healthy volunteers. Cmax and AUC tau of OPC-13213 were significantly higher, 173 and 209%, respectively, than those in the healthy volunteers. The accumulation ratios were not significantly different between the various renal function groups for cilostazol and its metabolites. The estimated pharmacological activity of cilostazol and its metabolites was similar between the normal volunteers and those with severe renal impairment. CONCLUSIONS: A dosage reduction in renally impaired patients is not supported by the pharmacokinetics of cilostazol and its metabolites in this patient group. PMID- 10702886 TI - Inhibition of CYP2D6 by quinidine and its effects on the metabolism of cilostazol. AB - OBJECTIVE: In vitro results are inconclusive as to whether cilostazol is metabolised by cytochrome P450 isoenzyme 2D6 (CYP2D6). The goals of this study were (1) to assure the dose of quinidine and timing relative to cilostazol used in this study were adequate to cause inhibition of CYP2D6, (2) to evaluate carryover effects of quinidine administration, and (3) to evaluate the effect of CYP2D6 deficiency and administration of quinidine (a CYP2D6 inhibitor) on the pharmacokinetics of a single 100 mg oral dose of cilostazol. DESIGN: This study was conducted as a single-centre, open-label, randomised sequence, 2-period, crossover pharmacokinetic trial. Water alone (treatment without quinidine) or two 200 mg oral doses of quinidine sulfate with water were administered 25 hours and 1 hour prior to a single 100 mg dose of cilostazol in period 1. Study participants were crossed over to opposite treatment in period 2. Metoprolol 25 mg, used as a positive control, was administered 1 hour after quinidine sulfate with water or using water alone to assess the magnitude of CYP2D6 inhibition by quinidine. STUDY PARTICIPANTS: 22 healthy nonsmoking Caucasian (14 male and 8 female) volunteers participated in the study. MAIN OUTCOME MEASURES: Serial blood and urine samples were collected at predose and after cilostazol administration to characterise cilostazol and its metabolite pharmacokinetics. Additional plasma samples were taken to assess the pharmacokinetics of quinidine. Urine samples were collected to measure metoprolol and hydroxymetoprolol. RESULTS: Administration of metoprolol with quinidine caused a significant (p < 0.001) decrease in the urinary 4-hydroxymetoprolol/metoprolol ratio compared with administration of metoprolol alone (42-fold decrease, 0.065 vs 2.707). Hence, quinidine effectively converted extensive metabolisers of CYP2D6 to poor metabolisers of CYP2D6. The 21-day washout period was adequate to have complete recovery from quinidine inhibition of CYP2D6. The analysis of variance demonstrated that the mean maximum plasma concentration (Cmax) for cilostazol, both adjusted and unadjusted for the free fraction, was higher in the control group than in the quinidine group (p = 0.023). However, the time to Cmax (p = 0.669), the area under the plasma concentration-time curve from time zero to infinity (AUC infinity; p = 0.133), and the apparent oral clearance (p = 0.135) were unchanged. The geometric mean ratios (90% confidence interval) comparing with quinidine (test) and without quinidine (reference) coadministration for Cmax and AUC infinity are 0.86 (0.77, 0.95) and 0.92 (0.84, 1.00), respectively. Similar patterns were observed for OPC-13015 and OPC-13213 with regard to Cmax, area under the plasma concentration-time curve from time zero to the last measurable concentration at time t, and AUC infinity (where determinable). The slight decrease in the systemic availability of cilostazol and its metabolites was thought to be a result of the increased gastrointestinal motility secondary to quinidine. CONCLUSIONS: Administration of quinidine sulfate 200 mg profoundly inhibited CYP2D6-mediated metabolism. The effects of quinidine inhibition of CYP2D6 metabolism were completely reversible during the 21-day washout period. Coadministration of quinidine with cilostazol had no substantial effect on cilostazol or its metabolites (OPC-13015 and OPC-13213). Hence, CYP2D6 does not have a significant contribution in the metabolic elimination of cilostazol. PMID- 10702887 TI - Effect of omeprazole on the metabolism of cilostazol. AB - OBJECTIVE: In vitro results suggest that cilostazol is metabolised by cytochrome P450 (CYP) isoforms 1A2, 2D6, 3A4 and 2C19. This study was designed to evaluate the effect of concomitant administration of omeprazole (a CYP2C19 inhibitor) on the pharmacokinetics of a single 100 mg oral dose of cilostazol. DESIGN: This study was conducted as a single-centre, open-label, nonrandomised, 2-period, crossover pharmacokinetic trial. A single 100 mg dose of cilostazol was administered orally on days 0 and 14. Oral omeprazole (40 mg every day) was administered on days 7 to 18. STUDY PARTICIPANTS: 20 healthy nonsmoking male and female volunteers. MAIN OUTCOME MEASURES: Serial blood samples were collected before and after cilostazol administration to characterise the pharmacokinetics of cilostazol and its metabolites. RESULTS: Following omeprazole coadministration, the increases in cilostazol maximum plasma concentration (Cmax) and area under the plasma concentration-time curve at time t (AUCt) were 18% (p = 0.062) and 26% (p < 0.001), respectively. For the 2 major circulating metabolites, OPC-13015 and OPC-13213, the OPC-13015 Cmax and AUCt increased by 29 and 69%, respectively (p < 0.001). However, for OPC-13213, the Cmax and AUCt decreased by 22 and 31%, respectively (p < 0.001). The plasma protein binding of cilostazol was unaffected by coadministration of omeprazole. CONCLUSIONS: Coadministration of cilostazol with omeprazole resulted in an increase in the systemic exposure of cilostazol and its active metabolite, OPC-13015, by 26 and 69%, respectively. For the other active metabolite, OPC-13213, systemic exposure decreased by 31% because of inhibition of cilostazol metabolism to this metabolite. These changes in systemic exposure were well tolerated. A dose of 50 mg cilostazol twice a day should be considered during coadministration of inhibitors of CYP2C19, such as omeprazole. PMID- 10702888 TI - Effects of CYP3A inhibition on the metabolism of cilostazol. AB - OBJECTIVE: In vitro results suggest that cilostazol is metabolised by cytochrome P450 (CYP) isoforms 1A2, 2D6, 3A and 2C19. This study investigated the role of CYP3A inhibition on the metabolism of cilostazol. DESIGN: The study was conducted as a single-centre, open-label, nonrandomised, 2-period, crossover pharmacokinetic trial. A single dose of cilostazol 100 mg was administered orally on days 1 and 15. Erythromycin (150 mg orally 3 times daily) was administered on days 8 to 20. 14C-erythromycin (3 microns Ci) was administered intravenously on days 1 and 15 one hour before cilostazol administration to determine baseline and the inhibitory effect of erythromycin treatment on CYP3A activity. STUDY PARTICIPANTS: 16 healthy nonsmoking male volunteers. MAIN OUTCOME MEASURES: Serial blood and pooled urine samples were collected before and after cilostazol administration to quantitate cilostazol and its metabolites. Serial exhalation samples were collected after intravenous 14C-erythromycin administration and radioactivity was quantitated by scintillation counting. Pharmacokinetics were determined by noncompartmental methods and compared before and after erythromycin administration. Tolerability assessments included adverse events, laboratory tests, vital signs and electrocardiographs. RESULTS: Following erythromycin coadministration, cilostazol maximum plasma concentration (Cmax), area under the plasma concentration-time curve at time t (AUCt), and area under the curve from zero to infinity (AUC infinity) increased significantly by 47, 87, and 73%, respectively, and an approximately 50% reduction in unbound clearance was observed for the major circulating metabolite of cilostazol, OPC-13015. Cmax decreased significantly (p < 0.001) by 24%, while AUCt increased by 8%; this increase was not significant. For the second major metabolite, OPC-13213, the Cmax and AUCt increased by 29 and 141%, respectively (p < 0.001). CONCLUSIONS: In vivo results are in agreement with previous in vitro human microsome studies, indicating that cilostazol is metabolised to OPC-13015 via CYP3A. In addition, OPC-13213 concentrations increased after inhibition of CYP3A because of inhibition of sequential metabolism of OPC-13213 via CYP3A. A starting dose for cilostazol of 50 mg twice daily should be considered during coadministration of inhibitors of CYP3A. PMID- 10702889 TI - Effect of multiple cilostazol doses on single dose lovastatin pharmacokinetics in healthy volunteers. AB - OBJECTIVE: To assess the effects of cilostazol on lovastatin pharmacokinetics. DESIGN: This was a single-centre, open-label, multiple dose, sequential treatment study. Participants received single oral doses of lovastatin 80 mg on days 1, 7 and 9, as well as oral cilostazol 100 mg twice daily on days 2 to 8, followed by a single oral 150 mg cilostazol dose on day 9. STUDY PARTICIPANTS: 15 healthy, nonsmoking male or female volunteers (aged 18 to 60 years) were enrolled, and 12 completed the study. MAIN OUTCOME MEASURES: Pharmacokinetic parameters were calculated using plasma concentrations of lovastatin and its beta-hydroxy metabolite and of cilostazol and its metabolites. Differences in the pharmacokinetics of each drug when given alone or in combination were assessed by analysis of variance. RESULTS: The maximum observed plasma concentration (Cmax) of lovastatin or its metabolite did not differ significantly when lovastatin was given alone and when it was given with 100 mg of cilostazol. The mean ratios of the area under the plasma concentration-time curve from zero to the time of the last measurable concentration (AUCt) for lovastatin coadministered with 100 mg of cilostazol to that with lovastatin given alone were 1.6 for lovastatin and 1.7 for its metabolite. With 150 mg of cilostazol, lovastatin Cmax did not change, whereas Cmax of the metabolite increased 2.2-fold. The mean AUCt ratios for lovastatin given with 150 mg cilostazol/lovastatin given alone were 1.6 and 2.0 for lovastatin and its metabolite, respectively. All increases in lovastatin and metabolite AUC were statistically significant, except for the 1.6-fold increase in lovastatin AUC with 150 mg of cilostazol. Maximum steady-state plasma drug concentration (Cssmax) and AUC during a dosage interval (AUC tau) for cilostazol 100 mg twice daily decreased 14 and 15%, respectively, upon lovastatin coadministration. CONCLUSIONS: Lovastatin and metabolite exposure is increased only by up to 2-fold when cilostazol is coadministered, which is considerably less than that observed for potent CYP3A inhibitors such as itraconazole and grapefruit juice. Absorption of cilostazol decreased approximately 15% when it was given with lovastatin. No dosage adjustments are necessary for cilostazol when coadministered with lovastatin, whereas lovastatin dose reductions may be needed when the 2 drugs are given together. PMID- 10702890 TI - Effect of cilostazol on the pharmacokinetics and pharmacodynamics of warfarin. AB - OBJECTIVE: To evaluate the effect of cilostazol administration on warfarin pharmacokinetics and pharmacodynamics following a single 25 mg dose of warfarin. DESIGN: A randomised double-blind 2-period crossover with healthy volunteers receiving either 100 mg cilostazol twice daily for 13 days or matching placebo twice daily for 13 days, and the other treatment 21 days later. A single 25 mg dose of warfarin was given 14 days prior to the start of the study, and 7 days after the cilostazol and placebo treatments. STUDY PARTICIPANTS: 15 normal healthy male volunteers. OUTCOME MEASURES: Noncompartmental pharmacokinetic parameters for (R)- and (S)-warfarin, the area under the curve of the prothrombin time (AUCPT), activated partial thromboplastin time (AUCaPTT), Ivy bleeding times, unbound fraction (fu) of cilostazol, and warfarin were determined for each individual. RESULTS: For (R)- and (S)-warfarin, the 90% confidence intervals for the ratios of the geometric means (90% CI) of the maximum plasma concentration and area under the plasma concentration-time curve were between 0.88 to 1.03. The 90% CI for the AUCPT and AUCaPTT was between 0.95 and 1.06. For Ivy bleeding time, the 90% CI for the ratios of the geometric means ranged between 0.71 and 1.22. The fu of cilostazol did not differ significantly between the 2 treatments. There was a 17% increase in the fu of warfarin (p < 0.05), which was not clinically significant. CONCLUSIONS: Coadministration of warfarin with twice daily administration of cilostazol 100 mg did not alter (R)- and (S)-warfarin pharmacokinetics, prothrombin time, partial thromboplastin time, Ivy bleeding times, or cilostazol protein binding. PMID- 10702891 TI - Interaction potential and tolerability of the coadministration of cilostazol and aspirin. AB - OBJECTIVE: This study evaluated the effects of repeated oral drug administration with cilostazol alone and with aspirin (acetylsalicylic acid) on platelet aggregation, coagulation and bleeding time as well as the cilostazol-aspirin pharmacokinetic interaction in healthy males. DESIGN: This was a randomised, double-blind, placebo-controlled, crossover study. Participants received either cilostazol 100 mg or placebo twice a day for 10 days; aspirin 325 mg/day was coadministered for the last 5 days. After a 14-day washout period, participants received the alternative treatment. STUDY PARTICIPANTS: 12 healthy male volunteers were enrolled. MAIN OUTCOME MEASURES: Differences in bleeding times, platelet aggregation, prothrombin time (PT) and activated partial thromboplastin time (aPTT) between cilostazol with aspirin and cilostazol alone. Noncompartmental pharmacokinetic parameters were determined for each study participant. RESULTS: Cilostazol, with or without aspirin, caused no changes in PT, aPTT or bleeding time. There was a 23 to 35% increase in inhibition of ADP induced ex vivo platelet aggregation by cilostazol plus aspirin when compared with aspirin alone. There was no additive or synergistic effect on arachidonic acid-induced platelet aggregation. Statistically significant but clinically insignificant increases in the area under the plasma concentration-time curve to the last measurable plasma concentration and trough concentrations of cilostazol and its metabolites (OPC-13015 and OPC-13213) occurred after aspirin coadministration, with no differences observed in the maximum plasma concentration Drug-related adverse events were generally mild, the most frequent being headache. CONCLUSIONS: Cilostazol and aspirin coadministration did not cause clinically significant changes in PT, aPTT, bleeding time, platelet aggregation or plasma concentrations of cilostazol and its 2 active metabolites. Cilostazol was generally well tolerated with or without aspirin. PMID- 10702892 TI - Morphology and classification of acute myeloid leukemias. AB - The morphology and classification of AML are discussed. In addition to routine morphology and cytochemistry, however, it is now necessary to use newer modalities of immunocytochemistry or flow cytometry to confirm a diagnosis. These latter are essential for the diagnosis of the newer described entities of AML with minimal differentiation (FAB-M0) and acute megakaryoblastic leukemia (FAB M7). Cytogenetics and fluorescent-in-situ-hybridization techniques are also very important for diagnosis as in FAB-M3 (promyelocytic leukemia) but also for detecting those myeloid leukemias that are associated with a favorable or unfavorable response. PMID- 10702893 TI - Acute lymphoblastic leukemia. AB - Over the last two decades, great strides have been made in the treatment of acute lymphoblastic leukemia (ALL). This progress has been paralleled by advances in diagnosis. In addition to morphology and cytochemistry, the diagnostic and prognostic importance of immunophenotypic and genetic features is becoming increasingly apparent. This article reviews the clinical and morphologic features, immunophenotypic classification, and karyotypic abnormalities of ALL, and discusses how these aspects relate to the diagnosis of ALL. PMID- 10702894 TI - Special stains in the diagnosis of acute leukemia. AB - Special stains are used in the evaluation of bone marrow specimens to augment Wright-Giemsa preparations. This article details the common cytochemical stains available, and discusses their clinical use as indicators of hematopoietic lineage. Iron and reticulin stains, which are widely used in the general evaluation of the bone marrow, are also reviewed. PMID- 10702895 TI - Chromosomal abnormalities in acute leukemias. AB - Conventional cytogenetic techniques are the standard for the diagnosis and follow up of patients with AML and ALL. Some characteristic translocations associated with various groups of AML diagnoses, such as t(8;21), t(15;17), and inv(16) for M2, M3, and M4eo, respectively, have been recognized for years. The most common cytogenetic abnormality found in childhood ALL and hyperdiploid adult ALL is t(9;22). Future directions include increased use of FISH and molecular diagnostic techniques. The clinical cytogenetics laboratory plays a major role in the diagnosis and management of AML and ALL. PMID- 10702896 TI - Molecular mechanisms in myelodysplastic syndromes and implications for evolution to acute leukemias. AB - This article reviews the molecular lesions that occur in the clonal hematopoietic disorders classified as myelodysplastic syndromes (MDS). A systematization of these molecular lesions is attempted based on the types of molecular abnormalities. Characteristically, these molecular lesions affect pluripotent hematopoietic progenitors and therefore affect myeloid, monocytic, erythroid, and megakaryocytic lineages. Progression of MDS to acute myelogenous leukemia is common; transformation is considered the final stage in the multi-step process of accumulation of molecular lesions over a prolonged latency period of MDS evolution. Although no molecular lesion is MDS specific, multiple combinations of molecular lesions are common and a systematic approach based on the type of molecular abnormality may offer a better understanding of MDS pathogenesis and the basis for new therapeutic strategies. PMID- 10702897 TI - Therapy-related acute leukemia. AB - Acute leukemias that arise as a result of treatment with DNA-damaging agents exhibit distinctive molecular, genetic, and clinico-pathologic features. In this timely article, the authors dissect the pathogenetic basis of therapy-related leukemias, elucidating important molecular mechanisms through which DNA damage causes these disorders. The authors also discuss how these molecular aberrations translate into specific clinical syndromes, and in addition, point out potential molecular targets for the development of innovative treatment approaches. PMID- 10702898 TI - Acute promyelocytic leukemia. From morphology to molecular lesions. AB - Acute promyelocytic leukemia is a unique subtype of acute myelogenous leukemia characterized by distinct morphologic, cytogenetic, and clinical characteristics. The t(15;17) translocation, which is a hallmark of this disease, results in a transcriptionally active fusion gene-derived from the PML gene from chromosome 15 and the retinoic acid receptor alpha (RARa) gene from chromosome 17. The PML/RARa protein product is responsible for the leukemic phenotype in these patients, but is also able to respond to pharmacologic levels of retinoic acid and induce cell differentiation. Treatment of this leukemia by retinoic acid represents the first example of gene-directed differentiation therapy for acute leukemia and has lead to greater understanding of the pathogenesis of this disease at the molecular level. PMID- 10702899 TI - Minimal residual disease in acute promyelocytic leukemia. AB - In the last decade our understanding of acute promyelocytic leukemia (APL) has advanced tremendously. The recognition of all-trans retinoic acid (ATRA) as a powerful therapeutic agent paralleled the cloning of the t(15;17) breakpoint. RtPCR for the PML-RARA hybrid mRNA has become the hallmark of molecular diagnosis and molecular monitoring in APL. Current techniques are useful in predicting complete remission and a possible cure in many patients who repeatedly test negative by PCR. Standardizing techniques and improving the sensitivity of the assay are important. Doing this in a way so that clinically relevant minimal residual disease can be distinguished from "indolent disease" remains among the future challenges in APL. PMID- 10702900 TI - The acute erythroleukemias. AB - Acute erythroleukemia is an aggressive leukemia derived from a multipotential stem cell. Three subtypes have been described: (1) M6a with greater than or equal to 30% blasts of the nonerythrocytic component, (2) M6b with greater than or equal to 30% pronormoblasts of the erythrocytic elements, and (3) M6c with greater than or equal to 30% blasts and greater than or equal to 30% pronormoblasts by the aforementioned exclusion criteria. The poor prognosis associated with this disorder positively correlates with a high pronormoblast:myeloblast ratio; unfavorable cytogenetic aberrations; a high proliferative index; and the presence of P-glycoprotein expression (multidrug resistance phenotype). Chemotherapeutic regimens directed toward these specific parameters should be devised in order to improve the characteristically poor outcome of this patient population. PMID- 10702901 TI - Molecular diagnosis in pediatric acute leukemias. AB - This article summarizes the tremendous progress currently achieved in understanding the molecular basis of the pediatric acute leukemias. The article is organized from the perspective of the most frequently encountered pediatric acute leukemia genetic abnormalities in a molecular diagnostics laboratory setting. For each specific entity, the basic molecular biology, putative mechanisms of leukemogenesis, detection methods, and clinical significance are reviewed. Emphasis is placed on discussing the fusion genes generated from common nonrandom chromosomal translocations in B-lineage acute lymphoblastic leukemia (ALL), although brief summaries of T-lineage and myeloid leukemia, as well as the use of the antigen receptor gene rearrangement for residual disease monitoring in acute lympocytic leukemia are also presented. Finally, an overview of emerging technologies of potential importance in the laboratory diagnosis and evaluation of the pediatric acute leukemias is provided. PMID- 10702902 TI - Gene therapy of pediatric leukemia. AB - Gene therapy approaches offer the possibility of adding a treatment modality that is non-cross-reactive with chemotherapy to the therapeutic options for children with leukemia. Current strategies under evaluation in preclinical models include modification of leukemia cells to decrease tumorogenicity or increase immunogenicity and gene transfer to immune system cells to enhance immune function. Some of these approaches are now being evaluated in clinical trials. PMID- 10702903 TI - Bone marrow engraftment analysis after allogeneic bone marrow transplantation. AB - BME analysis of allo-BMT patients is used to confirm engraftment and detect MC after transplant. With frequent monitoring, the detection of MC by BME analysis may alert clinicians to a high risk of relapse and allow early intervention with a rapid taper of immunosuppression or DLI therapy. In pancytopenic patients BME analysis can help differentiate relapse from drug toxicity or infection. Although many methods have been used for BME analysis, PCR amplification of STR loci is the choice of many clinical laboratories because it is informative, quantitative, relatively rapid, and sensitive. The sensitivity of BME analysis is dependent upon many factors that need to be optimized by the laboratory performing the analysis. Although BME analysis is complex, the clinical significance for allo BMT patients warrants the effort to develop, validate, and perform BME analysis in support of the allo-BMT service. PMID- 10702904 TI - Attention deficit-hyperactivity disorder and its deceivers. AB - There is a myriad of disorders that can mimic ADHD. Often parents or teachers, through their own investigation, will determine the diagnosis for their child's school problems as ADHD, when in fact, the difficulties are unrelated to ADHD. A carefully taken history, observation, and interaction with the child are needed. An evaluation of the school situation will help to indicate if the child's primary problem is behavioral, academic, medical, psychiatric, social, or attentional. Psychologic and educational testing is necessary to completely delineate the child's problems and needs. The greatest service that a physician can give children with academic problems is to approach each child in a systematic, scientific, and professional manner to determine the best treatment for the child and to demonstrate the most favorable outcome. PMID- 10702905 TI - Geriatric rehabilitation following fractures in older people: a systematic review. PMID- 10702906 TI - Isolation of a Bacillus sp. capable of transforming isoeugenol to vanillin. AB - Natural aroma compounds are of major interest to the flavor and fragrance industry. Due to the limited sources for natural aromas, there is a growing interest in developing alternative sources for natural aroma compounds, and in particular aromatic aldehydes. In several microbial species aromatic aldehydes are detected as intermediates in the degradation pathway of phenylpropanoids. Thus, bioconversion of phenylpropanoids is one possible route for the production of these aroma compounds. The present work describes the isolation of microbial strains, capable of producing vanillin from isoeugenol. Bacterial strains isolated from soil, were screened for their ability to transform isoeugenol to vanillin. One of these strains, strain B2, was found to produce high amounts of vanillin when grown in the presence of isoeugenol, and was also capable of growing on isoeugenol as the sole carbon source. Based on its fatty acids profile, strain B2 was identified as a Bacillus subtilis sp. The bioconversion capabilities of strain B2 were tested in growing cultures and cell free extracts. In the presence of isoeugenol, a growing cultures of B. subtilis B2 produced 0.61 g l-1 vanillin (molar yield of 12.4%), whereas cell free extracts resulted in 0.9 g l-1 vanillin (molar yield of 14%). PMID- 10702907 TI - Isolation of antigen specific llama VHH antibody fragments and their high level secretion by Saccharomyces cerevisiae. AB - Recently the existence of 'heavy chain' immunoglobulins in Camelidae has been described. However, as yet there is no data on the binding of this type of antibody to haptens. In addition, it was not a priori predictable whether the binding domains (VHH) of these antibodies could be produced and secreted by the lower eukaryotic micro-organism Saccharomyces cerevisiae. In the present study these questions are addressed. Heavy chain immunoglobulins directed against two hapten molecules, the azo-dyes RR6 and RR120 as well as the (proteinaceous) human pregnancy hormone, have been raised in Lama glama. We were able to select specific VHH fragments for all three antigens by direct screening of Escherichia coli or yeast libraries, even without prior enrichment via bio-panning. This is the first example of the isolation of llama anti-hapten VHH domains. Surprisingly, the affinities of the llama VHHs for the RR6 hapten obtained in this way are in the low nM range. Furthermore, some of the antigen specific VHHs were secreted by S. cerevisiae at levels over 100 mg l-1 in shake flask cultures. These two findings extend the possible application areas for the llama VHH fragments significantly. PMID- 10702908 TI - Pathways of glutamine metabolism in Spodoptera frugiperda (Sf9) insect cells: evidence for the presence of the nitrogen assimilation system, and a metabolic switch by 1H/15N NMR. AB - 1H/15N and 13C NMR were used to investigate metabolism in Spodoptera frugiperda (Sf9) cells. Labelled substrates ([2-15N]glutamine, [5-15N]glutamine, [2 15N]glutamate, 15NH4Cl, [2-15N]alanine, and [1-13C]glucose) were added to batch cultures and the concentration of labelled excreted metabolites (alanine, NH4+, glutamine, glycerol, and lactate) were quantified. Cultures with excess glucose and glutamine produce alanine as the main metabolic by-product while no ammonium ions are released. 1H/15N NMR data showed that both the amide and amine-nitrogen of glutamine was incorporated into alanine in these cultures. The amide-nitrogen of glutamine was not transferred to the amine-position in glutamate (for further transamination to alanine) via free NH4+ but directly via an azaserine inhibitable amido-transfer reaction. In glutamine-free media 15NH4+ was consumed and incorporated into alanine. 15NH4+ was also incorporated into the amide position of glutamine synthesised by the cells. These data suggest that the nitrogen assimilation system, glutamine synthetase/glutamate synthase (NADH GOGAT), is active in glutamine-deprived cells. In cultures devoid of glucose, ammonium is the main metabolic by-product while no alanine is formed. The ammonium ions stem both from the amide and amine-nitrogen of glutamine, most likely via glutaminase and glutamate dehydrogenase. 13C NMR revealed that the [1 13C] label from glucose appeared in glycerol, alanine, lactate, and in extracellular glutamine. Labelling data also showed that intermediates of the tricarboxylic acid cycle were recycled to glycolysis and that carbon sources, other than glucose-derived acetylCoA, entered the cycle. Furthermore, Sf9 cell cultures excreted significant amounts glycerol (1.9-3.2 mM) and ethanol (6 mM), thus highlighting the importance of sinks for reducing equivalents in maintaining the cytosolic redox balance. PMID- 10702909 TI - Treatment of recycled kraft pulps with Trichoderma reesei hemicellulases and cellulases. AB - Effects of recycling ECF-bleached softwood kraft pulp on pulp properties were evaluated in the laboratory. The tensile strength, fiber flexibility and WRV lost during drying of the pulp were recovered by refining between the cycles which, however, resulted in deteriorated drainage properties. The recycled pulps were treated with purified Trichoderma reesei cellulases and hemicellulases and the changes in fiber properties due to enzymatic treatments were characterized. The endoglucanases (EG I and EG II) significantly improved pulp drainage already at low dosage levels, and EG II was found to be more effective at a given level of carbohydrate solubilization. Combining hemicellulases with the endoglucanase treatments increased the positive effects of the endoglucanases on pulp drainage. However, as a result of the endoglucanase treatments a slight loss in strength was observed. Combining mannanase with endoglucanase treatments appeared to increase this negative effect, whereas the impact of xylanase was not significant. Although the drainage properties of the pulps could be improved by selected enzymes, the water retention capacity of the dried hornified fibers could not be recovered by any of the enzymes tested. PMID- 10702910 TI - Synthesis and purification of a deleted human growth hormone, hGH delta 135-146: sensitivity to plasmin cleavage and in vitro and in vivo bioactivities. AB - Proteolytically cleaved human 22 kDa growth hormone (22K hGH) between the amino acid residues 134 and 150 by plasmin or other proteases in vitro has been reported to be most active in growth promoting activity. In this study a deleted mutant hGH lacking amino acid residues from 135 to 146 and having more sensitivity to plasmin digestion was produced using the inverse polymerase chain reaction method and the Escherichia coli expression system. The mutant, hGH delta 135-146, was folded and purified effectively and found to be more sensitive to plasmin cleavage to form the two-chain form in vitro. The biological activities of this plasmin sensitive hGH delta 135-146 were tested by in vitro cell proliferation assays and in vivo growth promoting assay. In Ba/F3-hGHR cells, which express receptors for hGH, hGH delta 135-146 showed 10-20% less growth promoting activity than 22K hGH, but expressed comparable quantities of IGF-I mRNA to that of 22K hGH. In Nb2 rat lymphoma cells, which proliferate in response to hGH via the lactogenic receptors, hGH delta 135-146 showed equivalent activities to those of 22K hGH at lower concentrations. By the body weight gain test using hypophysectomized rats, a lower dose (2.5 nmol kg-1) of hGH delta 135 146 exhibited an equivalent activity to that of wild type 22K hGH, but a higher dose (25 nmol kg-1) of the mutant showed less growth promoting activity than 22K hGH. These results indicated that the plasmin sensitive recombinant hGH delta 135 146 failed to show higher biological activity than the 22K hGH in vivo, suggesting the unsuccessful formation of the active two-chain form in vivo. PMID- 10702911 TI - Mass-spectral analysis of human interferon-gamma and chloramphenicol acetyltransferase I produced in two Escherichia coli strains. AB - Recombinant human interferon-gamma and chloramphenicol acetyltransferase I were isolated from two Escherichia coli strains, E. coli LE329 and E. coli XL1-blue and characterized by electrospray ionization mass spectrometry (ESI-MS). The ESI MS analysis showed higher masses in comparison with the theoretically calculated for both proteins as well as unexpected molecular heterogeneity. The ESI-MS spectral patterns of the proteins depended on the host strain used and were more heterogenous for the proteins isolated from E. coli LE392. One of the proteins (human interferon-gamma obtained from E. coli XL1-blue) was further subjected to BrCN cleavage. The ESI-MS analysis of the polypeptide mixture revealed shift in the molecular mass for two peptides including the last 26 amino acids of the human interferon-gamma molecule. PMID- 10702912 TI - Comparison of particulate and continuous-bed columns for protein displacement chromatography. AB - A conventional anion exchange column packed with porous particles (BioScale Q2), and a novel continuous-bed column (UNO Q1) were compared for displacement separation of dairy whey proteins with polyacrylic acid as displacer. The steric mass action model was investigated as a means to aid and accelerate this development. Characteristic charges and steric factors were measured for the proteins and the displacer according to the model, and used together with the affinity constant derived from the adsorption isotherms for simulations, as well as for the construction of the affinity and operating regime plots. If possible, the latter two were used to select conditions for the actual experiments. In the case of the particle-based column, experimental results and simulations did not agree. In addition, the operating regime plot could not be constructed. The affinity plot did predict the order in the displacement train correctly, but gave misleading information concerning the possible effect of a change in displacer concentration. This is taken to be a result of the porous nature of the particles, which handicaps, to some extent, the interaction of the proteins and the displacer molecules with the adsorptive surface. Results were considerably better in case of the continuous-bed column, where there is no intraparticulate surface. PMID- 10702913 TI - Overproduction and characterization of seleno-methionine xylanase T-6. AB - The extracellular xylanase from Bacillus stearothermophilus T-6 is a thermostable alkaline tolerant enzyme that was found to bleach pulp optimally at pH 9 and 65 degrees C, and was successfully used in a large-scale bio-bleaching mill trial. In an attempt to obtain a heavy atom derivative suitable for complete X-ray analysis, xylanase T-6 was labeled biosynthetically with seleno-methionine, resulting in a 'built-in' array of atoms with specific X-ray anomalous scattering signal. Optimization of growth conditions resulted in over 0.8 g of homogeneous seleno-methionine xylanase T-6 per liter culture. The seleno-methionine enzyme was shown to be fully active and produced single crystals suitable for complete multiple wavelength anomalous diffraction (MAD) structural analysis. PMID- 10702914 TI - Starting a new century. PMID- 10702915 TI - Optimization of the extraction and fractionation of corn bran oil using analytical supercritical fluid instrumentation. AB - Supercritical fluid extraction (SFE) is combined with supercritical fluid chromatography (SFC) in an analytical mode to develop a system for fractionating and enriching high value ferulate-phytosterol esters (FPE) contained in corn bran oil. Corn bran is initially extracted with neat supercritical carbon dioxide (SC CO2) at various pressures (13.8, 34.5, and 69 MPa) and temperatures (40, 60, and 80 degrees C) to see if the FPE can be enriched in the extracts. These initial studies show the greatest percentage of FPE could be extracted under two sets of conditions: 69 MPa at 80 degrees C and 34.5 MPa at 40 degrees C. Both sets of parameters yield an extract containing approximately 1.25% FPE. A stock supply of corn bran oil is then produced by scaled-up SFE at 34.5 MPa and 40 degrees C for subsequent chromatographic fractionation. The SFE-obtained corn bran oil is then applied to the head of a minichromatographic column containing an amino-propyl sorbent. SFC is than commenced using neat SC-CO2 at 69 MPa and 80 degrees C to remove the majority of the triglyceride-based oil. Pressure and temperature are then lowered to 34.5 MPa and 40 degrees C, respectively, and ethanol is added as a modifier. The modifier is added in an increasing stepwise gradient program, and fractions are collected at equal volume intervals. The resultant fractions are analyzed by analytical high-performance liquid chromatography with evaporative light-scattering detection and show that FPE could be enriched to a 14.5% (w) level. PMID- 10702917 TI - Forensic discrimination of automotive paint samples using pyrolysis-gas chromatography-mass spectrometry with multivariate statistics AB - Analytical pyrolysis-gas chromatography (Py-GC) has been a standard method for the forensic analysis of automotive paint for a number of decades. Automotive paints are often identified by visual comparison of pyrograms for peak presence and intensities; however, such analyses can be subjective and time consuming. A preliminary investigation based on Py-GC-mass spectrometric analysis of 100 automobile paint samples of five different colors is presented. Designed experiments are employed to select pyrolysis conditions for adequate discrimination. Pattern recognition techniques including principal component analysis and canonical variates analysis are used to visualize clustering of pyrograms to validate comparisons between different automotive paint pyrograms. These methods have the potential to ease the interpretation task for data sets involving a large number of comparisons. PMID- 10702916 TI - Examination of the enantiomeric distribution of certain monoterpene hydrocarbons in selected essential oils by automated solid-phase microextraction-chiral gas chromatography-mass selective detection. AB - A viable approach for the determination of sources of essential oils based on automatic injection solid-phase microextraction-chiral-gas chromatography-mass selective detection is demonstrated. With no sample preparation, it is shown that the source of essential oils such as peppermint, spearmint, and rosemary can be easily distinguished. Short fiber exposure times of approximately 6 s to the headspace above submicroliter quantities of the selected oils are all that is required to obtain both the required sensitivity and resolution to afford analyses with excellent reproducibilities (relative standard deviation values consistently less than 5.0%). PMID- 10702918 TI - Optimization of extraction conditions for low-molecular-weight analytes using solid-phase microextraction AB - A group of volatile analytes under a molecular weight of 90 and representing 11 organic classes are extracted using identical conditions with 6 different solid phase microextraction fiber coatings. The amount of each of the analytes extracted by the various fibers is shown. The effects of sample modifiers, such as pH and ionic strength, on the recovery of the analytes are presented. A comparison of headspace and immersion extraction techniques is shown. PMID- 10702919 TI - Analysis of tetrahydrofuran and methanol in distillation residue samples by automated headspace solid-phase microextraction-gas chromatography with flame ionization detection AB - Automated headspace solid-phase microextraction (SPME) coupled with gas chromatography and flame ionization detection is used to determine the amounts of methanol and tetrahydrofuran (THF) in distillation residue samples from a proprietary chemical reaction. A 65-micron polydimethylsiloxane/divinylbenzene SPME fiber is used to perform the extractions. Optimized extraction conditions for each analyte are determined using a parts-per-million-level methanol in water standard and a parts-per-billion-level THF in water standard. The amount of methanol and THF in distillation residue samples is quantitated by both standard addition and external standard calibration curve. The two methods of quantitation are compared. PMID- 10702920 TI - Quantitative analysis in field-flow fractionation using ultraviolet-visible detectors: an experimental design for absolute measurements AB - In previous works, it has been shown that a standard ultraviolet-visible detection system can be used for quantitative analysis of heterogeneous systems (dispersed supermicron particles) in field-flow fractionation (FFF) by single peak area measurements. Such an analysis method was shown to require either experimental measurements (standardless analysis) or an accurate model (absolute analysis) to determine the extinction efficiency of the particulate samples. In this work, an experimental design to assess absolute analysis in FFF through prediction of particles' optical extinction is presented. Prediction derives from the semiempirical approach by van de Hulst and Walstra. Special emphasis is given to the restriction of the experimental domain of instrumental conditions within which absolute analysis is allowed. Validation by statistical analysis and a practical application to real sample recovery studies are also given. PMID- 10702921 TI - Relative molar response factors for thermal conductivity detectors AB - This paper describes an investigation into the universal nature of relative molar response factors for thermal conductivity detectors. Relative molar response factors are measured on multiple gas chromatographs equipped with thermal conductivity detectors, and the values are compared with values in the literature. As was observed previously, relative molar responses obtained on a single instrument for a homologous series vary linearly with respect to the number of carbon atoms in the hydrocarbon chain. However, significant differences are observed for the slope of this line depending on the instrument studied. This contradicts previous literature results that demonstrated an indepedence of the relative molar response with regard to the detector. The current results show that the calibration of thermal conductivity detectors using literature values for relative molar response factors could produce significant errors in the concentrations measured by the laboratory chromatograph. PMID- 10702922 TI - Evaluation of the EVA descriptor for QSAR studies: 3. The use of a genetic algorithm to search for models with enhanced predictive properties (EVA_GA). AB - The EVA structural descriptor, based upon calculated fundamental molecular vibrational frequencies, has proved to be an effective descriptor for both QSAR and database similarity calculations. The descriptor is sensitive to 3D structure but has an advantage over field-based 3D-QSAR methods inasmuch as structural superposition is not required. The original technique involves a standardisation method wherein uniform Gaussians of fixed standard deviation (sigma) are used to smear out frequencies projected onto a linear scale. The smearing function permits the overlap of proximal frequencies and thence the extraction of a fixed dimensional descriptor regardless of the number and precise values of the frequencies. It is proposed here that there exist optimal localised values of sigma in different spectral regions; that is, the overlap of frequencies using uniform Gaussians may, at certain points in the spectrum, either be insufficient to pick up relationships where they exist or mix up information to such an extent that significant correlations are obscured by noise. A genetic algorithm is used to search for optimal localised sigma values using crossvalidated PLS regression scores as the fitness score to be optimised. The resultant models were then validated against a previously unseen test set of compounds and through data scrambling. The performance of EVA_GA is compared to that of EVA and analogous CoMFA studies; in the latter case a brief evaluation is made of the effect of grid resolution upon the stability of CoMFA PLS scores particularly in relation to test set predictions. PMID- 10702923 TI - Statistical relationships among docking scores for different protein binding sites. AB - This report describes the existence of statistical relationships among scores computed with the DOCK program for a library of small molecules and a panel of protein binding sites. Multivariate relationships are observed in docking scores computed for a constant set of ligands in different binding sites of proteins that are dissimilar in structure and function. The structural basis for the correlations found among scores is analyzed in terms of size, shape and charge characteristics of the binding sites considered. Interestingly, these results parallel a growing body of evidence demonstrating the promiscuous behavior of small molecules in their interactions with macromolecules that could have an impact in future efforts in drug design. PMID- 10702924 TI - A molecular-field-based similarity study of non-nucleoside HIV-1 reverse transcriptase inhibitors. 2. The relationship between alignment solutions obtained from conformationally rigid and flexible matching. AB - An analysis of the relationship among alignment solutions obtained from field based matching of a representative set of rigid conformers of three non nucleoside HIV-1 reverse transcriptase inhibitors and solutions obtained from flexible matching of the same conformers is presented. In some cases, different alignment solutions obtained from rigid matching converge to the same solution when conformational rigidity is relaxed, indicating that a reduced set of conformers per molecule may be sufficient in many field-based similarity studies. Furthermore, the results also indicate the importance of going beyond the pairwise similarity level to obtain consistent solutions in flexible-matching studies. In this respect, the best conformationally flexible multi-molecule alignment obtained is found to be in good agreement with the relative binding geometry and orientation found experimentally from protein-ligand crystal structures. The rms separation between corresponding atoms in computed and 'experimental' sets of three inhibitor structures is 0.94 A. PMID- 10702925 TI - Modeling the interactions of a peptide-major histocompatibility class I ligand with its receptors. I. Recognition by two alpha beta T cell receptors. AB - A three-dimensional model of the complex between an Influenza Hemagglutinin peptide, Ha255-262, and its restricting element, the mouse major histocompatibility complex (MHC) class I molecule, Kk, was built by homology modeling and subsequently refined by simulated annealing and restrained molecular dynamics. Next, three-dimensional models of two different T cell receptors (TCRs) both specific for the Ha255-262/Kk complex were generated based on previously published TCR X-ray structures. Finally, guided by the recently published X-ray structures of ternary TCR/peptide/MHC-I complexes, the TCR models were successfully docked into the Ha255-262/Kk model. We have previously used a systematic and exhaustive panel of 144 single amino acid substituted analogs to analyze both MHC binding and T cell recognition of the parental viral peptide. This large body of experimental data was used to evaluate the models. They were found to account well for the experimentally obtained data, lending considerable support to the proposed models and suggesting a universal docking mode for alpha beta TCRs to MHC-peptide complexes. Such models may also be useful in guiding future rational experimentation. PMID- 10702926 TI - Modeling the interactions of a peptide-major histocompatibility class I ligand with its receptors. II. Cross-reaction between a monoclonal antibody and two alpha beta T cell receptors. AB - The recombinant antibody, pSAN13.4.1, has a unique T cell like specificity; it binds an Influenza Hemagglutinin octapeptide (Ha255-262) in an MHC (H-2Kk) restricted manner, and a detailed comparison of the fine specificity of pSAN13.4.1 with the fine specificity of two Ha255-262-specific, H-2Kk-restricted T cell hybridomas has supported this contention. A three-dimensional model of pSAN13.4.1 has been derived by homology modeling techniques. Subsequently, the structure of the pSAN13.4.1 antibody in complex with the antigenic Ha-Kk ligand was derived after a flexible and automated docking of the MHC-peptide pair into the Fab combining site. Interestingly, the most energetically favored binding mode shows numerous analogies to the recently determined recognition of class I MHC-peptide complexes by alpha beta T cell receptors (TCRs). The pSAN13.4.1 also binds diagonally across the MHC binding groove but is more deeply anchored to the peptide-MHC (pep/MHC) ligand than TCRs, notably through numerous interactions of its heavy chain. The present model accounts well for the experimentally determined binding affinity of a set of 144 single amino acid substituted Ha analogues and the observed shared specificity between the pSAN antibody and two different T cell receptors for the Ha-Kk antigenic ligand. Analogies and differences between Fab and TCR recognition are explained by dissecting the binding role of each chain of the immune receptors as well as the contribution of all peptide amino acids. PMID- 10702927 TI - Refinement of modelled structures by knowledge-based energy profiles and secondary structure prediction: application to the human procarboxypeptidase A2. AB - Knowledge-based energy profiles combined with secondary structure prediction have been applied to molecular modelling refinement. To check the procedure, three different models of human procarboxypeptidase A2 (hPCPA2) have been built using the 3D structures of procarboxypeptidase A1 (pPCPA1) and bovine procarboxypeptidase A (bPCPA) as templates. The results of the refinement can be tested against the X-ray structure of hPCPA2 which has been recently determined. Regions miss-modelled in the activation segment of hPCPA2 were detected by means of pseudo-energies using Prosa II and modified afterwards according to the secondary structure prediction. Moreover, models obtained by automated methods as COMPOSER, MODELLER and distance restraints have also been compared, where it was found possible to find out the best model by means of pseudo-energies. Two general conclusions can be elicited from this work: (1) on a given set of putative models it is possible to distinguish among them the one closest to the crystallographic structure, and (2) within a given structure it is possible to find by means of pseudo-energies those regions that have been defectively modelled. PMID- 10702929 TI - [When the cloak of history beckons--ophthalmology in changing times]. PMID- 10702928 TI - Theoretical investigation of substrate specificity for cytochromes P450 IA2, P450 IID6 and P450 IIIA4. AB - Three-dimensional models of the cytochromes P450 IA2, P450 IID6 and P450 IIIA4 were built by means of comparative modeling using the X-ray crystallographic structures of P450 CAM, P450 BM-3, P450 TERP and P450 ERYF as templates. The three cytochromes were analyzed both in their intrinsic structural features and in their interaction properties with fifty specific and non-specific substrates. Substrate/enzyme complexes were obtained by means of both automated rigid and flexible body docking. The comparative analysis of the three cytochromes and the selected substrates, in their free and bound forms, allowed for the building of semi-quantitative models of substrate specificity based on both molecular and intermolecular interaction descriptors. The results of this study provide new insights into the molecular determinants of substrate specificity for the three different eukaryotic P450 isozymes and constitute a useful tool for predicting the specificity of new compounds. PMID- 10702930 TI - [B-scan is the procedure of choice in optic disk drusen]. PMID- 10702931 TI - [Cataract surgery in Great Britain]. PMID- 10702932 TI - [New strategies in retinoblastoma therapy]. PMID- 10702933 TI - [Controversy about cataract operation in diabetic patients]. PMID- 10702934 TI - [Recurrent erosions of the cornea--possible cause? how to manage?]. PMID- 10702935 TI - [Contrast sensitivity and lighting in night time traffic]. PMID- 10702936 TI - [Planning sample size in ophthalmologic studies]. AB - An essential aspect in the cooperation of clinic and biometry consists in designing of studies, e.g. during the preparation of grant applications or for review by official drug surveillance institutions. A central aspect in study planning is the design-adequate and well-documented prediction of sample size, which should be recommended for any intended study. Based on several examples for sample size planning in study designs, which are of common relevance for ophthalmology, guidelines are derived to enable clinical researchers to perform sample size planning on their own. The latter can be based on the various available software packages for sample size prediction. PMID- 10702937 TI - [Merkel cell tumor of the eyelids: review of the literature and report of 2 patients]. AB - BACKGROUND: The Markel cell carcinoma is a rare malignant skin tumor. The tumor was first described in 1972 by Toker and he named it trabecular carcinoma. We had the opportunity to treat two patients with a Merkel cell carcinoma of the upper and lower eyelid and reviewed the literature using the "Medline" database concerning Merkel cell carcinomas of the ocular adnexa. HISTORY AND SIGNS: A 76 year-old female patient was referred to our hospital because of an inflammatory tumor of the left upper eyelid present for two months. A 91-year-old male patient noticed for four weeks a painless itching lesion at the left lower eyelid. The remaining ophthalmologic examination in these patients was unremarkable. THERAPY AND OUTCOME: Both tumors were excised. Histological and immunohistochemical examination verified a Merkel cell carcinoma in both cases. The 76-year-old female patient exhibited no recurrent tumor after a follow-up of 18 months. The 91-year-old male patient had a recurrent tumor inferior to the temporal lower eyelid 7 months after tumor excision, however, lymph node metastasis is not present as of yet. The patient underwent radiation therapy with cobalt of the left orbit with a total dose of 60 Gy. CONCLUSION: Merkel cell carcinomas can occur everywhere on the skin; the eyelids are affected in 10% of all cases. Best histochemical markers are cytokeratin 20 and neurospecific enolase. A review of the literature revealed 31 patients with Merkel cell carcinomas involving the eyelids. Female patients were more often affected than male patients. The upper eyelid was more frequently involved than the lower eyelid. Recurrent disease is frequent. PMID- 10702938 TI - [Carcinoma-associated retinopathy: a review with clinical examples]. AB - Cancer-associated retinopathy (CAR) is a rare paraneoplastic syndrome. In this survey we report about two further patients with CAR, who were referred to the University Eye Hospital of Tuebingen within a few months. The most common primary tumor associated with CAR is small cell carcinoma of the lung. Case reports about rhabdomyosarcoma, carcinoma of the endometrium, prostate and mamma were also described. The exact pathogenesis of CAR is still unknown. Specific autoantibodies were found against the photoreceptor protein recovering (23-kd retinal CAR antigen). However, this reaction is not present in all patients, and probably other antigens are also involved. Most of the patients experience symptoms of CAR before the primary tumor is detected. Besides glare sensitivity and flashing lights, a rapidly progressive, often asymmetric visual loss may occur. Although paracentral and mid-peripheral scotomas can be found frequently, visual field defects are often quite heterogeneous. Typically, the responses in the electroretinogram (ERG) are markedly reduced, but normal ERGs were also described. The fundus picture in CAR shows sheathing of the retinal vessels, narrowing of the arterioles and clumbing of the retinal pigment epithelium. The prognosis is poor. Frequently there is progression to bilateral loss of vision within a few months. Treatment of the primary tumor does not seem to alter the ocular prognosis. Systemic corticosteroids may be helpful in some patients. Nevertheless, no proven therapeutic regimen is currently available. PMID- 10702939 TI - [Complications after external retinal surgery in pseudophakic retinal detachment- are scleral buckling operations still current?]. AB - BACKGROUND: After conventional buckling procedures for pseudophakic retinal detachment the primary reattachment rate is 60 to 90% according to literature. Primary vitrectomy may be advantageous in these cases. The purpose of this study was to investigate the spectrum and role of potential side effects like anisometropia and diplopia, to find arguments for or against primary vitrectomy. Also risk factors for pseudophakic retinal detachment were analyzed in our study group, because they may be different from former studies, which were mostly undertake at the time of ec cataract extraction. PATIENTS AND METHODS: The data from 115 patients (120 eyes) with pseudophakic retinal detachment, who undergone buckling procedure between 1991 and 1996 were retrospectively reviewed. We analyzed the retinal reattachment rate, choroidal detachment, postoperative visual acuity, refraction, occurrence of diplopia, anisometropia and metamorphopsia. RESULTS: The primary retinal reattachment rate was 83.3%. There was a retinal redetachment rate of 16.7%. The primary PVR-rate was 4.2%. After the first reoperation an reattachment rate of 91.6% could be achieved, after the second one an overall rate of 95%. A choroidal detachment occurred in 29.2%. Postoperatively (26.5 months +/- 17.2) we found a cellophane maculopathy in 26.5% with consecutive metamorphopsia in 12.1%. 13 patients (15.9%) complained about diplopia, which had to be corrected by operation in 2 cases and prismatic glasses in 4 cases. Anisometropia could be deserved in 16.4%. The buckling procedures caused refractive changes of -1.80 dpt +/- 1.78 after an encircling band and +0.38 dpt +/- 1.01 after a plombage (average -1.05 +/- 1.86). 20% of patients treated with an encircling band developed anisometropia and only 8.3% of patients with a plombage. Postoperatively the visual acuity raised significantly from 0.3 to 0.6 (median). Best visual recovery could be observed in patients, who underwent only one operation. CONCLUSIONS: The primary retinal reattachment rate in pseudophakic retinal redetachment after conventional buckling procedures in our patients are comparable to those in the literature. In spite of phacoemulsification the risk-factors for pseudophakic retinal detachment has not changed. A considerable number of patients complained about postoperative complications like anisometropia and diplopia, which could be avoided by primary vitrectomy. To compare anatomic and functional results of both procedures a randomized prospective study should be undertaken. PMID- 10702940 TI - [Clinical results of intravitreal administration of tissue-type plasminogen activator (tPA) and gas for removal of subretinal hemorrhage in senile macular degeneration]. AB - BACKGROUND: Subretinal hemorrhage in age related macular degeneration (AMD) usually causes acute visual loss and is associated with poor visual prognosis. In order to prevent retinal damage and to perform laser treatment of the underlying choroidal neovascularization (CNV) the subretinal hemorrhage has to be removed from the macular region. This could be achieved by intravitreal injection of tissue plasminogen activator (tPA) and gas. PATIENTS AND METHODS: In 8 consecutive patients, suffering from a massive macular hemorrhage (duration of visual problems: mean 9 days), tissue plasminogen activator (tPA) (40 micrograms in 400 microliters BSS) and SF6-gas (0.75 ml) was transsclerally injected into the vitreous cavity to achieve liquification and displacement of the hemorrhage. RESULTS: In all patients liquification and displacement of the hemorrhage out of the macular region was achieved during follow up. During the first week after operation a significant increase of visual acuity was noticed in all patients, however ophthalmoscopically there was just little reduction of the hemorrhage in the foveolar area. After successful removal of the blood the choroidal neovascularization was treated successfully by laser coagulation in one patient. No laser treatment was performed in the other patients because of the subfoveal location of the neovascularisation or because of disciform scar. Visual acuity increased 4 lines after surgery. In one case the procedure was complicated by a persistent vitreous hemorrhage and vitrectomy had to be performed in another patient due to an endophthalmitis. CONCLUSION: Intravitreal injection of tPA assisted gas displacement of subretinal hemorrhage due to AMD leads to a significant increase of visual acuity during the first week after operation. Although a nearly complete removal of the hemorrhage out of the macular area could be achieved, it was difficult to differentiate this from the spontaneous course. Laser photocoagulation could be performed in only few cases. PMID- 10702941 TI - [Autoregulation of retinal arterioles in patients with diabetes mellitus and normal probands]. AB - BACKGROUND: Measurement of myogenic autoregulation of retinal arterioles was demonstrated by the use of the Retinal-Vessel-Analyzer (Carl Zeiss, Jena). The purpose of the presented study was to find a significant difference of this myogenic response in a group of healthy individuals vs. a group of patients with pathologic conditions. PATIENTS AND METHODS: As a group of patients with known pathology in microcirculation patients with type 1 diabetes were chosen. By isometric exercise an identical rise in mean arterial blood-pressure was provoked in 20 patients with type-1 diabetes and 20 matched healthy volunteers. The myogenic response of retinal arterioles was measured in both groups by the use of the Retinal-Vessel-Analyzer. The Wilcoxon test was used for statistical analysis within the two groups whereas both groups were compared with each other by the use of the Mann-Whitney test. RESULTS: Having the same age and same blood pressure rise (p = 0.624) a significant better myogenic response was found in the healthy subjects vs. the diabetic group (p = 0.008). In the diabetic group no correlation was found between myogenic response and duration of diabetes (p = 0.982) or HbA1c values (p = 0.83). CONCLUSIONS: In this study significant loss of autoregulation in patients with type 1 diabetes is demonstrated. By the use of the Retinal-Vessel-Analyzer noninvasive testing of the function of autoregulation of retinal arterioles is possible. This method might prove to be of great value in early detection of diabetic vessel pathology. PMID- 10702942 TI - [Adamantiadis-Behcet syndrome: fluorescein angiography and choroid ischemia]. AB - BACKGROUND: Adamantiadis-Behcet's disease is a chronically progressing multisystemic disorder. The underlying disease mechanism is an obliterative vasculitis of unknown etiology. Main clinical symptoms are oral and genital aphthous ulcers and intraocular inflammations. Additionally, cutaneous, rheumatoid, neural, gastrointestinal, or cardiovascular manifestations may be observed. Diagnosis is based on clinical features since currently no specific laboratory tests or pathognomonic histopathological features are available. Ocular changes may provide important diagnostic clues to the systemic disease. PATIENTS AND METHODS: In a retrospective fashion 196 patients of the Department of Ophthalmology of the University of Erlangen-Nurnberg between 1988 and 1998 with retinal vasculitis seen by fluorescence angiography were evaluated according to the diagnostic criteria of the "Behcet's Disease Research Committee of Japan". RESULTS: Among 196 patients there were 12 patients with Adamantiadis-Behcet's disease. Apart from retinal vasculitis, angiographic features included capillary dropout in 64% (9/14), swelling of the optic disc in 79% (11/14) and irregularly delayed areolar filling of choriocapillaris in the early phase of fluorescence angiography in 43% (6/14) of eyes. There was no statistically significant relationship between severity of the systemic disorder and the activity of the ocular disease. CONCLUSION: Apart from retinal vasculitis, 43% of eyes in patients with Adamantiadis-Behcet's disease presented a delayed choroidal filling in fluorescence angiography as a sign of choroidal involvement of occlusive vasculitis. We observed leakage of fluorescein from the optic disc, which could be due to a secondary inflammation of the ciliary circulation. Inflammatory involvement predominantly of choroidal vessels, as visualized in fluorescence angiography, may be a diagnostic lead in Adamantiadis-Behcet's disease. PMID- 10702943 TI - [Vertical vergence in convergence]. AB - PURPOSE: To report on a 7-year-old boy with a small left-over-right deviation ( VD) which increased when the head was tilted to the left shoulder and during convergence. METHODS: The squint angles were measured by the unilateral and alternate prism cover test at distance and near fixation when the head was in ortho-position and when it was tilted. RESULTS: At distance fixation (D) there was a latent deviation of-VD 3 degrees. With near fixation (N) at 0.3 m the vertical phoria increased to-VD 18 degrees. The angle of deviation was not influenced by (N) convex lenses in front of the fixating eye despite an adequate dis-accommodation. The-VD was fairly comittant in right and left gaze. At 45 degrees head tilt to the right shoulder the-VD decreased to (D) 2 degrees and (N) 12 degrees. At 45 degrees head tilt to the left shoulder the-VD increased to (D) 18 degrees and (N) 26 degrees. A dissociated vertical deviation was excluded by the dark red glass test and by the reversed fixation test. CONCLUSION: The disturbance can be explained by a, presumably congenital, supranuclear misinnervation and has to be differentiated from other types of vertical deviation. PMID- 10702944 TI - [Bilateral papillary edema in cerebrospinal syphilis]. AB - BACKGROUND: Nowadays luetic infections are rarely seen by ophthalmologists. We report on an immunocompetent ophthalmologically asymptomatic patient with bilateral papilledema due to perineuritis optici in lues cerebrospinalis. PATIENT: A 47-year old female patient presented with presbyopic complaints. Additionally she reported occasional dizziness with nausea and hearing loss with tinnitus. Visual acuity measured 16/20. There was a bilateral prominent optic disc with indistinct margins and papillary hemorrhagies on the right side and corresponding enlargement of the blind spot in the visual field. Echography revealed bilateral optic drusen. Serological examination suggested lues (TPHA 1:5120, IgM-FTA-Abs-Test 1:320, Cardiolipin 1:640). Cerebrospinal fluid examination indicated an inflammatory process in the CNS without proof of an autochthonous antibody production. CONCLUSION: Even nowadays lues cerebrospinalis must be suspected in patients with bilateral papilledema without visual loss. The ophthalmologist holds an important diagnostic position, because adequate treatment is able to prevent disease progression. PMID- 10702945 TI - [Eye injury caused by superglue]. AB - BACKGROUND: Eye lesions caused by superglue (instant glue) are relatively rare, and hence therapeutical experiences are limited. PATIENT: A female intended to treat a mild conjunctivitis by herself with anti-inflammatory eye drops. Inadvertently she took a tube of instant glue which then filled most of the conjunctival sac. After intensive rinsing the hardened glue was mechanically removed as far as possible. Remaining glue was spontaneously rejected within a few hours. After four days the resulting subtotal corneal erosion had healed, and visual function had returned almost to normal. CONCLUSIONS: Inspite of the dramatic, acute aspect the clinical course after superglue lesions is usually benign. Therapeutically, vigorous manipulations seem to be more harmful than useful. Spontaneous rejection of the glue should be awaited instead. PMID- 10702946 TI - [Influence of the presence of hot flashes during menopause on the metabolism of nitric oxide. Effects of hormonal replacement treatment]. AB - BACKGROUND: To asses if the hot flushes during the menopause are associated to alterations in the metabolism of nitric oxide, total antioxidant activity and other biochemical parameters. To evaluate the effect of hormone replacement therapy (HRT) in the metabolism of nitric oxide, total antioxidant activity and biochemistry in menopausal women with and without hot flushes. PATIENTS AND METHODS: Experimental study. This study included 29 healthy menopausal women with hot flushes (12 with HRT) and 20 without them (6 with HRT), 45-55 years old, with duration of menopause ranging from 1 to 5 years. Control group included 14 fertile women. Determinations of estradiol, nitrite-nitrate, total antioxidant activity and basic analytic, were performed before previous and after 4-6 month of HRT therapy. RESULTS: Previously to HRT, hot flushes were associated to smaller level of total antioxidant activity (p < 0.05) in plasma, without differences in nitrite-nitrate concentrations. HRT was associated to increase in total antioxidant activity level and nitrite-nitrate concentrations (p < 0.05) in menopausal women, with and without hot flushes. CONCLUSIONS: Hot flushes during the menopause indicate a high level of oxidative stress and a higher cardiovascular risk. HRT decreased the oxidative stress level and increased the nitric oxide derivate metabolites in menopausal women with and without hot flushes. PMID- 10702947 TI - [Clinical and analytic diagnostic evaluation of deep venous thrombosis of the lower limbs]. AB - BACKGROUND: Diagnosis of deep-vein thrombosis (DVT) of lower limbs has changed in recent years. The objective of our study was to analyze the diagnostic accuracy of a combination of clinical and epidemiological data and the D-Dimer plasma levels in this entity. PATIENTS AND METHODS: Clinical (symptoms and signs) and epidemiological data, personal and family history, and D-dimer plasma levels or positivity were reviewed, on the admittance, in 108 patients to whom a phlebography was performed due to a suspected DVT. RESULTS: Phlebography was positive in 76 cases (70.37%). Logistic regression analysis determined a prediction model of the diagnostic of DVI including a combination of both D-dimer plasma levels or positivity and pain along the deep venous involved area. CONCLUSION: Combination of D-dimer testing and pain along the distribution of the deep venous area is useful as an initial diagnostic approach to the DVI of the lower limbs. PMID- 10702948 TI - [Clinical manifestations of heterozygote familial hypercholesterolemia in Spain. Study of 301 cases from the central and northern areas]. AB - RATIONALE: To characterize clinical manifestations of familial hypercholesterolemia (FH) in Spain. PATIENTS AND METHODS: A group of 301 cases of FH from central and north regions of Spain. RESULTS: With a mean (SD) cholesterol level of 346 (58) mg/dl, only 7.6% of the patients have xanthomas and 20% ischaemic heart disease. 51% have a familial history of ischaemic heart disease. CONCLUSIONS: Different from the results of literature, xanthomas are very infrequent in FH in Spain, so diagnosis should be suspected from other data. The high prevalence of familial history of ischaemic heart disease supports the usefulness of this feature as a marker for diagnosis and prevention. PMID- 10702949 TI - [Hot flashes, menopause, nitric oxide and hormone replacement therapy: "one for all and all for one"]. PMID- 10702950 TI - [New directions in the diagnosis of deep venous thrombosis of the lower limbs]. PMID- 10702951 TI - [Identification of 2 families with hereditary nonpolyposis colonic cancer (HNPCC) and the Amsterdam criteria. Relevance of the genealogic tree and follow-up]. AB - Hereditary nonpolyposis colorectal cancer (HNPCC) diagnosis is based either on the so-called "Amsterdam 1 criteria" or "Amsterdam 2 criteria", which includes extracolonic neoplasms associated with Lynch II syndrome. Many families are suspected of having a hereditary predisposition to cancer and may benefit from close surveillance. We describe a family (family 1) with suspected HNPCC at the beginning who fulfilled the Amsterdam 1 criteria over the course of its follow up. We also describe an Amsterdam 2 family (family 2) with a very young affected individual. Both of them received genetic counseling and screening recommendations. A total colonoscopy was done to an asymptomatic member of family 1 and he was diagnosed with an early-stage colon cancer. He underwent subtotal colectomy because of the high risk of metachronous lesion. Screening recommendations must be the same in Amsterdam 2 families as in Amsterdam 1. Both families show the importance of considering the family history when hereditary criteria are suspected. PMID- 10702952 TI - [Report on travel to the Albert Schweitzer Hospital in Haiti]. PMID- 10702953 TI - [Training policies in biomedical research in Spain]. PMID- 10702954 TI - [Interferon and plasma lipids]. PMID- 10702955 TI - [Hyperlipemia and glycoprotein IIIa polymorphism]. PMID- 10702956 TI - [Reduction of hospitalization in community-acquired pneumonia by the use of therapeutic strategy based on short-term mortality]. PMID- 10702957 TI - [Participation of storage mites in mite-induced allergic respiratory diseases]. PMID- 10702958 TI - [Recurrent aseptic meningitis associated with amoxicillin-clavulanic acid]. PMID- 10702959 TI - Combination acetaminophen and doxapram potentiated hepatotoxicity in mouse primary cultured hepatocytes. AB - Doxapram-induced potentiation of acetaminophen-induced reductions in cell viability and apoptosis was examined in mouse primary cultured hepatocytes. Loss of viability following exposure of acetaminophen and/or doxapram in cultured hepatocytes was assessed by monitoring [3H]-thymidine incorporation and mitochondrial activity, and apoptosis of hepatocytes was determined by nuclear microscopic observation and from detection of a ladder-like DNA fragmentation pattern in agarose gel electrophoresis. The combination of acetaminophen (5 mM) and doxapram (10, 20, 50 or 100 microM) potentiated the reduction in cell viability and increased lipid peroxide levels of hepatocytes. Hepatocytes exposed for 24 h to acetaminophen (5 mM) plus doxapram (100 microM) showed atrophy of nuclei including chromatin condensation and a ladder-like DNA fragmentation pattern characteristic of apoptosis. Antioxidant (N-acetylcysteine), iron chelator (deferoxamine), intracellular calcium ion chelator (quin 2-AM), endonuclease inhibitor (aurintricarboxylic acid) and poly (ADP-ribose) polymerase inhibitor (3-aminobenzamide) all improved the viability of cells and eliminated the ladder-like DNA fragmentation in cells exposed to acetaminophen plus doxapram. In conclusion, the combination acetaminophen and doxapram potentiated the reduction in cell viability and apoptosis in mouse primary cultured hepatocytes. We suggest that careful observation for hepatotoxicity is recommended when acetaminophen and doxapram are prescribed simultaneously. PMID- 10702960 TI - Effect of amikacin on cytotoxic activity and hydrophobicity of Acinetobacter baumannii. AB - Effect of amikacin at suprainhibitory (2 or 4 x MIC) or supra-subinhibitory concentrations (2 or 4 x MIC + 0.2 x MIC) on bacterial growth, cytotoxicity and cell surface hydrophobicity of three Acinetobacter baumannii strains was studied. Amikacin at suprainhibitory concentrations induced postantibiotic effects (PAEs; suppression of bacterial growth after short time exposure of bacteria to the antibiotic) against all A. baumannii strains. PAEs ranged from 1.2 to 2.9 h (2 x MIC) and 3.5 to 6.3 h (4 x MIC). Supra-subinhibitory concentrations of amikacin (2 x MIC + 0.2 x MIC) manifested a more significant delay of bacterial regrowth (PA SMEs) for two strains (6.6 or 7.5 h) in comparison with PAEs. One strain under these conditions as well as all strains treated with amikacin at 4 x MIC + 0.2 x MIC did not show any regrowth. Amikacin at all concentrations tested significantly reduced cytotoxic activity of A. baumannii evaluated on alveolar epithelial type II cells. Survival of type II cells after application of antibiotic-treated A. baumannii was in the range of 88 to 101% of the control cells. Cell surface hydrophobicity of amikacin-treated bacteria was practically unchanged varying between 94 and 100.9% as compared to the controls. PMID- 10702961 TI - Comparison of the isolated human and guinea pig gallbladder strip models in the assessment of antispasmodic drugs. AB - The objective of this study was to compare the antispasmodic activities of atropine, verapamil, (-)scopolamine n-butyl bromide and propinox in the isolated human and guinea pig gallbladder strip models. Concentration-response curves for each of the agents were obtained in both models following administration of carbachol. Atropine was the only drug to show marked activity in the guinea pig gallbladder model (ED50 = 2.75 x 10(-7) M); the remaining drugs elicited less inhibition of a similar order of magnitude (ED50 = 1.65 x 10(-5), 4.18 x 10(-6) and 2.71 x 10(-5) M for verapamil, [-]scopolamine n-butyl bromide and propinox, respectively). In contrast, results obtained from the human gallbladder strip model revealed differences among the drugs (ED50 = 5.03 x 10(-8), 1.34 x 10(-6), 6.63 x 10(-6) and 5.45 x 10(-5) M for atropine, propinox, verapamil and [ ]scopolamine n-butyl bromide, respectively). Based on these results, propinox showed a relative potency in the human gallbladder that was 20.22-fold higher than that in the guinea pig model followed by atropine (5.47-fold) and verapamil (2.49-fold), whereas (-)scopolamine n-butyl bromide was 0.07 times more potent in the guinea pig model. Regression analysis of ED50 values showed a lack of correlation between the two models (r = 0.44). Considering interspecies variations, further studies in human tissues are needed to evaluate the efficacy of antispasmodic drugs. PMID- 10702962 TI - Effects of epinastine on the antitussive and rewarding effects of dihydrocodeine in mice. AB - The effects of chlorpheniramine and epinastine on dihydrocodeine were examined in mice. Orally administered dihydrocodeine (3-30 mg/kg) dose-dependently inhibited the number of capsaicin-induced coughs. The dose-dependent antitussive effects of dihydrocodeine were enhanced by each corresponding dose of chlorpheniramine or epinastine delivered at a ratio generally similar to that found in over-the counter antitussive preparations (dihydrocodeine:histamine H1 antagonist = 3:1). The ED50 value of dihydrocodeine in combination with chlorpheniramine was nearly the same as that for dihydrocodeine in combination with epinastine. On the other hand, while combination treatment with dihydrocodeine (3 mg/kg i.p.) and chlorpheniramine (1 mg/kg s.c.) significantly potentiated place preference, no potentiation was observed with the combination of dihydrocodeine (3 mg/kg i.p.) and epinastine (1 mg/kg s.c.). These results suggest that epinastine may be a useful constituent opioid-containing antitussive preparation that would not enhance the potential for psychological dependence. PMID- 10702963 TI - Effects of the calcium antagonists verapamil and nitrendipine on carbamazepine withdrawal. AB - This study was aimed at examining the effects of two frequently used Ca2+ antagonists, nitrendipine and verapamil, on withdrawal after cessation of long term treatment with the anticonvulsant drug carbamazepine in rats. The 48-h interruption of long-term (21 days) carbamazepine treatment led to the appearance of withdrawal characterized by increases in seizure intensity, the percentage of rats with tonic seizures and mortality. Oral treatment with the two calcium antagonists in combination with carbamazepine abolished the signs of carbamazepine withdrawal. Seizure intensity, the percentage of rats with tonic seizures and mortality in the groups treated with the combinations of carbamazepine + verapamil and carbamazepine + nitrendipine were significantly lower than those of the group of rats treated with carbamazepine alone. In conclusion, some Ca2+ antagonists could attenuate the manifestations of anticonvulsant withdrawal and thus could be used as adjuvants in long-term anticonvulsant therapy. PMID- 10702964 TI - A phase I clinical trial of dextromethorphan in intractable partial epilepsy. AB - We conducted an open-label pilot study of dextromethorphan (DM) in intractable partial epilepsy with the following objectives: a preliminary evaluation of the drug's safety and efficacy in the epileptic patient and a definition of a concentration range which can be safely achieved in future studies. Sixteen patients with drug-resistant, localization-related epilepsies entered the trial. After an 8-week baseline period, DM was added to the existing antiepileptic drugs at a dose of 40 and 50 mg every 6 h (160 and 200 mg/day). Each treatment period lasted 8 weeks. Seizure control improved after administration of DM, especially in the group of intermediate and slow metabolizers. Two patients, however, experienced increased seizure frequency and withdrew from the study. Adverse effects during DM administration were mild and transient. DM was well tolerated even in patients with high plasma levels of the drug (up to 15020 ng/dl). Our results indicate that DM is safe and effective in the treatment of comedicated patients with intractable partial epilepsies. PMID- 10702965 TI - Effect of grapefruit juice on the pharmacokinetics of theophylline in healthy male volunteers. AB - The effect of grapefruit juice (GFJ) on the pharmacokinetics of a single dose of theophylline was examined in an open crossover study. Healthy male volunteers were given sustained release theophylline (300 mg) along with 300 ml of either water or GFJ. Blood samples (2 ml) were collected at different time points from 0 to 48 h. Plasma was assayed for theophylline by HPLC and various pharmacokinetic parameters were calculated. Theophylline levels were lower at all time points with GFJ coadministration as compared to water but were significantly lower only during the absorption phase from 1 to 4 h. Although the pharmacokinetic parameters were not significantly different between the two groups, all values were reduced except Tmax which was slightly increased. The results indicate that patients may be advised not to consume GFJ when taking slow-release theophylline and monitoring of plasma theophylline levels in patients consuming GFJ might be helpful in better management of patient care. PMID- 10702966 TI - Study of the effects of aging on macromolecular synthesis in various organ systems using microscopic radioautography. AB - In order to elucidate the effects of aging on macromolecular synthesis such as DNA, RNA, proteins, glucides and lipids in various organ systems of experimental animals and humans, systematic studies using light and electron microscopic radioautography in various organ systems including skeletal, muscular, digestive, respiratory, urinary, reproductive, endocrine, circulatory, nervous and sensory systems were studied after incorporation with macromolecular precursors. The experimental animals used were mainly ddY strain mice from embryo to postnatal days 1 and 3, weeks 1 and 2 or months 1, 2 and 6 months up to 1 and 2 years senescent stages. Animals were injected with [3H]-thymidine for DNA, [3H]-uridine for RNA, [3H]-amino acids for proteins, [3H]-glucose, [3H]-glucosamine and [35S]O4 for glucides, [3H]-glycerol for lipids and some low molecular target tracers such as hormones, inorganic substances and drugs. Results demonstrate that these precursors when incorporated into various cell types in various organs showed specific patterns of macromolecular synthesis as observed in perinatal to juvenile, mature and senescent stages. These effects of aging could answer some of the questions as to how but not why we get old. PMID- 10702967 TI - TEM/CBED determination of strain in silicon-based submicrometric electronic devices AB - The convergent beam electron diffraction technique (CBED) of the transmission electron microscopy (TEM) has been employed to determine the strain distribution along a cutline parallel to the padoxide/Si interface in a 0.80 micron wide recessed-LOCOS structure. The values of the components of the strain tensor so obtained have been compared with those computed by two simulator codes. It has been found that both the LOCOS morphology and the strain distribution deduced from TEM images and TEM/CBED patterns, respectively, were in agreement with the simulation results, if some oxidation-related parameters were modified. PMID- 10702968 TI - Dynamical simulation of LACBED patterns in cross-sectioned heterostructures AB - The Large Angle Convergent Beam Electron Diffraction (LACBED) technique has been applied to determination of the tetragonal mismatch in coherent Si/Si1-xGex/Si heterostructures. Two-dimensional (2D) dynamical simulation of the LACBED patterns has been performed and compared with the corresponding experimental ones. A good agreement is found in the whole simulated area, particularly as regards the splitting of the Bragg contours, due to the strain field present in the TEM cross-sections. PMID- 10702969 TI - Antiphase disorder in GaAs/Ge heterostructures for solar cells AB - Antiphase disorder in metal organic vapour phase epitaxy grown GaAs/(100)Ge heterostructures has been studied both in as-grown materials and in GaAs solar cells by chemical etching, transmission electron microscopy, and cathodoluminescence. All the samples are single domains at the surface due to the self-annihilation of antiphase domains whose size decreases as the misorientation angle increases. Completely antiphase domain-free epitaxy has been achieved for substrate miscuts greater than 3 degrees off towards [111]. A reversal in sublattice location has been found in the GaAs layers varying the misorientation angle and the growth temperature. A model to explain this result has been proposed based on the role of surface steps in the nucleation process. Strong interaction between antiphase boundaries and misfit dislocations has been found in all the heterostructures. In solar cells antiphase domains have been observed in high densities in the initial layer of GaAs deposited on Ge. The successful realisation of high efficiency solar cells is due to the overgrowth of these domains by single phase material over most of the wafer area. PMID- 10702970 TI - TEM characterisation of porous silicon AB - In this work, the results of a structural investigation by transmission electron microscopy of porous silicon and porous silicon-based devices is reported. This investigation covers a wide range of porous silicon materials going from photoluminescent micro- or meso-porous silicon to macro-porous layers as well as some recent applications of this material for the fabrication of porous silicon and Si/SiO2 superlattices, suspended thick membranes and gas sensor devices. It appears that the structural investigation technique here employed, is a fundamental tool for the optimisation of the technological processing of this material. From the results reported in this work, a quite comprehensive view of the potentialities and limitations of the applications of porous silicon in the field of optical and gas sensor devices should also emerge. Finally, some solutions to the problems usually met during the technological processing of this material are proposed. PMID- 10702971 TI - Electron microscopy of reverse biased p-n junctions AB - The aim of this paper is to present and discuss the recent results obtained in the investigation of reverse-biased p-n junctions by means of the out-of-focus method. It will be shown how the interpretation of the experimental images is not in agreement with the expectations based on the bulk p-n junction theory. The possible causes of this discrepancy will be discussed: among them a significant reason could be the finite specimen thickness with the associated surface states, which influences the width and shape of the depletion layer in the thinned specimen. PMID- 10702972 TI - High resolution transmission electron microscopy to study very thin crystalline layers buried at an amorphous-crystalline interface AB - Structure characterisation of interfaces is a field of widespread application of high resolution transmission electron microscopy for its very high spatial resolution. Specimen thickness and electron optical condition have a deep influence on the high resolution electron transmission microscopy image contrast. Hence, in many cases, the real structure of the sample can be understood from experimental images only by comparison with the relevant simulation. Moreover, the understanding of the contrast variation of a few A at an interface is a task in which even the use of simulation could not produce an unequivocal solution of the experimental result. In this paper high resolution transmission electron microscopy image simulations show that two monolayers of crystalline material buried at an amorphous-crystalline interface can be successfully revealed and interpreted. The simulated images reproduce the experimental results as obtained from the Al/Si-As/n-GaAs (001) heterostructure. PMID- 10702973 TI - Correlation between shape and electronic states in nanostructures AB - We show how the electronic states of quantum wires and quantum dots can be evaluated exactly starting from the profile of the nanostructure observed by transmission electron microscopy, scanning tunneling microscopy and atomic force microscopy. The calculated quantization energies reproduce the energy position of the luminescence resonances in the optical spectra of different samples, without fitting parameters. PMID- 10702974 TI - Synthesis and structural characterisation of CdS nanoparticles prepared in a four components "water-in-oil" microemulsion AB - Nanostructured semiconductor particles are currently under intense investigation because of their enhanced photoreactivity and photocatalytic properties due to the quantum-size effect and the dependence of the photophysical and photochemical properties on their size as it approaches the exciton diameter. This increasing interest has led to the development of several synthetic procedures to prepare and stabilise uniform crystallites. In this paper, we report a novel synthetic pathway to obtain cadmium sulphide (CdS) nanoparticles in a quaternary "water-in oil" microemulsion formed by a cationic surfactant cetyltrimethylammonium bromide (CTAB), pentanol, n-hexane and water. The synthesis of CdS in this system is achieved by mixing two microemulsions containing Cd(NO3)2 and Na2S, respectively. The nanocrystals have been characterised by using UV--visible spectroscopy and Transmission Electron Microscopy to investigate the influence of various parameters of the particles' formation and stability in solution. Capping of nanoparticles with suitable organic molecules has been performed in order to increase their stability and afford solubility in a wide range of solvents. PMID- 10702975 TI - Electron microscopy of life-tested semiconductor laser diodes AB - Electron Microscopy on life-tested 980 nm SL SQW InGaAs/AlGaAs laser diodes is able to find and analyze lattice defects responsible for the detected failures. Anyway, the origin and evolution of those defects remains questionable. Only the comparative analysis of life-test measurements, EBIC-FIB/TEM images, and charge transport physics is able to point out a coherent framework for complete decoding of the failure kinetics. Minority-carrier diffusion and their enhanced recombination at defective lattice points are indicated, as the energy supply required for defect reaction and growth. The rules of charge diffusion drive both the reaction model, the interpretation of EBIC images and the expected electrical and optical effects. Strain release at the ultimate propagation of defects into the strained InGaAs quantum layer is then easily related to the final state of the failed devices. PMID- 10702976 TI - Low-temperature spectrally resolved cathodoluminescence study of degradation in opto-electronic and microelectronic devices AB - This study reports on the microcharacterization of devices for optoelectronic and for microelectronic applications using low temperature (T = 5 and 77 K) spectrally resolved cathodoluminescence (SCL). The mechanisms leading to compositional inhomogeneities in the regrowth regions of InP-based butt-coupled laser-waveguide devices for semiconducting optical amplifiers (SOAs) and for defect generation in the active and cladding layers of GaAs based pump lasers for erbium-doped optical fibre amplifiers (EDFAs) were studied. Beryllium outdiffusion in the base regions of GaAs-based heterojunction bipolar transistors (HBTs) after bias ageing was also studied. By comparing the CL results with TEM, SIMS and HRXRD studies and with the existing literature, the observed growth and operation induced defects were attributed, respectively, to the following mechanisms: recombination-enhanced defect glide (REDG) in the pump lasers, recombination enhanced impurity diffusion (REID) in the HBTs and electrostatically induced growth flux instabilities in the butt-coupled laser waveguide devices. PMID- 10702977 TI - Computation of the strain field generated by dislocations with a position dependent Burgers' vector distribution AB - A new phenomenon of strain relaxation will be presented. In a series of InxGa1 xAs graded composition buffer layers grown on well cut (001) GaAs substrates, a curvature of the epilayer lattice has been found, i.e. a tilt of the epilayer lattice orientation with respect to the substrate which varies coherently along the sample surface on the scale of several mm. The most recent data analysis performed on a buffer layer compositionally graded with a six-step profile shows also a thickness functional dependence of the curvature. The epilayer lattice curvature has been attributed to a coherent lateral distribution of the Burgers' vectors. An analytical model has been developed in the framework of the continuum elasticity theory to compute the related strain field. The results show small but unexpected contributions to the parallel strain. PMID- 10702978 TI - Metastability of Si1-yCy epilayers under 2 MeV alpha-particle irradiation AB - In this work we present some recent results concerning the alpha-particles irradiation of Si1-yCy alloy epitaxially grown on silicon. The study of the damage process is interesting because of the extensive use of backscattering technique as a tool of characterisation of this kind of materials and because of the possibility of adding information about the transformations that this metastable material undergoes. We point out that the irradiation damage process causes a change in the material structure different from that due to the thermal treatments. The irradiation damage occurs at a rate much higher than in Si, however it involves only a silicon atom fraction that appears to be proportional to the substitutional carbon content. PMID- 10702979 TI - Nitridation of gate and tunnel oxides employed in CMOS-ULSI technology AB - Nitridation treatments are very important in CMOS technology because of their capability of improving the gate and tunnel oxide reliability. In this work we report on N2O and NO annealing of pre-oxidised samples showing physical and electrical characteristics of the thin oxides. The difference between the physical behaviours of N2O and NO oxides is evidenced and related to their different electrical properties. PMID- 10702980 TI - Morphological and structural effects of excimer laser treatment of amorphous silicon AB - The excimer laser irradiation of thin film amorphous silicon (a-Si) precursors on glass is a suitable method for obtaining high-performance polycrystalline silicon (p-Si) active layers for devices and circuits. By changing the experimental conditions, the recrystallization method generates a variety of microstructures that have direct impact on the material performance. An additional reason for microstructural characterization is introduced by the methods for spatially locating the recrystallization nuclei, used in more ergonomic concepts of device fabrication. Metal and SiO2 strip overlayers have been applied here, on a-Si to fix the position of the solidification seeds after laser melting. The control of many aspects of the thin film microstructure can be achieved with a collection of a few inspection techniques like AFM, SEM, EC contrast, TEM, X-ray diffraction (XRD), some of which require preliminary grain decoration treatment, and some do not. The results of different irradiation experiments, are herein illustrated, enlightened by the above characterization techniques, for providing information on surface morphology, grain arrangement, preferred orientation. PMID- 10702981 TI - Self-assembling of In(Ga)As/GaAs quantum dots on (N11) substrates: the (311)A case AB - We have investigated the In(Ga)As island formation, in the Stranski-Krastanov growth mode, on (311)A GaAs substrates. The surface topography of InAs and InGaAs strained epilayers was studied by contact microscopies. The different substrate affects the overgrown island shape. In(Ga)As grown on (311)A gives rise to quantum wire-like islands. Quantum dots (QDs), but with highly anisotropic shapes, are the outcomes of InAs deposition. QD samples were also characterized by photoluminescence (PL) measurements. Correlation between optical and morphological properties was observed. PMID- 10702982 TI - Strain-driven morphology of Si1-xGex islands grown on Si(100) AB - A study has been carried out on the morphology and structure of three-dimensional (3D) SiGe islands grown by molecular beam epitaxy (MBE) on Si(100) substrates. Samples of Si1-xGex alloys have been prepared to investigate the effects either of the alloy composition or of the growth temperature. Atomic force microscopy (AFM) evidenced the growth of 3D islands and transmission electron microscopy (TEM) demonstrated wetting layer growth on Si(100), independently on the deposition conditions. Energy dispersive spectroscopy (EDS) micro-analyses carried out on cross-sections of large Si1-xGex islands with defects allowed a measurement of the Ge distribution in the islands. To the best of our knowledge, these have been the first experimental evidences of a composition change inside SiGe islands. The interpretation of the experimental results has been done in terms of strain-enhanced diffusion mechanisms both of the growing species (Si and Ge) and of small islands. PMID- 10702983 TI - [Essays on modern molecular genetics. Essay 8. Genome diseases and new molecular genetics. Part 3. Cancer is a genome disease. Transcription and cancer (the beginning of the lecture)]. PMID- 10702984 TI - [Molecular characteristics of multiresistant clinical strains of Mycobacterium tuberculosis isolated in Russia]. AB - The efficiency of tuberculosis control programs is largely determined by methods for rapid diagnosis of the agent. In comparison with the traditional methods, new molecular technologies for characterization of mycobacteria appear to be more promising, because the result can be obtained in almost no time. Sixty-five strains of M. tuberculosis isolated in various regions of Russia were investigated. Drug resistance and strain appurtenance of this sample were determined by classical (absolute concentrations method, IS6110-RFLP) and modern molecular genetic methods (detection of mutations in rpo B gene, DRE-PCR). The spectrum of mutations of the rpoB gene associated with rifampicin resistance was evaluated by direct sequencing. Mutations involving codons 531 (62.7%), 526 (18.6%), and 516 (10.2%) of rpoB gene predominated in the studied sample. The studied strains were discriminated into 52 individual strains by IS6110-RFLP and DRE-PCR typing. Analysis of the resultant genetic variants showed the predominance of M. tuberculosis family W. The efficiency of combined approach to screening for M. tuberculosis is discussed. PMID- 10702985 TI - [Changes in the virulence factor expression level in Listeria monocytogenes under various environmental conditions]. AB - Effects of chelators Chelex-100 and activated charcoal on the production of proteins responsible for virulence of Listeria monocytogenes, facultative intracellular parasite were studied. Bivalent cation chelator Chelex-100 stimulates the production of only thiol-dependent hemolysin listeriolysin O. The presence of activated charcoal, a nonspecific chelator, in culture medium stimulated the expression of listeriolysin O and other main virulence factors by increasing the level of their transcription. PMID- 10702986 TI - [Transposition and inheritance of Bordetella Tn-element in Escherichia coli K12]. AB - B. pertussis genetically mobile element TnBp3 integrates the plasmid in E. coli chromosome. During culturing under nonselective conditions the majority of cells of some E. coli strains lose the kanamycin resistance marker, which indicates the instability of TnBp3 inheriting. The stability of inheriting the integrated structure is higher in E. coli cells with recB-21 recC-12 sbcB-2 mutations. The role of RecBC recombination system in extrusion of TnBp3 is discussed. PMID- 10702987 TI - [New rare cutting restriction endonuclease SmII from Streptococcus milleri recognises 5'-ATTTAAAT-3']. AB - New restriction endonuclease (restrictase) Smil of type II was detected in the bacterial strain Streptococcus milleri. Cellular lysate enzyme cut T7 and adenovirus-2 DNAs at site 5'-ATTT decreases AAAT-3' but not lambda DNA which does not contain this sequence. Intense aeration inhibited the growth of S. milleri. The content of restrictase in the cells was the greatest during the logarithmic growth phase. A total of 20,000 units of Smil were isolated from 4 g of cells by cellular extract fractionation with ammonium sulfate and subsequent chromatography on columns with Bio Gel A 0.5 m, heparin agarose, and phosphocellulose. Purified enzyme cut the synthetic oligonucleotide duplex in the center of the recognized site 5'-ATTT decreases AAAT-3'. Smil restrictase is a true isoschisomer of rare-cutting Swal enzyme. Smil belongs to a small group of enzymes which recognize octanucleotide sites and can be used for large-block fragmentation of DNA. Comparison of specificities of rare-cutting and other restrictases suggests that the enzymes recognizing octanucleotides can evolutionally originate from enzymes recognizing both hexanucleotides and tetranucleotides. PMID- 10702988 TI - [DNA homology in various strains of nitrogen-fixing bacteria]. AB - Melting temperature and GC content were evaluated for DNA of some nitrogen-fixing bacteria of Rhizobium leguminosarum and Onobrychis spp. (Adans). The degree of homology between strains of the same species was determined. A combination of thermal denaturing and molecular hybridization can serve as a rapid test for evaluating the genome homology of the organisms compared. PMID- 10702990 TI - [Various genetic and molecular and biologic aspects of the melioidosis causative organism]. PMID- 10702989 TI - [Sequence analysis of hexon gene from adenovirus KR95 inducing hydropericardium syndrome in chickens]. AB - The nucleotide sequence of a part of the HindIII-D fragment (3300 b.p.) of adenovirus KR95 DNA has been determined. Analysis of the nucleotide sequence disclosed a continuous ORF for hexon gene (2814 b.p.) coding the 937 residue protein, part of ORF for the C-terminal region of pVI polypeptide, including 114 residues and the beginning of ORF coding 25 N-terminal residues for viral endoproteinase. Comparison of predicted KR95 hexon sequence and 8 mammalian and avian adenovirus hexon sequences revealed the highest homology between KR95 strain and avian adenoviruses FAV10 and FAV1 (91.1 and 80.1%, respectively). The results were used for creating a test system on the basis of the polymerase chain reaction. The system was used in analysis of fowl samples obtained from 12 poultry farms in Russia. The sequences of hexon gene amplified fragments in the isolated strains and similar fragments of other mammalian and avian adenoviruses have been compared. PMID- 10702991 TI - Effects of soybean (Glycine max) germination on biologically active components, nutritional values of seeds, and biological characteristics in rats. AB - We briefly reviewed the effects of soybean germination on biologically active components, nutritive value of seed and biological characteristics in rats. The purpose of this review is to evaluate the effects of soybean germination on nutritional values of seeds and the potential importance for the use of germinated soybeans, from a contemporary conception, in food preparation as well as on soybean possible influence in optimal health. Germination induced a substantial increase in the content of saponin, oestrogenic compounds and almost all phytosterols, particularly beta-sitosterol of seeds. Lecithin content increased slightly and gradually during germination process. Lipase and alpha galactosidase activities increased whereas lipoxygenase activities reduced after a short period of germination (< or = 72 h). Therefore, the substantial odour and flavour scores of germinated soybean flour were improved. Germinated seeds were also beneficial to heat penetration, their thermolabile antinutritional factors were easier to inhibit than those of dry beans, also the seeds did not require a long cooking time to be palatable. Duration of the germination process greatly influenced the nutritional value and palatability of seeds and biological characteristics in rats. In rat bioassay, one-day germination of soybeans induced a significant increase of daily body weight gain, daily protein intake in rats and protein efficiency ratio (PER) of seed meal. Palatability of seeds was also improved whereas a 5-day germination resulted in a decrease of PER of seed meal (less than the value of unheated seeds) and induced thyroid enlargement in rats. A vapour thermal treatment (100 degrees C, 20 min) eliminated thyroid-active agents and improved PER of seed meal, food intake and final weight of rats. Well prepared germinated soybeans can be used as a good alternate to animal proteins for more balanced nutritional diet. Development of food products from germinated soybean may be another way to further increase the versatility and utility of soybeans for both developing and industrialized countries, as germination induced the modification of certain specific biologically active components, palatability and nutritive value of seeds. PMID- 10702992 TI - Recovery and characterization of Balanites aegyptiaca Del. kernel proteins. Effect of defatting, air classification, wet sieving and aqueous ethanol treatment on solubility, digestibility, amino acid composition and sapogenin content. AB - In order to find alternative protein sources in African regions where protein deficiency in nutrition is prevailing, solubility, in-vitro digestibility, amino acid composition and chemical score of Balanites aegyptiaca Del. kernel proteins were investigated as a function of different processing steps including defatting, air classification, wet sieving and aqueous ethanol treatment. Air classification delivered a fine fraction of 58.1% of the total protein. Applying a wet sieving process, a protein concentrate of 72.9% protein content was achieved but the recovery was very low (35.6%). However, in case of isoelectric precipitation followed by aqueous ethanol treatment both protein content (78.2%) and recovery (53.7%) were high. Data concerning the chemical score revealed, that lysine content of the defatted kernel flour amounted to 74.2% of the recommended FAO/WHO standard level. In-vitro protein digestibility was found to be higher than of legume proteins. The digestible protein of the full fat flour, defatted flour, air classified and wet sieved fine fractions and protein concentrate were 91.9, 93.7, 82.0, 86.4 and 94.2%, respectively. The sapogenin content per 100 g protein of the investigated protein preparations was significantly lower (46% to 62%) than of the initial material (oilcake). PMID- 10702993 TI - Microbiological, nutritional and sensory evaluation of long-time stored amaranth biscuits produced from irradiation-treated amaranth grain. AB - The paper presents some results achieved by the evaluation of microbiological (total bacterial count, coliform bacteria, aerobic sporeforming bacteria, yeasts and moulds), nutritional (lysine) and sensory (shape, surface, colour consistency, taste, odour, the profiling of tastiness) quality and of the aw values of amaranth-based biscuits produced from the amaranth grain irradiated by various ionizing radiation doses (1.5, 3 and 5 kGy, source 60Co) and stored for the period of 12 months at the laboratory temperature (20-25 degrees C). The irradiation dose providing the biscuits maximum hygienic, nutritional and sensory quality maintained up to the end of the one-year storage was 5 kGy. PMID- 10702994 TI - Effect of processing on potato starch: in vitro availability and glycaemic index. AB - The content of digestible starch (DS) and resistant starch (RS) in processed potatoes was assessed. In addition, the effect of domestic cooking on the in vitro digestibility of starch in this tuber, which may influence the glycaemic response, was studied. Resistant starch in raw potato is high, however different RS values were obtained when processed, ranging from 1.18% in boiled potatoes to 10.38% in retrograded flour. In general, cooked potatoes have high levels of DS. Starch digestibility is improved after processing and it is affected by the cooking methods. Boiled and mashed potatoes showed the highest rate of digestion, on the contrary raw potato was hardly digested. The estimated Glycaemic Index (GI) from the degree of starch hydrolysis within 90 min was in accordance with the reported GI values, for potatoes processed in the same way. PMID- 10702995 TI - Changes in biogenic amines in mature and germinating legume seeds and their behavior during cooking. AB - Ungerminated legume seeds (broad bean, chick pea and lupine) were contained all tested biogenic amines. Tryptamine (TRY) was the main biogenic amine detected, and its concentration considerably increased during the germination. beta Phenylethylamine (PHE) was detected in small amounts and its concentration slowly increased during germination. The concentration of tyramine (TYR) showed a fluctuation pattern of changes during germination in all tested legumes. The concentrations of cadaverine (CAD) and putrescine (PUT) increased during the germination period in all tested grains. However, histamine (HIS) showed a fluctuated pattern of changes in both broad bean and lupine, and a gradual increase in chick pea. Spermidine (SPD) and spermine (SPM) contents of broad bean and chick pea showed a fluctuation pattern of change, while, a decrement trend of change was recorded for lupine along the germination period. By cooking, legume samples became free of biogenic amines which appeared in the boiling water. Heat treatment seems to have little effect on the concentration of biogenic amines in legume sprouts. The amounts of biogenic amines detected in the boiling water are less than the initial amounts of the sprouts (expected amounts). PMID- 10702996 TI - Determination of biogenic amines in cheese by ion exchange chromatography. AB - A sensitive, fully automated method, yielding reproducible results, was developed for the determination of relevant biogenic amines in cheese. Histamine, tyramine, putrescine, cadaverine, tryptamine, agmatine, spermidine and spermine were separated in the ion exchanger of OSTION LG ANB column of the automatic amino acid analyser Mikrotechna 339 T. Elution was carried out at 60 degrees C using a system of two buffers. The samples were extracted and precipitated by trichloroacetic acid and purified, after removal of fat, by membrane filtration. The recovery for individual amines in cheese ranged between 86% and 108%. The detection limit was of 1-5 mg of the respective amine per 1 kg of cheese and the method was linear within the dose range of 0.1-10 micrograms (for tryptamine 0.5 10 micrograms). PMID- 10702997 TI - Quality changes of onion (Allium cepa L.) as affected by the drying process. AB - Thin layer drying experiments of sliced onion were carried out under different controlled conditions using a laboratory dryer. Quality changes of the dried product were evaluated by analysis of colour, pyruvate, chemical and sensory parameters. The results obtained proved that drying temperatures above 65 degrees C exert a pronounced influence on colour changes. The pyruvate content decreased with increasing of temperature and slice thickness. The sugar content was also found to be significantly influenced by the drying temperature. The rate of ascorbic acid degradation decreased with increasing temperature and slice thickness. Significant correlations were obtained between chemically determined pyruvate content and sensory evaluated odour of the dried onion. PMID- 10702998 TI - Antioxidative activity of rosemary extract in lipid fraction of minced meat balls during storage in a freezer. AB - The influence of ethanol rosemary extract on lipid fraction of minced meat balls during storage in the freezer was studied. The quality of stored products was evaluated sensorically by the scale method and profiling of taste and aroma. Oxidative changes were investigated on the base of the changes in malonaldehyde content. It was observed that the addition of rosemary extract delayed the oxidation of lipid fraction of products. The antioxidative effect was related to the concentration of this extract in the product. The sensory changes and their intensity in products during storage depended on the amount of the extract added and the time of storage. The addition of the extract delayed the appearance of rancid taste in products. PMID- 10702999 TI - Influence of dietary spices and their active principles on pancreatic digestive enzymes in albino rats. AB - A few common spices or their active principles were examined for their possible influence on digestive enzymes of pancreas in experimental rat. Groups of animals were maintained for 8 weeks on the following spice diets: curcumin (0.5%), capsaicin (15 mg%), piperine (20 mg%), ginger (50 mg%), cumin (1.25%), fenugreek (2%), mustard (250 mg%) and asafoetida (250 mg%). Dietary curcumin, capsaicin, piperine, ginger, fenugreek and asafoetida prominently enhanced pancreatic lipase activity. Curcumin, capsaicin, piperine, ginger, cumin and asafoetida also stimulated pancreatic amylase. Trypsin was significantly stimulated by curcumin, capsaicin, piperine, ginger and cumin, while chymotrypsin was stimulated by all the spices tested except mustard. This stimulatory influence of test spices on the pancreatic digestive enzymes was however not observed when their intake was restricted to a single oral dose. The positive influences on the pancreatic digestive enzymes exerted by a good number of spices consumed in diet could be a factor contributing to the well recognised digestive stimulant action of spices. PMID- 10703000 TI - Evaluation of enzymes produced from yeast. AB - Pichia pinus was found to be capable of growing on mango wastes, producing pectinase (pectin lyase, EC-4.2.2.10) and lactase (beta-galactosidase, EC 3.2.1.23) enzymes. The two enzymes were successively purified by precipitation with ammonium sulfate followed by chromatography on Sephadex G-120. The purification procedure provided 1,846 and 929 fold purification with 20.6 and 24% yield recovery of pectinase and lactase, respectively. the km value of pectinase was 0.33% for pectin at pH 4.5 and that for lactase was 0.166% for lactose at pH 7.0. The purified enzymes, pectinase and lactase are stable up to 50 degrees C for 60 and 45 min, respectively, with 20 and 35% loss of their activity. Gel filtration on Sephadex G-200 indicated that the molecular weights of the purified pectinase was 90 x 10(3) Dalton and of lactase 115 x 10(3) Dalton. On the basis of the evaluation tests done, the enzymes were considered to have a potential technological interest as treating mango pastes (residues left after mango juice preparation) with the two prepared enzymes resulted in an increase of the colour intensity, total carbohydrate content and juice yield. Treating milk with the purified lactase also showed an increase in the total carbohydrate and reducing sugar produced. PMID- 10703001 TI - The effect of Allium sp. on the extension of lipolysis and proteolysis in Van herby cheese during maturation. AB - The aim of this study was to determine the effect of herb (Allium sp.) on biochemical changes of herby cheese produced in Turkey. Raw cows' milk was used for cheese manufacture. Five groups of cheeses, containing 0 (as control), 0.5, 1, 2, and 3% herb, were produced and coded as K, A, B, C, and D respectively. All cheese groups were ripened at 8 degrees C for 90 days. Samples were taken from cheeses after 3, 15, 30, 60 and 90 days, and analysed for lipolysis (as acid degree value) and proteolysis (water-soluble nitrogen, TCA-soluble nitrogen, and PTA-soluble nitrogen). It was found that lipolysis in herby cheeses increased with increasing herb addition, and the increase in lipolysis degree was significant (P < 0.05) in cheese D. Water-soluble N, TCA-soluble N, and PTA soluble N as indicator of proteolysis degrees were affected significantly (P < 0.05) by increasing herb ratios. PMID- 10703002 TI - [A method for non-destructure estimation of volatile metabolites of fruit after harvest]. PMID- 10703003 TI - [Determination of ochratoxin A in beer with automatic purification of samples on immunoaffinity columns]. PMID- 10703005 TI - Contents of metals in cultivated mushrooms. PMID- 10703004 TI - Antibacterial activity of green tea polyphenols against Escherichia coli K 12. PMID- 10703006 TI - [Therapy of generalized tonic-clonic status epilepticus in adulthood]. AB - If continuous seizure activity lasts longer than 5 minutes generalized tonic clonic seizures require prompt treatment, if significant morbidity and mortality are to be avoided. The mortality varies (mean: 20%) depending on patient age and etiology. Control of status epilepticus is achieved by benzodiazepines in about 80% of cases: Lorazepam is recommended due to its longer-acting effects on the central nervous system. To maintain the anticonvulsive effect phenytoin is usually administered intravenously. Fosphenytoin (not approved in Germany) has advantages over phenytoin, because it can be given three times more rapidly and produces fewer side effects. The IV use of valproic acid in status epilepticus seems to be promising, but needs further evaluation. There is no generally accepted treatment protocol for the therapy of persistent seizure activity lasting more than 60 minutes (i.e., refractory status epilepticus). Usually phenobarbital, or general anesthesia with thiopental or pentobarbital are treatment recommendations. In recent reports, the administration of midazolam or propofol proved to be effective and well-tolerated. PMID- 10703007 TI - [Inhibition of catechol-O-methyltransferase. Optimizing dopaminergic therapy in idiopathic Parkinson syndrome with entacapone]. AB - Registration of the inhibitor of the catechol-O-methyltransfersase (COMT) tolcapone has been stopped due to the possible relationship of tolcapone treatment to three cases of fatal hepatitis. As a result, strong uncertainty has emerged among neurologists about the principle of COMT inhibition itself. We review data, especially on the remaining COMT inhibitor, entacapone, with regard to pre-clinical and clinical efficacy and safety. PMID- 10703008 TI - ["Spreading depression" and peri-infarct depolarizations. Relevant pathological events in migraine and stroke?]. AB - Spreading depression is a fascinating phenomenon that can be provoked by chemical, electrical or mechanical stimuli of the cortex. Spreading depression like transient depolarizations are observed in the peri-infarct tissue after focal ischemia. The reduction in electrical activity and a negative direct current potential shift propagating over the cortex with a rate of 2-5 mm/minute are the physiological hallmarks. It is thought that spreading depression and peri infarct depolarizations might play a role in the pathogenesis of migraine and stroke. However, these events have never been detected in humans. This paper reviews the physiological characteristics of spreading depression and peri infarct depolarizations and discusses their potential role in migraine and stroke. PMID- 10703009 TI - [Diffusion-weighted MRI in patients with Creutzfeldt-Jakob disease]. AB - Today the diagnosis of Creutzfeldt-Jakob disease (CJD) is proven only postmortem or by evidence of neuropathology. During the patient's lifetime EEG recordings or cerebrospinal fluid analysis may support the diagnosis. In most cases, T2-MRI scans show hyperintensities of the basal ganglia. A new imaging technique called diffusion-weighted MRI (DWI) has recently been established. The sensitivity of DWI was evaluated in five patients suspected of CJD. All five cases showed hyperintense signal changes in the basal ganglia on DWI sequences. These findings were more pronounced in DWI than in T2, FLAIR, or PD-weighted images. Thus, DWI seems to be the most sensitive sequence for detecting changes in patients with suspected CJD. Moreover, its short scanning time ensures that fewer artifacts occur, especially in the case of myoclonus. PMID- 10703010 TI - [Progressive multifocal leukoencephalopathy in AIDS. Overview and retrospective analysis of 17 patients]. AB - We describe retrospectively the course of 17 AIDS patients with progressive multifocal leukoencephalopathy (PML) and give a review of their clinical symptoms and survival times. The relative frequency of PML in our cohort of AIDS patients was 2.6%. The mean of CD4-positive cells was 80.5 +/- 82.5/microliter. CD4 positive cells were > 200/microliter only in one patient and increased significantly under a combination of three antiretroviral drugs whereas, with the other patients, CD4-positive cells did not increase with a maximum of two antiretroviral drugs. The mean survival time was 6.6 (1.5-20) months and correlated positively with the number of CD4-positive cells. The diagnosis of PML can be regarded as confirmed when JC-virus DNA is detectable in cerebrospinal fluid, typical changes can be seen in MRI and typical clinical symptoms occur. No effective therapy is known to date. Single case reports on therapy with cidofovir, as in one of the cases described here, showed positive results. PMID- 10703011 TI - [Comparison of the annual data of 2 stroke units in neurological clinics of acute hospitals]. AB - The Stroke Unit concept of the German Neurological Society differs from its precedents in Scandinavia or the United Kingdom. Hallmarks of the German concept are very early onset of diagnosis and treatment, continuous surveillance of vital functions by bedside monitors and specialized care by a multiprofessional team. This comparison of two Stroke Units in Minden and Munich-Harlaching, working according to the new German concept, shows broad similarities in admission intervals, diagnostic procedures, treatment modalities and short term prognosis, with an exceedingly low in-hospital mortality of 3.4-5.6% and a high proportion of patients (64-69%) leaving the hospital with Barthel scores above 70. Ongoing studies will show how this kind of Stroke Unit treatment compares to general wards. PMID- 10703012 TI - [Generalized brain edema in non-purulent meningoencephalitis. The anti-edema effect of therapy with dexamethasone]. AB - We report about two female inpatients aged 58 and 24, suffering from non-purulent meningoencephalitis, in the first case caused by varizella-zoster-virus, in the second case probably due to viral infection. Both patients developed a diffuse brain edema associated with a progressive loss of consciousness. The adjunctive treatment with dexamethasone led to rapid improvement of clinical symptoms. Computed tomography revealed a significant reduction of brain edema. These results of these two cases support the efficacy of corticosteroids as adjunctive treatment of diffuse brain edema caused by non-purulent meningoencephalitis. The pathophysiological mechanisms are discussed. PMID- 10703013 TI - [Spontaneous spinal hemorrhage. Outcome after surgical therapy of epidural hematomas]. AB - Eight patients with spontaneous spinal epidural hemorrhages are presented. All eight initially had suffered from severe neck or back pain. In seven cases, progressive neurological deficits had developed, some of which led to complete paraplegia. On all eight patients operations were performed. After an average of 11 weeks' follow-up, full recovery from the preoperative pain and neurological deficits could be seen in 6 of the patients. If spinal hemorrhage is treated by decompression at an early stage, there is a good prognosis with respect to pain and neurological deficits. It was found that neurological deficits sometimes showed complete improvement, even if they were older than 36 hours before surgery was performed. In cases of severe local complaints in combination with progressive neurological deficits a spinal hemorrhage always must be considered. The best diagnostic method is magnetic resonance imaging. PMID- 10703014 TI - [Somatoparaphrenia. A positive variant of anosognosia for hemiplegia]. AB - Anosognosia for hemiplegia (AHP), i.e., unawareness of motor deficits and associated disorders, has been frequently reported, pre-dominantly following right hemispheric lesions. To a smaller extent, there are case reports of patients who give accounts of a feeling of strangeness concerning the contralesional limbs and sometimes attribute them to other persons. This "positive-variant" of AHP has been labeled "somatoparaphrenia" (SP). We report a case of SP in a 85-year-old woman with infarction of the right posterior cerebral artery and posterior parts of the right thalamus. She showed AHP and described her left side alternatively as her handicapped nephew and a clumsy cat. Misidentification of her daughter also occurred. With respect to the literature the predominant neuroanatomical features involved are lesions including right parietal cortex and/or posterior parts of the thalamus. Theories concerning the pathogenesis of this phenomenon comprise a denial of the illness, a lack of awareness caused by reduced sensory feedback and neglect, a misidentification or disturbance of the active discovery process considered necessary for realizing one's disorder. PMID- 10703015 TI - [Descending paralysis caused by wound botulism. A case report]. AB - We report on the history and clinical findings of an injecting drug abuser in the Canton of Zurich who presented with multiple deep abscesses in the arms and legs. A diagnosis of wound botulism was made based on his clinical presentation with a rapidly progressing descending paralysis starting at the cranial nerves, a neuromuscular junction disorder on neurophysiologic testing, and normal findings on lumbar puncture. Several cases of wound botulism have occurred in i.v. drug abuse in Switzerland since 1997. We suspect subcutaneous injections of contaminated heroin containing Clostridium spores as sites of entry. Wound botulism caused by Clostridium botulinum is a rare cause of rapidly progressing, generalized, flaccid paralysis and should be considered in patients with a history of i.v. drug abuse presenting with descending paralysis. PMID- 10703016 TI - [Prevention of meningococcal meningitis]. AB - In Germany, the incidence of meningococcal disease is approximately 1/100,000 and has not risen during recent years. Transmission occurs by direct contact with respiratory droplets, mostly from asymptomatic carriers and less frequently from patients with meningococal disease. The incubation period can vary from 2-10 days but usually is 3-4 days. Incidence is highest in children and decreases with age. The mortality from meningococcal disease is approximately 10%. In case meningococcal meningitis is clinically suspected antibiotic treatment (in Germany with penicillin G or a cephalosporin) should not be delayed. Patients must be isolated for at least 24 hours after the institution of antibiotic therapy. For early detection of local outbreaks, public health authorities should be quickly informed of suspected cases. Persons in close contact should be treated with antimicrobial chemoprophylaxis. In addition to rifampin, ciprofloxacin or ceftriaxone can be used for chemoprophylaxis. For control of local outbreaks of serogroup C meningococcal disease, the meningococcal vaccine can be used. PMID- 10703017 TI - ["Inner perilymph fistula" of the anterior semicircular canal. A new disease picture with recurrent attacks of vertigo]. AB - In 1998 Minor et al. described a new variant of perilymphatic fistula: the "superior canal dehiscence syndrome". This syndrome is clinically characterized by recurrent attacks of vertigo and oscillopsia induced by loud noises or stimuli that result in changes in intracranial or middle ear pressure. It is caused by a dehiscence of bone overlying the superior (anterior) semicircular canal. Due to this dehiscence, a third, mobile window (in addition to the round and oval windows) is formed, and changes in pressure are pathologically transduced to the anterior semicircular canal. Although the superior canal dehiscence syndrome is not a rare condition, no other cases have yet been reported. Therefore, we describe a typical patient who suffered for many years from recurrent attacks of vertigo and oscillopsia induced by coughing and Valsalva's maneuvers. High resolution temporal bone CT scan showed a defect in the bone overlying the left anterior semicircular canal. Three-dimensional eye movement recordings using the search coil technique and subsequent vector analysis demonstrated that the eye movements were induced by excitation of the left anterior semicircular canal. We conclude that superior canal dehiscence syndrome is an important differential diagnosis in patients suffering from symptoms of a perilymphatic fistula, especially since it can be successfully treated by "plugging" of the affected semicircular canal. Such patients are thus spared unnecessary surgery of the middle ear. PMID- 10703018 TI - [Comments on the review by R. Kaiser. Paraneoplastic neurological syndromes, diagnostic and pathogenetic significance of autoantibodies]. PMID- 10703019 TI - [Comment on the contribution by C. B. Ostertag and P. C. Warnke. Neuronavigation. Computer-assisted neurosurgery]. PMID- 10703021 TI - A detoxification dialogue. PMID- 10703020 TI - [Gunter Baumgartner 1924-1991 Freiburg and Zurich. Comments on the title picture]. PMID- 10703022 TI - Influencing handwashing behavior. PMID- 10703023 TI - Caring for the woman with migraine headaches. AB - Approximately 16% of American women experience migraine headaches. These debilitating headaches cause lost time from family, social activities, and work. Although migraines are thought to be a result of shifting menstrual and perimenopausal hormones, a physiologic connection has not been well established. Despite the lack of certainty regarding migraine cause, several theories have been postulated and a significant amount of literature has been published addressing the management of premenstrual migraines. Fewer articles have been published regarding the management of perimenopausal migraines, which are treated somewhat differently. This article approaches both premenstrual and perimenopausal migraine headaches from a chronic disease perspective, focusing on self-care and the use of prescription and nonprescription therapies. Implications for practice and future research are also discussed. PMID- 10703024 TI - Drugs, devices, diagnostics, and policies: the year's review. AB - Throughout 1999, the Food and Drug Administration approved several new drugs and medical devices. The agency also took an active role in defining public policies regarding pharmacologic issues for the American consumer. This article provides an overview of these topics. PMID- 10703025 TI - Listening to the patient's natural alarm system. PMID- 10703026 TI - Treating syphilis effectively. PMID- 10703027 TI - Dosing by dropperful. PMID- 10703028 TI - Providing clear patient instructions. PMID- 10703029 TI - Ethnic differences in the growth of low-birthweight infants. AB - Differences in growth were investigated among ethnic groups in low-birthweight babies (< 2500 g or < 32 weeks gestation) at birth and at 2-3 years. This prospective study was based on data for all 3091 low-birthweight live births in the South East Thames Region, UK, over a 1-year period, surviving to discharge from hospital. Weights were recorded at birth and at 2-3 years for 998 babies, and head circumferences for 859. These were compared with the UK 1990 reference standards. Ethnic differences were adjusted for parity, multiple birth, smoking and alcohol during pregnancy, mother's height, weight and age, marital status, partner's support and social class. At 2-3 years, there was substantial average catch-up growth only for the weight of infants of > or = 32 weeks' gestation. Babies < 32 weeks gestation had fallen behind. Head circumferences had failed to keep up or had fallen behind for both groups. The ethnic groups had similar birthweight standard deviation scores (SDS). At 2-3 years, Black babies of < 32 weeks' gestation had gained in weight and head circumference compared with White babies (adjusted difference in weight SDS: 0.71, [95% CI 0.28, 1.13]). Asian babies of at least 32 weeks' gestation had smaller heads than White, a difference that increased with time. It was concluded that ethnic differences in the growth of low-birthweight infants are related to gestational age. Although most of the babies born at < 28 weeks' gestation were close to their birthweight reference standards, only the Black infants had maintained their position at 2-3 years. Black infants, particularly when born preterm, tend to put on more weight than White. PMID- 10703030 TI - Maternal obesity and glucose intolerance during pregnancy among Mexican Americans. AB - Low birthweight is uncommon among Mexican-American infants, despite the substantial proportion of mothers who live in poverty. This apparent paradox has generated studies of factors protecting fetal growth, but may have masked other important health problems in the Mexican-American community. Obesity, impaired glucose tolerance and diabetes are common among Mexican-American women of childbearing age and during pregnancy. Prevalence of these conditions is two to four times higher in Mexican-American than in non-Hispanic white women. As obesity and glucose intolerance during pregnancy are associated with fetal overgrowth and increased risk of subsequent obesity and type 2 diabetes in mother and child, the adequacy of birthweight as a measure of maternal and infant risk may be obscured in populations with a high prevalence of these conditions. Their possible contribution to the increasing incidence of obesity and type 2 diabetes in Mexican-American children, adolescents and young adults has not been examined. Appropriate preconception, prenatal and follow-up care may identify high-risk women, improve weight and metabolic status and reduce the severity and impact of diabetes and its complications. However, late or no prenatal care is common among Mexican-American women and the frequency of follow-up care is unknown. As low birthweight is a major public health indicator of maternal and neonatal health, perceived 'good birth outcomes' have reduced health policy, programme and research attention to Mexican-American mothers and infants. Studies of the impact of obesity and glucose intolerance during pregnancy on the birthweights of Mexican-American infants should be undertaken, along with systematic assessment of the subsequent health status and preventive health-care needs of women and children in this population. PMID- 10703031 TI - What explains away the increased risk of histological chorioamnionitis in African American mothers of very-low-birthweight infants? Developmental Epidemiology Network Investigators. AB - We sought explanations for African-American mothers' increased risk of chorioamnionitis by sequentially adjusting for confounder variables both individually and in groups. We searched for a subset of covariates that had the most influence on the chorioamnionitis odds ratio (OR) of these women. The sample consisted of 305 African-American and 520 White mothers who gave birth to a very low-birthweight (< or = 1500 g) infant between 1991 and 1993, whose placenta was examined according to protocol and whose hospital chart was reviewed. Histologically proven chorioamnionitis was present in 43% of the placentas from African-American women and in 27% of those from Whites (crude OR 2.1, 95% confidence interval 1.5, 2.8). Singleton status appeared to be the most important effect modifier, with significant crude ORs of 1.5 among singletons and 3.4 among non-singletons. Using logistic regression models in the whole sample and in subgroups, we sought to 'explain away' this increased risk. Indeed, addition of information about confounder variables resulted in considerable reduction in the ORs to 1.1 among singletons and 1.9 among non-singletons. Particularly important among the confounders were singleton birth, Medicaid insurance, duration of ruptured membranes and gestational age. We discuss the possibility that this set of confounding variables conveys, in part, the same information as the variable African-American, and also perhaps information about the availability and/or utilisation of prenatal health care. PMID- 10703032 TI - Factors associated with preterm births in southeast Brazil: a comparison of two birth cohorts born 15 years apart. AB - An increase in preterm deliveries in Ribeirao Preto stimulated an analysis of possible explanatory factors. Two cohorts of singleton livebirths were studied, the first based on 6746 births in 1978-9 and the second based on 2846 births in 1994. A logistic regression was carried out to assess the association of preterm birth with several sociodemographic, behavioural and clinical variables, including year of survey. Delivery in private settings compared with a public setting, maternal age of < or = 17 compared with any other age group, and mothers who had had previous abortions and previous stillbirths were associated with greater rates of preterm birth. Although there was an increase in preterm birth rates regardless of mode of delivery, the increase was greater in the caesarean section group than in the vaginal delivery group. Over the study period, deliveries in private hospitals and caesarean section operations increased markedly (from 4% to 36% and from 30% to 51% respectively). Caesarean section may be the main contributor to the increase of preterm birth rate in this study. It is essential to ensure that health-care staff, especially those in private facilities, are properly educated and audited. PMID- 10703033 TI - Stillbirths and neonatal encephalopathy in Kathmandu, Nepal: an estimate of the contribution of birth asphyxia to perinatal mortality in a low-income urban population. AB - We describe a prospective cross-sectional survey over a 12-month period in the principal maternity hospital of Kathmandu, Nepal, where over 50% of the local population deliver. The study aim was to estimate the contribution of birth asphyxia to perinatal mortality in this setting. During 1995, there were 14,371 livebirths and 400 stillbirths, a total stillbirth rate of 27 per 1000 total births. The fresh term (2000 g or more) stillbirth rate was 8.5 per 1000 total births [95% CI 7.1, 10.1]. Ninety-two cases of neonatal encephalopathy (NE) affecting term infants were detected (excluding those due to congenital malformations, hypoglycaemia and early neonatal sepsis). The birth prevalence of NE was 6.4 per 1000 livebirths [95% CI 5.2, 7.8]. There was evidence of intrapartum compromise in 63 (68%) of the cases of NE and 65 (76%) of the stillbirths, but only in 12 (12%) of controls. The cause-specific early neonatal mortality rate for NE was 2.1 per 1000 livebirths [95% CI 1.4, 3.0]. Combining the NE deaths and fresh stillbirths gives an upper estimate for term birth asphyxia perinatal mortality rate of 10.8 per 1000 total births [95% CI 9.2, 12.6], 24% of all perinatal deaths before hospital discharge. This study suggests that birth asphyxia remains an important cause of perinatal mortality in developing countries. The paper discusses the pros and cons of different strategies to reduce birth asphyxia in low-income countries. PMID- 10703034 TI - Lower respiratory tract infections in an ethnic and social context. AB - Family size and smoking during pregnancy were studied as mediating factors for social and ethnic variation of lower respiratory tract infection (LRI) in hospital discharge data. The study population consisted of all children aged 0-4 years in the three largest metropolitan areas of Sweden during 1990-94. Maternal smoking during pregnancy increased the risk of children being admitted to hospital for LRI during their first 3 years of life, with an adjusted odds ratio (OR) of 1.3 for the age-group 0-1 years. The risk attributed to smoking during pregnancy was the same in children of mothers in ethnic groups in which smoking during pregnancy was related to social adversity as in those in which it was not. Having at least one sibling increased the risk of being admitted to hospital for LRI in the age group 0-1 years (adjusted OR 2.2). This risk was lower in children in families in which the mother was born in southern Europe, Africa, Asia or Latin America, suggesting a contextual relation to ethnicity for this risk factor. It is concluded that family size and smoking during pregnancy are important mediators of the risk for LRI related to social adversity and ethnicity in Swedish children below 2 years of age. PMID- 10703035 TI - Epidemiology of holoprosencephaly in Hawaii, 1986-97. AB - Holoprosencephaly is a birth defect affecting the medial structures of the brain and face. This investigation examined the epidemiology of holoprosencephaly in Hawaii between 1986 and 1997, using data obtained from a birth defects registry, and compared the results with those of other population-based studies. Twenty five cases were identified, producing a prevalence of 1.09 per 10,000 livebirths. There were 17 (68.0%) livebirths, two (8.0%) fetal deaths and six (24.0%) elective terminations. Ten (58.8%) of the livebirths died before the age of 1 year. Seven (28.0%) had a known chromosomal abnormality, most often trisomy 13. Prevalence rates were higher for maternal age > 39 years, maternal race/ethnicity of Far East Asian or Filipino, females and residence in Maui County. The small number of cases limited the statistical significance of the study. However, this report confirms many of the findings from the previous studies and adds new findings, particularly the higher prevalence among Far East Asians and Filipinos. PMID- 10703036 TI - Vitamin A deficiency in healthy children aged 6-59 months in Izmir Province of Turkey. AB - Vitamin A deficiency even at subclinical levels is associated with increased childhood mortality. There have been few studies related to vitamin A status of children in Turkey. The aim of this study was to assess vitamin A status of children aged 6-59 months in Izmir, Turkey, and to evaluate the relationship of these levels with nutritional status. One hundred and sixty children were selected for the study using the cluster sampling method. Serum retinol levels were measured by high-performance liquid chromatography (HPLC) and ranged from 9.8 to 59.2 micrograms/dL (mean 29.3 +/- 9.5 micrograms/dL). Levels were below the lower limit of the normal range in 15.6% of the children. Deficient and marginal serum retinol among stunted children were observed in 16% and 42% respectively. There was a statistically significant relationship between low serum retinol and stunting (P < 0.05). Although xerophthalmia and other clinical signs of vitamin A deficiency are rarely seen, subclinical vitamin A deficiency is a public health problem in Izmir, Turkey. PMID- 10703037 TI - Aetiology of childhood vision impairment, metropolitan Atlanta, 1991-93. AB - Data from the population-based Metropolitan Atlanta Developmental Disabilities Surveillance Program (MADDSP) were used to describe the underlying causes of vision impairment (VI; corrected visual acuity in the better eye of 20/70 or worse) in young children (n = 228) in metropolitan Atlanta in 1991-93. Children with VI were identified through record review at multiple educational and medical sources. Children were categorised as having isolated VI or multiple disabilities (i.e. VI plus one or more of four additional developmental disabilities) and as having low vision (visual acuity 20/70-20/400) or blindness (visual acuity worse than 20/400). Medical conditions abstracted from MADDSP sources were reviewed to determine the probable aetiology of a child's VI. Aetiologies were assigned to one of three developmental time periods: prenatal, perinatal, or postnatal. Prenatal aetiologies were identified in 43% of the children; 38% of the prenatal aetiologies were genetic. Perinatal aetiologies were found in 27% of the children. Postnatal aetiologies were rare. Prenatal aetiologies were more common in children with isolated VI; perinatal and postnatal aetiologies were more common in children with multiple disabilities. Children with prenatal aetiologies tended to have less severe vision loss than did children with perinatal or postnatal aetiologies. The distribution varied by birthweight, but did not differ significantly by sex or race. PMID- 10703038 TI - Active and passive smoking during pregnancy and risk of central nervous system tumours in children. AB - The effects of maternal active and passive smoking during pregnancy on childhood central nervous system (CNS) tumours were assessed in a population-based case control study. The mothers of 244 children aged 0-15 years with CNS tumours and 502 control mothers were interviewed about their smoking habits. All families were resident in the region of Lombardy, Italy. Risk estimates were calculated by unconditional logistic regression, adjusted for age, sex and area of residence. Active smoking by the parents before pregnancy was not associated with increased risk of CNS tumours in the children. Active smoking by the mother during early pregnancy (approximately the first 5 weeks) was associated with a slightly increased risk of the child developing a CNS tumour (odds ratio [OR] 1.5 [95% CI 1.0, 2.3]). An increased risk of CNS tumours was found in the children of non smoking mothers exposed regularly to tobacco smoke both in early pregnancy (OR 1.8 [95% CI 1.2, 2.6]) and in late pregnancy (OR 1.7 [95% CI 1.2, 2.5]). Although this study was retrospective, the results confirm our previous findings and suggest an association between the risk of developing CNS tumours in children and regular passive smoking by the mother during pregnancy. PMID- 10703039 TI - Using genetic epidemiology to study Rett syndrome: the design of a case-control study. AB - Rett syndrome is a neurological disorder that is seen almost exclusively in females. Although generally considered to have a genetic basis, the underlying mechanism remains obscure. One favoured hypothesis is that the syndrome is an X linked dominant disorder, lethal or non-expressed in males. Genealogical research has also suggested that the mode of transmission in Rett syndrome may involve a premutation which over several generations is converted to a full mutation. Geographical clustering has been reported, and it has also been proposed that Rett syndrome is a clinically variable condition and that other neurological disorders may be occurring more commonly in families with Rett syndrome. Other studies have found an apparent increase in intellectual disability and seizures in the extended families of girls with Rett syndrome. The science of genetic epidemiology can be used to identify familial aggregation, which is the clustering of a disorder within a family. We have used a case-control study design to investigate both fetal wastage and familial aggregation of other disorders in families of girls with Rett syndrome. The Australian Rett Syndrome Database provided the source of cases, and control probands were girls of a similar age with normal development. This paper describes the methodology for a case-control study of this rare condition using pedigree data and discusses issues in the collection and evaluation of such data. The use of a control population is an important feature. Both the strengths and the shortcomings of our design are identified, and recommendations are made for future research. PMID- 10703040 TI - Brain structure-function relationships: advances from neuroinformatics. PMID- 10703041 TI - Uniformity, specificity and variability of corticocortical connectivity. AB - In many studies of the mammalian brain, subjective assessments of connectivity patterns and connection strengths have been used to subdivide the cortex into separate but linked areas and to make deductions about the flow of information through the cortical network. Here we describe the results of applying statistical analyses to quantitative corticocortical connection data, and the conclusions that can be drawn from such quantitative approaches. Injections of the tracer WGA-HRP were made into different visual areas either side of the middle suprasylvian sulcus (MSS) in 11 adult cats. Retrogradely labelled cells produced by these injections were counted in selected coronal sections taken at regularly spaced intervals (1 mm) through the entire visual cortex, and their cumulative sums and relative proportions in each of 16 recognized visual cortical areas were computed. The surface dimensions of these areas were measured in each cat, from contour lines made on enlarged drawings of the same sections. A total of 116,149 labelled neurons were assigned to all visual cortical areas in the 11 cats, with 5212 others excluded because of their uncertain location. The distribution of relative connection strengths, that is, the percentage of labelled cells per cortical area, was evaluated using non-parametric cluster analyses and Monte Carlo simulation, and relationships between connection strength and area size were examined by linear regression. The absolute size of each visual cortical area was uniform across individual cats, whereas the strengths of connections between the same area pairs were extremely variable for injections in different animals. The overall distribution of labelling strengths for corticocortical connections was continuous and monotonic, rather than inherently clustered, with the highest frequencies presented by the absent (zero density) and the very-low-density connections. These two categories could not, on analytical grounds, be separated from each other. Thus it seems that any subjective description of corticocortical connectivity strengths by ordinal classes (such as 'absent', 'weak', 'moderate' or 'strong') imposes a categorization on the data, rather than recognizes a structure inherent in the data themselves. Despite the great variability of connections, similarities in the distribution profiles for the relative strengths of labelled cells in all areas could be used to identify clusters of different injection sites in the MSS. This supported the conclusion that there are four connectionally distinct subdivisions of this cortex, corresponding to areas 21a, PMLS and AMLS (in the medial bank) and to area PLLS (in the lateral bank). Even for tracer deposits in the same cortical subdivision, however, the strength of connections projecting to the site from other cortical areas varied greatly across injection in different individual animals. We further demonstrated that, on average, the strength of connections originating from any given cortical area was positively and linearly correlated with the size of its surface dimensions. When analysed by specific injection site location, however, this relationship was shown to hold for the individual connections to the medial bank MSS areas, but not for connections leading to the lateral bank area. The data suggest that connectivity of the cat's visual cortex possesses a number of uniform global features, which are locally organized in such a way as to give each cortical area unique characteristics. PMID- 10703042 TI - On variability in the density of corticocortical and thalamocortical connections. AB - Variability is an important but neglected aspect of connectional neuroanatomy. The quantitative density of the 'same' corticocortical or thalamocortical connection may vary by over two orders of magnitude between different injections of the same tracer. At present, however, the frequency distribution of connection densities is unknown. Therefore, it is unclear what kind of sampling strategies or statistical methods are appropriate for quantitative studies of connectivity. Nor is it clear if the measured variability represents differences between subjects, or if it is simply a consequence of intra-individual differences resulting from experimental technique and the exact placement of tracers relative to local spatial and laminar variation in connectivity. We used quantitative measurements of the density of a large number of corticocortical and thalamocortical connections from our own laboratories and from the literature. Variability in the density of given corticocortical and thalamocortical connections is high, with the standard deviation of density proportional to the mean. The frequency distribution is close to exponential. Therefore, analysis methods relying on the normal distribution are not appropriate. We provide an appendix that gives simple statistical guidance for samples drawn from exponentially distributed data. For a given corticocortical or thalamocortical connection density, between-individual standard deviation is 0.85 to 1.25 times the within-individual standard deviation. Therefore, much of the variability reported in conventional neuroanatomical studies (with one tracer deposited per animal) is due to within-individual factors. We also find that strong, but not weak, corticocortical connections are substantially more variable than thalamocortical connections. We propose that the near exponential distribution of connection densities is a simple consequence of 'patchy' connectivity. We anticipate that connection data will be well described by the negative binomial, a class of distribution that applies to events occurring in clumped or patchy substrates. Local patchiness may be a feature of all corticocortical connections and could explain why strong corticocortical connections are more variable than strong thalamocortical connections. This idea is supported by the columnar patterns of many corticocortical but few thalamocortical connections in the literature. PMID- 10703043 TI - Coordinate-independent mapping of structural and functional data by objective relational transformation (ORT). AB - Neuroscience has produced an enormous amount of structural and functional data. Powerful database systems are required to make these data accessible for computational approaches such as higher-order analyses and simulations. Available databases for key data such as anatomical and functional connectivity between cortical areas, however, are still hampered by methodological problems. These problems arise predominantly from the parcellation problem, the use of incongruent parcellation schemes by different authors. We here present a coordinate-independent mathematical method to overcome this problem: objective relational transformation (ORT). Based on new classifications for brain data and on methods from theoretical computer science, ORT represents a formally defined, transparent transformation method for reproducible, coordinate-independent mapping of brain data to freely chosen parcellation schemes. We describe the methodology of ORT and discuss its strengths and limitations. Using two practical examples, we show that ORT in conjunction with connectivity databases like CoCoMac (http://www.cocomac.org) is an important tool for analyses of cortical organization and structure-function relationships. PMID- 10703044 TI - Analysis of the connectional organization of neural systems associated with the hippocampus in rats. AB - The hippocampus of the rat enjoys a central significance for researchers interested in the neural mechanisms of memory and spatial information processing. Many of the theoretical models advanced to explain function in this system, however, do not reflect the wealth of information on the connectivity of these structures, and employ greatly simplified treatments of its complex connectivity. We were interested in whether a more analytical approach, which begins with analysis of the connectivity of the system, might provide insights complementary to those derived by synthetic models. Accordingly, we collated detailed neuroanatomical information about the connectivity of the hippocampal system in the rat, and analysed the resulting data. Analyses of connectivity based on a variety of different analytical techniques have recently been used to elucidate the global organization of other systems in the macaque and cat, and have given rise to successful predictions. We applied non-metric multidimensional scaling and non-parametric cluster analysis to our summary matrix of connection data. The analyses produced organizational schemes that were consistent with known physiological properties and provided the basis for making tentative predictions of the further structures that may contain 'place' and 'head-direction' cells, which structures we identify. The consistency between the analyses of connectivity and the distribution of physiological properties across the system suggests that functional relationships are constrained by the organization of the connectivity of the system, and so that structure and function are linked at the systems level. PMID- 10703045 TI - Hierarchical organization of macaque and cat cortical sensory systems explored with a novel network processor. AB - Neuroanatomists have described a large number of connections between the various structures of monkey and cat cortical sensory systems. Because of the complexity of the connection data, analysis is required to unravel what principles of organization they imply. To date, analysis of laminar origin and termination connection data to reveal hierarchical relationships between the cortical areas has been the most widely acknowledged approach. We programmed a network processor that searches for optimal hierarchical orderings of cortical areas given known hierarchical constraints and rules for their interpretation. For all cortical systems and all cost functions, the processor found a multitude of equally low cost hierarchies. Laminar hierarchical constraints that are presently available in the anatomical literature were therefore insufficient to constrain a unique ordering for any of the sensory systems we analysed. Hierarchical orderings of the monkey visual system that have been widely reported, but which were derived by hand, were not among the optimal orderings. All the cortical systems we studied displayed a significant degree of hierarchical organization, and the anatomical constraints from the monkey visual and somato-motor systems were satisfied with very few constraint violations in the optimal hierarchies. The visual and somato-motor systems in that animal were therefore surprisingly strictly hierarchical. Most inconsistencies between the constraints and the hierarchical relationships in the optimal structures for the visual system were related to connections of area FST (fundus of superior temporal sulcus). We found that the hierarchical solutions could be further improved by assuming that FST consists of two areas, which differ in the nature of their projections. Indeed, we found that perfect hierarchical arrangements of the primate visual system, without any violation of anatomical constraints, could be obtained under two reasonable conditions, namely the subdivision of FST into two distinct areas, whose connectivity we predict, and the abolition of at least one of the less reliable rule constraints. Our analyses showed that the future collection of the same type of laminar constraints, or the inclusion of new hierarchical constraints from thalamocortical connections, will not resolve the problem of multiple optimal hierarchical representations for the primate visual system. Further data, however, may help to specify the relative ordering of some more areas. This indeterminacy of the visual hierarchy is in part due to the reported absence of some connections between cortical areas. These absences are consistent with limited cross-talk between differentiated processing streams in the system. Hence, hierarchical representation of the visual system is affected by, and must take into account, other organizational features, such as processing streams. PMID- 10703046 TI - Anatomical connectivity defines the organization of clusters of cortical areas in the macaque monkey and the cat. AB - The number of different cortical structures in mammalian brains and the number of extrinsic fibres linking these regions are both large. As with any complex system, systematic analysis is required to draw reliable conclusions about the organization of the complex neural networks comprising these numerous elements. One aspect of organization that has long been suspected is that cortical networks are organized into 'streams' or 'systems'. Here we report computational analyses capable of showing whether clusters of strongly interconnected areas are aspects of the global organization of cortical systems in macaque and cat. We used two different approaches to analyse compilations of corticocortical connection data from the macaque and the cat. The first approach, optimal set analysis, employed an explicit definition of a neural 'system' or 'stream', which was based on differential connectivity. We defined a two-component cost function that described the cost of the global cluster arrangement of areas in terms of the areas' connectivity within and between candidate clusters. Optimal cluster arrangements of cortical areas were then selected computationally from the very many possible arrangements, using an evolutionary optimization algorithm. The second approach, non-parametric cluster analysis (NPCA), grouped cortical areas on the basis of their proximity in multidimensional scaling representations. We used non-metric multidimensional scaling to represent the cortical connectivity structures metrically in two and five dimensions. NPCA then analysed these representations to determine the nature of the clusters for a wide range of different cluster shape parameters. The results from both approaches largely agreed. They showed that macaque and cat cortices are organized into densely intra-connected clusters of areas, and identified the constituent members of the clusters. These clusters reflected functionally specialized sets of cortical areas, suggesting that structure and function are closely linked at this gross, systems level. PMID- 10703048 TI - Global relationship between anatomical connectivity and activity propagation in the cerebral cortex. AB - Anatomical connectivity is a prerequisite for cooperative interactions between cortical areas, but it has yet to be demonstrated that association fibre networks determine the macroscopical flow of activity in the cerebral cortex. To test this notion, we constructed a large-scale model of cortical areas whose interconnections were based on published anatomical data from tracing studies. Using this model we simulated the propagation of activity in response to activation of individual cortical areas and compared the resulting topographic activation patterns to electrophysiological observations on the global spread of epileptic activity following intracortical stimulation. Here we show that a neural network with connectivity derived from experimental data reproduces cortical propagation of activity significantly better than networks with different types of neighbourhood-based connectivity or random connections. Our results indicate that association fibres and their relative connection strengths are useful predictors of global topographic activation patterns in the cerebral cortex. This global structure-function relationship may open a door to explicit interpretation of cortical activation data in terms of underlying anatomical connectivity. PMID- 10703049 TI - Nonlinear PCA: characterizing interactions between modes of brain activity. AB - This paper presents a nonlinear principal component analysis (PCA) that identifies underlying sources causing the expression of spatial modes or patterns of activity in neuroimaging time-series. The critical aspect of this technique is that, in relation to conventional PCA, the sources can interact to produce (second-order) spatial modes that represent the modulation of one (first-order) spatial mode by another. This nonlinear PCA uses a simple neural network architecture that embodies a specific form for the nonlinear mixing of sources that cause observed data. This form is motivated by a second-order approximation to any general nonlinear mixing and emphasizes interactions among pairs of sources. By introducing these nonlinearities principal components obtain with a unique rotation and scaling that does not depend on the biologically implausible constraints adopted by conventional PCA. The technique is illustrated by application to functional (positron emission tomography and functional magnetic resonance imaging) imaging data where the ensuing first- and second-order modes can be interpreted in terms of distributed brain systems. The interactions among sources render the expression of any one mode context-sensitive, where that context is established by the expression of other modes. The examples considered include interactions between cognitive states and time (i.e. adaptation or plasticity in PET data) and among functionally specialized brain systems (using a fMRI study of colour and motion processing). PMID- 10703047 TI - Computational analysis of functional connectivity between areas of primate cerebral cortex. AB - Recent analyses of association fibre networks in the primate cerebral cortex have revealed a small number of densely intra-connected and hierarchically organized structural systems. Corresponding analyses of data on functional connectivity are required to establish the significance of these structural systems. We therefore built up a relational database by systematically collating published data on the spread of activity after strychnine-induced disinhibition in the macaque cerebral cortex in vivo. After mapping these data to two different parcellation schemes, we used three independent methods of analysis which demonstrate that the cortical network of functional interactions is not homogeneous, but shows a clear segregation into functional assemblies of mutually interacting areas. The assemblies suggest a principal division of the cortex into visual, somatomotor and orbito-temporo-insular systems, while motor and somatosensory areas are inseparably interrelated. These results are largely compatible with corresponding analyses of structural data of mammalian cerebral cortex, and deliver the first functional evidence for 'small-world' architecture of primate cerebral cortex. PMID- 10703050 TI - On imputing function to structure from the behavioural effects of brain lesions. AB - What is the link, if any, between the patterns of connections in the brain and the behavioural effects of localized brain lesions? We explored this question in four related ways. First, we investigated the distribution of activity decrements that followed simulated damage to elements of the thalamocortical network, using integrative mechanisms that have recently been used to successfully relate connection data to information on the spread of activation, and to account simultaneously for a variety of lesion effects. Second, we examined the consequences of the patterns of decrement seen in the simulation for each type of inference that has been employed to impute function to structure on the basis of the effects of brain lesions. Every variety of conventional inference, including double dissociation, readily misattributed function to structure. Third, we tried to derive a more reliable framework of inference for imputing function to structure, by clarifying concepts of function, and exploring a more formal framework, in which knowledge of connectivity is necessary but insufficient, based on concepts capable of mathematical specification. Fourth, we applied this framework to inferences about function relating to a simple network that reproduces intact, lesioned and paradoxically restored orientating behaviour. Lesion effects could be used to recover detailed and reliable information on which structures contributed to particular functions in this simple network. Finally, we explored how the effects of brain lesions and this formal approach could be used in conjunction with information from multiple neuroscience methodologies to develop a practical and reliable approach to inferring the functional roles of brain structures. PMID- 10703051 TI - Purification and inactivation by substrate of an allene oxide synthase (CYP74) from corn (Zea mays L.) seeds. AB - The allene oxide synthase (AOS) was purified from corn (Zea mays) seeds to homogeneity and characterized partially. The corn AOS was a hemoprotein cytochrome P450 with a molecular weight and pI of 53,000 and 6.0, respectively. The corn AOS was found to be irreversibly inactivated by a substrate, 13 hydroperoxyoctadienoic acid. The rate of the enzyme inactivation was higher at low pHs. PMID- 10703052 TI - Purification and properties of urease from the leaf of mulberry, Morus alba. AB - Urease was purified from leaves of mulberry (Morus alba, L.) by ammonium sulfate fractionation, acetone fractionation and sequential column chromatography including Q-Sepharose HP, Phenyl-Sepharose HP, Superdex 200 HR and Mono Q. The enzyme was purified 5700-fold to apparent homogeneity with a recovery of 3.6%. The molecular mass of the enzyme was determined to be 90.5 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis and 175 kDa by gel filtration, indicating that the enzyme was a homodimer. In the western blot analysis, 90.5 kDa subunit of the mulberry leaf urease cross-reacted with antiserum raised against jack bean seed urease. The N-terminal sequence of the first 20 residues of the enzyme revealed that it has a high similarity (80-90%) to ureases from other plant sources, suggesting that the mulberry leaf urease is closely related to other plant ureases. However, the mulberry leaf enzyme showed an optimum pH for activity of 9.0, while the optimum pH of most ureases isolated from plants and bacterial is neutral. In addition, the K(m) value for urea was 0.16 mM, which is lower than those of ureases from other sources. It is also proposed that urease activity ingested by browsing silkworm releases ammonia that is subsequently used in silkworm protein synthesis. PMID- 10703053 TI - Variability of phenylpropanoid precursors in the biosynthesis of phenylphenalenones in Anigozanthos preissii. AB - Feeding experiments using 13C labelled precursors and NMR spectroscopic studies revealed general biosynthetic incorporation of phenylalanine and variable incorporation of cinnamic acid, p-coumaric acid, caffeic acid and ferulic acid into phenylphenalenones in root cultures of Anigozanthos preissii. Evidence was obtained for parallel pathways of phenylphenalenone biosynthesis, with respect to the left phenylpropanoid unit, and a sequence involving utilisation of p-coumaric acid with late generation of an intermediate catechol moiety in the right phenylpropanoid unit. PMID- 10703054 TI - Droserone from cell cultures of Triphyophyllum peltatum (Dioncophyllaceae) and its biosynthetic origin. AB - The growth and droserone content of callus cultures of Triphyophyllum peltatum grown in liquid 1/5 Linsmaier and Skoog medium was studied. During a lag phase in growth, droserone concentrations in the medium reached a value of 2.1 mg g-1 fr. wt. After this maximum value the concentration decreased slightly to 1.8 mg g-1 fr. wt., while the growth of the calli was enhanced (25% increase in fr. wt. within 7 days). Plumbagin and isoshinanolone were likewise present in the medium. By feeding 13C2-labelled acetate to the cultures the biosynthesis of droserone was elucidated. The incorporation of whole C2-units unambiguously shows its acetogenic origin and fits well in the biosynthetic scheme suggested for the structurally--and biogenetically--related naphthylisoquinoline alkaloids. PMID- 10703055 TI - Stimulation of the production of hypericins by mannan in Hypericum perforatum shoot cultures. AB - Shoot organ cultures were established from callus derived from anthers of Hypericum perforatum flowers and the effect of elicitors on hypericin and pseudohypericin production in shoot organ cultures was investigated. Mannan stimulated pseudohypericin production up to four fold (0.82 mg/g dry wt) and hypericin production up to two fold (0.04 mg/g dry wt.) beta-1,3-glucan and pectin slightly stimulated pseudohypericin production (ca. two fold), but had no effect on hypericin production. On the other hand, yeast extract showed no stimulatory effect, on either hypericin or pseudohypericin production. PMID- 10703056 TI - Anthracenone ABA analogue as a potential photoaffinity reagent for ABA-binding proteins. AB - An anthracenone analogue of abscisic acid (ABA) was synthesized as a potential photoaffinity reagent and tested for biological activity. Reaction between 10,10' dimethoxy-9-anthrone with two equivalents of the lithiated dianion of cis-3 methylpent-2-en-4-yn-1-ol afforded an acetylenic alcohol key intermediate. Subsequent reduction of the triple bond, functional group manipulation of the side chain alcohol and deprotection of the dimethoxy protected anthrone provided anthracenone ABA analogue 7 as a potential photoaffinity reagent for ABA-binding proteins. The effect of natural ABA and the potential photoaffinity anthracenone ABA 7 on corn cell growth was determined at various concentrations. The results show that anthracenone ABA 7 is perceived as ABA-like, although producing less inhibition than ABA itself. For example, 7 at 33 microM produces approximately the same inhibition as ABA at 10 microM. PMID- 10703057 TI - Galloylglucoses and riccionidin A in Rhus javanica adventitious root cultures. AB - Adventitious root cultures of Rhus javanica L. produced large amounts of galloylglucoses (gallotannins) and an anthocyanidin, riccionidin A, formerly found only in liverworts. Production of both galloylglucoses and riccionidin A in the adventitious root culture system was suppressed by light. The Rhus root culture showed the highest productivity for those secondary metabolites in a modified Linsmaier-Skoog (LS) liquid medium containing 30 mM NH4+ and 30 mM NO3- as nitrogen sources in the presence of 10(-6) M 3-indoleacetic acid (IAA). PMID- 10703058 TI - Altered lignin composition in phenylalanine ammonia-lyase-inhibited radish seedlings: implications for seed-derived sinapoyl esters as lignin precursors. AB - We earlier reported that when phenylalanine ammonia-lyase (PAL) activity in radish seedlings was inhibited by the competitive inhibitor 2-aminoindan-2 phosphonic acid (AIP), soluble sinapoyl esters carried over from the seed were converted to wall-bound esters in young cotyledons. We now report that these soluble sinapoyl esters may also be converted into lignin in the cotyledons. When radish seedlings were grown in the presence of 100 microM AIP, lignin formation (determined as lignothioglycolic acid) was inhibited ca. 74% in the cotyledons and ca. 80% in hypocotyls plus roots. The syringyl to guaiacyl (S/G) ratio in the lignin of AIP-grown plants, as determined by alkaline cupric oxidation and from Fourier-transform infrared (FT-IR) spectra, was higher in cotyledons, but lower in hypocotyls plus roots, as compared to plants grown on distilled water. These results support the view that soluble sinapoyl esters preformed in seeds may contribute to the syringyl moiety of lignin in cotyledons during early seedling development and that there is no appreciable transport of soluble sinapoyl esters from cotyledons to the hypocotyls and roots. PMID- 10703059 TI - Two insecticidal tetranortriterpenoids from Azadirachta indica. AB - Two new triterpenoids, 6 alpha-O-acetyl-7-deacetylnimocinol [24,25,26,27-tetra norapotirucalla-(apoeupha)-6 alpha-acetoxy-7 alpha-hydroxy-1,14,20,22-tetraen 21,23-epoxy-3-one] (1) and meliacinol [24,25,26,27-tetranorapotirucalla (apoeupha)-1 alpha-trimethylacryloxy-21,23-6 alpha,28-diepoxy-16-oxo-17-oxa 14,20,22-trien-3 alpha,7 alpha-diol] (2) were isolated from the methanolic extract of the fresh leaves of Azadirachta indica (neem). Their structures have been elucidated through spectral studies, including 2D-NMR (COSY-45, NOESY, HMQC and HMBC). The bioactivity of these as well as of nimocinol, reported earlier from the same source, is reported. The first compound and nimocinol showed toxicity on fourth instar larvae of mosquitoes (Aedes aegypti) with LC50 values of 21 and 83 ppm, respectively. The second compound had no effect upto 100 ppm. PMID- 10703060 TI - Uptake of radiolabelled ochratoxin A from soil by coffee plants. AB - [3H, 14C] Ochratoxin A, prepared biosynthetically, was applied in dilute NaHCO3 solution to the soil in which coffee plants had grown to four pairs of leaves. Three weeks later the compound, isolated from dilute NaHCO3 extract of leaves by immunoaffinity chromatography, was detected by scintillation counting as a 1-2 ppm component of leaf dry weight, greatly exceeding the trace (ppb) occurrence of ochratoxin A in some green coffees, which therefore might arise in the field directly from fungal activity in soil rather than from fungal infection of cherries or processed green coffee. PMID- 10703061 TI - Polyhydroxylated pyrrolidine and piperidine alkaloids from Adenophora triphylla var. japonica (Campanulaceae). AB - Adenophora triphylla var. japonica (Campanulaceae) yielded two new alkaloids, the 6-C-butyl derivative of 2R,5R-bis(hydroxymethyl)-3R,4R-dihydroxypyrrolidine (DMDP) and alpha-1-C-ethyl-fagomine, together with the known alkaloids 1,4 dideoxy-1,4-imino-D-arabinitol, 1-deoxynojirimycin, and 1-deoxymannojirimycin. 6 C-Butyl-DMDP showed inhibitory activity toward almond beta-glucosidase (IC50 = 68 microM), whereas alpha-1-C-ethyl-fagomine inhibited bovine liver beta galactosidase (IC50 = 29 microM). PMID- 10703062 TI - Screening of the needles of different yew species and cultivars for paclitaxel and related taxoids. AB - The needles of several yew species and cultivars were analysed by high-pressure liquid chromatography for paclitaxel, 10-deacetylpaclitaxel, cephalomannine, baccatin III, 10-deacetylbaccatin III and brevifoliol. About 750 samples were collected from five different locations in the Netherlands and the UK. The results of this screening show a large variation in taxane content between the different species and cultivars. The content of paclitaxel and 10 deacetylbaccatin III varied from 0 to 500 micrograms/g and 0 to 4800 micrograms/g dried needles, respectively. Brevifoliol was found in a very high concentration in Taxus brevifolia. 10-Deacetylpaclitaxel, cephalomannine and baccatin III were found in concentrations ranging from 0 to 500 micrograms/g dried needles. PMID- 10703063 TI - Importance of a pyrogallol-type structure in catechin compounds for apoptosis inducing activity. AB - Several catechin compounds were examined for their ability to induce apoptosis in human histiocytic lymphoma U937 cells. Catechins with a pyrogallol-type structure in a B-ring induced apoptosis and a 3-O-gallate group in cis-relationship to the B ring enhanced the activity. Catechins without a pyrogallol-type structure in a molecule lacked activity. These data suggest the important role of the 5'(3') hydroxyl group in the B-ring and that a pyrogallol-type structure in a molecule is a minimum requirement for apoptosis induction by catechin compounds. PMID- 10703064 TI - The biotransformation of 18-hydroxy-9-epi-ent-pimara-7,15-diene by Gibberella fujikuroi. AB - Incubation of 18-hydroxy-9-epi-ent-pimara-7,15-diene with the fungus Gibberella fujikuroi gave the compounds 18-hydroxy-7 alpha,8 alpha-epoxy-9-epi-ent-pimara-15 ene, 18-hydroxy-7-oxo-ent-pimara-15-ene, 6 beta, 18-dihydroxy-7 alpha, 8 alpha epoxy-9-epi-ent-pimara-15-ene, 9 beta,18-dihydroxy-7 alpha, 8 alpha-epoxy-ent pimara-15-ene and 6 beta, 14 alpha, 18-trihydroxy-9-epi-ent-pimara-7,15-diene. Oxidation of C-19, which is characteristic of the biosynthesis pathway of the gibberellins is not produced. PMID- 10703065 TI - Chalcanol glucosides from seeds of Trifolium alexandrinum. AB - Three new chalcanol glucosides have been isolated from the seeds of Trifolium alexandrinum, of which the first two are alpha'-chalcanol-alpha, beta-epoxides and the third one is an alpha, beta-dihydroxy-alpha'-chalcanol. The structures of the isolated compounds were verified by means of MS and 2D NMR spectral analyses. PMID- 10703066 TI - Phenolic compounds from the leaves of Cornus controversa. AB - Two novel phenolic compounds from the leaves of Cornus controversa (Cornaceae) were characterized as (-)-2,3-digalloyl-4-(E)-caffeoyl-L-threonic acid and (-)-2 galloyl-4-(E)-caffeoyl-L-threonic acid, using spectroscopic methods. PMID- 10703067 TI - Tropane alkaloids from Erythroxylum zeylanicum O.E. Schulz (Erythroxylaceae). AB - Six tropane alkaloids were isolated from the Sri Lankan endemic plant Erythroxylum zeylanicum O.E. Schulz (Erythroxylaceae) and structurally elucidated by NMR and MS measurements. Three of them, erythrozeylanines A [1R,3R,5S,6R-6 acetoxy-3-(3',4',5'-trimethoxybenzoyloxy)tropane], B [cis-3 beta (cinnamoyloxy)tropane], and C [cis-6 beta-acetoxy-3 alpha-(cinnamoyloxy)tropane] are new, whereas the others have already been found in other Erythroxylum species. For the first time, the absolute configuration of a tropane alkaloid (erythrozeylanine A) has been determined by quantum chemical CD calculations. PMID- 10703068 TI - Steroidal alkaloids from Cryptolepis obtusa. AB - Two novel diglycosylated steroidal alkaloids of 5 delta-pregnene nucleus, named obtusine-20(R)-O-[beta-thevetopyranosyl-(1-->4)-beta-cyma ropyranoside] and obtusolactam-20(R)-O-[beta-thevetopyranosyl-(1-->4)-beta- cymaropyranoside], together with the known beta-sitosteryl-3-O-beta-glucopyranoside were isolated from the roots of Cryptolepis obtusa N. E. Br. PMID- 10703069 TI - [Listeriosis: an old or a current health problem?]. PMID- 10703070 TI - [Outcome of patients with systemic rheumatic diseases admitted to intensive care units: a retrospective study of 39 cases]. AB - PURPOSE: Patients with systemic rheumatic diseases are rarely admitted in intensive care units and very few studies focusing on the prognosis of those patients have been published. METHODS: Retrospective study over seven years in two intensive care units. RESULTS: Among 33 patients with systemic disease diagnosed 90 +/- 133 months before admission in the intensive care unit, who were aged 50 +/- 21 years and represented a total of 39 stays in the intensive care unit, the main cause of admission was acute respiratory failure (33%). Mean simplified acute physiology score (SAPS II) was 47 +/- 22. Two-thirds of the patients were under mechanical ventilation. Infection was diagnosed in 33% of the cases and exacerbation of the systemic rheumatic disease in 26%. Nosocomial infection was found in 19 patients (49%). Ten patients died during their stay in the intensive care unit, six from infection, three from an exacerbation of the systemic rheumatic disease, one from an unidentified cause. CONCLUSION: Even if severity scores of patients suffering from systemic diseases are higher at admission in intensive care units than those of other patients, there is no relevant reason to refuse critical care to these patients. PMID- 10703071 TI - [Hemorrhagic syndromes related to selective serotonin reuptake inhibitor (SSRI) antidepressants. Seven case reports and review of the literature]. AB - PURPOSE: Selective serotonin reuptake inhibitors (SSRIs) are widely prescribed. Since their release unexpected adverse effects such as bleeding disorders have been described. METHODS: Thirty patients with either hematoma or muco-cutaneous bleeding have been selected from case reports of the Saint-Etienne Pharmacovigilance center and from a literature review. RESULTS: The female/male sex-ratio was 3:4 and the mean age 42 years. Two newborns who had been exposed in utero to SSRIs were also included in the study. Eleven patients presented an underlying disease or were at risk. Various adverse effects such as bruising, hematoma, petechiae or purpura, epistaxis, and more rarely intestinal hemorrhage, ocular bleeding or cerebral hemorrhage were encountered. Symptoms were sometimes associated with prolonged bleeding time and platelet aggregation disorders and usually resolved within two days to four months after treatment discontinuation. CONCLUSION: Hematoma and muco-cutaneous bleeding would therefore be related to treatment, including selective serotonin reuptake inhibitors. However, these adverse effects are still poorly known and rarely reported. The main mechanism suggested would be a decrease in serotonin platelet leading to a defect in platelet aggregation. However, an increase in capillary fragility or susceptibility related to the patient's condition might be involved. Study of hemostasis history in patients requiring treatment with SSRIs might be of value. PMID- 10703072 TI - [Clinical or biological symptoms leading to the search for amyloidosis]. AB - INTRODUCTION: The extracellular, multifocal, disseminated or diffuse localization of amyloidosis accounts for the diversity of clinical presentations and late diagnosis. CURRENT KNOWLEDGE AND KEY POINTS: However, early diagnosis is important, as new drugs have been recently introduced. Both clinical picture and biology help guide diagnosis, including either kidney, heart, skin or neurologic involvement associated with monoclonal gammapathy (primary idiopathic amyloidosis AL); underlying inflammatory or infectious disease, familial Mediterranean fever with proteinuria (secondary amyloidosis AA); cardiac, neurologic or ocular involvement (heredofamilial amyloidosis); carpal tunnel syndrome, joint pain (amyloidosis of hemodialysis). Furthermore, amyloid fibrils are identified on salivary gland biopsy. FUTURE PROSPECTS AND PROJECTS: Due to the introduction of new specific drugs aimed at curing various amyloidoses, early diagnosis is important. Chemotherapy and hematopoietic stem cell transplantation are promising regarding AL amyloidosis, while liver transplantation has proven remarkably successful in heredofamilial amyloidosis. Progress in molecular biology should allow identification of various forms of familial amyloidosis. PMID- 10703073 TI - [Serum p53 antibody assay: evaluation in colorectal cancer]. AB - INTRODUCTION: Alterations of the tumor suppressor gene p53 and its protein synthesis is the most commonly observed genetic feature in human cancers. Direct diagnosis of the gene mutation using sequencing is the gold standard method. However, it requires advanced technology and is only performed in specialized research units. CURRENT KNOWLEDGE AND KEY POINTS: Demonstration of intratumoral p53 protein accumulation using immunohistochemistry is a routine diagnostic technique. Serum detection of p53 antibodies using ELISA has been recently developed. It is an easily feasible and reproducible method for the diagnosis of p53 alterations due to self-immunization in some patients in response to intratumoral p53 protein overexpression. This phenomenon is inconstant (about one third of the patients with a p53 gene mutation produce antibodies) and its mechanism is unclear. p53 Antibodies are found in 25% of the patients with colorectal cancer, independently of traditional tumor markers (carcinoembryonic antigen and carbohydrate antigen 19.9). The presence of these antibodies is not linked to the tumor stage. Since their ratios vary during the treatment, they might constitute a new tumor marker. FUTURE PROSPECTS AND PROJECTS: Early appearance of p53 serum antibodies during tumor development should make them useful for the detection of malignant transformation in patients with preneoplastic disease such as ulcerous colititis. Whether the presence of p53 antibodies in colorectal cancer patients has a prognostic significance requires further assessment. PMID- 10703074 TI - [Viral quantitation of hepatitis C virus: present and future]. AB - INTRODUCTION: Hepatitis C virus (HCV) plasmatic load, also called HCV viremia, is expressed in copies per milliliter or viral particle equivalents per milliliter. CURRENT KNOWLEDGE AND KEY POINTS: The methods used to measure HCV viremia are based on two principles: the quantitation of viral proteins (measure of antigenemia using monoclonal antibodies), and the quantitation of viral nucleic acid using PCR after reverse transcription (RT-PCR), or signal amplification (branched DNA). There is no relationship between viremia and liver histologic activity. FUTURE PROSPECTS AND PROJECTS: The use of HCV quantitative viremia in a treatment algorithm, including other factors such as age, genotype, degree of viral genetic heterogeneity, and liver fibrosis score will help adapt therapeutic associations for each individual. PMID- 10703075 TI - [Delusional parasitosis or Ekbom's syndrome]. AB - INTRODUCTION: Ekbom's syndrome or delusional parasitosis is a disorder in which the patient believes that he/she is infected by a parasite. Epidemiologic, nosologic, psychopathologic and therapeutic data can barely be interpreted, as delusional parasitosis has mostly been described in either isolated cases or small cohorts. An extensive literature review is recommended to better understand common features associated with this syndrome. EXEGESIS: Ekbom's syndrome is a chronic disorder that may occur at any age but is more common in the elderly, particularly in females. International classifications have included this syndrome in non-schizophrenic delusions. However, it has also been reported in schizophrenia, affective disorders, and organic or induced psychosis. Treatment is based on antipsychotic agents, psychotherapy, and cooperation between dermatologists and psychiatrists. CONCLUSION: Pimozide is currently the most effective treatment. It requires careful monitoring, as it has several adverse effects. For patients with concomitant depressive symptoms, the use of antidepressants is recommended. PMID- 10703076 TI - [Atypical presentation of tuberculosis in the elderly: a case report of pleuropericarditis]. AB - INTRODUCTION: The incidence of tuberculosis is increasing, particularly in the elderly, and has various clinical presentations. EXEGESIS: We describe the case of a 78-year-old woman who presented tuberculous pleuropericarditis. This case is atypical, due to infection localisation, negativity of the tuberculin skin test, and mixed pleural effusion. Following antituberculosis antibiotic therapy and corticotherapy, the outcome was favorable. CONCLUSION: Due to atypical and non specific clinical presentation, diagnosis of tuberculosis may be particularly difficult in the elderly. PMID- 10703077 TI - [Risk of serious acute asthma due to lupine flour associated with peanut allergy]. AB - Lupine flour (lupinus albus), recently authorized in France in human food, cross reacts with peanuts. We report a case of acute asthma in a patient with allergy to peanuts. EXEGESIS: This patient has a severe allergy to peanuts, presenting as acute asthma. Skin prick-tests to raw and cooked lupine flour were positive. The level of specific-IgE (Allerbio, France) to lupine flour were high. Oral challenge test induced acute asthma at a dose of 965 mg of lupine flour. This quantity may be included in 100 g of bread. CONCLUSION: This case report points out the fact that lupine flour is a high-risk allergen in patients presenting allergy to peanuts. It is necessary to evaluate the allergenic risk of new foods before their introduction into human daily food intake and to establish a network of allergy vigilance. PMID- 10703078 TI - [Point of inquiry in the head! Metastasis of the skull in lung cancer following breast cancer]. PMID- 10703080 TI - [A rare complication of lumbar puncture: the spinal subdural hematoma]. PMID- 10703079 TI - [Atypical chest pain]. PMID- 10703081 TI - [Cytomegalovirus infection of the submandibular gland in a patient with HIV infection]. PMID- 10703082 TI - [Discovery of testicular microlithiasis in the course of Ureaplasma urealyticum urethritis]. PMID- 10703083 TI - [Salmonella montevideo hypoxemic pleuropneumonia]. PMID- 10703084 TI - [Association of vitiligo and laryngeal carcinoma: a case report]. PMID- 10703085 TI - [Cryptogenic chronic cytolysis: think about celiac disease]. PMID- 10703086 TI - Scottish medicine. The past and the future. PMID- 10703087 TI - The development of medical ethics for the new millennium. PMID- 10703088 TI - The development of stroke services: entering the new millennium. PMID- 10703089 TI - Revisiting established traditions. New horizons for geriatric medicine in the 21st century. PMID- 10703090 TI - Renal replacement therapy. PMID- 10703091 TI - The development of aviation medicine: towards greater passenger safety. PMID- 10703092 TI - Pulmonary tuberculosis in elderly people in Scotland. PMID- 10703093 TI - A medical bouquet. Poppies, cinchona and willow. AB - The poppy, the cinchona tree and the willow tree have all provided medicines of undoubted value during past millennium and form a "medical bouquet" worthy of consideration as this century ends. The use of these plants by ancient peoples has gradually evolved into a more informed understanding and application of the compounds initially isolated and purified from their extracts. There has been a Scottish contribution to the medical history of all three plants. PMID- 10703094 TI - Developments in imaging. An historical review. AB - Following the discovery of X-rays by Roentgen in 1895, it was 70 years later that the specialty of imaging really took off, with ultrasound, CT, Nuclear medicine, and MRI now an integral part of all large departments. The recent progress has been accelerated by advances in computer technology which aided the development of imaging techniques that do not use ionising radiation. Some imaging techniques are replacing other more hazardous diagnostic procedures, (e.g. MRCP v ERCP, and MRA v catheter angiography) while some are able to provide almost perfect anatomical detail e.g. brain MRI. Techniques using radioisotopes provide more functional information which can be used to complement the morphological studies. We have witnessed the birth and maturation of Interventional Radiology. This is a highly skilled subspecialty enabling some surgical procedures to be replaced by minimally invasive methods using image guidance. These range from simple drainage procedures and guided biopsies to complex aortic bifurcation stent graft insertions. The potential for digital storage of all images is already established and the filmless radiology department in the more developed countries could be the norm by 2020. PMID- 10703095 TI - Newly emerging infections. A long look back to the future. PMID- 10703096 TI - Early aspects of the medical care of the poor in Scotland. PMID- 10703097 TI - Introducing new technology: a stepwise algorithm. PMID- 10703098 TI - The anatomic relation among the nerve roots, intervertebral foramina, and intervertebral discs of the cervical spine. AB - STUDY DESIGN: An anatomic study of the cervical intervertebral foramina, nerve roots, and intradural rootlets performed using a surgical microscope. OBJECTIVES: To investigate the anatomy of cervical root compression, and to obtain the anatomic findings related to cervical foraminotomy for the treatment of cervical radiculopathy. SUMMARY OF BACKGROUND DATA: Cervical foraminotomy is a procedure performed frequently for the management of cervical radiculopathy. However, anatomic studies of cervical foraminotomy have not been fully elucidated. METHODS: In this study, 18 cadavers were obtained for the study of the cervical spine. All the soft tissues were dissected from the cervical spine. Thereafter, laminectomy and facetectomy were performed on C4 through T1 using a surgical microscope. The nerve roots and surrounding anatomic structures, including intervertebral discs and foramina, were exposed. In addition, the intradural rootlets and their intersegmental connections were observed. RESULTS: The shape of the intervertebral foramina approximated a funnel, the entrance zone being the most narrow part and the root sleeves conical, with their takeoff points from the central dural sac being the largest part. Therefore, compression of the nerve roots occurred at the entrance zone of the intervertebral foramina. Anteriorly, compression of the nerve roots was caused by protruding discs and osteophytes of the uncovertebral region, whereas the superior articular process, the ligamentum flavum, and the periradicular fibrous tissues affected the nerve posteriorly. The C5 nerve roots were found to exit over the middle aspect of the intervertebral disc, whereas the C6 and C7 nerve roots were found to traverse the proximal part of the disc. The C8 nerve roots had little overlap with the C7-T1 disc in the intervertebral foramen. The C6 and C7 rootlets passed two disc levels in the dural sac. Also, a high incidence of the intradural connections between the dorsal rootlets of C5, C6, and C7 segments was found. CONCLUSIONS: This study demonstrated the anatomy of the nerve roots, rootlets, and intervertebral foramina, and may aid in understanding the pathology of cervical radiculopathy. The presence of intradural connections between dorsal nerve roots and the relation between the course of the nerve root and the intervertebral disc may explain the clinical variation of symptoms resulting from-nerve root compression in the cervical spine. To perform cervical foraminotomy for cervical radiculopathy, it is necessary to understand the detailed anatomy of the intervertebral foramina thoroughly. PMID- 10703099 TI - Internal architecture of the sacrum in the elderly. An anatomic and radiographic study. AB - STUDY DESIGN: A description of the internal architecture of the sacrum, including its trabecular arrangement, cortical thickness, and overall bone density. OBJECTIVES: To determine the strong and weak areas in the sacrum to understand more clearly the sacral structure and its clinical implications. METHODS: First, seven cadaveric sacral specimens were sectioned in different planes. Horizontal sections were performed at the upper S1, middle S1, S2, S3, and S4. Sagittal sections were made through the median sacral crest, the sacral foramina, and medial to the articular surface. A coronal section through the whole length of a sacral specimen was produced. All sections were studied radiographically, and the trabecular pattern was analyzed. In the second part of the study, axial computed tomography scans of 40 dry sacrum specimens were analyzed by using the National Institutes of Health Image 1.61 program. The cortical thickness and bone density were determined. RESULTS: In the upper sacrum, three distinctive distributions of bony trabeculae were noted, one extending from the center of the sacral body anterolaterally, and the other two extending from the pedicle toward the auricular surface. A condensation zone was observed at the intersection of these trabeculae and was located at the anterior cortex of the foraminal zone. The junction between S2 and S3 represented a weak area with abrupt disappearance of the condensation zone. Analysis of the bone density of the sacrum using the plot analysis demonstrated that, at S1 and S2, the anterior cortex of the foraminal zone (condensation zone) is the most compact part of the sacrum. CONCLUSION: These results suggest that the strongest part of the sacrum is the anterior cortex above the foramina in S1 and S2. The weakest point of the sacrum was found to lie at the level of the junction of S2 and S3. PMID- 10703100 TI - Experimental study on the dynamics of lumbosacral nerve root circulation. AB - STUDY DESIGN: Experimental investigation of the dynamics of nerve root circulation. OBJECTIVES: To study the dynamics of lumbosacral nerve root circulation by using seriography in dogs. SUMMARY OF BACKGROUND DATA: The vascular distribution to the nerve root has been discussed mainly from the morphologic aspects, and no adequate elucidation has been presented concerning the kinetics of the blood supply to the nerve root. METHODS: To investigate the direction of blood flow in the nerve roots, a series of photographs of the cauda equina were taken using a motor-driven camera immediately after 3 mL of india ink was injected through the aortic catheter manually. The changes in the blood flow direction caused by compression of the nerve root also were observed. After the dog was killed, the nerve roots were cleared by the Spalteholz technique to identify the vessels observed during the experiment. RESULTS: The blood flow in the radicular arteries was descending in the proximal part and ascending in the distal part of the nerve roots. This observation supports the suggestion that there is a so-called watershed of the blood flow in the radicular arteries themselves. However, when the ascending radicular artery of the nerve root was cramped, the radicular blood flow on the proximal side was downward. The microangiograms also showed that there were abundant anastomoses of intrinsic vessels in the nerve roots. CONCLUSIONS: There is no relatively hypovascular region in the nerve root that is vulnerable in the course of degenerative changes in the lumbosacral spine. Therefore, it is unlikely that the watershed represents a weak point of the blood flow in the nerve root. PMID- 10703101 TI - Biomechanical studies of a dynamized anterior thoracolumbar implant. AB - STUDY DESIGN: An in vitro investigation into the biomechanical properties of a dynamized anterolateral compression implant that allows controlled subsidence. OBJECTIVES: To determine the extent to which both modes of the anterolateral compression implant (controlled collapsing and rigid) are able to reestablish the stability of the lumbar spine after L4 corpectomy. SUMMARY OF BACKGROUND DATA: Over time, anterior and posterior spinal implants have been associated with progressive angulation, and occasionally implant failure and breakage. To circumvent this occurrence and provide better graft loading, dynamized or collapsing devices for clinical use have been developed. METHODS: Eight fresh calf spines (L1-L6) were placed in a biomechanical testing frame. Pure moments of 6 Nm were loaded onto the intact spine in six directions: flexion, extension, right and left lateral bending, and right and left axial rotation. A total L4 corpectomy then was performed, and the defect grafted with a wooden dowel. Loading was repeated after the specimens were stabilized using the two modes of the anterolateral compression implant in succession. RESULTS: The results showed that both modes of the implant (the rigid mode in particular) restore the stiffness of the unstable spine to normal levels of flexion, extension, and right and left lateral bending, even to levels exceeding normal. These devices, however, fall short of achieving normal stability in right and left axial rotation. CONCLUSION: In the cadaveric calf spine after L4 corpectomy, restoration of stability with a dynamized anterior spinal implant is possible in flexion, extension, and right and left lateral bending, but not in axial rotation. PMID- 10703102 TI - Viscoelastic finite-element analysis of a lumbar motion segment in combined compression and sagittal flexion. Effect of loading rate. AB - STUDY DESIGN: A study using a validated viscoelastic finite-element model of a L2 L3 motion segment to identify the load sharing among the passive elements at different loading rates. OBJECTIVE: To enhance understanding concerning the role of the loading rate (i.e., speed of lifting and lowering during manual material handling tasks) on the load sharing and safety margin of spinal structures. SUMMARY OF BACKGROUND DATA: Industrial epidemiologic studies have shown that jobs requiring a higher speed of trunk motion contribute to a higher risk of industrial low back disorders. Consideration of the dynamic loading characteristics, such as lifting at different speeds, requires modeling of the viscoelastic behavior of passive tissues. Detailed systematic analysis of loading rate effects has been lacking in the literature. METHODS: Complex flexion movement was simulated by applying compression and shear loads at the top of the upper vertebra while its sagittal flexion angle was prescribed without constraining any translations. The lower vertebra was fixed at the bottom. The load reached its maximum values of 2000 N compression and 200 N anterior shear while L2 was flexed to 10 degrees of flexion in the three different durations of 0.3, 1, and 3 seconds to represent fast, medium, and slow movements, respectively. The resisted bending moment, gross load-displacement response of the motion segment, forces in facet joints and ligaments, stresses and strains in anulus fibrosus, and intradiscal pressure were compared across different rates. RESULTS: The distribution of stress and strain was markedly affected by the loading rate. The higher loading rate increased the peak intradiscal pressure (12.4%), bending moment (20.7%), total ligament forces (11.4%), posterior longitudinal ligament stress (15.7%), and anulus fiber stress at the posterolateral innermost region (17.9%), despite the 15.4% reduction in their strain. CONCLUSIONS: Consideration of the time-dependent material properties of passive elements is essential to improving understanding of motion segment responses to dynamic loading conditions. Higher loading rate markedly reduces the safety margin of passive spinal elements. When the dynamic tolerance limits of tissues are available, the results provide bases for the guidelines of safe dynamic activities in clinics or industry. PMID- 10703103 TI - Spinous process strength. AB - STUDY DESIGN: Mechanical testing of cadaveric lumbar spines and dual energy radiograph absorptiometry scanning were performed. OBJECTIVES: To devise a technique to measure the strength of lumbar spinous processes and to determine the bone mineral density of the vertebrae used. SUMMARY OF BACKGROUND DATA: The spinous process has been identified as the weakest part of the anatomy to which a flexible fixation device can be attached. It was unknown if the spinous processes could withstand the forces applied by the device. METHODS: A hook was fitted to the spinous process of 32 lumbar vertebrae. A custom-built rig was designed to secure a vertebra to a materials testing machine. A loop of cord was passed over a bar mounted on the crosshead of the machine and around the two bollards of the hook. As the crosshead was raised, a tension was applied to the cord. Each vertebra was tested to failure. The bone mineral density of each vertebra was then measured using dual energy radiograph absorptiometry. RESULTS: Failure of the specimens occurred by failure of the spinous process, pedicles, or vertebral body. The logarithm (base 10) of the load (N) at which failure occurred was 2.53 +/- 0.3, which corresponded to a mean failure load of 339 N. The bone mineral density of each vertebral body varied between 0.263 and 0.997 g/cm2. A significant linear correlation was found between bone strength and bone mineral density (P < 0.0001). CONCLUSIONS: Specimens with a bone mineral density in the range of 0.263-0.997 g/cm2 failed at a mean load of 339 N when the load was applied through the spinous process hook of a flexible fixation device. PMID- 10703104 TI - Criterion validity of the cervical range of motion (CROM) goniometer for cervical flexion and extension. AB - STUDY DESIGN: This study used a validity protocol. OBJECTIVE: To estimate the criterion validity of the Cervical Range of Motion goniometer using a healthy population. SUMMARY OF BACKGROUND DATA: The results of the 1994 study by Mayo et al show that there are no validated tools currently available for clinically measuring the cervical range of motion. Numerous decisions regarding patient status and treatment are based wholly or in part on joint motion measurements. Because of current budgetary restrictions, clinicians are being asked to justify their interventions objectively, and to do so, they will need validated tools. METHODS: The population consisted of 31 healthy participants ranging in age from 18 to 45 years. None had experienced cervical problems in the previous 3 months or were pregnant. Data collection took place at the radiology department. After participants were positioned on a stool, the cervical range of motion goniometer frame was set on their head by the physiotherapist. With the participant in this neutral position, the physiotherapist took the first Cervical Range of Motion measurement. The radiograph technologist obtained the radiograph immediately afterward. This procedure was repeated with the participant in fully flexed and fully extended positions. RESULTS: A Pearson's r correlation test was used to evaluate the criterion validity of the Cervical Range of Motion goniometer versus the radiographic method. The two measurements proved to be highly correlated (flexion: r = 0.97, P < 0.001; extension: r = 0.98, P < 0.001). CONCLUSIONS: For this population of healthy participants, the Cervical Range of Motion goniometer was found to be valid for measurements of cervical flexion and extension. Further research is needed on the validity of this instrument for other cervical spine movements. PMID- 10703105 TI - Central cord injury complicating acute cervical disc herniation in trauma. AB - STUDY DESIGN: A retrospective study on 24 patients with acute central cervical cord injury caused by traumatic disc herniation. OBJECTIVES: To determine the correlation of disc herniation with central cord injury and to evaluate the role of anterior cervical decompression and interbody fusion in management of this injury. SUMMARY OF BACKGROUND DATA: Acute cervical disc herniation has been documented as a causative factor in spinal cord injury but has been infrequently reported with central cord syndrome. METHODS: Between 1989 and 1994, 24 patients with acute cervical disc herniation and central cord syndrome were studied. These patients underwent anterior decompression and fusion, and were followed for 2 to 7 years, with an average follow-up period of 3 years and 8 months. The degree of disc herniation and neurologic scores were rated. RESULTS: During follow-up period, the American Spinal Injury Association motor score in 24 patients was increased to 86.46 +/- 10.22 from 47.79 +/- 19.66. The age of the patients was very negatively correlated with recovery rate (P < 0.01), but no correlation was observed between severity of cord compression and neurologic scores (P > 0.05). Postoperative neurologic improvement in patients with fracture or dislocation was very significantly slower (P < 0.01) than in those without these injuries. CONCLUSIONS: Far more common than previously expected, acute disc herniation in cervical spine injury is the one of principal cause for central cord syndrome. Magnetic resonance imaging assessment and surgical intervention are required. PMID- 10703106 TI - C1-C2 intra-articular screw fixation for atlantoaxial posterior stabilization. AB - STUDY DESIGN: A trial of a new posterior stabilization technique for atlantoaxial instability and a report of preliminary results. OBJECTIVES: To describe a new posterior stabilization technique for atlantoxial instability. SUMMARY OF BACKGROUND DATA: Magerl's transarticular screw fixation is an accepted technique for rigid atlantoaxial stabilization, which reportedly has yielded many good clinical results. However, the technique is technically demanding and poses a risk of injury to the nerves and veins. METHODS: Eleven patients who had been treated with intra-articular screw fixation in combination with Halifax interlaminar clamp (OSTEONICS, Allendale, NJ) for atlantoaxial instability were observed. Results of their clinical examinations and biomechanical studies using resinous bones of a cervical spine model were reviewed. RESULTS: In all patients, occipital pain, neck pain, and neural deficit improved, and bony fusion with no correction loss was shown on radiography. To date, no vascular or neural complications have been found, and no instrumentation failures have occurred. In the biomechanical study, the Halifax with transarticular screw fixation had significantly greater flexion stiffness than the Halifax only or the Halifax with intra-articular screw fixation, but the torsion stiffness of the Halifax with intra-articular screw fixation was significantly greater than that of the other Halifax combinations. CONCLUSION: The preliminary results showed that this technique was effective in strengthening the rotational stability of the atlantoaxial fixation and was considered useful for atlantoaxial posterior stabilization. PMID- 10703107 TI - Progression of lumbosacral isthmic spondylolisthesis in adults. AB - STUDY DESIGN: A retrospective clinical and radiographic review of adult patients with progressive isthmic lumbosacral spondylolisthesis. OBJECTIVES: To describe the clinical presentation of adult-onset progression of isthmic spondylolisthesis and to analyze its causes. SUMMARY OF BACKGROUND DATA: Until recently, progression of lumbosacral spondylolisthesis in adults was rarely reported. On the contrary, although slip progression before skeletal maturity has been widely recorded, its occurrence in adults has been doubted. Only sporadic case reports of adult slip progression and only brief notes on the subject in clinical studies describing other aspects of spondylolisthesis have been published. METHODS: Patients with isthmic lumbosacral spondylolisthesis who had serial radiographs of the lumbar spine on which slip progression during adult life was noted were evaluated. The amount of vertebral slip was calculated in millimeters from decubitus lateral spinal radiographs. The calculation was expressed as the percentage of slipped vertebral body length. RESULTS: From 1989 to 1995, 18 patients (9 women and 9 men), ages 32 to 55 years, with documented adult isthmic slip progression were identified at the Spinal Surgery Unit of the Hadassah University Hospital. All patients reported incapacitating low back pain, accompanied in most by significant sciatica. Documented slip progression ranged from 9% to 30% (average, 14.6%), and occurred during a period of 2 to 20 years (average duration, 6.8 years). Slip progression started after the third decade of life and coincided with marked disc degeneration at the olisthetic level. Slip progression associated with disc degeneration (i.e., intervertebral space narrowing and the formation of spondylophytes and vacuum phenomenon) brought about severe clinical symptomatology related to segmental instability and spinal stenosis. Of the 18 patients in this study, 14 were treated with surgery. All these patients except 1 underwent decompression, pedicle screw fixation, and bilateral lateral fusion. One patient underwent posterolateral fusion without instrumentation. Immediate postoperative complications were observed in three patients, including two superficial wound infections and one transient foot drop. Solid fusion was obtained in 11 of the 14 patients who underwent surgery. CONCLUSIONS: The concurrent occurrence of disc degeneration at the slip level and adult slip progression explains how an asymptomatic developmental lesion, present for at least two to three decades, may become symptomatic. PMID- 10703108 TI - Medical realities of cauda equina syndrome secondary to lumbar disc herniation. AB - STUDY DESIGN: An analysis of 44 cauda equina syndrome cases. OBJECTIVES: To determine the neurologic outcome of cauda equina syndrome cases, in light of the significant medical implications of this disorder. SUMMARY OF BACKGROUND DATA: Cauda equina syndrome from lumbar disc herniation accounts for up to 1% of all disc herniations. Most of the literature supports surgery within 24 hours as a means of improving the outcome. METHODS: A retrospective chi 2 analysis was performed of 44 patients surgically treated for lumbar disc herniation who initially sought treatment for cauda equina syndrome. RESULTS: In 20 patients, diagnosis was made and surgery performed within 48 hours of the cauda equina syndrome onset, including 18 patients (90%) who underwent surgery within 24 hours. In 24 patients, surgery was performed more than 48 hours after the onset of cauda equina syndrome, with a mean delay of 9 days, including 17 patients (71%) with a mean delay of 3.7 days. Causes for delay were patient-related in 4 cases (17%) and physician-related in 20 cases (83%). According to chi 2 analysis, a greater chance of persistent bladder/sphincter problem (P = 0.008), persistent severe motor deficit (P = 0.006), persistent pain (P = 0.025), and sexual dysfunction (P = 0.006) existed with delayed surgery. CONCLUSION: The data strongly support the management of cauda equina syndrome from lumbar disc herniation as a diagnostic and surgical emergency. PMID- 10703109 TI - Variation in bone mineral density of the sacrum in young adults and its significance for sacral fixation. AB - STUDY DESIGN: Bone mineral density variations throughout the sacrum were measured and correlated with sacral screw insertion torque. OBJECTIVE: To quantify bone mineral density variations within the S1 body and ala of young human specimens, especially along the pathways of sacral screws, and to examine the relation between sacral screw fixation and bone mineral density. SUMMARY OF BACKGROUND DATA: Vertebral bone quality is an essential factor in anterior or posterior screw fixation of the spine. Several studies have been conducted regarding bone mineral density variations in the cervical and thoracolumbar spine. However, such variations in bone mineral density in the sacrum have not been well documented. METHODS: The bone mineral density of 13 sacral specimens from young male cadavers (mean age, 31 years) was measured using highly accurate quantitative computed tomography. Variations in bone mineral density were measured in five transverse layers and seven vertical columns within the S1 body, and in four transverse layers and six vertical columns within the ala. The sacral screw insertion torque was measured (unicortical and bicortical), and the correlation with bone mineral density was calculated. RESULTS: The mean bone mineral density of the S1 body was 381.9 +/- 59 mg/cm3, which was 31.9% higher than that of the sacral ala (mean, 296.9 +/- 86 mg/cm3) (P < 0.05). Bone mineral density of the superior sacral endplate was higher than that of any other transverse layer. Columns near the lateral posterior and lateral anterior of the S1 body had the highest bone mineral density. In the ala, bone mineral density values of the internal columns (pedicle) were the highest. Screw insertion torque for bicortical purchase along the S1 pedicle correlated well with the bone mineral density of the S1 body (r = 0.67, P < 0.05). CONCLUSION: This study quantified the volumetric bone mineral density variations within the S1 body and ala, and a significant linear correlation between the screw insertion torque and bone mineral density was found. Optimal sacral screw insertion pathways were also outlined based on bone mineral density values. PMID- 10703110 TI - The interspinous ligament of the lumbar spine. Magnetic resonance images and their clinical significance. AB - STUDY DESIGN: A preliminary study of magnetic resonance features of the interspinous ligament in degenerative lumbar spine. OBJECTIVES: To classify the magnetic resonance imaging features of the interspinous ligaments in relation to the patient's age, disc degeneration, and radiographic instability. Magnetic resonance imaging also was correlated with the histologic findings of the interspinous ligaments. SUMMARY OF BACKGROUND DATA: As reported, rupture of the interspinous ligament frequently is found in the degenerative lumbar spine. However, little information is available in the literature on imaging assessment of the interspinous ligament in degenerative lumbar disorders. METHODS: In this study, 24 interspinous ligaments at L1-L2 or L2-L3 from 15 patients with nondegenerated discs were selected to represent normal magnetic resonance features of the interspinous ligament, and 38 patients with the mean age of 49 years underwent functional radiography and magnetic resonance imaging. The magnetic resonance features of the interspinous ligament were classified into five categories according to their signal intensities: Type 1A (low intensity on T1- and T2-weighted images without hypertrophy of the spinal process); Type 1B (same signal pattern as in Type 1A with hypertrophy of spinal process); Type 2 (low intensity on T1- and high intensity on T2-weighted images); Type 3 (high intensity on T1-weighted images); and Type 4 (others). Seven patients with variable patterns of the interspinous ligament were selected to undergo histologic examinations. RESULTS: Of the interspinous ligaments considered normal, 80% were classified as Type 1A. There were 14 Type 1A, 30 Type 1B, 19 Type 2, 16 Type 3, and 20 Type 4 ligaments. The mean age and disc degeneration grade of the patients with the Type 1B ligaments was significantly higher. Instability was found to be associated with Type 2 interspinous ligaments (7 of 19), whereas instability rarely was noted in Types 1A (1 of 14) and 1B (1 of 30) ligaments. The histologic examination revealed that chondrometaplasia and necrotization of fiber bundle predominated in Type 1B, proliferation of cells and vascular invasion in Type 2, fatty degeneration in Type 3 ligaments. CONCLUSIONS: The magnetic resonance imaging characteristics may be helpful in assessing normal or pathologic changes in the interspinous ligaments. PMID- 10703111 TI - A radiostereometric analysis of movements of the sacroiliac joints during the standing hip flexion test. AB - STUDY DESIGN: The standing hip flexion test was evaluated by using a radiostereometric analysis. OBJECTIVES: To evaluate whether the commonly used standing hip flexion test reflects movement in the sacroiliac joints, or whether the increased load of one sacroiliac joint also reduces the mobility of the other sacroiliac joint according to the theory of form and form closure in the sacroiliac joints. SUMMARY OF BACKGROUND DATA: The standing hip flexion test, used frequently to analyze sacroiliac joint mobility, is advocated as a test for study of normal or impaired motion in the sacroiliac joint. METHODS: In this study, 22 patients considered to have sacroiliac pain were analyzed with radiostereometric analysis when standing and when performing the standing hip flexion test on the right and left sides. RESULTS: Very small movements were registered in the sacroiliac joints. When provoking one side, the rotations were small on both sides. CONCLUSIONS: The small movements registered support the theory of form and force closure in the sacroiliac joints. The self-locking mechanism that goes into effect when the pelvis is loaded in a one-leg standing position probably obstructs the movements in the sacroiliac joints. Therefore, the standing hip flexion test cannot be recommended as a diagnostic tool for evaluating joint motion in the sacroiliac joints. PMID- 10703112 TI - Physical and psychosocial factors related to low back pain during a 24-year period. A nested case-control analysis. AB - STUDY DESIGN: A retrospective nested case-control study. OBJECTIVES: To identify occupational factors related to low back pain, and to study how interactions between psychosocial and physical factors, and between work-related and leisure related factors affect low back pain in women and men. SUMMARY OF BACKGROUND DATA: A cohort of 484 subjects drawn from the general population was examined in 1969 and 1993, with a focus on occupational working conditions and musculoskeletal disorders. METHODS: Information about the physical and psychosocial working conditions and low back pain during the period 1970 to 1993 was collected retrospectively. Odds ratios and confidence intervals were calculated for different potential risk factors. RESULTS: During the 24-year period, 46% of the subjects became patients with low back pain. Among women, heavy physical workload, sedentary work, smoking, and the combination of whole body vibrations and low influence over work conditions were associated with an excess risk of low back pain. Among men, excess risk for low back pain was seen in heavy physical workload, sedentary work, high perceived load outside work, and the combination of poor social relations and overtime. CONCLUSIONS: Factors at work were seen to be risk indicators for low back pain among both genders. Low influence over work conditions among women and poor social relations at work among men, in combination with other factors, seem to be of high relevance for the occurrence of low back pain. PMID- 10703113 TI - The use of rhBMP-2 in interbody fusion cages. Definitive evidence of osteoinduction in humans: a preliminary report. AB - STUDY DESIGN: A prospective randomized controlled human clinical pilot trial. OBJECTIVES: To determine the feasibility of using rhBMP-2/collagen as a substitute for autogenous bone graft inside interbody fusion cages to achieve arthrodesis in humans. SUMMARY OF BACKGROUND DATA: Preclinical studies have shown rhBMP-2 to be an effective substitute for autogenous bone graft, but there are no studies to date documenting such efficacy for human spine fusion. METHODS: Fourteen patients with single-level lumbar degenerative disc disease refractory to nonoperative management were randomized to receive lumbar interbody arthrodesis with a tapered cylindrical threaded fusion cage filled with rhBMP 2/collagen sponge or autogenous iliac crest bone. Patients were evaluated with radiographs, sagittally reformatted computed tomography scans, and Short Form-36 and Oswestry outcome questionnaires. RESULTS: All 11 patients who received rhBMP 2 were judged by three independent radiologists to have solid fusions (at 6, 12, and 24 months postimplantation), whereas only 2 of the 3 control patients, who received the standard treatment of autogenous iliac crest bone, were deemed to be fused. The Oswestry Disability Questionnaire scores of the rhBMP-2 group improved sooner (after 3 months) than those of the autograft group, with both groups demonstrating similar improvement at 6 months. Short Form 36 scores continued to improve up to 24 months. CONCLUSION: The arthrodesis was found to occur more reliably in patients treated with rhBMP-2-filled fusion cages than in controls treated with autogenous bone graft, although the sample size was limited. There were no adverse events related to the rhBMP-2 treatment. This study is one of the first to show consistent and unequivocal osteoinduction by a recombinant growth factor in-humans. PMID- 10703114 TI - Management of chronic discogenic low back pain with a thermal intradiscal catheter. A preliminary report. AB - STUDY DESIGN: A prospective nonrandomized clinical trial. OBJECTIVE: To determine the outcome in a group of patients with chronic, function-limiting low back pain who met the criteria for interbody fusion surgery, but were instead treated with an intradiscal thermal catheter (SpineCath, Oratec Interventions, Inc., Menlo Park, CA). SUMMARY OF BACKGROUND DATA: This study represents the first reported trial of treatment for chronic discogenic low back pain with a novel thermal intradiscal catheter. METHODS: The authors evaluated 25 consecutive patients. The minimum duration of nonoperative care with the authors was 6 months. The visual analog pain scores, sitting tolerance times, and SF-36 summary scores were tabulated. RESULTS: The mean follow-up period was 7 months, and the mean duration of symptoms 58.5 months. Of the 25 patients, 20 (80%) reported a reduction of at least 2 points in visual analog pain scores, and 18 (72%) reported an improvement in sitting tolerance as well as reduction or discontinuance of analgesic medication. Visual analog pain scores improved by a mean reduction of 3.74, a 51% change (P = 0.0001). On the SF-36 physical function subscale, 72% of the patients improved by a mean increase of 15 points (P = 0.001), a mean change of 38%, and by a mean increase of 14 points on the bodily pain subscale (P = 0.004), a mean change of 48%. CONCLUSIONS: A statistically significant improvement in functional outcome was obtained in patients with chronic discogenic low back pain treated thermally by the SpineCath. PMID- 10703115 TI - Spine update. Lumbar foraminal stenosis. AB - Lumbar foraminal stenosis is an important pathologic entity to identify in the patient being treated for radicular symptoms. This update reviews the anatomy, clinical presentation, neuroradiographic evaluation, and treatment of pathology located in the intervertebral foramen. Patients with significant leg pain refractory to conservative treatment and concordance between the demonstrated area of stenosis and radicular symptoms and signs are candidates for the decompressive procedures discussed. The role of arthrodesis and spinal instrumentation in the management of foraminal stenosis also is addressed. PMID- 10703116 TI - Jefferson fracture resulting in Collet-Sicard syndrome. AB - STUDY DESIGN: A case report and review of the literature. OBJECTIVE: To increase awareness of and add to the spectrum of injury that can result from Jefferson fractures, to suggest a possible mechanism of injury, and to give a brief review of pertinent facts regarding C1 burst fractures and the Collet-Sicard Syndrome. SUMMARY OF BACKGROUND DATA: To the author's knowledge, this is the first reported case of a Jefferson fracture resulting in Collet-Sicard Syndrome. It represents only the second reported case of cranial nerve palsy caused by Jefferson fracture. METHODS: A 56-year-old man sustained a C1 burst fracture in a rollover motor vehicle accident. Repeated neurologic examinations over the ensuing days revealed lesions of cranial nerves IX, X, XI, and XII on the left side. RESULTS: Two weeks of traction, 10 weeks in a halo vest, and 2 weeks in a cervical collar resulted in adequate fracture healing and almost complete resolution of the patient's neurologic symptoms. CONCLUSION: Although this is the first reported case of Collet-Sicard Syndrome caused by Jefferson fracture, the authors' review of the literature suggests that cranial nerve injuries may go unrecognized in some patients with C1 burst fractures. The importance of a thorough neurologic examination, including examination of the cranial nerves, in all cases of cervical spine injury cannot be overemphasized. PMID- 10703117 TI - Members. American Ophthalmological Society. PMID- 10703118 TI - Air bags and ocular injuries. AB - PURPOSE: This investigation retrospectively examined ocular injuries associated with air bag deployment to gain a better appreciation of potential risk factors in motor vehicle accidents. National statistics regarding the efficacy of air bags were reviewed. METHODS: Review of the literature from 1991 to 1998 identified 44 articles describing 97 patients with air-bag-induced ocular injuries. Variables extracted from each case were age, sex, height, position in the car, eye wear, vehicle impact speed, visual acuity, and specific ocular injuries. RESULTS: Corneal abrasions occurred in 49% of occupants, hyphemas in 43%, vitreous or retinal hemorrhages in 25%, and retinal tears or detachments in 15%. The globe was ruptured in 10 patients. Patients involved in higher-speed accidents (over 30 mph) sustained a greater percentage of vitreous or retinal hemorrhages and traumatic cataracts, while those at slower speeds were more prone to retinal tears or detachments. In a subset of 14 patients with serious ocular injuries, the impact speed of 11 patients was recorded at 30 mph or less. Slower speed may be a risk factor for some ocular injuries. Occupant height was not a significant factor. National statistics confirm that air bags reduce fatalities in motor vehicle accidents. However, children sitting in the front seat without a seat belt and infants in passenger-side rear-facing car seats are at risk for fatal injury. CONCLUSION: Air bags combined with seat belts are an effective means of reducing injury and death in adults during motor vehicle accidents. However, this study has documented a wide variety of ocular injuries associated with air bag deployment. It is hoped that researchers can develop modifications that continue to save lives while minimizing additional harm. PMID- 10703119 TI - Mechanisms of orbital floor fractures: a clinical, experimental, and theoretical study. AB - PURPOSE: The purpose of this study was to investigate the two accepted mechanisms of the orbital blow-out fracture (the hydraulic and the buckling theories) from a clinical, experimental, and theoretical standpoint. METHODS: Clinical cases in which blow-out fractures resulted from both a pure hydraulic mechanism and a pure buckling mechanism are presented. Twenty-one intact orbital floors were obtained from human cadavers. A metal rod was dropped, experimentally, onto each specimen until a fracture was produced, and the energy required in each instance was calculated. A biomathematical model of the human bony orbit, depicted as a thin walled truncated conical shell, was devised. Two previously published (by the National Aeronautics Space Administration) theoretical structural engineering formulas for the fracture of thin-walled truncated conical shells were used to predict the energy required to fracture the bone of the orbital floor via the hydraulic and buckling mechanisms. RESULTS: Experimentally, the mean energy required to fracture the bone of the human cadaver orbital floor directly was 78 millijoules (mj) (range, 29-127 mj). Using the engineering formula for the hydraulic theory, the predicted theoretical energy is 71 mj (range, 38-120 mj); for the buckling theory, the predicted theoretical energy is 68 mj (range, 40-106 mj). CONCLUSION: Through this study, we have experimentally determined the amount of energy required to fracture the bone of the human orbital floor directly and have provided support for each mechanism of the orbital blow-out fracture from a clinical and theoretical basis. PMID- 10703120 TI - Ocular safety of Viagra, (sildenafil citrate). AB - To date, sildenafil citrate (Viagra) gives every evidence of being a safe drug for the eye despite a series of expressed concerns. A review of how its ocular safety profile has been identified offers insights into the strengths and weaknesses of present systems and resources for judging the ocular safety of Viagra or, for that matter, of any new drug. Such insights include: The great value of careful, informed assessment of preclinical information gleaned from laboratory experiments. By and large, such assessments point the way toward appropriate clinical evaluation. For Viagra, early in its development it was noted that besides exerting a major inhibitory effect on the intended target, the vascular-associated enzyme phosphodiesterase 5 (PDE5), the drug also exerts a lesser but definite inhibitory effect on the closely related PDE6, located in the retina. For this reason, preclinical evaluation of the drug included electroretinography plus postmortem histology. In addition, an extended eye examination was incorporated into clinical protocols. The often chaotic but invaluable information stream that becomes available once marketing approval has been gained and large populations begin to use a drug. False alarms, misattribution, and erroneous information are the order of the day. Nevertheless, as information accumulates, patterns of response clarify and the true nature of special susceptibility for subpopulations, if any, becomes apparent. A role for the astute clinician remains: Subtle changes or unusual risks for subpopulations can be missed entirely for long periods of time. A manifest need for improvement in evaluation of postmarketing side-effects. This need has led to the establishment of a new discipline: pharmacoepidemiology. In ophthalmology, the National Registry of Drug Induced Ocular Side-Effects maintains a constant and invaluable surveillance. Examples are supplied to illustrate each of these major points: Our presentation will include data gleaned from clinical trials plus postmarketing information on the incidence, duration, and type of color vision defects observed at different doses of Viagra. PMID- 10703121 TI - Ophthalmology's future in the next decade: a historical and comparative perspective. AB - PURPOSE: To gain a historical and comparative perspective about the future of ophthalmology within the profession of medicine. METHODS: A literature search is made of disciplines other than medicine (history, sociology, philosophy, economics, and ethics) in order to assess factors responsible for survival and healthiness of a profession. The "learned" professions (medicine, law, and theology) are assessed. Other "professional" careers valued by society (sports and classical music) are reviewed. RESULTS: From the perspective of other disciplines, the future of ophthalmology is seen as vulnerable and fragile. Survival of professions, be they classically or economically defined, is linked to societal needs, a profession's unique commitment and ability to provide services to society, and the profession's maintenance of knowledge as well as skill-based services. Historical evidence has shown erosion of a profession's power consequent to capitalist influences, government influences, access of skills by less trained individuals, and elitist posturing by a profession. Comparative evidence has shown societal acceptance of an escalation of salaries for designated superstars, increasing roles and influence of managerial personnel, and trivialization of values other than economic ones. CONCLUSION: Attention to historical and comparative trends by individual ophthalmologists as well as associations representing ophthalmologists is mandatory if ophthalmology as we know it is to survive within the profession of medicine. PMID- 10703122 TI - Indeterminate melanocytic proliferations of the conjunctiva. AB - PURPOSE: The purpose of this study is to test the hypothesis that a subset of conjunctival melanocytic proliferations exists that cannot be reproducibly classified as benign, malignant, or indeterminate. METHODS: Three groups of excisional biopsy specimens of conjunctival melanocytic proliferations were evaluated by 5 ophthalmic pathologists. These groups included lesions that were considered by the authors to represent benign (Group 1, n = 5), malignant (Group 2, n = 5) and indeterminate melanocytic proliferations (Group 3, n = 5). The panel classified the same sections in all 3 groups in a randomized, masked fashion, first without and then with a clinical history of patient age, sex and race. The kappa statistic (k) was used to quantify the degree of agreement among observers. RESULTS: There was strong concordance among the panel for both Group 1 (benign, k = 0.76) and Group 2 (malignant, k = 0.70) melanocytic proliferations. There was no concordance of the panel for Group 3 (indeterminate) lesions (k = 0.045). The concordance for Groups 1 and 2 and lack of concordance for Group 3 lesions were independent of knowledge of clinical history of age, sex, and race. CONCLUSIONS: A subset of melanocytic proliferations of the conjunctiva exists that cannot be reproducibly classified by pathologists as benign, malignant, or indeterminate. PMID- 10703123 TI - Combined nevi of the conjunctiva. AB - PURPOSE: To report the clinical and histologic features of combined nevi of the conjunctiva, a type of nevus that is not uncommon in the skin but has rarely been reported in the conjunctiva. METHODS: Conjunctival nevi and melanomas from the files of the University of California, San Francisco, eye pathology laboratory were reviewed from 1984 to 1999 for the presence of features of both standard nevocytic nevi and blue nevi. Clinical histories and, when available, clinical photographs were obtained. RESULTS: Thirty-one combined nevi were discovered during the 15-year period between 1984 and 1999. One case before 1984 had been incorrectly diagnosed as a junctional nevus. The dendritic and spindle-shaped blue nevus cells had been overlooked because they were not recognized as distinct from the standard nevocytic nevus cells. The recognition of a blue as well as a brown color, a deep as well as a superficial component in the lesion, or a history of pigmentation since birth may help to establish the correct clinical diagnosis and prevent an unnecessarily deep surgical resection. Although growth of the lesion or "satellites" in some patients may favor a clinical diagnosis of melanoma, none of the lesions in this series were malignant. CONCLUSION: Despite a paucity of reports of combined nevi of the conjunctiva in the medical literature, this type of nevus--a combination of a nevocytic and a blue nevus--is common and has been overlooked in the past. PMID- 10703124 TI - Acute hydrops in the corneal ectasias: associated factors and outcomes. AB - PURPOSE: To identify factors associated with the development of hydrops and affecting its clinical outcome. METHODS: Chart review of all patients with acute hydrops seen by a referral cornea service during a 2.5-year period between June 1996 and December 1998. RESULTS: Twenty-one patients (22 eyes) with acute hydrops were seen. Nineteen patients had keratoconus, 2 had pellucid marginal degeneration, and 1 had keratoglobus. Twenty-one of 22 (95%) eyes had seasonal allergies and 20 of 22 (91%) eyes had allergy-associated eye-rubbing behavior. Six of 22 (27%) had a diagnosis of Down's syndrome. Six patients were able to identify a traumatic inciting event: vigorous eye rubbing in 4 and traumatic contact lens insertion in 2. The affected area ranged from 7% to 100% of the corneal surface area and was related to disease duration and final visual acuity. Proximity of the area of edema to the corneal limbus ranged from 0 to 2.3 mm and was also related to prognosis. Three serious complications were observed: a leak, an infectious keratitis, and an infectious keratitis and coincidental neovascular glaucoma. Various medical therapies did not differ significantly in their effect on outcome, and ultimately 4 (18%) of 22 patients underwent penetrating keratoplasty. Best-corrected visual acuity was equal to or better than prehydrops visual acuity in 5 of the 6 patients in whom prehydrops visual acuity was known, without corneal transplantation. CONCLUSIONS: Allergy and eye-rubbing appear to be important risk factors in the development of hydrops. Visual results are acceptable in some patients without surgery. Close observation allows for the early detection and treatment of complications such as perforation and infection. PMID- 10703125 TI - Giant papillary conjunctivitis in frequent-replacement contact lens wearers: a retrospective study. AB - PURPOSE: A retrospective study was done of 47 patients who wore frequent replacement contact lenses on a daily basis and replaced them every 1 day to 12 weeks. The incidence of giant papillary conjunctivitis (GPC) was determined, and potential risk factors that may predispose frequent-replacement contact lens wearers to develop GPC were assessed. METHODS: The records of patients who were fitted with frequent-replacement contact lenses with no prior contact lens experience (September 1993 to February 1997) were reviewed. RESULTS: Forty-seven of 260 patients met the requirement for inclusion in this study. Ten (21.27%) of the patients developed GPC. The incidence varied according to how often the contact lenses were replaced. Incidence was 36% in patients who replaced their lenses at 4 weeks or longer and 4.5% in patients who replaced their lenses at less than 4 weeks. Lenses were coated more often in patients who replaced their lenses at 4 weeks or longer (pi = .23). A significantly greater number of patients in the GPC group incorporated enzyme into their contact lens care system (pi = .0004). A history of allergy was present, significantly more often in patients who developed GPC (pi = .012). There was no significant difference between the groups for age, sex, average daily wearing time, Food and Drug Administration classification of contact lens material, time in contact lenses from fitting to diagnosis or last follow-up period, or the parameters and fitting characteristics of the contact lenses. CONCLUSION: The frequency of contact lens replacement appears to be an important variable in development of GPC. Although frequent-replacement contact lenses do not eliminate GPC, patients on a 1-day to 3-week replacement cycle had a significantly lower risk of developing GPC than patients who replaced their lenses at longer intervals. Coating was present less often on lenses replaced every 1 day to 3 weeks. In patients who are at high risk for GPC, replacing lenses at intervals of 1 day to 2 weeks appears to offer a better strategy in avoiding GPC than incorporating enzymatic cleaning into their care system. PMID- 10703126 TI - Keratocyte density in vivo after photorefractive keratectomy in humans. AB - PURPOSE: To determine changes in keratocyte density in central human corneas in vivo after photorefractive keratectomy (PRK). METHODS: Fifteen patients (25 eyes) received excimer PRK (VISX Star) with epithelial removal by laser-scrape (43 microns ablation followed by manual scrape) to correct myopia between -1.5 D and 7.25 D. Corneas were examined by using confocal microscopy in vivo before PRK and at 1 day, 5 days, 1 month, and 3 months after PRK. A custom automated image processing algorithm identified bright objects (keratocytes) against a dark background and estimated keratocyte density by using the number and size of the objects. Cell density was quantified in anteroposterior stromal regions after PRK and compared to cell density in corresponding pre-PRK regions. RESULTS: One day after PRK, keratocyte density increased 9% in the anterior third of the stroma (pi = .003), was unchanged in the middle third of the stroma (pi = .481), and decreased 6% in the posterior third of the stroma (pi = .038). Keratocyte density remained elevated in the anterior stroma to 3 months after PRK; at this time, there was a 13% increase in keratocyte density throughout the full-thickness stroma (pi < .001). CONCLUSIONS: Keratocyte density was increased in the anterior stroma immediately after PRK in humans. Three months later, keratocyte density was increased in all anteroposterior stromal regions, suggesting that PRK affects keratocytes throughout the entire central cornea. PMID- 10703127 TI - Six-month results of hyperopic and astigmatic LASIK in eyes with primary and secondary hyperopia. AB - PURPOSE: To assess the safety and efficacy of laser in situ keratomileusis (LASIK) for hyperopia and hyperopic astigmatism and develop a LASIK nomogram for primary hyperopia or hyperopia secondary to myopic refractive surgery using the VISX STAR S2. METHODS: Prospective evaluation of LASIK in 46 primary eyes and 29 secondary eyes with fogged manifest sphere from +0.5 diopters (D) to +6.0 D and cylinder from 0 to +5.0 D. RESULTS: Mean manifest spherical equivalent (SE) in patients with primary hyperopia was +2.50 D +/- 0.93 preoperatively and +0.70 D +/- 1.19 at 6 months. At 6 months, 79% of primary hyperopes had uncorrected visual acuity (UCVA) of 20/40 or better; 63% were within +/- 1 D of emmetropia. One primary hyperope lost 2 lines of best spectacle-corrected vision (BCVA) at 1 month. Complications included transient epithelial defect (6.5%), epithelial cells in the interface (4.3%), diffuse lamellar keratitis (4.3%), haze (2.2%), and mild irregular astigmatism (2.2%). In those with secondary hyperopia, mean manifest SE was +1.70 D +/- 0.82 preoperatively and -0.27 D +/- 0.95 at 6 months. At 6 months, 83% of secondary hyperopes had UCVA of 20/40 or better; 74% were within +/- 1 D of emmetropia. No secondary hyperope lost > or = 2 lines of BCVA. Complications included intraoperative bleeding (3.4%), intraoperative epithelial defect (3.4%), transient interface debris (3.4%), significant dry eye (3.4%), blood in interface (3.4%), irregular astigmatism (6.9%), slight decentration (6.9%), trace haze (6.9%), mild epithelial ingrowth not requiring removal (3.4%), or corneal irregularity (3.4%). CONCLUSION: These early data suggest that LASIK for hyperopia from +0.5 to +6 D and astigmatism from 0 to +5 D using the VISX STAR S2 benefits from a nomogram adjusted for preoperative refraction, age, and prior refractive surgery and is safe and effective. Patients with secondary hyperopia achieved more correction than those with primary hyperopia, although the accuracy and predictability of LASIK in both groups has improved with the nomogram adjustments. PMID- 10703128 TI - Two clinical trials of an intraocular steroid delivery system for cataract surgery. AB - PURPOSE: To determine the safety and efficacy of an intraocular dexamethasone drug delivery system (Surodex) in the treatment of inflammation following cataract surgery. METHODS: Surodex is a biodegradable polymer that releases dexamethasone for 7 to 10 days after placement in the anterior segment. Study 1 was a prospective, randomized, double-masked Phase II clinical trial of 90 cataract surgical patients that compared treatment with Surodex to treatment with a placebo drug delivery system and to no anti-inflammatory drug treatment at all. Study 2 was a separate prospective, randomized, double-masked study of 60 cataract surgical patients that compared treatment with Surodex to topical dexamethasone (eye drop) therapy. RESULTS: In the first study, Surodex was superior to placebo in suppressing postsurgical inflammation throughout the 60 day postoperative period, as judged by masked-evaluator, slit-lamp grading of cell and flare. The differences were statistically significant from postoperative day 3 through postoperative week 3. The majority of Surodex patients did not require topical steroid by 2 weeks after surgery (93%) or by 2 months after surgery (88%). In the second study, Kowa laser flare meter readings were lower in Surodex patients throughout the 90-day postoperative period. The results were statistically significant at 4, 8, and 15 days following surgery. There were no significant adverse complications of Surodex in either study. CONCLUSION: Surodex was safe and effective in suppressing postcataract surgery inflammation and appears to be a promising alternative to topical steroids. PMID- 10703129 TI - Visual outcomes after anterior vitrectomy: comparison of ECCE and phacoemulsification. AB - PURPOSE: To determine whether vitrectomy instrumentation improved outcomes when vitreous loss occurred during either extracapsular cataract extraction (ECCE) or phacoemulsification (PE) with posterior chamber lens implantation (PCIOL). METHODS: A consecutive series of ECCE + PCIOL (group 1: 1985-1989) and PE + PCIOL (group 2: 1993-1997) surgeries by a single surgeon was reviewed retrospectively. RESULTS: In group 1, 14 of 647 patients (2.2%) and in group 2, 9 of 665 patients (1.4%) experienced vitreous loss. In group 1, final visual acuity averaged 20/83; in group 2, 20/25 (P = .005). Average follow-up was 5.7 years (group 1) and 3.2 years (group 2). Uveitis, glaucoma, corneal problems, and retinal problems were assessed. CONCLUSIONS: Anterior vitrectomy reduced complications from vitreous loss. Fewer vitreous losses occurred with PE than ECCE. Patients with vitreous loss after PE attained better vision. PMID- 10703131 TI - Experience with early internal obstruction of the filtration site following filtration surgery. PMID- 10703132 TI - Achieving success with the silicone expander for overacting superior obliques. AB - PURPOSE: To report the results of and complications with silicone expander surgery for the overacting superior oblique. METHODS: A total of 26 patients with bilateral overaction of the superior oblique and A-pattern strabismus and 5 patients with unilateral overacting superior oblique secondary to inferior oblique palsy were treated with a 7 mm silicone expander. Care was taken not to enter the sub-Tenon's space. RESULTS: The group that underwent bilateral superior oblique surgery had an average preoperative pattern of 37.42 diopters (D) and an average correction of 35.37 D. Three patients had a severe unilateral postoperative inflammatory incident that was successfully treated with oral and topical corticosteroids. One of these patient developed Brown's syndrome. Another patient, who had no postoperative inflammatory incident, also developed Brown's syndrome. In these 4 patients, the sub-Tenon's space was inadvertently entered during surgery. CONCLUSION: The silicone expander surgery has a very high success rate in treating the A-pattern associated with the bilateral overacting superior oblique. This procedure also works well for the unilateral superior oblique that overacts owing to an inferior oblique palsy. No cyclotorsion symptoms occurred after this surgery. However, 4 patients had complications because the sub-Tenon's space was exposed during surgery. With this procedure, there is a learning curve to obtain the skill not to enter the sub-Tenon's space. PMID- 10703130 TI - Combined exfoliation and pigment dispersion: an overlap syndrome. AB - PURPOSE: To describe a series of patients with combined pigment dispersion syndrome (PDS) and exfoliation syndrome (XFS) and to introduce a concept, the overlap syndrome, to aid in assessing multiple risk factors for glaucomatous damage. METHODS: A retrospective review of the records of all patients on our database who carried a diagnosis of both PDS/pigmentary glaucoma (PG) and XFS/exfoliative glaucoma (XFG). RESULTS: We identified 26 patients as having both XFS/XFG and PDS/PG. The average age was 64.3 +/- 9.8 years and 19 of 26 were men. All patients had bilateral PDS/PG. Bilateral XFS/XFG was present in 9 of 26 patients, and of the 17 patients with unilateral involvement, the left eye was affected in 13. CONCLUSION: Both XFS and PDS are common. Middle-aged patients with known PDS/PG should be suspected of having the onset of XFS if 1 eye escapes intraocular pressure control. Patients presenting with unilateral XFG may also have signs of PDS/PG, often remitted. We define an overlap syndrome as the appearance of a new co-morbidity for glaucomatous damage in a patient with a pre existing risk factor, which then changes the course, and prognosis of the disease. This concept should be particularly useful in dealing with secondary and normal-tension glaucoma. PMID- 10703133 TI - A value analysis model applied to the management of amblyopia. AB - PURPOSE: To assess the value of amblyopia-related services by utilizing a health value model (HVM). Cost and quality criteria are evaluated in accordance with the interests of patients, physicians, and purchasers. METHODS: We applied an HVM to a hypothetical statistical ("median") child with amblyopia whose visual acuity is 20/80 and to a group of children with amblyopia who are managed by our practice. We applied the model to calculate the value of these services by evaluating the responses of patients and physicians and relating these responses to clinical outcomes. RESULTS: The consensus value of care for the hypothetical median child was calculated to be 0.406 (of 1.000). For those children managed in our practice, the calculated value is 0.682. Clinically, 79% achieved 20/40 or better visual acuity, and the mean final visual acuity was 0.2 logMAR (20/32). Value appraisals revealed significant concerns about the financial aspects of amblyopia related services, particularly among physicians. Patients rated services more positively than did physicians. CONCLUSIONS: Amblyopia care is difficult, sustained, and important work that requires substantial sensitivity to and support of children and families. Compliance and early detection are essential to success. The value of amblyopia services is rated significantly higher by patients than by physicians. Relative to the measured value, amblyopia care is undercompensated. The HVM is useful to appraise clinical service delivery and its variation. The costs of failure and the benefits of success are high; high-value amblyopia care yields substantial dividends and should be commensurately compensated in the marketplace. PMID- 10703135 TI - Evidence-based medicine regarding the prevention of retinal detachment. AB - PURPOSE: To assess the quality of information in the literature regarding the prevention of retinal detachment in an effort to establish appropriate practice guidelines. METHODS: A panel of vitreoretinal experts performed a literature review of all publications in the English language for articles about prevention of retinal detachment. These article were then used to prepare recommendations for patient care in an American Academy of Ophthalmology "Preferred Practice Pattern" (PPP). Each recommendation was rated according to its importance in the care process and the strength of evidence supporting the given recommendation. RESULTS: Most recommendations were given a rating of "A" (most important to patient care). Only a single publication was graded as Level I (providing strong evidence in support of a recommendation), and this was not a prospective trial. Of the few publications rated as Level II (substantial evidence), most were studies documenting a lack of treatment benefit. Because of an absence of Level I and Level II studies in the literature, Level III (consensus of expert opinion) was the basis for most recommendations in the PPP. CONCLUSION: The current literature regarding prevention of retinal detachment does not provide sufficient information to strongly support prophylactic treatment of lesions other than symptomatic flap tears. Prospective randomized trials of prophylactic therapy are indicated. Eyes highly predisposed to retinal detachment should be considered for such studies. PMID- 10703134 TI - The therapy of amblyopia: an analysis comparing the results of amblyopia therapy utilizing two pooled data sets. AB - CONTEXT: We previously presented the results of an original pooled data set of 961 amblyopic patients who underwent patching therapy for amblyopia from 1965 to 1994 (study group 1). Three types of amblyopia were considered: anisometropic, anisometropic-strabismic, and strabismic. Analysis of this group's success was related to the age at which therapy was initiated, the type of amblyopia, and the depth of visual loss before treatment was begun. The purpose of the current study is to test the validity of these findings on a second group of 961 amblyopes employing the data set used by Woodruff and associates in their publications (study group 2). These 2 data sets, after adjustment to conform to the definitions of age, amblyopia, anisometropia, and similar items utilized in common between the 2 study groups, will be compared for the risk factors predictive of successful occlusion therapy. OUTCOME: As in the previous study, the success of occlusion therapy is defined as a visual acuity of 20/40 or better at the end of treatment. RESULTS: Success by the 20/40 criteria was achieved in 73.7% in study group 1 and in 59.9% in study group 2. By category, the rate of success in study group 1 was 77.2% in strabismic amblyopia, 67.2% in anisometropic-strabismic amblyopia, and 66.0% in anisometropic amblyopia. In study group 2, success was 61.2% in strabismic amblyopia, 51.2% in anisometropic strabismic amblyopia, and 63.0% in anisometropic amblyopia. Study group 1 univariate analysis related success in each group to the age at which therapy was initiated, the type of amblyopia, and the depth of visual loss before treatment in each group. In study group 2, univariate analysis related success of occlusion therapy to age and the depth of visual loss before treatment. Type of amblyopia was not related to outcome success in this group. When the 2 data sets were pooled, the risk factors for success were age and depth of visual loss at onset of treatment. CONCLUSIONS: Factors that appeared closely related to a successful outcome of patching therapy were patient age and depth of visual loss before treatment. These conclusions further support the value of early detection and screening for amblyopia, its prevention, where possible, and its adequate and vigorous treatment when it is detected and diagnosed. PMID- 10703136 TI - Transpupillary thermotherapy as primary treatment for small choroidal melanomas. AB - PURPOSE: To report short-term follow-up of eyes containing small choroidal melanomas that were treated with transpupillary thermotherapy (TTT). METHODS: Twenty eyes with suspected small choroidal melanomas were treated with TTT using infrared light delivered from the diode laser. RESULTS: The age of patients ranged from 26 to 82 years. In 14, there was documented growth of the melanoma prior to TTT. Tumor thicknesses ranged from less than 1 to 3.2 mm. Seven tumors were treated more than once. Follow-up ranged from 6 months to more than 3 years. Following treatment, tumor thicknesses decreased in all cases, usually within 2 months. Progressive atrophy of tumor mass and loss of pigmentation within the tumor continued beyond 1 year of follow-up in some eyes. Complications included field defects, vascular changes, and macular abnormalities. CONCLUSIONS: Transpupillary thermotherapy of small choroidal melanomas is usually followed by early tumor shrinkage but is complicated by dense scotomas, nerve fiber bundle defects, and occasionally macular abnormalities. Short-term follow-up suggests that TTT may arrest growth of selected small melanomas. PMID- 10703137 TI - Retinopathy progression and visual outcomes after phacoemulsification in patients with diabetes mellitus. AB - PURPOSE: To determine the rate of progression of diabetic retinopathy following phacoemulsification surgery and to determine if surgeon experience and/or surgical duration adversely affect visual outcome. METHODS: A retrospective review of 150 eyes of 119 diabetic patients who underwent phacoemulsification surgery over a 5-year period was performed. Data collected included patient age, sex, type and duration of diabetes, diabetic control, associated systemic health factors, preoperative visual acuity and retinopathy grade, duration of surgery, intraoperative complications, and postoperative course. The effect of these factors on visual outcome and rate of retinopathy progression was studied. Resident and private cases were compared. RESULTS: The visual acuity improved by two or more lines in 117 eyes (78%). Ninety-three eyes (62%) had a final visual acuity greater than or equal to 20/40. Retinopathy progression was seen in 37 eyes (25%) followed up for 6 to 10 months. Preoperative nonproliferative diabetic retinopathy, prolonged surgical duration, and limited surgical experience were statistically associated with retinopathy progression. CONCLUSIONS: The visual results and rate of retinopathy progression after phacoemulsification surgery in our series do not appear to differ significantly from those reported using other techniques. Nonproliferative diabetic retinopathy, longer surgical duration, and surgical inexperience resulted in an increased rate of retinopathy progression. PMID- 10703138 TI - Juvenile retinoschisis: a model for molecular diagnostic testing of X-linked ophthalmic disease. AB - BACKGROUND AND PURPOSE: X-linked juvenile retinoschisis (RS) provides a starting point to define clinical paradigms and understand the limitations of diagnostic molecular testing. The RS phenotype is specific, but the broad severity range is clinically confusing. Molecular diagnostic testing obviates unnecessary examinations for boys at-risk and identifies carrier females who otherwise show no clinical signs. METHODS: The XLRS1 gene has 6 exons of 26-196 base-pair size. Each exon is amplified by a single polymerase chain reaction and then sequenced, starting with exons 4 through 6, which contain mutation "hot spots." RESULTS: The 6 XLRS1 exons are sequenced serially. If alterations are found, they are compared with mutations in our > 120 XLRS families and with the > 300 mutations reported worldwide. Point mutations, small deletions, or rearrangements are identified in nearly 90% of males with a clinical diagnosis of RS. XLRS1 has very few sequence polymorphisms. Carrier-state testing produces 1 of 3 results: (1) positive, in which the woman has the same mutation as an affected male relative or known in other RS families; (2) negative, in which she lacks the mutation of her affected male relative; and (3) uninformative, in which no known mutation is identified or no information exists about the familial mutation. CONCLUSIONS: Molecular RS screening is an effective diagnostic tool that complements the clinician's skills for early detection of at-risk males. Useful outcomes of carrier testing depend on several factors: (1) a male relative with a clear clinical diagnosis; (2) a well-defined inheritance pattern; (3) high disease penetrance; (4) size and organization of the gene; and (5) the types of disease-associated mutations. Ethical questions include molecular diagnostic testing of young at-risk females before the age of consent, the impact of this information on the emotional health of the patient and family, and issues of employability and insurance coverage. PMID- 10703139 TI - Vision and quality-of-life. AB - OBJECTIVE: To determine the relationship of visual acuity loss to quality of life. DESIGN: Three hundred twenty-five patients with visual loss to a minimum of 20/40 or greater in at least 1 eye were interviewed in a standardized fashion using a modified VF-14, questionnaire. Utility values were also obtained using both the time trade-off and standard gamble methods of utility assessment. MAIN OUTCOME MEASURES: Best-corrected visual acuity was correlated with the visual function score on the modified VF-14 questionnaire, as well as with utility values obtained using both the time trade-off and standard gamble methods. RESULTS: Decreasing levels of vision in the eye with better acuity correlated directly with decreasing visual function scores on the modified VF-14 questionnaire, as did decreasing utility values using the time trade-off method of utility evaluation. The standard gamble method of utility evaluation was not as directly correlated with vision as the time trade-off method. Age, level of education, gender, race, length of time of visual loss, and the number of associated systemic comorbidities did not significantly affect the time trade-off utility values associated with visual loss in the better eye. The level of reduced vision in the better eye, rather than the specific disease process causing reduced vision, was related to mean utility values. The average person with 20/40 vision in the better seeing eye was willing to trade 2 of every 10 years of life in return for perfect vision (utility value of 0.8), while the average person with counting fingers vision in the better eye was willing to trade approximately 5 of every 10 remaining years of life (utility value of 0.52) in return for perfect vision. CONCLUSIONS: The time trade-off method of utility evaluation appears to be an effective method for assessing quality of life associated with visual loss. Time trade-off utility values decrease in direct conjunction with decreasing vision in the better-seeing eye. Unlike the modified VF-14 test and its counterparts, utility values allow the quality of life associated with visual loss to be more readily compared to the quality of life associated with other health (disease) states. This information can be employed for cost-effective analyses that objectively compare evidence-based medicine, patient-based preferences and sound econometric principles across all specialties in health care. PMID- 10703140 TI - Collagen type I and III synthesis by Tenon's capsule fibroblasts in culture: individual patient characteristics and response to mitomycin C, 5-fluorouracil, and ascorbic acid. AB - PURPOSE: This study was performed to better understand the differences between patients in specific components of wound healing as it may pertain to glaucoma filtration surgery, including the use of antimetabolites. METHODS: Human Tenon's capsule fibroblasts were obtained at the time of glaucoma filtering surgery and established in individual cell cultures from 35 glaucoma patients. The dose response to 5-fluorouracil (5FU) and mitomycin C (MMC) was determined. The individual cell lines were exposed to the antimetabolites and ascorbic acid with measurement of collagen type I and III production by an ELISA-type dot blot assay. These results were then statistically compared to the individual patient characteristics including age, race, previous surgery and medications, and type of glaucoma. RESULTS: 5-FU had little effect on collagen type I and III production or protein synthesis. MMC had an inhibitory effect on collagen secretion and total protein synthesis with increasing concentration. Photomicrographs of the cells after each treatment condition revealed characteristic morphologic changes when compared to controls. There was a large range of collagen type I and III production with correlation between the amounts of each collagen type secreted in response to the antimetabolites. However, there was no correlation with accepted risk factors for filtration failure. CONCLUSION: These antimetabolites act similarly on different cell lines in a nonspecific manner. The results suggest that the increased risk of filtration failure due to age, race, diagnosis, and previous conjunctival surgery is not due to differences in secretion of collagen types I and III by Tenon's capsule fibroblasts. PMID- 10703141 TI - A 12-year ophthalmologic experience with the shaken baby syndrome at a regional children's hospital. AB - PURPOSE: To examine the ophthalmologic experience with the shaken baby syndrome (SBS) at one medical center, including clinical findings, autopsy findings, and the visual outcome of survivors. METHODS: One hundred sixteen patients admitted from 1987 to 1998 for subdural hematomas of the brain secondary to abuse were included. RESULTS: Retinal hemorrhages were detected in 84% of the children, but this important finding had been missed often by nonophthalmologists. Poor visual response, poor pupillary response, and retinal hemorrhage correlated strongly with demise of the child. One child who died had pigmented retinal scars from previous abuse, a condition not previously observed histopathologically. The clinical and autopsy findings varied somewhat, probably because of the differing conditions for examination. No correlation could be made between computerized tomography scans done during life and the subdural hemorrhage of the optic nerve found on autopsy. Half of the surviving patients were known to have good vision. One fourth of the patients had poor vision, largely due to cerebral visual impairment from bilateral injury posterior to the optic chiasm. Severe neurologic impairment correlated highly with loss of vision. CONCLUSION: This series provides information on the frequency of eye findings in SBS patients. No fundus finding is pathognomonic for SBS. When retinal hemorrhages are found in young children, the likelihood that abuse occurred is very high. The difficulty that nonophthalmologists have in detecting retinal hemorrhage may be an important limiting factor in finding these children so they may be protected from further abuse. PMID- 10703142 TI - Scar remodeling after strabismus surgery. AB - PURPOSE: Patients with overcorrected strabismus (and several patients with undercorrection after extraocular muscle resection) underwent exploration of previously operated muscles, with the intention of advancing their tendons to prevent the need for surgery on additional muscles. Unexpectedly, it was found that, in many cases, an elongated scar segment of variable length was interposed between the muscle and its insertion site on the sclera. Laboratory investigations were carried out to elucidate the underlying mechanism(s) and to create an animal model of the disorder. METHODS: Lengthened scars were repaired on 198 muscles during 134 procedures performed on 123 patients. The scars consisted of amorphous connective tissue interposed between the globe and normal tendon. Repair was accomplished by excision of the scar and reattachment of the muscle to sclera, using absorbable sutures in 64 cases and nonabsorbable sutures in 70 cases. Histopathologic examination was performed on 82 clinical specimens, and tissue culture studies were performed on 7 specimens. To develop an animal model, 10 New Zealand white rabbits underwent bilateral superior rectus resection. Half of the eyes received sub-Tenon's injections of collagenase over the operative site during weeks 2, 3, 5, and 6 postoperatively; the other half received saline solution injections on the same schedule. At 10 weeks, half the sites were studied histologically, and the other half underwent collagen creep analysis. In a second study, the use of absorbable versus nonabsorbable sutures was compared in the rabbit model. RESULTS: In the clinical cases, the mean length of the elongated scar segments was 4.2 mm. A total of 105 of the 134 repair procedures were judged successful. Thirty-one procedures resulted in recurrence of the original overcorrection; 7 of these had documented restretches. Factors that distinguished patients with stretched scars from patients with classic slipped muscles included minimal or no limitation of versions, less separation of the tendons from sclera, and thicker appearance of the scar segments. The use of nonabsorbable sutures in the repair procedure reduced the recurrence rate. Histologic examination of the clinical stretched scar specimens showed dense connective tissue that was less well organized compared with normal tendon. In the tissue culture studies, cells cultured from the stretched scar specimens grew rapidly and were irregularly shaped. A high-molecular-weight protein was identified in the culture medium. By contrast, cells cultured from normal tendon (controls) grew more slowly and regularly, stopped growing at 4 days, and produced less total protein than cultured stretched scar specimens. In the animal model studies, the collagenase-treated sites showed elongated scars with increased collagen between the muscle and the sclera, as well as increased collagen creep rates, compared with the saline-treated controls. The use of nonabsorbable sutures in collagenase-treated animal model surgery sites was associated with shorter, thicker scars compared with similar sites sutured with absorbable sutures. CONCLUSIONS: A lengthened or stretched, remodeled scar between an operated muscle tendon and sclera is a common occurrence and is a factor contributing to the variability of outcome after strabismus repair, even years later. This abnormality may be revealed by careful exploration of previously operated muscles. Definitive repair requires firm reattachment of tendon to sclera with nonabsorbable suture support. PMID- 10703143 TI - Childhood blindness and visual loss: an assessment at two institutions including a "new" cause. AB - PURPOSE: This study was initiated to investigate the causes of childhood blindness and visual impairment in the United States. We also sought a particular etiology--congenital lymphocytic choriomeningitis virus (LCMV)--which has been considered exceedingly rare, in a fixed target population of children, the severely mentally retarded. METHODS: We undertook a library-based study of the world literature to shed light on the causes of childhood blindness internationally and to put our data in context. We prospectively examined all consented children (159) at 2 institutions in the United States to determine their ocular status and the etiology of any visual loss present. One of the institutions is a school for the visually impaired (hereafter referred to as Location V), in which most of the students have normal mentation. The other is a home for severely mentally retarded, nonambulatory children (hereafter referred to as Location M). This institution was selected specifically to provide a sample of visual loss associated with severe retardation because the handful of cases of LCMV in the literature have been associated with severe central nervous system insults. Histories were obtained from records on site, and all children received a complete cyclopleged ophthalmic examination at their institution performed by the author. Patients at Location M with chorioretinal scars consistent with intrauterine infection (a possible sign of LCMV) had separate consents for blood drawing. Sera was obtained and sent for standard TORCHS titers, toxoplasmosis titers (Jack S. Remington, MD, Palo Alto, Calif), and ELISA testing for LCMV (Centers for Disease Control and Prevention, Atlanta, Ga). RESULTS: The diagnoses at Location V were varied and included retinopathy of prematurity (19.4%), optic atrophy (19.4%), retinitis pigmentosa (14.5%), optic nerve hypoplasia (12.9%), cataracts (8.1%), foveal hypoplasia (8.1%), persistent hyperplastic primary vitreous (4.8%), and microphthalmos (3.2%). The most common diagnosis at Location M was bilateral optic atrophy, which was found in 65% of the patients examined who had visual loss. Of these, the insults were most often congenital (42.6%), with birth trauma, prematurity, and genetics each responsible for about 15% of the optic atrophy. The second most common diagnosis was cortical visual impairment (24%), followed by chorioretinal scars (5%), which are strongly suggestive of intrauterine infection. Of 95 patients examined at Location M, 4 had chorioretinal scars. Two of these had dramatically elevated titers for LCMV, as did one of their mothers. One of the other 2 children died before serum could be drawn, and the fourth had negative titers for both TORCHS and LCMV. CONCLUSIONS: At both locations studied, visual loss was most often due to congenital insults, whether genetic or simply prenatal. The visual loss at Location V was twice as likely as that at Location M to be caused by a genetic disorder. The genetic disorders at Location V were more often isolated eye diseases, while those among the severely retarded at Location M were more generalized genetic disorders. Our study identified optic atrophy as a common diagnosis among the severely mentally retarded with vision loss, a finding that is supported by previous studies in other countries. In our population of severely retarded children, the target etiology of lymphocytic choriomeningitis virus was responsible for half the visual loss secondary to chorioretinitis from intrauterine infection. This is more common than would be predicted by the few cases previously described in the literature, and strongly suggests that LCMV may be a more common cause of visual loss than previously appreciated. We believe that serology for LCMV should be part of the workup for congenital chorioretinitis, especially if the TORCHS titers are negative, and that perhaps the mnemonic should be revised to "TORCHS + L." Childhood blindness and visual impairment are tragic and co PMID- 10703144 TI - Ocular torticollis. PMID- 10703145 TI - Expanding the scope of lamellar keratoplasty. AB - PURPOSE: To investigate whether applications of current technology, such as cryolathe and excimer laser, might improve outcomes and increase use of lamellar keratoplasty. METHODS: Six studies were performed, beginning with animals and progressing to human subjects. The first study compared cryolathed with hand dissected rabbit corneas to ascertain which created a smoother donor interface. The second animal pilot study was done to determine whether thickness of donor cornea resection could be accurately predicted with the cryolathe. A prospective animal trial was then undertaken to compare lamellar keratoplasty outcomes using cryolathed versus hand-dissected tissue. The fourth work extrapolated previous animal findings to lamellar keratoplasty in human disease. Finally, two ongoing studies are described. The first explores the possibility of sutureless lamellar keratoplasty. The second utilizes the excimer laser to dissect the recipient stromal bed. RESULTS: The initial animal pilot study demonstrated a clearer stromal surface in cryolathed versus hand-dissected corneal tissue. The second pilot showed that plano-powered donor tissue could be generated to predetermined thickness. The prospective animal trial revealed that clear grafts of intended thickness could be obtained with cryolathing. Human studies suggested that lamellar keratoplasty using cryolathe-prepared donor tissue may offer superior results to free-hand dissection. Finally, one ongoing study indicates that sutureless lamellar keratoplasty is untenable, and the other shows that clear grafts can be obtained by combining cryolathed donor tissue with recipient photoablation. CONCLUSION: This body of work demonstrates that use of new lamellar keratoplasty technology may offer expanded scope and better outcomes than traditional lamellar keratoplasty techniques. PMID- 10703146 TI - Studies of intrastromal corneal ring segments for the correction of low to moderate myopic refractive errors. AB - PURPOSE: Intrastromal corneal ring segments (ICRS) were investigated for safety and reliability in the correction of low to moderate myopic refractive errors. METHODS: Initially, 74 patients with spherical equivalent refractive errors between -1.00 and -4.25 diopters (D) received the ICRS in 1 eye. After 6 months, 51 of these patients received the ICRS in the contralateral eye. The total number of eyes investigated was 125. The outcome measures were uncorrected and best corrected visual acuity, predictability and stability of the refraction, refractive astigmatism, contrast sensitivity, and endothelial cell morphology. RESULTS: The 89 eyes with 12-month follow-up showed significant improvement with uncorrected visual acuities of 20/16 or better in 37%, 20/20 or better in 62%, and 20/40 or better in 97%. Cycloplegic refraction spherical equivalents showed that 68% of the eyes were within +/- 0.50 D and 90% within +/- 1.00 D of the intended correction. Refractive stability was present by 3 months after the surgery. Only 1 patients had a loss greater than 2 lines or 10 letters of best spectacle-corrected visual acuity, but the patient's acuity was 20/20. Refractive cylinder, contrast sensitivity, and endothelial cell morphology were not adversely affected. The ICRS was removed from the eyes of 6 patients. Three removals were prompted by glare and double images occurring at night; 3 were for nonmedical reasons. All patients returned to within +/- 1.00 D of their preoperative refractive spherical equivalent, and no patients lost more than 1 line of best corrected visual acuity by 3 months after ICRS removal. CONCLUSION: The ICRS safely and reliably corrects myopic refractive errors between -1.00 and 4.50 D. PMID- 10703148 TI - The association between anisometropia, amblyopia, and binocularity in the absence of strabismus. AB - PURPOSE: First, to determine if thresholds exist for the development of amblyopia and subnormal binocularity with various types of anisometropia and to confirm or refute existing guidelines for its treatment or observation. Second, to delineate any association between the degree or type of anisometropia and the depth of amblyopia and severity of binocular sensory abnormalities. METHODS: Four hundred eleven (411) patients with various levels of anisometropia, no previous therapy, and no other ocular pathology were evaluated. The effect of anisometropia (both corrected and uncorrected) on monocular acuity and binocular function was examined. RESULTS: Spherical myopic anisometropia (SMA) of > 2 diopters (D) or spherical hypermetropic anisometropia (SHA) of > 1 D results in a statistically significant increase in the incidence of amblyopia and decrease in binocular function when compared to non anisometropic patients. Increasing levels of SMA and SHA beyond these thresholds were also associated with increasing depth (and in the case of SHA, incidence as well) of amblyopia. Cylindrical myopic anisometropia (CMA) or cylindrical hyperopic anisometropia (CHA) of > 1.5 D results in a statistically significant increase in amblyopia and decrease in binocular function. A clinically significant increase in amblyopia occurs with > 1 D of CMA or CHA. Increasing levels of CMA and CHA beyond > 1 D were also associated with an increased incidence (and in the case of SMA, depth as well) of amblyopia. CONCLUSIONS: This study provides guidelines for the treatment or observation of anisometropia and confirms and characterizes the association between the type and degree of anisometropia and the incidence and severity of amblyopia and subnormal binocularity. PMID- 10703147 TI - Cultured corneal epithelia for ocular surface disease. AB - PURPOSE: To evaluate the potential efficacy for autologous and allogeneic expanded corneal epithelial cell transplants derived from harvested limbal corneal epithelial stem cells cultured in vitro for the management of ocular surface disease. METHODS: Human Subjects. Of the 19 human subjects included, 18 (20 procedures) underwent in vitro cultured corneal epithelial cell transplants using various carriers for the epithelial cells to determine the most efficacious approach. Sixteen patients (18 procedures on 17 eyes) received autologous transplants, and 2 patients (1 procedure each) received allogeneic sibling grafts. The presumed corneal epithelial stem cells from 1 patient did not grow in vitro. The carriers for the expanded corneal epithelial cells included corneal stroma, type 1 collagen (Vitrogen), soft contact lenses, collagen shields, and amniotic membrane for the autologous grafts and only amniotic membrane for the allogeneic sibling grafts. Histologic confirmation was reviewed on selected donor grafts. Amniotic membrane as carrier. Further studies were made to determine whether amniotic membrane might be the best carrier for the expanding corneal epithelial cells. Seventeen different combinations of tryspinization, sonication, scraping, and washing were studied to find the simplest, most effective method for removing the amniotic epithelium while still preserving the histologic appearance of the basement membrane of the amnion. Presumed corneal epithelial stem cells were harvested and expanded in vitro and applied to the amniotic membrane to create a composite graft. Thus, the composite graft consisted of the amniotic membrane from which the original epithelium had been removed without significant histologic damage to the basement membrane, and the expanded corneal epithelial stem cells, which had been applied to and had successfully adhered to the denuded amniotic membrane. Animal model. Twelve rabbits had the ocular surface of 1 eye damaged in a standard manner with direct removal of the presumed limbal stem cells, corneal epithelium, and related epithelium, followed by the application of n-heptanol for 60 seconds. After 6 weeks, all damaged eyes were epithelialized and vascularized. Two such treated eyes were harvested without further treatment, to be used for histologic study as damaged controls. The remaining 10 rabbits received composite grafts (consisting of amniotic membrane with expanded allogeneic rabbit corneal epithelial cell transplants) applied to the ocular surface in a standard manner followed by the application of a contact lens. At 16 days following transplantation, 5 of the rabbits were sacrificed and the corneal rims were removed for histologic study. At 28 days, the remaining rabbits were sacrificed and the previously damaged eyes were harvested for histologic and immunohistochemical study. RESULTS: Human subjects. Of the 19 total patients admitted to the study, the presumed corneal epithelial stem cells of 1 patient did not grow in vitro. Of the remaining 18 patients (20 procedures, 19 eyes), 3 patients had unsuccessful results (3 autologous procedures), 1 patient had a partially successful procedure (allogeneic procedure), and 1 patient had a procedure with an undetermined result at present (allogeneic procedure). One unsuccessful patient had entropion/trichiasis and mechanically removed the graft and eventually went into phthisis. The other 2 unsuccessful patients suffered presumed loss of autologous donor epithelium and recurrence of the ocular surface disease (pterygium). The partially successful patient receiving an allogeneic transplant had infectious keratitis delay of his re epithelialization; he has only minimal visual improvement but has re epithelialized. The patient receiving the second allogeneic graft lost his donor epithelium at day 4. Additional donor epithelium was reapplied, but the result is undetermined at present. Amniotic membrane as carrier. The in vitro preparation of the amniotic membrane with corneal epithelial stem cell graft overlay was successful. Histology documented removal of the amniotic epithelium and reapplication of corneal epithelial cells. Animal model. The 2 rabbits that had no reparative surgery following standard ocular surface injury had histology and immunopathology consistent with incomplete corneal epithelial stem cell failure with vascularization and scarring of the ocular surface. Light microscopy and immunohistologic staining with AE5 confirmed the conjunctival phenotype of the ocular surface repair but also documented the incomplete model. The allogeneic stern cell transplants had varying results. One rabbit had a suppurative infection and lost the graft. Reparative surgery failed in 2 of the rabbits, failed partially in 3 of the rabbits, was partially successful in 3 others, and was successful in 1 rabbit at 28 days. Histologic and immunopathologic study documented successful growth of corneal epithelium onto the recipient surface. CONCLUSIONS: 1. Presumed corneal epithelial stem cells can be harvested safely from the limbus and expanded successfully in vitro. 2. Expanded corneal epithelial cell cultures can be grown onto various carriers, but currently denuded amniotic membrane seems to be the best carrier for ocular surface repair. 3. Expanded corneal epithelial cell transplants appear to resurface damaged ocular surfaces successfully, but cellular tracking and further confirmation are required. 4. Expanded allogeneic corneal epithelial cell transplants are technically possible and may represent alternative treatment modalities for selected ocular surface problems. 5. These techniques potentially offer a new method of restoring a normal ocular surface while minimizing the threat of damage or depletion to the contralateral or sibling limbal corneal epithelial stem cells. 6. The rabbit model was probably incomplete and should be interpreted with caution. The complete eradication of all corneal epithelial stem cells from any eye is difficult, making confirmation of such work challenging. 7. The results of the rabbit model suggest that allogeneic grafts may restore a nearly normal ocular epithelial surface to certain ocular surface injuries. PMID- 10703151 TI - [Pneumatosis intestinalis in abdominal infection]. PMID- 10703150 TI - [An apheresis method in treatment of familial hypercholesterolemia]. PMID- 10703152 TI - [Polyethylene glycol (Macrogol)--an overview of its use in diagnosis and therapy of gastrointestinal diseases]. AB - The pharmacological rationale for the use of polyethylene glycol (PEG) in gastroenterology is its inverse relation between molecular mass and intestinal absorbability, with practically no intestinal absorption at molecular masses exceeding 3000, its lack of intestinal enzymatic degradation or bacterial metabolism, and its water binding capacity. PEG is used as a nonabsorbable marker in the evaluation of small intestinal and colonic absorption and secretion, as a marker for intestinal permeability studies, as an essential component of colonic lavage solutions used for the preparation of the colon for diagnostic and therapeutic interventions, and for the treatment of fecal impaction or chronic constipation. Since PEG has been used for decades, there is extensive data documenting its safety in short term use. Although animal experiments have also proved the safety of chronic PEG administration, the increasing use of PEG for the treatment of chronic constipation in the long term requires further surveillance to establish its safety. PMID- 10703149 TI - Brown's syndrome: diagnosis and management. AB - PURPOSE: To better understand the various etiologies of Brown's syndrome, define specific clinical characteristics of Brown's syndrome, describe the natural history of Brown's syndrome, and evaluate the longterm outcome of a novel surgical procedure: the silicone tendon expander. Also, to utilize a computer model to simulate the pattern of strabismus seen clinically with Brown's syndrome and manipulate the model to show potential surgical outcomes of the silicone tendon expander. METHODS: Charts were reviewed on patients with the diagnosis of Brown's syndrome seen at a children's hospital ophthalmology clinic from 1982 to 1997, or seen in the author's private practice. Objective fundus torsion was assessed in up gaze, down gaze, and primary position in 7 Brown's syndrome patients and in 4 patients with primary superior oblique overaction. A fax survey was taken of members of the American Association of Ophthalmology and Strabismus (AAPOS) listed in the 1997-1998 directory regarding their results using the silicone tendon expander procedure for the treatment of Brown's syndrome. A computer model of Brown's syndrome was created using the Orbit 1.8 program by simulating a shortened superior oblique tendon or by changing stretch sensitivity to create an inelastic muscle. RESULTS: A total of 96 patients were studied: 85 with Brown's syndrome (38 with congenital and 47 with acquired disease), 6 with masquerade syndromes, 1 with Brown's syndrome operated on elsewhere, and 4 with primary superior oblique overaction in the torsion study. Three original clinical observations were made: 1. Significant limitation of elevation in abduction occurs in 70% of Brown's syndrome cases surgically verified as caused by a tight superior oblique tendon. Contralateral pseudo-inferior oblique overaction is associated with limited elevation in abduction. 2. Traumatic Brown's syndrome cases have larger hypotropias than nontraumatic cases (P < .001). There was no significant hypotropia in primary position in 56 (76%) of 74 congenital and nontraumatic acquired cases despite severe limitation of elevation. 3. Of 7 patients with Brown's syndrome, 6 had no significant fundus torsion in primary position, but had significant (+2 to +3) intorsion in up gaze. Spontaneous resolution occurred in approximately 16% of acquired nontraumatic Brown's syndrome patients. The silicone tendon expander was used on 15 patients, 13 (87%) were corrected with 1 surgery and 14 (93%) with 2 surgeries. The only failure was a Brown's syndrome not caused by superior oblique pathology. Five of the silicone tendon expander patients had at least 5 years follow-up (range, 5 to 11 years). Four (80%) of the 5 patients had an excellent outcome with 1 surgery, final results graded between 9 and 10 (on a scale of 1-10, 10 is best). The fifth patient had a consecutive superior oblique paresis and a good outcome after a recession of the ipsilateral inferior oblique muscle. The AAPOS survey had a mean outcome score of 7.3, with 65% between 8 and 10. There were 9 (6%) complications reported: 4 related to scarring and 5 extrusions of the implant. Three of the 5 extrusions were reported from the same surgeon. The computer model of an inelastic superior oblique muscle-tendon complex best simulated the motility pattern of Brown's syndrome with severe limitation of elevation in adduction, mild limitation of elevation in abduction, minimal hypotropia in primary position, no superior oblique overaction, and intorsion in up gaze. CONCLUSIONS: The presence of mild to moderate limitation of elevation in abduction is common, and its presence does not eliminate the diagnosis of Brown's syndrome. The majority of Brown's syndrome patients have a pattern of strabismus consistent with an inelastic superior oblique muscle-tendon complex that does not extend, but can contract normally; not the presence of a short tendon. The presence of inelastic or tethered superior oblique muscle-tendon can be diagnosed without forced duction testing by observing the pattern of strabismus including torsion. Because of the chance for spontaneous resolution, conservative management, not surgery, should be the first line of treatment for acquired Brown's syndrome. If surgery is indicated, a novel procedure, the silicone tendon expander, is an effective option with excellent long-term outcomes. PMID- 10703153 TI - Direct adsorption of lipoproteins (DALI) from whole blood: first long-term clinical experience with a new LDL-apheresis system for the treatment of familial hypercholesterolaemia. AB - BACKGROUND: The DALI (direct adsorption of lipoproteins) LDL-apheresis system is a novel device for the removal of lipoproteins from whole blood. METHODS: We report the first long-term treatment experience (16.7 +/- 12.6 months; 57 +/- 43 treatments/patient) using different DALI adsorber sizes (DALI-500, DALI-750, DALI 1000) in seven patients with homozygous (n = 1) and severe heterozygous familial hypercholesterolaemia. For each treatment, 1.6 fold of the calculated blood volume was processed. Treatments were scheduled at weekly or two-weekly intervals. RESULTS: The smallest DALI-500 configuration was unable to achieve sufficient removal of LDL cholesterol, with the adsorber being exhausted already at desorption of 65% of the calculated blood volume. In contrast, both larger adsorber systems effectively removed lipoproteins until the end of treatment. Therefore, the DALI-750 device was used for long-term treatment. LDL cholesterol (mean pretreatment value: 179 +/- 44 mg/dl) was reduced by 73.4 +/- 7.7% and Lp(a) levels (mean pretreatment value: 43 +/- 33 mg/dl) by 69.5 +/- 8.3%. HDL cholesterol (mean pretreatment value: 47 +/- 15 mg/dl) was reduced by 16.3 +/- 8.0% during the treatment. In the long term, LDL cholesterol was reduced by 54.0 +/- 10.5%--from 259 +/- 101 mg/dl to 119 +/- 19 mg/dl. No serious side effects occurred during the treatment. Long-term evaluation of other laboratory parameters showed a reduction in haemoglobin due to treatment-associated blood loss despite frequent iron supplementation. CONCLUSION: Sufficient reductions in LDL cholesterol and Lp(a) were achieved using the DALI-750 system and the treatment was well tolerated. The easy use and short period of 153 +/- 22 minutes required for each treatment are the major advantages of the DALI system as compared to other available LDL-apheresis devices. Potential particle release from the adsorber into the circulation must be ruled out before the system can be introduced in clinical routine. PMID- 10703154 TI - [Long-term results of laparoscopic partial posterior fundoplication in patients with esophageal reflux and disorders of esophageal peristalsis]. AB - BACKGROUND: Gastrosophageal reflux disease (GERD) of long duration is frequently associated with impaired esophageal body motility. This condition has been considered unsuitable for antireflux surgery. METHODS: In order to investigate the outcome of antireflux surgery in the presence of impaired esophageal peristalsis, we studied 67 consecutive GERD patients with poor esophageal body function who underwent laparoscopic partial posterior fundoplication. A standardized questionnaire, upper GI endoscopy, esophageal manometry and 24-hour pH monitoring were performed preoperatively and at a median of 28 months (range, 6-54 months) postoperatively. Esophageal motility was analyzed for contraction amplitudes in the distal two thirds of the esophagus (level 3, 4, and 5), frequency of peristaltic, simultaneous and interrupted waves and total number of defective propagations. In addition, parameters defining the function of the lower esophageal sphincter (LES) were-evaluated. RESULTS: Following antireflux surgery 65 patients (97%) were free of heartburn and regurgitation and had no esophagitis on endoscopy, confirmed by histology. The rate of dysphagia was reduced from 49% preoperatively to 9% postoperatively (p < 0.001). There was significant improvement in esophageal peristalsis after the antireflux procedure. The median DeMeester reflux score was reduced from 33.3 to 1.1 (p < 0.001). Lower esophageal sphincter pressure and intra-abdominal length were normal after surgery. CONCLUSIONS: Partial posterior fundoplication provides an effective antireflux barrier in patients with impaired esophageal body motility in the long term. Postoperative dysphagia is avoided by improving esophageal body function. PMID- 10703155 TI - [Central nervous system activation in opioid dependent patients, evaluation with Fourier analysis of pupillary oscillations]. AB - The aim of the present investigation was to determine whether Fourier analysis of pupillary oscillations permits detection of differences in the activation of the central nervous system of opioid-addicted patients. We analysed pupillary oscillations during the recording period of static pupillometry, which lasted 25.6 s. Using Fourier analysis, the spectrum was divided into five frequency bands (0.0-0.20, 0.21-0.40, 0.41-0.60, 0.61-0.80, 0.81-1.0 Hz); the total spectrum (0-1 Hz) was also assessed. Three groups of patients were selected: the group addicted to heroin (consuming exclusively heroin) consisted of 26 patients with a mean age of 25.0 +/- 6.3 years, the methadone substitution group of 20 patients with a mean age of 30.9 +/- 8.2 years, and the morphine substitution group of 20 patients with a mean age of 33.2 +/- 4.6 years. The 3 patient groups were compared with normal controls of similar age (25.1 +/- 4.6 years). In the frequency band of 0.0-0.20 Hz the morphine group showed significantly lower amplitudes than the heroin group. Also in the frequency band of 0.41-0.60 Hz the morphine group differed significantly from the other groups concerning lower amplitudes, reflecting deactivation. In the total spectrum of 0 to 1 Hz the differences between these two groups were significant. Comparison with normal controls also showed significant differences. The groups were further divided according to dose (high/low): Patients of the heroin group as well as those of the methadone and morphine groups who had consumed higher doses showed greater activation of the central nervous system. In conclusion the morphine group was more deactivated than the methadone and heroin group and patients who received higher doses of the substances showed greater central nervous activation. Thus, the measurement of central nervous activation by means of Fourier analysis of pupillary oscillations might be useful in monitoring substitution therapy. PMID- 10703156 TI - [Computerized tomography detection of portal vein air accumulation in severe gastroenteritis during chemotherapy]. AB - Portal venous gas is caused by various pathological processes, both iatrogenic (complications of endoscopy) and non-iatrogenic (bowel ischemia, obstruction, perforated gastric ulcer, septicaemia). We report an immunocompromised patient suffering from metastatic cancer of the breast who developed severe gastroenteritis. Plain radiographs of the abdomen showed branching tubular lucencies that extended from the porta hepatis widely over the liver. A computed tomography performed to exclude air in the biliary tract demonstrated portal venous gas. Although the finding of portal venous gas has been associated with a high mortality rate and usually necessitates surgery, our patient survived without surgical intervention. PMID- 10703157 TI - [Pulsating tinnitus]. AB - Tinnitus is a frequent symptom but a tinnitus that is rhythmic and synchronous with the patient's heartbeat is rare. The symptom "pulsating noise in the ear" may be due to various cause but most frequently, by glomus tumors, intracranial hypertension and atherosclerosis of the carotid arteries. Pulsatile tinnitus can often present a serious diagnostic problem. The diagnostic evaluation includes physical examination, audiologic assessment and imaging techniques (ultrasonic examination of cervical vessels, high-resolution computed tomography of the temporal bones, nuclear magnetic resonance, angiography of the carotid arteries and magnetic resonance angiography). Evaluation should be individualized but must include a thorough ENT work up. The evaluation of the patient includes otomicroscopy, palpation and auscultation of ears and cervical region and the head positioning test. The cause of pulsatile tinnitus may be even identified on otoscopic examination. Further investigations by other specialities (neurology, internal medicine, ophthalmology) may become necessary. Life-threatening causes such as arteriovenous shunts or carotid artery stenosis must be ruled out. Nevertheless, in several cases it will not be possible to determine the etiology of tinnitus. Finally, therapeutic options which occasionally can include surgical techniques or interventional radiology are discussed. PMID- 10703159 TI - [New developments in vascular surgery]. PMID- 10703158 TI - [Informed consent and responsibility for patient education in oncology. Review of Austrian and German jurisprudence]. AB - Informed consent is currently an ethical, medical and legal requirement. Increasing public discussion concerning real or supposed malpractice has caused patients to adopt a critical attitude and has caused courts to increasingly demand informed consent for patients. Unfortunately, the legal requirements for informed consent have developed from atypical situations involving dissatisfied and injured patients rather than from the more common occurrence of physicians helping patients and having satisfied patients. In addition, the law has failed to establish explicit guidelines for physicians. We review the elements of informed consent based on current Austrian and German jurisdiction in the particular field of oncology and summarise the legal and medical realities with the aim of delineating specific criteria for decision making. PMID- 10703160 TI - [Surgical and interventional therapeutic possibilities in aneurysms of the subclavian artery]. AB - Aneurysms of the subclavian artery are extremely rare and most commonly caused by arteriosclerosis, trauma or thoracic outlet syndrome. Less frequently seen causes also include syphilis, cystic media necrosis or tuberculosis or congenital anomalies. The presence of a subclavian aneurysm can give rise to various symptoms such as a pulsating supraclavicular mass, peripheral embolism or brachial plexus compression. Generally, surgical intervention is undertaken involving ligation and extirpation of the aneurysm followed by interposition of either a saphenous vein- or synthetic vascular graft. Recent diversifications in potential therapeutic strategies include the clinical application of transluminally positioned stents for the treatment of vascular lesions. In the literature review we found more than 260 published cases of surgically treated subclavian aneurysms and additional 17 subclavian aneurysms treated by endoluminal stent application. From 1992-1997 5 subclavian aneurysms were resected in our hospital. In four cases a vein graft of the vena saphena magna and in one case a PTFE graft were used. The sensory ischaemic deficit regressed in the further follow up in four of the five cases. Patency was checked postoperatively by ultrasound sonography, angiography or MR-angiography. PMID- 10703161 TI - [Intermediate-term complications and problems after endovascular aortic stent prostheses]. AB - From August 1994 to December 1998 321 patients were treated with endovascular stentgrafts for aortic aneurysm exclusion in our hospital. Primary exclusion rate was 91% (primary leakage 8.7%) and hospital mortality was 3.7%. 6 different types of grafts were used, 5 of them commercially available. Midterm complications are due to configuration changes of the grafts, material deterioration, side branch reperfusion and changes in aortic morphology. The common pattern of clinical expression of these complications is secondary leakage (60 cases) and graft limb occlusion (37 occurrences in 30 patients). 50% of the secondary endoleaks have been treated up to now leaving the remaining patients under a thorough follow-up. Graft limb thrombosis was treated in all but three, well compensated, patients. Midterm results of the first commercially available endografts are not yet satisfying in contrast to conventional aortic repair. The recently available grafts are promising as they have a better kink resistance, no graft sutures and fewer modular components. PMID- 10703162 TI - [Interleukin pattern, procalcitonin level and cellular immune status after endovascular aneurysm surgery]. AB - Endovascular repair of AAA's using stent grafts is considered to be a minimally invasive procedure. However, in some cases deleterious inflammatory reactions, e.g., flu-like postinterventional symptoms are observed. A few patients even develop a fatal "postimplantation syndrome". It is not clear whether these postoperative complications result from a) the inflammatory and immune response to the inserted graft material, b) alterations of the vascular endothelium during the implantation procedure, c) residual thrombotic material, or d) a combination of all these causes. This clinical trial aimed to prospectively investigate the association between inflammatory mediators like interleukin-1 receptor antagonist (IL-1RA), IL-6, and HLA-DR expression on monocytes and clinical outcome in patients after repair of abdominal aortic aneurysms (AAA). Fifteen patients treated with endovascular stent grafts for abdominal aortic aneurysm (AAA-E) were compared with 15 selected control patients who underwent a conventional surgical procedure (AAA-K) during the same period. Prior to intervention, there were no significant differences in marker levels. One hour postoperatively, IL-6 (421 pg/ml vs. 21 pg/ml) and IL-1RA (10,061 pg/ml versus 407 pg/ml) were significantly increased in the AAA-K-group, whereas in AAA-E patients, these parameters increased more gradually during the first postoperative day and did not reach the same level as in the control group. There was only a slight reduction of HLA-DR expression in both groups compared with baseline and no signs indicating a postimplantation syndrome were found. No excessive inflammatory response or complicated final outcome were observed. It is unclear if this can be explained by the prophylactic use of indometacin. PMID- 10703163 TI - [Stent wire fractures and mesh loosening in explanted endovascular prosthesis]. AB - Vascular grafts are available since the middle of the 50's. Explant retrieval studies still reveal not published material degradation today, after over 40 years of product development. Endovascular grafts exist since the beginning of the 90's. New developments and modifications have lead to numerous devices. The importance of explant retrieval studies grows. MATERIALS AND METHODS: Among 33 stentgrafts, retrieved 5 to 43 months after implantation, the majority were 18 MinTec devices (17 Stentor, 1 Cragg). These 18 explants were examined by endoscopy, stereomicroscopy and scanning electron microscopy. RESULTS: The following material degradation was observed. The textile coating showed holes along the longitudinal seam, the ligatures in-between the stentframes burst, allowing the frames to dislocate. Occasional fractures of the stent wire were seen accompanied by bowl-shaped alterations of the surface. CONCLUSION: The Stentor device is, since its modification 1996, not available any longer. It was one of the most applied devices world-wide. The possible material deficiencies must be known by those performing patient follow-up. The occurrence of alterations within 43 months shows the importance of a continued follow-up besides clinical trials. PMID- 10703164 TI - [Percutaneous therapy of catheter-induced traumatic vascular lesions with Dacron coated nitinol stents]. AB - OBJECTIVE: Percutaneous peripheral interventional procedures as well as coronary interventions can be complicated by dissections and traumatic lesions of peripheral arteries. The aim of this study was to evaluate the efficacy of fabric covered endoprostheses for percutaneous repair of traumatic peripheral arterial lesions. PATIENTS AND METHODS: In this study we used the EndoPro 1/Passager device (Boston Scientific, USA), which is a self-expanding nitinol stent covered by an ultrathin layer of dacron fabric. In 27 patients a total number of 31 endoprostheses (mean length 7.3 cm) were implanted in iliac (n = 20), femoral (n = 6) and popliteal arteries (n = 1). Indications for stenting were large dissections (n = 24), arterial perforations (n = 2) and one traumatic arterio venous fistula. RESULTS: An immediate exclusion of the lesion could be achieved in all cases. There were no major procedural complications. However, within 24 hours after implantation 14 patients (51.9%) developed fever. WBC and CRP were elevated in 13 (48.1%) and 17 (63%) patients, respectively. Repeated blood cultures could not show any bacterial growth. The primary patency after a mean follow-up of 19 (5 to 31) months was 85.2%. In two cases with markedly impaired peripheral run-off subacute graft occlusions occurred. In 2 other cases the angiography revealed relevant restenoses (> 75%). The patency could be restored in 2 of these 4 cases leading to a secondary patency rate of 92.6%. CONCLUSIONS: The EndoPro 1/Passager endoprosthesis seems to be safe and effective to seal large dissections and traumatic lesions of peripheral arteries, showing a high long-term patency rate. PMID- 10703165 TI - [Perforations in recanalization of arterial occlusions of the femoropopliteal area]. AB - Interventional recanalizations may induce wall perforations. The aim of this study was to evaluate the incidence and the management of this complication in chronic femoropoliteal occlusions. 512 cases undergoing interventional recanalization (PTA or Excimer laser-assisted angioplasty) of femoropopliteal occlusions between 5 and 32 cm in length (mean: 13.8 cm) were included. Perforations were immediately examined by angiography or colour-coded ultrasonography (7.5 MHz). Perforations were observed in 22/512 cases (4.3%). The event was coincident with guide-wire manipulations in 16/512 cases (3.1%), laser catheter advancement without wire guidance in 3/58 cases (5.2%), and balloon dilatation in 3/431 cases (0.7%), while lasering via wire guidance (n = 408) did not induce perforation. Combining treatment with prolonged balloon dilatation (success: 6/8, recurrent bleeding: 4/6), ultrasound-guided compression (UGC; success: 20/20) and regulation of anticoagulation, all active bleedings could be terminated. Due to perforation, 12/512 cases (2.3%) required repeated interventions for successful recanalization. Except hematomas of 15-320 ml volume (ultrasound measurement) and prolonged hospital stays for 24-72 h, no further clinical sequelae resulted. In conclusion, perforations occurring during recanalization of long and chronic femoropopliteal occlusions have to be considered as primary interventional failure. However, with immediate treatment including UGC, the risk of serious clinical sequelae is low. PMID- 10703167 TI - [Success and failure with the Demers catheter in dialysis]. AB - Between January 1995 and January 1999 54 Demers atrial catheters were implanted in 48 uraemic patients. Indications for implantation were: urgent need for haemodialysis with missing vascular access (39), fistula occlusion (7), low shunt flow (3) and problems with a previously implanted catheter (5). We observed 7 catheter infections, 5 catheter occlusions, 1 intraoperative air embolism, 3 haematomas and 1 dacron socket dislocation. The average period of use of an atrial catheter was 170 days, the longest period almost 2 years. The majority of catheters were explanted without any dysfunction. The long time of availability makes Demers atrial catheters an alternative to fistula for multimorbid patients on dialysis with poor long-term survival. PMID- 10703166 TI - [Outcome of treatment of iatrogenic vascular lesions]. AB - The incidence and etiology of iatrogen vascular injuries have changed during the last years due to the fact that many vascular procedures, including invasive catheter procedures, minimally invasive interventions and osteosynthetic operations are more frequently performed. 66 patients who were treated between 1993 and 1997 were retrospectively analysed. The most common cause of surgical repair were lesions after catheter procedures (n = 47; 71.2%) followed by various other causes (traumatology n = 6, orthopedics n = 5, gynecology n = 4, general surgery n = 4). In patients with complicated catheterizations and need for acute vascular-surgical repair, the limb could only be preserved in 12 of 21 cases (57.1%). Vascular lesions on account of other operative specialities finally required an amputation in 3 patients. One patient in this group died. Besides the seriousness of injury one main reason for an unfavourable outcome was a delayed diagnosis with subsequent exceeding of the ischemic tolerance in 18 patients (27.3%). Some rare and serious complications are especially discussed. PMID- 10703168 TI - [Experiences with an arterialized venous flap for intraoral defect reconstruction]. AB - This clinical investigation should try out the suitability of arterialized venous forearm flaps for immediate reconstruction of intraoral defects after excision of an oral squamous cell carcinoma. In contrast to the free radial forearm flap there is no need for either sacrificing a peripheral artery or jeopardizing motor nerves. As the exact function of arterialized venous flaps is still unknown we had to take into account the possible loss of the flap. Therefore we used this flap in patients with small or medium sized defects only. All skin fat flaps were raised out of the right forearm using two different flap types. In 34 patient we used a flap with a superficial vein passing through (type I), in 5 patients we took a flap with two parallel proximal veins (type II). After the flap had been sutured into the intraoral defect, in flap type I the original distal end of the vein was anastomosed to an artery and the original proximal end to a vein. In flap type II there was no specific differentiation between the veins both. 18 (46.2%) of the flaps survived completely, 9 (23%) had superficial epithelial loss or some marginal necrosis and 12 (30.8%) became completely necrotic. Areas of partial loss developed slowly and formed stable granulation tissue. The flap donor sites were either closed primarily (n = 19) or were covered with split thickness skin graft (n = 20). There were no functional problems of the donor forearms. These results contrast with the high success rates achieved with orthodox free forearm flaps. Further research into venous flaps is essential. PMID- 10703169 TI - [Catheter ablation of the inferior epigastric artery. An conservative method for conditioning the pedicled TRAM flap]. AB - The pedicled transverse rectus abdominis musculocutaneous (TRAM) flap is a recognized, reliable method of autogenous tissue breast reconstruction after mastectomy. However, the blood supply to the distal part of the flap is often precarious after ligation of the the main feeder, the inferior epigastric artery (AEI), at time of the operation. We describe our clinical experience with a new technique to augment the superior blood stream (superior epigastric artery, AES) by selective embolization of the inferior epigastric artery some weeks prior to plastic surgery. One hundred and eleven embolization procedures were performed until now. Technique of crossover and ipsilateral epigastric spiral embolization is described, requiring minimal time and material as an in- or outpatient procedure. The anatomical situation of the m. rectus abdominis blood supply is discussed as well as possible complications of the procedure. PMID- 10703170 TI - [Clinical results and color-coded duplex ultrasound findings 4 years after conditioned TRAM flap-plasty]. AB - Presented is a new technique in preoperative conditioning of the pedicled TRAM flap employing an interventional-radiologic procedure, selective embolization of the deep inferior epigastric artery (DIEA). During a four year period in 40 patients with a mean age of 48.4 (31-66) years breast reconstruction was performed by a superiorly pedicled TRAM-flap following preoperative conditioning. 30 of 40 patients were eligible for follow-up one to five years postoperatively. The mean interval between embolization and surgery amounted to 3.6 months. In 25 of 30 cases embolization of the DIEA was performed bilaterally, in 5 of 30 cases unilaterally. 14 of 30 patients underwent preoperative radiotherapy for breast cancer. Applying CCDS the peak flow values were determined in the superior epigastric arteries (TRAM/contralateral side). Pre-embolization values (54.9 cm/s/55.8 cm/s), post-embolization values (57.2 cm/s/57.9 cm/s) and late postoperative values (61.0 cm/s/61.6 cm/s) proved a statistically significant effect of selective embolization on peak flow without relevant difference between TRAM and contralateral side (p < 0.05). Postoperative flap complications consisted of partial necrosis in 2 of 30, fat necrosis in 1 of 30, impaired would healing in 5 of 30 and postoperative bleeding in 2 of 30 cases. Abdominal would healing complications occurred in 5 of 30 cases, abdominal wall weakness was found in 8 of 30 and hernia formation in 4 of 30 cases. Corrective surgery was performed at the breast (TRAM-flap) in 22 of 30 and at the abdomen (donor site) in 9 of 30 cases. Patient acceptance concerning selective embolization and TRAM flap surgery was very high. 29 of 30 patients confirmed that they would again choose this type of breast reconstruction. The pedicled TRAM-flap following preoperative conditioning by selective embolization of the DIEA constitutes a safe and reliable method of breast reconstruction with autogenous tissue. It is superior to the pedicled TRAM-flap without delay and offers definite advantages compared to alternative techniques of enhanced flap vascularization. PMID- 10703171 TI - [Revascularization of the corpora cavernosa in ischemia-induced erectile dysfunction]. AB - Revascularization of the cavernous bodies (CB) has its place in a highly selected patient population as the only causal therapy for erectile dysfunction (ED) and provides an alternative to the implantation of alloplastic erectile aids. The indication for arterialization of the CB is currently only established when conservative treatment has failed. The most important criteria for the intervention are controversial: an age of under 50, an incidence of not more than two classical risk factors for impotence and the exclusion of diabetes mellitus. The leading revascularization procedure in German-speaking countries comprises arteriovenous shunting of the inferior epigastric artery with the dorsal vein and artery of the penis. A retrospective analysis of results is problematic due to the nonuniform indicational criteria, the multiplicity of applied revascularization procedures and a nonstandardized follow-up. Subjective assessment of improved erectility is the sole target criterion in the majority of studies. Therapeutic results range from 33 to 87% with regard to subjective success rates. Attempts to objectify the treatment results were made in only few of the studies and disclosed bypass patency in 44 to 92% one year after the intervention. The conclusions drawn at the last "Consensus Development Conference on Impotence" (CDCI) of the National Institutes of Health (NIH) in Washington have not lost their topicality in view of the great number of still unanswered questions. There the recommendation was made in 1992 to perform penile arterialization only in controlled prospective clinical trials. The European Urological Association (EUA) is currently organizing a Consensus Conference that will provide the framework for prospective studies that can serve as a basis for clarifying the open questions. PMID- 10703172 TI - [Value of laparoscopy in acute abdomen]. AB - The correct and early diagnosis and treatment are essential for the outcome of the patient with an acute abdomen. In 520 patients with the initial diagnosis of acute abdomen early laparoscopy revealed certain advantages. 183 (35.2%) patients with 11 different diseases could be treated laparoscopically, 129 (24.8%) underwent a laparotomy. In 96.7% of our patients the initial diagnosis was correct, so that these patients received a suitable therapy. The method is economically justified because we have equal costs both with CT/szintiscanning and laparoscopy, whereas the costs of MRI are nearly 400 DM higher. PMID- 10703173 TI - [Laparoscopic appendectomy in advanced stages of appendicitis]. AB - The laparoscopic stapling appendectomy (LA) is a safe and efficient procedure for all forms of appendicitis. Overall, LA was performed in 51.8% of 280 patients with advanced appendicitis, in 1998 in more than 85%, due to increasing experience. In these cases, the appendix was dissected in double-staple technique and extracted in an endo-bag. The postoperative outcome was uncomplicated even in advanced appendicitis, a reoperation had to be done in 2 patients (1.4%). Wound infections occurred only in 3.5% of patients with LA compared to 8.2% with open appendectomy, the mortality was 0%. PMID- 10703174 TI - [Surgery of chronic pancreatitis (1)]. PMID- 10703175 TI - [Effect of crown margin on the condition of the gingiva]. AB - Laser Doppler flowmetry (LDF) is a useful non-invasive technique to monitor tissue blood flow. The aim of our investigation was to study the effect of prosthetic rehabilitation on gingival condition. The gingival index and gingival blood flow (GBF) measured by LDF were studied on a female patient before, during and after the replacement of four resin-faced, nickel-chromium alloy upper front crowns to all-ceramic ones. Prior to the treatment procedure (control phase) the gingival indexes at five measuring points were calculated to 1. At the end of the rehabilitation they turned to zero. As to the relation of the gingival indexes and LDF readings a significant correlation (r = 0.37, n = 49, p < 0.01) was found during the treatment procedure. Our results suggest that the gingival blood flow values measured by LDF may reflect fairly well the condition of marginal gingiva. PMID- 10703176 TI - [The value of beta-tricalcium-phosphate (CERASORB) in pre-prosthetic surgery]. AB - Short- and medium-term experience with a pure-phase beta-tricalcium phosphate bone-substitute material (Cerasorb) is reported on the basis of clinical, radiological (panorama, 2-3D CT records) and histological examination on 52 patients. The treatment modes on these patients included the filling of cysts, sinus grafting, augmentation and the filling of parodontal lesions. Disturbance of wound healing was not observed in any of the cases, and both the radiological and the histological examinations revealed that transformation of the implanted beta-tricalcium phosphate into bone could be detected as early as in the second month. During the 10-month follow-up period, this transformation was continuous, but was not complete; completion is to be expected only after 12 months. It is important that load-bearing tissue had already developed after 4-6 months. Studies to date suggest that autologous bone is not necessary for either sinus grafting or the filling of large cysts: Cerasorb alone is suitable for this purpose. PMID- 10703177 TI - [Regional epidemiologic examination of dental health status]. AB - This study examined the oral status of the adult population in South-West Hungary using panoramic radiographs. The results indicate that in this population group 5.2% of the teeth were carious and 38.6% were missing, 3.87% were endodontically treated. Around the apices of the endodontically treated teeth in this survey in 40%, radiographic signs of periapical pathology were observed. Only 18% of the missing teeth had fixed prosthodontic restoration. The results are indicating a substantial future need for dental treatment. PMID- 10703178 TI - In memory of Professor Janusz A. Indulski (1930-1999). PMID- 10703179 TI - Medical causes of female sickness absence during economic transition in Poland. AB - The current transformation of property relations and economic restructuring, along with many other factors, influence the health condition of workers. The present study was undertaken to investigate the trends in the rate and causes of female sickness absence during the period of economic transition in Poland, based on the absence analysis in one of the largest (before the process of restructuring) plants of the motor car industry. Vital for the current trends in the workers' absenteeism is the reduction in the rate of employment. The group investigated was composed of 3215 women and 5373 men employed during the years 1989-94. The main variable examined was the reason for the worker's discharge: (1) quitting the job because of health problems, (2) retirement, (3) termination of work contract due to economic problems of the plant, (4) change of affiliation agreed between the former and present employers. The structural transformations in the plant under study brought about the discharge of about 77% of workers during the period between 1989 and 1994, mostly owing to the termination of work contracts for economic reasons, and earlier retirement. The increase in the rate of sickness absence involved to a higher extent female workers; it amounted to 30% compared to 12% for male workers. Among the workers quitting employment, the rate of sickness absence was twice as high as that for the workers still under employment. The largest differences were noted for the female population; they concerned cardiovascular diseases, neoplasms and complications of pregnancy, delivery and the puerperium. The economic transformation processes make a considerable impact on the occurrence of sick absenteeism in workplaces. Workers leaving their jobs because of health problems, as well as those discharged for economic reasons belong to the highest risk group. PMID- 10703180 TI - Evaluation of doses to patients in X-ray diagnostics in Poland. AB - The study includes a detailed analysis of patients' exposure to ionising radiation resulting from diagnostic radiology in Poland. Data concerning the number and types of examinations were collected by wide-range questionnaire surveys conducted in 1986 and 1995. As the result, the number of examinations per 1000 inhabitants was found to be 572 in 1986 and 715 in 1995. Most of the examinations were the basic, conventional ones (mostly chest and spine radiography). A great part of this study includes detailed description of the methodology of evaluating doses received by patients from X-ray procedures. The theoretical Monte Carlo simulation with the original author's computer code was used for this purpose. As the result, the doses to patients in six age groups (< 1, 1-4, 5-9, 10-14, 15-19 and the adults) were estimated. In simulation process the examined persons were represented by the appropriate mathematical human phantoms. The doses from particular examinations performed in exposure conditions routinely used in Polish radiology departments are presented. The mean effective doses for adult patients from typical examinations are: 0.11 mSv from chest, 3.0 mSv from thoracic spine, and 4.3 mSv from L-S spine radiography. Significantly higher doses were received by patients undergoing fluoroscopy procedures: 23 mSv from barium enema and 14 mSv from the stomach examination. In conclusion, the estimated exposure levels in Poland are as follows: the average effective dose per examination: (1.4 +/- 0.6) mSv in 1986, (1.2 +/- 0.5) mSv in 1995, the average effective dose per capita: (0.8 +/- 0.3) mSv in 1986 and 1995. PMID- 10703182 TI - Economic status and occupational correlates of stomach cancer in the rubber industry. AB - Associations between stomach cancer and occupational exposure to rubber remain uncertain, and thus far only a few studies have been carried out to investigate risks of stomach cancer in different jobs in the rubber industry, and confounding of the risk/exposure relationship by non-occupational risk factors, especially by the economic status. This study was aimed to explore these questions. Following a case-cohort design, we used the data of 36 stomach cancer deaths in 1973-95 and a random sample (sub-cohort) of 188 from among 1598 subjects employed in a rubber plant in Shanghai, China. We analysed stomach cancer risks by the economic status, smoking, alcohol consumption, and residential exposure to coal burning related pollution, etc., and assessed by jobs and years, unadjusted and adjusted. The rate ratios for stomach cancer were found to be 1.70 for 1-19 yr of employment and 1.79 for 20 or more yr of employment in the inner tire tube department. After the economic status was adjusted, the rate ratios were 1.54 and 1.64, respectively. Although the front processing of rubber was also included in our analysis no evidence of any excess risk of stomach cancer was found. Rate ratios adjusted for four job groups were the highest in the low income group. The association followed a pattern of a dose-response relationship--increasing rate ratios correlated with decreasing annual income (statistic trend: chi 2(1) = 8.35, p = 0.004). The data obtained suggest that excess risks of stomach cancer in the rubber workers were mainly related to their economic status and only slightly (some 1.6) to the talc powder exposure. Results of this study fail to provide any statistically significant evidence to support the hypothesis that there was high risk of death from stomach cancer among the rubber workers employed in high dusty departments. PMID- 10703181 TI - Effects of acute exposure to aromatic hydrocarbons C 9 on locomotor activity in rats. Trimethylbenzene isomers. AB - This study was performed to find out whether in acute exposure to trimethylbenzene (TMB) isomers the dose effect relationship is linear or biphasic. In experiments performed on rats, the effect of four solvents was studied: three TMB isomers: 1,3,5-TMB (mesitylene), 1,2,3-TMB (hemimellitene), and 1,2,4-TMB (pseudocumene), and toluene, known for its biphasic activity, was used as a reference compound. The solvents were dissolved in olive oil and administered to rats orally at the doses of 0.008, 0.016, and 0.032 mol/kg. Spontaneous locomotor activity was assessed with the open-field test. Solvent concentrations in peripheral blood were determined parallelly by gas chromatography on separate groups of animals. Statistics employed a two-way analysis of variance (ANOVA) and Tukey's test. The results showed that oral administration of toluene at a dose of 0.008 mol/kg induced biphasic changes in the animal locomotor activity. It was found that TMB at applied doses increased slightly the animal locomotor activity, but the magnitude of changes did not indicate their stimulating effect. Contrary to toluene, no time-effect relationship was observed after administration of trimethylbenzene isomers. The mean blood concentrations of solvents were dose-related. The highest concentrations were observed after toluene administration. PMID- 10703183 TI - Innovative multidisciplinary research in environmental epidemiology: the challenges and needs. AB - The ability of epidemiology to determine the relationships between health and environmental insults has become exceedingly difficult. The multifactorial nature of disease and the diversity of the insults, which include biologic, physical, social and cultural factors, combined with genetic susceptibility, suggest the need to develop better models of multidisciplinary epidemiologic investigation. This paper highlights the needs of an environmental epidemiologic team, discusses ways to incorporate new ideas across disciplines and to integrate constructs and paradigms of social, ecological, cultural and population determinants with individual-based exposure assessments. Innovation will be the key to the survival and increasing importance of epidemiology in addressing the public health needs of the future, but what are the ways to enhance and encourage creativity in environmental epidemiology? The process of self-renewal and continuing education will be highlighted. Additionally, the complexity of the problems and the need for clear supervision and control of multidisciplinary research efforts require a forum of communication and an 'information-processing approach' beyond those in traditional epidemiologic studies. New approaches in data management and medical informatics must be incorporated into the epidemiologic investigative framework. Methods to be included should focus on opportunities for computer-supported sharing of ideas. Such capabilities minimise the geographic distances and the disparate knowledge and training of the investigators and bring the team closer to the objectives and functions inherent in multidisciplinary investigation. PMID- 10703184 TI - Organic solvents and time-dependent sensitization. AB - The nervous system is main target of the toxic action of most of organic solvents. There is little doubt that occupational solvent exposure may result in persisting neurobehavioural disturbances--the organic solvent syndrome. Recently, the solvents are quoted among possible causes of the abnormal condition referred to as multiple chemical sensitivity (MCS), and which is characterized by a psychosomatic over-reactivity to a variety of chemicals present in food or ambient air. According to some authors, MCS is a manifestation of the time dependent sensitization (TDS), a phenomenon of progressive increase in responsiveness to chemical agents following acute or intermittent exposure, and related to some functional aberrations within limbic structures. TDS is commonly induced by psychostimulant drugs. The purpose of the present paper was to show, based on the literature data, that under circumstances of acute and repeated exposure, some solvents (mainly toluene) exert effect on behaviour and on the functional state of some neurotransmitter systems similar to that exerted by drugs known to induce TDS. Of special importance is the fact that in case of solvents the behavioural and biochemical changes suggestive of sensitization appear after exposure at levels close to those admissible in the occupational exposure, and that the concentration-effect relationship is nonlinear (an inverted U curve). To date, however, only a few of the existing data may be regarded as a direct evidence of the solvent-induced TDS. It is mainly due to the fact that the experimental protocol of a TDS study does not match the experimental routine of neurotoxicity assessment. Some data suggest that some solvents are possibly unable to induce TDS. The necessity to assess the commonly used solvents for their ability to induce TDS has been emphasized. PMID- 10703185 TI - Polish bibliography of occupational medicine, 1998. Part 2. PMID- 10703186 TI - The ACE procedure: a surgical cure for overflow fecal incontinence. PMID- 10703187 TI - Public opinion and legislators' views on tobacco policy. AB - We explored the relationship between public opinion and Kentucky state legislators' views on increasing the cigarette excise tax to curb smoking, local option to pass stricter youth access to tobacco laws, and smoking restrictions in public places. The relationship of gender, education, political party affiliation, tobacco use, and tobacco allotment ownership to public and legislators' opinions was examined using logistic regression. Data from the random, statewide University of Kentucky Public Opinion Poll (n = 628 Kentucky adults) and a Delphi study of Kentucky legislators (n = 116 members of the Kentucky General Assembly) were used in this study. Controlling for the demographic differences in gender, age, ethnicity, education, and tobacco allotment ownership between the public opinion and legislator samples, legislators were far less likely than the public to support workplace or restaurant smoking restrictions. Participants with a college education were twice as likely to favor cigarette tax hikes and four to five times more likely to favor workplace and restaurant smoking restrictions than were those without a college degree. Tobacco allotment owners and tobacco users were less likely to support raising cigarette taxes and local option to curb teen tobacco use compared to nonowners and nonusers. Findings of this study suggest that Kentucky legislators are not keeping up with public opinion about tobacco control, particularly in regard to smoking restrictions in workplaces and restaurants. Health professional organizations can play a role by educating both their membership and lawmakers about public support for tobacco control policy. PMID- 10703188 TI - Medicine--the art of metamorphosis. PMID- 10703189 TI - Physician unionization: to be or not to be. PMID- 10703190 TI - [Shift work and health]. AB - Shiftwork, in particular night work, causes disruption of biological rhythms, perturbation of social and family life, with a negative influence on performance efficiency, health and social well-being. Deterioration of health can manifest in the short-term as sleep disorders, jet-lag syndrome and accidents; in the long term there is an increased risk of gastrointestinal, psychoneurotic and cardiovascular diseases, and impairment of the female reproductive function. The evaluation of a worker's fitness for shift and night work should be strictly connected with a careful job analysis, as the primary requisite is to arrange shift schedules according to ergonomic principles and to assure suitable compensative measures. Occupational health physicians should advise shiftworkers about proper coping strategies and carefully evaluate health disorders with absolute or relative contraindications. Health checks should be aimed at detecting early signs of intolerance and their frequency should be set in relation to specific working conditions, individual characteristics, and social factors known to influence tolerance to shift work. PMID- 10703191 TI - [Mental health and work: integrated technical actions between services for preventive hygiene and worksite safety and mental health centers]. AB - We analyzed occupational and mental health activities in an occupational health service and in a mental health service using the Method of Organizational Congruences (MOC). No technical actions in either services were dedicated to mental health at work although this is prescribed by the Italian law (833/76) and has a demand among the local shared users identified in this study. We propose integrated technical action for mental health in public health services to address the risk of stress, burnout and mobbing in the workplace. Attention is drawn to the need for further research on health services in the field of organization and mental well-being. PMID- 10703193 TI - [Asthma in hairdressers: a report of 5 cases]. AB - The paper reports 5 cases of bronchial asthma in hairdressers exposed to bleaching dusts containing potassium and ammonium persulphate. All subjects complained of asthmatic symptoms at diagnosis, and underwent measurement of non specific bronchial hyperresponsiveness to methacholine, skin prick tests for common allergens, PEF monitoring during 2 weeks at work, specific bronchial challenge (SBC) test with bleaching dust, and assessment of airway inflammation by induced sputum technique. All subjects were reassessed during a follow-up of 1 to 5 years. All subjects were negative for skin prick tests, but 3 showed an abnormal PEF variability at work (Maximal Amplitude > 10%, in at least half of the monitoring period). All subjects showed a positive airway response to SBC with bleaching dust, and 4 subjects did not react to the control tests with lactose dust. One subject only showed a high percentage of eosinophils (> 3%) in the induced sputum, while all were hyperreactive to methacholine (PD20FEV1 < 0.3 mg). During the follow-up, 2 subjects stopped working and 4 were treated by inhaled corticosteroids and bronchodilators. All subjects reported a significant improvement in asthmatic symptoms, related partly to the reduction of occupational exposure in the workplace and to the efficacy of anti-inflammatory treatment. In conclusion, similar findings were observed in these 5 cases of hairdresser asthma: absence of atopy, positive response to SBC, mild changes in PEF and variable percentages of eosinophils in induced sputum. Pharmacological treatment, associated with reduction of occupational exposure, could improve asthmatic symptoms, despite continuing the job. PMID- 10703192 TI - [The incidence of malignant mesothelioma (1977-1996) and asbestos exposure in the population of an area neighboring Lake Iseo, northern Italy]. AB - The study was stimulated by the occurrence of malignant mesotheliomas among the workers of two adjacent factories located in Sarnico, near Lake Iseo (province of Brescia, northern Italy), one of which manufactured crocidolite and chrysotile ropes and gaskets until 1993. The aim of the study was: identification of malignant mesotheliomas occurring between 1977 and 1996 among the residents of 11 villages, which constituted the recruitment area of the work-force; estimation of the incidence of malignant pleural mesothelioma; collection of working histories of all cases to evaluate previous exposure to asbestos and radiation therapy. 21 cases of mesothelioma were detected (20 pleural, 1 peritoneal; 9 among males), and 20 were supported by histopathologic diagnosis. The incidence (x 100,000 person-years, standard: European population) was 2.5 (0.7-4.2) and 2.8 (1.2-4.3) among males and females, respectively, corresponding to a three-fold increase among males and a more than ten fold increase among women in comparison with the incidence reported by the Lombardy Cancer Registry. No cases had been exposed to radiation therapy, whereas all cases had been occupationally exposed to asbestos. Occupational exposure to asbestos had occurred in work on the production of crocidolite and chrysotile ropes and gaskets (6 males); in work in a textile factory producing cotton garments that was adjacent to and polluted by the former, where, in addition, chrysotile blankets were used for fireproofing in the weaving area and pipes were insulated using amosite-containing materials (10 cases, 6 among females); 5 cases occurred among women working in silk factories, where asbestos exposure was possible because of the presence of pipes insulated with asbestos and because women were handling temperature-controlled trays insulted with asbestos. In conclusion, the study demonstrated that the occurrence of mesothelioma was higher among females than males in the study area and that all cases of mesotheliomas had been occupationally exposed to asbestos. PMID- 10703194 TI - [The epidemiological trend of brucellosis in the provinces of Sicily]. AB - The epidemiological trend of brucellosis in Italy has been uneven over the last few years since there was a decrease in incidence in some regions and an increase in others, including Sicily. The peak was reached in 1997 when 59% of the cases were reported in Sicily alone. Appropriate intervention strategies are therefore needed both as regards the general population and exposed workers in order to reduce the spread of this disease. PMID- 10703195 TI - [100 years of lead poisoning studies from a reading of articles published in La Medicina del Lavoro]. AB - In preparing this paper we considered the articles published in La Medicina del Lavoro from 1901, its first year of publication. This scientific journal was founded in Milan, when an animated debate arose in Italy on the necessity of treating and, above all, preventing occupational diseases. In the same city, the "Clinica del Lavoro" (i.e. Institute of Occupational Medicine) was inaugurated in 1910. Its founder, Professor Luigi Devoto, had to overcome numerous obstacles caused by the hostility of the Rector of the University of Pavia--the future Nobel prize winner Camillo Golgi--and the clinicians of the main hospital of Milan, founded by Francesco Sforza in the XV century. From reading a century of articles which appeared in La Medicina del Lavoro, it is clear that for occupational physicians lead is an exemplary topic by which to evaluate the evolution of research in the field of occupational diseases. The numerous pathological features of lead poisoning, the successive therapeutic responses of physicians, and the gradual development of preventive techniques constitute a paradigm that has subsequently been applied to all other fields of industrial toxicology. Reading the papers of 100 years gives a clear picture of the evolution of clinical syndromes over the decades. The pathological picture of lead poisoning gradually became less serious and progressively changed into aspecific, subclinical manifestations. The categories of workers in which lead poisoning had a high incidence changed over the years: painters, printers and munition makers had the highest incidence in the first three decades of this century; afterwards, those engaged in lead smelting, alloy production, painters, and in the last few decades those employed in battery, ceramic and PVC production. Prevention consisted mainly of early diagnosis of lead poisoning and instruction in proper hygiene measures. Later, in 1929, insurance of occupational diseases was made compulsory in Italy, and among the few risk factors covered by law were lead and its compounds. This law was a great advance not only in the diagnostic and insurance fields but also for prevention. Two aspects of occupational lead poisoning are particularly instructive: treatment on the one hand and the use of laboratory analysis on the other. In treatment, the initial approach was mainly empirical and physicians insisted on evacuation of the bowel. Laboratory analysis started in the 20's with analysis of erythrocytes with basophilic stippling and continued with the study of urinary porphyrins. This was followed by the determination of lead in blood and urine. These tests were used initially as diagnostic tools, and only since the 60's they have been used for biological monitoring of workers for preventive purposes. The identification of indicators of dose, of critical/subcritical effect, and of critical organ started with studies on lead poisoning. Since then, following this model, biological monitoring has been applied to numerous other metals, solvents, and pesticides. The evaluation of the Italian scientific literature on lead over one hundred years in La Medicina del Lavoro has been a very exciting experience. It suggests that knowledge of the evaluation of lead poisoning and lead exposure should be taught to medical students and young physicians, thereby stimulating them to put into practice the maxim that was engraved on the foundation stone of the Clinica del Lavoro: in aliis vivimus, movemur et sumus. PMID- 10703196 TI - [A century of industrial medicine in Italy: the opportunity of a debate for relaunching the discipline]. PMID- 10703197 TI - [Health legislation of the Russian Federation on the prevention of parasitic diseases (new standards and methodological documents)]. AB - The paper summarizes the results of the 1994-1998 activities in preparing sanitary legislative documents and standard guidelines that define strategies and tactics to prevent parasitic diseases in the Russian Federation under the present conditions. Moreover, it lists the guidelines that are being developed and planned for approval in 1999-2000. PMID- 10703198 TI - [Current problems in medical entomology]. AB - The major problems facing medical entomology as a science and practical health care facilities in the Russian Federations allows to outline the tasks to be solved in order of their priority and significance. These include the study and monitoring of tick-borne infections, resurrecting malaria, gnat-induced diseases, acariases, allergosis and pediculosis. It is emphasized that medical entomology as a science cannot develop since the man-made changes of the environment and the predicted global warming of the Earth climate are not taken into account. The present status of medical entomological service is considered to be poor. Governmental support is required. PMID- 10703199 TI - [Is not plague a "protonosis"? (the role of Protozoa in the epizootiology of plague)]. AB - The author expounds the idea that soil protozoa, whose vegetative forms and cysts can harbor the plague agent for fairly prolonged periods of time, can be a major player in the epizootiology of plague. It is also postulated that the symbiotic protozoa of the digestive tract of rodents and lagomorpha can also be a reservoir of the plague agent. If this is so, among apparent epizootic cycles in mammalians in wild plague foci one should look for Yersinia pestis in the protozoa from the burrows of their primary and secondary carriers. Because parasitism of bacteria in one-celled animals is essentially epizootic, plague epizootics are presumed to be a permanent process. PMID- 10703200 TI - [The biodiversity dynamics of the causative agents of diseases transmitted by ticks in the genus Ixodes: an analysis of multiyear data]. AB - Fauna of pathogen's met in the organism of the primary tick-borne disease vectors -Ixodes persulcatus Schulze and Ixodes ricinus (L.) was observed. Prevalence of Borrelia mono- and poly-infection in the I. persulcatus ticks within a season of the vector activity was analyzed and increase of the number of the dual infected specimens during the season was demonstrated. The first determination of Ehrlichia infected I. ricinus and I. persulcatus collected in the Baltic region of Russia was stated. The triple infection of Ixodes ticks in was proved: infection by the two species of Borrelia and Ehrlichia; infection by the three species of Borrelia and infection by the tick-borne encephalitis virus and two species of Borrelia. The first determination of the tick-borne encephalitis virus in I. ricinus in the recreational zone of Kaliningrad Province (Courland [correction of Curonian] Spit) was described. Dipetalonema sp. was detected in the St. Petersburg population of I. persulcatus. The prevalence of poly-infection among I. persulcatus ticks was stated. PMID- 10703201 TI - [Tick-borne encephalitis and Lyme disease: the epizootiological parallels and monitoring]. PMID- 10703202 TI - [An experimental study of the capacity of the rat mite Ornithonyssus bacoti (Hirst, 1913) to ingest, maintain and transmit Borrelia]. AB - For the first time a possibility of the gamasina mites' O. bacoti participation in Lyme disease spirochetes' circulation has been demonstrated. It has been experimentally shown that Borrelia burgdorferi s.l. are received by O. bacoti, survive in them for at least 21 days and are transmitted to white mice through mites' bites. Mice's infestation has occurred in 23% of cases. It is suggested that other bloodsucking gamasina mites inhabiting the Lyme borreliosis reservoir rodents nests may be capable of participating in borrelia circulation in the Lyme disease endemic areas. PMID- 10703203 TI - [The biological regional division by biting midges of the territory of western Kazakhstan]. PMID- 10703204 TI - [Glycyphagus cadaverum housedust mites--a powerful source of allergens in Uzbekistan]. AB - The house-dust mites, Glycyphagus cadaverum (Schrk.) and Dermatophagoides pteronyssinus (Trt.) are prevalent species in the houses of Uzbekistan. Their prevalence rates are 57.8 and 28.5%, respectively. G. cadaverum was detected in 62. 13% of the houses. It numbered as many as 1013 per g of dust. The tick, house dust and feeding media were extracted and allergic reactions to the extracts were examined by three immunological assays: passive hemagglutination test, indirect degranulation test, and common anaphylactic reaction: The study demonstrated that the extract from G. cadaverum had a higher allergenic potency, which is commonly used in Uzbekistan for the diagnosis of asthma. PMID- 10703205 TI - [The action of anticoagulants on the transmission of the causative agent of plague]. AB - The anticoagulants 4-hydrocoumarine and 1,3-indandione derivatives produce a varying action on the formation of a gizzard block in plague-infected fleas. The feeding of fleas on the animals having typical signs of severe intoxication with ratindane, zoocoumarine, and tomorine reduces the time and incidence of gizzard block formation in the fleas. The individuals with blocks retain their high infectability and the development of an infectious process is mainly generalized in rodents. In this connection, the package of eradication measures should involve the concurrent use of insecticides and zoocides which can reduce the size of both rodents and fleas. PMID- 10703206 TI - [A comparative analysis of the potential epidemiological importance of mosquitoes in the genus Anopheles under the anthropogenic landscapes in the Chu Valley, Kyrgyz Republic. III. The age composition of the females in natural populations of the dominant malarial mosquito species]. AB - The age composition of female malaria mosquitoes prevailing under the conditions of the man-made areas of the piedmont zone of the Chu River Valley, Republic of Kirghizia. That of the natural An. claviger and An. messeae populations was examined throughout the period of mosquito activity in the season of 1992. The physiological age of female malaria mosquitoes was determined by the expansion of ovarioles with terminal legs (Polovodova, 1947; Detinova, 1949), the maximum quantities of the expansions being borne in mind (Anufrieva, Artem'ev, 1981). A total of 2,606 females were dissected. In the piedmont zone of the Chu River Valley An. claviger and An. messeae were found to make at least five and six cycles, respectively, which is suggestive of their potential epidemiological significance. PMID- 10703207 TI - [The effect of antihormones on the susceptibility of Anopheles sacharovi Favre. mosquitoes to the causative agent of malaria Plasmodium gallinaceum Brumpt]. AB - A laboratory model of circulation of the malaria causative agent P. gallinaceum has been used to show that the effect of precocene (antijuvenoid) leads to a statistically significant reduction in the proportion of infected females developing eggs after blood suction. The females failing to develop eggs are not infected. Trichopol (antiexdisone) inhibits vitellogenesis The females undeveloping eggs become susceptible to the causative agent though to a lesser degree than those developing them. The findings suggest that there is an association of the mosquito susceptibility to the malaria causative agent with the balance of hormones in the body of disease the carrier. PMID- 10703208 TI - [Imported malaria in the Kabardino-Balkarian Republic]. AB - Forty-three cases of imported vivax malaria were notified in the Republic of Kabardino-Balkaria in 1981-1997. These included 40 military men serving in Afghanistan, a citizen from this country, a student of the Kabardino-Balkaria State University, a student of a Nal'chik technological college, and a newcomer from Armenia. Among the patients, urban and rural residents were 27.9 and 72.1%, respectively. 37.2% were detected in the season of effective mosquito infection; new cases of malaria were identified in 41.9%, relapses were found in 58.1% of patients. Analyzing the reasons of late diagnosis of imported malaria suggests that the patients visited health facilities too late due to their poor awareness of a risk for malaria. Out of 43 patients, 62.8% referred to in the first 3 days after the onset, 18.6, 9.3, 9.3, and 2.3% did on days 4-6, 7-15, 16, and after a month, respectively. Their physicians made diagnostic errors in 53% of cases. Acute respiratory disease, influenza, pneumonia, and hepatitis were most commonly diagnosed. They were found in 27.9, 11.6, 4.7, and 4.7%, respectively. PMID- 10703209 TI - [The importance of fluid perfusion for the outcome of the disease in malarial coma due to Plasmodium falciparum in adult Africans]. AB - Malaria coma induced by P.falciparum was diagnosed in 51 of 390 adult African patients who had been admitted to the therapeutical unit of the Donk Central Hospital and were receiving parenteral quinine at day 1 of the onset of coma. Examining some clinical and laboratory manifestations of malaria coma indicated that fatal outcome was significantly recorded among the patients with severe concomitant anemia and among the patients who had not or had received inadequate liquid parenterally on the first day of coma. The occurrence of acute renal failure in 4 patients with malaria coma resulted in 3 deaths. No great impact on the prognosis of malaria "hyperparasitemia", the severity of fever, the values of blood pressure was found. Whether it is advisable to use the parameters characterizing the opportuneness and scope of health care delivered to patients with severe malaria is discussed. PMID- 10703210 TI - [Anthropourgic foci of diphyllobothriasis in the basins of large lakes in the European north of Russia]. PMID- 10703211 TI - [A vaccine for the prevention of echinococcosis in sheep in Uzbekistan]. PMID- 10703212 TI - [Argasid tick-borne borreliosis]. PMID- 10703213 TI - Against the odds: growing up with parents who have learning difficulties. AB - For this article we drew on material from a study in which we explored how people who were brought up in a family headed by a parent or parents with learning difficulties managed the transition to adulthood. Using evidence from in-depth interviews, we provided an assessment of how the now-adult children came through what would generally be seen as a risk-filled upbringing. Despite the problems they encountered in their childhood, many of which originated outside the home, most of the informants had maintained a valued relationship with their family and remained close to their mother. PMID- 10703214 TI - Decision-making by adults with mental retardation in simulated situations of abuse. AB - The ability of women and men with mental retardation to suggest prevention focused decisions in response to simulated social interpersonal situations of abuse was investigated. Decision-making performance across three types of abusive situations (physical, sexual, psychological/verbal) was examined. Participants were able to suggest direct prevention-focused decisions aimed at resisting or stopping abuse 45% of the time and other-dependent prevention-focused decisions in the form of reporting 20% of the time. Prevention-focused decision-making was higher in situations of physical abuse (59%) than in situations of sexual (51%) or psychological/verbal abuse (26%). Women and men did not differ significantly in their decision-making responses. PMID- 10703215 TI - Intensive outpatient treatment of persons with mental retardation and psychiatric disorder: a preliminary study. AB - The impact of intensive outpatient mental health interventions (in a dual diagnosis clinic) on the hospitalization rate and length of stay was examined for 28 adults with mental retardation and severe psychiatric disorder. They were selected on the basis of frequent use of mental, medical, and social services. Charts were reviewed for the 12-month periods before and after referral to the program to compare service utilization. A single group pretest-posttest design with no control group was employed. Correlated t tests comparing the pre- and post-program number of hospitalizations and lengths of stay indicated significant decreases in both hospitalizations and lengths of stay after program entry, which may result in significant reductions in hospital costs. PMID- 10703216 TI - Caring for people with developmental disabilities: survey of nurses about their education and experience. AB - Over 500 nurses in New Jersey responded to a survey on education and training in the area of developmental disabilities. Respondents provided information on their work experience, experience with patients who have developmental disabilities, and opportunities for continuing medical education. Results showed that although many nurses thought educational activities related to developmental disabilities were important, only about 10% said that they received "a lot" of training. Most respondents (almost 60%) said that they received little or no training in the area, and most received no specific training on developmental disabilities since receiving their licenses or in their current job. Implications of these findings in light of the movement of people with developmental disabilities into community living and managed care plans are discussed. PMID- 10703217 TI - Motor task persistence of children with and without mental retardation. AB - Task persistence by 31 children with and without mental retardation during two challenging motor tasks was investigated. We used a 2 (group) x 2 (gender) MANOVA to analyze trials and seconds per trial. A main effect was found for group affiliation: Children without mental retardation attempted more trials over three sessions. No significant differences were found for seconds per trial, which indicated that all study participants experienced a comparable level of failure in regard to seconds completed before failure. Findings support the hypothesis that children with mental retardation are less persistent at challenging motor tasks than are peers without disabilities. These findings have both theoretical and practical implications. PMID- 10703218 TI - Leaving home at an early age: parents' decisions about out-of-home placement for young children with complex medical needs. AB - Qualitative methodology was used to examine the circumstances leading to the realization of the need for out-of-home placement. A series of indepth, semi structured interviews were conducted with 5 parents who had made the decision for out-of-home placement. Results indicated that these parents went through a similar process: initial feelings of excitement turn to weariness, first thoughts about the need for additional assistance in caring for their child, realization of the need for out-of-home placement based on minimal social supports, difficulties with services, overwhelming medical care, and mounting financial concerns ("triggering events"); need for approval for the decision; and reflections on the decision. Conclusions and implications for practice, policy, and future research are provided. PMID- 10703219 TI - Word from Washington. PMID- 10703220 TI - The power of mental retardation: reflections on the value of people with disabilities. PMID- 10703221 TI - [Pathology of alcoholic liver disease]. AB - The history of alcohol consumption has been nearly as long as the history of mankind. Alcohol-related diseases represent a serious problem all over the world and they show a gradually increasing tendency. It can be stated that the frequency of occurrence, severity and mortality of alcohol-related hepatic diseases are in direct correlation with the amount of alcohol consumed. The direct hepatotoxic effect of alcohol and its metabolites has become obvious by now. In addition to this, other mechanisms also play a part in the development of hepatic diseases: their occurrence and severity are significantly influenced by genetic and environmental factors. The rather wide spectrum of alcohol-related hepatic diseases includes steatosis, perivenular fibrosis, alcohol-related hepatitis, occlusive venous lesions, cirrhosis and hepatocellular cancer. All of these disorders are characterized by clearly defined, characteristic but non specific changes, which need to be supplemented by histological diagnostic criteria. Cirrhosis, which must still be regarded as an irreversibly lethal condition, is thought to develop in two ways. A well-known and widely accepted assumption is that episodes of alcohol-related hepatitis aggravated by progressive fibrosis sooner or later lead to cirrhosis. Another possible explanation is that steatosis facilitating the development and spreading of perivenular fibrosis--even without episodes of hepatitis--may lead to cirrhosis. Thus, alcohol-related hepatic conditions have characteristic pathohistological features, none of which, however, are pathognomonic at the same time. Therefore, the definitive diagnosis of any form of alcohol-related hepatic disorders needs to take evidence of alcohol consumption into account. PMID- 10703222 TI - [T3-thyrotoxicosis: incidence, significance and correlation with iodine intake]. AB - In a part of patients with thyrotoxicosis the serum triiodothyronine concentration increases only while free thyroxin level remains in the normal range (T3-thyrotoxicosis). This condition occurs in patients with untreated or treated thyrotoxicosis and in some other but rare thyroid disease, respectively. In this study occurrence and importance of T3-thyrotoxicosis were discussed. This form occurred in 11% of untreated thyrotoxicosis (53/480). The majority of patients suffering from T3-thyrotoxicosis have autonomous thyroid function (i.e. toxic uninodular and multinodular goiter: 45/53, 85%). In case of suppressed TSH and normal free thyroxin T3-thyrotoxicosis can be expected in 40% (53/140). The rate of free-triiodothyronin elevation is highest in Graves' disease with autoimmune origin. We consider the possibilities of development of triidothyronine increasing and the importance of iodine deficiency. Our patients are living in an area where mild iodine deficiency can be proved on the basis of decreased iodine excretion in urine. The results show the diagnostic importance of free-T3 determination. PMID- 10703223 TI - [Neuronavigation in pediatric neurosurgery]. AB - The precise orientation in the intracranial space is essential for the minimal invasive neurosurgical interventions. The CT and MR based neuro-navigation permits small, targeted exposures on the skull, and intraoperatively gives exact graphic-interactive guidance to the targeted intracranial lesions. The use of neuro-navigation can shorten the time of surgery and diminish the surgical mortality and morbidity. The favourable experiences of the first 21 neuro navigation aided operations in pediatric patients performed in the National Institute of Neurosurgery (Budapest, Hungary) with the Vector Vision Neuro navigation System (BrainLAB Gmbh, Germany) are discussed. PMID- 10703224 TI - [The origin, development and aging of hematopoietic stem cells]. AB - The past few years provided a number of challenges to our expectations regarding hematopoietic stem cell biology. Evidence has emerged that hematopoietic stem cells arise intraembryonally before they can be detected in the yolk sac. A number of genes that may regulate the formation, self-renewal, or differentiation of stem cells have been identified. Although different groups have attributed different properties to hematopoietic stem cells, it now appears that the differences may be explained by the existence of various stem cell populations, each with different potencies ranging from totipotent to more specialized, exist and may persist into adult life. Finally, we propose that the number of hematopoietic stem cells available to an individual is finite and the 'quality', although currently a subjective parameter, is of newly appreciated importance. PMID- 10703225 TI - [Prostatic tissue in a dermoid cyst of the ovary]. AB - The finding of mature prostatic tissue in a mature cystic teratoma (dermoid cyst) of the right ovary of a young girl is reported. Prostatic tissue rarely occurs in association with a normal phenotype in females. Reports of prostatic tissue in the female genital tract and their possible aetiology are also summarized in the paper. PMID- 10703226 TI - Eating disorders in adolescent girls. AB - Eating disorders are common in contemporary society. New information is emerging on the pathogenesis of anorexia nervosa and bulimia nervosa and includes psychologic, biologic, family, environmental, genetic and social factors. The physician providing care to adolescents is challenged to carry out a careful evaluation and monitor the patient for complications, especially loss of bone mass. Treatment requires a multidisciplinary team. PMID- 10703227 TI - Unfavorable lipid profiles in mild obesity with excess body fat percentage. AB - BACKGROUND: The aim of the present study was to investigate the usefulness of subclassifications of overweight children using the body fat percentage (Fat%) to predict the serum lipid profile. METHODS: School children (431, 236 boys and 195 girls) aged 9-12 years were divided into three obesity groups (non-, mild and advanced obesity) and were further divided into two subgroups according to the Fat% measured by bioelectrical impedance analysis. The mean fasting serum lipid levels were also evaluated. RESULTS: In the non-obesity and the advanced obesity groups, the Fat%-based subclassification demonstrated no essential differences in lipid profiles or in the prevalence of hyperlipidemia between the two subgroups. However, in the mild obesity group, the levels of low-density lipoprotein cholesterol and triglyceride and the atherogenic index were significantly higher and the high-density lipoprotein cholesterol level was significantly lower in the adipositic subgroup (Fat% > or = age/sex-specific cut-off value) than in the non adipositic subgroup. Multiple comparison of lipid levels among all six categories of children indicated that the adipositic subgroup of mild obesity had no advantage over the advanced obesity group with respect to the atherogenic potential and that the non-adipositic subgroup of mild obesity showed no additional risks compared to the non-obesity group. Moreover, the prevalence of hyperlipidemia in the adipositic subgroup of mild obesity (50.0%) was significantly different from that in its non-adipositic counterpart (13.3%) and was equivalent to that in the advanced obesity group. CONCLUSIONS: These results suggest that Fat% evaluation is useful to divide mildly obese children into two distinct subtypes based on serum lipid profiles and that the excess Fat% in mildly obese school children is a predictor of atherogenesis. PMID- 10703228 TI - Comparison of the fatty acid composition of total lipids and phospholipids in breast milk from Japanese women. AB - Fatty acid (FA) composition of total lipids (TL) and phospholipids (PL) in breast milk obtained from 20 normal delivery healthy women in Tokyo, Japan was analyzed. Total lipids were extracted from the samples and then PL, consisting of phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine (PS), sphingomyelin (Sph) and phosphatidylinositol (PI), were separated by two dimensional thin-layer chromatography. The FA composition of TL and PL was analyzed by gas liquid-chromatography. Compared with previous reports, the contents of eicosapentaenoic acid (EPA; 20:5n-3) and docosahexaenoic acid (DHA; 22:6n-3) in TL from Japanese women were higher than those from Chinese and Canadian women, which may be caused by different dietary habits and food types consumed by those populations. The contents of arachidonic acid (AA; 20:4n-6), EPA and DHA in PE and PI were much higher than those in PC. In addition, no significant correlation of EPA or DHA content was found between TL and PL. The findings indicate that PL especially PE and PI in human milk may be a source of EPA and DHA for infants in the rapid developmental stage. These results should be considered in infant formula production. PMID- 10703229 TI - Analysis of IgG subclasses in chronic active Epstein-Barr virus infection. AB - BACKGROUND: Although elevated serum levels of immunoglobulins are frequently observed in patients with chronic active Epstein-Barr virus (EBV) infection, there have been no reports concerning levels of IgG subclasses. METHODS: Serum levels of IgG subclasses were measured by the enzyme-linked immunosorbent assay (ELISA) in 30 children with severe chronic active EBV infection. RESULTS: Serum levels of IgG1 were elevated in most patients, except for one who showed an abnormally low level of IgG1 and progressive hypogammaglobulinemia. Serum levels of IgG2, IgG3 and IgG4 in the patients were comparable to those in control children, while abnormally low levels of IgG2, IgG3 and IgG4 were observed in six, three and four cases, respectively. CONCLUSION: Although not always susceptible to bacterial infections, low levels of IgG2 were frequently observed in patients with chronic active EBV infection and elevated IgG1 is responsible for the increase of serum IgG in these patients. PMID- 10703230 TI - Physiologic significance of nitric oxide and endothelin-1 in circulatory adaptation. AB - BACKGROUND: The purpose of the present paper was to evaluate the physiologic significance of nitric oxide (NO) and endothelin (ET)-1 in circulatory adaptation in the neonate. METHODS: The serum levels of NO metabolites (NOx; the sum of nitrites and nitrates) and the plasma level of ET-1 were determined in 14 healthy full-term infants at 0-6 h, 24 h and 5 days after birth. We measured the heart rate, the mean systemic blood pressure and the mean pulmonary arterial pressure, estimated by pulsed Doppler echocardiography, at each time point. RESULTS: The serum concentration of NOx was lowest at birth and increased with age. The plasma concentration of ET-1 was highest at birth and decreased with age. The ratio of NOx to ET-1 was inversely related to the estimated mean pulmonary arterial pressure in the early neonatal period. The ratio of NOx to ET-1 was not correlated with the systemic blood pressure. CONCLUSION: Increased NO synthesis and decreased production of ET-1 during the early neonatal period may contribute to the decrease in pulmonary arterial pressure. PMID- 10703231 TI - Relationship of insulin-like growth factor-I, insulin-like growth factor binding protein-3, insulin, growth hormone in cord blood and maternal factors with birth height and birthweight. AB - BACKGROUND: To determine whether the following factors are related to birthweight or birth height, we measured insulin-like growth factor (IGF)-I, insulin-like growth factor binding protein (IGFBP)-3, insulin and growth hormone (GH) levels in cord blood and also observed the relationship between birthweight, birth height and maternal factors. METHODS: One hundred and ninety-four cord bloods were collected, 106 from males and 88 from females. Three newborns were small for gestational age (SGA), 168 were appropriate (AGA) and 23 were large (LGA); 21 newborns were preterm and 172 were term. RESULTS: Levels of IGF-I and IGFBP-3, measured by enzyme-linked immunosorbent assay, were significantly lower in preterm babies (35.3 +/- 15.1 and 1025.6 +/- 562.8 ng/mL, respectively) than in term babies (61.6 +/- 39.5 and 1252.6 +/- 403.2 ng/mL, respectively; P < 0.01), but neither insulin nor GH levels, measured by radioimmunoassay, showed any significant difference between the two groups (P > 0.05). Among term babies, IGF I and IGFBP-3 levels were significantly higher in the LGA group (96.1 +/- 34.1 and 1544.7 +/- 418.1 ng/mL, respectively) than in the AGA group (56.4 +/- 37.6 and 1212.8 +/- 383.4 ng/mL, respectively; P < 0.01). Levels of IGF-I and IGFBP-3 showed significant correlation with birthweight and length, respectively (P < 0.01), although GH and insulin levels did not (P > 0.05). There was a significant correlation between IGF-I and IGFBP-3 levels (P < 0.01, r = 0.64), but IGF-I and IGFBP-3 levels showed no relationship with GH or insulin levels. Birthweight correlated significantly with prepartum maternal weight, maternal weight gain and maternal height (P < 0.05), but birth length correlated significantly only with maternal height (P < 0.05). CONCLUSIONS: Our results suggest that fetal growth depends on fetal levels of IGF-I and IGFBP-3 and maternal factors, not on insulin or GH. Levels of IGF-I and IGFBP-3 may not be regulated by insulin alone, but by the complex interactions between several factors, such as insulin, GH and maternal factors. PMID- 10703232 TI - Effect of socioeconomic status on the blood pressure in children living in a developing country. AB - BACKGROUND: Lower socioeconomic status has been reported to favor higher blood pressure both during childhood and adulthood, because obesity is more prevalent among this population. The aim of the present study was to evaluate the effect of socioeconomic status on blood pressure and prevalence of obesity among children living in a developing country. METHODS: Prepubertal primary school children (total number 1024, male/female ratio 513/511, mean age 10.32 +/- 0.60 years) living in Izmir, a metropolis of Turkey, were enrolled in the present study. The children were classified into three groups according to the locations of their schools as those from the well-developed (WD; n = 290), moderately developed (MD, n = 356) and underdeveloped (UD, n = 378) areas. Body mass index (BMI), ratio of BMI to the 50th percentile value of BMI for that age (BMI%) and systolic (SBP) and diastolic blood pressures (DBP) were determined in all children. Each socioeconomic group was subdivided into three subgroups with respect to BMI% as those with less than 90% (SG(< 90)), 90-110% (SG(90-110)), and more than 110% (SG(> 110)). Then, mean SBP and DBP in each subgroup of the WD, MD and UD groups were compared with the corresponding blood pressure values of each other to evaluate the effect of socioeconomic status on the blood pressure. In addition, the SBP and DBP of each subgroup were compared with other subgroups within that group to evaluate the effect of BMI on blood pressure. RESULTS: The BMI and BMI% of the UD group were significantly lower than that of the WD and MD groups (P < 0.05). In addition, the number of children with a BMI of more than 95% for their age was significantly lower in the UD group compared with the WD and MD groups (1.7, 1.9 and 0.5% in the WD, MD and UD groups, respectively, P < 0.05). The SBP and DBP were positively correlated with BMI% in each group and in all of the subjects cumulatively (r = 0.26, P < 0.001 for SBP and r = 0.34, P < 0.001 for DBP). The SBP and DBP were significantly higher in SG(> 110) than in SG(90-110) and were also higher in SG(90-110) than in SG(< 90) in each group. Mean DBP values in all subgroups of the MD and UD groups were significantly lower than the respective subgroups of the WD group. The SBP of SG(< 90) and SG(90-110) of the UD group were significantly lower than those of the corresponding subgroups of the WD and MD groups. CONCLUSION: Obesity is not more prevalent among the children of lower socioeconomic classes in Turkey as a developing country. In addition, independent of anthropometric structure, DBP and SBP were shown to be related to the socioeconomic status in childhood age groups and both values were determined to decrease in accordance with a decrease in the socioeconomic level. PMID- 10703233 TI - Expression of tumor necrosis factor-alpha protein in the myocardium in fatal myocarditis. AB - BACKGROUND: Tumor necrosis factor (TNF)-alpha is the most studied cytokine in the failing human heart and in experimental murine myocarditis. We have investigated the expression of TNF-alpha in the myocardium in human myocarditis. METHODS: We examined endomyocardial biopsy (n = 4) and autopsy (n = 5) tissues obtained from nine patients diagnosed with myocarditis by the Dallas criteria. Expression of TNF-alpha in the hearts was immunohistochemically studied using monoclonal antibodies against human TNF-alpha. RESULTS: Tumor necrosis factor-alpha protein was expressed in the myocardium of six of the nine patients studied. Four of five fatal patients showed intense immunoreactivity for TNF-alpha compared with survivors. Furthermore, left ventricular systolic function was reduced in patients with TNF-alpha-positive hearts. CONCLUSIONS: These findings may support the suggestion that TNF-alpha plays an important role in cardiac dysfunction and myocytic damage in fatal human myocarditis. PMID- 10703234 TI - Evaluation by exercise testing of children with mild and moderate valvular aortic stenosis. AB - BACKGROUND: The aim of the present paper was to determine the factors related to sudden death in aortic stenosis. METHODS: The factors related to sudden death were investigated in 40 asymptomatic children with mild and moderate aortic stenosis by treadmill testing. RESULTS: The QT interval of aortic stenosis cases were significantly longer than those of healthy children with increasing heart rates during exercise. CONCLUSIONS: A longer QT interval of aortic stenosis cases compared to normal children during exercise is the first sign of myocardial ischemia and leads to fatal ventricular arrhythmias and sudden death. For this reason we recommend that exercise testing should be performed frequently in aortic stenosis patients and that close follow up is necessary for patients with long QT segments that can be a marker for severe arrhythmias. PMID- 10703235 TI - Spectrum of renal osteodystrophy in children on continuous ambulatory peritoneal dialysis. AB - BACKGROUND: The prevalence of different types of bone disease in chronic renal failure (CRF) has changed significantly during the last decade. The aim of the present study is to evaluate the spectrum of bone disease in children with CRF undergoing continuous ambulatory peritoneal dialysis (CAPD). METHODS: Seventeen children with CRF on CAPD aged 7-20 years were evaluated. All patients had received regular vitamin D and calcium carbonate therapy during the 6 months preceding the bone biopsy. Serum calcium, phosphate, alkaline phosphatase and immunoreactive parathyroid hormone (iPTH) levels were measured and hand X-rays were performed. Transiliac bone biopsies were analyzed for histologic diagnosis. RESULTS: High turnover renal osteodystrophy (ROD) was the most common bone disease, present in eight patients (47%). Five patients (29%) had low turnover bone disease, and four (24%) had mixed ROD. The mean age of the high turnover ROD group was higher than that of the low turnover group (14 +/- 3 vs. 11 +/- 3 years, P < 0.05). Seven of the nine patients who had tubulo-interstitial nephritis were found to have high turnover bone disease. In contrast, none of the patients with glomerulonephritis exhibited high turnover bone lesions. Mean serum calcium levels were found to be significantly higher in the low turnover group compared with the patients with high turnover bone disease (P < 0.001). A serum iPTH level > 200 pg/mL was 100% sensitive and 66% specific in identifying patients with high turnover ROD. CONCLUSION: The spectrum of bone disease of the children with CRF undergoing CAPD seems to depend on the rate of CRF and primary disease. The risk of developing overt hyperparathyroid bone disease is high in children with slowly progressing forms of renal pathology and especially in those with tubulo-interstitial disease. In contrast, children with glomerular diseases who had a more rapidly progressive course may have a lesser risk of developing high turnover bone disease. The results of the present study indicate that even routinely prescribed regular vitamin D therapy early in the course of disease may lead to low turnover bone lesion in small children who have CRF due to rapidly progressive forms of renal pathology. PMID- 10703236 TI - Time-course changes of eosinophil counts in premature infants: no effects of medical manipulation, except erythropoietin treatment, on eosinophilia. AB - In order to determine the factors responsible for eosinophilia during the neonatal period, we counted the eosinophils of premature infants every week and compared the medical profiles of infants with eosinophil counts above the 95th percentile and those below that percentile during the course of study. Medical treatments such as mechanical ventilation, antibiotics administration and intravenous catheterization had no significant effects on the increase of eosinophils. Furthermore, the incidence of eosinophil counts above the 95th percentile was not different between breast-fed and formula-fed infants. The infants treated with erythropoietin had greater eosinophil counts than those with no treatment. It is probable that medical manipulation using foreign bodies such as intratracheal tube, intravenous catheter, antibiotics and artificial formula had no significant effects on the increase of eosinophil counts, except for exogenous erythropoietin. PMID- 10703237 TI - Delay of liver maturation as a cause of transient neonatal galactosemia. AB - BACKGROUND: Because a large amount of serum alpha-fetoprotein (alpha-FP) is synthesized in the liver of the fetus or premature newborn, high concentrations or delayed degradation of serum alpha-FP during the neonatal period may reflect hepatic immaturity. METHODS: In order to evaluate the relationship between transient neonatal galactosemia and delay of liver maturation, the concentration and half-life of serum alpha-FP during the neonatal period were measured in patients with transient galactosemia and in normal neonates. RESULTS: No significant differences were observed in the serum concentration of alpha-FP between normal and galactosemic patients less than 1 month of age. However, the half-life of serum alpha-FP was significantly longer in galactosemic patients between 15 and 60 days of age compared with age-matched normal neonates. CONCLUSION: Based on these results, we hypothesize that delay of liver maturation during the neonatal period, especially during the first 2 months after birth, can be a cause of transient neonatal galactosemia. PMID- 10703238 TI - Late hemorrhagic disease of the newborn. AB - BACKGROUND: Late hemorrhagic disease of the newborn (HDN) may occur without an underlying disorder or as a secondary manifestation of an underlying disorder. It may be seen in fully breast-fed infants without a routine supplementation of vitamin K. In contrast, idiopathic late HDN is defined as HDN without the presence of any risk factor, such as gastroenteritis or use of antibiotics. Severe hemorrhagic symptoms frequently occur. METHODS: Between March 1987 and May 1997, we evaluated 15 infants with idiopathic late HDN, who were diagnosed by detailed history, physical examination and laboratory findings. RESULTS: The age (mean +/- SD) at onset of symptoms was 62.4 +/- 33.9 days. All children were breast-fed infants and were born at term from healthy mothers. The delivery histories were uneventful. There was no history of vitamin K administration at birth. Signs and symptoms of the patients were convulsions (47%), feeding intolerance and poor sucking (47%), irritability (33%) and pallor (20%). In physical examination; there was bulging or full fontanel in 10 patients (67%), diminished or absent neonatal reflexes in nine patients (60%) and ecchymosis in three patients (20%). Before administration of vitamin K, prothrombin time (PT) was 76.1 +/- 43.0 s and partial thromboplastin time (PTT) was 123.4 +/- 68.8 s. Six to 12 h after administration of vitamin K, PT was 15.6 +/- 1.8 s and PTT was 33.4 +/- 1.0 s. Neurologic, gastrointestinal and skin hemorrhagic findings were found in 11 (73%), three (20%) and three patients (20%), respectively. There were both neurologic and skin bleeding symptoms in two patients. The mortality in the present study was 33%. CONCLUSIONS: Late HDN results in severe hemorrhage, especially hemorrhage in the central nervous system. Administration of vitamin K (1 mg, i.m.) at the birth can reduce these severe complications. PMID- 10703239 TI - Nasal mupirocin treatment of pharynx-colonized methicillin resistant Staphylococcus aureus: preliminary study with 10 carrier infants. AB - BACKGROUND: Nosocomial methicillin-resistant Staphylococcus aureus (MRSA) infection in infants has become a serious concern and a new means of preventing the transmission of MRSA in the community needs to be considered. METHODS: We performed nasal mupirocin treatment on 10 infants who were MRSA-positive either in the nose or the pharynx and evaluated the effect of mupirocin on the eradication of MRSA. RESULTS: Eradication of MRSA from the nose was successful in two cases and eradication from the pharynx in six (66.6%) of nine cases. The number of treatments required to achieve eradication varied; within three courses for nose carriers and from one to seven courses for pharynx carriers. Eradication was unsuccessful even after five to seven treatments in three pharynx-limited carriers. CONCLUSIONS: These data suggest that the effect of nasal mupirocin treatment on pharynx-colonized MRSA is limited and that repetitive treatment is necessary in some cases. However, in view of the possibility of preferential pharyngeal colonization of Staphylococcus aureus in infancy, nasal mupirocin treatment deserves further evaluation for eradication not only of nose- but also of pharynx-colonized MRSA. PMID- 10703240 TI - Acute rheumatic fever in Konya, Turkey. AB - BACKGROUND: Patients with acute rheumatic fever (ARF), who were admitted to Pediatric Cardiology Unit of Selcuk University Faculty of Medicine from July 1993 to 1998, were studied retrospectively to verify the clinical profile of the disease and to compare the results with those from other countries. METHODS: All patients were examined by one of the two pediatric cardiologists in our study group. Every patient had a chest X-ray, electrocardiogram and an echocardiographic investigation. Throat culture, antistreptolysin O test, C reactive protein and sedimentation rates were investigated for each patient. RESULTS: During the study period, 274 cases with ARF were identified among patients admitted to the present institution. There were 8032 visits during the study period, giving an occurrence rate of 3.4%. Arthritis was the most common major manifestation (81.4%). It was followed by carditis (60.9%) and chorea (17.9%). Subcutaneous nodules (0.7%) and erythema marginatum (0.4%) were both seen in patients with carditis. The mitral valve was the most commonly affected valve (95.8%), followed by the aortic valve (40.1%). Two patients died and regurgitation disappeared in 21% of patients with mitral regurgitation. Fifteen patients (14%) with isolated arthritis and pure chorea had mitral regurgitation demonstrated by echocardiographic investigation but without any significant murmur. CONCLUSION: The present study indicates that ARF is still a significant problem in Konya and that recurrences can be prevented by administering a 3-week benzathine penicillin G regimen. PMID- 10703241 TI - Psychological characteristics and biofeedback mitigation in preadolescents with eating disorders. AB - BACKGROUND: Anorexia and bulimia nervosa are considered to be the most serious eating disorders in female adolescents, with a multifactorial etiology and severe medical complications. It is interesting to investigate the specific relationships of these diseases to emotional stress, coping mechanisms and biofeedback mitigation. METHODS: The study comprised 76 obese and 27 anorectic girls, as well as 35 healthy girls as a control group. Psychological assessment was performed using the Eysenck Personality Questionnaire, Emotional Profile Index, General Anxiety Scale, Minnesota Multiphasic Personality Inventory and Cornell Medical Index. The therapy was multimodal, including a biofeedback relaxation system based on electrodermal response (EDR). RESULTS: Scores obtained from psychometric instruments, in particular concerning basic anxiety, intrafamily conflicts, self-defense and depression, showed that anorexia nervosa and hyperphagia are specifically stress related. Personality characteristics and the environment, as models for habits, modified the manner of coping with stress differently for anorexia and hyperphagia. The EDR biofeedback was shown to be an effective support for mitigation of eating disorders in preadolescents, with better results for anorectic girls. The correlated difference in personality profiles suggested the use of specific animated sequences. CONCLUSIONS: The results confirmed the hypothesis that obesity and anorexia nervosa could be related to different types of emotional stress and coping mechanisms, and accordingly be differently mediated by biofeedback technique. PMID- 10703242 TI - Fibrinolytic treatment of portal vein thrombosis after umbilical catheterization using systemic urokinase. PMID- 10703243 TI - Further delineation of Char syndrome. PMID- 10703244 TI - Two patients with trisomy 9 mosaicism. PMID- 10703245 TI - Phenylketonuria and cystic fibrosis in the same patient. PMID- 10703246 TI - Henoch-Schonlein purpura nephritis associated with human parvovirus B19 infection. PMID- 10703247 TI - Pseudotumor cerebri manifesting as a symptom of acute promyelocytic leukemia. PMID- 10703248 TI - Assessment of congenital nasal stenosis by shaded surface display reconstruction images. PMID- 10703249 TI - Detection of human herpesvirus 7 DNA in the cerebrospinal fluid of a child with exanthem subitum. PMID- 10703250 TI - Allogeneic CD34-selected peripheral stem cell transplant for acute non lymphocytic leukemia (FAB M5a) developed from juvenile chronic myelogenous leukemia. PMID- 10703251 TI - Survey of the current state of pediatric drug use in Japan (1994-6). Drug Therapy Committee of the Japan Pediatric Society. AB - BACKGROUND: A survey of the current state of pediatric drug therapy in Japan was conducted using a questionnaire completed by committee members and the former president of the Drug Therapy Committee of the Japan Pediatric Society. METHODS: Questionnaires were mailed to the members of the Drug Committee, who represent the various specialties of the Japan Pediatric Society, and to its former chairperson. Questionnaires contained eight items. RESULTS: This survey demonstrated that, although use of drug therapy in adults is clearly defined, pediatric drug therapy in Japan is extremely imprecise, with the potential to lead to undesirable consequences. CONCLUSION: The Japan Pediatric Society demands that the standardization of drug information labels and dosages specifically for children, including premature and full-term neonates, should be included under all aspects of National Health Insurance. PMID- 10703252 TI - The perceptual magnet effect in Australian English vowels. AB - Recent research (Kuhl, 1991) has suggested that the internal structure of vowel categories is graded in terms of stimulus goodness. It has been proposed that a best instance stimulus reflects a central point or prototype, which effectively renders within-category members perceptually more similar. Discrimination experiments suggest a nonlinear relationship between acoustic and perceptual space near category centers (Iverson & Kuhl, 1995b). This phenomenon has been described as the perceptual magnet effect. The present study investigated the presence of the perceptual magnet effect in five Australian vowel categories. Australian English speakers identified, rated, and discriminated between a pool of 32 vowel stimuli that varied in F1 and F2 values. The results from Experiments 1 and 2 showed that subjects were able to judge the quality and identify of each stimulus and that a general grading of stimulus quality was reported. This was not symmetrical, and the subjects' responses varied considerably. In Experiment 3, closer control of the methodology in the discrimination task and of contextual factors influencing the test materials was exercised. Despite this, evidence of the warping of perceptual space in discrimination data was not found. In general, these results do not provide support for the existence of the perceptual magnet effect, and explanations for this finding are discussed. PMID- 10703253 TI - Perceptual parsing of acoustic consequences of velum lowering from information for vowels. AB - Three experiments were designed to investigate how listeners to coarticulated speech use the acoustic speech signal during a vowel to extract information about a forthcoming oral or nasal consonant. A first experiment showed that listeners use evidence of nasalization in a vowel as information for a forthcoming nasal consonant. A second and third experiment attempted to distinguish two accounts of their ability to do so. According to one account, listeners hear nasalization in the vowel as such and use it to predict that a forthcoming nasal consonant is nasal. According to a second, they perceive speech gestures and hear nasalization in the acoustic domain of a vowel as the onset of a nasal consonant. Therefore, they parse nasal information from a vowel and hear the vowel as oral. In Experiment 2, evidence in favor of the parsing hypothesis was found. Experiment 3 showed, however, that parsing is incomplete. PMID- 10703254 TI - Adapting to remapped auditory localization cues: a decision-theory model. AB - This paper describes a model of adaptation to remapped auditory localization cues that is based on previous decision-theory models of psychophysical performance. The present model extends earlier work by explicitly assuming that past experience affects subject perception and by quantifying how training causes subjects' responses to evolve over time. The model makes quantitative predictions of total sensitivity, bias, and resolution for subjects involved in experiments investigating spatial auditory adaptation. One assumption of the model is that subjects cannot adapt to nonlinear rearrangements of localization cues, which is consistent with previous experimental reports in both audition (Shinn-Cunningham, Durlach, & Held, 1998b) and vision (Bedford, 1993). The model assumes that, in spatial adaptation experiments, subjects learn to interpret a continuous internal decision variable differently than normal; they do not learn to associate discrete stimulus-response pairs. This view is consistent with previous analyses of results from experiments investigating adaptation to visual rearrangement, as well as with the McCullough effect in vision (Bedford, 1993, 1995). PMID- 10703255 TI - Evaluation of response methods for the localization of nearby objects. AB - Four response methods for indicating the perceived locations of nearby objects were evaluated: the direct-location (DL) method, where a response pointer is moved directly to the perceived location of the target; the large-head (LH) and small-head (SH) methods, where the pointer is moved to the target location relative to a full-scale or half-scale manikin head; and the verbal report (VR) method, where the spherical coordinates of the target location are indicated verbally. Measurements with a visual target indicated that the DL method was relatively unbiased and considerably more accurate than the other methods, which were all roughly equivalent. Correcting for bias improved accuracy in the LH, SH, and VR responses, but not to the level of the uncorrected DL responses. Replacing the visual target with an acoustic stimulus approximately doubled the errors with the DL response but indicated similar performance in the front and rear hemispheres. The results suggest that DL is the most appropriate response method for close-range localization experiments. PMID- 10703256 TI - Perception of musical tension for nontonal orchestral timbres and its relation to psychoacoustic roughness. AB - Can tension in nontonal music be expressed without dynamic or rhythmic cues? Perceptual theories of tonal harmony predict that psychoacoustic roughness plays an important role in the perception of this tension. We chose a set of orchestrated chords from a nontonal piece and investigated listeners' judgments of musical tension and roughness. Paired comparisons yielded psychophysical scales of tension and roughness. Two experiments established distinct levels of these two attributes across chords. A model simulation reproduced the experimental roughness measures. The results indicate that nontonal tension could be perceived consistently on the basis of timbral differences and that it was correlated with roughness, the correlation being stronger as the perceptual salience of other attributes (such as high-pitched tones or tonal intervals) was reduced. PMID- 10703257 TI - Evaluating frequency proximity in stream segregation. AB - Consecutive sounds of similar structure that are close in frequency or pitch are more likely to be perceived as part of the same sequence than those at greater frequency separations. The principle of grouping into such perceptual sequences, or auditory streams, is known as frequency proximity. However, the metric by which one frequency difference is judged to be greater or less than another in complex auditory scenes is not yet known. Two experiments explored the metric for frequency proximity. We presented repeating three-tone stimulus patterns at a rate where they are normally heard as two streams, one containing the highest tone and one containing the lowest. The middle tone joined one stream or the other depending on its frequency. Subjects reported the perceived allocation of the variable tone by responding on a 5-point scale. The frequency at which either of these two percepts was equally probable was found to be lower than a logarithmic midpoint or the midpoints on a cochlear map or the Mel scale; that is, it was unlike metrics arrived at by direct comparisons of tones. Further, the midpoint for high and low tones presented synchronously was lower than that for the same tones presented sequentially, demonstrating that in addition to a proximity factor, some additional factor or factors must operate differently when the lower and upper fixed tones are, or are not, presented simultaneously. PMID- 10703258 TI - Induction and impairment of saturated yaw and surge vection. AB - A flight simulator was used to investigate the perception of self-motion and visual scene motion during the induction of saturated 10 deg/sec yaw and 50 m/sec surge vection, and during subsequent impairment of saturated vection by inertial motions. The subjects (n = 5) did not perceive any self-acceleration or visual scene deceleration during the induction of saturated vection but perceived a rather sudden change in self-velocity and visual scene velocity. The mean group times to saturated vection were 3.0 sec for yaw and 2.7 sec for surge. Above certain inertial motion amplitudes, the subjects reported additional self-motion from the applied inertial motions while experiencing saturated vection. To impair saturated yaw vection, these amplitudes were 0.6 m/sec2, 0.4 m/sec2, 8 deg/sec2, and 5 deg/sec2, for surge, sway, roll and yaw motions, respectively. To impair saturated surge vection, these amplitudes were 0.6 m/sec2, 0.3 m/sec2, 5 deg/sec2, and 4 deg/sec2, respectively. The results indicate that saturated vection is more robust for translations than for rotations because the rotational inertial amplitudes were closer to the amplitudes at which the applied inertial motion was perceived than the translational inertial amplitudes. PMID- 10703259 TI - Positive and negative compatibility effects. AB - This paper reports a series of four experiments that established a negative compatibility effect (NCE) by which compatible distractors led to slower and less accurate target performance than did incompatible ones (Experiment 1). This effect is interpreted as an early perceptual effect that delays the attribution of visual attention over the target location in the compatible condition. This view predicted that the NCE should be observed only when attention has to be selectively attributed to the target location. In Experiments 2 and 3, this prediction was tested by manipulating the perceptual load in the display. High perceptual load displays are known to require selective attention (Lavie, 1995). Accordingly, reliable NCEs were observed when high-load displays were used. In contrast, reduced NCEs were found in displays that did not require selective attention. Experiment 4 established that the manifestation of the NCE was influenced by low-level visual cues, such as brightness and contrast. Overall, these experiments indicated that the NCE can be understood as an early perceptual effect, which arises from a conflict between the cues that guide the distribution of attention when the task requires selective attention. PMID- 10703260 TI - The psychophysics of imagery. AB - A series of experiments considers the extent to which the interrelations among subjective magnitudes aroused by images corresponds to those for subjective magnitudes aroused by physical stimuli. In Experiment 1, 68 undergraduates typed phrases in response to graded categories regarding the imagined magnitude of lights, sounds, and smells. In Experiment 2, 5 undergraduates and, in Experiment 3, 3 graduate students then magnitude estimated the image intensity aroused by each of these stimulus phrases. In Experiments 4 and 5, the same subjects performed cross-modality matches between phrases arousing images for different attributes (light, sound, and smell). Statistical analysis indicates that estimates based on images display many of the same patterns as those based on physical stimuli. The major exception involves sequence effects, present for actual stimuli but not for images. PMID- 10703261 TI - The Pieron function in the threshold region. AB - The Pieron function (Pieron, 1914, 1920, 1952) describes the decay of reaction time (RT) when the intensity of the stimulus is increased. It is generally demonstrated within a suprathreshold range of intensities. However, in some studies, for the lowest range of intensities, the exponent of the function is clearly greater than that for the upper ranges of intensities. Such an increase in the exponent for the lowest intensities is assumed to result from a combined effect of stimulus intensity and of stimulis uncertainty in detection. Our first experiment used luminance levels that covered all the scotopic range and a spatial two-alternative forced-choice task in which both accuracy and RT were measured. It demonstrated a drastic increase in the exponent in the Pieron function when the intensities reached the threshold region. Since the estimates of the threshold region may have been biased by the use of a much larger range of luminances, a second experiment was conducted using luminances that covered only the threshold region. This experiment confirmed the previous estimates for the threshold region. PMID- 10703262 TI - Constrained scaling: the effect of learned psychophysical scales on idiosyncratic response bias. AB - We report seven experiments in which subjects were trained to respond with numbers to the loudness of 1000-Hz pure tones according to power functions with exponents of 0.60, 0.30, and 0.90. Subjects were then presented with stimuli from other continua (65-Hz pure tones or 565-nm lights varying in amplitude) and were asked to judge the subjective magnitude of these stimuli on the same numerical scale. Stimuli from the training continuum were presented, with feedback, on every other trial in order to maintain the trained scale. Except for the 0.90 scale, subjects readily learned the predetermined scales and were able to use them to judge the non-training stimuli with group results consistent with those usually reported. Also, in contrast to the usual magnitude estimation results, these results produced extremely low levels of intersubject variability. We argue that such learned scales can be used as "rulers" for measuring perceived magnitudes, according to a common unit. PMID- 10703264 TI - Concavities as basic features in visual search: evidence from search asymmetries. AB - Concave cusps and negative curvature minima play an important role in many theories of visual shape perception. Cusps and minima are taken to be part boundaries, used to segment an object into parts. Because of their important role in determining object structure and because there is some evidence that object structure is processed in parallel, it might be expected that concave cusps and negative curvature minima are processed preferentially. We tested this conjecture in several visual search experiments. Visual search for a target with a concave cusp among totally convex distractors yields nearly flat slopes (< 10 msec/item) for both present and absent trials. Reversing the roles of the target and the distractor results in inefficient search. The same asymmetry is found when the concave cusp is replaced by other types of concavity. We conclude, therefore, the concavities can serve as basic features in visual search experiments. This conclusion implies that the unit of selection in a visual search task is an object, rather than a location. PMID- 10703263 TI - The labeled dissimilarity scale: a metric of perceptual dissimilarity. AB - Fundamental to the concept of psychological distance is the idea that confusability allows discovery of the perceptual relationships between objects, which provides understanding of the underlying principles that govern the functioning of a system. Thus, judgments of dissimilarity (conceptually proportional to the inverse of confusability) may provide insight into the elusive underlying quality-coding mechanisms in that sensory system. In the present experiments, a labeled dissimilarity scale (LDS) that reflects the magnitude of odorant dissimilarity was developed in a fashion similar to that reported by Green (Green, Shaffer, & Gilmore, 1993). This scale was produced by rating the perceptual intensity implied by adverbs describing different levels of dissimilarity, and then attaching those descriptors to appropriate locations on a numerical scale. The usefulness of the scale was demonstrated by its ability to produce visual color space with ratings of dissimilarity of Munsell color chips. The stability and reliability of the LDS was evaluated by comparing it with the traditional scaling technique of magnitude estimation (ME). It was found that the scales produced similar ratings of odorant dissimilarity and showed a similar susceptibility to the effects of contrast convergence. However, the coefficients of variation of dissimilarities rated with ME were much higher than those produced with the LDS. The subjects also dealt with the LDS without the anxiety that usually accompanies first-time users of ME. The LDS provided stable ratings of odorant dissimilarity and preserved the inferred ratio scale properties of ME. PMID- 10703265 TI - Attention and spatial selection: electrophysiological evidence for modulation by perceptual load. AB - Behavioral data have suggested that perceptual load can modulate spatial selection by influencing the allocation of attentional resources at perceptual level processing stages (Lavie & Tsal, 1994). To directly test this hypothesis, event-related potentials (ERPs) were recorded for both low- and high-perceptual load targets in a probabilistic spatial cuing paradigm. The results from three experiments showed that, as measured by the lateral occipital P1 and N1 ERP components, the magnitude of spatially selective processing in extrastriate visual cortex increased with perceptual load. Furthermore, these effects on spatial selection were found in the P1 at lower levels of perceptual load than in the N1. The ERP data thus provide direct electrophysiological support for proposals that link perceptual load to early spatial selection in visual processing. However, our findings suggest a relatively broader model--where perceptual load is but one of many factors mediating early selection. PMID- 10703266 TI - Dissociating the effects of featural and conceptual interference on multiple target processing in rapid serial visual presentation. AB - Attentional blink (AB) describes the finding that, when subjects attend to a specified target in a rapidly presented visual stream, they show a decreased ability to process a subsequent probe item for up to 600 msec. In the present study, the roles of featural and conceptual interference in the processing of targets and probes in a rapid serial visual presentation stream were examined. In Experiment 1, featurally more complex T + 1 items produced larger AB even when the physical energy of the stimulus (e.g., the number of pixels) was held constant. In Experiment 2, the conceptual category of the T + 1 item affected target identification but not AB magnitude. These result suggest that featural interference is a major determinant of AB magnitude, whereas featural and conceptual interference both affect target identification. PMID- 10703267 TI - Do spelling variations affect associative and phonological priming by pseudohomophones? AB - A nonword prime can sound like a target word or one of the target's associates, and it can look like either without sounding like either. These pseudohomophones and pseudohomographs can vary in the number of letters shared with the target or its associate. In an associative priming experiment in which targets were named and prime duration was 125 msec within a mask-prime-mask-target sequence, pseudohomophones primed and pseudohomographs did not, with the pseudoassociative priming being only weakly affected by spelling differences. In three further experiments, prime homophony and homography were defined in respect to the target. Prime durations were 125 and 21 msec within a mask-prime-mask-target sequence and 57 msec within a mask-prime-target sequence. The superior priming by pseudohomophones was relatively insensitive to spelling. Results are discussed in terms of the phonological coherence hypothesis and the roles for orthographic information implied by the hypothesis. PMID- 10703269 TI - The importance of the convex hull for human performance on the traveling salesman problem: a comment on MacGregor and Ormerod (1996) AB - MacGregor and Ormerod (1996) have presented results purporting to show that human performance on visually presented traveling salesman problems, as indexed by a measure of response uncertainty, is strongly determined by the number of points in the stimulus array falling inside the convex hull, as distinct from the total number of points. It is argued that this conclusion is artifactually determined by their constrained procedure for stimulus construction, and, even if true, would be limited to arrays with fewer than around 50 points. PMID- 10703268 TI - Response latencies to the onset and offset of visual stimuli. AB - Simple response times (RTs) are known to be slower to the offset of a visual stimulus than to its onset. This is called the onset advantage. In the first of four experiments, we discovered that a spurious onset advantage can be produced by the long persistence of P31 phosphor. In the remaining three experiments, we found that offset RTs were slower only when they were made in a context in which responses to the abrupt onset of some stimuli had to be suppressed. We tested this hypothesis of response suppression in two ways: (1) by mixing regular onset trials with other trials on which a response to an onset had to be suppressed, and (2) by ramping the emergence of "offset" stimuli over time, so that offsets were the only abrupt events in the display. In both cases, we found that the onset advantage depended critically on whether the responses were made in a context of response suppression. We conclude that the onset advantage is mediated not by sensory factors such as visible persistence, but by response programming factors that are strongly affected by contextual events. PMID- 10703270 TI - Fertility and marital rates in first-onset schizophrenia. AB - BACKGROUND: Many studies have demonstrated that in schizophrenia there are decreased rates of marriage, fertility and marital fertility. However, it is not clear whether this finding occurs as a social consequence of having the illness or is an inherent part of the illness. One would expect it to vary across cultures if it were being mediated by social and cultural factors. METHOD: We investigated this by reviewing the marital and fertility data from a multi-ethnic first-contact group of patients in London with CATEGO broadly defined schizophrenia, and comparing this with similar data from a group of controls who were matched for age, gender and ethnicity. RESULTS: Our sample comprised 38 White British, 38 Caribbean and 24 Asian subjects. The Asian group were significantly older (mean age 38, P < 0.003) and had a greater percentage of females (50%). When compared to controls, the White and Caribbean patients, but not the Asians, had decreased marital and stable relationship rates. There was also decreased fertility and marital fertility as evidenced by number of children among the Whites and Caribbeans, but again significantly not among the Asians. Marital status did not predict fertility rates, particularly among the Caribbeans. Regression analysis demonstrated an effect for age on the number of children but not on marital status. CONCLUSIONS: These findings suggest that marital and reproductive behaviour are reduced in schizophrenia, but this effect may be mediated by social and cultural factors and therefore not apply consistently across ethnicity. Further research is needed to prospectively investigate populations to determine whether impairments of this nature are inherent or consequential to the illness. PMID- 10703271 TI - Age at onset of schizophrenia and neuroleptic dosage. AB - BACKGROUND: Lower age at onset of schizophrenia has been traditionally associated with poorer response to treatment and less favourable prognosis. The aim of the study was to find out whether age at onset of schizophrenia is related to the dosage of typical neuroleptics in outpatients. METHOD: Age at onset was defined as age at first seeking of psychiatric help. Demographic, social and disease related characteristics were studied in a group of 200 stable outpatients with schizophrenia (100 males and 100 females). Psychopathological symptoms were assessed with the Krawiecka Scale. Neuroleptic dosage was converted to milligrams of chlorpromazine equivalents and logarithmically transformed to obtain normal distribution. RESULTS: Onset of schizophrenia occurred earlier in males than in females. The average dosage was 251.7 (SD 303.9) mg chlorpromazine equivalents. In a multivariate linear regression model, lower age at onset and higher sum of symptoms were related to the drug dosage. CONCLUSION: The results confirm the findings of other authors that patients with lower age at onset are less responsive to typical neuroleptics. Some of the patients with early onset would be more appropriately treated with atypical neuroleptics, which may have better therapeutic efficacy. PMID- 10703272 TI - Mortality among involuntarily admitted psychiatric patients: a survival analysis. AB - BACKGROUND: Only a few studies have examined mortality among committed psychiatric patients, and most of them suffer methodologically from selected populations, small samples, and inappropriate methods of data analysis. The purpose of this study was to determine whether involuntarily admitted psychiatric inpatients are a high risk group for mortality compared to a group of voluntarily admitted psychiatric inpatients. METHOD: A retrospective cohort design was used with a maximum 9-year variable follow-up. A multistage sampling procedure was used to generate the sample, which consisted of 1064 involuntarily admitted patients and 1078 voluntarily admitted patients. RESULTS: During the follow-up period, 107 deaths were identified, 58 involuntary and 49 voluntary [chi 2 (1) = 0.9255, P = 0.336]. No significant differences were observed between the cohorts when survival analysis was used to examine survival experiences in the community. CONCLUSIONS: Compared to voluntary patients, involuntary patients do not appear to be at a higher risk for mortality. The high standard mortality ratios observed in both cohorts, however, are consistent with previous findings of high mortality among psychiatric patients in general, and support the need for intensive follow up in the community following discharge from a psychiatric inpatient unit. PMID- 10703274 TI - Life events in suicide and undetermined death in south-east Scotland: a case control study using the method of psychological autopsy. AB - BACKGROUND: Adverse life events have been associated with increased risk of suicide. Mental disorders are also major risk factors for suicide. Matching cases and controls for mental disorder is thus appropriate in studies of suicide. This procedure was used to study the degree to which excess adversity was more common in cases who committed suicide as opposed to living controls matched for mental disorder. METHODS: The study formed part of a retrospective case-control comparison of cases of suicide/undetermined death with living controls using psychological autopsy in south-east Scotland. Cases and controls were matched for age, sex and mental disorder. Informants were those closest to cases and controls. Cases and controls were assessed for life events using the Interview for Life Events. The subjects were 45 cases of suicide/undetermined death and 40 living controls. RESULTS: Cases and controls did not differ significantly in severity of mental disorder. Adverse interpersonal events within the family (P = 0.01) with an odds ratio (OR) of 9.0 (95% CI, 1.3-399) and adverse physical health-related events (OR 5.0, 95% CI 1.1-47, P = 0.04) were significantly more common in cases than controls. CONCLUSIONS: Cases had significantly more adverse life events than controls overall. The categories accounting for these differences were interpersonal family adversity and physical ill-health. There were no significant differences in either the number or severity of ongoing difficulties between cases and controls. Recent adverse life events contribute to the increased risk of suicide even when age, sex and mental disorder are controlled for. Future research should examine interactions between social support and acute and chronic adversity. PMID- 10703273 TI - Suicide attempts by psychiatric patients in acute inpatient, long-stay inpatient and community care. AB - BACKGROUND: This study examined rates of and risk factors associated with suicide attempts by psychiatric patients under active care. It was especially focussed on the relative rates across three standard treatment settings: acute inpatient care, long-stay inpatient care and community-based care. METHODS: A total of 12,229 patients in 13,632 episodes of care were rated on the Health of the Nation Outcome Scales (HoNOS) Item 2. For the purposes of the current investigation, a score of 4 was deemed to indicate a suicide attempt. RESULTS: Incidence densities per 1000 episode days were 5.4 (95% CI = 4.8-6.1) for patients under care in acute inpatient settings, 0.6 (95% CI = 0.5-0.8) for patients under care in long stay inpatient settings, and 0.5 (95% CI = 0.5-0.6) for patients under care in community-based arrangements. Predictors varied by treatment setting. Risk was elevated for personality disorders across all settings: 22.7 attempts per 1000 episode days (95% CI = 17.2-30.0) in acute inpatient care; 2.1 (95% CI = 1.0-4.5) in long-stay inpatient care; and 2.3 (95% CI = 1.7-3.0) in community-based care. This effect remained after adjustment for demographics. CONCLUSION: Rates of suicide attempts among psychiatric patients are a major issue facing contemporary mental health care systems, and risk factors vary across different treatment settings. PMID- 10703275 TI - A decade of spontaneous long-term course of psychogenic impairment in a community population sample. AB - BACKGROUND: Our epidemiological study demonstrates the spontaneous long-term course of predominantly psychosocially influenced ("psychogenic") disorders (neurotic spectrum disorders, personality disorders, stress reactions and somatoform disorders) in a representative community sample of the normal adult population of Mannheim, an industrial and university town in Germany. The natural spontaneous course of these disorders in a population sample over a long period remains largely unknown. METHOD: Beginning in 1979 (nt1 = 600) a random population sample was investigated three times over a mean period of approximately 11 years. The last follow-up study ended in 1994 (nt3 = 301). The follow-up sample was representative of the t1 sample. Psychodynamically trained and clinically experienced interviewers used a semi-structured interview and standardized clinical and psychometric instruments. Psychogenic impairment was assessed using a standardized expert rating (Impairment Score, IS). RESULTS: The mean sum-score of psychogenic impairment after 11 years exceeded the value at t1. The case rate (point prevalence, ICD diagnosis + clinical cut-off/IS) increased from 21.6% at t1 to 26.2% at t3 in the investigated follow-up sample. Intra individual correlation of psychogenic impairment between t1 and t3 was high (r = 0.55). We found strong evidence for an unfavorable long-term course of psychogenic impairment and only a weak tendency (23.1%) for spontaneous remission of clinically relevant psychogenic impairment. Within a regression model clinical variables, childhood development conditions and personality traits at t1 predicted psychogenic impairment at t3. CONCLUSION: All clinical variables conclusively indicate an unfavorable spontaneous long-term course of psychogenic impairment. Together with the well-known high prevalence of psychogenic disorders in the normal population, this underlines the need for early therapeutic and preventive intervention. PMID- 10703276 TI - Parent-adolescent agreement on emotional and behavioral problems. AB - BACKGROUND: The aim of the study is to report parent/youth self-report agreement on emotional and behavioral symptoms among 15- to 16-year-old adolescents. METHODS: A completed Child Behavior Checklist and Youth Self-Report were obtained from 580 subjects. RESULTS: Adolescents reported significantly more problem behaviors than their parents. Adolescent girls reported a significantly higher level of distress than boys in most symptom domains. The discrepancies between parent reports and youth self-reports were greater, especially for internalizing symptoms, for girls than for boys. CONCLUSIONS: Many adolescents in need of psychiatric assessment do not receive appropriate help because their problems remain unnoticed by adults. Internalizing problems among girls seem especially likely to remain unrecognized by adults. PMID- 10703277 TI - [Forensic medical evaluation of flaws in medical care rendered at public health institutions in the Russian Federation]. AB - The authors analyze the flaws in medical care rendered at various public health institutions by the results of 2436 expert committee conclusions, verified at bureaus of forensic medical expert evaluations. The essence and causes of these defects at the hospital and prehospital stages of medical care, rendered by physicians of different specializations and nurses, are analyzed. PMID- 10703278 TI - [Diagnostic role of acute microscopic changes in myocardium]. AB - Forensic medical diagnosis of death from coronary heart disease, acute ethanol poisoning, alcoholic cardiomyopathy, closed cardiac injuries, mechanical injuries incompatible with life which may be directly caused by acute cardiac failure, requires identification and evaluation of diagnostic complexes of acute myocardial changes. The diagnostic significance of such complexes of myocardial changes is characterized for the first time. A method for evaluation of such changes, addressed to expert histologists, is presented. PMID- 10703279 TI - [Pathogenesis of Vishnevskii's spots in death from excessive cooling]. AB - Histochemical successions of the gastric mucosa reaction are described, their role and the impact of the kallikrein-kinin system are evaluated, and pathophysiological aspects of development of Vishnevsky's spots as a pathognomonic sign of death from excessive cooling are described. Analysis of 34 deaths from excessive cooling helped determine the correlation between the size, volume, and depth of the gastric mucosa involvement, on the one hand, and duration of periods of cooling and freezing, on the other. The technique of expert evaluation of Vishnevsky's spots is described. PMID- 10703280 TI - [Computer analysis of face and skull size by "quantitative verbal portrait" method]. AB - The aim of this study was to develop a "quantitative verbal portrait" method on the basis of universal measured signs of the face and skull. The database includes 90 cases with expert evaluation of photocompatibility with proven identity. Four groups of signs were investigated: DX and DY--the width and height between accurately fixed concrete craniometrical (facial) points and the so called RX and RY indices--the ratio between distances within the framework of the same direction. The proposed scheme of evaluation helps compare the photograph of a live human and the skull. For evaluating the degree of identity, one should have a summary characterization of all selected signs for the skull and compare it with the sum on the photograph; sometimes it is possible to rule out the identity of the photograph and the skull by the absolute size, e.g., a very large skull cannot belong to a human with a very little face. The score for each group of signs is used for analysis of the information for detecting the similarities. Accumulation and comparative analysis of two information flows are possible: database on portraits of lost subjects and database on graphic images of the skulls of unidentified corpses. PMID- 10703281 TI - [Participation of forensic medics in anthropological examinations of the Orthodox church burials]. AB - Examinations of the rudiments of the Russian Orthodox Saints are reviewed. The burial of holy people in Russia is described. The results of examinations of rudiments of the Saints, Reverend Iosaph of Belgorod, Cornilai Padansky, and Arsenii Konevsky, are presented. A detailed algorithm of expert steps is suggested. PMID- 10703283 TI - [Use of Adobe Photoshop software in medical criminology]. AB - Describes the method of comparative analysis of various objects in practical medical criminology and making of high-quality photographs with the use of Adobe Photoshop software. Options of the software needed for expert evaluations are enumerated. PMID- 10703282 TI - [Chemical toxicological analysis of urinary cannabinoids by chromatography mass spectrometry]. AB - A chromato-mass-spectrometric method for identification of cannabinoids and 9 carboxy-11-nor-delta 9-tetrahydrocannabinol by their methyl derivatives is proposed. Use of alkaline hydrolysis in pretreatment of samples ensures the optimal conditions for complete isolation of 9-carboxy-11-nor-delta 9 tetrahydrocannabinol. Derivation with methyl iodide in absolute acetone in the presence of potassium carbonate was used. Preliminary conclusion on the presence of cannabinoids in the urine can be made from the results of enzyme immunoassay. PMID- 10703284 TI - [Discrimination between functions of an expert bureau and a burial firm (topic for discussion)]. AB - The authors suggest measures aimed at discrimination of the functions performed by forensic medical experts and the services rendered by burial firms. The responsibilities of pathology departments of hospitals and bureaus of forensic medical expert evaluations, which should be reflected in official documents, should be confined to manipulations needed to solve the professional problems and removal of sections. Cases when utilization of certain chemicals or manipulations aimed at restoration of the body are prohibited should be listed in detail. The order of transfer of the body from pathology department or forensic medical bureau to the burial firm and to relatives should be regulated with consideration for the results of autopsy. The moment of the end of autopsy studies is to be determined. PMID- 10703285 TI - [Forensic medical service in Brazil and exemplified by San Paolo state]. PMID- 10703286 TI - The occupational health nurse as case manager. PMID- 10703287 TI - Evaluating case management services for injured workers. Use of a quality assessment model. AB - An increasing number of employers and third party administrators are choosing case management as a strategy to coordinate services for workers who sustain an occupational injury or illness. The successful delivery of case management services requires the service provider possess special skills and knowledge. Occupational health nurses are uniquely qualified to provide case management services to injured and ill workers. The effectiveness of case management services is generally described in terms of costs and quality of services; thus, determining the effectiveness of a case management program requires an evaluation of costs and service quality. PMID- 10703288 TI - Injured workers' perceptions of case management services. A descriptive study. AB - This article describes the findings from a study of injured workers conducted as part of a multifaceted evaluation study of a case management program. The sample consisted of workers who filed a workers' compensation claim between January 1 and September 30, 1995. Data collection consisted of written surveys (n = 45), personal interviews (n = 27), and telephone interviews (n = 16). The findings from this study provided many insights into the injured workers' personal and work experiences, and, in particular, their perceptions of their experience with the nurse case management program. Workers satisfied with services described the nurse case manager (NCM) as having the ability to see the "big picture," to develop appropriate goals, and to anticipate client needs. Dissatisfied workers reported feeling unimportant in terms of service provision. They reported feeling that "the system" did not respond to their needs, and that the NCM was uninterested and disrespectful. This vivid portrayal of workers' experiences and perceptions of case management services provides valuable information about the world view of the injured worker. PMID- 10703289 TI - Return to work experience of injured workers in a case management program. AB - A major goal of case management programs is the worker's timely return to work. Few studies have examined return to work from the perspective of the injured worker. This article describes the findings from the case management evaluation that describe the return to work experience of workers who sustained catastrophic injuries, or who had secondary conditions or complications following the injury occurrence. Among the factors determined to affect the return to work experience were structural factors (i.e., psychosocial variables including job satisfaction and relationship with employer and coworkers, financial pressures, and system issues such as securing benefits) and process factors (i.e., interaction with service providers and with the workers' compensation system). Outcomes are described in terms of satisfaction with services and return to work. PMID- 10703290 TI - Occupational exposure to Mycobacterium tuberculosis. Legal issues in workers' compensation. AB - Occupational exposure to TB remains a significant threat in select high risk occupations despite 5 years of declining disease incidence rates in the United States. TB kills more people on a global scale than any other infectious disease. One third of the global population is currently infected with TB. Workers' compensation insurance may be inadequate to cover lost wages and medical bills in cases of occupational exposure to TB if the source patient is unknown. There is a need to reform state laws for workers' compensation so TB infections in high risk employees are presumed to be work related unless a community exposure to the disease is identified. PMID- 10703291 TI - Business writing: a necessary skill for occupational and environmental health nurses. PMID- 10703292 TI - Comprehensive education plan for a discrete stroke population: needs, considerations, and gaps. AB - The patient population which has the most to gain from comprehensive stroke education is the one which has experienced a stroke with minimal deficit and carries an increased risk for experiencing a second more severe stroke. This is the patient population the author selected, yet little published information is available. Given that in Canada stroke carries the third highest rate of mortality and the highest risk of morbidity there is a role for an advanced practice nurse to develop and deliver a comprehensive stroke education program for this population. This is of special significance given the understanding that stroke is considered to be one of the most preventable diseases. The practice environment for this patient population is shifting out of the hospital and into the community. The learning needs and issues as well as the identification of appropriate patient focussed outcomes needs to be examined. A comprehensive plan for the delivery of patient focussed education which includes content, setting, and timing for this target population needs to be developed, piloted and reported on. Nurses have been identified as appropriate for delivering patient education. An advanced practice nurse is poised to address this need and develop a model for others to incorporate in their practice. PMID- 10703293 TI - Continuous intrathecal baclofen infusions. An introduction and overview. AB - Continuous intrathecal baclofen infusion (CIBI) is a relatively new treatment modality for severe spasticity of spinal cord origin. Literature review suggests relief of severe spasms and rigidity is proven with CIBI, in patients with spinal cord injury and multiple sclerosis, while ongoing research exists for patients with acquired brain injury and cerebral palsy. Criteria for patient selection, the screening trial process, an outline of the surgical procedure, and generalities of maintenance therapies, will be reviewed broadly as per literature, as well as specifically to the Vancouver experience with adults. Additionally, reported patient outcomes and implications for nursing will be shared. PMID- 10703294 TI - Grant reviews: how to do them well. PMID- 10703295 TI - Oncology nursing education within a supportive care framework: an evidence-based undergraduate course. AB - Addressing supportive care needs of individuals affected by cancer is a crucial role for oncology nurses. If these needs are not identified and addressed, then individuals and their families risk experiencing biopsychosocial distress. This paper describes how a group of interested nurses with research and/or cancer knowledge developed, implemented and evaluated an evidence-based university undergraduate course in oncology nursing. The purposes of this course are to provide nurses with specialized knowledge about supportive care in oncology throughout the cancer care continuum and to assist registered nurses in preparing for oncology nursing certification. Theoretical, practical and research-based issues related to the scope of supportive care are incorporated into the assessment, nursing diagnosis, planning and evaluation of client care. PMID- 10703296 TI - Humanistic reflections of a research nurse in a longitudinal study: a personal essay. AB - The purpose of this paper, first, is to share my 'lived experience' as a research nurse, in a longitudinal survey design study, who followed lung cancer patients and their family caregivers throughout the illness duration. I embraced an overarching 'humanistic' philosophy to 'come to terms' with my intra- and interpersonal reactions arising from my role as a human research tool. These reactions were best captured by Paterson and Zderad's (1976) humanistic concepts of authenticity, vulnerability, relating, disclosure and enclosure, and presence. Second, I will describe how I dealt with these concepts within the context of a research nurse role. PMID- 10703297 TI - Practical support for women with breast cancer. AB - Although emotional, social and informational support are well recognized needs among women with breast cancer, practical support has not attracted serious research attention. However, consumer advocates and researchers studying the subjective experience of breast cancer note that practical support may be a particular concern for some women. In order to document some aspects of the practical support issue and begin to render this element of the experience visible, a community-based consumer group conducted a preliminary survey of practical support needs of women who were living with or had experienced breast cancer. Among the 100 women who responded to this survey, there was considerable evidence that practical support may well remain an unmet need in the current health care climate. PMID- 10703298 TI - High dose rate brachytherapy endobronchial treatments: potential equipment problems and emergencies. PMID- 10703299 TI - The ultimate one-stop for cancer patients with bone metastases: new combined bone metastases clinic. PMID- 10703300 TI - Health care, nurses and other motherhood issues. PMID- 10703301 TI - Reviewing a manuscript for publication: how do I do this? PMID- 10703302 TI - The point of no return: beyond sexual functioning to sexual health assessment in oncology nursing. AB - Disseminating research findings in a meaningful way is often a challenge. The topic of sexual health creates an even greater challenge. The work that numerous teams conducted within our research department had profound effects that altered the way in which we were practising cancer care. The Schering Lectureship provided the forum for our patients' collective voices to be heard. Artists in the world of music who crossed the spectrum from classical to country helped us to focus. The sexual health message was entwined in their words. That message became a personal interpretation we had not been able to convey with words alone. Through the songs, each nurse found his/her own meaning. To our surprise, this lectureship became a powerful medium for patients to speak out and nurses to learn. The nurses conducting the sexual health studies thank CANO and Schering Canada. To each oncology nurse who spoke about the message heard, please know that your words touched our hearts. While it has been a challenge to put this lectureship into a manuscript, it is hoped the written words will convey a similar message. PMID- 10703303 TI - Descriptive study to compare patient recall of information: nurse-taught versus video supplement. AB - An important goal in oncology nursing is to provide outpatients receiving chemotherapy with adequate information about their treatment so they will be able to cope with treatment reactions and make appropriate decisions about seeking early medical attention when potentially serious side-effects occur. The purpose of the present study was to evaluate patient teaching strategies at one cancer centre. A comparative descriptive study design was employed. A group of patients receiving one-to-one nurse/patient teaching was compared to a group of patients receiving one-to-one nurse/patient teaching plus a take-home instructional chemotherapy video. The patient groups were compared with respect to: a) level of recall of chemotherapy information; b) the sources of information used; and c) preferred information sources. When the mean scores achieved on the chemotherapy knowledge questionnaire were compared, no statistically significant differences were found between the two groups. In fact, both groups showed a "high" level of information recall. Both patient groups reported using a variety of information sources to learn about their chemotherapy, however, for both groups the preferred sources of information were their direct health care providers. The results of the study raise interesting issues about the feasibility of developing "high tech" patient education strategies. PMID- 10703304 TI - Nursing challenges in cancer pain assessment and management: setting an agenda for the CANO Pain Initiative. AB - The purpose of this inventory was to determine what oncology nurses perceived to be the challenges and priorities in pain assessment and management in Canada. It was recognised that there are a variety of practices in Canada, and if we are to establish an educational framework, there is a need to know what some of the issues are and what assessment tools are being used. Sixty-six survey questionnaires were sent to centres with an oncology or palliative care focus. Thirty-one surveys were returned with a great deal of information. The findings of this descriptive inventory of cancer pain assessment and management hold several key implications for future directions of the CANO Pain Initiative. The findings provide further support for ongoing efforts to educate and inform health professionals about the nature of cancer pain and associated assessment and management issues. A number of other issues have been identified with this survey which continues to support the efforts of the CANO Pain Initiative in developing educational modules, comprehensive care plans and practical ways to document pain interventions. PMID- 10703305 TI - High dose rate brachytherapy endobronchial treatments: adjunctive medications and discharge instructions. PMID- 10703307 TI - Palliative care nursing education: "in the classroom" and "from a distance". PMID- 10703306 TI - Developing a peripherally inserted central catheter service with registered nurses. PMID- 10703308 TI - Politics and power: nursing in Canada. PMID- 10703309 TI - Sailing into the millennium: new waters, new realities. PMID- 10703310 TI - Breast cancer: so much more than just a perioperative experience. AB - This paper's focus is the various types of breast cancer, the predisposing factors for the disease, treatment options, treatment modalities, conventional and unconventional therapies, surgical reconstruction, genetics and hormone replacement. The treatment of breast cancer is regarded as a team effort requiring the dedication and expertise of all of the members of the care team to ensure a favorable patient outcome. Within this team the author recommends an expanded new role for the perioperative nurse, that of the nurse navigator. PMID- 10703311 TI - Coral from Australian reef used to replace bone. PMID- 10703312 TI - ORNAC approved post basic OR education programs. PMID- 10703313 TI - Endovascular insitu saphenous vein bypass. AB - First endovascular insitu saphenous vein bypass procedure performed in Canada was in November 1998 at North York General Branson Division. This article describes the pathophysiology of lower limb occlusion, the perioperative nursing care and the endovascular approach to insitu saphenous vein bypass graft. PMID- 10703314 TI - Sterile O.R. repack: collaboration in action. PMID- 10703315 TI - Natural rubber latex allergy update. "Don't let latex irritate you". PMID- 10703316 TI - Poco-a-Poco: an OR nurse's experience of a lifetime. PMID- 10703317 TI - Hand washing awareness: a community health initiative. AB - Handwashing, so simple but forgotten in this busy, hectic, modern world. As perioperative nurses, handwashing is part of our everyday routine. The Ottawa Regional Operating Room Nurses Association brought this important message to the community through our "Back to Basics Health Awareness" Campaign described herein. PMID- 10703318 TI - I know handwashing is important, but... PMID- 10703319 TI - Conquering your mountain. PMID- 10703320 TI - Extravasation of infusate via implanted ports: two case studies. AB - Over the past two decades, implanted ports have become widely used infusion therapy devices. Although these devices have revolutionized the care of patients with cancer and are used routinely to administer various treatments, complications still can occur. Nurses must be vigilant in identifying potential and actual port-related problems and aware that radiological studies may not immediately reveal a problem. Two unique case studies are described in which extravasation complications occurred despite negative initial catheter dye studies. A clinical algorithm is presented that outlines the management of a suspected port extravasation. PMID- 10703321 TI - Recognizing and managing anemia. AB - Anemia is not an uncommon problem. Knowledge of a few common laboratory tests can be helpful when caring for patients with anemia. By reviewing approaches to classifying, evaluating, and managing patients with common anemias, nurses will be prepared to care for patients with an anemia more confidently. PMID- 10703322 TI - Urinary continence issues in oncology. AB - Urinary incontinence may occur with many types of cancer and cancer therapies. Symptoms of urinary incontinence negatively may affect patients' lifestyles, self esteem, and quality of life. Although urinary incontinence is not a new problem, nurses often underestimate its impact on patients. Knowledge of the physiology of micturition and causes of urinary incontinence in patients with cancer may help nurses identify patients at risk and provide a framework for conducting targeted assessments of urinary function. PMID- 10703323 TI - Nursing management of soft tissue sarcomas of the extremities. AB - Soft tissue sarcomas of the extremities are uncommon malignancies that require combined modality treatment. Treatments include extensive surgical procedures, usually radiation therapy, and, in some cases, chemotherapy. Nursing care requires an understanding of surgical and reconstructive procedures, brachytherapy implants, radiation safety regulations, and chemotherapeutic agents. The combination of extensive surgery and radiation has greatly reduced the need for amputation, but the complexity of the treatments requires skilled and knowledgeable nursing care. PMID- 10703324 TI - Care for the caregiver: strategies for avoiding "compassion fatigue". PMID- 10703325 TI - Port access and placement technique issues. PMID- 10703326 TI - SIADH. PMID- 10703327 TI - Cancer pain management. PMID- 10703328 TI - Fluorouracil. PMID- 10703329 TI - Gift from the East ... Chinese medicine. PMID- 10703330 TI - The Mini Mental State Examination (MMSE). PMID- 10703331 TI - Preventing workplace violence: looking at your hiring and termination processes. PMID- 10703332 TI - "Nursing component" of Medicare skilled nursing facility payment spurs lobbying by nursing home industry and strong concerns of nursing and resident advocacy groups. PMID- 10703333 TI - Pain on hold! PMID- 10703334 TI - Liability, UAPs and you. AB - Supervising unlicensed employees increases your malpractice risks. To protect yourself, learn what courts and juries have had to say when patients sue. PMID- 10703335 TI - Dealing with emotional outbreaks of Alzheimer's clients on a special unit. AB - Staff who work exclusively on Alzheimer's disease units need to develop strategies to deal with emotional outbreaks of these clients. This project demonstrated that staff is not helpless in dealing with Alzheimer clients, and it reinforced the staff by validating that they, too, are experts in coping with daily problems of Alzheimer's clients. Another asset was the inclusion of the staff in this study. It was a learning experience for all to see the completion of this work. PMID- 10703336 TI - Homeopathy. PMID- 10703337 TI - Magnet therapy. PMID- 10703338 TI - So you want to be a Director of Nursing ... now what! AB - The authors believe that even an experienced DON might need at least two months to adequately adjust and/or professionally realign problematic areas of concern in a new position. This opinion is in direct contrast with Maun (1998). Maun states "If a DON is moving to this job from another similar position, the overall program may take a week, not including any other corporate or facility specific orientations. If a person is taking a DON job for the first time, the process could take two weeks to allow for identifying areas of deficiency." The position of Director of Nursing is dynamic and fluid; it is challenging and exciting. The key to success is communication, forward planning, flexibility and the willingness to grow with the demanding changes in the arena of long term care. PMID- 10703339 TI - Advance directives. PMID- 10703340 TI - HCFA teleconference satellite broadcast. "Overview of the new nursing home initiatives". April 19, 1999. PMID- 10703341 TI - Laughter: Rx for healthcare professionals. PMID- 10703342 TI - Aromatherapy. PMID- 10703343 TI - The Geriatric Depression Scale (GDS). PMID- 10703344 TI - The evolving role of certified nursing assistants in long-term care facilities. 3. PMID- 10703345 TI - Poor staffing--poor care nursing organizations need to consider taking action. PMID- 10703346 TI - Examining economic improvements in managing diabetes in the nursing home. AB - In 1997, two-thirds of the medical cost of diabetes was borne by the elderly and nursing home care attributable to diabetes accounted for a third of that financial burden. The development of the Insulin Delivery Pen system can provide cost efficiency, and concurrently reduce the potential for contamination, free up nursing time, improve the administration process, and maintain dosing accuracy. The insulin pen delivery system (vs. the traditional vial and syringe method) is an attractive and cost effective option in the treatment of diabetes mellitus for residents in nursing facilities. PMID- 10703347 TI - Cancer and the elderly. PMID- 10703348 TI - The marriage: geriatrics and oncology. AB - Cancer always has been a disease of greater incidence and morbidity in the elderly, and sociodemographic trends predict this fact to escalate in the decades ahead. Optimum care of older adults with cancer can be enhanced through the marriage of two disciplines that have matured in recent years: geriatrics and oncology. This article reviews issues of cancer care for the elderly, including the strengths and challenges of this special population, quality of life considerations, and future directions. PMID- 10703349 TI - Pain, cancer, and older adults. AB - Pain is the most common symptom associated with cancer. Despite the fact that appropriate pain management exists, cancer pain often is inadequately controlled. Sixty percent to 85% of individuals with advanced cancer have either severe or chronic pain. Issues associated with cancer pain and pain management in later life include the effect of pain on the person, friends, and family; the influence of hope; the perception that older adults are less sensitive to painful stimuli; the special problems of drug management in older adults; and the unique issues of older women affected by cancer. This article concludes with implications for nurses. Future research needs to be directed to pain management at home, the ethics of pain management, pain in the cognitively impaired person, and the specific needs of older women. PMID- 10703350 TI - Assessment and management of cancer pain in the cognitively impaired elderly. AB - Cancer pain in the cognitively impaired elderly is a challenging problem for clinicians. Nursing and medical literature on the subject is sparse because the problem has not been adequately studied. This article presents strategies for assessment and management when the older adult has cancer and cognitive impairment. Traditional, pharmacologic, and alternative therapies are reviewed. Common etiologies for pain and confusion also are presented. PMID- 10703351 TI - Three good reasons to see a dentist BEFORE cancer treatment. PMID- 10703352 TI - Older women and mammography: factors influencing their attitudes. AB - Breast disease is a leading factor in the morbidity of older women. Unfortunately, this population segment receives less education on breast screening methods and has a lower rate of mammography use than others. This article focuses on the misconceptions concerning elderly breast care and the reasons why mammography use is lowest in this population segment. PMID- 10703353 TI - Promotion of advance care planning in the nonhospitalized elderly. AB - Factors that influence the execution or lack of execution of advance directives in 162 nonhospitalized older adults were examined. Significant results were found, and implications for geriatric nurses are presented. PMID- 10703354 TI - HIV & AIDS in older adults. AB - Many Americans mistakenly believe that older adults are not at risk for HIV/AIDS. Older people do not perceive themselves to be at risk for HIV infection, either. In reality, approximately 10% of AIDS cases are among people older than 50. Many health care providers lack an awareness of the risk of HIV/AIDS in the elderly population, and as a result, many older people with these conditions are misdiagnosed with other ailments. Major manifestations of HIV/AIDS in elderly adults include Pneumocystis carinii pneumonia, herpes zoster, tuberculosis, cytomegalovirus, oral thrush, Mycobacterium avium complex, and HIV dementia. Elderly HIV-positive women have special health concerns, such as cervical cancer. Nurses and nurse practitioners can heighten their colleagues' awareness of the existence of HIV/AIDS in the elderly population and educate their older patients on HIV/AIDS. Furthermore, information about sexuality and sexual practices of older adults should be incorporated into all health science curricula. Additional research is needed to determine the extent of the problem and how health care providers can best serve their older patients' needs. PMID- 10703355 TI - Pain and quality of life of older adults: Betty R. Ferrell, PhD, FAAN. Interview by Ann Schmidt Luggen. PMID- 10703356 TI - Essentials of home-based cancer care of the elderly. PMID- 10703357 TI - Cancer treatment compromises nutrition. PMID- 10703358 TI - ANA leads effort to protect nurses and patients. PMID- 10703359 TI - Mentoring in the academic setting. PMID- 10703360 TI - Tips to surviving nursing school. PMID- 10703361 TI - Resource books for nursing students. PMID- 10703362 TI - Rural family practice and the nurse practitioner. PMID- 10703363 TI - Being a nurse attorney. PMID- 10703364 TI - Research careers in nursing. PMID- 10703365 TI - A late bloomer gets her second chance. PMID- 10703366 TI - Care management in a managed care world. PMID- 10703367 TI - Saving your career in the 21st century. AB - This article addresses the fundamental and dramatic changes that case managers must undergo internally to keep pace with a rapidly and radically changing work environment as we move into the next millennium. The work paradigm is transforming from a model of well-defined job descriptions and clearly articulated career ladders within organizations to a fluid workforce in which individuals must now view themselves as a mobile portfolio of skills responding to particular needs within organizations Hence, case managers must retool their thinking, unlearn old beliefs that hinder success, and learn to manage their careers as microbusinesses within their organizations. This new model is founded on self-responsibility, entrepreneurial aptitude, vision and personal empowerment. Taking charge of one's career and consciously directing it is a dramatic departure from the norm for most individuals. There is a widely held tendency in our culture to define ourselves by our job titles. This is both an antiquated and myopic view that needs to be discarded to succeed in the future. Health care is a dynamic and evolving industry that requires forward-thinking, flexible, solutions-oriented people. The time is upon case managers to undergo a personal renaissance to artfully position themselves for success in the next millennium. PMID- 10703368 TI - The clinical case management of clients with major depression. AB - In the field of mental health, clinical case management represents a set of interventions that feature ongoing support to persons with chronic mental disorders. In this paper a specific case management approach to working with persons having major depression is outlined. The causes of depression may be primarily biological, but psychological and social conditions also have impact on the course of the disorder. The clinical case management approach includes a variety of interventions which attend to both counseling and environmental work and accommodate the changing severity of the client's depression over time. The interventions described include ego support, crisis intervention, interpersonal therapy, and cognitive-behavioral techniques. Case examples are included as illustrations. PMID- 10703369 TI - Case management of the HIV/AIDS client. AB - One of the most difficult issues a case manager has to deal with is how to stay knowledgeable and current in the face of rapidly changing medical advances. As treatment protocols for diseases change, so must the case management plans for clients. In the case of HIV/AIDS, this is especially true. This article examines the many opportunities case managers have to make an impact on clients' lives and long-term outcomes. As case managers increase their knowledge of disease processes and treatment protocols, they assist their clients in becoming empowered and becoming partners in their treatment plans, at the same time showing them how they can once again gain control of their lives. PMID- 10703370 TI - Caregiver well-being: a strengths-based case management approach. AB - Family caregiving creates challenges and gains for professionals and family members involved in the caregiving arrangement. Caregiver well-being, in particular, is a complex and multidimensional concept for case managers as they engage in assessment, measurement and intervention planning. This article describes a blending of concepts--case management, strengths model and caregiver well-being--and presents applications of this integrated practice framework. The Caregiver Well-Being Scale is discussed as a tool for use in case management with elders and their family caregivers. Case scenarios derived from a strengths-based practice perspective are presented as examples of ways in which the Scale can be integrated into case management practice. Implications for programmatic use are also highlighted. PMID- 10703371 TI - One-year outcomes of older adults referred for aging and mental health services by community gatekeepers. AB - One-year outcomes of older adults referred for community aging and mental health services through the Gatekeeper Model were examined in this study. Outcomes included level of social, physical, psychological, and economic isolation, physical health problems, service need, and service utilization. Findings indicate that individuals referred by gatekeepers were more likely to live alone and to be socially isolated but less likely to have physical health problems. They were also less likely to have a physician at referral, but at 1 year this difference was not found. Cognitive problems had a significant impact on the lives of clients referred by gatekeepers at referral but not after one year. At referral, those referred by gatekeepers had greater service needs, but after 1 year they did not use more services than those referred by other sources. Implications of these findings are discussed. The findings from this study indicate that the adoption of the Gatekeepers model does not result in high service utilization. The Gatekeeper model is inexpensive to implement and can benefit communities through increased collaboration among service providers. PMID- 10703372 TI - Research directions for case management. AB - Case management is a worldwide phenomenon and depending on the particular country's organizational framework of the health and social system, many models have been developed. The definitions of case management vary, but all of the models being developed aim to provide holistic quality care that is cost effective. Reviewing the various components of case management is important because they contribute to the success or failure of providing quality, cost effective care. The geographical setting where case managers work (i.e., hospitals or community/home), and the level of fiscal authority given to the case managers are two important components of case management models. Whether there is a conflict between case managers acting as advocates of clients while at the same time being the gatekeeper of funds is an area that needs exploration. The question is can the two notions be "married" as opposed to being inherently conflicting, and can case managers act in the best interests of the clients. There are many research directions for case management, including which aspects of the model ultimately have positive results for the client and health and social system. PMID- 10703373 TI - Varied perspectives of case management. PMID- 10703374 TI - Developing standards and quality measurements for case management practice. AB - Case management is a service offered in most health care and social service settings, but a universal definition and standards for practice, applicable to the various case management models, do not exist. Without nationally accepted standards and quality measures for community-based long-term-care case management, many agencies are scrambling to create them in order to justify their services to payers. This article describes the method followed by one county agency serving low-income elders and people with disabilities to create process standards and quality measures for case management practice. The project involved experienced case management staff who created measurement tools encompassing measures of 12 case management activities. Their goal was to justify the importance of the service being offered, to set minimum practice standards, and to raise the awareness and level of quality of case management practice. PMID- 10703375 TI - Tracking case management accountability: a systems approach. AB - A review of the evaluation of case management programs in the last two decades reveals a lack of consistency in showing outcome effectiveness. In addition, program evaluation models do not reflect the most prevalent theoretical foundation of case management practice, systems theory. It was the purpose of this article to conceptually establish an evaluation model that reflects systems theory in case management practice, to develop evaluation instruments congruent to collecting data in this model, and then to field-test this evaluation model. PMID- 10703376 TI - Care for the case manager: balancing your wheel of life. AB - The case manager's role in our complex health care system is demanding and draining without some self-reflective attention. The Wheel of Life is a key tool for individuals to assess how well they are leading a fully balanced life. The eight aspects of a balanced life--values, self-care, work, relationships, leisure, relaxation, exercise, and centering--are explained and discussed. A self reflective activity is presented that encourages readers to assess their current life balance. This focused clarification of personal and professional life will facilitate a more fully balanced life with rewards for case managers as individuals, and for their family, clients, and the health care organization. PMID- 10703377 TI - The importance of the case management approach: perceptions of multidisciplinary team members. AB - This article attempts to determine the importance of the case management approach as perceived by health professionals working in multidisciplinary teams. The case management approach is reported to streamline care and contain cost. The literature calls for continuing education in a multidisciplinary forum for all health professions; however, data on perceived or actual educational needs is scant. One hundred forty-one health practitioners working in clinical teams rated four case management components on 100 mm scales: assessing patient needs; educating caregivers; community agency liaison; and, cost monitoring. Statistical significance of differences was determined by Mann Whitney U and Wilcoxon rank sum testing. All scores were high, and of all variables tested, varied only by profession (p < .01). Administrators and nurses had relatively higher scores than physicians and physiotherapists. Patient needs were valued above education, education above liaison, and liaison above cost monitoring (p < .01, < .01, < .01, respectively). Continuing education providers should note that health professionals value case management and may be receptive to education. Further study is required to design educational modules for multidisciplinary use, test impact on knowledge and skills, and determine if transdisciplinary case management education can improve quality of care while containing costs. PMID- 10703378 TI - Embracing the continuum of care: an Australian private hospital's experience. AB - Continuity of care throughout the home, community, and hospital settings is essential in providing quality health care. A continuum-of-care model assists in improving communication between all stakeholders, decreases confusion, and ensures appropriate provision of resources so that patient/client care needs are met. Historically, continuity of care has been difficult to provide because care delivery has taken place in separate settings. In addition, appropriate processes, incentives and resources have not been in place to ensure sufficient interaction between care providers. In Australia the concept of case management has been inviting because it acknowledges the importance of continuity of care. It also addresses pressures on the health care system to ensure quality, cost effective service provision. The extent to which continuity of care is provided depends on the flexibility of the case management model including the flexibility of care providers to interact between settings. This article presents an Australian private hospital's experience utilizing a case management model. Three specialties (Home Care, Oncology, and Medical) will be used as examples demonstrating how the model incorporates the continuum of care concept. Their challenges in providing continuity of care beyond the hospital walls are explored. PMID- 10703379 TI - Case management changes and challenges. PMID- 10703380 TI - Case management from urban and suburban perspectives. AB - The purpose of this research, commissioned by an Area Agency on Aging in Pennsylvania, is to identify the factors that impact on the process of case management in urban and suburban settings. Random samples of clients receiving the same service in an urban and a suburban area were compared as well as direct observations made of case managers in these same locations. The results indicate that clients in the urban environment face individual and structural barriers that would increase the difficulty for case managers to access services on the clients' behalf. Workers in the two settings also face different environmental circumstances that impact on how they carry out the case management process. PMID- 10703381 TI - Helping older adults to live better with hearing and vision losses. AB - Because vision and hearing impairments increase in prevalence as age increases, professionals who work with older adults in community settings often encounter people with a wide range of difficulties with their vision and/or hearing. These problems can range from locating financial support to purchasing glasses or hearing aids to obtaining in-home training and devices that will make it possible for the individual with a sensory disability to continue living independently. Meeting the needs of these people requires that professionals be able to recognize sensory losses, accommodate for them, and help older adults to understand and cope with them. PMID- 10703382 TI - Emerging models of care management for older people and those with mental health problems in the United Kingdom. AB - Care management has emerged as a central component in the development of community-based care in many countries. It has been government policy for providers of social services to develop care management systems in the United Kingdom since 1993. This paper examines the extent to which it is possible to begin to discern models of care from the different care management arrangements which are now emerging. First, the background to changes in policy and the role of care management in the UK social care system are discussed; second, evidence from the early phases of care management development in the UK is also examined; and third, based upon the pilot phase of a major national study of care management, the key dimensions of variation in care management through which models may be constituted are identified. PMID- 10703383 TI - Doing well by doing good. The case for objective feedback in case management. AB - Social service programs that do not generate sufficient revenues will not survive in a Fee-For-Service (FFS) system. Yet a concern about finances is alien to many social workers' client-centered orientation. This article presents findings from a study that evaluated the effect of an objective feedback intervention on social workers' productivity in an FFS HIV/AIDS case management program. Results showed a substantial increase in billable hours (13.4% year-to-year; 6.4% pre- to-post intervention) which enable the program to reverse its operating deficit and raise staff salaries. PMID- 10703384 TI - Home care case management. Perspectives from the home front. AB - Focus groups were held with home care case managers in two cities in Canada which provided information on the role of the case manager, factors that influence decision making, recent changes that have taken place in case management, and the different and positive aspects of home care case management. Factors which influence decision making were grouped into organizational, client, family, other professionals, and case management factors. The differences in case managers' preparation and functions in the two sites are discussed. The difficult aspects of case management included making tough decisions related to client resources, and client, work life, and management issues. The positive aspects included the personal interaction with clients, the opportunity to follow through on services to clients, the diversity of case management work, and the opportunity for relatively independent professional practice. PMID- 10703385 TI - Generic drugs in transplantation: new responsibilities for clinical transplant coordinators. PMID- 10703386 TI - Model organ description protocols for completion by transplant surgeons using organs procured from medical examiner cases. AB - A significant number of donor organs emanate from the medical examiner's or coroner's offices, because victims of head injuries from vehicular accidents, falls, assaults, gunshot injuries, and unattended cerebrovascular accidents fall under the jurisdiction of the medical examiner or coroner. Unfortunately, many organ procurement organizations may not fully understand the legal responsibilities of the medical examiner. Most of the medical examiner or coroner cases could be used without compromising the medical-legal responsibilities of the medical examiner or coroner if a reliable description of the respective organ could be made after surgical removal by either having a pathologist present or by having the surgeon prepare a description of the respective organ. The objective of this paper is to present a series of protocols that have been designed to describe the heart, lungs, liver, kidney, and spleen for use by organ transplant surgeons. These protocols have proven to be highly successful in making more organs available for transplantation. PMID- 10703387 TI - When donor families and organ recipients meet. AB - Medical decisions about organ donation and transplantation are considered by a growing number of individuals. The complex issue of whether and to what extent organ recipients and donor families should interact or communicate has gained increasing public awareness, thereby creating an area of major ethical and legal concern for the transplant community. Communication issues have traditionally been decided by transplant coordinators and guided by personal beliefs, agency guidelines, and organizational policies. Organizations are often inconsistent in their practices, and this in turn causes frustration and confusion for both donor families and transplant recipients. This study explored how the experience of meeting the recipient(s) of a loved one's organ affected the grieving process of donor families and altered their lives. The information from this study might be useful to transplant professionals to develop guidelines and policies that lessen the confusion and frustration felt by those involved with the transplant process. PMID- 10703388 TI - Organizational characteristics of solid-organ donor hospitals and nondonor hospitals. AB - CONTEXT: Efforts to increase organ donation include serious attempts in hospital settings, where unrealized donation potential exists. Research on hospital donation must include understanding organizational as well as patient-specific influences on the donation process. OBJECTIVE: To identify organizational characteristics that distinguish hospitals producing organ donations from those that do not, and to estimate the number of nondonor hospitals with donor potential. DESIGN: Data from the American Hospital Association's 1992 annual survey of hospitals were matched to Organ Procurement and Transplantation Network information from the United Network for Organ Sharing regarding the number of solid-organ donors in 1992. Hospitals with donation capability were identified, based on bed size and factors necessary to produce successful donor maintenance and organ recovery. Based on statistical analyses, organizational characteristics distinguishing donor hospitals from nondonor hospitals were identified. We also compared the number of donors and the number of donor hospitals in 1992 and 1996. SETTING: United States. RESULTS: Among all hospitals affiliated with the American Hospital Association (n = 5607), 1214 (22%) were identified as donor hospitals (> or = 1 donation in 1992). Of 2333 hospitals with procurement capability, 1268 (54%) produced no donors in 1992. Based on a multiple logistic regression model, donor hospitals differed from nondonor hospitals by hospital ownership, with municipally owned hospitals more likely and federally owned hospitals less likely to produce donation, compared with for-profit and not-for-profit hospitals. Other organizational characteristics associated with donor hospitals were level of trauma services, whether the hospital had a transplant surgery program or a hospital ethics committee, and whether it was located in the South Atlantic, Southwest Central, or Pacific regions of the United States. CONCLUSIONS: Among hospitals not currently producing organ donations, there is a sizable subgroup with donor potential. This area merits further attention. PMID- 10703389 TI - A comparison of OPO pulsatile machine preservation practices and results. AB - CONTEXT: Kidney preservation has been performed by either ice (static) or machine pulsatile perfusion. Ice storage is simple, with only 1 methodology. Machine perfusion, on the other hand, is accomplished using multiple methodologies. This article delineates the different methodologies of pumping centers throughout the country. OBJECTIVE: Pulsatile machine perfusion is again being viewed as the preservation method of choice for kidneys from non-heart-beating cadaver donors and cadaver kidneys from marginal donors. To develop indices to predict the viability of cadaver kidneys for transplant, a review of the organ procurement organizations, specific perfusion techniques, and a comparison of the delayed graft function and graft survival rates were considered. METHODS: A survey, asking for specifics on perfusion parameters, pulsatile machine perfusion experience, and criteria for perfusion implementation and graft survival results, was mailed to all organ procurement organizations in the United States. RESULTS: Of the 44 centers that responded to the survey, 12 used pulsatile machine perfusion (11 used the Waters perfusion machine), 6 pumped marginal cadaver kidneys, and the remaining 6 pumped all cadaver kidneys. Minimum perfusion criteria, pulse rates, perfusate composition, pressures, renal resistance, and renal pressure and flow were considered. Vasodilators and other machine additives were used to improve flow. The variance in each center's number of cadaver kidneys pumped each year, as well as the differences in pump times, was noted. CONCLUSION: Twelve centers use pulsatile machine perfusion. A variety of techniques are used to perform pulsatile machine perfusion, but 11 of 12 have less delayed graft function than those programs employing ice storage preservation. PMID- 10703390 TI - Outcome of kidney transplantation under tacrolimus-based immunosuppression in elderly patients. AB - Kidney transplantation has become a reasonable treatment option for selected patients aged 60 years or older, and a number of different immunosuppressive drug protocols have been described. This article concerns 230 recipients who were aged 60 years or older and who were undergoing kidney-only transplantation at the University of Pittsburgh between January 1990 and April 1997. All recipients were treated with a tacrolimus-based immunosuppression regimen. The median follow-up was 31.5 months (range, 1-86). The 1-, 3-, and 5-year actuarial patient survival rates were 90%, 83%, and 76%, respectively. There were 42 (19%) deaths, cardiovascular disease (50%) and infection (38%) being the main causes. Death with a functioning kidney occurred in 28 (67%) patients. The 1-, 3-, and 5-year actuarial graft survival rates were 84%, 74%, and 64%, respectively. The delayed graft function rate was 33%. Rejection was seen in 57 (25%) elderly patients. The mean serum creatinine was 2.6 +/- 2.7 mg/dL and the serum urea nitrogen was 35 +/ 22 mg/dL. The mean tacrolimus level was 8.5 +/- 3.8 ng/mL. These results suggest that renal transplantation in older recipients under tacrolimus-based immunosuppression is associated with reasonable outcomes, and can be offered to appropriately selected patients. PMID- 10703391 TI - Compliance and noncompliance in kidney transplant patients: cues for transplant coordinators. AB - Maximizing kidney transplant patients' long-term compliance with immunosuppressants is a major challenge to transplant coordinators. Although previous research has found substantial proportions of recipients to be noncompliant, predictors of noncompliance and characteristics of noncompliers remain unclear. In this study of more than 1400 kidney transplant patients, we found noncompliance to be associated with patient and transplant characteristics and with patient beliefs concerning the efficacy of immunosuppressants. Three distinct profiles of noncompliers were identified: accidental noncompliers, invulnerables, and decisive noncompliers. This information can be used by transplant coordinators to recognize cues that predict noncompliance and to work with at-risk patients to forestall or remedy noncompliant behavior. PMID- 10703392 TI - Interventions in a heart transplant recipient with a histrionic personality disorder. AB - Organ transplantation is a psychosocially demanding process. Patients must undergo a comprehensive evaluation to await a donor organ that may never become available. After transplantation, recipients must deal with the acceptance of a new organ and comply with a medical regimen that includes numerous medications, follow-up exams, and procedures. Emotional well-being is monitored throughout the transplant process. However, despite the best of efforts and thorough pretransplant bio-psycho-social evaluations, it is possible for patients to have significant psychopathology that remains undetected. Following the stress of transplantation, such patients may present with exacerbation of symptomatology, which has the potential to negatively affect compliance and long-term outcome. PMID- 10703393 TI - New strategies using 'low-dose' mycophenolate mofetil to reduce acute rejection in patients following kidney transplantation. AB - CONTEXT: Tacrolimus, microemulsion cyclosporine (Neoral), and mycophenolate mofetil (MMF) at 2 and 3 grams daily have demonstrated superior immunosuppressive properties in several recent clinical trials involving solid-organ transplants. An effective immunosuppression may be maintained with lower doses of MMF administered with either tacrolimus or microemulsion cyclosporine. OBJECTIVE: To compare tacrolimus plus "low-dose" MMF-based immunosuppressive regimen (TMBIR) with Neoral plus "low-dose" MMF-based immunosuppressive regimens (NMBIR) among kidney transplant recipients. DESIGN: Prospective, randomized study. PATIENTS: 53 consecutive adult recipients of kidney transplant. Both groups (TMBIR and NMBIR) were equally matched on demographic characteristics. INTERVENTIONS: Participants were randomized to receive orally either tacrolimus (0.08 mg/kg twice daily) (n = 27) or Neoral (4 mg/kg twice daily) (n = 26). Both regimens were started before surgery and continued when allograft demonstrated no postoperative acute tubular necrosis. Both groups received similar "low-dose" MMF (500 mg twice daily) and prednisone (2 mg/kg/day to taper off after 1 year). Switch from tacrolimus to Neoral or vice versa was allowed after refractory rejection or serious adverse events. MAIN OUTCOME MEASURE: Acute rejection and patient and graft survival 1 year following kidney transplant. RESULTS: One-year patient survival rates were 88.9% for the TMBIR group and 100% for the NMBIR group; 1-year graft survival rates were 88.9% for the TMBIR group and 96.1% for the NMBIR group. No significant differences were found in the incidence of biopsy-confirmed acute rejection (14.8% TMBIR vs 23% NMBIR). Steroid-resistant rejections requiring cytolytic antibody therapy were higher in the NMBIR group (50% vs 25%). Three patients crossed over from NMBIR to TMBIR for refractory rejections and 1 patient crossed over from TMBIR to NMBIR for new onset seizure. Three episodes of cytomegalovirus infection were observed in the TMBIR group. Other adverse events were similar in both groups. CONCLUSIONS: Both tacrolimus and microemulsion cyclosporine combined with "low-dose" MMF and corticosteroids provide effective immunosuppression and have similar adverse events in kidney transplant recipients. PMID- 10703394 TI - Bone densitometry should be included in the evaluation of candidates for lung transplantation. AB - Bone loss and fractures are common complications of heart and liver transplantation, and are likely related to high-dose immunosuppressive therapy. We have previously demonstrated that many patients with end-stage lung disease already have osteoporosis and may be at even greater risk for fracture after lung transplantation. The purpose of this study is to determine the incidence of fracture in lung transplant recipients on osteoporosis prevention regimens, the relationship of fracture to pretransplant bone mineral density, and the impact of fracture on quality of life after lung transplantation. Twenty-one lung transplant candidates were prospectively evaluated with spine radiographs and bone mineral densitometry. Bone density was expressed as T scores, the number of standard deviations from the mean bone density of a young normal population of the same gender. Of 21 patients, 8 (38%) fractured during the first year. The mean pretransplant lumbar spine T score was significantly lower in the fracture patients (P = .03). Four of the 7 surviving fracture patients and 1 of the 10 patients who survived without fracture believed that chronic pain diminished their quality of life (X2 = 4.408; P = .04). These findings suggest that bone mineral density should be routinely included in the evaluation of lung transplant candidates. Patients with extremely low bone density or osteoporotic fracture should be counseled about the increased risk of fracture after transplantation. PMID- 10703395 TI - Quality of life after transplantation. PMID- 10703396 TI - Issues in cyclosporine drug substitution: implications for patient management. AB - Substantial improvements in short-term and long-term outcomes for kidney transplant recipients have resulted from better use of existing immunosuppressive agents and newer treatment options. Calcineurin inhibitors (e.g., cyclosporine and tacrolimus) remain the foundation of immunosuppressive therapy. These agents are considered critical-dose drugs because of their narrow therapeutic range, variable pharmacokinetics, formulation-dependent bioavailability, and negative clinical consequences of underdosing or overdosing. With the recent introduction of a new cyclosporine formulation, concern exists that current bioequivalence guidelines for generic approval may not provide adequate assessment of the safety and efficacy of critical-dose drugs. Transplant experts at 2 recent conferences recommended more rigorous criteria for bioequivalence testing of critical-dose drugs and adoption of consistent drug substitution practices. Additional recommendations included specifying the intended formulation and instituting appropriate monitoring whenever formulations are switched. A summary of the outcomes of these conferences and practice implications for transplant coordinators is discussed. PMID- 10703397 TI - Rare combined heart and kidney transplant in a pediatric patient: a case study. AB - Multiple reports of successful combined heart and kidney transplants adults suggest that this may be a viable option for a small subset of patients with coexisting end-stage heart and kidney failure. A review of the literature, however, reveals that few combined heart and kidney transplants have been reported in children. This article presents the case of a 13-year-old boy who underwent unsuccessful palliative surgery for a congenital heart defect. The patient developed heart failure with subsequent acute renal failure, and ultimately required a combined heart and kidney transplant. The combined procedure was successful in this patient and he is alive and well 27 months postoperatively. PMID- 10703398 TI - Medication nonadherence and its relation to financial restriction. AB - The question of patient nonadherence has always been an important factor in determining candidate suitability for organ transplantation. Data that explore the association of financial problems and posttransplant medication nonadherence are limited. Findings suggest that medication nonadherence was more likely to occur when recipients did not have insurance coverage and had to rely on Medicaid or indigent drug programs. Our center developed a formalized program within the outpatient pharmacy, including a full-time medication counselor who helped recipients secure resources to pay for pre- and posttransplant medications. To determine whether the availability of posttransplant medications could reduce medication nonadherence, we conducted a survey with 50 consecutive liver transplant recipients in the outpatient clinic. Nonadherence rates were significantly reduced from 25% to 10% (P < .01) compared with recipients who had been transplanted before the development of our drug program. These results suggest that optimum medication adherence can be obtained when recipients are provided guidance in securing their necessary medications without financial restriction. PMID- 10703399 TI - Liver donation by a trauma patient: a case study in placement. AB - Currently, more than 64,000 people are awaiting transplants in the United States. Transplant coordinators must do everything possible to ensure that viable organs from consented donors are transplanted. To evaluate donors and organ function, transplant coordinators rely on a multitude of diagnostic tests to determine donor organ suitability. How reliable are the results of these tests? The following case study presents an incident in which diagnostic test results were not accurate; as a result, transplant centers deferred what turned out to be a normal, atraumatic organ. The end result was that this organ was placed, but only after actual visualization in the operating room and the granting of full waivers to the transplanting center. PMID- 10703400 TI - Self-care guidelines: finding a common ground. AB - Pressure to reduce overall transplant costs is one of the factors which has led to earlier hospital discharge and increased patient management challenges in outpatient and home care settings. Earlier discharge often contributes to decreased opportunity to provide and ensure comprehension of critical patient and family education, resulting in challenges for home care clinicians who are committed not only to patient and environmental assessments, but to helping assure patient and family understanding of and compliance with critical posttransplant responsibilities and regimens. This article describes a transplant patient education tool that was developed for use with all types of solid organ transplant recipients discharged from multiple centers yet managed by a single home care organization. The tool provides patient education information that can be realistically reviewed and reinforced during the home visit. The resource focuses on key self-care issues to promote wellness and graft survival and help prevent adverse outcomes. PMID- 10703401 TI - Outpatient housing following kidney transplantation. AB - The purpose of this article is to describe the efforts of a medium-sized kidney transplant program in the southeastern United States to reduce the inpatient length of stay by moving toward the outpatient arena, thereby reducing the overall cost of the transplant experience. An outpatient transplant unit was created to house patients who were medically stable but still required further monitoring and education prior to being discharged. The development and implementation of the transplant outpatient unit significantly reduced inpatient length of stay following kidney transplantation from 14 to 5 days. The benefits and outcomes of the transplant outpatient unit were impressive. PMID- 10703402 TI - Family communication coordination: a program to increase organ donation. AB - To improve organ donation performance, the Medical College of Virginia Hospitals implemented a comprehensive family support and communication program, consisting of a standard family communications protocol, a hospital-based team from the Department of Pastoral Care, targeted staff education, and an ongoing quality assurance measuring and monitoring system. The 3 best-demonstrated request practices, private setting, "decoupling," and collaboration in the request between the organ procurement organization and hospital staff, were incorporated into the program. Improvement in the consent and donation rate was evident in the second calendar year of the program; the consent rate was 72% and the donation rate was 50%. During the second year, there was also a positive correlation between "decoupling," appropriate requestor, and the consent rate. Implementation of a hospital-based team and a standard protocol facilitated the clarification of roles and responsibilities toward clearer and more consistent family communication and support. Data suggest that staff experience is a major contributor to a positive donation outcome. PMID- 10703403 TI - Nerve transplantation: a father's final gift. AB - Offering the option of organ and tissue donation to grieving families may seem stressful, but asking the question may provide a positive means to extend care to the bereaved family and help others in return. Many donor families have said donation was an opportunity to make some sense out of a senseless situation and to relieve some of the grief they experienced. This article presents a case that started with such a discussion by ICU nurses in one of our donor hospitals, and ended with successful organ and tissue recovery and transplantation. As "routine" as this may sound, it was anything but routine--it made history. PMID- 10703404 TI - Comparison of consent rates between hospital-based designated requestors and organ procurement coordinators. AB - Recent legislation in our service area has put restrictions on who is allowed to approach families about donation. Many hospitals have therefore requested training for designated requestors to speak with families. The organ procurement organization followed the consent rate of 2 hospitals to evaluate the effectiveness of the designated requestors in the consent process. This article compares the success of the hospital-based designated requestor program in obtaining consent with that of another hospital, which relies solely on the organ procurement coordinator to approach families. Thirteen staff members in hospital A received the 8-hour designated requestor training. A decrease in the consent rate at hospital A prompted the organ procurement organization to interview the 13 requestors. Possible reasons for the decline ranged from lack of experience to the lack of identification as donor family advocate. Both hospitals were encouraged to consider a multidisciplinary approach, which includes the organ procurement coordinator in the request process. PMID- 10703405 TI - Utilization of donated lungs in Australia: 1989-1997. AB - Lung and heart-lung transplantation was first undertaken in Australia in the late 1980s and early 1990s. Although detailed data are available on Australian lung transplantation outcomes, little data are available regarding the utilization of donated lungs. This study examines donated lung utilization rates and considers various factors that may affect these rates. Australian donation and transplantation data were analyzed for the years 1989 through 1997. Results showed that 24% of overall donors were lung donors. The percentage of donors from whom at least 1 lung was transplanted increased from 6% in 1989 to 36% in 1997. Heart-lung transplantation rates changed little (2%-9%), whereas bilateral lung transplantation increased from 1% to 23% of donors. Single-lung donors accounted for 32% of lung donors in 1997. Uniform basic donor criteria and management guidelines, simple allocation mechanisms, and cooperative retrieval have evolved during this time. Close collaboration at the time of donation between units, coordinators, and ICUs has allowed early retrieval from well-managed donors. PMID- 10703406 TI - Estimating the non-heart-beating donor potential at a trauma center. AB - The number of people awaiting a life-saving transplant far outweighs the number of organs available for transplantation. Our organ procurement organization sought to identify the non-heart-beating donor potential at a large urban level 1 trauma center. We modified the death-record reviews for a 2-year period and determined that a total of 23 patients who expired were suitable for non-heart beating donation. Therefore, it can be hypothesized that if all large urban level 1 trauma centers participated in non-heart-beating donation, more organs would be available for transplantation. PMID- 10703407 TI - The importance of the words we use. PMID- 10703408 TI - Improved cognitive knowledge of midwives practising in the eastern Cape Province of the Republic of South Africa through the study of a self-education manual. AB - OBJECTIVES: To determine whether the Maternal Care manual of the Perinatal Education Programme (PEP) is effective in improving the cognitive knowledge of midwives. DESIGN: A prospective controlled trial in a region where PEP was not previously used. The midwifery knowledge of all midwives caring for pregnant women in the three towns was tested before the commencement of the study. The Maternal Care manual was then introduced to the midwives in he study town and they worked through the programme. Following the completion of the manual, all midwives were tested again with the same test. The time interval between the pre and post testing was 12 months. SETTING: Three towns on the Eastern Cape Province of South Africa. PARTICIPANTS: All Midwives caring for pregnant women in three towns. INTERVENTIONS: The Maternal Care manual of PEP was studied by the midwives in the study town. MEASUREMENTS AND FINDINGS: Changes in cognitive knowledge were tested with multiple choice questions. A significant improvement (p < 0.0001) was achieved in the study town. The mean score (maximum 70) improved by 22.4 (32%) marks, from 35.9 (51%) to 58.3 (83%). KEY CONCLUSIONS: The cognitive knowledge of midwives who completed the Maternal Care manual improved significantly. IMPLICATIONS FOR PRACTICE: These findings are to particular importance as the method by which the Maternal Care manual of PEP was applied conforms to the method suggested for the its national use. PMID- 10703409 TI - A survey of women's experiences of vaginal loss from 24 hours to three months after childbirth (the BLiPP study). AB - OBJECTIVE: To describe the range of normal vaginal loss as reported by women from 24 hours after delivery until three months postnatally. SETTING: Two health districts in the south of England. METHODS: A prospective survey of women's experiences and expectations of the duration, amount and colour of vaginal loss after childbirth. The term vaginal loss includes all types of fluid loss from the vagina following childbirth. FINDINGS: Five hundred and twenty-four women were recruited to the survey in 1995. Vaginal loss, as reported by the women, was considerably more varied in duration, amount and colour than descriptions in current midwifery textbooks. The median number of days reported for the duration of vaginal loss was 21 days and the interdecile range (10th to 90th percentile) was 10-42 days. For colour of lochia, women overall reported their vaginal loss to be more predominantly red/brown in colour and the traditional descriptions of the timing and colour phases of lochia rubra, serosa and alba are not supported by the majority of the women's experiences. Primiparous women were significantly more likely to report feelings of surprise or shock about their experiences of vaginal blood loss after the birth (odds ratio 4 [95% Confidence Interval 2-9]). Seven primiparous women (2%) were unaware that they would have a blood loss at all after the birth. IMPLICATIONS FOR PRACTICE: The findings from this survey have been used to develop information leaflets for women and health professionals about vaginal loss following childbirth. These leaflets include descriptions of normal ranges for the colour, amount and duration of vaginal loss in the first three months after childbirth. PMID- 10703410 TI - Midwives' lived experiences of being supportive to prospective mothers/parents during pregnancy. AB - OBJECTIVE: To elucidate midwives' narrated experiences of being supportive to prospective mothers or parents during pregnancy. DESIGN: Phenomenological hermeneutic analysis of transcribed text from seven tape-recorded interviews. SETTING: Midwifery clinics in five health centres in the context of Swedish primary health care. PARTICIPANTS: Seven midwives working in antenatal care. FINDINGS: The interpretation of the text showed that through perception and intuition the midwives seemed to become aware that some women needed increased support. The situations of these prospective mothers were often characterised by difficult social problems or fears. The prospective fathers were mostly absent in the narratives. The midwives acted on both a personal and a professional level with ethical perspectives in mind, when they were advocating their clients' rights to receive proper care. The comprehensive understanding of the interpretation revealed that the midwife sometimes perceived herself as being metaphorically 'The Good Mother'. KEY CONCLUSIONS: Having the role of 'The Good Mother' could be understood as a way for the midwife to establish a fruitful relationship with prospective mothers/parents. IMPLICATIONS FOR PRACTICE: The findings provide a basis for reflection on the mothering and supportive function of midwives when providing antenatal care. PMID- 10703411 TI - Feeding outcome in breast-fed term babies supplemented by cup or bottle. AB - OBJECTIVE: To describe the feeding method at discharge from midwifery care of term babies supplemented either by cup or bottle while in hospital. DESIGN: A retrospective review of the obstetric and midwifery records of 531 consecutively born babies. SETTING: A large maternity unit, with an integral General Practitioner Unit, in an inner city in the south of England. PARTICIPANTS: 63 term breast-feeding babies; 30 supplemented by cup and 33 supplemented by bottle. MAIN OUTCOME MEASURE: Breast feeding on discharge from midwifery care. FINDINGS: There were no significant differences between the bottle and cup supplementation groups in relation to feeding outcome (OR 1 94 95% CI 0.61, 6.31), or in the length of time from the beginning of supplementation to leaving hospital (median difference 1 95% CI 0, 1) or discharge from midwifery care (median difference 0 95% CI, -1, 1). Babies who received supplements of expressed breast milk, as opposed to artificial milk, were more likely to be supplemented by cup (OR 4 29, 95% CI 0.9, 26.91; p = 0.05), but were not more likely to be discharged from midwifery care breast feeding (OR 3.79, 95% CI 0.69, 38.36). CONCLUSION: Owing to the small scale and retrospective nature of this survey, its results must be viewed with caution. However, given the apparent lack of evidence in this area, prospective work should be undertaken to examine the most appropriate method of supplementation for term babies. Generalisation from work related to babies in special care baby units is no longer acceptable. PMID- 10703412 TI - Indigenous first feeding practices in newborn babies. AB - OBJECTIVE: To describe the common practices in indigenous first feeding of the newborn babies in countries of the continents of Africa, Asia and Latin America. METHODS: Anthropological, medical and nursing publications were searched for indigenous customs and beliefs concerning the first feed given to newborn babies. FINDINGS: From a nutritional point of view the first food is just symbolic but it has a function in either purifying and clearing the 'dirty' throat and bowel or to prepare the baby for adult life. CONCLUSIONS: The general withholding of colostrum should be abandoned and the indigenously strange custom of feeding water with sugar should be discouraged. PMID- 10703413 TI - A place for the partner? Expectations and experiences of support during childbirth. AB - AIM: To explore the expectations primigravidae have concerning the support that they hoped to have and would need from their partner during childbirth, and whether these kinds of support were actually provided by their partner. Additionally, to explore the thoughts and feelings of male partners concerning their supporting role, and, in retrospect, how well they felt they had managed. PARTICIPANTS: Eight couples living in Hampshire, UK, who were interviewed six weeks before the birth and approximately 12 weeks following labour and delivery. METHODS: Semi-structured interviews were taped, transcribed and analysed. An ethnographic approach was used to identify concepts and themes. FINDINGS: Support provided by the male partner evoked very positive responses from the women. The fathers perceived that they were very helpful to their partner during childbirth. Though the women mostly found childbirth straightforward some fathers, nevertheless, found the experience stressful. CONCLUSIONS: The father's needs and role should be regularly assessed during childbirth. PMID- 10703414 TI - Maintaining equilibrium: a grounded theory study of the processes involved when women make informed choices during pregnancy. AB - OBJECTIVE: To map the processes involved when women make informed choices during pregnancy. DESIGN: A grounded theory approach was used. Data were collected by means of focused interviews and observation. SETTING: Naturalistic, in antenatal clinics and participants' homes. PARTICIPANTS: Pregnant women receiving care in a variety of maternity settings in England. KEY FINDINGS: The core category was named Maintaining Equilibrium, whereby the woman attempted to make choices that would preserve the balance of her and her family's life. Substantive categories were Regulating, Contextualising and Actioning. IMPLICATIONS FOR PRACTICE: The core and substantive categories are discussed in relation to midwifery practice, with particular reference to how women judged the trustworthiness of the information and its source, and the strategies they used to operationalise their choices. The need is stressed for midwives to be sensitive and flexible regarding meeting the information needs of women, in order that women may reach their own decisions about how best to maintain their equilibrium. PMID- 10703415 TI - The prevalence of stress incontinence during pregnancy and following delivery. AB - OBJECTIVES: To examine the variation in findings from epidemiological studies which describe the prevalence of stress incontinence during and after pregnancy, and to undertake a prospective survey of the prevalence of stress incontinence during pregnancy and following childbirth in order to provide clarification of the findings presented in the literature. DESIGN: A review of the literature was undertaken using the Medline and Popline CD Rom. A postal questionnaire was sent to a sample of women when they reached 34 weeks' gestation and repeated at 8 weeks postpartum. PARTICIPANTS: 1008 women were recruited to the study when they attended the antenatal clinic at two hospitals in the north west of England. Seven hundred and seventeen (71%) women responded to the first questionnaire and 572 (57%) completed the second questionnaire. FINDINGS: The prevalence of stress incontinence during pregnancy reported in the literature ranges from 20 to 67%. Following delivery the reported prevalence is between 6 and 29%. In the present study 59% of women reported stress incontinence during pregnancy, and 31% following delivery. Ten per cent of the women had daily episodes of incontinence during their pregnancy, 2% of all women reported daily incontinence following delivery. An association was found between parity and stress incontinence, with women of higher parity being more likely to experience the condition. No difference in the prevalence of stress incontinence was found between women who had a normal delivery and those having an instrumental delivery. A caesarean section was found to be associated with a lower incidence of stress incontinence compared with a normal spontaneous delivery. KEY CONCLUSION: A high proportion of women experienced stress incontinence during pregnancy and/or following delivery. Some women reported severe symptoms, with leakage on a daily basis. Women of higher parity were more likely to suffer from the condition. Whilst women who had a normal spontaneous delivery or an instrumental delivery reported a similar level of stress incontinence, women who had a caesarean section were less likely to have the condition. PMID- 10703417 TI - Privacy is an issue that merits much more attention in nursing ethics. PMID- 10703416 TI - The significance of 'hierarchy' in a research project on adoption and adaptation. AB - In the course of phenomenological research project focusing on the midwife's care of the mother who relinquishes her baby for adoption, hierarchy emerged repeatedly as a significant theme. This concept manifested itself both methodologically as well as thematically. In methodological terms, hierarchy first became apparent during the planning phase in the selection of the research design: at this point it took the form of the distinction between quantitative and qualitative research design. In the course of applying for research access the same attitudes re-emerged in the gatekeepers' responses to the research design which had been chose. Later, during the field work, hierarchy appeared in the course of the interviews, in the form of the unequal balance of power between the interviewer and the informant. Specific tactics were required to overcome the likelihood of hierarchy affecting the data at this stage. Eventually this concept was raised by the informants, who included both relinquishing mothers an midwives, and became a major theme. The informants' use of hierarchy to explain the unselfishness of relinquishment proved to be one of a number of strategies which facilitated coping either with the experience of relinquishment or with caring for a women going through that experience. The conclusion which emerges is that hierarchy features in many aspects of life. While it may be beneficial when applied to certain situations to facilitate coping, in other circumstances it may be less than positive. PMID- 10703418 TI - Academic tension and nursing ethics research. PMID- 10703419 TI - Recovering ethics after 'technics': developing critical text on technology. AB - Much modern science and ethics debate is on high-profile problems such as animal organ transplantation, genetic engineering and fetal tissue research, in discourse that assumes technical tones. Other work, such as narrative ethics, expresses the failed promise of technology in the vivid detail of human experience. However, the essential nature of contemporary technology remains largely opaque to our present ethical lens on health care and on society. The limited controversies of modern science and ethics perpetuate 'technics', a technical, problem-solving mindset that fails to grapple successfully with the complexity of technology. A critical dialectic between practice and scholarship widens the ethical conversation in nursing to consider technology as an ongoing set of daily and fundamental moral choices on how we live. Critical text on technology recovers ethics from the limits of technics, and assists nurses to develop an inherent knowledge of technology that is needed to provide ethical care in a technological world. There are overlooked ethical challenges in the mundane, everyday routine activities of professional practice, and these have gone largely unexamined. Ethical behavior is not the display of one's moral rectitude in times of crisis. It is the day-to-day expression of one's commitment to other persons and the ways in which human beings relate to one another in their daily interactions. PMID- 10703421 TI - Researching the bereaved: an investigator's experience. AB - The issues discussed in this article concern the process of interviewing the bereaved relatives of organ donors, the personal impact, and the potentially painful nature of such research. Narrative interviews were carried out with 24 donor relatives. The relatively small number of donating families and their anonymity mean that little is understood about the experience of having a relative in a critical care situation that ends in donation. The purpose of this study was to develop a theory that explained the organ donation process for relatives of 'major organ' donors. A central concern of implementing the investigation was the possible threat it posed to the participants and myself. The sensitive nature of the research made access to relatives difficult. Undoubtedly, my nursing background and personal attributes had an impact on interactions with participants and the pursuance of the research agenda. PMID- 10703420 TI - 'Healthy viewing?': experiencing life and death through a voyeuristic gaze. AB - Recent times have witnessed a groundswell in the number of British television programmes that deal with the 'real life' experiences of people in various health care settings. Such programmes tend to focus upon the two interrelated strands of the experience of those who deliver professional care and those who are at the receiving end of it. The usual rationale given for such programmes is that they offer insights about the delivery of health care that are not readily accessible to members of the public. This article will look beneath the rationale and reasons offered by programme makers for the existence of such documentaries. It will explore insidious and questionable elements that go beyond revealing the 'lived experience' of professional carers and those for whom they care. Emerging from this is the challenging notion that such programmes deliver the opportunity to experience the vulnerability, suffering and even death of others through a voyeuristic gaze. PMID- 10703422 TI - The morality of treating patients with depot neuroleptics: the experience of community psychiatric nurses. AB - The aim of this qualitative study was to gain an understanding of the meaning that community psychiatric nurses impart to their everyday interactions with patients in depot neuroleptic treatment situations. Nine experienced community psychiatric nurses were interviewed using semistructured, open-ended questions. Data analysis was by the phenomenological descriptive method according to Giorgi. Four themes were identified, highlighting aspects of the moral meaning of treating patients with depot neuroleptics: (1) 'benevolent justification' occurs when nurses perceive that the patient's welfare is at stake; (2) 'inability to advocate the patients' best interest' occurs when nurses feel they are at a disadvantage; (3) 'accommodative interactions' occur when nurses are able to respond to a patient's expressed needs; and (4) 'acceptable advocacy' occurs when physicians are sensitive to nurses' suggestions on patients' treatment. The findings indicate that treatment care planning involving both patients and nurses is essential to enhance patients' autonomy, which is a precondition for satisfactory interactions. This phenomenological study describes the meaning that nurses give to administering depot neuroleptic injections to patients in the context of community psychiatric clinics. The phenomenon of concern was identified as the moral aspect in the interactions with individual patients in the treatment situation. PMID- 10703424 TI - Case study. Whose power, whose ethics? A student nurse's narrative. PMID- 10703423 TI - Women physicians' narratives about being in ethically difficult care situations in paediatrics. AB - This study is part of a comprehensive investigation of ethical thinking among male and female physicians and nurses. Nine women physicians with different levels of expertise, working in various wards in paediatric clinics at two of the university hospitals in Norway, narrated 37 stories about their experience of being in ethically difficult care situations. All of the interviewees' narrations were concerned with problems relating to both action ethics and relation ethics. The main focus was on problems in a relation ethics perspective. The most common themes in an action ethics perspective were overtreatment and withholding treatment. The more experienced physicians reasoned differently from the group of less experienced physicians and they coped with pressure in different ways. The less experienced physicians disclosed their professional experience yet seemed uncertain, while putting on an air of certainty, but the more experienced physicians disclosed both their professional and personal experience of caregiving and they seemed to allow themselves to feel uncertain in their certainty. Both groups emphasized a need for deep discussion between colleagues about their being in ethically difficult care situations. PMID- 10703425 TI - Project Romania: education in ethics and elaboration of guidelines for ethics in nursing for Romanian nurses. PMID- 10703426 TI - The code of ethics for nurses: Romanian Nursing Association. PMID- 10703428 TI - A dose of reality. PMID- 10703427 TI - The need to refocus nursing and nurse leadership on the possible. PMID- 10703429 TI - Value added decisions. PMID- 10703430 TI - Informed choice. PMID- 10703431 TI - Risking harm. PMID- 10703432 TI - Voice of the people. PMID- 10703433 TI - Crisis management. PMID- 10703434 TI - Northern Ireland new horizons. Interview by Tom Keighly. PMID- 10703435 TI - Implementing clinical supervision. PMID- 10703436 TI - Partnership. The challenge for nursing. PMID- 10703437 TI - Making a difference and our healthier nation. PMID- 10703438 TI - The phantom menace. PMID- 10703439 TI - Internet recruitment. PMID- 10703440 TI - Learning the art of management. PMID- 10703441 TI - Beverly Malone. Interview by Ian McMillan. PMID- 10703442 TI - Educating nurses. Interview by Tom Keighley. PMID- 10703443 TI - The informal process. Discipline at work. Part One. PMID- 10703444 TI - Rehabilitation. PMID- 10703445 TI - Widening entry routes. PMID- 10703446 TI - Openness. PMID- 10703447 TI - If numbers are to be the gold standard to measure and assess practice, health care is in real danger. PMID- 10703448 TI - At home blues. PMID- 10703449 TI - What respect? PMID- 10703450 TI - Palliative care for all. PMID- 10703451 TI - Home but not alone. PMID- 10703452 TI - Inequality in care for older people. PMID- 10703453 TI - Children first. PMID- 10703454 TI - Are you in control? PMID- 10703455 TI - Family-friendly policies: a panacea for the NHS? PMID- 10703456 TI - Partnership. Professional regulation. PMID- 10703457 TI - Proteomics and nutrition--a science for the first decade of the new millennium. PMID- 10703458 TI - Cobalt-deficiency-induced hyperhomocysteinaemia and oxidative status of cattle. AB - In ruminants, Co is required for the synthesis of vitamin B12, which in turn is needed for the resynthesis of methionine by methylation of homocysteine and thus, cobalamin deficiency may induce hyperhomocysteinaemia which is brought into context with perturbations of the antioxidative-prooxidative balance. The present study was conducted to explore whether Co deficiency in cattle is also associated with homocysteine-induced disturbances of oxidative status. Co deficiency was induced in cattle by feeding two groups of animals on either a basal maize-silage based diet that was moderately low in Co (83 micrograms Co/kg DM), or the same diet supplemented with Co to a total of 200 micrograms Co/kg DM, for 43 weeks. Co deficiency was apparent from a reduced vitamin B12 status in serum and liver and an accumulation of homocysteine in plasma which was in excess of 4.8 times higher in Co-deprived cattle than in controls. The much increased level of circulating homocysteine did not indicate severe disturbances in antioxidant-prooxidant balance as measured by individual markers of lipid peroxidation, protein oxidation, and the antioxidative defence system. There were no quantitative difference in plasma thiol groups, nor were there significant changes in concentrations of alpha-tocopherol, microsomal thiobarbituric acid-reactive substances and carbonyl groups in liver. However, there was a trend toward increased plasma carbonyl levels indicating a slight degradation of plasma proteins in the hyperhomocysteinaemic cattle. Analysis of the hepatic catalase (EC 1.11.1.6) activity revealed an 11% reduction in Co-deficient cattle relative to the controls. These results indicate that long-term moderate Co deficiency may induce a severe accumulation of plasma homocysteine in cattle, but considerable abnormalities in oxidative status failed to appear. PMID- 10703459 TI - Altering the temporal distribution of energy intake with isoenergetically dense foods given as snacks does not affect total daily energy intake in normal-weight men. AB - The objectives of the present study were to examine the effects of (1) ingesting mandatory snacks v. no snacks and (2) the composition of isoenergetically-dense snacks high in protein, fat or carbohydrate, on food intake and energy intake (EI) in eight men with ad libitum access to a diet of fixed composition. Subjects were each studied four times in a 9 d protocol per treatment. On days 1-2, subjects were given a medium-fat maintenance diet estimated at 1.6 x resting metabolic rate (RMR). On days 3-9, subjects consumed three mandatory isoenergetic, isoenergetically dense (380 kJ/100 g) snacks at fixed time intervals (11.30, 15.30 and 19.30 hours). Total snack intake comprised 30% of the subjects' estimated daily energy requirements. The treatments were high protein (HP), high carbohydrate (HC), high fat (HF) and no snack (NS). The order was randomized across subjects in a counterbalanced, Latin-square design. During the remainder of the day, subjects had ad libitum (meal size and frequency) access to a covertly manipulated medium-fat diet of fixed composition (fat:carbohydrate:protein, 40:47:13 by energy), energy density 550 kJ/100 g. All foods eaten were investigator-weighed before ingestion and left-overs were weighed after ingestion. Subjective hunger and satiety feelings were tracked hourly during waking hours using visual analogue scales. Ad libitum EI amounted to 13.9 MJ/d on the NS treatment compared with 11.7, 11.7 and 12.2 MJ/d on the HP, HC and HF diets respectively (F(3,21) 5.35; P = 0.007, SED 0.66). Total EI values were not significantly different at 14.6, 14.5, 15.0 and 14.2 MJ/d respectively. Snack composition did not differentially affect total daily food intake or EI. Average daily hunger was unaffected by the composition of the snacks. Only at 12.00 hours did subjects feel significantly more hungry during the NS condition, relative to the other dietary treatments (F(3,18) 4.42; P = 0.017). Body weight was unaffected by dietary treatment. In conclusion, snacking per se led to compensatory adjustments in feeding behaviour in lean men. Snack composition (with energy density controlled) did not affect the amount eaten of a diet of fixed composition. Results may differ in real life where subjects can alter both composition and amount of food they eat and energy density is not controlled. PMID- 10703460 TI - Total fluoride intake and urinary excretion in 4-year-old Iranian children residing in low-fluoride areas. AB - Knowledge of levels of fluoride ingestion and excretion is important in planning optimum fluoride therapy for young children. In previous literature, it has been assumed that only about one-third of ingested fluoride is excreted in young children. The aims of the present study were (a) to measure total fluoride intake, urinary fluoride excretion and fluoride balance, and (b) to investigate the effect of air temperature on fluoride intake and urinary fluoride excretion, in young children. Children (4 years old) living in a city, a small town and rural areas of Fars province, Iran, where drinking water contained 0.30-0.39 mg F/l, were invited to participate. Selection of subjects was by random sampling of kindergartens or health centres. The children were surveyed twice, once in summer and once in winter. Diet was obtained by 3 d diaries with interview. Samples of most foods and drinks were analysed for fluoride content. Ingestion of fluoride from toothpaste was estimated for each child. Each child's urine was collected over 24 h and analysed for fluoride content. Seventy-eight of the 116 volunteers completed all aspects of the study, which was conducted in 1995-6. For all children, the mean fluoride ingestion from diet was 0.390 (SD 0.122) mg/d or 0.028 (SD 0.008) mg/kg body weight per d. Fluoride ingestion from diet was higher in summer and higher in rural areas. The mean ingestion of fluoride from all sources was 0.426 (SD 0.126) mg/d and the mean fluoride urinary excretion was 0.339 (SD 0.100) mg/d. The difference between ingestion and urinary excretion was +0.087 (SD 0.143) mg, equivalent to 80% excretion. Faecal excretion was not estimated. The results indicate fluoride retention at 4 years to be much lower than previously assumed. PMID- 10703461 TI - Zinc intake and status in Australian vegetarians. AB - Vegetarians have a lower incidence of many chronic diseases than omnivores. However, vegetarian diets could potentially result in lower intakes of some minerals, particularly Zn. In a cross-sectional study, dietary Zn intake was measured using 12 d weighed records in ninety-nine vegetarians (ten vegans) aged 18-50 years and forty-nine age- and sex-matched omnivores. In men, the mean daily Zn intake and Zn density values were similar in omnivores, ovolactovegetarians and vegans, but in women they were significantly lower in vegetarians (mean intake 6.8 mg v. 8.4 mg in omnivores) and few achieved the recommended intake. Significantly more vegetarian than omnivorous women had a daily Zn intake < 6 mg (44% v. 13%). Mean serum Zn concentrations were similar in female omnivores and vegetarians, despite the differences in intake. However, omnivorous men had a lower mean serum Zn concentration (0.85 microgram/ml v. 0.95 microgram/ml) and more subjects had levels below the reference range of 0.72-1.44 micrograms/ml than ovolactovegetarians (P < 0.01). Overall more women than men had low Zn concentrations; and these women generally had intakes below 6 mg/d. There was a significant correlation between serum Zn concentration and dietary Zn density in vegetarians, especially females (P < 0.001), but not in omnivores. Ovolactovegetarians did not have a significantly greater risk of low Zn status than omnivores. PMID- 10703462 TI - Estimation of the energy costs of locomotion in the Iberian pig (Sus mediterraneus). AB - The energy cost of locomotion of four Iberian pigs was measured in two experiments conducted when the animals averaged 41.3 (SE 0.1) kg (first experiment) and 84.1 (SE 0.1) kg (second experiment). The heat production of the pigs was determined when standing or walking at a speed of 0.555 m/s on a treadmill enclosed in a confinement-type respiration chamber, on different slopes (-10.5, 0, and +10.5% in the first experiment, and -5.25, 0 and +10.5% in the second experiment). The energy costs of locomotion, estimated from the coefficients of linear regressions of heat production per kg body weight (BW) on distance travelled, were in the first experiment 2.99, 3.31 and 5.88 J/kg BW per m for -10.5, 0, and +10.5% inclines respectively, and 2.56, 2.84 and 7.13 J/kg BW per m for -5.25, 0 and +10.5% inclines respectively, in the second experiment. The net energy cost of locomotion on the level appeared to be independent of live weight, attaining a value of 2.98 J/kg BW per m. Also, it was found that within experiments the net energy cost of walking on negative slopes was similar to that for locomotion on the level, indicating that no energy was recovered on vertical descent. Mean values were 3.11 and 2.72 kJ/kg BW per m for the light and heavy pigs respectively. The energy cost of raising 1 kg BW one vertical metre was found to be 27.1 J/kg BW per m in the first experiment and 40.0 J/kg BW per m in the second experiment. Correspondingly, the calculated efficiency for upslope locomotion appeared to decline with increasing BW, resulting in average values of 36.2 and 24.5%. PMID- 10703463 TI - Interactions among the branched-chain amino acids and their effects on methionine utilization in growing pigs: effects on nitrogen retention and amino acid utilization. AB - An experiment was conducted to investigate the effects of branched-chain amino acid (BCAA) interactions on their utilization by growing pigs and the effects of excessive amounts of BCAA (leucine, isoleucine, valine) on the utilization of methionine. A semipurified diet containing 100 g crude protein/kg with a balanced amino acid pattern was prepared using casein supplemented with free amino acids. Three further diets were made by reducing the concentration of methionine + cyst(e)ine, valine or isoleucine by 20%. Each of these four diets was then supplemented with leucine (50% excess) or a mixture of BCAA (50% excess of each but excluding the limiting amino acid). All diets were isoenergetic and were made isonitrogenous by replacement of glutamic and aspartic acids. The twelve diets were given to twenty-four growing pigs (30-40 kg) in three periods according to a randomized block design. Each period lasted 8 d and N retention was measured during the last 5 d of each period. Reducing dietary methionine, valine or isoleucine reduced the utilization of N (N retained/N digested) by approximately 20% (P < 0.05). Adding leucine to the isoleucine-limiting diet decreased the utilization of N by 9% (P < 0.05). This was reversed by simultaneous addition of valine. Excess leucine in a valine-deficient diet did not significantly reduce N utilization. In methionine-limiting diets an excess of either leucine alone or of all three BCAA increased the utilization of N by 8% (P < 0.05). PMID- 10703464 TI - Interactions among the branched-chain amino acids and their effects on methionine utilization in growing pigs: effects on plasma amino- and keto-acid concentrations and branched-chain keto-acid dehydrogenase activity. AB - The present experiment was designed to elucidate the mechanism of the methionine sparing effect of excess branched-chain amino acids (BCAA) reported in the previous paper (Langer & Fuller, 2000). Twelve growing gilts (30-35 kg) were prepared with arterial catheters. After recovery, they received for 7 d a semipurified diet with a balanced amino acid pattern. On the 7th day blood samples were taken before (16 h postabsorptive) and after the morning meal (4 h postprandial). The animals were then divided into three groups and received for a further 7 d a methionine-limiting diet (80% of requirement) (1) without any amino acid excess; (2) with excess leucine (50% over requirement); or (3) with excesses of all three BCAA (leucine, isoleucine, valine, each 50% over the requirement). On the 7th day blood samples were taken as in the first period, after which the animals were killed and liver and muscle samples taken. Plasma amino acid and branched-chain keto acid (BCKA) concentrations in the blood and branched-chain keto-acid dehydrogenase (BCKDH; EC 1.2.4.4) activity in liver and muscle homogenates were determined. Compared with those on the balanced diet, pigs fed on methionine-limiting diets had significantly lower (P < 0.05) plasma methionine concentrations in the postprandial but not in the postabsorptive state. There was no effect of either leucine or a mixture of all three BCAA fed in excess on plasma methionine concentrations. Excess dietary leucine reduced (P < 0.05) the plasma concentrations of isoleucine and valine in both the postprandial and postabsorptive states. Plasma concentrations of the BCKA reflected the changes in the corresponding amino acids. Basal BCKDH activity in the liver and total BCKDH activity in the biceps femoris muscle were significantly (P < 0.05) increased by excesses of leucine or all BCAA. PMID- 10703465 TI - The effect of rumen adaptation to oxalic acid on selection of oxalic-acid-rich plants by goats. AB - Rumen microbial degradation is an important route for detoxification of secondary plant compounds encountered in the diets of free-grazing ruminants. Exposure to diets containing particular secondary plant compounds can lead to increased rates of secondary compound degradation in the rumen. An experiment was conducted to determine whether rumen adaptation to oxalic acid would influence the diet selection of goats offered choices between plant species differing in their oxalic acid content. Twelve adult female goats were divided into two groups of six animals each. One group received a daily oral dose, in gelatin capsules, of 0.6 mmol oxalic acid/kg live weight per d throughout the experiment while the other group received placebos consisting of empty gelatin capsules. After an adaptation period of 8 d, the animals were allowed to graze a mixture of spinach (rich in oxalic acid) and cabbage (low in oxalic acid) for 7 h/d on two consecutive days per week during four consecutive 1-week periods. Intervening days were spent on grass pasture. Diet composition and intake were measured using cuticular wax n-alkanes as internal markers. Results showed that adapted goats included a higher proportion of spinach in their diet (P < 0.05) although absolute intakes of spinach were the same for the two groups. Goats in the oxalic acid-adapted group consumed less cabbage than control animals (P < 0.05) suggesting that adaptation to oxalic acid at the rumen level may have interfered with detoxification of cabbage-derived secondary plant compounds. Voluntary intake increased progressively through the four experimental periods (P < 0.001) with a tendency for higher intakes among control than among adapted animals (P < 0.1). The experiment demonstrates how differences in the rate of degradation of secondary plant compounds may influence diet selection in ruminants. PMID- 10703466 TI - Determination of reticulo-rumen and whole-stomach digestion in lactating cows by omasal canal or duodenal sampling. AB - Four ruminally and duodenally cannulated multiparous Finnish Ayrshire cows were fed on diets consisting of grass silage (0.6 kg/kg DM) and one of four concentrates: barley, barley + urea, barley + rapeseed meal and barley + rapeseed cake. The objective of the present study was to compare omasal canal and duodenal digesta flows. Values for digesta flow into the omasal canal and duodenum were determined using a triple-marker method based on Co-EDTA, Yb-acetate and indigestible neutral-detergent fibre (NDF) markers. Microbial non-NH3 N (NAN) flow was assessed by purine flow. Microbial samples to determine the bacterial purine:N ratio were harvested from the rumen, omasum and duodenum. Organic matter flow was significantly lower into the omasum than the duodenum, indicating an endogenous organic matter secretion into the abomasum. In contrast, NDF and acid detergent fibre flows were significantly higher into the omasum indicating digestion of fibre in the omasum. Microbial NAN flows were significantly different (P < 0.001) when estimates were based on bacterial samples harvested from different sites. Differences in total NAN, microbial NAN and dietary NAN flows entering the omasal canal and duodenum were non-significant. The results indicated that the omasal sampling technique provides a promising alternative to the duodenal sampling technique to investigate forestomach digestion in dairy cows and offers an alternative means to study rumen N metabolism. PMID- 10703467 TI - Evidence of progressive deterioration of renal function in rats exposed to a maternal low-protein diet in utero. AB - Intrauterine growth retardation associated with maternal undernutrition is proposed to play a significant role in the aetiology of hypertension and CHD. Animal experiments suggest that the kidney, which is extremely vulnerable to the adverse effects of growth-retarding factors, may play an important role in the prenatal programming of hypertension. Maintenance of renal haemodynamic functions following structural impairment in fetal life is proposed to require adaptations which raise systemic blood pressure and promote a more rapid progression to renal failure. Rats were fed on diets containing 180 g casein/kg (control) or 90 g casein/kg (low protein) during pregnancy. The offspring were studied in terms of blood pressure, creatinine clearance, blood urea N, plasma and urinary albumin, renal morphometry and metabolic activity at 4, 12 and 20 weeks of age. Blood pressure was elevated at all ages in the low-protein-exposed offspring, relative to control rats. Rats (4 weeks old) exposed to the low-protein diet had smaller kidneys which were shorter and wider than those of control animals. Creatinine clearance was significantly reduced in 4-week-old rats exposed to the low-protein diet. Renal morphometry and creatinine clearance at older ages were not influenced by prenatal diet. Blood urea N, urinary output and urinary albumin excretion were, however, significantly greater in low-protein-exposed rats than in control rats at 20 weeks of age. These findings are suggestive of a progressive deterioration of renal function in hypertensive rats exposed to mild maternal protein restriction during fetal life. This is consistent with the hypothesis that adaptations to maintain renal haemodynamic functions following impairment of fetal nephrogenesis result in an accelerated progression towards glomerulosclerosis and increased intrarenal pressures mediated by rising vascular resistance. PMID- 10703468 TI - Effect of thyme oil and thymol dietary supplementation on the antioxidant status and fatty acid composition of the ageing rat brain. AB - The present study measured changes in antioxidant enzyme activity in, and the phospholipid fatty acid composition of the ageing rat brain and tested whether dietary supplementation with thyme oil or thymol could provide beneficial effects. There were significant declines in superoxide dismutase (EC 1.15.1.1) and glutathione peroxidase (EC 1.11.1.9) activities and the total antioxidant status in the untreated rats with age, while thyme-oil- and thymol-fed rats maintained significantly higher antioxidant enzyme activities and total antioxidant status. The proportions of 18:2n-6, 20:1n-9, 22:4n-6 and 22:5n-3 in the brain phospholipids resulting from all three dietary treatments were significantly higher in 28-month-old rats than in 7-month-old rats. Only 20:1n-9 levels in 28-month-old thyme-oil- and thymol-treated rats were significantly higher than in the age-matched control. The proportion of 22:6n-3 in brain phospholipids, which declined with age in control rats, was also significantly higher in rats given either supplement. This latter finding is particularly important as optimum levels of 22:6n-3 are required for normal brain function. These results highlight the potential benefit of thyme oil as a dietary antioxidant. PMID- 10703469 TI - Design and use of internal receiver coils for magnetic resonance imaging. AB - This review describes coils for MRI that are inserted into the body through natural orifices. It covers the design and implementation of small internal receiver coils for use in the pelvis and gastrointestinal tract. Normal anatomy delineated by the high resolution obtained by using these coils and the appearances in a number of disease states for each clinical application are described. PMID- 10703470 TI - A comparison of 14 and 12 gauge needles for core biopsy of suspicious mammographic calcification. AB - This study was carried out to compare the efficacy of 14 vs 12 G needles in stereotactic core biopsy of mammographic calcification. A consecutive series of 100 impalpable mammographic calcifications, without an associated mass and requiring stereotactic core biopsy were randomly allocated to either 14 G or 12 G needle sampling. All biopsies were performed using an upright stereotactic digital unit (Senovision GE) and a Bard automated biopsy gun. Core biopsy results were categorized as either normal, benign, atypical ductal hyperplasia, suspicious of ductal carcinoma in situ (DCIS), DCIS or invasive cancer. The radiographic calcification retrieval rates, complete and absolute sensitivity for malignancy of DCIS and DCIS with an invasive focus were obtained by comparison of core results with surgical histology. Radiographic calcification retrieval was achieved in 86% when using 14 G and 12 G needles. The absolute sensitivity and complete sensitivity for diagnosing DCIS were the same with 12 G and 14 G needles (72% versus 71% and 93% versus 94%, respectively). The use of 12 G needles does not appear to confer benefit over the use of 14 G needles in the diagnosis of mammographic calcification. PMID- 10703471 TI - Differentiation of recurrent rectal cancer and scarring with dynamic MR imaging. AB - The accuracy of dynamic contrast enhanced magnetic resonance (MR) imaging in the differentiation of malignant and benign pelvic lesions during follow-up of patients with treated colorectal tumours was evaluated prospectively. 19 patients (11 men, 8 women; age range 35-70 years; mean 57 years) with suspected local recurrence of colorectal malignancy were evaluated with MR imaging. Dynamic MR imaging with axial Turbo-FLASH gradient echo imaging and bolus injection of contrast medium was performed. Dynamic images, each consisting of one slice in the same location, were acquired at 5, 10, 15, 20 and 30 s, and at 1, 2, 3, 4, 5 and 10 min. The maximum change in signal intensity (Emax), the acceleration rate of the time-intensity curve (TIC) and the ratio of the signal intensity of the lesions to the signal intensity of the iliac artery (SIL/SIA) were used as the enhancement parameters. The TIC and SIL/SIA ratio at 60 s were found to be valuable in the differential diagnosis; Emax had no significance in differentiating benign and malignant lesions. Sensitivity was 83% for each calculated parameter. SIL/SIA has the highest specificity and accuracy among the parameters. PMID- 10703472 TI - Pneumothorax post CT-guided lung biopsy: a comparison between detection on chest radiographs and CT. AB - Pneumothorax is reported to be a more common complication of lung biopsy performed under computed tomography (CT) than under fluoroscopic guidance. This may simply reflect the greater sensitivity of CT over chest radiographs (CXRs) in the detection of small pneumothoraces. This study aimed to determine the incidence of pneumothorax detected by CXR and by CT after CT-guided biopsy of non pleurally based pulmonary masses, and to compare these incidences with previous reports in the literature of pneumothorax incidence post fluoroscopic biopsy. 88 consecutive CT-guided lung biopsies of masses not abutting the pleural surface were included. Immediate post-biopsy CT images, and 1 and 4 h CXRs were assessed independently by two observers for the presence and size of pneumothorax. 72 biopsies were fine needle aspirations (FNAs) performed with 22 G spinal needles only, seven were cutting needle biopsies (CNBs) performed with 18 G cutting needles only, and nine were both. 37 patients (42%) developed a pneumothorax. 35 were detected on CT (40%) and 22 on CXR (25%). None required tube drainage. Of the patients in whom CT demonstrated a pneumothorax, the average depth of this was significantly greater for those in whom CXR also detected a pneumothorax compared with those in whom CXR was negative (7.3 mm versus 3.4 mm, p < 0.05). The incidence of pneumothorax detected on CXR post CT-guided biopsy is similar to the reported incidence post fluoroscopic biopsy. PMID- 10703473 TI - Colour Doppler ultrasound for the evaluation of bowel wall thickening. AB - We performed colour Doppler ultrasound to evaluate bowel wall thickening and to determine the effectiveness of this modality. 42 patients (aged 8-83 years old, mean age 43.5 years) with bowel disease underwent both grey scale and colour Doppler ultrasound examinations. The diagnoses were classified into three categories: inflammation, vasculitis or ischaemia. The bowel wall thickness, wall echotexture and location of the involved portion were all recorded by grey scale ultrasound, while the presence of an intramural colour Doppler flow and arterial signal was evaluated by colour Doppler ultrasound. The colour Doppler flow was graded as "absent", "mild", or "abundant", and the resistive index was also calculated. Bowel wall thickening was observed in the bowel diseases demonstrating inflammation, vasculitis and ischaemia. Patients with ischaemia were significantly older than those with inflammation. The difference in bowel wall thickness was not significant among the disease categories. Differences in bowel wall echotexture, colour Doppler flow, arterial signal and resistive index among the disease categories were significant. The absence of a colour Doppler flow and of an arterial signal suggested ischaemia, while in younger patients, an abundant colour Doppler flow and a stratified echotexture suggested inflammation. The mean resistive index in the ischemic group was significantly higher than that in the inflammatory group. In conclusion, both grey scale and colour Doppler ultrasound are considered to provide useful information for evaluating and differentiating bowel wall thickening in various bowel diseases. PMID- 10703474 TI - High resolution imaging of transitional cell carcinoma with optical coherence tomography: feasibility for the evaluation of bladder pathology. AB - Significant challenges regarding patient morbidity and mortality remain in the management of transitional cell carcinoma (TCC). Among the most important of these challenges is the inability to identify early neoplastic changes and to assess the degree of tumour invasion into the bladder wall in vivo. Optical coherence tomography (OCT) has been recently developed to provide in situ, high resolution, catheter/endoscope based imaging. This study explored the feasibility of OCT for the evaluation of bladder pathology. Both in vitro and in vivo studies were performed. In vitro imaging of pathological human bladder was performed and compared with normal specimens and histopathology. In vivo imaging of normal rabbit bladder was also performed with our current catheter/endoscope based systems. In the in vitro studies, OCT was able to delineate normal microstructure of the bladder, such as the mucosa, submucosa and muscularis layers. This was in contrast to specimens of invasive carcinoma, where a disruption of the normal bladder wall architecture was seen. The in vivo experiment demonstrated current limitations of the catheter/endoscope based systems and provided valuable information for developing an improved system for bladder imaging. The ability of OCT to delineate microstructure of the bladder wall suggests feasibility for endoscopic based imaging. In particular, there is a potential role envisioned for OCT in the management of TCC, identifying pre-malignant states and the depth of tumour invasion. PMID- 10703475 TI - Contrast enhanced dynamic MRI of cervical carcinoma during radiotherapy: early prediction of tumour regression rate. AB - This prospective study investigated the relationship between changes in the MRI dynamic enhancement of cervical carcinoma early during radiotherapy, and tumour regression rate throughout radiotherapy. A total of 36 MRI examinations was performed in seven patients with cervical carcinoma, including a T2 weighted sequence weekly during radiotherapy and also a multislice dynamic Gd-DTPA enhanced sequence before and after the first 2 weeks of radiotherapy. Tumour enhancement was determined on dynamic images using a region of interest and signal-to-noise ratio method. Serial tumour volumes over time on T2 weighted images were estimated using the Cavalieri method of modern design-based stereology to obtain tumour regression rate. It was found that peak and mean enhancement prior to radiotherapy ranged from 3.0 to 13.3, and from 1.9 to 12.2, respectively. After 2 weeks of radiotherapy, peak and mean enhancement ranged from 7.5 to 13.0, and from 6.3 to 10.6, respectively. The change in peak and mean tumour enhancement between dynamic scans ranged, respectively, from -2.0 to 8.4 and from -4.5 to 8.5. Tumour volume decreased exponentially with time (p < 0.01). Tumour regression rates ranged from 2.0% to 15.2% per day, and correlated positively with changes of both peak and mean tumour enhancement (p < 0.01). It is concluded that MRI dynamic enhancement during the first 2 weeks of radiotherapy may provide early prediction of tumour regression rate, and therefore be of value in designing treatment schedules for cervical carcinoma. PMID- 10703476 TI - Patient specific doses used to analyse the optimum dose delivery in barium enema examinations. AB - The exposure and geometrical data for 89 barium enema examination patients were recorded manually in five hospitals in Finland. From the recorded data, organ and primary exit doses as well as effective individual doses were calculated for each patient using the ODS-60 program, which is capable of adjusting the calculation phantom according to a patient's size and sex. The mean (and standard deviation, SD) and median effective individual doses for the patients were 9.3 (5.7) and 6.8 mSv, respectively. Conversion functions from dose-area product to relevant organ doses and to effective individual dose were presented as a function of patient sex and weight. Mean primary exit dose values were calculated for each exposure. These were used to compare a theoretically justified exposure control (EC) function with the function of the automatic exposure (rate) control, AEC (AERC), at different hospitals. According to the analysis of primary exit doses, the implementation of the EC was far from optimum. With EC function proposed in this study the SD of effective individual doses to patients could be lowered considerably. PMID- 10703477 TI - An experimental study of radiation-induced cognitive dysfunction in an adult rat model. AB - The objectives of this study were to establish an adult rat model for the late onset of radiation-induced cognitive dysfunction and to compare behavioural dysfunction with histopathological changes. While under anaesthesia, 30 rats (experimental group) were irradiated with a total dose of 40 Gy, given as eight fractions in 24 days. Another 30 rats (control group) underwent sham irradiation. The cognitive functions of all rats were evaluated at 6, 9 and 12 months after irradiation using the Morris water maze and passive avoidance tasks. Histopathological examination of these rats was carried out after the evaluation of cognitive functions was complete. At 6 and 9 months after irradiation there were no significant differences between the control and irradiated groups in passive avoidance and water maze tests. At 12 months after irradiation, the passive avoidance task revealed a deterioration of cognitive function in the experimental group. Histopathological observations revealed no abnormal findings in the irradiated brains at the light microscope level. Late onset cognitive dysfunction following cranial irradiation was observed in an adult rat model. Pathological investigations showed no abnormalities in the irradiated brains. These findings indicate that radiation-induced cognitive dysfunction can precede morphological changes in the brain or that they arise without them. The present model seems useful for elucidating the pathogenesis of radiation-induced cognitive dysfunction and for developing methods for therapy and prophylaxis. PMID- 10703478 TI - Improved trigeminal and facial nerve tolerance following fractionated stereotactic radiotherapy for large acoustic neuromas. AB - The purpose of this study was to demonstrate improved cranial nerve tolerance following fractionated stereotactic radiotherapy for large acoustic neuromas, defined as tumours with pons-petrous distance (A) > 1 cm and midporous transverse diameter (A + Y) > 2 cm. Of 28 patients with acoustic neuromas treated with fractionated stereotactic radiotherapy, 19 had large tumours at high risk for radiosurgery-induced cranial neuropathy. Six patients received 36 Gy in six, weekly, fractions and 13 patients received 30 Gy in six, weekly, fractions. 15 patients had evaluable trigeminal nerve function and 16 had evaluable facial nerve function. At a median follow-up of 4.5 years, tumour shrinkage was seen in 10 patients and tumour size was stable in nine. None of the patients developed any evidence of transient, or permanent, trigeminal or facial neuropathy at any time during their follow-up period. Fractionated stereotactic radiotherapy may offer a superior therapeutic ratio to single fraction stereotactic radiosurgery in the management of large acoustic neuromas, as evidenced by the absence of post treatment trigeminal and facial neuropathy. PMID- 10703479 TI - Subclavian artery aneurysm with oesophagoarterial fistula. AB - Aneurysms of the subclavian artery are rare. Fistula formation between the subclavian artery and the oesophagus has been described in aberrant subclavian artery and oesophageal foreign body. However, a fistula between a non-aberrant subclavian artery aneurysm and the oesophagus has not been previously reported. In this report, an unusual case of subclavian artery aneurysm with a fistula to the oesophagus causing intractable haematemesis is presented with the angiographic findings. PMID- 10703480 TI - Lung metastasis invading the left atrium--CT diagnosis. AB - A case of metastatic colonic adenocarcinoma invading the left atrium is reported in a patient with clinical signs of cardiac tamponade. The intracavitary extension of the tumour was clearly demonstrated by contrast enhanced CT. As CT plays an important role in the evaluation of patients with intrathoracic masses, intravenous contrast medium is recommended in those cases with associated clinical symptoms of heart disease or pericardial effusion. Its use may establish the diagnosis of cardiac involvement. PMID- 10703482 TI - Ocular metastasis of choriocarcinoma. AB - Metastatic tumours of the eye commonly occur from primaries of the breast or lung. The prognosis is poor and the mean life expectancy is 6.5 months from diagnosis. Although metastatic tumours of the eye are not a rarity, ocular metastasis from choriocarcinoma has not been reported previously. We report a case of gestational choriocarcinoma with ocular metastasis. PMID- 10703481 TI - MR findings in degenerated ovarian leiomyoma. AB - Leiomyoma is one of the rarest solid tumours of the ovary. We report a case of a degenerated ovarian leiomyoma associated with pregnancy. MR findings are identical to those of degenerated uterine leiomyoma and it is difficult to differentiate between them. Ovarian leiomyoma should therefore be included in the differential diagnosis of subserosal uterine leiomyoma. PMID- 10703483 TI - Acute hyperthermia following stereotactic radiosurgery for pituitary adenoma. AB - A 71-year-old male presented with a large pituitary adenoma with superior extension into the optic chiasm and suprasellar cistern. He was treated with stereotactic radiosurgery to a dose of 16 Gy. Approximately 1 h after radiosurgery he developed fever; his temperature peaked at 105.1 degrees F and normalized about 20 h later. This case demonstrates that acute hyperthermia is a potential complication following high dose stereotactic radiosurgery for large pituitary tumours. PMID- 10703484 TI - The risks of treating keloids with radiotherapy. AB - The risk of carcinogenesis from radiation exposure is well known. It has been questioned for some time therefore, whether it is wise to treat benign disease with radiotherapy. We report a case of a patient who developed bilateral breast carcinomas almost 30 years after treatment of chest wall keloids with radiotherapy. There are only anecdotal reports in the literature of malignancies following treatment of keloids with radiotherapy. We review these reports and discuss the safety of this approach to the management of keloid scars. PMID- 10703485 TI - Rhenium-186 HEDP as a boost to external beam irradiation in osteosarcoma. AB - In this case report we demonstrate the usefulness of targeted radiotherapy in the form of rhenium-186 HEDP as a method for dose escalation in the treatment of osteosarcoma. PMID- 10703486 TI - Imaging appearances of Sister Mary Joseph nodule. AB - Sister Mary Joseph nodules are metastatic deposits in the periumbilical area. The purpose of this review is to demonstrate umbilical anatomy, discuss modes of metastatic spread, the various imaging appearances and their relevance to the radiologist. PMID- 10703487 TI - A perianal problem. PMID- 10703488 TI - The two cultures of medicine: objective facts versus subjectivity and values. PMID- 10703489 TI - Medical neutrality. PMID- 10703490 TI - A national database of medical error. PMID- 10703491 TI - Shaken baby (shaken impact) syndrome: non-accidental head injury in infancy. PMID- 10703492 TI - Photodynamic therapy. PMID- 10703493 TI - Herpes simplex virus vectors for gene therapy in Parkinson's disease and other diseases of the nervous system. PMID- 10703494 TI - Variations on the musical brain. PMID- 10703495 TI - The limits of pressure sore prevention. AB - Pressure sore prevalence and incidence were assessed in 275 patients who were admitted to a well-staffed internal medicine ward during a 12-month study period or who were present on day 1. Pressure sore risk was assessed by use of the Braden scale and patients scoring 16 or less were provided with intensive preventive care. During the study period 5.1% (95% confidence interval 2.7-7.8) of 275 patients had pressure sores (prevalence) and 4.4% (1.9-6.9) developed sores (incidence). None of the 239 patients who were assessed as not being at risk developed a sore. 36 patients were assessed as being at risk at some time during the study and 12 of these developed sores despite receiving high-quality preventive care. The results suggest that not all pressure sores can be prevented in severely ill patients. We believe that the 4.4% incidence of sores in this study approaches the current limit of prevention. PMID- 10703496 TI - Acupuncture for vulvodynia. AB - Vulvodynia is the sensation of burning and/or pain of the vulva in the absence of abnormal clinical findings. We offered acupuncture to twelve patients with this syndrome. All had experienced severe distress and impairment of sexual function and usual treatments had failed. The patients attended weekly for acupuncture and progress was monitored at each visit by enquiry, a questionnaire and a visual analogue scale for pain. Half had treatment for the first five weeks only, the other half for the second five weeks only. Side-effects were negligible. Two patients felt so much improved that they declared themselves 'cured'; three believed their symptoms had improved and wished to continue acupuncture; four felt slightly better and judged acupuncture more effective than any other treatment; and three noted no effect at all. Acupuncture is time-consuming and a large part of its beneficial effect in this study may have come from the regular specialist contact. However, in view of the patients' lack of response to other measures their satisfaction with the acupuncture was surprisingly high. PMID- 10703497 TI - Ascites and apparent renal failure treated with a Foley catheter. PMID- 10703498 TI - Deep upper limb and jugular venous thrombosis in dilated cardiomyopathy. PMID- 10703499 TI - Sleep disturbances and cardiac arrhythmia after treatment of a craniopharyngioma. PMID- 10703500 TI - Repair of a massive inguinal hernia. PMID- 10703501 TI - Abdominal neurofibromatosis. PMID- 10703502 TI - Jaundice associated with flavoxate. PMID- 10703503 TI - Genocidal doctors. PMID- 10703504 TI - The health of the Cornish tin miner 1840-1914. PMID- 10703505 TI - [The role of molecular genetic studies in the diagnosis of solid tumors]. PMID- 10703506 TI - [Polychemotherapy-induced changes in the antioxidant state of blood in patients with inoperable cancer of the stomach]. AB - Changes in antioxidant characteristics of blood in patients with inoperable tumors of the stomach were followed before and after combination polychemotherapy which comprised intraperitoneal and endolymphatic injection of drugs. General strengthening therapy was given prior to treatment. It was found that enzymatic activity as a component of antioxidant defenses offered by lymphocytes is low in such patients which adversely affects the structural and functional integrity of lymphocyte membranes. After chemotherapy, said function improved significantly showing a tendency towards normalizing. This effect correlated with clinical improvement and better quality of life in patients with inoperable gastric tumors. PMID- 10703507 TI - [Cytokines in the treatment and prevention of postoperative infectious complications in patients with colorectal cancer]. AB - Surgery for colorectal carcinoma has supplemented the following cytokine-based procedures: (1) extracorporeal immunotherapy (EIT) with autologous rIL2-activated mononuclear cells for treatment of patients with infectious complications, and (2) extracorporeal infusion immunotherapy with native cytokines (perfusate of xenospleen) to prevent postoperative infectious complications. EIT was followed by an effective correction of T-cell immunodeficiency and rehabilitation of monocytic function; rapid decrease in symptoms of endotoxicity; improved response to conventional therapy, and, as a result, a fall in mortality rates (from 38.9 to 11.7%). Immunotherapy with native cytokines brought about a significant decrease in postoperative complication incidence (36.4-15.8%), the average length of stay in hospital falling by 6.8 days. Immunological correction with cytokines was instrumental in raising the efficacy of surgical treatment of colorectal tumors. PMID- 10703508 TI - [EFR-like peptides and their receptors as prognostic factors for the survival of patients with non-small-cell lung cancer]. AB - The levels of EGFR and its ligands have been assayed in tumor and adjacent lung tissues in NSCLC patients. Both EGFR and EGF-like peptides were found in tumor more frequently than in unaltered tissue. It was shown (Kaplan-Meyer) that simultaneous expression of EGFR and its ligands in tumor and adjacent lung tissues was associated with lower overall and relapse-free survival in NSCLC patients. PMID- 10703509 TI - [Incidence and risk factors of papillomavirus infection and cervical dysplasia in sexually active adolescent girls]. AB - A cytological examination of 425 sexually active females aged 13-17 established cervical HPV infection incidence at 30.3 +/- 2.2% while that of cervical dysplasia--4.5 +/- 1.0%. The risk factors for HPV infection included low education and income level (p < 0.05), number of sexual partners exceeding three (p < 0.05), poor hygienic standards of woman (p < 0.05) and those of partner (p < 0.001) and smoking of more than 5 cigarettes per day (p < 0.001). The risk factors of cervical epithelial dysplasia were number of partners of 5 or more (p < 0.001), poor hygienic standards of male partner (p < 0.001) and smoking of 10 cigarettes and more per day (p < 0.001), etc. PMID- 10703510 TI - [The role of tumor-related papillomavirus infection of the genitals in the genesis of background pathology of the ectocervix, dysplasia and preinvasive cancer of the cervix uteri]. PMID- 10703511 TI - [Specific changes in the parameters of blood antioxidant status in patients with cervical carcinoma in the course of cancer therapyi]. AB - Parameters of the blood and plasma antioxidant systems were investigated at different stages of specific therapy for T3NxM0 cervical tumors in 23 patients to identify mechanisms of "relative" drug resistance. Treatment included preoperative telegammatherapy (10 Gy), aregional and regional lymphinfusion with low and high doses of cytostatics, respectively, complemented with endolymphatic injection of tocopherol acetate 100 ng. As a result, surgery after Wertheim became feasible in 14 out of 23 patients following external exposure to 30-40 Gy. Morphological examination detected metastases into the regional lymph nodes in 3 patients. The advantages offered by the treatment modality include adequate response to therapy as well as access to control of free-radical processes taking place in membranes of immunocompetent cells which in turn assures good clinical results. PMID- 10703512 TI - [Nucleolar organizers and mitotic conditions in endometrial hyperplasia and carcinomai]. AB - The study deals with the levels of nucleolar organizers (NO) and mitotic conditions in 85 samples of the endometrium (proliferative stage of menstrual cycle--5; glandular hyperplasia--10; adenomatosis--15; atypical hyperplasia--25 and adenocarcinoma--25). These findings point to a significant increase in NO number in atypical hyperplasia and especially in adenocarcinoma. The latter showed an inverse correlation between the index under study and cell differentiation stage. Endometrial mitosis displayed a higher mitotic index, a larger fraction of pathological mitoses and cells passing through metaphase as well as a variety of pathological forms of karyokinesis. A high correlation between NO number and mitotic index was observed for different conditions of the endometrium. PMID- 10703513 TI - [Sarcoma arising from Langerhans cellsi]. AB - A rare case of tumor arising from Langerhans cells in the tongue and neck area in a 37 year-old man is presented. It was a polymorphocellular sarcoma with bean like twisted nuclei. Electron microscopy identified granules of Langerhans (Bierbeck), multiple tubulo-vesicular structures, ring-like plates, Golgi apparatus, lysosomes and dendritic processes. Total leukocytic antigen was assayed in tumor cells but no expression of S-100 protein found. PMID- 10703514 TI - [Effect of ionizing radiation on functional activity of macrophagesi]. AB - It was shown that in vitro irradiation (8 Gy) of murine peritoneal macrophages suppressed their spontaneous cytotoxity and induced growth-stimulating activity. Exposure to 4 Gy induced mRNA proximal factors--TGF-beta and TNF-alpha and boosted growth-stimulating activity. These effects should be considered when evaluating efficacy of radiotherapy for tumors. PMID- 10703515 TI - [Effect of cycloferon on dissemination of Lewis lung carcinoma in mice and rhabdomyosarcoma ra-23 in rats]. AB - Tumor growth and proliferative activity of tumor cells were suppressed and the number of pulmonary metastases in C57B16 mice decreased 3.3-fold following seven injections of cycloferon (100 mg/kg body) to induce interferon production. Injections were carried out 1-16 days after subcutaneous transplantation of Lewis lung carcinoma. After mice were immunized with ovine red blood cells, cycloferon administration raised thymus-dependent humoral immune response. After eight injections of cycloferon (50 mg/kg body) into rats, from day of intravenous transplantation of rhabdomyosarcoma RA-23 until day 20, no significant effect on metastasizing into the lung was recorded. However, single injection of cyclophosphamide 50 mg/kg inhibited metastasis formation. The highest suppressor effect was registered with combination cycloferon-cyclophophamide treatment: mean weight of metastasis decreased by half, as compared with treatment with cyclophosphamide alone. Both drugs caused karyotypical abnormalities to occur in metastatic cells. Tumor growth and spreading suppression after cycloferon should be attributed to cytotoxic antitumor action, cell proliferation inhibition and immunomodulating effect. PMID- 10703516 TI - [The effect of dexamethasone on the functional features in rats with Pliss lymphosarcoma transplant]. AB - A computer-assisted method of video imaging the adhesive, permeability and thrombogenic properties of microvessels was used to study the effect of dexamethasone on the functional activity of the mesenteric microvascular endothelium in rats with Pliss' lymphosarcoma. Dexamethasone-21 Na-phosphate (0.1 mg/ml) was supplied to the desired mesenteric area and the necessary measurements were carried out. The latter showed that tumor growth was associated with microcirculation disturbances, namely, raised leukocyte adhesion to the mesenteric microvascular endothelium, microvascular permeability and thrombogenicity. Enhanced microvascular sensitivity to dexamethasone pointed to the important role played by changes in the adhesive properties of the endothelium among general paraneoplastic disorders of microcirculation. PMID- 10703517 TI - [The use of Neupogen in prevention of purulent septic complications after radical surgery for esophageal cancer]. AB - To prevent purulent complications, a granulocyte-colony stimulating factor- filgrastim-neupogen (200 or 480 mg) was injected during radical surgery and for the following 6 weeks in 26 patients with esophageal tumors and gastric cancer disseminated to the esophagus. A significant rise in leukocyte levels which potentiated antibacterial therapy was recorded in all patients. Purulent septic complication was registered in one case. PMID- 10703518 TI - [Radio- and thermoradiotherapy of patients with aggressive fibromatosis]. PMID- 10703519 TI - [Preoperative thermoradiotherapy in the combined treatment of rectal tumors is the inferior ampullar segmento]. AB - Data on the examination of 260 radically-treated patients with rectal tumors in the inferior ampullar segment are presented. 107 patients received surgery alone, another 75--preoperative radiotherapy, while still another 78--preoperative thermoradiotherapy. Combined treatment of stage III tumors significantly reduced recurrence incidence and was followed by significant increase in recurrence-free survival rates. In cases of tumor disseminated to the regional lymph nodes, recurrence-free survival was observed after preoperative thermoradiation only. PMID- 10703520 TI - [Comparison of magnetic resonance tomography and transvaginal ultrasonography in the comprehensive diagnosis of endometrial cancere]. PMID- 10703521 TI - [The potential and feasibility of conservative therapy for the early stages of ovarian canceri]. AB - Improvement in techniques of early diagnosis of ovarian cancer (OC) has highlighted the importance of development of methods of conservative surgery. Numerous researchers have failed so far to reach consensus on working out strategies of treatment. A retrospective analysis of clinico-morphological data on 176 patients with bordar-line and OC stage I (conservative surgery--46; radical--130) (1980-1985) showed that efficacy depended on tumor pattern rather than method of treatment. Given high risk of recurrence in unremoved ovary (2 out of 28 reproductive females), bilateral salpingo-oophorectomy without removal of the uterus is discussed as an alternative procedure of conservative therapy. Modern procedures of this therapy requiring accurate staging and monitoring can be carried out at large medical centers only. PMID- 10703522 TI - [Ten-year experience with intraoperative radiotherapy]. AB - An analysis of the end results of combined treatment for stage III non-small cell lung cancer, stage III gastric cancer and soft-tissue sarcoma has been carried out. Radical surgery was supplemented with intraoperative radiation therapy (IORT) with a single dose of 10-20 Gy. The radiation source was an original small size betatron installed in the operating room. IORT did not interfere with wound healing nor did it involve increase in postoperative lethality. The 5-year survival rates for lung cancer patients who received IORT or surgery alone were 34.1 and 24.4%, respectively. The same indices for gastric cancer in IORT patients were 34.2% while in surgical cases--21.7%. Two-year non-relapse survival in patients with soft-tissue sarcomas showed a rise due to reduced recurrence incidence in irradiated areas. PMID- 10703523 TI - [The results of cytologic diagnosis of bone tumors]. AB - The results of bone tumor cytological diagnosis in 968 patients examined and treated at the Institute Clinic (1988-1997) were analyzed. Histological examinations were performed in 479 cases. The share of cytologically inconclusive findings was significantly higher with benign lesions (17.5%) than malignant tumors (9.0%; p(0.05). The sensitivity and specificity of cytological analysis were 94.7 and 93.4%, respectively; positive predictive value--96.7, negative- 89.8 and overall diagnostic accuracy--94.3%. Histological pattern of tumor was correctly identified in 73.7% of malignancies. PMID- 10703524 TI - [Ulrasound diagnosis of melanoma metastasis into the gallbladder]. AB - A clinico-ultrasound description of presentation of melanoma metastases to the gallbladder in 8 patients is discussed. Secondary lesions to the bladder presented as exophytic outgrowths extending from the bladder wall into the lumen. Echo generated by the lesions was of medium intensity and low intensity around the edge of the elastic wall. They revealed dopplergaphic signs of blood flow. Complex application of echo- and dopplergraphy offered a means of differential diagnosis of metastatic lesions and other pathologies of the gallbladder presenting as exophytic outgrowths in the bladder wall in melanoma patients. PMID- 10703525 TI - [Mucinous adenocarcinoma of the prostatel]. PMID- 10703526 TI - [A case of uterine sarcoma in postmenopause]. PMID- 10703527 TI - [Problems of radiotherapy for locally advanced prostate neoplasms]. PMID- 10703528 TI - [Society and cancer. Search for mutual understanding]. PMID- 10703529 TI - [Arkadij Mihailovich Merkov--founder of oncologic statistics in the SSSR the 100th anniversary of his birth]. PMID- 10703530 TI - Trace minerals in human growth and development. AB - Trace mineral deficiencies may affect several biological functions in humans, including physical growth, psychomotor development and immunity. We have reviewed the mechanisms whereby several trace mineral deficiencies may affect these biological functions at different ages (fetal life, infancy, childhood and adolescence), as well as the evidence supporting this association. We describe the effects of zinc deficiency on the hormonal regulation of growth and sexual development in both humans and animal models. We provide data regarding the effects of iron deficiency on growth and psychomotor development. We mention the effects of copper, manganese, selenium and iodine deficiencies on growth and development. We conclude that iron deficiency may affect psychomotor development, but does not appear to affect growth. Zinc deficiency may cause growth retardation and psychomotor delay. PMID- 10703531 TI - Oral health in children and adolescents with IDDM--a review. AB - Children with insulin-dependent diabetes mellitus have a lower salivary flow rate, pH and buffer capacity, but a higher glucose content and peroxidase, IgA, magnesium and calcium concentration, in comparison with healthy children. Nevertheless the incidence of caries is lower than normal in diabetic children with good metabolic control. Periodontal disease usually starts at puberty as mild gingivitis with bleeding and gingival recession, and it may develop into severe periodontitis, especially in children with poor control of diabetes. Microangiopathy, impaired immune response, different bacterial microflora and collagen metabolism are involved in the pathogenesis of diabetic periodontal disease. The gingival flora is mostly composed of Gram-negative, anaerobic bacteria, while collagen has a lower solubility and is atrophic and inadequate to support the occlusion forces. For these reasons, prevention of periodontitis is important in diabetic children; they should receive oral hygiene instruction and visit a dentist at least twice a year. PMID- 10703532 TI - Influence of growth hormone treatment on pubertal timing and pubertal growth in children with idiopathic short stature. Dutch Growth Hormone Working Group. AB - We studied the influence of recombinant human growth hormone (rhGH) on pubertal timing and pubertal growth in children with idiopathic short stature (ISS), and evaluated whether this was different between children with and without intra uterine growth retardation (IUGR). Twenty-six (18 M, 6 IUGR; 'treated') subjects were treated with rhGH (6-7 days/week, dosage: 14-28 IU/m2 per week [i.e. 0.2-0.3 mg/kg per week]). Fifty-eight subjects (31 M, 9 IUGR; 'controls') were not treated. All subjects attained final height. Prepubertal height gain was significantly larger in the treated children compared to control children (M: 0.66 SDS, 95% confidence interval [CI] 0.41 to 0.92; F. 0.92 SDS, CI 0.58 to 1.26). Pubertal height gain, peak height velocity and duration of puberty were similar for the treated and control subjects. rhGH advanced the age at peak height velocity by 0.7 years (CI 0.3 to 1.0) in boys, and the age at onset of puberty by 1.1 years (CI 0.3 to 1.9) in girls. The gain in final height was 2-3 cm. Age and height SDS at start were the most important predictors for pubertal height gain, total height gain and final height in a multivariate regression analysis. Total height gain of treated subjects with IUGR was less than that of treated subjects without IUGR. In conclusion, rhGH did not affect pubertal growth in children with ISS, and slightly improved their final height. rhGH treatment should be started early to improve height as much as possible before the onset of puberty. PMID- 10703533 TI - Endogenous growth hormone secretion does not correlate with growth in patients with Turner's syndrome. Italian Study Group for Turner Syndrome. AB - We investigated in Turner's syndrome patients whether the decrease in growth hormone (GH) secretion is frequent or sporadic, whether or not reduced GH secretion contributes to insufficient growth, and whether age, spontaneous presence of telarche and/or pubarche, karyotype and weight influence GH secretion decrease. We evaluated GH reserve in 301 patients by classical stimulation tests and in 68 of these patients mean nocturnal spontaneous secretion was also measured. Spontaneous telarche and/or pubarche were present in 33% of girls aged > 9 years. In 11% of patients, weight was more than 20% above levels appropriate for height. In 36.2% of patients (low-responders), we observed a reduction of the GH reserve (peak < or = 10 micrograms/l during two stimulation tests). Moreover, we noted reduced mean nocturnal spontaneous secretion (< or = 3.3 micrograms/l) in 61.8% of patients. Karyotype and the presence/absence of spontaneous telarche and/or pubarche did not influence either GH reserve or mean nocturnal spontaneous secretion. GH secretion (both GH reserve and mean nocturnal spontaneous secretion) did not influence height, yet low-responders had a significantly higher chronological age than normal-responders. Obese Turner's girls were low responders and showed reduced mean nocturnal spontaneous secretion more frequently than normal weight girls; body mass index was significantly higher in patients with reduced GH secretion when compared to patients with normal GH secretion. We conclude that impairment of GH secretion is frequent in Turner's syndrome patients, especially if obese; that GH secretion impairment is not related to karyotype or spontaneous telarche and/or pubarche; that GH secretion is irrelevant to growth in these, patients and, therefore, its evaluation is unnecessary. PMID- 10703534 TI - Insulin-like growth factor-I (IGF-I) and IGF-binding protein-3 (IGFBP-3) concentrations compared to stimulated growth hormone (GH) in the evaluation of children treated for malignancy. AB - OBJECTIVE: The aim of this investigation was to evaluate the utility of IGF-I and IGFBP-3 determinations in screening for GH deficiency (GHD) in children previously submitted to treatment for childhood malignancy. PATIENTS AND METHODS: We compared the GH responses to two pharmacological tests (arginine and levo dopa) with the IGF-I and IGFBP-3 levels in 48 patients (29 boys) who had undergone bone marrow transplantation (BMT) (36 patients) or treatment for a solid cranial tumor (12 patients). RESULTS: 22 patients (45.8%) showed GHD (i.e. GH peak < 8 ng/ml in both tests), and only three (13.6%) of the GHD patients had concomitant low IGF-I levels (i.e. -2 SD below the normal mean) and only one (4.5%) an abnormal IGFBP-3 value (i.e. -2 SD below the normal mean). Among the 26 children with normal GH secretion, 21 (80.8%) also showed normal IGF-I and IGFBP 3 levels, three (11.5%) had a concomitant low IGF-I value and two (7.7%) a concomitant low IGFBP-3 value. A significant correlation was found between GH secretion and age at diagnosis (r = 0.26, P < 0.05), and between IGF-I and IGFBP 3 (r = 0.52, P < 0.0001), but not between GH and IGF-I or IGFBP-3. Comparing the growth pattern of these patients from diagnosis to the first year after therapy or BMT, we found that while individual height changes did not correlate with the GH peak, a significant correlation was found between height SDS decrease and IGF I (r = 0.31, P < 0.05) or IGFBP-3 SDS (r = 0.37, P < 0.01). CONCLUSION: Our results indicate that the cut-off of -2 SD for IGF-I and IGFBP-3 was insensitive in screening for GHD. A normal value did not exclude a subnormal GH response to provocative tests and therefore although IGF-I and IGFBP-3 levels may be indicators of the growth pattern, they cannot be used alone as a tool for identifying GHD children after treatment for childhood malignancy. PMID- 10703535 TI - Leptin in African-American children. AB - Leptin, the protein product of the obesity gene, produced by adipose tissue, regulates body weight and energy expenditure through CNS feedback mechanisms. In obesity, leptin levels are elevated suggestive of leptin resistance. Because of increased prevalence of obesity in African-Americans, the aim of this study was to assess leptin and its relationship to adiposity in African-American children. We measured plasma leptin levels in 42 African-American children (23 M, 19 F), age 11.8 +/- 0.3 yr, and compared them with 30 American-White children matched for age, body composition and puberty. Body composition was assessed by bioelectrical impedance and plasma leptin by RIA. Data are presented as means +/- SEM and statistical significance is implied by p < 0.05. There was no racial difference in plasma leptin levels (Blacks: 9.8 +/- 1.6, Whites 9.8 +/- 1.9 ng/ml). Leptin correlated with %BF in Black (r = 0.75, p = 0.005) and White (r = 0.79, p = 0.005) children. There were no gender or puberty related differences in leptin levels in African-American children. We concluded that leptin levels are comparable between African-American and American White children of similar body composition. The major determinant of serum leptin levels in these children is degree of adiposity with no gender or puberty related differences. Longitudinal studies are needed to assess leptin's role during puberty in both genders. PMID- 10703536 TI - Nodular goiter and thyroid carcinoma in children and adolescents in a moderate endemic area (lower Silesia-Sudeten endemia) in the last twelve years. AB - Although thyroid carcinoma is more common in the adult population, the risk of a nodule being malignant is greater in children. The aim of our present investigation was to ascertain the percentage of malignancy in nodular goiter observed in patients from the Lower Silesia region in the last 12 years. The examination included 60 children (12 boys and 48 girls) treated in our clinic from 1987 to June 1998. Age varied from 7 to 18 years (mean 14.8 +/- 2.4), most of them in the age group between 13 and 18 years. The following investigations were performed: TSH, T3, T4, thyroid ultrasonography, fine needle aspiration biopsy and Tc99 scintigraphy of the thyroid. Most of the patients were euthyroid; two children demonstrated pressure symptoms. All the patients were treated by operation. Histological examinations revealed the following: nodular goiter in 19 patients, cystic nodular goiter in 5, follicular adenoma in 20, fetal adenoma in 3, nodular goiter and follicular adenoma in 6, papillary carcinoma in 6, and follicular carcinoma in 1 patient. We concluded that an increased incidence of thyroid cancer has been noted in children with nodular goiter in Lower Silesia during the last 12 years. Thyroid cancer was observed mostly in patients with single nodules and was associated with a high risk of malignancy. PMID- 10703538 TI - Pregnancy outcome in patients with insulin dependent diabetes mellitus and diabetic nephropathy treated with ACE inhibitors before pregnancy. AB - The preconception and intrapregnancy parameters that are relevant to outcome in women with insulin dependent diabetes mellitus (IDDM) and diabetic nephropathy remain controversial. We analyzed the types and frequencies of maternal and neonatal complications in 24 IDDM patients with diabetic nephropathy (24 pregnancies), all with preserved to mildly impaired renal function. All patients received treatment with captopril for at least six months prior to planned pregnancy and were maintained under strict glycemic control from at least three months before pregnancy to delivery. A successful pregnancy outcome (live, healthy infant without severe handicaps two years after delivery) was observed in 87.5% of the patients. Preexisting hypertension was the only parameter found to be significantly predictive of an unsuccessful outcome (p = 0.0004). We conclude that in patients with IDDM complicated by diabetic nephropathy, pre-pregnancy captopril treatment combined with strict glycemic control offers a prolonged protective effect against possible renal deterioration and probably improves pregnancy outcome. PMID- 10703537 TI - Mass screening program for congenital hypothyroidism in south-eastern Poland. AB - The success of a screening program depends in great measure on effective organization. The purpose of the screening program is the commencement of treatment of selected patients within two weeks of life. In total in our laboratory we have tested 461,479 newborns during 1985-1996. The introduction of bar codes in 1995 (recommended by the Institute of Mother and Child in Warsaw) improved our work in the laboratory and departments of obstetrics and reduced the possibility of errors. The screening tests for congenital hypothyroidism appear to serve the monitoring of iodine deficiency disorders. The good organization of the screening program is indispensable and needs permanent control. PMID- 10703539 TI - Glutamic acid decarboxylase and islet cell antibodies in healthy Estonian children. AB - The prevalence of antibodies to the 65 kDa isoform of glutamic acid decarboxylase (GADA) was compared with that of islet cell antibodies (ICA) in 614 non-diabetic Estonian children (314 males) aged 3-18 years representing the general population. GADA were analyzed with a radioligand assay, and ICA with a standard immunofluorescence method with a detection limit of 2.5 Juvenile Diabetes Foundation (JDF) units. Fourteen subjects (2.3%, 95% confidence interval [CI] 1.1 3.5%) tested positive for GADA (median level 10.8 relative units [RU], range 7.7 154.2 RU), while 10 (1.6%, CI 0.6-2.6%) had ICA (median levels 34 JDF units, range 3-97 JDF units). Five subjects (0.8%, CI 0.1-1.5%; p = 0.03 vs GADA and 0.15 vs ICA) were double positive. The individual with the second highest GADA level (129.3 RU) and the highest ICA level (97 JDF units) presented with type 1 diabetes 4 months later. A follow-up sample was obtained approximately 3-4 years after the first sampling in 14 subjects initially positive for ICA and/or GADA. Four of the nine initially ICA-positive children remained positive, but their levels decreased from a median of 42 to 18 JDF units (p = 0.06). Only two of the nine retested subjects initially positive for GADA remained positive in the second sample. These observations suggest that the prevalence of GADA in non diabetic children is of the same magnitude as that of ICA. Combined positivity for both GADA and ICA is less prevalent than single antibody specificities, indicating that double autoantibody positivity may have a higher predictive value for future type 1 diabetes in the general population than either antibody separately. The evanescent character of diabetes-associated autoantibodies in a proportion of the unaffected children implies that more subjects may experience self-restricted beta-cell damage than the number progressing to actual disease. PMID- 10703540 TI - Efficacy of long-term growth hormone treatment in Turner's syndrome. European Study Group. AB - The efficacy of long-term growth hormone treatment and the optimal treatment modalities in Turner's syndrome are still controversial. Studies have shown widely divergent results. While there is still need for results of randomized controlled trials, large uncontrolled trials remain a valuable source of preliminary information. This study of 136 girls with Turner's syndrome (TS) from the European Lilly Turner Study shows that the treatment regimens followed by girls with TS who start GH treatment at a relatively late age (12.9 +/- 2.2 yr) lead at best to a small average gain in final height. Mean gain over initial projected final height was 3.7-4.7 cm, depending on the chosen Turner-specific height-for-age reference data. There was a considerable variation in gains over initial projected final height (range -10.6 to +17.2 cm), which may be partly explained by the existence of good and bad responders. Whether there is a genetic/molecular basis for this and how responsiveness can best be predicted remains an important challenge. PMID- 10703541 TI - Spatial clustering of childhood IDDM prevalence in Slovakia 1985-95: a socio economic issue? PMID- 10703542 TI - Tamoxifen treatment of progressive precocious puberty in a patient with McCune Albright syndrome. AB - Treatment of progressive precocious puberty in patients with McCune-Albright syndrome (MAS) has traditionally been with aromatase inhibitors, such as testolactone. However, the use of these agents has been characterized by problems with both efficacy and compliance. We report a case of MAS in which tamoxifen proved to be a successful alternative in the treatment of progressive precocious puberty. An African-American female presented with MAS at 2-5/12 years. Frequent menses, skeletal maturation and growth acceleration prompted initiation of therapy with testolactone at 22 mg/kg/d. Over the next 13 months, the patient's puberty advanced unchecked, despite progressive increases in the dose of testolactone. At age 4 years, medication was discontinued due to treatment failure. At 4-6/12 years, bone age was 10 years, predicted adult height was 137 cm, and monthly bleeding continued. Tamoxifen was then begun on an experimental basis. In response, the patient experienced immediate cessation of menses, and had an abrupt decrease in the rates of pubertal progression and linear growth. This patient has now been maintained on tamoxifen for over three years with no apparent adverse effects. GnRH analogue therapy was begun when the onset of central precocious puberty was noted. Predicted adult height has improved to 154 cm and growth velocity and skeletal maturation remain stable. Our results suggest that tamoxifen may have a valuable role in the treatment of precocious puberty in patients with MAS and may lead to superior results compared with those achieved with aromatase inhibitors. PMID- 10703543 TI - Gender identity reversal in an adolescent with mixed gonadal dysgenesis. AB - We describe a patient who was assigned female at birth because of genital ambiguity without performing further diagnostic procedures and presented at the age of 13-1/2 years because of her strong desire to change her legal sex. Karyotype was 46,XY; clinical, endocrinological, radiological and surgical work up revealed hypergonadotropic hypogonadism and mixed gonadal dysgenesis. Gender identity reversal was performed after extensive psychological testing and adaptation of living circumstances resulting in a successful integration as a male with normal psychological and social functioning. In several surgical procedures, the streak gonad, the nonfunctional testis, and the rudimentary uterus were removed, and a penis was reconstructed from a penisoid with chorda and hypospadias. Our patient supports the idea that gender identity is imprinted prenatally by hitherto poorly understood mechanisms and that sex assignment in infants with ambiguous genitalia needs careful consideration of not solely endocrinological and anatomical data. PMID- 10703544 TI - Ketoacidosis and hyperosmolarity as first symptoms of type 1 diabetes mellitus following ingestion of high-carbohydrate-containing fluids. AB - The concomitant occurrence of diabetic ketoacidosis and hyperosmolarity is reported in two children, as early symptoms of misdiagnosed type 1 diabetes mellitus. The precipitating factor for both severe metabolic abnormalities was the ingestion of a large amount of high-carbohydrate-containing fluids, a few days before admission. A similar situation has never been reported before in the literature. A successful therapeutic scheme is described. PMID- 10703545 TI - Severe hypoglycemia and reduction of insulin requirement in a girl with insulin dependent diabetes mellitus: first sign of a craniopharyngioma. AB - A girl with a history of insulin-dependent diabetes mellitus since 5.5 years, and Hashimoto's thyroiditis since 12 years, developed episodes of severe hypoglycemia from the age of 12 years. This was associated with falling insulin requirements, from 0.78 U/kg/day at 11 years to 0.34 U/kg/day at 16 years. At 16 years she was found to have GH, gonadotropin, ACTH, and probably also TSH deficiency with hyperprolactinemia. MRI scan revealed a cystic intrasellar craniopharyngioma with moderate suprasellar extension. In spite of cortisol replacement at 17 years, insulin requirement fell further to 0.25 U/kg/day at 18 years. In this girl, decreasing insulin requirements represented an early manifestation of combined growth hormone and cortisol deficiency. PMID- 10703546 TI - Special issue on gene conservation: identification and management of genetic diversity. PMID- 10703547 TI - Preserving genes, species, or ecosystems? Healing the fractured foundations of conservation policy. AB - The scientific foundations of conservation policy are the subject of a recent tripolar debate, with systematists arguing for the primacy of phylogenetic rankings, ecologists arguing for protection at the level of populations or ecosystems, and evolutionary biologists urging more attention for the factors that enhance adaptation and biodiversity. In the field of conservation genetics, this controversy is manifested in the diverse viewpoints of molecular systematists, population biologists, and evolutionary (and quantitative) geneticists. A resolution of these viewpoints is proposed here, based on the premise that preserving particular objects (genes, species, or ecosystems) is not the ultimate goal of conservation. In order to be successful, conservation efforts must preserve the processes of life. This task requires the identification and protection of diverse branches in the tree of life (phylogenetics), the maintenance of life-support systems for organisms (ecology), and the continued adaptation of organisms to changing environments (evolution). None of these objectives alone is sufficient to preserve the threads of life across time. Under this temporal perspective, molecular genetic technologies have applications in all three conservation agendas; DNA sequence comparisons serve the phylogenetic goals, population genetic markers serve the ecological goals, quantitative genetics and genome explorations serve the evolutionary goals. PMID- 10703548 TI - First policy then science: why a management unit based solely on genetic criteria cannot work. AB - In contrast to the goals of the symposium from which this series of papers originated, we argue that attempts to apply unambiguously defined and general management unit criteria based solely on genetic parameters can easily lead to incorrect management decisions. We maintain that conservation genetics is best served by altering the perspective of data analysis so that decision making is optimally facilitated. To do so requires accounting for policy objectives early in the design and execution of the science. This contrasts with typical hypothesis testing approaches to analysing genetic data for determining population structure, which often aspire to objectivity by considering management objectives only after the analysis is complete. The null hypothesis is generally taken as panmixia with a strong predilection towards avoiding false acceptance of the alternative hypothesis (the existence of population structure). We show by example how defining management units using genetic data and standard scientific analyses that do not consider either the specific management objectives or the anthropogenic risks facing the populations being studied can easily result in a management failure by losing local populations. We then use the same example to show how an 'applied' approach driven by specific objectives and knowledge of abundance and mortality results in appropriate analyses and better decisions. Because management objectives stem from public policy, which differs among countries and among species groups, criteria for defining management units must be specific, not general. Therefore, we conclude that the most productive way to define management units is on a case-by-case basis. We also suggest that creating analytical tools designed specifically to address decision making in a management context, rather than re-tooling academic tools designed for other purposes, will increase and improve the use of genetics in conservation. PMID- 10703549 TI - Genetic structure and gene flow among Komodo dragon populations inferred by microsatellite loci analysis. AB - A general concern for the conservation of endangered species is the maintenance of genetic variation within populations, particularly when they become isolated and reduced in size. Estimates of gene flow and effective population size are therefore important for any conservation initiative directed to the long-term persistence of a species in its natural habitat. In the present study, 10 microsatellite loci were used to assess the level of genetic variability among populations of the Komodo dragon Varanus komodoensis. Effective population size was calculated and gene flow estimates were compared with palaeogeographic data in order to assess the degree of vulnerability of four island populations. Rinca and Flores, currently separated by an isthmus of about 200 m, retained a high level of genetic diversity and showed a high degree of genetic similarity, with gene flow values close to one migrant per generation. The island of Komodo showed by far the highest levels of genetic divergence, and its allelic distinctiveness was considered of great importance in the maintenance of genetic variability within the species. A lack of distinct alleles and low levels of gene flow and genetic variability were found for the small population of Gili Motang island, which was identified as vulnerable to stochastic threats. Our results are potentially important for both the short- and long-term management of the Komodo dragon, and are critical in view of future re-introduction or augmentation in areas where the species is now extinct or depleted. PMID- 10703550 TI - Genetic structure of harbour porpoise Phocoena phocoena populations in the northwest Atlantic based on mitochondrial and nuclear markers. AB - The harbour porpoise, Phocoena phocoena, experiences high levels of nonnatural mortality owing to interactions with commercial fisheries throughout its range. To accurately evaluate the significance of this bycatch, information on population structure is required. We have examined the population structure of this species in the northwest Atlantic Ocean using mitochondrial DNA (mtDNA) sequence and nuclear microsatellite data. Samples from four previously proposed summer breeding populations--the Gulf of Maine, eastern Newfoundland, the Gulf of St Lawrence and West Greenland--were analysed. Control-region sequences revealed a significant partitioning of genetic variation among most of these summer populations, indicating that northwest Atlantic harbour porpoises should not be considered one panmictic population. Analysis of females alone yielded the highest levels of population subdivision, suggesting that females are more philopatric than males. At least three management units may be defined for harbour porpoises in the northwest Atlantic based on these data. Analysis of six microsatellite loci failed to detect significant population subdivision. Male mediated gene flow may maintain homogeneity among nuclear loci, while female philopatry is sufficient to produce a signal of population subdivision in the maternally inherited mtDNA genome. mtDNA analyses also indicate that winter aggregations of harbour porpoises along the US mid-Atlantic states comprise animals from more than one summer breeding population. PMID- 10703551 TI - Genetic structure of fragmented populations of red squirrel (Sciurus vulgaris) in the UK. AB - The relationships among 207 squirrels from 12 locations in the UK and three in mainland Europe were examined using mitochondrial DNA (mtDNA) control region sequence. Twenty-six haplotypes were detected, many of which were population specific. Eighty per cent of the populations analysed contained two or more haplotypes. Hierarchical analysis of molecular variance showed the majority of genetic variation to be partitioned among populations. Genetic diversity varied considerably within the UK, and conformed to no obvious geographical trend. The populations in Argyll and Spadeadam Forest showed the highest levels of variation in the UK. However, the greatest genetic diversity was seen in Bavaria, southern Germany where six unique alleles were detected in a sample of 10 individuals. Phylogenetic analysis revealed no evolutionary divergence between UK and mainland European haplotypes. We conclude that, within the UK, the genetic patterns observed are most likely to be explained by the effects of genetic drift which has occurred since the isolation of populations during the past few hundred years, hence we cannot detect any underlying phylogeographic pattern. Therefore, the use of larger, geographically distinct populations within the UK for augmentation of small isolated populations is unlikely to pose problems of genetic incompatibility. Further, the role that demographic factors may have in complicating the application of current genetically based management unit criteria is likely to need further attention. PMID- 10703552 TI - Intraspecific phylogeography of Lasmigona subviridis (Bivalvia: Unionidae): conservation implications of range discontinuity. AB - A nucleotide sequence analysis of the first internal transcribed spacer region (ITS-1) between the 5.8S and 18S ribosomal DNA genes (640 bp) and cytochrome c oxidase subunit I (COI) of mitochondrial DNA (mtDNA) (576 bp) was conducted for the freshwater bivalve Lasmigona subviridis and three congeners to determine the utility of these regions in identifying phylogeographic and phylogenetic structure. Sequence analysis of the ITS-1 region indicated a zone of discontinuity in the genetic population structure between a group of L. subviridis populations inhabiting the Susquehanna and Potomac Rivers and more southern populations. Moreover, haplotype patterns resulting from variation in the COI region suggested an absence of gene exchange between tributaries within two different river drainages, as well as between adjacent rivers systems. The authors recommend that the northern and southern populations, which are reproductively isolated and constitute evolutionarily significant lineages, be managed as separate conservation units. Results from the COI region suggest that, in some cases, unionid relocations should be avoided between tributaries of the same drainage because these populations may have been reproductively isolated for thousands of generations. Therefore, unionid bivalves distributed among discontinuous habitats (e.g. Atlantic slope drainages) potentially should be considered evolutionarily distinct. The DNA sequence divergences observed in the nuclear and mtDNA regions among the Lasmigona species were congruent, although the level of divergence in the COI region was up to three times greater. The genus Lasmigona, as represented by the four species surveyed in this study, may not be monophyletic. PMID- 10703553 TI - Disparate phylogeographic patterns of molecular genetic variation in four closely related South American small cat species. AB - Tissue specimens from four species of Neotropical small cats (Oncifelis geoffroyi, N = 38; O. guigna, N = 6; Leopardus tigrinus, N = 32; Lynchailurus colocolo, N = 22) collected from throughout their distribution were examined for patterns of DNA sequence variation using three mitochondrial genes, 16S rRNA, ATP8, and NADH-5. Patterns between and among O. guigna and O. geoffroyi individuals were assessed further from size variation at 20 microsatellite loci. Phylogenetic analyses using mitochondrial DNA sequences revealed monophyletic clustering of the four species, plus evidence of natural hybridization between L. tigrinus and L. colocolo in areas of range overlap and discrete population subdivisions reflecting geographical isolation. Several commonly accepted subspecies partitions were affirmed for L. colocolo, but not for O. geoffroyi. The lack of geographical substructure in O. geoffroyi was recapitulated with the microsatellite data, as was the monophyletic clustering of O. guigna and O. geoffroyi individuals. L. tigrinus forms two phylogeographic clusters which correspond to L.t. oncilla (from Costa Rica) and L.t. guttula (from Brazil) and which have mitochondrial DNA (mtDNA) genetic distance estimates comparable to interspecific values between other ocelot lineage species. Using feline-specific calibration rates for mitochondrial DNA mutation rates, we estimated that extant lineages of O. guigna diverged 0.4 million years ago (Ma), compared with 1.7 Ma for L. colocolo, 2.0 Ma for O. geoffroyi, and 3.7 Ma for L. tigrinus. PMID- 10703554 TI - Captive breeding, reintroduction, and the conservation genetics of black and white ruffed lemurs, Varecia variegata variegata. AB - A character-based phylogenetic species concept approach was used to examine conservation unit status for three wild populations of black and white ruffed lemurs, Varecia vareigata variegata, from Betampona (N = 3), Manombo (N = 6), and Ranomafana (N = 14), Madagascar. Population aggregation analysis was performed on 548 bp from the control region (D-loop) of the mitochondrial DNA (mtDNA). Twenty one diagnostic sites were found to differentiate the Betampona (northern) population from the Manombo/Ranomafana (southern) populations. Additionally, individuals from the North American captive population (N = 11) and from Parc Ivoloina, Madagascar (N = 6) were examined for the same mtDNA fragment. The captive animals more closely resembled the southern populations and the Parc Ivoloina animals were more similar to the northern population. However, the inclusion of these ex situ animals reduced the number of diagnostic sites differentiating the northern and southern populations. Our genetic data were used to assess the ongoing management strategy for reintroducing individuals into the Betampona population and for introducing new founders into the ex situ population. This study demonstrates the utility of combining genetic information with a consideration of conservation priorities in evaluating the implementation of management strategies. PMID- 10703555 TI - Molecular genetic analysis among subspecies of two Eurasian sturgeon species, Acipenser baerii and A. stellatus. AB - Two species, the Siberian sturgeon, Acipenser baerii, and stellate sturgeon, A. stellatus, were studied using mitochondrial DNA (mtDNA) (D-loop, cytochrome b (cyt-b) and ND5/6 genes) sequencing to determine whether traditionally defined subspecies correspond to taxonomic entities and conservation management units. Initially, several mtDNA regions for each taxon (A. baerii: 737 bp D-loop, 750 bp ND5, 200 bp ND6, and 790 bp cyt-b; A. stellatus: 737 bp D-loop and 600 bp ND5) were examined. The D-loop was the most variable region and was sequenced for 35 A. baerii and 82 A. stellatus individuals. No fixed, diagnostic differences were found between any of the subspecies. Geographical structuring of haplotypes was observed within A. baerii, and gene flow estimates suggest isolation of the A. baerii baicalensis subspecies and the Yenisie and Lena River populations. No intraspecific subdivisioning was found within the genetic data for A. stellatus. The use of the phylogenetic criterion (fixed diagnostic differences) for identifying conservation units is compared to the rationale and results of other methods. Overall, morphologically and geographically based subspecies designations within Acipenseridae may not directly correspond to the biological entities appropriate for management and should not be used for conservation programmes without genetic support. PMID- 10703556 TI - Microsatellite diversity and conservation of a relic trout population: McCloud River redband trout. AB - Rainbow trout native to the McCloud River, California, USA (Oncorhynchus mykiss stonei) are thought to represent a relic, nonanadromous trout adapted to harsh, fragmented environments. These fish, commonly named McCloud River 'redband' trout, survive in their most primitive form in a small, spring-fed stream, Sheepheaven Creek, in the upper McCloud River drainage. Turn-of-the-century fisheries records document both coastal anadromous steelhead and freshwater resident trout within the McCloud River drainage. The phylogenetic position of the McCloud River redband trout within O. mykiss has been debated for over 50 years. Based on phenotypic evidence, these fish were first reported as 'southern Sierra golden trout' by Wales in 1939. Behnke (1970) considered them a relic subspecies of nonanadromous, fine-scaled trout. Allozyme and mitochondrial DNA evidence suggested a coastal lineage. In this study, we examined within- and among-basin genetic associations for Sheepheaven Creek redband trout using 11 microsatellite loci. Within-basin analyses supported unique genetic characteristics in Sheepheaven Creek's trout in comparisons with other McCloud River rainbow trout. Microsatellite data supported significant independence between Sheepheaven Creek fish and hatchery rainbow trout. Inter-basin genetic distance analyses positioned Sheepheaven Creek fish with samples collected from Lassen Creek, a geographically proximate stream containing inland redband trout. California's redband trout shared a close genetic association with Little Kern River golden trout (O.m. whitei) and isolated rainbow trout from Rio Santo Domingo, Baja, Mexico (O.m. nelsoni), suggesting a vicariant distribution of microsatellite diversity throughout the southern range of this species. PMID- 10703557 TI - Marine population structure in an anadromous fish: life-history influences patterns of mitochondrial DNA variation in the eulachon, Thaleichthys pacificus. AB - Due to the apparent decline in size of a number of populations, eulachon, Thaleichthys pacificus, have recently become the focus of a conservation movement in the northeast Pacific. Little is known of the marine life-history phase of this anadromous fish, and although it has been suggested that eulachon spawning in different rivers may form distinct populations, nothing is known of their population structure. Molecular genetic data were used to investigate population structure and possible management schemes. Mitochondrial DNA genotypes, determined through restriction fragment length polymorphisms (RFLP) analysis, were resolved in fish from several rivers throughout the geographical range of eulachon. Our data support the idea that extant eulachon populations result from postglacial dispersal from a single Wisconsinan glacial refuge. Further, while three of the 37 haplotypes recovered account for approximately 79% of the samples, many private haplotypes were observed, suggesting possible regional population structure. While a great deal of genetic variation was observed (37 haplotypes in 315 samples), an AMOVA showed that > 97% of the total variation was detected within populations. As yet, it is unclear whether genetically distinct populations of eulachon exist, or if these fish may be treated as one or a few large populations. Results were tested against predictions made from hypotheses concerning the origin and persistence of subdivided populations in marine species, and seem to be more consistent with the Member-Vagrant hypothesis than isolation by distance. Eulachon present an interesting situation that illustrates the difficulties involved in defining management units in organisms with high levels of gene flow. PMID- 10703558 TI - A study of stress in medical students at Seth G.S. Medical College. AB - BACKGROUND: It is usually observed that medical students undergo tremendous stress during various stages of the MBBS course. There is a high rate of suicide among them. METHODS: To determine incidence of stress and factors controlling stress in medical students at various stages of MBBS course at Seth G S Medical college, 238 students (First year 98, Second 76, Third 64) were asked to complete a questionnaire on personal data (gender, stay at hostel, mode of travel, time spent in travel every day, medium of study in school, place of school education.), Stress inducing factors, Zung's depression scale, ways of coping, stress relievers, perceived social support and personality type. Statistical tests used were ANOVA, critical ratio and Student's 't' test. RESULTS: Majority of medical students (175/238--73%) perceived stress. Stress was found to be significantly more in Second and Third MBBS students rather than First MBBS levels (p < 0.05). Stress was not found to differ significantly on the basis of sex, stay at hostel, model of travel, time spent in travel every day, medium of study in school, place of school education. Stress was found to be significantly more in students having more than 95% of marks at 12th Standard as compared to others. Academic factors were greater perceived cause of stress in medical students. There was no significant difference in the students at different levels of MBBS regarding academic factors and social factors as a stress inducing factors. Physical factors were found to be significantly more in Second and Third MBBS students as compared to First MBBS students. Emotional factors were found to be significantly more in First MBBS students as compared to Second & Third MBBS students. Stress was more common in medical students who have dominant strategy of coping as positive reappraisal, accepting responsibility and planful problem solving. Stress was less common in medical students at Seth G S Medical College who have dominant strategy of coping as escaping and distancing from difficult situation. Family and Friend as perceived social supports were more in Second MBBS than First MBBS medical students. Stress was not found to be significantly more in students having their personality factor contributing to stress (Type A- 52/67) as compared to others (Type B--123/171). This indicates that the stress was not trait oriented but was process oriented (p = NS). CONCLUSION: Stress in medical students is common and is process oriented. It is more in second and third year. Academic factors are greater perceived cause of stress in medical students at Seth G S medical college. Emotional factors are found to be significantly more in First MBBS. It is dependent on person's ways of coping and social support. PMID- 10703559 TI - Bacterial profile and antimicrobial susceptibility pattern in catheter related nosocomial infections. AB - This prospective study was carried out over a period of 6 months in the Paediatric Intensive Care Unit (PICU) of a tertiary care teaching hospital. The aim of the study was to determine the organisms causing catheter related nosocomial infections in the PICU and to study their antimicrobial susceptibility pattern. Patients with endotracheal intubation, indwelling urinary catheters and central venous catheters (CVC)/venous cutdown catheters were included in the study. Colonization of the endotracheal tube, urinary catheter related infections (UCRI) and colonization of the CVC/venous cutdown catheters was studied. E. coli was the commonest organism colonizing the endotracheal tube tip with maximum susceptibility to cefotaxime and amikacin. E. coli was also was the commonest organism causing UCRI with maximum susceptibility to nitrofurantoin and amikacin. Acinetobacter was the commonest organism colonizing the CVC/venous cutdown catheters with maximum susceptibility to ciprofloxacin. All these sites of catheter related infections considered together, E. coli and Klebsiella were the commonest nosocomial organisms. Both had maximum susceptibility to amikacin. Methicillin resistant Staphylococcus aureus (MRSA) was isolated only from one culture. All the organisms had a poor susceptibility to cefazolin and amoxycillin. A knowledge of the resident microbial flora and their antimicrobial susceptibility pattern is necessary for formulating a rational antibiotic policy in an ICU. PMID- 10703560 TI - Traumatic transverse fracture of sacrum with cauda equina injury--a case report and review of literature. AB - Fractures of the sacrum are rare and generally associated with fracture of the pelvis. Transverse fractures of the sacrum are even less frequent and neurological deficit may accompany these fractures. A case of transverse fracture sacrum with cauda equina injury treated by sacral laminectomy and root decompression, is reported. PMID- 10703561 TI - Jejunal angiomatoses causing small bowel obstruction in a patient with Down syndrome: a case report. AB - Gastrointestinal vascular anomalies are extremely uncommon. We describe a patient with Down syndrome who presented with acute abdominal pain due to a mixed capillary and venous vascular malformation involving the proximal jejunum. PMID- 10703562 TI - Circumcaval ureter. AB - We report a case of circumcaval ureter diagnosed preoperatively by 'fish-hook' appearance on intravenous pyelogram. At surgery, patient was treated by 'Anderson Hones' pyeloplasty leaving the retrocaval segment in-situ. PMID- 10703563 TI - Anaesthetic management of bilateral alveolar proteinosis for bronchopulmonary lavage. AB - The most hazardous manifestation of pulmonary alveolar proteinosis is progressive hypoxia for which bronchopulmonary lavage (BPL) is the single most effective treatment. Unfortunately this procedure under general anesthesia itself increases the risk of hypoxia due to the need for one lung ventilation. It was therefore considered interesting to report the successful anaesthetic management of a patient with pulmonary alveolar proteinosis for Bronchopulmonary lavage. PMID- 10703564 TI - Renal arterial aneurysm--an incidental finding at autopsy. AB - Herein we describe a rare case of saccular renal artery aneurysm seen as an incidental autopsy finding in an elderly, hypertensive female. The aneurysm was seen as a small exophytic mass with calcified wall and lumen occluded by recanalized thrombus. PMID- 10703565 TI - Coronary artery disease in Asian Indians. AB - Coronary Heart Disease should new be considered an important public health problem in India. It is a part of the epidemiological transition characterized by changing lifestyles and a probable genetic predisposition. The interplay of factors with regards to their existence, causality and attributable weight-age needs to be understood in the context of management of an individual patient as well as strategic planning for control and prevention. PMID- 10703566 TI - Non enzymatic glycosylation of alpha-1-proteinase inhibitor of human plasma. AB - Human plasma contains inhibitors, which control the activity of proteolytic enzymes. Alpha-1-proteinase inhibitor and alpha-2-macroglobulin are two of them present in high concentration in human plasma, which inhibit action of trypsin among other proteinases. The trypsin inhibitory capacity (TIC) of human plasma is observed to be decreased in pathological conditions like diabetes mellitus. The mechanisms of decrease in TIC was due to nonenzymatic glycosylation of alpha-1 proteinase inhibitor (A1PI). A1PI was partially purified from normal human plasma by steps involving ammonium sulphate precipitation, DEAE Sepharose CL6B chromatography, Concanavalin A Sepharose Chromatography and Sephadex G-100 Gel filtration. Purified inhibitor was glycosylated in vitro by incubating it with varying glucose concentrations, under nitrogen for different periods of time in reducing conditions. After glycosylation, the molecular weight of inhibitor increased from 52 kDa to 57 KDa because of binding with glucose molecules. The percent free amino groups in the protein decreased with increasing glucose concentration and days of incubation. The TIC of such modified inhibitor decreased significantly. Decrease in TIC was dependent on the glucose concentration and period of incubation used during in-vitro glycosylation of native inhibitor. PMID- 10703567 TI - Urinary catheter related nosocomial infections in paediatric intensive care unit. AB - The present prospective study was carried out in the Paediatric Intensive Care Unit (PICU) of a tertiary care teaching hospital in Mumbai. The objective was to determine the incidence, risk factors, mortality and organisms responsible for urinary catheter related infections (UCRI). Colonization and/or bacteriuria was labelled as urinary catheter related infection (UCRI). Forty-four patients with 51 urinary catheters were studied. Incidence of UCRI was 47.06%. Age, female sex and immunocompromised status did not increase the risk of UCRI. Duration of catheter in-situ and duration of stay in the PICU were associated with higher risk of UCRI. The mortality was not increased by UCRI. Commonest organism isolated in UCRI was E. coli, which had maximum susceptibility to nitrofurantoin and amikacin. PMID- 10703569 TI - Difficult intubation in a case of ankylosing spondylitis: a case report. AB - A case of severe ankylosing spondylitis involving the entire spine was to be operated for lumbar osteotomy. She had fixed rigidity of the cervical spine with minimal rotational movement, inability to lie down supine and severe restrictive lung disease with hypoxemia (pO2 = 65 mmHg). An awake intubation was performed and the patient was operated under general anaesthesia in the prone position. Intraoperative "wake-up" test was performed to judge whether extent of straightening was excessive. Postoperatively, she was electively ventilated and extubated uneventfully after 24 hours. PMID- 10703568 TI - Primary linitis plastica of the rectosigmoid in a thirteen year old boy. AB - Childhood malignant neoplasms of the gut are extremely rare. The reported incidence of colorectal cancers in patients under 20 years of age is 1 in 10 million. The low index of suspicion for this tumour in children results in advanced disease at diagnosis and subsequently a poor prognosis. A rare case of a primary linitis plastica of the rectum occurring in a 13 year old boy is reported with review of pertinent literature. PMID- 10703570 TI - Plasma cell leukaemia--a report of two cases. AB - Two cases of plasma cell leukaemia--a rare form of leukaemia are described. Both cases presented with anaemia and hepatosplenomegaly. Investigations revealed leucocytosis with increased plasma cells (> 20%). Skeletal survey revealed a few osteolytic lesions in both cases. PMID- 10703571 TI - Poor insight in schizophrenia: neurocognitive basis. AB - Poor insight in schizophrenia has been recently thought to be a reflection of prominent and enduring neurocognitive impairments. Reports supporting this theory have implicated prefrontal and parietal lobe functions, among other parameters. The results of other studies have negated the role of neuropsychological abnormalities in poor insight. The analogy between poor insight in schizophrenia and anosognosia in neurological illness as proposed by one set of workers has been elucidated in this review and it appears quite promising. However, the drawing of definite conclusions from all this work has been deferred by us, because of the need for more uniform and standardized methodologies for research on the subject. Nevertheless, attempts to improve the cognitive processes, which affect insight in schizophrenia, may be done to gain better treatment outcome in this disorder. PMID- 10703572 TI - Comparison of two different approaches for internal jugular vein cannulation in surgical patients. AB - We compared the anterior approaches of internal jugular venous cannulation in 200 surgical patients, vis-a-vis the ease of cannulation and threading, number of attempts required and the incidence of complications following each route. The technique of posterior approach used in this study was found to have a higher rate of success in cannulation and lower rate of complication such as carotid puncture. The posterior approach was also a safe alternate route in obese or short necked patients. PMID- 10703573 TI - Anaerobic bacteraemia: a review of 17 cases. AB - Of 93 blood cultures received with a suspicion of anaerobic bacteraemia over a period of two years, only 17 (18.3%) showed anaerobic growth. Twelve grew anaerobes alone while five had a polymicrobial flora. Seven of these patients (4.3%) had pre-existing heart disease while others had history of prior surgery, diabetes mellitus or urinary tract infection. Oropharynx was the commonest portal of entry, followed by gastrointestinal tract. The anaerobes isolated were anaerobic streptococci, Bacteroides fragilis group and Bilophila and Eubacterium species. Fifteen patients developed major complications such as congestive cardiac failure, systemic embolisation, and perforative peritonitis. The mortality rate among the cases with anaerobic bacteraemia was 23.5% in this study. PMID- 10703574 TI - Effect of forced breathing on ventilatory functions of the lung. AB - Ventilatory functions were studied in 36 male and 35 female subjects (mean age 18.5 years), who underwent six weeks course in forced breathing. Ventilatory functions were studied in the form of Forced Vital Capacity (FVC), Forced Expiratory Volume at the end of one second as % of FVC (FEV1%), Maximum Voluntary Ventilation (MVV), Peak expiratory flow rate (PEFR) and Breath Holding Time. Some of these ventilatory functions were found to be increased after a course of forced breathing. PMID- 10703575 TI - Successful management of spontaneous pneumothorax during general anaesthesia in a patient with eosinophilia. AB - A 10-year-old male patient posted for left elbow arthrolysis developed pneumothorax during general anaesthesia. He had history of upper respiratory tract infection and high eosinophil count, which remained high in spite of treatment. In such patients, it is advisable to use steroid pre-operatively & intraoperatively to produce transient eosinopenia so that complications of eosinophilia are avoided. PMID- 10703576 TI - Malignant melanotic neuroectodermal tumour of infancy affecting the occipital squama. AB - An unusual case of a melanotic neuroectodermal tumour of the occipital squama, which underwent malignant transformation in a nine-month-old infant is reported and pertinent literature reviewed. PMID- 10703577 TI - Bilateral glaucomatocyclitic crisis in a patient with Holmes Adie syndrome. AB - A patient with pre-existing bilateral tonic pupils presented with simultaneous bilateral glaucomatocyclitic crisis. Deep tendon reflexes were absent although they were documented to be present 6 years ago. A possibility of a progressive autonomic dysfunction in both these conditions is discussed. PMID- 10703578 TI - Haemangiopericytoma of kidney: a report of 2 cases. AB - Haemangiopericytoma is a rare neoplasm of the kidney. There are no unique radiological or clinical identifiers that can aid in preoperative diagnosis. Surgery is the only reliable therapy, as both chemotherapy and radiotherapy have proven ineffective in several series. The outcome is difficult to predict, the only reliable predictor is presence or absence of metastasis. The rarity of this lesion prompts the report of these two cases. PMID- 10703579 TI - Acute otitis media in children--treatment options. PMID- 10703580 TI - Preparation of a potent anti-scorpion-venom-serum against the venom of red scorpion (Buthus tamalus). AB - A number of children and adults, especially pregnant women succumb to the sting by red Scorpion (Buthus tamalus) in Konkan region--particularly on the coastal line. No specific antiserum or any other antidote is available to treat a victim of scorpion bite and hence the need to prepare a potent antiserum. Red Scorpion (B. tamalus) venom is a mixture of a number of protein moieties and neurotoxins of low molecular weight. Therefore, the venom is poor in antigenic composition and it is difficult to get antibodies specific to neutralise lethal factor/factors. Using Bentonite as an adjuvant and extending the period of immunization a potent antiserum has been prepared capable of neutralising the lethal factor/factors. In vivo testing carried out in albino mice, guinea pigs, dogs and langurs confirms this finding and shows that the antiserum is quite effective in neutralising the scorpion venom to save the life of envenomated animals. PMID- 10703581 TI - Effect of honey on multidrug resistant organisms and its synergistic action with three common antibiotics. AB - A total of 15 bacterial strains (7 Pseudomonas & 8 Klebsiella species) isolated from various samples which showed multi-drug resistance were studied to verify in vitro antibacterial action of honey on the principle of Minimum Inhibitory Concentration (MIC) & its synergism with 3 common antibiotics--Gentamicin, Amikacin & Ceftazidime. The MIC of honey with saline for both organisms was found to be 1:2. The synergistic action was seen in the case of Pseudomonas spp. and not with Klebsiella spp. PMID- 10703582 TI - Pancreatic carcinoid: an unusual tumour in an uncommon location. AB - Primary pancreatic carcinoid is an extremely rare pancreatic neoplasm. It differs from other primary pancreatic tumours in cytoarchitecture, immunocytochemistry and biologic behaviour. Recognition of this rare entity is of vital importance having considerable therapeutic and prognostic implications. We report a case of an exophytic, pancreatic body carcinoid tumour in a man who presented with abdominal pain. The diagnosis was established by histopathological examination of the core biopsy specimen. A surgical resection of the lesion was done successfully and the patient made a satisfactory recovery from the operation. PMID- 10703583 TI - Cavitary pulmonary infarction--a rare cause of spontaneous pneumothorax. AB - A 14-year old female was admitted to the hospital with a diagnosis of resolving myocarditis and dilated cardiomyopathy. She developed spontaneous right-sided pneumothorax. Autopsy, revealed rupture of cavitary pulmonary infarction to be the cause of the pneumothorax, a rare finding. PMID- 10703584 TI - Cri-du-chat syndrome: clinical profile and prenatal diagnosis. AB - Prenatal diagnosis of cri-du-chat syndrome is described in 2 pregnancies. In Case 1, the mother was a balanced translocation carrier and had 2 previously affected off springs. Prenatal diagnosis by chorion villus sampling and cordocentesis was successful in diagnosing an affected conceptus and the pregnancy was electively terminated. Case 2 was referred for nonimmune foetal hydrops and cordocentesis revealed deletion 5p. This second case was noteworthy for the fact that deletion 5p has not been reported to cause foetal hydrops. PMID- 10703585 TI - [Radiation-induced arterial disease]. AB - Arterial occlusive disease developed after radiation therapy in three patients: a woman aged 56 had a sensation of heaviness in her right arm and bluish-black discolorations of fingers 3 years after radiation therapy for breast carcinoma, and two men aged 46 and 45 years had intermittent claudication 23 and 14 years, respectively, after radiation therapy for testicle malignancy. After creation of a bypass, the symptoms subsided. Radiation-induced arterial disease is a rare complication after radiotherapy and usually presents after a latent period of several years. Arterial lesions induced by radiotherapy may be distinguished from atherosclerotic lesions by their atypical localization, limited to the irradiated sites, and occurrence at an earlier age. Symptoms may be masked by pain, limitation of motion, nerve root damage and lymphoedema and may therefore not always be recognized. Indications for treatment are identical to those for atherosclerotic occlusive disease, but due to increased risk of restenosis, rethrombosis and graft infection, percutaneous transluminal angioplasty (with or without stent), endarterectomy or venous bypass surgery should be preferred to prosthetic bypass material. PMID- 10703586 TI - [Why is malaria in Vietnam under control, but Africa is threatened with a malaria disaster?]. AB - In Africa malaria parasites are increasingly developing resistance to the 3 affordable and tolerable drugs: chloroquine, amodiaquine and sulfadoxine pyrimethamine. Alternative products are much more expensive and more toxic. A malaria disaster is looming. On the contrary, in Vietnam a disaster appears to have been averted. Data on malaria epidemiology, on the mosquito, the parasite and the host, man, give insight into the differences and the possibilities of control. Artemisinin derivatives can play an important role in malaria control, also in Africa. Without improvement of care which will require considerable investment and attention, the prospects are bleak. PMID- 10703587 TI - [Deep venous thrombosis of the arm: etiology, diagnosis and treatment]. AB - Thrombosis of the upper extremity is frequently (30-52%) related to the use of an indwelling venous catheter, but it can also occur in healthy individuals after exercise. In the past it was considered a relatively benign thrombotic event, which was treated conservatively, sometimes even without anticoagulant therapy. Recent studies have shown that complications of deep venous thrombosis of the upper extremity occur frequently: pulmonary embolism (8-36%), recurrence thrombosis after cessation of anticoagulant treatment (2-15%) and post-thrombotic syndrome (up to 50%). Therefore when thrombosis of the upper extremity is clinically suspected, it should be objectively diagnosed by compression echography followed if negative by phlebography, with anticoagulant treatment directly afterward, preferably with low-molecular heparin and then acenocoumarol or phenprocoumon. PMID- 10703588 TI - [A new option for skin rejuvenation: CO2-laser resurfacing]. AB - Resurfacing denotes mechanical, chemical or physical removal of the superficial layers of aging skin, after which collagenesis is reactivated with improved epidermal architecture as a result. CO2 laser resurfacing is a modern technique for the treatment of skin aging. With CO2 laser resurfacing it is possible to perform accurate ablation of layers of the skin. Besides the cosmetic indications, this technique is a good treatment modality for a broad spectrum of dermatological diseases. PMID- 10703589 TI - [Gonadotrophin-releasing hormone antagonists: application in ovary-stimulating and sex-steroid dependent disorders]. AB - The hypothalamic gonadotrophin-releasing hormone (GnRH) stimulates synthesis and secretion of luteinizing hormone (LH) and follicle stimulating hormone (FSH) by the gonadotrophic cells of the pituitary. The mechanisms of action of GnRH antagonists and of agonists is completely different. Due tot competitive blockage of GnRH receptors by antagonist administration, LH (and to a lesser extent FSH) levels drop rapidly. Moreover pituitary function normalizes immediately following cessation of medication. The direct and rapid action of GnRH antagonists, the dose dependent suppression of LH and FSH and the rapid restoration of hypophyseal function after cessation of the use of antagonists may shorten and simplify in vitro fertilization, with less chance of side effects or complications. Further studies are required to decide whether antagonists can usefully be applied for other gynecological indications such as the polycystic ovary syndrome. The possibilities of profitable long term treatment will increase considerably if it proves possible to develop a sustained action formulation. PMID- 10703590 TI - ['Trigger thumb' in 38 children: good short-term and long-term results from surgery]. AB - OBJECTIVE: To determine the direct and long-term effectivity of incision of the pulley in a trigger thumb (tendovaginitis stenosans). DESIGN: Retrospective study with follow-up. METHODS: In the period 1984-1995, 38 children (24 boys and 14 girls) were diagnosed and operated on 45 trigger thumbs in the Onze Lieve Vrouwe Gasthuis (Amsterdam) and Medisch Centrum Alkmaar, the Netherlands. Data were obtained from notes, operation reports and review in follow-up, at least 2 years after the operation. RESULTS: The mean age of the children at the moment of surgery was 3 years and 2 months (range: 11 months-10.33 years). 39 out of 45 thumbs were reviewed. Four thumbs had limited postoperative function. Two of these had a revision operation. There was 1 thumb with a postoperative superficial infection. At follow-up all thumbs had maximal function. The nodule in the tendon, which was palpable in 43 out of 45 thumbs preoperatively, had (almost) disappeared in all 39 thumbs at follow-up. There was a family history of trigger thumb in 33% of the 33 patients with follow up. 18% had bilateral involvement of the thumbs. The 6 digits not included in follow-up had a normal function according to the last notes. CONCLUSION: The results of surgery in the short term are good, in the long term excellent. Few complications occur. Based on the findings, it seems advisable to operate on children with a trigger thumb if there is no spontaneous recovery within half a year. Trigger thumb is the result of a congenital tight pulley. PMID- 10703591 TI - [Experiences of guardians of psychogeriatric patients regarding decision not to treat]. AB - OBJECTIVE: To obtain an impression of how representatives of psychogeriatric patients experience their role in non-treatment decisions. DESIGN: Retrospective. METHOD: The names of the 50 most recently deceased patients in psychogeriatric departments of a nursing home and of their representatives were obtained from the register of the nursing home. Of the representatives 28 (67%) agreed to an interview. RESULTS: 24 of the 28 respondents had participated in some form of medical decision-making. There were 34 decisions of which 33 non-treatment decisions. In most cases the decision was to give symptomatic treatment only. In 27 of 34 cases the representative did not regard it as a burden to be involved in the decision-making, on the contrary they considered it self-evident and were pleased to be involved. CONCLUSION: In general, representatives of psychogeriatric nursing home patients did not regard it as a burden to be involved in end-of-life decision-making. PMID- 10703592 TI - [Increase of malaria in the Dominican Republic]. AB - In November 1999 nine European cases of malaria caused by Plasmodium falciparum reportedly were imported by tourists from the Dominican Republic. The incidence of malaria has increased since the hurricane George and since building activities for tourist centres have favoured growth of the mosquito population. Travellers to the Dominican Republic are advised to take precautions against mosquitoes and to take proguanil prophylaxis also malaria should be considered in patients with fever who have visited the area, if only as tourists. PMID- 10703593 TI - [Clinical thinking and decision making in practice. A nurse with acute pain between shoulder blades]. PMID- 10703594 TI - [Clinical thinking and decision making in practice. A nurse with acute pain between shoulder blades]. PMID- 10703595 TI - [No Friday afternoon peak in the number of patients referred to the emergency room at De Weezenlanden Hospital of Zwolle, May/June 1997]. PMID- 10703596 TI - [Laparoscopic inguinal hernia operation]. PMID- 10703597 TI - [A dispute about the concept of 'disease' in Dutch Journal of Medicine, 1863]. PMID- 10703598 TI - [Transmission of hepatitis B virus by a surgeon]. PMID- 10703599 TI - [Cervical smears taken by physicians' assistants are of lesser quality than smears taken by family physicians, but almost as good as the national average]. PMID- 10703600 TI - [Comments on separate consultations for second opinions]. PMID- 10703601 TI - Heterogeneity of Taiwan's indigenous population: possible relation to prehistoric Mongoloid dispersals. AB - Taiwan's 9 indigenous tribes (Tsou, Bunun, Paiwan, Rukai, Atayal, Saisiat, Ami, Puyuma, Yami) are highly homogeneous within each tribe, but diversified among the different tribes due to long-term isolation, most probably since Taiwan became an island about 12,000 years ago. Homogeneity of each tribe is evidenced by many HLA A,B,C alleles having the world's highest ever reported frequencies, e.g. A24 (86.3%), A26 (18.8%), Cw10 (36.8%), Cw7 (66%), Cw8 (32.1%), B13 (27.9%), B62 (37.4%), B75 (18%), B39 (53.5%), B60 (33.3%), and B48 (24%). Also, all of these tribes have HLA class I haplotype frequencies greater than 10%, with A24-Cw7-B39 in Saisiat (44.5%) being the highest, suggesting Taiwan's indigenous tribes are probably the most homogeneous ( the "purest") population in the world. A24-Cw8 B48, A24-Cw10-B60 and A24-Cw9-B61 found common to many Taiwan indigenous tribes, have also been observed in Maori, Papua New Guinea Highlanders, Orochons, Mongolians, Inuit, Japanese, Man, Buryat, Yakut, Tlingit, Tibetans and Thais. These findings suggest Taiwan's indigenous groups are more or less genetically related to both northern and southern Asians. Principal component analysis and the phylogenetic tree (using the neighbor-joining method) showed close relationship between the indigenous groups and Oceanians. This relationship supports the hypothesis that Taiwan was probably on the route of prehistoric Mongoloid dispersals that most likely took place along the coastal lowland of the Asian continent (which is under the sea today). Cultural anthropology also suggests a relationship between Taiwan's indigenous tribes and southern Asians and to a lesser extent, northern Asians. However, the indigenous groups show little genetic relationship to current southern and northern Han Chinese. PMID- 10703602 TI - Analysis of tumor necrosis factor-alpha promoter polymorphism in type 1 diabetes: HLA-B and -DRB1 alleles are primarily associated with the disease in Japanese. AB - Polymorphisms in the 5'-flanking region of the tumor necrosis factor (TNF)-alpha gene were examined to study the genetic background of type 1 diabetes in Japanese. Five different biallelic polymorphisms were examined in 136 type 1 diabetic patients and 300 control subjects. The frequencies of individuals carrying TNF-alpha-857T allele (designated as TNFP-D allele) or -863A/-1,031C allele (designated as TNFP-B allele) were significantly increased in the patients as compared with the controls. Since these TNF-alpha alleles are in linkage disequilibria with certain DRB1 and HLA-B alleles, two-locus analyses were carried out. The TNFP-D allele did not increase the risk in either the presence or absence of the DRB1*0405 or HLA-B54 allele, while the DRB1*0405 and HLA-B54 alleles per se could confer susceptibility in both the TNFP-D allele-positive and -negative populations. Moreover, an odds ratio was remarkably elevated in the population carrying both DRB1*0405 and HLA-B54. Similarly, the TNFP-B allele did not show significant association with the disease in either the HLA-B61-positive or -negative population, while the HLA-B61 allele could significantly increase the risk in the TNFP-B allele-positive population. These data suggest that the associations of TNFP-D and -B alleles may be secondary to their linkage disequilibria with the susceptible HLA class I and class II alleles. Because HLA B and DRB1 genes were independently associated, both of these genes may be contributed primarily to the pathogenesis of type 1 diabetes in Japanese. PMID- 10703604 TI - Molecular modeling of the minor histocompatibility antigen HA-1 peptides binding to HLA-A alleles. AB - Mismatch of the minor histocompatibility antigen HA-1 has been shown to correlate with graft-versus-host disease in HLA-matched sibling marrow transplants. The HA 1H peptide (VLHDDLLEA) that generates this response is known to be presented by HLA-A*0201. In order to understand the interaction of HA-1 peptides with other HLA-A alleles, we have used the LOOK interface to construct molecular models of both HA-1H peptide (VLHDDLLEA) and HA-1R peptide (VLRDDLLEA) binding with 103 HLA A alleles. The results show that in addition to A*0201, 21/103 other HLA-A alleles should be able to bind HA-1H peptide but not HA-1R peptide. Based on the modeled predictions, HLA alleles can be categorised into 4 groups with respect to their interaction with HA-1 peptides: Group 1 - bind HA-1H peptide but not HA-1R peptide; Group 2 - bind HA-1R peptide but not HA-1H peptide; Group 3 - bind both HA-1H and HA-1R peptides; Group 4 - bind neither peptide. These predicted binding patterns of HA-1 peptides will be useful as an aid for defining a wider pool of HLA-A alleles in which HA-1 disparities among donor-recipient pairs can be investigated. PMID- 10703603 TI - Association of DRB1*1501 with disseminated Mycobacterium avium complex infection in North American AIDS patients. AB - The HLA class II allele, DR2 (DRB1*1501), has been repeatedly found to be associated with development of tuberculosis and leprosy. We searched for associations of these and other class II alleles with disseminated Mycobacterium avium complex infection (DMAC) in North American Caucasian homosexual AIDS patients. Molecular typing for HLA-DRB1 and -DQB1 alleles in 176 cases of DMAC and 176 matched controls showed an association of accelerated onset of disease with DRB1*1501 (and the closely linked DQB1*0602) that was stronger upon adjustment for the degree and duration of CD4+ cell deficiency (P=0.04) and in multivariate analysis (P=0.02) than in unadjusted analysis. A similar trend was seen with DRB1*0701, and no other allele showed a relationship of similar magnitude. M. avium complex organisms may more effectively evade host defenses in individuals carrying an HLA polymorphism identical to that associated with M. tuberculosis and M. leprae. PMID- 10703605 TI - Identification of the null HLA-A2 allele, A*0232N. AB - We have identified a null HLA-A*02 allele, HLA-A*0232N, by using a combination of serology, flow cytometry, polymerase chain reaction using sequence-specific primers (PCR-SSP) and full-length sequencing. The null HLA-A2 allele was identified in an Asian individual originally typed by serology as an apparently homozygous HLA-A3, B51. Subsequent genotyping by PCR-SSP identified the genotype as HLA-A*0201, *0301, B*51, Cw*1402. The serological type and lack of detectable HLA-A2 was confirmed using monoclonal antibody typing reagents. Flow cytometry studies failed to identify any cell surface HLA-A2 expression on the patient's peripheral blood lymphocytes. Genotyping using a PCR-SSP set designed to detect null alleles revealed the mutation had not been previously described. Full-length sequencing of the allele identified an allele which was subsequently named HLA A*0232N. This allele is identical to HLA-A*0201 except for a novel point mutation (T for C) at position 493 which creates a premature stop codon. The sequencing enabled the development of a monospecific A*0232N PCR-SSP reaction which was used to screen 973 DNA samples: no further examples of A*0232N were identified. PMID- 10703606 TI - Presence of the DRB4*0103102N null allele in different DRB1*04-positive individuals. AB - The DRB4 gene encoding the DR53 antigen is present in DRB1*04-, DRB1*07- and DRB1*09-positive individuals. Eight allelic variants of DRB4 have been recognized, 5 resulting in an expressed DR53 antigen and 3 belonging to the null alleles. So far the DRB4*0103102N null allele had been found exclusively in individuals carrying the haplotype DR7,-DQ9. High-resolution typing of HLA class II by polymerase chain reaction using sequence-specific primers (PCR-SSP) and/or sequence-based typing of kidney patients and their families revealed the presence of the DRB4*0103102N null allele segregating with DRB1*04 and DQB1*03 in 4 different families. Three different haplotypes on which the null allele was located, were recognized by family studies: DRB1*0401, DQB1*0301; DRB1*0402, DQB1*0302 and DRB1*0404, DQB1*0302. Determination of the DR53 specificity of antisera reacting with DR53-positive individuals has always been difficult due to the simultaneous presence of DR4, 7 or 9. Identification of DR4-positive DR53 negative individuals as described here, provided the serological reactions with DR53-antisera and revealed the antibody specificities in the antisera used. PMID- 10703607 TI - NRAMP1 gene polymorphisms in patients with rheumatoid arthritis. AB - Rheumatoid arthritis (RA) is a chronic inflammatory joint disease associated with HLA-DR genes that share amino acid sequence motif QKRAA/QRRAA from position 70 to 74 in the third hypervariable region of DR1 molecule. The contribution of HLA in RA is however about 37%, suggesting a role for other genes. One such candidate is the gene that encodes natural resistance-associated macrophage protein (NRAMP1), which plays a crucial role in inflammation and tissue destruction. In the present study, we examined the role of NRAMP1 gene polymorphisms in susceptibility to RA. The results show that variation at position 543 in exon 15, which involves substitution of negatively charged aspartic acid (D) by uncharged asparagine (N), and the deletion of TGTG in the 3' UTR may confer protection from development of RA. PMID- 10703608 TI - The relative frequencies of HLA-DRB1*01 alleles in the major US populations. AB - The frequencies of alleles in the HLA-DRB1*01 family were determined in each of five US populations from a database of 82,979 individuals. Individuals typed as DR1 (or DRB1*01) comprised between 7.6%-21.3% of the individuals in each population group. Fifty-nine DR1 individuals were randomly selected from each group and subjected to high-resolution DNA typing by polymerase chain reaction using sequence-specific oligonucleotide probes. DRB1*0101 was the most common allele in the Caucasian, Asian/Pacific Islander, and Native American groups while the DRB1*0102 allele was found in the majority of African Americans and Hispanics. DRB1*0103 was present at a similar frequency in all populations. DRB1*0104, DRB1*0105, and DRB1*0106 alleles were not observed. PMID- 10703609 TI - HLA-DRB and -DQB1 polymorphism in the Macedonian population. AB - HLA-DRB1, DRB3/4/5 and DQB1 polymorphism has been studied in a population of 80 unrelated healthy Macedonians using molecular methods. Twenty-five different DRB1 alleles were identified of which DRB1*1104, *1501, *1601, and *1101 were found most frequently. Among the 15 identified DQB1 alleles, two were predominant: DQB1*0301 and *0502. The most frequent three-locus haplotypes were DRB1*1104 DRB3*02-DQB1*0301 (18%/), DRB1*1101-DRB3*02-DQB1*0301 (9%) and DRB1*1601-DRB5*02 DQB1*0502 (10%). Polymorphism for DRB1*04, *13 and *15 haplotypes was extensive. Eleven different DR2-related haplotypes were found, some of which were unusual for European populations: DRB1*1501-DRB5*0102-DQB1*0502, DRB1*1501-DRB5*02 DQB1*0502, DRB1*1501-DRB5*0102-DQB1*0601. PMID- 10703610 TI - Strong association between HLA-Cw*0706 and HLA-B*44032 in the Bubi population from Equatorial Guinea. AB - Unrelated Bubi, native to the island of Bioko (Equatorial Guinea), were previously typed by low-resolution polymerase chain reaction using sequence specific primers (PCR-SSP) and serology for HLA-A, -B and -C. HLA-B*44 was found frequently and associated with Cw*07. We have studied the HLA subtypes of 20 B*44pos/Cw*07pos Bubi individuals. HLA-B and -C were typed by sequencing exons 2 and 3. To distinguish the alleles Cw*1701/02/03, Cw*07011/012/06 and Cw*1801/02 additional sequencing of exon 1 or 5 was performed. All 20 B*44pos/Cw*07pos individuals of the Bubi population were typed Cw*0706 positive. Nineteen of them carried the B*44032 allele and one B*4407. In addition, 19 B*44neg/ Cw*07pos Bubi individuals were typed for HLA-C and none of them proved Cw*0706 positive. To determine whether the association between Cw*0706 and B*44032 was limited to the Bubi, 19 individuals from Dutch Caucasian families were typed in which B44 and Cw7 segregated on one haplotype. None of these individuals showed the presence of B*44032 or Cw*0706. The haplotypes found in the Dutch Caucasians were B*4402 Cw*0704, B*44031-Cw*07011 and B*44031-Cw*0702. The present observation indicates a strong association between B*44032 and Cw*0706 in the Bubi population. PMID- 10703611 TI - An HLB-B null allele (B*0808N) caused by a nucleotide deletion in exon 3, found in the family of a bone marrow transplant recipient. AB - We have identified a variant HLA-B allele, B*0808N, segregating through two generations of healthy individuals, whilst HLA typing the family of a bone marrow patient. Serological typing identified a disparity between the father (A1, A3 B7 DR7) and the brother (A1, A2 B56 DR1, DR7) of the patient. Low/medium resolution polymerase chain reaction using sequence-specific primers (PCR-SSP) revealed a B*08 allele undetectable by serological methods. High resolution DNA typing by polymerase chain reaction-sequencing based typing (PCR-SBT), revealed a nucleotide deletion at position 131 (C) in exon 3, the only difference between the new allele and B*0801. The deletion results in a frame shift in the protein coding sequence, introducing a premature termination codon (TGA) in exon 4. Although a B*08 allele is present in these individuals, the deletion prevents correct expression of the antigen on the cell surface. PMID- 10703612 TI - A novel HLA-A24 (A*2420) allele identified in the Atayal tribe of Taiwan. AB - The Taiwan indigenous population groups are classified into different tribes according their linguistic classification and cultural anthropology. One of the tribes, the Atayal, showed a high frequency of A24 alleles by SSOP analysis. High resolution sequencing based typing identified a A*2402 variant "A*2420" which was found in 6 unrelated individuals. High-resolution typing is required to identify HLA polymorphism in the Taiwanese minority groups. PMID- 10703614 TI - Identification of an HLA-B7 serological variant and its characterization by sequencing based typing. AB - We have identified an HLA-B*07 variant allele, B*0716, in a Caucasoid cadaver kidney donor. The HLA class I type by polymerase chain reaction using sequence specific primers (PCR-SSP) was A*01, 32; B*07, 08; Cw*07. Serological typing, using monoclonal and polyclonal anti-HLA antisera, gave disparate results for the B antigens. Monoclonal antibodies identified B7 and B8 antigens but polyclonal antisera recognised only the B8 antigen. PCR using sequencing based typing (PCR SBT) confirmed the presence of both B*0703 and B*0801 alleles but with a mutation in one of the alleles. The HLA-B*07 allele was isolated by allele-specific PCR and was shown to have a mutation, G-->T, at 292 in exon 2. This mutation changes codon 74, which encodes aspartic acid (GAC) present in all previously identified B*07 alleles, to tyrosine (TAC) in the variant. The serological results suggest that codon 74 is a crucial part of a B7 antigen-specific epitope recognised by tissue typing polyclonal antisera. PMID- 10703613 TI - Complete sequence analysis of the A*1103 allele. AB - Here we report the full-length sequence of a novel A*11 variant. The variant was identified by ARMS-PCR and serology, the sequence was confirmed by cloning and subsequent sequencing. This variant, A*1103, found in a family of oriental origin, resembles the A*1101 sequence in exon 2 but differs in exon 3 with regard to codons 151 and 152. The polymorphism's result in two amino acid substitutions (one conserved (His->Arg), one introducing a negative charge (Ala->Glu)) located in the alpha2 helical region. The arginine at amino acid position 151 is rare amongst Alocus alleles and is besides A*1103 only observed in A*29 variants, the glutamine at amino acid position 152 is shared with A*0301, A*25, *26, *34 variants and the A*02 subtypes A*0203, *0213 and *0226. In fact, the amino acid motif comprising codons 151 and 152 is unique to A*1103 among Alocus alleles, but is common to C-locus alleles. PMID- 10703615 TI - Identification of the novel allele HLA-B*0809 in a Caucasian individual: estimation of allogeneic potential between B*08 variants. AB - The identification of the new allele HLA-B*0809, which was found in a Caucasian individual, is described. In the sequence analysis the new allele differs from B*0801 by six nucleotides located in exon 3. As the structure is identical to a variety of other HLA-B alleles, it is likely that the new allele originated by gene conversion. At the protein level, the new allele has two amino acid differences compared to B*0801. Comparing the residues of the alpha1 and alpha2 domains of the B*08 variants to each other, a high degree of polymorphism was found. Three alleles differ from B*0801 by only one amino acid residue whereas the other comparisons revealed at least two disparities. B*0809 differs from the other B*08 variants by at least two amino acid residues, suggesting that mismatching may provoke alloreactivity and thus impair clinical outcome of bone marrow transplantation. Among the B*08 alleles with a single amino acid difference, the mismatch combinations B*0801 vs. B*0805 and B*0801 vs. B*0807 appear to be the most compatible based on structural data. PMID- 10703616 TI - Identification of a novel allele, B*15012, by polymerase chain reaction using sequence-specific primers (PCR-SSP) and sequence-based typing. AB - We report here a new allele, B*15012, identified by polymerase chain reaction using sequence-specific primers (PCR-SSP) and sequence-based typing (SBT). B*15012 differs to B*15011 (previously named B*1501) by a single nucleotide, at position 435 of codon 120. B*15011 encodes AAG and B*15012 encodes AAA at codon 120, this difference does not result in an amino acid change. PMID- 10703617 TI - Identification of the novel allele HLA-B*1545: assessment of alloreactivity in case of mismatch with other B*15 alleles. AB - The novel allele HLA-B*1545, which has been serologically typed as B62, was identified in a male Caucasian. In the sequence analysis the new allele differs from B*1507 by nucleotide 419 which is located in exon 3. Its structure suggests that it may have originated by a point mutation or by gene conversion with a variety of HLA-B alleles. At the protein level, the nucleotide substitution results in an amino add exchange compared to B*1507 (Tyr116Ser). Due to probably substantial differences to B*1507 and the other B*15 variants with regard to peptide binding, a mismatch is likely to impair clinical outcome of bone marrow transplantation. PMID- 10703618 TI - Identification of the novel allele HLA-B*1546 which belongs to the serological B72 type: implications for bone marrow transplantation. AB - The identification of the novel allele HLA-B*1546, which has serological B50 and B72 reactivity, was found in two members of a family of Turkish origin. In the sequence analysis the new allele differs from B*1501 by four nucleotides in exon 2. Its structure suggests that it may have originated by gene conversion with B*40, B*41, B*44, B*4501, B*47, B*4901 or B*50. At the protein level, the new allele has three amino acid differences compared to B*1501. Sequence alignment demonstrates that amino acid residue 46 is crucial for serological B72 reactivity. Due to substantial differences with other B*15 variants a possible mismatch may impair clinical outcome of bone marrow transplantation. PMID- 10703619 TI - Identification of a novel HLA-Cw*07 variant (Cw*0714) in a German Caucasian family. AB - We report herein the identification of a new HLA-Cw*07 allele in two members of a German Caucasian family. This novel allele, designated as Cw*0714, differs from Cw*07011 and Cw*0706 by two nucleotide changes: one at codon 66 (AAC-->AAG) in the exon 2, leading to an amino acid change from Asn to Lys; and another silent substitution at codon 99 (TAT-->TAC) in the exon 3. The latest substitution (T- >C at the third position of codon 99) was not seen in any of the HLA-Cw*07 alleles reported so far, thus being characteristic to the new HLA-Cw*0714 allele. PMID- 10703620 TI - New DRB1* alleles (HLA-DRB1*1135, DRB1*1430 and DRB1*1433) and a confirmatory sequence (DRB1*1133). AB - Three new DRB1 alleles (DRB1*1135, DRB1*1430 and DRB1*1433) and a confirmatory sequence (DRB1*1133) have been identified after following up unusual or novel polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) patterns during routine typing of the DRB1*03,*08,*11,*12,*13 and *14 allele groups. Of the new alleles found and described in this paper, two alleles were initially detected by the PCR-RFLP method which produced unexpected restriction polymorphism (DRB1*1133 and DRB1*1135) while the remaining two were found after following up rare allele typings from this technique (DRB1*1430 and DRB1*1433). PMID- 10703621 TI - Detection of four novel alleles: DRB1*1130, DRB1*13072, DRB1*1315 and DRB1*1331. AB - Four new DR52-associated DRB1 alleles are described. One allele, DRB1*1130, is a hybrid between a DRB3*02 allele and a DRB1*11011 allele. The other alleles, DRB1*13072, DRB1*1315, and DRB1*1331, are simple reshufflings of known polymorphic motifs. PMID- 10703622 TI - Tumor necrosis factor receptor II (TNFRII) exon 6 polymorphism in systemic lupus erythematosus. AB - Systemic lupus erythematosus (SLE) is a complex autoimmune disease that exhibits extensive clinical heterogeneity. Several studies have suggested a role for tumor necrosis factor alpha (TNFalpha) in SLE and recently, the locus encompassing the TNF receptor II (TNFRII), which is a mediator of TNF effect, was amongst the candidate loci suggested by genetic linkage studies of multi-case SLE families. Komata et al. reported an association between a polymorphism at position 196 (R allele) of TNFR II and SLE in Japanese patients. We have typed SLE patients from two different ethnic populations, Spanish and UK Caucasoids, for this polymorphism using a polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP)-based technique. No significant differences in allele or genotype frequencies were found between cases and matched controls in either population. The TNFRII 196R allele does not appear to be associated with SLE susceptibility in either Spanish or UK populations. PMID- 10703623 TI - Nomenclature for factors of the HLA system, update September 1999. Marsh for the WHO Nomenclature Committee for Factors of the HLA System. PMID- 10703624 TI - Corticosteroid cross-reactions: an alternative view. AB - An alternative view of corticosteroid cross-reactivity is proposed, based on 2 immune recognition sites on the corticosteroid molecule, 1 influenced by C 6/9 substitution and 1 by C 16/17 substitution. A case report is adduced in support of such a hypothesis. PMID- 10703625 TI - Provocative use testing of methyldibromo glutaronitrile in a cosmetic shampoo. AB - Methyldibromo glutaronitrile (1,2-dibromo-2,4-dicyanobutane, MDGN) is a component of Euxyl K400, a broad-spectrum preservative increasingly used in cosmetics over the last 10 years. Contact allergy to MDGN is not uncommon, with leave-on cosmetics and moistened toilet tissue being the most important sources. Rinse-off products have rarely been reported to elicit allergic reactions in patients sensitized to MDGN and shampoos have only been implicated in hairdressers. To determine whether subjects sensitized to MDGN would react in a provocative use test with a shampoo containing the preservative, we asked 12 pre-sensitized subjects to use a shampoo containing 0.02% MDGN for a period of 9-13 weeks. During this time, none of the subjects showed any skin reactions indicative of contact allergy. The results of this study provide evidence to support that a shampoo product containing 0.02% MDGN may safely be used by most individuals who are presensitized to MDGN. PMID- 10703626 TI - Increase in sensitization to oil of turpentine: recent data from a multicenter study on 45,005 patients from the German-Austrian Information Network of Departments of Dermatology (IVDK). AB - Contact allergy to oil of turpentine was reported to have become rare. However, the evaluation of standardized data of 45,005 patients tested 1992-1997 in 30 Dermatological Centers associated with the German-Austrian Information Network of Departments of Dermatology (IVDK) showed an increase in positive patch test reactions to turpentine from 0.5% during the years 1992-1995, up to 1.7% in 1996 and 3.1% in 1997. In particular, 17,347 patients tested in 1996-1997 were evaluated in detail by comparing 431 individuals with positive patch test reactions with the rest of the group found negative to turpentine. Using the so called MOAHLFA index, the following characteristics were shown. Turpentine allergy (a) was found to be significantly less frequent in men and in patients with occupational dermatitis, (b) showed no difference in its association with atopic dermatitis, (c) patients with turpentine allergy had significantly less symptoms of the hands, more symptoms of the legs or in the face and (d) were significantly more often aged over 60 years. Also, patients sensitized to turpentine had increased rates of additional sensitizations. The definite reason for the increase in turpentine sensitization in the population tested here is not clear. Therefore, a detailed exposure analysis is necessary; the new increase in turpentine allergies may be due to popular topical remedies or household chemicals. PMID- 10703627 TI - Type III and type IV hypersensitivity reactions due to mitomycin C. AB - A 71-year-old man developed an exfoliative dermatitis of the palms of the hands and soles of the feet, and a generalized itch, during treatment with intravesical instillations of mitomycin C for an undifferentiated carcinoma of the bladder. Patch tests with mitomycin C 0.03%, 0.1% and 0.3% aq. were positive. Because of the serious consequences of this finding, the patient was retested with mitomycin C in pet. (same concentrations), a more stable preparation. This showed clear positive reactions. During this last series of patch tests, he developed palpable purpura on the legs. We postulated that this reaction was an immune-complex mediated reaction, caused by the 2nd series of patch tests with mitomycin C. To prove this, we performed histopathological and immunofluorescence investigations, and these showed the reaction to be consistent with Henoch-Schonlein-type purpura. We therefore conclude that this patient developed systemic reactions to mitomycin C, characterized by an eczematous dermatitis as well as purpuric reactions. The intravesical installations with mitomycin C have been stopped. The patient's skin problems (the purpura as well as the eczema) have completely resolved and have not recurred. PMID- 10703628 TI - Efficacy of a barrier cream and its vehicle as protective measures against occupational irritant contact dermatitis. AB - The actual advantage of barrier creams over bland emollients for skin protection is still hotly debated. In a randomized, double-blinded study, a newly-introduced barrier cream and its moisturizing vehicle were compared regarding their skin compatibility, efficacy and resulting acceptance. Thus, 2 panels of 25 hospital nurses with mild signs of skin irritation were asked to use 1 of the test products provided (verum or vehicle) over a period of 4 weeks. Effects of both types of preparations were studied weekly by clinical examination and the instrumental assessment of bioengineering parameters. Results showed no significant differences between barrier cream and vehicle. In both groups, clinical skin status improved and stratum corneum hydration increased significantly during the study period. Both preparations were tolerated and accepted well, thus showing both skin protection and skin care. These results contribute to the debate as to whether a strict distinction between "skin care" and "skin protection" products is justified. The vehicle alone is capable of positively influencing skin status. Emphasis must be laid on regular, frequent, and correct application of a product for it to be effective. PMID- 10703629 TI - Regulation of the cutaneous allergic reaction by humidity. AB - Humidity is 1 of the environmental factors which regulate skin conditions. Effects of humidity on the cutaneous immune reaction were examined. Contact hypersensitivity to 2,4,6-trinitrochlorobenzene was elicited in C57BL/6 mice. The reaction was greater in mice housed under low humidity conditions (about 10%) for 2 days, at either the induction or elicitation phase, than in mice housed under rather high humidity conditions (80%). After housing under controlled humidity for 2 days, the number of I-A positive cells was 16% higher in the epidermis exposed to the dry condition. The increased population of FITC-positive cells were in regional lymph nodes after painting of FITC during housing under lower humidity. Our study demonstrated that the cutaneous immune reaction is regulated by environmental humidity and suggested 2 possible mechanisms, i.e., increase in Langerhans cells and increased penetration of allergen with low humidity. PMID- 10703630 TI - Effect of an antioxidant (quercetin) on sodium-lauryl-sulfate-induced skin irritation. AB - Quercetin is a bioflavonoid with antioxidant and anti-inflammatory activity. The purpose of this study was to examine the effect of quercetin on acute skin irritation, with special interest in the skin barrier function recovery. Acute irritant contact dermatitis was induced in 15 patients by 24-h occlusion of 2% sodium lauryl sulfate (SLS) (day (D) 1). The influence of application on SLS irritated skin of topical quercetin for 5 consecutive Ds, compared to vehicle and controls, was studied. Parameters measured were transepidermal water loss (TEWL) and erythema index. Final measurements were taken on D 7 after a 1-D rest period. TEWL and the erythema index continued to rise 2 D after application of SLS and 1 D after treatment with quercetin, vehicle or controls. Both TEWL and erythema values at D 7 did not return to values before the SLS barrier disruption at all the test sites. Therefore, quercetin topically applied after induction of irritant contact dermatitis does not appear to increase the recovery of barrier function and erythema caused by SLS. PMID- 10703632 TI - Irritant thresholds in subjects with colophony allergy. AB - The factors which determine individual susceptibility to the development of allergic contact dermatitis (ACD) are not well defined. Since it is known that the presence of skin irritation is well-documented as a promoter of sensitization, the sensitivity to irritation of subjects with ACD has been compared with a normal control group. Whilst 78% of those with a positive patch test reaction to the contact allergen colophony (colophonium) responded to a concentration of no higher than 2.5% sodium lauryl sulfate (SLS), only 44% of the matched control group reacted at this level of SLS. Thus, the results are consistent with the hypothesis that individuals with enhanced susceptibility to skin irritation are also more liable to develop allergic skin reactivity. PMID- 10703631 TI - The results of ingredient patch testing in contact dermatitis elicited by povidone-iodine preparations. AB - 10 cases of contact dermatitis which began during the application of povidone iodine preparations were examined with patch tests using 2 kinds of povidone iodine preparations and their ingredients, i.e., povidone-iodine, polyoxyethylene nonylphenyl ether and glycerin, and also the components of povidone-iodine, i.e., iodine and polyvinylpyrrolidone. All 10 cases reacted positively to the povidone iodine preparations and povidone-iodine, 3 out of the 10 to polyoxyethylene nonylphenyl ether, 1 out of the 9 tested to iodine, while no positive response was found to glycerin or polyvinyl-pyrrolidone. It was difficult to distinguish between allergic responses from irritation, as responses to patches of povidone iodine and its preparations usually include irritation at high frequencies. Based on comparison of results with a control group, however, those showing + or stronger reactions to 2% povidone-iodine at days 3 to 5 were considered to be allergic. Thus, 4 out of the 10 cases were considered as sensitization to povidone-iodine. Another 3 cases were found to be polyoxyethylene nonylphenyl ether sensitized, and another 1 iodine sensitized, while the patch test reactions of the other 2 were considered to have been elicited by irritation. PMID- 10703634 TI - Semicircular lipoatrophy: 18 cases in the same company. PMID- 10703633 TI - Strontium nitrate suppresses chemically-induced sensory irritation in humans. AB - Skin care products are complex formulations that may cause sensory irritation symptoms, characterized by stinging, burning, and itching. Substances capable of counteracting sensory irritation are of great practical interest. Strontium salts have been demonstrated to inhibit sensory irritation and inflammation when applied topically. In this double-blind study, we evaluated the efficacy of strontium nitrate in reducing chemically-induced skin sensory irritation in 8 subjects. In a random order, 20% strontium nitrate in 70% glycolic acid (pH=0.6) (mixture) was applied to the volar aspect of the forearm and a positive control (70% glycolic acid, pH=0.6) to the contralateral forearm. The irritation sensation was evaluated each min for the first 20 min after topical application using a scale from 0-4. The duration of the irritation sensation in min was also recorded. Strontium nitrate mixed with glycolic acid, in comparison with glycolic acid alone, markedly (p<0.01) shortened the duration of the irritation sensation from 24.4+/-4.1 (mean+/-SEM) min to 8.9+/-3.7 (mean+/-SEM) min, and significantly (p<0.05) reduced the mean magnitude of the irritation sensation at all time points (overall). The study demonstrated that strontium nitrate potently suppresses the sensation of chemically-induced irritation. PMID- 10703635 TI - Nickel allergy from orthodontic appliances. PMID- 10703636 TI - Allergic contact dermatitis due to monovalent sensitization to the oxidation hair dye intermediate oxamitol (2-aminomethyl-p-aminophenol-2HCl) without cross sensitivity to haptens of the para-group. PMID- 10703637 TI - Leukonychia from 2-ethyl-cyanoacrylate glue. PMID- 10703638 TI - Textile contact dermatitis presenting as lichen amyloidosus. PMID- 10703639 TI - Allergic contact dermatitis from diallyl disulfide. PMID- 10703640 TI - Allergic contact dermatitis from imidazolidinyl urea in an ultrasonic gel. PMID- 10703641 TI - Bullous contact dermatitis from nasturtium. PMID- 10703642 TI - 2 cases of allergic contact cheilitis from sodium lauryl sulfate in toothpaste. PMID- 10703643 TI - Allergic contact dermatitis from topical carmustine. PMID- 10703644 TI - Occupational airborne allergic contact dermatitis from sawdust in livestock sheds. PMID- 10703645 TI - Contact urticaria syndrome from mustard in anchovy fillet sauce. PMID- 10703646 TI - Recurrent allergic contact dermatitis and cheilitis due to castor oil. PMID- 10703647 TI - Occupational allergic contact dermatitis from glyoxal, glutaraldehyde and neomycin sulfate in a dental nurse. PMID- 10703648 TI - Lichenoid reaction to temporary tattoo. PMID- 10703649 TI - Airborne nickel dermatitis. PMID- 10703650 TI - Irritant contact dermatitis of the hands following thoracic sympathectomy. PMID- 10703651 TI - Antimicrobial resistance in the subgingival microflora in patients with adult periodontitis. A comparison between The Netherlands and Spain. AB - BACKGROUND: The widespread use of antibiotics for prophylaxis and treatment of bacterial infections has lead to the emergence of resistant human pathogens. Great differences have been documented between European countries in the use of systemic antibiotics. In parallel, significant differences in levels of resistant pathogens have been documented. AIM: To investigate whether differences in antibiotic use influence the level of antimicrobial resistance of the subgingival microflora of untreated patients with adult periodontitis in The Netherlands and Spain. METHOD: Blood agar plates containing breakpoint concentrations of penicillin, amoxicillin, amoxicillin and clavunalate, metronidazole, erythromycin, azithromycin, clindamycin and tetracycline were used to determine the proportion of bacteria from the subgingival plaque that was resistant to these antibiotics. In the Spanish patients, statistically significant higher mean levels of resistance were found for penicillin, amoxicillin, metronidazole, clindamycin and tetracycline. The mean number of different bacterial species growing on the selective plates was higher in the Spanish patients, as was the % of resistant strains of most periodontal pathogens. A striking difference was observed in the frequency of occurrence of tetracycline-resistant periodontal pathogens. In Spain, 5 patients had > or =3 tetracycline resistant periodontal pathogens, whereas this was not observed in any of the Dutch patients. CONCLUSIONS: The widespread use of antibiotics in Spain is reflected in the level of resistance of the subgingival microflora of adult patients with periodontitis. PMID- 10703652 TI - Radiographic periodontal attachment loss as an indicator of death risk in the elderly. AB - OBJECTIVES: Oral infections have been associated with serious systemic diseases and an increased risk of death. Our aims were to investigate whether radiographically-observed apical periodontitis lesions, carious teeth, periodontal attachment loss (horizontal bone loss, furcation lesions, number of teeth with infrabony periodontal pockets, the extent of infrabony periodontal pockets) and the sum of all these findings have any relationships with all-cause mortality within 4-year follow-up. MATERIAL AND METHODS: 292 community-dwelling elderly persons aged 76, 81 and 86 years. The number of deaths within 4 years was 54 (18.5%). In the dentate 169 subjects, of whom 32 (18.9%) deceased within 4 years, the mean number of teeth was 15.5 in men and 13.2 in women. The imaging method used was panoramic radiography supplemented by intraoral radiographs. RESULTS: 51% of the dentate subjects had infrabony pockets (mean 1.5, s.d. 2.2), and 40% had periapical periodontitis lesions (mean 1.0, s.d. 1.6). After controlling for age and gender, vertical bone loss judged as advanced infrabony pockets was associated with 4-year all-cause mortality (Odds ratio 2.2,1.0-4.7). Other associations were statistically insignificant. CONCLUSION: Periodontal attachment loss may indicate an increased risk of death in the elderly. PMID- 10703653 TI - The accuracy of the Vivacare true pressure-sensitive periodontal probe system in terms of probing force. AB - BACKGROUND: The Hunter TPS Vivacare periodontal probe was invented to perform consistent, accurate and reliable periodontal examinations "with controlled pressures". AIMS: The aims of the present investigation are 3: (1) what is the accuracy of the probing force when various probe heads are used in a correct operation position; (2) what is the effect of over- and under-reading of the operation position on the probing force; (3) what is the accuracy of the probing force when different probe handles are used. The Hunter TPS probe consists of a tip connected to a special spring mechanism, which controls the pressure extended to the probe tip. According to the manufacturer, the force indicator lines coincide at approximately 20 g force. METHOD: The test apparatus consisted of an electronic balance, and an electronic caliper. 12 TPS probes tips and 3 handles were selected to test whether there were differences in force between probes. Each probe tip was adapted to the same handle and tested 10 times. In a 2nd test, the TPS handles and over-, accurate-, and under-readings were analyzed as to how they affect the probing force. The 12 TPS probe tips were connected to each of the 3 handles and tested 10x for each of 3 handles and levels. RESULTS: The range in force between TPS probe tips was 8.4 g (p<0.001). For the handles as well as for each of the readings of the operating positions, the differences were small and non-significant. However, between over- and under-readings, there were statistical significant differences. CONCLUSION: The conclusion is that the variation in force between probe tips is high and always above the manufacturer's "approximately" 20 g. As long as the same handle and the same probe tip are used together, and the examiner is reading the markings correct, the TPS probe is adequate. PMID- 10703654 TI - Studies in vitro of abrasion by different manual toothbrush heads and a standard toothpaste. AB - BACKGROUND: Loss of dentine at the buccal cervical region of teeth has a multifactorial aetiology. However, a considerable amount of circumstantial evidence, supported by laboratory experiments, implicates toothbrushing with toothpaste as a consistent factor. Most interest has centred around the abrasivity of toothpastes, particularly since a toothbrush alone has negligible effects on dentine. The influence of filament stiffness on toothpaste abrasion was the subject of some studies, mostly at least 2 decades ago, and produced conflicting conclusions. Numerous changes to toothbrush design and construction have taken place in recent years. AIMS: The aim of this study was to measure the abrasion of a standard substrate, acrylic, by a standard toothpaste carried on modern brands of toothbrushes classified by manufacturers as hard (3 brands), medium (3 brands) and soft (6 brands). METHOD: The substrate was brushed for 20,000 strokes with at least 6 heads from each brand. Measurements of substrate loss were made at 5000 stroke increments by profilometry. RESULTS: Substrate loss for all brushes showed a pattern of abrasion which was to a first approximation linear. Overall, hard brushes caused least abrasion and soft brushes the most, with differences between groups being significant. Within-group differences between brands reached significance for soft and medium brushes but not hard brushes. CONCLUSIONS: The results could be explained by increased retention of toothpaste by smaller diameter filaments and denser tufts on soft brushes and the greater flexion of filaments increasing the area of contact with the surface. Calculations on the clinical outcome of these data in vitro indicate that toothbrushing with toothpaste alone would produce minimal damage to dentine even over many years. Differences between brushes therefore are probably of little clinical significance. Certainly, the data do not support the use of hard brushes, particularly in view of the potential detrimental effects to gingival tissues. PMID- 10703655 TI - A retrospective radiographic outcome assessment study of intra-bony defects treated by osseous surgery or by bone graft procedures. AB - BACKGROUND: Intra-bony defects remain a significant therapeutic problem in periodontal therapy. Various non-surgical and surgical treatment modalities are being used. The long-term stability following treatment of intra-bony defects is poorly documented. OBJECTIVES: To assess changes in intra-bony defects after either osseous surgery or open flap debridement in combination with grafting procedures with demineralized freeze-dried bone allografts (DFDBA). METHOD: Pre- and post-surgical computer digitized images of intra-oral radiographs from 60 patients who had received periodontal surgery to manage intrabony defects were analyzed by linear measurements. RESULTS: 36 patients were treated with osseous surgery and 24 had received flap procedures and grafting with DFDBA. Post surgical radiographs were obtained on average after 4.8 years (SD+/-2.8) and after 9.6 years (SD+/-3.6). A minor mean bone fill of 0.0 mm (SD+/-0.8) for osseous surgery sites and 0.5 mm (SD+/-0.9) for DFDBA sites, was noticed, but this gain was within the margin of measurement errors. Osseous surgery and modified Widman flap procedures with DFDBA resulted in crestal resorption, on average 1.7 mm (SD+/-1.5) and 1.5 mm (SD+/-1.5) and remaining mean defect depth of 2.0 mm (SD+/-1.4) and 2.5 mm (SD+/-1.6), respectively. CONCLUSIONS: Bone changes following bone graft procedures with DFDBA did not differ from those following osseous surgery, and neither procedure resulted in defect resolution with bone fill. It was also concluded that over the study period, stable treatment results were obtained as a result of both osseous surgery and modified Widman flap procedures with adjunct DFDBA. PMID- 10703656 TI - The calcium channel blocker used with cyclosporin has an effect on gingival overgrowth. AB - BACKGROUND/AIMS: To investigate whether the choice of calcium channel blocker, used in conjunction with cyclosporin A, affected the prevalence of gingival overgrowth. METHOD: A cohort of 135 renal transplant recipients who had been medicated with cyclosporin A in combination with either nifedipine (89) or amlodipine (46) since transplant, took part in the study. The inclusion criteria were that eligible subjects had been in receipt of a kidney transplant for at least 12 months, had at least 10 teeth and had not received specialist periodontal treatment. The age, gender, current drug regimen and dosage were recorded for each participant and alginate impressions taken of both arches. The presence and severity of gingival overgrowth were scored from plaster models. RESULTS: A higher proportion (72%) of the amlodipine group were categorised as having gingival overgrowth compared with only 53% of the nifedipine group, chi square=4.5, p<0.05. Logistic regression analysis was used to explore the relationship between the presence or absence of gingival overgrowth (dependent variable) and age, gender, time since transplant, dose of cyclosporin A, centre in which the patient was treated, and the calcium channel blocker used (independent variables). Independent predictors of gingival overgrowth in this multivariate analysis were whether the individual was treated with amlodipine or nifedipine (p=0.01) and whether the individual was young or old (p=0.01). Within the multivariate analysis, the odds ratio for amlodipine to be associated with gingival overgrowth compared with nifedipine was 3.0 (confidence interval 1.3 6.9). CONCLUSIONS: The prevalence of gingival overgrowth in renal transplant recipients maintained on cyclosporin A and nifedipine is lower than those treated with cyclosporin A and amlodipine. PMID- 10703657 TI - Lack of antimicrobial effect on periodontopathic bacteria by ultrasonic and sonic scalers in vitro. AB - BACKGROUND: The purpose of this study was to assess the antimicrobial effects of a sonic and ultrasonic scaler generally used for subgingival scaling on gram negative and gram-positive periodontopathic bacteria. METHOD: Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Campylobacter rectus, or Peptostreptococcus micros were suspended in Schaedler's broth medium and treated by a sonic or a magnetostrictive ultrasonic scaler for 30 s and 150 s in vitro. Bacterial suspensions treated by an ultrasonic cell disruptor served as a positive control and untreated bacterial suspensions served as a negative control. Following sonication, samples were serially diluted, streaked on blood agar plates and incubated for 2-5 days at 37 degrees C. RESULTS: Treatment by the sonic or ultrasonic scaler for up to 150 s did not reduce the viability of any of the tested periodontal pathogens. Compared to untreated controls, the viability of A. actinomycetemcomitans and P. gingivalis was significantly (p<0.05) reduced only following ultrasonication with the cell disruptor after 30 s (0.72 and 0.54 log CFU/ml, respectively) and of A. actinomycetemcomitans, P. gingivalis, C. rectus, and P. micros after 150 s (1.98, 1.34, 1.95 and 1.98 log CFU/ml, respectively). CONCLUSION: The data of the study may indicate that the assessed sonic and ultrasonic scaler used for subgingival debridement do not result in killing of the tested periodontal pathogens. PMID- 10703658 TI - Guided tissue regeneration procedure with bioresorbable membranes versus conventional flap surgery in the treatment of infrabony periodontal defects. AB - AIM: Claims are being made that clinical results of periodontal flap surgery are enhanced when membranes are employed to aid GTR in intrabony pockets. It was the aim of our study to determine whether this assumption was true for a certain bioresorbable membrane (Guidor Matrix Barrier). METHOD: 44 intrabony defects were treated in 16 patients. In 21 lesions, conventional flap surgery only was performed, while in 23, similar lesion membranes were placed as an additional treatment task. Results were evaluated over a time span of 12 months. RESULTS: In all surgical areas, treatment resulted in significant improvement of parameters such as attachment levels and probing depths, as well as index values for plaque and bleeding on probing. This occurred whether membranes had been used or not, without any significant differences when comparing the collective results of both treatment groups. CONCLUSIONS: Placement of membranes during periodontal surgery for the enhancement of tissue regeneration in intrabony pockets is often both difficult and time consuming. In the light of our clinical results with resorbable membranes, such extra effort seems hardly warranted. PMID- 10703659 TI - Long-term evaluation of osseointegrated dental implants in the treatment of partly edentulous patients. AB - AIM: The aim of this study was to evaluate the clinical, radiographic and microbiological status of implants after 10 years of functional load in patients treated for partial edentulism. METHOD: 15 patients, each successfully treated with 2-6 implants ad modum Branemark placed in free-standing fixed prostheses, were included in the study. RESULTS: Clinical evaluation revealed similar degrees of inflammation around teeth and implants. The probing pocket depth (PPD) was significantly greater around implants than around teeth. The mean marginal bone loss during 10 years of functional load was comparable to that found at the time of the 5-year follow-up. 74% of the implants remained free of marginal bone loss exceeding 1 mm. Marginal bone loss exceeding 2 mm, was found at only 5 sites. No marked differences in bacteria were present between teeth and implants. T. denticola, S. intermedia and P. micros were the commonest organisms detected around teeth and implants. The periodontal pathogens A. actinomycetemcomitans, P. gingivalis, P. intermedia, B. forsythus, and T. denticola, were found at implants with a marginal bone loss of more than 2 mm. CONCLUSION: Our study shows that the long-term results with implants in partially dentate patients are similar to those seen in edentulous patients and that no significant change occurred after 5 year follow-up over an additional period of 5 years. PMID- 10703660 TI - Clinical benefits of oral irrigation for periodontitis are related to reduction of pro-inflammatory cytokine levels and plaque. AB - BACKGROUND: Although a growing body of evidence indicates that oral irrigation with water has therapeutic benefits in periodontitis, the mechanisms of action have not been elucidated. The aims of this study were: (1) to analyze the effects of oral irrigation (Water Pik Oral Irrigator) on the clinical signs of adult periodontitis (AP) and on the levels of interleukin-1 beta (IL-beta), prostaglandin-E2 (PGE2), interleukin-10 (IL-10) and interferon-gamma (IFN-gamma) in GCF, and (2) to analyze the influence of the periodontitis-related IL-1 genotype (IL-1GT) on these variables. METHOD: A single-center, blinded study in otherwise healthy humans (n= 52) with localized mild to moderate AP was carried out, using the following groups: group A (n= 12), no oral hygiene for 14 days; group B (n=20), routine oral hygiene (ROH) for 14 days; group C (n=20), supra gingival oral irrigation plus ROH for 14 days. Group A patients were crossed-over to group C for 14 days (=day 28) after a professional prophylaxis. Group assignment was randomized by a coin toss, with the exception of group A subjects, who were self-selected as per recommendations of the internal review board for human subjects. GCF was sampled from 3 study teeth per patient and analyzed for IL-1 beta, PGE2, IL-10 and IFN gamma by ELISA on days 0, 7, 14 and 28. Probing pocket depths (PPD), clinical attachment levels (CAL), bleeding on probing (BOP), gingival index (GI) and plaque index (PI) were measured by a calibrated examiner (TWS) on days 0, 14 and 28. Analysis of covariance was performed using SAS 6.12 and Proc Mixed with group and IL-1GT as the factors and the baseline levels as the covariate, with output being least squares means and least significant difference (LSD). Significant differences were declared if the p-value for the F statistic was < or =0.05. RESULTS: Oral irrigation plus ROH resulted in a significant reduction in PPD, BOP, GI and PI, as well as IL-beta levels by 7 days and PGE2 levels by 14 days, relative to ROH or no oral hygiene. Interestingly, decreased IL-1 beta levels in patients using oral irrigation plus ROH was accompanied by a trend for increased levels of the "anti-inflammatory" cytokine IL-10. ROH reduced GI, BOP and PI, and PGE2 levels by 14 days, but had no effect on IL-1 beta or IL-10 levels relative to no oral hygiene. The effects of no oral hygiene were reversed by a prophy followed by oral irrigation plus ROH for 14 days. No clinical differences were evident between IL-1 GT (+) patients (n= 1) and GT (-) patients (n=40), but the former had significantly elevated levels of GCF IL-10 and borderline increases in IL-1 beta (p=0.07). CONCLUSIONS: Oral irrigation with water for 14 days had an improved therapeutic benefit for AP over that of routine oral hygiene alone and this improvement was accompanied by a down modulation of the pro-inflammatory cytokine profile in GCF. PMID- 10703661 TI - Reduction in gingival overgrowth associated with conversion from cyclosporin A to tacrolimus. AB - BACKGROUND: Unsightly gingival overgrowth affects many individuals immunosuppressed with cyclosporin A (CsA). Current management involves repeated periodontal surgery and intensive hygienist support. Tacrolimus is an effective alternative immunosuppressive agent for renal transplantation which does not appear to produce gingival enlargement. AIMS: The purpose of the present study was to monitor the gingival response of 4 renal transplant patients (RTPs), with clinically significant CsA-induced gingival overgrowth, after their immunosuppressive therapy was switched to tacrolimus. METHODS: Intra-oral photographs and alginate impressions were taken both prior to the drug conversion and again, 6 to 9 months later. Gingival overgrowth scores were determined, from plaster models on both these occasions. RESULTS: All of the RTPs experienced significant resolution of their gingival enlargement within the time period studied; however, only one had complete regression. CONCLUSION: It is concluded that conversion of RTPs with gingival overgrowth from CsA to tacrolimus may provide an effective management strategy for this clinical problem. PMID- 10703662 TI - Cytotoxicity of antiviral nucleotides adefovir and cidofovir is induced by the expression of human renal organic anion transporter 1. AB - The transport of organic anions in proximal convoluted tubules plays an essential role in the active secretion of a variety of small molecules by the kidney. In addition to other anionic substrates, the human renal organic anion transporter 1 (hOATI) is capable of transporting the nucleotide analogs adefovir and cidofovir. To investigate the involvement of hOATI in the mechanism of nephrotoxicity associated with these two clinically important antiviral agents, Chinese hamster ovary (CHO) cells were stably transfected with hOATI cDNA. The resulting CHOhOAT cells showed probenecid-sensitive and pH-dependent uptake of p-aminohippurate (Km = 15.4 FtM, V,,, ..ax = 20.6 pmol/106 cells min), a prototypical organic anion substrate. In addition, the stably expressed hOATI mediated efficient transport of adefovir (Km, = 23.8 tLM, V, a,, = 46.0 pmol/106 cells min) and cidofovir (K, = 58.0 /iM, Vt,ax = 103 pmol/106 cells * min) such that the levels of intracellular metabolites of both nucleotides were > 1 00-fold higher in CHOh OAT cells than in parental CHO. Consequently, adefovir and cidofovir were approximately 500-fold and 400-fold more cytotoxic, respectively, in CHOh OAT cells compared to CHO. The cytotoxicity of both drugs in CHOh OAT cells was markedly reduced in the presence of hOATI inhibitors. The cyclic prodrug of cidofovir, which exhibits reduced in vivo nephrotoxicity, was a poor substrate for hOATI and showed only marginally increased cytotoxicity in CHOh OAT cells. In conclusion, these studies demonstrate that hOATI plays a critical role in the organ-specific toxicity of adefovir and cidofovir, and indicates that CHOh OAT cells may represent a useful in vitro model to investigate the potential nephrotoxicity of clinically relevant organic anion agents. PMID- 10703663 TI - Molecular homology and the luminal transport of Hg2+ in the renal proximal tubule. AB - The aim of this study was to define mechanisms involved in the luminal uptake of inorganic mercury in the kidney using isolated perfused straight (S2) segments of the proximal tubule. When mercuric conjugates of glutathione (GSH), cysteinylglycine. or cysteine (containing 203Hg2+) were perfused through the lumen, the rates of luminal disappearance flux (JD) of inorganic mercury were approximately 39, 53, and 102 fmol/min per' min, respectively. Thus, the rates of luminal uptake of mercury are greater when the mercury is in the form of a mercuric conjugate of cysteine than in the form of a mercuric conjugate of cysteinylglycine or GSH. Addition of acivicin to the perfusate, to inhibit activity of the y-glutamyltransferase, caused significant reductions in the J,, for mercury in tubules perfused with mercuric conjugates of GSH. Addition of cilastatin, an inhibitor of dehydropeptidase- l (cysteinylglycinase) activity, caused significant reductions in the uptake of mercury in tubules perfused with mercuric conjugates of cysteinylglycine. These findings indicate that a significant amount of the luminal uptake of mercury, when mercuric conjugates of GSH are present in the lumen, is dependent on the activity of both y glutamyltransferase and cysteinylglycinase. Finally, the JD for mercury in tubules perfused with mercuric conjugates of cysteine was reduced by approximately 50% when 3.0 mM L-lysine or 5.0 mM cycloleucine was added to the perfusate. It is concluded that these findings indicate that at least some of the luminal uptake of mercuric conjugates of cysteine occurs at the site of one or more amino acid transporters via a mechanism involving molecular homology. PMID- 10703664 TI - Glomerular expression of dystroglycans is reduced in minimal change nephrosis but not in focal segmental glomerulosclerosis. AB - Extensive flattening of podocyte foot processes and increased permeability of the glomerular capillary filter are the major pathologic features of minimal change nephrosis (MCN) and focal segmental glomerulosclerosis (FSGS). Adhesion proteins anchor and stabilize podocytes on the glomerular basement membrane (GBM), and presumably are involved in the pathogenesis of foot process flattening. Thus far, ao3 P,-integrin was localized to basal cell membrane domains. In this report, ao- and 3-dystroglycan (DG) were detected at precisely the sa-ne location by immunoelectron microscopy. and the presence of ac- and /-DG chains was confirmed by immunoblotting on isolated human glomeruli. Because the major DG binding partners in the GBM (laminin, agrin, perlecan), and the intracellular dystrophin analogue utrophin are also present in glomeruli, it appears that podocytes adhere to the GBM via DG complexes, similar to muscle fibers in which actin is linked via dystrophin and DG to the extracellular matrix. As with muscle cells, it is therefore plausible that podocytes use precisely actin-guided DG complexes at their "soles" to actively govern the topography of GBM matrix proteins. Expression of the a//3-DG complex was reported to be reduced in muscular dystrophies. and therefore a search for similar pathologic alterations in archival kidney biopsies from patients with MCN (it = 16) and FSGS (ni = 8) was conducted by quantitative immunoelectron microscopy. The density of a-DG on the podocyte's soles was significantly reduced to 25% in MCN, whereas it was not different in normal kidneys and FSGS. The expression of 3-DG was reduced to >50% in MCN, and was slightly increased in FSGS. Levels of DG expression returned to normal in MCN after steroid treatment (7 = 4). Expression of /3-integrin remained at normal levels in all conditions. These findings point to different potentially pathogenic mechanisms of foot process flattening in MCN and FSGS. PMID- 10703665 TI - Interleukin-4 and interleukin-13 act on glomerular visceral epithelial cells. AB - In minimal change nephrosis (MCN), proteinuria is associated with structural changes of the glomerular visceral epithelial cells (GVEC). The occurrence of MCN has been associated with 2 lymphocyte-dependent conditions. To examine a direct role for type 2 cytokines in GVEC injury, the expression of interleukin (IL)-4/IL 13 receptors by GVEC and direct effects of IL-4 and IL-13 on GVEC were studied. Reverse transcription-PCR showed that isolated human and rat glomeruli and cultured human and rat GVEC expressed mRNA for IL-4Ralpha, IL-13Ralpha1, and IL 13Ralpha2. Protein expression of [L-4Ralpha and IL-13Ralpha2 by GVEC in human kidney biopsies and by cultured human GVEC was detected by immunohistochemistry. Western blotting demonstrated phosphorylation of STAT6 in cultured GVEC upon incubation with IL-4 or IL-13. This indicated signal transduction via the heterodimeric receptor complex IL-4R2, which is composed of the IL-4Ralpha and the IL-13Ralpha1. Direct effects on GVEC function were examined in monolayer experiments. IL-4 and IL-13 dose-dependently decreased transepithelial electrical resistance of monolayers of rat GVEC to approximately 30 and 40% of baseline values, respectively. The transepithelial electrical resistance decrease was associated with a significant increase in short-circuit current, whereas no changes were observed in the transmonolayer flux of the macromolecules horseradish peroxidase (molecular weight, 44 kD) and 14C-mannitol (molecular weight, 182 Da). No changes in cell structure were observed with electron microscopy. It is concluded that by binding to specific IL-4/ IL-13 receptors, IL 4 and IL-13 can exert specific effects on GVEC function, which could be of pathogenetic relevance for glomerular injury in MCN. PMID- 10703666 TI - Expression of profilin, an actin-binding protein, in rat experimental glomerulonephritis and its upregulation by basic fibroblast growth factor in cultured rat mesangial cells. AB - Profilin binds to actin monomer to regulate actin polymerization, and to phosphatidylinositol 4,5-bisphosphate to inhibit hydrolysis by phospholipase Cgamma1. This study investigated the expression of profilin in rat anti-Thy-1.1 mesangial proliferative glomerulonephritis (GN) and examined the effect of growth factors on its expression in cultured rat mesangial cells. Profilin mRNA was constitutively expressed in isolated glomeruli of untreated rats. However, in glomeruli of anti-Thy-1.1 GN rats, its expression was upregulated beginning on day 1, reaching a peak level on day 4 (3.9-fold versus control glomeruli), and decreased on day 14, as determined by competitive reverse transcription-PCR. Increased expression of profilin protein was confirmed using immunoblotting and immunohistochemistry. Immunoelectron microscopy revealed the presence of profilin in plasma membrane and the rough endoplasmic reticulum of mesangial cells, indicating that profilin was produced in mesangial cells. In cultured rat mesangial cells, expression of profilin mRNA and protein was upregulated by basic fibroblast growth factor but not by platelet-derived growth factor or transforming growth factor-beta. Suppression of profilin expression using an antisense oligonucleotide against profilin inhibited [3H]thymidine uptake. These findings indicated the involvement of profilin in anti-Thy-1.1 GN and suggest that the upregulation of profilin might be involved in the progression of anti Thy-1.1 GN possibly by affecting cell growth. PMID- 10703667 TI - Serum concentrations of laminin-P1 in thrombotic microangiopathy: usefulness as an index of activity and prognostic value. AB - Laminin is the main noncollagenous constituent of the basement membrane, and its serum levels could reflect the metabolic changes that occur in the basement membrane. Severe endothelial injury with thickening of basement membrane is a characteristic feature of thrombotic microangiopathy (TMA). With this background, the aim of the study was to investigate in a prospective way (1) the relationship among serum Lam-P1, the extent of renal histopathologic lesions, and the biochemical parameters commonly used as markers of TMA activity, and (2) the usefulness of serum Lam-P1 concentrations as a renal outcome prognostic index. To this end, 18 consecutive patients with active biopsy-proven TMA with renal involvement were studied. One hundred and twenty-one healthy control subjects, 20 patients with systemic scleroderma without renal involvement, and 35 patients with systemic lupus erythematosus (20 without nephropathy and 15 with diffuse proliferative type 4 lupus nephritis) were used as control groups. In addition, to analyze the influence of either renal failure or hemodialysis therapy on serum Lam-P1 levels, 91 patients on regular hemodialysis therapy and 81 patients with predialysis chronic renal failure of different etiologies were included in the study. Serum Lam-P1 was determined by RIA at admission, on days 10 and 30 of follow-up in all patients, and after 6 and 12 mo of follow-up in all surviving patients. Serum lactate dehydrogenase, haptoglobin, platelet count, hemoglobin, and serum creatinine were determined as markers of endothelial dysfunction and hemolysis. At admission, serum levels of Lam-P1 were significantly higher in patients with TMA than in healthy control subjects (3.39 +/- 0.56 U/ml versus 1.40 +/- 0.18 U/ml; P < 0.0001). In addition, patients with TMA had significantly higher serum Lam-P1 levels than the other groups included in the study. At the first control, Lam-P1 correlated with lactate dehydrogenase (P = 0.006) and hemoglobin (P = 0.002). During follow-up, platelet count and hemolysis indicators normalized in all patients, while serum Lam-P1 decreased only in patients with renal function recovery. In multivariate analysis, serum creatinine and Lam-P1 at day 10 were the only independent predictors of renal outcome (r2 = 0.94; P < 0.0001) and also correlated with indices of histopathologic damage (P < 0.001). Serum Lam-P1 normalized in all patients with chronic renal failure in the samples obtained at 6 and 12 mo of regular hemodialysis after solving active TMA, thus suggesting that histopathologic lesions, but not renal function itself, would be mainly responsible for the high Lam-P1 serum concentrations detected in TMA. In conclusion, serum Lam-P1 concentrations are increased in patients with active TMA. Furthermore, patients with poor renal outcome show a prolonged increase of serum Lam-P1 that is related to the extent of renal histologic lesions. Unlike the biochemical markers of hemolysis commonly used to assess TMA activity, the sequential determination of serum Lam-P1 provides valuable information about long term renal prognosis in patients with TMA. PMID- 10703668 TI - Telomere shortening in kidneys with age. AB - The histology and function of the kidney deteriorates with age and age-related diseases, but the mechanisms involved in renal aging are not known. In vitro studies suggest that telomere shortening is important in replicative senescence, and is accelerated by stresses that increase replication. This study explored the relationship between age and telomere length in surgical samples from 24 human kidneys, which were either histologically normal (17) or displayed histologic abnormalities (7). Telomere loss was assessed by two independent methods: Southern blotting of terminal restriction fragments (TRF) and slot blotting using telomere-specific probes. The results of these methods correlated with each other. The mean TRF length determined by Southern blotting in cortex was about 12 kb pairs (kbp) in infancy and was shorter in older kidneys. The slope of the regression line was about 0.029 kbp (0.24%, P = 0.023) per year. Telomere DNA loss in cortex by the slot blot method was 0.25% per year (P = 0.011). By both methods, the telomere loss in medulla was not significant and was less than in cortex. Comparisons of TRF length from 20 paired samples from cortex and medulla showed that TRF was greater in cortex than medulla, with the differences being greater in young kidneys and lessening with age due to telomere loss in cortex. These findings indicate that telomeres shorten in an age-dependent manner in the kidney, either due to developmental factors or aging, particularly in renal cortex. PMID- 10703669 TI - Relationship between expression of Bcl-2 genes and growth factors in ischemic acute renal failure in the rat. AB - The promotion of cell survival and regeneration in acute renal failure (ARF) is important for restitution of renal function. This study analyzes the temporal and spatial relationship between expression of pro- and anti-apoptotic members of the Bcl-2 gene family (Bcl-2, Bcl-X(L), Bax) and epidermal growth factor (EGF), insulin-like growth factor- (IGF-1), and transforming growth factor-beta (TGF beta), growth factors that are thought to be reparative in ARF. A rat model of ischemic ARF involving 30 min of bilateral renal artery occlusion followed by reperfusion for 0 to 14 d was used. Apoptosis and mitosis were quantified and qualitative assessment was made of other cellular damage including necrosis and loss of cellular adhesion. Locality and level of expression of the Bcl-2 and growth factor proteins were determined using immunohistochemistry. Apoptosis peaked between 4 and 14 d postischemia in both proximal and distal tubules. Mitosis peaked at 2 d in proximal tubules and 4 to 14 d in the distal tubules. A spatio-temporal relationship was observed between anti-apoptotic Bcl-2 gene family members and growth factors after ischemia-reperfusion. In control kidneys, expression of Bcl-2, Bcl-X(L) was low in epithelium of distal tubules, Bax had low-to-moderate expression in the proximal tubule and had low expression in the distal tubule, EGF and IGF-1 had low-to-moderate expression in the distal tubule, and TGF-beta had low expression in the proximal tubule. In contrast, within 24 h of reperfusion, distal tubules showed a marked increase in expression of Bcl-2 and a moderate increase in Bcl-X(L) and Bax. Proximal tubules showed a marked increase in Bax expression and a moderate increase in Bcl-X(L). Twenty-four hours after expression of the Bcl-2 proteins was increased, IGF-1 and EGF protein levels were increased in the distal tubule, similar to the Bcl-2 anti-apoptotic proteins, and were also detected in the adjacent proximal tubules, suggestive of paracrine action in these tubules. TGF-beta expression was moderately increased in regenerating proximal tubules, but no relationship was seen with the pattern of expression of the Bcl-2 genes. An explanation of these results is that the distal tubule is adaptively resistant to ischemic injury via promotion of survival by anti-apoptotic Bcl-2 genes, and its survival allows expression of growth factors critical not only to the maintenance and regeneration of its own cell population (autocrine action), but also to the adjacent ischemia-sensitive proximal tubular cells (paracrine action). PMID- 10703670 TI - Downregulation of SPARC expression is mediated by nitric oxide in rat mesangial cells and during endotoxemia in the rat. AB - Nitric oxide (NO) has been implicated in several forms of glomerulonephritis. In this study, a low stringency reversed transcription/PCR protocol was used to evaluate the action of NO on the mRNA expression pattern in rat mesangial cells (MC). To mimic the state of glomerular inflammation, MC were stimulated by exposure to the cytokines interleukin-1beta and tumor necrosis factor-alpha into producing high levels of NO via expression of inducible nitric oxide synthase (NOS). To detect NO-mediated effects, the resulting expression pattern was compared to that of MC stimulated by the cytokines in the presence of the NOS inhibitor N(G)-monomethyl-L-arginine (L-NMMA). Computer analysis of a differentially expressed cDNA fragment resulted in a 100% homology to the recently characterized mRNA of SPARC (secreted protein acidic and rich in cysteine). Further characterization of SPARC regulation revealed a cytokine- and cAMP-dependent decrease in SPARC mRNA and protein levels. Blocking NO formation by L-NMMA reversed the effects of cytokines and cAMP on SPARC expression, suggesting an NO-mediated mechanism. The NO donors S-nitroso-N-acetyl penicillamine and diethylenetriamine/NO further reduced SPARC expression in cytokine-treated MC as well as in controls. Moreover, downregulation of SPARC mRNA and protein expression in whole kidneys obtained from rats treated with endotoxin was observed. This downregulation of SPARC was reversed by treatment with L-N6-l (iminoethyl) lysine dihydrochloride, a potent and highly selective inhibitor of inducible NOS. These data characterize SPARC as an NO-regulated gene. This observation may be important in the context of tissue remodeling in chronic inflammatory kidney diseases. PMID- 10703671 TI - Effect of angiotensin-converting enzyme inhibition on glomerular basement membrane permeability and distribution of zonula occludens-1 in MWF rats. AB - The mechanism(s) by which angiotensin-converting enzyme (ACE) inhibitors prevent glomerular membrane loss of permselective function is still not understood. In male MWF rats, which develop spontaneous proteinuria with age, ACE inhibitors prevent proteinuria and increase glomerular ultrafiltration coefficient. These renoprotective effects are not associated with ultrastructural changes of capillary wall components. This study was undertaken to investigate whether ACE inhibitors modulate functional properties of glomerular basement membrane (GBM) and/or of epithelial cells, both of which have been suggested to play a role in the maintenance of the glomerular filtration barrier. The hydraulic and macromolecular permeability of the GBM were determined, by an in vitro filtration system, in untreated or lisinopril-treated rats and in Wistar rats taken as controls. By indirect immunofluorescence and immunoelectron microscopy, glomerular distribution of the tight junction protein zonula occludens- (ZO-1), a component of the slit diaphragm, was also studied. Results document that spontaneous proteinuria in MWF rats develops without significant changes in the permeability of the GBM to water and albumin, or in the ultrastructure of the podocyte foot processes, but is associated with an important alteration in the distribution of ZO-1 at the glomerular level. Lisinopril, which prevented proteinuria, also prevented glomerular redistribution of the protein. Thus, renoprotective effects of ACE inhibitors are not associated with changes in intrinsic functional properties of GBM, or ultrastructural changes of the epithelial cells, but rather with preservation of glomerular ZO-1 distribution and slit diaphragm function, which are essential for maintaining the filtration barrier. PMID- 10703672 TI - Chronic angiotensin II infusion but not bradykinin blockade abolishes the antiproteinuric response to angiotensin-converting enzyme inhibition in established adriamycin nephrosis. AB - Angiotensin-converting enzyme (ACE) inhibition reduces proteinuria in established adriamycin nephrosis. To investigate whether the reduction in proteinuria is due to decreased generation of angiotensin II (AngII) or to decreased degradation of bradykinin, four series of experiments in established adriamycin nephrosis were performed. In the first series, 2 mg/kg lisinopril reduced BP from 117 +/- 4 to 67 +/- 2 mmHg and proteinuria from 335 +/- 66 to 57 +/- 10 mg/24 h after 2 wk of treatment. Subsequent continuous intraperitoneal infusion of AngII (250 ng/kg per min) for 2 wk partially restored proteinuria to 180 +/- 42 mg/24 h, whereas BP increased to 97 +/- 3 mmHg. Subsequent withdrawal of AngII restored the antiproteinuric effects of lisinopril, whereas subsequent withdrawal of lisinopril restored proteinuria to pretreatment values. In the second series, AT1 receptor blockade induced a fall in BP and proteinuria similar to that by lisinopril. In the third series, lisinopril reduced BP from 121 +/- 5 to 68 +/- 2 mmHg and proteinuria from 355 +/- 90 to 101 +/- 10 mg/24 h. Subsequent intraperitoneal infusion of bradykinin antagonist (HOE 140; 1 mg/kg per 24 h) for 2 wk did not affect BP (72 +/- 2 mmHg) or proteinuria (92 +/- 15 mg/24 h). In the fourth series, bradykinin (3 mg/kg per 24 h) was infused for 2 wk to mimic decreased bradykinin breakdown. This did not affect proteinuria, but induced a fall in BP from 114 +/- 3 to 93 +/- 4 mmHg. The BP-lowering effect of exogenous bradykinin was completely reversed by 1 wk infusion of HOE 140 (93 +/- 4 to 113 +/- 4 mmHg), while proteinuria remained unchanged. In conclusion, the antiproteinuric effect of ACE inhibition appears to be independent of bradykinin in this model, supporting a main role for reduction of AngII in the antiproteinuric action of ACE inhibition. PMID- 10703673 TI - Functional and structural correlates of glomerulosclerosis after renal mass reduction in the rat. AB - Previously, it was shown that 5/6 renal mass reduction by surgical excision (RK NX) results in a marked reduction of glomerulosclerosis (GS) at 6 wk compared with the conventional 5/6 renal ablation by infarction (RK-I) model. To determine the pathogenetic correlates of the striking differences in GS, radiotelemetrically measured BP; single nephron function; glomerular volume; the temporal expression of mRNA for renin, transforming growth factor-beta, and platelet-derived growth factor-B; and plasma renin concentration were compared between RK-NX, RK-I, and sham-operated control rats. Hypertension only developed in the RK-I model, was present at 3 d after infarction, and was correlated with both an increased expression of renin mRNA by Northern analysis and elevated plasma renin concentration. Structural (glomerular volume) and functional (single nephron blood flow and GFR) indices of the compensatory adaptive response were significantly but similarly increased in the RK-NX and RK-I rats compared with sham-operated controls, indicating that these adaptations per se are not responsible for the initiation of GS after 5/6 renal mass reduction. Glomerular capillary pressure (P(GC)) was also significantly increased in both RK-I (56 +/- 2 mmHg) and RK-NX rats (50 +/- 0.9 mmHg) compared with controls (46 +/- 0.8 mmHg, P < 0.01), but the increase was significantly greater in RK-I versus RK-NX rats (P < 0.05) consistent with the higher BP in RK-I rats. These data indicate that differences in renin probably account for the early divergence of BP (and P(GC)) responses between RK-I and RK-NX models. Transforming growth factor-beta and platelet-derived growth factor-B mRNA expression in pooled RNA from kidneys from each group showed increases at 21 d along with early evidence of glomerular injury in the RK-I group but not in the RK-NX group, consistent with their postulated roles as molecular mediators of GS, but only in rats with pathologic glomerular hypertension. PMID- 10703674 TI - Lipoprotein(a) in the nephrotic syndrome: molecular analysis of lipoprotein(a) and apolipoprotein(a) fragments in plasma and urine. AB - Plasma levels of lipoprotein(a) (Lp(a)), an atherogenic particle, are elevated in kidney disease, which suggests a role of this organ in the metabolism of Lp(a). Additional evidence for a role of the kidney in the clearance of Lp(a) is provided by the fact that circulating N-terminal fragments of apolipoprotein(a) (apo(a)) are processed and eliminated by the renal route. To further understand the mechanism underlying such renal excretion, the levels of apo(a) fragments in plasma and urine relative to plasma Lp(a) levels were determined in patients with nephrotic syndrome (n = 15). In plasma, the absolute (24.7 +/- 20.4 versus 2.16 +/- 2.99 microg/ml, P < 0.0001) as well as the relative amounts of apo(a) fragments (4.6 +/-3.4% versus 2.1 +/- 3.3% of total Lp(a), P < 0.0001) were significantly elevated in nephrotic patients compared with a control, normolipidemic population. In addition, urinary apo(a) excretion in patients with nephrotic syndrome was markedly elevated compared with that in control subjects (578 +/- 622 versus 27.7 +/- 44 ng/ml per mg creatinine, P < 0.001). However, the fractional catabolic rates of apo(a) fragments were similar in both groups (0.68 +/- 0.67% and 0.62 +/- 0.47% in nephrotic and control subjects, respectively), suggesting that increased plasma concentrations of apo(a) fragments in nephrotic subjects are more dependent on the rate of synthesis rather than on the catabolic rate. Molecular analysis of apo(a) immunoreactive material in urine revealed that the patterns of apo(a) fragments in nephrotic patients were distinct from those of control subjects. Full-length apo(a), large N-terminal apo(a) fragments similar in size to those present in plasma, as well as C-terminal fragments of apo(a) were detected in urine from nephrotic patients but not in urine from controls. All of these apo(a) forms were in addition to smaller N-terminal apo(a) fragments present in normal urine. This study also demonstrated the presence of Lp(a) in urine from nephrotic patients by ultracentrifugal fractionation. These data suggest that in nephrotic syndrome, Lp(a) and large fragments of apo(a) are passively filtered by the kidney through the glomerulus, whereas smaller apo(a) fragments are secreted into the urine. PMID- 10703675 TI - Naltrexone does not relieve uremic pruritus: results of a randomized, double blind, placebo-controlled crossover study. AB - Improvement of uremic pruritus was reported under short-term administration of the mu-receptor antagonists naltrexone and naloxone. The aim of the present study was to confirm the efficacy and safety of the oral mu-receptor antagonist naltrexone during a 4-wk treatment period in patients on hemodialysis and peritoneal dialysis. A placebo-controlled, double-blind crossover study of uremic patients with persistent, treatment-resistant pruritus was performed. Of 422 patients screened between December 1997 and June 1998, 93 suffered from pruritus and 23 were eligible for the study. Patients were started either with a 4-wk naltrexone sequence (50 mg/d) or matched placebo. This was followed by a 7-d washout, and patients continued with a 4-wk sequence of the alternate medication. Pruritus intensity was scored daily by a visual analogue scale (VAS) and weekly by a detailed score assessing scratching activity, distribution of pruritus, and frequency of pruritus-related sleep disturbance. Sixteen of 23 patients completed the study. During the naltrexone period, pruritus decreased by 29.2% (95% confidence interval [CI], 18.7 to 39.6) on the VAS and by 17.6% (95% CI, 4.2 to 31.1) on the detailed score. In comparison, pruritus decreased by 16.9% (95% CI, 6.8 to 26.9) on the VAS and by 22.3% (95% CI, 9.3 to 35.2) on the detailed score during the placebo period. The difference between the naltrexone and the placebo treatment period was not statistically significant. Nine of 23 patients complained of gastrointestinal disturbances during the naltrexone period compared with only one of 23 patients during the placebo period (P < 0.05). These results show that treatment of uremic pruritus with naltrexone is ineffective. In addition, a high incidence of adverse effects was observed during treatment with naltrexone. PMID- 10703676 TI - Validation of comorbid conditions on the end-stage renal disease medical evidence report: the CHOICE study. Choices for Healthy Outcomes in Caring for ESRD. AB - Since 1995, the Medical Evidence Report for end-stage renal disease (Form 2728) has been used nationally to collect information on comorbid conditions. To date, these data have not been validated. A national cross-sectional study of 1005 incident dialysis patients (734 hemodialysis and 271 peritoneal dialysis) enrolled between October 1995 and June 1998 was conducted using clinical data to validate 17 comorbid conditions on Form 2728. Sensitivity and specificity were calculated for each condition. The relationship between patient characteristics and sensitivity was assessed in multivariate analysis. Sensitivity was fairly high (0.67 to 0.83) for HIV disease, diabetes, and hypertension; intermediate (0.40 to 0.52) for peripheral vascular disease, neoplasm, myocardial infarction, cerebrovascular disease, coronary artery disease, cardiac arrest, and congestive heart failure; and poor (<0.36) for dysrhythmia, ambulation status, pericarditis, chronic obstructive pulmonary disease, and smoking. Sensitivity did not change significantly over calendar time. The sensitivity of Form 2728 averaged across all 17 conditions was 0.59 (95% confidence interval, 0.43 to 0.75). The average sensitivity was 0.10 greater in peritoneal dialysis than hemodialysis patients. 0.11 greater in diabetic patients than nondiabetic patients, and 0.04 less with each added comorbid condition. The specificity was very good for hypertension (0.91) and excellent (>0.95) for the other 16 conditions. Comorbid conditions are significantly underreported on Form 2728, but diagnoses are not falsely attributed to patients. Scientific research, quality of care comparisons, and payment policies that use Form 2728 data should take into account these limitations. Considerable effort should be expended to improve Form 2728 coding if it is to provide accurate estimates of total disease burden in end-stage renal disease patients. PMID- 10703677 TI - Optimization of epoetin therapy with intravenous iron therapy in hemodialysis patients. AB - Iron deficiency limits the efficacy of recombinant human erythropoietin (rhEPO) therapy in end-stage renal disease (ESRD) patients. Functional iron deficiency occurs with serum ferritin >500 ng/ml and/or transferrin saturation (TSAT) of 20 to 30%. This study examines the effects of a maintenance intravenous iron dextran (ivID) protocol that increased TSAT in ESRD hemodialysis patients from conventional levels of 20 to 30% (control group) to those of 30 to 50% (study group) for a period of 6 mo. Forty-two patients receiving chronic hemodialysis completed a 16- to 20-wk run-in period, during which maintenance ivID and rhEPO were administered in amounts to achieve average TSAT of 20 to 30% and baseline levels of hemoglobin of 9.5 to 12.0 g/dl. After the run-in period, 19 patients randomized to the control group received ivID doses of 25 to 150 mg/wk for 6 mo. Twenty-three patients randomized to the study group received four to six loading doses of ivID, 100 mg each, over a 2-wk period to achieve a TSAT >30% followed by 25 to 150 mg weekly to maintain TSAT between 30 and 50% for 6 mo. Both regimens were effective in maintaining targeted hemoglobin levels. Fifteen patients in the control group and 17 patients in the study group finished the study in which the primary outcome parameter by intention to treat analysis was the rhEPO dose needed to maintain prestudy hemoglobin levels. Maintenance ivID requirements in the study group increased from 176 to 501 mg/mo and were associated with a progressive increase in serum ferritin to 658 ng/ml. Epoetin dose requirements for the study group decreased by the third month and remained 40% lower than for the control group, resulting in an overall cost savings in managing the anemia. Secondary indicators of iron-deficient erythropoiesis were also assessed. Zinc protoporphyrin did not change in either group. Reticulocyte hemoglobin content increased only in the study group from 28.5 to 30.1 pg. It is concluded that maintenance of TSAT between 30 and 50% reduces rhEPO requirements significantly over a 6-mo period. PMID- 10703678 TI - Vitamin E attenuates oxidative stress induced by intravenous iron in patients on hemodialysis. AB - Intravenous iron application to anemic patients on hemodialysis leads to an "oversaturation" of transferrin. As a result, non-transferrin-bound, redox-active iron might induce lipid peroxidation. To test the hypothesis that vitamin E attenuates lipid peroxidation in patients receiving 100 mg of iron(III) hydroxide sucrose complex intravenously during a hemodialysis session, 22 patients were investigated in a randomized cross-over design, either with or without a single oral dose of 1200 IU of all-rac-alpha-tocopheryl acetate taken 6 h before the hemodialysis session. Blood was drawn before and 30, 60, 90, 135, and 180 min after the start of the iron infusion, and areas under the curve (AUC0-180 min) of ratios of plasma malondialdehyde (MDA) to cholesterol and plasma total peroxides to cholesterol (two markers of lipid peroxidation) were determined as the outcome variables. At baseline of the session without vitamin E supplementation, plasma alpha-tocopherol concentrations (27.6 +/- 1.8 micromol/L) and ratios of alpha tocopherol to cholesterol (5.88 +/- 1.09 mmol/mol) were normal, plasma MDA concentrations were above normal (1.20 +/- 0.28 micromol/ L), and bleomycin detectable iron (BDI), indicating the presence of redox-active iron, was not detectable. Upon iron infusion, BDI and MDA concentrations increased significantly (P < 0.001). BDI concentrations explained the increase over baseline in MDA concentrations (MDA = 1.29 +/- 0.075 x BDI). Vitamin E supplementation, leading to a 68% increase in plasma alpha-tocopherol concentrations, significantly reduced the AUC0-180 min of MDA to cholesterol (P = 0.004) and peroxides to cholesterol (P = 0.002). These data demonstrate that a single oral dose of vitamin E attenuates lipid peroxidation in patients on hemodialysis receiving intravenous iron. Given that intravenous iron is applied repeatedly to patients on hemodialysis, this therapeutic approach may protect against oxidative stress-related degenerative disease in the long term. PMID- 10703679 TI - Effect of intravenous fluids on blood pressure course during hemodialysis in hypotensive-prone patients. AB - Hypertonic and hyperoncotic solutions are generally used as acute treatment for symptomatic hypotension during dialysis. Administration of hydroxyethylstarch (HES) was recently shown to be an effective substitution fluid in preserving blood volume (BV) and systolic BP (SBP) in a group of stable dialysis patients during dialysis. In this study, in nine cardiac-compromised dialysis patients with frequent symptomatic hypotensive episodes, the efficacy of three fluids (hypertonic saline [3%], albumin [20%], and HES [10%]) was assessed during three treatment sessions with combined ultrafiltration and hemodialysis, which only differed in the type of fluid administered intravenously. Changes in SBP and relative BV were compared. Fluids were given when SBP was less than 100 mmHg or when the decrease in SBP was more than 25 mmHg versus the start of the treatment. The ultrafiltration was continued at the same rate. When comparing SBP at the end of the dialysis session (t = end) with that at the time of infusion (t = iv), SBP decreased with saline, increased with albumin, and increased significantly with HES. The change in SBP in t = end versus t = iv was significantly greater when using saline compared with HES, and tended to decrease more when using saline compared with albumin (P = 0.09). Between albumin and HES there were no significant differences. BV decreased significantly (t = end) versus baseline (t = 0) during ultrafiltration and hemodialysis in all three treatment sessions. The decrease was significantly higher when using saline compared with albumin and saline compared with HES. Between albumin and HES there were no significant differences. When the values at t = end were compared with those at t = iv, BV decreased, although not significantly, with saline and albumin, but remained unchanged with HES. It is concluded that HES is an effective fluid in maintaining SBP and preserving BV in hypotensive-prone dialysis patients, comparable to albumin but superior to hypertonic saline. PMID- 10703680 TI - Predictors of loss of residual renal function among new dialysis patients. AB - Residual renal function (RRF) in end-stage renal disease is clinically important as it contributes to adequacy of dialysis, quality of life, and mortality. This study was conducted to determine the predictors of RRF loss in a national random sample of patients initiating hemodialysis and peritoneal dialysis. The study controlled for baseline variables and included major predictors. The end point was loss of RRF, defined as a urine volume <200 ml/24 h at approximately 1 yr of follow-up. The adjusted odds ratios (AOR) and P values associated with each of the demographic, clinical, laboratory, and treatment parameters were estimated using an "adjusted" univariate analysis. Significant variables (P < 0.05) were included in a multivariate logistic regression model. Predictors of RRF loss were female gender (AOR = 1.45; P < 0.001), non-white race (AOR = 1.57; P = <0.001), prior history of diabetes (AOR = 1.82; P = 0.006), prior history of congestive heart failure (AOR = 1.32; P = 0.03), and time to follow-up (AOR = 1.06 per month; P = 0.03). Patients treated with peritoneal dialysis had a 65% lower risk of RRF loss than those on hemodialysis (AOR = 0.35; P < 0.001). Higher serum calcium (AOR = 0.81 per mg/dl; P = 0.05), use of an angiotensin-converting enzyme inhibitor (AOR = 0.68; P < 0.001). and use of a calcium channel blocker (AOR = 0.77; P = 0.01) were independently associated with decreased risk of RRF loss. The observations of demographic groups at risk and potentially modifiable factors and therapies have generated testable hypotheses regarding therapies that may preserve RRF among end-stage renal disease patients. PMID- 10703681 TI - Time dependency of factors affecting renal allograft survival. AB - The function of renal transplants can deteriorate at any time posttransplant, but the risks and mechanisms may differ at different times posttransplant. Survival of 522 consecutive cadaveric renal transplant recipients followed for at least 6 mo were analyzed, with patient death censored. The overall risk factors in univariate analysis were acute rejection requiring antibody therapy (AR), delayed graft function, elevated serum creatinine at 6 mo, high panel-reactive antibodies, and donor age > or =55 yr, with borderline effects of recipient age and female gender. These risks were studied in each of three intervals posttransplantation: < or =6 mo, 6 mo to 5 yr, and >5 yr. Of the 135 graft failures, 53 occurred < or =6 mo, 61 between 6 mo and 5 yr, and 21 beyond 5 yr. By multivariate analysis. the risks for graft failure in interval < or =6 mo were AR (hazard ratio (HR) = 4.86, P < 0.001); delayed graft function (HR = 1.47, P = 0.06): and high panel-reactive antibodies (HR = 2.04, P = 0.0(3). Between 6 mo and 5 yr, the risks for graft loss were AR (HR = 2.87, P < 0.001) and serum creatinine at 6 mo > or =150 micromol/L (HR = 3.69, P < 0.001). Beyond 5 yr the risk factors were donor age > or =55 yr (HR = 5.87, P = 0.002), with a borderline effect of kidneys from female donors (HR = 2.28, P = 0.07). HLA-A, -B, and -DR matching and presensitization had most of their effect through early AR and impaired function. The results indicate that risks for graft loss are time dependent: early losses correlate with injury and rejection, but late events correlate with donor age and possibly workload. PMID- 10703683 TI - Long-term complications in renal transplantation. PMID- 10703682 TI - Matrix metalloproteinases in renal development and disease. PMID- 10703684 TI - Role of the kidney in erythropoiesis. 1957. PMID- 10703685 TI - Challenging issues associated with organ transplantation for Jehovah's Witness individuals. PMID- 10703686 TI - Implantable left ventricular assist devices can successfully bridge adolescent patients to transplant. AB - BACKGROUND: Left ventricular assist devices (LVAD) have been used successfully as a life-sustaining bridge to transplantation in adults with end-stage heart failure. Long-term implantable cardiac assist devices for smaller adolescent patients are not yet available in the United States. METHODS: This study reviews the experience with patients less than 21 years old that received HeartMate LVADs (TCI) at our institution. Twelve patients were implanted with 13 LVADs. The patients ranged in age from 11 to 20 years (mean 16 years). Body surface area ranged from 1.4 to 2.2 m2 (mean 1.8 m2). Patients were selected for LVAD placement based on eligibility for heart transplant and evidence of end-organ dysfunction. Device placement in small patients was facilitated with prosthetic graft abdominal wall closure. No patient received systemic anticoagulation. RESULTS: The duration of LVAD support ranged from 0 to 397 days (mean 123 days). Seven of the 8 patients eligible for discharge from the hospital with a vented electric LVAD were supported at home while awaiting transplantation. Outcomes of LVAD support were: LVAD explantation in 2 cases (15%), expiration with LVAD in place in 3 cases (23%), and successful transplantation in 8 cases (62%). Complications included 4 patients with systemic infection, 3 re-operations for hemorrhage, 1 embolic event, and 1 intraoperative air embolus that proved fatal. One explanted patient required a subsequent LVAD and the other expired 4 months after explantation. Six of the 8 transplanted patients are alive and well with follow-up ranging from 8 to 43 months. CONCLUSIONS: Adolescent patients with heart failure can be successfully supported on a long-term basis to heart transplantation with the HeartMate LVAD. The wearable device allows for discharge home while awaiting transplantation. Device explantation without subsequent transplantation can be unpredictable. The incidence of thromboembolism remains low despite the absence of systemic anticoagulation. The technique of prosthetic graft closure of the abdominal wall facilitates the use of this device in smaller patients. PMID- 10703687 TI - ACE inhibitor dosage at the time of listing predicts survival. AB - BACKGROUND: To be listed for heart transplantation (HTx), optimization of the dosage of angiotensin converting enzyme (ACE) -inhibitors is recommended worldwide even though this issue has not been thoroughly investigated in the pre transplantation cohort. OBJECTIVE: The aim of this database study was to analyze the prognostic impact of a pre-defined high vs a low ACE inhibitor dose range at the time of listing for elective HTx in addition to various previously established prognostic factors. METHODS: Medical records from 237 patients (84% male, mean age 54 years) admitted between January 1995 and January 1998 from 25 different centers in Austria were reviewed. Forty-seven percent were taking > or =75 mg captopril, > or =20 mg enalapril, > or =20 mg lisinopril or > or =5 mg ramipril daily ("high-dose" group) and 53% received smaller doses ("low-dose" group). RESULTS: No significant differences between groups were detected at baseline except that patients with higher ACE inhibitor doses were more likely to take nitrates, beta-blockers and amiodarone, received higher furosemide doses and had higher serum gamma-glutamyl transferase levels. Follow-up was 328 days (248 SD) with 16% deaths in the "high-dose" group vs 288 days (270 SD) with 25% deaths in the "low-dose" group. Kaplan-Meier survival curves demonstrated a significant difference over time between the two treatment groups (P = 0.03). Furthermore, dichotomized ACE inhibitor treatment at the time of listing was the strongest independent single predictor of mortality (P = 0.01) with only blood pressure (P = 0.02), alanine transaminase (P = 0.02) and left ventricular end diastolic diameter (P = 0.02) providing additional prognostic information. To explain these findings several factors have to be considered a) greater benefit with higher ACE inhibitor doses b) sicker patients receiving lower ACE inhibitor doses and c) more experienced heart failure care of the "high-dose" group. CONCLUSIONS: Heart transplantation candidates who, for whatever reason, receive ACE-inhibitors below the recommended dosages, are at increased mortality risk and thus merit greater scrutiny. PMID- 10703688 TI - Absence of enteroviral RNA in hearts explanted from patients with dilated cardiomyopathy. AB - BACKGROUND: The role of enterovirus infection in the pathogenesis of dilated cardiomyopathy (DCM) remains unclear. The objective of this study was to determine the prevalence of enterovirus in hearts explanted from patients with DCM and to compare it with enterovirus prevalence in hearts explanted from patients with other etiologies and in healthy donor hearts. METHODS: A total of 138 cardiac samples were analyzed, 70 from heart donors and 68 from transplant recipients (22 with DCM). A highly sensitive enterovirus-specific nested RT-PCR was used to test for enterovirus. RESULTS: All tests were negative except for one positive result that was attributed to carryover because sequencing of the amplification product showed it to be identical to the positive control. CONCLUSIONS: In this study the sample of explanted hearts nested RT-PCR showed no evidence of the presence of enteroviral RNA. This suggests that if enterovirus had a role in the genesis of DCM, it does not require or lead to the persistence of the virus in myocardial tissue. PMID- 10703689 TI - Nitric oxide synthase II mRNA expression in cardiac tissue of patients with heart failure undergoing cardiac transplantation. AB - OBJECTIVES: To examine whether inducible nitric oxide synthase is expressed in myocardial tissue of patients with heart failure. BACKGROUND: There is increasing evidence that alterations in nitric oxide synthesis are of pathophysiologic importance in heart failure. Nitric oxide (NO) can exert negative inotropic and cytotoxic effects on cardiomyocytes. A number of studies have shown altered nitric oxide production by the endothelial constitutive isoform of nitric oxide synthase (NOS III), but there is little information on the role of NOS II. Expression of NOS II could lead to excessive production of NO in the myocardium and affect cardiac contractility. METHODS: NOS II mRNA expression in myocardial tissue of 18 patients with idiopathic dilated cardiomyopathy (DCM), 7 patients with ischemic cardiopathy and severe ventricular dysfunction (ISCH), 4 patients with acute myocardial infarction (AMI) and 11 controls. Serum concentration of NO2-/NO3- (NOx) was also measured. RESULTS: NOS II gene expression occurred in all the patients with DCM, in 1 out of the 7 ISCH patients, in 2 out of the 4 patients with AMI and in none of the controls. Moreover, DCM patients showed a significant 6-fold increase in NOx concentration (253+/-47 nm/ml) as compared to controls (40+/-2 nm/ml) P < 0.001, a phenomenon not observed in ISCH patients (56+/-3 nm/ml). CONCLUSIONS: NOS II expression occurs in failing human cardiac myocytes and can play an specific role in the pathogenesis of DCM. PMID- 10703690 TI - Accuracy of echocardiographic right ventricular parameters in patients with different end-stage lung diseases prior to lung transplantation. AB - BACKGROUND: Because there are few data available on the accuracy of 2D echocardiography to assess right ventricular (RV) size and function in patients with far-advanced lung disease, in this prospective study, we compared various echocardiographic RV parameters with RV volumes derived from magnetic resonance imaging (MRI). METHODS: In 32 patients (18 male, 17 female) presenting for lung transplantation, we measured RV end-diastolic and end-systolic area as well as derived RV fractional area change, long-axis diameter, short-axis diameter, tricuspid valve anulus diameter (using 2D apical or sub-costal 4-chamber view), and RV end-diastolic diameter (using M-mode in the parasternal short-axis view). These values were compared with RV end-diastolic and end-systolic volumes derived by MRI, serving as the gold standard. RESULTS: Right ventricular end-diastolic area was the most accurate echocardiographic parameter of RV size (correlation to MRI: r = 0.88, p < 0.001), followed by RV end-diastolic short-axis diameter (r = 0.75, p < 0.001), long axis diameter (r = 0.66, p < 0.001), and tricuspid valve anulus diameter (r = 0.63, p < 0.001). In contrast, M-mode measurement of RV end diastolic diameter was possible in only 24/35 (68%) patients and showed a weak correlation to MRI-derived RV end-diastolic volume (r = 0.56, p = 0.004). Right ventricular fractional area change correlated well with MRI-derived RV ejection fraction (r = 0.84, p < 0.0001). In a sub-group analysis, patients with vascular lung disease showed best agreement between both methods for RV end-diastolic area and RV fractional area change compared with patients with restrictive or obstructive lung disease. CONCLUSION: This study shows that in patients with far advanced lung diseases, RV end-diastolic area demonstrated the best correlation with MRI-derived measurement of RV end-diastolic volume, and RV fractional area change compared favorably with MRI-derived ejection fraction. Despite reduced image quality, especially in patients with obstructive lung disease, these parameters can yield clinically valuable information. PMID- 10703691 TI - High dose rate brachytherapy to prevent recurrent benign hyperplasia in lung transplant bronchi: theoretical and clinical considerations. AB - BACKGROUND: Significant anastomotic stenosis and malacia is reported to affect 7% to 15% of lung transplant recipients. Laser debridement, dilation and stenting can be used effectively to treat the majority of these patients. However, persistent, as well as reactive hyperplastic tissue reaction, will occur in some of these patients, requiring multiple bronchoscopic interventions. The experience of 2 patients who received intraluminal brachytherapy irradiation to prevent recurrence of hyperplastic tissue causing airway obstruction is reported. Both had failed multiple attempts of local control, including wall stent, laser ablation and balloon dilation. They suffered from shortness of breath and progressive decrease in quality of life because of airway obstruction. METHODS: Two patients received intraluminal irradiation immediately following removal of severe post-lung transplant obstruction. Both patients developed airway obstruction 3 to 4 months after left lung transplantation. High Dose Rate (HDR) brachytherapy (192Ir). Afterloader was used to treat Patient 1 on two occasions. Patient 2 required a single treatment. The radiation dose of 3Gy/fraction was calculated at 1 cm from the catheter for all applications. RESULTS: Follow up for both patients included bronchoscopy at 3 weeks, 3 months and 6 months after radiation therapy. Follow up for Patient 1 is 7 months, and patient 2 is 6 months. Each patient had an initial complete response after radiation. There were no treatment-related complications, and both patients experienced significant improvement in respiratory function. CONCLUSIONS: Symptomatic benign airway obstruction from hyperplastic tissue in the bronchus after lung transplantation can be successfully treated with intraluminal radiation therapy. Patients who develop recurrent benign granulation tissue after stent and laser therapy may be considered for this type of treatment. PMID- 10703692 TI - Apoptosis and formation of peroxynitrite in the lungs of patients with obliterative bronchiolitis. AB - BACKGROUND: Obliterative bronchiolitis (OB) is the principal long-term complication of lung and heart-lung transplantation. OB is characterized histologically by inflammation, epithelial cell loss, fibrosis, and obliteration of the terminal airways. The contribution of apoptosis and peroxynitrite formation in OB was examined and assessed whether immunohistochemical markers of these reactions in transbronchial biopsy specimens were predictive of OB development. METHODS: Pulmonary tissue samples from lung transplant recipients with OB (n = 5) or without OB (control group; n = 7) were investigated by in situ terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) and nitrotyrosine immunohistochemistry. Furthermore, TUNEL and nitrotyrosine expression was compared between matched transbronchial biopsy specimens from the two patient groups. RESULTS: Sections with active OB displayed a significantly increased number of TUNEL-positive epithelial cells and macrophages compared with very little TUNEL in control specimens. TUNEL was almost absent in inactive OB. Nitrotyrosine was detected in all samples of pulmonary tissue, but nitrotyrosine expression was more intense in patients with active OB. There was no apparent temporospatial correlation of TUNEL and nitrotyrosine expression, and in matched transbronchial specimens, these immunohistochemical markers failed to identify patients with imminent risk of developing OB. CONCLUSIONS: Apoptosis contributes to the pathophysiology of active OB but is apparently not directly paralleled by tissue peroxynitrite formation. In transbronchial biopsy specimens, markers of apoptosis and peroxynitrite formation are not valid predictors of OB and more studies are required to deliniate the role of these mechanisms in pulmonary allograft rejection. PMID- 10703693 TI - Intravascular ultrasound imaging after cardiac transplantation: advantage of multi-vessel imaging. AB - BACKGROUND: Intravascular ultrasound is a sensitive tool to study transplant vasculopathy. However, there is no consensus regarding the methodology for imaging protocol. The impact of single versus multiple epicardial vessel imaging in determining the prevalence of transplant vasculopathy has not been determined. This study examines the benefit of three-vessel imaging versus one-vessel imaging in detecting transplant vasculopathy. METHODS AND RESULTS: One hundred eleven transplant recipients with intravascular ultrasound imaging at baseline (within 2 months of transplantation) were studied: 107 at 1-year, 53 at 2-year and 41 at 3 year follow-up. A total of 222 arteries, 519 segments and 772 sites were analyzed (94 LAD, 65 LCX and 65 RCA). The prevalence of transplant vasculopathy lesions was 27%, 41% and 58% at 1 year, 39%, 55% and 71% at 2 years and 39%, 55% and 74% at 3 years for patients with one-, two- and three-vessel imaging, respectively. Single- or two-vessel disease was present in 23% (7) and 32% (10) patients with three-vessel imaging, leading to the potential mislabeling of these 17 (55%) patients as "disease free" if they underwent only single-vessel imaging. CONCLUSIONS: Multivessel imaging is more sensitive in detecting the transplant vasculopathy lesions compared to single-vessel imaging. This important variable should be considered when designing and interpreting trials utilizing intravascular imaging derived end-point. PMID- 10703694 TI - 8-Br-cyclic GMP given during reperfusion improves post-transplant lung edema and free radical injury. AB - Substitution of the NO-pathway reduces ischemia/reperfusion injury following lung transplantation. 8-Br-cGMP is a membrane permeable analogue of cGMP, the second messenger of NO. In this study the effect of continuous administration of 8-Br cGMP on early graft function was evaluated. METHODS: Unilateral left lung transplantation was performed in 10 weight-matched pigs (23-30 kg). Donor lungs were flushed with 1.51 cold (1 degree C) LPD solution and preserved for 20 hours. In Group I (n = 5), 8-Br-cGMP (0.2 mg/kg/h) was given continuously over the entire observation time starting 15 min before reperfusion. Group II served as control, no 8-Br-cGMP was administered. In both groups, 250 microg PGE1 was injected into the pulmonary artery (PA) before flush. One hour after reperfusion the recipients contralateral right PA and bronchus were ligated to assess isolated graft function only. Extravascular lung water index (EVLWI), pulmonary vascular resistance, mean PA pressure, mean systemic arterial pressure and gas exchange were assessed during a 5-hour observation period. Lipid peroxidation as indicator for free radical mediated injury and neutrophil migration to the allograft were measured at the end of the assessment. RESULTS: EVLWI was significantly reduced in animals treated with 8-Br-cGMP (overall difference P = 0.024) with a peak 2 hours after reperfusion (Group I, 8.2+/-0.3 mg/ml vs Group II, 10.1+/-0.6 mg/ml; P = 0.039). Also in Group I the free radical mediated tissue injury was significantly lower when compared to Group II (Group I, 61.8+/ 12.3 pmol/g vs Group II, 120.7+/-7.2 pmol/g; P = 0.006). A tendency towards a reduced neutrophil migration after 8-Br-cGMP infusion was shown; however, the changes in comparison to the control animals were not statistically significant (Group I, 1.0+/-0.2 deltaOD/mg/min vs Group II, 1.7+/-0.3 deltaOD/mg/min; P = 0.13). Pulmonary- and systemic hemodynamics, and allograft gas exchange did not differ between groups. CONCLUSIONS: The results indicate that substitution of the NO pathway by administration of the second messenger cGMP at the time of reperfusion improves post-transplant lung allograft function. PMID- 10703695 TI - Differential time scale of fluid and solute permeability following hypothermic lung preservation. AB - BACKGROUND: Assessment of the quality of lung graft preservation by simple functional measures in some laboratory models may fail to detect endothelial injury. The effects of hypothermic preservation in isolation were investigated by measuring the pulmonary capillary filtration coefficient (Kf) and the albumin surface area product (PS) at various cold ischemic intervals. METHODS: Rat lungs were flushed with University of Wisconsin solution at 4 degrees C. Following storage at 4 degrees C, lungs for Kf measurement were subjected to a change in pulmonary arterial pressure. Kf was calculated from the change in rate of weight gain as a function of hydrostatic stress. PS lungs were exposed to Tris buffered Ringer's solution containing 1125 albumin (20 microM) in an isogravimetric state. Following a vascular flush the lungs were homogenized and underwent scintillation counting. Using the Kedem-Katchalsky equation PS was calculated. RESULTS: The Kf for the control, 4-hour, and 7-hour groups were 0.778, 1.816, 4.853 g/ cm H2O/min/100 g wet lung tissue, respectively. There was a significant increase in Kf with each time increment (P,0.01). The Kf for the 24-hour group was 5.587 g/cm H2O/min/100 g wet lung tissue; not an additional significant increase. PS for the control and 4-hour groups (0.0115 and 0.0101 cm3/g wet lung tissue/minute, respectively) were not significantly different. After 7 hours there was a significant increase to 0.171 cm3/g wet lung tissue/min. PS could not be measured after 24 hours. CONCLUSIONS: Significant endothelial injury occurs after 4 hours of cold ischemic preservation. There is progressive injury with time. Increase in water permeability is not secondary to increase in albumin permeability. PMID- 10703696 TI - Beneficial effects of novel nitric oxide donor (FK409) on pulmonary ischemia reperfusion injury in rats. AB - BACKGROUND: Nitric oxide (NO) seems to play an important role in tissue injury during reperfusion of the lung. FK409 is the first spontaneous NO donor that increases plasma guanosine 3':5'-cyclic monophosphate. It is reported that FK409 prevented myocardial infarction following occlusion and reperfusion in rat coronary arteries. In this study, we evaluated the effects of FK409 on pulmonary ischemia-reperfusion injury in an in situ warm ischemia model of rats. METHODS: Animals were divided into 2 groups: the FK409 study group that was administered FK409 (0.4 mg/kg) before reperfusion and the control group, administered a saline vehicle only. Following a thoracotomy, the bronchus, pulmonary artery and vein were separately clamped for 1 hour. Arterial oxygen tension (PaO2), arterial oxygen saturation (SaO2), and endothelin-I (ET-I) were measured after 2 hours of reperfusion. Histologic and immunohistochemical studies were performed; polymorphonuclear neutrophils (PMNs) were counted after 2 hours of reperfusion. RESULTS: PaO2, SaO2, ET-I after 2 hours of reperfusion and the 7-day survival rate were significantly (p < 0.05) better in the FK409 group than the control group. Histologic damage was reduced in the FK409 group compared with the control group. PMN infiltration was also significantly (p < 0.05) lower in the FK409 group than in the control group. CONCLUSION: FK409 seems to protect against ischemia-reperfusion injury of the lung. This effect may be related to a homeostatic effect on pulmonary vascular beds and prevention of PMN sequestration. PMID- 10703697 TI - Prevention of small airway obliteration in a swine heterotopic lung allograft model. AB - BACKGROUND: In our swine model of obliterative bronchiolitis preventing obliteration by the standard immunosuppression with cyclosporine, methylprednisolone, and azathioprine was not successful. The purpose of this study was to test the ability of a new immunosuppressive regimen to prevent alloimmune reaction and obliteration of the allografts. This regimen includes the novel macrolide SDZ RAD, i.e., 40-O-(2hydroxyethyl)-rapamycin. METHODS: Donor lung allografts of 1 cm3 were implanted sub-cutaneously into 11 random-bred non related domestic pigs receiving daily oral cyclosporine (10 mg/kg) and methylprednisolone (20 mg). In addition, the animals received either oral azathioprine (2 mg/kg) (Group 1) or oral SDZ RAD (1.5 mg/kg) (Group 2). Histologic alterations were graded from 0 to 3 based on repeatedly removed implants during a follow-up period of 3 months. RESULTS: Total epithelial destruction and permanent luminal obliteration occurred within 37 days in Group 1. After an initial grade of 2.3+/-0.3 destruction, epithelial recovery was evident in Group 2 (P < 0.01), and the bronchi stayed patent. Cartilaginous destruction was milder in Group 2 (P < 0.05) than in Group 1, but chondrocytic proliferation was more intense (P < 0.05). Alveolar tissue and native structures of the bronchial wall were destroyed in Group 1, but preserved in Group 2 with total recovery after a mild-grade initial necrosis. CONCLUSIONS: Unlike the standard triple therapy, SDZ RAD combined with cyclosporine and methylprednisolone preserves the pulmonary allografts and prevents epithelial destruction and subsequent luminal obliteration. This suggests that this regimen might efficiently suppress obliterative bronchiolitis and improve long-term results in lung transplant recipients. PMID- 10703698 TI - Is the prognosis poorer in heart transplanted patients who develop a right bundle branch block? AB - BACKGROUND: Currently studies conflict on the impact on mortality of right bundle branch block development after transplantation. Most studies conclude that right bundle branch block does not affect patient survival. However, no distinction is made between patients in whom right bundle branch block progresses and those in whom it remains unchanged during follow-up. The objective of this study is to assess clinical or survival differences between patients who develop right bundle branch block and those who do not, and also to analyze these differences depending on progression of this conduction abnormality. MATERIALS AND METHODS: Ninety-seven consecutive heart transplant recipients with more than 1 year's survival were analyzed. Twelve-lead standard ECGs were performed during the first week after transplantation, which allowed for classification of patients depending on the presence or absence of right bundle branch block. Subsequently, throughout the first year, 2 groups were identified, depending on increase of the conduction defect. The groups were compared and factors determining the presence of right bundle branch block and progression of the conduction defect were found. Survival curves for the conduction defect were also compared. RESULTS: Fifty percent of the patients developed right bundle branch block after transplantation; it was progressive in 10. Progressive right bundle branch block was related to greater renal dysfunction (odds ration [OR] = 10.8; confidence interval [CI] = 2-58; p = 0.006), a larger number of rejections (p = 0.01), and a greater death rate (OR = 12.8; CI = 2.5-64; p = 0.002). The presence of progressive right bundle branch block was an independent predictor of long-term mortality (OR = 27.9; CI = 4.2-186.3; p = 0.0006). CONCLUSIONS: The development of right bundle branch block after transplantation is related to intraoperative factors and to a greater number of rejections. The presence of this conduction disorder, particularly if it progresses during the first year, identifies a sub group of patients with a poorer long-term prognosis. PMID- 10703699 TI - Clinical features and treatment of Malassezia folliculitis with fluconazole in orthotopic heart transplant recipients. AB - Orthotopic heart transplant recipients need immunosuppressive treatment and are at an increased risk for opportunistic infections such as Malassezia folliculitis. During a 4-month period (July to October 1990), 11 such cases were identified and treated; all were male with mean age of 43+/-9 years and on standard triple immunosuppressive therapy. Skin scrapings in potassium hydroxide (KOH) preparation with microscopy and/or culture identified either Malassezia furfur or Malassezia pachydermatis as the etiologic agent. A treatment with topical preparation (clotrimazole 1% and selenium sulfide lotion) was effective in 6 patients, whereas the rest received systemic fluconazole treatment with satisfactory outcome; all lesions were resolved within 3 weeks. Fluconazole appears to be an effective agent with excellent therapeutic outcome when administered for 3 weeks. PMID- 10703700 TI - Morbidity and mortality related to the native lung in single lung transplantation for emphysema. AB - It has been advocated that a major drawback of single lung transplantation (SLT) is the risk of serious complications arising from the native lung. The morbidity and mortality related to the native lung in 46 patients who underwent SLT for pulmonary emphysema in Clichy from 1988 to 1997 were reviewed retrospectively. In particular, infectious complications and native lung hyperinflation were searched. Complications arising from the native lung are not unusual after SLT for subjects with emphysema, and it was concluded they are not responsible for a substantial mortality. PMID- 10703701 TI - Life supporting function for over one month of a transgenic porcine heart in a baboon. AB - BACKGROUND: Inhibition of hyperacute rejection (HAR) and sustained graft survival have been demonstrated in a pig-to-primate model of heterotopic cardiac xenotransplantation using pigs transgenic for human Decay Accelerating Factor (hDAF). Building on this work, an orthotopic model has been developed. This case records 39-day cardiac xenograft function in a life-supporting capacity with clinically applicable immunosuppression. METHODS: Using a heart from an hDAF transgenic pig, an orthotopic cardiac transplant was performed on an adult baboon. The immunosuppressive regimen consisted of induction with a short course of cyclophosphamide, followed by maintenance therapy with cyclosporine A, mycophenolate mofetil and a tapering course of corticosteroids. Post-operative monitoring included daily anti-pig hemolytic antibody titer surveillance and endomyocardial biopsy. RESULTS: The animal survived 39 days and was active and energetic throughout its postoperative course, remaining free of signs of cardiopulmonary failure. Endomyocardial biopsy performed on post-operative Day 36 revealed only patches of sub-endocardial fibrosis with no signs of active rejection. The baboon succumbed to an acute cardiopulmonary decompensation immediately following administration of medication via oral gavage. Post-mortem histopathology demonstrated well-preserved myocardial architecture with small foci of mild humoral rejection. CONCLUSIONS: This case documents the longest survival recorded to date of a discordant orthotopic cardiac xenograft and illustrates that the hDAF transgene combined with a clinically acceptable maintenance immunosuppressive regimen enables sustained, life-supporting function of porcine cardiac xenografts in non-human primates. The inhibition of hyperacute rejection and the subsequent control of humoral and cellular rejection for over 1 month demonstrated in this experiment represent significant progress in the development of a viable strategy for clinical xenotransplantation. PMID- 10703702 TI - Synergism between porcine reproductive and respiratory syndrome virus (PRRSV) and Salmonella choleraesuis in swine. AB - Porcine reproductive and respiratory syndrome virus (PRRSV) and Salmonella choleraesuis are two leading causes of economic loss in the swine industry. While respiratory disease is common in both S. choleraesuis and PRRSV infections, the factors that contribute to its development remain largely undefined. We investigated the interaction of PRRSV, S. choleraesuis, and stress in 5-week-old swine. All combinations of three factors (inoculation with S. choleraesuis on Day 0, PRRSV on Day 3, and treatment with dexamethasone on Days 3-7) were used to produce eight treatment groups in two independent trials. Fecal samples, tonsil and nasal swabs, serum samples and postmortem tissues were collected for bacteriologic and virologic examinations. No clinical signs were observed in pigs inoculated with only PRRSV or only S. choleraesuis. In contrast, pigs which were dually infected with S. choleraesuis and PRRSV exhibited unthriftiness, rough hair coats, dyspnea, and diarrhea. The pigs which received all three treatment factors were the most severely affected and 43% (three of seven) of the animals in this group died. Individuals in this group shed significantly higher quantities of S. choleraesuis in feces and had significantly higher serum PRRSV titers compared to other treatments (p < or = 0.05). In addition, S. choleraesuis and PRRSV were shed longer and by more pigs in this group than other groups and S. choleraesuis was recovered from more tissues in this group on Day 21 post inoculation. These results suggested that PRRSV, S. choleraesuis, and dexamethasone acted synergistically to produce a syndrome similar to that observed in the field. PMID- 10703704 TI - Expression and antigenic characterization of recombinant Mycoplasma agalactiae P48 major surface protein. AB - The gene encoding the P48 major surface lipoprotein of M. agalactiae has been recently characterised. Since its product plays an important role in the immune response of infected animals, in this study we analysed a recombinant P48 expressed in E. coli. Multiple point mutations were introduced by site directed mutagenesis in order to convert four tryptophan TGA codons, which are a typical feature of the mycoplasma genetic code, into the standard TGG. The mutated p48 gene was subcloned into pGex-2T and expressed in fusion with glutathione-S transferase. Following purification steps, P48 was eluted from carrier protein by thrombin digestion and used in Western blot and indirect ELISA using well characterised sheep sera. Results demonstrate that specific antibodies against P48 are detected 3 weeks after onset of clinical disease and the recombinant P48 is a diagnostically relevant marker of M. agalactiae infection. PMID- 10703703 TI - Antibiotic susceptibility of canine Bordetella bronchiseptica isolates. AB - The antimicrobial sensitivities of 78 recent (1995-1998) canine isolates of Bordetella bronchiseptica from 13 separate sources were determined. Minimum inhibitory concentrations were assessed using the E-test method or by agar dilution. All 78 isolates were sensitive to tetracycline, doxycycline, enrofloxacin, and amoxycillin/clavulanic acid; the majority were sensitive to ampicillin (63/78; 81%), trimethoprim (57/78; 73%), and sulphadiazine (63/78; 81%). Plasmids were detected in 14 out of the 24 isolates tested. There was no correlation between the presence of plasmids and antibiotic resistance, but there was some correlation between the presence of plasmids and the origin of the isolates. Three sizes of plasmid were found: 20, 14, and 5.5 kb. Eight of the isolates contained all three plasmids, the remainder one or two, Thirteen isolates demonstrated beta-haemolysis, of which six produced a soluble haemolysin. Except for one isolate, haemolysin production correlated with plasmid carriage. Pulsed-field gel electrophoresis showed that all except one isolate could be grouped in the same genotype. Within this genotype isolates could be divided into three subtypes, generally corresponding to their place of origin. PMID- 10703705 TI - Virulence factors and genetic relatedness of Escherichia coli strains isolated from pigs with post-weaning diarrhea. AB - Forty-six Escherichia coli strains isolated from post-weaning diarrhea of pigs were analysed for their phenotypic and genotypic properties. The isolates were of serogroups O138, O139, and O141 and most of them possessed hemolytic activities. PCR analysis showed that 34 of the isolates harboured the genes for shiga toxin 2e and 32 strains possessed the genes for heat-stable enterotoxins I and II. Ten strains had the fedA gene of F18 fimbriae. The genetic relationships among all isolates were tested by random amplified polymorphic DNA (RAPD) and enterobacterial repetitive intergenic consensus (ERIC) PCR analyses. Using the RAPD test with two different primers, six fingerprints were distinguished whereas the ERIC analysis revealed only three DNA patterns. Some strains possessing identical phenotypic and genotypic virulence determinants exhibited distinct RAPD profiles and some isolates with different pathogenic markers showed the same RAPD and ERIC pictures. Thus, RAPD, and to a less extent ERIC techniques, revealed intra- and interserogroup genotypic variations among the E. coli strains analyzed. PMID- 10703706 TI - The F4 fimbrial antigen of Escherichia coli and its receptors. AB - F4 or K88 fimbriae are long filamentous polymeric surface proteins of enterotoxigenic Escherichia coli (ETEC), consisting of so-called major (FaeG) and minor (FaeF, FaeH, FaeC, and probably FaeI) subunits. Several serotypes of F4 have been described, namely F4ab, F4ac, and F4ad. The F4 fimbriae allow the microorganisms to adhere to F4-specific receptors present on brush borders of villous enterocytes and consequently to colonize the small intestine. Such ETEC infections are responsible for diarrhea and mortality in neonatal and recently weaned pigs. In this review emphasis is put on the morphology, genetic configuration, and biosynthesis of F4 fimbriae. Furthermore, the localization of the different a, b, c, and d epitopes, and the localization of the receptor binding site on the FaeG major subunit of F4 get ample attention. Subsequently, the F4-specific receptors are discussed. When the three variants of F4 (F4ab, F4ac, and F4ad) are considered, six porcine phenotypes can be distinguished with regard to the brush border adhesiveness: phenotype A binds all three variants, phenotype B binds F4ab and F4ac, phenotype C binds F4ab and F4ad, phenotype D binds F4ad, phenotype E binds none of the variants, and phenotype F binds F4ab. The following receptor model is described: receptor bcd is found in phenotype A pigs, receptor bc is found in phenotype A and B pigs, receptor d is found in phenotype C and D pigs, and receptor b is found in phenotype F pigs. Furthermore, the characterization of the different receptors is described in which the bcd receptor is proposed as collection of glycoproteins with molecular masses ranging from 45 to 70 kDa, the bc receptor as two glycoproteins with molecular masses of 210 an 240 kDa, respectively, the b receptor as a glycoprotein of 74 kDa, and the d receptor as a glycosphingolipid with unknown molecular mass. Finally, the importance of F4 fimbriae and their receptors in the study of mucosal immunity in pigs is discussed. PMID- 10703707 TI - Immunological characterization of the major outer membrane protein of Haemophilus somnus. AB - Antigens and molecular mass diversity of the Haemophilus somnus major outer membrane protein (MOMP) were investigated. The molecular mass of the MOMP of 53 strains of H. somnus varied from 43 to 33 kDa and four MOMP MAb reactivity patterns were detected in immunoblot analysis and immunodot assay. The molecular mass and MAb reactivity data were used for preliminary grouping of H. somnus strains. Disease strains fell into groups 1 and 3, including two of three Group 3 subgroups, whereas strains from asymptomatic carriers were found in all the four groups and three subgroups. Immunoblot analysis with convalescent phase serum showed strain specific reactivity with MOMPs from three isolates used to reproduce disease in cattle. The reaction with the MOMP was only detectable at dilutions of 1:100 or less, whereas the same convalescent sera showed strong reactivity at dilutions of 1:1000 (or more) with other H. somnus antigens. The data suggest that the bovine immune response to the MOMP during infection is weak and is directed to antigenically variable determinants in a strain-specific manner. This may be important in evaluating the role of the antibody response to MOMPs in protective immunity. PMID- 10703708 TI - Active oral immunization of suckling piglets to prevent colonization after weaning by enterotoxigenic Escherichia coli with fimbriae F18. AB - Immunoprophylaxis of porcine oedema disease and post-weaning diarrhoea caused by strains of Escherichia coli expressing fimbriae F18 is an unsolved problem. The study was designed to examine whether vaccination with a live F18ac vaccine of unweaned pigs born to sows with F18ac antibody in the colostrum requires preformed fimbriae in the vaccine, and whether protection against the heterologous fimbrial variant F18ab is induced as well. Genetically susceptible pigs were vaccinated orally on three consecutive days, beginning 10 days before weaning with 10(11) CFU of an F18ac culture. Challenge with a dose of 10(7) CFU of E. coli F18 on three consecutive days was initiated 9 or 11 days after weaning. Eighteen pigs given the fimbriated F18ac vaccine and challenged with a strain of the homologous fimbrial variant were protected against colonization; mean faecal viable counts of the challenge strain were >3 log10 lower than those from the 17 non-vaccinated control pigs. The vaccinated pigs developed a significant rise of F18ac IgA serum antibodies. The 23 pigs which had received the non-fimbriated vaccine showed no significant protection and exhibited much lower serum F18ac IgA ELISA reactivities. Eighteen pigs vaccinated with the fimbriated F18ac and challenged with an F18ab strain had faecal viable counts nearly as high as those from 16 non-vaccinated control pigs. It is concluded that only oral vaccines having preformed fimbriae induce protection limited to the homologous fimbrial variant. PMID- 10703709 TI - A tissue culture system to study respiratory ciliary epithelial adherence of selected swine mycoplasmas. AB - An in vitro culture system for swine tracheal epithelial cells was developed to study the adherence of swine mycoplasmas. Swine tracheal epithelial cells were isolated by enzymatic digestion and cultured on microporous membranes. Growth medium was placed under the membrane support to create air-liquid interface feeding resulting in the cells growing cilia and microvilli on the apical surface. Two strains of Mycoplasma hyopneumoniae (pathogenic strain 91-3 and non pathogenic type strain J) and two strains of Mycoplasma flocculare (type strain Ms42 and field isolate 7160T) were used in this study. The morphology of the cultured tracheal cells was evaluated by transmission electron microscopy. Adherence of M. hyopneumoniae and M. flocculare and damage to the cilia were demonstrated using scanning electron microscopy. The pathogenic M. hyopneumoniae strain 91-3 adhered to cilia inducing obvious damage. The non-pathogenic M. hyopneumoniae strain J did not adhere to mature cilia. Both M. flocculare strains Ms42 and 7160T adhered to mature and budding cilia. No obvious ciliary damage was observed with strain Ms42. Minimal damage consisting of a slight tangling of the cilia occurred after adherence by strain 7160T. This model will enable us to further study the role of adherence of mycoplasmas on the pathogenesis of swine pneumonia. PMID- 10703710 TI - Antigenic differences in the H proteins of canine distemper viruses. AB - Antigenic properties between new Japanese field isolates and vaccine strains of canine distemper virus (CDV) have been compared using four monoclonal antibodies (MAbs) (JD-5, JD-7, JD-11 and d-7) against the hemagglutinin (H) proteins of CDV. JD-5, JD-7 and JD-11 are newly established antibodies. Three MAbs, namely d-7, JD 5 and JD-11, reacted similarly to all the CDV strains examined. However, JD-7 reacted much more strongly with the vaccine strains and an old field isolate than the recent field isolates in immunofluorescence, radio immunoprecipitation and virus neutralization assays. These results indicate that an antigenic region in the H protein, concerned with neutralization and recognized by JD-7, has been altered in the recent field isolates. PMID- 10703711 TI - Improvement of the identification of staphylococci isolated from bovine mammary infections using molecular methods. AB - Fifty-six Staphylococcus strains isolated from cases of bovine mammary infections were identified by using phenotypic and genotypic methods. Twenty-eight strains (50%) were identified at the species level according to their phenotypic characteristics, whereas the remaining 28 strains presented atypical or unreliable profiles. A combination of phenotypic and genotypic methods allowed the 56 strains studied to be classified. Internal transcribed spacer-polymerase chain reaction (ITS-PCR) based on the polymorphism of the 16S-23S rDNA spacer region appeared as a rapid and reliable method for the classification of bovine staphylococcal isolates at the species and subspecies levels. PMID- 10703712 TI - Effect of 7-methoxytacrine and L-carnitine on the activity of choline acetyltransferase. AB - Changes of choline acetyltransferase (ChAT) activity in the hippocampus and the basal ganglia were studied in rats treated i.p. with L-carnitine (CRT) and 7 methoxytacrine (7-MEOTA) (i.m.) separately or 3-days treated with L-carnitine and then with one administration of 7-MEOTA. Both compounds increased ChAT activity when administered separately. 3-day treatment of CRT followed by administration of 7-MEOTA normalized ChAT activity. PMID- 10703713 TI - Reactive oxygen species as mediators of tissue protection and injury. AB - Extensive research efforts during the last three decades resulted in a large body of experimental evidence that suggests an important role of the disbalance between generation and elimination of the oxygen and xenobiotic derived free radicals in physiological and pathological processes. Reactive oxygen species (ROS) are generated in many metabolic pathways, and are entering the organisms from exogenous sources, dominantly via airways and gut. ROS induced injuries, e.g. thermal, chemical, radiation, ischaemia/reperfusion, inflammation, hyperoxia, etc., result in diseases like atherosclerosis, ulcerative colitis, autoimmune diseases, asthma, etc. The current paper is designed to provide an overview of the effects ROS may exert in various tissues. Because of the effective defense systems, the tolerance of viable human cells to ROS is relatively high. The oxidant stress induced dysfunction of various systems, such as the gut, airways, nervous, cardiovascular system, etc., involve both direct and indirect mechanisms. Understanding of these molecular mechanisms is essential for a rational antioxidant therapy. PMID- 10703714 TI - Role of reactive oxygen and nitrogen species in etiopathogenesis of rheumatoid arthritis. AB - Rheumatoid arthritis (RA) is a chronic disease affecting up to 3% of the population in most countries. The causes of RA have not been completely elucidated. This paper aims to review the role of reactive oxygen and nitrogen species in the etiopathogenesis of RA. Reactive oxygen species (ROS), such as superoxide radical, hydrogen peroxide, hydroxyl radical and hypochlorous acid, as well as reactive nitrogen species (RNS), such as nitric oxide and peroxynitrite, contribute significantly to tissue injury in RA. Several mechanisms are involved in the generation and action of ROS and RNS. Superoxide radical, hydrogen peroxide and nitric oxide do not directly damage the majority of biological molecules. They are however converted into the highly reactive hydroxyl radical, which reacts with almost all molecules in living cells. The resulting chronic inflammation process can be reduced with antioxidant therapy. To date, scavenging, preventive, and enzyme antioxidants are available. The most important mode is scavenging of the hydroxyl radical and of hypochlorous acid. Another important way is to inhibit production of RNS and ROS by neutrophils, monocytes, and macrophages. The control of inflammation in arthritic patients by natural as well as synthetic antioxidants could become a relevant component of antirheumatic prevention and therapy. PMID- 10703715 TI - Neuro-immuno-teratogenicity of drugs used in neonatal pharmacotherapy in relation to the ontogenic stage at the time of their administration. AB - The risk of functional teratogenicity of two drugs used in neonatal pharmacotherapy was studied: indomethacin (INDO) and dexamethasone (DEX). Model experiments were carried out in Wistar strain rats, breed Konarovice, which received single subcutaneous drug injection (INDO 2 mg/kg, DEX 1 mg/kg) on postnatal day 4 (PD:4; model of human fetus/preterm newborn of 6-7-month gestational age) or on postnatal day 9 (PD:9; model of full-term human neonate). The rats were followed up during development (body weight, maturation) till late adulthood (age 6-8 months) using tests of cognition, immune reactivity and biochemical brain analysis. The results evaluated by comparing treated and control litter-mates indicated that the functional teratogenic risk was significantly higher in DEX than in INDO. DEX-rats revealed disorganization of developmental processes: retardation of body growth, but acceleration of sensory development (pinna and eye opening), retarded male sexual maturation. Adult DEX rats (age 6 months) of both series (PD:4, PD:9) had deficit of short-term memory (social recognition test). Disturbances of immune reactivity (decrease of humoral and rise of cell-mediated immune response) appeared both in adult INDO and DEX rats (age 7 months), but only in the PD:9 series i.e. when the drugs were administered at a higher stage of the ontogenic development simulating neonatal period in humans. This finding may be warning from the clinical point of view for the neonatological practice. PMID- 10703716 TI - The influence of alpha-lipoic acid on the toxicity of cadmium. AB - Alpha-lipoic acid (alpha-LA) is an important antioxidant drug with chelating properties. In experiments performed in male mice (CD-1, Charles River) the effects of cadmium on lipid peroxidation (LP), GSH level, the activity of catalase and glutathione peroxidase (GSH-Px) in liver homogenates were studied. Mice were injected with CdCl2 x 2.5 H2O at a dose of 40 micromol x kg(-1) s.c. Alpha-LA was administered simultaneously i.p. at the dose corresponding to alpha LA-to-Cd molar ratio of 5:1. The experiments were completed at 24 h. Cadmium increased LP to 200.7% of controls. This effect was prevented by alpha-LA treatment (p < or = 0.05). GSH level was decreased to 81.7% of controls and it was not affected by alpha-LA. GSH-Px activity diminished by Cd administration was corrected by alpha-LA (p < 0.001). Catalase activity decreased by Cd remained unaffected. The administration of alpha-LA alone enhanced LP and the activity of catalase. As estimated by AAS, Cd content in the liver, the kidneys, the brain and the testes remained unaffected by alpha-LA treatment. In the acute toxicity experiment, the mortality associated with cadmium was decreased by alpha-LA administration. The results suggest that the toxicity of Cd was decreased mainly by the antioxidant activity of alpha-LA rather than by cadmium removal from tissues. PMID- 10703717 TI - The role of mitochondria in apoptosis induced in vitro. AB - Cell death remains the focus of in vitro toxicology. Xenobiotics are capable of bringing about two types of cell death: apoptosis and necrosis. From our previous study we know that cells treated with xenobiotics showed very dynamic changes in their morphology, particularly vigorous movement of the plasma membrane. Such changes probably depend on adequate energy supply. This observation stands in contradiction with published data showing that generation of ATP in mitochondria is altered very early in apoptosis. In this study we analysed the relationship between mitochondrial activity and cell death induced by Etoposide, a selective inhibitor of topoisomerase II, treatment (10 microg/ml). As a model system we used stabilised cell line Hep2. Several markers of apoptosis, including typical cell morphology and DNA ladder formation were measured. The dynamics of morphological changes was recorded by the time-lapse videomicroscopy. We measured mitochondrial membrane potential with a specific fluorochrome DASPMI, quantification was done by microfluorometric assessment. Our data show that mitochondrial activity was maintained during the first 6 hours after the treatment with Etoposide, at the same time substantial changes in cell morphology as well as typical DNA fragmentation were observed. PMID- 10703718 TI - Antioxidant stobadine and neurobehavioural development of the rat offspring. AB - Stobadine (STO) is a potential neuro- and cardioprotective drug with high antioxidative properties. The presented study investigated the effects of oral STO administration (5, 15 and 50 mg/kg/d) during pregnancy and lactation to dams on neurobehavioural development of their offspring (body growth and maturation, sensory functions, neuromotor and reflex development, levels of activity and emotional reactivity, memory and learning processes). The results of our experiments showed that long-term administration of STO had no adverse effects on the course of pregnancy and lactation in dams and on the neurobehavioural development of offspring. PMID- 10703719 TI - Ecotoxicology of metals related to freshwater benthos. AB - The toxicity of Cu, Mn, Mo, Ni, V, As, Pb, Cr, Hg and Sn on the behaviour and survival of benthic worms Tubifex tubifex was studied. All tested metals were dissolved and determined in tap water under standardized conditions. The adverse effects of the metals were evaluated as acute toxicological effects upon exposure expressed as LC50 value with 95% confidence interval. On the basis of the LC50 values, the toxicity of the metals after an incubation for 96 h was ranked as follows: Cu(II)> Cu(I) > V > Hg > or = Mn > Ni > Cd > Cr > Mo > Pb > Sn(IV) = Sn(II) > As. From this sequence it is evident that copper was the most toxic metal ion. In addition, differences were found between Cu(II) and Cu(I) ions toxicity, the former being 2.5 times more toxic. In contrast, no differences could be confirmed between acute toxic effects of Sn(II) and Sn(IV). Arsenic showed the weakest toxicity of the tested metals. The LC50 value for As was 10,000 times higher than those for both copper ions. PMID- 10703720 TI - Streptozotocin-induced experimental diabetes in male Wistar rats. AB - The aim of the present work was to test the sensitivity of young male Wistar rats, Dobra Voda (Dv:WI) to the diabetogenic effect of streptozotocin (STZ) with regard to their health condition and mortality rates. Eight-week-old rats, weighing from 200 to 230 g, were randomised into five groups of eight animals. Streptozotocin was administered by i.v. injection in doses of 40, 50, 60 and 70 mg/kg body weight. The animals were kept on a standard diet with free access to water for 4 months. The highest STZ dose (70 mg/kg) was lethal to the animals, the doses of 50 and 60 mg/kg induced persistent hyperglycaemia with glucose levels above 20 mM. Body weights of STZ treated rats from all experimental groups were significantly lower than those of control animals. Considerable polyuria was observed in all STZ treated rats. About 40% of the STZ treated animals were found to develop overt cataract between days 90 and 100. At the end of the experiment, significant albuminuria was observed in the experimental groups administered 50 and 60 mg/kg STZ doses. We conclude that young male Wistar rats, Breeding Facility Dobra Voda (Dv:WI), Slovakia, treated by a single i.v. STZ dose of 50 or 60 mg/kg developed a persistent disease state characterised by severe hyperglycaemia with major clinical signs of diabetes mellitus. PMID- 10703721 TI - The use of cell culture systems for the assessment of general cellular toxicity and to detect the nature and location of free radical damage. AB - In the process of developing compounds to counteract the damaging effects of free radicals in biological systems it is important to determine the type and the intracellular location of specific toxic events. For that purpose cultured cells are used, because a properly designed cell culture system allows to asses specific damage at the cellular and subcellular level and can contribute to an evaluation of the biochemistry of free radical damage. A wide range of changes in cellular activities resulting from oxidative injury in vitro have been demonstrated. Data from many laboratories indicate that cell culture system coupled with appropriate analytical techniques can be used to explore cellular and biochemical details of damage induced by free radicals. The type of reactive oxygen species used to generate the radicals, the rate of radical production, and the location of action of the toxic species must be taken into account to understand the biochemistry of the system. An effort is made to analyse the considerable progress made in the development of appropriate in vitro models and end-points for use in testing and characterising the nature and location of free radical cytotoxicity. PMID- 10703723 TI - Isolation and cultivation of lung epithelial type 2 cells. The effect of cadmium. AB - The effects of cadmium exposure on rat lung type 2 cells were evaluated by morphological and biochemical examinations. The results showed dose dependent reduction in the marker enzyme (alkaline phosphatase), changes in cell membranes and in the antioxidant status. PMID- 10703722 TI - Use of single cell gel electrophoresis (comet assay) modifications for analysis of DNA damage. AB - Single cell gel electrophoresis (SCGE) or comet assay is a rapid and sensitive fluorescent microscopic method which allows measurement of DNA strand breaks in individual cells. Modifications of SCGE conditions permitted to detect different types of DNA damage. In order to characterize DNA damage induced by N-methyl-N' nitro-N-nitrosoguanidine (MNNG) and hydrogen peroxide (H2O2) in Chinese hamster V79 cells, two approaches were used: (1) two pH values of unwinding and electrophoresis solutions (pH > or = 13.0 and pH = 12.1) to specify the type of DNA lesions [the alkali-labile sites and true DNA single-strand breaks (ssb)] and (2) DNA glycosylases [endonuclease III (EndoIII) and formamidopyrimidine-DNA glycosylase (FaPy)] or DNA inhibitors [hydroxyurea (HU) + 1-(beta-D arabinofuranosyl)cytosine (AraC)] to characterize the types of DNA damage. Our results showed that the lesions induced by H2O2 represented mainly the true DNA ssb, while MNNG formed predominantly alkali-labile sites, which were converted to DNA ssb under strong alkaline conditions (pH > or = 13.0). The effects of DNA repair enzymes and DNA inhibitors were more significant under lower pH (pH = 12.1) of unwinding and electrophoresis solution. Both, DNA glycosylases and DNA inhibitors increased the level of DNA ssb. PMID- 10703724 TI - Experimental Goettingen minipig and beagle dog as two species used in bioequivalence studies for clinical pharmacology (5-aminosalicylic acid and atenolol as model drugs). AB - Due to proven similarities in biotransformation between man and minipig, minipig seems to be the experimental animal of choice for preclinical pharmacokinetic studies when an experiment with a drug exhibiting a great first pass bioelimination (like 5-aminosalicylic acid) is to be realised. On the other hand, both minipig and dog may be suitable species for a pharmacokinetic study with a drug characterized by a small extent of first pass biotransformation (like atenolol). PMID- 10703725 TI - Comparison of isoproterenol-induced changes in lysosomal enzyme activity in vivo and in vitro. AB - Isoproterenol was used as a drug which, when administered in high doses, is able to induce lysosomal enzyme activity changes in in vivo conditions. We correlated lysosomal enzyme activity in the absence and presence of isoproterenol, obtained in whole animals and in HeLa and HepG2 cells in tissue culture. In vivo experiments: male Wistar rats (270-300 g) were treated subcutaneously with isoproterenol in various doses. Effect of isoproterenol on lysosomal enzyme activity was assayed in the heart after differential centrifugation. In vitro experiments: Isoproterenol in concentrations 0.1-100 microg/ml was added to HeLa and HepG2 cells and the activity of lysosomal enzyme was measured in the cell homogenate. In the sedimentable and nonsedimentable fractions of the rat myocardium, the isoproterenol-induced changes in the activity of lysosomal enzyme were time-and dose-dependent. In HeLa cells, isoproterenol administration caused a dose-dependent increase of lysosomal enzyme activity, while in HepG2 cells the activity remained unchanged. Thus the isoproterenol-induced changes in lysosomal enzyme activity in the rat myocardium were comparable with the results found in vitro in HeLa cells. PMID- 10703726 TI - Genetic risk assessment of acid waste water containing heavy metals. AB - The mutagenic/cancerogenic potential of acid-mine water from the Slovak mining area Rudnany containing a high load of toxic metals was evaluated after its application to three model test organisms (bacteria Salmonella typhimurium, yeast Saccharomyces cerevisiae and plant Vicia sativa L.). The results obtained from the modified preincubation Ames assay proved that 1000-fold diluted waste water exhibited mutagenic effect in three (TA97, TA98, TA102) of four bacterial strains. In the test on yeast the toxicity and genotoxicity increased as a function of the concentration. At the highest concentration used (0.06%) the frequency of revertants increased 6 times and convertants increased 4.5 times above the control level. In the simultaneous phytotoxicity and clastogenicity assay, concentration dependent toxicity and statistically significant clastogenicity was proved. We can conclude that heavy metals might be responsible for the genotoxic/cancerogenic potential of the test water. However, we do not entirely exclude the possibility that its genotoxicity might be promoted by its high acidity. PMID- 10703728 TI - Toxicological evaluation of the new calcium antagonist VULM 993. AB - The tolerance of the new calcium antagonist VULM 993 was investigated in a series of toxicological studies. The following results were obtained: the maximum tolerated oral dose in acute toxicity was 10,000 mg/kg for mice and 6600 mg/kg for rats, for venous administration it was 26.1 mg/kg in mice and 32.2 mg/kg in rats. In subacute oral toxicity test in rats, VULM 993 showed no toxic effect up to 300 mg/kg/d. The drug was not teratogenic in rats (5, 50 or 250 mg/kg/d, p.o.). VULM 993 did not show any positive response in tests for genotoxicity in vitro. Transplacental study of VULM 993 in rabbits indicated active placental barrier function in the late stage of pregnancy. The toxicological profile of VULM 993 is characterised by a high tolerance in all relevant species of experimental animals, and no biologically significant mutagenic potential was recorded. PMID- 10703727 TI - Effect of chloridazone on the animal organism. AB - The acute toxic effect of the herbicide chloridazone and mitochondrial respiration were investigated and typical clinical signs of intoxication were described in rats (Wistar), pheasants (Phasianus colchicus) and sheep (Slovak Merino). The LD50 of chloridazone was calculated to be for rats 800 mg/kg bw (range 552 to 1160 mg/kg bw) and for pheasants 3684 mg/kg bw (range 1768 to 7677 mg/kg bw). According to WHO chloridazone is moderately toxic for rats and slightly toxic for pheasants. The LD50 for sheep is 161 mg/kg bw (range 76 to 340 mg/kg bw). Chloridazone thus presents an acute risk for ruminants, which is in coincidence with the WHO classification characterising it as a very toxic compound. The following clinical features of intoxication were observed after p.o. administration of chloridazone: apathy, dyspnoea, hyperventilation, hypersalivation (sheep - foam hypersalivation), paralysis, tonic-clonic convulsions and death in clonic convulsions. Very quick rigor mortis. Chloridazone interfered with mitochondrial respiration in the liver of rats yet its mode of action was different from that of succinate substrate or glutamate malate. Succinate dependent respiration was significantly decreased in both states (3 and 4) of respiration. Glutamate-malate respiration was not changed in state 4, though it significantly increased in state 3 after ADP administration. RCP (respiration control proportion) value was increased on using either of the substances. PMID- 10703729 TI - Protective effect of the antioxidant stobadine against cyclophosphamide and irradiation induced oxidative stress. AB - The antioxidant stobadine was tested for its efficiency against oxidative stress in model experiments with ICR nonpregnant mice exposed either to cyclophosphamide (80 mg/kg) or whole body 60Co (6.5 Gy) irradiation. In a teratological experiment, pregnant mice were exposed to cyclophosphamide (10 mg/kg) from day 11 to 17 of gestation. Toxicity was measured by determining the lysosomal enzymes acid phosphatase and N-acetyl-beta-D-glucosaminidase. Cyclophosphamide and irradiation caused a significant increase in acid phosphatase and N-acetyl-beta-D glucosaminidase activity in the spleen of nonpregnant mice. In the liver, lysosomal enzyme activities were unchanged and no changes in protein levels were recorded. In pregnant mice, acid phosphatase and N-acetyl-beta-D-glucosaminidase activities were increased in the spleen. An increase in foetal acid phosphatase liver activity was found. Pretreatment with stobadine prior to cyclophosphamide and irradiation significantly diminished the biochemical changes in both nonpregnant and pregnant mice. We conclude that stobadine is able to protect mice against cyclophosphamide- or irradiation-induced oxidative stress. PMID- 10703730 TI - Nonreceptor interactions in the pharmacology of blood platelets. AB - On the basis of values corresponding to concentrations exhibiting 50% inhibition of platelet aggregation induced with different stimuli and 50% inhibition of arachidonic acid liberation and thromboxane generation, we compared the antiplatelet effect of two cationic amphiphilic drugs--chloroquine and dithiaden. Compared to chloroquine, dithiaden was much more effective in inhibiting platelet aggregation, A-23187 induced arachidonic acid liberation and thromboxane generation. Chloroquine, on the other hand, was more effective in the inhibition of thrombin-induced arachidonic acid liberation. PMID- 10703731 TI - Adverse drug reactions to antibiotics and major antibiotic drug interactions. AB - Adverse drug reaction and drug interactions represent negative consequences of pharmacotherapy. A thorough knowledge of how the adverse reactions and drug interactions occur will aid in our ability to provide the best patient care possible. Adverse drug reactions and drug interactions are defined and mechanisms of their occurrence are discussed; ways to identify, assess, manage, and report these reactions are presented. The importance of postmarketing surveillance is highlighted. A thorough understanding of the mechanism of adverse drug reactions and drug interaction is essential for the health care practitioner. Improved patient care can be achieved by applying this knowledge to the individual patient. When drugs are administered to patients, many things can go wrong. The magnitude of this problem warrants spending sufficient resources to establish programs to identify, assess, and manage these reactions. Having effective reporting programs will help prevent many reactions and will provide earlier detection of adverse reactions to new drugs. PMID- 10703732 TI - Detection of apoptotic changes in HeLa cells after treatment with paracetamol and sodium fluoride. AB - Apoptosis is genetically programmed cell death, an irreversible process of cell senescence with characteristic features (cell shrinkage, chromatin condensation, DNA fragmentation, apoptotic bodies) different from necrosis. Several effective in vitro methods for qualitative and quantitative detection of apoptotic events have been developed. Chromatin degradation, reductions in cell volume and other apoptosis-associated changes in cell morphology and physiology can be quickly analysed by multiparametric flow cytometry (FC) using small numbers of intact cells. One further method used for morphological determination of apoptotic changes is the fluorescent microscopic technique (FM) based on labeling of cells with fluorochromes acridine orange and ethidium bromide. In our experiments FC was used for determination of DNA changes in HeLa cells based on staining of DNA by fluorochrome propidium iodide (PI). Among the tested chemicals (paracetamol and sodium fluoride) apoptotic process could only be detected in paracetamol treated cells. Apoptosis was induced mainly in cells treated with paracetamol (concentration range 4-5 mg/ml) for 8 h and following incubation for 18 h in fresh medium without paracetamol. The results obtained by the FM method correlated with the results obtained by FC. PMID- 10703733 TI - Detection of apoptosis in a heterogenous cell population using flow cytometry. AB - Apoptosis induced in human leukemic cells (promyelocytic human leukemic cells HL 60, multidrug-resistant subline HL-60/VCR) and human ovarian carcinoma cells (A2780 and multidrug-resistant subline A2780/ADR) in vitro was detected by flow cytometric analysis or DNA electrophoresis. The cytofluorometric techniques utilized, i. e. detection of phosphatidylserine exposed at the outer surface of the plasma membrane, identification of cells with "sub-G0" DNA content or increased light side scatter (cell internal structure) correlated with the electrophoretic determination of DNA fragmentation ("DNA ladder"). Detection of the 34 kDa mitochondrial protein recognized by the monoclonal antibody Apo2.7 yielded elevated percentages of apoptotic cells, suggesting that this technique detecting both early and late apoptosis in digitonin-fixed cells might not be restricted to the specific detection of programmed cell death. PMID- 10703734 TI - Streptozotocin diabetes-induced changes in aorta, peripheral nerves and stomach of Wistar rats. AB - In rats with diabetes induced by streptozotocin (STZ), we studied the reactivity of the aorta in response to vasoconstrictor and vasorelaxant agents, changes in conduction velocity in the sciatic nerve, and glutathion (GSH) content in the gastric mucosa as well as the occurrence of spontaneous gastric lesions. STZ induced diabetes was found to be accompanied by endothelial injury, exhibited by diminished endothelium-dependent relaxation and by increased noradrenaline- and H2O2-induced contraction. Conduction velocity in the nerves from STZ-treated animals was significantly lower compared to that in nerves from control animals. Moreover, gastric hyperaemia, occasional gastric lesions, and a significant depletion of GSH in the gastric mucosa were observed in STZ-treated rats. Our experiments confirmed the suitability of Wistar rats for the model of STZ-induced diabetes. PMID- 10703735 TI - Subchronic preexposure to carbon monoxide modifies heart response to a combined CO + isoproterenol challenge in rats. AB - The effect of repeated exposure to carbon monoxide (CO) on the response of middle age rats to an acute CO exposure combined with a low dose of a sympathomimetic agent was studied. A group of 12 rats (male albino, Wistar, age 9 months) without ECG abnormalities was divided into two subgroups matched for weight, heart rate and ECG: one subgroup was exposed to 500 ppm CO for 6 h/d, 5 d/w, for 6 weeks (peak COHb 31.5%, SD 3.5); the other one (controls) was exposed to fresh air. Two or three days after the last exposure both groups underwent combined challenge with 0.025 mg/kg isoproterenol s.c. and 90 minute exposure to CO in a concentration increasing from 500 to 1500 ppm; ECG was recorded continuously. The hearts were examined morphometrically and histologically. The CO-preexposed subgroup had, as compared to controls: 1) significantly higher blood hemoglobin (by 25%), erythrocyte count (by 28%) and volume (by 6%), and hematocrit (by 33%); 2) the same peak COHb; 3) lower basic heart rate and an earlier decrease after isoproterenol, 4) significantly smaller increase in ECG abnormalities and arrhythmias after isoproterenol and during CO exposure; 5) nonsignificantly higher heart weight indexes; 6) a nonsignificantly lower score of histological abnormalities. The global score of ECG pathology during CO exposure (abnormal pattern or arrhythmia) correlated best (multiple corr. coef. >0.9) with end exposure free (non CO bound) hemoglobin (negatively) and with mean heart rate during exposure (positively): the lower score in the preexposed subgroup was attributable primarily to the increased hemoglobin. Six-week intermittent CO exposure induced marked compensatory processes (hematological) but only a tendency to adaptational changes in the heart (by gross morphometry), and decreased the ECG response to CO+ isoproterenol challenge at the same COHb. PMID- 10703737 TI - Dual effect of pseudorabies virus growth factor (PRGF) displayed on actin cytoskeleton. AB - Pseudorabies virus growth factor (PRGF) was shown to possess transforming activity as well as transformation repressing activity in in vitro systems. In order to better understand these phenomena we studied actin cytoskeleton and its alterations induced by PRGF using normal human fibroblasts VH-10 and transformed cell line HeLa. For specific detection of filamentous actin cells were stained with phalloidin conjugated with fluorescein isothiocyanate (FITC)-phalloidin. PRGF was applied to VH-10 cells for various length of time from 10 min up to 48 h. The effect was very fast and changes in actin filament composition could be detected already after 10 min. In comparison to untreated cells the staining of treated cells was more diffuse and a number of actin microfilaments in individual stress fibers became reduced. After 30 min thick short actin bundles appeared in the perinuclear region. A 24-h exposure resulted in a large reduction of actin bundles. After additional 24 h a partial restoration of actin cytoskeleton in cells was observed. In transformed HeLa cells PRGF induced opposite process than in normal cells: the number of filamentous actin structures increased. We hypothesise that PRGF may act as a transcription-like factor and may initiate changes in gene expression which consequently result in actin cytoskeleton alterations. PMID- 10703736 TI - Teratological study of the antioxidant stobadine in rats. AB - The potential teratogenicity of the antioxidant stobadine (STO) was studied in Wistar rats. Daily oral doses of 5, 15 and 50 mg/kg STO were given from the 6th day of gestation up to weaning of pups--day 21 post partum. No significant differences between the STO treated groups and the control group were found in litter size, pre- and postimplantation losses and foetal body weight. External, skeletal and internal examinations of the foetuses revealed no evidence of teratogenesis. The offspring from the STO treated dams exhibited a high survival rate in their postnatal development. It can be concluded that STO had no adverse effects on the pre- and postnatal development of the offspring in rats. PMID- 10703739 TI - On public opinion, a scientist's mental state, and other trivialities. PMID- 10703738 TI - Developmental defects and chromosomal aberrations in spontaneous abortions and stillbirths. AB - The authors analysed 1488 cases of spontaneous abortions and stillbirths in Bratislava. They focused on the course of human embryogenesis and the chromosomal constitution. A high mean frequency rate of both developmental defects (14.4%) and chromosomal aberrations (33.6%) was revealed and both were found to be in close relation with the length of gestation. The most severe developmental defects occurred mostly in early stages of human embryogenesis, i.e. in the 1st trimester of gestation. PMID- 10703740 TI - Receptor-ligand interaction and molecular modelling. AB - A number of computational methods for the description of the ligand-receptor interaction that were developed in the past decade are reviewed in this paper. The central two sections introduce the methods that are already established as useful tools for the qualitative and quantitative description of ligand-receptor complexes, either when the detailed atomic structure of the receptor is known or not. The following section gives two examples of the application of the described methodology on two limiting cases. PMID- 10703741 TI - Temporal patterns recognized by a network of coordinated time delays and coincidence detectors. AB - A computational model of a neuronal network is described which performs a fundamental task of general perception: recognition of temporal patterns in continuous and uncued neuronal spike trains. The presented network is able to recognize each pattern element (100 ms interval composed of sets of 10, 20, 30 and 40 ms interspike intervals combined in linear order) as it arrives. Its operation is based upon biologically plausible filtering mechanisms and population neurodynamics. PMID- 10703742 TI - Temperature dependence of delayed fluorescence induction curve transients. AB - The temperature dependence of delayed millisecond fluorescence (DF) induction curve transients was investigated. The transients were obtained by keeping intact maize leaf segments in the dark for 30 to 300 s before illumination. The temperature dependence of DF induction revealed abrupt changes in activation energy of the recombination process. Those changes highly overcome the lowest energy barriers for certain types of chemical reactions, implying significant possible alterations in the DF mechanism itself. On the other hand, the particular transients responses expressed some specificities. Both the electrochemical gradient controlled changes, and the temperature induced changes contribute to it. PMID- 10703743 TI - Inhibition and stimulation of K+ transport across the frog erythrocyte membrane by furosemide, DIOA, DIDS and quinine. AB - Frog erythrocytes were incubated in iso- or hypotonic media containing 10 mmol/l Rb+ and 0.1 mmol/l ouabain and both Rb+ uptake and K+ loss were measured simultaneously. Rb+ uptake by frog red cells in iso- and hypotonic media was reduced by 30-60% in the presence of 0.01-0.1 mmol/l [(dihydroindenyl)oxy] alkanoic acid (DIOA) or 0.5-1.0 mmol/l furosemide. Furosemide inhibited K+ loss from frog erythrocytes incubated in hypotonic media but did not affect it in isotonic media. DIOA at a concentration of 0.05 mmol/l inhibited of K+ loss from frog erythrocytes in both iso- and hypotonic media. At the concentrations of 0.01 and 0.02 mmol/l DIOA significantly suppressed K+ loss in a K+-free chloride medium but not in a K+-free nitrate medium. The Cl(-)-dependent K+ loss was completely blocked at a concentration of 0.1 mmol/l DIOA and the concentration required for 50% inhibition of K-Cl cotransport was approximately 0.015 mmol/l. However, the inhibitory effect of DIOA on K-Cl cotransport was masked by an opposite stimulatory effect on K+ transport which was also observed in nitrate medium. Quinine in a concentration of 0.2-1.0 mmol/l was able to inhibit Rb+ uptake and K+ loss only in hypotonic media. In isotonic media, quinine produced a stimulation of Rb+ uptake and K+ loss. A three to five-fold activation of Rb+ uptake and K+ loss was consistently observed in frog erythrocytes treated with 0.05-0.2 mmol/l 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS). In contrast, another stilbene derivative 4-acetamido-4'-isothiocyanatostilbene-2,2' disulphonic acid (SITS) had no effect on K+ transport in the cells. Thus, of these drugs tested in the present study only DIOA at low concentrations may be considered as a selective blocker of the K-Cl cotransporter in the frog red blood cells. PMID- 10703744 TI - Relationship between hydrophobicity parameters and the strength and selectivity of phytotoxicity of sulfosuccinic acid esters. AB - The strength and selectivity of the phytotoxicity of 11 sulfosuccinic acid ester surfactants were determined on the leaves of Tradescantia bicolor, and the data were evaluated by multivariate mathematical-statistical methods. Spectral mapping technique combined with stepwise regression analysis indicated that both the strength and the selectivity of the effect depend significantly on the specific hydrophobic surface area of the anionic surfactants determined in the presence of ions. The significant relationship between this hydrophobicity parameter and phytotoxic activity suggests the involvement of apolar (hydrophobic) forces in the plant-surfactant interaction. It was assumed that the apolar alkyl chains of the surfactants may insert in the hydrophobic part of the phospholipid bilayers causing membrane disorder and malfunction. PMID- 10703745 TI - Effects of a new hypoglycemic agent A-4166 on glucose metabolism in rat adipocytes and muscle tissues. AB - The effects of the oral administration of a non-sulfonylurea hypoglycemic agent, the phenylalanine derivative A-4166, on serum insulin and glucose levels and glucose metabolism in isolated rat adipocytes and slices of muscle tissues were studied. An increase in serum insulin and a decrease in glucose levels were observed 30 minutes after A-4166 administration to rats fed basal or high fat diet. No changes in basal glucose transport in isolated fat cells were observed after the administration of A-4166. The effect of in vitro added insulin was, however, stronger in rats fed basal diet and treated with A-4166. An elevation of the membrane glucose transporter GLUT 4 was observed in rats treated with A-4166. An increase of basal lipogenesis, measured by incorporation of radiocarbon labeled glucose into lipids, was noted in adipocytes from rats fed high fat diet. The addition of insulin was followed by stimulation of lipogenesis in rats fed basal diet, however, this hormone had no effect in rats fed high fat diet. The administration of A-4166 did not affect the basal or insulin stimulated lipogenesis. Basal glucose oxidation in the diaphragm was not influenced by high fat diet or by A-4166 treatment. In the soleus muscle, basal glucose oxidation was decreased in rats fed high fat diet, and treatment with A-4166 increased the glucose oxidation up to values observed in the control basal diet fed rats. These results indicate that the administration of A-4166 can affect glucose metabolism in muscle tissue and the sensitivity of adipocytes to insulin. PMID- 10703746 TI - Folding-unfolding of immunoglobulin domains in titin: a simple two-state model. AB - The folding-unfolding reaction rate process in the giant protein titin is studied within a simple two-state model. The molecule is assumed to be stretched by an external force which modulates the potential barrier associated with the folded state. A two-state model for this process is assumed (i.e., the immunoglobulin domains are considered to be either folded or unfolded, with no intermediate states at all). Simple calculations yield a relation between the force and the pulling speed that agrees fairly well with data from experiments and Monte Carlo simulations performed recently. Moreover, in a regime involving ultrafast pulling, the results show that the detailed form of the potential barrier is irrelevant, a conclusion that agrees with the current theoretical work on molecular dynamics. PMID- 10703747 TI - Are intestinal tumours in Apc+/-mice a suitable model of colorectal carcinoma in man? AB - In snap frozen sections of the duodenum, jejunum, ileum, the right and left colon of APC+/-mice mucosubstances, activities of brush border glycosidases and proteases, immunoreactivity of sucrase and activities of some enzymes of pericellular proteolysis were studied. Multiple adenomas (tubular or tubulovillous) the numbers of which decreased in the aboral direction occurred in the small intestine. Two tubulovillous adenomas with dysplastic nuclei but with no invasion were found in the right colon. The morphological and histochemical findings resembled those of human colorectal tumours. Activities of brush border enzymes and sucrase immunoreactivity were decreased to various extent or were not present at all. The findings fluctuated even within the same section. Activities of enzymes of pericellular proteolysis were slightly increased in comparison with non affected mucosa. This model is suitable and deserves further studies. PMID- 10703748 TI - The arterial supply of the cubital joint in the Bactrian camel (Camelus bactrianus). AB - The arterial supply of the six cubital joints from bactrian camels was studied. There were 15 arterial branches supplying the joint. The branches that arose from the transverse cubital artery were the medial anterior, superior anterior, lateral anterior, middle anterior, inferior anterior, superior lateral, middle lateral, inferior lateral, superior posterior and superior lateral posterior cubital branches. The branches originating from the collateral ulnar artery were the medial, middle posterior, inferior posterior, medial posterior and inferior lateral posterior cubital branches. These arteries united with each other around the cubital joint. PMID- 10703749 TI - Effect of acute hyperglycaemia on the serum fructosamine and blood glycated haemoglobin concentrations in canine samples. AB - The effect was studied of an acute and non-persistent hyperglycaemia on the serum fructosamine and blood glycated haemoglobin concentrations in canine samples. Five dogs were given glucose solution intravenously and blood samples were taken from each dog before and at 5, 15, 30, 60 and 120 min and 24 h after the infusion. There was an intense hyperglycaemia 5 min after the injection was given, but no statistically significant differences in the serum fructosamine and glycated haemoglobin were observed. It was concluded that an acute and transient hyperglycaemia does not cause significant changes in the glycated haemoglobin and fructosamine concentrations in healthy dogs. PMID- 10703751 TI - A practical method for non-surgically inserting intra-arterial catheters in European wild boars (Sus scrofa). AB - In endocrinological studies, the usual methods for taking blood samples are impractical because of the need for frequent sampling, and catheterization is required. The catheterization methods described for domestic pigs are not suitable for use with European wild boars (Sus scrofa) under field conditions. We describe a method for inserting an intra-arterial catheter into the European wild boar. The animals were sedated with a combination of medetomidin (Domitor 1 mg/ml, Orion-Farmos, Finland) and zolazepam and tiletamine (Zoletil forte vet 50 mg/ml + 50 mg/ml, Virbac Laboratories, France). Four animals were catheterized and sampled at 2-h intervals for 48 h in March under winter conditions and five in June under summer conditions. In March, all the catheters remained in place for the entire sampling period. In June, one animal managed to displace the catheter from its artery, but it was recatheterized and the rest of the samples were collected without problems. This catheterization method is relatively easy to carry out and requires no special facilities. PMID- 10703750 TI - Overestimation of the predictive value of positives by the usual calculations of the specificity of diagnostic tests. AB - The specificity of a diagnostic test for a given disease, i.e. the percentage of true negatives, can be calculated from either the results from a group of healthy animals or possibly from a group which also contains diseased animals which are free of the particular disease for which the test has been performed. The specificity may be much lower in the latter case and the predictive value of positives thus greatly reduced. In the example of creatine kinase being used for the diagnosis of muscle diseases in dogs, the specificity at the thresholds of 105 and 150 U/L (upper limits of the 95% and 99.7% interquantiles) decreased from 0.98 and 1.0 to 0.66 and 0.78, respectively. PMID- 10703752 TI - Strategic control of gastrointestinal nematodes in dairy calves in Florestal, Minas Gerais, Brazil. AB - Following epidemiological studies of gastrointestinal helminthiasis in dairy cattle in Florestal County, Minas Gerais, 80 Swiss and crossbred Zebu x Holstein calves, 8-10-months old, were selected to test the efficacy of three treatment protocols using ivermectin for helminth control. The calves were treated in Brachiaria grass paddocks, naturally infected with Haemonchus, Cooperia, Oesophagostomum and Trichostrongylus species, and then divided into four groups of 20 animals each: group 1 was treated with 200 microg/kg body weight ivermectin in April (at the end of the rainy season) and October (beginning of the rainy season); group 2 was treated in April, August (middle of the dry season) and October; group 3 was treated in April, August, October and December (middle of the rainy season); and group 4 was left untreated as a control. The treatments effectively eliminated the worm burden only in groups 2 and 3 (p < 0.05), although the calves continued to excrete Cooperia eggs after each treatment with ivermectin. PMID- 10703753 TI - The prevalence and intensity of helminth and coccidial infections in dairy cattle in central Kenya. AB - A survey of gastrointestinal parasite infections of young (< 6 months old), immature (6-12 months old) and adult (> 12 months old) dairy cattle on 16 farms in Kiambu District, Kenya was conducted during a dry season (September 1991 to January 1992) and during a wet season (March to July 1992). The survey was based on monthly coproparasitological examination of cohorts and worm counts in tracer calves. The effects of age, sex, farm and season on the prevalence and intensity of helminth and coccidial infections were determined. Faecal egg and oocyst counts revealed that the overall prevalences were: strongyles (including trichostrongyles) (85.5%), liver flukes (Fasciola gigantica) (34.0%), coccidia (30.9%) and tapeworms (9.6%). Eight species of the protozoan Eimeria were identified, the most prevalent species being E. bovis and E. zuernii. The most prevalent nematode genera were Haemonchus, Cooperia, Oesophagostomum and Trichostrongylus. Season, farm and age of the animals had a significant (p<0.05) influence on the intensity of infection with strongyles, liver flukes and coccidia, whereas the sex of the animals had no significant (p>0.05) effect on the prevalence or intensity of infections. A higher intensity of infection with strongyles and coccidia was found in the wet season than in the dry season (p<0.05). The age-specific intensity was in the following order: for strongyles, immature animals of 6-12 months of age had the highest egg counts, followed by young calves and adults. Calves had significantly (p<0.05) higher oocyst counts than immatures or adults. Liver fluke egg counts did not differ significant (p>0.05) between immatures and adult cattle. PMID- 10703754 TI - Screening for Indian isolates of predacious fungi for use in biological control against nematode parasites of ruminants. AB - Four isolates of predacious fungi, two each of Arthrobotrys oligospora isolated from a sheep and a male crossbred calf and of Duddingtonia flagrans isolated from a sheep and a female buffalo in western India, were studied for their suitability as biocontrol agents against parasitic nematodes of ruminants, using growth assay, predatory activity, germination potential and ability to survive passing through the ruminants gut as criteria. The study showed that isolates of D. flagrans grew well in artificial media, had encouraging predatory activity, produced profuse chlamydospores that germinated easily at 25 degrees C and could survive passage through the ruminant gut. The ovine isolate of D. flagrans was superior in all respects to the isolate from buffalo and was the most promising candidate for biological control of nematode parasites of ruminants. PMID- 10703757 TI - Role of norepinephrine in depression. AB - This article reviews the role of norepinephrine (NE) and serotonin (5-HT) in depression and the therapeutic effects of antidepressant drugs from the perspective of human neurotransmitter depletion studies. The data reviewed suggest that both noradrenergic and serotonergic systems are involved in antidepressant action, but the specific impairment that underlies depression is unclear and is likely to vary among patients. Results from neurotransmitter depletion studies in depressed patients who have responded to treatment suggest that, while interactions between NE and 5-HT are likely, neither of these 2 neurotransmitter systems is the final common pathway for the therapeutic effect of antidepressant drugs. NE-selective antidepressant drugs appear to be primarily dependent on the availability of NE for their effects. Likewise, 5-HT-selective antidepressants appear to be primarily dependent on the availability of 5-HT for their effects. Antidepressants that cause effects on both noradrenergic and serotonergic systems-such as mirtazapine-may be dependent on the availability of both neurotransmitters for their effects. Neither 5-HT nor NE depletion induced clinical depression in healthy subjects or worsened depression in unmedicated symptomatic patients with major depression. This finding suggests that the cause of depression is more complex than just an alteration in the levels of 5-HT and/or NE. For some patients, depression may be more directly caused by dysfunction in brain areas or neuronal systems modulated by monoamine systems. We propose that antidepressant drugs may enhance neurotransmission in normal noradrenergic or serotonergic neurons and, through a time-dependent but as yet undiscovered process, restore function to brain areas modulated by monoamine neurons. Future research should focus on understanding the adaptive changes that follow enhancement of synaptic levels of monoamines in neuronal circuits of the frontal cortex, amygdala, and hippocampus. Research investigating the neurobiology of depression may be more informed if the focus is shifted to investigating areas of the brain modulated by monoamine systems rather than the monoamine systems themselves. PMID- 10703755 TI - Confirmation of the prophylactic value of paromomycin in a natural outbreak of caprine cryptosporidiosis. PMID- 10703758 TI - Noradrenergic approaches to antidepressant therapy. AB - A decade of remarkable research in neuroscience has given us a much more complete picture of how the central nervous system works and, in some instances, how the brain does not work when patients develop depression. Preclinical and clinical studies have shown that stimulation of the serotonergic system leads to noradrenergic effects and vice versa, confirming that the serotonin and norepinephrine systems are intimately connected in the central nervous system. Although medications that target the serotonergic neurotransmitter system have recently dominated antidepressant therapy, atypical antidepressants--with either mixed serotonergic and noradrenergic effects or exclusively noradrenergic effects -have been shown to be clinically efficacious medications. This increased understanding of the interrelationship between neurotransmitter systems has renewed interest in the role of neurotransmitters other than serotonin in the treatment of depression. With the introduction of reboxetine, a very selective norepinephrine reuptake inhibitor, researchers have had an opportunity to study the unique effects of norepinephrine in the etiology and treatment of depression. Ultimately, differences in neurotransmitter profiles may influence therapeutic potentials of antidepressants. For example, influencing norepinephrine may affect the expression of energy and interest, while influencing serotonin may affect impulse control and influencing dopamine may affect drive. Clinicians now have a range of antidepressants with variable neurotransmitter effects, different side effect profiles, and some interesting differences in functional utility in their armamentarium for treating depression. PMID- 10703759 TI - Treatment of severe depression. AB - A significant minority of depressed outpatients and a vast majority of those hospitalized for depression can be considered severely ill. Severe depressive states are quite heterogeneous and include the more classical subforms (e.g., melancholia or psychotic depression) as well as various comorbid presentations. This article reviews the classification, phenomenology, pathophysiology, and treatment of severe depression. PMID- 10703760 TI - New approaches to the treatment of refractory depression. AB - Although the majority of patients with depression respond well to their initial pharmacologic treatment, as many as 30% to 45% fail to achieve an adequate response. In addition to the more traditional lithium and thyroid hormone augmentation strategies, a number of new pharmacotherapeutic approaches are currently being used to help manage refractory depression, including the addition of another agent or a switch to another antidepressant. Augmentation and switching strategies are often selected in order to obtain a different neurochemical effect (e.g., adding a relatively noradrenergic agent to a relatively serotonergic antidepressant). In particular, several studies have suggested that depressed patients refractory to treatment with selective serotonin reuptake inhibitors (SSRIs) may show a good response to newer agents that have a pharmacologic profile distinct from the SSRIs. Furthermore, preliminary studies have shown that the addition of SSRIs to either noradrenergic drugs such as the tricyclic antidepressants (TCAs) or dopaminergic agents may be efficacious, even though concerns about drug-drug interactions and tricyclic cardiac toxicity have limited the use of TCA-SSRI combinations. The introduction of reboxetine, a relatively selective norepinephrine reuptake inhibitor, may increase the use of the latter therapeutic approach because of its improved safety profile compared with the TCAs. The review of treatment options for refractory depression that follows will outline the advantages, disadvantages, and level of support for a number of new treatment strategies. PMID- 10703761 TI - Social functioning and the treatment of depression. AB - For economic and scientific reasons, there has been a recent increase in the use of social functioning as an outcome measure in clinical trials of psychotropic drugs. The new antidepressants are more expensive than the older agents, and improvement in social functioning, e.g., return to work, may justify their use. New assessments (e.g., vitality, motivation, and performance) that go beyond symptom reduction may also capture a broader spectrum of outcomes for the newer drugs. This article presents the historical background and rationale for interest in social functioning as an outcome of treatment with psychotropic medications, presents recent examples of measures of social functioning from clinical trials of new antidepressants, discusses several of the methods for assessing social functioning, and suggests how these assessments can be used in clinical practice. PMID- 10703762 TI - A mother's prayer. PMID- 10703763 TI - AIDS researchers explore new drug options. PMID- 10703764 TI - Abuse of prescription drugs: is a patient ailing or addicted? PMID- 10703765 TI - From the Food and Drug Administration. PMID- 10703766 TI - From the Centers for Disease Control and Prevention. Outbreaks of Salmonella serotype Enteritidis infection associated with eating raw or undercooked shell eggs--United States, 1996-1998. PMID- 10703767 TI - From the Centers for Disease Control and Prevention. Tobacco use among middle and high school students--United States, 1999. PMID- 10703768 TI - Violence prevention and concealed weapons laws. PMID- 10703769 TI - Residual HIV-1 RNA after highly active antiretroviral therapy. PMID- 10703770 TI - Safety of antidepressant medications during pregnancy. PMID- 10703771 TI - Evaluating clinical studies of drug efficacy. PMID- 10703772 TI - Health consequences of exemptions from immunization laws. PMID- 10703773 TI - Health consequences of exemptions from immunization laws. PMID- 10703774 TI - Association between thymoma and second neoplasms. PMID- 10703775 TI - Rapid heroin detoxification under general anesthesia. PMID- 10703776 TI - Intravenous tissue-type plasminogen activator for treatment of acute stroke: the Standard Treatment with Alteplase to Reverse Stroke (STARS) study. AB - CONTEXT: Tissue-type plasminogen activator (tPA) is the only therapy for acute ischemic stroke approved by the Food and Drug Administration. OBJECTIVE: To assess the safety profile and to document clinical outcomes and adverse events in patients treated with intravenous tPA for acute stroke in clinical practice. DESIGN AND SETTING: Prospective, multicenter study of consecutive patients enrolled between February 1997 and December 1998 at 57 medical centers in the United States (24 academic and 33 community). INTERVENTION: Intravenous tPA (recombinant alteplase). PATIENTS: Three hundred eighty-nine patients with a mean age of 69 years (range, 28-100 years); 55% were men. MAIN OUTCOME MEASURES: Time intervals between stroke symptom onset, hospital arrival, and treatment with tPA; pretreatment computed tomographic scan results, intracerebral hemorrhage, and major systemic bleeding. The modified Rankin Scale score was used to assess clinical outcomes at 30 days. RESULTS: Median time from stroke onset to treatment was 2 hours 44 minutes, and the median baseline National Institutes of Health Stroke Scale score was 13. The 30-day mortality rate was 13%. At 30 days after treatment, 35% of patients had very favorable outcomes (modified Rankin score, 0 1) and 43% were functionally independent (modified Rankin score, 0-2). Thirteen patients (3.3%) experienced symptomatic intracerebral hemorrhage, including 7 who died. Twenty-eight patients (8.2%) had asymptomatic intracerebral hemorrhage within 3 days of treatment with tPA. Protocol violations were reported for 127 patients (32.6%), and included treatment with tPA more than 3 hours after symptom onset in 13.4%, treatment with anticoagulants within 24 hours of tPA administration in 9.3%, and tPA administration despite systolic blood pressure exceeding 185 mm Hg in 6.7%. A multivariate analysis found predictors of favorable outcome to be a less severe baseline National Institutes of Health Stroke Scale score, absence of specific abnormalities (effacement or hypodensity of >33% of the middle cerebral artery territory or a hyperdense middle cerebral artery) on the baseline computed tomographic scan, an age of 85 years or younger, and a lower mean arterial pressure at baseline. CONCLUSIONS: This study, conducted at multiple institutions throughout the United States, suggests that favorable clinical outcomes and low rates of symptomatic intracerebral hemorrhage can be achieved using tPA for stroke treatment. PMID- 10703777 TI - Use of tissue-type plasminogen activator for acute ischemic stroke: the Cleveland area experience. AB - CONTEXT: Little is known regarding outcomes after intravenous tissue-type plasminogen activator (IV tPA) therapy for acute ischemic stroke outside a trial setting. OBJECTIVE: To assess the rate of IV tPA use, the incidence of symptomatic intracerebral hemorrhage (ICH), and in-hospital patient outcomes throughout a large urban community. DESIGN: Historical prospective cohort study conducted from July 1997 through June 1998. SETTING: Twenty-nine hospitals in the Cleveland, Ohio, metropolitan area. PATIENTS: A total of 3948 patients admitted to a study hospital with a primary diagnosis of ischemic stroke (International Classification of Diseases, Ninth Revision, Clinical Modification code 434 or 436). MAIN OUTCOME MEASURES: Rate of IV tPA use and occurrence of symptomatic ICH among patients treated with tPA; proportion of patients receiving tPA whose treatment deviated from national guidelines; in-hospital mortality among patients receiving tPA compared with that among ischemic stroke patients not receiving tPA and with mortality predicted by a model. RESULTS: Seventy patients (1.8%) admitted with ischemic stroke received IV tPA. Of those, 11 patients (15.7%; 95% confidence interval [CI], 8.1%-26.4%) had a symptomatic ICH (of which 6 were fatal) and 50% (95% CI, 37.8%-62.2%) had deviations from national treatment guidelines. In-hospital mortality was significantly higher among patients treated with tPA (15.7%) compared with patients not receiving tPA (5.1%, P<.001) and compared with the model's prediction (7.9%; P<.006). CONCLUSIONS: A small proportion of patients admitted with acute ischemic stroke in Cleveland received tPA; they experienced a high rate of ICH. Cleveland community experience with tPA for acute ischemic stroke may differ from that reported in clinical trials. PMID- 10703778 TI - Selective referral to high-volume hospitals: estimating potentially avoidable deaths. AB - CONTEXT: Evidence exists that high-volume hospitals (HVHs) have lower mortality rates than low-volume hospitals (LVHs) for certain conditions. However, few employers, health plans, or government programs have attempted to increase the number of patients referred to HVHs. OBJECTIVES: To determine the difference in hospital mortality between HVHs and LVHs for conditions for which good quality data exist and to estimate how many deaths potentially would be avoided in California by referral to HVHs. DESIGN, SETTING, AND PATIENTS: Literature in MEDLINE, Current Contents, and First-Search Social Abstracts databases from January 1, 1983, to December 31, 1998, was searched using the key words hospital, outcome, mortality, volume, risk, and quality. The highest-quality study assessing the mortality-volume relationship for each given condition was identified and used to calculate odds ratios (ORs) for in-hospital mortality for LVHs vs HVHs. These ORs were then applied to the 1997 California database of hospital discharges maintained by the California Office of Statewide Health Planning and Development to estimate potentially avoidable deaths. MAIN OUTCOME MEASURES: Deaths that potentially could be avoided if patients with conditions for which a mortality-volume relationship had been treated at an HVH vs LVH. RESULTS: The articles identified in the literature search were grouped by condition, and predetermined criteria were applied to choose the best article for each condition. Mortality was significantly lower at HVHs for elective abdominal aortic aneurysm repair, carotid endarterectomy, lower extremity arterial bypass surgery, coronary artery bypass surgery, coronary angioplasty, heart transplantation, pediatric cardiac surgery, pancreatic cancer surgery, esophageal cancer surgery, cerebral aneurysm surgery, and treatment of human immunodeficiency virus (HIV)/acquired immunodeficiency syndrome (AIDS). A total of 58,306 of 121,609 patients with these conditions were admitted to LVHs in California in 1997. After applying the calculated ORs to these patient populations, we estimated that 602 deaths (95% confidence interval, 304-830) at LVHs could be attributed to their low volume. Additional analyses were performed to take into account emergent admissions and distance traveled, but the impact of loss of continuity of care for some patients and reduction in the availability of specialists for patients remaining at LVHs could not be assessed. CONCLUSIONS: Initiatives to facilitate referral of patients to HVHs have the potential to reduce overall hospital mortality in California for the conditions identified. Additional study is needed to determine the extent to which selective referral is feasible and to examine the potential consequences of such initiatives. PMID- 10703779 TI - Effect of breastfeeding and formula feeding on transmission of HIV-1: a randomized clinical trial. AB - CONTEXT: Transmission of human immunodeficiency virus type 1 (HIV-1) is known to occur through breastfeeding, but the magnitude of risk has not been precisely defined. Whether breast milk HIV-1 transmission risk exceeds the potential risk of formula-associated diarrheal mortality in developing countries is unknown. OBJECTIVES: To determine the frequency of breast milk transmission of HIV-1 and to compare mortality rates and HIV-1-free survival in breastfed and formula-fed infants. DESIGN AND SETTING: Randomized clinical trial conducted from November 1992 to July 1998 in antenatal clinics in Nairobi, Kenya, with a median follow-up period of 24 months. PARTICIPANTS: Of 425 HIV-1-seropositive, antiretroviral naive pregnant women enrolled, 401 mother-infant pairs were included in the analysis of trial end points. INTERVENTIONS: Mother-infant pairs were randomized to breastfeeding (n = 212) vs formula feeding arms (n = 213). MAIN OUTCOME MEASURES: Infant HIV-1 infection and death during the first 2 years of life, compared between the 2 intervention groups. RESULTS: Compliance with the assigned feeding modality was 96% in the breastfeeding arm and 70% in the formula arm (P<.001). Median duration of breastfeeding was 17 months. Of the 401 infants included in the analysis, 94% were followed up to HIV-1 infection or mortality end points: 83% for the HIV-1 infection end point and 93% to the mortality end point. The cumulative probability of HIV-1 infection at 24 months was 36.7% (95% confidence interval [CI], 29.4%-44.0%) in the breastfeeding arm and 20.5% (95% CI, 14.0%-27.0%) in the formula arm (P = .001). The estimated rate of breast milk transmission was 16.2% (95% CI, 6.5%-25.9%). Forty-four percent of HIV-1 infection in the breastfeeding arm was attributable to breast milk. Most breast milk transmission occurred early, with 75% of the risk difference between the 2 arms occurring by 6 months, although transmission continued throughout the duration of exposure. The 2-year mortality rates in both arms were similar (breastfeeding arm, 24.4% [95% CI, 18.2%-30.7%] vs formula feeding arm, 20.0% [95% CI, 14.4%-25.6%]; P = .30). The rate of HIV-1-free survival at 2 years was significantly lower in the breastfeeding arm than in the formula feeding arm (58.0% vs 70.0%, respectively; P = .02). CONCLUSIONS: The frequency of breast milk transmission of HIV-1 was 16.2% in this randomized clinical trial, and the majority of infections occurred early during breastfeeding. The use of breast milk substitutes prevented 44% of infant infections and was associated with significantly improved HIV-1-free survival. PMID- 10703780 TI - Prevention of mother-to-child HIV transmission in resource-poor countries: translating research into policy and practice. AB - Each year, an estimated 590,000 infants acquire human immunodeficiency virus type 1 (HIV) infection from their mothers, mostly in developing countries that are unable to implement interventions now standard in the industrialized world. In resource-poor settings, the HIV pandemic has eroded hard-won gains in infant and child survival. Recent clinical trial results from international settings suggest that short-course antiretroviral regimens could significantly reduce perinatal HIV transmission worldwide if research findings could be translated into practice. This article reviews current knowledge of mother-to-child HIV transmission in developing countries, summarizes key findings from the trials, outlines future research requirements, and describes public health challenges of implementing perinatal HIV prevention interventions in resource-poor settings. Public health efforts must also emphasize primary prevention strategies to reduce incident HIV infections among adolescents and women of childbearing age. Successful implementation of available perinatal HIV interventions could substantially improve global child survival. PMID- 10703781 TI - Peripartum cardiomyopathy: National Heart, Lung, and Blood Institute and Office of Rare Diseases (National Institutes of Health) workshop recommendations and review. AB - OBJECTIVE: Peripartum cardiomyopathy (PPCM) is a rare life-threatening cardiomyopathy of unknown cause that occurs in the peripartum period in previously healthy women. In April 1997, the National Heart, Lung, and Blood Institute (NHLBI) and the Office of Rare Diseases of the National Institutes of Health (NIH) convened a Workshop on Peripartum Cardiomyopathy to foster a systematic review of information and to develop recommendations for research and education. PARTICIPANTS: Fourteen workshop participants were selected by NHLBI staff and represented cardiovascular medicine, obstetrics, immunology, and pathology. A representative subgroup of 8 participants and NHLBI staff formed the writing group for this article and updated the literature on which the conclusions were based. The workshop was an open meeting, consistent with NIH policy. EVIDENCE: Data presented at the workshop were augmented by a MEDLINE search for English-language articles published from 1966 to July 1999, using the terms peripartum cardiomyopathy, cardiomyopathy, and pregnancy. Articles on the epidemiology, pathogenesis, pathophysiology, diagnosis, treatment, and prognosis of PPCM were included. RECOMMENDATION PROCESS: After discussion of data presented, workshop participants agreed on a standardized definition of PPCM, a general clinical approach, and the need for a registry to provide an infrastructure for future research. CONCLUSIONS: Peripartum cardiomyopathy is a rare lethal disease about which little is known. Diagnosis is confined to a narrow period and requires echocardiographic evidence of left ventricular systolic dysfunction. Symptomatic patients should receive standard therapy for heart failure, managed by a multidisciplinary team. If subsequent pregnancies occur, they should be managed in collaboration with a high-risk perinatal center. Systematic data collection is required to answer important questions about incidence, treatment, and prognosis. PMID- 10703782 TI - Thrombolytic therapy for ischemic stroke: from clinical trials to clinical practice. PMID- 10703783 TI - High-risk surgery--follow the crowd. PMID- 10703784 TI - Gun carrying and homicide prevention. PMID- 10703785 TI - The application of preventive medicine to the control of violence. PMID- 10703786 TI - Helping medical students help survivors of domestic violence. PMID- 10703787 TI - Physician groups and the war in Kosovo: ethics, neutrality, and interventionism. PMID- 10703788 TI - Disintegrating health services and resurgent tuberculosis in post-Soviet Tajikistan: an example of structural violence. PMID- 10703789 TI - Youth violence prevention: the physician's role. PMID- 10703790 TI - Effect of a ban on carrying firearms on homicide rates in 2 Colombian cities. AB - CONTEXT: Homicide is a leading cause of death in Colombia, with much of the fatal interpersonal violence concentrated in the country's largest cities. Firearms are involved in as much as 80% of homicides in Colombia. OBJECTIVE: To evaluate the effect of an intermittent police-enforced ban on carrying firearms on the incidence of homicide in urban Colombia. DESIGN: Interrupted time-series study with multiple replications. SETTING: Cali, Colombia, during 1993 and 1994 and Bogota, Colombia, from 1995 through August 1997. PARTICIPANTS: The populations of Cali and Bogota. INTERVENTION: Carrying of firearms was banned on weekends after paydays, on holidays, and on election days. Enforcement included establishment of police checkpoints and searching of individuals during traffic stops and other routine law enforcement activity. MAIN OUTCOME MEASURE: Homicide rates during intervention days were compared with rates during similar days without the intervention; estimates were based on comparisons within the same month, day of week, and time of day. RESULTS: There were 4078 homicides in Cali during 1993 and 1994 (114.6 per 100,000 person-years). In Bogota, 9106 homicides occurred from 1995 through August 1997 (61 per 100,000 person-years). The incidence of homicide was lower during periods when the firearm-carrying ban was in effect compared with other periods (multivariate-adjusted rate ratio, 0.86 [95% confidence interval [CI], 0.76-0.97] for Cali, and 0.87 [95% CI, 0.77-0.98] for Bogota). CONCLUSION: An intermittent citywide ban on the carrying of firearms in 2 Colombian cities was associated with a reduction in homicide rates for both cities. PMID- 10703791 TI - JAMA Patient Page: newborn care. PMID- 10703792 TI - Medicine's response to Jorg Haider. PMID- 10703793 TI - Role of WHO-MONICA Project in unravelling of the cardiovascular puzzle. PMID- 10703794 TI - Towards post-genomic investigation of colorectal cancer. PMID- 10703795 TI - Limitations of tests for human chorionic gonadotropin. PMID- 10703796 TI - Impact of malaria on the brain and its prevention. PMID- 10703797 TI - Awareness of awareness during general anaesthesia. PMID- 10703798 TI - Jelly-beans, only a colourful distraction from gestational glucose-challenge tests. PMID- 10703799 TI - Estimation of contribution of changes in classic risk factors to trends in coronary-event rates across the WHO MONICA Project populations. AB - BACKGROUND: From the mid-1980s to mid-1990s, the WHO MONICA Project monitored coronary events and classic risk factors for coronary heart disease (CHD) in 38 populations from 21 countries. We assessed the extent to which changes in these risk factors explain the variation in the trends in coronary-event rates across the populations. METHODS: In men and women aged 35-64 years, non-fatal myocardial infarction and coronary deaths were registered continuously to assess trends in rates of coronary events. We carried out population surveys to estimate trends in risk factors. Trends in event rates were regressed on trends in risk score and in individual risk factors. FINDINGS: Smoking rates decreased in most male populations but trends were mixed in women; mean blood pressures and cholesterol concentrations decreased, bodymass index increased, and overall risk scores and coronary-event rates decreased. The model of trends in 10-year coronary-event rates against risk scores and single risk factors showed a poor fit, but this was improved with a 4-year time lag for coronary events. The explanatory power of the analyses was limited by imprecision of the estimates and homogeneity of trends in the study populations. INTERPRETATION: Changes in the classic risk factors seem to partly explain the variation in population trends in CHD. Residual variance is attributable to difficulties in measurement and analysis, including time lag, and to factors that were not included, such as medical interventions. The results support prevention policies based on the classic risk factors but suggest potential for prevention beyond these. PMID- 10703800 TI - Estimation of contribution of changes in coronary care to improving survival, event rates, and coronary heart disease mortality across the WHO MONICA Project populations. AB - BACKGROUND: The revolution in coronary care in the mid-1980s to mid-1990s corresponded with monitoring of coronary heart disease (CHD) in 31 populations of the WHO MONICA Project. We studied the impact of this revolution on coronary endpoints. METHODS: Case fatality, coronary-event rates, and CHD mortality were monitored in men and women aged 35-64 years in two separate 3-4-year periods. In each period, we recorded percentage use of eight treatments: coronary-artery reperfusion before, thrombolytics during, and beta-blockers, antiplatelet drugs, and angiotensin-converting-enzyme (ACE) inhibitors before and during non-fatal myocardial infarction. Values were averaged to produce treatment scores. We correlated changes across populations, and regressed changes in coronary endpoints on changes in treatment scores. FINDINGS: Treatment changes correlated positively with each other but inversely with change in coronary endpoints. By regression, for the common average treatment change of 20, case fatality fell by 19% (95% CI 12-26) in men and 16% (5-27) in women; coronary-event rates fell by 25% (16-35) and 23% (7-39); and CHD mortality rates fell by 42% (31-53) and 34% (17-50). The regression model explained an estimated 61% and 41% of variance for men and women in trends for case fatality, 52% and 30% for coronary-event rates, and 72% and 56% for CHD mortality. INTERPRETATION: Changes in coronary care and secondary prevention were strongly linked with declining coronary endpoints. Scores and benefits followed a geographical east-to-west gradient. The apparent effects of the treatment might be exaggerated by other changes in economically successful populations, so their specificity needs further assessment. PMID- 10703801 TI - Effect of phenobarbital on seizure frequency and mortality in childhood cerebral malaria: a randomised, controlled intervention study. AB - BACKGROUND: Seizures commonly complicate cerebral malaria and are associated with an increased risk of death and neurological sequelae. We undertook a randomised study to assess the efficacy of intramuscular phenobarbital in preventing seizures in childhood cerebral malaria. METHODS: Children with cerebral malaria admitted to one hospital in Kilifi, Kenya, were randomly assigned a single intramuscular dose of phenobarbital (20 mg/kg) or identical placebo. Clinical tolerance was assessed at the start of the trial, with particular reference to respiratory depression and hypotension. Seizures were timed and recorded, and treated in a standard way. Plasma phenobarbital concentrations were measured. Analyses were by intention to treat. FINDINGS: 440 children with cerebral malaria were admitted to the hospital; 100 were not recruited to the study. Of the remaining 340, 170 received phenobarbital and 170 placebo. The drug was adequately absorbed and well tolerated. Seizure frequency was significantly lower in the phenobarbital group than in the placebo group (18 [11%] vs 46 [27%] children had three or more seizures of any duration; odds ratio 0.32 [95% CI 0.18 0.58]) but mortality was doubled (30 [18%] vs 14 [8%] deaths; 2.39 [1.28-4.64]). The frequency of respiratory arrest was higher in the phenobarbital group than in the placebo group, and mortality was greatly increased in children who received phenobarbital plus three or more doses of diazepam (odds ratio 31.7 [1.2-814]). INTERPRETATION: In children with cerebral malaria, phenobarbital 20 mg/kg provides highly effective seizure prophylaxis but is associated with an unacceptable increase in mortality. Use of this dose cannot, therefore, be recommended. PMID- 10703802 TI - Awareness during anaesthesia: a prospective case study. AB - BACKGROUND: Patients who are given general anaesthesia are not guaranteed to remain unconscious during surgery. Knowledge about the effectiveness of current protective measures is scarce, as is our understanding of patients' responses to this complication. We did a prospective case study to assess conscious awareness during anaesthesia. METHODS: 11785 patients who had undergone general anaesthesia were interviewed for awareness on three occasions: before they left the post anaesthesia care unit, and 1-3 days and 7-14 days after the operation. FINDINGS: We identified 18 cases of awareness and one case of inadvertent muscle blockade that had occurred before unconsciousness. Incidence of awareness was 0.18% in cases in which neuromuscular blocking drugs were used, and 0.10% in the absence of such drugs. 17 cases of awareness were identified at the final interview, but no more than 11 would have been detected if an interview had been done only when the patients left the post-anaesthesia care unit. Four non-paralysed patients recalled intraoperative events, but none had anxiety during wakefulness or had delayed neurotic symptoms. This finding contrasts with anaesthesia with muscle relaxants, during which 11 of 14 patients had pain, anxiety, or delayed neurotic symptoms. After repeated discussion and information, the delayed neurotic symptoms resolved within 3 weeks in all patients. Analysis of individual cases suggests that a reduced incidence of recall of intraoperative events would not be achieved by monitoring of end-tidal anaesthetic gas concentration or by more frequent use of benzodiazepines. INTERPRETATION: The inability to prevent awareness by conventional measures may advocate monitoring of cerebral activity by neurophysiological techniques. However, the sensitivity of such techniques is not known, and in the light of our findings, at least 861 patients would need to be monitored to avoid one patient from suffering due to awareness during relaxant anaesthesia. PMID- 10703803 TI - False diagnosis and needless therapy of presumed malignant disease in women with false-positive human chorionic gonadotropin concentrations. AB - BACKGROUND: 12 women were diagnosed of having postgestational choriocarcinoma on the basis of persistently positive human chorionic gonadotropin (hCG) test results in the absence of pregnancy. Most of the women had extirpative surgery or chemotherapy, or both, without significant diminution in hCG titre. Our aim was to assess whether the hCG concentrations were false-positive test results. METHODS: Samples were tested for hCG, hCG free beta subunit, and hCG beta-core fragment. Assay kinetics were also assessed, and samples were tested independently by competitive RIA. False-positive hCG concentrations were identified by two criteria: detection of hCG in serum and lack of detection of hCG and its degradation products in urine; and wide variations in results for different hCG assays. We corroborated false-positive hCG values by the lack of parallel changes in hCG results when serum was diluted, by false detection of other antigens, and by failure to detect hCG with in-house assays. FINDINGS: All 12 women met both criteria for false-positive hCG, and all had corroborating findings. In all 12 cases, a false diagnosis had been made, and most of the women had been subjected to needless surgery or chemotherapy. Assay kinetics indicated that heterophilic antibodies were responsible for the false-positive results. As a result of our findings all further therapy was stopped. INTERPRETATION: Current protocols for the diagnosis and treatment of choriocarcinoma should be modified to include a compulsory test for hCG in urine. PMID- 10703804 TI - Expression of pituitary-tumour transforming gene in colorectal tumours. AB - BACKGROUND: Basic fibroblast growth factor promotes angiogenesis and mitogenesis in colon carcinomas. Pituitary-tumour transforming gene (PTTG1) causes in-vitro and in-vivo transformation, regulates secretion of basic fibroblast growth factor, and inhibits chromatid separation. Most normal tissues show little or no PTTG1 expression but cancer cells express the gene abundantly. We postulated that PTTG1 expression in colorectal tumours is related to tumour invasiveness. METHODS: PTTG1 gene and protein expression were assessed in 68 colorectal tumours and compared with invasive characteristics, such as lymph-node invasion, evidence of metastases, tumour vessel density, and expression of basic fibroblast growth factor. PTTG1 expression is given in terms of the fold-increase over that in normal-adjacent colorectal tissue. FINDINGS: PTTG1 was overexpressed in all of 48 colon carcinomas (median fold-increase 2.2 [IQR 1.8-3.3]) and in 19 of 20 colonic polyps (2.2 [1.6-3.1]) compared with normal colonic tissue. Invasion of surrounding lymph nodes was associated with higher PTTG1 expression than in carcinomas limited to the bowel wall (3.4 [2.1-5.9] vs 1.9 [1.7-2.4], p=0.007), and higher PTTG1 expression was seen in more vascular than in less vascular tumours (2.6 [1.9-5.1] vs 1.9 [1.8-2.5], p=0.04). INTERPRETATION: Increased tumour PTTG1 expression may be a marker of invasive colorectal carcinoma and could represent a new therapeutic target. PMID- 10703805 TI - Reactive arthritis after Helicobacter pylori eradication. PMID- 10703806 TI - Long-term prognosis in anorexia nervosa: lessons from a 21-year follow-up study. AB - In a prospective long-term follow-up of 84 patients 21 years after first hospitalisation for anorexia nervosa, we found that 50.6% had achieved a full recovery, 10.4% still met full diagnostic criteria for anorexia nervosa, and 15.6% had died from causes related to anorexia nervosa. Predictors of outcome included physical, social, and psychological variables. PMID- 10703807 TI - Tolerability and side-effects of post-exposure prophylaxis for HIV infection. AB - A study of HIV post-exposure prophylaxis in 28 recipients showed that indinavir containing regimens were poorly tolerated. This finding has implications for compliance and efficacy of the currently recommended combinations. PMID- 10703808 TI - Paediatric living related intestinal transplantation between two monozygotic twins: a 1-year follow-up. AB - We report on a syngeneic living related intestinal transplant in a paediatric setting with a 1-year follow-up. This procedure has allowed progressive growth and weight gain of a recipient patient and a resumption of normal activities with full social and familial reintegration. PMID- 10703809 TI - Risk of subsequent invasive breast cancer after breast carcinoma in situ. AB - The standardised incidence rates for invasive breast cancer were estimated in a cohort of 3455 women with a primary lobular or ductal carcinoma in situ of the breast. PMID- 10703811 TI - Chromosomal damage in long-term residents of houses contaminated with cobalt-60. AB - Chromosomal translocations in people who have lived in houses contaminated with radiation were substantially raised compared with controls. Retrospective biological dosimetry indicated cumulative exposures less than 1.0 Gy, which were lower than values derived from physical measurements. PMID- 10703810 TI - Hypertensive cardiovascular damage in patients with primary autonomic failure. AB - Patients with primary autonomic failure have left-ventricular hypertrophy probably as a result of night-time hypertension. PMID- 10703812 TI - Juan Rodes: pulling basic and clinical research together. PMID- 10703813 TI - US accelerates investment in biomedical research. PMID- 10703814 TI - Gene therapy HIV scare proves to be a false alarm. PMID- 10703815 TI - Therapeutic monoclonal antibodies. AB - The therapeutic potential of monoclonal antibodies (mAb) was quickly realised after the hybridoma technique allowed their development in the mid 1970s. Chimeric humanised and fully humanised mAb can now be made by recombinant engineering. About a quarter of all biotech drugs in development are mAb, and around 30 products are in use or being investigated. Licensed products are available for inhibition of alloimmune and autoimmune reactivity, and for antitumour, antiplatelet, or antiviral therapy. Short-term side-effects are tolerable and as expected, although long-term safety remains to be elucidated. The cost-effectiveness and quality-of-life benefits of the use of mAb in patients who are usually seriously and chronically ill also needs studying. The therapeutic use of mAb is now established, and is perhaps the first example of how the "new biology" and the understanding of underlying molecular mechanisms has benefited patients. PMID- 10703816 TI - Prenylation inhibitors in renal disease. AB - Members of the superfamily of Ras GTPase signalling proteins (monomeric G proteins) require post-translational carboxy-terminal prenylation to function. Prenylation is the covalent attachment of a hydrophobic prenyl group (either farnesyl or geranylgeranyl), which localises the GTPase to cell membranes. Ras proteins exert substantial control on cell proliferation and gene-transcription events, and prenylation inhibitors are now included in clinical trials for cancer. Many renal diseases are highly proliferative and are driven by a range of profibrotic cytokines. We hypothesise that inhibition of prenylation could be of substantial therapeutic benefit in such diseases, providing greater selectivity against abnormal cytokine-driven proliferation and fibrogenesis than current treatments available to nephrologists. PMID- 10703817 TI - That's what it is. PMID- 10703818 TI - Screening mammography re-evaluated. PMID- 10703819 TI - Screening mammography re-evaluated. PMID- 10703820 TI - Screening mammography re-evaluated. PMID- 10703821 TI - Screening mammography re-evaluated. PMID- 10703822 TI - Screening mammography re-evaluated. PMID- 10703823 TI - Screening mammography re-evaluated. PMID- 10703824 TI - Screening mammography re-evaluated. PMID- 10703825 TI - Screening mammography re-evaluated. PMID- 10703826 TI - Screening mammography re-evaluated. PMID- 10703827 TI - Screening mammography re-evaluated. PMID- 10703828 TI - Pharmacological control of bleeding during cardiac surgery. PMID- 10703829 TI - Renal side-effects of cyclo-oxygenase-type-2 inhibitor use. PMID- 10703830 TI - Male impotence. PMID- 10703831 TI - Relevance to PSP and MSA to clinical practice. PMID- 10703832 TI - Elective or selective use of abciximab? PMID- 10703833 TI - Sleep disturbances and tryptophan in patients with Alzheimer's disease. PMID- 10703834 TI - Sickness and sex of child. PMID- 10703835 TI - Sickness and sex of child. PMID- 10703836 TI - The QUORUM statement. PMID- 10703837 TI - Amyloid-beta junkies. PMID- 10703838 TI - Medicolegal aspects of international teleconsultancy. PMID- 10703839 TI - Shaking-impact syndrome and lucidity. PMID- 10703840 TI - Skull and crossbones. PMID- 10703841 TI - New research initiative on plant-based antimalarials. PMID- 10703842 TI - Shopping at the contraceptive supermarket. PMID- 10703843 TI - An air pollution model for use in epidemiological studies: evaluation with measured levels of nitrogen dioxide and benzene. AB - The aim of the study was to evaluate the predictions derived from the Danish Operational Street Pollution Model (OSPM) when the input data are obtained by simple methods that could be used in large-scale epidemiological studies. The model calculations were thus compared with passive sampler measurements of nitrogen dioxide and benzene at 103 street locations in Copenhagen, Denmark, and at 101 locations in rural areas. Data on traffic and street configuration were collected by means of a simple registration scheme in which forms were filled out by local municipal authorities. Meteorological data were derived from routine measurements at Copenhagen airport, and data on background air pollution were based on a simple empirical model. Differences in air pollution levels between rural areas and Copenhagen and differences in nitrogen dioxide concentrations at various locations in Copenhagen were well reproduced by the OSPM. The correlation coefficients (r) between the measured and the predicted half-year average concentrations of nitrogen dioxide in Copenhagen were between 0.75 and 0.80 for various degrees of precision of the input data for the model. The results indicate that the OSPM used with the presented methods for generation of input data might be useful in assessing long-term exposure to air pollutants in epidemiological studies. PMID- 10703844 TI - Indoor/outdoor PM10 and PM2.5 in Bangkok, Thailand. AB - Twenty-four-hour averaged PM10 and PM2.5 concentrations were obtained by using 4 liter-per-minute-pumps and impactors in microenvironments of a busy shopping district and a university hospital campus. In both areas, most people live directly adjacent to their worksites--minimizing the need to measure commuting exposure as part of total daily exposure. Co-located samplers were set in indoor microenvironments, the near-ambient zone of the households, and at nearby streetside central ambient monitoring stations. Smoking and use of other indoor PM sources were recorded daily via questionnaires. Consistent with previous studies, smoking and the use of charcoal stoves increased indoor particulate matter levels. The sampled air-conditioned hospital area had substantially lower particle concentrations than outdoors. A simple total exposure model was used to estimate the human exposure. The averaged ratios of co-located PM2.5/PM10 concentrations in various microenvironments are reported for each location. A single daily indoor average PM10 concentration for all households measured in a given sampling day is calculated for correlation analysis. Results showed that day-to-day fluctuations of these calculated indoor PM10 levels correlated well with near-ambient data and moderately well with ambient data collected at the nearby central monitoring site. This implies that ambient monitors are able to capture the daily variations of indoor PM levels or even personal exposure and may help explain the robust association of ambient PM levels and health effects found in many epidemiological studies. Absolute PM exposures, however, were substantially underestimated by ambient monitors in the shopping district, probably because of strong local sources. PMID- 10703845 TI - Evaluation of annual external radiation doses at values near minimum detection levels of dosimeters at the Hanford nuclear facility. AB - In epidemiological studies of workers exposed to ionizing radiation, recognition of the limitations of available radiation dosimetry data is important to interpretation of a study's findings. This paper provides an investigation of external radiation dosimetry data for workers at the Hanford nuclear facility, focusing on changes over time in practices for recording dosimetry measurements that were between zero and the minimum detectable level of a radiation dosimeter. Reported annual external radiation doses for the years 1944-1989 were examined for 33,459 workers; these records are the sum of periodic dosimetry measurements within a calendar year. For each year, the proportion of annual external doses with values in the range of minimum detectable level was examined. Contrary to previous researchers, we conclude that there is evidence that dosimetry measurements with values between zero and the minimum detectable level may have been recorded as zero in some historical periods. This conclusion is supported by evidence drawn from historical documentation about radiation dosimetry practices at the Hanford facility. Although workers at the Hanford facility have relatively complete and detailed external radiation dosimetry data compared to some other nuclear facilities that began operation in the 1940s, these data may suffer limitations related to dosimetry recording practices at the facility. PMID- 10703846 TI - Determination of exposure to environmental tobacco smoke in restaurant and tavern workers in one US city. AB - Approximately 173 subjects employed as waiters, waitresses, or bartenders in the Knoxville, TN, Standard Metropolitan Statistical Area collected a sample of air from their breathing zone while at their workplace for one shift. In addition, area samples were placed near the work spaces of many of the subjects. Collected samples were analyzed for respirable suspended particulate matter (RSPM), ultraviolet-absorbing and fluorescing particulate matter, solanesol, 3-ethenyl pyridine (3-EP), and nicotine. Saliva samples were collected from the subjects prior to and within 24 h following their work shift, to confirm their non-smoking status. The range of concentrations of environmental tobacco smoke (ETS) constituents encountered was considerable, e.g., for nicotine, from undetectable to more than 100 microg/m3. However, the highest RSP levels observed were considerably lower than OSHA workplace standards. Distributions of ETS concentrations suggest that there are two "ETS exposure" types of bartenders: those that work in single room bars and those that work in larger, multiroom restaurant/bars. Personal exposure to ETS of the former group was ca. 10x greater than those of the latter group, who were exposed to ETS levels more comparable to those encountered by wait staff. Exposure (concentration x duration) differences between wait staff and workers in other types of unrestricted smoking environments reported in other studies suggest that exposures in the restaurant environment may be more difficult to assess than originally considered. Salivary cotinine levels indicated that for those subjects living in smoking homes, ETS exposures outside the workplace are at least as important as those in the workplace. PMID- 10703847 TI - Biomonitoring and whole body cotton dosimetry to estimate potential human dermal exposure to semivolatile chemicals. AB - Current methods of estimating absorbed dosage (AD) of chemicals were evaluated to determine residue transfer from a carpet treated with chlorpyrifos (CP) to humans who performed a structured exercise routine. To determine the dislodgeability of residue, a California Department of Food and Agriculture (CDFA) roller was applied to a flat cotton cloth upon a treated carpet. Levels ranged from 0.06 to 0.99 microg CP/cm2. Cotton whole body dosimeters (WBD) were also used to assess residue transfer. The dosimeters retained 1.5 to 38 mg CP/person. Urine biomonitoring (3 days) for 3,5,6-trichloro-2-pyridinol (TCP) of persons who wore only swimsuits revealed a mean AD of 176 microg CP equivalents/person. The results show that the AD depends on the extent of contact transfer and dermal absorption of the residue. Default exposure assessments based upon environmental levels of chemicals and hypothetical transport pathways predict excessive exposure. The cotton WBD retains chemical residues and may be effectively used to predict dermal dose under experimental conditions. PMID- 10703848 TI - Air quality monitoring during indoor Monster Truck and car demolition shows. AB - This article describes the results of air quality monitoring in an indoor ice skating rink during three Monster Truck and car demolition exhibitions, and the public health study that was carried out. The exposure of the people present to carbon monoxide and nitrogen dioxide was continuously monitored in order to determine the time-weighted average concentrations and the maximum peaks. Nitrogen dioxide concentrations were generally under the limit of detection of the device (0.5 ppm). However, carbon monoxide levels exceeded standards for workers. Maximum time-weighted average concentrations during the exhibitions were 100 parts per million with several peaks exceeding 200 parts per million (maximum value: 1600 parts per million). Recommendations were made and during a subsequent event, the carbon monoxide concentrations were reduced to protect health. Indoor exhibitions of motorized vehicles generate significant amounts of combustion gases, which can be a health hazard. There must be sufficient ventilation and the carbon monoxide and nitrogen dioxide concentrations must be monitored. In addition, the motors of the most polluting vehicles should be adjusted before the events in order to limit the emission of combustion products. If these steps are not met, the events should be held outdoors. PMID- 10703849 TI - Residence location as a measure of environmental exposure: a review of air pollution epidemiology studies. AB - Residence location has long been used to indicate environmental exposure in many epidemiological studies. This indicator is easy to establish, requires little exposure or monitoring data, and is potentially applicable to many types of investigations. The validity, accuracy and utility of residence location as an exposure indicator, however, is challenged by current concerns regarding multiple exposure pathways, persistent and toxic contaminants, and cumulative exposures from non-point, mobile and point sources. This paper reviews 45 epidemiological studies that use residence location to identify study populations and estimate air pollution exposures. Thirteen (29%) of the studies determined environmental exposures based on "proximity" measures, usually the distance from a subject's residence to a pollutant source. Other studies used "zones" presumed to have equal pollutant levels. Several studies combined zone and proximity approaches. Exposures were quantified using monitoring data in 27 (60%) studies and dispersion modeling in two (4%) studies. Sixteen (36%) studies did not use any environmental data to quantify exposure. A total of 31 (69%) of the studies reported significant associations between health endpoints and the pollutant exposures represented by residence location. In general, comprehensive and systematic approaches to identify and estimate population exposures were not used, and the exposure estimates were therefore deemed likely to have great uncertainty. Unless exposure levels among groups are verified, it cannot be determined whether nonsignificant associations between exposures and health endpoints indicate a lack of measurable health effects, or are merely a result of exposure misclassification. Site-specific and quantitative exposure assessments are needed to better quantify and confirm exposures within such studies, as well as to permit interpretations and comparisons across studies. PMID- 10703850 TI - Conceptual basis for multi-route intake dose modeling using an energy expenditure approach. AB - This paper provides the conceptual basis for a modeling logic that is currently being developed in the National Exposure Research Laboratory (NERL) of the U.S. Environmental Protection Agency (EPA) for use in intake dose assessments involving substances that can enter the body via multiple routes of exposure. NERL is simultaneously developing a consolidated human activity database that will provide much of the data needed to parameterize equations used to implement the logic. EPA's Office of Air Quality Planning and Standards (OAQPS) is, in part, using the logic in its on-going review of the carbon monoxide national ambient air quality standard. PMID- 10703851 TI - Abdominal adhesions to prosthetic mesh evaluated by laparoscopy and electron microscopy. AB - BACKGROUND: Most adhesion experiments involve observations at a single time point. We developed a method to evaluate abdominal adhesions to surgical mesh by sequential laparoscopy. STUDY DESIGN: An abdominal wall defect was created in rats and repaired with polypropylene mesh. Sequential laparoscopic evaluation of adhesion formation was performed in each animal. The percentage of mesh area involved was scored (0% to 100%). At various time intervals animals were sacrificed and samples were obtained for light and scanning electron microscopy. RESULTS: Adhesions were already present on day 1, increased by day 7, and did not progress thereafter. Mesh surfaces free of adhesions were covered with a confluent mesothelial cell layer, first seen by scanning electron microscopy on day 5 and complete by day 7. CONCLUSIONS: Intraabdominal adhesions are best studied by sequential laparoscopy. Adhesions develop within 1 day of prosthesis placement. Adhesion-free surfaces are carpeted with mesothelial cells by day 7 and remain free thereafter, for duration of study. PMID- 10703852 TI - Gut epithelial apoptosis after severe burn: effects of gut hypoperfusion. AB - BACKGROUND: Severe cutaneous burn causes transient mesenteric vasoconstriction and altered gut mucosal integrity. We recently showed that burn also increases gut epithelial cell death by apoptosis. The goal of this study was to determine whether changes in gut perfusion after burn contribute to burn-associated gut apoptosis. STUDY DESIGN: We first correlated superior mesenteric artery blood flow with measurement of gut perfusion at the tissue level by laser doppler in four nonburned rats before, during, and after arterial clamping to validate our measurements of gut perfusion. We then characterized gut perfusion sequentially over time after burn; gut perfusion was measured 3 cm from the ligament of Treitz before burn and hourly for 6 hours. A group of control rats underwent the exact same protocol without the burn to exclude effects of anesthesia and laparotomy on tissue perfusion (n = 4). We studied a third group of rats with hypoperfusion of the same duration and magnitude induced mechanically without burn (n = 7). Sections of the proximal gut from all three groups (control without burn, burn, and hypoperfusion without burn) were examined for epithelial apoptosis. RESULTS: Linear regression analysis demonstrated a strong correlation between superior mesenteric artery blood flow and intestinal tissue perfusion measured by laser doppler under both low and high flow conditions (r = 0.85). Laser doppler measurements of gut perfusion after burn showed deceased gut perfusion that was maximal at 2 hours postburn (p < 0.05), and that persisted for 4 hours (p < 0.05). By 6 hours, gut perfusion returned to baseline. Apoptosis increased significantly in the burn group (2.11 +/- 0.17%) compared with control (0.52 +/- 0.2%) and the mechanically decreased perfusion group (0.51 +/- .03) (p < 0.001). CONCLUSIONS: We conclude that burn-induced gut hypoperfusion is insufficient to cause burn-related increased gut epithelial apoptosis. We speculate that the signal for increased gut epithelial apoptosis is primarily related to proinflammatory mediators induced by the burn wound. PMID- 10703853 TI - Survival after emergency department thoracotomy: review of published data from the past 25 years. AB - BACKGROUND: Emergency department thoracotomy (EDT) has become standard therapy for patients who acutely arrest after injury. Patient selection is vitally important to achieve optimal outcomes without wasting valuable resources. The aim of this study was to determine the main factors that most influence survival after EDT. STUDY DESIGN: Twenty-four studies that included 4,620 cases from institutions that reported EDT for both blunt and penetrating trauma during the past 25 years were reviewed. The primary outcomes analyzed were in-hospital survival rates. RESULTS: EDT had an overall survival rate of 7.4%. Normal neurologic outcomes were noted in 92.4% of surviving patients. Factors reported as influencing outcomes were the mechanism of injury (MOI), location of major injury (LOMI), and signs of life (SOL). Survival rates for MOI were 8.8% for penetrating injuries and 1.4% for blunt injuries. When penetrating injuries were further separated, the survival rates were 16.8% for stab wounds and 4.3% for gunshot wounds. For the LOMI, survival rates were 10.7% for thoracic injuries, 4.5% for abdominal injuries, and 0.7% for multiple injuries. If the LOMI was the heart, the survival rate was the highest at 19.4%. The third factor influencing outcomes was SOL. If SOL were present on arrival at the hospital, survival rate was 11.5% in contrast to 2.6% if none were present. SOL present during transport resulted in a survival rate of 8.9%. Absence of SOL in the field yielded a survival rate of 1.2%. There was no clear single independent preoperative factor that could uniformly predict death. CONCLUSIONS: The best survival results are seen in patients who undergo EDT for thoracic stab injuries and who arrive with SOL in the emergency department. All three factors-MOI, LOMI, and SOL-should be taken into account when deciding whether to perform EDT. Uniform reporting guidelines are needed to further elucidate the role of EDT taking into account the combination of MOI, LOMI, and SOL. PMID- 10703854 TI - Diagnostic value of ultrasound-guided fine-needle aspiration biopsy, core-needle biopsy, and evaluation of combined use in the diagnosis of breast lesions. AB - BACKGROUND: To investigate whether ultrasound-guided core-needle biopsy (US-CNB) has more diagnostic value for breast tumors than ultrasound-guided fine-needle aspiration biopsy (US-FNAB) and to evaluate their combined use in patients with breast tumors. STUDY DESIGN: US-FNAB was carried out in 233 patients with breast tumors (254 lesions); both US-FNAB and US-CNB (combined biopsy) were performed in 81 of these patients (82 lesions). The diagnosis obtained by US-FNAB and US-CNB was compared with the surgical findings and the diagnostic value of US-CNB and combined biopsy were retrospectively evaluated. RESULTS: The sensitivity of US FNAB was 86.9%, the specificity was 78.6%, and the accuracy was 84%. In contrast, the sensitivity of US-CNB was 86.2%, the specificity was 95.8%, and the accuracy was 89%. The specificity of US-CNB was significantly higher than that of US-FNAB and the inadequate biopsy rate of US-CNB was significantly lower than that of US FNAB. For combined biopsy, the sensitivity, specificity, and accuracy were all 100%. The sensitivity, specificity, and accuracy of combined biopsy were significantly higher than those of US-FNAB. CONCLUSIONS: These findings suggest that US-CNB is more useful than US-FNAB, and that a combination of US-CNB and US FNAB can markedly improve the preoperative diagnosis of breast cancer. PMID- 10703855 TI - Esthetic reconstruction after mastectomy for inflammatory breast cancer: is it worthwhile? AB - BACKGROUND: Because inflammatory breast cancer (IBC) has been viewed as a malignancy with a poor likelihood of longterm survival, few women have been offered esthetic reconstruction after mastectomy for IBC. Recent advances in multimodality therapy have improved the outcomes for women with this disease. The purpose of this review was to assess the results of esthetic breast reconstruction in the population with IBC. STUDY DESIGN: Review of medical records at the City of Hope National Medical Center for the 10-year period ending in May 1997, revealed 23 women who underwent elective esthetic breast reconstruction after mastectomy for IBC. The records of these patients were reviewed retrospectively. Patients requiring reconstruction for large surgical chest wall defects were not included in the review. RESULTS: Treatment for IBC included mastectomy in all patients, chemotherapy in 22, and chest wall radiation therapy in 14. Immediate reconstruction was performed at the time of mastectomy (n = 14) or was delayed (n = 9). The types of reconstruction included transverse rectus abdominis musculocutaneous flap (n = 18), latissimus dorsi flap (n = 2), or prosthetic mammary implant reconstruction (n = 3). Seven women chose to undergo additional reconstruction procedures (ie, nipple reconstruction) after their initial reconstruction. With a median followup of 44 months for survivors, 16 patients developed recurrence after reconstruction. Of these, 6 were local recurrences and 10 were distant failures. Seven patients are currently alive with no evidence of disease, 4 are currently alive with disease, and 12 have died as a result of breast cancer. The median disease-free survival after reconstruction was 19 months. The median overall survival after reconstruction for all patients was 22 months. The only negative predictor of survival was a positive surgical margin at mastectomy. CONCLUSIONS: The significant emotional and esthetic benefits of breast reconstruction should be available to women with IBC. In light of the improving prognosis of IBC with current aggressive multimodality treatment, reconstructive procedures should be offered as part of comprehensive therapy. PMID- 10703856 TI - Functional and morphologic changes of the ileal mucosa after ileoanal pouch procedure. AB - BACKGROUND: Restorative proctocolectomy is used widely for treatment of ulcerative colitis and familial polyposis coli. Limited information is available regarding the morphologic and functional adaptation of the mucosa in a functioning ileoanal pouch. STUDY DESIGN: Ileal pouch specimens from patients who underwent pouch reconstruction (mean 7.5 years postcolectomy, n = 12) were compared with normal ileum (n = 15) and normal colon (n = 5). Amino oligopeptidase (AOP) and maltase activity were measured as parameters of normal ileal function. Histologic samples were examined for the presence of neutrophils and plasma cells, the villus to crypt height ratio, and the degree of crypt hyperplasia, villus blunting, and goblet cell mass. Data were analyzed by analysis of variance. RESULTS: The AOP activity in the normal ileum was 73 +/- 32 units of enzymatic activity per gram of mucosal protein; the AOP activities of the pouch and colon were 21 +/- 22 and 16 +/- 10, respectively. The maltase activity of the normal ileum measured 254 +/- 116 units of enzymatic activity per gram of mucosal protein, and the maltase activities of the pouch and colon were 57 +/- 71 units and 29 +/- 25 units, respectively. The ileal pouch mucosa demonstrated little acute inflammation and varying degrees of chronic inflammation. Morphologically, the ileal pouch mucosa demonstrated a range of adaptations, including villus blunting and crypt hyperplasia. Several specimens contained immature epithelial cells. CONCLUSIONS: The AOP and maltase activities in mucosa from ileoanal pouches and colon were significantly lower than those in normal ileal mucosa. Ileoanal pouch mucosa from humans undergoes adaptive changes to resemble colonic mucosa both morphologically and functionally. PMID- 10703857 TI - Treatment of acute colonic pseudo-obstruction with neostigmine. AB - BACKGROUND: Colonic pseudo-obstruction is a poorly understood syndrome, described by Ogilvie, and characterized by signs of large-bowel obstruction, without a mechanical cause. An imbalance in the autonomic nerve supply to the colon has been suggested as the pathophysiology. Recently, promising results with pharmacologic manipulation with neostigmine have been described. STUDY DESIGN: A prospective study was undertaken with 11 consecutive patients with clinical and radiologic signs of colonic pseudo-obstruction, in one general hospital, over a 1 year period. Patients were treated primarily with 2.5 mg of neostigmine in 100 mL of saline for 1 hour, under cardiac monitoring. Results were assessed by the clinical and radiologic responses. RESULTS: Rapid and effective spontaneous decompression of the colon was achieved in 8 patients after a single dose of neostigmine, within a mean of 90 minutes from the beginning of treatment. In another two patients decompression occurred only after a second dose was administered 3 hours after the first dose. In one patient, no changes were observed and colonoscopic decompression was performed. No significant bradycardia was observed in any of the patients. CONCLUSIONS: Neostigmine is a simple, safe, and effective therapy for treatment of colonic pseudo-obstruction. PMID- 10703858 TI - Is there a role for surgical resection in the treatment of early-stage pancreatic lymphoma? AB - BACKGROUND: Pancreatic lymphoma is a rare neoplasm. The role of surgical resection in curing this disease is poorly defined. STUDY DESIGN: From March 1983 to July 1997, eight patients with stage I or II primary pancreatic lymphoma were identified and retrospectively reviewed. All patients received chemotherapy, five patients received radiotherapy, and three patients also underwent surgical resection. A review of the published pancreatic lymphoma experience in the English-language literature was also undertaken. RESULTS: Three patients underwent pancreaticoduodenectomy with successful resection of the lymphoma and are disease free at 64, 62, and 53 months followup. Five patients were treated with nonresectional therapy. Three are disease free at 128, 51, and 24 months. Two patients died of disease at 9 and 37 months. A review of the pancreatic lymphoma experience in the English-language literature identified 122 cases of pancreatic lymphoma. Fifty-eight of these cases represented stage I or II lymphoma, which was treated without surgical resection with a 46% cure rate. Fifteen patients who had surgical resection for localized disease have been reported with a 94% cure rate. CONCLUSIONS: Based on both our single institution experience and the literature, it is suggested that surgical resection may play a beneficial role in the treatment of localized pancreatic lymphoma, although selection factors cannot be absolutely excluded. PMID- 10703859 TI - Clinicopathologic features and postoperative prognosis of multicentric small hepatocellular carcinoma. AB - BACKGROUND: Assessment of clinicopathologic characteristics and postoperative prognoses for patients with multicentric hepatocellular carcinoma (HCC) is important to determine not only a need to operate, but also an appropriate treatment after hepatic resection. STUDY DESIGN: Between May 1990 and April 1998, among 116 patients with an initial hepatectomy for HCC measuring 3 cm or less in maximum diameter, 34 patients had multicentric HCC (MC group), and 82 patients had single nodular HCC (SN group). To clarify the clinicopathologic features of patients in the MC group versus the SN group, we compared both the clinicopathologic parameters and the postoperative prognosis after curative hepatectomy between the two groups. RESULTS: The percentages of patients positive for hepatitis B surface antigen and hepatitis C virus antibody were not significantly different between the two groups. No differences were noted in pathologic characteristics of the main tumor or tumor markers. On the other hand, in the MC group, the percentage of patients evaluated in a Child's classification as either B or C was significantly higher (p < 0.05) than that of patients in the SN group, indicating that patients with multicentric HCC have a poor hepatic functional reserve. Both survival and disease-free survival of patients in the MC group who underwent a curative hepatectomy did not differ statistically from those in the SN group. CONCLUSIONS: Our results indicate that hepatic resection is useful, even for patients with multicentric HCC, if a curative hepatectomy can be performed and liver function can be saved, despite their poor hepatic functional reserve. PMID- 10703860 TI - Adding laparoscopy to experimental adhesion study protocols. PMID- 10703861 TI - Lessons from a life in surgery: close doesn't count. PMID- 10703862 TI - Should we regionalize major surgery? Potential benefits and policy considerations. PMID- 10703863 TI - Neoadjuvant chemotherapy in women with invasive breast carcinoma: conceptual basis and fundamental surgical issues. PMID- 10703864 TI - Irradiation for intimal hyperplasia: implications for peripheral arterial bypass. AB - Irradiation has been shown to inhibit postangioplasty intimal hyperplasia ("restenosis") in unbranched tubes. It seems likely that irradiation will similarly be able to inhibit intimal hyperplasia after a surgical anastomosis at a biochemical and cellular level, but whether it will produce a clinically relevant or even clinically detectable difference is unproved. One possibility is that no clinical effect may occur; the search for a "cure" for intimal hyperplasia has been long and, as yet, unsuccessful. On the other hand, if a strong effect without insurmountable logistical problems could be produced, one major cause of bypass graft failure would be preventable. Not only would the incidence of late graft occlusion, need for reoperation, and limb loss be reduced, but, if patency of prosthetics could be sufficiently improved, the initial operation could be made much easier, faster, and perhaps safer. PMID- 10703865 TI - Abdominal compartment syndrome from intractable constipation. PMID- 10703866 TI - Endoscopic cervico-thoraco-abdominal esophagectomy. PMID- 10703867 TI - Posterior transsacral approach: an alternative for the resection and reconstruction of severe ileoanal anastomotic strictures. PMID- 10703868 TI - Carcinoma of the gallbladder associated with anomalous junction of the pancreaticobiliary duct. PMID- 10703869 TI - The angiotensin type 2 receptor: variations on an enigmatic theme. AB - Since its discovery and molecular characterization, the angiotensin AT2.receptor has been enigmatic with respect to signalling pathways and function. Evidence now emerges that angiotensin II exerts actions through the AT2 receptor which are directly opposed to those mediated by the AT1 receptor. This can be exemplified e.g. by mutually antagonizing effects on cell growth. Upregulated by the endogenous agonist itself, as well as by several growth- and differentiating factors in development and tissue injury, the AT2 receptor appears to act as a modulator of complex biological programmes involved in embryonic development, cell differentiation, tissue protection and regeneration, as well as in programmed cell death. Research on the AT2 receptor has thus unveiled hitherto unknown functions of the renin-angiotensin system extending far beyond the classical role of this old hormonal system in cardiovascular control. PMID- 10703870 TI - Beneficial interactions between pharmacological, pathophysiological and hypertension research. AB - The treatment of essential hypertension continues to be carried out by drugs, combined with the adaptation of life style. The development of various types of antihypertensive drugs has not only greatly improved the management of hypertension, but also offered significant methodological sophistication of the pharmacological and pathophysiological sciences. Antihypertensive drugs and related experimental agents have been widely used in pharmaco-logical and pathophysiological research. The beneficial effects of such agents will be illustrated by means of several examples, emphasizing the sympathetic nervous system, the renin-angiotensin-aldosterone system, and calcium homeostasis as major targets. As pharmacological tools, which are also antihypertensives, we discuss various types of centrally acting antihypertensives, ganglionic and peripheral neuronal blocking agents, alpha- and beta-adrenoceptor antagonists, angiotensin converting enzyme (ACE)-inhibitors, renin-inhibitors, angiotensin II receptor antagonists (AT1-blockers) and calcium antagonists. Finally, a few remarks will be made concerning the beneficial therapeutic effects of classic and newer antihypertensive drugs, such as beta-blockers, diuretics, calcium antagonists, ACE-inhibitors and AT1-blockers. PMID- 10703871 TI - Relationship between dental pain perception and 24 hour ambulatory blood pressure: a study on 181 subjects. AB - OBJECTIVE: To investigate dental pain perception in a large group of essential hypertensive subjects. METHODS: A total of 130 hypertensive patients together with 51 normotensive subjects were submitted to tooth-electrical stimulation to determine the dental pain threshold (occurrence of pulp sensation) and tolerance (time when the subject asked for the test to be stopped). Blood pressure was measured at rest, before pain perception evaluation, and during a 24 h period by ambulatory monitoring. RESULTS: The normotensive and hypertensive subjects differed with regard to pain threshold (P = 0.002) and tolerance (P = 0.01). Pain perception variables were significantly correlated with both resting blood pressure and 24 h, diurnal and nocturnal arterial pressures, the correlation between pain threshold and 24 h systolic blood pressure being the most significant (r = 0.31, P < 0.0001). By contrast, parameters indicating 24 h blood pressure variability (percentage of nocturnal blood pressure reduction and 24 h blood pressure variation coefficients) were not associated with pain perception. Moreover, among the hypertensives only, a significant relationship was observed between pain sensitivity and both baseline and 24 h pressures. No association was found when pain perception and blood pressure were correlated in the normotensive group. CONCLUSIONS: The correlation between both baseline and 24 h blood pressure and pain perception has been confirmed in a large group study of normotensive and hypertensive subjects. Moreover, even among the hypertensive range of blood pressure, the higher the blood pressure is, the lower the sensitivity to pain is. These findings strengthen the hypothesis of a role of the degree of blood pressure elevation in modulating pain sensitivity. PMID- 10703872 TI - Autonomic nervous system activity in dipper and non-dipper essential hypertensive patients. What about sex differences? AB - OBJECTIVES: To compare the autonomic nervous system activity indexes obtained from photoplethysmography in dipper and non-dipper hypertensive patients and to seek a potential influence of sex on the relation between autonomic nervous system and the nocturnal decrease in blood pressure. METHODS: We studied 245 hypertensive patients, who underwent 24 h ambulatory blood pressure monitoring (ABPM), photoplethysmographic blood pressure recording, and echocardiography. Non dipping patients were defined as those whose nocturnal decrease in systolic blood pressure (SBP), diastolic blood pressure (DBP), or both was less than 10% of the daytime blood pressure. Spectral powers of SBP, DBP and heart rate were obtained from photoplethysmographic recordings over three main frequency bands: very low frequency (0.005-0.05 Hz), low frequency (0.05-0.14 Hz) and high frequency (0.14 0.40 Hz). RESULTS: Because their ABPM were normal (less than 135/85 mmHg; n = 33), of poor quality (n = 22) or performed at a period too far from the photoplethysmographic recording (n = 17), 66 patients were excluded from the analysis. The remaining 179 patients comprised 117 dippers and 62 non-dippers. The groups did not differ regarding clinical and echocardiographic characteristics, irrespective of sex. Low-frequency spectral powers were significantly lower in non-dippers than in dippers, whatever the signal, whereas high-frequency spectral powers did not differ significantly between the groups. The nocturnal decrease in blood pressure increased with increasing low-frequency spectral powers, but was negatively correlated with high-frequency spectral powers. Multivariate linear regression analysis identified low-frequency spectral power of SBP and clinic DBP as independent factors determining the decrease in blood pressure. After adjustment for all significant covariates, the odds of being a non-dipper did not differ between men and women. CONCLUSION: A non-dipper profile seemed to be associated, in both men and women, with lower low-frequency spectral powers compared with those in dippers, suggesting impaired sympathetic arterial modulation. PMID- 10703873 TI - Use of computerized neuropsychological tests (CANTAB) to assess cognitive effects of antihypertensive drugs in the elderly. Cambridge Neuropsychological Test Automated Battery. AB - OBJECTIVE: To establish reliability and ease of use of the Cambridge Neuropsychological Test Automated Battery (CANTAB) in assessing changes in cognitive function induced by antihypertensive drugs. DESIGN AND METHODS: Standard neuropsychological testing was combined with CANTAB in a double-blind 18 week cross-over study in elderly hypertensives taking perindopril or hydrochlorothiazide/triamterene (HT). Cognitive effects were assessed by employing tests of attention, visuospatial and verbal memory, learning, reasoning, planning, problem solving, speed and coordination. Affect was assessed using two different depression-rating scales. RESULTS: Perindopril and the diuretic had no adverse effects on the various aspects of cognitive function. Mood, as assessed by the Hamilton Depression Rating Scale and the Beck Depression Inventory, was improved on Perindopril, and the error rate in the motor screening test was lower. Ambulatory blood pressure monitoring showed both drugs achieved effective 24-h control. CONCLUSIONS: The ease of use and the ability to adjust the level of testing to the requirements of individual patients, together with the reliability of longitudinal test/re-test results, indicates that CANTAB is an important addition to the methods available to quantitate adverse central nervous system drug effects. The other purpose of the study was to assess any adverse cognitive effects of perindopril against a drug HT believed to have no adverse central nervous system effects. In this context, perindopril was free of adverse effects in all the objective tests employed. In addition, there was a benefit seen in two independent assessments of depression (the Hamilton and the Beck rating scales). PMID- 10703874 TI - Aortic pulse wave velocity in young normotensives with a family history of hypertension. AB - OBJECTIVE: To assess the effect of selected clinical and biochemical parameters, with particular consideration of familial hypertension, on the pulse wave velocity (PWV) in young normotensives. SUBJECTS AND METHODS: Seventy voluntary students were enrolled (mean age 22.3+/-2.1 years), 39 men and 31 women, with normal blood pressure. A history was obtained with respect to diabetes mellitus, ischaemic heart disease, lipid disorders and arterial hypertension in the family. The subjects were subdivided into two groups: those with (n = 33) and without (n = 37) a family history of arterial hypertension, and blood pressure and heart rate were measured three times and total cholesterol and its subfractions determined in plasma. The carotid to femoral PWV was measured using an automatic computerized recorder and analysed by the Complior program. RESULTS: The subjects with a family history of arterial hypertension had higher blood pressure levels (systolic and diastolic blood pressure, pulse pressure and mean arterial pressure), as well as mean body mass index and low-density lipoprotein (LDL) cholesterol. The PWV in this group did not differ from that in the subjects without a family history of arterial hypertension (9.69+/-2.8 versus 9.32+/-2.0). However, the PWV was significantly higher in males than females (10.62+/-2.2 versus 7.86+/-1.13, P < 0.0001) and there was a significant positive correlation between male gender and PWV. CONCLUSIONS: Familial arterial hypertension does not significantly affect aortic stiffness in terms of PWV. Male gender in this population of young healthy subjects is one of the most important factors associated with central arterial stiffness. PMID- 10703875 TI - Carotid wall inertial index increase is related to intima-media thickening in hypertensive patients. AB - OBJECTIVE: The aim of this study is to evaluate the relationship between carotid intima-media thickness (IMT) and arterial wall inertial behaviour. METHODS: The simultaneous and noninvasive assessment of the intima-media complex and arterial diameter waveform was performed using high-resolution ultrasonography. The common carotid artery of eleven normotensive subjects (NTA) and eleven mild-to-moderate essential hypertensive patients (HTA) were measured noninvasively using tonometry and an automatic densitometric analysis of B-mode images to obtain IMT and instantaneous pressure (P) and diameter (D) loops. A linear discrete time model was used to estimate the inertial index (K(M)) using a system modelling identification approach. RESULTS: In NTA K(M) was 0.333+/-0.256 (mmHg x s2/mm) and IMT 0.643+/-0.061 (mm), whereas in HTA K(M) was 0.798+/-0.590 (P < 0.05) and IMT 0.760+/-0.034 (P < 0.025). When all data of K(M) versus IMT of NTA and HTA were pooled in a linear regression analysis, a correlation coefficient of r = 0.61 (P < 0.05) was obtained. CONCLUSION: Wall inertia increase was associated with a higher IMT, suggesting that the intima-media thickening might be partially related to vascular hypertrophy manifested as increase of inertial behaviour. PMID- 10703876 TI - The effect of weight loss with or without exercise training on large artery compliance in healthy obese men. AB - BACKGROUND: Obesity is an independent risk factor for cardiovascular morbidity and mortality. Large artery compliance is thought to be associated with cardiovascular risk. The effect of weight loss on large artery compliance is not yet clarified. OBJECTIVE: To investigate the effect of weight loss, with or without exercise, on vessel wall properties in healthy obese men. DESIGN: This was a pair-matched randomized intervention study. All subjects were on an energy restricted diet. One subject from each pair was also on an exercise programme. Measurements were performed before and at the end of the study period. The study lasted for 3 months. METHODS: The vessel wall properties of the brachial and common carotid artery were assessed using a vessel wall movement detector system in combination with applanation tonometry. RESULTS: The mean body mass index was 32.3+/-0.4 kg/m2 and decreased (P < 0.001) to 27.6+/-0.4 kg/mm2 during the study. The mean blood pressure decreased (P < 0.001) by 6%. At operating pressures, carotid artery distensibility was 27.5+/-1.7 x 10(-3)/kPa at the start of the study and 31.1+/-1.8 x 10(-3)/kPa (P < 0.04) at the end of the study. Brachial and carotid artery compliances were 0.11+/-0.01 and 1.35+/-0.08 mm2/kPa at the start of the study and tended to increase to 0.12+/-0.001 (P = 0.06) and 1.48+/ 0.08 mm2/kPa (P = 0.057), respectively, at the end of the study. Isobaric compliance did not change. The diet-and-exercise group did not differ statistically from the only-diet group in the effects on weight loss, blood pressure and arterial compliance. CONCLUSION: This study shows that weight loss increased carotid artery distensibility at operating pressures, but not under isobaric conditions. This increase is probably due to the decrease in blood pressure. The addition of exercise did not result in an additional effect within 3 months. PMID- 10703877 TI - Comparative effects of different dihydropyridines on the expression of adhesion molecules induced by TNF-alpha on endothelial cells. AB - OBJECTIVE: Lacidipine has already been demonstrated to reduce the expression of some adhesion molecules induced by pro-oxidant signals on endothelial cells. In order to verify if this effect is a peculiarity of this molecule, or belongs to other dihydropyridinic compounds (DHPs), the activity of lacidipine was compared with that of lercanidipine, amlodipine, nimodipine and nifedipine. DESIGN AND METHODS: The compounds were incorporated in human umbilical vein endothelial cells (HUVECs) using native low-density lipoprotein as a carrier. The drug concentrations in HUVECs were measured by mass spectrometry. Human recombinant tumour necrosis factor-alpha was then incubated with HUVECs for 7 h at 37 degrees C for adhesion molecule expression. RESULTS: The cellular amount of lacidipine, lercanidipine and amlodipine was similar, while nimodipine and nifedipine were almost undetectable or undetectable, respectively. Lacidipine, at any concentration, determined a dose-dependent significant decrease of the expression of intercellular adhesion molecule-1 (ICAM-1) ICAM-1, vascular cell adhesion molecule-1 (VCAM-1) VCAM-1 and E-selectin (P < 0.01). Lercanidipine and amlodipine determined variable decreases of adhesion molecules at the intermediate and highest concentrations. Nimodipine and nifedipine determined no effect on ICAM-1, VCAM-1 and E-selectin. The lowest IC50, i.e. the concentration determining the 50% reduction of ICAM-1, VCAM-1 and E-selectin expression was obtained with lacidipine for all the adhesion molecules considered (P < 0.01). CONCLUSIONS: It is concluded that the effect of the DHPs used in this study on adhesion molecule expression is determined first by their lipophilicity and then by their intrinsic antioxidant activity. PMID- 10703878 TI - Bosentan reduces blood pressure and the target-organ damage induced by a high fructose diet in rats. AB - BACKGROUND: Rats fed a high-fructose diet develop hyperinsulinaemia, hypertriglyceridaemia, hypertension, renal changes similar to those in diabetic rats and left ventricular hypertrophy with deposition of collagen. Bosentan is an antagonist of endothelin receptors. Other authors have demonstrated that bosentan is effective in preventing the increase in blood pressure induced by a high fructose diet but, until now, the effect of the drug on the target organs has not been investigated. OBJECTIVE: To evaluate whether bosentan is effective, not only in reducing blood pressure, but also in limiting the renal and cardiac changes induced by a high-fructose diet METHODS: Forty Wistar-Kyoto (WKY) male rats were divided into four groups: groups 1 and 2 received a high-fructose diet, groups 3 and 4 received a standard diet for 1 month. Thereafter, the following treatments were administered: group 1, high-fructose diet plus bosentan 100 mg/kg per day; group 2, high-fructose diet plus placebo; group 3, standard diet plus bosentan 100 mg/kg per day; group 4, standard diet plus placebo. After a further 1 month, all animals were killed. A morphometric analysis was performed by examining 100 glomeruli for each animal. Renal deposits of collagen and fibronectin and cardiac deposits of collagen III were measured by means of immunochemistry. RESULTS: By the end of the study, bosentan had completely reversed the increase in blood pressure induced by a high-fructose diet, without modifying the blood pressure in normotensive rats. Moreover, bosentan reduced glomerular hypertrophy and deposits of collagen and fibronectin in the kidney and cardiac deposits of collagen III. CONCLUSIONS: The results of this study demonstrate that bosentan not only normalizes blood pressure, but also protects target organs in rats receiving a high-fructose diet. PMID- 10703879 TI - Differential brain atrial natriuretic peptide expression co-segregates with occurrence of early stroke in the stroke-prone phenotype of the spontaneously hypertensive rat. AB - OBJECTIVE: To determine how the downregulation of atrial natriuretic peptide (ANP) gene expression, previously demonstrated to occur only in the brain of the stroke-prone spontaneously hypertensive rat (SHRsp), in contrast to the stroke resistant SHR (SHRsr), co-segregates with stroke occurrence in SHRsp/SHRsr F2 descendants in order to study the 'protective' role towards stroke previously demonstrated in SHRsp for the quantitative trait locus STR2 that also carries the ANP gene. DESIGN AND METHODS: Eight male SHRsp, eight male SHRsr and 16 male SHRsp/SHRsr F2-intercross animals (progeny of brother/sister mated F1 hybrids from an original cross between F0 SHRsp and SHRsr) were selected for this study. All rats were exposed to a stroke-permissive Japanese-style diet starting at the age of 6 weeks. Half of the F2 animals had early strokes; the remainder had late strokes. Blood pressure was measured before sacrifice. Analysis of brain ANP expression using an RNase protection assay was performed in all animals. RESULTS: Downregulation of brain ANP in the stroke-prone phenotype was found to co segregate with the occurrence of early strokes in the F2 rats independently of blood pressure levels. CONCLUSIONS: The observed lower expression of ANP in the brains of stroke-prone rats appears to be the result of an inhibitory effect by another gene or genes. It seems unlikely that this specific trait represents a primary protective mechanism. PMID- 10703880 TI - Hypertension aggregates in families of kidney stone patients with high urinary excretion of uric acid. AB - OBJECTIVE: To determine whether kidney stone disease (KSD) and hypertension (HTN) share a common familial component that is determined by a specific urinary biochemical abnormality. DESIGN: Familial aggregation study. PATIENTS: Two hundred and twelve KSD patients, aged 18-50 years, collected a 24-h urine sample to measure the urinary excretion of uric acid, calcium, oxalate, magnesium and citrate, and were interviewed about the occurrence of HTN among first-degree relatives. OUTCOME: Positive family history (FHx) of HTN defined as two or more relatives with HTN, and HTN occurring in the fathers, mothers and siblings. RESULTS: Positive FHx of HTN was significantly associated with increasing urinary excretion of uric acid (P = 0.03) but not with the excretion of the other substances. When the patients were divided into those with and without hyperuricosuria, the adjusted odds ratio (OR) for positive FHx of HTN in a hyperuricosuric KSD patient was 3.8 (95% CI, 1.22-11.66). Separate analysis on the occurrence of HTN in the fathers, mothers and siblings of the probands indicated that hyperuricosuria is positively related to HTN occurring in the siblings of the patients (P < 0.001) but not in the fathers or in the mothers. The adjusted OR for HTN occurring in siblings of hyperuricosuric patients compared with siblings of non-hyperuricosuric patients was 3.8 (2.12-6.67). CONCLUSION: Siblings of KSD patients with hyperuricosuria had a significantly increased prevalence of HTN that could not be accounted for by age, family size, body-mass index and personal history of HTN of the probands. Additional studies need to be undertaken to determine whether this familial clustering has a genetic or environmental origin. PMID- 10703881 TI - Sibling resemblance of erythrocyte ion transporters in French-Canadian sibling pairs affected with essential hypertension. AB - OBJECTIVES: Erythrocyte Na+/Li+ countertransport and Na+,K+ cotransport are increased in some Caucasians with essential hypertension. This study examines the relative contributions of genetic and shared environmental factors to the activity of these ion carriers in French-Canadian sibling-pairs affected with essential hypertension. DESIGN: The activity of Na+/Li+ countertransport and Na+,K+ cotransport (rate of Na+ o-dependent Li+ efflux and bumetanide-sensitive 86Rb influx, respectively) was measured in 122 French-Canadian siblings with essential hypertension, including 36 brother/brother and 48 sister/sister pairs. Sibling/sibling correlations were estimated using the FCOR program of the S.A.G.E. package. RESULTS: Na+/Li+ countertransport and Na+,K+ cotransport were respectively higher by 27% (P = 0.002) and 42% (P = 0.0009) in erythrocytes from men compared with women. Intra-individual correlation analysis did not reveal a significant effect of age on the activity of these ion transporters in both males and females, and an influence of plasma lipids (triglycerides, cholesterol, low density lipoprotein, high-density lipoprotein) in females. In males, Na+,K+ cotransport was correlated with the level of serum triglycerides only (P = 0.01). Familial correlation analysis showed that sibling resemblance of Na+/Li+ countertransport and Na+,K+ cotransport was higher in men (r = 0.26 and 0.39) than in women (r = 0.01 and 0.03, respectively). CONCLUSION: The present data indicate that different factors contribute to the regulation of monovalent ion carriers in erythrocytes from Caucasian men and women with essential hypertension. The activity of erythrocyte Na+/Li+ countertransport and Na+,K+ cotransport appears to be more strongly determined by inheritable factors in men than in women. PMID- 10703882 TI - Angiotensin I to angiotensin II conversion in the human forearm and leg. Effect of the angiotensin converting enzyme gene insertion/deletion polymorphism. AB - OBJECTIVE: The angiotensin-converting enzyme (ACE) gene I/D polymorphism accounts for part of the variation in ACE concentration; subjects with one or two D alleles have approximately 25 and 50% higher ACE levels, respectively, than subjects with two I alleles. Data from studies on the pressor effects of angiotensin (Ang) I in DD compared with II subjects are inconsistent, because enhanced conversion in DD subjects may have been masked by a decreased responsiveness to Ang II. Here we quantify ACE genotype-related Ang I to Ang II conversion in the human forearm and leg using non-pressor 125I-Ang I infusions. DESIGN AND METHODS: Infusions were given to 12 women and 17 men (age 24-67 years) who were undergoing renal vein sampling followed by renal angiography for diagnostic purposes. 125I-Ang I was infused for 20 min into the right antecubital vein, and blood samples for the measurement of 125I-labelled and endogenous Ang I and Ang II were taken from the aorta, the left antecubital vein and a femoral vein under steady-state conditions. Genotype frequencies were determined by polymerase chain reaction. RESULTS: Fractional conversion (i.e. the percentage of arterially delivered 125I-Ang I that is converted to 125I-Ang II) in the forearm (38+/-4, 30+/-3 and 31+/-6% in 8 II, 16 ID and 5 DD subjects, respectively; mean +/- SEM) and leg (52+/-4, 48+/-3 and 42+/-5%) was similar in all three groups. In addition, no genotype-related differences in plasma Ang II/I ratio (a measure of ACE activity) were observed at the three sampling sites. CONCLUSIONS: Regional Ang I to Ang II conversion does not parallel the previously described D allele related differences in ACE concentration, suggesting that effects other than enhanced conversion may underlie the reported associations between the D allele and various cardiovascular diseases. PMID- 10703883 TI - Modulation of the AT2 subtype receptor gene activation and expression by the AT1 receptor in endothelial cells. AB - OBJECTIVE: To investigate whether angiotensin II type 2 (AT2) receptor (AT2-r) promoter activity and expression are modulated by angiotensin II (Ang II), and whether the AT1 receptor (AT1-r) is involved in this effect. DESIGN AND METHODS: Primary endothelial cells obtained from NEONATAL rat aorta, expressing both receptors, were transfected with the rat AT2-r promoter region cloned into a pCAT reporter vector. The reporter-expression study was performed in a transient transfection assay system. Transfected cells were studied following angiotensin converting enzyme inhibition to prevent endogenous formation of Ang II. Cells were subsequently stimulated for 6 h with Ang II, either alone or in combination with the AT1-r antagonist DuP753. AT2-r mRNA was assessed by RNase protection assay during the same pharmacological stimuli. RESULTS: Stimulation with Ang II caused an increase in promoter activity (+50%, P < 0.05 versus baseline), whereas mRNA expression was reduced by 50% (P < 0.05 versus baseline). Concomitant treatment with DuP753 and Ang II was associated with a 98% increase in promoter activity (P < 0.05 versus baseline). DuP753 also prevented the reduction in mRNA; it actually produced a 100% increase in AT2-r mRNA accumulation (P < 0.01 versus baseline). Studies with the AT2-r antagonist PD123319 indicate that the AT2-r is also involved in the regulation of AT2-r gene promoter activity. CONCLUSIONS: These data indicate that Ang II increases AT2-r promoter activity and decreases AT2-r mRNA accumulation in endothelial cells. The AT1 subtype receptor is involved in the modulation of both effects of Ang II. These findings suggest that changes in the expression of AT2 receptors may occur during treatment with AT1-r antagonists, and they indicate the existence of a cross-talk between AT1 and AT2 receptors. PMID- 10703884 TI - No vasoactive role of the angiotensin II type 2 receptor in normotensive Wistar rats. AB - OBJECTIVE: To investigate the vasoactive consequences of angiotensin II type 2 receptor stimulation in vivo. DESIGN AND METHODS: Three consecutive 10 min intravenous infusions of angiotensin (Ang) II (100, 300 and 1000 ng/kg per min) were given to 20 pentobarbitone-anaesthetized normotensive Wistar rats (weight 330+/-6 g, mean +/- SEM). The rats had been pretreated with saline (n = 8), the angiotensin II type 1 receptor antagonist, irbesartan (100 microg/kg per min for 30 min, n = 6), or the angiotensin II type 2 receptor antagonist, PD123319 (20 microg/kg per min for 30 min followed by continuous infusion throughout the entire experiment, n = 6). Regional haemodynamic effects of Ang II were studied using the radioactive microsphere method. RESULTS: Ang II increased mean arterial blood pressure (MAP) and heart rate by, maximally, 44+/-9 and 26+/-6%, respectively (P < 0.05 compared with baseline), and decreased cardiac output and systemic vascular conductance (cardiac output/MAP) by, maximally, 24+/-8 and 47+/ 4%, respectively (P < 0.05 compared with baseline). The Ang II-induced decrease in systemic vascular conductance was caused by decreases in vascular conductances (regional flow/MAP) of the gastrointestinal tract (52+/-4%), kidney (63+/-3%), skeletal muscle (39+/-8%), skin (63+/-4%), mesentery + pancreas (32+/-11%), adrenal (27+/-11%) and spleen (57+/-6%) (all P < 0.05 compared with baseline). Irbesartan increased baseline vascular conductances in adrenal, brain and kidney, and inhibited all haemodynamic responses induced by Ang II. PD123319 affected neither baseline values nor the Ang II-induced haemodynamic responses. CONCLUSIONS: Ang II-induced systemic and regional haemodynamic effects in normotensive Wistar rats are mediated exclusively via angiotensin II type 1 receptors. No evidence for angiotensin II type 2 receptor-mediated vasoactive responses was obtained. PMID- 10703885 TI - Cardiac interstitial fluid levels of angiotensin I and II in the pig. AB - OBJECTIVE: To study whether cardiac interstitial fluid levels of angiotensin I and II (Ang I and II) can be monitored in vivo, using the microdialysis technique, and to assess the contribution of plasma-derived angiotensins to the interstitial fluid levels of these peptides. DESIGN AND METHODS: Microdialysis probes were placed in the left ventricular (LV) myocardium of eight anaesthetized pigs, three of which were untreated and five treated with the angiotensin II type 1 (AT1) receptor antagonist L-158,809 (10 mg intracoronary). All pigs were given a 1 h intracoronary infusion of 125I-Ang II. Aortic and coronary venous blood samples were taken under steady-state conditions, and interstitial dialysate was collected during the entire infusion period. Immediately after stopping the infusion, LV tissue pieces were obtained at various time points. RESULTS: L 158,809 did not affect the levels of endogenous Ang I and II or the levels of plasma 125I-Ang II. Aortic Ang I and II levels (22 and 16 fmol/ml; geometric mean of eight pigs) were comparable to coronary venous Ang I and II levels, whereas the coronary venous 125I-Ang II levels (6650 c.p.m./ml) were approximately 30 times higher than those in the aorta. Tissue Ang I and II levels were 5 and 17 fmol/g, respectively. In untreated animals, the 125I-Ang II levels per g LV tissue were similar to the levels per ml coronary venous plasma, and the ex vivo half-life of tissue 1251-Ang II was > 30 min. In treated animals, tissue 125I-Ang II was < 5% of coronary venous 125I-Ang II and became undetectable within 15 min. 125I-Ang II, Ang I and Ang II levels in the interstitial fluid were close to or below the detection limit (200 c.p.m., 60 fmol and 20 fmol per ml, respectively) in all animals. CONCLUSIONS: Plasma and myocardial interstitial fluid angiotensin levels are of the same order of magnitude. Plasma Ang II does not contribute to the interstitial fluid level of Ang II, most likely because of its rapid metabolism in the vascular wall. Binding to AT1 receptors protects Ang II against metabolism. PMID- 10703886 TI - Effects of an auditory startle stimulus on blood pressure and heart rate in humans. AB - OBJECTIVE: To describe the effects of an auditory startle stimulus on blood pressure (BP) and heart rate (HR) in humans. DESIGN AND METHODS: Twenty-five volunteers, including nine untreated hypertensive subjects, were studied in the supine position. Polygraphic recordings were obtained for finger BP, R-R interval using ECG, respiratory movements using a thoracoabdominal belt and for electrooculomyogram using adhesive electrodes. Haemodynamic estimations were derived by modelling flow from the noninvasive BP signal. A background noise of 55 dB was administered through headphones and two acoustic startle stimuli (110 dB, 1-20 kHz, 0.15 s) were generated at 5-min intervals during the tele expiratory phase. The sham stimulation (0 dB, event marker) was compared with the effects of the noise stimulus (one-way ANOVA with repeated measures followed by a protected t test for multiple comparisons). RESULTS: A biphasic cardiovascular profile was observed in response to noise stimulation. Blood pressure and HR increases were combined in the early response (0-10 s) observed after the immediate motor contraction (blink). The average systolic BP rise was 18.7+/-2.7 mmHg (peak at 5.1 s) and the average HR increase was 10.8+/-1.1 bpm (peak at 3.4 s) for the first stimulus. These effects were highly significant compared with the sham response (P < 0.01). The second stimulus elicited BP and HR rises of a lesser amplitude (P < 0.01). The delayed response (10-30 s) corresponded with a moderate BP decrease. The haemodynamic indexes suggest that the early rise in blood pressure reflects a rise in total peripheral resistance. CONCLUSION: This is the first description of the BP response to an acute loud noise in humans. The early (within 10 s) BP and HR rises may depend upon the autonomic component of the startle reflex. One application of this test could be the discrimination of the different classes of antihypertensive drugs according to their sites of action. PMID- 10703888 TI - Altered blood pressure variability in patients with congestive heart failure. AB - OBJECTIVE: Congestive heart failure (CHF) is characterized by sympathetic overactivity but reduced variability of heart interval and sympathetic nerve activity; little information exists, however, about the alterations in blood pressure variability in this syndrome, especially during excitatory manoeuvres such as tilting or exercise. DESIGN AND METHODS: Nine patients with CHF (age 62+/ 1 years, NYHA class II-III, ejection fraction 33+/-1%, peak VO2 14.1+/-3.2 ml/min per kg body weight [mean +/- SEM]) and eight healthy control subjects (age 58+/-1 years) with normal left ventricular function were studied. Blood pressure (Finapres), R-R interval (ECG) and respiration (nasal thermistor) were recorded during 15-min periods of supine rest, 70 degree head-up tilting, submaximal bicycling exercise and post-exercise recovery. Total variance and the power of the spectral components of blood pressure (HF, respiratory-related; LF, 0.03-0.14 Hz; and VLF, 0.02-0.003 Hz) were measured. RESULTS: Compared with control subjects, CHF patients have, first, a normal overall blood pressure variability during supine rest but a failure to increase this variability in response to head up tilt and exercise; second, a suppressed LF spectral component of blood pressure at rest and in response to head-up tilt and exercise; and third, reappearance of LF blood pressure power during postexercise recovery. CONCLUSIONS: In CHF patients, overall blood pressure variability and its LF spectral component are altered at rest and during sympathoexcitatory manoeuvres. Somewhat paradoxically, however, the depressed LF blood pressure power is partially restored during a 15-min recovery period, indicating that at least part of the CHF-related alterations of blood pressure variability have the potential to revert back towards normal under appropriate physiological circumstances. PMID- 10703887 TI - Cardiac autonomic adjustments to normal human pregnancy: insight from spectral analysis of R-R interval and systolic arterial pressure variability. AB - OBJECTIVE: To assess the adaptation in autonomic control mechanisms that accompanies the marked haemodynamic changes, such as increases in cardiac size and output, that occur in the course of normal human pregnancy. DESIGN: We studied 14 healthy pregnant women (aged 30+/-1 years) before the 6th week (early stage) and within weeks 32-34 (late stage) of pregnancy, while they were at rest or in a state of active orthostatism (standing), which enhances sympathetic activity. METHODS: We used echocardiography to assess cardiac volumes and mass, and spectral analysis of the R-R interval and systolic arterial pressure variability to obtain indices of autonomic regulation of the circulation. This non-invasive methodology, recently validated with direct recordings of muscle sympathetic nerve activity, furnishes quantitative markers of sympathetic modulation of the sino-atrial node (low frequency component, LF in normalized units, nu), vagal modulation (high frequency component, HF in normalized units, nu) and the overall arterial pressure-heart rate baroreflex gain (alpha index). RESULTS: Late pregnancy was characterized by an increase in cardiac size and volumes and by a reduction of R-R interval, R-R interval variance and the alpha index, together with an increase in the LF/HF ratio (from 1.4+/-0.4 to 5.6+/ 1.9). Changes in markers of autonomic modulation of the sino-atrial node normally induced by the standing position were blunted. CONCLUSIONS: The late stage of normal human pregnancy appears to be characterized by alterations in the autonomic control of the circulation and by attenuated responsiveness to active standing, possibly as a consequence of the accompanying increase in cardiac size. PMID- 10703889 TI - Transcapillary escape rate of albumin is increased and related to haemodynamic changes in normo-albuminuric type 1 diabetic patients. AB - OBJECTIVE: An increase in urinary albumin excretion (UAE) in type 1 diabetic patients might reflect changes in vascular permeability and/or local haemodynamic factors. Indeed, transcapillary escape of albumin (TERalb), a measure of systemic capillary efflux, is increased in diabetic patients, even in those with a modest increase of albuminuria. In normo-albuminuric type 1 diabetic patients, systemic capillary and glomerular flow is increased. We hypothesized that these haemodynamic changes contribute to an elevated TERalb, even in the phase preceding micro-albuminuria. METHODS: We measured TERalb in 39 normo-albuminuric type 1 diabetic patients and 46 healthy controls. TERalb was calculated from the disappearance curve of 125I-albumin. Renal and systemic haemodynamics were measured by standard techniques. Forearm blood flow (FBF) was measured by plethysmography. Endothelial function was assessed by intra-arterial infusion of acetylcholine. The structural integrity of the vessel wall was determined by the post-occlusive reactive hyperaemia test. RESULTS: TERalb was increased in diabetic patients (5.53+/-0.40 versus 4.39+/-0.21 %/h, P = 0.01). Patients were divided into tertiles with respect to their TERalb. There were no differences in UAE, blood pressure, metabolic parameters, endothelial function or maximal vasodilatation after occlusion between the groups. However, filtration fraction and FBF were significantly increased in the group of diabetic patients with the highest levels of TERalb. Overall, in diabetic patients, FBF was significantly correlated with TERalb. CONCLUSIONS: TERalb is increased in normo-albuminuric type 1 diabetic patients. In these patients with an increased capillary permeability, there is no evidence of endothelial dysfunction or vessel wall damage. However, both FBF and filtration fraction are increased. Therefore, the increased vascular permeability in the early phase of type 1 diabetes is associated with general haemodynamic alterations. Notably, such an increase in vascular permeability is not necessarily reflected by abnormal UAE. This could be due to either a lack of change in glomerular permeability or due to the fact that the threshold for tubular reabsorption of albumin has not been exceeded. PMID- 10703890 TI - Prospective comparison of therapeutical attitudes in hypertensive type 2 diabetic patients uncontrolled on monotherapy. A randomized trial: the EDICTA study. AB - OBJECTIVE: To compare the anti-hypertensive effect of combination therapy versus a single drug regimen schedule (dose-titration or switching to a different drug class) in type 2 diabetic hypertensive patients with inadequate blood pressure (BP) control on monotherapy. DESIGN: Prospective, randomized, open-fashion, parallel study of two therapeutic strategies during an 8-week period. SETTING: Primary care centers in Spain. PARTICIPANTS: A total of 898 men and women with type 2 diabetes mellitus and hypertension, receiving antihypertensive treatment with one single drug and whose BP was > 140 and/or 90 mmHg. INTERVENTION: Patients were randomized to a fixed combination therapy (verapamil 180 mg plus trandolapril 2 mg; Knoll AG, Ludwigshafen, Germany) or continued on a single drug regimen, either increasing the dose of the current drug or switching to a different drug class. MAIN OUTCOME MEASURE: Absolute BP reduction in the two groups of treatment, and the percentage of normalized patients (< 140/90 mmHg) in each group. RESULTS: The diastolic BP (DBP) decrease (5.6 mmHg) was significantly greater in patients treated with combination therapy, compared to patients on monotherapy (2.9 mmHg). The decrease in systolic BP (SBP) was not significantly different (11.1 versus 10.0 mmHg). In addition, a significantly higher number of patients treated with combination therapy (82% versus 74%) reached diastolic BP normalization (< 90 mmHg). CONCLUSIONS: In type 2 hypertensive patients with uncontrolled BP despite anti-hypertensive monotherapy, the change to combination therapy was more effective in attaining DBP control than any monotherapy schedule (either increasing the dose or switching to another different drug class). PMID- 10703891 TI - Control of glomerular hyperfiltration and renal hypertrophy by an angiotensin converting enzyme inhibitor prevents the progression of renal damage in hypertensive diabetic rats. AB - OBJECTIVE: Glomerular hyperfiltration and renal hypertrophy are both considered important in the progression of diabetic nephropathy. The aim of this study was to compare the effects of an equivalent reduction in blood pressure produced by the angiotensin-converting enzyme (ACE) inhibitor spirapril (SPI) and an antihypertensive triple drug combination of hydralazine, reserpine and hydrochlorothiazide (HRH) on kidney function, proteinuria and renal structure in hypertensive diabetic rats. DESIGN AND METHODS: Four groups of animals were evaluated in short-term and long-term studies. In both studies one group served as a non-diabetic hypertensive control (H). The other three groups were rendered diabetic and were allocated to one of the following groups: the first diabetic group received no specific therapy (HD), the second diabetic group was treated with SPI (HD-SPI) and the third diabetic group was treated with HRH (HD-HRH). In each of the two studies the systolic blood pressure (SBP), 24 h urinary total protein, glomerular filtration rate (GFR), glomerular area, proximal tubular area and glomerular sclerosis were evaluated. RESULTS: The blood pressure reduction was equal in rats receiving either SPI or HRH. The GFR, proteinuria, glomerular area and tubular area were significantly increased in the HD group, both in the short-term and the long-term study. In the HD-SPI group the diabetic hyperfiltration and renal hypertrophy responses were prevented. In the HD-HRH group the GFR and proteinuria were slightly reduced in the later phases of diabetes, while the glomerular area and tubular area were not affected. Semiquantitative analysis of renal lesions showed that SPI was more effective than HRH in the prevention of the development of glomerulosclerosis. CONCLUSIONS: The results of this study suggest that the control of early adaptive hyperfiltration and renal hypertrophy by SPI may be relevant in the prevention of glomerulosclerosis. PMID- 10703892 TI - Low-density lipoprotein-cholesterol determines vascular responsiveness to angiotensin II in normocholesterolaemic humans. AB - OBJECTIVE: Both LDL-cholesterol and angiotensin II have been shown to increase the risk for and severity of cardiovascular disease. In hypercholesterolaemia, experimental studies have demonstrated an increased angiotensin type 1 (AT1) receptor expression on vascular smooth muscle cells and an increased vascular responsiveness to vasopressors has been documented in humans. We investigated in a normocholesterolaemic young population whether vascular responsiveness to angiotensin II (Ang II) infusion depends on LDL-cholesterol serum levels in the systemic and renal circulation. DESIGN AND METHODS: Changes in systolic and diastolic blood pressure (deltaBP) to Ang II infusion (0.5 and 3.0 ng/kg per min) were investigated in 103 normocholesterolaemic (LDL-cholesterol < 160 mg/dl) young white men (26+/-3 years; 24 h BP: 128+/-10/75+/-7 mmHg) without cardiovascular disease. According to their LDL-cholesterol levels, participants were classified into tertiles (lower tertile < 85 mg/dl, middle tertile 85-111 mg/dl, upper tertile > 111 mg/dl). RESULTS: Blood pressure (BP) responses to Ang II infusion 3.0 ng/kg per min were enhanced in the group with the highest LDL cholesterol levels (delta systolic BP: +12.8+/-6.7, +13.2+/-8.6, +17.9+/-9.6, P < 0.02; delta diastolic BP: +11.1+/-5.8, +11.5+/-6.5, +16.5+/-8.3, P < 0.01, for the lower, middle and upper tertiles, respectively). This holds true when baseline BP was taken into account as a confounding covariable (P < 0.015). BP responses to Ang II infusion were related to LDL-cholesterol serum levels (delta systolic BP: r = 0.26, P = 0.01; delta diastolic BP: r = 0.32, P = 0.001). In multiple stepwise regression analysis, LDL-cholesterol emerged as the strongest determinant of vascular responsiveness to Ang II (delta systolic BP: P < 0.01; delta diastolic BP: P < 0.001). CONCLUSION: In young male subjects, responsiveness to Ang II is determined by the LDL-cholesterol serum level even in the normal range of LDL-cholesterol, thereby potentially contributing to the cardiovascular risk of LDL-cholesterol even within the so-called normal range. PMID- 10703893 TI - Cholesterol enhances contractile responses in isolated small mesenteric arteries of normotensive and spontaneously hypertensive rats. AB - OBJECTIVE: In order to examine possible mechanisms by which hypercholesterolemia may contribute to the development of cardiovascular disease, we investigated the effect of cholesterol enrichment on contractility in isolated small rat mesenteric arteries. DESIGN: Contractile responses of cholesterol-enriched isolated small mesenteric arteries of normotensive Wistar-Kyoto (WKY) rats and spontaneously hypertensive rats (SHR) were compared with control groups. METHODS: First- to second-order mesenteric arteries (327-349 microm internal lumen diameter) were dissected from the mesenteric bed of 10-20-week-old male WKY rats and SHR, and incubated in cholesterol-free and cholesterol-rich (150 microg/ml) medium. Isolated arteries were mounted on a Mulvany-Halpern myograph for measurement of isometric tension. RESULTS: Cholesterol significantly increased active wall tension and active wall pressure in WKY rat arteries and active wall tension in SHR arteries in response to potassium chloride, norepinephrine and serotonin (P < 0.05). In addition, contractile responses to all agonists were significantly higher in cholesterol-enriched SHR arteries compared with cholesterol-enriched WKY rat vessels (P < 0.05). CONCLUSIONS: These findings suggest that elevated cholesterol content enhances agonist-stimulated contractility in small mesenteric resistance arteries, providing a possible mechanism by which hypercholesterolemia may contribute to the development of hypertension. PMID- 10703894 TI - Contrasting blood pressure effects of obesity in leptin-deficient ob/ob mice and agouti yellow obese mice. AB - OBJECTIVE: Recent advances in understanding the neuroendocrine pathways regulating appetite, metabolism and body weight afford an opportunity to explore further the mechanisms by which obesity influences arterial pressure. ob/ob(Lep(ob)/Lep(ob)) mice have a mutation in the ob gene and are leptin deficient. Leptin possesses pressor actions and has been shown to increase arterial pressure when infused chronically or over-expressed transgenically. In contrast, agouti yellow obese(Ay) mice have overexpression of an agouti peptide that blocks melanocortin receptors. Stimulation of melanocortin receptors by alpha-melanocyte-stimulating hormone decreases arterial pressure. DESIGN AND METHODS: This study measured arterial pressure in leptin-deficient ob/ob mice, agouti yellow obese mice and their lean controls to test the hypothesis that the effects of obesity on arterial pressure are importantly influenced by the genetic and neuroendocrine mechanisms causing the obesity. We measured arterial pressure directly in conscious ob/ob mice (n = 14), agouti yellow obese mice (n = 6) and the same number of lean littermates. RESULTS: Body weight was nearly twice as high in ob/ob mice as in their lean controls, but mean arterial pressure was significantly lower in ob/ob mice (92+/-3 mmHg) compared with their lean controls (106+/-2 mmHg; P = 0.00017). In contrast, mean arterial pressure was significantly higher in agouti yellow obese mice (124+/-3 mmHg) than in their lean controls (99+/-1 mmHg; P = 0.000002) despite the fact that the agouti mice had milder obesity. CONCLUSIONS: This study prompts three conclusions: (1) leptin deficient ob/ob mice and agouti yellow obese mice have contrasting blood pressure responses to obesity, (2) obesity does not invariably increase arterial pressure in mice, and (3) the arterial pressure response to obesity may depend critically on the underlying genetic and neuroendocrine mechanisms. PMID- 10703895 TI - Reliability and limitations of echocardiographic measurement of left ventricular mass for risk stratification and follow-up in single patients: the RES trial. Working Group on Heart and Hypertension of the Italian Society of Hypertension. Reliability of M-mode Echocardiographic Studies. AB - OBJECTIVE: To investigate the clinical reliability of repeated measurements of left ventricular mass in a single patient. DESIGN: We used test-retest reliability analysis, within-class correlation and interval of agreement measures. METHODS: Two M-mode tracings (three consecutive cycles) were recorded in the same session and 3-10 days apart (5+/-2 days; mean +/- SD) in 261 participants (age 45+/-13 years, body mass index 24.7+/-3.6 kg/m2; 131 hypertensive and 130 normotensive; 50% of each group women) in 16 centres in Italy. The two tracings were read by two observers in each centre, after classification by a three-order quality score (1 = poor, 2 = sufficient, 3 = optimal). RESULTS: The average quality score was 2.11+/-0.71 (21% poor, 50% sufficient, 29% optimal). Left ventricular mass values ranged from 56 to 419 g (170+/-61 g). On the same day, within-observer 90% interval of agreement between tracing 1 and tracing 2 was -28 to +22 g (-17 to +11% of tracing 1). For day-to day test-retest within-observer variability (average three cycles), the 90% interval of agreement was -30 to +35 g (-18 to +18%). This variability decreased to -13 to +12% at the 80% interval of agreement and -12 to +11% at the 75% interval of agreement. The 90% interval of agreement of test- retest between observer variability was -26 to 30 g (-19 to +15%). A negligible regression toward the mean was identified. Categorical consistency of retest in the identification of hypertensive patients with left ventricular hypertrophy, classified in the first study, was 87% (k = 0.87). CONCLUSIONS: Measurement of left ventricular mass in single patients allows reliable risk stratification on the basis of the presence of left ventricular hypertrophy. The probability of a true change in left ventricular mass over time is maximized for a single-reader difference greater than 18% of the initial value, although differences of 10-13% might also have clinical relevance. PMID- 10703896 TI - Blood pressure-independent cardiac hypertrophy in acromegalic patients. AB - OBJECTIVE: Acromegaly is frequently associated with an increase in left ventricular mass, even in the absence of systemic hypertension. Pathological studies on acromegalic hearts have shown an extensive interstitial fibrosis, suggesting the existence of a specific acromegalic cardiomyopathy. The aim of this study was to assess left ventricular wall structure in acromegaly by ultrasonic tissue characterization. DESIGN AND METHODS: We studied 10 untreated acromegalic patients and 10 age-matched healthy control subjects. The echo patterns of two-dimensional long-axis end-diastolic echocardiograms were assessed by colour-scale analysis of the interventricular septum, with estimates of the mean colour scale value, the broad band (Bb) and the derived collagen volume fraction (dCVF). We also measured electrocardiographic QT interval dispersion (QTd) as a marker of dyshomogeneous ventricular repolarization. RESULTS: Seven patients had left ventricular hypertrophy according to the sex-independent criteria; of these, two had arterial hypertension. None of our patients had echocardiographic evidence of diastolic or systolic dysfunction. All patients showed significantly increased myocardial echoreflectivity (Bb = 106.4+/-12.1 versus 79.3+/-6.5; dCVF% = 2.78+/-0.53 versus 1.58+/-0.29; P < 0.0001) and QTd (66+/-13 ms versus 54+/-8 ms, P < 0.05). A significant correlation was found between dCVF and the duration of acromegaly (r = 0.80; P = 0.005). CONCLUSIONS: Left ventricular remodelling observed in acromegaly is not related to the presence of arterial hypertension; we hypothesize that the increased echoreflectivity and QTd are long-term consequences of cardiac hypertrophy and prolonged exposure to high levels of growth hormone and insulin-like growth factor-I. PMID- 10703897 TI - Regional and systemic haemodynamic response to aortography in hypertensives. AB - OBJECTIVE: To study the effects of aortography and of aortic counterflow bolus injection per se on regional and systemic haemodynamics in hypertensives in comparison to normotensive matched controls. DESIGN AND METHODS: Mean blood velocity (MBV) and pulsatility index (PI)--as an index of regional vascular resistance--by the Doppler technique, at the femoral, common carotid and brachial arteries, finger arterial pressure and electrocardiographic R-R' interval were monitored beat-by-beat, before, during and for 3 min following counterflow bolus injections into the abdominal aorta of 40 ml/2.6 s of iopamidol (I), iso-osmolar mannitol (M) and 0.9 N saline (S), in 11 hypertensive and nine normotensive patients. RESULTS: After bolus injection of iopamidol, MBV increased to a peak at 35+/-5 s, both in normotensive (deltaMBV versus baseline +16.7+/-9.9 cm/s; P < 0.01) and in hypertensive subjects (deltaMBV versus baseline: +13.9+/-6.6 cm/s; P < 0.01). At the same time, the PI decreased both in normotensive (deltaPI versus baseline: -4.05+/-2.49; P < 0.01) and in hypertensive subjects (deltaPI versus baseline: -3.02+/-2.25; P < 0.01). After M boluses, the haemodynamic changes were of the same direction and magnitude as I for both groups, while after S the magnitude was approximately 50% lower. No significant differences were observed between normotensive and hypertensive subjects. In other vascular circulations, a 15% increase of the early diastolic backflow in the brachial artery, in phase with the femoral artery haemodynamic changes, was the only evidence of the procedure. Mean arterial pressure decreased and heart rate increased in phase with flow changes of the femoral artery. CONCLUSIONS: (1) The regional flow and systemic pressure changes observed during aortography seem, at least partially, to be due to the hydrodynamic perturbation induced by bolus injection per se. (2) The physical and chemical properties of the contrast media and therefore the probable different shear-stress modifications induced by the fluid injected could explain why the haemodynamic changes were greater after I compared to S and were more similar to M. (3) Hypertensive subjects did not show a different vasoreactive response in comparison to normotensive subjects during aortography. PMID- 10703898 TI - Prognostic significance of exercise versus resting blood pressure in patients with chronic heart failure. AB - BACKGROUND: Results on the prognostic value of exercise blood pressure differ among studies; this may be related to the characteristics of the studied population. OBJECTIVE: To assess the prognostic significance of blood pressure measured during exercise in patients with chronic heart failure being considered for heart transplantation. DESIGN AND METHODS: Symptom-limited bicycle exercise testing with measurement of blood pressure and respiratory gas analysis was performed in 274 potential candidates for heart transplantation. They were then followed up for mortality and cardiovascular events. RESULTS: Results are given as the mean +/- SD. The age of the patients was 51.5+/-11.0 years, the resting blood pressure was 114+/-20/75+/-12 mmHg, the peak work load was 91+/-33 W and the peak oxygen uptake was 15.1+/-5.0 ml/min per kg. The systolic blood pressure increased to 128+/-21 mmHg at 30 W and to 133+/-23 mmHg at 50% of the peak work load. During the total follow-up time of 513 years, 55 patients died and 145 suffered at least one cardiovascular event. After controlling for age, gender and body mass index, mortality and incidence of events were inversely related to the systolic pressure at 30 W or at 50% of the peak work load, or to both (P < 0.05). The inverse associations of outcome with the systolic pressure at 50% of the peak work load persisted after additional adjustment for resting pressure and for peak oxygen uptake. CONCLUSION: The data indicate that a lower exercise systolic pressure, particularly at 50% of the peak work load, is associated with a higher mortality and a greater incidence of cardiovascular events. PMID- 10703899 TI - Effects of alacepril and amlodipine on the renal injury induced by a high cholesterol diet in rats. AB - BACKGROUND: A high-cholesterol (HC) diet increases blood pressure and induces renal injury in rats. We compared the effects of alacepril, an ACE inhibitor, and amlodipine, a Ca antagonist, on the renal injury induced by an HC diet in rats. DESIGN AND METHODS: Male Sprague-Dawley rats were given either an HC diet only (n = 5), an HC diet and amlodipine (n = 10) or an HC diet and alacepril (n = 10). The control rats (n = 5) were given a normal diet Systolic blood pressure (SBP) was measured by a tail-cuff method. Serum lipids, malondialdehyde (MDA) as a parameter for lipid peroxidation and urinary protein excretion were determined at 0, 4 and 8 weeks. The renal injury was evaluated histologically by the glomeruli sclerosing score. RESULTS: The HC diet increased SBP. Amlodipine lowered SBP more significantly than alacepril. Serum total cholesterol was increased by the HC diet and was not affected by either anti-hypertensive agent. HDL-cholesterol was similarly decreased in the three HC diet groups. Alacepril, but not amlodipine, completely attenuated the MDA elevation induced by the HC diet. Urinary protein excretion was decreased by the two anti-hypertensive agents at a similar rate. The renal histological injury assessed by the sclerosing score was ameliorated more significantly by alacepril than by amlodipine. CONCLUSIONS: Both amlodipine and alacepril decreased blood pressure and urinary protein, and ameliorated the renal injury induced by the HC diet in rats. The renal effect of alacepril seems to be mediated by the decrease in oxidative stress as well as by reduction of blood pressure, since alacepril lowered the sclerosing score more than amlodipine and completely attenuated MDA, although the blood pressure reduction by alacepril was less than that by amlodipine. PMID- 10703900 TI - Adenosine mediates nitric-oxide-independent renal vasodilation by activation of A2A receptors. AB - OBJECTIVE: Adenosine dilates rabbit renal arteries by an endothelium-dependent, nitric oxide (NO)- and prostaglandin-independent mechanism. The aim was to identify the responsible P1-purinoceptor subtype and to investigate the involvement of K+-channels. METHODS: Rabbit renal arteries were perfused with medium containing indomethacin (10 micromol/l). After preconstriction with noradrenaline (0.4 micromol/l), changes in vessel diameter by P1-purinoceptor agonists were measured with a photoelectric device. The P1-receptor subtype was characterised by selective antagonists. RESULTS: Adenosine caused concentration dependent dilation (EC50 approximately 7 micromol/l). The mRNA for A1, A2A and A3 receptors were demonstrated by reverse transcription-polymerase chain reaction from total RNA of renal arteries. The agonists CPCA (A2) and CGS21680 (A2A) dilated renal arteries (EC50 approximately 0.1 micromol/l), and CPA (A1) was ineffective. As demonstrated by experiments using two arteries in sequence, CPCA induced release of an endothelium-derived relaxing factor. NO synthase inhibition by NG-nitro-L-arginine methyl ester (L-NAME) had no effect on CPCA-induced dilation. The concentration-response curves of adenosine, CPCA and CGS21680 were shifted to the right by the A2A antagonist ZM241385 (1 micromol/l), but not by the A1 and A3 antagonists DPCPX (1 micromol/l) and MRS1220 (1 micromol/l). Iberiotoxin (0.1 micromol/l), a blocker of Ca2+-activated K+-channels, slightly shifted the dose- response curve of CPCA. Arteries preconstricted by KCl showed dilation to CPCA, but not to acetylcholine chloride (ACh). CONCLUSION: Adenosine induces dilation of rabbit renal arteries through activation of A2A receptors. This effect depends on the release of an endothelium-derived relaxing factor, which is not NO. Dilation by ACh in the presence of L-NAME is likely to be mediated by K+ as an endothelium-derived relaxing factor. However, in the A2A receptor-induced dilation of rabbit renal arteries, K+ does not play this role, suggesting the involvement of a further soluble factor in the receptor-induced dilatory function of the endothelium. PMID- 10703901 TI - Deficiency of renal dopaminergic-dependent natriuretic response to acute sodium load in black salt-sensitive subjects in contrast to salt-resistant subjects. AB - OBJECTIVE: To evaluate the involvement of the renal dopaminergic system in the natriuretic responses to acute saline load in salt-resistant (SR) and salt sensitive (SS) black normotensive (NT) and hypertensive (HT) subjects. DESIGN AND METHODS: We studied the relationship between the urinary excretion of dopa, dopamine (DA) and its metabolite DOPAC and the natriuretic responses to acute volume expansion (2 l NaCl 0.9% over 2 h) in 20 black NT subjects (12 SR and 8 SS) and 19 black HT subjects (10 SS and 9 SR). Subjects received a low salt (LS) diet (40 mmol sodium/day) for 1 week and a high salt (HS) diet (300 mmol sodium/day) for 1 week; the sequence of the dietary regimens was randomized. Comparisons were made between the results before the saline infusion (baseline) and the results 2 h after the infusion. RESULTS: In all the groups saline infusion induced significant increases in urinary volume (ml/4 h) of two- to three-fold and in urinary sodium excretion (mmol/4 h) of three- to ten-fold; these increases were significantly greater during the HS diet than during the LS diet. Saline infusion significantly increased the mean arterial pressure (MAP) by 5 mmHg in HT-SS subjects and by 4-5 mmHg in NT-SS subjects, but the MAP did not changed in the NT-SR and HT-SR groups. Under the LS diet, saline infusion changed the DA excretion (in nmol/4 h) by -49+/-89 in HT-SS subjects, by 17+/-52 in NT-SS subjects, by 235+/-72 in HT-SR subjects and by 220+/-86 in NT-SR subjects (P < 0.05 between SR and SS subjects). The saline infusion-induced changes in DA excretion correlated significantly with the increases in urinary sodium excretion (r = 0.71, P < 0.01) in the NT-SR and HT-SR subjects under the LS diet, but not in the SR groups on the HS diet nor in the SS groups (HT and NT) on either diet. Saline infusion significantly reduced the DA/dopa ratio in SS (NT and HT) but not SR (NT and HT) subjects, whereas the DA/DOPAC (dihydroxyphenylacetic acid) ratios were similar in all the groups. CONCLUSIONS: The urinary dopaminergic system may participate in the natriuretic responses to acute sodium load only in SR subjects (NT and HT) and only under LS diets, but not in SS subjects (NT and HT). This strongly suggests that black NT- and HT-SS subjects have an underlying impairment in the activity of the renal dopaminergic system which may be associated with a reduced decarboxylation of dopa into DA. PMID- 10703902 TI - More lessons from migraine. PMID- 10703903 TI - Homeopathic treatment of migraine: a double blind, placebo controlled trial of 68 patients [see comment]. AB - To evaluate the efficacy of homeopathy in preventing migraine attacks and accompanying symptoms, a randomised, double-blind, placebo-controlled clinical trial was conducted. There was a one-month registration period without treatment, followed by four months individualised homeopathic treatment or identical placebo. Patients were stratified for common or classical migraine. Seventy-three patients were randomised, 68 completed the trial. Baseline values were similar in the two groups. Both the homeopathy and placebo groups had reduction in attack frequency, pain intensity and drug consumption, with a statistically non significant difference favouring homeopathy. Migraine diaries showed no difference between groups. The neurologists' trial evaluation showed a statistically significant reduction in attack frequency in the homeopathy group (P= 0.04) and non-statistically significant trends in favour of homeopathy for pain intensity and overall evaluation. Further research, with improved trial design, on the possible role of homeopathy in migraine prophylaxis is justified. PMID- 10703904 TI - Efficacy of homeopathic treatment of skin reactions during radiotherapy for breast cancer: a randomised, double-blind clinical trial. AB - The aim of this study was to assess the effects of Belladonna 7cH and X-ray 15cH associated in the treatment of acute radiodermatitis. A randomized double-blind placebo-controlled clinical trial involving 66 patients who had been operated on for breast cancer and were undergoing radiotherapy was conducted. The following parameters were assessed over ten weeks: breast skin colour, warmth, swelling and pigmentation. The efficacy of the treatment was assessed by the comparison of these parameters taken individually and by calculating an Index of Total Severity (sum of the scores of the four parameters) during radiotherapy, and during recovery, 15 and 30 d after the end of the radiotherapy. The differences of the scores of the Index of Total Severity during Radiotherapy were not statistically significant, but showed a trend towards a better activity of the homoeopathic medicine compared to placebo. Analysis of the data on Total Severity during recovery, showed a statistically significant benefit of the active medicines over placebo. The homeopathic medicines had particular effectiveness on the heat of the skin. The limited number of patients observed and the posology employed could have interfered with the significance of the results. Chemotherapy and hormonotherapy do not seem to affect the results. PMID- 10703905 TI - Homeopathy and general practice: an urban perspective. AB - Complementary medicine appears to be an increasingly popular option amongst both doctors and patients. General practitioners (GPs) in more affluent parts of Britain have showed considerable interest in its use. Our objectives were to ascertain the use of and attitudes towards homeopathy amongst GPs working in a socio-economically deprived urban area such as Liverpool. A postal questionnaire survey was carried out of all general practice principals in Liverpool, using freepost envelopes and one reminder after three weeks. With respect to eight common complementary therapies in general and homeopathy in particular, respondents were asked whether they treat with, refer to or endorse each therapy; and their views were questioned on NHS funding, effectiveness, adverse reactions, training needs and theoretical validity, for each therapy. The response rate was 131/252 (52%), and was higher amongst women and doctors aged under 40 y. During the previous week 37 (28%) GPs had been involved in homeopathy with their patients: 6.5% had treated directly, 18.5% had referred to, and 7% had endorsed homeopathy. 31% of GPs reported successful outcomes by homeopathic treatment compared with 14% reporting adverse effects. Respondents were generally uncertain about the validity of the theoretical basis of homeopathy; only 23% considered it to have a valid basis. PMID- 10703906 TI - The Blackie Memorial Lecture 1999: homeopathy versus orthodoxy--the current state of play. PMID- 10703907 TI - The Doctrine of Signatures: a historical, philosophical, scientific view (II). AB - The first part of this article discussed the history of the Doctrine of Signatures. Bach developed a series of Flower Remedies for emotional states. On the basis of the Doctrine of Signatures, these can liberate the vital force from an alien disease process or 'archeus'. An understanding of the role of the active principle of a homeopathic medicine, in the physiology of its source, may aid understanding of a curative action. PMID- 10703908 TI - Idiopathic demyelinating neuropathy. PMID- 10703909 TI - Boy with hyperactivity and clumsiness. PMID- 10703910 TI - Carcinoma of breast and panic attacks. PMID- 10703911 TI - Baby with eczema and behaviour problems. PMID- 10703912 TI - The case of the ultimate pessimist--a moral tale showing how homeopathy treats patients discarded by conventional medicine as untreatable. PMID- 10703913 TI - Pony with skin allergy. PMID- 10703915 TI - Proposals for new international system of nomenclature for homeopathic medicines. PMID- 10703914 TI - 20 years ago: The British Homeopathic Journal, January 1980. PMID- 10703916 TI - Official recognition of homeopathy in Belgium. PMID- 10703917 TI - Was Kent a Hahnemannian? PMID- 10703918 TI - Interactions between homeopathy and drug treatment. PMID- 10703919 TI - Bone matters: are density increases necessary to reduce fracture risk? PMID- 10703920 TI - Insulin-like growth factor I production is essential for anabolic effects of thyroid hormone in osteoblasts. AB - Thyroid hormone (T3) and insulin-like growth factor I (IGF-I) are critical regulators of skeletal function. T3 increases IGF-I production in bone. To assess the potential role of IGF-I as a mediator of T3 actions, we characterized phenotypic markers of osteoblast activity in two osteoblast models, normal mouse osteoblasts and MC3T3-E1 cells, exposed to T3 alone or under conditions that interfere with IGF-I actions. T3 significantly increased osteoblast 3H-proline incorporation, alkaline phosphatase (ALP), and osteocalcin. Both alphaIR3, a neutralizing monoclonal antibody to the IGF-I receptor, and JB1, an IGF-I analogue antagonist, attenuated the stimulatory effects of T3. T3 effects also were decreased in cells transfected with antisense oligonucleotide (AS-ODN) to the IGF-I receptor gene. Both IGF-I and T3 had mitogenic effects that were inhibited by the antagonists. IGF-I by itself did not stimulate 3H-proline incorporation, ALP, and osteocalcin in the models used, revealing that although IGF-I is essential for the anabolic effects of T3, it acts in concert with other factors to elicit these phenotypic responses. PMID- 10703921 TI - A role for N-cadherin in the development of the differentiated osteoblastic phenotype. AB - Cadherins are a family of cell surface adhesion molecules that play an important role in tissue differentiation. A limited repertoire of cadherins has been identified in osteoblasts, and the role of these molecules in osteoblast function remains to be elucidated. We recently cloned an osteoblast-derived N-cadherin gene from a rat osteoblast complementary DNA library. After in situ hybridization of rat bone and immunohistochemistry of human osteophytes, N-cadherin expression was localized prominently in well-differentiated (lining) osteoblasts. Northern blot hybridization in primary cultures of fetal rat calvaria and in human SaOS-2 and rat ROS osteoblast-like cells showed a relationship between N-cadherin messenger RNA expression and cell-to-cell adhesion, morphological differentiation, and alkaline phosphatase and osteocalcin gene expression. Treatment with a synthetic peptide containing the His-Ala-Val (HAV) adhesion motif of N-cadherin significantly decreased bone nodule formation in primary cultures of fetal rat calvaria and inhibited cell-to-cell contact in rat osteoblastic TRAB-11 cells. HAV peptide also regulated the expression of specific genes such as alkaline phosphatase and the immediate early gene zif268 in SaOS-2 cells. Transient transfection of SaOS-2 cells with a dominant-negative N-cadherin mutant (NCADdeltaC) significantly inhibited their morphological differentiation. In addition, aggregation of NCTC cells derived from mouse connective tissue stably transfected with osteoblast-derived N-cadherin was inhibited by either treatment with HAV or transfection with NCADdeltaC. Together, these results strongly support a role for N-cadherin, in concert with other previously identified osteoblast cadherins, in the late stages of osteoblast differentiation. PMID- 10703922 TI - Functional gap junctions between osteocytic and osteoblastic cells. AB - Morphological evidence shows that osteocytes, bone cells that exist enclosed within bone matrix, are connected to one another and to surface osteoblasts via gap junctions; however, it is unknown whether these gap junctions are functional. Using a newly established murine osteocytic cell line MLO-Y4, we have examined functional gap junctional intercellular communication (GJIC) between osteocytic cells and between osteocytic and osteoblastic cells. In our hands, MLO-Y4 cells express phenotypic characteristics of osteocytic cells including a stellate morphology, low alkaline phosphatase activity, and increased osteocalcin messenger RNA (mRNA) compared with osteoblastic cells. Northern and Western blot analysis revealed that MLO-Y4 cells express abundant connexin 43 (Cx43) mRNA and protein, respectively. Lucifer yellow dye transferred from injected to adjacent cells suggesting that osteocytic cells were functionally coupled via gap junctions. Functional GJIC between osteocytic and osteoblastic (MC3T3-E1) cells was determined by monitoring the passage of calcein dye between the two cell types using a double labeling technique. The ability of bone cells to communicate a mechanical signal was assessed by mechanically deforming the cell membrane of single MLO-Y4 cells, cocultured with MC3T3-E1 cells. Deformation induced calcium signals in MLO-Y4 cells and those elicited in neighboring MC3T3-E1 cells were monitored with the calcium sensitive dye Fura-2. Our results suggest that osteocytic MLO-Y4 cells express functional gap junctions most likely composed of Cx43. Furthermore, osteocytic and osteoblastic cells are functionally coupled to one another via gap junctions as shown by the ability of calcein to pass between cells and the ability of cells to communicate a mechanically induced calcium response. PMID- 10703923 TI - Crucial involvement of the EP4 subtype of prostaglandin E receptor in osteoclast formation by proinflammatory cytokines and lipopolysaccharide. AB - Prostaglandin E2 (PGE2) exerts its effects through the PGE receptor that consists of four subtypes (EP1, EP2, EP3, and EP4). Osteoclast formation in the coculture of primary osteoblastic cells (POB) and bone marrow cells was enhanced more by 11 deoxy-PGE1 (an EP4 and EP2 agonist) than by butaprost (an EP2 agonist) and other agonists, which suggests that EP4 is the main factor in PGE2-induced osteoclast formation. PGE2-induced osteoclast formation was not observed in the coculture of POB from EP4-deficient (EP4 k/o) mice and spleen cells from wild-type (w/t) mice, whereas osteoclasts were formed in the coculture of POB from w/t mice and spleen cells from EP4-k/o mice. In situ hybridization (ISH) showed that EP4 messenger RNA (mRNA) was expressed on osteoblastic cells but not on multinucleated cells (MNCs) in w/t mice. These results indicate that PGE2 enhances osteoclast formation through its EP4 subtype on osteoblasts. Osteoclast formation by interleukin 1alpha (IL-1alpha), tumor necrosis factor alpha (TNF-alpha), basic fibroblast growth factor (bFGF), and lipopolysaccharide (LPS) was hardly observed in the coculture of POB and bone marrow cells, both from EP4-k/o mice, which shows the crucial involvement of PG and the EP4 subtype in osteoclast formation by these molecules. In contrast, osteoclast formation by 1,25-hydroxyvitamin D3 (1,25(OH)2D3) was not impaired and that by parathyroid hormone (PTH) was only partially impaired in EP4-k/o mice, which may be related to the fact that EP4-k/o mice revealed no gross skeletal abnormalities. Because it has been suggested that IL-1alpha, TNF-alpha, bFGF, and LPS are involved in inflammatory bone loss, our work can be expected to contribute to an understanding of the pathophysiology of these conditions. PMID- 10703924 TI - 1,25-Dihydroxyvitamin D3 hypersensitivity of osteoclast precursors from patients with Paget's disease. AB - Our previous studies suggested that increased osteoclast formation and activity in Paget's disease may be related in part to increased responsiveness of highly purified osteoclast precursors to 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3]. However, the basis for this enhanced sensitivity to 1,25-(OH)2D3 is unclear. To address this question, we examined 24-hydroxylase and 1,25-(OH)2D3 receptor (VDR) messenger RNA (mRNA) expression during human osteoclast differentiation from normal subjects and patients with Paget's disease in response to 1,25-(OH)2D3 as well as VDR content and affinity. Reverse-transcription polymerase chain reaction (RT-PCR) analysis of granulocyte-macrophage colony-forming unit (GM-CFU), the earliest identifiable osteoclast precursor, derived from patients with Paget's disease demonstrated 24-hydroxylase mRNA expression in response to 1,25-(OH)2D3 was induced at concentrations of 1,25-(OH)2D3 that were at least one log less than that required for normal GM-CFU. VDR mRNA and VDR protein were detected in both immature and more differentiated osteoclast precursors, as well as in osteoclast-like multinucleated cells (MNCs). However, VDR expression was lower in MNCs than the mononuclear precursor cells. Osteoclast precursors and MNCs from patients with Paget's disease had levels of VDR expression similar to those of normal subjects but showed increased VDR affinity for 1,25-(OH)2D3. Because the effects of 1,25-(OH)2D3 are in part mediated by induction of expression of RANK ligand on marrow stromal cells, which in turn stimulates osteoclast formation, we examined expression of RANK ligand mRNA by marrow stromal cell lines derived from patients with Paget's disease and normal subjects in response to 1,25-(OH)2D3. RT PCR analysis showed no difference in sensitivity of marrow stromal cells to 1,25 (OH)2D3 from normal subjects or patients with Paget's disease although the Paget's stromal cells expressed increased basal levels of RANK ligand mRNA. These results show that VDR protein is expressed in early and more differentiated osteoclast precursors, that expression levels of VDR decline with osteoclast differentiation, and that 1,25-(OH)2D3 has direct effects on osteoclast precursors. The enhanced sensitivity to 1,25-(OH)2D3 is an intrinsic property of osteoclast precursors from patients with Paget's disease that distinguishes them from normal osteoclast precursors. Furthermore, our results suggest that an increased affinity of VDR for 1,25-(OH)2D3 may be responsible for the enhanced 1,25-(OH)2D3 sensitivity of osteoclast precursors in patients with Paget's disease compared with normal subjects. PMID- 10703925 TI - Biological activity of CD-ring modified 1alpha,25-dihydroxyvitamin D analogues: C ring and five-membered D-ring analogues. AB - Nonsteroidal analogues of 1alpha,25(OH)2D3, lacking either the full five-membered D ring (C-ring analogues) or the full six-membered C ring (D-ring analogues) are more potent inhibitors of cell proliferation or inducers of cell differentiation than is 1alpha,25(OH)2D3. Maximal superagonistic activity was seen for the C-ring analogue with a 24(R)-hydroxyl group in the side chain [30- to 60-fold the activity of 1alpha,25(OH)2D3]. The 19-nor-16-ene-26,27-bishomo C-ring analogue showed the best ratio of antiproliferative to calcemic effects (1275-fold better than 1alpha,25(OH)2D3 and severalfold better than all vitamin D analogues so far described). The analogues are able to stimulate specific vitamin D-dependent genes and are active in transfection assays using an osteocalcin promoter VDRE. Low binding affinity to the vitamin D binding protein, differences in metabolism, or affinity for the vitamin D receptor (VDR) are not the most important explanations for the enhanced intrinsic activity. However, the analogues are able to induce conformational changes in the VDR, which makes the VDR-ligand complex more resistant against protease digestion than is 1alpha,25(OH)2D3. In contrast to 20-epimer steroidal vitamin D analogues, 20-epimer C-ring analogues were less potent than analogues with a natural C-20 configuration. In conclusion, several nonsteroidal vitamin D analogues are superagonists of 1alpha,25(OH)2D3 despite lower receptor affinity and, for the C-ring analogues, higher flexibility of the side chain; moreover, they have a better selectivity profile than all analogues yet published. PMID- 10703926 TI - Inhibition of chondrogenesis by parathyroid hormone in vivo during repair of full thickness defects of articular cartilage. AB - We studied the effects of parathyroid hormone (PTH) on differentiation of chondroprogenitor cells during the repair of full-thickness articular cartilage defects. Three-millimeter cylindrical full-thickness articular cartilage defects, which are small enough to be resurfaced spontaneously by hyaline cartilage, were created in the femoral trochlea of the rabbit knee. Recombinant human PTH(1-84) (hPTH[1-84]) (25 ng/h) then was administered into the joint cavity with an osmotic pump, or in control animals, saline alone was administered. The animals were killed at 1, 2, 4, and 8 weeks. At 1 week, the defects were filled with undifferentiated cells, regardless of the PTH treatments. By 8 weeks, well developed cartilage covered the defects with reconstitution of subchondral bone up to the original bone-articular cartilage junction. In contrast, no evidence of chondrogenic differentiation was seen at any time during the experimental period in the defects treated with PTH. The reparative tissues also were examined immunohistochemically using anti-proliferating cell nuclear antigen (PCNA) and anti-PTH/PTH-related peptide (PTHrP) receptor antibodies. Interestingly, the chondroprogenitor cells that filled the defects expressed PTH/PTHrP receptor, suggesting that these cells are capable of responding to PTH/PTHrP signaling before overt chondrogenesis. Application of PTH did not interfere with proliferation but inhibited chondrogenic differentiation of the cells resulting in the formation of fibrous tissue that lost the expression of PTH/PTHrP receptor within 4 weeks. PMID- 10703927 TI - Regulatory roles of zinc in matrix vesicle-mediated mineralization of growth plate cartilage. AB - Zinc (Zn2+) has long been known to play important roles in mineralization and ossification of skeletal tissues, but the mechanisms of Zn2+ action are not well understood. In this study we investigated the effects of Zn2+ on mineralization in a cell culture system in which terminal differentiation and mineralization of hypertrophic growth plate chondrocytes was induced by retinoic acid (RA) treatment. Addition of Zn2+ to RA-treated cultures decreased mineralization in a dose-dependent manner without affecting alkaline phosphatase (APase) activity. Characterization of matrix vesicles (MVs), particles that initiate the mineralization process, revealed that vesicles isolated from RA-treated and RA/Zn2+-treated cultures showed similar APase activity, but vesicles from RA/Zn2+ treated cultures contained significantly less Ca2+ and Pi. MVs isolated from RA treated cultures were able to take up Ca2+ and mineralize in vitro, whereas vesicles isolated from RA/Zn2+-treated cultures were not able to do so. Detergent treatment, which ruptures the MV membrane and exposes preformed intravesicular Ca2+-Pi-phospholipid complexes, did not restore the Ca2+ uptake abilities of MVs isolated from RA/Zn2+-treated cultures, suggesting that vesicles from RA/Zn2+ treated cultures did not contain functional Ca2+-Pi-phospholipid complexes. Zn2+ treatment did not affect the content of annexins II, V, and VI in MVs or the Ca2+ dependent, EDTA-reversible binding of these molecules to the membrane surface. However, Zn2+ treatment did affect the EDTA-nonreversible binding of these molecules to the MV membrane, suggesting that Zn2+ interferes with the assembly of annexins in the MV membrane. In addition, Zn2+ inhibited annexin II-, V-, and VI-mediated Ca2+ influx into liposomes. In conclusion, Zn2+ inhibits the mineralizing competence of intravesicular Ca2+-Pi-phospholipid complexes and function of annexin channels, thereby controlling Ca2+ influx into MVs, the formation of the first crystal phase inside the vesicles and initiation of mineralization. PMID- 10703928 TI - Bone structural and mechanical properties are affected by hypotransferrinemia but not by iron deficiency in mice. AB - Hypotransferrinemia is a genetic defect in mice resulting in <1% of normal plasma transferrin (Tf) concentrations; heterozygotes for this mutation (+/hpx) have low circulating Tf concentrations. We used this mutant mouse in conjunction with dietary iron deficiency to study the influence of Tf and iron on bone structural and mechanical properties. Twenty-one weanling wild-type BALB/cj +/+ mice and 21 weanling +/hpx mice were fed iron-deficient or iron-adequate diets for 8 weeks. Twelve hpx/hpx mice were fed the iron-adequate diet. Hypotransferrinemia resulted in increased tibia iron and calcium concentrations, lower femur failure load, and extrinsic stiffness. Because the femurs of the hpx/hpx mice were disproportionately small, these bones actually had increased tissue material properties (ultimate stress [US] and modulus of elasticity) than those of wild type mice. This is the first report on the effect of dietary iron deficiency on bone structural and mechanical properties. Dietary iron deficiency in +/+ and +/hpx mice decreased tibia iron concentrations but had no effect on tibia calcium and phosphorus concentrations or femur structural or mechanical properties. Because the bones of the hpx/hpx mice were small, but had superior tissue mechanical properties, we conclude that Tf is important for normal bone mineralization. PMID- 10703929 TI - Risk factors for vertebral deformities in men: relationship to number of vertebral deformities. European Vertebral Osteoporosis Study Group. AB - Recent epidemiological studies suggest a similar overall prevalence of vertebral deformity in men to that in women, though the influence of increasing age on the prevalence of vertebral deformity is less marked in men. However, most affected men have only a single or two vertebral deformities, which may be unrelated to osteoporosis. The aim of this study was to examine the role of risk factors, previously demonstrated to be associated with vertebral osteoporosis in females, in men with single/dual deformities compared to those with multiple deformities. Age stratified random samples of men aged 50 years and over were recruited from population registers in 30 European centers as part of the European Vertebral Osteoporosis Study (EVOS). Subjects had a lateral spinal radiograph and the presence of vertebral deformity was determined using the McCloskey algorithm. Lifestyle and other risk factor data were obtained from an interviewer administered questionnaire. In all 6937 men with a mean age of 64.4 (SD = 8.5) years were studied of whom 738 (10.6%) subjects had one or two deformities, and 109 (1.6%) subjects had three or more deformities. There was a marked increase in the prevalence of multiple vertebral deformities with increasing age, but only a modest effect of age on the prevalence of single deformities. Associations between various risk factors for osteoporosis and vertebral deformity were analyzed separately in men with single/dual vertebral deformity from those with three or more deformities using logistic regression. After adjustment for age, there were statistically significant associations between the following risk factors and multiple deformities: previous hip fracture (odds ratio [OR] 10.5), lack of regular physical activity (OR 2.9), low body mass (OR 2.5), and previous steroid use (OR 2.3). By contrast, there were only weak associations with these same variables in males with single/dual deformities and, apart from poor self reported general health, all of the 95% confidence intervals spanned unity. There was no difference in the reporting of very heavy levels of physical activity under the age of 50 years between men with single/dual deformities and those with multiple deformities. In conclusion, men with multiple deformities showed a similar pattern of risk factor association to those seen in women with vertebral deformity, in contrast to men with single/dual deformities. PMID- 10703930 TI - The mechanical properties of cancellous bone in the proximal tibia of ovariectomized rats. AB - The "mature rat model" is an effective and often-used surrogate for studying mechanisms and characteristics of estrogen-deficient osteopenia. The purpose of this study was to extend our understanding of this animal model to include the mechanical properties of cancellous bone in the proximal tibia. Female Sprague Dawley rats were divided into two groups (n=13 each) at 14 weeks of age: an ovariectomized group (OVX) and a sham-operated control group (sham). The study terminated after a duration of 5 weeks. Specimens 2 mm long were cut from the proximal tibial metaphysis just below the growth plate and tested using two methods: (1) "whole-slice" compression, in which the entire specimen is loaded between two larger flat platens and (2) "reduced-platen" compression (RPC), which uses platens sized and aligned to load only the cancellous bone in the center of the sample. Three-point bending tests also were conducted on the femur. The short duration of estrogen deficiency yielded only minimal differences (< 10%) in femoral cortical bone but dramatic reductions (approximately 60%) in cancellous bone properties as determined by the RPC method. Ultimate stress was 7.23 MPa +/- 1.97 MPa for OVX versus 18.1 MPa +/- 5.21 MPa for sham; and elastic modulus was 252 MPa +/- 104 MPa for OVX versus 603 MPa +/- 180 MPa for sham. These changes in mechanical properties are similar in many respects to the dramatic effects reported in histomorphometric studies. For the whole-slice method, differences in mechanical properties between the two groups were not as large because the test directly loads both cancellous and cortical bone, and the latter is not affected as severely by estrogen deficiency. In this case, ultimate stress and elastic modulus were only 30% (or less) lower for the OVX group. PMID- 10703931 TI - Growth hormone treatment in adults with adult-onset growth hormone deficiency increases iliac crest trabecular bone turnover: a 1-year, double-blind, randomized, placebo-controlled study. AB - The effects of growth hormone (GH) substitution on bone metabolism were evaluated by dynamic histomorphometry on iliac crest bone biopsies. Twenty-nine patients, aged 21-61 years (mean 45.5 years), with adult-onset GH deficiency (GHD) were randomized to receive subcutaneous injections with GH (2 IU/m2/day = 0.67 mg/m2/day) or placebo for 12 months. Serum insulin-like growth factor I (IGF-I) levels increased 263 +/- 98% (mean +/- SD) during GH treatment (p < 0.0001). In the GH group, osteoid surface increased during treatment from 11% (3-15%) (median [25-75 percentiles]) to 21% (10-27%) (p = 0.01) and mineralizing surface from 4% (1-8%) to 11%(7-16%) (p = 0.04). Moreover, erosion surface tended to increase in the GH group from 2% (1-3%) to 4% (3-5%) (p = 0.07). The quiescent surface decreased in the GH group from 87% (83-96%) to 74% (68-87%) (p = 0.01). The adjusted appositional rate, mineral apposition rate, bone formation rate, bone erosion rate, mineralization lag time, and osteoid thickness remained unchanged during treatment. Erosion depth showed a trend toward increase in the GH group (p = 0.09), whereas wall thickness was unchanged. Bone balance at the remodeling unit level and activation frequency were unchanged. At the tissue level, bone erosion rate increased significantly from 26% (17-36%)/year to 39% (23-72%)/year (p = 0.03). Similarly, the bone formation rate at the tissue level tended to increase, from 24% (15-31%)/year to 36% (17%-63%)%/year (p = 0.06). Finally, bone balance at the tissue level decreased significantly from 1% (-2-2%)/year to -5% ( 13-1%)/year (p = 0.01). No significant difference in change was seen in the cancellous bone volume. We conclude that 12 months of GH substitution therapy increases trabecular bone turnover. Moreover, our data suggest that bone balance at the bone multicellular unit level is not changed to positive. PMID- 10703932 TI - Is BMU-coupling a strain-regulated phenomenon? A finite element analysis. AB - Histologically, two types of bone reconstruction are distinguished: modeling and remodeling. Modeling changes the amount of bone and determines its geometrical form in relation to the prevailing mechanical loads and their resulting deformation (strain). Remodeling renews existing bone in a sequence of resorption and formation. However, in both processes the cells responsible for resorption and formation are the same: osteoclasts and osteoblasts. We studied if there is a relation between the activity of these cells and the deformation of the local bone tissue during remodeling. Two finite element models were built on a microscopic, supracellular level: (1) a secondary osteon in cortical bone and (2) a Howship's lacuna in a trabecula. Both models were loaded in the "natural," that is, longitudinal direction. Equivalent strains were determined as a measure for the deformation of the bone tissue. In the first model, the strain field around the osteon showed a region of decreased deformation in front of the tunnel, just where osteoclasts excavate cortical bone tissue. Behind the cutting cone, elevated strain levels appear in the tunnel wall at locations where osteoblasts are active. The second model showed that a local excavation of a loaded trabecula leads to higher strains at the bottom of the lacuna, where resorption is stopped and osteoblasts are recruited to refill the gap. However, in the direction of loading reduced strain levels appear, just where resorption continues to proceed along the trabecular surface. We conclude that at the tissue level, strain distributions occur during the remodeling process that show a relationship to the activity of osteoblasts and osteoclasts. This suggests that BMU coupling, that is, the subsequent activation of osteoclasts and osteoblasts during remodeling, is a strain-regulated phenomenon. PMID- 10703933 TI - Apolipoprotein E gene polymorphism and bone loss: estrogen status modifies the influence of apolipoprotein E on bone loss. AB - The identification of genes that contribute to bone mineral density (BMD) and bone loss has widespread implications for the understanding and prevention of osteoporosis. The objective of this study was to examine the relationship between the presence and absence of the apolipoprotein E*4 (APOE*4) allele and both BMD and annualized percentage rate of change in BMD at the lumbar spine and hip in a population of 392 healthy, pre-, peri-, and postmenopausal white women participating in the Women's Healthy Lifestyle Project. APOE genotype was analyzed by restriction enzyme analysis from genomic DNA. BMD at the lumbar spine and hip was measured at baseline and after a mean of 2.5 years using dual-energy X-ray absorptiometry (DXA). In premenopausal women, there were no significant differences in BMD or in the annualized percentage rate of change in BMD at the spine or hip when comparing women with and without the APOE*4 allele. In contrast, spine bone loss was significantly greater in peri- and postmenopausal women having an APOE*4 allele than in women without this allele (-1.75 + 1.5% per year vs. -0.98 +/- 1.4% per year, respectively, p = 0.018). Among peri- and postmenopausal women currently using hormone replacement therapy (HRT), there were no differences in the annualized percentage rate of change in spine BMD; whereas, among non-HRT users, there was a 2-fold higher rate of spine bone loss in women with an APOE*4 allele compared with women without this allele (-2.31 +/- 1.5% per year vs. -1.27 +/- 1.3% per year, respectively, p = 0.033; APOE*4 x HRT interaction, p = 0.076). In conclusion, this study shows the importance of APOE*4 allele in spine bone loss in peri- and postmenopausal women and, more importantly, it provides evidence for a genetic and lifestyle interaction in modulating spine bone loss. PMID- 10703934 TI - Early postmenopausal bone loss is associated with PvuII estrogen receptor gene polymorphism in Finnish women: effect of hormone replacement therapy. AB - Genetic factors regulate bone mineral density (BMD) and possibly the development of osteoporosis. An association between estrogen receptor (ER) polymorphism, BMD, and postmenopausal hormone replacement therapy (HRT) has not been established. Therefore, we studied the influence of the ER genotype on BMD before and after a 5-year HRT in a placebo-controlled, population-based, randomized group of 322 early postmenopausal women. The participants were randomized into two treatment groups: the HRT group (n = 145) received a sequential combination of 2 mg estradiol valerate and 1 mg CPA with or without vitamin D3, 100-300 IU + 500 mg calcium lactate/day (equal to 93 mg Ca2+), and the non-HRT group (n = 177) received calcium lactate, 500 mg alone or in combination with vitamin D3, 100-300 IU/day. PvuII restriction fragment length polymorphism (RFLP) of the ERalpha was determined using polymerase chain reaction (PCR). BMDs of the lumbar spine (L2-4) and proximal femur were measured by using dual-energy X-ray absorptiometry (DXA). At the baseline, there were no significant differences in the lumbar or femoral neck BMDs between the three ER PvuII genotype groups (PP, Pp, pp). After 5 years, the BMD of the femoral neck remained unaltered and that of the lumbar spine increased by 1.7% in the HRT group, whereas both BMDs were decreased by 4-5% in the non-HRT group. The ER genotype did not modulate the femoral neck BMD change during the follow-up. In contrast, in the non-HRT-group the lumbar spine BMD decreased more in subjects with the ER genotypes PP (6.4%) and Pp (5.2%) than in subjects with the pp genotype (2.9%) (p = 0.002). In the HRT group, the relative changes of the lumbar spine BMD were similar in all three ER genotype groups. Thus without HRT, the pp genotype was associated with a smaller decrease in the lumbar spine BMD than the Pp and PP genotypes. Long-term HRT seemed to eliminate the ER genotype-related differences in the BMD. We conclude that subjects with the ER PvuII genotypes PP and Pp may have a greater risk of relatively fast bone loss after menopause than those with the pp genotype and that they may preferentially derive benefit from HRT. PMID- 10703935 TI - Osteoporosis in elderly men and women: effects of dietary calcium, physical activity, and body mass index. AB - Dietary calcium intake and physical activity are considered practical measures for prevention of osteoporosis. However, their associations with bone mineral density (BMD) in the elderly are not clear. The present study examined the association between osteoporosis and these two factors in relation to body mass index (BMI) in a cross-sectional, epidemiological study involving 1075 women and 690 men, aged 69 +/- 6.7 years (mean +/- SD). Dietary calcium intake (median of 580 mg/day) was inversely related to age (p = 0.01), positively related to physical activity index (PAI) (p = 0.01), femoral neck BMD (p = 0.01) in women, and higher lumbar spine (p = 0.003) and femoral neck BMD (p = 0.03) in men. Quadriceps strength was negatively associated with age (p < 0.0001) and positively related to BMI (p < 0.0001) and BMD (p < 0.0001) in both men and women. The PAI was associated with quadriceps strength (p < 0.0001) and femoral neck and lumbar spine BMD in women (p < 0.001) and with femoral neck BMD in men (p = 0.04); however, these associations were not significant after adjusting for age, BMI, quadriceps strength, and dietary calcium. Women in the top tertile of quadriceps strength (> or =23 kg) and dietary calcium intake (> or =710 mg/day) had 15% higher BMD than those in the lowest tertiles (< or =15 kg and < or =465 mg/day); the difference was comparable in men (11%). Among subjects with the lowest tertiles of BMI (< or =23 kg/m2 for women and < or =24 kg/m2 for men), quadriceps strength (< or =15 kg for women and < or =28 kg for men), and dietary calcium intake (< or =465 mg/day), 64% and 40% of women and men, respectively, were classified as having osteoporosis (based on a 2.5-SD reduction from the young-normal mean). The prevalence was only 12% in women and 1.5% in men among those in the highest tertiles of the three factors. Adequate dietary calcium intake and maintaining a physically active lifestyle in late decades of life could potentially translate into a reduction in the risk of osteoporosis and hence improve the quality and perhaps quantity of life in the elderly population. PMID- 10703936 TI - Enhanced expression of osteocalcin mRNA in human osteoarthritic trabecular bone of the proximal femur is associated with decreased expression of interleukin-6 and interleukin-11 mRNA. AB - Few studies have investigated the factors or mechanisms that may lead to structural changes in OA bone. This study examines the in vivo expression of messenger RNA encoding the osteoclastogenic cytokines interleukin-6 (IL-6) and interleukin-11 (IL-11), together with the osteoblastic marker osteocalcin (OCN) and the calcitonin receptor (CTR), which in bone is exclusively expressed by osteoclasts. Total RNA was isolated from intertrochanteric trabecular bone from OA patients, and from controls taken at autopsy. The patterns of mRNA expression of IL-6, IL-11, OCN, and CTR were examined using reverse-transcription polymerase chain reaction (RT-PCR) by determining the relative ratios of the amplified products with respect to glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Both IL-6 and IL-11 mRNA were significantly less abundant in OA than in the control group. Expression of IL-11 mRNA decreased significantly with age for both groups. OCN mRNA expression was significantly more abundant in OA, and there was no significant difference for CTR mRNA between the two groups. For both OCN and CTR in OA, expression increased significantly with increasing age. These differences in expression between the OA and control groups are consistent with an hypothesis that biochemical and genetic factors in bone can contribute or perhaps underlie the degenerative joint changes seen in OA. PMID- 10703937 TI - Nitric oxide modulates fracture healing. AB - The role of the messenger molecule nitric oxide has not been evaluated in fracture healing. NO is synthesized by three kinds of nitric oxide synthase (NOS): inducible NOS (iNOS), endothelial (eNOS), and neuronal (bNOS). We evaluated the role of these enzymes in a rat femur fracture-healing model. There was no messenger RNA (mRNA) expression, immunoreactivity, or enzymatic activity for NOS in unfractured femoral cortex. After fracture, however, mRNA, protein, and enzymatic activity for iNOS were identified in the healing rat femoral fracture callus, with maximum activity on day 15. The mRNA expression for eNOS and bNOS was induced slightly later than for iNOS, consistent with a temporal increase in calcium-dependent NOS activity that gradually increased up to day 30. mRNA expression for the three NOS isoforms also was found in six of six human fracture callus samples. To study the effect of suppression of NO synthesis on fracture healing, an experimental group of rats was fed an NOS inhibitor, L nitroso-arginine methyl ester (L-NAME), and the control group was fed its inactive enantiomer, D-nitroso-arginine methyl ester (D-NAME). An 18% (p < or = 0.01) decrease in cross-sectional area and a 45% (p < or = 0.05) decrease in failure load were observed in the NOS-inhibited group on day 24 after fracture. Furthermore, the effect of NO supplementation to fracture healing was studied by delivering NO to the fracture site using carboxybutyl chitosan NONOate locally. On day 17 after fracture, there was a 30% (p < or = 0.05) increase in cross sectional area in the NO-donor group compared with the NOS inhibition group. These results show for the first time that NO is expressed during fracture healing in rats and in humans, that suppression of NOS impairs fracture healing, and that supplementation of NO can reverse the inhibition of healing produced by NOS inhibitors. PMID- 10703938 TI - Study of the nonresorptive phenotype of osteoclast-like cells from patients with malignant osteopetrosis: a new approach to investigating pathogenesis. AB - Osteopetrosis manifests as failure of osteoclastic bone resorption. The cause of the disease lies either in the hematopoietic lineage or in the bone marrow stromal microenvironment. It has not been possible to define the cell type involved in the various forms of the human disease because of the inability to form human osteoclasts in vitro. Using the recently described method for generating human osteoclasts from peripheral blood in coculture with rat osteoblastic UMR 106 cells, we demonstrate that a defect lies in the mature osteoclast-like cells in four cases of this disease. Control and osteopetrotic cocultures generated large numbers of osteoclast-like cells (calcitonin and vitronectin receptor positive, and F-actin ring-positive cells) with similar morphology. Bone resorption did not occur in three of the four osteopetrotic cultures. In case 1, in which bone resorption was identified, the area of resorption was negligible compared with the number of osteoclast-like cells in the culture and was detected only by scanning electron microscopy. In contrast, up to 20% of the bone surface in controls was resorbed. The normal and osteopetrotic osteoclast-like cells had a similar phenotype except that two of the osteopetrotic cases did not express CD44 and two expressed CD44 weakly, whereas CD44 was strongly expressed in the controls. This study shows that it is possible to reproduce in vitro the pathological features of human osteopetrosis, and the assay provides a means of acquiring a greater understanding of the pathogenesis of human osteopetrosis. PMID- 10703939 TI - Platelet-osteosarcoma cell interaction is mediated through a specific fibrinogen binding sequence located within the N-terminal domain of thrombospondin 1. AB - Approximately 20% of patients with osteosarcoma have metastatic disease in lungs or bones at diagnosis. The requirement of platelets in hematogenous dissemination of metastatic cells is now well established. Tumor cells interact with platelets and induce platelet aggregation. In this respect, metastatic potential of tumor cells correlates with their capacity to aggregate platelets in vitro. We have previously shown that thrombospondin 1 (TSP-1) is synthesized and expressed on the surface of MG-63 osteosarcoma cells and mediates platelet-osteosarcoma cell interaction. However, active sites mimicking the function of TSP-1 during platelet-osteosarcoma cell interaction are not known. In this study, a panel of antibodies directed against the N-terminal and C-terminal domains and type 1, type 2, and type 3 repeats of TSP-1 were first used to delineate the structural requirement for the binding of osteosarcoma cell surface-associated TSP-1 to platelets. A drastic inhibition of the platelet-aggregating activity of MG-63 cells was obtained in the presence of a monoclonal antibody directed against the N-terminal domain of TSP-1. Among a series of 16 synthetic peptides spanning the whole N-terminal domain of TSP-1, only synthetic peptide N12/I encompassing amino acid residues 151-164 of the N-terminal domain of TSP-1 inhibited the platelet aggregating activity of MG-63 cells. Electron microscopy studies showed that peptide N12/I strongly inhibited platelet-osteosarcoma cell interaction. A polyclonal antibody directed against peptide N12/I specifically bound to the surface of MG-63 cells, recognized TSP-1 and drastically inhibited the platelet aggregating activity of MG-63 cells. In addition, peptide N12/I specifically bound to fibrinogen and inhibited TSP-1/fibrinogen interaction. Overall, our results provide evidence that a fibrinogen-binding sequence located within the N terminal domain of TSP-1 mediates the binding of osteosarcoma cell surface associated TSP-1 to platelet-bound fibrinogen. PMID- 10703940 TI - Assessment of significant change in BMD: a new approach. PMID- 10703941 TI - Polycystic bone disease. PMID- 10703942 TI - The use of arsenic trioxide (As2O3) in the treatment of acute promyelocytic leukemia. AB - Arsenic trioxide (As2O3) has been shown to be even more effective than all-trans retinoid (ATRA) in the treatment of acute promyelocytic leukemia (APL). The combination of induction of apoptosis, induction of differentiation and inhibiting the proliferation, and killing of APL cells could be the main cellular mechanisms of As2O3 in APL treatment. As2O3 may affect APL cells by disturbing the activities of some important intracorporal enzymes, regulating the related gene expression and arresting the progression of cell cycle. PMID- 10703943 TI - The IFN-alpha-inducing capacity of Sendai Virus in human leukocytes is independent from the hemagglutinating and hemolytic activities and from the viral proteins of the Sendai Virus preparations. AB - About 30 Sendai Virus (SV) preparations, examined for their capacity to induce natural human interferon alpha from fresh human leukocytes (Le-IFN-alpha) of healthy donors, were characterized for hemagglutinating (HA) and hemolytic (HemA) activities and for SDS-PAGE proteic pattern. The SV preparations were produced by a single passage or by serial propagations through eggs in different conditions of multiplicity of infection (m.o.i.). The produced SV subpopulations showed variable IFN-inducing capacity, the values of which are distributed over a 6-fold range resembling a Gaussian distribution. No detectable difference was observed comparing the SV preparation obtained by serial propagations with those obtained by a single passage. The variability of the measured HA activity and HemA activity and as well as the SDS-PAGE proteic pattern of the SV preparations did not correlate with the induced amount of IFN per cell. In the same experimental conditions to produce Le-IFN-alpha, u.v.-treated SV preparations were unable to induce interferon depending on the u.v.-treatment. So it can be concluded that the distinct nHu-IFN-alpha-inducing capacity of SV subpopulation could be mainly associated with divergent compositions of the viral RNAs rather than with a different contents of viral proteins, among those detectable in SDS-PAGE and those responsible for HA activity and for HemA activity. PMID- 10703944 TI - Inhibition of CDK2, CDK4 and cyclin E and increased expression of p27Kip1 during treatment with interferon-alpha in carcinoid tumor cells. AB - IFN-alpha has presented antitumor effect in about 50% of carcinoid tumor patients, though the antitumor mechanism of IFN-alpha is still to be elucidated. In this study we demonstrated that IFN-alpha could result in accumulation of S phase population and upregulation of cyclin-dependent kinase inhibitor (CKI), p27. Moreover, IFN-alpha inhibits DNA synthesis assessed by [3H] thymidine incorporation and colony formation on soft agar. Immunodepletion of p27 increased CDK2- and cyclin E-associated kinase activities. These data suggest that IFN alpha exerts its antiproliferative effects in the neuroendocrine differentiated cell lines by: 1) inhibition of DNA synthesis and colony formation, 2) upregulation on the mRNA and protein expressions of the CKI, p27. PMID- 10703945 TI - Circulating dendritic cells in early and advanced cancer patients: diminished percent in the metastatic disease. AB - Despite the well demonstrated fundamental role of dendritic cells (DC) in generating antitumor immunity in experimental conditions, to date there are only few preliminary studies which investigate the percent of DC in the peripheral blood of cancer patients. Several cell surface markers have now been described which are specific to cultured DC, however their expression in vivo is still controversial. Recently, however, two DC subsets, consisting of immature and mature DC, have been shown to be present in peripheral blood, which can be recognized as CD123+ and CD11c+ cells, respectively. On this basis, we decided to investigate the presence of both mature and immature DC in the peripheral blood of early or advanced cancer patients. The study included 40 solid tumor patients, 18 of whom had a locally limited disease, while the other 22 showed distant organ metastases. CD123+ and CD11c+ cells were detected by FACS using monoclonal antibodies, and expressed as the percent of total leukocytes. The control group consisted of 50 healthy subjects. The mean percent of both CD123+ and CD11c+ cells was significantly lower in cancer patients than in controls. Moreover, the mean percent of both DC subsets was significantly lower in metastatic patients than in the non-metastatic ones. This study, demonstrating significantly lower percents of both immature and mature DC in the peripheral blood of cancer patients, particularly in those with distant organ metastases, suggests that DC deficiency may play a role in inducing cancer-related immunosuppression. Therefore, the demonstration of a diminished percent of DC in peripheral blood may represent a new interesting biological marker predicting a poor prognosis in human neoplasms, as with lymphocytopenia, the unfavourable prognostic significance of which has been well demonstrated. PMID- 10703946 TI - Anti-intercellular substance antibody log titres are correlated with serum concentrations of interleukin-6, interleukin-15 and tumor necrosis factor-alpha in patients with Pemphigus vulgaris relationships with peripheral blood neutrophil counts, disease severity and duration and patients' age. AB - Pemphigus vulgaris is a rare dermatosis of autoimmune origin, characterized by autoantibodies directed against intercellular substance (AICS) and presenting with intra-epidermal blisters and/or erosions of the skin and mucous membranes. The aim of this paper is to analyze the relationships between serum AICS titers (after log transformation) and: patients' age, disease duration and disease activity; serum cytokine (IL-6, IL-7, IL-15 and TNF-alpha) concentrations and peripheral blood cell counts (namely neutrophils, lymphocytes and natural killer cells). Fifteen consecutive subjects affected with PV were enrolled. Diagnosis was supported by histological examination as well as by direct and indirect immunofluorescence tests. Cytokine determinations were made by means of commercially available ELISA kits. This study shows for the first time that AICS titers have a significant correlation with age of PV patients (R=0.57, p=0.031) and with the disease duration (R=0.73, p=0.002). A correlation between blood neutrophils count and log (AICS) titres was observed (R=0.6, p=0.021). Furthermore, significant correlations were observed between log (AICS) titres and serum IL-15 (R=0.54, p=0.048), serum IL-6 (R=0.53, p=0.05) or serum TNF-alpha concentrations (R=0.53, p=0.05). These data, taken together, show that there are several connections between the log (AICS) titres, some proinflammatory cytokines, peripheral blood neutrophil counts and the numbers of individuals' lesions, suggesting a relationship between AICS production and lesion development. PMID- 10703947 TI - The CD nomenclature--a brief historical summary of the CD nomenclature, why it exists and how CDs are defined. PMID- 10703948 TI - CD35: complement receptor type 1. PMID- 10703949 TI - CD44. PMID- 10703950 TI - CD66a. PMID- 10703951 TI - CD66b. PMID- 10703952 TI - CD66c. PMID- 10703953 TI - CD66d. PMID- 10703954 TI - CD66e. PMID- 10703955 TI - CD66f. PMID- 10703956 TI - RP-HPLC tryptic mapping of IgG1 proteins with post-column fluorescence derivatization. AB - Peptide mapping is an important analytical technique widely used to study the primary structure of proteins. In quality control settings, it is employed as an identity test to probe for small changes in protein primary structure. A great challenge in peptide mapping is to minimize the detection limit for peptides due to the low detectability of smaller peptides based on their ultraviolet absorbance. The detection of peptide fragments can be enhanced by pre-or post column derivatization with fluorescent tags. The use of post-column o pthalaldehyde (OPA) and fluorescamine chemistries for on-line derivatization of peptide fragments from the RP-HPLC tryptic maps of several IgG1 monoclonal antibodies was explored. This paper describes the simple and sensitive peptide mapping technique for structural confirmation of proteins using picomoles of samples by post-column fluorescence derivatization. A comparison of UV and fluorescence detection of a peptide map is also presented. The method includes post column OPA derivatization of tryptic peptides from RP-HPLC tryptic maps with fluorescence detection. The conclusion reached that fluorescence detection gave relative detectability for tryptic peptides that range from 10- to 100-fold better than those observed with UV detection. The sensitivity of the peptide map increased by about 200-500 fold, i.e. peptide maps could be obtained using 2-5 pmol of digest instead of 1 nmol of digest. A roughly equal fluorescence response for all peptides (equal peak areas) was generally observed. PMID- 10703957 TI - Application of numerical taxonomy techniques to the choice of optimum mobile phase in high-performance thin-layer chromatography (HPTLC). AB - A total of eight solvent systems used for the separation of 1,4-benzodiazepine mixtures were investigated. The classification of solvent systems was carried out by the numerical taxonomy technique. The selection of optimum solvent system was accomplished on the basis of information quantity and objective function. The method was found to be a rapid and efficient tool in the choice of optimum system solvent. PMID- 10703958 TI - Ultrasensitive determination of beta-carotene in fish oil-based supplementary drugs by HPLC-TLS. AB - This research study demonstrates that thermal lens spectrometry (TLS) combined with efficient isocratic HPLC separation provides an excellent analytical means to profile and quantify carotenes in fish body oils and possibly also in vegetable oils. In particular, a highly sensitive, selective, simple and rapid method for the determination of ultratraces of beta-carotene in fish oil-based supplementary drugs has been proposed. The analyte could be determined reliably and precisely in the presence of a complex matrix including other fat-soluble vitamins (A, D and E), polyunsaturated fatty acids, sterols and other pigments by matrix-matched calibration in cod liver oil. The suitability of the method is also evidenced by favourable analytical figures of merit for beta-carotene such as a linearity range of 1-120 ng ml(-1), LOD of 0.58 ng ml(-1), recovery of 101.4+/-3.3% and measurement repeatability of 4.1%. PMID- 10703959 TI - Titrimetric determination of Cremophor EL in aqueous solutions and biofluids: part 2: ruggedness of the method with respect to biofluids. AB - A titration method for Cremophor EL, as a multicomponent mixture commonly used as non-ionic emulgent for manufacturing certain parenteralia, was developed for quantitative routine analysis in biofluids. A coated wire electrode is used as the end-point indicator in potentiometric titrations of Cremophor EL with sodium tetraphenylborate. The method tolerates a broad pH range, addition of alkanols and components of drug formulations and is sufficiently rugged. Reliable results are obtained at 20 degrees C. Disturbing ions from biofluid matrices can be masked or complexed by addition of formaldehyde, ethylenediaminetetraacetic acid and sodium fluoride. Sodium hydroxide is used for the required adjustment of the samples to pH 10. Cremophor EL spiked urine samples can be determined directly, whereas the true value of the emulgent content in the case of Cremophor EL spiked plasma samples is achieved by means of a conventional method. PMID- 10703960 TI - Stability, compatibility and plasticizer extraction of quinine injection added to infusion solutions and stored in polyvinyl chloride (PVC) containers. AB - The stability of quinine was determined in various diluents and in polyvinyl chloride (PVC) containers. The release of diethyhexyl phthalate (DEHP) from PVC bags into intravenous infusions of quinine was also measured. We used an injection of two doses of quinine; quiniforme at 500 mg and quinimax at 400 mg in either 250- or 500-ml PVC infusion bags containing 5% dextrose, to give initial nominal concentrations of 2 or 1 mg ml(-1) quiniforme and 1.6 or 0.8 mg ml(-1) quinimax, the mean concentrations commonly used in clinical practice. Samples were assayed by stability-indicating high-performance liquid chromatography (HPLC) and the clarity was determined visually. Experiments were conducted to determine whether the stability and compatibility of quinine would be compromised, and whether DEHP would be leached from PVC bags and PVC administration sets during storage and simulated infusion. There was no substantial loss of quiniforme and quinimax over 1- or 2-h simulated infusion irrespective of the diluent, and storage during 8 h at 22 degrees C, 48 or 72 h at 4 degrees C and 96 h at 45 degrees C. Leaching of DEHP was also detected during simulated infusion delivery using PVC bags and PVC administration sets. The quantity was less than 2 microg ml(-1). During storage at 4 degrees C and room temperature the leaching of DEHP was low, but when the temperature was 45 degrees C the quantity was high, 21 microg ml(-1). To minimise patient exposure to DEHP, quinine solutions with all drugs should be infused immediately or stored for a maximum of 48 h at 4 degrees C. PMID- 10703961 TI - Interference of magnetic resonance imaging contrast agents with the serum calcium measurement technique using colorimetric reagents. AB - A possible interaction between either linear (Gd-DTPA-BMA and Gd-DTPA) or macrocyclic (Gd-DOTA) gadolinium complexes used as magnetic resonance imaging (MRI) contrast agents and colorimetric technique reagents for the measurement of serum calcium was evaluated on human serum pools, and its mechanism was investigated by means of UV spectrometry and electro-spray ionization mass spectrometry (ESI-MS). The highest concentration tested was 2.5 mM (corresponding to a putative strictly intravascular distribution of the compound) and the lowest dose was 0.2 mM (i.e. about two elimination half lives). Serum calcium was dosed in duplicate by conventional colorimetric techniques involving o-cresol-phthalein complexone (OCP) or methylthymol blue (MTB) as reagents. No interference was detected when mixing Gd DOTA with serum, whatever the concentration. Gd DTPA (2.5 mM) did not interfere with the colorimetric technique either. Conversely, the Gd DTPA-BMA solution induced a concentration-related variation in apparent calcium levels. In the UV experiments, solutions of 2.5 mM MRI contrast media were mixed with OCP or MTB and UV absorption spectra were recorded between 400 and 800 nm. For Gd-DOTA/OCP and Gd-DOTA/MTB, no significant variations in the absorbance were detected. However, in the presence of Gd DTPA BMA, the absorbance of OCP and MTB showed substantial and immediate variations over time. The ESI-MS studies showed a complete displacement of Gd3+ ion in the case of Gd-DTPA BMA. In the presence of OCP, we observed the disappearance of Gd-DTPA BMA and the formation of the free ligand DTPA BMA and a new complex Gd OCP with an original stoichiometry of 2/2. Such a phenomenon did not occur in the case of Gd DOTA and Gd DTPA. The decomplexation of Gd-DTPA BMA in the presence of OCP can probably be explained by the weaker thermodynamic stability of Gd-DTPA BMA compared to that of Gd-DOTA and Gd DTPA. PMID- 10703962 TI - Validation of a rat pheochromocytoma (PC12)-based cell survival assay for determining biological potency of recombinant human nerve growth factor. AB - A method has been validated, according to the Guidelines of the International Conference on Harmonization (ICH), for precise quantitation of the biological activity of recombinant human nerve growth factor (rhNGF) for lot release testing. The assay is based on the survival of a subclone of rat pheochromocytoma PC12 cells (PC12-CF) in response to rhNGF. Cell survival is measured by monitoring the reduction, by living cells, of the alamarBlue dye into a red form which is highly fluorescent. The assay is simple, has high throughput (performed in 96-well microtiter plates) and shows reproducible dose-response curves in the concentration range of 0.2-50 ng/ml. The method was validated for its linearity, accuracy, precision, robustness, and to meet current regulatory requirements. The assay demonstrated good linearity, yielding a coefficient of determination of 0.9902. Sample recovery studies demonstrated an accuracy ranging from 96 to 98%. The repeatability of the assay and intermediate precision had coefficients of variation (CV) of <9%. The assay was stability-indicating since it was able to detect changes in rhNGF samples degraded by protease treatment and in a number of isolated rhNGF variants. Robustness was demonstrated by the relative insensitivity of the assay to small deliberate changes in key method parameters. The validation data, provided in this manuscript, indicate that the newly described bioassay for rhNGF is robust, accurate, precise, and suitable for lot release potency testing of rhNGF. PMID- 10703963 TI - Simultaneous determination of tinidazole, furazolidone and diloxanide furoate in a combined tablet preparation by second-derivative spectrophotometry. AB - A second-derivative spectrophotometric procedure has been developed for the simultaneous determination of tinidazole (TD), furazolidone (FD) and diloxanide furoate (DF) in a commercial preparation. The method consists of the utilization of second-derivative absorption spectra of tablet extract in distilled water and then determination of the analyte concentration in the mixture was carried out using zero-crossing (ZC) and ratio-compensation (RC) techniques. Calibration graphs constructed at their wavelengths of determination were linear in the concentration range of TD (5-20 microg ml(-1)), FD (2.5-10 microg ml(-1)) and DF (7.5-15 microg ml(-1)). The results were found to be accurate and free from interference. The details of the statistical treatment of analytical data are also presented. PMID- 10703964 TI - Fluorimetric determination of procaine in pharmaceutical preparations based on its reaction with fluorescamine. AB - A simple spectrofluorimetric analysis of a local anaesthetic named procaine using a specific labelling reagent for primary amino groups, has been developed. Because procaine shows very weak native fluorescence, the technique of spectrofluorimetry has been very much limited for its determination. A detail study of the variables affecting the derivatisation reaction (pH, fluorescamine (FC) concentration, temperature, reaction time), have minuciously been studied. The minimum detectable quantity is estimated as 7.7 ng ml(-1), with a relative standard deviation of 2.16% (ten determinations) for a procaine concentration of 100 ng ml(-1). The present method can be employed for the analysis of procaine by direct fluorescence measurements, without the interference from other compounds. The applicability of the present methodology have been demonstrated analysing three pharmaceutical preparations containing the analyte with satisfactory results. PMID- 10703965 TI - Molecular structures of metal complexes with mefenamic acid. AB - The infrared spectra of Na and Ca complexes of mefenamic acid were investigated in the region between 4000 and 400 cm(-1). These spectra were compared with X-ray powder diffraction patterns of complexes. It is shown that the proposed structure for these complexes obtained from the infrared spectra was supported by X-ray powder diffraction measurements. Bands due to v(as)(COO)- and v(s)(COO)- stretching vibrations appear at about 1580 cm(-1) and in the range 1390-1400 cm( 1) in the infrared spectra of the samples, respectively. The values of NH deformation vibrations do not change in the spectra of mefenamic acid and its metal complexes. On the other hand, molecular mechanic calculations and conformational analysis of three compounds were also established in the present work. As a result of these theoretical studies, the atomic planes and the peak assignments of the powder diffraction patterns were determined. As a result of these experimental and theoretical investigations, it may be concluded that metal atoms are connected to the carboxyl group of the mefenamic acid. PMID- 10703966 TI - Simultaneous determination of paracetamol, caffeine and acetylsalicylic acid by means of a FI ultraviolet pls multioptosensing device. AB - A simple and rapid analytical procedure is proposed for the simultaneous determination of caffeine (CF), acetylsalicylic acid (ASA) and paracetamol (PCT) in pharmaceutical preparations by partial least-squares (PLS) treatment of a flow through multisensor based on the integration of the retention and UV detection of the analytes on a solid support. Diode-array spectrophotometry has been used to obtain spectra (240-350 nm) of the analytes retained on C18 bonded phase beads packed in a flow cell. By using a 0.5% pH 1 HClO4 solution as the carrier, the multisensor responds linearly in the measuring range without requiring additional reagents or derivatization processes and the active microzone is regenerated by using methanol as eluting agent. Spectra of the corresponding analytes were used to provide multivariate data for the multivariate procedure. The statistical parameters obtained by the application of PLS methods at different reaction times were analysed, from which the optimum reaction time for the simultaneous determination of the analytes was selected. In the analysis of real and synthetic samples, precise and accurate values were obtained. PMID- 10703967 TI - Voltammetric, spectrofluorimetric and spectrophotometric methods to determine flufenamic acid. AB - Simple, rapid and sensitive voltammetric, spectrofluorimetric and spectrophotometric methods for determination of flufenamic acid (FF) in bulk powder and capsule dosage form are presented. The methods are based on the cyclisation reaction of FF with concentrated sulphuric acid to produce the corresponding acridone derivative. The voltammetric method is based on the adsorptive stripping differential pulse (DP) technique. The acridone derivative is determined over the concentration range of 8-60 ng ml(-1) using adsorptive preconcentration at the hanging mercury drop electrode (HMDE). The lower detection limit was found to be 1.02 ng ml(-1). The fluorimetric and spectrophotometric methods are based on the measurement of the fluorescence intensity at 450 nm (lambda(ex) = 400 nm)and peak-to-peak measurements of the first- (D1) and second-derivative (D2) curves, respectively. Beer's law is obeyed over the concentration ranges of 2-20 ng ml(-1) and 0.2-8.0 microg ml(-1) for the fluorimetric and spectrophotometric measurements, respectively. The three methods were proved to be accurate and reproducible as indicated by a relative standard deviation of <2%. PMID- 10703968 TI - Validation of a reversed-phase LC method for quantitative analysis of intravenous admixtures of ciprofloxacin and metronidazole. AB - The admixture ciprofloxacin-metronidazole is nowadays well known as a clinical tool when anaerobic organisms are involved. The stability of ciprofloxacin with metronidazole depends on several variables and this has been scarcely studied. Thus, the aim of this study was the search for a specific, precise and accurate method that would allow the quantification of ciprofloxacin in the presence of metronidazole. The reversed-phase liquid chromatography method is the analytical technique that has proved to be the most adequate for stability and compatibility studies of this mixture. This method was shown to be linear (r>0.999) in the range of concentrations of this study chosen, between 80 and 120% of the concentration of the commercial formulation. It showed precision with a repeatability (within-day) of 0.62% and reproducibility (between-day) of 1.14%, accuracy expressed as recovery percentage. PMID- 10703969 TI - An ELISA for quantification of murine IgG in rat plasma: application to the pharmacokinetic characterization of AP-3, a murine anti-glycoprotein IIIa monoclonal antibody, in the rat. AB - An enzyme-linked immunosorbent assay (ELISA) has been developed to determine concentrations of murine IgG in rat plasma. Specifically, the assay was developed to measure a murine anti-glycoprotein IIIa antibody (AP-3) in rat plasma to facilitate future investigations of AP-3 pharmacokinetics and pharmacodynamics in the rat. The working range of the assay is 15-100 ng ml(-1), corresponding to a limit of quantification of 1.5 microg ml(-1) in rat plasma. The assay was validated with respect to accuracy, precision, and cross-reactivity with both pooled rat and mouse IgG. Intra-assay recoveries of AP-3 in rat plasma ranged from 93 to 103%, with CV% values ranging from 5.2 to 8.5%. Inter-assay recoveries of the plasma AP-3 samples ranged from 107 to 119% with CV% values ranging from 17.7 to 25.1%. The assay has no appreciable cross reactivity with pooled rat IgG and full cross reactivity with pooled mouse IgG, making this an ideal assay to determine plasma pharmacokinetics of mouse antibodies in the rat. The assay was used to determine the pharmacokinetics of AP-3 in a Sprague-Dawley rat. PMID- 10703970 TI - Simultaneous determination of benazepril hydrochloride and hydrochlorothiazide by micro-bore liquid chromatography. AB - A micro-bore liquid chromatographic method was developed for the simultaneous determination of benazepril hydrochloride and hydrochlorothiazide in pharmaceutical dosage forms. The use of a BDS C-18 micro-bore analytical column, results in substantial reduction in solvent consumption and increased sensitivity. The mobile phase consisted of a mixture of 0.025 M sodium dihydrogen phosphate (pH 4.8) and acetonitrile (55:45, v/v), pumped at a flow rate of 0.40 ml min(-1). Detection was set at 250 nm using an ultraviolet detector. Calibration graphs are linear (r better than 0.9991, n = 5), in concentration range 5.0-20.0 microg ml(-1) for benazepril hydrochloride and 6.2-25.0 microg ml( 1) for hydrochlorothiazide. The intra- and interday R.S.D. values were <1.25% (n = 5), while the relative percentage error (Er) was <0.9% (n = 5). The detection limits attained according to IUPAC definition were 0.88 and 0.58 microg ml(-1) for benazepril hydrochloride and hydrochlorothiazide, respectively. The method was applied in the quality control of commercial tablets and content uniformity test and proved to be suitable for rapid and reliable quality control. PMID- 10703971 TI - Adaptation of solid phase extraction to an automated column switching method for online sample cleanup as the basis of a facile and sensitive high-performance liquid chromatographic assay for paclitaxel in human plasma. AB - An improved method for assaying paclitaxel in human plasma by high-performance liquid chromatography (HPLC) with UV detection at 227 nm has been developed by adapting previously reported sample preparation methods and chromatographic conditions to facilitate semi-automated sample cleanup using a column switching technique. Manual sample manipulations were limited to isolating the drug and internal standard from plasma (1.0 ml) by liquid-liquid extraction using tert butyl methyl ether. The sample extract was initially loaded onto a short cartridge column containing a cyanopropyl stationary phase. During the predetermined time interval that the drug and internal standard eluted from the cartridge, 1.50-2.20 min, a fully automated 6-position switching valve was used to direct the effluent onto an octylsilica analytical column. The same mobile phase, composed of acetonitrile-methanol-ammonium acetate buffer (pH 5.0; 20 mM) (76:19:105, v/v/v) and delivered at flow rate of 1.0 ml/min, was used for both separations. The overall retention times of paclitaxel and the internal standard were 10.9 and 18.1 min, respectively. The analytical method was thoroughly validated for quantitating paclitaxel in plasma at concentrations ranging from 6 to 586 nM (5-500 ng/ml). The lowest concentration of paclitaxel measured with acceptable day-to-day accuracy (100.2%) and precision (RSD 11.7%, n = 21, 5 months) was 6 nM (5 ng/ml). The sensitivity and selectivity of the assay proved to be more than adequate for monitoring steady-state plasma concentrations of the drug when administered to cancer patients as a 96 h continuous intravenous infusion in combination with other anticancer agents, such as doxorubicin and topotecan. Moreover, the heart-cutting procedure prevented the problematic introduction of interfering nonpolar plasma components onto the analytical column, thereby enhancing sample throughput while decreasing the technical demands of the assay. The method was found to be extremely reproducible and robust during extended use for the routine analysis of plasma specimens acquired from several clinical trials. PMID- 10703972 TI - Binding analysis of nilvadipine to plasma lipoproteins by capillary electrophoresis-frontal analysis. AB - Capillary electrophoresis coupled with frontal analysis (HPCE/FA) was applied to the ultramicro analysis of enantioselective binding of nilvadipine (NV), a calcium channel blocker, to plasma lipoproteins. The drug lipoprotein mixed solution was hydrodynamically introduced into a non-coated fused silica capillary for capillary electrophoresis. Since NV has no electric charge in the run buffer (pH 7.4), the unbound NV moved towards the cathodic end by electroosmotic flow, which was faster than the electrophoretic migrations of negatively charged lipoproteins and the bound NV. Once unbound NV migrated apart from lipoprotein, and bound NV was quickly released from the protein to maintain the binding equilibrium. Thus, NV migrated as a zone with a plateau region. The concentration of NV in this plateau region appearing on the electrophorogram was the same as the unbound NV concentration in the initial sample solution. It was found that the binding of NV to high-density lipoprotein (HDL), low-density lipoprotein (LDL) and oxidized LDL was non-specific and not enantioselective. Partition-like binding to the lipid part of these lipoproteins seemed to occur dominantly. The total binding affinities of NV to LDL were about seven times stronger than those to HDL, and the oxidation of LDL enhanced the binding affinity significantly. PMID- 10703973 TI - Liquid chromatographic determination of urinary 5-methyl-2'-deoxycytidine and pseudouridine as potential biological markers for leukaemia. AB - A simple reversed-phase liquid chromatographic (LC) method for the determination of urinary 5-methyl-2'-deoxycytidine (m5dCyd), recently claimed (on the basis of an imuno-technique) to be a potential marker for leukaemia, has been developed. Sample pre-treatment is based on a microcolumn clean-up step with an average recovery of 79% and a RSD of 3%. Detection limit was 0.2 microg/ml which is about tenfold lower than levels previously measured by an ELISA method in urine of healthy individuals. The creatinine (Cre) excretion, necessary for normalising the m5dCyd excretion, was evaluated by ion-pair liquid chromatography which permitted the simultaneous determination of pseudouridine (psi), a modified nucleoside also potentially useful as a marker for leukaemia. The described LC procedures were applied to the analysis of urine samples from healthy individuals and leukaemia patients. While the urinary psi/Cre ratio was found significantly increased for leukaemia patients, the urinary m5dCyd levels in healthy individuals were below the detection limits and did not increase in presence of the malignant disease. PMID- 10703974 TI - Mass spectrometric (HPLC/ESI--MS/MS) quantification of pyrimido[1,3-a]purin 10(3H)-one, a guanine adduct formed by reaction of malondialdehyde with DNA. AB - A high performance liquid chromatography/electrospray ionization tandem mass spectrometric (HPLC/ESI MS/MS) method has been developed for quantification of pyrimido[1,2-a]purin-10(3H)-one adducts from DNA. The method is based on acid catalyzed cleavage of the adducts from DNA and the use of [2,3a,10 13C3]pyrimido[1,2-a]purin-10(3H)-one as an internal standard in the analysis. For this purpose the latter compound was prepared. Rate constants for the acid catalyzed cleavage of pyrimido[1,2-a]purin-10(3H)-one from the corresponding 2' deoxyribonucleoside were determined, and its hydrolytic stability and possible formation by a cross reaction between guanine and [2,3a,10]pyrimido[1,2-a]purin 10(3H)-one were studied. PMID- 10703975 TI - Determination of ephedrine, theophylline and phenobarbital in a tablet dosage form by capillary electrophoresis. AB - A capillary electrophoresis method was developed to separate and quantitate ephedrine (ED), theophylline (TP) and phenobarbital (PB) in a tablet dosage form. Tablets were ground and extracted with methanol using ultrasonication. Aliquots of standard stock solution were hydrodynamically injected for 5 s at the anodic end. Separation was performed on a fused silica capillary (72 cm x 50 microm i.d.; 50 cm to detector) at an applied voltage of 20 kV with a phosphate run buffer (pH 8.0, 50 mM). Analysis was performed at ambient temperature (23+/-1 degrees C) and the total run time was 9 min with detection at 220 nm. Calibration curves were prepared for ED, TP and PB with methyl p-hydroxy benzoate as internal standard. For each analyte, the correlation coefficients were >0.999 (n = 4). The RSD% of ten replicate injections for each analyte were <1%. The method was applied to the quantitation of ED, TP and PB in a commercial tablet dosage form. PMID- 10703976 TI - Determination of norfloxacin spectrophotometrically using 2,4 dinitrofluorobenzene. PMID- 10703977 TI - Determination of fludarabine in a pharmaceutical formulation by LC. PMID- 10703978 TI - Anodic adsorptive stripping voltammetric determination of the anesthetic drug: methohexital sodium. AB - Methohexital (MS) determination is based on the formation of insoluble mercury salt on a hanging mercury drop electrode after preaccumulation by adsorption. This property was exploited in developing a highly sensitive stripping voltammetric procedure for the determination of the drug. The anodic current of adsorbed compound is measured by linear sweep anodic stripping voltammetry (LSASV), preceded by a period of preconcentration. The effect of various parameters such as supporting electrolyte composition, pH, initial potential, scan rate, accumulation time and ionic strength are discussed to characterize the interfacial and redox behavior. The detection limit was found to be 2x10(-7) M (56.8 ppb) with 180-s accumulation time. The interference of some amino acids, ascorbic acid and some metal ions was investigated. The application of this method was tested in the determination of methohexital in spiked urine samples. The precision of the method is satisfactory with a relative standard deviation of 2.5%. PMID- 10703979 TI - Use of experimental design to optimise a flow injection analysis assay for L-N monomethylarginine. AB - A flow injection analysis (FIA) method to determine L-N-monomethylarginine, based on the reaction with ortho-phthalaldehyde in the presence of a suitable thiol group, was optimised using experimental design. Two different approaches were followed wherein, (i) critical factors were identified in a screening design, and (ii) the simplex algorithm was used for further optimisation. In the first approach, the chemical reaction was optimised off-line and the optimal chemical conditions were transferred to the FIA-system. In the second approach the reaction and the FIA-system parameters were optimised together. The on-line approach is preferred. PMID- 10703980 TI - Simultaneous determination of benazepril hydrochloride and hydrochlorothiazide in tablets by second-order derivative spectrophotometry. AB - A second-order derivative spectrophotometric method for the simultaneous determination of benazepril hydrochloride and hydrochlorothiazide in pharmaceutical dosage forms is described. The determination of benazepril hydrochloride in the presence of hydrochlorothiazide was achieved by measuring the second-order derivative signals at 253.6 and 282.6 nm, while the second-order derivative signal at 282.6 nm was measured for the determination of hydrochlorothiazide. The linear dynamic ranges were 14.80-33.80 microg ml(-1) for benazepril hydrochloride and 18.50-42.20 microg ml(-1) for hydrochlorothiazide, the correlation coefficient for the calibration graphs were better than 0.9998, n = 5, the precision (%RSD) was better than 1.43% and the accuracy was satisfactory (Er < 0.99%). The detection limits were found to be 2.46 and 1.57 microg ml(-1) for benazepril hydrochloride and hydrochlorothiazide, respectively. The method was applied in the quality control of commercial tablets and proved to be suitable for rapid and reliable quality control. PMID- 10703981 TI - The multicomponent automated dissolution system: an alternative in the development and pharmaceutical analysis of generic polydrugs. AB - The necessity of assuring the quality of polydrugs, especially those with low aqueous solubility and in vivo absorption, has led to the development and evaluation of new techniques that can reduce the time and cost of analysis. This study examines the efficiency and accuracy of an automated dissolution system, fitted with an integrated multicomponent detector, for analysis of generic polydrugs using multiple linear regression (MLR). Trimethoprim and sulphamethoxazole were chosen as model drugs for this study and comparison was made with a conventional analysis based on HPLC. Both analytical systems under study gave reproducible and accurate results. Analysis of variance showed that there was no significant statistical difference between the methods of analysis, nor any statistical difference between the measured amounts of drug in the three different formulations. We have demonstrated that low cost instrumentation coupled with MLR data processing provides entirely satisfactory drug analysis to standard at least as good as that achieved using HPLC and provides an opportunity to reduce the time and analysis cost of other generic formulations. PMID- 10703982 TI - Preliminary study on the effect of miniaturisation and use of volatile mobile phases in LC for the on-line LC-MS analysis of basic pharmaceuticals. AB - To enhance to compatibility of the on-line coupling of liquid chromatography (LC) with mass spectrometry (MS) for the analysis of basic pharmaceuticals, the use of volatile mobile phase systems in combination with miniaturised LC was investigated. Multifactor analysis of variance (MANOVA) was used to evaluate the data obtained for the various variables (modifier, stationary phase, buffer, buffer pH and buffer concentration) on the resolution, peak symmetry and retention of four basic compounds analysed using LC columns with internal diameters (I.D.) of 0.3, 1.0 and 4.6 mm (conventional). Preliminary results obtained with the investigated micro and conventional columns showed similar behaviour with respect to ruggedness. The various investigated variables showed that miniaturisation by simply downscaling dimensions can result in varying selectivity and peak shapes for basic compounds. When comparing volatile mobile phases (containing ammonium acetate or ammonium citrate) and a conventional non volatile mobile phase (containing sodium phosphate) under pH 3 conditions, similar separation performances were observed. In the present study, ammonium citrate as the buffering salt, a high buffer concentration and methanol as the modifier showed the best peak symmetry. PMID- 10703983 TI - Comparison of derivative spectrophotometric and liquid chromatographic methods for the determination of omeprazole in aqueous solutions during stability studies. AB - A first derivative spectrophotometric method was developed for the determination of omeprazole in aqueous solutions during stability studies. The derivative procedure was based on the linear relationship between the omeprazole concentration and the first derivative amplitude at 313 nm. The first derivative spectra was developed between 200 and 400 nm (deltalambda = 8). This method was validated and compared with the official high-performance liquid chromatography (HPLC) method of the USP. It showed good linearity in the range of concentrations studied (10-30 microg ml(-1)), precision (repeatability and inter-day reproducibility), recovery and specificity in stability studies. It also seemed to be 2.59 times more sensitive than the HPLC method. These results allowed to consider this procedure as useful for the rapid analysis of omeprazole in stability studies since there was no interference with its decomposition products. PMID- 10703984 TI - Physico-chemical characterisation of the modifications I and II of (R,S) propranolol hydrochloride: solubility and dissolution studies. AB - The crystallisation conditions and the physicochemical properties of the modifications I and II of (R,S) propranolol hydrochloride were investigated. Detailed methods of preparation of the two forms were described. Data from FTIR spectroscopy, X-ray powder diffraction, thermal analysis, solubility and dissolution studies were used for the identification and the characterisation of the two forms. The forms I and II were easily differentiated by their IR spectra, X-ray patterns and thermal behaviour. The two polymorphs were found to be enantiotropically related to each other. Their stability was followed at room temperature over a period of 1 year and under different conditions of temperature, grinding and compression to verify the tendency to solid solid transition and to study the existence range of the two forms. The equilibrium solubilities of the two polymorphs in n-octanol were determined as well as their dissolution profiles as pellets in aqueous medium. These studies showed that form I, the less thermodynamically stable, was more soluble (by more than 34%) and dissolved faster than form II in agreement with the thermodynamic rules (A. Burger, R. Ramberger, Mikrochim. Acta II (1979) 259-271). PMID- 10703985 TI - Indirect determination of paracetamol in pharmaceutical formulations by inhibition of the system luminol-H2O2-Fe(CN)3-(6) chemiluminescence. AB - After a large drug scanning, the system Luminol-H2O2-Fe(CN)6(3-) is proposed for first time for the indirect determination of paracetamol. The method is based on the oxidation of paracetamol by hexacyanoferrate (III) and the subsequent inhibitory effect on the reaction between luminol and hydrogen peroxide. The procedure resulted in a linear calibration graph over the range 2.5-12.5 microg ml(-1) of paracetamol with a sample throughput of 87 samples h(-1). The influence of foreign compounds was studied and, the method was applied to determination of the drug in three different pharmaceutical formulations. PMID- 10703986 TI - The voltammetric behavior of nizatidine and its determination in biological fluids. AB - The voltammetric behavior of nizatidine (a newly introduced antiulcer drug) was studied using direct current (DCt), alternating current and differential pulse polarography (DPP). Well-defined cathodic waves were obtained over the whole pH range in Britton-Robinson buffers, in addition to 0.1 and 1 M HCl media. The main reduction wave was characterized as being irreversible and diffusion-controlled, although adsorption phenomena played a limited role in the electrode process. The current-concentration relationship was found to be rectilinear over the range 1x10(-5)-6x10(-4) and 2x10-6) -2x10(-4) M using DCt and DPP modes respectively, with a minimum detectability (S/N = 2) of 2x10(-7) M using the latter technique. The number of electrons involved in the reduction process was established, and the mechanism of electrode reaction was verified. The proposed method was successfully applied to determination of nizatidine in spiked human plasma and urine and the percentage recoveries were 96.12+/-0.40 and 97.12+/-0.17, respectively. PMID- 10703987 TI - LC determination of octreotide acetate in compound formulations of Sandostatin and diamorphine hydrochloride. AB - The determination of octreotide acetate in compound formulations of Sandostatin and diamorphine hydrochloride by RP-LC is described. Octreotide acetate, diamorphine hydrochloride and their respective degradants, [des-Thr-ol8] octreotide and 6-O-acetylmorphine, were baseline resolved using a Lichrospher-60 RP-select B column with a mobile phase composition of acetonitrile/phosphate buffer (pH 7.4, 20 mM) (35:65 v/v) with UV detection at 210 nm. The method is simple, selective, precise and suitable for the determination of octreotide acetate in admixture. PMID- 10703988 TI - Determination of barbiturates in urine by micellar liquid chromatography and direct injection of sample. AB - A liquid chromatographic procedure for the determination of six barbiturates (barbital, diallyl barbituric acid, phenobarbital, butabarbital, amobarbital and pentobarbital) in urine samples is described. The proposed system uses a Spherisorb octadecyl-silane ODS-2 C18 analytical column and a guard column of similar characteristics. The UV detector was set at 240 nm. A study to select adequate composition of the micellar mobile phase for the separation of these compounds in urine samples is performed. Maximum resolution was achieved with a 0.07 M sodium dodecylsulphate-0.3% propanol at pH 7.4 eluent. Limits of detection at 240 nm were ranged between 0.13 microg ml(-1) for diallyl barbituric acid and 2.7 microg ml(-1) for amobarbital. The procedure allows for the determination of these compounds in 20 minutes, it does not require prior a sample preparation step and it can be very useful to the investigation of intoxication. PMID- 10703989 TI - Applications of modulated differential scanning calorimetry in preformulation studies. AB - Characterization of the thermal properties of active pharmaceutical ingredients is critical in the selection of appropriate physical forms for development and defining proper manufacturing, handling and storage conditions of those chemical entities. Modulated differential scanning calorimetry (MDSC) has proven to be an effective tool in the thorough characterization of thermal behavior of compounds in preformulation studies. Selected applications of MDSC for various preclinical compounds are presented, thereby demonstrating the utility of this analytical method in the determination of glass transitions, characterization of desolvation and degradation processes as well as in the study of polymorphic transformations and crystallizations. PMID- 10703990 TI - Microbiological bioassay of erythromycin thiocyanate: optimisation and validation. AB - The validation of an analytical method for the quantitative determination of erythromycin thiocyanate formulated in an antibiotic preparation for veterinary use was carried out. This method is based on the microbiological method described in the European Pharmacopoeia to analyze erythromycin thiocyanate as a raw material. This erythromycin thiocyanate preparation is presented as a powder for oral administration after mixing with feed. For that reason, it was planned to validate the method for the quantitative determination of erythromycin thiocyanate incorporated both in the medicated premix and the mixture with feed. The microbiological method followed a linear model and was not proportional. The number of replicates needed to obtain a valid result was less than four in all cases. The small difference in concentration, expressed in natural logarithm detected by the method, was 0.1. PMID- 10703991 TI - Study on the charge-transfer reaction between 7,7,8,8-tetracyanoquinodimethane and drugs. AB - The charge-transfer (CT) reaction between 7,7,8,8-tetracyanoquinodimethane (TCNQ) as a pi-electron acceptor and cinnarizine, analgin, norfloxacin as electron donors have been studied by spectrophotometric method. The charge transfer complexes between TCNQ and these drugs have stable blue color, therefore a simple, rapid, accurate and sensitive method for determination of these drugs has been developed. The optimization of the experimental conditions is described. Beer's law is obeyed in the ranges 2-18, 2-18 and 4-32 microg/ml for cinnarizine, analgin and norfloxacin, respectively. The apparent molar absorptivity of CT complexes at 743 nm is 1.58x10(4), 1.71x10(4) and 8.91x10(3) l/mol per cm, respectively. The composition of all these CT complexes are found to be 1:1 by different methods. The relative SDs are less than 3% (n = 10). The proposed method has been applied to the determination of these drugs in their each pharmaceutical dosage forms with satisfactory results. PMID- 10703992 TI - Post-column reaction detection based on fluorescence energy transfer in the far red spectral region. AB - Post-column reaction detection may result in enhanced analytical sensitivity and selectivity. This paper describes an on-line HPLC with post-column fluorescence energy transfer assay using biotin as a model analyte. Biotin labeled with R phycoerythrin was used as the donor labeled ligand and streptavidin labeled with an indodicarbocyanine dye (Cy5), the acceptor labeled binder protein. The use of these labels provided a detection wavelength in the far red spectral region which is more selective for biological samples. In the on-line system, biotin was injected into the HPLC system followed by Cy5 labeled streptavidin and R phycoerythrin labeled biotin, post-column. The mixture was incubated on-line in an open tubular reactor coil maintained at 37 degrees C. The measured response was the sensitized emission of Cy5 due to fluorescence energy transfer from R phycoerythrin labeled biotin measured at 670 nm. Excitation was at 488 nm, which provided a large Stokes shift for reduction of scatter interference. The system was optimized with regard to the post-column reagents to obtain the minimum detectable concentration while maintaining appropriate dynamic range for the analysis of biotin. Biotin spiked in 0.01 M phosphate buffer, pH 7.4, showed a dynamic range of 304.0 pg/ml-122.20 ng/ml with a correlation coefficient of 0.993. The limit of detection for this assay was 304.0 pg/ml. The precision calculated at the blank (n = 6) was 4.14%. PMID- 10703993 TI - Validation of an automated liquid chromatographic method for omeprazole in human plasma using on-line solid-phase extraction. AB - An automated system using on-line solid-phase extraction and HPLC with UV detection has been validated in order to determine omeprazole in human plasma. The extraction was carried out using C18 cartridges. After washing, omeprazole was eluted from the cartridge with mobile phase onto an Inertsil ODS-2 column. The developed method was selective and linear for drug concentrations ranging between 5 and 500 ng ml(-1). The recovery of omeprazole ranged from 88.1 to 101.5%, and the limit of quantitation (LOQ) was 5 ng ml(-1). The intraday accuracy ranged from 93.1 to 106.2% and the interday accuracy varied from 95.4 to 105.1%. For the LOQ, good values of precision (8.7 and 17.5% for intraday and interday, respectively) were also obtained. This automated system has been applied to determine omeprazole in human plasma samples from bioequivalence studies. PMID- 10703994 TI - A comparative bioavailability study of different aspirin formulations using on line multidimensional chromatography. AB - A multi-dimensional column chromatographic method employing UV spectrometric detection was optimised and successfully used in a comparative bio-availability study of aspirin obtained from different commercially available oral dosage forms. Sample clean-up was achieved by on-line solid-phase extraction. In this study, the bioavailability of aspirin was compared in plain aspirin tablets, chewed tablets, effervescent tablets and Enteric-coated aspirin tablets. Blood samples were taken at frequent intervals after single dosing in ten healthy volunteers, the plasma samples were first treated with physostigmine sulphate to minimise enzymatic hydrolysis of aspirin to salicylate. The results showed the measured Tmax, Cmax, and AUC was significantly higher for soluble aspirin than for the other formulations and the t1/2 was shorter. This indicates the rapid absorption of aspirin from a soluble formulation compared with that from the other formulations. These differences suggest that the soluble formulation could be the aspirin of choice to treat patients suspected to be at high risk of myocardial infarction. The method performs, in a single step, an efficient extraction and clean-up of aspirin from human plasma. The calibration graph was linear over the calibration range 0.2-12 microg ml(-1) plasma with a limit of detection of 0.1 microg ml(-1). The intra- and inter-assay coefficients of variation were less than 6% and the recoveries ranged from 86 to 98%. The proposed method combines the advantages of being simple and selective in the presence of other potential interfering drugs and is suitable for routine analyses to obtain valuable information about the clinical effects of the drug and its use in prevention treatments of acute myocardial infarction. The whole procedure takes 7 min and is in agreement with other conventional methods. PMID- 10703995 TI - Photodegradation of inclusion complexes of isradipine with methyl-beta cyclodextrin. AB - The paper presents results of the studies on photochemical decomposition of isradipine (IS) and its liquid inclusion complexes with methyl-beta-cyclodextrin (M-betaCD). The process of photodegradation was assessed by the methods of UV spectrophotometry, HPLC (reverse-phase) and HPTLC (normal phase) chromatographic methods. The process of photodegradation of IS was analysed in the conditions of version I of the document of International Chemical Harmonization (ICH)-HBO-200 lamp. Quantitative evaluation of the photochemical decomposition was performed on the basis of the calculated photodegradation rate constant (k), half-life period (t0.5) and time of degradation of 10% of the compound (t0.1). Formation of inclusion complexes of IS with M-betaCD was proved to increase twice the photostability of the drug. The analytical methods used were subjected to a validation procedure in which the limits of detectability and determinability as well as specificity, precision and sensitivity of the method were determined. PMID- 10703996 TI - An isocratic LC method for the simultaneous determination of vitamins A, C, E and beta-carotene. AB - An isocratic liquid chromatographic method for the separation and simultaneous determination of retinyl acetate, propionate or palmitate (esters of vitamin A), beta-carotene (pro-vitamin A), ascorbic acid (vitamin C) and DL-alpha-tocopherol (vitamin E) is described. Samples are analysed by means of a reversed-phase column (LiChrospher 100 RP-18), using methanol as mobile phase. The UV Vis detector used was set at a wavelength of 300 nm and switched to 450 nm at 17 min, allowing the determination of beta-carotene. These vitamins were separated within 25 min and the detection limits ranged from 7 (beta-carotene) to 65 ng ml(-1) (ascorbic acid). PMID- 10703997 TI - Discrimination of herbal medicines according to geographical origin with near infrared reflectance spectroscopy and pattern recognition techniques. AB - Herbal medicines have an important role in clinical therapy in Asian countries such as Korea, Japan, and China. The objective of this study is to develop a nondestructive and accurate analytical method to discriminate herbal medicines according to geographical origin. Even though they are the same species, their qualities are different by growing conditions such as climate and soil. Near infrared (NIR) reflectance spectroscopy and a pattern recognition technique were applied for discrimination of herbal medicines according to geographical origin (Korea and China). Astragali Radix (AR), Ganoderma, and Smilacis Rhizoma (SR) were examined. It is shown that the representative NIR reflectance spectra in each group are different according to geographical origin after second derivatization to enhance spectral features. Also, the NIR reflectance spectra of Chinese and Korean samples were differentiated using principal component (PC) score plots. To establish the discrimination rule, Mahalanobis distance and discriminant analysis with PLS2 were utilized. PMID- 10703998 TI - Potentiometric sensors for the selective determination of sulbutiamine. AB - Five novel polyvinyl chloride (PVC) matrix membrane sensors for the selective determination of sulbutiamine (SBA) cation are described. These sensors are based on molybdate, tetraphenylborate, reineckate, phosphotun gestate and phosphomolybdate, as possible ion-pairing agents. These sensors display rapid near-Nernstian stable response over a relatively wide concentration range 1x10( 2)-1x10(-6) M of sulbutiamine, with calibration slopes 28 32.6 mV decade(-1) over a reasonable pH range 2-6. The proposed sensors proved to have a good selectivity for SBA over some inorganic and organic cations. The five potentiometric sensors were applied successfully in the determination of SBA in a pharmaceutical preparation (arcalion-200) using both direct potentiometry and potentiometric titration. Direct potentiometric determination of microgram quantities of SBA gave average recoveries of 99.4 and 99.3 with mean standard deviation of 0.7 and 0.3 for pure SBA and arcalion-200 formulation respectively. Potentiometric titration of milligram quantities of SBA gave average recoveries of 99.3 and 98.7% with mean standard deviation of 0.7 and 1.2 for pure SBA and arcalion-200 formulation, respectively. PMID- 10703999 TI - Optimisation of parameters for determination of rubidium in spent CAPD fluids by flame and electrothermal atomic absorption spectrometry. AB - An analytical procedure is reported for the determination of rubidium in spent continuous ambulatory peritoneal dialysis (CAPD) fluids by flame and electrothermal atomic absorption spectrometry (FAAS, ETAAS). Samples of spent CAPD fluids were collected as 5 ml aliquots in polyethylene tubes and stored in a freezer at -20 degrees C. Before analysis, samples were equilibrated to room temperature and analysed within 8 h. A total of 2 mg ml(-1) of caesium was added to each sample and standard solution to overcome interferences from ionisation. An air-acetylene flame was applied in FAAS determinations. Analysis was performed against aqueous standards. The calibration graph was linear from 30.0 up to 5000 microg l(-1) Rb, while the limit of detection (3 s) was found to be 20.0 microg l(-1) rubidium. Good repeatability of measurement (RSD 1%) was obtained. Parameters were also optimised for determination of rubidium in spent CAPD fluids by ETAAS. Ten-fold diluted samples (3.5% nitric acid) were analysed applying standard addition calibration. The calibration graph was linear from 2.0 up to 30.0 microg l(-1) rubidium, while the limit of detection (3 s) was found to be 1.0 microg l(-1) rubidium (sample volume 10 microl). Good repeatability of measurement (RSD 5%) was obtained. The results of direct determination by FAAS and ETAAS were compared to those obtained after acid digestion of samples in Parr bombs. The accuracy of the procedure for direct determination was checked by spiking samples. In 73% of samples analysed, the differences between the results obtained by the two techniques, either for direct determinations of samples or for samples digested in a Parr bomb did not exceed +/-10%. PMID- 10704000 TI - Application of derivative-differential UV spectrophotometry and ratio derivative spectrophotometric determination of mephenoxalone and acetaminophen in combined tablet preparation. AB - A method is presented for the direct determination of mephenoxalone and acetaminophen in combined pharmaceutical dosage forms without prior separation. The first method, derivative-differential spectrophotometry, comprised of measurement of the difference absorptivities derivatized in the first order (deltaD1) of a tablet extract in 0.1 N NaOH relative to that of an equimolar solution in methanol at wavelengths of 289.6 and 252.6 nm respectively. The second method is based on ratio derivative spectrophotometry. The amplitudes in the first derivative of the ratio spectra at 233.5 and 288.9 nm were selected to determine mephenoxalone and acetaminophen in the mixture. The proposed methods, which give thoroughly comparable data, are simple and rapid, and allow one to obtain precise and accurate results. PMID- 10704001 TI - Utilization of carbon disulphide for the analytical determination of betahistine hydrochloride and captopril in their pharmaceutical preparations. AB - Spectrophotometric, atomic absorption spectrometric and high performance liquid chromatographic (HPLC) procedures have been developed for the determination of betahistine hydrochloride and captopril. The three procedures are based on the reaction of the drugs with carbon disulphide in alkaline medium with the formation of the dithiocarbamate or the trithiocarbonate derivative of betahistine (BHT) and captopril (CAP), respectively, then subsequent chelation with divalent metals. The absorbance measurement of the formed chelates or of the metal moiety of chelates was used as the basis for the spectrophotometric and atomic absorption spectrometric determinations. The formed complexes have been used as pre-column derivatizing procedure for the HPLC determination of the two drugs. The different experimental conditions were optimized. The calibration graphs were linear over the applicable concentration ranges. The proposed procedures were applied successfully for the determination of the two investigated drugs in their tablets dosage form. PMID- 10704002 TI - Densitometric determination of impurities in drugs: Part IV. Determination of N (4-aminobenzoyl)-L-glutamic acid in preparations of folic acid. PMID- 10704003 TI - Extractional-spectrophotometric determination of famotidine in pharmaceutical formulations. PMID- 10704004 TI - Quantitative analysis of methocarbamol in solid dosage forms with 1H-NMR spectroscopy. PMID- 10704005 TI - Determination of vitamin D3 metabolites: state-of-the-art and trends. AB - The steps involved in the methods for the determination of vitamin D3 metabolites (namely, 25-hydroxyvitamin D3, 1,25-dihydroxyvitamin D3, 24,25-dihydroxyvitamin D3) mainly in clinical samples are critically reviewed. Sample pretreatment (e.g. deproteinization, saponification, liquid liquid and liquid solid extraction, etc.) as a function of both type of sample and detection system, quantitation based on protein saturation and liquid as well as gas chromatography are discussed. The chemical principles on which the methods are based and the derivatization procedures, which facilitate separation and/or detection, are also commented upon. Finally, the future prospects of the research on methods for the determination of these metabolites are outlined. PMID- 10704006 TI - The non-aqueous titrimetric assay of the selected anti-inflammatory agents using tetra-n-butylammonium hydroxide as titrant. AB - A potentiometric titration method in non-aqueous media is proposed for the determination of some commonly used anti-inflammatory agents. The direct potentiometric titration of three anti-inflammatory agents, namely mefenamic acid, fenbufen and ibuprofen; and the indirect potentiometric titration of diclofenac sodium was carried out in acetonitrile solvent using tetra-n butylammonium hydroxide as titrant, at 25 degrees C and under a nitrogen atmosphere. The method was found to be highly accurate and precise, having a relative standard deviation of <1.0% for all anti-inflammatory agents studied. Also, it was shown that the method could be successfully applied to the assay of commercial pharmaceuticals containing the above-mentioned anti-inflammatory agents. The validity of the method was tested by the recovery studies of standard addition to pharmaceuticals and the results were found to be satisfactory. The proposed method is simple, rapid and sufficiently precise for quality control purposes. PMID- 10704007 TI - Monitoring of a film coating process for tablets using near infrared reflectance spectrometry. AB - A process analytical chemical method using near infrared diffuse reflectance spectrometry was developed for the determination of the amount of tablet coating on single tablets. This method is based on calibration of the spectra versus the added mass of coating solution. The tablet core was composed of two halves of different chemical composition and spectra were recorded from both sides of the tablets. The calibration was carried out using the chemometric methods principal component analysis (PCA), partial least squares (PLS), and multiplicative signal correction (MSC). The PLS-model utilised spectra obtained from both sides, pretreated with MSC, and ordered into one object. This method can be used in process analytical chemistry at-line. Additional characterisation of the measurements was obtained by calibrating the spectra versus coating thicknesses obtained from optical microscopy. Using PCA, it was possible to roughly estimate the maximum depth in the coating material that returns chemical information, the 'information depth', which was 0.1-0.2 mm. PMID- 10704008 TI - Sample pooling to enhance throughput of brain penetration study. AB - The advent of combinatorial synthesis and high throughput screening in pharmaceutical research has inevitably given rise to a large number of interesting prelead compounds that requires rapid analytical throughput for kinetic characterization. The traditional approach of one-compound-at-a-time bioanalysis has not been able to meet the demand for high productivity of pharmacokinetic screening. This report demonstrates the application of sample pooling in expediting the pharmacokinetic screening, including assessment of brain penetration, of six NK1 receptor antagonists in rats: CAM 6108 (C1), CAM 6122 (C2), CAM 6178 (C3), CAM 5825 (C4), CAM 6182 (C5), and CAM 6121 (C6). The approach was adopted to avoid complications associated with cocktail dosing where multiple compounds are administered to one animal. The present investigation features individualized dosing (one compound per animal), followed by sample pooling of brain and plasma and bioanalysis via a conventional LC-fluorescence method. Rats were dosed intravenously with each of the six NK1 receptor antagonists and blood and brain samples were harvested at suitable post-dose time intervals. Plasma or brain homogenate samples from the same time points were pooled into two groups (C1-C3 and C4-C6) for assay. Drug compounds in plasma or brain were extracted by protein precipitation and quantitated using a validated gradient HPLC/fluorescence method, which was made feasible for both groups of compounds with a modification in gradient scheme. Plasma assay precision and accuracy for C1-C6 were < or =4.7% and within +/-9.8%, respectively. Brain homogenate assay accuracy for C1-C6 was within +/-7.0%. Brain penetration of these compounds was evaluated as the AUC of brain and plasma and their respective brain/plasma AUC ratio. The sample pooling approach helped to quickly identify C1 as the NK1 receptor antagonist with the greatest extent of brain penetration, followed by C2, C6, C4, C5, and C3 in that order. By employing sample pooling approach, pharmacokinetic parameters and brain penetration of all six compounds were obtained in a fraction of the time required by conventional single compound dosing and analysis. PMID- 10704009 TI - Determination of ABT-089 in human plasma by high performance liquid chromatography using in situ precolumn derivatization with 7-fluoro-4-nitrobenzo 2-oxa-1,3-diazole. AB - ABT-089 is a potent, selective neuronal cholinergic channel modulator with cognition enhancing activity in several animal paradigms. A simple and sensitive chromatographic method for the specific determination of ABT-089 in human plasma has been developed and validated. The method utilizes in situ precolumn fluorescence derivatization of the sample with 7-fluoro-4-nitrobenzo-2-oxa-1,3 diazole (NBD-F) prior to liquid-liquid extraction followed by reverse phase HPLC and fluorescence detection (lambda(ex) 495 nm, lambda(em) 533 nm). The described method significantly simplifies sample preparation. The derivatized product was separated from interference using a YMC ODS-AQ, 5 microm, 250x4.6 mm i.d. column using a mobile phase consisting of 30:5:65 (v:v:v) acetonitrile/methanol/aqueous buffer at a flow rate of 0.75 ml min(-1). The aqueous buffer consisted of 0.01 M tetramethylammonium perchlorate, 0.1% (v:v) trifluoroacetic acid, pH 3.0. An Alltech Absorbosphere CN, 5 microm, cartridge guard column was also used before the analytical column. Plasma samples were alkalinized with 0.1 M NaHCO3, 300 microl of a 1 mg ml(-1) ethanolic solution of NBD-F was added and the samples were heated in a water bath for 10 min at 50 degrees C. The samples were then extracted with tert-butylmethylether, evaporated to dryness and then reconstituted in mobile phase. For 1 ml of plasma, a limit of quantitation (LOQ) of 1.6 ng ml(-1) was obtained. The method was linear from 1.6 to 836 ng ml(-1). Inter and intra-day assay RSD (n = 6) were less than 9%. Accuracy determinations showed the quality control samples to range between 88-114% of the theoretical concentration. PMID- 10704010 TI - Determination of linezolid in plasma by reversed-phase high-performance liquid chromatography. AB - An HPLC-UV method was developed for assay of linezolid in dog, rat, mouse, and rabbit plasma. Linezolid and the internal standard were extracted on a solid phase cartridge (SPE) and separated on a reversed-phase column (C8, 4.6x150 mm, 5 microm) with 20% acetonitrile in water as mobile phase. The SPE quantitatively recovered linezolid and the internal standard from plasma samples. The chromatographic peak height ratio or peak area ratio based on UV absorbency at 251 nm was used for quantitative analysis. The assay procedures were simple and the assay was specific and had adequate precision and accuracy. Calibration standards with concentrations over the range of 0.01 20 microg/ml were validated for routine sample analysis to support the pharmacokinetic and toxicology studies with linezolid in dog, rat, mouse, and rabbit. Analysis of quality control samples showed the coefficients of variation were usually <10% and the measured and theoretical concentrations differed by <10% in most assays. Linezolid in the plasma samples was stable when stored at below -20 degrees C for at least 63 days, at room temperature (22-23 degrees C) for up to 24 h, and after three freeze-thaw cycles. This HPLC method has been successfully used in multiple laboratories to assay plasma samples from pharmacokinetic and toxicology studies with linezolid in the animal species. PMID- 10704011 TI - Application of LC-NMR and LC-MS to the identification of degradation products of a protease inhibitor in dosage formulations. AB - LC-NMR and LC-MS were applied to the characterization of six degradation products of a protease inhibitor, N-hydroxy-1,3-di-[4-ethoxybenzenesulphonyl]-5,5-dimethyl [1,3]c yclohexyldiazine-2-carboxamide, in a dosage formulation. A reversed-phase HPLC method was developed for the separation of the parent compound and its six degradation products. LC-MS was then utilized to obtain the molecular weight and fragmentation information using an electrospray ionization (ESI) interface in the positive ion mode. LC-NMR was employed to acquire detailed structural information using a selective solvent suppression pulse sequence in the stop flow mode. This work demonstrated the usefulness of this integrated approach for the rapid and unambiguous identification of drug compounds and their degradation products in dosage formulations. PMID- 10704012 TI - HPLC-UV method for the quantitation of nevirapine in biological matrices following solid phase extraction. AB - Nevirapine (VIRAMUNE) is a non-nucleoside reverse transcriptase inhibitor with activity against human immunodeficiency virus type 1 (HIV-1), currently marketed for the treatment of HIV-1 infected adults. A reverse phase HPLC-UV method was optimized and validated for the determination of nevirapine in human plasma, serum, milk and cerebrospinal fluid. The analyte was extracted from 250 microl of biofluid using a bonded silica solid phase extraction column, and resolved chromatographically on a reversed-phase, 15x0.46 cm i.d. 5 microm particle Supelco LC-8 analytical column with an isocratic mobile phase of 63% phosphate buffer (0.025 M, pH 6.0) with 1-butanesulfonic acid as anion-pair reagent: 21.5% methanol: 15.5% acetonitrile. The peaks were detected at a flow rate of 1.0 ml min(-1), at a wavelength of 280 nm, with a run time of 10 min. The assay was linear over a range of 25 to 10000 ng ml(-1). This method has been used for the clinical development of nevirapine. PMID- 10704013 TI - Determination of miconazole in pharmaceutical creams using internal standard and second derivative spectrophotometry. AB - A simple method is proposed for miconazole determination in pharmaceutical creams, based on extraction and second derivative spectrophotometry. In the presence of sodium lauryl sulfate (0.5%) and sulphuric acid (0.4 mol l(-1)), the miconazole and internal standard (IS) (methylene blue) were extracted to 100 microl of methylene chloride. The organic phase was evaporated in the nitrogen stream and the dry residue was dissolved in methanol (1.5 ml). The analytical signal was obtained as the ratio between second derivative absorbances measured at 236.9 nm (miconazole) and at 663.2 nm (IS). The use of IS in such multi-stage procedure enabled quite good analytical performance in calibration range 50.0 400 mg l(-1): linear correlation coefficient 0.9995, precision (measured as CV for ten replicates) at 50.0 mg l(-1) and at 400 mg l(-1) of miconazole was 1.5 and 0.5% respectively. Four commercial pharmaceutical creams were analyzed and the results obtained were in good agreement with the results obtained by reversed phase high performance liquid chromatography (HPLC). PMID- 10704014 TI - Determination of copolymer ratios of poly(lactide-co-glycolide) using near infrared spectroscopy. AB - The near infrared (NIR) spectroscopic technique was used to determine copolymer ratios of polylactide-co-glycolide samples. Appropriate quantities of DL polylactic acid and lactic-co-glycolic acid polymers with 86:14, 75:25, 64:36 and 52:48 lactide to glycolide ratios were dissolved in methylene chloride to obtain 5% (w/w) solutions. NIR spectra of the samples were obtained from the solutions using a Polyol Analyzer operated in the transmittance mode. Linear regression calibration models were generated at 2130 and 2288 nm from the second derivative spectral data obtained from the NIR technique. The lowest and highest standard errors of calibration (SEC) at 2130 nm were 1.29 and 1.63%, whereas those obtained from the calibration models generated at 2288 nm were 2.00 and 2.03%, respectively. Partial least squares (PLS) calibration models were also generated from the second derivative spectral data from 1100 to 2500 nm. The lowest and the highest SEC for the models were 1.46 and 1.53%, respectively. The calibration models were then used to predict the lactide contents of the unknown (test) samples. The highest percent error of prediction was 2.56% for samples with 86% lactide content when the linear regression calibration at 2130 nm was used, whereas the highest percent error of prediction was 1.56% for samples with 64% lactide content when the linear regression calibration at 2288 nm was used. The highest percent error of prediction was 1.73% for samples with 75% lactide content when the two-factor PLS calibration model was used. PMID- 10704015 TI - Monitoring in vitro experiments using microdialysis sampling on-line with mass spectrometry. AB - A method has been developed for the real-time analysis of components in in vitro reactions by the on-line combination of microdialysis sampling (MD) with tandem mass spectrometry (MS/MS) and single stage mass spectrometry (MS). Apparatus and parameters associated with the integration have been studied. Analytical figures of merit for the drug gepirone have been determined. The qualitative 'limit of identification' was found to be 100 ng/ml and 200 ng/ml for methods using thermospray and electrospray MS interfaces, respectively. Using this approach, monitoring of in vitro experiments involving drug metabolites, enzymatic reactions, and ligand-protein binding interactions were performed. PMID- 10704016 TI - Identification of losartan degradates in stressed tablets by LC-MS and LC-MS/MS. AB - Three unknown peaks were observed in the severely stressed losartan tablets (at 40 degrees C and 75% relative humidity for 3 years) analyzed by a stability indicating HPLC method. The sample solutions were subjected to liquid chromatography-tandem mass spectrometric (LC-MS/MS) analysis to obtain the chemical identities of these potential degradates. High accuracy and sensitivity of losartan and its degradates were obtained by using atmospheric pressure chemical ionization (APCI) technique in the LC-MS/MS analysis. Three trace level degradates (< or = 0.1%) were found to be the aldehyde and dimeric derivatives of losartan. The structural assignment was further confirmed by comparing the tandem mass spectra of the unknowns with those of the authentic materials of each corresponding degradate. Finally, the mechanistic pathways for the formation of the dimers are discussed. PMID- 10704017 TI - Feasibility of lovastatin analysis by packed column supercritical fluid chromatography with ultraviolet detection. AB - A reliable supercritical fluid chromatography (SFC) method was developed for the analysis of lovastatin, a hypocholesterolaemic drug, from MEVACOR. Methanol modified carbon dioxide was shown to elute the drug, and its dehydrolovastatin and hydroxy acid lovastatin degradation products from a Hypersil silica column. However, the hydroxy acid lovastatin was found to tail in this mobile phase. The phenomena was eliminated by the addition of trifluoroacetic acid [Haouck, S. Thomas, D. K. Ellison, Talanta 40 (1993) 491] to the mobile phase which permitted the drug and its two main degradation products to all elute from the Hypersil silica column in under 6 min with symmetrical peak shape. Chromatographic limit of detection (LOD) and limit of quantification (LOQ), linear dynamic range (LDR), and injection precision were obtained in order to assess the chromatographic performance of the SFC system for the lovastatin separation. PMID- 10704018 TI - Radioimmunoassay for a novel benzodiazepine inverse agonist, S-8510, in human plasma and urine. AB - A radioimmunoassay (RIA) was developed for a new benzodiazepine inverse agonist, 2-(isoxazol-3-yl)-3,6,7,9-tetrahydroimidazo [4,5-d] pyrano [4,3-b] pyridine monophosphate monohydrate (S-8510), in human plasma and urine. For competitive RIA, three amino derivatives of S-8510 were labelled by the Bolton and Hunter method and rabbit antisera were prepared using three immunogens, conjugates of three carboxyl derivatives of S-8510 with bovine serum albumin. All combinations of the labelled antigens and antisera were examined and homologous combinations were selected for the competitive RIA. One of the three homologous combinations had the best selectivity after investigations of cross-reactivity using 12 related compounds and was very sensitive for S-8510. Next, a pretreatment for biological samples was developed using mixed mode solid-phase extraction (SPE) column followed by the RIA (SPE/RIA). The assay recoveries for human plasma and urine were both excellent and the limits of quantitation were extremely low, 80 and 200 pg ml(-1), respectively. Human plasma samples and urine samples after administration of this drug were successfully measured by the SPE/RIA. No cross reactive metabolites were detected in any fractions after RP-HPLC separation of the plasma samples. The RIA using carefully selected antiserum and labelled antigen was highly specific for unchanged S-8510. To simplify the RIA procedure, a scintillation proximity assay (SPA) using the same labelled antigen and antiserum was developed for analyzing S-8510 in human plasma and found to be very promising. PMID- 10704019 TI - Determination of active ingredients in the pharmaceutical formulations containing hydrochlorothiazide and its binary mixtures with benazepril hydrochloride, triamterene and cilazapril by ratio spectra derivative spectrophotometry and vierordt's method. AB - Procedures were developed for the simultaneous determination of pharmaceuticals in binary mixtures, containing hydrochlorothiazide-benazepril hydrochloride, hydrochlorothiazide triamterene and hydrochlorothiazide cilazapril by ratio spectra derivative spectrophotometry and Vierordt's method. Mean recoveries, relative standard deviations and linearity ranges in calibration graphs of the methods were compared. These procedures were successfully applied to three pharmaceutical formulations for the determination of active ingredients. PMID- 10704020 TI - Simultaneous determination of alpha-fetoprotein, human chorionic gonadotropin and estriol in serum of pregnant women by time-resolved fluoroimmunoassay. AB - We have developed a simple and rapid time-resolved fluoroimmunoassay (TR-FIA) for simultaneous determination of alpha-fetoprotein (AFP), human chorionic gonadotropin (hCG) and estriol (E3) using europium and samarium ion chelate. In the proposed method, we used a combination of a 96-well microtiter plate for the AFP and hCG assay and transferable solid phase plate for the E3 assay. Therefore, these analytes could be measured simultaneously. The measurable ranges for AFP, hCG and E3 by the proposed method were 3.91-1000 ng ml(-1), 877-250000 IU l(-1) and 0.39 100 ng ml(-1), respectively. The proposed method which utilized characteristics of a rare earth ion chelate, was convenient (unnecessary diluting samples), quick (96 assays for 2 h), and required only a small quantity sample (50 microl). The principle of this proposed method is applicable to other antigens. PMID- 10704021 TI - Separation of morphine and its oxidation products by capillary zone electrophoresis. AB - Separation of morphine and its oxidation products (10-S-hydroxymorphine, pseudomorphine and morphine N-oxide) by capillary zone electrophoresis in Tris borate buffer in the presence of cyclodextrins was studied. Pyridoxine was used as an internal standard. PMID- 10704022 TI - Spectrophotometric and spectrofluorimetric determination of etodolac and aceclofenac. AB - Two simple, sensitive and reproducible spectrophotometric and spectrofluorimetric methods were adopted for the analysis of the anti-inflammatory drugs, etodolac and aceclofenac. The first method is based on the formation of coloured complexes between the drugs and p-dimethylaminobenzaldehyde reagent (PDAB) in the presence of sulfuric acid and ferric chloride. Measurement of the absorbances was carried out at 591.5 and 545.5 nm for etodolac and aceclofenac, respectively. Regression analysis of Beer's plots showed good correlation in the concentration ranges 10 80 and 8-55 microg ml(-1), respectively. The second was the spectrofluorimetric method in which samples of etodolac in ethanol showed native fluorescence at a lambda = 345 nm when excitation was at 235 nm and samples of aceclofenac in the phosphate buffer pH 8 showed native fluorescence at lambda = 355 nm when excitation was at 250 nm. The calibration graph was rectilinear from 96 to 640 ng ml(-1) for etodolac and from 2 to 8 microg ml(-1) for aceclofenac. The proposed methods are applied successfully for the determination of the two drugs in bulk powder with a mean accuracy of 100.48+/-0.85 and 100.03+/-0.38 in the PDAB method and of 100.61+/-0.79 and 99.88+/-0.45 in the spectrofluorimetric method. Applicability of the proposed methods was examined by analysing dosage forms of the investigated drugs. Recoveries were 98.77-101.46 and 98.65-102.10% for the two methods, respectively and RSD values were 0.6-0.7 and 0.35-1.06% respectively. PMID- 10704023 TI - Simultaneous GC-MS determination of nicotine and cotinine in plasma for the pharmacokinetic characterization of nicotine in rats. AB - A gas liquid chromatography/mass spectrometry assay method was developed for the simultaneous determination of nicotine and its major metabolite, cotinine, in rat plasma. Of particular interest was improving the low and variable extraction recovery for the parent drug and the metabolite. In addition, the feasibility of this assay method for pharmacokinetic studies of nicotine and cotinine after intravenous (i.v.), oral, and intraperitoneal (i.p.) administration of 1 mg kg( 1) of nicotine was tested. The low (30 and 48% for nicotine and cotinine, respectively) and variable (25 and 22% coefficient of variation for nicotine and cotinine, respectively) extraction recovery for nicotine and cotinine into dichloromethane was significantly improved by the addition of NaCl to the plasma. As a result, the recoveries for nicotine and cotinine were improved to 68 and 65%, respectively. The coefficient of variation was less than 10% in the 50-500 ng ml(-1) range and less than 16.58% at 10 ng ml(-1) for both nicotine and cotinine, indicating that the reproducibility of the assay was also improved by the extraction procedure. When injected intravenously at a dose of 1 mg kg(-1), the temporal profile of plasma concentration for nicotine followed a bi exponential decline. Moment analysis revealed that pharmacokinetic parameters for nicotine (i.e. Cl, 46.30 ml min(-1) kg(-1); Vss, 2.77 1 kg(-1)) was similar to those reported in studies using 14C-nicotine. Absolute bioavailabilities of nicotine for i.p. and oral administration were 87.0 and 80.4%, respectively. The concentration of the metabolite increased up to 4 h to reach Cmax after i.p. and oral administrations and then declined slowly with time. These results indicate that this convenient analytical procedure is readily applicable to pharmacokinetic studies of nicotine and cotinine involving small laboratory animals with a sensitivity comparable with that reported for studies using 14C nicotine. PMID- 10704024 TI - Assay of beta-hematin formation by malaria parasite. AB - Novel leads are urgently required for designing antimalarials due to the reduced efficacy of presently available drugs. The malaria parasite has a unique reaction of heme polymerization, which has attracted much attention in the recent past as a target for the design of antimalarial drugs. The process is hampered by non availability of a proper assay method. Currently available methods are cumbersome and require advanced instrumentation or radioactive substrates. Here, we are describing an assay for hemozoin formation that is simple and reproducible. This assay has routinely been used by us for the identification of potential compounds with antimalarial activity. PMID- 10704025 TI - Simultaneous determination of propafenone and 5-hydroxypropafenone enantiomers in plasma by chromatography on an amylose derived chiral stationary phase. AB - An enantioselective liquid chromatography method was developed for the simultaneous determination of propafenone (PPF) and 5-hydroxypropafenone (PPF 5OH) enantiomers in plasma. After liquid liquid extraction with dichloromethane, the enantiomers were resolved on a Chiralpak AD column using hexane-ethanol (88:12, v/v) plus 0.1% diethylamine as the mobile phase and monitored at 315 nm. Under these conditions the enantiomeric fractions of the drug and of its metabolite were analysed within 20 min. The extraction procedure resulted in absolute recoveries of 62.9 and 61.3% for (R)- and (S)-PPF, respectively, and of 57.6 and 56.5% for (R)- and (S)-PPF-5OH, respectively. This procedure was efficient in removing endogenous interferents as well the interference of an other PPF metabolite, N-despropylpropafenone (PPF-NOR). The calibration curves were linear over the concentration range 25-1250 ng/ml. Low values of the coefficients of variation were demonstrated for both within-day and between day assays. The method described in this paper allows the determination of PPF and PPF-5OH enantiomers at plasma levels as low as 25 ng/ml and can be used in clinical pharmacokinetic studies. PMID- 10704026 TI - Interlaboratory study comparing the microbiological potency of spiramycins I, II and III. AB - An interlaboratory study has been performed to determine the relative potencies of spiramycins (SPMs) I, II and III by diffusion or/and turbidimetric assays with Bacillus subtilis or Staphylococcus aureus as the test organisms. Six laboratories from three countries participated. Experimental procedures were according to the European Pharmacopoeia, 3rd ed. The activity of SPM I is markedly higher than that of SPM II and III. By diffusion, the activities of SPM II and III relative to SPM I were found to be 57 and 72%, respectively. The interlaboratory relative standard deviations (RSD) varied from 3.6 to 16.3%. By turbidimetry, the activities of SPM II and III relative to SPM I were found to be 45 and 52%, respectively. The interlaboratory RSD values varied from 2.6 to 7.7%. The results of the study were analyzed according to the ISO 5725-2 guidelines to determine the repeatability, the between-laboratory and the reproducibility variances of both methods. PMID- 10704027 TI - Characterisation of the rat lung lavage model as biological assay for testing the activity of surfactant preparations. AB - The present report describes how pharmacological assays may be validated and sets a basis for a discussion on the validation of biological test systems. The note for guidance on the validation of analytical procedures published by the European agency for the evaluation of medicinal products was adapted to the validation of a pharmacological test system. The presently described rat lung lavage test (RLL test) is an animal model that has great similarities to the pathophysiology of the acute respiratory distress syndrome of humans. In this RLL-test, the activity of surfactants can be tested in a standardised fashion. The usefulness of the point estimator and the corresponding confidence intervals (CI) as a statistical test procedure for equivalence was demonstrated. A validation can be based on the above mentioned guidance but should be adjusted to pharmacological needs. Based on the presented experiences, it can be concluded that a specific guideline for validation of pharmacological or biological tests is desirable. PMID- 10704028 TI - Application of the quartz crystal microbalance to the monitoring of Staphylococcus epidermidis antigen-antibody agglutination. AB - The change in solution properties due to the agglutination of an antigen with its specific antibody has previously been used as a marker of infection. This method has been modified to allow the binding activity between species to be followed using the frequency response of a quartz crystal microbalance (QCM). The Bayston agglutination plate assay for Staphylococcus epidermidis has been modified to allow the electrode of a QCM to act as a direct sensor for the change in solution properties as agglutination occurs. Antibody and antigen were introduced to the crystal surface and the agglutination process was followed as a change in crystal resonant frequency. Serum, known to be infected with the organism, gave a titre of 3.9x10(-2)% v/v (-118 Hz, +/-12 SD, N = 9) matching that given by triplicate plate assay. Uninfected serum gave no frequency changes at this concentration, yielding a titre of 2.5x10(-2)% v/v again matching the plate titre (N = 3). Infected serum gave responses 40 times faster then those of the uninfected serum. The piezoelectric quartz crystal method gave a positive or negative diagnosis in <15 min compared with the 24 h required for the plate assay. PMID- 10704029 TI - A rapid and selective solid-phase UV spectrophotometric method for determination of ascorbic acid in pharmaceutical preparations and urine. AB - Ascorbic acid was determined by a solid-phase UV spectrophotometry technique through the sorption of this on a dextran-type anion-exchange resin, Sephadex QAE A-25 and posterior direct measurement of its absorbance on the resin at 267 and 400 nm, packed in a 1-mm cell. The calibration graph was linear over the range 0.3-5.0 microg ml(-1). The sensitivity obtained is more than 50 times higher than that of the corresponding solution method. The detection limit was 0.05 microg ml(-1) and the relative standard deviation 0.74% (n = 10). This method is very rapid and highly selective for determining ascorbic acid in the presence of other species absorbing in the ultraviolet region and which are normally encountered with it. The one-step method proposed was successfully applied in the determination of ascorbic acid in pharmaceutical preparations and urine and the results were of comparable accuracy as indicated by a statistical analysis of the data, using both t- and F-tests. PMID- 10704030 TI - Simultaneous multivariate spectrophotometric analysis of paracetamol and minor components (diphenhydramine or phenylpropanolamine) in tablet preparations. AB - The use of multivariate spectrophotometric calibration is reported for the analysis of two decongestant tablets, where paracetamol is the principal component and diphenhydramine or phenylpropanolamine are the minor components. The resolution of these mixtures has been accomplished without prior separation or derivatisation, by using partial least-squares (PLS-1) regression analysis of electronic absorption spectral data. Although the molar ratios of paracetamol to the minor components were 38:1 and 25:1 respectively, the latter have been determined with high accuracy and precision, and with no interference from tablet excipients. PLS is able to take into account small deviations of paracetamol from linearity in the studied concentration range. The application of classical least squares (CLS) analysis yields unsatisfactory results, due to the low absorbances of the minor components within the range where all components obey Beer's law. PMID- 10704031 TI - Flow injection analysis of mercury(II) in pharmaceuticals based on enzyme inhibition and biosensor detection. AB - An enzymatic amperometric procedure for measurement of mercury(II) in pharmaceuticals, based on the inhibition of invertase and on a glucose electrode was studied. Analytical parameters for measurements in batch and flow injection analysis (FIA) have been optimised. Mercury(II) was detected in the 10-60 ppb range with RSD < or =2%. A sample throughput of 6 h(-1) for batch and 15 h(-1) for FIA was obtained. The total mercury(II) from thimerosal (thiomersal, sodium ethylmercurithiosalicylate) in eye-drop samples was measured with the amperometric procedure after oxidative cleavage treatment. Results for both batch and FIA procedures correlated well with atomic absorbtion spectroscopy (AAS) data. PMID- 10704032 TI - Analysis and pharmacokinetics of quetiapine and two metabolites in human plasma using reversed-phase HPLC with ultraviolet and electrochemical detection. AB - A sensitive and specific HPLC assay for the measurement of the antipsychotic compound quetiapine in human plasma has been developed and validated. The assay employs a three-step liquid liquid extraction of quetiapine and its 7 hydroxylated and 7-hydroxylated, N-dealkylated metabolites from human plasma, and utilizes ultraviolet (UV) detection of quetiapine and electrochemical detection of the metabolites. The method provides a linear response from a quantitation limit of 2.50 to 500 ng ml(-1) for each analyte using 0.4 ml plasma. The assay is applicable from 500 to 5000 ng ml(-1) by sample dilution with de-ionized water. The inter-assay precision of quetiapine in plasma calibration standards across 4 validation days averaged 11.9% relative standard deviation (RSD) over the range 2.50 to 500 ng ml(-1), with intra-assay precision averaging 16.0% RSD and mean accuracy of 98.6% of theory. Similarly, the inter-assay precision of the 7 hydroxylated metabolite in plasma calibration standards across 4 validation days averaged 13.7% RSD over the range 2.50 to 500 ng ml(-1), with intra-assay precision averaging 17.6% RSD and mean accuracy of 109% of theory. The 7 hydroxylated, N-dealkylated metabolite demonstrated inter-assay precision of 16.2% RSD, intra-assay precision of 19.9% RSD, and mean accuracy of 104% of theory over the range 2.50 to 500 ng ml(-1). The present assay method was used to support a study comparing the pharmacokinetic profile of quetiapine with the time course of dopamine D2 and serotonin 5-HT2 receptor occupancy in the brain using positron emission tomography (PET). We describe in this paper the bioanalytical method and the plasma concentrations of quetiapine and its metabolites resulting from this study. PMID- 10704033 TI - Stability study of piroxicam and cinnoxicam in solid pharmaceuticals. AB - The pseudo-first order rate constant for the hydrolysis of cinnoxicam as a function of temperature was obtained by variable-temperature kinetic experiments. The method used is on a generalization of non-isothermal analysis, and takes advantage of the capabilities of modern data collection and processing systems. A spectrophotometric method under non isothermal conditions was carried out. The results obtained are identical to those obtained under the same conditions by using traditional constant-temperature kinetic runs. This provides the possibility of reducing the amount of time spent and chemicals usually used in collecting kinetic data in mechanistic studies in solution by an order of magnitude. PMID- 10704034 TI - Multiwavelength spectrophotometric determination of acid dissociation constants: Part III. Resolution of multi-protic ionization systems. AB - A multiwavelength spectrophotometric (WApH) titration method was applied to study several multi-protic histamine H2-receptor antagonists which involved four acid dissociation constants (pKa values) over the pH range of 2-10. Specifically, UV absorption spectra of the drug solution were acquired in the course of a pH metric titration using an optical device based on a fibre optics dip probe, a light source and a diode array detector. Target factor analysis was utilized to deduce the pKa values from the spectral data recorded at different pH. It was noted that some of the pKa values were within mid pH range which were difficult to obtain because of insufficient absorption spectra acquired in the un-buffered region of the titration curve. With the aid of the WApH technique coupled with an optically transparent buffer, all pKa values have been successfully determined and were in excellent agreement with those measured using a conventional pH metric method. PMID- 10704035 TI - 1H and 13C nuclear magnetic resonance studies of the sites of protonation in itraconazole and fluconazole. AB - 1H and 13C nuclear magnetic resonance spectra of itraconazole were measured in neutral solution, and upon addition of varying amounts of deuterated hydrochloride acid. The comparison of the chemical shift in the different solutions revealed the high proton affinity of the piperazine nitrogen N26. Upon addition of a surplus of acid, the triazole ring was protonated. Corresponding observations were made for fluconazole. PMID- 10704036 TI - Extraction and spectrophotometric determination of copper(II) with S,S-bis(2 aminophenyl)oxalate. AB - A new selective reagent, S,S'-bis(2-aminophenyl)oxalate (H2L), for the extractive spectrophotometric determination of copper has been prepared. The ligand, H2L, forms a 1:1 complex with copper(II) in methanol. The molar absorptivity of Cu(II) S,S'-bis(2-aminophenyl)oxalate complex in methanol is 5365 M(-1) cm(-1) at 504 nm. The method has been applied for the determination of copper in pharmaceutical formulations, environmental and foodstuff samples. PMID- 10704037 TI - Simultaneous spectrophotometric determination of rutin, quercetin and ascorbic acid in drugs using a Kalman Filter approach. AB - A UV-spectrophotometric analysis of severely overlapped spectra yielded by anti capillary fragility pharmaceutical materials, viz. rutin, quercetin and ascorbic acid, in authentic mixtures and in tablets or soft elastic capsules was proposed. The method was based on Kalman Filter calibration of either orthogonal experimental design, i.e. standard solutions containing only one component at a time, or non-orthogonal experimental design, i.e. standard solutions containing more than one component. This calibration was followed by quantitative determinations of two- and three-component mixtures in synthetic mixtures or in oral dosages within the concentration range 2-10 microg ml(-1). A statistical analysis of the results was reported. PMID- 10704038 TI - Monitoring of oxytetracycline in ovine milk by high-performance liquid chromatography. AB - A method for 'in vivo' determination of the oxytetracyclin residues in ovine milk at low levels is described. Two groups of Sardinian breed sheep were treated with a dose of oxytetracycline by intramammary infusion and intramuscular administration, respectively. Oxytetracycline residues in extracts obtained from a preliminary cleanup procedure, were detected by an isocratic high-performance liquid chromatography (HPLC) method. Linear calibration plots were obtained over a large concentration range of 1 mg ml(-1)-10 ng ml(-1), with correlation coefficients higher than 0.996. Recoveries between 85.8 and 98.9% were obtained. Limit of detection (LOD) and limit of determination (LOQ) were 5.2 and 17.5 ng ml(-1), respectively. This method would be useful for routine monitoring of oxytetracycline residues in ovine dairy milk. PMID- 10704039 TI - Rapid liquid chromatographic assay for the determination of acetaminophen in plasma after propacetamol administration: application to pharmacokinetic studies. AB - A simple method for the rapid estimation of acetaminophen in plasma is described here. p-Propionamidophenol was used as internal standard. The assay involved a single ethyl acetate extraction and liquid chromatographic analysis at a wavelength of 242 nm using a reversed-phase encapped column, with a mobile phase of acetonitrile and 0.005 M potassium dihydrogen phosphate adjusted at pH 3.00. The limit of quantitation of acetaminophen by this method was 0.05 microg ml(-1), only 0.1 ml of the plasma sample was required for the determination. The calibration graph was linear from 0.05 to 100 microg ml(-1). Intra and inter-day precision (CV) did not exceed 8.93%. Mean recoveries of 90.31% with a CV of 1.38% were obtained. Applicability of the method was demonstrated by a pharmacokinetic study in normal volunteers who received 2 mg propacetamol. PMID- 10704040 TI - Spectrophotometric determination of silicate traces in hemodialysis solutions. AB - Reliable methods for the analysis of silicon are of great importance, because it seems that the silicate anion can reduce aluminum bioavailability in patients undergoing dialysis. Thus, a simple and sensitive spectrophotometric method is described for the determination of silicate traces in dialysis solutions. The method is based on the reaction between silicate ions and excess ammonium molybdate reagent to give a yellow silico-molybdic complex. This complex is then reduced to the heteropoly blue compound by means of ascorbic acid. Absorbance values are measured at 830 nm, and are stable for more than 2 h. A good linearity was obtained up to 300 ng ml(-1) of silicon concentration. The accuracy and the precision of the method were good; relative standard deviation values of 2% intraday and of 3.9% interday for six replicates on 40 ng ml(-1) standard silicate solutions were found. Results of the analysis of some commercial hemodialysis solution samples, obtained by means of the 'standard additions' method, are provided. PMID- 10704041 TI - Spectrophotometric methods for the determination of benazepril hydrochloride in its single and multi-component dosage forms. AB - Three sensitive and accurate methods are presented for the determination of benazepril in its dosage forms. The first method uses derivative spectrophotometry to resolve the interference due to formulation matrix. The second method depends on the color formed by the reaction of the drug with bromocresol green (BCG). The third one utilizes the reaction of benazepril, after alkaline hydrolysis, with 3-methylbenzothialozone (MBTH) hydrazone where the produced color is measured at 593 nm. The latter method was extended to develop a stability-indicating method for this drug. Moreover, the derivative method was applied for the determination of benazepril in its combination with hydrochlorothiazide. The proposed methods were applied for the analysis of benazepril in the pure form and in tablets. The coefficient of variation was less than 2%. PMID- 10704042 TI - Solid-phase extraction for the determination of caffeine in traditional Chinese medicinal prescriptions containing Theae folium by high performance liquid chromatography. AB - A high performance liquid chromatographic method in combination with C-18 reverse phase solid-phase extraction (SPE) was developed for determination of caffeine (CA) in traditional Chinese medicinal prescriptions which contain Theae folium. The frequently used prescriptions include Shin-Yi-San, Chuan-Chyong-Char-Tyau San, Tsang-Eel-San, San-Hwang-Shyr-Gau-Tang, Tzy-Shenn-Ming-Mu-Tang and Shiang Chyong-San. The present HPLC system uses a Merck RP-select B column by isocratic elution with methanol and 1% (v/v) acetic acid (1:4) as the mobile phase and detected at UV 270 nm. 8-Chlorotheophylline was used as an internal standard. The extracts of prescriptions were treated by Supelclean LC-18 SPE tube for eliminating interferences. Blank decoctions of each prescription were also examined as a test for interferences. The recoveries of caffeine from the Chinese medicinal prescriptions ranged from 88.5 to 92.3%. The relative standard deviations of caffeine ranged between 0.86 and 1.97% (intraday) and 1.04 and 3.90% (interday). The contents of caffeine in standard decoctions ranged from 13.98 to 19.62 mg g(-1). PMID- 10704043 TI - Flow-injection fluorimetric determination of 1,4-benzodiazepines in pharmaceutical formulations after acid hydrolysis. AB - A simple, rapid and fully automated flow injection method with fluorimetric detection after hydrolysis with H2SO4 in ethanolic or methanolic medium at room temperature has been developed for the determination of 1,4-benzodiazepines (oxazepam, diazepam and nitrazepam) in pharmaceutical formulations. The calibration curves are linear in the ranges (mg ml(-1)) of oxazepam (0.025 0.150), diazepam (0.010-0.125) and nitrazepam (0.010-0.150), with detection limits of 0.01, 0.005 and 0.005 mg ml(-1), respectively, and RSD (1% (n = 10). The measurement throughput is 60 h(-1) using a 200-microl sample volume obtained by the direct dissolution of formulations in alcohol. PMID- 10704044 TI - Simultaneous determination of ephedrine, pseudoephedrine, norephedrine and methylephedrine in Kampo medicines by high-performance liquid chromatography. AB - A simultaneous high-performance liquid chromatographic method for the determination of ephedrine, pseudoephedrine, norephedrine and methylephedrine (ephedrine alkaloids) in Kampo medicines which contain Ephedrae Herba was established. The analysis can be accomplished within 25 min with a Wakosil-II 5C18 HG column by isocratic elution using a mixture of water, acetonitrile and sodium dodecyl sulfate (65:35:0.4) as the mobile phase at a flow-rate of 1.0 ml min(-1), and detection at 210 nm. The detection limits of ephedrine alkaloids are 0.37-1.06 microM per injection (5 microl). This method was applied to analyze the quantities in eight Kampo decoctions; Mao-to, Makyo-yokukan-to, Makyo-kanseki-to, Yokuinin-to, Sho-seiryu-to, Keima-kakuhan-to, Kakkon-to and Kakkon-to-ka-senkyu sin'i. The concentration (per Ephedrae Herba gram) of ephedrine alkaloids was higher in the Makyo-kanseki-to decoction than in the others. Calcium sulfate from Gypsum Fibrosum raised ephedrine alkaloids dissolution in the Makyo-kanseki-to decoction. PMID- 10704045 TI - A comparison of gas-liquid chromatography, NMR spectroscopy and Raman spectroscopy for determination of the substituent content of general non-ionic cellulose ethers. AB - This paper describes and compares three techniques that can be used to characterize the substituent content of hydroxypropylcellulose (HPC and L-HPC) and hydroxypropyl methylcellulose (HPMC): gas-liquid chromatography (GLC) with a BP1 column and FI detection, 13C-NMR spectroscopy of hydrolysed samples, and Raman spectroscopy. GLC and 13C-NMR spectroscopy both allow independent quantification of hydroxypropoxyl and methoxyl contents. 13C-NMR spectroscopy, though requiring lengthier sample preparation, has the advantage of also quantifying the degree of substitution at each substitutable glucopyranose hydroxyl. Raman spectroscopy may be useful for rapid approximate estimation of hydroxypropoxyl content. PMID- 10704046 TI - Utility of microcalorimetry in the characterization of the browning reaction. AB - An isothermal microcalorimeter was utilized to characterize a model solid-state interaction. The degradation of the HIV protease inhibitor, DMP 450, in a binary mixture with hydrous lactose was followed in the presence of 5% additional water. Heat produced in the microcalorimeter sample vessel from either chemical or physical change is channeled through extremely sensitive thermopile blankets and is measured as it flows into infinite heat sinks. Solid-state 1:1 mixtures of DMP 450 and hydrous lactose each with 5% water added were analyzed in the microcalorimeter at 50, 60 and 65 degrees C. The resulting heat flow profiles were consistent with an autocatalytic rate law. An activation energy of 26.12 kcal mol(-1) for the DMP 450:lactose mixture was determined from the slope of the Arrhenius plot of the microcalorimetry heat flow maximum value versus the reciprocal of the absolute temperature. The activation energy determined by the traditional method with HPLC analysis was found to be in excellent agreement with the microcalorimetry value at 26.38 kcal mol(-1). PMID- 10704047 TI - A modified simple and rapid reversed phase liquid chromatographic method for quantification of diazepam and nordiazepam in plasma. PMID- 10704048 TI - Increased luminescence of the tetracycline-europium(III) system following oxidation by hydrogen peroxide. PMID- 10704049 TI - High performance thin layer chromatographic determination of tolperisone hydrochloride. PMID- 10704050 TI - Validation of a reversed-phase liquid chromatographic method for the determination of hydrocortisone phosphate disodium in a gel formulation. PMID- 10704052 TI - Production of secretory leucocyte protease inhibitor (SLPI) in human pancreatic beta-cells. AB - Secretory leucocyte protease inhibitor (SLPI) is a potent inhibitor of granulocyte elastase and cathepsin G, and also an inhibitor of pancreatic enzymes like trypsin, chymotrypsin and pancreatic elastase. SLPI has also been shown to inhibit HIV-1 infections by blocking viral DNA synthesis. Since SLPI is an inhibitor of pancreatic proteases we wished to investigate whether SLPI was also actually produced in the pancreas. M-RNA from human pancreatic tissue showed evidence of SLPI production using the reverse transcriptase polymer chain reaction technique (RT-PCR). Using immunohistochemical methods SLPI was demonstrated in the beta-cells of the islets of Langerhans. The function could be local protease/antiprotease regulation or antiviral/antibacterial defence in the close vicinity of the cell surface, or even inside the beta-cell itself. PMID- 10704051 TI - Hyperresponsiveness in the human nasal airway: new targets for the treatment of allergic airway disease. AB - Allergic rhinitis is a condition which affects over 15% of the population in the United Kingdom. The pathological process involves two stages: nasal inflammation, and the development of nasal airway hyperresponsiveness (AHR) to allergen and a number of other stimuli. This results in the amplification of any subsequent allergic reaction, contributing to the chronic allergic state. A number of different hypotheses have been proposed to explain the underlying mechanism of AHR, including a role for eosinophil-derived proteins, free radicals and neuropeptides. While there may be a number of independent pathways which can result in AHR, evidence obtained from both animal models and in vivo experiments in humans indicate that some mediators may interact with one another, resulting in AHR. Further research into these interactions may open new avenues for the pharmacological treatment of chronic allergic rhinitis, and possibly other allergic airway diseases. PMID- 10704053 TI - Reduced airway hyperresponsiveness by phosphodiesterase 3 and 4 inhibitors in guinea-pigs. AB - The aim of the present study was to compare the effects of selective phosphodiesterase (PDE) 3, 4 and 5 inhibitors on antigen-induced airway hyperresponsiveness in sensitized guinea-pigs. When the sensitized guinea-pigs were orally pre-treated with the selective PDE4 inhibitor, Ro 20-1724 (30 mg/kg), and studied 48h after OA, a significant reduction (P<0.01) of the leftward shift of the dose-response curve to ACh was noted, whereas it was ineffective at the lower dose (10 mg/kg). Administration of the selective PDE3 inhibitor, milrinone (30 mg/kg) also elicited a significant reduction (P<0.01) of the airway hyperresponsiveness, whereas the PDE5 inhibitor zaprinast (30 mg/kg) was ineffective. These results show that both PDE3 and PDE4 inhibitors are able to inhibit the antigen-induced airway hyperresponsiveness in sensitized guinea-pigs and support the potential utility of selective PDE inhibitors in the treatment of asthma. PMID- 10704054 TI - Inflammatory markers in cystic fibrosis patients with lung Pseudomonas aeruginosa infection. AB - Chronic endobronchial inflammation and bacterial infection are the main causes of morbidity and mortality in cystic fibrosis (CF), an autosomal recessive genetic disorder associated with improper function of chloride channels. Inflammation in CF lung is greatly amplified after Pseudomonas aeruginosa infection. In this study the relationship between P. aeruginosa status and inflammatory markers has been investigated. Seventeen CF children in acute lung exacerbation were examined. CF patients without P. aeruginosa infection were characterized by elevated activity of sputum elastase, reduced response of peripheral blood lymphocytes to PHA and significant resistance to the antiproliferative action of glucocorticoids. These parameters were normalized after antibiotic treatment. The patients with prolonged P. aeruginosa infection demonstrated extremely high levels of elastase activity and elevated amounts of sputum IL-8 and TNF-alpha. Although antibiotic treatment resulted in clinical improvement, it failed to suppress excessive immune response in the lung. The data indicate that CF patients with prolonged P. aeruginosa need the modified treatment, which should include immunomodulating drugs and protease inhibitors as well as antibacterial therapy. PMID- 10704055 TI - Extracorporeal circulation causes release of neutrophil gelatinase-associated lipocalin (NGAL). AB - Extracorporeal circulation (ECC) used during cardiac surgery causes activation of several inflammatory systems. These events are not fully understood but are responsible for complications during the immediate postoperative period. Neutrophil gelatinase-associated lipocalin (NGAL), a member of the expanding lipocalin family, has recently been described as an inflammatory protein. In this study, the release of NGAL into the circulation in 41 patients undergoing heart surgery with ECC was evaluated. A 4- to 5-fold elevation of the concentration of NGAL in plasma was observed during the immediate postoperative course with a rapid elimination during the first postoperative day. Four patients undergoing lung surgery (without ECC) were also studied. The plasma concentration of NGAL only increased with a factor of 1.1-2.2 over the operation. We conclude that NGAL is released into the circulation during heart surgery, probably as a result of the inflammatory activation of leukocytes initiated by the extracorporeal circulation. PMID- 10704056 TI - Inhibitory effect of ferulic acid and isoferulic acid on the production of macrophage inflammatory protein-2 in response to respiratory syncytial virus infection in RAW264.7 cells. AB - We investigated the effect of ferulic acid (FA) and isoferulic acid (IFA), which are the main active components of the rhizoma of Cimicifuga heracleifolia (CH), an anti-inflammatory drug used frequently in Japanese traditional medicine, on the production of macrophage inflammatory protein-2 (MIR-2) in a murine macrophage cell line, RAW264.7, in response to respiratory syncytial virus (RSV) infection. Following the exposure of cells to RSV for 20h, the MIP-2 level in condition medium was increased to about 20 ng/ml, although this level in mock infected cells was negligible. In the presence of either FA or IFA, RSV-infected cells reduced MIP-2 production in a dose-dependent manner. These data suggest that FA and IFA might be responsible, at least in part, for the anti-inflammatory drug effect of CH extract through the inhibition of MIP-2 production. PMID- 10704057 TI - Novel co-operation between eotaxin and substance-P in inducing eosinophil-derived neurotoxin release. AB - Eosinophils, chemokines, and neuropeptides are thought to play effector roles in the pathogenesis of allergic diseases such as rhinitis. Eotaxin is a novel C-C chemokine with a potent and relatively specific eosinophil chemoattractant activity that binds selectively to CCR3 receptor, however, its activity in inducing eosinophil granules proteins release is poorly characterized. This study was performed to determine whether eotaxin primes eosinophil exocytosis and whether this co-operates with the sensory neuroimmune-axis. In the present communication, we show that 10 ng/ml eotaxin primed normal human eosinophil for exaggerated eosonophil-derived neurotoxin (EDN) release stimulated by 10(-8) M Substance-P (SP). This novel priming was blocked by; 7B11 and Herbimycin A (HA), the CCR3 antagonist and the tyrosine kinase inhibitor, respectively. SDS-Page studies showed significant tyrosine phosphorylation of several protein residues induced by 10(-8) M SP only after priming with 10 ng/ml eotaxin. These results demonstrate a novel co-operation between eotaxin and SP in inducing eosinophil cytotoxicity, which at least in part involves tyrosine kinases pathway(s). PMID- 10704058 TI - BN 52021 decreases alveolar macrophage-mediated lung injury in experimental extrinsic allergic alveolitis. PMID- 10704060 TI - Changes in pulmonary histology and exfoliated bronchoalveolar cells induced by in vivo introduction of the tumor necrosis factor-alpha gene. AB - An inflammatory cytokine, tumor necrosis factor (TNF)-alpha, has been implicated in the pathogenesis of inflammatory lung diseases such as interstitial pneumonia (IP). To clarify the role of the inflammatory cytokine in the pathogenesis of lung inflammation, we introduced a murine TNF-alpha gene into murine lungs by the hemagglutinating virus of Japan (HVJ)-liposome method. Seven days after the TNF alpha gene introduction resulted in marked cellular infiltration of alveoli, and mild histological change was observed 28 days after the gene introduction. Electron microscopic analysis revealed minimal deposition of collagen fibrils. Analysis of the BAL revealed that the total cell number was markedly increased 3 and 7 days after the gene introduction, and more than 90% of the cells were macrophages. The increase in the cell number was returned to below the normal level 28 days after the gene introduction. During the development of IP, TNF alpha may regulate pathologic change of the pulmonary interstitium and alveolar cells. PMID- 10704059 TI - Loxosceles deserta spider venom induces the expression of vascular endothelial growth factor (VEGF) in keratinocytes. AB - Evenomation by arachnids of the genus Loxosceles frequently results in disfiguring necrotic skin lesions. The cellular and molecular mechanisms which contribute to lesion development are incompletely defined but appear to involve participation of several pro-inflammatory mediators. We have recently observed that Loxosceles deserta venom induces the production of chemokines in human umbilical vein endothelial cells (HUVECs) and human pulmonary epithelial cells. In the present study we observed that Loxosceles deserta venom induces the expression of vascular endothelial growth factor (VEGF) in human keratinocytes but little in smooth muscle cells and none in pulmonary epithelial cells. A potent endothelial cell-specific mitogen, VEGF induces angiogenesis and vascular permeability in vivo. RNase protection assay data indicate that VEGF mRNA concentrations in keratinocytes are significantly increased at 2 h following venom exposure. These data suggest that keratinocyte-derived VEGF may contribute to the vasodilation, edema and erythema which occur following Loxosceles evenomation. PMID- 10704061 TI - The influence of retinoic acid and retinoic acid derivatives on beta2 integrins and L-selectin expression in HL-60 cells in vitro. AB - A decreased expression of the beta2-integrin CD11b molecules on peripheral neutrophils from patients with pustular psoriasis occurred during treatment with retinoid compounds. Since this effect could not be mimicked in vitro with isolated peripheral neutrophils, the effect of retinoid compounds on cell differentiation was investigated. The promyelocytic cell line, HL60, was used to study what effect different retinoid compounds had on the cell surface expression of CD11b and L-selectin (CD62L) molecules, complement-mediated phagocytosis, adhesion and the oxidative burst. Retinoid-differentiated cells showed a significantly lower expression of CD11b and CD62L, and a decreased phagocytosis and oxidative burst compared to DMSO-differentiated HL60 cells or peripheral blood neutrophils. The diminished expression of beta2-integrins or L-selectin did not affect their adhesion to non-activated or lipopolysaccharide-activated endothelial cells in vitro but may however affect adhesion to vascular endothelium under shear forces during blood flow. These results suggest that retinoid treatment could affect several early steps in the inflammatory process. PMID- 10704062 TI - Lactoferrin protects gut mucosal integrity during endotoxemia induced by lipopolysaccharide in mice. AB - The hypothesis that lactoferrin protects mice against lethal effects of bacterial lipopolysaccharide (LPS) is the subject of experimental investigations described in this article. Lipopolysaccharide is a powerful toxin produced by gram negative bacteria that when injected into humans or experimental animals reproduce many of the pathophysiologic and immune responses caused by live bacteria. Lactoferrin administered intraperitoneally 1 hr prior to injection of LPS significantly enhanced the survival of mice, reducing LPS-induced mortality from 83.3% to 16.7%. Changes in locomotor and other behavioral activities resulting from LPS injection were not present in mice treated with lactoferrin. Also, histological examination of intestine revealed remarkable resistance to injury produced by LPS if mice were pretreated with lactoferrin. Severe villus atrophy, edema and epithelial vacuolation were observed in LPS-treated animals but not in lactoferrin-treated counterparts. Electrophysiological parameters were used to assess secretory and absorptive functions in the small intestine. In mice treated with LPS, transmural electrical resistance was reduced and absorption of glucose was increased. Lactoferrin treatment had no significant influence on basal electrophysiological correlates of net ion secretion or glucose absorption nor on changes induced by LPS. Collectively, these results suggest that lactoferrin attenuates the lethal effect of LPS and modulates behavioral and histopathological sequela of endotoxemia. PMID- 10704063 TI - Ligneous conjunctivitis: biochemical evidence for hypofibrinolysis. AB - Ligneous conjunctivitis (LC) is a rare disease of unknown etiology characterized by the growth of "woody" plaques on ocular and extraocular mucosa. These lesions are comprised of fibrin and both direct and indirect evidence implicates hypofibrinolysis as the primary defect in LC. To further elucidate the pathophysiology of LC we investigated the biochemical aspects of ligneous lesions with respect to the fibrinolytic system. Ligneous lesions were obtained from the right eye of a 15 year-old female patient with longstanding LC since age 2.5 year. Ligneous conjunctivitis in this patient has exhibited a chronic recurrent coarse and has involved multiple muscosal sites. Samples analyzed included an abundant mucoid thread from the conjunctival fornix and the ligneous plaque attached to the inferior tarsus. Samples were analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) to characterize protein profiles and by a variety of zymographic methods to visualize fibrinolytic enzymes. We found that mucoid and ligneous samples were distinct entities. Specifically, ligneous samples contained polypeptides with electrophoretic profiles characteristic of intact fibrin, and were replete in fibrin-bound tissue plasminogen activator (t-PA). Despite the presence of ample t-PA, ligneous samples were essentially devoid of fibrinolytic activity. In contrast, neither proteins nor t-PA could be detected in mucoid samples when fractionated by 7.5 15% SDS-PAGE or analyzed by fibrin zymography, respectively. Despite the absence of t-PA, mucoid samples were replete in fibrinolytic activity. This activity was plasminogen independent, heterogenous and inhibited by PMSF. Degradation profiles suggested that this activity represented in part alpha-chymotrypsin, consistent with this patient's treatment regime, as well as plasmin, elastase and an unidentified neutrophil-derived activity. Interestingly, ligneous samples contained both latent and activated forms of matrix metalloproteinase-9 (MMP-9), whereas mucoid samples contained predominantly activated forms of MMP-9. LC is characterized by defective fibrinolysis, despite the presence of ample t-PA and intact fibrin, and by an abundant mucoid thread which binds both endogenous and exogenous enzymes including serine protease(s) and collagenase(s). The implications of these results with respect to a role for exuberant mucus production or abnormal mucins in the development of a relative mucosal-site specific plasmin(ogen) deficiency is discussed. PMID- 10704064 TI - The impact of eotaxin- and IL-5-induced adhesion and transmigration on eosinophil activity markers. AB - Eosinophils accumulate at sites of allergic inflammation, and play important roles in asthma/allergic disorders. The mechanism of eosinophil recruitment into tissues is not fully understood. In this study, we evaluated whether adhesion and/or transmigration, in the presence of IL-5 and eotaxin, alter the expression of CD9, CD11b, the beta1alpha4-integrin, and the EG2-epitope on intracellular ECP. We also investigated whether CD9 is involved in the adhesion process. With flow cytometry the surface expression of CD9, CD11b and the beta1alpha4-integrin, and the intracellular expression of EG2, were analyzed before, and after transmigration/adhesion to fibronectin. To evaluate the eventual role of CD9 in adhesion, eosinophils were preincubated with monoclonal antibodies to CD9. We observed decreased expression of CD9, and increased expression of CD11b on eosinophils, after adhesion and transmigration. The transmigration did not change the expression of the beta1alpha4-integrin or EG2, whereas the adhesion resulted in a decreased EG2 expression. Antibodies to CD9 decreased the adhesion property of eosinophils. The eosinophils are activated after both adhesion and transmigration by means of decreased CD9 and increased CD11b expression. The expression of the EG2-epitope on intracellular ECP was decreased when eosinophils adhered to fibronectin, probably due to degranulation. Our results also indicate that CD9 is involved in the adhesion of eosinophils to fibronectin. PMID- 10704066 TI - Report of the Food and Drug Administration Subcommittee on Xenotransplantation: meeting of 3 and 4 June, 1999, Center for Biologics Evaluation and Research. PMID- 10704065 TI - Human neutrophils specifically interact with human monocyte-derived macrophage monolayers. AB - Neutrophils and macrophages express on their membrane molecules which may, in principle, interact with each other, promote specific cell to cell adhesion, affect cell function and finally, as a consequence, modulate the progression of the inflammatory process. We tested therefore if human neutrophils specifically adhere to human monocyte-derived macrophage monolayer (MDMM). Our findings show that neutrophils significantly adhere to 4-day old MDMM and that the extent of adhesion is increased by LPS-activation of MDMM. The specificity of the interaction was shown by the very low extent of adhesion of neutrophils either to freshly prepared monocyte or other types of cell monolayers and by the low percent of adhesion showed by eosinophils exposed to 7-day old MDMM. A role for beta2 integrins, CD31 and PAF-receptor in the mechanism of neutrophil-MDMM interaction is suggested by specific antagonists. We suggest that the adhesion between the two cell types could lead to an increase in concentration of neutrophil- or macrophage released factors in the interaction site and in a mutual modulation of phagocyte functions. PMID- 10704067 TI - Pig xenogeneic antigen modification with green coffee bean alpha-galactosidase. AB - Green coffee bean alpha-galactosidase can cleave the terminal alpha-galactose (alphaGal) on oligosaccharides that form the major antigen on pig endothelial cells recognized by primate-specific antibodies. Studies have been made of the conditions under which it is functional (e.g. temperature, pH) and of its biochemical and immunologic effects. Pig-to-rhesus monkey vein transplants were studied to identify the efficiency of the enzyme in delaying hyperacute rejection. When a graft became occluded, biopsies were taken for light microscopy (hematoxylin and eosin), scanning electron microscopy (SEM) and immunostaining with Griffonia simplicifolia IB4 lectin (GSIB4), and for IgM, IgG and C3. alpha Galactosidase was stable for 72-96 h and was effective at 4 degrees C and pH 6.9 (conditions of human liver graft storage), although better function was obtained at 20 degrees C and pH 6.5. Using the porcine PK15 cell assay, the cytotoxicity of human serum was reduced after treatment of the pig cells with the enzyme. In vitro studies demonstrated that porcine veins treated with alpha-galactosidase lost endothelial expression of the Gal epitope within 30 min. SEM, however, demonstrated endothelial damage beginning within 2 h, probably caused by the alpha-galactosidase, as no damage was found in phosphate-buffered saline-treated veins, where the Gal epitope was preserved for >3 h. No change was found in either group on light microscopy. In vivo studies demonstrated that patency of the alpha-galactosidase-treated veins (mean 2.5 h) was longer than that of untreated veins (0.23 h) (P < 0.01). Biopsies showed no GSIB4 lectin staining for alpha-Gal epitopes and much less IgM and C3 deposition in the treated group. Light microscopy and SEM demonstrated more severe endothelial damage, hemorrhage, and fibrin formation in the untreated group. Galactosidase is effective in removing the terminal alphaGal and delays the onset of hyperacute rejection of pig veins transplanted into monkeys. However, its effect is temporary and, on its own, its use is unlikely to prolong survival of pig organs transplanted into primates sufficiently to be of clinical value. PMID- 10704068 TI - Immunohistologic evaluation of mechanisms mediating hyperacute lung rejection, and the effect of treatment with K76-COOH, FUT-175, and anti-Gal column immunoadsorption. AB - Although most investigators agree that lung dysfunction occurs rapidly in various pig-to-primate hyperacute lung rejection (HALR) models, the basic mechanisms mediating this phenomenon remain in question. Here we describe an immunohistochemical method for assessment of mechanisms driving HALR. Using an established model wherein piglet lungs are perfused ex vivo with human blood, six experimental groups (K76 COOH; FUT-175; K76 with FUT; anti-alpha-Gal column adsorption; column with FUT; and column with K76) and two control groups (unmodified human blood; autologous pig blood) were studied. Each lung was biopsied serially during perfusion, and assessed using an immunohistochemical technique, with vWF staining as an internal control to quantitate binding of human IgM, IgG, C3, C5b-9, properdin, and C1q. The effect of each treatment and subsequent lung perfusion on IgG and IgM anti-alpha-Gal titers(by ELISA) and on pig endothelial cell cytotoxicity were correlated with histologic findings. We found that [1] the classical complement activation pathway was activated, as has been shown for other pig organs in primate or human blood environments [2]; alternative complement pathway activation is also seen, which has not been described for other organs in pig-to-primate models, but only in the context of classical pathway activation; and [3] anti-Gal column absorption, pharmacologic inhibition of complement, or combination therapy each was associated with histologic evidence of partial protection, consistent with what would be predicted for each intervention. Further, immunohistologic differences correlated with physiologic outcomes [8] and with antibody assay results, and revealed that treatments used were incompletely effective. Our data suggest that more complete inhibition of antibody- and complement-driven pathways than was achieved in these experiments will be necessary to prevent the antibody and complement-mediated facets of hyperacute lung rejection. This immunohistologic technique may also help us identify additional pathogenic mechanisms important to eventual clinical application of pig-to-human lung xenografts. PMID- 10704069 TI - The effect of macrophage depletion on delayed xenograft rejection: studies in the guinea pig-to-C6-deficient rat heart transplantation model. AB - The purpose of this study was to investigate the effect of macrophage depletion, using liposome-encapsulated dichloromethylene diphosphonate (Lip-Cl2MDP), on delayed xenograft rejection (DXR) in the guinea pig-to-C6-deficient rat heart transplantation model. In this model, hyperacute rejection does not occur, but, in untreated recipients, xenografts are still destroyed by DXR within 1-2 days. Graft survival was 68 +/- 8.4 h in splenectomized control rats, 77 +/- 16.3 h with Lip-Cl2MDP alone, 99 +/- 10.4 h with deoxysperguarlin (DSG; P < 0.01 vs. controls), and 127 +/- 19.4 h with Lip-Cl2MDP plus DSG (P < 0.01 vs. DSG alone). Treatment with DSG alone or in combination with Lip-Cl2MDP resulted in significant reductions in serum IgM levels at rejection. Immunohistological studies showed that Lip-Cl2MDP alone or in combination with DSG reduced infiltration of grafts by both EDI + and ED2 + macrophages. These experiments support the concept that macrophages play an important role in DXR and suggest that strategies targeting macrophages may be useful in controlling DXR. PMID- 10704070 TI - Xenograft rejection of fetal porcine islet-like cell clusters in the rat: effects of active and passive immunization. AB - The process of islet xenograft rejection is still poorly understood. To elucidate further possible mechanism(s) involved in xenograft rejection, the effect of different immunization protocols was investigated. Fetal porcine islet-like cell clusters (ICCs) were transplanted under the kidney capsule in otherwise untreated rats, rats pre-immunized by s.c. injections of ICCs and in rats passively immunized with immune serum. The rejection process was evaluated with regard to antibody and complement deposition in the graft, as well as to morphology and phenotype of the infiltrating cells. In otherwise untreated animals, a moderate perigraft mononuclear cell infiltrate was seen after 3 days. Graft destruction became evident on day 6 with marked intragraft infiltration by macrophages (ED1 positive), whereas T cells were in the minority and mainly located in the perigraft area. In contrast to the findings in non-immunized rats, the rejection process in pre-immunized rats was characterized by marked intragraft infiltration by macrophages 3 days after transplantation. Moreover, both T cells and macrophages heavily infiltrated the adjacent kidney parenchyma, and major histocompatibility complex (MHC) class II expression in surrounding kidney tubular cells was concomitantly enhanced. Syngeneic rat islets mixed with porcine ICCs escaped the rejection process in non-immunized rats but were affected in pre immunized animals. Thus, the specificity of the rejection process in non immunized animals seems to be lost in pre-immunized animals. The early macrophage infiltration was also accelerated in rats passively immunized with immune serum, but no early switch from perigraft to intragraft infiltration or subsequent cellular infiltration in the adjacent kidney parenchyma was seen. Circulating xenoreactive antibodies of the IgG isotype increased after transplantation in normal and otherwise untreated rats. No distinct IgG deposition in the ICC xenografts was observed until day 12 after transplantation in untreated rats, whereas perigraft deposition of IgG was found 1 day after transplantation in pre immunized rats and in rats given immune serum. No deposition of complement was observed within the ICC xenograft in any of the groups during the observation period. The dependence on T cells, the massive infiltration of macrophages with a unique phenotype, the cellular distribution, and the loss of specificity (bystander killing) of the rejection process in immunized rats suggest that ICC xenograft rejection shares some of its main characteristics with a delayed type hypersensitivity-like (DTH) immune response. PMID- 10704071 TI - Literature update 1999, part 2. PMID- 10704072 TI - The mechanisms of action of interleukin-1 on rabbit intestinal epithelial cells. AB - Interleukin-1 (IL-1) is an inflammatory mediator that increases Cl- secretion in intestinal epithelial cells. To identify the signal transduction pathway(s) involved in IL-1's action, cells were treated with IL-1 and the levels of cyclooxygenase (COX) enzymes, prostaglandin E2 (PGE2) and phospholipase A2 activating protein (PLAP), and the activity of phospholipase A2 (PLA2) were measured. IL-1 caused concentration- and time-dependent increases in the levels of PLA2 activity, and/or in the levels of PLAP, COX-2 and PGE2. The IL-induced increase in PGE2 levels was biphasic, with the first peak due to the increase in PLAP levels, and the second peak due to the increase in COX-2 levels. This increase in PGE2 levels may provide a mechanism for acute and chronic inflammation in the intestine. PMID- 10704073 TI - Suppressive activity of a macrolide antibiotic, roxithromycin, on pro inflammatory cytokine production in vitro and in vivo. AB - This study was designed to examine the influence of a macrolide antibiotic, roxithromycin (RXM), on the production of pro-inflammatory cytokines, interleukin (IL)-1beta and tumor necrosis factor (TNF)-alpha. In the first experiments, we examined the effect of RXM on in vitro cytokine production from lipopolysaccharide (LPS)-stimulated human peripheral blood monocytes. The monocytes were cultured in the presence of various doses of the agent. After 24 h, the culture supernatants were obtained and assayed for IL-1beta and TNF-alpha contents by enzyme-linked immunosorbent assay. RXM suppressed the in vitro production of IL-1beta and TNF-alpha in response to LPS stimulation. This was dose dependent and first noted at a concentration of as little as 0.05 microg/ml, which is much lower than therapeutic blood levels. In the second part of the experiments, we examined the influence of RXM on the appearance of IL-1beta and TNF-alpha in mouse lung extract induced by LPS inhalation. RXM was administered orally into BALB/c mice at a single dose of 2.5 mg/kg once a day for 5-12 weeks. These mice were then instilled with LPS into the trachea and examined for the presence of cytokines in aqueous lung extracts. Pretreatment of mice with RXM for 5 weeks did not influence of the appearance of both IL-1beta and TNF-alpha in aqueous lung extracts. However, pretreatment for more than 7 weeks dramatically suppressed the cytokine appearance in the extracts. PMID- 10704074 TI - Studies on the biological effects of ozone: 10. Release of factors from ozonated human platelets. AB - In a previous work we have shown that heparin, in the presence of ozone (O3), promotes a dose-dependent platelet aggregation, while after Ca2+ chelation with citrate, platelet aggregation is almost negligible. These results led us to think that aggregation may enhance the release of platelet components. We have here shown that indeed significantly higher amount of platelet-derived growth factor (PDGF), transforming growth factor beta1 (TGF-beta1) and interleukin-8 (IL-8) are released in a dose-dependent manner after ozonation of heparinised platelet-rich plasma samples. These findings may explain the enhanced healing of torpid ulcers in patients with chronic limb ischemia treated with O3 autohaemoteraphy (O3-AHT). PMID- 10704075 TI - A possible involvement of ion transporter in tumor necrosis factor alpha and cycloheximide-induced apoptosis of endothelial cells. AB - We examined the tumor necrosis factor alpha (TNFalpha)-induced apoptosis of vascular endothelial cells from the standpoint of ion channels. Cultured vascular endothelial cells from bovine carotid artery were used. Apoptosis was determined by a propidium iodide assay. Treatment of the endothelial cells with TNFalpha and cycloheximide for 6 h induced nuclear fragmentation in a TNFalpha dose-dependent manner (1-10 ng/ml). Concomitant treatment of endothelial cells with TNFalpha at a dose of 10 ng/ml and cycloheximide at a dose of 10 microg/ml elicited endothelial cell apoptosis as high as 23.4+/-4.1% at 6 h after administration. However, 10 ng/ml TNFalpha alone elicited a little apoptosis at 6 h after its administration (% apoptosis=4.1+/-0.8%). Cycloheximide (10 microg/ml) did not induce apoptosis at all. Concomitant treatment of endothelial cells with 1 mmol/l of 4,4-diisothiocyanatostilbene-2,2-disulfonic acid, which is a chloride bicarbonate exchanger blocker, partially inhibited the TNFalpha and cycloheximide induced endothelial cell apoptosis. On the other hand, endothelial cell apoptosis due to TNFalpha and cycloheximide was completely inhibited by benzyloxycarbonyl Asp-CH2OC(O)-2,6-dichlorobenzene (50 micromol/l), an inhibitor of caspase. Moreover, pyrrolidine dithiocarbanate, an inhibitor of nuclear factor kappa B (NF kappaB), also suppressed endothelial cell apoptosis induced by TNFalpha and cycloheximide completely. These findings suggest that the endothelial cell apoptosis induced by TNFalpha and cycloheximide is closely related to not only chloride ions, but also both NF-kappaB and caspase activation. That is to say, there is a possibility that chloride ions or bicarbonate (pH) may play an important role in signal transduction such as NF-kappaB and caspase activation in the apoptosis induced by TNFalpha and cycloheximide. PMID- 10704076 TI - Aspirin and some other nonsteroidal anti-inflammatory drugs inhibit cystic fibrosis transmembrane conductance regulator protein gene expression in T-84 cells. AB - Cystic fibrosis (CF) is caused by mutations in the CF gene, which encodes CF transmembrane conductance regulator protein (CFTR), a transmembrane protein that acts as a cAMP-regulated chloride channel The disease is characterized by inflammation but the relationship between inflammation, abnormal transepithelial ion transport, and the clinical manifestations of CF are uncertain. The present study was undertaken to determine whether three nonsteroidal anti-inflammatory drugs (NSAIDs) (aspirin, ibuprofen, and indomethacin) modulate CFTR gene expression in T-84 cells. Treatment with NSAIDs reduced CFTR transcripts, and decreased cAMP-stimulated anion fluxes, an index of CFTR function. However, the two phenomena occurred at different concentrations of both drugs. The results indicate that NSAIDs can regulate both CFTR gene expression and the function of CFTR-related chloride transport, and suggest that NSAIDs act via multiple transduction pathways. PMID- 10704077 TI - Time dependent production of cytokines and eicosanoids by human monocytic leukaemia U937 cells; effects of glucocorticosteroids. AB - In the present study the human monoblast cell line U937 has been used as a model to study the function of human mononuclear phagocytes in asthma. The kinetics of the production of eicosanoids and cytokines, which are thought to play a role in the pathogenesis of asthma, were studied. In addition, the effects of glucocorticosteroids were investigated, as these drugs are of great importance for the treatment of asthmatic patients. After stimulation with phorbol-12 myristate acetate (PMA) for 24 h, U937 cells were cultured in the absence or presence of lipopolysaccharide (LPS: 1 and 5 microg ml(-1)) and glucocorticosteroids (budesonide, fluticasone propionate and prednisolone: 10( 11), 10(-9) and 10(-7) M) for 96 h. The production of interleukin-1beta (IL 1beta), interleukin-6 (IL-6), prostaglandin E2 (PGE2) and thromboxane B2 (TxB2) gradually increased in time after stimulation with LPS, whereas the transient production of tumor necrosis factor alpha (TNF-alpha) reached its maximum between 6 and 12 h. Interferon-gamma (IFN-gamma), interleukin-10 (IL-10) and leukotriene B4 (LTB4) were not detectable. All three glucocorticosteroids (budesonide, fluticasone propionate and prednisolone) completely inhibited the production of both eicosanoids and cytokines. The production of eicosanoids was more sensitive to these glucocorticoids than the production of cytokines. The observed differences in the kinetics of the production of eicosanoids and cytokines stress the importance of time course experiments in studies on the effect of drugs on mononuclear cells. PMID- 10704078 TI - Lymphocytes apoptosis in patients with acute exacerbation of asthma. AB - Asthma is characterized by airway inflammation, which can be now assessed by the analysis of induced sputum. Ten patients with asthma were investigated during acute exacerbation for the quantification of apoptosis, for Bcl-2 and Fas expression, in induced sputum lymphocytes. They were compared to 12 patients with chronic obstructive pulmonary disease (COPD), and 10 healthy controls. Spontaneous apoptosis was determined by staining nuclei with propidium iodide, and analyzed with a FACScan. Bcl-2 was measured by Western blotting, and results were obtained by densitometric scanning, done by the gel proanalyser. The investigation of Fas was performed using the streptavidin-biotin preroxidase complex method. Patients with asthma and patients with COPD exhibited a significant increase of cellularity, percentage of neutrophils, eosinophils and lymphocytes when compared to healthy controls. Apoptosis in induced sputum mononuclear cells was found decreased in patients with asthma compared to COPD patients and healthy controls. The quantification of apoptosis was measured after exposure to anti-cytokine antibodies. Anti-TNF-alpha antibody blocked the apoptosis in both patients groups and healthy controls, suggesting that TNF-alpha acted as an inducer of apoptosis. Anti-IL-10 blocked apoptosis completely exclusively in patients with asthma. Bcl-2 expression was found to be increased in induced sputum mononuclear cells from patients with asthma, compared to healthy controls and patients with COPD. Expression of Fas could be detected in patients with asthma, at a lower level than COPD patients and healthy controls. Distinct mechanisms of apoptosis were found in patients with asthma and patients with COPD, characterized by different levels of Bcl-2 and Fas expression. Induction of apoptosis should be a beneficial process in allergic inflammation traduced in induced sputum mononuclear cells. The apoptosis process is assumed by two different mechanisms in asthma and COPD. Our findings indicated that in asthmatic patients, activated lymphocytes accumulate in the bronchi; because of their prolonged survival that maintains inflammation. PMID- 10704080 TI - Cyclosporin A decreases human macrophage interleukin-6 synthesis at post transcriptional level. AB - In addition to its well-established effect on T cells, cyclosporin A (CsA) also inhibits inflammatory cytokine production by macrophages. However, little is known about the mechanism of action of CsA on macrophage cytokine production. We measured the effect of CsA on basal and phorbol-myristate-acetate (PMA) stimulated production of interleukin-6 using the human monocyte cell line U937 differentiated with dimethylsulfoxide (DMSO). Interleukin-6 levels were measured in supernatant and cell lysates using specific enzyme-linked immunosorbent assays. We found that CsA decreases not only IL-6 release but also cytokine synthesis. The concentration of CsA used did not affect either cell viability or proliferation. Three possibilities may be advanced to explain the CsA-due decrease in IL-6 production by macrophages: (a) inhibition of the synthesis of an early common regulatory protein, (b) inhibition of cytokine gene transcription, or (c) modulation of post-transcriptional events. The first possibility was tested by measuring the effect of cycloheximide on the experimental system during the first 3 hours of culture. Although cycloheximide decreased total cytokine synthesis, the pattern of cytokine modulation by CsA persisted. These data suggest that CsA-mediated macrophage cytokine inhibition is not mediated by an early common regulatory protein. To further explore the inhibition mechanism, we measured IL-6 mRNA levels by Northern blot. IL-6 mRNA levels were unaffected by CsA both in resting and PMA-stimulated cells. We conclude that in human macrophages CsA diminishes IL-6 production at post-transcriptional level. PMID- 10704081 TI - Airway responsiveness: role of inflammation, epithelium damage and smooth muscle tension. AB - The purpose of this study was the effect of epithelium damage on mechanical responses of airway smooth muscles under different resting tension. We performed acetylcholine (ACh) (10(-5) M)-induced contraction on tracheal strips from 30 rabbits in five groups (0.5, 1, 1.5, 2 and 2.5 g) before and after epithelium removal. At low resting tension (0.5-1.5 g), the epithelium removal decreased the ACh-induced contractions. At 2 g resting tension, the epithelium removal increased the ACh-induced contractions of airways with intact epithelium about 20%. At 2.5 g resting tension, the elevation of contraction is about 25% (P<0.01). Consequently, after epithelium loss, the resting tension determines the airway smooth muscles responsiveness. In asthma, mediators such as ACh act on already contracted inflammatory airways, which results in additional increase of contraction. In contrast, low resting tension, a condition that simulates normal tidal breathing, protects from bronchoconstriction even when the epithelium is damaged. PMID- 10704079 TI - Interrelationship of the kinin system, nitric oxide and eicosanoids in the antigen-induced arthritis in rabbits. AB - The aim of the present study was to investigate the interrelationship of the kinin system, nitric oxide and eicosanoids in the acute phase of antigen-induced arthritis (AIA) in rabbits. The arthritis was induced in immunized rabbits and the following parameters were evaluated 24 hours later: leukocyte influx (total and differential white cell count), vascular permeability (Evans's blue method), and synovial PMN cell infiltrate. PGE2 and LTB4 (radioimmunoassay) levels were quantified in the synovial fluid. The animals were pre-treated with 20mg/kg/day during 14 days with L-NAME or D-NAME and/or Enalapril (0.12 mg/kg/day-14 days), and/or the B2 antagonist of Bradykinin HOE 140 (0.9 mg/kg). Our results showed that L-NAME was effective in the prevention of AIA with reduction of all Inflammatory parameters analyzed. Enalapril partially reverted the L-NAME anti inflammatory effects. The simultaneous treatment with HOE 140 abolished this reversion and returned the inflammatory parameters to the levels observed in L NAME treated animals. Our results suggest that pressoric alterations induced by L NAME could not account for all its anti-inflammatory action in this model of experimental arthritis. Additionally the contribution of the kinin system in AIA was characterized as well as its interaction with eicosanoids and nitric oxide. PMID- 10704082 TI - The immunoregulatory abilities of polymorphonuclear neutrophils in multiple sclerosis. PMID- 10704083 TI - Inflammation and CFTR: might neutrophils be the key in cystic fibrosis? AB - The aim of this hypothesis is to provide new insights into the still unclear mechanisms governing airway inflammation in cystic fibrosis. Although the genetic basis of cystic fibrosis as well as the molecular structure of cystic fibrosis transmembrane regulator (CFTR), the mutated protein which causes the disease, have been well defined, a clear relationship between the genetic defect and the pulmonary pathophysiology, especially chronic infections and neutrophil-dominated airway inflammation has not been established. Cystic fibrosis is thus a unique pathological situation in that neutrophils can be depicted as both an antiinfectious and a proinflammatory cell. In cystic fibrosis there is an emerging picture of an imbalance between these two roles with both a reduction in the antiinfectious efficacy and an augmentation of the proinflammatory functions. Better knowledge of fundamental defects in neutrophil function in cystic fibrosis as well as a novel cellular function of CFTR, which will be reviewed, will allow identification of potentially new clinical targets and aid selective therapeutic action aimed at counteracting the lethal neutrophil-induced airway inflammation. The rationale for colchicine therapy is a significant example of a drug which might act both at the molecular levels on CFTR expression in epithelial cells and on neutrophils to mediate antiinflammatory effects. Preliminary results are presented in this issue (Med Inflamm 1999; 8: 13-15). PMID- 10704084 TI - Interest of colchicine for the treatment of cystic fibrosis patients. Preliminary report. AB - Cystic fibrosis (CF) lung disease is characterized by persistent inflammation. Antiinflammatory drugs, such as corticosteroids and ibuprofen, have proved to slow the decline of pulmonary function although their use is limited because of frequent adverse events. We hypothesized that colchicine could be an alternative treatment because of its antiinflammatory properties and upregulatory effect on cystic fibrosis transmembrane regulator (CFTR) closely related proteins. We herein present results obtained in an open study of eight CF children treated with colchicine for at least 6 months. Clinical status was better in all patients and respiratory function tests significantly improved in five. Median duration of antibiotherapy decreased significantly. These preliminary results support our hypothesis of a beneficial effect of colchicine in CF patients and stress the need for a controlled therapeutic trial. PMID- 10704085 TI - Prevention of antigen-induced bronchial hyperreactivity and airway inflammation in sensitized guinea-pigs by tacrolimus. AB - We examined the effect of the immunosuppressive agent, tacrolimus (FK506), on antigen-induced bronchial hyperreactivity to acetylcholine and leukocyte infiltration into the airways of ovalbumin-challenged guinea-pigs. Subcutaneous injection of 0.5 mg/kg of FK506, 1 h before and 5 h after intra-nasal antigen challenge prevented bronchial hyperreactivity to aerosolized acetylcholine, eosinophilia in bronchoalveolar lavage (BAL) fluid and bronchial tissue and the invasion of the bronchial wall by CD4+ T-lymphocytes. FK506 also suppressed ovalbumin-induced increase in the number of leukocytes adhering to the pulmonary vascular endothelium and expressing alpha4-integrins. Inhibition by FK506 of antigen-induced bronchial hyperreactivity in sensitized guinea-pigs may thus relate to its ability to prevent the emergence of important inflammatory components of airway inflammation, such as eosinophil accumulation, as well as CD4+ T-lymphocyte infiltration into the bronchial tissue. PMID- 10704086 TI - Regulation of inflammatory responses to Bordetella pertussis by N(G)-monomethyl-L arginine in mice intranasally infected. AB - To investigate effect of MMLA, an inhibitor of nitric oxide (NO) production, on regulation of inflammatory responses to Bordetella pertussis infection, mice were infected intranasally, and treated with various concentrations of MMLA. Ten days after infection, mice treated with MMLA at dosage of 100 mg/kg, given intraperitoneally in a single dose or for 5 consecutive days, showed at histopathologic examination, a significant decrease of intensity of inflammation (scores, 0.6 +/- 0.2 and 0.9 +/- 0.5 respectively). A decrease of cellular accumulation of neutrophils and lymphocytes in the bronchoalveolar lavage (BAL) fluid was observed in infected mice treated with MMLA, especially at dosage of 10 mg/kg, given in a single dose intraperitoneally. In addition, BP-infected mice treated with MMLA (100 mg/kg, intraperitoneally) for 5 consecutive days showed higher mortality rate than untreated mice infected with B. pertussis, and the number of B. pertussis in lungs of mice treated with MMLA was significantly increased. However, MMLA treatment of infected mice had some effect on levels of IFN-gamma and nitrite/nitrate (end-stable products of NO) in the BAL fluid. This study indicates that NO may play a role either as microbiocidal agent or as a modulator of immune regulation, inasmuch as it may upregulate tissue inflammatory response to B. pertussis. PMID- 10704087 TI - Arachidonic acid and freshly isolated human bone marrow mononuclear cells. AB - Arachidonic acid (AA), a fatty acid found in the human bone marrow plasma, is the precursor of eicosanoids that modulate bone marrow haematopoiesis. To further our understanding of the role of AA in the bone marrow physiology, we have assessed its incorporation in human bone marrow mononuclear cells. Gas chromatography analysis indicates the presence of AA in their fatty acid composition. In bone marrow mononuclear cells, [3H]-AA is incorporated into triglycerides and is later delivered into phospholipids, a result not observed with blood mononuclear cells. Prelabelling-chase experiments indicate a trafficking of labelled AA from phosphatidylcholine to phosphatidylethanolamine. Stimulation of prelabelled bone marrow mononuclear cells with granulocyte-macrophage colony-stimulating factor (GM-CSF) results in the release of a part of the incorporated labelled AA. Finally, exogenous AA (up to 1 microM) has no significant effect on cell growth. In conclusion, human bone marrow mononuclear cells participate to the control of marrow AA concentrations by incorporating AA into phospholipids and triglycerides. In turn, bone marrow mononuclear cells can release AA in response to the potent haematopoietic growth factor GM-CSF. PMID- 10704088 TI - Prevention of renal injury after induction of ozone tolerance in rats submitted to warm ischaemia. AB - On the basis that ozone (O3) can upregulate cellular antioxidant enzymes, a morphological, biochemical and functional renal study was performed in rats undergoing a prolonged treatment with O3 before renal ischaemia. Rats were divided into four groups: (1) control, a medial abdominal incision was performed to expose the kidneys; (2) ischaemia, in animals undergoing a bilateral renal ischaemia (30 min), with subsequent reperfusion (3 h); (3) O3 + ischaemia, as group 2, but with previous treatment with O3 (0.5 mg/kg per day given in 2.5 ml O2) via rectal administration for 15 treatments; (4) O2 + ischaemia, as group 3, but using oxygen (O2) alone. Biochemical parameters as fructosamine level, phospholipase A, and superoxide dismutases (SOD) activities, as well as renal plasma flow (RPF) and glomerular filtration rate (GFR), were measured by means of plasma clearance of p-amino-hippurate and inulin, respectively. In comparison with groups 1 and 3, the RPF and GFR were significantly decreased in groups 2 and 4. Interestingly, renal homogenates of the latter groups yielded significantly higher values of phospholipase A activity and fructosamine level in comparison with either the control (1) and the O3 (3) treated groups. Moreover renal SOD activity showed a significant increase in group 3 without significant differences among groups 1, 2 and 4. Morphological alterations of the kidney were present in 100%, 88% and 30% of the animals in groups 2, 4 and 3, respectively. It is proposed that the O3 protective effect can be ascribed to the substantial possibility of upregulating the antioxidant defence system capable of counteracting the damaging effect of ischaemia. These findings suggest that, whenever possible, ozone preconditioning may represent a prophylactic approach for minimizing renal damage before transplantation. PMID- 10704089 TI - Evaluation of inflammatory cytokine secretion by human alveolar macrophages. AB - The alveolar macrophage (AM) secretes interleukin 1beta (IL-1beta), tumour necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6) and interleukin-8 (IL-8), all of them inflammatory cytokines involved in the pathogenesis of many lung diseases. The aim of the present work was to evaluate the basal and stimulated secretion of these cytokines by human AMs. Human AMs were collected by bronchoalveolar lavage (BAL) from four healthy controls and 13 patients with diffuse interstitial lung disease (five cases of sarcoidosis, three of hypersensitivity pneumonitis and five of idiopathic pulmonary fibrosis). AMs were cultured in the presence or absence of different concentrations of lipopolysaccharide (LPS), phorbolmyristate and gamma-interferon. IL-1beta, TNF alpha, IL-6 and IL-8 levels were measured in BAL fluid and culture supernatant using specific enzyme-linked immunosorbent assays. The substance found to stimulate the secretion of inflammatory cytokines to the greatest extent was LPS at a concentration of 10 microg/ml. Regarding the secretion of IL-1beta, four observations were of interest: basal secretion was very low; LPS exerted a potent stimulatory effect; considerable within-group variability was observed; and there were no significant differences in the comparisons among groups. With respect to TNF-alpha secretion, the results were similar. The only striking finding was the higher basal secretion of this cytokine with respect to that of IL-1beta. Regarding the secretion of IL-6, the same pattern followed by TNF-alpha was found. However, it should be stressed that the increase induced by LPS was smaller than in the two previous cytokines. Regarding the secretion of IL-8, three findings were patent: the strong basal secretion of this cytokine; the moderate increase induced by LPS; and the existence of significant differences among the different groups with respect to the stimulated secretion of this cytokine, which reached maximum values in patients with idiopathic pulmonary fibrosis. Finally, it should be noted that the pattern of cytokines observed in the BAL fluid was similar to that found in cultured AM supernatants. The pattern of inflammatory cytokine secretion by AMs differs from that of other cells of the mononuclear phagocyte system (MPS). In this sense. AMs secrete low amounts of IL 1, moderate amounts of TNF-alpha and IL-6, and high quantities of IL-8. Adherence is an important stimulus in the secretion of these molecules and LPS elicits an increased secretion inverse to the basal secretion. There is considerable individual variability in the secretion of inflammatory cytokines by the AMs of patients with interstitial lung disease and the AMs of these patients are primed in vivo for the secretion of these cytokines. The results of our study, carried out in vitro, can be extrapolated to the in vivo setting. PMID- 10704090 TI - Differential modulation of annexin I binding sites on monocytes and neutrophils. AB - Specific binding sites for the anti-inflammatory protein annexin I have been detected on the surface of human monocytes and polymorphonuclear leukocytes (PMN). These binding sites are proteinaceous in nature and are sensitive to cleavage by the proteolytic enzymes trypsin, collagenase, elastase and cathepsin G. When monocytes and PMN were isolated independently from peripheral blood, only the monocytes exhibited constitutive annexin I binding. However PMN acquired the capacity to bind annexin I following co-culture with monocytes. PMN incubation with sodium azide, but not protease inhibitors, partially blocked this process. A similar increase in annexin I binding capacity was also detected in PMN following adhesion to endothelial monolayers. We propose that a juxtacrine activation rather than a cleavage-mediated transfer is involved in this process. Removal of annexin I binding sites from monocytes with elastase rendered monocytes functionally insensitive to full length annexin I or to the annexin I-derived pharmacophore, peptide Ac2-26, assessed as suppression of the respiratory burst. These data indicate that the annexin I binding site on phagocytic cells may have an important function in the feedback control of the inflammatory response and their loss through cleavage could potentiate such responses. PMID- 10704091 TI - Photodegradation of folic acid in aqueous solution. AB - A study of the photodegradation of folic acid by ultraviolet radiation in aqueous solution has been made. Folic acid is photolysed by an apparent first-order kinetics and the log k-pH profile shows a gradual decrease in rate in the pH range 2.0-10.0. The profile indicates the appearance of three steps which reflect the participation of different ionic species of folic acid (pKa(1) 2.3, pKa(2) 8.3) in the photolysis reaction. The rate of photodegradation varies from 0.1550 x 10(-3) min(-1) (pH 10.0) to 5.04 x 10(-3) min(-1) (pH 2.5) in the pH range studied. The photolysis of folic acid shows that it is degraded to pterine-6 carboxylic acid and p-amino-benzoyl-L-glutamic acid. A maximum yield of these products is obtained at 3-8 h, depending upon the pH. An HPLC method has been used for the assay of folic acid and its degradation products. PMID- 10704092 TI - Qualitative and quantitative aspects of the degradation of several tripeptides derived from the antitumour peptide antagonist [Arg(6), D-Trp(7,9), MePhe(8)] substance P[6-11]. AB - The tripeptides Arg-Trp-Phe, Arg-Trp-Phe-NH2, Phe-Trp-Arg and Phe-Trp-Arg-NH2 were subjected to a degradation study to get a more detailed insight into the degradation processes of the antitumor hexapeptide antagonist [Arg(6), D Trp(7,9), MePhe(8)] substance P?6-11? which was investigated in earlier research. Degradation kinetics as well as identities of degradation products of the tripeptides emerging in alkaline and acidic media were studied. The amidated forms (Arg-Trp-Phe-NH2, Phe-Trp-Arg-NH2) appear to be less stable than the carboxylic forms (Arg-Trp-Phe, Phe-Trp-Arg). Deamidation of the amide C-terminus, racemization of the Phe and Arg residues, ornithine formation, hydrolysis of the peptide backbone and diketopiperazine formation with elimination of the N terminal fragments were the major degradative processes. Comparing these reactions with the reactions of antagonist [Arg(6), D-Trp(7,9), MePhe(8)] substance P?6-11? it appeared that racemization of Phe and Arg, hydrolysis of the peptide backbone and diketopiperazine formation did not occur in detectable amounts in the hexapeptide. probably due to lower reaction rates of these reactions compared to the overall degradation rate of antagonist [Arg(6), D Trp(7,9) MePhe(8)] substance P?6-11?. PMID- 10704093 TI - Quantitative determination of dimethicone in commercial tablets and capsules by Fourier transform infrared spectroscopy and antifoaming activity test. AB - Fourier transform infrared (FTIR) spectroscopy and antifoaming activity test have been employed for the quantitative analysis of dimethicone. Linearity, accuracy and precision are presented for both methods. These methods have been also used to compare different dimethicone-containing proprietary medicines. FTIR spectroscopy has shown to be adequate for quantitation of dimethicone in commercial tablets and capsules in order to comply with USP requirements. The antifoaming activity test is able to detect incompatibilities between dimethicone and other constituents. The presence of certain enzymes in some medicinal products increases the defoaming properties of these formulations. PMID- 10704094 TI - Pharmacokinetic and pharmacodynamic analyses of trazodone in rat striatum by in vivo microdialysis. AB - The aim of this study was to investigate the brain pharmacokinetics and pharmacodynamics of trazodone. Sensitive microbore high-performance liquid chromatographic methods with electrochemical detection (LC-ED) were developed for the determination of trazodone, serotonin (5-HT), and their respective metabolites. The feasibility of microdialysis coupled with LC-ED system for direct analysis of these compounds in the rat striatum was investigated. Striatal dialysates were automatically injected onto a cyano microbore column, through an on-line injector, for the determination of trazodone and its metabolite or onto a reversed phase microbore column for the determination of 5-HT and its metabolite. A monophase phenomenon with a first-order elimination rate constant was observed for trazodone. The brain pharmacokinetics of trazodone appear to conform to a one compartment model. Surprisingly, no significant changes in striatal 5-HT or its metabolite were observed following the same dosage and time course. The present results suggest that brain microdialysis methods may be applicable to pharmacokinetic and pharmacodynamic studies of psychotrophic agents. PMID- 10704095 TI - HPLC/atmospheric pressure chemical ionization-mass spectroscopy of eight regulated sulfonamides. AB - Reversed phase high performance liquid chromatography coupled with on-line atmospheric pressure chemical ionization mass spectrometry, HPLC,APCI-MS, has been applied to a mixture of eight sulfonamides. In full scan mode, extracted ion chromatograms produced minimum detectable quantities (MDQ) of 0.8 ng on column, for six of the eight regulated sulfonamides investigated. Selected ion monitoring yielded a 50 pg MDQ for sulfamerazine, sulfadiazine and sulfamethazine, while, the other compounds presented higher values. Analysis of supercritical fluid extracts of chicken liver containing sulfadimethoxine were found to be easily detected by HPLC/APCI-MS. In extracts of chicken liver spiked with 25 microg/kg( 1) (25 ppb) of sulfadimethoxine this compound could be detected in selected ion mode, while 100 pg/microl(-1) was detectable in either full scan or single ion modes. The analysis method for extracted sulfadimethoxine also demonstrated good linearity and reproducibility in both single ion and scan mode. PMID- 10704096 TI - Determination of chloral hydrate metabolites in human plasma by gas chromatography-mass spectrometry. AB - Chloral hydrate (CH) is a widely used sedative. Its pharmacological and toxicological effects are directly related to its metabolism. Prior investigations of CH metabolism have been limited by the lack of analytical techniques sufficiently sensitive to identify and quantify metabolites of CH in biological fluids. In this study a gas chromatography mass spectrometry (GC/MS) method was developed and validated for determining CH and its metabolites, monochloroacetate (MCA), dichloroacetate (DCA), trichloroacetate (TCA) and total trichloroethanol (free and glucuronidated form, TCE and TCE-Glu) in human plasma. Of these, DCA and MCA are newly identified metabolites in humans. The drug, its plasma metabolites and an internal standard, 4-chlorobutyric acid (CBA), were derivatized to their methyl esters by reacting with 12% boron trifluoride methanol complex (12% BF3-MeOH). The reaction mixture was extracted with methylene chloride and analyzed by GC/MS, using a selected ion monitoring (SIM) mode. The quantitation limits of MCA, DCA, TCA, and TCE were between 0.12 and 7.83 microM. The coefficients of variation were between 0.58 and 14.58% and the bias values ranged between -10.03 and 14.37%. The coefficients of linear regression were between 0.9970 and 0.9996. PMID- 10704097 TI - Quantitation of a novel metalloporphyrin drug in plasma by atomic absorption spectroscopy. AB - A bioanalytical method to quantify cobalt mesoporphyrin (CoMP), a novel therapeutic agent, in plasma has been developed and validated. The approach involves atomic absorption spectroscopy to determine total cobalt in a sample and a back-calculation of the amount of compound present. Endogenous plasma cobalt concentrations were small ( <0.2 ng/ml(-1) Co in rat plasma) in comparison to the quantitation limit (4.5 ng/ml(-1) Co). The inter-day imprecision of the method was 10.0% relative standard deviation (RSD) and the inter-day bias was +/- 8.0% relative error (RE) over a standard curve range of 4.5- 45.0 ng/ml(-1) Co. Because it quantifies total cobalt, the method cannot differentiate between parent drug and metabolites, but negligible metabolism allows reliable estimates of the actual parent drug concentration. A correlation study between the atomic absorption method and 14C-radiometry demonstrated excellent agreement (r = 0.9868, slope = 1.041 +/- 0.028, intercept = 223.7 +/- 190.0) and further substantiated the accuracy of the methods. Methodology was successfully applied to a pharmacokinetic study of CoMP in rat, with pharmacokinetic parameter estimation. The elimination half-lives, after intra-muscular and subcutaneous administration, were 7.7 and 8.8 days, respectively. PMID- 10704098 TI - Determination of bis(tributyltin) oxide by GC--MS with on-line hydride derivatization: application to drug substance analysis. AB - We report the determination of residual bis(tributyltin) oxide in a drug substance by GC-MS after extraction and on-line conversion to tributyltin hydride. Gas chromatography was performed using a 15 m x 0.25 mm i.d. DB-5 HT column with a temperature program from 100 to 160 degrees C at 15 degrees C min( 1). A mass range of 165-185 amu was monitored with the MS detector. Hydride generation is performed by placing a small amount of solid sodium borohydride in the injection port of a gas chromatograph and injecting samples and standards through this material. Conversion to tributyltin hydride is shown to be quantitative and linear for levels of bis(tributyltin) oxide between 1 and 100 ppm in the drug substance. The use of GC-MS provides sensitive and selective detection of tin containing species and the tin isotope pattern allows for confirmation of the presence of tin in chromatographic peaks. Recovery at 6 ppm was 89% with an injection precision of 6%. The limit of detection for bis(tributyltin) oxide in drug substance is 1 ppm. PMID- 10704099 TI - A time-resolved fluorescence immunoassay for insulin in rodent plasma. AB - We describe a time-resolved fluoroimmunoassay (TR-FIA) for quantification of insulin in rodent serum and plasma in the picomolar levels typical of these samples. The method is a solid-phase, sequential saturation assay based on competition of unlabeled insulin and biotinamidocaproyl-labeled insulin for anti insulin antibody. Europium-labeled streptavidin allows the DELFIA system (Wallac) to be used for detection. The assay is sensitive (0.1 fmol detection limit, EC50 = 58 +/- 3 pM), accurate ( > 95% recovery of 88-880 pM insulin added to the samples), and simple enough to be automated in a 96-well microtiter plate format. Blood samples of 5 microl can be quickly processed and analyzed within a working concentration range of 40-200 pM, allowing direct measurement of insulin levels in rodents from a tail bleed. We used the TR-FIA to assess insulin levels in mouse and rat samples. In studies of streptozotocin-induced diabetes, as well as glucose load experiments, the assay gave results consistent with known literature. The measured insulin levels correlated significantly with values obtained by radioimmunoassay (R2 = 0.996). The intra-assay and inter-assay coefficients of variation were 2.3% and 15%, respectively. We compared results of this assay with an enzyme-linked immunosorbent assay (ELISA) method. The TR-FIA method was comparable to the ELISA but had higher sensitivity and required only one-tenth as much sample. The assay can be performed using commercially available reagents that allow for high sensitivity and practicability. PMID- 10704100 TI - Direct determination of stability of protease inhibitors in plasma by HPLC with automated column-switching. AB - Automated column-switching HPLC methods were developed and utilized for the direct analyses of three hydroxamic acid based metalloprotease inhibitors in rat plasma. These column-switching methods involved the use of a restricted-access media (RAM) precolumn and a column-switching valve, allowing the complete automation of sample preparation and HPLC. The plasma samples were directly injected onto a precolumn packed with SPS/ODS stationary phase and then backflushed onto an ODS analytical column using a 6-port column-switching device. The drug stability in rat plasma was determined using both the automated and traditional HPLC methods. The results obtained from the automated column switching methods were in good agreement with those from traditional methods that involve sequential protein precipitation, liquid extraction, solvent evaporation, and sample reconstitution. In addition to the elimination of labor-intensive and time-consuming sample preparation procedures, the column-switching methods allowed on-line analyte enrichment and accurate determination of drug stability in plasma with detection limits in the range of 10-20 ng/ml(-1). This work represents, for the first time, a drug stability study in plasma by automated column-switching HPLC technique with the use of a RAM column. Our column switching methods can be readily adapted to any existing HPLC system with minimal hardware modification. PMID- 10704101 TI - Nondestructive tablet hardness testing by near-infrared spectroscopy: a new and robust spectral best-fit algorithm. AB - A new algorithm using common statistics was proposed for nondestructive near infrared (near-IR) spectroscopic tablet hardness testing over a range of tablet potencies. The spectral features that allow near-IR tablet hardness testing were evaluated. Cimetidine tablets of 1-20% potency and 1-7 kp hardness were used for the development and testing of a new spectral best-fit algorithm for tablet hardness prediction. Actual tablet hardness values determined via a destructive diametral crushing test were used for construction of calibration models using principal component analysis/principal component regression (PCA/PCR) or the new algorithm. Both methods allowed the prediction of tablet hardness over the range of potencies studied. The spectral best-fit method compared favorably to the multivariate PCA/PCR method, but was easier to develop. The new approach offers advantages over wavelength-based regression models because the calculation of a spectral slope averages out the influence of individual spectral absorbance bands. The ability to generalize the hardness calibration over a range of potencies confirms the robust nature of the method. PMID- 10704102 TI - A capillary gas chromatographic assay with nitrogen phosphorus detection for the quantification of topiramate in human plasma, urine and whole blood. AB - An accurate and robust method involving liquid liquid extraction and capillary gas chromatographic (GC) assay with nitrogen phosphorus detection (NPD) was developed and validated for the quantitative determination of topiramate [2,3:4,5 bis-O-(-1-methylethylidene)-beta-D-fructopyranose sulfamate], Topamax, an anticonvulsant drug, in human plasma, urine, and whole blood. The galactopyranose analog of topiramate was used as the internal standard. A DB-5, fused silica capillary column (J&W Scientific, Folsom, CA) was used, yielding typical retention times of 4.95 min for topiramate and 5.32 min for the internal standard in human plasma. The assay involved organic extraction with methyl t-butyl ether (MTBE) from base, a back extraction into acid and a second extraction in MTBE. The organic solvent was evaporated, and the residue was redissolved and injected for analysis. The standard curve was validated from 0.5 to 50 microg/ml(-1) for human plasma and whole blood, and from 1.0 to 50 microg/ml(-1) for urine. Peak area ratios of drug to internal standard were determined and used to construct a standard curve. The resulting chromatograms showed no endogenous interfering peaks with the respective blank human fluids. Chromatograms corresponding to topiramate and the internal standard produced sharp peaks that were well resolved. This assay showed precision and accuracy of < or = 5%. Two minor human metabolites of topiramate did not interfere with the assay. This assay was successfully applied to determine the pharmacokinetics of topiramate during the development of this drug. PMID- 10704103 TI - Isolation and identification of process impurities in trimethoprim drug substance by high-performance liquid chromatography, atmospheric pressure chemical ionization liquid chromatography/mass spectrometry and nuclear magnetic resonance spectroscopy. AB - Twenty-two lots of recently synthesized trimethoprim drug substance, from five different manufacturers, in three different countries of origin, China, Israel and the United States, were investigated for the presence of impurities. A liquid chromatographic system, using gradient elution, and a mobile phase consisting of 0.25% TEA/0.1% formic acid (pH 5.8)--acetonitrile, was used to separate and detect two significant, recurring impurities in trimethoprim drug substance. The two impurities were isolated by preparative liquid chromatography and identified, using a combination of liquid chromatography/mass spectroscopy and nuclear magnetic resonance, as 2,4-diamino-5-(4-ethoxy-3,5-dimethoxybenzyl) pyrimidine and 2,4-diamino-5-(3-bromo-4,5-dimethoxybenzyl) pyrimidine. These impurities were not detected by the compendial method and were present at significant levels in 17 of the lots tested. Total impurity concentrations were in the range of 0.1 2.1%. PMID- 10704104 TI - Characterization of the complexation of diflunisal with hydroxypropyl-beta cyclodextrin. AB - The equilibrium dialysis method was applied to the determination of drug cyclodextrin stability constants using diflunisal and 2-hydroxypropyl-beta cyclodextrin (HPBCD) as a model system. Analysis of the data showed the existence of a linear Scatchard plot, indicative of the formation of a 1:1 diflunisal:HPBCD complex. The mean complexation constant (Kc) +/- S.D. was 3,892 +/- 360 M(-1). The stoichiometry of the complex was verified using the appropriate mass action law equation. The diflunisal:HPBCD complex was also investigated using titration microcalorimetry. A Kc of 3,394 M(-1) was obtained together with an enthalpy change (deltaH) of -20.76 kJ/mol(-1). The Kc values obtained here using the equilibrium dialysis and microcalorimetric methods were comparable to one reported previously using a potentiometric method (5,564 +/- 56 M(-1)). PMID- 10704105 TI - Equilibrium distribution of HIV antiviral drugs into human peripheral blood mononuclear cells (PBMC) is controlled by free drug concentration in the extracellular medium. AB - Effect of protein binding on the equilibrium distribution of selected HIV antiviral drugs into isolated human peripheral blood mononuclear cells (PBMC, mainly lymphocytes) was investigated. Human PBMC from a single healthy human donor were isolated, purified, and cryopreserved. Uptake of non-peptide HIV-1 protease inhibitors PNU-96988 and PNU-103017 by these cells in vitro was evaluated as a function of increasing concentration of human serum in the cell incubation media. Both PNU-96988 and PNU-103017 were extensively bound to serum proteins. Uptake/efflux kinetics were very rapid such that accumulation by the cells was thermodynamically, not kinetically, controlled. Accumulation by human PBMCs in vitro was directly proportional to the free and not the total drug concentration in the media. For comparative purposes, the serum protein binding effect on the distribution of two HIV reverse transcriptase (RT) inhibitors, delavirdine (RESCRIPTOR) and zidovudine (AZT), was also evaluated. Like the HIV-1 protease inhibitors, delavirdine was found to be extensively associated with serum proteins and its accumulation by human PBMCs in vitro to be proportional to the free and not total drug concentration. In contrast, AZT was not bound to serum proteins to any significant extent. The uptake of this drug by human PBMCs in vitro was independent of serum concentration. However, the intrinsic cellular accumulation of PNU-96988, PNU-103017 and delavirdine were all greater than AZT. Thus, the extent to which drugs uptake by cells is affected by serum appears proportional to the binding affinity of the serum proteins for the drug. PMID- 10704106 TI - In vivo on-line HPLC-microdialysis: simultaneous detection of monoamines and their metabolites in awake freely-moving rats. AB - An on-line microbore high performance liquid chromatographic--electrochemical (HPLC--EC) detection method has been developed for the simultaneous measurement of dopamine (3,4-dihydroxyphenethylamine, DA), its metabolites 3,4 dihydroxyphenylacetic acid (DOPAC), and homovanillic acid (3-methoxy-4 hydroxyphenylacetic acid, HVA), as well as serotonin (5-hydroxytryptamine, 5-HT) and its metabolise 5-hydroxyindole-3-acetic acid (5-HIAA) in the striatum of awake and freely moving rats. Our method was capable of detecting DOPAC, 5-HIAA, HVA, DA and 5-HT at retention time of 3, 5, 6.5, 9, and 24 min, respectively, in repeated on-line microdialysis sampling. Analysis was performed using a 150 x 1 mm 5 microm C18 microbore column attached directly to a thin-layer radial flow electrochemical cell (UniJet) comprising a 6 mm glassy carbon working electrode. In order to accommodate signal outputs (due to a varying level in the neurochemicals under investigation), an amperometric detector was equipped with a sensitivity programmable controlling software for automatically switching sensitivity inattentively and repeatedly for each collection cycle. PMID- 10704107 TI - Determination of MKT-077, a novel antineoplastic agent, in plasma samples by high performance liquid chromatography and its application to pharmacokinetics in rats. AB - A simple high-performance liquid chromatographic method was developed for determination of a novel antineoplastic agent MKT-077 in plasma. MKT-077 was extracted from 50 microl of plasma with acetonitrile containing 1 ml trifluoroacetic acid per liter. Chromatographic separation was achieved within 13.5 min using a reverse-phase Puresil C18 analytical column. A visible detector operated at 490 nm was used. The linearity of the calibration curve was obtained (r2 = 0.99986) over the analytical range of 10-500 ng/ml(-1). The intra- and inter-assay precision was in the range of 0.9-11.1 and 0.3-4.4%, respectively. The intra- and inter-assay bias ranged from -7.3 to 11.1% and from 0.4 to 11.6%, respectively. The utility of this assay was demonstrated after the administration of a single dose of MKT-077 to rats. The plasma elimination half-life of MKT-077 was 1.8-4 h. PMID- 10704108 TI - Quantitative determination of an extremely polar compound allantoin in human urine by LC-MS/MS based on the separation on a polymeric amino column. AB - Quantitative determination of allantoin in biological matrix poses a challenge to bioanalysis due to the extreme polarity of allantoin. The molecule exhibits virtually no retention on any of the available hydrophobic HPLC packing materials. In this study, an assay was developed for the LC-MS/MS quantitation of allantoin in human urine using a polymeric amino column to achieve the necessary separation. The urine samples were prepared by a single-step solid phase extraction (SPE) procedure. The extracted samples were analyzed on a jordi-gel DVB polyamine column (operated under an acetonitrile/water gradient with water as the stronger mobile phase) interfaced with a Finnigan TSQ 7,000 mass spectrometer. Negative electrospray ionization (ESI) was employed as the ionization source. Allantoin and its internal standard (13C1, 15N1-allantoin) were detected by use of single reaction monitoring (SRM) mode. The method was validated in the concentration range of 14.6-213 microg/ml(-1), with within and between run accuracy and precision both < 7%. The method has been successfully applied to clinical sample analysis. PMID- 10704109 TI - Determination of silymarine in the extract from the dried silybum marianum fruits by high performance liquid chromatography and capillary electrophoresis. AB - Silybine (SBN), isosilybine (ISBN), silycristine (SCN), silydianine (SDN), and taxifoline (TXF) are the main active flavanoids, generally found in the dried fruits of silybum marianum. The concentrations of these compounds, excepted TXF, are all together usually expressed as silymarine content. In this paper the determination of the silymarine titre was made by high performance liquid chromatography (HPLC), and high performance capillary electrophoresis (HPCE). Two reversed stationary phases, RP-18 and RP-8, were observed comparing the resolutions of all considered flavanoids with each stationary phases. The HPCE was carried out considering the possible improvement in the resolution of SBN, CN, SDN and TXF using, 8-cyclodextrines or organic modifier. The qualitative and quantitative data obtained by HPLC and HPCE were compared. PMID- 10704110 TI - Use of artificial neural networks to predict quaternery phase systems from limited experimental data. AB - The aim of the present work was to develop a method for predicting the phase behaviour of four component systems consisting of oil, water and two surfactants from a limited number of screening experiments. Investigations were conducted to asses the potential of artificial neural networks (ANNs) with back-propagation training algorithm to predict the phase behaviour of four component systems. Three inputs only (percentages of oil and water and HLB of the surfactant blend) and four outputs (oil in water emulsion, water in oil emulsion, microemulsion, and liquid crystals containing regions) were used. Samples used for training represented about 15% of the sampling space within the tetrahedron body. The network was trained by performing optimization of the number and size of the weights for neuron interconnections. The lowest error was obtained with 15 hidden neurons and after 4,500 training cycles. The trained ANN was tested on validation data and had an accuracy of 85.2-92.9%. In most cases the errors in the prediction were confined to points lying along the boundaries of regions and for the extrapolated predictions outside the ANNs 'experience'. This approach is shown to be highly successful and the ANNs have proven to be a useful tool for the prediction of the phase behaviour of quaternary systems with less experimental effort. PMID- 10704111 TI - Tissue distribution of polaprezinc in rats determined by the double tracer method. AB - The tissue distribution of polaprezinc (an insoluble zinc complex of L-carnosine) in rats was studied by the double tracer method using [U-14C-histidine]-, 65Zn polaprezinc. The 65Zn-radioactivity was measured with an auto-gamma counter, and the 14C containing 65Zn was converted to an absolute count according to the calibration curve for quenching with a liquid scintillation counter with the spill-over method. After the administration of 14C-, 65Zn-polaprezinc to rats, the excretion ratio and time courses in the tissues of the 14C-and 65Zn radioactivity were different each other. We found that polaprezinc was metabolized as endogenous amino acid or zinc after dissociation in the body. The zinc concentration in plasma reached its maximum at 1 h and decreased slowly, returning to the endogenous level at 11 h after the administration of non-labeled polaprezinc. The concentrations of zinc in liver, kidney, testis, prostate, and cerebrum remained rather constant. The replacement ratios of 65Zn to zinc in the tissues at its maximum percentage were 40% in plasma, 16-20% in liver, kidney, blood, and prostate. The low replacement ratios in testis and cerebrum (2-3%) suggested that zinc uptakes in testis and brain were regulated by the blood testis-barrier and blood-brain-barrier, respectively. PMID- 10704112 TI - Automated determination of 'Ecstasy' and amphetamines in urine by SPME and capillary gas chromatography after propylchloroformate derivatisation. AB - The determination of amphetamines and their methylenedioxylated analogs in urine by propylchloroformate derivatisation and automated solid-phase microextraction is described. The urine sample was adjusted to pH 10.8 and added propylchloroformate reagent and an internal standard. Derivatisation resulted in water-stable carbamates which were automatically extracted by solid-phase microextraction. A fiber coated with polydimethylsiloxane was inserted into the urine matrix and agitated for 16 min. The fibre with the extracted carbamates was injected into the heated split-splitless injection port of the gas chromatograph where the analytes were evaporated at 300 degrees C, separated on a methylsilicone capillary column and detected by either a nitrogen phosphorous detector or by mass spectrometry. The method was shown to be highly reproducible and robust with respect to variations in the urine matrices. The detection limits were 5 ng/ml(-1) of methamphetamine, MDMA and MDEA and 15 ng/ml(-1) of amphetamine and MDA in urine. The method is a solvent free, automated alternative to traditional methods for determination of the amphetamine and their methylendioxylated analogs in urine. PMID- 10704113 TI - Simultaneous determination of hydrochlorothiazide and amiloride hydrochloride by ratio spectra derivative spectrophotometry and high-performance liquid chromatography. AB - Rapid, precise, accurate and specific ratio spectra derivative spectrophotometry and high-performance liquid chromatographic procedures were described for the simultaneous determination of hydrochlorothiazide and amiloride hydrochloride in combined pharmaceutical dosage forms. For the first method, ratio spectra derivative spectrophotometry, the signals were measured at 285.7 nm for hydrochlorothiazide and at 302.5 nm for amiloride hydrochloride in the mixture, in the first derivative of the ratio spectra. The second method is based on high performance liquid chromatography (HPLC) on LiChrosorb RP-C18 column (5 microm, 20 cm x 4.6 mm) using 0.025 M orthophosphoric acid (adjusted to pH 3.0 with triethylamine (TEA)), acetonitrile (84:16 v/v) as a mobile phase at a flow rate of 1.2 ml/min(-1). Detection was carried out using a UV detector at 278.0 nm. Commercial sugar-coated and laboratory-prepared mixtures containing both drugs in different proportions were assayed using the developed methods. PMID- 10704114 TI - Enantiomeric separation of optically active pyridazinone derivatives by chiral HPLC. AB - Several 4,5-disubstituted 3(2H)-pyridazinone derivatives containing 2 hydroxymethylpyrrolidino moiety as a chiral building block were synthetized. Separation of enantiomers was carried out by chiral HPLC on Chiralcel OJ and OF columns. Mobile phases consisted of hexane, ethanol and 2-propanol. Chiralcel OJ column was capable of separating most of the enantiomeric pairs. For one type of compound. Chiralcel OF column was used for separation. PMID- 10704116 TI - Study of solution equilibria between aluminum(III) ion and ofloxacin. AB - The complex formation equilibria between aluminium(III) ion and ofloxacin in 0.1 mol/l(-1) ionic medium at 298 K were studied by glass electrode pH-metric and UV spectrophotometric measurements. Within ofloxacin to aluminium mole ratio ranging from 2:1 to 25:1 and in pH interval from 2.5 to 10.5, the obtained experimental results were explained by the formation of the following complexes: AI(Hoflo) (log beta1,1,1 = 15.93 +/- 0.03), Al(oflo)2 (log beta1,2,0= 14.84 +/- 0.07), Al(oflo) (log beta1,1,0 = 10.20 +/- 0.04) as well as several other mixed and pure hydrolytic complexes. The structure and mechanism of the formation of the complexes and their possible implications on aluminum toxicity were discussed. PMID- 10704115 TI - Determination of TA-0201, a novel orally active non-peptide endothelin antagonist, in rat plasma and tissues by a liquid chromatography--electrospray ionization--tandem mass spectrometry system. AB - N-[6-[2-[(5-bromo-2-pyrimidinyl)oxy]ethoxy]-5-(4-methylphenyl)-4-pyrimid inyl]-4 (2-hydroxy-1, 1-dimethylethyl) benzenesulfonamide sodium salt (TA-0201) is a novel orally active non-peptide antagonist for endothelin (ET) receptors. A sensitive and simultaneous determination method of TA-0201 and its major metabolites by liquid chromatography tandem mass spectrometry (LC--MS/MS) in a selected reaction monitoring mode using [2H6]TA-0201 as the internal standard was developed, and the plasma and tissue concentrations of TA-0201 and its major metabolites were determined in a pharmacokinetic study. The lower limit of determination of plasma TA-0201 concentrations by this method was 0.1 ng/0.5 ml, and the between- and within-run accuracy and precision were both less than 5.1%, in the calibration curve range of 0.1-50 ng/0.5 ml. This method was applied to determine the concentrations of TA-0201 and its metabolites in plasma and various target tissues, i.e. the heart, lung and kidney, after oral administration of TA 0201 (0.1 mg/kg(-1)) to male rats. TA-0201 and its major metabolite of a carboxylic acid form were detected in plasma and all the tissues 24 h after administration, their tissue concentrations being higher than those in plasma and still detectable at 72 h. Thus, this method could successfully be applied to study pharmacokinetic properties of TA-0201 in rats. PMID- 10704117 TI - Impurity profiling in bulk pharmaceutical batches using 19F NMR spectroscopy and distinction between monomeric and dimeric impurities by NMR-based diffusion measurements. AB - The impurity profile of production batches of fluorine-containing drugs can be characterised efficiently using 19F NMR spectroscopy. This yields the number and proportions of impurities in the bulk drug to a level of approximately equal 0.1 mole% in a few minutes of NMR experiment time. The approach has been exemplified using a partially purified batch of the steroidal product fluticasone propionate, the impurities in which include a number of dimeric species. Further distinction between the monomer and dimer impurities has been achieved through high resolution chemical shift-resolved NMR measurement of molecular diffusion coefficients on the intact mixture using 19F NMR spectroscopy. The ability of NMR based diffusion coefficient determination to distinguish between monomeric and dimeric substances was validated using a standard mixture of authentic materials containing both monomers and dimers. PMID- 10704118 TI - Direct enantioseparation of adrenergic drugs via thin-layer chromatography using molecularly imprinted polymers. AB - Molecularly imprinted polymers (MIPs) of (-)-pseudoephedrine and (-)-norephedrine were prepared to use as chiral stationary phases (CSPs) in thin layer chromatography (TLC). The resolution of the enantiomers of adrenergic drugs, including pseudoephedrine, ephedrine, norephedrine, and epinephrine were investigated on these CSPs. In preparation of MIPs, two monomers: (1) methacrylic acid and (2) itaconic acid were employed as functional monomers. Mobile phase system of either methanol or acetonitrile was used and the effects of acetic acid content of the mobile phases were also investigated. The best resolution was achieved for enantioseparation of norephedrine on plates based on MIP of (-) norephedrine using itaconic acid as functional monomer (alpha = 5.1) in mobile phase 1% acetic acid in methanol. Moreover, these MIPs were able to resolve the racemates of compounds whose structures corresponded to print molecule. The results obtained showed that TLC based on MIPs could succeed the direct separation of enantiomers of adrenergic drugs as a method of separation. The method offers a rapid, sensitive and reliable method for quality control of optically active compounds. PMID- 10704119 TI - Modelling of conditions for the enantiomeric separation of beta2-adrenergic sympathicomimetics by capillary electrophoresis using cyclodextrins as chiral selectors in a polyethylene glycol gel. AB - A two-factor central composite design was used to determine a mathematical model for prediction of the optimal conditions for the separation of the enantiomers of some widely used beta2-sympathicomimetic drugs (beta2-agonists) by capillary electrophoresis using cyclodextrins (CD) as a chiral selector in a polyethylene glycolgel. The effects of the chemical structure of these drugs along with the addition of polyethylene glycol to the cyclodextrin solution on the resolution of their enantiomers were studied. To allow impurity studies down to 0.1% (distomer eutomer) a resolution of 2.5 should be warranted. Those beta2-agonists containing two hydroxylic groups in the aromatic ring structure show the highest enantiomeric separation, due to the fact that one of their enantiomers has a better geometric structure to fit into the beta-cyclodextrin cavity. PMID- 10704120 TI - LTB4 as marker of 5-LO inhibitory activity of two new N-omega-ethoxycarbonyl-4 quinolones. AB - The supposed 5-LO inhibitory activity of two N-omega-ethoxycarbonyl-4-quinolones was tested determining leukotriene B4 (LTB4) in RBL-1 cell cultures, pretreated with the two compounds of interest. LTB4, obtained by solid-phase extraction (SPE) from cell cultures supernatants, was determined by micellar electrokinetic chromatography (MEKC). The analysis was performed using an uncoated capillary, filled with borate buffer at pH 8.3, containing 12.5 mM SDS as micelles generator. Therefore, following the decreasing of LTB4 it was possible to verify the 5-LO inhibitory activity of two quinolone derivatives. To asses the suitability of the use of LTB4 as marker of the activity of the new compounds, the analysis was repeated using quercetin, a well known 5-LO inhibitor. PMID- 10704121 TI - The determination of busulphan in dissolution samples by flow injection analysis. AB - A robust colourimetric flow injection analysis (FIA) procedure is described for the determination of busulphan in dissolution samples of a 2 mg tablet formulation. The sample solution is injected directly into a reagent stream containing 4-(4-nitrobenzyl)pyridine/potassium hydrogen phthalate. An on-line heating stage allows the formation of a coloured pyrridinium salt species, which following stabilisation is detected spectrophotometrically at 570 nm. The method has been fully validated and is linear over the concentration range 0.004-0.024 mg of busulphan ml(-1). The method can also been applied to uniformity of content and bulk assay testing. PMID- 10704122 TI - Direct high-performance liquid chromatographic determination of the enantiomeric purity of levodopa and methyldopa: comparison with pharmacopoeial polarimetric methods. AB - Chiral high-performance liquid chromatography was employed for determination of the enantiomeric purity of levodopa and methyldopa. The determination of D-DOPA in levodopa was accomplished using a chiral ligand-exchange chromatograpy with an ordinary C18 column and a chiral mobile phase containing N,N-dimethyl-L phenylalanine and Cu(II) acetate or by means of LC on a teicoplanin column in conjunction with ethanol-water (65:35, v/v). Both methods gave good performance, however, the latter was faster and more convenient and suitable for routine analyses. For the determination of D-methyldopa a LC method based on the use of a teicoplanin column in polar organic mode with methanol-acetic acid-triethylamine (1,000:0.05:0.05, v/v/v) mobile phase was developed. The precision, accuracy, linearity and selectivity were satisfactory. In comparison with pharmacopoeial polarimetric methods (according to the European Pharmacopoeia and the Pharmacopoea Bohemoslovaca), the LC methods proved to be much more sensitive giving detection limits 0.04% of D-DOPA and 0.3% of D-methyldopa. PMID- 10704123 TI - A method for routine analysis of recombinant immunoglobulins (rIgGs) by capillary isoelectric focusing (cIEF). AB - A capillary isoelectric focusing (cIEF) method was developed for routine analysis of recombinant immunoglobulins (rlgGs). The cIEF method used a dimethyl siloxane coated capillary and a separation matrix of 2% ampholytes in 0.4% methylcellulose (MC). The rIgGs, and internal pI marker protein standards, were mixed with carrier ampholyte in MC, focused using high voltage, and then the protein bands were mobilized past a UV detector by simultaneous application of low pressure and voltage. Qualitatively and quantitatively equivalent rIgG focusing profiles were obtained via cIEF and gel-based IEF, with individual isoform peak area percentages and calculated peak pI values being comparable for the same samples. Linear relationships were obtained for peak area response versus sample concentration, and for the pI gradient developed between the internal pI marker standards. The relative standard deviation (RSD) in rIgG peak areas was less than 2% intra-day and less than 8% inter-day (72 h). The RSD for the mobilization times of rIgG peaks was less than 1% intra-day and less than 3% inter-day (72 h). There was no observed decrease in the performance of the capillary over 150 analyses. cIEF offers several important advantages over gel IEF, e.g. direct, quantitative detection of proteins by intrinsic UV absorbance at 280 nm, rapid analyses ( < or = 30 min), capability of automation, and one-step, electronic data analysis and archival. These data demonstrate the superiority of the cIEF method for routine analysis of rIgGs. PMID- 10704124 TI - Mexiletine-sensitive membrane electrode for medical application. AB - Response characteristics of mexiletine-sensitive membrane electrodes based on crown ether and ion-exchanger were examined in a physiological saline in order to find an electrode suitable for determining concentrations of this drug under physiological conditions. Among various crown ethers screened, 4',4"(5")-di-tert butyldicyclohexano-18-crown6 showed the highest sensitivity to mexiletine in physiological saline containing 0.15 M NaCl and 5 mM 4-(2-hydroxyethyl)-2 piperazineethanesulfonic acid (Hepes) NaOH (pH 7.4). However, the detection limit of 30 microM was 10 times higher than that of 3 microM observed with the electrode based on an ion-exchanger, sodium tetrakis[3,5-bis(2-methoxyhexafluoro 2-propyl)phenyl]borate. Having high selectivity against inorganic cations such as Na+ or K+, the electrode using the ion-exchanger enabled us to determine the level of mexiletine in saliva, the monitoring of which is quite effective for controlling the dose of this drug noninvasively. The mexiletine concentrations determined with the mexiletine electrode compared favourably with those determined by high-performance liquid chromatography which requires an additional procedure to extract mexiletine from saliva. PMID- 10704125 TI - Determination of risperidone and 9-hydroxyrisperidone in human plasma by high performance liquid chromatography: application to therapeutic drug monitoring in Japanese patients with schizophrenia. AB - A high-performance liquid chromatographic method has been developed for the simultaneous determination of risperidone and its major active metabolite 9 hydroxyrisperidone in plasma. Risperidone and 9-hydroxyrisperidone in plasma were extracted using a CN bonded-solid phase cartridge, followed by, C4 reversed-phase HPLC separation. Risperidone, 9-hydroxyrisperidone and trazodone as an internal standard were detected by ultraviolet absorbance at 280 nm. It was possible to determine risperidone in the concentration range of 1.0-100.0 ng/ml(-1) and 9 hydroxyrisperidone at a range of 2.0-200.0 ng/ml(-1). The detection limits of risperidone and 9-hydroxyrisperidone were 0.5 and 1.0 ng/ml(-1), respectively. The mean recoveries of risperidone and 9-hydroxyrisperidone added to plasma were less than 92.0 and 92.6%, with a coefficient of variation of less than 10.6 and 10.5%, respectively. This method has been used for the simultaneous determination of steady-state plasma concentration (Css) of risperidone and 9 hydroxyrisperidone in schizophrenic patients treated with 3-, 6-, and 12-mg risperidone oral doses per day. PMID- 10704127 TI - Automated solid-phase extraction workstations combined with quantitative bioanalytical LC/MS. AB - An automated solid-phase extraction workstation was used to develop, characterize and validate an LC/MS/MS method for quantifying a novel lipid-regulating drug in dog plasma. Method development was facilitated by workstation functions that allowed wash solvents of varying organic composition to be mixed and tested automatically. Precision estimates for this approach were within 9.8% relative standard deviation (RSD) across the calibration range. Accuracy for replicate determinations of quality controls was between -7.2 and +6.2% relative error (RE) over 5-1,000 ng/ml(-1). Recoveries were evaluated for a wide variety of wash solvents, elution solvents and sorbents. Optimized recoveries were generally > 95%. A sample throughput benchmark for the method was approximately equal 8 min per sample. Because of parallel sample processing, 100 samples were extracted in less than 120 min. The approach has proven useful for use with LC/MS/MS, using a multiple reaction monitoring (MRM) approach. PMID- 10704126 TI - Simultaneous high-performance liquid chromatographic determination of puerarin, daidzin, paeoniflorin, liquiritin, cinnamic acid, cinnamaldehyde and glycyrrhizin in Kampo medicines. AB - We report a high-performance liquid chromatographic method to determine the quantities of puerarin, daidzin, paeoniflorin, liquiritin, cinnamic acid, cinnamaldehyde and glycyrrhizin in Kampo medicine. All seven compounds were separated in less than 30 min with a Wakosil-II 5C18 AR column by linear gradient elution using 0.01% (v/v) phosphoric acid acetonitrile (0 min 90:10, 10 min 88:12, 22 min 70:30, 30 min 30:70) as the mobile phase at a flow-rate of 1.0 ml/min(-1), and detection at 250 nm. The detection limits of these compounds are 0.15-0.3 microM with response linearity. This method was applied to determine the quantities in eight Kampo decoctions; Mao-to, Makyo-yokukan-to, Makyo-kanseki-to, Yokuinin-to, Sho-seiryu-to, Keima-kakuhan-to, Kakkon-to and Kakkon-to-ka-senkyu sin'i. Glycyrrhizin content was lower in both the decoction and the methanol diluted decoction of Sho-seiryu-to compared with the others. Low pH due to organic acids of Schisandrae fructus in the decoction caused inhibition for glycyrrhizin dissolution in Sho-seiryu-to. PMID- 10704128 TI - Simultaneous determination of oxalic, fumaric, maleic and succinic acids in tartaric and malic acids for pharmaceutical use by ion-suppression reversed-phase high performance liquid chromatography. AB - A reliable method for the simultaneous determination of oxalic, fumaric, maleic, and succinic acids in tartaric and malic acids for pharmaceutical use by reversed phase ion-suppression high performance liquid chromatography is presented. HPLC was achieved on a Nova-Pak C18 column by isocratic elution using water adjusted to pH 2.10-2.15 with perchloric acid, and detection was by UV adsorption at a wavelength of 210 nm. This method was found to be superior to previous liquid chromatography as well as other classical assay, and to be an attractive choice for the analysis of these compounds. PMID- 10704129 TI - Effects of surfactants on the spectral behaviour of calcein (II): a method of evaluation. AB - The spectral behavior of calcein, a water-soluble self quenching fluorescent marker often used in biomedical analysis, can be considerably affected by the presence of surfactants. With this study we intend to obtain further information on the photophysical properties of calcein, in the presence of surfactants and in the concentration range commonly used to investigate the release of such marker from vesicle dispersions. The experiments were carried out both in water and in a physiological buffer (HEPES, pH 7.5), in the presence of Triton X-100, sodium dodecyl sulphate and centyltrimethylammonium bromide, both below and above their critical micelle concentration (c.m.c.). The obtained results confirm that calcein fluorescence can be affected by the presence of surfactants. Thus, environmental conditions must always be carefully checked for the actual quantitative evaluation of this dye. Furthermore, this study sheds some light on the nature and mechanism of calcein quenching. PMID- 10704130 TI - Enzymatically modified ion-selective electrodes for flow injection analysis. AB - The application of potentiometric biosensors based on membrane ion-selective electrodes for flow injection analysis (FIA) is presented. The biosensors have been obtained by covalent binding of enzyme molecules directly to surface of ion selective membranes. The biosensor/FIA systems enable the determination of urea and penicillin in the millimolar ranges of concentration. About 30 samples per hour can be analysed in the performed FIA systems. The operational stability of the bioanalytical systems exceeds 1 month. PMID- 10704131 TI - UV-spectrophotometry and square wave voltammetry at nafion-modified carbon-paste electrode for the determination of doxazosin in urine and formulations. AB - By using several electrochemical techniques, the study of electroanalytical behaviour of antihipertensive Doxazosin at Nafion modified carbon paste electrode (NMCPE) has been carried out. The voltammetric peak is very pH dependent, reaching the maximum i(p) at pH 6.8 (Ep -0.17 V), the reduction process being quasi-reversible and fundamentally controlled by adsorption. A method based on the control of adsorptive preconcentration of the Doxazosin on the NMCPE, before its voltammetric determination, is proposed. The detection limit reached using square wave voltammetry (SWV) as redissolution technique was 2.33x10(-11) M and the variation coefficient at 2x10(-9) M level was 3.54%. A spectrophotometric study of Doxazosin has also been made and two waves at 244 and 329 nm (pH 1.7), were obtained. The wave at 329 nm changes its height and position with the pH, allowing the pKa determination (6.94+/-0.21) using different methods. The obtained detection limit was 0.5x10(-6) M, and the variation coefficient at 1.5x10(-5) M level was 0.99%. The UV spectrophotometric method is sufficiently accurate and precise to be applied in the Carduran tablets assay, while the voltammetric method (AdS-SWV) can in addition be used to determine the drug at trace level in human urine samples with good recoveries. PMID- 10704132 TI - PH-metric log P 11. pKa determination of water-insoluble drugs in organic solvent water mixtures. AB - The apparent acid dissociation constants (p(s)Ka) of two water-insoluble drugs, ibuprofen and quinine, were determined pH-metrically in acetonitrile water, dimethylformamide water, dimethylsulfoxide water, 1,4-dioxane-water, ethanol water, ethylene glycol-water, methanol water and tetrahydrofuran water mixtures. A glass electrode calibration procedure based on a four-parameter equation (pH = alpha + SpcH + jH[H+]+jOH[OH-]) was used to obtain pH readings based on the concentration scale (pcH). We have called this four-parameter method the Four Plus technique. The Yasuda Shedlovsky extrapolation (p(s)Ka + log [H2O] = A/epsilon + B) was used to derive acid dissociation constants in aqueous solution (pKa). It has been demonstrated that the pKa values extrapolated from such solvent water mixtures are consistent with each other and with previously reported measurements. The suggested method has also been applied with success to determine the pKa values of two pyridine derivatives of pharmaceutical interest. PMID- 10704133 TI - Selective spectrofluorimetric determination of phenolic beta-lactam antibiotics through the formation of their coumarin derivatives. AB - A simple, sensitive and selective spectrofluorimetric procedure was developed for the determination of amoxycillin, cefadroxil and cefoperazone. The method is based on the reaction between these drugs and ethyl acetoacetate, in acidic medium, to give yellow fluorescent products with excitation wavelengths ranging from 401 to 467 nm and emission wavelengths ranging from 465 to 503 nm. The reaction conditions were studied and optimized. The reaction obeyed Beer's law over the range of 10.0-20.0, 1.5-1.0 and 50.0-100.0 microg ml(-1) for amoxycillin, cefadroxil and cefoperazone, respectively. Interference's from other antibiotics, drugs and dosage forms additives, in capsules and vials dosage forms, were investigated. The proposed method was applied to the analysis of pharmaceutical formulations (capsules and vials) containing the above antibiotics, either alone or in combination with other antibiotics or drugs. The validity of the method was tested by the recovery studies of standard addition which were found to be satisfactory. The results of the proposed method demonstrated that the method is equally accurate, precise and reproducible as the official methods (USP XXIII) and those published for the non-official binary mixtures. PMID- 10704134 TI - Spectrophotometric and electrochemical study of protolytic equilibria of some oximes-acetylcholinesterase reactivators. AB - Newly synthesized oximes, mono and bis imidazole derivatives, which promise to be more effective acetylcholinesterase reactivators than standard antidotes used, were investigated by spectrophotometric and electrochemical methods. The electrochemical investigations confirmed the existence of overlapping equilibria, obtained by spectrophotometric methods. Dissociation constants of those oximes were also obtained by numerical treatment of overlapping equilibria, using the Lavendberg Marquardt least square method, and when compared with the same for some similar compounds, were found to be very effective acetylcholinesterase reactivators. The distribution of ionic forms of the investigated oximes, as a dependence of pH values, was calculated from the obtained values of dissociation constants. The results indicated that many oxime anions will be available at physiological pH 7.4 and a relative increased ability to reactivate inhibited acetylcholinesterase could be expected. PMID- 10704135 TI - Simultaneous determination of some multicomponent dosage forms by quantitative thin layer chromatography densitometric method. AB - In this study, simultaneous determination of naproxen with diflunisal (mixture I), paracetamol with chlorzoxazone (mixture II) and chlorphenoxamine hydrochloride with 8-chlorotheophylline and caffeine (mixture III) in multicomponent mixtures was conducted by a thin layer chromatography densitometric method. The mobile phase ethyl acetate: methanol: ammonia 25% (85:15:5 v/v) was used for the separation of the components of mixtures (I) and (II) with Rf values of 0.16 for naproxen, 0.4 for diflunisal, 0.77 for paracetamol and 0.32 for chlorzoxazone. Efficient separation of the components of mixture (III) was attained using ethyl acetate as mobile phase with Rf values of 0.12, 0.62 and 0.42 for chlorphenoxamine hydrochloride, 8-chlorotheophylline and caffeine, respectively. Linearity ranges, mean recoveries and relative standard deviations in calibration graphs of the proposed method were calculated. The method has been successfully applied to pharmaceutical formulations, sugar-coated tablets, capsules and suppositories. The results obtained were statistically compared with those obtained by applying the reported alternate methods. PMID- 10704136 TI - Determination of some antihistaminic drugs by atomic absorption spectrometry and colorimetric methods. AB - Atomic absorption spectrometry (AAS) and colourimetric methods have been developed for the determination of pizotifen (I), ketotifen (II) and loratadine (III). The first method depends on the reaction of the three drugs (I); (II) and (III) with cobalt thiocyanate reagent at pH 2 to give ternary complexes. These complexes are readily extracted with organic solvent and estimated by indirect atomic absorption method via the determination of the cobalt content in the formed complex after extraction in 0.1 M hydrochloric acid. It was found that the three drugs can be determined in the concentration ranges from 10 to 74, 12 to 95 and 10 to 93 microg ml(-1) with mean percentage recovery of 99.71+/-0.87, 99.70+/ 0.79 and 99.62+/-0.75%, respectively. The second method is based on the formation of orange red ion pairs as a result of the reaction between (I); (II) and (III) and molybdenum thiocyanate with maximum absorption at 469.5 nm in dichloromethane. Appropriate conditions were established for the colour reaction. Under the proposed conditions linearity was obeyed in the concentration ranges 3.5-25, 5-37.5 and 2.5-22.5 microg ml(-1) with mean percentage recovery of 99.60+/-0.41, 100.11+/-0.43 and 99.31+/-0.47% for (I): (II) and (III), respectively. The third method depends on the formation of radical ion using 2,3 dichloro-5,6-dicyano-p-benzoquinone (DDQ). The colour formed was measured at 588 nm for the three drugs (I); (II) and (III), respectively. The method is valid in concentration range 10-80 microg ml(-1) with mean percentage recovery 99.75+/ 0.44, 99.94+/-0.72 and 99.17+/-0.36% for (I); (II) and (III), respectively. The proposed methods were applied to the analysis of pharmaceutical preparations. The results obtained were statistically analysed and compared with those obtained by applying the official and reference methods. PMID- 10704137 TI - Simultaneous determination of rifampicin, isoniazid and pyrazinamide in tablet preparations by multivariate spectrophotometric calibration. AB - The use of multivariate spectrophotometric calibration is presented for the simultaneous determination of the active components of tablets used in the treatment of pulmonary tuberculosis. The resolution of ternary mixtures of rifampicin, isoniazid and pyrazinamide has been accomplished by using partial least squares (PLS-1) regression analysis. Although the components show an important degree of spectral overlap, they have been simultaneously determined with high accuracy and precision, rapidly and with no need of nonaqueous solvents for dissolving the samples. No interference has been observed from the tablet excipients. A comparison is presented with the related multivariate method of classical least squares (CLS) analysis, which is shown to yield less reliable results due to the severe spectral overlap among the studied compounds. This is highlighted in the case of isoniazid, due to the small absorbances measured for this component. PMID- 10704138 TI - In vitro metabolic products of RWJ-34130, an antiarrythmic agent, in rat liver preparations. AB - The in vitro metabolism of RWJ-34130, an antiarrhythmic agent, was conducted using rat hepatic 9000 x g supernatant (S9) and microsomes in an NADPH-generating system, and the rat liver perfusion. The 100 and 20 microg ml(-1) concentrations of RWJ-34130 aqueous solution were used for microsomal incubation and liver perfusion, respectively. Unchanged RWJ-34130 (approximately 77-78% of the sample in both S9 and microsomes) plus a major metabolite, RWJ-34130 sulfoxide (20% of the sample in both S9 and microsomes) were profiled, isolated and identified from both hepatic S9 and microsomal incubates (60 min) using HPLC and mass spectrometry (MS), and by comparison to a synthetic RWJ-34130 sulfoxide, which was synthesized by reacting RWJ-34130 with MCPBA (meta-chloroperoxy benzoic acid). No unchanged RWJ-34130 was detected in the 3 h liver perfusate, however, 1 phenyl-2-oxo-pyrrolidine was profiled, isolated and identified as a major hydrolyzed metabolite of liver perfusate. RWJ-34130 is not extensively metabolized in vitro in rat hepatic S9 and microsomes. All HPLC metabolic profiles of hepatic S9 and microsomal samples (30 min, 60 min) were qualitatively and nearly quantitatively identical. PMID- 10704139 TI - A rapid and reliable solid-phase extraction--LC/MS/MS assay for the determination of two novel human leukocyte elastase inhibitors, SYN-1390 and SYN-1396, in rat plasma. AB - A rapid and rugged solid-phase extraction-liquid chromatographic-electrospray tandem mass spectrometric method has been developed to quantitate two novel human leukocyte elastase inhibitors, SYN-1390 and SYN-1396, in rat plasma. A reversed phase column and an isocratic mobile phase consisting of acetonitrile-water formic acid (70:30:0.2, v/v/v) were used. The mass spectrometer was operated in the multiple reaction monitoring mode. For both analytes, standard curves were linear over a working range of 0.1-20 microg ml(-1) (r> or =0.995) and the limit of quantitation was 0.1 microg ml(-1) with a 150 microl plasma volume. This assay proved to be useful for the determination of SYN-1390 and SYN-1396 in plasma samples from pharmacokinetic study. PMID- 10704140 TI - Development and validation of an HPLC assay for fentanyl and related substances in fentanyl citrate injection, USP. AB - The stability indicating properties of the USP method for the assay of fentanyl in fentanyl citrate injection were evaluated [1] by analyzing fentanyl drug substance and product after acid, hydrogen peroxide, heat, and light treatment. N phenyl-N-(4-piperidinyl)propionamide (PPA), which is a known degradation product/process impurity of fentanyl, was not adequately resolved from the fentanyl peak, and mobile phase adjustments did not improve the resolution (Fig. 1). Therefore, the USP method did not meet the requirements for a stability indicating assay. In addition, the wavelength in the USP method was too high (230 nm) to provide adequate levels for the quantitation of the related substances of fentanyl and, in addition, the acetate ions in the mobile phase could interfere with a lower wavelength detection. An isocratic, reversed phase, stability indicating, high performance liquid chromatographic (HPLC) method for the assay of fentanyl and related substances in fentanyl citrate injection, USP has been developed and validated. The chromatographic conditions employed an Inertsil C8, 5 column (25 cm x 4.6 mm), a mobile phase of aqueous perchloric acid [0.23%, w/v] acetonitrile [65:35, v/v], and ultraviolet (UV) detection at 206 nm. Under the chromatographic conditions of the method, PPA and seven other known process impurities were separated from the active. Degradation studies showed that the active eluted as a spectrally pure peak resolved from its degradation products. PMID- 10704141 TI - A reverse phase ion-pairing HPLC method for the stability monitoring of sulphacetamide ophthalmic preparations. PMID- 10704142 TI - Polarographic determination of benznidazole in DMSO. PMID- 10704143 TI - Inhibitors of complement activity in human breast-milk: a proposed hypothesis of their physiological significance. AB - Several natural components abundant in the fluid phase of human breast-milk have been shown to be inhibitors of complement activation in vitro, particularly the classical pathway. These include lysozyme, lactoferrin, lactalbumin alpha and other ligand chelators, complement regulator proteins and other specific soluble inhibitors of complement activation. Their physiological significance probably resides in their ability to restrict in vivo complement activation to specialized (compartmentalized) sites on the cellular membrane structures in human milk, represented by the abundant surface area of the milk fat globule membranes. This would serve to prevent inflammatory-induced tissue damage of the delicate immature gastrointestinal tract of the newborn as well as the mammary gland itself. A number of recognized and potential inhibitors of complement activity in human milk and other biological fluids are hereby reviewed, with a proposal of their physiological significance. PMID- 10704144 TI - Upregulation of ICAM-1 expression on J774.2 macrophages by endotoxin involves activation of NF-kappaB but not protein tyrosine kinase: comparison to induction of iNOS. AB - This study compares the signal transduction pathway which leads to the upregulation of intercellular adhesion molecule-1 (ICAM-1) expression with that of the increase in the expression of inducible nitric oxide synthase (iNOS) protein and activity caused by endotoxin in cultured J774.2 macrophages. Treatment of J774.2 cells with lipopolysaccharide E. coli (LPS) induced a concentration-dependent increase in the expression of ICAM-1 on the cell surface within 4 h and an increase in iNOS protein and activity at 24 h. The upregulation of ICAM-1 expression on J774.2 macrophages caused by LPS was significantly inhibited by pretreatment of the cells with inhibitors of the activation of the nuclear transcription factor NF-kappaB, such as L-1-tosylamido-2 phenylethylchloromethyl ketone (TPCK), pyrrolidine dithiocarbamate (PDTC), rotenone or calpain inhibitor I, but not by the tyrosine kinase inhibitors, tyrphostin AG126 or genistein. In contrast, genistein or tyrphostin AG126 also prevented the induction of iNOS protein and activity in J774.2 macrophages elicited by LPS. Thus, the increase in the expression of ICAM-1 on J774.2 macrophages by endotoxin involves the activation of NFkappaB, but not of protein tyrosine kinase. PMID- 10704145 TI - The destructive action of IL-1alpha and IL-1beta in IDDM is a multistage process: evidence and confirmation by apoptotic studies, induction of intermediates and electron microscopy. AB - Using the rat beta-cell RIN-5AH insulinoma line as a means for studying insulin dependent diabetes mellitus (IDDM), it is shown that interleukin-1 (IL-1) induces beta-cell damage initiated by early apoptotic signals. This action is demonstrated by DNA fragmentation, as assessed by specific BrdU labeling, surface expression of Fas and nitric oxide (NO) production. In addition, the interplay between NO and Fas is shown, while scanning electron microscopy (SEM) confirms apoptosis by revealing the degree and type of cellular damage which, in the case of IL-1alpha, can be reversed by an inhibitor to NO synthesis. Apoptosis is also reconfirmed by transmission electron microscopy (TEM) by observing condensed nuclear chromatin after IL-1 exposure. Thus, treatment of insulinoma cells with IL-1alpha and IL-1beta seems to initiate a number of signals, including PKC activation as published previously, that ultimately lead to beta-cell destruction. Each IL-1 isoform, however, definitely follows a different pathway of action. PMID- 10704146 TI - Estimation of SLE activity based on the serum level of chosen cytokines and superoxide radical generation. AB - We estimated the serum levels of IL-6, TNF-alpha and IL-10, and generation of superoxide radicals, as well as their mutual dependence, in 63 SLE patients at various stages of disease activity. Our results indicate a statistically significant increase of the serum levels studied, and an increase of superoxide anion generation by granulocytes, in correlation with SLE activity. These results indicate that oxygen metabolism and the examined cytokines play an important role in pathogenesis of SLE. The assessment of these parameters can be useful in the estimation of disease activity. PMID- 10704147 TI - Expression of Bcl-2 in inflammatory sites from patients with active Behcet's disease. AB - Behcet's disease (BD) is a current systemic vasculitis of unknown aetiology. Eyes, skin, joints, the oral cavity, genital system, blood vessels, central nervous system and lung are usually involved. Defective regulation of programmed cell death (apoptosis) may play a role in the development of (BD), and the proto oncogene Bcl-2 is involved in the control of apoptosis in immunocompetent cells. We therefore wished to investigate the expression of Bcl-2 in the peripheral lymphocytes and in two inflammatory sites of patients with active BD: bronchoalveolar lavage (BAL) and cerebrospinal fluid (CSF) lymphocytes. Levels of Bcl-2 expression in the lymphocytes of patients with BD and, for comparison, in the lymphocytes of healthy controls and non-inflammatory neurological diseases (NIND), were studied by two-colour cytofluorography and RNA analysis. In BD patients, a significant proportion of T cells expressed increased amounts of Bcl 2 protein, both in peripheral blood and in inflammatory sites. Mononuclear cells of patients with BD showed increased amount of Bcl-2 messenger RNA. The in vitro incubation of T lymphocytes with IL-10, significantly increased the Bcl-2 expression, specifically in T lymphocytes from inflammatory sites. In active BD, stimulation of HSV-1 T lymphocytes slightly increased Bcl-2 expression, not significantly different from unstimulated HSV-1 T cells. The occurrence of circulating T lymphocytes with abnormally high Bcl-2 expression in peripheral circulation and in inflammatory sites may be explained in part by the increased in vivo activation levels, and by aetiopathological agent(s): our findings seem to indicate an important role in the chronic inflammation in BD. PMID- 10704148 TI - Leguminous lectins as tools for studying the role of sugar residues in leukocyte recruitment. AB - The natural physiological ligands for selectins are oligosaccharides found in glycoprotein or glycolipid molecules in cell membranes. In order to study the role of sugar residues in the in vivo lectin anti-inflammatory effect, we tested three leguminous lectins with different carbohydrate binding affinities in the peritonitis and paw oedema models induced by carrageenin in rats. L. sericeus lectin was more anti-inflammatory than D. virgata lectin, the effects being reversed by their specific binding sugars (N-acetylglucosamine and alpha methylmannoside, respectively). However, V. macrocarpa, a galactose-specific lectin, was not anti-inflammatory. The proposed anti-inflammatory activity of lectins could be due to a blockage of neutrophil-selectin carbohydrate ligands. Thus, according to the present data, we suggest an important role for N acetylglucosamine residue as the major ligand for selectins on rat neutrophil membranes. PMID- 10704149 TI - Incorporation and effect of arachidonic acid on the growth of human myeloma cell lines. AB - The objectives of this work are to investigate the incorporation of arachidonic acid (AA) in the human myeloma cell lines OPM2, U266 and IM9, and to assess the effect of AA and lipoxygenase products of AA on their growth. The kinetics of acylation of [3H]AA indicates that myeloma cells incorporate AA into their membrane phospholipids and triglycerides. PLA2-treatment and base hydrolysis experiments confirm that [3H]AA is incorporated unmodified in U266, IM9 and OPM2 phospholipids, and is linked by an ester bond. Prelabeling-chase experiments indicate no trafficking of labeled AA among the various phospholipid species. Addition of AA and lipoxygenase products of AA (leukotriene B4 and C4, lipoxin A4 and B4, 12- and 15-hydroxyeicosatetraenoic acid) have no effect on U266, IM9 and OPM2 proliferation assessed by [3H]thymidine incorporation into DNA. In conclusion, while human myeloma cells readily incorporate AA in their membrane phospholipids and triglycerides, AA and lipoxygenase products are not important modulators of their proliferation. PMID- 10704150 TI - Chronic mild prenatal stress exacerbates the allergen-induced airway inflammation in rats. AB - The effects of chronic mild prenatal stress on leukocyte infiltration into the airways was investigated in rat offspring. The chronic prenatal stress consisted of transitory and variable changes in the rat's living conditions. Offspring at adult age were actively sensitized (day 0) and intratracheally challenged (day 14) with ovalbumin. Bronchoalveolar lavage was performed in the offspring at 48 h after intratracheal challenge with ovalbumin. A significant increase in total leukocyte infiltration was observed in the non-stressed offspring group and this was associated with a marked recruitment of eosinophils without a significant effect on the influx of neutrophils and mononuclear cells. In the prenatal stressed offspring, the counts of both total leukocyte and eosinophils, as well as mononuclear cells, was increased by 50% compared to the non-stressed offspring. We provide here the first experimental evidence that chronic mild unpredictable prenatal stress produces a marked increase in the allergen-induced airway inflammation in the rat offspring. PMID- 10704151 TI - Cell cycle regulation and RNA polymerase II. AB - The cell cycle and transcription by RNA polymerase II (RNAP II) are closely related. They utilize shared components. RNAP II transcriptional activity is modulated during the cell cycle. Cell cycle dependent changes in the phosphorylation status of the carboxyl-terminal domain (CTD) of the largest subunit of RNAP II (RNAP II-LS) alter transcription. Several CTD kinases are members of the cyclin-dependent kinase (cdk) superfamily, including p34cdc2 (cdk1), cdk7, cdk8, and cdk9. Each of these cdks, with their respective cyclin partners, have been linked to cell cycle regulatory events. Other CTD kinases such as casein kinase II (CKII) and c-abl have also been implicated in cell cycle dependent modifications of the CTD. In addition, the stalling of RNAP II complexes at DNA lesions helps stimulate p53 accumulation which largely determines the cell's DNA damage response, including cell cycle arrest. Alzheimer's disease pathology results partially from activation of mitotic cdks in postmitotic neurons which can phosphorylate RNAP II-LS and other targets. PMID- 10704152 TI - Visual attention and aging. AB - The present review of visual attentional processes and aging focuses on definitions of attention that emphasize some aspect of the control of information processing (selective attention) or the processing resources needed to drive these control processes (attentional capacity). Emphasis is placed on how increased adult age affects attentional mechanisms and how these age differences in attention affect overall information processing. Past research has emphasized that selective attention appears to be resistant to age-related decline. Age related deficits in attentional capacity or processing resources, however, have been found. A review of more recent psychological research demonstrates the extension of the investigation of attention with emphasis on further defining what is selected in selective attention, and on reexamining the processing resources or capacity issue. Finally, developments in cognitive neuroscience are reviewed in terms of their relevance to attention and aging. PMID- 10704153 TI - Modulations of primary visual cortex activity representing attentive and conscious scene perception. AB - In the visual cortex, information is transferred from one area to the next by means of feedforward connections. These connections shape the receptive field properties of neurons in subsequent visual areas. Horizontal and feedback connections modulate this neuronal activity, resulting in the phenomenon of contextual modulation. In area V1, where receptive field properties reflect only low level processing, contextual modulation can be observed that represents fully evaluated perceptual saliency of the features within the receptive field. Here, we discuss to what extent these modulations are related to high level visual processes like perceptual organization, attention and visual awareness. Contextual modulation appears to reflects a process very distinct from receptive field based processing. This process seems to integrate information from distant areas in visual cortex to neurophysiologically 'highlight' those neurons that represent image elements or features of objects that stand out perceptually. Moreover, similar modulations are observed in relation to whether objects are attended to or not. Finally, these modulations are only present when subjects are aware of the visual input. PMID- 10704154 TI - Mitogen-activated signaling and cell cycle regulation in airway smooth muscle. AB - Increased airway smooth muscle mass has been demonstrated in patients with bronchopulmonary dysplasia and asthma. These data highlight the need for a precise understanding of the events involved in airway smooth muscle mitogenesis. To that end, investigators have developed cell culture systems adopting tracheal and bronchial myocytes from different species. A growing body of literature suggests that common signal transduction pathways regulate airway smooth muscle cell cycle entry across species lines. This review summarizes what is known about mitogen-activated signal transduction in airway smooth muscle cells. The extracellular signal regulated kinase (ERK) and phosphatidylinositol 3-kinase (PI 3-kinase) pathways appear to be major positive regulators of airway smooth muscle proliferation. It is also conceivable that growth factor stimulation of airway smooth muscle simultaneously elicits signaling through negative regulatory pathways such as the p38 mitogen-activated protein (MAP) kinase pathway, perhaps as a safeguard against excessive growth. PMID- 10704155 TI - Interactions of HIV-1 NEF with cellular signal transducing proteins. AB - Nef is a 27 - 34 kD myristoylated protein unique to primate lentiviruses. A functional Nef gene is important for development of high viremia and simian AIDS in SIV infected rhesus macaques (1). In a transgenic mouse model expression of Nef protein alone when expressed under a CD4-promoter is sufficient to cause an AIDS like disease (2). A critical role for Nef in development of AIDS in humans is suggested by the observation that some individuals with a long-term nonprogressive HIV-1 infection are infected with viruses carrying naturally occurring Nef deletions (3-5). The mechanism of Nef action remains incompletely understood, but multiple lines of evidence point out to a role in modulation of cellular signaling pathways via physical and functional interactions with host cell proteins. PMID- 10704156 TI - Circulation online only : march 7, 2000 PMID- 10704157 TI - From bench to bedside: the future is now. Presented at the 72nd Scientific Sessions of the American Heart Association, Atlanta, Georgia. PMID- 10704158 TI - Predicting restenosis: bigger is better but not best. PMID- 10704159 TI - Long-term follow-up of patients with mild coronary artery disease and endothelial dysfunction. AB - BACKGROUND: Coronary endothelial dysfunction is characterized by vasoconstrictive response to the endothelium-dependent vasodilator acetylcholine. Although endothelial dysfunction is considered an early phase of coronary atherosclerosis, there is a paucity of information regarding the outcome of these patients. Thus, this study was designed to evaluate the outcome of patients with mild coronary artery disease on the basis of their endothelial function. METHODS AND RESULTS: Follow-up was obtained in 157 patients with mildly diseased coronary arteries who had undergone coronary vascular reactivity evaluation by graded administration of intracoronary acetylcholine, adenosine, and nitroglycerin and intracoronary ultrasound at the time of diagnostic study. Patients were divided on the basis of their response to acetylcholine into 3 groups: group 1 (n=83), patients with normal endothelial function; group 2 (n=32), patients with mild endothelial dysfunction; and group 3 (n=42), patients with severe endothelial dysfunction. Over an average 28-month follow-up (range, 11 to 52 months), none of the patients from group 1 or 2 had cardiac events. However, 6 (14%) with severe endothelial dysfunction had 10 cardiac events (P<0.05 versus groups 1 and 2). Cardiac events included myocardial infarction, percutaneous or surgical coronary revascularization, and/or cardiac death. CONCLUSIONS: Severe endothelial dysfunction in the absence of obstructive coronary artery disease is associated with increased cardiac events. This study supports the concept that coronary endothelial dysfunction may play a role in the progression of coronary atherosclerosis. PMID- 10704160 TI - Randomized trial comparing intravenous nitroglycerin and heparin for treatment of unstable angina secondary to restenosis after coronary artery angioplasty. AB - BACKGROUND: The treatment of unstable angina targets the specific pathophysiological thrombotic process at the site of the active culprit lesion. In unstable angina due to a restenotic lesion, smooth muscle cell proliferation and increased vasoreactivity may play a more important role than thrombus formation. Therefore, the relative benefits of nitroglycerin and heparin might differ in unstable angina associated with restenosis compared with classic unstable angina. METHODS AND RESULTS: We randomized 200 patients hospitalized for unstable angina within 6 months after angioplasty (excluding those with intracoronary stents) to double-blind administration of intravenous nitroglycerin, heparin, their combination, or placebo for 63+/-30 hours. Recurrent angina occurred in 75% of patients in the placebo and heparin-alone groups, compared with 42.6% of patients in the nitroglycerin-alone group and 41.7% of patients in the nitroglycerin-plus-heparin group (P<0.003). Refractory angina requiring angiography occurred in 22.9%, 29.2%, 4. 3%, and 4.2% of patients, respectively (P<0.002). The odds ratios for being event free were 0.24 (95% CI, -0.13 to 0.45, P=0.0001) for nitroglycerin versus no nitroglycerin and 0.98 (95% CI, -0.55 to 1. 73, P=NS) for heparin versus no heparin. No patient died or suffered myocardial infarction. CONCLUSIONS: Intravenous nitroglycerin is highly effective in preventing adverse ischemic events (recurrent or refractory angina) in patients with unstable angina secondary to restenosis, whereas heparin has no effect. PMID- 10704161 TI - Prediction of restenosis after coronary angioplasty by use of a new index: TIMI frame count/minimal luminal diameter ratio. AB - BACKGROUND: It has been shown recently that postangioplasty coronary flow reserve and the degree of residual stenosis have a modest predictive value for short- and long-term clinical outcomes after coronary angioplasty. Corrected TIMI frame count (CTFC) is a simple quantitative index of coronary blood flow. Its relationship with Doppler coronary flow velocity and clinical outcome after coronary angioplasty has not been fully clarified. The aim of this study was to identify clinical, angiographic, and functional predictors of clinical and angiographic restenosis after conventional coronary angioplasty. METHODS AND RESULTS: We studied 70 consecutive patients in whom intracoronary Doppler flow velocity measurements were performed before and after angioplasty. Patients were evaluated for restenosis by clinical follow-up, exercise stress test/(201)Tl scintigraphy, and follow-up angiography, which was performed at 10. 5+/-10.3 months in 63 patients. According to the results of univariate analysis, a new index, postangioplasty CTFC/minimal luminal diameter (MLD) ratio, was created. Multivariate analysis revealed that CTFC/MLD ratio was the only independent predictor of angiographic (OR 2.02; 95% CI 1.37 to 2.97; P<0.0004) and clinical (OR 1.60; 95% CI 1.15 to 2.21; P<0.005) restenosis. The receiver operating characteristic curve area of this index was 79% for angiographic and 73% for clinical restenosis. The optimal CTFC/MLD ratio cutoff values were 7.88 for angiographic and 7.94 for clinical restenosis, respectively. CONCLUSIONS: Our data indicate that postangioplasty CTFC/MLD ratio, which incorporates both the angiographic and functional features of coronary lesions, is a reliable, objective, and inexpensive index for prediction of angiographic and clinical restenosis after conventional coronary angioplasty. PMID- 10704162 TI - Acute myocardial infarction complicated by atrial fibrillation in the elderly: prevalence and outcomes. AB - BACKGROUND: Although atrial fibrillation (AF) is a common complication of acute myocardial infarction (MI), patient characteristics and association with outcomes remain poorly defined in the elderly. METHODS AND RESULTS: We evaluated 106 780 Medicare beneficiaries > or =65 years of age from the Cooperative Cardiovascular Project treated for acute MI between January 1994 and February 1996 to determine the prevalence and prognostic significance of AF complicating acute MI in elderly patients. Patients were categorized on the basis of the presence of AF, and those with AF were further subdivided by time of AF (present on arrival versus developing during hospitalization). AF and non-AF patients were compared by univariate analysis, and logistic regression modeling was used to identify clinical predictors of AF. The influence of AF on outcomes was evaluated by unadjusted Kaplan-Meier survival curves and logistic regression models. AF was documented in 23 565 patients (22. 1%): 11 510 presented with AF and 12,055 developed AF during hospitalization. AF patients were older, had more advanced heart failure, and were more likely to have had a prior MI and undergone coronary revascularization. AF patients had poorer outcomes, including higher in-hospital (25.3% versus 16.0%), 30-day (29.3% versus 19.1%), and 1-year (48.3% versus 32.7%) mortality. AF remained an independent predictor of in-hospital (odds ratio [OR], 1. 21), 30-day (OR, 1.20), and 1-year (OR, 1.34) mortality after multivariate adjustment. Patients developing AF during hospitalization had a worse prognosis than patients who presented with AF. CONCLUSIONS: AF is a common complication of acute MI in elderly patients and independently influences mortality, particularly when it develops during hospitalization. PMID- 10704163 TI - Insulin-resistant prediabetic subjects have more atherogenic risk factors than insulin-sensitive prediabetic subjects: implications for preventing coronary heart disease during the prediabetic state. AB - BACKGROUND: Subjects who convert to type 2 diabetes mellitus have increased cardiovascular risk factors relative to nonconverters. However, it is not known whether these atherogenic changes in the prediabetic state are predominantly due to insulin resistance, decreased insulin secretion, or both. METHODS AND RESULTS: We examined this issue in the 7-year follow-up of the San Antonio Heart Study, in which 195 of 1734 subjects converted to type 2 diabetes. At baseline, converters had significantly higher body mass index, waist circumference, triglyceride concentration, and blood pressure and lower HDL cholesterol than nonconverters. Atherogenic changes in converters were markedly attenuated (and no longer significant) after adjustment for the homeostasis model assessment of insulin resistance (HOMA IR, a surrogate for insulin resistance); in contrast, the differences in risk factors between converters and nonconverters increased after adjustment for the ratio of early insulin increment to early glucose increment (DeltaI(30-0)/DeltaG(30-0)) during an oral glucose tolerance test (a surrogate for insulin secretion). We also compared converters who had a predominant insulin resistance (high HOMA IR and high DeltaI(30-0)/DeltaG(30-0)) (n=56) and converters who had a predominant decrease in insulin secretion (low HOMA IR and low DeltaI(30-0)/DeltaG(30-0)) (n=31) with nonconverters (n=1539). Only the converters who were insulin resistant had higher blood pressure and triglyceride levels and lower HDL cholesterol levels than nonconverters. CONCLUSIONS: Our data suggest that atherogenic changes in the prediabetic state are mainly seen in insulin-resistant subjects and that strategies to prevent type 2 diabetes might focus on insulin-sensitizing interventions rather than interventions that increase insulin secretion because of potential effects on cardiovascular risk. PMID- 10704164 TI - Fast determination of regional myocardial strain fields from tagged cardiac images using harmonic phase MRI. AB - BACKGROUND: Tagged MRI of the heart is difficult to implement clinically because of the lack of fast analytical techniques. We investigated the accuracy of harmonic phase (HARP) imaging for rapid quantification of myocardial strains and for detailed analysis of left ventricular (LV) function during dobutamine stimulation. METHODS AND RESULTS: Tagged MRI was performed in 10 volunteers at rest and during 5 to 20 microg(-1). kg(-1). min(-1) dobutamine and in 9 postinfarct patients at rest. We compared 2D myocardial strains (circumferential shortening, Ecc; maximal shortening, E(2); and E(2), direction) as assessed by a conventional technique and by HARP. Full quantitative analysis of the data was 10 times faster with HARP. For pooled data, the regression coefficient was r=0.93 for each strain (P<0.001). In volunteers, Ecc and E(2) were greater in the free wall than in the septum (P<0.01), but recruitable myocardial strain at peak dobutamine was greater in the LV septum (P<0.01). E(2) orientation shifted away from the circumferential direction at peak dobutamine (P<0.01). HARP accurately detected subtle changes in myocardial strain fields under increasing doses of dobutamine. In patients, HARP-determined Ecc and E(2) values were dramatically reduced in the asynergic segments as compared with remote (P<0.001), and E(2) direction shifted away from the circumferential direction (P<0.001). CONCLUSIONS: HARP MRI provides fast, accurate assessment of myocardial strains from tagged MR images in normal subjects and in patients with coronary artery disease with wall motion abnormalities. HARP correctly indexes dobutamine-induced changes in strains and has the potential for on-line quantitative monitoring of LV function during stress testing. PMID- 10704165 TI - Inspiratory impedance during active compression-decompression cardiopulmonary resuscitation: a randomized evaluation in patients in cardiac arrest. AB - BACKGROUND: Blood pressure is severely reduced in patients in cardiac arrest receiving standard cardiopulmonary resuscitation (CPR). Although active compression-decompression (ACD) CPR improves acute hemodynamic parameters, arterial pressures remain suboptimal with this technique. We performed ACD CPR in patients with a new inspiratory threshold valve (ITV) to determine whether lowering intrathoracic pressures during the "relaxation" phase of ACD CPR would enhance venous blood return and overall CPR efficiency. METHODS AND RESULTS: This prospective, randomized, blinded trial was performed in prehospital mobile intensive care units in Paris, France. Patients in nontraumatic cardiac arrest received ACD CPR plus the ITV or ACD CPR alone for 30 minutes during advanced cardiac life support. End tidal CO(2) (ETCO(2)), diastolic blood pressure (DAP) and coronary perfusion pressure, and time to return of spontaneous circulation (ROSC) were measured. Groups were similar with respect to age, gender, and initial rhythm. Mean maximal ETCO(2), coronary perfusion pressure, and DAP values, respectively (in mm Hg), were 13.1+/-0.9, 25.0+/-1.4, and 36.5+/-1.5 with ACD CPR alone versus 19.1+/-1.0, 43.3+/-1.6, and 56.4+/-1.7 with ACD plus valve (P<0.001 between groups). ROSC was observed in 2 of 10 patients with ACD CPR alone after 26.5+/-0.7 minutes versus 4 of 11 patients with ACD CPR plus ITV after 19.8+/-2.8 minutes (P<0.05 for time from intubation to ROSC). Conclusions Use of an inspiratory resistance valve in patients in cardiac arrest receiving ACD CPR increases the efficiency of CPR, leading to diastolic arterial pressures of >50 mm Hg. The long-term benefits of this new CPR technology are under investigation. PMID- 10704166 TI - Identification of the substrate of atrial vulnerability in patients with idiopathic atrial fibrillation. AB - BACKGROUND: Experimental studies have shown that atrial fibrillation (AF) causes remodeling, which facilitates AF perpetuation. AF may also, however, occur in patients without remodeling and underlying structural cardiac disease. The substrate for enhanced vulnerability in these patients is unknown. METHODS AND RESULTS: We studied 43 patients without structural heart disease: 18 patients with documented sporadic paroxysmal AF and 25 control patients without AF. In each patient, a decapolar catheter was positioned against the right atrial free wall, and a quadripolar catheter was positioned in the right atrial appendage. Unipolar electrograms were recorded. Atrial vulnerability was assessed according to an increasingly aggressive stimulation protocol. Mean local fibrillatory interval (FI) was used as an index of local refractoriness. Spatial dispersion of refractoriness was assessed through the calculation of the coefficient of dispersion (CD), which was defined as the SD of mean local FI expressed as a percentage of the mean FI. In the AF group, AF was induced with a single extrastimulus in 16 of 18 patients; the CD was 5.4+/-2.6, and the mean FI was 164+/-29 ms. In the control group, AF could be induced only with more aggressive pacing in 23 of the 25 patients; the CD was 1.4+/-0.7 (P<0.0001), and the mean FI was 175+/-26 ms (NS). CONCLUSIONS: Patients with idiopathic AF showed increased dispersion of refractoriness, which may be the substrate for the observed enhanced inducibility and spontaneous occurrence of AF. PMID- 10704167 TI - Hemodialysis impairs endothelial function via oxidative stress: effects of vitamin E-coated dialyzer. AB - BACKGROUND: Patients who undergo hemodialysis experience accelerated atherosclerosis and premature death. Recent evidence suggests that endothelial dysfunction proceeds to and exacerbates atherosclerosis. It remains unknown whether hemodialysis per se causes endothelial dysfunction. METHODS AND RESULTS: We evaluated endothelial function estimated by flow-mediated vasodilation during reactive hyperemia using high-resolution ultrasound Doppler echocardiography before and after a single session in patients on maintenance hemodialysis. Several studies have shown that the imbalance between pro-oxidant and antioxidant activities in hemodialyzed patients results in high oxidative stress, which causes lipid peroxidation and endothelial injury. Accordingly, we investigated the effects of antioxidative modification during hemodialysis on endothelial function using a vitamin E-coated cellulose membrane dialyzer. Nonspecific endothelium-independent vasodilation was measured after administration of a sublingual glyceryl trinitrate spray (0.3 mg). A single session of hemodialysis by noncoated dialyzer impaired flow-mediated vasodilation (P<0.05) associated with increased plasma levels of oxidized LDL (P<0.05), an index of oxidative stress. Hemodialysis by vitamin E-coated membrane prevented dialysis-induced endothelial dysfunction and increases in oxidized LDL. Plasma levels of oxidized LDL were inversely correlated with the magnitudes of flow-mediated vasodilation (r=-0.53, P< 0.001). Hemodialysis by noncoated or vitamin E-coated membrane did not affect glyceryl trinitrate-induced endothelium-independent vasodilation. CONCLUSIONS: Our findings indicate that hemodialysis per se impairs endothelial function, possibly by increasing oxidative stress. PMID- 10704168 TI - Asymptomatic peripheral arterial disease is independently associated with impaired lower extremity functioning: the women's health and aging study. AB - BACKGROUND: We report the implications of asymptomatic lower extremity peripheral arterial disease (PAD) for lower extremity functioning among participants in the Women's Health and Aging Study, an observational study of disabled women > or = 65 years of age living in and around Baltimore. METHODS AND RESULTS: The ankle brachial index (ABI) and measures of upper and lower extremity functioning were measured among study participants. Of 933 women with ABI < or =1. 50, 328 (31%) [corrected] had an ABI <0.90, consistent with PAD. Sixty-three percent of PAD participants had no exertional leg pain. Among participants without exertional leg pain, lower ABI levels were associated with slower walking velocity, poorer standing balance score, slower time to arise 5 times consecutively from a seated position, and fewer blocks walked per week, adjusting for age, sex, race, cigarette smoking, and comorbidities. ABI was not associated independently with measures of upper extremity functioning. CONCLUSIONS: Asymptomatic PAD is common and is independently associated with impaired lower extremity functioning. In addition to preventing cardiovascular morbidity and death, further study is warranted to identify effective interventions to improve functioning among the growing number of men and women with asymptomatic PAD. PMID- 10704169 TI - Platelet GP IIIa Pl(A) polymorphisms display different sensitivities to agonists. AB - BACKGROUND: Both inherited predisposition and platelet hyperreactivity have been associated with ischemic coronary events, but mechanisms that support genetic differences among platelets from different subjects are generally lacking. Associations between the platelet Pl(A2) polymorphism of GP IIIa and coronary syndromes raise the question as to whether this inherited variation may contribute to platelet hyperreactivity. METHODS AND RESULTS: In this study, we characterized functional parameters in platelets from healthy donors with the Pl(A) (HPA-1) polymorphism, a Leu (Pl(A1)) to Pro (Pl(A2)) substitution at position 33 of the GP IIIa subunit of the platelet GP IIb/IIIa receptor (integrin alpha(IIb)beta(3)). We studied 56 normal donors (20 Pl(A1,A1), 20 Pl(A1,A2), and 16 Pl(A2,A2)). Compared with Pl(A1,A1) platelets, Pl(A2)-positive platelets showed a gene dosage effect for significantly greater surface-expressed P selectin, GP IIb/IIIa-bound fibrinogen, and activated GP IIb/IIIa in response to low-dose ADP. Surface expression of GP IIb/IIIa was similar in resting platelets of all 3 genotypes but was significantly greater on Pl(A2,A2) platelets after ADP stimulation (P=0.003 versus Pl(A1,A1); P=0.03 versus Pl(A1,A2)). Pl(A1,A2) platelets were more sensitive to inhibition of aggregation by pharmacologically relevant concentrations of aspirin and abciximab. CONCLUSIONS: Pl(A2)-positive platelets displayed a lower threshold for activation, and platelets heterozygous for Pl(A) alleles showed increased sensitivity to 2 antiplatelet drugs. These in vitro platelet studies may have relevance for in vivo thrombotic conditions. PMID- 10704170 TI - Crucial role of endogenous interleukin-10 production in myocardial ischemia/reperfusion injury. AB - BACKGROUND: The anti-inflammatory cytokine interleukin-10 (IL-10) has been detected in the plasma of patients with myocardial ischemia/reperfusion. The aim of our study was to investigate the role of endogenously produced IL-10 in myocardial ischemia/reperfusion. METHODS AND RESULTS: In the present study, we used wild-type and IL-10-deficient mice subjected to myocardial ischemia/reperfusion. Significant levels of IL-10 were produced in wild-type mice at 2 to 6 hours after myocardial reperfusion. The genetic deletion of IL-10 enhanced neutrophil infiltration into the reperfused tissues at 6 hours after reperfusion and increased infarct size and myocardial necrosis. Furthermore, in the absence of IL-10, an enhancement of the inflammatory response was seen, as demonstrated by increased plasma levels of tumor necrosis factor-alpha, nitrite/nitrate (breakdown products of NO), and increased tissue expression of intercellular adhesion molecule-1. Reperfusion for 24 hours was associated with a 75% mortality rate in IL-10-deficient mice, whereas no deaths occurred in the wild-type animals. CONCLUSIONS: The present findings provide the first direct evidence that endogenous IL-10 inhibits the production of tumor necrosis factor alpha and NO and serves to protect the ischemic and reperfused myocardium through the suppression of neutrophil recruitment. PMID- 10704171 TI - Gene transfer of endothelial nitric oxide synthase improves relaxation of carotid arteries from diabetic rabbits. AB - BACKGROUND: Diabetes mellitus is associated with impairment of NO-mediated vascular relaxation. The purpose of this study was to determine whether adenovirus-mediated gene transfer of endothelial NO synthase (eNOS) or Cu/Zn superoxide dismutase (SOD1) improves responsiveness to acetylcholine in alloxan induced diabetic rabbits. METHODS AND RESULTS: After 8 weeks, plasma glucose was greater in diabetic rabbits (418+/-35 mg/dL) (mean+/-SEM) than in normal rabbits (105+/-4 mg/dL). Carotid arteries were removed and cut into ring segments. Arteries were incubated for 2 hours with adenoviral vectors driven by a CMV promoter expressing beta-galactosidase (beta-gal), eNOS, SOD1, or vehicle. After incubation with virus, arteries were incubated for an additional 24 hours to allow transgene expression. Vascular reactivity was examined by recording isometric tension. After precontraction with phenylephrine, responses to the endothelium-independent vasodilator sodium nitroprusside were similar in diabetic and normal arteries. Endothelium-dependent relaxation to acetylcholine (3x10(-6) mol/L) was significantly less in arteries from diabetic animals (68+/-5%) than in normal vessels (90+/-3%). Adenoviral transfection of arteries with eNOS improved relaxation in response to acetylcholine in diabetic (EC(50) eNOS=0.64+/-0.12x10( 7) mol/L versus vehicle =1. 70+/-0.43x10(-7) mol/L) but not normal arteries. Vasorelaxation in response to acetylcholine was inhibited by N(omega)-nitro-L arginine (100 micromol/L) in all groups. Responses to acetylcholine were unchanged after gene transfection of SOD1 or beta-gal in arteries from diabetic or normal rabbits. CONCLUSIONS: Adenovirus-mediated gene transfer of eNOS, but not SOD, improves impaired NO-mediated relaxation in vessels from diabetic rabbits. PMID- 10704172 TI - Attenuated cardiac allograft vasculopathy in mice with targeted deletion of the transcription factor STAT4. AB - BACKGROUND: To study transcription factor signaling pathways that mediate cardiac allograft vasculopathy, we used mice with targeted gene deletion of signal transducer and activator of transcription (STAT)4 and STAT6 as recipients in our mouse cardiac transplant model of chronic rejection. METHODS AND RESULTS: At day 55 after transplantation, cardiac grafts placed into STAT4 -/- (n=10) had reduced frequency (24+/-2%) and severity (9+/-4%) of vascular occlusion compared with wild-type controls (n=7, frequency 70+/-12% [P<0.001], severity 25+/-6% [P<0.05]). This decrease was associated with reduced intragraft expression ((32)P RT-PCR and immunohistochemistry) of the Th1 signature cytokines interferon-gamma (P<0.001) and interleukin (IL)-2 (P<0.001). Furthermore, cardiac grafts in STAT4 /- had fewer infiltrating CD45(+) mononuclear cells (99+/-27 cells/mm(3) compared with 551+/-168 cells/mm(3) in wild-type controls [P<0.05]) and reduced expression of P-selectin (P<0.001) and E-selectin (P<0.01) ligand, recently shown to regulate Th1 cell recruitment. In contrast, in grafts placed into STAT6 -/- (n=11), the development of cardiac allograft vasculopathy (frequency 62+/-8%, severity 28+/-6%) and Th2 cytokine profiles (IL-4, IL-10) were comparable to those in wild-type controls. CONCLUSIONS: Hence, we show that immune responses mediated by STAT4, but not STAT6, contribute to the development of cardiac allograft vasculopathy. We speculate that when present, STAT4-mediated signaling pathways may promote cardiac allograft vasculopathy by directing Th1-specific lymphocyte recruitment, activation, and effector functions. PMID- 10704173 TI - Improved mechanoenergetics and cardiac rest and reserve function of in vivo failing heart by calcium sensitizer EMD-57033. AB - BACKGROUND: Myofilament Ca(2+) sensitizers enhance contractility but can adversely alter diastolic function through sensitization to low intracellular Ca(2+) concentration. Concomitant phosphodiesterase III inhibition (PDE3I) may offset diastolic changes but limit the mechanoenergetic benefits. We tested whether selective Ca(2+) sensitization in vivo with the use of EMD-57033 enhances both systolic and diastolic function in failing hearts at minimal energetic cost. METHODS AND RESULTS: Pressure-dimension data were measured with sonomicrometry/micromanometry in conscious dogs before (CON, n=9) and after tachycardia-induced heart failure (HF, n=11). In contrast to blunted dobutamine (DOB) responses in HF, low-dose EMD-57033 (0.4 mg. kg(-1). min(-1) for 20 minutes) markedly enhanced contractility, doubling end-systolic elastance and raising fractional shortening similarly in CON-treated and HF hearts. EMD-57033 effects were achieved at a reduced heart rate, without vasodilation. EMD-57033 augmented blunted heart rate-dependent contractility responses in HF at a rate of twice that of DOB, despite matched basal inotropic responses. EMD-57033 also improved diastolic function, lowering left ventricular end-diastolic pressure and increasing the filling rate. At equipotent inotropic doses and matched preload, EMD-57033 lowered the oxygen cost of contractility by -11.4+/-5.8%, whereas it rose 64+/-18% with DOB (P=0.001) and 28+/-11% with milrinone. Doubling EMD-57033 dose further augmented positive inotropy in CON and HF, accompanied by vasodilation, increased heart rate, and other changes consistent with PDE3I coactivity, but the oxygen cost of contractility remained improved compared with the use of DOB. CONCLUSIONS: Selective Ca(2+) sensitization with minimal PDE3I in vivo is achieved with the use of EMD-57033, improving basal and rate-stimulated contractility and mechanoenergetics of HF without compromising diastolic function. Despite PDE3I activity at higher doses, energetic benefits persist. PMID- 10704174 TI - Proximal atrioventricular bundle, atrioventricular node, and distal atrioventricular bundle are distinct anatomic structures with unique histological characteristics and innervation. AB - BACKGROUND: Direct 3D analysis (ie, stereotaxic analysis of 3 planes) has shown that the atrioventricular (AV) node (AVN) is continuous with only specialized myocardium of the proximal AV bundle (PAVB) and distal AV bundle (DAVB) or His bundle. The purpose of the present study was to determine whether the PAVB, AVN, and DAVB possess histological features distinct from each other and from the ordinary myocardium. METHODS AND RESULTS: A protocol that preserves the cytoplasmic and interstitial integrity of the tissue and permits serial sections of the AV junction region to be made in 3 orthogonal planes showed that the PAVB, AVN, and DAVB are characterized by myocardium aggregated into fascicles containing approximately 8 myofibers. Myofibers within the fascicles are coiled or spiraled about each other; and spiraling is most compact in the PAVB. Collagen encases individual fascicles and segregates primary fascicles into secondary fascicles. Fascicles, and not myofibers, are in parallel array in the PAVB, interwoven in the AVN, and parallel in the DAVB. Narrow junctions of parallel fascicles separate the AVN from the PAVB and DAVB. Myocytes, which are largest in DAVB, possess clear perinuclear regions; thin finger-like end processes, which are most numerous in the AVN; uniform, delicate cross-striations; and intercalated disks, which are broader in the PAVB and form short stacks in the AVN. Sheaves of nerve terminals are found, including boutons as in skeletal muscle [corrected]. CONCLUSIONS: The PAVB, AVN, and DAVB have distinct histological features. Collagen septation of primary and secondary fascicles presents natural barriers within the tissues and to surrounding myocardium and structures. These findings confirm that the AV junction region contains a specialized conduction system that is anatomically isolated from ordinary myocardium. PMID- 10704175 TI - Heterogeneous sympathetic innervation influences local myocardial repolarization in normally perfused rabbit hearts. AB - BACKGROUND: Heterogeneity of sympathetic innervation is thought to contribute to the potential for fatal arrhythmia. However, little is known about the effects of heterogeneous innervation on repolarization. METHODS AND RESULTS: To assess this relationship, we measured activation recovery intervals (ARIs) from 64 epicardial sites in 11 rabbits studied 2 weeks after regional denervation produced by phenol and 4 sham-operated rabbits. ARI results were compared with the distribution of sympathetic innervation measured from 3D reconstructions of serial autoradiographs of [(125)I]metaiodobenzylguanidine and (99m)Tc-sestamibi. ARIs were recorded during baseline sinus rhythm, norepinephrine (NE) infusion (0.1 microg. kg(-1). min(-1)), and left stellate ganglion stimulation (SS). NE shortened ARI in 98% of electrodes in the denervated region. The degree of ARI shortening and dispersion increased (P<0.001 and P<0.01, respectively) as denervation became more severe. SS shortened ARI in 30% of electrodes in the denervated area, with increased shortening and dispersion related to increased severity of denervation (P<0.01). SS prolonged ARI in 70% of electrodes in the denervated area, with no correlation with severity of denervation. CONCLUSIONS: The magnitude and dispersion of local repolarization responses are related to the severity of denervation, as well as the type of stimulation: neural (SS) versus humoral (NE). The differences may relate to the concentration of NE released. PMID- 10704176 TI - Economic outcomes of implantable cardioverter-defibrillators. AB - BACKGROUND: We reviewed the literature pertaining to the cost-effectiveness of implantable cardioverter-defibrillator (ICD) therapy in the management of ventricular fibrillation and tachycardia. Discussed are the methodology, advantages, and limitations of economic-outcomes analyses as related to ICD therapy; the impact of new technology and physician practice patterns; and methodological recommendations for future studies. METHODS AND RESULTS: Articles published between 1990 and 1997 were screened for cost-effectiveness analyses of ICD versus antiarrhythmic drug therapy. Randomized clinical trials, prospective and retrospective studies, and economic models were included. These studies report incremental cost-effectiveness ratios ranging from cost savings of $13 975 per life-year saved (LYS) to an incremental cost of $114 917 per LYS for ICD therapy. Differences were due to study type, cost-reporting methodology, ICD technology used, and length of follow-up. Assuming current technology and physician practice patterns, we find that ICD total therapy costs may break even in 1 to 3 years. CONCLUSIONS: Recent literature suggests that ICDs are a cost effective therapy for management of life-threatening ventricular tachyarrhythmias. The advent of new technology and patient management practices should further improve the cost-effectiveness of ICD therapy. Future studies of ICD cost-effectiveness should address the implications of truncated follow-up periods and quality of life. PMID- 10704177 TI - Images in cardiovascular medicine. Saphenous vein graft aneurysm. PMID- 10704178 TI - Dynamic magnetic resonance images of murine cardiac function at physiologic heart rates. PMID- 10704180 TI - Good Blood Sugar Control Keeps Patients Healthier for Years. PMID- 10704179 TI - Subclinical Hypothyroidism : A Heart Disease Risk? PMID- 10704181 TI - Treating high-risk asthma populations. PMID- 10704182 TI - President proposes steps to prevent medical mistakes. PMID- 10704183 TI - Patient bill of rights on the agenda again. PMID- 10704184 TI - Meeting highlights: the 11th Annual Symposium on Transcatheter Cardiovascular Therapeutics. PMID- 10704185 TI - Hereditary deafness. PMID- 10704186 TI - Autosomal recessive nonsyndromic hearing loss. AB - Nearly all genes for autosomal recessive nonsyndromal inherited hearing loss (ARNSHL) localized thus far cause prelingual severe to profound or profound hearing impairment. Of the 25 reported loci, most have been identified using single consanguineous families. Six of these genes have been cloned and encode a variety of proteins, including ion channels, extracellular matrix components, cytoskeletal components, and proteins essential for synaptic vesicular trafficking. One of these genes appears to be responsible for approximately 50% of all congenital severe to profound or profound hearing loss in many world populations, and mutations in two other genes can lead to either syndromic or nonsyndromic forms of deafness. The identification of additional genes that cause ARNSHL and elucidation of their function will refine our understanding of auditory physiology at the molecular level. PMID- 10704187 TI - Clinical phenotype and mutations in connexin 26 (DFNB1/GJB2), the most common cause of childhood hearing loss. AB - Mutations in the gene for connexin 26, GJB2, are the most common cause of hearing loss in American and European populations, with a carrier rate of about 3%-a rate similar to that for cystic fibrosis. A single mutation, 35delG, is responsible for most of this autosomal recessive hearing loss, DFNB1. A broad spectrum of mutations in GJB2 has been found to be associated with hearing loss, including another deletion mutation, 167delT, which has a carrier rate of about 4% in the Ashkenazi Jewish population. Mutations in GJB2 have also been found to be associated with dominant nonsyndromic hearing loss, DFNA3. Clinical studies have shown that the recessive hearing loss can vary from mild to profound, even within the same sibship. This type of hearing loss is nonsyndromic and is accompanied by normal vision, vestibular responses, and no malformations of the inner ear detectable by computed tomography scanning. Progressive and asymmetrical hearing loss has been noted in some cases, but it accounts for fewer than one-third of the cases of this type of hearing loss. The discovery of mutations in GJB2 that cause hearing loss has profound implications in the early diagnosis of hearing loss in general. The relative ease of diagnosis by genetic testing of Cx26 permits early identification of children with GJB2/DFNB1 hearing loss. This testing, coupled with hearing loss diagnosed by infant auditory brainstem response audiometry, will ensure that hearing-impaired children and their parents receive proper medical, audiologic, genetic, and educational counseling. Am. J. Med. Genet. (Semin. Med. Genet.) 89:130-136, 1999. PMID- 10704188 TI - Jervell and Lange-Nielsen syndrome: a Norwegian perspective. AB - Jervell and Lange-Nielsen syndrome (MIM 220400; JLNS), is a rare form of profound congenital deafness combined with syncopal attacks and sudden death due to prolonged QTc; it is an autosomal recessive trait. After its first description in Norway in 1957, later reports from many other countries have confirmed its occurrence. Nowhere is the prevalence so high as in Norway, where we estimate a prevalence of at least 1:200,000. The KCNQ1 and KCNE1 proteins coassemble in a potassium channel, and mutations in either the KCNQ1 gene or the KCNE1 gene disrupt endolymph production in the stria vascularis in the cochlea, causing deafness. KCNQ1 seems to be the major gene in JLNS. Long QT syndrome (LQTS) is a separate disorder of either autosomal dominant or recessive inheritance caused by mutations in four different ion channel genes; KCNQ1 is the one most frequently involved. Some heterozygous carriers of JLNS mutations in either gene may suffer from prolonged QTc and be symptomatic LQTS patients with a need for appropriate medical treatment to prevent life-threatening cardiac arrhythmia. In general, frameshift/stop mutations cause JLNS, and missense/splice site mutations cause LQTS, but a precise genotype-phenotype correlation in LQTS and JLNS is not established, which complicates both genetic counseling and clinical risk evaluation in carriers. We review JLNS from a Norwegian perspective because of the unusually high prevalence, the genetic homogeneity associated with considerable mutational heterogeneity, and some evidence for recurrent mutational events as well as one founder mutation. We outline the clinical implications for investigation of deaf children and cases of sudden infant death syndrome as well as careful electrocardiographic monitoring of identified mutation carriers to prevent sudden death. Am. J. Med. Genet. (Semin. Med. Genet.) 89:137-146, 1999. PMID- 10704189 TI - Unconventional myosins and the genetics of hearing loss. AB - Mutations of the unconventional myosins genes encoding myosin VI, myosin VIIA and myosin XV cause hearing loss and thus these motor proteins perform fundamental functions in the auditory system. A null mutation in myosin VI in the congenitally deaf Snell's waltzer mice (Myo6(sv)) results in fusion of stereocilia and subsequent progressive loss of hair cells, beginning soon after birth, thus reinforcing the vital role of cytoskeletal proteins in inner ear hair cells. To date, there are no human families segregating hereditary hearing loss that show linkage to MYO6 on chromosome 6q13. The discovery that the mouse shaker1 (Myo7(ash1)) locus encodes myosin VIIA led immediately to the identification of mutations in this gene in Usher syndrome type 1B; subsequently, mutations in this gene were also found associated with recessive and dominant nonsyndromic hearing loss (DFNB2 and DFNA11). Stereocilla of sh1 mice are severely disorganized, and eventually degenerate as well. Myosin VIIA has been implicated in membrane trafficking and/or endocytosis in the inner ear. Mutant alleles of a third unconventional myosin, myosin XV, are associated with nonsyndromic, recessive, congenital deafness DFNB3 on human chromosome 17p11.2 and deafness in shaker2 (Myo15(sh2)) mice. In outer and inner hair cells, myosin XV protein is detectable in the cell body and stereocilia. Hair cells are present in homozygous sh2 mutant mice, but the stereocilia are approximately 1/10 of the normal length. This review focuses on what we know about the molecular genetics and biochemistry of myosins VI, VIIA and XV as relates to hereditary hearing loss. Am. J. Med. Genet. (Semin. Med. Genet.) 89:147-157, 1999. Published 2000 Wiley-Liss, Inc. PMID- 10704190 TI - The usher syndromes. AB - Mutations in the gene (MYO7A) encoding myosin-VIIa, a member of the large superfamily of myosin motor proteins that move on cytoplasmic actin filaments, and in the USH2A gene, which encodes a novel protein resembling an extracellular matrix protein or a cell adhesion molecule, both cause Usher syndrome (USH), a clinically heterogeneous autosomal recessive disorder comprising hearing and visual impairment. Patients with USH1 have severe to profound congenital hearing impairment, vestibular dysfunction, and retinal degeneration beginning in childhood, while those with USH2 have moderate to severe hearing impairment, normal vestibular function, and later onset of retinal degeneration. USH3 is characterized by progressive hearing loss and variable age of onset of retinal degeneration. The phenotype resulting from MYO7A and USH2A mutations is variable. While most MYO7A mutations cause USH1, some cause nonsyndromic hearing impairment, and one USH3 phenotype has been described. USH2A mutations cause atypical USH as well as USH2. MYO7A is on chromosome region 11q13 and USH2A is on 1q41. Seven other USH genes have been mapped but have not yet been identified. USH1A, USH1C, USH1D, USH1E, and USH1F have been assigned to chromosome bands 14q32, 11p15.1, 10q, 21q21, and 10, respectively, while USH2B is on 5q, and USH3 is at 3q21-q25. Myosin VIIa mutations also result in the shaker-1 (sh1) mouse, providing a model for functional studies. One possibility is that myosin-VIIa is required for linking stereocilia in the sensory hair bundle; another is that it may be needed for membrane trafficking. The ongoing studies of myosin-VIIa, the USH2A protein, and the yet to be identified proteins encoded by the other USH genes will advance understanding of the Usher syndromes and contribute to the development of effective therapies. Am. J. Med. Genet. (Semin. Med. Genet.) 89:158-166, 1999. PMID- 10704191 TI - Autosomal dominant nonsyndromic hearing impairment. AB - Nearly all genes that have been localized for autosomal dominantly inherited hearing impairment are characterized by postlingual hearing loss that is progressive in nature. This auditory phenotype is in contrast to that of genes localized for autosomal recessive hearing impairment, which generally cause nonprogressive severe-to-profound or profound prelingual hearing loss. In most cases, extended pedigrees have been used to localize autosomal dominant deafness genes, To date, 22 autosomal dominant loci have been mapped, and 10 of these genes have been cloned. The functions of these deafness-causing genes are diverse and include transcription factors, extracellular matrix components, ion channels, cytoskeletal components, and unknown functions. Interesting findings include the unexpected expression pattern of some of these genes and the discovery that in some genes, allele variants can cause either isolated hearing loss or syndromic deafness. The greatest challenge for future research will be identifying additional deafness-causing genes and elucidating their function in the inner ear. Am. J. Med. Genet. (Semin. Med. Genet.) 89:167-174, 1999. @ 2000 Wiley-Liss, Inc. PMID- 10704192 TI - Effects of subunit occupancy on partitioning of an intermediate in thymidylate synthase mutants. AB - Experimental evidence for a 5-exocyclic methylene-dUMP intermediate in the thymidylate synthase reaction was recently obtained by demonstrating that tryptophan 82 mutants of the Lactobacillus casei enzyme produced 5-(2 hydroxyethyl)thiomethyl-dUMP (HETM-dUMP) (Barret, J. E., Maltby, D. A., Santi, D. V., and Schultz, P. G. (1998) J. Am. Chem. Soc. 120, 449-450). The unusual product was proposed to emanate from trapping of the intermediate with beta mercaptoethanol in competition with hydride transfer from H(4)folate to form dTMP. Using mutants of the C-terminal residue of thymidylate synthase, we found that the ratio of HETM-dUMP to dTMP varies as a function of CH(2)H(4)folate concentration. This observation seemed inconsistent with the conclusion that both products arose from a common intermediate in which CH(2)H(4)folate was already bound to the enzyme. The enigma was resolved by a kinetic model that allowed for differential partitioning of the intermediate formed on each of the two subunits of the homodimeric enzyme in forming the two different products. With three C terminal mutants of L. casei TS, HETM-dUMP formation was consistent with a model in which product formation occurs upon occupancy of the first completely bound subunit, the rate of which is unaffected by occupancy of the second subunit. With one analogous E. coli TS mutant, HETM-dUMP formation occurred upon occupancy of the first subunit, but was inhibited when both subunits were occupied. With all mutants, dTMP formation occurs from occupied forms of both subunits at different rates; here, binding of cofactor to the first subunit decreased affinity for the second, but the reaction occurred faster in the enzyme form with both subunits bound to dUMP and CH(2)H(4)folate. The model resolves the apparent enigma of the cofactor-dependent product distribution and supports the conclusion that the exocyclic methylene intermediate is common to both HETM-dUMP and dTMP formation. PMID- 10704193 TI - L-Vinylglycine is an alternative substrate as well as a mechanism-based inhibitor of 1-aminocyclopropane-1-carboxylate synthase. AB - L-Vinylglycine (L-VG) has been shown to be a mechanism-based inhibitor of 1 aminocyclopropane-1-carboxylate (ACC) synthase [Satoh, S., and Yang, S. F. (1989) Plant Physiol. 91, 1036-1039] as well as of other pyridoxal phosphate-dependent enzymes. This report demonstrates that L-VG is primarily an alternative substrate for the enzyme. The L-VG deaminase activity of ACC synthase yields the products alpha-ketobutyrate and ammonia with a k(cat) value of 1.8 s(-1) and a K(m) value of 1.4 mM. The k(cat)/K(m) of 1300 M(-1) s(-1) is 0.17% that of the diffusion controlled reaction with the preferred substrate, S-adenosyl-L-methionine. The enzyme-L-VG complex partitions to products 500 times for every inactivation event. The catalytic mechanism proceeds through a spectrophotometrically detected quinonoid with lambda(max) of 530 nm, which must rearrange to a 2-aminocrotonate aldimine to yield final products. Alternative mechanisms for the inactivation reaction are presented, and the observed kinetics for the full reaction course are satisfactorily modeled by kinetic simulation. The inactive enzyme is an aldimine with lambda(max) of 432 nm. It is resistant to NaBH(3)CN but is reduced by NaBH(4). ACC synthase is now expressed in Pichia pastoris with an improved yield of 10 mg/L. PMID- 10704194 TI - Structure and binding specificity of the second N-terminal cellulose-binding domain from Cellulomonas fimi endoglucanase C. AB - The 1,4-beta-glucanase CenC from Cellulomonas fimi contains two cellulose-binding domains, CBD(N1) and CBD(N2), arranged in tandem at its N-terminus. These homologous CBDs are distinct in their selectivity for binding amorphous and not crystalline cellulose. Multidimensional heteronuclear nuclear magnetic resonance (NMR) spectroscopy was used to determine the tertiary structure of CBD(N2) in the presence of saturating amounts of cellopentaose. A total of 1996 experimental restraints were used to calculate an ensemble of 21 final structures for the protein. CBD(Nu2) is composed of 11 beta-strands, folded into two antiparallel beta-sheets, with a topology of a jellyroll beta-sandwich. On the basis of patterns of chemical shift perturbations accompanying the addition of cellooligosaccharides, as well as the observation of intermolecular protein-sugar NOE interactions, the cellulose-binding site of CBD(N2) was identified as a cleft that lies across one face of the beta-sandwich. The thermodynamic basis for the binding of cellooligosaccharides was investigated using isothermal titration calorimetry and NMR spectroscopy. Binding is enthalpically driven and consistent with a structural model involving hydrogen bonding between the equatorial hydroxyls of the glucopyranosyl rings and polar amino acid side chains lining the CBD(N2) cleft. Affinity electrophoresis was used to determine that CBD(N2) also binds soluble beta-1,4-linked polymers of glucose, including hydroxyethylcellulose and beta-1,3-1,4-glucans. This study complements a previous analysis of CBD(N1) [Johnson, P. E., Joshi, M. D., Tomme, P., Kilburn, D. G., and McIntosh, L. P. (1996) Biochemistry 35, 14381-14394] and demonstrates that the homologous CBDs from CenC share very similar structures and sugar binding properties. PMID- 10704196 TI - Cysteine scanning mutagenesis of helices 2 and 7 in GLUT1 identifies an exofacial cleft in both transmembrane segments. AB - Cysteine scanning mutagenesis in conjunction with site-directed chemical modification of sulfhydryl groups by p-chloromercuribenzenesulfonate (pCMBS) or N ethylmaleimide (NEM) was applied to putative transmembrane segments (TM) 2 and 7 of the cysteine-less glucose transporter GLUT1. Valid for both helices, the majority of cysteine substitution mutants functioned as active glucose transporters. The residues F72, G75, G76, G79, and S80 within helix 2 and G286 and N288 within helix 7 were irreplaceable because the mutant transporters displayed transport activities that were lower than 10% of Cys-less GLUT1. The indicated cluster of glycine residues within TM 2 is located on one face of the helix and may provide space for a bulky hydrophobic counterpart interacting with another transmembrane segment or lipid side chains. Characteristic for helix 7, three glutamine residues (Q279, Q282, and Q283) played an important role in transport activity of Cys-less GLUT1 because an individual replacement with cysteine reduced their transport rates by about 80%. ParaCMBS-sensitivity scanning of both transmembrane segments detected several membrane-harbored residues to be accessible to the extracellular aqueous solvent. The pCMBS reactive sulfhydryl groups were located exclusively in the exofacial half of the plasma membrane and, when presented in a helical model, lie along one side of the helices. Taken together, transmembrane segments 2 and 7 form clefts accessible to the extracellular aqueous solvent. The lining residues are however excluded from interaction with intracellular solutes, as justified by microinjection of pCMBS into the cytoplasm of Xenopus oocytes. PMID- 10704195 TI - Effect of synthetic sialyl 2-->1 sphingosine and other glycosylsphingosines on the structure and function of the "glycosphingolipid signaling domain (GSD)" in mouse melanoma B16 cells. AB - Mouse melanoma B16 cells are characterized by a high concentration of GM3 ganglioside, which has been identified as a melanoma-associated antigen and is present as a clustered microdomain organized with major signal transducers, c Src, small G-protein (Rho A), and focal adhesion kinase (FAK), to form a "glycosphingolipid signaling domain" or "glycosignaling domain" (GSD) separable from cholesterol- and caveolin-enriched microdomain, "caveolae." Cholesterol binding reagents, filipin and nystatin, disrupt the structure and function of caveolae, but have no effect on GSD function [Iwabuchi, K., et al. (1998) J. Biol. Chem. 273, 33766-33773]. In this study, we searched for compounds which disrupt the structure and function of GSD in B16 cells. Such compounds should have structural features analogous to those of GM3, destroy or reduce clustering of GM3 in GSD, and inhibit GM3-dependent adhesion and signaling. The simplest compound so far found with these properties is sialyl alpha2-->1 sphingosine (Sph). We describe the synthesis of this compound and its analogues, and their effects on GM3 expression pattern and GSD function, in comparison with effects of lyso-GM3 and other lyso compounds, in B16 cells. Incubation of B16 cells with 0.5 10 microM sialyl alpha2-->1 Sph or 1-5 microM lyso-GM3 reduced GM3 clustering and GM3-dependent adhesion, and inhibited adhesion-dependent cellular FAK activity. The c-Src activation response of GSD isolated from B16 cells was inhibited strongly by sialyl alpha2-->1 Sph. Substitution of the Sph amino group with a chloroacetyl or N,N-dimethyl group strongly reduced the inhibitory effect of sialyl alpha2-->1 Sph on GM3-dependent adhesion, FAK, and c-Src response. Other lyso compounds such as lyso-phosphatidylcholine, galactosyl-Sph (psychosine), and lactosyl-Sph at 0.5-10 microM did not show the same effect as sialyl alpha2-->1 Sph. Thus, adhesion coupled with signal transduction, initiated by clusters of GM3 in GSD, is blocked by sialyl alpha2-->1 Sph or lyso-GM3. Analogues with N substitution of Sph in sialyl alpha2-->1 Sph, other lyso-phospholipids, and galactosyl- or lactosyl-Sph did not block such adhesion, coupled with activation of c-Src and FAK. PMID- 10704197 TI - Annexin XII E105K crystal structure: identification of a pH-dependent switch for mutant hexamerization. AB - Annexins are a family of calcium- and phospholipid-binding proteins involved with numerous cellular processes including membrane fusion, ion channel activity, and heterocomplex formation with other proteins. The annexin XII (ANXB12) crystal structure presented evidence that calcium mediates the formation of a hexamer through a novel intermolecular calcium-binding site [Luecke et al. (1995) Nature 378, 512-515]. In an attempt to disrupt hexamerization, we mutated a conserved key ligand in the intermolecular calcium-binding site, Glu105, to lysine. Despite its occurrence in a new spacegroup, the 1.93 A resolution structure reveals a hexamer with the Lys105 epsilon-amino group nearly superimposable with the original intermolecular calcium position. Our analysis shows that the mutation is directly involved in stabilizing the hexamer. The local residues are reoriented to retain affinity between the two trimers via a pH-dependent switch residue, Glu76, which is now protonated, allowing it to form tandem hydrogen bonds with the backbone carbonyl and nitrogen atoms of Thr103 located across the trimer interface. The loss of the intermolecular calcium-binding site is recuperated by extensive hydrogen bonding favoring hexamer stabilization. The presence of this mutant structure provides further evidence for hexameric annexin XII, and possible in vivo roles are discussed. PMID- 10704198 TI - Homology modeling, molecular dynamics simulations, and analysis of CYP119, a P450 enzyme from extreme acidothermophilic archaeon Sulfolobus solfataricus. AB - The recent characterization of a thermophilic and barophilic CYP119 from Sulfolobus solfataricus offers a new opportunity to identify the origin of its stability by comparing it with mesophilic P450s with known structures. Since the three-dimensional structure of CYP119 is not yet available, homology modeling techniques were used to build model structures for this enzyme. The overall quality and stability of the models were assessed using three protein analysis programs and by monitoring structural stability during 1 ns of molecular dynamics simulations at 300 and 390 K. The results show the CYP119 models to be of good quality. Possible origins of the thermo- and barostability of CYP119 were then investigated by examining the amino acid compositions and the three-dimensional structure of CYP119 compared with the five mesophilic templates. Three possible factors were identified that could contribute to the enhanced stability of CYP119. The first was the higher relative population of salt bridges and the presence of a few unique salt bridges found in CYP119 that were absent in all five template CYP450s. The second factor was a decreased population of Ala and an increased population of Ile found in the interior of CYP119, which are likely to improve packing in CYP119. The third factor was a more extensive aromatic cluster seen in CYP119 which was not found in all five template P450s. In addition, the model CYP119 three-dimensional structures were also used to determine key properties related to its function. Specifically, binding site residues and surface residues for redox partner interactions were identified. These residues identified together with those residues found that might contribute to the increased stability are suggested for future mutagenesis studies. The results obtained from these experimental studies shall then provide further validation of the suggested origins of stability and the structure-function relationships derived from the model structures of this enzyme. PMID- 10704199 TI - Structures of the superoxide reductase from Pyrococcus furiosus in the oxidized and reduced states. AB - Superoxide reductase (SOR) is a blue non-heme iron protein that functions in anaerobic microbes as a defense mechanism against reactive oxygen species by catalyzing the reduction of superoxide to hydrogen peroxide [Jenney, F. E., Jr., Verhagen, M. F. J. M., Cui, X. , and Adams, M. W. W. (1999) Science 286, 306 309]. Crystal structures of SOR from the hyperthermophilic archaeon Pyrococcus furiosus have been determined in the oxidized and reduced forms to resolutions of 1.7 and 2.0 A, respectively. SOR forms a homotetramer, with each subunit adopting an immunoglobulin-like beta-barrel fold that coordinates a mononuclear, non-heme iron center. The protein fold and metal center are similar to those observed previously for the homologous protein desulfoferrodoxin from Desulfovibrio desulfuricans [Coelho, A. V., Matias, P., Fulop, V., Thompson, A., Gonzalez, A., and Carrondo, M. A. (1997) J. Bioinorg. Chem. 2, 680-689]. Each iron is coordinated to imidazole nitrogens of four histidines in a planar arrangement, with a cysteine ligand occupying an axial position normal to this plane. In two of the subunits of the oxidized structure, a glutamate carboxylate serves as the sixth ligand to form an overall six-coordinate, octahedral coordinate environment. In the remaining two subunits, the sixth coordination site is either vacant or occupied by solvent molecules. The iron centers in all four subunits of the reduced structure exhibit pentacoordination. The structures of the oxidized and reduced forms of SOR suggest a mechanism by which superoxide accessibility may be controlled and define a possible binding site for rubredoxin, the likely physiological electron donor to SOR. PMID- 10704200 TI - High resolution solution structure of the 1.3S subunit of transcarboxylase from Propionibacterium shermanii. AB - Transcarboxylase (TC) from Propionibacterium shermanii, a biotin-dependent enzyme, catalyzes the transfer of a carboxyl group from methylmalonyl-CoA to pyruvate to form propionyl-CoA and oxalacetate. Within the multi-subunit enzyme complex, the 1.3S subunit functions as the carboxyl group carrier and also binds the other two subunits to assist in the overall assembly of the enzyme. The 1.3S subunit is a 123 amino acid polypeptide (12.6 kDa) to which biotin is covalently attached at Lys 89. The three-dimensional solution structure of the full-length holo-1.3S subunit of TC has been solved by multidimensional heteronuclear NMR spectroscopy. The C-terminal half of the protein (51-123) is folded into a compact all-beta-domain comprising of two four-stranded antiparallel beta-sheets connected by short loops and turns. The fold exhibits a high 2-fold internal symmetry and is similar to that of the biotin carboxyl carrier protein (BCCP) of acetyl-CoA carboxylase, but lacks an extension that has been termed "protruding thumb" in BCCP. The first 50 residues, which have been shown to be involved in intersubunit interactions in the intact enzyme, appear to be disordered in the isolated 1.3S subunit. The molecular surface of the folded domain has two distinct surfaces: one side is highly charged, while the other comprises mainly hydrophobic, highly conserved residues. PMID- 10704201 TI - Porphyrin interactions with wild-type and mutant mouse ferrochelatase. AB - Ferrochelatase (EC 4.99.1.1), the terminal enzyme of the heme biosynthetic pathway, catalyzes Fe(2+) chelation into protoporphyrin IX. Resonance Raman and UV-vis absorption spectroscopies of wild-type and engineered variants of murine ferrochelatase were used to examine the proposed structural mechanism for iron insertion into porphyrin. The recombinant variants (i.e., H207N and E287Q) are enzymes in which the conserved amino acids histidine-207 and glutamate-287 of murine ferrochelatase were substituted with asparagine and glutamine, respectively. Both of these residues are at the active site of the enzyme as deduced from the Bacillus subtilis ferrochelatase three-dimensional structure. On the basis of changes in the UV-vis absorption spectrum, addition of free-base or metalated porphyrins to wild-type ferrochelatase and H207N variant yields a 1:1 complex, most likely a monomeric protein-bound species at the active site. In contrast, the addition of porphyrin (either free base or metalated) to E287Q is substoichiometric, as this variant retains bound porphyrin in the active site during isolation and purification. The specificity of porphyrin binding is confirmed by the narrowing of the structure-sensitive lines and the vinyl vibrational mode in the resonance Raman spectra. Shifts in the resonance Raman lines of free-base and metalated porphyrins bound to the wild-type ferrochelatase indicate a nonplanar distortion of the porphyrin macrocycle. However, the magnitude of the distortion cannot be determined without first defining the specific type of deformation. Significantly, the extent of the nonplanar distortion varies in the case of H207N- and E287Q-bound porphyrins. In fact, resonance Raman spectral decompositions indicate a homogeneous ruffled deformation for the nickel protoporphyrin bound to the wild-type ferrochelatase, whereas both planar and ruffled conformations are present for the H207N-bound porphyrin. Perhaps more revealing is the unusual resonance Raman spectrum of the endogenous E287Q-bound porphyrin, which has the structure-sensitive lines greatly upshifted relative to those of the free-base protoporphyrin in solution. This could be interpreted as an equilibrium between protein conformers, one of which favors a highly distorted porphyrin macrocycle. Taken together, these findings suggest that distortion occurs in murine ferrochelatase for some porphyrins, even without metal binding, which is apparently required for the yeast ferrochelatase. PMID- 10704202 TI - Heteronuclear NMR and soft docking: an experimental approach for a structural model of the cytochrome c553-ferredoxin complex. AB - The combination of docking algorithms with NMR data has been developed extensively for the studies of protein-ligand interactions. However, to extend this development for the studies of protein-protein interactions, the intermolecular NOE constraints, which are needed, are more difficult to access. In the present work, we describe a new approach that combines an ab initio docking calculation and the mapping of an interaction site using chemical shift variation analysis. The cytochrome c553-ferredoxin complex is used as a model of numerous electron-transfer complexes. The 15N-labeling of both molecules has been obtained, and the mapping of the interacting site on each partner, respectively, has been done using HSQC experiments. 1H and 15N chemical shift analysis defines the area of both molecules involved in the recognition interface. Models of the complex were generated by an ab initio docking software, the BiGGER program (bimolecular complex generation with global evaluation and ranking). This program generates a population of protein-protein docked geometries ranked by a scoring function, combining relevant stabilization parameters such as geometric complementarity surfaces, electrostatic interactions, desolvation energy, and pairwise affinities of amino acid side chains. We have implemented a new module that includes experimental input (here, NMR mapping of the interacting site) as a filter to select the accurate models. Final structures were energy minimized using the X-PLOR software and then analyzed. The best solution has an interface area (1037.4 A2) falling close to the range of generally observed recognition interfaces, with a distance of 10.0 A between the redox centers. PMID- 10704203 TI - Systematic mutational mapping of sites on human interferon-beta-1a that are important for receptor binding and functional activity. AB - A systematic mutational analysis of human interferon-beta-1a (IFN-beta) was performed to identify regions on the surface of the molecule that are important for receptor binding and for functional activity. The crystal structure of IFN beta-1a was used to design a panel of 15 mutant proteins, in each of which a contiguous group of 2-8 surface residues was mutated, in most instances to alanine. The mutants were analyzed for activity in vitro in antiviral and in antiproliferation assays, and for their ability to bind to the type I IFN (ifnar1/ifnar2) receptor on Daudi cells and to a soluble ifnar2 fusion protein (ifnar2-Fc). Abolition of binding to ifnar2-Fc for mutants A2, AB1, AB2, and E established that the ifnar2 binding site on IFN-beta comprises parts of the A helix, the AB loop, and the E helix. Mutations in these areas, which together define a contiguous patch of the IFN-beta surface, also resulted in reduced affinity for binding to the receptor on cells and in reductions in activity of 5 50-fold in functional assays. A second receptor interaction site, concluded to be the ifnar1 binding site, was identified on the opposite face of the molecule. Mutations in this region, which encompasses parts of the B, C, and D helices and the DE loop, resulted in disparate effects on receptor binding and on functional activity. Analysis of antiproliferation activity as a function of the level of receptor occupancy allowed mutational effects on receptor activation to be distinguished from effects on receptor binding. The results suggest that the binding energy from interaction of IFN-beta with ifnar2 serves mainly to stabilize the bound IFN/receptor complex, whereas the binding energy generated by interaction of certain regions of IFN-beta with ifnar1 is not fully expressed in the observed affinity of binding but instead serves to selectively stabilize activated states of the receptor. PMID- 10704204 TI - Fibrils formed in vitro from alpha-synuclein and two mutant forms linked to Parkinson's disease are typical amyloid. AB - Two missense mutations in the gene encoding alpha-synuclein have been linked to rare, early-onset forms of Parkinson's disease (PD). These forms of PD, as well as the common idiopathic form, are characterized by the presence of cytoplasmic neuronal deposits, called Lewy bodies, in the affected region of the brain. Lewy bodies contain alpha-synuclein in a form that resembles fibrillar Abeta derived from Alzheimer's disease (AD) amyloid plaques. One of the mutant forms of alpha synuclein (A53T) fibrillizes more rapidly in vitro than does the wild-type protein, suggesting that a correlation may exist between the rate of in vitro fibrillization and/or oligomerization and the progression of PD, analogous to the relationship between Abeta fibrillization in vitro and familial AD. In this paper, fibrils generated in vitro from alpha-synuclein, wild-type and both mutant forms, are shown to possess very similar features that are characteristic of amyloid fibrils, including a wound and predominantly unbranched morphology (demonstrated by atomic force and electron microscopies), distinctive dye-binding properties (Congo red and thioflavin T), and antiparallel beta-sheet structure (Fourier transform infrared spectroscopy and circular dichroism spectroscopy). alpha-Synuclein fibrils are relatively resistant to proteolysis, a property shared by fibrillar Abeta and the disease-associated fibrillar form of the prion protein. These data suggest that PD, like AD, is a brain amyloid disease that, unlike AD, is characterized by cytoplasmic amyloid (Lewy bodies). In addition to amyloid fibrils, a small oligomeric form of alpha-synuclein, which may be analogous to the Abeta protofibril, was observed prior to the appearance of fibrils. This species or a related one, rather than the fibril itself, may be responsible for neuronal death. PMID- 10704205 TI - Solution structure of the sixth LDL-A module of the LDL receptor. AB - The low-density lipoprotein receptor (LDLR) is the primary mechanism for uptake of plasma cholesterol into cells and serves as a prototype for an entire class of cell surface receptors. The amino-terminal domain of the receptor consists of seven LDL-A modules; the third through the seventh modules all contribute to the binding of low-density lipoproteins (LDLs). Here, we present the NMR solution structure of the sixth LDL-A module (LR6) from the ligand binding domain of the LDLR. This module, which has little recognizable secondary structure, retains the essential structural features observed in the crystal structure of LDL-A module five (LR5) of the LDLR. Three disulfide bonds, a pair of buried residues forming a hydrophobic "mini-core", and a calcium-binding site that serves to organize the C-terminal lobe of the module all occupy positions in LR6 similar to those observed in LR5. The striking presence of a conserved patch of negative surface electrostatic potential among LDL-A modules of known structure suggests that ligand recognition by these repeats is likely to be mediated in part by electrostatic complementarity of receptor and ligand. Two variants of LR6, identified originally as familial hypercholesterolemia (FH) mutations, have been investigated for their ability to form native disulfide bonds under conditions that permit disulfide exchange. The first, E219K, lies near the amino-terminal end of LR6, whereas the second, D245E, alters one of the aspartate side chains that directly coordinate the bound calcium ion. After equilibration at physiologic calcium concentrations, neither E219K nor D245E folds to a unique disulfide isomer, indicating that FH mutations both within and distant from the calcium-binding site give rise to protein-folding defects. PMID- 10704206 TI - Solution structure of the PDZ2 domain from human phosphatase hPTP1E and its interactions with C-terminal peptides from the Fas receptor. AB - The solution structure of the second PDZ domain (PDZ2) from human phosphatase hPTP1E has been determined using 2D and 3D heteronuclear NMR experiments. The binding of peptides derived from the C-terminus of the Fas receptor to PDZ2 was studied via changes in backbone peptide and protein resonances. The structure is based on a total of 1387 nonredundant experimental NMR restraints including 1261 interproton distance restraints, 45 backbone hydrogen bonds, and 81 torsion angle restraints. Analysis of 30 lowest-energy structures resulted in rmsd values of 0.41 +/- 0.09 A for backbone atoms (N, Calpha, C') and 1.08 +/- 0.10 A for all heavy atoms, excluding the disordered N- and C-termini. The hPTP1E PDZ2 structure is similar to known PDZ domain structures but contains two unique structural features. In the peptide binding domain, the first glycine of the GLGF motif is replaced by a serine. This serine appears to replace a bound water observed in PDZ crystal structures that hydrogen bonds to the bound peptide's C-terminus. The hPTP1E PDZ2 structure also contains an unusually large loop following strand beta2 and proximal to the peptide binding site. This well-ordered loop folds back against the PDZ domain and contains several residues that undergo large amide chemical shift changes upon peptide binding. Direct observation of peptide resonances demonstrates that as many as six Fas peptide residues interact with the PDZ2 domain. PMID- 10704207 TI - Adenosine deaminase prefers a distinct sugar ring conformation for binding and catalysis: kinetic and structural studies. AB - Several recent X-ray crystal structures of adenosine deaminase (ADA) in complex with various adenosine surrogates have illustrated the preferred mode of substrate binding for this enzyme. To define more specific structural details of substrate preferences for binding and catalysis, we have studied the ADA binding efficiencies and deamination kinetics of several synthetic adenosine analogues in which the furanosyl ring is biased toward a particular conformation. NMR solution studies and pseudorotational analyses were used to ascertain the preferred furanose ring puckers (P, nu(MAX)) and rotamer distributions (chi and gamma) of the nucleoside analogues. It was shown that derivatives which are biased toward a "Northern" (3'-endo, N) sugar ring pucker were deaminated up to 65-fold faster and bound more tightly to the enzyme than those that preferred a "Southern" (2' endo, S) conformation. This behavior, however, could be modulated by other structural factors. Similarly, purine riboside inhibitors of ADA that prefer the N hemisphere were more potent inhibitors than S analogues. These binding propensities were corroborated by detailed molecular modeling studies. Docking of both N- and S-type analogues into the ADA crystal structure coordinates showed that N-type substrates formed a stable complex with ADA, whereas for S-type substrates, it was necessary for the sugar pucker to adjust to a 3'-endo (N-type) conformation to remain in the ADA substrate binding site. These data outline the intricate structural details for optimum binding in the catalytic cleft of ADA. PMID- 10704208 TI - Mechanistic studies of rat protein farnesyltransferase indicate an associative transition state. AB - Protein farnesyltransferase is a zinc metalloenzyme that catalyzes the transfer of a 15-carbon farnesyl group to a conserved cysteine residue of a protein substrate. Both electrophilic and nucleophilic mechanisms have been proposed for this enzyme. In this work, we investigate the detailed catalytic mechanism of mammalian protein farnesyltransferase by measuring the effect of metal substitution and/or substrate alterations on the rate constant of the chemical step. Substitution of cadmium for the active site zinc enhances peptide affinity approximately 5-fold and decreases the rate constant for the formation of the thioether product approximately 6-fold, indicating changes in the metal-thiolate coordination in the catalytic transition state. In addition, the observed rate constant for product formation decreases for C3 fluoromethyl farnesyl pyrophosphate substrates, paralleling the number of fluorines at the C3 methyl position and indicating that a rate-contributing transition state has carbocation character. Magnesium ions do not affect the affinity of either the peptide or the isoprenoid substrate but specifically enhance the observed rate constant for product formation 700-fold, suggesting that magnesium coordinates and activates the diphosphate leaving group. These data suggest that FTase catalyzes protein farnesylation by an associative mechanism with an "exploded" transition state where the metal-bound peptide/protein sulfur has a partial negative charge, the C1 of FPP has a partial positive charge, and the bridge oxygen between C1 and the alpha phosphate of FPP has a partial negative charge. This proposed transition state suggests that stabilization of the developing charge on the carbocation and pyrophosphate oxygens is an important catalytic feature. PMID- 10704209 TI - The action of soybean lipoxygenase-1 on 12-iodo-cis-9-octadecenoic acid: the importance of C11-H bond breaking. AB - Previous work has demonstrated that the ferric form of soybean lipoxygenase-1 will catalyze an elimination reaction on 12-iodo-cis-9-octadecenoic acid (12 IODE) to produce 9, 11-octadecadienoic acid and iodide ion. Elimination is accompanied by irreversible inactivation of the enzyme on 1 out of 10 turnovers. In the present work, 11,11-dideuterio-12-IODE (D(2)-12-IODE) was synthesized and used to demonstrate that both the elimination reaction and inactivation of the enzyme exhibit very large kinetic isotope effects. The rates with the deuterated compound are so low that the isotope effects are difficult to quantify, but they appear to be comparable to the isotope effects previously observed for the normal reaction catalyzed by lipoxygenase and much larger than can be explained by zero point energy considerations. ESR spectroscopy was used to demonstrate that 12 IODE can reduce ferric lipoxygenase to the ferrous form, and a large isotope effect on this process was observed with D(2)-12-IODE. It is proposed that the pathway leading to reduction and inactivation by 12-IODE is initiated by homolytic cleavage of the C(11)-H bond. Elimination could be initiated either by homolytic or by heterolytic cleavage of this bond. The results suggest that very large isotope effects may be a general feature of C-H bond cleavages catalyzed by this enzyme. PMID- 10704210 TI - C5 cytosine methylation at CpG sites enhances sequence selectivity of mitomycin C DNA bonding. AB - We have established that UvrABC nuclease is equally efficient in cutting mitomycin C (MC)-DNA monoadducts formed at different sequences and that the degree of UvrABC cutting represents the extent of drug-DNA bonding. Using this method we determined the effect of C5 cytosine methylation on the DNA monoalkylation by MC and the related analogues N-methyl-7 methoxyaziridinomitosene (MS-NMA) and 10-decarbamoylmitomycin C (DC-MC). We have found that C5 cytosine methylation at CpG sites greatly enhances MC and MS-NMA DNA adduct formation at those sites while reducing adduct formation at non-CpG sequences. In contrast, although DC-MC DNA bonding at CpG sites is greatly enhanced by CpG methylation, its bonding at non-CpG sequences is not appreciably affected. These cumulative results suggest that C5 cytosine methylation at CpG sites enhances sequence selectivity of drug-DNA bonding. We propose that the methylation pattern and status (hypo- or hypermethylation) of genomic DNA may determine the cells' susceptibility to MC and its analogues, and these effects may, in turn, play a crucial role in the antitumor activities of the drugs. PMID- 10704211 TI - Probing the relationship between RNA-stimulated ATPase and helicase activities of HCV NS3 using 2'-O-methyl RNA substrates. AB - The hepatitis C virus (HCV) NS3 protein contains an amino terminal protease (NS3 aa. 1-180) and a carboxyl terminal RNA helicase (NS3 aa. 181-631). NS3 functions as a heterodimer of NS3 and NS4A (NS3/4A). NS3 helicase, a nucleic acid stimulated ATPase, can unwind RNA, DNA, and RNA:DNA duplexes, provided that at least one strand of the duplex contains a single-stranded 3' overhang (this strand of the duplex is referred to as the 3' strand). We have used 2'-O-methyl RNA (MeRNA) substrates to study the mechanism of NS3 helicase activity and to probe the relationship between its helicase and RNA-stimulated ATPase activities. NS3/4A did not unwind double-stranded (ds) MeRNA. NS3/4A unwinds hybrid RNA:MeRNA duplex containing MeRNA as the 5' strand but not hybrid duplex containing MeRNA as the 3' strand. The helicase activity of NS3/4A was 50% inhibited by 40 nM single-stranded (ss) RNA but only 35% inhibited by 320 nM ss MeRNA. Double stranded RNA was 17 times as effective as double-stranded MeRNA in inhibiting NS3/4A helicase activity, while the apparent affinity of NS3/4A for ds MeRNA differed from ds RNA by only 2.4-fold. However ss MeRNA stimulated NS3/4A ATPase activity similar to ss RNA. These results indicate that the helicase mechanism involves 3' to 5' procession of the NS3 helicase along the 3' strand and only weak association of the enzyme with the displaced 5' strand. Further, our findings show that maximum stimulation of NS3 ATPase activity by ss nucleic acid is not directly related to procession of the helicase along the 3' strand. PMID- 10704212 TI - Importance of terminal base pair hydrogen-bonding in 3'-end proofreading by the Klenow fragment of DNA polymerase I. AB - We describe studies aimed at evaluating the physical factors governing the rate of 3'-end proofreading by the Klenow fragment of E. coli DNA polymerase I. Two nonpolar deoxynucleoside isosteres containing 2,4-difluorotoluene (F) and 4 methylbenzimidazole (Z), which are non-hydrogen-bonding shape mimics of thymine and adenine, respectively, are used to investigate the effects of base pair geometry and stability on the rate of this exonuclease activity. Steady-state kinetics measurements show that complementary T.A base pairs at the end of a primer-template duplex are edited 14-40-fold more slowly than mismatches. By contrast, a 3'-end T residue in a T. Z pair is edited at a rate equivalent to that of natural base mismatches despite the fact that it resembles a T.A pair in structure. Similarly, the A in an A.F pair is edited as rapidly as a mismatched pair despite its close structural mimicry of an A.T pair. Interestingly, when the base pairs are reversed and F or Z is located at the 3'-end, they are edited more slowly, possibly implicating specific interactions between the exonuclease domain and the base of the nucleotide being edited. Finally, thermal denaturation studies are carried out to investigate the relationship between editing and the ease of unwinding of the duplex. The rapid editing of bases opposite F or Z residues at the duplex terminus seems to correlate well with the stability of these base pairs when placed in a context resembling a primer-template duplex. In general, the rate of 3'-end editing appears to be governed by the rate of fraying of the DNA terminal pair, and base pair geometry appears to have little effect. PMID- 10704213 TI - Reevaluation of transcriptional regulation by TATA-binding protein oligomerization: predominance of monomers. AB - The TATA-binding protein (TBP) plays an important role in transcriptional initiation by all three nuclear RNA polymerases. TBP contains a conserved C terminal domain (cTBP) that binds DNA. Crystallographic studies of cTBP (i.e., TBP without the N-terminal domain) from various species and molecular biology studies of cTBP and mixed cTBP/TBP species have led to the view that DNA binding by TBP is regulated by TBP dimerization. Using sedimentation equilibrium, we show that yeast cTBP forms dimers in solution at 5 degrees C with a dissociation constant of 7 +/- 1 microM. This observation of cTBP dimers in solution is in accord with the dimeric state observed in crystal structures of cTBP. In contrast, physiologically relevant, full-length yeast TBP is monomeric at 5 degrees C and forms dimers at 30 degrees C with a dissociation constant of 51 +/- 16 microM. This dissociation constant precludes formation of stable full-length TBP dimers at physiological concentrations. In addition, we tested for yeast TBP oligomerization in the presence of TBP-associated factors in the context of TFIID. No evidence for TBP oligomers was found using immunoprecipitation techniques from yeast whole-cell extracts. We conclude that yeast TBP is predominantly monomeric under physiological conditions, arguing against a role for TBP dimerization in the regulation of transcriptional initiation. PMID- 10704214 TI - The P5abc peripheral element facilitates preorganization of the tetrahymena group I ribozyme for catalysis. AB - Phylogenetic comparisons and site-directed mutagenesis indicate that group I introns are composed of a catalytic core that is universally conserved and peripheral elements that are conserved only within intron subclasses. Despite this low overall conservation, peripheral elements are essential for efficient splicing of their parent introns. We have undertaken an in-depth structure function analysis to investigate the role of one of these elements, P5abc, using the well-characterized ribozyme derived from the Tetrahymena group I intron. Structural comparisons using solution-based free radical cleavage revealed that a ribozyme lacking P5abc (E(DeltaP5abc)) and E(DeltaP5abc) with P5abc added in trans (E(DeltaP5abc).P5abc) adopt a similar global tertiary structure at Mg(2+) concentrations greater than 20 mM [Doherty, E. A., et al. (1999) Biochemistry 38, 2982-90]. However, free E(DeltaP5abc) is greatly compromised in overall oligonucleotide cleavage activity, even at Mg(2+) concentrations as high as 100 mM. Further characterization of E(DeltaP5abc) via DMS modification revealed local structural differences at several positions in the conserved core that cluster around the substrate binding sites. Kinetic and thermodynamic dissection of individual reaction steps identified defects in binding of both substrates to E(DeltaP5abc), with > or =25-fold weaker binding of a guanosine nucleophile and > or =350-fold weaker docking of the oligonucleotide substrate into its tertiary interactions with the ribozyme core. These defects in binding of the substrates account for essentially all of the 10(4)-fold decrease in overall activity of the deletion mutant. Together, the structural and functional observations suggest that the P5abc peripheral element not only provides stability but also positions active site residues through indirect interactions, thereby preferentially stabilizing the active ribozyme structure relative to alternative less active states. This is consistent with the view that peripheral elements engage in a network of mutually reinforcing interactions that together ensure cooperative folding of the ribozyme to its active structure. PMID- 10704215 TI - Kinetic parameters for tmRNA binding to alanyl-tRNA synthetase and elongation factor Tu from Escherichia coli. AB - Aminoacylation and transportation of tmRNA to stalled ribosomes constitute prerequisite steps for trans-translation, a process facilitating the release of stalled ribosomes from 3' ends of truncated mRNAs and the degradation of incompletely synthesized proteins. Kinetic analysis of the aminoacylation of tmRNA indicates that tmRNA has both a lower affinity and a lower turnover number than cognate tRNA(Ala) for alanyl-tRNA synthetase, resulting in a 75-fold lower k(cat)/K(M) value. The association rate constant of Ala-tmRNA for elongation factor Tu in complex with GTP is about 150-fold lower than that of Ala-tRNA(Ala), whereas its dissocation rate constant is about 5-fold lower. These observations can be interpreted to suggest that additional factors facilitate tmRNA binding to ribosomes. PMID- 10704216 TI - In vitro reconstitution of the Schizosaccharomyces pombe alternative excision repair pathway. AB - Schizosaccharomyces pombe alternative excision repair has been shown genetically and biochemically to be involved in the repair of a wide variety of DNA lesions. AER is initiated by a damage-specific endonuclease (Uve1p) that recognizes UV induced photoproducts, base mispairs, abasic sites, and platinum G-G diadducts and cleaves the DNA phosphodiester backbone 5' to a lesion. Several models exist that employ various mechanisms for damage removal based on the activities of Rad2p, a nuclease thought to be responsible for damage excision in AER. This study represents the first report of the biochemical reconstitution of the AER pathway. A base mispair-containing substrate is repaired in a reaction requiring S. pombe Uve1p, Rad2p, DNA polymerase delta, replication factor C, proliferating cell nuclear antigen, and T4 DNA ligase. Surprisingly, damage is removed exclusively by the 5' to 3' exonuclease activity of Rad2p and not its "flap endonuclease" activity and is absolutely dependent upon the presence of the 5' phosphoryl moiety at the Uve1p cleavage site. PMID- 10704217 TI - Probing the photoreaction mechanism of phytochrome through analysis of resonance Raman vibrational spectra of recombinant analogues. AB - Resonance Raman spectra of native and recombinant analogues of oat phytochrome have been obtained and analyzed in conjunction with normal mode calculations. On the basis of frequency shifts observed upon methine bridge deuteration and vinyl and C(15)-methine bridge saturation of the chromophore, intense Raman lines at 805 and 814 cm(-)(1) in P(r) and P(fr), respectively, are assigned as C(15) hydrogen out-of-plane (HOOP) wags, lines at 665 cm(-)(1) in P(r) and at 672 and 654 cm(-)(1) in P(fr) are assigned as coupled C=C and C-C torsions and in-plane ring twisting modes, and modes at approximately 1300 cm(-)(1) in P(r) are coupled N-H and C-H rocking modes. The empirical assignments and normal mode calculations support proposals that the chromophore structures in P(r) and P(fr) are C(15) Z,syn and C(15)-E,anti, respectively. The intensities of the C(15)-hydrogen out of-plane, C=C and C-C torsional, and in-plane ring modes in both P(r) and P(fr) suggest that the initial photochemistry involves simultaneous bond rotations at the C(15)-methine bridge coupled to C(15)-H wagging and D-ring rotation. The strong nonbonded interactions of the C- and D-ring methyl groups in the C(15) E,anti P(fr) chromophore structure indicated by the intense 814 cm(-1) C(15) HOOP mode suggest that the excited state of P(fr) and its photoproduct states are strongly coupled. PMID- 10704218 TI - Intensity of the polarized Raman band at 1340-1345 cm-1 as an indicator of protein alpha-helix orientation: application to Pf1 filamentous virus. AB - Raman spectra of oriented alpha-helical protein molecules exhibit a prominent band near 1340-1345 cm(-)(1), the intensity of which is highly sensitive to molecular orientation. Polarization of the 1340-1345 cm(-)(1) marker is evident in Raman spectra of alpha-helical poly-L-alanine (alphaPLA) and alpha-helical poly-gamma-benzyl-L-glutamate (alphaPBLG). Corresponding polarization is also observed in Raman spectra of the filamentous virus Pf1, which is an assembly of alpha-helical coat protein molecules. In alphaPLA and alphaPBLG, we assign the band to a normal mode of symmetry type E(2) and specifically to a vibration localized in the (O=C)-C(alpha)-H linkages of the main chain peptide group. Although strict helical symmetry does not apply to coat subunits of filamentous viruses, an approximate E(2)-type mode may be presumed to account for a corresponding Raman band of Pf1 and fd filamentous viruses. Spectroscopic studies of N-methylacetamide and isotopically-edited fd viruses support the present assignment of the 1340-1345 cm(-)(1) band. Polarization anisotropy indicates that this band may be exploited as a novel indicator of protein alpha-helix orientation. Application of this approach to the polarized Raman spectrum of Pf1 suggests that, on average, the axis of the alpha-helical coat protein subunit in the native virion structure forms an angle of 20 +/- 10 degrees with respect to the virion axis. PMID- 10704219 TI - Dos, a heme-binding PAS protein from Escherichia coli, is a direct oxygen sensor. AB - A direct sensor of O(2), the Dos protein, has been found in Escherichia coli. Previously, the only biological sensors known to respond to O(2) by direct and reversible binding were the FixL proteins of Rhizobia. A heme-binding region in Dos is 60% homologous to the O(2)-sensing PAS domain of the FixL protein, but the remainder of Dos does not resemble FixL. Specifically, the C-terminal domain of Dos, presumed to be a regulatory partner that couples to its heme-binding domain, is not a histidine kinase but more closely resembles a phosphodiesterase. The absorption spectra of Dos indicate that both axial positions of the heme iron are coordinated to side chains of the protein. Nevertheless, O(2) and CO bind to Dos with K(d) values of 13 and 10 microM, respectively, indicating a strong discrimination against CO binding. Association rate constants for binding of O(2) (3 mM(-)(1) s(-)(1)), CO (1 mM(-)(1) s(-)(1)) and even NO (2 mM(-)(1) s(-)(1)) are extraordinarily low and very similar. Displacement of an endogenous ligand, probably Met 95, from the heme iron in Dos triggers a conformational change that alters the activity of the enzymatic domain. This sensing mechanism differs from that of FixL but resembles that of the CO sensor CooA of Rhodospirillum rubrum. Overall the results provide evidence for a heme-binding subgroup of PAS-domain proteins whose working range, signaling mechanisms, and regulatory partners can vary considerably. PMID- 10704220 TI - Movement of the Rieske iron-sulfur protein in the p-side bulk aqueous phase: effect of lumenal viscosity on redox reactions of the cytochrome b6f complex. AB - Based on the atomic structures of the mitochondrial cytochrome bc(1) complex, it has been proposed that the soluble domain of the [2Fe-2S] Rieske iron-sulfur protein (ISP) must rotate by ca. 60 degrees and translate through an appreciable distance between two binding sites, proximal to cytochrome c(1) and to the lumen side quinol binding site. Such motional freedom implies that the electron transfer rate should be affected by the lumenal viscosity. The flash-induced oxidation of cytochrome f, the chloroplast analogue of cytochrome c(1), was found to be inhibited reversibly by increased lumenal viscosity, as was the subsequent reduction of both cytochrome b(6) and cytochrome f. The rates of these three redox reactions correlated inversely with lumenal viscosity over a viscosity range of 1-10 cP. Reduction of cytochrome b(6) and cytochrome f was not concerted. The rate of cytochrome f reduction was observed to be approximately half that of cytochrome b(6) regardless of the actual viscosity, implying that the path length traversed by the ISP in reduction of cytochrome f is twice that of cytochrome b(6). This suggests that upon initiation of electron transfer by a light flash, cytochrome b(6) reduction requires movement of reduced ISP from an initial position predominantly proximal to cytochrome f, apparently favored by the reduced ISP, to the quinol binding site at which the oxidant-induced reduction of cytochrome b(6) is initiated. Subsequent reduction of cytochrome f requires the additional movement of the ISP back to a site proximal to cytochromef. There is no discernible viscosity dependence for cytochrome b(6) reduction under oxidizing conditions, presumably because the oxidized ISP preferentially binds proximal to the quinone binding niche. The dependence of the cytochrome redox reaction on ambient viscosity implies that the tethered diffusional motion of the ISP is part of the rate limitation for charge transfer through the b(6)f complex. PMID- 10704221 TI - Analysis of the electron paramagnetic resonance properties of the [2Fe-2S]1+ centers in molybdenum enzymes of the xanthine oxidase family: assignment of signals I and II. AB - Molybdoenzymes of the xanthine oxidase family contain two [2Fe-2S](1+,2+) clusters that are bound to the protein by very different cysteine motifs. In the X-ray crystal structure of Desulfovibrio gigas aldehyde oxidoreductase, the cluster ligated by a ferredoxin-type motif is close to the protein surface, whereas that ligated by an unusual cysteine motif is in contact with the molybdopterin [Romao, M. J., Archer, M., Moura, I., Moura, J. J. G., LeGall, J., Engh, R., Schneider, M., Hof, P., and Huber, R. (1995) Science 270, 1170-1176]. These two clusters display distinct electron paramagnetic resonance (EPR) signals: the less anisotropic one, called signal I, is generally similar to the g(av) approximately 1.96-type signals given by ferredoxins, whereas signal II often exhibits anomalous properties such as very large g values, broad lines, and very fast relaxation properties. A detailed comparison of the temperature dependence of the spin-lattice relaxation time and of the intensity of these signals in D. gigas aldehyde oxidoreductase and in milk xanthine oxidase strongly suggests that the peculiar EPR properties of signal II arise from the presence of low-lying excited levels reflecting significant double exchange interactions. The issue raised by the assignment of signals I and II to the two [2Fe-2S](1+) clusters was solved by using the EPR signal of the Mo(V) center as a probe. The temperature dependence of this signal could be quantitatively reproduced by assuming that the Mo(V) center is coupled to the cluster giving signal I in xanthine oxidase as well as in D. gigas aldehyde oxidoreductase. This demonstrates unambiguously that, in both enzymes, signal I arises from the center which is closest to the molybdenum cofactor. PMID- 10704222 TI - Intrinsic structural disorder of the C-terminal activation domain from the bZIP transcription factor Fos. AB - The bZIP proto-oncoprotein c-Fos activates transcription of a wide variety of genes involved in cell growth. The C-terminal activation domain of c-Fos is functionally independent of the remainder of the protein. Fos-AD corresponds to the C-terminal activation domain of human c-Fos (residues 216-380). Fos-AD suppresses (squelches) transcription in vitro, as expected for a functional activation domain lacking a DNA-binding domain. Fos-AD is unstructured and highly mobile, as demonstrated by circular dichroism spectra indicative of unfolded proteins, a lack of (1)H chemical shift dispersion, and negative (1)H-(15)N heteronuclear nuclear Overhauser effects. The hydrodynamic properties of Fos-AD are also consistent with an extended structure. We conclude that the C-terminal domain of human c-Fos is biologically active yet intrinsically disordered. Our results suggest that conformational disorder is an integral aspect of the diverse contributions to transcriptional regulation by c-Fos. PMID- 10704223 TI - Transient structure of the amyloid precursor protein cytoplasmic tail indicates preordering of structure for binding to cytosolic factors. AB - The cytoplasmic tail of the amyloid precursor protein (APP) appears to play two important roles in the cell through participation in intracellular signaling and proteolytic processing of APP. Hence, knowledge of the structure of the 47 residue cytoplasmic tail of APP is important for understanding the molecular interactions involved in normal cell function as well as in the pathogenesis of Alzheimer's disease. Multidimensional solution NMR spectroscopy has been applied to examine the structural features of a 49-residue peptide (APP-C) containing two N-terminal residues (GS) and the APP cytoplasmic tail, over the pH range of 4.2 7.1. Although the peptide does not adopt a stable folded structure, regions of unstable structure exist over the pH range examined and have been characterized by a combination of H(alpha) chemical shifts, NOE analysis, and (3)J(HNH)(alpha) coupling constants and by identification of transient hydrogen bonds between amide protons and titrating carboxylate groups. These studies extend the work of others [Kroenke et al. (1997) Biochemistry 36, 8145-8152] by identifying an additional nascent helix and a hydrophobic cluster within the N-terminal 20 amino acid residues and by further characterizing the TPEE turn as a helix capping box. The transient structure of APP-C provides insight into the importance of preordering of this cytoplasmic tail in governing specificity and affinity for cytosolic binding partners. PMID- 10704224 TI - Action of quinolones against Staphylococcus aureus topoisomerase IV: basis for DNA cleavage enhancement. AB - Topoisomerase IV is the primary cellular target for most quinolones in Gram positive bacteria; however, its interaction with these agents is poorly understood. Therefore, the effects of four clinically relevant antibacterial quinolones (ciprofloxacin, and three new generation quinolones: trovafloxacin, levofloxacin, and sparfloxacin) on the DNA cleavage/religation reaction of Staphylococcus aureus topoisomerase IV were characterized. These quinolones stimulated enzyme-mediated DNA scission to a similar extent, but their potencies varied significantly. Drug order in the absence of ATP was trovafloxacin > ciprofloxacin > levofloxacin > sparfloxacin. Potency was enhanced by ATP, but to a different extent for each drug. Under all conditions examined, trovafloxacin was the most potent quinolone and sparfloxacin was the least. The enhanced potency of trovafloxacin correlated with several properties. Trovafloxacin induced topoisomerase IV-mediated DNA scission more rapidly than other quinolones and generated more cleavage at some sites. The most striking correlation, however, was between quinolone potency and inhibition of enzyme-mediated DNA religation: the greater the potency, the stronger the inhibition. Dose-response experiments with two topoisomerase IV mutants that confer clinical resistance to quinolones (GrlA(Ser80Phe) and GrlA(Glu84Lys)) indicate that resistance is caused by a decrease in both drug affinity and efficacy. Trovafloxacin is more active against these enzymes than ciprofloxacin because it partially overcomes the effect on affinity. Finally, comparative studies on DNA cleavage and decatenation suggest that the antibacterial properties of trovafloxacin result from increased S. aureus topoisomerase IV-mediated DNA cleavage rather than inhibition of enzyme catalysis. PMID- 10704225 TI - Factors determining vesicular lipid mixing induced by shortened constructs of influenza hemagglutinin. AB - The HA2 subunit of influenza hemagglutinin is responsible for fusion of the viral and host-cell membranes during infection. An N-terminal 127 amino acid construct of HA2, FHA2-127, is shown to induce lipid mixing of large unilamellar vesicles under endosomal low pH conditions. Thus, FHA2 could serve as a good model system for biophysical studies of membrane fusion. With FHA2, we began to develop a mechanistic model which could explain how this short construct facilitates membrane fusion. In this endeavor, we studied the possible role of the kinked loop region (amino acids 105-113). A construct missing this loop, FHA2-90, although able to induce lipid mixing, has lost the sharp pH-dependent transition seen with FHA2-127 and native HA. In addition, FHA2-127 promotes extensive vesicle aggregation more effectively than FHA2-90 upon acidification. These data suggest that the kinked loop may play a pH-dependent regulatory role. To test this, we compared bis-ANS binding to the two constructs and observed that binding to FHA2-127 increases at a faster rate than FHA2-90 as the pH is decreased, indicating that the kinked loop not only is an ANS-binding site, but that it binds better at low pH. The pH dependence of this transition directly correlates with that observed in lipid mixing. Further, cysteine mutations of acidic residues in the kinked region are both fusion inactive and bind much less ANS, whereas a similar mutation of a threonine residue had little effect on fusion activity or ANS binding. This evidence lends further support to our idea that the kinked loop serves a regulatory role. To test the physiological relevance of the FHA2-127 fusion mechanism, we studied the effects of a G1E mutation, known to abolish fusion in native HA. We found that G1E-127 is fusion inactive as expected. This evidence indirectly suggests that the mechanism of FHA2-127 is perhaps physiologically relevant and from its study, we can learn much about the mechanism of native HA. PMID- 10704226 TI - The relationship between the effect of lysine analogues and salt on the conformation of lipoprotein(a). AB - Lipoprotein(a) [Lp(a)] exhibits many of the same properties as plasminogen, owing to a similar structural makeup from a composite of multiple kringle domains. Shared behavior includes induction of an expanded conformation by lysine analogues, inhibition of this effect, and creation of a compact conformation by NaCl. Here, we examine in detail the independent and mutual effects of NaCl and 6 aminohexanoic acid (6-AHA) on the structure of Lp(a) and the relationship between the binding of the two ligands. We find that NaCl promotes the compact conformation while binding to Lp(a) homogeneously. In the absence of salt, 6-AHA leads to the complete unfolding of Lp(a), a process that is accompanied by cooperative binding. Reversal of conformation and weakening of binding occurred when one ligand was added to Lp(a) in the presence of the other, suggesting competitive binding. High concentrations of NaCl completely reversed the expansion of Lp(a) in 100 mM 6-AHA, and high concentrations of 6-AHA unfolded Lp(a) in the presence of 100 mM NaCl, but only by 30% in the case of the 15 kringle IV Lp(a) studied. Induction of the compact form of Lp(a) appears to be an effect in common with all salts examined and cannot be attributed solely to the anion, as in the case of plasminogen. The results were summarized in terms of a model of Lp(a) depicting the conformational alterations of apo(a) caused by the binding of the two ligands. In the compact conformation in NaCl, apo(a) is apposed to the particle surface. The fully expanded form in 6-AHA results from release of both the variable and constant kringle domains. In the intermediate form in water and in a solution containing both NaCl and 6-AHA, only the variable domain is released from the particle surface. PMID- 10704227 TI - Nitrogen monoxide activates iron regulatory protein 1 RNA-binding activity by two possible mechanisms: effect on the [4Fe-4S] cluster and iron mobilization from cells. AB - The iron-regulatory protein 1 (IRP1) regulates the expression of several molecules involved in iron (Fe) metabolism by reversibly binding to iron responsive elements (IREs) in the untranslated regions (UTR) of particular mRNA transcripts. Several studies have indicated that nitrogen monoxide (NO) may influence IRP1 RNA-binding activity by a direct effect on the [4Fe-4S] cluster of the protein. It has also been suggested that NO may act indirectly on IRP1 by affecting the intracellular Fe pools that regulate the function of this protein [Pantopoulous et al. (1996) Mol. Cell. Biol. 16, 3781-3788]. There is also the possibility that NO may S-nitrosate sulfhydryl groups that are crucial for mRNA binding and decrease IRP1 activity by this mechanism. We have examined the effect of a variety of NO donors [e.g., S-nitroso-N-acetylpenicillamine (SNAP), spermine NONOate (SperNO), and S-nitrosoglutathione (GSNO)] on IRP1 RNA-binding activity in both LMTK(-) fibroblast lysates and whole cells. In cell lysates, the effects of NO at increasing RNA-binding activity were only observed when cells were made Fe-replete. Under these circumstances, IRP1 contains an [4Fe-4S] cluster that was susceptible to NO. In contrast, when lysates were prepared from cells treated with the Fe chelator desferrioxamine (DFO), NO had no effect on the RNA-binding activity of IRP1. The lack of effect of NO under these conditions was probably because this protein does not have an [4Fe-4S] cluster. In contrast to the NO generators above, sodium nitroprusside (SNP) decreased IRP1 RNA binding when cells were incubated with this compound. However, SNP had no effect on IRP1 RNA binding activity in lysates, suggesting that the decrease after incubation of cells with SNP was not due to S-nitrosation of critical sulfhydryl groups. Apart from the direct effect of NO on IRP1 in Fe-replete cells, we have shown that NO generated by SNAP, SperNO, and GSNO could also mobilize Fe from cells. When NO generation was induced in RAW 264.7 macrophages, an increase in IRP1 RNA-binding activity occurred but there was only a small increase in Fe release. Our results suggest that NO could activate IRP1 RNA-binding by two possible mechanisms: (1) its direct effect on the [4Fe-4S] cluster and (2) mobilization of (59)Fe from cells resulting in Fe depletion, which then increases IRP1 RNA-binding activity. PMID- 10704228 TI - Backbone dynamics of Tet repressor alpha8intersectionalpha9 loop. AB - A set of single Trp mutants of class B Tet repressor (TetR), in which Trp residues are located from positions 159 to 167, has been engineered to investigate the dynamics of the loop joining the alpha-helices 8 and 9. The fluorescence anisotropy decay of most mutants can be described by the sum of three exponential components. The longest rotational correlation time, 30 ns at 10 degrees C, corresponds to the overall rotation of the protein. The shortest two components, on the subnanosecond and nanosecond time scale, are related to internal motions of the protein. The initial anisotropy, in the 0.16-0.22 range, indicates the existence of an additional ultrafast motion on the picosecond time scale. Examination of physical models for underlying motions indicates that librational motions of the Trp side chain within the rotameric chi(1) x chi(2) potential wells contribute to the picosecond depolarization process, whereas the subnanosecond and nanosecond depolarization processes are related to backbone dynamics. In the absence of inducer, the order parameters of these motions, about 0.90 and 0.80 for most positions, indicate limited flexibility of the loop backbone. Anhydrotetracycline binding to TetR induces an increased mobility of the loop on the nanosecond time scale. This suggests that entropic factors might play a role in the mechanism of allosteric transition. PMID- 10704229 TI - The process of amyloid-like fibril formation by methionine aminopeptidase from a hyperthermophile, Pyrococcus furiosus. AB - Amyloid is associated with serious diseases including Alzheimer's disease and senile-systemic amyloidosis due to misfolded proteins. In the course of study of the denaturation process of methionine aminopeptidase (MAP) from the hyperthermophile P. furiosus, we found that MAP forms amyloid-like fibrils, and we then investigated the mechanism of amyloid fibril formation. The kinetic experiments on denaturation monitored by CD at 222 nm indicated that MAP in the presence of 3.37 M GuHCl at pH 3.31 changed to a conformation containing a considerable content of beta-sheet structure after the destruction of the alpha helical structure. MAP in this beta-rich conformation was highly associated, and its stability was remarkably high: the midpoint of the GuHCl denaturation curve was 4.82 M at pH 3.0, and a thermal transition was not observed up to 125 degrees C by calorimetry. The amyloid-like fibril formation of MAP was confirmed by Congo red staining with a typical peak at 542 nm in the difference spectrum, showing a cross-beta X-ray diffraction pattern with a clear sharp reflection at 4.7 A and a characteristic unbranched fibrillar appearance with a length of about 1000 A and a diameter of about 70 A in the electron micrographs. Present results indicate that the amyloid-like form of MAP appears just after the protein is almost completely denatured, and even highly stable proteins can also form amyloid-like conformation under conditions where the denatured state of the protein is abundantly populated. PMID- 10704230 TI - Escherichia coli ATP synthase alpha subunit Arg-376: the catalytic site arginine does not participate in the hydrolysis/synthesis reaction but is required for promotion to the steady state. AB - The three catalytic sites of the F(O)F(1) ATP synthase interact through a cooperative mechanism that is required for the promotion of catalysis. Replacement of the conserved alpha subunit Arg-376 in the Escherichia coli F(1) catalytic site with Ala or Lys resulted in turnover rates of ATP hydrolysis that were 2 x 10(3)-fold lower than that of the wild type. Mutant enzymes catalyzed hydrolysis at a single site with kinetics similar to that of the wild type; however, addition of excess ATP did not chase bound ATP, ADP, or Pi from the catalytic site, indicating that binding of ATP to the second and third sites failed to promote release of products from the first site. Direct monitoring of nucleotide binding in the alphaR376A and alphaR376K mutant F(1) by a tryptophan in place of betaTyr-331 (Weber et al. (1993) J. Biol. Chem. 268, 20126-20133) showed that the catalytic sites of the mutant enzymes, like the wild type, have different affinities and therefore, are structurally asymmetric. These results indicate that alphaArg-376, which is close to the beta- or gamma-phosphate group of bound ADP or ATP, respectively, does not make a significant contribution to the catalytic reaction, but coordination of the arginine to nucleotide filling the low-affinity sites is essential for promotion of rotational catalysis to steady-state turnover. PMID- 10704231 TI - Activation of Zap-70 tyrosine kinase due to a structural rearrangement induced by tyrosine phosphorylation and/or ITAM binding. AB - The protein tyrosine kinase ZAP-70 is implicated in the early steps of the T-cell antigen receptor (TCR) signaling. Binding of ZAP-70 to the phosphorylated immunoreceptor tyrosine-based activation motifs (ITAMs) of the TCR zeta chain through its two src-homology 2 (SH2) domains results in its activation coupled to phosphorylation on multiple tyrosine residues, mediated by Src kinases including Lck as well as by autophosphorylation. The mechanism of ZAP-70 activation following receptor binding is still not completely understood. Here we investigated the effect of intramolecular interactions and autophosphorylation by following the kinetics of recombinant ZAP-70 activation in a spectrophotometric substrate phosphorylation assay. Under these conditions, we observed a lag phase of several minutes before full ZAP-70 activation, which was not observed using a truncated form lacking the first 254 residues, suggesting that it might be due to an intramolecular interaction involving the interdomain A and SH2 region. Accordingly, the lag phase could be reproduced by testing the truncated form in the presence of recombinant SH2 domains and was abolished by the addition of diphosphorylated ITAM peptide. Preincubation with ATP or phosphorylation by Lck also abolished the lag phase and resulted in a more active enzyme. The same results were obtained using a ZAP-70 mutant lacking the interdomain B tyrosines. These findings are consistent with a mechanism in which ZAP-70 phosphorylation/autophosphorylation on tyrosine(s) other than 292, 315, and 319, as well as engagement of the SH2 domains by the phosphorylated TCR, can induce a conformational change leading to accelerated enzyme kinetics and higher catalytic efficiency. PMID- 10704232 TI - Self-assembly of recombinant prion protein of 106 residues. AB - The central event in the pathogenesis of prion diseases is a profound conformational change of the prion protein (PrP) from an alpha-helical (PrP(C)) to a beta-sheet-rich isoform (PrP(Sc)). The elucidation of the mechanism of conformational transition has been complicated by the challenge of collecting high-resolution biophysical data on the relatively insoluble aggregation-prone PrP(Sc) isoform. In an attempt to facilitate the structural analysis of PrP(Sc), a redacted chimeric mouse-hamster PrP of 106 amino acids (MHM2 PrP106) with two deletions (Delta23-88 and Delta141-176) was expressed and purified from Escherichia coli. PrP106 retains the ability to support PrP(Sc) formation in transgenic mice, implying that it contains all regions of PrP that are necessary for the conformational transition into the pathogenic isoform [Supattapone, S., et al. (1999) Cell 96, 869-878]. Unstructured at low concentrations, recombinant unglycosylated PrP106 (rPrP106) undergoes a concentration-dependent conformational transition to a beta-sheet-rich form. Following the conformational transition, rPrP106 possesses properties similar to those of PrP(Sc)106, such as high beta-sheet content, defined tertiary structure, resistance to limited digestion by proteinase K, and high thermodynamic stability. In GdnHCl-induced denaturation studies, a single cooperative conformational transition between the unstructured monomer and the assembled beta-oligomer was observed. After proteinase K digestion, the oligomers retain an intact core with unusually high beta-sheet content (>80%). Using mass spectrometry, we discovered that the region of residues 134-215 of rPrP106 is protected from proteinase K digestion and possesses a solvent-independent propensity to adopt a beta-sheet-rich conformation. In contrast to the PrP(Sc)106 purified from the brains of neurologically impaired animals, multimeric beta-rPrP106 remains soluble, providing opportunities for detailed structural studies. PMID- 10704233 TI - Inhibitors of kinesin activity from structure-based computer screening. AB - Kinesin motor proteins use ATP hydrolysis for transport along microtubules in the cell. We sought to identify small organic ligands to inhibit kinesin's activity. Candidate molecules were identified by computational docking of commercially available compounds using the computer program DOCK. Compounds were docked at either of two sites, and a selection was tested for inhibition of microtubule stimulated ATPase activity. Twenty-two submillimolar inhibitors were identified. Several inhibitors appeared to be competitive for microtubule binding and not for ATP binding, and three compounds showed 50% inhibition down to single-digit micromolar levels. Most inhibitors grouped into four distinct classes (fluoresceins, phenolphthaleins, anthraquinones, and naphthylene sulfonates). We measured the binding of one inhibitor, rose bengal lactone (RBL), to kinesin (dissociation constant 2.5 microM) by its increase in steady-state fluorescence anisotropy. The RBL binding site on kinesin was localized by fluorescent resonance energy transfer (FRET) using a donor fluorophore (coumarin) covalently attached at unique, surface-exposed cysteine residues engineered at positions 28, 149, 103, 220, or 330. RBL was found to bind in its original docked site: the pocket cradled by loop 8 and beta-strand 5 in kinesin's three-dimensional structure. These results confirm this region's role in microtubule binding and identify this pocket as a novel binding site for kinesin inhibition. PMID- 10704235 TI - FK506 binding protein from a thermophilic archaeon, methanococcus thermolithotrophicus, has chaperone-like activity in vitro PMID- 10704234 TI - Fusion of beta 2-adrenergic receptor to G alpha s in mammalian cells: identification of a specific signal transduction species not characteristic of constitutive activation or precoupling. AB - The forward and antegrade interactions that comprise the agonist receptor-G protein complex were studied in Chinese hamster fibroblasts transfected to express the beta(2)-adrenergic receptor (beta(2)AR), the beta(2)AR and the alpha subunit of its cognate G protein (G(s)), and a protein consisting of the beta(2)AR fused at its carboxy terminus with G(alpha)(s) (beta(2)AR-G(s)). Expression levels were matched at approximately 600 fmol/mg. Basal adenylyl cyclase activities were increased with the fusion receptor membranes compared to coexpressed receptor plus G(alpha)(s), and to wild-type beta(2)AR (20.5 +/- 1.8 vs 9.0 +/- 0.88 vs 8.7 +/- 0.93 pmol min(-)(1) mg(-)(1)), confirming in mammalian cells that the fusion of beta(2)AR and G(alpha)(s) results in a state not attained by expression of unfused components. However, agonist-stimulated activities were not increased proportionally, such that the stimulation over basal of the beta(2)AR-G(s) fusion protein (1. 5-fold) was less than wild-type beta(2)AR (2.1-fold). Agonist competition studies performed in the absence of guanine nucleotide exhibited high-affinity binding sites with a lower K(H) (1.75 vs 8. 47 nM) and greater %R(H) (51% vs 44%) for beta(2)AR-G(s), but GppNHp failed to convert most of these to the low-affinity state. Functional studies with the inverse agonist ICI 118551 did not show enhanced efficacy or potency with the fusion protein. Adenylyl cyclase studies with three partial agonists with diverse structures (dobutamine, ritodrine, and phenylephrine) showed no enhancement of efficacy with beta(2)AR-G(s) and a minor trend toward enhanced potency. Taken together, these results indicate that the tethering of G(alpha)(s) to the beta(2)AR causes a conformational change in the receptor that stabilizes a species "trapped" between the non-guanine nucleotide-bound state and the GTP bound form. Functionally the receptor is not characterized by a consistent pattern of properties ascribed to other states such as constitutive activation or precoupling, but rather represents a unique state in the transition from high- to low-affinity forms. PMID- 10704236 TI - The unitary evoked potential at the frog nerve-muscle junction results from synchronous gating of fusion pores at docked vesicles. AB - Exocytosis of a single vesicle has been proposed as the mechanism which determines quantal size by releasing a prepackaged and standard amount of acetylcholine. As first described by del Castillo and Katz (1954) the endplate potential is composed of 100 unitary events and the small variance suggests a binomial release from 100 "discrete patches of membrane". However, exocytosis of 100 vesicles selected randomly from 5000 docked vesicles would yield a variance that is 7 times greater than observed values. We propose that the presynaptic ridge with its compliment of docked vesicles functions as the "discrete patch of membrane" such that arrays of calcium activated fusion pores meter transmitter to form the unit of release. A model based on the synchronous flicker of a large number of fusion pores produces the small variance of both miniature end plate potentials and unitary end plate potentials. Release from a single locus (fusion pore) would generate the sub-MEPP. This model permits vesicle trafficking and vesicular content depletion during tetanic stimulation and explains the frequency dependency of MEPP amplitudes and changes in sub-MEPP to bell-MEPP class ratios. PMID- 10704237 TI - Transient apoptosis elicited by insulin in serum-starved glioma cells involves Fas/Fas-L and Bcl-2. AB - The expression of fas gene in glioma cells varies with growth stage. When insulin elicited transient apoptosis of glioma cells was in progress, the expression of fas gene increased at both transcriptional and translational levels. In contrast, the expression of fas-L gene in glioma cells remained constant. Apoptosis occurred in the cells having high level of surface Fas protein. When the expression of Fas-L in U-373MG cells was suppressed by ribozyme, the insulin elicited transient apoptosis vanished. Overexpression of Bcl-2 in U-373MG cells did not alter significantly the cell cycle progression and the expression of fas gene. However, these cells were resistant to insulin-trigged death. Therefore, insulin-elicited apoptosis involved Fas-related death signal, and which could be prevented by the protective effect of Bcl-2. PMID- 10704238 TI - Apoptosis during iron chelator-induced differentiation in F9 embryonal carcinoma cells. AB - We have previously demonstrated that three potent iron chelators, hinokitiol, dithizone and deferoxamine, induce differentiation of F9 embryonal carcinoma cells, as do other well-known morphogens such as retinoic acid (RA) and sodium butyrate (NaB). In this study, we compared the patterns of cell proliferation, cell death and cell cycle arrest during the process of differentiation induced by these five agents. When F9 cells were cultured with the agents at their individual differentiation-inducing concentrations, cell proliferation was rapidly inhibited by treatment with the iron chelators and NaB. In contrast, RA did not influence the rate of increase of cell number at the concentration of 1 microm. The three chelators also caused a marked reduction in cell viability, and the treated cells exhibited internucleosomal DNA fragmentation, whereas cells treated with NaB showed no apoptotic characteristics. RA induced apoptosis weakly at 1 microm and strongly at higher concentrations. In addition, all the iron chelators hindered cell cycle progression, resulting in an arrest at the G1-S interface or S phase. The phenomena observed in chelator-treated cells were considerably different from those in RA- or NaB-treated cells. It is concluded that the three iron chelators cause both severe apoptotic cell death and cell cycle arrest of proliferating F9 cells via cellular iron deprivation, and that this apoptotic change may be independent of the process of differentiation. PMID- 10704239 TI - Cloning and characterization of the gene encoding the highly expressed ribosomal protein l3 of the ciliated protozoan Tetrahymena thermophila. Evidence for differential codon usage in highly expressed genes. AB - We have cloned and characterized the cDNA and the macronuclear genomic copy of the highly conserved ribosomal protein (r-protein) L3 of Tetrahymena thermophila. The r-protein L3 is encoded by a single copy gene interrupted by one intron. The organization of the promoter region exhibits features characteristic of ribosomal protein genes in Tetrahymena. The codon usage of the L3 gene is highly biased. A thorough analysis of codon usage in Tetrahymena genes revealed that genes could be categorized into two classes according to codon usage bias. Class A comprises r-protein genes and a number of other highly expressed genes. Class B comprises weakly expressed genes such as the conjugation induced CnjB and CnjC genes, but surprisingly, this class also contains abundantly expressed genes such as the genes encoding the surface antigens SerH3 and SerH1. Codon usage is slightly more restricted in class A than in class B, but both classes exhibit distinct and different codon usage biases. Class A genes preferentially use C and U in the silent third codon positions, whereas class B genes preferentially use A and U in the silent third codon positions. The analysis suggests that two different strategies have been employed for optimization of codon usage in the A+T-rich genome of Tetrahymena. PMID- 10704240 TI - Microfilament-dependent modulation of cytoplasmic protein binding to TNFalpha mRNA AU-rich instability element in human lymphoid cells. AB - Cytoplasmic proteins with binding capability to AU-rich instability determinant sequences (ARE) of tumour necrosis factor alpha (TNFalpha) mRNA 3' untranslated region (3'UTR) were assessed in human lymphoid cells. In vitro label transfer experiments using wild type as well as mutant sequences in which the 70 nucleotide-long AUUUA pentamer-containing portion of the 3'UTR had been deleted conferred binding specificity to five major activities of 22/25-, 38/40-, 50-, 60 and 80-kDa proteins in cytoplasmic extracts of peripheral blood mononuclear cells (PBMCs). Cytochalasin-induced disarrangement of the F-actin-based microfilament system led to a Triton X-100-insoluble to soluble redistribution of these binding activities. No such changes were observed in Jurkat tumour cells. Combination of in vivo UV-crosslinking and in vitro label transfer experiments revealed considerable differences in RNA association between proteins of the same cell type as well as between proteins of identical molecular weight (Mw) derived from either PBMCs or Jurkat cells. Our findings may explain some aspects of differential regulation of interleukin 2 (IL-2) and TNFalpha mRNA stability upon microfilament disruption in human PBMCs observed in an earlier study. These results also suggest that the physical state of cytoplasmic structural environment might contribute to important regulatory processes regarding key elements of eukaryotic mRNA metabolism, such as modulation of stability. Finally, these data highlight the possibility that the often observed disorganization of the cytoskeleton in tumour cells may partly be responsible for the maintenance of the neoplastic state, a phenomenon that potentially involves ARE-AUBP interactions. PMID- 10704241 TI - Cell fractionation analysis of human CD8 glycoprotein transport between endoplasmic reticulum, intermediate compartment and Golgi complex in tissue cultured cells. AB - We have set up an analytical cell fractionation procedure to dissect, by a non morphological method, the anterograde transport of proteins from endoplasmic reticulum, intermediate compartment and Golgi complex in tissue cultured cells. Using this procedure after pulse-chase labelling of cells expressing human CD8 glycoprotein, we obtained results that: (1) support the view that the intermediate compartment is a distinct station in the export from the endoplasmic reticulum to the Golgi complex; and (2) strongly suggests that the O glycosylation process starts after the intermediate compartment, presumably in the cis -Golgi complex. PMID- 10704242 TI - S-adenosylmethionine synthesis in Leishmania infantum promastigotes. AB - Methionine adenosyltransferase (MAT), S -adenosylmethionine (AdoMet), and S adenosylhomocysteine (AdoHcy), have been analysed at different time-points during the growth curve of Leishmania infantum. MAT activity and AdoMet content peaked in the lag and early log phases, whereas higher levels of AdoHcy were found in stationary phase cells. MAT activity of cell extracts displayed hyperbolic kinetics for both its substrates, l -methionine and ATP, with km values of 35 microm and 5 m m, respectively. MAT has an absolute requirement for divalent cations, and is dependent on sulfydryl protective agents. Unlike other sources, L. infantum MAT activity seems to be transcriptionally regulated, with an accumulation of MAT-mRNA during rapid growth periods of promastigotes. PMID- 10704243 TI - Cytotoxicity by stored human breast-milk: possible contribution of complement system. AB - Human milk stored over some period in vitro possesses certain cytotoxic properties, which require further studies. Cytolysis induced by stored human milk has now been further characterized, using rabbit red blood cells as targets, to determine the contribution of other components, particularly the complement system. Cytolysis was found to be temperature dependent, greatly enhanced by low concentrations of magnesium and calcium ions, but inhibited by moderate to excessive amounts of calcium ions, and by heating at 56 degrees C. PMID- 10704244 TI - Novel chicken CXC and CC chemokines. AB - Upon stimulation with lipopolysaccharide (LPS) the chicken macrophage cell line HD-11 secretes factors with cytokine activity. To characterize these molecules, representational difference analysis with RNA of LPS-induced and uninduced HD-11 cells was performed. Two cDNA clones were isolated that code for polypeptides with structural features of chemokines. cDNA K60 codes for a novel CXC chemokine of 104 residues including a putative signal peptide of 20 amino acids at the N terminus. It is 67% identical to the previously cloned chicken chemokine 9E3/CEF4. K60 exhibits a similar degree of sequence identity to human interleukin 8 and other related CXC chemokines (about 50%), rendering straight-forward predictions of its biological properties difficult. cDNA K203 codes for a novel CC chemokine of 89 amino acids including a putative N-terminal signal peptide of 21 residues. It is 43% identical to a previously characterized chicken protein with homology to mammalian macrophage inflammatory protein 1beta (MIP-1beta). K203 exhibits about 50% sequence identity to human MIP-1beta and other related CC chemokines. PMID- 10704245 TI - Determinants of the functional interaction between the soluble GM-CSF receptor and the GM-CSF receptor beta-subunit. AB - The GM-CSF receptor consists of a GM-CSF specific low affinity alpha-subunit (GMRalpha) and a beta-subunit (betac) that associates with GMRalpha in the presence of GM-CSF to form a high-affinity complex. A splice variant soluble isoform of GMRalpha (solalpha) consists of the extracellular domain of GMRalpha and a unique 16-amino acid C-terminal domain. Exogenously administered solalpha is unable to associate with betac on the cell surface either in the presence or absence of GM-CSF. However, paradoxically, co-expression of solalpha with betac results in the ligand-independent association of solalpha with betac on the cell surface via the C-terminal domain of solalpha. To study the interaction and functional characteristics of the solalpha-betac complex we engineered a soluble betac-subunit (ECDbeta) and expressed it alone and with solalpha. Co-expressed but not independent sources of solalpha and ECDbeta could be co-precipitated in the absence of ligand demonstrating the extracellular domain of betac was sufficient for association with solalpha upon co-expression. However, independent sources of solalpha could associate with ECDbeta in the presence of GM-CSF as could a C-terminal deficient solalpha mutant (ECDalpha) and the addition of ECDbeta to ECDalpha and GM-CSF was associated with a conversion from a low- to high-affinity ligand-receptor complex. PMID- 10704246 TI - Correlation between DNA methylation and murine IFN-gamma and IL-4 expression. AB - In order to determine the possible role of DNA methylation as a regulatory mechanism for the restricted pattern of lymphokine production among differentiated Th(1)and Th(2)cells, we examined the extent of methylation of the interferon gamma (IFN-gamma) and the interleukin 4 (IL-4) genes in fresh activated murine Th(0), Th(1)and Th(2)cells, unstimulated naive T cells, B cells, bone marrow derived non-B non-T cells, thymocytes and liver. All of the CpG dinucleotides examined in the IL-4 and the IFN-gamma genes, were fully methylated over the body of the gene in all of the examined cells. However, analysis of the promoter regions of these genes revealed a different pattern. While the IL-4 promoter is fully methylated in all of the examined cells, two adjacent CpG dinucleotides near the initiation point of the IFN-gamma gene were unmethylated in all T cells, including 17-day-old fetal thymocytes. In contrast, B cells, bone marrow non-B non-T cells and liver cells displayed a full methylated profile of the IFNgamma promoter. These results suggest that the mutually exclusive pattern of IFNgamma and IL-4 production in Th(1)and Th(2)cells is not regulated by differential demethylation of these two genes. PMID- 10704247 TI - Differential induction of JE/MCP-1 in subclones from a murine macrophage cell line, RAW 264.7: role of kappaB-3 binding protein. AB - The JE/MCP-1 gene is an immediate-early gene, and its product is a CC chemokine that attracts monocytes, basophils and T lymphocytes. JE/MCP-1 gene expression is induced by various inflammatory stimuli, but its transcriptional mechanism is not fully understood. To address this question, we obtained two subclones from a parental RAW264.7 cell line, one subline with low JE/MCP-1-producing capacity (named RAW.c11) and the other with high JE/MCP-1-producing capacity (named RAW.c25), in response to lipopolysaccharide (LPS). These subclones have no significant differences in CD14 expression, nitric oxide production, or production of other cytokines, including TNF-alpha or IL-1alpha/beta. In electrophoretic mobility shift assays (EMSA), there were no significant differences in DNA binding to the NF-kappaB-consensus sequence and interferon regulatory factor (IRF)-1,2 binding sequences. However, significantly higher binding activity to the NF-kappaB-like sequence (kappaB-3), which is located in the promoter region of the JE/MCP-1 gene, was shown by a high producer subclone than by a low producer subclone. Transient transfection analysis using deletion mutants of a 0.5-kb region from -467 to +59 identified an LPS-responsive region in a kappaB-3 site (from -169 to -132) in the high producer subclone. Mutation of this site markedly reduced sensitivity to LPS in the high producer subclone. These data suggest that a yet undefined nuclear factor may be involved in differential JE/MCP-1 gene transcription. PMID- 10704248 TI - Divergent regulation of the murine CC chemokine C10 by Th(1) and Th(2) cytokines. AB - Chemokines are typically found as products of acute stimulation of host defence cells. In contrast, the mouse CC chemokine C10 was previously shown to be a delayed, stably induced product of macrophages treated with interleukin 3 (IL-3), IL-4 or GM-CSF. We investigated the possibility that C10 is differentially regulated by cytokines associated with Th(1)and Th(2)cells. Northern blot analysis of bone marrow-derived macrophages showed that, in addition to IL-4, the Th(2)-specific cytokines IL-10 and IL-13 upregulated C10 over a 48-h period in a dose-dependent manner. In contrast, MIP-1alpha and MCP-1/JE were induced by IL-3 or GM-CSF at 48 h and this induction was inhibited by IL-4. Interferon gamma, a Th(1)-specific product, abolished the induction of C10 mRNA and protein by either IL-3 or granulocyte-macrophage colony-stimulating factor (GM-CSF) in either bone marrow-derived or peritoneal macrophages. The inhibition of C10 production by interferon gamma was not NO dependent. Finally the GM-CSF-mediated induction of C10 in peritoneal macrophages was eliminated when these cells presented antigen to established T cells of Th(1)phenotype. The findings are consistent with a potential role for C10 in the modulation of immune reactions of Th(2)type. PMID- 10704249 TI - Activation of neutral sphingomyelinase by IL-1beta requires the type 1 interleukin 1 receptor. AB - The cytokine interleukin 1beta (IL-1beta) plays an important role in host defence reactions and neuro-immune interactions but it is still not clear which of the two interleukin 1 receptor subtypes is coupled to activation of neutral sphingomyelinase (nSMase) by IL-1beta. To investigate involvement of neutral sphingomyelinase (nSMase) in central IL-1beta effects we used P(2)fractions of brain cerebral cortex from wild-type mice and mice deficient in the type 1 IL-1 receptor. IL-1beta (human, recombinant) was shown to activate, in a dose dependent manner, nSMase in the P(2)brain fraction of the wild-type mice while in the knock-out mice the stimulatory effect of IL-1beta on nSMase was absent. In the presence of an IL-1 receptor antagonist (IL-1ra), IL-1beta did not activate nSMase either in the cortex of wild-type or knock-out mice. These data suggest that nSMase, a key enzyme of the sphingomyelin signal transduction pathway, might be involved in IL-1beta signalling in the brain and that activation of the enzyme requires the IL-1 receptor type 1. PMID- 10704250 TI - The interferon-induced gene ISG12 is regulated by various cytokines as the gene 6 16 in human cell lines. AB - The gene for ISG12 (originally designated p27) was isolated as an oestrogen induced gene. The authors undertook a comprehensive study using quantitative RT PCR, in which we delineate the regulation of ISG12 by seven different cytokines including interferons and poly(I). poly(C) in seven human cell lines of different origin. In all cell lines ISG12 is strongly induced by IFN-alpha and only slightly by IFNgamma. Poly(I).poly(C) induces ISG12 in a cell line-dependent manner, whereas none of the other cytokines tested elicited a response. Comparing the induction pattern of ISG12 to that of 6-16 a high degree of similarity was found. The induction levels varied, however, between cell lines. PMID- 10704251 TI - Sesquiterpene lactones are potent inhibitors of interleukin 8 gene expression in cultured human respiratory epithelium. AB - Sesquiterpene lactones, derived from Mexican-Indian medicinal plants, are known to have potent anti-inflammatory properties but the mechanisms of this effect are not completely understood. Recent data demonstrated that sesquiterpene lactones were potent inhibitors of the pro-inflammatory transcription factor NF-kappaB. Because activation of NF-kappaB is involved in the regulation of the chemokine interleukin 8 (IL-8), we hypothesized that the sesquiterpene lactones, isohelenin and parthenolide, would inhibit IL-8 gene expression in cultured human respiratory epithelium. Incubating A549 cells with tumour necrosis factor alpha (TNF-alpha) induced IL-8 mRNA expression and secretion of immunoreactive IL-8. Pretreatment with either isohelenin or parthenolide inhibited TNF-alpha-mediated IL-8 gene expression in a concentration-dependent manner. Pretreatment with either compound inhibited TNF-alpha mediated activation of the IL-8 promoter and TNF-alpha-mediated nuclear translocation of NF-kappaB. In addition, pretreatment with isohelenin or parthenolide inhibited TNF-alpha-mediated degradation of the NF-kappaB inhibitory protein, I-kappaBalpha. We conclude that sesquiterpene lactones are potent in vitro inhibitors of IL-8 gene expression in cultured human respiratory epithelium. The most proximal mechanism of inhibition appears to involve inhibition of I-kappaBalpha degradation. Stabilization of cytoplasmic I kappaBalpha leads to inhibition of NF-kappaB nuclear translocation and of subsequent IL-8 promoter activation. The ability of sesquiterpene lactones to modulate IL-8 gene expression may explain, in part, their anti-inflammatory effects. PMID- 10704252 TI - High altitude increases circulating interleukin-6, interleukin-1 receptor antagonist and C-reactive protein. AB - Hypoxic pulmonary vasoconstriction is associated with but may not be sufficient for the development of high-altitude pulmonary oedema (HAPO). Hypoxia is known to induce an inflammatory response in immune cells and endothelial cells. It has been speculated that hypoxia-induced inflammatory cytokines at high altitude may contribute to the development of HAPO by causing capillary leakage in the lung. We were interested if such an inflammatory response, possibly involved in a later development of HAPO, is detectable at high altitude in individuals without HAPO. We examined the plasma levels of interleukin 6 (IL-6), interleukin 1 receptor antagonist (IL-1ra) and C-reactive protein (CRP) in two independent studies: study A, Jungfraujoch, Switzerland, three overnight stays at 3458 m, n=12; study B: Capanna Regina Margherita, Italy, 3 overnight stays at 3647 m and one overnight stay at 4559 m, n=10. In both studies, probands showed symptoms of acute mountain sickness but no signs of HAPO. At the Jungfraujoch, IL-6 increased from 0.1+/-0.03 pg/ml to 2. 0+/-0.5 pg/ml (day 2, P=0.03), IL-1ra from 101+/-21 to 284+/-73 pg/ml (day 2, P=0.01), and CRP from 1.0+/-0.4 to 5.8+/-1.5 micrograms/ml (day 4, P=0.01). At the Capanna Margherita, IL-6 increased from 0. 5+/-0.2 pg/ml to 2.0+/-0.8 pg/ml (P=0.02), IL-1ra from 118+/-25 to 213+/-28 pg/ml (P=0.02), and CRP from 0.4+/-0.03 to 3.5+/-1.1 micrograms/ml (P=0.03). IL-8 was below the detection limit of the ELISA (<25 pg/ml) in both studies. The increase of IL-6 and IL-1ra in response to high altitude was delayed and preceded the increase of CRP. We conclude that: (1) circulating IL-6, IL-1ra and CRP are upregulated in response to hypobaric hypoxic conditions at high altitude, and (2) the moderate systemic increase of these inflammatory markers may reflect considerable local inflammation. The existence and the kinetics of high altitude induced cytokines found in this study support the hypothesis that inflammation is involved in the development of HAPO. PMID- 10704253 TI - Local regulation of immune responses: corneal endothelial cells alter t cell activation and cytokine production. AB - Corneal endothelial cells (CE cells) inhibit antigen- and mitogen-activated lymphocyte proliferation assays, although interleukin 2 receptor (IL-2R) expression and responsiveness to exogenous IL-2 are unaffected. To examine this activity further, co-cultures of CE cells and T cell clones were studied. CE cells inhibited IL-2 and IL-4 production by T cells stimulated with Ag and APC, but not IL-5 or IL-6 production. CE cells also inhibited NFAT-driven lacZ reporter gene production following Ag stimulation of transfected KZO T hybridoma cells. Conversely, stimulation of IL-2 production by ionomycin, with or without PMA, was unaffected by the CE cells. Preincubation of KZO hybridoma or Jurkat cells with CE cells, or CE cell-conditioned culture supernatant, inhibited the intracellular calcium ([Ca(2+)](i)) increase induced by TCR ligation, but not the [Ca(2+)](i)increase induced by ionomycin or thapsigargin. The inhibitory effect was independent of APC and did not act by blocking costimulation, since IL-2 production stimulated by immobilized anti-CD3 alone was also inhibited by CE cells. The supernatant factor was heat labile. This novel activity is unlike other immunoregulatory molecules, including transforming growth factor beta (TGF beta) and may contribute to local immune privilege. PMID- 10704254 TI - Renal synthesis of leukaemia inhibitory factor (LIF), under normal and inflammatory conditions. AB - Leukaemia inhibitory factor (LIF) is a pleiotropic cytokine that is particularly involved in nephrogenesis and repair of the extracellular matrix. Transgenic mice overexpressing LIF have mesangial proliferative glomerulonephritis. Also, during local inflammatory reactions, such as kidney graft rejection or urinary tract infections, urinary LIF excretion is enhanced. The aim of the study therefore was to study LIF production by normal and inflammatory diseased kidneys (glomerulonephritis or graft rejection), maintained in short cultures. To determine the responsibility of the kidney itself in LIF synthesis, we measured LIF secretion into the culture supernatants of human mesangial or renal tubular epithelial cells. Fragments from diseased kidneys, whether grafts or not, released more LIF than normal human kidney fragments, mesangial or renal tubular epithelial cells. However, LIF production was delayed in renal transplants compared to glomerulonephritic samples taken from untreated patients. In every case, LIF production was enhanced by interleukin 1beta (IL-1beta) and inhibited by IL-4 or dexamethasone, except in two severe rejection episodes. So, LIF appeared to respond to pro- and anti-inflammatory stimuli, in vitro and in vivo. Considering its biological effects, LIF could play a role in inflammatory renal diseases. PMID- 10704255 TI - Stem cell factor (SCF) can regulate the activation and expansion of murine intraepithelial lymphocytes. AB - Murine intraepithelial lymphocytes (IEL) express c-kit, the receptor for stem cell factor (SCF). SCF induced a low but significant proliferative response in IEL, but not in splenic T cells. SCF stimulation of IEL resulted in an expansion of the c-kit(+), TCRgammadelta(+)cell population. SCF-induced proliferation was dependent upon SCF-c-kit interactions, since antibody to c-kit blocked this response, and IEL obtained from c-kit mutant (W/W(v)) mice failed to respond to SCF. SCF acted synergistically with anti-TCRgammadelta and with concavalin A (Con A) to induce proliferation and interferon gamma (IFN-gamma) production in IEL. Finally, mice injected with SCF had a significant increase in the number of IEL in the small intestine. SCF-treated mice had increased numbers of TCRalphabeta(+)and TCRgammadelta(+)cell populations, as well as increased numbers of c-kit(+)and c-kit(-)IEL. These data suggest that SCF-c-kit interactions play an important role in regulating IEL expansion and activation. PMID- 10704256 TI - Tumour necrosis factor alpha (TNF-alpha) interferes with Fas-mediated apoptotic cell death on rheumatoid arthritis (RA) synovial cells: a possible mechanism of rheumatoid synovial hyperplasia and a clinical benefit of anti-TNF-alpha therapy for RA. AB - To investigate the mechanism of rheumatoid synovial hyperplasia (RASH), the influence of tumour necrosis factor alpha (TNF-alpha) on Fas-mediated apoptotic cell death (Fas-ACD) was examined on cultured rheumatoid synovial cells (RASCs). RASCs were obtained from the synovial tissues of eight patients with rheumatoid arthritis (RA) and SCs from eight patients with osteoarthritis (OA) were used as a control. To examine the influence of TNF-alpha on Fas-ACD, SCs were cultured with anti-Fas antibody (CH11) for 16 h in the absence or presence of different doses of recombinant TNF-alpha. ACD was determined by electron microscopic analysis and the percentage of apoptotic cells was calculated by trypan blue staining. In addition, the expression of Fas and Bcl-2 on RASCs was examined by flow cytometry. As a result, RASCs were more susceptible to Fas-ACD in vitro than OASCs. TNF-alpha interfered with Fas-ACD on RASCs in a dose-dependent manner. Moreover, removal of TNF-alpha activity by a neutralizing anti-TNF-alpha antibody (cA2) restored Fas-ACD. Flow cytometric analysis showed no significant changes in either Fas or Bcl-2 expression on RASCs after the culture with TNFalpha. These results suggest the following: (1) Fas-ACD might be diminished in vivo by local excessive TNF-alpha and this might contribute in part to RASH. (2) The inhibition of Fas-ACD on RASCs by TNF-alpha might not be associated with changes in the expression of Fas or Bcl-2. (3) In addition, considering a magnetic resonance imaging (MRI) finding of marked reduction in the RASH after cA2 treatment, blockade of TNF-alpha activity could restore Fas-ACD in RA synovium, implicating a clinical benefit of anti-TNF-alpha therapy for RA. PMID- 10704257 TI - Retinoic acid suppresses interleukin 6 production in normal human osteoblasts. AB - Systemic long-term retinoid therapy for chronic skin diseases significantly reduced bone turnover markers within days and led to bone abnormalities. Retinoic acid (RA) plays a key role in the regulation of mouse bone cell proliferation, differentiation and functions. Meanwhile, there is little information of RA effect on human osteoblast and osteoclast cell development and function. Interleukin 6 (IL-6) is a pleiotropic cytokine with profound effects on bone metabolism. Thus, the present study examined the RA effect on cell differentiation, alkaline phosphatase and osteocalcin production as well as IL-6 production in normal human osteoblasts. The number of large differentiated osteoblast cells decreased in RA-treated cultures P<0.05. The production of bone specific markers, alkaline phosphatase and osteocalcin, was also reduced in RA treated cultures. Normal human osteoblasts produced 31.0+/-4.8 pg IL-6 per ml in control cultures. Within 24 h, RA at all four concentrations reduced Il-6 production from normal human osteoblasts. The pharmacological concentration of 10(-5) M RA suppressed 90% of IL-6 production. The present study shows for the first time that RA profoundly inhibits IL-6 production in normal human osteoblasts within 24 h and in a dose-dependent manner. RA was shown previously to inhibit IL-6 production in several other normal and malignant human cell types. The associated decrease in osteoblast cell differentiation, alkaline phosphatase and osteocalcin production could result from the rapid RA-inhibition of IL-6 production. Thus, RA inhibition of IL-6 production in normal human osteoblasts may contribute to the bone abnormalities seen after systemic long term retinoid therapy in some patients. PMID- 10704258 TI - Amlodipine inhibits the production of cytokines induced by ouabain. AB - Recent studies have suggested that cytokines are capable of modifying cardiovascular function and that drugs used in the treatment of heart failure have various modulating properties on the production of cytokines. More recently, we have found that ouabain induces the production of cytokines. This study was performed to examine the effects of calcium channel blockers on the production of cytokines induced by a cardiac glycoside. Human peripheral blood mononuclear cells (PBMC) were obtained from healthy volunteers. PBMC were cultured in 0.1, 1, 10, and 30 micromol/l amlodipine, diltiazem, and nifedipine in presence of 1 micromol/l ouabain. After 24 h of incubation, IL-1alpha, IL-1beta, IL-6, and TNF alpha were measured in the culture supernatants by enzyme-linked immunosorbent assay. Ouabain induced the production of IL-1alpha, IL-1beta and IL-6, but not of TNF-alpha. Induction of IL-1beta was most prominent. The production of IL-1alpha, and IL-6 was inhibited by amlodipine in a concentration-dependent manner and was significantly decreased at a concentration of 10 micromol/l. IL-1beta production was also inhibited by 30 micromol/l amlodipine. In contrast, neither diltiazem nor nifedipine inhibited the production of these cytokines. The unique property of amlodipine to inhibit the production of IL-1alpha, IL-1beta and IL-6 may contribute to its beneficial effects in heart failure patients. PMID- 10704259 TI - TANAX(T-61): an overview. AB - In this overview the authors describe the use of Tanax (T-61) for euthanasia. Tanax is a solution with three components (embutramide, mebenzonium iodide and tetracaine hydrochloride) used for painless death in pets and laboratory animals. It is also used for malicious intoxication in animals and for suicide attempts in humans. After a description of the modality and outcome of intoxication, the authors report the secondary toxic effects evoked by N, N -dimethyl-formamide, the solvent employed to dissolve the three components of Tanax. Finally, the analytical methods used to identify Tanax components in biological fluids and tissues are described. PMID- 10704260 TI - New in vitro approaches to explore cellular and molecular events related to carcinogenesis. PMID- 10704261 TI - Effect of D-002 on acetic acid-induced colitis in rats at single and repeated doses. AB - D-002 is a natural mixture of higher aliphatic primary alcohols purified from beeswax, with mild anti-inflammatory and effective antiulcer effects experimentally proved. Furthermore, it reduces leukotriene (LTB(4)) in the exudate of carrageenan-induced pleurisy and has a protective effect on the pre ulcerative phase of carrageenan-induced colonic ulceration in the guinea pig. This study was conducted to determine the effect of D-002 on acetic acid-induced colitis in rats at single and repeated doses. In a first series, D-002 was orally administered at 25 and 50 mg kg(-1), 24 h before the induction of colitis, meanwhile, in a second series, it was administered 24 h after the induction of colitis. Two other series (III and IV) examined the protective and therapeutic effect of D-002 administered for 7 days at the same doses, before or after colitis induction. Significant reductions in wet weight, macroscopic injury, polymorphonuclear infiltration and wall thickness were observed in colonic mucosa of D-002-treated animals compared with controls in both protective and therapeutic alternatives. It is concluded that D-002 was effective to protect or prevent the damage associated to acetic acid-induced colitis. PMID- 10704262 TI - Actions of Mn(2+) in the presence of ionophore A23187 in taenia coli of guinea pig. AB - In this work, we have investigated whether Mn(2+)enters the cytoplasm in the presence of ionophore for divalent cations, A23187 and induces contraction of taenia coli. In Ca(2+)-free, 60 m m K(+)medium, the application of 5 m m Mn(2+)evoked contraction and a concomitant increase in Mn(2+)influx into the cytoplasm. However, in Ca(2+)-free medium, the application of 5 m m Mn(2+)in the presence of 1x10(-5)m A23187 did not evoke both contraction and Mn(2+)influx. These results suggest that Mn(2+)penetrates through Ca(2+)channels in a state of membrane depolarization with K(+)and activates the contractile proteins in taenia coli. However, Mn(2+)does not enter the cytoplasm in the presence of ionophore A23187 and then does not induce contraction. PMID- 10704263 TI - Protective effects of vitamin c against cisplatin-induced nephrotoxicity and lipid peroxidation in adult rats: a dose-dependent study. AB - Cisplatin is one of the most active cytotoxic agents in the treatment of cancer, but its clinical use is associated with nephrotoxicity. In the present study we report the effects of different amounts of vitamin C (50, 100 or 200 mg kg( 1)body wt.) in rat kidneys treated with cisplatin (5 mg kg(-1)body wt.), using single doses of both compounds. Cisplatin administration induced lipid peroxidation which was accompanied by a decrease in renal glutathione level in animals killed 7 days after treatments. Furthermore, an increase in serum creatinine has been observed. Treatment of animals with vitamin C 10 min prior to the cisplatin inhibited cisplatin-mediated damage. Seven days after vitamin C plus cisplatin treatments, the depleted level of glutathione and changes in the creatinine clearance recovered to significant levels (P<0.05). Similarly, the enhanced serum creatinine levels which are indicative of renal injury showed a significant reduction (P<0.05) with the three doses of vitamin C tested. The protective effect of vitamin C was dose-dependent. The results suggest that vitamin C is an effective chemoprotective agent against nephrotoxicity induced by the antitumoral cisplatin in Wistar adult rats. PMID- 10704264 TI - Protective effect of triterpenes on calcium oxalate crystal-induced peroxidative changes in experimental urolithiasis. AB - Naturally occurring pentacyclic triterpenes of plant origin have been identified as possessing a wide range of pharmacological effects. Lupeol (Lupa-21,20(29) dien, 3beta-ol) has been found to be efficient in reducing the risk of stone formation in animals by way of preventing crystal-induced tissue damage and dilution of urinary stone-forming constituents. In the present study, two structurally related triterpenes, lupeol and betulin (Lupa-20(29)ene-3,28 diol) were assessed for their antilithiatic effect. Foreign body implantation method followed by supplementation of ammonium oxalate was adapted to induce stone formation in the bladder. This led to elevated lipid peroxidation and depleted antioxidant status in the renal tissues. Both the triterpenes were equally efficient in minimizing crystal-induced renal peroxidative changes measured in terms of malondialdehyde and subsequent tissue damage. The antioxidant status, comprising of the enzymatic and non-enzymatic components, was found to be significantly depleted in the kidney and bladder of stone-forming animals. Both lupeol and betulin were comparable in their ability to restore the thiol status and the antioxidant enzymes like superoxide dismutase, catalase and glutathione peroxidase. The mechanism by which the two compounds render protection against oxalate-induced toxic manifestations and free radical production may involve the inhibition of calcium oxalate crystal aggregation and enhancement of the body defence systems. 2000 Academic Press. PMID- 10704265 TI - How safe is neem extract with respect to thyroid function in male mice? AB - In this investigation we attempted to find out the hitherto unstudied adverse effects of neem (Azardirachta indica) leaf extract on the thyroid function of male mice. Neem leaf extract was orally administered in two different doses (40 mg and 100 mg kg(-1)day(-1)for 20 days). The extract exhibited differential effects. While the higher dose decreased serum tri-iodothyonine (T(3)) and increased serum thyroxine (T(4)) concentrations, no significant alterations of levels were observed in the lower dose group, indicating that the high concentrations of neem extract can be inhibitory to thyroid function, particularly in the conversion of T(4)to T(3), the major source of T(3)generation. A concomitant increase in hepatic lipid peroxidation (LPO) and a decrease in glucose-6-phosphatase (G-6-Pase) activity in the higher dosed group also indicated the adverse effect of neem extract despite an enhancement in the activities of two defensive enzymes, superoxide dismutase (SOD) and catalase (CAT). Thus, it appears that the higher concentration of neem extract may not be safe with respect to thyroid function and lipid peroxidation. PMID- 10704266 TI - The effects of carbamazepine and valproic acid on the erythrocyte glutathione, glutathione peroxidase, superoxide dismutase and serum lipid peroxidation in epileptic children. AB - Some epileptic drugs may change antioxidant enzyme activities in humans and experimental animals. Recent studies suggest that membrane lipid peroxidation may be causally involved in some forms of epilepsy, and the differences are reported in free radical scavenging enzyme levels. GSHpX, SOD, GSH are important parameters of antioxidant defence mechanisms. This study was undertaken to evaluate the effects of valproic acid and carbamazepine (CBZ) therapy on erythrocyte glutathione (GSH), glutathione peroxidase (GSHpX), superoxide dismutase (SOD) and lipid peroxidation. During the treatment with VPA or CBZ, the erythrocyte GSHpX and GSH levels of epileptic children were significantly changed as compared to those of health control subjects. The mean levels of serum lipid peroxidation and erythrocyte superoxide dismutase were not statistically different from controls. The methods used for investigation of glutathione peroxidase, superoxide dismutase, glutathione and serum lipid peroxidation were all based on spectrophotometric measurement. PMID- 10704267 TI - Hydrogen peroxide-induced oxidative damage and apoptosis in cerebellar granule cells: protection by Ginkgo biloba extract. AB - The ability of oxidative stress to induce apoptosis and the protective effects of Ginkgo biloba extract (EGb761) against this induction were studied in cultures of rat cerebellar granule cells. Cells were exposed to oxidative stress by treatment with 50 microm hydrogen peroxide+100 microm ferrous sulphate which generates hydroxyl radicals by Fenton reaction. Both morphological observation and biochemical analysis revealed that H(2)O(2)/FeSO(4)treatment induced apoptotic cell death in cerebellar granule cells, which was characterized by chromatin condensation and DNA fragmentation. During this process, the fluidity of the cell membrane decreased markedly, and the conformation of membrane proteins altered significantly. Pretreating cerebellar granule cells with the antioxidant EGb761 (Ginkgo biloba extract) effectively attenuated oxidative damage induced by H(2)O(2)/FeSO(4), and prevented cells from apoptotic cell death. The results suggested that EGb761 might be used as a potential drug for neuronal diseases associated with the excessive production of reactive oxygen species. PMID- 10704268 TI - Evidence for different interactions between beta(1)- and beta(2)-adrenoceptor subtypes with adenylyl cyclase in the rat brain: a concentration-response study using forskolin. AB - The aim of this study was to investigate beta(1)- and beta(2)-adrenoceptor signalling systems in the rat brain studying the synergistic effects between beta adrenoceptor agonists and forskolin- induced activation of adenylyl cyclase. Experiments were performed in slices from cerebral cortex and cerebellum because they contain mainly beta(1)- and almost exclusively beta(2)- adrenoceptors, respectively. Five beta-adrenergic agonists were used, clenbuterol, flerobuterol, isoproterenol, salbutamol, and tulobuterol. All agonists stimulated cyclic AMP accumulation in the cerebral cortex but flerobuterol was inactive in the cerebellum. Forskolin amplified the generation of cyclic AMP. Forskolin potentiation was observed in glial cells but not in neurons and was not dependent on the number of beta-adrenoceptors. In return the amplitude of the potentiation was highly dependent on the intrinsic activity of the agonist in the cerebral cortex whereas it was constant whatever the agonist tested in the cerebellum. To analyse this difference we developed a modelling approach using a concentration response study. Isoproterenol and forskolin stimulations of cyclic AMP production were studied either alone or in combination with increasing concentrations of forskolin and isoproterenol, respectively. In the cerebral cortex isoproterenol and forskolin were both able to potentiate the cyclic AMP accumulation induced by the other compound, whereas, in the cerebellum, isoproterenol was unable to increase the stimulation induced by forskolin. The results support the hypothesis that beta(1)- and beta(2)-adrenoceptors display distinct mechanisms of action in the signalling system by which they stimulate the accumulation of cyclic AMP. PMID- 10704269 TI - Sensitivity analysis of pharmacokinetic parameters in one-compartment models. AB - The quantitative properties of a compartmental model can be characterized by the pharmacokinetic parameters. The sensitivity analysis of pharmacokinetic parameters was studied in six kinds of one-compartment models. The sensitivity functions with respect to different pharmacokinetic parameters for these one compartment models were deduced using the partial differentiation. The pharmacokinetic parameters were ranked according to their sensitivity functions. PMID- 10704270 TI - Study of the evolution of blood and striatal levels of methyldopa: a microdialysis study in sinaortic denervated rats. AB - Pharmacokinetic of methyldopa (50 mg kg(-1)i.p.) was studied in anesthetized sham operated and sinoaortic denervated (SAD) rats by using the microdialysis technique. Vascular shunt probe was inserted into the carotid artery and concentric probe was placed in the striatum. Levels of methyldopa were measured by HPLC-EC. The number of animals in each group was six and normal distribution of the variables of the study was assumed. Peak concentrations in arterial blood of methyldopa were similar in both groups of rats but the elimination rate constant was 0.31+/-0.09 h(-1)for sham rats (n =6) and 1.28+/-0.31 h(-1)for SAD rats (n =6, P<0.05). Low levels of methyldopa and a more pronounced decrease were seen in striatum of sinoaortic denervated rats. In conclusion, by using a microdialysis technique, different kinetic profiles of methyldopa were observed in sham operated and sinoaortic denervated rats. PMID- 10704271 TI - Pharmacokinetics of the peripheral benzodiazepine receptor antagonist, PK 11195, in rats. The effect of dose and gender. AB - In spite of the extensive use of the peripheral benzodiazepine (BZ) receptor antagonist, PK 11195 (PK), in pharmacological studies, there is a lack of information of its pharmacokinetics in the rat. In this study the pharmacokinetics of PK were determined after bolus intravenous (i.v.) administration in rat. The effects of dose and gender were evaluated in Sprague Dawley age-matched male and female rats after the injection of PK (5, 10, 20 mg kg(-1)). Plasma was collected at 5-300 min. Levels of PK in plasma and brain were determined by a novel HPLC method. The stability of PK in blood in vitro was determined. PK is stable in rat blood in vitro. The pharmacokinetics of PK are described by a two-compartment model. The half-lives for distribution ( approximately 0.14 h) and elimination ( approximately 5.4 h) are not related to dose. The large volume of distribution (9-24 l kg(-1)) indicates an extensive distribution outside plasma. Total plasma clearance increases with increasing dose (23-42 ml min(-1)kg(-1)). The brain/plasma ratio ( approximately 3) is not related to dose. These finding suggest that the pharmacokinetics of PK are related to dose (5-20 mg kg(-1)) and gender in rat. PMID- 10704273 TI - The hamster heart is resistant to calcium paradox. AB - Reintroduction of Ca(2+)or modification of internal Ca(2+)stores by caffeine results in widespread irreversible injury. The adult golden hamster, however, is immune to such insult and the present report investigates the phenomenon. Isolated Langendorff perfused hamster and rat heart were subjected to 15 min Ca(2+)-free perfusion followed by 30 min of Ca(2+)perfusion at 37 degrees C. Caffeine was introduced during Ca(2+)-free perfusion in a number of experiments. Papillary muscles were processed for the ultra-structural study. The hamster heart did not exhibit the calcium paradox state whereas rat heart did. Hamster heart treated with caffeine either throughout or 5 min after starting Ca(2+)-free perfusion showed 70%+/-8 or 65%+/-8. 42 recovery, respectively, when Ca(2+)reperfusion was performed. Ultrastructure of muscle from both groups showed relaxed myocytes with slight disorientation of the sarcomere register. This disorientation was not seen in hamster hearts undergoing the conventional calcium paradox protocol. The hamster cardiac muscle is remarkably tolerant to [Ca(2+)]()i loading either induced by Ca(2+)reperfusion or caffeine-induced sarcoplasmic reticulum Ca(2+)release. Structural and functional characterization of Ca(2+)depletion and repletion in the hamster heart have been discussed. PMID- 10704272 TI - In vitro and in vivo vasodilating activity of nitroso derivatives gem substituted with electron-withdrawing groups. AB - A series of nitroso compounds gem -substituted with electron-withdrawing groups (R(2)C(X)NO, R=alkyl, X=NO(2), CN, Cl), were studied for their in vitro and in vivo vasodilating properties as well as for their ability to activate soluble guanylate cyclase (sGC) in RFL-6 cells. All the compounds, with the sole exception of chloro derivative, display good in vitro vasodilating action and are able to increase the basal level of cGMP. Their potencies as vasodilators decrease in the presence of oxyhaemoglobin, a scavenger of nitric oxide (NO). The haemodynamic profile of the most interesting compounds, assessed in anaesthetized pigs, is also in line with a release of NO from these compounds. PMID- 10704274 TI - The diuretic chlorthalidone normalizes baroreceptor and Bezold-Jarisch reflexes in DOCA-salt hypertensive rats. AB - The contributions of arterial baroreceptor and Bezold-Jarisch reflexes, and atrial natriuretic factor (ANF) to the anti-hypertensive effect of the diuretic chlorthalidone were investigated in rats with deoxycorticosterone acetate (DOCA) salt-induced hypertension. Chlorthalidone (8 mg rat(-1)day(-1)added to food) was given to one group during all 20 days of DOCA (8 mg kg(-1)s.c. twice per week) administration (preventive regimen) and, to another group, 20 days after DOCA treatment was initiated until the 40th day (therapeutic regimen). DOCA caused a significant increase in mean arterial pressure, reduced arterial baroreflex, and increased both the Bezold-Jarisch reflex and pro-ANF converting enzyme activity. Chlorthalidone reversed or prevented the DOCA-salt-induced hypertension, which was accompanied by the normalization of both the arterial baroreflex and the Bezold-Jarisch reflex. Additionally, both preventive and therapeutic regimens with chlorthalidone did cause normalization of the plasma sodium concentration and pro-ANF converting enzyme activity in the left atrium that follows DOCA-salt hypertension. Although it is difficult to determine the relative importance of each of the above regulatory mechanisms altered by chlorthalidone treatment, these data indicate that they may account for the prevention or decrease of DOCA salt-induced hypertension in rats. PMID- 10704275 TI - Does remote organ ischaemia trigger cardiac preconditioning during coronary artery surgery? AB - The aim of this study is to provide biochemical evidence of the occurrence of cardiac preconditioning via remote organ ischaemia on the patients undergoing coronary artery surgery. Eight male patients were randomly allocated into two groups. Blood samples were collected via coronary perfusion catheter immediately before cardiopulmonary bypass (point 0), prior to declamping aorta (point 1) and 5 min after declamping the aorta (point 2) to determine creatinine phosphokinase (CPK), CPK-MB and lactate dehydrogenase (LDH) levels in the control group. A tourniquet wrapped around the right upper extremity of the patient was inflated and deflated twice to perform 3 min of ischaemia separated with 2 min of reperfusion in the preconditioning group. Blood samples were withdrawn as described for the control group. Only LDH levels at point 2 were found to be significantly higher than the control group's. These data implied that preconditioning appeared to protect myocardium by enhancing anaerobic glycolysis. PMID- 10704276 TI - Somatostatin inhibits the release of noradrenaline induced by electrical stimulation of the rat mesenteric artery. AB - Somatostatin, a peptide with antisecretory and antiproliferative effects, coexists with noradrenaline in sympathetic neurons. Octreotide, a stable somatostatin analogue, prevents hypertension and cardiovascular structural changes induced by prolonged infusion of DPSPX (1,3-dipropyl-8 sulfophenylxanthine, a non-selective adenosine receptor antagonist) in rats. In the present work we investigated the effect of somatostatin and its analogue octreotide on the release of [(3)H]noradrenaline from sympathetic nerves in the rat mesenteric artery. Rat mesenteric arteries were incubated for 60 min with [(3)H]noradrenaline (0.2 microm), mounted in perifusion chambers, washed out for 90 min and electrically stimulated (2 Hz, 5 min, 50 mA). Radioactivity was measured in the tissue and in the perifusion fluid before, during and after stimulation. Both somatostatin and octreotide inhibited tritium release evoked by electrical stimulation of in vitro preparations of rat mesenteric arteries preloaded with [(3)H]noradrenaline. The maximal effects produced by octreotide and somatostatin were a 56 and 70% inhibition of noradrenaline release, respectively. For somatostatin an EC(50)=0. 18 n m (0.01 n m-2.2 n m;n =16) was calculated. When used alone, the somatostatin receptor antagonist, cyclo(7 aminoheptanoyl-Phe- d -Trp-Lys-Thr[BZL]) (CYCAM; 1 microm), had no effect on noradrenaline release induced by electrical stimulation. However, it was able to significantly antagonize the inhibitory effects of octreotide and somatostatin. These results are compatible with a negative modulatory role of somatostatin on sympathetic neurotransmission. PMID- 10704277 TI - Two genes are responsible for Griscelli syndrome at the same 15q21 locus. AB - Griscelli syndrome is a rare autosomal recessive disease characterized by pigment dilution, variable cellular immunodeficiency, and an acute phase of uncontrolled T lymphocyte and macrophage activation. We previously mapped the disease locus to 15q21 and showed that a MyoVa gene (HGMW-approved symbol MYO5A) defect leads to Griscelli syndrome. We report a second MyoVa mutation in a new patient, confirming this first finding. However, in four other Griscelli syndrome patients analyzed, the MYOVA protein is expressed, and no mutation can be detected in the MyoVa gene coding sequence, even in the alternatively spliced region for which exon-intron boundaries were characterized. Linkage analysis performed in 15 Griscelli families thus far studied confirms the first localization. However, fine haplotype analysis in three families strongly suggests the existence of a second gene at the same locus for Griscelli syndrome less than 7.3 cM distant from the MyoVa gene. PMID- 10704278 TI - Molecular cytogenetic resources for chromosome 4 and comparative analysis of phylogenetic chromosome IV in great apes. AB - We have generated a panel of 55 somatic cell hybrids retaining fragments of human chromosome 4. Each hybrid has been characterized cytogenetically by FISH and molecularly by 37 STSs, evenly spaced along the chromosome. The panel can be exploited to map subregionally DNA sequences on chromosome 4 and to generate partial chromosome paints useful in the characterization of chromosomal rearrangements involving this chromosome. Furthermore, a panel of 84 YACs mapping on chromosome 4 has been characterized by FISH. A subset of this panel is recognized by STSs used in the somatic cell hybrid characterization. In this way a correlation between the genetic and the physical maps can be established. These resources have been used to investigate the conservation of the phylogenetic chromosome IV in great apes. The results indicate that all the pericentric inversions that differentiate chromosome IV in these species are distinct and that one of the breakpoints frequently lies very close to the centromere. In 4 instances, the YAC containing the breakpoint was identified. The breakpoint in IVq of PTR and MMU lies in the same YAC, suggesting that this breakpoint has been utilized twice in the evolutionary history of this chromosome. PMID- 10704279 TI - Lop12, a mutation in mouse Crygd causing lens opacity similar to human Coppock cataract. AB - A new cataract mutation was discovered in an ongoing program to identify new mouse models of hereditary eye disease. Lens opacity 12 (Lop12) is a semidominant mutation that results in an irregular nuclear lens opacity similar to the human Coppock cataract. Lop12 is associated with a small nonrecombining segment that maps to mouse Chromosome 1 close to the eye lens obsolescence mutation (Cryge(Cat2-Elo)), a member of the gamma-crystallin gene cluster (Cryg). Using a systemic candidate gene approach to analyze the entire Cryg cluster, a G to A transition was found in exon 3 of Crygd associated with the Lop12 mutation and has been designated Crygd(Lop12). The mutation Crygd(Lop12) leads to the formation of an in-frame stop codon that produces a truncated protein of 156 amino acids. It is predicted that the defective gene product alters protein folding of the gamma-crystallin(s) and results in lens opacity. PMID- 10704280 TI - Human BAC ends quality assessment and sequence analyses. AB - End sequences from bacterial artificial chromosomes (BACs) provide highly specific sequence markers in large-scale sequencing projects. To date, we have generated >300,000 end sequences from >186,000 human BAC clones with an average read length of >460 bp for a total of 141 Mb covering approximately 4.7% of the genome. Over 60% of the clones have BAC end sequences (BESs) from both ends representing more than fivefold coverage of the human genome by the paired-end clones. Our quality assessments and sequence analyses indicate that BESs from human BAC libraries developed at The California Institute of Technology (CalTech) and Roswell Park Cancer Institute have similar properties. The analyses have highlighted differences in insert size for different segments of the CalTech library. Problems with the fidelity of tracking of sequence data back to physical clones have been observed in some subsets of the overall BES dataset. The annotation results of BESs for the contents of available genomic sequences, sequence tagged sites, expressed sequence tags, protein encoding regions, and repeats indicate that this resource will be valuable in many areas of genome research. PMID- 10704281 TI - Organization and parent-of-origin-specific methylation of imprinted Peg3 gene on mouse proximal chromosome 7. AB - Peg3 is the first imprinted gene to be identified on mouse proximal chromosome 7; the human PEG3 homologue is on chromosome 19q13.4. Peg3 encodes a C(2)H(2)-type zinc finger protein that is expressed only from the paternal allele in embryos and adult brain. The gene has been shown to regulate maternal behavior and offspring growth and has been implicated in the TNF-NFkappaB signal pathway. Here we show that Peg3 consists of nine exons spanning 26 kb. The 5' region of the gene contains a region rich in repeated sequences and a CpG island. Analysis of expressed sequence tags revealed a transcript present upstream of the island and on the strand opposite to Peg3. These structural features and DNA sequences are conserved in mouse and human. The 5' region of Peg3 is preferentially methylated on the inactive maternal allele, as shown by comparing embryos with paternal (PatDp. prox7) and maternal (MatDp.prox7) duplication of proximal chromosome 7. Recently, a new maternally expressed Zim1 gene located downstream of Peg3 was identified, which suggested that another imprinted cluster is present on proximal chromosome 7. PMID- 10704282 TI - Expression profile viewer (ExProView): a software tool for transcriptome analysis. AB - A software tool, Expression Profile Viewer (ExProView), for analysis of gene expression profiles derived from expressed sequence tags (ESTs) and SAGE (serial analysis of gene expression) is presented. The software visualizes a complete set of classified transcript data in a two-dimensional array of dots, a "virtual chip," in which each dot represents a known gene as characterized in the transcript databases Expressed Gene Anatomy Database or UniGene. The virtual chip display can be changed between representations of different conceptual systems for gene/protein classification and grouping. Four alternative projections are currently available: (i) cellular role, (ii) subcellular compartment, (iii) chromosome localization, and (iv) total UniGene display. However, the chip can be adapted to any other desired layout. By selecting dots, further information about the represented genes is obtained from the local database and WWW links. The software thus provides a visualization of global mRNA expression at the descriptive level and guides in the exploration of patterns of functional expression, while maintaining direct access to detailed information on each individual gene. To evaluate the software, public EST and SAGE gene expression data obtained from the Cancer Genome Anatomy Project at the National Center for Biotechnology Information were analyzed and visualized. A demonstration of the software is available at http://www.biochem.kth. se/exproview/. PMID- 10704283 TI - Identification of 40 genes on a 1-Mb contig around the IL-4 cytokine family gene cluster on mouse chromosome 11. AB - Five related cytokine genes, interleukin 3 (Il3), interleukin 4 (Il4), interleukin 5 (Il5), interleukin 13 (Il13), and granulocyte-macrophage colony stimulating factor (Csfgm or Csf2), are tightly linked on mouse chromosome 11. We now describe a 1-Mb transcript map of this cytokine cluster. Genomic clones obtained by screening mouse bacterial artificial chromosome (BAC) and P1-derived artificial chromosome (PAC) libraries were subcloned into the pSPL3 expression vector and transfected into COS7 cells for exon trapping. In total, 118 distinct, putative exons were sequenced and characterized, mapping up to 29 distinct genes to the mouse cluster, including Il4 and Csf2. Northern blot and RT-PCR analyses indicate that all of these genes are expressed. Analysis of 1 Mb of published sequence from the region of conserved synteny on human chromosome 5q31-q33 identified 45 gene candidates, including 35 expressed genes in the human IL-4 cytokine gene cluster. Probes for 20 human genes were tested for cross hybridization to murine BAC and PAC clones, thereby mapping 11 additional genes to the mouse complex. Thus, a total of 40 genes including 6 cytokine genes have been physically mapped within 1 Mb of mouse chromosome 11. Gene order in this complex is similar, but not identical, between human and mouse. The integrated physical and transcript maps should prove valuable as a complement to genomic sequencing and expression-dependent transcript maps of this segment of the genome. PMID- 10704284 TI - Comparative sequence analysis of 634 kb of the mouse chromosome 16 region of conserved synteny with the human velocardiofacial syndrome region on chromosome 22q11.2. AB - Mouse genomic DNA sequence extending 634 kb on proximal mouse chromosome 16 was compared to the corresponding human sequence from chromosome 22q11.2. Haploinsufficiency for this region results in velocardiofacial syndrome (VCFS) in humans. The mouse region is rearranged into three conserved blocks relative to human, but gene content and position are highly conserved within these blocks. Examination of the boundaries of one of these blocks suggested that the evolutionary chromosomal rearrangement occurred in the mouse lineage, resulting in inactivation of the mouse orthologue of ZNF74. Sequence analysis identified 21 genes and 15 ESTs. These include 2 novel genes, Srec2 and Cals2, and previously undescribed splice variants of several other genes. Exon discovery was carried out using GRAIL2, MZEF, or comparative analysis across 491 kb of conserved mouse and human sequence. Sequence comparison was highly effective, identifying every gene and nearly every exon without the high frequency of false-positive predictions seen when algorithmic methods were used alone. In combination, these procedures identified every gene with no false-positive predictions. Comparative sequence analysis also revealed regions of extensive conservation among noncoding sequences, accounting for 6% of the sequence. A library of such sequences has been established to form a resource for generalized studies of regulatory and structural elements. PMID- 10704285 TI - Genomic structure and DNA binding properties of the human zinc finger transcriptional repressor AP-2rep (KLF12). AB - We recently cloned a novel murine transcriptional repressor, the Kruppel-like zinc finger protein AP-2rep (HGMW-approved symbol KLF12), that binds to a regulatory element in the AP-2alpha gene promoter. In the present study, we characterize the human AP-2rep homolog and describe expression patterns in human urogenital and lymphoma cell lines. The predicted human protein of 402 amino acids exhibits 95.8% identity and 98.5% similarity to the murine AP-2rep peptide. The genomic locus of human AP-2rep consists of seven exons and was assigned to chromosome 13q22 by fluorescence in situ hybridization to metaphase chromosomes. Human AP-2rep repressed both reporter expression from a transiently transfected AP-2alpha promoter and the endogenous AP-2alpha gene and inversely was negatively regulated by AP-2alpha. The consensus motif CAGTGGG was identified by an in vitro binding site selection assay. In summary, our data further point to an important role of AP-2rep as a transcriptional silencer and reveal reciprocal regulation of AP-2alpha and AP-2rep. PMID- 10704286 TI - Mapping and developmental expression analysis of the WD-repeat gene Preb. AB - We have isolated from mouse a novel WD-motif-containing gene designated Preb. This gene encodes a predicted protein of 416 amino acids and has significant homology with other members of the WD-motif gene superfamily that play a role in cell fate determination. Preb maps to the proximal end of chromosome 5 in mouse, near the Hmx1 homeobox gene. Preb is detectable in early stage embryos in the peripheral nervous system, developing liver, and surface ectoderm. Later, Preb is expressed in the anterior portion of Rathke's pouch, which gives rise to the anterior pituitary, the organ responsible for the production of prolactin and other hormones. In midgestation embryos, the most extensive expression of Preb is observed in the perichondrium of the craniofacial, axial, and appendicular skeleton. The expression pattern of Preb in murine embryos suggests a potential role in the specification of multiple cell types, in particular, the fetal skeleton. PMID- 10704287 TI - Characterization, chromosomal assignment, and tissue expression of a novel human gene belonging to the ARF GAP family. AB - We have identified and characterized a novel human ADP-ribosylation factor GTPase activating protein (ARFGAP1) gene that is related to other members of the ARF GAP family. The full-length cDNA for human ARFGAP1 was cloned following the identification of an EST obtained by large-scale cDNA library sequencing through a Blast search of public databases. Structurally, ARFGAP1 encodes a polypeptide of 516 amino acids, which contained a typical GATA-1-type zinc finger motif (CXXCX(16)CXXC) with the four cysteine residues that are highly conserved among other members of the ARF GAP family. The conserved ARF GAP domain may emphasize the biological importance of this gene. The ARFGAP1 gene, which contained 16 exons ranging from 0.5 to 9.3 kb, was mapped to human chromosome 22q13.2-q13.3 using radiation hybridization and in silico analyses. ARFGAP1 is strongly expressed in endocrine glands and testis. Interestingly, the expression of ARFGAP1 in testis is about sixfold higher than that in ovary, indicating a possible role of ARFGAP1 in the physiological function of sperm. Expression of ARFGAP1 in four human fetal tissues and seven cancer cell lines was also detected. PMID- 10704288 TI - A sequence-ready map of the Usher syndrome type III critical region on chromosome 3q. AB - Usher syndrome type 3 (USH3; MIM 276902) is an autosomal recessive disorder associated with progressive hearing loss and retinal degeneration. We recently refined the localization of USH3 to a 1-cM genetic interval between markers D3S1299 and D3S3625. We have now constructed a bacterial artificial chromosome contig over the region. Novel polymorphic markers were generated and physically fine-mapped, allowing further narrowing of the critical interval to a 250-kb genomic fragment. Of seven ESTs mapping to the initial critical region, WI-11588 and SHGC-133 represent the human SIAH2 gene, which was excluded as a candidate for USH3 by sequencing and subsequently, by its position. KIAA0001 and D3S3882 derive from the transcript of a putative G-protein-coupled receptor gene that was excluded as a candidate by sequencing of patient DNA. These data provide a basis for the sequencing and final characterization of the USH3 region and isolation of the disease gene. PMID- 10704289 TI - Deletions within the mouse beta-globin locus control region preferentially reduce beta(min) globin gene expression. AB - The mouse beta-globin gene cluster is regulated, at least in part, by a locus control region (LCR) composed of several developmentally stable DNase I hypersensitive sites located upstream of the genes. In this report, we examine the level of expression of the beta(min) and beta(maj) genes in adult mice in which HS2, HS3, or HS5,6 has been either deleted or replaced by a selectable marker via homologous recombination in ES cells. Primer extension analysis of RNA extracted from circulating reticulocytes and HPLC analysis of globin chains from peripheral red blood cells revealed that all mutations that reduce the overall output of the locus preferentially decrease beta(min) expression over beta(maj). The implications of these findings for the mechanism by which the LCR controls expression of the beta(maj) and beta(min) promoters are discussed. PMID- 10704290 TI - A transcript map of a 10-Mb region of chromosome 19: a source of genes for human disorders, including candidates for genes involved in asthma, heart defects, and eye development. AB - Several projects have produced maps of the physical position of genes within the human genome, either on a genome-wide scale or of a more detailed subsection of a chromosome. However, these maps largely rely on the mapping of expressed sequences (cDNAs and ESTs) back onto physical maps by their localization onto specific fragments of DNA within the radiation hybrid panels. In this report we present a gene map of a section of chromosome 19 that has been derived by combining the use of a method of gene identification (exon trapping) that does not rely on expression patterns, with data available in the genome databases to produce a fine-detailed transcript map. This map also provides several potential candidates for disorders that map to this region of the genome. Details of the maps and more detailed descriptions of cosmid contigs, exon sequences, and expression patterns for the 96 exons that form the basis of this transcript map are available on a series of Web pages that are referenced in this report. These Web pages can be accessed from http://www.nottingham.ac.uk/ pdzmgh/tm/livemap19q. html. PMID- 10704291 TI - Polymorphic markers for the arylsulfatase A gene reveal a greatly expanded meiotic map for the human 22q telomeric region. AB - Two microsatellite markers, D22S1743 and D22S1744, were developed for the arylsulfatase A (ARSA) region of chromosome 22q. Linkage analysis for 171 families, using nine reference markers covering all of 22q, placed these new markers 2.0 Kosambi cM distal to D22S526, making them more distal than any microsatellite markers currently on the Genethon or Marshfield linkage maps. Recombination between proximal markers D22S270/D22S683 and D22S446/D22S311 exhibited increased rates of female meiotic recombination compared to male recombination (P < 0.01). In contrast, the region encompassing sJCW16, D22S526, D22S1743, and D22S1744 exhibited relatively greater recombination in males (1.1 cM for females and 7.5 cM for males; chi(2); P < 0.005). These four distal markers lie in a region of hyperrecombination having a sex-averaged recombination ratio of between 8.3 (D22S1843/D22S1744) and 12 cM (sJCW16/D22S526) per megabase. PMID- 10704292 TI - The human CYP2C locus: a prototype for intergenic and exon repetition splicing events. AB - In human there are four known CYP2C genes that have been mapped to chromosome 10q24 with the order Cen-2C18-2C19-2C9-2C8-Tel. Previously we have shown that splicing events joining exons from the neighboring 2C18 and 2C19 genes occur in human liver and epidermis. Here evidence is presented that the terminal genes of this cluster, 2C18 and 2C8, are also involved in intergenic splicing. Most interestingly, several of these 2C18/2C8 RNAs were composed of all nine exons, thus conceivably having the potential for coding functional proteins. Moreover, chimeric RNA species consisting of exons originating not only from the CYP2C8 and CYP2C18 genes, but also from the CYP2C19 gene were detected. In all cases the exons from the different CYP2C genes were joined at the correct canonical splice sites. However, the closely linked RBP4 gene is not participating in intergenic splicing with the CYP2C genes. In addition, CYP2C8 gene expression was found to generate a variety of scrambled RNA molecules including species that contained repetitions of certain exons. PMID- 10704293 TI - Pollen coupling of forest trees: forming synchronized and periodic reproduction out of chaos. AB - Many of the tree species in mature forests show masting; their reproductive activity has a large variance between years and is often synchronized between different individuals. In this paper, we analyse a globally coupled map model in which trees accumulate photosynthate every year, produce flowers when the energy reserve level exceeds a threshold, and set seeds and fruits at a rate limited by pollen availability. Without pollen limitation, the trees in the forest show independent chaotic fluctuation. Coupling of trees via pollen exchange results in reproduction being synchronized partially or completely over the forest. The whole forest shows diverse dynamical behaviors determined by the values of two essential parameters; the depletion coefficient k and the coupling strength beta. We find perfectly synchronized periodic reproduction, synchronized reproduction with a chaotic time series, clustering phenomena, and chaotic reproduction of trees without synchronization over individuals. There are many parameter windows in which synchronized reproduction of trees shows a stable periodic fluctuation. For perfectly synchronized forests, we can calculate all the Lyapunov exponents analytically. They show that synchronized reproduction of all the trees in the forest can only occur when trees flower at low (but positive) levels in a significant fraction of years, resulting in small fruit sets due to outcrossed pollen limitation. This is consistent with the observation that the distinction between mast years and non-mast years is often not clear cut. PMID- 10704294 TI - The evolution of parasite virulence and transmission rate in a spatially structured population. AB - If the transmission occurs through local contact of the individuals in a spatially structured population, the evolutionarily stable (ESS) traits of parasite might be quite different from what the classical theory with complete mixing predicts. In this paper, we theoretically study the ESS virulence and transmission rate of a parasite in a lattice-structured host population, in which the host can send progeny only to its neighboring vacant site, and the transmission occurs only in between the infected and the susceptible in the nearest-neighbor sites. Infected host is assumed to be infertile. The analysis based on the pair approximation and the Monte Carlo simulation reveal that the ESS transmission rate and virulence in a lattice-structured population are greatly reduced from those in completely mixing population. Unlike completely mixing populations, the spread of parasite can drive the host to extinction, because the local density of the susceptible next to the infected can remain high even when the global density of host becomes very low. This demographic viscosity and group selection between self-organized spatial clusters of host individuals then leads to an intermediate ESS transmission rate even if there is no tradeoff between transmission rate and virulence. The ESS transmission rate is below the region of parasite-driven extinction by a finite amount for moderately large reproductive rate of host; whereas, the evolution of transmission rate leads to the fade out of parasite for small reproductive rate, and the extinction of host for very large reproductive rate. PMID- 10704295 TI - Heat transfer in a microvascular network: the effect of heart rate on heating and cooling in reptiles (Pogona barbata and Varanus varius). AB - Thermally-induced changes in heart rate and blood flow in reptiles are believed to be of selective advantage by allowing animal to exert some control over rates of heating and cooling. This notion has become one of the principal paradigms in reptilian thermal physiology. However, the functional significance of changes in heart rate is unclear, because the effect of heart rate and blood flow on total animal heat transfer is not known. I used heat transfer theory to determine the importance of heat transfer by blood flow relative to conduction. I validated theoretical predictions by comparing them with field data from two species of lizard, bearded dragons (Pogona barbata) and lace monitors (Varanus varius). Heart rates measured in free-ranging lizards in the field were significantly higher during heating than during cooling, and heart rates decreased with body mass. Convective heat transfer by blood flow increased with heart rate. Rates of heat transfer by both blood flow and conduction decreased with mass, but the mass scaling exponents were different. Hence, rate of conductive heat transfer decreased more rapidly with increasing mass than did heat transfer by blood flow, so that the relative importance of blood flow in total animal heat transfer increased with mass. The functional significance of changes in heart rate and, hence, rates of heat transfer, in response to heating and cooling in lizards was quantified. For example, by increasing heart rate when entering a heating environment in the morning, and decreasing heart rate when the environment cools in the evening a Pogona can spend up to 44 min longer per day with body temperature within its preferred range. It was concluded that changes in heart rate in response to heating and cooling confer a selective advantage at least on reptiles of mass similar to that of the study animals (0. 21-5.6 kg). PMID- 10704296 TI - A classification of possible routes of Darwinian evolution. AB - A classification of four possible routes of Darwinian evolution is presented. These are serial direct evolution, parallel direct evolution, elimination of functional redundancy, and adoption from a different function. This classification provides a conceptual framework within which to investigate the accessibility by Darwinian evolution of complex biological structures. PMID- 10704297 TI - How to make a biological switch. AB - Some biological regulatory systems must "remember" a state for long periods of time. A simple type of system that can accomplish this task is one in which two regulatory elements negatively regulate one another. For example, two repressor proteins might control one another's synthesis. Qualitative reasoning suggests that such a system will have two stable states, one in which the first element is "on" and the second "off", and another in which these states are reversed. Quantitative analysis shows that the existence of two stable steady states depends on the details of the system. Among other things, the shapes of functions describing the effect of one regulatory element on the other must meet certain criteria in order for two steady states to exist. Many biologically reasonable functions do not meet these criteria. In particular, repression that is well described by a Michaelis-Menten-type equation cannot lead to a working switch. However, functions describing positive cooperativity of binding, non-additive effects of multiple operator sites, or depletion of free repressor can lead to working switches. PMID- 10704298 TI - Ecological symmetry breaking can favour the evolution of altruism in an action response game. AB - The evolution of altruistic behaviour is studied in a simple action-response game with a tunable degree of conflict of interest. It is shown that for the continuous, mixed-medium approach no stable polymorphism favours altruism. Ecological dynamics are explored with the addition of a spatial dimension and a local energy variable. A continuous spatial model with finite local range does not introduce any substantial difference in the results with respect to the level of altruism. However, the model illustrates how ecological coupling may lead to the formation of stable spatial patterns in the form of discrete and isolated clusters of players as a consequence of inverse density dependence. A discrete, individual-based model is built in which local interactions are also modelled as occurring within a finite neighbourhood of each individual and spatial positions are not restricted as in lattice models. This model shows substantially different results. A high level of altruism is observed for low (but positive) degrees of conflict and this level decreases linearly for higher degrees of conflict. The evolution of altruism is explained by studying the broken symmetries introduced by the spatial clusters themselves, mainly between their central and peripheral regions which, in combination with the discrete and the stochastic nature of the model, result in the stabilization of strategies in which players behave altruistically towards the same type. As a consequence of the activity of the players, energy resources at the centre of an altruistic cluster are very depleted; so much so that, for low conflict, fitter non-altruistic mutants may initially invade only to become locally extinct due to their less efficient use of energy as their numbers increase. In peripheral regions invader may subsist; however, for geometrical reasons long-lasting genealogies tend to originate only at the centre of a cluster. PMID- 10704299 TI - Mathematical analysis of the relative contributions of decreased production and increased peripheral destruction in idiopathic thrombocytopenic purpura and implications in splenectomy. AB - We utilize a model of platelet concentration kinetics and bone marrow production based on three terms (a constant loss term, a random loss term and a higher order loss term) to compare a hypoplastic bone marrow patient and a patient with Idiopathic Thrombocytopenic Purpura (ITP) for the same platelet concentration. We compare this model to published data and show that in many ITP patients there is an overall decrease in platelet production. However, for almost all cases of ITP there is an increase in peripheral platelet destruction, even in those cases where total bone marrow production is less than that in a normal individual or is severely depressed. We are able to graphically depict the variable contributions of decreased production and increased peripheral destruction in patients with ITP and hence give insight into their relative contributions in a given patient. We apply a unique feature of our model, the newly postulated destruction term proportional to the platelet concentration squared (the higher order loss term), to explain cases of antibody negative ITP. Application of our model to data on patients splenectomized as treatment for ITP shows promise in predicting which patients are likely to respond. PMID- 10704300 TI - Modeling the evolution of genetic architecture: A continuum of alleles model with pairwise AxA epistasis. AB - A continuum of alleles model with pair-wise AxA epistasis is proposed and its transmission genetic, and variational properties are analysed. The basic idea is that genes control the values of underlying variables, which affect the genotypic value of phenotypic characters proportional to a "scaling factor". Epistasis is the influence of one gene on the average effect of another gene. In this model, epistasis is introduced as a mutational effect of one gene on the scaling factors of another gene. In accordance with empirical results, the model assumes that the average direct effect of mutations is zero, as is the average epistatic effect. The model predicts that, on average, a mutation at one locus increases the expected mutational variance of mutations at another interacting locus. The increase in mutational variance is predicted to be equal to the variance of the pair-wise epistatic effects. This result is consistent with the observation that mutant phenotypes tend to be more variable than the wildtype phenotype. Another generic result of this model is that the frequency of canalizing mutations can at most be equal to the frequency of de-canalizing mutations. Furthermore, it is predicted that the mutational variance of a character increases at least linearly with the size of the character; hence this model is scale variant. In the case of two characters it is shown that the dimensionality of the locus-specific mutational effect distribution is invariant, i.e. the rank of the mutational covariance matrix M is invariant. While in additive models the mutational covariance matrix is always and entirely invariant, the invariance in the case of epistatic models is unexpected. Epistatic interactions can change the magnitude of the mutational (co)variances at a locus and can thus influence the structure of the mutational covariance matrix. However, in the present model the dimensionality of the mutational effect distribution remains the same. A consequence of this result is that, in this model, the genetic architecture of a set of characters is always evolvable i.e. no hard constraints can evolve. PMID- 10704301 TI - A dynamical model for the growth and size distribution of multiple metastatic tumors. AB - Metastasis is the spread of tumors culminating in the establishment of one or more secondary tumors at remote sites. In deciding the best treatment for cancer therapy, estimations of the colony size of metastatic tumors and predictions of the future spread of colonies are needed. A dynamical model for the colony size distribution of multiple metastatic tumors is presented here. The dynamics is described by equations that incorporate both the colonization by metastasis and the growth of each colony. When the colony growth is subject to the Gompertz function, the explicit solution obtained tends to an asymptotic stable distribution that shows a monotonically decreasing or U-shaped pattern according to the values of clinically significant parameters, such as the colonization coefficient and the fractal dimension of blood vessels. This predicted colony size distribution agrees well with successive data of a clinically observed size distribution of multiple metastatic tumors of liver cancer. The combined analysis of the theoretical colony size distribution and clinical data will give useful information on the diagnosis and the therapy for cancer patients. PMID- 10704302 TI - Variable zinc coordination in endostatin. AB - Endostatin is a proteolytic fragment of collagen XVIII that potently inhibits angiogenesis and tumour growth. Human endostatin contains a zinc ion, bound near the N terminus, which was not observed in the original structure of mouse endostatin at pH 5. Controversial data exist on the role of this zinc ion in the anti-tumour activity. We report two new crystal structures of mouse endostatin at pH 8.5 with bound zinc. One crystal form shows a metal ion coordination similar to that in human endostatin (His132, His134, His142, Asp207), but the conformation of the N-terminal segment is different. In the other crystal form, Asp136 replaces His132 as a zinc ligand. Site-directed mutagenesis of zinc binding residues demonstrates that both coordination geometries occur in solution. The large degree of structural heterogeneity of the zinc-binding site has implications for endostatin function. We conclude that zinc is likely to play a structural rather than a critical functional role in endostatin. PMID- 10704303 TI - Mot protein assembly into the bacterial flagellum: a model based on mutational analysis of the motB gene. AB - The 308 residue MotB protein anchors the stator complex of the Escherichia coli flagellar motor to the peptidoglycan of the cell wall. Together with MotA, it comprises the transmembrane channel that delivers protons to the motor. At the outset of the mutational analysis of MotB described here, we found that the non motile phenotype of a DeltamotAB strain was rescued better by a pmotA(+)B(+) plasmid than the non-motile phenotype of a DeltamotB strain was rescued by a pmotB(+) plasmid. Transcription in each case was from the inducible tac promoter but relied on the native ribosome-binding site (RBS). This result confirms that translational coupling to motA is important for normal translation of the motB mRNA, since overproduction of MotA in trans did not improve complementation by pmotB. However, introduction of an optimized RBS into pmotB (to generate pmotB(o)) did. To dissect the function of the periplasmic domain of MotB, site directed mutagenesis was used to replace Gln, Ser, and Tyr codons scattered throughout motB with amber (UAG) codons. Plasmid-borne motB(am) genes were introduced into sup(o), supE, and supF strains to see what motility defects were imposed by particular amber mutations and whether the defects could be suppressed by amber-suppressor tRNAs inserting the native or heterologous amino acids. Amber mutations at codon 268 or earlier in pmotB, and at codon 261 or earlier in pmotB(o) or pmotAB, eliminated motility. Thus, in agreement with the deletion analysis of motB by another laboratory, we conclude that the portion of MotB carboxyl-terminal to its peptidoglycan-binding motif (residues 161 to 264) is not essential. In strains containing supE or supF alleles, motility defects associated with motB(am) mutations were suppressed weakly, if at all, in pmotB. In contrast, motility defects conferred by most motB(am) mutations in pmotB(o) or pmotAB could be suppressed to a significant extent. However, the S18(am), Q100(am), Q112(am), Q124(am), Y201(am), and Y208(am) mutations were still suppressed extremely poorly. Full-length MotB was present at very low levels in suppressor strains containing the first four mutations, but Y201(am) and Y208(am) were suppressed efficiently at the translational level. We suggest that a translational pause by suppressor tRNAs reading UAG at these two positions may divert the nascent polypeptide into an alternative folding pathway that traps MotB in a non-functional conformation. We further propose that MotA and MotB form a stable pre-assembly complex in the membrane. In this complex, MotB exists in a form that cannot associate with peptidoglycan and blocks the proton-conducting channel. Opening of the channel and attachment to the cell wall may occur when the complex collides with a flagellar basal body and MotA makes specific contacts with the C ring and/or the MS ring. PMID- 10704304 TI - A minimal system for Tn7 transposition: the transposon-encoded proteins TnsA and TnsB can execute DNA breakage and joining reactions that generate circularized Tn7 species. AB - In the presence of ATP and Mg(2+), the bacterial transposon Tn7 translocates via a cut and paste mechanism executed by the transposon-encoded proteins TnsA+TnsB+TnsC+TnsD. We report here that in the presence of Mn(2+), TnsA+TnsB alone can execute the DNA breakage and joining reactions of Tn7 recombination. ATP is not essential in this minimal system, revealing that this cofactor is not directly involved in the chemical steps of recombination. In both the TnsAB and TnsABC+D systems, recombination initiates with double-strand breaks at each transposon end that cut Tn7 away from flanking donor DNA. In the minimal system, breakage occurs predominantly at a single transposon end and the subsequent end joining reactions are intramolecular, with the exposed 3' termini of a broken transposon end joining near the other end of the Tn7 element in the same donor molecule to form circular transposon species. In contrast, in TnsABC+D recombination, breaks occur at both ends of Tn7 and the two ends join to a target site on a different DNA molecule to form an intermolecular simple insertion. This demonstration of the capacity of TnsAB to execute breakage and joining reactions supports the view that these proteins form the Tn7 transposase. PMID- 10704305 TI - Mutational analysis of archaeal histone-DNA interactions. AB - Site-specific mutagenesis of the hmfB gene cloned from the archaeon Methanothermus fervidus, followed by expression in Escherichia coli, has been used to generate approximately 90 recombinant (r) variants of the archaeal histone HMfB. The abilities of these variants to form stable archaeal nucleosome containing complexes with linear pBR322 DNA, and with an 89 bp restriction fragment of this DNA have been determined. Variants that failed to form such complexes, based on negative gel-shift assays, had substitutions at the N terminus or within the alpha1, L1 and L2 regions of the rHMfB histone fold, at sites predicted to be homologous to eucaryal histone fold residues that contact the DNA in the eucaryal nucleosome. Variants that failed to give gel shifts were further assayed for their abilities to facilitate ligase-catalyzed circularization of a linear 88 bp DNA molecule, and to reduce the ellipticity of a DNA solution at 275 nm (theta(275)). Consistent with cooperative but independent sites of DNA binding, a combination of three residue substitutions, one each in alpha1, L1 and L2, was required to generate a rHMfB variant with no detectable DNA binding based on gel shift, circularization and theta(275) reduction assays. PMID- 10704306 TI - Roles of the ccoGHIS gene products in the biogenesis of the cbb(3)-type cytochrome c oxidase. AB - In many bacteria the ccoGHIS cluster, located immediately downstream of the structural genes (ccoNOQP) of cytochrome cbb(3) oxidase, is required for the biogenesis of this enzyme. Genetic analysis of ccoGHIS in Rhodobacter capsulatus demonstrated that ccoG, ccoH, ccoI and ccoS are expressed independently of each other, and do not form a simple operon. Absence of CcoG, which has putative (4Fe 4S) cluster binding motifs, does not significantly affect cytochrome cbb(3) oxidase activity. However, CcoH and CcoI are required for normal steady-state amounts of the enzyme. CcoI is highly homologous to ATP-dependent metal ion transporters, and appears to be involved in the acquisition of copper for cytochrome cbb(3) oxidase, since a CcoI-minus phenotype could be mimicked by copper ion starvation of a wild-type strain. Remarkably, the small protein CcoS, with a putative single transmembrane span, is essential for the incorporation of the redox-active prosthetic groups (heme b, heme b(3 )and Cu) into the cytochrome cbb(3) oxidase. Thus, the ccoGHIS products are involved in several steps during the maturation of the cytochrome cbb(3) oxidase. PMID- 10704307 TI - Hyperstable stacked-disk structure of tobacco mosaic virus protein: electron cryomicroscopy image reconstruction related to atomic models. AB - The stacked disk aggregate of tobacco mosaic virus protein is an intriguing object due to its high degree of stability, in spite of indications that the aggregate is held together to a great extent by water-mediated interactions between adjacent protein rings. Here, we present a set of models that were constructed using the atomic coordinates of the four-layer aggregate, and compare these with a three-dimensional reconstruction of the stacked disk obtained by means of cryoelectron microscopy and helical image processing. The comparison of the four possible models of the stacked disk with the data shows that there is a better correlation of the data with the left-handed model built from the A-A ring pair coordinates than with the two models involving the A-B ring pair, or with the right-handed model of the A-A ring pair. This establishes that the packing of the protein subunits in the stacked disk is different from that previously believed. We also note some differences between the observed structure and A-A ring pair model in the region of the flexible loop at small radius that might be an indication of conformational differences that give rise to the stability of the aggregate. PMID- 10704308 TI - Structural studies of soluble oligomers of the Alzheimer beta-amyloid peptide. AB - Recent studies have suggested that non-fibrillar soluble forms of Abeta peptides possess neurotoxic properties and may therefore play a role in the molecular pathogenesis of Alzheimer's disease. We have identified solution conditions under which two types of soluble oligomers of Abeta40 could be trapped and stabilized for an extended period of time. The first type of oligomers comprises a mixture of dimers/tetramers which are stable at neutral pH and low micromolar concentration, for a period of at least four weeks. The second type of oligomer comprises a narrow distribution of particles that are spherical when examined by electron microscopy and atomic force microscopy. The number average molecular mass of this distribution of particles is 0.94 MDa, and they are are stable at pH 3 for at least four weeks. Circular dichroism studies indicate that the dimers/tetramers possess irregular secondary structure that is not alpha-helix or beta-structure, while the 0.94 MDa particles contain beta-structure. Fluorescence resonance energy transfer experiments indicate that Abeta40 moieties in amyloid fibrils or protofibrils are more similar in structure to those in the 0.94 MDa particles than those in the dimers/tetramers. These findings indicate that soluble oligomeric forms of Abeta peptides can be trapped for extended periods of time, enabling their study by high resolution techniques that would not otherwise be possible. PMID- 10704309 TI - Identification of natural ligands for SH2 domains from a phage display cDNA library. AB - The cytoplasmic domain of the Fc gamma receptor IIB (FcgammaRIIB) can be successfully displayed on the surface of filamentous phage, and after phosphorylation in vitro, can interact specifically with the SH2 domains of SHP 2, a cytoplasmic tyrosine phosphatase. When full-length FcgammaRIIB is expressed on phage, however, this interaction is greatly compromised, illustrating that characteristics of the full-length sequence are not well tolerated by the phage display system. Many associations in cell physiology are driven by similar interactions involving small modular binding domains or ligands, and so a fragmented cDNA library will facilitate display of such domains free of sequences which compromise their expression. A fragmented leukocyte cDNA display library of 10(8) clones was constructed. This library was phosphorylated in vitro with fyn kinase and was selected against the tandem SH2 domains of SHP-2 in the search for additional ligands. A depletion strategy to remove non-specific clones was employed, using SHP-2 Sepharose, prior to in vitro phosphorylation and selection. This permitted the emergence of clones encoding the cytoplasmic domain of PECAM 1, another natural ligand for SHP-2. The importance of dual phosphorylation of tyrosine residues at positions 663 and 686 was confirmed in competition ELISA experiments using phosphorylated phage and synthetic peptides. Thus, phage display of fragmented cDNA libraries permits the identification and characterisation of phosphorylated ligands of modular binding domains based on their functional interaction. PMID- 10704310 TI - Pulse-chase analysis of the in vivo assembly of the bacteriophage T4 tail. AB - The in vivo assembly pathway of the complex tail of bacteriophage T4 virus was determined using pulse-chase analysis as a non-invasive alternative to the in vitro experiments previously used to map assembly. Bacteriophage T4 mutants defective in head assembly were used to infect cultures of Escherichia coli in order to study tail assembly in isolation. Beginning with the onset of late protein synthesis, the cultures were labeled continuously with [(3)H]leucine to normalize against subsequent sample losses. After completed tails had begun to accumulate at a constant rate, the cultures were pulsed with [(35)S]methionine, and then chased. Completed tails were purified at one minute intervals for the next 30 minutes and their proteins separated electrophoretically and counted by liquid scintillation. Total (35)S incorporation into each protein rose and then leveled off as the chase of unlabeled methionine flushed the label through the pools of soluble proteins and assembly intermediates and into completed tails. The inflection point in the sigmoidal (35)S-incorporation curve of each protein marks the maximal uptake of (35)S within that pool just before the effect of the chase becomes apparent and the curve begins to level off. The length of the delay in the apparent chase time reflects the position of that protein in the pathway. The closer the assembly point to the end of the pathway, the sooner the chase appears, revealing the relative order of assembly. As predicted, tail completion proteins such as gp18 (tail sheath) and 19 (tail tube) show the earliest inflection, while those earlier in the pathway take longer to chase. Of the 17 tail proteins analyzed, 14 are in agreement with the established in vitro pathway. The other three, gp15, gp10 and gp53, have helped us to develop a model that offers a plausible explanation for their altered chase times. PMID- 10704311 TI - Conformation of the RNA polymerase II C-terminal domain: circular dichroism of long and short fragments. AB - The C-terminal domain (CTD) of the largest subunit of RNA polymerase II consists of tandemly repeated copies of a heptapeptide with the Y(1)S(2)P(3)T(4)S(5)P(6)S(7) consensus sequence. This repeat contains two overlapping SPXX motifs that can adopt a beta-turn conformation. In addition, each CTD repeat contains the PXXP sequence characteristic of the left-handed helix of polyproline II (P(II)) found in SH3 domain ligands and the PXY sequence that is the target for WW domains. We have studied CTD fragments using circular dichroism (CD) to characterize the conformation of the CTD in water and in the hydrogen bond-promoting solvent trifluoroethanol (TFE). In water, an eight-repeat fragment is predominantly unordered, but at 32 degrees C has P(II) and beta-turn contents estimated to be about 15 % and less than 10 %, respectively. In 90 % TFE, the beta-turn fraction is estimated to be about 75 %, the remainder being unordered and P(II) conformations. The Tyr side-chains are ordered to a significant extent in 90 % TFE. Replacement of the fully conserved Pro residues by alpha-aminoisobutyric acid leads to a large increase in beta-turn. Replacement of Ser2 by Ala does not substantially alter the CTD conformation in water or TFE. Ser5 replacement by Ala increases the P(II) content in water and affects the conformation in TFE-rich solutions. Phosphorylation of Ser2 and Ser5 has little effect in water, but Ser2 affects the conformation in TFE-rich solution in much the same way as Ser5-->Ala substitution. The CD of the full-length murine CTD in water is similar to that of the eight-repeat fragment, indicating little difference in conformation with increasing chain length beyond eight repeats. The roles of P(II) and beta-turn in the interaction of CTD with its target proteins (mediator and RNA-processing components) are discussed. The most likely interactions are between P(II) and WW or SH3 domains, or with some unknown P(II) binding motif. PMID- 10704312 TI - Gene sequence and crystal structure of the aldehyde oxidoreductase from Desulfovibrio desulfuricans ATCC 27774. AB - The aldehyde oxidoreductase (MOD) isolated from the sulfate reducer Desulfovibrio desulfuricans (ATCC 27774) is a member of the xanthine oxidase family of molybdenum-containing enzymes. It has substrate specificity similar to that of the homologous enzyme from Desulfovibrio gigas (MOP) and the primary sequences from both enzymes show 68 % identity. The enzyme was crystallized in space group P6(1)22, with unit cell dimensions of a=b=156.4 A and c=177.1 A, and diffraction data were obtained to beyond 2.8 A. The crystal structure was solved by Patterson search techniques using the coordinates of the D. gigas enzyme. The overall fold of the D. desulfuricans enzyme is very similar to MOP and the few differences are mapped to exposed regions of the molecule. This is reflected in the electrostatic potential surfaces of both homologous enzymes, one exception being the surface potential in a region identifiable as the putative docking site of the physiological electron acceptor. Other essential features of the MOP structure, such as residues of the active-site cavity, are basically conserved in MOD. Two mutations are located in the pocket bearing a chain of catalytically relevant water molecules. As deduced from this work, both these enzymes are very closely related in terms of their sequences as well as 3D structures. The comparison allowed confirmation and establishment of features that are essential for their function; namely, conserved residues in the active-site, catalytically relevant water molecules and recognition of the physiological electron acceptor docking site. PMID- 10704313 TI - Structure-fluorescence correlations in a single tryptophan mutant of carp parvalbumin: solution structure, backbone and side-chain dynamics. AB - Heterogeneous fluorescence intensity decays of tryptophan in proteins are often rationalized using a model which proposes that different rotameric states of the indole alanyl side-chain are responsible for the observed fluorescence lifetime heterogeneity. We present here the study of a mutant of carp parvalbumin bearing a single tryptophan residue at position 102 (F102W) whose fluorescence intensity decay is heterogeneous and assess the applicability of a rotamer model to describe the fluorescence decay data. We have determined the solution structure of F102W in the calcium ligated state using multi-dimensional nuclear magnetic resonance (NMR) and have used the minimum perturbation mapping technique to explore the possible existence of multiple conformations of the indole moiety of Trp102 of F102W and, for comparison, Trp48 of holo-azurin. The maps for parvalbumin suggest two potential conformations of the indole side-chain. The high energy barrier for rotational isomerization between these conformers implies that interwell rotation would occur on time-scales of milliseconds or greater and suggests a rotamer basis for the heterogeneous fluorescence. However, the absence of alternate Trp102 conformers in the NMR data (to within 3 % of the dominant species) suggests that the heterogeneous fluorescence of Trp102 may arise from mechanisms independent of rotameric states of the Trp side-chain. The map for holo-azurin has only one conformation, and suggests a rotamer model may not be required to explain its heterogeneous fluorescence intensity decay. The backbone and Trp102 side-chain dynamics at 30 degrees C of F102W has been characterized based on an analysis of (15)N NMR relaxation data which we have interpreted using the Lipari-Szabo formalism. High order parameter (S(2)) values were obtained for both the helical and loop regions. Additionally, the S(2) values imply that the calcium binding CD and EF loops are not strictly equivalent. The S(2) value for the indole side-chain of Trp102 obtained from the fluorescence, NMR relaxation and minimum perturbation data are consistent with a Trp moiety whose motion is restricted. PMID- 10704314 TI - The folding of an immunoglobulin-like Greek key protein is defined by a common core nucleus and regions constrained by topology. AB - TNfn3, the third fibronectin type III domain of human tenascin, is an immunoglobulin-like protein that is a good model for experimental and theoretical analyses of Greek key folding. The third fibronectin type III domain of human tenascin folds and unfolds in a two-state fashion over a range of temperature and pH values, and in the presence of stabilising salts. Here, we present a high resolution protein engineering analysis of the single rate determining transition state. The 48 mutations report on the contribution of side-chains at 32 sites in the core and loop regions. Three areas in the protein exhibit high Phi-values, indicating that they are partially structured in the transition state. First, a common-core ring of four positions in the central strands B, C, E and F, that are in close contact, form a nucleus of tertiary interactions. The two other regions that appear well-formed are the C' region and the E-F loop. The Phi-values gradually decrease away from these regions such that the very ends of the two terminal strands A and G, have Phi-values of zero. We propose a model for the folding of immunoglobulin-like proteins in which the common-core "ring" forms the nucleus for folding, whilst the C' and E-F regions are constrained by topology to pack early. Folding characteristics of a group of structurally related proteins appear to support this model. PMID- 10704315 TI - Molecular clocks reduce plasmid loss rates: the R1 case. AB - Plasmids control their replication so that the replication frequency per plasmid copy responds to the number of plasmid copies per cell. High sensitivity amplification in replication response to copy number deviations generally reduces variation in copy numbers between different single cells, thereby reducing the plasmid loss rate in a cell population. However, experiments show that plasmid R1 has a gradual, insensitive replication control predicting considerable copy number variation between single cells. The critical step in R1 copy number control is regulation of synthesis of a rate-limiting cis-acting replication protein, RepA. De novo synthesis of a large number of RepA molecules is required for replication, suggesting that copy number control is exercised at multiple steps. In this theoretical kinetic study we analyse R1 multistep copy number control and show that it results in the insensitive replication response found experimentally but that it at the same time effectively prohibits the existence of only one plasmid copy in a dividing cell. In combination with the partition system of R1, this can lead to very high segregational stability. The R1 control mechanism is compared to the different multistep copy number control of plasmid ColE1 that is based on conventional sensitivity amplification. This implies that while copy number control for ColE1 efficiently corrects for fluctuations that have already occurred, R1 copy number control prevents their emergence in cells that by chance start their cycle with only one plasmid copy. We also discuss how regular, clock-like, behaviour of single plasmid copies becomes hidden in experiments probing collective properties of a population of plasmid copies because the individual copies are out of phase. The model is formulated using master equations, taking a stochastic approach to regulation, but the mathematical formalism is kept to a minimum and the model is simplified to its bare essence. This simplicity makes it possible to extend the analysis to other replicons with similar design principles. PMID- 10704316 TI - Thermal unfolding of an intermediate is associated with non-Arrhenius kinetics in the folding of hen lysozyme. AB - A variety of techniques, including quenched-flow hydrogen exchange labelling monitored by electrospray ionization mass spectrometry, and stopped-flow absorbance, fluorescence and circular dichroism spectroscopy, has been used to investigate the refolding kinetics of hen lysozyme over a temperature range from 2 degrees C to 50 degrees C. Simple Arrhenius behaviour is not observed, and although the overall rate of folding increases from 2 to 40 degrees C, it decreases above 40 degrees C. In addition, the transient intermediate on the major folding pathway at 20 degrees C, in which the alpha-domain is persistently structured in the absence of a stable beta-domain, is thermally unfolded in a sigmoidal transition (T(m) approximately 40 degrees C) indicative of a cooperatively folded state. At all temperatures, however, there is evidence for fast ( approximately 25 %) and slow ( approximately 75 %) populations of refolding molecules. By using transition state theory, the kinetic data from various experiments were jointly fitted to a sequential three-state model for the slow folding pathway. Together with previous findings, these results indicate that the alpha-domain intermediate is a productive species on the folding route between the denatured and native states, and which accumulates as a consequence of its intrinsic stability. Our analysis suggests that the temperature dependence of the rate constant for lysozyme folding depends on both the total change in the heat capacity between the ground and transition states (the dominant factor at low temperatures) and the heat-induced destabilization of the alpha-domain intermediate (the dominant factor at high temperatures). Destabilization of such kinetically competent intermediate species is likely to be a determining factor in the non-Arrhenius temperature dependence of the folding rate of those proteins for which one or more intermediates are populated. PMID- 10704317 TI - A reassessment of the factors affecting microtubule assembly and disassembly in vitro. AB - Current models of microtubule assembly from pure tubulin involve a nucleation phase followed by microtubule elongation at a constant polymer number. Both the rate of microtubule nucleation and elongation are thought to be tightly influenced by the free GTP-tubulin concentration, in a law of mass action dependent manner. However, these basic hypotheses have remained largely untested due to a lack of data reporting actual measurements of the microtubule length and number concentration during microtubule assembly.Here, we performed simultaneous measurements of the polymeric tubulin concentration, of the free GTP-tubulin concentration, and of the microtubule length and number concentration in both polymerizing and depolymerizing conditions. In agreement with previous work we find that the microtubule nucleation rate is strongly dependent on the initial GTP-tubulin concentration. But we find that microtubule nucleation persists during microtubule elongation. At any given initial tubulin-GTP concentration, the microtubule nucleation rate remains constant during polymer assembly, despite the wide variation in free GTP-tubulin concentration. We also find a remarkable constancy of the rate of microtubule elongation during assembly. Apparently, the rate of microtubule elongation is intrinsic to the polymers, insensitive to large variations of the free GTP-tubulin concentration. Finally we observe that when, following assembly, microtubules depolymerize below the free GTP-tubulin critical concentration, the rate-limiting factor for disassembly is the frequency of microtubule catastrophe. At all time-points during disassembly, the microtubule catastrophe frequency is independent of the free GTP-tubulin concentration but, as the microtubule nucleation rate, is strongly dependent on the initial free GTP tubulin concentration. We conclude that the dynamics of both microtubule assembly and disassembly depend largely on factors other than the free GTP-tubulin concentration. We propose that intrinsic structural factors and endogenous regulators, whose concentration varies with the initial conditions, are also major determinants of these dynamics. PMID- 10704318 TI - Structural and mechanistic basis of porphyrin metallation by ferrochelatase. AB - Ferrochelatase, the enzyme catalyzing metallation of protoporphyrin IX at the terminal step of heme biosynthesis, was co-crystallized with an isomer mixture of the potent inhibitor N-methylmesoporphyrin (N-MeMP). The X-ray structure revealed the active site of the enzyme, to which only one of the isomers was bound, and for the first time allowed characterization of the mode of porphyrin macrocycle distortion by ferrochelatase. Crystallization of ferrochelatase and N-MeMP in the presence of Cu(2+) leads to metallation and demethylation of N-MeMP. A mechanism of porphyrin distortion is proposed, which assumes that the enzyme holds pyrrole rings B, C and D in a vice-like grip and forces a 36 degrees tilt on ring A. PMID- 10704319 TI - Assessing annotation transfer for genomics: quantifying the relations between protein sequence, structure and function through traditional and probabilistic scores. AB - Measuring in a quantitative, statistical sense the degree to which structural and functional information can be "transferred" between pairs of related protein sequences at various levels of similarity is an essential prerequisite for robust genome annotation. To this end, we performed pairwise sequence, structure and function comparisons on approximately 30,000 pairs of protein domains with known structure and function. Our domain pairs, which are constructed according to the SCOP fold classification, range in similarity from just sharing a fold, to being nearly identical. Our results show that traditional scores for sequence and structure similarity have the same basic exponential relationship as observed previously, with structural divergence, measured in RMS, being exponentially related to sequence divergence, measured in percent identity. However, as the scale of our survey is much larger than any previous investigations, our results have greater statistical weight and precision. We have been able to express the relationship of sequence and structure similarity using more "modern scores," such as Smith-Waterman alignment scores and probabilistic P-values for both sequence and structure comparison. These modern scores address some of the problems with traditional scores, such as determining a conserved core and correcting for length dependency; they enable us to phrase the sequence-structure relationship in more precise and accurate terms. We found that the basic exponential sequence-structure relationship is very general: the same essential relationship is found in the different secondary-structure classes and is evident in all the scoring schemes. To relate function to sequence and structure we assigned various levels of functional similarity to the domain pairs, based on a simple functional classification scheme. This scheme was constructed by combining and augmenting annotations in the enzyme and fly functional classifications and comparing subsets of these to the Escherichia coli and yeast classifications. We found sigmoidal relationships between similarity in function and sequence, with clear thresholds for different levels of functional conservation. For pairs of domains that share the same fold, precise function appears to be conserved down to approximately 40 % sequence identity, whereas broad functional class is conserved to approximately 25 %. Interestingly, percent identity is more effective at quantifying functional conservation than the more modern scores (e.g. P-values). Results of all the pairwise comparisons and our combined functional classification scheme for protein structures can be accessed from a web database at http://bioinfo.mbb.yale.edu/alignCopyright 2000 Academic Press. PMID- 10704320 TI - Experimental and theoretical studies of the effects of deoxyribose substitutions on the stability of the UUCG tetraloop. AB - Experimental and theoretical thermodynamic studies of the consequences of 2' hydroxyl substitution in the RNA UUCG tetraloop show distinct position dependence consistent with the diverse structural contexts of the four-loop ribose hydroxyls in this motif. The results suggest that even for simple substitutions, such as the replacement of the ribose hydroxyl (2'-OH) with hydrogen (2'-H), the free energy change reflects a complex interplay of hydrogen bonding and solvation effects and is influenced by the intrinsic pucker preferences of the nucleotides. Furthermore, theoretical studies suggest that the effect of these mutations in the single-strand state is sequence dependent, in contrast to what is commonly assumed. Free energy perturbation simulations of ribose-deoxyribose mutations in a single-strand dodecamer and in trinucleotide models suggest that in the denatured state, the magnitude of the free energy change for deoxyribose substitutions is determined to a larger extent by the identity of the nucleotide (A, C, G or U) rather than its structural context. Single-strand mutational effects must be considered when interpreting mutational studies in molecular terms. PMID- 10704321 TI - Evidence for the involvement of two areas of the zebra finch forebrain in sexual imprinting. AB - Sexual imprinting in male zebra finches is a two-step process, including an acquisition period early in life and a stabilization process normally occuring during the first courtship attempts of the male. During the acquisition period, a young male learns about its social environment. During stabilization, which can be delayed experimentally until day 100, it develops a preference for the appropriate object for courtship behavior on the basis of its previous and acute experience. Thereafter, this preference cannot be altered again. Exploring the physiological basis for imprinting, we have previously shown that the neurons of two forebrain areas (ANC and HAD) increase their spine density in the course of the stabilization process, while in two other areas (MNH and LNH) a decrease of spine density can be observed. With the present experiments, we tested the idea that the spine density decrease in MNH and LNH is the anatomical manifestation of the imprinting process. Previous behavioral experiments have shown that exposure to a nestbox after 100 days of age stabilizes the sexual preference of a zebra finch male as well as does exposure to a female. The present study shows that nestbox exposure also reduces the spine density in MNH and LNH, but has no effect on ANC and HAD. It has also been shown previously that treating males with an antiandrogen between days 40 and 100 affects the final preference of a male. The present experiment indicates that the same treatment affects spine growth during development in MNH and LNH and prevents the increase of spine density within HAD and ANC normally induced by exposure to a female. The results are interpreted as strong evidence for the involvement of MNH and LNH in sexual imprinting. PMID- 10704322 TI - Development of neuronal responsiveness in the mediorostral neostriatum/hyperstriatum ventrale during auditory filial imprinting in domestic chicks. AB - Chronic electrophysiological recordings of slow field potentials from tone imprinted chicks show significantly enhanced fast Fourier transform (FFT) power during playback of rhythmic 400-Hz imprinting tone stimuli in the presence of a surrogate mother. The FFT power was already significantly higher during the very first imprinting session, when the chick was exposed to the imprinting tone stimuli in the presence of the surrogate mother compared to spontaneous activity (EEG recordings). During discrimination tests, where individual chicks were exposed to the imprinting tone stimuli in alternation to rhythmic 700-Hz tone stimuli (discrimination tone stimuli), the FFT power was significantly higher during playback of the imprinting tone stimuli than the FFT power during playback of the discrimination tone stimuli. Chicks which were imprinted in the absence of the surrogate mother also show enhanced FFT power in the course of the imprinting sessions; however, in contrast to the first group, they did not show significant differences in the FFT power during playback of either the imprinting or discrimination tone stimuli in the discrimination tests. Our results suggest that the high FFT power of a potential imprinting stimulus or situation, which is expressed in newborn (still naive) chicks, is maintained only when the chicks form an association between the tone stimuli and a positive emotional situation (represented by the surrogate mother). PMID- 10704323 TI - Posttraining inactivation of excitatory afferent input to the locus coeruleus impairs retention in an inhibitory avoidance learning task. AB - These experiments examined whether the nucleus paragigantocellularis (PGi) contributes to memory storage processing via its ascending excitatory influence on locus coeruleus (LC) neuronal activity. Activation of the LC leads to memory enhancement and also results in a widespread release of norepinephrine in target structures, such as the amygdala and hippocampus. Infusion of norepinephrine into either structure also improves memory for several types of learned responses. Thus, the capacity for norepinephrine to modulate memory within limbic structures may be contingent upon the functional connections between PGi and the LC. To examine this hypothesis, male Sprague-Dawley rats were implanted with cannula aimed above PGi (Experiments 1 and 2) or 1.5 mm dorsal or medial to PGi (Experiment 3). Immediately following inhibitory avoidance training (0.45 mA, 0. 5 s), phosphate-buffered saline, lidocaine (Experiment 1), or 12.5 or 25 nmol/0.5 microl of the GABA agonist muscimol (Experiment 2) was infused into PGi. On a retention test given 48 h later, the latency to reenter the footshock compartment was significantly shorter for subjects given either lidocaine or 12.5 or 25.0 nmol of muscimol compared to controls. In Experiment 3, infusion of lidocaine or muscimol into areas 1.5 mm dorsal or medial to PGi did not significantly alter retention, indicating that the memory impairment observed in Experiments 1 and 2 was site specific and not due to the spread of drug to cell groups surrounding PGi. These findings suggest that PGi may serve a vital function in relaying biologically relevant information to forebrain structures involved in memory via its excitatory influence on the LC. PMID- 10704325 TI - The effect of cholinergic, GABAergic, serotonergic, and glutamatergic receptor modulation on posttrial memory processing in the hippocampus. AB - Though the hippocampus is widely recognized as important in learning and memory, most of the evidence for this comes from animal lesion and human pathological studies. Due to the relatively small number of drugs that have been tested in the hippocampus for their ability to alter posttrial memory processing, there is a general impression that memory processing involves only a few neurotransmitters. We have evaluated the effects of cholinergic, GABAergic, serotonergic, and glutamatergic receptor agonists and antagonists for their ability to facilitate or impair retention. CD-1 mice received acute intrahippocampal drug infusion following footshock avoidance training in a T-maze. Retention was tested 1 week after training and drug administration. The results indicate that receptor agonists of acetylcholine and glutamate improved retention, while antagonists impaired retention. However, scopolamine did not impair retention, but M1 and M2 antagonists did. Receptor agonists of serotonin and GABA impaired retention, while antagonists improved retention. Drugs acting on 5-HT-1 and 5-HT-2 as well as GABA(A) and GABA(B) receptor subtypes did not differentially effect retention. PMID- 10704324 TI - Short- and long-term memory are differentially affected by metabolic inhibitors given into hippocampus and entorhinal cortex. AB - Rats were implanted with cannulae in the CA1 area of the dorsal hippocampus or in the entorhinal cortex and trained in one-trial step-down inhibitory avoidance. Two retention tests were carried out in each animal, one at 1.5 h to measure short-term memory (STM) and another at 24 h to measure long-term memory (LTM). The purpose of the present study was to screen the effect on STM of various drugs previously shown to affect LTM of this task when given posttraining at the same doses that were used here. The drugs and doses were the guanylyl cyclase inhibitor LY83583 (LY, 2.5 microMg), the inhibitor of Tyr-protein kinase at low concentrations and of protein kinase G (PKG) at higher concentrations lavendustin A (LAV, 0.1 and 0.5 microMg), the PKG inhibitor KT5823 (2.0 microMg), the protein kinase C (PKC) inhibitor staurosporin (STAU, 2.5 microMg), the inhibitor of calcium/ calmodulin protein kinase II (CaMKII) KN62 (3.6 microMg), the protein kinase A (PKA) inhibitor KT5720 (0.5 microMg), and the mitogen-activated protein kinase kinase (MAPKK) inhibitor PD098059 (PD, 0.05 microMg). PD was dissolved in saline; all the other drugs were dissolved in 20% dimethyl sulfoxide. In all cases the drugs affected LTM as had been described in previous papers. The drugs affected STM and LTM differentially depending on the brain structure into which they were infused. STM was inhibited by KT5720, LY, and PD given into CA1 and by STAU and KT5720 given into the entorhinal cortex. PD given into the entorhinal cortex enhanced STM. LTM was inhibited by STAU, KN62, KT5720, KT5823, and LAV (0.5 microMg) given into CA1 and by STAU, KT5720, and PD given into the entorhinal cortex. The results suggest that STM and LTM involve different physiological mechanisms but are to an extent linked. STM appears to require PKA, guanylyl cyclase, and MAPKK activity in CA1 and PKA and PKC activity in the entorhinal cortex; MAPKK seems to play an inhibitory role in STM in the entorhinal cortex. In contrast, LTM appears to require PKA and PKC activity in both structures, guanylyl cyclase, PKG, and CaMKII activity in CA1, and MAPKK activity in the entorhinal cortex. PMID- 10704326 TI - Effects of intraseptal zolpidem and chlordiazepoxide on spatial working memory and high-affinity choline uptake in the hippocampus. AB - Injection of GABA(A)/benzodiazepine receptor ligands into the medial septum (MS) alters the activity of cholinergic neurons that innervate the hippocampus and can produce bidirectional modulation of spatial memory. Recent evidence suggests that two subtypes of the GABA(A) receptor are differentially localized to either GABAergic (alpha(1)/beta(2)/gamma(2)) or cholinergic (alpha(3)/beta(3)/gamma(2)) neurons within the MS. The present studies characterized the dose-related behavioral and neurochemical effects of intraseptal infusions of two benzodiazepine (BDZ) agonists that appear to exhibit different profiles of pharmacological specificity for these receptor subtypes. Male Sprague-Dawley rats were cannulated and then artificial CSF, chlordiazepoxide (CDP: 8 or 12 microg), or zolpidem (4, 8, or 12 microg) was injected into the MS. Spatial working memory was assessed in a delay radial-arm maze task and the activity of cholinergic neurons in the MS was evaluated by high-affinity choline uptake (HA-ChU) in the hippocampus. Intraseptal injection of either CDP or zolpidem produced dose related impairments in spatial working memory and decreases in hippocampal HAChU. Both BDZ agonists were found to produce retrograde memory deficits and a decrease in HAChU following the highest dose tested (12 microg). However, intraseptal injection of 8 microg of zolpidem produced a behavioral deficit comparable to the high dose of CDP, but did not alter HAChU within the HPC. Although the cholinergic component of the septohippocampal pathway has been shown to be important in modulating hippocampal physiology and spatial memory processes, data from the present experiments suggest that the GABAergic component may also play an important role in the behavioral functions of the septohippocampal pathway. PMID- 10704328 TI - Strain-dependent interactions between MK-801 and cocaine on retention of C57BL/6 and DBA/2 mice tested in a one-trial inhibitory avoidance task: involvement of dopaminergic mechanisms. AB - Two sets of experiments were carried out with C57BL/6 (C57) and DBA/2 (DBA) mice tested in a one-trial inhibitory avoidance task. In the first set C57 and DBA mice were injected posttraining with saline or with the D1 DA receptor antagonist SCH 23390 and then with saline, cocaine (5 mg/kg), MK-801 (0.1 mg/kg), or with a combination of these two drugs. Cocaine enhanced retention in the C57 strain and impaired it in the DBA strain, and MK-801 potentiated the effects of cocaine in both strains. Furthermore, pretreatment with SCH 23390 completely antagonized the potentiation of the effects of cocaine exerted by MK-801. In the second set of experiments mice belonging to these same two strains were injected posttraining with vehicle or with the D2 DA receptor antagonist (-)-sulpiride and then with saline, cocaine (5 mg/kg), MK-801 (0.1 mg/kg), or with a combination of these two drugs. Pretreatment with the D2 DA receptor antagonist completely antagonized in both strains the potentiation of the effect of cocaine exerted by MK-801. The results of the present research show that the noncompetitive NMDA receptor antagonist MK-801 enhances the effect of cocaine on retention performance in C57 and DBA mice and that dopaminergic mechanisms are involved in this potentiation. PMID- 10704327 TI - Estradiol enhances the induction of homosynaptic long-term depression in the CA1 region of the adult, ovariectomized rat. AB - An ovarian steroid-dependent cycle of synaptogenesis and synapse shedding occurs naturally in the hippocampus of the adult female rat. The newly formed axospinous synapses in CA1 may differ functionally from extant axospinous synapses, e.g., in terms of their modifiability. Here we assess whether estradiol alters the induction of homosynaptic long-term depression of the Schaffer collateral-CA1 synapses in vitro. Sprague-Dawley rats were bilaterally ovariectomized and, beginning 6-8 days later, received a series of injections of either 17beta estradiol or sesame oil sc. Field potentials were recorded in hippocampal slices. In estradiol-treated animals, asynchronous, low-frequency stimulation led to significant long-term depression of the activated synapses in CA1 s. radiatum and no change of the inactive synapses in s. oriens. In contrast, this conditioning stimulation did not significantly alter any CA1 responses in oil-treated control animals. Subsequent high-frequency conditioning stimulation significantly potentiated the activated s. radiatum synapses in both estradiol- and oil-treated animals. Thus, given the stimulation conditions used here, estradiol enables the induction of homosynaptic long-term depression at the CA3-CA1 synapses in adult females. PMID- 10704329 TI - HIV and pandemic influenza virus: two great infectious disease challenges. PMID- 10704330 TI - Eradication of polio by vaccination. PMID- 10704331 TI - Enhancing DNA immunization. PMID- 10704332 TI - Detection of an RNA-dependent RNA polymerase in mitochondria from a mitovirus infected isolate of the Dutch Elm disease fungus, Ophiostoma novo-ulmi. AB - RNA-dependent RNA polymerase (RdRp) activity has been detected in mitochondria from an isolate of Ophiostoma novo-ulmi infected with O. novo-ulmi mitovirus 6 (OnuMV6). The reaction products corresponded to the double-stranded and single stranded forms of OnuMV6 RNA. Western blot analysis using antibodies raised against a conserved RdRp region has detected a protein of ca. 80 kDa in OnuMV6 infected mitochondria, close to the predicted size of the OnuMV6 RdRp. No RdRp activity or protein was detected in mitochondria from an uninfected O. novo-ulmi isolate. This is the first detection of a virus RdRp in fungal mitochondria and the results are consistent with the use of UGA tryptophan codons in its synthesis. PMID- 10704333 TI - Effective construction of DNA vaccines against variable influenza genes by homologous recombination. AB - We demonstrate the potential of cloning by homologous recombination as a rapid method to construct DNA molecules encoding newly developing hemagglutinins (HA) of influenza A virus. The variable parts of the HA genes were cloned into a basic construct containing the HA gene from an H3N2 strain. The recombinant DNAs thus created encode different variable domains with neutralising epitopes from four recently circulating influenza A H3 strains. The technology allows rapid production of DNA constructs for vaccines that can induce antibody and, particularly, cellular immune responses. These new constructs were also capable of conferring protection to challenge in mice. The technology may hence be a valuable tool for rapid adaptation of influenza vaccines to changes in the circulating influenza strains. PMID- 10704334 TI - Enhancement of the basal-level activity of HIV-1 long terminal repeat by HIV-1 nucleocapsid protein. AB - Two HIV-1 proteins, Tat and NCp7 (NC), have zinc finger-like structures. NC is a virion protein and has been shown to accumulate in the nucleus 8 h postinfection. Since transcription factors with zinc fingers assist the transcriptional activity of both RNA polymerases II and III, we examined the effect of NC on HIV-1 LTR directed gene expression. The HIV-1 NC binds to the HIV-1 LTR and results in a mobility shift in polyacrylamide gel electrophoresis. Competition assays with cold probes revealed that the binding of NC and formation of a DNA-protein complex could be prevented by the addition of excess unlabeled LTR self-probe, but not the HIV-1 V3 envelope gene. The DNase I footprint analysis showed that NC binds to six regions within HIV-1 LTR, four of which are near the transcription start site. The NC alone enhances LTR basal-level activity in RNA runoff experiments. When the general transcription factors (GTFs) were added in the assay, NC enhances NF-kappaB, Sp1, and TFIIB-induced HIV-1 LTR-directed RNA transcription. RNA transcription directed by the adenovirus major late promoter, however, is not significantly affected by NC in the cell-free system. Transient transfection of human T lymphocytes with the plasmids containing HIV-1 nc or gag showed enhancement of LTR-CAT activity. Moreover, transfection of HIV-1 provirus containing mutations in NC zinc-finger domains dramatically decreases the enhancement activity in human T cells, in which HIV-1 LTR is stably integrated into the cellular genome. These observations show that NC binds to HIV-1 LTR and cooperatively enhances GTFs and NF-kappaB induced HIV-1 LTR basal-level activity. NC may play the role of a nucleation protein, which binds to LTR and enhances basal-level transcription by recruiting cellular transcription factors to the HIV 1 promoter in competition with cellular promoters. PMID- 10704335 TI - Sequence requirements for translation initiation of Rhopalosiphum padi virus ORF2. AB - Rhopalosiphum padi virus (RhPV) is an aphid-infecting virus with a 10-kb ssRNA genome that contains two large open reading frames (ORFs). ORF1 and ORF2 encode the nonstructural and structural polyproteins, respectively. Both ORFs are preceded by noncoding regions of 500 nt that could function as internal ribosome entry segments (IRESes). The sequence for ORF2 lacks an obvious initiation codon, but an out-of-frame AUG codon is present that could translate ORF2 through a +1 frameshift. To investigate the mechanisms of translation initiation of ORF2, a series of point and deletion mutations were constructed and transcribed and translated in vitro. A bicistronic plasmid containing two copies of the RhPV intergenic region translated both ORFs efficiently, indicating that the region functioned as an IRES in vitro. Deletion analysis showed that the region required for activity of the IRES extended from 178 nt upstream and 6 nt downstream of the 5' border of ORF2. Changes in the out-of-frame AUG codon had little effect on translation initiation, but mutations that included the first and second codons of ORF2 or that disrupted a putative pseudoknot structure near the 3' end of the IRES failed to initiate protein synthesis. Sequence analysis of the in vitro synthesized proteins showed that the first amino acid of the polyprotein corresponded to the second codon of ORF2. These results show that in vitro the noncoding region upstream of RhPV ORF2 functions as an IRES that contains a pseudoknot that directs translation initiation at a non-AUG codon. If the in vitro synthesized proteins have not been processed by an aminopeptidase, translation is initiated at the second (GCA) codon of ORF2. PMID- 10704336 TI - Primate cytomegaloviruses encode and express an IL-10-like protein. AB - An open reading frame (ORF) with homology to interleukin-10 (IL-10) has been identified in rhesus cytomegalovirus (RhCMV). The IL-10-like protein is generated from a multispliced, polyadenylated early gene transcript encompassing part of the corresponding UL111A ORF of human CMV (HCMV). Immunological analyses confirm expression of the IL-10-like protein both in tissue culture and in RhCMV-infected rhesus macaques. Conserved ORFs were subsequently identified in human, baboon, and African green monkey CMV, and a fully processed transcript has been mapped in fibroblasts infected with the Towne strain of HCMV. The conservation of this previously unrecognized ORF suggests that the protein may play an essential role in primate CMV persistence and pathogenesis. PMID- 10704337 TI - Synthesis of viral DNA and late capsid protein L1 in parabasal spinous cell layers of naturally occurring benign warts infected with human papillomavirus type 1. AB - We investigated human papillomavirus type 1 (HPV1)-specific transcription, viral DNA replication, and viral protein expression in naturally occurring benign tumors by in situ hybridization, 5-bromodeoxyuridine (BrdU) incorporation, and immunohistochemistry and obtained results different from other HPV-infected benign tumors characterized so far. Moderate amounts of transcripts with a putative coding potential for E6/E7, E1, and E2 were demonstrated from the first subrabasal cell layer throughout the stratum spinosum and granulosum. In addition very large amounts of E4 and L1 transcripts were present in the same epithelial layers. This finding was substantiated by the demonstration of L1 and E4 protein already in the bottom-most spinous cell layer. Furthermore massive amplification of the viral DNA as measured by BrdU incorporation and different methods of in situ hybridization took place in the lowest 5 to 10 suprabasal cell layers. These findings are in contrast to the assumption that late gene expression and viral DNA synthesis are restricted to the more differentiated cell layers of the epithelium and point to differences in the regulation of the vegetative life cycle between different papillomavirus types. PMID- 10704339 TI - Phenotyping of Evi1, Evi11/Cb2, and Evi12 transformed leukemias isolated from a novel panel of cas-Br-M murine leukemia virus-infected mice. AB - Cas-Br-M murine leukemia virus (MuLV) is a slow-transforming retrovirus that potently induces leukemias in mice and therefore is well suited for retroviral insertional mutagenesis. We used Cas-Br-M MuLV in NIH/Swiss mice to establish a new panel of mainly myeloid leukemias. All tumors found in leukemic animals were classified by gross pathology, morphology, and immunophenotype, as well as the incidence of known common virus integration sites (VISs) in MuLV-induced myeloid malignancies (i.e., Evi1, Evi11/Cb2, Evi12, Fli1, and c-Myb). Interestingly, male mice were more susceptible than females to the induction of leukemia by Cas-Br-M MuLV. Seventy-four of the Cas-Br-M MuLV-inoculated mice developed a severe splenomegaly, sometimes in association with a thymoma. Although most of the immunophenotyped Cas-Br-M MuLV tumors were of myeloid origin (58%), numerous T cell leukemias (21%) and mixed myeloid/T-cell leukemias (21%) were found. The myeloid leukemias and myeloid compartment of the mixed leukemias were further characterized by immunophenotyping with stem cell-, myeloid-, and erythroid specific antibodies. The known Cas-Br-M MuLV common VISs (Evi1, Evi11/Cb2, and Evi12) were demonstrated in 19%, 12%, and 20% of the cases, respectively, whereas no Fli1 and c-Myb rearrangements were found. Integrations into Evi1 were restricted to myeloid leukemias, whereas those in Evi11/Cb2 and Evi12 were identified in myeloid as well as T-lymphoid leukemias. This panel of well characterized Cas-Br-M MuLV-induced hematopoietic tumors may be useful for the isolation and characterization of new proto-oncogenes involved in myeloid or T cell leukemias. PMID- 10704338 TI - Proline residues in the HIV-1 NH2-terminal capsid domain: structure determinants for proper core assembly and subsequent steps of early replication. AB - Recent analyses suggest that the p24 capsid (p24(CA)) domain of the HIV-1 group specific antigen (Gag) may be divided into two structurally and functionally distinct moieties: (i) an amino-terminal portion, previously shown to bind the cellular chaperone cyclophilin A, and (ii) a carboxy-terminal domain, known to contribute to the interaction of the Gag and Gag-Pol precursors during the early assembly process. In order to gain deeper insight into the role of the amino terminal domain of the p24(CA) protein during viral replication, eight highly conserved proline residues known to promote turns and to terminate alpha-helices within the p24 tertiary structure were replaced by a leucine residue (P-position L). Following transfection of the proviral constructs in COS7 cells, the majority of the mutants resembled wild-type viruses with respect to the assembly and release of virions. However, although the released particles contained wild-type levels of genomic viral RNA, the mature products of the Gag and Gag-Pol polyproteins as well as the Env glycoproteins-all of them, except mutant P225L were either noninfectious or severely affected in their replicative capacity. Entry assays monitoring the process of viral DNA synthesis led to the classification of selected provirus mutants into four different phenotypes: (i) mutant P225L was infectious and allowed complete reverse transcription including formation of 2-LTR circles; (ii) mutants P149L, P170L, and P217L failed to form 2 LTR circles; (iii) mutant P222L displayed a severe defect in binding and incorporating cyclophilin A into virions, was delayed with respect to DNA polymerization, and failed to form a 2-LTR replication intermediate; and (iv) mutant P133L was unable even to synthesize a first-strand cDNA product. All replication-defective mutants were characterized by severe alterations in the stability of virion cores, which were in two cases reflected by visible changes in the core morphology. These results suggest that proline residues in the NH(2) terminal capsid domain represent critical structure determinants for proper formation of functional virion cores and subsequent stages of early replication. PMID- 10704340 TI - Association of histone deacetylase with COUP-TF in tumorigenic Ad12-transformed cells and its potential role in shut-off of MHC class I transcription. AB - Chicken ovalbumin upstream promoter-transcription factor (COUP-TF) is an orphan nuclear receptor that represses transcription of many genes. In adenovirus type 12 (Ad12) transformed cells, a high level of binding activity of COUP-TF to the major histocompatibility complex (MHC) class I enhancer correlates with the down regulation of class I transcription, which, in turn, contributes to tumorigenesis. The mechanism by which COUP-TF represses transcription has yet to be elucidated. Here we show that COUP-TF represses transcription through its association with histone deacetylase. This was demonstrated using reciprocal binding assays that determined that the interaction between COUP-TF and histone deacetylase requires the COUP-TF C-terminal repression domain. Moreover, a histone deacetylase enzymatic activity was found to be associated with COUP-TF in Ad12-transformed cells. Transfection experiments further revealed that exogenous histone deacetylase facilitates transcriptional repression by COUP-TF. Also, supershift assays suggest that the transcriptional corepressor N-CoR, which is known to associate with histone deacetylases, is a part of the COUP-TF complex bound to the MHC class I enhancer R2 site. Finally, we provide evidence that inhibition of histone deacetylases relieves the repression of MHC class I expression in Ad12-transformed cells. Taken together these results support the notion that deacetylation of histones, mediated through COUP-TF, serves to down regulate MHC class I transcription in Ad12-transformed cells. PMID- 10704341 TI - Caspase-dependent apoptosis of cells expressing the chemokine receptor CXCR4 is induced by cell membrane-associated human immunodeficiency virus type 1 envelope glycoprotein (gp120). AB - Human immunodeficiency virus type 1 (HIV-1) envelope glycoproteins interact with CD4 and chemokine receptors on T cells to deliver signals that trigger either activation, anergy, or apoptosis. However, the molecular mechanisms driving these responses remain poorly understood. In this study we demonstrate that apoptosis is induced upon HIV-1 envelope binding to the chemokine receptor CXCR4. Cells expressing a mutant form of CXCR4 with a C-terminal deletion were also sensitive to HIV-1 envelope-mediated apoptosis, indicating that the cytoplasmic tail of CXCR4 is not required to induce the apoptotic pathway. The specificity of this process was analyzed using several inhibitors of gp120-CD4-CXCR4 interaction. Monoclonal antibodies directed against the gp120-binding site on CD4 (ST4) and against CXCR4 (MAB173) prevented the apoptotic signal in a dose-dependent manner. The cell death program was also inhibited by SDF-1alpha, the natural ligand of CXCR4, and by suramin, a G protein inhibitor that binds with a high affinity to the V3 loop of HIV-1 gp120 envelope protein. These results highlight the role played by gp120-binding on CXCR4 to trigger programmed cell death. Next, we investigated the intracellular signal involved in gp120-induced apoptosis. This cell death program was insensitive to pertussis toxin and did not involve activation of the stress- and apoptosis-related MAP kinases p38(MAPK) and SAPK/JNK but was inhibited by a broad spectrum caspase inhibitor (z-VAD.fmk) and a relatively selective inhibitor of caspase 3 (z-DEVD.fmk). Altogether, our results demonstrate that HIV induces a caspase-dependent apoptotic signaling pathway through CXCR4. PMID- 10704342 TI - Analysis of cis-acting sequences involved in plus-strand synthesis of a turnip crinkle virus-associated satellite RNA identifies a new carmovirus replication element. AB - Satellite RNA C (satC) is a 356-base subviral RNA associated with turnip crinkle virus (TCV). A 3'-proximal element (3'-UCCCAAAGUAU) located 11 bases from the 3' terminus of satC minus strands can function as an independent promoter in an in vitro RNA-dependent RNA polymerase (RdRp) transcription system. Furthermore, in the absence of a 5'-proximal element, the 3'-proximal element is required for complementary strand synthesis in vitro. Site-directed mutagenesis was conducted to investigate the functional significance of this element and the 3' minus strand terminal sequence "3'-OH-CCCUAU," which contains the minus-strand 3'-end sequence "3'-OH-CC(1-2)(A/U)(A/U)(A/U)" found in all carmovirus RNAs. Single mutations in the 3'-terminal sequence, which we have named the carmovirus consensus sequence (CCS), suppressed satC plus-strand synthesis to undetectable levels in protoplasts while still permitting some minus-strand synthesis. However, single and multiple mutations introduced into the 3'-proximal element had little or no effect on satC accumulation in protoplasts. In vivo genetic selection (SELEX) of the minus-strand 3'-terminal 21 bases revealed that all satC species accumulating in plants contained the 3' CCS. In addition, the 3'-proximal element preferentially contained a sequence similar to the CCS and/or polypurines, suggesting that this element may also contribute to accumulation of satC in vivo. PMID- 10704343 TI - Requirement of a 5'-proximal linear sequence on minus strands for plus-strand synthesis of a satellite RNA associated with turnip crinkle virus. AB - Viral RNA replication begins with specific recognition of cis-acting RNA elements by the viral RNA-dependent RNA polymerase (RdRp) and/or associated host factors. A short RNA element (3'-AACCCCUGGGAGGC) located 41 bases from the 5' end of minus strands of satellite RNA C (satC), a 356-base subviral RNA naturally associated with turnip crinkle virus (TCV), was previously identified as important for plus strand synthesis using an in vitro RdRp assay (H. Guan, C. Song, A. E. Simon, 1997, RNA 3, 1401-1412). To examine the functional significance of this element in RNA replication, mutations were introduced into the consecutive C residues in the element. A single mutation of the 3'-most C residue resulted in undetectable levels of satC plus strands when transcripts were assayed in protoplasts and suppressed transcription directed by the element in vitro. However, satC minus strands were detectable at 6 h postinoculation (hpi) of protoplasts, accumulating to about 10% of wild-type levels at 24 hpi. This mutation, when in the plus-sense orientation, had little or no effect on minus-strand synthesis from full-length satC plus strands in vitro, suggesting that the 5'-proximal RNA element is required for satC plus-strand synthesis. In addition, in vivo genetic selection revealed a strict requirement for 10 of the 14 nucleotides of the element, indicating that the primary sequence is essential for RNA accumulation. PMID- 10704344 TI - Epitope-tagging approach to determine the stoichiometry of the structural and nonstructural proteins in the virus particles: amount of Vpr in relation to Gag in HIV-1. AB - We used an epitope-tagging approach to determine the ratio of Gag (structural) to Vpr (nonstructural) in the virus particles directed by human immunodeficiency virus type 1. For this purpose, chimeric Gag and Vpr expression plasmids were constructed with the Flag epitope (DYKDDDDK), and the sequences corresponding to the chimeric protein were introduced into human immunodeficiency virus type 1 proviral DNA (NL4-3) to determine the ratio in the virus particles when these proteins are expressed in cis. In addition, NL4-3 DNA was modified to disrupt Vpr synthesis to determine the extent of incorporation of Vpr-FL when it is expressed in trans through a heterologous promoter. The analysis of virus particles generated by transfection of proviral DNA into RD cells indicated that (1) the ratio of Gag to Vpr in virus particles, when Vpr-FL is expressed in cis (in the context of proviral DNA), is in the range of 150-200:1 (14-18 molecules of Vpr per virion) and (2) the expression of Vpr-FL in trans showed efficient incorporation with a Gag to Vpr ratio of 5-7:1 (392-550 molecules of Vpr). These results suggest that the presence of the same epitope on different viral proteins may provide an accurate comparison of these proteins in the virus particles. PMID- 10704345 TI - The E7 oncoprotein of human papillomavirus type 16 interacts with F-actin in vitro and in vivo. AB - We report here that E7 oncoprotein of human papillomavirus type 16 (HPV-16) forms a complex in vivo and in vitro with actin, one of the components of the cellular cytoskeleton. The in vivo interaction was detected by immunofluorescent staining and confocal microscopic examination of normal human oral keratinocytes (NHOK) and CV-1 cells after transient expression of E7 employing the vaccinia virus-T7 RNA polymerase system and by coimmunoprecipitation from an immortalized, nontumorigenic cell line obtained after transfecting NHOK with the cloned HPV-16 DNA genome. The in vitro interaction was detected by cosedimentation of bacterially expressed E7 phosphorylated with rabbit reticulocyte lysate or purified casein kinase II (CKII) prior to incubation with F-actin. This interaction was inhibited if E7 phosphorylation by the rabbit reticulocyte lysate was prevented with heparin, a CKII inhibitor, or if the amino acids Ser-31 and Ser-32 in E7, which are phosphorylated by CKII, were replaced with amino acids that cannot be phosphorylated. Interestingly, a decrease in the amount of polymerized actin occurred in cells expressing E7. PMID- 10704346 TI - Heparan sulfate glycosaminoglycans are involved in adenovirus type 5 and 2-host cell interactions. AB - Gene therapy vectors derived from subgroup C adenoviruses of the serotype 5 (Ad5) and 2 (Ad2) resulted in inefficient infection of well differentiated respiratory cells, both in vitro and in vivo. The level of expression and localization of the primary receptor for Ad5 and Ad2, termed CAR, do not completely explain why the infection efficiency varies greatly in different experimental conditions. The possibility that additional receptors like proteoglycans are involved in the infection of Ad5 and Ad2 was investigated, because several pathogenic microorganisms use heparan sulfate-glycosaminoglycans (HS-GAGs) as coreceptors for multistep attachment to target cells. The HS-GAG analog heparin decreased Ad5 and Ad2-mediated infection and binding starting from the concentration of 0.1 microgram/ml, up to a maximum of 50%. A similar reduction in Ad5 binding and infection was obtained by treatment of cells with heparin lyases I, II, and III but not with chondroitin ABC lyase. The effect of heparin on Ad5 binding has not been observed in surface GAG-defective Raji cells and after treating A549 cells with heparin lyases I, II,and III. The binding of Ad5 was completely abolished when both CAR was blocked with RmcB antibody and HS-GAGs were competitively inhibited by heparin. Parallel experiments demonstrate that HS-GAGs are irrelevant to binding and infection of the subgroup B adenovirus type 3. Collectively, these results demonstrate for the first time that HS-GAGs expressed on the cell surface are involved in the binding of Ad5 and Ad2 to host cells. PMID- 10704347 TI - Site-specific integration of adeno-associated virus-based plasmid vectors in lipofected HeLa cells. AB - Adeno-associated virus (AAV) integrates specifically into a site (AAVS1) on human chromosome 19q13.3-qter. Similarly, there is accumulating evidence that this site specific integration occurs by transfection of AAV-based plasmid vectors. In order to further define the process of plasmid integration events, we constructed some AAV plasmids, introduced them into HeLa cells by lipofection, and isolated chromosomal integrants. One of such plasmids, pTH-5, contained the rep and neomycin-resistant (neo(r)) genes flanked by the 5'- and 3'-inverted terminal repeats of AAV and the hygromycin-resistant (hyg(r)) gene located in the plasmid backbone. Southern blot analysis revealed that among 36 G418-resistant (G418(r)) clones isolated, 22 (61%) showed site-specific integration into AAVS1. Further structural and functional analyses on the expression of the hyg(r) gene in the site-specific clones and the LacZ gene in clones generated with plasmid pTH-2 indicated that, together with the AAV sequence, the plasmid backbone was integrated into the AAVS1 site and thus the neo(r) and hyg(r) genes remained linked at high frequencies in the targeted integrants compared with random integrants. Sequence analysis of integration junctions between pTH-5 and AAVS1 revealed that the junctions occurred in the p5 promoter region of the plasmid while mainly in the partial cDNA coding region of the AAVS1 site. We also found that plasmid pTH-1 linearized in the backbone before lipofection gave a significantly lower frequency of site-specific integration (26%) than the circular form (60%). This finding may support the involvement of the double stranded, circular form of infected AAV in the integration process. Our results may help to understand the process and mechanism of site-specific integration of lipofected AAV plasmid vectors. PMID- 10704348 TI - Interaction with human immunodeficiency virus (HIV) type 2 predicts HIV type 1 genotype. AB - In West Africa, India, and certain regions of Europe, both human immunodeficiency viruses types 1 and 2 (HIV-1 and HIV-2) are known to cocirculate. To investigate the HIV-1 subtypes involved in dual HIV-1 and HIV-2 infections, we sequenced the envelope C2-V3 region from 29 dually infected female commercial sex workers from Senegal. The majority of women (23 of 29) were infected by HIV-1 subtype A. Within the HIV-1 subtype A sequences, 14 of 23 (60.8%) clustered with the West African associated A/G recombinant form (IbNG), and 9 of 23 (39.2%) formed a separate cluster distinct from the A/G IbNG. In contrast, in HIV-1 singly infected individuals, non-IbNG subtype A was found in only 13 of 98 (13.3%). Therefore, the lack of protection and/or interaction with HIV-2 was associated with a distinct HIV-1 A genotype. These results suggest differences in the biological properties of HIV-1 genotypes and their in vivo interaction with HIV 2. PMID- 10704349 TI - Vaccination to treat persistent viral infection. AB - Persistent infections caused by such agents as the human immunodeficiency virus, hepatitis B virus, Epstein-Barr virus, etc., present formidable medical problems. A defining characteristic of these infections is that anti-viral cytotoxic T lymphocytes (CTL) may be lost or, if present, fail to clear the infection. Here we report a vaccination strategy which was successful in generating lytic CTL in persistently infected mice. Vaccination with an immunodominant CTL epitope derived from the nucleoprotein of lymphocytic choriomeningitis virus (LCMV) delivered in the form of a lipopeptide incorporating a universal CD4 helper epitope successfully induced lytic MHC-restricted CTL in mice persistently infected with LCMV since birth. However, induction of such CTL did not eliminate the virus, most likely because the CTL were generated at low frequencies and had 2 to 3 logs lower affinity than CTL generated in uninfected mice inoculated with the vaccine. Both CTL populations from either uninfected or persistently infected mice produced significant and similar amounts of interferon-gamma and IL-6. Vaccine-induced low-affinity CTL were still inadequate at complete removal of the virus when combined with LCMV-specific CD4 helper T lymphocytes. Thus, our results establish that CTL can be generated in persistently infected mice and that a crucial factor for clearing viral infection is the affinity of the CTL. PMID- 10704350 TI - In vivo infection of ramified microglia from adult cat central nervous system by feline immunodeficiency virus. AB - Infection of microglial cells by the human immunodeficiency virus (HIV) is supposed to play an important role in the pathogenesis of AIDS-related central nervous system (CNS) complications. So far, however, experimental data about interactions between HIV and ramified microglia from the adult CNS were only occasionally reported, making it difficult to understand the exact nature of pathogenic events contributing to HIV-encephalopathy. Therefore, we used the animal model of feline immunodeficiency virus (FIV) infection of domestic cats to establish an experimental system which is suitable for studying the relationships between an immunodeficiency virus and the mature ramified microglia of the central nervous system. By means of density gradient centrifugation approximately 95% pure microglial cells could be isolated from adult feline brain that were characterized by their CD45(low) phenotype. Resident microglia extracted from the CNS of experimentally infected cats harbored FIV-specific DNA and cocultivation with mitogen-activated, but uninfected peripheral blood mononuclear cells (PBMC) resulted in recovery of high-titered infectious virus. Double labeling of brain cell monocultures explanted from persistently infected animals for both microglia and FIV markers disclosed less than 1% of viral antigen expressing microglial cells. This suggests that during the subclinical phase of the infection only a small number of brain-resident macrophages are productively infected. However, interaction of FIV-infected microglia and inflammatory lymphocytes may promote viral replication, thus supporting viral spread in brain tissue. PMID- 10704351 TI - The human papillomavirus type 11 E1E4 protein is phosphorylated in genital epithelium. AB - The most abundant viral transcript in human papillomavirus (HPV) 11-infected xenograft tissue has been shown to encode the E1(wedge)E4 protein. The function of E1(wedge)E4 protein has not been determined. Several potential phosphorylation sequence motifs were identified in the HPV 11 E1(wedge)E4 protein, including potential sites of phosphorylation by mitogen-activated protein kinase (MAPK), cAMP-dependent protein kinase (PKA), casein kinase II, and protein kinase C. To test phosphorylation of the HPV 11 E1(wedge)E4 protein, a soluble maltose binding protein (MBP) fusion was produced in Escherichia coli. Only MAPK and PKA phosphorylated the E1(wedge)E4 protein. Phosphoamino acid analysis showed that one or more threonine residues were phosphorylated by MAPK, and both serine and threonine residues were phosphorylated by PKA. MBP-E1(wedge)E4 mutant proteins were designed to delineate the E1(wedge)E4 phosphoacceptor residues. MAPK was shown to phosphorylate E1(wedge)E4 on threonine 53 within a MAPK consensus phorphorylation sequence motif. PKA was shown to phosphorylate E1(wedge)E4 at two residues: threonine 36 within a consensus motif and serine 44 within a variant of the PKA consensus phosphorylation sequence motif. HPV 11-infected human genital tissue grown as a xenograft in an athymic mouse was labeled with [(32)P]orthophosphate. Phosphoamino acid analysis of E1(wedge)E4 protein immunoprecipitated from (32)P-labeled tissue revealed that both serine and threonine residues were phosphorylated. Analysis by liquid chromatography-mass spectrophotometry was consistent with phosphorylation of residues within the PKA and MAPK phosphorylation sequence motifs. Expression of E1(wedge)E4 protein containing phosphorylation substitution mutations showed that the PKA mutant did not differ from wild-type E1(wedge)E4 protein in intracellular distribution. In contrast, the MAPK mutant did not localize exclusively to the cytoplasm nor did it colocalize with wild-type E1(wedge)E4 protein. We conclude that HPV 11 E1(wedge)E4 protein is phosphorylated in vitro and in vivo. Our data are consistent with phosphorylation of HPV 11 E1(wedge)E4 protein by MAPK and PKA in infected tissue. PMID- 10704352 TI - Generation of intersubtype human immunodeficiency virus type 1 recombinants in env gene in vitro: influences in the biological behavior and in the establishment of productive infections. AB - The occurrence of human immunodeficiency virus type 1 (HIV-1) recombinant genomes belonging to different subtypes is a common event in regions where more than two subtypes cocirculate. Although there are accumulating data toward an increase in the number of intersubtype recombinants, little has been addressed about the biological behavior of such mosaic genomes. This work reports the biological characterization of engineered in vitro HIV-1 intersubtype recombinants in the gp120 region. The recombinants possess the entire gp120 of B or F Brazilian isolates in the Z6 (subtype D) backbone. Here we show that this type of recombinant structure results in profound impairment to the establishment of productive infections in CD4-positive cells. The characterization of biological properties of those recombinant viruses demonstrated viral production occurring only during a transient peak early on infection and that they are not able to down-regulate the expression of CD4 receptor on the cell surface. We also report the phenotype reversion of one recombinant virus studied here, after 62 days in culture. Two amino acid substitutions in highly constant gp120 regions (C1 and C4) were identified in the revertant virus. The mutation occurring in the C4 region is localized near two amino acid residues critical for gp120/CD4 interaction. Based on these data, we suggest that failure in CD4 down-modulation by recombinant viruses can be due to a structural dysfunction of gp160 protein unable to block CD4 at the endoplasmic reticule. The possibilities that the establishment of latent infections can be directly related to the continuous expression of CD4 on the infected cell surface and that the occurrence of mutations in amino acid nearby residues critical for gp120/CD4 interaction can restore the fully productive infectious process are discussed. PMID- 10704353 TI - Phosphorylation of the RAP74 subunit of TFIIF correlates with Tat-activated transcription of the HIV-1 long terminal repeat. AB - Transcription from the HIV-1 long terminal repeat (LTR) is regulated by the viral transactivator Tat, which increases RNA polymerase II (RNAP II) processivity. Previous reports have demonstrated that phosphorylation of the RNAP II carboxy terminal domain by TFIIH and P-TEFb is important for Tat transactivation. Our present results demonstrate that phosphorylation of the RAP74 subunit of TFIIF is also an important step in Tat transactivation. Interestingly, while the general transcription factor TFIIF is required for both basal and Tat-activated transcription, phosphorylation of the RAP74 subunit occurs in the presence of Tat and correlates with a high level of transcription activity. Using a biotinylated DNA template transcription assay, we provide evidence that RAP74 is phosphorylated by TAF(II)250 during Tat-activated transcription. Depletion of RAP74 from the HeLa nuclear extract inhibited HIV-1 LTR-driven basal transcription and Tat transactivation. The addition of TFIIF, reconstituted from recombinant RAP30 and RAP74, to the depleted HeLa nuclear extract resulted in restoration of Tat transactivation. Of importance, the exogenous RAP74 was rapidly phosphorylated in the presence of Tat. These results suggest that RAP74 phosphorylation is one important step, of several, in the Tat transactivation cascade. PMID- 10704354 TI - "Local rules" theory applied to polyomavirus polymorphic capsid assemblies. AB - The papovaviruses are nonenveloped dsDNA viruses whose capsids are characterized by a non-quasi-equivalent bonding pattern in which 72 pentameric capsomeres occupy positions having either five or six neighboring capsomeres. The local rules theory of Berger et al. (1994, Proc. Natl. Acad. Sci. USA, 91, 7732-7736), previously developed to explain aspects of icosahedral capsid assembly, has been applied to the papovavirus geometry. Local rules describe capsid symmetry patterns in terms of the local interactions of assembly units, such as coat proteins or capsomeres. Polymorphic assemblies, including T = 1 icosahedral, dodecahedral, spiral, and tubular structures of the polyomavirus VP1 protein, can be induced by specific mutations or changes in the solvent conditions during in vitro assembly of the recombinant coat protein. Local rules models were developed to model the wild-type capsid and several polymorphic assemblies. Some assemblies corresponded to structures modeled by small deviations from wild-type local rules. We conclude that aspects of polyomavirus assembly are consistent with local rules models, although they do not explain all polymorphisms. These results may provide insights into the nature of papovavirus assembly, constraints on assembly pathways, and strategies for disrupting assembly. PMID- 10704355 TI - Vaccinia virus-related events and phenotypic changes after infection of dendritic cells derived from human monocytes. AB - The in vitro interactions between vaccinia virus (VV) and monocyte-derived human dendritic cells (DC) have been studied to gain a better understanding of the mechanisms involved in the induction of an immune response by VV. This work showed that VV binds to DC less efficiently than to HeLa cells (HeLa). Capping of viral antigens on the DC surface and electron microscopic examinations suggested that VV enters into DC mainly by endocytosis instead of fusion as for HeLa. Early viral-encoded proteins were expressed in DC but late viral proteins and viral DNA synthesis did not occur. Nevertheless, when successfully infected, DC expressed a similar amount of a foreign, viral-encoded protein, as HeLa, if the early component of the p7.5 promoter was used. VV infection did not lead to DC maturation as determined by following the level of several cell surface markers associated with maturation, but an inhibition of the expression of the costimulatory molecule CD80 was noticed. The proliferation of allogeneic peripheral blood lymphocytes (PBL) was stimulated by VV-infected DC or inhibited depending on the particular donor lymphocytes employed. PBL from VV-vaccinated individuals with good memory responses to VV antigens proliferated in the presence of infected autologous DC. PBL from individuals with poor memory responses to VV and one unvaccinated individual also proliferated, albeit to a lower level, in the presence of infected autologous DC. These results suggest that VV-infected DC could both stimulate memory cells and prime naive cells in vitro. PMID- 10704356 TI - Quantitation of virus-specific classes of antibodies following immunization of mice with attenuated equine herpesvirus 1 and viral glycoprotein D. AB - The antibody responses of CBA/J mice infected intranasally (i.n.) with either the attenuated KyA strain or the pathogenic RacL11 strain of equine herpesvirus 1 (EHV-1) or immunized with recombinant glycoprotein D (rgD) were investigated using the ELISPOT assay to measure EHV-1-specific antibody-secreting cells (ASC) in the regional lymphoid tissue of the respiratory tract. IgG, IgA, and IgM ASC specific for EHV-1 were detected in the mediastinal lymph nodes (MLN) and lungs 2 weeks after i.n. infection with EHV-1 strain KyA or RacL11, or immunization with heat-killed KyA or rgD. EHV-1-specific ASC were present in the MLN and lungs at 4 and 8 weeks, but declined in frequency by fivefold in the lung at 8 weeks. However, i.n. immunized (2 x 10(6) pfu KyA or 50 microgram rgD/mouse) mice infected at 8 weeks with pathogenic EHV-1 RacL11 resisted challenge and showed eight- and tenfold increases in MLN ASC and lung ASC, respectively, by 3 days after challenge. In contrast to the intranasal route of immunization, intraperitoneal immunization yielded ASC frequencies in the MLN and lungs that were only slightly above those of nonimmunized control mice. These data indicate that immunization with infectious or heat-killed EHV-1 KyA, or rgD, induces significant levels of virus-specific ASC both in the MLN and lungs, a specific memory B-cell response, and long-term protective immunity. The finding that the numbers of ASC induced by the pathogenic strain versus the attenuated strain of EHV-1, which were virtually identical, indicated that the ability to generate a B cell response is independent of and does not contribute to EHV-1 virulence. PMID- 10704357 TI - Interaction between virion-bound host intercellular adhesion molecule-1 and the high-affinity state of lymphocyte function-associated antigen-1 on target cells renders R5 and X4 isolates of human immunodeficiency virus type 1 more refractory to neutralization. AB - The oligomeric nature of the viral envelope proteins has been partly held responsible for the observed differences in neutralization sensitivity between primary and laboratory-adapted strains of human immunodeficiency virus type 1 (HIV-1). However, recent evidence suggests that host factors can also modify the sensitivity of HIV-1 particles to neutralization. Having previously demonstrated that the acquisition of host-encoded intercellular adhesion molecule (ICAM)-1 proteins by newly formed viruses has a functional significance for the life cycle of HIV-1, we investigated whether the acquisition of host-derived ICAM-1 by HIV-1 could affect the virus sensitivity to neutralization. In this study, we have first shown that the physical presence of host cell membrane ICAM-1 on HIV-1 was not modifying virus sensitivity to neutralization by either two different anti gp120 monoclonal antibodies (0.5beta and 4.8D) or soluble CD4. However, the ability of the F105 anti-gp120 monoclonal antibody (specific for the CD4-binding site) to neutralize ICAM-1-bearing virions was diminished when target cells were pretreated with an lymphocyte function-associated antigen-1 (LFA-1)-activating antibody. Interestingly, ICAM-1/POS progeny viruses were found to be slightly more resistant to neutralization by individual human sera in target cells expressing a low-affinity form of LFA-1 than viruses devoid of host-encoded ICAM 1 proteins. This resistance was markedly enhanced when target cells expressed an activated LFA-1 form on their surface. These results suggest that the interaction between virally embedded host ICAM-1 and target cell surface LFA-1 should be considered a factor modulating neutralization sensitivity of HIV-1 by human sera from HIV-1-infected individuals. PMID- 10704359 TI - Functional significance of alternate phosphorylation in Sendai virus P protein. AB - Phosphorylation of the negative-sense RNA virus phosphoproteins is highly conserved, implying functional significance. Sendai virus (SV) phosphoprotein (P) is constitutively phosphorylated at S249. Abrogation of the SV P primary phosphorylation causes phosphorylation of P at alternate sites, creating a problem in determining the function of phosphorylation. We have now identified the alternate phosphorylation sites using two-dimensional phosphopeptide analysis of several deletion and point mutants of the P protein. The alternate phosphorylation sites were mutagenized to create P with (S249combo) or without (combo) primary phosphorylation. The combo protein has less than 10% phosphorylation compared with the wild-type P or S249combo. Functional analysis of the mutant proteins using a Sendai virus minigenome replication system showed that the combo P protein was as proficient in supporting minigenome replication as the wild-type P in cell cultures. These studies suggest that like the primary, the alternate phosphorylation of the P protein is also dispensable for virus replication in cell cultures. Interestingly, the ability of the multiple site mutant of P (combo mutant has eight serine residues changed to alanine residues) to support efficient virus RNA synthesis suggests that the P protein has a high flexibility at least in its sequence and perhaps also in structure. PMID- 10704358 TI - U94, the human herpesvirus 6 homolog of the parvovirus nonstructural gene, is highly conserved among isolates and is expressed at low mRNA levels as a spliced transcript. AB - Human herpesvirus 6 variants A and B (HHV-6A and HHV-6B, respectively) encode homologs (U94) of the parvovirus nonstructural gene, ns1 or rep. Here we describe the HHV-6B homolog and analyze its genetic heterogeneity and transcription. U94 nucleotide and amino acid sequences differ by approximately 3.5% and 2.5%, respectively, between HHV-6A and HHV-6B. Among a collection of 17 clinically and geographically disparate HHV-6 isolates, intravariant nucleotide and amino acid sequence divergence was less than 0.6% and 0.2%, respectively; all 13 HHV-6B isolates had identical amino acid sequences. The U94 transcript is spliced to remove a 2.6-kb intron and is expressed at very low levels relative to other HHV 6B genes, reaching approximately 10 copies per cell 3 days after infection. The mRNA has several small AUG-initiated open reading frames upstream of the U94 open reading frame, a hallmark of proteins expressed at low levels. Consistent with this, the U94-encoded protein was immunologically undetectable in HHV-6B-infected cells. The high degree of sequence conservation suggests that the gene function is nearly intolerant of sequence variation. The low abundance of U94 transcripts and the presence of encoded inefficient translation initiation suggest that the U94 protein may be required only in small amounts during infection. PMID- 10704360 TI - Role of matrix protein in the type D retrovirus replication cycle: importance of the arginine residue at position 55. AB - We previously reported that a mutant of Mason-Pfizer monkey virus (M-PMV), which has an amino acid substitution in the matrix (MA) protein at position 55, MA R55W, showed altered viral morphogenesis, reduced glycoprotein incorporation, and loss of infectivity. In this report, we show that two additional amino acid substitutions at this site in MA, R55F and R55Y, also result in similar altered morphogenesis, Env incorporation, and infectivity, demonstrating that these changes are not specific for the substitution of tryptophan in place of arginine 55. Attempts to isolate second site infectious revertants from cells transfected with the R55W mutant genome resulted only in the recovery of infectious viruses in which the codon at position 55 had reverted to one encoding arginine. In contrast, no revertants were obtained from the phenylalanine and tyrosine mutants in which three nucleotide changes had been engineered into the arginine codon. These results confirm that the arginine residue at position 55 is critical for intracellular targeting of M-PMV Gag molecules and support the concept that as part of a cytoplasmic transport retention signal R55 interacts with cellular trafficking components rather than other regions of Gag. PMID- 10704361 TI - Positive and negative regulation of TGF-beta signaling. AB - Cytokines of the transforming growth factor beta (TGF-beta) superfamily, including TGF-betas, activins and bone morphogenetic proteins (BMPs), bind to specific serine/threonine kinase receptors and transmit intracellular signals through Smad proteins. Upon ligand stimulation, Smads move into the nucleus and function as components of transcription complexes. TGF-beta and BMP signaling is regulated positively and negatively through various mechanisms. Positive regulation amplifies signals to a level sufficient for biological activity. Negative regulation occurs at the extracellular, membrane, cytoplasmic and nuclear levels. TGF-beta and BMP signaling is often regulated through negative feedback mechanisms, which limit the magnitude of signals and terminate signaling. Negative regulation is also important for formation of gradients of morphogens, which is crucial in developmental processes. In addition, other signaling pathways regulate TGF-beta and BMP signaling through cross-talk. Nearly 20 BMP isoforms have been identified, and their activities are regulated by various extracellular antagonists. Regulation of TGF-beta signaling might be tightly linked to tumor progression, since TGF-beta is a potent growth inhibitor in most cell types. PMID- 10704362 TI - The ran decathlon: multiple roles of Ran. AB - The Ran GTPase system affects many cellular processes, including the regulation of cell cycle progression, nuclear envelope structure and function, and nucleocytoplasmic transport. The biochemical basis for the involvement of Ran in nuclear import and export has been well documented, but the direct targets of Ran in other cellular processes have not yet been identified. There is, however, mounting evidence that Ran directly affects at least some of these other cellular processes by mechanisms independent of its role in transport. In this Commentary we discuss evidence linking Ran to different aspects of cell function, and how these multiple facets of Ran's activity may relate to each other. PMID- 10704363 TI - Trachynilysin mediates SNARE-dependent release of catecholamines from chromaffin cells via external and stored Ca2+. AB - Trachynilysin, a 159 kDa dimeric protein purified from stonefish (Synanceia trachynis) venom, dramatically increases spontaneous quantal transmitter release at the frog neuromuscular junction, depleting small clear synaptic vesicles, whilst not affecting large dense core vesicles. The basis of this insensitivity of large dense core vesicles exocytosis was examined using a fluorimetric assay to determine whether the toxin could elicit catecholamine release from bovine chromaffin cells. Unlike the case of the motor nerve endings, nanomolar concentrations of trachynilysin evoked sustained Soluble N-ethylmaleimide sensitive fusion protein Attachment Protein REceptor-dependent exocytosis of large dense core vesicles, but only in the presence of extracellular Ca2+. However, this response to trachynilysin does not rely on Ca2+ influx through voltage-activated Ca2+ channels because the secretion was only slightly affected by blockers of L, N and P/Q types. Instead, trachynilysin elicited a localized increase in intracellular fluorescence monitored with fluo-3/AM, that precisely co-localized with the increase of fluorescence resulting from caffeine-induced release of Ca2+ from intracellular stores. Moreover, depletion of the latter stores inhibited trachynilysin-induced exocytosis. Thus, the observed requirement of external Ca2+ for stimulation of large dense core vesicles exocytosis from chromaffin cells implicates plasma membrane channels that signal efflux of Ca2+ from intracellular stores. This study also suggests that the bases of exocytosis of large dense core vesicles from motor nerve terminals and neuroendocrine cells are distinct. PMID- 10704364 TI - Progesterone regulates the accumulation and the activation of Eg2 kinase in Xenopus oocytes. AB - Xenopus prophase oocytes reenter meiotic division in response to progesterone. The signaling pathway leading to Cdc2 activation depends on neosynthesized proteins and a decrease in PKA activity. We demonstrate that Eg2 protein, a Xenopus member of the Aurora/Ipl1 family of protein kinases, accumulates in response to progesterone and is degraded after parthenogenetic activation. The polyadenylation and cap ribose methylation of Eg2 mRNA are not needed for the protein accumulation. Eg2 protein accumulation is induced by progesterone through a decrease in PKA activity, upstream of Cdc2 activation. Eg2 kinase activity is undetectable in prophase and is raised in parallel with Cdc2 activation. In contrast to Eg2 protein accumulation, Eg2 kinase activation is under Cdc2 control. Furthermore, by using an anti-sense strategy, we show that Eg2 accumulation is not required in the transduction pathway leading to Cdc2 activation. Altogether, our results strongly suggest that Eg2 is not necessary for Cdc2 activation, though it could participate in the organization of the meiotic spindles, in agreement with the well-conserved roles of the members of the Aurora family, from yeast to man. PMID- 10704365 TI - Unscheduled re-entry into the cell cycle induced by NGF precedes cell death in nascent retinal neurones. AB - During their early postmitotic life, a proportion of the nascent retinal ganglion cells (RGCs) are induced to die as a result of the interaction of nerve growth factor (NGF) with the neurotrophin receptor p75. To analyse the mechanisms by which NGF promotes apoptosis, an in vitro culture system consisting of dissociated E5 retinal cells was established. In this system, NGF-induced apoptosis was only observed in the presence of insulin and neurotrophin-3, conditions that favour the birth of RGCs and other neurones expressing the glycoprotein G4. The pro-apoptotic effect of NGF on the G4-positive neurones was evident after 10 hours in vitro and was preceded by a significant upregulation of cyclin B2, but not cyclin D1, and the presence of mitotic nuclei in these cells. Brain-derived neurotrophic factor prevented both the increase of cyclin B2 expression in the G4-positive neurones and the NGF-induced cell death. Finally, pharmacologically blocking cell-cycle progression using the cyclin-dependent kinase inhibitor roscovitine prevented NGF-induced cell death in a dose-dependent manner. These results strongly suggest that the apoptotic signalling initiated by NGF requires a driving stimulus manifested by the neuronal birth and is preceded by the unscheduled re-entry of postmitotic neurones into the cell cycle. PMID- 10704366 TI - Chromosome condensation induced by geminivirus infection of mature plant cells. AB - Tomato golden mosaic virus (TGMV) is a geminivirus that replicates its single stranded DNA genome through double-stranded DNA intermediates in nuclei of differentiated plant cells using host replication machinery. We analyzed the distribution of viral and plant DNA in nuclei of infected leaves using fluorescence in situ hybridization (FISH). TGMV-infected nuclei showed up to a sixfold increase in total volume and displayed a variety of viral DNA accumulation patterns. The most striking viral DNA patterns were bright, discrete intranuclear compartments, but diffuse nuclear localization was also observed. Quantitative and spatial measurements of high resolution 3-dimensional image data revealed that these compartments accounted for 1-18% of the total nuclear volume or 2-45% of the total nuclear FISH signals. In contrast, plant DNA was concentrated around the nuclear periphery. In a significant number of nuclei, the peripheral chromatin was organized as condensed prophase-like fibers. A combination of FISH analysis and indirect immunofluorescence with viral coat protein antibodies revealed that TGMV virions are associated with the viral DNA compartments. However, the coat protein antibodies failed to cross react with some large viral DNA inclusions, suggesting that encapsidation may occur after significant viral DNA accumulation. Infection by a TGMV mutant with a defective coat protein open reading frame resulted in fewer and smaller viral DNA containing compartments. Nevertheless, nuclei infected with the mutant virus increased in size and in some cases showed chromosome condensation. Together, these results established that geminivirus infection alters nuclear architecture and can induce plant chromatin condensation characteristic of cells arrested in early mitosis. PMID- 10704367 TI - Clonal mesenchymal progenitors from human bone marrow differentiate in vitro according to a hierarchical model. AB - Bone marrow stromal cells can give rise to several mesenchymal lineages. The existence of a common stem/progenitor cell, the mesenchymal stem cell, has been proposed, but which developmental stages follow this mesenchymal multipotent progenitor is not known. Based on experimental evidence, a model of mesenchymal stem cell differentiation has been proposed in which individual lineages branch directly from the same progenitor. We have verified this model by using clonal cultures of bone marrow derived stromal fibroblasts. We have analyzed the ability of 185 non-immortalized human bone marrow stromal cell clones to differentiate into the three main lineages: osteo-, chondro- and adipogenic. All clones but one differentiated into the osteogenic lineage. About one third of the clones differentiated into all three lineages analyzed. Most clones (60-80%) displayed an osteo-chondrogenic potential. We have never observed clones with a differentiation potential limited to the osteo-adipo- or to the chondro adipogenic phenotype, nor pure chondrogenic and adipogenic clones. How long the differentiation potential of a number of clones was maintained was assessed throughout their life span. Clones progressively lost their adipogenic and chondrogenic differentiation potential at increasing cell doublings. Our data suggest a possible model of predetermined bone marrow stromal cells differentiation where the tripotent cells can be considered as early mesenchymal progenitors that display a sequential loss of lineage potentials, generating osteochondrogenic progenitors which, in turn, give rise to osteogenic precursors. PMID- 10704368 TI - Interaction between the cytodomains of the alpha 3 and beta 1 integrin subunits regulates remodelling of adhesion complexes on laminin. AB - The first step of laminin 1-induced signal transduction is initiated by the formation of alpha 6 beta 1 integrin-specific adhesion complexes. In contrast, on other laminin isoforms the adhesion complexes are alpha 3 beta 1 integrin specific due to a transdominant regulation of the alpha 6 beta 1 integrin by the alpha 3 beta 1 integrin. To determine the mechanism of this regulation, peptides representing the cytoplasmic domain of the alpha 3 or alpha 6 integrin subunits were microinjected together with recombinant enhanced green fluorescence protein into live fibroblasts. Microinjection of the alpha 3 integrin peptide to laminin 1-adherent cells displaying alpha 6 beta 1 integrin-specific adhesion complexes resulted in the disengagement of the alpha 6 beta 1 integrin, while microinjection of green fluorescence protein alone or in combination with the alpha 6 integrin cytodomain had no effect. Further surface plasmon resonance studies revealed that the cytodomain of the beta 1 integrin subunit interacts with low affinity with the cytoplasmic tail of the alpha 3 integrin subunit, but not with that of several other alpha subunits including alpha 6. These results imply that the cytoplasmic tails of the integrin alpha subunits play a critical role in the regulation of integrin-induced signal transduction. In particular, the intracellular tail of the alpha 3 integrin subunit controls the formation of adhesion complexes in cells adhering to laminins. PMID- 10704369 TI - Rho and Rac exert antagonistic functions on spreading of macrophage-derived multinucleated cells and are not required for actin fiber formation. AB - Multinucleated giant cells (MNGC) derived from avian blood monocytes present, like osteoclasts, an unusual cytoskeletal organization characterized by (1) cortical rings of actin filaments, (2) unique adhesion structures called podosomes and (3) vinculin containing focal complexes which are not visibly connected to F-actin structures. The Rho family of small GTPases plays an essential role in the regulation and organization of cellular cytoskeletal structures including F-actin and vinculin associated structures. Using bacterial toxins such as modified exoenzyme C3 (C3B) and toxin B or overexpression of constitutively active Rac and Rho proteins fused to the green fluorescent protein (GFP), we show that Rac and Rho play antagonistic roles in regulating the morphology of osteoclast-like cells. Inhibition of Rho by C3B triggered MNGC spreading whereas activated Rho promoted cell retraction. However, inhibition or activation of Rho led to complete disorganization of fibrillar actin structures, including podosomes. Toxin B inhibition of Rho, Rac and Cdc42 induced a time dependent F-actin and vinculin reorganization. Initially, actin fibers with associated adhesion plaques formed and disappeared subsequently. Finally, only small focal complexes remained at the MNGC periphery before retracting. At the time when actin fibers formed, we observed that Rac was already inhibited by toxin B. By combining C3B treatment and overexpression of a dominant negative form of Rac (N17Rac), we show that the formation of these focal adhesion and actin fiber structures required neither Rho nor Rac activity. Moreover, our results show that podosomes are extremely unstable structures since any modifications of Rho or Rac activity resulted in their dissociation. PMID- 10704370 TI - Autophagic and apoptotic types of programmed cell death exhibit different fates of cytoskeletal filaments. AB - Programmed cell death comprises several subtypes, as revealed by electron microscopy. Apoptosis or type I programmed cell death is characterized by condensation of cytoplasm and preservation of organelles, essentially without autophagic degradation. Autophagic cell death or type II programmed cell death exhibits extensive autophagic degradation of Golgi apparatus, polyribosomes and endoplasmatic reticulum, which precedes nuclear destruction. In the present study, we analysed the fate of cytokeratin and F-actin during autophagic cell death in the human mammary carcinoma cell line MCF-7 because recent studies suggest that an intact cytoskeleton is necessary for autophagocytosis. Programmed cell death was induced by 10(-)(6) M tamoxifen. For quantitative light microscopic analysis, autophagic vacuoles were visualized by monodansyl cadaverin, which stains autophagic vacuoles as distinct dot-like structures. In control cultures, the number of monodansylcadaverin-positive cells did not exceed 2%. Tamoxifen induced a dramatic increase 2-4 days after treatment to a maximum of 60% monodansylcadaverin-positive cells between days 5 and 7. Cell death, as indicated by nuclear condensation, increased more gradually to about 18% of all cells on day 7. In cells with pyknotic nuclei cytokeratin appeared disassembled but retained its immunoreactivity; actin was still polymerized to filaments, as demonstrated by its reaction with phalloidin. Western blot analysis showed no significant cleavage of the monomeric cytokeratin fraction. For comparison, apoptotic or type I cell death was studied using the human colon cancer cell HT29/HI1 treated with the tyrosine kinase inhibitor tyrphostin A25 as a model. Cleavage of cytokeratin was already detectable in early morphological stages of apoptosis. F-actin was found to depolymerize; its globular form could be detected by antibodies; western blot analysis revealed no products of proteolytic cleavage. In conclusion, in our model of apoptosis, early stages are associated with depolymerization of actin and degradation of intermediate filaments. In contrast, during autophagic cell death intermediate and microfilaments are redistributed, but largely preserved, even beyond the stage of nuclear collapse. The present data support the concept that autophagic cell death is a separate entity of programmed cell death that is distinctly different from apoptosis. PMID- 10704371 TI - The Saccharomyces cerevisiae SDA1 gene is required for actin cytoskeleton organization and cell cycle progression. AB - The organization of the actin cytoskeleton is essential for several cellular processes. Here we report the characterization of a Saccharomyces cerevisiae novel gene, SDA1, encoding a highly conserved protein, which is essential for cell viability and is localized in the nucleus. Depletion or inactivation of Sda1 cause cell cycle arrest in G(1) by blocking both budding and DNA replication, without loss of viability. Furthermore, sda1-1 temperature-sensitive mutant cells arrest at the non-permissive temperature mostly without detectable structures of polymerized actin, although a normal actin protein level is maintained, indicating that Sda1 is required for proper organization of the actin cytoskeleton. To our knowledge, this is the first mutation shown to cause such a phenotype. Recovery of Sda1 activity restores proper assembly of actin structures, as well as budding and DNA replication. Furthermore we show that direct actin perturbation, either in sda1-1 or in cdc28-13 cells released from G(1) block, prevents recovery of budding and DNA replication. We also show that the block in G(1) caused by loss of Sda1 function is independent of Swe1. Altogether our results suggest that disruption of F-actin structure can block cell cycle progression in G(1) and that Sda1 is involved in the control of the actin cytoskeleton. PMID- 10704372 TI - Apical endocytosis of ricin in MDCK cells is regulated by the cyclooxygenase pathway. AB - Addition of arachidonic acid or stimulation of arachidonic acid production by secretory phospholipase A2 selectively upregulated apical endocytosis of ricin in MDCK cells without affecting basolateral endocytosis. Electron microscopic studies revealed that MDCK cells treated with secretory phospholipase A2 and incubated with horseradish peroxidase had an increased number of normal appearing peroxidase-labeled endosomes and no sign of membrane ruffling. Moreover, inhibition of basal arachidonic acid release, either by decreasing the cytosolic phospholipase A(2) activity or the diacylglycerol lipase activity, reduced the rate of apical endocytosis. Furthermore, indomethacin, an inhibitor of the cyclooxygenase pathway, counteracted the stimulation of endocytosis seen with both secretory phospholipase A2 and arachidonic acid, suggesting that formation of eicosanoids such as prostaglandins could be essential for the regulation. This idea was supported by the finding that prostaglandin E2, the predominant prostaglandin formed in kidney, also upregulated ricin uptake. The regulatory effect of the cyclooxygenase pathway on apical endocytosis of ricin was found to be independent of protein kinases A and C, which are known to selectively control apical clathrin-independent endocytosis in polarized cells. PMID- 10704373 TI - A checkpoint that monitors cytokinesis in Schizosaccharomyces pombe. AB - Cell division in Schizosaccharomyces pombe is achieved through the use of a medially positioned actomyosin ring. A division septum is formed centripetally, concomitant with actomyosin ring constriction. Genetic screens have identified mutations in a number of genes that affect actomyosin ring or septum assembly. These cytokinesis-defective mutants, however, undergo multiple S and M phases and die as elongated cells with multiple nuclei. Recently, we have shown that a mutant allele of the S. pombe drc1(+)/cps1(+) gene, which encodes a 1,3-(beta) glucan synthase subunit, is defective in cytokinesis but displays a novel phenotype. drc1-191/cps1-191 cells are capable of assembling actomyosin rings and completing mitosis, but are incapable of assembling the division septum, causing them to arrest as binucleate cells with a stable actomyosin ring. Each nucleus in arrested cps1-191 cells is able to undergo S phase but these G(2) nuclei are significantly delayed for entry into the M phase. In this study we have investigated the mechanism that causes cps1-191 to block with two G(2) nuclei. We show that the inability of cps1-191 mutants to proceed through multiple mitotic cycles is not related to a defect in cell growth. Rather, the failure to complete some aspect of cytokinesis may prevent the G(2)/M transition of the two interphase-G(2) nuclei. The G(2)/M transition defect of cps1-191 mutants is suppressed by a mutation in the wee1 gene and also by the dominant cdc2 allele cdc2-1w, but not the cdc2-3w allele. Transient depolymerization of all F-actin structures also allowed a significant proportion of the cps1-191 cells to undergo a second round of mitosis. We conclude that an F-actin and Wee1p dependent checkpoint blocks G(2)/M transition until previous cytokinesis is completed. PMID- 10704374 TI - Morphology and behaviour of dinoflagellate chromosomes during the cell cycle and mitosis. AB - The morphology and behaviour of the chromosomes of dinoflagellates during the cell cycle appear to be unique among eukaryotes. We used synchronized and aphidicolin-blocked cultures of the dinoflagellate Crypthecodinium cohnii to describe the successive morphological changes that chromosomes undergo during the cell cycle. The chromosomes in early G(1) phase appeared to be loosely condensed with numerous structures protruding toward the nucleoplasm. They condensed in late G(1), before unwinding in S phase. The chromosomes in cells in G(2) phase were tightly condensed and had a double number of arches, as visualised by electron microscopy. During prophase, chromosomes elongated and split longitudinally, into characteristic V or Y shapes. We also used confocal microscopy to show a metaphase-like alignment of the chromosomes, which has never been described in dinoflagellates. The metaphase-like nucleus appeared flattened and enlarged, and continued to do so into anaphase. Chromosome segregation occurred via binding to the nuclear envelope surrounding the cytoplasmic channels and microtubule bundles. Our findings are summarized in a model of chromosome behaviour during the cell cycle. PMID- 10704375 TI - Microtubules, but not actin filaments, drive daughter cell budding and cell division in Toxoplasma gondii. AB - We have used drugs to examine the role(s) of the actin and microtubule cytoskeletons in the intracellular growth and replication of the intracellular protozoan parasite, Toxoplasma gondii. By using a 5 minute infection period and adding the drugs shortly after entry we can treat parasites at the start of intracellular development and 6-8 hours prior to the onset of daughter cell budding. Using this approach we found, somewhat surprisingly, that reagents that perturb the actin cytoskeleton in different ways (cytochalasin D, latrunculin A and jasplakinolide) had little effect on parasite replication although they had the expected effects on the host cells. These actin inhibitors did, however, disrupt the orderly turnover of the mother cell organelles leading to the formation of a large residual body at the posterior end of each pair of budding parasites. Treating established parasite cultures with the actin inhibitors blocked ionophore-induced egression of tachyzoites from the host cells, demonstrating that intracellular parasites were susceptible to the effects of these inhibitors. In contrast, the anti-microtubule drugs oryzalin and taxol, and to a much lesser extent nocodazole, which affect microtubule dynamics in different ways, blocked parasite replication by disrupting the normal assembly of the apical conoid and the microtubule inner membrane complex (IMC) in the budding daughter parasites. Centrosome replication and assembly of intranuclear spindles, however, occurred normally. Thus, daughter cell budding per se is dependent primarily on the parasite microtubule system and does not require a dynamic actin cytoskeleton, although disruption of actin dynamics causes problems in the turnover of parasite organelles. PMID- 10704376 TI - Actin filaments and microtubules play different roles during bristle elongation in Drosophila. AB - Developing bristles in Drosophila pupae contain 7-11 bundles of crosslinked actin filaments and a large population of microtubules. During bristle growth the rate of cell elongation increases with bristle length. Thin section EM shows that bundle size is correlated with the amount of cytoplasm at all points along the bristle. Thus, as the bristle elongates and tapers, fewer actin filaments are used. To ensure penetration of inhibitors we isolated thoraces and cultured them in vitro; bristles elongate at rates identical to bristles growing in situ. Interestingly, inhibitors of actin filament assembly (cytochalasin D and latrunculin A) dramatically curtailed bristle elongation while a filament stabilizer (jasplakinolide) accelerated elongation. In contrast, inhibitors of microtubule dynamics (nocodazole, vinblastine, colchicine and taxol) did not affect bristle elongation. Surprisingly, the bristle microtubules are stable and do not turn over. Furthermore, the density of microtubules decreases as the bristle elongates. These two facts coupled with calculations and kinetics of elongation and the fact that the microtubules are short indicate that the microtubules are assembled early in development and then transported distally as the bristle grows. We conclude that actin assembly is crucial for bristle cell elongation and that microtubules must furnish other functions such as to provide bulk to the bristle cytoplasm as well as playing a role in vesicle transport. PMID- 10704377 TI - Ezrin links syndecan-2 to the cytoskeleton. AB - The syndecan family of heparan sulfate proteoglycans is known to associate with the actin cytoskeleton, possibly transducing signals from the extracellular matrix. In the search for proteins that could mediate the association of syndecan 2 with the actin cytoskeleton we found that ezrin, a protein which links membrane receptors to the cytoskeleton, coimmunoprecipitated with syndecan-2 in COS-1 cells. In vitro assays indicated a direct association between the amino-terminal domain of ezrin and the cytoplasmic domain of syndecan-2. Confocal microscopy showed colocalization of ezrin and syndecan-2 in actin-rich microspikes in COS-1 cells. The syndecan-2/ezrin protein complex was resistant to 0.2% Triton X-100 extraction but the syndecan-2/amino-terminal domain of ezrin complex was not, which indicated that carboxi-terminal domain of ezrin is involved in the cytoskeleton anchorage of this protein complex. Additionally we observed that the activation of rhoA GTPase increased syndecan-2 insolubility in 0.2% Triton X-100 and syndecan-2/ezrin association. Taken together, these results indicate that ezrin connects syndecan-2 to the actin cytoskeleton. PMID- 10704378 TI - Bidirectional transcytosis of IgG by the rat neonatal Fc receptor expressed in a rat kidney cell line: a system to study protein transport across epithelia. AB - The neonatal Fc receptor, FcRn, transports immunoglobulin G (IgG) across cellular barriers between mother and offspring. FcRn also protects circulating IgG from catabolism, probably during transport across the capillary endothelium. Only one cell culture model of transcytosis has been used extensively, the transport of IgA from the basolateral to the apical surface of Madin-Darby canine kidney cells by the polymeric immunoglobulin receptor (pIgR). We report that rat inner medullary collecting duct (IMCD) cells transfected with DNA encoding the (alpha) subunit of rat FcRn specifically and saturably transport Fc when grown as polarized monolayers. Using this system, we have found that transcytosis by FcRn, like transcytosis by the pIgR, depends upon an intact microtubule system. FcRn differs most strikingly from the pIgR in its ability to transport its ligand in both the apical to basolateral and basolateral to apical directions. The phosphatidylinositol 3-kinase inhibitors wortmannin and LY294002 inhibited basolateral to apical transport by FcRn more than apical to basolateral transport, suggesting that there are differences in the mechanisms of transport in the two directions. Lastly, we found that transcytosis by FcRn depends upon vesicular acidification. We anticipate that the IMCD cell culture model will allow further elucidation of the mechanism of IgG transport by FcRn. PMID- 10704379 TI - Ca2+-dependent myosin II activation is required for uropod retraction during neutrophil migration. AB - Buffering of intracellular Ca2+ transients in human neutrophils leads to reduced motility due to defective uropod detachment on fibronectin and vitronectin-coated surfaces. Since one potential target of a rise in [Ca2+]i is the activation of myosin II, we characterized the role of myosin II during motility. Treatment of neutrophils with a myosin inhibitor (2,3-butanedione monoxime), or myosin light chain kinase inhibitors (ML-7, ML-9, or KT5926) resulted in impaired uropod retraction and a dose-dependent decrease in chemokinesis following stimulation with N-formyl-Met-Leu-Phe (fMLP). Treatment with ML-9 resulted in a redistribution of F-actin and talin to the non-retracted uropods, mimicking the redistribution observed during [Ca2+]i buffering. Impairment of uropod retraction and redistribution of F-actin and talin by myosin II inhibition was only observed on adhesive substrates such as fibronectin and not on poorly adhesive substrates such as human serum-coated glass. At higher concentrations of ML-9, cell polarization was inhibited and pseudopod extension occurred radially. Using an antibody specific for serine 19-phosphorylated regulatory light chain of myosin II, regions of activated myosin II were found at the leading edge as well as the uropod in motile fMLP-stimulated cells. [Ca2+]i depletion caused a 50% decrease in the level of serine 19-phosphorylated myosin II suggesting that activation of myosin II by intracellular Ca2+ transients may be an essential step in establishing a polarized pseudopod and providing the force required for uropod retraction during PMN motility on adhesive surfaces. PMID- 10704380 TI - Mucin-like molecules form a negatively charged coat that protects Trypanosoma cruzi trypomastigotes from killing by human anti-alpha-galactosyl antibodies. AB - In the presence of sialic acid donors Trypanosoma cruzi acquires up to 10(7) sialic acid residues on its surface, in a reaction catalyzed by its unique trans sialidase. Most of these sialic acid residues are incorporated into mucin-like glycoproteins. To further understand the biological role of parasite sialylation, we have measured the amount of mucin in this parasite. We found that both epimastigote and trypomastigote forms have the same number of mucin molecules per surface area, although trypomastigotes have less than 10% of the amount of glycoinositol phospholipids, the other major surface glycoconjugate of T. cruzi. Based on the estimated surface area of each mucin, we calculated that these molecules form a coat covering the entire trypomastigote cell. The presence of the surface coat is shown by transmission electron microscopy of Ruthenium Red stained parasites. The coat was revealed by binding of antibodies isolated from Chagasic patients that react with high affinity to alpha-galactosyl epitopes present in the mucin molecule. When added to the trypomastigote, these antibodies cause an extensive structural perturbation of the parasite coat with formation of large blebs, ultimately leading to parasite lysis. Interestingly, lysis is decreased if the mucin coat is heavily sialylated. Furthermore, addition of MgCl2 reverses the protective effect of sialylation, suggesting that the sialic acid negative charges stabilize the surface coat. Inhibition of sialylation by anti trans-sialidase antibodies, found in immunized animals, or human Chagasic sera, also increase killing by anti-alpha-galactosyl antibodies. Therefore, the large amounts of sialylated mucins, forming a surface coat on infective trypomastigote forms, have an important structural and protective role. PMID- 10704381 TI - A model for anteroposterior patterning of the vertebrate limb based on sequential long- and short-range Shh signalling and Bmp signalling. AB - It has been proposed that digit identity in chick limb bud is specified in a dose dependent fashion by a long-range morphogen, produced by the polarising region. One candidate is Sonic hedgehog (Shh) protein, but it is not clear whether Shh acts long or short range or via Bmps. Here we dissect the relationship between Shh and Bmp signalling. We show that Shh is necessary not only for initiating bmp2 expression but also for sustaining its expression during the period when additional digits are being specified. We also show that we can reproduce much of the effect of Shh during this period by applying only Bmp2. We further demonstrate that it is Bmps that are responsible for digit specification by transiently adding Noggin or Bmp antibodies to limbs treated with Shh. In such limbs, multiple additional digits still form but they all have the same identity. We also explored time dependency and range of Shh signalling by examining ptc expression. We show that high-level ptc expression is induced rapidly when either Shh beads or polarising regions are grafted to a host limb. Furthermore, we find that high-level ptc expression is first widespread but later more restricted. All these data lead us to propose a new model for digit patterning. We suggest that Shh initially acts long range to prime the region of the limb competent to form digits and thus control digit number. Then later, Shh acts short range to induce expression of Bmps, whose morphogenetic action specifies digit identity. PMID- 10704382 TI - Control of hindbrain motor neuron differentiation by the homeobox gene Phox2b. AB - Motor neurons are a widely studied model of vertebrate neurogenesis. They can be subdivided in somatic, branchial and visceral motor neurons. Recent studies on the dorsoventral patterning of the rhombencephalon have implicated the homeobox genes Pax6 and Nkx2.2 in the early divergence of the transcriptional programme of hindbrain somatic and visceral motor neuronal differentiation. We provide genetic evidence that the paired-like homeodomain protein Phox2b is required for the formation of all branchial and visceral, but not somatic, motor neurons in the hindbrain. In mice lacking Phox2b, both the generic and subtype-specific programs of motoneuronal differentiation are disrupted at an early stage. Most motor neuron precursors die inside the neuroepithelium while those that emigrate to the mantle layer fail to switch on early postmitotic markers and to downregulate neuroepithelial markers. Thus, the loss of function of Phox2b in hindbrain motor neurons exemplifies a novel control point in the generation of CNS neurons. PMID- 10704383 TI - Netrin 1 acts as an attractive or as a repulsive cue for distinct migrating neurons during the development of the cerebellar system. AB - Netrin 1 is a long-range diffusible factor that exerts chemoattractive or chemorepulsive effects on developing axons growing to or away from the neural midline. Here we used tissue explants to study the action of netrin 1 in the migration of several cerebellar and precerebellar cell progenitors. We show that netrin 1 exerts a strong chemoattractive effect on migrating neurons from the embryonic lower rhombic lip at E12-E14, which give rise to precerebellar nuclei. Netrin 1 promotes the exit of postmitotic migrating neurons from the embryonic lower rhombic lip and upregulates the expression of TAG-1 in these neurons. In addition, in the presence of netrin 1, the migrating neurons are not isolated but are associated with thick fascicles of neurites, typical of the neurophilic way of migration. In contrast, the embryonic upper rhombic lip, which contains tangentially migrating granule cell progenitors, did not respond to netrin 1. Finally, in the postnatal cerebellum, netrin 1 repels both the parallel fibres and migrating granule cells growing out from explants taken from the external germinal layer. The developmental patterns of expression in vivo of netrin 1 and its receptors are consistent with the notion that netrin 1 secreted in the midline acts as chemoattractive cue for precerebellar neurons migrating circumferentially along the extramural stream. Similarly, the pattern of expression in the postnatal cerebellum suggests that netrin 1 could regulate the tangential migration of postmitotic premigratory granule cells. Thus, molecular mechanisms considered as primarily involved in axonal guidance appear also to steer neuronal cell migration. PMID- 10704384 TI - Ligand endocytosis drives receptor dissociation and activation in the Notch pathway. AB - Endocytosis of the ligand delta; is required for activation of the receptor Notch during Drosophila development. The Notch extracellular domain (NotchECD) dissociates from the Notch intracellular domain (NotchICD) and is trans endocytosed into delta;-expressing cells in wild-type imaginal discs. Reduction of dynamin-mediated endocytosis in developing eye and wing imaginal discs reduces Notch dissociation and Notch signalling. Furthermore, dynamin-mediated delta endocytosis is required for Notch trans-endocytosis in Drosophila cultured cell lines. Endocytosis-defective delta proteins fail to mediate trans-endocytosis of Notch in cultured cells, and exhibit aberrant subcellular trafficking and reduced signalling capacity in Drosophila. We suggest that endocytosis into delta expressing cells of NotchECD bound to delta plays a critical role during activation of the Notch receptor and is required to achieve processing and dissociation of the Notch protein. PMID- 10704385 TI - Murine forkhead/winged helix genes Foxc1 (Mf1) and Foxc2 (Mfh1) are required for the early organogenesis of the kidney and urinary tract. AB - The murine genes, Foxc1 and Foxc2 (previously, Mf1 and Mfh1), encode forkhead/winged helix transcription factors with virtually identical DNA-binding domains and overlapping expression patterns in various embryonic tissues. Foxc1/Mf1 is disrupted in the mutant, congenital hydrocephalus (Foxc1/Mf1(ch)), which has multiple developmental defects. We show here that, depending on the genetic background, most Foxc1 homozygous mutants are born with abnormalities of the metanephric kidney, including duplex kidneys and double ureters, one of which is a hydroureter. Analysis of embryos reveals that Foxc1 homozygotes have ectopic mesonephric tubules and ectopic anterior ureteric buds. Moreover, expression in the intermediate mesoderm of Glial cell-derived neurotrophic factor (Gdnf), a primary inducer of the ureteric bud, is expanded more anteriorly in Foxc1 homozygous mutants compared with wild type. These findings support the hypothesis of Mackie and Stephens concerning the etiology of duplex kidney and hydroureter in human infants with congenital kidney abnormalities (Mackie, G. G. and Stephens, F. G. (1975) J. Urol. 114, 274-280). Previous studies established that most Foxc1(lacZ )Foxc2(tm1) compound heterozygotes have the same spectrum of cardiovascular defects as single homozygous null mutants, demonstrating interaction between the two genes in the cardiovascular system. Here, we show that most compound heterozygotes have hypoplastic kidneys and a single hydroureter, while all heterozygotes are normal. This provides evidence that the two genes interact in kidney as well as heart development. PMID- 10704386 TI - Complementary expression of transmembrane ephrins and their receptors in the mouse spinal cord: a possible role in constraining the orientation of longitudinally projecting axons. AB - In the developing spinal cord, axons project in both the transverse plane, perpendicular to the floor plate, and in the longitudinal plane, parallel to the floor plate. For many axons, the floor plate is a source of long- and short-range guidance cues that govern growth along both dimensions. We show here that B-class transmembrane ephrins and their receptors are reciprocally expressed on floor plate cells and longitudinally projecting axons in the mouse spinal cord. During the period of commissural axon pathfinding, B-class ephrin protein is expressed at the lateral floor plate boundaries, at the interface between the floor plate and the ventral funiculus. In contrast, B-class Eph receptors are expressed on decussated commissural axon segments projecting within the ventral funiculus, and on ipsilaterally projecting axons constituting the lateral funiculus. Soluble forms of all three B-class ephrins bind to, and induce the collapse of, commissural growth cones in vitro. The collapse-inducing activity associated with B-class ephrins is likely to be mediated by EphB1. Taken together, these data support a possible role for repulsive B-class Eph receptor/ligand interactions in constraining the orientation of longitudinal axon projections at the ventral midline. PMID- 10704387 TI - Expression of achaete-scute homologues in discrete proneural clusters on the developing notum of the medfly Ceratitis capitata, suggests a common origin for the stereotyped bristle patterns of higher Diptera. AB - The stereotyped positioning of sensory bristles in Drosophila has been shown to result from complex spatiotemporal regulation of the proneural achaete-scute genes, that relies on an array of cis-regulatory elements and spatially restricted transcriptional activators such as Pannier. Other species of derived schizophoran Diptera have equally stereotyped, but different, bristle patterns. Divergence of bristle patterns could arise from changes in the expression pattern of proneural genes, resulting from evolution of the cis-regulatory sequences and/or altered expression patterns of transcriptional regulators. Here we describe the isolation of achaete-scute homologues in Ceratitis capitata, a species of acalyptrate Schizophora whose bristle pattern differs slightly from that of Drosophila. At least three genes, scute, lethal of scute and asense have been conserved, thus demonstrating that gene duplication within the achaete-scute complex preceded the separation of the families Drosophilidae and Tephritidae, whose common ancestor goes back more than 100 million years. The expression patterns of these genes provide evidence for conservation of many cis-regulatory elements as well as a common origin for the stereotyped patterns seen on the scutum of many Schizophora. Some aspects of the transcriptional regulation have changed, however, and correlate in the notum with differences in the bristle pattern. The Ceratitis pannier gene was isolated and displays a conserved expression domain in the notum. PMID- 10704388 TI - Notch signalling is required for cyclic expression of the hairy-like gene HES1 in the presomitic mesoderm. AB - Somitic segmentation provides the framework on which the segmental pattern of the vertebrae, some muscles and the peripheral nervous system is established. Recent evidence indicates that a molecular oscillator, the 'segmentation clock', operates in the presomitic mesoderm (PSM) to direct periodic expression of c hairy1 and lunatic fringe (l-fng). Here, we report the identification and characterisation of a second avian hairy-related gene, c-hairy2, which also cycles in the PSM and whose sequence is closely related to the mammalian HES1 gene, a downstream target of Notch signalling in vertebrates. We show that HES1 mRNA is also expressed in a cyclic fashion in the mouse PSM, similar to that observed for c-hairy1 and c-hairy2 in the chick. In HES1 mutant mouse embryos, the periodic expression of l-fng is maintained, suggesting that HES1 is not a critical component of the oscillator mechanism. In contrast, dynamic HES1 expression is lost in mice mutant for Delta1, which are defective for Notch signalling. These results suggest that Notch signalling is required for hairy like genes cyclic expression in the PSM. PMID- 10704389 TI - A new function of BMP4: dual role for BMP4 in regulation of Sonic hedgehog expression in the mouse tooth germ. AB - The murine tooth development is governed by sequential and reciprocal epithelial mesenchymal interactions. Multiple signaling molecules are expressed in the developing tooth germ and interact each other to mediate the inductive tissue interactions. Among them are Sonic hedgehog (SHH), Bone Morphogenetic Protein-2 (BMP2) and Bone Morphogenetic Protein-4 (BMP4). We have investigated the interactions between these signaling molecules during early tooth development. We found that the expression of Shh and Bmp2 is downregulated at E12.5 and E13.5 in the dental epithelium of the Msx1 mutant tooth germ where Bmp4 expression is significantly reduced in the dental mesenchyme. Inhibition of BMP4 activity by noggin resulted in repression of Shh and Bmp2 in wild-type dental epithelium. When implanted into the dental mesenchyme of Msx1 mutants, beads soaked with BMP4 protein were able to restore the expression of both Shh and Bmp2 in the Msx1 mutant epithelium. These results demonstrated that mesenchymal BMP4 represents one component of the signal acting on the epithelium to maintain Shh and Bmp2 expression. In contrast, BMP4-soaked beads repressed Shh and Bmp2 expression in the wild-type dental epithelium. TUNEL assay indicated that this suppression of gene expression by exogenous BMP4 was not the result of an increase in programmed cell death in the tooth germ. Ectopic expression of human Bmp4 to the dental mesenchyme driven by the mouse Msx1 promoter restored Shh expression in the Msx1 mutant dental epithelium but repressed Shh in the wild-type tooth germ in vivo. We further demonstrated that this regulation of Shh expression by BMP4 is conserved in the mouse developing limb bud. In addition, Shh expression was unaffected in the developing limb buds of the transgenic mice in which a constitutively active Bmpr-IB is ectopically expressed in the forelimb posterior mesenchyme and throughout the hindlimb mesenchyme, suggesting that the repression of Shh expression by BMP4 may not be mediated by BMP receptor-IB. These results provide evidence for a new function of BMP4. BMP4 can act upstream to Shh by regulating Shh expression in mouse developing tooth germ and limb bud. Taken together, our data provide insight into a new regulatory mechanism for Shh expression, and suggest that this BMP4-mediated pathway in Shh regulation may have a general implication in vertebrate organogenesis. PMID- 10704390 TI - Conditional inactivation of VEGF-A in areas of collagen2a1 expression results in embryonic lethality in the heterozygous state. AB - VEGF-A has been implicated in regulating the initial angiogenic invasion events that are essential for endochondral bone formation. VEGF-A mRNA expression was indeed found in the sclerotome of the developing somite and in the limb-bud mesenchyme at E10.5 in mouse development but declined during chondrogenesis and became upregulated in hypertrophic chondrocytes prior to angiogenic invasion. To determine the functional importance of VEGF-A expression in the developing chondrogenic tissues, VEGF-A was conditionally inactivated during early embryonic development using Collagen2a1-Cre transgenic lines. Deletion of a single VEGF-A allele in Collagen2a1-Cre-expressing cells results in embryonic lethality around E10.5. This lethality is characterized by aberrant development of the dorsal aorta and intersomitic blood vessels, along with defects in the developing endocardial and myocardial layers of the heart. A small percentage of VEGF(Flox)/+, Collagen2a1-Cre fetuses survive until E17.5, show aberrant endochondral bone formation and develop a heart phenotype resembling a dilated form of ischemic cardiomyopathy. These results provide insights into the function of VEGF-A in heart and endochondral bone formation and underscore the importance of tightly controlled levels of VEGF-A during development. PMID- 10704391 TI - HNF1(beta) is required for mesoderm induction in the Xenopus embryo. AB - XHNF1(&bgr;) is a homeobox-containing gene initially expressed at the blastula stage in the vegetal part of the Xenopus embryo. We investigated its early role by functional ablation, through mRNA injection of an XHNF1(beta)/engrailed repressor fusion construct (XHNF1(beta)/EngR). Dorsal injections of XHNF1(beta)/EngR mRNA abolish dorsal mesoderm formation, leading to axial deficiencies; ventral injections disrupt ventral mesoderm formation without affecting axial development. XHNF1(beta)/EngR phenotypic effects specifically depend on the DNA-binding activity of its homeodomain and are fully rescued by coinjection of XHNF1(beta) mRNA. Vegetal injection of XHNF1(beta)/EngR mRNA blocks the mesoderm-inducing ability of vegetal explants. Both B-Vg1 and VegT maternal determinants trigger XHNF1(beta) expression in animal caps. XHNF1(beta)/EngR mRNA blocks B-Vg1-mediated, but not by eFGF-mediated, mesoderm induction in animals caps. However, wild-type XHNF1(beta) mRNA does not trigger Xbra expression in animal caps. We conclude that XHNF1(beta) function is essential, though not sufficient, for mesoderm induction in the Xenopus embryo. PMID- 10704392 TI - The C. elegans par-4 gene encodes a putative serine-threonine kinase required for establishing embryonic asymmetry. AB - During the first cell cycle of Caenorhabditis elegans embryogenesis, asymmetries are established that are essential for determining the subsequent developmental fates of the daughter cells. The maternally expressed par genes are required for establishing this polarity. The products of several of the par genes have been found to be themselves asymmetrically distributed in the first cell cycle. We have identified the par-4 gene of C. elegans, and find that it encodes a putative serine-threonine kinase with similarity to a human kinase associated with Peutz Jeghers Syndrome, LKB1 (STK11), and a Xenopus egg and embryo kinase, XEEK1. Several strong par-4 mutant alleles are missense mutations that alter conserved residues within the kinase domain, suggesting that kinase activity is essential for PAR-4 function. We find that the PAR-4 protein is present in the gonads, oocytes and early embryos of C. elegans, and is both cytoplasmically and cortically distributed. The cortical distribution begins at the late 1-cell stage, is more pronounced at the 2- and 4-cell stages and is reduced at late stages of embryonic development. We find no asymmetry in the distribution of PAR 4 protein in C. elegans embryos. The distribution of PAR-4 protein in early embryos is unaffected by mutations in the other par genes. PMID- 10704393 TI - Depolarisation causes reciprocal changes in GFR(alpha)-1 and GFR(alpha)-2 receptor expression and shifts responsiveness to GDNF and neurturin in developing neurons. AB - GDNF and neurturin are structurally related neurotrophic factors that promote the survival of many different kinds of neurons and influence axonal and dendritic growth and synaptic function. These diverse effects are mediated via multicomponent receptors consisting of the Ret receptor tyrosine kinase plus one of two structurally related GPI-linked receptors, GFR(alpha)-1 and GFR(alpha)-2. To ascertain how the expression of these receptors is regulated during development, we cultured embryonic neurons under different experimental conditions and used competitive RT/PCR to measure the levels of the mRNAs encoding these receptors. We found that depolarising levels of KCl caused a marked increase in GFR(alpha)-1 mRNA and a marked decrease in GFR(&agr;)-2 mRNA in sympathetic, parasympathetic and sensory neurons. These changes were accompanied by increased responsiveness to GDNF and decreased responsiveness to neurturin, and were inhibited by L-type Ca(2+) channel antagonists, suggesting that they were due to elevated intracellular free-Ca(2+). There was no consistent effect of depolarising levels of KCl on ret mRNA expression, and neither GDNF nor neurturin significantly affected receptor expression. These results show that depolarisation has marked and opposing actions on the expression of GFR(&agr;)-1 and GFR(&agr;)-2, which are translated into corresponding changes in neuronal responsiveness to GDNF and neurturin. This provides evidence for a mechanism of regulating the neurotrophic factor responses of neurons by neural activity that has important implications for structural and functional plasticity in the developing nervous system. PMID- 10704394 TI - Increased cell death and delayed development in the cerebellum of mice lacking the rev-erbA(alpha) orphan receptor. AB - The rev-erbA(alpha) gene, belonging to the steroid receptor superfamily of transcription factors, is highly conserved during evolution but little is known so far about its functions in development or in adult physiology. Here, we describe genetically altered mice lacking the rev-erbA(alpha) gene. These animals do not show any obvious phenotype in either fat tissue or skeletal muscle, despite the known regulation of rev-erbA(alpha) expression during adipocyte and myotube differentiation in vitro. However, during the second week of life, the cerebellum of rev-erbA(alpha) mutants presents several unexpected abnormalities, such as alterations in the development of Purkinje cells, delay in the proliferation and migration of granule cells from the external granule cell layer and increased apoptosis of neurons in the internal granule cell layer. Interestingly, the expression pattern of rev-erbA(alpha) suggests that the abnormalities observed in the external granule cell layer could be secondary to Purkinje cell alterations. Taken together, our data underline the importance of rev-erbA(alpha)expression for the appropriate balance of transcriptional activators and repressors during postnatal cerebellar development. PMID- 10704395 TI - A dual role for homothorax in inhibiting wing blade development and specifying proximal wing identities in Drosophila. AB - The Drosophila wing imaginal disc gives rise to three body parts along the proximo-distal (P-D) axis: the wing blade, the wing hinge and the mesonotum. Development of the wing blade initiates along part of the dorsal/ventral (D/V) compartment boundary and requires input from both the Notch and wingless (wg) signal transduction pathways. In the wing blade, wg activates the gene vestigial (vg), which is required for the wing blade to grow. wg is also required for hinge development, but wg does not activate vg in the hinge, raising the question of what target genes are activated by wg to generate hinge structures. Here we show that wg activates the gene homothorax (hth) in the hinge and that hth is necessary for hinge development. Further, we demonstrate that hth also limits where along the D/V compartment boundary wing blade development can initiate, thus helping to define the size and position of the wing blade within the disc epithelium. We also show that the gene teashirt (tsh), which is coexpressed with hth throughout most of wing disc development, collaborates with hth to repress vg and block wing blade development. Our results suggest that tsh and hth block wing blade development by repressing some of the activities of the Notch pathway at the D/V compartment boundary. PMID- 10704396 TI - Local, possibly contact-mediated signalling restricted to homotypic neurons controls the regular spacing of cells within the cholinergic arrays in the developing rodent retina. AB - In the vertebrate retina neurons of the same type commonly form non-random arrays, assembled by unknown positional mechanisms during development. Computational models in which no two cells are closer than a minimal distance, simulate many retinal arrays. These findings have important biological implications, since they suggest that cells are determined as neurons of specific types before entering their arrays, and that local, possibly contact-mediated interactions acting exclusively among the elements of an array account for its assembly. This is here verified by combining experimental manipulations in normal and transgenic models with computational analysis for the cholinergic mosaics, the only arrays so far for which the development of spatial ordering is known quantitatively. When generalised, these findings suggest a plan for vertebrate retinal patterning, where homotypic interactions organise retinal arrays first, then local interactions between synaptic partners suffice to establish the topographical connections that support retinal processing. PMID- 10704397 TI - split ends encodes large nuclear proteins that regulate neuronal cell fate and axon extension in the Drosophila embryo. AB - split ends (spen) encodes nuclear 600 kDa proteins that contain RNA recognition motifs and a conserved C-terminal sequence. These features define a new protein family, Spen, which includes the vertebrate MINT transcriptional regulator. Zygotic spen mutants affect the growth and guidance of a subset of axons in the Drosophila embryo. Removing maternal and zygotic protein elicits cell-fate and more general axon-guidance defects that are not seen in zygotic mutants. The wrong number of chordotonal neurons and midline cells are generated, and we identify defects in precursor formation and EGF receptor-dependent inductive processes required for cell-fate specification. The number of neuronal precursors is variable in embryos that lack Spen. The levels of Suppressor of Hairless, a key transcriptional effector of Notch required for precursor formation, are reduced, as are the nuclear levels of Yan, a transcriptional repressor that regulates cell fate and proliferation downstream of the EGF receptor. We propose that Spen proteins regulate the expression of key effectors of signaling pathways required to specify neuronal cell fate and morphology. PMID- 10704398 TI - Transcriptional regulation of atonal required for Drosophila larval eye development by concerted action of eyes absent, sine oculis and hedgehog signaling independent of fused kinase and cubitus interruptus. AB - Bolwig's organ is the larval light-sensing system consisting of 12 photoreceptors and its development requires atonal activity. Here, we showed that Bolwig's organ formation and atonal expression are controlled by the concerted function of hedgehog, eyes absent and sine oculis. Bolwig's organ primordium was first detected as a cluster of about 14 Atonal-positive cells at the posterior edge of the ocular segment in embryos and hence, atonal expression may define the region from which a few Atonal-positive founder cells (future primary photoreceptor cells) are generated by lateral specification. In Bolwig's organ development, neural differentiation precedes photoreceptor specification, since Elav, a neuron specific antigen, whose expression is under the control of atonal, is expressed in virtually all early-Atonal-positive cells prior to the establishment of founder cells. Neither Atonal expression nor Bolwig's organ formation occurred in the absence of hedgehog, eyes absent or sine oculis activity. Genetic and histochemical analyses indicated that (1) responsible Hedgehog signals derive from the ocular segment, (2) Eyes absent and Sine oculis act downstream of or in parallel with Hedgehog signaling and (3) the Hedgehog signaling pathway required for Bolwig's organ development is a new type and lacks Fused kinase and Cubitus interruptus as downstream components. PMID- 10704399 TI - A developmentally regulated GAGA box-binding factor and Sp1 are required for transcription of the hsp70.1 gene at the onset of mouse zygotic genome activation. AB - We have investigated the onset of zygotic genome transcription in early two-cell mouse embryos by analyzing the regulation of hsp70.1, one of the first genes expressed after fertilization. The transcriptional activation of both an episomic hsp70 promoter and the endogenous hsp70.1 gene requires the contiguity of the GC box proximal to the TATA box with a GAGA box and involves GC box- and GAGA box binding factors. In vivo transcription factor titrations with double-stranded oligodeoxyribonucleotides and antibodies pinpoint these factors as Sp1 and a novel murine GAGA box-binding factor, which is structurally related to the Drosophila GAGA factor and acts as transcriptional coactivator/potentiator of Sp1. Mouse unfertilized eggs and one-cell and two-cell embryos display a GAGA box binding activity of maternal origin that disappears at the four-cell stage and is also abundant in the gonads, but is barely detectable in other adult tissues. In light of the well-established nucleosome-disruption role of the Drosophila GAGA factor, these findings suggest a novel mechanism of enhancer-independent gene derepression in early mouse embryos. PMID- 10704400 TI - Cell death: drosophila Apaf-1 - no longer in the (d)Ark. AB - The recent discovery and characterization of Ark, the Drosophila homolog of the mammalian cell-death adaptor protein Apaf-1, have revealed that, like Apaf-1, this protein is important in multiple apoptosis pathways. The new findings also suggest that cell death in flies is very similar to that in mammals after all. PMID- 10704401 TI - Planar polarity: out of joint? AB - Epithelial structures, such as the wing hairs and ommatidia in Drosophila, are aligned in the plane of the epithelium. This planar polarity requires the transmembrane receptor Frizzled. Recent studies have shed new light on mechanisms that could be involved in generating or transducing the polarity signal. PMID- 10704402 TI - Left-right development: the roles of nodal cilia. AB - Cilia on the ventral side of the mouse node have been implicated in initiating the left-right axis during embryonic development, but how cilia relate to other factors in the left-right pathway and the mechanism by which cilia convey patterning information remain uncertain. PMID- 10704403 TI - Bacterial sporulation: pole-to-pole protein oscillation. AB - Sporulating bacteria need to temporally coordinate DNA replication, chromosome partitioning and sporulation initiation. Recent work has shown that one aspect of this coordination lies with the interdependent subcellular localization of two proteins, Spo0J and Soj, and in the Spo0J-dependent spatial oscillation of Soj. PMID- 10704404 TI - Odorant receptors: axon contact-mediated diversity. AB - To make the most of a small number of neurons, the nematode olfactory system includes neurons that are bilaterally symmetrical in morphology, but differ in the sets of genes they express. An intriguing recent example is the axon contact mediated asymmetry in expression of the str-2 odorant receptor gene. PMID- 10704405 TI - Cortical development: receiving reelin. AB - Recent genetic and biochemical studies indicate that lipoprotein receptors are components of the neuronal receptor for Reelin, mediating the glycoprotein's essential function in cortical development. At least eight cadherin-related neuronal receptors may also play a part in this signalling system. PMID- 10704406 TI - Membrane traffic: do cones mark sites of fission? AB - Membrane fission occurs in eukaryotic cells whenever a vesicle is produced or a larger subcellular compartment is divided into smaller discrete units. Recent evidence suggests this fission event is promoted by enzymes that generate phosphatidic acid and thereby cause a distortion of the lipid bilayer. PMID- 10704407 TI - Gene silencing: shrinking the black box of RNAi. AB - The mysterious mechanism whereby the mere presence of double-stranded RNA can inactivate specific genes is yielding its secrets. Recent results identifying cellular components required for RNAi in Caenorhabditis elegans indicate that the mechanism is conserved, ancient and may provide a defense against selfish DNA. PMID- 10704408 TI - Visuo-motor control: giving the brain a hand. AB - Sensory information is acquired in spatial coordinate systems linked to sense organs, yet movement must be executed in coordinate systems linked to motor effector organs. Neurophysiological experiments are yielding new insights into how the brain transforms coordinate systems to facilitate movement. PMID- 10704409 TI - Ectopic hypermethylation of flower-specific genes in Arabidopsis. AB - BACKGROUND: Arabidopsis mutations causing genome-wide hypomethylation are viable but display a number of specific developmental abnormalities, including some that resemble known floral homeotic mutations. We previously showed that one of the developmental abnormalities present in an antisense-METHYLTRANSFERASEI (METI) transgenic line resulted from ectopic hypermethylation of the SUPERMAN gene. RESULTS: Here, we investigate the extent to which hypermethylation of SUPERMAN occurs in several hypomethylation mutants, and describe methylation effects at a second gene, AGAMOUS. SUPERMAN gene hypermethylation occurred at a high frequency in several mutants that cause overall decreases in genomic DNA methylation. The hypermethylation pattern was largely similar in the different mutant backgrounds. Genetic analysis suggests that hypermethylation most likely arose either during meiosis or somatically in small sectors of the plant. A second floral development gene, AGAMOUS, also became hypermethylated and silenced in an Arabidopsis antisense-METI line. CONCLUSIONS: These results suggest that ectopic hypermethylation of specific genes in mutant backgrounds that show overall decreases in methylation may be a widespread phenomenon that could explain many of the developmental defects seen in Arabidopsis methylation mutants. This resembles a phenomenon seen in cancer cells, which can simultaneously show genome wide hypomethylation and hypermethylation of specific genes. Comparison of the methylated sequences in SUPERMAN and AGAMOUS suggests that hypermethylation could involve DNA secondary structures formed by pyrimidine-rich sequences. PMID- 10704410 TI - Clb/Cdc28 kinases promote nuclear export of the replication initiator proteins Mcm2-7. AB - BACKGROUND: In the budding yeast Saccharomyces cerevisiae, the cyclin-dependent kinases of the Clb/Cdc28 family restrict the initiation of DNA replication to once per cell cycle by preventing the re-assembly of pre-replicative complexes (pre-RCs) at replication origins that have already initiated replication. This assembly involves the Cdc6-dependent loading of six minichromosome maintenance (Mcm) proteins, Mcm2-7, onto origins. How Clb/Cdc28 kinases prevent pre-RC assembly is not understood. RESULTS: In living cells, the Mcm proteins were found to colocalize in a cell-cycle-regulated manner. Mcm2-4, 6 and 7 were concentrated in the nucleus in G1 phase, gradually exported to the cytoplasm during S phase, and excluded from the nucleus by G2 and M phase. Tagging any single Mcm protein with the SV40 nuclear localization signal made all Mcm proteins constitutively nuclear. In the absence of functional Cdc6, Clb/Cdc28 kinases were necessary and sufficient for efficient net nuclear export of a fusion protein between Mcm7 and the green fluorescent protein (Mcm7-GFP), whereas inactivation of these kinases at the end of mitosis coincided with the net nuclear import of Mcm7-GFP. In contrast, in the presence of functional Cdc6, which loads Mcm proteins onto chromatin, S-phase progression as well as Clb/Cdc28 kinases was required for Mcm GFP export. CONCLUSIONS: We propose that Clb/Cdc28 kinases prevent pre-RC reassembly in part by promoting the net nuclear export of Mcm proteins. We further propose that Mcm proteins become refractory to this regulation when they load onto chromatin and must be dislodged by DNA replication before they can be exported. Such an arrangement could ensure that Mcm proteins complete their replication function before they are removed from the nucleus. PMID- 10704411 TI - Dopamine modulates acute responses to cocaine, nicotine and ethanol in Drosophila. AB - BACKGROUND: Drugs of abuse have a common property in mammals, which is their ability to facilitate the release of the neurotransmitter and neuromodulator dopamine in specific brain regions involved in reward and motivation. This increase in synaptic dopamine levels is believed to act as a positive reinforcer and to mediate some of the acute responses to drugs. The mechanisms by which dopamine regulates acute drug responses and addiction remain unknown. RESULTS: We present evidence that dopamine plays a role in the responses of Drosophila to cocaine, nicotine or ethanol. We used a startle-induced negative geotaxis assay and a locomotor tracking system to measure the effect of psychostimulants on fly behavior. Using these assays, we show that acute responses to cocaine and nicotine are blunted by pharmacologically induced reductions in dopamine levels. Cocaine and nicotine showed a high degree of synergy in their effects, which is consistent with an action through convergent pathways. In addition, we found that dopamine is involved in the acute locomotor-activating effect, but not the sedating effect, of ethanol. CONCLUSIONS: We show that in Drosophila, as in mammals, dopaminergic pathways play a role in modulating specific behavioral responses to cocaine, nicotine or ethanol. We therefore suggest that Drosophila can be used as a genetically tractable model system in which to study the mechanisms underlying behavioral responses to multiple drugs of abuse. PMID- 10704412 TI - EGO-1 is related to RNA-directed RNA polymerase and functions in germ-line development and RNA interference in C. elegans. AB - BACKGROUND: Cell-fate determination requires that cells choose between alternative developmental pathways. For example, germ cells in the nematode worm Caenorhabditis elegans choose between mitotic and meiotic division, and between oogenesis and spermatogenesis. Germ-line mitosis depends on a somatic signal that is mediated by a Notch-type signaling pathway. The ego-1 gene was originally identified on the basis of genetic interactions with the receptor in this pathway and was also shown to be required for oogenesis. Here, we provide more insight into the role of ego-1 in germ-line development. RESULTS: We have determined the ego-1 gene structure and the molecular basis of ego-1 alleles. Putative ego-1 null mutants had multiple, previously unreported defects in germ-line development. The ego-1 transcript was found predominantly in the germ line. The predicted EGO-1 protein was found to be related to the tomato RNA-directed RNA polymerase (RdRP) and to Neurospora crassa QDE-1, two proteins implicated in post transcriptional gene silencing (PTGS). For a number of germ-line-expressed genes, ego-1 mutants were resistant to a form of PTGS called RNA interference. CONCLUSIONS: The ego-1 gene is the first example of a gene encoding an RdRP related protein with an essential developmental function. The ego-1 gene is also required for a robust response to RNA interference by certain genes. Hence, a protein required for germ-line development in C. elegans may be a component of the RNA interference/PTGS machinery. PMID- 10704413 TI - A conserved function for Arabidopsis SUPERMAN in regulating floral-whorl cell proliferation in rice, a monocotyledonous plant. AB - Studies of floral organ development in two dicotyledonous plants, Arabidopsis thaliana and Antirrhinum majus, have shown that three sets of genes (A, B and C) can pattern sepals, petals, stamens and carpels [1] [2]. Mechanisms that define boundaries between these floral whorls are unclear, however. The Arabidopsis gene SUPERMAN (SUP), which encodes a putative transcription factor, maintains the boundary between stamens and carpels [3] [4] [5], possibly by regulating cell proliferation. By overexpressing SUP cDNA in rice, we examined whether its effects on whorl boundaries are conserved in a divergent monocotyledonous species. High-level ectopic SUP expression in transgenic rice resulted in juvenile death or dwarf plants with decreased axillary growth. Plants with lower levels of SUP RNA were vegetatively normal, but the flowers showed ubiquitous ventral carpel expansion. This was often coupled with reduced stamen number, or occurrence of third-whorl stamen-carpel mosaic organs. Additionally, proliferation of second-whorl ventral cells produced adventitious lodicules, and flowers lost the asymmetry that is normally inherent to this whorl. We predict that SUP is a conserved regulator of floral whorl boundaries and that it affects cell proliferation. PMID- 10704414 TI - Olfactory neurons are interdependent in maintaining axonal projections. AB - In mice, individual olfactory neurons express one of the thousand distinct olfactory receptor genes [1] [2] [3]. Neurons that express a given receptor converge on distinct loci in the olfactory bulb to form structures called glomeruli [4] [5] [6]. The olfactory receptor is involved in an instructive manner in this axonal convergence [6] [7] but little is known about the mechanisms involved in maintaining convergence. We have previously created a transgenic olfactory receptor locus that functions independently of the endogenous loci [8]. Here, we show that, although the projections of neurons expressing this ectopic transgenic olfactory receptor always converge in newborn mice, surprisingly, in adult mice, convergence is not always maintained. Moreover, in adult mice there is a positive correlation between the number of neurons expressing the transgenic receptor and the probability of maintaining convergence. These observations, taken together with the variability observed in wild-type [4] [6] and genetically manipulated mice ([6] and our unpublished observations), suggest that olfactory neurons require the presence of other similar axons to maintain a glomerulus. We call this phenomenon interdependence. PMID- 10704415 TI - Selective high-affinity transport of aspartate by a Drosophila homologue of the excitatory amino-acid transporters. AB - Excitatory amino-acid transporters (EAATs) are structurally related plasma membrane proteins that mediate the high-affinity uptake of the acidic amino acids glutamate and aspartate released at excitatory synapses, and maintain the extracellular concentrations of these neurotransmitters below excitotoxic levels [1] [2] [3] [4]. Several members of the EAAT family have been described previously. So far, all known EAATs have been reported to transport glutamate and aspartate with a similar affinity. Here, we report that dEAAT2 - a nervous tissue specific EAAT homologue that we recently identified in the fruit fly Drosophila [5] - is a selective Na(+)-dependent high-affinity aspartate transporter (K(m) = 30 microM). We found that dEAAT2 can also transport L-glutamate but with a much lower affinity (K(m) = 185 microM) and a 10- to 15-fold lower relative efficacy (V(max)/K(m)). Competition experiments showed that the binding of glutamate to this transporter is much weaker than the binding of D- or L-aspartate. As dEAAT2 is the first known EAAT to show this substrate selectivity, it suggests that aspartate may play a specific role in the Drosophila nervous system. PMID- 10704416 TI - NURD-complex genes antagonise Ras-induced vulval development in Caenorhabditis elegans. AB - Chromatin-modifying complexes are important for transcriptional control, but their roles in the regulation of development are poorly understood. Here, we show that components of the nucleosome remodelling and histone deacetylase (NURD) complex [1] [2] [3] [4] [5] antagonise vulval development, which is induced by the Ras signal transduction pathway. In three of the six equivalent vulval precursor cells, the Ras pathway is active, leading to the production of vulval fates [6]; in the remaining three, the Ras pathway is inhibited and vulval fates repressed. Inhibition of Ras signaling occurs in part through the action of the synthetic multivulval (synMuv) genes, which comprise two functionally redundant pathways (synMuvA and synMuvB) [7]. We found that five Caenorhabditis elegans members of the NURD chromatin remodelling complex inhibit vulval development through both the synMuvA and synMuvB pathways (hda-1, rba-1, lin-53, chd-3 and chd-4); a further two members, the MTA1-related genes egr-1 and egl-27, act only in the synMuvA pathway. We propose that the synMuvA and synMuvB pathways function redundantly to recruit or activate a core NURD complex, which then represses vulval developmental target genes by local histone deacetylation. These results emphasise the importance of chromatin regulation in developmental decisions. Furthermore, inhibition of Ras signaling suggests a possible link between NURD function and cancer. PMID- 10704417 TI - Ectopic G-protein expression in dopamine and serotonin neurons blocks cocaine sensitization in Drosophila melanogaster. AB - Sensitization to repeated doses of psychostimulants is thought to be an important component underlying the addictive process in humans [1] [2] [3] [4]. In all vertebrate animal models, including humans [5], and even in fruit flies, sensitization is observed after repeated exposure to volatilized crack cocaine [6]. In vertebrates, sensitization is thought to be initiated by processes occurring in brain regions that contain dopamine cell bodies [2] [7]. Here, we show that modulated cell signaling in the Drosophila dopamine and serotonin neurons plays an essential role in cocaine sensitization. Targeted expression of either a stimulatory (Galpha(s)) or inhibitory (Galpha(i)) Galpha subunit, or tetanus toxin light chain (TNT) in dopamine and serotonin neurons of living flies blocked behavioral sensitization to repeated cocaine exposures. These flies showed alterations in their initial cocaine responsiveness that correlated with compensatory adaptations of postsynaptic receptor sensitivity. Finally, repeated drug stimulation of a nerve cord preparation that is postsynaptic to the brain amine cells failed to induce sensitization, further showing the importance of presynaptic modulation in sensitization. PMID- 10704418 TI - Autoimmune-prone mice share a promoter haplotype associated with reduced expression and function of the Fc receptor FcgammaRII. AB - Human autoimmune diseases thought to arise from the combined effects of multiple susceptibility genes include systemic lupus erythematosus (SLE) and autoimmune diabetes. Well-characterised polygenic mouse models closely resembling each of these diseases exist, and genetic evidence links receptors for the Fc portion of immunoglobulin G (FcR) with their pathogenesis in mice and humans [1] [2] [3]. FcRs may be activatory or inhibitory and regulate a variety of immune and inflammatory processes [4] [5]. FcgammaRII (CD32) negatively regulates activation of cells including B cells and macrophages [6]. FcgammaRII-deficient mice are prone to immune-mediated disease [7] [8] [9]. The gene encoding FcgammaRII, Fcgr2, is contained in genetic susceptibility intervals in mouse models of SLE such as the New Zealand Black (NZB) contribution to the (NZB x New Zealand White (NZW)) F1 strain [1] [10] [11] and the BXSB strain [12], and in human SLE [1] [2] [3]. We therefore sequenced Fcgr2 and identified a haplotype defined by deletions in the Fcgr2 promoter region that is present in major SLE-prone mouse strains (NZB, BXSB, SB/Le, MRL, 129 [13]) and non-obese diabetic (NOD) mice but absent in control strains (BALB/c, C57BL/6, DBA/2, C57BL/10) and NZW mice. The autoimmune haplotype was associated with reduced cell-surface expression of FcgammaRII on macrophages and activated B cells and with hyperactive macrophages resembling those of FcgammaRII-deficient mice, and is therefore likely to play an important role in the pathogenesis of SLE and possibly diabetes. PMID- 10704419 TI - Binocular rivalry and perceptual coherence. PMID- 10704421 TI - Good eggs and bad eggs. PMID- 10704420 TI - Hard cases. PMID- 10704422 TI - The amygdala. PMID- 10704424 TI - Chips 'n' DNA. PMID- 10704423 TI - The U box is a modified RING finger - a common domain in ubiquitination. PMID- 10704425 TI - Frankencells and mirth. PMID- 10704426 TI - Recent advances in the understanding of genetic causes of congenital heart defects. AB - The clinical approach to children with congenital heart defects (CHD) has been revolutionized during the past four decades by developments in diagnostics and therapeutics. In contrast, a profound understanding of the causes of the majority of CHD has only begun to emerge within the past few years. Prior epidemiological studies suggested that Mendelian disorders constituted a very small percentage of CHD and that polygenic inheritance was responsible for the majority of cases. Recent discoveries, largely achieved with molecular genetic studies, have provided new insights into the genetic basis of heart malformations. These studies have shown that CHD caused by single gene or single locus defects is more common than had been suspected. In addition, a higher percentage of heart malformations occur in the context of familial disease than was evident previously. In this review, molecular genetic studies of specific heart lesions and syndromes with CHD are reviewed. Progress on the Human Genome Project has accelerated identification of genes for Mendelian traits with heart defects, and it is anticipated that disease genes for most single gene traits will be known within a few years. Future challenges include utilizing this emerging genetic information to improve diagnosis and treatment of children with CHD, and harnessing the power of genomics to analyze isolated heart defects with complex inheritance patterns. PMID- 10704427 TI - Cyclin D1 in parathyroid disease. AB - Primary hyperparathyroidism (HPT), most commonly due to parathyroid adenoma, is a disorder characterized by excessive secretion of PTH. So far, abnormalities in two genes, cyclin D1 and MEN1, have been implicated in the development of parathyroid adenomas. Cyclin D1, now an established Oncogene involved in numerous human cancers, was first identified and recognized as an Oncogene in the study of parathyroid tumors. A subset of parathyroid adenomas contains a clonal rearrangement that places the PTH gene's regulatory sequences in proximity to the cyclin D1 Oncogene causing its overexpression, and 20-40% of parathyroid adenomas overexpress the cyclin D1 protein. Transgenic animal models have further confirmed the role of cyclin D1 as a driver of abnormal parathyroid cell proliferation. Future studies on the mechanism of cyclin D1's oncogenicity and its interactions with other parathyroid growth regulators will further our understanding of parathyroid cell biology and may prove useful clinically. PMID- 10704428 TI - Spongy degeneration of the brain, Canavan disease: biochemical and molecular findings. AB - Canavan disease is a severe progressive leukodystrophy characterized by swelling and spongy degeneration of the white matter of the brain. It is an autosomal recessive disease found more frequently among Ashkenazi Jews. The clinical features are those of severe mental retardation with inability to gain developmental milestones. Hypotonia, head lag and macrocephaly are characteristic of Canavan disease and become apparent after 5-6 months of age. Massive excretion in the urine of N-acetylaspartic acid is the biochemical marker for Canavan disease, which is caused by deficiency of the enzyme aspartoacylase. This discovery allowed for accurate diagnosis of Canavan disease, while prior to that, a brain biopsy was needed. The gene for aspartoacylase has been cloned and two mutations predominate among Ashkenazi Jewish individuals with Canavan disease and account for more than 98% of the Ashkenazi Jewish patients. The mutations among other ethnic groups are more diverse. The carrier frequency for the two common mutations among Ashkenazi Jews was found to be surprisingly high, 1:37. Screening for carriers is now common practice for this population. A knock-out mouse for Canavan disease is being genetically engineered in our laboratory. The mouse model will allow for development of strategies for gene therapy. PMID- 10704429 TI - Molecular genetics of peroxisomal disorders. AB - Twenty five human peroxisomal disorders have been defined at this time. They are subdivided into two major categories: 1) the disorders of peroxisome biogenesis, in which the organelle fails to form normally, and there are defects that involve multiple peroxisomal functions; and 2) disorders that affect single peroxisomal enzymes. During the last five years the molecular defects have been identified in nearly all. These recent advances have several important implications. They have facilitated diagnosis of affected patients. The improved capacity to provide prenatal diagnosis and heterozygote identification has been of great value for genetic counseling and disease prevention. Study of genotype-phenotype correlations has led to a new and more rational classification system. The identification of the molecular defects and the development of animal models have increased understanding of pathogenetic mechanisms, and have led to novel therapeutic approaches. PMID- 10704430 TI - Molecular genetics of holoprosencephaly. AB - Holoprosencephaly (HPE) is a common developmental defect of the human forebrain and midface. Pathological studies have identified different categories of severity of the brain and craniofacial malformations observed in HPE, although the variable clinical spectrum of HPE extends in unbroken sequence from alobar HPE and cyclopia to clinically unaffected carriers in familial HPE. The etiology of HPE is extremely heterogeneous including both environmental and genetic causes. Here we focus on molecular aspects of HPE in light of the recent identification of some of the genes causing human HPE and other candidate genes involved in forebrain development, through different approaches, such as positional cloning and functional cloning, based on animal models. These approaches will aid in the identification of additional genes involved in HPE and in a better understanding of the molecular genetics of brain development. PMID- 10704431 TI - Genetics of Carney complex and related familial lentiginoses, and other multiple tumor syndromes. AB - Carney complex is a multiple endocrine neoplasia (MEN) syndrome that affects the adrenal cortex, the pituitary and thyroid glands, and the gonads. The complex is also associated with skin and mucosa pigmentation abnormalities and myxoid and other neoplasms of mesenchymal and neural crest origin. Thus, this syndrome also belongs to another group of genetic disorders, the lentiginoses (or lentigenoses), which include the Peutz-Jeghers, LEOPARD, arterial dissections and lentiginosis, and Laugier-Hunziker syndromes, Cowden disease and Ruvalcaba-Myhre Smith (Bannayan-Zonana) syndrome and the centrofacial, benign patterned and segmental lentiginoses, all of which can be associated with a variety of developmental defects. The inheritance of Carney complex, just like that of the other MENs and the lentiginoses, is autosomal dominant. Genetic loci or genes have been identified for Carney complex, Peutz-Jeghers and Ruvalcaba-Myhre-Smith syndromes, but not for other lentiginoses. Elucidation of the molecular defects responsible for these disorders is expected to shed light on aspects of early neural crest differentiation, the regulation of pigmentation, the development of autonomous endocrine function, and endocrine and nonendocrine tumorigenesis. PMID- 10704432 TI - Progress in molecular genetics of autosomal dominant polycystic kidney disease. AB - Among the prevalent human genetic disorders, human autosomal dominant polycystic kidney disease is certainly one of the most challenging, both from a clinical and a fundamental perspective. In the recent years, important progress opened novel research avenues to elucidate the genetic basis, the cellular pathophysiologic mechanisms and the molecular function of genes and proteins involved in autosomal dominant polycystic kidney disease. PMID- 10704433 TI - Luteinizing hormone receptor mutations in disorders of sexual development and cancer. AB - Human male sexual development is regulated by chorionic gonadotropin (CG) and luteinizing hormone (LH). Aberrant sexual development caused by both activating and inactivating mutations of the human luteinizing hormone receptor (LHR) have been described. All known activating mutations of the LHR are missense mutations caused by single base substitution. The most common activating mutation is the replacement of Asp-578 by Gly due to the substitution of A by G at nucleotide position 1733. All activating mutations are present in exon 11 which encodes the transmembrane domain of the receptor. Constitutive activity of the LHR causes LH releasing hormone-independent precocious puberty in boys and the autosomal dominant disorder familial male-limited precocious puberty (FMPP). Both germline and somatic activating mutations of the LHR have been found in patients with testicular tumors. Activating mutations have no effect on females. The molecular genetics of the inactivating mutations of the LHR are more variable and include single base substitution, partial gene deletion, and insertion. These mutations are not localized and are present in both the extracellular and transmembrane domain of the receptor. Inactivation of the LHR gives rise to the autosomal recessive disorder Leydig cell hypoplasia (LCH) and male hypogonadism or male pseudohermaphroditism. Severity of the clinical phenotype in LCH patients correlates with the amount of residual activity of the mutated receptor. Females are less affected by inactivating mutation of the LHR. Symptoms caused by homozygous inactivating mutation of the LHR include polycystic ovaries and primary amenorrhea. PMID- 10704434 TI - Precursor cell transitions in oligodendrocyte development. PMID- 10704435 TI - Untying the Gordian knot of cytokinesis. Role of small G proteins and their regulators. PMID- 10704436 TI - Nuclear export of MAP kinase (ERK) involves a MAP kinase kinase (MEK)-dependent active transport mechanism. AB - In response to extracellular stimuli, mitogen-activated protein kinase (MAPK, also known as ERK), which localizes to the cytoplasm in quiescent cells, translocates to the nucleus and then relocalizes to the cytoplasm again. The relocalization of nuclear MAPK to the cytoplasm was not inhibited by cycloheximide, confirming that the relocalization is achieved by nuclear export, but not synthesis, of MAPK. The nuclear export of MAPK was inhibited by leptomycin B (LMB), a specific inhibitor of the nuclear export signal (NES) dependent transport. We have then shown that MAP kinase kinase (MAPKK, also known as MEK), which mostly localizes to the cytoplasm because of its having NES, is able to shuttle between the cytoplasm and the nucleus constantly. MAPK, when injected into the nucleus, was rapidly exported from the nucleus by coinjected wild-type MAPKK, but not by the NES-disrupted MAPKK. In addition, injection of the fragment corresponding to the MAPK-binding site of MAPKK into the nucleus, which would disrupt the binding of MAPK to MAPKK in the nucleus, significantly inhibited the nuclear export of endogenous MAPK. Taken together, these results suggest that the relocalization of nuclear MAPK to the cytoplasm involves a MAPKK dependent, active transport mechanism. PMID- 10704437 TI - Reduced loading of intracellular Ca(2+) stores and downregulation of capacitative Ca(2+) influx in Bcl-2-overexpressing cells. AB - The mechanism of action of the oncogene bcl-2, a key regulator of the apoptotic process, is still debated. We have employed organelle-targeted chimeras of the Ca(2+)-sensitive photoprotein, aequorin, to investigate in detail the effect of Bcl-2 overexpression on intracellular Ca(2+) homeostasis. In the ER and the Golgi apparatus, Bcl-2 overexpression increases the Ca(2+) leak (while leaving Ca(2+) accumulation unaffected), hence reducing the steady-state [Ca(2+)] levels. As a direct consequence, the [Ca(2+)] increases caused by inositol 1,4,5 trisphosphate (IP3)-generating agonists were reduced in amplitude in both the cytosol and the mitochondria. Bcl-2 overexpression also reduced the rate of Ca(2+) influx activated by Ca(2+) store depletion, possibly by an adaptive downregulation of this pathway. By interfering with Ca(2+)-dependent events at multiple intracellular sites, these effects of Bcl-2 on intracellular Ca(2+) homeostasis may contribute to the protective role of this oncogene against programmed cell death. PMID- 10704438 TI - In situ biochemical demonstration that P-glycoprotein is a drug efflux pump with broad specificity. AB - While P-glycoprotein (Pgp) is the most studied protein involved in resistance to anti-cancer drugs, its mechanism of action is still under debate. Studies of Pgp have used cell lines selected with chemotherapeutics which may have developed many mechanisms of resistance. To eliminate the confounding effects of drug selection on understanding the action of Pgp, we studied cells transiently transfected with a Pgp-green fluorescent protein (GFP) fusion protein. This method generated a mixed population of unselected cells with a wide range of Pgp GFP expression levels and allowed simultaneous measurements of Pgp level and drug accumulation in living cells. The results showed that Pgp-GFP expression was inversely related to the accumulation of chemotherapeutic drugs. The reduction in drug concentration was reversed by agents that block multiple drug resistance (MDR) and by the UIC2 anti-Pgp antibody. Quantitative analysis revealed an inverse linear relationship between the fluorescence of Pgp-GFP and MDR dyes. This suggests that Pgp levels alone limit drug accumulation by active efflux; cooperativity between enzyme, substrate, or inhibitor molecules is not required. Additionally, Pgp-GFP expression did not change cellular pH. Our study demonstrates the value of using GFP fusion proteins for quantitative biochemistry in living cells. PMID- 10704439 TI - MAD3 encodes a novel component of the spindle checkpoint which interacts with Bub3p, Cdc20p, and Mad2p. AB - We show that MAD3 encodes a novel 58-kD nuclear protein which is not essential for viability, but is an integral component of the spindle checkpoint in budding yeast. Sequence analysis reveals two regions of Mad3p that are 46 and 47% identical to sequences in the NH(2)-terminal region of the budding yeast Bub1 protein kinase. Bub1p is known to bind Bub3p (Roberts et al. 1994) and we use two hybrid assays and coimmunoprecipitation experiments to show that Mad3p can also bind to Bub3p. In addition, we find that Mad3p interacts with Mad2p and the cell cycle regulator Cdc20p. We show that the two regions of homology between Mad3p and Bub1p are crucial for these interactions and identify loss of function mutations within each domain of Mad3p. We discuss roles for Mad3p and its interactions with other spindle checkpoint proteins and with Cdc20p, the target of the checkpoint. PMID- 10704440 TI - In vitro formation of the endoplasmic reticulum occurs independently of microtubules by a controlled fusion reaction. AB - We have established an in vitro system for the formation of the endoplasmic reticulum (ER). Starting from small membrane vesicles prepared from Xenopus laevis eggs, an elaborate network of membrane tubules is formed in the presence of cytosol. In the absence of cytosol, the vesicles only fuse to form large spheres. Network formation requires a ubiquitous cytosolic protein and nucleoside triphosphates, is sensitive to N-ethylmaleimide and high cytosolic Ca(2+) concentrations, and proceeds via an intermediate stage in which vesicles appear to be clustered. Microtubules are not required for membrane tubule and network formation. Formation of the ER network shares significant similarities with formation of the nuclear envelope. Our results suggest that the ER network forms in a process in which cytosolic factors modify and regulate a basic reaction of membrane vesicle fusion. PMID- 10704441 TI - Mechanism of residence of cytochrome b(5), a tail-anchored protein, in the endoplasmic reticulum. AB - Endoplasmic reticulum (ER) proteins maintain their residency by static retention, dynamic retrieval, or a combination of the two. Tail-anchored proteins that contain a cytosolic domain associated with the lipid bilayer via a hydrophobic stretch close to the COOH terminus are sorted within the secretory pathway by largely unknown mechanisms. Here, we have investigated the mode of insertion in the bilayer and the intracellular trafficking of cytochrome b(5) (b[5]), taken as a model for ER-resident tail-anchored proteins. We first demonstrated that b(5) can acquire a transmembrane topology posttranslationally, and then used two tagged versions of b(5), N-glyc and O-glyc b(5), containing potential N- and O glycosylation sites, respectively, at the COOH-terminal lumenal extremity, to discriminate between retention and retrieval mechanisms. Whereas the N-linked oligosaccharide provided no evidence for retrieval from a downstream compartment, a more stringent assay based on carbohydrate acquisition by O-glyc b(5) showed that b(5) gains access to enzymes catalyzing the first steps of O-glycosylation. These results suggest that b(5) slowly recycles between the ER and the cis-Golgi complex and that dynamic retrieval as well as retention are involved in sorting of tail-anchored proteins. PMID- 10704442 TI - Regulation of HMG-CoA reductase degradation requires the P-type ATPase Cod1p/Spf1p. AB - The integral ER membrane protein HMG-CoA reductase (HMGR) is a key enzyme of the mevalonate pathway from which sterols and other essential molecules are produced. HMGR degradation occurs in the ER and is regulated by mevalonate-derived signals. Little is known about the mechanisms responsible for regulating HMGR degradation. The yeast Hmg2p isozyme of HMGR undergoes regulated degradation in a manner very similar to mammalian HMGR, allowing us to isolate mutants deficient in regulating Hmg2p stability. We call these mutants cod mutants for the control of HMG-CoA reductase degradation. With this screen, we have identified the first gene of this class, COD1, which encodes a P-type ATPase and is identical to SPF1. Our data suggested that Cod1p is a calcium transporter required for regulating Hmg2p degradation. This role for Cod1p is distinctly different from that of the well characterized Ca(2+) P-type ATPase Pmr1p which is neither required for Hmg2p degradation nor its control. The identification of Cod1p is especially intriguing in light of the role Ca(2+) plays in the regulated degradation of mammalian HMGR. PMID- 10704443 TI - The Emp24 complex recruits a specific cargo molecule into endoplasmic reticulum derived vesicles. AB - Members of the yeast p24 family, including Emp24p and Erv25p, form a heteromeric complex required for the efficient transport of selected proteins from the endoplasmic reticulum (ER) to the Golgi apparatus. The specific functions and sites of action of this complex are unknown. We show that Emp24p is directly required for efficient packaging of a lumenal cargo protein, Gas1p, into ER derived vesicles. Emp24p and Erv25p can be directly cross-linked to Gas1p in ER derived vesicles. Gap1p, which was not affected by emp24 mutation, was not cross linked. These results suggest that the Emp24 complex acts as a cargo receptor in vesicle biogenesis from the ER. PMID- 10704444 TI - PEX19 binds multiple peroxisomal membrane proteins, is predominantly cytoplasmic, and is required for peroxisome membrane synthesis. AB - Peroxisomes are components of virtually all eukaryotic cells. While much is known about peroxisomal matrix protein import, our understanding of how peroxisomal membrane proteins (PMPs) are targeted and inserted into the peroxisome membrane is extremely limited. Here, we show that PEX19 binds a broad spectrum of PMPs, displays saturable PMP binding, and interacts with regions of PMPs required for their targeting to peroxisomes. Furthermore, mislocalization of PEX19 to the nucleus leads to nuclear accumulation of newly synthesized PMPs. At steady state, PEX19 is bimodally distributed between the cytoplasm and peroxisome, with most of the protein in the cytoplasm. We propose that PEX19 may bind newly synthesized PMPs and facilitate their insertion into the peroxisome membrane. This hypothesis is supported by the observation that the loss of PEX19 results in degradation of PMPs and/or mislocalization of PMPs to the mitochondrion. PMID- 10704445 TI - Dynein-mediated cargo transport in vivo. A switch controls travel distance. AB - Cytoplasmic dynein is a microtubule-based motor with diverse cellular roles. Here, we use mutations in the dynein heavy chain gene to impair the motor's function, and employ biophysical measurements to demonstrate that cytoplasmic dynein is responsible for the minus end motion of bidirectionally moving lipid droplets in early Drosophila embryos. This analysis yields an estimate for the force that a single cytoplasmic dynein exerts in vivo (1.1 pN). It also allows us to quantitate dynein-mediated cargo motion in vivo, providing a framework for investigating how dynein's activity is controlled. We identify three distinct travel states whose general features also characterize plus end motion. These states are preserved in different developmental stages. We had previously provided evidence that for each travel direction, single droplets are moved by multiple motors of the same type (Welte et al. 1998). Droplet travel distances (runs) are much shorter than expected for multiple motors based on in vitro estimates of cytoplasmic dynein processivity. Therefore, we propose the existence of a process that ends runs before the motors fall off the microtubules. We find that this process acts with a constant probability per unit distance, and is typically coupled to a switch in travel direction. A process with similar properties governs plus end motion, and its regulation controls the net direction of transport. PMID- 10704446 TI - Phosphorylation of tyrosine residues 31 and 118 on paxillin regulates cell migration through an association with CRK in NBT-II cells. AB - Identification of signaling molecules that regulate cell migration is important for understanding fundamental processes in development and the origin of various pathological conditions. The migration of Nara Bladder Tumor II (NBT-II) cells was used to determine which signaling molecules are specifically involved in the collagen-mediated locomotion. We show here that paxillin is tyrosine phosphorylated after induction of motility on collagen. Overexpression of paxillin mutants in which tyrosine 31 and/or tyrosine 118 were replaced by phenylalanine effectively impaired cell motility. Moreover, stimulation of motility by collagen preferentially enhanced the association of paxillin with the SH2 domain of the adaptor protein CrkII. Mutations in both tyrosine 31 and 118 diminished the phosphotyrosine content of paxillin and prevented the formation of the paxillin-Crk complex, suggesting that this association is necessary for collagen-mediated NBT-II cell migration. Other responses to collagen, such as cell adhesion and spreading, were not affected by these mutations. Overexpression of wild-type paxillin or Crk could bypass the migration-deficient phenotype. Both the SH2 and the SH3 domains of CrkII are shown to play a critical role in this collagen-mediated migration. These results demonstrate the important role of the paxillin-Crk complex in the collagen-induced cell motility. PMID- 10704447 TI - Long-term culture of purified postnatal oligodendrocyte precursor cells. Evidence for an intrinsic maturation program that plays out over months. AB - Oligodendrocytes myelinate axons in the vertebrate central nervous system (CNS). They develop from precursor cells (OPCs), some of which persist in the adult CNS. Adult OPCs differ in many of their properties from OPCs in the developing CNS. In this study we have purified OPCs from postnatal rat optic nerve and cultured them in serum-free medium containing platelet-derived growth factor (PDGF), the main mitogen for OPCs, but in the absence of thyroid hormone in order to inhibit their differentiation into oligodendrocytes. We find that many of the cells continue to proliferate for more than a year and progressively acquire a number of the characteristics of OPCs isolated from adult optic nerve. These findings suggest that OPCs have an intrinsic maturation program that progressively changes the cell's phenotype over many months. When we culture the postnatal OPCs in the same conditions but with the addition of basic fibroblast growth factor (bFGF), the cells acquire these mature characteristics much more slowly, suggesting that the combination of bFGF and PDGF, previously shown to inhibit OPC differentiation, also inhibits OPC maturation. The challenge now is to determine the molecular basis of such a protracted maturation program and how the program is restrained by bFGF. PMID- 10704448 TI - Massive idiosyncratic exon skipping corrects the nonsense mutation in dystrophic mouse muscle and produces functional revertant fibers by clonal expansion. AB - Conventionally, nonsense mutations within a gene preclude synthesis of a full length functional protein. Obviation of such a blockage is seen in the mdx mouse, where despite a nonsense mutation in exon 23 of the dystrophin gene, occasional so-called revertant muscle fibers are seen to contain near-normal levels of its protein product. Here, we show that reversion of dystrophin expression in mdx mice muscle involves unprecedented massive loss of up to 30 exons. We detected several alternatively processed transcripts that could account for some of the revertant dystrophins and could not detect genomic deletion from the region commonly skipped in revertant dystrophin. This, together with exon skipping in two noncontiguous regions, favors aberrant splicing as the mechanism for the restoration of dystrophin, but is hard to reconcile with the clonal idiosyncrasy of revertant dystrophins. Revertant dystrophins retain functional domains and mediate plasmalemmal assembly of the dystrophin-associated glycoprotein complex. Physiological function of revertant fibers is demonstrated by the clonal growth of revertant clusters with age, suggesting that revertant dystrophin could be used as a guide to the construction of dystrophin expression vectors for individual gene therapy. The dystrophin gene in the mdx mouse provides a favored system for study of exon skipping associated with nonsense mutations. PMID- 10704449 TI - Sphingolipid-cholesterol rafts diffuse as small entities in the plasma membrane of mammalian cells. AB - To probe the dynamics and size of lipid rafts in the membrane of living cells, the local diffusion of single membrane proteins was measured. A laser trap was used to confine the motion of a bead bound to a raft protein to a small area (diam < or = 100 nm) and to measure its local diffusion by high resolution single particle tracking. Using protein constructs with identical ectodomains and different membrane regions and vice versa, we demonstrate that this method provides the viscous damping of the membrane domain in the lipid bilayer. When glycosylphosphatidylinositol (GPI) -anchored and transmembrane proteins are raft associated, their diffusion becomes independent of the type of membrane anchor and is significantly reduced compared with that of nonraft transmembrane proteins. Cholesterol depletion accelerates the diffusion of raft-associated proteins for transmembrane raft proteins to the level of transmembrane nonraft proteins and for GPI-anchored proteins even further. Raft-associated GPI-anchored proteins were never observed to dissociate from the raft within the measurement intervals of up to 10 min. The measurements agree with lipid rafts being cholesterol-stabilized complexes of 26 +/- 13 nm in size diffusing as one entity for minutes. PMID- 10704450 TI - Schwann cell myelination requires timely and precise targeting of P(0) protein. AB - This report investigated mechanisms responsible for failed Schwann cell myelination in mice that overexpress P(0) (P(0)(tg)), the major structural protein of PNS myelin. Quantitative ultrastructural immunocytochemistry established that P(0) protein was mistargeted to abaxonal, periaxonal, and mesaxon membranes in P(0)(tg) Schwann cells with arrested myelination. The extracellular leaflets of P(0)-containing mesaxon membranes were closely apposed with periodicities of compact myelin. The myelin-associated glycoprotein was appropriately sorted in the Golgi apparatus and targeted to periaxonal membranes. In adult mice, occasional Schwann cells myelinated axons possibly with the aid of endocytic removal of mistargeted P(0). These results indicate that P(0) gene multiplication causes P(0) mistargeting to mesaxon membranes, and through obligate P(0) homophilic adhesion, renders these dynamic membranes inert and halts myelination. PMID- 10704451 TI - P(0) glycoprotein overexpression causes congenital hypomyelination of peripheral nerves. AB - We show that normal peripheral nerve myelination depends on strict dosage of the most abundantly expressed myelin gene, myelin protein zero (Mpz). Transgenic mice containing extra copies of Mpz manifested a dose-dependent, dysmyelinating neuropathy, ranging from transient perinatal hypomyelination to arrested myelination and impaired sorting of axons by Schwann cells. Myelination was restored by breeding the transgene into the Mpz-null background, demonstrating that dysmyelination does not result from a structural alteration or Schwann cell extrinsic effect of the transgenic P(0) glycoprotein. Mpz mRNA overexpression ranged from 30-700%, whereas an increased level of P(0) protein was detected only in nerves of low copy-number animals. Breeding experiments placed the threshold for dysmyelination between 30 and 80% Mpz overexpression. These data reveal new points in nerve development at which Schwann cells are susceptible to increased gene dosage, and suggest a novel basis for hereditary neuropathy. PMID- 10704452 TI - A dual role of erbB2 in myelination and in expansion of the schwann cell precursor pool. AB - Neuregulin-1 provides an important axonally derived signal for the survival and growth of developing Schwann cells, which is transmitted by the ErbB2/ErbB3 receptor tyrosine kinases. Null mutations of the neuregulin-1, erbB2, or erbB3 mouse genes cause severe deficits in early Schwann cell development. Here, we employ Cre-loxP technology to introduce erbB2 mutations late in Schwann cell development, using a Krox20-cre allele. Cre-mediated erbB2 ablation occurs perinatally in peripheral nerves, but already at E11 within spinal roots. The mutant mice exhibit a widespread peripheral neuropathy characterized by abnormally thin myelin sheaths, containing fewer myelin wraps. In addition, in spinal roots the Schwann cell precursor pool is not correctly established. Thus, the Neuregulin signaling system functions during multiple stages of Schwann cell development and is essential for correct myelination. The thickness of the myelin sheath is determined by the axon diameter, and we suggest that trophic signals provided by the nerve determine the number of times a Schwann cell wraps an axon. PMID- 10704453 TI - Syndapin isoforms participate in receptor-mediated endocytosis and actin organization. AB - Syndapin I (SdpI) interacts with proteins involved in endocytosis and actin dynamics and was therefore proposed to be a molecular link between the machineries for synaptic vesicle recycling and cytoskeletal organization. We here report the identification and characterization of SdpII, a ubiquitously expressed isoform of the brain-specific SdpI. Certain splice variants of rat SdpII in other species were named FAP52 and PACSIN 2. SdpII binds dynamin I, synaptojanin, synapsin I, and the neural Wiskott-Aldrich syndrome protein (N-WASP), a stimulator of Arp2/3 induced actin filament nucleation. In neuroendocrine cells, SdpII colocalizes with dynamin, consistent with a role for syndapin in dynamin mediated endocytic processes. The src homology 3 (SH3) domain of SdpI and -II inhibited receptor-mediated internalization of transferrin, demonstrating syndapin involvement in endocytosis in vivo. Overexpression of full-length syndapins, but not the NH(2)-terminal part or the SH3 domains alone, had a strong effect on cortical actin organization and induced filopodia. This syndapin overexpression phenotype appears to be mediated by the Arp2/3 complex at the cell periphery because it was completely suppressed by coexpression of a cytosolic COOH-terminal fragment of N-WASP. Consistent with a role in actin dynamics, syndapins localized to sites of high actin turnover, such as filopodia tips and lamellipodia. Our results strongly suggest that syndapins link endocytosis and actin dynamics. PMID- 10704455 TI - Integrin dynamics and matrix assembly: tensin-dependent translocation of alpha(5)beta(1) integrins promotes early fibronectin fibrillogenesis. AB - Fibronectin matrix assembly is a multistep, integrin-dependent process. To investigate the role of integrin dynamics in fibronectin fibrillogenesis, we developed an antibody-chasing technique for simultaneous tracking of two integrin populations by different antibodies. We established that whereas the vitronectin receptor alpha(v)beta(3) remains within focal contacts, the fibronectin receptor alpha(5)beta(1) translocates from focal contacts into and along extracellular matrix (ECM) contacts. This escalator-like translocation occurs relative to the focal contacts at 6.5 +/- 0.7 microm/h and is independent of cell migration. It is induced by ligation of alpha(5)beta(1) integrins and depends on interactions with a functional actin cytoskeleton and vitronectin receptor ligation. During cell spreading, translocation of ligand-occupied alpha(5)beta(1) integrins away from focal contacts and along bundles of actin filaments generates ECM contacts. Tensin is a primary cytoskeletal component of these ECM contacts, and a novel dominant-negative inhibitor of tensin blocked ECM contact formation, integrin translocation, and fibronectin fibrillogenesis without affecting focal contacts. We propose that translocating alpha(5)beta(1) integrins induce initial fibronectin fibrillogenesis by transmitting cytoskeleton-generated tension to extracellular fibronectin molecules. Blocking this integrin translocation by a variety of treatments prevents the formation of ECM contacts and fibronectin fibrillogenesis. These studies identify a localized, directional, integrin translocation mechanism for matrix assembly. PMID- 10704454 TI - Physiological role of gap-junctional hemichannels. Extracellular calcium dependent isosmotic volume regulation. AB - Hemichannels in the overlapping regions of apposing cells plasma membranes join to form gap junctions and provide an intercellular communication pathway. Hemichannels are also present in the nonjunctional regions of individual cells and their activity is gated by several agents, including calcium. However, their physiological roles are unknown. Using techniques of atomic force microscopy (AFM), fluorescent dye uptake assay, and laser confocal immunofluorescence imaging, we have examined the extracellular calcium-dependent modulation of cell volume. In response to a change in the extracellular physiological calcium concentration (1.8 to 100 Hz. Here, we show that such networks, to which interneurons have been added along with chemical synaptic interactions between respective cell types, can generate population ripples superimposed on afferent input-induced intracellular depolarizations. During simulated ripples, interneurons fire at high rates, whereas pyramidal cells fire at lower rates. The model oscillation is generated by the electrically coupled pyramidal cell axons, which then phasically excite interneurons at ripple frequency. The oscillation occurs transiently because rippling can express itself only when axons and cells are sufficiently depolarized. Our model predicts the occurrence of spikelets (fast prepotentials) in some pyramidal cells during sharp waves. PMID- 10704479 TI - Molecular basis of symbiotic promiscuity. AB - Eukaryotes often form symbioses with microorganisms. Among these, associations between plants and nitrogen-fixing bacteria are responsible for the nitrogen input into various ecological niches. Plants of many different families have evolved the capacity to develop root or stem nodules with diverse genera of soil bacteria. Of these, symbioses between legumes and rhizobia (Azorhizobium, Bradyrhizobium, Mesorhizobium, and Rhizobium) are the most important from an agricultural perspective. Nitrogen-fixing nodules arise when symbiotic rhizobia penetrate their hosts in a strictly controlled and coordinated manner. Molecular codes are exchanged between the symbionts in the rhizosphere to select compatible rhizobia from pathogens. Entry into the plant is restricted to bacteria that have the "keys" to a succession of legume "doors". Some symbionts intimately associate with many different partners (and are thus promiscuous), while others are more selective and have a narrow host range. For historical reasons, narrow host range has been more intensively investigated than promiscuity. In our view, this has given a false impression of specificity in legume-Rhizobium associations. Rather, we suggest that restricted host ranges are limited to specific niches and represent specialization of widespread and more ancestral promiscuous symbioses. Here we analyze the molecular mechanisms governing symbiotic promiscuity in rhizobia and show that it is controlled by a number of molecular keys. PMID- 10704483 TI - Exacerbated responses to oxidative stress by an Na(+) load in isolated nerve terminals: the role of ATP depletion and rise of [Ca(2+)](i). AB - We have explored the consequences of a [Na(+)](i) load and oxidative stress in isolated nerve terminals. The Na(+) load was achieved by veratridine (5-40 microM), which allows Na(+) entry via a voltage-operated Na(+) channel, and oxidative stress was induced by hydrogen peroxide (0.1-0.5 mM). Remarkably, neither the [Na(+)](i) load nor exposure to H(2)O(2) had any major effect on [Ca(2+)](i), mitochondrial membrane potential (Deltapsim), or ATP level. However, the combination of an Na(+) load and oxidative stress caused ATP depletion, a collapse of Deltapsim, and a progressive deregulation of [Ca(2+)](i) and [Na(+)](i) homeostasis. The decrease in the ATP level was unrelated to an increase in [Ca(2+)](i) and paralleled the rise in [Na(+)](i). The loss of Deltapsim was prevented in the absence of Ca(2+) but unaltered in the presence of cyclosporin A. We conclude that the increased ATP consumption by the Na,K-ATPase that results from a modest [Na(+)](i) load places an additional demand on mitochondria metabolically compromised by an oxidative stress, which are unable to produce a sufficient amount of ATP to fuel the ATP-driven ion pumps. This results in a deregulation of [Na(+)](i) and [Ca(2+)](i), and as a result of the latter, collapse of Deltapsim. The vicious cycle generated in the combined presence of Na(+) load and oxidative stress could be an important factor in the neuronal injury produced by ischemia or excitotoxicity, in which the oxidative insult is superimposed on a disturbed Na(+) homeostasis. PMID- 10704484 TI - cAMP-dependent plasticity at excitatory cholinergic synapses in Drosophila neurons: alterations in the memory mutant dunce. AB - It is well known that cAMP signaling plays a role in regulating functional plasticity at central glutamatergic synapses. However, in the Drosophila CNS, where acetylcholine is thought to be a primary excitatory neurotransmitter, cellular changes in neuronal communication mediated by cAMP remain unexplored. In this study we examined the effects of elevated cAMP levels on fast excitatory cholinergic synaptic transmission in cultured embryonic Drosophila neurons. We report that chronic elevation in neuronal cAMP (in dunce neurons or wild-type neurons grown in db-cAMP) results in an increase in the frequency of cholinergic miniature EPSCs (mEPSCs). The absence of alterations in mEPSC amplitude or kinetics suggests that the locus of action is presynaptic. Furthermore, a brief exposure to db-cAMP induces two distinct changes in transmission at established cholinergic synapses in wild-type neurons: a short-term increase in the frequency of spontaneous action potential-dependent synaptic currents and a long-lasting, protein synthesis-dependent increase in the mEPSC frequency. A more persistent increase in cholinergic mEPSC frequency induced by repetitive, spaced db-cAMP exposure in wild-type neurons is absent in neurons from the memory mutant dunce. These data demonstrate that interneuronal excitatory cholinergic synapses in Drosophila, like central excitatory glutamatergic synapses in other species, are sites of cAMP-dependent plasticity. In addition, the alterations in dunce neurons suggest that cAMP-dependent plasticity at cholinergic synapses could mediate changes in neuronal communication that contribute to memory formation. PMID- 10704485 TI - Axonal transport of microtubule-associated protein 1B (MAP1B) in the sciatic nerve of adult rat: distinct transport rates of different isoforms. AB - Cytoskeletal proteins are axonally transported with slow components a and b (SCa and SCb). In peripheral nerves, the transport velocity of SCa, which includes neurofilaments and tubulin, is 1-2 mm/d, whereas SCb, which includes actin, tubulin, and numerous soluble proteins, moves as a heterogeneous wave at 2-4 mm/d. We have shown that two isoforms of microtubule-associated protein 1B (MAP1B), which can be separated on SDS polyacrylamide gels on the basis of differences in their phosphorylation states (band I and band II), were transported at two different rates. All of band I MAP1B moved as a coherent wave at a velocity of 7-9 mm/d, distinct from slow axonal transport components SCa and SCb. Several other proteins were detected within the component that moved at the velocity of 7-9 mm/d, including the leading wave of tubulin and actin. The properties of this component define a distinct fraction of the slow axonal transport that we suggest to term slow component c (SCc). The relatively fast transport of the phosphorylated MAP1B isoform at 7-9 mm/d may account for the high concentration of phosphorylated MAP1B in the distal end of growing axons. In contrast to band I MAP1B, the transport profile of band II was complex and contained components moving with SCa and SCb and a leading edge at SCc. Thus, MAP1B isoforms in different phosphorylation states move with distinct components of slow axonal transport, possibly because of differences in their abilities to associate with other proteins. PMID- 10704486 TI - Presynaptic P2X receptors facilitate inhibitory GABAergic transmission between cultured rat spinal cord dorsal horn neurons. AB - The superficial layers of the spinal cord dorsal horn (DH) express P2X2, P2X4, and P2X6 subunits entering into the formation of ionotropic (P2X) receptors for ATP. Using a culture system of laminae I-III from neonatal rat DH, we show that ATP induced a fast nonselective cation current in 38% of the neurons (postsynaptic effect). ATP also increased the frequency of miniature IPSCs (mIPSCs) mediated by GABA(A) receptors or by glycine receptors in 22 and 9%, respectively, of the neurons tested (presynaptic effect) but had no effect on glutamatergic transmission. The presynaptic effect of ATP on GABAergic transmission was not significantly affected by thapsigargin (1 microM) but was completely dependent on Ca(2+) influx. Presynaptic and postsynaptic effects were inhibited by suramin, pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid, and reactive blue and were not reproduced by uridine 5'-triphosphate (UTP) or adenosine 5'-O-(2-thiodiphosphate) (ADP-beta-S), suggesting the implication of ionotropic P2X rather than of metabotropic P2Y receptors. alphabeta-methylene-ATP (100 microM) did not reproduce the effects of ATP. ATP reversibly increased the amplitude of electrically evoked GABAergic IPSCs and reduced paired-pulse inhibition or facilitation without affecting IPSC kinetics. This effect was preferentially, but not exclusively, observed in neurons coreleasing ATP and GABA. We conclude that in cultured DH neurons, the effects of ATP are mediated by P2X receptors having a pharmacological profile dominated by the P2X2 subunit. The presynaptic receptors might underlie a modulatory action of ATP on a subset of GABAergic interneurons involved in the spinal processing of nociceptive information. PMID- 10704487 TI - The neuronal monoamine transporter VMAT2 is regulated by the trimeric GTPase Go(2). AB - Monoamines such as noradrenaline and serotonin are stored in secretory vesicles and released by exocytosis. Two related monoamine transporters, VMAT1 and VMAT2, mediate vesicular transmitter uptake. Previously we have reported that in the rat pheochromocytoma cell line PC 12 VMAT1, localized to peptide-containing secretory granules, is controlled by the heterotrimeric G-protein Go(2). We now show that in BON cells, a human serotonergic neuroendocrine cell line derived from a pancreatic tumor expressing both transporters on large, dense-core vesicles, VMAT2 is even more sensitive to G-protein regulation than VMAT1. The activity of both transporters is only downregulated by Galphao(2), whereas comparable concentrations of Galphao(1) are without effect. In serotonergic raphe neurons in primary culture VMAT2 is also downregulated by pertussis toxin-sensitive Go(2). By electron microscopic analysis from prefrontal cortex we show that VMAT2 and Galphao(2) associate preferentially to locally recycling small synaptic vesicles in serotonergic terminals. In addition, Go(2)-dependent modulation of VMAT2 also works when using a crude synaptic vesicle preparation from this brain area. We conclude that regulation of monoamine uptake by the heterotrimeric G proteins is a general feature of monoaminergic neurons that controls the content of both large, dense-core and small synaptic vesicles. PMID- 10704488 TI - Changes in activating protein 1 (AP-1) composition correspond with the biphasic profile of nerve growth factor mRNA expression in rat hippocampus after hilus lesion-induced seizures. AB - In adult brain, nerve growth factor (NGF) gene expression is generally upregulated by neuronal activity. However, a single episode of hilus lesion (HL) induced limbic seizures stimulates a biphasic increase in NGF mRNA expression with peaks at 4-6 and 24 hr after lesion and an intervening return to control levels at 10-12 hr after lesion. In vitro studies suggest that NGF transcription is regulated via an activating protein 1 (AP-1) binding site in the first intron of the NGF gene. To examine the relationship between seizure-induced AP-1 binding and NGF gene expression in this paradigm, NGF mRNA levels and AP-1 binding were examined after HL seizures. Furthermore, to gain insight into the functional composition of the AP-1 complex, supershift analysis was performed to characterize which Fos and Jun family members are included in the AP-1-binding complex at the different time points analyzed. Solution hybridization analysis verified the biphasic increase in NGF mRNA content of the dentate gyrus after HL seizures. After an initial increase, AP-1 binding slowly declined in a stepwise manner that encompassed, but did not correspond with, the two phases of NGF mRNA expression. However, supershift analyses demonstrated that the relative contributions of JunD and JunB to the AP-1 complex exhibited positive and negative correlations, respectively, with the phases of increased NGF expression after HL. These results suggest that AP-1 complexes containing JunD promote NGF transactivation and that transient changes in the relative contributions of JunD and JunB to AP-1 binding underlie the biphasic increase in NGF gene expression induced by HL seizures. PMID- 10704489 TI - Expression of Bcl-2 protects against photoreceptor degeneration in retinal degeneration slow (rds) mice. AB - The retinal degeneration slow or rds gene encodes rds/peripherin, an integral membrane glycoprotein in the outer segments of rod and cone photoreceptors. Mice homozygous for a null mutation in rds fail to develop outer segments and undergo subsequent degeneration of photoreceptors by the apoptotic pathway. Mutations in the human RDS gene are responsible for several forms of inherited blindness including autosomal-dominant retinitis pigmentosa and macular degeneration. Here, we examined the effects of ectopic Bcl-2 expression in transgenic photoreceptors on the rate of retinal degeneration in rds mutant mice. We observed an approximately twofold preservation of photoreceptors compared with nontransgenic rds mutant mice at 3 months. Immunoblot analysis showed similar levels of Bcl-2 in 2-, 3-, and 4-week-old transgenic mice. Expression of Bcl-2 in the rds mouse did not lead to outer segment formation and did not induce cell death. These results suggest that Bcl-2 expression may be an effective therapeutic strategy in humans with mutations in RDS or other genes that affect the integrity of photoreceptor outer segments. PMID- 10704490 TI - Effects of neurotrophins on cortical plasticity: same or different? AB - Neurotrophins are important regulators of visual cortical plasticity. It is still unclear, however, whether they play similar or different roles and which are their effects on the electrical activity of cortical neurons in vivo. We therefore compared the effects of all neurotrophins, nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin-4 (NT-4), and neurotrophin 3 (NT-3) on visual cortical plasticity and on cell spontaneous and visually evoked activity. Rats were monocularly deprived for 1 week at the peak of the critical period, and neurotrophins were infused intracortically. The main finding is that, with the exception of NT-3, all neurotrophins affect the outcome of monocular deprivation, but there are clear differences in their mechanisms of action. In particular, NT-4 and NGF counteract monocular deprivation effects without causing detectable alterations either in spontaneous or visually evoked neuronal activity. BDNF is less effective on ocular dominance plasticity and, in addition, strongly affects spontaneous and visually evoked activity in cortical neurons. PMID- 10704491 TI - Regulation of kinetic properties of GluR2 AMPA receptor channels by alternative splicing. AB - The four subunits of the AMPA-type glutamate receptor (GluR1-GluR4 or GluR-A-GluR D) exist in two distinct forms, flip and flop, generated by alternative splicing of a 115 bp region. The GluR2 subunit plays a key role in determining the functional properties of the AMPA receptor channel. In this study, we examined the differences in kinetic properties between the flip and flop splice variants of the GluR2 subunit expressed in Xenopus oocytes using fast agonist application techniques. Glutamate was applied to outside-out patches from oocytes with piezo driven double-barreled application pipettes. Because homomeric receptor channels composed of the edited form of GluR2 (GluR2R) produce no appreciable current responses, we expressed the unedited form of GluR2 (GluR2Q) in oocytes, which produced large current responses sufficient for analysis of the kinetic properties. The time constant for desensitization during application of 1 mM glutamate was 5.89 +/- 0. 17 msec (n = 50) in flip and 1.18 +/- 0.05 msec (n = 37) in flop. The deactivation time constant was 0.62 +/- 0.06 msec (n = 10) in flip and 0.54 +/- 0.05 msec (n = 10) in flop. The steady-state nondesensitizing current was 6.8 +/- 0.4% (n = 53) of the peak current in flip, whereas it was almost negligible in flop, being only 1.1 +/- 0.1% (n = 36). The slower desensitization kinetics and larger steady-state current responses in the flip variant were also observed in heteromeric receptors assembled from GluR2Q/GluR2R. Thus, desensitization occurred much more prominently in the flop variant in the recombinant GluR2 receptor channels. PMID- 10704492 TI - Functional GluR6 kainate receptors in the striatum: indirect downregulation of synaptic transmission. AB - Kainate receptors (KARs) are abundantly expressed in the basal ganglia, but their function in synaptic transmission has not been established. In the present study, we show that the GluR6 subunit of KARs is expressed in both substance P- and enkephalin-containing GABAergic projection neurons of the mouse striatum. Using whole-cell voltage-clamp recordings in brain slices, we demonstrate the presence of functional KARs in the dorsal striatum activated by low concentrations of the AMPA/KAR agonist domoate in wild-type but not GluR6-deficient mice. Despite the abundance of KARs, we found no evidence for synaptic activation of these receptors after single or repetitive stimulation of glutamatergic afferents. Domoate induces a transient increase in the frequency of spontaneous IPSCs of small amplitude and a sustained depression of large IPSCs evoked by minimal electrical stimulation within the striatum in wild-type mice but not in GluR6 deficient mice. This depressant effect is inhibited in presence of adenosine A(2A) receptor antagonists, ZM-241385 and SCH-58261. These data strongly suggest that, in striatal neurons, KARs depress GABAergic synaptic transmission indirectly via release of adenosine acting on A(2A) receptors. PMID- 10704493 TI - Multiple G-protein betagamma combinations produce voltage-dependent inhibition of N-type calcium channels in rat superior cervical ganglion neurons. AB - Activation of several G-protein-coupled receptors leads to voltage-dependent (VD) inhibition of N- and P/Q-type Ca(2+) channels via G-protein betagamma subunits (Gbetagamma). The purpose of the present study was to determine the ability of different Gbetagamma combinations to produce VD inhibition of N-type Ca(2+) channels in rat superior cervical ganglion neurons. Various Gbetagamma combinations were heterologously overexpressed by intranuclear microinjection of cDNA and tonic VD Ca(2+) channel inhibition evaluated using the whole-cell voltage-clamp technique. Overexpression of Gbeta1-Gbeta5, in combination with several different Ggamma subunits, resulted in tonic VD Ca(2+) channel inhibition. Robust Ca(2+) channel modulation required coexpression of both Gbeta and Ggamma. Expression of either subunit alone produced minimal effects. To substantiate the apparent lack of Gbetagamma specificity, we examined whether heterologously expressed Gbetagamma displaced native Gbetagamma from heterotrimeric complexes. To this end, mutant Gbeta subunits were constructed that differentially modulated N-type Ca(2+) and G-protein-gated inward rectifier K(+) channels. Results from these studies indicated that significant displacement does not occur, and thus the observed Gbetagamma modulation can be attributed directly to the heterologously expressed Gbetagamma combinations. PMID- 10704494 TI - Organization of ionotropic glutamate receptors at dendrodendritic synapses in the rat olfactory bulb. AB - Dendrodendritic synapses between mitral (or tufted) and granule cells of the olfactory bulb play a major role in the processes of odor discrimination and olfactory learning. Release of glutamate at these synapses activates postsynaptic receptors on the dendritic spines of granule cells, as well as presynaptic NMDA receptors in the mitral cell membrane. However, immunocytochemical studies have failed to demonstrate the presence of ionotropic glutamate receptors in granule cell dendrites. By using a postembedding immunogold procedure, we describe here the precise organization of neurotransmitter receptors at dendrodendritic synapses. We show that there is a selective localization of glutamate and GABA receptors at asymmetric and symmetric synaptic junctions, respectively. In addition, we demonstrate that NMDA and AMPA receptors are clustered at postsynaptic specializations on granule cell spines and that they are extensively colocalized. Conversely, glutamate receptors do not appear to be concentrated in clusters on mitral cell dendrites, suggesting that the presynaptic effects of glutamate are mediated by a small complement of extrasynaptic receptors. By analyzing the subsynaptic distribution of the NR1 and GluR2/3 subunits, we show that they are distributed along the entire extent of the postsynaptic specialization, indicating that both NMDA and AMPA receptors are available for dendrodendritic signaling between mitral and granule cells. These results indicate that the principles recently found to underlie the organization of glutamate receptors at axospinous synapses also apply to dendrodendritic synapses. PMID- 10704495 TI - Functional correlation of GABA(A) receptor alpha subunits expression with the properties of IPSCs in the developing thalamus. AB - GABA(A) receptor alpha1 and alpha2 subunits are expressed differentially with ontogenic period in the brain, but their functional roles are not known. We have recorded GABA(A) receptor-mediated IPSCs from laterodorsal (LD) thalamic relay neurons in slices of rat brain at various postnatal ages and found that decay times of evoked IPSCs and spontaneous miniature IPSCs undergo progressive shortening during the first postnatal month. With a similar time course, expression of transcripts and proteins of GABA(A) receptor alpha2 subunit in LD thalamic region declined, being replaced by those of alpha1 subunit. To further address the causal relationship between alpha subunits and IPSC decay time kinetics, we have overexpressed GABA(A) receptor alpha1 subunit together with green fluorescent protein in LD thalamic neurons in organotypic culture using recombinant Sindbis virus vectors. Miniature IPSCs recorded from the LD thalamic neurons overexpressed with alpha1 subunit had significantly faster decay time compared with control expressed with beta-galactosidase. We conclude that the alpha2-to-alpha1 subunit switch underlies the developmental speeding in the decay time of GABAergic IPSCs. PMID- 10704496 TI - Mice lacking G-protein receptor kinase 1 have profoundly slowed recovery of cone driven retinal responses. AB - G-Protein receptor kinase 1 (GRK1) ("rhodopsin kinase") is necessary for the inactivation of photoactivated rhodopsin, the light receptor of the G-protein transduction cascade of rod photoreceptors. GRK1 has also been reported to be present in retinal cones in which its function is unknown. To examine the role of GRK1 in retinal cone signaling pathways, we measured in mice having null mutations of GRK1 (GRK1 -/-) cone-driven electroretinographic (ERG) responses, including an a-wave component identified as the field potential generated by suppression of the circulating current of the cone photoreceptors. Dark-adapted GRK1 -/- animals generated cone-driven ERGs having saturating amplitudes and sensitivities in both visible and UV spectral regions similar to those of wild type (WT) mice. However, after exposure to a bright conditioning flash, the cone driven ERGs of GRK1 -/- animals recovered 30-50 times more slowly than those of WT mice and similarly slower than the cone-driven ERGs of mice homozygously null for arrestin (Arrestin -/-), whose cone (but not rod) response recoveries were found to be as rapid as those of WT. Our observations argue that GRK1 is essential for normal deactivation of murine cone phototransduction and provide the first functional evidence for a major role of a specific GRK in the inactivation of vertebrate cone phototransduction. PMID- 10704497 TI - Proliferation and differentiation of progenitor cells throughout the intact adult rat spinal cord. AB - The existence of multipotent progenitor populations in the adult forebrain has been widely studied. To extend this knowledge to the adult spinal cord we have examined the proliferation, distribution, and phenotypic fate of dividing cells in the adult rat spinal cord. Bromodeoxyuridine (BrdU) was used to label dividing cells in 13- to 14-week-old, intact Fischer rats. Single daily injections of BrdU were administered over a 12 d period. Animals were killed either 1 d or 4 weeks after the last injection of BrdU. We observed frequent cell division throughout the adult rodent spinal cord, particularly in white matter tracts (5-7% of all nuclei). The majority of BrdU-labeled cells colocalized with markers of immature glial cells. At 4 weeks, 10% of dividing cells expressed mature astrocyte and oligodendroglial markers. These data predict that 0.75% of all astrocytes and 0.82% of all oligodendrocytes are derived from a dividing population over a 4 week period. To determine the migratory nature of dividing cells, a single BrdU injection was given to animals that were killed 1 hr after the injection. In these tissues, the distribution and incidence of BrdU labeling matched those of the 4 week post injection (pi) groups, suggesting that proliferating cells divide in situ rather than migrate from the ependymal zone. These data suggest a higher level of cellular plasticity for the intact spinal cord than has previously been observed and that glial progenitors exist in the outer circumference of the spinal cord that can give rise to both astrocytes and oligodendrocytes. PMID- 10704498 TI - Distinct roles for ionotropic and metabotropic glutamate receptors in the maturation of excitatory synapses. AB - We used the single-cell culture preparation to study the role of activity in the development of glutamatergic synapses in vitro. Rat hippocampal cells grown in isolation on glial islands formed functional autaptic connections and continued to elaborate new synapses throughout the 2 week investigation, resulting in increases in both the evoked AMPA receptor (AMPAR) and NMDA receptor (NMDAR) components of the EPSC. Synaptogenesis was not prevented by chronic blockade of sodium channels or all of the known glutamate receptors. Analysis of miniature EPSCs revealed that AMPAR quantal size doubled over time in vitro whereas NMDAR quantal size remained constant. However, the proportion of synaptic responses mediated only by NMDARs increased over time in vitro. The increase in AMPAR quantal size was prevented by TTX and ionotropic glutamate receptor antagonists, whereas the increase in the proportion of NMDAR-only synapses was prevented by metabotropic glutamate receptor antagonists. Notably, chronic NMDAR blockade incubation did not block the formation of the AMPAR EPSC, indicating that NMDAR dependent plasticity is not necessary for the onset of AMPAR synaptic transmission in this system. We conclude that action potentials and ionotropic glutamate receptor activation are necessary for the developmental increase in AMPAR quantal size and that metabotropic glutamate receptor activation is required for the production of NMDAR-only synapses, but none of these is essential for synapse formation. PMID- 10704499 TI - Neural cell adhesion molecule-stimulated neurite outgrowth depends on activation of protein kinase C and the Ras-mitogen-activated protein kinase pathway. AB - The signal transduction pathways associated with neural cell adhesion molecule (NCAM)-induced neuritogenesis are only partially characterized. We here demonstrate that NCAM-induced neurite outgrowth depends on activation of p59(fyn), focal adhesion kinase (FAK), phospholipase Cgamma (PLCgamma), protein kinase C (PKC), and the Ras-mitogen-activated protein (MAP) kinase pathway. This was done using a coculture system consisting of PC12-E2 cells grown on fibroblasts, with or without NCAM expression, allowing NCAM-NCAM interactions resulting in neurite outgrowth. PC12-E2 cells were transiently transfected with expression plasmids encoding constitutively active forms of Ras, Raf, MAP kinase kinases MEK1 and 2, dominant negative forms of Ras and Raf, and the FAK-related nonkinase. Alternatively, PC12-E2 cells were submitted to treatment with antibodies to the fibroblast growth factor (FGF) receptor, inhibitors of the nonreceptor tyrosine kinase p59(fyn), PLC, PKC and MEK and an activator of PKC, phorbol-12-myristate-13-acetate (PMA). MEK2 transfection rescued cells treated with all inhibitors. The same was found for PMA treatment, except when cells concomitantly were treated with the MEK inhibitor. Arachidonic acid rescued cells treated with antibodies to the FGF receptor or the PLC inhibitor, but not cells in which the activity of PKC, p59(fyn), FAK, Ras, or MEK was inhibited. Interaction of NCAM with a synthetic NCAM peptide ligand, known to induce neurite outgrowth, was shown to stimulate phosphorylation of the MAP kinases extracellular signal-regulated kinases ERK1 and ERK2. The MAP kinase activation was sustained, because ERK1 and ERK2 were phosphorylated in PC12-E2 cells and primary hippocampal neurons even after 24 hr of cultivation on NCAM-expressing fibroblasts. Based on these results, we propose a model of NCAM signaling involving two pathways: NCAM-Ras-MAP kinase and NCAM-FGF receptor-PLCgamma-PKC, and we propose that PKC serves as the link between the two pathways activating Raf and thereby creating the sustained activity of the MAP kinases necessary for neuronal differentiation. PMID- 10704500 TI - Late retinal progenitor cells show intrinsic limitations in the production of cell types and the kinetics of opsin synthesis. AB - The seven major cell classes of the vertebrate neural retina arise from a pool of multipotent progenitor cells. Several studies suggest a model of retinal development in which both the environment and the progenitor cells themselves change over time (). To test this model, we used a reaggregate culture system in which a labeled population of progenitor cells from the postnatal rat retina were cultured with an excess of embryonic retinal cells. The labeled cells were then assayed for their cell fate choices and their kinetics of rod differentiation, as measured by opsin synthesis. The kinetics of opsin synthesis remained unchanged, but fewer postnatal cells adopted the rod cell fate when cultured with embryonic cells. There was an increase in the percentage of bipolar cells produced by postnatal progenitor cells, indicating a possible respecification of fate. The increase in bipolar cells could occur even after progenitor cells had completed their terminal mitoses. These alterations in cell fates appeared to be caused at least in part by a secreted factor released by the embryonic cells that requires the LIFRbeta/gp130 complex for signaling. Finally, although surrounded by 20-fold more embryonic cells, the postnatal cells did not choose to adopt any fates normally produced only by embryonic cells. PMID- 10704501 TI - Acute changes in maternal thyroid hormone induce rapid and transient changes in gene expression in fetal rat brain. AB - Despite clinical evidence that thyroid hormone is essential for brain development before birth, effects of thyroid hormone on the fetal brain have been largely unexplored. One mechanism of thyroid hormone action is regulation of gene expression, because thyroid hormone receptors (TRs) are ligand-activated transcription factors. We used differential display to identify genes affected by acute T(4) administration to the dam before the onset of fetal thyroid function. Eight of the 11 genes that we identified were selectively expressed in brain areas known to contain TRs, indicating that these genes were directly regulated by thyroid hormone. Using in situ hybridization, we confirmed that the cortical expression of both neuroendocrine-specific protein (NSP) and Oct-1 was affected by changes in maternal thyroid status. Additionally, we demonstrated that both NSP and Oct-1 were expressed in the adult brain and that their responsiveness to thyroid hormone was retained. These data are the first to identify thyroid hormone-responsive genes in the fetal brain. PMID- 10704502 TI - Growth cones are not required for initial establishment of polarity or differential axon branch growth in cultured hippocampal neurons. AB - Hippocampal neurons developing in culture exhibit two types of differential, seemingly competitive, process outgrowth in the absence of external cues. During the initial acquisition of polarity, one of several equivalent undifferentiated minor neurites preferentially grows to become the axon. Once the axon has formed, it typically branches, and the branches grow differentially rather than concurrently. In axons with only two branches, growth alternates between branches. In both axon establishment and branch growth alternation, growth among sibling processes or branches must be differentially regulated. We found that elaborate and dynamic growth cones were associated with growth, whereas diminished growth cones were associated with nongrowing processes or branches. To test whether growth cones were necessary for differential growth, growth cone motility was eliminated by application of cytochalasin E. Although cytochalasin treatment before axon formation yielded longer processes overall, a similar percentage of both treated and untreated neurons had one process that grew more rapidly and became much longer than its sibling processes. Immunostaining to visualize dephospho-tau, an axonal marker, demonstrated that these single dominant processes were axons. Axons that formed in cytochalasin were thicker and showed more intense anti-tubulin staining than their sibling processes. Branched axons deprived of growth cones retained a pattern of differential growth and often included alternation. These results indicate that neither formation of a single axon nor differential growth of branches are dependent on growth cone motility and suggest that the neuron can regulate neurite elongation at sites other than at the growth cone. PMID- 10704503 TI - Application of neutralizing antibodies against NI-35/250 myelin-associated neurite growth inhibitory proteins to the adult rat cerebellum induces sprouting of uninjured purkinje cell axons. AB - The myelin-associated proteins NI-35/250 exert a powerful inhibition on axon regeneration, but their function exerted on intact neurons is still unclear. In the adult CNS these proteins are thought to regulate axon growth processes to confine plasticity within restricted regions and to prevent the formation of aberrant connections. We have recently shown that application of neutralizing IN 1 antibody Fab fragment against NI-35/250 proteins to the adult cerebellum induces the expression of injury/growth-associated markers in intact Purkinje cells. Here, we asked whether these cellular modifications are accompanied by growth phenomena of Purkinje neurites. A single intraparenchymal application of IN-1 Fab fragment to the adult cerebellum induces a profuse sprouting of Purkinje axons along their intracortical course. The newly formed processes spread to cover most of the granular layer depth. A significant axon outgrowth is evident 2 d after injection; it tends to increase at 5 and 7 d, but it is almost completely reversed after 1 month. No axonal modifications occur in control Fab-treated cerebella. The IN-1 Fab fragment-induced cellular changes and axon remodeling are essentially reproduced by applying affinity-purified antibody 472 raised against a peptide sequence of the recombinant protein NI-220, thus confirming the specificity of the applied treatments on these myelin-associated molecules. Functional neutralization of NI-35/250 proteins induces outgrowth from uninjured Purkinje neurites in the adult cerebellum. Together with previous observations, this suggests that these molecules regulate axonal plasticity to maintain the proper targeting of terminal arbors within specific gray matter regions. PMID- 10704504 TI - DM-GRASP is necessary for nonradial cell migration during chick diencephalic development. AB - Cell migration is fundamental to normal CNS development. Radial migration, along radial glial fibers, has been the principal pathway studied, however, nonradial or tangential cell migration has increasingly been identified at all levels of the CNS. Receptors, cell adhesion molecules, and extracellular matrix molecules have all been shown to participate in radial cell migration. In contrast, the molecular basis of nonradial cell migration has only recently begun to be elucidated. Using replication defective retroviral vectors we have determined the location and time when nonradial cell migration begins in the developing chick diencephalon. We have identified three molecules that are expressed in spatially and temporally restricted domains that are consistent with them playing a role in nonradial cell migration. One of these molecules, DM-GRASP, a transmembrane protein with five extracellular Ig domains, is expressed on the nonradially migrating cells in addition to axons. To test the hypothesis that DM-GRASP participates in guiding nonradial cell migration, we injected a replication defective retroviral vector used for lineage tracing followed by a DM-GRASP blocking antibody. Embryos injected with the blocking antibody showed a near complete block in nonradial cell migration specifically where DM-GRASP is expressed. Furthermore, morphological analyses revealed disruption of the normal architecture of the diencephalon indicating nonradial cell migration is necessary for normal morphological development of the brain. Our data indicate that DM GRASP is necessary for nonradial cell migration in the chick diencephalon and have provided a system to further explore the function of nonradial cell migration during CNS development. PMID- 10704505 TI - The flathead mutation causes CNS-specific developmental abnormalities and apoptosis. AB - We describe a new mutation, flathead (fh), that arose spontaneously in an inbred colony of Wistar rats. The mutation is autosomal recessive, and the behavioral phenotype of fh/fh rats includes spontaneous seizures, tremor, impaired coordination, and premature death. A striking feature of the fh mutation is a dramatic reduction in brain size (40% of normal at birth). In contrast, no abnormalities are evident in the peripheral nervous system or in other tissues outside of the CNS. Although bromodeoxyuridine incorporation assays indicate that the rate of cell proliferation in the fh/fh cortex is similar to that of unaffected animals, in situ terminal deoxynucleotidyl transferase-mediated dUTP biotin end-labeling assays reveal a dramatic increase in apoptotic cell death beginning after embryonic day 16 (E16). At E18 there is a 20-fold increase in cell death in the ventricular zone of fh/fh neocortex, and at postnatal day 1 (P1), the number of apoptotic cells is still two times that of normal. However, by P8 the extent of cell death in fh/fh is comparable to that of unaffected littermates, indicating that the reduction in brain growth is caused by abnormally high apoptosis during a discrete developmental period. Late-developing structures such as the cerebellum, neocortex, hippocampus, and retina are most severely affected by the fh mutation. Within these structures, later-generated neuronal populations are selectively depleted. Together, these results suggest that the flathead gene is essential for a developmental event required for the generation and maturation of late-born cell populations in the brain. PMID- 10704506 TI - Organization of cortical activities related to movement in humans. AB - The extent and function of synchronization of oscillatory elements in the human sensorimotor cortex during movement remains unclear. Here we determine whether synchronization is distributed in both the spatial and frequency domains and whether it changes according to task. Electrocorticographic (ECoG) signals were recorded from presumed nonpathological areas simultaneously with electromyographic (EMG) signals from upper limb muscles during isometric and phasic movement tasks in humans with subdural electrodes in situ for investigation of epilepsy. Functional mapping of the sensorimotor cortex was performed by previous electrical stimulation through the same ECoG electrodes used for recording. Significant coherence between ECoG and EMG was seen at discrete frequencies in the range of 7-100 Hz. There was no predilection for coherence within a given frequency band to be associated with cortical sites that had been functionally defined as producing contralateral arm motor responses on stimulation. However, coherence with muscle in the 7-14 and 15-30 Hz band tended to be associated with ECoG sites that lay close to or within the central sulcus as determined intraoperatively. The spatial pattern and frequency of coherence changed with different tasks, although similarities in the coherence pattern remained for tasks that shared common features. These findings provide support for the hypothesis that that synchronization at specific frequencies links cortical activities into a functional ensemble during voluntary movement. PMID- 10704507 TI - Spectral-temporal receptive fields of nonlinear auditory neurons obtained using natural sounds. AB - The stimulus-response function of many visual and auditory neurons has been described by a spatial-temporal receptive field (STRF), a linear model that for mathematical reasons has until recently been estimated with the reverse correlation method, using simple stimulus ensembles such as white noise. Such stimuli, however, often do not effectively activate high-level sensory neurons, which may be optimized to analyze natural sounds and images. We show that it is possible to overcome the simple-stimulus limitation and then use this approach to calculate the STRFs of avian auditory forebrain neurons from an ensemble of birdsongs. We find that in many cases the STRFs derived using natural sounds are strikingly different from the STRFs that we obtained using an ensemble of random tone pips. When we compare these two models by assessing their predictions of neural response to the actual data, we find that the STRFs obtained from natural sounds are superior. Our results show that the STRF model is an incomplete description of response properties of nonlinear auditory neurons, but that linear receptive fields are still useful models for understanding higher level sensory processing, as long as the STRFs are estimated from the responses to relevant complex stimuli. PMID- 10704508 TI - Amphetamine withdrawal alters bistable states and cellular coupling in rat prefrontal cortex and nucleus accumbens neurons recorded in vivo. AB - Repeated amphetamine administration is known to produce changes in corticoaccumbens function that persist beyond termination of drug administration. We have found previously that long-term alteration in dopamine systems leads to changes in gap junction communication, expressed as dye coupling, between striatal neurons. In this study, the cellular bases of amphetamine-induced changes were examined using in vivo intracellular recordings and dye injection in ventral prefrontal-accumbens system neurons of control and amphetamine-treated rats. Rats that had been withdrawn from repeated amphetamine displayed a significant increase in the incidence of dye coupling in the prefrontal cortex and nucleus accumbens, which persisted for up to 28 d after withdrawal. The increased coupling was limited to projection neurons in both prefrontal cortical and accumbens brain regions, as identified by their axonal trajectory or the absence of interneuron-selective immunocytochemical markers. These changes occurred with no substantial loss of tyrosine hydroxylase-immunoreactive terminals in these cortical and accumbens regions, ruling out dopamine degeneration as a precipitating factor. Previous studies showed that nitric oxide plays a role in the regulation of coupling; however, amphetamine-withdrawn rats had fewer numbers of neurons and processes that stained for nitric oxide synthase immunoreactivity. In amphetamine-treated rats, a higher proportion of cortical cells fired in bursts, and a larger proportion of accumbens and prefrontal cortical neurons exhibited bistable membrane oscillations. By increasing corticoaccumbens transmission, amphetamine withdrawal may lead to neuronal synchronization via gap junctions. Furthermore, this adaptation to amphetamine treatment persists long after the drug is withdrawn. PMID- 10704509 TI - Functional specificity of callosal connections in tree shrew striate cortex. AB - Although callosal connections have been shown to link extensive regions of primary visual cortex, the distribution of these connections with respect to the map of visual space and the map of orientation preference remains unclear. Here we combine optical imaging of intrinsic signals with injection of fluorescent microspheres to assess the functional specificity of callosal connections in the tree shrew. By imaging both hemispheres simultaneously while presenting a series of spatially restricted stimuli, we find that a substantial region of visual space is represented bilaterally. Each hemisphere includes a representation of the ipsilateral visual field that is highly compressed relative to that of the contralateral visual field and is most extensive in the lower visual field, where approximately 30(o) of central visual space are represented bilaterally. Callosal connections extend throughout the region of bilateral representation but terminate in a spatially restricted manner that links visuotopically corresponding sites in the two hemispheres. In contrast, callosal connections appear to terminate without regard for the map of orientation preference, showing little sign of the orientation-specific modular and axial specificity that is characteristic of long-range horizontal connections. By coordinating the activity in the two hemispheres in a way that preserves nearest neighbor relationships, callosal connections may best be viewed as elements of local circuits that operate within a single bilateral representation of visual space. PMID- 10704510 TI - Curvature of visual space under vertical eye rotation: implications for spatial vision and visuomotor control. AB - Most models of spatial vision and visuomotor control reconstruct visual space by adding a vector representing the site of retinal stimulation to another vector representing gaze angle. However, this scheme fails to account for the curvatures in retinal projection produced by rotatory displacements in eye orientation. In particular, our simulations demonstrate that even simple vertical eye rotation changes the curvature of horizontal retinal projections with respect to eye-fixed retinal landmarks. We confirmed the existence of such curvatures by measuring target direction in eye coordinates in which the retinotopic representation of horizontally displaced targets curved obliquely as a function of vertical eye orientation. We then asked subjects to point (open loop) toward briefly flashed targets at various points along these lines of curvature. The vector-addition model predicted errors in pointing trajectory as a function of eye orientation. In contrast, with only minor exceptions, actual subjects showed no such errors, showing a complete neural compensation for the eye position-dependent geometry of retinal curvatures. Rather than bolstering the traditional model with additional corrective mechanisms for these nonlinear effects, we suggest that the complete geometry of retinal projection can be decoded through a single multiplicative comparison with three-dimensional eye orientation. Moreover, because the visuomotor transformation for pointing involves specific parietal and frontal cortical processes, our experiment implicates specific regions of cortex in such nonlinear transformations. PMID- 10704511 TI - Striatonigrostriatal pathways in primates form an ascending spiral from the shell to the dorsolateral striatum. AB - Clinical manifestations in diseases affecting the dopamine system include deficits in emotional, cognitive, and motor function. Although the parallel organization of specific corticostriatal pathways is well documented, mechanisms by which dopamine might integrate information across different cortical/basal ganglia circuits are less well understood. We analyzed a collection of retrograde and anterograde tracing studies to understand how the striatonigrostriatal (SNS) subcircuit directs information flow between ventromedial (limbic), central (associative), and dorsolateral (motor) striatal regions. When viewed as a whole, the ventromedial striatum projects to a wide range of the dopamine cells and receives a relatively small dopamine input. In contrast, the dorsolateral striatum (DLS) receives input from a broad expanse of dopamine cells and has a confined input to the substantia nigra (SN). The central striatum (CS) receives input from and projects to a relatively wide range of the SN. The SNS projection from each striatal region contains three substantia nigra components: a dorsal group of nigrostriatal projecting cells, a central region containing both nigrostriatal projecting cells and its reciprocal striatonigral terminal fields, and a ventral region that receives a specific striatonigral projection but does not contain its reciprocal nigrostriatal projection. Examination of results from multiple tracing experiments simultaneously demonstrates an interface between different striatal regions via the midbrain dopamine cells that forms an ascending spiral between regions. The shell influences the core, the core influences the central striatum, and the central striatum influences the dorsolateral striatum. This anatomical arrangement creates a hierarchy of information flow and provides an anatomical basis for the limbic/cognitive/motor interface via the ventral midbrain. PMID- 10704512 TI - Peripheral odor coding in the rat and frog: quality and intensity specification. AB - In mammals, two recent studies have shown recently that one odor molecule can be recognized by several molecular olfactory receptors (ORs), and a single OR can recognize multiple odor molecules. In addition, one olfactory receptor neuron (ORN) may respond to different stimuli chosen as representative of distinct odor qualities. The aim of the present study was to analyze quality and intensity coding abilities of rat single ORNs, comparing them with previous extensive data gathered in the frog to get insight into the generality of olfactory coding mechanisms over vertebrates. Response properties of 90 rat ORNs to different odors or to one odor at different concentrations were analyzed. In the rat and the frog, odor quality appears to be specified through the identity of activated ORNs. However, rat ORNs have higher response thresholds. This lower sensitivity may be interpreted as an increase in selectivity of rat ORNs for low or medium odor intensities. In these conditions, the lower proportion of activated ORNs could be counterbalanced by their number, as well as by their higher glomerular convergence ratio in the olfactory bulb. From amphibians to mammals, the olfactory system appears to use universal mechanisms based on a combinatorial coding mode that may allow quasi-infinite possibilities of adaptation to various olfactory environments. PMID- 10704513 TI - Response characteristics of an identified, sexually dimorphic olfactory glomerulus. AB - Partitioning of synaptic neuropil into glomeruli is a common feature of primary olfactory centers in most animal species. The functional significance of glomeruli, however, is not yet well understood. The present study is part of our effort to test the hypothesis that each glomerulus is a functional unit dedicated to processing information about a particular odorant or attribute of odor molecules and that the glomerular array constitutes a map of "odor space." We investigated the physiological and morphological features of uniglomerular projection neurons (PNs) associated with an identified glomerulus in each antennal lobe of the female sphinx moth, Manduca sexta. This "lateral large female glomerulus" (latLFG) is sexually dimorphic and therefore may play a female specific role, such as processing of information about one or more odorants important for orientation of a female to host plants for oviposition. Together with the medial LFG (medLFG), the latLFG resides outside the array of spheroidal ordinary glomeruli, near the entrance of the antennal (olfactory) nerve. Each LFG is innervated by four to five PNs. Using intracellular recording and staining, we examined the responses of latLFG-PNs to odorants that represent major classes of volatiles released by host plants of M. sexta. All latLFG-PNs were excited when the ipsilateral antenna was stimulated with low concentrations of the monoterpenoid linalool. Dose-response analysis showed that neither other monoterpenoids nor representatives of other classes of host plant volatiles were similarly stimulatory to latLFG-PNs. These findings are consistent with the idea that each glomerulus has a characteristic, limited molecular receptive range. PMID- 10704514 TI - Transformations of an auditory temporal code in the medulla of a sound-producing fish. AB - The fish auditory system provides important insights into the evolution and mechanisms of vertebrate hearing. Fish have relatively simple auditory systems, without a cochlea for mechanical frequency analysis. However, as in all vertebrates, the primary auditory afferents of fish represent sounds as stimulus entrained spike trains. Thus, fish provide important models for studying how temporal spiking patterns are used in higher level neural computations. In this paper we demonstrate that one of the fundamental transformations of information in the auditory system of a sound-producing fish, Pollimyrus, takes place in the auditory medulla. We discovered a class of neurons in which evoked spiking patterns were relatively independent of the stimulus fine structure and appeared to reflect intrinsic properties of the neurons. These neurons generated sustained responses but were poorly phase-locked to tones compared with the primary afferents. The interval histograms showed that spike timing was regular. However, in contrast to primary afferents, the mode of the interspike interval distribution was independent of the period of tonal stimuli. The tuning of the neurons was broad, with best sensitivity in the same spectral region where these animals concentrate energy in their communication sounds. The physiology of these neurons was similar to that of the chopper neurons known in the auditory brainstem of mammals. Our findings suggest that this medullary transformation, from phase-locked afferent input to chopper-like physiology, is basic to vertebrate auditory processing, even within lineages that have not evolved a cochlea. PMID- 10704515 TI - Novel role for the NMDA receptor redox modulatory site in the pathophysiology of seizures. AB - Redox-active compounds modulate NMDA receptors (NMDARs) such that reduction of NMDAR redox sites increases, and oxidation decreases, NMDAR-mediated activity. Because NMDARs contribute to the pathophysiology of seizures, redox-active compounds also may modulate seizure activity. We report that the oxidant 5, 5' dithio-bis(2-nitrobenzoic acid) (DTNB) and the redox cofactor pyrroloquinoline quinone (PQQ) suppressed low Mg(2+)-induced hippocampal epileptiform activity in vitro. Additionally, in slices exposed to 4-7 microM bicuculline, DTNB and PQQ reversed the potentiation of evoked epileptiform responses by the reductants dithiothreitol and Tris(2-carboxyethyl)phosphine (TCEP). NMDA-evoked whole-cell currents in CA1 neurons in slices were increased by TCEP and subsequently decreased by DTNB or PQQ at the same concentrations that modulated epileptiform activity. However, DTNB and PQQ had little effect on baseline NMDA-evoked currents in control medium, and PQQ did not alter NMDAR-dependent long-term potentiation. In contrast, in slices returned to control medium after low Mg(2+) induced ictal activity, DTNB significantly inhibited NMDAR-mediated currents, indicating endogenous reduction of NMDAR redox sites under this epileptogenic condition. These data suggested that PQQ and DTNB suppressed spontaneous ictal activity by reversing pathological NMDAR redox potentiation without inhibiting physiological NMDAR function. In vivo, PQQ decreased the duration of chemoconvulsant-induced seizures in rat pups with no effect on baseline behavior. Our results reveal endogenous potentiation of NMDAR function via mass reduction of redox sites as a novel mechanism that may enhance epileptogenesis and facilitate the transition to status epilepticus. The results further suggest that redox-active compounds may have therapeutic use by reversing NMDAR-mediated pathophysiology without blocking physiological NMDAR function. PMID- 10704516 TI - Modest neuropsychological deficits caused by reduced noradrenaline metabolism in mice heterozygous for a mutated tyrosine hydroxylase gene. AB - Tyrosine hydroxylase (TH) is the initial and rate-limiting enzyme for the biosynthesis of catecholamines that are considered to be involved in a variety of neuropsychiatric functions. Here, we report behavioral and neuropsychological deficits in mice carrying a single mutated allele of the TH gene in which TH activity in tissues is reduced to approximately 40% of the wild-type activity. In the mice heterozygous for the TH mutation, noradrenaline accumulation in brain regions was moderately decreased to 73-80% of the wild-type value. Measurement of extracellular noradrenaline level in the frontal cortex by the microdialysis technique showed a reduction in high K(+)-evoked noradrenaline release in the mutants. The mutant mice displayed impairment in the water-finding task associated with latent learning performance. They also exhibited mild impairment in long-term memory formation in three distinct forms of associative learning, including active avoidance, cued fear conditioning, and conditioned taste aversion. These deficits were restored by the drug-induced stimulation of noradrenergic activity. In contrast, the spatial learning and hippocampal long term potentiation were normal in the mutants. These results provide genetic evidence that the central noradrenaline system plays an important role in memory formation, particularly in the long-term memory of conditioned learning. PMID- 10704517 TI - Anticipatory biasing of visuospatial attention indexed by retinotopically specific alpha-band electroencephalography increases over occipital cortex. AB - Alpha-band (8-14 Hz) oscillatory EEG activity was examined with high-density scalp electrical recording during the cue-stimulus interval of an endogenous spatial cueing paradigm. In different blocks, cued spatial locations (left or right) were in either the upper or lower visual field, and attended stimuli were either oriented Ts or moving dots. Distractor stimuli were equally likely in the uncued hemifield. Sustained focal increases of alpha-band activity were seen over occipital cortex contralateral to the direction of the to-be-ignored location (ipsilateral to the cued direction of attention) before onset of the to-be attended stimulus. The focus of alpha-band activity also moved depending on whether cued locations were in the upper or lower field. Results are consistent with active gating of uncued spatial locations. PMID- 10704518 TI - The left hemisphere's role in hypothesis formation. AB - In a probability guessing experiment, subjects try to guess which of two events will occur next. Humans tend to match the frequency of previous occurrences in their guesses. Animals other than humans tend to maximize or always choose the option that has occurred the most frequently in the past. Investigators have argued that frequency matching results from the attempt of humans to find patterns in sequences of events even when told the sequences are random. There is independent evidence that the left hemisphere of humans houses a cognitive mechanism that tries to make sense of past occurrences. We performed a probability guessing experiment with two split-brain patients and found that they approximated frequency matching in their left hemispheres and approached maximizing in their right hemispheres. We obtained a conceptual replication of that finding on patients with unilateral damage to either the left or right hemisphere. We conclude that the neural processes responsible for searching for patterns in events are housed in the left hemisphere. PMID- 10704519 TI - Methylphenidate enhances working memory by modulating discrete frontal and parietal lobe regions in the human brain. AB - The indirect catecholamine agonist methylphenidate (Ritalin) is the drug treatment of choice in attention deficit/hyperactivity disorder (AD/HD), one of the most common behavioral disorders of childhood (DSM-IV), although symptoms may persist into adulthood. Methylphenidate can enhance cognitive performance in adults and children diagnosed with AD/HD (Kempton et al., 1999; Riordan et al., 1999) and also in normal human volunteers on tasks sensitive to frontal lobe damage, including aspects of spatial working memory (SWM) performance (Elliott et al., 1997). The present study investigated changes in regional cerebral blood flow (rCBF) induced by methylphenidate during performance of a self-ordered SWM task to define the neuroanatomical loci of the beneficial effect of the drug. The results show that the methylphenidate-induced improvements in working memory performance occur with task-related reductions in rCBF in the dorsolateral prefrontal cortex and posterior parietal cortex. The beneficial effects of methylphenidate on working memory were greatest in the subjects with lower baseline working memory capacity. This is to our knowledge the first demonstration of a localization of a drug-induced improvement in SWM performance in humans and has relevance for understanding the treatment of AD/HD. PMID- 10704520 TI - Chronic jet lag produces cognitive deficits. AB - Traveling across time zones causes disruption to the normal circadian rhythms and social schedules because of travelers' shift in time. As the endogenous circadian timing system adapts slowly to new time cues, the phase relationship between biological rhythms and external time cues are out of synchronization for a period of time. This disturbance of circadian rhythms has been shown to impair physical and psychological health (Winget et al., 1984). To test the effects of repeated jet lag on mental abilities, airline cabin crew were compared with ground crew. Salivary cortisol was used as a physiological marker for circadian disruption. The cabin crew group, who had a history of repeated jet lag, had significantly higher salivary cortisol levels in an average working day. In addition, this elevated level of cortisol was only seen in the same subjects when the cabin crew were on transmeridian flights but not domestic flights. Cabin crew also exhibited cognitive deficits, possibly in working memory, that became apparent after several years of chronic disruption of circadian rhythms. PMID- 10704521 TI - Glycosylated major urinary protein of the house mouse: characterization of its N linked oligosaccharides. AB - A minor component of the major urinary protein complex of the house mouse was chromatographically isolated and ascertained to be a previously suspected glycoprotein. Using highly sensitive mass-spectrometric techniques for sequencing and linkage analysis, the N-linked oligosaccharides of this glycoprotein were characterized. They were determined to be of the complex type with a wide heterogeneity. The heterogeneity was due to both the degree of sialylation and the presence of galactose residues in either beta(1-3) or beta(1-4) linkages. The biantennary structures were the most pronounced glycans, while tri- and tetraantennary entities were minor. PMID- 10704522 TI - Interaction between galectin-3 and FcgammaRII induces down-regulation of IL-5 gene: implication of the promoter sequence IL-5REIII. AB - Our previous work demonstrated the capacity of galectin-3 (a beta-galactoside binding animal lectin) to inhibit IL-5 gene expression in different cell types, but the interaction of lectin with the cells and the pathways for the inhibition process are unknown. One of the purposes of this work was to study the cellular ligand for galectin-3. We have demonstrated that galectin-3 can bind to the low affinity IgG receptor (FcgammaRII or CD32) by using different experimental approaches, such as flow cytometry, fusion protein GST technology, and with a model of FcgammaRII-deficient mice. To further analyze the interaction between FcgammaRII and galectin-3, and its implication in IL-5 gene down-regulation we used FcgammaRII-deficient mice. When PBMC from these mice were incubated with galectin-3, the expression of the IL-5 gene was unchanged. However, when PBMC from wild type mice and FcgammaRIII-deficient mice were incubated with galectin 3, IL-5 gene expression was down-regulated. Finally, we studied the implication of the negative regulatory sequence in the IL-5 gene promoter. In the presence of galectin-3, a DNA-protein complex was formed with the IL-5REIII region. This complex was not observed when unrelated oligonucleotide was used. So, galectin-3 induces a pathway, which activates a transcription factor that binds to IL 5REIII. This interaction is capable of inhibiting IL-5 gene transcription. PMID- 10704523 TI - Protein structure controls the processing of the N-linked oligosaccharides and glycosylphosphatidylinositol glycans of variant surface glycoproteins expressed in bloodstream form Trypanosoma brucei. AB - The variant surface glycoproteins (VSGs) of Trypanosoma brucei are a family of homodimeric glycoproteins that adopt similar shapes. An individual trypanosome expresses one VSG at a time in the form of a dense protective mono-layer on the plasma membrane. VSG genes are expressed from one of several polycistronic transcription units (expression sites) that contain several expression site associated genes. We used a transformed trypanosome clone expressing two different VSGs (VSG121 and VSG221) from the same expression site (that of VSG221) to establish whether the genotype of the trypanosome clone or the VSG structure itself controls VSG N-linked oligosaccharide and GPI anchor glycan processing. In gel release and fluorescent labeling of N-linked oligosaccharides and on-blot fluorescent labeling and release of GPI anchor glycans were employed to compare the carbohydrate structures of VSG121 and VSG221 when expressed individually in wild-type trypanosome clones and when expressed together in the transformed trypanosome clone. The data indicate that the genotype of the trypanosome clone has no effect on the N-linked oligosaccharide structures present on a given VSG variant and only a minor effect on the GPI anchor glycans. The latter is most likely an effect of changes in inter-VSG packing when two VGSs are expressed simultaneously. Thus, N-linked oligosaccharide and GPI anchor processing enzymes appear to be constitutively expressed in bloodstream form African trypanosomes and the tertiary and quaternary structures of the VSG homodimers appear to dictate the processing and glycoform microheterogeneity of surface-expressed VSGs. PMID- 10704524 TI - Human alpha-N-acetylgalactosaminidase: site occupancy and structure of N-linked oligosaccharides. AB - Human alpha-N-acetylgalactosaminidase (alpha-GalNAc; also known as alpha galactosidase B) is the lysosomal exoglycohydrolase that cleaves alpha-N acetylgalactosaminyl moieties in glycoconjugates. Mutagenesis studies indicated that the first five (N124, N177, N201, N359, and N385) of the six potential N glycosylation sites were occupied. Site 3 occupancy was important for enzyme function and stability. Characterization of the N-linked oligosaccharide structures on the secreted enzyme overexpressed in Chinese hamster ovary cells revealed highly heterogeneous structures consisting of complex (approximately 53%), hybrid (approximately 12%), and high mannose-type (approximately 33%) oligosaccharides. The complex structures were mono-, bi-, 2,4-tri-, 2,6-tri-, and tetraantennary, among which the biantennary structures were most predominant (approximately 53%). Approximately 80% of the complex oligo-saccharides had a core-region fucose and 50% of the complex oligosaccharides were sialylated exclusively with alpha-2,3-linked sialic acid residues. The majority of hybrid type oligo-saccharides were GalGlcNAcMan(6)GlcNAc-Fuc(0-1)GlcNAc. Approximately 54% of the hybrid oligosaccharide were phosphorylated and one-third of these structures were further sialylated, the latter representing unique phosphorylated and sialylated structures. Of the high mannose oligosaccharides, Man(5 7)GlcNAc(2) were the predominant species (approximately 90%) and about 50% of the high mannose oligosaccharides were phosphorylated, exclusively as monoesters whose positions were determined. Comparison of the oligosaccharide structures of alpha-GalNAc and alpha-galactosidase A, an evolutionary-related and highly homologous exoglycosidase, indicated that alpha-GalNAc had more completed complex chains, presumably due to differences in enzyme structure/domains, rate of biosynthesis, and/or aggregation of the overexpressed recombinant enzymes. PMID- 10704525 TI - Identification of a novel UDP-Glc:GlcNAc beta1-->4-glucosyltransferase in Lymnaea stagnalis that may be involved in the synthesis of complex-type oligosaccharide chains. AB - Several studies suggest, that the snail Lymnaea stagnalis contains glycoproteins whose oligosaccharide side chains have structural features not commonly found in mammalian glycoproteins. In this study, prostate glands of L. stagnalis were incubated in media containing either [(3)H]-mannose, [(3)H]-glucosamine, or [(3)H]-galactose, and the metabolically radiolabeled protein-bound oligosaccharides were analyzed. The newly synthesized diantennary-like complex type asparagine-linked chains contained a considerable amount of glucose, next to mannose, GlcNAc, fucose, galactose, and traces of GalNAc. Since glucose has not been found before as a constituent of diantennary N-linked glycans as far as we know, we assayed the prostate gland of L. stagnalis for a potential glucosyltransferase activity involved in the biosynthesis of such structures. We report here, that the prostate gland of L. stagnalis contains a beta1-->4 glucosyltransferase activity that transfers glucose from UDP-glucose to acceptor substrates carrying a terminal N-acetylglucosamine. The enzyme prefers substrates carrying a terminal GlcNAc that is beta6 linked to a Gal or a GalNAc, structures occurring in O-linked glycans, or a GlcNAc that is beta2 linked to mannose, as is present in N-linked glycans. Based on combined structural and enzymatic data, we propose that the novel beta1-->4-gluco-syltransferase present in the prostate gland may be involved in the biosynthesis of Glcbeta1-->4GlcNAc units in complex type glycans, in particular in N-linked diantennary glycans. PMID- 10704526 TI - Microanalysis of enzyme digests of hyaluronan and chondroitin/dermatan sulfate by fluorophore-assisted carbohydrate electrophoresis (FACE). AB - Hyaluronan and chondroitin/dermatan sulfate are glycosaminoglycans that play major roles in the biomechanical properties of a wide variety of tissues, including cartilage. A chondroitin/dermatan sulfate chain can be divided into three regions: (1) a single linkage region oligosaccharide, through which the chain is attached to its proteoglycan core protein, (2) numerous internal repeat disaccharides, which comprise the bulk of the chain, and (3) a single nonreducing terminal saccharide structure. Each of these regions of a chondroitin/dermatan sulfate chain has its own level of microheterogeneity of structure, which varies with proteoglycan class, tissue source, species, and pathology. We have developed rapid, simple, and sensitive protocols for detection, characterization and quantitation of the saccharide structures from the internal disaccharide and nonreducing terminal regions of hyaluronan and chondroitin/dermatan sulfate chains. These protocols rely on the generation of saccharide structures with free reducing groups by specific enzymatic treatments (hyaluronidase/chondroitinase) which are then quantitatively tagged though their free reducing groups with the fluorescent reporter, 2-aminoacridone. These saccharide structures are further characterized by modification through additional enzymatic (sulfatase) or chemical (mercuric ion) treatments. After separation by fluorophore-assisted carbohydrate electrophoresis, the relative fluorescence in each band is quantitated with a cooled, charge-coupled device camera for analysis. Specifically, the digestion products identified are (1) unsaturated internal Deltadisaccharides including DeltaDiHA, DeltaDi0S, DeltaDi2S, DeltaDi4S, DeltaDi6S, DeltaDi2,4S, DeltaDi2,6S, DeltaDi4,6S, and DeltaDi2,4,6S; (2) saturated nonreducing terminal disaccharides including DiHA, Di0S, Di4S and Di6S; and (3) nonreducing terminal hexosamines including glcNAc, galNAc, 4S-galNAc, 6S galNAc, and 4, 6S-galNAc. PMID- 10704527 TI - Adaptation of FACE methodology for microanalysis of total hyaluronan and chondroitin sulfate composition from cartilage. AB - Protocols for analyzing the fine structure of hyaluronan and chondroitin sulfate using fluorophore-assisted carbohydrate electrophoresis of 2-aminoacridone derivatized hyaluronidase/chondroitinase digestion products were adapted for direct analysis of previously characterized cartilage-derived samples. The chondroitin sulfate disaccharide compositions for fetal and 68 year human aggrecan from FACE analyses were DeltaDi4S (50%), DeltaDi6S (43%), and DeltaDi0S (7%); and DeltaDi4S (3%), DeltaDi6S (96%), and DeltaDi0S (1%), respectively. The nonreducing terminal structures included predominantly 4S-galNAc with minor amounts of 6S-galNAc and Di6S for the fetal aggrecan sample and, in addition, included 4,6S-galNAc in the 68 year aggrecan sample. FACE analysis of a proteinase K digest of rat chondrosarcoma tissue gave an internal disaccharide composition for its chondroitin sulfate chains of DeltaDi0S (7%) and DeltaDi4S (93%) with no DeltaDi6S and DeltaDi4, 6S detected, while DeltaDiHA from hyaluronan was 5% of the total. Analysis of nonreducing terminal structures indicated the presence of 4S-galNAc (51%), galNAc (27%), and Di4S (22%) with no 4,6S-galNAc or Di6S detected. Unexpectedly, FACE analysis detected putative linkage oligosaccharide structures from the chondroitin sulfate chains including both unsulfated (85%) and 4-sulfated (15%) linkage oligosaccharides. Finally, the number averaged chain length estimated from the ratio of the molar fluorescence of the Deltadisaccharides to that of the nonreducing termini or the linkage oligosaccharide structures was calculated as approximately 16 kDa. A tissue glucose concentration of 0.72 g/l was also measured. These results for both samples as determined by FACE analysis were similar to results previously reported, using more labor and time intensive procedures, validating the FACE protocols. PMID- 10704528 TI - Characterization of the N-linked oligosaccharides of megalin (gp330) from rat kidney. AB - Megalin (gp 330) is a large cell surface receptor expressed on the apical surfaces of epithelial tissues, that mediates the binding and internalization of a number of structurally and functionally distinct ligands. In this paper we report the first detailed structural characterization of megalin-derived oligosaccharides. Using strategies based on mass spectrometric analysis, we have defined the structures of the N-glycans of megalin. The results reveal that megalin glycoprotein is heterogeneously glycosylated. The major N-glycans identified belong to the following two classes: high mannose structures and complex type structures, with complex structures being more abundant than high mannose structures. The major nonreducing epitopes in the complex-type glycans are: GlcNAc, Galbeta1-4GlcNAc (LacNAc), NeuAcalpha2-6Galbeta1-4GlcNAc (sialylated LacNAc), GalNAcbeta1-4[NeuAcalpha2-3]Galbeta1-4GlcNAc (Sd(a)) and Galalpha1 3Galbeta1-4GlcNAc. Most complex structures are characterized by the presence of (alpha1,6)-core fucosylation and the presence of a bisecting GlcNAc residue. PMID- 10704529 TI - Ultrastructural localization of sulfated and unsulfated keratan sulfate in normal and macular corneal dystrophy type I. AB - Keratan sulfate (KS) proteoglycans are of importance for the maintenance of corneal transparency as evidenced in the condition macular corneal dystrophy type I (MCD I), a disorder involving the absence of KS sulfation, in which the cornea becomes opaque. In this transmission electron microscope study quantitative immuno- and histochemical methods have been used to examine a normal and MCD I cornea. The monoclonal antibody, 5-D-4, has been used to localize sulfated KS and the lectin Erythrina cristagalli agglutinin (ECA) to localize poly N acetyllactosamine (unsulfated KS). In normal cornea high levels of sulfated KS were detected in the stroma, Bowman's layer, and Descemet's membrane and low levels in the keratocytes, epithelium and endothelium. Furthermore, in normal cornea, negligible levels of labeling were found for N -acetyllactosamine (unsulfated KS). In the MCD I cornea sulfated KS was not detected anywhere, but a specific distribution of N -acetyllactosamine (unsulfated KS) was evident: deposits found in the stroma, keratocytes, and endothelium labeled heavily as did the disrupted posterior region of Descemet's membrane. However, the actual cytoplasm of cells and the undisrupted regions of stroma revealed low levels of labeling. In conclusion, little or no unsulfated KS is present in normal cornea, but in MCD I cornea the abnormal unsulfated KS was localized in deposits and did not associate with the collagen fibrils of the corneal stroma. This study has also shown that ECA is an effective probe for unsulfated KS. PMID- 10704530 TI - A novel viral alpha2,3-sialyltransferase (v-ST3Gal I): transfer of sialic acid to fucosylated acceptors. AB - The substrate specificity of an alpha2,3-sialyltransferase (v-ST3Gal I) obtained from myxoma virus infected RK13 cells has been determined. Like mammalian sialyltransferase enzymes, the viral enzyme contains the characteristic L- and S sialyl motif sequences in its catalytic domain. Analysis of the deduced amino acid sequences of cloned sialyltransferases suggests that v-ST3Gal I is closely related to mammalian ST3Gal IV. v-ST3Gal I catalyzes the transfer of sialic acid from CMP-NeuAc to Type I (Galbeta1-3GlcNAcbeta) II (Galbeta1-4GlcNAcbeta) and III (Galbeta1-3GalNAcbeta) acceptors. In addition, the viral enzyme also transfers sialic acid to the fucosylated acceptors Lewis(x) and Lewis(a). This substrate specificity is unlike any sialyltransferases described to date, though it is most comparable with those of mammalian ST3Gal IV enzymes. The products from reactions with fucosylated acceptors were characterized by capillary zone electrophoresis, (1)H-NMR spectroscopy and mass spectrometry. They were shown to be 2,3-sialylated Lewis(x) and 2,3-sialylated Lewis(a), respectively. PMID- 10704531 TI - Cloning of the human cDNA which can complement the defect of the yeast mannosyltransferase I-deficient mutant alg 1. AB - The assembly of the lipid-linked oligosaccharide, Glc(3)Man(9)GlcNAc(2)-P-P-Dol, occurs on the rough ER membrane in an ordered stepwise manner. The process is highly conserved among eukaryotes. In order to isolate the human mannosyltransferase I (MT-I) gene involved in the process, we used the Saccharomyces cerevisiae MT-I gene ( ALG1 ), which has already been cloned. On searching the EST database with the amino acid sequence of the ALG1 gene product, we detected seven related human EST clones. A human fetal brain cDNA library was screened by PCR using gene-specific primers based on the EST nucleotide sequences and a 430 bp cDNA fragment was amplified. The cDNA library was rescreened with this 430 bp cDNA, and two cDNA clones (HR1-3 and HR1-4) were isolated and sequenced. On a homology search of the EST database with the nucleotide sequence of HR1-3, we detected a novel human EST clone, AA675921 (GenBank accession number). Based on the nucleotide sequences of AA675921 and HR1-4, we designed gene-specific PCR primers, which allowed to amplify a 1.8 kb cDNA from human fetal brain cDNA. This cDNA was cloned and shown to contain an ORF encoding a protein of 464 amino acids. We designated this ORF as Hmat-1. The amino acid sequence deduced from the Hmat-1 gene showed several highly conserved regions shared with the yeast and nematode MT-I sequences. Furthermore, this 1.8 kb cDNA successfully complemented the S. cerevisiae alg1-1 mutation, indicating that the Hmat-1 gene encodes the human MT-I and that the function of this enzyme was conserved between yeast and human. PMID- 10704532 TI - Glycosaminoglycan conformation: do aqueous molecular dynamics simulations agree with x-ray fiber diffraction? AB - Glycosaminoglycan-protein interactions are biologically important and require an appreciation of glycan molecular shape in solution, which is presently unavailable. In previous studies we found strong similarity between aqueous molecular dynamics (MD) simulations and published x-ray diffraction refinements of hyaluronan. We have applied a similar approach here to chondroitin and dermatan, attempting to clarify some of the issues raised by the x-ray diffraction literature relating to chondroitin and dermatan sulfate. We predict that chondroitin has the same beta(1-->4) linkage conformation as hyaluronan, and that their average beta(1-->3) conformations differ. This is explained by changes in hydrogen-bonding across this linkage, resulting from its axial hydroxyl, causing a different sampling of left-handed helices in chondroitin (2.5- to 3.5 fold) as compared with hyaluronan (3.0- to 4.0-fold). Few right-handed helices, which lack intramolecular hydrogen-bonds, were sampled during our MD simulations. Thus, we propose that the 8-fold helix observed in chondroitin-6-sulfate, represented in the literature as an 8(3) helix (right-handed), though it has never been refined, is more likely to be 8(5) (left-handed) helix. Molecular dynamics simulations implied that (4)C(1) and (2)S(O), but not (1)C(4), forms of iduronate could be used in refinements of dermatan x-ray fiber diffraction patterns. Current models of 8-fold dermatan sulfate chains containing (4)C(1) iduronate refine to right-handed helices, which possess no intramolecular hydrogen-bonds. However, MD simulations predict that models containing (2)S(O) iduronate could provide better (8(5) helix) starting structures for refinement. Thus, the 8-fold dermatan sulfate refinement (8(3) helix) could be in error. PMID- 10704533 TI - Sleepiness, fatigue, and impaired alertness. PMID- 10704534 TI - Neuroanatomical and neurophysiological aspects of sleep: basic science and clinical relevance. AB - This is an exciting period for basic sleep research, because we are now beginning to understand some of the mechanisms controlling the changes in consciousness associated with sleep and wakefulness. Witness the recent discoveries of a probable genetic involvement in narcolepsy, the identification of hypothalamic structures promoting sleep, and the mounting evidence that adenosine is an endogenous sleep factor. We review these and other recent developments that help us understand the neuroanatomical and neurophysiological basis of some sleep disorders. For a detailed discussion of specific sleep disorders and clinical issues, the reader is referred to other sources. Overviews are also available covering rapid eye movement (REM) and non-REM sleep physiology, the role of humoral factors in sleep, and the relationship between the immune system and sleep. In fact, where appropriate, here we draw directly on material from our earlier summaries of work in the field. We begin with a brief review of the organization of sleep and wakefulness to provide the background for the subsequent discussions of the anatomy and neurophysiology of the neural control of different vigilance states and associated sleep disorders. For example, a brief description of the neural mechanisms of REM sleep will be followed by an outline of selected sleep disorders related to REM sleep. In summary, we make no attempt here to include all sleep disorders and only review a few selected examples, those in which there is an understanding based on knowledge of central nervous system physiology. Unfortunately we are not now able to include sleep apnea, because the discovery of sleep apnea not only was a defining moment for clinical sleep research, but to this day remains the principal presenting complaint at some sleep disorder clinics. PMID- 10704535 TI - Interactions of sleep and Parkinson's disease. AB - Sleep disruption represents an important, and clinically relevant facet of Parkinson's Disease [PD]. This review attempts to integrate the current knowledge regarding sleep alterations in PD by examining following: the nature of sleep disturbance in PD, the influence of antiparkinson medication on sleep parameters, the interaction of psychological conditions such as depression and anxiety with sleep, and possible beneficial aspects of sleep in PD. Special emphasis is placed on rapid eye movement sleep behavior disorder and the evidence of it heralding PD and related disorders. PMID- 10704536 TI - Genetics and sleep disorders. AB - This review describes the recent growth of our knowledge in the genetics of these sleep disorders and reports some of our preliminary molecular studies in restless legs syndrome. PMID- 10704537 TI - Melatonin, sleep, and circadian rhythm disorders. AB - In the 1970s and early 1980s, neuroendocrinology was viewed by many neuroscientists as a ""window to the brain" to an understanding of brain function." In psychiatry, many have viewed sleep physiology as a window in biological psychiatry. This is, in part, because sleep is one of the few easily quantifiable functions of interest to psychiatrists. Melatonin is a hormone with powerful effects on behavior particularly circadian and sleep behavior. In contrast with other hormones, the pathophysiology and pathology of abnormal melatonin secretion is poorly understood. In this article, we document the well established phase-shifting and sleep-promoting effects of melatonin and discuss some implications for neuropsychiatrists when the neurophysiology of melatonin goes array. It is both striking and in some ways not surprising that the majority of patients with phase delay syndrome described in our research studies have been misdiagnosed as having depression. The reason for this is elucidated in this article and the information concerning this condition may be helpful to many who are relatively unfamiliar with this particular sleep disorder. We can anticipate that patients with specific neurological disorders may have changes in their melatonin secretion and future research, for example in patients with head injury and conditions such as retinitis pigmentosa may be the basis for reviews a few years hence. PMID- 10704538 TI - Sleep and fatigue. PMID- 10704539 TI - The measurement of fatigue and contributing neuropsychiatric factors. PMID- 10704540 TI - Anterior chamber depth and complications during cataract surgery in eyes with pseudoexfoliation syndrome. AB - PURPOSE: To look for associations of preoperative A-scan ultrasound ocular dimensions with complications during phacoemulsification in eyes with pseudoexfoliation. METHODS: A total of 174 eyes with pseudoexfoliation of 135 patients undergoing planned cataract surgery were included in a prognostic study based on the review of a clinical database. Preoperatively, A-scan ultrasound examination with measurement of anterior chamber depth, lens thickness, and total axial length was performed. Phacoemulsification with implantation of a posterior chamber intraocular lens was performed by a total of five surgeons. Intraoperative complications (zonular dialysis and/or vitreous loss) were correlated with preoperative findings including ultrasound dimensions. Multivariate logistic regression analysis with a generalized estimating equations method was used for statistical analysis. RESULTS: Intraoperative complications occurred in 12 eyes (6.9%) of 11 patients. The anterior chamber was significantly shallower in eyes with than in eyes without complications (mean, 2.36 +/- 0.44 mm vs 2.74 +/- 0.52 mm; P =.013). The differences in lens thickness (4.93 +/- 0.55 mm vs 4.72 +/- 0.54 mm; P =.30) and the differences in axial length (22.92 +/- 1.09 mm vs 23.66 +/- 1.36 mm; P =.07) between the two groups did not reach statistical significance. In eyes with pseudoexfoliation, an anterior chamber depth of less than 2.5 mm was associated with a risk of 13.4% for intraoperative complications compared with an overall incidence of intraoperative complications of 6.9% and an incidence of 2.8% for an anterior chamber depth of 2.5 mm or more. CONCLUSIONS: A small anterior chamber depth may indicate zonular instability in eyes with pseudoexfoliation syndrome and should alert the cataract surgeon to the possibility of intraoperative complications. PMID- 10704541 TI - Phacoemulsification and lens implantation after scleral buckling surgery. AB - PURPOSE: To determine the intraoperative and postoperative complications and best corrected visual acuity outcomes of eyes undergoing phacoemulsification and intraocular lens implantation after retinal detachment repair by the scleral buckling technique. METHODS: The charts of all patients who underwent phacoemulsification and intraocular lens implantation between July 1991 and May 1998 in two surgical practices were reviewed to identify eyes with a history of retinal detachment repaired by the scleral buckling technique. Eyes with a history of pars plana vitrectomy were excluded. Demographic and surgical data, preoperative and postoperative best-corrected visual acuity, and intraoperative and postoperative complications were recorded. RESULTS: We identified 34 eyes of 32 patients. The mean interval from retinal detachment repair to phacoemulsification was 12.4 years. The mean interval from phacoemulsification to final examination was 20 months. Risk factors for retinal detachment included isolated myopia (82%), myopia with lattice retinal degeneration (5.9%), and myopia with trauma (8.8%). One eye (2.9%) had no identifiable risk factors. Final best-corrected visual acuity of 20/40 or better was attained in 29 (85%) of 34 eyes and 20/20 or better in 18 (53%) of the eyes. Of the five eyes with the lowest best-corrected visual acuity, three had a macula-off retinal detachment; one had a posterior capsule opacity, epiretinal membrane, and corneal edema secondary to ocular ischemia; and one had advanced glaucoma. All five eyes still experienced an improvement in best-corrected visual acuity. With regard to complications, one eye had a posterior capsular tear with vitreous loss and another developed a postoperative retinal tear. Posterior capsule opacification requiring laser capsulotomy developed in 13 eyes (38%). No eye developed a retinal redetachment. CONCLUSION: Phacoemulsification and intraocular lens implantation can be performed safely after scleral buckling surgery and excellent best-corrected visual acuity results can be attained in most eyes. No modification of surgical technique is necessary. No retinal redetachment occurred in this series. PMID- 10704542 TI - Prospective intrapatient comparison of extracapsular cataract extraction and lens implantation with and without trabeculectomy. AB - PURPOSE: To determine whether extracapsular cataract extraction and posterior chamber lens implantation combined with trabeculectomy provides better long-term results than extracapsular cataract extraction and lens implantation alone in a group of patients with primary open-angle glaucoma and cataract. METHODS: In a prospective, randomized clinical trial, 35 patients with bilateral, symmetric, primary open-angle glaucoma and visually disabling cataracts were randomly selected to undergo surgery with trabeculectomy in one eye and without in the other. All procedures were performed by a single surgeon in a private practice setting with follow-up for more than 5 years in all cases. RESULTS: After an average of 87 months of follow-up, extracapsular cataract extraction and posterior chamber lens implantation reduced intraocular pressure by 4.4 +/- 3.3 mm Hg, reduced the number of medications by 1.28 +/- 0.86, increased diopter vector of astigmatism by 1.49, and was associated with visual field loss in six of 35 eyes. After an average of 80 months of follow-up, extracapsular cataract extraction and posterior chamber lens implantation with trabeculectomy reduced intraocular pressure by 8.2 +/- 4.6 mm Hg (P =.0001), reduced the number of medications by 1.76 +/- 0.82 (P=.0002), increased diopter vector of astigmatism by 1.14, and was associated with visual field loss in one eye (P =.05). Both groups had similar improvement in visual acuity and perioperative complications. CONCLUSIONS: Extracapsular cataract extraction and posterior chamber lens implantation with trabeculectomy was beneficial in the long-term control of intraocular pressure and in prevention of visual field loss. This procedure should be considered for patients in whom long-term pressure control at a lower level would be beneficial in preventing further optic nerve damage. PMID- 10704544 TI - Intraocular pressure and progression of glaucomatous visual field loss. AB - PURPOSE: To evaluate the relationship between intraocular pressure and visual field progression in patients with primary open-angle glaucoma. METHODS: We prospectively followed 113 patients with early to moderate glaucomatous field damage. Conventional automated static perimetry, high-pass resolution perimetry, and intraocular pressure measurements were carried out at 6-month intervals. The mean and the highest intraocular pressure in the follow-up were compared in stable and progressing patients with each perimetric technique. RESULTS: The mean (+/- SD) follow-up was 4.5 +/- 0.9 years. The mean (+/- SD) intraocular pressure in patients remaining stable with conventional perimetry [18.2 +/- 3.3 mm Hg, n = 81 (71.7%)] was not significantly different (P =.65) from those in whom it progressed (17.9 +/- 3.3 mm Hg, n = 32 [28.3%]). The mean intraocular pressure in patients remaining stable with high-pass resolution perimetry (17. 9 +/- 3.5 mm Hg, n = 63 [55.8%]) was not significantly different (P =.33) from those in whom it progressed (18.5 +/- 3.0 mm Hg, n = 50 [44.2%]). The mean (+/- SD) of the highest (single or three highest) pressure during follow-up for stable and progressing patients with conventional perimetry was not significantly different (22.6 +/- 5.0 and 23.0 +/- 4.6 mm Hg, respectively, P =.76). However, for high pass resolution perimetry, the difference was highly significant (21.6 +/- 4.5 and 24.1 +/- 4.9 mm Hg, respectively, P <. 01). Furthermore, patients who progressed with high-pass resolution perimetry had more damaged baseline fields compared with those who remained stable (P <.01). CONCLUSIONS: The mean level of intraocular pressure does not differentiate glaucoma patients with progressive visual field loss from ones who remained stable. Baseline visual field status and peak intraocular pressure of patients who progress with high-pass resolution perimetry are significantly different from those who remain stable. PMID- 10704543 TI - The effect of brimonidine tartrate on retinal blood flow in patients with ocular hypertension. AB - PURPOSE: To study the effects of topical brimonidine tartrate 0.2%, an alpha(2) agonist ocular hypotensive drug, on retinal capillary blood flow in patients with ocular hypertension. METHODS: The study was a double-masked, randomized, placebo controlled trial set in a tertiary eye center. Ocular hypertensive patients with repeatable intraocular pressures greater than 21 mm Hg and normal visual fields and optic disks were consecutively recruited. After an eye examination, baseline retinal blood flow measurements were made with confocal scanning laser Doppler flowmetry in one study eye. Patients were then randomly assigned to receive either brimonidine or placebo (saline) twice daily for 8 weeks. Blood flow and intraocular pressure measurements were then repeated after 4 and 8 weeks. RESULTS: Seventeen patients were randomly assigned to receive brimonidine, and 14 received placebo. One patient in each group failed to complete the study. The mean group differences in baseline age and intraocular pressure were not statistically significant (59. 23 [+/-10.24] and 52.23 [+/-16.46] years, respectively, and 24.84 [+/-2.08] and 24.56 [+/-2.85] mm Hg, respectively). Brimonidine reduced intraocular pressure by 17.90% and 16.17% at 4 and 8 weeks, respectively, with a significant difference in treatment effect compared with the placebo group (P <.007). The group difference in treatment effect in any of the three hemodynamic parameters velocity, volume, and flow was within 8% and not significantly different at 4 or 8 weeks (P.360). Based on a type I error of 0.05, our study had a power greater than or equal to 75% to detect group differences in treatment effect of greater than or equal to 15% to 20%. CONCLUSIONS: Brimonidine reduces intraocular pressure without altering retinal capillary blood flow in patients with ocular hypertension. PMID- 10704545 TI - Comparison of long-term variability for standard and short-wavelength automated perimetry in stable glaucoma patients. AB - PURPOSE: To quantify and compare, on a point-by-point basis, the long-term variability of standard and short-wavelength automated perimetry in a group of stable glaucoma patients. METHODS: From a group of 53 glaucoma patients experienced in visual field testing, we identified one eye, randomly chosen, from each of 25 glaucoma patients whose condition was found to be stable, based on both standard and short-wavelength automated perimetry visual field criteria. On each of three visits during a period of up to 3 months, each patient performed one standard and one short-wavelength automated perimetry 24-2 visual field in a random order on a Humphrey visual field analyzer. The long-term variability (also referred to as test-retest variability) was defined as the SD of the three threshold decibel values at each test location. The long-term variability for each test point (mean +/- SD) was determined separately for both standard visual fields and short-wavelength automated perimetry. RESULTS: With all 52 test locations of the 24-2 field averaged, the global long-term variability, mean (+/- SD) for standard visual fields and short-wavelength automated perimetry was 2.37 +/- 2.03 dB (95% confidence interval, 2.26-2.48 dB) and 2.92 +/- 2.03 dB (95% confidence interval, 2.81-3.03 dB), respectively (P <.0001). In 16 of the 52 visual field locations, long-term variability on short-wavelength automated perimetry was significantly higher than long-term variability on standard visual fields. In addition, the long-term variability increased with greater distance from the point of fixation for both standard visual fields and short-wavelength automated perimetry. The long-term variability decreased closer to fixation, more for standard visual fields than for short-wavelength automated perimetry. CONCLUSIONS: In a group of stable glaucoma patients, mean long-term variability was 0.55 dB higher for short-wavelength automated perimetry than for standard visual fields. This needs to be taken into consideration when serial visual fields are evaluated for change. PMID- 10704546 TI - Frequency doubling technology perimetry for detection of glaucomatous visual field loss. AB - PURPOSE: To evaluate the ability of frequency doubling technology perimetry to detect early, moderate, and advanced glaucomatous visual field loss. METHODS: In a prospective study, frequency doubling technology perimetry (C-20 full threshold) was performed in the right eye of 254 normal control subjects and 230 patients with early (n = 85), moderate (n = 114), or advanced (n = 31) glaucomatous visual field loss. Previous Humphrey Field Analyzer test results were used to classify glaucomatous visual field loss as early (mean deviation no worse than -6 dB), moderate (mean deviation between -6 and -12 dB) or advanced (mean deviation between -12 and -22 dB). RESULTS: Receiver operating characteristic curves showed 100% sensitivity and specificity (area under the curve, 1.0) for detecting advanced glaucomatous visual field loss, approximately 96% sensitivity and 96% specificity (area under the curve, 0.9751) for detecting moderate glaucomatous visual field loss, and approximately 85% sensitivity and 90% specificity (area under the curve, 0.9261) for early glaucomatous visual field loss. CONCLUSIONS: Frequency doubling technology perimetry demonstrates high sensitivity and specificity for detection of early, moderate, and advanced glaucomatous visual field loss. PMID- 10704547 TI - Algorithm for interpreting the results of frequency doubling perimetry. AB - PURPOSE: To evaluate an algorithm for the identification of glaucomatous visual field defects with the screening mode of frequency doubling technology. METHODS: Screening-mode frequency doubling technology and Swedish interactive threshold algorithm perimetry were performed on 137 of 150 consecutive patients referred to a glaucoma specialist. We created an algorithm for the frequency doubling technology that gave increased importance to both more severe defects and defects closer to fixation. These values were then compared with the results of the Swedish interactive threshold algorithm visual fields evaluated by the glaucoma hemifield test, two masked glaucoma specialists, and a published definition of glaucomatous damage to determine sensitivity and specificity of the frequency doubling technology screening mode for detecting glaucoma. PMID- 10704548 TI - Comparison of frequency doubling perimetry with humphrey visual field analysis in a glaucoma practice. AB - PURPOSE: To determine the sensitivity and specificity of frequency doubling perimetry with Humphrey visual field testing used as the gold standard. METHODS: Frequency doubling perimetry and Humphrey visual field testing (24-2) were performed on 29 consecutive patients in a glaucoma practice. Data for the right eye were used to calculate sensitivity, specificity, and receiver operating characteristic curves. RESULTS: For the frequency doubling perimetry in screening mode, and with an abnormal glaucoma hemifield test used as the gold standard, the area under the receiver operating characteristic curve was 89.3%, 81.5%, or 75.0% for the presence of mild, moderate, or severe relative defects, respectively. Similar results were found with the use of mean deviation (P <.05) to define Humphrey visual field defects. For frequency doubling perimetry in threshold mode, the area under the receiver operating characteristic curve was 93.4% with the presence of any defect (P <.05) used as the criterion for an abnormal case, and an abnormal glaucoma hemifield test as the gold standard. In all cases, the threshold mode detected defects better than the screening mode. PMID- 10704549 TI - Combined pars plana vitrectomy and glaucoma drainage implant placement for refractory glaucoma. AB - PURPOSE: To report visual acuity and intraocular pressure outcomes among patients who have undergone combined pars plana vitrectomy and placement of a glaucoma drainage implant. METHODS: The medical records of all patients who underwent combined pars plana vitrectomy and placement of a glaucoma drainage implant at the Bascom Palmer Eye Institute by one of the authors between January 1, 1990, and February 28, 1998, were reviewed. Forty patients (40 eyes) were identified, including 14 patients with neovascular glaucoma secondary to proliferative diabetic retinopathy or central retinal vein occlusion, 15 patients with other posterior segment disease, seven patients with secondary angle-closure glaucoma, and four patients with aphakia with ruptured anterior hyaloid face. Main outcome measures included visual acuity and intraocular pressure at 1 year postoperatively. RESULTS: At 1 year postoperatively, 31 (77.5%) of 40 patients had stable or improved visual acuity; three eyes (7. 5%) had a final visual acuity of no light perception and three additional eyes (7.5%) were enucleated (because of chronic pain in two eyes and endophthalmitis in one eye). Mean preoperative intraocular pressure was 34 mm Hg and the median number of preoperative antiglaucoma medications was two. At 1 year postoperatively, mean intraocular pressure was 13 mm Hg and the median number of antiglaucoma medications was zero. Twenty-two patients (55.0%) achieved an intraocular pressure greater than 5 mm Hg and less than or equal to 21 mm Hg without antiglaucoma medication, and an additional seven patients (17.5%) achieved this level of intraocular pressure control with medication. Only one patient (2.5%) underwent further glaucoma surgery for uncontrolled intraocular pressure. CONCLUSIONS: Although combined pars plana vitrectomy and placement of a glaucoma drainage implant is often a successful management option in selected patients with refractory glaucoma, visual outcome may be poor because of severe underlying ocular disease and postoperative complications. PMID- 10704550 TI - Ultrasound biomicroscopic study of ciliary body thickness in eyes with narrow angles. AB - PURPOSE: To determine the ciliary body thickness and other biometric findings in eyes with narrow angles. METHODS: Eighteen otherwise normal eyes with narrow angles in 18 Japanese patients and 18 normal control eyes with open angles in 18 age-matched and sex-matched Japanese patients were studied. A-scan ultrasonography was performed to measure anterior chamber depth, lens thickness, axial length, and relative lens position. Ultrasound biomicroscopy was also performed to obtain measurements of the anterior ocular structures, including anterior chamber depth and ciliary body thickness at sites 1 mm and 2 mm posterior to the scleral spur (positions 1 and 2, respectively). RESULTS: Compared with normal control eyes, the narrow-angle eyes showed a shallower anterior chamber (narrow angle, 1.87 +/- 0.27 mm; control, 2.69 +/- 0.26 mm; P <.0001), a thicker lens (4.97 +/- 0.49 mm, 4.26 +/- 0.53 mm; P <.0001), a more anteriorly located lens (2. 21 +/- 0.13, 2.35 +/- 0.14; P <.0001), a shorter axial length (22.70 +/- 0.97 mm, 23.41 +/- 0.86 mm; P =.012), and a thinner ciliary body (position 1: 454 +/- 107 microm, 602 +/- 86 microm; P <.0001; position 2: 203 +/- 50 microm, 321 +/- 68 microm; P <.0001). Lens thickness was significantly correlated with ciliary body thickness at positions 1 (R(2) = 0.34; P =.0001) and 2 (R(2) = 0.43; P <.0001). Anterior chamber depth was significantly correlated with ciliary body thickness at positions 1 (R(2) = 0.48; P <.0001) and 2 (R(2) = 0.56; P <.0001). CONCLUSION: Thinning of the ciliary body may be one of the important factors associated with the anterior location of the lens, the increased lens thickness, and the decreased anterior chamber depth in eyes with a narrow angle. PMID- 10704551 TI - Serum laminin as a marker of diabetic retinopathy development: a 4-year follow-up study. AB - PURPOSE: The usefulness of laminin as a serum marker of diabetic retinopathy is a topic that generates conflicting views. The aim of the present study was to investigate the effect of diabetic retinopathy on serum laminin-P1, the larger pepsin resistant fragment of laminin, and to elucidate whether serum laminin-P1 could be an indicator of the risk for development of diabetic retinopathy. METHODS: In a prospective study, 97 consecutive diabetic patients (35 type 1 and 62 type 2) without diabetic retinopathy and a urinary albumin excretion rate lower than 20 microg per minute were enrolled in a 4-year follow-up study. Patients who developed microalbuminuria during the study were excluded in order to avoid the influence of diabetic nephropathy on serum laminin-P1. At the end of follow-up, data from ophthalmologic studies and serum laminin-P1 were evaluated in the 66 normoalbuminuric diabetic patients who completed the study. RESULTS: No statistical differences were observed in baseline laminin-P1 serum concentrations between patients who developed diabetic retinopathy (n = 15) and patients who remained without it during follow-up (n = 51). However, serum laminin-P1 levels obtained at the end of the study were significantly higher in patients who developed diabetic retinopathy (1.75 +/- 0.33 U/ml versus 1.47 +/- 0. 27 U/ml; P =.002). Furthermore, statistical difference was observed when initial and final values of serum laminin-P1 were compared in patients who developed diabetic retinopathy (1.56 +/- 0.27 U/ml versus 1.75 +/- 0.33 U/ml; P =.001). Remarkably, an increase in serum laminin-P1 concentration was detected in all but two of the patients who developed diabetic retinopathy. The relative risk of development of diabetic retinopathy in patients who showed an increase in serum laminin-P1 during follow-up was 5.4 (95% confidence interval, 1.32 to 22.13). CONCLUSIONS: Serum laminin-P1 is a marker and a risk indicator of diabetic retinopathy but is not an early predictor of its development. PMID- 10704552 TI - Lutetium texaphyrin (Lu-Tex): a potential new agent for ocular fundus angiography and photodynamic therapy. AB - PURPOSE: To investigate the suitability of lutetium texaphyrin (lu-tex) as a fluorescence imaging agent in the delineation of retinal vascular and choroidal vascular diseases. The utilization of an efficient fluorescent molecule that is also a photosensitizer represents a unique opportunity to couple diagnosis and therapy. METHODS: Fundus fluorescence angiography comparing lu-tex (motexafin lutetium, Optrin, Pharmacyclics Inc, Sunnyvale, California) with the conventional angiographic dyes, sodium fluorescein, and indocynanine green (ICG), was performed on the eyes of normal and laser-injured New Zealand white rabbits. Plasma pharmacokinetic data and plasma protein binding were assessed in addition to light microscopy of the retina in both imaged and laser-injured eyes. RESULTS: Normal retinal and choroidal vasculature was well delineated by lu-tex angiography. Experimentally induced choroidal and retinal vascular lesions were enhanced by lu-tex and demonstrated different staining patterns than fluorescein or ICG, particularly at the margins of the lesions. Lu-tex cleared rapidly from the plasma, with 39.7% bound to the high-density lipoprotein (HDL) fraction while 15.8% was bound to the low-density lipoprotein (LDL) fraction. No evidence of retinal toxicity after dye administration was observed by either ophthalmoscopy and fundus photography or by light microscopy. CONCLUSION: Lu-tex angiography is a potentially valuable method for retinal vascular and choroidal vascular evaluation, and it has advantages over fluorescein and ICG angiography. The same agent could conceivably be used for both the identification of abnormal vasculature and subsequent photodynamic treatment. PMID- 10704553 TI - Tissue plasminogen activator to treat impending pupillary block glaucoma in patients with acute fibrinous HLA-B27 positive iridocyclitis. AB - PURPOSE: To report the use of intracameral tissue plasminogen activator to dissolve fibrinous membranes and break posterior synechiae in patients with acute HLA-B27-positive iridocyclitis with impending pupillary block. METHODS: Two patients with severe acute fibrinous iridocyclitis and seclusio pupillae were identified. Because of the concern of impending pupillary block, intracameral tissue plasminogen activator (12.5 microg in 0.1 ml, Activase; Genentech, Inc, South San Francisco, California) was injected with a 25-gauge needle through the corneal limbus. RESULTS: Both patients showed complete dissolution of fibrin with disruption of posterior synechiae. There were no adverse events after injection. Neither patient required further invasive intervention, and both fully recovered with medical management. CONCLUSIONS: Intracameral tissue plasminogen activator is a safe and effective agent for patients with severe acute iridocyclitis and pupillary seclusion. Patients with clinical signs suggestive of impending pupillary block glaucoma may be considered for tissue plasminogen activator injection to avoid the possible need for emergency surgical iridectomy and synechiolysis. PMID- 10704554 TI - Aspirin use and the prevention of acute ischemic cranial nerve palsy. AB - PURPOSE: To assess the relationship of aspirin use and ischemic cranial nerve palsies among patients with diabetes mellitus and hypertension. METHODS: This retrospective case-control study involved 100 patients with ischemic cranial nerve palsies in association with diabetes, hypertension, or both (palsy cases) and 163 age-matched and sex-matched patients with diabetes, hypertension, or both but without ischemic cranial nerve palsies (nonpalsy control subjects). Comparisons were made with respect to duration of diabetes, dose and duration of aspirin use, dose and duration of tobacco use, and presence of cardiac or cerebrovascular disease. RESULTS: There were 20 oculomotor, 33 trochlear, 37 abducens, and 10 facial nerve palsy cases. The median duration of diabetes was 6 years for cases and 7 years for control subjects. There were 34 cases (34%) who had used aspirin for a mean duration of 5.5 years before the onset of the cranial nerve palsy and 49 control subjects (30.1%) who had used aspirin for a mean duration of 4.3 years. There were no significant differences between cases and control subjects for duration of diabetes (P =.94); aspirin use (P =.51), duration (P =.50), and dosage (P =.89); tobacco use (P =.73) and consumption (P =.45); and proportion of cardiac disease (P =.17). Cerebrovascular disease was significantly less common among palsy cases than nonpalsy control subjects (P<.001). There was no significant difference in the odds of a patient having cranial nerve palsy in the aspirin group compared with the nonaspirin group (odds ratio, 1.12; 95% confidence interval, 0.70-2.04). CONCLUSION: Aspirin use was not associated with a reduced rate of ischemic third, fourth, sixth, and seventh nerve palsies among patients with diabetes mellitus and hypertension. Aspirin appears to be ineffective in preventing ischemic third, fourth, sixth, and seventh cranial nerve palsies. Patients with ischemic cranial nerve palsy have a significantly lower rate of strokes and transient ischemic attacks than patients who have diabetes or hypertension but who do not have a history of cranial nerve palsy. PMID- 10704555 TI - Epstein-Barr virus dacryoadenitis. AB - PURPOSE: To describe the clinical features of lacrimal gland inflammation associated with Epstein-Barr virus infection. METHODS: The clinical records, laboratory data, and radiographs of patients who had inflammation of one or both lacrimal glands that had begun less than 4 weeks previously were reviewed. RESULTS: Sixteen patients with dacryoadenitis were encountered between 1980 and 1996, a cumulative frequency of approximately one case per 10,000 new ophthalmic outpatients. Six individuals had serologic or other evidence of recent Epstein Barr virus infection and were distinguished by the presence of regional lymphadenopathy, no purulent discharge, and a duration of symptoms of 6 weeks. CONCLUSION: Epstein-Barr virus is a probable cause of unilateral and bilateral dacryoadenitis in young adults. PMID- 10704556 TI - Fine lattice lines on the corneal surface after laser in situ keratomileusis (LASIK). AB - PURPOSE: To present an example of a pattern of lines resembling fine lattice on the corneal surface subsequent to laser in situ keratomileusis (LASIK). This subtle phenomenon may be relatively common and may affect visual outcome. METHOD: Case report. RESULTS: A 41-year-old year old man with high myopia and best corrected visual acuity of 20/20 +2 in each eye underwent laser in situ keratomileusis (LASIK). No operative or postoperative complications occurred. No striae were evident on slit-lamp examination with direct illumination and retroillumination at the time of surgery or in the postoperative period. Postoperative uncorrected visual acuity was 20/25 with a best-corrected spectacle correction of 20/25 in both eyes. Fine lines in a lattice pattern were seen only with fluorescein dye in the precorneal tear film as areas of "negative stain" within the LASIK flap. With tear film supplementation, the lines were less evident and visual acuity improved. One year postoperatively, his uncorrected visual acuity was 20/25 in both eyes. The best-corrected spectacle visual acuity was RE: 20/20 -2, LE: 20/25. The fine lines were still present within the flap. A soft contact lens improved visual acuity to 20/20 in both eyes. Although all four puncta were occluded, he had no epiphora. CONCLUSION: Fine lines in a lattice pattern that may represent folds in the epithelium or Bowman layer may be present within the flap after LASIK and may adversely affect visual acuity. They may be visible as areas of negative stain with fluorescein dye in the precorneal tear film in the absence of any striae visible in the flap. These superficial lines have been seen more in patients with high degrees of correction and in patients with dry eye. If visual acuity is affected, it may be improved with punctal occlusion, tear supplements, or a contact lens. PMID- 10704557 TI - Diffuse interface keratitis after laser in situ keratomileusis (LASIK): a nonspecific syndrome. AB - PURPOSE: To characterize the presentation of diffuse interface keratitis after laser in-situ keratomileusis (LASIK). METHODS: Case report. RESULTS: Diffuse interface keratitis occurred in the left eye of a postoperative LASIK patient after central epithelial debridement without exposure of the flap margin or elevation of the flap. CONCLUSION: Diffuse interface keratitis is a nonspecific presentation of corneal inflammation after LASIK, with accumulation of inflammatory cells in the potential space of the interface. Diffuse interface keratitis after LASIK may have multiple causes. PMID- 10704558 TI - Mycobacterium chelonae keratitis after laser in situ keratomileusis successfully treated with medical therapy and flap removal. AB - PURPOSE: To report a case of Mycobacterium chelonae keratitis after laser in situ keratomileusis successfully treated with medical therapy and flap removal. METHODS: Case report. A 36-year-old white woman in good health developed a paracentral keratitis in her right eye 1 month after bilateral laser in situ keratomileusis. Initial treatment included topical steroids and then intensive Ocuflox (ofloxacin ophthalmic solution; Allergan, Inc, Irvine, California) without success. Cultures were negative. The keratitis worsened, and she was referred to our institution. Interface infiltration was noted, and the flap was lifted to obtain adequate laboratory studies. Cultures were positive for M chelonae. RESULTS: The keratitis was treated with intensive topical amikacin sulfate 1%, topical clarithromycin 1%, and Ciloxan (ciprofloxacin HCL; Alcon Laboratories, Inc, Fort Worth, Texas) with minimal improvement in her clinical condition. She developed a toxic reaction to amikacin 1%. In order to improve antibiotic penetration, the hazy, ulcerated corneal flap was removed. The keratitis then resolved with intensive topical clarithromycin 1% and Ocuflox over 5 weeks. The patient now has visual acuity without correction of 20/50, despite superficial corneal haze. CONCLUSION: M chelonae is a rare and insidious cause of infection after laser in situ keratomileusis. Diagnosis can be difficult and is often delayed. Aggressive medical management, with flap removal, if needed, may lead to resolution of infection. PMID- 10704559 TI - Caterpillar setae in the deep cornea and anterior chamber. AB - PURPOSE: To report a case of caterpillar setae embedded in the deep cornea and anterior chamber. METHODS: A 26-year-old man was struck in his right eye by a caterpillar (Dendrolimus punctatus walker). Severe conjunctival injection, chemosis, and erosion of the cornea developed immediately. Numerous setae fragments were found to be embedded into the palpebral conjunctiva and deep cornea, extending into the anterior chamber near the anterior iris surface. RESULTS: After partial removal of the setae under a microscope, the inflammation subsided and visual acuity improved to RE: 20/20. CONCLUSION: Caterpillar setae are sharp enough to penetrate the cornea and extend into the anterior chamber. PMID- 10704560 TI - A new surgical technique for management of conjunctivochalasis. AB - PURPOSE: To present a new surgical technique for severe, symptomatic conjunctivochalasis and our hypothesis of the pathogenesis of this condition. METHODS: Six eyes of three patients with conjunctivochalasis (average age +/-SD, 70.0 +/- 9.6 years; range, 56-78 years) were treated with a conjunctival fixation to sclera with three 6-0 Vicryl (Johnson & Johnson, New Brunswick, New Jersey) stitches. RESULTS: With a mean follow-up period of 209.5 days (range, 181-219 days), we achieved successful treatment in all eyes, with no recurrence of conjunctival folds. CONCLUSION: We successfully treated conjunctivochalasis with conjunctival fixation to sclera, which strongly suggests that conjuctival folds are caused by the folding and the elevating of loosely adherent bulbar conjunctiva of the lower eyelid. PMID- 10704561 TI - Pars plana vitrectomy during cataract surgery for prevention of aqueous misdirection in high-risk fellow eyes. AB - PURPOSE: To report the use of pars plana vitrectomy as a prophylactic measure during cataract surgery for prevention of aqueous misdirection in high-risk fellow eyes. METHODS: Chart review of two patients with severe aqueous misdirection in their first eye at the time of cataract surgery that only responded to pars plana vitrectomy. RESULTS: In both patients, cataract extraction with posterior chamber intraocular lens implantation was initially performed after pars plana vitrectomy, with the creation of hyaloido-capsulo iridotomy to establish a communication between the vitreous cavity and the anterior chamber. CONCLUSION: Pars plana vitrectomy as a prophylactic measure during cataract surgery may have a beneficial role in fellow eyes at high risk for developing aqueous misdirection. PMID- 10704562 TI - Postoperative endophthalmitis caused by sequestered Acinetobacter calcoaceticus. AB - PURPOSE: To describe postoperative endophthalmitis caused by sequestered Acinetobacter calcoaceticus. METHOD: Case report. A 40-year-old woman developed recurrence of inflammation after extracapsular cataract extraction with intraocular lens (IOL) implantation. At last recurrence, the capsular bag was studded with white deposits. Intraocular lens was removed along with capsular bag during pars plana vitrectomy. RESULTS: The capsular bag, when cultured, grew A calcoaceticus. The media remained clear with no evidence of recurrence of infection over a 3-month follow-up. CONCLUSION: Postoperative endophthalmitis similar to that caused by sequestered Propionibacterium acnes can be caused by A calcoaceticus. PMID- 10704563 TI - Intralenticular Candida species abscess in a premature infant. AB - PURPOSE: To report the clinical and histopathologic findings of a premature infant with severe retinopathy of prematurity complicated by the development of an intralenticular fungal abscess. METHODS: Case report and literature review. RESULTS: A markedly premature infant developed Candida septicemia at 29 weeks postconception. Over the ensuing 10 weeks, cataract and intraocular inflammation developed sequentially in each eye, as did progressive retinopathy of prematurity with tractional retinal detachment. Pars plana vitrectomy and lensectomy revealed intralenticular Candida species abscess. CONCLUSION: Progressive cataract and intraocular inflammation in a low birth weight infant may be caused by endogenous intraocular infection secondary to systemic candidiasis. Cataract secondary to retinopathy of prematurity is rare. PMID- 10704564 TI - Laser iridocystotomy for bilateral acute angle-closure glaucoma secondary to iris cysts. AB - PURPOSE: To report laser iridocystotomy for bilateral acute angle-closure glaucoma secondary to peripheral iris cysts. METHOD: Case report. RESULTS: In a 55-year-old man with increased bilateral intraocular pressure, gonioscopy revealed varied angle narrowing. Bilateral angle-closure glaucoma secondary to peripheral iris cysts was diagnosed by ultrasound biomicroscopy. The peripheral iris cysts could not be seen in mydriasis by gonioscopy. Therefore, we decided to perform laser iridocystotomy with argon and Nd:YAG laser. Collapse of the cysts after laser treatment was demonstrated by ultrasound biomicroscopy. At follow-up, 9 months after laser treatment, intraocular pressure had dropped below 20 mm Hg in both eyes without further therapy. The iris cysts did not recur, which was demonstrated by ultrasound biomicroscopy. CONCLUSIONS: Peripheral iris cysts may produce angle closure and may cause secondary angle-closure glaucoma. If transpupillary laser cystotomy is not possible, laser iridocystotomy may produce collapse of the iris cysts and correction of secondary angle closure. PMID- 10704565 TI - Tissue plasminogen activator and gas for diabetic premacular hemorrhage. AB - PURPOSE: To report the effect of intravitreal injection of tissue plasminogen activator and gas for the treatment of diabetic premacular hemorrhage. METHODS: Five eyes of five consecutive patients with fresh diabetic premacular hemorrhage and previous panretinal photocoagulation were treated with intravitreal injection of 50 microg of tissue plasminogen activator, and 0.3 ml of sulfur hexafluoride. After intravitreal injection, patients maintained a prone position for 3 days and were followed for an average of 7 months (range, 6 to 8 months). RESULTS: All eyes had clearing of the premacular hemorrhage with improved visual acuity. Three eyes with no visible fibrovascular proliferation required 3 weeks to resorb the hemorrhage, and two eyes with active fibrovascular proliferation needed 2 months to resorb the intravitreal hemorrhage. CONCLUSION: Intravitreal injection of tissue plasminogen activator and gas may be an effective way to treat fresh diabetic premacular hemorrhage. PMID- 10704566 TI - Rearrangement of immunoglobulin gene in metastatic Waldenstrom macroglobulinemia to the vitreous. AB - PURPOSE: To report metastatic Waldenstrom macroglobulinemia cells with immunoglobulin heavy chain gene rearrangement in the vitreous and the blood. METHODS: A 58-year-old man with Waldenstrom macroglobulinemia developed bilateral vitreitis. Diagnostic vitrectomy was performed on the left eye. The vitreous cells and the peripheral blood lymphocytes were analyzed using microdissection and polymerase chain reaction amplification. RESULTS: Vitrectomy specimen of the left eye contained a few degenerated cells. Molecular analysis showed immunoglobulin heavy chain gene rearrangement at the third complementary determining region of the vitreal infiltrating cells and peripheral blood lymphocytes. CONCLUSIONS: Waldenstrom macroglobulinemia rarely metastasizes to the vitreous. Molecular detection of the immunoglobulin heavy chain gene third complementary determining region rearrangement is helpful in the diagnosis of the malignancy. Microdissection combined with polymerase chain reaction is a useful and innovative tool for molecular pathological investigation. PMID- 10704567 TI - Siblings of retinoblastoma patients: are we underestimating their risk? AB - PURPOSE: To describe the clinical presentation of probable germ-line mosaicism in four retinoblastoma kindreds. METHODS: Review of 255 retinoblastoma patients and their family records in a University of California, San Francisco-Bascom Palmer database to identify those with potential germ-line mosaicism. Parents and siblings of retinoblastoma patients were given comprehensive ophthalmologic examinations. RESULTS: Four kindreds were identified, wherein retinoblastoma was diagnosed in two siblings and both parents demonstrated no evidence of retinoblastoma or retinocytoma. CONCLUSION: Clinical appearance of germ-line mosaicism is demonstrated in our retinoblastoma patient populations. We recommend routine clinical screening of all parents and siblings of retinoblastoma patients to provide more accurate genetic counseling and to allow earlier examination and treatment of children at presymptomatic disease stages. Germ-line mosaicism must be considered as a genetic transmission pattern in these patients, and genetic counseling should specifically recognize this possibility. If a parent is germ line mosaic, the possibility of bearing a second child with retinoblastoma is clearly higher than conventionally believed. PMID- 10704568 TI - Cox-2 expression in retinoblastoma. AB - PURPOSE: Cox-2, a prostaglandin synthase, is overexpressed in colorectal cancers and is involved in angiogenesis as well as in tumorigenesis. In this study, we investigate the expression of Cox-2 in retinoblastoma. METHODS: Twenty-nine formalin-fixed retinoblastoma specimens were examined by the labeled-streptavidin biotin method using anti-Cox-2 antibody. RESULTS: Cox-2 positive immunoreactions were observed in 28 (96%) of 29 retinoblastomas specimens. CONCLUSION: This preliminary study suggests the overexpression of Cox-2 in both differentiated and undifferentiated retinoblastoma and its possible role in tumorigenesis. PMID- 10704569 TI - Bilateral juxtafoveolar telangiectasis in monozygotic twins. AB - PURPOSE: To report the clinical and angiographic features of two monozygotic twins affected by bilateral group 2 idiopathic juxtafoveolar telangiectasis. METHOD: Case reports. RESULTS: Two 64-year-old women, who were identical twins, were suffering from visual loss. One twin had suffered from visual loss for 1 year and had a visual acuity of 20/25 in both eyes, and the other twin had suffered for 2 years and had a visual acuity of 20/30 in both eyes. Fluorescein angiography disclosed similar fundus features. An analogous area of capillary telangiectasis and leakage was observed in the right macula, where no intraretinal pigment was seen; the left macula showed a similar amount of intraretinal pigment associated with tiny right-angle venules and minimal leakage. CONCLUSION: This observation raises the issue of genetic influences in the pathogenesis of this disease. PMID- 10704570 TI - High prevalence of herpes simplex virus type 2 in acute retinal necrosis syndrome associated with herpes simplex virus in Japan. AB - PURPOSE: To determine the type of herpes simplex virus in acute retinal necrosis syndrome associated with herpes simplex virus. METHODS: Herpes simplex virus type 1, herpes simplex virus type 2, varicella-zoster virus, Epstein-Barr virus, and cytomegalovirus were examined by polymerase chain reaction in intraocular specimens from 16 patients with acute retinal necrosis syndrome. Anti-herpes simplex virus type 1 and anti-herpes simplex virus type 2 type-specific antibodies in serum from the patients were detected by enzyme immunoassay. RESULTS: Of 16 patients with acute retinal necrosis syndrome, seven were polymerase chain reaction positive for herpes simplex virus type 2 and nine were positive for varicella-zoster virus. Anti-herpes simplex virus type 2 antibody was positive and anti-herpes simplex virus type 1 antibody was negative in the sera of the seven patients with herpes simplex virus type 2 DNA-positive acute retinal necrosis syndrome. In contrast, anti-herpes simplex virus type 2 antibody was absent in all nine varicella-zoster virus DNA-positive acute retinal necrosis syndrome patients. CONCLUSION: Herpes simplex virus type 2 has been demonstrated to be the major causative agent in acute retinal necrosis syndrome associated with herpes simplex virus by molecular biological and serological methods. Negative preexisting anti-herpes simplex virus type 1 antibody may play an important role in acute retinal necrosis syndrome associated with herpes simplex virus type 2. PMID- 10704571 TI - Pseudopapilledema in neurofibromatosis type 2. AB - PURPOSE: To report a case of neurofibromatosis type 2 with pseudopapilledema secondary to a prepapillary gliotic membrane. METHOD: Case report. Results of an ocular examination and fluorescein angiography of a patient are described. RESULTS: Fundus examination of a 14-year-old male with neurofibromatosis type 2 revealed an irregular elevation of the optic nerve and a perifoveal epiretinal membrane in the right eye. Fluorescein angiography demonstrated no autofluorescence nor leakage in the area of the optic nerve. CONCLUSION: The patient has pseudopapilledema secondary to an epiretinal membrane overlying the optic disk of the right eye. The possibility of pseudopapilledema should be considered when evaluating patients with neurofibromatosis type 2 and abnormal optic nerves. PMID- 10704572 TI - Estrogen and visual hallucinations in a patient with Charles Bonnet syndrome. AB - PURPOSE: To describe the occurrence of visual hallucinations in a patient with Charles Bonnet syndrome associated with estrogen intake. METHOD: Case report. RESULTS: An 84-year-old woman with poor visual acuity secondary to bilateral, nonexudative, age-related macular degeneration had nonthreatening visual hallucinations 2 weeks after starting oral estrogen for osteoporosis. The estrogen was stopped, and the hallucinations subsided. The patient was given estrogen twice more and each time the hallucinations recurred. CONCLUSION: We report a case of Charles Bonnet syndrome associated with estrogen intake in an 84 year-old woman. Estrogen may have promoted release phenomena and triggered the hallucinatory episodes in our patient. PMID- 10704574 TI - Correction PMID- 10704573 TI - Bilateral sixth nerve palsy associated with MDMA ("ecstasy") abuse. AB - PURPOSE: To report the association of methylenedioxymetamphetamine (MDMA, "ecstasy") abuse and bilateral sixth nerve palsy. METHODS: Case report. RESULTS: A 17-year-old male presented with horizontal diplopia in all directions of gaze after having taken MDMA tablets at 5-day to 7-day intervals during a 2-month period. Examination showed bilateral sixth nerve palsy. Ocular motility returned to normal within 5 days without use of MDMA and with no other treatment. CONCLUSION: Methylenedioxymetamphetamine "ecstasy" abuse should be considered in the differential diagnosis in otherwise unexplained sixth nerve palsy. PMID- 10704575 TI - Epiphyseal fractures of the proximal tibia. AB - Fractures of the proximal tibial epiphysis and apophysis are rare. Data of ten patients seen in the last 17 yr with an epiphyseal (6) or apophyseal (4) fracture of the proximal tibia were reviewed. Three patients with an epiphyseal fracture were treated by open reduction and fixation, the other three by closed reduction. All apophyseal fractures were treated by open reduction and internal fixation. No major complications occurred. The final results were good. PMID- 10704576 TI - Femoral head measurement in hemiarthroplasty: assessment of interobserver error using 3 measuring systems. AB - We have assessed the consistency of measurement of femoral head circumference using 3 standard measuring instruments used in hemiarthroplasty of the hip. Fifty femoral heads were independently sized by ten independent observers using a caliper, a half circular measuring template, each allowing measurement to the nearest millimeter. We found significantly greater variance of the results using the half-circular templates (p=0.001) and the calipers (p=0. 011) compared with the full circular measuring templates. Measurement with calipers underestimated femoral head size by 0.72 mm compared with full circular measuring templates (p=0.02). Insertion of an undersized hemiarthroplasty head is a cause of increased point loading of the acetabulum and may result in increased rates of acetabular erosion. The use of full circular measurement templates is recommended as the most consistent method for head sizing in hemiarthroplasty of the hip. PMID- 10704577 TI - Treatment of isolated complex distal femoral fractures by external fixation. AB - Thirteen patients with isolated distal femoral fractures were treated by external fixation. There were seven males and six females with an average age of 45 years. Four were Type A3 fractures, one Type C1, five Type C2 and three Type C3 fractures. Seven of these were open. In seven cases the articular surface was first reduced and fixed. The fixation was extended across the knee to supplement the distal fixation in six severe cases. The average follow up was 30 months. There was one non-union in the study with the average time to union in the other patients being six months. Using the Mize criteria for assessing clinical results we found that nine patients obtained a good to excellent score and four were classed as failures. The average range of movement of the knee in the study was 100 degrees. Apart from the single non-union all the fractures healed and there were no other serious complications. Considering the severity of the fractures we did not find any evidence to suggest that temporary fixation of the lateral soft tissues by fixator pins was detrimental. The results suggest that external fixation may be used to treat these difficult fractures without the risk of serious complications. PMID- 10704578 TI - Ipsilateral hip and distal femoral fractures. AB - We tried to find the trauma mechanism and treatment rationale of ipsilateral concomitant hip and distal femoral fractures involving the articular surface. Between 1988 and 1995, 15 cases of ipsilateral hip (confined to neck or trochanteric areas of the femur) and distal (confined to supra- and intercondylar area of the femur) femoral articular fractures were collected. The hip fractures consisted of 10 trochanteric fractures and five neck fractures, which were managed with reduction and fixation in 14 (Knowles' pin in eight, DHS in four and standard Gamma nail in two), and primary bipolar hemiarthroplastry in one. The distal femoral articular fractures were open in 11; these were managed with radical debridement, implantation of Septopal chains and immediate internal fixation, followed by prophylactic autogenous bone grafting 6 weeks later in the recent six cases (five Judet plates, four dynamic condylar screws and two condylar plates). The other four closed distal femoral fractures were managed with early reduction and internal fixation (two Judet plate, one dynamic condylar screw and one condylar plate). The union time was 20.3 (12-48) weeks for proximal fractures and 23.7 (12-36) weeks for distal fractures. Early infection developed in three cases. Nonunion of a femoral neck fracture developed in one case. The other complications were implant failure in one, coxa vara in one, refracture in one, delayed union in one and knee stiffness in one. PMID- 10704579 TI - Treatment of extensive bone and soft tissue defects of the lower limb by traction and free-flap transfer. AB - Twenty patients with extensive bone and soft tissue defects and posttraumatic osteomyelitis were treated between 1983 and 1995. In all cases an external fixator was used for bone fixation. Bone defects were managed with the Ilizarov intercalary bone transport. Two types of traction were used: the Ilizarov type and a 'new' Ljubljana type. The results of treatment were compared between the two types of traction. In all cases delayed bony union was observed. Osteomyelitis never reactivated. All patients were satisfied with treatment. They were all independent except for one amputee. The Ljubljana traction method was found to have the following advantages: no discrepancy in leg length, no orthopaedic support was needed, the aesthetic outcome was better, the traction time was reduced and there was less soft tissue damage during bone traction. PMID- 10704580 TI - Evaluation of the role of pin fixation versus collar and cuff immobilisation in supracondylar fractures of the humerus in children. AB - Long term results of children with supracondylar humeral fractures treated with manipulation and strapping and manipulation followed by pin fixation were evaluated. Forty patients were regarded as Gartland type II injuries. 33 of these were treated with closed reduction and collar and cuff immobilisation and 7 with closed reduction and percutaneous pinning. Two cases of cubitus varus were reported one from each treatment modality. Forty-four patients were included as Gartland type III injuries. Of these 14 were treated with closed reduction and collar and cuff immobilisation, 25 with closed reduction and percutaneous pinning and five with open reduction and pinning. There were two cases of cubitus varus and one case of cubitus valgus following pin fixation. In addition one case of extension lag and one significant ulnar nerve neurapraxia was recorded following pin fixation. One case of cubitus varus was seen following manipulation and collar and cuff treatment. There was no statistical difference between either treatment modality in terms of predicting a better outcome (p0.05). We conclude that pin fixation has no advantages over simple immobilisation in certain Gartland II and III type injuries. Although pin fixation is beneficial in unstable injuries collar and cuff immobilisation continues to have an important role in the treatment of stable supracondylar fractures. PMID- 10704581 TI - Management of distal femoral fractures in elderly patients using retrograde titanium supracondylar nails. AB - We present our experience with a retrograde supracondylar nail used for the management of fractures of the distal femur in elderly patients. Eighteen fractures of the distal femur in 18 patients were treated with AIM titanium supracondylar nails. Sixteen patients with a median age of 83 years (62-100 years) were finally available for review. All 16 fractures were classified as extra-articular type A according to the AO classification. The average operative time was 58 min. Follow up ranged between 4 and 35 months (average 20.4 months). Fifteen fractures (93.7%) united in an average duration of 3.6 months. The average range of motion achieved at the knee was 100. 6 degrees. There were no implant failures, knee sepsis or wound healing problems. One non-union and two stress fractures of the femur above the nail were the main complications in this series. We concluded that the AIM titanium supracondylar nail is a useful alternative implant for the management of the osteoporotic fractures of the distal femur particularly the extra-articular AO type A fracture in the elderly population. PMID- 10704582 TI - The treatment of lateral clavicle fractures. AB - This study assesses the results of surgical treatment of 15 displaced Neer type II fractures of the lateral clavicle in 15 patients, which occurred between November 1988 and March 1995 and which were followed up for a mean period of 4.6 years (range 2-9 years). The patients fell into two groups, one 'acute group' and one 'non-union' group. Patients treated initially by a non-operative approach had suffered prolonged morbidity and time off work prior to and after surgery. The ultimate result was good. The fixation used was a Dacron arterial graft as a sling around the clavicle and coracoid process. Delayed (non-union) cases were augmented with bone graft and inter-fragmentary screw fixation. All fractures eventually united. We question the place of prolonged non-operative management in the treatment of displaced Neer type II fractures of the lateral clavicle. PMID- 10704583 TI - Surgical treatment of displaced acetabular fractures - 72 cases followed for 10 (6-14) years. AB - Seventy-two displaced acetabular fractures managed surgically were evaluated retrospectively. The follow-up period was 10 (6-14) yr. The commonest fractures were posterior wall (28) and both columns (10). The surgical approaches were Kocher-Langenbeck (47), ilioinguinal (19) and extended iliofemoral (6). No neural monitoring was used in operations and no preventive agents for heterotopic ossification or thromboembolism were used perioperatively. Reduction was rated congruent in 59 (81.9%) and noncongruent in 13 (18.1%). The early postoperative complications were 1 vascular injury, 1 iatrogenic sciatic nerve injury, 1 deep vein thrombosis and 2 wound infections. The late complications were heterotopic ossification in 20 patients, avascular necrosis of the femoral head in 4 and symptomatic arthritis in 10. Functional outcomes were rated as excellent in 31, good in 23, fair in 7 and poor in 11. Our results show that traditional management is effective enough for displaced acetabular fractures. PMID- 10704584 TI - The use of interlocked 'customised' blade plates in the treatment of metaphyseal fractures in patients with poor bone stock. AB - Balanced and stable fixation in metaphyseal fractures and nonunion can be difficult because of osteoporosis, disuse osteopenia, comminution, joint proximity or malignant infiltration. Five patients with nonunion and four with comminuted or pathological acute fractures of metaphyseal areas of the tibia or humerus were treated with a 'customized' interlocked blade plate. The plates are standard AO dynamic compression plates of a suitable length that are bent to an acute angle in an industrial vice and 'interlocked'. The nine patients with a mean age of 62.2 years (range 30-91) were followed up for a mean of 7.2 months. At follow-up all fractures had healed with a single complication of subacromial impingement in a patient with a proximal humeral fracture. PMID- 10704585 TI - Primary surgical treatment of war injuries of the foot. AB - Presented are the results of primary surgical treatment of war injuries of the foot in 250 patients wounded in the territory of former Yugoslavia in the period June 1991-October 1995. Total number of feet treated was 275. Aetiologically, the most frequent injuries were those inflicted by the effect of mine blasts (65.2%) and by bullets from firearms (30.8%). Injuries to a single foot were present in 140 (56.0%), combined injuries in 107 (42.8%) and associated with burn injuries in three (1.2%) patients. Soft tissue injuries were present in 25 (9.1%) and injuries to bone structures and joints in 250 (90.9%) feet. Injuries resulting from solid blasts were found in nine (3.2%) feet. Postoperative bone fragment stabilization was required in 115 (41.8%) feet. Stabilization was by plaster of Paris in 87 (75.6%) and by external fixation in 23 (20. 0%) feet. Amputations were performed in 73 (26.5%) feet. Covering of soft tissue and bone defects was required in 84 (33.6%) patients. PMID- 10704586 TI - Nonoperative management of penetrating liver trauma. PMID- 10704587 TI - Massive haemothorax following thoracic vertebral fracture. PMID- 10704588 TI - Cannulated screw fixation of fractured capitellum: surgical technique through a limited approach. PMID- 10704589 TI - Implanted piece of allogeneic femoral bone and late sciatic nerve compression. PMID- 10704590 TI - Attempted nail-gun suicide: fluid management in penetrating cardiac injury. PMID- 10704591 TI - Acute anterior compartment syndrome of the forearm following lifting a heavy load. PMID- 10704592 TI - Overcoming codon bias: a method for high-level overexpression of Plasmodium and other AT-rich parasite genes in Escherichia coli. AB - Parasite genes often use codons which are rarely used in the highly expressed genes of Escherichia coli, possibly resulting in translational stalling and lower yields of recombinant protein. We have constructed the "RIG" plasmid to overcome the potential codon-bias problem seen in Plasmodium genes. RIG contains the genes that encode three tRNAs (Arg, Ile, Gly), which recognise rare codons found in parasite genes. When co-transformed into E. coli along with expression plasmids containing parasite genes, RIG can greatly increase levels of overexpressed protein. Codon frequency analysis suggests that RIG may be applied to a variety of protozoan and helminth genes. PMID- 10704593 TI - Molecular characterisation of the actin gene of the filarial parasite Wuchereria bancrofti. AB - Wuchereria bancrofti is the major cause of lymphatic filariasis in humans. Although it is responsible for this immensely morbid and debilitating disease, very little is known of the basic molecular biology of this parasite, and there is a vast lack of knowledge on its gene organisation. In this study, the actin gene of W. bancrofti has been characterised by sequencing a clone isolated from a genomic DNA library of this parasite. The 5' flanking region had a potential TATA box and a putative mRNA initiation site. The gene had five exons encoding 376 amino acids, and four introns ranging in size from 109 to 190bp. The 3' flanking region had a potential polyadenylation signal with the sequence ATTAAA which is a common natural variant of the conventional sequence AATAAA. The gene was AT-rich, with a GC content of 37.2%. Southern blot analysis of W. bancrofti genomic DNA indicated that the gene is possibly found as a single copy. The actin amino acid sequence of W. bancrofti showed a high degree of homology to the actin of many organisms of different taxonomic groups, but the highest homology was observed with the free-living nematode Plectus acuminatus. This suggests that P. acuminatus may bear a close evolutionary relationship to W. bancrofti. PMID- 10704594 TI - Cloning and characterisation of a peroxiredoxin from the swine roundworm Ascaris suum. AB - Antioxidant enzymes in parasites play an important role in protection against the oxygen radicals by generating during aerobic metabolism, as well as in defence against host immune cell assault. Here we report the cloning and characterisation of a cDNA encoding peroxiredoxin from Ascaris suum (AsPrx). AsPrx is 776bp long and contains the nematode 22bp splice leader sequence at the 5' end and polyadenylation signal followed by poly(A) tail at the 3' end. AsPrx codes a full length protein with a predicted molecular mass of 22. 6kDa, and possesses two cysteine residues at amino acid 49 and 168 that are conserved among Prx proteins. GenBank() analysis showed that the deduced amino acid sequence had significant similarity to parasite and mammalian Prx at the amino acid level. DNA nicking revealed that Escherichia coli-expressed recombinant AsPrx (rAsPrx) is enzymatically inhibited to form oxidative-nicking of supercoiled plasmid DNA. Two dimensional immunoblot analysis with mouse anti-rAsPrx serum reacted two major constituent protein spots in extracts of adult female worms, suggesting that the native AsPrx might be function as a major antioxidant enzyme in Ascaris suum. PMID- 10704595 TI - Immune and pathophysiological responses to different strains of Giardia duodenalis in neonatal mice. AB - Numerous studies have demonstrated various strain differences between Giardia isolates, but little is known about the immunology and pathogenesis of infections. This study aimed to compare host responses to strains of Giardi duodenalis differing in levels of virulence and pathogenicity and, by doing so, elucidate the mechanisms via which pathogenic strains establish infections. Marked differences were found in the infection dynamics, histopathological responses and serum antibody responses of neonatal mice infected with either G. duodenalis strain BRIS/83/HEPU/106 (isolated from a human) or BRIS/95/HEPU/2041 (isolated from a sulphur-crested cockatoo, Cacatua galerita). Infections with the bird strain were more intense (6.7-times greater) and persisted longer (by 14days) than infections with the human strain. The bird strain was more pathogenic and caused greater pathophysiological alteration to the gut mucosa, including increased villous atrophy, hyperplasia of goblet cells and vacuolated epithelial cells. Mice infected with the bird strain produced less serum anti Giardia IgA and IgM, but more total (non-specific) serum IgA than those infected with the human strain of Giardia. This suggests that avian G. duodenalis strains are infective for mammalian hosts and may contribute to zoonotic infections. Furthermore, infection of mice with BRIS/95/HEPU/2041 serves as a good experimental model to provide further insight into the mechanisms via which G. duodenalis causes disease. PMID- 10704596 TI - PCR assay for the specific amplification of Oesophagostomum bifurcum DNA from human faeces. AB - Oesophagostomiasis in humans due to infection with Oesophagostomum bifurcum (nodule worm) is of major human health significance in northern Togo and Ghana where the human hookworm, Necator americanus, also exists at high prevalence. Accurate diagnosis of O. bifurcum infection in humans is central to studying the epidemiology and controlling the parasite. To overcome limitations of current copro-diagnostic methods, we have developed an alternative, molecular approach. Utilising genetic markers in the second internal transcribed spacer (ITS-2) of ribosomal DNA, we have established a two-step, semi-nested PCR method for the specific amplification of minute amounts (fg) of O. bifurcum DNA from human faecal samples. Using a panel of 155 well-defined faecal and DNA samples, the assay achieved a sensitivity of 94.6% and a specificity of 100%. This PCR assay will be useful for the diagnosis of O. bifurcum infection and as a molecular tool for elucidating the epidemiology of human oesophagostomiasis. PMID- 10704597 TI - Histopathological investigation of experimental ocular toxocariasis in gerbils. AB - Male gerbils were inoculated intragastrically with embryonated Toxocara canis eggs. They were euthanised, and their eyes were excised at different days p.i. to identify the number of larvae as well as lesions resulting in these organs. In most animals, larvae were detected from day 5 to day 60 p.i. (end of the study). From days 10 to 20 p.i., larvae and haemorrhage were observed in the choroid and in the ciliary process. At days 30 and 40 p.i., some eyes had larvae surrounded by an infiltrate of neutrophils, oedema, haemorrhages and tissue damage in the retina or the ciliary process. On day 60 p.i., there were granulomatous lesions in the retina. This study provides a model for the study of ocular toxocariasis. PMID- 10704598 TI - Measurement of the efficacy of vaccines and antimicrobial therapy against infection with Toxoplasma gondii. AB - To facilitate studies of vaccines and antimicrobial agents effective against Toxoplasma gondii infection, an assay system was developed to semi-quantitate parasitaemia using PCR amplification of T. gondii DNA obtained from the blood of mice infected with the parasite. A competitive internal standard DNA fragment of the B1 gene of T. gondii was generated and used in PCR so that the amplified product could be semi-quantitated and false negative results could be avoided. The PCR assay system was used to analyse the levels of parasitaemia in immunised and antimicrobial agent treated mice at various times after infection with T. gondii. The results of these studies indicate that this highly sensitive detection method is a rapid and reliable procedure that can be employed to assess the abilities of vaccines or antimicrobial agents to provide protection early following T. gondii infection. PMID- 10704599 TI - ITS-1 ribosomal DNA sequence variants are maintained in different species and strains of Echinococcus. AB - This study investigated sequence heterogeneity in the first internal transcribed spacer (ITS-1) of ribosomal DNA within and among species and strains of Echinococcus. Different ITS-1 sequence variants exist in Echinococcus granulosus and Echinococcus multilocularis, which represent at least four evolutionary lineages: (1) a sheep strain-lineage of E. granulosus, (2) a sister lineage of a cervid and camel E. granulosus ITS-1 variants, (3) a lineage including the ITS-1 variants representing horse, bovine and camel strains of E. granulosus, as well as variants from E. multilocularis, Echinococcus oligarthrus and Echinococcus vogeli and (4) a distinct lineage of ITS-1 variants including E. granulosus strains from sheep and cervid, and E. multilocularis. At least two of the species (E. granulosus and E. multilocularis) were paraphyletic for ITS-1. Divergent ITS 1 variants from these two species shared distinct evolutionary lineages. The sequence data provided evidence that at least two turnover mechanisms, namely slippage and unequal crossing over/transposition, have led to the divergence and maintenance of sequence variants in Echinococcus species and strains. PMID- 10704600 TI - Phylogeny of the monopisthocotylea and Polyopisthocotylea (Platyhelminthes) inferred from 28S rDNA sequences. AB - This study focuses on the phylogenetic relationships within the Polyopisthocotylea and Monopisthocotylea, two groups that are often grouped within the monogeneans, a group of disputed paraphyly. Phylogenetic analyses were conducted with multiple outgroups chosen according to two hypotheses, a paraphyletic Monogenea or a monophyletic Monogenea, and with three methods, namely maximum parsimony, neighbour joining and maximum likelihood. Sequences used were from the partial domain C1, full domain D1, and partial domain C2 (550 nucleotides, 209 unambiguously aligned sites) from the 28S ribosomal RNA gene for 16 species of monopisthocotyleans, 26 polyopisthocotyleans including six polystomatids, and other Platyhelminthes (61 species in total, 27 new sequences). Results were similar with outgroups corresponding to the two hypotheses. Within the Monopisthocotylea, relationships were: ?[(Udonella, capsalids), monocotylids], (diplectanids, ancyrocephalids)?; each of these families was found to be monophyletic and their monophyly was supported by high bootstrap values in neighbour joining and maximum parsimony. Within the Polyopisthocotylea, the polystomatids were the sister-group of all others. Among the latter, Hexabothrium, parasite of chondrichthyans, was the most basal, and the mazocraeids, mainly parasites of clupeomorph teleosts, were the sister-groups of all other studied polyopisthocotyleans, these, mainly parasites of euteleosts, being polytomous. PMID- 10704601 TI - Internal transcribed spacer rDNA can be used to infer the phylogenetic relationships of species within the genus Nematodirus (Nematoda: molineoidea). AB - Sequences of the first internal transcribed spacer rDNA were characterised for four veterinary important species of gastrointestinal nematodes from the genus Nematodirus. The sequence data were combined with previously published data of the second internal transcribed spacer to determine whether these rDNA regions provided a suitable number of informative characters to determine the phylogenetic relationships of species within the genus. A total of 32 alignment positions of the first internal transcribed spacer data set and 33 characters from the second internal transcribed spacer data set were informative in phylogenetic analyses. Irrespective of whether the data from each spacer were analysed separately or combined, only one most parsimonious tree was produced, with the relationships of the four species fully resolved. In addition, several regions of conservatism in the first internal transcribed spacer sequence among the four Nematodirus species suggests that this rDNA region may also provide phylogenetic information for higher taxonomic levels within the Molineoidea. PMID- 10704602 TI - Temporal effects of protein nutrition on the growth and immunity of lambs infected with Trichostrongylus colubriformis. AB - The aim of this research was to determine whether metabolisable protein supply during the early period of infection with Trichostrongylus colubriformis influenced resilience and the later stages in the development and magnitude of host resistance in previously nematode-naive lambs. Eighty TexelxGreyface lambs were fed pelleted feeds calculated to provide grossly different amounts of metabolisable protein. Sixty of the lambs received a trickle infection of T. colubriformis and 20 lambs were kept as uninfected controls. There were four initial groups, namely infected or uninfected and fed either a moderate or a high protein feed. After 5weeks of infection, a further four groups were established by changing the feed of half of the animals fed the moderate protein feed to the high protein feed and of half of the animals fed high protein to the moderate protein feed. Live weight gain and feed conversion ratio were greatest for lambs fed the high protein feed and were reduced by infection. Faecal egg counts, worm burdens and per capita fecundity of adult female nematodes were unaffected by changes to metabolisable protein supply. Decreasing metabolisable protein supply following 5weeks of infection reduced live weight gain without any effect on resistance to T. colubriformis. Haematological variables, indicative of improved resistance, were also largely unaffected by metabolisable protein supply. It is concluded that the requirements of immune function probably had priority over those of growth and that the metabolisable protein supply provided by the moderate protein feed was sufficient to account for the requirements for the expression of immunity. It is probable that the potential for metabolisable protein supply to enhance resistance to infection from T. colubriformis is dependent on the levels and magnitude (i.e. in relation to maintenance requirements) of metabolisable protein supply being compared and the demand of other competing physiological functions. PMID- 10704603 TI - High molecular mass glycans are major structural elements associated with the laminated layer of in vitro cultivated Echinococcus multilocularis metacestodes. AB - The laminated layer of the larval stage (metacestode) of the cestode parasite Echinococcus multilocularis is composed largely of carbohydrates, which form a tight microfibrillar meshwork around the entire metacestode. Since this laminated layer is the only parasite structure which is in constant contact with host immune and non-immune cells, and appears largely resistant to physiological and immunological reactions of the host, it most likely carries out important functions with regard to host-parasite interactions. In infected hosts, the metacestode is usually concentrically covered by host connective tissue cells and large amounts of collagen, causing a dense scar-like fibrosis, and it is likely that host-derived components are incorporated into the laminated layer at the host-parasite interface. Therefore, in order to obtain information on the molecular composition of this structure, we used parasite larvae which were generated through in vitro cultivation and thus were largely devoid of interfering host components. Lectin fluorescence on section-labelling of metacestodes embedded in LR-White suggested that the laminated layer is largely composed of N-acetyl-beta-D-galactosaminyl, and alpha- and beta-D-galactosyl residues, as well as of the core structure of O-linked carbohydrate chains, N acetylgalactosamine-beta-1.3-galactose, while N-linked glycopeptides and alpha-D mannosyl residues and/or glucosyl residues were found mainly within the germinal layer, and within the cellular mass and the surface of developing protoscoleces. Lectin-gold EM confirmed these findings. The laminated layer was isolated from in vitro cultivated metacestodes by urea extraction, and the ultrastructure of the purified laminated layer was assessed comparatively with respect to the laminated layer of intact parasites. The glycan composition was determined using SDS-PAGE and lectin blotting. This work has laid the basis for a more detailed dissection of the molecular composition of the laminated layer. PMID- 10704604 TI - Infectivity, persistence, and antibody response to domestic and sylvatic Trichinella spp. in experimentally infected pigs. AB - Groups of pigs were inoculated with genotypes of Trichinella belonging to: Trichinella spiralis, Trichinella nativa, Trichinella britovi, Trichinella pseudospiralis (from Caucasus), T. pseudospiralis (from USA), Trichinella murrelli, Trichinella sp. (from North America), and Trichinella nelsoni. The pigs were sacrificed between 5 and 40weeks p.i., and the number of muscle larvae per gram (l.p.g.) of tissue was determined as an average of 18 muscles. All Trichinella genotypes were infective for pigs, but both their infectivity and persistence varied: 5weeks p.i., T. spiralis muscle larvae were present in high numbers (mean=427l.p.g.), while T. britovi, T. nelsoni, and T. pseudospiralis larvae were present in moderate numbers (means=24-52l.p.g.); larvae of the remaining genotypes were recovered only in low numbers (means=0.05-5. 00l.p.g.). The total larval burden (live weight of pigxl.p.g.) was constant over time for T. spiralis, T. britovi, and T. nelsoni, but declined significantly (P<0.05) for the other genotypes. Antibody responses could be detected 3-4weeks p.i. by seven different Trichinella ES antigens, but the antibody levels and dynamics differed significantly among the experimental groups. In pigs inoculated with T. spiralis, T. britovi, or T. nelsoni, the antibody level increased rapidly between weeks 3 and 5 p.i. and was stable or increased slightly throughout the experimental period. In pigs inoculated with T. nativa, T. murrelli, or Trichinella (T6) (from North America), a rapid increase was detected between weeks 3 and 5 p.i., but for these genotypes a reduction in the antibody levels was seen thereafter. In the pigs inoculated with T. pseudospiralis, the antibody level increased more gradually over a period from week 3 p. i. to weeks 15-20 p.i., and decreased thereafter. In general, all species of Trichinella were detected by any of the seven ES antigens, which points to the potential use of one common antigen for surveillance and epidemiological studies on both domestic and sylvatic Trichinella in pigs. Homologous ES antigens were slightly more sensitive in detecting antibodies to the corresponding Trichinella species. PMID- 10704605 TI - Genetic markers in ribosomal DNA for the identification of members of the genus Anisakis (Nematoda: ascaridoidea) defined by polymerase-chain-reaction-based restriction fragment length polymorphism. AB - Polymerase-chain-reaction-based restriction fragment length polymorphism analysis was performed to establish genetic markers in rDNA, for the identification of the three sibling species of the Anisakis simplex complex and morphologically differentiated Anisakis species, i.e. Anisakis physeteris, Anisakis schupakovi, Anisakis typica and Anisakis ziphidarum. Different restriction patterns were found between A. simplex sensu stricto and Anisakis pegreffii with two of the restriction endonucleases used (HinfI and TaqI), between A. simplex sensu stricto and A. simplex C with one endonuclease (HhaI), and between A. simplex C and Aniskis pegreffii with three endonucleases (HhaI, HinfI and TaqI), while no variation in patterns was detected among individuals within each species. The species A. physeteris, A. schupakovi, A. typica and A. ziphidarum were found to be different from each other and different from the three sibling species of the A. simplex complex by distinct fragments using 10-12 of the endonucleases tested. The polymorphisms obtained by restriction fragment length polymorphisms have provided a new set of genetic markers for the accurate identification of sibling species and morphospecies. PMID- 10704607 TI - Corrigendum to "A nested PCR for the ssrRNA gene detects trypanosoma binneyi in the platypus and trypanosoma sp. in wombats and kangaroos in Australia" PMID- 10704608 TI - Absence of atherosclerosis in human intramyocardial coronary arteries: a neglected phenomenon. AB - Atherosclerosis is absent in human intramyocardial (buried) coronary arteries but atherosclerosis may be severe in superficial segments of the same vessels. The development of atherosclerosis has three phases: a plasma phase, a transfer phase and an intramural phase. The transfer phase involves the transfer of low-density lipoproteins and macrophages from the plasma into the arterial wall. Efficiency of transfer is low where plasma flow near the wall is rapid. Eddy currents caused by arterial branches produce low flow near the arterial wall. Plasma recalculates and moves slowly in these eddy currents and thus prolongs contact of LDL and macrophages with the wall, increasing the occurrence of atherosclerosis. Absence of atherosclerosis in buried vessels appears due to the effects of myocardial contraction on the transfer phase. Contraction of the myocardium surrounding buried arterial vessel compresses these vessels and moves the plasma, LDL and macrophages away from the wall. This will decrease transfer into the wall and act to prevent the development of atherosclerosis. Similar but less striking effects occur where bridges of myocardium cross arterial vessels. Possible applications to human disease are discussed briefly. PMID- 10704609 TI - Macrophage plasma membrane chondroitin sulfate proteoglycan binds oxidized low density lipoprotein. AB - Lipoprotein interactions with macrophage proteoglycans (PGs) is believed to play an important role in the cellular uptake of lipoproteins and in macrophage cholesterol accumulation. Recently, we have shown the participation of macrophage plasma membrane glycosaminoglycans (GAGs) in the cellular uptake of oxidized LDL (Ox-LDL). The aim of the present study was to identify the specific cell surface proteoglycans involved in this interaction. J-774 A.1 macrophage-like cell line plasma membrane proteoglycans were isolated by anion exchange chromatography from cells that were prelabeled with [35S]sodium sulfate. Using Sepharose 6B chromatography, cell surface major proteoglycans were identified as chondroitin sulfate (CS) proteoglycans (77%) and heparan sulfate (HS) proteoglycans (23%). Binding rates of these 35S-labeled proteoglycans to Ox-LDL and to native LDL were analyzed by their ability to bind lipoproteins coupled to a CnBr-activated Sepharose CL-4B chromatography. Of the total labeled cell surface proteoglycans added to the column, 57% were bound to the Sepharose-coupled Ox-LDL, whereas 73% of the cell surface proteoglycans were bound to the Sepharose-coupled native LDL. Binding of the plasma membrane macrophage 35S-labeled proteoglycans to Ox-LDL was inhibited by adding increasing concentrations of non-labeled chondroitin sulfate, or by pretreatment of the 35S-labeled proteoglycans fraction with chondroitinase ABC. In contrast, neither the addition of non-labeled heparan sulfate, nor pretreatment of the labeled proteoglycans fraction with heparinase III, had any significant effect on proteoglycan binding to Ox-LDL. These findings were further supported by using mutant cells characterized by specific glycosaminoglycan deficiencies. Ox-LDL binding and degradation by mutant 745 CHO cells which are characterized by a deficiency in both heparan sulfate and chondroitin sulfate, was decreased by 28 and 27% respectively, compared to the binding of Ox-LDL to the wild-type CHO cells. Ox-LDL binding and degradation by mutant 677 CHO cells, which lack heparan sulfate but have increased levels of chondroitin sulfate, however, was found to be increased by 29 and 19%, respectively, compared to Ox LDL binding to the wild-type CHO cells. Finally, analysis of the cell surface proteoglycans in macrophages that were subjected to oxidative stress, by their preincubation with angiotensin II, exhibited a 51-59% increase in their cell surface proteoglycan content, with a major effect on chondroitin sulfate proteoglycans. The present study thus demonstrated that Ox-LDL can specifically bind to macrophage surface chondroitin sulfate proteoglycans, and the macrophage content of this proteoglycan is increased under oxidative stress. The interaction between macrophage chondroitin sulfate proteoglycans and Ox-LDL can contribute to enhanced uptake of Ox-LDL with the formation of cholesterol-loaded foam cells, and accelerated atherosclerosis. PMID- 10704610 TI - Endothelium-dependent and -independent functions are impaired in patients with coronary heart disease. AB - Endothelium plays a pivotal role in the development of atherosclerosis. Endothelial dysfunction participates in the course of acute coronary event. Using high-resolution ultrasound technique, endothelial dysfunction has been demonstrated in patients with atherosclerosis and risk factors for coronary disease, such as hypertension, diabetes mellitus, hypercholesterolemia and being smokers. In the present study, using this non-invasive method, the endothelial function of the brachial artery of patients with coronary heart disease (CHD) (n = 71) and control subjects (n = 34) was investigated. The results showed that endothelium-dependent and -independent vasodilatation were impaired in patients with CHD (2.61+/-2.91 vs. 8.10+/-7.81%, 17.20+/-7.93 vs. 23.19+/-8.89%, respectively) (P<0.001). Flow-mediated dilation (FMD) was significantly positively correlated with nitroglycerine-induced dilation (P<0.001). On univariate and multivariate analysis, the extent of FMD was significantly correlated with serum HDL-C levels (P<0.01). In conclusion, our study indicates both endothelial and underlying smooth muscle functions were impaired in patients with CHD. Decreased HDL-C levels may impair endothelial function. PMID- 10704611 TI - Caco-2 cells secrete two independent classes of lipoproteins with distinct density: effect of the ratio of unsaturated to saturated fatty acid. AB - Polarized Caco-2 cells can synthesize two distinct density classes of lipoproteins, i.e. chylomicron/VLDL (d<1.006 g/ml) or IDL/LDL density (1.0093.36 mmol/l consumed each of four diets for 32-day periods. The diets contained 30% energy as fat of which 2/3 was either corn oil or beef tallow with and without 115 mg/4.2 MJ of supplemental cholesterol in the form of cooked egg yolk. The susceptibility of LDL to oxidation was assessed during a challenge with hemin and hydrogen peroxide, and results are expressed as lag time to oxidation in minutes. Addition of moderate amounts of cholesterol to either the corn oil or beef tallow enriched diet resulted in increased susceptibility of LDL to oxidation (decreased lag time): 69+/-22 min versus 96+/-24 min in the corn oil diet with versus without supplemental cholesterol, respectively, P = 0.006; 82+/-20 min versus 96+/-26 min in the beef tallow diet with versus without supplemental cholesterol, respectively, P = 0.025. A stepwise equation indicated that as plasma oleic acid concentrations increased and/or linoleic acid concentrations decreased, lag time increased (decreased susceptibility to oxidation), whereas as dietary cholesterol concentrations increased, lag time decreased (increased susceptibility to oxidation). In conclusion, these data suggest that addition of a moderate amount of dietary cholesterol to a reduced fat diet rich in polyunsaturated or saturated fatty acids increased the in vitro susceptibility of LDL to oxidation. PMID- 10704619 TI - Common paraoxonase gene variants, mortality risk and fatal cardiovascular events in elderly subjects. AB - Recent studies indicate that the enzyme paraoxonase may be an important modulator of cardiovascular disease risk because of its ability to protect LDL from oxidation. We tested for association between two functional variants of the paraoxonase gene (Met-55/Leu and Gln-192/Arg) and both all-cause mortality and fatal cardiovascular disease. This was done within a population-based study among subjects aged 85 years and over in a cross-sectional and a prospective design. In the cross-sectional analysis, the distribution of both paraoxonase genotypes was found to be similar in the subset of 364 elderly subjects who were born in Leiden, The Netherlands, as compared with 250 young subjects whose families originated from the same geographical region. The polymorphisms were in strong linkage disequilibrium (P<0.00001) and the frequency of the haplotype carrying both risk alleles was not lower in the elderly than in the young (0.313 vs. 0.284). The complete cohort of 666 elderly subjects was followed over 10 years. The risk of all-cause and cardiovascular mortality was not increased in elderly subjects with the paraoxonase Leu/Leu (RR, 1.1 [95% CI, 0.9-1.5] and 1.3 [95% CI, 0.8-2.0], respectively) or the Arg/Arg genotype (RR, 0. 9 [95% CI, 0.7-1.2] and 0.7 [95% CI, 0.4-1.3], respectively). In a subset of patients with diabetes, the all-cause mortality risk was elevated in Arg/Arg carriers (RR, 2.1 [95% CI, 0.8 5.8]) but this did not reach statistical significance. Analysis of genotype combinations did not yield significant associations with mortality. The paraoxonase gene variants, previously associated with coronary artery disease, are thus not likely to have a major effect on the risk of fatal cardiovascular disease in the population at large. Adverse effects of the gene variants might be observed in subjects exposed to factors that enhance oxidative stress such as diabetes. PMID- 10704620 TI - ICAM-1 expression by vascular smooth muscle cells is phenotype-dependent. AB - Atherosclerosis is an inflammatory disease characterised by increased expression of adhesion molecules for leukocytes on both the surface of dysfunctional endothelium and on smooth muscle cells (SMC) within the lesion. It is also characterised by altered SMC phenotypic expression, indicated by a decreased volume fraction of myofilaments (V(v)myo) [1,2] and changes in gene expression [3]. The present study used an in vitro model to investigate, by immunofluorescence staining and flow cytometry, the influence of phenotype on vascular SMC expression of the adhesion molecule for leukocytes, intracellular adhesion molecule-1 (ICAM-1), and the regulatory mechanisms involved in this process. Smooth muscle cells with a high V(v)myo, freshly isolated from rat aortic media, expressed little or no ICAM-1 and this could not be induced by interleukin-1beta (IL-1beta). As SMC modulated phenotype, indicated by decreasing V(v)myo over the first 5 days of culture, there was a concomitant increase in ICAM-1 expression. At day 9 of primary culture, when SMC cultures had returned to the high V(v)myo phenotype, ICAM-1 expression was markedly lower. However, these cells retained the capacity to express ICAM-1 in response to IL-1beta. After several passages in culture, cells (with a low V(v)myo) constitutively expressed ICAM-1, with levels further up-regulated in response to IL-1beta. These changes in ICAM-1 expression were not related to proliferative state, since similar results were obtained with growth arrested SMC. Investigation of signalling pathways involved in regulating ICAM-1 expression by primary vascular SMC suggested a complex regulatory mechanism. Activation of adenyl cyclase (with forskolin) caused a significant increase in cells expressing ICAM-1. Treatment with inhibitors of protein kinase C (chelerythrine chloride), protein tyrosine kinase (genistein), or the transcription factor NF-kappaB (PDTC) had no significant effect on IL-1-induced ICAM-1 expression. However, in the presence of serum, both genistein and PDTC caused a significant increase in basal expression. The results indicate that ICAM-1 expression by SMC is phenotype-dependent, with expression evident only after cells have modulated to a low V(v)myo phenotype. They also indicate the existence of complex regulatory mechanisms, possibly involving the SMC cytoskeleton. PMID- 10704621 TI - Inhibition of tissue-factor-mediated thrombin generation by simvastatin. AB - A previous study has shown that simvastatin reduces in vivo clotting activation and monocyte tissue factor (TF) expression. This effect, however, was only in part attributable to the reduction of serum cholesterol, suggesting that more than one mechanism may be involved. Furthermore, it was not investigated if the inhibition of clotting activation was dependent upon the reduced expression of monocyte TF. In order to assess if simvastatin directly affects clotting activation, we developed an in vitro method in which clotting system is activated by monocytes stimulated with LPS. Monocytes were prepared from blood taken from healthy volunteers or patients with hypercholesterolemia and incubated with heparinized plasma plus either simvastatin (0.01-10 microM) or medium as control. Samples were then stimulated with LPS (4 microg/ml) and after 6 h the rate of thrombin generation, assessed by prothrombin fragment (F) 1+2, was measured. In separate experiments, we measured the expression of TF by monocytes which were incubated with simvastatin and then stimulated with LPS. The study showed that compared to control, LPS-stimulated monocytes induced abundant formation of F1+2, which was inhibited by simvastatin in a dose-dependent manner. Simvastatin also inhibited dose dependently the monocyte expression of TF. This study suggests that simvastatin inhibits the rate of thrombin generation by directly interfering with the monocyte expression of TF. PMID- 10704622 TI - Serum N-acetyl-beta-D-gulucosaminidase activity increases in association with insulin resistance in patients with coronary artery disease. AB - N-acetyl-beta-D-glucosaminidase (NAG) is released from lysosomes, but the clinical significance of its serum activity in the pathogenesis of coronary artery disease has not been well understood. We measured serum NAG activity by a colorimetric method in consecutive 168 patients suspected of having coronary artery disease who underwent diagnostic coronary angiography. In addition, we evaluated the relationship between the activity and severity of coronary artery disease, as well as various coronary risk factors. Serum NAG activity was higher in the multi-vessel disease group than in the no stenotic lesion group (9.2+/-2.3 vs. 7.8+/-1.8 U/l, P<0.01) and in the single-vessel disease group (vs. 8.2+/-2.2 U/l, P<0.05). In all patients, Gensini score was closely correlated with the serum NAG activity (r = 0.39, P<0.001). Multiple regression analysis showed that serum NAG activity was correlated with plasma insulin level (r = 0.49, P<0.01), but not correlated with other coronary risk factors. In 126 patients without apparent diabetes mellitus, serum NAG was also correlated with plasma insulin level (r = 0.37, P<0.01) and additionally with insulin resistanc determined by homeostasis model assessment (r = 0.47, P<0.01). Our results suggested that serum NAG activity correlates with the severity of coronary artery disease in relation to plasma insulin level and insulin resistance, and thus can be an indicator of coronary artery disease based upon abnormalities of glucose metabolism. PMID- 10704623 TI - Safety profile of atorvastatin-treated patients with low LDL-cholesterol levels. AB - Data pooled from 21 atorvastatin clinical trials have been analyzed to establish the safety of reducing low density lipoprotein cholesterol (LDL-C) levels below currently recommended minimum targets in hypercholesterolemic patients. Safety data for atorvastatin-treated patients with at least one LDL-C value < or =80 mg/dl (2.1 mmol/l) (n = 319) during treatment (mean LDL-C level throughout treatment was 91 mg/dl [2.4 mmol/l]) were compared to those from all atorvastatin treated patients (n = 2502) and patients treated with lovastatin, simvastatin or pravastatin (n = 742). The frequency of treatment-associated adverse events (AEs) in the atorvastatin LDL-C < or =80 mg/dl (2.1 mmol/l) subgroup (24%) was comparable to the frequencies observed for all atorvastatin-treated patients (20%) and for patients receiving the other statins (24%). Patient withdrawals due to treatment-associated AEs (constipation, dyspepsia and flatulence being the most common) were consistent and low across treatment groups. No treatment associated deaths occurred in any group. Safety data for 21 atorvastatin-treated patients with LDL-C < or =50 mg/dl (1.3 mmol/l) were also analyzed and found to be similar to all atorvastatin-treated patients and patients treated with the other statins. While recognizing the short-term nature of the data (all patients who received atorvastatin were treated for < or =1 year and approximately 30% were treated for < or =6 months), this analysis suggests that reducing LDL-C levels below 80 (2.1 mmol/l) or 50 mg/dl (1.3 mmol/l) with atorvastatin does not alter its safety profile, as measured by frequency of AEs, which remains similar to those of other statins. PMID- 10704624 TI - Polygenic influence on plasma homocysteine: association of two prevalent mutations, the 844ins68 of cystathionine beta-synthase and A(2756)G of methionine synthase, with lowered plasma homocysteine levels. AB - A moderately elevated plasma total homocysteine (tHcy), whether measured during fasting or post-methionine load (PML), is increasingly being recognized as a risk factor for coronary artery diseases (CAD). However, etiologies for moderately elevated plasma tHcy, particularly with regard to the role of genetic influence on plasma tHcy levels, are still not well understood. In the current investigation, we studied 1025 individuals with respect to the effect of the 68 bp insertion (844ins68 variant) of the cystathionine beta-synthase (CBS) gene, the A(2756)G transition of the B(12)-dependent methionine synthase (MS) gene and the C(677)T transition of the methylenetetrahydrofolate reductase (MTHFR) gene on fasting and 4 h PML tHcy. Of these individuals, 153 (14.9%) were heterozygous for the 68-bp insertion, 329 (32.1%) were heterozygous for the G(2756) allele and 122 (11.9%) were homozygous for the C(677)T transition. Individuals heterozygous for the insertion had significantly lower PML increase in tHcy concentrations, while individuals homozygous for the A(2756)G transition had significantly lower fasting tHcy levels. A 2-way ANOVA showed that there was no interaction between the 844ins68 and the A(2756)G transition for either fasting tHcy or PML increase in tHcy, confirming the fact that the effect of these two genotypes on plasma tHcy levels are additive. The effects are opposite but additive with the C(677)50% of all individuals in this study carried polymorphic traits, which predisposed them to either higher or lower plasma tHcy concentrations, thus providing new evidence of the importance of genetic influences as determinants of tHcy levels. PMID- 10704625 TI - C-reactive protein concentration in children: relationship to adiposity and other cardiovascular risk factors. AB - Whether or not C-reactive protein (CRP) predicts heart disease in adults because it is a marker of damage or atherosclerosis is difficult to assess. In children, there is no confounding with coronary disease or active smoking. We measured CRP in 699 children aged 10-11 years. CRP levels were 47% higher in girls than boys, and rose with age by 15%/year. CRP levels were 270% (95% CI, 155-439%) higher in the top fifth than the bottom fifth of Ponderal index (weight/height(3)). After adjustment, CRP levels remained 104% (95% CI, 23-236%) higher in the 56 children of South Asian origin. CRP was unrelated to: birth weight, height, social class, Helicobacter pylori infection or passive smoke exposure. CRP was correlated with several cardiovascular risk factors, but only fibrinogen (r = 0.33, P = 0.0001), HDL-cholesterol (r = -0.13, P = 0.0006), heart rate (r = 0.12, P = 0.002) and systolic blood pressure (r = 0.08, P = 0.02) remained statistically significant after adjustment. We conclude that adiposity is the major determinant of CRP levels in children while physical fitness has a small independent effect. The strong relationships with fibrinogen and HDL-cholesterol suggest a role for inflammation throughout life in the development of atherosclerosis and cardiovascular disease. Longitudinal studies are needed to determine whether these associations reflect long term elevations of these risk factors in some individuals, or short term fluctuations in different individuals. PMID- 10704626 TI - G protein beta3 subunit polymorphism and risk of thrombosis and restenosis following coronary stent placement. AB - C825T polymorphism in the G protein beta3 subunit gene (GNB3) has been associated with arterial hypertension, coronary artery disease and myocardial infarction. On the cellular level, C825T polymorphism is associated with altered transmembrane signaling via adenylyl cyclase inhibiting (G(i)) G proteins. This study was designed to test whether C825T polymorphism has an impact on the processes leading to restenosis and thrombosis following coronary stenting. The primary endpoint of the study was angiographic restenosis (> or =50% diameter stenosis) at 6-month follow-up. Secondary endpoint was angiographically proven stent thrombosis within 30 days of implantation. In the 562 consecutive patients C825T genotype was CC, 46.1%, CT, 45.2% and TT, 8.7%. The incidence of angiographic restenosis was 32.7% in homozygous carriers of the C allele, 28.2% in CT patients and 33.3% in homozygous carriers of the T allele (P = 0.563). C825T genotype distribution in 34 consecutive patients with subacute stent thrombosis (44.0% CC, 50.0% CT, and 6.0% TT) was not different from that of 451 patients with angiographically patent stented vessel (45.4% CC, 44.6% CT, 10.0% TT; P = 0.644). In conclusion, C825T polymorphism has no appreciable impact on the mechanisms leading to thrombosis and restenosis following coronary stent placement. PMID- 10704627 TI - Insulin-like growth factor binding protein-1 (IGFBP-1) and IGF-I during oral and transdermal estrogen replacement therapy: relation to lipoprotein(a) levels. AB - Low levels of insulin-like growth factor binding protein-1 (IGFBP-1) have recently been associated with several risk factors for cardiovascular disease. The effects of estrogen replacement therapy (ERT) on plasma IGFBP-1 levels are, however, unclear. A double-blind, placebo-controlled study for 6 months was conducted in 73 hysterectomized postmenopausal women randomized into two groups: oral estradiol (E2) valerate, 2 mg/day (n = 35) and transdermal E2 gel, 1 mg/day (n=38). Plasma IGFBP-1, insulin-like growth factor-I (IGF-I) and lipoprotein(a) (Lp(a)) were determined at baseline, 3 and 6 months. The groups were similar for age and BMI. The baseline levels of estrone (E1), E2, IGFBP-1, IGF-I and Lp(a) did not differ between the groups. During treatment, serum estradiol concentrations increased in both groups. During oral ERT, IGFBP-1 levels increased by 104% (P<0.001), whereas IGF-I levels decreased by 13% (mean, P<0.05). IGF-I and IGFBP-1 levels remained unchanged in the transdermal group. Lp(a) levels decreased by 23% (median, P<0.001) in the oral group, but were unaffected by transdermal therapy. The change in IGFBP-1 concentrations during oral ERT showed an inverse correlation to that in Lp(a) (r = -0.40, P<0.05, Spearman correlation). In conclusion, oral ERT seems to enhance plasma levels of IGFBP-1, which may be one reason for the reduced Lp(a) levels. PMID- 10704628 TI - Increased plasma homocysteine after menopause. AB - Besides genetic defects in the enzymes involved in homocysteine metabolism and nutritional deficiencies in vitamin cofactors, sex steroid hormones may modulate plasma homocysteine levels. The post-menopausal state has been found to be associated with higher plasma homocysteine levels, but data are inconsistent and studies published so far did not adjust for age, which is an important confounding factor in studying the effect of menopause. In the present study total plasma homocysteine levels were measured in a meticulously selected population in which the contrast in estrogen status between pre- and postmenopausal women of the same age was maximized. The study comprised 93 premenopausal and 93 postmenopausal women of similar age (range 43-55 years). Women were selected from respondents to a mailed questionnaire on menopause, which was sent to all women aged 40-60 years in the Dutch town of Zoetermeer (n = 12675). Postmenopausal women who were at least three years after menopause or whose menses had stopped naturally before age 48 were age-matched with premenopausal women with regular menses and without menopausal complaints. Plasma homocysteine levels in the fasting state were related to menopausal status; the age-adjusted geometric mean was 10.7 micromol/l in premenopausal and 11.5 micromol/l in postmenopausal women (difference of 7%, 95% confidence interval 0.3 14%, P = 0.04). Additional adjustment for plasma creatinine, body mass index, smoking habit (yes, no) and alcohol intake did not influence this difference. The results of this population-based study indicate that plasma homocysteine is affected by menopause. PMID- 10704629 TI - Chylomicron processing in familial dysbetalipoproteinemia and familial combined hyperlipidemia studied with vitamin A and E as markers: a new physiological concept. AB - In previous work we identified a transfer/diffusion process occurring in the postprandial state that more or less contributes to the accumulation of beta-VLDL in familial dysbetalipoproteinemia (FD). Here we present a new theoretical concept underlying chylomicron processing developed on the basis of extended quantitative analyses of fat loading experiments, with both vitamins A and E, performed in patients with familial combined hyperlipidemia (FCH) in comparison to patients with FD and control subjects. Recovery of triglycerides from the fat load in the plasma triglyceride pool was <4%, indicating a very effective lipolysis process with an active remnant generation. Vitamin A from the fat load was, over 48 h, quantitatively recovered in the plasma lipoprotein pool; vitamin E was recovered to 2241%. Nevertheless, transfer/diffusion of both vitamins showed similar patterns. At equilibrium, their contents correlated strongly with the lipoprotein concentrations, the slopes being similar for control subjects and both groups of patients. Only in those FD patients with the highest lipid values, did the vitamin A/lipoprotein mass ratio in the Sf>100 fraction deviate from the total group mean. In the Sf 15-100 fraction, most specific for 'remnants', vitamin A/cholesterol ratios for all subjects were uniform proving that beta-VLDL formation is a thermodynamic process regulated by concentration gradients and the lipophilicity of lipoprotein constituents, not a typical feature for patients with FD. In patients with FD, vitamin A in the plasma pool was recovered excessively (276%) in line with recognition in various pools as a result of the transfer/diffusion process in plasma. PMID- 10704630 TI - The relationship of oxidized lipids to coronary artery stenosis. AB - A total of 1200 patients with angina were cardiac catheterized establishing that 63% had 70-100% stenosis, 12% had 10-69% stenosis of one or more of their coronary arteries and 25% had microvascular angina listed as 0% stenosis. Prior to catheterization 10 ml of blood was drawn and the plasma subjected to analysis for the concentration of cholesterol, lipid peroxides (LPX), total antioxidant capacity (TAOC), fibrinogen (FB), ceruloplasmin (CP) and activation of polymorphonuclear leukocytes (PMNLs). Comparisons were made to non-smoking controls without angina. Significant differences in LPX were found between the patients with 0 and 10-69% stenosis (P<0.001), with 10-69 and 70-100% stenosis (P<0.001), and with 0 and 70-100% stenosis (P<0.001). Under 70 years of age there was a significant difference in LPX between patients with all levels of stenosis and age and sex matched controls (P<0.001). Differences in the mean plasma cholesterol concentration for different levels in the degree of stenosis were not significant, indicating that LPX provided consistent data on the severity of stenosis while the plasma cholesterol concentration did not. Compared with controls an increase in activation of PMNLs (P<0.01), an increase in concentration of both FB and CP (P<0.01) and a decrease in total antioxidant capacity were noted in the plasma of catheterized patients. In summary the concentration of oxidation products rather than the concentration of cholesterol in the plasma identified stenosis in cardiac catheterized patients. PMID- 10704631 TI - An excess concentration of oxysterols in the plasma is cytotoxic to cultured endothelial cells. AB - To test if there is an excess concentration of oxysterols in the plasma of the patients with cardiovascular disease, we analyzed the oxysterol content in the plasma from 105 cardiac catheterized patients with angina and 80+/-8% stenosis in their coronary arteries. The result showed that the plasma contained a significantly higher concentration of oxysterols than did plasma from 105 age- and sex-matched, non-catheterized and angina-free controls (P<0.05). We used endothelial cells (ECs) cultured in medium containing either [3H]thymidine, [3H]mevalonolactone or 45Ca(2+) to determine how the plasma from the patients influences cell growth and function. We found that less [3H]thymidine (P<0.05), less [3H]mevalonolactone (P<0.05) and more 45Ca(2+) (P<0.001) was incorporated into ECs cultured in the plasma from 36 patients with 83+/-4% stenosis than from the 36 controls. When synthetic 7beta-hydroxycholesterol, cholesterol 5beta,6beta epoxide, cholesterol 5alpha,6alpha-epoxide and 7-ketocholesterol were added to the plasma from the controls, the influx of 45Ca(2+) into ECs then equaled that in the plasma of patients. The enhanced incorporation of 45Ca(2+) into the ECs cultured in the plasma both from the patients and from controls with added synthetic oxysterols substantiates in vitro the hypothesis that oxysterols increase the influx of calcium into cells. These data indicated that an excess of oxysterols in the plasma of the patients was cytotoxic to the cultured cells. PMID- 10704632 TI - Achievement of target plasma cholesterol levels in hypercholesterolaemic patients being treated in general practice. AB - A total of 1028 hypercholesterolaemic men and women aged 18-75 participated in an open label, randomised, parallel group, 6-month treatment-to-target study conducted in 240 general practices throughout Australia. The study compared atorvastatin monotherapy with simvastatin monotherapy or, if necessary, with the combination of simvastatin and cholestyramine in terms of their abilities to achieve a plasma total cholesterol target of<5.0 mmol/l. The initial daily dose of each drug was 10 mg. If the target was not achieved, the dose was doubled at 6 week intervals to a maximum daily dose of 80 mg atorvastatin or 40 mg simvastatin, with the simvastatin supplemented if necessary with 4 g cholestyramine. The percentage of patients achieving the target at 10 and 20 mg doses of atorvastatin were comparable to 20 and 40 mg of simvastatin, respectively. Despite relatively high baseline levels of plasma total cholesterol (mean levels of 7.41 and 7.31 mmol/l in the atorvastatin and simvastatin groups, respectively) the majority of patients in each group achieved the plasma total cholesterol target of<5.0 mmol/l. Treatment with atorvastatin achieved the target in 83% of patients, while simvastatin (or simvastatin plus cholestyramine) achieved the target in 66% of the patients (P<0.005). The target was achieved with 10 mg atorvastatin in 38% of patients and with 10 mg simvastatin in 26% of cases (P<0.005). In patients whose baseline cholesterol levels were between 5.6 and 6.5 mmol/l, 95% of the atorvastatin group and 86% of the simvastatin group reached the target. Even with baseline cholesterol levels between 7.6 and 8.5 mmol/l, the target was reached in 78% of the atorvastatin group and 61% of the simvastatin group. It is thus realistic for general practitioners to expect the majority of their at risk patients to achieve target plasma cholesterol levels that have been shown in population studies to be associated with relatively low rates of coronary heart disease. These targets are achieved in significantly more patients and at lower mg doses with atorvastatin than simvastatin. PMID- 10704633 TI - Community-living nonagenarians in northern ireland have lower plasma homocysteine but similar methylenetetrahydrofolate reductase thermolabile genotype prevalence compared to 70-89-year-old subjects. AB - This cross-sectional study assessed relationships between plasma homocysteine, 'thermolabile' methylenetetrahydrofolatereductase (MTHFR) genotype, B vitamin status and measures of renal function in elderly (70-89 years) and nonagenarian (90+ years) subjects, with the hypothesis that octo/nonagenarian subjects who remain healthy into old age as defined by 'Senieur' status might show reduced genetic or environmental risk factors usually associated with hyperhomocysteinaemia. Plasma homocysteine was 9.1 micromol/l (geometric mean [GM]) for all elderly subjects. Intriguingly, homocysteine was significantly lower in 90+ (GM; 8.2 micromol/l) compared to 70-89-year-old subjects (GM; 9.8 micromol/l) despite significantly lower glomerular filtration rate (GFR) and serum B12 in nonagenarian subjects and comparable MTHFR thermolabile (TT) genotype frequency, folate and B6 status to 70-89-year-olds. For all elderly subjects, the odds ratio and 95% confidence intervals for plasma homocysteine being in the highest versus lowest quartile was 4.27 (2.04-8.92) for age <90 compared >90 years, 3.4 (1.5-7.8) for serum folate <10.7 compared >10.7nmol/l, 3.0 (0.9-10.2) for creatinine >140 compared <140 umol/l and 2.1 (1.0-4.4) for male sex. This study shows that plasma homocysteine does not invariably increase with age. Compared to similarly enlisted 70-89-year-olds, apparently well, mentally alert, community-living 90+ year olds approximating 'Senieur' status, show lower homocysteine, which is unexplained by renal function, TT genotype and B vitamin status, suggesting that lower homocysteine may be associated with survival. PMID- 10704635 TI - Dynamical analysis of density-dependent selection in a discrete one-island migration model. AB - A system of non-linear difference equations is used to model the effects of density-dependent selection and migration in a population characterized by two alleles at a single gene locus. Results for the existence and stability of polymorphic equilibria are established. Properties for a genetically important class of equilibria associated with complete dominance in fitness are described. The birth of an unusual chaotic attractor is also illustrated. This attractor is produced when migration causes chaotic dynamics on a boundary of phase space to bifurcate into the interior of phase space, resulting in bistable genetic polymorphic behavior. PMID- 10704636 TI - A mathematical model to study the effects of drug resistance and vasculature on the response of solid tumors to chemotherapy. AB - A mathematical model is developed that describes the reduction in volume of a vascular tumor in response to specific chemotherapeutic administration strategies. The model consists of a system of partial differential equations governing intratumoral drug concentration and cancer cell density. In the model the tumor is treated as a continuum of two types of cells which differ in their proliferation rates and their responses to the chemotherapeutic agent. The balance between cell proliferation and death within the tumor generates a velocity field which drives expansion or regression of the spheroid. Insight into the tumor's response to therapy is gained by applying a combination of analytical and numerical techniques to the model equations. PMID- 10704637 TI - Contact tracing in stochastic and deterministic epidemic models. AB - We consider a simple unstructured individual based stochastic epidemic model with contact tracing. Even in the onset of the epidemic, contact tracing implies that infected individuals do not act independent of each other. Nevertheless, it is possible to analyze the embedded non-stationary Galton-Watson process. Based upon this analysis, threshold theorems and also the probability for major outbreaks can be derived. Furthermore, it is possible to obtain a deterministic model that approximates the stochastic process, and in this way, to determine the prevalence of disease in the quasi-stationary state and to investigate the dynamics of the epidemic. PMID- 10704638 TI - Qualitative behavior of a variable-yield simple food chain with an inhibiting nutrient. AB - A simple food chain which consists of nutrient, prey and predator in which nutrient is growth limiting at low concentrations but growth inhibiting at high concentrations is investigated in this study. It is assumed that the nutrient concentration is separated into internal and external nutrient concentration and only the internal nutrient level is capable of catalyzing cell growth. It is shown that the dynamics of the system depend on thresholds R(0) and R(1). With inhibition, there exist initial conditions for which the predator becomes extinct but not the prey when R(0)<1. If R(0),R(1)1, the system is uniformly persistent even in the inhibited environment. PMID- 10704639 TI - Distributions of cherries for two models of trees. AB - Null models for generating binary phylogenetic trees are useful for testing evolutionary hypotheses and reconstructing phylogenies. We consider two such null models - the Yule and uniform models - and in particular the induced distribution they generate on the number C(n) of cherries in the tree, where a cherry is a pair of leaves each of which is adjacent to a common ancestor. By realizing the process of cherry formation in these two models by extended Polya urn models we show that C(n) is asymptotically normal. We also give exact formulas for the mean and standard deviation of the C(n) in these two models. This allows simple statistical tests for the Yule and uniform null hypotheses. PMID- 10704640 TI - The density of the extinction probability of a time homogeneous linear birth and death process under the influence of randomly occurring disasters. AB - Under the influence of randomly occurring disasters, the eventual extinction probability, q, of a birth and death process, Z, is a random variable. In this paper, we obtain an integral expression for the probability density function g(x) of q under the assumption that the population process Z is a time homogeneous linear birth and death process and the disasters occur according to an arbitrary renewal process so that its interarrival times have a density. An example is provided to demonstrate how to evaluate the integral numerically. PMID- 10704641 TI - The use of biological stains in the analysis of late Palaeozoic pteridosperm cuticles. AB - The use of biological stains in the cuticular analysis of late Palaeozoic pteridosperms based on specimens from the Stephanian Blanzy-Montceau Basin (Central France) is discussed. Bismarck Brown, Malachite Green G, Methylene Blue, Methyl Green, Neutral Red, Safranin T, and a double staining with Neutral Red and Malachite Green G, were tested. Bismarck Brown, Malachite Green G, Methylene Blue, and Neutral Red increase contrast and emphasize differences in cutinization. The double staining in Neutral Red and Malachite Green G enhances the three-dimensional morphology of complex epidermal structures. Safranin T increases contrast, emphasizes cutinization differences, and enhances the three dimensional morphology of complex epidermal features. The colour photography of cuticles is normally not affected by the presence of stains, but some stains mask black-and-white half-tones. PMID- 10704642 TI - Morphology and ultrastructure of orbicules in the subfamily Ixoroideae (Rubiaceae). AB - Orbicules were studied in 43 species belonging to 32 genera of the five tribes of the Ixoroideae (Rubiaceae). The orbicules were investigated with scanning electron microscopy (SEM), light microscopy (LM), and transmission electron microscopy (TEM). Orbicules are present in all species investigated of the tribes Pavetteae, Octotropideae and Coffeeae. In the tribes Gardenieae and Aulacocalyceae orbicules were found in some species, while they were absent in others. 15 species out of 11 genera lack orbicules. Three orbicule types (III, V, and VI) could be distinguished, mainly on the basis of general morphological and ultrastructural variations. Orbicules that belong to type III (0.50-1.29um) have perforations in their wall, a regular or irregular shape and two or three electron transparent cores. This orbicule type, exclusively found in all Pavetteae species investigated, can be divided into two subtypes. Orbicules of subtype IIIa are present in all genera related to Ixora, and orbicules of subtype IIIb in those genera related to Tarenna. Orbicules of type V (0.97-1.86um) are present in Himalrandia tetrasperma (Aulacocalyceae), and in all Octotropideae genera investigated, except Feretia. Complexes of more than three individual orbicules characterize this type. They are irregularly shaped and have many perforations as well as sporopollenin granules on the orbicule wall. In all species investigated of tribe Coffeeae, type VI orbicules (0.56-1.60um) are present. These orbicules are characterized by their embedded position towards the tapetal membrane, their aggregated form and by the presence of perforations as well as sporopollenin granules on their orbicule wall. In the tribe Gardenieae different types of orbicule were found (V, VI and orbicules that cannot be classified in our typology). Our results suggest that orbicule characters in the Ixoroideae may be systematically useful on tribal level. PMID- 10704643 TI - Modern pollen rain from the Biligirirangan-Melagiri hills of southern Eastern Ghats, India. AB - A total of 39 soil surface samples collected between 11 degrees 30'N 76 degrees 45'E and 12 degrees 45'N 78 degrees 15'E from the mainly deciduous forests in the Biligirirangan-Melagiri hills of the southern Eastern Ghats were analysed for their pollen content. The samples are distributed among four different deciduous and evergreen vegetation types between 210 and 1700m altitudes and fall within three distinct rainfall regimes. The aims of this paper are to provide new data on the modern pollen rain from the Southern Eastern Ghats, a region characterized by a unique and complex climate and vegetation, and to interpret these data using multivariate statistics and the diagram of pollen percentages. We could distinguish first between the deciduous and the evergreen forests and then also between different types of deciduous forest. The distinction between the evergreen and deciduous forests was based on a humidity gradient and that among the deciduous forests on a physiognomic gradient identified through correspondence analysis. The above analysis also allowed us to identify a set of 14 pollen taxa markers and 11 associated pollen taxa that help differentiate the evergreen from deciduous forests. Similarly, a set of 12 pollen taxa markers and six associated pollen taxa was demarcated to help distinguish woodland formations from scrub and thicket formations, among the deciduous vegetation. We could also differentiate amongst the four distinct vegetation types sampled, on the basis of distinct associations of both tree and herb pollen taxa according to their relative abundance in the pollen diagram as well as on the proportion of total arboreal pollen. The ground cover of grasses and other herbaceous plants in the deciduous forests is effectively demonstrated by percentages of non-arboreal pollen varying between 40 and 70%. The 1000m altitude limit reflecting a gradient of humidity and the physiognomic gradient among deciduous forests seem to be important in this region. PMID- 10704644 TI - Modern pollen-representation of some boreal species on islands in a large lake in Canada. AB - Studies of pollen source areas of closed-canopy sites are contradictory. Some authors found that closed-canopy sites mainly collect local pollen while others found more distant sources. This dichotomy might stem from the use of canopies of varying degrees of closure, and from variations in the pollen productivity of the local vegetation and the background pollen rain. Here, 30 islands were used to evaluate the pollen sources of closed-canopy sites. We compared pollen with the forest inventory in three quadrat sizes: 100, 400m(2) and on the whole island. Regression analyses showed that most pollen of Picea spp., Pinus spp., and Betula spp. comes from within the 400m(2) quadrat. Abies balsamea and Thuja occidentalis showed no relationship with vegetation in any of the quadrats considered, suggesting a more regional source. Insularity and island size are important factors influencing the pollen source area; correlations were stronger on islands located 120050ha. These results suggest that closed-canopy sites on islands may be useful in stand-level vegetation history reconstruction through pollen analysis, but that caution must be exercised in separating the local and regional signals. PMID- 10704645 TI - Lower Cretaceous dinoflagellate cyst and acritarch stratigraphy of the Cismon APTICORE (Southern Alps, Italy). AB - A pelagic sedimentary succession, virtually complete from the Upper Hauterivian to the Upper Aptian and unconformably overlain by the Middle-Upper Albian p.p., was continuously cored in the Belluno Basin (southern Alps, NE Italy) as part of the APTICORE Program. APTICORE at Cismon Valley penetrated 131.8m of limestones, marls and black shales, with 100% recovery of good quality cored material.One hundred and forty-six samples recovered from the marl and shale beds of the Cismon core were processed and analyzed for palynomorphs. Most of them yielded relatively rich and fairly well preserved assemblages of marine and terrestrially derived palynomorphs.The results of a qualitative study of dinoflagellate cysts and acritarchs are presented and discussed. The distributions of 150 taxa are tabulated against the chronostratigraphy independently established on the basis of original litho-, bio-, chemo-, magnetostratigraphic investigations and of correlations with extensively studied sections outcropping in the vicinity of the Cismon drill site.The acritarch Pinocchiodinium erbae gen. et sp. nov. is described. Due to its distinctive morphology and extremely constant occurrence also in the black shales of the Selli Level, it is proposed as a marker species for the Aptian sediments of the Tethys.The dinoflagellate cysts Kallosphaeridium dolomiticum sp. nov. and Nexosispinum hesperus brevispinosum subsp. nov. are described from the Upper Hauterivian. Additional taxonomic remarks are made about other dinoflagellate cyst species, including the emendations of Tanyosphaeridium magneticum Davies 1983 and Bourkidinium granulatum Morgan 1975.The biostratigraphic value of selected taxa is discussed and compared with data known both from the Tethyan and Boreal realms. In particular, the extinction of Bourkidinium granulatum emend. is proposed as the best dinoflagellate cyst event for the delimitation of the Hauterivian-Barremian boundary in the Northern Hemisphere. The first appearance datums of Prolixosphaeridium parvispinum and Odontochitina operculata, and the slightly younger last appearance datum of Nexosispinum vetusculum are confirmed as useful biohorizons for recognition of the lower part of the Upper Barremian and hence for the approximation of the Lower-Upper Barremian boundary. The last occurrences of Rhynchodiniopsis aptiana and Phoberocysta neocomica are calibrated in the basal Aptian. PMID- 10704646 TI - Effects of extracorporeal circulation and heparin on the phenotype of platelet surface antigens following heart surgery. AB - In this prospective study, the time-dependent effects of extracorporeal circulation and heparin-mediated effects on platelet surface antigens in vitro were investigated using whole blood flow cytometry. Blood samples were drawn prior to and following extracorporeal circulation in 89 patients. The response of surface antigen expression (glycoprotein IIb/IIIa, P-selectin, and glycoprotein Ib) with and without in vitro stimulation was measured. A significant correlation of the duration of extracorporeal circulation with the postoperative response of glycoprotein IIb/IIIa, glycoprotein Ib, and P-selectin to in vitro activation was found. Postoperative P-selectin and glycoprotein Ib expression stimulated with ADP correlated to blood loss. Heparin in vitro significantly reduced glycoprotein Ib expression. Heparin, as well as the duration of extracorporeal circulation, independently correlated to phenotypic changes of platelets following extracorporeal circulation. The significant correlation of these variables to postoperative blood loss demonstrates their relevance to platelet function in vivo. PMID- 10704647 TI - A prospective study on the incidence and clinical relevance of heparin-induced antibodies in patients after vascular surgery. AB - The heparin-platelet factor 4-antibody assay, polyanion-platelet factor 4 antibody assay and heparin-induced platelet activation test are used for laboratory diagnosis of the immune form of heparin-induced thrombocytopenia. Fifty consecutive patients receiving heparin treatment for more than 5 days after vascular surgery were prospectively screened for heparin-induced thrombocytopenia antibodies, thrombocytopenia (daily platelet counts), deep-vein thrombosis (color coded duplex sonography), and arterial reocclusion (clinical assessment). None of the patients developed thrombocytopenia or thrombosis in association with formation of heparin-induced thrombocytopenia antibodies. Despite the absence of clinical evidence of heparin-induced thrombocytopenia, many patients formed heparin-induced thrombocytopenia antibodies: 34% of the patients were positive in the heparin-platelet factor 4-antibody assay, 28% in the polyanion-platelet factor 4-antibody assay, 14% in the heparin-induced platelet activation test, and 54% with any of these tests. Patients predominantly developed IgM (24%) and IgA antibodies (16%), whereas IgG antibodies were found in 12% of patients. Whereas the majority of patients with positive ELISA assays had IgM and IgA antibodies, patients with a positive functional assay (heparin-induced platelet activation test) predominantly had IgG antibodies. We conclude that a high percentage of patients develop heparin-induced antibodies after vascular surgery without any clinical symptoms of heparin-induced thrombocytopenia. None of the assays therefore is predictive of the clinical manifestation of heparin-induced thrombocytopenia in asymptomatic patients. Therefore, the diagnostic specificity of both antigen and activation assays for heparin-induced thrombocytopenia appears to be relatively low in the vascular surgery patient population. PMID- 10704648 TI - Optimal dosing of subcutaneous unfractionated heparin for the treatment of deep vein thrombosis. AB - Twice-daily, inpatient, subcutaneous unfractionated heparin is at least as effective and safe as continuous intravenous unfractionated heparin for the treatment of acute deep vein thrombosis. Subcutaneous unfractionated heparin therefore may be suitable for outpatient treatment of deep vein thrombosis. The purpose of this study was to develop a dosing nomogram for a dose each 12 hours (2 doses per day) 12-hourly subcutaneous unfractionated heparin that is suitable for outpatient treatment of acute deep vein thrombosis. A cohort study was performed in patients with acute deep vein thrombosis in two phases. In both phases, the first subcutaneous loading dose of unfractionated heparin was 317 U/kg, and the second dose was 231 U/kg. The activated partial thromboplastin time was measured daily, 6 hours after the morning injection, and subsequent doses of unfractionated heparin were adjusted according to a nomogram, which was modified for phase II. Warfarin was started with unfractionated heparin. In phase I (14 outpatients), activated partial thromboplastin time results were frequently subtherapeutic (9:14) the day after starting unfractionated heparin (day 1), and were frequently supratherapeutic (27:40) after the first 2 days of unfractionated heparin therapy. In phase II (21 patients), to explain the frequently subtherapeutic activated partial thromboplastin time results that were obtained on day 1, the activated partial thromboplastin time results were measured after the initial loading dose. Mean activated partial thromboplastin time results of 86 and 61 seconds were obtained 6 and 12 hours after this dose, suggesting that 317 U/kg is a suitable subcutaneous loading dose. In contrast to phase I, in phase II, unfractionated heparin dose was not increased on day 1 in response to a low activated partial thromboplastin time result. This reduced the frequency of supratherapeutic activated partial thromboplastin time results during the early days of therapy without increasing the frequency of subtherapeutic results. Warfarin therapy had a substantal effect on the activated partial thromboplastin time that appeared to account for a high frequency of supratherapeutic results during the later days of unfractionated heparin therapy; the activated partial thromboplastin time increased by 20 seconds (95% CI, 14-27 seconds) with each increase in the International Normalized Ratio of 1.0. We had limited success at developing a dosing nomogram for subcutaneous unfractionated heparin that reliably achieved activated partial thromboplastin time results within the therapeutic range. However, as oral anticoagulants directly increased activated partial thromboplastin time results, we suggest that adjusting unfractionated heparin dose to achieve prespecified activated partial thromboplastin time results may not be appropriate in patients who are receiving concomitant warfarin therapy. PMID- 10704649 TI - Fibrinogen fractions in the third trimester of pregnancy and in puerperium. AB - The concentration of fibrinogen, its fractions, and the concentration of C reactive protein were determined in 45 healthy pregnant women before and after vaginal (27) or caesarean section 18 delivery. The control group consisted of 33 blood donors. In pregnancy, increased concentration of total fibrinogen and its fractions and a normal concentration of C-reactive protein were noted. Three days after vaginal delivery the concentration of total and high molecular weight fibrinogen fraction decreased slightly and the ratio of high to low molecular weight fibrinogen increased. After caesarean section both total and high molecular weight fibrinogen and the ratio of high to low molecular weight fibrinogen increased. The C-reactive protein concentration increased after either type of delivery. The degree of augmentation, however, was ten times as strong after caesarean section. In all women at the end of puerperium the concentrations of the compounds studied returned to normal values. The results suggest that the mechanisms leading to the increase of fibrinogen during pregnancy and after delivery are different. The increase of all fibrinogen fractions in pregnancy may depend first of all on hormones, whereas the increased proportion of high molecular weight to low molecular weight fibrinogen after delivery depends on the acute phase reaction. The degree of this reaction depends on the type of delivery. PMID- 10704651 TI - Investigation of the relocation of cytosolic phospholipase A2 and annexin V in activated platelets. AB - Cytosolic phospholipase A(2) is a Ca(2+)-dependent enzyme that acts on membrane phospholipids to release arachidonic acid, which in platelets is converted to thromboxane A(2). Annexin V is a Ca(2+)-dependent, phospholipid-binding protein, which is proposed to regulate inflammation by inhibiting cytosolic phospholipase A(2). Here, we have studied the association of cytosolic phospholipase A(2) and annexin V with platelet membranes after thrombin stimulation. In a time-dependent manner, an exact correlation was found between the membrane association of cytosolic phospholipase A(2) and annexin V. Calcium from the intracellular stores was sufficient for the relocation of intracellular annexin V and cytosolic phospholipase A(2) to platelet membranes. Activation in the presence of arginyl glycyl-aspartyl-serine (RGDS), which inhibits binding of fibrinogen to its adhesive ligand, does not alter the amount of cytosolic phospholipase A(2) or annexin V that binds to membranes. When activation-induced actin polymerisation was prevented by cytochalasin E, the recovery of both annexin V and cytosolic phospholipase A(2) remained unchanged. However, complete depolymerisation of the cytoskeleton with DNase I almost abolished the association of cytosolic phospholipase A(2) with the membranes, and it completely abolished the relocation of annexin V to platelet membranes. Finally, we show that cytosolic phospholipase A(2) can be specifically purified from platelet membranes by affinity chromatography on GST-annexin V and that immunoprecipitation using antibodies against cytosolic phospholipase A(2) copurify annexin V and cytosolic phospholipase A(2) from activated platelets. These findings suggest that following platelet activation with thrombin, both cytosolic phospholipase A(2) and annexin V, relocate to platelet membranes where they interact. An intact cytoskeleton seems to be a prerequisite for the interaction of cytosolic phospholipase A(2) and annexin V with platelet membranes. The incorporation of cytosolic phospholipase A(2) into the membrane fraction of thrombin-activated platelets parallels that of annexin V, which suggests an interaction between the two proteins. PMID- 10704650 TI - Successful eradication of Helicobacter pylori as determined by ((13))C-urea breath test does not alter fibrinogen and acute phase response markers. AB - In this study we examined in 100 patients testing positive for Helicobacter pylori infection whether successful eradication therapy with pantoprazole, clarithromycin, and metronidazole alters fibrinogen and other acute phase response markers. Of 100 patients, only 11 showed a fibrinogen level above 300 mg/dL. Successful eradication proven by the 13C-urea breath test does not alter acute phase response markers. These findings indicate that Helicobacter pylori infection is unlikely to affect atherosclerosis unfavourably via acute phase response. PMID- 10704652 TI - A new approach to detect reticulated platelets stained with thiazole orange in thrombocytopenic patients. AB - Recent studies have shown that reticulated platelets stained with Thiazole Orange (T.O.) are useful markers for thrombopoiesis. The percentage of T.O. positive platelets tends to be inconsistent using the original method, especially when the peripheral blood platelet count is very low. We measured T.O. positive platelet levels in patients with severe thrombocytopenic disorders, using concentrated platelet-rich plasma and carrying out a two-color analysis involving T.O. and an anti-glycoprotein IIb/IIIa monoclonal antibody. This method allowed us to obtain consistent T.O. positive platelet rates in patients with thrombocytopenia whose platelet counts were below 20 approximately 30x10(9)/l. By this method, the T.O. positive rates of platelets from idiopathic thrombocytopenic purpura patients were found to be significantly higher than in the control group. The T.O. positive rates of other thrombocytopenic disorders were similar to those of the control group. These results are consistent with those previously reported. We conclude that our technique of measuring of T.O. positive platelets using platelet-rich plasma is useful for analyzing severe thrombocytopenic disorders. PMID- 10704653 TI - Stimulating effect of biologically modified low density lipoproteins on ADP induced aggregation of washed platelets persists in absence of specific binding. AB - Oxidized low density lipoproteins are closely associated with atherosclerosis and also might be directly involved in thrombosis because they have been shown to mediate a stimulating effect on human platelets. In this work, we used biologically modified low density lipoproteins (i.e., low density lipoproteins sufficiently oxidized to show specificity for the macrophage scavenger receptor system) to examine if specific binding of the oxidized apolipoprotein moiety to the platelet surface is a prerequisite for the platelet-stimulating effects reported by other authors. We find that biologically modified low density lipoproteins show specific binding to human platelets (K(d)=5.83+/-0.4 microg/mL, 3850+/-620 sites/platelet) and strongly augment both ADP- and thrombin-induced aggregation of washed platelets. Maleylated albumin, an antagonist of oxidized low density lipoproteins binding to all currently classified scavenger receptors, is able to reduce platelet oxidized low density lipoproteins binding to background levels. Nevertheless, maleylated albumin is not able to exert any kind of normalizing effect on the augmented ADP-induced aggregation response observed in the presence of biologically modified low density lipoproteins. From these data, we conclude that specific binding of oxidatively modified apolipoprotein B to the platelet surface is not essential to the process of platelet stimulation. Therefore, we conclude that these stimulating effects may be mediated by unidentified compounds formed in the lipid phase of the lipoproteins. PMID- 10704654 TI - Effect of cryopreservation on endothelin-1 productions of human mammary artery. AB - The aim of this study was to determine the ability of human mammary artery cells to maintain a metabolic activity by measuring the artery concentration of two vasoactive substances, endothelin-1 (ET-1) and cyclic guanosyl monophosphate (cGMP), and a neurohumoral substance-neuropeptide Y (NPY)-prior to and following cryostorage. Ten distal segments of internal mammary arteries were obtained at the time of surgery in patients who had undergone coronary artery bypass grafting. One ring of each vessel served as a fresh control for the other ring that was used in cryopreservation experiments. The arteries were frozen with liquid nitrogen at a controlled rate down to -130 degrees C with an automatic freezing machine and were then stored in a liquid nitrogen vapor at -150 degrees C. After mammary artery extraction, ET-1, cGMP, and NPY concentrations were studied before and after cryopreservation. Cryopreserved, compared to fresh arteries, exhibited an increase in ET-1 (11.11+/-1.61 vs. 3. 09+/-0.06 pg/mg; p=0.004) and a decrease in cGMP (9.88+/-2.04 vs. 8. 55+/-2.07 p moles/mg; p<0.02), whereas there was no significant NPY variation. An increase in ET-1 and decrease in cGMP was found in 10 out of 10 and 6 out of 10 of cryopreserved artery specimen, respectively. There was no significant correlation between ET-1 and cGMP production in fresh or in cryopreserved arteries. The present method of cryostorage is effective in preserving "hormonal" mammary artery activity. However, the particularly high ET-1 concentration without associated cGMP concentration may be deleterious by increasing smooth-muscle cell proliferation and vascular tone of cryopreserved arteries. PMID- 10704655 TI - A rapid method to visualize von willebrand factor multimers by using agarose gel electrophoresis, immunolocalization and luminographic detection. AB - A highly sensitive and rapid clinical method for the visualization of the multimeric structure of von Willebrand Factor in plasma and platelets is described. The method utilizes submerged horizontal agarose gel electrophoresis, followed by transfer of the von Willebrand Factor onto a polyvinylidine fluoride membrane, and immunolocalization and luminographic visualization of the von Willebrand Factor multimeric pattern. This method distinguishes type 1 from types 2A and 2B von Willebrand disease, allowing timely evaluation and classification of von Willebrand Factor in patient plasma. It also allows visualization of the unusually high molecular weight multimers present in platelets. There are several major advantages to this method including rapid processing, simplicity of gel preparation, high sensitivity to low concentrations of von Willebrand Factor, and elimination of radioactivity. PMID- 10704656 TI - A new sensitive chromogenic substrate assay of tissue factor pathway inhibitor type 1. AB - The present assay is a modification of our previously published two-stage chromogenic substrate assay of tissue factor pathway inhibitor type-1 (TFPI) activity [1]. In the first stage, the reaction mixture was made with factor VIIa (FVIIa) molecules in excess of tissue factor (TF) binding sites and contained diluted plasma, recombinant FVIIa (10 nM), recombinant TF (1/400 vol/vol), bovine factor Xa (1,1 nM), I-2882(R) (100 microg/ml), and CaCl(2) (10 mM). The fibrin polymerisation inhibitor I-2882(R) was added to the reaction mixture to prevent formation of cross-linked fibrin. In the second stage, residual TF/FVIIa catalytic activity was measured by the addition of a substrate mixture that contained bovine factor X and a chromogenic substrate (S-2222(R)). Standard curves were constructed using serial dilutions (0-1%) of pooled normal plasma. The dose-response relationship for serial dilutions of plasma was linear. The intra-assay coefficient of variations (CVs) for pre- and postheparin plasma samples (i.e., normal and high TFPI levels) were 1.7% and 9.9%, respectively; the inter-assay CVs were 10.0% and 19. 7%, respectively. The effect of variation in antithrombin activity on the assay was approximately 5%. The present assay correlated fairly well with our previously published assay (r=0.82, n=100) and with a commercial TFPI activity assay (Actichrome(R) TFPI Activity Assay, American Diagnostica, Greenwich, CT, USA; r=0.90, n=100), as well as with an antigen assay for TFPI total antigen (Imubind(R), American Diagnostica; r=0,96, n=100). Altman and Bland plots revealed that our previous assay underestimated TFPI activity at high TFPI levels (i.e., postheparin TFPI samples) compared with the other methods. PMID- 10704657 TI - Singlet oxygen inactivates fibrinogen, factor V, factor VIII, factor X, and platelet aggregation of human blood. AB - Activated polymorphonuclear leukocytes participate in hemostasis. These phagocytes generate up to 5 mmol/l of oxidants of the HOCl- and chloramine-type. The present study shows, for the first time, that physiological concentrations of NaOCl or chloramines act as anticoagulants in human plasma. Prothrombin time, activated partial thromboplastin time, and thrombin time at chloramine concentrations greater than 1 mmol/l are prolonged proportional to the oxidant concentration. Plasmatic coagulation factors sensible to oxidation are fibrinogen, factor V, factor VIII, and factor X with a 50% effective dose of 2-3 mmol/l NaOCl or taurine-chloramine. Chloramines or chloramine-like agents (e.g., chloramine T(R) or vancomycin) also inactivate platelet aggregation (in whole blood or platelet-rich plasma) at an 50% effective dose of about 1.0 mmol. This irreversible oxidation of the hemostasis components is inhibited by addition of methionine, cysteine, ascorbic acid, or azide in 10-fold molar excess prior to oxidation. The oxy-radical inhibitors mannitol, superoxide dismutase, or catalase do not antagonize the action of NaOCl or chloramines. Therefore, the oxidant here involved has reaction characteristics of singlet oxygen (1O(2)), a nonradical, excited (i.e., light-emitting) oxidant. The hemostasis factors sensible to oxidation might dispose of oxidizable, for their function critical, methionine or cysteine residues. In conclusion, blood coagulation factors I, V, VIII, X and thrombocytes are sensible to nonradical oxidants of activated phagocytes. Via 1O(2) generation, polymorphonuclear leukocytes can generate a local pericellular zone of anticoagulation. The data suggest that the cell signal 1O(2) in physiological amounts is an antithrombotic agent. PMID- 10704658 TI - Whale high-molecular-weight and low-molecular-weight kininogens. AB - The expression of high-molecular-weight and low-molecular-weight kininogen mRNAs in the whale liver was examined by reverse transcription-polymerase chain reaction. The nucleotide sequences of the high-molecular-weight and low-molecular weight kininogen cDNAs were analyzed and deduced to the amino acid sequences. The high-molecular-weight kininogen composed of 609 amino acid residues with 18 signal peptides possessed the consensus sequences of the cysteine protease inhibitor domains I and II, the bradykinin domain, the histidine-rich region, and the prekallikrein-binding region. Except for the histidine-rich region, the overall homologies with bovine, human, and rat high-molecular-weight kininogens were 81%, 76%, and 62%, respectively. The low-molecular-weight kininogen is composed of 408 amino acid residues. The nucleotide sequence down to C(1200) as well as the amino acid sequence till Ile(382) is identical to that of the high molecular-weight kininogen. The remaining low-molecular-weight kininogen-specific carboxy-terminal portion possessed an amino acid sequence similar to that of the land mammals. The overall homologies with bovine, human, and rat low-molecular weight kininogens were 82%, 79%, and 64%, respectively. The amino acid sequences of both whale high-molecular-weight and low-molecular-weight kininogens are most similar to those of the bovine among the land mammals analyzed so far. An incubation of dolphin/whale plasma with human plasma kallikrein, or with bovine trypsin, in the presence of carboxypeptidase inhibitors generated bradykinin antigen as well as the spasmogenic activity to the estrous rat uterus. The amount of bradykinin released by the latter enzyme was almost double of the former, indicating that the dolphin/whale plasma contained similar concentrations of low molecular-weight and high-molecular-weight kininogens. PMID- 10704659 TI - LDL-Apheresis removes serum amyloid P and A in hypercholesterolemic patients. PMID- 10704660 TI - Comparison between routine laboratory prothrombin time measurements and fingerstick determinations using a near-patient testing device (Pro-Time). PMID- 10704661 TI - Thrombelastography in healthy volunteers, patients with stable angina and acute chest pain. PMID- 10704663 TI - Effect of temperature and incubation time on D-dimer serum levels in healthy subjects. PMID- 10704662 TI - Receptor expression for IgG constant fraction in human umbilical vein endothelial cells. PMID- 10704664 TI - Plasma concentrations of von Willebrand factor in patients with angina pectoris secondary to coronary atherosclerosis or cardiac syndrome X. PMID- 10704665 TI - Digital analysis of light microscope immunofluorescence: high-resolution co localization of synaptic proteins in cultured neurons. AB - A protocol is presented for determining the subcellular distribution of fluorescently labeled proteins in neurons using deconvolved images gathered with a wide-field microscope. The protocol includes optimal settings for the numerical algorithm used to deconvolve the images and an objective method for thresholding the deconvolved images to retain only high-intensity, specific labeling. The effectiveness of the protocol is demonstrated using a fluorescent antibody stain directed towards the alpha1 subunit of the GABA(A) receptor in cultured neurons. We also show, using an antibody against the presynaptic vesicular protein synaptophysin, that the technique can detect presumptive regions of synaptic contact between neurons. Double-labeling with the anti-alpha1 and anti synaptophysin antibodies in a cultured neuron reveals regions of both synaptic and non-synaptic alpha1 labeling. Thus, numerical postprocessing of wide-field images can be used to efficiently locate receptor proteins in neurons in relation to functionally important structures. This confocal-like functionality is attained without the excessive bleaching and phototoxicity associated with the intense laser excitation light used in confocal techniques. PMID- 10704666 TI - Computer simulation approach to the quantification of immunogold labelling on plasma membrane of cultured neurons. AB - Cell culture is a convenient model system to study the expression of plasma membrane-bound proteins in nerve cells. Analysing it with an ultrastructural detail researchers often apply transmission electron microscopy together with immunogold labelling. Plasma membrane profiles are one-dimensional (1D) and provide little information about the topography of membrane-bound proteins. In order to convert 1D estimates of spatial arrangement for preembedding immunogold labelled proteins into two-dimensional (2D) quantities, namely the 2D pattern and density of labelling, this paper presents a simple computer simulation technique. This technique is based on a mathematical model permitting a simulated immunogold labelled membrane to be sampled in a way similar to microtome sectioning. An interlabel distance (ILD) estimate is used to define the position of immunogold particles in membrane profiles. In order to interpret experimental ILD measurements the simulated distribution best fit to the experimental data is selected and the corresponding 2D density and pattern of particle scattering are considered to explain the real situation. Various parameters including a cell section thickness, immunogold particle size etc can be adjusted to suit the demands of a particular experiment. The technique was applied to quantify the NCAM preembedding immunogold labelling in the plasma membrane of cultured rat hippocampal neurons. PMID- 10704667 TI - Introduction of macromolecules into synaptosomes using electroporation. AB - Synaptic terminals are sites of high metabolic activity and thus are particularly vulnerable to oxidative stress. Oxidative damage to proteins can be toxic to neurons and may cause irreversible cell damage and neurodegeneration. A neuroprotective mechanism used by cells to combat oxidative damage is to selectively degrade damaged proteins. Therefore, it is of interest to study the mechanism of degradation of oxidatively damaged proteins in synaptosomes. One way of oxidizing synaptosomal proteins in vitro is by incubating intact synaptosomes in the presence of an oxidizing agent. A problem with this approach is that it may also cause oxidative damage to the machinery required to recognize and degrade oxidized proteins. We have, therefore, introduced a fluorescent macromolecule into synaptosomes to assess the feasibility of using this technique to study how oxidized proteins are degraded and removed from synaptic terminals. Synaptosomes were subjected to electroporation in the presence of FITC labelled dextran with an average molecular weight of 70000 (FD-70) and non-specific binding was determined by running parallel experiments in lysed synaptosomes. Following extensive washing, synaptosomes were assayed for the presence of intra synaptosomal FD-70 by measuring fluorescence in a microplate fluorescence reader. Significant differences in fluorescence were found between intact and lysed synaptosomes with maximal uptake at 100 V/ 1500 microF (approx. 36 pmol/mg protein). To determine if membrane transport was compromised by electroporation, uptake of 3H-arginine was compared in control and electroporated synaptosomes. While untreated electroporated synaptosomes showed a loss of 22% in the ability to transport arginine, preincubation in the presence of 1 mM ATP resulted in a complete restoration of arginine transport. These results show that electroporation is a potentially useful technique for introducing a specific oxidized protein, into synaptic terminals so its metabolic fate can be examined. PMID- 10704668 TI - Recording ionic events from cultured, DiI-labelled myenteric neurons in the guinea-pig proximal colon. AB - To date investigations of enteric neurons by patch clamping/calcium imaging have been limited by studying unidentified heterogeneous populations of neurons. In DiI-labelled colonic myenteric neurons, the feasibility of recording ionic events was determined by applying DiI either to the mucosa or the circular muscle, dispersing neurons after 48 h organotypic culture, and patch-clamping/calcium imaging labeled neurons after 3-7 days in culture. Myenteric neurons with diffuse DiI fluorescence were typically smooth and agranular. Neurons labeled after DiI was applied to circular muscle, fired in either a phasic or a tonic manner, and exhibited fast afterhyperpolarizations (100-300 ms duration) at the end of a depolarizing pulse. They expressed a fast inward current and at least three different outward currents. Action potentials elicited in DiI-labeled sensory neurons were followed by a prolonged afterhyperpolarization (AH, 4-6 s). The offset of a suprathreshold depolarizing step elicited a prolonged outward tail current that approximated the timecourse of the prolonged AH. In addition, in response to membrane depolarization in DiI-labeled neurons loaded with fura-2, robust Ca(2+) transients were recorded using the perforated patch technique. These results demonstrate that DiI labeling of cultured myenteric neurons is feasible, and patch clamp/Ca(2+) fluorescence recordings can be made from specific populations of cultured DiI-labeled colonic myenteric neurons. PMID- 10704670 TI - Intrafascicular electrodes for stimulation and recording from mudpuppy spinal roots. AB - This paper presents a technique for stimulating and recording from multiple intact spinal roots in the in vitro mudpuppy (Necturus maculatus) spinal cord forearm preparation using fine wire electrodes, a modified intrafascicular electrode. We found that multiple spinal roots of the preparation could be implanted with these modified electrodes for independent stimulation or recording of the roots without inducing mechanical vibrations, disrupting conduction, or obscuring the view of or access to the spinal cord. Recording and stimulation performance using these electrodes was compared with results obtained using conventional hook electrodes. We found that intrafascicular electrodes were more efficient than hook electrodes for stimulating nerve fibers, being able to produce equivalent levels of activation using stimulation levels that were an order of magnitude smaller. Compound action potential signals recorded from electrodes implanted in the spinal roots were found to be larger than those from hook electrodes placed around the corresponding spinal nerve, showing that intrafascicular electrodes are more efficient at recording activity in the nerve. Moreover, it was possible to record evoked activity from cutaneous mechanoreceptors, even though the signal to noise ratio was low. Rough estimates of the conduction velocities for the fastest components in the compound action potentials were calculated and found to be around 17.5 m/s for both dorsal and ventral roots. PMID- 10704669 TI - Fluorescence microscopy of stimulated Zn(II) release from organotypic cultures of mammalian hippocampus using a carbonic anhydrase-based biosensor system. AB - We demonstrate here that electrical stimulation of organotypic cultures of rat hippocampus results in the prompt release of significant amounts of Zn(II) by a fluorescence microscopic method. The fluorescence imaging of free Zn(II) is achieved using a highly selective biosensing indicator system consisting of human apo-carbonic anhydrase II (apoCAII) and a fluorescent aryl sulfonamide inhibitor of the enzyme, ABD-N. The apoenzyme and ABD-N in the absence of Zn(II) exhibit weak, reddish fluorescence typical of the ABD-N alone; when Zn(II) is added it binds to the apoenzyme (K(D) = 4 pM), which strongly promotes binding of ABD-N to the holoenzyme (K(D) = 0.9 microM). Binding of ABD-N to the holoenzyme results in a 9-fold increase in apparent quantum yield, significant blue shifts in excitation and emission, an increase in average fluorescence lifetime, a 4-fold increase in the ratio of intensities at 560 and 680 nm, and a large increase in anisotropy. Prior to stimulation, cultures immersed in phosphate-buffered saline with glucose and apoCAII with ABD-N emitted negligible fluorescence, but within 20 s after electrical stimulation a diffuse cloud of greenish fluorescence emerged and subsequently covered most of the culture, indicating release of zinc into the extracellular medium. PMID- 10704671 TI - RT-PCR amplification of mRNA from single brain neurospheres. AB - A method is described that allows cDNA production from individual brain cell clones or 'neurospheres'. These culture-generated spheres of stem, progenitor, and differentiated cells have been the focus of interest because they represent an in vitro model of neurogenesis. However, because neurospheres are somewhat resistant, in part due to their enclosure by a dense extracellular matrix, to methods attempting to disrupt them and isolate nucleic acids, there is a need for new technology that affords the simple and efficient RT-PCR for studies of neural gene expression and discovery. A method is described here that uses sonication and an all-in-one approach for the construction of cDNA from single neurospheres. The generation of cDNA from individual adult brain stem/progenitor cell neurospheres is useful for future studies of neurogenic gene expression. PMID- 10704672 TI - Multiple components in the agonist concentration-response relationships of neuronal nicotinic acetylcholine receptors. AB - Assessing the potential of nicotinic agonists as therapeutic agents has frequently relied upon single component EC(50) values obtained from studies of nicotinic receptors expressed in Xenopus oocytes. We have evaluated the validity of this approach using voltage clamp techniques. In general, agonist concentration-response plots for the alpha3beta2, alpha3beta4, alpha4-1beta2, alpha4-1beta4 and alpha7 combinations were poorly fitted by a single component Hill-equation. Improved fits were obtained with the sum of two components, although only in the case of alpha3beta4 and alpha4-1beta2 was the improvement significant regardless of the weighting method used. For the acetylcholine (ACh) concentration-response relationships of the alpha4-1beta2 combination, the two EC(50) values were 0.3 and 58.3 microM. For the alpha3beta4 combination, the two EC(50) components were 39 and 2919 microM. The 39 microM component of alpha3beta4 represented 36% of the sum of the maximum responses of both curves. This shows that for some combinations, the secondary components represent a well-separated, major population of receptors. Therefore, published EC(50) values which assume that only a single subtype of functional receptor is present may not accurately describe agonist action may therefore need to be re-evaluated. PMID- 10704673 TI - Comparison of eight clinical rating scales used for the assessment of MPTP induced parkinsonism in the Macaque monkey. AB - The most valuable model of Parkinson's disease available at present is the primate model treated with 1-methyl-4-phenyl-1,2,3, 6-tetrahydropyridine (MPTP), frequently used to study response to new drugs or surgical treatments. The evaluation of such therapies requires clinical rating scales which measure precisely motor behaviour in both normal and parkinsonian monkeys. It is obvious that such evaluation can only be valid if parallel studies are carried out under similar experimental conditions with well-defined objective criteria. Hence the need to compare and assess the different rating scales in use if we want to be able to compare the results of clinical studies. In order to give rise to some fresh thinking on the necessity of a certain uniformity of assessment, this study compares eight clinical rating scales and considers their capacity to express in quantitative terms both the severity of MPTP intoxication in five cynomolgus monkeys and the alleviation afforded by levodopa. None of the eight scales reaches all the criteria despite the Kurlan scale would appear as an interesting working basis for a further consensual definition of a worldwide used parkinsonian monkey clinical rating scale PMID- 10704674 TI - A method for activating neurons using endogenous synaptic waveforms. AB - Neuronal input-output functions are traditionally studied using rectangular or ramp waveforms of injected current. These waveforms are easy to produce and responses to them easy to quantify; thus they have been central to our understanding of the roles that membrane properties play in controlling repetitive firing. However, since smooth rectangular step and ramp waveforms lack the dynamic features of endogenous synaptic input, their use has the potential to underemphasize the importance of input patterns in controlling physiological patterns of neuronal output. To activate neurons with current waveforms that replicate natural synaptic input, we developed a method for acquiring, digitally manipulating and reinjecting endogenous synaptic currents. We demonstrate, by applying this technique to phrenic motoneurons (PMNs) in rhythmically-active in vitro preparations from neonatal rats, that stimulation of neurons with endogenous current waveforms produces responses that mimic those produced by spontaneous synaptic inputs. Acquired waveforms can be reinjected repeatedly to produce consistent responses, and can also be amplified or filtered prior to reinjection to yield a range of information including standard descriptors of firing behavior such as frequency/current plots. This technique provides a valuable tool for analysing characteristics of the synaptic waveform important in generating neuronal output and how synaptic factors interact with membrane properties to control repetitive firing. PMID- 10704676 TI - Interphase detection of t(4;14)(p16.3;q32.3) by in situ hybridization and FGFR3 overexpression in plasma cell malignancies. AB - The immunoglobulin (Ig) genes are frequently involved in chromosomal rearrangements with a wide variety of partner loci in multiple myeloma (MM). However, several partner chromosomes have not been detected by conventional cytogenetic methods; for example, 4p16.3 (FGFR3), 6p25.3 (IRF4), and 16q23 (c maf). To clarify the incidence of t(4;14)(p16.3;q32.3) in primary tumors of MM and to evaluate possible correlations with specific manifestations of the disease, G-banding, double-color fluorescence in situ hybridization (DC-FISH), and/or reverse-transcriptase polymerase chain reaction (RT-PCR) were performed on 40 patients with MM-two with plasmacytoma (PCM) and three with plasma cell leukemia (PCL). All patients were studied by DC-FISH; 40 were studied by G banding and 36 were studied by RT-PCR. The FISH probes consisted of a cosmid pC385.12 containing the FGFR3 gene, a YAC Y6 containing VH, and a phage Iggamma1 10 containing the gamma1 constant region (Cgamma). We identified eight patients with either FGFR3/Cgamma fusion or FGFR3 overexpression: six patients with both FGFR3/Cgamma fusion and FGFR3 overexpression, one patient with FGFR3/Cgamma, and one with FGFR3 overexpression. FGFR3/Cgamma fusion was demonstrated at a frequency of 19% to 38% on interphase nuclei in seven of the 45 patients. Lytic bone lesions were found to be associated with FGFR3 overexpression. Interphase FISH with FGFR3 and Cgamma probes combined with RT-PCR proved to be an effective tool for detection of this fully cryptic translocation, thus facilitating the characterization of clinical features of MM patients with t(4;14). PMID- 10704677 TI - Genetic alterations of gastric cancer: comparative genomic hybridization and fluorescence In situ hybridization studies. AB - Genetic changes leading to the development of gastric cancers are still in dispute. In the following study, we used comparative genomic hybridization (CGH) to screen for DNA copy number changes along all chromosomes in 37 gastric carcinomas, and fluorescence in situ hybridization (FISH) with the C-MYC and TP53 probes in 14 cases for comparison. The aim of this study was to identify those chromosome regions that contain genes important for the development of gastric carcinomas and to identify genetic markers associated with tumor progression. The most often involved gains were 2q, 7pq, 8pq, 13q, 17q, 18q, and 20pq. The most commonly deleted regions were 17p. The pattern of genetic changes was different depending on the existence of nodal metastasis and histologic types. Gains in 8q and losses in 17p were the most common features of the CGH changes. However, only 3 among the available 10 cases (30%) showed an amplification of the C-MYC gene by FISH. Allelic loss of TP53 was found in 2 of 4 cases (50%). This difference might be due to another rearrangement of these 2 genes which cannot be detected by FISH, or other possible genes in that area may be involved in the tumorigenesis and nodal metastasis of gastric carcinomas. PMID- 10704678 TI - Complex karyotype and N-RAS point mutation in a case of acute megakaryoblastic leukemia (M7) following a myelodysplastic syndrome. AB - The development of acute megakaryoblastic leukemia (ANLL-M7) following myelodysplastic syndrome (MDS) has been described only in a few reports, and the mutations necessary for this transformation are still unknown. In this study, we describe a case of ANLL-M7 with a previous history of MDS presenting a complex karyotype 46,XX, t(4;11)(q21;q23),del(5)(q13q33),t(12;13)(p13;q21) and N-RAS point mutation. During MDS, the patient showed a hypercellular myelogram with dysplasia of the three myeloid lineages and the clinical symptoms characteristic of the 5q- syndrome. During the follow-up, we observed the appearance of two additional subclones, one with an isochromosome 17q and another with polyploidy. The presence of an isochromosome 17q in one subclone and polyploidy in another is probably due to the genetic instability generated by the malignant transformation. PMID- 10704679 TI - Cytogenetic analysis of esophageal squamous cell carcinoma cell lines by comparative genomic hybridization: relationship of cytogenetic aberrations to in vitro cell growth. AB - Cancer is characterized by autonomous growth of cells, and it is widely accepted that cell proliferation is primarily influenced by individual cell genetics. To elucidate the mechanisms of cancer cell proliferation, we studied differences in genetic aberrations for different type of tumors with different proliferation characteristics. We employed comparative genomic hybridization (CGH) to detect genetic aberrations in six cell lines of esophageal squamous cell carcinoma (ESCC). Three cell lines (YES-1, -2, and -3) grow in culture without fetal calf serum (group A), while others require serum to be maintained in vitro (group B). Both groups showed very similar cytogenetic aberrations: over-representations of 11q13 (6/6), 8q23-qter (5/6), Xq25-qter (5/6), 3q26-qter (4/6), 5p (4/6), 7p15 pter (4/6), 8q21.3-q22 (4/6), 17p (4/6), and 20q13 (4/6), and under representations of 18q21-qter (6/6), 4q28-q33 (4/6), and 9p21 (4/6). Six amplification loci were mapped to chromosomal regions of 6q23 (1 case), 7p12 (2 cases), 9p21 (1 case), 11p11.2-12 (3 cases), 11q13 (2 cases), and 17p12 (2 cases). However, some differences were detected. DNA copy number increases at 7p12-p13, 11q14-q22, and 11q22-qter and under-representations of 4p, 8p, and 11p14-pter. In contrast, gains at 12p and 20p, and losses at 3p and 5q were detected only in group-B cell lines. These observations suggest that cytogenetic differences between the two groups may be linked to differences in cell growth characteristics in vitro, and that the genes in these chromosomal regions may play important roles in cell proliferation. PMID- 10704680 TI - Primary macroglobulinemia with t(11;18)(q21;q21). AB - These are the first cases of primary macroglobulinemia (PMG) with t(11;18)(q21;q21) reported in the literature. The first case was a 77-year-old man with macroglobulinemia (serum IgM: 8.36 g/dL). Abnormal lymphoid cells were detected in the blood and bone marrow. Immunologic and karyotypic analyses revealed that abnormal cells were positive for surface IgM-k, CD19, and CD20, negative for CD5 and CD10, and all had a t(11;18)(q21;q21). The second case was a 57-year-old woman with macroglobulinemia (serum IgM: 12.0 g/dL). Abnormal lymphoid cells were detected in blood and marrow, and cells were positive for surface IgM-lambda, CD19, and CD20, and negative for CD5 and CD10. Plasma cells bearing cytoplasmic IgM-lambda were increased in pleural fluid. Karyotyping demonstrated t(2;11;18)(q21-23;q21;q21). Rearrangements within BCL2 and YES genes located at 18q21 were not detected. Sixteen other cases with t(11;18)(q21;q21) have been reported in marginal zone B-cell lymphoma. Therefore, our report is in agreement with the finding that part of primary macroglobulinemia is a variant of marginal zone B-cell lymphoma. PMID- 10704681 TI - Alterations of P19ARF in rodent hepatoma cell lines but not in human primary liver cancer. AB - The tumor suppressor gene CDKN2A is functionally inactivated, through mutations, deletions, or methylation, in a large variety of primary neoplasms as well as tumor cell lines. The CDKN2A locus gives rise to two distinct transcripts. P16INK4 and P19ARF. Because it has been shown that the disruption of only P19arf coding sequences in mice is sufficient for tumor development, this transcript most likely also encodes a tumor suppressor. We have analyzed the two CDKN2A transcripts in fifteen human primary liver carcinomas, two human hepatoma cell lines, and five rodent hepatoma cell lines. No homozygous deletions of P19ARF and P16INK4 were found in these samples, whereas the normal P19arf transcript was absent in two of the five rodent cell lines (nonexpressed in one case and mutated in another). These results suggest that functional abrogation of P19ARF is not a primary event in hepatocarcinogenesis. PMID- 10704682 TI - Fanconi anemia: myelodysplasia as a predictor of outcome. AB - The adverse potential of the development of myelodysplastic syndrome (MDS) in Fanconi anemia (FA) was examined in a retrospective study of 41 FA patients who had bone marrow morphology and chromosomes reviewed by a single group. Thirty three patients had adequate cytogenetic studies, and 16 (48%) had one or more abnormal studies: nine initially, and seven more on follow-up. Cytogenetic clonal variation was frequent, including disappearance of clones, clonal evolution, and appearance of new clones. The estimated five-year survival with a cytogenetic clone is 0.40, compared to 0.94 without a clone. Morphologic myelodysplasia (MDS), independent of a cytogenetic clone, was found in 13/41 patients (32%). The estimated five-year survival with MDS is 0.09, versus 0.92 without MDS. Leukemia developed in three patients whose initial cytogenetic clones prior to leukemia were t(1;18), t(5;22) and monosomy 7; the one with t(1;18) also had MDS. Our results focus on marrow morphology, and suggest that morphologic MDS may be more important than classical cytogenetics in prediction of an adverse outcome. PMID- 10704683 TI - Trisomy 15, sex chromosome loss, and hematological malignancy. AB - We report 6 patients with myelodysplasia who, on routine cytogenetic studies, demonstrated trisomy 15. Four of these also had sex chromosome loss. A review of the literature revealed 6 other cases of trisomy 15 with sex chromosome loss and 22 cases of trisomy 15 as the sole chromosomal abnormality. All cases had hematologic malignancy or myelodysplasia. Trisomy 15 is uncommon but tends to be associated with myelodysplasia in older subjects, and with sex chromosome loss in about one third of cases. PMID- 10704684 TI - High frequency of loss of heterozygosity for 1p35-p36 (D1S247) in Wilms tumor. AB - We analyzed the loss of heterozygosity (LOH) for 1p in 18 Wilms tumors using a panel of 11 polymorphic markers. Loss of heterozygosity was identified in 56% of the tumors. The smallest region of overlap was defined for marker D1S247, underlying the 1p35-1p36.1 locus. This is the highest LOH frequency for 1p, or for the well-defined 11p13 and 11p15.5 loci. Based on the fact that tumors of all stages, with both favorable and unfavorable histology, exhibited LOH, we suggest that the 1p35-1p36.1 locus is involved in the etiology of Wilms tumor. PMID- 10704685 TI - Translocation (5;9)(q22;q34) in a case of acute lymphoblastic leukemia with multiple bone involvement: effectiveness of donor lymphocyte infusion for relapse after allogeneic stem cell transplantation. AB - A case of acute lymphoblastic leukemia with a new translocation, t(5;9)(q22;q34) is reported with special reference to the clinical features and the response to treatment. This case exhibited several unique clinical features, including expression of the myeloid antigen on the early pre-B-cell phenotype, multiple bone involvement, and favorable response to donor lymphocyte infusion despite early relapse after allogeneic hematopoietic stem cell transplantation. PMID- 10704686 TI - Trisomy 4 in a case of gynecomastia. AB - Gynecomastia is an anomaly associated with alterations in the levels of hormones, especially estrogens. Sufferers of gynecomastia present a high risk of developing carcinomas. Only two cytogenetic descriptions of this type of mammary proliferation are available. In the present study, we analyzed chromosomally a sample of breast tissue from a patient with gynecomastia following short-term culture. The only clonal chromosomal alteration encountered was trisomy of chromosome 4. This alteration has not been described previously for samples of gynecomastia or breast cancer in males. We believe that the alterations in hormone levels in the male breast tissue that lead to this type of cellular proliferation induce the formation of chromosomal abnormalities, making the cells more susceptible to becoming malignant. PMID- 10704687 TI - Pentasomy 8 in acute monoblastic leukemia. AB - We report a case of pentasomy of chromosome 8 in 30-year-old man with de novo acute monoblastic leukemia (FAB AML M5a). PMID- 10704688 TI - Characterization of a chromosomally complex lung cancer cell line using multiwell fluorescence in situ hybridization. AB - The chromosomal characterization of a non-small cell lung cancer cell line (NCIH358) is described. This characterization was achieved using a simple, cheap and technically straightforward multiwell fluorescence in situ hybridization (FISH) method. The many and complex chromosome rearrangements identified by this method could not be defined using conventional G-banded chromosome analysis, and have not been previously described. For the detailed characterization of complex cell lines, multiwell FISH has many advantages over more technically demanding and expensive FISH techniques, and opens up the possibility of screening for consistent rearrangements, leading to the identification of unique fusion genes. PMID- 10704689 TI - Comparative genomic hybridization reveals extensive variation among different MCF 7 cell stocks. AB - Comparative genomic hybridization (CGH) allows the detection of DNA sequence copy number changes on a genome-wide scale in a single hybridization reaction. The ability of CGH to be applied to formalin-fixed, paraffin-embedded tumor samples has lead to its widespread application in the cytogenetic analysis of archival material. When setting up CGH in the laboratory, rigorous control experiments must be carried out to ensure that the losses and gains are scored correctly. Groups interested in breast cancer frequently use the MCF-7 cell line as a positive control in these experiments, comparing the results to previously described genetic alterations. Here we present the results of CGH carried out with three stocks of MCF-7 cells. The cells differ widely in their proliferative response to 17-beta estradiol and show extensive variation in copy number changes affecting specific chromosomal regions. We suggest that care must be taken, therefore, when choosing a cell line as a positive control for CGH experiments. PMID- 10704690 TI - Uneven frequencies of secondary chromosomal abnormalities in acute myeloid leukemias with t(8;21), t(15;17), and inv(16). AB - The types and incidences of secondary chromosomal abnormalities were analyzed in three subtypes of leukemia with recurrent abnormalities, translocations t(8;21), t(15;17), and inversion inv(16). The main types of clonal secondary abnormalities were similar to those described in the literature, loss of sex chromosome associated with t(8;21), trisomy 8 with t(15;17), and trisomies 8 or 22 with inv(16). On the whole, the incidence of clonal abnormalities was significantly higher in t(8;21) leukemia than in the two other subtypes. This difference was not related to a chromosomal instability peculiar to this leukemia subtype, because the incidence of nonclonal abnormalities was the same in the three types of leukemia studied. The significance of secondary clonal abnormalities remains speculative. A careful comparative analysis of structural rearrangements of the chromosomes usually involved in secondary abnormalities must be carried out as a first step to identify the key genes altered. PMID- 10704691 TI - Microsatellite instability in ovarian and other pelvic carcinomas. AB - Twenty-six cases of ovarian carcinoma and six cases of other pelvic neoplasms were analyzed for microsatellite instability (MSI) using frozen specimens, fluorescence technology, and four selected markers (D2S123 on chromosome 2, D18S58 on chromosome 18, BAT26 on chromosome 2, and BAT40 on chromosome 1). This procedure also allowed the detection of loss of heterogeneity (LOH) at the four selected loci. One of the cases of ovarian carcinoma exhibited MSI and this was evident at three loci. Of 44 informative loci, 7 exhibited LOH representing 3 cases of ovarian carcinoma, 3 of 4 cases of primary peritoneal carcinoma, and one case of unknown primary. These data support other findings that MSI is not a frequent occurrence in ovarian cancer; however, LOH is a more frequent event and may be a target for the development of diagnostic/prognostic procedures for ovarian and primary peritoneal carcinoma. PMID- 10704692 TI - New variant translocation (8;20)(q22;q13) in bone marrow cells of extramedullary blast crisis in chronic myelogenous leukemia. PMID- 10704693 TI - Additive effects of estrogen deficiency and diabetes on bone mineral density in rats. AB - We investigated the combined effects of estrogen deficiency and diabetes on bone mineral density (BMD) and bone metabolism in rats. Ten-week-old, female rats were randomly divided into four groups: controls (C), an ovariectomized group (O), a streptozotocin-induced diabetic group (S), and a combined ovariectomy and streptozotocin-induced diabetic group (OS). The BMD of the lumbar spine and the femur were measured before grouping and at 23 weeks old. At the end of the experiment, blood samples were obtained via cardiac puncture, and bone gla protein (BGP), tartrate-resistant acid phosphatase (TRAP) and 1,25 dihydroxyvitamin D levels were measured. The rats in the C, O, S, and OS groups, in that order, had higher levels of BMD of the lumbar spine and femur at 23 weeks of age. The BGP levels in the S and OS groups were significantly lower than in C and O groups. Significantly higher 1,25-dihydroxyvitamin D was observed in the O group compared with the C, S and OS groups. No differences were obtained in TRAP among four groups. Our data suggest that the combined effects of estrogen deficiency and diabetes on BMD are not synergistic or counteractive but additive. PMID- 10704694 TI - Self reported attitude and behavior of young diabetics about discussing their disease. AB - A performa-guided survey was conducted among 47 young patients of diabetes mellitus (onset of diabetes <30 years). Questions included were regarding the type of treatment, health status information about diabetes, and the assumptions and experiences of the patients on certain psychosocial behavior. A total of 59.6% subjects said that they could disclose everything about their disease to their friends and acquaintances. Twenty-seven percent felt that they could divulge only partial information and 12.8% did not want to discuss their disease with their friends and acquaintances. Subjects who said that they could disclose about their disease felt that they could do so because they were putting a lot of effort into achieving better control of their blood glucose. One of the fears expressed about not discussing their disease was that in doing so people would treat them differently or perceive them as sick. However only 38% experienced such a change in the behaviour of their acquaintances. Seventy-three percent of them had received unsolicited advice from others about food and dietary restrictions. Forty-three percent of the subjects had received instructions from acquaintances to stop all treatment and shift to household remedies. Hypoglycemia could be a motivating factor to help patients to discuss their illness with the acquaintances. PMID- 10704695 TI - Clinical evaluation of muscle strength in 20-79-years-old obese Japanese. AB - It is well known that obesity is closely related to non-insulin-dependent diabetes mellitus, hyperlipidemia, hypertension and cardiovascular disease, and the insulin resistance associated with obesity is supposed to play a central role for the development of these diseases. Thus, effective prevention and treatment of obesity need to be explored. In 357 obese (body mass index > or =26.4) subjects, aged 20-79 years, grip and leg strength were determined and compared with age- and sex-matched 1683 nonobese control subjects. Age-dependent alteration of body composition, evaluated by waist-hip ratio and the relative fat mass volume, was also compared. Finally, the relationship between the number of risk factors related to atherosclerosis and muscle strength was evaluated. Grip and leg strength in obese subjects were obviously stronger than controls under the age of 60 in both sexes. However, in the subjects over 60 years old, muscle strength was similar between obese subjects and controls. Weight bearing index (WBI) (leg strength (kg)/body weight (kg)) in obese subjects was remarkably lower than that in controls in all generations. In obese subjects, the waist-hip ratio and relative percentage of fat increased with aging, and obese subjects with multiple risk factors had higher waist-hip ratio and a tendency for lower muscle strength. Reduced WBI was considered to be a fundamental feature of obese subjects, and obese subjects increased fat composition with aging, which may be linked with low muscle strength. Thus, we need to design the most effective protocols to maximize and maintain quantitative and qualitative properties of muscle. PMID- 10704696 TI - Serum levels of carboxy-terminal propeptide of human type I procollagen are an indicator for the progression of diabetic nephropathy in patients with type 2 diabetes mellitus. AB - The finding that glomerular mesangial cells produce human type I collagen suggests that the serum levels of carboxy-terminal propeptide of human type I procollagen (P1CP) may reflect the severity of diabetic nephropathy. We therefore investigated the relationship between serum P1CP levels and the extent of diabetic complications in 100 patients (46 males and 54 females) with Type 2 diabetes and in 64 healthy subjects. Serum P1CP was determined by radioimmunoassay. In diabetes, we defined P1CP levels less than 142 ng/ml as a normal P1CP group (group A), whereas we defined them as equal to or greater than 142 ng/ml as a high P1CP group (group B). The diabetic patients had significantly elevated serum P1CP levels compared with the controls. The prevalence of hypertension, proliferative diabetic retinopathy or macroalbuminuria was significantly higher in group B than in group A. Serum P1CP levels showed a significant positive correlation with urinary albumin excretion, but not with fasting blood glucose, glycosylated hemoglobin A(1c) or serum osteocalcin. Macroalbuminuric patients showed significantly higher P1CP levels than the normoalbuminuric patients. In patients in the absence of diabetic nephropathy, no significant differences of P1CP levels were found among the severity of diabetic retinopathy. The present results suggest that serum P1CP levels reflect the progression of diabetic nephropathy in patients with Type 2 diabetes. PMID- 10704697 TI - Mitochondrial gene mutations in gestational diabetes mellitus. AB - Mitochondrial DNA mutations have been implicated in many diseases including diabetes mellitus. Although gestational diabetes mellitus (GDM) has been suggested to have genetic determinant and to be etiologically indistinct with non insulin-dependent diabetes mellitus (NIDDM), its association with mitochondrial gene mutations is still unknown. In this study, 137 patients with GDM and 292 non diabetic pregnant controls were examined for mitochondrial DNA mutations from the nucleotide 3130-4260 encompassing tRNA-Leu gene and adjacent NADH dehydrogenase 1 gene by polymerase chain reaction, single-stranded conformation polymorphism, restriction fragment length polymorphism and DNA sequencing. One heteroplasmic mutation at the position of 3398 (T-C), which changed a highly conserved methionine to threonine in NADH dehydrogenase subunit 1, was identified in 2.9% GDM patients but not in the controls, indicating its association with GDM (P = 0.01). Two novel mutations, a heteroplasmic C3254A and a homoplasmic A3399T, were also found in GDM subjects, the functional meaning of which merits further investigation. G3316A and T3394C mutations implicated in NIDDM, were seen at higher frequencies in patients with GDM than the controls. Our results suggest that mitochondrial DNA mutations may contribute to the development of GDM in some patients. PMID- 10704698 TI - Expenditure on health care incurred by diabetic subjects in a developing country- a study from southern India. AB - The objective of the study was to estimate the direct costs of diabetes care to patients attending secondary care facilities in Madras, India. A total of 596 subjects were studied, at the Private Hospital for Diabetes Mellitus (PHD) (n = 422), and at the Government General Hospital (GGH) (n = 174). A simple interview schedule enabled a face to face interaction with the patients by the research investigator which elicited a frank and true response. The validity of the data collected was established by independent scrutiny of financial records in a sub sample. Payment bills for expenses of 140 subjects chosen on a random basis from the total sample of 422 PHD patients were compared with the costs reported by the subjects. There were no statistically significant differences both in the inpatient and the outpatient cases between the reported cost and actual cost. Median bill value (total costs)=Rs.1010 (range 195-16700) reported value=880 (110 20355) Z = -0.97, P = 0.33 and, for outpatients, median bill value=Rs.800 (195 4560) reported value=Rs. 740 (110-6320) Z = -1.56, P = 0.12. For inpatients, median bill value = Rs. 4235 (1289-16700) reported value=Rs.5459 (1285-20355), Z = -1.27, P5 years duration of diabetes spent more than those who had <5 years of duration; Rs.5570 (360-75200) and Rs.3220, (460-25600), respectively. All differences between these sub-groups were statistically significant. Within the ambit of economic aspects of the population in a developing country, the direct cost on diabetes health care is very high for many people. PMID- 10704699 TI - Factors associated with microalbuminuria in type 1 diabetes mellitus: a cross sectional study. AB - In order to determine the prevalence of microalbuminuria in people with Type 1 diabetes mellitus (Type 1 DM) and identify factors associated with microalbuminuria, we studied 312 Type 1 DM patients attending in three hospitals in two Spanish regions over 6 months. Clinical characteristics, micro- and macro vascular complications, blood pressure, 24-h urine albumin excretion, lipid profile, HbA1(c) levels, smoking habits, and family history of hypertension and diabetic nephropathy were recorded. Univariate analysis and multiple logistic regression were used to examine associations between these variables and the prevalence of microalbuminuria. We detected microalbuminuria in 29% of the patients. The prevalence of microalbuminuria was high during the second decade of diabetes and declined thereafter. Univariate analysis showed dyslipidaemia (P<0. 002), previously diagnosed hypertension (P<0.001), family history of hypertension (sibling alone P<0.006; mother alone P<0.05), family history of diabetic nephropathy (P<0.001), and laser-treated retinopathy (P<0.03) to be factors associated with the presence of microalbuminuria. Multiple logistic regression revealed an association between microalbuminuria and family history of nephropathy (OR 7.6, 3.6-16). In conclusion, in our sample the frequency of microalbuminuria seems to be related to the presence of dyslipidaemia, hypertension, and to a family history of hypertension or nephropathy. PMID- 10704700 TI - Vascular complications in young Asian Indian patients with type 1 diabetes mellitus. AB - To determine the prevalence of micro vascular and macro vascular complications in Asian Indian Type 1 diabetic subjects. There has been no major report on the prevalence of vascular complications in Type 1 diabetic patients in India. This study was done in Type 1 diabetic patients, aged < or =20 years at diagnosis of diabetes (n=617, M:F 322:295) with a minimum of 3 year follow-up. Standard diagnostic methodologies were used to test for micro vascular and macro vascular complications of diabetes. Retinopathy was detected in 13. 4% (background diabetic retinopathy 11.2%, proliferative diabetic retinopathy 1.9%, preproliferative 0.31%, maculopathy was seen in 13.3% of retinopathy cases), nephropathy in 7.1%, sensory neuropathy in 3.0%, ischaemic heart disease in 0.5% and peripheral vascular disease in 0.5% of the study subjects. Duration of diabetes showed positive association with retinopathy, nephropathy and neuropathy. Average glycosylated haemoglobin values, at follow up showed an association with retinopathy. Although the glycaemic control was suboptimal in the study group, prevalences of all complications, especially macro vascular complications were lower in Type 1 diabetic patients in this ethnic group, in comparison with the European or American counterparts. PMID- 10704701 TI - Postprandial hypertriglyceridaemia in treated type 2 diabetic subjects --the role of dietary components. AB - Postprandial hyperlipidaemia is a risk factor for cardiovascular diseases (CVD). This study was done (a) to evaluate whether postprandial hypertriglyceridaemia was common in Indian type 2 diabetic patients on treatment and (b) to see whether the high carbohydrate content of the diet was a cause of the lipid abnormality. Two hundred type 2 diabetic subjects (M:F, 137:63; mean age 51.6+/-10.2 years, mean BMI 25.5+/-3.1 kg/m(2)) with diabetes duration of 7.6+/-5.6 years were studied. Fasting and 2 h post prandial responses of plasma glucose and triglycerides (TG) were measured using a breakfast meal, usually consumed by the patient with the intake of usual hypoglycaemic drugs. Patients with a post prandial TG value greater than 15% of the corresponding fasting TG value were designated as group 2 and the remaining subjects as group 1. Dietary composition of the breakfast were calculated. Among the 200 subjects, 52 (26%) had post prandial TG higher than the fasting values. This was seen in patients who were consuming lower percentage of carbohydrates and higher percentage of fats than prescribed. Therefore the postprandial rise in TG was probably due to the high fat content of the diet and due to a lower insulin sensitivity. This study highlights the facts that postprandial hypertriglyceridaemia is seen only in a small proportion of the treated patients and the high carbohydrate diet does not produce hypertriglyceridaemia, either in the fasting or post prandial state. The minority who show an increased TG value at 2 h have been taking lower carbohydrate with higher fat content in the meal. This could have produced a lower insulin sensitivity in these patients. PMID- 10704702 TI - Fetal growth is not associated with early onset of severe retinopathy in type 1 diabetes mellitus. AB - Reduced fetal growth has been suggested as a possible risk factor for diabetic nephropathy. The aim of the present study was to examine whether there could be an association also with rapidly progressing severe retinopathy in younger type 1 diabetic patients. Maternal pregnancy, as well as birth parameters of 27 type 1 diabetic patients with severe retinopathy diagnosis at a median age of 25 years, were studied retrospectively. The control group consisted of 22 type 1 diabetic patients with mild background retinopathy and with similar age, age at onset, and duration of diabetes. Mothers of the subjects with severe retinopathy had a higher body mass index (P = 0.03) but similar age, blood pressure levels, and weight gain during pregnancy as those of the control group. All but four babies, two in each group, were born after 37 completed gestational weeks. There were no differences regarding birth weight or of relative birth weight corrected for gestational length. Head circumference, birth length, and placenta weight were similar. The results indicate that fetal growth is not a factor of major importance for the development of severe retinopathy in younger type 1 diabetic patients. PMID- 10704703 TI - Small cell lung cancer in women: risk associated with smoking, prior respiratory disease, and occupation. AB - Small cell carcinoma of the lung (SCLC) occurs most frequently in heavy smokers, yet exhibits a lesser predominance among men than other smoking-associated lung cancers. Incidence rates have increased more rapidly in women than men and at a faster rate among women than other cell types. To investigate the importance of smoking and other risk factors, a case-control study of SCLC in women was conducted. A total of 98 women with primary SCLC and 204 healthy controls, identified by random-digit dialing and frequency matched for age, completed telephone interviews. Data collected include demographics, medical history, family cancer history, residence history, and lifetime smoking habits. Odds ratios (ORs) and 95% confidence intervals (95% CI) were calculated using logistic regression analysis. Risk for small cell carcinoma in women is strongly associated with current use of cigarettes. Ninety-seven of 98 cases had smoked cigarettes; 79% of cases were current smokers and 20% were former smokers at the time of diagnosis compared to 13% current and 34% former smokers among controls. The ORs associated with smoking are 108.7 (95% CI 14.8-801) for ever-use of cigarettes, 278.9 (95% CI 37.0-2102) for current smoking, and 31.5 (95% CI 4. 1 241) for former smoking. Risk increases steeply with pack-years of smoking and decreases with duration of smoking cessation. After adjusting for age, education, and lifetime smoking history, medical history of physician-diagnosed respiratory disease including chronic bronchitis, emphysema, pneumonia, tuberculosis, asthma, and hay fever is not associated with a significant increase in lung cancer risk. Employment in blue collar, service, or other high risk occupations is associated with a two to three-fold non-significant increase in risk for small cell carcinoma after adjusting for smoking. PMID- 10704704 TI - The impact of overall treatment time on outcomes in radiation therapy for non small cell lung cancer. AB - PURPOSE: A retrospective study was carried out to evaluate the impact of overall treatment time (OTT) on the results of radiation therapy for non-small cell lung cancer (NSCLC). MATERIALS AND METHODS: From Jan. 1990 to Dec. 1996, 256 patients with stages I-IIIb NSCLC entered this analysis. All patients received definitive radiotherapy. Biologically effective dose (BED) was used to standardize the irradiation effects. The correlation between OTT and local progression-free survival was analyzed by linear-regression and Cox proportional hazard models. The prognostic variables for survival and distant metastasis were also briefly studied. RESULTS: OTT had been shortened in 64 patients because of an accelerated hyperfractioned irradiation, while OTT was prolonged i n 114 patients due to interruptions of irradiation courses. The main ca uses of interruption were machine breakdown or delayed preparations of c errobend block for boost fields (55%), holidays (11%) and treatment toxi city and side effects (34%). Patients tre ated with prolonged OTT (> 45 days) had significant poorer local progression free survival than whom with OTT of T transition mutation on codon 531 (Gln-531- >Amber-531) of src gene. This mutation caused a prematured translation termination and up-regulated the kinase activity. To examine whether this mutation could be a risk factor for colon carcinoma in the Chinese population, we used the same PCR-based assay to analyze src genotypes of 131 colon cancers and other various types of carcinoma. No mutation was detected in all specimens that were screened in this study. Thus, mutation at Gln-531 of src gene does not seem to be involved in the development of colon cancer in Chinese ethnicity. PMID- 10704744 TI - A comparative study of normal and reverse phase high pressure liquid chromatography for analysis of porphyrins accumulated after 5-aminolaevulinic acid treatment of colon adenocarcinoma cells. AB - Primary adenocarcinoma cells of the rectosigmoid colon (WiDr-cells) were treated with 5-aminolevulinic acid (5-ALA). Cellular porphyrins were separated and quantified by high performance liquid chromatography (HPLC), both as free porphyrin acids after an easy extraction method with a subsequent reverse phase technique, and then as porphyrin esters after a more laborious extraction method and subsequent normal phase technique. The porphyrins were detected by means of a fluorescence detector. Analysis by normal phase HPLC indicated that 81% (739 pmol/mg protein) of the total amounts of fluorescing porphyrins accumulated was protoporphyrin IX, while similar analysis by reverse phase HPLC indicated that PpIX constituted 91% (622 pmol/mg protein) of the accumulated porphyrins. In addition to protoporhyrin IX, copro-, hexa-, hepta- and uroporphyrins were observed in extracts from 5-ALA-treated cells by both methods. The discrepancy between the two methods increased with increasing hydrophilicity of the analysed porphyrins, with uroporphyrin estimated to be 6-fold higher (63 vs. 10 pmol/mg protein) by normal than by reverse phase HPLC. PMID- 10704745 TI - Complex-type chromosomal exchanges in blood lymphocytes during radiation therapy correlate with acute toxicity. AB - The new method of chemical-induced premature chromosome condensation combined with fluorescence in situ hybridization was used to analyze chromosomal damage in peripheral blood mononuclear lymphocytes of patients undergoing radiation treatment for esophageal cancer with high-energy X-rays or accelerated carbon ions at the National Institute of Radiological Sciences (Chiba, Japan). Total number of aberrant cells correlated with radiation field size, but no correlation was found with acute toxicity. A high frequency of complex-type exchanges were also recorded. This aberration type presented a high individual variability, and correlated well with the acute morbidity. Cytogenetic analysis by interphase chromosome painting is proposed as a useful tool for monitoring normal tissue effects during radiotherapy. PMID- 10704746 TI - Effects of riluzole on the electrophysiological activity of pallidal neurons in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-treated monkey. AB - The effect of riluzole administration, an antiglutamatergic compound, on the electrophysiological activity of the pallidal complex of 1-methyl-4-phenyl 1,2,3,6-tetrahydropyridine (MPTP)-treated monkeys is compared with those induced by two dosages of levodopa (L-DOPA), the first affording the best clinical alleviation, the second sufficient to induce dyskinesias. Both dosages of L-DOPA reduced sharply the firing frequency of globus pallidus pars internalis (GPi) neurons (respectively, 43.8+/-23.0 and 27.4+/-20.2 vs. 111. 2+/-31.4 Hz), decreased the percentage of bursting cells (respectively, 60.7 and 50.0 vs. 80.3%) and augmented the number of regular cells (respectively, 6.5 and 33.0 vs. 4.8%). Riluzole restored the firing frequency (75.0+/-26.9 Hz) and the firing pattern of the GPi (39.7% bursting, 9.5% regular and 50.8% irregular). These results suggest that the emergence of dyskinesia may well be due to a modification of the neuronal messages transmitted from the GPi to the motor nuclei of the thalamus. Riluzole would represent an interesting alternative to dopamine therapy in Parkinson's disease since it regularizes firing but does not cause dyskinesia. PMID- 10704747 TI - Neuroprotective effects of chronic estradiol benzoate treatment on hippocampal cell loss induced by status epilepticus in the female rat. AB - Neuroprotective properties of estrogen are supported by extensive experimental evidence. In this study, the effects of estrogen were examined on the neurodegeneration secondary to status epilepticus induced by kainic acid in the rat. Chronic supplementation of ovariectomized rats with estradiol benzoate (20 microg/day) did not modify the expression of seizures monitored by electroencephalography, but significantly reduced cellular loss in the hippocampus. This neuroprotection was in particular observed in the dentate hilus and CA3 pyramidal layer when treatment with estradiol benzoate was started five days before status epilepticus induction. These findings suggest that estrogen can exert neuroprotective effects in a model of status epilepticus, in the absence of anti-epileptic properties. PMID- 10704748 TI - Acute application of interleukin-1beta induces Ca(2+) responses in human microglia. AB - The effects of the pro-inflammatory cytokine interleukin-1-beta (IL-1beta) on levels of intracellular calcium [Ca(2+)](i) in cultured human microglia have been studied using the fluorescent Ca(2+) indicator fura-2. IL-1beta (2 ng/ml) caused a slow, progressive increase in [Ca(2+)](i) in standard Ca(2+)-containing physiological solution (PSS). A similar effect was observed in separate studies using Ca(2+)-free PSS, however, the mean rate of increase was significantly lower than that measured with PSS. Similar results were obtained in a separate protocol, where cells were exposed to both IL-1beta in Ca(2+)-free PSS and PSS. The slope of the IL-1beta induced increase of [Ca(2+)](i) in Ca(2+)-free PSS was not altered when adenosine triphosphate was added prior to application of the cytokine. These results suggest that IL-1beta-induced responses in human microglia involve both a Ca(2+) entry pathway and a mechanism of intracellular increase other than from IP(3)-sensitive stores. PMID- 10704749 TI - The control of human locomotor pointing under restricted informational conditions. AB - Since a perception-action coupling type of control (Kugler, P.N. and Turvey, M.T., Information, Natural Law, and Self-Assembly of Rhythmic Movements, Erlbaum, Hillsdale, 1987, 481 pp.) continuously operates during locomotor pointing tasks (e.g. long jumping) (Montagne, G., Cornus, S., Glize, D., Quaine, F. and Laurent, M., A 'perception-action coupling' type of control in long-jumping. J. Motor Behav., (2000) in press), the information sources underlying this control have to be dealt with. Under the assumption that subjects use information about the first order time remaining before they pass the target, we identify in the literature four different sources of information that specify this physical property. Only one of these sources is inevitably present under all possible environmental conditions containing at least a continuously visible target on the floor. This study aimed to test its sufficiency to perform a locomotor pointing task. The use of a virtual reality set-up permitted us to compare locomotor pointing executed with all four information sources or only with the aforementioned one. The likeness found between those two conditions, as far as the pointing performance and the mode of control are concerned, expresses the evoked sufficiency. PMID- 10704750 TI - Maintenance of tricarboxylic acid cycle kinetics in Brown-Norway Fischer 344 rats may translate to longevity. AB - Glucose metabolism and tricarboxylic acid cycle (TCA) kinetics have been shown to decline in brain with age in various animal species. This study examined TCA cycle kinetics and age in Brown-Norway Fischer 344 rats. Using [1-(13)C]glucose infused over 10, 30, 60 or 100 min, and following the label through the TCA cycle using (1)H?(13)C? spin-echo difference magnetic resonance spectroscopy, groups of 2 (n=18), 12 (n=16), and 24 (n=16) month old rats were evaluated. Unexpectedly, TCA cycle kinetics did not change with age. Observed decreases in glutamate, glutamine and N-acetyl aspartate levels are consistent with an age-related decrease in neuronal numbers. The possible link between this observation and increased longevity, together with a decreased incidence of neoplasia in the Brown-Norway Fischer 344 rat is discussed. PMID- 10704751 TI - Diminution of N-methyl-D-aspartate-induced perturbation of neurotransmission by dexmedetomidine in the CA1 field of rat hippocampus in vitro. AB - The effects of alpha(2)-adrenoceptor activation on N-methyl-D-aspartic acid (NMDA)-induced perturbation of neurotransmission and normal NMDA-receptor dependent function (long-term potentiation, (LTP)) were investigated in the hippocampal CA1 field in vitro. Bath perfusion of dexmedetomidine hydrochloride (50 nM), which was initiated before NMDA (100 microM) exposure, enhanced the extent of recovery of extracellular field excitatory postsynaptic potentials after NMDA infusion. On the other hand, the induction and early maintenance of LTP was normal in the presence of dexmedetomidine. Thus, dexmedetomidine can diminish acute NMDA-induced perturbation of neurotransmission while the same dose of this drug does not influence the normal activation of NMDA receptors. PMID- 10704752 TI - Vestibular, visual, and somatosensory contributions to human control of upright stance. AB - We investigated the changes of human posture control of upright stance which occur when vestibular cues (VEST) are absent and visual and somatosensory orientation cues (VIS, SOM) are removed. Postural responses to sinusoidal tilts of a motion platform in the sagittal plane (+/-2 degrees, f=0.05, 0.1, 0.2 and 0.4 Hz) were studied in normal subjects (Ns) and patients with bilateral vestibular loss (Ps). We found that absence of VEST (Ps, visual reference) and removal of VIS (Ns, no visual reference) had little effect on stabilization of upright body posture in space. In the absence of both VEST and VIS (Ps, no visual reference) somatosensory graviception still provided some information on body orientation in space at 0.05 and 0.1 Hz. However, at the higher frequencies Ps qualitatively changed their behavior; they then tended to actively align their bodies with respect to the motion platform. The findings confirm predictions of a novel postural control model. PMID- 10704753 TI - Increased amygdala volumes in female and depressed humans. A quantitative magnetic resonance imaging study. AB - The amygdala are thought to play an important role in emotional information processing. First studies indicate a link between amygdala atrophy, fear and aggression and between amygdala hypertrophy and depression. To investigate a possible relationship between amygdala volumes, aggression and depression, we measured the amygdala of 62 patients with temporal lobe epilepsy (TLE) with and without aggressive behavior or depression and 20 healthy volunteers using quantitative magnetic resonance imaging (MRI). Amygdala volumes of female patients (n=26) were significantly larger than those of males (n=36) (left side: P=0.001; right side P=0.05). Depressed patients displayed significant enlargement of both amygdala (left side: P=0.008; right side: P=0.001) There was no significant finding relating to the factor aggression neither was there any significant interaction between aggression, dysthymia and gender. PMID- 10704754 TI - Leukemia inhibitory factor null mice: unhampered in vitro outgrowth of sensory axons but reduced stimulatory potential by nerve segments. AB - Leukemia inhibitory factor (LIF) is locally up-regulated after peripheral nerve injury and may be involved in the subsequent regeneration. Here, adult mice with or without LIF gene deletions were used to study the role of LIF in regeneration. The results show that axonal regeneration in vitro from dorsal root ganglia (DRGs) was unaffected by LIF deletion. However, segments from wild type mice promoted DRG axonal outgrowth better than segments from LIF deleted animals when in vivo-injured sciatic nerve segments were co-cultured with DRGs from normal adult mice. Addition of LIF could not restore the deficit. This suggests that LIF is engaged in the local regulation of regeneration but not in the regenerative events occuring at the cell body level. PMID- 10704755 TI - Post-treatment with nicotinamide (vitamin B(3)) reduces the infarct volume following permanent focal cerebral ischemia in female Sprague-Dawley and Wistar rats. AB - Delayed treatment with nicotinamide (NAm) protects male rats against cerebral ischemia. Since the preponderant use of male animals in stroke research may produce results not applicable to female stroke patients due to gender-related differences, we examined whether delayed NAm treatment could protect female rats against focal cerebral ischemia using a model of permanent middle cerebral artery occlusion (MCAo). NAm (500 mg/kg) given intravenously, 2 h after MCAo, significantly reduced the infarct volume of female Sprague-Dawley (55%, P<0.05) and Wistar rats (60%, P<0.05) rats when compared with saline-injected controls. These studies confirm that NAm is neuroprotective specifically at the dose of 500 mg/kg in rats. The novel findings are that this neuroprotection occurs in female, as well as male rats, and that the neuroprotection observed is more robust when administered as an intravenous bolus compared with intraperitoneal administration. PMID- 10704756 TI - Arborization of dewlap motoneurons in the green anole lizard (Anolis carolinensis) is not sexually dimorphic. AB - Male anoles extend a bright red throat fan, called a dewlap, during both courtship and aggressive encounters. Female anoles perform this behavior less often than males and only in aggression towards both sexes. The cartilage, muscle fibers, and motoneuron somata controlling the display are larger in males than females. In the present study, we used the Golgi technique in an effort to characterize more completely the morphology of these dewlap motoneurons, and to investigate whether the dendritic arborization is different between the sexes. In addition to describing the morphology, we report that the length of processes, and numbers of primary processes and branch points are comparable in males and females. This similarity in arborization represents an intriguing contrast to other sexually dimorphic neuromuscular systems. PMID- 10704757 TI - Neon colour spreading in three-dimensional illusory objects in humans. AB - We studied whether neon spreading can be induced within three-dimensional illusory triangles. Kanizsa triangles were induced by black pacman disks consisting of red sectors with curved sides. Viewing our stimuli monocularly produced two-dimensional illusory contours and surfaces as well as neon spreading in each figure. Triangles appeared concave or convex under stereoscopical viewing. Neon colour spreading was induced within illusory figures bending in three-dimensional space, suggesting that neural contour completion and surface filling-in interact across depth. Surprisingly, neon spreading was induced above the intervening surface even when the inducers were below the surface. Neon colour and illusory configuration were preserved behind the intervening surface only when it appeared transparent. PMID- 10704758 TI - Partial neuroprotection of in vivo excitotoxic brain damage by chronic administration of the red wine antioxidant agent, trans-resveratrol in rats. AB - The antioxidant compound trans-resveratrol, is found in substantial amount in several types of red wine and is considered one of the substances responsible for the lower incidence of coronary heart diseases among regular consumers of such wines, an effect also known as the French paradox. It has also been proposed that resveratrol may have beneficial effects against neurodegenerative diseases. We report here that chronic administration of resveratrol to young-adult rats, significantly protects from the damage caused by systemic injection of the excitotoxin kainic acid, in the olfactory cortex and the hippocampus. The same treatment, however, is not able to give any significant protection in an ex vivo model of simulated ischemia on hippocampal slices in vitro. This first evidence of a partial neuroprotective action of chronic administration of resveratrol in vivo, suggests that other models of neurodegenerative injury, and in particular of excitotoxic brain damage, should be investigated in order to assess the potentiality for resveratrol to be used as a pharmacological tool for neuroprotection. PMID- 10704759 TI - Activation of gamma-aminobutyric acid(A) receptors in the paraventricular nucleus of the hypothalamus reduces apomorphine-, N-methyl-D-aspartic acid- and oxytocin induced penile erection and yawning in male rats. AB - The effect of muscimol and baclofen injected into the paraventricular nucleus of the hypothalamus on penile erection and yawning induced by apomorphine, oxytocin and N-methyl-D-aspartic acid (NMDA) was studied in male rats. Muscimol (20-200 ng), but not baclofen (200 ng), injected into the paraventricular nucleus of the hypothalamus 10 min before apomorphine (50 ng), oxytocin (10 ng) or NMDA (50 ng) reduced penile erection and yawning induced by the above compounds given into the paraventricular nucleus. Bicuculline (250 ng) injected into the paraventricular nucleus 5 min before muscimol (100 ng) prevented the inhibitory effect of muscimol on penile erection and yawning induced by apomorphine, oxytocin and NMDA. The present results show that gamma-aminobutyric acid (GABA) inhibits penile erection and yawning by acting on GABA(A) receptors in the paraventricular nucleus of the hypothalamus. PMID- 10704760 TI - Acute inflammation differentially alters the activity of two classes of rat spinal visceral nociceptive neurons. AB - Quantitative neurophysiological studies have identified the presence of at least two spinal neuron populations (ABRUPT and SUSTAINED) which are excited by the noxious visceral stimulus colorectal distension (CRD). The present study examined the effects of acute colorectal inflammation on the activity of dorsal horn neurons in decerebrate, cervical spinal cord-transected male rats. Extracellular recordings were made using tungsten microelectrodes and inflammation was produced by intracolonic instillation of turpentine (25% solution). The total activity of SUSTAINED neurons during CRD increased starting one hour after turpentine instillation whereas the total activity of ABRUPT neurons during CRD, as a group, was unaffected during the two hours of study. Increases in total activity during CRD correlated with increases in spontaneous activity. These observations further support that visceral nociception travels by a dual pathway and suggest a predominant role for SUSTAINED neurons in the signaling of visceral pain-related events. PMID- 10704761 TI - Effects of kindling in wheat germ agglutinin-horseradish peroxidase [corrected] labeling in neurons of the interamygdaloid pathway in rats. AB - This paper describes in kindled rats an increment in wheat germ agglutinin horseradish peroxidase labeling in anterior commissure, bed nuclei of stria terminalis and amygdala. Three groups of animals were analyzed: control, sham operated and kindled animals with ten convulsive generalized seizures. Results show that kindled animals have an increase in fiber labeling in anterior commissure and in the bed nuclei of stria terminalis, as well as a greater number of labeled neurons in amygdala. This label enhancement is related to the hyperexcitability of neurons produced by epilepsy, and could be associated to the propagation and formation of secondary foci and related plastic changes occurring during kindling. PMID- 10704762 TI - Brain angiotensin type 1 receptor expression and function in the Zucker obese rat. AB - The Zucker obese rat is a model with predisposition for hypertension. There is evidence that angiotensin II (ANG II) may play a role in the maintenance of this hypertension. However, the potential role of brain ANG II in this regard has been largely unexplored. The aim of the present study was to compare the pressor response produced by i. c.v. injection of ANG II in Zucker obese and lean rats, and to determine if functional differences could be correlated to changes in brain AT1 receptor protein and/or mRNA expression. The Zucker obese rat had a significantly greater increase in blood pressure after i.c.v. injection of ANG II compared to the lean rat. AT1 receptor protein expression was greater in the brainstem, but not the hypothalamus, of the obese rat. These data raise the possibility that increased central responsiveness to ANG II may play a role in the predisposition of the Zucker obese rat to hypertension. PMID- 10704763 TI - Middle cerebral artery occlusion produces secondary, remote impairment in hippocampal plasticity of rats - involvement of N-methyl-D-aspartate receptors? AB - There is increasing evidence that focal cerebral ischaemia produces remote functional alterations that may substantially contribute to the post-stroke neurological outcome. Changes initially limited to peri-infarct areas may evolve and spread via transneuronal connections to other structures. In the present study we investigated whether focal ischaemia produced by 2-h occlusion of the middle cerebral artery (MCAo) in SD rats may influence the physiological function of the hippocampus. Three days later in vitro long-term potentiation (LTP) was studied in hippocampal slices from ipsi- and contralateral hemispheres. In rats with MCAo LTP was not-inducible in the ipsilateral hippocampus, while the contralateral side expressed stable potentiation (6.6+/-4.1 vs. 35. 0+/-8.0%, respectively). Treatment with 6-h i.v. infusion of an uncompetitive N-methyl-D aspartate (NMDA) receptor antagonist MRZ 2/579 starting at reperfusion not only preserved but additionally enhanced ipsilateral LTP, while a slight insignificant decrease was observed in the contralateral side (77.0+/-18.4 vs. 20.8+/-6.5%). The study demonstrates post-stroke functional changes in the hippocampus that can be modulated by NMDA receptor antagonists. PMID- 10704764 TI - Expression of growth hormone receptor in the human brain. AB - This study was designed to investigate the presence of growth hormone receptor (GHR) expression in the human brain tissue, both normal and tumoral, as well as in the human glioblastoma cell line U87MG. Reverse transcription-polymerase chain reaction revealed the presence of GHR mRNA in all brain samples investigated and in U87MG cells. GHR immunoreactivity was also detected in this cell line using both immunocytochemistry and western blotting. All together, our data demonstrate the existence of GHR expression within the central nervous system (CNS), thus supporting a possible role for GH in the CNS physiology. PMID- 10704765 TI - Odorants suppress voltage-gated currents in retinal horizontal cells in goldfish. AB - Odorants are known to suppress non-selectively voltage-gated currents in olfactory receptor cells. We found that odorants also suppress voltage-gated currents in neurons of outside of the olfactory system. Under voltage clamp, odorants such as amyl acetate, limonene, and acetophenone suppressed non selectively voltage-gated currents (a Ca(2+) current, a delayed rectifier K(+) current, a fast transient K(+) current, and an anomalous rectifier K(+) current) in horizontal cells from the goldfish retina. An amyl acetate puff completely and immediately suppressed the Ca(2+) current (I(Ca)) and the delayed rectifier K(+) current induced by repetitive depolarizations, suggesting that amyl acetate is a closed-channel blocker. Odorants did not change significantly the activation curve of I(Ca), but made the slope of inactivation curve of I(Ca) gentler and shifted its half-inactivation voltage toward a negative voltage. These results are similar to the effects of odorants on voltage-gated currents in olfactory receptor cells. This suggests that odorants may suppress the voltage-gated currents in retinal horizontal cells by the same mechanism described in olfactory receptor cells. PMID- 10704767 TI - Menstrual changes in sleep, rectal temperature and melatonin rhythms in a subject with premenstrual syndrome. AB - We studied a sighted woman with premenstrual syndrome who showed menstrual changes in circadian rhythms. She showed alternative phase shifts in the sleep rhythm in the menstrual cycle: progressive phase advances in the follicular phase and phase delays in the luteal phase. Rectal temperature rhythm also showed similar menstrual changes, but the phase advance and delay started a few days earlier than changes in sleep-wake rhythm so that the two rhythms were dissociated around ovulation and menstruation. These results suggest that her circadian rhythms in sleep and temperature are under the control of ovarian steroid hormones and that these two rhythms have different sensitivity to the hormones. PMID- 10704766 TI - Ca(2+) channel-mediated currents in retinal glial (Muller) cells of the toad (Bufo marinus). AB - Whole-cell voltage-clamp recordings were used to detect voltage-gated Ca(2+) channels in freshly isolated retinal glial (Muller) cells of the toad (Bufo marinus). Using Ca(2+) ions (2 mM) as charge carriers (in the presence of 1 mM Mg(2+)), no inwardly directed currents could be observed during the application of depolarizing voltage steps. However, after omitting the divalent cations from the bath solution, large-amplitude inwardly directed currents were evoked that were carried by Na(+) ions, and were mediated by at least two different kinds of Ca(2+) channels, transient low voltage-activated (LVA) channels and sustained high voltage-activated (HVA) channels. While the LVA currents activated at potentials positive to -90 mV and peaked at -40 mV, the HVA currents activated positive to -60 mV and peaked at -20 mV. It is concluded that Muller glial cells of the toad express distinct types of voltage-gated Ca(2+) channels that may be activated, under certain conditions, close to physiological membrane potentials. PMID- 10704768 TI - Distribution of prostaglandin EP(3) and EP(4) receptor mRNA in the rat parabrachial nucleus. AB - By using in situ hybridization, the distribution of mRNA for the PGE(2) receptors EP(3) and EP(4) was examined in the rat parabrachial nucleus (PB), a major brain stem relay for autonomic and nociceptive processing. EP(3) receptor mRNA was present in most subnuclei, with the densest labeling in the external lateral, dorsal lateral, superior lateral, central lateral and Kolliker-Fuse nuclei. EP(4) receptor mRNA expressing cells had a more restricted distribution, largely being confined to the superior lateral and adjacent parts of the dorsal and central lateral nuclei in a pattern complementary to that for EP(3) receptor mRNA. These findings suggest that EP(3) and EP(4) receptors in PB have distinct functional roles that include nociceptive processing, blood pressure regulation and feeding behavior. PMID- 10704769 TI - Pramipexole inhibits lipid peroxidation and reduces injury in the substantia nigra induced by the dopaminergic neurotoxin 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine in C57BL/6 mice. AB - Pramipexole has been showed to protect cultured dopaminergic (DAergic) cells against free radical-induced cytotoxicity. To test if pramipexole is protective against 1-methyl-4-phenyl-1,2,3, 6-tetrahydropyridine (MPTP)-mediated nigral DAergic injury in vivo and if such protection is related to inhibition of lipid peroxidation, DAergic function and lipid peroxidation were determined in MPTP treated C57BL/6 mice. We reported that MPTP administration induced a 38.1% increase of lipid peroxidation product thiobarbituric acid reactive substance (TBARS) in nigra, a 46.7% decrease of tyrosine hydroxylase -positive nigral DAergic neurons and a 59.4% reduction of striatal DA levels. However, pramipexole treatment significantly inhibited the TBARS production by 76%, and attenuated the MPTP-induced decreases in nigral DAergic neurons and striatal DA levels by about 50%. This study suggests that pramipexole can inhibit free radical-mediated lipid peroxidation and protect MPTP-induced nigral injury. PMID- 10704770 TI - Transthyretin binds amyloid beta peptides, Abeta1-42 and Abeta1-40 to form complex in the autopsied human kidney - possible role of transthyretin for abeta sequestration. AB - The deposition of amyloid beta protein (Abeta), a proteolytic cleavage product of amyloid precursor protein (APP), is an invariable pathological feature of the Alzheimer's disease brain, while APP gene is widely expressed in all neuronal and non-neuronal tissues with the highest levels of expression in the brain, and kidney. To understand the role transthyretin (TTR) plays in the sequestration mechanism of Abeta in the kidney, we have investigated interactions of TTR with Abeta1-40 and Abeta1-42 molecules by an immunoprecipitation method, in vitro binding studies, and overlay assay. These in vivo and in vitro biochemical experiments showed that TTR bound Abeta1-42 preferentially, and Abeta1-40 only to a limited extent, to form TTR-monomer and -dimer-Abeta complexes in the normal human kidney. We provide new evidence supporting the hypothesis that TTR, an Abeta binding protein, plays an important role in the sequestration of Abeta and prevents amyloid formation in the kidney. PMID- 10704771 TI - The inhibitory influence of an acquired escape strategy on subsequent avoidance learning in gerbils. AB - In gerbils an acquired non-avoidance strategy as pre-experience in a shuttle-box was studied in its influence on the learning of a standard avoidance paradigm. Thereby the same cue stimuli, frequency modulated tones (CS) and electric footshocks (US) were used in different behavioral paradigms. In the preexperience sessions the interstimulus interval between CS and US and on the other hand the escapability or inescapability of the US was varied. It was found that the experience with a relatively long interstimulus interval led to a prolonged maintenance of an acquired escape strategy during subsequent standard avoidance learning. This effect increased with the preexperience of an inescapable US. Additional insights into the temporal inhibition of avoidance learning by the pre experience were provided by behavioral events like attention responses and orienting responses reflecting components of information processing. PMID- 10704772 TI - CPI-1189 inhibits interleukin 1beta-induced p38-mitogen-activated protein kinase phosphorylation: an explanation for its neuroprotective properties? AB - The p38 mitogen-activated protein kinase (p38-MAPK) is a central enzyme in one of the major protein kinase cascades that regulate proapoptotic and proinflammatory signal transduction. p38-MAPK is activated by receptor/ligand recognition events or by exposure to extracellular stressors, including oxidative stress. Activation of p38-MAPK is affected by dual phosphorylation on a specific inhibitory domain. Dual phosphorylation causes a structural change in the p38-MAPK enzyme which allows binding of ATP and target substrate. Agents which block ATP docking to phosphoactivated p38-MAPK are being investigated for treatment of inflammatory diseases and neurodegenerative pathologies. An alternative strategy for p38-MAPK antagonism would be the inhibition of p38-MAPK phosphoactivation. We now report potent inhibition of p38-MAPK phosphorylation by a synthetic benzamide (CPI-1189) which displays protective action against tumor necrosis factor-alpha (TNFalpha) induced neurodegeneration. In primary astrocytes treated with interleukin 1beta (IL1beta), CPI-1189 inhibits p38-MAPK phosphorylation at concentrations of 10 nM or less. While the precise molecular target of CPI-1189 remains unknown, these findings suggest a novel mechanism for the neuroprotective properties of the compound. These findings also indicate that antagonism of the p38-MAPK may be achieved through pharmacological inhibition of p38-MAPK phosphorylation, a strategy that is conceptually distinct from direct inhibition of ATP binding to the active enzyme. PMID- 10704773 TI - Protective effect of topiramate against hippocampal neuronal damage after global ischemia in the gerbils. AB - The purpose of this study was to examine whether topiramate would reduce neuronal damage after transient global ischemia in the gerbils because topiramate blocks voltage sensitive sodium channels and non-N-methyl-D-aspartate receptors and enhances gamma-aminobutyric acid-mediated inhibitory transmission. Both common carotid arteries were occluded for 3 min with microaneurysmal clips. The gerbils were treated with topiramate (50, 100, or 200 mg/kg, i.p.) immediately after ischemia. Neuronal cell damage in the hippocampal CA1 region was evaluated quantitatively 7 days after ischemia. Topiramate at the dose of 50 mg/kg failed to reduce hippocampal neuronal damage. However, topiramate when administered at the dose of 100 or 200 mg/kg significantly reduced hippocampal neuronal damage in dose-dependent manner (P<0.001 and P<0.0005, respectively). These results suggest that topiramate has a neuroprotective effect against neuronal damage following global ischemia in the gerbils. PMID- 10704774 TI - The spatial pattern of the vacuolation in patients with sporadic Creutzfeldt Jakob disease. AB - 1600 microm in diameter and, in the majority of tissue sections, the vacuole clusters were distributed with regular periodicity parallel to the tissue boundary. The size of the vacuole clusters was positively correlated with patient age in the lower laminae of the occipital cortex and the inferior temporal gyrus (ITG) and negatively correlated with age in the hippocampus. In addition, the size of the vacuole clusters was positively correlated with disease duration in the upper laminae of the ITG. The size and distribution of the vacuole clusters suggests that the vacuolation in CJD reflects the degeneration of specific brain pathways and supports the hypothesis that prion pathology may spread through the brain along well defined anatomical pathways. PMID- 10704776 TI - GABAergic projection from the arcuate nucleus to the supraoptic nucleus in the rat. AB - Electrical stimulation of the neurones in the hypothalamic arcuate nucleus results in a transient inhibition followed by a marked post-stimulus excitation of magnocellular neurones of the supraoptic nucleus. Microdialysis administration of the gamma-aminobutyric acid agonist (GABA(A)), muscimol, directly into the supraoptic nucleus inhibited both oxytocin and vasopressin neurones and these actions were fully reversed by the GABA(A) antagonist bicuculline. In addition, bicuculline administration blocked the inhibition induced by arcuate stimulation, but had no effect on the post-stimulus excitation. Thus, part of the inhibitory pathway arising from or passing through the arcuate nucleus to the supraoptic nucleus is mediated by the neurotransmitter GABA. However, the post-inhibitory excitation induced by arcuate stimulation is not a rebound response, but appears to involve an independent excitatory pathway. PMID- 10704775 TI - Doxapram accentuates white matter injury in neonatal rats following bilateral carotid artery occlusion. AB - The effect of the respiratory stimulant, doxapram, on white matter damage was investigated in neonatal rats under cerebral ischemia. Five-day-old rats underwent bilateral carotid artery occlusion with or without 50 mg/kg i.p. of doxapram. Their brains were neuropathologically examined 48 h later. Doxapram induced about a 20% decrease of PCO(2) for 90 min, but did not cause any neuropathological abnormalities. Bilateral carotid artery occlusion resulted in mild cerebrocortical lesions in 67% of pups, and white matter lesions in the internal capsule in 44%. Doxapram, in addition to bilateral carotid artery occlusion, produced more severe white matter injury in the internal capsule (injury score; 0.67+/-0.87 vs. 1.70+/-0.48, P<0.05) and in the subcortical white matter (0.33+/-0. 67 vs. 1.10+/-0.54, P<0.05). These results demonstrated that the use of doxapram under an ischemic condition accentuates white matter damage in neonatal rats. PMID- 10704777 TI - Inducement of fluent speech in persons who stutter via visual choral speech. AB - A novel phenomenon of fluency enhancement via visual gestures of speech in the absence of traditional auditory feedback is reported herein. The effect on visual choral speech on stuttering frequency was investigated. Ten participants who stuttered recited memorized text aloud under two conditions. In a visual choral speech (VCS) condition participants were instructed to focus their gaze on the face, lips and jaw of a research assistant who 'silently mouthed' the text in unison. In a control condition, participants recited memorized text to the research assistant who sat motionless. A statistically significant (P=0.0025) reduction of approximately 80% in stuttering frequency was observed in the VCS condition. As visual linguistic cues are sufficient to activate the auditory cortex, one may speculate that VCS induces fluency in a similar yet undetermined manner as altered auditory feedback does. PMID- 10704778 TI - Distribution of luteinizing hormone-releasing hormone in the upper brainstem and diencephalon of the cat: an immunocytochemical study. AB - The distribution of luteinizing hormone-releasing hormone (LH-RH)-immunostained cell bodies and fibres was studied in the brainstem and diencephalon of the cat using an indirect immunoperoxidase technique. The brainstem and the thalamus were devoid of immunostained cell bodies, whereas in the hypothalamus immunopositive perikarya were observed in the supraoptic nucleus, the anterior hypothalamus, the preoptic region and in the arcuate nucleus. Our findings also showed that the hypothalamus is richer in immunostained fibres, and that in this region such fibres are more widely distributed than in the thalamus and upper brainstem. No immunopositive fibres were observed in the lower brainstem. Our results point to a more widespread distribution of LH-RH-immunostained perikarya in the cat hypothalamus than that previously reported in the cat; a similar distribution to that found in the rat, and a more restricted distribution than in primates. Additionally, our study shows a more widespread distribution of immunostained fibres in the cat brainstem and diencephalon than that previously described for other mammals. In this context, our results describe for the first time in the mammals central nervous system fibres containing LH-RH located in the stria medullaris of the thalamus, the supramammillary decussation, the laterodorsal and lateroposterior thalamic nuclei, the nucleus reuniens, the supraoptic nucleus, and the optic chiasm. Thus, our findings reveal that LH-RH-immunostained structures are widely distributed in the upper brainstem and in the diencephalon of the cat, suggesting that the peptide may be involved in several physiological functions. PMID- 10704779 TI - A multielectrode array for intrafascicular recording and stimulation in sciatic nerve of cats. AB - The feasibility of implanting an array of penetrating electrodes into peripheral nerves is studied in acute experiments in the cat sciatic nerve. A novel, silicon based array of microelectrodes, the Utah Electrode Array, was used, which contains 25 or 100 1-mm long electrodes that project out from a silicon substrate. Electrode arrays of this complexity, when inserted in the peripheral nerve, could cause significant compression of the nerve and block the conduction of action potentials. Using a high velocity insertion technique, the electrode array was implanted into the sciatic nerve. Compound action potentials were evoked by and recorded with cuff electrodes. Compound action potentials recorded 1 h after insertion were only slightly altered from those recorded before insertion. Single units were readily extracted from evoked multiunit neural recordings in response to cutaneous stimulation and limb rotation around joints. Current injections into the nerve through the electrodes evoked muscle twitches with currents in the 10 microA range. Recording and stimulation stability were demonstrated for periods of up to 60 h. We have shown that high density arrays of electrodes can be inserted into the peripheral nerve and can provide a stable recording and stimulating interface to individual peripheral nerve axons. Such an array may be useful in future neuroscience research and potential neuroprosthetic applications. PMID- 10704780 TI - A cautionary note: the actions of adenosine agonists and antagonists may be reversed under certain conditions in primary cultures. AB - It is now generally accepted that adenosine has a neuroprotective role in the central nervous system. Agonists of adenosine such as 2-chloroadenosine (2-ClA) have been shown to be neuroprotective, while antagonists such as 8 phenyltheophylline (8-PT) increase neurotoxicity. However, paradoxical results have been reported with adenosine analogues, especially with respect to length of time of administration. We observe similarly contradictory findings with respect to 2-ClA and 8-PT actions in primary hippocampal cultures exposed to glutamate or kainic acid. We found 8-PT and 2-ClA had antagonist and agonist actions, respectively, only with acute (1 h) treatment; with chronic treatment (24 h), 2 ClA had no effects, while 8-PT had significant agonist actions. We also show that with variations in the type of culturing system, concentration, and pH that 8 PT's neurotoxic antagonist actions could be dramatically changed. We, therefore, present this paper as a cautionary note in experimenting with adenosine analogues. PMID- 10704781 TI - In vivo electron spin resonance spectroscopy on signal decay of intrastriatal nitroxide radical after acute administration of haloperidol in rats. AB - Sequential changes in the electron spin resonance (ESR) signal intensity of nitroxide radical perfused in the striatum of rats treated with haloperidol (HPD) were evaluated using a 700-MHz ESR spectrometer. Nitroxide radical was perfused in the striatum by in vivo microdialysis. Nitroxide used was 3-carbamoyl-2,2,5, 5 tetramethylpyrrolidine-1-oxyl. Following 6-h perfusion of the nitroxide radical by dialysis at the rate of 2 microl/min through the radical introducer that had been stereotaxically implanted in the rat's striatum, HPD or saline was injected intraperitoneally into the rats in the resonator. The sequential changes in the ESR spectrum of the nitroxide radical were then evaluated. Spectra were successively observed in all animals. The half-life, which was estimated on the basis of the exponential decay in signal intensity, was used as a parameter of decay rate of the ESR signal intensity of nitroxide radical. The half-life in the rats injected with HPD was significantly longer than that in controls. This finding suggests that the reducing ability of the striatal extracellular space of a rat acutely treated with HPD was decreased in comparison with that of the control. PMID- 10704782 TI - Hippocampal norepinephrine-like voltammetric responses following infusion of corticotropin-releasing factor into the locus coeruleus. AB - Intracerebroventricular (i.c.v.) administration of corticotropin-releasing factor (CRF) increases the activity of noradrenergic neurons in the locus coeruleus (LC) assessed by electrophysiological and neurochemical studies. It has been suggested that this effect of i.c.v. CRF is exerted directly on LC noradrenergic (LC-NE) neurons. Infusion of CRF directly into the LC increases cortical and hippocampal release of norepinephrine (NE) as indicated by in vivo microdialysis studies, but the electrophysiological studies have shown both increases and decreases. The present study used in vivo voltammetry to study changes in the extracellular concentrations of NE in the rat hippocampus in response to infusion of CRF (100 ng) into the LC. When the infusion cannula was located in or very close to the LC, the immediate response to CRF was a small decrease in the NE-like oxidation current, followed by a robust increase after about 6-7 min. The oxidation current reached a peak around 13 min and returned to baseline by about 30 min after CRF infusion. By contrast with CRF, infusion of glutamate into the LC increased the oxidation current with a delay of around 30 s and a peak within 90 s. The responses to LC infusion of CRF in rats treated with DSP-4 to deplete hippocampal NE were substantially smaller than those in untreated rats, suggesting that the oxidation signals in untreated rats reflected changes in concentrations of NE. The response to glutamate was markedly augmented by pretreatment with the NE reuptake inhibitor, desmethylimipramine, suggesting that the observed responses reflected changes in NE. Infusion of the same dose of CRF into brain structures outside the LC did not elicit consistent changes in oxidation current in the hippocampus. The time course of the responses to CRF is compatible with previously reported electrophysiological responses of LC-NE neurons to CRF and with neurochemical evidence indicating that CRF can affect the activity of LC-NE neurons. The results indicate that CRF may act in or close to the LC to induce release of hippocampal NE, but the delayed response to CRF compared with that to glutamate, suggests that CRF does not directly activate LC-NE neurons. PMID- 10704783 TI - Increase in plasma adenosine during brain ischemia in man: a study during transient ischemic attacks, and stroke. AB - Adenosine is a "retaliatory metabolite" which accumulates during experimental brain ischemia and has vasodilatory and putative neuroprotective effects. The aim of this study was to assess whether human cerebral ischemia and necrosis evaluated in the clinical models of transient ischemic attack (TIA) and stroke, respectively-acutely raise plasma adenosine levels. We studied 20 patients: 10 with TIA and 10 with stroke. In all, blood was serially sampled for assessment of plasma adenosine by an high-performance liquid chromatography method. Sampling occurred on peripheral blood during TIA and stroke upon admission, and serially thereafter every day up to 7 days and every other day up to 20 days. We found that in TIA and stroke patients, peripheral adenosine levels were increased to a similar extent upon admission (TIA = 264 +/- 53 vs. stroke = 257 +/- 60 nM, p = ns), peaked on the day 2 for TIA (300 +/- 60) and on day 3 for stroke (289 +/- 43) patients, and steadily decreased towards the normal range, reached by all TIA patients by day 5 and by stroke patients by day 15. Stroke and TIA are associated with a rapid increase in circulating plasma adenosine concentration in man, detectable in peripheral vein. The adenosine surge likely mirrors an increased production from the ischemic brain, and it lasts days (for TIA) and weeks (for stroke) after the acute event. PMID- 10704784 TI - Age and insulin-like growth factor-1 modulate N-methyl-D-aspartate receptor subtype expression in rats. AB - N-Methyl-D-aspartate (NMDA) receptors have been reported to have an important role in synaptic plasticity and neurodegeneration. Two major subtypes of these receptors, NMDAR1 and NMDAR2, are present in brain and heterogeneity of these receptors have been reported to define specific functional responses. In this study, the effects of age and chronic insulin-like growth factor-1 (IGF-1) administration on NMDA receptor density and subtype expression were investigated in frontal cortex, CA1, CA2/3 and the dentate gyrus of the hippocampus of young (10 months), middle-aged (21 months) and old (30 months) male Fisher 344xBrown Norway (F1) rats. No age-related changes in (125)I-MK-801 binding or NMDAR1 protein expression were observed in hippocampus or frontal cortex. However, analysis of NMDAR2A and NMDAR2B protein expression in hippocampus indicated a significant decrease between 21 and 30 months of age and administration of IGF-1 increased these receptor subtypes. In cortex, NMDAR2A and NMDAR2B protein expression were not influenced by age or IGF-1 treatment, although NMDAR2C protein expression decreased with age and this decline was not ameliorated by IGF 1 administration. These data demonstrate that NMDA receptor subtypes are altered with age in a regional and subtype specific manner. We conclude that both age and IGF-1 regulate the expression of NMDA receptor subtypes and suggest that age related changes in NMDA receptor heterogeneity may result in functional changes in the receptor that have relevance for aging. PMID- 10704786 TI - Sodium overload through voltage-dependent Na(+) channels induces necrosis and apoptosis of rat superior cervical ganglion cells in vitro. AB - Using the failure to exclude trypan blue as a criterion for cell death, we found that veratridine, the voltage-dependent Na(+) channel activator, exerted its toxicity to cultured sympathetic neurons in a dose-dependent manner (half-maximal toxicity occurred at 2 microM). The co-presence of tetrodotoxin completely reversed the toxicity only at concentrations of veratridine < 20 microM. Veratridine neurotoxicity was due to the influx of Na(+); a medium low in Na(+) (36 mM) completely abolished its neurotoxicity, whereas a Ca(2+)-free medium did not attenuate its neurotoxicity. Furthermore, the buffering action of 1, 2-Bis-(2 aminophenoxy)ethane-N,N,N',N',-tetraacetate (BAPTA) on veratridine-induced increase in intracellular Ca(2+) levels neither blocked veratridine neurotoxicity in normal medium, nor attenuated the low Na(+) effect. Elevated K(+) effectively blocked veratridine neurotoxicity in a Ca(2+)-dependent manner. Cytoplasmic pH measurements using a fluorescent pH indicator demonstrated that cellular acidification (from pH 7.0 to pH 6.5) occurred upon treatment with veratridine. Both veratridine-induced acidification and cell death were ameliorated by 5-(N ethyl-N-isopropyl)amiloride, the specific inhibitor of the Na(+)/H(+) exchanger (IC(50) = 0.5 microM). Finally, necrosis occurred predominantly in veratridine neurotoxicity, but both staining with bis-benzimide and TUNEL analysis showed nuclear features of apoptosis in sympathetic neurons undergoing cell death. PMID- 10704785 TI - Brain AT(2) receptor mediate vasodepressor response to footshocks: role of kinins and nitric oxide. AB - In the present study the role of brain AT(2) receptor in the cardiovascular response to stress was investigated in conscious rats. Footshock-stress increased mean arterial pressure (MAP) and heart rate (p < 0.0001). Intracerebroventricular (i.c.v.) administration of losartan (100 microg/5 microl), a specific angiotensin AT(1) receptor antagonist, not only attenuated the pressor response to footshocks, but also resulted in a consistent vasodepressor response (-10 mmHg, p < 0.02). Meanwhile, heart rate response was not altered. Given alone, PD 123319 (3 microg/5 microl, i.c.v.), a specific angiotensin AT(2) receptor antagonist, did not alter the hemodynamic response to footshocks. However, simultaneous block of brain AT(1) and AT(2) receptors by combined administration of losartan and PD 12319, eliminated the vasodepressor response unmasked after footshocks in rats i.c.v.-pretreated with losartan alone. In addition, we studied the role of brain kinins and nitric oxide (NO) in the vasodepressor response observed after footshocks in losartan i.c.v.-treated rats. Intracerebroventricular administration of icatibant (20 pmol/5 microl), a selective B(2) receptor antagonist, or N(G)-nitro-L-arginine methyl ester (100 microg/5 microl), a selective NO-synthase inhibitor, abolished the vasodepressor response to footshocks in losartan-treated rats. Our data suggest that the vasodepressor response to footshocks in the presence of AT(1) antagonist is triggered by activation of AT(2) receptor. Brain NO and kinins appear to contribute in this effect. PMID- 10704787 TI - Resolution of two [(35)S]GTP-gamma-S binding sites and their response to chronic morphine treatment: a binding surface analysis. AB - The mechanisms by which prolonged exposure to morphine leads to tolerance are not fully understood. We investigated the effects of etorphine (ET) on [(35)S]guanosine 5'-(-thio)-triphosphate ([(35)S]GTP-gamma-S) binding in brains of rats made tolerant to morphine via the implantation of morphine (or placebo) pellets. Binding surface analysis was used to characterize the interactions of ET, Gpp(Np)H and GTP-gamma-S with sites labeled by [(35)S]GTP-gamma-S. Data sets were fitted to one- and two-site binding models using the nonlinear least squares curve fitting program MLAB-PC (Civilized Software, Bethesda, MD, USA). Two binding sites were readily resolved. Chronic morphine significantly increased the B(max) and K(d) of the high affinity binding site. ET stimulated [(35)S]GTP-gamma S binding in placebo membranes via an increase in the B(max) of the high affinity binding site. In contrast, ET stimulated [(35)S]GTP-gamma-S in chronic morphine membranes via a large decrease in the K(d) of the high affinity site. These results suggest that chronic morphine treatment alters the mechanism by which ET stimulates [(35)S]GTP-gamma-S binding to G-proteins. Since proper G protein/receptor coupling increases [(35)S]GTP-gamma-S binding via an increase in B(max) values, these results suggest that opioid receptors in chronic morphine membranes are not normally coupled to G-proteins. These findings corroborate earlier studies that reported changes in G-protein function in morphine tolerant animals. PMID- 10704788 TI - Mechanisms by which cyclodextrins modify drug release from polymeric drug delivery systems. AB - For many drug candidates a modified in vivo drug release is desired to improve efficacy, sustain effect or minimise toxicity. Polymeric delivery systems, such as microspheres, nanospheres and polymeric films, have been extensively researched in an attempt to achieve modified drug release. Cyclodextrins offer an alternative approach. These cyclic oligosaccharides have the ability to form non covalent complexes with a number of drugs and in so doing alter their physicochemical properties. In addition, the primary and secondary hydroxyl groups of the native (alpha, beta, gamma-) cyclodextrins are potential sites for chemical modification. It follows that the incorporation of these agents into polymeric drug delivery systems, as physical mixtures, covalently bound conjugates or cross-linking agents, frequently permits a greater degree of control of drug release. This paper reviews the incorporation of various cyclodextrins into polymeric formulations. The mechanisms by which cyclodextrin/polymer formulations act to modify drug release are considered. PMID- 10704789 TI - Carrier-enhanced human growth hormone absorption across isolated rabbit intestinal tissue. AB - Small molecular weight alpha acid derivatives are able to enhance the intestinal absorption of human growth hormone through isolated rabbit intestinal tissue. The enhancement is not through the usual tissue modification associated with traditional penetration enhancers nor is it through an active transport process. Rather these small molecules associate with human growth hormone in solution to make it more transportable through intestinal tissue. It is shown that the enhancer has specificity for a particular protein and the enhancer and human growth hormone must be in solution together to be effective, i.e. pretreating the tissue with enhancer and then adding the protein does not increase tissue permeability. Moreover, the enhancer does not increase the permeability of mannitol or progesterone, thus providing additional evidence of specificity and establishing that these agents are not classical penetration enhancers. PMID- 10704790 TI - A mechanism on the drug release into a perfect sink from a coated planar matrix with a super-saturation loading in the core. AB - A comprehensive model is proposed to accurately describe drug release kinetics from a coated plane sheet when drug loading in the core is above its saturation level. The general solutions are acquired in a dimensionless form by the Laplace transform and the solution for the special case-a perfect sink condition, is derived from these general ones. On the basis of the model calculations, the effects of the diffusion ratios and thickness ratios of the coatings to the core, and drug loading entrapped in the core during the release processes have been discussed over wide range of variables. To validate the model equations proposed, the coated systems, 5-fluoracilum/ethylene-vinyl alcohol copolymer (5-Fu/EVAL) core matrix coated with various polymeric materials such as EVAL, cellulose acetate (CA), poly(2-hydroxyethyl methacrylate) (PHEMA) and poly( methyl methacrylate) (PMMA), having different diffusivities, are designed, experimentally investigated and graphically and quantitatively compared with the theoretical values. The results show a good correlation between the theory and experiment. PMID- 10704791 TI - Interactions during aqueous film coating of ibuprofen with aquacoat ECD. AB - During the development of a coated ibuprofen formulation a sticking tendency occurred when applying Aquacoat ECD. This interaction indicated the formation of a eutectic mixture. The compatibility of the components of Aquacoat ECD with ibuprofen was investigated by differential scanning calorimetry. Cetyl alcohol, a stabilizing excipient in Aquacoat, was found to form a eutectic system with ibuprofen. It was characterized by the construction of a phase diagram with 33 mol% ibuprofen and an onset temperature of 40.5 degrees C. Wide-angle X-ray diffraction was used to identify the polymorphic forms of cetyl alcohol. The results confirmed the amorphous state in the aqueous dispersion in contrast to the beta(0)- and gamma(4)-polymorphs of solid cetyl alcohol. PMID- 10704792 TI - The influence of lactose carrier on the content homogeneity and dispersibility of beclomethasone dipropionate from dry powder aerosols. AB - Dry powder formulations for inhalation usually comprise a mixture of coarse lactose (CL), employed as a carrier, and micronized drug. It was the aim of this study to determine the effects of fine lactose (FL), blended as a tertiary component on the mixing homogeneity and dispersibility of a model hydrophobic drug, beclomethasone dipropionate (BDP). BDP particles (volume median diameter (VMD) 4.6 microm) existed mainly as agglomerates, the majority of which were not dispersed into primary particles after aerosolization at a high shear force (4.7 psi). The resultant particle size distribution of BDP was multi-modal with VMD varying between 4.7 and 30.2 microm. Ternary interactive mixtures were prepared to consist of CL, FL and BDP with a fixed ratio of lactose to BDP of 67.5:1 w/w, but two concentrations of FL, i.e. 2.5 and 5%, w/w. The mixing was carried out using different sequences of adding the three components for two mixing times (15 and 60 min). Binary mixtures composed of CL and BDP were prepared for both mixing times as the controls, and these exhibited a coefficient of variation (COV) in BDP content <= 5%. Addition of FL to the binary formulations greatly reduced the content uniformity of BDP if the final powder were prepared by first mixing CL with FL before mixing with the drug (COV>20%, after mixing for 15 min). However, the mixtures, prepared using other mixing sequences, had a similar uniformity of BDP content to the binary mixtures. All ternary mixtures containing 2.5% FL consistently produced a significantly higher (ANOVA P<0.01) fine particle fraction (FPF, 3.1--6.1%) and fine particle dose (FPD, 13.6--30.1 microg) of BDP than the binary mixtures (FPF, 0.3-0.4%; FPD, 1.6-2.1 microg) after aerosolization at 60 l min(-1) via a Rotahaler into a twin stage liquid impinger. The mixing sequences exerted a significant (P<0.05) effect on the dispersion and deaggregation of BDP from the formulations prepared using a mixing time of 15 min but such an effect disappeared when the mixing time was lengthened to 60 min. The dispersibility of BDP was always higher from the ternary mixtures than from the binary mixtures. BDP delivery from dry powder inhalers was improved markedly by adding FL to the formulation, without substantial reduction in the content uniformity of the drug. PMID- 10704793 TI - Chitosans as nasal absorption enhancers of peptides: comparison between free amine chitosans and soluble salts. AB - A total of three free amine chitosans (CS J, CS L and CS H) and two soluble chitosan salts (CS G and CS HCl) were evaluated for their efficacy and safety as nasal absorption enhancers of peptides based on in situ nasal perfusion and subacute histological evaluation in rat. At 0.5% w/v, all chitosans were effective in enhancing the nasal absorption of [D-Arg(2)]-Kyotorphin, an enzymatically stable opioid dipeptide. The enhancing effect of the free amine chitosans increased as the pH was decreased from 6.0 to 4.0 (P<0.05). However, the pH effect was not significant for the two chitosan salts (P0.05), suggesting that their adjuvant activity may be less pH-dependent than the free amine form. CS J and CS G were subsequently selected for further studies. At only 0.02% w/v, their enhancing effect was already significant and comparable to that of 5% w/v hydroxypropyl-beta-cyclodextrin (HP-beta-CD). Both chitosans at 0.1% caused minimal release of total protein and phosphorus from the rat nasal mucosa, with the values similar to that of 5% HP-beta-CD. At 0. 5% the two chitosans also stimulated smaller release of lactate dehydrogenase, an intracellular enzyme used as marker of nasal membrane damage, than 1.25% dimethyl-beta-cyclodextrin. Morphological evaluation of the rat nasal mucosa following 2-week daily administration indicated that the two chitosans (1.0%) produced only mild to moderate irritation. In conclusion, both the free amine and the acid salt forms of chitosans are effective in enhancing the nasal absorption of [D-Arg(2)] Kyotorphin and have potential for further studies as a safe and effective nasal absorption enhancer of peptide drugs. PMID- 10704794 TI - Transdermal iontophoretic delivery of timolol maleate in albino rabbits. AB - The use of transdermal iontophoresis is a promising technique for the systemic delivery of water soluble and ionic drugs of relatively large molecular size. The present study investigates the skin pre-treatment with Azone (laurocapram) and iontophoresis on the pharmacodynamic effect of timolol maleate (TM) in vivo in albino rabbits. The pharmacodynamic effect of TM was evaluated by transdermal delivery and compared with an intravenous (i.v.) administration. Iontophoresis of TM (0.1 mg/ml) produced a significant inhibition in the isoprenaline (ISP) induced tachycardia. Iontophoresis with higher concentration of TM (1 mg/ml) produced a 100% inhibition of the ISP induced tachycardia. Pre-treatment of skin with Azone (3% v/v emulsion) eliminated the lag time and prolonged the duration of action of iontophoresis from 4 to 6 h. The AUC of Azone treated group was significantly higher than that of the untreated group (P<0.05). Further, the AUC with iontophoretic delivery and pre-treatment of Azone was comparable to that of intravenous TM (30 microg/kg). In conclusion, iontophoresis in combination with Azone can increase the transdermal delivery of TM, whereby the required transport rate can be achieved with a lower drug concentration. PMID- 10704795 TI - The dermal delivery of lignocaine: influence of ion pairing. AB - The purpose of the present study was to determine the significance of ion pairing on the permeation of lignocaine. Results of diffusion studies through polydimethylsiloxane (PDMS) at different pH values 4. 0, 6.0, 7.0, 8.0 indicated that lignocaine hydrochloride (L-HCl) flux significantly increased with the amount of unionized base. In order to see if similar results could be obtained using human skin, permeation runs were performed with human skin at pH of 4.0, 5.5 and 7.0. These values were chosen to simulate an appropriate range of physiological conditions. Results of the experiments with human epidermis showed increasing L-HCl flux with increasing pH, confirming the trends seen with PDMS membranes. A linear relationship was found between the apparent partition coefficient and the steady state flux. Further experiments were conducted at donor pH 4.0 to minimise the contribution of the unionized species. Although an excess of different ions such as nitrate, mesylate and bromide increased the apparent partition coefficient, the steady state flux was not significantly increased. The steady state lignocaine flux was increased up to 2.45-fold using different counter ions. The highest flux was measured from lignocaine morpholinopropane sulfonate (L-mps). It is possible to enhance the flux of salts across lipophilic membranes by using an ion pair approach. The degree to which this is possible depends on the lipophilicity of the counter ion, the medium in which the ion pair forms, and the ionic strength. PMID- 10704796 TI - Effect of buffer pH, buffer concentration and skin with or without enzyme inhibitors on the stability of [Arg(8)]-vasopressin. AB - The stability of [Arg(8)]-vasopressin (AVP) as a function of buffer pH, buffer concentration, salt concentration, temperature, and skin with and without enzyme inhibitors was investigated. AVP was analyzed by reverse-phase high-performance liquid chromatography. The results indicated that the buffer's pH affected the degradation rate of AVP. Buffer ions (H(2)PO(4)(-) and HPO(4)(2-)) and salt concentrations had no effect on the degradation of AVP. Maximum stability was achieved at pH 3.35 among pH values tested. The activation energy for the overall reaction was 21.5 kcal mol(-1) at pH 3.35. From the Arrhenius equation, the shelf life of AVP at 25 degrees C and pH 3.35 was calculated to be 1.38 years. The degradation rate of AVP in the skin (area: 9 cm(2), thickness: 0.5 mm) was 0.22 h(-1). Bestatin (an aminopeptidase inhibitor) had the best stabilizing effect on the degradation of AVP by skin among the three enzyme inhibitors (i.e. aprotinin, bestatin, and leupeptin) studied. The degradation rate of AVP in the skin was reduced to 0. 059 h(-1) in the presence of bestatin in comparison with no inhibitor (0.22 h(-1)). PMID- 10704797 TI - Improved compression properties of propyphenazone spherical crystals. AB - Spherical propyphenazone crystals were produced by an agglomeration technique using a three solvents system. After selecting the best propyphenazone solvent (ethyl alcohol), non-solvent (demineralized water) and bridging liquid (isopropyl acetate), several of their ratios were tested by a Sheffe ternary diagram. Micromeritic properties of agglomerates such as flowability, were improved and their compression behavior was investigated and compared to that of raw crystals. By compression and densification studies, along with tablet SEM analysis, we have been able to explain the compression mechanism of propyphenazone spherical crystals and have shown that their better tablet/ability can be due to the small size of individual particles in the agglomerates PMID- 10704798 TI - Evaluation of low-substituted hydroxypropylcelluloses (L-HPCs) as filler-binders for direct compression. AB - The aims of this study were to assess the potential value of low-substituted hydroxypropylcelluloses (L-HPCs) as excipients of direct compression, and to investigate relationships between the chemical and physical properties of the polymers and (a) the powder rheological behavior and (b) drug release profiles from direct compressed tablets elaborated with (1:1) theophylline:L-HPC mixtures. Experiments were performed with five L-HPC varieties of different nominal particle sizes and degree of substitution. The products were characterized with regard to the moisture content, density, IR and Raman spectroscopy, hydroxypropyloxy content, heat of hydration, particle size, specific surface and porosity, and important differences were found in relation with all these properties. The differences in specific surface principally determine the flow and compaction properties of the powders, and the mechanical and microstructural properties of the tablets. The control of the hydroxypropyloxy content and the particle size of the L-HPCs allow the theophylline release profile to be regulated. PMID- 10704799 TI - Methods of studying aging and stabilization of spray-congealed solid dispersions with carnauba wax. 1. Microcalorimetric investigation. AB - Rapidly cooled materials are often unstable as a result of changes in their physical properties due to imperfect crystallization. In the process of spray congealing, melted material is atomized into droplets which very quickly solidify. This increases the possibility of the material crystallizing in different metastable forms. In this study it is shown that isothermal microcalorimetry can be used to observe the change in the thermodynamic state of spray-congealed carnauba wax during storage. In order to accelerate the thermodynamic change in the spray-congealed wax, three annealing procedures have been developed and compared using isothermal microcalorimetry. By means of annealing, a spray-congealed product closer to a thermodynamically stable state has been achieved. PMID- 10704800 TI - Dependency of cyclosporine tissue distribution and metabolism on the age and gender of rats after a single intravenous dose. AB - In a previous study we demonstrated the dependency of cyclosporine (CyA) pharmacokinetics on the age and gender of Wistar rats given 10 mg/kg intravenously. The present study has been conducted under the same experimental conditions (10 mg/kg as a single intravenous dose) to identify the mechanisms behind such differences. On the one hand, drug distribution was studied by measuring the CyA levels in blood, liver, kidney, spleen, adipose tissue, skin and muscle at 48 h post-treatment by using a specific fluorescence polarization immunoassay (m-FPIA, Abbott Laboratories). Drug blood and tissue levels in male rats were significantly higher than the female counterparts except for adipose tissue where the concentrations were 2-fold higher in females. In males, the highest CyA concentrations were observed in the liver, followed in rank order by kidney and spleen, fat, skin, muscle, then blood. On the contrary, females showed the highest drug levels in fat, followed by liver, kidney, spleen, skin, muscle and blood. Age exerted a significant influence on CyA tissue levels in males but no effect was observed in females. The potential differences in drug metabolism were established by measuring (HPLC) the amounts of CyA and its metabolites accumulated in faeces after hepatic biotransformation and biliary excretion. The amounts of circulating metabolites in blood as well as those accumulated and excreted in the liver and urine were also estimated by using specific (m-FPIA) and non-specific fluorescence polarization immunoassay (p-FPIA, Abbott Laboratories), respectively. The analysis of faeces revealed that AM9 was the major identified metabolite with females excreting lower amounts of unchanged CyA than males. In addition, the comparison of the AUC values corresponding to parent CyA and total CyA derivatives suggested that blood concentrations of CyA metabolites were higher in females indicating higher biotransformation rates. Therefore, both CyA distribution and metabolism are responsible for the sex associated differences in drug pharmacokinetics previously found in rats. PMID- 10704801 TI - Determination of optimal combination of surfactants in creams using rheology measurements. AB - The effect of surfactant on the rheological properties of some cream formulations was studied. Two surfactants from two different series were combined to determine the combination which yielded the most viscoelastic structure for creams. The surfactants were the soybean derivatives soya sterol, polyethylene glycol 10 soya sterol and polyethylene glycol 25 soya sterol and the sorbitol derivatives sorbitan monooleate and sorbitan trioleate. The rheological properties of the creams were studied using oscillation stress sweep, oscillation frequency sweep and viscosity tests. Droplet size distribution and conductivity of the creams were also determined. The combination polyethylene glycol 10 soya sterol and sorbitan trioleate yielded the most viscoelastic structure with linearly viscoelastic behaviour. PMID- 10704802 TI - Investigation of the chemical stability of an erythromycin-tretinoin lotion by the use of an optimization system. AB - A combination of 2% erythromycin and 0.05% tretinoin in an alcohol-isopropanol lotion was prepared. Two parameters were investigated for their influence on the stability of erythromycin and/or tretinoin, namely pH and the concentration of butylhydroxytoluene (BHT) as antioxidant. To investigate these two parameters, an optimization technique was used with two factors (pH and concentration of BHT) at two levels. Accelerated stability analysis was performed at 45 degrees C in the dark to exclude isomerization of tretinoin. To analyse erythromycin and tretinoin in the combination preparation, a TLC method, previously developed in the laboratory, was used. The degradation of erythromycin seemed to be much faster than the tretinoin degradation. Optimal stability is shown in the pH range of 8.2 8.6 for erythromycin and 7.2-8.2 for tretinoin while the concentration of BHT had no significant influence. PMID- 10704804 TI - Effect of pectinolytic enzymes on the theophylline release from pellets coated with water insoluble polymers containing pectin HM or calcium pectinate. AB - Theophylline pellets were coated with cellulosic (Aquacoat ECD 30, Surelease clear) or acrylic (Eudragit NE30D, RS30D) polymer aqueous dispersions, containing 10% (related to the insoluble polymer content) of pectin HM or calcium pectinate, using a Uni-Glatt fluidized-bed coating apparatus. When commercial pectinolytic enzymes were added to the dissolution media (0.05 M acetate - phosphate buffer, pH 6.0), the release of theophylline from the coated pellets was generally slower than that observed in the media without enzymes. The enzymatic slowing down of the drug release, depending on the type of the aqueous polymer dispersion used, is more important with mixed Eudragit NE/calcium pectinate coated pellets. The results obtained have been examined with regard to the validity of the approach based on the combination of pectins and the insoluble polymer aqueous dispersions intended for specific-delivery of drugs to the colon. The mechanism of the hydrophilic drug release from pellets coated with insoluble polymer aqueous dispersions containing an aqueous gel-forming polymer has been also discussed. PMID- 10704803 TI - In vitro release studies of methylmethacrylate liberation from acrylic cement powder. AB - Bone cement or polymethylmethacrylate (PMMA) is commonly used for anchoring cemented prosthesis to the bone. Cytotoxic effect of culture media exposed to PMMA powder may be related with long term problems associated with acrylic cement application, being the monomer (methylmethacrylate) one of the cement's component partly responsible for the cytotoxic effect. The present work reports the studies of monomer release from acrylic bone cement powder under different experimental conditions: setting time of PMMA (in solution and air) and different culture media composition. High-performance liquid chromatography was used for the determination of residual monomer. Mathematical models were applied to experimental dissolution data revealing that monomer release is lightly affected by the studied variables. The monomer release seems to be a surface phenomena, suggesting that the possible actions of monomer will mainly be due to the initial loss of non polymerized monomer rather than to further depolymerization of the already polymerized cement. PMID- 10704805 TI - Studies of pectin HM/Eudragit RL/Eudragit NE film-coating formulations intended for colonic drug delivery. AB - Theophylline pellets were coated with Eudragit NE30D aqueous dispersions, containing various pectin HM/Eudragit RL30D ionic complexes, using an Uni-Glatt fluidized-bed apparatus. Dissolution studies were then carried out on the coated pellets at pH 6.0, in absence and in presence of commercial pectinolytic enzymes. The theophylline release from the coated pellets, after an initial latency phase, occurred linearly as a function of time. The theophylline release rate was dependent on the pectin HM content of the complexes incorporated in the coatings. The lowest theophylline release from the coated pellets was obtained when the pectin HM content of the complexes was 20.0% w/w (related to Eudragit RL), i.e. when the complexation between pectin HM and Eudragit RL is optimal. The theophylline release from the coated pellets was slower in presence of the pectinolytic enzymes when the pectin content of complexes is higher than 20% w/w. On the other hand, the effect of the enzymes induced an increase of the theophylline release when the pectin HM content of the coatings ranged between 10.0 and 15.0% w/w (related to Eudragit RL). PMID- 10704807 TI - Epidermal permeability-penetrant structure relationships: 4, QSAR of permeant diffusion across human stratum corneum in terms of molecular weight, H-bonding and electronic charge. AB - Principal components analysis (PCA) and multivariate regression analysis (MRA) are used to assess the predictors of permeant diffusion across human stratum corneum. Log(D/h), was estimated from logk(p)+0.024-0.59 logK(oct), where D=diffusion coefficient (cm(2)/h), h=path length (cm), k(p) permeability coefficient (cm/h), K(oct)=partition coefficient (octanol/water). Molecular weight (MW) with (1) scaled H-bonding parameters alpha and beta, or (2) summed modulus of partial charge from molecular modelling were tested as predictors of (D/h). Charge may be computed for any molecule, whilst alpha and beta values are generally unavailable for molecules of biological interest. PCA suggests a dominant permeation pathway since 93% of data variation is in PC1 of log(D/h), MW and charge and 82% in PC1 of log(D/h), MW, alpha and beta. MRA using MW, alpha and beta is unsatisfactory because of collinearity amongst predictors. The best predictor was the product MW*charge. Similarity of the eigenvectors in PCA and normalised coefficients in MRA indicates that charge and MW are equally important predictors of diffusion. PMID- 10704806 TI - Effect of surfactants on human stratum corneum: electron paramagnetic resonance study. AB - Electron paramagnetic resonance (EPR) spectra of nitroxide spin probes are useful for studying biological membranes and chemical-membrane interactions. Recently, we established a stripping method to remove stratum corneum (SC) for this purpose. To assess this stripping method with EPR and correlate with standard methods, we quantified the irritant effects of three types of surfactants by measurements of visual score and transepidermal water loss (TEWL), SC hydration and chromametry and studied EPR spectra measurements of surfactant-treated cadaver SC (C-SC) and stripped off SC (S-SC) on patch tested sites. 5-Doxyl stearic acid was the spin label. The order parameter S obtained from the spectra of S-SC correlated with those of C-SC and TEWL values. The results suggest that this method is capable of evaluating the fluidity of SC and correlates with the above bioengineering parameters. PMID- 10704808 TI - Pharmacokinetics and in-situ absorption studies of a new anti-allergic compound 73/602 in rats. AB - Compound 73/602 (AA) is a structural analogue of vasicinone, an alkaloid present in the leaves and roots of Adhatoda vasica (Acanthaceae). It possesses potent antiallergic activity in mice, rats and guinea pigs. The pK(a) of AA was determined to be 2.87+/-0. 19 by UV spectrophotometry. The absorption kinetics of this compound were studied in-situ using a rat gut technique at pH 2.6 and 7.4. The rate of absorption at pH 2.6 (0.0288+/-0.004 min(-1)) was slightly less than at pH 7.4 (0.035+/-0.0008 min(-1)). This characteristic behavior was attributed to the low pK(a) of AA, a weekly basic compound, where nearly 35% of the compound remained in the unionized form at pH 2.6. Also, the return of compound into the mucosal lumen from the blood capillaries over a period of 2 h after administering a 2 mg dose in tail vein was less than 0.3%. Hence it was concluded that entero enteric circulation of AA did not contribute significantly to the in-situ absorption rates. Pharmacokinetic parameters of AA were determined in male rats after administering a single 10 mg/kg intravenous dose (i.v.) and 50 mg/kg oral bolus dose. Following i.v. administration the initial decline in serum concentration was rapid with half-life of 20.2 min. After a single oral dose the concentration-time data of AA in rats was best described by a one-compartment model with equal first order absorption and apparent elimination rate constants. The half-life of the decline in serum concentration of AA following oral administration was 50.6 min, indicating absorption rate limiting disposition at the high dose given. Comparison of AUC of oral and i. v. data indicates that only about 60% of the oral dose reach the systemic circulation. PMID- 10704809 TI - Physical stability and in-vitro gene expression efficiency of nebulised lipid peptide-DNA complexes. AB - The lower respiratory tract provides a number of disease targets for gene therapy. Nebulisation is the most practical system for the aerosolisation of non viral gene delivery systems. The aerosolisation process represents a significant challenge to the maintenance of the physical stability and biological activity of the gene vector. In this study we investigate the role of a condensing polycationic peptide on the stability and efficiency of nebulised lipid-DNA complexes. Complexes prepared from the cationic lipid 1, 2-dioleoyl-3 trimethylammonium propane (DOTAP) and plasmid DNA (pDNA) at mass (w/w) ratios of 12:1, 6:1 and 3:1, and complexes prepared from DOTAP, the polycationic peptide, protamine, and pDNA (LPD) at 3:2:1 w/w ratio were nebulised using a Pari LC Plus jet nebuliser. Samples from the nebuliser reservoir (pre- and post-nebulisation) and from the aerosol mist were collected and investigated for changes, including: particle diameter, retention of in-vitro transfection activity and the relative concentration and nature of the complexed pDNA remaining after the nebulisation procedure. The process of jet nebulisation adversely affected the physical stability of lipid:pDNA complexes with only those formulated at 12:1 w/w DOTAP:pDNA able to maintain their pre-nebulisation particle size distribution (145+/-3 nm pre-nebulisation vs. 142+/-2 nm aerosol mist) and preserve significant pDNA integrity in the reservoir (35% of pre-nebulisation pDNA band intensity). The LPD complexes were smaller (102+/-1 nm pre-nebulisation vs. 113+/ 2 nm aerosol mist) with considerably greater retention of pDNA integrity in the reservoir (90% of pre-nebulisation pDNA band intensity). In contrast the concentration of pDNA in the aerosol mist for both the 12:1 w/w DOTAP:pDNA and LPD complexes were significantly reduced (10 and 12% of pre-nebulised values, respectively). Despite reduced pDNA concentration the transfection (% cells transfected) mediated by aerosol mist for the nebulised complexes was comparatively efficient (LPD aerosol mist 26 vs. 40% for pre-nebulised complex; the respective values for 12: 1 w/w DOTAP:pDNA were 12 vs. 28%). The physical stability and biological activity of nebulised lipid:pDNA complexes can be improved by inclusion of a condensing polycationic peptide such as protamine. The incorporation of the peptide precludes the use of potentially toxic excesses of lipid and charge and may act as a platform for the covalent attachment of peptide signals mediating sub-cellular targetting. PMID- 10704810 TI - Gene expression in an intact ex-vivo skin tissue model following percutaneous delivery of cationic liposome-plasmid DNA complexes. AB - The skin represents an attractive site for the localised gene therapy of dermatological pathologies and as a potential antigen bioreactor following transdermal delivery. Potential also exists for the gene therapy of skin as a cosmetic intervention. The most exploited non-viral gene delivery system involves the complexation of cationic liposomes with plasmid DNA (pDNA) to form lipid:pDNA vectors that protect the DNA from nuclease-mediated degradation and improve transgene-cell interactions. Despite numerous studies examining the potential for these vectors in delivering genes to a variety of keratinocyte models, investigations into the topical application of such complexes to intact skin tissue is limited. This ex-vivo study, conducted with intact skin tissue derived from hairless mice, provides quantitative confirmation that topical administration of cationic lipid:pDNA complexes can mediate uptake and expression of reporter pDNA (33-fold higher compared with control) in viable epidermal tissue. The ex-vivo study design provides for intact skin tissue that has not been subjected to depilatory procedures of potential detriment to stratum corneum barrier function, and can be utilised for the quantitative and efficient examination of a potentially wide range of non-viral gene vectors designed for epidermal expression. PMID- 10704811 TI - Polyamidoamine Starburst dendrimers as solubility enhancers. AB - The solubility of the hydrophobic drug ibuprofen has been compared in an aqueous solution of polyamidoamine (PAMAM) G4 dendrimer and sodium dodecyl sulphate (SDS). The PAMAM G4 dendrimer solution significantly enhanced the solubility of ibuprofen compared to 2% SDS solution. It was found that the solubility of ibuprofen in dendrimer solution was directly proportional to dendrimer concentration and inversely proportional to temperature. The influence of dendrimer solution pH on the solubility enhancement of ibuprofen suggests that it involves an electrostatic interaction between the carboxyl group of the ibuprofen molecule and the amine groups of the dendrimer molecule. PMID- 10704812 TI - Allelic distribution of nine short tandem repeat (STR), HLA-DQA1, and polymarker loci in an Omani sample population. AB - Allele frequency distributions of nine short tandem repeat (STR) loci, D3S1358, vWA, FGA, D8S1179, D21S11, D18S51, D5S818, D7S820, and D13S317, HLA-DQA1 and polymarker (PM) loci were studied in a sample population from Sultanate of Oman, Middle East. Blood samples were collected from 162 unrelated individuals. For all STR, HLA-DQA1 and PM loci, no deviations from Hardy-Weinberg equilibrium, based on the exact test, were observed. The most discriminating loci were D18S51 (PD=0.966) and FGA (PD=0.964), while the least informative locus is GYPA (PD=0.585). The allele frequency data may be useful in forensic case work. PMID- 10704813 TI - Study of the reaction mechanism of 1,8-diazafluoren-9-one with the amino acid, L alanine. AB - This paper describes the study of the reaction between 1, 8-diazafluoren-9-one (DFO) with the amino acid L-alanine in methyl alcohol. Particular interest was paid to the possible role of the solvent which appears to react with the DFO to form a hemiketal that is the reactive species. A potential reaction pathway is proposed based on the current data collected. The reaction intermediates and products were identified by a variety of spectroscopic methods including mass spectrometry (MS), nuclear magnetic resonance (NMR) spectroscopy, and X-ray crystallography. PMID- 10704814 TI - A statistical study of air bubbles on athletic shoesoles. AB - Comparisons of a shoemark with a shoesole (and standards) sometimes lead to associations based on air bubbles (among other manufacturing or acquired characteristics). Today, the assessment of the evidential value of air bubbles coincidences relies largely upon the examiner's experience and/or follows sometimes a verification based on the examination of a small number of analogous pairs collected for the case at hand. Statistical data related to the occurrence and characteristics of air bubbles on shoesoles in an attempt to model the potential variability have been gathered. Seventy-one pairs of shoes with the same design, brand, model and size were obtained. Right and left soles were photographed. An image-processing algorithm was developed to allow the systematic acquisition of data such as: (1) the number of air bubbles on the sole and around given structural elements; (2) the measure of air bubbles characteristics such as their surface and position. These data allow a discussion of the assessment of the probability of finding on shoesoles (same design, brand, model and size) a certain number of air bubbles on a surface with the same positions and morphology. PMID- 10704815 TI - Pulmonary surfactant-associated protein A levels in cadaveric sera with reference to the cause of death. AB - Pulmonary surfactant-associated protein A (SP-A) is an exclusively lung specific protein, and is considered to leak into the blood stream in alveolar septal damage. In this study we examined the serum SP-A level in forensic autopsy materials using an enzyme immunoassay with monoclonal antibodies to assess the postmortem level in relation to the cause and mode of death. Although a gradual postmortem degradation should be taken into consideration, topological relationship of serum level seemed to be fairly stable (arterial> or =venous blood in most cases), indicating no evident influence of postmortem diffusion. Significant elevation of serum SP-A (76.7-250 ng/ml in left heart blood) was observed in hyaline membrane diseases from various causes independent of the postmortem intervals (<30 h). However, mean SP-A levels in postmortem heart blood were usually low in asphyxia including hanging, strangulation and choking (left, 25.5 ng/ml; right, 22.3 ng/ml), polytrauma (left, 13.1 ng/ml; right, 9.0 ng/ml) and stab wound to the neck (left, 34.1 ng/ml; right, 29.4 ng/ml). Prominent elevation was noted in a case of fatal strangulation with complication of idiopathic interstitial pneumonia, and also in some deaths due to drowning, burns in fires, blunt and gunshot chest injuries. These findings indicated that postmortem elevation of serum SP-A levels was associated with alveolar septal damage due to inflammation, mechanical and physical stresses, which caused leakage of SP-A into the bloodstream. PMID- 10704816 TI - Homicide by sharp force in two Scandinavian capitals. AB - In the Oslo and Copenhagen capital areas, 141 homicides by sharp force were committed in the ten-year period 1985-1994. This method accounted for 33% of the homicides in this period. Thirty-five percent of the victims were female, and most of the victims were between 20 and 50 years of age. The majority of the male victims were killed by an acquaintance, the females by their spouse. Sixty-five percent of the male and 37% of the female victims had alcohol in their blood. The majority of the female victims had lesions in 3-4 anatomical regions, while the males most often had lesions in only one anatomical region. Seventy-nine percent of the females and 36% of the males had self-defence injuries in the upper extremities. In 21 cases (15%) the offender was a woman, 19 of their victims being male; the weapon in these cases was most often a kitchen knife. Seventy eight percent of the females and 49% of the males were killed in their own home. The most common circumstance was family row when the victim was female, while a fight was the most common circumstance when the victim was male. Three offenders committed suicide after having committed homicide(s) (seven victims; three offenders). PMID- 10704817 TI - Chronological study of diazinon in putrefied viscera of rats using GC/MS, GC/EC and TLC. AB - A qualitative and qualitative weekly study of diazinon in experimental rats after a lethal dose is described. GC/MS and TLC were used for qualitative, and GC/EC for quantitative analysis. The replicate content of diazinon in stomach and intestine (S/I) revealed a high rate of decrease during the first month. The liver (L) content fluctuates through a general trend of decrease. Immediate post mortem content of 34.5 mg in summer and 94 in winter was found in S/L samples, while it was 0.79 and 0.63 respectively for L-samples. The respective remaining amount after 2 months was 1. 16, 4.40 (S/I), 0.61 and 0.66 mg (L). A round figure of 4% remains in S/L samples. The chronological plots indicate the possibility of detection at longer periods. Interpretation of data is outlined regarding relative contents of organs and the factors affecting persistence of diazinon in putrefied viscera. PMID- 10704818 TI - Fatal intoxication due to dothiepin. AB - An ingestion of an unknown quantity of Harmomed (dothiepin and diazepam) capsules in a suicide is described. The authors report a new and fast method of analysing and determining the dothiepin concentration in postmortem specimens. Quantitation of dothiepin, and its metabolite desmethyldothiepin was performed by ethyl acetate extraction from alkalinized body fluids before GC-MS analysis. The analyses were performed without any complex sample clean-up steps and with little sample material. Postmortem concentrations of dothiepin, desmethyldothiepin, diazepam and desmethyldiazepam in body fluids are given. The proposed method is a rapid procedure for analysis in cases of deliberate poisoning with the antidepressant drug dothiepin. PMID- 10704819 TI - RAFTK/Pyk2-mediated cellular signalling. AB - Intracellular signal transduction following extracellular ligation by a wide variety of surface molecules involves the activation and tyrosine phosphorylation of protein tyrosine kinases (PTKs). Tyrosine phosphorylation, controlled by the coordinated actions of protein tyrosine phosphatases (PTPs) and tyrosine kinases, is a critical regulatory mechanism for various physiological processes, including cell growth, differentiation, metabolism, cell cycle regulation and cytoskeleton function. The focal adhesion PTK family consists of the focal adhesion kinase (FAK) and the RAFTK/Pyk2 kinase (also known as CAK-beta and CADTK). RAFTK/Pyk2 can be activated by a variety of extracellular signals that elevate intracellular calcium concentration, and by stress signals. RAFTK/Pyk2 is expressed mainly in the central nervous system and in cells derived from hematopoietic lineages, while FAK is widely expressed in various tissues and links transmembrane integrin receptors to intracellular pathways. This review describes the role of RAFTK/Pyk2 in various signalling cascades and details the differential signalling by FAK and RAFTK/Pyk2. PMID- 10704820 TI - Structure and function of phosphatidylinositol-3,4 kinase. AB - Activation of phosphatidylinositol (PI)-kinase is involved in the regulation of a wide array of cellular activities. The enzyme exists as a dimer, consisting of a catalytic and a regulatory subunit. Five isoforms of the regulatory subunit have been identified and classified into three groups comprising respectively 85-kDa, 55-kDa, and 50-kDa proteins. Structural differences in the N-terminal regions of the different group members contribute to defining their binding specificity, their subcellular distributions, and their capacity to activate the 110-kDa catalytic subunit. Two widely distributed isoforms of the catalytic subunit have been identified-p110alpha and p110beta. Despite the fact that they bind to the p85alpha regulatory subunit similarly, p110alpha and p110beta appear to have separate functions within cells and to be activated by different stimuli. Moreover, although p85/p110 PI-kinase almost exclusively phosphorylates the D-3 position of the inositol ring in phosphoinositides when purified PI is used as a substrate in vitro, it appears to phosphorylate the D-4 position with similar or higher efficiency in vivo. Thus, it is highly probable that p85/p110 PI-kinase transmits signals to downstream targets via both D-3- and D-4-phosphorylated phosphoinositides. PMID- 10704822 TI - The GTP-binding protein G(ialpha) translocates to kinetochores and regulates the M-G(1) cell cycle transition of Swiss 3T3 cells. AB - The receptor-generated signals that are responsible for driving the cell cycle are incompletely characterised in mammalian cells. It is clear, however, that the cellular messenger systems that stimulate DNA synthesis and mitosis are separable. These are interwoven with biochemical checkpoints that ensure that processes, such as chromosomal replication and microtubule attachment to duplicated chromosomes, are complete before the following phase of the cell cycle is initiated. In some cells, activation of DNA synthesis by factors such as LPA and serum has been shown to require the GTP-binding protein G(i). We have found that G(i) plays an additional role in mitosis activated by both 7-transmembrane receptors and tyrosine kinase receptors, and that this involves the translocation of the alpha-subunit of G(i) (G(ialpha)) to the nucleus. Here we show by confocal microscopy that G(ialpha)migrates to the nucleus near the onset of mitosis in serum-activated Swiss 3T3 cells and binds to the kinetochore region of replicated chromosomes. Inhibition of G(i) function with pertussis toxin had no effect on the induction of DNA synthesis by serum, but cell proliferation was inhibited. Flow cytometric analysis showed that this resulted from retardation of the transition through mitosis and into G(1). Additionally, pertussis toxin impaired the activity of p34(cdc2), a cyclin-dependent kinase involved in the transition from M-phase to G(1), but not the S-phase cyclin, cyclin E. These data show that the G-protein G(i) has a key role in the regulation of mitosis in fibroblasts. PMID- 10704821 TI - Activation of stat3 and stat1 DNA binding and transcriptional activity in human brain tumour cell lines by gp130 cytokines. AB - In this study we examine the activation of the latent Stat family of transcription factors by the gp130 family of cytokines in cell lines derived from human brain tumours. Of the cytokines tested, oncostatin M resulted in the most dramatic induction of Stat1 and Stat3 in all cell lines analysed, as assessed by the formation of protein/DNA complexes. Interleukin-6, leukemia inhibitory factor, and ciliary neurotrophic factor also induced Stat complexes more selectively and to a lesser magnitude than oncostatin M. The kinetics of Stat1 and Stat3 activation was rapid and transient; the nuclear accumulation of DNA binding-proficient Stat protein was detected in the nucleus within minutes of cytokine induction. The transcriptional potential of the oncostatin M-activated Stat molecules was demonstrated in two glioma cell lines (U87-MG, SNB-19) by transient transfection experiments using a Stat-responsive reporter plasmid. Oncostatin M-dependent transcription from this reporter plasmid was reduced to uninduced levels by the inclusion of a dominant-negative Stat3 molecule, demonstrating that Stat molecules were responsible for the induction. These studies demonstrate that oncostatin M is the most potent activator of Stat molecules in a variety of brain tumour-derived cell lines, an observation that could have implications affecting the balance between proliferation/apoptosis of these cells. PMID- 10704823 TI - Tyrosine phosphorylation and association of FcgammaRII and p72(Syk) are not limited to the FcgammaRII signalling pathway. AB - The tyrosine kinase p72(Syk) plays a critical role in platelet signal transduction. It associates with the platelet receptor for the Fc domain of IgGs, FcgammaRII, following stimulation by FcgammaRII cross-linking. Here, we show that p72(Syk) and FcgammaRII tyrosine phosphorylation and association occured following platelet stimulation by: (1) two monoclonal antibodies, which form a bridge between a target antigen and FcgammaRII, and (2) the G-protein-coupled receptor agonist thrombin. The kinetics of the p72(Syk)/FcgammaRII association depended on the signalling pathway (i.e., the antigen targeted or the thrombin receptor). We established a direct relationship between the level of FcgammaRII phosphorylation and the detection of its association with p72(Syk). Inhibition of p72(Syk) by piceatannol resulted in partial or total inhibition of FcgammaRII phosphorylation, after immunological activation or addition of thrombin, respectively, suggesting that p72(Syk) participates in FcgammaRII phosphorylation. The results provide evidence that p72(Syk)/FcgammaRII association is not restricted to immunological activation. PMID- 10704824 TI - A role for phosphoinositides in tyrosine phosphorylation of p125 focal adhesion kinase in rat pancreatic acini. AB - Previous studies have shown that different agonists increase tyrosine phosphorylation of the focal adhesion related proteins p125(FAK), p130(Cas), and paxillin in different cell types and that tyrosine phosphorylation depends on the integrity of the actin cytoskeleton. Because phosphoinositides are important for the maintenance of the cytoskeleton, the role of phosphoinositides in the tyrosine phosphorylation of these proteins in response to occupancy of m3 muscarinic and CCK(A) receptors has been investigated in pancreatic acini. Addition of carbachol or CCK-8 to pancreatic acini resulted in rapid increases in the tyrosine phosphorylation of p125(FAK), p130(Cas), and paxillin. Pretreatment of pancreatic acini with LY294002 or wortmannin resulted in a concentration dependent inhibition of tyrosine phosphorylation of p125(FAK), p130(Cas), and paxillin stimulated by carbachol or CCK-8. Carbachol- or CCK-8-stimulated tyrosine phosphorylation of these proteins was not inhibited by rapamycin, PD 98059 or SB 203580, and thus it was dissociated from the activation of p70 S6 or MAP kinases. These results indicate that m3 muscarinic and CCK(A) receptor mediated increase in p125(FAK), p130(Cas), and paxillin tyrosine phosphorylation in pancreatic acini depends on the ability of these cells to synthesise phosphoinositides. PMID- 10704825 TI - Shc associates with the IL-3 receptor beta subunit, SHIP and Gab2 following IL-3 stimulation. Contribution of Shc PTB and SH2 domains. AB - p46(Shc) and p52(Shc) become heavily tyrosine phosphorylated in response to interleukin 3 (IL-3) treatment. We have investigated the potential of Shc to integrate IL-3 signalling pathways and demonstrate that Shc associates with the beta subunits of the human (betac) and murine (Aic2A) IL-3 receptors, SHIP and Gab2 following IL-3 stimulation. The interaction between Shc and the IL-3 receptor beta chains was direct, mediated by both the SH2 and PTB domains. Interaction with SHIP was via the Shc PTB domain and the Shc SH2 domain mediated the interaction with Gab2. Phosphopeptide competition studies suggest that the SH2 domain interacts primarily with tyrosine 612 of betac (610 of Aic2A), and the PTB domain with tyrosine 577 of betac (575 of Aic2A). PTB binding to IL-3R beta chains was of highest affinity, and appeared to play the primary role in binding. These findings suggest that Shc may play an important role in coordinately integrating IL-3 signalling pathways. PMID- 10704826 TI - The LIM domain: regulation by association. AB - The LIM domain is a zinc finger structure that is present in several types of proteins, including homeodomain transcription factors, kinases and proteins that consist of several LIM domains. Proteins containing LIM domains have been discovered to play important roles in a variety of fundamental biological processes including cytoskeleton organization, cell lineage specification and organ development, but also for pathological functions such as oncogenesis, leading to human disease. The LIM domain has been demonstrated to be a protein protein interaction motif that is critically involved in these processes. The recent isolation and analysis of more LIM domain-containing proteins from several species have confirmed and broadened our knowledge about LIM protein function. Furthermore, the identification and characterization of factors that interact with LIM domains illuminates mechanisms of combinatorial developmental regulation. PMID- 10704827 TI - Surface ectoderm is necessary for the morphogenesis of somites. AB - The paraxial mesoderm of the neck and trunk of mouse embryos undergoes extensive morphogenesis in forming somites. Paraxial mesoderm is divided into segments, it elongates along its anterior posterior axis, and its cells organize into epithelia. Experiments were performed to determine if these processes are autonomous to the mesoderm that gives rise to the somites. Presomitic mesoderm at the tailbud stage was cultured in the presence and absence of its adjacent tissues. Somite segmentation occurred in the absence of neural tube, notochord, gut and surface ectoderm, and occurred in posterior fragments in the absence of anterior presomitic mesoderm. Mesodermal expression of Dll1 and Notch1, genes with roles in segmentation, was largely independent of other tissues, consistent with autonomous segmentation. However, surface ectoderm was found to be necessary for elongation of the mesoderm along the anterior-posterior axis and for somite epithelialization. To determine if there is specificity in the interaction between ectoderm and mesoderm, ectoderm from different sources was recombined with presomitic mesoderm. Surface ectoderm from only certain parts of the embryo supported somite epithelialization and elongation. Somite epithelialization induced by ectoderm was correlated with expression of the basic-helix-loop-helix gene Paraxis in the mesoderm. This is consistent with the genetically defined requirement for Paraxis in somite epithelialization. However, trunk ectoderm was able to induce somite epithelialization in the absence of strong Paraxis expression. We conclude that somitogenesis consists of autonomous segmentation patterned by Notch signaling and nonautonomous induction of elongation and epithelialization by surface ectoderm. PMID- 10704828 TI - Function of the spalt/spalt-related gene complex in positioning the veins in the Drosophila wing. AB - Spalt and Spalt-related encode conserved Zn-finger proteins that mediate the function of the TGF-beta molecule Decapentaplegic during the positioning of veins in the Drosophila wing. Here we show that Spalt and Spalt-related regulate the vein-specific expression of the transcription factors of the knirps and iroquois gene complexes, delimiting their domains of expression in the wing pouch. The effects of spalt/spalt-related mutations on knirps and iroquois expression are cell-autonomous, suggesting that they could be direct. The regulation of iroquois involves transcriptional repression by Spalt and Spalt-related, whereas the regulation of knirps involves a combination of transcriptional activation and repression mediated by the same genes. We suggest that the regulation of the iroquois and knirps gene complexes by Spalt and Spalt-related translates the Decapentaplegic morphogenetic gradient into precisely spaced pattern elements. PMID- 10704829 TI - Interaction between Otx2 and Gbx2 defines the organizing center for the optic tectum. AB - Otx2 is expressed in the mesencephalon and prosencephalon, and Gbx2 is expressed in the rhombencephalon around stage 10. Loss-of-function studies of these genes in mice have revealed that Otx2 is indispensable for the development of the anterior brain segment, and that Gbx2 is required for the development of the isthmus. We carried out gain-of-function experiments of these genes in chick embryos with a newly developed gene transfer system, in ovo electroporation. When Otx2 was ectopically expressed caudally beyond the midbrain-hindbrain boundary (MHB), the alar plate of the metencephalon differentiated into the optic tectum instead of differentiating into the cerebellum. On the other hand, when Gbx2 was ectopically expressed at the mesencephalon, the caudal limit of the tectum shifted rostrally. We looked at the effects of misexpression on the isthmus- and tectum-related molecules. Otx2 and Gbx2 interacted to repress each other's expression. Ectopic Otx2 and Gbx2 repressed endogenous expression of Fgf8 in the isthmus, but induced Fgf8 expression at the interface between Otx2 and Gbx2 expression. Thus, it is suggested that interaction between Otx2 and Gbx2 determines the site of Fgf8 expression and the posterior limit of the tectum. PMID- 10704830 TI - Ectopic pigmentation in Xenopus in response to DCoH/PCD, the cofactor of HNF1 transcription factor/pterin-4alpha-carbinolamine dehydratase. AB - DCoH, the dimerization cofactor of the HNF-1 homeodomain proteins (hepatocyte nuclear factor-1alpha and beta), is involved in gene expression by associating with these transcription factors. The protein also called PCD for pterin-4alpha carbinolamine dehydratase is a bifunctional factor as it catalyzes also the regeneration of tetrahydrobiopterin. This coenzyme is used by the enzyme phenylalanine hydroxylase, which generates tyrosine, the precursor of catecholamines and melanin. DCoH/PCD presumably cooperates with other partners, because it is expressed earlier than HNF1 and phenylalanine hydroxylase (PAH) in early vertebrate development. It is also found in cells lacking HNF1 and PAH like skin, brain and the pigmented epithelium of the eye suggesting a yet unknown function. We show that the overexpression of DCoH/PCD in Xenopus induces the formation of ectopic pigment cells in the epidermis, that are visible earlier than the endogenous pigmentation and broader distributed. This ectopic pigmentation is accompanied by an increase in tyrosinase activity and the amount of melanin. Overexpression of DCoH/PCD induces the appearance of pigment cells also in animal cap explants, that normally differentiate into atypical epidermis. DCoH/PCD mutants with impaired carbinolamine dehydratase activity retain the potential to induce pigmentation and we propose therefore that DCoH/PCD is not simply an essential enzyme for melanin biosynthesis, but also a regulator for the differentiation of pigment producing cells. PMID- 10704831 TI - The onset of germ cell migration in the mouse embryo. AB - Mouse primordial germ cells (PGCs) are specified between embryonic day 6.5 (E6.5) and E7.5, when they have been visualized as an alkaline phosphatase-positive (AP+) cell population in the developing allantois. By E8.5, they are embedded in the hind-gut epithelium. Previous experiments have suggested different sites for PGCs' origin, and it is unclear how they reach the gut epithelium. We have used transgenic mice expressing GFP under a truncated Oct4 promoter to visualize living PGCs. We find GFP+/AP+ cells in the posterior end of the primitive streak as a dispersed population of cells actively migrating into the allantois, and directly into the adjacent embryonic endoderm. Time-lapse analysis shows these cells to be actively migratory from the time they exit the primitive streak. PMID- 10704832 TI - The control of Xenopus embryonic primary neurogenesis is mediated by retinoid signalling in the neurectoderm. AB - In Xenopus, the primary neurons form in three domains either side of the midline in the posterior neurectoderm. At the late neurula stage there are approximately 120 primary sensory neurons on each side of the embryo. Co-injecting synthetic mRNA encoding retinoic acid receptor alpha (NR1B1) and retinoid X receptor beta (NR2B2) results in an increase in the number of primary neurons and this is further enhanced by the addition of retinoic acid indicating that elevated retinoid signalling promotes an increase in the number of cells undergoing primary neurogenesis. However, primary neurogenesis remains confined to the three domains that normally give rise to primary neurons indicating that not all regions of the neurectoderm respond equivalently to elevated retinoid signalling. The inhibition of retinoid signalling with a dominant negative retinoid receptor or treatment with citral, an inhibitor of retinoid metabolism, inhibits the formation of primary neurons. However, the lateral extent of the neurectoderm does not differ following these experimental manipulations suggesting that changes in primary neuron cell number, in response to changes in retinoid signalling, cannot be accounted for by significant gains or losses of neurectoderm. In addition, two lines of evidence are presented to suggest that retinoid signalling affects primary neurogenesis by acting directly on the neurectoderm. First, animal caps neuralized by noggin undergo primary neurogenesis in response to retinoid signalling and second primary neurogenesis is elevated in neural conjugates in which the ectodermal, but not the mesodermal, component has been co-injected with RAR/RXR mRNA. PMID- 10704833 TI - Xenopus Xenf: an early endodermal nuclear factor that is regulated in a pathway distinct from Sox17 and Mix-related gene pathways. AB - We report a novel zygotic gene encoding a Xenopus endodermal nuclear factor, Xenf. Expression of Xenf starts at the late blastula stages and is decreased after gastrulation. Xenf shows no structural homology to any known proteins. When GFP-tagged Xenf is overexpressed in Xenopus cells, Xenf protein is localized to the nucleus, associating closely with the chromosomes. In animal cap assays, Xenf expression is strongly activated by mRNA injection of Vg1 and VegT, maternal vegetal genes that can induce endodermal differentiation. In contrast, Xenf is not induced by endoderm-inducing zygotic transcription factors such as Sox17 and Mix-related genes. In turn, Xenf does not activate expression of Sox17, Mixer or Milk. Thus, Xenf is regulated by maternal vegetal positional information in a parallel manner to Sox17 and Mix-related gene pathways. PMID- 10704834 TI - The early expression of VAChT and VIP in mouse sympathetic ganglia is not induced by cytokines acting through LIFRbeta or CNTFRalpha. AB - Sympathetic ganglia consist of noradrenergic and cholinergic neurons. The cholinergic marker protein vesicular acetylcholine transporter (VAChT) and the neuropeptide vasoactive intestinal peptide (VIP), co-expressed in mature cholinergic sympathetic neurons, are first detectable during embryonic development of rat sympathetic ganglia. However, the subpopulation of cholinergic sympathetic neurons which innervates sweat glands in mammalian footpads starts to express VAChT and VIP during the first postnatal weeks, under the influence of sweat gland-derived signals. In vitro evidence suggests that the sweat gland derived cholinergic differentiation factor belongs to a group of neuropoietic cytokines, including LIF, CNTF and CT-1, that act through a LIFRbeta-containing cytokine receptor. To investigate whether the embryonic expression of cholinergic properties is elicited by a related cytokine, the expression of VAChT and VIP was analyzed in stellate ganglia of mice deficient for the cytokine receptor subunits LIFRbeta or CNTFRalpha. The density of VAChT- and VIP-immunoreactive cells in stellate ganglia of new-born animals was not different in LIFRbeta(-/-) and CNTFRalpha(-/-) ganglia as compared to ganglia from wild-type mice. These results demonstrate that the early, embryonic expression of VAChT and VIP is not induced by cytokines acting through LIFRbeta- or CNTFRalpha-containing receptors. PMID- 10704835 TI - Overlapping and specific functions of Braf and Craf-1 proto-oncogenes during mouse embryogenesis. AB - The three mammalian Raf serine/threonine protein kinases mediate the transduction of proliferative and differentiative signals from cell surface receptors to the nucleus. In vertebrates, Raf signaling has been implicated in the progression of mouse embryos through the two-cell stage and in the induction of posterior mesoderm. However, mouse embryos mutant for each of the Raf genes exhibit no developmental defects before mid-gestation. Here we describe the phenotype of mouse mutants with different combinations of mutant Craf-1 and Braf alleles. Our results show that Raf signaling is indeed indispensable for normal development beyond the blastocyst stage. However, due to a significant redundancy between Craf-1 and Braf, either gene is sufficient for normal development until mid gestation. The molecular and developmental mechanisms for this redundancy were investigated by monitoring the expression of Raf genes throughout embryogenesis and by biochemical studies in mutant cell lines. PMID- 10704836 TI - Regulatory gene expression patterns reveal transverse and longitudinal subdivisions of the embryonic zebrafish forebrain. AB - To shed light on the organization of the rostral embryonic brain of a lower vertebrate, we have directly compared the expression patterns of dlx, fgf, hh, hlx, otx, pax, POU, winged helix and wnt gene family members in the fore- and midbrain of the zebrafish. We show that the analyzed genes are expressed in distinct transverse and longitudinal domains and share expression boundaries at stereotypic positions within the fore- and midbrain. Some of these shared expression boundaries coincide with morphological landmarks like the pathways of primary axon tracts. We identified a series of eight transverse diencephalic domains suggestive of neuromeric subdivisions within the rostral brain. In addition, we identified four molecularly distinct longitudinal subdivisions and provide evidence for a strong bending of the longitudinal rostral brain axis at the cephalic flexure. Our data suggest a strong conservation of early forebrain organization between lower and higher vertebrates. PMID- 10704837 TI - XTIF2, a Xenopus homologue of the human transcription intermediary factor, is required for a nuclear receptor pathway that also interacts with CBP to suppress Brachyury and XMyoD. AB - Ligand-bound nuclear receptors (NRs) recruit cofactors such as members of the p160 family and CREB-binding protein (CBP) to activate transcription. We have cloned the Xenopus homologue of the human transcription intermediary factor 2 (TIF2), a member of the p160 family of cofactors. Xenopus TIF2 (XTIF2) mRNA is expressed homogeneously during late blastula-early gastrula stages and later becomes highly expressed in the notochord. To study the function of XTIF2 during development, we have used two dominant negative constructs, one encompassing the NR-binding domain and the other the CBP interacting region of XTIF2. Overexpression of the XTIF2 dominant negative mRNAs causes ectopic expression of Xenopus Brachyury (Xbra) and MyoD in all tissue layers. Moreover, ectopic expression of the dominant negative construct that contains the CBP-binding region produces strong phenotypes at hatching stage such as loss of head structures, shortened trunks and open blastopores, which can be rescued by XTIF2 coexpression. These observed defects are due, at least in part, to repression of dorsal mesoderm and endoderm genes. Our data suggest the existence of a NR pathway that requires XTIF2 and CBP to repress Xbra and XMyoD. PMID- 10704838 TI - Requirement of Xmsx-1 in the BMP-triggered ventralization of Xenopus embryos. AB - Signaling triggered by polypeptide growth factors leads to the activation of their target genes. Several homeobox genes are known to be induced in response to polypeptide growth factors in early Xenopus development. In particular, Xmsx-1, an amphibian homologue of vertebrate Msx-1, is well characterized as a target gene of bone morphogenetic protein (BMP). Here, using a dominant-negative form of Xmsx-1 (VP-Xmsx-1), which is a fusion protein made with the virus-derived VP16 activation domain, we have examined whether Xmsx-1 activity is required in the endogenous ventralizing pathway. VP-Xmsx-1 induced a secondary body axis, complete with muscle and neural tissues, when overexpressed in ventral blastomeres, suggesting that Xmsx-1 activity is necessary for both mesoderm and ectoderm to be ventralized. We have also examined the epistatic relationship between Xmsx-1 and another ventralizing homeobox protein, Xvent-1, and show that Xmsx-1 is likely to be acting upstream of Xvent-1. We propose that Xmsx-1 is required in the BMP-stimulated ventralization pathway that involves the downstream activation of Xvent-1. PMID- 10704839 TI - The role of cadherins during primordial germ cell migration and early gonad formation in the mouse. AB - Primordial germ cells (PGCs) are the founder cells of the gametes. In mammals, PGCs migrate from the hindgut to the genital ridges, where they coalesce with each other and with somatic cells to form the primary sex cords. We show here that, in both sexes, PGCs express P- and E-cadherins during and after migration, and N-cadherin at post-migratory stages. E-Cadherin is not expressed by PGCs whilst in the hindgut, but is upregulated as they leave. Blocking antibodies against E-, but not P-cadherin cause defective PGC-PGC coalescence, and in some cases, ectopic PGCs. PMID- 10704840 TI - Severe developmental defects in Dictyostelium null mutants for actin-binding proteins. AB - The actin cytoskeleton is implicated in many cellular processes, such as cell adhesion, locomotion, contraction and cytokinesis, which are central to any development. The extent of polymerization, cross-linking, and bundling of actin is regulated by several actin-binding proteins. Knock-out mutations in these proteins have revealed in many cases only subtle, if any, defects in development, suggesting that the actin system is redundant, with multiple proteins sharing overlapping functions. The apparent redundancy may, however, reflect limitations of available laboratory assays in assessing the developmental role of a given protein. By using a novel assay, which reproduces conditions closer to the natural ones, we have re-examined the effects of disruption of many actin-binding proteins, and show here that deletion of alpha-actinin, interaptin, synexin, 34 kDa actin-bundling protein, and gelation factor affect to varying degrees the efficiency of Dictyostelium cells to complete development and form viable spores. No phenotypic defects were found in hisactophilin or comitin null mutants. PMID- 10704841 TI - Interaction between the bHLH-PAS protein Trachealess and the POU-domain protein Drifter, specifies tracheal cell fates. AB - bHLH-PAS proteins represent a class of transcription factors involved in diverse biological activities. Previous experiments demonstrated that the PAS domain confers target specificity (Zelzer et al., 1997. Genes Dev. 11, 2079-2089). This suggested an association between the PAS domain and additional DNA-binding proteins, which is essential for the induction of specific target genes. A candidate for interaction with Trh is Drifter/Ventral veinless, a POU-domain protein. A dual requirement for Trh and Drifter was identified for the autoregulation of Trh and Drifter expression. Furthermore, ectopic expression of both Trh and Dfr (but not each one alone) triggered trh autoregulation in several embryonic tissues. A direct interaction between Drifter and Trh proteins, mediated by the PAS domain of Trh and the POU domain of Drifter, was demonstrated. PMID- 10704842 TI - Ectopic expression of Msx2 in chick retinal pigmented epithelium cultures suggests a role in patterning the optic vesicle. AB - During the initial stages of vertebrate retinogenesis, cells of the optic vesicle adopt one of two alternate cell fates. Cells in the distal-most part of the vesicle, immediately beneath the surface ectoderm, undergo neural differentiation; cells in the proximal part differentiate into retinal pigmented epithelial cells. The mechanisms that establish this pattern of differentiation are poorly understood. In the mouse embryo, Msx2, a homeobox-containing transcription factor, is expressed in cells of the optic vesicle that will form the neural retina, whilst the developing retinal pigmented epithelium (RPE) does not express this gene. Msx2 could therefore be involved in patterning the optic vesicle into neural and pigmented domains. To explore this possibility we ectopically expressed mouse Msx2 in cultures of chick RPE cells. Compared with cultures transfected with a control construct, Msx2-transfected cultures contained fewer cells expressing the RPE marker, Mitf, and more cells expressing class III beta-tubulin, a neuronal marker. In addition a small proportion of Msx2 transfected cells acquired a neural-like morphology. These results show that Msx2 can suppress the differentiated state of RPE cells and promote their differentiation into neural cell types. We suggest that Msx2 may pattern the optic vesicle into neural and pigmented domains by affecting the balance between RPE and neural retina differentiation. PMID- 10704843 TI - The Drosophila gene abstrakt, required for visual system development, encodes a putative RNA helicase of the DEAD box protein family. AB - The molecular mechanisms underlying axonal pathfinding are not well understood. In a genetic screen for mutations affecting the projection of the larval optic nerve we isolated the abstrakt locus. abstrakt is required for pathfinding of the larval optic nerve, and it also affects development in both the adult visual system and the embryonic CNS. Here we report the molecular characterization of abstrakt. It encodes a putative ATP-dependent RNA helicase of the DEAD box protein family, with two rare substitutions in the PTRELA and the RG-D motifs, thought to be involved in oligonucleotide binding: serine for threonine, and lysine for arginine, respectively. Two mutant alleles of abstrakt show amino acid exchanges in highly conserved positions. A glycine to serine exchange in the HRIGR motif, which is involved in RNA binding and ATP hydrolysis, results in a complete loss of protein function; and a proline to leucine exchange located between the highly conserved ATPase A and PTRELA motifs results in temperature sensitive protein function. Both the broad requirement for abstrakt gene function and its ubiquitous expression are consistent with a molecular function of the abstrakt protein in mRNA splicing or translational control. PMID- 10704844 TI - gcm and pointed synergistically control glial transcription of the Drosophila gene loco. AB - In Drosophila lateral glial cell development is initiated by the transcription factor encoded by glial cells missing. glial cells missing activates downstream transcription factors such as repo and pointed which subsequently control terminal glial differentiation. The gene loco has been identified as a potential target gene of pointed and is involved in terminal glial differentiation. It encodes an RGS domain protein expressed specifically by the lateral glial cells in the developing embryonic CNS. Here we analyzed the loco promoter and the control of the glial-specific transcription pattern. Using promoter-reporter gene fusions we identified a 1.9 kb promoter element capable of directing the almost complete loco gene expression pattern. Sequence analysis suggested the presence of gcm and pointed DNA binding sites. Following in vitro mutagenesis of these sites we demonstrated their relevance in vivo. The expression of loco is initially dependent on gcm. During subsequent stages of embryonic development gcm and pointed appear to activate loco transcription synergistically. In addition, at least two other factors appear to repress loco expression in the ectoderm and in the CNS midline cells. PMID- 10704845 TI - knot is required for the hypopharyngeal lobe and its derivatives in the Drosophila embryo. AB - The genetic control of segmentation in the Drosophila larval head has been only partially elucidated. We report that the knot (kn) gene of Drosophila is required for the formation of the hypopharyngeal lobe in the germ-band stage embryo and the ventral arms and lateralgrate of the larval head skeleton. Rearrangement mutations of knot disrupt the previously characterized gene, collier, which encodes a COE-family transcription factor. kn is required for the activation of cnc (cap'n'collar, a bZIP homeotic selector gene) in the progenitors of hypopharyngeal lobe. kn is also required for the activation of an EST marker for the cephalic mesoderm, CK01299. Based on the relative expression of kn and the posterior compartment-specific gene hedgehog, we provide additional evidence that in Drosophila, unlike many other insects, the hypopharyngeal lobe arises from part of the mandibular segment. PMID- 10704846 TI - The design and analysis of a homeotic response element. AB - We have shown that the 26 bp bx1 element from the regulatory region of Distal less is capable of imposing control by the homeotic genes Ultrabithorax and abdominal-A on a general epidermal activator in Drosophila. This provides us with an assay to analyze the sequence requirements for specific repression by these Hox genes. Both the core Hox binding site, 5'-TAAT, and the adjacent EXD 5'-TGAT core site are required for repression by Ultrabithorax and abdominal-A. The Distal-less bx1 site thus fits with the model of Hox protein binding specificity based on the consensus PBX/HOX-family site TGATNNAT[g/t][g/a], where the key elements of binding specificity are proposed to lie in the two base pairs following the TGAT. A single base pair deletion in the bx1 sequence generates a site, bx1:A(-)mut, that on the consensus PBX/HOX model would be expected to be regulated by the Deformed Hox gene. We observed, however, that the bx1:A(-)mut site was regulated predominantly by Sex combs reduced, Ultrabithorax and abdominal-A. The analysis of this site indicates that the specificity of action of Hox proteins may depend not only on selective DNA binding but also on specific post-binding interactions. PMID- 10704847 TI - Regulation of early expression of Dlx3, a Xenopus anti-neural factor, by beta catenin signaling. AB - The ectoderm of the pre-gastrula Xenopus embryo has previously been shown to be at least partially patterned along the dorsal-ventral axis. The early expression of the anti-neural homeodomain gene Dlx3 is localized to the ventral ectoderm by a mechanism that occurs prior to gastrulation and is independent of the Spemann organizer. The repression of Dlx3 is mediated by signaling though beta-catenin, but is probably not dependent on the induction of the Xnr3 or chordin genes by beta-catenin. We propose a model in which this early regulation of Dlx3 accounts for the pro-neural bias of dorsal ectoderm. PMID- 10704848 TI - Characterization of the regulatory elements controlling neuronal expression of the A-isoform of the ecdysone receptor gene of Drosophila melanogaster. AB - During the development of the adult central nervous system (CNS) of the fruitfly Drosophila melanogaster, the A-isoform of the ecdysone receptor (EcR-A), a typical nuclear hormone receptor, is expressed at high levels in the Type II neurons, a set of neurons that die shortly after the emergence of the adult. To understand the role that transcriptional regulation of nuclear receptor genes plays in CNS development, we have dissected the region controlling the transcription of EcR-A by analyzing the ability of this region to drive the expression of reporter genes in transgenic animals. These analyses have demonstrated that the Type II neurons are a heterogeneous collection of neurons that utilize different regulatory elements to coordinate the expression of the same transcript. PMID- 10704849 TI - Brachyury is a target gene of the Wnt/beta-catenin signaling pathway. AB - To identify target genes of the Wnt/beta-catenin signaling pathway in early mouse embryonic development we have established a co-culture system consisting of NIH3T3 fibroblasts expressing different Wnts as feeder layer cells and embryonic stem (ES) cells expressing a green fluorescent protein (GFP) reporter gene transcriptionally regulated by the TCF/beta-catenin complex. ES cells specifically respond to Wnt signal as monitored by GFP expression. In GFP positive ES cells we observe expression of Brachyury. Two TCF binding sites located in a 500 bp Brachyury promoter fragment bind the LEF-1/beta-catenin complex and respond specifically to beta-catenin-dependent transactivation. From these results we conclude that Brachyury is a target gene for Wnt/beta-catenin signaling. PMID- 10704850 TI - BMP2 is required for early heart development during a distinct time period. AB - BMP2, like its Drosophila homologue dpp, is an important signaling molecule for specification of cardiogenic mesoderm in vertebrates. Here, we analyzed the time course of BMP2-requirement for early heart formation in whole chick embryos and in explants of antero-lateral plate mesoderm. Addition of Noggin to explants isolated at stage 4 and cultured for 24 h resulted in loss of NKX2.5, GATA4, eHAND, Mef2A and vMHC expression. At stages 5-8 the individual genes showed differential sensitivity to Noggin addition. While expression of eHAND, NKX2.5 and Mef2A was clearly reduced by Noggin vMHC was only marginally affected. In contrast, GATA4 expression was enhanced after Noggin treatment. The developmental period during which cardiac mesoderm required the presence of BMP signaling in vivo was assessed by implantation of Noggin expressing cells into stage 4-8 embryos which were then cultured until stage 10-11. Complete loss of NKX2.5 and eHAND expression was observed in embryos implanted at stages 4-6, and expression was still suppressed in stages 7 and 8 implanted embryos. GATA4 expression was also blocked by Noggin at stage 4, however increased at stages 5, 6 and 7. Explants of central mesendoderm, that normally do not form heart tissue were employed to study the time-course of BMP2-induced cardiac gene expression. The induction of cardiac lineage markers in central mesendoderm of stage 5 embryos was distinct for different genes. While GATA4, -5, -6 and MEF2A were induced to maximal levels within 6 h after BMP2 addition, eHAND and dHAND required 12 h to reach maximum levels of expression. NKX2.5 was induced by 6 h and accumulated over 48 h. vMHC and titin were induced at significant levels only after 48 h of BMP2 addition. These results indicate that cardiac marker genes display distinct expression kinetics after BMP2 addition and differential response to Noggin treatment suggesting complex regulation of myocardial gene expression in the early tubular heart. PMID- 10704851 TI - Cross-regulatory interactions among tracheal genes support a co-operative model for the induction of tracheal fates in the Drosophila embryo. AB - The Drosophila tracheal system arises from clusters of ectodermal cells that invaginate and migrate to originate a network of epithelial tubes. Genetic analyses have identified several genes that are specifically expressed in the tracheal cells and are required for tracheal development. Among them, trachealess (trh) is able to induce ectopic tracheal pits and therefore it has been suggested that it would act as an inducer of tracheal cell fates; however, this capacity appears to be spatially restricted. Here we analyze the expression of the tracheal specific genes in the early steps of tracheal development and their cross-interactions. We find that there is a set of primary genes including trh and ventral veinless (vvl) whose expression does not depend on any other tracheal gene and a set of downstream genes whose expression requires different combinations of the primary genes. We also find that the combined expression of primary genes is sufficient to induce some downstream genes but not others. These results indicate that there is not a single master gene responsible for the appropriate expression of the tracheal genes and support a model where tracheal cell fates are induced by the co-operation of several factors rather than by the activity of a single tracheal inducer. PMID- 10704852 TI - Direct interaction of two homeoproteins, homothorax and extradenticle, is essential for EXD nuclear localization and function. AB - The Drosophila Homothorax (HTH) and Extradenticle (EXD) are two homeoproteins required in a number of developmental processes. EXD can function as a cofactor to Hox proteins. Its nuclear localization is dependent on HTH. In this study we present evidence of in vivo physical interaction between HTH and EXD, mediated primarily through an evolutionarily conserved MH domain in HTH. This interaction is essential for the mutual stabilization of both proteins, for EXD nuclear localization, and for the cooperative DNA binding of the EXD-HTH heterodimer. Some in vivo functions require both EXD and HTH in the nucleus, suggesting that the EXD-HTH complex may function as a transcriptional regulator. PMID- 10704853 TI - Cooperative roles of Bozozok/Dharma and Nodal-related proteins in the formation of the dorsal organizer in zebrafish. AB - In vertebrates, specification of the dorso-ventral axis requires Wnt signaling, which leads to formation of the Nieuwkoop center and the Spemann organizer (dorsal organizer), through the nuclear accumulation of beta-catenin. Zebrafish bozozok/dharma (boz) and squint (sqt), which encode a homeodomain protein and a Nodal-related protein, respectively, are required for the formation of the dorsal organizer. The zygotic expression of boz and sqt in the dorsal blastoderm and dorsal yolk syncytial layer (YSL) was dependent on the maternally derived Wnt signal, and their expression at the late blastula and early gastrula stages was dependent on the zygotic expression of their own genes. The dorsal organizer genes, goosecoid (gsc) and chordin (din), were ectopically expressed in wild-type embryos injected with boz or sqt RNA. The expression of gsc strictly depended on both boz and sqt while the expression of din strongly depended on boz but only partially depended on sqt and cyclops (cyc, another nodal-related gene). Overexpression of boz in embryos defective in Nodal signaling elicited the ectopic expression of din but not gsc and resulted in dorsalization, implying that boz could induce part of the organizer, independent of the Nodal proteins. Furthermore, boz; sqt and boz;cyc double mutants displayed a severely ventralized phenotype with anterior truncation, compared with the single mutants, and boz;sqt;cyc triple mutant embryos exhibited an even more severe phenotype, lacking the anterior neuroectoderm and notochord, suggesting that Boz/Dharma and the Nodal-related proteins cooperatively regulate the formation of the dorsal organizer. PMID- 10704854 TI - Mammary gland patterning in the AXB/BXA recombinant inbred strains of mouse. PMID- 10704855 TI - Differential expression of the Groucho-related genes 4 and 5 during early development of Xenopus laevis. AB - Recently, we demonstrated that the Xenopus Wnt effector XTcf-3 interacts with Groucho-related transcriptional repressors (Roose et al., 1998. Nature 395, 608 612). A long form of the Groucho-related genes, XGrg-4, was shown to repress axis formation in the Xenopus embryo, whereas a short form, XGrg-5, acted as a potentiator. In this study, the temporal and spatial expression of XGrg-4 and XGrg-5 is described in Xenopus laevis embryos. Both genes are maternally expressed. In the gastrula, transcripts of both genes are present in the animal as well as the vegetal region. At later stages, XGrg-4 and XGrg-5 show specific patterns of expression in the central nervous system (CNS), cranial ganglia, eyes, otic vesicles, stomodeal-hypophyseal anlage, cement gland, head mesenchyme, branchial arches, neural crest and derivatives, somites, pronephros, pronephric duct, heart and tailbud. Differences in the expression of XGrg-4 and XGrg-5 were found in the CNS, cranial ganglia, olfactory placodes, stomodeal-pharyngeal anlage, cement gland, head mesenchyme and ectoderm. PMID- 10704856 TI - Expression of two novel mouse Iroquois homeobox genes during neurogenesis. AB - Members of the Drosophila Iroquois homeobox gene family are implicated in the development of peripheral nervous system and the regionalization of wing and eye imaginal discs. Recent studies suggest that Xenopus Iroquois homeobox (Irx) genes are also involved in neurogenesis. Three mouse Irx genes, Irx1, Irx2 and Irx3, have been previously identified and are expressed with distinct spatio-temporal patterns during neurogenesis. We report here the cloning and expression analysis of two novel mouse Irx genes, Irx5 and Irx6. Although Irx5 and Irx6 proteins are structurally more related to one another, we find that Irx5 displays a developmental expression pattern strikingly similar to that of Irx3, whereas Irx6 expression resembles that of Irx1. Consistent with the notion that Mash1 is a putative target gene of the Irx proteins, all four Irx genes display an overlapping expression pattern with Mash1 in the developing CNS. In contrast, the Irx genes and Mash1 are expressed in complementary domains in the developing eye and olfactory epithelium. PMID- 10704857 TI - Male specific expression suggests role of DMRT1 in human sex determination. AB - Sex determination in mammals is controlled by various transcription factors. Following the identification of SRY on the Y chromosome, several other factors have been identified. They can normally be identified as being involved in sex determination by the identification of sex reversal mutations or deletions, functional studies, and also by male-specific expression patterns in embryos. Here, it is shown that DMRT1, recently demonstrated to be deleted in 9p monosomies associated with sex reversal, is specifically expressed during sex determination in the genital ridge of human male, but not female, embryos, similar to SRY. PMID- 10704858 TI - Cloning and expression of xSix3, the Xenopus homologue of murine Six3. AB - The vertebrate Six family of transcription factor genes are homologues of the fruitfly gene sine oculis (so). Human, murine, avian and fish (medaka, zebrafish) homologues have recently been cloned. We report the cloning and developmental pattern of expression of xSix3, the Xenopus laevis homologue of Six3. In addition, we have compared all the known sequences of vertebrate Six3 genes. xSix3 is very homologous to Six3 in other vertebrates in terms of amino acid sequence. The reported developmental pattern of expression of Six3 in chick and mouse includes not only the developing eyes and the ventral diencephalic tissue between them, but also a large, sagittally-oriented telencephalic region. The distribution of xSix3, however, is virtually restricted to the eyes and ventral diencephalon, showing only a very small territory of expression in the telencephalon. PMID- 10704859 TI - The Ets family member Erg gene is expressed in mesodermal tissues and neural crests at fundamental steps during mouse embryogenesis. AB - The Erg gene belongs to the Ets family encoding a class of transcription factors. To gain new insight on the in vivo functional specificity of the Erg gene within the wide Ets family, we used in situ hybridization to determine its expression pattern during murine embryogenesis. We found that the Erg gene expression predominates in mesodermal tissues, including the endothelial, precartilaginous and urogenital areas. A specific Erg gene expression was also identified in migrating neural crest cells. A comparison with Fli-1, the most closely Erg related gene, revealed that both gene expressions partially overlap, suggesting that they may contribute to related functions in these tissues. Like other Ets family genes, Erg seems involved in several fundamental developmental steps in murine embryogenesis, including epithelio-mesenchymal transition, cell migration, settlement and differentiation. PMID- 10704860 TI - Developmental expression of the hemichordate otx ortholog. AB - The phylogenetic location of hemichordates is unique because they seem to fill an evolutionary gap between echinoderms and chordates. We report here characterization of Pf-otx, a hemichordate ortholog of otx, with its embryonic and larval expression pattern. Pf-otx is initially expressed in the vegetal plate of the blastula. Expression remains evident in the archenteron through gastrulation and then disappears. A new expression domain appears near the mouth along the preoral and postoral ciliated bands in the early tornaria larva. PMID- 10704861 TI - Expression patterns of Twist and Fgfr1, -2 and -3 in the developing mouse coronal suture suggest a key role for twist in suture initiation and biogenesis. AB - Sutural growth depends on maintenance of a balance between proliferation of osteogenic stem cells and their differentiation to form new bone, so that the stem cell population is maintained until growth of the skull is complete. The identification of heterozygous mutations in FGFR1, -2 and -3 and TWIST as well as microdeletions of TWIST in human craniosynostosis syndromes has highlighted these genes as playing important roles in maintaining the suture as a growth centre. In contrast to Drosophila, a molecular relationship between human (or other vertebrate) TWIST and FGFR genes has not yet been established. TWIST mutations exert their effect via haploinsufficiency whereas FGFR mutations have a gain-of function mechanism of action. To investigate the biological basis of FGFR signalling pathways in the developing calvarium we compared the expression patterns of Twist with those of Fgfr1, -2 and -3 in the fetal mouse coronal suture over the course of embryonic days 14-18, as the suture is initiated and matures. Our results show that: (1) Twist expression precedes that of Fgfr genes at the time of initiation of the coronal suture; (2) in contrast to Fgfr transcripts, which are localised within and around the developing bone domains, Twist is expressed by the midsutural mesenchyme cells. Twist expression domains show some overlap with those of Fgfr2, which is expressed in the most immature (proliferating) osteogenic tissue. PMID- 10704862 TI - Identification and developmental expression of par-6 gene in Xenopus laevis. AB - The par genes (partitioning defective) are required to establish polarity in the Caenorhabditis elegans embryo. We have identified the Xenopus homologue of C. elegans PAR-6 (XPAR-6). XPAR-6 is a protein of 377 amino acids with one PDZ domain which is involved in mediating protein-protein interactions. It shares 59% and 58% amino acid identity with the mouse and Drosophila PAR-6, respectively, and 54% overall identity with C. elegans PAR-6. Xpar-6 is expressed both maternally and zygotically. Xpar-6 is first detected in the animal half of the egg, and this pattern of expression persists into the cleavage and blastula stages. At the gastrula stage, the message is detected in animal pole area and in a broad domain of ventral region, but is excluded from dorsal region. With the onset of neurulation, the localized expression of Xpar-6 becomes more obvious, leading to it being enriched in the dorsolateral region along the lateral edges of neural plate and anterior presumptive head region surrounding the anterior border of neural plate. At late tailbud stage, Xpar-6 transcripts show localized expression throughout the head, labeling the branchial arches, eyes, otic vesicles and brain, while more posteriorly Xpar-6 labels the somites, pronephros, tail tip and proctodeum. Therefore, this analysis suggests that Xpar-6 has a regionalized pattern of expression during Xenopus early embryogenesis. PMID- 10704863 TI - Dynamic expression of lunatic fringe during feather morphogenesis: a switch from medial-lateral to anterior-posterior asymmetry. AB - Expression of Lunatic fringe mRNA was studied during feather morphogenesis and showed three stages of dynamic expression pattern. (1) Lunatic fringe was first expressed in the epithelium as a ring bordering the feather primordium when it was initially induced. (2) Shortly after, it showed a polarized pattern, first toward the lateral side of the feather primordium and then made a 90 degrees C switch toward the posterior side of the short bud. It then becomes weakly expressed in the long bud stage. (3) Finally, it is expressed in the marginal plate epithelia of feather filaments. In contrast, Radical fringe is weakly expressed in the feather bud, but is also present in the marginal plate epithelia of feather filaments. PMID- 10704864 TI - Isoform-specific expression of BAG-1 in mouse development. AB - BAG-1 is a family of proteins with diverse activities that range in cultured cells from protection against programmed cell death through to regulation of steroid hormone action. At least three proteins (BAG-1L, BAG-1M and BAG-1) are encoded by the Bag-1 mRNA through the use of alternative translation-initiation sites. To assess the in vivo function of these factors, we have used in situ hybridization and immunohistochemical techniques to determine the distribution of Bag-1 transcript and proteins during mouse development. Bag-1 mRNA was identified in several organs with cartilaginous tissues showing the highest expression levels. The level of expression at some of these sites was downregulated during the course of development. In the immunohistochemical studies, antibodies directed against the BAG-1 proteins stained all the sites identified in the in situ hybridization studies although isoform-specific differences were observed. BAG-1L specific antibody showed ubiquitous staining as early as day 10.5 post coitum but there was a progressive restriction during subsequent stages of embryogenesis. On the contrary, an antibody that preferentially recognized the other isoforms only stained the mouse myocardium in the early developmental stages before finally recognizing additional organs later on in development. These results demonstrate a stage- and site-specific expression of the BAG-1 isoforms during mouse development. PMID- 10704865 TI - Expression of sprouty2 during early development of the chick embryo is coincident with known sites of FGF signalling. AB - The Drosophila sprouty protein is a recently-identified intracellular modulator of FGF and EGF receptor tyrosine kinase activity which antagonises ras/MAP kinase signalling. In a differential display analysis to identify genes involved in patterning the mid/hindbrain region of the chick neural tube, we have identified a sprouty orthologue, sprouty2. Here we report expression of sprouty2 transcripts in the developing chick embryo. We find a close correlation with known sites of FGF activity but little correlation with expression patterns of members of the EGF family. Initially, transcripts are associated with the primitive streak. During the period of neural tube patterning expression is detected in the anterior neuropore, in the isthmic region and in neural plate and posterior spinal cord. Transcripts are also detected in the otic placode, tail bud, mesoderm of the branchial arches, somitic myotome, retina, limb buds and gut mesenchyme; all known sites of FGF action. PMID- 10704866 TI - Appropriate expression of the mouse H19 gene utilises three or more distinct enhancer regions spread over more than 130 kb. AB - H19 and Igf2 are closely linked, reciprocally imprinted genes which lie on distal chromosome 7 in the mouse. Data suggests that common elements are used for expression and imprinting of both genes, and simple models have been proposed based on the presence of a single set of enhancers located downstream of H19. In this study we have investigated the H19 expression pattern from a 130 kb YAC transgene, which imprints H19 appropriately at ectopic loci. However, we show that while enhancers for expression in many cell types are present on the YAC, those for expression in mesodermal components of the heart, kidney, lung and thymus are located at a greater distance. Based on the available evidence, we conclude that regulation of H19 is complex, requiring contribution from at least three different sets of cell-type specific enhancers. Thus, the mechanism of reciprocal imprinting of H19 and Igf2 utilises different regulatory elements in different cell types during mouse development. PMID- 10704868 TI - Identification of chick frizzled-10 expressed in the developing limb and the central nervous system. AB - We have identified chick frizzled (Fz)-10, encoding a Wnt receptor, and examined the expression pattern during embryogenesis. Fz-10 is expressed in the region posterior to the Hensen's node at stage 6. Fz-10 expression is detected in the dorsal domain of the neural tube and the central nervous system of the developing embryo. In the developing limb, Fz-10 expression starts at stage 18 in the posterior-dorsal region of the distal mesenchyme, and gradually expands to the anterior-distal region. Fz-10 is also expressed in the feather bud and branchial arch. Implantation of Sonic hedgehog (Shh)-expressing cells into the anterior margin of the limb bud resulted in the induction of Fz-10 expression in anterior dorsal mesenchyme. PMID- 10704867 TI - Transient cardiac expression of the tinman-family homeobox gene, XNkx2-10. AB - In Drosophila, the tinman homeobox gene is absolutely required for heart development. In the vertebrates, a small family of tinman-related genes, the cardiac NK-2 genes, appear to play a similar role in the formation of the vertebrate heart. However, targeted gene ablation of one of these genes, Nkx2-5, results in defects in only the late stages of cardiac development suggesting the presence of a rescuing gene function early in development. Here, we report the characterization of a novel tinman-related gene, XNkx2-10, which is expressed during early heart development in Xenopus. Using in vitro assays, we show that XNkx2-10 is capable of transactivating expression from promoters previously shown to be activated by other tinman-related genes, including Nkx2-5. Furthermore, Xenopus Nkx2-10 can synergize with the GATA-4 and SRF transcription factors to activate reporter gene expression. PMID- 10704869 TI - Correlation of asymmetric Notch2 expression and mouse incisor rotation. AB - Notch signaling defines an evolutionarily conserved cell communication mechanism, which enables neighboring cells to adopt different fates. Furthermore, Notch signaling may create boundaries that direct both the growth and patterning of the developing organs. Here we report on the expression of Notch receptors during the development of rodent incisors. Before the acquisition of their characteristic shape, incisors rotate antero-posteriorly and become asymmetric at their labial lingual axis. Notch2 is expressed only in the anterior part of the developing incisors, well before their rotation, while Notch2 expression was symmetrical in the developing molars. This is the first demonstration of an asymmetric gene expression pattern during the rotation of the rodent incisors. PMID- 10704870 TI - Co-localization of FAM and AF-6, the mammalian homologues of Drosophila faf and canoe, in mouse eye development. AB - The Drosophila fat facets and canoe genes regulate non-neural cell fate decisions during ommatidium formation. We have shown previously that the FAM (fat facets in mouse) de-ubiquitinating enzyme regulates the function of AF-6, (mammalian canoe homologue), in the MDCK epithelial cell line (Taya et al., 1998. The Ras target AF-6 is a substrate of the fam de-ubiquitinating enzyme. J. Cell Biol. 142, 1053 1062). We report here that the expression of the FAM and AF-6 proteins overlaps extensively in the mouse eye from embryogenesis to maturity, especially in the non-neural epithelia including the retinal pigment epithelium, subcapsular epithelium of the lens and corneal epithelium. Expression is not limited to the epithelia however, as FAM and AF-6 also co-localize during lens fibre development as well as in sub-populations of the neural retina. PMID- 10704871 TI - Differential expression of glycogen synthase kinase 3 genes during zebrafish embryogenesis. AB - Glycogen synthase kinase 3 (GSK-3) belongs to a highly conserved family of protein serine/threonine kinase whose members in high eukaryotes are involved in hormonal regulation, nuclear signaling, and cell fate determination. We have identified two zebrafish homologues related to mammalian GSK-3, ZGSK-3alpha and ZGSK-3beta. ZGSK-3alpha was expressed uniformly from cleavage onward, and later was found in many but not all tissues, especially in the central nervous system, spinal cord, somites and pronephric ducts. ZGSK-3beta was also transcribed maternally but the transcripts were not uniformly distributed during early cleavage stage. Most signals were concentrated in the inner part of the blastomeres. From midblastula stage onward, the ZGSK-3beta transcripts remained confined to inner parts of the deep cell layer. During shield stage, both epiblast and hypoblast expressed the transcripts. After late gastrulation, the signals were detected ubiquitously. During segmentation, prominent ZGSK-3beta signal was detected in head portion of the neural system. In the trunk, the expression was maintained in the neural tube and paraxial mesoderm and then became prominent in adaxial cells, followed by expression at the posterior region of somites. In pharyngula period ZGSK-3beta transcripts were distributed in similar regions as those of ZGSK-3alpha, namely, neural tissues of the head portion, spinal cord and somites. PMID- 10704872 TI - Expression patterns of two new members of the Semaphorin family in Drosophila suggest early functions during embryogenesis. AB - We report the sequence and expression analysis of two new Drosophila members of the Semaphorin family. Both proteins show the presence of Semaphorin domains and transmembrane domains. Both genes are expressed maternally and in embryos, and reveal distinct expression patterns much earlier than the onset of neurogenesis. We also present an overview of the domain structure of all so far known semaphorins in Drosophila. Furthermore, we compared all Drosophila and C. elegans Semaphorins and discuss them in the light of their evolution. PMID- 10704873 TI - Dynamic Lunatic fringe expression is correlated with boundaries formation in developing mouse teeth. AB - The formation of boundaries is a fundamental organizing principle during development. The Notch signalling pathway regulates this developmental patterning mechanism in many tissues. Recent data suggest that Notch receptors are involved in boundary determination during odontogenesis. It remains, however, uncertain if other components of the Notch pathway are also important for compartmental lineage restrictions in teeth. Here we report on the expression of the Lunatic fringe gene, which encodes a secreted signalling molecule regulating the Notch pathway, during the development of mouse teeth. Lunatic fringe is expressed in both epithelial and mesenchymal components of the developing molar. The expression pattern of Lunatic fringe in the epithelium is complementary to that of the Notch receptors. Lunatic fringe is asymmetrically expressed in the incisor epithelium during its antero-posterior rotation. This expression pattern defines the lingual comportment of the incisor epithelium whereas the labial comportment is defined by Notch2 expression. PMID- 10704874 TI - SRY, SOX9, and DAX1 expression patterns during human sex determination and gonadal development. AB - SRY, SOX9, and DAX1 are key genes in human sex determination, by virtue of their associated male-to-female sex reversal phenotypes when mutated (SRY, SOX9) or over-expressed (DAX1). During human sex determination, SRY is expressed in 46,XY gonads coincident with sex cord formation, but also persists as nuclear protein within Sertoli cells at 18 weeks gestation. High-level SOX9 expression in the sex cords of the testis parallels that seen during mouse development, however in humans, SOX9 transcripts also are detected in the developing ovary. Low-level DAX1 expression predates peak SRY expression by at least 10 days, and persists in Sertoli cells throughout the entire sex determination period. In Dosage Sensitive Sex reversal, the anti-testis properties of DAX1 over-expression could act prior to the peak effects of SRY and continue during the period of SOX9 expression. These findings highlight expression differences for the SRY, SOX9, and DAX1 genes during sex determination in humans and mice. These results provide a direct framework for future investigation into the mechanisms underlying normal and abnormal human sex determination. PMID- 10704875 TI - Zebrafish wnt4b expression in the floor plate is altered in sonic hedgehog and gli-2 mutants. AB - The floor plate of the neural tube serves an important function as a source of signals that pattern cell fates in the nervous system as well as directing proper axon pathfinding. We have cloned a novel zebrafish wnt family member, wnt4b, which is expressed exclusively in the floor plate. To place wnt4b in the context of known regulators of midline development, its expression was analyzed in the zebrafish mutants cyclops (cyc), floating head (flh), you-too (yot), and sonic you (syu). wnt4b expression in the medial and lateral floor plate are shown to be regulated independently: medial floor plate expression occurs in the absence of a notochord, while lateral floor plate expression requires a functional notochord, sonic hedgehog and gli-2. PMID- 10704876 TI - Expression of chick BMP-1/Tolloid during patterning of the neural tube and somites. AB - The expression pattern described here is that of the chick BMP-1/Tolloid family of secreted metalloproteinases during early stages of development. BMP-1/Tolloid transcripts are expressed in the blastoderm, at gastrulation stages and as the neural plate forms and neural tube folds, BMP-1/Tolloid is found at the neural plate/ectodermal transition. Expression is maintained in the premigratory neural crest, and transiently in the migrating cephalic neural crest cells. BMP 1/Tolloid is also expressed in the caudal, but not in the anterior notochord, and in the ventral neural tube at the time of dorso-ventral patterning. Further sites of BMP-1/Tolloid expression are the lateral plate mesoderm and the dermotome and the myotome of the somites. PMID- 10704877 TI - Developmental expression of a novel Ftz-F1 homologue, ff1b (NR5A4), in the zebrafish Danio rerio. AB - The Ftz-F1 genes encode orphan receptors of the nuclear receptor superfamily. The mammalian Ftz-F1 homologue, SF-1, has been found to be essential for the proper development of the adrenal-gonadal axis and it also plays a critical role in mammalian sex-determination. We report here the isolation and characterisation of a novel zebrafish Ftz-F1 gene, ff1b. Whole-mount in situ hybridization revealed onset of expression in the developing rostral diencephalon at 22 h post fertilization (h.p.f.). Later, at 30 h.p.f., transcripts could be detected in the anterior regions of the pancreatic anlagen. Expression in both locations peaks at 36 h.p.f. and disappears at around 48 h.p.f. PMID- 10704878 TI - The Wingless target gene Dfz3 encodes a new member of the Drosophila Frizzled family. AB - Here we report the identification of Dfz3, a novel member of the Frizzled family of seven-pass transmembrane receptors. Like Dfz2, Dfz3 is a target gene of Wingless (Wg) signalling, but in contrast to Dfz2, it is activated rather than repressed by Wg signalling in imaginal discs. We show that Dfz3 is not required for viability but is necessary for optimal Wg signalling at the wing margin. PMID- 10704879 TI - Conserved and divergent expression of T-box genes Tbx2-Tbx5 in Xenopus. AB - We report here the identification of four members of T-box family genes, Xltbx2 Xltbx5, in Xenopus. Two of them are probable pseudovariant genes of XTbx5 and ET, a putative Xenopus ortholog of Tbx3. We compared their expression patterns in both embryos and limbs. In embryos, expression of Xltbx2 and Xltbx3 showed novel diversities, such as Xltbx2 in the neural crest cells and Xltbx3 in the ventral spinal cord, together with mutual similarities in the following regions: dorsal retina, proctoderm, lateral line organ, cement gland and cranial ganglia. The patterns in limbs were highly conserved with mouse and chick orthologs, including the limb-type specific expression of Xltbx4 and Xltbx5. In addition, RT-PCR analysis showed that they are expressed weakly even in adult limbs as previously reported in the newt. PMID- 10704880 TI - The ActR-I activin receptor protein is expressed in notochord, lens placode and pituitary primordium cells in the mouse embryo. AB - ActR-I is a type I serine/threonine kinase receptor which has been shown to bind activin and bone morphogenetic proteins (BMPs). To study the function of ActR-I, we have generated novel monoclonal antibodies that specifically recognize the extracellular domain of mouse ActR-I. We examined the level of ActR-I protein during mouse development by immunohistochemistry. We found that in the embryonic body, ActR-I protein first appears in a restricted part of the primitive streak region and is present throughout the length of notochord. Furthermore, ActR-I protein is expressed in the facial sensory organ primordia, including eye area, otic vesicle and olfactory placode, which all contain invaginating ectoderm. In addition, ActR-I is produced in pituitary primordium (Rathke's pouch), mammary buds and the epithelial layer of branchial arches. Interestingly, in the lens placodes and in early Rathke's pouch, ActR-I protein is transiently localized at the apical surface of the epithelial cells, indicating the presence of an apical basal asymmetry in these cells. PMID- 10704881 TI - Vertebrate orthologues of the Drosophila region-specific patterning gene teashirt. AB - In Drosophila the teashirt gene, coding for a zinc finger protein, is active in specific body parts for patterning. For example, Teashirt is required in the trunk (thorax and abdomen) tagmata of the embryo, parts of the intestine and the proximal parts of appendages. Here we report the isolation of vertebrate cDNAs related to teashirt. As in Drosophila, human and murine proteins possess three widely spaced zinc finger motifs. Additionally, we describe the expression patterns of the two murine genes. Both genes show regionalized patterns of expression, in the trunk, in the developing limbs and the gut. PMID- 10704882 TI - GFP-tagged balancer chromosomes for Drosophila melanogaster. AB - We constructed green fluorescent protein (GFP)-expressing balancer chromosomes for each of the three major chromosomes of Drosophila melanogaster. Expression of GFP in these chromosomes is driven indirectly by a Kruppel (Kr) promoter, via the yeast GAL4-UAS regulatory system. GFP fluorescence can be seen in embryos as early as the germ band extension stage, and can also be seen in larvae, pupae, and adults. We show the patterns of GFP expression of these balancers and demonstrate the use of the balancers to identify homozygous progeny. PMID- 10704883 TI - Preface PMID- 10704884 TI - Salivary gland development in Drosophila melanogaster. AB - The Drosophila salivary gland is proving to be an excellent experimental system for understanding how cells commit to specific developmental programs and, once committed, how cells implement such decisions. Through genetic studies, the factors that determine where salivary glands will form, the number of cells committed to a salivary gland fate, and the distinction between the two major cell types (secretory cells and duct cells) have been discovered. Within the next few years, we will learn the molecular details of the interactions among the salivary gland regulators and salivary gland target genes. We will also learn how the early-expressed salivary gland genes coordinate their activities to mediate the morphogenetic movements required to form the salivary gland and the changes in cell physiology required for high secretory activity. PMID- 10704885 TI - Reiterative signaling and patterning during mammalian tooth morphogenesis. AB - Mammalian dentition consists of teeth that develop as discrete organs. From anterior to posterior, the dentition is divided into regions of incisor, canine, premolar and molar tooth types. Particularly teeth in the molar region are very diverse in shape. The development of individual teeth involves epithelial mesenchymal interactions that are mediated by signals shared with other organs. Parts of the molecular details of signaling networks have been established, particularly in the signal families BMP, FGF, Hh and Wnt, mostly by the analysis of gene expression and signaling responses in knockout mice with arrested tooth development. Recent evidence suggests that largely the same signaling cascade is used reiteratively throughout tooth development. The successional determination of tooth region, tooth type, tooth crown base and individual cusps involves signals that regulate tissue growth and differentiation. Tooth type appears to be determined by epithelial signals and to involve differential activation of homeobox genes in the mesenchyme. This differential signaling could have allowed the evolutionary divergence of tooth shapes among the four tooth types. The advancing tooth morphogenesis is punctuated by transient signaling centers in the epithelium corresponding to the initiation of tooth buds, tooth crowns and individual cusps. The latter two signaling centers, the primary enamel knot and the secondary enamel knot, have been well characterized and are thought to direct the differential growth and subsequent folding of the dental epithelium. Several members of the FGF signal family have been implicated in the control of cell proliferation around the non-dividing enamel knots. Spatiotemporal induction of the secondary enamel knots determines the cusp patterns of individual teeth and is likely to involve repeated activation and inhibition of signaling as suggested for patterning of other epithelial organs. PMID- 10704886 TI - Kidney morphogenesis: cellular and molecular regulation. AB - Development of an organ is directed by cell and tissue interactions and these also occur during the formation of functional kidney. During vertebrate development inductive signalling between mesenchyme and epithelium controls the organogenesis of all three kinds of kidneys: pronephros, mesonephros and metanephros. In higher animals the metanephros differentiates into the permanent kidney and in this review we will mainly concentrate on its development. Molecular interactions currently known to function during nephrogenesis have primarily been based on the use of knockout techniques. These studies have highlighted the role for transcription factors, signalling molecules, growth factors and their receptors and also for extracellular matrix components in kidney development. Finally in this review we will represent our own model for kidney development according to the knowledge of the genes involved in the development of the functional excretory organ, kidney. PMID- 10704887 TI - Pax genes and the differentiation of hormone-producing endocrine cells in the pancreas. AB - Despite the pivotal role of the pancreas in hormonally-regulated pathways in the body, e.g. glucose homeostasis, the genetic mechanisms defining it have for many years remained largely enigmatic. After years out of the spotlight, pancreas development has once again come to centre stage. To a large extent, this is due to recent advances made through the detailed analysis of transgenic mice which have been engineered to carry mutations in specific developmental control genes. This review specifically focuses on the specification of the endocrine pancreas lineage and in particular on the role of the developmental control genes Pax4 and Pax6 in the generation of specific endocrine cell types. The comparison of various phenotypes of different mouse mutants affecting endocrine development supports a model in which Pax4 and Pax6 are required for the differentiation of certain endocrine cell lineages and implies a potential for acting at different levels of endocrine development. PMID- 10704888 TI - The molecular basis of lung morphogenesis. AB - To form a diffusible interface large enough to conduct respiratory gas exchange with the circulation, the lung endoderm undergoes extensive branching morphogenesis and alveolization, coupled with angiogenesis and vasculogenesis. It is becoming clear that many of the key factors determining the process of branching morphogenesis, particularly of the respiratory organs, are highly conserved through evolution. Synthesis of information from null mutations in Drosophila and mouse indicates that members of the sonic hedgehog/patched/smoothened/Gli/FGF/FGFR/sprouty pathway are functionally conserved and extremely important in determining respiratory organogenesis through mesenchymal-epithelial inductive signaling, which induces epithelial proliferation, chemotaxis and organ-specific gene expression. Transcriptional factors including Nkx2.1, HNF family forkhead homologues, GATA family zinc finger factors, pou and hox, helix-loop-helix (HLH) factors, Id factors, glucocorticoid and retinoic acid receptors mediate and integrate the developmental genetic instruction of lung morphogenesis and cell lineage determination. Signaling by the IGF, EGF and TGF-beta/BMP pathways, extracellular matrix components and integrin signaling pathways also directs lung morphogenesis as well as proximo distal lung epithelial cell lineage differentiation. Soluble factors secreted by lung mesenchyme comprise a 'compleat' inducer of lung morphogenesis. In general, peptide growth factors signaling through cognate receptors with tyrosine kinase intracellular signaling domains such as FGFR, EGFR, IGFR, PDGFR and c-met stimulate lung morphogenesis. On the other hand, cognate receptors with serine/threonine kinase intracellular signaling domains, such as the TGF-beta receptor family are inhibitory, although BMP4 and BMPR also play key inductive roles. Pulmonary neuroendocrine cells differentiate earliest in gestation from among multipotential lung epithelial cells. MASH1 null mutant mice do not develop PNE cells. Proximal and distal airway epithelial phenotypes differentiate under distinct transcriptional control mechanisms. It is becoming clear that angiogenesis and vasculogenesis of the pulmonary circulation and capillary network are closely linked with and may be necessary for lung epithelial morphogenesis. Like epithelial morphogenesis, pulmonary vascularization is subject to a fine balance between positive and negative factors. Angiogenic and vasculogenic factors include VEGF, which signals through cognate receptors flk and flt, while novel anti-angiogenic factors include EMAP II. PMID- 10704889 TI - Liver specification and early morphogenesis. AB - The classically defined induction of the liver from the endoderm, elicited by the cardiac mesoderm, has recently been discovered to involve signaling by fibroblast growth factors (FGFs). Multiple FGFs induce hepatic gene expression independent of an effect on growth. A subset of these FGFs cooperates with other factors to promote morphogenesis of the newly specified hepatocytes. Subsequent to the formation of the liver bud, distinct mesenchymal signals and hepatic response pathways stimulate further growth and differentiation of the hepatic parenchymal cells and prevent apoptosis. The initial stages of hepatogenesis are therefore beginning to be understood, and serve as a paradigm for the development of other tissues from the endoderm. PMID- 10704890 TI - The battle of the sexes. AB - The sex determining gene, Sry, determines the sex of the organism by initiating development of a testis rather than an ovary from the cells of the bipotential gonad. In the 10 years since the discovery of Sry, new genes and cellular pathways that operate in the establishment of the gonadal primordium and the initiation of testis development have been discovered. Experiments defining mechanisms downstream of Sry are providing clear examples of how a regulatory transcription factor initiates cellular processes including proliferation and cell migration, which in turn influence architectural patterning, fate commitment, and differentiation of cells within an organ. PMID- 10704891 TI - Gastric pH profiles of beagle dogs and their use as an alternative to human testing. AB - Gastric pH levels were measured in samples of gastric aspirates from eight fasted beagle dogs. The gastric pH in fasting dogs fluctuated from 2.7 to 8.3, with a mean of 6.8+/-0.2 (SE). Each dog received the following four treatments in randomly-assigned order: (A) distilled water; (B) a placebo capsule; (C) pentagastrin, and (D) ranitidine. The gastric pH remained relatively constant after distilled water administration. In contrast, the treatments with pentagastrin and placebo capsule each lowered gastric pH. Pretreatment with pentagastrin was more successful in lowering gastric pH than that with placebo capsule. On the other hand, the pH rose above 7.0 in all dogs by the first hour after treatment with ranitidine. This animal model may be helpful in evaluating the biopharmaceutics of drugs exhibiting pH-dependent dissolution or decomposition. PMID- 10704892 TI - Increase of the duration of the anticonvulsive activity of a novel NMDA receptor antagonist using poly(butylcyanoacrylate) nanoparticles as a parenteral controlled release system. AB - A novel non-competitive NMDA receptor antagonist MRZ 2/576 is a potent but rather short-acting (5-15 min) anticonvulsant following intravenous administration to mice as estimated by the prevention of maximal electroshock induced convulsions. This is most probably due to a rapid elimination of the drug from the central nervous system by transport processes that are sensitive to probenecid. Intravenous administration of the drug bound to poly(butylcyanoacrylate) nanoparticles coated with polysorbate 80 prolongs the duration of the anticonvulsive activity in mice up to 210 min and after probenecid pre-treatment up to 270 min compared to 150 min with probenecid and MRZ 2/576 alone. The results of this study demonstrate that polysorbate 80 coated poly(butylcyanoacrylate) nanoparticles used so far as a delivery system to the brain for drugs that do not freely penetrate the blood brain barrier can also be used as a parenteral controlled release system to prolong the CNS availability of drugs that have a short duration of action. PMID- 10704893 TI - Enhancing effect of 5 alpha-cyprinol sulfate on mucosal membrane permeability to sodium ampicillin in rats. AB - Effect of 5 alpha-cyprinol sulfate, a bile alcohol sulfate specific to carp bile, on rectal membrane permeability to sodium ampicillin (AMP Na) was examined in rats. AMP Na is not easily absorbed through rat rectal membrane without aid. 5 alpha-Cyprinol sulfate significantly enhanced the rectal membrane permeability to AMP Na even at a low concentration (6.25 mM), though sodium taurocholate needed a higher concentration (25 mM). Co-administration of phosphatidylcholine significantly suppressed the enhancing action of both sodium taurocholate and 5 alpha-cyprinol sulfate. On the other hand, calcium ion did not suppress the action of 5 alpha-cyprinol sulfate, although it did clearly suppress the action of sodium taurocholate. In conclusion, 5 alpha-cyprinol sulfate was found to have a potent enhancing effect on mucosal membrane permeability to water-soluble compounds. The enhancing mechanism of 5 alpha-cyprinol sulfate appeared to be different from that of sodium taurocholate. PMID- 10704894 TI - Improvement of availability of allopurinol from pharmaceutical dosage forms I - suppositories. AB - Solid dispersion and crystallization of a very slightly water-soluble drug, allopurinol, were prepared using urea, sodium salicylate and beta-cyclodextrin (beta-CD) as carriers. The spectroscopic infra-red (IR), differential scanning calorimetry (DSC) and powder X-ray diffractometry (PXRD) data indicate a role of these carriers in decreasing the crystallinity of allopurinol and complexing abilities. Solid dispersion and crystallization of the drug with these carriers were used in suppository formulations to investigate their role in enhancement of drug release through the membrane barrier. The bases used included Suppocire AM and the mixture of polyethylene glycols (PEGs). The release rates of allopurinol from lipophilic and hydrophilic suppository bases were examined and compared with those obtained for their inclusion compounds incorporated in the same bases. The prepared suppositories were evaluated for in-vitro drug release, when fresh and on storage. The release of pure allopurinol from the lipophilic base was remarkably higher than that from the hydrophilic one. The release of allopurinol from lipophilic as well as hydrophilic bases was significantly enhanced by crystallization of the drug from 5% w/v of sodium salicylate. Allopurinol crystallized from sodium salicylate, showed enhanced release reaching about 100% in 1 h from the Suppocire AM base. The obtained data from these experiments proved the superiority of the PEG formulations containing coevaporates of the drug to sodium salicylate, ratio 1:1, or of the drug to beta-CD, ratio 1:2; T(90%),12 and 36 min, respectively. A significant decrease of uric acid excretion in rabbits was observed after rectal administration of suppositories containing allopurinol crystallized from sodium salicylate. PMID- 10704895 TI - Effect of dose on cyclosporine-induced suppression of hepatic cytochrome P450 3A2 and 2C11. AB - Cyclosporine is a potent immunosuppressive drug that undergoes extensive hepatic metabolism catalyzed primarily by the cytochrome P450 (P450) 3A enzyme family. Cyclosporine alters its own metabolism by selective suppression of specific P450 isoforms after chronic therapy in rats. Modulation of hepatic P450 by chronic cyclosporine dosing is associated with increased blood concentrations leading to nephropathy. However, the relationship between cyclosporine dose and hepatic enzyme suppression is not known. The purpose of this study was to examine the effect of escalating doses of cyclosporine on P450 regulation and metabolic activity in the rat. Following 1 week of a low-salt diet, rats were given cyclosporine 5, 15, 30 or 50 mg/kg per day or an equal volume of vehicle for 2 weeks via oral gavage. At the end of the dosing period, livers were removed and hepatic microsomes prepared. Hepatic P450 proteins were measured using Western blot analysis and catalytic activity determined by in vitro testosterone hydroxylation. Cyclosporine dosing suppressed both P450 3A2 and 2C11 protein expression and catalytic activity in a dose-dependent manner. Catalytic activity of two other P450 isoforms, 2A1 and 2B1, were unchanged by cyclosporine administration. Thus, the selective suppression of hepatic microsomal P450 by cyclosporine is not only dependent on the length of therapy, but also the dose administered. PMID- 10704896 TI - Allopurinol encapsulated in polycyanoacrylate nanoparticles as potential lysosomatropic carrier: preparation and trypanocidal activity. AB - The activity of allopurinol-loaded polyethylcyanoacrylate nanoparticles against Trypanosoma cruzi was compared to that of free allopurinol using in vitro cultures of epimastigotes. Ethylcyanoacrylate nanoparticles were prepared by an emulsion polymerization process, and formulations containing different concentrations of allopurinol, polyethylcyanoacrylate and surfactants were investigated and analyzed in size and amount of drug entrapped. The nanoparticles obtained were less than 200 nm in size, as measured by electron microscopy and cytometry. The peak amount of allopurinol entrapped in the nanoparticles was 62.8+/-1.9 microg mg(-1) of nanoparticles using 400 microl of polyethylcyanoacrylate, 200 microl of surfactant (Tween 20) and 20 mg of allopurinol in 50 ml of polymerization medium and the association efficiency was 100.7%. After 6 h of incubation at pH 7.4 the release of allopurinol from the nanoparticles was 7.4%, while at pH 1.2 only 3.1% was released after 4-6 h (t=42.8, P<0.0001). The in vitro studies, using cultures of T. cruzi epimastigotes, demonstrated considerable increases in the trypanocidal activity of the allopurinol-loaded nanoparticles in comparison with a standard solution of allopurinol (91.5 vs. 45.9%) at an allopurinol concentration of 16.7 microg ml( 1). In addition, it was shown that the unloaded nanoparticles, by mechanisms not completely elucidated, had a trypanocidal activity similar to that of standard solutions of allopurinol. To study cytotoxicity, increasing concentrations of unloaded nanoparticles were incubated on vero-line cell cultures. The concentration that killed 50% cells was 200 microg ml(-1), four times higher than that necessary to kill 50% of T. cruzi. It is concluded that the polyethylcyanoacrylate nanoparticles constitute a good carrier of drugs against the T. cruzi. The allopurinol loaded-nanoparticles significantly increased the trypanocidal activity in comparison to the free drug. PMID- 10704897 TI - The influence of alkali fatty acids on the properties and the stability of parenteral O/W emulsions modified with solutol HS 15. AB - Arachis oil based parenteral O/W emulsions were prepared using soya bean phosphatidylcholine (SPC) and different combinations of co-emulsifiers containing polyethylene glycol fatty acid esters (Solutol HS 15) and alkali fatty acids (sodium laurate, sodium stearate). The parameters measured were droplet size (both by photon correlation spectroscopy and laser diffractometry), pH and zeta potential. All emulsions were subjected to autoclaving. The addition of polyethylene glycol 12-hydroxy stearate (Solutol HS 15) led to a significant decrease of mean oil droplet size. For long-term stability the amount added turned out to be the most important factor. With increased amounts of Solutol HS 15 the packing density of the emulsifier layer and the zeta potential decreased leading to instability. The optimum load of Solutol HS 15 was found to be 15 micromol/ml. Alkali fatty acids markedly improved the physical stability of the emulsions. Improved stability properties conferred to emulsions by alkali fatty acids could be attributed to the zeta potential increase even in the presence of Solutol HS 15. Consequently a mixed emulsifier film was established in which the ionized fatty acids determined the interface charge. In addition to this a strengthening of the molecular interactions occurring between phospholipid and Solutol HS 15 emulsifier in the presence of ionized fatty acids at the O/W interface can be assumed (L. Rydhag, The importance of the phase behaviour of phospholipids for emulsion stability, Fette Seifen Anstrichm. 81 (1979) 168-173). Different co-emulsifier mixtures were shown to have a pronounced impact on the plasma protein adsorption onto emulsion droplets. PMID- 10704898 TI - A new theoretical approach to tablet strength of a binary mixture consisting of a well and a poorly compactable substance. AB - The objective of this study was to analyse the tensile strength of a well and a poorly compactable substance in a tablet mixture. Recent developments in the theory of percolation were taken into account and two power laws are proposed, one for the tensile strength as a function of the relative density of the mixture, and the other for the relationship between the strength and compaction pressure. Both equations are assumed to be valid in a comparatively low pressure range. A universal testing instrument Zwick UPM 1478 was used for the manufacture and testing of the compacts. Mixtures of Avicel PH101 and paracetamol at different ratios were chosen as model systems. The experimental results showed that the proposed model equations fitted the experimental data reasonably well for all mixture ratios. It was observed that the critical solid fraction of the mixture, i.e. the strength percolation threshold, increased with rising amounts of the drug. We investigated the strength threshold not only in terms of the solid fraction, but also in terms of the mass fraction (excipient percolation threshold). It is assumed that a tablet can only be produced with a certain minimal amount of the well compactable substance that is needed to build a percolating cluster in the tablet. An interpretation is therefore provided for the dilution capacity of a direct tableting excipient with a poorly compactable drug. The dilution capacity was experimentally determined according to the method of Minchom and Armstrong (Br. Pharm. Conf. (1987) 69 pp.). Our experimental estimate of 79.9% drug is in perfect agreement with our proposed theoretical calculation of 79.7%. These estimates are, however, much higher than the one reported in a recent study (Y. Habib, L. Augsburger, G. Reier, Th. Wheatley, R. Shangraw, Dilution potential: a new perspective, Pharm. Dev. Tech. 1 (2) (1996) 205-212) where the dilution capacity of the same mixture was investigated. This discrepancy can be explained based on the different pressure ranges and extrapolation techniques that were used. As a conclusion, concepts of the percolation theory can successfully be applied to the kind of mixture studied in this paper. It is conceivable that the theoretical tools presented can also be applied to mixtures of more than two substances if they consist of a single well compactable excipient and several poorly compactable components. Such mixtures are relevant for the development of direct compressible tableting formulations. PMID- 10704899 TI - Diffusional characteristics of freeze/thawed poly(vinyl alcohol) hydrogels: applications to protein controlled release from multilaminate devices. AB - The incorporation of a model protein, bovine serum albumin (BSA), into poly(vinyl alcohol) (PVA) hydrogel films during the freezing and thawing process and its subsequent release behavior were investigated. The effect of the number of freezing and thawing cycles as well as the stability of BSA were examined. BSA release profiles were not significantly different from gels prepared after 3 or 5 cycles. However, the rate and overall amount of PVA dissolution were considerably higher for gels prepared after 3 cycles. These observations were then applied to the development of novel, freeze/thawed PVA laminates. Laminates containing gel layers prepared after 3 or 5 cycles were successfully prepared with good stability over a 6 month swelling period. These structures, containing distinct layers of very specific properties, could be used to achieve zero-order release behavior. PMID- 10704900 TI - Diffractive optical element based sensor for surface quality inspection of concave punches. AB - Optical surface quality of concave shaped punches was investigated with a diffractive optical element based sensor. Image information of the present sensor was studied for the purpose of surface roughness and reflectance inspection. The robust sensor is proposed for off-line optical inspection of concave punches. PMID- 10704901 TI - The automatic micrometer screw. AB - A new analytical method - the automatic micrometer screw - has been established to measure the edge height of tablets. The equipment offers many advantages compared with other methods. The precision is slightly increased compared to the traditional micrometer screw and the measurement with a small punch and a linear voltage transducer. No longer any touch of the tablet is necessary and influences results. The method works automatically and continuously, no manual measurement of the tablets is necessary. Up to ten tablets can be analyzed at the same time because of a rotary table on which they are positioned. Thus the method is not personal intensive. By combining the results from the measurement of punch displacement which means tablet height in the die and the results of the measurement with the automatic micrometer screw which means tablet height outside the die, a convenient measurement for the decompression process is possible. PMID- 10704902 TI - Modeling of theophylline release from different geometrical erodible tablets. AB - The aim of this study is to reveal statistically how various geometrical shapes such as triangle, cylinder, half-sphere affect the release rate of the active substance called theophylline in erodible hydrogel matrix tablets. We have tried to indicate these changes in the release rate of theophylline by supporting our aim with the mathematical equations developed by Hopfenberg and Katzhendler et al. The model developed by Hopfenberg assumes that drug release occurs from the primary surface area of the device but Katzhendler et al. (I. Katzhendler, A. Hoffman, A. Goldberger, M. Friedman, Modelling of drug release from erodible tablets, J. Pharm. Sci. 86 (1997) 110-115), described a situation where the erosion rates of the tablet are different in the radial and axial directions. Hydrogel matrix tablets were prepared with hydroxypropylmethylcellulose (HPMC E(50)) possessing different geometrical shapes as triangular, cylindrical and half-spherical using experimental design. When the dissolution results have been evaluated, it has been observed theophylline release from different geometrical erodible tablets fitted with that of the Katzhendler et al., equation. This equation which was suggested for cylindrical tablets was also used to interpret half-spherical and triangular tablets. According to the above stated equation, n has been determined as 4 for triangular tablets and 1.5 for half-spherical tablets and we have also suggested that, these n values could be used in the kinetic programs. PMID- 10704903 TI - Interaction of Bay t 3839 coprecipitates with insoluble excipients. AB - Bay t 3839 is an extremely sparingly soluble drug and results in severe problems with regard to oral bioavailability. In order to improve the dissolution performance a coprecipitate formulation approach was investigated and the necessary ratio polyvinylpyrrolidone (pvp) to drug substance was found to be approximately 15 in order to achieve a sufficient supersaturation in 900 ml 0.1 N HCl, which is considered to be an appropriate as well as challenging in vitro test method. Surprisingly substantial differences in the dissolution characteristics of coprecipitate granulates were revealed when the coprecipitate was formed on excipients which was not the case when pure pvp/drug coprecipitates were submitted to dissolution testing. As substantial deterioration of the supersaturation capability was detected only in the case of insoluble excipients it is concluded that solid/liquid interfaces during the dissolution step act as recrystallization inducers. In case of soluble fillers/adsorbents, like mannitol, the supersaturation capability could be maintained also after the granulation step. Therefore, it is likely that in the case of extremely challenging drug candidates like, e.g. Bay t 3839, the supersaturated solution is less stable than in the case of more soluble compounds. These more susceptible coprecipitates can be stabilized by proper selection of the granulation excipients. Additionally the admixture of crystallization inhibiting agents like sodium dodecyl sulfate (SDS) offers the opportunity to decrease the necessary ratio pvp/Bay t 3839 considerably (to a ratio of approximately 7). PMID- 10704904 TI - In vitro and in vivo characterization of biodegradable enoxacin microspheres. AB - The in vitro release and plasma concentration profiles of sustained release enoxacin microspheres intended for the treatment of bone and systemic infections due to sensitive strains of bacteria were investigated. Microspheres of enoxacin were prepared by using poly(glycolic acid-co-DL-lactic acid) (PLGA) by the emulsion solvent evaporation technique and characterized by in vitro release in an incubator, and in vivo release in the rat subcutaneous model. The microspheres were spherical in nature, and particle size range had a significant influence on the in vitro release. The enoxacin plasma concentration 2 h after the administration of treatments was two-fold higher in animals who received the free drug compared with those who received microspheres of size range 125-250 microm. The plasma of animals who received the free drug was depleted of enoxacin by the end of the first day. However, the plasma concentration of enoxacin in the animals who received microspheres was sustained above 0.5 microg/ml for about 8 days. The results show that biodegradable microspheres of enoxacin can be prepared which release the antibiotic in vivo for days following a subcutaneous administration. This should provide a means for the sustained treatment of infections due to sensitive strains of bacteria. PMID- 10704905 TI - The truncated estrogen receptor alpha variant lacking exon 5 is not involved in progesterone receptor expression in meningiomas. AB - Human meningioma tissues are mostly estrogen receptor (ER) negative and progesterone receptor (PR) positive in ligand binding and enzyme immuno assays. To explain this apparently ER independent PR expression, we investigated the existence of a 'hidden' ER variant, which would be capable of activating transcription of the PR gene. Total RNA of seven meningiomas, two breast cancer tissues and of MCF7 cells was analyzed by RT-PCR using primers situated in exon 4 and exon 6. Differential hybridization of the PCR transcripts with probes in exon 4 and 5 respectively, revealed a wild type ER (wtER) fragment and an exon 5 deleted ER variant (ERDelta5). PCR products of two meningiomas were cloned for sequence analysis. The result confirmed the existence of a wtER and ERDelta5.RT PCR followed by Southern analysis was performed on mRNA of 23 meningiomas to determine the amount of ERDelta5 relative to wtER, which was compared to the PR content of the tissues. In contrast to our initial hypothesis and literature data on breast cancer, there was no relationship between the ERDelta5/wtER ratio and PR protein concentration. It is therefore concluded that ERDelta5 mRNA does not play the dominant role in PR synthesis in meningioma tissue. PMID- 10704906 TI - 19-oxygenations of 3-deoxy androgens, potent competitive inhibitors of estrogen biosynthesis, with human placental aromatase. AB - Aromatase is a cytochrome P450 enzyme complex that catalyzes the conversion of androst-4-ene-3,17-dione (AD) to estrone through three sequential oxygenations of the 19-methyl group. To gain insight into the ability of 3-deoxy derivative of AD, compound 1, and its 5-ene isomer 4, which are potent competitive inhibitors of aromatase, to serve as a substrate, we studied their 19-oxygenation by human placental aromatase and the metabolites isolated were analyzed by gas chromatography-mass spectrometry. Inhibitors 1 and 4 were found to be oxygenated with aromatase to produce the corresponding 19-hydroxy derivatives 2 and 5 and 19 oxo derivatives 3 and 6 as well as the 17beta-reduced 19-hydroxy compounds 7 and 8. Kinetic studies indicated that the 5-ene steroid 4 was surprisingly a good substrate for the aromatase-catalyzing 19-oxygenation with the V(max) value of 45 pmol/min per mg prot which was approx. four times higher than that of the other. The relative K(m) value for steroids 1 and 4 obtained in this study is opposite from the relative K(i) value obtained previously in the inhibition study. The results reveal that there is a difference between a binding suitable for serving as an inhibitor of aromatase and a binding suitable for serving as a substrate of the enzyme in the 3-deoxy steroid series and the C-3 carbonyl group of AD is essential for a proper binding as a substrate to the active site of aromatase. PMID- 10704907 TI - Differential cross-talk of estrogen and growth factor receptors in two human mammary tumor cell lines. AB - Cultured human mammary MCF7 and T47D tumor cell lines were used to test the interference of the partial antiestrogen 4'-hydroxytamoxifen (4-OH-TAM) and the pure antiestrogen ZM 182780 with growth factor (IGF-I, heregulin) signaling pathways. Growth of both cell lines was stimulated by IGF-I (20 ng/ml) or heregulin (3 nM). ZM 182780 effectively blocked growth factor induced as well as basal proliferation of MCF7 cells while the compound was ineffective in interfering with growth factor mitogenic activity in T47D cells. On both cell lines the IGF-I or heregulin- induced proliferation was enhanced further by 4-OH TAM. This synergism could be inhibited dose-dependently by ZM 182780. When cells were grown in the presence of estradiol plus growth factors, the antiestrogenic potencies of both compounds and the efficacy of ZM 182780 were unaffected, while the efficacy of 4-OH-TAM was reduced. Our data show cell type specific cross-talk between the receptor for estrogen and that for IGF-I or heregulin, which is different in MCF7 and T47D cells, respectively. In MCF7 cells with demonstrable cross-talk, a clear superiority exists for a pure antiestrogen over a partial agonist in interfering with growth factor mitogenic activity. PMID- 10704909 TI - Uptake of dehydroepiandrosterone-3-sulfate by isolated trophoblasts from human term placenta, JEG-3, BeWo, Jar, BHK cells, and BHK cells transfected with human sterylsulfatase-cDNA. AB - The human placenta lacks the enzyme 17alpha-hydroxylase/17-20-lyase, and is thus unable to convert cholesterol into estrogens. Therefore estrogen synthesis of trophoblast cells depends on the supply of precursors such as dehydroepiandrosterone-3-sulfate (DHEA-S) and 16alpha-hydroxy dehydroepiandrosterone-3-sulfate by maternal and fetal blood. To investigate the cellular internalisation of these anionic hydrophilic precursors, the uptake of [(3)H]-/[(35)S]-DHEA-S and [(3)H]-taurocholate by isolated cytotrophoblasts, cells of choriocarcinoma cell lines (JEG-3, BeWo, Jar), BHK and BHK cells transfected with human sterylsulfatase-cDNA (BHK-STS cells) was studied. Furthermore, the activity of sterylsulfatase of these cells in suspension and in corresponding cell homogenate was measured. During the first 5 min of incubation with [(3)H]-DHEA-S or [(35)S]-DHEA-S, radioactivity of cytotrophoblasts increased significantly, while radioactivity of JEG-3, Jar, BHK and BHK-STS cells did not increase. Radioactivity of BeWo cells increased slightly. For all cell types, there was no significant difference for uptake of either substrate. During incubation with [(3)H]-taurocholate, radioactivity of cytotrophoblasts did not increase. Sterylsulfatase activity of cytotrophoblast homogenate was significantly lower than that of cytotrophoblast suspension. Sterylsulfatase activity of BHK-STS, JEG-3 or BeWo cell homogenate was significantly higher than that of the corresponding cell suspension. In BHK and Jar cells sterylsulfatase activity was not detectable. Cytotrophoblasts take up DHEA-S without prior hydrolysis. BHK, BHK-STS, JEG-3, and Jar cells do not take up and BeWo cells slowly take up DHEA-S. In cytotrophoblasts extracellular DHEA-S rapidly gains access to intracellular sterylsulfatase, while in choriocarcinoma and BHK-STS cells access of DHEA-S to sterylsulfatase is limited. Our results indicate, that uptake by cytotrophoblasts is mediated by a carrier which is not expressed in choriocarcinoma or BHK cells and which is different from the known taurocholate transporting organic anion transporting polypetides. PMID- 10704908 TI - Bovine adrenal 3beta-hydroxysteroid dehydrogenase (E.C. 1.1.1. 145)/5-ene-4-ene isomerase (E.C. 5.3.3.1): characterization and its inhibition by isoflavones. AB - The isoflavones daidzein, genistein, biochanin A and formononetin inhibit potently and preferentially the gamma-isozymes of mammalian alcohol dehydrogenase (gammagamma-ADH), the only ADH isozyme that catalyzes the oxidation of 3beta hydroxysteroids. Based on these results, we proposed that these isoflavones might also act on other enzymes involved in 3beta-hydroxysteroid metabolism. Recently, we showed that they indeed are potent inhibitors of a bacterial beta hydroxysteroid dehydrogenase (beta-HSD). To extend this finding to the mammalian systems, we hereby purified, characterized and studied the effects of isoflavones and structurally related compounds on, a bovine adrenal 3beta-hydroxysteroid dehydrogenase (3beta-HSD). This enzyme catalyzes the oxidation of 3beta hydroxysteroids but not 3alpha-, 11beta- or 17beta-hydroxysteroids. The same enzyme also catalyzes 5-ene-4-ene isomerization, converting 5-pregnen 3, 20-dione to progesterone. The K(m) values of its dehydrogenase activity determined for a list of 3beta-hydroxysteroid substrates are similar (1 to 2 microM) and that of its isomerase activity, determined with 5-pregnen 3, 20-dione as a substrate, is 10 microM. The k(cat) value determined for its isomerase activity (18.2 min(-1)) is also higher than that for its dehydrogenase activity (1.4-2.4 min(-1)). A survey of more than 30 isoflavones and structurally related compounds revealed that daidzein, genistein, biochanin A and formononetin inhibit both the dehydrogenase and isomerase activity of this enzyme. Inhibition is potent and concentration dependent. IC(50) values determined for these compounds range from 0.4 to 11 microM, within the plasma and urine concentration ranges of daidzein and genistein of individuals on vegetarian diet or semi-vegetarian diet. These results suggest that dietary isoflavones may exert their biological effects by inhibiting the action of 3beta-HSD, a key enzyme of neurosteroid and/or steroid hormone biosynthesis. PMID- 10704910 TI - 7alpha-methyl-19-nortestosterone, a synthetic androgen with high potency: structure-activity comparisons with other androgens. AB - CNNT. There was a good correlation between bioactivity and binding affinity to AR for the 7alpha-substituted androgens compared to T. In contrast, relative to their binding affinity to AR, the androgenic potency of DHT and 19-NT was lower compared to T. The reason for the lower in vivo androgenic activity of 19-NT is attributable to its enzymatic conversion to 5alpha-reduced-19-NT in the prostate. In the case of DHT, the lower bioactivity could be attributed to its faster metabolic clearance rate relative to T. The correlation was further investigated in vitro by co-transfection of rat ARcDNA expression plasmid and a reporter plasmid encoding the chloramphenicol acetyl transferase (CAT) gene driven by an androgen inducible promoter into CV-1 cells. All the androgens led to a dose dependent increase in the CAT activity. MENT was found to be the most potent followed by DHT, 19-NT, T, and CNNT. The specificity of the androgenic response was confirmed by its inhibition with hydroxyflutamide, an antiandrogen. Thus, there was a good correlation between binding affinity and in vitro bioactivity in the transient transfection assay for the androgens. This suggests that the in vivo bioactivity of androgens could be influenced not only by binding affinity to receptors but also by factors such as absorption, binding to serum proteins and metabolism. However, the high potency of MENT is primarily related to its higher affinity to AR. PMID- 10704911 TI - Estrogen synthesis in human colon cancer epithelial cells. AB - Epidemiological and experimental data suggest an involvement of estrogen in the development and progression of colorectal cancer. In order to determine whether local synthesis of estrogen occurred in human colonic cancer cells, two colorectal cancer cell lines, HCT8 and HCT116, were evaluated for gene expression and enzyme activity of cytochrome P450 aromatase. In addition, the effect on aromatase expression of charcoal-stripped fetal calf serum, of quercetin and genistein and of tamoxifen and raloxifene was investigated in both cell lines. RT PCR analysis revealed that colorectal adenocarcinoma cell lines contain aromatase as a major component. The conversion of [(3)H]-androstenedione to estrone and labeled water was dose-dependently inhibited by 4-hydroxyandrostenedione and obeyed Michaelis-Menten kinetic with apparent Km values of approximately 20 nM and V(max) values of approx. 200 and 500 fmol/mg protein/h for HCT8 and HCT116 cells, respectively. After 24 h incubation, genistein (1 microM) significantly increased aromatase activity in HCT8 cells, with no effect on HCT116 cells. In accord with previous observation in reproductive tissues, quercetin (1 microM) significantly inhibited the enzyme activity in both cell lines. Also tamoxifen (100 nM) acted as inhibitor, while raloxifene (10 nM) decreased the enzyme activity only in HCT116 cells. The aromatase gene expression modulation by these effective agents was consistent with their effects on enzyme activity. These findings demonstrate for the first time that colorectal adenocarcinoma cell lines express aromatase. Interestingly, the enzyme activity was inhibited by quercetin, one major dietary flavonoid, by tamoxifen, a hormonal therapeutic agent for breast cancer, and by raloxifene, used in the prevention of postmenopausal osteoporosis. PMID- 10704912 TI - Immunoassay of 7-hydroxysteroids: 2. Radioimmunoassay of 7alpha-hydroxy dehydroepiandrosterone. AB - High sensitivity radioimmunoassay of 3beta, 7alpha-dihydroxy-5-androsten-17-one (7alpha-OH-DHEA) has been developed and evaluated. The method is based on polyclonal rabbit antisera raised against 19-O-(carboxymethyl)oxime bovine serum albumin conjugate and bridge- and position homologous [(125)I]iodotyrosine methyl ester as a tracer. Sensitivity of the assay amounted to 3.12 fmol (0.95 pg)/tube, precision as a mean intra- and interassay coefficient of variation was 7.1 and 10.6%, respectively, and the average recovery of the analyte added to steroid free serum was 110%. Out of the steroids occurring in human serum which may interfere with the assay, the only important cross-reactants were dehydroepiandrosterone and 3beta, 7beta-dihydroxy-5-androsten-17-one (7beta-OH DHEA) with cross-reactivities of 1.95 and 1.16%, respectively. The levels of free (unconjugated) 7alpha-OH-DHEA have been determined in 29 sera from healthy volunteers (23 females and 6 males), and from 48 patients (43 females and 5 males) in which dehydroepiandrosterone and its sulfate (DHEA/S) had been measured for various endocrinopathies. The levels in healthy subjects ranged from 0.21 to 6.57 (mean 2.33+/-1.50) nM, while those of the patients from 0 to 5. 99 (mean 1.46+/-1.52) nM. The levels of 7alpha-OH-DHEA in patients significantly correlated with those of DHEA and its sulfate. PMID- 10704913 TI - Pregnenolone metabolized to 17alpha-hydroxyprogesterone in yeast: biochemical analysis of a metabolic pathway. AB - The cDNA coding for the human 3beta-hydroxy-5-ene steroid dehydrogenase/5-ene-4 ene steroid isomerase (3beta-HSD) has been expressed in yeast. When expressed from identical vectors except for the coding sequence, the specific activity of the type I is lower than that of the type II enzyme. A mutant of the human 3beta HSD type II lacking the putative membrane spanning domain 1 was generated by site directed mutagenesis: its apparent K(m) for pregnenolone (PREG) is significantly increased and its V reduced to the level of the type I enzyme. The influence of the kinetic properties of 3beta-HSD in the accumulation of 17alpha hydroxyprogesterone was probed by co-expression of the bovine 17alpha-hydroxylase cytochrome P450 (P45017alpha) cDNA. The metabolism of PREG was followed with time using the membrane fraction. Kinetic properties of the 3beta-HSD were modulated such that its activity was in excess, limiting or balanced with respect to the activity of the P45017alpha and the accumulation of intermediates and products recorded. Conditions for the generation of the by-products resulting from the 17,20-Lyase activity of the P45017alpha were found. The potential applications of the system are discussed. PMID- 10704914 TI - Rectangular solid domains in ceramide-cholesterol monolayers - 2D crystals. AB - Very small rectangular domains were observed by atomic force microscopy in binary monolayers of synthetic ceramides and cholesterol. When the cholesterol content is increased the domains are bigger although the rectangular shape is retained. The almost perfect shape of the domains indicates two-dimensional single ceramide crystals. Lipid domains in monolayers of this particular shape and size have to our knowledge not been reported in the literature previously. PMID- 10704915 TI - Expression of rat, renal NHE2 and NHE3 during postnatal development. AB - The current studies were designed to characterize the expression of sodium hydrogen exchangers NHE2 and NHE3 during rat, renal ontogeny. NHE2 mRNA and immunoreactive protein were more highly expressed at 2 and 3 weeks of age, with declining levels into adulthood. In situ hybridization of NHE2 mRNA localized the message to the renal inner cortex and outer medullary regions and suggested higher mRNA levels in suckling animals as compared to adults. Immunohistochemical analysis of rat kidney with the NHE2 antiserum showed specific staining of the distal convoluted tubules. In contrast, NHE3 mRNA expression was lowest in 2-week animals and higher in older rats, while NHE3 immunoreactive protein showed constant expression levels during development. Additionally uptake experiments utilizing HOE694 showed no change in NHE2 or NHE3 functional protein expression in 2-week-old rats versus adults. We conclude that the developmental increase in NHE2 mRNA and immunoreactive protein expression cannot be detected by functional assays, which suggests that NHE2 does not play a role in sodium absorption by the renal tubules (as has been previously suggested). Additionally, molecular changes seen in NHE3 mRNA expression do not affect functional protein activity, suggesting increased mRNA translational efficiency or protein stability in suckling rats. PMID- 10704916 TI - Cold shock in Bacillus subtilis: different effects of benzyl alcohol and ethanol on the membrane organisation and cell adaptation. AB - A temperature shift-down of Bacillus subtilis from 40 to 20 degrees C induces an 80 min growth lag. Benzyl alcohol reduced this period to 51 min, whereas ethanol prolonged it up to 102 min. The effect of the two alcohols on the membrane state was investigated by measuring the steady-state fluorescence anisotropy and analysing the lifetime distribution of diphenylhexatriene (DPH) and its polar derivative, TMA-DPH. As followed from the fluorescence anisotropy, the two alcohols exerted similar (fluidizing) effects on the cytoplasmic membranes of B. subtilis. However, benzyl alcohol significantly shortened the main DPH lifetime component and widened its distribution, while ethanol had no effect. The benzyl alcohol activity was interpreted in terms of an increased membrane hydration due to disordering of the membrane structure. Such an effect imitates the cold shock induced synthesis of unsaturated fatty acids in B. subtilis. The fatty acid analysis revealed that ethanol hindered this adaptive synthesis of fatty acids. At the same time, its effect on the membrane state (membrane order) was very low and could not substitute the physiological response as was the case with benzyl alcohol. It can thus be concluded that the adaptation of the membrane physical state contributes significantly to the cold shock response of B. subtilis. PMID- 10704917 TI - Location of tryptophan residues in free and membrane bound Escherichia coli alpha hemolysin and their role on the lytic membrane properties. AB - alpha-hemolysin (HlyA) is an extracellular protein toxin secreted by Escherichia coli that acts at the level of plasma cell membranes of target eukaryotic cells. Previous studies showed that toxin binding to the bilayers occurs in at least two ways, a reversible adsorption and an irreversible insertion. Studies of HlyA insertion into bilayers formed from phosphatidylcholine show that insertion is accompanied by an increase in the protein intrinsic fluorescence. In order to better define structural parameters of the membrane-bound form, the location of tryptophan residues was studied by means of quenchers of their intrinsic fluorescence located at 7, 12 and 16 positions of the acyl chain of phosphatidylcholine. The quenching was progressively weaker suggesting an interfacial location of the Trp. In parallel, HlyA was subjected to oxidation with N-bromosuccinimide to study the role of Trp residues exposed to aqueous media in its structure-function relationship. In the folded toxin molecule, a single residue was susceptible to oxidation with NBS, whereas incubation with LUV of the toxin prior modification prevents its oxidation, suggesting that Trp residue(s) are directly involved in toxin binding and insertion. Finally, the modification of residues exposed to solvent resulted in a complete impairment of the lytic activity. It was concluded that the modification-sensitive Trp residues are essential for the structure and function of native HlyA. These results are consistent with the model proposed by Soloaga et al. (Mol. Microbiol. 31 (1999) 1013-1024) according to which HlyA is bound to a single monolayer through a number of amphipathic instead of inserted transmembrane helices. PMID- 10704918 TI - Fluorescently labeled neomycin as a probe of phosphatidylinositol-4, 5 bisphosphate in membranes. AB - Phosphatidylinositol-4,5-bisphosphate (PI(4,5)P(2)), a minor component of the plasma membrane, is important in signal transduction, exocytosis, and ion channel activation. Thus fluorescent probes suitable for monitoring the PI(4,5)P(2) distribution in living cells are valuable tools for cell biologists. We report here three experiments that show neomycin labeled with either fluorescein or coumarin can be used to detect PI(4,5)P(2) in model phospholipid membranes. First, addition of physiological concentrations of PI(4,5)P(2) (2%) to lipid vesicles formed from mixtures of phosphatidylcholine (PC) and phosphatidylserine (PS) enhances the binding of labeled neomycin significantly (40-fold for 5:1 PC/PS vesicles). Second, physiological concentrations of inositol-1,4,5 trisphosphate (10 microM I(1,4,5)P(3)) cause little translocation of neomycin from PC/PS/PI(4,5)P(2) membranes to the aqueous phase, whereas the same concentrations of I(1,4,5)P(3) cause significant translocation of the green fluorescent protein/phospholipase C-delta pleckstrin homology (GFP-PH) constructs from membranes (Hirose et al., Science, 284 (1999) 1527). Third, fluorescence microscopy observations confirm that one can distinguish between PC/PS vesicles containing either 0 or 2% PI(4, 5)P(2) by exposing a mixture of the vesicles to labeled neomycin. Thus fluorescently labeled neomycin could complement GFP-PH constructs to investigate the location of PI(4,5)P(2) in cell membranes. PMID- 10704919 TI - Hemolysis of human erythrocytes induced by tamoxifen is related to disruption of membrane structure. AB - Tamoxifen (TAM), the antiestrogenic drug most widely prescribed in the chemotherapy of breast cancer, induces changes in normal discoid shape of erythrocytes and hemolytic anemia. This work evaluates the effects of TAM on isolated human erythrocytes, attempting to identify the underlying mechanisms on TAM-induced hemolytic anemia and the involvement of biomembranes in its cytostatic action mechanisms. TAM induces hemolysis of erythrocytes as a function of concentration. The extension of hemolysis is variable with erythrocyte samples, but 12.5 microM TAM induces total hemolysis of all tested suspensions. Despite inducing extensive erythrocyte lysis, TAM does not shift the osmotic fragility curves of erythrocytes. The hemolytic effect of TAM is prevented by low concentrations of alpha-tocopherol (alpha-T) and alpha-tocopherol acetate (alpha TAc) (inactivated functional hydroxyl) indicating that TAM-induced hemolysis is not related to oxidative membrane damage. This was further evidenced by absence of oxygen consumption and hemoglobin oxidation both determined in parallel with TAM-induced hemolysis. Furthermore, it was observed that TAM inhibits the peroxidation of human erythrocytes induced by AAPH, thus ruling out TAM-induced cell oxidative stress. Hemolysis caused by TAM was not preceded by the leakage of K(+) from the cells, also excluding a colloid-osmotic type mechanism of hemolysis, according to the effects on osmotic fragility curves. However, TAM induces release of peripheral proteins of membrane-cytoskeleton and cytosol proteins essentially bound to band 3. Either alpha-T or alpha-TAc increases membrane packing and prevents TAM partition into model membranes. These effects suggest that the protection from hemolysis by tocopherols is related to a decreased TAM incorporation in condensed membranes and the structural damage of the erythrocyte membrane is consequently avoided. Therefore, TAM-induced hemolysis results from a structural perturbation of red cell membrane, leading to changes in the framework of the erythrocyte membrane and its cytoskeleton caused by its high partition in the membrane. These defects explain the abnormal erythrocyte shape and decreased mechanical stability promoted by TAM, resulting in hemolytic anemia. Additionally, since membrane leakage is a final stage of cytotoxicity, the disruption of the structural characteristics of biomembranes by TAM may contribute to the multiple mechanisms of its anticancer action. PMID- 10704920 TI - Dual effect of NO on K(+)(ATP) current of mouse pancreatic B-cells: stimulation by deenergizing mitochondria and inhibition by direct interaction with the channel. AB - Nitric oxide (NO) is assumed to contribute to the impairment of B-cell function in type 1 diabetes mellitus (IDDM). In the present paper we show that in mouse B cells with intact metabolism authentic NO (20 microM) led to a biphasic effect on the K(+)(ATP) current, namely a transient increase and a consecutive almost complete inhibition. This resembles closely the effect that we have observed previously with the NO donor S-nitrosocysteine (SNOC, 1 mM) suggesting that merely NO caused both phases of this effect. We now demonstrate that the rise in the current amplitude was accompanied by a depolarization of the mitochondrial membrane potential DeltaPsi and a concomitant reduction in the ATP/ADP ratio. Thus, it seems likely that the increase in current amplitude is due to the interference of NO with cell metabolism. The subsequent inhibition of the K(+)(ATP) current is assumed to be caused by a direct effect on the channel since K(+)(ATP) single channel current activity measured in excised patches was strongly reduced by authentic NO and SNOC. Our data reveal new insights into the mechanisms underlying the biphasic action of NO on K(+)(ATP) channels in pancreatic B-cells. PMID- 10704921 TI - A synthetic channel-forming peptide induces Cl(-) secretion: modulation by Ca(2+) dependent K(+) channels. AB - A synthetic Cl(-) channel-forming peptide, C-K4-M2GlyR, applied to the apical membrane of human epithelial cell monolayers induces transepithelial Cl(-) and fluid secretion. The sequence of the core peptide, M2GlyR, corresponds to the second membrane-spanning region of the glycine receptor, a domain thought to line the pore of the ligand-gated Cl(-) channel. Using a pharmacological approach, we show that the flux of Cl(-) through the artificial Cl(-) channel can be regulated by modulating basolateral K(+) efflux through Ca(2+)-dependent K(+) channels. Application of C-K4-M2GlyR to the apical surface of monolayers composed of human colonic cells of the T84 cell line generated a sustained increase in short circuit current (I(SC)) and caused net fluid secretion. The current was inhibited by the application of clotrimazole, a non-specific inhibitor of K(+) channels, and charybdotoxin, a potent inhibitor of Ca(2+)-dependent K(+) channels. Direct activation of these channels with 1-ethyl-2-benzimidazolinone (1-EBIO) greatly amplified the Cl(-) secretory current induced by C-K4-M2GlyR. The effect of the combination of C-K4-M2GlyR and 1-EBIO on I(SC) was significantly greater than the sum of the individual effects of the two compounds and was independent of cAMP. Treatment with 1-EBIO also increased the magnitude of fluid secretion induced by the peptide. The cooperative action of C-K4-M2GlyR and 1-EBIO on I(SC) was attenuated by Cl(-) transport inhibitors, by removing Cl(-) from the bathing solution and by basolateral treatment with K(+) channel blockers. These results indicate that apical membrane insertion of Cl(-) channel-forming peptides such as C-K4-M2GlyR and direct activation of basolateral K(+) channels with benzimidazolones may coordinate the apical Cl(-) conductance and the basolateral K(+) conductance, thereby providing a pharmacological approach to modulating Cl( ) and fluid secretion by human epithelia deficient in cystic fibrosis transmembrane conductance regulator Cl(-) channels. PMID- 10704922 TI - Establishment of plasma membrane polarity in mammary epithelial cells correlates with changes in prolactin trafficking and in annexin VI recruitment to membranes. AB - Mammary epithelial cells (MEC) of lactating animals ferry large amounts of milk constituents in vesicular structures which have mostly been characterized by morphological approaches (Ollivier-Bousquet, 1998). Recently, we have shown that under conditions of lipid deprivation, perturbed prolactin traffic paralleled changes in the membrane phospholipid composition and in the cytosol versus membrane distribution of annexin VI (Ollivier-Bousquet et al., 1997). To obtain additional information on the membrane events involved in the vesicular transport of the hormone to the apical pole of the cell, we conducted a biochemical study on prolactin-containing vesicles in MEC at two different stages of differentiation. We first showed that MEC of pregnant and lactating rabbits exhibited membrane characteristics of non-polarized and polarized cells respectively, using annexin IV and the alpha-6 subunit of integrin as membrane markers. Incubation of both cell types with biotinylated prolactin for 1 h at 15 degrees C, followed by a 10-min chase at 37 degrees C revealed that prolactin transport was activated upon MEC membrane polarization. This was confirmed by subcellular fractionation of prolactin-containing vesicles on discontinuous density gradients. In non-polarized MEC, (125)I-prolactin was mainly recovered in gradient fractions enriched with endocytotic vesicles either after incubation at 15 degrees C or after a 10-min chase at 37 degrees C. In contrast, in polarized MEC, the hormone switched from endocytotic compartments to a fraction enriched in exocytotic clathrin-coated vesicles during the 10-min chase at 37 degrees C. Association of annexin VI to prolactin carriers was next studied in both non polarized and polarized cells. Membrane compartments collected at each gradient interface were solubilized under mild conditions by Triton X-100 (TX100) and the distribution of annexin VI in TX100-insoluble and TX100-soluble fractions was analyzed by Western blotting. Upon MEC polarization, the amount of annexin VI recovered in TX100-insoluble fractions changed. Quite interestingly, it increased in a membrane fraction enriched with endocytotic clathrin-coated vesicles, suggesting that annexin VI may act as a sorting signal in prolactin transport. PMID- 10704923 TI - Systemic administration of cationic phosphonolipids/DNA complexes and the relationship between formulation and lung transfection efficiency. AB - Performances of cationic lipid formulations for intravenous gene delivery to mouse lungs have been previously reported. We report in this study that cationic phosphonolipids, when appropriately formulated, can be good synthetic vectors for gene delivery to lung after intravenous administration. One of our reagents, GLB43, was capable of mediating a 500-fold higher expression in the lungs of mice than could be obtained with free pDNA alone (P=0.018). We demonstrate that the most important parameters for cationic phosphonolipid transfection activity after systemic administration are the chemical structure of the cationic phosphonolipid, the lipid to DNA charge ratio and the inclusion of co-lipid in the formulation. We report using a luciferase reporter gene that transfection activity in vivo 24 h after cationic phosphonolipid systemic administration could not be predicted from in vitro analysis. In contrast to in vitro studies, cationic phosphonolipids including the oleyl acyl chains (GLB43) were more effective than its analogue with the myristyl acyl chains (GLB73). Using pathological analysis of animal livers, we demonstrate that the toxicity level was correlated with the lipoplex formulation and the lipid to DNA ratio. PMID- 10704924 TI - Solution conformations of short-chain phosphatidylcholine. Substrates of the phosphatidylcholine-preferring PLC of Bacillus cereus. AB - The phosphatidylcholine (PC)-preferring phospholipase C (PLC) from Bacillus cereus (PLC(Bc)) hydrolyzes various 1,2-diacyl derivatives of PC at different rates. Substrates with side chains having eight or more carbons are present in micellular form in aqueous media and are processed most rapidly. The catalytic efficiency (k(cat)/K(m)) for the hydrolyses of short-chain PCs at concentrations below their respective critical micelle concentrations also decreases as the side chains become shorter, and this loss of efficiency owes its origin to increases in K(m). In order to ascertain whether the observed increases in K(m) might arise from conformational changes in the glycerol backbone, nuclear magnetic resonance (NMR) experiments were performed in D(2)O to determine the (3)J(HH) and (3)J(CH) coupling constants along the glycerol subunit of 1, 2-dipropanoyl-sn-glycero-3 phosphocholine (K(m)=61 mM), 1, 2-dibutanoyl-sn-glycero-3-phosphocholine (K(m)=21.2 mM) and 1, 2-dihexanoyl-sn-glycero-3-phosphocholine (K(m)=2.4 mM). Using these coupling constants, the fractional populations for each rotamer about the backbone of each of substrate were calculated. Two rotamers, which were approximately equally populated, about the sn-1-sn-2 bond of each substrate were significantly preferred, and in these conformers, the oxygens on the sn-1 and sn 2 carbons of the backbone were synclinal to optimize intramolecular hydrophobic interactions between the acyl side chains. There was greater flexibility about the sn-2-sn-3 bond, and each of the three possible staggered conformations was significantly populated, although there was a slight preference for the rotamer in which the oxygen bearing the phosphate head group was synclinal to the oxygen at the sn-2 carbon and to the sn-1 carbon; in this orientation, the head group is folded back relative to the side chains. These studies demonstrate that there is no significant change in the conformation about the glycerol backbone as a function of side chain length in short-chain phospholipids. Thus, prior organization of the substrate seems an unlikely determinant of the catalytic efficiency of PLC(Bc), and other factors such as hydrophobic interactions or differential solvation/desolvation effects associated with the complexation of the substrate with PLC(Bc) may be involved. PMID- 10704925 TI - Submolecular organization of DMPA in surface monolayers: beyond the two-layer model. AB - A new approach to the data refinement of X-ray reflection measurements from lipid surface monolayers, applied to DMPA on pure water, reveals the structural organization of the lipid in unprecedented detail and provides new insights into headgroup conformation and hydration as a function of lateral pressure. While conventional box models are incapable of modeling the experimental data at high momentum transfer satisfactorily, a quasimolecular composition-space refinement approach using distribution functions to map the spatial organization of submolecular headgroup fragments yields a much better description and overcomes inherent difficulties of box models. Upon going from the fluid liquid-expanded (LE) phase to the hexatic liquid-condensed (LC) phase, the orientation of the headgroup is tightly coupled to the ordering of the acyl chains. Headgroups tilt toward the surface normal to accommodate for the large reduction in available area per lipid molecule. The spread of the headgroup fragment distribution is considerably larger than the global interface roughness and increases slightly with compression. In distinction to earlier work on DMPE using the two-box approach, we find that the phosphate hydration stays essentially constant across the whole isotherm. The discrepancy between the results observed with the different models is attributed to intrinsic deficiencies of the box model. PMID- 10704926 TI - The N-terminal region of the plasma membrane Ca(2+) pump does not separate from the main catalytic fragments after proteolysis. AB - The purified plasma membrane Ca(2+) pump (PMCA) was digested with trypsin, and the proteolytic products were identified by immunoblotting with monoclonal antibodies JA9 or 5F10 directed against the extreme N-terminal segment and the central portion of the molecule, respectively. After a short treatment with low concentrations of the protease, JA9 reacted predominantly with a peptide of 35 kDa whereas 5F10 detected a peptide of 90 kDa. The trypsin cut leading to the production of these fragments had no effect on the maximal activity of the enzyme. At higher concentrations of trypsin, JA9 detected a main fragment of 33 kDa and smaller fragments of 19 and 15 kDa. The persistence of fragments reacting with JA9 indicates that the N-terminal region containing its epitope (residues 51 75) was not easily accessible to the protease in the native PMCA. However, the reactivity with JA9 was rapidly lost during proteolysis of the denatured protein. The passage of the mixture of PMCA fragments through a calmodulin-Sepharose column resulted in the retention of the N-terminal 35 kDa fragment together with that of 90 kDa, despite the fact that only the latter binds calmodulin. The ethylenediaminetetraacetic acid (EDTA) eluate, which contained about equal amounts of both fragments, had a Ca(2+) ATPase activity similar to that of the intact enzyme. The tight association between the two peptides was evidenced by the fact that concentrations of polyoxyethylene 10 lauryl ether (C(12)E(10)), sodium dodecyl sulfate (SDS) high enough for inactivating the enzyme and dissociate the pump from calmodulin were unable of breaking the interaction between the 35 and 90 kDa fragments. Altogether, these results show that after digestion with trypsin, the N-terminal portion of the PMCA, including the extreme N-terminal segment, remains part of a fully functional catalytic complex. PMID- 10704927 TI - Squalamine is not a proton ionophore. AB - Squalamine, an aminosterol antibiotic isolated from the dogfish shark, creates relatively large defects in phospholipid bilayers, allowing the unrestricted translocation of small molecules across these compromised membranes (B.S. Selinsky, Z. Zhou, K.G. Fotjik, S. R. Jones, N.R. Dollahon, A.E. Shinnar, Biochim. Biophys. Acta 1370 (1998) 218-234). However, an aminosterol structurally similar to squalamine was found to act as a proton ionophore in anionic phospholipid vesicles. In contrast with squalamine, gross membrane disruption was not observed with this synthetic analog (G. Deng, T. Dewa, S.L. Regen, J. Am. Chem. Soc. 118 (1996) 8975-8976). In this report, the ionophoric activity of squalamine was tested in anionic and zwitterionic phospholipid vesicles. No ionophoric activity was observed for squalamine in vesicles comprised of phosphatidylglycerol (PG), phosphatidylcholine (PC), or a mixture of the two lipids. Experiments using radiolabeled squalamine indicated that all of the squalamine added to PG vesicles remained with the vesicles, while approximately one-half of the squalamine added to PC vesicles was incorporated. We have synthesized the aminosterol analog of squalamine possessing ionophoric activity, and its ionophoric activity in PG vesicles was confirmed. The synthetic compound possessed no measurable lytic activity when added to preformed phospholipid vesicles. As both compounds possess significant antimicrobial activity, these results suggest that either multiple mechanisms for the antimicrobial activity of aminosterols exist, depending upon the aminosterol structure, or possibly an unrelated common mechanism for antimicrobial activity remains to be discovered. PMID- 10704928 TI - Anion modulation of calcium current voltage dependence and amplitude in salamander rods. AB - Hofmeister anions were used to investigate the ability of Cl(-) replacement to produce inhibition and a hyperpolarizing activation shift in L-type Ca(2+) currents (I(Ca)) of rod photoreceptors. Inhibition of I(Ca) largely followed the Hofmeister sequence: Cl(-)=Br(-)4) linked. Treatment with proteases suggests that the protein is efficiently shielded from digestion. All the extracts and residues contain major amounts of (glyco)proteins and/or proteins, in agreement with a previous suggestion that they are of major importance in the structure of the cell wall. PMID- 10704983 TI - Enthalpy and enzyme activity of modified histidine residues of adenosine deaminase and diethyl pyrocarbonate complexes. AB - Kinetic and thermodynamic studies have been made on the effect of diethyl pyrocarbonate as a histidine modifier on the active site of adenosine deaminase in 50 mM sodium phosphate buffer pH 6.8, at 27 degrees C using UV spectrophotometry and isothermal titration calorimetry (ITC). Inactivation of adenosine deaminase by diethyl pyrocarbonate is correlated with modification of histidyl residues. The number of modified histidine residues complexed to active site of adenosine deaminase are equivalent to 4. The number and energy of histidine binding sets are determined by enthalpy curve, which represents triple stages. These stages are composed of 3,1 and 1 sites of histidyl modified residues at diethyl pyrocarbonate concentrations, 0.63, 1.8, 3.3 mM. The heat contents corresponding to the first, second and third sets are found to be 18000, 22000 and 21900 kJ mol(-1) respectively. PMID- 10704984 TI - Magnetic nanostructured composites using alginates of different M/G ratios as polymeric matrix. AB - Alginate extracted from Sargassum fluitans and Macrocystis pyrifera with different molecular weights and mannuronic/guluronic ratios, M/G, were used as gel matrixes in order to obtain magnetic nanostructured composites. Magnetic nanocrystalline particles of iron oxides were formed inside the alginate matrix by in situ alkaline oxidation of ferrous ions. The magnetic materials obtained were subjected to several oxidative cycles and the increment in iron content was determined after each cycle. X-ray diffraction, magnetometry and Mossbauer spectroscopy were used to examine the materials. The high magnetic response, the absence of hysteresis, and the centered paramagnetic doublet in the Mossbauer spectra indicate the presence of nanocrystalline particles with a superparamagnetic behavior. X-ray diffractograms show peaks that correspond to maghemite. After the first cycle, Sargassum had four times the magnetic response of Macrocystis, which had more than twice the M/G ratio. PMID- 10704985 TI - Phase equilibria and mechanical properties of gel-like water-gelatin-locust bean gum systems. AB - The establishment of phase equilibrium in aqueous gelatin-locust bean gum (LBG) systems in the process of cooling from 313 to 291 K in specific conditions, passes ahead of the gelation process(.) This allows the suggestion that macrostructure and mechanical properties of the system can be predicted on the basis of knowledge of its phase diagram, obtained for the liquid gelatin-LBG systems comprising gelatin molecular aggregates. Textural and rheological analysis of gel-like gelatin-LBG systems demonstrate the effect of two factors determining their mechanical properties and acting opposite each other when the concentration of LBG in the system increases: concentration of gelatin by LBG in the process of phase separation, and decrease in the density of the gel network. PMID- 10704986 TI - A novel heparan sulphate with high degree of N-sulphation and high heparin cofactor-II activity from the brine shrimp Artemia franciscana. AB - With the aid of heparinase and heparitinases from Flavobacterium heparinum and 13C and IH NMR spectroscopy it was shown that the heparan sulphate isolated from the brine shrimp Artemia franciscana exhibits structural features intermediate between those of mammalian heparins and heparan sulphates. These include an unusually high degree of N-sulphation (with corresponding very low degree of N acetylation), a relatively high content of iduronic acid residues (both unsulphated and 2-O-sulphated) and a relatively low degree of 6-O-sulphation of the glucosamine residues. The major sequences (glucuronic acid-->N-sulphated glucosamine and glucuronic acid-->N, 6-disulphated glucosamine) are most probably arranged in blocks. Although exhibiting negligible anticlotting activity in the APTT and anti-factor Xa assays the A. franciscana heparan sulphate has a high heparin cofactor-II activity (about 1/3 that of heparin). PMID- 10704987 TI - Synthesis and structural characterization of poly(LGGVG), an elastin-like polypeptide. AB - Poly(LGGVG) a potential elastin-like biomaterial has been synthesized and studied both in solution (by circular dicroism and nuclear magnetic resonance) and in the aggregated state (by transmission electron microscopy). For sake of comparison, also the conformation of the protected (Boc-LGGVG-OEt) and free (H(2)(+)-LGGVG OH) 'monomers' has been investigated. While in the latter ones the presence has been evidenced of more or less stable type II beta-turns, the polymer showed a conformational ensemble, possibly comprising type II beta-turns, type I beta turns and open (unordered) structures. At supramolecular level, twisted-rope aggregates were observed by transmission electron microscopy for the polymer. Thus, the title compound has shown to possess, at both molecular and supramolecular level, physico-chemical properties very similar to those of elastin, so to give some confidence that it could really constitute the precursor of an artificial substitute of elastin itself. PMID- 10704988 TI - A new bacterial polysaccharide (YAS34). I. Characterization of the conformations and conformational transition. AB - This paper concerns the study of the conformational transition of a new exopolysaccharide (YAS34) using experimental techniques such as optical rotation, conductimetric and microcalorimetric measurements as a function of temperature. The behaviors of this polysaccharide in the acid or sodium salt form are compared; a deacetylated sample is also prepared to demonstrate the role of substituents. For the native structure (never heated), a conformational transition is observed but the deacetylated polysaccharide exhibits no ordered conformation. Multidetection size exclusion chromatography (SEC) analyses and conductimetric experiments allowed to determine the nature of each conformation and the molecular dimensions. From these results, it is suggested that the native conformation is a double helix which by heating over T(m) (temperature corresponding to half conformational transition) dissociates into disordered single chains. In the acid and sodium salt forms, by cooling below T(m), an ordered conformation is restored. This conformation seems to be an intramolecular double helix 'hairpin-like turn' (called renatured conformation). Nevertheless an irreversible denaturation is obtained progressively in the sodium salt form when the time of heating over T(m) increases. The conformation of the deacetylated polysaccharide corresponds to that of a single flexible chain (disordered conformation). The conformational transition for the native conformation was studied also in relation to the polyelectrolytic character of the polysaccharide: stability as a function of salt nature and salt and polymer concentrations was investigated for the polymer initially in the sodium and acid forms. PMID- 10704989 TI - A new bacterial exopolysaccharide (YAS34). II. Influence of thermal treatments on the conformation and structure. Relation with gelation ability. AB - This paper describes an original mechanism of evolution of a polysaccharide system occurring during thermal treatments. Under its native conformation, the YAS34 polymer presents a solution character in the dilute and semi-dilute regimes. However, the zero shear rate viscosity indicates existence of interchain interactions which disappear on the deacetylated polymer. Thermal treatments over the temperature of conformational change produce a progressive and irreversible evolution of the physical properties when the polymer is under its sodium salt form. This evolution was related to a modification of the arrangement of acetyl substituents. The heated polysaccharide gives thermoreversible gel which is very elastic. A gelation mechanism is proposed based on formation of helical segments connecting the network. PMID- 10704990 TI - On the temperature-induced coil to globule transition of poly-N isopropylacrylamide in dilute aqueous solutions. AB - Poly-N-isopropylacrylamide (PNIPAM) is a chemical isomer of poly-leucine, having the polar peptide group in the side-chain rather than in the backbone. It has been demonstrated experimentally that PNIPAM dissolved in aqueous solution undergoes a collapse transition from coil to globule on increasing temperature above the θ-point. By a careful reviewing of existing experimental data, we emphasize that such coil to globule collapse has to be considered an intramolecular first-order transition, analogous to the cold renaturation of small globular proteins. The main theoretical approaches to the coil to globule collapse in homopolymers are discussed briefly, and a critical comparison between the existing models is performed. We point out that, as a general result, the coil to globule collapse is expected to be a first-order transition for rigid and semi-rigid macromolecules. Finally, taking advantage of the analogy between the coil to globule collapse of PNIPAM and the cold renaturation of small globular proteins, we try to clarify some important and intriguing aspects of protein thermodynamics. This leads to the conclusion that the amphiphilic nature of polypeptide chain plays the fundamental role for the existence of two temperature induced conformational transitions. PMID- 10704991 TI - Structure of insect chitin isolated from beetle larva cuticle and silkworm (Bombyx mori) pupa exuvia. AB - Chitin samples in a alpha-form structure were isolated from beetle larva cuticle and silkworm (Bombyx mori) pupa exuvia by treatment with 1 N HCl and 1 N NaOH. Chitosan was prepared by treating them in 40% NaOH containing NaBH(4). Chitin and chitosan were analyzed by X-ray, [13C]CP/MAS NMR, [13C]FT-NMR, and scanning electron microscopy (SEM) methods. Insect chitin degraded more readily than shrimp chitin when treated with 6 N HCl and the enzyme-chitinase. After treatment with 2 N HCl at 100 degrees C, the insect chitin crystallinity increased. N deacetylation of insect chitin was easier than that of crustaceous chitin, and about 94% of the N-acetyl groups were removed in one treatment with 40% NaOH for 4 h at 110 degrees C. After treatment with 2 N HCl, 55% of the N-acetyl groups of silkworm chitin were removed under the same conditions. Beetle chitin showed a higher affinity for chitinase than shrimp chitin. PMID- 10704992 TI - Development of tailor-made glycidyl methacrylate-divinyl benzene copolymer for immobilization of D-amino acid oxidase from Aspergillus species strain 020 and its application in the bioconversion of cephalosporin C. AB - A tailor-made glycidyl methacrylate-divinyl benzene (GMA-DVB) copolymer PC-3 was evolved by studying the effect of synthesis variables on binding and expression of D-amino acid oxidase (DAAO) from Aspergillus species strain 020. Almost quantitative binding (100%) and a high yield of immobilization per unit of enzyme loaded was achieved. Optimum pH, optimum temperature and K(m)95% was achieved by using 3% (w/v) solution of ceph C, 48 U of DAAO per g of ceph C, keeping dissolved oxygen level above 50%, maintaining the pH between 7.6 and 7.8 and temperature at 24 degrees C. The immobilized DAAO was used for 60 cycles in a stirred tank reactor. PMID- 10704993 TI - Hydroxyl radical generation by an extracellular low-molecular-weight substance and phenol oxidase activity during wood degradation by the white-rot basidiomycete Trametes versicolor. AB - One-electron oxidation activity, as measured by ethylene generation from 2-keto-4 thiomethylbutyric acid, phenol oxidase activity, and the generation of hydroxyl radical were examined in cultures of the lignin-degrading white-rot basidiomycete fungus, Trametes (Coriolus) versicolor. The activity levels of specific lignin degrading enzymes and cellulases, as well as the rate of wood degradation, also were examined. The fungus secreted a low-molecular-weight substance (M(r) 1000 5000) that catalyzed a redox reaction between molecular oxygen and an electron donor, to produce the hydroxyl radical via hydrogen peroxide. During wood decay, T. versicolor also produced significant amounts of laccase and lignin peroxidase, carboxymethyl cellulase, and Avicelase. The roles of the hydroxyl radical, phenol oxidases, and cellulases in wood degradation by white-rot fungi are discussed. That the hydroxyl radical produced by the low-molecular-weight substance secreted by T. versicolor results in new phenolic substructures on the lignin polymer, making it susceptible to attack by laccase or manganese peroxidase is suggested. PMID- 10704994 TI - Development of a cold resistant mutant of plant growth promoting Pseudomonas fluorescens and its functional characterization. AB - A cold resistant mutant of plant growth promoting Pseudomonas fluorescens GRS(1), was isolated following N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) treatment. The mutant was able to grow and promote root and shoot elongation of wheat both at 25 and 10 degrees C, a temperature at which the wild type was unable to proliferate and function. In an effort to determine the mechanistic basis for this behavior, it was observed that following growth at 10 degrees C, the mutant synthesized some new proteins. SDS-polyacrylamide gel electrophoresis of the protein synthesized by the mutant revealed the presence of one major protein with a molecular mass of 13.5 kDa. However, at this point the function of this protein in not known. PMID- 10704995 TI - Nervous system proteoglycans as modulators of neurite outgrowth. AB - The proteoglycans are multifunctional macromolecules composed of a core polypeptide and a variable number of glycosaminoglycan chains. The structural diversity and complexities of proteoglycan expression in the developing and adult Nervous System underlies the variety of biological functions that these molecules fulfill. Thus, in the Nervous System, proteoglycans regulate the structural organisation of the extracellular matrix, modulate growth factor activities and cellular adhesive and motility events, such as cell migration and axon outgrowth. This review summarises the evidences indicating that proteoglycans have an important role as modulators of neurite outgrowth and neuronal polarity. Special emphasis will be placed on those studies that have shown that proteoglycans of certain subtypes inhibit neurite extension either during the development and/or the regeneration of the vertebrate Central Nervous System. PMID- 10704996 TI - Phosphorylation of microtubule-associated protein 2 (MAP2) and its relevance for the regulation of the neuronal cytoskeleton function. AB - Neurons, the basic information processing units of the nervous system, are characterized by a complex polar morphology which is essential for their function. To attain their precise morphology, neurons extend cytoplasmatic processes (axons and dendrites) and establish synaptic connections in a highly regulated way. Additionally, neurons are also subjected to small plastic changes at the adult stage which serve to regulate synaptic transmission. Every step of neuronal development is genetically controlled by endogenous determinants, as well as by environmental signals including intercellular contacts, extracellular matrix and diffusible signals. Cytoskeletal components are among the main protein targets modified in response to most of those extracellular signals which ultimately determine neuronal morphology. One of the major mechanisms controlling the neuronal cytoskeleton is the modification of the phosphorylation state of cytoskeletal proteins via changes in the relative activities of protein kinases and phosphatases within neurons. In particular, the microtubule-associated protein 2 (MAP2) family of proteins is an abundant group of cytoskeletal components which are predominantly expressed in neurons and serve as substrates for most of protein kinases and phosphatases present in neurons. MAP2 phosphorylation seems to control its association with the cytoskeleton and it is developmentally regulated. Moreover, MAP2 may perform many functions including the nucleation and stabilization of microtubules (and maybe microfilaments), the regulation of organelle transport within axons and dendrites, as well as the anchorage of regulatory proteins such as protein kinases which may be important for signal transduction. These putative functions of MAP2 have also been proposed to play important roles in the outgrowth of neuronal processes, synaptic plasticity and neuronal cell death. Thus, MAP2 constitutes an interesting case to understand the regulation of neuronal function by the alteration of the phosphorylation state of cytoskeletal proteins in response to different extracellular signals. Here we will review the current knowledge about the regulation of MAP2 function through phosphorylation/dephosphorylation and its relevance in the broader context of neuronal functions. PMID- 10704997 TI - Wind-up of spinal cord neurones and pain sensation: much ado about something? AB - Wind-up is a frequency-dependent increase in the excitability of spinal cord neurones, evoked by electrical stimulation of afferent C-fibres. Although it has been studied over the past thirty years, there are still uncertainties about its physiological meaning. Glutamate (NMDA) and tachykinin NK1 receptors are required to generate wind-up and therefore a positive modulation between these two receptor types has been suggested by some authors. However, most drugs capable of reducing the excitability of spinal cord neurones, including opioids and NSAIDs, can also reduce or even abolish wind-up. Thus, other theories involving synaptic efficacy, potassium channels, calcium channels, etc. have also been proposed for the generation of this phenomenon. Whatever the mechanisms involved in its generation, wind-up has been interpreted as a system for the amplification in the spinal cord of the nociceptive message that arrives from peripheral nociceptors connected to C-fibres. This probably reflects the physiological system activated in the spinal cord after an intense or persistent barrage of afferent nociceptive impulses. On the other hand, wind-up, central sensitisation and hyperalgesia are not the same phenomena, although they may share common properties. Wind-up can be an important tool to study the processing of nociceptive information in the spinal cord, and the central effects of drugs that modulate the nociceptive system. This paper reviews the physiological and pharmacological data on wind-up of spinal cord neurones, and the perceptual correlates of wind-up in human subjects, in the context of its possible relation to the triggering of hyperalgesic states, and also the multiple factors which contribute to the generation of wind-up. PMID- 10704998 TI - The p75 neurotrophin receptor and neuronal apoptosis. AB - Although evidence continues to accumulate for the apoptosis-inducing role of the p75 neurotrophin receptor, several outstanding questions remain. One of these concerns the signal transduction pathway of p75, which continues to be elusive. The evidence for the roles of ceramide, c-jun kinase and NF-kappaB is discussed: none of these are able to account satisfactorily for p75 death signalling. Negative modulation of Trk signalling by p75 could account for part of the pro apoptotic effect, but is unlikely to be a major component. Although recent evidence indicates that the juxtamembrane region is critical for causing cell death, p75 has a well-conserved death domain. This may be important for functions other than killing. In glial cells and some neurons that express p75 but not TrkA, p75 causes cell death in response to nerve growth factor (NGF) binding. In sensory neurons and PC12 cells, p75 appears to signal constitutively. In cholinergic forebrain neurons, p75 expression leads to atrophy and downregulation of cholinergic markers, rather than cell death. The major challenges in p75 research are to define its signalling pathways, and particularly the intracellular proteins with which it interacts. Another major challenge is to develop a model that reconciles the different facets of p75, such as its ability in some situations to assist TrkA to rescue NGF-dependent neurons, but to stimulate apoptosis in others. PMID- 10704999 TI - Internal mammary lymph nodes: to treat or not to treat. PMID- 10705000 TI - Should internal mammary lymph nodes in breast cancer be a target for the radiation oncologist? AB - PURPOSE: The elective treatment of internal mammary lymph nodes (++IMNs) in breast cancer is controversial. Previous randomized trials have not shown a benefit to the extended radical mastectomy or elective IMN irradiation overall, but a survival benefit has been suggested by some for subgroups of patients with medial tumors and positive axillary lymph nodes. The advent of effective systemic chemotherapy and potential for serious cardiac morbidity have also been factors leading to the decreased use of IMN irradiation during the past decade. The recent publishing of positive trials testing postmastectomy radiation that had included regional IMN irradiation has renewed interest in their elective treatment. The purpose of this study is to critically review historical and new data regarding IMNs in breast cancer. METHODS AND MATERIALS: The historical incidence of occult IMN positivity in operable breast cancer is reviewed, and the new information provided by sentinel lymph node studies also discussed. The results of published randomized prospective trials testing the value of elective IMN dissection and/or radiation are analyzed. The data regarding patterns of failure following elective IMN treatment is studied to determine its impact on local-regional control, distant metastases, and survival. A conclusion is drawn regarding the merits of elective IMN treatment based on this review of the literature. RESULTS: Although controversial, the existing data from prospective, randomized trials of IMN treatment do not seem to support their elective dissection or irradiation. While it has not been shown to contribute to a survival benefit, the IMN irradiation increases the risk of cardiac toxicity that has effaced the value of radiation of the chest wall in reducing breast cancer deaths in previous randomized studies and meta-analyses. Sentinel lymph node mapping provides an opportunity to further evaluate the IMN chain in early stage breast cancer. Biopsy of "hot" nodes may be considered in the future to select patients who are most likely to benefit from additional regional therapy to these nodes. CONCLUSIONS: Irradiation of the IMN chain in conjunction with the chest wall and supraclavicular region should be considered only for those with pathologically proven IMNs with the goal of improving tumor regional control. PMID- 10705001 TI - The modified tangential irradiation technique for breast cancer: how to cover the entire axillary region. AB - PURPOSE: The two-portal tangential irradiation technique has usually been applied to breast cancer patients after breast-conserving surgery (1, 2) and is expected to irradiate the axillary lymph node region to some extent (3). We investigated the range of the axillary region covered by this technique and tried to devise an optimal irradiation technique (modified tangential irradiation) that would cover the axillary lymph node region properly. METHODS AND MATERIALS: We checked the status of the surgical clips left at axillary lymph node sites by reviewing the simulator films and planning CT scans of 63 patients who underwent axillary dissection of level I, I-II, or I-III lymph nodes. Then we created the modified tangential irradiation technique and applied this technique to 16 patients and checked the irradiation volume by CT scans. RESULTS: We found that all of the surgical clips on lateral-view simulator films were on the ventral side of the dorsal edge line of the humeral head. All but one clip were on the caudal side of the caudal edge line of the humeral head. Accordingly, it is possible to irradiate almost all axillary lymph node regions by setting the dorsal edge of the irradiation field on lateral-view simulator films at the dorsal edge of the humeral head and the cranial edge at the caudal edge of the humeral head. CONCLUSIONS: All breast tissue and the entire axillary lymph node region can be covered by the modified tangential irradiation technique without increasing the lung volume irradiated. PMID- 10705002 TI - Dose selection for prostate cancer patients based on dose comparison and dose response studies. AB - PURPOSE: To better define the appropriate dose for individual prostate cancer patients treated with three-dimensional conformal radiation therapy (3D CRT). METHODS AND MATERIALS: Six hundred eighteen patients treated with 3D CRT between 4/89 and 4/97 with a median follow-up of 53 months are the subject of this study. The bNED outcomes were assessed by the American Society for Therapeutic Radiology and Oncology (ASTRO) definition. The patients were grouped into three groups by prostate-specific antigen (PSA) level (<10 ng/ml, 10-19.9 ng/ml, and 20+ ng/ml) and further subgrouped into six subgroups by favorable (T1, 2A and Gleason score < or =6 and no perineural invasion) and unfavorable characteristics (one or more of T2B, T3, Gleason 7-10, perineural invasion). Dose comparisons for bNED studies were made for each of the six subgroups by dividing patients at 76 Gy for all subgroups except the favorable <10 ng/ml subgroup, which was divided at 72.5 Gy. Five-year bNED rates were compared for the median dose of each dose comparison subgroup. Dose response functions were plotted based on 5-year bNED rates for the six patient groupings, with the data from each of the six subgroups divided into three dose groups. The 5-year bNED rate was also estimated using the dose response function and compares 73 Gy with 78 Gy. RESULTS: Dose comparisons show a significant difference in 5-year bNED rates for three of the six subgroups but not for the favorable <10 ng/ml, the favorable 10-19.9 ng/ml, or the unfavorable > or =20 ng/ml subgroups. The significant differences ranged from 22% to 40% improvement in 5-year bNED with higher dose. Dose response functions show significant differences in 5-year bNED rates comparing 73 Gy and 78 Gy for four of the six subgroups. Again, no difference was observed for the favorable <10 ng/ml group or the unfavorable > or =20 ng/ml group. The significant differences observed in 5-year bNED ranged from 15% to 43%. CONCLUSIONS: Dose response varies by patient subgroup, and appropriate dose can be estimated for up to six subdivisions of prostate cancer patients. The appropriate use of high dose with 3D CRT results in 5-year cure rates that equal or exceed other treatments. The national practice must be upgraded to allow the safe administration of 75-80 Gy with 3D CRT. PMID- 10705003 TI - Quality of life in T1-3N0 prostate cancer patients treated with radiation therapy with minimum 10-year follow-up. AB - PURPOSE: To describe patient-reported quality of life using a validated survey in a cohort of patients who are long-term survivors of definitive radiotherapy for T1-3N0 prostate cancer. METHODS AND MATERIALS: Survivors of a previously reported cohort of prostate cancer patients treated with staging pelvic lymphadenectomy and definitive radiotherapy between November 1974 and August 1988 were queried using a questionnaire incorporating the RAND 36-Item Health Survey and the University of California, Los Angeles Prostate Cancer Index. Responses were reviewed and analyzed. Of the 146 N0 patients, 88 have survived for 10 years postdiagnosis. Fifty-six (64%) of these patients were still alive with valid addresses and were mailed copies of the questionnaires, of which 46 (82%) responded. Median potential follow-up from date of diagnosis was 13.9 years, with a median age of responders of 80 years. RESULTS: The mean sexual function score was 15.4, with a bother score of 42. The mean urinary function score was 65, with a bother score of 61. The mean bowel function score was 72.6, with a bother score of 64.8. The amount of patient bother reported in the sexual category is similar to that previously reported for cohorts of prostate cancer patients undergoing radiotherapy or observation. This is despite the fact that sexual function was similar to that previously reported for patients postprostatectomy. Patient reported function and bother scores in urinary and bowel categories were somewhat more severe than a previously reported radiotherapy cohort with shorter follow up. CONCLUSIONS: With long follow-up, most patients who underwent radiotherapy for prostate cancer in the era described exhibit somewhat worse bladder, bowel, and erectile function than recently published controls without prostate cancer. In this cohort of older men with long follow-up, erectile function is similar to reported prostatectomy series. However, patient bother related to erectile function is similar to that of controls in earlier published radiotherapy series. Worse urinary and bowel function may be due to progressive symptoms with aging and longer follow-up, or to the radiotherapy techniques performed during the era in question. PMID- 10705004 TI - Palladium-103 brachytherapy for prostate carcinoma. AB - PURPOSE: A report of biochemical outcomes for patients treated with palladium-103 (Pd-103) brachytherapy over a fixed time interval. METHODS AND MATERIALS: Two hundred thirty patients with clinical stage T1-T2 prostate cancer were treated with Pd-103 brachytherapy and followed with prostate-specific antigen (PSA) determinations. Kaplan-Meier estimates of biochemical failure on the basis of two consecutive elevations of PSA were utilized. Multivariate risk groups were constructed. Aggregate PSA response by time interval was assessed. RESULTS: The overall biochemical control rate achieved at 9 years was 83.5%. Failures were local 3.0%; distant 6.1%; PSA progression only 4.3%. Significant risk factors contributing to failure were serum PSA greater than 10 ng/ml and Gleason sum of 7 or greater. Five-year biochemical control for those exhibiting neither risk factor was 94%; one risk factor, 82%; both risk factors, 65%. When all 1354 PSA determinations obtained for this cohort were considered, the patients with a proportion of PSAs < or = 0.5 ng/ml continued to increase until at least 48 months post-therapy. These data conformed to a median PSA half-life of 96.2 days. CONCLUSIONS: Prostate brachytherapy with Pd-103 achieves a high rate of biochemical and clinical control in patients with clinically organ-confined disease. PSA response following brachytherapy with low-dose-rate isotopes is protracted. PMID- 10705005 TI - Normal tissue dosimetric comparison between HDR prostate implant boost and conformal external beam radiotherapy boost: potential for dose escalation. AB - PURPOSE: To compare the dose and volume of bladder and rectum treated using high dose-rate (HDR) prostate implant boost versus conformal external beam radiotherapy boost, and to use the dose-volume information to perform a critical volume tolerance (CVT) analysis and then estimate the potential for further dose escalation using HDR brachytherapy boost. METHODS AND MATERIALS: Using CT scan data collected before and after patients underwent HDR prostate implant, a 7 field conformal prostate-only external beam treatment plan and HDR brachytherapy treatment plan were constructed for each patient. Doses to the normal structures were calculated. Dose-volume histograms (DVH) were plotted for comparison of the two techniques. Wilcoxon signed rank test was performed at four dose levels to compare the dose to normal structures between the two treatment techniques. The acute and late effects of HDR brachytherapy were calculated based on the linear quadratic (LQ) model. CVT analyses were performed to calculate the potential dose gain (PDG) using HDR brachytherapy boost. RESULTS: The volume of bladder and rectum receiving high dose was significantly less from implant boost. On the average, 0.19 cc of the bladder received 100% of the brachytherapy prescription dose, compared with 5.1 cc of the bladder receiving 100% of the prescription dose in the 7-field conformal external beam radiotherapy boost. Similarly, 0.25 cc of the rectum received 100% of the dose with the implant boost, as compared to 2.9 cc in the conformal external beam treatment. The implant also delivered higher doses inside the prostate volume. On average, 47% of the prostate received > or =150% of the prescription dose. The CVT analysis revealed a range of PDG using the HDR brachytherapy boost which depended on the following variables: critical volume (CV), critical volume tolerance dose (CVTD), number of HDR fractions (N), and the dose of external beam radiotherapy (XRT) delivered with brachytherapy boost. The PDG varied from -3.45% to 10.53% for tumor with an alpha-beta ratio of 10 and 7.14% to 64.6% for tumor with an alpha-beta ratio of 1.5 based on the parameters used for calculation in this study. CONCLUSIONS: HDR brachytherapy can provide better sparing of rectum and bladder while delivering a higher dose to the prostate. Even with the increased late effects of high dose per fraction, there is still a potential for dose escalation beyond external radiotherapy limits using HDR brachytherapy. PMID- 10705006 TI - Improved long-term survival with combined modality therapy for pediatric nasopharynx cancer. AB - PURPOSE: Nasopharynx cancer is a rare malignancy in childhood. This study aims to determine the role of chemotherapy, the optimal dose of radiation, and the long term outcome for children with locoregional disease. METHODS AND MATERIALS: Thirty-three patients [median age 14 (range: 12-20) years] were treated for Stage I-IVB nasopharynx cancer. Thirteen patients (39%) received radiotherapy alone and 20 patients (61%) had chemotherapy and radiotherapy. The median radiation dose to the primary tumor was 66 Gy (range: 54-72 Gy). The median follow-up time for surviving patients was 8.4 years (range: 0.5-23.6 years). RESUL TS: The actuarial 10-year locoregional relapse-free survival, distant metastases-free survival, and overall survival rates were 77%, 68%, and 58% , respectively. Locoregional control was improved for patients treated with radiation doses > 60 Gy compared to those receiving < or = 60 Gy (93% vs. 60%, p < 0.03). The addition of chemotherapy had no significant effect on locoregional control but did reduce the development of distant metastases (16% vs. 57%, p = 0.01). Combined modality therapy improved 10-year disease-free survival (84% vs. 35%, p < 0.01) and survival (78% vs. 33%, p < 0.05) over radiation alone. The 10-year actuarial rate of severe complications was 24%.60 Gy are used for gross disease. The addition of chemotherapy decreases the risk of distant metastases and increases survival. PMID- 10705007 TI - Total biological effect on late reactive tissues following reirradiation for recurrent nasopharyngeal carcinoma. AB - PURPOSE: To assess the additional damage of normal tissues attributable to reirradiation and the magnitude of partial recovery following the initial course. METHODS AND MATERIALS: Symptomatic late complication rates (excluding xerostomia) in 3635 patients receiving one course (Group 1) and 487 patients receiving two courses of external radiotherapy (Group 2) for nasopharyngeal carcinoma were retrospectively analyzed and compared. RESULTS: Group 2 had significantly lower actuarial complication-free survival rates than Group 1: 48% versus 81% at 5 years. The post-retreatment incidence was significantly affected by biologically effective dose (BED) (assuming an alpha/beta ratio of 3 Gy) of the first course: hazard ratio (HR) = 1.04 per Gy(3) (p = 0.01), but only marginally by that of the second course: HR = 1.01 per Gy(3) (p = 0.06). If the summated BED was taken as the dose unit, it was estimated that a total BED of 143 Gy(3) would induce a 20% incidence at 5 years, while the corresponding dose projected from Group 1 was 111 Gy(3). The gap effect was insignificant in the overall analyses, but a trend of decreasing risk with increasing interval was observed in patients with gap > or = 2 years: HR = 0.86 per year (p = 0.07). CONCLUSION: The major determinant of post retreatment complication is the severity of damage during the initial course. The sum of total doses tolerated is higher than that expected with a single-course treatment, suggesting occurrence of partial recovery (particularly in those reirradiated after an interval of 2 years or more). PMID- 10705008 TI - Investigation of the comparative toxicity of 5-FU bolus versus 5-FU continuous infusion circadian chemotherapy with concurrent radiation therapy in locally advanced rectal cancer. AB - PURPOSE: To compare the relative toxicities of bolus versus infusional 5-FU chemotherapy administrated concurrently during external beam irradiation in patients with locally advanced rectal cancer following surgical extirpation. METHODS: A total of 26 eligible patients were retrospectively identified as having been treated for rectal adenocarcinoma at the Stratton VAMC between 1989 and 1997. A comparative analysis of treatment dose intensities, treatment delays and toxicities in these patients was performed. RESULTS: Significantly less WBC toxicity was observed in the patients receiving infusional 5-FU chemotherapy. The other toxicities, with the exception of skin toxicity, were generally less frequent in the 5-FU infusional group. When the toxicities were corrected for 5 FU dose intensity, to yield toxicity per mg of 5-FU, statistically significant differences were found for hematological toxicity (WBC and platelets), and for gastrointestinal toxicity (frequency and severity of diarrhea and weight loss). The majority of patients receiving infusional 5-FU therapy were treated using a circadian pattern of treatment peaking around the time of the radiation therapy. Patients receiving infusional 5-FU were able to tolerate over twice the dose intensity as those receiving bolus administration. Local recurrence rate in all patients was 3.8% comparing favorably to other reported studies. Distant recurrence frequency was also acceptable at 34.6% for the group. CONCLUSION: Infusional 5-FU chemotherapy compared with bolus therapy during pelvic radiation minimizes toxicity to the patient while maximizing the dose of 5-FU that can be delivered. As infusional 5-FU therapy during radiation has previously been shown to increase disease free duration and survival, infusional 5-FU should be considered as an acceptable standard of care to prevent local recurrence of rectal adenocarcinoma following its resection. Shaping this infusional 5-FU chemotherapy within the day so that most of the daily dose is delivered around the time of the radiation therapy may further modify the toxic therapeutic ratio of combined modality therapy. PMID- 10705009 TI - Preoperative chemoradiation in fixed distal rectal cancer: dose time factors for pathological complete response. AB - PURPOSE: Preoperative chemoradiation is being utilized extensively in the treatment of rectal cancer. However, a variety of dose time factors in both delivery of chemotherapy and irradiation remain to be established. This study was undertaken to examine the impact of dose time factors on pathological complete response (pCR) rates following preoperative chemoradiation for fixed rectal cancer. METHODS AND MATERIALS: Thirty-three patients with fixed rectal cancers were treated with combined 5-fluorouracil (5-FU) chemotherapy and pelvic radiation. Twenty-one patients received bolus 5-FU during the first 3-5 days of radiation and repeated on days 28-33 of their radiation treatment. Twelve patients were treated with continuous infusion (CI) 5-FU, 225 mg/m(2) for the duration of the pelvic radiation. Fifteen patients received a planned total radiation dose of 45 to 50 Gy and 18 patients received a dose of 55 to 60 Gy. Surgical resection was then carried out 6-8 weeks after completion of treatment. RESULTS: Diarrhea was the most frequent acute toxicity. Grade 3 diarrhea was observed in 6 patients requiring treatment interruption and was not related to the chemotherapy regimen. There was no Grade 4 or 5 toxicity. pCR was observed in 2 of 21 (10%) patients treated with bolus 5-FU as compared to 8 of 12 (67%) for patients treated with CI (p = 0.002). pCR were observed in 8 of 18 (44%) patients receiving radiation dose > or = 5500 cGy as compared to 2 of 15 (13%) patients treated to a dose < or = 5000 cGy (p = 0.05). In the high-dose radiation (> or = 5500 cGy) group, a significant difference in pCR rate was observed in patients treated with CI, 8 of 12 (67%) (p = 0.017) as compared with bolus 5-FU (0 of 6). There was no significant difference in operative morbidity or in wound healing between patients treated with bolus 5-FU or CI or within the groups treated with low or high doses of radiation. Three patients have developed local recurrence at 14 and 24 months, two in the low-dose group treated with bolus 5-FU and one patient in the CVI group. The overall 5-year survival for the whole group is 71%. CONCLUSION: Dose intensity of 5-FU and dose of radiation correlate significantly with the likelihood of achieving a pCR. Continuous infusion 5-FU (CI) and a preoperative radiation dose of 5500 cGy or higher can achieve pCR rates of approximately 50%, even in fixed cancers of the rectum. PMID- 10705010 TI - Paclitaxel and concurrent radiation for gastric cancer. AB - PURPOSE: To determine the activity and toxicity of paclitaxel and concurrent radiation for gastric cancer. METHODS AND MATERIALS: Twenty-seven patients were studied. Twenty-five had proximal gastric cancers, two had distal cancers. Eight had esophageal extension, 6 had celiac adenopathy, and 7 had retroperitoneal adenopathy. Patients received paclitaxel, 50 mg/m(2) by 3-hour intravenous (IV) infusion, weekly, on days 1, 8, 15, 22, and 29. Radiation was administered concurrently to a total dose of 45.0 Gy, in 1.80 Gy fractions, for 25 treatments. Patients who were medically or surgically inoperable received a sixth week of paclitaxel with a radiation boost to 50.4 Gy. RESULTS: Esophagitis and gastritis were the most important toxicities, Grade 3 in four patients (15%), and Grade 4 in three patients (11%). Five patients (19%) had Grade 3 nausea. The overall response rate was 56%, including three patients (11%) with a complete response. The 2-year progression-free and overall survival rates were 29% and 31%, respectively. CONCLUSION: Concurrent paclitaxel and radiation demonstrates substantial local-regional activity in gastric cancer. Future investigations combining paclitaxel and radiation with other local-regional and systemic treatments are warranted. PMID- 10705011 TI - Operative and conservative management of primary gastric lymphoma: interim results of a German multicenter study. AB - PURPOSE/OBJECTIVE: Biology and appropriate management of gastrointestinal (GI lymphomas are matters of an ongoing controversial debate. To evaluate histological features, sites of involvement and management of primary GI lymphomas, a prospective multicentric study was initiated in 10/1992. Aim of study was the further standardization of operative and conservative treatment modalities. MATERIALS AND METHODS: Study started 10/1992 and was closed 11/1996. A total of 381 evaluable patients had been accrued then. Standardized diagnostic workup included endoscopic and radiological evaluation of the complete GI-tract as well as a central histological review. Diagnosis was established after Lewin, stage classification was made after Musshoff, and histological classification was made after Isaacson. Treatment decision concerning operative or conservative management was due to the initially acting physician. Patients with resection of low grade lymphoma received total abdominal irradiation 30 Gy + 10 Gy boost to incompletely resected areas. After resection of high grade lymphoma CHOP chemotherapy (4 cycles for stage IE, 6 cycles for higher stages) after McKelvy was followed by total abdominal irradiation 30 Gy for stage IE respectively involved field irradiation 30 Gy for higher stages with 10 Gy boost to incompletely resected areas. Primary conservative- treatment consisted of six cycles COP chemotherapy after Bagley for low grade lymphomas stage > IE and total abdominal irradiation 30 Gy + 10 Gy boost to involved areas for all stages. Patients with high grade lymphomas received 4 x CHOP followed by total abdominal irradiation 30 Gy + 10 Gy boost to involved areas or 6 x CHOP plus involved field radiation therapy with 40 Gy. 257 patients are considered for analysis due to exclusion criteria of the study, 190 of them were suffered from gastric lymphoma. Their median observation time is 29 months, maximum observation time is 68 months. RESULTS: Sites of involvement were stomach in 73.4%, small bowel 9.6%, ileocoecal region 6.9%, and other sites 3.2% More than one GI site was involved in 6.9%. Gastric lymphomas achieved a survival probability of 89% after 3 years. Though surgical and conservative treatment was not randomized, outcome was analyzed in gastric NHL stages I and II (histologic subtype not considered showing no significant influence). At 3 and 5 years survival is 88% in resected cases vs. 94% and 86% in conservatively treated patients (p = 0.350). Analyzing only stages I + II(1) surgery also seems of no advantage even considering only RO resections. There was one acute gastrointestinal bleeding under primary chemotherapy for a high grade lymphoma. Toxicities of grade III and IV WHO were rarely seen during treatment. All other acute toxicities were not more than grade II WHO. CONCLUSION: Conservative treatment in this setting is feasible. The operative approach seems not to be advantageous compared to conservative treatment and should be critically reconsidered. PMID- 10705012 TI - Combined radiotherapy and chemotherapy (cisplatin and 5-fluorouracil) as palliative treatment for localized unresectable or adjuvant treatment for resected pancreatic adenocarcinoma: results of a feasibility study. AB - PURPOSE: To evaluate a cisplatin-containing chemoradiotherapy (CRT) regimen followed by chemotherapy for unresectable (locally advanced group, n = 32) and resected (adjuvant group, n = 10) pancreatic adenocarcinoma. The quality of palliation and percentage of secondary resections were also studied for unresectable disease. METHODS AND MATERIALS: The protocol comprised CRT (45 Gy over 5 weeks), combined with 5-fluorouracil and cisplatin during the first and fifth weeks, followed, 3 weeks later, by 4 cycles of the same chemotherapy plus leucovorin. RESULTS: All patients completed CRT but only 50% of each group finished the entire protocol. Gastrointestinal toxicity and weight loss were the major side effects during CRT. Enhanced hematological toxicity limited the post CRT chemotherapy. For the locally advanced group, median survival was 9 months; 1 and 2-year survival rates were 31 and 12. 5%, respectively. The overall response rate was 16% and 50% had stable disease. A lasting palliative effect defined as improved performance status and decreased analgesic consumption, was recorded for 43% of the patients. Only three secondary resections have been performed. For the adjuvant group, median survival was 17 months. CONCLUSIONS: Although toxic in advanced disease, this regimen significantly lowered pain and analgesic consumption, but had poor impact on secondary resectability. In an adjuvant setting, although equally toxic, this series was too small to allow conclusions to be drawn. PMID- 10705013 TI - Combined modality treatment in unresectable extrahepatic biliary carcinoma. AB - PURPOSE: Cancers of the extrahepatic biliary tract are rare. Surgical resection is considered the standard treatment, but is rarely feasible. Several reports of combined modality therapy, including external beam radiation, often combined with chemotherapy and intraluminal brachytherapy, have been published. The purpose of this study was to evaluate the effect of chemoradiation plus intraluminal brachytherapy on response, local control, survival, and symptom relief in patients with unresectable or residual extrahepatic biliary carcinoma. METHODS AND MATERIALS: From February 1991 to December 1997, 20 patients (14 male, 6 female; mean age 61 +/- 12 years; median follow-up 71 months) with unresectable (16 patients) or residual (4 patients), nonmetastatic extrahepatic bile tumors (common bile duct, 8; gallbladder, 1; Klatskin, 11) received external beam radiation (39.6-50.4 Gy); in 19 patients, 5-fluorouracil (96-h continuous infusion, days 1-4 at 1,000 mg/m(2)/day) was also administered. Twelve patients received a boost by intraluminal brachytherapy using (192)Ir wires of 30-50 Gy, prescribed 1 cm from the source axis. RESULTS: During external beam radiotherapy, 8 patients (40%) developed grade 1-2 gastrointestinal toxicity. Four patients treated with external-beam plus intraluminal brachytherapy had a clinical response (2 partial, 2 complete) after treatment. For the total patient group, the median survival and time to local progression was 21.2 and 33.1 months, respectively. Distant metastasis occurred in 10 (50%) patients. Two patients who received external beam radiation plus intraluminal brachytherapy developed late duodenal ulceration. Two patients with unresectable disease survived more than 5 years. CONCLUSION: Our data suggest that chemoradiation plus intraluminal brachytherapy was relatively well-tolerated, and resulted in reasonable local control and median survival. Further follow-up and additional research is needed to determine the ultimate efficacy of this regimen. New chemoradiation combinations and/or new treatment strategies (neoadjuvant chemoradiation) may contribute, in the future, to improve these results. PMID- 10705014 TI - Treatment of superficial esophageal cancer by external radiation therapy alone: results of a multi-institutional experience. AB - PURPOSE: To assess the effectiveness and toxicity of external radiation therapy for superficial esophageal cancer. METHODS AND MATERIALS: During the period from March 1979 to November 1996, 78 patients with superficial esophageal cancer received radiation therapy without intracavitary irradiation at nine radiotherapy institutions in Japan. All patients had histologically-proven squamous cell carcinoma. Endoscopic ultrasonography was performed in 34 patients to discriminate mucosal from submucosal cancer. Most of the patients had received radiation therapy using conventional fractionation at an average dose of 65.5 Gy. RESULTS: The survival rates at 1, 2, and 5 years were 88%, 73%, and 45%, respectively. The local control rates at 1, 2, and 5 years were 85%, 79%, and 66%, respectively. Although the difference was not significant, the survival rate of cancer patients with a tumor invading the submucosa was lower than that of the other patients. In 6 mucosal cancer patients, local recurrence was observed in 1 patient with extensive cancer. Regional lymph node recurrence and distant failure were not observed in mucosal cancer patients, while in 28 submucosal cancer patients, the 5-year survival rate and relapse free rate were only 49% and 43%, respectively. Univariate and multivariate analysis identified age as the only significant prognostic factor. Severe late injury, such as esophageal ulcer, perforation, and bleeding, was not observed. CONCLUSION: External radiation therapy is effective for mucosal cancer. However, further investigation is needed to establish a better standard treatment protocol for submucosal cancer. PMID- 10705015 TI - Stage III thymoma: pattern of failure after surgery and postoperative radiotherapy and its implication for future study. AB - PURPOSE: With the conventional approach of surgery and postoperative radiotherapy for patients with Masaoka Stage III thymoma, progress has been slow for an improvement in the long-term survival rate over the past 20 years. The objective of this study was to evaluate the pattern of failure and survival after surgery and postoperative radiotherapy in Stage III thymoma and search for a new direction for better therapy outcome. METHODS AND MATERIALS: Between 1975 and 1993, 111 patients with thymoma were treated at Massachusetts General Hospital. Of these, 32 patients were determined to have Masaoka Stage III thymoma. The initial treatment included surgery for clinically resectable disease in 25 patients and preoperative therapy for unresectable disease in 7 patients. Surgical procedure consisted of thymectomy plus resection of involved tissues. For postoperative radiotherapy (n = 23), radiation dose consisted of 45-50 Gy for close resection margins, 54 Gy for microscopically positive resection margins, and 60 Gy for grossly positive margins administered in 1.8 to 2.0 Gy of daily dose fractions, 5 fractions a week, over a period of 5 to 6.6 weeks. In preoperative radiotherapy, a dose of 40 Gy was administered in 2.0 Gy of daily dose fractions, 5 days a week. For patients with large tumor requiring more than 30% of total lung volume included in the target volume (n = 3), a preoperative radiation dose of 30 Gy was administered and an additional dose of 24-30 Gy was given to the tumor bed region after surgery for positive resection margins. RESULTS: Patients with Stage III thymoma accounted for 29% (32/111 patients) of all patients. The median age was 57 years with a range from 27 to 81 years; gender ratio was 10:22 for male to female. The median follow-up time was 6 years. Histologic subtypes included well-differentiated thymic carcinoma in 19 (59%), high-grade carcinoma in 6 (19%), organoid thymoma in 4 (13%), and cortical thymoma in 3 (9%) according to the Marino and Muller-Hermelink classification. The overall survival rates were 71% and 54% at 5 and 10 years, respectively. Ten of the 25 patients who were subjected to surgery as initial treatment were found to have incomplete resection by histopathologic evaluation. The 5- and 10-year survival rates were 86% and 69% for patients (n = 15) with clear resection margins as compared with 28% and 14% for those (n = 10) with incomplete resection margins even after postoperative therapy, p = 0.002. Survival rates at 5 and 10 years were 100% and 67% for those with unresectable disease treated with preoperative radiation (n = 6) and subsequent surgery (n = 3). Recurrence was noted in 12 of 32 patients and 11 of these died of recurrent thymoma. Recurrences at pleura and tumor bed accounted for 77% of all relapses, and all pleural recurrences were observed among the patients who were treated with surgery initially. CONCLUSION: Incomplete resection leads to poor results even with postoperative radiotherapy or chemoradiotherapy in Stage III thymoma. Pleural recurrence is also observed more often among patients treated with surgery first. These findings suggest that preoperative radiotherapy or chemoradiotherapy may result in an increase in survival by improving the rate of complete resection and reducing local and pleural recurrences. PMID- 10705016 TI - Changes in tumor oxygen tension during radiotherapy of uterine cervical cancer: relationships to changes in vascular density, cell density, and frequency of mitosis and apoptosis. AB - PURPOSE: Changes in oxygen tension (pO(2)) during the early phase of fractionated radiotherapy were studied in 22 patients with uterine cervical cancer. The aims were to investigate (a) whether possible changes in pO(2) differed among and within tumors and (b) whether the changes could be attributed to changes in vascular density, cell density, and frequency of mitosis and apoptosis. METHODS AND MATERIALS: The pO(2) was measured polarographically in four regions of the tumors before treatment and after 2 weeks of radiotherapy. The vascular density, cell density, and frequency of mitosis and apoptosis were determined from biopsies taken from the tumor regions after each pO(2) measurement. RESULTS: The changes in pO(2) during therapy differed among the tumors and were correlated to pO(2) before treatment (p < 0.001). The direction of the changes was consistent throughout the tumors; all regions in tumors with increased oxygenation had increased or no change in pO(2) and vice versa. The tumors with increased pO(2) (n = 10) had a large decrease in cell density and a significant increase in apoptotic frequency. In contrast, the tumors with decreased pO(2) (n = 10) had a smaller decrease in cell density (p = 0.014) and no significant increase in apoptotic frequency. Vascular density and mitotic frequency showed no change during therapy; however, vascular damage other than decreased vascular density was observed. CONCLUSION: These results indicate that the oxygenation of cervix tumors generally changes during the early phase of radiotherapy. The change depends on the balance between the factor leading to an increase and that leading to a decrease in oxygenation; i.e., decreased cell density and vascular damage, respectively. Increased apoptotic frequency may contribute to a large decrease in cell density and hence increased oxygenation during therapy. PMID- 10705017 TI - Dosimetry and dose-response relationships in newly diagnosed patients with malignant gliomas treated with iodine-131-labeled anti-tenascin monoclonal antibody 81C6 therapy. AB - PURPOSE: The objective of this study was to perform the dosimetry and evaluate the dose-response relationships in newly diagnosed patients with malignant brain tumors treated by direct injections of (131)I-labeled 81C6 monoclonal antibody (MAb) into surgically created resection cavities (SCRCs). METHODS AND MATERIALS: Absorbed doses to the 2-cm-thick shell as measured from the margins of the resection cavity interface were estimated for 42 patients with primary brain tumors. MR images were used to assess the enhanced-rim volume as a function of time after radiolabeled MAb therapy. Biopsy samples were obtained from 15 patients and 1 autopsy. RESULTS: The average absorbed dose [range] to the 2-cm shell region was 32 [3-59] Gy. For the endpoint of minimal time to MR contrast enhancement, the optimal absorbed dose and initial dose-rate were 43 +/- 16 Gy and 0. 41 +/- 0.10 Gy/h, respectively. There was a correlation between the absorbed dose and dose rate to the shell region and biopsy outcome (tumor recurrence, radionecrosis, and tumor recurrence and/or radionecrosis). In this Phase I study, the maximum tolerated dose (MTD) was 120 mCi. At this MTD, the estimated average absorbed dose and initial dose rate to the 2-cm shell were 41 [9-89] Gy and 0.51 [0.24-1.13] Gy/h, respectively. These values are in agreement with the optimal values based on the time to MR lesion rim enhancement. CONCLUSIONS: The average absorbed dose to the 2-cm shell region varied considerably and mainly depended on cavity volume. In future clinical trials, the administered activity of (131)I-labeled 81C6 MAb may be adjusted based on cavity volume in order to deliver the optimal absorbed dose of 43 Gy rather than giving a fixed administered activity. PMID- 10705018 TI - Parenchymal pineal tumors: a clinicopathological study of 76 cases. AB - PURPOSE: The aim of this study was to identify factors that could lead to optimization of the management of pineal parenchymal tumors (PPT) which remains equivocal and controversial. METHODS AND MATERIALS: In order to determine factors that influence PPT prognosis, a series of 76 consecutive patients from 12 European centers with histologically proven tumors was retrospectively reviewed. The clinical records and material for histologic review were available in all cases. Follow-up was achieved in 90% of cases. RESULTS: According to WHO classification, there were 19 pineocytomas, 28 intermediate and mixed PPT, and 29 pineoblastomas. According to a four-grade institutional classification, there were 11 Grade 1, 27 Grade 2, 20 Grade 3, and 18 Grade 4. Surgical resection was attempted in 44 patients, whereas 30 had biopsy only. In one case, diagnosis was made at autopsy and in another on spinal deposits. Forty-four patients were irradiated following surgery, 15 patients received chemotherapy. Forty-one patients were alive (median follow-up: 85 months); 9 patients died perioperatively; 26 patients relapsed. Univariate analysis showed a good outcome correlated with age above 20 years, tumor diameter less than 25 mm, and low-grade histology. Multivariate analysis confirmed histology and tumor volume to be significant independent prognostic factors. The extent of surgery and radiotherapy had no clear influence on survival. CONCLUSIONS: This review highlights the prognostic features of PPT and may help to determine treatment strategies based on radiologic and pathologic characteristics. PMID- 10705019 TI - Local control with multimodality therapy for stage 4 neuroblastoma. AB - PURPOSE: To evaluate the efficacy of 21 Gy hyperfractionated radiotherapy for local control in conjunction with surgery and intensive systemic therapy for patients with Stage 4 neuroblastoma. METHODS AND MATERIALS: After achieving a partial or complete remission, 47 children, ages 1-10 years, with Stage 4 neuroblastoma were treated on four consecutive institutional protocols (N4-N7) with dose-intensive multi-agent chemotherapy, maximal surgical debulking, and hyperfractionated radiotherapy (1.5 Gy twice a day to 21 Gy). Radiotherapy fields encompassed the initial tumor volume and regional lymph nodes plus a 3-cm margin. This was followed by consolidation with either autologous bone marrow transplantation (N4 and N5) or immunotherapy (N6 and N7). RESULTS: Forty-five of 47 patients had a complete response to surgery and chemotherapy prior to radiotherapy. Five-year actuarial rates of local control, progression-free survival, and overall survival were 84%, 40%, and 45%, respectively. Among 26 patients who relapsed, 1 failed only at the primary site, 22 developed distant metastases exclusively, and 3 had both local and distant failures. There were no acute complications of radiotherapy. CONCLUSION: Hyperfractionated radiotherapy to 21 Gy, in conjunction with dose-intensive systemic therapy and aggressive surgical resection, is well tolerated and is associated with durable local control for most patients with Stage 4 neuroblastoma. PMID- 10705020 TI - A role for radiotherapy in neuropathic bone pain: preliminary response rates from a prospective trial (Trans-tasman radiation oncology group, TROG 96.05) AB - PURPOSE: Radiotherapy (RT) has a proven role in palliation of pain from bone metastases with numerous randomized trials obtaining response rates (RRs) of typically 70-80% regardless of the fractionation employed. However RT for neuropathic bone pain (NBP), i.e., pain with a radiating cutaneous component due to compression/irritation of nerves by tumor has not previously been studied, and its role is thus uncertain. METHODS AND MATERIALS: In February 1996, the Trans Tasman Radiation Oncology Group (TROG) initiated a multicenter randomized trial comparing a single 8 Gy fraction with 20 Gy in 5 fractions for NBP with an accrual target of 270. Formal interim analyses were planned at 90 and 180 patients. The 90th patient was accrued in June 1998, and data from the first interim analysis with both arms combined form the basis of this report. RESULTS: Forty-four patients were randomized to a single 8 Gy, 46 to 20 Gy in 5 fractions. The commonest primary sites were prostate (34%), lung (28%) and breast (10%). Median age was 68 years (range 37-89). The index site was spine (86%), rib (13%), base of skull (1%). On an intention-to-treat basis, the overall RR was 53/90 = 59% (95% CI = 48-69%), with 27% achieving a complete response and 32% a partial response. The overall RR for eligible patients was 49/81 = 60% (95% CI = 49-71%) with 27% and 33% achieving complete and partial responses respectively. Estimated median time to treatment failure was 3.2 months (95% CI = 2.1-5.1 months), with estimated median survival of 5.1 months (95% CI = 4.2-7.2 months). To date, six spinal cord/cauda equina compressions and four new or progressive pathological fractures have been detected at the index site after randomization, although one cord compression occurred before radiotherapy was planned to commence. In February 1999, the Independent Data Monitoring Committee strongly recommended continuation of the trial. CONCLUSION: Although these results are preliminary, it seems clear that there is indeed a role for RT in the treatment of NBP. Analysis of outcome by treatment arm awaits completion of the randomized trial. PMID- 10705021 TI - Normal tissue complication probabilities: dependence on choice of biological model and dose-volume histogram reduction scheme. AB - PURPOSE: To evaluate the impact of dose-volume histogram (DVH) reduction schemes and models of normal tissue complication probability (NTCP) on ranking of radiation treatment plans. METHODS AND MATERIALS: Data for liver complications in humans and for spinal cord in rats were used to derive input parameters of four different NTCP models. DVH reduction was performed using two schemes: "effective volume" and "preferred Lyman". DVHs for competing treatment plans were derived from a sample DVH by varying dose uniformity in a high dose region so that the obtained cumulative DVHs intersected. Treatment plans were ranked according to the calculated NTCP values. RESULTS: Whenever the preferred Lyman scheme was used to reduce the DVH, competing plans were indistinguishable as long as the mean dose was constant. The effective volume DVH reduction scheme did allow us to distinguish between these competing treatment plans. However, plan ranking depended on the radiobiological model used and its input parameters. CONCLUSIONS: Dose escalation will be a significant part of radiation treatment planning using new technologies, such as 3-D conformal radiotherapy and tomotherapy. Such dose escalation will depend on how the dose distributions in organs at risk are interpreted in terms of expected complication probabilities. The present study indicates considerable variability in predicted NTCP values because of the methods used for DVH reduction and radiobiological models and their input parameters. Animal studies and collection of standardized clinical data are needed to ascertain the effects of non-uniform dose distributions and to test the validity of the models currently in use. PMID- 10705022 TI - Acute and late toxicity of patients with inflammatory bowel disease undergoing irradiation for abdominal and pelvic neoplasms. AB - PURPOSE: Little data exists in the medical literature describing the response of patients with inflammatory bowel disease (IBD) to abdominal and pelvic irradiation. To clarify the use of this modality in this setting, this study assesses the short- and long-term tolerance of 28 patients with IBD to abdominal and pelvic irradiation. METHODS AND MATERIALS: From 1970 to 1999, 28 patients with IBD (10 patients-Crohn's disease, 18 patients-ulcerative colitis) were identified and underwent external beam abdominal or pelvic irradiation. Mean follow-up time after radiation therapy was 32 months. Patients were treated either by specialized techniques (16 patients) to minimize small and large bowel irradiation or by more conventional approaches (12 patients). Acute and late toxicity was scored. RESULTS: The overall incidence of severe toxicity was 46% (13/28 patients). Six of 28 patients (21%) experienced severe acute toxicity necessitating cessation of radiation therapy. Late toxicity requiring hospitalization or surgical intervention was observed in 8 of 28 patients (29%). One patient experienced both an acute as well as late toxicity. For patients undergoing radiation therapy by conventional approaches, the 5-year actuarial rate of late toxicity was 73%. This figure was 23% for patients treated by specialized techniques (p = 0.02). CONCLUSIONS: Because of the potentially severe toxicity experienced by patients with IBD undergoing abdominal and pelvic irradiation, judicious use of this modality must be employed. Definition of IBD location and activity as well as careful attention to irradiation technique may allow treatment of these patients with acceptable rates of morbidity. PMID- 10705023 TI - New scoring system identifies kidney outcome with radiation therapy in acute renal allograft rejection. AB - PURPOSE: To evaluate the role of radiation therapy for acute refractory renal rejection after failure of medical intervention, and to identify risk factors that influence graft survival following radiation therapy. METHODS: Between June 1989 and December 1995, 53 renal transplant recipients (34 men and 19 women) were treated with localized radiation therapy for acute renal allograft rejection. Graft rejection was defined as an increase in serum creatinine with histologic evidence of rejection on renal biopsy. Ninety-one percent were cadaveric transplant recipients. The majority of patients who experienced acute graft rejection initially received corticosteroid therapy, except for 25% who were referred for radiation therapy and steroids for the first rejection. In more recent years, patients with moderate or severe steroid-resistant or recurrent rejection received OKT3, a polyclonal antilymphocyte antibody (ATGAM), tacrolimus (FK506), or mycophenolate mofetil (MMF). Patients who failed to respond to medical treatment were then referred for radiation therapy. Ultrasound was performed for kidney localization. Treatment consisted of a dose of 600 cGy given in 3 or 4 fractions using 6 MV photons, delivered AP or AP/PA. RESULTS: The overall actuarial graft survival from the initiation of RT was 83% at 1 month, 60% at 1 year, and 36% at 5 years. The median follow-up from the date of transplant to the last follow-up was 22 months. The median time from the date of transplant to the initiation of radiotherapy was 3 months, and the median time from the initiation of radiotherapy to the last follow-up was 10 months. Variables evaluated were as follows: human leukocyte antigen matching on HLA-A, HLA-B, and HLA-DR, the transplant panel-reactive antibodies (PRA) at transplantation, number of acute rejection episodes, interval from the date of the transplant to the first rejection, serum creatinine levels at the time of the first radiation treatment, number of transplants, and concomitant immunosuppressive therapy. Independent factors examined by Cox regression modeling were: gender (p = 0.005), creatinine levels (p = 0.000), HLA-DR (p70% (p = 0.014). Each factor was scored using integral coefficients to generate four different groups. The Kaplan-Meier survival analyzed by group produces an interpretable separation of the risk factors for graft loss. CONCLUSIONS: The outcome in patients treated with radiation therapy for acute renal graft rejection can be predicted by a novel scoring system. Patients with scores of three or less are able to achieve 100% renal graft salvage, while patients who have scores of 12 or higher are not able to be salvaged with the current radiation therapy regimen. Future studies should be directed toward identifying more effective treatment for patients who have a high score based on our criteria. The scoring system should be utilized to identify patients at risk who could benefit from radiation therapy. Further study with a randomized trial utilizing this scoring system is needed to confirm the validity of the scoring system in predicting graft survival and the efficacy of radiation in patients who receive radiation therapy for acute graft rejection. PMID- 10705024 TI - Hypoxia and necrosis in rat 9L glioma and Morris 7777 hepatoma tumors: comparative measurements using EF5 binding and the Eppendorf needle electrode. AB - PURPOSE: The purpose of this study was to assess the presence of tumor hypoxia using two independent techniques: binding of the 2-nitroimidazole EF5 and Eppendorf needle electrode measurements. The distribution of tumor hypoxia was assessed with respect to tumor necrosis in corresponding histological studies. METHODS AND MATERIALS: Each of several rats bearing a subcutaneous 9L glioma or Morris 7777 hepatoma tumor was given EF5 i.v. to a final, whole-body concentration of 100 microM. About 2.5 h later, each rat was anesthetized, and needle electrode measurements were made in the tumor along 1-5 tracks (30-200 individual measurements). At 3 h post-EF5 injection, the tumor was excised and frozen. Frozen sections were analyzed for the presence and distribution of binding of EF5 and necrosis using immunohistochemical techniques followed by staining with hematoxylin and eosin (H&E). The histochemical analysis and electrode readings in similar regions of the tumor were compared. RESULTS: Electrode measurements were taken at 0.4-mm intervals along one-dimensional tracks, whereas EF5 binding measurements from tissue sections contained two dimensional information at high spatial resolution ( approximately 2.5 micro). The EF5 measurements showed greater spatial heterogeneity than did the electrode measurements. In tumor regions with minimal necrosis, needle tracks with relatively high pO(2) readings were usually found to contain relatively low EF5 binding, and vice versa. Because EF5 binding is inversely related to tissue pO(2), this result was expected. The expected inverse correlation of the two techniques was most disparate in necrotic tumor regions (confirmed by H&E staining), where needle electrode measurements showed low to zero pO(2) values, but little or no EF5 binding was found. CONCLUSION: The two methods compared in this study operate in fundamentally different ways and provide substantially different information. EF5 binding provided detailed spatial information on the distribution of hypoxia in viable tumor tissue. There was no EF5 binding in necrotic tumor tissue because cells in such tissue were unable to metabolize the drug. In contrast, output from the needle electrode method appeared to represent a "track-average" tissue pO(2) and did not distinguish between extreme hypoxia and either macroscopic or microscopic necrosis. At the present time, the importance of tumor necrosis in determining treatment response is unknown. However, our data suggest that the Eppendorf needle electrode technique will overestimate the presence of hypoxia. Both techniques are potentially limited by sampling errors in tumors with heterogeneous distributions of hypoxia. PMID- 10705025 TI - The 90-day coronary vascular response to (90)Y-beta particle-emitting stents in the canine model. AB - PURPOSE: Long-term preclinical studies using continuous, low-dose-rate vascular brachytherapy with (32)P beta-emitting stents have yielded largely disappointing results. In contrast, a shorter half-life, higher dose-rate (90)Y beta-emitting stent more closely mimics the delivery dose rate characteristics of clinically effective beta- and gamma-wire and balloon brachytherapy devices. We evaluated the dose response characteristics of a (90)Y beta-emitting stent in the canine coronary injury model and hypothesized that this device would reduce neointimal formation. METHODS: Seventy-seven (90)Y beta-emitting coronary stents (15 mm BXTM, 3.0- and 3.5-mm diameter) were implanted in 26 normal dogs (20-25 kg) using a randomized, blinded study design. Stent activity included nonradioactive controls (n = 24), 4.5 microCi (n = 15), 8 microCi (n = 12), 16 microCi (n = 18), and 32 microCi (n = 8). Histologic endpoints were assessed at 3 months. RESULTS: Luminal stenosis and neointimal area were similar in control stents and low activity (4.5 and 8 microCi) (90)Y stents. Higher activity stents (16 and 32 microCi) were associated with significant adverse effects. Frequent total occlusions (5 of 18 stents, 28%; p = 0.008) and a 40% increase in neointimal area (p = 0.024 vs. control) occurred in the 16 microCi group. Incomplete neointimal healing and a trend for reduced neointimal cell density were evident only in the 16- and 32-microCi group. CONCLUSION: Despite unique characteristics (2.7 day half-life and a higher dose rate) of (90)Y beta-emitting coronary stents, they have an adverse effect on neointimal formation, including frequent total occlusions at high activity levels. Incomplete healing, present 90 days (33 half lives) after stent placement, indicates prolonged recovery from radiation injury. PMID- 10705026 TI - Radiation doses to the cell nucleus in single cells and cells in micrometastases in targeted therapy with (131)I labeled ligands or antibodies. AB - PURPOSE: The aim of this study was to theoretically investigate how the radiation dose to cell nuclei depends on the subcellular position of (131)I. The influence of the size of the cells and crossfire irradiation in clusters of cells was also studied. METHODS AND MATERIAL: Using data describing the dose rate around a point source of (131)I, we calculated the dose distributions inside and around cell models of different sizes. The assumed positions of (131)I were on the cellular or nuclear membrane, in the cytoplasm, in the nucleus, or spread in the whole cell. The mean doses to the nucleus of the targeted cell and to the nuclei of its neighbors were calculated using the dose distributions. RESULTS: The dose distributions inside a single targeted cell showed very different distribution profiles depending on the subcellular position of the (131)I. Targeting the nucleus instead of the cellular membrane could increase the dose to the nucleus 10-fold. Crossfire irradiation can be the major contributor to the nuclear dose in clusters of more than six cells. CONCLUSIONS: Dosimetry without microscopic considerations is inadequate for targeted radionuclide therapy of disseminated or clustering tumor cells exposed to (131)I. Therapeutic doses could be achieved, even in single cells, when (131)I was positioned near, or inside the cell nucleus, or when the clusters were large enough. PMID- 10705027 TI - Improvement of combined modality therapy with cisplatin and radiation using electroporation of tumors. AB - PURPOSE: To evaluate whether a local drug delivery method, i.e., electroporation of tumors, increases the radiosensitizing effect of cisplatin. METHODS AND MATERIALS: Subcutaneous Ehrlich-Lettre ascites (EAT) tumors in CBA mice were treated either by cisplatin, electric pulses, or ionizing radiation. In electrochemotherapy protocol, electric pulses were given to the tumor 3 min after intravenous injection of cisplatin. The interval between electrochemotherapy and irradiation was 20 min. Treatment effectiveness was evaluated by tumor growth delay and local tumor curability. RESULTS: Electrochemotherapy of EAT tumors proved to be effective treatment, resulting in 12% tumor cures, whereas treatment with cisplatin or electric pulses alone did not yield any tumor cures. As expected, injection of cisplatin 20 min prior to irradiation, increased radioresponse of tumors from 27% to 73% tumor cures. Electroporation of tumors also increased radiation response of tumors to 54% tumor cures. Electrochemotherapy given prior to irradiation increased radioresponsiveness of tumors, resulting in 92% tumor cures. CONCLUSIONS: This study shows that delivery of cisplatin into the cells by electroporation of tumors increases the radiosensitizing effect of cisplatin. However, some effect may also be ascribed to application of electric pulses to the tumors that in our study also predisposed tumor cells to radiation damage. PMID- 10705028 TI - Beta versus gamma for catheter-based intravascular brachytherapy: dosimetric perspectives in the presence of metallic stents and calcified plaques. AB - PURPOSE: Both beta and gamma emitters are currently used in the catheter-based intravascular brachytherapy. The dosimetric effects due to the presence of metallic stents and calcified plaques have not been fully addressed. This work compares these effects for two most commonly used beta and gamma sources ( (90)Sr and (192)Ir). MATERIALS AND METHODS: An EGS4 Monte Carlo package was used to calculate dose in water for a (90)Sr (supplied by NOVOSTE) and an (192)Ir (Supplied by BEST) source, with or without the presence of a calcified plaque or a metallic stent. Plaques of different shape (shell and disk), size and density, and two types of stainless-steel stents (ring or mesh stent) were studied. The ring stent consists of identical rings stacked along the long axis of the sources. The gap between two rings is 0.3 mm. The mesh stents are made of identical square (0.1 x 0.1 or 0.2 x 0.2 mm(2)) holes separated from each other by stainless-steel wire. The cross section of wire for both ring and mesh stents is 0.1 x 0.1 mm(2). A dose perturbation factor (DPF), defined as the ratio of the doses with and without the presence of a plaque or a stent, was introduced to quantify the effects. A carefully chosen set of EGS4 transport parameters for the small geometry in question was used in the calculation. RESULTS: The radial and axial dose distributions calculated in water were found to agree with the published measurements to within 3%. The dose perturbations due to the presence of calcified plaques or metallic stents were found far more significant for the (90)Sr source than those for the (192)Ir source. Up to 30% dose reduction behind a plaque were observed for the (90)Sr source, while the dose reduction for the (192)Ir source was found to be negligible. The dose enhancement inside a plaque was as high as 10% for the beta source or 6% for the gamma source. In the presence of a stent, the DPF was in the range of 1.15-0.75 for the beta source, while it was almost equal to 1.0 for the gamma source. CONCLUSION: The dose perturbation due to the presence of a calcified plaque or a metallic stent is significant for the beta source. The dose reduction in the region beyond a plaque or a stent could be more than 20%. For the gamma source, the dose effect behind a plaque or a stent is practically negligible. These dosimetric differences between the beta and gamma sources in the presence of a calcified plaque or metallic stent should be considered in the dose prescription of intravascular brachytherapy. PMID- 10705029 TI - Mucosal dose prescription in endobronchial brachytherapy: a study based on CT dosimetry. AB - PURPOSE: To investigate the consequences of using different dose prescription methods for endobronchial brachytherapy (EB), both with and without the use of a centered applicator. MATERIALS AND METHODS: A CT scan was performed during EB procedures in 13 patients after insertion of the lung applicator. A dosimetric analysis was subsequently performed in five of these patients using a 3D brachytherapy treatment planning system (PLATO v13.3, Nucletron). RESULTS: Dose prescription to the mucosa yields uniform dose distributions to the bronchial mucosa when a centrally positioned applicator is used. When non-centrally positioned applicators are used, mucosal dosing results in a significant underdosage to parts of the target volume. Due to the rapid dose fall-off in EB, dose prescription to the mucosa resulted in inadequate coverage of the outer portion of the bronchial wall and adjacent peribronchial space. When compared to mucosal dose prescription, prescription to the outer aspect of the bronchial wall appears to improve target coverage while limiting the hyperdose (i.e., 200%) volume. The diameters of the different bronchial segments, as determined by CT measurements in 13 patients, correlated well with calculated values based upon the tracheal diameter. CONCLUSIONS: Mucosal dose prescription should only be used in combination with centered EB applicators. Given the rapid dose fall-off in EB mucosal dose prescription should be used with caution in curative treatments where EB, without additional external radiotherapy, is used as the sole treatment modality. In curative EB, both improved target coverage and a limited hyperdose volume can be achieved by dose prescription to the outer aspect of the bronchial wall. PMID- 10705030 TI - A glass compensator filter to improve breast image quality in radiation therapy simulation. AB - PURPOSE: To improve the image quality of simulation films in tangential radiotherapy for breast cancer, we have designed a new compensator filter for the variation of breast contour using high-density-glass material. METHODS AND MATERIALS: The measurements and analyses of the body contour were done using CT scans, taken in the treatment position, of 20 breast cancer patients. The maximum tissue deficit that needed to be compensated for was 8 cm, and the authors fabricated the compensator system using high-density-glass material to maintain transparency. The glass compensator can be attached to the accessory mount of the simulator head and its position can be easily adjusted according to breast shape and position. The image qualities of simulation films taken with and without the glass compensator in tangential breast radiotherapy field were compared and the film densitometry was performed using the humanoid phantom. RESULTS: Using this compensator system, the overall image quality improved, resulting in enhanced contrast and resolution of the breast simulation image. The delineator wires for the beam margins were also well depicted, and the surgical clips within the breast tissue can be easily demonstrated. The film densitometry resulted in much less saturation over the breast tissue when using the glass compensator. CONCLUSION: Using the glass compensator system, the geographical miss may be reduced with the virtue of the improved image quality. PMID- 10705031 TI - Use of an electron reflector to improve dose uniformity at the vertex during total skin electron therapy. AB - PURPOSE: The vertex of the scalp is always tangentially irradiated during total skin electron therapy (TSET). This study was conducted to determine the dose distribution at the vertex for a commonly used irradiation technique and to evaluate the use of an electron reflector, positioned above the head, as a means of improving the dose uniformity. METHODS AND MATERIALS: Phantoms, simulating the head of a patient, were irradiated using our standard procedure for TSET. The technique is a six-field irradiation using dual angled electron beams at a treatment distance of 3.6 meters. Vertex dosimetry was performed using ionization methods and film. Measurements were made for an unmodified 6 MeV electron beam and for a 4 MeV beam obtained by placing an acrylic scattering plate in the beam line. Studies were performed to examine the effect of electron scattering on vertex dose when a lead reflector, 50 x 50 cm in area, was positioned above the phantom. RESULTS: The surface dose at the vertex, in the absence of the reflector, was found to be less than 40% of the prescribed skin dose. Use of the lead reflector increased this value to 73% for the 6 MeV beam and 99% for the degraded 4 MeV beam. Significant improvements in depth dose were also observed. The dose enhancement is not strongly dependent on reflector distance or angulation since the reflector acts as a large source of broadly scattered electrons. CONCLUSION: The vertex may be significantly underdosed using standard techniques for total skin electron therapy. Use of an electron reflector improves the dose uniformity at the vertex and may reduce or eliminate the need for supplemental irradiation. PMID- 10705032 TI - Assessment of large single-fraction, low-energy X-ray dose with radiochromic film. AB - PURPOSE: To investigate the accuracy of in vivo dosimetry using radiochromic film for large single-fraction, low-energy irradiations. METHODS AND MATERIALS: Gafchromic MD-55-2 radiochromic film and LiF thermoluminescent dosimeters (TLDs) were placed in vivo on 25 patients to ascertain their effectiveness for assessment of dose. All patients received 10 Gy single fractions at energies ranging from 100 kVp (half-value layer [HVL] = 3.5 mm Al) up to 250 kVp (HVL = 2.3 mm Cu). Effects of small air gaps were also investigated using LiF TLDs and radiochromic film. RESULTS: Radiochromic film adequately measured applied dose for 25 patients in vivo with a standard deviation of 5.5% from prescribed dose. LiF TLDs recorded a standard deviation of 4.1% from measured to applied dose. Small air gaps which can be created under the film or TLDs during in vivo dosimetry were shown to have a measurable but minimal effect on results for gaps less than 5 mm. CONCLUSIONS: Gafchromic film has adequately measured applied dose in vivo at low energy for large 10 Gy single-fraction irradiation. PMID- 10705034 TI - Genetic approaches to the study of replicative senescence. AB - Genetic analyses of replicative senescence have revealed the dominance of the senescent phenotype since whole cell fusion of normal with immortal cells yields hybrids having limited division potential. We exploited the recessive nature of immortality by fusing different immortal human cell lines with each other and identified four complementation groups for indefinite division. This allowed for a focussed approach involving microcell mediated chromosome transfer that led to the implication of chromosomes 1, 4 and 7 as loci for cell senescence genes. More recently we have cloned the gene on chromosome 4, MORF 4. It is a member of a family of genes with motifs suggestive of transcriptional regulators. Characterization of this novel gene family should lend further insights into the phenomenon of replicative cell senescence. PMID- 10705035 TI - Genetics of aging: lessons from centenarians. AB - Aging is a universal phenomenon that affects nearly all animal species. It can be considered as the product of an interaction between genetic, environmental and lifestyle factors, which in turn influence longevity that varies between and within species. Several studies have been focused in healthy centenarians, because these exceptional individuals represent the best example of successful aging. These studies have shown that centenarians have escaped the major age associated diseases, they have well conserved several immune parameters, and at least one gene allele has been identified and linked with longevity. In parallel, studies at cellular level have identified several genes that influence, positively or negatively, normal replicative in vitro life-span. The ability of these genes to regulate aging in vitro, in conjunction with the telomeres shortening hypothesis have raised the intriguing question of the existence of a molecular clock that counts and thus may modulates human aging and longevity. This review article will discuss these issues, focusing in the nature of the genetic factors that associate with these phenomena. PMID- 10705036 TI - Epigenetic aspects of cellular senescence. AB - The limited proliferative potential of normal cells in culture has been proposed as a model for cellular aging in vivo. It is clear that cellular aging has a genetic component but epigenetic processes could also be involved. Insight gained during years of intensive study suggests cellular aging is a multi-step process and that cells possess a counting mechanism that determines the number of doublings the cells can complete. In this paper, we review evidence suggesting a role for epigenetic processes in cell senescence and discuss the possible insights that might be provided by experiments designed to induce a premature senescent like state. PMID- 10705037 TI - Inhibin and reproductive aging. PMID- 10705038 TI - Involvement of prolactin in breast cancer: redefining the molecular targets. AB - The mammary gland is the major target tissue of prolactin (PRL) in mammals. Although this pituitary hormone has been long suspected to be involved in the progression of human breast cancer, the failure of clinical improvement by treatment with dopamine agonists (which lower circulating levels of PRL) rapidly reduced the interest of oncologists concerning a potential role of PRL in the development of breast cancer. Within the last few years, however, several studies reported first, that PRL is also synthesized by mammary epithelial cells, and second that it may exert a proliferative action in an autocrine/paracrine manner. In agreement with a recent epidemiological study, these observations have led to a reconsideration of the role of PRL as an active participant in breast cancer, and are an impetus to redefine the molecular targets of anti-prolactin strategies since dopamine analogs are assumed to be inefficient on extrapituitary PRL synthesis. In this review, we briefly summarize the current knowledge of PRL effects on both normal and tumor mammary cells, and we discuss the most relevant articles supporting the autocrine-paracrine action of PRL in the breast. With the aim of defining putative new molecular targets, we propose an overview of the main PRL receptor signaling cascades known to be triggered by PRL in mammary epithelial cells or, when not available, in other cell types. Finally, because proteolytic fragments of rat PRL have been shown to inhibit the angiogenic process, which may be relevant for preventing the progression of solid tumors such as breast tumors, we discuss the hypothesis that the enzymatic cleavage of human PRL could also represent a new molecular target in the search for alternative strategies in the treatment of breast cancer. PMID- 10705039 TI - Epidermal differentiation, apoptosis, and senescence: common pathways? AB - Regulation and execution of epidermal terminal differentiation, apoptosis and cellular senescence share some molecular and cellular features. The three phenomena result in suppression of proliferation and in some instances they seem to overlap. The boundaries between keratinocyte terminal differentiation and senescence are unclear and the former has been proposed to be a form of apoptosis. However, cumulative evidence argues that they are alternative, independent responses to different stimuli. I summarize in this review classical and recent evidence underlying the molecular control of these biological processes and propose a rationale to understand their nature and function in epidermis. PMID- 10705040 TI - The influence of oxygen toxicity on yeast mother cell-specific aging. AB - The effect of deleting both catalase genes and of increased oxygen as well as paraquat (a pro-oxidant) on the replicative life span of yeast mother cells has been investigated to test the so-called oxygen theory of aging. This is well established in higher organisms, but has not been extensively tested in the unicellular yeast model system. Life span determinations were performed in ambient air or in a controlled atmosphere (55% oxygen) and an isogenic series of strains deleted for one or both yeast catalases was used and compared with wild type. In the absence of cellular catalase, increased oxygen caused a marked decrease in life span that could be completely reversed by adding 1 mM GSH, a physiological antioxidant, to the yeast growth medium. In a second unrelated strain, the effects were similar although even the wild type showed a decrease in life span when oxygen was increased. The effect could again be compensated by addition of extracellular GSH. Our results show that manipulating the detoxification of reactive oxygen species has a profound effect on yeast aging. These findings are discussed in the light of recent results relating to oxygen toxicity in the aging process of higher organisms. PMID- 10705041 TI - Testing the heterogeneity theory of late-life mortality plateaus by using cohorts of Drosophila melanogaster. AB - Variation among individuals in robustness has been posed as a general explanation for the lack of increase in late-life mortality rates. Here, we test corollaries of this heterogeneity theory. One is that populations that have undergone strong laboratory selection for differentiated stress resistance should show significant differences in their late-life mortality schedules. To test this corollary, we employed 40 410 flies from three groups of Drosophila melanogaster populations that differ substantially in their resistance to starvation. No significant differences between these groups were found for late-life mortality. Another corollary of the heterogeneity theory is that there should be late-life plateaus in stress resistance that coincide with the plateau stage of the mortality curve. In 20 994 flies from six replicate outbred laboratory populations, we measured mortality rates every other day and starvation and desiccation resistance every 7 days. Both male and female starvation and desiccation resistance clearly decreased with time overall. There was no late-life plateau in male desiccation resistance. A late-life plateau in male starvation resistance may exist, however. Together, these two experiments generally constitute evidence against heterogeneity as a major contributor to the phenomenon of late-life mortality plateaus. PMID- 10705042 TI - Aging of human palatal salivary glands: a histomorphometric study. AB - The aim of the present study was to examine age-related changes in the parenchymal and stromal components of palatal salivary glands of healthy subjects. Palatal salivary gland biopsies were obtained from 120 autopsies and were divided into young, adult, and old age groups. Histomorphometric measurements were performed on hematoxylin and eosin (H&E) stained slides. Parenchymal components included acini and ducts, and stromal components included connective tissue, blood and lymphatic vessels, inflammatory infiltrate, and adipose tissue. The mean volume fraction of each component in each age group was calculated. Statistical analysis was performed by one-way ANOVA and Tamhane tests. The mean volume fraction of the acinar component demonstrated a significant age-related decrease of 48% (P < 0.001). The mean volume fractions of the ducts and of all the stromal components demonstrated a significant age related increase (P < 0. 001). The inflammatory infiltrate component had the highest increase with aging (1471%), followed by the ducts (177%), blood and lymphatic vessels (138%), adipose tissue (130%), and connective tissue (60%). These age-related changes, the first to be reported in palatal salivary glands from healthy subjects, are different from those described in the labial salivary glands, especially in regard to the significant increase in the parenchymal ductal component, as well as in the stromal inflammatory infiltrate and adipose tissue components. It can be suggested that these changes could have important implications regarding the age-related function of these glands. PMID- 10705043 TI - The effects of aging on enzyme activities and metabolite concentrations in skeletal muscle from sedentary male and female subjects. AB - Aging affects the metabolic capacity of skeletal muscle, in particular the glycolytic and respiratory capacities. The purpose of this study was to quantify biochemical alterations due to aging in muscular metabolic capacity in human skeletal muscles in sedentary subjects. The activities of various marker enzymes and metabolites related to glycolysis, Krebs' cycle and the electron transfer chain and high energy phosphate compounds were measured in muscle biopsies from the rectus abdominis, vastus lateralis, and gluteus maximus muscles of 76 sedentary subjects (32 males and 44 females) between 15 and 91 yr. No significant differences between males and females were found, but changes related to age were: a decrease in hexokinase and lactate dehydrogenase activities in the rectus abdominis; a decrease in citrate synthase activity and citrate in the vastus lateralis; an increase in pyruvate kinase activity and a decrease in ATP and creatine phosphate concentrations in the gluteus maximus. These data suggest that distinct muscles may respond differently to aging regardless of sex in sedentary subjects. PMID- 10705044 TI - The accelerated occurrence of age-related changes of organism in Chernobyl workers: a radiation-induced progeroid syndrome? AB - The rate of aging was studied in 306 persons working at Chernobyl Atomic Power Station after the accident by means of integral and partial biological age assessment. An accelerated rate of aging was found in 81% of men and in 77% of women in comparison with a control random population sample of Kiev. Persons younger than 45 years appeared to be more vulnerable to radiation. The biological age of persons who worked in the contaminated zone immediately after the disaster exceeded the biological age in those who arrived in Chernobyl 4 months later. The biological age in the investigated persons exceeded its average populational value for 5 years (the integral biological and partial cardiopulmonary age) and for 11 years for the partial psychological age. These data may underlie the concept of radiation progeroid syndrome as the form of accelerated aging. PMID- 10705045 TI - High sensitivity and specificity of a laboratory-based serological test, pylori DTect ELISA, for detection of Helicobacter pylori infection. AB - A number of commercial ELISA kits are now available for detection of Helicobacter pylori infection. Generally, whereas the manufacturers have claimed high sensitivity and specificity, independent studies have often failed to confirm the results. The aim of this study was to independently evaluate the pylori DTect ELISA, a commercial kit for detection of H. pylori infection, in Australian patients with dyspepsia and reflux symptoms. Two hundred and nine consecutive patients (102 males and 107 females, mean age 52.8 years) who were referred for endoscopy due to upper gastrointestinal symptoms, but had not received anti-H. pylori therapy were enrolled. A 10 mL blood sample was obtained from each subject and used to evaluate the kit. The absorbance index (AI) was calculated from the mean of two readings of optical density (OD) of each serum sample. Eight biopsies from the gastric antrum (x3), body (x2), fundus (x2), and incisura (x1) were obtained from each patient for CLO-testing (x1), culture (x3), and histological examination (x4) for H. pylori. Overall, 84 (40.2%) patients were infected with H. pylori as determined by the biopsy-based "gold standard." The AIs ranged from 0 to 1.86; 0.12 to 1.86 in H. pylori positive patients and 0 to 1.49 in negative patients. The pylori DTect ELISA obtained an accuracy of 94 to 95% under AI ranges between 0.20 to 0.40, with the highest accuracy being 95% under AIs of 0.25 and 0.35. An AI of 0.25 was recommended as the best cut-off AI, with a sensitivity of 96.4%, specificity of 93.6%, positive predictive value of 91% and negative predictive value of 97.5%. It is concluded that the pylori DTect ELISA is accurate for detecting H. pylori infection in patients with dyspepsia and reflux symptoms in Australia, when an AI of 0.25 is taken as the cut-off value. PMID- 10705046 TI - Molecular characterization and LD(50) identify virulent isolates of Staphylococcus epidermidis from adult sepsis. AB - Staphylococcus epidermidis plays an important role in infections of patients with implanted prosthetic devices. The exact clinical significance of recovered S. epidermidis from clinical specimens is difficult to assess, as they are inhabitants of the normal skin. In this study, 11 adults with clinical sepsis and blood cultures that grew only S. epidermidis were the host population. Bacterial virulence in vivo was determined by using the mouse LD(50) assay where the intravenous lethality was determined for each patient isolate. Bacterial dose (CFU x 10(9)) that produced lethality in 50% of the animals at 12 h was the value used for comparison. Restriction fragment length polymorphism (RFLP) analysis of chromosomal DNA by pulsed-field gel electrophoresis (PFGE) was used for identification of individual strains and their clonal organization. Confirmation of species assignment was done by RFLP analysis of 16S + 23S rRNA gene regions (ribotyping). Plasmid profile analysis was also conducted. Four of 11 blood isolates from adults with S. epidermidis sepsis had indistinguishable or closely related DNA patterns and were considered clone A. The same clone was previously seen to account for the majority of sepsis in a neonatal intensive care unit. There were significant differences in virulence characteristics of the S. epidermidis isolates. Clone A isolates produced lethality by LD(50) in mice at a dose averaging 2.35; clone B isolate at a dose of 2.54, and the remaining isolates, representing six distinct clones, were lethal to mice at significantly larger doses (3.51-5.17, average 4.16). These data suggest that individual clones of S. epidermidis isolated from septic adults have detectable differences in virulence as defined by an animal bioassay, and the more virulent clone is widespread. PMID- 10705047 TI - Streptococcus pneumoniae from ophthalmic infections: serotype distribution and penicillin susceptibility. AB - Streptococcus pneumoniae is one of the pathogens causing infection of the conjunctiva and the uveal tract. The present study began with the observation of two ophthalmic S. pneumoniae isolates showing intermediate resistance to penicillin. Among the 25 isolates of S. pneumoniae from 617 ophthalmic specimens, during the period of 14 months, four were found to exhibit an intermediate resistance to penicillin. Minimum Inhibitory Concentration values ranging from 0.125 microg/mL to 0.25 microg/mL was observed. No multidrug resistant strains were isolated. Serogrouping/typing of the S. pneumoniae revealed the following serogroups/types; 6A (n = 3), 6B (n = 2), 22 (n = 3), 14 (n = 3), 23A (n = 2), and 1 each of 23B, 19A, 7B, 32, 9, 42, 21, 39, 10, 3, and 34. One strain showed cross reaction in pool 29, 35, and 47. These findings represent the first such observation of ophthalmic isolates from India. PMID- 10705048 TI - Assessment of the FAN anaerobic bottle for culture of continuous ambulatory peritoneal dialysis fluid using the BacT/Alert system. AB - The purpose of this study was to determine if the newly available FAN anaerobic bottle (FANAN) alone would be comparable to the combination of the FAN aerobic (FANAE) plus the standard BacT/Alert anaerobic (REGAN) bottles for culture of continuous ambulatory peritoneal dialysis (CAPD) fluid from patients with CAPD peritonitis. CAPD fluid (10 mL) was injected into each bottle, which was then monitored by the BacT/Alert instrument by using a 7-day protocol. Aerobic and anaerobic terminal subculture were performed on all bottles before they were classified as being culture negative. There were 181 effluents received that were suitable for analysis. Growth was detected in 76 (42%) effluents by at least one method. FANAE was the single best medium detecting 84/96 (88%) of all organisms whereas the FANAN and REGAN each detected 69/96 (72%). The combination of FANAE and REGAN bottles detected 92/96 (96%) isolates, which was significantly better than the FANAN or FANAE alone for isolate recovery (p < 0.001). The isolates that were missed by the FANAN but that were recovered by either FANAE or REGAN were all facultative anaerobes commonly detected in CAPD fluids. Terminal subculture revealed otherwise undetected pathogens in 3.9% of positive effluents, usually Pseudomonas aeruginosa. Based on our data, FANAE was the single best bottle for detection of CAPD peritonitis and, in combination with an anaerobic bottle, detected growth from the most effluents. FANAN alone could not substitute for the FANAE/REGAN combination. Although terminal subculture remains controversial, we recommend routine aerobic subculture to ensure that no P. aeruginosa isolates are missed. PMID- 10705049 TI - Comparison of several methods to determine methicillin-resistance in Staphylococcus aureus with focus on borderline strains. AB - We compared the performance of several phenotypic tests to detect methicillin resistant Staphylococcus aureus, with special focus on borderline strains. The reliability of the agar screen oxacillin and BBL Crystal tests was asserted for all methicillin-susceptible (n = 25), -resistant (n = 29) and borderline beta lactamase-hyperproducer (n = 10) strains. Whereas these tests failed to detect 4 of 5 rare borderline strains containing few cells with high-level methicillin resistance (i.e., a frequency of 10(-7)-10(-8)), a "two-temperature" disk diffusion method, performed simultaneously at 35 and 42 degrees C, detected all of such strains. PMID- 10705050 TI - Insufficient diagnostic accuracy of imported serological kits for Helicobacter pylori infection in Japanese population. AB - Although there are many reports of the high diagnostic accuracy of commercially available serologic kits for Helicobacter pylori infection in Western countries, they rarely has been investigated in oriental population. Accordingly we examined their usefulness in 492 Japanese patients with dyspeptic symptoms. Diagnostic accuracy of 4 imported serologic kits (HEL-p TEST, HM CAP, G.A.P IgG, Helico G2) was investigated using the (13)C-urea breath test as the gold standard. When intermediate results were excluded, the sensitivity, specificity and accuracy of these serologic tests ranged from 88.6% to 97.8%, 67.9% to 85.9%, and 87.9% to 91.4%, respectively, which were comparable with reported median accuracy in the Western population. However, there were many intermediate results in these tests, ranging from 5.3% to 23.0%. Their usefulness seemed to be limited in our patient population because of the large number of intermediate results. PMID- 10705051 TI - In vitro pharmacodynamic characteristics of flucytosine determined by time-kill methods. AB - Two Candida albicans isolates, three non-albicans Candida isolates (Candida glabrata, Candida krusei, and Candida tropicalis), and one Cryptococcus neoformans isolate were evaluated by time-kill methods to characterize the relationship of flucytosine concentrations to antifungal activity and the duration of the post-antifungal effect (PAE). Against Candida and Cryptococcusisolates, flucytosine at concentrations > 1 x MIC exhibited fungistatic ( 5 h), suggests lower daily dosing may possible without loss of antifungal efficacy. PMID- 10705052 TI - High prevalence of antibiotic resistance of common pathogenic bacteria in Taiwan. The Antibiotic Resistance Study Group of the Infectious Disease Society of the Republic of China. AB - We analyzed the antimicrobial susceptibilities of all clinical isolates of 14 common pathogenic bacteria recovered from patients in eight medical centers in Taiwan during 1995 and 1996. Susceptibility to commonly used antimicrobial agents was tested by the disk diffusion method as recommended by the National Committee for Clinical Laboratory Standards. Of the Staphylococcus aureus isolates, 59.3% and 62% were oxacillin-resistant in 1995 and 1996, respectively, whereas 63.2% of the coagulase-negative staphylococci isolates during the study period were oxacillin-resistant. The rate of penicillin-resistance among Streptococcus pneumoniae isolates was 39.7% in 1995 and 53.7% in 1996. Macrolide-resistance was found in 71.4%, 42.1%, and 46.7% of S. pneumoniae, beta-hemolytic streptococci, and viridans streptococci, respectively, in 1996. Less than 2% of the enterococcal isolates were vancomycin resistant, but 77% of them were gentamicin resistant. Resistance to gentamicin was also common in Enterobacteriaceae, Pseudomonas aeruginosa, and Acinetobacter baumannii. Various degrees of resistance to ampicillin, piperacillin, cephalosporins, aztreonam, and ciprofloxacin were detected in Enterobacteriaceae, P. aeruginosa, and A. baumannii. More than 55% of Haemophilus influenzae isolates were ampicillin resistant. In summary, resistance to many antimicrobial agents in various common pathogenic bacteria is very common in Taiwan. Our results implicate that antibiotic resistance in the developing countries need to be monitored closely. PMID- 10705053 TI - Screening and confirmatory testing for extended spectrum beta-lactamases (ESBL) in Escherichia coli, Klebsiella pneumoniae, and Klebsiella oxytoca clinical isolates. AB - Escherichia coli and Klebsiella spp. were screened for ESBL based on routine susceptibility testing results. Isolates with intermediate or resistant susceptibilities for extended spectrum cephalosporins or aztreonam were reported as probable ESBL producers. By using the NCCLS proposed ESBL confirmatory method, we tested 61 screen-positive isolates from 42 patients, 30 randomly selected susceptible isolates, and 12 isolates with previously characterized beta lactamases. Ceftazidime contributed to 97% of screen-positive isolates, whereas aztreonam added a single patient isolate. An ESBL was confirmed in 86% of K. pneumoniae, 100% of K. oxytoca, and 20% of E. coli screen-positive single patient isolates. None of the susceptible isolates were shown to produce ESBL. Based on these findings a comment regarding the presence of ESBL seems sufficient for Klebsiella spp. but confirmatory testing is indicated for E. coli. 0.25 microg/mL was used to indicate the presence of ESBL, the specificity of the assay increased to 100%. The NCCLS ESBL phenotypic confirmatory method was reproducible and accurate enough to be used in the clinical laboratory. PMID- 10705054 TI - Comparison of phenotypic methods to identify enterococci intrinsically resistant to vancomycin (VanC VRE). AB - Clinical laboratories must be able to differentiate between enterococci possessing acquired resistance to vancomycin (vanA and vanB genotypes) from those that are inherently resistant (vanC1 and vanC2/C3 genotypes). We compared several routine phenotypic tests to determine the species identity of clinical isolates of enterococci and a PCR assay for the van ligase genes was used to confirm identification of VanC VRE. The Vitek Gram Positive Identification card identified 53/60 (88%) Enterococcus faecalis and E. faecium isolates and 81/141 (57%) VanC VRE without additional testing. Another 32 of the VanC VRE required additional testing (e.g., motility and pigmentation) for correct identification. However, 7 of these 32 VanC VRE were nonmotile. The rapid ID 32 STREP strips identified 50/60 (83%) E. faecalis and E. faecium isolates and 102/141 (72%) VanC VRE. All E. faecalis and E. faecium isolates were nonmotile and did not acidify 1% methyl-alpha-D-glucopyranoside (MGP). Only 93/115 (81%) E. gallinarum and 21/26 (81%) E. casseliflavus/E. flavescens were motile but all 141 VanC VRE acidified MGP. MGP acidification can accurately differentiate VanC VRE from E. faecalis and E. faecium. Because some VanC VRE isolates are nonmotile, MGP acidification is preferred as a simple and less costly test for identification of these isolates. PMID- 10705055 TI - Comparison of the activity of two broad-spectrum cephalosporins tested against 2,299 strains of Pseudomonas aeruginosa isolated at 38 North American medical centers participating in the SENTRY Antimicrobial Surveillance Program, 1997 1998. AB - Pseudomonas aeruginosa is an important nosocomial pathogen. Resistance to certain beta-lactam antimicrobial agents among P. aeruginosa is increasing. The SENTRY Antimicrobial Surveillance Program was designed to employ a network of hospitals in the United States, Canada, Latin America, and Europe to monitor the predominant bacterial and fungal pathogens and antimicrobial susceptibility patterns associated with community-acquired and nosocomial bloodstream, respiratory tract, wound, and urinary tract infections. The purpose of this analysis of SENTRY results was to extract information on the current North American susceptibility pattern of P. aeruginosa for two antipseudomonal cephalosporins, ceftazidime, and cefepime. Clinical isolates were provided by 30 centers in the United States (grouped into five regions) and eight centers in Canada. Susceptibility testing was performed at a central reference laboratory by using broth microdilution methods and interpretive criteria specified by the National Committee for Clinical Laboratory Standards. Of the 34, 530 North American bacterial isolates tested during 1997 and 1998, 2299 (6.7%) were P. aeruginosa. There were no substantial differences in regional rates of P. aeruginosa susceptibility to ceftazidime (range 78.8-81.9%) or cefepime (range 80.0-83.4%) The percentage of resistant isolates among the 1784 United States isolates was 13.3% for ceftazidime versus 7.1% for cefepime (p < 0.05). It is essential to continue surveillance of the in vitro efficacy of these and other beta-lactam agents against P. aeruginosa because of the clinical importance of these safe and broad-spectrum cephems used alone or in combination in current clinical practice. PMID- 10705056 TI - Oral fusidic acid fails to eradicate methicillin-resistant Staphylococcus aureus colonization and results in emergence of fusidic acid-resistant strains. AB - Carriers of methicillin-resistant Staphylococcus aureus (MRSA) in hospital constitute a reservoir of infections and increase the risk of bacteremia and wound infection. In this prospective randomized trial, we tested the effectiveness of oral fusidic acid for eradication of MRSA colonization. From March 1997 through February 1998, patients with MRSA colonization in medical intensive care units in a large urban teaching hospital were randomly assigned to receive fusidic acid 500 mg q8h orally for 7 days or no anti-staphylococcal treatment. Twenty-three MRSA carriers were found during the study period and 16 were eligible for evaluation; six of them received fusidic acid. MRSA colonization was cleared in only two of the six patients with fusidic acid treatment, and later recurred in one of them. MRSA disappeared for 1, 2, 7, 7, and 8 weeks, respectively, in five of the 10 patients without treatment. MRSA persisted in the other five cases. Although all MRSA isolates found in the initial surveillance culture were susceptible to fusidic acid (MIC /= 256 microg/mL). Pulsed field gel electrophoresis pattern analysis showed that the resistant strains were genetically identical to the susceptible strains isolated from the same patient before fusidic acid treatment, in both cases. However, genetically distinct strains colonized in the same individual during follow-up were found in four out of 16 cases. We conclude that oral fusidic acid alone is not suitable for eradication of MRSA colonization, and may lead to the emergence of resistant strains. PMID- 10705057 TI - Rapid identification of clinical yeast isolates using the colorimetric AUXACOLOR system. AB - The AUXACOLOR colorimetric system (Sanofi Diagnostics Pasteur, Marnes-la Coquette, France) for the identification of clinical yeast isolates, was compared in its identification of 100 yeast strains to conventional identification methods. Of the 94 correctly identified isolates, 47% (n = 44) were identified by 24 h, and 100% (n = 94) were identified by 48 h. AUXACOLOR is a simple, rapid and accurate method for the identification of yeast pathogens. PMID- 10705058 TI - Biofilm production by Staphylococcus epidermidis isolates associated with catheter related bacteremia. AB - The mean biofilm production of 22 Staphylococcus epidermidis isolates associated with catheter related bacteremia was significantly higher than that of 32 nose isolates from healthy individuals. This difference was due to seven catheter related isolates. These findings do not show a clear association between biofilm production and virulence. PMID- 10705059 TI - Knowledge and attitudes of high school pupils towards children with special health care needs: an Israeli exploration. AB - One hundred seventy-one pupils of a high school in Holon (Israel) were questioned regarding their attitudes towards chronically disabled individuals who have special health-care needs. The level of pupils' knowledge concerning the etiology, symptoms and complications of these chronic conditions was approximately 72%; it did not increase in four years of study. Previous personal knowledge of a child or adolescent with a chronic disease or handicap influenced the understanding of that particular ailment. The pupils' attitude towards handicapped individuals become more positive and tolerant with increasing age (and class level). A correlation was found between the level of knowledge about special medical-health needs and tolerant attitudes toward chronic patients. Most of the pupils think that the following occupational and recreational interests for the disabled will help to increase their self-image and improve their feelings of acceptance: computers, music (playing and listening), sports and domestic pets. The pupils believe the topics of chronic diseases and patients should be studied as a regular part of the school curriculum and these subjects should be taught with the participation of both physicians and teachers. The pupils stressed the role of the teacher in handling children with the chronic disease. Only half of the pupils think that their class peers should also know about the chronic disease of this individual pupil. The information sources of pupils concerning chronic disease are mainly TV, newspapers, adolescent periodicals, books, physicians and nurses, and lastly, the school. Conclusions are presented concerning health promotion activities in the school. PMID- 10705060 TI - Coping with new treatments for cancer: a feasibility study of daily diary measures. AB - This article describes a study of the feasibility and value of using daily diary measures to assess coping, adjustment and symptoms in patients participating in phase I clinical trials of new anticancer drugs. Ten patients (six women, four men) with advanced metastatic cancer were studied during a four-week phase I trial. Measures of psychological well-being, mental adjustment and symptoms were determined prior to treatment, and participants also completed daily ratings of psychological coping responses, mood and symptoms. Completion rates for diaries were high, and the amount of missing data averaged only 3.2% per patient. Overall, the most frequently endorsed coping responses were 'acceptance' and 'positive reinterpretation and growth'. There were systematic variations in coping by seeking social support across the trial, with more frequent use during phases of hospitalisation. Idiosyncratic fluctuations in patterns of coping were also observed. Positive daily mood was greater among patients who coped by ignoring their condition, and was also correlated with fighting spirit. Daily symptoms were inversely associated with positive mood and with fighting spirit. It is concluded that the daily diary approach is feasible, and may help to increase understanding of the experience of patients taking part in experimental anticancer drug trials. PMID- 10705061 TI - Psychooncological provision in a general hospital: the Herford model. AB - Since October 1995 a Department of Psychooncology exists at the Herford Community Hospital. It was founded by a private foundation with the aim of supporting patients who suffer from cancer. The psychooncological section is an independent unit. This state of independence proved to be very useful and functional. The Herford Model is based on four compartments: psychosocial support for the patients; support for the patients' relations; the education of doctors and nurses; the evaluation of the activities of the team members, one music therapist and three psychologists. During the past 2.5 years 846 patients have had contact with the Department of Psychooncology. The mean age was about 62 years within a range from 15 to 92 years. Of the patients, 48% were men and 52% were women. The maximum number of meetings was 49: 29% of the patients had between four and eight therapeutical meetings, 12% had more than nine sessions. It is the aim of this report to show how the section is organized. The daily work and the rate of provision is shown and an outlook of the team's work in the future is given. PMID- 10705062 TI - Pre- and postoperative information needs. AB - The purpose of this study was to investigate patients' need for pre- and postoperative information. Fifty patients admitted to open cholecystectomy were included in the study, 37 women and 13 men. Their median age was 49.5 years, ranging 17-76. The patients answered one questionnaire both at admission and at discharge. In the questionnaire, 48 statements had to be answered on a five point, Likert scale. Our data show that patients admitted for cholecystectomy want a lot of information both at admission and at discharge. The most requested information was related to anxiety-creating factors such as pain and post operative symptoms after surgery. Thirty per cent of the patients wanted both written and verbal information. This result focuses on the need to develop and share with the patient both written and verbal information. PMID- 10705063 TI - Antihypertensive treatment and patient autonomy--the follow-up appointment as a resource for care. AB - Since hypertension is a chronic condition which generally requires long-term commitment to pharmacological therapy as well as alterations of patient lifestyle, the patient-physician communication in the clinical setting is an important determinant of the quality of care and health outcome. The aim of the present study was to explore the structure and content of the communication between the patient and the physician, and the process of decision-making at a routine follow-up appointment for hypertension. The study was based on 51 audio recordings of authentic consultations. Most patients had a passive role in the consultations, and initiated few topics of conversation. The few topics that the patients initiated were usually not about hypertension. Patients' questions about medication mainly referred to unwanted effects of the drugs. Little time was invested in discussing risks related to hypertension. A collaborative shared decision-making was seldom observed in the consultations. PMID- 10705064 TI - Patient satisfaction with the information provided at a psychiatric emergency unit. AB - OBJECTIVE: Evaluation of patient satisfaction with information at a psychiatric emergency unit. DESIGN: Patient survey. SETTING: Psychiatric patients assessed information provided by staff on illness, symptoms, treatment alternatives, treatment design, medication, time schedule for treatment and the expected therapeutic response. PARTICIPANTS: The sample included 100 subjects (63% response rate). OUTCOME MEASURE: Patient satisfaction. RESULTS: 59% were women. Mean age was 43 years. 87% were Swedish. 30% had psychotic, 35% bipolar and 35% anxiety disorders. 87% were admitted voluntarily. Almost 80% were satisfied with the patient-staff relationship. Questions on information, except medication, scored low. Patients with non-psychotic disorder were more satisfied with information on symptoms, treatment alternatives and treatment design, and voluntary patients with information about medication. Patients born in Sweden and voluntary patients awarded influence on treatment planning higher scores. CONCLUSIONS: Psychiatric patients requiring emergency care did understand information. The staff provided satisfactory information only when knowledgeable. PMID- 10705065 TI - Physician expressions of uncertainty during patient encounters. AB - Uncertainty is inherent in clinical medicine and may contribute to variability in physician practice patterns, patient satisfaction, and exchange of information. However, research on physician disclosure of uncertainty to patients is sparse. We measured the frequency of physician expressions of uncertainty to patients using audiotapes of visits to 43 physicians by 216 continuity patients in a university-affiliated general medicine clinic. We also analyzed the audiotapes using Roter Interaction Analysis. Physicians completed Gerrity's Physician's Reaction to Uncertainty scale and patients completed the Kranz Health Opinion Survey and a standardized satisfaction questionnaire. Physicians made verbal expressions of uncertainty in 71% of clinic visits. Physicians with greater self rated reluctance to disclose uncertainty to patients made fewer expressions. Physicians who made more uncertainty expressions also used more positive talk and partnership building, and gave more information to patients. Physicians also expressed more uncertainty to patients with more education, greater desire for information, and more questions. Physician uncertainty expression were associated with greater patient satisfaction, but not independently of other physician verbal behaviors that were also associated with satisfaction. PMID- 10705066 TI - Missed opportunities to impact fast response to AMI symptoms. AB - The potential for reducing cardiovascular disease mortality rates lies both in prevention and treatment. The earlier treatment is administered, the greater the benefit. Thus, duration of time from onset of symptoms of acute myocardial infarction to administration of treatment is important. One major factor contributing to failure to receive efficacious therapy is the delay time from acute myocardial infarction (AMI) symptom onset to hospital arrival. This paper examines the relationship of several factors with regard to intentions to seek care promptly for symptoms of AMI. A random-digit dialed telephone survey (n = 1294) was conducted in 20 communities located in 10 states. People who said they would wait until they were very sure that symptoms were a heart attack were older, reported their insurance did not pay for ambulance services, and reported less confidence in knowing signs and symptoms in themselves. When acknowledging symptoms of a heart attack, African-Americans and people with more than a high school education reported intention to act quickly. No measures of personal health history, nor interaction with primary care physicians or cardiologists were significantly related to intention to act fast. The study confirms the importance of attribution and perceived self-confidence in symptom recognition in care seeking. The lack of significant role of health history (i.e. those with chronic conditions or risk factors) and clinician contact highlights missed opportunities for health care providers to educate and encourage patients about their risk and appropriate action. PMID- 10705067 TI - Concepts of mental health--a survey of attendees at a mental health promotion conference. AB - This survey of attendees at a Mental Health Promotion Conference in Hull, Yorkshire, England in the summer of 1996 was undertaken in part fulfilment of an MSc in Health Promotion. Of the 120 delegates, 99 attended the conference. There were 77 completed questionnaires. The most outstanding result of the survey was the very strong agreement among respondents that sense of self-worth is the most important attribute of being mentally healthy. The respondents were also strongly in favour of enablement strategies but on the whole envisioned these as having primarily an educational rather than a societal focus. About a third of the respondents tended to conceptualise mental health promotion within a clinical framework. The survey demonstrates the need to strengthen concepts of positive mental health and for mental health professionals to identify and work more closely with non-health workers in the promotion of positive mental health. PMID- 10705068 TI - Pilot study of a home-based asthma health education program. AB - OBJECTIVE: Caring for a child with asthma can affect the parent's coping and well being and coping strategies. This study examined the influence of a home-based asthma health education program on parental coping and quality of life. DESIGN: Randomized controlled non-blinded clinical trial. SETTING: Northern community pediatrician's office. PATIENTS: Families whose children, under the age of 11, had chronic stable asthma, and who presented to the pediatrician's office for continuing care; those with an acute exacerbation of asthma were excluded. INTERVENTIONS: Families were randomly assigned to receive either a single two hour, standardized home-based asthma health education session or a booklet representing conventional care. MAIN OUTCOME MEASURES: One and three-months following the intervention, assessments were obtained for coping measured by Hymovich's Parent Perception Inventory (PPI), quality of life measured by the Pediatric Asthma Caregiver Quality of Life Questionnaire (PACQLQ) and change in asthma measured by the Caregiver Perception of Change (CPC) survey. RESULTS: Forty families were recruited and randomized; baseline characteristics were similar between groups. At the final follow-up, reduction in parental need for asthma information (p = 0.04), reduction in parental concerns (p = 0.02) and increased use of coping strategies (p = 0.04) were observed in the home-based care group. Improvement was noted in the parent's perception of their child's asthma in the home-based asthma education group (p = 0.01). Quality of life as measured by the PACQLQ remained unchanged over the intervention period (p > 0.05). CONCLUSIONS: These results suggest the use of a one-time, flexible, home based intervention to assist families caring for children with asthma should be considered and appears effective. PMID- 10705069 TI - Effect of high ambient temperature on contamination and physical stability of one liter ready-to-hang enteral delivery systems. AB - The effect of high ambient temperature on the physical stability and bacterial contamination of 1-L, prefilled, closed enteral feeding systems was examined under simulated clinical conditions. One hundred Jevity Ready-to-Hang enteral feeding systems (Abbott Park, IL, USA) were placed in a 37 degrees C incubator for 24 h. The Ready-to-Hang formula containers were visually inspected at 0 and 24 h. Formula samples were collected from the containers at 24 h and plated on trypticase soy agar. Two samples had insignificant bacterial growth of one colony forming unit per milliliter that was not demonstrated in repeat culture. No growth was observed for any other sample. Additional samples collected from the two apparently contaminated delivery sets showed no growth. No set showed signs of formula instability, such as coagulation, clumping, or curdling. These findings suggest that, even at a high ambient temperature of 37 degrees C, the risk of bacterial contamination or compromised physical integrity is very low with the use of 1-L, prefilled, closed enteral feeding systems. PMID- 10705070 TI - Vasodilatory actions of the dietary peptide carnosine. AB - The objective of this study was to test the hypothesis that the dietary dipeptide carnosine (beta-alanine-L-histidine) causes direct decreases in arterial tone. Isolated descending thoracic aortic rings from male Sprague-Dawley rats were used for all studies. Preconstriction of vessels was accomplished with phenylephrine. Carnosine (0.625-20 mM) produced dose-dependent vascular relaxation (P < 0.05) that was independent of endothelium. The constituent amino acid L-histidine did not produce any significant relaxation over the same dose range, whereas beta alanine actually produced dose-dependent vasoconstriction (P < 0.05). The soluble guanylate cyclase inhibitor methylene blue (10(-5) M) significantly decreased the relaxation produced by carnosine (P < 0.05). Measurement of cyclic GMP in the presence and absence of methylene blue after carnosine and phenylephrine exposure was also done. Methylene blue 10(-5) M resulted in a decrease in cyclic GMP levels from 65.3 +/- 15.6 fmol/mg protein to 8.6 +/- 0.9 fmol/mg of protein (P = 0.001). We conclude that carnosine produces relaxation of isolated rat aorta independent of endothelium. The effect of carnosine is at least in part mediated via cyclic GMP production and is not reproduced by its constituent amino acids, L histidine and beta-alanine. PMID- 10705071 TI - Energy metabolism and substrate oxidation in patients with Crohn's disease. AB - Weight loss and malnutrition are common features in patients with Crohn's disease. This study was designed to evaluate diet-induced thermogenesis and substrate oxidation in patients with Crohn's disease. Twenty-three patients (17 women, 6 men; age 34 +/- 2 y) and 17 healthy control subjects (13 women, 4 men; age 36 +/- 3 y) were studied. Resting energy expenditure and fasting substrate oxidation were measured by indirect calorimetry in the morning after an overnight fast. After a standard homogenized test meal (10 kcal/kg), indirect calorimetry was performed every 30 min for 3 h to measure the diet-induced thermogenesis and the postprandial substrate oxidation. In the fasting state, resting energy expenditure was significantly higher in patients than in control subjects (1433 +/- 43 versus 1279 +/- 53 kcal/24 h). Lipid oxidation was higher in patients with Crohn's disease than in control subjects (1.17 +/- 0. 07 versus 0.61 +/- 0.11 mg. kg(-1). min(-1), P < 0.01). Postprandially, diet-induced thermogenesis was significantly lower in patients with Crohn's disease than in control subjects (4.6% +/- 0.5 versus 6.3% +/- 0.5 of energy intake, P < 0.01). Lipid oxidation was significantly higher in patients with Crohn's disease than in control subjects (0.78 +/- 0.05 versus 0.56 +/- 0.08 mg. kg(-1). min(-1), P < 0.05), and glucose oxidation was lower in patients with Crohn's disease than in control subjects. In patients with Crohn's disease, lipid oxidation positively correlates with the disease activity evaluated by the Crohn's Disease Activity Index (r = 0.48, P150), fasting and postprandial lipid oxidation was significantly higher than in patients with inactive Crohn's disease (P < 0.05). In conclusion, patients with Crohn's disease have increased fat oxidation, which correlates with disease activity and this may explain the reduced fat stores in patients with Crohn's disease. PMID- 10705072 TI - The role of exercise in the treatment of obesity. AB - The prevalence of obesity in the USA has increased dramatically in the past decade. This foreshadows an increase in the rates of morbidity and mortality from obesity-related diseases and increases in the number of individuals undergoing weight-loss therapy. Although exercise has long been recommended for inclusion in such therapy, the present review has found that it has had little or no effect on weight loss per se when the exercise is limited to the typically prescribed 3-5 h/wk of moderate or vigorous activity. However, further review has shown that exercise helps to preserve and at times even increase fat-free mass during weight loss. At the same time, fat loss is generally increased. Neither type nor amount of exercise appears to have much effect during treatment, with the possible exception of resistance training, which is associated with the best outcome for fat-free mass. The most important role of exercise, however, is in the maintenance of the weight loss. In this respect, the volume of exercise seems to be important because several lines of evidence have indicated that exercise must expend roughly 2500 kcal/wk to maintain weight loss. Studies of weight maintenance, however, have generally not included randomized controls; thus, further research is required to solidify these conclusions. PMID- 10705073 TI - The association between specific nutritional antioxidants and manifestation of colorectal cancer. AB - Determination of specific antioxidants for examination of oxidative balance and immune responses may be of value in the early diagnosis of colorectal cancer. In the present report, we investigated urinary excretion of zinc, copper, and neopterin and serum levels of vitamins A, C, and E in 30 patients (age = 64 +/- 12 y) with colorectal cancer at the time of diagnosis and in 30 control subjects (age = 61 +/- 11 y) with benign disorders not associated with a systemic inflammatory response. Urinary excretion of zinc, copper, and neopterin was significantly elevated, and serum concentration of vitamin A was decreased in patients with colorectal cancer; these changes are characteristics of systemic immune activation. These phenomena may be of use for the detection of tumor progression and immune response to neoplasm. PMID- 10705074 TI - Metabolic abnormalities associated with skeletal myopathy in severe anorexia nervosa. AB - The aim of this study was to characterize the metabolic disturbance associated with the skeletal myopathy resulting from extreme weight loss in anorexia nervosa. Muscle function was examined in eight female patients with severe (40%) weight loss due to anorexia nervosa and histologically confirmed myopathy. A wide range of biochemical and hematologic investigations were carried out, including serum enzymes and the response of plasma lactate to ischemic exercise of forearm muscles. All patients showed proximal muscular weakness. A diminished lactate response to ischemic exercise was a consistent finding, and a reduction of serum carnosinase activity was also found. There were no other consistent biochemical or hematologic abnormalities apart from lymphopenia of no clinical consequence. These findings contribute to our understanding of severe protein-energy malnutrition on the musculoskeletal system. The resulting disorder is a metabolic myopathy from which the patients recover rapidly as their nutrition improves. Although the patients admitted to a variety of abnormal eating behaviors, no correlation was found between a specific type of abnormal eating behavior and subsequent biochemical abnormalities. Reinstating appropriate eating behavior will treat the myopathy. PMID- 10705075 TI - Influence of short-term protein malnutrition of mice on the phenotype and costimulatory signals of lymphocytes from spleen and Peyer's patches. AB - The objective of this study was to investigate the impact of short-term protein malnutrition (PM) on immunoglobulin A (IgA) production and on the number and phenotype of lymphocytes in Peyer's patches (PP) and in the spleen. Balb/c mice were fed for 4, 7, or 10 d with a protein-deficient diet (0.1% protein). We determined B lymphocytes (CD40(+)), T lymphocytes (CD3(+)), T-helper (CD4(+)), and T-suppressor (CD8(+)) cells and the expression of costimulatory signals B7.1 (CD80) and B7.2 (CD86) on B cells and their counter receptors CD28 and CTLA-4 on T cells by fluorescence-activated cell-sorting analysis. Luminal IgA concentration in the small intestine was determined by an enzyme-linked immunosorbent assay. Four days of PM caused a significant reduction in the number of mononuclear cells in the spleen (5.6 x 10(7) +/- 1 x 10(7) versus 2. 4 x 10(7) +/- 0.5 x 10(7), P < 0.001) and the PP (13 x 10(6) +/- 3 x 10(6) versus 8.6 x 10(6) +/- 2 x 10(6), P < 0.01). There was a relative increase of T cells in the spleen and a relative increase of B cells in the PP. Luminal IgA content of small intestine was significantly reduced after 4 d of PM (242 +/- 55 microg versus 173 +/- 39 microg, P < 0.05) and remained at about this level until day 10 of PM. Four days after PM, the costimulatory signals B7.1 and B7. 2 on B cells were upregulated in the PP but markedly downregulated in the spleen, which was inversely related to the expression of the counter receptor CD28 on T-helper cells. We conclude that short-term PM increases the activation of B cells in the PP but reduces the relative number and activation state of splenic B cells. Only 4 d of PM caused a systemic and intestinal immunodepression, as indicated by a markedly decreased content of mononuclear cells in the PP and the spleen. PMID- 10705076 TI - Fatty-acid synthase and human cancer: new perspectives on its role in tumor biology. AB - This review documents the changing perspectives on the function of fatty-acid synthase and fatty-acid synthesis in human tumor biology. With the recent discovery that human cancer cells express high levels of fatty-acid synthase and undergo significant endogenous fatty-acid synthesis, our understanding of the role of fatty acids in tumor biology is expanding. Once considered largely an anabolic-energy-storage pathway, fatty-acid synthesis is now associated with clinically aggressive tumor behavior and tumor-cell growth and survival and has become a novel target pathway for chemotherapy development. These findings will ultimately enhance our understanding of fatty acids in tumor biology and may provide new diagnostic and therapeutic moieties for patient care. PMID- 10705077 TI - Iron and copper in human milk. AB - The reported concentrations of iron and copper in breast milk show a wide variation. Research published over the past 50 y has reported median values of 0.47 and 0.32 mg/L for iron and copper, respectively. The levels of both metals decrease with the progress of lactation. The calculated iron-to-copper ratio of reported means differs from 0.25 to 6.29 (median = 1.18). Maternal constitutional variables such as undernutrition, iron and copper body reserves, stage of lactation, adolescent motherhood, gestation length, and infection and environmental variables such as iron and copper dietary intake, in addition to supplementation, smoking, vegetarianism, and prolonged use of hormonal contraceptives before and during lactation did not consistently affect the concentrations of iron and copper in breast milk. Extreme cases of either low or high levels of body metal availability or altered metabolism due to chelating therapy or illnesses such as Wilson's disease and infections did not affect metal transfer from blood serum to breast milk. There is no clinical or scientific support for the need of extra iron or copper, besides the quantities provided by milk in the full-term breast-fed infant, at least during the first 6 mo. PMID- 10705079 TI - Nutrient-gene interactions during intrauterine life and lactation. PMID- 10705080 TI - Breakfast cereal, caffeinated coffee, mood, and cognition. PMID- 10705078 TI - Insulin reduces leucine oxidation and improves net leucine retention in parenterally fed humans. AB - Protein metabolism during parenteral feeding was measured with and without euglycemic hyperinsulinemic clamping in five healthy human subjects using the primed continuous infusion of [(13)C]leucine (LEU) tracer methodology. All subjects underwent two periods of protein measurement. Subjects were randomized to have the clamp first or second. Two subjects had the clamp first, and they participated in another study in which the measurement period after the clamp was increased by another 2 h. Insulin reduced LEU oxidation (from 26 to 22 microM. kg(-1). h(-1); P < 0.05) and improved net LEU balance (from 3 to 7 microM. kg( 1). h(-1); P < 0. 05). The order of the clamp influenced the effects of insulin. With clamping performed second, insulin reduced LEU flux (from 152 to 134 microM. kg(-1). h(-1); P < 0.01), endogenous LEU rate of appearance (Ra; from 125 to 107 microM. kg(-1). h(-1); P < 0.01), and non-oxidative LEU disappearance (NOLD; from 128 to 113 microM. kg(-1). h(-1); P < 0.01). With clamping performed first, NOLD decreased after insulin was stopped (from 129 to 121 microM. kg(-1). h(-1); P < 0.05), but no change was seen in the flux and LEU Ra, despite the return of plasma insulin and amino acid concentrations to basal levels. The reduction in NOLD was accentuated with time and did not reach plateau even after 6 h and indicated a prolonged carry-over effect for NOLD and Ra. This effect was not seen for leucine oxidation. PMID- 10705081 TI - Transferrin modulates tumor necrosis factor-alpha secretion by cultured human mononuclear cells: influence of iron status. PMID- 10705082 TI - Theophile Roussel and the elimination of pellagra from 19th century France. PMID- 10705083 TI - Light protection during parenteral nutrition infusion: is it really necessary? PMID- 10705084 TI - Pushing the envelope of nutrition support: complementary therapies. PMID- 10705085 TI - A trawl through the current nutrition literature. PMID- 10705086 TI - Ameloblastic fibroma and related lesions: current pathologic concept. AB - Ameloblastic fibroma (AF) is a true mixed tumor, in which the epithelial and the ectomesenchymal elements are neoplastic. There are two rare variants of AF; granular cell AF and peripheral AF. Ameloblastic fibrosarcoma is a rare tumor, and is regarded as the malignant counterpart of the benign AF. Recent immunohistochemical study using MIB-1 shows labelling indices in the mesenchymal component of the recurrent AF and ameloblastic fibrosarcoma are quite high, in contrast with the conventional AF. Ameloblastic fibrodentinoma is a histologic variant of AF in which dentin or dentinoid tissue has formed, but there is no eveidence that ameloblastic fibrodentinoma exhibit a different biologic behavior than ordinary AF. Ameloblastic fibro-odontoma is a lesion similar to AF, but also showing inductive changes that lead to the formation of both dentin and enamel. Some lesions diagnosed as ameloblastic fibro-odontoma are probably developing odontoma, but the others should not be considered as hamartomatous in nature, since there are rare cases of ameloblastic fibro-odontoma showing true neoplastic behavior, and since the existence of malignant variant is evident. In revised WHO's classification of odontogenic tumors, the terms "ameloblastic fibrodentinoma" and "dentinoma" are used synonymously, however, there are histologic difference between several cases reported previously as "dentinoma" and ameloblastic fibrodentinoma. PMID- 10705087 TI - Prognostic value of apoptotic index in cutaneous basal cell carcinomas of head and neck. AB - Basal cell carcinoma (BCC) is the most common type of human cancer, often locally invasive, and following a benign clinical course. However, a proportion of BCCs do recur after treatment, causing extensive local tissue destruction, seldom metastasizing. Morphological methods to unequivocally distinguish the aggressive forms of these tumors (BCC2) from the ordinary ones (BCC1) have so far been lacking. Apoptosis, or programmed cell death, is thought to be important for the death of tumor cells in various stages of carcinogenesis. We analyzed the extent of apoptosis in BCCs of head and neck in a morphological, morphometric, and electron-microscopic study, to estabilish on a retrospective basis, the relative frequency of recurrence of tumors showing different apoptotic rates. We found that BCC1 showed lower apoptotic index (AI) than BCC2 [BCC1: AI from 2.03 to 10.45% (mean value: 5.98%) BCC2: AI from 21. 91 up to 43.82% (mean value: 39.82%)]. The morphometric analysis of both BCC1 and BCC2 revealed significant differences between the values concerning nuclear area, length, perimeter, and roundness of the apoptotic cells with respect to the 'viable' neoplastic cells. Electron-microscopy confirmed that the features of morphological apoptotic cells were characteristic of programmed cell death. We hypothesized that low apoptotic rates in BCC1 could be indicative of a good prognosis. In fact, this corresponded to an 'expansive' but not still invasive neoplastic state. In this phase, however, the tumor cells may constitute the target for genetic changes triggered by enviromental physical or chemical mutagenic agents, such as UV rays. BCC2, then, could be the result of newly selected mutated neoplastic cellular clones, with more aggressive biological behavior. The high apoptotic level found in BCC2 could thus be used as an indirect alarm signal from pathologists. This hypothesis seems to be supported by most of the current data in the literature and by the clinical outcome of BCC2 of our series. In our opinion, routine evaluation of apoptosis in BCCs could be proposed to facilitate their sub-classification, contributing toward the evaluation of the prospective outcome of the individual patients. PMID- 10705088 TI - Up-regulation of Fas ligand and down-regulation of Fas expression in oral carcinogenesis. AB - An important molecule involved in delivering the death signal that initiates apoptosis is called Fas, or Apo-1, which sits on the cell surface. When another molecule called the Fas ligand (FasL) binds to it, Fas triggers a series of events inside the cell that leads to apoptosis. In order to investigate the mechanism of immune escape and the expression of Fas and FasL in oral premalignant lesions (OPLs) and oral squamous cell carcinomas (OSCCs), a total of 64 samples were evaluated by an immunohistochemical method using a labelled streptavidin-biotin assay. These samples comprised nine hyperkeratotic and 24 oral premalignant lesions (nine of mild, moderate, and six of severe dysplastic lesions), and 24 OSCCs, together with seven healthy controls. The results demonstrated that the majority of invasive OSCCs showed down-regulation of Fas expression but up-regulation of FasL expression. These phenomena were also detected in OPLs. The results indicate that the expression of Fas and FasL is involved in oral carcinogenesis and this may be a mechanism by which the cancer cells evade the host immune assault. Perhaps, in future, Fas/FasL system may be used as a prognostic biomarker in predicting the behavior of oral premalignant lesions. PMID- 10705090 TI - Low incidence of H-, K- and N-ras oncogene mutations in cytological specimens of laryngeal tumours. AB - Laryngeal cancer is a rare type of neoplasia, constituting approximately 2% of all human cancers. Mutations of the ras gene family is one of the main activating mechanisms in human cancer. Their involvement in head and neck cancer has been mainly demonstrated at the level of the overexpression whereas ras mutations in these cancers are rare in the Western world. In the present study we explored the incidence of codon 12-point mutation in the H-, K- and N-ras genes, in 41 laryngeal cytological specimens. These specimens corresponded to 19 benign and 22 malignant lesions of the larynx. Only two specimens carried a codon 12-point mutation in the K-ras gene (4.8%) while no mutation was detected in the H- and N ras genes. K-ras mutations were detected in one benign and one malignant specimen. These results indicate low incidence of ras oncogene mutations in laryngeal cytological specimens. PMID- 10705089 TI - Effects of an anti-angiogenic agent, TNP-470, on the growth of oral squamous cell carcinomas. AB - Anti-tumor effects of an anti-angiogenic agent, TNP-470, on the growth of oral squamous cell carcinomas (SCCs) were evaluated in vivo and in vitro. The growth of an oral SCC cell line, HSC-2, inoculated subcutaneously in severe combined immuno-deficiency (SCID) mice was inhibited in a dose-dependent manner by the treatment with this agent. A reduction of microvessels surrounding tumor tissues treated with TNP-470 was observed by immunohistochemistry. Significant side effects were not observed except for weight loss during the period of treatment with high dose (50 mg/kg) of TNP-470. The direct effects of TNP-470 on oral SCC cell lines were also evaluated in culture. The growth of all eight SCC cell lines tested was inhibited by TNP-470, but the sensitivity of the oral SCC cell lines to TNP-470 was about 1700 times less than that of endothelial cells. These results suggest that TNP-470 inhibits the growth of oral SCC by anti-angiogenic activities and that it may be effective as a new therapy of oral cancer. PMID- 10705091 TI - Larynx cancer in Slovakia and the role of anatomical subsites. AB - During the time period 1968-92, 6958 laryngeal cancers (6602 in men and 356 in women) were diagnosed in Slovakia and notified to the National Cancer Registry. We analysed long-term trends in incidence, mortality and survival. Mortality and incidence rates in Slovakian men rose rapidly until 1980 and more slowly subsequently. The mortality-to-incidence ratio initially was 40% and increased to 70% in the period 1985-88. A log-linear model showed that the more recent generations experienced the slowest increase in incidence. Incidence and mortality rates in women remained stable and did not exceed 1 per 100,000. The 5 year survival probability from invasive larynx cancer was 47%. Survival rates had shown no particular trend by year and age at diagnosis. The main finding was that 5-year survival from supraglottis cancer is 20% poorer than survival from glottis cancer. Supraglottis is the prevalent larynx subsite in countries with high larynx, oropharynx and oesophagus incidence rates. This supports the hypothesis that supraglottis cancer is more strongly linked to a synergistic effect of smoking and alcohol than glottis cancer. PMID- 10705092 TI - Risk of second primary cancer following treatment of squamous cell carcinoma of the lower lip. AB - The risk of second primary cancers (excluding skin cancers) was evaluated among 56 patients who underwent treatment for a squamous cell carcinoma of the lower lip. The mean follow-up was 5.5 years. Ten patients (17.8%) developed at least one new primary cancer. The prevalence of second primary cancers within the respiratory and upper digestive tract, and elsewhere in the body, was 19.4 and 12.9 per 1000 person-years of follow-up, respectively. Patients were at risk for a second primary cancer at a steady rate of 2.7% per year during at least 5 years. PMID- 10705093 TI - The experimental evaluation of an oral cancer information leaflet. AB - The aims of our series of studies were: (1) to prepare an information leaflet about oral cancer for the general public; (2) to design a questionnaire to assess the level of knowledge about oral cancer; and (3) to evaluate the newly prepared leaflet. A study was conducted for each aim. For study 1, multiple drafts of oral cancer leaflets were distributed for comment and editing. For study 2, a large number of statements (100) about oral cancer were compiled and judged. Redundant items were withdrawn. The final version was administered to criterion groups (students, dentists and the public) to assess reliability and validity. For study 3, a multigroup pre- and post-test randomised design (comprising an experimental and two control groups) was adopted to evaluate knowledge improvement from exposure to the oral cancer leaflet. The participants in each study were as follows: in study 1, questions were judged by the authors, colleagues (n=2) and a non-patient group (n=10); in study 2, undergraduate psychology students (n=41), medical students (n=60), dental students (n=82), members of the public (n=54) and general dental practitioners (n=22) were respondents for the reliability/validity assessment; and in study 3, the dental students and members of the public from the previous study participated. The measure was a true/false 36-item questionnaire with additional demographic questions. In study 1 the Flesch readability index for the leaflet equalled approximately 80, i.e. 'fairly easy'. In study 2 the Kuder-Richardson 20 reliability coefficient of the questionnaire equalled 0.76. Criterion validity was confirmed with general dental practitioners scoring greater knowledge than members of the public (p<0.05). In study 3 participants who had access to the leaflet showed a significant increase in knowledge at post-test in comparison to pre-test. Control group results were supportive of a positive effect from leaflet exposure. In conclusion, the design of a health information leaflet, the assessment of reliability and validity of an associated knowledge questionnaire and evaluation of the influence of such a leaflet should be conducted in a series of planned steps. This approach can be used for other health-related issues which require dissemination of information. Three studies have demonstrated the successful application of this design and evaluation model to the area of oral cancer education. PMID- 10705094 TI - Prognostic significance of Ki-67 (MIB1), PCNA and p53 in cancer of the oropharynx and oral cavity. AB - Up to now results concerning the prognostic value of tumor proliferation markers in squamous cell head and neck carcinoma have been equivocal. Beside biological reasons, different treatment modalities are hypothetically responsible for contradictory findings. The aim of this study was to investigate the relationship between proliferative capacity, represented by the immunohistochemical labeling index of proliferation markers Ki-67, PCNA and p53 status, and treatment failure in a matched-pair study design of recurrent and non-recurrent carcinoma initially treated with primary surgery combined with curative post-operative radiation. From a group of 239 patients with T1-T3 carcinoma of the oropharynx or oral cavity, 28 patients with recurrent disease were selected and matched with 28 patients with non-recurrent disease regarding stage and location of tumor as well as age and therapy. All patients received primary surgery combined with post operative radiation. Immunohistochemistry determined the p53 status and the PCNA and MIB1 (Ki-67) labeling index. The Ki-67 labeling index was significantly (p=0.032) higher in tumors from patients suffering from treatment failure (mean=59. 1%) than in non-failures (mean=50.5%). Carcinoma with a Ki-67 (MIB1) labeling index above the median (53.7%) of the general study population showed a mean time to relapse of 45 months (n=25), whereas mean time-to-relapse was 61.7 months for those cases (n=31) below the median of the general study population (p=0.029). The PCNA labeling index did not correlate significantly with tumor recurrence (mean=50.2% for treatment failures, 45% for non-failures, p=0.31), nor with time-to-relapse (p=0.26). Forty-six percent of tumors showed p53 over expression. However, there was no significant correlation between p53 over expression and tumor recurrence or time-to-relapse. We present the largest series of oropharyngeal and oral cavity carcinoma investigated by immunohistochemistry in a controlled study. We conclude that a high Ki-67 labeling index is an indicator for treatment failure in these patients. Like other investigations for different head and neck subsites, we found no relationship between p53 or PCNA status and tumor prognosis. Our data, obtained from a group of patients treated with a combination of surgery and post-operative irradiation, show that for squamous cell carcinoma of the oropharynx and oral cavity the detection of Ki-67 is an unfavorable prognostic factor. PMID- 10705095 TI - Surfing prognostic factors in head and neck cancer at the millennium. AB - The ability to reliably predict cancer outcome could tailor therapy to the aggressiveness of the tumour to achieve the best results in terms of loco regional control, overall survival and quality of life. Retrospective and prospective clinical trials involving large series of patients have validated some predictive clinical and pathological factors, whereas the utility of many other prognostic factors has not been established. This has led to some confusion in clinical practice. In order to clarify the significance, role and cost of these prognostic factors we carried out a Medline search of all papers published between 1993 and 1998 concerning the reliability and cost of markers with prognostic significance, in head and neck squamous cell carcinoma, and assessed the results according to a number of criteria relating to reliability and cost. Regarding reliability we classified prognostic factors into: (1) those with a proven significance based on the fact that they were unanimously reported as having an independent statistical correlation with outcome and prognosis; and (2) those for which results were not unanimous, and which significance is still controversial. Cost analysis showed a substantial difference between validated tests which are of low cost and experimental tests which are expensive. Based on these data regarding both the reliability and cost of each prognostic factor, we propose guidelines for their use in clinical practice in the year 2000. PMID- 10705096 TI - Preoperative concurrent chemoradiotherapy plus radical surgery for advanced squamous cell carcinoma of the oral cavity: an analysis of long-term results. AB - Locoregionally advanced squamous cell carcinomas of the head and neck continue to be a major clinical problem. We demonstrated in 1996 that preoperative concurrent cisplatin- or carboplatin-based chemotherapy and radiotherapy plus radical surgery in advanced oral cancer had minimal toxicity, had high clinical tumor response rates, was well tolerated and produced impressive complete response rates and a high 5-year survival rate. The purpose of the present study was the long-term follow-up of this treatment regimen for advanced oral carcinoma. Forty eight patients with squamous cell carcinoma of the oral cavity (including soft palate) were treated preoperatively with cisplatin- or carboplatin-based chemotherapy in combination with simultaneous irradiation to a target volume of 40 Gy, and 2-6 weeks later underwent curative surgery. All patients with advanced Stage II (n=7), Stage III (n=22) and Stage IV (n=19) were treated and followed for an average of 7.2 years (range: 61-144 months). The overall actuarial survival of all patients was 81.3% at 5 years and also at 10 years. Progression free survival at both 5 and 10 years was 84.8% for all patients, and 85.7% for Stage II, 90.0% for Stage III, and 78.9% for Stage IV patients. Progression-free survival rates according to the histopathologic regression grade of primary tumor following preoperative chemoradiotherapy at 10 years were 40. 0% for Grade IIa, 88.9% for Grade IIb, 100% for Grade III, and 87.5% for Grade IV. Patients who achieved good responses histopathologically (Grades IIb, III, IV) had superior survival rates in comparison to patients with extensive residual tumor (Grade IIa) in surgically resected specimens (p=0.0012). A better histologic regression grade was also associated with a higher survival rate even in the long-term analysis. This treatment regimen for advanced oral cancer produced high clinical and pathologic complete response and survival rates with an acceptable acute toxicity profile and lack of late therapeutic complications. The long-term follow up showed gratifying results even for advanced oral cancers without a substantial increase in distant metastasis and second primary malignancy. PMID- 10705097 TI - Brachytherapy using gold-198 foils in treatment of mouth tumors: case report. AB - The authors presents a clinical case treated with brachytherapy performed with special mold of gold-198 disc, with the purpose of evaluating the distribution of radiation dose, the viability of manufacturing the radioactivity prosthesis and its operational cost. In despite of being only one case, we can conclude that the prosthesis with gold-198 foils can be manufactured in acrylic with thickness thinner than those ones with cylinder of cesium-137, resulting lower operational costs, besides permitting better distribution of radiation dose on the lesion. PMID- 10705098 TI - Spontaneous regression of an anaplastic large cell lymphoma in the oral cavity: first reported case and review of the literature. AB - Lymphomas account for 2-5% of all oral malignancies and are the third most common in this site. This case report appears to be the first in the world literature describing spontaneous regression in the oral cavity of a subset of non-Hodgkins lymphomas known as Ki-1 anaplastic large cell lymphomas (ALCL). Ki-1 ALCL account for 2-7% of all non-Hodgkins lymphomas and the clinical presentation is variable; they may arise de novo or in the setting of a separate primary lymphoma and commonly present in the extra-nodal location. Disease severity is also variable with waxing and waning lesions at one extreme which may spontaneously regress to bone marrow involvement in around 12% of cases. This case is especially interesting since the patient is a farmer, given the recent evidence that there may be a link between non-Hodgkins lymphoma and this occupation. PMID- 10705099 TI - Body fat distribution, the menopause transition, and hormone replacement therapy. AB - Endocrine changes resulting from the menopause transition dramatically modify women's hormonal milieu. The consequences of these changes not only lead to cessation of reproduction and accompanying symptoms in women, but also dramatically impact long-term health. Loss of estrogen has been associated with the development of cardiovascular disease. Central distribution and accumulation of adipose tissue, and the concomitant insulin resistant dyslipidemic state have emerged as important components of a cluster of metabolic abnormalities that are strongly related to coronary heart disease. Thus, estrogen deficiency may affect cardiovascular disease risk by mediating changes in body fat distribution. This article is an update of the literature in the area of menopause, hormone replacement therapy, and body fat distribution. Cross-sectional studies using anthropometric measurements of abdominal fat distribution most often failed to detect an effect of the menopause transition that was independent of advancing age and degree of obesity. The use of radiologic techniques such as DEXA and computed tomography, however, led to the conclusion that the menopause transition accelerates the selective deposition of intra-abdominal fat. Available longitudinal data also support an increase in central body fatness occurring with menopause. Most intervention trials on hormone replacement therapy and body fat distribution showed that the treatment prevented the increase in central adiposity that was noted in postmenopausal women receiving no treatment or placebo. These results are supported by retrospective studies that showed a lower WHR in hormone users vs non-users. Mechanisms potentially explaining the menopause-related acceleration in abdominal fat accumulation include changes in regional adipose tissue metabolism in the face of a positive energy imbalance. As some inconsistencies were found among studies, further investigations using longitudinal and intervention designs, as well as more precise methodologies to measure body fat distribution, are needed to clearly establish the effects of menopause and hormone replacement on abdominal body fat distribution and the concomitant increase in cardiovascular disease risk. PMID- 10705100 TI - Chromium in the prevention and control of diabetes. AB - Chromium is an essential nutrient involved in the metabolism of glucose, insulin and blood lipids. Suboptimal dietary intake of chromium is associated with increased risk factors associated with diabetes and cardiovascular diseases. Within the past five years, chromium has been shown to improve glucose and related variables in subjects with glucose intolerance and type 1, type 2, gestational and steroid-induced diabetes. Severe neuropathy and glucose intolerance of a patient on total parenteral nutrition, who was receiving currently recommended levels of chromium, were reversed by additional supplemental chromium. Chromium increases insulin binding to cells, insulin receptor number and activates insulin receptor kinase leading to increased insulin sensitivity. Additional studies are urgently needed to elucidate the mechanism of action of chromium and its role in the prevention and control of diabetes. PMID- 10705101 TI - Psychological adjustment in a french cohort of type 1 diabetic children. PEDIAB Collaborative Group. AB - The aim of the study was to determine whether IDDM affects the course of psychological adjustment in youths. The study sample included 164 children with IDDM (mean age=10.2) and their parents compared to 164 healthy controls matched for age, sex and socioeconomic status. Adjustment was measured with the Child Behavior Checklist, a parental rating scale, validated and adapted for the French population. Two-way ANOVAs on CBCL scale scores showed that scores for both internalizing and externalizing problem behaviors and the total CBCL score were significantly raised in diabetic children (p<0.001). Further comparisons on the 8 narrow-band scale scores of the CBCL indicated increased scores for diabetic children on 6 dimensions. A significant Gender x Status (IDDM versus Controls) interaction was found, supportive of higher rates of aggressive behaviors amongst male diabetic children (p<0.01). Controlling for age, no correlation was found between CBCL total, internalizing and externalizing scores and duration of IDDM or HbA1c levels within the diabetic group. Psychological adjustment to chronic illness needs to be considered with respect to both normal developmental demands as well as in the context of the specific challenges posed by the disease. PMID- 10705102 TI - Normal and abnormal day-to-day variability of urinary albumin excretion in control and diabetic subjects. AB - Urinary albumin excretion (UAE) is very variable from day to day. This variability, more or less potent, might by itself have a patho-physiological significance. We analyzed day-to-day UAE in 207 elderly (60-75 years) inpatients (134 with and 73 without diabetes mellitus) attending the department of internal medicine of the Angers University hospital. Twenty-four-hour urine was collected 3 times during a 5-10 day hospitalization period. One-hundred-fifty-one patients (73%) displayed normoalbuminuria (UAE<30 mg/24 h in 2 or 3 measures) while 56 patients (27%) had microalbuminuria (UAE within 30-300 mg/24 h in 2 or 3 measures). As the raw data of UAE was not normally distributed, we transformed UAE into the variable z=log (log (k + UAE)) where k is an integer and looked for a k value for which z might be normally distributed. We found that z was actually normally distributed for k=2. Mean value and coefficient of variation of z in the 3 measurements were used to define the level and the temporal intra-individual variability of UAE. Expressed in term of z, the day-to-day intra-individual variability of UAE showed a potent change (from large variability to small variability) at the particular level z=1.25, corresponding to UAE=30.8 mg/24 h. This value is precisely the level currently used to define microalbuminuria in diabetic subjects. It is remarkable that the day-to-day variability of UAE collapses when UAE crosses the level which has been used to define microalbuminuria. PMID- 10705103 TI - Management of diabetic patients by general practitioners in France 1997: an epidemiological study. AB - To describe the characteristics of diabetic patients, the associated risk factors, the complications of the disease and its management by general practitioners (GPs) in France, a randomised sample of French GPs was asked to record data on all consecutive diabetic patients attending a regular visit within 3 months. Data were obtained by interview, clinical examination and usual follow up complementary examinations of the patients. Patients were classified into 3 groups:, patients treated with insulin and considered to have type 1 diabetes, [2i], insulin-treated patients expected to have type 2 diabetes, [2d], patients with type 2 diabetes and not treated with insulin. Data from 7540 diabetic out patients were recorded by 3084 GPs: 657 patients (8.7%) belonged to group 1, 1383 patients (18.3%) to group 2i and 5351 (71.0%) to group 2d. Patients, including 53.7%, [2i] 54.1%, and [2d] 56.5% of men, were (mean +/- SE) 58.8 +/- 0.7, [2i] 63.4 +/- 0.3, and [2d] 63.9 +/- 0.2 years old, respectively. Duration of diabetes was 15.9 +/- 0.4, [2i] 11.4 +/- 0.2, and [2d] 10.1 +/- 0.1 yr. The last fasting blood glucose level (laboratory assay) was 1.61 +/- 0.02, [2i] 1.68 +/- 0.01, and [2d] 1.61 +/- 0.01 g/L, and the last HbA1c 8.5 +/- 0.1, [2i] 8.1 +/- 0.1, and [2d] 7.8 +/- 0.1%, respectively. Tobacco smoking was observed in 19.2%, [2i] 13.1%, and [2d] 12.6% of the patients, hypertension in 39.6%, [2i] 55.9%, and [2d] 58.6%, micro- or macro-albuminuria in 18.6%, [2i] 11. 2%, and [2d] 9.5%, retinopathy in 31.1%, [2i] 12.9%, and [2d] 8.6%, and history of coronary artery disease in 16.3%, [2i] 15.0%, and [2d] 12.8%. Self-monitoring of blood glucose was performed by 93.2%, [2i] 37.9%, and [2d] 16.9% of the patients. During the previous 12 months, a visit had been performed with a diabetologist in 54.0%, [2i] 20.7%, and [2d] 12.9% of the patients, with an ophthalmologist in 62.9%, [2i] 51.5%, and [2d] 49.4%. These results underline the specific characteristics of French diabetic patients. A high prevalence of uncontrolled risk factors, mainly hypertension, contrasts with a relatively low frequency of micro- and macro-angiopathy, maybe underestimated by non-systematic routine follow-up. Closer collaboration between GPs and specialists should be developed to improve the management and care of diabetic patients in France. PMID- 10705104 TI - Mechanisms of postprandial hyperglycemia in liver transplant recipients: comparison of liver transplant patients with kidney transplant patients and healthy controls. AB - Impaired glucose tolerance or diabetes mellitus are frequent complications after organ transplantation, and are usually attributed to glucocorticoid and immunosuppressive treatments. Liver transplantation results in total hepatic denervation which may also affect glucoregulation. We therefore evaluated postprandial glucose metabolism in a group of patients with liver cirrhosis before and after orthotopic liver transplantation. Seven patients with liver cirrhosis of various etiologies, 6 patients having received a kidney transplant, and 6 healthy subjects were studied. Their glucose metabolism was evaluated in the basal state and over 4 hours after ingestion of a glucose load with 6.6 (2) H glucose dilution analysis. The patients with liver cirrhosis were studied before, and again 4 weeks (range 2-6) and 38 weeks (range 20-76, n=6) after orthotopic liver transplantation. Basal glucose metabolism was similar in liver and kidney transplant recipients. Impaired glucose tolerance was present in both groups, but postprandial hyperglycemia was exaggerated and lasted longer in liver transplant patients. Postprandial insulinemia was lower in liver transplant recipients, while C-peptide concentrations were comparable to those of kidney transplant recipients, indicating increased insulin clearance. Glucose turnover was not altered in both groups of patients during the initial 3 hours after glucose ingestion, but was higher in liver transplant early after transplantation during the fourth hour. Postprandial hyperglycemia remained unchanged in liver transplant recipients 38 weeks after liver transplantation, despite substantial reduction of immunosuppressive and glucocorticoid doses. We conclude that liver transplant recipients have severe postprandial hyperglycemia which can be attributed to insulinopenia (secondary, at least in part, to increased insulin clearance) and a late increased glucose turnover. These changes may be secondary to hepatic denervation. PMID- 10705105 TI - Insufficient adaptation of hypoglycaemic threshold for cognitive impairment in tightly controlled type 1 diabetes. AB - It is well known that hypoglycaemic thresholds for hormones and symptoms occur at lower plasma glucose levels in patients with strict glycaemic control. However, whether the threshold for cognitive impairment also shifts is still an unresolved question. We studied 19 type 1 diabetic patients, including 8 with hypoglycaemia unawareness, aged 37.0 +/- 7.4 y.r., with diabetes duration 15.2 +/- 10.7 yr, and HbA1c 7.6 +/- 1.1%. Hypoglycaemic thresholds for hormones, symptoms, awareness and cognitive function using the 4-choice reaction time test (4RT), were measured every 30 min during a 150 min stepped 4.4 to 2.2 mM hypoglycaemic hyperinsulinemic clamp. We found that 4RT- accuracy deteriorated earlier than 4RT time (3.2 and 2.7 mM, respectively, p<0.01), and that both correlated poorly with HbA1C before and after adjustment for age and diabetes duration (r=0.11, and 0.18, respectively). On the opposite, adrenaline, autonomic and neuroglycopenic symptoms, and awareness significantly correlated with HbA1c values (r=0.56, 0.70, 0.61, and 0.63, after adjustment, respectively). Furthermore, after allocating the patients into two subgroups according to HbA1c values (<8% n=12, and >=8% n=7), we found that, as opposed to other thresholds, accuracy and 4RT-time were minimally and not significantly influenced by glycaemic control, therefore exhibiting the smaller glucose thresholds shifts (- 0.2 and - 0.5 mM for accuracy and time, respectively, vs. 0.6 -0.8 for other thresholds). IN CONCLUSION: 1) the hypoglycaemic thresholds for cognitive dysfunction shift with strict glycaemic control, but not significantly and less than other thresholds, 2) as opposed to other reports, accuracy deteriorates earlier than speed during the 4RT test, and 3) these "maladapted" reactions may contribute to the higher risk for severe hypoglycaemia in subjects with tight glycaemic control. PMID- 10705106 TI - Clinical characteristics of type 2 diabetes in patients with mutations of HFE. AB - Genetic hemochromatosis (GH) is associated with two mutations of the HFE gene (Cys282Tyr and His63Asp). Heterozygosity for GH is associated with a mild increase in iron metabolism parameters, and increased iron stores are associated with abnormal glucose tolerance and decreased insulin sensitivity in the general population. We have previously shown that the frequency of the two HFE mutations is not increased in patients with type 2 diabetes. However, to assess whether the presence of HFE mutations modulates the clinical presentation of type 2 diabetes, we studied the clinical characteristics and iron metabolism indexes according to the presence of the two mutations in 266 patients with type 2 diabetes. The Cys282Tyr mutation and the His63Asp mutation were present in 9. 8% and 26% of the patients, respectively. Serum iron, transferrin saturation and ferritin concentrations were significantly increased in patients expressing either HFE mutations, compared to those without any mutation. There was no difference in the clinical characteristics in the two groups except that obesity was significantly less frequent in the patients with at least one mutation than in those without any mutation (27.6% vs 42.8%, p=0.02). This finding suggests that, in the absence of obesity, HFE mutations, through the insulin resistance associated with the increase in iron stores, may contribute to the onset of type 2 diabetes. PMID- 10705107 TI - [The practical audit: from theory to practice?]. PMID- 10705108 TI - [Health costs associated with the diabetic foot in developed countries.A plea for the creation of health care networks]. AB - COST OF DIABETIC FOOT: A limited number of epidemiological and economical studies on diabetic foot in France is available. Studies conducted in other countries, namely USA, Netherlands, UK and especially Sweden, have evidenced major direct and indirect costs that generally are underestimated in overall economical cost of diabetes forecast. The rapid increase of diabetes prevalence throughout the world, combined with diabetic population aging, allows to speculate that this situation may dramatically worsen in the future. In order to reduce the economical and human burden of this complication, it is an urgent public health necessity in France to set multidisciplinary organisations, dealing with both in patient care, which importance should be decreasing, and out-patient care, as well as prevention and foot care networks. The swedish experience figures an interesting model for our healthcare system. PMID- 10705109 TI - [Report of the experts of the ALFEDIAM (French Language Association for the Study of Diabetes and Metabolic Diseases) and SFN] Management of end-stage renal failure in diabetic patients. Short text]. PMID- 10705110 TI - [Limbal autograft transplantation, eight consecutive cases]. AB - BACKGROUND: Limbal autograft transplantation is the procedure of choice in the management of ocular surface disorders secondary to stem cells deficiency. The aim of our study was to investigate the indications, results and limits of this infrequent surgery. METHODS: Limbal autograft transplantation was performed in 8 patients and the mean follow-up period was 11 months. Limbal stem cell deficiency was due to chemical burns in 4 patients, history of prior surgery extending to the limbus in 2 patients, chronic limbitis with dystichiasis in one patient and persistent corneal epithelial defect after keratoplasty in one case. RESULTS: In 6 out of 8 cases, the stability of the ocular surface normalized and comfort significantly improved. Four of these patients had increased visual acuity after surgery. Two patients who presented with severe alkali burn did not respond well to limbal autotransplantation. CONCLUSION: Limbal autograft transplantation is a reliable and effective procedure in limbal stem cells deficiencies. New associate procedures such as amniotic membrane transplantation will however be necessary to improve the prognosis of very severe corneal surface disorders. PMID- 10705111 TI - [Alternating hemiplegia of childhoood and oculomotor anomalies]. AB - BACKGROUND: Alternating hemiplegia of childhood is a syndrome which begins in the first year of life. It is characterized by repeated attacks of uni-or bilateral hemiplegia or hemiparesia. In most cases paroxysmal manifestations are observed: movements or dystonia++ attacks, episodic nystagmus, abnormal eye movements and disturbance of the neurovegetative system, predominantly in the first year of life. ANALYSIS: In half of the cases, neurological anomalies begin during the neonatal period with a non characteristic aspect. Typical attacks take place after one year of life, sometimes associated with partial epilepsy. In a quarter of cases, the oculomotor anomalies have been known since early life. The diagnosis is made prior to one year on the basis of associated oculomotor anomalies and other symptoms without EEG arguments for epilepsy. Paroxysmal nystagmus is always found. One eye is affect in most cases, generally with horizontal and seldom with vertical movements of large variable pendular amplitude. One eye with nystagmus and the other with mydriasis is sometimes reported. Most attacks last from 30 sec to 3 min. Paroxysmal strabismus described in half of the cases seems to be generally unilateral internuclear transitory ophthalmoplegia. Finally, ocular deviations on the hemiparetic side are described. They are generally unique or sometimes associated with head deviation. Spontaneous blinking is reduced. CONCLUSION: Alternating hemiplegia of childhood is a non-epileptic sporadic, paroxysmal manifestation of unknown pathogenesis. Prognosis is poor. The presence of oculomotor signs suggests the diagnosis. PMID- 10705112 TI - [Toxicity of preserved and unpreserved beta-blocker eyedrops in an in vitro model of human conjunctival cells]. AB - PURPOSE: To compare the toxicity of a short-time application of timolol with benzalkonium chloride (timolol-BAC+) and unpreserved timolol (timolol-BAC-) in an in vitro model of human conjunctival cells. METHODS: Chang's conjunctival cell line (ATCC CCL 20.2) was treated for 15min. with 0.1%, 0.25% or 0.4% timolol BAC(+) or BAC(-) and then examined immediately or 24h later. Cell viability, chromatin condensation, mitochondrial mass and activity, free radicals production were studied by microplate cold light cytometry. Moreover, relative cell number was evaluated by crystal violet colorimetric test. In addition, cell size and the expression of an apoptotic marker Apo2.7 were studied by flow cytometry. RESULTS: Timolol-BAC(+) induced a rapid decrease in cell viability ranging from 40% immediately after treatment to 85% 24h later. A small, significantly less important decrease in cell viability was also observed with all tested concentrations of timolol-BAC(-). 24h after treatment with 0.25% timolol-BAC(+), the relative cell number was reduced by 55% whereas it did not vary after 0.25% timolol -BAC(-) treatment. Only timolol-BAC(+) induced chromatin condensation, decrease in mitochondrial membrane potential and cell size reduction. Moreover, cells treated with timolol-BAC(+) overexpressed the apoptotic marker Apo2.7. Also reactive oxygen species (ROS) production was significantly more important after cell exposure to timolol-BAC(+). CONCLUSION: In our model of conjunctival cells in vitro, timolol-BAC(+) induced irreversible cytotoxic damage with some characteristics of apoptosis. The active compound of timolol-BAC(-) could be responsible for ROS production and for cell viability variations. Oxidative stress could also play a role in timolol-BAC(+)-induced toxicity. In vitro toxic effects of antiglaucoma drugs could, in part, explain some ocular surface disorders in long-term treated patients. PMID- 10705113 TI - [High-dose intravitreal ganciclovir in CMV retinitis]. AB - PURPOSE: To assess the efficacy and tolerability of high-dose (2000microg) intravitreous ganciclovir in the maintenance therapy of CMV retinitis in AIDS patients. MATERIAL: and methods: Prospective open study in a single center. The maintenance therapy of CMV retinitis consisted of a single high-dose (2000microg) intravitreous injection per week. The evaluation criteria were clinical examinations, and photographs when necessary. RESULTS: Over a 44-week period, twenty-four eyes (15 patients) have been included. The relapse rate evaluated by the survival curves method was 5% at 44 weeks. No ocular toxicity was reported after a follow-up of 24 weeks. CONCLUSION: High-dose intravitreous ganciclovir induces a prolonged remission of CMV retinitis and is well tolerated. Therefore, we recommend the use of the 2000microg dosage per intravitreous injection instead of the classical dosage of 400microg. PMID- 10705114 TI - [Radiotherapy for age-related macular degeneration: risk factors of complications, prevention and treatment of side-effects]. AB - PURPOSE: To analyze the retinal and choroidal side-effects of radiotherapy given for age-related macular degeneration (ARMD) and to describe the risk factors of these complications and their treatment. MATERIAL: and methods: Two hundred and ninety five eyes in 270 patients with ARMD were treated using radiotherapy. Nineteen patients had diabetes. The doses were as follows: 15 Gy or less (4 eyes); 16 Gy/4 fractions (113 eyes); 18 Gy/5 fractions (35 eyes); 20 Gy/5 fractions (123 eyes); 24 Gy/6 fractions (2 eyes); 28.8 Gy/8 fractions (17 eyes); more than 28.8 Gy (1 eye). Patients had a regular follow-up visit with visual acuity, contrast sensitivity evaluation, biomicroscopic fundus examination, fluorescein and ICG angiographies every six months over a mean period of 15 months. RESULTS: Radiation retinopathy was noted in 15 eyes, a bilateral neovascular glaucoma in one patient, ischemic optic neuropathy in 5 eyes, choroidal telangiectasiae in 19 eyes, venous occlusion in 2 eyes, oedematous retinopathy with major exudation (ORME) in 31 eyes, and choroidal hematoma in 8 eyes. Radiation retinopathy, choroidal telangiectasiae and ORME were related to radiation dose. Radiation retinopathy was more severe and more frequent in patients with diabetes. Choroidal telangiectasiae were diagnosed with ICG angiography and were treated early with laser. CONCLUSION: Radiotherapy for ARMD should not be done in patients with diabetes. Hypofractionation is not recommended. ICG angiography should be considered essential in the follow-up of patients treated with radiotherapy. PMID- 10705115 TI - [B-scan ultrasonography and optical coherence tomography (O.C.T.) in epiretinal macular membranes: pre- and post-operative evaluation]. AB - PURPOSE: This study was designed to evaluate B-scan ultrasonography and optical coherence tomography in the pre- and post-operative morphologic evaluation of idiopathic epiretinal macular membranes. METHODS: Fifteen eyes were examined using B-scan ultrasonography and optical coherence tomography before and after surgical excision of the epiretinal macular membranes. RESULTS: Ultrasonography prior to surgery allowed an excellent evaluation of both the peripheral and the posterior vitreous and showed the membrane. The thickening of the retina and the retinal folds were equally visualised by either method. An improved observation of the macular oedema was obtained by the optical coherence tomography as well as the type of membrane adhesion. After surgery, both methods were able to detect membrane remnants. A better visualisation of retinal thickness and cystoid macular oedema was obtained by optical coherence tomography rather than ultrasonography. Serous sub-foveal retinal detachment was only revealed by optical coherence tomography. DISCUSSION: Ultrasonography is indispensable in case of media opacity or of optical inaccessibility of the posterior pole. Moreover, it allows an excellent global analysis of the anterior and posterior vitreous, which is very useful for the surgeon. The membrane is always showed by ultrasonography, but not always by optical coherence tomography in case of diffuse adherence. Both methods can detect the membrane and a cystoid macular edema. However, retinal analysis with the optical coherence tomography is more precise. It can even detect cysts of the macular oedema or serous sub-foveal retinal detachment. CONCLUSION: B-scan ultrasonography and optical coherence tomography give complementary information in pre- and post-operative examination of epiretinal macular membranes. PMID- 10705116 TI - [Complications of radiotherapy in patients with age-related macular degeneration (AMD). A study of 48 cases]. AB - PURPOSE: This study describes the ocular complications after teletherapy indicated for patients with age-related macular degeneration (AMD). MATERIAL: and methods: The study concerned 48 patients suffering from an exudative form of age related macular degeneration, non suitable for laser photocoagulation, and irradiated by high-energy electron beam therapy. RESULTS: With a mean follow-up time of 2 years, late effects were observed in 16 (33. 3%) patients: 9 (18.75%) radiation optic neuropathies, 9 radiation retinopathies (18.75%) and 2 neovascular glaucomas (4.1%). CONCLUSION: The frequent and serious complications observed led us to adapt the technique and dose of radiation therapy indicated in patients with age-related macular degeneration. PMID- 10705117 TI - [Pigmentosum retinis and tubulo-interstitial nephronophtisis in Sensenbrenner syndrome: a case report]. AB - PURPOSE: Sensenbrenner syndrome or cranio-ectodermal dysplasia is an extremely rare autosomal recessive condition (12 cases reported in literature). Our observation shows the possibility of both ocular and renal involvement associated with cranio-ectodermal abnormalities. PATIENTS: and method:We report the case of a girl who presented a typical cranio-ectodermal syndrome with dolicocephaly, short thorax, short limbs, short fingers and teeth abnormalities. At five years, she was found to have pigmentosum retinitis with amblyopy and moderate hyperopia. A chronic renal failure with uncontrollable hypertension underwent a cadaveric donor transplantation at the age of six years. RESULTS: Two years later, the pigmentosum retinitis was stable. The kidney histology revealed a tubulo interstitial nephronophtisis. The molecular analysis of the NPH 1 locus, which was associated with nephronophtisis, was negative. DISCUSSION: Our observation and two recent publications have in common ocular and renal abnormalities associated with cranio-ectodermal dysplasia. The underlying genetic defect would involve not only morphogenesis but also development and maturation of organs as eye and kidney. Sensenbrenner syndrome would thus be similar to certain disorders affecting the eye, kidney, skeleton and ectodermal structures such as the EEM, Senior-Loken, Mainzer-Saldino, and Jeune syndromes. CONCLUSION: The retinal dystrophy falls within the spectrum of clinical and genetic forms of pigmentosum retinitis. Our observation would confirm possible links between Sensenbrenner syndrome and oculorenal syndromes. PMID- 10705118 TI - [Medulloepithelioma of the ciliary body. A case report]. AB - Medulloepithelioma of the ciliary body is an uncommon intraocular tumor occurring during the first year of life. Malignant degeneration may occur. We report the case of a 4-year-old child who presented medulloepithelioma of the left eye disclosed by oesotropia at 2 years of age. Clinically, there was oesotropia, positive light perception and cataract with vascular membrane spreading to the nasal side of the irido-corneal angle. CT scan and ultrasound B revealed a ciliary body tumor involving the sclera and orbital fat. After exenteration, the pathology study reported malignant medulloepithelioma of the ciliary body with scleral extension. No local recurrence or metastasis has been observed at 8 months follow-up. We discuss the clinical, radiological and therapeutic features of this uncommon tumor. PMID- 10705119 TI - [Endogenous aspergillus endophthalmitis: a case repport]. AB - We report a case of endogenous aspergillus endophthalmitis. This infection occurred in a young immunocompromised boy of 6 years old. The localisation of the chorioretinitis was unusual because out of the posterior area. The evolution was favorable with recovering of the visual acuity under general treatment and intravitreous injections. PMID- 10705120 TI - [Fragmentation of hydrogel episcleral implants: a series of 4 cases]. AB - A histopathologic study was performed on 4 capsule specimens obtained after surgical removal of hydrogel episcleral implant previously sutured to the scleral surface aiming to reattach the retina. Fragments of hydrogel were found on the inner surface of the capsular fragments, each of them parts of the capsule, which coated the hydrogel implant on the scleral surface. Some fragments were surrounded with a foreign body giant cell granulomatous reaction. Changes in scleral curvature and scleral thinning were consequences of the buckling procedure. The hydrogel fragmentation previously observed in experimental and human specimens gave histologic evidence of degradation of this material after its implantation on the eye surface. This characteristic demonstrates the instability of the material after implantation. PMID- 10705122 TI - [Discrepancies and illegalities]. PMID- 10705121 TI - [Gorlin-Goltz phacomatosis: ophthalmological aspects in one case]. AB - We report the case of a 21-year-old girl who presented an eyelid tumor with retinal hamartoma. General examination revealed a basal cell nevus on the face, jaw cysts, skeletal malformations and brain calcifications. Histological examination of the eyelid lesion and of the skin nevus showed basal cell carcinoma. Familial investigation evidenced the hereditary nature of this disease. We review Gorlin-Goltz phakomatosis, an uncommon disease often unrecognized by ophthalmologists, and discuss nosological considerations. PMID- 10705123 TI - [Immunomodulators in the prevention and the treatment of corneal graft rejection]. PMID- 10705124 TI - [Surgical correction of ametropia after corneal grafts]. PMID- 10705125 TI - [Difficult cornea procurement: causes and consequences of the exceptional situation in France]. AB - Despite scientific advances, corneal grafting is still in a crucial situation in France in 1999. Over the last 3 years, the number of corneal grafts (2 903 in 1996, 3 213 in 1997, and 4 053 in 1998) has been insufficient to satisfy estimated needs (8 041 in 1996, 8 303 in 1997, 7 400 in 1998). The consequences are waiting lists, long time on waiting lists, and regional differences. This situation results more from difficulties in the cornea procurement system than from limited numbers of potential donors or restricted selection criteria. Progress will depend greatly on structural changes. PMID- 10705126 TI - [Torpedo maculopathy]. PMID- 10705127 TI - [ [In Process Citation] PMID- 10705128 TI - [ [In Process Citation] PMID- 10705129 TI - [ [In Process Citation] PMID- 10705130 TI - [Editorial] [In Process Citation] PMID- 10705131 TI - [Teaching targets in vascular medicine]. AB - The reference list presented here is a selection of educative objectives performed by the French "College des Enseignants de Medecine Vasculaire" for the different levels of the medical initial education course. It was produced through a collective procedure, after selecting the most relevant topics and setting up writing rules based upon docimology, and favoring practical rather than theoretical objectives. The main topics are peripheral obstructive arterial disease, polyatherosclerosis, atherosclerosis risk factors, venous thromboembolic disease, thrombophilia, chronic venous insufficiency, lymphatic insufficiency, leg ulcers, vascular acrosyndromes, cerebrovascular diseases and connective tissue diseases, vascular occupational diseases, vascular adverse effects of drugs, diabetic vascular disease, the vascular consequences of hypertension, vascular malformations and angiodysplasia, inflammatory arterial diseases, and vascular explorations. As a whole they include about 300 objectives for the five teaching levels. We hope that this list will help stimulate production of training courses and documents strongly needed in this field. PMID- 10705132 TI - [Tolerance and therapeutic results of iloprost in obliterative arteriopathy in lower limbs at the severe chronic ischemia stage. A retrospective study of 29 consecutive cases]. AB - The aim of this retrospective, study was to assess the tolerance and therapeutic effect of a stable prostacyclin (iloprost) analog in severe forms of permanent lower limb ischemia. Ninety consecutive unselected patients, in Leriche and Fontaine stages III or IV, turned down for vascular surgery after angiography and treated with iloprost for 28 days were enrolled in the study. Patients were followed up clinically (ischemic pain, trophic changes, walking distance) and with transcutaneous oxymetry (D28). Long-term assessment (mean 2 years) was expressed as rates of death, major amputation and "patients alive with limb". There were no manifestations of intolerance to iloprost. At two months, 42 out of 90 patients (47%) were considered as responders because of a lack (n=36) or significant decrease (n=6) in pain, reduction of trophic lesions and conservative walking. At long term (6 months, one and two years) we observed that 10 (11%), 17 (20%) and 22 (25%) patients respectively had died, 24 (27%), 26 (30%) and 28 (32%) patients underwent major amputation, but 60 (68%), 54 (62%) and 49 (56%) patients still alive with their limb and conservative walking. No predictive factors were noted, but diabetic patients without microangiopathy or recent bypass occlusions (respectively 43% and 56% out of patients were alive with limb at 6 months) were associated with bad results. This retrospective study, despite its limitations, underlines the good tolerance to, and effectiveness of iloprost in non surgical chronic critical ischemia. However, no predictive criterion of long-term effectiveness could be established, except initial clinical severity and clinical change one month after treatment. PMID- 10705133 TI - Proximal long saphenous vein valves in primary venous insufficiency. AB - OBJECTIVE: To verify some of the previous findings of venous valves described in the literature, their pathophysiological significance and clinical implications. MATERIALS AND METHODS: The elementary components of 65 proximal valves of the long saphenous vein and their interrelationships were subjected to histopathological examination. Valves were taken from patients subjected to long saphenous vein surgical removal for varicose veins of the lower limbs. Measurements and morphological evaluations were performed by optical microscopy. RESULTS: The valvular sinus, agger and proximal portion of the cusp underwent parallel variations of thickness. Thickening of the proximal portion of cusp was related to increase in smooth muscle cells in the agger and to elastic layer dissociation. Thickening of the distal portion of cusp depended on the collagen component; sometimes it was shortened, crumpled and led to the formation of a thickened border. The vein wall in a commissural aneurysm was usually thinner than in the valvular sinus. Alterations in the intima, in the elastic membrane and in the media were found in the 98% of the valvular annulus. Ectasis and asymmetry of the venous wall were mainly related to the muscular hypoplasia of the media. CONCLUSIONS: The development of primary venous insufficiency seems to be due to the following tissue alterations: dilatation of the valvular annulus and hypotrophy of the cusp. The hemodynamic mechanical injury increases the tissue damages of both annulus and cusps. This pathophysiologic interpretation of venous insufficiency suggests the need for detailed diagnostic procedures before reparative surgery of valves. PMID- 10705134 TI - [Assessment of the prevalence of atherosclerotic lower limb arteriopathy in France as a systolic index in a vascular risk population]. AB - OBJECTIVE: Obliterative arteriopathy of the lower limbs is a severe disease. History taking often underestimates prevalence. In studies using the Rose questionnaire and examining the prevalence of symptomatic arteriopathy defined by the presence of intermittent claudication, prevalence has been rather constant, around 2% in the general population in industrialized countries. A more clinical approach searching for physical anomalies (absence of distal pulse) generally gives higher rates. The most recent data led us to conduct a study focusing on screening for arterial disease using the systolic index (the systolic index is the ratio between the ankle and humeral systolic pressure). A systolic index below 0.90 would be a sign of defective perfusion, increasing in severity with poorly compensated arterial lesions. METHOD: A survey was performed in a random sample of 150 practitioners using an allocation procedure to the nearest colleague in case of refusal. A data sheet containing demographic data and the main risk factors was established for each consulting patient aged from 40 to 80 years. The systolic index was measured in each patient with at least one vascular risk or who consulted for pain in the lower limbs. A simple sequential non randomized patient recruitment scheme was used. RESULTS: The survey population included nearly 9,000 patients (8,987), 46% men and 54% women, mean age 64 years (table I). Patient risk factors including smoking, diabetes, hypertension, and physical exercise were adjusted for sex and age (table II-VI, IX). The systolic index was recorded in 41% of the population who had a vascular risk factor. Among these patients, nearly one-fourth had a systolic index under 0.90, giving a prevalence in the sample population of 11% (table VII). This rate was also assessed by age and sex (table VIII). Logistic regression evidenced a prognostic value (fig. 1) for smoking, hypertension and sedentary activity, and to a lesser extent, for age and sex. There was a significant difference by latitude, patients living in northern France having a higher risk than those living in southern France (table X). In addition, there was a relationship between the systolic index and spontaneous complaint, showing that this complaint is poorly specific for arterial disease (table XI, XII). CONCLUSION: This epidemiological approach to arteriopathy implemented by general practitioners searching for patients with a vascular profile and measuring systolic index is a novel method. It gave an estimation of prevalence at 11% in the French population, a rate compatible with that reported by other international teams. The method also allowed an analysis of factors affecting the rate. PMID- 10705135 TI - [Leukocyte adhesion on a fibrinogen-coated surface under static conditions: experimentation and creation of a model]. AB - The adhesion of polymorphonuclear leukocytes (PMNs) on the vascular endothelium is a complex process that occurs during different biological and pathological events and involves numerous molecules. The adhesion cascade is induced after PMN stimulation by various molecular or cellular signals. Fibrinogen is one of the substrates for CD11b/CD18 B2-integrins expressed at the PMN surface; fibrinogen neutrophil binding is induced by inflammatory reactions. In order to understand this process, we have carried out studies on the basis of preliminary experiments on red blood cells and synthetic particles. The modelization of quiescent PMNs adhesion on a fibrinogen substrate was investigated with a sedimentation cell chamber. Two different physiological conditions were tested: the activated state of PMN by a synthetic pro-inflammatory activator (FMLP). The activated state of PMNs was both quantified by flow cytometry and controlled by fluorescence microscopy. The results suggest that quiescent neutrophils deposit in accordance with the ballistic deposition model. This random adsorption model differs from random sequential adsorption (RSA) in that the cells arriving at the surface are able to roll along cells previously adsorbed introducing the notion of gravitational attraction of cells. The preliminary results obtained with stimulated PMN do not allow to choose between one of this two deposition models. Nevertheless, the qualitative and quantitative effects of FMLP on neutrophils were demonstrated by modifications of adhesion molecules expression. PMID- 10705136 TI - [Diabetes mellitus and fibrinogen. Hemorrheological and microcirculatory consequences]. AB - Fibrinogen level, erythrocyte aggregation and transcutaneous oxygen pressure (TcPO2) were measured in the two populations of 32 healthy volunteer subjects and 119 diabetic patients without any clinical sign of micro and/or macroangiopathy. Measured parameters were studied according to both the long term glycemic control (HbA1C) and the type of diabetes. Results showed a significant elevation of plasma fibrinogen in all diabetic patients even when they had good glycemic control. Hyperfibrinogenemia appeared to be related to both long term glycemic control and type of diabetes. Hyperfibrinogenemia in studied diabetic patients was associated with a tendency to erythrocyte hyperaggregation. Indeed, this hyperaggregation like fibrinogen level, seem to be related to both HbA1C level and the type of diabetes. In contrast, the TcPO2 was found to be significantly decreased in all patients irrespective to either HbA1C or the type of diabetes. Besides, TcPO2 was not correlated with fibrinogen nor erythrocyte aggregation. In conclusion, the decrease in TcPO2 as observed here, does not directly depend upon the type of diabetes, or the HbA1C level, i.e the quality of mid and/or long term glycemic control. PMID- 10705137 TI - [Neonatal catheterization, an underestimated cause of claudication in young patients? A case report]. AB - We report a case of isolated occlusion of the common iliac artery diagnosed at the age of 25 years in a woman, that was supposed to be related to occlusion of a perinatal arterial catheterization of the umbilical artery. This original observation opens discussion to: 1) the delays in the diagnosis of arterial disease in young subjects; 2) the need of an extensive history report; 3) the high frequency of arterial complications following arterial umbilical catheterization; 4) the problems of fertility in case of hypogastric arterial occlusion. PMID- 10705138 TI - [Spontaneous hematoma: thoughts about selective serotonin uptake inhibitors]. PMID- 10705139 TI - [Treatment of acute coronary syndrome]. PMID- 10705140 TI - [CT imaging of pelvic injuries in polytrauma patients]. AB - CT has become a major tool in the management of multiple trauma. Concerning pelvic trauma, visceral and bone injuries can be detected. We review here the different pelvic injuries focusing on CT diagnostic approach. PMID- 10705141 TI - [The new listing of medicals acts...is becoming!]. PMID- 10705142 TI - [3D-CT angiography with volume rendering technique in the intracerebral aneurysms]. AB - PURPOSE: Many techniques of 3D reconstruction (MIP, SSD) permit a good evaluation of the circle of Willis in order to detect cerebral aneurysms. More recently, the advent in the clinical practice of a calculation algorithm (VRT) adapted to the workstations for images treatment seems to improve evaluation of the characteristics regarding these aneurysms. MATERIALS AND METHODS: We report 4 cases with cerebral aneurysms studies with CT-angiography using the technique MIP and VRT. RESULTS: The VRT, using the totality of image informations, allows a better understanding than MIP about the intracranial cerebral aneurysms including their constitution and 3D localization. CONCLUSION: The VRT reconstruction permits to obtain quickly good quality and reproductive images, without relationship with threshold. PMID- 10705143 TI - [ROC analysis comparing screen film mammography and digital mammography]. AB - PURPOSE: To compare the diagnosis performances of radiologists on screen film versus digital mammography. MATERIAL: and methods: Two sets of 123 mammograms, screen film mammography and storage phosphor digital mammography, are studied comparatively with ROC analysis. RESULTS: Phantom study show that conventional method give better scores for usual tension but the detectability of smaller microcalcification is equivalent. To obtain with digital technic the same conventional score you have to increase the radiation dose. Roc Curves, simulated "detection" mode showed that radiologists performed with higher accuracy with conventional system but this difference is weekly statistically significant. ROC Curves, simulated "diagnostic" mode showed the same results wit no statistically significant difference but when the decision to go to the biopsy is the gold standard, ROC Curves were essentially equivalent for both film screen and digital mammography system. The readers consistently considered the digital mammograms to be less suspicious for cancer findings. The agreement study as proposed by the FDA indicate that probability of a positive digital mammograms given a positive screen film is 75% (threshold value 90%) and the probability of a negative digital mammograms given a negative analog film is 85% (threshold value 85%). CONCLUSION: Analysis of specific discrepancies indicate that spatial resolution is an essential limiting factor for digital method but high resolution phosphor plate are interesting in imaging treated breast, radioluscent lesion, fatty benign tumor, hamartoma, intramammary lymph node, breast with prosthesis. PMID- 10705144 TI - [Capsular retraction of shoulder: prospective study of imaging benefits in 20 patients]. AB - PURPOSE: To evaluate the contribution of principal imaging techniques in diagnosis and treatment in adhesive capsulitis of the shoulder. MATERIALS AND METHODS: In 20 patients presenting adhesive capsulitis of shoulder since mean of 6,7 months, the following examinations were performed: radiographies, angioscintigraphy, MRI as well as an opaque arthrography and a bursography associated with corticosteroid injection. Patients were followed during one year. RESULTS: The opaque arthrography was to affirm the adhesive capsulitis for the inclusion of the patients. Radiographies (patchy demineralization) and scintigraphy (hyperfixation) were often pathological. In MRI, T1 fat-saturated sequences after contrast injection almost always showed enhancement of the articular capsula, the synovia, the miscellaneous bone or the sub-acromial bursa. The latter was often modified and retracted at bursography. In 19 of 20 cases, a functional improvement was observed after the opacifications. CONCLUSION: Therapeutic effect of both arthrography and bursography is almost proved. Post contrast MRI confirms presence of vascular troubles in all the shoulder structures even at this advanced stage. PMID- 10705146 TI - [Small bowel volvulus due to midgut malrotation: value of Doppler sonography. A case report]. AB - A case of small bowel volvulus due to midgut malrotation in a 4 years-old child complaining of vomiting and abdominal pain. Diagnosis was made by a Doppler ultrasound showing the volvulus and the intestinal malrotation. We are discussing the interest of this technique. PMID- 10705145 TI - [Agenesis of the internal carotid artery: two cases report]. AB - The authors report two cases of agenesis of the internal carotid artery evaluated with MR angiography allows the diagnosis of the internal carotid artery agenesis and appreciates both the supply post. PMID- 10705147 TI - [Hydatid cyst of the mediastinum: 2 cases]. AB - Hydatid cysts of the mediastinum are very uncommon. We report two cases of mediastinal hydatic cyst. The first case was located in the cardiophrenic recess while the other was within both the heart and mediastinum. A review of the imaging findings and value of various techniques is presented. PMID- 10705148 TI - [A case of lymph nodes involvement in malakoplakia of the bladder]. AB - This case illustrates the involvement of lymph nodes in a patient with histologically proven malakoplakia of the urinary bladder. The nodes appeared hyperdense at CT and hyperintense at MR. PMID- 10705149 TI - [What is it? Interpretation: an aneurysm of a coronary-pulmonary fistula demonstrated by computed tomography]. PMID- 10705150 TI - [FluoroCT guided biopsies: a simplified technique]. AB - We report the use of a specific handle we designed for interventional procedures guided by fluoro-CT. This handle, easy to use and low cost, improves the procedures reducing dramatically the procedure time. PMID- 10705151 TI - [The exam request seen by the radiologist, the report seen by the clinician]. PMID- 10705152 TI - [ [In Process Citation] PMID- 10705153 TI - Aplastic anemia before bone marrow transplantation and antilymphocyte globulin. AB - In severe aplastic anemia, disease-dependent mortality was high before allogeneic bone marrow transplantation (BMT) and immunosuppressive therapies (IST) including antilymphocyte globulin became available. However, under supportive therapy alone, spontaneous remissions were observed in up to 20% of severe cases, reflecting the natural course of the disease. Therefore, in evaluating new forms of treatment, one has to keep in mind that remission is not necessarily response, and that final proof of utility of any new therapy still requires a randomized study design. Transition to leukemia or myelodysplasia was rarely observed if the initial diagnosis was accurate. The much higher incidence of leukemias in patients treated by IST, but not by BMT is probably due to the better life expectancy of patients at risk, rather than to a leukemogenic potential of IST itself. 'Outdated' therapeutic modalities, such as androgens or splenectomy, may still be justified as an adjuvant therapy in selected cases. PMID- 10705154 TI - What is the definition of cure for aplastic anemia? AB - Treatment with immune suppression and bone marrow transplantation has improved the response rates and survival of patients with aplastic anemia. Measurement of response requires that common endpoints be recorded at specific times. There has been no agreement on such parameters for patients with aplastic anemia. In this paper issues related to measurement of response are reviewed and criteria for response are proposed. Adoption of uniform criteria would facilitate comparisons of treatment efficacy. PMID- 10705155 TI - Current results of bone marrow transplantation in patients with acquired severe aplastic anemia. Report of the European Group for Blood and Marrow transplantation. On behalf of the Working Party on Severe Aplastic Anemia of the European Group for Blood and Marrow Transplantation. AB - We have analyzed 2,002 patients grafted in Europe between 1976 and 1998 from an identical twin (n = 34), from an HLA-identical sibling (n = 1,699) or from an alternative donor (n = 269), which included unrelated and family mismatched donors. The proportions of patients surviving in these three groups are, respectively, 91, 66 and 37%: major causes of failure were acute graft-versus host disease (GvHD) (11%), infection (12%), pneumonitis (4%), rejection (4%). In multivariate Cox analysis, factors predicting outcome were patient's age (p < 0.0001), donor type (p < 0.0001), interval between diagnosis and bone marrow transplantation (BMT) (p < 0.0005), year of BMT (p = 0.0005) and female donor for a male recipient (p = 0.02). Patients were then divided in two groups according to the year of BMT: up to or after 1990. The overall death rate dropped from 43 to 24% (p < 0.00001). Improvements were seen mostly for grafts from identical siblings (from 54 to 75%, p < 0.0001), and less so for alternative-donor grafts (from 28 to 35%; p = 0.07). Major changes have occurred in the BMT protocol: decreasing use of radiotherapy in the conditioning regimen (from 35 to 24%; p < 0.0001) and increasing use of cyclosporin (with or without methotrexate) for GvHD prophylaxis (from 70 to 98%; p < 0.0001). In conclusion, the outcome of allogeneic BMT for patients with severe aplastic anemia has considerably improved over the past two decades: young patients, grafted early after diagnosis from an identical sibling, have currently an over 80% chance of long-term survival. Transplants from twins are very successful as well. The risk of complications with alternative donor transplants is still high. PMID- 10705156 TI - Results of immunosuppression in aplastic anaemia. AB - The availability of immunosuppressive agents, such as antilymphocyte globulin and cyclosporin, has had a major impact on the outcome of patients with aplastic anaemia. The majority of patients will show a response to immunosuppressive therapy. For those who fail to respond to an initial course, a second or further courses of immunosuppression can be given. Improvement in survival may also be due to the improved quality of supportive care available today for patients with aplastic anaemia. The additional use of haemopoietic growth factors such as granulocyte colony stimulating factor with immunosuppressive therapy requires further prospective randomised studies in order to evaluate whether they are of benefit for patients with aplastic anaemia. Because of the rarity of this condition, the use of multicentre prospective randomised studies is crucial for further evaluation of treatment modalities in aplastic anaemia. PMID- 10705157 TI - Cell cycling stress in the monocyte line as a risk factor for progression of the aplastic anaemia/paroxysmal nocturnal haemoglobinuria syndrome to myelodysplastic syndrome. AB - Severe aplastic anaemia (SAA) causes permanent stem cell damage from which patients do not recover after treatment with antilymphocyte globulin (ALG). To produce peripheral blood values compatible with life, the few remaining stem and precursor cells are put under stress. We defined a 'stress factor' (SF) for various haematopoietic lines as the ratio of the corresponding peripheral blood (PB) value to the total colony number in short-term bone marrow cultures from 86 patients with different outcomes. Both values are expressed as percentage of normal, hence SF averages 1 in normal steady-state haematopoiesis. SF was elevated in all patients, from 2-to 40-fold, with wide variations in different patient groups and striking differences between haematopoietic lineages. In long term disease-free survivors after ALG (group 1) the mean total colony count was 19% of normal, with a significantly higher proportion of erythroid burst-forming units compared to normal. They had ineffective erythropoiesis with haemoglobin (Hb) values below, and reticulocyte counts above normal; platelet counts were 67% of normal. In contrast, monocyte counts were in the high normal range, resulting in a high SF (18.7 +/- 1.9) for monocytes. In patients who developed paroxysmal nocturnal haemoglobinuria (PNH) after ALG (group 2), ineffective erythropoiesis, reflecting haemolysis, was more pronounced and they had striking relative monocytosis, resulting in a significantly higher SF for monocytes (33.7 +/- 5.7) compared with group 1 (p < 0.0001). High monocyte counts most likely reflect the relative resistance of nucleated cells to complement, compared with red cells and platelets. Patients who developed myelodysplastic syndrome (MDS) or acute myeloid leukaemia (AML) after ALG, with or without PNH (group 3), had the highest SF for monocytes (39 +/- 10). They also had neutrophil counts in the upper range, or above normal, resulting in a high SF for neutrophils: 32 +/- 19. In patients with persisting or relapsing-remitting pancytopenia without a clinically detectable clonal disorder (group 4), all values were strikingly similar to those of the PNH group. In patients who achieved normal PB values after uncomplicated bone marrow transplantation (group 5), the SF averaged 3, but they also had ineffective erythropoiesis and mild relative monocytosis, a possible sign of occult PNH. We conclude that all patients after treatment of SAA have ineffective erythropoiesis and relative monocytosis, and that these abnormalities probably reflect PNH. We suggest that the resulting high SF for the leukocyte - particularly the monocyte line - predisposes to the development of MDS/AML. We discuss how these results may provide some of the missing pieces in the puzzle of SAA/PNH. PMID- 10705158 TI - Paroxysmal nocturnal haemoglobinuria: a replacement of haematopoietic tissue? AB - Acquired somatic mutations of the PIG-A gene lead to deficient expression of glycosyl-phosphatidyl-inositol-anchored proteins (GPI-AP) by haematopoietic cells and play a causative role in the pathogenesis of paroxysmal nocturnal haemoglobinuria (PNH). However, PIG-A mutations do not explain how the defective PNH clone can expand. It was hypothesized that a selection process conferring a relative advantage to the GPI-AP-deficient population is required. Since GPI-AP deficient cells are also detectable in a substantial proportion of patients with otherwise typical aplastic anaemia (AA), the mechanisms inducing bone marrow failure might selectively spare the GPI-deficient cells. In order to examine the growth characteristics of GPI-AP-deficient cells in more detail, we performed repeated analyses of GPI-AP expression on peripheral blood cells in 41 patients with AA. We observed four patterns of the course of GPI-AP-deficient populations: (1) 13 patients showed normal expression of GPI-AP in the first analysis and in at least two follow-up studies at a median time of 709 days after the first analysis. (2) Secondary evolution of a GPI-AP-deficient population was a rare event. Only 4 patients with initially normal GPI-AP expression developed a GPI-AP deficient population during follow up after immunosuppressive treatment. (3) Persistence of GPI-AP-deficient cells was observed in 16 patients during a median follow-up time of 774 days. However, in some patients, the size of the GPI-AP deficient population increased substantially. (4) Disappearance of a GPI-AP deficient population was observed in 8 patients. The time course of GPI-AP expression in relation to the treatment suggests that therapeutic interventions might modulate the ratio of normal versus GPI-AP-deficient haematopoiesis. Overall, these data argue against an 'absolute growth advantage' of GPI-AP deficient cells. Our data are consistent with the hypothesis that haematopoietic failure caused by damage to normal haematopoiesis allows the outgrowth of a GPI AP-deficient population. Thus, in at least some patients GPI-AP-deficient cells might pre-exist at a very low percentage and replace haematopoiesis after an insult to the normal cells. PMID- 10705159 TI - Cure from severe aplastic anemia in vivo and late effects. AB - Recent progress in the treatment of aplastic anemia has dramatically changed the previously grim prognosis for these patients. Improvements in bone marrow transplantation and immunosuppression have increased the number of long-term survivors so that immediate survival is no longer the sole concern. Here, were review the major clinical studies and summarize recent analyses of late malignant and nonmalignant complications following treatment of aplastic anemia. PMID- 10705160 TI - Predictive factors for cure after immunosuppressive therapy of aplastic anemia. AB - In a previous study, we evaluated efficacy of repeated antilymphocyte globulin (ALG) treatment for patients with severe aplastic anemia not responding to an initial ALG treatment or relapsing after initial response to ALG. We now searched in the same cohort of patients for differences between patients who responded to treatment and remained free of complications and those who relapsed or developed a clonal complication. From 107 patients surviving for more than 1 year after immunosuppression, 34 remained free from complications after the first course of ALG, and 73 presented an event defined as relapse of aplastic anemia, development of a clonal complication such as paroxysmal nocturnal hemoglobinuria, myelodysplastic syndrome or leukemia, or appearance of a solid tumor. We compared these two groups for survival, clinical performance and blood counts during follow-up. Survival probability was 93% for the event-free patients, and 55% for the patients with a complication event (p = 0.0003). Event-free patients had a higher incidence of complete remission (71%), were more often free of immunosuppressive treatment (79%) and independent of transfusions (100%), and had a higher Karnofsky score (91% with a score > or =90%) as compared to the group with events (29, 37, 67, 48%; p < or = 0.0002). At 1 and 3 years, event-free patients had significantly higher leukocyte and neutrophil counts, as compared to patients with a complication (p < 0.05). However, at 3 and 5 years, event-free patients had borderline higher platelet counts (p = 0.056, p = 0.078) and hemoglobin (p = 0. 097, p = 0.061). The coefficient of variation as an expression of the variability of the results in each group was systematically lower at 3, 5 and 10 years in the group of event-free patients. Despite some differences between the two groups, our data support the hypothesis that patients with long lasting remissions should not be considered as definitively cured of aplastic anemia. PMID- 10705161 TI - Neuropathological substrates of dementia and depression in vascular dementia, with a particular focus on cases with small infarct volumes. AB - The neuropathological substrates of dementia and depression were evaluated in 30 patients with cerebrovascular disease and significant cognitive impairment (VaD), with a particular focus on patients with small infarct volumes (<15 ml). VaD patients with small infarct volumes had a similar degree of cognitive impairment to those with larger infarct volumes (>15 ml) but were significantly more likely to be depressed and to have areas of microinfarction. A review of individual cases with small infarct volumes suggested that the combination of microinfarction, diffuse white matter disease and perivascular changes, or the overlap of neurodegenerative pathologies and microvascular changes were particularly important. Microinfarction was also significantly associated with major depression. PMID- 10705162 TI - Absorption of aluminium-26 in Alzheimer's disease, measured using accelerator mass spectrometry. AB - Although chromosomal abnormalities underpin some early onset cases of familial Alzheimer's disease (AD), most cases are sporadic and not associated with such abnormalities. Aluminium (Al) is a significant but controversial risk factor for sporadic AD, and studies have reported associations between Al and the principal pathological features of AD, senile plaques and neurofibrillary tangles. The present study measured gastrointestinal (GI) absorption of Al under normal dietary conditions using (26)Al tracer and accelerator mass spectrometry (AMS). Following overnight fast, 13 AD patients (aged 63-76 years) and 13 age-matched controls (aged 62-76 years) ingested a fruit drink containing 27 ng (26)Al. Plasma samples were obtained before and 1 h after the drink and from these the fraction of (26)Al absorbed across the GI tract was estimated. The GI tract rigorously excludes Al with only 0.06-0.1% of the ingested Al being absorbed. The mean fraction absorbed by AD subjects exceeded controls by a factor of 1.64 (p/=90% were attained by 75 and 85% of the implant group, respectively, but none of the control group. One third of implant patients, but none of the control patients, achieved >/=50% improvement in leakage severity. Over half of the implant patients (56%) were completely dry compared with 1 control patient (4%). Implant patients, but not control patients, exhibited significant improvement with respect to two quality of life measures. Neuromodulation resulted in increases of 220% (p < 0.0005) and 39% (p = 0.013), respectively, in urodynamically assessed bladder volume at first contraction and maximum fill. At 36 months the actuarial rate of treatment failure was 32.4% (95% CI, 17.0-56.0%). Adverse events most frequently involved pain at the implant site, and the incidence of serious complications was low. CONCLUSIONS: Neuromodulation is markedly more effective than conservative management in alleviating symptoms of refractory urge incontinence. Quality of life and urodynamic function are also improved by neuromodulation. The effects of neuromodulation are long-lasting, and associated morbidity is low. PMID- 10705195 TI - Increased nitric oxide production in the spermatic vein of patients with varicocele. AB - OBJECTIVE: To define the level of nitric oxide (NO) in the spermatic vein of patients with varicocele and its relation with male infertility. MATERIALS AND METHODS: Following physical and color Doppler ultrasonographic examination, whole blood samples were drawn from a peripheral vein and a dilated varicocele vein from fourteen patients with clinically palpable varicocele (G2-3) before ligation. NO levels in the serum were determined as total nitrite by Greiss reaction and results were compared with Mann-Whitney U test. RESULTS: NO levels in the internal spermatic vein were 36.05 +/- 8. 92 micromol/l, compared to 19.41 +/- 4.12 micromol/l in the peripheral vein and the difference was statistically significant (p < 0.01). CONCLUSION: In view of our results, increased NO levels in the dilated varicocele vein might be responsible for spermatozoa dysfunction. PMID- 10705196 TI - Immunohistochemical and quantitative study of mast cells in La Peyronie's disease. AB - OBJECTIVE: In order to investigate the possible association between mast cells (MCs) and the fibrous plaque of La Peyronie's disease, the number of MCs in normal penile tissue and in the fibrous plaque was determined. METHODS: The control group consisted of 5 total and 3 partial penectomies with no fibrotic lesions, while the study group consisted of 23 excisional biopsies from cases of La Peyronie's disease dating back to at least 2 years earlier and with no signs of activity. The biopsies included tissues from the tunica albuginea (TA), the areolar tissue (Br) between the tunica and the erectile tissue (CC) and from the latter. The number of MCs was counted with the aid of an image analysis program following staining the antibody antitryptase. RESULTS: In the cases of La Peyronie's disease the number of MCs/mm(2) was significantly higher in the TA and Br but lower in the CC. The MCs were related to fibroblasts and vasculonervous channels in the TA, and were concentrated around the fibrous plaques and granulation tissue between the TA and BR and between the latter and the CC. CONCLUSION: Our data indicate that MCs have a role in the genesis of the fibrous plaque in the TA and in the persistent inflammation in the Br. Medical treatment aimed at repressing MC activation and proliferation locally might be useful in La Peyronie's disease. PMID- 10705197 TI - History of 7,093 patients with lower urinary tract symptoms related to benign prostatic hyperplasia treated with alfuzosin in general practice up to 3 years. AB - OBJECTIVES: As we have previously published 4 articles reporting the treatment of 7,093 clinical benign prostatic hyperplasia (BPH) patients treated with alfuzosin in a 3-month open-labelled study which was subsequently extended to 12, 24, and 36 months, the objective of this article is to provide additional data on dropouts, acute urinary retention (AUR), progression to surgery, and safety under the natural conditions of general practice, paying special attention to the predictive factors. METHODS: 7,093 patients were initially enrolled by 1,812 centers for up to 3 months. Subsequently 1,508, 1,325, and 812 general practitioners agreed to extend the study up to 12, 24, and 36 months, respectively, which corresponds to 4 patient populations. RESULTS: The baseline symptom profile of patients who completed the study was identical to that of patients who dropped out (because the center resigned or during treatment). In the 4 patient populations, the percentage of patients per month who dropped out, experienced adverse effects, AUR and surgery were 0. 6-1.6, 0.1-0.5, 0.01-0.03, and 0.1-0.3%, respectively. The classes of symptom severity were not predictive for dropouts: 3.5, 12.6, 20, and 14.3% of the severe patients dropped out during treatment versus 4.2, 13.7, 22.9, and 14.0% of the moderate patients who dropped out up to 3, 12, 24, and 36 months, respectively. Safety was satisfactory regarding the number of adverse events and blood pressure measurement. No retrograde ejaculation was reported. CONCLUSION: Under the natural conditions of general practice the reasons for dropping out were not correlated with symptom severity. PMID- 10705198 TI - Community study of uncomplicated lower urinary tract symptoms among male Italien immigrants in Sydney, Australia. AB - OBJECTIVE: To determine the prevalence, levels of bother and self-reported management of lower urinary tract symptoms (LUTS) in Italian-born men aged 40-80 years. METHOD: 305 randomly selected men aged 40-80 years (72% response rate) participated in a community-based study (computer-assisted telephone survey) in early 1997 in Sydney, Australia. RESULTS: LUTS are common: 41% of men needed to wake up at least once at night to urinate; 35% indicated they had terminal dribbling 'sometimes' or 'frequently'; 31% experienced urgency although few (3%) had urge incontinence; 19% could retain urine in their bladder during the day for no more than 2 h, and 19% experienced hesitancy. Urinary symptoms correlated moderately/poorly with self-rated bother. The prevalence of bothersome frequency and urgency was significantly age-related. Only half (n = 49, 52%) of the men bothered by urinary symptoms had seen a general practitioner (GP) about these symptoms in the last 5 years: of these three quarters had been referred to a urologist and half of these had received surgical treatment. Anxiety about prostate cancer, but not the degree of bother from urinary symptoms, independently predicted attendance at a GP (adjusted odds ratio 6.4, p = 0.006). CONCLUSIONS: Although LUTS are common in Italian-born men, their experiences of bother do not correlate well with symptoms and do not appear to influence referral and treatment. Education is needed to improve men's understanding of the importance of 'bother' as an indicator for urological surgery. PMID- 10705199 TI - Risk factors in prostatectomy bleeding: preoperative urinary infection is the only reversible factor. AB - OBJECTIVES: To find out whether any of a preset battery of clinical and laboratory parameters has significant correlation with blood loss caused by transurethral prostatectomy (TURP). PATIENTS AND METHODS: All new patients undergoing a TURP over a 1-year period were included in the study. For each patient the following parameters were documented: (1) Pre-operatively: Age, mode of presentation, blood pressure, complete blood count, coagulation screening, calcification of abdominal blood vessels on plain X-ray, prostate size on digital rectal examination (DRE), pre-operative urine culture, presence of haematuria, ECG changes and drugs taken. (2) Intraoperatively: prostate morphology at cystoscopy, type of anaesthesia, operating surgeon, operating time, weight of resected prostate tissue and any blood transfusions. (3) Postoperatively: Length of irrigation time and prostate histology. All the intra-operative and postoperative blood loss was calculated. Their sum constituted total blood loss which constituted the dependent variable of the study. Statistical methods used included bivariate correlation, Students t test or the chi(2) test as appropriate. RESULTS: Of 140 TURPs, 121 were eligible for final analysis. The mean total blood loss was 552 ml (34 ml per gram of tissue) and the mean resected prostate weight was 18 g. Of the continuous variables, the one with strongest correlation with total blood loss was resected prostate weight (r = 0.47, p = 0.000) followed by prostate size on DRE (r = 0.45, p = 0.000) then operating time (r = 0.39, p = 0.000). Of the categorical variables, the one with the strongest correlation with total blood loss was preoperative urine culture (p = 0.01). CONCLUSIONS: The only reversible factor associated with blood loss caused by TURP is pre-operative urinary infection. Operating time is partly controllable, as it may improve (i.e. decrease) with experience of the operator. PMID- 10705200 TI - Comparison of an LH-RH analogue (Goeserelin acetate, 'Zoladex') with combined androgen blockade in advanced prostate cancer: final survival results of an international multicentre randomized-trial. International Prostate Cancer Study Group. AB - OBJECTIVE: The aim of this study was to compare the effects of the nonsteroidal antiandrogen flutamide plus the LH-RH analogue goserelin acetate (combined androgen blockade [CAB]) with goserelin acetate alone in patients with advanced prostate cancer. The original analyses at 25 and 56 months of follow-up have been reported previously, and here we report the final survival analysis after 10 years of follow-up. METHODS: 589 patients with advanced prostate cancer (55% with metastatic [M1] and 45% with locally advanced [M0] disease) were randomized to receive goserelin acetate 3.6 mg either alone or in combination with flutamide (250 mg three times daily). RESULTS: A total of 583 patients were included in the analysis. There was a small, but nonsignificant, benefit for CAB compared with goserelin acetate alone in all patients with respect to survival (hazard ratio 0.88, 95% CI 0.73, 1.06). Subgroup analysis of M0 and M1 patients showed similar results (M0: hazard ratio 0.92, 95% CI 0.68, 1.25; M1: hazard ratio 0.85, 95% CI 0.66, 1. 08). The treatment effect was not significantly different for M0 and M1 patients (p = 0.685). CONCLUSIONS: In this large randomized trial containing significant numbers of M0 patients, after 10 years there was a small but nonsignificant benefit for CAB over castration alone. PMID- 10705201 TI - Prognostic significance of Ki-67 expression in advanced prostate cancers in relation to disease progression after androgen ablation. AB - OBJECTIVE: This study was performed to examine the expressions of Ki-67 antigen, bcl-2 and p53 oncoprotein and to assess their capacity to predict the short-term response to the initial endocrine treatment and disease progression in prostate cancer. METHODS: Formalin-fixed and paraffin-embedded tumor tissues from a total of 73 patients with untreated prostate cancers were collected by transrectal ultrasound-guided biopsy. All patients with stage C or D prostate cancers had received continuous endocrine treatment. Ki-67 antigen, p53 and bcl-2 oncoprotein were detected by immunohistochemical methods. RESULTS: Short-term response to initial endocrine therapy judged at 3 months showed a significant correlation with Ki-67 labeling index (LI) and bcl-2 expression. Multivariate analysis showed that only a high Ki-67 LI was an independent potential predictor of prostate specific antigen failure or the appearance of new metastasis. CONCLUSIONS: The findings suggested that the Ki-67 LI was the most useful parameter to predict disease progression after the initial endocrine treatment. Additional studies must be performed to evaluate the prognostic significance of both cell proliferation and apoptosis in detail. PMID- 10705202 TI - Prostate-specific antigen levels and clinical response to flutamide as the second hormone therapy for hormone-refractory prostate carcinoma. AB - BACKGROUND: Some authors have recently reported that maximum androgen block (MAB), in which the nonsteroidal anti-androgen, flutamide, is used together with conventional hormone therapy such as castration or luteinizing hormone-releasing hormone analogue, is more effective for prostate cancer than conventional methods. However, others have reported that the effect of MAB on survival is minimal, and definite conclusions concerning MAB remain unclear. Conversely, using flutamide as a second-line hormone therapy after recurrence is also considered, but few authors have reported whether this therapeutic option is effective or for which patients it is effective. MATERIALS AND METHODS: 124 patients with prostate cancer were diagnosed and followed at Kobe City General Hospital between 1995 and 1997. Twenty-two of these cases developed recurrence during first-line hormone therapy, and flutamide was prescribed in these cases. The prognostic value and effectiveness of flutamide were evaluated by measurement of serum prostate-specific antigen (PSA) at diagnosis, posttreatment nadir PSA level, PSA at the time of flutamide use, histological grade, recurrence-free time after firstline hormone therapy and age at the time of diagnosis. RESULTS: Six of 9 cases whose post-treatment nadir PSA levels after initial hormone therapy were within the normal limit (<4 ng/ml) achieved complete remission (CR) with flutamide use, but no patient whose post-treatment nadir PSA level remained elevated achieved CR. PSA at diagnosis and PSA at the start of flutamide use were significantly lower for patients with CR. However, the results of multivariate logistic regression analysis demonstrated that only the post-treatment nadir PSA level was significantly correlated with prognosis of flutamide use. CONCLUSIONS: Flutamide use as second-line hormone therapy should be limited to cases in which first-line hormone therapy has been highly effective and for whom the post treatment nadir PSA level was within normal limits, and other patients should undergo other therapies. By limiting flutamide use to patients in whom the effect of flutamide is considered to be maximal, the incidence of complications and medication costs can be decreased. PMID- 10705203 TI - Urinary continence following radical prostatectomy assessed by a self administered questionnaire. AB - OBJECTIVES: A total aim of this study was to assess the incidence of urinary incontinence in patients following radical prostatectomy and determine the factors that may influence this incidence. METHODS: A total of 135 men underwent radical retropubic prostatectomy at our center between 1987 and 1997. 120 patients were sent a questionnaire regarding preoperative and postoperative voiding habits. Data collected included preoperative and postoperative continence status, interval to postoperative continence status, associated urinary symptoms, willingness to undergo radical prostatectomy again if need be and additional postoperative procedures. Patient age, date of surgery, number of neurovascular bundles resected at prostatectomy and duration of follow-up were also noted. RESULTS: Of the 120 patients, 116 (96.7%), a mean of 65.2 (range 48-76) years old, responded to the questionnaire. Mean follow-up was 4.3 years (range 1-10.8). Continence was defined as no regular use of pads. Our overall urinary incontinence rate was 14.4%. Of the respondents, 88. 8% (103/116) had achieved final continence status by 6 months postoperatively, and 95% (110/116) would undergo surgery again if need be. Of the patients considered incontinent postoperatively, 66. 6% had associated urgency. Age, year of surgery, number of neurovascular bundles resected at prostatectomy, preoperative urinary leakage of postvoiding dribbling, postoperative pelvic floor exercises, and anastomotic stricture had no significant impact on postoperative continence status. CONCLUSIONS: Using an anonymous self-administered questionnaire, we found a 14.4% incontinence rate after radical prostatectomy. These results allow patients to have realistic expectations when counseled prior to this operation. PMID- 10705204 TI - Identification of a novel microsatellite marker tightly linked to the KAI-1 gene for predicting prostate cancer progression. AB - BACKGROUND: We have mapped the human prostate-specific membrane antigen (PSM) gene to the chromosome 11p11.2 region at 62.5 cM, a region which also contains the prostatic cancer metastasis suppressor gene KAI-1. The genetic marker D11S1344 has been utilised for loss of heterozygosity (LOH) studies on the KAI-1 gene in a large series of prostate cancer specimens. The results were negative and it was concluded that deletions of the KAI-1 gene were not involved in the development of the metastatic phenotype in these tumours. One possible explanation for this result could be that D11S1344 is not sufficiently tightly linked to the KAI-1 gene to detect small deletions. OBJECTIVE: To attempt to identify a genetic marker more tightly linked to the KAI-1 gene than D11S1344. METHODS: Yeast artificial chromosome (YAC) clones containing the KAI-1 gene and the neighbouring marker D11S1344 were analysed by the fluorescent in situ hybridisation technique. The human genomic inserts in these novel clones were sized by pulsed field gel electrophoresis. For more accurate mapping of the KAI-1 gene, YACs containing it were screened for polymorphic markers (including D11S1344) from the 11p11.2 region. RESULTS: The novel YAC clones localised exclusively to the 11p11.2 region, with single hybridisation signals compared to the dual signals consistently obtained with nearby PSM-containing YACs. All the KAI-1 clones found had small inserts (<300 kb). The only known microsatellite which gave amplification products with these YACs was D11S986 which has been mapped at 61.3 cM on human chromosome 11. CONCLUSIONS: We have precisely localised KAI-1 at 61.3 cM on human chromosome 11. This is some 1.2 cM away from the previously utilised LOH microsatellite marker, D11S1344. We suggest that the very tightly linked microsatellite D11S986 may be a more accurate marker to assess LOH of the KAI-1 gene and thus predict progression of prostate cancer. The region of genetic duplication around the PSM gene does not extend as far distally on 11p as KAI-1. PMID- 10705206 TI - Erectile dysfunction and anejaculation PMID- 10705205 TI - Polymorphisms in the vitamin D receptor gene and the androgen receptor gene and the risk of benign prostatic hyperplasia. AB - OBJECTIVE: Little is known about risk factors for the development of benign prostatic hyperplasia (BPH). Recently, associations were observed between prostate cancer (CaP) risk and polymorphisms in the vitamin D receptor (VDR) gene and the androgen receptor (AR) gene. Since both receptors are relevant for prostate growth, the VDR and AR are also expected to be involved in the development of BPH. The objective of this study is to establish the relationship between the risk of BPH and a polymorphism in the number of CAG repeats in the AR gene and a TaqI restriction enzyme polymorphism in the VDR gene. METHODS: For this study, 98 patients who had been treated for BPH-related complaints and 61 convenience controls (predominantly bladder cancer patients) were recruited from the outpatient clinic. DNA was isolated from peripheral blood, and genotyping was performed with PCR-based methods. Means as well as odds ratios (ORs) with 95% confidence intervals (CI) were calculated using SPSS software. RESULTS: The mean number of CAG repeats in the AR gene in patients and controls was found to be similar: 21.8 (SD = 2.8) and 21.9 (SD = 2.9), respectively. In the subgroup of patients with a prostate volume of at least 50 cm(3), the mean number of repeats was 21.5 (SD = 2.6). The OR for BPH for individuals with homozygous presence of the VDR TaqI restriction fragment length polymorphism (RFLP) (tt) versus individuals with homozygous absence (TT) or heterozygotes (Tt) was found to be 1.0 (95% CI 0.4-2.4). For individuals with a prostate volume of at least 50 cm(3), the OR was 1.2 (95% CI 0.5-3. 2). CONCLUSION: Unlike earlier observations in prostate cancer, we did not find an association between the CAG repeat polymorphism in the AR gene and the TaqI RFLP polymorphism in the VDR gene and the risk of BPH. PMID- 10705207 TI - Treatment of heterotopic cervical and intrauterine pregnancy. AB - OBJECTIVE: To find a suitable technique to selectively terminate a cervically implanted embryo while maintaining viability of a concomitant intrauterine pregnancy. METHODS: A 34-year-old patient achieved a twin pregnancy after 4 IVF attempts. Ultrasound revealed a viable intrauterine and cervical pregnancy. Given our experience with KCl injection for fetal reduction, we offered the patient an attempt to reduce the cervical pregnancy. RESULTS: Best visualization in this case was obtained by transabdominal scanning. A 6-inch 20-gauge spinal needle was inserted transcervically and maneuvered into the thorax of the embryo. Fetal heart rate ceased even before KCl could be injected. Then 3 cm(3) of saline were injected to provide better visualization of the cervical fetus, and to confirm absence of heart beat. The patient had minor vaginal bleeding for several days. The intrauterine pregnancy progressed uneventfully through 36(1)/(2) weeks with delivery of a healthy, 2, 700-gram newborn. CONCLUSION: Cervical pregnancy is usually considered a life-threatening event. Other factors such as concomitant intrauterine pregnancy and the patient's infertility history generally would be secondary concerns. In this case, we were able to selectively terminate the cervical pregnancy, while preserving the intrauterine one, allowing this couple to have a healthy newborn. Further cases will be necessary to appropriately define risk rates for such an approach. PMID- 10705208 TI - Intra-uterine fetal demise caused by amniotic band syndrome after standard amniocentesis. AB - The amniotic band syndrome represents a prime example of exogenous disruption of an otherwise normal fetal development. It may be a sequel of invasive diagnostic procedures such as amniocentesis or fetal blood sampling. A 38-year-old gravida II, para II delivered a morphologically normal male stillborn at term. The pregnancy history had been unremarkable but for an early 2nd-trimester amniocentesis. Cause of the intra-uterine fetal demise was noted to be an amniotic band constricting the umbilical cord. An amniotic band is a rare but potentially fatal condition which may be induced by, e.g., invasive prenatal procedures. Such bands are not usually diagnosed prenatally; however, selected patients with augmented risk may profit from intensive ultrasound evaluation including Doppler studies. PMID- 10705209 TI - Intrauterine growth retardation and fetal cardiac function. AB - Intrauterine growth retardation is a pathology which is found in 3-10% of all pregnancies and it is associated with around 20-25% of all fetal intrauterine deaths and with long-term neurologic sequelae. It presents an increased risk of distress during labor and delivery and a greater risk of perinatal mortality. The causes of IUGR and the cardiac and venous Doppler in normal fetuses are analyzed, and the hemodynamic cardiac modifications in IUGR fetus are discussed. The fetal cardiac function in intrauterine growth retardation shows a redistribution of the fetal cardiac output, which tends to favor the left ventricle as the mechanism to compensate for the uteroplacental insufficiency. The Doppler velocity indices are modified as the fetal condition progressively deteriorates and they represent an important tool for the management of the complicated pregnancy. PMID- 10705210 TI - Biometry of the fetal heart between 10 and 17 weeks of gestation. AB - OBJECTIVES: Assessment of the dimensions of the cardiac chambers and the great arteries in the human fetus may be helpful in the prenatal diagnosis of congenital heart disease. The purpose of this prospective cross-sectional study was to compile normative data in fetal cardiac measurements in early pregnancy. The structure of the fetal heart was examined in 136 normal singleton fetuses between 10 and 17 weeks of gestation. METHODS: The transversal heart diameter, both ventricular dimensions, interventricular septal thickness, heart area, heart circumference, thoracic diameter, thoracic circumference and thoracic area were measured in the four-chamber view during diastole. Diameters of the pulmonary trunk and ascending aorta were obtained in the short axis and long axis view during systole. Ultrasound examinations were performed with a 5.0-MHz transvaginal and/or transabdominal phased-array sector scanner. RESULTS: The four chamber view and the cross-over of the pulmonary artery and the aorta were adequately visualized in 44% of the fetuses at 10 weeks of gestation, in 75% at 11 weeks of gestation, in 93% at 12 weeks of gestation and in 100% of the fetuses at 13-17 weeks of gestation. Before 14 weeks of gestation transvaginal sonography was superior to the transabdominal sonography in visualization of the fetal heart and great arteries. After 14 weeks of gestation transabdominal sonography accurately demonstrated the structure of the fetal heart. The ratio of right and left ventricle (RV/LV) and the ratio of the pulmonary trunk and aorta (PT/AO) were constant during this period of gestation (approximately 1.00 and 1. 10, respectively). The ratio of the cardiac and thoracic area showed only a slight increase with advancing gestational age, but with significant correlation. The fetal heart rate showed a slow decrease from 167 to 150 bpm in this period of gestation. The transversal heart diameter, both ventricular dimensions, interventricular septal thickness, heart area, cardiothoracic diameter ratio, aortic diameter and the pulmonary trunk diameter showed a highly significant linear correlation to the gestational age and the biparietal diameter. CONCLUSION: The advancing quality of ultrasound images allows fetal echocardiography in the first and early second trimester. Our normative data could be the basis of studying the development of cardiac structures in congenital heart disease and it might be helpful in the detection of some congenital heart defects in early pregnancy. PMID- 10705211 TI - Prenatal diagnosis of a subamniotic hematoma. AB - A case of a subamniotic hematoma was diagnosed at 34 weeks of gestation. Pregnancy and delivery were uneventful. The ultrasound features of a subamniotic hematoma, and the differential diagnosis with lesions of less favorable outcome are described. PMID- 10705212 TI - Effect of steroids on arterial Doppler in intrauterine growth retardation fetuses. AB - BACKGROUND: Antenatal steroid administration decreases fetal mortality and morbidity in preterm neonates. However, maternal steroids administration can cause a transient reduction in fetal heart rate variability and can also alter biophysical profile parameters. Other methods to assess fetal well-being in high risk fetuses are therefore needed, when steroids are given in high-risk pregnancies. OBJECTIVE: To study the effect of betamethasone and dexamethasone on placental and fetal arterial circulation in 40 growth-retarded fetuses. METHODS: Forty fetuses with intrauterine growth retardation were exposed in utero to a single course of betamethasone (n = 25) or dexamethasone (n = 15). Blood flow waveforms were recorded from uterine arteries, umbilical arteries, descending aorta and middle cerebral arteries and analyzed prospectively. Doppler recordings were compared before (baseline), during (24-48 h) and after treatment (4-7 days). RESULTS: No significant changes were found in the pulsatility indices (PI) in any of these vessels during the course and after the treatment was stopped. However, the PI in the middle cerebral arteries showed a trend to decrease 24-48 h and 4-7 days after steroids were given to the mother when compared to pretreatment values. CONCLUSION: Since antenatal steroids do not seem to affect Doppler measurements, they might be useful to assess well-being in high-risk fetuses during the course. It might allow to differentiate transient iatrogenic changes in fetal heart rate variability from the effect of hypoxemia. PMID- 10705213 TI - Sonographic measurement of the fetal iliac angle cannot be used alone as a marker for trisomy 21. AB - The fetal iliac wings angle was studied in 255 fetuses before amniocentesis at 16.7 weeks (+/- 1.3), using a sonographic axial view of the fetal pelvis. The measurement could be performed in 208 fetuses (81.6%), of whom 4 had trisomy 21 (T 21). The mean iliac angle was greater in fetuses with T 21 than in normal fetuses (69.8 degrees vs. 88.7 degrees; p = 0.03). This measurement is subject to significant intra- and interexaminer variability (interclass correlation coefficient: 0.65 and 0.23, respectively). When a 90 degrees value is used as a threshold, specificity, sensitivity, positive and negative predictive values are, respectively, 80, 75, 7. 0 and 99.4%. The 20% rate of false-positives rules out the use of this measurement as the sole criterion for the indication of amniocentesis for T 21 antenatal diagnosis. PMID- 10705214 TI - Severe oligohydramnios with intact membranes: an indication for diagnostic amnioinfusion. AB - OBJECTIVE: To quantify the improvement in ultrasonographic fetal imaging following diagnostic amnioinfusion for the indication of unexplained midtrimester oligohydramnios. METHODS: Patients referred for unexplained midtrimester oligohydramnios were retrospectively reviewed. Videotapes of those undergoing diagnostic antenatal amnioinfusion were analyzed for quality of visualization of routinely imaged structures before and after the infusion procedure. RESULTS: The overall rate of adequate visualization of fetal structures improved from 50.98 to 76.79% (p < 0.0001). In fetuses having preinfusion-identified obstructive uropathy, there was improvement in identification of associated anomalies from 11.8 to 31.3%. CONCLUSIONS: Several authors have suggested that diagnostic amnioinfusion can facilitate fetal imaging and increase diagnostic precision in the setting of unexplained severe oligohydramnios. We have quantified the improvement in the rate of optimal visualization of fetal structures which likely translates, in experienced hands, into this observed improved diagnostic precision. Of particular importance is the improvement in appreciation of associated anomalies in cases of obstructive uropathy in which such findings may determine whether or not invasive fetal therapy is indicated. PMID- 10705215 TI - Does amniotomy influence the prognosis of babies in cases with severe chorioamnionitis? Report of a twin pregnancy with varying outcome. AB - We report our experience in a woman with a twin pregnancy. The patient suffered severe Escherichia coli chorioamnionitis and the outcomes were different between the two babies after birth. The first baby had only a mild infection, but the second suffered sepsis and subsequent perinatal death. These differences in outcome appeared to be due to amniotomy performed for the first baby after late labor stage I to augment uterus contractions. Removal of infectious amniotic fluid from the amniotic cavity might thus have prevented the spread of the chorioamnionitis. E. coli sometimes causes severe infection during pregnancy and the perinatal period. In this case, a large number of enteropathogenic E. coli (serotype O-6) was cultured from blood, stool, pharyngeal swab, gastric juice and puncture fluid from the thoracic cavity of the second baby. O-6 is classified an enterotoxigenic strain mainly causing diarrhea because of endotoxin released from bacteria. O-6 has not hitherto been reported as a cause of severe infection in chorioamnionitis and perinatal sepsis. PMID- 10705216 TI - Antenatal diagnosis of cardiac malformation: a structural study. AB - The purpose of this study was to find out the functional and anatomical relationship between the various cardiac segments in congenital heart disease before and after birth. This study of cardiac pathology focused on 227 fetuses referred to fetal echocardiography because of suspicion of congenital heart defects (CHD) from 1986 to 1991. The heart was imaged and classified according to a division into 7 adjacent segments. Normality or abnormality of each segment and of the general state of the subject's heart was coded in a binary mode, in which 0 = normal and 1 = pathological. We also coded the extracardiac morphology, noting existence or absence of another extracardiac defect in the fetus. The binary segmental data were entered into a worksheet for further analysis. Observations were conducted prenatally and verified after birth or abortion. Fetal echocardiography was first assessed using conventional statistical analysis. The calculated accuracy and predictive values to diagnose any cardiac defect as well as for correctly coding abnormal or normal cardiac segments were excellent. Then, using HUDAP software (Hebrew University Data Analysis Package), we could demonstrate a perfect fit between the functional and pathological structure of the cardiac segments in the fetus. Moreover, using this unique type of data analysis, we found a multidimensional scaling confirmation of the embryogenesis theory of the heart. PMID- 10705217 TI - De novo partial duplication of 3q and distal deletion of 20p in a 15-week abort us with omphalocele. PMID- 10705218 TI - CpG DNA as a Th1 trigger. AB - Over the last few years, it has been recognized that along with structural components and products of bacteria, bacterial DNA is also capable of signaling infectious danger to cells of the innate immune system. Particular DNA sequences (CpG motifs), which are abundant in prokaryotic (bacterial) but not in mammalian DNA, cause the activation and stimulation of immune cells. Research has been catalyzed by the finding that certain synthetic oligodeoxynucleotides mimic the action of bacterial DNA. Immunostimulation induced by bacterial DNA or synthetic oligonucleotides not only contributes to our knowledge of the pathogen-host interrelationship during infection, but can also be used therapeutically to condition or modify ongoing immune responses of the adaptive immune system. Accordingly, CpG motifs have been used as vaccine adjuvants as well as instructing agents to selectively induce Th1-dominated immune responses. Hence, CpG motifs might be used in the future as adjuvants and/or immunomodulatory agents to treat or prevent undesired Th2-dominated immune responses, such as allergy. PMID- 10705219 TI - Latex allergy in children. AB - Natural rubber is a component of the latex of the tropical Hevea brasiliensis tree which is widely used in the manufacturing of medical devices and a large variety of articles for everyday use. Over a dozen allergens have been identified in the latex of H. brasiliensis. The allergens Hev b 1, Hev b 3, Hev b 6, and Hev b 7 are proteins that are involved in the biosynthesis of rubber or the coagulation of latex. Allergens that are part of the plant's defense system are represented by Hev b 2 and class I endochitinases. The allergens Hev b 4, Hev b 5, and Hev b 8-10 were classified as either structural or housekeeping proteins. Immediate-type hypersensitivity reactions to proteins present in Hevea latex were first described in 1927. Since then, natural rubber latex (NRL) allergy has become an important medical problem for an increasing number of individuals. Sensitization mainly occurs by wound or mucosal contact with NRL devices during surgery or by inhalation of airborne allergens released from powdered latex gloves. The number of surgical interventions and an atopic disposition are the most important risk factors for developing latex allergy, especially in children with spina bifida. Exposure to NRL products should be carefully avoided for individuals who belong to high-risk groups. Initial studies on establishing a latex-free environment for surgery in all spina bifida patients have reported on a decrease in sensitization and allergy to NRL. PMID- 10705220 TI - Suppression of specific IgE antibody responses by liposome-conjugated ovalbumin in mice sensitized with ovalbumin via the respiratory tract. AB - BACKGROUND: Previously we have shown that intranasal administration of ovalbumin (OVA) together with cholera toxin (CT) abrogates nasal tolerance to OVA, resulting in the induction of specific IgE antibody (Ab) responses, and that intraperitoneal injection of OVA coupled with liposomes (OVA-liposomes) induces a selective suppression of IgE Ab responses to OVA. Whether OVA-liposomes suppress anti-OVA IgE Ab responses in mice sensitized with CT-combined OVA via the respiratory tract remains to be clarified. METHODS: In some experiments, mice were given OVA, liposomes or OVA-liposomes with or without CT intranasally three times, at 2-week intervals (weeks 0, 2 and 4). In other experiments, mice were given OVA-liposomes intranasally 2 days before or 1 and 3 weeks after CT-combined OVA (week 0), which was administered intranasally three times, at 2-week intervals (weeks 0, 2 and 4). Two weeks after the third administration of CT combined OVA (week 0), nasal wash and serum IgA, IgG and IgE Ab responses were assayed. RESULTS: Pretreatment with OVA-liposomes suppressed IgE Ab responses to CT-combined OVA, with a significantly high production of both nasal IgA and serum IgG Abs. Moreover, treatment with OVA-liposomes 1 and 3 weeks after CT-combined OVA administration also suppressed IgE Ab responses. The suppression of anti-OVA IgE Ab production by OVA-liposomes was accompanied by a simultaneous enhancement of specific IgA and IgG (IgG1, and especially IgG2a) Ab production. CONCLUSIONS: Postimmunization treatment with OVA-liposomes, as well as preimmunization treatment, suppressed specific IgE Ab responses in mice sensitized intranasally with CT-combined OVA. Allergens conjugated to liposomes may be appropriate for preventing the development of allergies to inhaled or dietary antigens in humans. PMID- 10705221 TI - Effect of suplatast tosilate (IPD-1151T) on a mouse model of asthma: inhibition of eosinophilic inflammation and bronchial hyperresponsiveness. AB - BACKGROUND: Suplatast tosilate (IPD) is a newly developed 'anti-allergic' drug. It seems to be a unique compound because of its ability to suppress IgE but not IgG or IgM production in vivo and cytokine production from type 2 helper T cells (Th2) in vitro. However, information on its in vivo effect on an animal model of asthma is limited. METHOD: BALB/c mice sensitized to ovalbumin (3 times, 2-week interval) were challenged with ovalbumin by inhalation (50 mg/ml for 20 min, once a day for 6 days). In this study, we explored the influence of IPD on eosinophil infiltration into the airways, bronchial hyperresponsiveness (BHR) to methacholine, specific IgE antibody production, and cytokines in bronchoalveolar lavage fluid (BALF) using this murine model. RESULTS: Treatment with IPD significantly reduced the number of total cells and eosinophils in BALF (around 40%) and almost completely inhibited the development of antigen-induced BHR. Histological findings confirmed the reduction of submucosal cell infiltration in the lung, and disclosed the marked inhibition of bronchial epithelial cell damage. Ovalbumin-specific IgE was slightly but significantly reduced. The levels of IL-4, IL-5 and IL-13 in BALF were significantly decreased in mice treated with the compound compared to those in untreated mice. CONCLUSION: These results suggest that IPD is capable of inhibiting the production of Th2 cytokines, which inhibit eosinophil infiltration into the murine airway, IgE synthesis, and development of BHR, in a murine model of asthma. PMID- 10705222 TI - Once-daily theophylline reduces serum eosinophil cationic protein and eosinophil levels in induced sputum of asthmatics. AB - BACKGROUND: Because eosinophilic airway inflammation is a characteristic feature of bronchial asthma, the treatment of airway inflammation is important in the management of asthma. Theophylline has been reported to reduce airway inflammation, in addition to its well-known bronchodilating effect. OBJECTIVE: In order to evaluate the effects of theophylline on airway inflammation, we investigated 48 subjects with mild and moderate asthma. METHODS: The patients were randomly divided into two groups, with or without theophylline treatment (control n = 24; theophylline, n = 24). We examined the level of serum eosinophil cationic protein (ECP), induced sputum samples, and peak expiratory flow (PEF) and obtained spirograms before and after 4 weeks of treatment with once-daily theophylline (200-600 mg/day) of subjects with mild or moderate asthma. RESULTS: Theophylline significantly increased morning and evening PEF and significantly decreased the diurnal variation of PEF. After treatment with theophylline, both serum ECP and the percentage of eosinophils in induced sputum were significantly decreased. In contrast, peripheral blood eosinophil count was unchanged after treatment with theophylline. CONCLUSIONS: These findings suggest that theophylline reduces airway inflammation and the severity of asthma, presumably via suppression of both eosinophil activity and subsequent eosinophil infiltration of the airways. PMID- 10705223 TI - Increased eosinophil cation protein level in sensitized nonasthmatics is linked to subsequent hyperresponsiveness to methacholine. The Odense Schoolchild Study. AB - BACKGROUND: Increased levels of eosinophil cation protein (ECP) in sensitized subjects may reflect early stages of an ongoing inflammatory process and therefore precede asthma and bronchial hyperreactivity. AIM: To study whether nonasthmatic subjects with sensitization to allergens and increased ECP levels are at a higher risk for subsequent increased bronchial reactivity compared with sensitized nonasthmatics with normal ECP levels. METHODS: A prospective study of 240 schoolchildren with a mean age of 13.9 years (range: 12.6-15.9) who were followed up after 6.3 years. Bronchial reactivity was assessed by methacholine provocation testing. Sensitization was defined by one or more positive reactions (>3 mm wheal) to 10 common aeroallergens by skin prick testing. Increased ECP was defined as values above 20 microg/l. This separated the subjects into four categories: group 1: healthy controls without sensitization (n = 147); group 2: sensitized subjects with a serum ECP below 20 microg/l (n = 55); group 3: sensitized subjects with an ECP level at or above 20 microg/l (n = 16), and group 4: all asthmatics (n = 22). RESULTS: Bronchial reactivity was similar in subjects of groups 2 and 3 at baseline (p = 0.8). Six years later, subjects from group 3 were more responsive to methacholine compared with subjects from group 2 (median: 12.7 versus 20.5 micromol; p < 0.05). In a logistic regression with hyperresponsiveness to methacholine at follow-up as dependent variable, the odds ratios (OR) for the groups were, with group 1 as reference: group 2: OR = 2.2 (0.8-6.6: p = 0.2), group 3: 5.9 (1. 6-21.7: p < 0.01). CONCLUSION: Subjects with sensitization and increased ECP levels are subsequently more airway-responsive to methacholine compared with sensitized subjects with normal ECP levels. This supports the hypothesis that sensitization is linked to increased bronchial reactivity through a process in which markers of inflammation are involved. PMID- 10705224 TI - Specific IgE levels to Cicer arietinum (Chick pea) in tolerant and nontolerant children: evaluation of boiled and raw extracts. AB - The chick pea, Cicer arietinum, is a legume commonly consumed in Spain and other Mediterranean countries. The sera of 29 children (mean age: 8.4 years) with a current or past history of allergic reactions after ingestion of chick pea, and positive skin tests to this legume, were used to study the allergenic composition of raw and boiled chick pea extracts. The patient population was divided into 2 groups: group 1 consisted of 19 patients with clinical sensitivity confirmed by either positive oral challenges or a convincing recent history of anaphylaxis after eating chick peas, and group 2 consisted of 10 patients with clinical sensitivity in the past, but tolerant at the time of blood extraction. Six atopic children, not allergic to legumes, were included as controls. Specific IgE to the raw and boiled extracts was measured by ELISA. The allergenic composition of both extracts was analyzed by SDS-PAGE and immunoblots. There were no significant differences between specific IgE levels to the raw and boiled extracts (p = 0.23). The mean levels in group 1 were significantly higher than in group 2 and controls (p = 0.0001). Multiple IgE binding proteins/peptides were detected in both extracts in the molecular weight range of 10-106 kD. Only nontolerant patients recognized a similar number of bands in both extracts. Chick pea extracts contain a majority of heat-stable allergens, which could be responsible for the clinical sensitivity to chick pea. Patients with a current clinical allergy to chick pea have statistically higher specific IgE levels than tolerant patients and controls. PMID- 10705225 TI - IL-4 induces eotaxin production in corneal keratocytes but not in epithelial cells. AB - BACKGROUND: In severe allergic eye diseases, the breakdown of epithelial barrier function can lead to severe corneal damage such as erosions or ulcers which often resist treatment. Although eosinophils are thought to play a crucial role in corneal tissue damage in severe ocular allergy, the mechanisms of eosinophil recruitment to the cornea has not been fully clarified. Eotaxin has been found in tears of severe allergic patients with corneal ulcer. In this study, we investigated whether the Th2 cytokine interleukin-4 (IL-4) induces eotaxin production in human corneal epithelial cells and keratocytes. METHODS: Primary cultures of human corneal epithelial cells and keratocytes were incubated with IL 4 and/or TNF-alpha for 48 h. Released eotaxin was measured by ELISA, and the eotaxin proteins were visualized by immunocytochemistry. Eotaxin mRNA expression in cultured cells was analyzed by RT-PCR. RESULTS: IL-4 induced eotaxin production in keratocytes in a dose- and time-dependent manner which was enhanced by TNF-alpha. There was no detectable eotaxin produced by corneal epithelial cells (<5 pg/ml). The cytoplasm of keratocytes incubated with IL-4 stained positively against anti-eotaxin antibodies, while eotaxin mRNA was detected in keratocytes incubated with IL-4. CONCLUSIONS: Human corneal keratocytes, but not epithelial cells, are capable of producing eotaxin by stimulation with IL-4. Our results suggest that eotaxin production in keratocytes induced by IL-4 may play an important role in eosinophil recruitment to corneal ulcers in allergic ocular disease. Eotaxin production by keratocytes may explain the severity of allergic disease involving the corneal stroma. PMID- 10705226 TI - Beclomethasone decreases elevations in phosphodiesterase activity in human T lymphocytes. AB - BACKGROUND: We recently reported that CD4+ T cells that have been activated in vivo or in vitro contain elevated cyclic adenosine monophosphate (cAMP) phosphodiesterase (PDE) activity. Since both phosphodiesterase inhibitors and glucocorticoids have anti-inflammatory activity, we sought to investigate the effect of beclomethasone on PDE activity. METHODS: PDE activity was measured in CD4+ T cells after 24 h of culture with beclomethasone. Cells were obtained from the peripheral blood of nonatopic persons (nCells), pre-seasonal (pCells), seasonal (within the first 2 weeks; sCells) and mid-seasonal (mCells) allergic rhinitics and asymptomatic allergic asthmatics (aCells). In addition, the effect of beclomethasone on Th2 cell lines and cells that had been activated in vitro with PHA or interleukin (IL)-2 was determined. RESULTS: PDE activity was decreased in a concentration-dependent manner by incubation of mCells, Th2 lines and PHA or IL-2-activated CD4+ T cells with beclomethasone (p < 0.05). However, beclomethasone did not modulate PDE activity in nCells, pCells, sCells, or aCells. CONCLUSIONS: Beclomethasone only decreases cAMP PDE activity in CD4+ T cells when it is increased by cell activation either in vitro or in vivo. PMID- 10705227 TI - beta-estradiol suppresses T cell-mediated delayed-type hypersensitivity through suppression of antigen-presenting cell function and Th1 induction. AB - BACKGROUND: Although an immunomodulatory role for estrogens has long been demonstrated by experimental and clinical observations, the mechanism by which estrogens exert their effect on T cells has not been clearly defined. METHODS: In this study we analyzed the effects of beta-estradiol (E2), at its contraceptive dose, on the delayed-type hypersensitivity (DTH) to purified protein derivatives (PPD) and associated immune response in female mice. RESULTS: E2 treatment decreased PPD-specific DTH response, which coincided with a decrease in the leukocytes numbers in the draining lymph nodes (DLN) and spleen compared with control mice. E2 treatment also suppressed the in vitro PPD-specific proliferative response of DLN and spleen cells from PPD-primed mice. The analysis of production and gene expression of cytokines by DLN cells demonstrated that E2 treatment suppressed IL-2 and IFN-gamma production in response to PPD in vitro. In contrast, IL-4 and IL-10 gene expression by DLN cells of E2-treated mice, taken 24 h after in vivo restimulation of mice with PPD, was enhanced. Furthermore, we found that spleen APC from E2-treated mice failed to induce optimum proliferation of the PPD-primed T cells in response to PPD in vitro. The impaired APC function by E2 was not due to induction of suppressor cell activity because addition of the normal spleen APC to APC from E2-treated mice restored the proliferative response of the PPD-primed T cells in response to PPD. CONCLUSION: Our results suggest that the E2-mediated inhibition of DTH reaction is due to a combination of the down regulation of APC function and deviation of the immune response from Th1-type to Th2-type. PMID- 10705228 TI - Antibodies to HIV-1 gp41 recognize synthetic peptides of human IFN-alpha and IFN beta. AB - Based on our finding that a common epitope exists between HIV-1 gp41 and human type I interferons (IFN-alpha and IFN-beta), and increased levels of antibodies against human IFN-alpha and IFN-beta were observed in HIV-1-infected individuals, we tried to explain the mechanism of increased levels of antibodies. Mouse antisera recognizing HIV-1 recombinant soluble (rs) gp41 (aa 539-684) interacted with two synthetic peptides sequence-corresponding to the IFN-alpha/beta receptor binding site on human IFN-alpha and IFN-beta, while normal mouse serum (pooled normal sera) did not. The anti-rspg41 antisera after adsorption by IFN-beta sepharose column lost the activity of interaction with both synthetic peptides. In another experiment, rsgp41 could bind to sepharose column conjugated with anti IFN-beta polyclonal antibodies (IgG). These results indicate that the common epitope on gp41 and type I interferons could induce antibodies recognizing the receptor binding site on IFN-alpha and IFN-beta, suggesting that increased levels of antibodies against IFN-alpha and IFN-beta in HIV-1-infected individuals could be induced by gp41. PMID- 10705229 TI - CASCADE: a European collaborative study on vascular determinants of brain lesions. Study design and objectives. AB - Dementia is a highly prevalent disease that may have a cardiovascular component. White matter lesions and brain atrophy (brain abnormalities) are prevalent in dementia cases and might form part of the anatomical basis for the disease. We designed a multi-centre study, CASCADE (Cardiovascular Determinants of Dementia), to examine long-term (10-20 years) and short-term (5 years) cardiovascular risk factors for, and the cognitive consequence of, brain abnormalities. White matter lesions and atrophy are measured with magnetic resonance imaging. Cognitive function is measured with nine tests of memory and executive function. The studies included in CASCADE were ongoing and geographically spread throughout Europe to capture the cardiovascular risk gradient. In each study, a random sample of at least 100 subjects aged 65-75 years was selected who participated in the previous research examinations conducted by the respective centres. The objectives and design of the CASCADE project are described. PMID- 10705230 TI - Prevalence of cognitive impairment and dementia as defined by neuropsychological test performance. AB - The Canadian Study of Health and Aging (CSHA) provided a population-based estimate of the prevalence of dementia of 8% for those aged 65 and older. Other studies have produced both higher and lower prevalence estimates. Factors that may contribute to these differences include: the use of or the reliance on neuropsychological testing, the consideration of functional impairment as a criterion for dementia and the inclusion of the category of cognitive impairment without dementia in the diagnostic classification. We examined the impact of these methodological factors by reanalyzing the CSHA database for those individuals who completed neuropsychological testing. If the diagnosis of dementia required only impaired neuropsychological test performance, there was an increased prevalence of dementia relative to the clinical consensus diagnosis, but including the requirement of functional impairment for dementia reduced this discrepancy. The findings illustrate the need for clear operationalization of diagnostic criteria for cognitive impairment and dementia in neuroepidemiological studies. PMID- 10705231 TI - Recruitment procedures and their impact on the prevalence of dementia. Results from the Leipzig Longitudinal Study of the Aged (LEILA75+). AB - Recruitment procedures may exert a considerable influence on the outcome of health surveys in the elderly. Their impact on the prevalence of dementia will be measured in an epidemiological field study in a sample of 1,692 randomly selected individuals (75+). Face-to-face interviews were conducted using SIDAM (structured interview for the diagnosis of dementia of Alzheimer type, multi-infarct dementia and dementias of other etiology according to ICD-10 and DSM-III-R). Furthermore, proxy interviews were performed with relatives of fragile and functionally dependent individuals. Considering face- to-face interviews of community-dwelling individuals, a prevalence of moderate and severe dementia of 5.3% was found. When including information on respondents by proxy and institutionalized individuals, the prevalence rate increased to 6.3 and 10.5%, respectively. It will be argued that covering the whole population in question and ensuring high response rates are central issues to minimize selection bias. PMID- 10705232 TI - Relationships between cholesterol, apolipoprotein E polymorphism and dementia: a cross-sectional analysis from the PAQUID study. AB - This study assesses the cross-sectional relationship between serum cholesterol level and dementia, controlling for apolipoprotein E (apoE) genotype, in a nested case-control study of 334 elderly French subjects aged 73 and over who participated in the PAQUID study (37 demented subjects and 297 nondemented controls). A diagnosis of dementia was established by two-step screening: (1) psychometric testing and DSM-III-R criteria and (2) neurologist's confirmation. Cholesterol, its fractions and apoE genotype were determined from a blood sample. Elevated high-density lipoprotein cholesterol was associated with a significantly decreased risk of dementia, independent of apoE status and other potential confounding variables, suggesting that cholesterol fractions could be involved in both Alzheimer's disease and vascular dementia. PMID- 10705233 TI - Neurologic recovery after penetrating craniocerebral war injury. AB - From June 1, 1991, until December 31, 1992, 116 patients with penetrating craniocerebral war injuries inflicted at the east Slavonian front were treated at the Osijek University Hospital. There were 26 (22.4%) gunshot wounds and 90 (77.6%) wounds inflicted by explosive devices and projectiles. Four years after the injury, a study of the survivors' condition was carried out. No difference was recorded in the survival rate between the patients with gunshot wounds compared with explosive wounds. Rehabilitation produced good results in the survivors, in the prevention of both acute and chronic complications and permanent damage. Older age, lower Glasgow Coma Scale, and level of consciousness were found to be prognostic indicators of outcome. PMID- 10705234 TI - 10th italian congress of neuroepidemiology. taormina, march 23-26, 2000 PMID- 10705235 TI - The emerging role of cytokines in the treatment of advanced melanoma. For the EORTC Melanoma Cooperative Group. AB - Cytokine-based treatment regimens have been evaluated for advanced melanoma in a number of phase I and phase II trials within the last decade. Treatment with interleukin-2 (IL-2) as a single agent has resulted in response rates of approximately 15%, if a high dose of IL-2 is administered. Combination of interferon-alpha (IFNalpha) and high dose IL-2 yields response rates ranging from 10 to 41%. Response rates exceeding 50% have been reported with chemoimmunotherapy, if the treatment regimens included at least the three agents IL-2, IFNalpha and cisplatin. Recent randomized trials have evaluated the impact of these drugs on the survival of patients with advanced melanoma. The current 'state of the art' is discussed in this review. PMID- 10705236 TI - Clinical pathway for the management of resectable gastric cancer with peritoneal seeding: best palliation with a ray of hope for cure. AB - Peritoneal seeding from primary gastric cancer occurs in 10-20% of patients. The diagnosis of this advanced disease is usually not provided by clinical studies prior to abdominal exploration. From the data available in the literature, the surgeon is forced to make an intraoperative judgement concerning the risks and benefits of an aggressive management plan versus supportive care. A strategy has evolved that utilizes peritonectomy and extended gastrectomy to maximally cytoreduce tumor combined with perioperative intraperitoneal chemotherapy. In the current state of the art, the perioperative intraperitoneal chemotherapy is heated and manually distributed to provide uniform treatment to all peritoneal surfaces and the resection site. The pharmacologic parameters have been established and the results of phase II studies are reported. Five-year survival of patients in whom a complete cytoreduction was possible is approximately 10% with a median survival of 12 months. Gastrectomy with peritonectomy to eliminate all visible implants combined with perioperative intraperitoneal chemotherapy should be used in selected patients with primary gastric cancer and carcinomatosis. PMID- 10705237 TI - Analysis of prognostic and survival factors related to treatment of low-grade astrocytomas in adults. AB - Prognostic factors for low-grade astrocytomas have been proposed, but optimal treatment remains controversial. Eighty-eight consecutive adult patients with supratentorial low-grade astrocytomas were retrospectively reviewed to determine specific factors influencing outcome. All underwent craniotomy (43 radical resections, 45 nonradical resections). Sex, age at diagnosis, preoperative Karnofsky performance status (KPS), tumor location, estimated extent of resection, radiation, chemotherapy, histological type, p53 status, MIB-1 staining and the apoptotic index were assessed as parameters for prognostic significance. KPS (p = 0.03), tumor location (p < 0.001), extent of surgical resection (p < 0.001) and radiotherapy (p = 0.01) were significantly associated with longer survival rates by univariate analysis. Multivariate analysis also showed a significant correlation between radiation therapy after surgical removal and survival time (p < 0.001). p53 status was not of importance in determining the necessity for radiotherapy. Radical surgical removal is the most important factor in the management of low-grade astrocytomas. Radiation therapy appears to be effective in improving the prognosis regardless of the extent of surgical resection or the p53 status. PMID- 10705238 TI - Chromosomes 7,17 polysomies and overexpression of epidermal growth factor receptor and p53 protein in epithelial hyperplastic laryngeal lesions. AB - PURPOSE: To visualize directly a sequence of genetic changes underlying the entire spectrum of epithelial hyperplastic laryngeal lesions (EHLL) and laryngeal cancer by the use of non-isotopic in situ hybridization (ISH) for chromosomes 7 and 17 in correlation with overexpression of p53 protein and epidermal growth factor receptor (EGFR). The specific aim was to compare the results and prognostic significance between the two types of EHLL: isolated, mainly atypical hyperplasia or risky epithelium, and EHLL associated with squamous cell carcinoma (SCC). PATIENTS AND METHODS: 59 tissue specimens of EHLL obtained from 34 patients, graded according to the Ljubljana classification into simple (SH), abnormal (AbH) and atypical hyperplasia (AtH), and carcinoma in situ (CIS) were included in the study. Non-fluorescent ISH for chromosomes 7 and 17 was performed by biotinylated alpha-satellite DNA probes. Immunohistochemical staining for EGFR and p53 protein was analyzed on the same tissue samples. RESULTS: Polysomy for both chromosomes increased in correlation with progressive grades of EHLL. The most important finding was the statistically significant difference in chromosome copy numbers between the isolated AtH and AtH associated with SCC. Overexpression of EGFR and p53 protein was found in 61 (36/59) and 52% (31/59) of cases, respectively. The immunoreactivity for both markers increased with the grade of lesions, but the staining pattern was not so uniform in isolated EHLL. On the other hand, the immunoreactivity was expressed more constantly in EHLL adjacent to SCC. CONCLUSIONS: Numerical changes in chromosomes 7 and 17 might be associated with an upregulation of EGFR and p53 genes, and could contribute to critical events in laryngeal carcinogenesis. For daily practice, the cytogenetic and immunohistochemical analyses could be of assistance in distinguishing between low- and high-risk groups of AtH. However, the isolated forms of atypical hyperplasia need considerable further study by evaluating genetic changes with the described methods regarding their ultimate transformation to carcinoma. PMID- 10705239 TI - Expression of tumor suppressor gene p16(INK4) products in primary gastric cancer. AB - Recent studies have shown that the cyclin-dependent kinase (CDK) inhibitor p27(Kip1) represents an indicator for patients' outcome in several human malignancies including gastric cancer. However, the clinicopathologic value of another class of CDK inhibitor, p16(INK4), has not been determined. In a retrospective study, we examined the expression of p16(INK4) by immunohistochemical assay of 80 samples of primary gastric cancers and their adjacent nonneoplastic mucosas. Less than 10% of non-tumor gastric mucosal cells were p16(INK4) positive, whereas the expression of p16(INK4) in gastric cancer cells varied widely from 0 to 100% (mean, 24.5%). The expression of p16(INK4) was not seen in 11.3% (9/80) of the cancer cases, but in 65% (52/80) this protein was even overexpressed when compared with the nonneoplastic mucosa. A clinicopathologic survey indicated that a low or no expression of p16(INK4) was associated with poorly differentiated carcinoma (p = 0.0133), but the level of expression did not correlate with other parameters including patients' prognosis or with the expression of the pRb protein. In an effort to explore the underlying mechanism for the p16(INK4)-negative cases, a prospective study was also performed on 20 cases of gastric cancer to compare the level of the p16(INK4) protein with the methylation status of the p16(INK4) promoter. Gastric cancer tissues with methylation expressed significantly lower levels of the p16(INK4) protein (p = 0.0013) and two of them lacked p16(INK4) expression altogether, whereas all the cancer tissues without methylation expressed it. These findings suggest that the p16(INK4) protein may be associated with differentiation of gastric cancer tissues and that methylation of the p16(INK4) promoter may, in part, account for the loss of p16(INK4) expression. PMID- 10705240 TI - Risk factors of metastasis in oral squamous cell carcinomas. AB - Lymph node and distant metastasis were comparatively studied in 225 oral carcinomas, and factors predisposing toward metastasis were investigated using clinical and immunohistopathological approaches. Neither the sites of tumors nor T-stage was correlated with either type of metastasis. Tumor cell differentiation was weakly correlated with lymph node metastasis, and stromal reaction (the degree of cell infiltration) did not differ greatly between metastasis-positive and negative tumors, although natural killer (NK) activities were correlated with lymph node metastasis. However, the mode of tumor cell invasion was closely associated with both lymph node and distant metastases. In grade 4C and 4D tumors, distant and lymph node metastases were observed in 8 (16%) and 31 (62%) cases, respectively, while of 68 grade 1 and 2 tumors, distant metastasis was not observed in any, and lymph node metastasis occurred in only 15 (22. 1%). In addition, the expression of p53 protein was correlated with lymph node metastasis; of 70 tumors without p53 protein expression, 23 (32.9%) revealed lymph node metastasis, while it occurred in 54 out of 96 tumors positive for p53 protein. However, p53 protein expression was not associated with distant metastasis, and p24 protein, a cyclin-dependent kinase inhibitor, did not show any relationship with either type of metastasis. These results indicate that lymph node metastasis is correlated with multiple factors in the host and tumor cells, but distant metastasis is only correlated with the mode of tumor cell invasion, suggesting that the former can be highly accurately predicted by invasion mode, p53 protein expression and NK activity. PMID- 10705241 TI - Overexpression of the heat shock protein HSP70 family and p53 protein and prognosis for patients with gastric cancer. AB - The cell synthesis of heat shock proteins is increased by a variety of environmental and pathophysiological stressful conditions. The 70-kD heat shock protein family (HSP70 family) which constitutively expresses hsc70 and heat inducible hsp70 is thought to be involved in protein-protein interactions, including oncogene products. We investigated the HSP70 family expression and biological behavior of gastric cancer, and its relation to p53 overexpression. Expressions of HSP70 and p53 in 164 primary gastric tumors were determined immunohistochemically. Exploratory data were analyzed on a set of 164 primary gastric cancers, and we constructed in prognostic significance of the HSP70 expression level and the relation to p53 overexpression. Expression of HSP70 (hsc70 and hsp70) were detected in nuclei and/or cytoplasm of cancer cells. Western blotting analysis showed that hsc70 and hsp70 were both expressed in five gastric cancer cell lines. Immunohistochemically stained positive cells of HSP70 varied from 0 (very weak) to 100%, in each case. The median level of positive cell rate was 19.0%. A HSP70 expression of over 19.0% was related to the differentiated tissue type of gastric cancer, but not to other clinicopathological factors. There was no difference in survival rates in subjects with higher and lower groups of HSP70 expression. HSP70 expression was also not related to p53 overexpression in the nuclei and p53 overexpression related poorer prognosis. Our findings show that the expression of HSP70 is not associated with tumor advance-related characteristics or with the prognosis of gastric cancer. Measurements of HSP70 expression do not appear to be a useful prognostic marker. PMID- 10705242 TI - Loss of p27(KIP1) expression predicts poor prognosis in patients with esophageal squamous cell carcinoma. AB - Immunohistochemical studies of cell cycle-regulating proteins were conducted on tissue samples from 106 patients with esophageal squamous cell carcinoma. Reduction of p27(KIP1) was observed in 41 (39%) cases and was significantly associated with a poor prognosis by univariate and multivariate analyses (p = 0.015). In contrast, p16(INK4) expression was reduced in 55 (52%) of the cases and was not correlated with prognosis. p27(KIP1) was not correlated with the PCNA index, while p16(INK4) expression was inversely correlated with the PCNA index (p = 0.01). The expression of these two proteins was not correlated with the clinicopathological parameters of the patients. Nodal status was shown by univariate analysis (p = 0.01) to be a prognostic factor, and poorly differentiated tumors were significantly associated with a poor prognosis by multivariate analysis (p = 0.027). Thus, reduction of p27(KIP1) plays an important role in the progression of esophageal squamous cell carcinomas and is considered to be an independent prognostic indicator of this disease. PMID- 10705243 TI - A novel case of Wilms' tumor followed by colon cancer, both showing microsatellite instability. PMID- 10705245 TI - Thrombocytes are the major source for soluble vascular endothelial growth factor in peripheral blood. AB - Serum levels of vascular endothelial growth factor (VEGF-S) have been reported to correlate with tumor stage and prognosis in various human malignancies. The source of soluble VEGF in peripheral blood remains obscure. We therefore measured the concentration of immunoreactive VEGF in 241 serum samples and 61 plasma samples (VEGF-P) from 20 subjects undergoing myeloablative chemotherapy and from 3 normal platelet donors. A significant correlation between the peripheral blood platelet count (PC) and VEGF-S (r = 0.86) but not VEGF-P was found. VEGF-S levels were 58.43 +/- 42.50 pg/ml (mean +/- SD) in patients with a PC < 50 x 10(9)/l, 203.29 +/- 176.56 pg/ml for a PC of 50-150 x 10(9)/l, and 457.42 +/- 475.41 pg/ml for a PC > 150 x 10(9)/l. Interestingly, VEGF-P levels were substantially lower than the corresponding VEGF-S values, namely below the detection limit in most cases. Supernatants from platelet-rich plasma contained no VEGF, but after in vitro lysis of the platelets very high VEGF levels were found. The VEGF content per 10(9) platelets was calculated at 2.51 +/- 2.39 pg and was dependent on the mean platelet volume. In summary, VEGF release from platelets during blood clotting was found to be the main source of VEGF in serum samples. Cancer patients in clinical remission have negligible amounts of soluble VEGF in peripheral blood, and myeloablative chemotherapy causes a significant drop in VEGF-S levels corresponding to the decrease in PC. Thus, studies addressing the diagnostic and prognostic value of VEGF-S in cancer patients must be interpreted with caution. Our data provide the basis for predicting VEGF-S in relation to PC in vivo, and for reevaluating former studies of VEGF-S in patients with malignant or nonmalignant disease. PMID- 10705244 TI - Bcl-2, Bax, Bcl-x(L) and Bcl-x(S) expression in neoplastic and normal endometrium. AB - The two anti-apoptotic proteins Bcl-2 and Bcl-x(L), and the two pro-apoptotic proteins Bax and Bcl-x(S) were measured by Western blotting in 51 neoplastic and 8 normal endometrial samples. The corresponding mRNA levels were analyzed by semiquantitative reverse transcriptase-polymerase chain reaction in a subgroup of 19 endometrial carcinomas. Neoplastic tissues had higher amounts of Bcl-2 protein than normal tissues (p < 0.051). Bcl-x(L) followed the same trend since its levels were higher in tumor than in normal samples (p < 0.048). Interestingly, Bcl-2 and Bcl-x(L) protein content showed a trend towards an inverse correlation (r = 0.27, p < 0.052). mRNA and protein levels directly correlated only with Bcl 2 (r = 0.63, p < 0.0032). Despite the fact that the amounts of Bcl-2, Bax and Bcl x(L) proteins in the neoplastic population were not significantly differently distributed according to the clinicopathological features of the patients, the differences observed between normal and neoplastic samples suggest that these proteins may play a role in endometrial carcinoma: long-term follow-up studies will be required to confirm this hypothesis. PMID- 10705246 TI - Erythropoietin in radiation oncology - A review. 1st international conference, Freiburg, June 11-12, 1999. AB - The therapeutic potential of erythropoietin gains increasing attention among radiation oncologists because the prognosis is better for patients with high blood hemoglobin levels following radiotherapy. However, there is still a debate on how hemoglobin affects radiotherapy. Further, the means to manipulate the hemoglobin level, their indication and administration need to be clarified. Available experimental and clinical data on hypoxia, anemia and on their treatment with erythropoietin have been extensively discussed at an international conference in Freiburg, Germany, in June 1999. This report gives a summary reviewing the topic. PMID- 10705247 TI - Obsessive-compulsive disorder with delusions. AB - Obsessive-compulsive (OCD) and delusional disorders (DD) have been recognised with increased frequency in recent years, and the propensity of some OCD subjects to become deluded has become a focus of interest. This study reports illness specific demography along with measures of symptom severity and tests to assess schizotypal ideation, dysfunctional attitudes, attributional and attention bias in 30 patients with OCD, 29 with DD, 16 with OCD with delusions (OC-DD) and a 30 subject control group (CG). Obsessional features appeared before delusions in the OC-DD group, suggesting that OCD was the primary pathology. Delusions were more likely in subjects obsessional about one rather than multiple themes. There was some support for proposals that depression and schizotypy may bring out delusions in OCD and some evidence for the utility of categorising OCD according to the number of obsessions a subject has. PMID- 10705248 TI - Magico-religious beliefs in schizophrenia: a study from north India. AB - Psychiatric disorders in India are often attributed to influence of supernatural phenomena, and many patients are subjected to various kinds of 'magico-religious' treatments. We studied 40 cases of schizophrenia and ascertained magico-religious beliefs held by their key relatives. The effects of such magico-religious beliefs on psychopathology and treatment-seeking behaviour were explored. The sample were schizophrenia patients diagnosed according to ICD-10 of the World Health Organisation. Psychopathology was assessed on the 9th version of the Present State Examination (PSE-9). Supernatural Attitude Questionnaire was administered to the key relatives of the patients to ascertain their beliefs about various supernatural phenomena and magico-religious treatments. It was observed that the majority of the patients had undergone magico-religious treatment (n = 23). Nearly 74% of the patients who had symptoms coloured by cultural influences such as delusional explanation in terms of paranormal phenomena had undergone magico religious treatment. It was also seen that though many relatives denied personal conviction in such magico-religious beliefs, yet they sought some kind of magico religious treatment for the patients. The prevalence of culturally influenced delusions as defined by the PSE-9 was very low. Belief in supernatural influences is common in patients' relatives from urban background and with adequate education, and treatment based upon such beliefs is sought to a considerable extent in such cases. Local and community belief in such phenomena appeared to be a factor in influencing the decision to seek magico-religious treatment. PMID- 10705249 TI - Patterns of comorbidity among DSM-III-R personality disorders. AB - The aim of this study was to examine patterns of comorbidity among personality disorders (PDs) in a sample of 156 psychiatric inpatients. PDs were assessed with Semistructured Clinical Interview for DSM-III-R Personality Disorders. To determine significant co-occurrence among axis II diagnoses, odds ratio and the percent of co-occurrence of pairs of disorders were calculated. Both statistical methods revealed high rates of comorbidity: significance association was found for 36 pairs of disorders using the percent of co-occurrence, and for 22 pairs of disorders using the odds ratio. These results support the concept of 'apparent comorbidity' for most PDs, deriving from conceptual and definitional artifacts or from a 'state-biasing effect'. In light of these observations, a categorical approach to PDs, resulting in a list of diagnoses, appears useless in psychiatric practice. A dimensional classification is probably better suited for PDs, improving the understanding of personality psychopathology and its clinical implications. PMID- 10705250 TI - Naturalistic course of obsessive compulsive disorder and comorbid depression. Longitudinal results of a prospective follow-up study of 74 actively treated patients. AB - Seventy-four patients who met DSM-III-R criteria for obsessive compulsive disorder (OCD) were studied in a prospective follow-up study in order to investigate course and prognosis of OCD with or without comorbid depressive symptomatology. Subjects were examined three times: at admission (baseline), 6 months later (follow-up 1) and 12 months after follow-up 1 (follow-up 2). At admission, 51 (72.9%) OCD patients were assessed as depressive by the Hamilton Depression Scale score. Between admission and follow-up 1, all patients received behavior therapy and a serotonin reuptake inhibitor, between follow-up 1 and follow-up 2 they received different kinds of treatment in order to maximize therapeutic effects. A 25% Yale-Brown Obsessive-Compulsive Scale (Y-BOCS) score reduction from admission to follow-up 2 and in addition, a total Y-BOCS score of below 16 at follow-up 2 was defined as 'good prognosis course'. The results obtained showed that OCD patients who followed a good prognosis course, showed no significant depressive symptomatology at follow-up 2 (p = 0.001). These results imply that patients with a diagnosis of OCD may present depression at admission and/or follow-up 1; however, if OC symptomatology decreases longitudinally, depressive symptoms disappear too. We may assume that OCD is dominant over depression, and it seems that a comorbid depression does not have any major influence on the prognosis of OCD. PMID- 10705251 TI - Clinical dimensions of delusional beliefs: a factor-analytic study. AB - We investigated the factorial composition as well as the demographic, anamnestic and clinical correlates of 13 features of delusional beliefs in a sample of 127 psychiatric inpatients with active delusions at the time of their assessment: Six factors were extracted, jointly accounting for 67.3% of the variance, which were interpreted as the dimensions of cognitive disintegration, volitional dyscontrol, doxastic strength, distress, incongruence and expansiveness, respectively. Moreover, most factors exhibited distinctive profiles of demographic, anamnestic or clinical correlates. Overall, our findings provide supportive evidence for both the clinical multidimensionality of delusional beliefs at the factor analytic level and the - at least partial - external validity of the factorial solution obtained. PMID- 10705252 TI - Bipolar II disorder and its premorbid personality. AB - This study psychopathologically analyzes and compares the premorbid personalities associated with bipolar II disorder and unipolar depression. Using Cloninger's tridimensional personality theory, we evaluated 14 inpatients with bipolar II disorder and 14 inpatients with unipolar depression. The results indicate that the premorbid personality associated with bipolar II disorder is characterized as 'the reward-dependent, passive-avoidant/dependent tendency of personality' or 'the dependent tendency of personality'. We also clarified that atypical symptoms of bipolar II disorder, such as the hypomanic state and the mixed state, were induced by the mingling of this tendency with the melancholic personality type (Tellenbach) or the cycloid personality type (Kretschmer), both of which are based on syntony (Minkowski), categorized by the use of an obsessional defense mechanism to maintain stable social relations. When an insufficiency in the obsessional defense weakens the syntony, the dependent tendency engenders the symptoms stated above. Although few serious problems associated with the dependent tendency may have occurred previously, it may have resulted in conflicts with others after onset of the bipolar II disorder. Therefore, psychotherapy for these conflicts is necessary along with the administration of mood stabilizers. PMID- 10705254 TI - Atypical bipolar II depression compared with atypical unipolar depression and nonatypical bipolar II depression. AB - Aim of the study was to find out whether atypical bipolar II depression was distinct from both atypical unipolar depression and nonatypical bipolar II depression. Seventy-nine consecutive atypical bipolar II depressed outpatients were compared with 42 consecutive atypical unipolar depressed outpatients and with 53 consecutive nonatypical bipolar II depressed outpatients. Among the variables studied (age at intake, age at onset, female gender, duration of illness, psychosis, comorbidity, chronicity, recurrences, severity), age at intake and onset were significantly lower in the atypical bipolar II group than in the other groups. The other variables, apart from psychosis, were not significantly different. Findings suggest that atypical bipolar II depression may have an age at onset different from that of atypical unipolar depression and nonatypical bipolar II depression. As different ages at onset may identify distinct subtypes of depression, this finding might suggest that atypical bipolar II depression may be distinct from both atypical unipolar depression and nonatypical bipolar II depression. PMID- 10705253 TI - Use of the ICD-10 classification in psychiatry: an international survey. AB - On the background of some years of experience with ICD-10 psychiatric diagnoses in many countries of the world, an international comparison was performed to evaluate the frequency and use of the ICD-10 psychiatric diagnoses. For future revision of the ICD-10, it is important to know which diagnostic categories are either not used or are used possibly in an unspecific manner. Nineteen departments of psychiatry in 10 different countries took part in the study, presenting data on 33,857 treatment cases leading to a total of 25,470 ICD-10 main diagnoses. The analysis of data reveals that on a four-character level (Fxx.x), the 10 most often used diagnostic categories represent 40% of all main diagnoses, and 70% on a three-character level (Fxx.-). There are 32 specific diagnostic categories on a four-character level which are not used at all and 121 which are used less frequently than 0.1% in inpatient and outpatient treatment. The study shows that the ICD-10 classification is in use in a variety of treatment settings worldwide. Further results and limitations of this study are discussed against the background of transcultural differences. PMID- 10705255 TI - Pathogenesis of lung disease in cystic fibrosis. AB - Lung disease in cystic fibrosis is primarily due to a defect in the cystic fibrosis transmembrane regulating protein (CFTR). This results in abnormal chloride transfer across epithelial membranes causing an excessively viscid mucus lining of the airways. Bacterial invasion particularly with Staphylococcus aureus, Haemophilus influenzae and Pseudomonas aeruginosa stimulates a vigorous and excessive primarily neutrophil-driven inflammatory response throughout the lungs. Products of this inflammation not only damage incoming bacteria but also the host tissue itself. Over a period of years this chronic suppurative process results in permanent ongoing lung destruction principally manifested as bilateral bronchiectasis. PMID- 10705256 TI - Serum concentrations of lignocaine before, during and after fiberoptic bronchoscopy. AB - BACKGROUND: Lignocaine is commonly used for local anesthesia during fiberoptic bronchoscopy (FOB). Several studies have reported the peak serum concentration of lignocaine in relation to time, but most of them did not specify the administered dose of lignocaine gel and its possible correlation with peak serum concentration. OBJECTIVE: The aim of our study was to record the plasma concentrations of lignocaine before, during and after FOB and to evaluate whether the doses for nasal and tracheobronchial anesthesia have any correlation with the peak serum concentrations of the drug. METHODS: Twelve patients with no comorbid conditions undergoing FOB were studied. Lignocaine was administered as a 2% solution using a larynx syringe, 2% gel (mean dose 182.5 +/- 15 mg) and finally 2% solution through the bronchoscope (mean dose 339 +/- 12 mg). Total dose was 622 +/- 20 mg. Venous blood samples were taken before the beginning of local anesthesia and then at 5, 10, 20, 60, 90 and 120 min thereafter. RESULTS: Our results showed that peak plasma concentrations of lignocaine were observed in 8 patients 20 min after the beginning of local anesthesia, in 3 patients 30 min afterwards and in 1 patient 60 min afterwards (2.15 +/- 0.4 microg/ml, 1.9 +/- 0.3 microg/ml, 1. 81 microg/ml, respectively). None of our patients exceeded the critical level of toxicity (5 microg/ml). Both the total and tracheobronchial doses of lignocaine were significantly correlated with peak serum concentration (r = 0.63, p = 0.05 and r = 0.64, p = 0.02, respectively). No correlation was found between the dose for nasal anesthesia and peak serum concentration. No adverse reactions were observed. CONCLUSIONS: In conclusion our data show that although the amount of lignocaine used in this study exceeded the recommended highest dose (400 mg) in all patients, no toxic levels were observed. Peak plasma concentrations were found within 20-30 min from the beginning of local anesthesia. The dose for the anesthesia of nasal mucosa represented a significant percentage of the total dose, but did not correlate with the peak serum concentration of the drug. PMID- 10705257 TI - Light's criteria revisited: consistency and comparison with new proposed alternative criteria for separating pleural transudates from exudates. AB - OBJECTIVES: The first objective was to assess the diagnostic value of new biochemical criteria proposed to discriminate pleural transudates from exudates and to compare their efficiency with those of Light's criteria. The second objective of the study was to assess the interstudy variability of the parameters repeatedly determinated in two different groups of patients with pleural effusion. PATIENTS AND METHODS: We recorded clinical characteristics and final diagnoses and measured pleural fluid (PF) and serum levels of protein, LDH, cholesterol and cholinesterase of 243 patients with pleural effusion. RESULTS: Sixty-one (25%) pleural effusions were transudates and 182 were exudates. The sensitivity (99%) and accuracy (96%) of Light's criteria were higher than those of the other criteria tested, although the differences with those of the PF LDH cholesterol combination (96 and 93%) did not show statistical significance. Pleural LDH concentration was the criterion with the highest specificity (95%), being significantly higher (p < 0.05) than that of Light's criteria. The sensitivity, specificity and accuracy of most criteria tested did not vary when compared with those obtained in a study performed 5 years previously. CONCLUSIONS: Light's criteria remain the criteria of choice for segregating exudates from transudates. Based on cost-efficiency reasons, the PF LDH cholesterol combination appears as an alternative. Because both sets of criteria misdiagnose a substantial percentage of transudates, exceptions based on good clinical judgment and the complementary use of a more specific criterion, as the PF concentration of LDH, must be considered. PMID- 10705258 TI - Limited additive value of pleural fluid carcinoembryonic antigen level in malignant pleural effusion. AB - OBJECTIVE: To assess the additive value of pleural fluid carcinoembryonic antigen (CEA.PF) level in the diagnosis of malignant pleural effusion. METHODS: Thoracentesis and closed pleural biopsy were performed in consecutive patients with pleural effusions. CEA.PF, cell analysis, and biochemical, cytopathologic and microbiologic studies were carried out. Further diagnostic interventions were undertaken if initial tests were inconclusive. RESULTS: A total of 176 patients were evaluated. The effusions proved malignant in 78 patients (44%). Benign etiologies were diagnosed in 89 cases, comprising 51 tuberculous pleurisies, 12 empyemas, 26 others. The cause was unknown in 9 patients. Median (range) in ng/ml of CEA.PF were 233 (1-12,500) in malignant vs. 2.5 (0.3-9) in tuberculosis, 1.4 (0.1-2) in transudates, 19.4 (0.6-312) in empyemas, p < 0.001. Receiver operating characteristic curve identified 10 ng/ml as the best cut-off for CEA.PF, yielding a sensitivity of 0.77, a specificity of 0.94, a positive and negative predictive value of 0.92 and 0.82, respectively. Among the 78 patients with malignant effusions, CEA.PF was elevated but initial cytopathologic study was nondiagnostic in 14 patients (18%). Prompted by the raised CEA.PF, further diagnostic interventions were undertaken and secured the diagnosis of malignancy in all of these 14 patients. CONCLUSIONS: CEA.PF level adds limited value on cytopathologic study in the diagnosis of malignant pleural effusions. It potentially identifies 18% of patients with malignant effusions who require further investigations despite negative initial cytopathologic study. PMID- 10705260 TI - Co-expression of vascular endothelial growth factor and its receptor flt-1 in malignant pleural mesothelioma. AB - BACKGROUND: Vascular endothelial growth factor (VEGF) is a potent angiogenic protein with a selective mitogenic effect on endothelial cells known to be involved in many normal and pathological processes. Coexpression of VEGF and its receptor flt-1 has been reported in different types of malignant tumors. OBJECTIVE: In the present study we investigated the expression of VEGF and flt-1 in 90 cases of diffuse malignant pleural mesotheliomas. METHODS: VEGF and flt-1 expression was analyzed by immunohistochemistry and non-radioactive in situ hybridization. RESULTS: VEGF expression was visualized immunohistochemically in tumor cells. flt-1 expression correlated with histological differentiation (p < 0.013). Furthermore, expression of flt-1 was detected in tumor cells, macrophages and microvessels adjacent to tumor cells. VEGF and flt-1 expression were confirmed by in situ hybridization. CONCLUSION: There was a statistically significant correlation between VEGF and flt-1 expression (p < 0.001). The observed coexpression of VEGF and flt-1 possibly suggests a potential autocrine loop for malignant pleural mesothelioma cells. PMID- 10705259 TI - Significance of alpha-2-macroglobulin, alpha-1-acid glycoprotein, and C-reactive protein in pleural effusion differentiation. AB - BACKGROUND: The differentiation between exudates and transudates is fundamental when investigating the cause of pleural effusions. Acute-phase proteins could be potentially useful markers in this discrimination. OBJECTIVE: The present study was designed to evaluate whether the acute-phase proteins: alpha(2)-macroglobulin (AMG), alpha(1)-acid glycoprotein (AAG) and C-reactive protein (CRP) are useful in investigating the pleural effusions. METHODS: We prospectively measured the concentrations of the above proteins in the serum and pleural fluid of 84 consecutive patients with various diseases using a nephelometric assay. RESULTS: Pleural effusion AMG, AAG and CRP were all significantly elevated in the group of patients with exudates compared to patients with transudates (p < 0.001, p < 0.001 and p < 0.01, respectively). An AAG value >63 mg/dl in a pleural effusion is predictive of an exudate with a sensitivity of 90% and a specificity 85%. Similarly, an AMG value >44 mg/dl in a pleural effusion is predictive of an exudate with a sensitivity and a specificity of 90% and 60%, respectively. Moreover, pleural AAG was significantly higher in cancerous exudates than in exudates and transudates of all other cause taken together (p < 0.001). Finally, to differentiate the same pleural effusion, the cut-off value of 1.0 mg/dl of pleural CRP has a sensitivity and a specificity of 74% and 74%, respectively. CONCLUSIONS: We conclude that both AAG and AMG concentrations in pleural effusions have a high sensitivity and are therefore useful parameters in distinguishing exudates from transudates, but the latter is inferior due to its unacceptably low specificity. PMID- 10705261 TI - Lung function decline in 4-monthly repeated spirometric measurements: due to silt aerosol exposure or decreasing effort? AB - BACKGROUND: Workers on dredgers and lighters on rivers are exposed to the inhalation of aerosols and dusts. OBJECTIVE: The aim of this study was to investigate effects of river silt aerosol and dust exposure on the respiratory health of dredging employees. METHODS: Six examinations were performed over a period of 2 years at 4-monthly intervals in 54 seamen with higher silt aerosol exposure and 36 controls of the same employer. RESULTS: No significant differences could be observed between the groups at any time of the study but there was an unexpected significant decrease in the age-corrected expiratory vital capacity (FVC), forced expiratory volume in 1 s (FEV(1)) and midexpiratory flow rate (MMEF(25/75)) over the six series in both groups. This may indicate a loss of effort of the participants in re-examinations since biological and technical influences were highly unlikely to be the cause of these findings. CONCLUSIONS: Ignoring this possible decline of effort in frequently repeated measurements may result in overestimating potential effects of occupational exposure. PMID- 10705262 TI - Pulmonary function tests of Mycobacterium avium-intracellulare infection: correlation with bronchoalveolar lavage fluid findings. AB - BACKGROUND: The pulmonary Mycobacterium avium-intracellulare (MAI) infection in patients with neither predisposing lung disease nor immunodeficiency is recognized to occur predominantly in older or middle-aged and thin females, and has characteristic chest computed tomography (CT) findings with multiple nodules and bronchiectasis. OBJECTIVE: To investigate the pathophysiology with and individual of MAI infection with neither predisposing lung disease nor immunodeficiency. The bronchoalveolar lavage (BAL) fluid (BALF) was recovered directly from the affected segments identified by the chest CT. METHODS: We performed a pulmonary function test (PFT) and BAL in 15 patients (all females) with MAI infection and 5 normal female volunteers. RESULTS: The residual volume and the slope of phase III (DeltaN(2)) were significantly increased and V(25) was significantly decreased in the 15 patients compared with the normal control subjects. The patients' BALF showed significant increases in neutrophils and activated CD4 lymphocytes, proinflammatory cytokines, and neutrophil elastase (NE) compared to those in the controls. We analyzed the PFT and BALF findings, and observed a significant correlation between the DeltaN(2) and interleukin (IL) 8 concentration (r = 0.65, p < 0.01), IL-8/albumin ratio (r = 0.51, p < 0.05) and NE/albumin ratio (r = 0.57, p < 0.05) in the BALF and DeltaN(2), respectively. CONCLUSIONS: These findings suggest that small airway dysfunction in MAI infection without predisposing lung disease is related to the inflammation probably related to the neutrophil. PMID- 10705263 TI - Neutrophil inflammation and activation in bronchiectasis: comparison with pneumonia and idiopathic pulmonary fibrosis. AB - BACKGROUND: Pulmonary inflammation in bronchiectasis, pneumonia and idiopathic pulmonary fibrosis (IPF) is dominated by neutrophils. Pathophysiologic differences are seen in the degree of airway and tissue destruction. Neutrophil activation and neutrophil proteolytic activity might differ between bronchiectasis, pneumonia and IPF. OBJECTIVE: The aim of this study was to determine whether levels of inflammatory and protective markers in bronchoalveolar lavage (BAL) differed among cases of bronchiectasis, pneumonia and IPF. METHODS: We studied 11 bronchiectasis patients (group 1), 30 pneumonia patients (group 2), 15 IPF patients (group 3) and 12 healthy volunteers (group 4). In the bronchoalveolar lavage fluid, concentrations of alpha(1)-proteinase inhibitor, myeloperoxidase (MPO) and elastase-alpha(1)PI complex were determined using immunoluminometric assays. Elastase inhibition capacity (EIC) and elastase activity were determined using a colorimetric assay. RESULTS: No EIC, but free elastase activity, was found in 82% of group 1, 20% of group 2, 20% of group 3 and 0% of group 4. Median MPO concentration was highest in group 1: 7,951 ng/ml (16th-84th percentile [16-84%]: 256-36,342) vs. 692 ng/ml (106-2,279; group 2), 332 ng/ml (98-1,657; group 3), and 0.12 ng/ml (0.08-0.26; group 4). Bronchiectasis patients with bronchial Pseudomonas infection showed higher amounts of neutrophils (p < 0.01) and higher elastase activity (p < 0.05) than patients with sterile lavage. CONCLUSION: Bronchiectasis patients show a severe imbalance between neutrophil activity and protective molecules leading to possible lung destruction. Chronic Pseudomonas infection might trigger neutrophil activation. Future research and treatment strategies should focus on increased bacterial clearance and inhibition of neutrophil toxicity. PMID- 10705264 TI - Non-pulmonary effects induced by the addition of formoterol to budesonide therapy in patients with mild or moderate persistent asthma. AB - OBJECTIVE: The present study was designed to assess the non-pulmonary effects of a 2-week treatment with the addition of formoterol to budesonide therapy in 10 patients with mild or moderate asthma. METHODS: Each patient was invited to perform a screening visit which included spirometry before and after inhalation of 12 microg formoterol with a metered dose inhaler (MDI), measurement of arterial blood pressure, baseline electrocardiography and 24-hour Holter monitoring, and a test for evaluating upper limb tremor. Patients then began bronchodilating therapy with 12 microg formoterol MDI and 400 microg budesonide Turbuhaler b.i.d. Each patient was also given a peak flowmeter and a diary in which he had to record the morning and evening values measured before taking inhaled drugs. Two weeks later, the patients repeated the same examinations; the diary card was returned 2 months after the beginning of the study. RESULTS: Adding formoterol to budesonide therapy caused a significant improvement in lung function, but neither induced any statistically significant effect on mean heart rate, nor altered the circadian rhythm of autonomic regulation nor elicited significant alterations in cardiac morphology. However, the evaluation of upper limb tremor revealed a statistically significant increase (p = 0.02). CONCLUSIONS: This study shows that adding the recommended dose of formoterol to an inhaled corticosteroid therapy does not induce significant cardiac undesirable effects, although tremor, surely due to stimulation of beta(2) receptors of the skeletal muscles, may sometimes be a limiting effect. PMID- 10705265 TI - Quality of life and economic features in elderly asthmatics. AB - BACKGROUND: In the geriatric population, asthma tends to be overlooked. Moreover, typical symptoms of asthma may mimic chronic bronchitis and emphysema. OBJECTIVE: To compare the characteristics of asthma between elderly (>/=65 years) and adult (<65 years) asthma patients with regard to asthma severity, health-related quality of life, and direct expenditures for medical care generated by the disease. METHODS: A cross-sectional study was made in the asthmatic population older than 14 years in the area of Barcelona, Spain. Asthma severity was determined according to the International Consensus criteria of 1992. St. George's Respiratory Questionnaire (SGRQ) was used to measure the quality of life. Direct costs were calculated registering all costs generated by each patient per year. RESULTS: The study population consisted of 282 adult asthmatics and 51 elderly asthmatics. Asthma was more severe in the elderly group (mild 10%, moderate 35%, severe 55%) than in the adult group (mild 47%, moderate 35%, severe 18%). Elderly asthmatics had significantly higher total SGRQ scores (48 vs. 35, p < 0.001) than adult asthmatics, as well as significantly higher scores for all subscales. Asthma-derived direct costs in elderly asthmatics (mean USD 1,490 vs. USD 773) were double those in adult asthmatics, mainly due to higher costs of hospitalization and medication in the elderly. CONCLUSIONS: Asthma in elderly people as compared with asthma in adulthood was more severe and was associated with a worse health-related quality of life, and significantly higher expenditures for medical care. PMID- 10705266 TI - Inhaled corticosteroids may reduce neutrophilic inflammation in patients with stable chronic obstructive pulmonary disease. AB - BACKGROUND: Although both inhaled and oral corticosteroids have anti-inflammatory effects causing improvement in clinical symptoms and spirometry in the treatment of asthma, the role of corticosteroids in the management of chronic obstructive pulmonary disease (COPD) is controversial. OBJECTIVE: To evaluate the effects of inhaled corticosteroids on sputum neutrophilia in clinically stable COPD patients. METHODS: In total, 18 patients were enrolled in the study. During 2 months, 9 patients in group A inhaled fluticasone propionate (FP) 500 microg 3 times daily. In group B 9 patients received placebo. All of the patients continued to inhale both salbutamol and ipratropium bromide. In 9 patients, sustained-released theophylline was also administered. Blood samples, spirometric tests, blood gas analyses, and either spontaneous or induced sputum cultures were evaluated on entry into the study, after a 2 months of treatment and following the 6-week washout period. RESULTS: After the 2-month FP treatment, no significant changes in the number of peripheral blood neutrophils, blood gas and spirometry data were observed in both groups. In group A, the total cell number and the number of neutrophils decreased from a mean of 3. 4 +/- 1.3 x 10(6) cells/g and 0.6 +/- 0.3 x 10(6) neutrophils/g on entry into study to 1.9 +/- 0.6 x 10(6) cells/g and 0.02 +/- 0.01 x 10(6) neutrophils/g after 8-week treatment with FP, returning to 3.3 +/- 1.1 x 10(6) cells/g and 0.5 +/- 0.3 x 10(6) neutrophils/g following the washout period. The percentages of neutrophils were 55. 6 and 77.9% in groups A and B after 2 months of FP treatment. There was no significant change in group B values during the study. CONCLUSION: These data suggest that neutrophilic inflammation in sputum may be decreased by inhaled corticosteroids in clinically stable COPD patients. PMID- 10705267 TI - Therapeutic equivalence and acceptability of two multidose powder inhalers in the treatment of asthma. AB - BACKGROUND: Problems in using conventional inhalation aerosols, in addition to environmental reasons, have driven development of the dry powder inhalers. OBJECTIVE: To compare therapeutic equivalence and acceptability of two dry powder inhalers, Easyhaler (EH) and Diskhaler (DH), in the delivery of beclomethasone dipropionate (BDP) in the treatment of asthma. METHODS: Adult asthmatics (n = 185), previously treated with inhaled steroids, were recruited in this open parallel-group study. After a run-in period of 2 weeks during which the patients inhaled 800 microg/day of BDP via DH, the patients were randomly allocated into three groups: EH 200 group (62 patients) using EH 200 microg/dose inhaler (1 inhalation q.i.d.), EH 400 group (62 patients) using EH 400 microg/dose inhaler (1 inhalation b.i.d.), and DH group (61 patients) using DH 200 microg/dose inhaler (1 inhalation q.i.d.) for 12 weeks. The primary outcome variables were PEF and FEV(1). RESULTS: The 95% CI for treatment difference in morning PEF between the EH 200 and DH groups was -16 to 23 litres/min and between the EH 400 and DH groups -18 to 21 litres/min. There was an increase in the morning PEF of 13 litres/min (p < 0.05) in the EH 200 group, and 9 and 11 litres/min in the DH and EH 400 groups. No differences between the groups were seen in the lung function parameters, in the symptom scores, in the use of rescue medication or in the incidence of adverse events. The treatments had no effects on morning cortisol levels. The patients in the EH groups compared the inhalers by using an 11-item questionnaire. In 8 questions the majority of the patients rated EH superior to DH. CONCLUSION: The tested inhalers were therapeutically equal. However, based on the acceptability data, the EH was better accepted by the patients than DH. PMID- 10705268 TI - Immediate intraoral adaptation of mandibular advancing appliances of thermoplastic material for the treatment of obstructive sleep apnea. AB - BACKGROUND: In the treatment of obstructive sleep apnea (OSA), mandibular advancing devices (MAD) are usually individually fabricated on plaster casts of both jaws from polymethyl-methacrylate. The potential disadvantages of these devices are (1) the costs and (2) the time required to construct the device. OBJECTIVE: In this study, the efficacy and feasibility of a cheap MAD consisting of thermoplastic material (SnorBan((R))), which can be directly moulded intraorally, were evaluated. METHODS: In a prospective study, the effect of an MAD consisting of thermoplastic material was investigated in 22 consecutive patients with OSA [respiratory disturbance index (RDI) 32.6 +/- 18.4/h]. Polysomnographic sleep was recorded prior to treatment and after 3 months of treatment with the MAD. RESULTS: Three of the 22 patients who did not tolerate the MAD were excluded from the analysis, whereas 11 patients were classified as responders. In the responder group, the mean RDI decreased from 27.6 +/-7.3 to 7.3 +/- 2.9 (p < 0. 01), correspondingly the sleep quality and the Epworth Sleepiness Scale improved (p < 0.05). Eight patients proved to be non-responders without relevant changes for the measured parameters. CONCLUSIONS: In 50% (11 of 22) of the patients, the MAD improved the OSA to a clinically relevant degree. In contrast to the majority of established MAD, the MAD investigated is cheap and immediately adaptable and thus a feasible strategy to 'screen' the efficacy of this therapeutic principle. Thus the construction of unnecessary MAD is avoided. PMID- 10705269 TI - The silhouette sign revisited. PMID- 10705270 TI - Concurrence of sarcoidosis and lung cancer. A report of four cases. AB - Although sarcoidosis and lung cancer are both frequently encountered conditions, their simultaneous occurrence in the same patient is unusual. In this report, we describe 4 cases of their concurrence and discuss the possible pathogenic mechanisms of their concurrent appearance. In particular, in 2 of the cases, both diseases had coexisted for a long period (more than 6 and 4 years, respectively), showing a surprisingly slow growth of cancers. Although the chest computed tomography showed hilar and mediastinal lymphadenopathy, the histopathological findings of the excised lymph nodes of both cases revealed no metastasis. The causal relationship between sarcoidosis and lung cancer remains uncertain, but cases such as these may be helpful in elucidating its precise nature. PMID- 10705271 TI - Myasthenic inspiratory vocal cord dysfunction: efficacy of nasal continuous positive airway pressure treatment. AB - Myasthenic vocal cord dysfunction (VCD), presenting with severe inspiratory stridor, was successfully treated with nasal continuous positive airway pressure (nCPAP), thus giving the medical staff time to make the diagnosis and avoiding intubation or tracheostomy. An important sign leading to diagnosis was the very high MEF(50)/MIF(50) ratio calculated from the flow-volume loop. nCPAP treatment induced prompt remission of stridor and a sharp reduction in the MEF(50)/MIF(50) ratio from 9.90 to 1.36. A review of the literature has shown that VCD with inspiratory stridor is an unusual onset symptom of myasthenia gravis and that nCPAP treatment may avoid emergency oral/tracheal intubation and tracheostomy. After diagnosis, the patient underwent thymectomy, and today, 3 years later, he is well without any further therapy. PMID- 10705272 TI - Trepopnea due to recurrent lung cancer. AB - Trepopnea is a condition whereby breathing may be comfortable in one position but difficult or labored in another. A unique case with trepopnea due to recurrent lung cancer with right main pulmonary artery stenosis and left main bronchus obstruction is presented. The patient had developed trepopnea 3 months earlier, but developed orthopnea shortly before he was admitted to our hospital. An emergent wall stent implantation was performed via the right femoral vein in the sitting position with the patient's leg stretched out. The symptoms and respiratory function improved after stent implantation. PMID- 10705273 TI - Spontaneous coughing up of a polyp. AB - We report the case of an 18-year old female with right lower lobe atelectasis, who was admitted to our hospital because of a nonproductive cough. She underwent fiberoptic bronchoscopy that revealed a peduncular polyp in the right truncus intermedius or middle bronchus. Before admission for laser polypectomy, she spontaneously coughed up the tissue mass, and the right lower lobe atelectasis disappeared. We report a rare case of 'autopolypectomy' of a bronchial adenoma. PMID- 10705274 TI - Pulmonary nodules following kidney transplantation. PMID- 10705275 TI - Treatment of spontaneous pneumothorax: why not simple talc pleurodesis by medical thoracoscopy? elisabeth.voland@vs.admin.ch. PMID- 10705276 TI - Bronchodilating effect of formoterol but not of salmeterol in two asthmatic patients. PMID- 10705277 TI - Bird Fancier's disease incidentally diagnosed by thallium scanning: a case report. PMID- 10705278 TI - [Card no. 32: ATM (Ataxia Telangiectasia Mutated)]. PMID- 10705279 TI - [Nutrition, genetic polymorphism, and cancer]. PMID- 10705280 TI - [Can they heal p53]. PMID- 10705281 TI - [The inactivating methylation of the p16(INK4a) promoter is an early event in oncogenesis]. PMID- 10705282 TI - [MIF and P53: 2 major partners at the inflammation and cancer crossroad]. PMID- 10705283 TI - [Ski and SnoN: antagonistic proteins of TGFbeta signaling]. AB - Ski and SnoN are two proto-oncogenes that, at high cellular concentrations, are associated with tumors. Up to now, apart the fact that SnoN and Ski were known to bind to DNA indirectly, very little was known about the mechanism which enables these factors to induce tumorigenesis. We know now that SnoN and Ski interact with the SMAD proteins which are mediators of TGFbeta signaling. These SMADs enable recruitment to target gene promoters of SnoN and Ski as well as the histone deacetylase activity which is associated with them. Whereas physiologic concentrations of SnoN and Ski allow a feedback regulation of TGFbeta signaling, deregulation of SnoN or Ski expression leads to total inhibition of TGFbeta signaling and of the tumor suppressors Smad2 and Smad4, which can explain the role of SnoN and Ski as oncogenes. PMID- 10705285 TI - [Role of diagnostic and therapeutic tools in the definition of differentiated thyroid carcinoma (1800-1950)]. AB - This paper is an attempt to evaluate the role of diagnostic and therapeutic tools to define differentiated thyroid carcinoma. At the beginning of the last century, physicians described its clinical feature: hard and invasive goiter arising after 25 and leading to death. In 1860, the surgical revolution encouraged the surgeons of goiter to treat thyroid cancer: simple goiter was viewed as precancer. From 1880, cell and tumor theories led pathologists to define microscopically thyroid cancer. In 1920, they demonstrated that the two most constant indications of thyroid epithelial malignancy were invasion of the blood vessels and distant metastasis. In 1930, radiotherapists introduced the concept of prognostic classification which combines histological criteria and patient survival for thyroid cancer. From 1940, the medical use of radioiodine led to distinguish two groups of thyroid tumors: those which are able to concentrate radioiodine and those which are not. Physicians, specialised in thyroid endocrinology, established the rules of thyroid cancer treatment. Our purpose is to analyse the epistemological and historical context of this pathology definition. PMID- 10705284 TI - [Review of adjuvant breast cancer therapy in non-menopausal women including early results of medical castration with LH-RH analogs]. AB - Three treatment modalities have successively dominated adjuvant therapy of breast cancer in non-menopausal women, namely, castration, chemotherapy and tamoxifen administration. The benefits afforded by each of these modalities seem similar when the treatments are compared indirectly by meta-analysis. Once the anti tumour action of LH-RH analogues and their reversible action on ovarian function had been established, these analogues were used instead of surgical castration in direct comparisons of the three treatment modalities. Most of the patients in these trials had estrogen and/or progesterone receptor positive tumours. According to the current state-of-the-art and whilst awaiting the final results of ongoing trials, we can conclude that: The survival of surgically castrated patients is the same as that of patients who have received CMF-type chemotherapy. The survival of patients on tamoxifen is the same as that of patients who have received CMF-type chemotherapy if tamoxifen is administered for 5 years. It is lower if tamoxifen is given for only 2 years. In 2 out of 3 trials, patients receiving the combined treatment castration plus tamoxifen had improved recurrence-free survival rates compared to patients on chemotherapy (regardless of whether an anthracyclin was included or not. It is too early to comment on overall survival. Combining castration and chemotherapy seems to be advantageous in patients less than forty and in those in whom chemotherapy has not induced amenorrhea. Combining tamoxifen and chemotherapy markedly decreases the risks of disease recurrence and of death but the high standard deviations recorded mean that this statement has to be tempered. Finally, an arrest of ovarian function by LH-RH analogues that is only temporary apparently does not adversely impinge upon results. This has, however, to be proved in an ad hoc trial and the optimum duration of analogue administration has to be established. PMID- 10705286 TI - [Standards, Options and Recommendations (SOR): pathology in oncology]. PMID- 10705287 TI - [Standards, Options and Recommendations (SOR) for drafting of anatomic and surgical pathology reports or cytopathology reports in oncology]. AB - CONTEXT: The "Standards, Options and Recommendations" (SOR) project, started in 1993, is a collaboration between the Federation of the French Cancer Centres (FNCLCC), the 20 French Cancer Centres and specialists from French Public Universities, General Hospitals and Private Clinics. The main objective is the development of clinical practice guidelines to improve the quality of health care and outcome for cancer patients. The methodology is based on literature review and critical appraisal by a multidisciplinary group of experts, with feedback from specialists in cancer care delivery. OBJECTIVES: To develop practice guidelines according to the definitions of the Standards, Options and Recommendations project for the content of the anatomic and surgical pathology or cytopathology reports in field of oncology. METHODS: Data were identified either by searching on Medline or via members of the expert groups personal references lists. When the guidelines were defined, the document was submitted to 49 independent reviewers, and to the medical committees of the 20 French Cancer Centres. RESULTS: The main recommendations for the drafting of the anatomic and surgical pathology or cytopathology reports in oncology are: 1) The reports must contain the identification of the pathologist, of the patient and of the specimen, a gross description for the surgical specimen, eventually a microscopic description, the diagnosis, all the elements essential for establishing the prognosis and for the clinical care, and a conclusion. 2) The reports could contain some comments. 3) The reports must be brief, precise, clear, homogeneous and ideally standardised, in order to be comprehensible for all the clinicians and the pathologists. PMID- 10705288 TI - [Standards, Options and Recommendations (SOR): clinical practice guidelines for diagnosis, treatment and follow-up of cutaneous melanoma. Federation Nationale des Centres de Lutte Contre le Cancer]. AB - CONTEXT: The "Standards, Options and Recommendations" (SOR) project, started in 1993, is a collaboration between the Federation of the French Cancer Centres (FNCLCC), the 20 French Cancer Centres and specialists from French Public Universities, General Hospitals and Private Clinics. The main objective is the development of clinical practice guidelines to improve the quality of health care and outcome for cancer patients. The methodology is based on literature systematic review and critical appraisal by a multidisciplinary group of experts, with feedback from specialists in cancer care delivery. OBJECTIVES: To develop clinical practice guidelines according to the definitions of Standards, Options and Recommendations for the management of patients with cutaneous melanoma. METHODS: Data have been identified by literature search using Medline - until December 1998 - and the personal reference lists of the expert group. Once the guidelines were defined, the document was submitted for review to national and international independent reviewers and to the medical committees of the 20 French Cancer Centres. RESULTS: The main recommendations for the management of cutaneous melanoma (CM) are: 1) The primary prevention of melanoma is based on a reduction in exposure to ultraviolet rays (solar or artificial). 2) The diagnosis of CM requires the surgical removal and histological examination of the lesion (standard). 3) The pathological report must include the diagnosis of primary malignant melanoma, the maximum thickness of the tumour in millimeters (Breslow), the clearance of surgical margins, the level of invasion (Clark), the presence and extension of regression and the presence of any ulceration (standard). 4) The standard treatment of a primary melanoma without lymph node involvement is based on surgery that must ensure adequate margins depending on the thickness of the tumour (standard, level of evidence B). 5) After surgery of a stage I melanoma, there is no indication for additional treatment outside a prospective therapeutic study (standard, level of evidence B, French Consensus Conference). 6) For a local recurrence without node involvement, in the absence of other metastases, surgical excision is the standard treatment. 7) In the case of metastatic regional lymph nodes, a complete regional lymphadenectomy is required. There is no indication for additional treatment outside a prospective therapeutic study (standard, level of evidence B). The inclusion of these patients in controlled studies of immunotherapy is recommended. 8) There is no standard therapeutic strategy for metastatic melanoma. Conventional palliative treatment is chemotherapy with dacarbazine (level of evidence B). 9) Follow-up is based on physical examination (standard). Patient information must encourage self surveillance. Clinical surveillance and self-detection are indicated in all cases throughout life (standard). PMID- 10705290 TI - [First line chemotherapy for advanced ovarian cancer in 2000: standards and questions]. AB - Ovarian cancer is wellknown to be chemosensitive since more than thirty years. However, long term results of this disease remain low. That's why standard first line chemotherapy is evolving to attempt to increase disease free survival and overall survival. Before cisplatin, standard chemotherapy was an alkylant used alone, mainly melphalan. With cisplatin disponibility, cisplatin based chemotherapy like cisplatin-cyclophosphamide with or without doxorubicin (CP or CAP) is used. Carboplatin can replace cisplatin because theses two platinum compounds have the same tumoral efficacy. Carboplatin is less toxic and its administration is more easy; so carboplatin with cyclophosphamide is actually the standard combination for elderly patients. Paclitaxel-cisplatin or carboplatin became the new actual standard combination. However, questions are asked concerning first-line chemotherapy for advanced ovarian cancer. Some of them are resolved like optimal number of cycles (6 in average), intensity-dose of cisplatin (25 mg/m2/week or 75 mg/m2 every 3 weeks) or for carboplatin (300 mg/m2 every 3 weeks or dose calculation according to AUC of 5 to 7.5 mg/ml x min). Another questions are ongoing like the place of anthracyclins or new drugs in front-line, the use of intra-peritoneal way for cisplatin and the role of intensive chemotherapy or immunotherapy as consolidation. PMID- 10705289 TI - [Primary chemotherapy with the Rosen T10 protocol before conservative surgery in limb primitive osteosarcomas: results about 56 cases]. AB - We report the results of a prospective Tunisian study using primary chemotherapy followed by conservative surgery in primitive limb osteosarcoma. From January 1988 to January 1998, 56 patients affected by limb osteosarcoma entered in a prospective study of neoadjuvant chemotherapy with the T10 protocol before surgery with a conservative intent. Initial work-up include: clinical exam with tumor measurements, chest and limb X-rays, limb CT-scan or MRI, chest CT-scan, bone scintigraphy and hematological and renal biological exams. Patients receive pre- and post-operative chemotherapy according to the T10 modified protocol. Fifty-six patients (33 M/23 F) with a mean age of 19 years (8 to 28) are included. Mean clinical and radiological tumor size is around 14 cm. Main histologic type is classic osteosarcoma (50% of cases) and 10 patients (9%) presented with initial metastasis; 42 patients on 56 receive the whole pre operative protocol. Treatment is well tolerated excluding 18 episodes of mucositis, 29 of leucopenia (< grade 3), 7 of thrombopenia (< grade 3), 4 of cutaneous toxicity, 2 of pulmonary toxicity and 3 of nausea-vomiting. We observe 36% of good histological responders and 64% of bad responders to primary chemotherapy, 27 patients on 49 operated (53%) have a conservative surgery and 18 (47%) a radical surgery. With a median follow-up of 51 months (8 to 128), 29 patients remain alive free of disease (15/17 GR and 14/30 BR), 2 are alive with disease, 2 died by toxicity, 14 died by progressive disease and 9 are lost to follow-up with evolutive disease. Five year disease-free survival is 55% for the 46 non metastatic patients. In univariate analysis, seric alkaline phosphatase level (p = 0.0014) and histological response to chemotherapy (p = 0.0218) are significant factors for prognosis. PMID- 10705291 TI - [Descriptive epidemiology of upper aerodigestive tract cancers in the department of Somme]. AB - The aim of this study is to show the epidemiologic features of upper aerodigestive tract cancers in the department of Somme. This study focused on the 1984-1995 period. For men, the upper aerodigestive tract cancers are the most frequent cancers in the Somme area. The incidence rate of pharynx cancers has shown slightly a decline over the past 10 years. The mortality rate of larynx cancers for males fell from 18.2 in 1984-1987 to 13.8 in 1992-1995 per 100,000. 5 year survival rates are between 20 and 39% for men, and from 23 to 42% for women. France has the highest incidence of these cancers in Europe. Within the country, the Somme area has a high risk for incidences and mortality as well. The results of this study indicate that upper aerodigestive tract cancers represent a serious problem for public health. Further areas need to be researched before such factors can be causally implicated in the aetiology of the disease in order to make up the prevention. PMID- 10705293 TI - [Congres, enseignements, sites internet en oncologie] PMID- 10705292 TI - [41th meeting of the American Society for Therapeutic Radiology and Oncology. San Antonio, Texas, October 31-November 4, 1999]. PMID- 10705294 TI - Leptin and the immune system: how nutritional status influences the immune response. AB - Several observations suggest the presence of an interaction between immune and the endocrine systems. Leptin is an adipocyte-derived hormone, that belongs structurally to the long-chain helical cytokine family such as interleukin-2 (IL 2), interleukin-12 (IL-12), growth hormone (GH), and signals by a class I cytokine receptor (Ob-R). This cytokine represents an important link between fat mass on the one side and the regulation of energy balance and reproductive function on the other. Indeed, obese leptin-deficient ob/ob mice display low body temperature, hyperphagia, infertility and evidence of immune defects with lymphoid organ atrophy, mainly affecting thymic size and cellularity. Acute starvation, associated with decreased leptin levels, causes thymic atrophy and reduces the delayed type hypersensitivity (DTH) reaction to antigens in normal mice, resembling that observed in ob/ob mice. Leptin replacement reverses the immunosuppressive effects of acute starvation in mice. Leptin differentially affects the in vitro proliferation and cytokine production by naive and memory T cells, increasing IL-2 secretion and proliferation of naive T cells, while inducing IFN-g production in memory T cells with little effect on their proliferation. Presence of leptin seems to be necessary for the induction and maintenance of the pro-inflammatory Th1 immune response. These findings support the hypothesis that leptin plays a key role in linking nutritional state to the T cell function. According to this view, leptin might represent an important target for immune intervention in a variety of pathophysiological conditions. PMID- 10705295 TI - Interleukin-18: biological properties and clinical implications. AB - IL-18, originally identified as interferon-gamma inducing factor (IGIF), is related to the IL-1 family in terms of its structure, processing, receptor, signal transduction pathway and pro-inflammatory properties. IL-18 is also functionally related to IL-12, as it induces the production of Th1 cytokines and participates in cell-mediated immune cytotoxicity. This review summarizes the recent advances in the understanding of IL-18 structure, processing, receptor expression and immunoregulatory functions, and focuses on the role of IL-18 modulation in tumours, infections, and autoimmune and inflammatory diseases. PMID- 10705296 TI - The importance of shedding of membrane proteins for cytokine biology. AB - Most transmembrane proteins are subjected to limited proteolysis by cellular proteases. The recent molecular cloning of the TNF-a converting enzyme (TACE) revealed that this shedding enzyme belongs to a family of metalloproteinases which contain a disintegrin domain (ADAM family). The activity of these proteases seems to be tightly regulated. Mice lacking functional TACE are not viable demonstrating the importance of this enzyme for body homeostasis. This review describes the current knowledge of shedding enzymes, the ADAM protein family, the mechanism of shedding as well as physiological consequences of shedding of cytokines and cytokine receptors for cytokine biology. PMID- 10705297 TI - Regulation of lipopolysaccharide-induced NO synthase expression in the major organs in a mouse model. The roles of endogenous interferon-gamma, tumor necrosis factor-alpha and interleukin-10. AB - Elevated NO production mediated by activation of the enzyme iNOS is thought to play a central role in the development of tissue damage observed during septic shock. IFN-gamma, TNF-alpha and IL-10 have been shown to be involved in the regulation of LPS-induced serum levels of the NO-oxidation products nitrate and nitrite. Therefore, in the present study, we investigated the role of endogenous IFN-gamma, TNF-alpha and IL-10 in the regulation of LPS-induced tissue iNOS expression in the major organs. To this end, mice were pre-treated with anti-IFN gamma, anti-TNF-alpha, anti-IL-10 monoclonal antibodies, or combinations of these, two hours before intraperitoneal LPS-challenge. Immunohistochemical staining for iNOS and determination of iNOS activity indicated that iNOS expression was mainly upregulated in the small intestine, lung and heart, and that IFN-gamma, TNF-alpha as well as IL-10 are involved in the regulation of iNOS expression and enzyme activity. Whereas blocking either IFN-gamma or TNF-alpha did not affect iNOS expression, iNOS enzymatic activity seems to be inhibited. In contrast, blocking both mediators nearly completely prevents iNOS expression after LPS challenge, suggesting that the presence of either IFN-gamma or TNF alpha is essential for LPS-induced iNOS expression in these organs. Combined treatment of these monoclonal antibodies revealed that whereas on the one hand IL 10 inhibits LPS-induced iNOS expression, on the other hand IL-10 or an IL-10 inducible factor is also involved in the upregulation of iNOS expression after LPS challenge. PMID- 10705298 TI - Interleukin-18 stimulates HIV-1 replication in a T-cell line. AB - Interleukin-18 (IL-18) is a recently identified proinflammatory cytokine. Its ability to induce interferon-g suggests a potential virustatic effect. On the other hand, it stimulates NFkB - an activator of HIV replication. Recently, stimulation of HIV-1 in monocytic cells has been demonstrated. In the present study, the influence of IL-18 on HIV-1 replication in lymphatic cells was investigated. Hut78 cells were infected with HIV-1 in the presence of recombinant human IL-18 expressed either in E. coli or eucaryotically by baculovirus in Sf9 cells. HIV-1 replication was monitored by p24 ELISA and endpoint titration of culture supernatants on C8166 cells. The addition of IL-18 led to a 3- to 15-fold enhancement of HIV replication in Hut78 cells. By addition of neutralising monoclonal anti-IL-18 antibodies, this effect of IL-18 was reduced by 75%. Exposure of Hut78 to IL-18 prior to HIV infection could exclude the possibility that IL-18 promotes infection of cells. Taken together, these data provide direct evidence for an IL-18-mediated enhancement of HIV-1 replication in lymphatic cells. PMID- 10705299 TI - Cell-cell contact of human T cells with fibroblasts changes lymphocytic mRNA expression: increased mRNA expression of interleukin-17 and interleukin-17 receptor. AB - Using random arbitrarily primed-reverse transcribed-PCR and sequence analysis, we investigated changes in lymphocytic molecules after cell-cell contact with fibroblasts. An mRNA species which was upregulated in Jurkat T cells by cell-cell contact with MRHF cells (a human foreskin fibroblast line) was identified as coding for the human interleukin-17 receptor. This finding was confirmed by quantitative RT-PCR for the HUT78 and Jurkat T cell lines, for peripheral blood lymphocytes, and for tonsillar T cells. Furthermore, the interleukin-17 mRNA, coding for a proinflammatory cytokine, was also upregulated in peripheral blood lymphocytes and tonsillar T cells after cell-cell contact with fibroblasts. Supernatants obtained from cell-cell contact-stimulated peripheral blood lymphocytes enhance the production of interleukin-6 and interleukin-8 by fibroblast-like synoviocytes and this effect could be blocked by interleukin-17 antibodies. Changes in the mRNA levels of Jurkat T cells induced by cell-cell contact with adherent cells were also found for M-type pyruvate kinase, for tropomyosin TM30 and for the p54nrb gene product. PMID- 10705300 TI - Induction of natural killer cell activity and perforin and granzyme B gene expression following continuous culture of short pulse with interleukin-12 in young and old mice. AB - Anticancer immunotherapy with cytokines is often limited by the occurrence of severe toxicity, particularly in older age groups, which are characterized by a reduced tolerance to antineoplastic therapies. We, and others, have recently demonstrated the efficacy of pulsing procedures with IL-2 as a new therapeutic strategy to induce antitumor cytotoxic cells. The aim of this paper was to evaluate the effect of IL-12 on NK cell activity in young and old mice and to investigate the possibility of inducing NK cytotoxicity and perforin and granzyme B gene expression through a brief exposure of spleen lymphocytes from young and old mice to IL-12. Pulsed lymphocytes were compared with non-pulsed cells cultured continuously in IL-12. IL-12 was able to boost both endogenous and IL-2 induced NK cell activity in young and old mice; the levels of cytotoxicity were lower in old than in young animals although the relative increase of IL-12 plus IL-2 versus IL-2 alone was greater for old mice. Comparable levels of NK cell activity were obtained in pulsed (5 min-1 hour) and non-pulsed lymphocytes from both young and old mice after one or three days of culture. The efficacy of the pulsing procedure was evident in both endogenous and IL-2-induced NK cytotoxicity. The mRNA encoding perforin and granzyme B were markedly and similarly enhanced in both IL-12-pulsed and non-pulsed lymphocytes in comparison with control cells. The results demonstrate the effectiveness of IL-12 pulsing in inducing antitumor cytotoxic cells, suggesting the possibility of using IL-12 pulsing, alone or in combination with IL-2, in the immunotherapy of both young and old subjects. PMID- 10705301 TI - Interleukin-12 induced interferon-gamma increases inflammation in acute dextran sulfate sodium induced colitis in mice. AB - There is increasing evidence that IL-12 and Th1-cytokines play an important role in intestinal inflammation. We therefore examined the role of IL-12 and interferon-gamma (IFN-gamma) in our model of dextran sulfate-induced acute colitis in mice. Treatment of mice with rmIL-12 during colitis induction resulted in severe aggravation as demonstrated by a greater loss of body weight, an increase of the histological parameters, and reduction of myeloperoxidase activity in colonic biopsies. Depletion of neutrophils in mice also led to aggravation of colitis. Neutralization of IFN-gamma in IL-12-treated mice with colitis inhibited these effects of IL-12. Neutralization of endogenous IFN-gamma or IL-12 with specific antibodies in DSS-treated mice, however, had only weak ameliorating effects. Since IL-12 and IFN-gamma have been shown to mediate experimental chronic colitis we conclude that the transition from a macrophage/neutrophil determined response to a Th-cell response promotes chronic intestinal inflammation. PMID- 10705302 TI - Serum concentrations of granulocyte-colony stimulating factor in complicated Plasmodium falciparum malaria. AB - Involvement of neutrophils in the control of blood parasites in malaria has been reported. Both, mononuclear phagocytes and neutrophils are known to be stimulated by cytokines such as TNF-alpha in order to augment the defence potency against the parasites. Previously, it has been shown that serum-G-CSF concentrations are increased in patients with bacterial sepsis. In vitro studies have shown that P. falciparum - infected erythrocytes induce the release of G-CSF by several cells such as endothelial cells and monocytes, however, nothing is known about G-CSF serum concentrations during the clinical course of severe P. falciparum malaria. Thus, it was the aim of the present study to investigate the time course for G CSF serum concentrations in patients with complicated P. falciparum malaria, and to correlate these values with other mediators of inflammation and hematopoesis. Twenty-six patients suffering from complicated P. falciparum malaria were included in the study, and 20, age and sex matched, healthy volunteers were used as the negative control group. Serum samples for determination of G-CSF were taken on day 0, 7 and 14, and measured by ELISA. We found significantly increased serum concentrations of G-CSF in patients with complicated P. falciparum malaria on day 0, values decreasing to within the normal range by day 7. A significant correlation was found between G-CSF (d0) and procalcitonin, the parasite count, erythropoietin and macrophage inflammatory protein, however no correlation could be shown for the neutrophil count. In conclusion, on the day of hospital admission, elevated serum concentrations of G-CSF were detected in patients with complicated P. falciparum malaria, which might indicate a role of G-CSF in the acute defence mechanism against the parasites. PMID- 10705303 TI - Interleukin-1 receptor antagonist, soluble tumor necrosis factor-alpha receptor type I and II, and soluble E-selectin serum levels in multiple sclerosis patients receiving weekly intramuscular injections of interferon-beta1a. AB - BACKGROUND: interferon beta (IFN-beta) reduces relapse rate and disease progression in patients with the relapsing-remitting form of multiple sclerosis (RRMS). IFN-beta may act by upregulating the expression of anti-inflammatory components of the immune system. OBJECTIVES: To determine whether weekly intramuscular (i.m.) injection of IFN-beta1a had a short- or long-term effect on the expression of naturally occurring soluble factors that play an immunosuppressive role within the cytokine network. MATERIALS AND METHODS: serum levels of interleukin-1 receptor antagonist (IL-1Ra), soluble tumor necrosis factor alpha receptor type I and type II (sTNF-alphaRI and sTNF-alphaRII), and soluble E-selectin (sE-Sel) were followed over time in ten patients with RRMS who were treated with weekly i.m. injections of 30 mg (= 6 MU) of IFN-beta1a. Patient sera were sampled before, and 24, 48, 72, 96, and 168 hours after the first IFN beta1a injection (short-term), and then at 1, 3, 6, 9 and 12 months after therapy initiation (long-term); highly sensitive, commercially available ELISA tests were used. RESULTS: serum levels of IL-1Ra, sTNF-alphaRI and sTNF-alphaRII, but not sE Sel were significantly increased in both short- and long-term follow-up. Interestingly, IL-1Ra, sTNF-alphaRI and sTNF-alphaRII behaviors were completely different, suggesting that these naturally occurring immunoregulatory factors were differentially affected by IFN-beta1a. CONCLUSION: our study demonstrates that weekly i.m. injection of 30 mg of IFN-beta1a induces the expression of soluble mediators that may suppress the activities of pro-inflammatory cytokines such as IL-1 and TNF-alpha. PMID- 10705304 TI - Hepatocyte growth factor plasma levels after myocardial infarction are not affected by recombinant tissue-type plasminogen-activator therapy. AB - Hepatocyte growth factor (HGF), a cytokine involved in tissue regeneration, angiogenesis and lateral vessel growth, is secreted as a biological-inactive, single-chain precursor named pro-HGF. In case, of tissue injury pro-HGF is proteolytically cleaved at the extracellular locus by serine proteases. Results obtained from in vitro experiments showed that urokinase-type plasminogen activator (uPA) and tissue-type plasminogen activator (tPA) can cleave single chain HGF. In this study we measured serum HGF levels in patients with acute myocardial infarction (MCI). Two groups of patients were compared. One group (n = 7) was treated with a conventional therapy and the other group (n = 7) was subjected to a thrombolytic therapy with recombinant tissue-type plasminogen activator (rtPA). Serum samples were collected at time of admission and subsequently 12-16 hours, 20-30 hours and 50-60 hours after onset of chest pain. At admission and before administration of rtPA, serum HGF levels peaked at 16.8 +/- 2.2 ng/ml in the lysed group and at 20.7 +/- 6.5 ng/ml in the non-lysed group. Levels then continuously declined, reaching lowest values 50-60 hours after onset of chest pain (3.2 +/- 1.3 ng/ml in the group treated with rtPA versus 4.4 +/- 0.9 ng/ml in the non-lysed group). No statistical significant difference could be detected between the two groups at any time. We suggest that serine proteases other than tPA are involved in HGF activation in vivo. PMID- 10705305 TI - Randomized study of recombinant interleukin-2 after autologous bone marrow transplantation for acute leukemia in first complete remission. AB - Immunological control of acute leukemia may be achieved after allogeneic transplant. Despite promising preliminary results, the impact of immunotherapy with interleukin-2 (r-IL-2) on patients with acute leukemia (AL), in first complete remission (CR1) remains unclear. We conducted a prospective multicenter randomized trial to compare outcome in patients with AL in CR1, treated with autologous bone marrow transplantation (BMT) with or without postgraft r-IL-2. One hundred and thirty patients with AL in CR1 (myeloblastic (AML): N = 78; lymphoblastic (ALL): N = 52) were randomized at time of BMT to receive (N = 65) or not (N = 65) r-IL-2. r-IL-2 (RU 49637 from Roussel Uclaf) was started after hematological recovery, as a five cycle regimen (12 M IU/m2/day continuous infusion on day 1-5, 15-17, 29-31,43-45 and 57-59). The two groups were balanced for patient and transplant characteristics. Analysis was based on an intent to treat. Thirty-eight (59%) of the 65 patients randomized into the study group started r-IL-2 at a median of sixty-eight days (23-140) after transplant and received 77% (16-100) of the scheduled dosage. They received a median of 120 x 10(6) IU/m2 (25-156) over 10 (3-13) days during a total median period of 56 (3 78) days. With a median follow-up of 7 years (5.4-8.1 years), 79 patients relapsed (study group: 43 (66%); control group: 36 (55%): p = NS). Survival and leukemia-free survival estimates were 33% (23-45) versus 43% (22-52) and 29% (19 41) versus 36% (24-51) respectively for study and control groups (all p = NS). These results show that leukemic control after autologous BMT is not increased by r-IL-2 therapy. Further studies should investigate more appropriate r-IL-2 schedules and the possibilities offered by better antigen recognition and activated effector cells. PMID- 10705306 TI - Aerosolized interferon-alpha treatment in patients with multi-drug-resistant pulmonary tuberculosis. AB - Multi-drug-resistant tuberculosis (MDR-TB) has emerged as an obstacle to the control of tuberculosis. Recent data however, suggest that interferon-(IFN)-gamma and IFN-alpha may improve disease evolution in subjects affected with pulmonary tuberculosis caused by multi-resistant (IFN-gamma) and sensitive (IFN-alpha) strains. The mechanisms involved are not known, even though it has been reported that IFN-gamma-secreting CD4+ Th cells may possess antitubercular effects. In addition, IFN-alpha can induce IFN-gamma secretion by CD4+ Th cells, and both types of IFN may stimulate macrophage activities. The aim of this study was to explore the possibility that aerosolized IFN-alpha, administered concomitantly with conventional antitubercular chemotherapy, may improve the course of pulmonary tuberculosis. After six months of directly observed therapy (DOT), seven patients who were non-responders to a second line antitubercular therapy were given an IFN-alpha aerosol (3 MU, three times a week) for two months as adjunctive therapy. All strains were resistant to at least two first-line drugs. After IFN-alpha administration, the patients were followed up for a further six months with the same DOT. Sputum samples were collected monthly during the study period, with the exception of the IFN-alpha administration period, when the observations were performed weekly. High resolution computed tomography (HRCT) chest scans were performed before and after IFN-alpha inhalations. The analysis of the results showed that the mean number of Mycobacterium tuberculosis (Mt) had remained statistically unchanged (p = 0.80) during the first 6 months of DOT. During the following 2 months of IFN-alpha administration, 5 patients became negative (p = 0.02). After the end of treatment a progressive increase in Mt number was observed (p = 0. 02). Sputum cultures remained positive for all patients throughout the study period, although a significant decrease (p = 0.02) in the colony number per culture was observed after adjunctive treatment with IFN alpha. After stopping administration of IFN-alpha, a significant increase (p = 0.03) in the colony number per culture was noted as well as in Mt numbers. HRCT scans were slightly improved in all patients. These preliminary data suggest that aerosolized IFN-alpha may be a promising adjunctive therapy for patients with MDR TB. Optimal doses and schedules however, require further studies. PMID- 10705307 TI - In vitro release of interleukin-15 by broncho-alveolar lavage cells and peripheral blood mononuclear cells from patients with different lung diseases. AB - IL-15 shares several biological activities with IL-2 and uses the b and g chain of the IL-2 receptor. In addition to its T-cell stimulating capacity, IL-15 exhibits regulatory properties on macrophage proinflammatory cytokine release. IL 15 is released by non-lymphoid cells, e.g. muscle cells, fibroblasts and monocytes/macrophages. In many lung diseases alveolar macrophages (AM) are activated and release pro- inflammatory cytokines. We asked whether IL-15 is released ex vivo by AM and peripheral blood mononuclear cells (PBMC) from patients with inactive sarcoidosis (PSi), active sarcoidosis (PSa), tuberculosis (TB), hypersensitivity pneumonitis (HSP), cryptogenic fibrosing alveolitis (CFA) and pneumonia (PN). Additionally, we examined the kinetics of the IL-15 release of these cells. During 24 hours of culture, AM from controls (CO) released 3.8 +/ 1.9 pg/ml (mean +/- SD) of IL-15, which was significantly lower than in most of the patient groups (PSa: 8.7 +/- 3.9 pg/ml, TB: 8.4 +/- 1.9 pg/ml, CFA: 5.7 +/- 1.5 pg/ml, and PN: 7. 8 +/- 2.6 pg/ml) except PSi (4.0 +/- 2.6 pg/ml) and HSP (9.3 +/- 9.5 pg/ml). PBMC from patients with PSa released significantly more IL 15 than PBMC from CO (10.8 +/- 8.9 pg/ml versus 6.9 +/- 2.2 pg/ml) whereas PBMC IL-15 release of the other groups did not differ from CO (TB: 5.7 +/- 1.4 pg/ml; CFA: 4.6 +/- 1.6 pg/ml; HSP: 4.9 +/- 3.8 pg/ml). Kinetic studies revealed a minor peak after 5 hours and a major peak from 12 hours to 35 hours for AM and PBMC. In summary, AM from all patient groups but the PSi and the HSP group released increased levels of IL-15, although the total amount of this cytokine is very low. PMID- 10705308 TI - Distinct patterns of cytokine regulation in discrete clinical forms of Plasmodium falciparum malaria. AB - The pathogenesis of two of the most severe complications of Plasmodium falciparum malaria, cerebral malaria (CM) and severe malarial anaemia (SA) both appear to involve dysregulation of the immune system. We have measured plasma levels of TNF and its two receptors in Ghanaian children with strictly defined cerebral malaria (CM), severe malarial anaemia (SA), or uncomplicated malaria (UM) in two independent studies in an area of seasonal, hyperendemic transmission of P. falciparum. Levels of TNF, soluble TNF receptor 1 (sTNF-R1) and 2 (sTNF-R2) were found to be significantly higher in CM than in the other clinical categories of P. falciparum malaria patients. Levels of both receptors depended on clinical category, whereas only sTNF-R1 levels were significantly dependent on parasitemia. Detailed analysis of the interrelationship between these variables resolved this pattern further, and identified marked differences between the patient categories. While levels of TNF, sTNF-R1 and sTNF-R2 correlated with parasitemia in UM, this was not the case in CM and SA. Rather, there was a tendency towards high levels of TNF and its receptors in CM and low levels in SA without significant correlation to parasitemia in either category. This, and the fact that malaria-induced increases in plasma levels of IL-10 are much lower in SA compared to CM, suggest that distinct forms of dysregulation of the immune response to infection contribute to the pathogenesis of CM and SA. PMID- 10705310 TI - The cytokine network and immune functions, by jacques Theze, ed PMID- 10705309 TI - Interferon-gamma and its functional receptors overexpression in benign prostatic hyperplasia and prostatic carcinoma: parallelism with c-myc and p53 expression. AB - The therapeutic potential of IFN-gamma in prostatic cancer has been documented in several reports, although no immunohistochemical studies of this factor and its receptors in the prostate have been reported. The aim of the present study was to investigate the expression of IFN-gamma and its receptor components (IFN-gamma Ralpha and IFN-gamma-Rbeta) in normal prostate, benign prostatic hyperplasia (BPH) and prostatic cancer (PC), as well as the possible relationship between this factor and the products of the p53 gene (the wild and mutant forms) and the oncogene c-myc, by means of immunochemical techniques (Western blot, ELISA, and quantification of immunostaining in histological sections). In normal prostate, IFN-gamma and its two receptors were expressed in the basal cells of the epithelium and some stromal cells. In BPH specimens, immunostaining of basal epithelial cells was significantly increased for IFN-gamma and its a receptor, whereas stromal cell immunostaining was significantly increased for IFN-gamma and its b receptor. In addition, columnar epithelial cells immunostained for IFNbeta Rbeta. PC specimens differed from BPH specimens in the significantly increased immunostaining of epithelial cells for IFN-gamma and its two receptors, and the immunostaining of columnar epithelial cells for IFN-gamma-Ralpha. Immunodetection of wild-p53 was weak and limited to some stromal cells in the three types of specimens. Immunostainings for both mutant-p53 and c-myc were negative in normal prostate, and positive in the epithelium and stromal cells of both BPH and PC specimens. Immunostaining intensity in PC was significantly higher than in BPH. These observations suggest that the expression of both mutant-p53 and c-myc, together with other factors, might be involved in the development of prostatic hyperplasia and neoplasia, while the increased expression of IFN-gamma and its receptors could be regarded as an attempt, although insufficient, to inhibit the uncontrolled cell proliferation. PMID- 10705311 TI - Bacteria-cytokine interaction in health and disease PMID- 10705312 TI - [Typing of extended-spectrum beta-lactamase-producing Salmonella typhimurium strains isolated in a pediatric unit]. AB - Extended-spectrum b-lactamases (ESBLs) derive mainly from TEM and SHV b lactamases. These enzymes confer resistance to all oxyimino cephalosporins and monobactams except cephamycins and carbapems. ESBLs are often encoded by large plasmids that carry resistance determinants to multiple antibiotics and spread among the members of the Enterobacteriaceae. Since the first outbreak of Klebsiella pneumoniae expressing an extended-spectrum beta-lactamase reported in 1984, nosocomial infections due to Enterobacteriaceae species which produce ESBLs have been generally recovered from patients hospitalized in intensive care units. The most frequently isolated ESBL-producing strains belong to the genus Klebsiella, Escherichia, Enterobacter and Proteus; ESBLs are rarely associated with the genus Salmonella. The first Salmonella were detected in France in 1984 (Salmonella typhimurium), in Tunisia in 1988 (Salmonella wien) and in Argentina in 1991 (Salmonella typhimurium). In 1994, 10 isolates of Salmonella typhimurium expressing an extended-spectrum beta-lactamase were isolated for the first time from 10 children hospitalized in a pediatric unit of the hospital Ibn-Rochd, Casablanca. Previous study showed that all isolates belonged the same serotype, and biotype, and showed a resistance to oxyimino beta-lactams, gentamycin, tobramycin and trimethoprim-sulfamethoxazole but remained susceptible to tetracycline, chloramphenicol and quinolones. Oxyimino beta-lactams resistance determinant of all strains of Salmonella typhimurium was transferred by conjugation to Escherichia coli; Resistance to gentamycin and trimethoprim sulfamethoxazole was also cotransferred. In this study, we characterized the relationship between all isolates by comparing plasmid profiles and patterns of proteins because there appear to be the more effective method for evaluating epidemiologic relationship between Salmonella species, and the protein profiles method has been used for many bacterial species. These two methods have the advantages of speed and simplicity. All isolates presented the same plasmid pattern characterised by three plasmids and the same pattern of proteins composed of 36 bands. We concluded by combining results that this outbreak involved the spread of the same strain of Salmonella typhimurium between the ten children. As this type of resistance is easily transferred by these isolates to other bacterial species, the major risk would be its transfer to Salmonella typhi. PMID- 10705313 TI - [Induction of labor by misoprostol, an analog of PGE1. A prospective study of 200 cases]. AB - We carried out a prospective study of 200 pregnant women who required induction of labor at full term, at the Lalla Meryem maternity unit of the Ibn Rochd University Hospital, Casablanca, between January 1st 1996 and June 30th 1997. The aim of this study was to evaluate the efficacy, tolerance and acceptance of misoprostol (Cytotec) as a drug for inducing labor in unfavorable conditions (Bishop < 5). Misoprostol (a PGE1 analog) was administered to the women via the vagina, with a dose of 1/4 tb (50 mg) given every 6 hours, and a maximum of 3 doses (150 mg). If labor had not begun 18 h after the start of the protocol, misoprostol induction was considered to have failed. We found that misoprostol failed to induce labor in 5% of cases, Syntocinon was required in 40% of cases and the interval between misoprostol insertion and vaginal delivery was 13.3 + 11.1 h. The rate of delivery by cesarean section was 22% and the mean amount of misoprostol required was 1.3 doses (66 mg). The mean cost of labor induction was 0.6 FF, the frequency of uterine hyperstimulation was 3.5% and maternal, fetal and neonatal tolerance was good. Our results confirm that intravaginal misoprostol is very effective and well tolerated for the induction of labor in pregnant women at full term, in unfavorable obstetric conditions. PMID- 10705314 TI - [Intestinal parasitosis in the inhabitants of a suburban zone in which the groundwater is polluted by nitrates of fecal origin (Yeumbeul, Senegal)]. AB - This study was carried out in 1997 to 1998, to determine the prevalence of intestinal parasite infestations due to groundwater pollution at Yeumbeul, Senegal, and to follow the progression of parasite infestations following anti parasite treatment. The study included 705 people living in a suburban zone in which the water table was polluted with nitrates of fecal origin. These individuals consumed either well water or water from springs. The overall prevalence of parasite infestation was 42.26%, but varied significantly with age (p < 0.001). Individuals who consumed well water were more frequently infested than those who consumed spring water, but the difference was not significant (p > 0.3). Giardia and Entamoeba coli were the most frequently isolated parasites, with Giardia predominating in the 0 to 9 year age-group. There was no significant correlation between the prevalence of the various parasites and indicators of water pollution (R2 = 0.0566 for nitrates and 0.1086 for fecal coliform bacteria). Similarly, no correlation was found with water pollution factors such as the depth of the water table (R2 = 0.027) and the distance between the wells and the latrines (R2 = 0.00007). Following specific treatment, the prevalence of parasite infestation fell to 30.81%. This indicates the limitations of drug treatment, which is always used alone to combat intestinal parasites, in the face of possible reinfestation. PMID- 10705316 TI - ["Project Calmette": French medical cooperation in Cambodia]. PMID- 10705315 TI - [Porocephalosis: still a rare diagnosis]. AB - We report a case of porocephalosis in a 35-year-old Congolese adult who habitually ate large amounts of snake. The principal nonspecific clinical sign was abdominal pain. Diagnosis was based on X-ray examination of the abdomen without preparation and abdominal ultrasound scan. PMID- 10705317 TI - [Calmette Hospital, Phnom Penh, Cambodia. Assessment of the implementation of the Medical Information System (SIM). Global analysis of the 1998 results]. AB - Calmette is a national university hospital with 220 adult beds. It has emergency, surgical, medical and gynecology and obstetrics departments, along with a radiology unit, a laboratory for medical analyses, a central pharmacy and an outpatient clinic. This hospital has an unusual statute, with managerial autonomy and a system of cost recovery that currently provides 64% of the hospital's income. Since 1994, it has benefited from a French cooperation program. The French NGO, Medecins du Monde, has been present at Calmette since 1990, providing support for <>, the indigent sector of the medical department. The aim of the Medical Information System (SIM) is to develop a simple, reliable and reproducible system so that, for every action undertaken at the hospital (hospitalization, day hospital and outpatient clinic) the following pieces of information are recorded: 1) the disease; 2) the type of patient; 3) the type of management; 4) the means used to treat the patient; 5) the cost. Data are collected and analyzed using programs created with EPIINFO software (CDC, WHO), using the EPIGLUE module. In 1998, 10,814 admissions were recorded at Calmette Hospital, 7,811 (72.2%) of which were to the Emergency Department and 3,003 (27.2%) of which were direct admissions to other wards. We analyzed 10,603 (95%) computerized medical summaries (RMI). About 50% of beds were occupied in the maternity and gynecology ward whereas almost 90% of beds were occupied in the surgical and emergency wards. AIDS and tuberculosis were the conditions most frequently treated by the medical department, despite a marked increase in more specialized areas of medicine such as cardiology and diabetology. The surgical department reflected the concentration on emergency services of the hospital, with cranial traumatism the primary reason for admission for the hospital as a whole. The mean age of patients was 27 years for the maternity ward and 49 years for the medicine A ward. The mortality rate was about 5% for the medical wards (mainly due to AIDS) and almost 50% in the emergency department (cerebrovascular neurologic disease, cranial traumatism). The proportion of nonpaying patients was high (about 40% in terms of stays in hospital and about 50% of all days spent in hospital). The training of a Cambodian manager for the SIM is a key priority. The point of the SIM is to use the treated data it produces to improve management and decision-making. The data it produces should be used to define the profile of the patients treated, both from a medical point of view and in terms of their ability to pay. This is a fundamental step towards identifying activities that should receive priority as part of a development strategy for a structure evolving in a highly competitive environment. The SIM data are also invaluable for the short term management of the hospital through the contribution they make to the development of effective analytical accounting, making it possible to evaluate costs and to adjust charges appropriately. Finally, the involvement of the SIM in the setting up and functioning of the Comite de Lutte Contre les Infections Nosocomiales (CLIN; the Hospital-Acquired Infections Committee) in 1999 to 2000 is not utopia, it is the logical continuation of improvements in the overall quality of care It involves, in particular, the training of nurses and head nurses, initiated by nurses acting as technical assistants in the French cooperation program. The definition of the role of the hygiene nurse and the selection of such a nurse from the trained head nurses are also part of this process. PMID- 10705318 TI - Toxoplasmosis in cambodia : initial serological evaluation at phnom penh changes in the resistance of plasmodium falciparum to chemical malaria treatments in cambodia from 1991 to 1997 clinical aspects of AIDS at calmette hospital, phnom penh knowledges, attitudes and practices of university students regarding HIV infection in phnom-penh, cambodia, 1999 cambodia and AIDS : french support tropical lung infections PMID- 10705319 TI - In Memoriam Richard B. Caspari, M.D., 1942-2000. PMID- 10705320 TI - The future of orthopaedics in the United States: an analysis of the effects of managed care in the face of an excess supply of orthopaedic surgeons. AB - Recent technological advances in orthopaedic surgery have propelled both the volume of surgical cases and their complexity, resulting in increased costs, which should naturally result in higher incomes for surgeons. However, the transition from a fee-for-service model of physician compensation to a managed care model has resulted in major shifts in economic resource allocation. An economic model of this market based on imperfect competition shows that these changes have shifted market power from surgeons to the managed care organizations. Our model predicts that practicing surgeons will retire earlier, medical students will begin to select other specialties, and innovation will be slowed. Antitrust laws limit surgeons' ability to combat this trend through meaningful collective bargaining, creating the potential for future shortages as the baby boom generation reaches retirement age and the demand for orthopaedic services increases dramatically. PMID- 10705321 TI - Tourniquet versus no tourniquet use in routine knee arthroscopy: a prospective, double-blind, randomized clinical trial. AB - PURPOSE: The purpose of this study was to determine the effects of tourniquet use for routine knee arthroscopy based on both subjective and objective functional outcome measures. TYPE OF STUDY: The study was a prospective, double-blind, randomized clinical trial. MATERIALS AND METHODS: There were 120 patients randomized to tourniquet inflation (300 mm Hg) or no tourniquet inflation during routine knee arthroscopy. Patients recorded their average pain on a visual analog scale and their narcotic use for the previous 24 hours, for the first 5 postoperative days. Patients also completed a preoperative and postoperative (2 week, 6 week, 3 month) Western Ontario and McMaster University Osteoarthritis Index (WOMAC), 6-minute walk, 30-second stair climb, 1-leg standing vertical leap, range of motion, and isokinetic strength testing. Time to return to work and sport was documented. RESULTS: No statistically significant difference was found between tourniquet-up and tourniquet-down groups for the WOMAC quality of life measure, functional tests, isokinetic muscle strengthening, or time to return to work or sport (t test/repeated measures analysis of variance). However, there was a trend for less early postoperative pain and slightly better isokinetic strength testing at 2 weeks in the tourniquet-down group. Visualization was rated by surgeons to be 3 times better in the tourniquet-up group, although mean operative time did not differ between the groups. CONCLUSION: The use of a pneumatic tourniquet at 300 mm Hg does not significantly effect overall patient quality of life or functional outcome following routine knee arthroscopy. PMID- 10705322 TI - The effect of an exogenous fibrin clot on the regeneration of the triangular fibrocartilage complex: an in vivo experimental study in dogs. AB - The effect of an exogenous fibrin clot on the regeneration of the triangular fibrocartilage complex (TFCC) was examined in a dog model. In 12 mature dogs, bilateral TFCC resection was performed. The resulting defect was packed with an exogenous fibrin clot (experimental) while the contralateral side was left empty (control). Tissue regeneration was evaluated grossly and histologically at 6, 12, and 26 weeks. At each time period, the regenerated tissue in the fibrin clot filled defect appeared more mature and more congruent with the adjacent cartilaginous surfaces than did the control (empty) defect. At 26 weeks, the clot regenerated tissue had the histological appearance of a normal TFCC with a homogeneous fibrocartilaginous matrix, regularly oriented collagen fibers, and normal integration with the adjacent support structures of the joint. The results of this study indicate that an exogenous fibrin clot could be used to promote a fibrocartilaginous repair tissue for a resected TFCC. Such therapy could be used in the arthroscopic treatment of TFCC injury and resection in an effort to improve postoperative outcome. PMID- 10705323 TI - Mini-deltoid splitting rotator cuff repair: do results deteriorate with time? AB - To determine whether the results of arthroscopically assisted rotator cuff repair deteriorate with time, 60 shoulders were evaluated on 2 separate occasions. There were 7 small, 16 medium, 20 large, and 17 massive tears. Patients were evaluated with a detailed questionnaire that included the UCLA shoulder scale and physical examination. Average initial follow-up was at 21 months (range, 12 to 68 months), and the second follow-up was at an average of 62 months (range, 24 to 103 months); only 4 patients had a change of more than 3 points on the UCLA scoring scale. No statistically significant difference was found in pain, function, range of motion, strength score, satisfactory results (80% on both occasions), and UCLA score (30.8 v 31.4) at second follow-up. The results of our study indicate that there is no significant deterioration over time of results of rotator cuff repair using the mini-deltoid splitting technique. PMID- 10705324 TI - Addressing glenohumeral stiffness while treating the painful and stiff shoulder arthroscopically. AB - The shoulder can be primarily or secondarily stiff. Cadaveric cutting studies have shown increases in passive range of glenohumeral motion when certain portions of the capsule are released. This study has recorded the intraoperative gains made in passive range of motion for external rotation, flexion, abduction, and internal rotation with sequential release of the rotator interval, inferior capsule, and posterosuperior capsule, regardless of initial etiology, and followed-up over time. Thirty one of 60 shoulders, found clinically to have a loss of passive range of motion and having failed a nonoperative approach, underwent a capsular release. Eighteen patients underwent a partial capsular release (group 1) and 13 patients (group 2) underwent a complete capsular release. Thirty of 31 shoulders had statistically significant gains in passive range of motion with sequential release. In general, resection of the rotator interval contributed to gains in external rotation; resection of the inferior capsule (anteroinferior and posteroinferior) contributed gains to external rotation, forward flexion, and internal rotation; and resection of the posterosuperior capsule contributed to gains only in internal rotation. At a minimum of 18 months follow-up, 30 of 31 shoulders retained their intraoperative gains. There was no difference in the results between primarily and secondarily stiff shoulders for motion gains (P >.05). Arthroscopically addressing capsular tightness is beneficial in returning shoulders with a loss of passive glenohumeral motion to normal regardless of the etiology. PMID- 10705325 TI - Measurement of brake response time after right anterior cruciate ligament reconstruction. AB - Recommendations on safe driving after anterior cruciate ligament (ACL) reconstruction have been largely intuitive. This study evaluated 12 male patients who underwent ACL reconstruction with subsequent outpatient rehabilitation and compared them with 10 subjects who had no knee dysfunction. The following clinical measures were assessed every 2 weeks for 10 weeks: brake response time (BRT), 6-meter walk time (6MWT), knee range of motion (ROM), pain (visual analog scale), and joint effusion. Statistical testing was completed using analysis of covariance with repeated measures. The results from treatment group were compared with norms from the AAA Traffic Safety and Engineering Department. BRT showed significant differences over 10 weeks (P =.043) in the study group. There were no significant differences between the study and control group based on condition (ACL reconstruction v control) (P =.586). Pain and effusion were found to have no significant interaction effect on BRT. The treatment group's BRT increased from the 25th percentile (AAA normals) to the 87th percentile after 10 weeks of rehabilitation. Although treatment group BRT was equal to AAA normal population BRT at week 4, the large improvement from week 2 to week 4 meant that learning effects could not be ruled out until week 6. Significant differences were found for 6MWT between week 2 and all other weeks (P <.0001). The results suggest brake reaction time matches control times at 4 to 6 weeks. Thus, BRT might be used to establish return to driving criteria (in part) after ACL reconstruction if other driving impediments do not exist. PMID- 10705326 TI - Intra-articular mechanical blocks and full extension in patients undergoing anterior cruciate ligament reconstruction. AB - Patients with acute anterior cruciate ligament (ACL) rupture frequently present with a lack of full extension. Current literature is unclear whether arthroscopic debridement is necessary before reconstruction to achieve full extension postoperatively. This study examined the postoperative extension achieved in 153 knees that underwent ACL reconstruction within 12 weeks of index injury. All patients performed preoperative physical therapy to increase range of motion and control pain/swelling, regardless of presenting range of motion without prior aspiration or arthroscopy. Of the 153 knees, 103 had meniscal pathology, of which 73 were peripheral vertical tears; 96 of the 153 knees lacked >/=3 degrees extension preoperatively. Five of 96 knees had an intra-articular mechanical block to extension and all regained full extension after ACL reconstruction. This study documented that a true intra-articular mechanical block is unusual in primary ACL ruptures. Lack of full extension can be adequately dealt with during surgical reconstruction without a detrimental effect on knee extension postoperatively. PMID- 10705327 TI - Refilling of removal defects: impact on extensor mechanism complaints after use of a bone-tendon-bone graft for anterior cruciate ligament reconstruction. AB - In a prospective study, the impact of refilling removal defects at the tibial tuberosity and patella on extensor mechanism complaints was studied after anterior cruciate ligament reconstruction with a bone-tendon-bone ligament graft. A conventional cannulated drill was sued in patients of group 1 (n = 62), and an oscillating hollow saw was used in group 2 (n = 70) for drilling and removing bone material from the tibial tunnel. On completing the procedure in the patients in group 2, the bone material removed was inserted into the bony removal defects at the tibial tuberosity and the patella. In both groups, the extensor mechanism was closed using loose sutures. Both groups underwent the same postoperative therapy regimen. All patients underwent clinical and radiographic follow-up after approximately 34 months. In this follow-up, no differences with regard to knee function as assessed by the IKDC rating scale and KT-1000 measurements were found between the 2 groups. The rate of mild to severe patellofemoral complaints was 31% (41 patients). Complaints related to the removal defects as assessed by kneeling and squatting on the operated knee were described by 25 patients (18.9%). No significant difference between the groups with and without refilling of the removal defects was observed, and the overall subjective ratings on a visual analog scale (VAS) (0 = no pain, 10 = severe pain) for kneeling and squatting were 5.57 and 3.69, respectively. Follow-up time was identified as the critical parameter for problems related to the extensor mechanism, such as complaints in kneeling and squatting, with hardly any problems observed after 2 years. On the other hand, anterior knee pain correlated significantly with the residual limitation of motion. PMID- 10705328 TI - Notchplasty in anterior cruciate ligament reconstruction: an experimental animal study. AB - The purpose of this study was to evaluate the effects of a notchplasty on the biomechanical and histological properties of the anterior cruciate ligament (ACL) and changes in patellar articular cartilage. We used an in situ freeze-thaw model for the ACL reconstruction performed with or without notchplasty in 36 Japanese white rabbits. The cross-sectional area of the regenerated ACLs without a notchplasty (6.4 +/- 0.4 mm(2)) was statistically smaller than that of the ACLs with a notchplasty (7.0 +/- 0.3 mm(2)), and the cross-sectional area of the normal ACLs (5.4 +/- 0.5 mm(2)) was statistically smaller than that of both other types. However, the mechanical strength of the ACL with the notchplasty was identical to that of the ACL without a notchplasty. Although the notchplasty areas were covered with fibrous scar tissue, caliper measurement and histological examination showed no obvious osteochondral reconstitution in the notchplasty sites of any of the specimens. Regarding the deleterious effect of notchplasty on patellar articular cartilage, the extent of the slight degenerative changes was about the same in the group with a 1-mm notchplasty and in the control group. PMID- 10705329 TI - An unusual epidemic of Staphylococcus-negative infections involving anterior cruciate ligament reconstruction with salvage of the graft and function. AB - We performed a retrospective study of 13 patients who had postoperative clinical and laboratory signs of infection after autogenous bone-patellar tendon-bone (BPTB) anterior cruciate ligament (ACL) reconstructions. From January 1991 to November 1996 we experienced only 2 infected knees in 1,300 reconstructions, but between December 1996 and February 1997 10 patients in 70 ACL reconstructions developed a postoperative suspected infection. We found the origin of contamination (coagulase-negative Staphylococcus) in the supposedly sterile inflow cannula. When we changed this device, we had only 1 infection in the next 400 reconstructions. The diagnosis in these cases was derived from clinical signs and laboratory results, but only 2 of 11 samples of aspirated synovial fluid tested positive for Staphylococcus. The mean interval between the surgery and the onset of signs of infection and the start of antibiotic therapy was 7.7 days. All the patients had antibiotic association at the highest level. Six knees underwent arthroscopic debridement when the clinical signs indicated resistence to antibiotics. The normal postoperative rehabilitation program was modified but was not discontinued. Although recovery time was longer, overall results were similar to uncomplicated reconstructions. On the basis of our experience, we believe that when there is a notable increase in infection rates, a thorough search for contamination is indicated. Our source of infection was material that was thought to be sterile. Ultimately, early diagnosis and treatment is of critical importance to obtain good results. Even suspicion of infective postoperative complication should be sufficient cause to search for responsible microorganisms and begin antibiotic therapy. Arthroscopic debridement should be proposed to patients with resistence to antibiotics. PMID- 10705330 TI - A comparison of results in middle-aged and young patients after anterior cruciate ligament reconstruction. AB - The aim of this retrospective study was to compare the results after arthroscopic anterior cruciate ligament (ACL) reconstruction in middle-aged and young patients. From our database (including 604 patients with a follow-up rate of 95%), we extracted all the patients over 40 years of age (group A, n = 30) and compared them with a group of patients from the same material, aged between 20 and 24 years (group B, n = 37). The groups were comparable in terms of the male:female ratio and surgical techniques. The follow-up was performed by independent observers. The median follow-up period was 31 months (range, 22 to 60 months) in group A and 38 months (24 to 60 months) in group B (P =.014). Before injury, the Tegner activity level was 6 (4-9) in group A and 9 (4-9) in group B (P <.001). At follow-up, the Tegner activity level was 5 (3-9) in group A and 6 (3-9) in group B (P =.032). At the follow-up, there was no difference in terms of the Lysholm score, which was 91 (37-100) and 89 (38-100) points in group A and group B, respectively. Using the IKDC evaluation system, 10 patients (33%) were classified as normal, 12 (40%) as nearly normal, 6 (20%) as abnormal, and 2 (7%) as severely abnormal in group A, compared with 8 (22%) normal, 18 (48%) nearly normal, 10 (27%) abnormal, and 1 (3%) severely abnormal in group B (NS). The 1 leg hop quotient was 90% (52-167) in group A and 93% (70-118) in group B (P =.056). The KT-1000 measurement showed an anterior side-to-side laxity difference of 2.0 mm (-4 to 8.5 mm) in group A and 2.0 mm (-2.5 to 8.0 mm) in group B (not significant). The middle-aged patients were subjectively more pleased with the results than the younger patients. There were no differences in either early or late complications between the groups. At the index operation, 11 of 30 patients (37%) in group A and 1 of 37 (3%) in group B had cartilage lesions or degenerative changes (P <.001). Age does not appear to disqualify middle-aged patients with symptomatic ACL tears from undergoing reconstruction. PMID- 10705331 TI - Can local anesthesia be recommended for routine use in elective knee arthroscopy? A comparison between local, spinal, and general anesthesia. AB - Local anesthesia (LA) for outpatient knee arthroscopy is not a standard procedure at most hospitals. To evaluate the LA technique for knee arthroscopy on medically healthy patients, this study compared 3 anesthesia techniques. Four hundred patients were randomized to either local (n = 200), general (n = 100), or spinal (n = 100) anesthesia. Evaluated outcomes included the patient's subjective view of the procedure, and nausea and pain at rest and during active movement. All variables were recorded perioperatively and postoperatively. In addition, the performing surgeon's opinion of the degree of patient pain and the technical difficulty of the procedure were noted. Three hundred forty-two patients completed the study. In the group receiving local anesthesia (n = 180) the median visual analog scale pain score during surgery was 6 mm (mean, 17.5; SD, 23.2; range, 0 to 100 mm). Twenty-one LA patients would have preferred another form of anesthesia. In 29 patients, LA was not considered as the optimal anesthesia by the performing surgeon. Eight LA patients agreed with the surgeon that the anesthesia method used was not optimal, of these patients, 5 had synovitis (3%). In 5% of the LA patients there were technical problems. Thus, this study shows that elective knee arthroscopy can be performed under local anesthesia in 92% of the patients from a technical point of view. Excluding patients who do not choose local anesthesia and those who have hypertrophic synovitis preoperatively, knee arthroscopies can be performed as safely and effectively under local anesthesia as under any other form of anesthesia. For most patients, local anesthesia can be recommended as the standard procedure for outpatient knee arthroscopy. PMID- 10705332 TI - Autogenous tendon graft substitution for absent knee joint meniscus: a pilot study. AB - The purpose of this pilot study was to explore the potential of an autogenous tendon graft to substitute for an absent human knee joint meniscus. Based on the results of animal studies and human reports, it was hypothesized that autogenous tendon tissue would substitute for human knee joint meniscus: maintain mechanical integrity, convert to fibrocartilage, preserve the joint compartment, and provide symptomatic relief for the patient. Five patients, 2 men and 3 women, average age 41 years, had surgical absence of the lateral meniscus, genu valgum, and severe degenerative arthritis of the lateral compartment, but a stable knee. All patients were offered alternative treatments: do nothing, medication, arthroscopic debridement, osteotomy, and knee replacement. The operations were performed by arthroscopy. An accompanying arthroscopic debridement procedure was performed in the same compartment. In 4 cases, the donor graft was the semitendinosus tendon. In 1, the patellar tendon was used because the semitendinosus had been previously used in an anterior cruciate ligament reconstruction. Four of the 5 patients had a second-look arthroscopy and biopsy between 9 and 24 months. There was partial physical integrity to the tendon graft. The tendon graft did not completely convert to fibrocartilage. The joint surface was not preserved. Only 1 patient had minimal clinical improvement; the others were not improved. No patient was made worse. One patient had a total knee replacement 1 year later. Another had a knee fusion after 4 years. All other patients are considering future reconstructive surgery. The autogenous tendon graft as used in this pilot study was not successful as a substitute for an absent meniscus. The hypothesis was not realized. The observations from this pilot study should be helpful in future study protocol design. PMID- 10705333 TI - Arthroscopic treatment of post-traumatic cysts of the talus. AB - We report on 9 patients with persistent ankle pain and radiographic evidence of a cystic lesion on the talus. All had a history of an inversion-type of ankle injury. Radiographs were initially negative, but a cyst developed about 6 months after the injury. Arthroscopic debridement revealed extrusion of viscid gelatinous material from the cyst. The cavity was arthroscopically abraded to bleeding base. Follow-up of these patients at an average of 26 months showed statistically significant improvement in terms of pain, swelling, stiffness, limp, and activity level. Bone grafting may not be necessary in the treatment of post-traumatic cysts. PMID- 10705334 TI - Knot security in simple sliding knots and its relationship to rotator cuff repair: how secure must the knot be? AB - We sought to determine which simple sliding knot configurations would have adequate strength for rotator cuff repair. Four knot configurations were tied with both No. 1 polydioxanone suture and No. 2 Ethibond suture (Ethicon, Somerville, NJ) using 3 different tying techniques: hand-tie, standard knot pusher, and cannulated double-diameter knot pusher. The knots were then tested to failure on a materials testing system. The weakest standard knot configuration was S=S=S=S. The other 3 knot configurations (S//S//S//S, SxSxSxS, and S//xS//xS//xS) generally failed in the 35 to 50 N range. Ultimate strength in this range can be shown to be adequate to withstand, without suture failure, a maximal contraction of a repaired rotator cuff tear within the rotator crescent, assuming certain conditions are met (suture anchors placed 1 cm apart, 2 sutures per anchor). More complex knots are not necessary for adequate knot security. However, the same configuration with only 1 suture per anchor will not be strong enough because the suture will fail under maximum physiological load. This study shows that we can predict the adequacy of a given knot configuration under maximum physiological loading conditions. PMID- 10705335 TI - Preiser's disease: arthroscopic treatment of avascular necrosis of the scaphoid. AB - We report the case of a 50-year-old female patient with Preiser's disease (avascular necrosis of the scaphoid) who, after a 2-year history of wrist pain, underwent arthroscopic debridement of the necrotic scaphoid using a standard technique. Osteoarthritic changes of the articular cartilage, partial rupture of the scapholunate ligament, local synovitis, and loose fragments were documented. The patient reported significant improvement in pain relief and complete relief of mechanical symptoms at 31-month follow-up examination. Radiographs demonstrated no progression of collapse of the scaphoid or acceleration of degenerative changes in the wrist. Arthroscopy in Preiser's disease allows direct visualization and assessment of the exact pathology of the radiocarpal and midcarpal joint and the scaphoid cartilage. Arthroscopic debridement of the necrotic scaphoid increased wrist functional range of motion, provided excellent pain relief, and improved health-related quality of life. PMID- 10705336 TI - The pitcher's mound: a late sequela of posterior type II SLAP lesions. AB - Three patients with long-standing disabling shoulder pain underwent arthroscopic examination. Two of the 3 had preoperative magnetic resonance imaging scans showing complete rotator cuff tears confirmed at surgery. The third patient was found to have a partial-thickness bursal surface cuff tear. In addition, each patient was found to have a quite prominent posterior superior glenoid osteophyte located beneath an unstable type II SLAP lesion. PMID- 10705337 TI - The axial view in evaluating tibial translation in cases of insufficiency of the posterior cruciate ligament. AB - The purpose of this report is to present a new radiological method of diagnosis and evaluation of posterior instability using the patellofemoral axial view. During a period of 22 months, we performed clinical and radiological assessments on 20 patients (6 acute and 14 chronic) with isolated posterior instability caused by posterior cruciate ligament (PCL) rupture and on 20 patients with normal knees. The radiological examination included stress radiographs using the Telos device (Telos, Griesheim, Germany) as well as a modification of the routine axial patellofemoral view. Both diagnosis and quantification of the posterior tibial translation was possible in all cases by measuring, on the axial view, the distance between the anterior edge of the tibial plateau and the center of the femoral groove (trochlea). Clinical examination, conventional radiography, KT 1000 arthrometry, stress radiography at 90 degrees and at 20 degrees of flexion, and magnetic resonance imaging all assist in diagnosing a PCL tear. This new radiographic technique is simple, fast, and consistently effective both in patients with acute and those with chronic PCL tears, as well as in those who have undergone PCL reconstruction. PMID- 10705338 TI - Distraction in the lateral position in elbow arthroscopy. AB - To perform successful elbow arthroscopy, it is very important to have precise positioning and to distract the joint during arthroscopy. We have developed a method of distraction during elbow arthroscopy that enlarges the articular space while ensuring the stability of the elbow. Our method is simple and requires no expensive specialized instruments. Our distraction method using lateral positioning is useful and safe for elbow arthroscopy. Because the articular space is widened sufficiently, stability of the elbow is maintained and no complication was seen. PMID- 10705342 TI - Protein tyrosine phosphatase (PC12, Br7,S1) family: expression characterization in the adult human and mouse. AB - Protein tyrosine phosphatases (PTPs) play important roles in modulating signals transduced by tyrosine kinases. Certain phosphatases have been implicated as having important roles in embryonic development as well as in adult physiology. Although both kinases and phosphatases are equally important in regulating signal transduction, phosphatases as a group have not been well characterized. Thus, characterization of sequence, expression, and biological function for additional phosphatases is informative. PTPBr7/PC12 and PTPSl are mouse receptor PTPs sharing similar amino acid sequences. Northern blot analysis demonstrated expression of these genes in adult rodent brain and revealed previously uncharacterized transcripts in the brain and other tissues. Our results demonstrate that PTPBr7/PC12 and PTPSl are members of a larger family of PTPs. We have identified two novel family members as well as several novel transcriptional splice variants from both human and mouse colon cDNA libraries. Expression analysis demonstrated that the various mRNA transcripts are differentially expressed, with the highest levels found in the brain, intestinal tract, uterus, and placenta. In situ hybridization analysis of mouse brain and intestinal tissues established that each isoform has a unique expression pattern in specific cell populations as well as in tissue regions. Furthermore, these restricted patterns suggest that the encoded family of phosphatases may play roles in modulating signal transduction pathways important for specific cell types and biological processes. PMID- 10705343 TI - Development of Meissner-like and Pacinian sensory corpuscles in the mouse demonstrated with specific markers for corpuscular constituents. AB - The development of Meissner-like and Pacinian corpuscles was studied in mice [from postnatal day (Pd) 0 to 42] by using immunohistochemistry for specific corpuscular constituents. The battery of antigens investigated included PGP 9.5 protein and neurofilaments, as markers for the central axon; S100 protein, vimentin, and p75(LNGFR) protein, to show Schwann-related cells; and epithelial membrane antigen to identify perineurial-related cells. In Meissner-like corpuscles immunoreactivity (IR) for neuronal markers was found by Pd7 and later. The lamellar cells of these corpuscles expressed first S100 protein IR (Pd7 to Pd42), then vimentin IR (Pd12 to Pd42), and transitory p75(LNGFR) IR (Pd7 to Pd19 20). Vimentin IR, but not epithelial membrane antigen, was detected in the capsule-like cells of the Meissner-like corpuscles. On the other hand, the density of Meissner-like corpuscles progressively increased from Pd0 to Pd19-20. Pacinian corpuscles were identified by Pd7. From this time to Pd42 the central axon showed IR for neuronal markers, and the inner core cells were immunoreactive for S100 protein. Moreover, vimentin IR was detected in the inner core cells by Pd19 and later. Unexpectedly, the central axons displayed S100 protein IR (from Pd7 to P28), while p75(LNGFR) protein IR or epithelial membrane antigen IR were never detected. Taken together, and based on the expression of the assessed antigens alone, the present results suggest that the Meissner-like and the Pacinian corpuscles in mice become mature around Pd19-Pd28 and Pd20, respectively. Furthermore, these results provide a baseline timetable for future studies in the normal or altered development of sensory corpuscles in mice since specific sensory corpuscles are functionally associated with different subtypes of sensory neurons the development of which is selectively disturbed in genetically manipulated mice. PMID- 10705345 TI - Colloid in the pituitary pars distalis of viscacha (Lagostomus maximus maximus): ultrastructure and occurrence in relation to season, sex, and growth. AB - Randomly distributed extracellular colloidal accumulations were observed in the pars distalis of viscacha (Lagostomus maximus maximus). They were preferentially located in the peripheral zone of the gland and showed variability in shape and size. Two different types of colloidal accumulations were found by electron microscopy: 1) those surrounded by nongranulated follicular cells that correspond to characteristic follicles, and 2) those surrounded by granulated cells. In the follicles lined by nongranulated follicular cells, long, prominent microvilli and cytoplasmic processes protruded into the lumen. The frequency of these accumulations varies during the year in adult male animals, showing an increase in number during summer and a decrease during winter. The lowest value was registered in August (winter). The mean follicular diameter did not vary seasonally. The number of colloidal accumulations did not vary seasonally in adult female viscachas, but a significant difference in the mean follicular diameter between pregnant and non-pregnant females was observed. Pituitaries of immature animals contain fewer colloidal accumulations than those of adults. In fetuses, these accumulations were absent. The administration of melatonin provoked a decrease in the number of these structures. The numeric changes of the colloidal accumulations observed in this study are associated with: 1) the seasonal reproductive activity in adult males, and 2) the reproductive condition, body weight and sexual maturity in males and females. The fact that melatonin administration decreases the population of colloidal accumulations in males suggests participation of the pineal gland in these changes. PMID- 10705344 TI - Extensive glycosylation changes revealed by lectin histochemistry in morphologically normal prenatal tissues of the mouse mutant undulated (un/un). AB - Recently we observed that in human embryos and fetuses with a variety of malformations, not only malformed tissues, but also several non-malformed tissues displayed alterations in the glycosylation pattern. It was the aim of this work to investigate this more or less inexplicable phenomenon under experimental conditions. To this end, we studied a well known mouse model, the mouse mutant undulated, which has an exactly defined genetic defect (substitution in the pax-1 gene) leading to a localized malformation in the vertebral column. The glycosylation pattern was studied using lectin histochemistry. Distribution of binding sites for the lectins RCA I, Con A, SNA, SBA, PNA, LTA and WGA was studied during the organogenesis stages (i.e., days 11-18). It was striking that in mutants, changes in the glycosylation pattern were found not only in the malformed organ (i.e., vertebral anlage), but also in other embryonic tissues, which showed normal morphology. This suggests that the altered glycosylation seems to be a part of genetically determined phenomena throughout the entire organism. Our results show that a defect in a gene with a very restricted expression can cause universal changes in the glycosylation pattern during development. PMID- 10705346 TI - Characterization of nonmuscle cells present in the intima of normal adult bovine pulmonary artery. AB - In this study, the presence of cells in the intimal region of normal adult bovine pulmonary artery (BPA) was examined by analysis of longitudinal sections at the level of light and transmission electron microscopy. In addition, the morphological and immunohistochemical phenotype of these cells as well as the presence of particular extracellular matrix (ECM) components in this region were also determined. Since ECM production and cell proliferation have been demonstrated to be regulated by locally released growth factors such as transforming growth factor beta (TGFbeta), the presence of TGFbeta-1 in this region was also investigated. Our findings reveal the presence of immature or "nonmuscle" cells into the subendothelial space of normal adult BPA. These cells were characterized by the presence of abundant cytoplasmic organelles and scanty microfilaments. Such cells were negative to antibodies against smooth muscle alpha actin (SM alpha-actin), 1E12, and vWf, but not to vimentin. Similar cells have recently been detected in normal adult BPA and canine carotid arteries, but in the medial region. Because of their location in these elastic arteries, the nonmuscle cells are involved not only in the remodeling of the medial region, but also in the neointima or intimal thickening formation by migration from the media to the subendothelial space, where they proliferate and secrete ECM components. However, a limited number of morphological studies and the current investigation describe the presence of scattered nonmuscle cells within the intima of some normal elastic arteries. This would suggest an important role for these resident cells within the intima in normal and pathological processes as well. In addition, our results show the presence, in this region, of TGFbeta-1 and of ECM components that include collagen, elastin, fibronectin, and laminin which are present in normal conditions and during the intima formation in vivo. PMID- 10705347 TI - Immunohistochemical localization of carbonic anhydrases I, II, and VI in the developing rat sublingual and submandibular glands. AB - Carbonic anhydrase has been localized to the acini and ducts of mature rat salivary glands. This enzyme has been associated with ion transport, a prominent function of striated and excretory ducts in salivary glands, suggesting that it might be used as a marker of ductal differentiation. The purpose of this study was to immunohistochemically document developmental changes in carbonic anhydrase in the ducts of the rat sublingual and submandibular glands. Immunohistochemistry was performed with antibodies to human carbonic anhydrase isoenzymes I, II and VI on sections of sublingual and submandibular glands from rats at representative postnatal developmental ages. Reactions were weak in the ducts of both glands at 1 day, then progressively increased. By 42 days, reactions had the adult pattern of virtually none in the mucous or seromucous acini, moderate to strong in the striated and excretory ducts, and none to weak in the intercalated ducts. Weak to moderate reactions were observed in the granular convoluted tubules of the submandibular gland as they became recognizable at age 42 days. Reactions to carbonic anhydrase I and II antibodies also increased from none (1 day) to modest (42 days) in the demilunes of the sublingual gland. The order of reaction intensity of the antibodies was II > I > VI. When localized via these anti-human antibodies, carbonic anhydrase is a useful marker of the functional differentiation of the striated and excretory ducts of the developing rat sublingual and submandibular glands. PMID- 10705348 TI - Changes in peptidergic nerves in the atrioventricular valves of streptozotocin induced diabetic rats: a confocal microscopy study. AB - Several previous studies have described the distribution of neuropeptide Y (NPY) like and calcitonin gene related peptide (CGRP)-like immunoreactive nerve fibres in the atrioventricular valves of humans and various animals. It has been suggested that peptide-containing nerve fibres might have motor or sensory roles in valvular function. Although there is evidence that diabetic changes occur in the sympathetic (preganglionic and postganglionic), parasympathetic (vagal) and peptidergic nerves of rats, the changes of peptide-containing nerve fibres in the atrioventricular valves of the diabetic rat have not been studied. The distribution, relative density and staining intensity of NPY-like and CGRP-like immunoreactive nerve fibres in the mitral and tricuspid valves were studied in whole mount preparations using confocal microscopy with a computer-assisted image analysis system. Streptozotocin-induced diabetic and control rats were sacrificed at 12 and 24 months. The nerve staining intensity within the tricuspid valve was greater than the mitral valve in both control (P < 0.01) and diabetic (P < 0.001) rats. Nerve density in the anterior leaflet was greater than the posterior leaflet of the mitral valve. However, the anterior leaflet of the mitral and tricuspid valves showed a decreased number of nerve fibres, followed by drastic reduction in the staining intensities for both the peptides studied (P < 0.001) in the long-term diabetic rat. The decrease in the number of nerve fibres that follow the mechanical interruption of nerves raises the possibility that cycles of degeneration may occur. It is suggested that these peptide-containing nerve fibres in the atrioventricular valves may be involved in valvular dysfunction in the diabetic state. PMID- 10705349 TI - Spleen of Dasypus hybridus (Mammalia, dasypodidae): a light and electron microscopic study. AB - Armadillos are relictual mammals important as models for biomedical studies. They contain adaptative and primitive characteristics in both anatomical and physiological aspects. In this study we describe the splenic histology and cytology of the "mulita," Dasypus hybridus. Organ samples were processed for light and electron microscopy study. The microanatomy of the organ samples as well as their different cell types are described. The spleen is non-sinusoidal, with the typical arrangement for storage functions. White pulp is lightly diffuse. Red pulp is a meshwork of circulating, immunocompetent and hemopoietic cells. Differences with other studied members of the group are discussed. The general structure of the organ agrees with the semi-fossorial habit of the species. Persistence of myeloid activity in the adult suggests the existence of specific inductive functions of the stroma. Better knowledge of this fact may give further insight on the phylogeny of hemopoiesis. PMID- 10705350 TI - Remodeling of the vascular bed and progressive loss of capillaries in denervated skeletal muscle. AB - Very little is known regarding structural and functional responses of the vascular bed of skeletal muscle to denervation and about the role of microcirculatory changes in the pathogenesis of post-denervation muscle atrophy. The purpose of the present study was to investigate the changes of the anatomical pattern of vascularization of the extensor digitorum longus muscle in WI/HicksCar rats 1, 2, 4, 7, 12, and 18 months following denervation of the limb. We found that the number of capillaries related to the number of muscle fibers, i.e. the capillary-to-fiber ratio (CFR), decreased by 88%, from 1.55 +/- 0.35 to 0.19 +/- 0.04, during the first 7 months after denervation and then slightly declined at a much lower rate during the next 11 months of observation to 10% of the CFR in normal muscle. Between months 2 and 4 after denervation, the CRF decreased by 2.4 times, from 58% to 24% of the control value. The loss of capillaries during the first 4 months following nerve transection was nearly linear and progressed with an average decrement of 4.16% per week. Electron microscopy demonstrated progressive degeneration of capillaries following nerve transection. In muscle cells close to degenerating capillaries, the loss of subsarcolemmal and intermyofibrillar mitochondria, local disassembly of myofibrils and other manifestations of progressive atrophy were frequently observed. The levels of devascularization and the degree of degenerative changes varied greatly within different topographical areas, resulting in significant heterogeneity of intercapillary distances and local capillary densities within each sample of denervated muscle. Perivascular and interstitial fibrosis that rapidly developed after denervation resulted in the spatial separation of blood vessels from muscle cells and their embedment in a dense lattice of collagen. As a result of this process, diffusion distances between capillaries and the surfaces of muscle fibers increased 10-400 times. Eighteen months after denervation most of the capillaries were heavily cushioned with collagen, and on the average 40% of the muscle cells were completely avascular. Devascularization of the tissue was accompanied by degeneration and death of muscle cells that had become embedded in a dense lattice of collagen. Immunofluorescent staining for the vascular isoform of alpha-actin revealed preservation of major blood vessels and a greater variability in thickness of their medial layer. Hyperplastic growth of the medial layer in some blood vessels resulted in narrowing of their lumens. By the end of month 7 after denervation, large deposits of collagen around arterioles often exceeded their diameters. Identification of oxidative muscle fibers after immunostaining for slow-twitch myosin, as well as using ultrastructural criteria, has shown that after 2 months of denervation oxidative muscle fibers were less susceptible to atrophy than glycolytic fibers. The lower rate of atrophy of type I muscle fibers at early stages of denervation may be explained by their initially better vascularization in normal muscle and their higher capacity to retain capillaries shortly after denervation. Thus, degeneration and loss of capillaries after denervation occurs more rapidly than the loss of muscle fibers, which results in progressive decrease of the CFR in denervated muscle. The change of capillary number in denervated muscle is biphasic: the phase of a rapid decrease of the CFR during the first 7 months after nerve transection is followed by the phase of stabilization. The presence of areas completely devoid of capillaries in denervated muscle and the virtual absence of such areas in normal muscle indicate the development of foci of regional hypoxia during long-term denervation. The anatomical pattern of muscle microvascularization changes dramatically after nerve transection. Each muscle fiber in normal muscle directly contacts on average 3-5 capillaries. (ABSTRACT TRUNCATED) PMID- 10705351 TI - Cell death in denervated skeletal muscle is distinct from classical apoptosis. AB - Denervation of skeletal muscle is followed by the progressive loss of tissue mass and impairment of its functional properties. The purpose of the present study was to investigate the occurrence of cell death and its mechanism in rat skeletal muscle undergoing post-denervation atrophy. We studied the expression of specific markers of apoptosis and necrosis in experimentally denervated tibialis anterior, extensor digitorum longus and soleus muscles of adult rats. Fluorescent staining of nuclear DNA with propidium iodide revealed the presence of nuclei with hypercondensed chromatin and fragmented nuclei typical of apoptotic cells in the muscle tissue 2, 4 and to a lesser extent 7 months after denervation. This finding was supported by electron microscopy of the denervated muscle. We found clear morphological manifestations of muscle cell death, with ultrastructural characteristics very similar if not identical to those considered as nuclear and cytoplasmic markers of apoptosis. With increasing time of denervation, progressive destabilization of the differentiated phenotype of muscle cells was observed. It included disalignment and spatial disorganization of myofibrils as well as their resorption and formation of myofibril-free zones. These changes initially appeared in subsarcolemmal areas around myonuclei, and by 4 months following nerve transection they were spread throughout the sarcoplasm. Despite an increased number of residual bodies and secondary lysosomes in denervated muscle, we did not find any evidence of involvement of autophagocytosis in the resorption of the contractile system. Dead muscle fibers were usually surrounded by a folded intact basal lamina; they had an intact sarcolemma and highly condensed chromatin and sarcoplasm. Folds of the basal lamina around the dead cells resulted from significant shrinkage of cell volume. Macrophages were occasionally found in close proximity to dead myocytes. We detected no manifestations of inflammation in the denervated tissue. Single myocytes expressing traits of the necrotic phenotype were very rare. A search for another marker of apoptosis, nuclear DNA fragmentation, using terminal deoxyribonucleotidyl transferase mediated dUTP nick end labeling (the TUNEL method) in situ, revealed the presence of multiple DNA fragments in cell nuclei in only a very small number of cell nuclei in 2 and 4 month denervated muscle and to less extent in 7 month denervated muscle. Virtually no TUNEL reactivity was found in normal muscle. Double labeling of tissue denervated for 2 and 4 months for genome fragmentation with the TUNEL method and for total nuclear DNA with propidium iodide demonstrated co-localization of the TUNEL-positive fragmented DNA in some of the nuclei containing condensed chromatin and in fragmented nuclei. However, the numbers of nuclei of abnormal morphology containing condensed and/or irregular patterns of chromatin distribution, as revealed by DNA staining and electron microscopy, exceeded by 33-38 times the numbers of nuclei positive for the TUNEL reaction. Thus, we found a discrepancy between the frequences of expression of morphological markers of apoptosis and DNA fragmentation in denervated muscle. This provides evidence that fragmentation of the genomic DNA is not an obligatory event during atrophy and death of muscle cells, or, alternatively, it may occur only for a short period of time during this process. Unlike classical apoptosis described in mammalian thymocytes and lymphoid cells, non-inflammatory death of muscle fibers in denervated muscle occurs a long time after the removal of myotrophic influence of the nerve and is preceded by the progressive imbalance of the state of terminal differentiation. Our results indicate that apoptosis appears to be represented by a number of distinct isotypes in animals belonging to different taxonomic groups and in different cell lineages of the same organism. PMID- 10705352 TI - National Action Plan on Breast Cancer Workshop on Multicultural Aspects of Cancer Etiology. Washington, DC, USA. March 17-19, 1999. PMID- 10705353 TI - National Action Plan on Breast Cancer workshop on multicultural aspects of cancer etiology. A conceptual framework for the planning of ethno-oncology. AB - This presentation addresses the significance and implications of cancer research planning as it relates to the diversity of ethnic groups in the U.S. and indicates the limitations and potential benefits to be derived from such research. It stresses the importance of an adequate statistical data base and the relative importance of genetic and lifestyle factors in cancer etiology. PMID- 10705354 TI - Epidemiology, stage at diagnosis, and tumor biology of breast carcinoma in multiracial and multiethnic populations. AB - All women, regardless of their racial or ethnic origin or heritage, are at risk of developing breast cancer. Variations in breast carcinoma incidence rates among multicultural populations suggest that etiologic factors differ in their biologic expression and impact on disease outcome. Key among those factors that affect breast carcinoma development are the roles of genetics and the environment, the reproductive experience and the effects of endogenous and exogenous hormones in women, the change in immune status and host vulnerability, and the biologic determinants of breast carcinoma. Cultural dynamics, sociodemographic differences, and behavioral characteristics across population subgroups modulate how biologic disease is expressed among different races and ethnic groups. These interactions contribute to the observed variations in breast carcinoma incidence, mortality, and survival. Stage, a measure of disease status, is used to assess prognosis, plan treatment, and evaluate outcome. Numerous studies have reported a more advanced stage of breast carcinoma at diagnosis in racial/ethnic subgroups, especially among women from African American, Hispanic, American Indian, and native Hawaiian cultures. Factors associated with advanced stage at diagnosis in multicultural populations range from changes in the basic biological characteristics at the molecular and cellular level, to more complex behavioral attributes unique to a particular multicultural population, to societal issues such as access to care and socioeconomic conditions-all of which impact on the health measure called "stage at diagnosis." Rapid advancements in knowledge of cancer biology and of genetic markers and tumor products are providing new mechanisms for identifying etiologic pathways that can be utilized for better screening, detection, treatment and monitoring of disease. Further studies are needed that evaluate the biologic and molecular alterations in tumor development, progression, and response to therapy. Public health attention needs to be directed toward the societal influences that impact breast carcinoma development, as well as augmenting recognition of the need for culturally appropriate, broad based behavioral changes at the community level. In addition, continued efforts are needed to ensure the inclusion of multicultural population subgroups and minority investigators in all aspects of research-basic, clinical and applied. PMID- 10705355 TI - Migration patterns and breast carcinoma. AB - BACKGROUND: Many American Indian and Alaska Native women have lower incidence rates of breast carcinoma than other racial/ethnic groups in the United States. The rates in most areas, however, have increased in recent years. The author reviews the migration patterns and effects that might contribute to this change. METHODS: A review of the literature on migration and breast carcinoma incidence was conducted. RESULTS: Migration significantly impacts on breast carcinoma incidence in all groups of women studied. CONCLUSIONS: Research must be designed that will explore the components of host, life-styles, and environment on breast carcinoma rates in American Indian and Alaska Native women to elucidate mechanisms of breast carcinoma etiology. PMID- 10705356 TI - Urban native American health issues. AB - BACKGROUND: This article presents an overview of urban-dwelling American Indians and Alaska Natives, including a summary of data issues and a brief overview of historical and related social changes resulting in migration from reservations to urban areas. METHODS: A literature search was performed and documented focus groups were held; in addition, Native Sisters' field records from Los Angeles, California, and Denver, Colorado, were reviewed. RESULTS: Urban Indian communities are intertribal and represent over half of the Native American population in the U.S., yet they lack access to sufficient health services. Urban Indian clinics are greatly underfunded. CONCLUSIONS: A greater proportion of funding needs to be allocated to community-driven, culturally respectful, multiyear behavioral research to improve the screening, treatment, and survival of American Indian women with breast carcinoma. PMID- 10705357 TI - Changing the research paradigm: community involvement in population-based research. PMID- 10705358 TI - The excess burden of breast carcinoma in minority and medically underserved communities: application, research, and redressing institutional racism. AB - BACKGROUND: In 1998, the American Cancer Society, the National Cancer Institute, and the Centers for Disease Control and Prevention reported an overall downward trend in cancer incidence and mortality between 1990 and 1995 for all cancers combined. Many minority and medically underserved populations, however, did not share equally in these improvements. METHODS: A review of surveillance and other reports and recent literature on disparities in cancer incidence and mortality in minority and medically underserved communities was conducted 1) to ascertain the extent to which these communities bear an excess cancer burden, and 2) to explore the macrosocietal and microinstitutional barriers to equitable benefits in cancer health care delivery. RESULTS: Tragic disparities in cancer incidence and mortality in minority and medically underserved communities continue to be inadequately addressed. Overall improvements in U.S. cancer incidence and mortality rates are not shared equally by all segments of our society. While numerous individual and cultural barriers to optimal cancer control and care exist in minority and medically underserved communities, a major factor precluding these populations from sharing equally in advances in cancer research is prevailing societal and institutional racism. CONCLUSIONS: Immediate and equitable application of existing cancer control interventions and quality treatment options will significantly decrease cancer incidence and mortality. Enhanced surveillance efforts and a greater investment in targeted cancer research in those communities with the greatest disparities must be employed immediately if we are to achieve the goal of the president of the United States of eliminating racial and ethnic disparities in cancer and other diseases by 2010. Unless we acknowledge and redress institutionalized racism, the miscarriage of health justice will be perpetuated while celebrated advances in cancer research leading to declining incidence and mortality rates continue to evade our nation's minority and medically underserved communities. PMID- 10705359 TI - Ongoing research to identify environmental risk factors in breast carcinoma. AB - Environmental exposures, timing and duration of exposure, and one's genetic susceptibility all contribute to breast carcinoma and its progression. The purpose of this article was to identify known and suspected environmental causes of breast carcinoma, identify some environmental risk factors that may represent significant risk factors for certain groups, and describe current studies, supported by the National Institutes of Health/National Institute of Environmental Health Sciences, that clarify how environmental factors contribute to the development of breast carcinoma. Known and suspected environmental risk factors include organochlorine pesticides and other synthetic chemicals, hormonal factors (including exogenous endocrine disrupters), diet, tobacco and alcohol use, radiation, and magnetic fields. In at least 50% of breast carcinoma cases, none of the known risk factors apply. It is likely that an environmental component accounts for much of the unknown 50% of risk. Knowing the environmental factors for breast carcinoma development is an area that should be investigated intensely because it offers our best hope for prevention. Understanding why African-American women have a more aggressive form of breast carcinoma, whether they receive adequate follow-up treatment, and how these factors contribute to increased mortality rates requires further exploration. Data that demonstrate the lower incidence rate of breast carcinoma in Asian women, the relation to low fat diets and diets high in phytoestrogens, and how this might serve as a model for all women should be investigated. Finally, differences in behavioral and cultural attitudes, ethnicity, economic status, and life-style influences among different groups of women require further study to determine how these factors contribute to enhancing or reducing breast carcinoma risk. PMID- 10705360 TI - Breast carcinoma etiology: current knowledge and new insights into the effects of reproductive and hormonal risk factors in black and white populations. AB - BACKGROUND: A crossover in breast carcinoma incidence at ages 45-49 years has been observed between black and white women, with blacks experiencing higher incidence at younger ages and lower incidence after age 50 years. Can this phenomenon be partially explained by the differences in the distributions of reproductive risk factors? This article focuses on the effects and distributions of age at first full term pregnancy (FFTP), parity, and oral contraceptive (OC) use in younger versus older and black and white populations. Effects of hormone replacement therapy (HRT) are also summarized. METHODS: A literature review was conducted and information integrated on the effects and distributions of reproductive and hormonal risk factors in black and white populations, the crossover effect of parity, and the Pike model of "breast tissue age." RESULTS: Overall, early age at FFTP and higher parity decreased risk for both races. Distributions of age at FFTP and parity varied widely between the two races. Based on the effects and distributions of age at FFTP and parity, the authors formulated the hypothesis that a crossover in incidence curves between the two races would be expected, rather than be considered an anomaly. Regarding OC use, generally a stronger increase in risk was observed for younger women than for older women. Regarding HRT, a recent meta-analysis observed an increased risk of 1.35 for 5 years of use or more. CONCLUSIONS: To promote public health in diverse populations, and to provide further insight into breast carcinoma etiology, research needs to focus on multicultural differences and similarities in the relation of hormonal risk factors and breast carcinoma. PMID- 10705361 TI - Diet and breast carcinoma in multiethnic populations. PMID- 10705362 TI - Dietary and/or nutritional risk factors and breast carcinoma: consumer response. AB - The author is a former state legislator and feminist activist. Having traveled extensively in Asia, she questions the comparison of U.S. statistics regarding dietary/nutritional risk factors with those from countries whose public health standards are below that of the U.S. The author also notes that only recently has the U.S. had a critical mass of women in Congress that led to the funding of the first Women's Health Initiative. Countries in Asia have few women in positions of political power to advocate for public policies regarding women's health issues, especially concerning the gathering of data regarding breast carcinoma, which is considered a culturally sensitive topic. Although soy is considered a healthy staple of the majority of Asian diets because of its phytoestrogens, there is a mixed message to American consumers because cancer, rather than heart disease, is the leading cause of death among Asian females. PMID- 10705363 TI - Associations between energy balance and body mass index and risk of breast carcinoma in women from diverse racial and ethnic backgrounds in the U.S. AB - There is increasing epidemiologic evidence of an association between body mass index and energy expenditure and the risk of breast carcinoma. Women who are overweight or obese, especially women who gain weight throughout adulthood, are at an increased risk for developing breast carcinoma after menopause. Conversely, overweight women are at a reduced risk for developing breast carcinoma in the premenopausal years. The association between body mass index and breast carcinoma risk has been observed in women from several racial and ethnic backgrounds. Many studies have found an association between increased physical activity and reduced risk for breast cancer. Studies regarding physical activity and breast carcinoma risk have been conducted primarily with white women; therefore, the cross racial/ethnic patterns with this risk factor are unknown. This article reviews data regarding the associations between body mass index, physical activity, and breast carcinoma risk and presents potential mechanisms for the observed associations, such as sex hormones, reproduction, growth hormones, insulin, and immune function. It outlines challenges in measuring physical activity and body mass index across populations. Finally, the current study discusses limitations of the available data and suggests future research priorities. Obesity and a sedentary lifestyle may be two important risk factors for breast carcinoma that can be modified and thus may have significant public health impact in women from various racial and ethnic backgrounds. PMID- 10705364 TI - Socioeconomic factors and breast carcinoma in multicultural women. AB - Breast carcinoma is the most common cancer in women in the U.S. and the second leading cause of cancer death in women. Furthermore, there are racial differences in breast carcinoma incidence, mortality, and survival rates. Social and economic factors within racial/ethnic groups are being examined as risk factors not only for breast carcinoma mortality and survival but also as determinants of the rate of incidence. Social and economic factors have been associated in the literature predominantly with cancer mortality and survival. When socioeconomic status (SES) is considered, certain studies suggest that racial disparities in breast carcinoma are smaller than when social and economic factors are examined alone, but these disparities still persist. Sources of data for this discussion include the National Cancer Institute (NCI) (the Surveillance, Epidemiology, and End Results [SEER] program, a group of population-based cancer registries that cover up to 14% of the U.S. population. SEER reports cancer incidence, mortality, and survival rates), the U.S. Bureau of the Census, the National Center for Health Statistics (NCHS), and numerous articles from the scientific literature. Socioeconomic factors or SES can be considered "cross-cutting risk factors" (i.e., they can be related to the risk of developing breast carcinoma [rate of incidence] as well as to the risk of dying [mortality] from this disease). They also are the risk factors that "cut across" racial and ethnic populations. Socioeconomic factors are related to breast carcinoma mortality and survival rates in multicultural women. Racial disparities in breast carcinoma mortality and survival rates can be explained partially by stage distribution at the time of diagnosis, which may be related to SES. For example, African-American women present with more advanced stage distributions for breast carcinoma than white women. Similarly, women of lower SES present with higher stage disease than women of upper SES who present with more localized breast carcinoma. The lack of data regarding the SES of cancer patients limits our understanding of the contributions of SES to cancer incidence and mortality rates. SES appears to be related to breast carcinoma incidence, mortality, and survival rates. Breast carcinoma mortality is higher in women of lower SES. Additional research on SES, race, culture, and the relation of these factors to cancer incidence rate is needed. PMID- 10705365 TI - Multicultural aspects of breast cancer etiology workshop: concluding comments. PMID- 10705366 TI - Abstracts of poster presentations PMID- 10705367 TI - National Action Plan on Breast Cancer workshop on multicultural aspects of breast cancer etiology. Recommendations of the workshop. PMID- 10705368 TI - The GTP hydrolysis defect of the Saccharomyces cerevisiae mutant G-protein Gpa1(G50V). AB - The Saccharomyces cerevisiae haploid cell response to pheromone involves two seven-transmembrane-domain pheromone receptors that couple to a heterotrimeric G protein. The G50V mutation in the G protein alpha subunit (G(alpha)), Gpa1p, is analogous to the p21(ras) transforming mutation Gly-->Val 12, and has been extensively examined for the phenotypes it produces in yeast cells. Here we have characterized the Gpa1(G50V) mutant protein in vitro by examining GTPgammaS binding, GDP exchange, GTP occupancy and guanosine triphosphatase (GTPase) activity. Compared to wild-type (WT) Gpa1p, Gpa1(G50V)p was found to have a moderately reduced GTPase activity and increased GTP occupancy, while GTPgammaS binding and GDP exchange were not significantly altered. The yeast regulator of G protein Signalling (RGS) protein, Sst2p, was also expressed and purified, and found to have a significantly reduced ability to stimulate the initial rate of GTP hydrolysis of Gpa1(G50V)p compared to its effect on WT Gpa1p. Probing conformational transitions by a protease sensitivity assay suggested that Gpa1(G50V)p did not bind the transition state mimetic GDP/AlF(4)(-) as efficiently as the WT Gpa1p. These biochemical results can explain many of the known gpa1(G50V) yeast cell phenotypes. PMID- 10705369 TI - Identification of a putative DEAD-box RNA helicase and a zinc-finger protein in Candida albicans by functional complementation of the S. cerevisiae rok1 mutation. AB - We identified two novel genes, CHR1 and CSR1, of the fungal pathogen Candida albicans, by functional complementation of the Saccharomyces cerevisiae rok1 mutation. The Rok1 protein is a member of the DEAD protein family of ATP dependent RNA helicases. ROK1 is required for cell cycle progression and also for rRNA processing. The CHR1 gene product of 578 amino acids is highly homologous to the Rok1 protein (54% identity) and is considered to be a putative DEAD-box RNA helicase. We predict that the CSR1 gene encodes a 73 kDa protein of 612 amino acids with five zinc-finger motifs at the C-terminal region. CHR1 or CSR1 on a high-copy number plasmid showed a slow-growth phenotype in a condition where the ROK1 expression is turned on from the GAL1 promoter. This result is consistent with the lethality caused by the ROK1 overexpression. We conclude that CHR1 encodes a functional homologue of Rok1 protein and CSR1 is a heterologous suppressor of the rok1 mutation. PMID- 10705370 TI - High-frequency occurrence of chromosome translocation in a mutant strain of Candida albicans by a suppressor mutation of ploidy shift. AB - Significant occurrence of high-ploidy cells is commonly observed among many Candida albicans strains. We isolated two isogenic strains, STN21 and STN22, each from a half sector of a colony obtained after mild UV-irradiation of a Arg(-) derivative of CBS5736. The two strains were different from each other in ploidy states and chromosome organization. Although cells of STN22 were homogeneous in size and had a single nucleus, high-ploidy cells, with either a single large nucleus or several nuclei, were present together with apparently normal cells with a single nucleus in the cell population of STN21. Flow cytometry showed that STN22 was a stable diploid; however, STN21 seemed to be the mixture of different ploidy states, including diploid and tetraploid. The phenotype of STN21 containing high-ploidy cells is referred to here as the Sps(-) phenotype (suppressor of ploidy shift). STN22 showed a typical electrophoretic karyotype similar to strain 1006 in C. albicans. However, an extra chromosomal band appeared in some clones of STN21 at high frequency. By assignment of several DNA probes, this extra chromosome was shown to be a translocation of the 7F-7G portion of chromosome 7 with the 470 kb DNA segment containing H SfiI fragment from chromosome 4. Thus, this extra chromosome is a hybrid of 4H and 7F-7G. Since the isogenic Sps(+) strain STN22 exhibited no extra chromosome bands, a correlation is suggested between the Sps(-) phenotype and the occurrence of chromosome translocations. PMID- 10705371 TI - A mathematical model for yeast respiro-fermentative physiology. AB - A mechanistic model is presented that describes the respiro-fermentative physiology of yeast. The model assumes the presence of multiple types of glucose carriers and multiple assimilation pathways. Respiro-fermentative physiology is explained by the mechanistic response of the different types of carriers and assimilation pathways on the substrate concentration. At low substrate concentrations, glucose is taken up mainly via a high affinity carrier with a low maximum uptake rate. At high substrate concentrations, this carrier becomes saturated and the main pathway for glucose uptake is via a low affinity carrier with a high maximum uptake rate. The price to pay for the high uptake rate is a lowered assimilation efficiency, resulting in a low biomass yield. Product formation occurs via the pathway with the high uptake rate. The model explains the link between substrate concentration and product formation generally observed in the literature on yeast and bacteria. Model parameter values are estimated by fitting data from the literature. The model distinguishes itself from other models in that it does not rely on the presence of switches, such as the 'critical dilution rate', or on the assumption that the respiratory capacity reaches its maximum during respiro-fermentative metabolism. The present theory is not designed exclusively for the phenomenon of respiro-fermentative physiology: it describes the degradation of substances by heterotrophic micro-organisms in general. PMID- 10705372 TI - YHP1 encodes a new homeoprotein that binds to the IME1 promoter in Saccharomyces cerevisiae. AB - The IME1 gene is essential for initiation of meiosis in the yeast Saccharomyces cerevisiae. Transcription of IME1 is detected under conditions of starvation for nitrogen and glucose, and in the presence of the MATa1 and MATalpha2 gene products. In our previous work, we have shown that there are two elements acting as TUP1-dependent upstream repression sequence (URS) and tup1 mutation-dependent upstream activation sequence (UAS) between nt -915 and -621 of the IME1 promoter under nutritional conditions. The region from -915 to -621 has also been reported to harbour meiotic URS and UAS when a/alpha cells were transferred to sporulation conditions. To identify proteins that are able to bind to the region, we screened a cDNA library fused with the Gal4-activation domain by means of the one-hybrid system. We identified a previously unknown gene (YDR451c), which we designated YHP1, encoding a homeodomain protein of the Drosophila antennapedia type. The region for binding of Yhp1 was delimited to the 28 bp region between nt -702 and 675 of the IME1 promoter in vivo and in vitro, and the 28 bp region harboured a URS activity in a Yhp1-dependent manner under nutrient growth conditions. Although a yhp1 single-disruption mutation did not give rise to a scorable phenotype under nutritional and sporulation conditions, the level of the YHP1 transcript was significantly lower in the cells grown in acetate medium (presporulation medium) and sporulation medium than those grown in glucose medium, and the reduction of YHP1 transcription in acetate medium coincided with an increment of the IME1 transcript. We suggest that the homeoprotein Yhp1 that binds directly to the 28 bp region of the IME1 promoter is a new repressor acting under glucose growth conditions. PMID- 10705373 TI - Molecular cloning and characterization of a glucan synthase gene from the human pathogenic fungus Paracoccidioides brasiliensis. AB - 1,3-beta-D-glucan is a fungal cell wall polymer synthesized by the multi-subunit enzyme 1,3-beta-D-glucan synthase. A subunit of this integral membrane protein was first described as the product of the FKS1 gene from Saccharomyces cerevisiae using echinocandin mutants. Other FKS1 genes were also reported for Candida albicans, Aspergillus nidulans and Cryptococcus neoformans. Here, we report the nucleotide sequence of the first homologous FKS gene cloned from the pathogenic fungus Paracoccidioides brasiliensis. An open reading frame of 5942 bp was identified in the complete sequence, interrupted by two putative introns, the first close to the 5' end and the second close to the 3' end of the gene. A promoter region is also described containing consensus sequences such as canonical TATA and CAAT boxes and, possibly, multiple sites for glucose regulation by creA protein. The deduced sequence of 1926 amino acid show more than 85% similarity to FksAp from A. nidulans, and 71% to Fks1p and Fks2p from S. cerevisiae. Computational analysis of P. brasiliensis Fks1p suggests a similar structure to transmembrane proteins, such as FksAp, with the presence of two domains composed by hydrophobic helices that limit the putative highly hydrophilic catalytic domain within the cytoplasm. PMID- 10705374 TI - Anaerobic and aerobic batch cultivations of Saccharomyces cerevisiae mutants impaired in glycerol synthesis. AB - Glycerol is formed as a by-product in production of ethanol and baker's yeast during fermentation of Saccharomyces cerevisiae under anaerobic and aerobic growth conditions, respectively. One physiological role of glycerol formation by yeast is to reoxidize NADH, formed in synthesis of biomass and secondary fermentation products, to NAD(+). The objective of this study was to evaluate whether introduction of a new pathway for reoxidation of NADH, in a yeast strain where glycerol synthesis had been impaired, would result in elimination of glycerol production and lead to increased yields of ethanol and biomass under anaerobic and aerobic growth conditions, respectively. This was done by deletion of GPD1 and GPD2, encoding two isoenzymes of glycerol 3-phosphate dehydrogenase, and expression of a cytoplasmic transhydrogenase from Azotobacter vinelandii, encoded by cth. In anaerobic batch fermentations of strain TN5 (gpd2-Delta1), formation of glycerol was significantly impaired, which resulted in reduction of the maximum specific growth rate from 0.41/h in the wild-type to 0.08/h. Deletion of GPD2 also resulted in a reduced biomass yield, but did not affect formation of the remaining products. The modest effect of the GPD1 deletion under anaerobic conditions on the maximum specific growth rate and product yields clearly showed that Gdh2p is the important factor in glycerol formation during anaerobic growth. Strain TN6 (gpd1-Delta1 gpd2-Delta1) was unable to grow under anaerobic conditions due to the inability of the strain to reoxidize NADH to NAD(+) by synthesis of glycerol. Also, strain TN23 (gpd1-Delta1 gpd2-Delta1 YEp24-PGKp-cth PGKt) was unable to grow anaerobically, leading to the conclusion that the NAD(+) pool became limiting in biomass synthesis before the nucleotide levels favoured a transhydrogenase reaction that could convert NADH and NADP(+) to NADPH and NAD(+). Deletion of either GPD1 or GPD2 in the wild-type resulted in a dramatic reduction of the glycerol yields in the aerobic batch cultivations of strains TN4 (gpd1-Delta1) and TN5 (gpd2-Delta1) without serious effects on the maximum specific growth rates or the biomass yields. Deletion of both GPD1 and GPD2 in strain TN6 (gpd1-Delta1 gpd2-Delta1) resulted in a dramatic reduction in the maximum specific growth rate and in biomass formation. Expression of the cytoplasmic transhydrogenase in the double mutant, resulting in TN23, gave a further decrease in micromax from 0.17/h in strain TN6 to 0.09/h in strain TN23, since the transhydrogenase reaction was in the direction from NADPH and NADP(+) to NADH and NADP(+). Thus, it was not possible to introduce an alternative pathway for reoxidation of NADH in the cytoplasm by expression of the transhydrogenase from A. vinelandii in a S. cerevisiae strain with a double deletion in GPD1 and GPD2. PMID- 10705375 TI - Current awareness on yeast [bibliography]. PMID- 10705376 TI - Midgestation lethality in mice deficient for the RecA-related gene, Rad51d/Rad51l3. AB - Homologous recombination (HR) occurs in all organisms, and is important for repair of DNA damage, chromosome segregation during meiosis, and genetic diversification. Genes critical for recombinational DNA repair and meiotic recombination include members of the RecA/RAD51 family, of which seven have been identified in mammals. Here, we describe the disruption of Rad51d (recently designated Rad51l3) in mice and its phenotypic consequences. Rad51d-deficient mice die between 8.5 and 11.5 dpc. The affected embryos are smaller than littermates, posteriorly truncated, and developmentally delayed. Embryonic fibroblasts from mutant embryos could not be propagated more than one generation in culture. Rad51d-deficient blastocysts were not sensitive to gamma radiation or methylmethanesulfonate (MMS) in blastocyst outgrowth experiments. The variable and generalized developmental progression defects in Rad51d-deficient embryos suggests that mutant cells may undergo delayed or suboptimal repair of DNA damage, resulting in accumulated degrees of mutation and/or cell cycle perturbation that are incompatible with normal embryonic development. genesis 26:167-173, 2000. PMID- 10705377 TI - Temperature-dependent expression of turtle Dmrt1 prior to sexual differentiation. AB - Vertebrates employ varied strategies, both chromosomal and nonchromosomal, to determine the sex of the developing embryo. Among reptiles, temperature-dependent sex determination (TSD) is common. The temperature of incubation during a critical period preceding sexual differentiation determines the future sex of the embryo, presumably by altering the activity or expression of a temperature dependent regulatory factor(s). Here we examine the expression of the Dmrt1 gene, a candidate regulator of mammalian and avian sexual development, in the turtle. During the sex-determining period, Dmrt1 mRNA is more abundant in genital ridge/mesonephros complexes at male-promoting than at female-promoting temperatures. Dmrt1 is the first gene found to show temperature-dependent expression prior to sexual differentiation, and may play a key role in sexual development in reptiles. genesis 26:174-178, 2000. PMID- 10705378 TI - A development-specific histone H3 localizes to the developing macronucleus of Euplotes. AB - During the process of macronuclear development, the ciliate Euplotes crassus undergoes extensive programmed DNA rearrangement. Previous studies have identified a gene, H3(P), that is expressed only during sexual reproduction and is predicted to encode a variant histone H3 protein. In the current study, an antiserum to the H3(P) protein has been generated. The antiserum has been used to demonstrate that H3(P) is maximally expressed during the polytene chromosome stage of macronuclear development. Moreover, H3(P) is localized to the developing macronucleus, but not other nuclei present within the cell. Additional studies indicate that at least one additional variant histone is also present within the developing macronucleus. The results indicate that there are significant changes in nucleosome composition within the developing macronucleus, and provide additional support for the notion that changes in chromatin structure play a role in the DNA rearrangement processes of macronuclear development. genesis 26:179 188, 2000. PMID- 10705379 TI - Pc-G/trx-G and the SWI/SNF connection: developmental gene regulation through chromatin remodeling. AB - Pc-G and trx-G genes are responsible for maintenance of transcriptional regulation and provide a cellular memory mechanism throughout development. Studies in fly, yeast, mouse, and human have implicated modulation of higher order chromatin structure as an important component in this process. Specifically, connections between SWI/SNF complexes and trx-G genes have provided a mechanistic link between chromatin remodeling and transcriptional regulation. Here we discuss recent genetic and biochemical data that has shed light on the molecular mechanisms and pathways associated with Pc-G and trx-G function in developmental processes such as cell cycle control and hematopoiesis. genesis 26:189-197, 2000. PMID- 10705380 TI - A novel family of retrotransposons in Xenopus with a developmentally regulated expression. AB - SUMMARY: We have obtained a novel family of LTR-retrotransposons in Xenopus laevis, named Xretpos, from cDNA and genomic clones. Its long terminal repeats (LTRs) can be subdivided into U3, R, and U3 to U5 region, and are bounded by 6 bp inverted repeats. Xretpos contains primer binding site and polypurine tract, and multiple copies of Xretpos-related element are present in the genome. A long open reading frame (ORF) encodes the CCHC motif conserved in retroviral gag proteins and leucine zipper motif capable of forming the coiled-coil. However, no amino acid homology to usually conserved retroviral pol gene was revealed. We report that in vitro synthesized Xretpos complementary RNAs are translated to produce a predicted size of protein. We also show that zygotically activated Xretpos transcripts are restricted to ventro-posterior specific regions and induced by UV irradiation and BMP-4 overexpression in cycloheximide-dependent way. genesis 26:198-207, 2000. PMID- 10705381 TI - The balbiani body: asymmetry in the mammalian oocyte. PMID- 10705382 TI - Differential expression of VEGF isoforms and VEGF(164)-specific receptor neuropilin-1 in the mouse uterus suggests a role for VEGF(164) in vascular permeability and angiogenesis during implantation. AB - The mechanism(s) by which localized vascular permeability and angiogenesis occur at the sites of implantation is not clearly understood. Vascular endothelial growth factor (VEGF) is a key regulator of vasculogenesis during embryogenesis and angiogenesis in adult tissues. VEGF is also a vascular permeability factor. VEGF acts via two tyrosine kinase family receptors: VEGFR1 (Flt-1) and VEGFR2 (KDR/Flk-1). Recent evidence suggests that neuropilin-1 (NRP1), a receptor involved in neuronal cell guidance, is expressed in endothelial cells, binds to VEGF(165) and enhances the binding of VEGF(165) to VEGFR2. We examined the spatiotemporal expression of vegf isoforms, nrp1 and vegfr2 as well as their interactions in the periimplantation mouse uterus. We observed that vegf(164) is the predominant isoform in the mouse uterus. vegf(164) mRNA accumulation primarily occurred in epithelial cells on days 1 and 2 of pregnancy. On days 3 and 4, the subepithelial stroma in addition to epithelial cells exhibited accumulation of this mRNA. After the initial attachment reaction on day 5, luminal epithelial and stromal cells immediately surrounding the blastocyst exhibited distinct accumulation of vegf(164) mRNA. On days 6-8, the accumulation of this mRNA occurred in both mesometrial and antimesometrial decidual cells. These results suggest that VEGF(164) is available in mediating vascular changes and angiogenesis in the uterus during implantation and decidualization. This is consistent with coordinate expression of vegfr2, and nrp1, a VEGF(164)-specific receptor, in uterine endothelial cells. Their expression was low during the first 2 days of pregnancy followed by increases thereafter. With the initiation and progression of implantation (days 5-8), these genes were distinctly expressed in endothelial cells of the decidualizing stroma. Expression was more intense on days 6-8 at the mesometrial pole, the presumptive site of heightened angiogenesis and placentation. However, the expression was absent in the avascular primary decidual zone immediately surrounding the implanting embryo. Crosslinking experiments showed that (125)I-VEGF(165) binds to both NRP1 and VEGFR2 present in decidual endothelial cells. These results suggest that VEGF(164), NRP1 and VEGFR2 play a role in VEGF-induced vascular permeability and angiogenesis in the uterus required for implantation. genesis 26:213-224, 2000. PMID- 10705383 TI - The legacy of Karl Rokitansky. PMID- 10705385 TI - Comparative pathology: human disease counterparts in marine animals. AB - Marine animals offer unparalleled diversity in form, behavior, environments occupied, and other attributes. Furthermore, certain of these animals have evolutionary links to humans. These features make marine animals of special interest to comparative and human pathologists. This article focuses on comparative aspects of 4 major disorders, namely, tumors, immunologic disorders, vascular pathology, and anatomic malformations. PMID- 10705386 TI - The role of Clostridium septicum in paraneoplastic sepsis. AB - CONTEXT: Clostridium septicum infections are rare but often associated with serious if not fatal outcomes. Clostridium septicum infection does not appear to be associated with a single specific defect in cellular or humoral immunity. It has been associated with multiple medical problems, including but not limited to leukemia, malignancy of the bowel, other solid tumors, cyclic neutropenia with enterocolitis, diabetes mellitus, and severe arteriosclerosis. Most cases of C septicum are associated with malignancy, and mortality approaches 100% if care is not rendered within 12 to 24 hours. OBJECTIVES: To evaluate outcomes of patients with C septicum bacteremia, whether treated medically or surgically or both, and to note associated conditions. DESIGN: Retrospective evaluation of patients found to have C septicum bacteremia in the past 6 years. SETTING: Two teaching hospitals, Brooke Army Medical Center (250 beds) and Wilford Hall Medical Center (292 beds), were the source of our patients. PATIENTS: All patients found to have C septicum bacteremia during hospitalization or postmortem examination were included in the study. There were no exclusion criteria. MAIN OUTCOME MEASURE: Mortality associated with C septicum infection. RESULTS: In our case series, mortality was 33%, which is slightly lower than reported in prior studies (43% 70%). CONCLUSION: Presumptive identification based on Gram stain, awareness of C septicum infection as a paraneoplastic syndrome, and prompt, clear communication between laboratory personnel and clinicians are necessary for early diagnosis of C septicum infection. Early institution of antibiotic therapy improves prognosis. PMID- 10705387 TI - Effects of restructuring on the performance of microbiology laboratories in Alberta. AB - OBJECTIVE: To evaluate the error rates of organism identification and antibiotic susceptibility proficiency testing challenges before, during, and after microbiology laboratory restructuring in Alberta. METHODS: Alberta Health substantially reduced and redistributed laboratory funds to the regional health authorities in 1995, forcing a dramatic restructure of services. Many rural hospitals expanded their microbiology test menus, and urban centers consolidated microbiology testing into a centralized high-volume laboratory. The Laboratory Proficiency Testing Program of the College of Physicians and Surgeons of Alberta mailed regular test profile surveys to microbiology laboratories during the restructure period to determine the type and extent of changes in services. Based on the types of tests and the extent of analysis being done, most rural B-level and some C-level laboratories were reclassified to the A level. The Laboratory Proficiency Testing Program reviewed the error rates of proficiency challenges based on the performance of different levels of laboratories before and after the period of restructure. RESULTS: Overall performance has improved according to the number of errors documented on identification and susceptibility challenges for laboratories that remained at the same classification (ie, A or C). The number of major identification errors for laboratories that were reclassified increased, but the rate of major susceptibility errors decreased. More reclassified laboratories do not have dedicated registered technologist(s) who perform microbiology testing and are not supervised by an on-site pathologist and/or medical microbiologist compared with laboratories that remained at the same classification. CONCLUSIONS: Microbiology laboratory restructuring will have adverse effects on the quality of complex testing if experienced technologists are not retained and services are not medically supervised. PMID- 10705388 TI - A mouse model of aerosol-transmitted orthopoxviral disease: morphology of experimental aerosol-transmitted orthopoxviral disease in a cowpox virus-BALB/c mouse system. AB - OBJECTIVES: To determine the morphologic changes and disease progression of aerosolized cowpox virus infection in BALB/c mice and to ascertain the suitability of cowpox virus-infected BALB/c mice as a model of aerosol transmitted, orthopoxviral respiratory disease. METHODS: BALB/c mice were inoculated with cowpox virus, Brighton strain, by aerosol or intranasal route. Mice were killed at specified times after inoculation, necropsied, and tissues were collected for routine histology, immunohistochemistry, and electron microscopy. RESULTS: Inoculation by both routes resulted in disease and death. Immunolabeled viral antigen and lesions predominated in the tissues associated with the inoculation route, that is, lungs, airways, trachea, and nasal passages and sinuses. Tracheitis was evident in the intranasally infected group only. Lesions were generally necrotizing and hemorrhagic, neutrophilic, and increased in extent and severity in a time-dependent fashion. Viral intracytoplasmic inclusion bodies, immunolabeled viral antigen, or virions were readily seen in epithelial tissues, smooth muscle cells of airways and vessels, fibroblasts, periosteal cells, perineural cells, and macrophages. Although the extension of infection appeared to be primarily direct, lesions suggesting hematogenous dissemination were occasionally noted in bone marrow and skin. Transmission electron microscopy demonstrated features of cell injury or death, virion assembly and maturation, and both A-type and B-type inclusions. CONCLUSIONS: Aerosol inoculation of BALB/c mice with cowpox virus provides a reliable and facilitative model of aerosol-transmitted, orthopoxviral respiratory disease. PMID- 10705389 TI - Prophylactic mastectomy: pathologic findings in high-risk patients. AB - BACKGROUND: According to recently published data, prophylactic mastectomy (PM) appears to prevent about 90% of the expected malignant neoplasms in women with a family history of breast cancer. OBJECTIVES: To identify the frequency of high risk lesions in PM specimens and to determine occurrence of any new primary breast cancer following PM. DESIGN: We performed a retrospective study of women undergoing unilateral or bilateral PM. Medical charts and pathologic findings of 35 patients who underwent bilateral mastectomies at University Hospital, Syracuse, NY, from 1989 to 1996 were reviewed. Patients with biopsy-proven bilateral breast cancer were excluded. Patients were divided into 3 groups: (A) positive family history and no known breast cancer (n = 9), (B) positive family history and contralateral neoplasia (n = 13), and (C) negative family history and contralateral neoplasia (n = 13). These findings were compared with those found in reduction mammoplasty specimens from 10 women at standard risk of breast cancer. RESULTS: The mean age of the control group of women undergoing reduction mammoplasty was 38 years. The pathologic specimens demonstrated no significant pathologic findings in 9 and fibrocystic change in 1. In group A, the mean number of affected relatives was 3.1, and the mean age was 38 years. Two of these 9 women had atypical duct hyperplasia and 1 had atypical lobular hyperplasia in their breasts (ie, 33% with high-risk pathologic findings). Of the 13 group B women (mean age, 46.6 years; mean of 2.5 affected relatives and unilateral breast cancer), the contralateral PM specimen contained duct carcinoma in situ in one and invasive ductal cancer in a second (15% with occult malignant neoplasms). In 13 group C patients (mean age, 47.1 years), 3 (23.1%) of the contralateral PM specimens displayed atypical duct hyperplasia or atypical lobular hyperplasia. At a mean follow-up of 4.8 years, there have been no new breast malignant neoplasms in these 45 women. CONCLUSIONS: The occurrence of unilateral cancer in patients with family history of breast cancer is associated with a 15.4% probability of simultaneous occult malignant neoplasms in the contralateral breast. Patients with a strong family history but no evidence of breast cancer have a substantially similar rate of proliferative disease in their PM specimens as those women who have unilateral cancer but no significant family history. PMID- 10705390 TI - A unique method for mutation analysis of tumor suppressor genes in colorectal carcinomas using a crypt isolation technique. AB - BACKGROUND: Contamination of nontumor tissue makes genetic analysis difficult. For this reason, it is important to obtain pure tumor tissue to ensure accurate genetic analysis. OBJECTIVE: To accurately assess the incidence of mutation of tumor suppressor genes (p53: exon 5-8; APC: mutated cluster region; NF-2 gene: all exons) in 45 colorectal carcinomas. METHODS: We developed an application of the polymerase chain reaction-single-strand conformation polymorphism and DNA sequence by coupling them with crypt isolation. RESULTS: Mutations of p53 and APC genes were found in 24 and 22 of 45 colorectal carcinomas, respectively. No mutation of the NF-2 gene was observed in this cancer. Single-strand conformation polymorphism using a crypt isolation technique showed a clear migrating band and no false-positive data. CONCLUSIONS: The crypt isolation technique is a useful method for accurately analyzing genetic alterations. Furthermore, our proposed method confirmed the morphological findings obtained before the genetic analysis. PMID- 10705391 TI - Analysis of false-negative diagnoses on endoscopic brush cytology of biliary and pancreatic duct strictures: the experience at 2 university hospitals. AB - CONTEXT: Endoscopic brush cytology is a valuable technique for the diagnosis of pancreatobiliary malignancy. Despite its widespread use, the sensitivity of this method has been reported as approximately 50%. The specificity is usually higher than 95%. Few reports have systematically analyzed the reasons for this relatively low sensitivity. OBJECTIVES: To determine the rate and reasons for false-negative diagnoses in endoscopic brushing cytology of biliary and pancreatic ducts based on the results of sensitivity, specificity, accuracy, and positive and negative predictive values. DESIGN: Retrospective analysis of laboratory data and slide review of false-negative cases. SETTING: Two tertiary care state university hospitals. PATIENTS: A total of 183 pancreatobiliary brushing specimens obtained from patients undergoing endoscopic retrograde cholangiopancreatography for biliary or pancreatic duct disease for a 4- to 5 year period. INTERVENTION: Endoscopic retrograde cholangiopancreatography brushings. MAIN OUTCOME MEASURES: Determination of sensitivity, specificity, accuracy, and positive and negative predictive values. Analysis of false-negative results. RESULTS: The sensitivity, specificity, accuracy, and positive and negative predictive values, overall, were 48%, 98%, 79%, 92%, and 76%, respectively. Sampling error was a major cause of false-negative diagnoses (67%), followed by interpretive (17%) and technical errors (17%). CONCLUSIONS: Improvements in sensitivity and diagnostic accuracy for cancer of the pancreatobiliary tract can be achieved by optimizing slide preparatory techniques. Also, enhancement of the cytologist's diagnostic skills enables the identification of the morphologic features of premalignant lesions. Repeat brushings are indicated for suspicious or negative results not consistent with the clinical or radiologic findings. PMID- 10705392 TI - Expression of telomerase activity and telomerase RNA in human soft tissue sarcomas. AB - OBJECTIVE: To investigate whether telomerase is reactivated in soft tissue tumor and whether telomerase activity is regulated at the transcriptional level. DESIGN: Fresh tissue samples of 24 soft tissue sarcomas were analyzed for telomerase activity by a radioactive polymerase chain reaction-based telomeric repeat amplification protocol assay and for human telomerase RNA (hTR) by an in situ hybridization assay. SETTING: Tertiary care teaching hospital. PATIENTS: Twenty-four patients with soft tissue tumor were surgically treated. Twelve patients had malignant fibrous histiocytoma, 5 had liposarcoma, 6 had leiomyosarcoma, and 1 had rhabdomyosarcoma. RESULTS: Telomerase activity was detected in 4 sarcoma samples (17%), all of which were positive for hTR. Expression of hTR was demonstrated in 13 sarcomas (54%), 4 of which were positive for telomerase and 9 of which were negative for telomerase. One (50%) of 2 grade 1 tumors, 9 (50%) of 18 grade 2 tumors, and 3 (75%) of 4 grade 3 tumors showed hTR expression. CONCLUSIONS: The relatively low frequency of telomerase activity in soft tissue sarcomas suggests that telomerase may not play an important role in tumorigenesis in these tumors. Telomerase ladders were demonstrated only in association with tumors expressing hTR. It is noteworthy that half of the patients with grade 1 and 2 tumors expressed hTR, suggesting that telomerase RNA may be useful as a marker for identifying tumor aggressiveness earlier than the conventional histopathologic grading scale. PMID- 10705393 TI - Clinical usefulness of telomerase assay for the detection of lymph node metastasis in patients with oral malignancy. AB - OBJECTIVE: Telomerase is considered a diagnostic marker of malignancy. We investigated the usefulness of telomerase assay for the detection of lymph node micrometastasis. METHODS: Sixteen cervical lymph nodes with metastasis of oral cancer and 20 benign lymph nodes were studied. The oral cancer cell line was used to estimate the sensitivity for telomerase assay. Telomerase activity was measured by semiquantitative telomeric repeat amplification protocol. RESULTS: There was a significant difference between malignant and benign lymph nodes. The telomerase activity of 50 mg of lymph nodes with 103 or more cancer cells differed from that of control lymph nodes. Lymph nodes with 102 or fewer tumor cells expressed similar levels as benign lymph nodes. CONCLUSIONS: In addition to routine histologic examination, telomerase assay is considered a useful tool for the detection of lymph node metastasis in patients with oral malignancy. PMID- 10705394 TI - Salivary gland basal cell and canalicular adenomas: immunohistochemical demonstration of myoepithelial cell participation and morphogenetic considerations. AB - OBJECTIVE: To evaluate cellular composition of salivary gland adenomas using 3 monoclonal antibodies that recognize a smooth muscle phenotype confirmed to be sensitive for myoepithelial differentiation. DESIGN: Immunohistochemical evaluation of 25 salivary gland basal cell and canalicular adenomas. SETTING: Archival pathology material from the files of Henry Ford Hospital, Detroit, Mich, and the University of California at San Francisco. RESULTS: All basal cell adenoma variants exhibit some degree of myoepithelial cell participation with periductal, epithelioid, and spindled (stromal-like) morphologic structures. Only the canalicular adenomas, even if mixed with trabecular and solid patterns, are devoid of staining with these 3 antibodies, suggesting an adenoma composed exclusively of ductal luminal cells. CONCLUSIONS: There is an overlapping histomorphologic and common cellular composition of the basal cell adenoma variants with other recognized adenomas, such as pleomorphic adenoma and myoepithelioma. Relative differentiation toward 3 cell phenotypes (ductal luminal, basal, and myoepithelial) and the character of extracellular matrix production in varying proportions by the neoplastic myoepithelial cells distinguishes the spectrum of salivary gland adenomas identified in current classification schemes. PMID- 10705395 TI - Angiectatic nasal polyps that clinically simulate a malignant process: report of 2 cases and review of the literature. AB - BACKGROUND: Approximately 5% of inflammatory or allergic sinonasal polyps develop extensive vascular proliferation and ectasia with deposition of pseudoamyloid. These so-called angiectatic nasal polyps (ANPs) can grow rapidly and exhibit an aggressive clinical behavior that could simulate malignancy preoperatively. OBJECTIVE: To systematically address the differential histologic diagnosis of ANPs. METHODS: We evaluated by light microscopy, immunohistochemistry, and electron microscopy biopsy and resection specimens from 2 large ANPs (8 and 10 cm in diameter) that presented in 2 adult men with life-threatening epistaxis and facial deformity, respectively. RESULTS: The tumors were firm, lobulated, and covered by smooth, partially ulcerated mucosa. Histologically, clusters of dilated, thin-walled blood vessels embedded in pools of Congo red-negative eosinophilic material, associated with patchy necrosis and atypical stromal spindle cells, were seen. Electron microscopy and immunohistochemistry (CD34, factor VIII) confirmed the endothelial nature of the cells lining the spaces, whereas the atypical stromal cells were classified as myofibroblasts. CONCLUSIONS: These 2 cases represent extreme examples of ANPs that clinically simulate a malignant process. Awareness of the histological features of ANPs should prevent confusion of such lesions with other vascular or spindle cell lesions of the nasopharynx that would require different treatment and carry a different prognosis. PMID- 10705396 TI - Esophageal collision tumor (Large cell neuroendocrine carcinoma and papillary carcinoma) arising in a Barrett esophagus. AB - We report herein a unique case of an esophageal collision tumor composed of a papillary adenocarcinoma and a large cell neuroendocrine carcinoma arising in a Barrett esophagus. Hematoxylin-eosin and silver staining patterns, immunohistochemistry, and electron microscopy of the large cell neuroendocrine component are discussed. PMID- 10705397 TI - The cytomorphology of pleuropulmonary blastoma. AB - Pleuropulmonary blastoma is a rare, primitive primary neoplasm of the thorax in young children. The tumor, which is often but not always associated with cystic lung lesions, may arise in pulmonary parenchyma, the mediastinum, and pleura. Histologically, it is characterized by a biphasic neoplastic population of undifferentiated-appearing small round cells and larger spindle-shaped cells. A proportion of these cancers may also manifest more specific mesenchymal differentiation. In contrast to the pulmonary blastoma of adults, a malignant epithelial component does not occur. We present herein the third known case of a fine needle aspiration biopsy of a pleuropulmonary blastoma in a 5-year-old girl. The smears were moderately cellular and included an admixture of the characteristic small ovoid blastemal elements and scattered spindled mesenchymal tumor cells. PMID- 10705398 TI - Primary malignant melanoma of the common bile duct: a case report and review of the literature. AB - Primary malignant melanoma of the common bile duct is rare. To our knowledge, only 6 cases have been reported previously. The pathologic diagnosis of primary malignant melanoma in extracutaneous sites often requires the use of confirmatory immunohistochemical stains and electron microscopy studies, as well as tests to rule out other possible remote or concurrent primary sites. The presence of junctional activity adjacent to the tumor is another important requisite for the diagnosis of this entity. Nevertheless, absolute exclusion of a metastatic melanoma from an unknown occult site or regressed site is not entirely possible. We describe our observations in a case of primary malignant melanoma of the common bile duct in a 48-year-old man and discuss the criteria for diagnosis of primary melanoma. PMID- 10705399 TI - Pleomorphic hyalinizing angiectatic tumor of soft parts: immunohistochemical study including the expression of vascular endothelial growth factor. AB - We report morphologic, flow cytometric, and immunohistochemical findings in two cases of pleomorphic hyalinizing angiectatic tumor of soft parts. Both patients were middle-aged women with subcutaneous lesions located in the lower extremity. The tumors consisted of sheets of spindled and pleomorphic cells with frequent intranuclear pseudoinclusions associated with clusters of ectatic vessels surrounded by prominent perivascular hyaline material. Numerous, nonhyalinized vessels were also present, mostly in the peripheral areas of the lesions. Some of these vessels had their walls permeated by numerous small capillaries. Immunostaining for vascular endothelial growth factor (VEGF), a secreted protein that has been implicated in tumor-associated angiogenesis, demonstrated positive staining in both tumoral and endothelial cells. Tumor cells were also reactive to vimentin and CD34. Focal positivity for CD99 and factor XIIIa was also present. Flow cytometry yielded a diploid DNA histogram with S-phase fraction of 7%. Our findings corroborate those from previously reported cases. They further suggest that angiogenesis and the angiogenic factor VEGF may play a role in the development of this peculiar tumor. PMID- 10705400 TI - Pagetoid bowen disease: a report of 2 cases that express cytokeratin 7. AB - Bowen disease is a variant of squamous cell carcinoma in situ. In some cases a pagetoid growth pattern can be observed with cytologically atypical clear cells arranged singly and in nests. The differential diagnosis of pagetoid Bowen disease includes primarily Paget disease and malignant melanoma in situ, as well as other less common entities. Two cases of pagetoid Bowen disease are described, one in a 65-year-old man with a thigh lesion and the other in a 25-year-old man with a lesion in the penile/scrotal region. Neither patient had clinical evidence of an internal malignant neoplasm. In both cases, the neoplastic cells were positive for cytokeratin (CK) 7 and CK 19 and were negative for CK 18, CK 20, carcinoembryonic antigen, GCDFP-15, c-erbB2, S100, and HMB-45. In aggregate, these findings support the diagnosis of pagetoid Bowen disease. Previously, others have shown that CK 7 is an almost invariable marker of Paget disease. Thus, we report these two cases to illustrate that CK 7 can be expressed by pagetoid Bowen disease and should not be a cause of confusion in the differential diagnosis. PMID- 10705401 TI - Metastatic female adnexal tumor of probable Wolffian origin: a case report and review of the literature. AB - Female adnexal tumor of probable wolffian origin is a rare neoplasm that can present diagnostic difficulties. We report herein a case of a 60-year-old woman with female adnexal tumor of probable wolffian origin arising within the leaves of a broad ligament and, 5 years later, presenting with metastasis to the liver. The morphologic, immunohistochemical, ultrastructural, and DNA ploidy findings of the original and metastatic tumor, differential diagnoses, and the results of the English-language literature review are presented. PMID- 10705402 TI - Calcifying fibrous pseudotumor of the neck. AB - Calcifying fibrous pseudotumor is a rare lesion of uncertain histogenesis that has a unique histologic appearance. We report herein a case of a 24-year-old woman with a mass on the right posterior side of the neck. Magnetic resonance imaging with contrast showed a well-circumscribed mass between the right splenius and semispinalis cervicis muscles; the study suggested high collagen content. Simple excision was performed. The histologic findings were diagnostic of calcifying fibrous pseudotumor. Our review of 19 previously reported cases suggests that a good outcome is expected when a diagnosis of calcifying fibrous pseudotumor is made. PMID- 10705403 TI - Epithelioid leiomyosarcoma with osteoclast-like giant cells in the rectum. AB - We report a rare case of rectal epithelioid leiomyosarcoma with osteoclast-like giant cells. A 71-year-old Japanese man was admitted to a hospital with melena. Results of a colonoscopy test revealed a polypoid tumor in the rectum, and a biopsy specimen from the lesion showed a sarcoma; the patient underwent rectosigmoidectomy. At gross inspection, the tumor measured 8 x 7 x 4 cm and was polypoid with ulcerations. Necrotic and hemorrhagic foci were scattered. Microscopically, the tumor consisted of 2 cell types: malignant tumor cells with epithelioid features and benign-appearing osteoclast-like giant cells. The tumor cells were polygonal and epithelioid in shape and had eosinophilic or clear cytoplasms, with scattered giant tumor cells. Immunohistochemical examination revealed that the tumor cells were positive for vimentin, muscle actin, alpha smooth muscle actin, and desmin, whereas the osteoclast-like giant cells were positive for CD68, leukocyte common antigen, and lysozymes. We diagnosed this case as epithelioid leiomyosarcoma with osteoclast-like giant cells. To the best of our knowledge, this is the first case of rectal epithelioid leiomyosarcoma with osteoclast-like giant cells. PMID- 10705405 TI - Thyroid sclerosing mucoepidermoid carcinoma with eosinophilia: mimic of Hodgkin disease in nodal metastases. AB - We present the clinical and pathologic findings of a case of sclerosing mucoepidermoid carcinoma with eosinophilia of the thyroid in a 39-year-old woman. This particular case is notable because it initially presented as a cervical lymph node metastasis, and the initial clinical and histologic impression was Hodgkin disease, nodular sclerosis type. Sclerosing mucoepidermoid carcinoma with eosinophilia is a differentiated malignant neoplasm of the thyroid that can be confused with anaplastic carcinoma, medullary carcinoma, squamous cell carcinoma, or, as in this case, Hodgkin disease. A correct diagnosis of sclerosing mucoepidermoid carcinoma with eosinophilia involves awareness of this entity and appropriate immunohistochemical analysis. In this article, we briefly review the literature and stress the histologic and cytologic findings characteristic of sclerosing mucoepidermoid with eosinophilia of the thyroid. PMID- 10705404 TI - Fulminant parvovirus infection following erythropoietin treatment in a patient with acquired immunodeficiency syndrome. AB - We report the case of a 41-year-old black man with acquired immunodeficiency syndrome who developed a severe chronic anemia due to parvovirus infection. Bone marrow biopsy revealed erythroid aplasia. The infectious nature of the anemia was not recognized, and the patient was treated with erythropoietin. The patient's reticulocyte response was inadequate, however, and he remained anemic. A second bone marrow biopsy showed erythroid hyperplasia and prominent intranuclear parvovirus inclusions within erythroid progenitors. Erythropoietin was discontinued and was followed by a course of intravenous immunoglobulin, which resulted in rapid correction of anemia. To our knowledge, this is the first reported case of fulminant human parvovirus infection exacerbated by erythropoietin administration and documented by sequential bone marrow histologic examination. This case illustrates the critical importance of considering parvovirus in the etiology of chronic anemia with erythroid aplasia in immunocompromised patients. PMID- 10705406 TI - Carcinoid-related angiomatous polyposis simulating Crohn disease. AB - Polyposis associated with ileal carcinoid tumors is a rarely described pathologic mucosal transformation that may simulate inflammatory or neoplastic polyps. We describe a case with innumerable sessile polyps and groups of large filiform-like polyps of the terminal ileum associated with submucosal carcinoid tumors and a large mesenteric carcinoid tumor mass. The clinical, radiographic, and endoscopic presentation of the polyposis, together with the presence of multiple small bowel stenotic lesions, simulated Crohn disease. We propose the descriptive terminology angiomatous polyposis to describe the striking microscopic vascular proliferation that characterizes these polyps. The distribution of these lesions, with the most profuse polyposis in the immediate proximity of the carcinoid nests, and the immunohistochemical demonstration of growth factor substances, such as transforming growth factor alpha within neoplastic cells and adjacent polyps, suggest a tumor factor-mediated stromal proliferation. PMID- 10705407 TI - Basaloid squamous cell carcinoma occurring in the urinary bladder. AB - Basaloid squamous cell carcinoma is a recently described, distinct variant of squamous cell carcinoma that arises predominantly in the upper aerodigestive tract. Herein we report a case of basaloid squamous cell carcinoma arising in the urinary bladder. The patient was a 60-year-old woman who experienced intractable urinary tract infections following multiple corrective surgical procedures for incontinence. Biopsies of cystoscopically evident flat lesions were performed, and the patient subsequently underwent a radical cystectomy. Histologically, the lesions consisted of nests of basaloid cells with brisk mitotic activity, areas of squamous differentiation along with areas of squamous metaplasia, and squamous cell carcinoma in situ. These features are similar to those of basaloid squamous cell carcinoma described elsewhere in the body. To our knowledge, this is the first reported case of basaloid squamous cell carcinoma in the urinary bladder. PMID- 10705408 TI - Transformation of monocytoid B-cell lymphoma to large cell lymphoma associated with crystal-storing histiocytes. AB - We report a case of crystal-storing histiocytosis associated with large cell lymphoma in a patient with a history of monocytoid B-cell lymphoma 10 years previously. The cervical lymph node biopsy showed a diffuse proliferation of large lymphocytes with vesicular nuclear chromatin and distinct nucleoli. These lymphocytes were associated with numerous immunoglobulin lambda light-chain crystal-storing histiocytes, which morphologically resembled rhabdomyoblasts. Occasional lymphoid cells also showed large immunoglobulin crystals. This case establishes the association of crystal-storing histiocytes with lymphomas of mucosa-associated lymphoid tissue and emphasizes the need for immunophenotyping to distinguish these unusual cases from other tumors, particularly adult rhabdomyomas. PMID- 10705409 TI - Atypical lobular hyperplasia of the breast: a study of pathologists' responses in the College of American Pathologists Performance Improvement Program in Surgical Pathology. PMID- 10705410 TI - Benign intraneural epithelium in the breast. PMID- 10705411 TI - Pathologic quiz case: infection in a patient with a cervical arteriovenous malformation. PMID- 10705412 TI - Pathologic quiz case: an 11-day-old boy with lethargy, jaundice, fever, and melena. PMID- 10705417 TI - Lucas' pathology of tumors of the oral tissues PMID- 10705418 TI - The principles of clinical cytogenetics PMID- 10705419 TI - Tumors of the ovary, maldeveloped gonads, fallopian tube, and broad ligament PMID- 10705420 TI - Heroic nurses. PMID- 10705421 TI - Should suctioning be left to the nurse? PMID- 10705422 TI - Use of nursing resources and comfort of cancer patients with and without do-not resuscitate orders in the intensive care unit. AB - BACKGROUND: Little is known about the level of comfort experienced by cancer patients with do-not-resuscitate orders and how use of nursing resources affects their comfort. OBJECTIVE: To explore the relationship between use of nursing resources and comfort in cancer patients with and without do-not-resuscitate orders in the intensive care unit. METHODS: The sample consisted of 30 adult patients who had do-not-resuscitate orders and 30 randomly selected patients who did not. Pairs consisting of 1 patient from each group were admitted to the study simultaneously and were evaluated during the same observation period. Level of comfort was assessed by using the PACU Behavioral Pain Rating Scale. Data on use of nursing resources, determined with the Therapeutic Intervention Scoring System, and on demographics and severity of illness were abstracted from the medical records. RESULTS: Chi-square analyses revealed no significant relationship between comfort and use of nursing resources. Differences between the 2 types of patients in comfort and in use of resources were not significant. Evaluation of the multivariate relationship between comfort and use of resources, with do-not-resuscitate status added as a further predictor variable, revealed no significant relationships. Severity of illness and a patient's number of visitors were predictors of use of nursing resources. CONCLUSIONS: Despite high use of nursing resources, nurses continue to focus on comfort as an outcome of care irrespective of patients' do-not-resuscitate status. PMID- 10705423 TI - Critical care nurses' perceptions of obstacles and helpful behaviors in providing end-of-life care to dying patients. AB - BACKGROUND: Little is known about nurses' perceptions of obstacles or helpful behaviors ("helps") in providing end-of-life care in the intensive care setting. OBJECTIVE: To determine the importance of various obstacles and helps in providing end-of-life care as perceived by critical care nurses. METHODS: A questionnaire was mailed to 300 members of the American Association of Critical Care Nurses. Nurses were asked to rate obstacles and helps in giving end-of-life care, and additional obstacles and/or helps, and answer demographic questions. RESULTS: Six of the top 10 obstacles were related to issues with patients' families that make care at the end of life more difficult, such as the family's not fully understanding the meaning of life support, not accepting the patient's poor prognosis, requesting more technical treatment than the patient wished, and being angry. Added obstacles related mostly to problems with physicians' behavior. Most helps were ways to make dying easier for patients and patients' families, such as agreement among physicians about care, dying with dignity, and families' acceptance of the prognosis. Added helps included allowing music, pets, and so forth into the patient's room. CONCLUSIONS: Nurses have difficulties with patients' families and physicians concerning end-of-life issues, especially when the behaviors remove the nurses from caring for a patient or cause the patient pain or prolong suffering. Nurses do not acknowledge having difficulty providing care to dying patients aside from conflicts that arise because of patients' families and physicians. PMID- 10705424 TI - State of the science on parental stress and family functioning in pediatric intensive care units. AB - BACKGROUND: Critical illness of a child is stressful for parents and may affect family functioning. Most research on hospitalization in pediatric intensive care units has focused on the immediate responses of parents to the experience. OBJECTIVE: To critically review literature about pediatric intensive care units and to link those studies to a theoretical framework: McCubbin and McCubbin's resiliency model of family stress, adjustment, and adaptation. An updated synthesis of the literature is essential to prevent unnecessary duplication of research. METHODS: Guidelines presented by Ryan-Wenger were used to critique the scientific credibility and integrity of 38 research reports found by searching MEDLINE, the Cumulative Index to Nursing and Allied Health, and reference lists. The critique was organized according to the components of the research process, and then study results were reviewed according to the variables of the resiliency model. RESULTS: Most publications focused on variables in the adjustment phase, including stressors, resources, perceptions of stressors, and outcomes for patients' families. Obvious gaps in knowledge were related to families' vulnerability, type, and problem-solving and coping strategies. Many of the studies were biased toward white persons and toward mothers. CONCLUSIONS: Further research is warranted on (1) families of various ethnic backgrounds; (2) fathers and their low participation rates; (3) mother and father comparisons; (4) replication of interventional research with larger and more diverse samples; (5) exploratory and prospective, longitudinal research; and (6) research with children in pediatric intensive care units. PMID- 10705425 TI - Effects of 3 analgesic regimens on the perception of pain after removal of femoral artery sheaths. AB - BACKGROUND: Effective pain management after removal of femoral artery sheaths after percutaneous transluminal coronary angioplasty is highly individualized and requires frequent, accurate assessment and administration of analgesics as needed. OBJECTIVE: To determine which of 3 analgesic regimens is most effective in decreasing patients' perception of pain with the fewest side effects after removal of a femoral artery sheath. SAMPLE: 130 adult who had undergone percutaneous transluminal coronary angioplasty and were in an 8-bed cardiac short stay unit in a 1400-bed acute care hospital. METHOD: Patients were randomized to receive either intravenous morphine, intravenous fentanyl, subcutaneous lidocaine around the sheath site, or an intravenous placebo before sheath removal. Rescue analgesia (intravenous fentanyl) was made available to all groups. Patients used a visual analog scale to assess pain within 10 minutes before, 1 minute after, and 20 minutes after sheath removal. Pain levels, frequency of side effects, and use of rescue analgesia were compared among groups. RESULTS: Age, sex, number of stents, and frequency of hematomas did not differ significantly among groups. Pain ratings, use of rescue analgesia, and side effects (nausea, vomiting, or vasovagal symptoms) were not significantly different among treatment groups. Ratings of pain were slightly higher immediately after sheath removal in all groups. CONCLUSION: For most patients, removal of femoral artery sheaths and manual compression for hemostasis are relatively pain-free. Pain scores among patients given analgesia with subcutaneous lidocaine, intravenous morphine, or intravenous fentanyl were not significantly different from pain scores among control patients. PMID- 10705426 TI - Assessing dopamine concentrations: an evidence-based approach. AB - BACKGROUND: Both overmedication and undermedication can be potentially life threatening. If the actual volume of a 100-mL intravenous bag used to mix dopamine solutions is greater than the labeled volume, overdilution of medication can occur, resulting in an ineffective hemodynamic response in patients and thus an unintended adverse drug event. OBJECTIVES: To determine the actual fluid volumes of 100-mL intravenous bags, compare the actual volumes of 100-mL bags from the 3 major manufacturers of intravenous bags, and determine if the excess volume is sufficient to cause a clinically significant overdilution of dopamine. METHODS: A comparative descriptive design was used. The volumes of 162 intravenous bags of 100 mL of 5% dextrose in water (32 lot numbers with various expiration dates) were measured. Visual volume was confirmed by using a 250-mL graduated cylinder. Volume by weight was determined with a calibrated laboratory quality electronic scale. On the basis of a mathematical model, any overfill greater than 110 mL was considered clinically significant. RESULTS: The difference between actual and labeled volumes was statistically and clinically significant. Mean visual volume was 110.20 mL (range, 107-114 mL). Mean weighed volume was 109.26 mL (range, 106.15-112.09 mL). The fluid volumes among bags from the 3 major IV companies differed significantly (P < .001). CONCLUSIONS: The overfill in sufficient numbers of 100-mL intravenous bags was enough to cause clinically significant overdilution of dopamine. When dopamine or other vasoactive medications are mixed, either an in-line buret or premixed bags of the drugs should be used to prevent an unintended adverse drug event. PMID- 10705427 TI - Impact of a nurse-managed heart failure clinic: a pilot study. AB - BACKGROUND: One approach to optimize clinical and economic management of congestive heart failure is the use of multidisciplinary outpatient clinics in which advanced practice nurses coordinate care. One such clinic was developed in 1995 at a southeastern university hospital to enhance management of patients with chronic congestive heart failure. OBJECTIVES: To evaluate the effects of a multidisciplinary outpatient heart failure clinic on the clinical and economic management of patients with congestive heart failure. METHODS: Data on hospital readmissions, emergency department visits, length of stay, charges, and reimbursement from the 6 months before 15 patients joined a heart failure clinic were compared with data from the 6 months after the patients joined the clinic. RESULTS: The patients had a total of 38 hospital admissions (151 hospital days) in the 6 months before joining the clinic and 19 admissions (72 hospital days) in the 6 months afterward. The mean length of stay decreased from 4.3 days in the 6 months before joining to 3.8 days in the 6 months afterward, and the number of emergency department visits also decreased, although neither decrease was statistically significant. Mean inpatient hospital charges decreased from $10,624 per patient admission to $5893. Reimbursements were $7751 (73% collection rate) and $5138 (87% collection rate), respectively. CONCLUSIONS: Patients seemed to benefit from participation in the heart failure clinic. If a healthcare provider is available to manage early signs and symptoms of worsening heart failure, hospital readmissions may be decreased and patients' outcomes may be improved. PMID- 10705428 TI - The Framingham Heart Study: a pivotal legacy of the last millennium. AB - The life span at birth in the Greco-Roman era (200-300 BC) was 27 years. The life span in 1900 was 47 years, representing a gain of 20 years in more than 2000 years, or an increase of 1 year per century. At the close of the 20th century, the average life span was 77 years. This is a 30-year gain in life expectancy in only 1 century, or an average of 3 years for every decade in the last century. Compare that with only 1 year gain in life expectancy per century for the previous 20 centuries. It must be quite evident from this brief review that in the last century the Framingham Heart Study played a pivotal role in influencing physicians and the public to place major emphasis on the prevention of CV disease. To be sure, the control of epidemics, cure of infections with antibiotics, universal vaccination, and establishing food and environmental safety standards in the last century were instrumental in extending the life span. Nevertheless, major and direct influences on the explosive expansion in longevity during our life time were the contributions of the Framingham Heart Study. To paraphrase W. B. Kannel, a chief investigator in the Framingham Heart Study, "A cardiovascular event should be regarded as a medical failure rather than the first indication for the need to treat." PMID- 10705429 TI - Developing empathy in students. PMID- 10705430 TI - The Seventh International Conference on Intelligent Systems for Molecular Biology (ISMB'99) PMID- 10705431 TI - Genes regulated cooperatively by one or more transcription factors and their identification in whole eukaryotic genomes. AB - MOTIVATION: The question addressed here is how cooperative interactions among transcription factors (TFs), a very frequent phenomenon in eukaryotic transcriptional regulation, can be used to identify genes that are regulated by one or more TFs with known DNA binding specificities. Cooperativity may be homotypic, involving binding of only one transcription factor to multiple sites in a gene's regulatory region. It may also be heterotypic, involving binding of more than one TF. Both types of cooperativity have in common that the binding sites for the respective TFs form tightly linked 'clusters', groups of binding sites often more closely associated than expected by chance alone. RESULTS: A statistical technique suitable for the identification of statistically significant homotypic or heterotypic TF binding site clusters in whole eukaryotic genomes is presented. It can be used to identify genes likely to be regulated by the TFs. Application of the technique is illustrated with two transcription factors involved in the cell cycle and mating control of the yeast Saccharomyces cerevisiae, indicating that the results obtained are biologically meaningful. This rapid and inexpensive computational method of generating hypotheses about gene regulation thus generates information that may be used to guide subsequent costly and laborious experimental approaches, and that may aid in the assignment of biological functions to putative open reading frames. PMID- 10705433 TI - FingerPRINTScan: intelligent searching of the PRINTS motif database. AB - MOTIVATION: By identifying an unknown gene or protein as a member of a known family, we can infer a wealth of previously compiled information pertinent to that family and its members. RESULTS: This paper introduces a method that classifies sequences using familial definitions from the PRINTS database, allowing progress to be made with the identification of distant evolutionary relationships. The approach makes use of the contextual information inherent in a multiple-motif method, and has the power to identify hitherto unidentified relationships in mass genome data. We exemplify our method by a comparison of database searches with uncharacterized sequences from the Caenorhabditis elegans and Saccharomyces cerevisiae genome projects. This analysis tool combines a simple, user-friendly interface with the capacity to provide an 'intelligent', biologically relevant result. PMID- 10705432 TI - A new method to predict the consensus secondary structure of a set of unaligned RNA sequences. AB - MOTIVATION: To predict the consensus secondary structure, possibly including pseudoknots, of a set of RNA unaligned sequences. RESULTS: We have designed a method based on a new representation of any RNA secondary structure as a set of structural relationships between the helices of the structure. We refer to this representation as a structural pattern. In a first step, we use thermodynamic parameters to select, for each sequence, the best secondary structures according to energy minimization and we represent each of them using its corresponding structural pattern. In a second step, we search for the repeated structural patterns, i.e. the largest structural patterns that occur in at least one sequence, i.e. included in at least one of the structural patterns associated to each sequence. Thanks to an efficient encoding of structural patterns, this search comes down to identifying the largest repeated word suffixes in a dictionary. In a third step, we compute the plausibility of each repeated structural pattern by checking if it occurs more frequently in the studied sequences than in random RNA sequences. We then suppose that the consensus secondary structure corresponds to the repeated structural pattern that displays the highest plausibility. We present several experiments concerning tRNA, fragments of 16S rRNA and 10Sa RNA (including pseudoknots); in each of them, we found the putative consensus secondary structure. PMID- 10705434 TI - An analysis of the Protein Data Bank in search of temporal and global trends. AB - MOTIVATION: Biological databases, with their rapidly expanding contents, are indispensable tools in the quest to understand more about biological function. However, a serious user of a database that comprises a large collection of data, collected over a long period, will likely be struck by the inconsistency in reporting individual items of data. This paper takes a critical look at the Protein Data Bank (PDB) to explore the seriousness of the problem in one particular data set and to explore the implications to those actively engaged in comparative analysis of these data. RESULTS: Averaged over the complete corpus, the stereochemical quality of atomic models has, in the past few years, moved towards ideal values. At the same time, there are inconsistencies in how data are reported. Water content is not reported consistently and the percent of data collected when reporting the high-resolution shell varies, detracting from the value of resolution as a yardstick for assessing the quality of a structure. A more detailed analysis of these inconsistencies is hampered by the lack of machine-readable experimental data. To the user of macromolecular structure data, this suggests that structural details beyond the standard quality measures of resolution and R value should be considered when using coordinate sets for further derivation or in inferring biological function. To the curators of the PDB, this suggests the need to capture more of the experimental data associated with the experiment in a way that permits straightforward parsing. PMID- 10705435 TI - Homonyms and synonyms in the Dictionary of Interfaces in Proteins (DIP). AB - MOTIVATION: Should reports on molecular mimicry in particular cases, e.g. responsible for cross-reactivity, be considered as accidental or as a general principle in protein evolution? To answer this question, two types of similarity have to be considered: those in homologues (synonyms) and resemblance between patches from unrelated proteins (homonyms). RESULTS: All interfaces from known protein structures were collected in a comprehensive data bank [Dictionary of Interfaces in Proteins (DIP)]. A fast, sequence-independent, three-dimensional superposition procedure was developed to search automatically for geometrically similar surface areas. Surprisingly, we found a large number of structurally similar interfaces on the surface of unrelated proteins. Even patches from different types of secondary structure were found resembling each other. The putative functional meaning of homonyms is demonstrated with striking examples. PMID- 10705436 TI - bioWidgets: data interaction components for genomics. AB - MOTIVATION: The presentation of genomics data in a perspicuous visual format is critical for its rapid interpretation and validation. Relatively few public database developers have the resources to implement sophisticated front-end user interfaces themselves. Accordingly, these developers would benefit from a reusable toolkit of user interface and data visualization components. RESULTS: We have designed the bioWidget toolkit as a set of JavaBean components. It includes a wide array of user interface components and defines an architecture for assembling applications. The toolkit is founded on established software engineering design patterns and principles, including componentry, Model-View Controller, factored models and schema neutrality. As a proof of concept, we have used the bioWidget toolkit to create three extendible applications: AnnotView, BlastView and AlignView. PMID- 10705437 TI - ORBIT: an integrated environment for user-customized bioinformatics tools. AB - MOTIVATION: There are a large number of computational programs freely available to bioinformaticians via a client/server, web-based environment. However, the client interface to these tools (typically an html form page) cannot be customized from the client side as it is created by the service provider. The form page is usually generic enough to cater for a wide range of users. However, this implies that a user cannot set as 'default' advanced program parameters on the form or even customize the interface to his/her specific requirements or preferences. Currently, there is a lack of end-user interface environments that can be modified by the user when accessing computer programs available on a remote server running on an intranet or over the Internet. RESULTS: We have implemented a client/server system called ORBIT (Online Researcher's Bioinformatics Interface Tools) where individual clients can have interfaces created and customized to command-line-driven, server-side programs. Thus, Internet-based interfaces can be tailored to a user's specific bioinformatic needs. As interfaces are created on the client machine independent of the server, there can be different interfaces to the same server-side program to cater for different parameter settings. The interface customization is relatively quick (between 10 and 60 min) and all client interfaces are integrated into a single modular environment which will run on any computer platform supporting Java. The system has been developed to allow for a number of future enhancements and features. ORBIT represents an important advance in the way researchers gain access to bioinformatics tools on the Internet. PMID- 10705438 TI - MUTbase: maintenance and analysis of distributed mutation databases. AB - MOTIVATION: To develop tools for the submission of mutations to databases and maintenance of locus-specific mutation databases. Advanced, integrated computer systems are needed to store and organize the increasing mutation information. RESULTS: The MUTbase program suite provides an easy, interactive and quality controlled submission of information to mutation databases. For further study of the databases on the World Wide Web, a number of tools are provided. The program package also writes and updates a large number of Web pages, e.g. about the distribution and statistics of disease-causing mutations, and changes in restriction patterns. PMID- 10705439 TI - Completing the E. coli proteome: a database of gene products characterised since the completion of the genome sequence. AB - A database collating research on E. coli genes whose products have been characterised subsequent to in silico predictions from the completed genome sequence. PMID- 10705440 TI - MULTICLUSTAL: a systematic method for surveying Clustal W alignment parameters. AB - MULTICLUSTAL is a Perl script designed to automate the process of alignment parameter choice for Clustal W with the goal of generating high quality multiple sequence alignments. PMID- 10705441 TI - Role of conserved histidine residues in D-aminoacylase from Alcaligenes xylosoxydans subsp. xylosoxydans A-6. AB - D-Aminoacylase from Alcaligenes xylosoxydans subsp. xylosoxydans A-6 (Alcaligenes A-6) was strongly inactivated by diethylpyrocarbonate (DEPC). An H67N mutant was barely active, with a kcat/Km 6.3 x 10(4) times lower than that of the recombinant wild-type enzyme, while the H67I mutant lost detectable activity. The H67N mutant had almost constant Km, but greatly decreased kcat. These results suggested that His67 is essential to the catalytic event. Both H69N and H69I mutants were overproduced in the insoluble fraction. The kcat/Km of H250N mutant was reduced by a factor of 2.5 x 10(4)-fold as compared with the wild-type enzyme. No significant difference between H251N mutant and wild-type enzymes in the Km and kcat was found. The Zn content of H250N mutant was nearly half of that of wild-type enzyme. These results suggest that the His250 residue might be essential to catalysis via Zn binding. PMID- 10705442 TI - Screening for the in vitro anti-tumor-promoting activities of edible plants from Malaysia. AB - A total of 114 methanol extracts from 42 plant families of edible Malaysian plants were screened for their inhibitory activities toward tumor promoter 12-O hexadecanoylphorbol-13-acetate (HPA)-induced Epstein-Barr virus (EBV) activation in Raji cells. By testing at a concentration of 200 micrograms/ml, 74% of the 114 extracts inhibited EBV activation by 30% or more. This rate is comparable to those observed in the previous tests on edible Thai (60%) and Indonesian (71%) plants, and, importantly, much higher than that (26%) observed for Japanese edible plants. Approximately half of the Malaysian plants did not taxonomically overlap those from the other three countries, suggesting that Malaysian plants, as well as Thai and Indonesian plants, are an exclusive source of effective chemopreventive agents. Further dilution experiments indicated an extract from the leaves of Piper betle L. (Piperaceae) to be one of the most promising species. The high potential of edible Southeast Asian plants for cancer chemoprevention is collectively discussed. PMID- 10705443 TI - Generation of recombinant human large latent transforming growth factor-beta 1 and monoclonal antibodies to it. AB - Transforming growth factor beta 1 (TGF-beta 1) is a regulator of cell growth and differentiation. It is produced in various of cells and tissues as a biologically latent complex, whose significance is still unknown. We established a Chinese hamster ovary cells that produced recombinant human large latent TGF-beta 1. The growth factor was purified from serum-free conditioned medium of the cell line was purified to apparent homogeneity by four steps of column chromatography. The purified protein gave a single band with the apparent molecular weight of 210,000 on SDS-PAGE, and had four subunits, of 12.5, 40, 53, and 150-190 kDa. These components were identical to TGF-beta 1, the N-terminal remnant of pro-TGF-beta 1, pro-TGF-beta 1, and latent TGF-beta 1 binding protein, respectively. The purified growth factor had biological activity similar to that of the growth factor purified from human platelets. We prepared four monoclonal antibodies by immunization of mice with the recombinant protein. In western blotting, two of the antibodies bound to latent TGF-beta 1 binding protein. The two other antibodies reacted with the N-terminal remnant of pro-TGF-beta 1. Recombinant large latent TGF-beta 1 and its monoclonal antibodies could be used for detailed structural and functional studies of the large latent TGF-beta 1 complex. PMID- 10705444 TI - Screening for Drosophila proteins with distinct expression patterns during development by use of monoclonal antibodies. AB - Kc 167 is a cell line established from Drosophila embryonic hemocytes and has been shown to express many extracellular matrix (ECM) and other proteins important during development. We have screened monoclonal antibodies (mAbs) raised against heparin affinity purified proteins from conditioned medium of Kc 167 cells to identify novel proteins with important roles for development. One mAb recognized a protein expressed with temporary and tissue specific patterns during Drosophila embryogenesis and larval development. This approach is an alternative to screening of Expression Sequence Tag (EST) clones by in situ hybridization to initiate reverse genetics. In addition, a number of mAbs recognizing ECM proteins were also identified. These mAbs will be useful for biochemical and cell biological analyses of Drosophila ECM proteins. PMID- 10705445 TI - Gene encoding a dextransucrase-like protein in Leuconostoc mesenteroides NRRL B 512F. AB - A gene, dsrT, encoding a dextransucrase-like protein was isolated from the genomic DNA libraries of Leuconostoc mesenteroides NRRL B-512F dextransucrase like gene. The gene was similar to the intact open reading frames of the dextransucrase gene dsrS of L. mesenteroides NRRL B-512F, dextransucrase genes of strain NRRL B-1299 and streptococcal glucosyltransferase genes, but was truncated after the catalytic domain, apparently by the deletion of five nucleotides. dsrT mRNA was produced in this strain L. mesenteroides when cells were grown in a sucrose medum, but at a level of 20% of that of dsrS mRNA. The molecular weight of the dsrT gene product was 150,000 by SDS-PAGE. The product did not synthesize dextran, but had weak sucrose cleaving activity. The insertion of five nucleotides at the putative deletion point in dsrT resulted in an enzyme with a molecular weight of 210,000 and with dextransucrase activity. PMID- 10705446 TI - Induction and repression of a Streptomyces lividans chitinase gene promoter in response to various carbon sources. AB - Induction and repression of a gene for chitinase (chiA) in Streptomyces lividans was investigated using a catechol 2,3-dioxygenase gene (xylE) as the reporter gene. Of various substrates examined, expression of the promoter (PchiA) was observed after a delay when colloidal chitin or small chitin-oligosaccharides were added to the medium. N-acetylglucosamine completely repressed the chiA promoter. The duration of the delay in expression of PchiA differed with the inducer used, with chitobiose inducing the activity most rapidly. The minimum concentration of chitobiose needed for induction was 1 microM. It appears, therefore, that an efficient inducer of the gene for chitinase in S. lividans is chitobiose. PMID- 10705447 TI - Amino acid sequence of an unique ribonuclease with a C-terminus rich in O glycosylated serine and threonine from culture medium of Lentinus edodes. AB - The mushroom Lentinus edodes produces three base-non-specific and acid ribonucleases, RNases Le2, Le37, and Le45. The latter two are excreted from mycelia into the medium. The primary structure of RNase Le37, which had a molecular mass of 37 kDa, was sequenced. It was a member of the RNase T2 family, as is RNase Le2. RNase Le37 was some 30 amino acid residues longer at the C terminal end than RNase Le2. The C-terminal region of RNase LE37 was rich in O glycosylated serine and threonine. In fungal glucoamylases and chitinases, which hydrolyze raw-starch and chitin, respectively, have structures resembling the structure of the C-terminal of RNase Le37. PMID- 10705448 TI - Molecular modeling study of highly branching (1-->3)-alpha-D-glucan, a model polysaccharide for cariogenic glucan, using the N-H mapping method. AB - A systematic search for possible regular helical structures of a highly branching (1-->3)-alpha-D-glucan was done using the n-h mapping technique, combined with MM3-generated relaxed-residue energy map calculations with respect to the conformations of the backbone glycosidic linkages. The alpha-D-glucan, consisting of a (1-->3)-alpha-linked backbone with alpha-D-glucose side residues attaching to an O6 atom of every second backbone residue, was considered as a model polysaccharide of a branching part of the glucan produced by oral bacteria, which was known to be related to dental plaque formation and to contribute to dental caries. The potential energy surfaces of the trisaccharide repeating unit of the branching alpha-D-glucan indicated that (1-->6)-alpha-linked side residues did not appear to interfere significantly with the backbone stereochemistry, probably due to a further separation of the three-bond-linked side residue compared with an ordinary two-bond-linked residue. Based on the n-h maps of the branching alpha D-glucan, the side residues, when involved in a complete helix, mostly contributed additional stabilizations to particular helical structures. It was found by checking the typical helix models that formation of hydrogen bonds involving side residues was probably a major cause of the stabilization. This hydrogen bonding was expected to increase insolubility for the glucan chain--a typical, physical property observed for the bacterial alpha-D-glucan--by introducing its backbone stereochemistry as an additional stiff feature. PMID- 10705449 TI - Structure of ribulose 1,5-bisphosphate carboxylase/oxygenase gene cluster from a thermophilic hydrogen-oxidizing bacterium, Hydrogenophilus thermoluteolus, and phylogeny of the fructose 1,6-bisphosphate aldolase encoded by cbbA in the cluster. AB - Four genes, cbbO, cbbY, cbbA, and the pyruvate kinase gene (pyk), were found downstream of ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) genes, cbbLS, from a thermophilic hydrogen-oxidizing bacterium, Hydrogenophilus thermoluteolus (formerly Pseudomonas hydrogenothermophila). cbbO was similar to norD in the denitrification gene cluster, and cbbY was similar to cbbY from other autotrophic bacteria. cbbA encoded fructose 1,6-bisphosphate aldolase (FBP aldolase); however, CbbA was little similar to other CbbA proteins. When CbbA was overexpressed in Escherichia coli, overproduction of CbbA was detected by SDS PAGE. However, the cell extract had slightly higher activity than a cell extract of E. coli without cbbA. Phylogenetic analysis showed class II FBP aldolase divided into classes IIA and IIB, and that CbbA from H. thermoluteolus was in class IIA. Activities of RubisCO and FBP aldolase were examined under autotrophic, mixotrophic, and heterotrophic conditions. The activities of the two enzymes were regulated independently. PMID- 10705450 TI - Effects of hydrostatic pressure and temperature on growth and lipid composition of the inner membrane of barotolerant Pseudomonas sp. BT1 isolated from the deep sea. AB - A barotolerant member of the genus Pseudomonas was isolated from deep-sea sediment obtained from the Japan Trench, at a depth of 4418 m. The growth temperature was found to affect the hydrostatic pressure range in which the bacterium could grow; the optimum hydrostatic pressure for growth shifted to a higher pressure with increasing temperature. We examined the lipid composition of the inner membrane of cells grown at various hydrostatic pressures and temperatures. The fatty acid components of the inner membrane lipids were C16:0, C16:1, C18:0, and C18:1. The phospholipid components of the inner membrane were phosphatidylethanolamine, cardiolipin, phosphatidylglycerol, and phosphatidylserine. It is evident that the effects of elevated hydrostatic pressure are comparable to the effects of low temperature on both the fatty acid composition of the inner membrane lipids and the phospholipid composition of the inner membrane of this bacterium. PMID- 10705451 TI - Sequence of egV and properties of EgV, a Ruminococcus albus endoglucanase containing a dockerin domain. AB - The Ruminococcus albus F-40 egV gene, encoding endoglucanase V (EGV), consists of an open reading frame of 1,833 nucleotides and encodes 611 amino acids with a deduced molecular weight of 67,103. The deduced EGV is a modular enzyme composed of a catalytic domain of family 5 of glycosyl hydrolases, a domain of unknown function, and a dockerin domain responsible for cellulosome assembly, suggesting that R. albus F-40 produces a cellulosome, and EGV is a component of the cellulosome. A truncated form of EGV with an apparent molecular weight of 42,000 was purified from a recombinant Escherichia coli and characterized since EGV suffered from partial proteolysis by E. coli protease(s). The truncated EGV was active toward carboxylmethyl cellulose, xylan, lichenan, and acid-swollen cellulose. The pH and temperature optima of the enzyme were 7.0 and 40 degrees C, respectively. By Western blot analysis using the antiserum raised against the truncated enzyme, EGV was detected in the whole cells but not in the culture supernatant of R. alubus F-40, suggesting that EGV was located on the cell surface. PMID- 10705452 TI - Requirement for lysine-19 of the yeast mitochondrial ATPase inhibitor for the stability of the inactivated inhibitor-F1Fo complex at higher pH. AB - The ATPase inhibitor is a regulatory subunit of mitochondrial ATP synthase. In this study, the role of Lys19 of the yeast ATPase inhibitor was examined by site directed mutagenesis. Two amino acids (Gln and Glu) were substituted for the Lys19. The purified mutant inhibitor (Lys19-->Gln) had similar ATPase inhibitory activity to that of the wild-type inhibitor at pH 6.5, but was less active at pH 7.4. ATP synthesis in mutant mitochondria was normally activated by the addition of ADP and succinate, but the inactivated ATPase complex in the mutant mitochondria was activated more readily than that in control cells by raising pH. These results show that Lys19 of the yeast ATPase inhibitor is not essential for ATPase inhibitory activity, but increases the stability of the inhibitor-F1Fo complex at higher pH. PMID- 10705453 TI - Purification and characterization of a thermostable chitinase from Streptomyces thermoviolaceus OPC-520 and cloning of the encoding gene. AB - When Streptomyces thermoviolaceus OPC-520 was grown in a minimal medium with 1% chitin, three activity bands corresponding to proteins of 40 kDa (Chi40), 30 kDa (Chi30), and 25 kDa (Chi25) were detected. Among them, Chi30 was purified from the culture filtrate of the strain. The molecular mass was estimated to be 30 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and its isoelectric point was 3.8. The optimum pH and temperature of Chi30 were 4.0 and 60 degrees C, respectively. Chi30 was stable at pH 6-8 up to 60 degrees C. The gene encoding Chi30 (chi30) was cloned and its nucleotides sequenced. The open reading frame of chi30 encoded a protein consisting of 347 amino acids with a calculated molecular weight of 35,621. The mature Chi30 consisted of only a catalytic domain and showed a significant similarity with ChiA from S. coelicolor and ChiA from S. lividans. The existence of a 12-bp direct repeat sequence in the promoter region of chi30 was detected, which have been suggested to be involved in both chitin induction and glucose repression. PMID- 10705454 TI - Okaramines N, O, P, Q and R, new okaramine congeners, from Penicillium simplicissimum ATCC 90288. AB - Five new okaramine congeners, okaramines N, O, P, Q, and R, were isolated from Penicillium simplicissimum ATCC 90288. Their structures were determined by an analysis of spectroscopic data. The insecticidal activity of these new okaramines was evaluated against silkworms. PMID- 10705455 TI - New paralytic alkaloids, asperparalines A, B and C, from Aspergillus japonicus JV 23. AB - New paralytic alkaloids, asperparalines A (1), B (2) and C (3), were isolated from okara (the insoluble residue of whole soybean) that had been fermented with Aspergillus japonicus JV-23. Their structures were elucidated by spectroscopic methods and X-ray crystallography. These asperparalines showed paralytic activity against silk worms. PMID- 10705456 TI - High molecular weight transglutaminase inhibitor produced by a microorganism (Streptomyces lavendulae Y-200). AB - Transglutaminases catalyze the cross-linking and amine incorporation of proteins, and are implicated in various biological phenomena such as blood clotting, wound healing, apoptosis, and cell differentiation. Streptomyces lavendulae Y-200, isolated from soil, produced a substance that inhibited transglutaminases. The inhibitory substance was purified from the cultured medium by procedures of acid precipitation, deoxyribonuclease treatment, and gel filtration chromatography. The partially purified sample was dark brown. The inhibitory activity was stable under acidic, alkaline, and high temperature conditions, and resistant to the treatment with proteinases such as trypsin and Pronase. The molecular weight of the inhibitory substance was estimated to be between 10(4) and 10(5) from its permeability through ultrafilter membranes. The acid hydrolysate of the inhibitory substance contained amino acids and sugars. The inhibitory substance inhibited both calcium-dependent and calcium-independent transglutaminases in a competitive manner with a glutamine substrate. The extent of inhibition caused by the calcium-dependent transglutaminase increased with increasing calcium concentration. The results obtained here may help identify a novel regulatory substance of transglutaminase in biological systems. PMID- 10705457 TI - Aspirin and salicylic acid do not inhibit methyl jasmonate-inducible expression of a gene for ornithine decarboxylase in tobacco BY-2 cells. AB - Similar to the prostanoid-mediated inflammatory response in mammals, jasmonate mediated wound response in plant leaves is inhibited by salicylic acid (SA) or acetylsalicylate (aspirin). In tobacco BY-2 cells, expression of the gene for ornithine decarboxylase (ODC) involved in putrescine synthesis is rapidly inducible by methyl jasmonate (MeJA). A nuclear gene for ODC isolated from tobacco, gNtODC-1, was an intron-less gene and MeJA induced the expression of a GUS fusion gene with the gNtODC-1 promoter in transformed tobacco cells. Although SA alone did not induce the expression, 0.2 to 20 microM SA increased the MeJA induced expression of the fusion gene to about two-fold. A similar increase was observed with aspirin but not with 3- or 4-hydroxybenzoic acids. SA at concentrations up to 200 microM did not inhibit the MeJA-induction of mRNAs for the GUS fusion gene and the endogenous gene for ODC. PMID- 10705458 TI - Enzymatic synthesis of stable, odorless, and powdered furanone glucosides by sucrose phosphorylase. AB - Sucrose phosphorylase from Leuconostoc mesenteroides catalyzed transglucosylation from sucrose to 4-hydroxy-3(2H)-furanone derivatives. When 4-hydroxy-2,5-dimethyl 3(2H)-furanone (HDMF) and 2-ethyl-4-hydroxy-5-methyl-3(2H)-furanone or 5-ethyl-4 hydroxy-2-methyl-3(2H)-furanone (EHMF) were used as acceptors, their transfer ratios were more than 45%. In the case of glucosylation of HDMF, the major transfer product was identified as 2,5-dimethyl-3(2H)-furanone 4-O-alpha-D glucopyranoside (DMF-G). In the case of glucosylation of EHMF, two major transfer products were obtained, and their structures were identified as 2-ethyl-5-methyl 3(2H)-furanone 4-O-alpha-D-glucopyranoside (2E5MF-G) and 5-ethyl-2-methyl-3(2H) furanone 4-O-alpha-D-glucopyranoside (5E2MF-G) on the bases of spectrometric investigations. These glucosides were more stable than each aglycone. The glucosylated HDMF, DMF-G, was an odorless chemical, on the other hand, HDMF had a pineapple flavor. The glucosylated EHMF (EMF-G) were white odorless powders, though aglycone EHMF was a pale yellow syrup like a caramel with an intense sweet odor. Although DMF-G and EMF-G showed little radical-scavenging activity, hydrolyzates of these glucosides by an intestinal acetone powder from pigs had antioxidative activity as well as their aglycones. It was suggested that these glucosides improved some physical properties and may become prodrugs by glucosylation. PMID- 10705459 TI - Molecular characterization of a novel yeast cell-wall acid phosphatase cloned from Kluyveromyces marxianus. AB - A novel Kluyveromyces marxianus gene that encodes an acid phosphatase, Pho610, was cloned in Saccharomyces cerevisiae. The deduced amino acid sequence was distinct from S. cerevisiae phosphatases but similar to some fungal enzymes. A peculiar feature of the sequence is that it has hydrophobic stretches both at the N- and C-termini, which is a characteristic of the precursors of glycosylphosphatidylinositol(GPI)-anchored proteins. When the gene was expressed in S. cerevisiae, the active enzyme was recovered in the periplasmic fraction by glucanase digestion. The Pho610 polypeptide was highly glycosylated and a significant portion was covalently linked to the cell-wall glucan. The enzyme was secreted when the C-terminal region was truncated to remove the GPI signal. Therefore, Pho610 is a novel cell-wall protein having an enzyme activity. PMID- 10705460 TI - Identification of a 14-3-3 protein from Lentinus edodes that interacts with CAP (adenylyl cyclase-associated protein), and conservation of this interaction in fission yeast. AB - We previously identified a gene encoding a CAP (adenylyl cyclase-associated protein) homologue from the edible Basidiomycete Lentinus edodes. To further discover the cellular functions of the CAP protein, we searched for CAP interacting proteins using a yeast two-hybrid system. Among the candidates thus obtained, many clones encoded the C-terminal half of an L. edodes 14-3-3 homologue (designated cip3). Southern blot analysis indicated that L. edodes contains only one 14-3-3 gene. Overexpression of the L. edodes 14-3-3 protein in the fission yeast Schizosaccharomyces pombe rad24 null cells complemented the loss of endogenous 14-3-3 protein functions in cell morphology and UV sensitivity, suggesting functional conservation of 14-3-3 proteins between L. edodes and S. pombe. The interaction between L. edodes CAP and 14-3-3 protein was restricted to the N-terminal domain of CAP and was confirmed by in vitro co precipitation. Results from both the two-hybrid system and in vivo co precipitation experiments showed the conservation of this interaction in S. pombe. The observation that a 14-3-3 protein interacts with the N-terminal portion of CAP but not with full-length CAP in L. edodes and S. pombe suggests that the C-terminal region of CAP may have a negative effect on the interaction between CAP and 14-3-3 proteins, and 14-3-3 proteins may play a role in regulation of CAP function. PMID- 10705461 TI - Effect of a phosphorylated guar gum hydrolysate on increased calcium solubilization and the promotion of calcium absorption in rats. AB - The effect of a phosphorylated guar gum hydrolysate (P-GGH) on calcium solubilization and its influence on calcium absorption were studied in vitro and in vivo. P-GGH was prepared by chemically modifying a guar gum hydrolysate (GGH) with sodium metaphosphate. P-GGH inhibited the precipitation of calcium phosphate in vitro. The apparent calcium absorption and the amount of femur calcium were significantly higher in rats fed on the P-GGH diet (50 g/kg of diet) than in rats fed on the GGH diet (50 g/kg of diet) or the control diet. Moreover, the amount of soluble calcium in the ileal contents was significantly higher in the P-GGH fed rats than in the GGH-fed rats. These results indicate that P-GGH may inhibit calcium phosphate formation in the lower part of the small intestine, and thus increase calcium absorption. PMID- 10705462 TI - Phosphatidylserine synthesis required for the maximal tryptophan transport activity in Saccharomyces cerevisiae. AB - Saccharomyces cerevisiae cho1/pss mutants, which are severely impaired in phosphatidylserine (PS) synthesis, do not have detectable amounts of PS in their lipid fractions. Their derivatives with mutations that cause defects in tryptophan synthesis grew poorly in a medium containing 5 micrograms/ml of L tryptophan, a concentration that met the requirements of tryptophanauxotrophic CHO1/PSS strains. The rates of tryptophan uptake of trp1 cho1/pss mutants were low at low tryptophan concentrations. This defect in the use of tryptophan was restored either by expression of CHO1/PSS or by introduction of a gene encoding tryptophan transporter, TAT1 or TAT2. These results indicate that PS synthesis is required for the maximal tryptophan-transporting activity of S. cerevisiae at low tryptophan concentrations. PMID- 10705463 TI - Radical-capturing reaction of 5,7,3',4'-tetramethylquercetin with the AIBN radical initiator. AB - In order to clarify the mechanism for the radical-capturing reaction which is initiated at the C3-hydroxyl group of flavonols, 5,7,3',4'-tetramethylquercetin (TMQ) was reacted with the 2,2'-azobis-isobutyronitrile (AIBN) radical initiator in benzene. Six products, one depside and its two hydrolytic products, one nitrile adduct, and two others, were isolated from the reaction mixture, and their structures were determined by instrumental analyses. The quantitative change to the four main products against the reaction time was measured by an HPLC method. The radical-capturing reaction pathway for TMQ with AIBN is proposed from these products and their quantitative changes. The pathway dividing into two clearly reveals that one subpath formed the depside and its hydrolytic products, while the other formed the nitrile adduct. The reactivity of each two sub-path was nearly the same, different from the case of TMQ and the 2,2'-azobis-2,4 dimethylvaleronitrile (AMVN) radical initiator. PMID- 10705464 TI - Ultraviolet spectrometric determination of neutral monosaccharides by HPLC with ethanolamine. AB - The reaction between ethanolamine and sugars in a borate solution gave intense UV absorption at 310 nm. By adding ethanolamine in the mobile phase, simple and sensitive ion-exchange HPLC of sugar-borate complexes was achieved. This method can be used to measure as little as 100 pmole of seven monosaccharides from pectic polysaccharides. PMID- 10705465 TI - Purification of collagenase and specificity of its related enzyme from Bacillus subtilis FS-2. AB - A collagenase in the culture supernatant of B. subtilis FS-2, isolated from traditional fish sauce, was purified. The enzyme had a molecular mass of about 125 kDa. It degraded gelatin with maximum activity at pH 9 and a temperature of 50 degrees C. The purified enzyme was stable over a wide range of pH (5-10) and lost only 15% and 35% activity after incubation at 60 degrees C and 65 degrees C for 30 min, respectively. Slightly inhibited by EDTA, soybean tripsin inhibitor, iodoacetamide, and iodoacetic acid, the enzyme was severely inhibited by 2-beta mercaptoethanol and DFP. The protease from B. subtilis FS-2 culture digested acid casein into fragments with hydrophilic and hydrophobic amino acids as C terminals, in particular Asn, Gly, Val, and Ile. PMID- 10705467 TI - Phytotoxicity of indole-3-acetic acid produced by the fungus, Pythium aphanidermatum. AB - Pythium aphanidermatum causes the serious disease of Pythium red blight on bentgrass. IAA, one of the metabolites that has been isolated from this fungus, showed the same symptom of Pythium red blight on bentgrass at a concentration of 1,000 mg/1. The IAA content in the foliage of bentgrass infected by this fungus was about 200 times that of an untreated control. These results suggest that IAA produced by this fungus was the causal substance of Pythium red blight on bentgrass. PMID- 10705466 TI - Inactivation of human type A and B influenza viruses by tea-seed saponins. AB - The effects of a mixture of tea-seed saponins obtained from the seeds of Camellia sinensis var. sinesis on human influenza viruses types A and B were investigated. At the concentrations of 60, 80, and 100 micrograms/ml, respectively, the mixture inactivated viruses A/Memphis/1/71 (H3N2), B/Lee/40, and A/PR/8/34 (H1N1) almost completely. The mixture also inactivated type A virus A/PR/8/34 after inoculation at concentrations of 1-30 micrograms/ml dose-dependently. PMID- 10705468 TI - Dual CCK-A and CCK-B receptor antagonists (II). Preparation and structure activity relationships of 5-alkyl-9-methyl-1,4-benzodiazepines and discovery of FR208419. AB - In our continuing research for dual CCK-A and -B antagonists, according to our hypothesis that dual CCK-A and -B antagonists should be more efficacious than selective CCK-A antagonists for the treatment of pancreatitis, we have prepared various 5-alkyl-9-methyl-1,4-benzodiazepines. From the compounds prepared, 1 cyclohexyl-carbonylmethyl-5-ethyl-9-methyl-3- (m-tolylureido)-2-oxo-1,4 benzodiazepine, (40) was selected as a candidate for development due to its well balanced high affinity for both receptors. The R-enantiomer of 40, (R)-40 (FR 208419), had 27-fold higher affinity for the CCK-A receptor and 8-fold more potent CCK-B receptor binding activity than (S)-40. The biological activity after p.o. administration of (R)-40, estimated from the ID50 value (0.23 mg/kg p.o.) obtained by preliminary evaluation by gastric emptying effects, is considered to be high enough for further development. This compound is now undergoing further biological evaluations with a view to clinical development. PMID- 10705469 TI - Difference spatial distribution function analysis of aqueous solutions. II. Hydration structures of dimethyl ether, 180 degrees ethyl methyl ether and 0 degree ethyl methyl ether solutions. AB - Monte Carlo simulations are systematically presented to demonstrate the influence of the hydrophobic group's steric bulk on hydration structure. We have simulated a dimethyl ether (DME), two conformations for ethyl methyl ether (0 degree EME and 180 degrees EME), and 0 degree ethanol solutions. Spatial distribution function (SDF), goo(x,y,z) and difference SDF (DSDF), delta goo(x,y,z), obtained from MC simulation in an infinitely dilute aqueous solution of ether show the three-dimensional probability of an atom-atom pair distribution between solute and solvent atoms. Based on the results of SDF in an infinitely dilute aqueous solution of ether, the distribution of hydration water molecules can be divided into hydrogen acceptor (HA) and hydrophobic hydration (HH), regions, and the spatial orientation of the hydrogen-bonded water in the HA region is found to form a triple-layer structure, as it does in alcohol solutions. From the results of an analysis of the DSDF delta goo(x,y,z) between the SDFs of EME and DME, it is apparent that the distribution changes of hydration water molecules in ether solutions are essentially similar to those in the alcohol solutions. Further, we show that the hydration water molecules are distributed mainly in the stable area in the binding energy's (BE) contour maps for each region. PMID- 10705470 TI - Nonpeptide arginine vasopressin antagonists for both V1A and V2 receptors: synthesis and pharmacological properties of 4-(1,4,5,6-tetrahydroimidazo[4,5 d][1]benzoazepine-6-carbonyl)benzanili de derivatives and 4'-(5,6-dihydro-4H thiazolo[5,4-d][1]benzoazepine-6-carbonyl)benzanilid e derivatives. AB - Arginine vasopressin (AVP) has a dual action mainly in the periphery, i.e., vasoconstriction and water reabsorption via V1A and V2 receptors; it may play a role in a number of diseases, including congestive heart failure (CHF), hypertension, renal disease, edema, and hyponatremia. We have attempted to develop a new series of orally active AVP antagonists for both V1A and V2 receptors based on the hypothesis that the blockade of both V1A and V2 receptors might be beneficial to CHF patients. In this report, a series of compounds structurally related to 4'-(1,4,5,6-tetrahydroimidazo[4,5-d][1]benzoazepine-6- carbonyl)benzanilide and 4'-(5,6-dihydro-4H- thiazolo[5,4-d][1]benzoazepine-6 carbonyl)benzanilide were synthesized and examined for AVP antagonist activity for both V1A and V2 receptors. As a result, it was found that the 4'-(1,4,5,6 tetrahydroimidazo[4,5-d][1]benzoazepine-6-carbon yl)-2- phenylbenzanilide derivatives showed potent binding affinity for both V1A and V2 receptors. Especially, 4'-(2-methyl-1,4,5,6- tetrahydroimidazo[4,5-d][1]benzoazepine-6 carbonyl)-2-phe nylbenzanilide monohydrochloride (18, YM087 = conivaptan hydrochloride) exhibited potent binding affinity and AVP antagonist activity, after intravenous administration, for both V1A and V2 receptors. Furthermore, YM087 exhibited the most potent oral activity for the V2 receptor. Details of the synthesis and pharmacological properties of this series are presented. PMID- 10705471 TI - Lipid A and related compounds. XXXVII. Determination of favorable binding linkages of lipid A analog to antigen moiety for synthetic vaccines. AB - For the determination of favorable binding linkages of lipid A analog as a synthetic immunoadjuvant to the antigen moiety for synthetic vaccines, new N acylated L-serine-containing D-glucosamine analogs (Type A, B, C) were synthesized and their mitogenicities were examined. Among chemically synthesized compounds (6-15, 30), compound 8 for Type B exhibited the most potent mitogenicity. PMID- 10705472 TI - Mono and trinuclear complexes of alpha-oximinoacetoacetylpyridine-4 phenylthiosemicarbazone. AB - Complexes of several transition metal ions with alpha-oximinoacetoacetyl pyridine 4-phenylthiosemicarbazone (H3OAPT) have been prepared. Attempts were made to elucidate their geometries by elemental analysis, molar conductance, magnetic measurements and by some spectroscopic (IR, ESR and electronic) techniques. All the investigated metal ions form mononuclear complexes except for CuII, which forms mononuclear and trinuclear complexes with its chloride and acetate salts, respectively. The IR spectra show that the ligand behaves as a mono or binegative tridentate. Moreover, it acts as a trinegative hexadentate in the trinuclear CuII complex. The protonation constants (logK1H = 9.9 and log K2H = 6.0), as well as the stability constants of the metal complexes, are determined by the pH titration of H3OAPT and its metal(II) complexes against 0.01 M NaOH. CuII complexes possess square-planar stereochemistry while CoII and NiII have an octahedral one. The crystal field parameters of CoII and NiII complexes are evaluated. PMID- 10705473 TI - 2-Hydroxy-5-(ethanolamino)-3-(10'-Z-pentadecenyl)-1,4-benzoquinone, new microbial phase II metabolite of maesanin. AB - The use of microbial models for biotransformation of the natural benzoquinone, maesanin (1), resulted in the isolation of an ethanolamine conjugate (5) from the culture broth of Debaryomyces polymorphus ATCC 20280. Metabolite 5 was characterized as 2-hydroxy-5-(ethanolamino)-3-(10'-Z-pentadecenyl)-1,4-benzoq uinone. The production of 5 represents a new type of phase II conjugation reaction in microbial systems. The results of preliminary mammalian metabolism of 1 in rats were inconclusive. PMID- 10705474 TI - Interactions of cyclodextrins with dipalmitoyl, distearoyl, and dimyristoyl phosphatidyl choline liposomes. A study by leakage of carboxyfluorescein in inner aqueous phase of unilamellar liposomes. AB - The interaction of cyclodextrins (CDs) with L-alpha-dipalmitoyl phopsatidyl choline (DPPC), L-alpha-distearoyl phosphatidyl choline (DSPC), and L-alpha dimyristoyl phosphatidyl choline (DMPC) unilamellar liposomes was investigated by the leakage of carboxylfluorescein (CF) entrapped in the inner aqueous phase of liposomes, at 25 degrees C (DPPC and DSPC liposomes) and at 5 degrees C (DMPC liposomes). The efficiency of CDs for CF leakage was remarkable in the order of heptakis (2,6-di-O-methyl)-beta-CD (DOM-beta-CD) > alpha-CD > heptakis (2,3,6-tri O-methy)-beta-CD (TOM-beta-CD) from DPPC liposomes, in the order of DOM-beta-CD > TOM-beta-CD > alpha-CD from DSPC liposomes and in the order of alpha-CD > DOM beta-CD > TOM-beta-CD from DMPC liposomes. The other CDs used in the present studies, beta-CD, 2-hydroxylpropyl beta-CD, and gamma-CD scarcely induced the CF leakage from above the three liposomes. From the profiles of % CF leakage, together with measurements of differential scanning calorimetry, it was found that hydrophobic DOM-beta-CD penetrates the matrix of the liposomes to interact with them as well as TOM-beta-CD, and that less hydrophobic alpha-CD exists at the surface of the membrane to interact with the liposomes. Further, it was found that the interaction of CDs with liposomes changes depending not only on the length of fatty acid chain of phospholipid (condensation force and hydrophobicity) but also the hydrophobicity and the cavity size of CD. PMID- 10705475 TI - Structure and stereochemistry of the higher bile acid isolated from turtle bile: (22S,25R)-3 alpha,12 alpha,15 alpha,22-tetrahydroxy-5 beta-cholestan-26-oic acid. AB - The structure and stereochemistry of the higher bile acid, tetrahydroxyisosterocholanic acid (TISA), which was previously isolated from the bile of Amyda japonica (turtle) and proposed as a tetrahydroxyisosterocholanic acid, have been established as (22S,25R)-3 alpha,12 alpha,15 alpha,22 tetrahydroxy-5 beta-cholestan-26-oic acid by X-ray crystallographic analysis of its ethyl ester. PMID- 10705476 TI - The inclusion compounds of beta-cyclodextrin with 4-substituted benzoic acid and benzaldehyde drugs studied by proton nuclear magnetic resonance spectroscopy. AB - Inclusion compounds of some 4-substituted benzoic acid and benzaldehyde drugs with beta-cyclodextrin were prepared and characterized by IR spectroscopy, powder X-ray diffraction, thermogravimetry, and 1H-NMR spectroscopy. The thermal stability and chemical stability of these drugs were strikingly improved after inclusion. The effect of inclusion on the chemical-shifts of protons H-3 and H-5 in the NMR spectroscopy is discussed. Using the relative shift theory, the preferred inclusion mode was proposed. The center of the aromatic ring of the drug molecule was considered to be located in the cavity 1.2 A inside from the H 5 plane of beta-cyclodextrin. PMID- 10705477 TI - Structure-activity relationships of a new antifungal imidazole, AFK-108, and related compounds. AB - Fungicidal activity of widely used imidazole antifungal drugs in topical applications is not so strong in spite of their fungistatic activities against dermatophytes and pathogenic yeasts. In order to improve fungicidal activity of imidazole antifungal agents, a series of novel imidazole derivatives having a hydrophobic substituent derived from isoprenoid were synthesized. The efficacy of these compounds was evaluated with respect to direct cell-membrane damaging activity, ergosterol biosynthesis inhibition, minimum growth-inhibitory concentration (MIC) and therapeutic effect for experimental dermatophytosis of guinea pigs. Among the newly synthesized compounds, the geranyl derivative named AFK-108 (2a) showed the highest in vivo fungicidal activity with both cell membrane damaging activity and ergosterol biosynthesis inhibition in vitro. PMID- 10705478 TI - Chemical constituents of Glycosmis citrifolia (Willd.) Lindl. Structures of four new acridones and three new quinolone alkaloids. AB - The structures of two new dimeric acridone alkaloids, glycobismine-D (1) and -E (2), having a novel linkage as binary acridones, three monomeric acridones, glycocitrine-IV (3), -V (4), and -VI (5), and three quinolone alkaloids, glycocitlone-A (6), -B (7), and -C (8) from Glycosmis citrifolia (Willd.) Lindl. (Rutaceae) have been elucidated by spectrometric studies. PMID- 10705479 TI - Copper(II)-catalyzed oxidation of d-alpha-tocopherol by oxygen in aqueous solution. AB - alpha-Tocopherol (alpha-Toc) was solubilized in aqueous solutions using 13 solubilizing agents and the products of oxidation by oxygen in the presence and the absence of Cu(II) were analyzed by HPLC. In the presence of Cu(II), the oxidation was accelerated and 5-formyl-7,8-dimethyltocol and alpha-tocoquinone were the major oxidation products. Their yields greatly increased in the presence of Cu(II). The yields and the rates of formation of the products were dependent on the properties of solubilizing agents and other conditions as well as the presence of Cu(II) or other metal ions. It is suggested that slight changes in the structure of the solubilizing agents affect the course of the reaction. PMID- 10705480 TI - Three new sesquiterpenoid glucosides of Ficus pumila fruit. AB - As the glycosyl constituents of Ficus pumila L. fruits (Moraceae), three new sesquiterpenoid glucosides, pumilasides A, B and C were isolated together with benzyl beta-D-glucopyranoside, (E)-2-methyl-2-butenyl beta-D-glucopyranoside and rutin. Their structures were characterized as (1S,4S,5R,6R,7S,10S)-1,4,6 trihydroxyeudesmane 6-O-beta-D-glucopyranoside, (1S,4S,5S,6R,7R,10S)-1,4 dihydroxymaaliane 1-O-beta-D-glucopyranoside and 10 alpha, 11-dihydroxycadin-4 ene 11-O-beta-D-glucopyranoside by spectral and chemical methods. PMID- 10705481 TI - A new concise synthesis of arcyriacyanin A and its unique inhibitory activity against a panel of human cancer cell line. AB - The nucleophilic addition reaction of N-tosyl-4-oxo-4,5,6,7-tetrahydroindole (12) with the lithium salt of 1-methoxyindole (5), followed by dehydration with 2,3 dichloro-5,6-dicyano-1,4-benzoquinone (DDQ) gave the derivative of 2,4'-bi-1H indole (9) which provides a new concise synthetic method of an indole pigment of the slime mould, arcyriacyanin A. The compound was first demonstrated here to have unique inhibitory activity to a panel of human cancer cell lines and to inhibit protein kinase C and protein tyrosine kinase. PMID- 10705482 TI - Acridone alkaloids from the root bark of Severinia buxifolia in Hainan. AB - Eight new acridone alkaloids, buxifoliadines-A--H together with nine known acridone compounds, were isolated and characterized from the root bark of Severinia buxifolia which was collected in Hainan province, China. Their structures were determined by spectroscopic methods. The relationship between acridone alkaloids with collecting area is discussed. The 13C-NMR spectra of the prenyl substituents at C-2 and/or C-4 of N-unsubstituted acridone alkaloids are also discussed. PMID- 10705483 TI - A chiral synthesis of four stereoisomers of 1,3-dimethyl-1,2,3,4 tetrahydroisoquinoline, an inducer of Parkinson-like syndrome. AB - Four stereoisomers of 1,3-dimethyl-1,2,3,4-tetrahydroisoquinoline, an inducer of Parkinson-like syndrome, were synthesized by applying a new method of 1,2,3,4 tetrahydroisoquinoline (TIQ) synthesis utilizing the Pummerer reaction as a key step. The chiral centers at C-1 and C-3 were constructed by two routes starting from alaninol (3) and 1-phenylethylamine (4) as a chiral source. Enantiomerically pure 1,3-dimethyl-TIQs (1R,3S)-(2) [corrected], (1S,3R)-(ent-2) [corrected], (1S,3S)-(1) [corrected], and (1R,3R)-(ent-1) [corrected] were prepared in a stereochemically unambiguous manner from 3 in 11 steps (route I) and from 4 in 6 steps (route II). The conformations of tetrahydroisoquinoline ring in 1-methyl, 3 methyl, and 1,3-dimethyl-TIQs were discussed on the basis of their CD, 1H-NMR spectra, and steric energies. PMID- 10705484 TI - Steroidal glycosides from the aerial part of Asclepias incarnata L. II. AB - Thirty new steroidal glycosides were obtained from the aerial part of Asclepias incarnata L. (Asclepiadaceae). These glycosides were confirmed to have lineolon, isolineolon, 12-O-acetyllineolon, 12-O-(Z)-cinnamoyllineolon, metaplexigenin, 15 beta-hydroxylineolon, 15 beta-hydroxyisolineolon, 16 alpha-hydroxyisolineolon, 12 O-tigloyl-16 alpha-hydroxyisolineolon as the aglycone and 2,6-dideoxyhexopyranose as the sugar sequence by spectroscopic methods and chemical evidence. PMID- 10705485 TI - Total syntheses of novel cytocidal beta-carboline alkaloids, oxopropalines D and G. AB - A new type of beta-carboline nucleus, N-methoxymethyl-4-methyl-beta-carboline (4) was synthesized by thermal electrocyclic reaction of a 1-azahexatriene system, involving the indole 2,3-bond. The key compound N-methoxymethyl-1-methoxycarbonyl 4-methyl-beta-carboline (2) was then prepared in a four-step sequence. The total synthesis of oxopropaline G (1e) was achieved from this key compound in four steps. Furthermore, the enantioselective total syntheses of (+)-oxopropaline D (1c) and its enantiomer were also achieved by application of the Sharpless oxidation-procedure in nine steps from 2. PMID- 10705486 TI - The Dakin-West reaction of N-alkoxycarbonyl-N-alkyl-alpha-amino acids employing trifluoroacetic anhydride. AB - The Dakin-West reaction of N-alkoxycarbonyl-N-alkyl-alpha-amino acids (1a-j) with trifluoroacetic anhydride in the presence of pyridine gave alpha-amido trifluoromethyl ketones (2a-j), in which probable intermediates were mesoionic 1,3-oxazolium-5-olates (munchnones). The diastereoselective reduction of 2a-f with NaBH4 gave the threo-aminoalcohols (5a-f), which may be explained by the Felkin-Anh model. This was confirmed by converting 5a-f into trans-5 trifluoromethyl-2-oxazolidinones (6a-f) in good yields. PMID- 10705487 TI - Iron(III)picolinate-induced oxygenation and subsequent rearrangement of triterpenoid derivatives with hydrogen peroxide. AB - The reaction of oleanane triterpenoid 1b with a FeIII(PA; picolinate)3/H2O2/MeCN system (reagent system A), a simple model system for mono-oxygenase, gave the 11 alpha-hydroxyl derivative 3 as major product, along with 11-oxo derivative 4 and 12-oxo derivative 6. The reaction of lupane triterpenoid 2b with reagent system A gave only oxidative rearrangement compounds, (20R)-aldehyde 8 and (20S)-aldehyde 9 were epimeric isomers. Then, we have found that iron(III) picolinate complex, FeIII(PA)3 is efficient in effecting the rearrangement of triterpenoid epoxides 5 and 7 into the corresponding carbonyl compounds, 6, 8 and 9 with 1,2-shift of the hydride. PMID- 10705488 TI - A short-step synthesis of orally active carbapenem antibiotic CS-834. AB - An orally bioavailable carbapenem CS-834, which is a pivaloyloxymethyl (POM) ester-type prodrug and has (R)-5-oxopyrrolidin-3-ylthio moiety at the C-2 position of the 1 beta-methylcarbapenem skeleton, is currently under clinical trial. We accomplished a short-step synthesis of CS-834 by using phosphorus ylide from the intramolecular Wittig-type reaction in the key step for cyclization to the bicyclic carbapenem system. The POM ester group was found to be suitable for the cyclization conditions. PMID- 10705489 TI - Synthesis and antiinflammatory activity of 7-methanesulfonylamino-6 phenoxychromones. Antiarthritic effect of the 3-formylamino compound (T-614) in chronic inflammatory disease models. AB - A group of derivatives of 7-methanesulfonylamino-6-phenoxychromone (1) at the pyrone and phenoxy rings was synthesized starting with 4-chloro-3-nitroanisole and evaluated against acute and chronic inflammations in oral administration in animals. Significant potency in the rat models of carrageenin-induced edema (CPE) and adjuvant-induced arthritis (AA) was realized with 2'-fluoro and 2',4' difluoro derivatives (9a and 9d), and 3-formylamino derivative (19a) and its 2' fluoro and 2',4'-difluoro compounds (22a and 22d), displaying AA therapeutic effect of ED40 = 2.5-7.1 mg/kg/d for 7 d and AA prophylactic effect of 53-70% inhibition at the dosage of 3 mg/kg/d for 22 d. To identify a candidate for further pharmacological study, the five compounds were subjected to evaluation of their gastro-ulcerogenic liability, leading to selection of the fluorine-free compound 19a which did not cause acute ulceration at 300 mg/kg in oral administration in rats. Compound 19a (ED40 = 3.6 mg/kg in established AA) possessed good therapeutic efficacy against type II collagen-induced arthritis in DBA/1J mice with doses of 30 and 100 mg/kg, suggesting the development of 19a (designated T-614) as a prospective disease-modifying antirheumatic agent. In addition, a preparative-scale synthetic route to T-614 has been established. PMID- 10705490 TI - Studies on the number of contacts between ibuprofen and ethenzamide using thermal analysis. AB - We studied a method of estimating the number of contacts in a solid dosage form using thermal analysis. Ibuprofen (IB) and Ethenzamide (ET) were used as model actives. IB and ET were granulated and sieved with each other. We prepared mixtures of IB and ET using different diameter granules. The number of contacts between the samples was calculated by the expression method of Ouchiyama and Tanaka. By thermal analysis, we measured the quantity of endotherm up to 60-63 degrees C from 56 degrees C. The quantity of endotherm up to 61 degrees C from 56 degrees C was in good proportion to the number of contacts calculated by the expression. PMID- 10705491 TI - Isolation of iso-grayanotoxin II from leaves of Leucothoe grayana Max. Its X-ray crystallographic analysis and acute toxicity in mice. AB - The structure of iso-grayanotoxin II, a new diterpenoid from Leucothoe grayana MAX., has been determined as 3 beta,5 beta,6 beta,14 beta,16 alpha pentahydroxygrayanotox-9(10)-ene by spectroscopic and X-ray crystallographic analysis. The lethal dosage level of iso-grayanotoxin II in mice was lower than that of grayanotoxin III. PMID- 10705492 TI - Direct lithiation and alkylation of trifluoromethyl enol ethers having a beta sulfur substituent. AB - The direct lithiation and alkylation of beta-sulfur-alpha-trifluoromethyl substituted enol ethers 1-3 proceeded to give the alkylated products 4a-f, 5, 6a c in moderate to high yields. PMID- 10705493 TI - A new diterpene glycoside from Rabdosia rubescens. AB - A new ent-kaurene beta-D-glucoside was isolated from Rabdosia rubescens, together with the known compounds oridonin, ponicidin, and pedalitin. The structure of new compound was determined, on the basis of spectral data and X-ray crystallographic analysis, to be ent-7 beta,20-epoxy-kaur-16-ene-1 beta,6 alpha,7 alpha,14 alpha,15 alpha-pentanol 1-O-beta-D-glucopyranoside. PMID- 10705494 TI - Design, synthesis and biological activity of 7-O-(4-O-acetyl-3-iodo-2,3,6 trideoxy-alpha-L-arabino-hexopyranosyl) daunomycinone and 7-O-(3-chloro-2,3,6 trideoxy-4-O-propanoyl-alpha-L-lyxo- hexopyranosyl)daunomycinone. AB - The synthesis and pharmacological evaluation of 7-O-(4-O-acetyl-3-iodo-2,3,6 trideoxy-alpha-L-arabino-hexopyranosyl) daunomycinone and 7-O-(3-chloro-2,3,6 trideoxy-4-O-propanoyl-alpha-L-lyxo-hexopyrano syl) daunomycinone are described. Their cytotoxic activity was evaluated against normal and resistant cell lines. Both compounds exhibited activity against the adriamycin resistant cell line KB A1. These results support the hypothesis that the increased lipophilicity of the sugar part of anthracyclines is associated with their ability to overcome multidrug resistance (MDR). PMID- 10705495 TI - Marstomentosides O-T polyoxypregnane glycosides from Marsdenia tomentosa. AB - Thirteen pregnane glycosides were isolated from fresh leaves of Marsdenia tomentosa collected in the Fukuyama district. Of these, six were glycosides previously obtained from the same plant collected in the Fukuoka district and one from another Asclepiadaceous plant. The structures of the six new glycosides were determined by spectrometric method. PMID- 10705496 TI - Constituents of holothuroidea.9. Isolation and structure of a new ganglioside molecular species from the sea cucumber Holothuria pervicax. AB - A new ganglioside molecular species, HPG-7 (1) was obtained from the polar fraction of the chloroform/methanol extract of the sea cucumber, Holothuria pervicax. On the basis of chemical and spectroscopic evidence, the structure of 1 was determined, and the major component was 1-O-[alpha-L-fucopyranosyl-(1-->4)-(N acetyl-alpha-D-neuraminosyl) - (2-->11)-(N-glycolyl-alpha-D-neuraminosyl)-(2-->4) (N-acetyl -alpha-D- neuraminosyl)-(2-->6)-beta-D-glucopyranosyl]-(2S,3S,4R)-2 [(2R)-2- hydroxytetracosanoylamino]-14-methyl-hexadecane-1,3,4-triol. This is the first report on the isolation and structure elucidation of trisialo-ganglioside from sea cucumber. 1 showed neuritogenic activity toward the rat pheochromocytoma cell line, PC-12 cell. PMID- 10705497 TI - Reaction of 3-acetonyl-5-cyano-1,2,4-thiadiazole with phenylhydrazine hydrochlorides: indolization and phenylpyrazolation. AB - Treatment of 3-acetonyl-5-cyano-1,2,4-thiadiazole (1) with 4-methyl or 4 methoxyphenylhydrazine hydrochloride provided 5-cyano-3-(2,5-dimethylindol-3-yl) 1,2,4-thiadiazole (2) or 5-cyano-3-(5-methoxy-2-methylindol-3-yl)-1,2,4 thiadiazole (3) as the sole product, respectively. In contrast, treatment of 1 with phenylhydrazine hydrochloride resulted in the formation of 5-cyano-3-(2 methylindol-3-yl)-1,2,4-thiadiazole (4) and the unexpected 5-cyano-3-(3,5 dimethyl-1-phenylpyrazol-4-yl)-1,2,4-thiadiazole (5). In a similar manner, when 1 was treated with 4-chlorophenylhydrazine hydrochloride, indolization was suppressed by phenylpyrazolation giving rise to 5-cyano-3-(5-chloro-2-methylindol 3-yl)-1,2,4-thiadiazole (6) and 5-cyano-3-[1-(4-chlorophenyl)-3,5-dimethylpyrazol 4-yl]-1,2,4-thia diazole (7). The reaction mechanism is discussed. Compounds 4, 5 and 6 exhibited weak antimicrobial activity against Helicobacter pylori. PMID- 10705498 TI - A facile synthesis of 2-deoxy-2,3-didehydroneuraminic acid derivatives. AB - The 2-thio- or 2-selenoglycosides of N-acetylneuraminic acid methyl ester were transformed by successive treatment with dimethyl(methylthio)sulfonium triflate (DMTST) and 1,8-diazabicyclo[5.4.0]-7-undecene (DBU) to give the corresponding methyl 2-deoxy-2,3-didehydroneuraminates in excellent yields. Their acids and their analogues are sialidase inhibitors of pharmaceutical interest. PMID- 10705499 TI - A study of drug-carrier interactions in dry powder inhaler formulations using the Andersen cascade impactor, X-ray microanalysis and time of flight aerosol beam spectrometry (TOFABS). AB - The purpose of this study was to determine the in vitro deposition of both drug (albuterol sulfate) and carrier (lactose) particles in relation to each other from a dry powder inhaler formulation using an Andersen cascade impactor (ACI) and time of flight aerosol beam spectrometry (TOFABS). In addition, scanning electron microscopy (SEM) combined with X-ray microanalysis was employed to distinguish albuterol sulfate from lactose. Drug particles apparently penetrated deeper into the impactor than lactose particles contained in the formulation. In some certain stages of impactor, drug particles were separated from lactose particles. Although the TOFABS cannot distinguish between albuterol sulfate and lactose, the TOF spectra obtained from the Aerosizer would appear to be partly indicative of the interactions which exist between drug and carrier. One symmetrical TOF peak was obtained from drug or lactose alone. The TOF peak of the drug was always lower than the TOF of lactose. The times obtained for each powder between experiments were highly reproducible and typical of material and particle size. The use of SEM-X-ray microanalysis also allowed some qualitative characterization of shape and state of association of the two components. PMID- 10705500 TI - Multiple solubility maxima of oxolinic acid in mixed solvents and a new extension of Hildebrand solubility approach. AB - A new extension of the Hildebrand solubility approach which describes drug solubility in solvent mixtures showing multiple solubility peaks, the chameleonic effect, is proposed. The experimental solubilities of oxolinic acid were measured at 25 degrees C in solvent mixtures of ethanol-water and ethanol-ethyl acetate. A plot of the mole fraction of the drug against the solubility parameter (delta) of the solvent mixtures displays two peaks at delta = 30.78 MPa1/2 (80% v/v of ethanol in water) and at delta = 20.90 MPa1/2 (30% v/v of ethanol in ethyl acetate). The new extension proposed reproduces two solubility peaks. The thermograms of the solid phase before and after equilibration with the solvent mixtures did not show significant changes. The new extension was also tested with experimental data previously reported for drugs showing two solubility peaks of different height. The accuracy of other published models for describing two solubility maxima is also compared. PMID- 10705501 TI - Partial solubility parameters of lactose, mannitol and saccharose using the modified extended Hansen method and evaporation light scattering detection. AB - The modified extended Hansen method was tested for the first time to determine partial solubility parameters of non-polymeric pharmaceutical excipients. The method was formerly tested with drug molecules, and is based upon a regression analysis of the logarithm of the mole fraction solubility of the solute against the partial solubility parameters of a series of solvents of different chemical classes. Two monosaccharides and one disaccharide (lactose monohydrate, saccharose and mannitol) were chosen. The solubility of these compounds was determined in a series of solvents ranging from nonpolar to polar and covering a wide range of the solubility parameter scale. Sugars do not absorb at the UV-vis region, and the saturated solutions were assayed with a recent chromatographic technique coupled to an evaporative light scattering detector. This technique was suitable to determine the concentration dissolved in most solvents. The modified extended Hansen method provided better results than the original approach. The best model was the four parameter equation, which includes the dispersion delta d, dipolar delta p, acidic delta a and basic delta b partial solubility parameters. The partial solubility parameters obtained, expressed as MPa1/2, were delta d = 17.6, delta p = 28.7, delta h = 19, delta a = 14.5, delta b = 12.4, delta T = 32.8 for lactose, delta d = 16.2, delta p = 24.5, delta h = 14.6, delta a = 8.7, delta b = 12.2, delta T = 32.8 for mannitol and delta d = 17.1, delta p = 18.5, delta h = 13, delta a = 11.3, delta b = 7.6, delta T = 28.4 for saccharose. The high total solubility parameters delta T obtained agree with the polar nature of the sugars. The dispersion parameters delta d are quite similar for the three sugars indicating that the polar delta p and hydrogen bonding parameters (delta h, delta a, delta b) are responsible for the variation in the total solubility parameters delta T obtained, as also found for drugs. The results suggest that the method could be extended to determine the partial solubility parameters of other non-polymeric pharmaceutical excipients. PMID- 10705502 TI - Development of plasmin-selective inhibitors and studies of their structure activity relationship. AB - Various compounds were synthesized by combining three components at positions P1, P1' and P2'. Of these, N-(trans-4-aminomethylcyclohexanecarbonyl)-Tyr(O-2 bromobenzylo xycarbonyl)- octylamide inhibited plasmin selectively with IC50 values of 0.80 and 0.23 microM towards S-2251 and fibrin, respectively. This compound also inhibited plasma kallikrein, urokinase, thrombin and trypsin with IC50 values of 10, > 50, > 50 and 1.6 microM, respectively. PMID- 10705503 TI - Inhibition of human immunodeficiency virus type 1 reverse transcriptase and ribonuclease H activities by constituents of Juglans mandshurica. AB - From the stem-bark of Juglans mandshurica, two new naphthalenyl glucopyranosides, 1,4,8-trihydroxynaphthalene 1-O-[alpha-L-arabinofuranosyl-(1-->6)-beta-D glucopyranoside] (1) and 1,4,8-trihydroxynaphthalene 1-O-beta-D-[6'-O-(3",5" dihydroxy-4"-methoxybenzoyl)]glucopyranosi de (4), and two new alpha-tetralonyl glucopyranosides, 4 alpha,5,8-trihydroxy-alpha-tetralone 5-O-beta-D-[6'-O-(3",5" dihydroxy-4"-methoxybenzoyl)]glucopyranosi de (7) and 4 alpha,5,8-trihydroxy alpha-tetralone 5-O-beta-D-[6'-O-(3",4",5"-trihydroxybenzoyl)]glucopyranoside (8), were isolated together with three known naphthalenyl glucopyranosides (2, 3 and 5), one alpha-tetralonyl glucopyranoside (6), four flavonoids (9-12), and two galloyl glucopyranosides (13, 14). Amongst the isolated compounds, 1,2,6 trigalloylglucopyranose (13) and 1,2,3,6-tertagalloylglucopyranose (14) exhibited the most potent inhibition of reverse transcriptase (RT) activity with IC50 values of 0.067 and 0.040 microM, respectively, while the latter compound also inhibited ribonuclease H (RNase H) activity with an IC50 of 39 microM, comparable in potency to illimaquinone used as a positive control. 1,4,8-Trihydroxy naphthalene 1-O-beta-D-glucopyranoside (2), 1,4,8-trihydroxynaphthalene 1-O-beta D-[6'-O-(4"-hydroxy-3",5"-dimethoxybenzoyl)]glucopyranoside (3) and 8 showed moderate inhibition against both enzyme activities, and inhibitory potency of 2 against RNase H activity (IC50 = 156 microM) was slightly greater than that against the RT activity (IC50 = 290 microM). The inhibitory potencies of 4 alpha,5,8-trihydroxy-alpha-tetralone 5-O-beta-D-[6'-O-(4"-hydroxy-3",5" dimethoxybenzoyl)] glucopyranoside (6), 7 and 8 against RT activity increased accompanied by an increase in the number of free hydroxyls on the galloyl residues, as represented by the IC50 values of > 500, 330 and 5.8 microM, respectively. PMID- 10705504 TI - New monoterpene glycoside esters and phenolic constituents of Paeoniae radix, and increase of water solubility of proanthocyanidins in the presence of paeoniflorin. AB - Seven new monoterpene glycoside esters related to paeoniflorin were isolated from Paeoniae Radix, together with polymeric proanthocyanidins, polygalloylglucoses and 48 known compounds (a benzoylsucrose, seven aromatic acids, adenosine, nine monoterpene glycosides, eight flavan-3-ols, a catechin dimer formed by oxidation, seven proanthocyanidins, three galloylsucroses, five galloylglucoses, and six ellagitannins). The structures of the new compounds were determined by spectral investigation including two-dimensional NMR techniques. In addition, increased water solubility of polymeric proanthocyanidin in the presence of paeoniflorin was examined by n-octanol-water partition and 1H-NMR spectral experiments. PMID- 10705505 TI - Determination of hydration kinetics of sulfaguanidine anhydrate in aqueous solution by calorimetry. AB - A heat conduction microcalorimeter was used to evaluate the isothermal transition in water from anhydrate to monohydrate at 298 K. Sulfaguanidine (SGN) anhydrate was used as a model compound for the measurement of hydration kinetics in water. It is the well-known that SGN is very slightly soluble in water and capable of existing as the anhydrate or monohydrate form in the solid state. The transition rates of SGN anhydrate to monohydrate in tablets and granules were investigated. The hydration kinetics of tablets with controlled surface areas, obtained by coating the side with paraffin in aqueous solution, followed an apparent zero order mechanism. On the other hand, the transition mechanism of the granules involved a phase boundary-controlled contracting interface reaction. PMID- 10705506 TI - Reactions of N-hydroxysuccinimide esters of anthranilic acids with anions of beta keto esters. A new route to 4-oxo-3-quinolinecarboxylic acid derivatives. AB - A new approach for the synthesis of 4-oxo-3-quinolinecarboxylic acid derivatives is described. This methodology involves the C-acylation of the anions of appropriate beta-keto esters with novel N-hydroxysuccinimide esters of anthranilic acids. The intermediate C-acylation products 3 are spontaneously cyclized to afford 3-ethoxycarbonyl-4-oxoquinoline derivatives 4. The introduction of a variety of substituents at positions 1 and 2 of the quinoline ring is feasible with the selection of suitable anthranilic acids and beta-keto esters. The structure of the obtained 2-substituted 3-ethoxycarbonyl-4 oxoquinolines was confirmed by IR and NMR spectral data. PMID- 10705507 TI - Synthesis and pharmacokinetics of 1 alpha-hydroxyvitamin D3 tritiated at 22 and 23 positions showing high specific radioactivity. AB - A novel synthesis of a radioactive compound of 1 alpha-hydroxyvitamin D3 (1 alpha OHD3) (1) and its pharmacokinetics are described. Radioactive 1 alpha OHD3 tritiated at 22 and 23 positions ([22,23-(3)H4]1 alpha OHD3) (5) was prepared via key reactions of the reduction of acetylenic side chain in the ketone (12) with tritium gas in the presence of palladium-charcoal and the subsequent Wittig reaction with the A-ring synthon (16). [22,23-(3)H4]1 alpha OHD3 (5) showed high specific radioactivity (111.5 Ci/mmol) and was used successfully in pharmacokinetics studies with rats. In the pharmacokinetics studies, the plasma concentration level of the active form of vitamin D3, 1 alpha,25-dihydroxy vitamin D3 [1 alpha,25(OH)2D3], after oral or intravenous administration of [22,23-(3)H4]1 alpha OHD3 (5), showed longer half-life, lower maximum concentration, and lower area under the curve than those after treatment of 1 alpha,25(OH)2D3 tritiated at 26 and 27 positions (4). These results might suggest a beneficial therapeutic utility of 1 alpha OHD3 (1) over the treatment of 1 alpha,25(OH)2D3 (2). PMID- 10705508 TI - Fluorinative Beckmann fragmentation: fluorinative alpha-cleavage of cyclic ketoximes by diethylaminosulfur trifluoride. AB - Diethylaminosulfur trifluoride reacted with cyclic ketoximes bearing substituent(s) that can stabilize a carbocation to cause fluorinative fragmentation, affording fluorinated carbonitrile. Ketoximes lacking such substituents afforded complex mixtures. However, the introduction of a sulfur functionality, which can stabilize a carbocation and can be easily removed from the reaction products, into the ketoxime was effective for producing the fluorinative fragmentation. PMID- 10705509 TI - Mechanism of superoxide dismutase-like activity of Fe(II) and Fe(III) complexes of tetrakis-N,N,N',N'(2-pyridylmethyl)ethylenediamine. AB - The superoxide dismutase (SOD) activity of iron(II) tetrakis-N,N,N',N'(2 pyridylmethyl)ethylenediamine complex (Fe-TPEN) was reexamined using a pulse radiolysis method. In our previous study (J. Biol. Chem., 264, 9243-9249 (1989)), we reported that this complex has a potent SOD activity in a cyt. c (cytochrome c)-based system (IC50 = 0.8 microM) and protects E. coli cells against paraquat toxicity. The present pulse radiolysis experiment revealed that Fe(II)TPEN reacts stoichiometrically with superoxide to form Fe(III)TPEN with a second-order rate constant of 3.9 x 10(6) M-1 S-1 at pH 7.1, but superoxide did not reduce Fe(III)TPEN to Fe(II)TPEN. The reaction of Fe(III)TPEN and superoxide was biphasic. In the fast reaction, an adduct (Fe(III)TPEN-superoxide complex) was formed at the second-order rate constant of 8.5 x 10(5) M-1 S-1 at pH 7.4. In the slow one, the adduct reacted with another molecule of the adduct, regenerating Fe(III)TPEN. In the cyt. c method with catalase, this Fe(III)TPEN-superoxide complex showed cyt. c oxidation activity, which had led to overestimation of its SOD activity. Based on the titration data, the main species of complex in aqueous media at neutral pH was indicated to be Fe(III)TPEN(OH-). A spectral change after the reduction with hydrated electron indicates that the OH- ion coordinates directly to Fe(III) by displacing one of the pyridine rings. The X-ray analysis of [Fe(II)TPEN]SO4 supported this structure. From the above results we propose a novel reaction mechanism of FeTPEN and superoxide which resembles a proton catalyzed dismuting process, involving Fe(III)TPEN-superoxide complex. PMID- 10705510 TI - Molecular state of chlorpheniramine in resinates. AB - Chlorpheniramine (CPM) maleate was prepared as a series of resinates by the batch method. The several resinates were investigated by differential scanning calorimetry (DSC), X-ray powder diffraction (XRPD) and infrared (IR) spectrometry. The results from DSC and XRPD showed that the molecular state of the entrapped drug changed from the crystalline to amorphous state. IR spectra indicated that only CPM species was entrapped in the resinates. Moreover, it also showed that the positively charged amine group of the drug interacted with the sulfonate groups of the resin by ionic association. The dissolution of the drug and resinates was also studied where it was found that the dissolution of the resinates was retarded by their crosslinked structure and markedly affected by the quantity of resin. PMID- 10705511 TI - Mechanism of antioxidative activity of fluvastatin-determination of the active position. AB - In order to clarify the mechanism of action for the antioxidative activity of fluvastatin sodium (FLV, (+/-)-sodium (3RS, 5RS, 6E)-7-[3-(4-fluorophenyl)-1-(1 methylethyl)-1H-indol-2-yl]-3, 5-dihydroxy-6-heptanoate) and its derivatives, reaction of the corresponding methyl ester of FLV with di-tert-butyl diperoxyoxalate was examined, and the corresponding keto derivative was isolated from the reaction mixture. On the basis of this result, it was concluded that the active site is the allylic carbon conjugated with the indole ring. PMID- 10705512 TI - A facilitated cyclic ether formation and its potential application in solid-phase peptide and organic synthesis. AB - A "trimethyl lock" system has been known to facilitate lactonization reactions through what has been termed a stereopopulation control mechanism. We have found that a similar trimethyl lock system can also facilitate cyclic ether formation with the concomitant release of a carboxylic acid in the presence of anhydrous tetrabutylammonium fluoride. To study this base-mediated trimethyl lock facilitated cyclic ether formation, we synthesized fifteen model compounds. All model compounds underwent base-mediated cyclic ether formation in high yields at 0 degree C to room temperature (r.t.) with the concomitant release of the attached carboxylate. Such a system potentially could be used for the development of a two-dimensional linker for solid phase peptide and organic synthesis. PMID- 10705513 TI - [2-(omega-phenylalkyl)phenoxy]alkylamines.II: Synthesis and selective serotonin-2 receptor binding. AB - A series of [2-(omega-phenylalkyl)phenoxy]alkylamines was synthesized and their receptor binding affinity was examined in vitro. These compounds showed an affinity for serotonin-2 (5-HT2) and dopamine-2 (D2) receptors. [2-(2 phenylethyl)phenoxy]alkylamine derivatives with a pyrrolidine or piperidine moiety in the structure showed higher affinity for 5-HT2 receptors but lower affinity for D2 receptors. Among these compounds, (S)-2-[2- [2-[2-(3 methoxyphenyl)ethyl]phenoxy]ethyl]-1-methylpyrrolidine, (S)-27, exhibited the most potent and selective affinity for 5-HT2 receptors. Furthermore, (S)-27 was effective in inhibiting 5-HT-induced vasoconstriction in vitro and platelet aggregation both in vitro and ex vivo. PMID- 10705514 TI - Structural features for fluorescing present in methoxycoumarin derivatives. AB - Structural features of fluorescent methoxycoumarins were examined from the viewpoint of substituent effect and ring structure in connection with intramolecular charge-transfer (ICT). The fluorescence of methoxycoumarins depended primarily upon the ICT from a C6-electron-donating group to the substituents at the C3-position of the coumarin ring. Furthermore, the presence of a lactone ring itself, including a carbonyl group, cyclic ether oxygen and ethylenic bond as partial ring structures, was found to be essential for fluorescing in methoxycoumarins according to the fluorescent behaviors of chemically deformed model compounds. PMID- 10705515 TI - Scavengers for peroxynitrite: inhibition of tyrosine nitration and oxidation with tryptamine derivatives, alpha-lipoic acid and synthetic compounds. AB - The inhibitory effects of various endogenous and synthetic compounds on the nitration and oxidation of L-tyrosine by peroxynitrite were examined. Nitrating and oxidizing activities were monitored by the formation of 3-nitrotyrosine and dityrosine with a HPLC-UV-fluorescence detector system, respectively. Glutathione, serotonin and synthetic sulfur- and selenium-containing compounds inhibited both the nitration and oxidation reaction of L-tyrosine effectively. However, 5-methoxytryptamine, melatonin and alpha-lipoic acid only inhibited the nitration reaction, and enhanced the formation of an oxidation product. This is important evidence that there are different intermediates in the nitrating and oxidizing reactions of L-tyrosine by peroxynitrite. It was suggested that 5 methoxytryptamine, melatonin and alpha-lipoic acid reacted only with the nitrating intermediate of peroxynitrite and inhibited nitration of L-tyrosine. Actually, the DNA strand breakage, which is believed to be a typical reaction of hydroxyl radical-like species, caused by peroxynitrite was not effectively inhibited by 5-methoxytryptamine. 5-Methoxytryptamine, melatonin and alpha-lipoic acid were viewed as useful reagents for investigating the mechanisms of damage by peroxynitrite in vitro. PMID- 10705516 TI - Synthesis and structure studies of complexes of some second row transition metals with 1-(phenylacetyl and phenoxyacetyl)-4-phenyl-3-thiosemicarbazide. AB - The synthesis of the new complexes of 1-phenylacetyl-4-phenyl-3-thiosemicarbazide (H2papts) and 1-phenoxyacetyl-4-phenyl-3-thiosemicarbazide (H2Pxapts); [Ru(HL)2(H2O)2], [Rh(HL)3], [Ag(H2L)(H2O)2](NO3), trans [UO2(HL)(bipy)(AcO)(H2O)2] (H2L = H2papts, H2pxapts; bipy = 2,2'-bipyridyl), [Ag(H2papts)(bipy)]+ and [Pd-(Hpapts)(bipy)]+ is described. Characterization of these complexes by IR, electronic and 1H-NMR spectra, conductometric titrations and thermal analysis is included. The complexes [Ru(HL)2(H2O)2] were found to be efficient catalysts for the oxidation of primary alcohols to aldehydes and acids, secondary alcohols to ketones and aryl halides to aldehydes and acids in the presence of NaIO4 as co-oxidant. PMID- 10705517 TI - Enantioselective synthesis of (2R,4'R,8'R)-alpha-tocopherol (vitamin E) based on enzymatic function. AB - Syntheses of (S)-chroman-2-carboxaldehyde congener 1 and (S)-chiral isoprene unit 3 were achieved based on the enzymatic acetylation of (+/-)-chroman-2-methanol 6 and (+/-)-(2,3)-anti-2-methyl-3-(p-methoxyphenyl)-1,3-propane diol 12, respectively. Synthesis of the side-chain part corresponding to (3R,7R)-3,7,11 trimethyldodecan-1-ol 27 was achieved by the coupling reaction of (S)-3 and (R) 3,7-dimethyloctyl iodide 4. The Wittig reaction of (3R,7R)-phosphonium salt 2 derived from (3R,7R)-27 and (S)-1 gave the olefin 28 which was subjected to catalytic hydrogenation to afford (2R,4'R,8'R)-alpha-tocopherol. PMID- 10705518 TI - Synthesis and pharmacological activity of O-(5-isoxazolyl)-L-serine. AB - A novel isoxazole derivative, O-(5-isoxazolyl)-L-serine (OIS, 1), was synthesized by a Mitsunobu reaction of isoxazolin-5-one (4) with N-Boc-L-serine tert-butyl ester (5) and subsequent deprotection of the coupling product. Its structure was elucidated by spectroscopic analyses. The pharmacological activity of 1 was also examined with cloned glutamate receptors and transporters using a Xenopus oocyte expressing system showing substrate activity on an excitatory amino acid carrier 1 (EAAC 1) as a glutamate transporter. PMID- 10705519 TI - Acetylated and non-acetylated flavonol triglycosides from Galega officinalis. AB - Three flavonol triglycosides kaempferol 3-[2Gal-(4 acetylrhamnosyl)robinobioside], kaempferol 3-(2Gal-rhamnosylrobinobioside) and quercetin 3-(2G-rhamnosylrutinoside) have been isolated from a methanolic extract of Galega officinalis aerial parts. They are reported for the first time in the genus Galega; moreover, the acetylated triglycoside is a new natural product. PMID- 10705520 TI - Effect of polymer excipients on the enzyme activity of lyophilized bilirubin oxidase and beta-galactosidase formulations. AB - The effects of excipients on the protein stability during lyophilization as well as the storage stability of lyophilized bilirubin oxidase (BO) and beta galactosidase (GA) formulations were studied using four polymer excipients: dextran, polyvinylalcohol (PVA), poly(acrylic acid) (PAA), and alpha, beta-poly(N hydroxyethyl)-L-aspartamide (PHEA). Denaturation of BO and GA during lyophilization largely depended on the excipient used. Dextran appeared to cause severe damage to proteins, whereas PHEA protected proteins effectively from denaturation. Storage stability of BO and GA formulations also depended on the excipients, such that the formulations containing dextran and PAA were relatively unstable. Storage stability was improved by absorption of a small amount of water for all the formulations studied. Absorption of a larger amount of water, however, decreased the storage stability of the formulations containing PVA, PAA or PHEA. In contrast, the storage stability of formulations containing dextran did not decrease noticeably with increasing water. This may be because formulations containing dextran have a higher glass transition temperature than formulations containing PVA, PAA or PHEA when a large amount of water is absorbed. PMID- 10705521 TI - A new oleanene glucuronide obtained from the aerial parts of Melilotus officinalis. AB - A new oleanene glucuronide called melilotus-saponin O2 (1) was isolated together with three known ones (soyasaponin I, astragaloside VIII, wistariasaponin D) from the aerial parts of Melilotus officinalis (L.) Pallas (Leguminosae). The structure of 1 was determined to be 3-O-alpha-L-rhamnopyranosyl-(1-->2)-beta-D xylopyranosyl- (1-->2)-beta-D-glucuronopyranosyl melilotigenin by spectroscopic and chemical methods. PMID- 10705523 TI - Cardiac glycosides from Erysimum cheiranthoides. AB - Two new cardiac glycosides called cheiranthosides VI (2) and VII (3) were isolated together with a known one, glucoerysimoside (1) from the seeds of Erysimum cheiranthoides. Based on spectroscopic data, the structures of 2 and 3 were characterized as periplogenin 3-O-beta-D-glucopyranosyl(1-->4)-beta-D fucopyranoside and periplogenin 3-O-beta-D-glucopyranosyl(1-->4)-beta-D antiaropyranoside, respectively. PMID- 10705522 TI - An efficient one-pot method for the large-quantitative production of radiopharmaceutical ligand: diazadioximes. AB - An efficient one-pot procedure for the preparation of diazadioxime was described. Treatment of ketooximes with alkyldiamine followed by NaBH4 in dry ethanol afforded the corresponding d,l-diazadioximes in 56--74% yield without isolation of the intermediates. PMID- 10705524 TI - Design, synthesis, conformational analysis and biological activities of purine based 1,2-di-substituted carbocyclic nucleosides. AB - New 1,2-di-substituted carbocyclic nucleosides with 6-chloropurine, adenine and hypoxanthine bases were synthesized by construction of purine on the primary amino group of (+/-)-trans-2-aminocyclopentylmethanol. AM1 calculations showed close correspondence between the positions of the heteroatoms in the adenine derivative and dideoxyadenosine. The most active of the new compounds in antiviral assays and antitumoral assays against L1210/0, MOLT4/C8 and CEM/0 cells was the 6-chloropurine derivative. PMID- 10705525 TI - Synthesis of peptides with alpha,beta-dehydroamino acids. XIII photoisomerization of Ac-(Z)-delta Phe-NHMe: Ac-(E)-delta Phe-NHMe. AB - Easily accessible Ac-(Z)-delta Phe-NHMe was photoisomerized to so far unknown Ac (E)-delta Phe-NHMe. Some parameters of the process leading to a diastereomeric mixture of ratio 90(Z):10(E) have been tested and the photoisomerization has been carried out on a preparative milligram scale. The isomers were separated via crystallization followed by preparative HPLC. PMID- 10705526 TI - Surface active properties of simple cyclic and heterocyclic amines in water. AB - The surface tension of aqueous solutions of simple cyclic, heterocyclic and aromatic amines was measured with a Du Nouy tensiometer at 25 degrees C and the results discussed in terms of structure-aggregation relationships. The simple compounds used in this study were piperazine, piperidine, morpholine, 3 methylpyridine, cyclohexylamine and benzylamine, with carbon numbers ranging from four to seven. Piperazine, piperidine and morpholine did not form micellar associations but cyclohexylamine, benzylamine and 3-methylpyridine did, indicating that more than six carbons are necessary to form micellar associations, at least for compounds having a six-membered ring. PMID- 10705527 TI - Two new meliacarpinins from the roots of Melia azedarach. AB - Two new azadirachtin-type limonoids, 1-methacrylyl-3-acetyl-11 methoxymeliacarpinin (1) and 1-(2-methylpropanoyl)-3-acetyl-11 methoxymeliacarpinin (2), together with the known compounds, meliacarpinin D (3), melianin B (4) and 2 beta,3 beta-dihydroxy-5 alpha-pregn-17(20)-(Z)-en-16-one (5), were isolated from the roots of Melia azedarach. The structures of 1 and 2 were elucidated by analysis of spectroscopic data and comparison of their NMR data with those of 3. Compounds 1 and 5 exhibited significant activity in the brine shrimp lethality test (BST). PMID- 10705528 TI - Facile synthesis of optically active gamma-lactones via lipase-catalyzed reaction of 4-substituted 4-hydroxybutyramides. AB - Lipase-catalyzed transesterification of racemic 4-substituted 4 hydroxybutyramides with succinic anhydride proceeded enantioselectively to afford (S)-succinic acid monoester and unreacted (R)-4-hydroxybutyramide derivative, which were separated easily by treatment with an alkaline solution. Both enantiomers were converted easily to optically active gamma-substituted gamma butyrolactones. PMID- 10705529 TI - One-pot enantioselective synthesis of optically active homoallylic alcohols from allyl halides. AB - A one-pot, convenient method for the preparation of optically active homoallylic alcohols from allyl halides was developed. Allyltrichlorosilanes were generated in situ from allyl halides and trichlorosilane in the presence of cuprous chloride and tertiary amine. Without isolation of the allyltrichlorosilanes, benzaldehyde and chiral biquinoline N,N'-dioxide were introduced into the same flask, producing the corresponding homoallylic alcohols with good to high enantioselectivities. PMID- 10705530 TI - Synthesis of peptides mimicking chemokine receptor CCR5 and their inhibitory effects against HIV-1 infection. AB - Peptides mimicking chemokine receptor CCR5 were synthesized and their anti-HIV-1 activities evaluated. Prepared compounds, especially a sulfated derivatives, showed significant anti-HIV-1 activities. Furthermore, a hybrid molecule linked to an N-carbomethoxycarbonyl-prolyl-phenylalanine (CPF) moiety had a greater effect. PMID- 10705531 TI - Synthesis of J-111,347, a novel 1 beta-methylcarbapenem with broad-spectrum antibacterial activity. AB - Synthesis of J-111,347 (1), a new 1 beta-methylcarbapenem with broad-spectrum antibacterial activity including that against methicillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa, was achieved via diastereoselective preparation of a side-chain thiol 3 from an optically active (R)-3,4-dihydroxybutanal 4. PMID- 10705532 TI - New estrogenic antagonists bearing dicarba-closo-dodecaborane as a hydrophobic pharmacophore. AB - We have designed and synthesized estrogen antagonists bearing dicarba-closo dodecaborane (carborane) as a hydrophobic pharmacophore based on the structure of 1-(4-hydroxyphenyl)-1,12-dicarba-closo-dodecaborane, a potent estrogen agonist that we reported previously. Compounds with a long alkyl chain bearing an amide moiety on the carborane skeleton (6, 7) showed estrogen antagonistic activity in a luciferase reporter gene assay using COS-1 cells transfected with a rat ER alpha-expression plasmid and as an appropriate reporter plasmid. PMID- 10705533 TI - PCDDs/DFs emissions from crematories in Japan. AB - Concentrations of PCDDs and PCDFs in emission gases from 10 crematories were measured. The relationship between PCDDs/DFs and several factors such as structure, equipment and operational state of the crematory is discussed. Furthermore, emission of PCDDs/DFs from all crematories in Japan is estimated. The following results are obtained: (1) total concentration of PCDDs/DFs was 2.2 290 ng/N m3, whose TEQ concentration was 0.0099-6.5 ng TEQ/N m3; (2) total concentration of PCDFs was higher than that of PCDDs; (3) T4CDFs was the highest in the homologue pattern and 2,3,7,8-T4CDF was the highest in the isomer pattern; (4) emission of PCDDs/DFs was the largest in the first 20 min of cremation; (5) concentration of PCDDs/DFs was related to the existence of a secondary combustion chamber and a dust collector, and the ratio of the numbers of main and secondary combustion chambers; (6) total emission of PCDDs/DFs from crematories in Japan was estimated to be 8.9 g TEQ/yr. PMID- 10705534 TI - Improved extraction procedures for polychlorinated biphenyls in solid samples with aqueous sodium hydroxide under autoclave conditions. AB - The efficacy of the extraction of polychlorinated biphenyls (PCBs) from varnish infiltrated insulating papers as a model of solid materials with an aqueous sodium hydroxide (NaOH) by autoclaving at 121 degrees C for 30 min was compared with those for the conventional procedures, i.e., saponification with 1 N ethanolic NaOH in a boiling water bath for 60 min and extraction with benzene in a Soxhlet apparatus. The newly invented autoclaving method was found to be superior to the conventional ones, yielding approximately 5- to 6-fold cumulative PCB congeners without being accompanied by extended decomposition. Therefore, the autoclave-based sample treatment is recommended for more accurate determination of PCBs in the case of PCB-impregnated solid materials such as hardened oils and resin-coated or -infiltrated papers instead of being treated conventionally. PMID- 10705535 TI - Atmospheric deposition of PCDD/Fs near an old municipal solid waste incinerator: levels in soil and vegetation. AB - The levels of polychlorinated dibenzo-p-dioxins (PCDD) and polychlorinated dibenzofurans (PCDF) were determined in soil and vegetation samples taken from 24 sites in the vicinity of an old municipal solid waste incinerator (San Adria del Besos, Barcelona, Spain). Duplicate samples were collected within a radius of 3 km from the stack. PCDD/F concentrations in soils ranged from 1.22 to 34.28 ng I TEQ/kg (d.m.) with median and mean values of 9.06 and 12.24 ng I-TEQ/kg, respectively. In turn, the levels of PCDD/Fs in vegetation samples ranged from 0.33 to 1.98 ng I-TEQ/kg (d.m.), with median and mean values of 0.58 and 0.70 ng I-TEQ/kg, respectively. Although the present PCDD/F concentrations in soil samples were higher than those recently found in soils taken near other incinerators from Catalonia, they are of the same order of magnitude than the levels of these pollutants found in incinerators from other countries. By contrast, the concentrations of PCDD/Fs in herbage samples were comparable to those found in recent surveys carried out in Catalonia. PMID- 10705536 TI - Peak identification for PCDD/Fs in environmental samples at different temperature programs. AB - A method has been developed for peak recognition of 136 polychlorinated dibenzo-p dioxins (PCDDs) and polychlorinated dibenzofurans (PCDFs) at different temperature programs. Their retention behaviours are predicted on the basis of an identification database of retention values (A, B) of gas chromatography. By the retention times of 13C labelled 2,3,7,8-substituted PCDD/F internal standards, the retentions of all PCDDs and PCDFs can be calculated. After comparison with the retentions of practical environmental samples, the predicted values have been proved to be very accurate. PMID- 10705537 TI - Microtox testing of pentachlorophenol in soil extracts and quantification by capillary electrochromatography (CEC)--a rapid screening approach for contaminated land. AB - An approach to rapid soil testing which involved the use of simple solvent extraction methods was developed. The analytes of interest were priority pollutants of low water solubility which could not be readily removed from the soil using water. Direct toxicity testing of the soil samples by Microtox showed a high background toxicity which prevented realistic toxicity data from being obtained for the contaminants present. A range of different extraction solutions was used in an attempt to extract the contaminants while eliminating the matrix effects of the soil. It was necessary that the solvents selected for extraction of the soil samples were not of significant toxicity, as this could potentially mask the toxic effects of any compounds extracted from the soil. The extraction efficiencies of solvent systems were evaluated using pentachlorophenol (PCP) as a model compound of known toxicity in the Microtox assay. A rapid and cost effective method was developed in order to determine the amount of PCP recovered from the soil by the extraction solvents employed. This method consisted of a solid phase extraction (SPE) step followed by quantification using capillary electrochromatography (CEC). Recoveries were greater when a higher proportion of organic solvent (methanol) was used in the extraction process, and lowest when water was used. An extraction based on water could provide information on the potential for leaching of contaminants from the soil into nearby water bodies in an environmental setting. An organic solvent extraction method could indicate how much toxicity soil-dependent organisms might be exposed to through ingestion. Extraction based on 50% (v/v) methanol in water was considered to be the most suitable overall extraction solution for soil screening, given that this permitted extraction of the water-insoluble compound PCP at a level which was clearly toxic in the Microtox assay while also retaining the capability to extract water-soluble contaminants. PMID- 10705538 TI - Polychlorinated organic compounds in the Arctic cod liver: trends and profiles. AB - Polychlorinated organic compounds (POCs) have been measured in Arctic cod liver from Vestertana Fjord for a period of 1987-1998. Significant decrease was observed for DDD (p = 0.043), alpha-HCH (p = 0.001), and gamma-HCH (lindane; p = 0.001). Contents of DDE, 2,3,7,8-tetrachlorodibenzofuran, PCBs, chlordanes, chloronaphthalenes, hexachlorobenzene and polychlorodiphenyl ethers had no significant trend. Contents of three hexa- and two heptachlorodibenzofurans and octachlorodibenzofuran increased slightly from 1987 to 1994, but then at very high rate from 1994 to 1998. Trends of HCHs, profiles of PCBs and levels of chlordanes are in accordance with atmospheric long range transport. The hexa-, hepta- and octachlorodibenzofurans observed are major impurities in chlorophenol formulation Ky-5, which has been used as wood preservative and as fungicide/slimicide in industrial processes. Their profile in Vestertana cod was similar to that observed in Ky-5 contaminated fish. PMID- 10705539 TI - Polychlorinated dibenzo-p-dioxins and dibenzofurans in sediment, soil, fish, shellfish and crab samples from Tokyo Bay area, Japan. AB - Concentrations of tetra- to octa-chlorinated dibenzo-p-dioxins and dibenzofurans in samples collected in or near Tokyo Bay, Japan, with a densely inhabited catchment area, were congener-specifically determined and discussed. Analyzed in this study were samples of surface sediment covering the whole bay area, reference soil representing atmospheric impact, and fish, shellfish and crab commonly consumed as food. The range of concentrations were comparable to or higher than those in other parts of Japan. The origins of these compounds in the catchment area of the bay were investigated in terms of homolog and isomeric compositions in the sediment samples. Biota-sediment accumulation factors for benthic species declined as the degree of chlorination increased. PMID- 10705540 TI - Determination of polychlorinated dibenzo-p-dioxins and dibenzo-furans in solid residues from wood combustion by HRGC/HRMS. AB - PCDD/PCDF were determined in solid samples from wood combustion. The samples included grate ashes, bottom ashes, furnace ashes as well as fly and cyclone ashes. The solid waste samples were classified into bottom and fly ash from native wood and bottom and fly ash from waste wood. For each of the four classes concentration distribution patterns from individual congeners, the sums of PCDD/PCDF and the international toxicity equivalents (I-TEQ) values are given. The I-TEQ levels of fly ash from waste wood burning can be approximately up to two thousand times higher than the values from fly ashes of natural wood. The I TEQ levels in bottom ashes from waste wood combustion systems are as low as the corresponding ashes from the combustion of native wood. Grate ash samples from waste wood combustion systems with low carbon burnout show high levels of PCDD/PCDF. PMID- 10705541 TI - The deep-sea as a final global sink of semivolatile persistent organic pollutants? Part I: PCBs in surface and deep-sea dwelling fish of the north and south Atlantic and the Monterey Bay Canyon (California). AB - The understanding of the global environmental multiphase distribution of persistent organic pollutants (POPs) as a result of the physico-chemical properties of the respective compounds is well established. We have analysed the results of a vertical transport of POPs from upper water layers (0-200 m) to the deepwater region (> 800 m) in terms of the contamination of the biophase in both water layers. The contents of persistent organochlorine compounds like polychlorinated biphenyls (PCBs) in fish living in the upper water layers of the North Atlantic and the South Atlantic, and at the continental shelf of California (Marine Sanctuary Monterey Bay and its deep-sea Canyon) are compared to the levels in deep-sea or bottom dwelling fish within the same geographic area. The deep-sea biota show significantly higher burdens as compared to surface-living species of the same region. There are also indications for recycling processes of POPs--in this case the PCBs--in the biophase of the abyss as well. It can be concluded that the bio- and geo phase of the deep-sea may act similarly as the upper horizons of forest and grasslands on the continents as an ultimate global sink for POPs in the marine environment. PMID- 10705542 TI - The deep-sea as a final global sink of semivolatile persistent organic pollutants? Part II: Organochlorine pesticides in surface and deep-sea dwelling fish of the north and south Atlantic and the Monterey Bay Canyon (California). AB - The understanding of the global environmental multiphase distribution of persistent organic pollutants (POPs) as a result of the physico-chemical properties of the respective compounds is well established. We have analysed the results of a vertical transport of POPs from surface water to deepwater in terms of the contamination of the biota living in the respective environmental compartments. Samples were taken from the North and the South Atlantic and from the uprising water region of the continental shelf of California (Marine Sanctuary Monterey Bay and its Canyon). The contents of persistent organochlorine pesticides (DDTs, chlordanes, toxaphenes, HCHs, and HCB) in surface-living fish are compared to those in deepwater fish of the same geographic area. The deepwater biota show significantly higher burdens as compared to surface-living species of the same region. There are also indications for recycling processes of POPs of the class of organochlorine pesticides in the biophase of the abyss as well. It can be concluded that the bio- and geophase of the deep-sea may act as an ultimate global sink for persistent semivolatile contaminants in the marine environment like the soil on the continents. PMID- 10705543 TI - Organochlorine contaminants in Morelet's crocodile (Crocodylus moreletii) eggs from Belize. AB - Non-viable eggs of Morelet's crocodile (Crocodylus moreletii) were collected from Gold Button (GBL) and New River lagoons (NRL) in northern Belize and screened for organochlorine (OC) compounds using gas chromatography (GC) with electron capture detection (ECD). All egg samples from both lagoons (n = 24) tested positive for one or more OCs. Primary contaminants were p,p-DDE and methoxychlor, detected in 100% and 29% of the eggs examined, respectively. Concentrations of individual OC contaminants ranged from 1 ppb (ng chemical/g egg) to > 0.5 ppm (microgram chemical/g egg). Total concentrations of OCs (sum of all OCs) for one egg collected from a nest at GBL reached as high as 0.7 ppm. Sediment samples from both lagoons also tested positive for OCs (lindane, aldrin, methoxychlor, heptachlor epoxide, p,p-DDT, among others). Nest media (soil and plant material) collected from crocodile nests at GBL were positive for p,p-DDT, methoxychlor, aldrin, endosulfan II, and endrin aldehyde. Based on the 24 egg samples analyzed to date, crocodiles from both lagoons are being exposed to OCs. Such exposure may present a health threat to populations of crocodiles in Central America. PMID- 10705544 TI - Synthesis, isolation and purification of C10-C13 polychloro-n-alkanes for use as standards in environmental analysis. AB - Short chain (C10-C13) polychloro-n-alkanes (sPCAs) mixtures were synthesized by refluxing pure n-alkane (> 99%) with sulfuryl chloride (SO2Cl2) in the presence of UV-light (550 W). The free radical initiated reactions produced analogs containing approximately 4-9 chlorine atoms on each carbon chain. Purification of reaction products was achieved by adsorption chromatography on Florisil. The products were characterized by high-resolution gas chromatography/mass spectrometry (HRGC/MS) operated in the electron capture negative ionization (ECNI) and in electron ionization (EI) modes. Individual standards can now be combined to create standards whose profiles resemble that of environmental samples. Quantification of a known amount of the newly synthesized sPCAs mixture, using an industrial formulation as an external standard, resulted in an overestimation (approximately 28%) in its true value. PMID- 10705545 TI - A comparative study of polychlorinated alkanes as standards for the determination of C10-C13 polychlorinated paraffines in fish samples. AB - Fish samples of different origins have been analysed with short column gas chromatography-electron capture negative ion/mass spectrometry (SCGC/ECNI-MS). The influence of standards composed from different polychlorinated alkanes on the quantification of short chain polychlorinated paraffines (CP) was studied. Concentration values varied very differently with the response of the standards depending on their chlorination degree. The results show, that technical CP products should not be used as standard in most cases, because the composition of polychlorinated alkanes in fish is different from sample to sample. PMID- 10705546 TI - Drugs in the environment. PMID- 10705547 TI - Biodegradability of some antibiotics, elimination of the genotoxicity and affection of wastewater bacteria in a simple test. AB - Most antibiotics and their metabolites are excreted by humans after administration and therefore reach the municipal sewage with the excretions. Only little is known about their biodegradability in aquatic environments. It was recognised that genotoxic substances may represent a health hazard to humans but also may affect organisms in the environment. Therefore, the biodegradability of some clinically important antibiotic drugs (ciprofloxacin, ofloxacin, metronidazole) and hereby the elimination of their genotoxicity was investigated as the first step of an environmental risk assessment using the Closed Bottle test (CBT) (OECD 301 D) and the SOS chromotest. Additionally, to assess toxicity of the antibiotics tested against aquatic bacteria (i) a growth inhibition test (GIT) with Pseudomonas putida was conducted, (ii) a toxicity control was used in the CBT and (iii) the colony forming units (CFUs) were monitored in the test vessels. Worst case concentrations of the antibiotics in hospital effluents were estimated and compared with minimum inhibitory concentrations for susceptible pathogenic bacteria and with the genotoxic potency in the SOS chromotest. Both the concentrations calculated for hospital effluents and the adverse effects in bacteria were in the same order of magnitude. None of the test compounds were biodegraded. The genotoxicity was not eliminated. PMID- 10705548 TI - 1-octanol/water distribution coefficient of oxolinic acid: influence of pH and its relation to the interaction with dissolved organic carbon. AB - The distribution of oxolinic acid (OA) between 1-octanol and buffers at a broad range of pH values was studied and found to decrease with increasing pH. The distribution coefficient to dissolved organic carbon (DOC), log DDOC, was estimated and compared with an experimentally derived log DDOC, showing the experimental value to be almost three orders of magnitude higher. Because only the neutral molecule is assumed to distribute to 1-octanol, the interaction with DOC is considered to be electrostatic in character. PMID- 10705549 TI - Sorption and mobility of metronidazole, olaquindox, oxytetracycline and tylosin in soil. AB - Laboratory studies were conducted to characterise four different antibiotic compounds with regard to sorption and mobility in various soil types. Distribution coefficients (Kd values) determined by a batch equilibrium method varied between 0.5 and 0.7 for metronidazole, 0.7 and 1.7 for olaquindox and 8 and 128 for tylosin. Tylosin sorption seems to correlate positively with the soil clay content. No other significant interactions between soil characteristics and sorption were observed. Oxytetracycline was particularly strongly sorbed in all soils investigated, with Kd values between 417 in sand soil and 1026 in sandy loam, and no significant desorption was observed. Soil column leaching experiments indicated large differences in the mobility of the four antibiotic substances, corresponding to their respective sorption capabilities. For the weakly adsorbed substances metronidazole and olaquindox the total amounts added were recovered in the leachate of both sandy loam and sand soils. For the strongly adsorbed oxytetracyline and tylosin nothing was detected in the leachate of any of the soil types, indicating a much lower mobility. Results from defractionation and extraction of the columns (30 cm length) showed that 60-80% of the tylosin added had been leached to a depth of 5 cm in the sandy loam soil and 25 cm in the sand soil. PMID- 10705550 TI - Acute and chronic toxicity of veterinary antibiotics to Daphnia magna. AB - The acute and chronic toxicity of nine antibiotics used both therapeutically and as growth promoters in intensive farming was investigated on the freshwater crustacean Daphnia magna. The effect of the antibiotics metronidazole (M), olaquindox (OL), oxolinic acid (OA), oxytetracycline (OTC), streptomycin (ST), sulfadiazine (SU), tetracycline (TC), tiamulin (TI) and tylosin (TY) was tested in accordance to the ISO (1989) and OECD (1996) standard procedures. The acute toxicities (48-h EC50 value, mg/l) in decreasing order were OA (4.6), TI (40), SU (221), ST (487), TY (680) and OTC (approximately 1000). NOECs were 340 mg/l for TC and 1000 mg/l for M and OL. Toxic effect on reproduction occurred generally at concentrations, which were one order of magnitude below the acute toxic levels. The chronic toxicity (EC50 values, mg/l) in the D. magna reproduction test in decreasing order were TI (5.4), SU (13.7), TC (44.8) and OTC (46.2). The NOECs (mg/l) obtained in the reproduction test with OA, ST, TY and M were 0.38 for OA, 32 for ST, 45 for TY and 250 for M. The observed toxicity of OA to D. magna indicates that this substance, which is a commonly used feed additive in fish farms, has a potential to cause adverse effects on the aquatic environment. PMID- 10705551 TI - Algal toxicity of antibacterial agents used in intensive farming. AB - The growth inhibiting effects of eight antibiotics used either therapeutically or as growth promoters in intensive farming on two species of micro algae, Microcystis aeruginosa (freshwater cyanobacteria) and Selenastrum capricornutum (green algae) were investigated. The effects of the antibiotics benzylpenicillin (penicillin G) (BP), chlortetracycline (CTC), olaquindox (O), spiramycin (SP), streptomycin (ST), tetracycline (TC), tiamulin (TI) and tylosin (TY) were tested in accordance with the ISO 8692 (1989) standard protocol. Algal growth was measured as increase in chlorophyll concentration by extraction with ethanol followed by measurement of fluorescence. Results were quantified in terms of growth rates using the Weibull equation to describe the concentration response relationship. The toxicity (EC50 value, mg/l) in alphabetic order were BP (0.006); CTC (0.05); O (5.1); SP (0.005); ST (0.007); TC (0.09); TI (0.003) and TY (0.034) for M. aeruginosa. BP (NOEC = 100); CTC (3.1); O (40); SP (2.3); ST (0.133); TC (2.2); TI (0.165) and TY (1.38) for S. capricornutum. In this investigation M. aeruginosa is found to be about two orders of magnitude more sensitive than S. capricornutum. It was observed that most of the compounds were unstable during the test period due to hydrolysis and photolysis. PMID- 10705552 TI - Phytotoxicity to and uptake of flumequine used in intensive aquaculture on the aquatic weed, Lythrum salicaria L. AB - Phytotoxicity of Flumequine on the aquatic weed Lythrum salicaria L. was determined by two laboratory models: a single concentration test, by which the effects of 100 mg l-1 were evaluated after 10, 20, 30 days and a multiple concentration test, by which the effects of 5000-1000-500-100-50 micrograms l-1 were evaluated after 35-day exposure. 100 mg l-1 are highly toxic and significantly decrease the growth of plants; this effect increases with time. Concentrations between 5000 and 50 micrograms l-1 induced hormesis in plants, by significantly increasing mean number and dimension of leaves and secondary roots. The effect is the highest at 50 micrograms l-1 and decreases with increase in concentration. Both toxic effect and hormesis can be related to plant drug uptake, quite high, in the order of micrograms g-1. The ecological implication of Flumequine contamination in aquatic environments and the possible use of Lythrum salicaria for bioremediation and/or monitoring technique are discussed. PMID- 10705553 TI - Effects of the antibiotics oxytetracycline and tylosin on soil fauna. AB - Antibiotics may enter the terrestrial environment when amending soils with manure. A Note of Guidance on ecological risk assessment of veterinary medicines was issued in January 1998. Hardly any information about ecotoxicological effects of already existing substances are available. This study has tested the effects of two widely used antibiotics, tylosin and oxytetracycline, on three species of soil fauna: Earthworms, springtails and enchytraeids. Neither of the substances had any effect at environmentally relevant concentrations. The lowest observed effect concentration was 3000 mg kg-1 and in many cases no effect was seen even at the highest test concentration of 5000 mg kg-1. PMID- 10705554 TI - Stability of Tylosin A in manure containing test systems determined by high performance liquid chromatography. AB - Tylosin is a widely used antibiotic for the treatment of infections in swine. Tylosin consists of a mixture of Tylosin A, Tylosin B, Tylosin C and Tylosin D. All components contribute to the potency of tylosin but Tylosin A is by far the major component (usually about 90% and not less than 80%). A fast, robust and easily performed HPLC method has been developed for determination of Tylosin A in the presence of tylosin residues; Tylosin B, Tylosin C and Tylosin D in manure containing incubation media. The separation was performed using a YMC-Pack ODS-AQ column (250 x 4.6 mm i.d., 5 microns particle size) operated at 35 degrees C. The mobile phase consisted of 2.25% (w/v) sodium perchlorate pH 2.5-acetonitrile (60:40 v/v). Detection was performed by measuring the UV absorption at a wavelength of 290 nm. Calibration curves of tylosin made in the incubation medium containing 6.4% manure were linear in the range from 0.375 to 128.0 mg/l (R2 = 0.999). The limit of quantitation (at the RSD 20% level) for Tylosin A was found to be 0.4 mg/l in incubation media containing 6.4% manure. The recovery of Tylosin A was in the range from 100% to 108% depending on the concentration of manure. The reproducibility was good as the relative standard deviation (n = 4) in each matrix tested was in the range from 0.7 to 1.9 at the 25 mg/l level. The stability of Tylosin A was studied under methanogenic conditions and the half life was found to be less than two days. Studies under aerobic conditions showed that the degradation rate was found to increase with increasing concentrations of manure particles in the incubation medium. It is, however, not clear whether the decrease in the concentration of Tylosin A is caused by sorption, abiotic or biotic chemical degradation. The major degradation product of Tylosin A in methanogenic as well as aerobic incubation media has a UV-spectrum and a retention time corresponding to Tylosin B. Furthermore, Tylosin D is believed to be a minor degradation product. PMID- 10705555 TI - Biodegradability of antineoplastic compounds in screening tests: influence of glucosidation and of stereochemistry. AB - Some pharmaceuticals such as antineoplastics are carcinogenic, mutagenic, teratogenic and fetotoxic. Antineoplastics and their metabolites are excreted by patients into waste water. In laboratory testing the frequently used isomeric anti-tumour agents cyclophosphamide (CP) and ifosfamide (IF) were shown to be not biodegradable. They are not eliminated in municipal sewage treatment plants and therefore detected in their effluents. Structural related compounds are beta-D glucosylisophosphoramidmustard (beta-D-Glc-IPM; INN = glufosfamide) and beta-L glucosylisophosphoramidmustard (beta-L-Glc-IPM). beta-L-Glc-IPM has no antineoplastic effects whereas beta-D-Glc-IPM is active against tumours. In contrast to IF and CP and almost all other investigated antineoplastics beta-D Glc-IPM is inherently biodegradable. Improved biodegradability of beta-D-Glc-IPM compared to IF shows that reducing the impact of pharmaceuticals on the aquatic environment is feasible by changing the chemical structure of a given compound exerting a similar mode of action and therapeutic activity. Stereochemistry may be crucial for pharmaceutical activity of the compounds as well as for its biodegradability in the environment. PMID- 10705556 TI - The application of predictive models in the environmental risk assessment of ECONOR. AB - Environmental risk assessment of products requires information on the physico chemical properties, persistence and ecotoxicity of the product, its constituents and possible metabolic and degradation products. Experimental investigations are usually required to generate this information and consequently risk assessment can be costly and time consuming. One possible approach to minimising the amount of experimental testing is to supplement experimental data with data predicted using models such as quantitative structure-activity relationships (QSARs). Using these models, information can be generated based primarily on the knowledge of the chemical structure of the substance(s) under investigation. In this study predictive models were used to assess the environmental risk of the veterinary medicine, ECONOR which contains the active ingredient valnemulin. Available experimental data on the properties, degradability and ecotoxicity of valnemulin was supplemented with predicted data. Where possible, experimental data was used to validate the predicted approaches and this indicated that the predictions were accurate. Information on usage, properties and degradability was input to fate models to predict environmental concentrations (PECs) of valnemulin in soil, pore water and groundwater. Comparison of PECs with experimental and predicted ecotoxicity data for valnemulin indicated that that even under 'worst case' scenarios the environmental risk posed by valnemulin was low. PMID- 10705557 TI - Environmental risk assessment of human pharmaceuticals in Denmark after normal therapeutic use. AB - An environmental risk assessment is presented for the 25 most used pharmaceuticals in the primary health sector in Denmark. Predicted environmental concentrations (PECs) for the aquatic environment were calculated using conservative assumptions and all PECs exceeded 1 ng/l. Measured concentrations were in general within a factor of 2-5 of PECs and ranged from approximately 0.5 ng/l to 3 micrograms/l for nine of the pharmaceuticals reported in literature. The calculation of predicted no-effect concentration (PNEC) based on aquatic ecotoxicity data was possible for six of the pharmaceuticals. PEC/PNEC ratio exceeded one for ibuprofen, acetylsalicylic acid, and paracetamol. For estrogens the PEC/PNEC ratio approached one when non-standard test was used. The ratio was below one for estrogens (standard test), diazepam and digoxin. For the terrestrial compartment, toxicity data were not available, and no assessment was carried out. Comparisons of predicted concentrations of furosemide, ibuprofen, oxytetracycline and ciprofloxacin in sludge based on either preliminary experimental sludge-water partition coefficients (Kd), octanol-water coefficients (Kow) or acid-base constants (pKa) revealed large variations. PMID- 10705558 TI - The prevalence and prognostic significance of electrocardiographic abnormalities. PMID- 10705559 TI - Morbidity and Irish Catholic descent in Britain: relating health disadvantage to behaviour. AB - OBJECTIVES: This paper critically evaluates the evidence for two health-related stereotypes of the Irish, namely that behaviours such as smoking and heavy drinking explain their excess morbidity in Britain, and secondly that, in illness, this ethnic group behaves more stoically. DESIGN: Data are reported on over 850 respondents from each of three cohorts (aged 18, 39 and 58 in 1990/91) of the West of Scotland 20-07 Study, in which a small but pervasive excess of morbidity has been observed in those of Catholic background (in this area associated with Irish descent). Logistic regression was used to investigate any differences in drinking, smoking and participation in sport between those of Catholic and non-Catholic heritage, whilst controlling for sex and social class. Where a difference was observed, we looked for an association between health related behaviour and the Catholic morbidity excess for six measures of physical and mental health. Finally, illness behaviour at age 39 and 58 was investigated for those experiencing one of a number of common symptoms in the month prior to interview, by noting whether a general medical practitioner (GP) was consulted. RESULTS: The only difference in health-related behaviour was in the eldest cohort, where an excess of smoking was observed for the Catholics. However, except for lung power, smoking was not able to explain very much, if any, of the Catholic morbidity disadvantage. For most of the symptoms studied, GP consultation rates were similar, although there was a tendency towards Catholic over-consulting. CONCLUSION: This paper finds minimal evidence in favour of either stereotype: behaviours such as smoking and excess drinking were not strongly associated with the poor morbidity status of the Irish in the population we have studied; neither have the Irish been found to be more stoic in illness. Therefore the stereotypes are not an adequate explanation, nor a necessary correlate, of the frequent finding of raised morbidity in communities of Irish Catholic origin. PMID- 10705560 TI - High prevalence of type 2 diabetes in an urban settlement in Kerala, India. AB - BACKGROUND: Prevalence of type 2 or non insulin dependent diabetes mellitus is high among Indians living in India as well as abroad. Prevalence among persons of Indian origin in many countries is greater than that of people of other ethnic extraction. The Indian state of Kerala is distinguished by a high level of achievement in the health sector, characterised by both lower mortality rates and greater density of health care institutions that ensure access to most people. These attributes make the prevalence of diabetes and the pattern of its management in Kerala worth studying. OBJECTIVE: To estimate the prevalence of diabetes among persons 20 years or older in an urban housing settlement in Trivandrum city, the capital of Kerala, as well as study the management of the disease in subjects affected. DESIGN: Cross sectional survey for detecting diabetes and other chronic diseases in all willing residents of an urban housing settlement in Trivandrum, the capital city of Kerala, as part of a preventive campaign against lifestyle diseases. Fasting plasma glucose, serum triglycerides, cholesterol, height, weight and blood pressure were measured, and a detailed questionnaire administered to ascertain previous diabetic status and management. RESULTS: Overall prevalence of type 2 diabetes is 16.3%. In the 30-64 age group, age standardised prevalence is 13.7%. Gender differences in prevalence are negligible. Greater prevalence is associated with advancing age, body mass index above 24.99, sedentary habits, serum total cholesterol > 239, serum triglycerides > 149, hypertension and smoking. Compared to non-diabetics, diabetics have greater mean and range of fasting plasma glucose values (8.87 +/- 3.6 mM/l as against 4.34 +/- 0.53 mM/l). 32 out of 38 diabetics among the subjects (82.4%) were already diagnosed even before the survey; of them, 89% were on medication. 3% of subjects had impaired fasting glucose, or FPG level between 110-125 mg/dl. CONCLUSION: Prevalence of type 2 diabetes among a group of urban residents in Trivandrum city in Kerala is very high. This is associated also with a high detection rate and compliance to treatment. PMID- 10705561 TI - Using ethnicity as a classification variable in health research: perpetuating the myth of biological determinism, serving socio-political agendas, or making valuable contributions to medical sciences? AB - There is a need for a valid way to classify the human species consistently and reliably, be it to highlight similarities between human populations such as intelligence or physical capacity, to dispel myths about group differences, or to discover 'novel' risk factors for diseases. In contrast to racial divisions, which are usually based on differences in skin colour and physical features, ethnicity is a complex concept which has both socio-cultural and biological components. However, because of the relative vagueness of the term, the interpretation of the 'Ethnicity' construct is not simple, and its definition is often unique to the research project at hand. Therefore conducting ethnicity research necessitates being aware of the differences between the concept of ethnicity and race, acknowledgement of the complexity of the ethnicity construct, and requires that a clear definition of the use of this term be made explicit by the researcher. PMID- 10705562 TI - Taking a first puff: cigarette smoking experiences among ethnically diverse adolescents. AB - OBJECTIVES: To study the social contexts and physiological consequences of an initial cigarette smoking experience among adolescents from four ethnic groups (African American, European American, Hispanic, Native American) who vary by gender and locale (e.g. urban vs rural). METHOD: A qualitative study using individual interviews and focus groups. RESULTS: Results both amplify and reinforce conclusions about peer and family influences on adolescent smoking initiation reported in quantitative studies of teen smoking. Within the broader themes of peers and family, several important sub-themes emerged. The study findings suggest that peer influence can be characterized as social conformity or social acceptance. Males were more likely than females to describe experiences involving peers exerting strong messages to conform to smoking behaviors. Roles played by family members in the initiation process were complex and included those of initiator, prompter, accomplice, and inadvertent source of cigarettes. European American and Hispanic girls provided descriptions of parents/family members as instigators of their first smoking experience. Hispanic adolescents descripted instances in which family members prompted cigarette use at a young age by encouraging the young person to light the adult's cigarette. Finally, ethnic differences in the physiological responses to initial smoking suggest the need to further explore the role of brand preference and variations in inhaling among ethnically diverse adolescents. CONCLUSION: In order to design effective cigarette smoking prevention programs for adolescents, it is important to understand the meaning of smoking behaviors for adolescents from different ethnic and social backgrounds. PMID- 10705563 TI - The Transtheoretical Model and cervical screening: its application among culturally diverse communities in Queensland, Australia. AB - OBJECTIVES: To apply the Transtheoretical Model of Behaviour Change (TTM) to cervical cancer screening to determine and report on the level of support required by different language and cultural groups in Queensland to enhance participation. The model consists of six stages: Pre-contemplation (no intention to be screened, no past action), Contemplation (intention to be screened, no past action), Action (intention to be screened, initial screening), Maintenance (intention to be screened, regular screening), Relapse (no intention to be screened, initial screening) and Relapse Risk (no intention to be screened, regular screening). DESIGN: Focus groups and structured interviews were used to classify women in terms of the model and collect information regarding knowledge, health service contact barriers and enhancing factors and sources of information in relation to cervical cancer screening. The sample was recruited by bi-cultural workers for each community using snowball techniques. RESULTS: The interview sample consisted of Australian South Sea Islanders, Chinese, German, Greek and Moslem women. There was no evidence of significant differences in TTM stage according to ethnicity. Women who intended to be screened in the future were more likely to have positive decisional balance scores and higher knowledge scores than women who did not. Women who had had Pap tests were significantly more likely to have received information from their general practitioner (GP) than women who had not had Pap tests. Women in Action and Maintenance were also more likely to have had their last Pap test by a female GP compared to women in relapse categories. Women in Pre-contemplation were more likely than women who had Pap tests to agree that they would travel a long way to see a practitioner who spoke their own language. CONCLUSION: Classification based on the model was supported both by the decisional balance scale and measures of knowledge. Women in earlier stages of the model were more likely to express preferences for the provision of services in their own language and by a female. Cervical cancer screening among women in Action and Maintenance appeared to be better supported by GPs. Cervical cancer screening promotion for women of diverse cultures and ethnicities has tended to focus on Pre-contemplation and Contemplation stages, however, as most women in this sample were in Action or Maintenance, as are most Australian-born women, structuring cervical cancer screening promotion in terms of the TTM may significantly improve the effectiveness of interventions for women of diverse cultures and ethnicities. PMID- 10705564 TI - Interventions to improve uptake of breast screening in inner city Cardiff general practices with ethnic minority lists. AB - OBJECTIVE: To increase the uptake of breast screening in three inner city GP practices with a high proportion of ethnic minority patients. SETTING: The study was carried out in May and June 1997 in the South East Wales division of Breast Test Wales (BTW). Three inner city general practices in Cardiff, with a low uptake in the previous round of breast screening and a high proportion of ethnic minority women on their lists, were targeted to receive interventions to increase uptake. This preliminary study was not randomized but sought to offer insights into the interventions which may be worth pursuing and the groups that are harder to reach. DESIGN: INTERVENTIONS: identification of ethnic language groups; GP endorsement letter; translated literature including: multilingual leaflet, GP letter, screening invitation; transport to the screening centre; language support. RESULTS: Of 369 women invited, 187 attended for screening. This gives an uptake of 50.7% compared with an uptake of 35.2% in the previous screening round, a statistically significant increase of 15.5%. (95% CI +8.2% to +22.5%). CONCLUSION: Findings show that translated literature, GP endorsement letter and language support by linkworkers were beneficial. The provision of free transport was ineffective and under-utilized. Uptake was highest amongst Urdu and Gujarati speaking groups and lowest for Bengali and Somali speakers which are hardest to reach. There is scope for improving the attendance rate amongst ethnic minority groups but this can costly. PMID- 10705565 TI - Inter-ethnic differences in youth tobacco language and cigarette brand preferences. AB - OBJECTIVE: To describe and understand variations in cigarette brand preferences between adolescents from varying ethnic and gender groups around the US. DESIGN: A qualitative study where adolescents, both smokers and nonsmokers, were interviewed individually in depth. SETTING: Schools and recreation centers in four sites: urban Maryland (Baltimore), urban Texas (Houston), rural Alabama and rural New Mexico. PARTICIPANTS: 121 adolescent volunteers 13-19 years of age, representing African American, white, American Indian and Hispanic ethnic groups, from both genders. RESULTS: Considerable geographic and ethnic variation exists in terminology used by youth to refer to cigarettes and to their use. Clear patterns in brand preference by ethnic group were found that follow patterns of targeted marketing by ethnicity. White teens preferred Marlboro brand cigarettes, while African-American teens who smoke preferred Newports. Hispanic and American Indian teens were more likely to smoke Marlboro or Camel cigarettes. Hispanic teens were most likely to mention low price as a reason for choosing a particular brand or to state that the brand does not matter. Tobacco advertisements targeting ethnic groups and the use of promotional items to encourage teen smoking were also recognized as factors influencing brand preferences. CONCLUSIONS: These findings have implications for the design of intervention programs aimed at curbing teen smoking. When working with teens who already smoke, using youth language to target messages at perceived characteristics of commonly used brands may be more effective and meaningful than talking about cigarette use in general. Another implication of this work is to shed light on what impact an advertising ban would have on teen brand preferences, brand loyalty, and prevalence of smoking. PMID- 10705566 TI - Towards evidence-based health promotion and service provision for new migrants to Australia. AB - OBJECTIVES: To examine patterns of health utilization and health information dissemination among immigrants to Australia in the first 6 months of immigration using data from the Longitudinal Study of Immigrants to Australia (LSIA). METHODS: The population for the LSIA consists of 5178 principal applicants making their first arrival to Australia on a migrant visa between 1 September 1993 and 31 August 1995, inclusive, and who are aged 15 years or over at the time (96% of all principal applicants). The influence of immigration category, country of birth, health status and age on the likelihood of receiving information about health, sources of health information and use of general practitioners were explored using separate logistic regressions for men and women. RESULTS: Women who received health information were older and less likely to have a chronic illness than women who did not. Men who received health information were older and more likely to be in the Business visa category. Younger women, those in Preferential Family visa categories and bilingual women were more likely than other women to have received health information from an NGO. For men, the only significant predictor of source of health information was being in the Independent visa category. Women who used health services were younger, more likely to have a chronic illness, be proficient in English and less likely to be in the Independent visa category than women who did not. Men who used health services were older, more likely to have a chronic condition and have limited English than men who did not. Men who used health services were more likely to be in the Humanitarian visa class and less likely to be in the Concessional Family or Business visa categories than men who did not use health services. They were also more likely to have been born in Oceania, Middle East and North Africa and Africa. CONCLUSIONS: The results of this study indicate that there are important differentials in knowledge of and use of the health system and these differences are unlikely to be captured by using measures based on ethnicity or country of origin alone. Predictors of health service utilization were different for men and women. In particular, age and lack of English proficiency appeared to be barriers to health service use for women. Visa category and country of birth were more important determinants of health service use for men. PMID- 10705567 TI - [Environmental risk factor in Parkinson disease]. PMID- 10705568 TI - [The cerebellum oracle: the body's own virtual reality center]. PMID- 10705569 TI - [Neuropsychological deficits in depressive disorders]. AB - Neuropsychological deficits are considered a significant feature of depression both for their differential value in the diagnostics and the underlying pathophysiology of depressive disorders. The studies reviewed in the present paper predominantly focus on memory disorders as well as frontal-lobe associated dysfunctions such as deficits of attentional performance and executive functions. Despite heterogeneous results in the literature, there is emerging evidence that executive functions and anterograde memory performance are those domains predominantly affected in depressive disorders. The fact that not all depressive patients display neuropsychological deficits rather indicates a dominant role of moderating variables. We discuss the following variables possibly intervening with type and degree of neuropsychological dysfunction: [1] severity and remission of depressive disorders, [2] age of patients and age at the first manifestation of a depressive disorder, [3] psychological factors such as motivation and coping with failure, [4] type and efficacy of antidepressive drug treatment, [5] duration of the stay as inpatients, [6] number of depressive episodes, and [7] gender. Furthermore, we discuss the theory of a dysfunction of fronto-striatal circuits as an underlying mechanism of depressive disorders. The majority of neuropsychological findings seem to support this theory. PMID- 10705570 TI - [Anxiety as a concomitant symptom of depression in the elderly]. AB - Anxiety is a symptom frequently encountered in elderly depressed patients. It is supposed to have an unfavourable prognostic impact. We studied the frequency of psychic anxiety and its relationship to demographic, social and clinical variables in a group of 54 elderly depressed patients admitted to a gerontopsychiatric day-clinic. Severity of depression, cognitive performance, functional level, social situation and life satisfaction were studied by means of standardized instruments. In about half of the patients studied, we found significant psychic anxiety. Compared to the other depressed patients, patients with anxiety suffered more frequently from cognitive impairment and were more dissatisfied with the acceptance of their disease by other persons. They more frequently lived together with other persons in one household. When leaving psychic anxiety out of consideration as a depressive symptom, the severity of depression was not different in patients with and without anxiety. Anxious patients were equally responsive to day-clinic treatment as non-anxious patients. However, duration of treatment was considerably increased. Thus, elderly depressed patients with anxiety show particular psychosocial and clinical features and require special attention in diagnostics and treatment. PMID- 10705572 TI - [30 years multiple system atrophy concept: retrospect and overview of multiple system atrophy]. AB - Multiple system atrophy represents an enigmatic, clinico-pathologically defined neurodegenerative disease. According to findings of the last three decades, the historically derived, previously used terms olivopontocerebellar atrophy, striatonigral degeneration, and Shy-Drager-syndrome should be avoided in clinical use because of both the clinical and morphological overlap between these syndromes. Complex neurodegenerative syndromes other than multiple system atrophy according to recently developed criteria, should rather be referred to as multiple system degenerations. PMID- 10705571 TI - [New antidepressive drugs in comparison with classical tricyclics. Mechanism of action and clinical evaluation]. AB - During the last decade several new agents have been introduced in the treatment of depressive disorders. In comparison to classical tricyclic antidepressants, these agents have less side effects and a very low toxicity. Selective inhibitors of serotonin reuptake do not produce sedation; however, in the initial phase of treatment this is not always an advantage. In contrast, receptor antagonists, such as mirtazapin and nefazodone have moderate sleep-inducing properties. Selective agents are also characterized by certain pharmacologic interactions which can lead to considerable inhibition of the metabolism of other drugs. Classical antidepressants still play an important role in the treatment of severe depressive episodes. Different synaptic effects of new antidepressants lead to an increased availability of monoaminergic neurotransmitters. The article summarizes the most important neurobiological consequences of these acute synaptic effects which might be more closely associated with neurobiological correlates of depression. The authors conclude about the advantages and disadvantages of the different classes of antidepressants from a clinical point of view. PMID- 10705573 TI - [Physiology and methods for studying the baroreceptor reflex]. AB - The baroreflex is of major importance for the moment-to-moment maintenance of arterial pressure particularly during orthostatic stress. Blood pressure increase stimulates the receptors e.g. in the carotid sinuses and the aortic arch, and rapidly increases the receptor discharge rate. Blood pressure decrease induces arrest of impulse transmission to the nucleus of the solitary tract. The impulses are modulated by the nucleus ambiguous, the rostral ventrolateral medulla, the dorsal nucleus of the vagus nerve, parabrachial and paraventricular nuclei and other central structures. Blood pressure increase induces an increase of cardiovagal activity resulting in cardiodeceleration and a decrease of sympathetic peripheral vasoconstrictor outflow. The receptor firing rates show adaptation and resetting to longer lasting blood pressure changes, hysteresis, i.e. firing rates that are higher with rapid blood pressure increase than during the return to baseline pressure. The receptors interact with respiration, chemoreceptor stimulation, central stimuli, exercise and sleep, etc. Baroreceptor function and interaction e.g. with chemoreceptors is compromised in diseases such as diabetic autonomic neuropathy. Guillain-Barre syndrome, arterial hypertension, heart failure and probably in most stroke patients. Fatal complications may result from baroreceptor malfunction. Subtle analysis of the baroreflex is therefore crucial for a refined pathophysiological understanding of these diseases. Pharmacological testing and "neck chamber" negative pressure stimulation of the receptors are as useful as the non-invasive computerized analysis of the interaction of spontaneous blood pressure and heart rate fluctuations. PMID- 10705574 TI - Early events in M-CSF receptor signaling. AB - The M-CSF receptor (M-CSFR) is expressed in monocytes-macrophages and their progenitors, and drives growth and development of this blood cell lineage. The M CSFR is a member of a small family of growth factor receptors exhibiting related structures but distinct tissue-specific functions. This review discusses the early molecular events in the M-CSF signaling mechanisms, positive signals, negative signals, the possible organization of individual signaling pathways, and the problem of achieving specificity in the signal transduction mechanism. PMID- 10705575 TI - Mannose-6-phosphate/IGF-II receptors mediate the effects of IGF-1-induced latent transforming growth factor beta 1 on expression of type I collagen and collagenase in dermal fibroblasts. AB - We have previously shown that insulin-like growth factor-1 (IGF-1) induces the expression of latent transforming growth factor beta 1 (LTGF-beta 1) through activation of c-fos and c-jun oncogenes. In this study we investigated whether IGF-1 induced latent TGF-beta 1 has autocrine effects on dermal fibroblasts and described a possible mechanism. Human dermal fibroblasts were treated with either vehicle, IGF-1 alone, or IGF-1 with either anti-TGF-beta 1 neutralizing antibody or mannose-6-phosphate (M6P) and levels of mRNAs for TGF-beta 1, collagenase and the pro alpha 1(I) chain of type I collagen were then evaluated by Northern analysis. Conditioned medium was also collected from treated and untreated cells and assayed for TGF-beta 1 protein by enzyme-linked immunosorbent assay. The results of the Northern analysis revealed a differential effect on the expression of the pro alpha 1(I) chain of type I collagen and collagenase in dermal fibroblasts: mRNA for the former being significantly increased in response to IGF 1 treatment while that for collagenase was markedly suppressed. These effects of IGF-1 were blocked to a significant extent by TGF-beta 1 neutralizing antibody at a concentration of 0.5-2.0 micrograms/ml. As the TGF-beta 1 induced by IGF-1 is inactive in the traditionally used mink lung epithelial cell growth inhibition assay, we explored the possible role of IGF-II/M6P receptors in facilitating these autocrine effects. The results showed that the greater than two-fold increase (201.9 +/- 38 vs 81.8 +/- 13, p < 0.05) in mRNA for the pro alpha 1(I) chain of type I collagen induced by IGF-1 was at least 60% inhibited by M6P in a time-dependent fashion. A direct correlation between the expression of TGF-beta 1 and the pro alpha 1(I) chain of type I collagen was found in response to either IGF-1 alone or IGF-1 with M6P. Treatment of cell cultures with TGF-beta 1 neutralizing antibody mimicked the effect of M6P. In contrast to the effects on expression of type I collagen, the level of collagenase mRNA was markedly reduced by IGF-1 alone and was restored by the administration of M6P. The levels of TGF beta 1 in conditioned medium from treated and untreated cells showed a similar pattern to that of the mRNA detected by Northern analysis. These findings suggest that IGF-1 induces latent TGF-beta 1 and that the matrix-modulating autocrine effects of LTGF-beta 1 on dermal fibroblasts are facilitated by M6P/IGF-II receptors on these cells. PMID- 10705576 TI - Correlation between ALK-6 (BMPR-IB) distribution and responsiveness to osteogenic protein-1 (BMP-7) in embryonic mouse bone rudiments. AB - Osteogenic protein-1 (OP-1) or bone morphogenetic protein-7 (BMP-7) stimulates cartilage formation in mouse bone rudiments in vitro but arrests terminal differentiation of prehypertrophic chondrocytes into hypertrophic chondrocytes. In this study we report that these effects of OP-1 depend on the developmental stage of the bone rudiment, early stages (E14 and E15 metatarsals) being most responsive. E17 metatarsals that already contained a hypertrophic area that had initiated mineralization were no longer affected by OP-1. We then investigated whether the sensitivity of the early long bone rudiments to OP-1 correlated with high expression of the OP-1 binding type I serine/threonine kinase receptors (activin receptor-like kinase: ALK-2/ActR-I, ALK-3/BMPR-IA or ALK-6/BMPR-IB) at this early stage. We did not find any significant difference in overall mRNA levels of these ALKs between stages E14 through E17 as assessed by RNase protection assays. However, by immunohistochemistry we found that ALK-6 staining was strong in E14 early cartilage primordium and its future perichondrium but dropped sharply to low levels in these cell types until onset of chondrocyte (pre)hypertrophy at E16. By contrast, ALK-2 and ALK-3 immunostainings in E14 were barely detectable. We also examined by immunohistochemistry the local synthesis of OP-1. OP-1 was present in E14 early chondrocytes and forming perichondrium but in low amounts; however, production of OP-1 increased in these cell types with age. All three receptor types as well as OP-1 were present in significant amounts in prehypertrophic chondrocytes and late hypertrophic chondrocytes including those undergoing mineralization. The temporary high immunostaining for ALK-6 in the early proliferating chondrocytes and future perichondrium of E14 bone rudiments, and its absence in older bones correlated with the sensitivity of chondrocytes and perichondrium to (exogenous) OP-1. We therefore propose that the effects of OP-1 on these cells in vitro are mediated by ALK-6/BMPR-IB. We furthermore conclude that locally produced OP-1 is a potential autocrine/paracrine growth factor. Increased local production of OP-1 may be partially responsible for the age-related decrease in responsiveness to exogenous OP-1 with respect to hypertrophy and mineralization of cartilage. PMID- 10705577 TI - Expression of insulin-like growth factor-1 receptor (IGF-1R) and p27Kip1 in melanocytic tumors: a potential regulatory role of IGF-1 pathway in distribution of p27Kip1 between different cyclins. AB - Insulin-like growth factor-1 receptor (IGF-1R) has been shown to be important for melanoma cell growth and survival. In this study we first show, using immunohistochemistry, that progression from benign nevi to malignant melanoma is paralleled by an increased expression of IGF-1R and a down-regulation of the cyclin-dependent kinase inhibitor p27Kip1. Even though the expression of p27Kip1 was drastically reduced compared to benign tumors, detectable amounts of it could be assayed by Western blotting in cultured melanoma cells. To analyze whether there is a causative relationship between the IGF-1 pathway and p27Kip1 expression, melanoma cells were treated with alpha IR-3, an antibody blocking the IGF-1 binding to IGF-1R, or Tunicamycin, which inhibits the translocation of IGF 1R to the cell surface. From these studies we could conclude that the overall expression of p27Kip1 is independent of the IGF-1 pathway. In contrast, the association of p27Kip1 with the different cyclins was drastically affected. Both TM and alpha IR-3 decreased the binding of p27Kip1 to cyclin D1, whose expression was drastically reduced. On the other hand there was an increased binding of p27Kip1 to cyclin E and cyclin A. This redistribution of p27Kip1 may be a mechanism for growth arrest and induction of apoptosis following interruption of the IGF-1 pathway in melanoma cells. PMID- 10705578 TI - A soluble fibroblast growth factor receptor is released from HL-60 promyelocytic leukemia cells: implications for paracrine growth control. AB - The biological activities of fibroblast growth factors (FGF) are mediated by specific cell membrane receptors (FGFR), which have three immunoglobulin-like IgG domains in the extracellular region. The carboxy-terminal segment of the third IgG domain of FGFR1 could be encoded by different exons, designated IIIa, IIIb, or IIIc. While exons IIIb or IIIc encode receptor forms with both intracellular and extracellular domains, the FGF receptor becomes potentially a secreted form lacking the intracellular domain and the transmembrane region when exon IIIa is expressed. Using reverse transcription polymerase chain reaction, we have found that mRNAs encoding the nucleotide sequences of FGFR1-IIIa and FGFR1-IIIc are expressed in HL-60 cells. FGFR1-IIIa fragment was synthesized by a glutathione S transferase gene fusion system. The purified 33 kDa FGFR1-IIIa fragment fusion protein could bind [125I]-labelled FGF-2 in Western ligand blot analysis. Three species of proteins with the molecular weights of 82, 60, and 50 kDa were identified in serum-free, conditioned medium from HL-60 cells by Western blot using an antiserum against purified FGFR1-IIIa fragment fusion protein. Exposure to FGF-2 caused an increase in [3H]-thymidine incorporation into DNA of HL-60 cells and increased cell proliferation, but the addition of FGFR1-IIIa fragment fusion protein inhibited FGF-2-stimulated DNA synthesis and caused a dose dependent inhibition of FGF-2-stimulated cell proliferation. The effects on DNA synthesis were partly reversed by antibody against the FGFR1-IIIa fragment. These results indicate that both cell membrane spanning and secreted FGF receptors are expressed in HL-60 cells, and that the actions of FGFs as paracrine growth factors could be modulated by secreted FGF receptor forms. PMID- 10705579 TI - Basic fibroblast growth factor: serum levels in the female. AB - This study investigated serum levels of basic fibroblast growth factor (b FGF), a potent angiogenic factor, during distinct periods of the female life and compared them with corresponding levels in age-matched males. Healthy females (n = 59) and males (n = 53) were included in the study, divided into six groups: fetuses (cord blood), neonates, children, adults (females in proliferative and secretory phase), pregnant and "elderly" men and women. Serum b FGF levels were measured by an enzyme immunoassay. No statistically significant difference was found between both genders. Blood levels in fetuses and neonates were significantly increased as compared to adults (p = 0.01, p = 0.02, respectively). Restricting the analysis to females, all age groups, but fetuses (p = 0.05), demonstrated no difference when compared to proliferative phase adults. In conclusion, b FGF serum levels do not differ between males and females and are elevated in fetal and neonatal life, when growth and development are enhanced. PMID- 10705580 TI - Biological repair of thyroid cartilage defects by osteogenic protein-1 (bone morphogenetic protein-7) in dog. AB - The efficacy of human recombinant osteogenic protein-1 (OP-1; bone morphogenetic protein-7) in regeneration of dog larynx was examined by treating thyroid cartilage defects (1.5 cm2) in dogs with thyroid allografts covered with host perichondrium or fascia. Prior to implantation allografts were frozen, thawed and demineralized. The treatment groups were as follows: I--Allograft control implant (n = 3); II--Implants coated with 500 micrograms OP-1 (n = 4); III--Implants coated with 100 micrograms OP-1 (n = 3); IV--Implants coated with 500 micrograms OP-1 and covered with neck fascia (n = 3); and V--Implants extracted with 1 M NaCl and guanidine hydrochloride, and coated with 500 micrograms OP-1 (n = 4). Dogs were sacrificed four months following surgery. Each larynx was removed, carefully dissected and a three-dimensional reconstruction of the defect area was performed on serial sections. The results revealed that the implants of control dogs remained intact with no apparent reduction in size and new tissue formation. OP-1 enriched thyroid allografts, dose dependently induced bone, cartilage and ligament-like structures comprising up to 80% of the total regenerated defect area. Boundaries of the defects healed by formation of new bone when bone resided within the old thyroid cartilage layers. Old cartilage not containing bone within its layers healed by complete integration with newly formed cartilage. Both new bone and cartilage were embedded into layers of new ligament-like tissue which expressed specific morphologic and molecular markers. The three newly formed tissues were tightly connected into a "bone-cartilage-ligament continuum" of tissues, suggesting that OP-1 served as a multiple tissue morphogen in this specific microenvironment. PMID- 10705581 TI - Developmental differences in understanding of balance scales in the United States and Zimbabwe. AB - This cross-cultural, developmental study of understanding of the physics of a balance scale involved 2 age groups (5- and 22-year-olds) in 2 countries (the United States and Zimbabwe). A factorial design was used. Each participant solved 6 types of balance scale problems, and the patterns of errors for these problem types were analyzed to determine which rules and concepts the participant used (e.g., whether number of weights was considered). As expected, on average, older participants were more accurate than younger participants, although significant age differences in the Zimbabwean sample were not found for problems solved by weight or distance cues. One finding differing from earlier research was that problems in which the beam balanced were more difficult for all participants. Generally, understanding of the physics of a balance scale appeared fairly similar in the 2 countries. Some cultural practices may account for the findings- in particular, the availability of preschool and age at time of academic specialization in higher education. PMID- 10705582 TI - Tests of the relationship between children's temperament and asthma and of the reliability and validity of the Brief Scale of Temperament. AB - Two studies are reported. One examined the reliability and validity of a brief scale to measure children's temperament; the other tested the relationship of early temperament and the development of asthma. In Study 1, principal caretakers of 46 4- to 7-year-olds, usually the mothers, filled out questionnaires containing the Brief Scale of Temperament (BST) and the Behavioral Style Questionnaire. The caretakers did this twice, about 1 week apart. The first time the children's recent temperament was assessed, and the second time past (> 1 year) temperament was assessed. Internal consistency and correlational analyses revealed substantial reliability and validity for BST assessments of recent and past temperament. Study 2 incorporated the BST in a large-scale survey of 325 families, with parents providing data on their children's asthma and temperament. BST assessments of early (past) temperament were made retrospectively regarding the child's first few years. Comparisons of early temperament revealed no differences between children who developed asthma and those who did not in their overall scores on the BST or for any of the temperament characteristics it measures. In addition, correlational analyses of data for asthmatics showed that early temperament was not related to ratings of the severity of the asthma condition the children developed or the impacts of any of 12 specific asthma triggers, including any involving emotional states, such as stress or worry. PMID- 10705583 TI - Longitudinally foretelling drug use in the late twenties: adolescent personality and social-environmental antecedents. AB - This research focuses on the interrelation of the parent-child attachment, unconventionality, friends' drug use, and the young adult's use of drugs. Data were collected from participants at 4 points in time: early adolescence, late adolescence, early 20s, and late 20s. Data were collected from mothers at the 3 points in time that corresponded with the first 3 collections of data from their children. Both the youths and their mothers were individually interviewed. The findings indicated that the effect of parent-child mutual attachment was mediated through early adolescent personality attributes of greater responsibility, less rebelliousness, and intolerance of deviance. These non-drug-prone personality and behavioral attitudes, in turn, insulated the young adult from affiliating with drug-using peers, and these attitudes were related to less drug use in the early 20s and ultimately in the late 20s. The results suggest that interventions focused on enhancing parent-child mutual attachment should result in a reduction of the risk factors conducive to drug use during the late 20s. The fact that these findings cover a decade and a half, from early adolescence to the late 20s, underscores the significance of placing drug use in a perspective that includes familial and behavioral aspects. PMID- 10705584 TI - Children's level of participation in a false-belief task, age, and theory of mind. AB - The hide-and-seek deception task of M. Chandler, A. S. Fritz, and S. Hala (1989) was modified to provide a more precise estimate of the age at which children acquire and manifest a theory of mind. Two characters (good, bad) and two levels of involvement (pretend play, sociodramatic play) were incorporated into the research design, so that children's representational understanding of deception could be studied. Two-, 3-, and 4-year-olds (N = 90) participated in the study. The results indicated that 4-year-olds used significantly more deceptive strategies than 2- and 3-year-olds in pretend play and in sociodramatic play. There was no difference between 2- and 3-year-olds in the use of deceptive strategies; they used significantly fewer strategies in the bad roles than in the good roles. No significant differences were found in the 3 age groups' performances in the good-character tasks. The reality-masking hypothesis (P. Mitchell, 1994) accounts for the differences in performances on that task; thus, children younger than 4 years old do seem to have a theory of mind. PMID- 10705585 TI - Parental, peer, and teacher influences on the social behavior of Hong Kong Chinese adolescents. AB - Intensive interviews and self-report questionnaires were used to investigate parental, peer, and teacher influences on the prosocial and antisocial behaviors of Hong Kong Chinese adolescents. Students came from 5 academically different high schools. Results indicated that perceived parental influence was positively associated with frequency of prosocial behavior and negatively associated with frequency of delinquent behavior. Students with good relationships with their parents and peers showed lower frequencies of antisocial behaviors than did students with bad relationships. Adolescents in different identity statuses (achievement, moratorium, foreclosure, diffusion; E. H. Erikson, 1968) showed different patterns of prosocial and antisocial behaviors. For example, adolescents in the identity achievement group exhibited high frequencies of prosocial behaviors and low frequencies of antisocial behaviors, but those in the identity moratorium group exhibited quite high frequencies of both prosocial and antisocial behaviors. PMID- 10705586 TI - Cognitive socialization across ethnocultural contexts: literacy and cultural differences in intellectual performance and parent-child interaction. AB - A comparative study of parent-child interaction and its relation to children's intellectual achievement is presented. The question of cultural continuities in cognitive development was examined. The cross-national design also illustrates some of the problems encountered when such relationships are studied across social contexts in general. The results suggest that although interaction characteristics are related to children's intellectual achievement, that relation is moderated by context factors that may operate differently in each culture. The findings are discussed in terms of how literacy mediates parents' teaching styles in ways that remain culturally ingrained. Research issues and recommendations for future research and policy are discussed. PMID- 10705587 TI - Hemispheric processing deficits in patients with paranoid schizophrenia. AB - The purpose of the present study was to investigate hemispheric deficits in individuals with paranoid schizophrenia on four kinds of tasks: dichoptic viewing tasks involving verbal and nonverbal visual stimuli, and dichotic listening tasks involving verbal and nonverbal auditory stimuli. As dependent measures, both accuracy and speed of (correct) responding were measured. The sample recruited for this study consisted of 18 patients with paranoid schizophrenia, 15 outpatients with anxiety disorders, and 20 controls with no history of psychiatric disorders. Results indicated that, relative to the controls, the paranoid schizophrenic patients were less accurate and less efficient on auditory verbal tasks requiring right hemisphere processing. Unlike the controls the paranoid schizophrenic patients manifested a lateralized left hemisphere advantage. PMID- 10705588 TI - Stability of emotion talk in families from the toddler to the preschool years. PMID- 10705589 TI - Religious orientation and death obsession. PMID- 10705590 TI - Promoting isolationism in science: the unspoken section agenda. PMID- 10705591 TI - Neck retractions, cervical root decompression, and radicular pain. AB - STUDY DESIGN: Two-group repeated measures. OBJECTIVES: To evaluate the changes in the flexor carpi radialis H reflex after reading and neck retraction exercises and to correlate reflex changes with the intensity of radicular pain. BACKGROUND: Repeated neck retraction movements have been routinely prescribed for patients with neck pain. METHODS AND MEASURES: Ten nonimpaired subjects (mean age, 27 +/- 4 years) and 13 patients (mean age, 35 +/- 9 years) with C7 radiculopathy volunteered for the study. The flexor carpi radialis H reflex was elicited by electrical stimulation of the median nerve at the cubital fossa before and after 20 minutes of reading and after 20 repetitive neck retractions. Subjective intensity of the radicular pain was reported before and after each condition using an analog scale. RESULTS: For patients with radiculopathy, a repeated measures analysis of variance showed a significant decrease in the H reflex amplitude (from 0.81 +/- 0.4 to 0.69 +/- 0.39 mV), an increase in radicular symptoms after reading (from 4.2 +/- 1.3 to 5.6 +/- 1.4 on the visual analog scale), an increase in the H reflex amplitude (from 0.69 +/- 0.39 to 1.01 +/- 0.49 mV), and a decrease in pain intensity (from 5.6 +/- 1.4 to 1.5 +/- 1.3) after repeated neck retractions. There was an association between cervical root compression (smaller H reflexes) and increased pain during reading and between cervical root decompression (larger H reflex) and reduced pain (r = -0.86 to 0.60). Exacerbation of symptoms was found with a reading posture. There were no significant changes in the H reflex amplitude in the nonimpaired group. No changes were found in reflex latency for either groups. CONCLUSIONS: Neck retractions appeared to alter H reflex amplitude. These exercises might promote cervical root decompression and reduce radicular pain in patients with C7 radiculopathy. The opposite effect (an exacerbation of symptoms) was found with the reading posture. PMID- 10705592 TI - Tender point sensitivity, range of motion, and perceived disability in subjects with neck pain. AB - STUDY DESIGN: Descriptive analysis of impairment and disability measures in subjects with neck pain. OBJECTIVES: To identify discrete tender points and overall pressure sensitivity and assess relationships among palpation tenderness, active cervical range of motion, visual analog scale pain scores, and Sickness Impact Profile disability scores. BACKGROUND: Palpation tenderness and cervical range of motion are used to evaluate patients with neck pain, but their ability to predict patient-perceived pain and disability is unknown. METHODS AND MEASURES: We studied 45 women and 15 men with neck pain (mean age, 35 +/- 7 years). Group 1 included 30 persons who had not sought treatment, and group 2 included 30 persons who had just been referred for treatment. RESULTS: Subjects demonstrated low mean pressure pain thresholds of tender points (2.3 +/- 1.3 kg). Regression analysis showed that only neck flexion predicted pain (R2 = 0.23), with decreased flexion associated with higher pain levels. Sickness Impact Profile total score was predicted by neck rotation (R2 = 0.31), group (R2 = 0.16), tender point pressure pain threshold (R2 = 0.04), and neck retraction (R2 = 0.03). Decreased neck rotation, neck retraction, and pressure pain thresholds were associated with higher disability. CONCLUSIONS: Neither palpation tenderness nor cervical range of motion were strong predictors of pain and disability in subjects with neck pain. PMID- 10705593 TI - Defective running shoes as a contributing factor in plantar fasciitis in a triathlete. AB - STUDY DESIGN: Case study of a patient who developed plantar fasciitis after completing a triathlon. OBJECTIVES: To describe the factors contributing to the injury, describe the rehabilitation process, including the analysis of defective athletic shoe construction, and report the clinical outcome. BACKGROUND: Plantar fasciitis has been found to be a common overuse injury in runners. Studies that describe causative factors of this syndrome have not documented the possible influence of faulty athletic shoe construction on the symptoms of plantar fasciitis. METHODS AND MEASURES: The patient was a 40-year-old male triathlete who was followed up for an initial evaluation and at weekly intervals up to discharge 4 weeks after injury and at 1 month following discharge. Perceived heel pain, ankle strength, and range of motion were the primary outcome measures. Shoe construction was evaluated to assess the integrity of shoe manufacture and wear of materials by visual inspection of how shoe parts were glued together, if shoe parts were assembled with proper relationship to each other, if the shoe sole was level when resting on a level surface, and if the sole allowed unstable motion. RESULTS: The patient appeared to have a classic case of plantar fasciitis with a primary symptom of heel pain at the calcaneal origin of the plantar fascia. On initial evaluation, right heel pain was a 9 of 10, plantar flexion strength was a 3+/5, and ankle dorsiflexion motion was 10 degrees. One month after discharge, perceived heel pain was 0, plantar flexion strength was 5/5, and dorsiflexion motion was 15 degrees and equal to the uninvolved extremity. The right running shoe construction deficit was a heel counter that was glued into the shoe at an inward leaning angle, resulting in a greater medial tilt of the heel counter compared with the left shoe. The patient was taught how to examine the integrity of shoe manufacture and purchased a new pair of sound running shoes. CONCLUSIONS: A running shoe manufacturing defect was found that possibly contributed to the development of plantar fasciitis. Assessing athletic shoe construction may prevent lower extremity overuse injuries. PMID- 10705594 TI - [The "difficult" patient in community psychiatry]. AB - The "difficult patient" does not exist in psychiatric textbooks but rather in every day practice. A solely diagnostic description is not very useful, the categorisation into three separate subgroups by Sheets et al. (1982) on the contrary helps a lot. Three case studies describes typical treatment-situations with "difficult patients" in a community psychiatric setting. PMID- 10705595 TI - [Patient education regarding the diagnosis. Results of a survey of psychiatric patients]. AB - PURPOSE: Physicians are obliged to inform all patients about the diagnosis and course of their illness. Often difficulties occur in informing patients with psychiatric disorders. Psychiatrists often fear complications (worsening of the symptomatology) by the disclosure of the diagnosis, which facilitate the informed consent on the treatment. Up to now only a few data concerning this topic are available. METHODS: 255 psychiatric inpatients were given a short questionnaire about their illness, its course, and the wish for being informed about their psychiatric diagnosis and the prognosis of their psychiatric disorder. RESULTS: Most patients want to get informations about their diagnosis (88.6%) and the prognosis of their psychiatric disorder (80.4%). Only 27.8% of the patients described a fear of the disclosure of diagnosis, and some more (35.3%) were afraid to hear the prognosis of their psychiatric disorder. However, this fear did not reduce their wish to become fully informed. 60% of the patients reported a satisfactory information of their diagnosis and were able to give the correct medical diagnosis. Furthermore, our results revealed that those patients not willing to be treated were informed about their diagnosis insufficiently. CONCLUSION: The informing about their diagnosis should play a more important role in the treatment of psychiatric patients. PMID- 10705596 TI - [Substance dependence and suicide in hospitalized patients. A description]. AB - OBJECTIVE: The research group "Suicide and Psychiatric Hospital" has studied inpatient suicides in psychiatric hospitals in Baden-Wurttemberg and Bavaria (Germany) from 1970-1996. Whereas suicides in depression and schizophrenia are described very well, a description of addicted inpatients who committed suicide is still missing. PATIENTS AND METHODS: We studied the data of substance dependent patients of the collective of 726 inpatient suicides. RESULTS AND CONCLUSIONS: Among the generally rare suicides of addicted inpatiens--33 of 726- about 81% were alcohol dependent. The frequency of suicide in this group of patients decreased from 6.7% in the years 1970-1984 to 1.4% between 1990 and 1996. Whereas the age remained more or less stable in the middle of the fifth decade, the time from hospitalisation until suicide shortened from 192 days (mean) to 52 days. Suicide of alcohol dependent and poly-drug-abusing patients today almost exclusively takes place during treatment on an open station and among patients being in treatment on a voluntary basis. Suicide attempts in the anamnesis are extraordinarily rare in alcohol dependent suiciders. PMID- 10705597 TI - [Comparison of suicide attempts by male and female adolescents. Results of a 10 year patient population of a child and adolescent psychiatry clinic]. AB - OBJECTIVE: This study examines the way in which suicide attempts differ between male and female adolescents. METHODS: The characteristics of 173 female and 62 male adolescents who were treated at a clinic for child and adolescent psychiatry over a period of 10 years were investigated. RESULTS: Suicide attempts by adolescents in the patient population did not decrease in accordance with suicides in the same age-group among the general population. Male adolescents more often attempted suicide in a place where they were less likely to be discovered, but did not use a "harsher" method than female adolescents. The diagnosis rendered according to ICD-10 in 36% of cases was not always typical for the respective sex. CONCLUSIONS: The ways in which adolescents attempt suicide have clearly changed over the past years. Diagnoses that are untypical for the respective sex may represent a risk factor. PMID- 10705598 TI - [Homeless men in inpatient psychiatric treatment--a controlled study. 1: Health status and self assessment at intake]. AB - OBJECTIVE: This study examines the objective and the subjectively reported state of health, the social network and the utilisation of mental health services in a representative group of homeless men (n = 50) at time of admission to a psychiatric hospital and compares these results with a control group (matched by diagnosis) of non-homeless men. METHOD AND PATIENTS: The BPRS, the SF-12 Health Survey and a neglection index were administered. The main psychiatric diagnosis (ICD-10) were alcohol addiction (n = 29), drug addiction (n = 13), schizophrenia (n = 7) or personality disorder (n = 1). RESULTS: No differences were found according to sociodemographic basis data, but the homeless group had a smaller social network and less financial resources. There was a higher rate of involuntary admission in the homeless group, less contact to mental health services in the weeks before admission, more psychopathological symptoms and more physical neglection. Self-rating of mental and physical health, however, did not differ significantly. There was a positive correlation between thought disturbance and positive self-rating of mental health. CONCLUSION: The mental and physical health of the homeless patients was markedly worse. Beneath structural barriers symptoms, the extreme distress of their living situation and the decreased insight and motivation for treatment are characteristics of this group of patients which make them difficult to treat. PMID- 10705599 TI - [Homeless men in inpatient psychiatric treatment--a controlled study. 2: Effects of treatment]. AB - OBJECTIVE: This study examines the objective and the subjectively reported state of health in a regional representative sample of homeless men (n = 50) at time of admission into a psychiatric hospital and shortly before discharge and compares the results with a control group (matched by diagnosis) of non-homeless men. METHOD AND PATIENTS: The BPRS and the SF-12 Health survey were administered. The main psychiatric diagnosis (ICD-10) were alcohol or drug addiction (84%), schizophrenic disorders (14%) and personality disorders (2%). RESULTS: The median of hospitalisation was 26 days with no significant differences between the two groups. At discharge outpatient treatment was planned for only 16% of the homeless patients but for 40% of the controls. There was a significant improvement in symptoms and self reported state of health. On admission and discharge thought disturbance and a positive self-rating of mental health were significantly correlated. CONCLUSIONS: Against the wide-spread clinical prejudice that inpatient treatment of homeless mentally ill men is not effective our results show that the objective state of health as well as the self-perceived mental and physical health was improved by a regular psychiatric in-patient treatment. The results furthermore indicate the limitations of inpatient treatment the need for outpatient treatment programmes. PMID- 10705600 TI - [Assisted living for former long-term hospitalized psychiatric patients]. AB - OBJECTIVES: The aim of the study is the evaluation of essential characteristics of patients who entered the sheltered group homes of the Psychosozialen Dienst (PSD) in the city of Vienna after the establishment of sectorized psychiatric out patient care facilities. METHOD: Eighty patients who lived in these group homes on the first key day, June 30th 1993, were investigated. Any change in their living situation and rate of hospitalization was ascertained at follow-up, 3.5 years after the first key day. RESULTS: The patients had an average period of hospitalization of 240.5 days per year before entry to a group home, which decreased to 12.4 days per year after entry to a sheltered group home. At follow up more than half of all patients (65%) were still able to live in the community successfully. The number and the length of hospitalizations between the first key day and the follow-up were lowest for patients who had moved to private homes. CONCLUSIONS: Sheltered group homes play an essential role in the process of rehabilitation towards independent living within the community. The results demonstrate that rehabilitation in private apartments can be possible even after 5.7 years of residence in sheltered group homes. PMID- 10705601 TI - [Model creation in art therapy. A sculpture project at a psychiatric clinic]. AB - Therapeutic modelling has found little attention in the literature. For the first time, the present article reports about sculptural work in psychiatric art therapy. Since 1994 the Psychiatric Hospital Kilchberg/Zurich is carrying out a sculpture project, in which about 30 patients work with sand- or limestone for 2 1/2 hours daily, cared for by eight therapists and protected by detailed safety regulations. At the beginning and the end of each afternoon the patients stand in a circle to communicate inner mood and ideas about the work. An exposition at the end of the project gives the opportunity to demonstrate the sculptures to family, friends and clinical staff. Until now there have been only positive experiences, which speak in favour of a wider use of sculpture in psychiatric art therapy. Sculptural work stimulates the vitality of the patients, who are challenged to use their willpower adaptedly and rhythmically. Especially on patients with autodestructive tendencies sculptural work seems to exert a beneficial influence. PMID- 10705603 TI - ["Refuse hoarding syndrome"]. AB - OBJECTIVE: The "litter hoarding syndrome" is described only occasionally during the past decades. It seems to be rather unknown in the psychiatric literature. In the course of the syndrome the patients gather more and more litter in their homes until it becomes unhabitable. Physicians and social psychiatric services are often confronted with this manifestation of a psychiatric illness. METHODS: Because of the dramatic development, the extent and the specific circumstances this paper reports case of a young female patient with the litter hoarding syndrome. RESULTS: The term "litter hoarding syndrome" was first coined by Dettmering [3] during a lecture on 25.1.1984 in the Psychiatric Clinic of the Eppendorf University Hospital in Hamburg. In 1985 Klosterkotter et al. [7] described the "diogenes syndrome" which offered some nosological similarities. With the exception of this publications an the PhD thesis by Pastenaci [11] only a few reports have been published during the last 28 years throughout the world and no epidemiological data about the syndrome can be found. CONCLUSION: Based on this case some ideas about differential diagnosis and syndrome classification shall be presented. PMID- 10705602 TI - [Chronic and therapy refractory Fregoli syndrome]. AB - We present a 46-year old lady suffering from chronic paranoid schizophrenia accompanied by a chronic Fregoli's syndrome. The patient feels persecuted by a male person and identifies this persecutor in different persons of different shape, different age and different sex. The symptoms are very refractory to treatment despite various neuroleptic therapies. Historical, psychodynamic and neuropsychological aspects of this delusional syndrome are discussed. PMID- 10705604 TI - [Away from life into a trance. Story of an admission]. PMID- 10705605 TI - ["Hearing voices" by a congenitally deaf schizophrenic patient]. PMID- 10705606 TI - [Laboratory findings during the classic swine fever epidemic of 1997-1998]. AB - The results of the laboratory tests carried out by the Institute for Animal Science and Health (ID-Lelystad), the Netherlands, on samples collected during the Classical Swine Fever (CSF) epidemic 1997-1998 are summarized in this article. The relevance of the different laboratory tests and various samples collected on the eradication of CSF during an outbreak is evaluated. PMID- 10705608 TI - [Law on obviousness]. PMID- 10705607 TI - [Ornithobacterium rhinotracheale infections in poultry: a review]. AB - O. rhinotracheale is a relatively new bacterium. It is found in commercial fowl and wild birds throughout the world. O. rhinotracheale causes respiratory disease, presenting as pneumonia and air sacculitis. It is transmitted horizontally as well as vertically. O. rhinotracheale is difficult to isolate. Serologically, twelve serotypes can be distinguished, of which serotype A is the most prevalent. Treatment can be difficult, because acquired resistance against the regular antibiotics is common in O. rhinotracheale isolates. An inactivated vaccine for broiler breeders has been developed and for turkeys an inactivated autovaccine can be made. PMID- 10705609 TI - Effect of some antibiotics on pigmentation in Serratia marcescens. AB - Serratia marcescens is characterized by its ability to produce a red pigment called prodigiosin. It is well known that there are some substances affecting pigmentation in bacteria. Cefoxitin, erythromycin, tobramycin, co-trimoxazole, imipenem and nitrofurantoin were found to have an inhibitory effect on pigmentation in a S. marcescens strain isolated from urine. It was also shown that the LD50 dose determined by inoculation of eight-week-old BALB/c mice with serial dilutions of overnight cultures of pigmented and nonpigmented variants was lower (LD50 = 300 x 10(3.94)) in the nonpigmented variant than in the pigmented one (LD50 = 300 x 10(5.58)). In addition, the Sereny test showed that in contrast to the pigmented variant, the nonpigmented variant caused keratitis in guinea pig eye. PMID- 10705610 TI - Examination of polyclonal rabbit immune sera against serovars of Acinetobacter baumannii and genospecies 3 for cross-reactions with reference strains of other named/unnamed genospecies of Acinetobacter. AB - Polyclonal rabbit immune sera against 38 serovars of Acinetobacter baumannii and genospecies 13 capable of growth at 44 degrees C and against 26 serovars of genospecies 3 and genospecies 13 incapable of growth at 44 degrees C were examined for serological cross-reactivity with reference strains comprising 17 genospecies (among them 6 named species) of Acinetobacter. Checkerboard agglutination tests yielded very few cross-reactions. Specifically, genospecies 17 cross-reacted weakly with A. baumannii serovar 27; strains genospecies 13 (Bouvet), TU 14, and 'close to TU 13' were strongly agglutinated by antiserum against A. baumannii serovar 18. Genospecies 14 (Bouvet) yielded a very weak cross-reaction with genospecies 3 serovar 20, and genospecies TU 13 reacted weakly with anti-genospecies 3 serovar 15 serum. The 'between genospecies 1 and 3' reference strain proved to be A. baumannii serovar 5 as determined with absorption tests. PMID- 10705612 TI - Occurrence of siderophores in enterococci. AB - In 70 strains of the genus Enterococcus belonging to 16 species and isolated from clinical material, animals and the environment, a hydroxamic class of siderophores was detected with the aid of chemical and biological tests. A correlation between siderophore activity and species affiliation was found: E. faecalis strains showed the highest siderophore activity. A correlation between siderophore activity and pathogenicity could not be confirmed. PMID- 10705611 TI - Identification of Borrelia burgdorferi sensu stricto, Borrelia garinii and Borrelia afzelii in Ixodes ricinus ticks from southern Bohemia using monoclonal antibodies. AB - The prevalence of Borrelia burgdorferi sensu lato (s.l.) in 761 Ixodes ricinus ticks collected in different localities of southern Bohemia was investigated. The minimum infection rate of nymphs, males and females as determined by dark-field microscopy was 4.8% in 1997. The total B. burgdorferi prevalence determined in ticks in 1998 was 11.04%. A significant local variability of the prevalence of B. burgdorferi was not found. Nine isolates were identified at a genospecies level by immunoblotting with prepared monoclonal antibodies (Mabs). The most prevalent genospecies in ticks was B. burgdorferi sensu stricto (s.s.) (6 strains). Two tick isolates reacted with Mabs specific to both B. garinii and B. afzelii and one isolate was detected as a mixed culture of B. burgdorferi s.s. and B. garinii. Considerable variability in protein composition among the strains obtained was found to exist. Five B. burgdorferi genospecies-specific Mabs were prepared and used for the identification of spirochaete isolates. Mabs 9B8 and 6C5 reacted with OspA and OspB proteins of B. burgdorferi s.s., while 4D7 and 1D11 recognised OspA protein of B. garinii and 9D2 specifically reacted with the OspB protein of B. afzelii. In addition, Mab 5H4 reacted with flagellin of all the above-mentioned genospecies of B. burgdorferi s.l. PMID- 10705614 TI - Phenotypes of staphylococcal resistance to macrolides, lincosamides and streptogramin B (MLS) in a Turkish university hospital. AB - Resistance to macrolides, lincosamides and streptogramin B (MLS) which is expressed either constitutively or inducibly, is mediated by erm genes (erm A, erm B, and erm C in staphylococci). The transposon TN 554, harbouring the erm A gene also encodes spectinomycin resistance. In Turkey, data related to MLS resistance phenotypes of staphylococci are not available. In this study, we screened 500 consecutive clinical isolates of staphylococci isolated in Hacettepe University Hospital, for MLS and spectinomycin resistance by the standard disk diffusion method. All MLS-resistant isolates were further tested for spectinomycin susceptibility by the agar screening method. Of 500 staphylococcal isolates, 368 (73.6%) were susceptible and 132 (26.4%) were resistant to MLS antibiotics. Ninety-one (18.2%) of the resistant isolates exhibited a constitutive resistance pattern, whereas 40 were inducibly resistant. MS (resistance to macrolides and lincosamides only) resistance was detected in only one isolate (0.2%). Of 40 inducibly resistant isolates, 21 were found to be resistant to spectinomycin by both the disk diffusion and agar screening tests, probably indicating a presence of the erm A gene. These results suggest that MLS resistance has been considerably high among clinical isolates of staphylococci in our hospital. On the whole, constitutive resistance was the pattern most frequently encountered. In contrast, MS resistance was very rare. Further epidemiological and molecular investigations are required for clarification of the data presented. PMID- 10705613 TI - Aminoglycoside resistance mechanisms in clinical isolates of Pseudomonas aeruginosa from the Canary Islands. AB - Strains of Pseudomonas aeruginosa resistant to aminoglycoside antibiotics were selected from 152 clinical isolates. We identified two patterns of resistance correlating with the resistance mechanism characterized by changes in permeability, enzymatic modification due to the acetylating enzyme, AAC(6')-II, or a combination of both. We detected enzymatic activity of the phosphorylase enzyme, APH(3'), in all the isolates. We compared the mechanisms of resistance detected by three methods i.e., radioenzymatic assay, phenotype of resistance and DNA probes. The phenotype of resistance was tested using a kit developed by Schering-Plough Corp. Hybridization was made with 18 DNA probes for the most frequent genes encoding for aminoglycoside-modifying enzymes. All isolates with AAC(6') activity hybridized with the aac(6')-Ib probes and to a minor degree, with the aac(6')-IIb probe. None of the isolates showed hybridization with aph(3')-I, aph(3')-II, or aph(3')-III DNA probes. Serotyping of the strains showed that the O:11 serotype was the most frequent one in strains whose resistance was due to the AAC(6') enzyme. The O:6 serotype was associated with changes in permeability. Encoding of the resistance mechanism seemed to be chromosomal in all the strains. PMID- 10705615 TI - Complementary characterization of a hyaluronic acid splitting enzyme from Streptococcus agalactiae. AB - A hyaluronic acid splitting enzyme of Streptococcus agalactiae was characterized by splitting mechanism, Michaelis-constant and inhibition type for sulfated hyaluronic acid: The enzyme splits hyaluronic acid as a hyaluronate lyase [EC 4.2.2.1]. The Km = 8 x 10(-4) mg ml-1 was determined with the influence of substrate inhibition constant Kiu = 2 x 10(-6) mg ml-1. Sulfated hyaluronic acid inhibits the enzyme in a partially non-competitive way. The inhibition constant is Ki = 5.47 x 10(-4) mg ml-1. The GBS-hyaluronate lyase cleaves hyaluronic acid as an endoglycosidase. The work is related with the intention to establish a hyaluronate lyase of microbial origin as a therapeutical enzyme replacing bovine hyaluronidase. PMID- 10705616 TI - Enhancement of antiherpetic activity of cytarabine by hydroxyurea in human embryonic skin-muscle fibroblasts. AB - Pharmacological induction of low deoxyribonucleoside triphosphate (dNTP) levels is a novel approach to combine inhibition of virus replication. In accordance with this concept, the alteration of antiherpes activity of cytarabine (AraC) in combination with hydroxyurea (HU), an inhibitor of ribonucleotide reductase, was studied. The results were confirmed by virus yield assays in human embryonic skin muscle fibroblasts (HESMF). It was demonstrated that HU significantly enhanced the antiviral activity of AraC against HSV-1 and HSV-2 in HESMF cultures and this inhibition was reversed by an excess of deoxycytidine (dCyt). The observations suggest that the reduction in dCTP level accounts for the potentiating effect of HU on the antiviral activity of AraC. PMID- 10705617 TI - Modulation of calcium-induced clot formation of human plasma by Malassezia furfur. AB - The effect of Malassezia furfur on clot formation by human plasma was examined. The clotting time in the presence of M. furfur or Candida albicans was measured. M. furfur shortened the clotting time of calcium-induced clots by human plasma in a concentration-dependent manner. However, the protein content of the clots formed were not significantly different between the M. furfur-treated and the control group. The clotting time of clots triggered by thrombin or by placing plasma in glass tubes, which artificially activate the blood coagulation systems, were not affected by treatment with M. furfur. Moreover, acetone-treated M. furfur also shortens the calcium-induced clot formation time, while treatment with zymolyase, which causes decomposition of beta-glucan, did not shorten it. These results suggest that M. furfur activates the blood coagulation systems, and the beta-glucan portion of M. furfur plays a key role in shortening calcium induced clot formation time. PMID- 10705618 TI - Serotype, antimicrobial susceptibility and clone distribution of Pseudomonas aeruginosa in a university hospital. AB - To study the epidemiology of Pseudomonas (P.) aeruginosa, serotyping and antibiotic susceptibility testing were performed on 208 clinical isolates. Sixteen of these isolates were additionally examined by pulsed-field gel electrophoresis (PFGE) of chromosomal DNA. All 208 isolates belonged to 13 of the 16 described serotypes. Thirty isolates (14.4%) belonged to serotype O6, 75 (36%) to serotype O11 and 53 (25.6%) to other serotypes, 42 (20.2%) were polyagglutinating and eight (3.8%), autoagglutinating. Twenty-six per cent of isolates were resistant to piperacillin, 9.1% to ceftazidime, 9.6% to imipenem, 45.7% to ciprofloxacin, 39.9% to amikacin, 51% to gentamicin, 48.6% to netilmicin and 45.2% to tobramycin. Antibiotic resistance varied according to serotype and was highest in serotype O11. Sixteen isolates were analysed by PFGE; nine were multiresistant serotype O11 isolates recovered in four hospital units, while seven were susceptible serotype O6 or O11 isolates from a single unit. The multiresistant serotype O11 isolates had two PFGE patterns indicating that they were capable of spreading: one PFGE pattern was shared by the isolates recovered in spring and the other by those recovered in autumn 1997. The seven susceptible O6 and O11 isolates from a single unit had seven different PFGE patterns. Our results have shown that serotype O11 was the most prevalent P. aeruginosa serotype in our hospital and that its antibiotic resistance was high. The discriminatory power of serotyping is inadequate to permit the tracing of different strains. Macrorestriction analysis of chromosomal DNA was found to provide the best means of strain discrimination. PMID- 10705619 TI - Isolation of Mobiluncus species from the human vagina. AB - We report the results of a study concerning the characteristics of 52 strains of Mobiluncus spp. isolated from 982 vaginal secretions from patients with suspected bacterial vaginosis. 158 of these women presented the features of this bacterial infection. Of the strains isolated, 39 belonged to the species Mobiluncus curtisii, (25 of these which to M. curtisii subsp. curtisii and 14, to M. curtisii subsp. holmesii), and 13, to Mobiluncus mulieris. The vaginal isolates of Mobiluncus spp. were identified by comparing their biochemical profiles with those of the type strains M. curtisii subsp. holmesii (ATCC 35,242), M. curtisii subsp. curtisii (ATCC 35,241) and M. mulieris (ATCC 35,243). All strains of M. mulieris proved to be sensitive to the antimicrobial agents assayed, while strains of M. curtisii were seen to be resistant only to metronidazole. PMID- 10705620 TI - Molecular comparison of Mycoplasma hominis strains isolated from colonized women and women with various urogenital infections. AB - Twenty Mycoplasma hominis strains isolated from colonized women and women with various urogenital infections were investigated for genetic and antigenic homogeneity by different methods. Restriction fragment length polymorphism analysis demonstrated heterogeneity for all strains, with one exception. Two strains sequentially isolated from one patient showed identical patterns. Otherwise, no clonal clustering could be detected within the strains isolated from either of the diagnostic groups. In contrast, SDS-PAGE analysis and the comparison of the immunoblot pattern revealed antigenic similarities of strains isolated from patients with bacterial vaginosis, chorioamnionitis, premature rupture of membranes and preterm delivery as well as endometritis but showed obvious differences in comparison to strains isolated from colonized women. PMID- 10705621 TI - The future of human genetics is the faster continuum of the past. PMID- 10705622 TI - Plasma lipids, lipoprotein Lp(a), apolipoproteins, and hemostatic risk factors distributions in postmenopausal women. AB - In the present study, the effect of hormone replacement therapy on lipid metabolism, apolipoproteins and hemostatic risk factors for cardiovascular disease was assessed in 216 Croatian postmenopausal women. There were 156 current users divided in to two groups according to the duration of therapy. The short term study of < 10 months (X +/- SD 5.31 +/- 2.69) included 49 users, and long term study of > 11 months (X +/- SD 22.06 +/- 10.95) included 107 users of hormone replacement therapy. Sixty nonusers served as a control group. In the short-term study, current users had a significant increase in serum HDL cholesterol, apolipoprotein A-I, A-II and a decrease in total/HDL cholesterol ratio, apoB and antithrombin III (p < 0.05). No significant differences were recorded for total cholesterol, triglycerides, LDL cholesterol, lipoprotein Lp(a) and plasminogen. In the long-term study, a significant increase in HDL cholesterol, apo A-I and total/HDL cholesterol ratio, and a decrease in AT III were observed. Results of the study showed favorable effects of hormone replacement therapy on serum lipid profile and apolipoproteins as a protective regimen from cardiovascular disease in both treatment groups of postmenopausal women. There are conflicting reports regarding increased fibrinolytic activity. The clinical relevance of the observed changes in antithrombin III concentrations as an important coagulation inhibitor is doubtful and should be considered in a more extensive evaluation of the potential hemostatic risk factors for cardiovascular risk and thromboembolism. PMID- 10705623 TI - Bioelectric impedance in the estimation of hemodynamic and fluid status in dialysis patients. AB - In order to estimate the hemodynamic and fluid changes, "dry body weight" and intradialytic stability, electric bioimpedance cardiography was performed in 37 dialysis patients during dialysis procedure, i.e. before, at 2 h and after dialysis. The following parameters were estimated: systemic vascular resistance index-fl. Ohm/m2 (SVRI), mean arterial pressure-Torr (MAP), thoracic fluid conductivity/Ohm (TFC), cardiac index-L/min/m2 (CI), left cardiac work index-kg m/m2 (LCWI) and ejection fraction % (EF). Results were compared with changes in total body water estimated by the urea kinetic model (UKM). The patients were divided into three groups: normotensive (n = 12), hypertensive (n = 15) and hypotensive (n = 10). EF was increased in all the three groups, but only in hypotensives this change was significant (from 40.5 +/- 9.1 to 50.2 +/- 5.41, p < 0.01). The changes in other hemodynamic parameters (CI, LCWI, SVRI) did not reach statistical significance. TFC decreased significantly in all the three groups: normotensive from 0.056 +/- 0.009 to 0.048 +/- 0.009 (p < 0.001), hypotensive from 0.043 +/- 0.009 to 0.035 +/- 0.058 (p < 0.001) and hypertensive from 0.054 +/- 0.016 to 0.045 +/- 0.014 (p < 0.001). These changes were accompanied by a significant decrease in total body water (TBW): from 34.05 +/- 4.19 to 32.72 +/- 4.51 in the hypotensive group, from 34.06 +/- 7.18 to 32.91 +/- 7.27 in the normotensive group, and from 38.92 +/- 7.06 to 37.59 +/- 7.04 in the hypertensive group. The technique was found to be simple, noninvasive and helpful for the estimation of individual hemodynamic changes during dialysis procedure. PMID- 10705624 TI - Tumor neoangiogenesis in rebiopsied patients with prostatic carcinoma. AB - Tumor neoangiogenesis has proven its prognostic value in malignant tumors of different organs including prostatic adenocarcinoma. The aim of this retrospective study was to disclose the significance of neoangiogenesis in incidental prostatic carcinoma (ICP) after transurethral resection (TURP). The authors examined all patients having ICP diagnosed at the Institute of Pathology, Medical Faculty in Ljubljana during the 1985-1989 period, in whom rebiopsy was performed. Ten of 68 patients were examined to determine how microvessels correlate with the degree of tumor differentiation determined histopathologically, and with Gleason score, stage of disease, and survival time. Microvessels were highlighted by immunostaining endothelial cells for factor VIII related antigen and counted in a x200 microscope field (0.8012 mm2) in the most active areas of neovascularization. Neoangiogenesis and tumor differentiation degree were then correlated between primary tumors and rebiopsies. Tumor differentiation degree was also correlated with the time of the disease progression. Microvessel counts (MVC) showed no association with the degree of tumor differentiation, Gleason score, disease stage, or patients' survival. The time elapsed to the disease progression was associated with less differentiated tumors (p = 0.004; 0.006). Microvessel counts were significantly higher in rebiopsies compared to initial tumors (p = 0.006), but no differences could be observed in the degree of differentiation. The results showed that the determination of microvessel density in rebiopsied patients had a better prognostic value than the degree of tumor differentiation. Nevertheless, the latter showed an association with the time of the disease progression. The number of patients included in the study was too low to confirm the association of MVC and degree of tumor differentiation in primary tumors. PMID- 10705626 TI - Extracorporeal treatment for refractory hyperlipidemia. AB - Radical, non-pharmacological, methods of treatment should never be used until it has first been shown that conventional therapy either fails to control hyperlipidemia or cannot be tolerated by the patient. In general, the use of extracorporeal techniques will be restricted to patients with severe familial hypercholesterolemia, although occasionally they may be resorted to in other categories of hyperlipidemia. Seven different procedures are available today for routine clinical practice: unselective plasma exchange, semi-selective double filtration and its modifications as well as the highly selective procedures of immunoadsorption, chemo-adsorption onto dextran sulfate, heparin induced LDL precipitation lipoprotein(a) column, and LDL hemoperfusion (direct adsorption of lipids--DALI). Large-scale regression studies were performed with five highly selective treatment modalities. Control coronary angiograms obtained after about two years of treatment showed that atherosclerotic plaques on coronary arteries had not enlarged or had even been reduced in 80% to 90% of patients. PMID- 10705625 TI - Pantoprazole versus omeprazole in the treatment of reflux esophagitis. AB - Pantoprazole is a new proton pump inhibitor with a potent antisecretory activity, well defined pharmacokinetics and safety profile. The aim of this single blind, randomized clinical trial was to compare the efficacy of pantoprazole (PAN) 40 mg/day and omeprazole (OME) 20 mg/day in patients with grade I and II GERD (Savary-Miller classification). A total of 120 patients were included (PAN = 60 and OME = 60). In the per protocol/analysis, healing rates at 4 weeks were 76.3% PAN and 71.2% OME (ns), and at 8 weeks 94.7% PAN and 92.9% OME (ns). In the intention to treat analysis, healing rates at 4 weeks were 75% PAN and 70% OME (ns), and at 8 weeks 90% PAN and 86.6% OME (ns). Both pantoprazole and omeprazole were well tolerated with no serious drug related adverse events. Pantoprazole 40 mg/day was found to be safe and effective therapy comparable to omeprazole 20 mg/day in the short-term treatment for reflux esophagitis (grade I and II). PMID- 10705627 TI - Creutzfeldt-Jakob disease in a patient with a lyophilized dura mater graft. AB - A 37-year-old patient with Creutzfeldt-Jakob disease (CJD) is presented, who had received a cadaveric dura matter graft 12 year before the onset of neurologic symptoms. Initial clinical presentation included cerebellar symptoms, with dementia and myoclonus developing in later stages of the disease. EEG showed diffuse slowing with sporadic triphasic periodic activity. CT was normal in the early stage but pronounced cerebral and cerebellar atrophy with widened sulci were seen on MRI in the late stage of the disease. The prion protein (PrP) gene was homozygous for valin at the polymorphic codon 129. Cerebrospinal fluid analysis for 14-3-3 protein was positive. We believe that this patient is the first Croatian to acquire CJD by dural implant. Based on this case and a review of 66 cases from the literature, it is manifest that the awareness of iatrogenic transmission of CJD and adoption of preventive measures are the only effective way to stop the spread of CJD among surgically treated patients. PMID- 10705629 TI - Man and medicine--today and tomorrow. PMID- 10705628 TI - Nasal polyps in children. AB - During the 1980-1995 period, 957 patients were admitted for nasal polyposis, 59 (6.2%) of them children. Results were analyzed with respect to localization of the polyps, allergy, histologic picture, bacteriologic tests, radiologic findings of paranasal sinuses, cytology of nasal mucosa, therapeutic approach, and tendency to relapse. Forty children (22 boys and 18 girls) were analyzed. Their mean age was 11.1 years. Unilateral polyposis was found in 29, and bilateral in 11 children. Antrochoanal polyps (ACP) were found in 30, bilateral nasal polyps in 5, polyps in maxillary sinus in 5, and allergy in 8 children. Inflammation of the maxillary sinus was present in 93% of children with ACP. Their correlation between ACP and inflammation in maxillary sinus suggested that edema of the sinus mucosa have led to protrusion through the ostium and formation of the nasal polyp. PMID- 10705630 TI - Angiomyolipoma of the kidney--an immunohistochemical study. AB - The study included 25 patients with angiomyolipoma of the kidney. None of them had any signs of tuberous sclerosis. Serious clinical complications such as massive hemorrhage (2/25) and hydronephrosis (2/25) were documented in four patients. Concomitant adenoma of the adrenal gland and renal cell carcinoma were found in one patient each. All tumors consisted of a differing mixture of mature adipose tissue, smooth muscle cells and blood vessels, with thickened walls devoid of elastic lamina. Immunohistochemically, the smooth muscle cells stained strongly with antibodies against vimentin, desmin and actin, as well as HMB-45 and NKI/C3. Immunohistochemical staining for NKI/C3 was stronger and more diffusely distributed. These two markers proved to be very useful in the diagnosis of angiomyolipoma, when only needle biopsy specimens are available. PMID- 10705631 TI - Use of octreotide-acetate in preventing pancreatitis-like changes following therapeutic endoscopic retrograde cholangiopancreatography. AB - Acute pancreatitis is a serious complication of endoscopic retrograde cholangiopancreatography (ERCP) and endoscopic sphincterectomy (EST). In addition, serum pancreatic enzymes increase without clinical symptoms in up to 75% of patients undergoing endoscopic procedures. The aim of this trial was to investigate the effects of octreotide in the prevention of these possible complications in patients undergoing therapeutic ERCP. The study was carried out in 209 subjects who were randomly allocated to two groups (A and B). Group A received 0.5 mg of octreotide-acetate subcutaneously one hour prior to ERCP; group B was given placebo. Serum amylase and lipase values were measured before premedication and 1.5, 2, 6 and 24 hours following endoscopy. Following ERCP, the increase in both amylase and lipase values was significantly greater in the control (placebo) group, but this significance disappeared 24 hours following the procedure. Symptoms of acute pancreatitis developed in 4 (3.85%) patients who were given octreotide-acetate, compared to 10 (9.52%) patients in the control group. The results obtained in our study seem to indicate that octreotide could prevent the increase in serum pancreatic enzymes, but no significant difference was observed in the prevention of post-ERCP pancreatitis. PMID- 10705632 TI - Clinical interpretation of brainstem evoked response audiometry abnormalities in cochlear pathology. AB - This investigation involved 45 patients with sensorineural hearing loss (SNHL): 24 with Meniere's disease, 18 with acoustic trauma, and 3 with SNHL due to ototoxic drugs. They all underwent pure tone audiometry and standard brainstem evoked response audiometry (BERA). In patients without wave I in auditory brainstem response, electrocochelography (ECochG) was performed. The findings are presented showing that cochlear lesions (beside threshold elevation) cause latency prolongation of wave I, III and V relative to normal latencies at the actual click hearing level. At high stimulation levels, this effect is almost completely compensated for by the fact that cochlear recruiting ears exhibit steeper latency-intensity curves than do normal ears. But, at the same time this pathology does not cause latency prolongation of central conduction time (CCT). Beside this, cochlear lesions will cause, in some cases, deterioration of replicability (poor waveform resolution) of waves preceding wave V. In such cases, the authors strongly recommend electrocochleography (ECochG) to make wave I visible, because they think that it is the best way to verify the diagnosis of cochlear lesion using BERA. PMID- 10705633 TI - Evaluation of the enzyme test for the detection of clinically significant red blood cell antibodies during pregnancy. AB - In the Croatian transfusion medicine, no general agreement has yet been achieved whether red blood cell (RBC) Rhesus (Rh) antibodies detected during pregnancy only by enzyme tests can cause hemolytic disease of the newborn (HDN). Results of the detection of clinically significant RBC antibodies by low-ionic-strength additive solution antiglobulin test (LISS-IAT) and trypsin enzyme test in 22,947 pregnant women are presented. All pregnant women in whom clinically significant RBC antibodies (RBC-CSA) were detected by LISS-IAT and/or enzyme tests were followed and observed during pregnancy. The women who had enzyme-only anti-D antibodies in their serum were followed up during subsequent pregnancies. Out of 302 positive results obtained by both techniques, irregular clinically significant enzyme-only antibodies (anti-RhD and anti-RhE specificity) were detected in 14 (4.6%) pregnant women. None of 11 RhD positive newborns whose mothers had enzyme-only anti-D antibodies, had signs of HDN after delivery. In these 11 women, anti-D antibodies were detected by LISS-IAT in the first trimenon of subsequent pregnancy. Nine infants born from subsequent pregnancies to women who had previously had enzyme-only anti-D, had clinical signs of HDN. The authors concluded that there is no need for enzyme tests in prenatal testing because enzyme tests are not reliable in the prediction of HDN. PMID- 10705634 TI - Therapeutic plasma exchange. AB - Therapeutic plasma exchange is a treatment modality used for over two decades in a variety of diseases. The purpose of this review is to outline the general principles of, including its rationale, current indications, prescription, and complications. The treatment of complications and methods to reduce the risk of their occurrence are outlined. Apheresis technology developments offer new hopes and promises for the clinician team. PMID- 10705635 TI - Acute renal failure after open heart surgery: prevention and management. AB - The frequency of acute renal failure (ARF) in the setting of open heart surgery is high, because the patients are frequently elderly, may have atherosclerosis, diabetes mellitus, hypertension, or previous renal disease. The appropriate preoperative patient preparation might be the first step treatment. The choice of renal support used in the care of ARF patients may influence the ultimate outcome, and therefore dialysis treatment must be tailored to each individual patient. PMID- 10705636 TI - Biliary duct reoperations. AB - Conventional operations for cholelithiasis are rarely associated with postoperative complications. However, when the complications do occur, they are frequently life threatening and require reoperation. These reoperations are associated with a considerably higher risk than primary procedures, and require maximal caution and experience from the surgeon. Reoperations are performed in anatomically altered conditions, and in a patient psychically and physically exhausted and in fear from repeat procedure. Therefore, it is of utmost importance for each primary operation including conventional procedure for cholelithiasis to be carried out at a high professional level, using all technologic achievements available that facilitate and improve the surgeon's work safety. In this way, the need of undesired and hazardous reoperations is minimized. The rate of and indications for reoperation in 530 patients operated on by the conventional procedure for cholelithiasis during the 1994-1999 period are presented. PMID- 10705637 TI - The hundredth anniversary of the Slavonski Brod Hospital. AB - In 1998, Dr. Josip Bencevic General Hospital in Slavonski Brod celebrated 100 years of its work. The first Hospital building started working on April 1, 1898. At present, the Hospital has 650 beds, 1240 employees, 210 of them with university degree, most of them physicians and some other professionals, and about 550 nurses. All medical specializations have been developed, while an appropriate level of equipment ranks this medical institution high among county hospitals in Croatia. PMID- 10705638 TI - Inhibition of dopamine transport and binding of [3H] spiperone to dopamine D2 receptor by acute Pb-toxicity in vivo. AB - Alterations of dopamine transport and binding of [3H] spiperone to dopamine D2 receptor following acute lead intoxication in an in vivo experimental model in adult rats imitate the acute action of Pb2+ in occupationally exposed workers or in occasional incidents of poisoning. The results show that lead-intoxication causes an increased Pb level in blood, a decrease in dopamine uptake and release in synaptosomes and affects the dopamine D2 receptor. These data show that Pb intoxication may cause severe disturbances in the transport and receptor binding of dopamine, which is involved in the regulation of a variety of functions including locomotor activity, emotion and neuroendocrine secretion. PMID- 10705639 TI - Genetically determined vascular diseases. AB - Review of literature data concerning genetically determined blood vessel diseases is presented. These disorders are a cause of a process similar to Binswanger's disease but occurring familial. Recurrent TIA, ischemic strokes and other kind of blood supply disturbances lead to numerous and various intensity vasogenic brain tissue damage, particularly in the white matter. Progressive neurological symptoms and dementia form the picture of subcortical leukoencephalopathy in several members of family. Moyamoya disease, fibromuscular dysplasia, hereditary hemorrhagic telangiectasia, hereditary cerebral hemorrhage with amyloidosis, pseudoxanthoma elasticum, two types of subcortical encephalopathy in Japan, HERNS and CADASIL are described. PMID- 10705640 TI - Variability of localization and intensity of damage of the white matter of the brain and cerebellum in genetically conditioned diseases. AB - The investigations were based on 3 cases with Leigh, 5 cases with Krabbe's, 4 cases of Alpers, 2 cases with Sandhoff, 1 case with Alexander's disease and 1 case with metachromatic leukodystrophy. In 1 case included into the study we have diagnosed nonketotic hyperglycinemia II. All the diseases under examination are recognized as genetically conditioned or are supposed to be of genetic origin. Damage of the white matter in a more delineated form in certain regions was found in Leigh disease. The changes demonstrated a variable degree of intensity from demyelination to necrosis. More extensive lesions of white matter in gyri and semivoal centrum were found in diseases with simultaneously damaged gray matter e.g. in Alpers and Sandhoff disease. The most extensive changes of diffuse demyelination were found in Krabbe's and Alexander's disease. In these diseases demyelination was accompanied with specific morphological structures e.g. globoidal cells (Krabbe's disease) and Rosenthal fibers (Alexander's disease). The peculiar type of demyelination was characteristic for nonketotic hyperglycinemia of type II. It was expressed by demyelination with vacuolization. PMID- 10705641 TI - Uncompacted myelin in hereditary neuropathy with liability to pressure palsies with the 17 p11.2 deletion. AB - A 16-year-old girl with a typical features of hereditary neuropathy with liability to pressure palsies (HNPP) and deletion on chromosome 17p11.2 was described. In the mother who was asymptomatic the same genetic defect was found. In a sural nerve biopsy obtained from the girl myelin thickenings characteristic for this disease and de- and remyelination in nerve fibers were found. Special attention was paid to the occurrence of uncompacted myelin, which was present in diffuse and focal forms. It is concluded that high amount of uncompacted myelin is characteristic for HNPP and it is probably related to the under-expression of peripheral myelin protein 22. PMID- 10705642 TI - The comparison of microglia maturation in CNS of normal human fetuses and fetuses with Down's syndrome. AB - The study was performed on the tissues derived from the central nervous system (CNS) of 72 normal human fetuses between 8 and 22 week of gestation (GW) and 30 fetuses with genetically confirmed Down's syndrome between 17 and 22 GW. Histochemical, immunocytochemical and ultrastructural examinations of microglial cells in frontal lobe, mesencephalon and cerebellum were carried out. A quantitative evaluation of developing microglia was performed in comparison with astroglial cells by counting the mean number of cells per 1 mm2. The study indicated that microglial cells emerge at the same time in all structures under study, both in normal fetuses and in those with Down's syndrome. It was also found that ameboid microglia (AM) and ramified microglia (RM) emerge at the same time and show the same morphological structure in both groups of fetuses. It was revealed that in the CNS of fetuses with Down's syndrome, the number of ramified microglial cells increased significantly as compared with in normal fetuses. Astroglial cells outnumbered microglial cells in the normal fetal development. Due to the enhanced number of RM cells in the CNS of fetuses with Down's syndrome the quantitative difference between these cells obliterated, and microglial cells in the frontal lobe cortex even outnumbered astroglial cells. PMID- 10705643 TI - Hallervorden-Spatz disease in an adult patient. AB - Hallervorden-Spatz disease (HSD) is an extremely rare degenerative process. The familial studies point to inherited, autosomal recessive neurodegenerative disorder. Quite recently this disease gene has been identified to chromosome 20p12.3-p13. Clinical manifestations of HSD leading to death after several years of illness are most frequently observed in childhood. HSD in adults is very scarce. The case reported concerns a woman who at the age of 26 years began to suffer from slowly progressing psycho-organic syndrome with muscular rigidity, involuntary movements and dysarthria. The patient was hospitalized several times with successive diagnoses of multiple sclerosis, amyotrophic lateral sclerosis and Huntington's disease. Shortly before death magnetic resonance imaging (MRI) scan showed a decreased signal in both basal ganglia. The patient died at the age of 34 years after an eight-year illness. In the brain autopsy symmetric hyperpigmentation of globus pallidus (GP) and reticular part of substantia nigra (SN) was found. The microscopic observation revealed abundant deposits of brown pigment mostly in GP and SN. In addition, numerous spheroids disseminated in the basal ganglia, mesencephalon and medulla oblongata, as well as Lewy bodies in SN were noted. Pigment deposits expressed intensive iron positive reaction by Perls' Prussian-blue method. Based on the described neuropathological changes occurring mostly in GP and SN, Hallervorden-Spatz disease was diagnosed. PMID- 10705644 TI - Ultrastructural changes in substantia nigra and striatum observed on a mouse model of Parkinson's disease induced by MPTP administration. AB - The study was carried out on a mouse model of Parkinson's disease induced by the administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-neurotoxin which damages dopaminergic neurons in substantia nigra. Occurrence of dark degenerated neurons was the most prominent ultrastructural change. They were characterized by the progressive condensation of cytoplasm and nuclear chromatin as well as by the light mitochondria and dilated cisternae of Golgi apparatus. Dark degenerated neurons were found particularly often on the 7th day after toxication, however on the last day of the observation, only a few neurons showed the features of dark degeneration. It is likely that degenerative changes led to death in the part of neurons only. PMID- 10705645 TI - Microglia and neuritic plaques in familial Alzheimer's disease induced by a new mutation of presenilin-1 gene. An ultrastructural study. AB - The results of the ultrastructural study of the brains of two sisters with familial Alzheimer's disease (AD) induced by a new mutation of presenilin-1 (PS 1) gene who died at the young age (35 and 37 years) are presented. In both cases, the changes typical of AD with particularly large number of neuritic plaques (NPs) were found. Microglial cells were located between amyloid core and neurites. At the ultrastructural level, the content of microglial cytoplasm was differentiated (amyloid fibrils or/and phagocytic bodies). This may suggest that microglial cells participate in forming of amyloid fibrils and/or phagocytosis of amyloid. PMID- 10705647 TI - Morphology of experimental diabetes and cerebral ischemia in the rat brain. AB - Male Wistar rats were subjected to streptozotocin administration (85 mg/kg i.p.) and to cerebral air embolia with common carotids ligation. Light microscopy studies showed foci of neuronal loss and dark neurons especially in hippocampus, dentate gyrus, amygdaloid, thalamus and hypothalamus in the diabetic rats. Cerebral ischemia aggravated above changes and additionally, small hemorrhagic foci in putamen close to globus pallidus were observed. Our results indicate on chronic, generalized pathologic process in diabetic-rat's brain, which may be related to an oxidative stress and leads to a death of neurons in necrotic or apoptotic way. PMID- 10705646 TI - Effect of coenzyme Q10 (CoQ10) on superoxide dismutase activity in ET-1 and ET-3 experimental models of cerebral ischemia in the rat. AB - The aim of the work was to evaluate the influence of CoQ10 on superoxide dismutase (SOD) activity levels in the rat model of cerebral ischemia induced by endothelins (ET-1 or ET-3). ETs (20 pmol) were injected into the right lateral cerebral ventricle and immediately CoQ10 was given intraperitoneally (10 mg/kg b.w.). In the brains of experimental animals subjected both to ET-1 and ET-2 administration there was observed a decrease of SOD activity in the brain stem, in the cerebrallum and in the cerebral cortex at all time intervals. ET-1, as compared to ET-3 evoked longer lasting disturbances in SOD activity. In the cerebellum and in the cerebral cortex positive effect of CoQ10 and recovery to the control values was noted after 4 hours in the group subjected to ET-3 injection and after 24 hours in the ET-1 treated animal. Investigated brain areas showed different sensitivity to ETs. Above data may indicate on beneficial effect CoQ10 in the cerebral ischemia via decrease of free radicals concentration. PMID- 10705648 TI - Electron microscopy studies on experimental diabetes and cerebral ischemia in the rat brain. AB - Male Wistar rats were subjected to streptozotocin administration (85 mg/kg i.p.) and to cerebral air embolia with common carotids ligation. Electron microscope studies showed dark neurons, degeneration of endothelial cells and changes in basement membrane of brain capillaries, and changed astroglia in diabetic rats. Our results seem to support our previous findings in light microscopy and correspond with some others authors' suggestions that diabetes leads to chronic, generalized pathologic process in diabetic-rat brain, not-only dependent on vascular pathology, but which may be related to an oxidative/metabolic stress leading to a death of neurons in necrotic or apoptotic way. PMID- 10705649 TI - Border zone neovascularization in cerebral ischemic infarct. AB - Immunocytochemical and quantitative studies on vascular reaction (angiogenesis) in cortical border zone of infarct were undertaken. Intensity and temporal profile of angiogenesis was assessed in 60 patients aged between 48 and 69 (younger group), and between 70 and 92 (older group), with cerebral infarct in the area of middle cerebral artery vascularization, who died during the first six weeks following the stroke. We have found that angiogenesis was a multistage process in which four stages were distinguished: phase of primary activation of endothelial cells, two consecutive phases of active angiogenesis and final phase of only sporadic proliferation of vessels. The distinction of phases in a multiphase angiogenic cascade helped us to evaluate the correlation with survival time and the age of patients. The most pronounced intensification of angiogenesis and increased density of CD 31 positive capillaries in penumbra were observed in the second phase, especially in younger patients. The duration of the penumbral neovascularization decreased in the older age patient. Our results indicate that sprouting angiogenesis is a quantitatively significant source of vessels in the cerebral infarct border zone. However, non-therapeutically stimulated angiogenesis developed only 3-4 days after the stroke, that is beyond the period of reversible changes in ischemic penumbra recognized as a "therapeutical window" in the human brain. The angiogenic therapy opens a new way towards the revascularization of ischemic brain infarct. PMID- 10705650 TI - Neuropathological changes within the brain of rabbits with experimental model of antiphospholipid syndrome in different time after immunization. AB - The post-mortem neuropathological investigations were carried out on 20 female New Zealand rabbits. Two main types of changes were found: inflammatory, including meningeal and perivascular infiltrates, and thrombotic within the nervous tissue. The findings revealed that active process within the CNS persists at least 3 months after APS was evoked, however its intensity, especially necrotic changes and vessel wall thickening evidently diminish. The active APS after experiment had been finished was also confirmed in blood samples. PMID- 10705651 TI - The early stage of multiple sclerosis in the light of molecular and immunological studies. AB - The studied material consisted of patients in the first and second year from the onset of multiple sclerosis. In majority of the studied 18 cases the junctional repertoire of TCR in blood lymphocytes of patients in early phase of MS was restricted demonstrating the mono- or oligoclonal character of rearrangement in the spectrum V delta 1-J delta 1, V delta 5-J delta 1 and V delta 3-J delta 1 in contrast to overwhelming polyclonal picture in the control group. In majority of the cases oligoclonal bands were detected and IgG index was above normal level indicating on intrathecal IgG synthesis. The comparison of humoral immunological markers in the early phase of MS with the control group revealed several higher values in patients, but only concerning TNF alpha level in serum, IgG in CSF and IgG index the differences were statistically significant. After treatment with Prednisone the decrease of all studied markers was established, but significant only of free kappa chains to creatinine ratio in urine. The obtained results indicate, that the early phase of MS is characterised by the profound shift of gamma/delta TCR receptors in direction of mono- or oligoclonal bands in CSF, what may be explained by the oligoclonal expansion of certain B and T cells due to stimulation by an antigen related to MS pathogen. PMID- 10705652 TI - Molecular analysis of PRNP gene in Polish population and in Creutzfeldt-Jakob disease. AB - In our study we have examined allelic variation of codon 129 among the Polish population as well as Polish and Dutch CJD cases. The open reading frame of the PrP gene was amplified using the polymerase chain reaction (PCR). PCR product was digested with Nsp I and Mae II endonucleases and separated by 2% agarose gel electrophoresis and, finally, sequenced by the Sanger dideoxy-mediated chain termination method. To obtain population data we have screened 109 unrelated Polish adults. There were 45% of methionine homozygotes, 16% of valine homozygotes and 3% of heterozygotes. Among Polish CJD cases, 75% were methionine homozygous, 12.5% were valine homozygous and 12.5% were heterozygous, whereas among Dutch CJD cases it was 29% of Met/Met and 71% of Met/Val genotypes. PMID- 10705653 TI - Echigo-1: a panencephalopathic strain of Creutzfeldt-Jakob disease: ultrastructural studies of the optic nerve. AB - The Echigo-1 strain of CJD was isolated by Mori and colleagues (1989) from a case of 33-year-old female with a panencephalopathic type of CJD. An incubation period following intracerebral inoculation of hamsters with 10% cleared suspension of the Echigo-1-affected brain was approximately six months. We report here ultrastructural changes which are comparable with those in the white matter of another panencephalopathic type of CJD, the Fujisaki strain of CJD (GSS) passaged in mice. Vacuoles developed within myelinated axons: within axoplasm or within the myelin sheath and these were accompanied by exuberant reaction of macrophages and hypertrophic astrocytes. Axons underwent Wallerian degeneration and dystrophic neurites were also seen. Most important, we observed proliferation of inner mesaxons. Cross-sectional profiles of innumerable myelinated fibers contained membranous organelles which were continuous with the inner lamellae of the oligodendroglial cells. These unusual proliferations of inner mesaxon formed whorls and elaborated loops. In some axons, proliferation was so severe that loops of mesaxon filled the whole cross-section of the axon. Occasionally, we observed intrusion of the membranous tongue of the inner mesaxon into axoplasm. This study presents a second panencephalopathic model of CJD available in small laboratory rodents. It is important because this is the only such model in hamsters and it may be used for comparative studies of different strains of agent in the same host; thus far only mouse and hamster model have been available for comparative studies. PMID- 10705654 TI - Neuronal loss and apoptosis in experimental Creutzfeldt-Jakob disease in mice. AB - One of the hallmarks of prion disease--neuronal cell loss, may be accomplished by apoptosis. The aim of this study was to estimate the neuronal cell loss in mice brains with experimental Creutzfeldt-Jakob disease (CJD) and control mice in the comparison with the apoptosis appeared by in situ end labelling (TUNEL) in function of time of post-incubation period and developing of the spongiform changes. The number of neurons was considerably lower in terminally sick animals (20-21 week of incubation period) than in control mice. The mean value of loss of neuronal cell was 32%. The greatest loss (55%) of neurons was noted in the septal nuclei of the paraterminal body and the least lost (16%) in the hypothalamus. We report here, that apoptotic cells are readily detectable in CJD-affected mice brains in time-dependt manner after infection of Fujisaki strain, but the number of apoptotic cells detected by in situ end labelling does not well correlate with the extensiveness of neuronal loss. The degree of apoptosis corresponds to the well developed spongiform changes. PMID- 10705655 TI - Constitutive expression of cyclooxygenase-2 (COX-2) in developing brain. A. Choroid plexus in human fetuses. AB - It is believed that prostanoids produced by COX-1 activity are essential for the physiological functions of tissues while those produced by COX-2 lead to various pathological changes in these tissues. Brain is an exceptional organ where in some neurons COX-2 mRNA and its protein are constitutively expressed. Since some prostaglandins may play an important role in the control of blood-brain barrier and cerebral blood flow the purpose of the present study was to examine the COX-2 expression in choroid plexus, which participate in the nutrition of brain parenchyma of human fetuses. Study was performed on 25 brains of human fetuses from 12 to 38 weeks of gestation. In light and electron microscopy characteristic developmental transformation of choroid morphology was observed. In young fetuses from 12 to 20 week of gestation epithelial cells of choroid plexus are cuboidal, contain large amount of glycogen storage and their nuclei are COX-2 immunopositive. From 25 week of gestation until term the amount of glycogen in the choroid plexus diminishes, some apical nuclei are shifted toward central parts of the cells and number of cytoplasm organelles increases. In these cells expression of COX-2 protein is located in cytoplasm but epithelial nuclei are immunonegative. Our results provided evidence that COX-2 is constitutively expressed in the developing human choroid plexus. Different localization of COX-2 in choroid epithelial cells suggests that this enzyme may play a different role in various periods of the choroid plexus development. PMID- 10705656 TI - [The periodical "Das Gesundheitswesen" and changes in public health]. PMID- 10705657 TI - [Case management and expert assessment of work disability]. AB - More than 3 years ago the MDK in Bavaria established a new way of handling medical expertising concerning incapacitation for work. Until that time the physical examination of the patient was seen as an indispensable condition for the statement of capacitation for work. All former efforts to qualify the consultations together with the staff of the institutions of the social health insurance served the purpose of preparing and improving this physical examination oft the patient.--The new bavarian way here presented tried from the beginning to make those examinations unnecessary by improving the cooperation and communication with the treating physicians. PMID- 10705658 TI - [Role of the clinic in follow-up of schizophrenic illnesses]. AB - Schizophrenics have the longest hospital stays and incur the greatest costs with respect to psychosocial care compared to other psychiatric patients. The present study focuses on the relevance of inpatient treatment during the course of schizophrenia with regard to specific symptom characteristics. In addition, the role of the hospital in partnership with local community health care facilities for the care of schizophrenic patients is described. Central aspects of hospitalisation, such as the circumstances of first admission, involuntary treatment, discharge planning, and provision for readmission are analysed and linked to our own data. Schizophrenic patients comprised the largest group of involuntarily treated patients and only 50% said they would return voluntarily to hospital if their symptoms recurred. On discharge from hospital, schizophrenic patients did not differ from other psychiatric patients with regard to their attitudes to hospitalisation. Improvement in both work and interpersonal skills was evident after first inpatient treatment. To provide better care for this group of patients, patient-oriented health provision services with individual attention, treatment and care planning are needed, in contrast to the more traditional hospital-oriented approaches that have been tending to give priority to institutional needs. PMID- 10705659 TI - [Ambulatory detoxification of alcoholic patients--evaluation of a model project]. AB - We present the concept and first catamnestic results of a model project "outpatient alcohol detoxification" which was initiated in 1998. To date 48 patients (18 female, 29 male, mean age 44.5 years, mean duration of alcohol dependence 10.2 years) entered treatment, 43 (92%) patients could be successfully detoxified. In only 21 (44%) patients a medication was necessary. To avoid drugs with abuse potential, doxepine (34%) and clonidine (19%) were predominantly used. No severe medical complications were observed. A follow-up examination after 6 months showed that 26 (55%) patients were abstinent and still in outpatient treatment. Further implications of this project are discussed. PMID- 10705660 TI - [Initial effects of the 1998 revised narcotic law in substitute drug treatment of narcotic dependent patients--results of a physician survey]. AB - To study the consequences of the last revision of the German narcotics act (1998, which forbids prescribing codeines as substitutes), a survey was carried out several months after implementation with 639 physicians, who gave substitution treatment. It could be determined how far codeine patients were ready to change their substitute, how content physicians said their patients were after changing the substitute and which were the attitudes of physicians to the revision in question. RESULTS: The majority of patients (70%) who were formerly substituted by means of codeine accepted methadone as a substitute. After the method of substitution had been changed, 51% of patients had been classified as "very content" and 35% as "reasonably content". Especially those physicians who prescribed codeines in the past, unlike physicians who used methadone for substitution treatment only, criticised the revision in question and feared that many patients should be without medical care as a consequence of the new situation. PMID- 10705661 TI - [Legal principles and discretionary power regarding official autopsy order (administratively determined dissection)]. AB - In Germany, autopsies can be ordered by the representatives of the board of health according the Epidemics Law as so called administrative autopsy. If the autopsy is not absolutely necessary, it is mandatory to take into consideration the rights of the deceased persons as well as the rights of the relatives. The decisions of the legislature concerning clinical autopsies must also be considered. The juridical aspects and different types of organising the interrogation of the relatives are explained. PMID- 10705662 TI - [What do medical students know about the epidemiology of tetanus and tuberculosis in Germany?--A comparison of epidemiologic knowledge regarding a rare and a prevalent infectious disease]. AB - A Comparison of Epidemiological Knowledge About a Rare and a Common Infectious Disease: Tetanus and tuberculosis are still worldwide health problems. Due to an increasing number of insufficiently vaccinated persons, mainly in adults, tetanus in Germany requires more and more medical attention. At the same time, due to global migration and an increasing number of immunocompromised patients, the probability of getting infected with tuberculosis is on the increase. For preventing infectious diseases it is necessary to analyse to what extent medical students are informed about the epidemiology of tetanus and tuberculosis in Germany. This knowledge is a mainstay for later differential diagnosis. After completing medical training it is expected that they are capable of diagnosing both diseases properly and that they are trained to inform the population about necessary vaccinations. The following study analysed the incidence estimation of medical students (n = 472) on tetanus and tuberculosis morbidity in Germany. It is obvious that medical students still have gaps in their knowledge about the epidemiology of tetanus and tuberculosis. They overestimated the incidence of tetanus cases per year around 10-1,000 times. In comparison, they underestimated the incidence of tuberculosis. The study shows a distorted assessment of importance of medical problems by the students of medicine. Medical education should consider this and focus more on epidemiology of infectious diseases including tetanus and tuberculosis. PMID- 10705663 TI - [Chemical accident in Schonbeck--an assessment of the risk to health and environment]. AB - Travelling from Belgium to the BUNA works in Schkopau, ten of eighteen tank wagons filled with vinyl chloride (VC) derailed on the Magdeburg-Halle railway line just outside Schonebeck station. One wagon exploded and four others went up in flames. Buildings and trees in gardens located in the immediate vicinity of the track caught fire and burned. Four owners of garden plots suffered burns. A total of 28 people received inpatient treatment in a nearby hospital, another 268 people were treated as outpatients. The typical symptoms of fume poisoning such as headache, nausea, irritations of respiratory tract and eyes were the primarily diagnosed problems. The vegetation was damaged by flue gases and developing HCl causing fire and caustic burns. Fire brigades and special task forces succeeded to control the looming danger of health and environmental hazards by cooling the burning wagons and pumping the liquid gases from the tank wagons. Vinyl chloride which was released over several days was measured in residential areas to be 0.06 8 ml/m3 air. Vinyl chloride is a gas which is heavier than air. When exposed to light it will be degraded within a few days. A technical guide concentration of 3 ml/m3 air has been adopted for its cancerogenic potential. Dioxin values measured in soils and plants were in the natural range of 20 ng I-TE/kg DS. These values increased to 8300 ng at the very seat of the fire only. With the water used for fire fighting vinyl chloride penetrated into the groundwater revealing values of up to 73 mg/litre. A total of 292 urine samples taken from patients and members of the rescue forces, residents and a control group were tested for their contents of the VC metabolite thiodiacetic acid. However, this number does not allow to draw any conclusions with regard to a potential increase in the risk of cancer. With 0.27, 0.43 and 0.37 mg/litre of urine, the mean values are in the normal range for unexposed people. Only three cases showing values of up to 3.1 mg/litre indicated that a real exposure had taken place. The environmental and health authorities have evaluated the results of the measurements at site. They had set up a consultancy service and were integrated in civil forums. When the project "Extension of the vinyl chloride plant" for PVC production at Buna Sow Leuna Olefinverbund GmbH will be completed, deliveries of vinyl chloride by rail and road will be unnecessary in future. However, the problem of transporting dangerous goods and a complex training of task forces will continue to be of topical importance. PMID- 10705664 TI - [Co-pay regulation in the Netherlands health regulation: an evaluation of the effects]. PMID- 10705665 TI - [Evaluation of the revision of the federal cancer registry law and its long-term sequelae]. PMID- 10705666 TI - Psychological adaptation of adolescents with immigrant backgrounds. AB - In the present study, the author examined 3 theoretical perspectives--family values, acculturation strategies, and social group identity--as predictors of the psychological well-being of adolescents from immigrant backgrounds. The 3 perspectives share the view that immigrants' successful adaptation involves the balancing of their heritage culture and the culture of the society of settlement. The participants were 506 adolescents from 4 backgrounds--Vietnamese, Pakistani, Turkish, and Chilean--who were living in Norway. The 3 theoretical perspectives together accounted for between 12% and 22% of the explained variance of mental health, life satisfaction, and self-esteem. The predictive powers of the different perspectives, however, were dependent on which outcome was predicted. On the whole, social group identity showed the strongest predictive power. PMID- 10705667 TI - In-group favoritism among Native and immigrant Turkish Cypriots: trait evaluations of in-group and out-group targets. AB - The authors investigated whether in-group favoritism manifests itself as praise for the in-group- or as denigration of the out-group. A total of 450 Turkish Cypriots (248 native, 202 immigrant) judged the applicability of positive and negative trait words to in-group and out-group targets. Both the native and the immigrant groups judged the positive traits as more applicable to their respective in-groups than to the out-group. The native group evaluated the negative social traits as more applicable to the immigrant group. The immigrant group also judged the negative social traits as more applicable to themselves. The two groups did not differ in their judgments for more personal negative traits. PMID- 10705668 TI - Dispositional need for cognitive closure and self-enhancing beliefs. AB - In 3 studies, the author examined self-enhancing beliefs as a function of dispositional need for cognitive closure. The results of the 1st study revealed that fathers in the Netherlands believed that they devoted more time to their children than did average Dutch fathers; these beliefs were strongest for participants with a high need for closure. Results of Studies 2 and 3 replicated the findings in Study 1 in a controlled experimental context with approaches developed by S. T. Allison, D. M. Messick, and G. R. Goethals (1989) and by D. M. Messick, S. Bloom, J. P. Boldizar, and C. D. Samuelson (1985). PMID- 10705669 TI - Variables associated with adolescent alcohol use: a multiethnic comparison. AB - The authors examined the influence of sociodemographic variables on the frequency and intensity of alcohol use among a nationally representative sample of Black, Hispanic, and White adolescents who had participated in the 1991 National Household Survey on Drug Abuse (U.S. Department of Health and Human Services, 1993). The sample consisted of 8,756 U.S. adolescents aged 12 to 18 years. The authors found that (a) approximately 19% of the respondents had used alcohol in the last 30 days: (b) among the respondents who had used alcohol, 21% had consumed 1 or more drinks per drinking episode; and (c) there were important similarities as well as important differences in variables that promoted alcohol use among Black. Hispanic, and White adolescents. PMID- 10705670 TI - Beliefs about overcoming psychological problems among British and Japanese students. AB - This study was part of a series investigating lay attributions for the cure of psychological problems. Three groups of students--Japanese students in Japan, Japanese students studying in England, and British students--completed a questionnaire and rated the perceived efficacy of 24 different strategies for overcoming each of 5 psychological problems: agoraphobia, depression, smoking cessation, lack of confidence, and obesity. Factor analysis of the curative strategies revealed 5 interpretable factors, the first 3 of which were Professional Help, Inner Control, and Understanding. There were numerous cultural differences between the British group and both Japanese groups in their written ratings of strategies, particularly professional help, for each psychological problem. PMID- 10705671 TI - Relationship of family socialization processes to adolescent moral thought. AB - The author investigated the relationship between salient family processes and adolescent moral thought among a sample of 271 adolescents and their parents. Family-process variables measured were adaptability, cohesion, and parent adolescent communication; adolescent moral thought was measured by the influence the adolescent participants attributed to sources of moral authority. Perceptions of high family cohesion were associated with the greatest influence attributed to the family as a source of moral authority. Perceptions of high family adaptability were associated with greater influence attributed to all sources of moral authority. Parent-adolescent dyads who perceived high positive communication showed strong parent-adolescent agreement on the levels of influence attributed to all sources of moral authority. The findings (a) support the view that a strong relationship exists between family-socialization processes and the content of adolescent moral thought and (b) redress an empirical imbalance in research literature. PMID- 10705672 TI - Positive and negative responses to personal discrimination: does coping make a difference? AB - Although researches (e.g., K. L. Dion, K. K. Dion, & A. W.-p. Pak, 1992) have associated perceiving personal discrimination with negative psychological symptoms, group consciousness theorists (e.g., S. L. Bartky, 1977) have suggested that perceiving personal discrimination can be empowering. To attempt to reconcile these presumably opposing findings, the author suggested that the method of coping with perceiving personal discrimination would better predict whether the outcomes are negative or positive than would the perception of personal discrimination alone. Female university students (N = 262) in the United States completed questionnaires assessing perceptions of personal discrimination, psychological symptoms, and psychosocial behaviors. Coping mechanisms predicted psychosocial behaviors better than did personal discrimination: The more the participants used social support to cope, the more collective action and less helplessness behavior they reported. Also, the more the participants used avoidance to cope, the more helplessness behavior they reported. PMID- 10705673 TI - Self-focusing situations and depression. AB - The author examined the situations in which people turn their attention to the self and investigated differences between depressed and nondepressed persons in self-focusing situations. He developed a Self-Focusing Situation List containing 58 items and administered the list to 436 Japanese undergraduates. He also administered a depression scale to part of the sample (n = 365). A factor analysis extracted 6 factors, and cluster analyses of these 6 factors scores yielded 5 clusters. The participants who preferred self-focusing when alone and avoided self-focusing after positive events (Cluster 5) scored higher on the depression scale than did the participants in Clusters 1, 2, 3, and 4. PMID- 10705674 TI - Use of the self-concept in forming preferences by French students of different levels of academic achievement. AB - According to the results of recent research in France (D. Martinot & J. M. Monteil, 1995), only high-achieving students possessed well-structured academic self-concepts, which, in academic settings, should facilitate the use of a prototype-matching strategy (i.e., a decision-making strategy in which the self concept guides one's choices). In 2 studies, the authors examined the tendency among French students of different academic levels to use the prototype-matching strategy. In Study 1, the participants of high and average academic achievement, but not those of lower academic achievement, used prototype matching in forming preferences. In Study 2, all participants in the condition (experimental) that facilitated the accessibility of academic self-concepts used prototype matching; the participants in the control condition did not. PMID- 10705675 TI - Gender and conjugal differences in happiness. AB - The author examined conjugal congruence on 4 role experiences--spousal, parental, filial, worker--and on subjective well-being (SWB). According to purposive sampling strategy, 222 community adults (111 married couples) in Taiwan completed a research questionnaire. Conjugal congruence on role experiences was linked to conjugal congruence on SWB as well as personal well-being. Analyses showed that conjugal congruence on role experiences (except the worker role) and SWB was generally high. However, some conjugal discrepancies persisted: The husbands were more committed to the worker role, whereas the wives were more committed to the parental role. Furthermore, conjugal discrepancies in role experiences were related to conjugal discrepancies in SWB as well as to husbands' happiness. PMID- 10705676 TI - Creativity of small groups and of persons working alone. PMID- 10705677 TI - Osteoporosis: a new understanding of its impact and pathogenesis. AB - Osteoporosis is a major public health problem. It is a skeletal disorder characterized by low bone mass and an increased susceptibility to fractures. In the United States, it affects more than 25 million people, accounts for 1.5 million fractures annually, and costs the nation in excess of $10 billion. The consequences of osteoporosis include decreased functional independence and increased morbidity and mortality. Osteoporosis is a multifactorial disease. The pathogenesis of the disorder has genetic, hormonal, and environmental influences that affect peak bone mass, perimenopausal bone loss, and age-dependent bone loss. PMID- 10705678 TI - Evaluation of osteoporosis. AB - Osteoporosis is a silent epidemic that is preventable and treatable. Few people are currently having osteoporosis diagnosed early enough to receive the benefit of prevention. Therefore, most present with a fracture as the first clinical manifestation of the disease. The physician taking a history must have a high index of suspicion for osteoporosis. All women should be counseled about risk factors for osteoporosis. The diagnosis of this disease includes an evaluation of bone mineral density by radiologic technique. The osteopathic physician is uniquely well trained to recognize and diagnose osteoporosis. PMID- 10705680 TI - Management of osteoporosis in the new millennium. AB - Key to effect management of osteoporosis is early diagnosis. Careful assessment of risk and determination of bone mineral density can enable the clinician to recognize this disease before it manifests with fractures. Treatment strategies have been shown to reduce fracture rates and potentially improve bone mineral content. Effective nonpharmacologic treatment includes nutritional considerations, exercise, prevention of falls, behavioral alterations, and osteopathic manipulative treatment. Approved pharmacologic strategies include estrogen replacement therapy, alendronate, and calcitonin. Because of the multiple health benefits of estrogen replacement therapy, unless contraindicated, it should be used whenever possible. The effectiveness of combination therapy is yet to be determined. Serial determinations of bone mineral density and the use of urinary biomarkers are effective to monitor therapy. A "four-step approach" to management is proposed. The primary physician can play a key role in the early recognition and treatment of this potentially devastating disorder. PMID- 10705679 TI - Effective strategies for the prevention of osteoporosis across the life span. AB - Osteoporosis is generally considered an age-related reduction in the quantity and quality of bone, but it need not be considered an inevitable consequence of aging. Primary prevention of osteoporosis is clearly desirable, and all women should receive counseling regarding universal preventive measures such as dietary calcium and vitamin D, weight-bearing and resistance exercises, and smoking cessation. Perimenopausal and postmenopausal women should also be counseled about the potential benefits and risks of hormone prophylaxis. For individuals who are unable to take estrogens, other pharmacologic measures are available for prevention. Osteoporosis may also affect elderly men and patients receiving glucocorticoid therapy, and preventive measures should also be used for them. PMID- 10705681 TI - Doctors' complaints help forge insurance regulation. Payment due. AB - Arkansas doctors are banning to help draft a regulation that would require health maintenance organizations and insurance carriers to process health care claims within 45 calendar days after receipt of a claim. The proposed regulation would be administered by the Arkansas Department of Insurance. If additional information from a claimant is needed to process a claim, a carrier must notify a provider within 10 days of receipt of the claim. Once that information is received by the carrier, it has an additional 45 days to pay. PMID- 10705683 TI - MR cholangiopancreatography is important in evaluation of pancreatic and biliary systems. PMID- 10705682 TI - High-output cardiac failure related to hemodialysis arteriovenous fistula. AB - Among the multiple etiologies for congestive heart failure in patients with end stage renal disease, the contribution of hemodialysis fistulas to the myocardial work load is often overlooked. We recently cared for a patient with high-output cardiac failure from an arteriovenous fistula and review the features and pathophysiology of this condition. PMID- 10705684 TI - Physiologic effects of stellate ganglion block: a result of complete ganglion blockade or the vertical spread of local anesthetic? AB - Traditionally, stellate ganglion blockade has been used for the diagnosis and treatment of upper extremity sympathetic pain. However, this treatment has not been shown to provide adequate sympathetic blockade of the upper extremities. This study demonstrates that a carefully performed upper thoracic sympathetic block with imaging guidance can result in a successful sympathetic blockade of the upper extremities. This study furthermore demonstrates that the occurrence of a Horners syndrome is not a testimony to a successful sympathetic block of the upper extremities. PMID- 10705685 TI - Impact of modern technology on the advancement of medicine. PMID- 10705686 TI - Entering the gene therapy era. AB - Gene therapy is a new modality with the potential for treating a variety of diseases, inherited or acquired. Although the initial clinical trials, particularly those for cystic fibrosis, have not been successful, the preliminary results of more recent studies of gene therapy for myocardial infarction are encouraging. New plasmids, viral and nonviral vectors, and applications are being developed at a rapid pace. A switch is now inserted in the plasmids so that the therapeutic gene can be turned on and off as needed. A chimeric RNA/DNA oligonucleotide has also been designed so that mutated genes can be converted to normal sequences. These and other novel approaches soon will propel the gene therapy industry into reality. Healthcare providers and educators need to be prepared for the coming of the gene therapy era. PMID- 10705687 TI - Modulation of immune responses to DNA vaccines by codelivery of cytokine genes. AB - DNA vaccines containing genes for antigenic portions of viruses have recently been developed as a novel vaccination technology. Direct injection of plasmid DNA in vivo results in prolonged expression of viral proteins and may, thus, mimic the action of attenuated vaccines. An important advantage of this vaccination method is that in vivo-synthesized viral proteins can enter both major histocompatibility complex (MHC) class I and class II antigen-processing pathways to activate specific immunization. In many animal models for infectious diseases, DNA vaccines induced a broad range of immune responses, including antibody, CD8+ cytotoxic T lymphocytes (CTL) and CD4+ helper T (Th) lymphocyte responses, and protective immunity against challenge with the pathogen. The magnitude and nature of these immune responses to DNA vaccines can be further manipulated by codelivery of cytokine genes. Summarizing the many studies reported to date, we can draw conclusions regarding the adjuvant effects of these cytokine genes on DNA vaccines. Coadministration of granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin (IL)-2 genes induces higher antibody titers and T cell proliferation responses than other cytokine genes tested to date. In contrast, the CTL activity is only modestly increased by the GM-CSF and IL-2 genes. The IL-12 gene polarizes the immune responses to DNA vaccines toward Th1 cell development and stimulates the strongest CTL activity. In contrast, co injection of the IL-4 gene promotes the development of Th2 cells and increases production of antibodies, but suppresses CTL activity. Thus, the immune responses to DNA vaccines can be engineered by co-injection of an appropriate cytokine gene to favor the formation of either CTL or neutralization antibodies and, therefore, provide the best protection against a particular pathogen. PMID- 10705689 TI - Recent developments in CD8+ T-lymphocyte memory research. AB - Appropriately activated CD8+ T cells differentiate into cytotoxic effectors capable of eliminating cancer cells and virally infected cells. Successful generation and maintenance of effective CD8+ T-cell memory, through either natural infection or through vaccination, establishes long-term protection against various pathogenic agents and, therefore, contributes significantly to our health. This report is a review of recent advances in CD8+ T-cell memory research. The pool of memory CD8+ T cells is maintained through a mechanism of rapid turnover that is antigen-independent, class I major histocompatibility complex (MHC I) antigen-dependent, and, potentially, IL-15-dependent. Memory CD8+ T cells, in marked contrast to naive CD8+ T cells, constitutively express cytotoxic effector function in the absence of antigen stimulation. Furthermore, the vast majority of activated CD8+ T cells in mice infected with lymphocytic choriomeningitis virus are antigen-specific, prompting revision of the commonly held view that many antigen-nonspecific CD8+ T cells are activated in response to viral infection. These newly published results not only provide exciting insights into the inner workings of CD8+ memory maintenance, but they also establish a sound foundation for future investigation. As more detailed molecular and cellular mechanisms of the regulation of memory CD8+ T-cell survival emerge, the most exciting challenge will be to apply this understanding toward the rational design of vaccines and immunotherapies. These potentials are even more relevant in view of the critical nature of CD8+ T cells in combating viral infection and cancer, and the relative paucity of effective drugs against these diseases. PMID- 10705688 TI - Tumor immunology--when a cancer cell meets the immune cells. AB - Tumor immunology consists of two essential concepts: immune surveillance, which specifies the host immune reactions against tumor cells, and tumor immune escape, which refers to the tumor-cell evasion process against the host immune system. Effective antitumor immunity by the host immune surveillance is supposed to be dependent on the identification of tumor antigens. In the process of malignant transformation, genetic mutations with aberrant expression of cancer-related genes and protein products are potentially immunogenic, which may serve as rejection antigens. Several scenarios are proposed to be responsible for tumor immune-escape mechanisms. The elucidation of the immune deficit against cancer progression has been a difficult task and no solitary mechanism explains the complicated cancer-host immune interactions. Cancer cells may overcome immune surveillance through a common but effective pathway, either by changing the polarity of effector cells, thus down-regulating the proliferation of tumor specific cytotoxic T cells, or altering the effector compositions of immune cells within the tumor milieu, or both. Understanding the interaction between cancer cells and host immune cells within the tumor milieu is of importance for further clinical applications of immunotherapy in cancer treatment. PMID- 10705690 TI - Impact of magnetic resonance imaging on the advancement of medicine. AB - Magnetic resonance (MR) imaging has had a significant impact in many areas of modern medicine. Because of its good image resolution, tissue characterization, and functional assessment of various organs and systems, MR imaging has become an important modern technology in clinical practice and medical research. MR imaging has great flexibility in viewing anatomic structures in arbitrary imaging planes. With ultrafast MR imaging techniques, images of areas of interest can be obtained in a very short time, with elimination of physiologic motion artifacts. MR angiography obviates the need for catheterization and provides highly detailed images of the vascular anatomy, even of structures as small as the coronary artery. MR cholangiopancreatography provides results comparable to those of endoscopic retrograde cholangiopancreatography in many hepatobiliary diseases. Functional MR imaging is a new tool to evaluate physiologic function. MR spectroscopy has been used to explore the metabolic activity of normal and diseased organs and structures. The applications of MR imaging in clinical medicine and biomedical research are expanding. PMID- 10705691 TI - Technological progress in ultrasonography. AB - Technical progress in ultrasonography (US) is especially rapid now, due to continuing advances in transducer design, signal processing techniques, and Doppler technology. A number of important technical breakthroughs have been made in the past decade. Among these, multidimensional array transducers, harmonic imaging, miniaturized transducers, extended field-of-view imaging, hand-carried ultrasound units, three-dimensional (3-D) US, and ultrasound contrast agents are the most remarkable innovations. Improved spatial and contrast resolution allows delineation of anatomic details and increases diagnostic accuracy and confidence. Miniaturized transducers can be used to image tiny or superficial structures in the human body, and can guide the surgeon to the problem site. Extended field-of view imaging provides a larger field for demonstration of pathology in certain anatomic locations. Hand-carried US units are used widely in a physician's office or in remote areas, and bring high-quality medical imaging to the bedside. 3-D US has already shown significant benefits in perinatology; with further improvements in real-time imaging technology, 3-D US will emerge as an important adjunct to conventional 2-D US. In conjunction with harmonic imaging technology and Doppler technology, US contrast agents allow more powerful anatomic and functional evaluation of human organs. They may also play important therapeutic roles in the near future. Advancing computer technology is expected to lead to more important break-throughs in the next 5 to 10 years. PMID- 10705692 TI - Impact of nonsurgical, invasive endoscopy of the diagnosis and therapy of gastrointestinal diseases. AB - In the last 2 decades, endoscopy has become the most reliable means of diagnosing gut disorders. With the aid of magnifying endoscopy and staining methods, gastrointestinal cancers and malignant diseases can be detected in the early stages. Endoscopic ultrasonography is currently indicated for staging digestive cancers, assessing submucosal tumors, and diagnosing biliary and pancreatic diseases. During the past decade, endoscopy evolved from a solely diagnostic tool to a therapeutic modality. Endoscopic heater probes, bipolar electrocoagulation, and laser therapy are all of effective for achieving immediate hemostasis and preventing rebleeding in actively bleeding lesions. Various ligation methods and endoscopic injection sclerotherapy safely achieve high rates of hemostasis. Endoscopic mucosal resection allows curative treatment of gastrointestinal cancer in the mucosal layer, and removal of precancerous lesions. Endoscopic balloon dilators have been developed for dilating gut stenoses. Various endoprostheses are used for palliation of malignant stenoses. Endoscopic sphincterotomy, balloon dilation, lithotripsy, and endoprosthesis placement are alternatives for biliary tract and pancreatic disease therapy. Endoscopic photodynamic therapy is useful in palliation of esophageal cancer or in ablation of dysplastic lesions in Barrett's esophagus. With advances in modern biotechnology, endoscopy continues to show value in treating or preventing diseases, with less discomfort and lower cost than other methods. PMID- 10705693 TI - Impact of medical informatics on medical education. AB - In recent years, medical informatics has become a well-recognized branch of medicine. It is a multidisciplinary science that combines information technology and various specialties of medicine. The impact of medical informatics on medical education is advancing along with the rapid developments in computer science. Departments of medical informatics or similar divisions have appeared in schools of medicine in Taiwan in the past 5 years. At National Taiwan University College of Medicine, we offer curricula in basic computer concepts, network concepts, operating systems, word processing, database and data processing, computer media resources, multimedia computer statistics, intelligent health information systems, medical diagnostic support systems, and electronic medical record systems. Distance learning has also been favorably accepted on this campus. Recently, we proposed the concept of a virtual medical campus, which will break the physical barriers of time and space. We expect this revolution to influence every aspect of medicine, especially medical education. PMID- 10705694 TI - Recent advances in telemedicine. AB - With continuing advances in information technology, the applications of computers in medicine are increasing rapidly. Modern information technology not only affects the delivery of health care, but also significantly influences the doctor patient relationship. Since the 1990s, technologic developments in high-bandwidth telecommunications systems and digitizing devices have led to a surge of interest in telemedicine. In recent years, the Internet, with its powerful penetration and scalability, has become an increasingly popular medical information resource and a new platform for telemedicine. The impact of modern technology on the advancement of telemedicine in Taiwan started with the 1995 National Information Infrastructure project, which uses networks of different bandwidths for teleconsultation and distance education programs. In 1998, National Taiwan University and Taipei Medical College in Taiwan, and the University of Pittsburgh and the University of Iowa in the USA, began cooperation on a new Cyber Medical Center (CMC) project that integrates the technologies of multimedia, database management, a multiple-site videoconferencing system, and the World Wide Web. The aim of the CMC is to create a multimedia network system for the management of electronic patient records, teleconsultation, online continuing medical education, and information services on the Web. In the future, telemedicine systems in Taiwan are expected to combine the Internet and cable television to connect clinics, hospitals, insurance organizations, and public health administrations; and, finally, to extend health services to every household. PMID- 10705695 TI - Long-term survival of patients with mandibular osteosarcoma. AB - Osteosarcoma generally has a poor prognosis. Osteosarcoma of the mandible is rare and may have a less aggressive course. Three patients with osteosarcoma of the mandible were treated during the past 19 years at National Taiwan University Hospital. All were women, with an average age of 49 years. The patients were treated with radical excision of the tumor, with either pre- or postoperative radiotherapy. None of the patients received chemotherapy. Reconstruction with iliac osteocutaneous free flap, fibular osteoseptocutaneous free flap, or Leibinger reconstruction plate was performed to repair the facial defects resulting from tumor resection or radiation-induced necrosis. The three patients were alive and without evidence of recurrence at the time of writing, with follow up periods ranging from 7 to 18 years. All had acceptable facial contour and satisfactory oromandibular function after reconstruction. These results support the effectiveness of wide resection with radiotherapy for treatment of mandibular osteosarcoma. With the availability of microsurgical reconstruction and biocompatible reconstruction plates, we advocate extensive ablation of this tumor. Long-term survival, good functional recovery, and acceptable facial contour can be expected. PMID- 10705696 TI - Varicella pneumonia complicated by acute respiratory distress syndrome in an adult. AB - Primary varicella infection is uncommon in adults, but carries a higher rate of morbidity and mortality than in children. Pneumonia is the most common complication of primary varicella infection in adults. However, varicella pneumonia complicated with acute respiratory distress syndrome (ARDS) is very rare. We report a case of ARDS secondary to varicella pneumonia in a 26-year-old man with primary varicella. The patient was otherwise healthy and had no evidence of human immunodeficiency virus infection. The initial chest radiograph showed bilateral reticulonodular infiltrates, which quickly evolved to diffuse alveolar consolidations. Arterial blood gas analysis revealed a ratio of arterial partial pressure to fraction of inspired oxygen of 87. Abnormal liver function and thrombocytopenia were also noted. Treatment consisted of mechanical ventilatory support and intravenous administration of acyclovir. His pulmonary condition gradually improved and he was successfully weaned from the ventilator 1 week later. He was discharged on the 15th hospital day with a favorable outcome. His pulmonary function improved progressively, with normal spirometry and lung volumes, but persistent mild impairment of diffusing capacity, 6 months after discharge. PMID- 10705697 TI - Tuberculous meningitis in a Filipino maid. AB - Tuberculous meningitis, while not uncommon in Taiwan, has not been reported among foreign workers. We report the first case of tuberculous meningitis in a 37-year old Filipino maid in Taiwan, who presented with headache, fever and vomiting. She had been well before this episode and the small screening films of the chest radiograph obtained on her arrival in Taiwan 15 months previously, and every 6 months thereafter showed no evidence of tuberculosis. The suspicion of tuberculous meningitis was delayed until disturbance of consciousness manifested and a standard chest radiograph showed a diffuse miliary pattern in both lung fields. A cerebrospinal fluid sample that was sent for a polymerase chain reaction-based assay specific for Mycobacterium tuberculosis showed a positive result. The patient recovered with sequelae of mildly incoherent speech and urinary incontinence after antituberculous medication and short-course steroid treatment. Clinicians should be aware of the possibility of tuberculous meningitis in foreign workers with complaints of fever and headache. Because high quality chest radiographs are a prerequisite for early detection of pulmonary tuberculosis, we recommended that standard posterior-anterior chest radiographs should be obtained as part of the routine health examination for foreign workers. PMID- 10705698 TI - Mandibular osteomyelitis caused by Blastoschizomyces capitatus in a child with acute myelogenous leukemia. AB - A 6-year-old girl with acute myelogenous leukemia (AML) developed fungal mandibular osteomyelitis during chemotherapy. Blastoschizomyces capitatus was recognized histologically by its yeast-like morphology and formation of annelloconidia, and was confirmed by culture. The fungal osteomyelitis of the mandible was treated successfully with prolonged antifungal medication, extensive surgical debridement and an oral care program, without interrupting leukemia chemotherapy. B. capitatus osteomyelitis of the mandible may occur during chemotherapy in AML patients with poor dental condition. Successful treatment can be achieved by careful management without interruption of antineoplastic chemotherapy. PMID- 10705700 TI - Econometrics literature on the cost of graduate medical education PMID- 10705699 TI - Graduate medical education costs in nonacademic health center teaching hospitals: evidence from Maryland. AB - As managed care has grown, much concern has been expressed about the potential plight of the nation's 125 academic health centers (AHCs). Less concern has focused on non-AHC teaching hospitals, although most studies of graduate medical education (GME) costs include these hospitals in their estimates. While most studies have found that costs increase positively with various measures of "teaching intensity," some have concluded that hospitals with smaller programs have costs that are the same or less than comparable nonteaching hospitals. However, few studies have tested whether AHCs' cost structures are sufficiently similar to those of other hospitals to reliably include them in the same estimation. This article tests that assumption for Maryland hospitals, finds it violated, and presents results for non-AHC teaching hospitals. The results reveal that, at least in Maryland, even small teaching programs add to hospital costs. PMID- 10705701 TI - Does system membership enhance financial performance in hospitals? AB - While hospitals continue to join multi-institutional systems, empirical data on the benefits of system membership are ambiguous. This study examines the same 166 Florida hospitals in 1986 and 1992. System membership, in general, did not enhance financial returns (measured by operating margin, total margin, and return on assets) for the pooled data or for either year. In fact, a significant negative relationship is noted in 1986. However, when only hospitals affiliated with national systems (in this study, American Medical International, Hospital Corporation of America, or Humana) are analyzed, a positive statistically significant association is found for two of the above three profitability indicators for both the pooled data and for 1986. However, there was no statistically significant impact noted for 1992. Reasons for the apparent discrepancy in the impact of national versus local/regional systems on hospital financial performance and the apparent declining ability of national systems to generate above-average returns are explored. PMID- 10705702 TI - Alcoholism treatment offset effects: an insurance perspective. AB - This study investigates whether alcoholism treatment costs are offset by reductions in other medical treatment costs by comparing people treated for alcoholism with a matched comparison group. The alcoholism treatment group is defined by diagnoses of alcohol dependence, abuse, or psychoses from health insurance claims field between January 1980 and June 1987. A comparison sample was matched on age, gender, and insurance coverage. In this primarily methodological study, expected costs for nonalcoholism treatments were calculated from standardized regressions. Offset effects were measured from the insurer's perspective through differences in expected total nonalcoholism treatment costs in the periods preceding and following alcoholism treatment. Members of the alcoholism treatment group were more likely than the comparison group to be hospitalized and to need other (nonalcoholism) medical treatment, thus incurring higher total costs. Offset effects emerged for patients with alcohol abuse and without mental psychosis comorbidities. PMID- 10705703 TI - Limited English proficiency and Latinos' use of physician services. AB - Many Latinos have limited English proficiency and this may negatively affect their use of health care services. To examine this, the authors interviewed 465 Spanish-speaking Latinos and 259 English speakers of various ethnicities who presented to a public hospital emergency department with non-urgent medical problems to assess previous physician visits, sociodemographic characteristics, and level of English proficiency. The proportion of patients who reported no physician visit during the 3 months before study enrollment was not related to English proficiency. However, among the 414 patients who saw a physician at least once, Latinos with fair and poor English proficiency reported approximately 22 percent fewer physician visits (p = 0.020 and p = 0.015, respectively) than non Latinos whose native language was English, even after adjusting for other determinants of physician visits. The magnitude of the association between limited English proficiency and number of physician visits was similar to that for having poor health, no health insurance, or no regular source of care. PMID- 10705705 TI - Vulnerable populations and health insurance. AB - This study provided a national profile of health insurance of certain vulnerable populations including children, racial/ethnic minorities, low-income families, non-metropolitan statistical area (MSA) residents, and those with poor health status. The study shows an increase in the proportion of uninsured nonelderly population. While public insurance helped reduce the employment- and health related disparities in private coverage, it has not overcome other disparities related to vulnerable characteristics including race/ethnicity, wages, education, and area of residence. Comparison between health maintenance organization (HMO) and fee-for-service insurance indicates that younger although not much healthier people, racial/ethnic minorities, MSA residents, and those residing in the West and Northeast regions were more likely to have HMO coverage. To reduce significant disparities in health insurance coverage, policy makers will have to consider expanding public insurance coverage, targeting vulnerable groups, particularly those with multiple vulnerable characteristics rather than merely the economically distressed. Expecting managed care to achieve cost containment for services provided to vulnerable populations may be unrealistic. PMID- 10705704 TI - The cost of health conditions in a health maintenance organization. AB - In this retrospective cohort analysis of all adults who were members of Kaiser Permanente, Northern California, between July 1995 and June 1996 (N = 2,076,303), the authors estimated the prevalence, average annual costs per person, and percentage of total direct medical expenditures attributable to each of 25 chronic and acute conditions. Ordinary least squares regression was used to adjust for age, gender, and comorbidities. The costs attributable to the 25 conditions accounted for 78 percent of the health maintenance organization's total direct medical expense for this age-group. Injury accounted for a higher proportion (11.5 percent) of expenditures than any other single condition. Three cardiovascular conditions--ischemic heart disease, hypertension, and congestive heart failure--together accounted for 17 percent of direct medical expense and separately accounted for 6.8 percent, 5.7 percent, and 4.0 percent, respectively. Renal failure ($22,636), colorectal cancer ($10,506), pneumonia ($9,499), and lung cancer ($8,612) were the most expensive conditions per person per year. PMID- 10705706 TI - Patients with acute myocardial infarction in northern Italy are often infected by Helicobacter pylori. AB - BACKGROUND: The classical risk factors for acute myocardial infarction (AMI) fail to explain all the epidemiological variations of the disease. Among the new risk factors recently reported, several infectious agents appear to increase the risk of AMI. In particular, acute and chronic respiratory diseases due to Chlamydia pneumoniae, and Helicobacter pylori (H. pylori) infection seem to be strongly involved. The aim of this work is to determine the prevalence of H. pylori infection in a group of male patients with AMI, in a case-control study, where a group of blood donors matched for sex and age served as control. We searched for the classical risk factors in all patients. METHODS: We studied 212 consecutive male patients, aged 40-65 years, admitted for AMI at the Coronary Care Units at Hospitals in three towns of Northern Italy. H. pylori infection was assessed by the highly specific and sensitive 13C-urea breath test and by presence of antibodies (IgG) against H. pylori in circulation. Volunteer blood donors attending our Hospital Blood Bank served as controls. Among the patients we investigated the presence of hypertension, cholesterol and glucose levels in serum, fibrinogen in plasma and the smoking habit. RESULTS: H. pylori infection was present in 187/212 (88%) of the patients and in 183/310 (59%) of the control population (p < 0.0001). Classical risk factors for AMI did not differ among patients with and without H. pylori infection. CONCLUSION: Patients admitted to the Coronary Care Unit for acute myocardial infarction had a notably higher prevalence of H. pylori infection than the general population. The classical risk factors for coronary disease were equally present in all patients with AMI irrespective of H. pylori status. PMID- 10705707 TI - 31Phosphorus magnetic resonance spectroscopy to evaluate medical therapy efficacy in peripheral arterial disease. A pilot study. AB - BACKGROUND: Human and animal studies have shown that propionyl-L-carnitine, increasing carnitine content, improves the energy metabolism of ischemic skeletal muscle. The aim of the study was to evaluate the accuracy of Doppler continuous wave, Treadmill test and 31Phosphorus magnetic resonance spectroscopy in determining the efficacy of propionyl-L-carnitine in patients with peripheral arterial disease. METHODS: EXPERIMENTAL DESIGN: Prospective study. SETTING: University hospital. PATIENTS: Eighteen male patients with peripheral arterial disease (category 3) and 8 healthy volunteers form the basis of the study. Patients quit smoking, start physical training (2-3 Km walk per day) and were assigned to medical therapy consisting of propionyl-L-carnitine (8 patients) or placebo (10 patients). Patients were studied with Doppler continuous wave, Treadmill test and 31Phosphorus magnetic resonance spectroscopy at day 0 and at day 90. The following parameters were assessed by the principal component analysis: clinical (absolute claudication, ankle brachial index at rest and at 2, 5 and 10 minutes after completing Treadmill exercises) and biochemical (inorganic phosphorus/phosphocreatine ratio and pH profiles at 20% and 50% of the maximum load, the recovery half time of phosphocreatine, number of exercise steps and slope of linear relationship between muscle power and inorganic phosphorus/phosphocreatine ratio). RESULTS: Final evaluation showed a significant improvement of clinical and biochemical variables (p < 0.05 and p < 0.02 respectively). Breaking down the results on the basis of the two study arms, 31Phosphorus magnetic resonance spectroscopy showed a significant improvement of biochemical variables in the group of patients treated with propionyl-L-carnitine (p < 0.05) and was more sensitive in the evaluation of changes induced by 90-day treatment as compared with the other noninvasive examinations. CONCLUSIONS: 31P MRS permits the evaluation of muscle metabolic effect induced by PLC after a 90 day-period in patients affected by category 3 of peripheral arterial disease and it is a more sensitive tool in the evaluation of the pharmacological effects of medical therapy. PMID- 10705708 TI - Third-generation assays for hepatitis C antibodies: a four-year study of pattern changes in patients with chronic and past infection. AB - BACKGROUND: Many advances have been made in the sensitivity of assays for hepatitis C virus antibodies (HCV-Ab). Nevertheless, polymerase chain reaction (PCR) is still the best method to establish if infection has become chronic. In this study we utilised third-generation assays for HCV-Ab in a four-year follow up to determine the trend in antibody levels in currently and past infected patients. METHODS: Seventy-two multitransfused subjects were enrolled. All the patients were reactive at the first test with third-generation screening and confirmatory assays (ELISA-3 and RIBA-3) for HCV-Ab. They were subsequently retested in a follow-up ranging from 41 to 47 months. Viraemia was investigated with a standardised PCR kit; negative samples were reevaluated with nested PCR. Differences in antibody trend were calculated with the Wilcoxon signed-rank test. RESULTS: No statistical variation in antibody titre was found in the 41 HCV-RNA positive patients, although some of these showed a decrease in anti-c100p level. In contrast, anti-c22p, anti-c33c and anti-c100p levels decreased significantly in the 19 past infected patients. Twelve patients were HCV-RNA negative or intermittently positive with commercial PCR test, and consistently or intermittently positive in nested PCR: in these patients, antibody trend varied. CONCLUSIONS: Although resolving hepatitis is associated with a decrease in antibody titre, the trend should be observed for a long period to distinguish between chronic and past infection. However, the evaluation in a single patient can be unreliable. Since a doubtful response for HCV-RNA is in some cases obtained, further improvements in the diagnosis of chronic HCV infection are needed. PMID- 10705709 TI - Na+ channel blockers vs class III antiarrhythmic drugs in treating sustained ventricular tachycardia: reversing and preventing as different electrophysiological mechanisms. AB - METHODS: 169 selected patients with previous AMI and inducible sustained ventricular tachycardia (sVT) at electrophysiological study (EPS) were followed up prospectively for recurrent sVT during a five year period. At EPS, ventricular effective refractory period (VERP)/action potential duration (APd) ratio, ventricular conduction velocity, excitable gap, cycle length and QRS duration were measured. The patients with inducible sVT at basic programmed stimulation or after isoproterenol infusion (200 micrograms i.v.) underwent drug suppression tests (amiodarone, procamide, propanolol + procamide, amiodarone + propaphenone). On this basis, they were later assigned to 3 different groups: the amiodarone treated group (n = 112), the procamide-treated group (n = 22) and the nonresponder group (control group, n = 35). RESULTS: After procamide infusion (100 mg/min for 10 min), at fast pacing drive the VERP/APd ratio was significantly increased from baseline levels (p < 0.001), the conduction velocity (Vmax) was significantly depressed (by 25%, p < 0.005), the excitable gap was significantly reduced (p < 0.005, 23% of cycle length) and 71 patients were no longer inducible; during sinus rhythm, Vmax was significantly reduced from baseline values (by 14%, p < 0.05), VERP was only moderately increased and the excitable gap was slightly but significantly prolonged (p < 0.01, 53% of cycle length): 147 patients were still inducible, without significant difference from baseline values. After amiodarone infusion (300 mg/100 ml i.v. over 10 min), Vmax was significantly reduced from baseline levels at fast pacing drive (by 21%, p < 0.001), while the excitable gap did not reduce significantly (34%) from baseline levels and 91 patients were still inducible; during sinus rhythm, conduction velocity was moderately depressed (by 7%, p < 0.05), while VERP was significantly increased from baseline levels (p < 0.002) and the excitable gap was significantly reduced (p < 0.001, 29% of cycle length); only 35 patients were still inducible. In the remaining 134 patients the reset curve showed excitable gaps = 24 +/- 3% of the cycle length. The absolute and relative values of the excitable gap measured in the 35 patients who were still inducible were significantly higher than those measured in patients no longer inducible (p < 0.05). At follow-up, the sVT recurrence rate was: 28% in the amiodarone-treated group, 43% in the procamide-treated group and 36% in the control group: control data were significantly different from the former (p < 0.05) but not from the latter group. Significant correlation was reported between the plasmatic concentration of procamide and amiodarone and the percentage of reduction of Vmax at fast pacing drive (r = 0.79, p < 0.05; r = 0.83, p < 0.01). Resulting variability checked by the analysis of variance showed no significant difference between the two series (p < 0.12638). The development of conduction velocity depression was clearly dose-dependent. Drug suppression test with procamide + propanolol resulted in noninducibility of sVT in 115 patients, with strong significant difference from when the drug was used alone (p < 0.0015). Better results at EPS were obtained by the combination amiodarone + propaphenone, although no additional benefit was reported when compared with amiodarone alone. CONCLUSIONS: In conclusion, antiarrhythmic drugs with class I action may be highly effective in terminating sVI (lidocaine, procamide) but may be ineffective in preventing it or even arrhythmogenic. According to our data, drugs with class III antiarrhythmic action showed significantly different behavior: they were more effective in prolonging refractoriness and reducing the excitable gap at longer cycle lengths and, thus, capable of preventing, rather than terminating. Although statistical difference was reported between the amiodarone-treated group and the control group, the incidence of recurrent sVT remained too high to consider drug therapy as PMID- 10705710 TI - Nitrite plasma levels in normolipemic and hypercholesterolemic patients with peripheral occlusive arteriopathy. AB - BACKGROUND: The authors studied the nitrite plasma levels in a group of patients with peripheral obstructive arteriopathy. METHODS: The series consisted of 63 subjects (43 males, 20 females, mean age 64 +/- 9 years) suffering from peripheral arterial occlusive disease of the lower limbs, at II (55 cases) and III (8 cases) Fontaine stage; 21 subjects with total cholesterol (TC) lower than 200 mg/dl were considered as normolipemics, 24 subjects with TC values between 200 and 240 as mild hypercholesterolemics, 18 subjects with TC above 240 mg/dl as severe hypercholesterolemics. For each subject the determination of nitrite plasma levels was carried out, by the Gutman and Hollywood colorimetric method. RESULTS: In the normolipemic arteriopathics the basal value of nitrites was sharply reduced (p < 0.05) compared to the controls; in the mild hypercholesterolemics the mean basal value of nitrites was markedly higher compared to the controls; in the severe hypercholesterolemics the mean basal value of nitrites was statistically (p < 0.05) higher than that of the controls. In the arteriopathic patients, globally considered, the mean basal value of nitrites was superimposable on that of the normal control subjects. CONCLUSIONS: This study, carried out on the nitrite plasma levels in a group of arteriopathic patients allowed us to show the enhanced levels of nitric oxide due to the increase of LDL; this effect, previously observed in hypercholesterolemic diabetic and coronaropathic patients, leads us to the hypothesis of a stimulating effect of LDL upon NO endothelial synthesis; this would be a compensatory response to the damaging and vasoconstricting action of LDL. PMID- 10705711 TI - Bronchial reactivity in subjects with urticaria. AB - BACKGROUND: Respiratory symptoms (rhinitis, asthma) accompany some skin diseases with a proven immunological or allergic pathogenesis whereas the immunological and allergological findings are sometimes negative in subjects with chronic urticarial skin symptoms and no cause can be found. This paper explores the possibility of airway inflammation in cases of this kind. METHODS: Nonspecific bronchostimulation with methacholine was carried out in 12 subjects with a clinical history of urticaria and negative immunological and allergological findings. RESULTS: All subjects displayed normal bronchoreactivity (mean PD20 1400 micrograms). The bronchostimulation test (evaluated from the FEV1) was always negative. In the control group the mean PD20 was 720 micrograms. CONCLUSIONS: The fact that these eleven subjects did not display bronchial hyperreactivity suggests that monitoring of their respiratory function is not required. PMID- 10705712 TI - T-cell activity in HLA-associated autoimmune diseases. AB - BACKGROUND: The existence of T-helper-1 (Th1) and T-helper-2 (Th2) subsets has been implicated in the regulation of several immune responses, and alterations in the Th1/Th2 balance have been involved in autoimmunity. The present study investigates the relative influence of Th1 and Th2 patterns in autoimmune responses in patients with HLA-associated autoimmune diseases. METHODS: This study concerns 849 patients of both sexes, suffering from several autoimmune diseases. Tissue typing for HLA antigens of Class I (A, B, C) and Class II (DR, DQ) was carried out in all patients by conventional serologic methods, comparing results with frequencies detected in a normal population. Many immunological tests were also done. In particular, lymphocyte subsets (CD4+, CD8+, CD3-HLA-DR+, NK cells, sIg + B cells) were detected with monoclonal antibodies by a fluorescent cytometer. The changes in frequencies of T cell subsets were used to calculate the possible incidence of two effector phenotypes (TE-1; TE-2). RESULTS: The results of the immunogenetic analysis confirmed the significant HLA associations in several diseases. The essential T-cell changes were also exposed, thus defining the incidence of T-cell phenotypes (TE-1 = 56.3%; TE-2 = 34.8%). This finding suggested a major impact of cell-mediated immunity, as compared with that of antibody-mediated immunity. CONCLUSIONS: The anomalies of Th1/Th2 balance can impact autoimmune disease, and in many cases a Th2 response can prevent Th1 mediated autoimmunity, which is the most evident phenomenon in several HLA associated diseases. PMID- 10705713 TI - Cytotoxic effector function in HLA-associated autoimmune diseases. AB - BACKGROUND: The mechanisms by which cytotoxic-T-lymphocytes (CTLs) and natural killer (NK) cells recognize and kill target cells are the subject of intense research. Previous observations in patients with HLA-associated autoimmune diseases show a major impact of cell-mediated immunity as compared to antibody mediated immunity. This study analyzes the possible co-operation of NK cells in the autoimmune process, comparing their increase to that of cytotoxic-T lymphocytes. METHODS: This research examines the incidence of CD8+ T cells and NK cells increases in 1065 patients with various autoimmune diseases. Tissue typing for HLA antigens was carried out in all patients by conventional serologic methods. Lymphocyte subsets (CD4+, CD8+, CD3-HLA-DR+, NK cells, sIg+ B cells) were detected with monoclonal antibodies by a fluorescent cytometer. Results were statistically compared to normal control values by the Student's "t"-test. RESULTS: The analysis of results shows the incidence of CTLs and NK cell increases in patients with different autoimmune syndromes. In some diseases (uveitis, multiple sclerosis, and other neurological autoimmune disorders) raised frequencies of NK cell increase are most evident. These results suggest some possible relationships between CTLs and NK cells in the pathogenesis of the autoimmune process. CONCLUSIONS: The present results show that CTLs and NK cells are implicated in immune dysregulation, which characterizes the complex pathogenesis of autoimmune diseases. PMID- 10705714 TI - Effects of ATP-MgCl2 administration in hypovolemic dogs. AB - BACKGROUND: The aim of the study was to evaluate the efficacy of ATP-MgCl2 on myocardial insufficiency associated with hypovolemic shock in dogs. We designed the study as a controlled randomized study. METHODS: Six mixed-breed dogs weighing 22 +/- 3 kg were included in the control group and 20 +/- 3 kg in the ATP-MgCl2 group. After the animals were anesthetized 40 ml/kg of blood was withdrawn in 15 minutes. Animals were observed for 45 minutes after removal of blood. Six animals received 45 ml/kg of lactated Ringer's solution and the other animals were treated with 45 ml/kg of lactated Ringer's solution and ATP-MgCl2. All measurements were made before removal of blood, 45 min after exsanguination and at 1 hour intervals for 3 hours. The following parameters were measured; systemic and pulmonary arterial pressures, pulmonary capillary wedge pressure, central venous pressure, cardiac output, rectal temperature, arterial pH, PCO2 and PO2 and mixed venous hemoglobin oxygen saturation. In addition blood samples were collected for the analysis of lactate and tumor necrosis factor (TNF) concentrations. RESULTS: After hemorrhage, cardiac index (CI) decreased significantly from 122 +/- 9 to 52 +/- 9 ml/kg/min in the control group (p < 0.0001) and from 124 +/- 11 ml/kg/min to 50 +/- 6 ml/kg/min in the ATP-MgCl2 group, respectively (p < 0.0001). After volume replacement, Cl was 93 +/- 6 ml/kg/min in the control group and 111 +/- 4 ml/kg/min in the ATP-MgCl2 group 3 hours after the onset of reinfusion, respectively (p < 0.05). TNF was 36 +/- 5 pg/ml in the control group and 21 +/- 3 pg/ml in the ATP-MgCl2 group (p < 0.05). Three hours after the onset of hemorrhagic shock, oxygen consumption and delivery were 126 +/- 14 and 206 +/- 19 ml/min in the control group and 198 +/- 16 and 305 +/- 27 ml/min in the ATP-MgCl2 group, respectively. At the same time point the oxygen extraction ratio was 0.49 +/- 0.04 in the control group and 0.61 +/- 0.03 in the ATP-MgCl2 group (p < 0.01). CONCLUSIONS: Hemorrhagic shock causes TNF release which may cause multiple organ failure. Organ dysfunction still persists even after the appropriate treatment. ATP-MgCl2 attenuates the release of TNF which may improve the adverse effects of hemorrhagic shock. PMID- 10705715 TI - Reanimation of the heart after death. AB - BACKGROUND: The purpose of this study was to determine if hearts could be successfully reanimated after death in an extra corporeal circuit. METHODS: Reanimation of pig and dog heart was accomplished in an extra corporeal circuit after death by lethal injection of Pentothal (euthanasia). A total of 42 experiments were performed. RESULTS: The most successful combination included enhancement of the cardioplegic and reperfusion solutions with amino acids, calcium and magnesium ions, and using the animals own drug free blood in the reperfusion solution. Successful resuscitation of the heart was accomplished after ischemic cold storage as long as 9 hours after death. A beating heart was sustained by the extra corporeal circuit for over 10 hours (635 minutes). CONCLUSIONS: In summary, successfully reanimation of the heart in an extra corporeal circuit is possible when cold cardioplegic solution is administered 10 35 minutes after death by lethal injection of Pentothal. PMID- 10705716 TI - Free radicals: important cause of pathologies refer to ageing. AB - Free radical are highly reactive chemical species with an unpaired electron in an atomic or molecular orbital. In biological systems, the most important free radicals are superoxide anion and hydrogen peroxide; in the presence of transition metals such as iron, copper and manganese both these free radicals produce hydroxyl radicals. Free radicals attack proteins, nuclei acids and membranes containing large quantities of polyunsaturated fatty acids. Because of their toxicity, the organism has developed ways to deactivate them. The superoxide dismutase enzyme (SOD) catalyzes dismutation of the superoxide radical into hydrogen peroxide and oxygen hydrogen peroxide is in turn reduced to water and oxygen by peroxidase glutathione and catalase enzymes. The production of radicals in the brain is due to catecholamine metabolism such as dopamine and norepinephrine and is increased by the presence of transition metals and by a deficiency of antioxidant agents such as vitamin E. Two main groups of dementia exist in older age: the multi-infarctual dementias, caused by cerebrovascular disorders and the primary degenerative disorders such as Alzheimer, where no vascular disease is evident. Free radicals play an important role in Parkinson's disease, in Alzheimer's disease and in stroke. The value of SOD and CAT activity following the above mentioned degenerative diseases differ among the various studies carried out. In Alzheimer's disease, the value of SOD activity probably increases in the neuropathologically involved areas. In stroke, the SOD value does not vary either in the ischemic area or in the peri-infarctual one during the first 24 hrs after lesion, while the CAT value decreases. PMID- 10705717 TI - Shoulder arthroplasty. Indications, contraindications and complications. AB - Prosthetic substitution of the glenohumeral joint, begun at the end of the last century, has developed greatly in recent years. Today the most widely used shoulder prostheses are defined as "modular" because of their extensive adaptability. The capacity to adapt to anatomic variations must be incorporated within their structure, and normal articular biomechanics must be respected. The choice of prosthesis must be based on the condition of the joint surfaces, on the anatomic and functional condition of the rotator cuff. So endoprosthesis of the shoulder is indicated for avascular necrosis of the head of the humerus, fractures and pseudoarthrosis of the extreme proximal end of the humerus, arthropathy following rotator cuff rupture. Total shoulder prosthetization is indicated for glenohumeral osteoarthritis, rheumatoid arthritis and outcomes of endoprosthesis. The principal contraindications for shoulder replacement include an infection in progress, Charcot's arthropathy and severe neurological pathologies. The complications of shoulder prostheses include infection, dislocation, loosening of a component, periprosthetic humeral and glenoid fractures, nerve injuries, fractures of a prosthetic component and ectopic ossification. At present prosthetic substitution of the glenohumeral joint offers good results. Indispensable conditions for these results include anatomic and functional integrity of the musculature, good bone quality, correct positioning of the prosthetic components and a proper rehabilitation program. PMID- 10705718 TI - Congenital hypopituitarism in a 48-year old adult. Natural course, hormonal study and MRI evidence. AB - A case of Congenital Hypopituitarism (CH) in an untreated 48 yr-old-man is reported. The hormonal studies demonstrated a panhypopituitarism and MR imaging revealed absence of pituitary stalk, small anterior pituitary remnant on the sella floor and ectopic neurohypophysis at the tuber cinereum. The pattern of hormonal responsiveness suggests that CH encompasses findings typical of primary anterior pituitary disease and those of hypothalamic dysfunction. PMID- 10705719 TI - Takayasu's arteritis on steroid therapy. Seven years follow-up. AB - The authors report a 7 year follow-up of Takayasu's arteritis (TA) type III, group 1, in a young Italian woman. At diagnosis, at the age of 25, the echotomographic and angiographic studies showed narrow subclavian arteries, narrow abdominal aorta (diameter of 0.6-0.8 cm) below the renal arteries, stenotic left common carotid and renal arteries, and occluded upper mesenteric artery. With steroid therapy, (prednisone 50 mg/day per os), the erythrocyte sedimentation rate (ESR) normalized within 12 days. With a maintenance dosage of 7.5 mg/day per os, the patient achieved remission as documented by the absence of symptoms, the persistent normalization of ESR, and the improving of the diameter of the abdominal aorta (1.3-1.4 cm). On steroid therapy, the patient had a normal pregnancy and delivered a healthy baby girl. The disease has been stable for seven years. Recently, diabetes mellitus occurred and it has been treated with insulin therapy. The rising of ESR after tapering of steroid therapy (prednisone 5 mg per os on alternate days) suggests an alternative treatment with a cytotoxic agent. PMID- 10705720 TI - Pulmonary hamartoma. A rare case report. AB - Pulmonary hamartoma is a rare lung neoformation, usually symptomless and by chance discovered, of a probable dysontogenetic origin with prevailing cartilaginous tissue and adult, onset age. The Authors report a rare case of a 25 year-old student, symptomless and fortuitously found by means of a radiograph of the chest. Many interesting features characterize the case report: histological nature of the pulmonary hamartoma, mainly vascular, so much as to feign an angiosarcoma at the macroscopical examination, and with small peripheral calcifications as shown by lung CT scan; the measures (about 7 cm) plentifully above the parameters usually reported in the literature (from 2 cm to 4 cm); the young onset age (about 10 years old). We may consider a case exceptionally reported in the literature. Besides, on the base of a few studies and of our experience, the results of the pulmonary hamartoma growth rate and doubling time are reported. PMID- 10705721 TI - Extralymphonodal Castleman's disease. A case report. AB - Castleman's disease is a rare lymph node pathology characterized by angiofollicular hyperplasia. There are two forms of the disease: localized and systemic, with different features, symptoms and prognosis. Three are the histological types of disease: plasma cell, hyaline-vascular and mixed variants. We report the case of a 65-year-old female affected by localized plasma cell variant of Castleman's disease. The singularity of our case lies in its localization on the breast and monoclonal plasma cell proliferation inside the nodule. PMID- 10705722 TI - Tuberculosis and Pelger-Huet anomaly. Case report. AB - We describe a patient with pulmonary tuberculosis and a rare disturbance of leukocyte segmentation, known as "Pelger-Huet anomaly", which can be observed in various diseases such as malignancies and/or infections. The importance of this association is equivocal: some authors have related to the association the particular severity of tuberculosis or the death they observed; in the case reported we noted no evidence of such a relation, notwithstanding the presence of the homozygous form of the Pelger-Huet anomaly. We suggest therefore that, when Pelger-Huet anomaly is found, an underlying disease should be searched for; the course of this illness, however, might not be affected. PMID- 10705723 TI - The new techniques of gynaecologic laparoscopy. Gasless, open Hasson, optic trocar. AB - BACKGROUND: New techniques of laparoscopy: gasless, open Hasson, optic trocars allow to avoid the risks of vessel and bowel injuries. The objective of this study was to evaluate the capability of a retractor system as an alternative to conventional technique without pneumoperitoneum and to assess if the system facilitates the use of conventional surgical instruments during gynaecological surgery. METHODS: DESIGN: Prospective evaluation. SETTING: University-affiliated county hospital. PATIENTS: Gasless laparoscopy surgery was performed on 49 patients between December 1995 and July 1996 with a retractor system without pneumoperitoneum consisting of an intrabdominal retractor using conventional surgical and laparoscopic instruments and to enable a simultaneous vaginal approach. RESULTS: Gasless laparoscopy was successful in 44 (90%) of cases. A simultaneous vaginal approach was used in one third of indications including vaginal myomectomy and laparoscopic assisted hysterectomy. Conversion to laparotomy was required in 5% of cases. Mean procedure duration was 90 minutes and mean hospitalisation time was 5.7 days. CONCLUSIONS: The introduction of new techniques of laparoscopy: gasless, open Hasson, optic trocars has broadened the application of operative laparoscopy. Gasless technique in lieu of conventional laparoscopy can be performed reliably and safely for most gynaecological indications. The most outstanding benefit of this method is that it can be combined with a vaginal approach which is not possible using a pneumoperitoneum due to gas leakage. The place of gasless laparoscopy will depend on continuing development by instrument manufacturers, in order to achieve an instrument providing vision as good as that seen with the pneumoperitoneum. PMID- 10705724 TI - Flavonoids from Hypericum perforatum show antidepressant activity in the forced swimming test. AB - It has been shown recently that a flavonoid fraction (fraction II) obtained from a crude extract of Hypericum perforatum (St. John's Wort) was remarkably active in the forced swimming test (FST). Fraction II was further separated using MLCCC to give fractions IIa and IIb. Both fractions proved to be active in the FST at different dosages. Further separation of fraction IIa by preparative HPLC yielded fraction IIa1 which mainly was composed of hyperoside, isoquercitrin, miquelianin and quercitrin, and fraction IIa2 which contained small amounts of hyperoside and astilbin, while most compounds were not known. Both fractions were active after acute treatment in the FST. Isolates obtained from these fractions including hyperoside, isoquercitrin, quercitrin, miquelianin, the aglycone quercetin and astilbin, were tested for activity in the FST. Except for quercetin, quercitrin and astilbin all compounds were active. To exclude false positive results in the FST the validity was checked in open field experiments and in the FST after 12 days of daily treatment. PMID- 10705725 TI - Evidence for specific interactions between kavain and human cortical neurons monitored by fluorescence correlation spectroscopy. AB - Specific interactions between a tetramethylrhodamine-labeled kavain derivative and human cortical neurons were found for the first time using fluorescence correlation spectroscopy (FCS). Human cortical neurons, cultivated in Nunc chambers, were incubated with 1 nM of the dye-labeled kavain derivative. A total binding of 0.55 nM was found after an incubation period of 60 min. 50% of the total binding was specifically displaced in the presence of 1 microM non-labeled (+)-kavain. Evidence for these specific interactions was verified by a saturation experiment. Both the non-linear Scatchard plot and the n value of 1.58 +/- 0.07 in the sigmoid Hill plot indicate binding sites with different binding affinities. PMID- 10705726 TI - Inhibition of tyrosinase by flavonoids, stilbenes and related 4-substituted resorcinols: structure-activity investigations. AB - Several flavonoids, stilbenes and related 4-substituted resorcinols, obtained from Artocarpus incisus and other plants or synthesized, were tested for their inhibitory activity against tyrosinase. The structure-activity relationships suggested that specific natural or synthesized compounds having the 4-substituted resorcinol skeleton have potent tyrosinase inhibitory ability. Kinetic studies have indicated that specific compounds having the 4-substituted resorcinol skeleton exhibit competitive inhibition of the oxidation of DL-beta-(3,4 dihydroxyphenyl)alanine (DL-DOPA) by mushroom tyrosinase. These findings could lead to the design and discovery of new tyrosinase inhibitors. PMID- 10705727 TI - The 5 alpha-reductase inhibitory components from heartwood of Artocarpus incisus: structure-activity investigations. AB - The methanol extract of heartwood of Artocarpus incisus showed potent 5 alpha reductase inhibitory activity. We investigated the 5 alpha-reductase inhibitory effects of nine compounds isolated from A. incisus. Chlorophorin (IC50 = 37 microM) and artocarpin (IC50 = 85 microM) showed more potent inhibitory effects than did alpha-linolenic acid, which is known as a naturally occurring potent inhibitor. Structure-activity investigations suggested that the presence of an isoprene substituent (prenyl and geranyl) would enhance 5 alpha-reductase inhibitory effects. PMID- 10705728 TI - Lignin-carbohydrate complexes: intestinal immune system modulating ingredients in kampo (Japanese herbal) medicine, juzen-taiho-to. AB - An extract preparation (TJ-48) of a Japanese herbal (Kampo) medicine, Juzen-Taiho To has been found to show an enhancing activity on proliferation of bone marrow cells mediated by Peyer's patch cells (intestinal immune system modulating activity) (1). When TJ-48 was fractionated by MeOH- and water-extractions, EtOH precipitation and dialysis, water-soluble dialyzable (F-3) and polysaccharide fractions (F-5) both showed a significant intestinal immune system modulating activity in vitro; other fractions had no activity. Oral administration of F-3 (150 mg/kg) also showed the intestinal immune system modulating activity in C3H/HeJ mice. Two active substances were purified by gel filtration from F-3. Chemical analysis indicated that the active substances comprised lignin as well as a carbohydrate consisting mainly of arabinose, galactose, glucose and galacturonic acid. Activity was significantly reduced not only by NalO4 oxidation but also by NaClO2 treatment. Standard lignin showed no activity. Carbohydrate and lignin in the active substances were co-eluted by hydrophobic-interaction chromatography on phenyl-Sepharose, and the stimulating effect and lignin content of the substances did not change even after the hydrophobic-interaction chromatography. These results suggest that lignin-carbohydrate complexes are involved in intestinal immune system modulation by TJ-48. PMID- 10705729 TI - Effects of luteolin and other flavonoids on IgE-mediated allergic reactions. AB - The anti-allergic action of luteolin was investigated in the rodent experimental allergic models. In the present study, the effects of luteolin were compared to those of baicalein, quercetin, and prednisolone. Luteolin as well as baicalein, quercetin, and prednisolone inhibited the IgE antibody-mediated biphasic cutaneous reaction (immediate phase reaction and late phase reaction) in mice. However, these compounds did not affect the histamine-, serotonin-, and platelet activating factor-induced cutaneous reactions in rats. In an in vitro study, luteolin, baicalein, and quercetin inhibited IgE-mediated histamine release from bone marrow-derived cultured murine mast cells (BMMC) and rat peritoneal mast cells. These compounds also inhibited IgE-mediated TNF-alpha and IL-6 production from BMMC. From these results, luteolin inhibited the IgE-mediated biphasic cutaneous reaction mainly by the inhibition of histamine and cytokine release from mast cells, but not through mediator antagonistic effects. PMID- 10705730 TI - Antiplasmodial activity of Cryptolepis sanguinolenta alkaloids from leaves and roots. AB - The roots of Cryptolepis sanguinolenta have been investigated for their chemical composition since 1931 but so far no studies on the leaves have been reported although they are used in traditional medicine in Guinea-Bissau. Two new alkaloids identified as cryptolepinoic acid (1) and methyl cryptolepinoate (2) and the known alkaloids cryptolepine (4), hydroxycryptolepine (5/5a) and quindoline (6), were isolated from the ethanolic and chlorophormic leaf extracts. Aqueous and ethanolic extracts of the leaves and roots and seven alkaloids isolated from those extracts were tested in vitro against Plasmodium falciparum K1 (multidrug-resistant strain) and T996 (chloroquine-sensitive clone). All the extracts were shown to give 90% inhibition of P. falciparum K1 growth at concentrations < 23 micrograms/ml. Cryptolepine (4) was the most active alkaloid tested with IC50 values (0.23 microM to K1; 0.059 microM to T996) comparable with chloroquine (0.26 microM to K1; 0.019 microM to T996). The indolobenzazepine alkaloid cryptoheptine (7) was the second most active with IC50 values of 0.8 microM (K1) and 1.2 microM (T996). Cryptolepinoic acid (1) showed no significant activity while its ethyl ester derivative 3 was active against P. falciparum K1 (IC50 = 3.7 microM). All the indoloquinoline alkaloids showed cross-resistance with chloroquine but not the indolobenzazepine alkaloid 7. It was noticed that alkaloids with weakly basic characteristics were active whereas other structurally related alkaloids with different acid-base profiles were inactive. These observations are in agreement with the antimalarial mechanism of action for quinolines. PMID- 10705731 TI - Cardiovascular effects of visnagin on rats. AB - The present article describes the effects of visnagin on systolic blood pressure and heart rate in the anaesthetized rat. Intravenous administration of visnagin (0.3-5 mg kg-1) produced dose-related decreases in blood pressure with no significative changes in heart rate. Under nitric oxide synthase inhibition (L NAME, 50 mg kg-1) the hypotensive effects of visnagin (5 mg kg-1) were not affected. Visnagin (5 x 10(-6) M-10(-4) M) produced a weak decrease in the rate and amplitude of spontaneous contractions in right atria. Visnagin also caused a weak decrease in peak contractile force and the df/dtmax with no significant changes in the time to peak tension or the time for total contraction in left atria driven at a basal rate of 1 Hz. Visnagin (10(-5) M, 5 x 10(-5) M and 10(-4) M) concentration-dependently decreased pressor response to KCl (IC50 = 5.1 +/- 2.5 x 10(-5) M) and noradrenaline (IC50 = 2.6 +/- 0.9 x 10(-5) M) in rat isolated mesenteric beds. Visnagin (3 x 10(-7) M-10(-4) M) induced a concentration dependent relaxation of isolated mesenteric arteries contracted by noradrenaline (IC50 = 1.7 +/- 0.8 x 10(-5) M). The relaxant effects in the absence of functional endothelium were not significantly different (IC50 = 1.5 +/- 0.3 x 10( 5) M, P > 0.05) from those observed in segments with intact endothelium. In conclusion, the main mechanism responsible for the acute hypotensive effect of visnagin is the vasorelaxant response induced by this drug in resistance arteries. PMID- 10705732 TI - Beta-glucuronidase inhibitory activity and hepatoprotective effect of 18 beta glycyrrhetinic acid from the rhizomes of Glycyrrhiza uralensis. AB - An inhibitor of beta-glucuronidase from the rhizomes of Glycyrrhiza uralensis was isolated and its hepatoprotective activity on CCI4-induced hepatotoxicity of rats was investigated. From the water-soluble extract of G. uralensis, glycyrrhizin was isolated as a potent inhibitor of beta-glucuronidase. When glycyrrhizin was orally administered, it had a hepatoprotective activity. However, when glycyrrhizin was intraperitoneally administered, it did not have a hepatoprotective activity. 18 beta-Glycyrrhetinic acid, which is a major metabolite of glycyrrhizin by human intestinal bacteria, was also a potent inhibitor of beta-glucuronidase. When 18 beta-glycyrrhetinic acid was intraperitoneally administered, it also had some hepatoprotective activity. These results suggest that glycyrrhizin may be a natural prodrug for the observed hepatoprotective effect in rats and that serum beta-glucuronidase levels have implications for the liver injury, as reductions of its activity by administration of inhibitors such as G. uralensis or its derived products and silymarin correlate with reductions in biochemical indices of liver injury. PMID- 10705733 TI - Antiproliferative effect on human prostate cancer cells by a stinging nettle root (Urtica dioica) extract. AB - In the present study the activity of a 20% methanolic extract of stinging nettle roots (Urtica dioica L., Urticaceae) on the proliferative activity of human prostatic epithelial (LNCaP) and stromal (hPCPs) cells was evaluated using a colorimetric assay. A concentration-dependent and significant (p < 0.05) antiproliferative effect of the extract was observed only on LNCaP cells during 7 days, whereas stromal cell growth remained unaltered. The inhibition was time dependent with the maximum of growth reduction (30%) at a concentration of 1.0E-6 mg/ml on day 5 compared to the untreated control. On day 4 and 6, the reduction in proliferation of LNCaP cells showed the minimal effective dose at 1.0E-9 mg/ml. No cytotoxic effect of ME-20 on cell proliferation was observed. The antiproliferative effect of ME-20 of stinging nettle roots observed both in an in vivo model and in an in vitro system clearly indicates a biologically relevant effect of compounds present in the extract. PMID- 10705734 TI - Evidence for bioadhesive effects of polysaccharides and polysaccharide-containing herbs in an ex vivo bioadhesion assay on buccal membranes. AB - Aqueous extracts of polysaccharide-containing plants are widely used in therapy for irritated mucus membranes in the pharynx region. In order to prove the existence of mucilaginous effects of polysaccharide hydrocolloids on epithelia an ex vivo system based on porcine buccal membranes was established. The tissue culture was stable and there was no indication of cytolytic processes during the 5 hour incubation period. This was confirmed through histological studies and the respective LDH values as toxicity marker. The test system was shown to discriminate the adhesive effects of different raw polysaccharides, obtained from a variety of medicinal plants. While polysaccharides from Altheae officinalis, Plantago lanceolata, Malva moschata, or Tilia cordata showed only moderate bioadhesion to epithelial tissue, strong adhesive processes were observed with polysaccharides from Fucus vesiculosus and Calendula officinalis. The adhesive effects were concentration-dependent. Histological studies of membranes, incubated with a fluorescence-labelled rhamnogalacturonan, indicated the presence of distinct polysaccharide layers on the apical membrane surface. With these results, adsorption effects of certain polysaccharides on mucus membranes were shown for the first time. Such effects suggest that this may account, at least in part, for the therapeutic effects of mucilage-containing plants in the treatment of irritated buccal membranes. PMID- 10705735 TI - Alkamide levels in Echinacea purpurea: effects of processing, drying and storage. AB - The effects of processing, drying, and storage time and temperature on alkamide levels in Echinacea purpurea roots are described. Chopping altered the levels of some alkamides slightly, whereas drying had no effect. Levels of all alkamides fell by over 80% during storage at 24 degrees C for 64 weeks. Alkamide levels also dropped significantly during storage at -18 degrees C. PMID- 10705736 TI - Seasonal variation of artemisinin and its biosynthetic precursors in plants of Artemisia annua of different geographical origin: proof for the existence of chemotypes. AB - The time course of the levels of artemisinin, its biosynthetic precursors and the biosynthetically related sesquiterpenes was monitored during a vegetation period of Artemisia annua plants of different geographical origin. Considerable differences in contents of artemisinin and its direct precursors artemisinic acid and dihydroartemisinic acid were found between these A. annua's. For the first time the A. annua plants of different geographical origin were found to belong to different chemotypes. A chemotype with a high artemisinin level was found to have also a high dihydroartemisinic acid level but a relatively low artemisinic acid level. Reversibly, a chemotype with low levels of artemisinin and dihydroartemisinic acid contained a high artemisinic acid level. Artemisinic acid is considered to be the direct precursor of dihydroartemisinic acid in the biosynthetic pathway of artemisinin. The observed accumulation of artemisinic acid in one of the A. annua chemotypes may indicate the presence of a rate limiting step in the biosynthetic pathway of artemisinin. The enzymatic reduction of artemisinic acid into dihydroartemisinic acid is probably a "bottle neck" in the biosynthetic pathway of artemisinin in varieties with high artemisinic acid and consequentially low artemisinin levels. After a night-frost period, the level of artemisinin was increased, in the Vietnamese A. annua plants, while the dihydroartemisinic acid level was decreased. This phenomenon is in accordance with our hypothesis that stress triggers the conversion of dihydroartemisinic acid to artemisinin. It is suggested that the presence of high levels of dihydroartemisinic acid may be an adaptation to stress conditions (e.g., night frost), during which relatively high levels of 1O2 are formed. Dihydroartemisinic acid gives the plant protection by reacting with these reactive oxygen species yielding artemisinin as stable end-product. PMID- 10705737 TI - Cytotoxins, mycotoxins and drugs from a new deuteromycete, Acremonium neo caledoniae, from the southwestern lagoon of New Caledonia. AB - A new cytotoxic trichothecene sesquiterpene, verrol 4-acetate, was isolated, along with known macrocyclic trichothene mycotoxins and medicinal styrylpyrones, from cultures of a new deuteromycete Acremonium neo-caledoniae Roquebert et Dupont n. sp., taken from drifting wood in the southwestern lagoon of New Caledonia. PMID- 10705738 TI - Steroids from Harrisonia abyssinica. AB - In addition to the known sterols and ketosteroids beta-sitosterol (24 alpha ethylcholest-5-en-3 beta-ol), stigmasterol (24 alpha-ethylcholesta-5,22-dien-3 beta-ol), campesterol (24 alpha-methylcholest-5-en-3 beta-ol), beta-sitostenone (stigmast-4-en-3-one, 24 alpha-ethylcholest-4-en-3-one), stigmastenone (stigmasta 4,22-dien-3-one, 24 alpha-ethylcholesta-4,22-dien-3-one), campestenone (24 alpha methylcholest-4-en-3-one), and stigmasta-3,5-dien-7-one (24 alpha-ethylcholesta 3,5-dien-7-one), the new steroids stigmasta-3,5,22-trien-7-one (24 alpha ethylcholesta-3,5,22-trien-7-one), and campesta-3,5-dien-7-one (24 alpha methylcholesta-3,5-dien-7-one) were isolated from the stem bark of Harrisonia abyssinica and identified by NMR and mass spectrometry. PMID- 10705739 TI - Baicalein: an in vitro antigenotoxic compound from Scutellaria baicalensis. AB - Bioassay-guided fractionation of an H2O extract of the root of Scutellaria baicalensis has furnished an in vitro antigenotoxic flavonoid, baicalein (1) and 2',5,5',7-tetrahydroxy-6',8-dimethoxyflavone (2). Compound 1 exhibited a dose dependent inhibition of aflatoxin B1 (AFB1) and N-methyl-N'-nitro-N nitrosoguanidine mutagenicity in the Salmonella typhimurium bacterial mutation assay. In the chromosome aberration assay, compound 1, at a concentration of 5 microM, reduced the frequency of chromosome aberration induced by AFB1 but increased the clastogenic effect of AFB1 at a concentration of 50 microM. PMID- 10705740 TI - Flavonol glycoside gallate and ferulate esters from Persicaria lapathifolia as inhibitors of superoxide production in human monocytes stimulated by unopsonized zymosan. AB - Aerial parts of Persicaria lapathifolia S.F. Gray (Polygonaceae) exhibited an inhibitory effect on superoxide production in unopsonized zymosan-stimulated human monocytes. Two known compounds, quercetin 3-O-beta-(2"-galloyl) glucopyranoside and quercetin 3-O-beta-(2"-galloyl)-rhamnopyranoside, and a new compound, quercetin 3-O-beta-(6"-feruloyl)-galactopyranoside, were isolated as the inhibitors of superoxide production by activity-guided fractionation. IC50 values were shown at the concentrations of 2.1 microM by quercetin 3-O-beta-(2" galloyl)-glucopyranoside, 1.9 microM by quercetin 3-O-beta-(2"-galloyl) rhamnoypranoside, and 3.5 microM by quercetin 3-O-beta-(6"-feruloyl) galactopyranoside whose inhibitory potencies were similar to oxyphenylbutazone (IC50 = 1.9 microM) as a positive control. PMID- 10705741 TI - Potentiating effect of obacunone from Dictamnus dasycarpus on cytotoxicity of microtuble inhibitors, vincristine, vinblastine and taxol. AB - The limonoid triterpene, obacunone, was found to enhance the cytotoxicity of vincristine against L1210 cells by approximately 10-fold. Further, it was found that the cytotoxicity of other microtubule inhibitors such as vinblastine and taxol in drug-sensitive KB-3-1 cells as well as in multidrug-resistant KB-V1 cells was enhanced greatly in the presence of obacunone. On the other hand, there was no remarkable potentiating effect of obacunone on the cytotoxicity of other antineoplastic drugs such as adriamycin, cisplatin or 5-fluorouracil. From these results, it is implied that the potentiating action of obacunone may be limited to microtubule inhibitors. PMID- 10705742 TI - Hyaluronidase inhibitory active 6H-dibenzo[b,d]pyran-6-ones from the feces of Trogopterus xanthipes. AB - Bioassay-guided fractionation of the MeOH extract of Pteropi faeces (the feces of Trogopterus xanthipes Milne-Edwards) furnished three hyaluronidase inhibitory active 6H-dibenzo[b,d]-pyran-6-ones (1-3), together with a new compound, 3,8,10 trihydroxy-6H-dibenzo[b,d]pyran-6-one (4). Their structures were established on the basis of the spectroscopic methods. PMID- 10705743 TI - Aromatase and sulfatase inhibitors from Lepiota americana. AB - From an edible mushroom Lepiota americana Pk., (Agaricaceae), 2-aminophenoxazin-3 one that inhibited aromatase at IC50 = 5.7 microM and 3 beta-hydroxy-5,8 epidioxyergosta-6,22-diene that inhibited sulfatase at IC50 = 0.9 microM were isolated. Neither 2-aminophenoxazin-3-one was active against sulfatase nor was 3 beta-hydroxy-5,8-epidioxyergosta-6,22-diene active against aromatase. PMID- 10705744 TI - Furoquinolines with antiplatelet aggregation activity from leaves of Melicope confusa. AB - Using antiplatelet aggregation as a guide for fractionation, four furoquinoline type alkaloids, confusameline (1), skimmianine (2), kokusaginine (3), and O methylconfusameline (4), were isolated from the leaves of Melicope confusa. All compounds showed significant antiplatelet aggregation activity. PMID- 10705745 TI - Antihyperglycemic acetylenic glucosides from Bidens pilosa. AB - In vivo bioassay-guided fractionation of the aqueous alcohol extract of the aerial parts of Bidens pilosa Sch. Bip. var. radiata (Asteraceae) using C57 BL/Ks db/db mice as a model for type 2 diabetes, yielded two known polyacetylenic glucosides, identified as 2-beta-D-glucopyranosyloxy-1-hydroxy-5(E)-tridecene 7,9,11-+ ++triyne (1) and 3-beta-D-glucopyranosyloxy-1-hydroxy-6(E)-tetradecene 8,10,1 2-triyne (2). A 3:2 mixture of compounds 1 and 2 effected a significant drop in blood glucose. PMID- 10705746 TI - Antinociceptive activity of I3,II8-binaringenin, a biflavonoid present in plants of the guttiferae. AB - This paper describes the antinociceptive action of 13,118-binaringenin (GB-1a), a biflavonoid isolated from Clusia columnaris and present in several plants of the family Guttiferae, in a writhing test and a formalin test. It was found that it exhibits potent and dose-related antinociceptive action in both experimental models, with ID50 values of 22 mumol/kg against the writhing test and 28 mumol/kg against the second phase of the formalin test. It was more potent than some well known analgesic drugs used as reference. Based on a hot-plate test its mechanism of action seems to be unrelated with the opioid receptors. PMID- 10705747 TI - Activity of solidagenone and their semisynthetic derivatives on the glucocorticoid-mediated signal transduction. AB - The labdane diterpene solidagenone and four semisynthetic derivatives were assessed for effects on the glucocorticoid-mediated signal transduction. Solidagenone and the derivatives proved to be active with IC50 values between 1 and 25 micrograms/mL. All compounds were cytotoxic towards L 1210, BHK and COS 7 cells with IC50 from 10-100 micrograms/mL. PMID- 10705748 TI - Inhibitory effects of lignans and flavonoids in saiboku-to, a herbal medicine for bronchial asthma, on the release of leukotrienes from human polymorphonuclear leukocytes. AB - To identify the anti-allergic components contained in Saiboku-To, a herbal medicine for the treatment of bronchial asthma, we studied the effects of eight phenolic compounds, which have been identified as the major human metabolites of Saiboku-To, and three triterpenoids contained in Saiboku-To on the release of leukotriene (LT) from human polymorphonuclear leukocytes (PMLs) stimulated with Ca(2+)-ionophore A23,187. All phenolic compounds exhibited dose-dependent suppression on release of both LTB4 and LTC4, while triterpenoids did not show any effects, except for glycyrrhetinic acid, which selectively inhibited LTC4 release. The five phenolic compounds, magnolol, dihydroxydihydromagnolol, baicalein, medicarpine and davidigenin, were found to exert a marked inhibition on LTB4- and LTC4-release with IC50 values of 0.7-15.3 microM. The results suggest that the phenolic compounds contribute to the anti-allergic effects of Saiboku-To through suppression of LT-release from PMLs. PMID- 10705749 TI - Interaction of flavones from the roots of Scutellaria baicalensis with the benzodiazepine site. AB - A radioreceptor binding assay was used to guide the isolation of four chemical constituents in the organic solvent extracts of Scutellaria baicalensis Georgi capable of binding to the benzodiazepine site (BZD-S) of the GABAA receptor: wogonin (Ki = 2.03 +/- 0.24 microM), baicalin (Ki = 77.10 +/- 4.79 microM), baicalein (Ki = 5.69 +/- 0.95 microM), and scutellarein (Ki = 12.00 +/- 1.27 microM). All four compounds contain the flavonoid phenylbenzopyrone nucleus. Based on BZD-S binding, the order of affinity among these four compounds was wogonin > baicalein > scutellarein > baicalin. PMID- 10705750 TI - Antiplasmodial activity of the alkaloids of Peschiera fuchsiaefolia. AB - The tertiary and quaternary alkaloids isolated from the stem bark, root bark and seeds of Peschiera fuchsiaefolia are reported. The tertiary alkaloid crude extract from the stem bark was tested in vitro against Plasmodium falciparum on the basis of the antimalarial use of the plant. It showed good activity against both the D6 strain (IC50 = 495 ng/ml) and chloroquine-resistant W2 strain (IC50 = 817 ng/ml) and voacamine was the most active of the tested alkaloids (IC50 = 238 ng/ml for D6 and 290 ng/ml for W2). The tertiary alkaloid crude extract from the root bark of the same plant is more active than voacamine (IC50 = 179 ng/ml for D6 and 282 ng/ml for W2 strain), and is particularly rich in dimeric alkaloids (0.22% of the vegetable material). PMID- 10705751 TI - Quantitative determination of secoiridoid and gamma-pyrone compounds in Gentiana lutea cultured in vitro. AB - The production of secondary metabolites was studied in shoots, roots, and hairy roots of Gentiana lutea obtained in vitro. In shoots, both secoiridoid and gamma pyrone compounds were detected in amounts similar to those found in aerial parts of plants collected from nature. The most abundant secoiridoid was gentiopicrin while mangiferin was the main compound among the gamma-pyrones. The adventitious roots obtained in vitro showed a poor biosynthetic capacity. Upon infection with Agrobacterium rhizogenes, nine hairy root clones were established which differed in the amount of secondary metabolites. PMID- 10705752 TI - [Invasive or noninvasive ventilation? No either-or! On the value of noninvasive ventilation]. PMID- 10705753 TI - [Noninvasive ventilation in acute respiratory insufficiency]. AB - In many ICU's in these days non-invasive mask ventilation is a technique in addition to the traditional invasive ventilation via endotracheal tube if the patient is able to cooperate. If contraindication (mucus retention, swallowing disorders, acute neurologic disorders, circulatory instability) and definition of interruption criteria are regarded, the method is safe. Besides its use for exacerbation of chronic obstructive pulmonary disease, especially the cardiogenic, pulmonary oedema is an indication which is supported by several studies. Other conditions such as pneumonia and ARDS have only been observed in relatively small studies. According to pathophysiologic pathways the use of non invasive techniques seems limited to not-so-advanced stages of the disease in these cases. Considering the growing importance of this technique for the weaning procedure after long-term mechanical ventilation should be established at every ICU. PMID- 10705754 TI - [Noninvasive ventilation in ventilator weaning]. PMID- 10705755 TI - [Solitary fibrous pleural tumors--rare tumors with unpredictable clinical behavior]. AB - Solitary fibrous tumors of the pleura are rare tumors with unpredictable clinical behaviour. We report about two patients with an incidental finding of an intrathoracic tumor. Preoperative diagnosis was uncertain. In both patients, a solid tumor of the pleura was resected en bloc in combination with a wedge resection of the lung following anterolateral thoracotomy. The postoperative course was eventful. Six months after primary complete resection there were no signs of tumor recurrence. PMID- 10705756 TI - [Primary leiomyosarcoma of the pulmonary artery--a case report]. AB - A case of a primary pulmonary leiomyosarcoma originating in the right pulmonary artery in a 61-year-old woman is reported. Patient's complaints were non-specific and after non-invasive diagnostics a chronic thromboembolic event was suspected to have occurred a long time ago. During a heart catheter investigation prior to planned surgical embolectomy a small specimen was taken from the pulmonary artery. Histological examination revealed malignant the tumour and by immunohistological staining its smooth muscle differentiation was confirmed. Complete resection of the tumour was achieved by radical surgery, including pneumonectomy and extensive resection of pulmonary vessels with subsequent implantation of a Goretex prosthetic. There was no evidence of recurrence or metastasis of the primary leiomyosarcoma of the pulmonary artery in a 14-month follow-up period after operation. PMID- 10705757 TI - [Quartz and lung tumors--data and facts from the pathologist]. PMID- 10705758 TI - [Possibilities and limits in prevention of occupationally-induced obstructive respiratory tract diseases]. PMID- 10705759 TI - [Respiratory health risks in working with chickens]. PMID- 10705760 TI - [Flock worker's lung. Background for a new pulmonary disease picture in occupational medicine]. PMID- 10705761 TI - The newly founded Society for Psychophysiological Research. PMID- 10705762 TI - Independence of valence modulation and prepulse inhibition of startle. AB - This study sought to determine whether prepulse inhibition and valence modulation of startle are independent, both within and across individuals. Acoustic probes (105 dB) were delivered as 68 undergraduates viewed pleasant, neutral, and unpleasant pictures. Weak acoustic stimuli (8 dB above background) preceded half of the probes by 120 ms. As expected, startles were larger during unpleasant than during pleasant pictures, and smaller on prepulse than no-prepulse trials. In general, valence modulation and prepulse inhibition of startle were unrelated. That is, prepulse inhibition was consistent across affective states, valence modulation did not differ between no-prepulse and prepulse trials, and valence modulation and prepulse inhibition effects were uncorrelated across individuals. Analysis of raw and percent modification scores generally led to similar conclusions. It is concluded that valence modulation and prepulse inhibition are independent startle modulatory phenomena, although this conclusion is tempered by a finding of poor internal consistency reliability for valence modulation. PMID- 10705763 TI - Mixed-effects models in psychophysiology. AB - The current methodological policy in Psychophysiology stipulates that repeated measures designs be analyzed using either multivariate analysis of variance (ANOVA) or repeated-measures ANOVA with the Greenhouse-Geisser or Huynh-Feldt correction. Both techniques lead to appropriate type I error probabilities under general assumptions about the variance-covariance matrix of the data. This report introduces mixed-effects models as an alternative procedure for the analysis of repeated-measures data in Psychophysiology. Mixed-effects models have many advantages over the traditional methods: They handle missing data more effectively and are more efficient, parsimonious, and flexible. We described mixed-effects modeling and illustrated its applicability with a simple example. PMID- 10705764 TI - Evaluating group distributional characteristics: why psychophysiologists should be interested in qualitative departures from the normal distribution. AB - Real data often do not approximate the normal distribution. Under nonnormal conditions, psycho-physiologists who use parametric statistics may be testing with inadequate power and/or testing a measure of location (i.e., the mean) that does not represent the desired portion of the distribution for statistical comparison. The purpose of this paper is: first, to provide psycho-physiologists with a method to investigate group distributional characteristics; second, to evaluate the distributional characteristics of heart period and respiratory sinus arrhythmia in human adults and newborns; third, to demonstrate the increased statistical power that can accompany the selection of an alternative statistical analysis for nonnormal data. Suggestions are provided on how to analyze nonnormal data. PMID- 10705765 TI - Early attention effects in human auditory-evoked potentials. AB - A fundamental question in attention theory concerns the earliest processing stages that can be modulated by selective attention. A series of experiments is reported in which very early attention effects are found under specific conditions in the frequency-following potential (FFP), a brain stem response to low-frequency tone stimuli. In two experiments, stimuli of two different modalities were applied, and attention directed to one of the modalities. In two further experiments, only auditory stimuli were presented. In the first of these last two experiments, a dichotic paradigm with sustained attention to one ear was used, in the second a monotic paired-stimuli paradigm was used, in which the first stimulus served as reference for the second one. Only in the last experiment significant attention effects were found in the latency, but not in the amplitude of the FFP. The results show that a very early attention effect on the latency of the FFP can be demonstrated, but only under highly specific conditions. The size and preconditions of the attention effect suggest that it reflects subtle intramodal tuning mechanisms in the cochlea or in the lower brain stem. PMID- 10705766 TI - Error monitoring during reward and avoidance learning in high- and low-socialized individuals. AB - The error-related negativity (ERN) is a response-locked brain potential generated when individuals make mistakes during simple decision-making tasks. In the present study, we examined ERN under conditions of reward and punishment, among participants who scored extremely low or high on the socialization scale of the California Psychological Inventory (CPI). Participants completed a forced-choice task, and were rewarded for correct responses in half the trials, and punished for incorrect responses in the remaining trials. A significant interaction between socialization (SO) and condition revealed that low-SO participants produced smaller ERNs during the punishment task than during the reward task, whereas high-SO participants produced similar ERNs in both conditions. Reaction time and electromyogram data essentially bolster the interpretation that the ERN effects reflect differences in error salience for high-SO and low-SO participants, and are consistent with the avoidance-learning deficits seen in psychopathy. PMID- 10705767 TI - The effect of warning stimulus modality on blink startle modification in reaction time tasks. AB - The present study investigated the effects of lead stimulus modality on modification of the acoustic startle reflex during three reaction time tasks. In Experiment 1, participants (N = 48) were required to press a button at the offset of one stimulus (task relevant) and to ignore presentations of a second (task irrelevant). Two tones that differed in pitch or two lights served as signal stimuli. Blink startle was elicited during some of the stimuli and during interstimulus intervals. Skin conductance responses were larger during task relevant stimuli in both groups. Larger blink facilitation during task-relevant stimuli was found only in the group presented with auditory stimuli, whereas larger blink latency shortening during task-relevant stimuli was found in both groups. Experiment 2 (N = 32) used only a task-relevant stimulus. Blink magnitude facilitation was significant only in the group presented with tones, whereas blink latency shortening was significant in both groups. Experiment 3 (N = 80) used a go/nogo task that required participants to press a button if one element of a compound stimulus ended before the second, but not if the asynchrony was reversed. The offset asynchrony was varied between groups as a manipulation of task difficulty. Startle magnitude facilitation was larger during acoustic than during visual stimuli and larger in the easy condition. The present data indicate that startle facilitation in a reaction time task is affected by stimulus modality and by task demands. The effects of the task demands seem to be independent of lead stimulus modality. PMID- 10705768 TI - Development of multimodal attention in young infants: modification of the startle reflex by attention. AB - This study examined the effect of attention engagement to compound auditory visual stimuli on the modification of the startle blink reflex in infants. Infants at 8, 14, 20, or 26 weeks of age were presented with interesting audiovisual stimuli. After stimulus onset, at delays defined by heart rate changes known to be associated with sustained attention or attention disengagement, blink reflexes were elicited by visual or auditory stimuli. Blink amplitude to either visual or auditory stimuli was enhanced when the infants were engaged in attention to the foreground auditory-visual stimuli relative to control trials with no foreground patterns. This enhancement of the blink amplitude increased from 8 to 26 weeks of age. In contrast to selective modality enhancement for single-modality foreground stimuli, these results show that these multimodal stimuli engage both visual and auditory attention systems in this age range. PMID- 10705769 TI - Effects of fragrance on female sexual arousal and mood across the menstrual cycle. AB - The effects of fragrance on sexual response in women were investigated using subjective and physiological measures of sexual arousal and of mood. Responses were obtained from female participants in three different fragrance conditions (female fragrance, male fragrance, and a "blank" or neutral substance), as they viewed erotic and sexually neutral films, and fantasized about sexual situations. Each woman was tested twice: during the midfollicular and periovulatory phases of her menstrual cycle. Menstrual cycle phase effects were apparent; self-report data indicated greater sexual arousal and more positive mood during the periovulatory than during the follicular phase. Results demonstrated a positive effect of the male fragrance on genital arousal during erotic fantasy, but this finding was apparent only during the follicular phase testing session. This effect did not appear to be mediated by any effects of fragrance on mood. PMID- 10705770 TI - Comparison of electrodermal constant voltage and constant current recording techniques using the phase angle between alternating voltage and current. AB - If electrodermal activity is recorded with direct current, constant voltage and constant current measurements result in different dependencies of electrodermal reactions on the actual electrodermal level. The present study demonstrates empirically that such a problem does not exist when instead the phase angle changes between alternating current and voltage are obtained. Forty subjects were subjected to a 20-trial habituation series on two different occasions, in which electrodermal level variations were induced by room temperature changes. A multiplexing system was used to enable quasi-simultaneous constant current and constant voltage recording under both direct and alternating current measurement conditions. If the alternating current technique was applied and electrodermal responses were expressed as changes of phase angle between voltage and current, electrodermal recordings with constant voltage and with constant current provided equivalent results, even if electrodermal levels were considerably different. Therefore, using the phase angle method instead of the conventional direct current methods will finally resolve the problem of differential level dependency in electrodermal recording. A further advantage will be that electrode and skin polarization are prevented by the use of alternating current. PMID- 10705771 TI - Temporal stability of the emotion-modulated startle response. AB - In the present study, we examined the stability of one measure of emotion, the emotion-modulated acoustic startle response, in an undergraduate sample. Using the acoustic startle paradigm on two different occasions, we measured stability of affective modulation of the startle response during and following the presentation of pictures selected to be of positive, negative, or neutral emotional valence. The two assessments were separated by 4 weeks. Two groups of subjects were compared: one group that viewed the same pictures at each assessment and a second group that viewed different pictures at the second assessment. We found that viewing different pictures at two assessments separated by 4 weeks yielded moderate stability of the emotion modulation of startle magnitude, whereas subjects who viewed the same pictures at both assessments showed poor stability. Furthermore, this difference was due to the stability of responses to high versus low arousal pictures, not to differences in valence. PMID- 10705772 TI - Orienting response reinstatement and dishabituation: effects of substituting, adding, and deleting components of nonsignificant stimuli. AB - The prediction that orienting response (OR) reinstatement is negatively related to the measure of common features, shared by the stimulus input and representations of preceding events, and positively related to the measure of their distinctive features, was examined. A nonsignificant test stimulus (TS) was presented after nine repetitions of a standard stimulus (SS), followed by two additional repetitions of SS. TS was created by either substituting 0, 1, or 2 components of SS (Experiment 1), or by either adding or deleting 0, 1, or 2 components of SS (Experiment 2). Skin conductance changes to TS (OR reinstatement) and the subsequent SS (dishabituation) were used as dependent measures. The results of Experiment 1 supported the prediction that substituting components of neutral stimuli affects OR reinstatement, with a larger effect for between-categories than within-categories substitution. Experiment 2 demonstrated that adding and deleting components similarly affects OR reinstatement. PMID- 10705773 TI - Selective influence of the menstrual cycle on perception of stimuli with reproductive significance: an event-related potential study. AB - In this study, we examined changes in the event-related potential (ERP) to stimuli with and without reproductive significance occurring during the menstrual cycle. Eleven spontaneously cycling women were tested during three menstrual phases (menses, ovulatory phase, luteal phase) differing in plasma concentrations of gonadal hormones. ERPs were recorded while subjects were presented with slides showing pictures from four different stimulus categories (sexual stimuli, babies, people occupied with body care, ordinary people). Slides were presented randomly in the context of two tasks, requiring either affective processing (i.e., to judge the emotional content of a slide as positive, neutral, or negative) or structural processing (i.e., to estimate the number of parallel thin lines inserted in each picture). Menstrual phase primarily affected a late positive component (LPC) peaking 550-600 ms poststimulus. The effects were as follows: (i) During the ovulatory phase, amplitude of the LPC to sexual stimuli was larger than that evoked by the other stimulus categories. (ii) This relationship was not apparent during the other menstrual phases or (iii) during the ovulatory phase when the task required structural processing. The ovulatory increase in LPC positivity to sexual stimuli suggests a greater valence of these stimuli during a phase of increased sexual desire. The data indicate a specific effect of the menstrual cycle on the processing of sexual stimuli that increases with deeper emotional processing. PMID- 10705774 TI - EOG correction: which regression should we use? AB - Electrooculogram (EOG) correction is used to remove eye-movement-related contamination from electroencephalograms (EEG). Correction is reliant on the regression procedure, although when multiple EOG channels are used in the correction, the appropriate type of regression to use is not known. In the present study, we aimed to resolve this matter. Computer simulations were used to compare the simultaneous, multiple-stage, and single-channel regression methods of correction. EOG propagation was modeled on prior findings, under conditions of varying vertical and horizontal EOG (VEOG/HEOG) correlation. The dependent variable was the correlation between the uncontaminated and the corrected EEG. The simultaneous regression procedure gave the best correction, with its advantage increasing as a function of VEOG/HEOG correlation. It is recommended that the simultaneous regression procedure be used for EOG correction of the EEG. PMID- 10705775 TI - [Guidelines and responsibility of the physician]. PMID- 10705776 TI - [New indications for permanent cardiac pacing]. PMID- 10705777 TI - [Evaluation of the impact of a training program on the quality of hospital discharge record data]. PMID- 10705778 TI - [Efficacy and safety of clarithromycin in the treatment of community-acquired pneumonia]. AB - Community-acquired pneumonia (CAP) is a serious disease frequently treated empirically, which required the selection of an antibiotic that covers all common pathogens and achieves good pulmonary concentrations. The availability of intravenous (i.v.) formulations may also be helpful, permitting i.v./p.o. sequential therapy. From January 1992 to December 1997 we treated 290 CAP patients with clarithromycin (CL) 500 mg BID, first given i.v. in 250 or 500 ml of saline solution and then switched after 4-5 days to the same dosage given p.o. Of these 290 patients 163 were males (98 smokers) and 127 were females (41 smokers); 87 were over 65 years old and 203 had concomitant diseases (mainly cardiovascular), 172 patients were admitted after unsuccessful therapy (122 cephalosporins and 48 penicillins). Diagnosis was made based on clinical and radiological findings, therapy was initiated prior to microbiological diagnosis. Clinical and radiological improvement was achieved by 261/290 patients (90%) within 10-15 days. Mild adverse events occurred in 11 patients. This results indicate that CL is effective and safe: its antimicrobial spectrum and pharmacokinetic profile, the possibility of i.v./p.o. sequential administration, make it an ideal antibiotic for the treatment of CAP. PMID- 10705780 TI - [Treating also those who cannot recover]. PMID- 10705779 TI - [A rare case of cryptococcal meningitis unrelated to AIDS]. AB - It is presented the clinical case of a man 60 years old, heterosexual, suffering from chronic bronchopathy from old date, inveterate smoker, with previous diskotomy, herniotomy, who presents a symptomatology characterized from recurrent fever, productive cough, dyspnea, asthenia and headache for 6 month. He was admitted to hospital for fever and for a sensory slightly obnubilated. A series of investigations for typhus fever, cytomegalovirus, all with negative results were performed. He resulted negative also to the test to PPD as well as to markers of B and C hepatitis and the test for HIV. The study of the principal cancer markers also gave negative results, while the blood smears displayed leukopenia with monocytosis. The magnetic nuclear resonance of the brain showed the presence of multiple lesions of the brain and along the meninges: the examination of the liquor underlines the presence of the Cryptococcus neoformans, making to set the diagnostic of cryptococcal meningitis. The immunological study showed low values of CD4 in presence of normal values of CD8 and of a normal natural killer function. The exitus happened at 64th day. The interest of the case consists in the fact that in the medical Italian literature, unlike the international one, are not described cases of cryptococcal meningitis in patients not infected by HIV. PMID- 10705781 TI - [Current aspects of diagnosis of diabetes]. PMID- 10705782 TI - [Thyroid diseases: molecular diagnosis and therapeutic perspectives]. AB - The diagnostic algorithm of thyroid diseases, the most frequent dysendocrine condition, can be today integrated by the newly developed molecular methodologies. From the early diagnostic approaches, centered on the assessment of thyroid function, either by in vivo radioisotopic techniques, or by in vitro radioimmunological measurement of hormone plasma concentrations, it is nowadays possible to precisely define the molecular events triggered by the iodothyronine signal at the level of target tissues. In this brief review will be discussed the recent progresses on cloning and characterization of several genes involved in the regulation of thyroid differentiation, ability to trap iodine, synthesis and secretion of iodothyronines, regulation of thyroid function by TSH, transduction of the hormonal signal to subcellular structures involved in the translation of the hormone message in specific biological, effects such as those on metabolic homeostasis, cell proliferation and differentiation. It will be also discussed the most recent advancements on the genetics of thyroid diseases which have allowed to characterize the molecular basis of several thyropathies such as congenital hypothyroidism, thyroid hormone resistance syndrome, hyperthyroidism or hypothyroidism caused by TSH-receptor alterations, molecular abnormalities of oncogenes or tumor suppressor genes which are associated with benign or malignant thyroid cell transformation. The most recent developments of the diagnostic procedures of thyroid diseases, also in their pre-clinical stage, will be also reviewed together with a brief highlight on the most recent treatment options, centered on prophylactic therapeutic intervention or on the development of gene therapy strategies which will be possibly applied in a near future. PMID- 10705784 TI - [Patient-centered interviews in general practice]. AB - Many studies have confirmed that the interview approach adopted by the physician influences the quality of the doctor-patient relationship as well as the accuracy and validity of information which is elicited during the medical interview. A correct interview approach is also the basis for diagnosis and treatment. This paper summarizes the main characteristics of the patient-centred interview approach which has to integrate the doctor centred approach in order to render the medical interview efficient in terms of data collection, doctor-patient relationship and time. The first part of the interview with the patient has always to be patient-centred. The patient is facilitated to report all the information regarding his symptoms and their psychosocial context and to express his ideas and expectations, after which the interview style becomes more directive and doctor-led. In this phase the doctor remaining however attentive to patient's cues. Some data are presented which describe the prevailing interview approach of general practitioners without any formal training in patient-centred interview techniques. The analysis of doctors' verbal behaviour during the consultation confirmed the necessity of educational interventions focalized on the improvement of doctors' interview skills. PMID- 10705783 TI - [Meniere's disease: diagnosis and new treatment perspectives]. AB - Meniere's disease is characterized by attacks of vertigo and by sensorineural hearing loss, tinnitus, and ear fullness; these cochlear symptoms are unilateral in the majority of cases. Medical treatment tends to control vertigo and to cure the possible causes of endolymphatic hydrops (autoimmune disease, syphilis, metabolic and endocrine derangements, etc). In case of failure of medical treatment, surgery is considered. Endolymphatic mastoid shunt is indicated when hearing is normal or fluctuating. In all other cases, vestibular neurectomy or chemical vestibular labyrinthectomy, by means of intratympanic low concentration gentamicin (20 mg/mL), are indicated. Total or substantial regression of vertigo, at 2 years follow up, occurs in 65% of cases after endolymphatic-mastoid shunt, and in > 90% after vestibular neurectomy or chemical vestibular labyrinthectomy. PMID- 10705785 TI - [When bacteria communicate with each other: the significance of intercellular signals in acute Pseudomonas aeruginosa infections]. AB - In the opportunistic pathogen Pseudomonas aeruginosa a cell-to-cell signaling circuitry regulates the production of several major extracellular virulence factors. This interbacterial communication system allows P. aeruginosa to produce these virulence factors in a coordinated manner that overwhelms the host defense mechanisms. This article gives an overview of our present knowledge of the complex cell-to-cell signaling circuitry of P. aeruginosa and tries to outline its importance in the pathogenesis of acute P. aeruginosa infections. New therapeutic strategies, based on the blockage of this circuitry, are presented. PMID- 10705786 TI - [Immunologic effectors and Mycobacterium tuberculosis]. PMID- 10705787 TI - [Infective diarrhea]. PMID- 10705788 TI - [Lyme borreliosis: update]. PMID- 10705789 TI - [HIV: epidemiology and treatment --2000]. PMID- 10705790 TI - [Infective Chlamydia pneumoniae and coronary disease: where is the truth?]. AB - Chlamydia pneumoniae has been associated with atherosclerosis and coronary heart disease for several years. This association is based upon 4 research areas: 1. seroepidemiological studies, 2. detection of the pathogen in diseased arteries, 3. experimental studies in vitro and in animal models, and 4. human intervention trials with antibiotics. We review and discuss recent data from these four areas. Although the infectious hypothesis of atherosclerosis is not new, the bulk of evidence supporting a role for C. pneumoniae is important. However, a definite pathogenic role for initiating or accelerating the athreosclerotic process remains unproven. Several large clinical trials with antibiotic interventions in patients with coronary heart disease are ongoing. Treatment of C. pneumoniae infection and the perspectives for a vaccine remain, however, problematic. PMID- 10705792 TI - [Blastomycosis 30 years after living in Africa]. PMID- 10705791 TI - [Chronic meningococcemia]. PMID- 10705793 TI - [Alterations of liver tests and unexplained fever after returning from Haiti]. PMID- 10705794 TI - [Is there a role for infectious disease specialists in private practice?]. AB - For the last 20 years infectious diseases have gained increasing importance for hospital medicine. As a specialty, infectious diseases have been recognized only recently by the Swiss medical association. However, the precise role of infectious disease specialist operating in private practice remain to be defined. The medical community faces many challenges for which infectious disease specialist must provide answers. Knowledge in microbiology has progressed enormously and many very sophisticated and, partly, expensive diagnostic techniques are widely available. New treatment options are introduced while numerous microbial species demonstrate increasing resistance to antimicrobial agents. The intervention of infectious disease specialist could thus contribute to optimize treatment and limit the use of economic resources. Infectious disease specialist in private practice are also facing new activities such as parenteral outpatient treatment for severe infections and HIV infection, which clearly require a specialized professional approach. Infectious disease specialist in private practice will need great care to find a responsible equilibrium between clinical consultation and telephone consultation. PMID- 10705795 TI - [Biologic weapons: are they anecdotes of the past or a future reality?]. PMID- 10705796 TI - [Rheumatology expertise]. AB - Requests for expert appraisals in cases of osteoarthritic diseases increases continually, especially those addressed by the "Office de I'Assurance Invalidite (OAI)" or private insurances (PRIV). In this series of 413 cases, the mean age of the applicants was significantly lower than that of 10 years ago. There were more men than women, foreigners than Swiss natives and couples than people living alone but not significantly. 85% of complaints were for vertebral pain; fibromyalgia and depression were noted in 8% and 27% of cases respectively. Only 42% of those demanding time off work had heavy work; 66% did not continue their schooling beyond the minimum requirement. Less than 1/3 of work interruption for OAI and 1/6 for PRIV were confirmed; more than 1/2 of the patients for PRIV could not justify a work disability higher than 20%. 19% of our appraisals were disputed; 3% succeeded and 3% are still in progress. Chronic rheumatological complaints do not necessarily result in insurance compensations. Practitioners can contribute to chronification of rheumatological complaints by perpetuating work interruption for non medical reasons. PMID- 10705798 TI - [Vaud-Geneva: new ambitions for university links]. PMID- 10705797 TI - [Oral zolmitriptan (Zomig) in the treatment of migraine crisis]. PMID- 10705799 TI - [Diagnosis and treatment of pulmonary tuberculosis in asylum seekers and refugees in the Bern canton 1993-1997]. AB - As part of a preventive program asylum seekers and refugees are screened for tuberculosis. Necessary treatments have to be administered in the accepting canton. Aim of this cohort study was to assess the way of diagnosis and the outcome of treatment in 64 cases of pulmonary tuberculosis and positive culture in the Canton of Berne between 1993 and 1997. Results existed for 62 of them (96.9%). 34 cases of tuberculosis were discovered by screening and 23 more than 6 months afterwards. 87.1% of all cases were treated for more than 6 months. Among patients with unfavourable outcome four disappeared, three were transferred out. Efficacy of screening for tuberculosis depends also on follow-up during treatment which took place in the Canton of Berne. A strict organisation like the one applied in Berne or directly controlled treatment are necessary to warrant curative treatment. PMID- 10705800 TI - [Practical considerations in management of adult asthmatic patients]. AB - Asthma is a common, chronic inflammatory disease of the airways associated with pronounced health and economic consequences. Identification and control of asthma triggers and an adequate pharmacological therapy are main components. But for a successful asthma care consistent education is another critical aspect for management that has to be considered. In this article several practical considerations for managing asthma in adults, with emphasis on the importance of patient education and partnership in care are described. PMID- 10705801 TI - [Milieu therapy for patients with dementia. Appropriate, regular stimulation by pleasant experiences]. AB - The target group includes caregivers of demented persons at home and in institutions. The aim is a synopsis of different ways to cope with behavioural disturbances by milieu-therapy for demented persons. Appeals are made to their remaining resources, thus getting them more joy and less frustration by less excessive demands. Less boring activities avoid to feel under-challenged. More activity during day-time provides better sleep at night.--Consequently there are less behavioural disturbances with less stress for caregivers thus enabling them to keep the patients longer at home, leading to lower health costs. Behavioural disturbances of demented persons should always be treated by milieu-therapy achieving a response rate of up to 60%. With application of adjuvant medication, e.g. geriatric neuroleptics, a rate of 70% is within reach. Milieu-therapy is optimal for the prevention of behavioural disturbances. The better the adaptation of milieu-therapy to the individual patient's deficits and lifestyle and to the lifestyle of his caregiver, the better the effect. PMID- 10705802 TI - [Primary retroperitoneal seminoma--a rare germ cell neoplasm]. AB - A 44-year old male presented himself with left-sided abdominal pain. On clinical examination a painless, enlarged supraclavicular lymph node on the left side and small atrophic testes were detected. Ultrasonography revealed a huge retroperitoneal mass. By biopsy of the retroperitoneal tumor the diagnosis of a seminoma was made, but neither in the orchidectomy specimen of the right side nor in the testicular biopsy of the left side a primary tumor or a scar could be identified. Thus, the diagnosis of a extragonadal primary retroperitoneal seminoma was made. The patient responded well to the combined chemotherapy consisting of cisplatin, bleomycin and etoposide. For distinction from an occult and/or burned-out seminoma testicular biopsy or surgical exploration of the testes with histology are mandatory. PMID- 10705803 TI - [Giant cell arteritis]. PMID- 10705804 TI - [1899: the first mathematical description of the pressure-volume diagram by Otto Frank (1865-1944)]. AB - Not until 60 years after 1899 Otto Frank's mathematical formulation of the volume pressure diagram and his concept of the mechanism of the cardiac work of the left ventricle cardio-physiologists began to rediscover Frank's work systematically. Frank's scientific development proves his constant commitment to and deep interest in understanding and analysing physiological problems mathematically. Consequently Frank worked on the quality and theory of physiological instruments and the problems of measurement, to calculate the influence in experimental research of the cardio-vascular system. Besides Frank's publications on different types of manometers his theory of "Windkessel" function as a model of the mechanics of the left ventricle and the different energies of the cardiac work are of importance even today, although we know only few details of his life as a pupil of Carl Ludwig and Carl von Voit and as a professor of physiology in Giessen and Munich. PMID- 10705805 TI - [Preservation of the arousal sum. The physiologists Ernst W. Brucke, Sigmund Exner and Ernst Fleischl von Marxow as teachers of Sigmund Freud]. PMID- 10705807 TI - Tracking the Pain. Jue(a,b,c,) and the formation of a theory of circulating qi(d) through the channels. PMID- 10705806 TI - [Knowledge of the "Grafenberg zone" and female ejaculation in ancient Indian sexual science. A medical history contribution]. AB - Ancient Indian texts in sexology (kamasastra) from the 11th century onwards prove that their authors knew about the area later termed the "Grafenberg zone" in Europe, as well as about the female ejaculation connected with the stimulation of this area. The Grafenberg zone is a sexually arousable zone in the front part of the vagina, stimulation of which can lead to the discharge of liquid from the urethra, a phenomenon which is described as female ejaculation. The german gynaecologist Ernst Grafenberg, who worked in America, described this zone, situated beneath the clitoris, for the first time (at least in this century) in Western medicine in an article published in 1950. (There are, however, evidences, that the 17th-century anatomist Regnier de Graaf had knowledge about the mentioned erogenous zone as well as female ejaculation.) Since the 1980s the so called Grafenberg zone, popularly termed "G-spot", and female ejaculation have been controversially discussed medically as well as in popular science, first in the United States, then in Europe; both phenomena have meanwhile been accepted as facts in medical manuals and reference books (e.g. the "Pschyrembel"). Whereas the oldest and most well-known sexological-erotological work of Ancient India, the Kamasutra, dating probably from the third century A.D., apparently did not know the Grafenberg zone and female ejaculation, texts such as the Pancasayaka (11th century), Jayamangala (Yasodhara's commentary on the Kamasutra from the 13th century), the Ratirahasya (13th century), as well as the late kamasastra works Smaradipika and Anangaranga (16th century?) demonstrably describe both, the Grafenberg zone and female ejaculation, in great detail. The female ejaculation is described already in the 7th century in a non-kamasastra-text, in a work of the poet Amaru called the Amarusataka. PMID- 10705808 TI - [Early Egyptian forerunners of the Paranatellonta?]. AB - The term "paranatellonta" is well-known in greek astrological literature. It designates stars either rising together with the sun or being in other conspicuous positions to it. Tentatively, a forerunner of this conception is identified in an egyptian depiction attested several times from the 13th century BC onwards. There, "gods" are depicted who are defined by their positions in regard to the sun-god. It seems possible to connect their positions with the typical meanings of the word paranatellonta. Some reflections on the contribution of Egypt to hellenistic astrology are added, including some references to the largely unpublished corpus of demotic egyptian astrological texts. PMID- 10705809 TI - ["2 unpublished fragments by Hildegarde of Bingen copied by Gerhard von Hohenkirchen (1448)"]. PMID- 10705810 TI - [Thoughts on the illness of Hermann von Reichenau (1019-1054)]. AB - Hermann from Reichenau--Hermannus contractus--apparently suffered from a disease which led to considerable physical handicap leaving his outstanding intellectual talents undamaged. Various statements about his condition--an epileptic, suffering from spasticity, afflicted by poliomyelits--have never been reconsidered. Using the biography written by this disciple Berthold, the most important contemporary source about Hermanns' life, an approach to a correct diagnosis from a neurologists point of view was the aim of this study. By unbiased analysis of the symptoms described by Berthold a neurologic syndrome is worked out: it comprised a flaccid tetraparesis involving the bulbar area. The sensory as well as the autonomic nervous system were apparently not involved. Intellectual functions were unaffected. Considering this syndrome and other details of Hermanns' life as well as the beginning and course of his illness, a traumatic birth injury, an early childhood disease and a central nervous as well as an infectious disease are ruled out. Muscle disease is considered possible, but motor neuron disease--either amyotrophic lateral sclerosis or spinal muscular atrophy--seems to be the most convincing diagnosis. PMID- 10705811 TI - Neuroendocrine neoplasia. Current concepts. PMID- 10705812 TI - Pathologists' roles in clinical utilization management. A financing model for managed care. AB - In ancillary or laboratory utilization management, the roles of pathologists have not been explored fully in managed care systems. Two possible reasons may account for this: pathologists' potential contributions have not been defined clearly, and effective measurement of and reasonable compensation for the pathologist's contribution remains vague. The responsibilities of pathologists in clinical practice may include clinical pathology and laboratory services (which have long been well-defined and are compensated according to a resource-based relative value system-based coding system), laboratory administration, clinical utilization management, and clinical research. Although laboratory administration services have been compensated with mechanisms such as percentage of total service revenue or fixed salary, the involvement of pathologists seems less today than in the past, owing to increased clinical workload and time constraints in an expanding managed care environment, especially in community hospital settings. The lack of financial incentives or appropriate compensation mechanisms for the services likely accounts for the current situation. Furthermore, the importance of pathologist-driven utilization management in laboratory services lacks recognition among hospital administrators, managed care executives, and pathologists themselves, despite its potential benefits for reducing cost and enhancing quality of care. We propose a financial compensation model for such services and summarize its advantages. PMID- 10705813 TI - Thymic neuroendocrine carcinomas with combined features ranging from well differentiated (carcinoid) to small cell carcinoma. A clinicopathologic and immunohistochemical study of 11 cases. AB - We reviewed 11 cases of primary thymic neuroendocrine carcinomas with combined features ranging from well-differentiated to poorly differentiated neuroendocrine carcinoma. For 3 asymptomatic patients, tumors were discovered during routine examination. Presentation in the other patients was as follows: Cushing syndrome, 2 patients; chest pain, 3 patients; superior vena cava syndrome, 1 patient; and hypercalcemia and hypophosphatemia, 1 patient. No clinical data were available for the 11th patient. All tumors were located in the anterior mediastinum and treated by surgical excision. The lesions were large and well-circumscribed with areas of hemorrhage and necrosis. They were characterized by areas showing a proliferation of monotonous, round tumor cells adopting a prominent organoid pattern admixed with areas showing sheets of atypical cells with hyperchromatic nuclei, frequent mitoses, and extensive areas of hemorrhage and necrosis. Immunohistochemical studies performed in 6 cases showed strong CAM 5.2 low molecular-weight cytokeratin positivity in all cases, chromogranin and synaptophysin positivity in 4, Leu-7 in 3, and focal positivity for p53 in 2. Follow-up information for 9 cases showed that all patients died of their tumors between 1 and 4 years after diagnosis. The present cases highlight the heterogeneity of neuroendocrine neoplasms and reinforce the notion that these tumors form part of a continuous spectrum of differentiation. PMID- 10705814 TI - Mapping metastases in sentinel lymph nodes of breast cancer. AB - Localization of metastases within the sentinel lymph nodes (SLNs) of breast cancer has not been studied. Forty SLNs from 36 patients with operable primary breast cancers were identified by means of lymphatic mapping with patent blue dye. The junction between the patent blue-stained lymphatic vessel draining the tumor and the SLN was labeled with alcian blue. Metastases within the serially sectioned SLNs were assigned to the alcian blue-labeled side, to the opposite side of the virtually halved nodes, or both. Eight SLNs were negative for metastasis. Eleven SLNs had metastases only in the blue half. Only 4 cases had larger metastases in the nonblue half. Metastases are more likely to be located in the vicinity of the inflow junction of the identifiable lymphatic draining the tumor and the SLN. This should be considered when SLNs are examined, especially when they are halved for different studies. PMID- 10705815 TI - Expression of P-glycoprotein in hepatocellular carcinoma. A determinant of chemotherapy response. AB - To characterize the P-glycoprotein (Pgp) expression in human hepatocellular carcinoma (HCC), we studied 101 cases of HCC treated with surgical resection without prior treatment. Pgp expression was detected immunohistochemically using 2 monoclonal antibodies (C494, C219) and correlated with pathologic features, survival, and p53 expression. Chemotherapy response was analyzed in a separate group of patients with inoperable HCC treated with systemic chemotherapy. Positive immunostaining was seen in 92% and 80% of the tumors with C494 and C219, respectively; bile canalicular type staining was seen in all positive tumors. Pgp expression was less extensive in the tumors than in the corresponding nontumorous liver tissue. Tumor Pgp expression with either antibody had no association with cellular differentiation, aggressive pathologic features, survival, or p53 overexpression. In patients with inoperable HCC, the chemotherapy response was significantly inversely related to Pgp expression with C494 and C219. Pgp was expressed in human HCC but was patchy and less extensive than in the nontumorous tissue. Response to systemic chemotherapy was inversely related to the level of Pgp expression in patients with inoperable tumors. Pgp expression in tumors not treated with chemotherapy was not associated with a more aggressive tumor phenotype or p53 overexpression and did not influence survival. PMID- 10705816 TI - Estrogen and progesterone receptors in colon tumors. AB - Existing data suggest that there is a hormonal basis to the cause of colon cancer. In the present study, we evaluated the presence of estrogen receptors (ERs) and progesterone receptors (PRs) in colonic tumors from 156 women diagnosed with colon cancer in Utah from September 1991 through September 1994. Immunohistochemical staining with antibodies to ERs and PRs was performed on histologic sections prepared from paraffin blocks. None of the tumors were considered ER-positive; 1 tumor was PR-positive. Use of hormone replacement therapy was not associated with PR-positive tumors. These data do not support previous reports that suggest that colon tumors frequently have receptors for estrogen, progesterone, or both. PMID- 10705817 TI - Quantitative analysis of the decay of immunoreactivity in stored prostate needle biopsy sections. AB - Application of immunohistochemistry to assess the presence of prognostic tissue markers is used widely. The quantitation of these markers may be hampered by a time-related loss of antigenicity in formalin-fixed paraffin-embedded tissue stored on glass slides. Potential loss of immunohistochemical staining intensity was studied on prostatic needle biopsy sections stored for a maximum of 4 years with antibodies against p27kip1, CD-44s, MIB-1, and androgen receptor (AR). In benign tissue, the positive/total ratio for p27kip1 was determined, while CD-44s staining intensity was assessed semiquantitatively. For MIB-1 and AR, nuclear staining intensity was assessed using computed image analysis. An exponential and significant decay of immunoreactivity was seen for p27kip1, CD-44s, MIB-1, and AR, with half-lives of 587 days, 214 days, and 290 days for p27kip1, MIB-1, and AR, respectively. Immunohistochemical assessment of prognostic tissue markers on stored slides must be considered with care in research and clinical settings. PMID- 10705818 TI - Paraffin-section detection of CD10 in 505 nonhematopoietic neoplasms. Frequent expression in renal cell carcinoma and endometrial stromal sarcoma. AB - We tested 505 cases of nonhematopoietic neoplasms by immunohistochemistry using a newly characterized monoclonal antibody (clone 56C6) against the CD10 antigen. CD10 was expressed widely in neoplasms of the genitourinary tract, including 41 (89%) of 46 cases of renal cell carcinoma, 13 (54%) of 24 cases of transitional cell carcinoma, and 11 (61%) of 18 cases of prostatic adenocarcinoma. In addition, 5 (100%) of 5 endometrial stromal sarcomas, 3 (60%) of 5 rhabdomyosarcomas, 7 (50%) of 14 pancreatic adenocarcinomas, 5 (45%) of 11 cases of schwannoma, and 12 (40%) of 30 cases of malignant melanoma also were positive for CD10. Similar to normal tissue, CD10 positivity was restricted to the apical surface of malignant glandular cells of well-differentiated colonic, pancreatic, and prostatic adenocarcinoma, whereas in poorly differentiated adenocarcinoma and other tumors, such as melanoma, transitional cell carcinoma, renal cell carcinoma, and endometrial stromal sarcoma, the CD10 positivity showed diffuse cytoplasmic or membranous/Golgi patterns. The monoclonal antibody clone 56C6 is a reliable marker for CD10 in paraffin immunohistochemistry after heat-induced epitope retrieval. CD10 expression in renal cell carcinoma and endometrial stromal sarcoma may be a useful marker in the differential diagnoses of these tumors because both tumors otherwise lack specific markers. PMID- 10705819 TI - Coordinate expression of cytokeratins 7 and 20 in prostate adenocarcinoma and bladder urothelial carcinoma. AB - We studied the expression of cytokeratin (CK)-7 and CK-20 in prostate adenocarcinoma and urothelial carcinoma and evaluated their usefulness for distinguishing high-grade forms of these tumors. We examined prostate adenocarcinoma in 59 radical prostatectomy specimens and in 10 autopsy specimens showing metastatic disease, and urothelial carcinoma of the bladder in 28 cystectomy specimens. Immunohistochemical staining for CK-7, CK-20, and prostate specific antigen (PSA) was performed on paraffin sections. For prostate adenocarcinoma, 5 cases had only CK-7 positivity, 5 had only CK-20 focal positivity, 1 stained for both markers, and 48 were negative for both. PSA was positive in all but 1 poorly differentiated prostatic carcinoma. In the autopsy cases, PSA was expressed in the prostate and the metastatic tumors in most cases; few cases were focally positive for CK-7 or CK-20, but none was positive for both markers. For the urothelial tumors, CK-7 was the sole positive marker in 6 cases, and CK-20 in 1 case; 17 cases were positive for both, and 4 were negative for both. All urothelial carcinomas were PSA negative. Although PSA is useful for differentiating prostatic from urothelial carcinoma, CK-7 and CK-20 are helpful when both are positive, supporting the diagnosis of urothelial carcinoma. However, if only 1 marker is positive or both are negative, these markers have limited usefulness for distinguishing these carcinomas. PMID- 10705820 TI - Detection of hepatitis C virus RNA sequences in B-cell non-Hodgkin lymphoma. AB - Serologic testing shows that hepatitis C virus (HCV) may have a role in the pathogenesis of B-cell non-Hodgkin lymphomas (B-cell NHLs). We tried to demonstrate HCV RNA sequences in paraffin-embedded tissue from B-cell NHLs by reverse-transcription double polymerase chain reaction (RT-PCR) and Southern blotting. We studied 31 consecutive cases of B-cell NHLs; lymph nodes from 32 patients with diseases other than B-cell NHL were negative controls. Positive strand HCV RNA was tested with primers for the 5' untranslated region. Replicative negative strand HCV RNA was tested with strand-specific RT-PCR for the 5' untranslated region. Immunohistochemical staining for HCV was done using an antibody to HCV core protein. Positive-strand HCV RNA was detected in 8 patients with B-cell NHL; negative-strand HCV RNA was detected in 6 of these cases, indicating viral replication. All control cases were negative for HCV RNA. Immunohistochemistry showed no staining of lymphoma cells for HCV core proteins in any case. HCV and B-cell NHLs may be associated. RT-PCR on paraffin-embedded lymphoma tissue is an alternative method of testing for HCV. The value of immunohistochemistry could not be ascertained. The exact role of HCV in the pathogenesis of B-cell NHL needs to be studied further. PMID- 10705821 TI - Lack of expression of surface immunoglobulin light chains in B-cell non-Hodgkin lymphomas. AB - We describe 10 cases of B-cell non-Hodgkin lymphoma (NHL) that did not express immunoglobulin kappa or lambda light chains by dual-color flow cytometry. Cases were identified from 298 consecutive cases of B-cell NHL and included follicular center cell lymphoma, diffuse large B-cell lymphoma, small noncleaved cell lymphoma, and small lymphocytic lymphoma. One case did not express any immunoglobulin heavy chain (IgH) as well; however, isolated expression of IgG heavy chain was seen in another case. Immunoglobulin heavy chains were not part of the lymphoma panel in other cases. All 3 cases in which gene rearrangement studies were performed showed rearrangement of IgH genes, including the case that did not express surface IgH chains. Immunoglobulin kappa light chain genes were rearranged in 2 of 3 cases and were in germline configuration in the third. All 147 cases of benign lymph nodes analyzed by flow cytometry showed polyclonal expression of immunoglobulin kappa and lambda light chains. Because of the absence of surface immunoglobulin light chains, these tumors must be distinguished from precursor B-cell acute lymphoblastic leukemia, plasma cell tumors, and rare cases of florid follicular hyperplasia that do not express surface immunoglobulins. The absence of immunoglobulin expression on malignant B cells can result from defects at any level from gene transcription to translocation of fully assembled proteins to the cell surface. PMID- 10705822 TI - High concordance of karyotype analysis and RT-PCR for CBF beta/MYH11 in unselected patients with acute myeloid leukemia. A single center study. AB - Identification of the inversion 16 in patients with acute myeloid leukemia (AML) is of great practical value since these patients have a relatively favorable prognosis, especially when treated with high-dose cytarabine. We compared the results of cytogenetic analysis and reverse transcriptase-polymerase chain reaction (RT-PCR) for core binding factor (CBF) beta/myosin heavy chain (MYH11) in 241 unselected cases of AML. In contrast with other studies, we found a high concordance between these 2 methods. Eighteen of 241 patients showed a cytogenetic anomaly of the chromosome 16. We detected the fusion transcript by RT PCR in all 18 cases and in 2 additional patients with AML without any cytogenetic anomaly of chromosome 16. One patient had a normal diploid karyotype, and the second patient showed a trisomy 22 in karyotype analysis, which often is associated with inv(16). Only 8 of 20 CBF beta/MYH11-positive patients had M4Eo morphologic features. The much higher discrepancy between cytogenetic analysis and RT-PCR in other studies, especially in AMLs other than M4Eo, possibly indicates the necessity for PCR screening regardless of the French-American British classification. PMID- 10705823 TI - Precursor B-lymphoblastic transformation of grade I follicle center lymphoma. AB - Part of the natural history of follicle center lymphoma (FCL) is transformation to a more aggressive neoplasm, almost always a diffuse large B-cell lymphoma. We describe a rare example of a precursor B-lymphoblastic transformation of grade I FCL occurring in a 45-year-old woman 12 years after initial presentation and 3 years after successful treatment for a diffuse large cell transformation. The lymphoblastic lymphoma shared the same immunoglobulin heavy chain gene rearrangement as the FCL as assessed by polymerase chain reaction amplification and direct sequencing, as well as identical kappa light chain gene rearrangements by Southern blot analysis. The immunoglobulin heavy chain variable gene sequences of both tumors showed numerous identical base substitutions compared with germline sequences and 3 additional mutations in the lymphoblastic lymphoma not present in the low-grade FCL. These results indicate origin of the lymphoblastic process from the mature follicle center B-cell clone, rather than divergent origin of the 2 tumors from a common immature B-cell precursor. PMID- 10705824 TI - Appropriateness of prostate-specific antigen testing. AB - We established criteria for appropriate use of the prostate-specific antigen (PSA) assay and used them to evaluate PSA test utilization at 1 tertiary care institution. During a 6-month period, 2,330 PSA results were reported for outpatients and 95 for inpatients. We reviewed medical records for a random sample of 338 outpatient results (14.51%) and all 95 inpatient results, of which 21% (71/338) of outpatient and 17% (16/95) of inpatient results were inappropriate according to our test utilization criteria. Among outpatients, 52% of tests were done for screening and 19% for monitoring for cancer recurrence. For inpatients, workup for cancer (53/95 [56%]) was the most frequent indication for testing and screening the second (24/95 [25%]). Of tests failing the criteria, 66 (76%) of 87 resulted from excessively frequent and age-inappropriate screening. We assessed the potential effect on clinical outcome if these tests were not performed. Of the 87 tests considered inappropriate, only 1 test result influenced clinical management for patients younger than 75 years. By instituting simple limits on age and frequency, we estimate that 74% (64/87) of the inappropriate tests could have been eliminated. PMID- 10705826 TI - Testing for antineutrophil cytoplasmic antibodies. PMID- 10705825 TI - The preanalytic phase. An important component of laboratory medicine. AB - The preanalytic phase is an important component of total laboratory quality. A wide range of variables that affect the result for a patient from whom a specimen of blood or body fluid has been collected, including the procedure for collection, handling, and processing before analysis, constitute the preanalytic phase. Physiologic variables, such as lifestyle, age, and sex, and conditions such as pregnancy and menstruation, are some of the preanalytic phase factors. Endogenous variables such as drugs or circulating antibodies might interact with a specific method to yield spurious analytic results. The preanalytic phase variables affect a wide range of laboratory disciplines. PMID- 10705827 TI - Deficiencies in management training. PMID- 10705828 TI - It's not my patient. PMID- 10705829 TI - Who should get HIV PEP? PMID- 10705830 TI - Fibromyalgia onset. PMID- 10705831 TI - African Americans hardest hit by AIDS epidemic in U.S. PMID- 10705832 TI - Congress cares for the disabled. PMID- 10705833 TI - California nurses win landmark victory. PMID- 10705834 TI - Nurse cutbacks & patient death. PMID- 10705835 TI - Cutting it close. It's the right dose and the right drug--how can it be too much? PMID- 10705836 TI - When time is muscle. PMID- 10705837 TI - Demystifying cardiac markers. PMID- 10705838 TI - A new beat on an old rhythm. PMID- 10705839 TI - Removing a PICC. PMID- 10705840 TI - Educating staff about pain management. PMID- 10705841 TI - The measure of advocacy. PMID- 10705842 TI - Nursing international. PMID- 10705843 TI - Caring for unconscious patients. PMID- 10705844 TI - Books of the year. PMID- 10705845 TI - ADA coverage. Defining who is a 'qualified individual with a disability'. PMID- 10705846 TI - Assessing for occupational hazards. PMID- 10705847 TI - Prospects for a public health perspective on psychoactive drug use. PMID- 10705848 TI - Achieving the implausible in the next decade's tobacco control objectives. PMID- 10705849 TI - New NHLBI clinical guidelines for obesity and overweight: will they promote health? AB - OBJECTIVES: The purpose of this study was to assess the justification, on the basis of mortality, of the new National Heart, Lung, and Blood Institute (NHLBI) guidelines on obesity and overweight and to discuss the health implications of declaring all adults with a body mass index of 25 through 29 "overweight." METHODS: The relationships between NHLBI body mass index categories and mortality for individuals older than 31 years were analyzed for 6253 Alameda County Study respondents aged 21 through 75 years. Time-dependent proportional hazards models were used to adjust for changes in risk factors and weight during follow-up. RESULTS: Adjusted relative risks of mortality for 4 NHLBI categories compared with the category "normal" indicated that only being underweight or moderately/extremely obese were associated with higher mortality. Specific risk varied significantly by sex. CONCLUSIONS: Our results are consistent with other studies and fail to justify lowering the overweight threshold on the basis of mortality. Current interpretations of the revised guidelines stigmatize too many people as overweight; fail to account for sex, race/ethnicity, age, and other differences; and ignore the serious health risks associated with low weight and efforts to maintain an unrealistically lean body mass. PMID- 10705850 TI - Historical and cultural roots of drinking problems among American Indians. AB - Roots of the epidemic of alcohol-related problems among many Native North Americans are sought in cultural responses to European arrival, the role of alcohol in frontier society, and colonial and postcolonial policies. Evidence from the historical record is considered within the framework of current social science. Initially, Native American's responses to alcohol were heavily influenced by the example of White frontiersmen, who drank immoderately and engaged in otherwise unacceptable behavior while drunk. Whites also deliberately pressed alcohol upon the natives because it was an immensely profitable trade good; in addition, alcohol was used as a tool of "diplomacy" in official dealings between authorities and natives. The authors argue that further research into the origins of modern indigenous people's problems with alcohol would benefit from an interdisciplinary "determinants of health" approach in which biological influences on alcohol problems are investigated in the context of the cultural, social, and economic forces that have shaped individual and group drinking patterns. PMID- 10705851 TI - HIV incidence among injection drug users in New York City, 1992-1997: evidence for a declining epidemic. AB - OBJECTIVES: We assessed recent (1992-1997) HIV incidence in the large HIV epidemic among injection drug users in New York City. METHODS: Data were compiled from 10 separate studies (N = 4979), including 6 cohort studies, 2 "repeat service user" studies, and 2 analyses of voluntary HIV testing and counseling services within drug treatment programs. RESULTS: In the 10 studies, 52 seroconversions were found in 6344 person-years at risk. The observed incidence rates among the 10 studies were all within a narrow range, from 0 per 100 person years at risk to 2.96 per 100 person-years at risk. In 9 of the 10 studies, the observed incidence rate was less than 2 per 100 person-years at risk. The weighted average incidence rate was 0.7 per 100 person-years at risk. CONCLUSIONS: The recent incidence rate in New York City is quite low for a high seroprevalence population of injection drug users. The very large HIV epidemic among injection drug users in New York City appears to have entered a "declining phase," characterized by low incidence and declining prevalence. The data suggest that very large high-seroprevalence HIV epidemics may be "reversed." PMID- 10705852 TI - The dynamics of alcohol and marijuana initiation: patterns and predictors of first use in adolescence. AB - OBJECTIVES: This study, guided by the social development model, examined the dynamic patterns and predictors of alcohol and marijuana use onset. METHODS: Survival analysis and complementary log-log regression were used to model hazard rates and etiology of initiation with time-varying covariates. The sample was derived from a longitudinal study of 808 youth interviewed annually from 10 to 16 years of age and at 18 years of age. RESULTS: Alcohol initiation rose steeply up to the age of 13 years and then increased more gradually; most participants had initiated by 13 years of age. Marijuana initiation showed a different pattern, with more participants initiating after the age of 13 years. CONCLUSIONS: This study showed that: (1) the risk of initiation spans the entire course of adolescent development; (2) young people exposed to others who use substances are at higher risk for early initiation; (3) proactive parents can help delay initiation; and (4) clear family standards and proactive family management are important in delaying alcohol and marijuana use, regardless of how closely bonded a child is to his or her mother. PMID- 10705853 TI - The relationship between external threats and smoking in central Harlem. AB - OBJECTIVES: This study assessed the relationship between external risks, such as personal and neighborhood danger, and smoking by using a new theoretical framework based on competing mortality risk models. METHODS: Regression analyses of self-reported data from residents of Central Harlem, New York, surveyed from 1992 through 1994 (n = 695, response rate = 72%) were used to assess the relationship between smoking and 2 measures of external health threats: levels of neighborhood danger and lifetime trauma. RESULTS: Support for the framework was mixed. At the 95% confidence level, exposure to lifetime trauma was positively related to current smoking status but was not related to the number of cigarettes smoked, conditional on being a smoker. Living in a "somewhat unsafe neighborhood" was also statistically significantly related to current smoking status. CONCLUSIONS: Although the framework implies that policies directed at improving the physical and social environment might improve health through their indirect effects on behaviors, little supporting evidence was found. Smoking rates may decrease if exposure to violence and neighborhood danger is reduced. This framework needs to be tested on larger and more information-rich data sets. PMID- 10705854 TI - Trends in adult cigarette smoking in California compared with the rest of the United States, 1978-1994. AB - OBJECTIVES: This study compared trends in adult cigarette smoking prevalence in California and the remainder of the United States between 1978 and 1994. METHODS: We used data from National Health Interview Surveys and Behavioral Risk Factor Surveillance System surveys to compare trends in smoking prevalence among persons 18 years and older. RESULTS: In both California and the remainder of the United States, the estimated annual rate of decline in adult smoking prevalence accelerated significantly from 1985 to 1990: to -1.22 percentage points per year (95% confidence interval [CI] = -1.51, -0.93) in California and to -0.93 percentage points per year (95% CI = -1.13, -0.73) in the remainder of the nation. The rate of decline slowed significantly from 1990 to 1994: to -0.39 percentage points per year (95% CI = -0.76, -0.03) in California and to -0.05 percentage points per year (95% CI = -0.34, 0.24) in the remainder of the United States. CONCLUSIONS: The presence of an aggressive tobacco control intervention has supported a significant decline in adult smoking prevalence in California from 1985 to 1990 and a slower but still significant decline from 1990 to 1994, a period in which there was no significant decline in the remainder of the nation. To restore nationwide progress in reducing smoking prevalence, other states should consider similar interventions. PMID- 10705855 TI - The impact of an antismoking media campaign on progression to established smoking: results of a longitudinal youth study. AB - OBJECTIVES: We examined the impact of a statewide antismoking media campaign on progression to established smoking among Massachusetts adolescents. METHODS: We conducted a 4-year longitudinal survey of 592 Massachusetts youths, aged 12 to 15 years at baseline in 1993. We examined the effect of baseline exposure to television, radio, and outdoor antismoking advertisements on progression to established smoking (defined as having smoked 100 or more cigarettes), using multiple logistic regression and controlling for age; sex; race; baseline smoking status; smoking by parents, friends, and siblings; television viewing; and exposure to antismoking messages not related to the media campaign. RESULTS: Among younger adolescents (aged 12 to 13 years at baseline), those reporting baseline exposure to television antismoking advertisements were significantly less likely to progress to established smoking (odds ratio = 0.49, 95% confidence interval = 0.26, 0.93). Exposure to television antismoking advertisements had no effect on progression to established smoking among older adolescents (aged 14 to 15 years at baseline), and there were no effects of exposure to radio or outdoor advertisements. CONCLUSIONS: These results suggest that the television component of the Massachusetts antismoking media campaign may have reduced the rate of progression to established smoking among young adolescents. PMID- 10705856 TI - Characterizing and identifying "hard-core" smokers: implications for further reducing smoking prevalence. AB - OBJECTIVES: Some smokers may never quit. Depending on how many of these "hard core" smokers exist, tobacco control efforts could reach the limits of a minimum achievable smoking prevalence. We defined the hard core as heavy smokers with weak quitting histories who expect never to quit smoking. We compared them with other smokers and analyzed whether they represent a meaningful barrier to further reducing smoking prevalence. METHODS: We used data from the 1996 California Tobacco Surveys (18616 adults; response rate = 72.9%). RESULTS: In 1996, 5.2% of California smokers 26 years and older (1.3% of the California population) were hard-core smokers. Compared with other smokers, hard-core smokers were more likely to be retired non-Hispanic White males, with 12 years or less of education and incomes below $30,000 a year, who live alone. They began smoking at younger ages and attributed fewer negative health consequences to smoking than other smokers. CONCLUSIONS: Current tobacco control efforts have a long way to go before they "hit the wall." Nonetheless, the group of hard-core smokers represents a challenge because they appear to be largely unaffected by the messages of tobacco control. PMID- 10705857 TI - Maternal smoking and adverse birth outcomes among singletons and twins. AB - OBJECTIVES: This study assessed the effects of maternal smoking on birth outcomes among singletons and twins. METHODS: An algorithm was developed to link twins with their siblings in the 1995 Perinatal Mortality Data Set. A random-effects logistic regression model was then used to estimate the association between maternal smoking and several adverse outcomes for a random sample of singletons and for all twins with available maternal smoking information. RESULTS: The algorithm successfully linked sibling pairs for 91% of the twin sample. Maternal smoking was associated with a significantly increased risk of low birthweight, very low birthweight, and gestation of less than 33 weeks for both singletons and twins and with an increased risk of gestation of less than 38 weeks, infant mortality, and placental abruption for singletons. Among smokers, negative impacts on the risk of low birthweight, very low birthweight, and extreme premature delivery were significantly higher for women carrying twins. CONCLUSIONS: Some of the negative effects of smoking on low birthweight and preterm delivery are greater for twins than for singletons. Women carrying twins should be warned that smoking increases their already high risk of serious infant health problems. PMID- 10705858 TI - Smoking prevalence in 2010: why the healthy people goal is unattainable. AB - OBJECTIVES: This study examined the changes in smoking initiation and cessation needed to realize the Healthy People 2010 national adult smoking prevalence objective (13%). METHODS: Using data from the National Health Interview Surveys, we calculated smoking prevalence over time with a dynamic population demographics model, examining the effects of changes in smoking initiation and cessation. RESULTS: The draft objective is unattainable solely through decreases in smoking initiation. It could be achieved through smoking cessation alone only if cessation rates immediately increased by a factor of more than 3.5. Assuming plausible decreases in initiation and increases in cessation, the draft objective is virtually unattainable. CONCLUSIONS: The health objectives should challenge the status quo but be achievable. Formal analysis often can assist in establishing reasonable objectives. PMID- 10705859 TI - Predictors of continued smoking over 25 years of follow-up in the normative aging study. AB - OBJECTIVES: This study tested the hypothesis that high daily cigarette consumption and addiction to smoking are risk factors for the long-term continuation of smoking. METHODS: Using longitudinal data from 986 male smokers, we entered cigarettes per day, psychological addiction, age, and education into a survival analysis as predictors of continued smoking over a 25-year period. RESULTS: Younger men and those who smoked more cigarettes per day were more likely to remain smokers in the long term. Addiction and education level were not significant predictors of continued smoking. CONCLUSIONS: Heavier smokers are more at risk than lighter smokers for long-term smoking. It is therefore very important to provide smoking cessation treatments for heavy smokers as early as possible after the initiation of smoking. PMID- 10705860 TI - Tobacco marketing and adolescent smoking: more support for a causal inference. AB - OBJECTIVES: This prospective study examined the effect of tobacco marketing on progression to established smoking. METHODS: Massachusetts adolescents (n = 529) who at baseline had smoked no more than 1 cigarette were reinterviewed by telephone in 1997. Analyses examined the effect of receptivity to tobacco marketing at baseline on progression to established smoking, controlling for significant covariates. RESULTS: Adolescents who, at baseline, owned a tobacco promotional item and named a brand whose advertisements attracted their attention were more than twice as likely to become established smokers (odds ratio = 2.70) than adolescents who did neither. CONCLUSIONS: Participation in tobacco marketing often precedes, and is likely to facilitate, progression to established smoking. Hence, restrictions on tobacco marketing and promotion could reduce addiction to tobacco. PMID- 10705861 TI - Women and smoking in Hollywood movies: a content analysis. AB - OBJECTIVES: We analyzed the portrayal of smoking in Hollywood films starring 10 popular actressess. METHODS: Five movies were randomly sampled for each actress, for a total of 96 hours of film footage that was analyzed in 1116 5-minute intervals. RESULTS: Leading female actors were as likely to smoke in movies aimed at juvenile audiences (PG/PG-13) as in R-rated movies, whereas male actors were 2.5 times more likely to smoke in R-rated movies. PG/PG-13-rated movies were less likely than R-rated movies to contain negative messages about smoking. CONCLUSIONS: Smoking is highly prevalent in Hollywood films featuring popular actressess and may influence young audiences for whom movie stars serve as role models. PMID- 10705862 TI - Tobacco and alcohol use during pregnancy and risk of oral clefts. Occupational Exposure and Congenital Malformation Working Group. AB - OBJECTIVES: This study examined the relationship between maternal tobacco and alcohol consumption during the first trimester of pregnancy and oral clefts. METHODS: Data were derived from a European multicenter case-control study including 161 infants with oral clefts and 1134 control infants. RESULTS: Multivariate analyses showed an increased risk of cleft lip with or without cleft palate associated with smoking (odds ratio [OR] = 1.79, 95% confidence interval [CI] = 1.07, 3.04) and an increased risk of cleft palate associated with alcohol consumption (OR = 2.28, 95% CI = 1.02, 5.09). The former risk increased with the number of cigarettes smoked. CONCLUSIONS: This study provides further evidence of the possible role of prevalent environmental exposures such as tobacco and alcohol in the etiology of oral clefts. PMID- 10705863 TI - Maternal cigarette smoking during pregnancy and infant ponderal index at birth in the Swedish Medical Birth Register, 1991-1992. AB - OBJECTIVES: This study examined the effect of maternal smoking during pregnancy on infant body proportion. METHODS: The ponderal index, defined as birthweight divided by crown-heel length cubed, was examined in 207,607 infants from the Swedish Medical Birth Register for 1991 and 1992. RESULTS: Infant ponderal index was used as the outcome variable in an ordinary least squares continuous regression, which included early pregnancy smoking status, gestational age, and birthweight among the predictors. Ponderal index increased by 0.030 (+/- 0.0014) among infants of moderate smokers and by 0.040 (+/- 0.0017) among infants of heavy smokers, showing a dose response. CONCLUSIONS: Smoking differentially alters the trajectory of weight vs length growth in the fetus. PMID- 10705864 TI - Minimal smoking cessation interventions in prenatal, family planning, and well child public health clinics. AB - OBJECTIVES: This study assessed the prevalence and effectiveness of smoking cessation interventions for women of childbearing age in public health clinics. METHODS: Smokers in prenatal, family planning, and well-child services in 10 public health clinics (n = 1021) were interviewed 5 to 8 weeks after a medical visit to assess their exposure to smoking cessation interventions and smoking cessation outcomes. RESULTS: Depending on clinic service and intervention component (poster, video segment, provider advice, booklet), 16% to 63% of women reported exposure to an intervention component during their visit. Women in prenatal services received more interventions and had better outcomes than those in the other services. CONCLUSIONS: Exposure to more interventions increased readiness and motivation to quit and the number of actions taken toward quitting. PMID- 10705865 TI - The effects of race/ethnicity and income on early childhood asthma prevalence and health care use. AB - OBJECTIVES: Asthma is the most common chronic illness among US children and is most prevalent in low-income and minority groups. We used multivariate models to disentangle the effects of race/ethnicity, income, and other individual-level risk factors on asthma in a population-based sample of children aged 3 years. METHODS: Data are from the 1988 National Maternal and Infant Health Survey and 1991 Longitudinal Follow-Up. Odds ratios of asthma prevalence, hospitalization, and emergency room use were estimated, with control for socioeconomic characteristics, health behaviors, and insurance. RESULTS: Asthma prevalence, hospitalization, and emergency room use declined with increasing income for non Black but not Black children. CONCLUSIONS: Lifetime income and sociodemographic characteristics do not explain the excess risks of asthma and emergency health care use for asthma among young Black children. PMID- 10705866 TI - Quality of diabetes care in community health centers. AB - OBJECTIVES: This study assessed the quality of diabetes care in community health centers. METHODS: In 55 midwestern community health centers, we reviewed the charts of 2865 diabetic adults for American Diabetes Association measures of quality. RESULTS: On average, 70% of the patients in each community health center had measurements of glycosylated hemoglobin, 26% had dilated eye examinations, 66% had diet intervention, and 51% received foot care. The average glycosylated hemoglobin value per community health center was 8.6%. Practice guidelines were independently associated with higher quality of care. CONCLUSIONS: Rates of adherence to process measures of quality were relatively low among community health centers, compared with the targets established by the American Diabetes Association. PMID- 10705867 TI - Hospitalization of homeless persons with tuberculosis in the United States. AB - OBJECTIVES: This study assessed whether homeless patients are hospitalized for tuberculosis (TB) more frequently and longer than other patients and possible reasons for this. METHODS: We prospectively studied hospitalizations of a cohort of TB patients. RESULTS: HIV-infected homeless patients were hospitalized more frequently than other patients, while homeless patients who had no insurance or whose insurance status was unknown were hospitalized longer. Hospitalization cost $2000 more per homeless patient than for other patients. The public sector paid nearly all costs. CONCLUSIONS: Homeless people may be hospitalized less if given access to medical care that provides early detection and treatment of TB infection and disease and HIV infection. Providing housing and social services may also reduce hospital utilization and increase therapy completion rates. PMID- 10705868 TI - Reaching out to the underserved: a successful volunteer program. PMID- 10705869 TI - The effect of the law on underage drinking and driving. PMID- 10705870 TI - Vaccination strategies for targeted and difficult-to-access groups. PMID- 10705871 TI - A preliminary study of CA15-3, c-erbB-2, epidermal growth factor receptor, cathepsin-D, and p53 in saliva among women with breast carcinoma. AB - A panel of markers used to aid in the diagnosis and treatment of breast cancer was examined in the saliva of a cohort of healthy women, women with benign lesions of the breast, and women with diagnosed breast cancer. We found recognized tumor markers c-erbB-2 (erb), cancer antigen 15-3 (CA15-3), and tumor suppressor oncogene protein 53 (p53) in the saliva of all three groups of women. The levels of erb and CA15-3 in the cancer patients evaluated, however, were significantly higher than the salivary levels of healthy control subjects and benign tumor patients. Conversely, pantropic p53 levels were higher in control subjects compared with those women with breast cancer and those with benign tumors. Although cathepsin-D and epidermal growth factor receptor were detected, they did not demonstrate any clear correlation with disease status. The results of the pilot suggest that these markers have potential use in initial detection and/or follow-up screening for the detection of breast cancer in women. PMID- 10705872 TI - Utility of c-erbB-2 expression in tissue and sera of ovarian cancer patients. AB - In this study, expression of the c-erbB-2 gene in tumors and healthy tissue of patients with ovarian cancer was investigated. Serum c-erbB-2 protein levels were also determined. Elevated serum values were observed in 45% of patients. c-erbB-2 protein levels in the tumors were significantly higher than in healthy tissue. Overexpression of the protein was observed in 60% of patients. However, no association was found between the clinical variables and tumor c-erbB-2 expression. This is the first study in the literature investigating the c-erbB-2 oncoprotein levels in the normal and tumor tissue. We conclude that the role of the c-erbB-2 gene in ovarian cancer warrants further studies. PMID- 10705873 TI - Isolation and characterization of human breast adenocarcinoma cells made resistant to alpha-difluoromethylornithine. AB - Human breast adenocarcinoma cells MCF-7 were selected for resistance to ornithine decarboxylase (ODC) inhibitor, alpha-difluoromethylornithine (DFMO). Stepwise increments of the concentration of DFMO resulted in selection of MCF-7 cells that were capable of growing in the presence of 1.0 mM DFMO. This capacity was associated with a 10-fold increase in ODC activity and marked enhancement in the synthesis rate of ODC protein as verified by a 2-hr [35S]methionine labeling of cellular proteins followed by immunoprecipitation and SDS-PAGE. The resistant cells had much higher concentration of putrescine, spermidine, and spermine than the control cells. A 25-fold increase in ED50 (effective dose causing 50% inhibition) for the antiproliferative action of DFMO in these resistant cells was observed. The susceptibility of wild-type and resistant cell lines to other inhibitors of the polyamine biosynthetic pathway and adriamycin is also reported. PMID- 10705874 TI - Mutation analysis of the putative tumor suppressor gene PTEN/MMAC1 in hepatocellular carcinoma. AB - Loss of heterozygosity of chromosome 10q has been reported in hepatoma. Areas with a high rate of loss of genetic material could harbor putative tumor suppressor genes. PTEN/MMAC1, a candidate tumor suppressor gene located at chromosome 10q23.3, has recently been identified and found to be homozygously deleted or mutated in several different types of human tumors. To determine whether the PTEN/MMAC1 gene is a target of 10q loss of heterozygosity in hepatoma, we examined 42 primary hepatomas for mutations in PTEN/MMAC1 by using nested reverse transcriptase polymerase chain reaction (RT-PCR) of the RNA and single-stranded conformation polymorphism (SSCP) analysis of all genomic exons. Although 2 of 42 hepatoma tissues had aberrant transcripts, 5 matched noncancerous liver tissues also had aberrant transcripts. Southern blot analysis of the entire genomic DNA revealed no genomic change. Therefore, like the TSG101 or FHIT gene, aberrant transcripts of PTEN/MMAC1 using the nested RT-PCR method were a common phenomenon for both cancerous and noncancerous liver tissues, which may not be related to oncogenesis. None of the 42 cases had small deletions, point mutations, or insertions. Our results suggest that the PTEN/MMAC1 gene may not play a role in the pathogenesis of hepatoma. PMID- 10705875 TI - Lack of association between hyperprolactinemia and colon carcinoma. AB - Two recent studies reported that many patients with colorectal carcinoma have elevated serum prolactin (PRL) concentrations and have suggested ectopic PRL secretion as the cause. In the present study, serum PRL was minimally elevated in 16 of 116 colon cancer patients and 2 of 25 control subjects; medications or chemotherapy appeared to be responsible for the PRL elevations in 11 of 16 cancer patients. Serum PRL was not correlated with either plasma carcinoembryonic antigen or disease stage. Preoperative and postoperative serum PRL concentrations were similar in 26 evaluated patients. None of 19 colorectal tumors was positive for PRL staining by immunohistochemistry. Thus, we could not confirm previous reports of frequent hyperprolactinemia in patients with colorectal cancer; factors such as medications, anxiety, pain, and nausea may have raised serum PRL in these earlier studies. Serum PRL is not a useful marker for colon carcinoma, at least in patients in the United States. PMID- 10705876 TI - Fluorescence in situ hybridization: molecular probes for diagnosis of pediatric neoplastic diseases. AB - Fluorescence in situ hybridization (FISH) has become an important tool for diagnosing neoplasia in children. With probes designed to identify specific chromosomes and chromosomal regions, FISH is commonly used to detect the specific chromosomal abnormalities associated with hematologic diseases and solid tumors. Variations of FISH currently being investigated, such as comparative genomic hybridization, multicolor FISH, and microchip arrays, will probably result in additional uses of FISH in both research and clinical cytogenetic laboratories. Although FISH has disadvantages when compared with conventional cytogenetics and molecular methods, FISH will continue to be important in analyzing chromosomal abnormalities of tumors in children. PMID- 10705877 TI - p53 mutation and mitotic infidelity. AB - Chromosome instability (a high frequency of chromosomal loss and gain and genome doubling, often referred to as karyotypic instability) is one of the major characteristics of cancer cells. It facilitates carcinogenesis by increasing the chance of specific mutations responsible for malignant phenotypes. Chromosome instability in most cases reflects the occurrence of defective mitosis, including unequal distribution of chromosomes to daughter cells and failure to undergo cytokinesis, which leads to generation of aneuploid cells. Both in vivo and in vitro, chromosome instability has been shown to correlate with loss or mutation of the p53 tumor suppressor protein, the product of one of the most frequently mutated genes in cancer. The major function of p53 is to prevent cells from proceeding through the cell cycle when cells experience stress, insults, or errors that disturb the preprogrammed cell cycle progression. During the last several years, significant advances have been made in understanding how p53 is involved in the regulation of mitosis and how loss or mutation of p53 affects mitotic fidelity, which will be the subject of this review. PMID- 10705878 TI - Direct correlation between cell loss and depression of labeled thymidine incorporation after irradiation. I. Spleen. AB - The object of this investigation was to determine the relationship between depression of DNA synthesis and cell loss in the spleen after single small radiation doses and to determine whether DNA synthesis after exposure to different radiation modalities can be used as a measure of radiation sensitivity and relative biological effectiveness (RBE). The results presented illustrate, for the first time, that depression of DNA synthesis and cell loss in the spleen after small single doses of irradiation are directly correlated. This correlation indicates that depression of labeled thymidine incorporation and cell loss can be used to determine tissue sensitivity to irradiation. Advance determination of depression of labeled thymidine incorporation or cell loss, using small radiation doses from different radiation modalities, can establish cell killing efficiency and RBE values for each proliferating tissue. PMID- 10705879 TI - Comparative review of 5-HT3 receptor antagonists in the treatment of acute chemotherapy-induced nausea and vomiting. AB - Since their introduction, 5-HT3 receptor antagonists have become the agents of choice in the prevention of acute chemotherapy-induced nausea and vomiting and are generally superior to high-dose metoclopramide regimens. The availability of four different agents (ondansetron, granisetron, dolasetron, and tropisetron) within this class has prompted investigations into potential differences between the drugs, which appear to be few. More importantly, the results of recently conducted randomized comparative trials in patients receiving moderately or highly emetogenic chemotherapy have demonstrated similar efficacy. Although study designs and patient populations differed, seven large comparative trials in patients receiving highly emetogenic chemotherapy reported no significant differences in complete or complete plus major response rates among the agents. Similar results were generally reported in trials evaluating patients receiving moderately emetogenic chemotherapy. The safety and tolerability of these agents also appear to be similar. The most common adverse events include headache, gastrointestinal effects, lightheadedness, and sedation. All agents are available in both intravenous and oral dosage forms and may be administered as a single dose. PMID- 10705880 TI - Arsenic in drinking water and bladder cancer. PMID- 10705881 TI - Surgical diseases of the great vessels. PMID- 10705882 TI - [Prescribing behavior of primary care physicians in diabetes therapy: effect of drug budgeting]. AB - BACKGROUND AND OBJECTIVE: In spite of the great importance of diabetes in Germany, little is known about the medical treatment of diabetic patients by primary health care practices and the effects of the drug budget, introduced by the German Health Care Structure Reform Act (GSG) from 1993. PATIENTS AND METHODS: Computerized data (MediPlus, IMS HEALTH) on prescriptions of the most important drugs were analysed in 2892 diabetic patients of 362 primary care physicians for the period of July 1992 to December 1994. RESULTS: There was an initial decrease in prescriptions per treated patient of antidiabetic drugs and antihypertensive drugs according to the GSG, which was not maintained during the study period. Nevertheless, a cost saving per treated patient with respect to beta-blocker and ACE inhibitors was observed, mainly as a result of a change of preparations and a drop in drug company sales prices. When beginning of a new therapy with oral antidiabetics, the physicians increasingly used acarbose rather than less expensive sulphonylureas. A previous trend of increased use of ACE inhibitors and diuretics for antihypertensive treatment was maintained. In 1993 and 1994, the number of prescriptions and the prescription costs for lipid lowering drugs decreased compared to the values of the last six months of 1992. A global decrease in prescription use of drugs without proven efficacy observed in the first six months of 1993, did not persist. CONCLUSION: The data show, that the drug budget had no relevant long term impact on drug prescribing by internists and general practitioners for diabetic patients. PMID- 10705883 TI - [Factors modifying frequency of publications of clinical research results exemplified by medical dissertations]. AB - BACKGROUND AND OBJECTIVE: The problem of so-called "publication bias"--distortion of statements about an event by selective publication--is receiving growing attention. Assessment of the results of a given study can be affected by knowing the factors that favour or inhibit the publication of its results. METHODS: The data were based on an analysis of 371 medical doctoral dissertations on clinical research. Characteristic features of the dissertations were obtained by questionnaire, supplemented by MEDLINE search and, additionally, by personal enquiry about the publications. Bivariate and multivariate analysis of the characteristic features was used to evaluate the factors that influenced publication of a dissertation. RESULTS: There were mainly three factors that influenced the decision whether or not a dissertation was published: (1) a positive result of the study (odds ratio 2.5, 95% confidence interval 1.12-5.75); (2) the publishing activity of the dissertation's referee (odds ratio 3.8, 95% confidence interval 1.77-7.97; and (3) the quality of the statistical analysis of the dissertation's data (odds ratio 1.67, 95% confidence interval 0.99-2.80). CONCLUSION: A distinct publication bias can be demonstrated also in the case of doctoral dissertations in that positive results are clearly published more often than negative ones. PMID- 10705884 TI - [Diabetic muscle infarct]. AB - HISTORY AND CLINICAL FINDINGS: A 64-year-old diabetic man with secondary failure of treatment with oral hypoglycemic agents was admitted to our clinical department to initiate insulin therapy. The patient was otherwise in good health. Before his dismissal he acutely developed symptoms of pain in his left calf. In addition, the patient was unable to move his left leg and presented palpable pea sized nodules of his left calf. INVESTIGATIONS: Laboratory investigations revealed elevated serum creatine kinase levels on day two after the onset of clinical symptoms (peak value: 2238 U/I). There was evidence for antinuclear antibodies, c-reactive protein was normal. An ultrasound investigation showed a focal edema or bleeding located to the musculus gastrocnemius. Duplex-sonography excluded thrombosis or embolisation. Magnetic resonance imaging showed a diffuse enhancement of signal intensity within the musculus soleus and in areals of the musculus gastrocnemius, as signs of increased blood supply. All clinical findings were consistent with the diagnosis of diabetic muscle infarction. TREATMENT AND COURSE: Under symptomatic treatment with tramadol, diclofenac ointment, and fragmented heparin serum creatine kinase returned to normal levels (105 U/I) within 14 days. In accordance, symptoms of local pain disappeared completely. After two weeks of treatment the patient was able to move his leg without pain. CONCLUSION: This is the first presentation of a patient with diabetic muscle infarction from the onset of symptoms until full recovery. In addition, this case confirms previous descriptions that this condition can be diagnosed by clinical and sonographic findings in combination with magnetic resonance imaging without invasive histologic techniques. PMID- 10705885 TI - [Improvements in palliative treatment of colorectal carcinoma]. PMID- 10705886 TI - [Albert Schweitzer: physician, theologian, philosopher and musician. On his 125th birthday]. PMID- 10705887 TI - [Gynecologist, pathologist, practice partner: who is responsible for undetected pregnancy? Decision of the federal court 29 June 1999]. PMID- 10705888 TI - [Dislocation of a fragment of an indwelling venous catheter]. PMID- 10705889 TI - [Achieving publication as an indicator of high dissertation quality?]. PMID- 10705890 TI - [Actinomycosis of the hand]. PMID- 10705891 TI - Gatifloxacin and moxifloxacin: two new fluoroquinolones. PMID- 10705892 TI - [Neurological complications after roller coaster rides: an emerging new risk?]. AB - OBJECTIVE: To describe neurological complications occurring after roller-coaster rides. PATIENTS AND METHODS: We report 6 cases of complications occurring after roller-coaster rides and analyze published data. RESULTS: Complications seen our patients included 5 cervicoencephalic arterial dissections, one with brainstem dysfunction due to extending syringobulbia. Reported data include one cervicoencephalic arterial dissection, one case of carotid artery occlusion, 3 cases of subdural hematoma, one with subarachnoid hemorrhage, one with cerebrospinal fluid leak, and one with Brown-Sequard syndrome secondary to an enterogenous cyst of the spinal cord. In all patients, pain was the first symptom experienced. In 71.4% of cases, it occurred immediately after the trauma. Marfanis syndrome may be the only risk factor identifiable prior to exposure. The mechanisms of most complications are poorly understood but are likely to involve sudden head and neck flexion-extension movements. CONCLUSION: Neurological complications occurring after roller-coaster rides are highly uncommon, but may leave invalidating sequelae or be fatal. Clinicians should be aware of these complications so these patients can be given proper care early, particularly at the stage when pain is the only sign. Early management could help limit the consequences of these complications. PMID- 10705893 TI - [Importance of medicolegal data for national cause of death statistics]. AB - OBJECTIVE: The validity of French cause of death statistics largely depends on the transmission of the medical information reported on the dedicated part of the death certificate. Data referring to death causes are collected and analyzed at the Information Department on the cause of death (INSERM SC8). In major urban areas with one or more forensic institutions, it has been observed that the proportion of death certificates indicating ino particular cause of deathi is higher than the national average. We therefore examined deaths occurring in 1996 in the Lyons region. METHODS: This retrospective study involved 5,208 deaths recorded in Lyons France in 1996. Complementary information on these deaths was obtained from INSERM SC8 and Lyons Medicolegal Institute. RESULTS: It was observed that a large portion of violent deaths occurring in this city in 1996 (more than 30%) were not recorded appropriately in the national statistics due to a lack of communication after autopsy. CONCLUSION: Considering the importance of epidemiology and suicide prevention in the national public health strategy, improved information transmission must be developed in the near future. PMID- 10705895 TI - [In Process Citation] PMID- 10705894 TI - [Pseudotumoral abdominal granuloma concomitant with immune reconstitution after antiretroviral treatment]. AB - BACKGROUND: Use of powerful multiple-drug antiretroviral regimens can significantly raise CD4+ counts restoring immune function, but in certain cases, leading to inflammatory reactions. CASE REPORT: An HIV-infected patient developed a mycobacteriosis of the digestive tract when his CD4 count fell below 5/mm3. He was given antimycobacterial treatment in combination with an effective triple antiretroviral regimen. At two years, the clinical situation was controlled with persistent optimal response (CD4 = 338/mm3 HIV-RNA < 500 copies/ml); the antimycobacterial regimen was discontinued. One year later the patient still had a CD4+ count above 500/mm3 but developed a voluminous mesenteric mass invaded by a CD68+ histiocyte proliferation. No causal agent could be identified. The clinical course was favorable after reintroducing antimycobacterial treatment combined with short-term corticosteroid therapy. DISCUSSION: Reconstitution of the immune system after long-term use of the new antiretroviral therapies raises the question of whether anti-infectious prophylaxis should be maintained. However, possible reactions to earlier pathogens after restoration of specific immunity would warrant secondary prophylaxis even in patients responding to powerful antiretroviral combinations. PMID- 10705896 TI - [Does cannabis play a role in scuba diving accidents?]. PMID- 10705897 TI - [Course of hepatocarcinoma after alcoholization in patients with cirrhosis and partial portal vein thrombosis]. PMID- 10705898 TI - [Infarction: thrombolysis or angioplasty? That's the question...]. PMID- 10705899 TI - [Management of patients with chronic pain]. PMID- 10705900 TI - [Whiplash injury of the cervical spine]. AB - SOFT TISSUE INJURY: Whiplash injuries of the cervical spine are common, usually after traffic accidents. Soft tissue injuries may involve the muscles, ligaments, disks, and facet joints (excluding severe cases of fracture or dislocation not discussed here). CLINICAL MANIFESTATIONS: There are 2 cardinal signs: neck pain and headache. These signs may be associated or not with other symptoms. CLINICAL COURSE: Most cases follow a benign course and resolve within a few weeks or months. Symptoms persist however in 20 to 40% of the cases. Evolutive organ injury, sensitization of the central nervous system, and secondary psychological reactions (including desire for financial compensation) all contribute to the development of a chronic situation. MEDICOLEGAL ASPECTS: Excepting cases with over neurological complications, it is particularly difficult to assess the nature and seriousness of whiplash injuries with physical examination and imaging. This diagnostic difficulty explains the frequency of medicolegal procedures. It is particularly difficult to treat chronic forms. If a medicolegal procedure is in course, it is advisable that the expert reports and fair compensation be established as rapidly as possible. PMID- 10705901 TI - [Macrophagic myofasciitis. Study and Research Group on Acquired and Dysimmunity related muscular diseases (GERMMAD)]. AB - MACROPHAGIC MYOFASCIITIS: A most unusual inflammatory myopathy, first described by Germmad had been reported with increasing frequency since 1993 in the leading French myopathology centers. We present our experience with this new disease: macrophagic myofasciitis. CLINICAL FEATURES: By November 1999, 70 cases of macrophagic myofasciitis had been recorded since our first description. The first 22 patients (sex ratio M/F = 1:3) referred with the presumptive diagnosis of polymyositis (n = 11), polymyalgia rheumatica (n = 5), mitochondrial cytopathy (n = 4), and congenital myopathy or muscle dystrophy (n = 1 each). Symptoms included myalgia (91%), anthralgia (68%), marked asthenia (55%), muscle weakness (45%), and fever (32%). LABORATORY FINDINGS: Abnormal laboratory findings included elevated CK levels (50%), markedly increased erythrocyte sedimentation rate (37%), and myopathic EMG (35%). Muscle biopsy showed a unique myopathological pattern characterized by: i) centripetal infiltration of epimysium, perimysium and perifascicular endomysium by sheets of large cells of the monocyte/macrophage lineage (CD68+, CD1a-, S100-, with a PAS-positive content; ii) absence of necrosis, of both epithelioid and giant cells, and of mitotic figures; iii) presence of occasional CD8+ T-cells; iv) inconspicuous muscle fiber damage. The picture was easily distinguishable from sarcoid myopathy and fasciitis panniculitis syndromes. The infectious diseases know to be associated with reactive histiocytes, including Whippleis disease, Mycobacterium avium intracellulare infection and malakoplakia, could not be documented. Patients improved under corticosteroid therapy and/or immunomodulatory therapeutic CONCLUSION: A new inflammatory muscle disorder, characterized by a distinctive pathological pattern of macrophagic myofasciitis is emerging in France. PMID- 10705902 TI - [Extrahepatic manifestations of hepatitis C virus]. AB - ACCEPTED ASSOCIATIONS: Extrahepatic manifestations of the hepatitis C virus (HCV) have been reported in a large body of literature. It is now generally accepted that the HCV is strongly associated with so-called essential cryoglobulinemia, membrane proliferative glomerulonephritis with or without cryoglobulinemia, sporadic porphyria cutanea tarda, and with lymphocyte sialadenitis with or without a dry mouth syndrome. PROBABLE ASSOCIATIONS: Several epidemiological arguments suggest that HCV could play a role in certain types of non-Hodgkin lymphomas. HCV is also sometimes implicated in periarteritis nodosa and with peripheral neuropathies with lymphocyte vasculitis. UNPROVEN ASSOCIATIONS: The role of HCV in autoimmune dysthyroidism, lichen planus, diabetics mellitus, and antiphospholipid syndromes is a subject of debate. There have also many sporadic reports of manifestations of syndromes suggesting, though not proving, an association with the hepatitis C virus. PMID- 10705904 TI - [Antibiotic strategies: good practices in intensive care]. PMID- 10705903 TI - [Mode of action of glucocorticoids]. AB - RECEPTORS: The effect of glucocorticoids is mediated by a receptor mainly found in the cytoplasm. The glucocorticoid receptor is a member of the nuclear receptor superfamily. RECEPTOR ACTIVATION: When unstimulated, the glucocorticoid receptor is inactivated by its integration within a multiple-protein complex associating heat shock proteins, immunophilins and cyclophilins. When the hormone binds to its receptor, the complex dissociates and the receptor migrates to the nucleus. In the nucleus, the activated receptor provokes an upregulation or downregulation of target gene expression. GENE REGULATION: Gene expression may be regulated via an interaction between specific nucleic acid sequences of the glucocorticoid receptor and nuclear DNA (direct mechanism) or by protein-protein interactions (indirect mechanism). The expression of target genes is either inhibited or stimulated. PHARMACOLOGICAL CONSEQUENCES: The main pharmacological applications of glucocorticoids can be explained by these different mechanisms. The antiinflammatory action of glucocorticoids results from an inhibition of the transcription of the collagenase gene via an interaction with the AP-1 transcription factor. The anticancer action of glucocorticoids results from the induction of apoptotic cell death via a mechanism which would require the transcriptional activity of the glucocorticoid receptor. PMID- 10705905 TI - Mission-based management: tranquility or tumbrels of our universe? PMID- 10705906 TI - Hostility and the metabolic syndrome in older males: the normative aging study. AB - OBJECTIVE: Several studies have shown that hostility, as measured by the Minnesota Multiphasic Personality Inventory-derived Cook-Medley Hostility Scale (Ho), is positively associated with several cardiovascular risk factors, possibly accounting for the relationship between Ho scores and cardiovascular mortality. This study was undertaken to examine associations between hostility and cardiovascular risk factors representing the metabolic syndrome in 1,081 older men who participated in the Normative Aging Study. METHODS: Subjects included men who completed the Minnesota Multiphasic Personality Inventory in 1986 and who participated in a subsequent laboratory examination within 1 to 4 years. Total and subscale Ho scores were computed, and associations with anthropometric data, cigarette smoking, dietary information, serum lipids, blood pressure, and fasting glucose and insulin levels were examined. RESULTS: The total Ho score was positively associated with waist/hip ratio, body mass index, total caloric intake, fasting insulin level, and serum triglycerides. The Ho score was inversely related to education and high-density lipoprotein cholesterol concentration. Path analysis also suggested that the effects of hostility on insulin, triglycerides, and high-density lipoprotein cholesterol were mediated by its effects on body mass index and waist/hip ratio, which, in turn, exerted their effects on lipids and blood pressure through insulin. CONCLUSIONS: The results are consistent with those of prior research and also suggest that, in older men, hostility may be associated with a pattern of obesity, central adiposity, and insulin resistance, which can exert effects on blood pressure and serum lipids. Furthermore, effects of hostility on the metabolic syndrome appear to be mediated by body mass index and waist/hip ratio. PMID- 10705907 TI - Hostile attitudes predict elevated vascular resistance during interpersonal stress in men and women. AB - OBJECTIVE: Existing research indicates that hostility is associated with enhanced blood pressure responses during social stress, but little is known about the hemodynamic patterns underlying these blood pressure increases, particularly in women. The present study examined hemodynamic responses to a low-anger interpersonal stressor, testing the hypotheses that hostile individuals show enhanced vascular responses and that low hostile individuals show enhanced myocardial responses. METHODS: Eighty undergraduate men and women were categorized as high or low in hostility on the basis of median splits of Cook Medley Hostility Scale scores. Participants discussed a controversial topic with a confederate who disagreed with them, and hemodynamic responses were assessed with impedance cardiography. RESULTS: High hostile individuals exhibited greater increases in diastolic blood pressure and total peripheral resistance and smaller increases in cardiac output during an interpersonal stressor than did low hostile individuals. Systolic blood pressure and heart rate increases were greater among high hostile relative to low hostile females and comparable among low and high hostile males. Affective responses and task perceptions were generally similar for high and low hostile participants, but the relationship between task perception and hemodynamic responses varied on the basis of hostility level. CONCLUSIONS: These findings suggest that hostility in both men and women is associated with heightened vascular and dampened cardiac responsivity to interpersonal stress that is not deliberately anger provoking. Moreover, they indicate that the associations between task perception and hemodynamic responses vary between high and low hostile individuals. PMID- 10705908 TI - Intrusive traumatic recollections and comorbid posttraumatic stress disorder in depressed patients. AB - OBJECTIVE: Recent studies have found evidence of the presence and role of intrusive traumatic memories in depressed patients. In this study, we attempted to replicate these findings, examining the full range of early and later traumatic events, as well as comorbid posttraumatic stress disorder, in male and female depressed patients. METHODS: Sixty-nine outpatients meeting criteria of the Diagnostic and Statistical Manual of Mental Disorders, fourth edition, for major depressive episode were recruited from the outpatient department of an academic hospital. RESULTS: Seventy-five percent of the depressed patients were found to have had one or more early and/or more recent traumatic experiences. The symptom category of reexperiencing was diagnosed in 48% of these trauma-exposed respondents. Comorbid posttraumatic stress disorder was diagnosed in 13% of the total sample. CONCLUSIONS: The findings show that depressed patients are highly likely to have experienced traumatic events and intrusive traumatic recollections. Future research should focus on the direction of any causal relationship between trauma, reexperiencing, posttraumatic stress disorder, and depression. PMID- 10705909 TI - Trait anxiety and reactions to patient-centered and doctor-centered styles of communication: an experimental study. AB - OBJECTIVE: A patient-centered model of communication has often been advocated in preference to a doctor-centered model. The aim of the present study was to assess in an experimental setting how subjects' general level of anxiety affects their reactions to these two communication styles as measured by emotional reactions and satisfaction immediately after consultation. METHODS: Twenty students with low trait anxiety and 21 students with high trait anxiety each had a single consultation with a physician who performed the consultation using either a patient-centered or doctor-centered style of communication. Questionnaires about emotional state were completed by the students before and after the consultation, and a questionnaire about satisfaction was completed after the consultation. RESULTS: Students with low trait anxiety were significantly more satisfied with a patient-centered than a doctor-centered style of communication. There were no significant differences in emotional response to the two styles of communication. Students with high trait anxiety reacted emotionally more positively to a doctor centered communication style, with significant and nearly significant change scores for the emotions of tension/anxiety and vigor/activity, respectively. No significant difference was found between satisfaction scores. CONCLUSIONS: Data indicate that differences between subjects' emotional traits may be of importance for a differentiated response to patient-centered and doctor-centered communication styles. Subjects' trait anxiety seems to be a significant factor that should be taken into account when assessing the effects of different communication styles. PMID- 10705911 TI - The relationship between menstrual cycle phases and suicide attempts. AB - OBJECTIVE: The validity of prior studies on the menstrual cycle and suicide attempts assumes that suicidal women accurately describe their cycles. The three objectives of this study were 1) to explore whether prior inconsistencies are due to the effects of sample selection and method of assessment of the menstrual cycle, 2) to assess the relationship between the menstrual cycle phase and suicide attempts, and 3) to establish the role of sexual hormones in suicide attempts. METHODS: The original sample included 134 women who came to the emergency room of a general hospital after a suicide attempt. One hundred eight female blood donors were recruited as control subjects. The menstrual cycle was divided into follicular, midcycle, and luteal phases using two clinical methods and serum hormonal assessment. Dividing the follicular phase into menstrual and nonmenstrual phases was also considered. RESULTS: Two of 11 previously used sampling methods produced a sample size similar to that of the hormonal assessment. kappa values between the two clinical and the endocrinological methods were low (0.40-0.50). The number of suicide attempts during the follicular phase (particularly during the menstrual phase) was significantly higher than expected. CONCLUSIONS: Despite the inability to control for other variables and limitations, the results of this study suggest that sample selection could introduce biases and that studies relating psychiatric symptomatology and menstrual cycle phases need to use hormonal determinations. New studies are needed to verify that suicide attempts are more frequent during the follicular phase (particularly during the menstrual phase). PMID- 10705910 TI - Salivary MUC5B-mediated adherence (ex vivo) of Helicobacter pylori during acute stress. AB - OBJECTIVE: Biochemical host defenses at mucosal sites, such as the oral cavity, play a key role in the regulation of microbial ecology and the prevention of infectious disease. These biochemical factors have distinct features, some of which benefit the host and some that benefit bacteria. We investigated the effects of acute stress on the salivary levels of the carbohydrate structure sulfo-Lewis (sulfo-Le), which is linked to the mucosal glycoprotein MUC5B. Sulfo Le was recently identified as an adhesion molecule for Helicobacter pylori; therefore, we also measured saliva-mediated adherence (ex vivo) of H. pylori. The oral cavity is suspected to be involved in the transmission of H. pylori. METHODS: Saliva was collected from 17 undergraduates before (baseline), during (stress), and after (recovery) exposure to a video showing surgical procedures. In addition, blood pressure, an impedance cardiogram, and an electrocardiogram were recorded. RESULTS: During stressor exposure, participants reported increased state anxiety. In addition, stroke volume increased and heart rate decreased. The stressor induced a strong increase in salivary sulfo-Le concentration (U/ml), sulfo-Le output (U/min), sulfo-Le/total protein ratio (U/mg protein), and saliva mediated adherence (ex vivo) of H. pylori. As expected, sulfo-Le concentration correlated with the adherence of H. pylori (r = 0.72, p < .05). It was demonstrated that the observed adherence was induced by MUC5B and that the carbohydrate structure sulfo-Le contributed to this process. CONCLUSIONS: Our study demonstrated a direct link between stress-mediated biochemical changes and altered host-microbe interactions in humans. Increased bacterial adherence may be a contributing factor in the observed relationship between stress and susceptibility to infectious disease. PMID- 10705913 TI - Coping profile differences in the biopsychosocial functioning of patients with temporomandibular disorder. AB - OBJECTIVE: The objective of this study was to evaluate whether biopsychosocial functioning differences exist between samples of patients with temporomandibular disorder (TMD) who differ in coping profiles as assessed by the Multidimensional Pain Inventory. METHODS: A total of 322 patients who presented with TMD were administered a comprehensive biopsychosocial assessment battery, and the acute or chronic status of their disorder was determined. A follow-up evaluation was conducted 6 months later to determine the status of their pain. RESULTS: TMD patients with dysfunctional and interpersonally distressed coping profiles demonstrated more acute and chronic psychosocial difficulties than patients with adaptive coper profiles. The data also suggest that having a dysfunctional/distressed coping profile on the Multidimensional Pain Inventory has some predictive value in determining the likelihood of developing chronicity in the absence of treatment. CONCLUSIONS: The presence of a dysfunctional/distressed coping profile in patients with TMD is likely to provide clinicians with important information about the biopsychosocial functioning of those patients, which, in turn, will help to determine the most effective treatment modalities to use with TMD patients. PMID- 10705912 TI - Mood states associated with transitory changes in asthma symptoms and peak expiratory flow. AB - OBJECTIVE: This study examined the within-person relations between transitory changes in mood, asthma symptoms, and peak expiratory flow rate (PEFR). METHODS: Thrice-daily for 21 consecutive days, 48 adults with moderate to severe asthma entered information in palm-top computers about their mood and asthma symptoms. A multidimensional model of mood, ie, the mood circumplex, informed the assessment of mood arousal and mood pleasantness. At each observation, participants also recorded their PEFR with peak flow meters that stored blinded data. Albuterol doses were also monitored electronically. Before and after the 21-day study, spirometric measures of airways obstruction were taken under controlled conditions. RESULTS: Random effects regression models revealed a significant, but weak, within-person relation between symptoms and PEFR. Changes in mood vectors with an arousal component were significantly related to PEFR changes, whereas changes in mood vectors with a pleasantness component tracked changes in asthma symptom reports, even after adjustment for contemporaneous PEFR and after controlling for time of day and albuterol dosing. Comparison of spirometric assessments with unsupervised PEFR suggested that part of the relation between mood arousal and PEFR may be attributable to the "effort-dependence" of peak flow self-monitoring. CONCLUSIONS: Different dimensions of mood were associated with transitory changes in asthma symptoms and PEFR. This may be one reason why individuals with asthma misperceive the severity of their symptoms in relation to underlying airways obstruction. PMID- 10705914 TI - Emotional abuse, self-blame, and self-silencing in women with irritable bowel syndrome. AB - OBJECTIVE: The purpose of this study was to investigate the presence of emotional abuse and two psychosocial constructs (self-blame and self-silencing) in a sample of women diagnosed with irritable bowel syndrome (IBS) relative to a comparison sample of women diagnosed with inflammatory bowel disease (IBD). METHODS: Women diagnosed with IBS (N = 25) were compared with women diagnosed with IBD (N = 25) on measures of history of abuse, self-blame, and self-silencing. RESULTS: It was found that women in the IBS sample scored significantly higher on emotional abuse, self-blame, and self-silencing than did women in the IBD sample. These three variables were also found to be significantly intercorrelated in both the IBS and IBD samples. Finally, emotional abuse was significantly higher in IBS patients than in IBD patients beyond the differences accounted for by physical and/or sexual abuse history. CONCLUSIONS: These findings empirically demonstrate an association between IBS and emotional abuse, as well as a possible connection with psychosocial variables, that may mediate the connection between emotional abuse and functional bowel symptoms. We suggest that these variables be further evaluated in the context of clinically relevant research on IBS. PMID- 10705915 TI - Immunological effects of massage therapy during academic stress. PMID- 10705917 TI - Foundations of molecular therapies in breast carcinoma. PMID- 10705916 TI - Anxiety, depression, and heart rate variability. PMID- 10705918 TI - The ErbB receptors as targets for breast cancer therapy. AB - Breast carcinomas express high levels of ErbB receptors and their ligands, and their overexpression has been associated with a more aggressive clinical behavior. For these reasons therapies directed at these receptors have the potential to be useful anti-cancer treatments. A series of monoclonal antibodies (MAbs)3 directed against the EGF (ErbB1) receptor and the closely related HER2/Neu (ErbB2) receptor are currently under evaluation. These MAbs have shown promising preclinical activity and "chimeric" and "humanized" MAbs have been produced in order to obviate the problem of host immune reactions. These antibodies are currently being tested in clinical trials either alone or in combination with chemotherapeutic agents. Clinical activity with one of these antibodies, trastuzumab, a humanized anti-ErbB2 MAb, has been documented in patients with breast cancer in a series of clinical trials and has recently been approved for the therapy of patients with metastatic ErbB2 overexpressing breast cancer. In addition to antibodies, compounds that inhibit receptor tyrosine kinases have shown significant preclinical activity and are currently being evaluated in the clinic. PMID- 10705919 TI - Immunotherapeutic approaches for the treatment of breast cancer. AB - The application of immunotherapeutic principles to the treatment and prevention of breast cancer is a relatively new undertaking. Although cytokine infusions, cancer vaccines, and T cell therapy have been extensively studied in solid tumors such as melanoma and renal cell carcinoma, the therapeutic efficacy of these approaches is not well explored in breast cancer. The recent definition of tumor specific immunity in breast cancer patients and the identification of several breast cancer antigens has generated enthusiasm for the application of immune based therapies to the treatment of breast malignancies. In general, immunotherapies can be considered either non-specific, such as a general immunomodulator (e.g., a cytokine), or tumor-specific (e.g., a vaccine that targets breast cancer tumor antigens). This review describes three major immunotherapeutic strategies that have the potential to enhance or generate an anti-breast cancer T cell immune response: (i) cytokine therapy; (ii) cancer vaccines; and (iii) T cell therapy, and explores how each strategy has been applied to the treatment of breast cancer. PMID- 10705920 TI - Integrin function in breast carcinoma progression. AB - The differentiation and function of mammary epithelial cells is dependent upon the combined action of growth factor/hormone receptors and integrin receptors, which act in concert to control the signals required for normal cell function. It is now becoming clear that integrin receptors also contribute to carcinoma cell behavior and that alterations in expression and function during transformation have a large impact on breast carcinoma progression. The focus of this review is to discuss integrin-dependent functions that can be manipulated as targets for the therapeutic intervention of breast cancer. A combination of correlative and mechanistic studies have contributed to the identification of specific integrin receptors, namely alpha2beta1, alpha6beta1, and alpha6beta4, implicated in breast carcinoma progression. Although this field is still emerging and much remains to be learned, potential integrin-dependent signaling targets have been identified. PMID- 10705922 TI - Cell cycle checkpoints as therapeutic targets. AB - Most human breast tumors arise from multiple genetic changes which gradually transform differentiated and growth-limited cells into highly invasive cells that are unresponsive to growth controls. The genetic evolution of normal breast cells into cancer cells is largely determined by the fidelity of DNA replication, repair, and division. Cell cycle arrest in response to DNA damage is an important part of the mechanism used to maintain genomic integrity. The control mechanisms that restrain cell cycle transition after DNA damage are known as cell cycle checkpoints. This review will focus on cell cycle checkpoint signaling pathways commonly mutated in human breast tumors and suggest how different components of these checkpoint pathways offer the potential for chemotherapeutic intervention. PMID- 10705921 TI - Role of retinoid receptors in the prevention and treatment of breast cancer. AB - Retinoids are vitamin A-related compounds that have been found to prevent cancer in animals and humans. In this review, we discuss the role of retinoids and their receptors in the treatment and prevention of breast cancer. The retinoid receptors are expressed in normal and malignant breast cells, and are critical for normal development. In breast cells, when bound by retinoid hormones, these proteins regulate proliferation, apoptosis, and differentiation. The mechanism by which retinoids inhibit breast cell growth has not been completely elucidated, however, retinoids have been shown to affect multiple signal transduction pathways, including IGF-, TGFbeta-, and AP-1-dependent pathways. Retinoids have also been shown to suppress the growth and prevent the development of breast cancer in animals. These agents suppress tumorigenesis in carcinogen-treated rats and in transgenic mice, and inhibit the growth of transplanted breast tumors. These promising preclinical results have provided the rationale to test retinoids in clinical trials for the treatment and prevention of breast cancer. Several retinoids, including all trans retinoic acid and 9-cis retinoic acid, have been shown to have modest activity in the treatment of breast cancer, and these agents are now in clinical trials in combination with cytotoxic agents and anti estrogens. Another retinoid, 4-HPR, is currently being tested in a human cancer prevention trial. Preliminary results suggest that 4-HPR may suppress breast cancer development in premenopausal women. Future clinical trials will focus on testing new synthetic retinoids that have reduced toxicity and enhanced therapeutic and preventive efficacy. PMID- 10705923 TI - The estrogen receptor: a logical target for the prevention of breast cancer with antiestrogens. AB - A strategy for the prevention of breast cancer has been refined over the last century beginning with the first observation that oophorectomy caused disease regression in some patients, to the identification of the estrogen receptor some 60 years later, and finally to the synthesis of the first nonsteroidal antiestrogen. Tamoxifen was the first clinically useful antiestrogen and has been used for the treatment of breast cancer for the last twenty-one years in the United States. It is therefore a logical progression that antiestrogens are now recognized as useful agents for the prevention of breast cancer. We will discuss the estrogen receptor as a target for the treatment and now the prevention of breast cancer. Data from the National Surgical and Bowel Project (NSABP)4 tamoxifen prevention trial will be discussed with the preliminary results of two other European studies. The status of breast cancer prevention to date involves the comparison of the current standard of prevention, tamoxifen, with the osteoporosis prevention drug, raloxifene in an ongoing trial called Study of Tamoxifen and Raloxifene (STAR). PMID- 10705925 TI - Management of intrinsic gliomas of the tectal plate in children. A ten-year review. AB - The natural history, management, and long-term outcome for patients with benign, intrinsic tectal plate gliomas remain controversial in spite of their propensity to cause late-onset hydrocephalus. A 10-year retrospective review has identified 11 consecutive children with tectal plate lesions. Headache, vomiting, a decline in school performance, tremor, and complex partial seizures were common presenting symptoms. All patients presented with signs and symptoms of hydrocephalus. Magnetic resonance (MR) imaging delineated an intra-axial mass lesion of the midbrain primarily localized to the tectal plate which uniformly was hyperintense on T2-weighted imaging and had a more variable appearance on T1 weighted imaging and rare enhancement with gadolinium. No patient underwent surgical resection, chemotherapy, or radiotherapy. Three of 11 patients (27%) showed evidence of progression in size or a new focus of enhancement on MR imaging, which was clinically asymptomatic. In this series, no patient with a tectal plate lesion less than 1.5 cm in maximal diameter and without gadolinium enhancement showed any evidence of clinical or radiological progression. Although intrinsic tectal lesions in children are clinically indolent and the initial management consists of CSF diversion, these lesions may eventually progress and still warrant long-term follow-up with serial MR imaging. PMID- 10705924 TI - Angiogenesis as a target for breast cancer therapy. AB - Angiogenesis, the development of new blood vessels, is crucial for the growth of both primary tumors and metastases beyond a minimal size and the vasculature of tumors facilitates their metastatic spread. Inhibition of angiogenesis is thus seen as a potentially useful approach to anti-metastasis therapy, and is an area of active research and development. Here we discuss this therapeutic approach in the context of breast cancer. An overview of the contribution of angiogenesis to tumor development is provided and current treatment options for breast cancer are briefly summarized. Assessment of angiogenesis in primary breast tumors has been shown to provide independent prognostic information. There are opportunities for the application of anti-angiogenesis therapeutic strategies in the treatment of breast cancer. Clinical trial design must take into account the unique properties of anti-angiogenic agents to fully assess their potential clinical benefit. PMID- 10705926 TI - Spinal cord tethering associated with amniotic band syndrome. AB - Amniotic band syndrome (ABS) comprises fetal morphological abnormalities that may be associated with fibrous amniotic bands that damage developing fetal parts resulting in cutaneous scars, erosions and ulcerations, digital constricting bands, craniofacial and visceral anomalies. Multiple asymmetric encephaloceles and anencephaly are neural-tube-like defects previously reported with ABS. This is the first report of spinal dysraphism with dorsal spinal cord tethering associated with ABS. We examine the pathogenetic theories of ABS in light of this report. PMID- 10705927 TI - Effect of intrauterine myelomeningocele repair on central nervous system structure and function. AB - BACKGROUND: It has been postulated that intrauterine myelomeningocele repair might improve neurologic outcome in patients with myelomeningocele. A total of 59 such procedures have been performed at Vanderbilt University. Preliminary results suggested that the degree of hindbrain herniation is reduced by intrauterine repair. In an attempt to further quantify the possible benefits of this surgery, a subset of these patients was brought back to Vanderbilt for study. METHODS: A group of 26 patients who had undergone intrauterine myelomeningocele repair underwent an extensive evaluation which included manual muscle testing, MR imaging and precise determination of the anatomic level of their lesions as well as multiple other tests. The results of this analysis were compared to those in 2 groups of historical controls. RESULTS: In this group of patients intrauterine myelomeningocele repair substantially reduced the incidence of moderate to severe hindbrain herniation (4 vs. 50%). The incidence of shunt-dependent hydrocephalus was more modestly reduced (58 vs. 92%). The average level of leg function closely matched the average anatomic level of the lesion in both the fetal surgery and control groups. CONCLUSION: The most dramatic effect of intrauterine repair appears to be on hindbrain herniation. A less dramatic, but significant, reduction in shunt-dependent hydrocephalus is also seen. Prospective patients should be cautioned not to expect improvement in leg function as the result of this surgery. The potential benefits of surgery must be carefully weighed against the potential risks of prematurity. PMID- 10705928 TI - Is indomethacin harmful in spinal cord injury treatment? An experimental study. AB - This study was designed to analyze the effect of early indomethacin on the lipid peroxidation after spinal cord injury in rats. The use of anti-inflammatory drugs to affect delayed and secondary injury after trauma to the spinal cord has now become a matter of standard clinical practice. However, spinal cord injury remains an enormous clinical problem and research that may lead to improved treatment is to be encouraged and commended. Three experimental groups consisting of 40 rats each were formed. Using microsurgical technique, total laminectomy between T5 and T10 was performed. Spinal cord injury was achieved with an epidural aneurysm clip, and pharmacological treatment immediate after the injury was performed by injecting indomethacin intraperitoneally (i.p.) at a dose of 3 mg/kg to indomethacin-treated group. The three main groups were divided into subgroups of 8 rats each. It was planned to stop the biochemical reactions at a different time in each of these subgroups, by the application of liquid nitrogen to the spinal cord and paravertebral structures at the end of the 1st, 15th, 30th, 60th, and 90th minutes. All the spinal cords were removed and protected from further reactions by immersing in the liquid nitrogen tank. The lipid peroxidation levels were assessed by determining thiobarbituric acid reactive substances formation. The results of the study showed that the administration of 3 mg/kg indomethacin immediately after spinal cord injury induces lipid peroxidation to a significant degree (p<0.05 one-way ANOVA and Tukey HSD tests) when compared to the saline-treated group. This result suggests that early posttraumatic indomethacin treatment may be harmful in spinal cord injury. PMID- 10705929 TI - Propofol for sedation and control of intracranial pressure in children. AB - Following central nervous system insults, control of intracranial pressure may lessen the incidence of morbidity and mortality. Therapies to control intracranial pressure include osmolar agents, prevention of and control of seizures, drainage of cerebrospinal fluid, hypothermia, and barbiturates. Control of agitation and excessive patient movement are additional components in the management of ICP. Although opioids and benzodiazepines are generally effective, in a small subset of patients, alternative agents may be necessary. The authors present 2 children with increased ICP in whom propofol was used to provide sedation and control ICP. The use of propofol in this setting and its possible applications in the children with increased ICP are discussed. PMID- 10705930 TI - Intracranial nasal dermoid sinus cyst associated with colloid cyst of the third ventricle. Case report and new concepts. AB - A case of a 16-year-old male with both a nasal dermoid sinus cyst (NDSC) and a third ventricle colloid cyst is presented. The NDSC was excised via a single stage combined intracranial-extracranial approach and the third ventricle colloid cyst was resected endoscopically. The pathogenetic theories of NDSC and third ventricle colloid cyst are discussed, and an embryological explanation for the simultaneous development of the two lesions in this patient is explored. This case is best classified among congenital developmental malformations in a category we propose to call 'anterior neuropore corridor defects.' PMID- 10705931 TI - A case report of caudal regression syndrome associated with an intraspinal arachnoid cyst. AB - We report here a rare case of caudal regression syndrome associated with an intraspinal arachnoid cyst. The patient was a 6-month-old baby girl with multicomplex congenital abnormalities: sacrococcygeal dysgenesis and ventral curvature, large terminal cyst (myelocystocele), spinal arachnoid cyst, cerebellar hypertrophy (suspected), high imperforate anus, partial dysgenesis of the large intestine, omphalocele, atresia of the vagina, bilateral incomplete ureter duplication, incomplete pseudoduplicated bladder and bilateral talipes equinovarus. We performed plastic repair of the myelocystocele and perineal lesion for caudal regression syndrome and partial removal of the cyst wall for the intraspinal arachnoid cyst. She has been well for 3 years postoperatively, and her mental development is normal. PMID- 10705932 TI - Optic glioma with characteristic bilateral optic atrophy in a 3-year-old girl. AB - We report a case of optic glioma with bilateral optic atrophy. A 3-year-old girl presented with vomiting and left hemiparesis. She had hypothalamic dysfunction, right ptosis, right monocular nystagmus, left facial palsy, left hemiparesis, and left pes adductus. Neuroimaging studies showed obstructive hydrocephalus with a large suprasellar calcified tumor with a ring-like enhancement mimicking craniopharyngioma. Visual-evoked potentials showed delayed latency of N75 in the right occipital lead. The tumor, a pilocytic astrocytoma in the right optic tract and chiasma, was partially removed via a right frontotemporal craniotomy. The right optic nerve had shrunk to half the normal diameter and became twisted downwardly. Intracranial pressure (ICP) increased to 40 cm H2O. The fundus had bilateral optic atrophy without disc swelling. To our knowledge, this is the first report of a lamina/dot sign of the optic disc in a small child with a brain tumor and a normal neuroretinal fiber layer. These ocular findings may result from possible interruption of the axonal flow caused by the tumor and not increased ICP. PMID- 10705933 TI - Aneurysmal bone cyst of the temporal bone. AB - OBJECTIVE AND IMPORTANCE: Aneurysmal bone cysts (ABC) are benign bone neoplasms which typically involve the spine and long bones. We present a rare case of an ABC in the temporal bone with significant cerebellar compression. CLINICAL PRESENTATION: The patient was a young boy who presented with several weeks of left posterior auricular pain. Computed tomography, magnetic resonance imaging and angiogram showed an expansile bone neoplasm involving the left petrous temporal bone. INTERVENTION: A retrosigmoid-transmastoid craniectomy was performed, and total removal of the bone lesion was achieved. The pathological examination revealed the diagnosis of ABC. CONCLUSION: The postoperative course was uneventful, and the imaging studies demonstrated total removal of the neoplasm. This case represents the imaging and surgical management of a rare ABC in the temporal bone. This diagnosis should be considered in the differential of bone neoplasms in this region. PMID- 10705934 TI - Images in pediatric neurosurgery. Diastematomyelia. PMID- 10705935 TI - ACTH activates rapid eye movement-related phasic inhibition during REM sleep in patients with infantile spasms. AB - OBJECTIVES: Phasic inhibition index (PII) is the rate of the simultaneous occurrence of phasic chin muscle activity (PCMA) and rapid eye movement bursts during rapid-eye-movement sleep (REMS). In naive patients with infantile spasms (IS), the PII value was found to reflect their prognosis. We studied the effects of adrenocorticotropic hormone (ACTH) on REMS components including PII in IS. METHODS: REMS parameters were examined in 18 IS patients before and after ACTH treatment. The effects of corticosteroids (CSs) were examined in 3 patients with congenital adrenal hyperplasia (CAH) and 3 with nephrotic syndrome (NS). RESULTS: ACTH decreased PII and PCMA in IS patients. In CAH patients, physiological doses of CSs corrected the increased intrinsic ACTH level and increased PII. In NS patients, therapeutic doses of CSs suppressed PCMA without affecting PII. CONCLUSION: ACTH suppressed PCMA through CSs, and reduced PII directly. ACTH was hypothesized to eliminate IS through these dual modes of action. PMID- 10705936 TI - Risk of symptoms related to late effects of poliomyelitis. AB - OBJECTIVE: To compare the risk of developing symptoms related to late effects of poliomyelitis between polio patients and persons of similar age and sex without history of poliomyelitis. MATERIAL AND METHODS: The study comprised information on 148 patients with prior poliomyelitis and 115 persons with no history of poliomyelitis. Information was obtained by questionnaire and analyzed by multiple logistic regression method. RESULTS: The risk of experiencing two or more symptoms was significantly higher among the polio patients than among the persons without history of poliomyelitis. The elevation in risk was less pronounced in the nonparalytic group (OR = 2.35; 95% CI = 0.92-5.97) than the group with permanent muscular weakness (OR = 8.84; 95% CI =4.32-18.09). CONCLUSIONS: Although symptoms defined in the PPS are unspecific and may occur in the general population, the risk for developing such symptoms are higher among the polio victims. The difference in risk among nonparalytic and paralytic patients may depend on the extent of motor neuron damage in the acute stage. PMID- 10705937 TI - Impaired cytokine production by peripheral blood mononuclear cells and monocytes/macrophages in Parkinson's disease. AB - OBJECTIVE: Although the pathogenesis of Parkinson's disease (PD) is still unknown, several reports suggest the presence of immunological abnormalities in the patients with PD such as impaired T cell responses or cytokine production by the peripheral immune system. MATERIAL AND METHOD: In this study, we examined cytokine production by peripheral blood mononuclear cells (PBMC) and monocyte/macrophages (PBM) in the patients with idiopathic PD, using age-related healthy donors as a normal control and cerebrovascular diseases (CVD) as a disease control. RESULTS: Production of TNF-alpha, IL-1alpha, IL-1beta and IL-6 by PBMC and TNF-alpha by PBM were significantly lower in the patients with PD as compared to the control groups. IFN-gamma production by LPS-stimulated PBMC in the patients with PD was also significantly lower than that in control groups. Cytokine production by PBMC from the patients with CVD who had a similar disability as the patient group was not significantly different from those in normal controls. Thus, impaired production of inflammatory cytokines may not be due to the mental and physical stress caused by their disability. CONCLUSION: In the patients with PD, a significant negative correlation was noted in 1alpha 1beta, IL-1beta and IL-6 levels produced by LPS-stimulated PBMC and Hoehn Yahr disability score of the patients, suggesting that the impaired cytokine production may progress with disease progression. These abnormalities in cytokine production may not be primary but may affect the prognosis of PD. PMID- 10705938 TI - Dopamine D2 receptor imaging in Gilles de la Tourette syndrome. AB - OBJECTIVES: To examine postsynaptic dopamine D2 receptors in Tourette syndrome (TS). MATERIAL AND METHODS: Seventeen patients and a control group were investigated using single photon emission computed tomography (SPECT) and iodobenzamide (123I-IBZM). RESULTS: In neuroleptic treated patients (n = 7) 123I IBZM-binding was significantly reduced compared to both normal controls (P < 0.0001) and unmedicated patients (P < 0.001). In unmedicated patients (n = 10) mean binding ratio did not differ from that of control group. Patients in advanced stages of the disease, however, revealed significantly reduced relative striatal binding compared to patients in the early stages (P<0.005) and normal controls (P<0.0001). CONCLUSION: The results lend further support to the hypothesis that the dopamine receptor is involved in TS pathology. During the natural course of the disease tics often improve in early adulthood. It is suggested that this spontaneous recovery from tics may be associated with reduced receptor binding capacity. PMID- 10705939 TI - SPECT in Alzheimer's disease: features associated with bilateral parietotemporal hypoperfusion. AB - OBJECTIVE: To investigate why bilateral parietotemporal hypoperfusion, the typical SPECT pattern of Alzheimer's disease (AD), occurs in some but not in all patients with probable AD. METHODS: We reviewed the SPECT scans of 220 patients presenting with cognitive impairment. Among them, 104 patients fulfilled NINCDS ADRDA criteria for probable AD, 48 (32 women) with early onset (before age of 65) and 56 (40 women) with late onset of the symptoms. Dementia severity was assessed by the Mini-Mental State Examination. The SPECT scans were classified by visual inspection blind to clinical diagnoses. RESULTS: Bilateral parietotemporal hypoperfusion was more frequent in patients with severe AD, in those with early onset of the symptoms, and in men. Duration of symptoms, type of gamma-camera or radiopharmaceutical agent used were not associated with this SPECT pattern. CONCLUSION: These findings may be useful in the clinical setting and point to heterogeneity of AD according to age at onset. PMID- 10705940 TI - The clinical and genetic characteristics of spinocerebellar ataxia type 7 (SCA 7) in three Black South African families. AB - OBJECTIVES: Spinocerebellar ataxia type 7 is a rare autosomal dominant neurodegenerative disorder characterized by progressive cerebellar and retinal degeneration, described in various population groups in the literature. This is the first description from South Africa. The objective was to document the clinical and genetic characteristics of our patients and to determine concordance with other described cases. PATIENTS AND METHODS: The index cases were identified clinically on the basis of the typically described features of progressive ataxia with visual failure due to progressive cerebellar and retinal/macular degeneration. Associated neurological disturbances were documented. Where possible, and available, family members were assessed and pedigrees were delineated. Molecular tests for SCA expansions were determined in the index cases. RESULTS: Three pedigrees of SCA 7 were identified. The patients were all Black South Africans. The genetic and clinical characteristics are typical for SCA 7. CONCLUSION: SCA 7 is a rare distinct neurodegenerative disorder characterized by trinucleotide expansion. PMID- 10705941 TI - Screening for HIV-associated distal-symmetric polyneuropathy in CDC classification stages 1, 2, and 3. AB - OBJECTIVES: A total of 670 patients were screened for distal symmetric HIV associated polyneuropathy during CDC stages 1-3 and its correlation to immunological deterioration. MATERIAL AND METHODS: Clinical examinations of 670 patients admitted to the neurological outpatient clinic at the Department of Neurology, University of Munster. Neurophysiological investigations were performed on the sural and peroneal nerve for detection of axonal and myelin lesion. RESULTS: Clinical examination proved progressive clinical signs and symptoms indicating distal symmetric polyneuropathy from CDC 1 (32%) to CDC 3 (55%). At least one neurophysiological result was impaired in CDC 1 in 25% and in CDC 3 in 45%. Significant correlation between neurophysiological changes and CDC4(+)-cells and beta-microglobuline were detected for stage CDC 3 C. CONCLUSION: Results show stage related prevalence of distal symmetric polyneuropathy already in early stages. In late stages of HIV-infection prevalence of distal symmetric polyneuropathy seems to be directly correlated to immunodeficiency syndrome. The pathogenesis of distal symmetric polyneuropathy during HIV-infection is up to now incompletely understood, but results indicate a clear dependency between progressive immunological dysfunction and neuropathy. High active antiretroviral therapy in patients suffering from distal symmetric polyneuropathy is a main topic of future studies. PMID- 10705942 TI - Preoperative B-mode ultrasound plaque appearance compared with carotid endarterectomy specimen histology. AB - In carotid artery stenosis both the degree of the lesion and its plaque morphology are thought to be associated with the carrier's thromboembolic risk. In this study we evaluated the diagnostic preciseness of non-invasively B-mode ultrasound in predicting the histopathological plaque structure. We examined 44 patients with > 50% ICA stenosis by B-mode within 6 weeks prior to carotid endarterectomy. At the affected bifurcations, up to 10 different regions of interest (ROI) per artery were investigated. Plaque appearance was classified according to 6 subtypes considering different ultrasonic plaque features. Postoperatively, plaque specimens were examined histopathologically for their relative content of calcification, fibrous tissue and different soft tissue. B mode ultrasound was compared with histopathological features in ROI. A total of 265 regions of interest were evaluated. In mainly echolucent types of plaques, atheromatous debris was most frequently seen, whereas fibrosis was rare. Homogeneous echolucent plaques showed a high proportion of cholesterol and/or recent haemorrhage. Thrombosis at the plaque surface was often seen in "completely echolucent" plaque type (each P<0.001). Carotid B-mode ultrasonography is able to predict the histopathological components and the texture of carotid plaques. PMID- 10705943 TI - Visual neglect as a predictor of functional outcome one year after stroke. AB - OBJECTIVE: The aim was to study the role of visual neglect in acute right hemisphere brain infarct as a predictor of poor functional outcome during the first year after stroke. In particular, we were interested in the additional value of neglect measures besides hemiparesis, hemianopia, cognitive deficits and age. PATIENTS AND METHODS: A consecutive series of 57 patients with a neuroradiologically verified right hemisphere infarct was examined within 10 days of the stroke. Fifty patients were followed up for 1 year. Neglect was measured with the Conventional and the Behavioural subtests of the Behavioural Inattention Test (BITC and BITB, respectively). The predictors were determined at the 10-day examination. Functional outcome was assessed 3, 6 and 12 months after the onset with the Frenchay Activities Index. RESULTS: Neglect in BITB was the best single predictor, which together with high age formed the best combination of predictors for poor functional outcome at each follow-up. Hemiparesis was also included in this prediction model at the 3-month follow-up, but hemianopia, BITC, or visuoconstructional and memory deficits showed no additional predictive value. However, neglect usually recovered soon. When neurological and cognitive deficits were assessed at the same time as the outcome, hemiparesis rather than neglect was the strongest correlate of poor outcome. CONCLUSION: Neglect in acute stroke is an important predictor of poor functional recovery. Residual neglect, which could be compensated in the follow-up tests, may nevertheless restrict patients' real-life activities and hobbies. PMID- 10705944 TI - Auditory P300 event related potential in minor ischemic stroke. AB - OBJECTIVE: Various degenerative cerebral diseases and even depression may cause abnormalities of the cognitive event related potentials (ERPs). We conducted the present study to elucidate the effects of ischemic stroke on the P300 ERP component. MATERIAL AND METHODS: We recorded the P300 wave using an auditory oddball paradigm in 38 consecutive brain infarct patients with mild neurological deficits at 3 and 12 months post-stroke, and in 29 healthy control subjects. RESULTS: Brain infarction slightly prolonged the P300 latency, and the observed delay was related to the presence and degree of post-stroke depression evaluated with the Zung Depression Scale and the DSM-III criteria. Infarction did not affect the P300 amplitude or its distribution on the scalp. The results of the patients with hemispheric brain infarction and those of the patients with brainstem infarction were similar, and also the values of the patients with the left- and right-sided lesions. The normal physiological correlation between subject age and the P300 latency was absent at 3 months post-stroke, but was present at 12 months post-stroke. CONCLUSION: Brain infarction delays the P300 ERP and temporarily distorts its age-related physiology. The increase of the P300 latency seems to be associated with the post-stroke depression. PMID- 10705945 TI - Serum levels of coenzyme Q10 in patients with multiple sclerosis. AB - To elucidate whether serum coenzyme Q10 levels are related with the risk for multiple sclerosis (MS) or are a marker for the activity of the disease, we compared serum levels of coenzyme Q10 and the coenzyme Q10/cholesterol ratio, in 31 patients with MS (during exacerbations) and 19 matched controls using a high performance liquid chromatography technique. The mean serum coenzyme Q10 levels and the coenzyme Q10/cholesterol ratio did not differ significantly between the 2 study groups. The values did not correlate with age, age at onset, and duration of the disease. These results suggest that serum coenzyme Q10 concentrations are unrelated with the risk for MS and are not a useful marker of activity of the disease. PMID- 10705946 TI - Giant negative T waves in Guillain-Barre syndrome. AB - A Guillain-Barre syndrome patient showed giant negative T waves on electrocardiography at the height of the disease, with large left ventricular hypokinesis on echocardiography and extensive defects on 123I-meta iodobenzylguanidine myocardial scintigraphy. Gamma-globulin improved the neurological symptoms, and the above abnormalities resolved. We speculate that cardiac sympathetic nerve endings were transiently damaged, with consequent myocardial injury, due to norepinephrine toxicity. PMID- 10705948 TI - Plasma alpha-tocopherol level in diabetes mellitus. PMID- 10705947 TI - Localization of carotenoids in different eye tissues. PMID- 10705950 TI - Turnover of tocopherols in Atlantic salmon (Salmo salar, L.). PMID- 10705949 TI - Sesamin and alpha-tocopherol synergistically suppress lipid-peroxide in rats fed a high docosahexaenoic acid diet. AB - Docosahexaenoic acid (DHA) is an essential nutrient for human health, but has extremely high oxidative susceptibility. We examined the suppressing effect of sesamin, a sesame seed lignan, on lipidperoxides in rats fed a low alpha tocopherol and high DHA containing diet. Groups of rats were fed four experimental diets: low alpha-tocopherol (10 mg/kg diet) control diet, low alpha tocopherol + 0.2% sesamin diet, low alpha-tocopherol + 0.5% DHA diet and low alpha-tocopherol + 0.5% DHA + 0.2% sesamin diet. TBARS concentrations in plasma and liver were significantly increased by DHA, but were completely suppressed by sesamin. Alpha-tocopherol concentrations in plasma and liver decreased by addition of DHA, but with sesamin recovered to the control level. The addition of DHA into the diets caused remarkable increases of DHA concentrations in plasma and liver lipids. Sesamin caused a significant increase of DHA concentrations in the triacylglycerol of plasma. PMID- 10705951 TI - Determination of vitamin E in eggs and during the larval development of the sea bass, Dicentrarchus labrax by high performance liquid chromatography. PMID- 10705952 TI - Studies on the capability of vitamin C to induce SOS response and cytotoxicity in the absence and in the presence of vitamin E. PMID- 10705953 TI - Changes in erythrocyte antioxidant enzymes during prolonged submaximal exercise. PMID- 10705954 TI - Decreased serum ubiquinol-10 levels in healthy subjects during exercise at maximal oxygen uptake. PMID- 10705956 TI - Cloning and expression of rat glucocorticoid-receptor DNA-binding domain. AB - The inability to obtain sufficient quantities of purified glucocorticoid receptor (GR) causes difficulties in studying in vitro the interactions of GR with other proteins which contribute to the transcriptional activation of chromatin. Thus, we overexpressed the DNA-binding domains (DBD) of GR in E. coli using glutathione S-transferase (GST)-fusion protein expression vector. The DBD-GST protein (about 57 kDa), recovered in the soluble fraction, was purified by glutathione-agarose column. However, during the chromatography, more than 90% of the DBD-GST protein was cleaved into three species with molecular sizes of about 40, 35 and 30 kDa. The 40 kDa and 35 kDa proteins were immunostained with the anti-GR antibody, and the 30 kDa protein was stained with anti-GST antibody. Among these proteins, only the 57 kDa DBD-GST protein bound to the GRE-agarose. These data indicate that the properly folded 57 kDa protein has a DNA binding function and is resistant to intrinsic cleavage of the protein. PMID- 10705957 TI - Mitochondrial oxidative stress: a self-propagating process with implications for signaling cascades. PMID- 10705955 TI - An investigation into a possible relationship between vitamin D, parathyroid hormone, calcium and magnesium in a normally pigmented and an albino rural black population in the Northern Province of South Africa. AB - Vitamin D levels are important in the management of patients with various disorders of calcium metabolism associated with rickets, osteomalacia, osteodystrophy osteoporosis and hypoparathyroidism. 82 albinos and 58 normally pigmented children resident at the Siloe School for the Visually Impaired were sampled. Blood samples of fasting subjects were collected over a two-day period and analyzed for vitamin D, parathyroid hormone, plasma calcium and both plasma and red blood cell magnesium measurements. The height and weight of each subject was also recorded. The results are discussed in relation to the different skin pigmented groups, for specific age groups, sex and visual status. Statistical outliers were excluded from the results. It appears that the Albino group has significantly (p = 0.06) higher vitamin D levels against the background of a similar dietary intake and similar exposure to sunlight/day length. Thus black children/subjects require a significantly higher intake of vitamin D to attain the same level as their Albino counterparts. In spite of significantly higher vit D levels, the other homeostatic control mechanisms were not altered (i.e., PTH levels are similar in both groups). This study supports the postulate that a dark complexion predisposes to sub-optimal vit D status. PMID- 10705959 TI - Using homocysteine and related metabolites to diagnose vitamin deficiency states. PMID- 10705958 TI - Homocysteine and other thiol compounds in ageing. PMID- 10705960 TI - Pantothenic acid in maintaining thiol and immune homeostasis. PMID- 10705961 TI - Fatty acid cyclooxygenase induction accompanied by prostaglandin D synthesis in a human megakaryoblastic cell line CMK differentiated by phorbol ester. PMID- 10705962 TI - Redox regulation by thioredoxin and thioredoxin reductase. PMID- 10705963 TI - Tryparedoxin II from Crithidia fasciculata. PMID- 10705965 TI - Tryparedoxin and tryparedoxin peroxidase. PMID- 10705964 TI - Cloning and expression of tryparedoxin I from Crithidia fasciculata. PMID- 10705966 TI - Crystallisation of tryparedoxin I from Crithidia fasciculata. PMID- 10705967 TI - Biosynthesis of selenoproteins--an overview. PMID- 10705968 TI - The penultimate selenocysteine residue at the C-terminus of mammalian thioredoxin reductase plays an obligatory role in the NADPH-disulfide oxidoreductase catalytic mechanism. PMID- 10705969 TI - The influence of the cytokines Il-1beta and INFgamma on the expression of selenoproteins in the human hepatocarcinoma cell line HepG2. AB - Metabolic labeling of HepG2 cells with 75selenite indicated the expression of more than four selenoproteins in the 80 to 45 kDa range and more than four selenoproteins in the 24 to 14 kDa range. Although there were no significant changes in the total protein content, determined by the method of BioRad, the densitometric analysis of the autoradiogramms showed a reduced expression of all labeled selenoproteins in the cell lysates of HepG2 after cytokine treatment for 24 hours. A stronger reduction was observed after treatment with Il-1beta than with IFNgamma. Moreover, the inhibitory effects were more significant for those selenoproteins in the higher molecular mass range. Our data on the inhibition of selenoprotein synthesis and on repression of SeP promoter activity show that the expression of selenoproteins, especially of SeP, is influenced by acute phase reaction and pro-inflammatory reactions. PMID- 10705970 TI - Evidence that multiple proteins from Salmonella typhimurium are required for the biosynthesis of 5-methylaminomethyl-2-selenouridine in tRNAs. PMID- 10705971 TI - The role of 4'-phosphopantetheine in t' biosynthesis of fatty acids, polyketides and peptides. AB - The peptide part of CoA, 4'-phosphopantetheine, has been identified as an essential cofactor in in the biosynthesis of fatty acids, polyketides, depsipeptides, peptides, and compounds derived from both carboxylic and amino acid precursors, like rapamycin. The cofactor is attached to a unique protein moiety, referred to as acyl carrier protein, aminoacyl carrier protein, or peptidyl carrier protein. These carrier proteins are either associated to enzyme complexes (type II) or integrated within multifunctional polypeptide chains (type I). The cofactor is added in a post-translational modification reaction by highly specific transferases, acting on CoA. The functions of carrier proteins in directed condensation reactions are: (1) the selection of substrates to be attached as thioesters, (2) the stabilization of intermediates, (3) the presentation of intermediates for modification by associated enzyme activities, (4) facilitation of the directed condensation reactions of two adjacent intermediates, and (5) assistance in the termination reaction(s) leading to product release. PMID- 10705972 TI - Effect of dietary vitamin B6 contents on antibody production. AB - When mice were placed on diets extreme deficient in vitamin B6, ovalbumin dependent antibody productions (IgE, IgG1, IgG2a) were significantly suppressed, and alanine aminotransferase activity in the liver was also significantly decreased. In the case of pyridoxine excess (6 mg% = about ten times standard amount) in a 70% casein diet, ovalbumin-dependent antibody productions were also considerably suppressed. These responses were weaker in a low casein (5%) or normal casein (20%) diet than in a 70% casein diet. The administration of high doses of pyridoxine (6 mg%) resulted in the suppression of hepatic cathepsin B activity. Therefore, we conclude that ovalbumin-dependent antibody productions (IgG1, IgE) were suppressed by pyridoxine excess diet (6 mg%), because hepatic cathepsin B activity was suppressed by the excess pyridoxine in diet. PMID- 10705973 TI - Activation of the coenzyme at the early step of the catalytic cycle of tryptophanase. AB - Tryptophanase is catalytically more competent in alkaline pH even though the coenzyme exists as an inactive aldamine structure in this pH region. The binding of a substrate analog, 3-indolepropionate to the enzyme shifts the equilibrium from the substituted aldamine to the ketoenamine form in the entire pH region studied. The resultant ketoenamine form is favorable for transaldimination with the substrate amino group, a prerequisite for subsequent catalysis. This implies that the binding of tryptophan in alkaline pH, where the enzyme shows maximum activity, converts the inactive aldamine form of the coenzyme to the active ketoenamine form, which is favorable for undergoing the next step of the catalytic process. PMID- 10705974 TI - Search for a microbial biotin transporter. PMID- 10705975 TI - Control mechanism of expression of tyrosine phenol-lyase, vitamin B6 dependent L DOPA producing enzyme. PMID- 10705976 TI - A reactivating factor for coenzyme B12-dependent diol dehydratase. PMID- 10705977 TI - Biokinetic parameters and metabolism of S-benzoylthiamine-O-monophosphate. PMID- 10705978 TI - Effect of high-dosed thiamine hydrochloride and S-benzoyl-thiamine-O monophosphate on thiamine-status after chronic ethanol administration. PMID- 10705979 TI - Thermodynamic properties of L-lactate oxidase reconstituted with modified flavins. PMID- 10705981 TI - Biosynthesis of thiamin under anaerobic conditions in Saccharomyces cerevisiae. PMID- 10705980 TI - Inhibition of cathepsin activities by vitamin B6 derivatives. AB - We found that pridoxal phosphate (PLP), a coenzyme form of vitamin B6, inhibited the activity of cathepsins B, K, S and L in vitro. Cathepsins activities in cultured splenocytes were suppressed by the addition of pridoxal (PL) or pridoxine (PI) in to the culture medium. A newly synthesized artificial vitamin B6 derivative, a pridoxal propionate derivative, CLIK-164, showed selective inhibition of cathepsin O/K. PMID- 10705982 TI - Purification and properties of pyridoxine oxidase from Aureobacterium luteolum and pyridoxal reductase from Schizosaccharomyces pombe. PMID- 10705983 TI - Transketolase and protein oxidation of senescence accelerated mouse. PMID- 10705984 TI - Serum iron and copper and their relations to hepatocellular carcinoma in porphyria cutanea tarda and hemochromatosis patients--case report. PMID- 10705985 TI - Abnormalities in serum copper and iron concentrations in porphyria cutanea tarda patients and their relationships with other parameters. PMID- 10705986 TI - Zinc deficiency and normal contents of magnesium and calcium in metabolic X syndrome patients as assessed by the analysis of hair element concentrations. PMID- 10705988 TI - The role of the molybdenum cofactor in humans. PMID- 10705987 TI - Analysis of calcium, magnesium, and zinc levels in hair of healthy students. Screening of calcium or magnesium deficiency hazard. PMID- 10705990 TI - Vanadate induces apoptosis in epidermal JB6 P+ cells via hydrogen peroxide mediated reactions. AB - Apoptosis is a physiological mechanism for the control of DNA integrity in mammalian cells. Vanadium induces both DNA damage and apoptosis. It is suggested that vanadium-induced apoptosis serves to eliminate DNA-damaged cells. This study is designed to clarify a role of reactive oxygen species in the mechanism of apoptosis induced by vanadium. We established apoptosis model with murine epidermal JB6 P+ cells in the response to vanadium stimulation. Apoptosis was detected by a cell death ELISA assay and morphological analysis. The result shows that apoptosis induced by vanadate is dose-dependent, reaching its saturation level at a concentration of 100 microM vanadate. Vanadyl (IV) can also induce apoptosis albeit with lesser potency. A role of reactive oxygen species was analyzed by multiple reagents including specific scavengers of different reactive oxygen species. The result shows that vanadate-induced apoptosis is enhanced by NADPH, superoxide dismutase and sodium formate, but was inhibited by catalase and deferoxamine. Cells exposed to vanadium consume more molecular oxygen and at the same time, produce more H2O2 as measured by the change in fluorescence of scopoletin in the presence of horseradish peroxidase. This change in oxygen consumption and H2O2 production is enhanced by NADPH. Taken together, these results show that vanadate induces apoptosis in epidermal cells and H2O2 induced by vanadate plays a major role in this process. PMID- 10705989 TI - Hypoxia-induced bFGF gene expression is mediated through the JNK signal transduction pathway. AB - Although the synthesis of angiogenic factors in hypoxic regions of solid tumors is recognized as one of the critical steps in tumor growth and metastasis, the signal transduction pathway involved in hypoxic induction of basic fibroblast growth factor (bFGF) gene expression is still obscure. In the study described here, we investigated the intracellular responses to hypoxia and the mechanisms triggering the initiation of angiogenic activity in drug-resistant human breast carcinoma MCF-7/ADR cells. Northern blots showed an increase in the level of c jun, c-fos, and bFGF mRNA during hypoxia. Gel mobility-shift analysis of nuclear extracts from hypoxia-exposed cells showed an increase in AP-1 binding activity. In addition, hypoxic treatment strongly activated c-Jun N-terminal kinase 1 (JNK1), leading to phosphorylation and activation of c-Jun. Expression of a dominant negative mutant of JNK1 suppressed hypoxia-induced JNK1 activation as well as bFGF gene expression. Taken together, hypoxia-induced bFGF gene expression is mediated through the stress-activated protein kinase (SAPK) signal transduction pathway. PMID- 10705991 TI - Alterations in sarcoplasmic reticulum function in female vastus lateralis with eccentric exercise. AB - This study examined the alterations in sarcoplasmic reticulum (SR) Ca2+ sequestration function in homogenates during eccentric exercise and recovery and following additional eccentric exercise, and correlated these alterations with changes in force output. Eight healthy, untrained females, aged 20-25 years, cycled for a total of 60 min on an eccentric cycle ergometer (30 min at 66+/-3% VO2 peak and 30 min at 76+/-3% VO2 peak, determined during concentric exercise). Biopsies (extracted from the vastus lateralis) were taken before and after the exercise as well as on days 2, 6 and prior to and following identical exercise on day 14. Ca2+-uptake (nmol/min/mg protein) was unaffected (p > 0.05) following the first session of eccentric exercise; however, by day 2 a depression in uptake (p < 0.05) was observed which persisted throughout the remainder of the experiment. Maximal Ca2+-ATPase activity (nmol/min/mg protein) was elevated (p < 0.05) immediately following the first exercise session, remained elevated through day 2 and returned to pre-exercise levels by day 6 of recovery and increased again by day 14. No changes in either Ca2+-ATPase activity or Ca2+-uptake were observed with exercise on day 14. Both eccentric sessions, performed on days 0 and 14, resulted in similar depressions in force (p < 0.05) immediately following exercise. By day 2 force had recovered to pre-exercise levels. The results demonstrate that a prolonged alteration in SR Ca2+-uptake occurs following eccentric work that is unaccompanied by parallel changes in either SR Ca2+-ATPase activity or mechanical performance. PMID- 10705992 TI - Dexamethasone makes the gastric mucosa susceptible to ulceration by inhibiting prostaglandin synthetase and peroxidase--two important gastroprotective enzymes. AB - The plausible mechanism by which dexamethasone makes the gastric mucosa susceptible to ulceration has been studied. As acid aggravates ulcer, the role of dexamethasone on acid secretion was first investigated. Dexamethasone stimulates both basal and drug (mercaptomethylimidazole)-induced gastric acid secretion by 100 and 50% respectively in male Wister rats 24 h after intramuscular administration at the dose of 1 mg/kg body wt. This stimulated acid secretion is 93% blocked by cimetidine indicating increased liberation of histamine in the process. Pretreatment of dexamethasone before 24 h produces ulcer in 30% of the pylorus- ligated rats and aggravates the ulcer index by 82% in both pylorus and esophagus ligated rats. The incidence of ulceration in the latter cases is also increased by 25%. As mucosal prostaglandin synthetase and peroxidase play an important role in gastroprotection through biosynthesis of prostaglandin and by scavenging endogenous H2O2 respectively, the effect of dexamethasone on the activities of these gastroprotective enzymes were studied. Prostaglandin synthetase and peroxidase activities of the mucosa are significantly inhibited by 87 and 83% respectively by 24-h pretreatment with dexamethasone. The results indicate that dexamethasone makes the mucosa prone to ulceration by inhibiting the activity of prostaglandin synthetase to block the gastroprotective action of prostaglandin and also by inhibiting the peroxidase, thereby elevating the endogenous H2O2 level to generate more reactive hydroxyl radical responsible for the mucosal damage. PMID- 10705993 TI - Regulation of the apolipoprotein B in heterozygous hypobetalipoproteinemic knock out mice expressing truncated apoB, B81. Low production and enhanced clearance of apoB cause low levels of apoB. AB - Low levels of cholesterol are protective against development of coronary artery disease. Heterozygous hypobetalipoproteinemic individuals expressing truncated apolipoprotein (apo)B as a result of mutation in the apob gene have low levels of cholesterol and apoB in their plasma. To study the molecular mechanism of low levels of apoB in these individuals, we employed a previously reported knock out mouse model generated by targeted modification of the apob gene. The heterozygous, apoB-100/B-81, mice express full length and truncated apoB, B-81, and have 20 and 35% lower levels of total cholesterol and apoB, respectively, when compared to WT (apoB-100/B-100) mice. The majority of the truncated apoB, B 81, fractionated in the VLDL- density range. The mechanism of low levels of apoB in B-100/B-81 mice was examined. Total hepatic apoB mRNA levels decreased by 15%, primarily due to lower levels of apoB-81 mRNA. Since apoB mRNA transcription rates were similar in B-100/B-100 and B-100/B-81 mice, low levels of mutant apoB 81 mRNA occurred by enhanced degradation of apoB mRNA transcript containing premature translational stop codon. ApoB synthesis measured on isolated hepatocytes decreased in B-100/B-81 mice by 35%, while apoB-48, apoE, and apoAI syntheses remained unchanged. Metabolic studies using whole animal showed a 32% decrease in triglyceride secretion rates, consistent with the apoB secretion rates. Inhibition of receptor-mediated clearance of apoB-81-containing particles resulted in greater relative accumulation of apoB-81 in plasma than apoB-100, suggesting enhanced clearance of apoB-81-containing particles. These results demonstrate that low levels of apoB in heterozygous hypobetalipoproteinemic mice occurs by low rates of apoB secretion, and increased clearance of truncated apoB. Similar mechanisms appear to contribute to low levels of apoB in hypobetalipoproteinemic humans. PMID- 10705994 TI - Peroxide sensitivity of endothelin responses in coronary artery smooth muscle: ET(A) vs. ET(B) pathways. AB - Endothelins (ETs) contract de-endothelialized rings from left descending coronary artery via ET(A) or ET(B) receptors. Here we test the hypothesis that the actions of EA(A) and ET(B) receptors are similar in their sensitivities to damage by hydrogen peroxide. In Ca2+-containing Krebs' solution, 100 nM of the ET(B) agonist IRL1620 produced contractions with significantly smaller force (17.6+/ 1.7 mN) than 50 nM of the ET(A) + ET(B) agonist ET-1 (73.2+/-4.6 mN) (p < 0.05). In Ca2+-free solutions, the contractions due to both agents were significantly smaller (p < 0.05). Pretreating the tissues with peroxide inhibited the contractions produced by either agent. The IC50 values for peroxide were significantly higher (p < 0.05) using ET-1 (1.0+/-0.3 mM in Ca2+, 1.4+/-0.1 mM in Ca2+-free) than using IRL 1620 (0.32+/-0.08 in Ca2+, 0.25+/-0.01 mM in Ca2+ free). Pretreating microsomes isolated from the artery smooth muscle with up to 10 mM peroxide did not significantly affect 125I-ET-1 binding to ET(A) or ET(B) receptors (p > 0.05). In comparing the peroxide induced inactivation of the various processes in this artery and based on literature, we conclude that the actions of ET(A) may also involve a peroxide resistant Ca2+-independent pathway(s). PMID- 10705995 TI - Bcl-2, survivin and variant CD44 v7-v10 are downregulated and p53 is upregulated in breast cancer cells by progesterone: inhibition of cell growth and induction of apoptosis. AB - Progesterone inhibits the proliferation of normal breast epithelial cells in vivo, as well as breast cancer cells in vitro. But the biologic mechanism of this inhibition remains to be determined. We explored the possibility that an antiproliferative activity of progesterone in breast cancer cell lines is due to its ability to induce apoptosis. Since p53, bcl-2 and survivin genetically control the apoptotic process, we investigated whether or not these genes could be involved in the progesterone-induced apoptosis. We found a maximal 90% inhibition of cell proliferation with T47-D breast cancer cells after exposure to 10 microM progesterone for 72 h. Control progesterone receptor negative MDA-231 cancer cells were unresponsive to 10 microM progesterone. The earliest sign of apoptosis is translocation of phosphatidylserine from the inner to the outer leaflet of the plasma membrane and can be monitored by the calcium-dependent binding of annexin V in conjunction with flow cytometry. After 24 h of exposure to 10 microM progesterone, cytofluorometric analysis of T47-D breast cancer cells indicated 43% were annexin V-positive and had undergone apoptosis and no cells showed signs of cellular necrosis (propidium iodide negative). After 72 h of exposure to 10 microM progesterone, 48% of the cells had undergone apoptosis and 40% were annexin V positive/propidium iodide positive indicating signs of necrosis. Control untreated cancer cells did not undergo apoptosis. Evidence proving apoptosis was also demonstrated by fragmentation of nuclear DNA into multiples of oligonucleosomal fragments. After 24 h of exposure of T47-D cells to either 1 or 10 microM progesterone, we observed a marked down-regulation of protooncogene bcl-2 protein and mRNA levels. mRNA levels of survivin and the metastatic variant CD44 v7-v10 were also downregulated. Progesterone increased p53 mRNA levels. These results demonstrate that progesterone at relative high physiological concentrations, but comparable to those seen in plasma during the third trimester of human pregnancy, exhibited a strong antiproliferative effect on breast cancer cells and induced apoptosis. PMID- 10705996 TI - Expression of PDGF-beta receptor, EGF receptor, and receptor adaptor protein Shc in rat osteoblasts during spaceflight. AB - A number of studies have indicated that microgravity induces osteopenia and modulates functions of mammalian cells. However, the molecular mechanisms underlying these effects of microgravity are still unknown. Rat osteoblasts were cultured for 4 and 5 days during Shuttle-Spacelab flight, and fixed by guanidine isothiocyanate solution on board after treatment with 1alpha, 25 (OH)2 vitamin D3. The mRNA levels for platelet-derived growth factor (PDGF)-beta receptor, epidermal growth factor (EGF) receptor, the growth factor receptor adaptor protein Shc, and c-fos were determined using the method of quantitative reverse transcription-polymerase chain reaction. The mRNA levels for EGF receptor were not altered by microgravity. However, the mRNA levels for PDGF-beta receptor, Shc, and c-fos were decreased to 62, 55 and 25% on the 4th day of flight, and 47, 40, and 43% on the 5th day, respectively, as compared to the corresponding ground controls. Expression of the growth factor receptor and the receptor adaptor protein was modulated in rat osteoblasts during spaceflight. Data suggest that signal transduction via growth factor receptors in rat osteoblasts is impaired by microgravity. Dysfunction of osteoblasts might be involved in spaceflight-induced osteopenia. PMID- 10705997 TI - Calcium- and ADP-magnesium-induced respiratory uncoupling in isolated cardiac mitochondria: influence of cyclosporin A. AB - This study was designed to determine the effect of calcium and ADP-Mg on the oxidative phosphorylation in isolated cardiac mitochondria. The influence of cyclosporin A was also evaluated. The mitochondria were extracted from rat ventricles. Their oxidative phosphorylations were determined in two respiration media with different free Ca2+ concentrations. Respiration was determined with palmitoylcarnitine and either ADP or ADP-Mg. With elevated free Ca2+ concentrations and ADP-Mg, the transition state III to state IV respiration did not occurred. The ADP:O ratio was reduced. The phenomenon was not observed in the other experimental conditions (low free Ca2+ concentration with either ADP- or ADP-Mg or elevated free Ca2+ concentration with ADP-). Uncoupling was allied with a constant AMP production, which maintained an elevated ADP level in the respiration medium and prevented the return to state IV respiration. It was also observed in a respiration medium devoid of free Ca2+ when the mitochondria were pre-loaded with Ca2+. Uncoupling was inhibited by cyclosporin A. Furthermore, the Krebs cycle intermediates released from 14C-palmitoylcarnitine oxidation revealed that succinate was increased by elevated free Ca2+ and ADP-Mg. Succinate is a FAD linked substrate with low respiration efficiency. Its accumulation could account for the decreased ADP:O ratio. The Ca2+- and ADP-Mg-induced uncoupling might be partly responsible for the mechanical abnormalities observed during low-flow ischemia. PMID- 10705998 TI - Protein kinase C isoforms in pituitary cells displaying differential sensitivity to phorbol ester. AB - Investigations with protein kinase C (PKC) isoform-specific antisera, revealed distinct profiles of PKC isoform content amongst pituitary tissues. Western analysis revealed the alpha, beta, delta, epsilon, zeta and theta isoforms of PKC are present in rat anterior and posterior pituitary tissue as well as in the GH3 somatomammotrophic cell line. AtT-20/D16-V corticotrophic and alphaT3-1 gonadotrophic murine cell lines contained no PKC-delta. The gamma or eta isoforms were undetected in any pituitary tissue. PKC activity measurements revealed Ca2+ independent PKCs in alphaT3-1 and GH3 cells which were more sensitive to activation by phorbol-dibutyrate (PDBu) than the corresponding PKC activity found in COS cells. However, Ca2+-dependent PKC activities were of similar sensitivity to PDBu in GH5, alphaT3-1 and COS cells, indicating that functional differences observed in PDBu-sensitivity in these cells may be due to differential activation of Ca2+-independent PKC isoforms. Moreover, substrate-specificity of these PKCs were also compared indicating that the amount of Ca2+-dependency of the observed PKC activity from the same pituitary tissue is dependent upon the substrate utilized by the PKC isotypes present. These findings explain differential sensitivities of PKC-mediated actions that have previously been observed in a range of pituitary cells. PMID- 10705999 TI - Antioxidant activity of the flaxseed lignan secoisolariciresinol diglycoside and its mammalian lignan metabolites enterodiol and enterolactone. AB - The antioxidant activities of the flaxseed lignan secoisolariciresinol diglycoside (SDG) and its mammalian lignan metabolites, enterodiol (ED) and enterolactone (EL), were evaluated in both lipid and aqueous in vitro model systems. All three lignans significantly (p < or = 0.05) inhibited the linoleic acid peroxidation at both 10 and 100 microM over a 24-48 h of incubation at 40 degrees C. In a deoxyribose assay, which evaluates the non site-specific and site specific Fenton reactant-induced *OH scavenging activity, SDG demonstrated the weakest activity compared to ED and EL at both 10 and 100 microM; the greatest *OH scavenging for ED and EL was observed at 100 microM in both assays. The incubation of pBR322 plasmid DNA with Fenton reagents together with SDG, ED or EL showed that the inhibition of DNA scissions was concentration dependent. The greatest non site-specific activity of lignans was at 100 microM, thus, confirming the results of the deoxyribose test. In contrast, the protective effect of SDG and EL in the site-specific assay was lost and that of ED was minimal. Therefore, the results indicate a structure-activity difference among the three lignans with respect to specific antioxidant efficacy. All three lignans did not exhibit reducing activity compared to ascorbic acid, therefore, did not possess indirect prooxidant activity related to potential changes in redox state of transition metals. The efficacy of SDG and particularly the mammalian lignans ED and EL to act as antioxidants in lipid and aqueous in vitro model systems, at relatively low concentrations (i.e. 100 microM), potentially achievable in vivo, is an evidence of a potential anticarcinogenic mechanism of flaxseed lignan SDG and its mammalian metabolites ED and EL. PMID- 10706000 TI - Dexamethasone-induced decrease in HMG-CoA reductase and protein-farnesyl transferase activities does not impair ras processing in AR 4-2J cells. AB - Rat pancreatic acinar cells AR 4-2J respond to dexamethasone by differentiation and a decreased proliferation rate. Protein labelling by [3H]-mevalonolactone, used as a precursor of farnesyl and geranylgeranyl isoprenoid groups, was increased in the presence of dexamethasone. In these same conditions, dexamethasone decreased HMG-CoA reductase activity, leading to a diminished isotopic dilution of the mevalonate precursor. As ras proteins, known to be involved in the regulation of proliferation and differentiation, need to be farnesylated for full biological function, we also measured the level of farnesyl transferase activity and found a dose-dependent decrease in dexamethasone treated cells. Despite these negative effects of dexamethasone on mevalonate pathway, there was no appearance of non-isoprenylated forms of ras, indicating that the level of isoprenoid precursors and farnesyl transferase activity were not limiting in this model. PMID- 10706001 TI - Characterization of Saccharomyces cerevisiae strains expressing ira1 mutant alleles modeled after disease-causing mutations in NF1. AB - The 2818 amino acids of neurofibromin, the product of the human NF1 gene, include a 230 amino acid Ras-GAP related domain (GRD). Functions which may be associated with the rest of the protein remain unknown. However, many NF1 mutations in neurofibromatosis 1 patients are found downstream of the GRD, suggesting that the C-terminal region of the protein is also functionally important. Since the C terminal region of neurofibromin encompassing these mutations is homologous with the corresponding regions in the two Saccharomyces cerevisiae Ras-GAPs, Ira1p and Ira2p, we chose yeast as a model system for functional exploration of this region (Ira-C region). Three missense mutations that affect the Ira-C region of NF1 were used as a model for the mutagenesis of IRA1. The yeast phenotypes of heat shock sensitivity, iodine staining, sporulation efficiency, pseudohyphae formation, and GAP activity were scored. Even though none of the mutations directly affected the Ira1p-GRD, mutations at two of the three sites resulted in a decrease in the GAP activity present in ira1 cells. The third mutation appeared to disassociate the phenotypes of sporulation ability and GAP activity. This and other evidence suggest an effector function for Ira1p. PMID- 10706002 TI - Brain gamma-glutamyltranspeptidase: characteristics, development and thyroid hormone dependency of the enzyme in isolated microvessels and neuronal/glial cell plasma membranes. AB - The characteristics, cellular locus and regulation of the enzyme gamma glutamyltranspeptidase (gammaGT) in brain were examined. In rat brain homogenates, the activity of the enzyme exhibited tissue differences- kidney>>>brain==testis>>liver>>skeletal muscle=ventricular muscle and regional differences--brain stem>hippocampus=cerebellum>cerebral cortex, with no significant species/strain differences in the select group of mammals studied. Methods were developed for the isolation from brain of microvessels (MV) and plasma membranes from neuronal/glial cells (N/G PM) utilizing morphological indicators and marker analyses. GammaGT activity was >12 higher in MV than N/G PM; however the enzyme displayed: stability, heat-activation and inhibition with maleate to the same extent in both fractions. A comparative study indicated that in the N/G PM fraction, gammaGT activity was low in all animals studied; gammaGT activity in MV however, was barely detectable in amphibians and reptiles, very low in birds and very high in mammal -mirroring the phylogenetic development of a functional blood-brain barrier. In the rat, gammaGT in both MV and N/G PM displayed a pronounced postnatal increase in activity but the extent and the patterns were different--in all cases, that of the MV greatly exceeded that of the N/G PM and in the MV, the enzyme activity the exhibited the same pattern as the postnatal development of the blood-brain barrier. The induction of congenital hypothyroidism by propylthiouracil (PTU) had no effect on gammaGT in N/G PM but effected a one third reduction in the activity of gammaGT in MV. The normalization by thyroid hormone replacement indicated that MVgammaGT is under thyroid hormone control. The induction of hypothyroidism by PTU in the adult, however, was without effect on enzyme activity in either fraction. The implications of the thyroid hormone dependency of MVgammaGT in the neonatal period and the relationship of gammaGT to the function of the blood brain-barrier is discussed. PMID- 10706003 TI - Effects of bis(maltolato) oxovanadium (IV) on protein serine kinases in skeletal muscle of streptozotocin-diabetic rats. AB - The in vivo effects of bis(maltolato)oxovanadium (IV) (BMOV) on the activity of protein serine kinases in skeletal muscle of STZ-diabetic Wistar rats were studied. BMOV was administered to STZ-diabetic rats at a concentration of 0.75 mg/ml for 8 weeks. Chronic BMOV treatment completely normalized plasma glucose levels in the diabetic animals after 8 weeks of treatment. Insulin-stimulated ERK 1 and ERK-2 activity was markedly increased in STZ-diabetic rats. Chronic BMOV treatment normalized the activity of ERK-2 in the diabetic treated animals, whereas the activity of ERK-1 was unaffected. In contrast to ERK-1 and ERK-2, the activity of the ribosomal S6 kinase p90rsk was decreased in STZ-diabetic rats. BMOV treatment restored the activity to normal levels. Basal p70 S6K activity was increased about 2.5-fold in the untreated diabetic group and no further increase in activity was observed after insulin stimulation. BMOV treatment did not correct the changes in p70 S6K activity in either the basal or insulin-stimulated states. In conclusion (i) the activity of ERK-1, ERK-2 and p90rsk were altered in skeletal muscle of STZ-diabetic rats; (ii) the glucoregulatory effects of BMOV were accompanied by concurrent improvement in the activities of ERK-2 and p90rsk; and (iii) there appears to be a dissociation between the activation of ERK-2 and p90rsk, suggesting that the regulation of p90rsk may be much more complex in vivo. PMID- 10706004 TI - Changes of sodium and ATP affinities of the cardiac (Na,K)-ATPase during and after nitric oxide deficient hypertension. AB - In the cardiovascular system, NO is involved in the regulation of a variety of functions. Inhibition of NO synthesis induces sustained hypertension. In several models of hypertension, elevation of intracellular sodium level was documented in cardiac tissue. To assess the molecular basis of disturbances in transmembraneous transport of Na+, we studied the response of cardiac (Na,K)-ATPase to NO deficient hypertension induced in rats by NO-synthase inhibition with 40 mg/kg/day N(G)-nitro-L-arginine methyl ester (L-NAME) for 4 four weeks. After 4 week administration of L-NAME, the systolic blood pressure (SBP) increased by 36%. Two weeks after terminating the treatment, the SBP recovered to control value. When activating the (Na,K)-ATPase with its substrate ATP, no changes in Km and Vmax values were observed in NO-deficient rats. During activation with Na+, the Vmax remained unchanged, however the K(Na) increased by 50%, indicating a profound decrease in the affinity of the Na+-binding site in NO-deficient rats. After recovery from hypertension, the activity of (Na,K)-ATPase increased, due to higher affinity of the ATP-binding site, as revealed from the lowered Km value for ATP. The K(Na) value for Na+ returned to control value. Inhibition of NO synthase induced a reversible hypertension accompanied by depressed Na+-extrusion from cardiac cells as a consequence of deteriorated Na+-binding properties of the (Na,K)-ATPase. After recovery of blood pressure to control values, the extrusion of Na+ from cardiac cells was normalized, as revealed by restoration of the (Na,K)-ATPase activity. PMID- 10706006 TI - Balloon catheter vascular injury of the alloxan-induced diabetic rabbit: the role of insulin-like growth factor-1. AB - Neointimal thickening following catheter injury is characterized, in part, by growth factor-induced vascular smooth muscle cell (VSMC) proliferation. It was hypothesized that a reduction in serum insulin-like growth factor-1 (IGF-1), characteristic of chemically-induced diabetes, would result in decreased VSMC proliferation and attenuate neointimal thickening. It was found that alloxan treated New Zealand White rabbits exhibit varying degrees of glycemia. Rabbits classified as diabetic (glucose > or = 400 mg/dL) had significantly decreased serum concentration of IGF-1 (87.4+/-14 nmol/L vs. 170+/-14 nmol/L) and significantly decreased intimal/medial (I/M) ratios 2, 4, and 8 weeks after aortic injury compared to euglycemic rabbits (13.7+/-2, 21.1+/-2, 32.4+/-3 in euglycemics and 6.6+/-1, 14+/-2, 19+/-5 in diabetics, respectively). The I/M for high hyperglycemic animals (glucose 286-399 mg/dL) was comparable to diabetic animals yet their serum IGF-1 levels were normal rather than depressed. Vascular IGF-1 content similarly increased upon injury in both diabetic and euglycemic animals. In diabetic animals, proliferating cell nuclear antigen (PCNA) immunostaining was present by day 1 peaked by day 5 and returned to control by day 14. In euglycemic animals, staining by day 1 continued to increase through day 14. A similar increase in mitogen-activated protein kinase (MAPK) activity occurred from day 1 through day 5 in both diabetic and euglycemic animals. This is the first demonstration of an association between MAPK activity and VSMC proliferation following vascular injury in diabetic animals as previously reported in euglycemic animals. In conclusion, this study provides evidence against a direct effect of IGF-1 in the reduction in neointimal thickening, VSMC proliferation, and MAPK activity upon catheter injury in chemically-induced diabetic rabbits. PMID- 10706007 TI - Psychiatry and its rightful position in medicine. PMID- 10706005 TI - Molecular characterisation of a NADH ubiquinone oxidoreductase subunit 5 from Schistosoma mansoni and inhibition of mitochondrial respiratory chain function by testosterone. AB - Complementary DNA, encoding the mitochondrial enzyme NADH-ubiquinone oxidoreductase subunit 5 (SmND5) of the human parasite Schistosoma mansoni was isolated by screening a S. mansoni cDNA library with a human androgen receptor (hAR) cDNA probe. The complete nucleotide and deduced aminoacid sequences of SmND5 were determined. Southern blot analysis revealed the occurrence of a single copy gene for SmND5 and by means of RT-PCR, it was shown that sex- and stage specific expression of SmND5 occurred. In order to establish a functional relationship between the mitochondrial enzyme and the androgen receptor, the effects of testosterone were compared to those of classical respiratory chain inhibitors, using adult schistosome and beef heart submitochondrial particles. Physiological concentrations of testosterone were able to inhibit the maintenance of proton gradient across the mitochondrial membranes, as well as ATP synthesis. The steroid was found to be cytotoxic to the larvae, but not to adult schistosomes. A model is proposed to explain the observed in vivo testosterone related differences in worm burdens, in experimental chronic infections. PMID- 10706008 TI - Mental health economic studies from developing countries reviewed in the context of those from developed countries. AB - OBJECTIVE: Mental health economic studies from developing countries were reviewed in the context of such studies from developed countries. METHOD: Mental health economic studies were ascertained through a systematic Medline search, chasing references at the end of papers acquired from the initial medline search and details of studies furnished by members of the WHO collaborating centre. RESULTS: Only a small number of mental health economic studies from developing countries were identified. They were mainly cost-of-illness and cost-effectiveness studies. CONCLUSION: Psychiatric disorders impose a significant burden in developing countries. It is not always possible to extrapolate findings from developed countries to developing countries. Potential avenues for future research and development are discussed. PMID- 10706009 TI - Psychopathology in treated Wilson's disease determined by means of CPRS expert and self-ratings. AB - OBJECTIVE: To examine the occurrence and severity of psychopathological symptoms in patients with treated Wilson's disease (WD) and to evaluate the clinical utility of a self-assessment. METHOD: Twenty-six consecutive patients with confirmed WD were investigated using the Comprehensive Psychopathological Rating Scale (CPRS) and the CPRS Self-rating Scale. RESULTS: The total CPRS scores ranged from 2.5 to 59.0 (mean 29.4 +/- 15.5). Most common symptoms were: autonomic disturbances, observed muscular tension, fatiguability, reduced sexual interest, lack of appropriate emotion, concentration difficulties, reduced sleep, aches and pains, hostile feelings, apparent sadness and failing memory. Agreement between interview-based ratings and self-ratings was low. CONCLUSION: The patients with treated WD have prominent psychopathology in the same range as in patients with moderate to severe depressive disorders. A specific symptom profile has been identified. If confirmed, the identification of the typical symptom profile might be of great importance. The patients with WD tend to underestimate the presence of psychopathological symptoms. PMID- 10706010 TI - Co-occurrence of Axis I and Axis II disorders in substance abusers. AB - OBJECTIVE: This study examined the co-occurrence of anxiety/mood and personality disorders (PDs) in substance abusers, the impact of anxiety/ mood disorders on the symptom profiles of PDs, and the impact of anxiety/mood disorders and PDs on pre-treatment status. METHOD: Current anxiety/mood disorders and PDs and pre treatment status were assessed using semi-structured interviews in 370 treated substance abusers. RESULTS: Anxiety/mood disorders and PDs frequently co occurred, with the overall pattern of associations being non-specific. The strongest associations were of social phobia with avoidant and schizotypal PD, and of major depression with borderline PD. However, symptom profiles of PDs were not associated with anxiety/mood disorders. Finally, anxiety/mood disorders and PDs were both independently and differentially associated with poor pre-treatment characteristics. CONCLUSION: The findings suggest the clinical importance of obtaining both Axis I and Axis II diagnoses in treated substance abusers, and highlight the distinctiveness of the Axis I and Axis II disorders. PMID- 10706011 TI - Predicting medication adherence in severe mental disorders. AB - OBJECTIVE: This study explored the utility of the health belief model (HBM) in explaining medication adherence in subjects with severe and disabling mental disorders. METHOD: Six well-established measuring instruments, with confirmed reliability and validity, were used to assess each component of the HBM and medication adherence in 39 hospital-treated subjects with affective disorders (n = 27) or schizophrenia (n = 12). RESULTS: Highly adherent and partially adherent subjects differed significantly in their perception of illness severity, their beliefs about themselves and their control over the disorder, and their concerns about further hospitalization. Two components of the HBM (perceived severity of illness and perceived benefits of treatment) explained 43% of the variance in adherence behaviour. CONCLUSION: Although the study has a number of methodological limitations, the results suggest that clinical assessment of components of the HBM may improve the detection of patients at risk of medication non-adherence. PMID- 10706012 TI - Potential for misdiagnosis among Turkish migrants with psychotic disorders: a clinical controlled study in Germany. AB - OBJECTIVE: The elevated rate of schizophrenia among migrants has been explained in part by possible misdiagnosis. In this study an attempt is made to quantify the extent of potential misdiagnosis among migrants in comparison to non migrants. METHOD: One hundred patients of Turkish origin (Tr-Pat) and a control group of 50 patients of German origin (G-Pat), all with a paranoid-hallucinatory syndrome upon admission, were examined by an interviewer of Turkish origin (Tr Int), an interviewer of German origin (G-Int) and the clinician. The diagnostic evaluation was then compared. RESULTS: Nineteen per cent of Tr-Pat and 4% of G Pat showed diagnostic disagreement between the three raters, while in 8% of Tr Pat and 0% of G-Pat the two research diagnoses disagreed. In Tr-Pat with 'bad' German knowledge showed tendentially more (29%) diagnostic disagreement than Tr Pat with 'good' German knowledge (17%). CONCLUSION: The rate of potential misdiagnosis is higher among migrants, yet not strongly correlated to poor second language proficiency. PMID- 10706013 TI - Extrapyramidal symptoms and oestrogen. AB - OBJECTIVE: The present study aimed to investigate neuroleptic side-effect severity in women with psychosis, and to investigate their putative association with variations in sex steroids over the menstrual cycle. Based on the oestrogen hypothesis, which postulates a synergistic relationship between oestrogen and neuroleptics, it was hypothesized that oestrogen would exacerbate extrapyramidal symptoms. METHOD: Twenty-five psychotic women were assessed using the ESRS and blood hormone analysis. Testing was conducted twice, 2 weeks apart, in a randomized cross-over design. RESULTS: Contrary to expectation the results indicated that high levels of oestrogen reduce hyperkinetic symptoms in women with psychosis, and this effect appears to be further potentiated when both oestrogen and progesterone are high. CONCLUSION: On the basis of these findings, and receptor studies in animals, it was concluded that oestrogen has different effects on dopamine dynamics in the mesolimbic and mesostriatal pathways. PMID- 10706014 TI - Factors in the onset of schizophrenia: a comparison between London and Trinidad samples. AB - OBJECTIVE: Sociodemographic factors play an important role in the genesis of mental disorders. High rates of unemployment and other social factors have been reported previously among African-Caribbeans with schizophrenia in London. The aim of the present study was to compare these factors in Trinidad with London African-Caribbeans. METHOD: Using internationally defined criteria, patients with first-onset schizophrenia were recruited in both countries, and information on the onset of symptoms, help-seeking, pathways into care, premorbid personality and educational and employment status were collected. These two samples are compared on a number of these factors. A total of 56 cases of first onset of psychosis coming into contact with psychiatric services in Trinidad were studied. Of these, 46 cases were diagnosed as having schizophrenia using the CATEGO program. Over a period of 2 years, 38 African-Caribbean patients with schizophrenia were recruited in London. RESULTS: African-Caribbean patients with schizophrenia in London were more likely to be admitted for perceived threat to others and to have shown loss of interest and serious neglect and to have assaulted others. A lower proportion were admitted via a psychiatrist and a higher proportion by the police. The unemployment rate among the London sample of African-Caribbeans was much higher than that in the general population, whereas this was not the case for the Trinidad patients. CONCLUSION: These findings are discussed in the context of culture and aetiology of schizophrenia, and suggestions with regard to future research are made. PMID- 10706015 TI - Are neurological abnormalities in schizophrenic patients and their siblings the result of perinatal trauma? AB - OBJECTIVE: Previous findings of increased neurological abnormalities in schizophrenic patients and their non-psychotic siblings raise the question of possible causes. The purpose of the present study was to examine the role of perinatal trauma in the aetiology of neurological abnormality. METHOD: Obstetric information obtained from hospital records for 55 schizophrenic patients and 19 non-psychotic siblings was scored blindly and separately from a neurological assessment of hard and soft signs. RESULTS: Obstetric complications (OCs) were significantly increased in patients but not in siblings compared to their respective neonatal control groups. Neurological abnormalities were related to OCs in siblings but not in patients. Neurological abnormalities in patients were negatively related to reduced neonatal head circumference. CONCLUSION: Early somatic trauma may increase the probability of neurological abnormality in individuals who are genetically 'at risk'. PMID- 10706016 TI - Outcome of schizophrenia in a non-industrialized society: comparative study between Bali and Tokyo. AB - OBJECTIVE: The aim of the present study was to contrast the outcome of schizophrenic patients between Bali and Tokyo, the former being a non industrialized society and the latter an industrialized society in Asia. METHOD: A total of 51 Balinese schizophrenics and 40 schizophrenics in Tokyo were evaluated by five outcome measures at a 5-year follow-up. RESULTS: No significant difference was found in the mean scores of the Positive and Negative Syndrome Scale, Eguma's Social Adjustment Scale and the re-admission rates between the subjects in the two sites. The cumulative length of stay in hospital during the 5 year period was significantly shorter in Bali. The percentage of subjects on psychiatric medication at the follow-up was significantly lower in Bali than that in Tokyo. CONCLUSION: Although the clinical outcome of schizophrenics in Bali was not superior to that in Tokyo, the subjects in Bali tended to be able to live in society without neuroleptic medication. PMID- 10706017 TI - Prevalence and background factors of depression in first admitted schizophrenic patients. AB - OBJECTIVE: The aim of this study was to investigate the prevalence and background factors of depression in first admitted schizophrenic patients. METHOD: The study is an analysis of 998 consecutively admitted schizophrenic patients with their first hospitalization. Patient's characteristics were prospectively assessed using standardized instruments at the time of first admission and discharge. RESULTS: High prevalence rates of depressive symptoms were found. Depressed schizophrenic patients were more likely to have suicidal tendencies, were older, more frequently married, less frequently single and unemployed and had more family members with psychiatric disorders other than schizophrenia than the non depressed patients. Positive, negative and extrapyramidal symptoms do not have a substantial influence on depression in these patients. CONCLUSION: The study suggests that depression represents a distinct psychopathological dimension of the acute illness in first admitted schizophrenic patients. In particular, in light of the suicidal tendencies, recognition and treatment of depression is an important clinical task. PMID- 10706018 TI - Neuromorphological abnormalities in schizophrenic patients with good and poor outcome. AB - OBJECTIVE: The present study was designed specifically to assess the relationship between brain morphology and outcome in schizophrenia. METHOD: Fifty-six schizophrenic patients and a matched group of 32 healthy subjects were studied with magnetic resonance (MR) imaging scans. Clinical assessment included the Krawiecka-Manchester Scale (K-MS) and the Outcome scale by Strauss and Carpenter. RESULTS: Along several neuromorphological measures the patients differed from controls only for right and left ventricular volumes. The 'poor outcome' patients had a left and right ventricular enlargement when compared to the 'good outcome' patients and healthy controls. A regression analysis showed that right ventricle volume, left temporal lobe volume and left hippocampal volume entered into the regression equation, accounting for a 27% of the outcome measure. CONCLUSION: The outcome does not seem to be predicted by one particular morphological site but involves different brain regions; however, the ventricular enlargement identifies a subgroup of patients with poor outcome. PMID- 10706019 TI - Association between oestradiol and puerperal psychosis. AB - OBJECTIVE: Postpartum psychiatric disorders with long-lasting adverse sequelae are common during the childbearing years. These disorders can be severe and resistant to conventional psychiatric treatment methods. We present two consecutive cases with puerperal psychosis who were refractory to conventional treatment methods but responded successfully to oestrogen therapy. METHOD: Serum oestradiol concentration was measured by radioimmunoassay and the documented oestradiol deficiency replaced with physiological oestradiol sublingually. The treatment effect was evaluated by the Brief Psychiatric Rating Scale. RESULTS: In both cases the baseline oestradiol concentration was low (28 and 69 pmol/L). During the treatment with oestradiol, there was a concomitant elevation of the concentration of serum oestradiol, which coincided with the decline in psychotic symptoms. CONCLUSION: The observation of low serum oestradiol together with psychotic symptoms and successful treatment with oestradiol suggests that oestradiol may have a causal relevance to puerperal psychosis and significance in the treatment of this condition. PMID- 10706020 TI - Positive rolandic sharp waves in preterm infants with periventricular leukomalacia: their relation to background electroencephalographic abnormalities. AB - The aim of this study was to clarify the significance of positive rolandic sharp waves (PRS) in preterm infants with periventricular leukomalacia (PVL) and their relation to background electroencephalographic (EEG) abnormalities. We retrospectively evaluated EEG findings of 93 preterm infants; 31 infants in the PVL group and 62 in the control group. PVL was diagnosed on the basis of ultrasonographic findings. We evaluated the EEG within 3 weeks of life in this study. PRS were defined as sharp transients of positive polarity appearing in the rolandic regions with an amplitude of more than 100 microV, sharply differentiated from the background activities. The number of PRS per minute was calculated over each record. PRS were defined as present when their frequency was beyond 0.1 per minute. PRS were observed in 14 (45%) and disorganized patterns in 27 (87%) of 31 infants in the PVL group, but both were not recognized in any infants in the control group. PRS were always associated with disorganized patterns. In the first EEG, PRS were absent in 8 of 11 infants with more than two recordings, although at least one of acute or chronic stage EEG abnormalities were already present. PRS were observed in 9 of 10 infants with severe diplegia, in 5 of 11 infants with moderate diplegia and none in 10 infants with mild diplegia. The average age of the first appearance of PRS was 7.6 days. The average age of the first appearance of periventricular echodensity and cyst was 4.2 days and 21.1 days, respectively. In conclusion, PRS are related to severe deep white matter injury and could be an early marker of severe PVL. PRS appeared in combination with disorganized patterns and were considered to be incorporated into chronic-stage EEG abnormalities. Detailed evaluation of background EEG activities can be helpful in detecting PVL with a high sensitivity from the early neonatal period. PMID- 10706021 TI - Bone marrow transplantation in aspartylglucosaminuria--histopathological and MRI study. AB - This study comprised two patients with aspartylglucosaminuria (AGU), who were followed up for 4 and 7 years. The patients underwent allogeneic bone marrow transplantation (BMT) at the ages of 2 and 2.6 years. Both patients had abnormal speech development and gross motor clumsiness. At the time of the BMT, they were mentally retarded. We report on follow-up data of these patients obtained by MRI, in addition to the histopathological, biochemical and clinical investigations. MR images of six non-transplanted patients and seven healthy children served as controls. In the non-transplanted patients, MRI revealed evident delay of myelination in contrast to the two transplanted patients showing fair or evident grey- vs. white matter differentiation on T2-weighted images. The aspartylglucosaminidase (AGA) activity in blood leukocytes reached a heterozygous level. Urinary excretion of aspartylglucosamine and glycoasparagines slowly decreased but remained about a third of the pre-BMT level 5 years after BMT. Storage lysosomes in electron microscopic investigations were not decreased 6 months after BMT, but after 1.5-2 years, rectal mucosa samples showed a decrease in the storage vacuoles of different cells. Three years after BMT, no cells with storage vacuoles were present. Allogeneic BMT slowly normalises the pathological, biochemical and MRI findings in patients with AGU. PMID- 10706022 TI - Ocular motor abnormalities in Gaucher disease. AB - Gaucher disease (GD) without primary neurological involvement (GD1) is now treatable with exogenous enzyme replacement therapy (ERT). ERT does not halt the fatal neurological progression of type 2 (infantile) disease (GD2), but in type 3 disease (GD3), neurological progression may be slowed down or possibly halted in some cases by higher doses of ERT. Therefore, it is crucial to distinguish between GD1 and GD3 disease for appropriate treatment. Saccade initiation failure (SIF) (ocular motor apraxia, supranuclear gaze palsy) is often the earliest neurological sign in GD3. This sign can be difficult to detect clinically, but is readily revealed as missed quick-phases during induced optokinetic and vestibular nystagmus. We investigated whether objective ocular motor assessment could improve the detection of GD3 disease. Eight children, diagnosed enzymatically with GD, were tested using D.C.-electro-oculography and/or video. In 6 children, optokinetic and vestibular nystagmus showed marked paucity of quick-phases making the eyes "lock up" at the limit of gaze, thus indicating SIF. In 3 cases the diagnosis was revised from GD1 to GD3. The diagnosis of GD3 was made in two children who were too ill for clinical assessment of SIF. Only 1 child had previous clinical evidence of SIF, and so GD3 was confirmed. One child was normal and the diagnosis of another remains uncertain. These results show that the possibility of SIF and hence GD3 may not be excluded by clinical examination alone. Thus we recommend that, where possible, objective eye movement assessment be carried out in children with GD. PMID- 10706023 TI - Serial intelligence test scores in pediatric moyamoya disease. AB - Serial intelligence tests in 38 patients with childhood moyamoya disease were evaluated. A total of 98 tests were administered. The IQ scores were classified into three categories: tests performed between the time of onset of symptoms and 5 years after the onset of symptoms (n = 44), tests performed 5-10 years after the onset of symptoms (n=32), and tests performed more than 10 years after the onset of symptoms (n=22). When more than one test was performed during each period, the mean of the IQ scores was used. The IQ tests were administered two or more times to 10 patients in the onset-5-years category, and 5 of them exhibited lower IQ scores on later tests. The IQ scores were significantly lower in the 5 10 years category and in the more than 10 years category (76.8 +/- 23.1 and 73.9 +/- 31.1, respectively) than in the onset-5-years category (92.9 +/- 22.7). The IQ scores for the 5-10 years category and the more than 10 years category did not differ significantly. The IQ in pediatric moyamoya disease begins to decrease after the onset of symptoms, but the decline eventually stabilizes more than 10 years after the onset of symptoms. PMID- 10706024 TI - Unilateral thalamic lesions in premature infants: risk factors and short-term prognosis. AB - The aim of the study was to assess incidence, risk factors, clinical symptomatology and short-term outcome of unilateral thalamic lesions in preterm infants, as detected by ultrasound. Sixteen preterm infants, born after a gestational age of less than 35 weeks, with a unilateral thalamic lesion, but without additional significant cerebral lesions, were included. Their follow-up data were compared to those of a selected control group consisting of healthy premature infants. In addition, the neonatal clinical data of the patients with a thalamic lesion were compared to data of the healthy control group and of a general control group, consisting of a non-selected year-cohort of preterm infants. During the study period, the incidence of unilateral thalamic lesions was 5.3% among preterm infants. Ultrasound was not able to distinguish between hemorrhagic and ischemic lesions. The infants with a unilateral thalamic lesion had a more complicated respiratory course and were ventilated significantly longer than infants without such a lesion. The infants with a thalamic lesion had disturbances in tone, persisting throughout infancy, while the healthy control group showed only transient disturbances in tone. PMID- 10706025 TI - Neuro-cognitive development and epilepsy outcome in children with surgically treated hemimegalencephaly. AB - We performed a long-term follow-up of 10 patients with hemimegalencephaly and refractory epilepsy, after having treated them with hemispherectomy. Before surgery, 9 patients presented with delayed motor and cognitive development. Surgery was performed between age 5 months and 4 years and 8 months; the mean postsurgical follow-up was 5 years and 2 months. The epilepsy improved in most cases: 6 patients became seizure-free and 2 presented only dystonic fits. The cognitive outcome was less favourable, even though some improvement of cognitive competence was found in all. The neurological deficit did not increase after surgery, and the quality of life improved significantly. A good cognitive development before surgery, less severe morphological changes in neuroimaging, and functional and anatomical integrity of the "healthy" hemisphere seem to be associated with a better cognitive outcome. PMID- 10706026 TI - Asymmetrical myelination of the posterior limb of the internal capsule in infants with periventricular haemorrhagic infarction: an early predictor of hemiplegia. AB - AIM: To prospectively assess the predictive value of asymmetrical myelination on MRI of the posterior limb of the internal capsule (PLIC) in newborn infants with an intraventricular haemorrhage (IVH) associated with unilateral haemorrhagic parenchymal involvement (PI), for subsequent development of a hemiplegia. METHODS: 12 preterm infants (GA 25-36 wks) and 4 full-term infants were studied. Using cranial ultrasound (US), the pre-term infants were diagnosed to have an IVH with unilateral PI. The term infants presented with a porencephalic cyst (PC) on the first postnatal US, following an antenatal IVH with PI. MRI was performed at 40 wks postmenstrual age in the pre-term infants and during the first 2 weeks of life in the full-term infants, using a 1.5T magnet. Using an inversion recovery sequence, the myelination of the internal capsule was recorded as normal, abnormal or equivocal. Neurological assessment > or = 12 months disclosed the presence of a hemiplegia or asymmetry in tone pattern. RESULTS: All 4 cases with a normal internal capsule had a normal outcome in spite of the development of a PC. All 9 cases with an abnormal PLIC developed a hemiplegia, while 1 of the 3 cases with an equivocal PLIC is normal on neurological assessment, one developed a mild asymmetry in tone and 1 a mild hemiplegia. CONCLUSION: While a symmetrical signal intensity within the internal capsule on MRI, performed at 40 weeks PMA, in infants with an IVH and unilateral PI appears to be strongly related to a normal outcome, an asymmetrical PLIC is an early predictor of future hemiplegia. PMID- 10706027 TI - Hereditary prothrombin deficiency presenting as intracranial haematoma in infancy. AB - Hereditary deficiency of prothrombin is a rare autosomal recessive bleeding disorder, with severe bleeding diathesis in homozygotes, but rarely resulting in intracranial haematoma. We describe two infants of consanguineous parents, presenting with acute subdural haematoma. Because such haematomas in infancy are highly indicative of trauma caused by child battering and because the socio economic status of the family was unstable, there was a suspicion of child battering. However, further investigations revealed a bleeding diathesis due to a prothrombin deficiency. DNA analysis of the prothrombin gene showed homozygosity for a novel mutation, substituting Lys for Glu at codon 7 and resulting in decreased specific clotting activity. We discuss the probability of bleeding diathesis versus child battering in the aetiology of intracranial haematoma. PMID- 10706028 TI - Preterm birth in Sjogren-Larsson syndrome. AB - Sjogren-Larsson syndrome (SLS) was originally described as a triad of spasticity, mental retardation and congenital ichthyosis. The syndrome reflects an underlying deficiency of microsomal fatty aldehyde dehydrogenase (FALDH). We report on clinical data concerning pregnancy, labor and neonatal period in 15 patients. Pregnancies were uncomplicated, except for preterm rupture of membranes in three pregnancies, and the occurrence of preterm birth. Mean gestational age was 35.3 weeks (S.D. 2.4 weeks), and preterm birth was found in 73% of the children, while all children were born before or in the 38th week of gestation. Birth weight was normal for gestational age in all patients. The neonatal period was free from serious complications, apart from hemolytic disease in two patients. Preterm birth was found in 7% of the healthy siblings, reflecting the normal population. Prematurity and spasticity are intrinsic and concurrent parts of SLS, without causal relation. SLS should be considered in every neonate with congenital ichthyosis, especially if the child is born preterm. A possible explanation for preterm birth in SLS could be the defective inactivation of leukotriene B4 (LTB4), which recently has been demonstrated in patients with SLS. PMID- 10706029 TI - Traumatic basilar artery dissection in a child: need for anticoagulation? AB - Dissection of a cerebral blood vessel is a rare complication of acute neurotrauma with a high incidence of morbidity and mortality. We report on a case of a pediatric patient with severe neurological symptoms in whom angiography showed evidence of a basilar artery dissection. The patient was heparinized and recovered uneventfully. PMID- 10706031 TI - Organochlorine insecticides in substantia nigra in Parkinson's disease. AB - The concentrations of organochlorine (OC) compounds in the substantia nigra (SN) were compared in Parkinson's disease (PD) with concentrations in brain from cortical Lewy body dementia (CLBD), Alzheimer's disease (AD), and nondemented nonparkinsonian controls (CON). The levels of the gamma isomer of hexachlorocyclohexane (gammaHCH, lindane) were significantly higher in PD tissues (mean +/- SD: 0.56 +/- 0.434 microg/g lipid) than in the other three groups (CLBD 0.052 +/- 0.101 microg/g lipid; AD none detected; CON 0.125 +/- 0.195; all differences from PD significant at p < .05, Mann-Whitney U-test). Dieldrin (HEOD) was higher in PD brain than in AD or control brain, while 1,1'-(2,2 dichloroethenyl diene)-bis(4-chlorobenzene) (p,p-DDE) and total Aroclor-matched polychlorinated biphenyls (matched PCBs) were only higher in PD substantia nigra when these concentrations were compared with those of CLBD. These findings are not inconsistent with the hypothesis derived from epidemiological work and animal studies that organochlorine insecticides produce a direct toxic action on the dopaminergic tracts of the substantia nigra and may contribute to the development of PD in those rendered susceptible by virtue of cytochrome P-450 polymorphism, excessive exposure, or other factors. PMID- 10706030 TI - Lung injury, inflammation, and inflammatory stimuli in rats exposed to ozone. AB - The effects of ozone (O3) on airway epithelia, inflammation, and expression of inflammatory stimuli were investigated to delineate the mechanisms of inflammatory reactions relevant to lung injury. Because the airway responses to O3 develop gradually, this investigation included a time-sequence analysis. Rats exposed for 3 h to 1 ppm O3 were studied at 4-h intervals up to 20 h postexposure. Bronchoalveolar lavage fluid (BAL) was analyzed for albumin as an indicator of increased permeability, polymorphonuclear leukocytes (PMNs) to assess the inflammatory status, macrophage inflammatory protein-2 (MIP-2, an inflammatory chemokine), and cell adhesion molecules for their role in inflammation and PMN functions. The time-related increase in albumin was matched by a similar significant increase for PMNs, MIP-2, and intercellular adhesion molecule-1 (ICAM-1). However, no marked change occurred for beta-2 integrin (CD 18) and leukotriene B4 (LTB4). The results establish a temporal correlation of epithelial permeability with changes in inflammatory activity and stimuli responsible for PMN recruitment in the lung. The observations of elevated MIP-2 and ICAM-1 levels are consistent with their role in injury and inflammation. An early expression of MIP-2 mRNA in BAL cells, that is, immediately post O3 exposure, and the peak increase in BAL MIP-2 levels 4 h later support the chemotactic role of MIP-2 in PMN recruitment at 4- and 12-h time points. The rapid drop in MIP-2 and ICAM-1 levels appears to signal the termination of inflammatory cell recruitment, which is accompanied by an onset of recovery. PMID- 10706032 TI - Developmental toxicity of lead-contaminated sediment in Canada geese (Branta canadensis). AB - Sediment ingestion has recently been identified as an important exposure route for toxicants in waterfowl. The effects of lead-contaminated sediment from the Coeur d'Alene River Basin (CDARB) in Idaho on posthatching development of Canada geese (Branta canadensis) were examined for 6 wk. Day-old goslings received either untreated control diet, clean sediment (48%) supplemented control diet, or CDARB sediment (3449 microg/g lead) supplemented diets at 12%, 24%, or 48%. The 12% CDARB diet resulted in a geometric mean blood lead concentration of 0.68 ppm (ww), with over 90% depression of red blood cell ALAD activity and over fourfold elevation of free erythrocyte protoporphyrin concentration. The 24% CDARB diet resulted in blood lead of 1.61 ppm with decreased hematocrit, hemoglobin, and plasma protein in addition to the effects just described. The 48% CDARB diet resulted in blood lead of 2.52 ppm with 22% mortality, decreased growth, and elevated plasma lactate dehydrogenase-L (LDH-L) activity. In this group the liver lead concentration was 6.57 ppm (ww), with twofold increases in hepatic lipid peroxidation (thiobarbituric acid-reactive substances, TBARS) and in reduced glutathione concentration; associated effects included elevated glutathione reductase activity but lower protein-bound thiols concentration and glucose-6 phosphate dehydrogenase (G-6-PDH) activity. The kidney lead concentration in this group was 14.93 ppm with subacute renal tubular nephrosis in one of the surviving goslings. Three other geese in this treatment group exhibited calcified areas of marrow, and one of these displayed severe chronic fibrosing pancreatitis. Lead from CDARB sediment accumulated less readily in gosling blood and tissues than reported in ducklings but at given concentrations was generally more toxic to goslings. Many of these effects were similar to those reported in wild geese and mallards within the Coeur d'Alene River Basin. PMID- 10706033 TI - Aluminum toxicity in a molluscan neuron: effects of counterions. AB - Previous studies using the freshwater snail Lymnaea stagnalis have indicated significant accumulation of aluminum (Al) from simple salts (chloride or nitrate) or Al lactate [Al(lactate)3 preparations, but not from the Al maltol complex [Al(maltol)3]. This is in contrast to findings in mammalian systems, where uptake and neurotoxicity are greatest for the soluble and lipophilic Al(maltol)3 complex. This study was undertaken to investigate the direct effects of extracellular Al (100 microM) from three Al preparations [AlCl3, Al(lactate)3 and Al(maltol)3] on electrophysiological parameters of an identified neuron, the right parietal dorsal 1 (RPD1) neuron, of L. stagnalis in vitro. The effects of the corresponding counterion/ligand on the solubility and availability of Al in solution were also examined. Significant effects of Al on electrical properties, including membrane depolarization, increased firing activity, and abnormal firing patterns, were seen in the presence of AlCl3 and Al(lactate)3, which formed polyhydroxy and labile Al species in aqueous solution, but not with Al(maltol)3, which remained as the soluble monomeric complex. Qualitative differences were also observed between the response to AlCl3 and Al(lactate)3, despite their similar chemistry. The extent of action potential broadening was greater with Al(lactate)3, suggesting some interaction between Al and lactate in their cellular uptake and/or toxicity. It is suggested that polyhydroxy Al species are toxic to molluscan neurons, possibly via disruption of intracellular Ca2+ homeostasis. PMID- 10706034 TI - Dietary indole-3-carbinol alters immune functions in rats. AB - To further elucidate the physiological mechanisms that may contribute to the dichotomy of effect of indole-3-carbinol (I3C) on cancer development, we examined immune functions representative of the three major branches of the immune system in rats fed either a high (150 mg/kg) or low (50 mg/kg) dose of I3C. Animals fed the high dose of I3C daily for 7 wk had significantly reduced natural killer cell activity. In contrast, T-cell-mediated delayed-type hypersensitivity was significantly elevated. Antibody production in response to the antigen keyhole limpet hemocyanin was not significantly altered compared to controls. These results indicate that exposure to I3C may have differential effects on major immune responses. The significance of these immune function alterations in tumor development will require additional investigation of the effects of dietary I3C on immune functions in appropriate tumor models. PMID- 10706035 TI - Anti-HLA antibodies after solid organ transplantation. AB - We have cited more than 23 studies showing that de novo development of anti-HLA antibodies is associated with increased acute and chronic rejection and decreased graft survival in kidney, heart, lung, liver, and corneal transplants. Antibodies to both HLA class I and class II antigens seem to be detrimental. Antibodies of the IgG isotype and possibly the IgM isotype were clinically relevant. Most studies showed that donor-specific antibodies were associated with rejection and graft loss. Therefore, HLA antibodies provide a clinical readout for patient alloreactivity that may have the ability to distinguish graft dysfunction due to immunologic and nonimmunologic causes. Antibody may act as a critical trigger for rejection of allografts and may serve as an early indicator of a slowly smoldering chronic rejection that is not manifested at a given time by biochemical measures such as serum creatinine levels. The effectiveness of various drugs on chronic rejection should be evaluable by their effects on HLA antibody production. We predict that recently developed ELISA and flow cytometry techniques using purified HLA antigen will increase the clinical relevance of posttransplantation HLA antibody monitoring by (1) allowing the detection of low levels of donor antibody; (2) easily distinguishing the isotype and target (HLA class I or class II) of the antibodies; and (3) correlating the antibody with specific graft pathology. PMID- 10706036 TI - Ischaemia/reperfusion injury and inflammation. PMID- 10706037 TI - CD25-targeted therapy revisited. PMID- 10706038 TI - Functional responses of T cells blocked by anti-CD25 antibody therapy during cardiac rejection. AB - BACKGROUND: Despite anti-CD25 (interleukin [IL]-2 receptor alpha chain) monoclonal antibody (mAb) therapy, rejection can still occur. T-cell activation through the IL-2 receptor beta and gamma chains by IL-2 or other growth factors may contribute to this rejection. Recently, we have demonstrated that the T-cell growth factor IL-15 was abundantly present in rejecting cardiac grafts during anti-CD25 mAb treatment. METHODS: To test whether IL-2- and IL-15-responsive T cells play an active role in rejection during anti-CD25 mAb therapy, we measured the frequency of IL-2- and IL-15-proliferative T cells in peripheral blood from treated patients during rejection (n=12). Measurements were made by limiting dilution analysis in the absence and presence of extra in vitro-added mouse anti human CD25 mAb. RESULTS: In the absence of anti-CD25 mAb, the frequencies of peripheral T cells responding to recombinant human (rh)IL-2 and rhIL-15 from patients were lower than those measured in samples of healthy controls (n=7): median of IL-2-responding T cells 78 per 10(6) (range 31-210 per 10(6)) vs. 154 per 10(6) (122-484 per 10(6), P=0.008) and median of IL-15-responding T cells 62 per 10(6) (range 19-207 per 10(6)) vs. 129 per 10(6) (range 79-192 per 10(6), P=0.02), respectively. In the presence of extra in vitro-added anti-CD25 mAb, frequencies of IL-2-responding T cells from patients significantly decreased, although a considerable number of T cells still proliferated on rhIL-2 (median 85%, range 46-100%). In contrast, the frequencies of IL-15 T cells still responding remained stable (median 2%, range 0-50%, P<0.001). CONCLUSIONS: Treatment with anti-CD25 mAbs cannot provide complete suppression of T-cell function because significant numbers of IL-2- and IL-15-responsive T cells remain present in the peripheral blood of allograft recipients during anti-CD25 mAb treatment. PMID- 10706039 TI - In vivo efficacy of a bioartificial liver in improving spontaneous recovery from fulminant hepatic failure: a controlled study in pigs. AB - BACKGROUND: Bioartificial liver may be useful as a bridge to liver transplantation but there are no data of its efficacy in successfully bridging to spontaneous recovery in fulminant hepatic failure. The aim of our study was to evaluate the efficacy of a bioartificial liver in increasing the spontaneous recovery of pigs with hepatic failure. METHODS: The bioartificial liver consisted in a semipermeable dialyzer with 0.6 x 10(9) cryopreserved allogenic hepatocytes. Hepatic failure was induced by portacaval shunt plus 70% hepatectomy and 1 hour occlusion of the hepatic artery. Forty-one pigs were distributed 24 hr after liver failure induction to a group treated with the bioartificial liver (4 hr daily) until recovery or death (n=16), or to a control group (n=25). Intracranial pressure was monitored in 18 additional pigs, before and 4 hr after treatment with the bioartificial liver with (n=12) or without hepatocytes (n=6). RESULTS: Fifteen days after induction of hepatic failure, 44% of the treated animals had survived and recovered from liver failure versus 22% controls (P=0.030). Intracranial pressure decreased from 13.13+/-5.1 to 7.19+/-2.06 mmHg (P=0.02) in treated animals, and remained unchanged in sham-treated animals (14.08+/-1.92 to 12.54+/-3.82, ns). CONCLUSIONS: Bioartificial liver increases survival and allows spontaneous recovery in pigs with fulminant hepatic failure. PMID- 10706040 TI - Donor treatment with mycophenolate mofetil: protection against ischemia reperfusion injury in the rat. AB - BACKGROUND: Mycophenolic acid, the active metabolite of mycophenolate mofetil, inhibits the glycosylation of cell membrane glycoproteins. We hypothesized that impaired glycosylation of cell adhesion molecules on endothelial cells in vivo results in decreased susceptibility to inflammation or immunogenicity after allogeneic transplantation. METHODS: The expression of mannose residues on cultured rat endothelial cells was examined after stimulation with interleukin 1 in the presence or absence of mycophenolic acid using labeled Galanthus nivalis agglutinin. The in vitro adhesion of blood leukocytes to heart tissue was examined using peripheral blood leukocytes of recipient origin and sections of donor heart tissue exposed to ischemia-reperfusion injury after pretreatment with vehicle or mycophenolic mofetil. (LEWxBN)F1 donor rats were treated with 20 or 60 mg/kg/day of mycophenolate mofetil for 1 or 2 weeks followed by transplantation of the heart into Lewis recipients after storage in heparin-containing normal saline for either 10 min at 4 degrees C or 120 min at room temperature. RESULTS: Endothelial cells stimulated in vitro with interleukin 1 showed an increase in a population of strongly mannose-positive cells, which was prevented by the addition of mycophenolic acid during the culture. The in vitro adhesion of peripheral blood leukocytes to cardiac tissue sections exposed to prolonged storage and reperfusion was significantly less if the donor had been treated with mycophenolate mofetil. Treatment of cardiac graft donors with mycophenolate mofetil protected the graft against early graft failure after prolonged storage at room temperature, because the mean graft survival was 9.4+/-0.6 days for grafts that came from donors treated with mycophenolate mofetil versus 1.2+/-0.9 days (P<0.05) for grafts that came from vehicle-treated donors. Donor pretreatment with mycophenolate mofetil did not affect the survival time of heart grafts transplanted after 15 min of standard cold storage or the survival of grafts transplanted into presensitized recipients. CONCLUSION: Donor treatment with mycophenolate mofetil protects cardiac grafts against primary nonfunction after prolonged tepid storage, which may be related to the inhibition of glycosylation of cell adhesion molecules involved in ischemia-reperfusion injury. PMID- 10706041 TI - Viremia and excretion of TT virus in immunosuppressed heart transplant recipients and in immunocompetent individuals. AB - BACKGROUND: The TT virus (TTV) was discovered in patients with symptomatic posttransfusion hepatitis, but many viremic individuals are asymptomatic. Inadvertent transfusion-associated transmission must therefore be anticipated. We screened blood donors and heart transplant recipients for TTV infections. METHODS: Nested polymerase chain reaction was used to detect TTV DNA in plasma, serum, urine, and fecal samples from 600 blood donors, from 100 healthy individuals, and from 495 heart transplant recipients. RESULTS: A total of 3.2% of the blood donors, but 25% of the heart transplant recipients were viremic. TTV subtypes G1a/b and G2a/b were observed in both groups, but the subtype distributions were discrepant. A severe, acute infection with TTV subtype 3 was observed in one blood donor. The prevalence of TTV infections in heart transplant recipients was not correlated to transfusion frequency. Nine viremic heart transplant recipients and their 75 blood donors were studied in detail. Seven blood donors were viremic, but only two "pairs" of viremic blood donors and transfusion recipients had identical TTV isolates. TTV DNA was detected in the feces of 5% (5/100) of immunocompetent individuals (staff), in 46% (52/112) of viremic heart transplant recipients, and in the urine of 55% (20/36). TTV DNA was detected in six of six batches of pooled "virus-inactivated" plasma (solvent/detergent treated), and in none of eight batches of commercial immunoglobulins. CONCLUSION: Although TTV is transfusion-transmissible, the parenteral transmission rate may have been overestimated. Many TTV infections are apparently acquired by nonparenteral routes. Immunoglobulins are safe but pooled plasma is not safe regarding TTV transmission. PMID- 10706042 TI - The economic impact of cytomegalovirus infection after liver transplantation. AB - BACKGROUND: We studied the economic impact of cytomegalovirus (CMV) disease and its effective reduction with antiviral prophylaxis in liver transplant recipients. METHOD: Analysis of institutional charge data accumulated during a prospective, randomized, controlled trial comparing oral acyclovir 800 mg four times daily for 120 days (ACV) and intravenous ganciclovir 5 mg/kg every 12 h for 14 days followed by ACV for 106 days (GCV) was performed. RESULTS: Liver transplant recipients who developed CMV disease had significantly higher charges (median: $148,300) than those who developed asymptomatic CMV infection ($119,600) or experienced no CMV infection ($114,100) (P<0.01). A multiple linear regression analysis indicated that CMV disease is associated with a 49% increase in charges, independent of other factors influencing increased hospitalization charges. In CMV-seronegative patients who received a CMV-seropositive donor organ, GCV prophylaxis was associated with a significant reduction in charges, as compared to ACV prophylaxis ($113,900 vs. $153,300, respectively; P=0.02). CONCLUSIONS: CMV disease is an independent risk factor for increased resource utilization associated with liver transplantation. The use of an effective prophylactic antiviral regimen provides savings in health care resources, particularly in patients at high risk for developing CMV disease. PMID- 10706043 TI - Disaccharidase activities and fat assimilation in pediatric patients after intestinal transplantation. AB - BACKGROUND: Intestinal transplantation has become an accepted therapy for short bowel syndrome and other types of intestinal failure. In order to assess digestive capabilities and feeding practices in a group of 22 pediatric patients after intestinal transplantation, we assessed mucosal disaccharidase activities and assimilation of total dietary lipid and vitamin E. Twelve of the patients had undergone contemporaneous liver transplantation. METHODS: Mucosal biopsies were assayed for disaccharidase activities between 15 and 412 days after transplantation in 7 of the 22 when all were receiving some enteral nutrition and were free of rejection. Coefficients of lipid absorption were determined in those patients receiving total enteral feeding (two-thirds polymeric/one-third elemental) between 43 and 1032 days after transplantation; oral vitamin E tolerance tests were done at about the same time. RESULTS: Activities of lactase, sucrase, maltase, and palatinase consistently exceeded reference ranges (P<0.05). Mean coefficient of lipid absorption equaled 86+/-12% and was not influenced by duration of time after transplantation. No patient required dietary lipid restriction. No significant absorption of vitamin E was demonstrated until 160 days after transplantation. Vitamin E absorption did correlate with length of time elapsed after surgery (r=0.64, P<0.0011). CONCLUSIONS: The results of this investigation show that, in the absence of histologic or clinical indications of allograft rejection, pediatric intestinal transplant recipients do not have primary disaccharidase deficiencies. Similarly, absorption of usual dietary lipid content is adequate once weaning from parenteral nutrition is complete. In contrast, early assimilation of vitamin E is poor. Vitamin E absorption subsequently improves, but the mechanism is obscure. PMID- 10706044 TI - Early increase of peripheral B cell levels in kidney transplant recipients with CMV infection or reactivation. AB - BACKGROUND: Cytomegalovirus (CMV) infection or reactivation is a frequent complication of renal transplantation. Diagnosis of these conditions relies on the detection of circulating antigen, or of specific IgM and/or IgG, which develop over several weeks. Evocative clinical features may be detected earlier, but lack specificity. Rapid and early changes in the partition of lymphocyte subsets could be an additional indication of pending CMV infection. METHODS: A systematic follow-up of peripheral B lymphocytes identified immunophenotypically by the determination of surface immunoglobulins (sIg), performed in 97 kidney transplant recipients, allowed to identify transient increases apparently predictive of CMV primo-infection or reactivation over the next 3 months. To better define the nature of these B cells, an extended investigation was performed for 14 prospective patients. In addition to surface Ig, membrane CD19, HLA-DR, and CD80 expression were explored. The cytoplasmic presence of mu, kappa, and lambda chains was also examined. B cell function was investigated using the ELISPOT technique, which allows an enumeration of the populations of IgG, IgA, and IgM secreting B cells. RESULTS: Retrospective analysis of the clinical outcome of the cohort of 97 patients evidenced that early transient increases in B cell levels were significantly (P<0.0001) associated with CMV infection. The same trend was noted in the smaller series of patients who benefited from a more extensive investigation of B cells, 10 of whom presented clinical or biological signs of CMV infection. Mature B cells, expressing surface Ig, CD19, DR, and CD80 are those presenting transient increases. No significant variation of preB (cmu+/kappalambda-) or activated (spot-forming) cells was evidenced in these patients. CONCLUSION: Individual examination of each patient's immune reconstitution profile allows to detect transient peaks of mature B cell during the initial immunosuppressive therapy, that appear to be predictive of oncoming CMV infection or reactivation. PMID- 10706045 TI - The domino transplant: transplant recipients as organ donors. PMID- 10706046 TI - Conversion to mycophenolate mofetil in conjunction with stepwise withdrawal of cyclosporine in stable renal transplant recipients. AB - BACKGROUND: Mycophenolate mofetil (MMF) is now part of standard immunosuppression in the first phase after renal transplantation. A relevant question is if it can replace drugs such as cyclosporine (CsA) in the maintenance treatment, improving cardiovascular risk profile. METHODS: In 17 patients with a stable renal function (at least 6 months) posttransplantation, we studied the effect of CsA replacement by MMF. After starting MMF (1 g b.i.d.), CsA dosage was reduced from regular to low (median trough level 130 microg/L, respectively, 45 microg/L), followed by complete withdrawal, while prednisone (7.5 mg daily) was continued. We measured ambulatory blood pressure, glomerular filtration rate, renal plasma flow, renal vascular resistance, and metabolic factors at start and after 8 weeks on regular, low-dose CsA, respectively, no CsA with MMF and prednisone. RESULTS: Two patients dropped out after the switch to low-dose CsA/MMF, due to diarrhea in one and a steroid responsive rejection in the other. The complete switch from CsA to MMF was successful in all 15 patients and accompanied by a decrease in 24 hr systolic blood pressure (from 152+/-13 to 145+/-13 mmHg; P<0.01), diastolic blood pressure (93+/-9 to 89+/-12 mmHg; P<0.05), RVR (0.29+/-0.06 to 0.25+/-0.09 mmHg.ml/min; P<0.05), and an increase in glomerular filtration rate (46.6+/-8.8 to 58.0+/-10.5 ml/min; P<0.01) and renal plasma flow. Intermediate low density lipoprotein cholesterol decreased (0.79+/-0.37 to 0.41+/-0.16 mmol/L; P<0.01). High density lipoprotein-cholesterol decreased, but remained in the safe range. After 1 year two patients stopped the MMF; one because of Kaposi's sarcoma and one because of recurrent infections CONCLUSIONS: The stepwise switch from CsA to MMF was safe and mostly successful, and had beneficial effects on blood pressure, glomerular hemodynamics, and lipid profile. Beneficial trends were already present after partial withdrawal of CsA. PMID- 10706047 TI - Late-onset renal failure after liver transplantation: role of posttransplant alcohol use. AB - BACKGROUND: Late-onset renal failure is being increasingly recognized as a complication in patients undergoing liver transplantation for hepatitis C virus (HCV). However, its precise incidence, predisposing risk factors, and impact on outcome after liver transplantation, have not been defined. METHODS: The development of late-onset renal failure (defined as serum creatinine persistently >2.0 mg/dl, occurring more than 6 months posttransplant) was assessed in 120 consecutive liver transplant recipients who survived at least 6 months after transplantation. Fifty-seven percent (68/120) of the patients had undergone transplantation for liver disease due to HCV. The median follow-up was 5 years. RESULTS: Late-onset renal failure developed in 28% (33/120)of the patients. Posttransplant alcohol use (P=0.0001), posttransplant diabetes (P=0.0042), and recurrent HCV hepatitis (P=0.019) were significantly associated with late onset renal failure. In multivariate analysis, alcohol use (O.R. 10.7, 95%; CI 2.4 35.9, P=0.001) and diabetes (O.R. 2.1, 95%; CI 1.1-9.9, P=.03) were independently significant predictors of late onset renal failure. When only patients transplanted for HCV were analyzed, posttransplant alcohol use (P=0.004) was the only significant independent predictor of late-onset renal failure. HCV genotype 1b, as compared with other HCV genotypes, was associated with a higher rate of late-onset renal failure in patients with HCV; 70% of the patients with genotype 1b versus 32% of those with 1a and 33% of those with 2b, developed late onset renal failure (P=0.03). At a median follow up of 5 years, mortality in patients with HCV with late-onset renal failure was 52% as compared with 2% in those without renal failure (P=.0001). CONCLUSION: Late-onset renal failure in patients with HCV portended a grave outcome. Alcohol use was an independent predictor of late-onset renal failure in patients with HCV and represents a potentially modifiable risk factor for late-onset renal failure in these patients. PMID- 10706048 TI - Dosing of intravenous ganciclovir for the prophylaxis and treatment of cytomegalovirus infection in solid organ transplant recipients. AB - BACKGROUND: The optimal regimen for the prevention and treatment of cytomegalovirus (CMV) disease in solid organ transplant recipients remains to be defined, particularly for patients with abnormal or changing renal function. METHODS: A prospective trial was conducted in patients receiving i.v. ganciclovir using a standardized dosing nomogram that corrects for renal function. Steady state peak (P) and trough (T) serum levels were determined by high-performance liquid chromatography and correlated with therapeutic outcomes and toxicities attributable to ganciclovir. RESULTS: Over the study period, 44 individuals received ganciclovir prophylaxis (5 mg(kg/day) and 25 patients were treated (5 mg/kd q12 hr) for symptomatic CMV disease. Ganciclovir levels (microg/ml+/-SD) achieved in prophylaxis were P: 7.98+/-3.34, T: 3.03+/-2.63; and in treatment were P: 9.00+/-3.72, T: 2.65+/-1.82. Despite corrections for renal dysfunction, undialyzed patients with serum creatinine >3.0 mg/dl had trough levels in excess of the population mean (T: range 3-8 microg/ml). Failure of prophylaxis (disease) or therapy (relapse) occurred in 14 patients; 8 of these were at risk for primary infection (donor CMV seropositive, recipient seronegative, P<0.01). Patients at greatest risk for relapse after treatment of CMV disease were liver transplant recipients, patients with ganciclovir-resistant viral isolates, and renal patients with six antigen MHC donor-recipient mismatches. CONCLUSIONS: This trial demonstrates the efficacy of a nomogram for ganciclovir dosing during renal dysfunction; reduced doses can be used for prophylaxis for undialyzed patients with renal dysfunction (1.25 mg/kg/day for Cr > or =3.0, 1.25 mg/kg QOD for Cr > or =5.0). Some groups of transplant recipients may require more intensive anti CMV regimens. PMID- 10706049 TI - Cross-species compatibility of intercellular adhesion molecule-1 (CD54) with its ligands. AB - BACKGROUND: The molecular interactions of intercellular adhesion molecule-1 (ICAM 1; CD54) are potentially important in several situations in the context of pig-to human xenotransplantation. If porcine bone marrow is to be used for the induction of xenograft tolerance in humans, the role that has been suggested for ICAM-1 in the interactions of haematopoietic stem cells makes its cross-species compatibility important. Similarly, the potential role of ICAM-1 interactions in graft rejection makes it an important molecule to study. METHODS: An in vitro static cell-to-cell adhesion study was used to look at the successful interaction of ICAM-1 with its ligands across the pig-human species barrier in both directions. A second in vitro system, the standard long-term bone marrow culture (LT-BMC), was used to study the functional role of ICAM-1 in haematopoiesis. RESULTS: Human ICAM-1 was able to adhere to ligands on porcine cells, including one or more ligand that contains CD18. Conversely, human CD18-containing ligands mediated adherence to porcine cells. Using the long-term bone marrow culture system, there was no evidence that blocking the interactions of ICAM-1 inhibited hematopoiesis, either in the human-human or pig-human combinations of precursor cells and marrow stroma. CONCLUSIONS: ICAM-1 is able to interact with at least some of its ligands across the species barrier, in both pig-human and human-pig combinations. However, the interactions of ICAM-1 do not appear to be central to hematopoiesis, at least in the model system used. PMID- 10706050 TI - Enhanced T cell cytokine gene expression in mouse airway obliterative bronchiolitis. AB - BACKGROUND: Obliterative bronchiolitis (OB), chronic allograft rejection of the lung, is a major cause of morbidity and mortality after lung transplantation. Previous studies using the heterotopic mouse trachea model of chronic airway rejection have shown a T cell infiltrate composed of CD4+ and CD8+ T cells. The goal of these experiments was to characterize the pattern of T lymphocyte cytokines during chronic airway rejection using the heterotopic mouse trachea model. METHODS: Isografts (BALB/c into BALB/c) and allografts (BALB/c into C57BL/6) were implanted into cyclosporin-treated animals and harvested 2, 4, 6, and 10 weeks posttransplant. Cytokine mRNA expression in these grafts was determined using reverse transcription polymerase chain reactions. Expression of Th1 cytokines, interleukin- (IL) 2 and gamma-interferon, and Th2 cytokines, IL-4, and IL-10 were analyzed, as well as the cytotoxic lymphocyte product granzyme B and expressed relative to beta-actin gene expression. RESULTS: In allografts, expression of IL-2 (P=0.002), gamma-interferon (P=2x10(-6)), granzyme B (P=0.003), IL-4 (P=0.06), and IL-10 (P=8x10(-6)) were 2- to 10-fold higher compared to isografts throughout the time-course of graft injury. Th1 and cytotoxic lymphocyte gene expression were increased to a greater extent than Th2 cytokines in allografts compared with isografts, and both Th1 and Th2 cytokine gene expression persisted at 6-10 weeks. CONCLUSIONS: These data suggest that Th1, Th2, and cytotoxic lymphocyte subtypes all contribute to the development of obliterative bronchiolitis in the heterotopic mouse trachea model. Efforts to reduce the development of obliterative bronchiolitis may require the antagonism of multiple T cell pathways. PMID- 10706051 TI - Activation of inflammatory mediators in rat renal isografts by donor brain death. AB - BACKGROUND: Brain death (BD) has been thought to influence the early course of transplanted organs by triggering a series of nonspecific inflammatory events that in turn may increase the kinetics and intensity of the immunological host responses. In this study early nonspecific, cellular, and molecular changes occurring in kidney isografts from BD donors are compared with those from normal anesthetized, ventilated controls. METHODS: After induction of brain death, the animals were mechanically ventilated for 6 hr before organ removal. Only rats with stable blood pressure (mean arterial pressure >80 mmHg) were included. Serum creatinines were measured daily. Representative grafts were harvested 6 hr after brain death and between 1 hr and 5 days after engraftment for morphology, immunohistology, and reverse transcriptase-polymerase chain reaction. The presence of serum cytokines was assessed by enzyme linked immunoabsorbant assay. RESULTS: Serum creatinine levels rose slightly in recipients from BD donors. Serum interleukin-1beta levels increased within 6 hr versus controls (P<0.05). mRNA levels of interleukin-1beta and macrophage inhibitory protein-1 in the kidneys were up-regulated transiently before engraftment (6 hr after BD) and 1 hr after revascularization (P<0.05). By immunohistology, numbers of infiltrating polymorphonuclear leukocytes peaked at 24 hr in parallel with intragraft induction of P- and E-selectin, complement, and other proinflammatory chemokines and cytokines. At 5 days, the isografts from BD donors were highly infiltrated by host leukocyte populations associated with intense up-regulation of their products. In contrast, those from control donors remained relatively normal through this initial follow-up period. CONCLUSIONS: The intense nonimmune inflammation produced in isografts after donor BD may represent the initial stages of a continuum between an initial nonspecific and later immune reactivity, when placed in the context of allotransplantation. PMID- 10706052 TI - The importance of H2 haplotype sharing in the induction of specific unresponsiveness by pretransplant blood transfusions. AB - BACKGROUND: The beneficial effect on graft survival achieved by pretransplant blood transfusions is well established. Previous studies have shown that the degree of major histocompatibility complex (MHC) (mis)-match between the transfusion donor and the recipient plays a determining role. However, other factors are also involved. In this study, we explored the hypothesis that, in addition to sharing of MHC antigens between the transfusion donor and the recipient, the MHC type of the organ donor is also of importance. METHODS: To mimic the human situation, F1 mice, rather than inbred strains, were pretreated with haplotype-shared allogeneic whole blood transfusions and transplanted with hearts of organ donors with different matched or mismatched H2 haplotypes. RESULTS: When a heart was transplanted 1 week after donor-specific transfusion (DST; blood transfusion donor=organ donor), an excellent prolongation of graft survival was obtained (median survival time: 77 days vs. 9 days in untreated mice). However, this was only the case when a haplotype was shared with the recipient; a DST given with no match between organ donor (=BT donor) and recipient did not induce any prolongation. Furthermore, in order to obtain the optimal beneficial effect of a haplotype-shared blood transfusion, the other haplotype of the transfusion donor had to be mismatched with the recipient. The immunogenetic studies showed that haplotype-shared blood transfusions in combinations where the H2 type of the organ donor differed from that of the transfusion donor are less efficient in inducing prolongation of graft survival. CONCLUSIONS: These results demonstrate that haplotype-shared blood transfusions can induce a significantly prolonged survival of cardiac allografts in F1 mice. The immunogenetic studies suggest that presentation of alloantigen-derived peptides in the context of self MHC (the indirect pathway of allorecognition) is essential for the beneficial effect of haplotype-shared blood transfusions. PMID- 10706053 TI - Attenuation of cytomegalovirus-induced endothelial intercellular adhesion molecule-1 mRNA/protein expression and T lymphocyte adhesion by a 2'-O methoxyethyl antisense oligonucleotide. AB - BACKGROUND: Intercellular adhesion molecule-1 (ICAM-1) is strongly induced under inflammatory conditions associated with allograft rejection, thereby promoting leukocyte recruitment and activation at the site of inflammation. Enhancement of ICAM-1 expression can also be the result of viral infection, in particular human cytomegalovirus (CMV), a frequent source of complications in the transplant recipient. In vitro studies have shown that CMV infection of endothelial cells (EC) results in the direct enhancement of ICAM-1 expression and consequent leukocyte adhesion/activation suggesting mechanisms by which CMV exacerbates graft vascular disease. Although treatment of EC with ICAM-1-specific antisense oligonucleotides has been shown to attenuate ICAM-1 induction under simulated inflammatory conditions (i.e., TNF-alpha), no studies have addressed their effectiveness on virally-induced ICAM-1 expression. RESULTS: In the current investigation, we show that the progressive increase in endothelial ICAM-1 protein expression that follows inoculation with CMV correlates with a progressive accumulation of ICAM-1 mRNA. Furthermore, we demonstrate that treatment of EC with a partially 2'-O-methoxyethyl modified ICAM-1-specific antisense oligonucleotide before viral inoculation significantly reduces CMV associated induction of ICAM-1 protein and mRNA expression. Finally, we show that antisense-mediated attenuation in ICAM-1 expression results in a significant reduction of T lymphocyte adhesion to CMV-infected EC monolayers, an interaction that has been implicated in allogeneic T lymphocyte activation, in viral transmission to transiently adherent leukocytes and subsequent hematogenous dissemination. CONCLUSIONS: These findings demonstrate for the first time that antisense oligonucleotides can effectively reverse virally-induced host cellular protein expression, specifically ICAM-1, as well as consequent T lymphocytes adhesion, thus broadening the potential clinical utility of antisense oligonucleotides. PMID- 10706054 TI - A model of gradual onset brain death for transplant-associated studies in rats. AB - BACKGROUND: The relatively few studies that have examined the systemic events after brain death have primarily involved large animals. For more precise definition of the physiology of this central catastrophe and its influence on peripheral organs, we have established a reproduceable model of gradual onset brain death in rats. METHODS: The central injury is induced by graded inflation of a Fogarty catheter placed intracranially under EEG and blood pressure monitoring. The rats were mechanically ventilated for 6 hr before removal of their kidneys. Complications and mortality are discussed. RESULTS: The majority (83%) of the 100 experimental animals could be used as organ donors. After a transient period of autonomic storm, the mean arterial blood pressure remained consistently between 80-100 mmHg, not appreciably different from controls. Despite normotension, the transplanted kidneys from brain dead donors showed a significantly longer interval to regain uniform cortical color and turgor than kidneys from control animals. CONCLUSIONS: We describe a controlled model of gradual onset brain death in the rat in which normotension can be sustained for several hours before the kidneys are removed for transplantation. Despite stable donor blood pressure, ischemia of peripheral organs may explain in part the increased incidence of delayed graft function of cadaver kidneys compared with those from living donors. This model is suitable for transplant-related studies involving organs from donors with irreversible central injury. PMID- 10706055 TI - Treatment of posttransplant lymphoproliferative disorder with the anti-CD20 monoclonal antibody rituximab alone in an adult after liver transplantation: a new drug in therapy of patients with posttransplant lymphoproliferative disorder after solid organ transplantation? AB - BACKGROUND: Posttransplant lymphoproliferative (PT-LPD) disorder is a life threatening complication with an incidence of 1-10%. Uniform treatment, so far, does not exist. METHODS: In December 1996, 5 months after a liver transplant, a 43-year-old patient developed a PT-LPD with para-aortal lymphomas and splenomegaly. Histological investigations revealed a PT-LPD of a diffuse large B cell type of the centroblastic variant. The patient received three cycles of a modified cyclophosphamide, doxorubicin, vincristine, and prednisone-regimen, resulting in complete remission but the patient withdrew from further treatment. In February 1998, the patient had a recurrence of PT-LPD with gastric involvement and parasplenic lymphomas. The patient rejected cytotoxic treatment because of her fear of drug-induced progressive myopathy, so we conducted treatment with the monoclonal antibody--directed against CD20-rituximab. RESULTS AND CONCLUSIONS: After 2 doses of rituximab, clinical symptoms had disappeared and after 6 doses, gastroscopy revealed no residual disease. At this time, the patient remains in remission, with a follow up of > or =6 months. Anti-CD20 monoclonal antibody rituximab is a new, well-tolerated drug for the treatment of lymphomas. In addition, this drug may offer an additional treatment option for patients with PT LPDs. PMID- 10706056 TI - Stimulated response of peripheral lymphocytes may distinguish cyclosporine effect in renal transplant recipients receiving a cyclosporine+rapamycin regimen. AB - BACKGROUND: Clinically, cyclosporine (CSA, Neoral) is titrated to concentrations, and not to pharmacological effect. METHODS: Intracellular interleukin- (IL) 2 was measured in phorbol myristic acid-ionomycin-stimulated peripheral lymphocytes by flow cytometry, after isolation from 14 renal transplant recipients receiving CSA+prednisone, and double-blind rapamycin (rapamycin:placebo=4:1). RESULTS: The proportion (%) of CD4+IL-2+ lymphocytes corresponding to CSA levels (mean+/-SD ng/ml) measured preoperatively (TO=O), and on postoperative day 8, before (356+/ 63), and 2 hr after the morning dose (Cmax=1567+/-669), decreased from 39+/-16 to 15+/-8 and 3+/-1.6, respectively. Reciprocally, unresponsive lymphocytes (%CD4+IL 2-) increased with increasing CSA levels and predicted an EC50 of 249 ng/ml (CSA concentration at which CD4+IL-2- cells increased by 50% over baseline) in an Emax pharmacodynamic model. CONCLUSIONS: Clinically, the pharmacological effect of CSA is quantifiable, and lies in the upper end of the predicted range. In our Neoral treated sample population, Cmax was associated with the least variable "cyclosporine effect." Such information could potentially individualize immunosuppression, and lead to rational dosing strategies. PMID- 10706057 TI - Influenza virus immunization effectivity in kidney transplant patients subjected to two different triple-drug therapy immunosuppression protocols: mycophenolate versus azathioprine. AB - BACKGROUND: Due to possible complications and treatment limitations, the prevention of influenza in renal transplant (RT) patients is highly indicated. METHODS: Forty-nine patients with a 1-year functioning RT subjected to two different immunosuppressive regimens and 37 healthy relatives (HR) were administered the anti-influenza vaccine as recommended for 1996 to 1997. Anti influenza antibody, creatinine, and immunological markers were estimated at 1 and 3 months after vaccination. RESULTS: Three months after vaccination, 46.2% of the RT patients and 69% of the HR (P=0.06) showed protective antibody titers to influenza A (relative risk [RR]; 0.67; 95% confidence interval: 0.44-1.02). A total of 20.5% of the RT patients and 44.8% of the HR showed antibodies to influenza B (P=0.03). Despite these differences, the incidence of illness was similar. The immunosuppressive regimen had no effect on the antibody response. CONCLUSIONS: Although the RT patients showed a reduced antibody response, no negative effects on graft outcome were observed. PMID- 10706058 TI - Renal allograft rejection with normal renal function in simultaneous kidney/pancreas recipients: does dissynchronous rejection really exist? AB - BACKGROUND: Between July 1, 1994 and December 1, 1998, 147 simultaneous kidney/pancreas transplantations were performed at our center. Of 95 patients who experienced at least one acute renal allograft rejection episode after transplantation, 7 (7.4%) developed rejection in the presence of stable and normal or near-normal renal function. METHODS: The indication for renal allograft biopsy was a rising serum lipase, i.e., suspected pancreatic rejection. All seven patients were treated with steroids and augmentation of the tacrolimus dose, with a fall in the serum lipase and no change in the serum creatinine. RESULTS: The serum creatinine levels just before, at the time of, 1 week after the biopsy, and at most recent follow-up were 1.4+/-0.4, 1.3+/-0.3, 1.2+/-0.2, and 1.2+/-0.2 mg/dl. The serum lipase levels just before, at the time of, 1 week after the biopsy, and at most recent follow-up were 1022+/-1157 mg/dl, 874+/-996 mg/dl, 243+/-260 mg/dl, and 94+/-75 mg/dl. The tacrolimus dosages and levels at the time of the biopsy and 1 week later were 14.9+/-5.0 mg/day and 15.0+/-4.0 ng/ml, and 16.4+/-6.3 mg/day and 15.1+/-6.8 ng/ml. CONCLUSIONS: These findings suggest that, in patients undergoing simultaneous kidney/pancreas transplantation, the entity of dissynchronous pancreatic allograft rejection without renal allograft rejection may not really exist. These data also make an additional fundamental point that acute rejection may occur in patients with normal and stable renal function. PMID- 10706059 TI - Cardiac allograft vascular disease after orthotopic heart transplantation: methylenetetrahydrofolate reductase gene polymorphism C677T does not account for rapidly progressive forms. AB - BACKGROUND: Recently, homocysteine (HCY) levels have been suggested to be a risk factor in cardiac allograft vascular disease (CAVD). As plasma levels are partially under genetic control, we investigated the influence of the methylenetetrahydrofolate reductase (MTHFR) polymorphism on HCY levels and development of CAVD in heart transplant (HTX) recipients. METHODS: Genotyping and assessment of fasting HCY levels were performed in a cohort of 146 HTX recipients and correlated to the onset and progression of CAVD, assessed by serial angiography. RESULTS: Actuarial freedom from CAVD did not differ significantly between the genotypes. However, patients positive for CAVD presented with higher HCY levels than CAVD-negative individuals (21.0+/-9.4 vs. 18.2+/-6.6 micromol/L, P=0.046). CONCLUSIONS: There is some evidence that plasma HCY might be involved in development of CAVD. However, polymorphism of the MTHFR gene could not be shown to be related to severity of allograft vascular disease. PMID- 10706060 TI - Graft-versus-host-disease-associated thymic damage results in the appearance of T cell clones with anti-host reactivity. AB - BACKGROUND: We studied whether T-cell clones, which appear in the periphery as a result of the failure of thymic negative selection during graft-versus-host disease (GVHD), have any in vivo anti-host reactivity and can cause GVHD in an adoptive transfer model. METHODS: We performed our studies in a murine model (B10.BR into CBA/J) for allogeneic bone marrow transplantation with major histocompatibility complex-matched and minor histocompatibility antigen mismatched unrelated donors and unique Vbeta T-cell deletion patterns in donors and recipients. RESULTS: GVHD resulted in the appearance of Vbeta6+ T cells as a result of a loss of negative selection. We found that Vbeta6+ T cells from normal donors proliferated in vitro and in vivo. Depletion of Vbeta6+ T cells from the donor T-cell inoculum resulted in less GVHD morbidity and a decrease in the loss of thymic cellularity. To test the anti-host reactivity of de novo generated Vbeta6+ T cells in animals with GVHD, we developed an adoptive transfer model of splenic T cells from CBA/J host animals with GVHD into sublethally irradiated CBA/J recipients Depletion of Vbeta6+ T cells from the splenic T cells before adoptive transfer could significantly decrease the transient GVHD morbidity in the sublethally irradiated hosts. CONCLUSIONS: Our data indicate that GVHD associated thymic damage results in a loss of thymic negative selection, which leads to the appearance of T-cell clones with anti-host reactivity in vitro and in vivo. PMID- 10706062 TI - A closed door opens: organ transplantation from brain dead donors. PMID- 10706061 TI - Cytotoxic T lymphocyte antigen 4-immunoglobulin fusion protein combined with methotrexate/cyclosporine as graft-versus-host disease prevention in a canine dog leukocyte antigen-nonidentical marrow transplant model. AB - BACKGROUND: We studied whether blocking of the T cell costimulatory signal from B7-->CD28 by cytotoxic T lymphocyte antigen 4-immunoglobulin fusion protein would, either by itself or when added to methotrexate/cyclosporine, result in improved graft-versus-host disease prevention after dog leukocyte antigen nonidentical canine hematopoietic stem cell transplantation after 920 cGy total body irradiation. RESULTS AND CONCLUSIONS: Survivals of cytotoxic T lymphocyte antigen 4-immunoglobulin fusion protein-treated dogs were only slightly prolonged over controls. It appeared that the addition of cytotoxic T lymphocyte antigen 4 immunoglobulin fusion protein failed to induce graft-host tolerance in this model beyond that achieved with methotrexate/cyclosporine alone. PMID- 10706063 TI - Allorecognition of artificial nerve guides filed with human Schwann cells: an in vitro piloot study. PMID- 10706064 TI - Carotid atherosclerosis in renal transplant recipients. PMID- 10706065 TI - Hepatitis B, the hidden danger in cadaveric organ donors. PMID- 10706066 TI - Renal transplantation without a pre-transplant crossmatch. PMID- 10706067 TI - Transplantation tolerance induced by "mega dose" CD34+ cell transplants. AB - Early studies in murine models and more recent clinical data in heavily pretreated leukemia patients have shown that escalation of hematopoietic progenitor cells can overcome major genetic barriers and enable rapid and durable engraftment of haploidentical 3-loci mismatched transplants without graft-versus host disease. In vitro studies suggest that veto cells within the progenitors population most likely mediate this facilitating effect. Leukemia relapse is relatively low in patients with acute myeloid leukemia (AML) but is greater in adults with acute lymphoblastic leukemia (ALL). Donor NK cells most likely mediate the resistance to relapse in patients with AML who are recipients of haploidentical transplants. Immune reconstitution in adults but not in children is slow as in adult recipients of HLA matched unrelated bone marrow transplants. The "mega dose" concept was also shown recently to be useful for tolerance induction in sublethally irradiated mice, so as to effectively overcome the marked resistance presented by the large number of lymphocytes surviving the sublethal conditioning. Thus, allogeneic chimeras generated by transplantation of large doses of Sca1+Lin- cells, permanently accept allogeneic donor type skin grafts. However, the numbers required to attain this desirable goal may not be easily collected from human donors. Nonalloreactive T cells synergize with murine Sca1+Lin- cells and might, therefore, enable achievement of engraftment of haploidentical transplants in sublethally conditioned patients. PMID- 10706068 TI - Modulation of hematopoietic stem/progenitor cell engraftment by transforming growth factor beta. AB - OBJECTIVE: To investigate if cell cycle progression plays a role in modulating the engraftment potential of mouse hematopoietic stem and progenitor cells (HSPC). MATERIALS AND METHODS: HSPC were isolated from adult mouse bone marrow, cultured in vitro under conditions promoting cell cycle arrest, and subsequently were evaluated for cell cycle status, clonogenic activity, and transplant potential. RESULTS In the presence of steel factor (STL) as a survival cytokine, transforming growth factor beta (TGF-beta) increased the G0/G1 fraction of cycling progenitor cells (Rh(high)) after a 20-hour culture. Clonogenic activity of quiescent long-term repopulating (Rh(low)) HSPC was unaffected by this culture, whereas clonogenic potential of Rh(high) cells decreased by about 30%. In competitive repopulation assays, Rh(low) cells cultured in STL + TGF-beta engrafted better than cells cultured in STL alone. However, culture in STL + TGF beta did not overcome the failure of Rh(high) cells to engraft after transplant. We also utilized a two-stage culture system to first induce proliferation of Rh(low) HSPC by a 48-hour culture in STL + interleukin 6 + Flt-3 ligand, followed by shifting the culture to STL + TGF-beta for 24 hours to induce cycle arrest. A competitive repopulation assay demonstrated a relative decrease in repopulating potential in cultures that were cycle arrested compared to those that were not. CONCLUSION: Cell cycle progression by itself cannot account for the decrease in repopulating potential that is observed after ex vivo expansion. Other determinants of engraftment must be identified to facilitate the transplantation of cultured HSPC. PMID- 10706069 TI - Analysis of the surface expression of c-kit and occurrence of the c-kit Asp816Val activating mutation in T cells, B cells, and myelomonocytic cells in patients with mastocytosis. AB - OBJECTIVE: The Asp816Val c-kit activating mutation is detectable in the peripheral blood cells of some patients with mastocytosis and in lesional skin biopsies obtained from adult patients with urticaria pigmentosa. These observations led to the conclusion that this mutation is present in mast cells and mast cell precursors that express c-kit. However, the distribution of the Asp816Val mutation among hematopoietic lineages is unknown. To determine the distribution of the Asp816Val mutation among hematopoietic lineages and to explore its relationship to clinical disease, we examined cells bearing differentiation markers for myelomonocytic cells as well as T and B lymphocytes, in both peripheral blood and bone marrow obtained from patients with mastocytosis. MATERIALS AND METHODS: The presence of Asp816Val c-kit mutation in cells magnetically sorted from peripheral blood or bone marrow according to surface differentiation markers was studied by reverse transcriptase polymerase chain reaction (RT-PCR) restriction fragment length polymorphism (RFLP) analysis. The surface expression of c-kit was determined by flow cytometry. RESULTS: The mutation was detectable by RT-PCR in at least one cell lineage in the bone marrow in 7 of 7 patients examined and in the peripheral blood of 11 of 11 adult patients with urticaria pigmentosa and indolent disease. The mutation was identified most frequently in B cells and myeloid cells. Flow cytometric analysis demonstrated that the differentiated cells expressing mutated c-kit were negative for surface KIT. CONCLUSION: These results are consistent with the conclusion that the c-kit Asp816Val mutation occurs in an early progenitor cell and is carried by myelomonocytic cells, T cells, and B cells in addition to mast cells. However, unlike mast cells, these myelomonocytic cells, T cells, and B cells do not concomitantly express surface c-kit and thus may be less susceptible to the effects of this mutation. PMID- 10706070 TI - Establishment of stromal cell line from an MDS RA patient which induced an apoptotic change in hematopoietic and leukemic cells in vitro. AB - OBJECTIVE: We previously reported on the heterogeneity of bone marrow stromal cell function in supporting hematopoietic cell proliferation and differentiation in vitro among refractory anemia (RA) of myelodysplastic syndrome (MDS) patients. Interestingly, stromal cells from some MDS RA patients induced an apoptotic change in CD34+ hematopoietic cells. However, the mechanism responsible for this action was unclear. MATERIALS AND METHODS: In the present study, we established a cloned stromal cell line (LS801) from an MDS RA patient by introducing recombinant SV40-adenovirus vector containing the SV40 early gene. RESULTS: LS801 induced an apoptotic change in CD34+ cells from normal subjects and cloned leukemic cells in a coculture system. When hematopoietic cells were cocultured but kept separate from LS801 by a 0.45-microm Millipore membrane to prevent their attachment, the action of LS801 in inducing apoptosis of hematopoietic cells was inhibited. Additionally, no production of fas ligand, tumor necrosis factor alpha or interferon gamma in LS801 was observed. CONCLUSION: These findings suggest that the novel stromal cell line LS801 will shed light on research into the mechanism underlying the apoptotic changes induced by stromal cells in hematopoietic cells. PMID- 10706071 TI - Replication status as a marker for predisposition for lymphoma in patients with chronic hepatitis C with and without cryoglobulinemia. AB - OBJECTIVE: Essential mixed cryoglobulinemia (EMC) type II is associated with hepatitis C virus (HCV) in 90% of the patients with this disorder. A significant subset of these patients is at risk to develop non-Hodgkin lymphoma (NHL). The objective of this study was to examine whether the presence of EMC, a presumably premalignant step of lymphoproliferation, is associated with changes in the replication state of normal structural genes. MATERIALS AND METHODS: The study group included three subgroups: (1) seven patients with HCV without EMC; (2) eight patients with HCV associated with EMC. 3. Seven patients with follicular lymphoma; and (3) six healthy individuals served as control group. Monocolor fluorescent in situ hybridization (FISH) with probes to p53, RB-1, and 21q22 was applied to leukocytes nuclei for the evaluation of replication timing. RESULTS: Asynchronous replication (SD) rate was similar in patients with NHL and those with HCV associated with EMC and both are significantly higher when compared to patients with HCV without EMC and to normal controls (p < 0.01) for each comparison. This held true for all studied loci (21q22, RB-1, and p53). Patients infected by HCV (but without EMC) had a significantly higher rate of asynchronous pattern in comparison with healthy controls (p < 0.01). CONCLUSIONS: Patients with a "premalignant" clinical condition HCV with EMC already demonstrate asynchronous type of replication which is similar to patients who already have an established malignant disease (i.e., NHL). In the future, replication may be used to assess the risk of malignant transformation in patients with "benign" proliferation. PMID- 10706072 TI - Expression and induction of costimulatory and adhesion molecules on acute myeloid leukemic cells: implications for adoptive immunotherapy. AB - OBJECTIVE: Previously, we observed an increased recognition of malignant cells by cytotoxic T lymphocytes (CTL) when the target cells were cultured in vitro for 24 hours. In this study, we analyzed the expression of costimulatory and adhesion molecules on acute myeloid leukemia (AML) cells and determined whether 24-hour culture of the cells was associated with upregulation of these molecules. We analyzed whether this incubation period improved recognition of AML cells by CTL. MATERIALS AND METHODS: Expression of costimulatory and adhesion molecules on leukemic blasts of 34 patients comprising each AML FAB subclassification were analyzed directly and after 24 hours of culture, and the recognition of these AML cells by an HLA-A2 restricted CTL clone was determined. Blocking studies were performed with antibodies against CD54, CD58, and CD11a. RESULTS: Immunophenotyping showed a low expression of CD80 and CD40 and a variable CD86 expression on most AML cells. CD54 expression was generally low, CD58 expression was high, and CD11a expression was variable, with a higher expression in AML M0, M1, M4, and M5. Twenty-four hours of culture resulted in a significant upregulation of CD40, CD54, and CD58. Impaired recognition of AML cells by the HLA-A2 restricted CTL clone was enhanced 100-200% by 24 hours of preincubation of the leukemic cells. Blocking studies showed the importance of multiple adhesion molecules on the AML cells. CONCLUSION: Low expression of multiple costimulatory and adhesion molecules on AML could be upregulated by 24 hours of culture, which was associated with increased recognition of the AML blasts by CTL. Blocking multiple adhesion molecules completely abolished CTL recognition, showing the importance of the combination of these molecules for T-cell interaction with AML. PMID- 10706073 TI - The benzene metabolite, hydroquinone, selectively induces 5q31- and -7 in human CD34+CD19- bone marrow cells. AB - OBJECTIVE: Chronic exposure to high concentrations of benzene is associated with an increased incidence of myelodysplastic syndrome and acute myelogenous leukemia. Acute myelogenous leukemia developing in patients treated with alkylating agents for other cancers or occupationally exposed to benzene exhibit a pattern of cytogenetic aberrations predominantly involving loss of all or part of chromosomes 5 and/or 7. In contrast, trisomy 8 is observed equally in both de novo and secondary acute myelogenous leukemia. Studies using peripheral lymphocytes or lymphoblastoid cell lines have observed dose-dependent loss of chromosomes 5, 7, and 8 following treatment with the benzene metabolite, hydroquinone. The purpose of this study was to determine the dose response and specificity of hydroquinone-induced aberrations on chromosomes 5, 7, and 8 using human CD34+CD19 bone marrow cells. MATERIALS AND METHODS: Fluorescence in situ hybridization analysis was performed on CD34+CD19- bone marrow cells using the locus-specific probes, 5q31, 5p15.2, and centromeric probes specific for human chromosomes 7 and 8 following hydroquinone exposure. RESULTS: Hydroquinone exposure results in -7, selective deletion of 5q31 but not chromosome 5 and no loss or gain of chromosome 8 in human CD34+CD19- cells. CONCLUSION: CD34+ bone marrow cells are more susceptible and show a different pattern of cytogenetic aberrations as a result of hydroquinone exposure compared to lymphocytes. CD34+ bone marrow cells exhibit unique susceptibility to the development of specific chromosome aberrations that have been identified as the earliest structural changes occurring in the development of secondary myelodysplastic syndrome and acute myelogenous leukemia. PMID- 10706074 TI - Human adult tonsil xenotransplantation into SCID mice for studying human immune responses and B cell lymphomagenesis. AB - OBJECTIVE: To generate a human-mouse xenochimeric model where human cells remain clustered in the animal to optimize their interactions and recovery. MATERIALS AND METHODS: Severe combined immune deficient mice (SCID) were xenografted subcutaneously with human adult tonsil pieces (hu-ton-SCID mice). Such animals were: (a) compared with those receiving tonsil cells in suspension, and (b) immunized with de novo and recall antigens. RESULTS: Human tonsil pieces survived a long period of time in SCID mice, while polyclonal human T- and B-lymphocytes persisted in close vicinity within the implantation area; however, little or no graft-versus-host disease was detectable. Not surprisingly, local development of lymphoproliferative disease was often observed in animals receiving lymphoid implants from donors previously infected by the Epstein-Barr virus. One month after surgery, higher serum levels of human IgG were found in SCID mice transplanted with tonsil pieces (2x10(7) cells/animal) than in animals injected with 5x10(7) tonsil cells in suspension (1.9 vs. 0.3 mg/mL, p < 0.002). Importantly, the production of human IgG in hu-ton-SCID mice remained polyclonal for at least 6 months and was linked to the presence of cells within the implants. Immunization of hu-ton-SCID mice with hepatitis B core, a de novo antigen, did not produce a significant IgG immune response; however immunization with tetanus toxoid (TT), a thymus-dependent recall antigen, yielded high (> 700 fold increase in anti-TT IgG levels) and long-lasting (> 6 months) secondary immune responses. CONCLUSION: The hu-ton-SCID mouse xenochimeric model described in this report may improve our understanding of human lymphoid cell interactions, secondary immune responses, and lymphomagenesis. PMID- 10706075 TI - Proliferation of dendritic cell progenitors in long term culture is not dependent on granulocyte macrophage-colony stimulating factor. AB - A unique long term culture (LTC) system has been developed which supports the production of dendritic cells (DC). Cell production is dependent on a stromal cell layer derived from murine spleen. This LTC system produces a high turnover of non-adherent cells that express DC morphology, cell-surface markers, and antigen-presenting capacity. OBJECTIVE: The long term production of these cells suggests that the LTC system supports hemopoiesis. It was of interest to examine the number and nature of hemopoietic progenitors present in LTC. MATERIALS AND METHODS: A combination of approaches, including FACS analysis, spleen colony forming unit assays, and in vitro colony assays were undertaken. RESULTS: Pluripotent haemopoietic stem cells are not detectable among the non-adherent cell population produced in LTC. Instead, LTC support a replicating c-kit+ progenitor population, which generates only dendritic-like colonies in in vitro colony assays. In addition, this population does not respond to combinations of growth factors thought to stimulate DC proliferation, including granulocyte macrophage-colony stimulating factor (GM-CSF) and Flt3L. Production of DC occurs only in the presence of LTC-derived culture supernatant or a confluent stromal cell layer. CONCLUSIONS: These results suggest that LTC contain a dendritic progenitor that is dependent upon the stromal cell network for proliferation and differentiation. The development of only DC within LTC allows easy collection of cells for experimentation. This, in combination with the fact that DC development occurs in the absence of exogenous growth factors, makes the LTC system a practical model for the study of DC function and development. PMID- 10706076 TI - Thrombopoietin can influence mature megakaryocytes to undergo further nuclear and cytoplasmic maturation. AB - OBJECTIVE: We studied the effects of TPO on the nuclear and cytoplasmic maturation of mature megakaryocytes. MATERIALS AND METHODS: We prepared mature megakaryocytes by velocity sedimentation method and investigated the maturational effect of TPO on such mature megakaryocytes from the aspects of DNA ploidy and cytoplasmic features using a flow cytometry and Br-dU incorporation. We also studied the effects of TPO on expression of GpIIb and NF-E2 transcripts by RT-PCR and on proplatelet formation. RESULTS: DNA content increased when megakaryocytes were cultured for 2 days with TPO, resulting in generation of megakaryocytes with a DNA content of 32N or 64N. SCF had a similar, but weaker, effect, while IL-6 did not influence mature megakaryocytes. The proportion of megakaryocytes incorporating Br-dU into their nuclei was 44.3% +/- 6.9% after culture with TPO, 19.5% +/- 4.6% with SCF, 7.5% +/- 3.5% with IL-6, and 5.2% +/- 2.7% in control culture. Flow cytometry showed that TPO did not generate larger megakaryocytes with more complex intracellular structures when compared with cells of the same DNA content class (16N) cultured without TPO. In addition, TPO also did not enhance the expression of NF-E2 transcripts, but it delayed proplatelet formation by cultured megakaryocytes. CONCLUSION: TPO first affected endomitosis by mature megakaryocytes and then altered their cytoplasmic maturation, with both maturation processes remaining proportionate to each other. PMID- 10706077 TI - Autografting with Ph-negative progenitors in patients at diagnosis of chronic myeloid leukemia induces a prolonged prevalence of Ph-negative hemopoiesis. AB - OBJECTIVE: In many patients with chronic myeloid leukemia (CML), a residual population of primitive normal (Ph-negative) progenitors persists despite the marked expansion of the leukemic (Ph-positive) clone. These cells may be found in the blood of patients studied soon after diagnosis or during the period of endogenous hematopoietic recovery that follows myeloreductive therapy. Based on those observations, we have developed a clinical protocol that allows collection of Ph-negative peripheral blood progenitor cells (PBPC) with transplantable hematopoietic regenerative potential. The aim of this study is to examine changes that occur in the percentage of Ph-negative- and Ph-positive-committed progenitor cells and to determine the relationship between changes and clinical outcome. MATERIALS AND METHODS: We followed 15 patients with CML, mobilized and autografted soon after diagnosis with 85%-100% Ph-negative PBPC for a median time of 28 months (range 18-50) after transplant. At 6 months, 1 year, 2 years, and last follow-up, cytogenetic analyses were performed on fresh bone marrow cells and on colony-forming cells (CFC). RESULTS: Autologous transplantation induces a reduction in the proportion of Ph-positive CFC, from 70%-100% to 0%-25% in the majority of patients (78%). After autografting, 8 of 15 patients achieved a long lasting cytogenetic remission (median, 24 months; range, 21-43) with a Ph positivity ranging between 0% and 20% at the level of mature mononuclear cells and colony-forming cells (CFC). In some patients, the majority of CFC remained Ph negative, whereas the majority of the mature cells were Ph-positive. Other patients (5/15) developed cytogenetic relapse (100% Ph-positive), although they were in hematological remission. We found that detection of Ph-positive long-term culture initiating cells (LTC-IC) in the marrow at diagnosis was the only factor significantly associated with recurrence of the disease (p < 0.01); on the other hand, the number of Ph-negative LTC-IC infused showed a significant correlation with a better outcome (p < 0.03). CONCLUSION: We have shown that a prolonged period of complete or almost complete Ph-negative hemopoiesis can be achieved in patients with CML who undergo autografting with Ph-negative progenitors. Longer follow-up study will be needed to assess whether these changes are associated with improved survival. PMID- 10706078 TI - Peripheral blood progenitor cell mobilization in healthy donors receiving recombinant human granulocyte colony-stimulating factor. AB - OBJECTIVE: We analyzed the incidence of primitive (LTC-IC) and committed (CFU mix, BFU-E, CFU-GM) hematopoietic progenitors detected under steady-state conditions and upon progenitor cell mobilization in a cohort of healthy donors receiving recombinant human granulocyte colony-stimulating factor (rhG-CSF). MATERIALS AND METHODS: Healthy donors (n = 30) of HLA-mismatched or -matched stem cell transplants were mobilized with rhG-CSF (8 microg/Kg body weight subcutaneously twice daily until completion of leukapheresis). PBPC collections were started after 4 days of rhG-CSF therapy. RESULTS: Steady-state incidence of bone marrow LTC-IC, but not committed progenitors, significantly correlated with the numbers of mobilized CD34+ cells (r = 0.6, p = 0.004), CFU-GM (r = 0.79, p = 0.0005) and CFC (r = 0.76, p = 0.001) detected after 4 days of rhG-CSF therapy. Statistically significant correlations were also found between steady-state blood CFU-GM and peak numbers of CD341 cells (r = 0.68, p = 0.001), numbers of day 4 CD341 cells (r = 0.52, p = 0.005), CFU-GM (r = 0.63, p = 0.002), and CFC (r = 0.61, p = 0.003). CONCLUSION: Our data show that in normal volunteers baseline marrow LTC-IC and blood CFU-GM correlate with rhG-CSF-mobilized PBPC. The potential clinical relevance of these findings in the identification of poor mobilizers will be tested in a prospective study. PMID- 10706079 TI - A calcitriol analogue, EB1089, inhibits the growth of LNCaP tumors in nude mice. AB - Limited options for the treatment of prostate cancer have spurred the search for new therapies. One innovative approach is the use of 1alpha,25-dihydroxyvitamin D3 (calcitriol) analogues to inhibit cancer growth. We demonstrate here that the calcitriol analogue, EB1089, extensively inhibits the growth of LNCaP prostate cancer cells in culture and causes the cells to both accumulate in G0-G1 and undergo apoptosis. Importantly, we found that EB1089 inhibits the growth of LNCaP tumor xenografts in nude mice. Because of these antiproliferative properties in vivo, EB1089 is a potential new therapeutic agent for the treatment of prostate cancer. PMID- 10706080 TI - Osteoprotegerin prevents and reverses hypercalcemia in a murine model of humoral hypercalcemia of malignancy. AB - Osteoprotegerin (OPG), a novel, secreted tumor necrosis factor receptor family member that inhibits osteoclast formation and activity was examined for its activity in a syngeneic tumor model of humoral hypercalcemia of malignancy. Normal mice bearing Colon-26 tumors develop increases in both parathyroid hormone related protein (PTHrP) expression and plasma PTHrP, marked hypercalcemia, and increased bone resorption. OPG, given either at the onset of hypercalcemia or after it had occurred, blocked tumor-induced increases in bone resorption and hypercalcemia and rapidly normalized blood ionized calcium. In tumor-bearing mice, OPG treatments reduced osteoclast activity from approximately 2-fold above normal into the subphysiological range but had no effects on tumor size, tumor induced cachexia, or PTHrP levels. The potent effects of OPG in this humoral hypercalcemia of malignancy model suggest a potential therapeutic role for OPG in the prevention and treatment of this disorder. PMID- 10706081 TI - Adenoviral Bak overexpression mediates caspase-dependent tumor killing. AB - One of the most promising strategies in cancer gene therapy is adenoviral transfer of proapoptotic genes. We therefore evaluated the novel strategy of adenovirally overexpressing the proapoptotic Bak gene. Our results showed marked apoptosis in cancer cells in vivo and in vitro after Bak gene transfer via a binary adenoviral vector system. This effect was not seen in a caspase 3 defective cell line (MCF-7) and was abrogated in Bak-sensitive tumors after administration of the caspase inhibitor z-DEVD-fmk. Our results suggest that adenoviral-mediated overexpression of Bak provides a novel therapeutic strategy for cancer therapy, but this process appears to be caspase dependent. PMID- 10706082 TI - A new variant anaplastic lymphoma kinase (ALK)-fusion protein (ATIC-ALK) in a case of ALK-positive anaplastic large cell lymphoma. AB - Anaplastic lymphoma kinase (ALK)-positive lymphomas ("ALKomas") constitute a distinct molecular and clinicopathological entity within the heterogeneous group of CD30-positive large cell lymphomas. In 80-85% of cases tumor cells express a Mr 80,000 hybrid protein comprising the nucleolar phosphoprotein nucleophosmin (NPM) and the ALK. We report here the cloning and expression of a novel ALK fusion protein from an ALK-positive lymphoma. This case was selected for molecular investigation because of (a) the absence of NPM-ALK transcripts; (b) the atypical staining patterns for ALK (cytoplasm-restricted) and for NPM (nucleus-restricted); and (c) the presence of a Mr 96,000 ALK-protein differing in size from NPM-ALK. Nucleotide sequence analysis of ALK transcripts isolated by 5'-rapid amplification of cDNA ends revealed a chimeric mRNA corresponding to an ATIC-ALK in-frame fusion. ATIC is a bifunctional enzyme (5-aminoimidazole-4 carboxamide ribonucleotide transformylase and IMP cyclohydrolase enzymatic activities) that catalyzes the penultimate and final enzymatic activities of the purine nucleotide synthesis pathway. Expression of full-length ATIC-ALK cDNA in mouse fibroblasts revealed that the fusion protein (a) possesses constitutive tyrosine kinase activity; (b) forms stable complexes with the signaling proteins Grb2 and Shc; (c) induces tyrosine-phosphorylation of Shc; and (d) provokes oncogenic transformation. These findings point to fusion with ATIC as an alternative mechanism of ALK activation. PMID- 10706083 TI - Detection of gene amplification by genomic hybridization to cDNA microarrays. AB - Gene amplification is one of the major mechanisms of oncogene activation in tumorigenesis. To facilitate the identification of genes mapping to amplified regions, we have used a technique based on the hybridization of total genomic DNA to cDNA microarrays. To aid detection of the weak signals generated in this complex hybridization, we have used a tyramide-based technique that allows amplification of a fluorescent signal up to 1000-fold. Dilution experiment suggests that amplifications of 5-fold and higher can be detected by this approach. The technique was validated using cancer cell lines with several known gene amplifications, such as those affecting MYC, MYCN, ERBB2, and CDK4. In addition to the detection of the known amplifications, we identified a novel amplified gene, ZNF133, in the neuroblastoma cell line NGP. Hybridization of NGP cDNA on an identical array also revealed over expression of ZNF133. Parallel analysis of genomic DNA for copy number and cDNA for expression now provides rapid approach to the identification of amplified genes and chromosomal regions in tumor cells. PMID- 10706084 TI - The DNA mismatch repair genes Msh3 and Msh6 cooperate in intestinal tumor suppression. AB - Repair of mismatches in DNA in mammalian cells is mediated by a complex of proteins that are members of two highly conserved families of genes referred to as MutS and MutL homologues. Germline mutations in several members of these families, MSH2, MSH6, MLH1, and PMS2, but not MSH3, are responsible for hereditary non-polyposis colorectal cancer. To examine the role of MSH3, we generated a mouse with a null mutation in this gene. Cells from Msh3-/- mice are defective in repair of insertion/ deletion mismatches but can repair base-base mismatches. Msh3-/- mice develop tumors at a late age. When the Msh3-/- and Msh6 /- mutations are combined, the tumor predisposition phenotype is indistinguishable from Msh2-/- or Mlh1-/- mice. These results suggest that MSH3 cooperates with MSH6 in tumor suppression. PMID- 10706085 TI - Expression of the Wilms' tumor suppressor gene, WT1, reduces the tumorigenicity of the leukemic cell line M1 in C.B-17 scid/scid mice. AB - The Wilms' tumor suppressor gene, WT1, encodes a transcription of the Cys2-His2 zinc finger type. Loss of WT1 gene function has been implicated in the development of malignancies including Wilms' tumor and acute leukemias. We have shown previously that ectopic expression of WT1 +KTS isoforms in murine M1 leukemic cells spontaneously induces monocytic differentiation without the requirement for external differentiation-inducing stimuli. To determine whether these observed effects in vitro corresponded to a reduction in tumorigenicity in vivo, parental M1, control M1.Neo, and M1.WT1 +KTS cells were transplanted into C.B-17 scid/scid mice, and the growth and metastatic behavior of the cell lines were monitored for a period of 20 weeks. Mice inoculated either s.c. on the flank or directly into the peritoneal cavity, with M1 cells stably expressing WT1 +KTS isoforms exhibited a marked decrease in tumor formation compared with control groups. Moreover, tumors arising in mice after the injection of M1.WT1 +KTS cells exhibited a loss in ectopic WT1 protein expression. Confirmation that the tumors arose from M1.WT1 +KTS cells was achieved by the amplification of the introduced transgene from tumor samples and indicates that the tumorigenicity of leukemic M1 cells in these animals correlates with a loss in WT1 expression. This investigation is the first to demonstrate the tumor-suppressive effects of WT1 expression in a leukemic cell line, further advancing the notion that WT1 acts as a differentiation-promoting gene during hematopoiesis and that loss of functional WT1 expression may contribute to leukemogenesis in vivo. PMID- 10706086 TI - Enhanced nuclear diacylglycerol kinase activity in response to a mitogenic stimulation of quiescent Swiss 3T3 cells with insulin-like growth factor I. AB - Results from several laboratories have established the existence in the nucleus of an autonomous polyphosphoinositide cycle, which is involved in both cell proliferation and differentiation. A key step of intranuclear polyphosphoinositide metabolism is the phospholipase C-mediated generation of diacylglycerol (DAG). In insulin-like growth factor (IGF)-I-stimulated Swiss 3T3 cells, a transient elevation of intranuclear DAG levels is essential for attracting the alpha isoform of protein kinase C (PKC) to the nucleus. Previous evidence has shown that the nucleus also contains DAG kinase, i.e., the enzyme that yields phosphatidic acid from DAG, thus terminating PKC-mediated signaling events. Here we show that IGF-I treatment of quiescent Swiss 3T3 cells results in the stimulation of nuclear DAG kinase activity. Time course analysis showed an inverse relationship between nuclear DAG mass and DAG kinase activity levels. After IGF-I treatment, maximal enhancement of DAG kinase activity was measured in the internal matrix domain of the nucleus. PKC-alpha remained within the nuclear compartment, even when nuclear DAG mass returned to basal levels. This was conceivably due to interactions with specific nuclear PKC-binding proteins, some of which were identified as lamins A, B, and C and protein C23/nucleolin. Treatment of cells with two DAG kinase inhibitors, R59022 and R59949, blocked the IGF-I-dependent rise in nuclear DAG kinase activity and maintained elevated intranuclear levels of DAG. The two inhibitors also markedly potentiated the mitogenic effect of IGF-I. These results suggest that nuclear DAG kinase plays a key role in regulating the levels of DAG present in the nucleus and that DAG is a key molecule for the mitogenic effect that IGF-I exerts on Swiss 3T3 cells. PMID- 10706087 TI - FasL:Fas ratio--a prognostic factor in breast carcinomas. AB - Programmed cell death (apoptosis) is primarily mediated by Fas ligand (FasL; CD95L) and the Fas receptor (Fas; CD95). In this study, FasL was detected by immunohistochemical analysis in 85% of breast carcinomas and 14% of fibroadenomas randomly chosen, indicating that high expression of FasL might play a role in tumor pathology. FasL and Fas levels as well as FasL:Fas ratios were further ascertained in 215 human breast tumors, including 199 invasive ductal carcinomas, by real-time quantitative reverse transcription-PCR and compared with expression levels and ratios found in 25 normal human tissues, in 37 fibroadenomas, and in 5 normal breast tissues. Among breast carcinomas, high FasL mRNA expression seems to be positively correlated with histological grading (n = 212; P<0.0001). A ratio of FasL:Fas mRNA transcripts >1 is found to be significantly associated with decreased patient's disease-free survival (n = 211; P<0.03) and increased mortality (n = 211; P = 0.19). A FasL:Fas ratio >1 is related to tumor progression scored by histological grading (n = 212; P<0.02). The selection process leading to highly aggressive breast tumor variants might be enhanced by FasL-mediated tumor fratricide, eventually a possible target for novel therapeutic strategies. PMID- 10706088 TI - Dendritic cell-based immunotherapy of prostate cancer: immune monitoring of a phase II clinical trial. AB - We assessed both non- and peptide-specific immune responses in prostate cancer patients before and after immunotherapy with dendritic cells exogenously pulsed with the prostate-specific membrane antigen-derived peptides, PSM-P1 and PSM-P2. For all subjects, we observed that clinical responses were strongly associated with two indicators of immunocompetence: skin test responses to recall antigens and cytokine secretion by T cells after nonspecific stimulation. In a subset of responders, we observed cytokine secretion or cytotoxicity against the immunizing peptides or an immunodominant epitope from an influenza recall antigen. The clinical results support the use of monitoring for overall immunocompetence to help determine why a patient has or has not responded to therapy. Moreover, it could be useful as an inclusion criterion to select those more likely to benefit from treatment. PMID- 10706089 TI - Overexpression of insulin-like growth factor-binding protein-2 results in increased tumorigenic potential in Y-1 adrenocortical tumor cells. AB - Increased concentrations of insulin-like growth factor-binding protein-2 (IGFBP 2) have been observed in human malignancies including adrenocortical carcinomas. To elucidate the functional consequences of IGFBP-2 overexpression, we have stably transfected the cDNA of murine IGFBP-2 in mouse adrenocortical tumor cells (Y-1). Long-term overexpression of IGFBP-2 was associated with significant morphological alterations, enhanced cell proliferation, and increased cloning efficiency as compared with mock transfected control cells. The enhanced proliferation of IGFBP-2 secreting clones was independent of exogenous insulin like growth factors (IGFs). These data suggest that elevated levels of IGFBP-2 may contribute to the highly malignant phenotype of adrenocortical cancer by a thus far unknown, presumably IGF-independent, mechanism. PMID- 10706090 TI - Susceptibility to prostate cancer: interaction between genotypes at the androgen receptor and prostate-specific antigen loci. AB - The androgen receptor (AR) regulates gene transcription by binding to androgen response elements in target gene promoters. The prostate-specific antigen (PSA) gene has a polymorphic androgen response element sequence with two alleles, A and G. We hypothesize that allelic differences in AR-driven PSA expression may influence prostate cancer risk. To test this hypothesis, we assayed PSA genotype for 57 prostate cancer cases and 156 controls from our previous pilot study in which prostate cancer risk was associated with the AR "CAG-short" genotype. Odds ratios (ORs) were estimated relating prostate cancer risk to AR and PSA genotypes, singly and in combination. Subjects with the PSA GG genotype were at significantly increased risk for advanced, but not for localized, prostate cancer (OR, 2.90; 95% confidence interval, 1.24-6.78). When cross-classifying subjects by AR and PSA genotypes, subjects with either a CAG-short allele (and not PSA GG) or with the PSA GG genotype (and not CAG-short) had a modest, statistically insignificant increase in prostate cancer risk overall. However, subjects with both a short CAG allele and PSA genotype GG had a more than 5-fold increase in prostate cancer risk (OR, 5.08; 95% confidence interval, 1.59-16.25). All of the ORs were substantially greater for advanced prostate cancer. Studies with larger numbers of advanced cases will be needed to confirm these results. These results indicate that polymorphism in the PSA gene promoter influences prostate cancer risk, and that the allelic variation in promoter activity may be androgen dependent. Furthermore, these results support a multigenic etiology for prostate cancer. PMID- 10706091 TI - IDN5109, a taxane with oral bioavailability and potent antitumor activity. AB - IDN5109 is a new taxane, derived from 14beta-hydroxy-10-deacetylbaccatin III, selected for its lack of cross-resistance in tumor cell lines expressing the multidrug resistant phenotype. Because, unlike paclitaxel, IDN5109 is a poor substrate for P-glycoprotein, we hypothesized that IDN5109 given p.o. could improve bioavailability compared with paclitaxel. Here, we studied the p.o. and i.v. pharmacokinetics of IDN5109 together with its antitumor activity. Using a high-performance liquid chromatography method, the bioavailability of IDN5109 was determined to be 48% after oral delivery. IDN5109 given p.o. was highly active against the two human ovarian carcinoma xenografts 1A9 and HOC18 (90-100% tumor regressions) and showed significant activity on the paclitaxel-resistant MNB-PTX1 xenograft (10% tumor regressions). The p.o. administration was as active as the i.v. route at doses reflecting the pharmacokinetic data. IDN5109 is the first taxane with good oral bioavailability and potent antitumor activity and represents a potential candidate for clinical investigation. PMID- 10706092 TI - Increased death receptor 5 expression by chemotherapeutic agents in human gliomas causes synergistic cytotoxicity with tumor necrosis factor-related apoptosis inducing ligand in vitro and in vivo. AB - The intractability of malignant gliomas to multimodality treatments plays a large part in their extremely poor prognosis. Tumor necrosis factor-related apoptosis inducing ligand (TRAIL) is a novel member of the tumor necrosis factor (TNF) family that induces apoptosis preferentially in tumor cells through binding to its cognate death receptors, DR4 and DR5. Here we show that the DNA-damaging chemotherapeutic drugs, cis-diamminedichloroplatinum(II) (CDDP) and etoposide, elicited increased expression of DR5 in human glioma cells. Exposure of such cells in vitro to soluble human TRAIL in combination with CDDP or etoposide resulted in synergistic cell death that could be blocked by soluble TRAIL neutralizing DR5-Fc or the caspase inhibitors, Z-Asp-CH2-DCB and CrmA. Moreover, systemic in vivo administration of TRAIL with CDDP synergistically suppressed both tumor formation and growth of established s.c. human glioblastoma xenografts in nude mice by inducing apoptosis without causing significant general toxicity. The combination treatment resulted in complete and durable remission in 29% of mice with the established s.c. xenografts and also significantly extended the survival of mice bearing intracerebral xenografts. These results provide preclinical proof-of-principle for a novel therapeutic strategy in which the death ligand, TRAIL, is safely combined with conventional DNA-damaging chemotherapy. PMID- 10706093 TI - Length and loss of heterozygosity of an intron 1 polymorphic sequence of egfr is related to cytogenetic alterations and epithelial growth factor receptor expression. AB - Overexpression of epithelial growth factor receptor (EGFR) is correlated with a poor prognosis and reduced steroid receptor expression. Recently, it was demonstrated that the length of a CA repeat in the intron 1 of EGFR correlated with the expression of EGFR in vitro. We investigated 112 cases of cancerous and noncancerous breast tumor samples for loss of heterozygosity (LOH) in intron 1 of the egfr gene and determined the intratumoral EGFR content and genetic alterations by comparative genomic hybridization. Heterozygous tumors with short CA repeats showed elevated EGFR expression in contrast to tumors with longer CA repeats. Tumors with LOH in intron 1 of egfr revealed higher EGFR expression when the longer allele was lost compared with loss of the shorter allele. Additionally, tumors with a loss of the long allele showed more chromosomal alterations, especially a higher frequency of amplifications. We conclude that the CA repeat status in intron 1 of the egfr gene also modulates the intratumoral EGFR content in vivo. Furthermore, LOH at the CA repeat is associated with genetically advanced tumors. Therefore, allele-specific gene expression due to LOH of the CA repeat could be assumed to be an important event in invasive breast cancer development. PMID- 10706094 TI - PART-1: a novel human prostate-specific, androgen-regulated gene that maps to chromosome 5q12. AB - Genes regulated by androgenic hormones are of critical importance for the normal physiological function of the human prostate gland, and they contribute to the development and progression of prostate carcinoma. We used cDNA microarrays containing 1500 prostate-derived cDNAs to profile transcripts regulated by androgens in prostate cancer cells. This study identified a novel gene that we have designated PART-1 (prostate androgen-regulated transcript 1), which exhibited increased expression upon exposure to androgens in the LNCaP prostate cancer cell line. Northern analysis demonstrated that PART-1 is highly expressed in the prostate gland relative to other normal human tissues and is expressed as different transcripts using at least three different polyadenylation signals. The PART-1 cDNA and putative protein are not significantly homologous to any sequences in the nonredundant public sequence databases. Cloning and analysis of the putative PART-1 promoter region identified a potential binding site for the homeobox gene PBX-la, but no consensus androgen response element or sterol regulatory element binding sites were identified. We used a radiation hybrid panel and fluorescence in situ hybridization to map the PART-1 gene to chromosome 5q12, a region that has been suggested to harbor a prostate tumor suppressor gene. These results identify a new gene involved in the androgen receptor regulated gene network of the human prostate that may play a role in the etiology of prostate carcinogenesis. PMID- 10706095 TI - Methylation patterns in human androgen receptor gene and clonality analysis. AB - Tumor clonality, an important issue in tumor biology, has been analyzed using X chromosome inactivation studies based on the differential methylation patterns of active and inactive alleles. Recently, a PCR-based androgen receptor gene (AR) analysis method was developed that takes advantage of highly polymorphic CAG repeats and nearby HpaII and HhaI sites in exon 1 of AR at the Xq13 region. However, the data from this assay, which is now widely used, are sometimes uninterpretable and irreproducible for some currently unclear reason. To determine that reason, we analyzed a panel of lung cancer cell lines, using HpaII or HhaI restriction enzymes, methylation-specific PCR, and bisulfite genomic sequencing of the polymorphic CAG repeat site of AR exon 1, including nearby CpG sites. We found direct evidence of a variable methylation pattern at the restriction sites that prevented proper enzyme cleavage in two lung cancer cell lines (NCI-H292 and NCI-H1944) obtained from female patients who had a polymorphic CAG repeat in AR exon 1. Our data suggest that methylation patterns at the CpG sites of AR exon 1 are complicated and vary among different individuals. Therefore, the reliability of the PCR-based clonality analysis may require further evaluation. PMID- 10706096 TI - The glucocorticoid receptor mediates a survival signal in human mammary epithelial cells. AB - Complex autocrine and paracrine signaling pathways control the multiple cycles of epithelial cell proliferation and involution characteristic of the human mammary gland. Activation of these pathways can lead to cell division, cell cycle arrest, apoptosis, or survival; their aberrant regulation often contributes to malignant transformation. In this report, we show that glucocorticoid signals a potent survival pathway in the immortalized human mammary epithelial cell line MCF10A. Withdrawal of glucocorticoid from defined media triggers apoptosis, despite the presence of epidermal growth factor and insulin. Apoptosis is accelerated by ectopic expression of c-Myc and blocked by overexpression of Bcl2. Although MCF10A cells can undergo apoptosis after CD95/Fas receptor activation, cell death caused by glucocorticoid withdrawal is independent of CD95/ Fas receptor signaling. The mechanism through which glucocorticoid inhibits apoptosis is also independent of phosphatidylinositol 3-kinase activity and its downstream target Akt, thus establishing the existence of a novel epithelial cell survival pathway mediated by glucocorticoids. PMID- 10706097 TI - A single nucleotide polymorphism in the E-cadherin gene promoter alters transcriptional activities. AB - E-cadherin plays a critical role in many aspects of cell adhesion, epithelial development, and the establishment and maintenance of epithelial polarity. The loss of the adhesive function of E-cadherin is a critical step in the promotion of epithelial cells to a more malignant phenotype. We identified a C/A single nucleotide polymorphism at -160 from the transcriptional start site of the E cadherin gene promoter. Transient transfection experiments showed that the A allele of this polymorphism decreased the transcriptional efficiency by 68% compared with the C allele (P<0.001). Electrophoretic mobility shift and footprinting assays revealed that the C allele had a stronger transcriptional factor binding strength than the A allele. These results indicate that the -160 C/A polymorphism has a direct effect on E-cadherin gene transcriptional regulation. This allelic variation may be a potential genetic marker that can help identify those individuals at higher risk for invasive/metastatic diseases. PMID- 10706098 TI - Human MT6-matrix metalloproteinase: identification, progelatinase A activation, and expression in brain tumors. AB - The localization of proteolytic enzymes at the cell surface is a widely used strategy for facilitating tumor invasion. In this study, we have cloned a new member of the membrane-type subfamily of matrix metalloproteinases (MT-MMPs), a group of enzymes associated with tumor progression. The cloned cDNA encodes a protein of 562 amino acids with a domain organization similar to that of other MT MMPs, including a prodomain with a cysteine switch, a catalytic domain with the zinc-binding site, a hemopexin-like domain, and a COOH-terminal extension rich in hydrophobic residues. The predicted protein sequence also contains a short insertion of basic residues located between the propeptide and the catalytic domain and involved in the proteolytic activation of MT-MMPs by furin-like enzymes. Furthermore, immunofluorescence and Western blot analysis of COS-7 cells transfected with the isolated cDNA revealed that the encoded protein is localized at the cell surface. Based on these properties, this novel human matrix metalloproteinase has been called MT6-MMP because it is the sixth identified member of this subfamily of matrix metalloproteinase. Cotransfection of expression plasmids encoding MT6-MMP and progelatinase A resulted in activation of COS-7-secreted progelatinase A, as demonstrated by gelatin zymography. In contrast, transfection of progelatinase A cDNA alone did not lead to the activation of the proenzyme. Northern blot analysis of polyadenylated RNAs isolated from human tissues demonstrated that MT6-MMP is predominantly expressed in leukocytes, lung, and spleen. MT6-MMP was also detected at high levels in SW480 colon carcinoma cells as well as in some anaplastic astrocytomas and glioblastomas, but not in normal colon or brain or in meningiomas. On the basis of these results, we propose that MT6-MMP may facilitate tumor progression through its ability to activate progelatinase A at the membrane of cells from colon carcinomas or brain tumors. PMID- 10706099 TI - Candidate genes for the hypoxic tumor phenotype. AB - In this study, we have analyzed changes induced by hypoxia at the transcriptional level of genes that could be responsible for a more aggressive phenotype. Using a series of DNA array membranes, we identified a group of hypoxia-induced genes that included plasminogen activator inhibitor-1 (PAI-1), insulin-like growth factor-binding protein 3 (IGFBP-3), endothelin-2, low-density lipoprotein receptor-related protein (LRP), BCL2-interacting killer (BIK), migration inhibitory factor (MIF), matrix metalloproteinase-13 (MMP-13), fibroblast growth factor-3 (FGF-3), GADD45, and vascular endothelial growth factor (VEGF). The induction of each gene was confirmed by Northern blot analysis in two different squamous cell carcinoma-derived cell lines. We also analyzed the kinetics of PAI 1 induction by hypoxia in more detail because it is a secreted protein that may serve as a useful molecular marker of hypoxia. On exposure to hypoxia, there was a gradual increase in PAI-1 mRNA between 2 and 24 h of hypoxia followed by a rapid decay after 2 h of reoxygenation. PAI-1 levels were also measured in the serum of a small group of head and neck cancer patients and were found to correlate with the degree of tumor hypoxia found in these patients. PMID- 10706100 TI - EMMPRIN (CD147), an inducer of matrix metalloproteinase synthesis, also binds interstitial collagenase to the tumor cell surface. AB - Extracellular matrix metalloproteinase inducer (EMMPRIN), also known as basigin or CD147, is a glycoprotein that is enriched on the surface of tumor cells and stimulates production of several matrix metalloproteinases by adjacent stromal cells. In this study, we have found that EMMPRIN not only stimulates the production of interstitial collagenase (MMP-1) but also forms a complex with MMP 1 at the tumor cell surface. Complex formation was demonstrated by phage display, affinity chromatography, and immunocytochemistry. Presentation of MMP-1 complexed to EMMPRIN at the tumor cell surface may be important in modifying the tumor cell pericellular matrix to promote invasion. PMID- 10706101 TI - Gene promoter hypermethylation in tumors and serum of head and neck cancer patients. AB - Promoter hypermethylation is an important pathway for repression of gene transcription in cancer cells. We analyzed aberrant DNA methylation at four genes in primary tumors from 95 head and neck cancer patients and then used the presence of this methylation as a marker for cancer cell detection in serum DNA. These four genes were tested by methylation-specific PCR and included: p16 (CDKN2A), O6-methylguanine-DNA-methyltransferase, glutathione S-transferase P1, and death-associated protein kinase (DAP-kinase). Fifty-five % (52 of 95) of the primary tumors displayed promoter hypermethylation in at least one of the genes studied: 27% (26/95) at p16, 33% (31 of 95) at O6-methylguanine-DNA methyltransferase; and 18% (17 of 92) at DAP-kinase. No promoter hypermethylation was observed at the glutathione S-transferase P1 gene promoter. We detected a statistically significant correlation between the presence of DAP-kinase gene promoter hypermethylation and lymph node involvement (P = 0.014) and advanced disease stage (P = 0.016). In 50 patients with paired serum available for epigenetic analysis, the same methylation pattern was detected in the corresponding serum DNA of 21 (42%) cases. Among the patients with methylated serum DNA, 5 developed distant metastasis compared with the occurrence of metastasis in only 1 patient negative for serum promoter hypermethylation (P = 0.056). Promoter hypermethylation of key genes in critical pathways is common in head and neck cancer and represents a promising serum marker for monitoring affected patients. PMID- 10706102 TI - p53 phosphorylation and association with murine double minute 2, c-Jun NH2 terminal kinase, p14ARF, and p300/CBP during the cell cycle and after exposure to ultraviolet irradiation. AB - p53 phosphorylation and association with proteins is implicated in its stability and activity. We have compared the association of DNA-bound and overall pools of p53 with murine double minute 2 (Mdm2), c-Jun NH2-terminal kinase (JNK), p300/CBP, and p14ARF during cell cycle progression. Whereas DNA-bound p53 associates with JNK at G0-G1 and with Mdm2 and p300 during S and G2-M phases, the general pool of p53 was found in complex with JNK and Mdm2 almost throughout the cell cycle. Phosphorylation of p53 at serines 9, 15, and 20 is at the highest levels at G1 and at serines 37 and 392 during G2-M phase. Whereas a high dose of UV irradiation was required for phosphorylation of serines 15 and 392 between 8 and 24 h after treatment, a low dose caused immediate phosphorylation on serines 9, 20, and 372. These dynamic changes in the phosphorylation of p53 are expected to play a pivotal role in p53 association, stability, and function. PMID- 10706103 TI - A germ-line p53 mutation accelerates pulmonary tumorigenesis: p53-independent efficacy of chemopreventive agents green tea or dexamethasone/myo-inositol and chemotherapeutic agents taxol or adriamycin. AB - Recent evidence indicates that individuals with a p53 germ-line mutation (Li Fraumeni syndrome) have a 50% risk of developing lung cancer by age 60. In this study, p53 heterozygous knockout mice and p53 transgenic mice carrying a dominant negative mutant were crossed with the A/J mouse, which is highly susceptible to lung tumor induction, to investigate whether a p53 germ-line mutation is a predisposing gene for carcinogen-induced pulmonary adenomas in mice. The number of lung tumors was not significantly increased in (TSG-p53 x A/J)F1 p53 heterozygous knockout mice as compared with that in (TSG-p53 x A/J)F1 wt mice 16 weeks after exposure to N-nitrosomethylurea (MNU). In contrast, an average of 22 lung tumors were observed in (UL53-3 x A/J)F1 mice carrying a mutant p53 transgene (135Valp53) compared with an average of 7 lung tumors seen in (UL53-3 x A/J)F1 wt mice after treatment with N-nitrosomethylurea. Similar enhancement of lung tumor multiplicity (approximately 3-fold) was seen when mutant versus wt mice were treated with the tobacco-related carcinogens benzo[a]pyrene or 4 (methylnitrosamino)-1-(3-pyridyl)-1-butanone. These results suggest that the mutant p53 transgene may have a dominant negative effect on the wt p53. The potential usefulness of this new mouse model in lung cancer chemoprevention and chemotherapy was examined. The chemopreventive efficacy of the green tea or a combination of dietary dexamethasone and myoinositol and the chemotherapeutic efficacy of Taxol or Adriamycin was examined in wt mice or mice with a mutation in the p53 gene. Mice treated with dexamethasone/myo-inositol and green tea displayed an average of 70 and 50% inhibition of lung tumors, respectively, regardless of p53 status. Similarly, when mice bearing established lung adenomas were treated with Taxol or Adriamycin, a decrease in tumor volume of approximately 70% was observed independent of p53 mutation status. Thus, the (UL53-3 x A/J)F1 p53 transgenic mouse seems to be an excellent model for human carriers of p53 germ-line mutations (Li-Fraumeni syndrome). Furthermore, the lung adenomas generated in this model possess mutations in both the K-ras proto oncogene and the p53 tumor suppressor gene. This model should prove directly useful for chemoprevention and chemotherapy studies. PMID- 10706104 TI - Genotoxic polycyclic aromatic hydrocarbon ortho-quinones generated by aldo-keto reductases induce CYP1A1 via nuclear translocation of the aryl hydrocarbon receptor. AB - Procarcinogenic polycyclic aromatic hydrocarbons (PAHs) induce their own metabolism and activation by binding to the cytosolic aryl hydrocarbon receptor (AhR), which then translocates to the nucleus and activates CYP1A1 gene transcription via xenobiotic response elements (XREs). Although the AhR demonstrates a strict specificity for planar aromatics, nonplanar (+/-)-trans-7,8 dihydroxy-7,8-dihydrobenzo(a)pyrene also induced CYP1A1 expression in HepG2 cells over a delayed timecourse (approximately 6-12 h), suggesting a requirement for (+/-)trans-7,8-dihydrobenzo(a)pyrene metabolism. Aldo-keto reductase (AKR) inhibitors blocked this effect, suggesting that benzo(a)pyrene-7,8-dione (BPQ), a planar PAH o-quinone generated by AKRs, was the downstream inducer. BPQ was found to be a potent and rapid inducer of CYP1A1, with an EC50 value in HepG2 cells identical to that of the parent benzo(a)pyrene. BPQ was a more potent inducer of CYP1A1 when compared with the 1,6-, 3,6-, and 6,12-benzo(a)pyrene-diones. Multiple PAH o-quinones caused induction of CYP1A1, demonstrating that this was a general property of AKR-generated PAH o-quinones. HepG2-101L cells stably transfected with a XRE-luciferase construct showed that BPQ activated CYP1A1 transcription via a XRE-dependent mechanism. BPQ failed to induce CYP1A1 in AhR deficient and AhR nuclear translocator-deficient murine hepatoma cell lines and confirmed that induction of CYP1A1 was AhR and AhR nuclear translocator dependent. Electrophoretic mobility shift assays demonstrated the specific appearance of BPQ-activated AhR in the nucleus, and immunofluorescence studies confirmed that BPQ mediated nuclear translocation of the AhR. Classical bifunctional inducers elevate CYP1A1 expression via a XRE and are subsequently converted by CYP1A1 to electrophiles that induce phase II enzymes via an electrophilic response element/antioxidant response element PAH o-quinones represent a novel class of bifunctional inducer because they are electrophiles produced by phase II enzymes that simultaneously induce phase I enzymes via a XRE and phase II enzymes via a electrophilic response element/antioxidant response element (see also M. E. Burczynski et al., Cancer Res., 59: 607-614, 1999). This study shows that the AhR provides the only known mechanism by which genotoxic PAH o-quinones generated in the cytosol can be targeted to the nucleus with specificity. PMID- 10706105 TI - High lactate levels predict likelihood of metastases, tumor recurrence, and restricted patient survival in human cervical cancers. AB - Pathophysiological parameters such as vascular density and tissue oxygen pressure can influence tumor malignancy and patient survival. Observations from our group showed that metastatic spread of carcinomas of the uterine cervix and of head and neck cancers was closely correlated with the lactate concentration in the primary lesion. Because these results were obtained in a low number of patients, the present investigation was performed to verify such a correlation in a larger population. Cryobiopsies were taken at first diagnosis of cervical cancer from 34 patients. Tissue concentrations of ATP, glucose, and lactate in viable tumor regions of these biopsies were measured microscopically using the technique of imaging bioluminescence. There was no correlation between stage or grade and any of the metabolic parameters measured. ATP and glucose concentrations were not significantly different in metastatic and nonmetastatic primary tumors (P>0.05). However, lactate concentrations were significantly higher (P = 0.001) in tumors with metastatic spread (mean +/- SD, 10.0+/-2.9 micromol/g; n = 20) compared with malignancies in patients without metastases (6.3+/-2.8 micromol/g; n = 14). The majority of patients who suffered a recurrence of the disease (17 of a total of 22 patients) or died (15 of 20) within the observation period of up to 8 years belonged to the metastatic, i.e., high lactate group. A Kaplan-Meier analysis of the data showed that the overall and disease-free survival probabilities of patients having low tumor lactate values were significantly higher compared with patients with high tumor lactate concentrations (P = 0.015 and 0.014, respectively). We conclude that tumor lactate content may be used as a prognostic parameter in the clinic. Furthermore, these findings are in accordance with data from the literature showing that the presence of hypoxia in cervical tumors is associated with a poorer patient survival. PMID- 10706106 TI - A novel SMAD4 gene mutation in seminoma germ cell tumors. AB - Transforming growth factor (TGF)-beta is known as an antiproliferative factor in the majority of mammalian cells, including stem germ cells. Lack of TGF-beta induced growth inhibition has been associated with disruptions of TGF-beta receptors and SMADs. In the present study, we performed a mutational analysis of the TGF-beta signaling system, including TGF-beta receptor type I and type II and SMADs (SMAD1-SMAD7), in 20 seminoma germ cell tumors. Using reverse transcription PCR, single-strand conformational polymorphism, and sequencing analysis, the COOH terminal domain of SMAD4 was found to be mutated: a single thymine was inserted between nt 1521 and 1522 in 2 of 20 tumors analyzed. This addition of a thymine creates a frameshift and a new stop signal at codon 492, which leads to premature termination of the encoded protein. Such a mutation potentially abrogates signaling from TGF-beta as well as the other TGF-beta family members, including activin and bone morphogenetic protein, which all use the SMAD pathway. Immunohistological analysis confirmed the loss of expression of SMAD4 protein in the seminoma tissues with the insertional mutation. To our knowledge, this is the first description of a novel SMAD4 insertional mutation in seminoma testicular germ cell tumors. This mutational inactivation of SMAD4/COOH-terminal domain may cause TGF-beta unresponsiveness. It could thus provide a basis for understanding the potential role of the TGF-beta system in germ cell tumorigenesis. PMID- 10706107 TI - Down-regulation of transforming growth factor beta receptors by androgen in ovarian cancer cells. AB - Steroid hormones have been implicated in the etiology and/or progression of epithelial ovarian cancer. As ovarian surface epithelial cells are growth inhibited by transforming growth factor beta (TGF-beta), we tested whether steroid hormones could regulate the expression of TGF-beta1 or its receptors in ovarian cancer cells, as assessed by quantitative reverse transcription-PCR. Treatment of ovarian cancer HEY cells with 500 nM 5alpha-dihydrotestosterone (DHT), but not estradiol-17beta or progesterone, for 60 h down-regulated the expression of mRNA for TGF-beta receptors I and II (TbetaR-I and TbetaR-II), betaglycan, and endoglin but had no effect on TGF-beta1 mRNA levels. Androgen receptor (AR) mRNA expression in HEY cells was compared to other ovarian cancer cell lines. OVCAR-3 cells expressed AR mRNA levels similar to that of androgen responsive LNCaP prostate cancer cells, whereas SKOV-3 and HEY cells expressed only 3 and 0.01%, respectively. Western blot analysis and saturation binding assays confirmed the expression of AR protein in these three cell lines, but at the limit of detection in SKOV-3 and HEY cells. Treatment of SKOV-3 and HEY cells for 24 h with 1-50 nM DHT resulted in a dose-dependent down-regulation of TbetaR II mRNA. The AR antagonist hydroxyflutamide did not reverse the effect of DHT on SKOV-3 cells but by itself down-regulated TbetaR-II mRNA. This apparent androgen mimetic action of hydroxyflutamide and the ability of SKOV-3 and HEY cells to respond to DHT may be due to their expression of AR-associating protein 70, an AR co-activator reported to amplify AR transactivation and to result in agonist activity of AR antagonists. DHT was able to reverse TGF-beta1 growth-inhibitory action in SKOV-3 cells and in a primary culture of ovarian cancer cells derived from ascites. Thus, androgens may promote ovarian cancer progression in part by decreasing TGF-beta receptor levels, thereby allowing ovarian cancer cells to escape TGF-beta1 growth inhibition. PMID- 10706108 TI - The 4-pregnene and 5alpha-pregnane progesterone metabolites formed in nontumorous and tumorous breast tissue have opposite effects on breast cell proliferation and adhesion. AB - Progesterone is required for the full proliferative activity of the breasts and may be directly or indirectly involved in either stimulating or inhibiting breast cancer. To determine whether the effects on breast cancer are attributable to progesterone metabolites, we compared the capacity of nontumorous and tumorous breast tissue to convert progesterone and then tested the effects of these metabolites on breast cell proliferation and anchorage. Tissues from the operated breasts of six patients with infiltrating duct carcinomas were incubated with [14C]progesterone for 2, 4, and 8 h, and the metabolites were identified and quantified. The identified metabolites (equal to >95% of recovered radioactivity) can be divided into those that retain the double bond of progesterone in the carbon-4 position of ring A (4-pregnenes) and those that are 5alpha-reduced (5alpha-pregnanes). The results show that tumorous breast tissue has elevated 5alpha-reductase activity, which results in significantly higher total levels of 5alpha-pregnanes, especially 5alpha-pregnane-3,20-dione (5alphaP), whereas normal (nontumorous) breast tissue produces more 4-pregnenes, especially 3alpha-hydroxy 4-pregnen-20-one (3alphaHP). 5alphaP and 3alphaHP are each one enzymatic step removed from progesterone, resulting from the action of either 5alpha-reductase or 3alpha-hydroxysteroid oxidoreductase (3alpha-HSO), respectively. The ratio of 5alpha-pregnanes:4-pregnenes is >5-fold greater and the ratio of 5alphaP:3alphaHP is nearly 30-fold greater in tumorous than nontumorous breast tissue incubates. In vitro studies with three breast cell lines (MCF-7, MCF-10A, and ZR-75-1) show that 3alphaHP dose dependently inhibits, whereas 5alphaP significantly stimulates, proliferation. Additional studies with MCF-7 and MCF-10A cells indicate that each of the 4-pregnenes isolated from breast tissue suppresses, whereas each respective 5alpha-reduced product stimulates, cell proliferation. Studies of cell anchorage were conducted using MCF-7 cells and various concentrations of 5alphaP or 3alphaHP. The number of cells attached to the substrate was significantly (P<0.05) decreased by treatment with > or =30 nM 5alphaP and increased by treatment with > or =50 nM 3alphaHP. Conversely, the number of cells detached from the substrate after partial trypsin exposure was significantly increased by treatment with > or =40 nM 5alphaP and decreased by treatment with > or =30 nM 3alphaHP. The results suggest that a change in in situ progesterone metabolism, resulting in an increased 5alpha-pregnane:4-pregnene (especially 5alphaP:3alphaHP) ratio, may promote breast cancer by promoting increased cell proliferation and detachment, whereas increases in 4-pregnenes may retard these tumorigenic processes. These studies suggest that endogenous progesterone metabolites may provide a new hormonal basis for breast cancer. PMID- 10706109 TI - Androgen receptor mutations in prostate cancer. AB - We analyzed the frequency and relevance of mutations in the coding region of the androgen receptor (AR) in genomic DNA extracted from 137 specimens of prostate cancer. The specimens were obtained from the primary tumors of patients affected by stage B disease [15 nonmicrodissected (group 1A) and 84 microdissected (group 1B)] and from the metastatic deposits of individuals with stage D1 disease [8 nonmicrodissected (group 2A) and 30 microdissected (group 2B)] who had not undergone androgen ablation therapy. The study was conducted by PCR-single strand conformational polymorphism (SSCP) analysis of exons 2-8 in the four groups and direct sequence analysis of exon 1 in group 1B. As positive and negative controls, we used genomic DNA extracted from genital skin fibroblasts of patients affected by various forms of androgen resistance with known mutations in the AR. To control for genetic instability, PCR-SSCP analysis of exon 2 of the human progesterone receptor was carried out on each specimen. The overall number of mutations detected was 11 (8%). No mutations were detected in any of the 99 patients with stage B disease. Eleven mutations were detected in exons 2-8 in 8 of the 38 patients with stage D1 disease (all in group 2B). Simultaneous analysis of exon 2 of the progesterone receptor was carried out, and no SSCP changes were identified. These data suggest that AR mutations are rare and presumably do not play a role in the initial phase of prostatic carcinogenesis. The presence of a significant number of AR mutations in metastatic disease indicates that mutations of this molecule may play a role in the most advanced phases of the natural history of this disease, either by facilitating growth or acquisition of the metastatic phenotype. PMID- 10706110 TI - Genetic polymorphisms in uridine diphospho-glucuronosyltransferase 1A1 and association with breast cancer among African Americans. AB - We examined the role of constitutional genetic variation at the UDP glucuronosyltransferase (UGT) 1A1 locus in breast cancer susceptibility. The UGT1A1 enzyme is a major UGT involved in estradiol glucuronidation. To date, four UGT1A1 variant alleles characterized by a variation in the number of TA from five through eight repeats in the atypical TATA box region have been described in the African-American population. Functional analyses of the transcriptional activity in breast and liver cells revealed that the transcription activation of a reporter gene is inversely correlated with the number of repeats. Reverse transcription-PCR analysis confirmed the expression of UGT1A1 in human liver in the hepatocarcinoma cell line HepG2 and provided evidence of the expression of UGT1A1 in breast cancer tissue, where a positive signal was observed in 11 of 12 breast cancer cell lines tested. The population-based case-control study involved 200 women with breast cancer and 200 female controls of African ancestry. We postulated that breast cancer cases might have a higher prevalence of low activity allele-containing genotypes than controls (alleles presenting seven and eight repeats in the A(TA)nTAA motif of the TATA box). The age-adjusted odds ratio (OR) for breast cancer comparing women with seven and eight allele containing genotypes versus 5/5, 5/6, and 6/6 genotypes was 1.8 [95% confidence interval (CI), 1.0-3.1; P = 0.06] in premenopausal women and 1.0 (95% CI, 0.5 1.7; P = 0.9) in postmenopausal women. The observed 1.8-fold elevated risk in premenopausal women with invasive breast cancer is highly suggestive of a possible interaction between UGT genotype and hormones. Additional analyses suggested a stronger association of UGT1A1 genotype with estrogen receptor (ER) negative breast cancer. Among premenopausal women, the association was stronger for ER- breast cancer (OR, 2.1; 95% CI, 1.0-4.2; P = 0.04) than ER+ breast cancer (OR, 1.3; 95% CI, 0.6-3.0; P = 0.5). The OR was slightly stronger among women who used oral contraceptives, and the association remained null in postmenopausal women, regardless of whether they took hormone replacement therapy. Our current findings suggest that further investigations are warranted to elucidate the role of UGT1A1 in breast cancer risk. PMID- 10706111 TI - Chemoprevention of aflatoxin B1 hepatocarcinogenesis by coumarin, a natural benzopyrone that is a potent inducer of aflatoxin B1-aldehyde reductase, the glutathione S-transferase A5 and P1 subunits, and NAD(P)H:quinone oxidoreductase in rat liver. AB - Structurally diverse compounds can confer resistance to aflatoxin B1 (AFB1) hepatocarcinogenesis in the rat. Treatment with either phytochemicals [benzyl isothiocyanate, coumarin (CMRN), or indole-3-carbinol] or synthetic antioxidants and other drugs (butylated hydroxyanisole, diethyl maleate, ethoxyquin, beta naphthoflavone, oltipraz, phenobarbital, or trans-stilbene oxide) has been found to increase hepatic aldo-keto reductase activity toward AFB1-dialdehyde and glutathione S-transferase (GST) activity toward AFB1-8,9-epoxide in both male and female rats. Under the conditions used, the natural benzopyrone CMRN was a major inducer of the AFB1 aldehyde reductase (AFAR) and the aflatoxin-conjugating class alpha GST A5 subunit in rat liver, causing elevations of between 25- and 35-fold in hepatic levels of these proteins. Induction was not limited to AFAR and GSTA5: treatment with CMRN caused similar increases in the amount of the class-pi GST P1 subunit and NAD(P)H: quinone oxidoreductase in rat liver. Immunohistochemistry demonstrated that the overexpression of AFAR, GSTA5, GSTP1, and NAD(P)H:quinone oxidoreductase affected by CMRN is restricted to the centrilobular (periacinar) zone of the lobule, sometimes extending almost as far as the portal tract. This pattern of induction was also observed with ethoxyquin, oltipraz, and trans stilbene oxide. By contrast, induction of these proteins by beta-naphthoflavone and diethyl maleate was predominantly periportal. Northern blotting showed that induction of these phase II drug-metabolizing enzymes by CMRN was accompanied by similar increases in the levels of their mRNAs. To assess the biological significance of enzyme induction by dietary CMRN, two intervention studies were performed in which the ability of the benzopyrone to inhibit either AFB1 initiated preneoplastic nodules (at 13 weeks) or AFB1-initiated liver tumors (at 50 weeks) was investigated. Animals pretreated with CMRN for 2 weeks prior to administration of AFB1, and with continued treatment during exposure to the carcinogen for a further 11 weeks, were protected completely from development of hepatic preneoplastic lesions by 13 weeks. In the longer-term dietary intervention, treatment with CMRN before and during exposure to AFB1 for a total of 24 weeks was found to significantly inhibit the number and size of tumors that subsequently developed by 50 weeks. These data suggest that consumption of a CMRN containing diet provides substantial protection against the initiation of AFB1 hepatocarcinogenesis in the rat. PMID- 10706112 TI - ZD4190: an orally active inhibitor of vascular endothelial growth factor signaling with broad-spectrum antitumor efficacy. AB - There is evidence that vascular endothelial growth factor (VEGF) contributes to solid tumor growth through the promotion of both angiogenesis and tumor vascular permeability. To abrogate VEGF signaling, we developed a small molecular weight inhibitor of VEGF receptor tyrosine kinase (RTK) activity that was compatible with chronic oral administration. ZD4190, a substituted 4-anilinoquinazoline, is a potent inhibitor of KDR and Flt-1 RTK activity, and VEGF stimulated HUVEC proliferation in vitro. Chronic once-daily oral dosing of ZD4190 to young rats produced a dose-dependent increase in the femoral epiphyseal growth plate area, which may be attributed to the inhibition of VEGF signaling in vivo because vascular invasion of cartilage is a prerequisite to the process of ossification. Once-daily oral dosing of ZD4190 to mice bearing established (approximately 0.5 cm3) human tumor xenografts (breast, lung, prostate, and ovarian) elicited significant antitumor activity and at doses that would not be expected to have any direct antiproliferative effect on tumor cells. Prolonged tumor cytostasis was further demonstrated in a PC-3 xenograft model with 10 weeks of ZD4190 dosing, and upon withdrawal of therapy, tumor growth resumed after a short delay. These observations are entirely consistent with the proposed mode of action. ZD4190 is one of a series of VEGF RTK inhibitors that may have utility in the treatment of a range of histologically diverse solid tumor types. PMID- 10706113 TI - Retroviral immunotoxin gene therapy of acute myelogenous leukemia in mice using cytotoxic T cells transduced with an interleukin 4/diphtheria toxin gene. AB - The potential benefit of immunotoxin (IT) for cancer therapy has mostly been limited by the fact that only a small portion of injected dose ever reaches the cancer target. Thus, we set out to determine whether antigen-specific CTLs would be suitable vehicles to deliver IT to the site of cancer cells in vivo. A retroviral vector was constructed for gene therapy with (interleukin 4) IL-4 positioned downstream of its 20-amino-acid leader sequence that permitted cotranslational protein synthesis of IT along with truncated diphtheria toxin (DT). IL-4 was chosen as a ligand based on the expression of IL-4 receptor (IL 4R) on most acute myeloid leukemia cases. The first-time expression and secretion of a cytokine/DT fusion toxin was accomplished in mammalian NIH.3T3 cells, and then a retroviral vector was assembled. The retroviral IT was used to transiently transduce T15, a CD8+ CTL T cell line that specifically recognized C1498 (a lethal murine acute myeloid leukemia cell line). Transduced T15 T cells expressed intracellular DT and IL-4 as determined by immunofluorescence. Secreted IT supernatants collected from T15 had enzymatic activity and killed IL-4R+ C1498 cells, but not IL-4R- EL4 cells. Intravenous injection of transduced T15, but not nontransduced T15, into mice with s.c. tumors significantly inhibited tumor growth. In contrast, systemic therapy with a bacterial preparation of the same IL 4 IT given at its maximum tolerated dose did not protect. Retroviral IT-treated mice showed no sign of the renal or hepatic toxicity that is common to this class of IT. Together, these data indicate that retroviral IT may solve problems relating to systemic IT therapy by delivering reagent more directly to the site of cancer in vivo and may impart new anticancer defense mechanisms to antigen specific T cells. PMID- 10706114 TI - Microtubule disruption induced in vivo by alkylation of beta-tubulin by 1-aryl-3 (2-chloroethyl)ureas, a novel class of soft alkylating agents. AB - We have previously reported that 4-tert-butyl-[3-(2-chloroethyl)ureido] benzene (4-tBCEU), a potent cytotoxic agent, modulates the synthesis of tubulins, suggesting that its cytotoxicity may be mediated through an antimicrotubule mechanism. Indeed, 4-tBCEU and its 4-iso-propyl (4-isopropyl [3-(2 chloroethyl)ureido] benzene) and 4-sec-butyl (4-sec-butyl [3-(2 chloroethyl)ureido] benzene) homologues induced disruption of the cytoskeleton and arrest of the cell cycle in G2 transition and mitosis. To better understand the mechanisms responsible for microtubule disruption by 1-aryl-3-(2 chloroethyl)ureas (CEU), we first examined their cytotoxicity on Chinese hamster ovary cells resistant to vinblastine and colchicine due to the expression of mutated tubulins (CHO-VV 3-2). These cells showed resistance to CEU, e.g., 4 tBCEU having an IC50 of 21.3+/-1.1 microM as compared with an IC50 of 11.6+/-0.7 microM for wild-type cells, suggesting a direct effect of the drugs on tubulins. Western blot analysis confirmed the disruption of microtubules and evidenced the formation of an additional immunoreactive beta-tubulin with an apparent lower molecular weight on SDS polyacrylamide gel. Incubation of MDA-MB-231 cells with [urea-14C]-4-tBCEU revealed the presence of a radioactive protein that coincided with the additional beta-tubulin band, indicating that CEU could covalently bind to the beta-tubulin. The 4-tBCEU-binding site on beta-tubulin was identified by competition of the CEU with colchicine, vinblastine, and iodoacetamide, a specific alkylating agent of sulfhydryl groups of cysteine residues. Colchicine, but not vinblastine, prevented the formation of the additional beta-tubulin band, suggesting that 4-tBCEU alkylates either Cys239 or Cys354 residues near the colchicine-binding site. To determine the cysteine residue alkylated by 4-tBCEU, we incubated the radiolabeled drug with human neuroblastoma cells (SK-N-SH) that overexpress the betaIII-tubulin, an isoform where Cys239 is replaced by a serine residue. The results clearly showed that betaIII-tubulin is not alkylated by [urea-14C]-4-tBCEU, suggesting that cysteine 239 residue is essential for the reactivity of 4-tBCEU with beta-tubulin. Taken together, these findings indicate that the mechanism of cytotoxicity of CEU involves microtubule depolymerization through alkylation of beta-tubulin. PMID- 10706115 TI - Evaluation of HSV-tk gene therapy in a rat model of chemically induced hepatocellular carcinoma by intratumoral and intrahepatic artery routes. AB - Transfer of the herpes simplex virus-thymidine kinase (HSV-tk) gene followed by the administration of ganciclovir (GCV) into hepatocellular carcinoma (HCC) derived cell lines either in vitro or transplanted into nude mice has been shown to provide a potential strategy for HSV-tk-based gene therapy of HCC. We report herein an analysis of the antitumoral efficacy of two recombinant adenoviruses (Ads), Ad.CMVtk and Ad.AFPtk, in a relevant model of multifocal hepatic lesions induced in rats by a potent alkylating chemical carcinogen, diethylnitrosamine. Two routes of administration of the Ad were studied: intratumoral and intrahepatic artery injections. Both recombinant Ads, Ad.CMVtk and Ad.AFPtk, express the HSV-tk gene under the control of the early enhancer/promoter cytomegalovirus and alpha-fetoprotein regulatory gene sequences, respectively. The antitumor response was assessed by magnetic resonance imaging and by autopsy and histological analysis following postmortem. Tumor growth cessation was demonstrated by magnetic resonance imaging in large tumor nodules of size 5-8 mm treated by intratumoral administration of 2x10(9) pfu Ad.CMVtk plus i.p. treatment with GCV. We also show an antitumor efficacy in small tumor nodules of size <3 mm treated with 2x10(9) pfu Ad.CMVtk plus GCV by the intrahepatic artery route, albeit associated with an adverse toxicity. In vivo targeting of the HSV tk gene to diethylnitrosamine-induced HCC cells with the recombinant Ad.AFPtk suppresses the hepatic toxicity in the nontumoral liver. The lower antitumor response would argue for the use of multiple injections of such adenoviral constructs. These observations may lead to potential approaches for designing gene therapy destined for early treatment of dysplastic nodules or advanced HCC in cirrhosis. PMID- 10706116 TI - Inhibition of human prostate cancer proliferation in vitro and in a mouse model by a compound synthesized to block Ca2+ entry. AB - Accelerated Ca2+ entry may be one component of the pathway regulating the proliferative phenotype of some types of cancer. Thus, a pharmacological agent with the ability to retard Ca2+ influx in susceptible cancers might inhibit proliferation of them by a cytostatic mechanism rather than by inducing cytotoxicity. We have developed a chemical synthetic scheme that has produced a small library of novel compounds that block Ca2+ entry induced by occupancy of the P2 receptor in two prostate cancer cell lines and inhibit proliferation of these cells in vitro. One of the agents, named TH-1177, was used to treat severe combined immunodeficient mice inoculated with the human prostate cancer line PC 3. Although the doses used and treatment schedule were chosen arbitrarily, treatment extended the mean life span of mice bearing tumors by up to 38%. Treatment of mice without cancer at doses 18 times that used in mice with tumors was not associated with any obvious toxicity, either grossly or on histological examination. These results suggest that novel cytostatic agents with efficacy against human prostate cancer cells can be developed by chemical synthesis of agents directed at the Ca2+ entry pathway. PMID- 10706117 TI - ONYX-015 works synergistically with chemotherapy in lung cancer cell lines and primary cultures freshly made from lung cancer patients. AB - p53 mutations and loss of heterozygosity (LOH) have been detected in >50% of lung cancers. Wild-type p53 can prevent replication of damaged DNA and promote apoptosis of cells with abnormal DNA. A human adenovirus, ONYX-015, which has a deletion in the E1B region, has shown tumor-specific cytolytic effect in tumor cells with nonfunctional p53 and antitumor efficacy that can be augmented by chemotherapeutic agents. A recent report from an independent group, however, indicates that wild-type p53 is necessary for the infection of this replicating virus, and it is in direct contradiction to previous observations of the ONYX group. In this study, we carried out cytopathic effect (CPE) assays using ONYX 015 on five human lung cancer cell lines with known p53 status. Two of these cell lines, NCI-H522 and NCI-H1703, have mutations and LOH in their p53 gene. Both lines were lysed in a dose-dependent manner and showed 100% cytolysis at a multiplicity of infection of 0.1. Two additional cell lines, NCI-H2347 and NCI H838, both of which have wild-type p53 gene, showed near complete lysis at a multiplicity of infection of 1. We demonstrate here that the lung cancer cells with nonfunctional p53 are at least 10 times more sensitive to ONYX-015 cytolysis than the lung cancer cells with wild-type p53. In addition, standard chemotherapeutic agents (paclitaxol and cisplatin) showed a synergistic effect when combined with ONYX-015, and this effect was p53 mutant dependent. Furthermore, we tested the cytolytic effect of ONYX-015 on a panel (n = 7) of primary first-passage cultures made from freshly resected lung cancers. ONYX-015 lysed primary lung cancer cells in six of seven (86 %) primary cultures. Two of four primary cultures treated with chemotherapeutic agents had a synergistic effect with ONYX-015. Our data indicate that wild-type p53 is not required for the infection of this replicating virus, and also we demonstrate that ONYX-015 is effective alone and works synergistically with chemotherapeutic agents in lung cancer cell lines and primary cultures. This study suggests that ONYX-015 may be effective, especially in combination with conventional chemotherapy, in the treatment of patients with lung cancer. PMID- 10706118 TI - Apoptosis induction of human myeloid leukemic cells by ultrasound exposure. AB - Therapeutic ultrasound (ULS) and the resulting cavitation process has been shown to induce irreversible cell damage. In this study, we wanted to further investigate the mechanism of ULS-induced cell death and to determine whether apoptosis is involved. High intensity focused pulsed ULS sonication at a frequency of 750 KHz was delivered to HL-60, K562, U937, and M1/2 leukemia cell line cultures. ULS exposure used with induction of transient cavitation in the focal area was delivered with an intensity level of 103.7 W/cm2 and 54.6 W/cm2 spatial-peak temporal-average intensity. As a control, ULS of lower intensity was delivered at 22.4 W/cm2 spatial-peak temporal-average intensity, presumably without generation of cavitation. Our results indicated that DNA damage induced by ULS cavitation did not involve generation of free radicals in the culture media. Morphological alterations observed in cells after exposure to ULS included: cell shrinkage, membrane blebbing, chromatin condensation, nuclear fragmentation, and apoptotic body formation. Apoptotic cells were evaluated by fluorescence microscopy and detected using the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay, which identifies DNA breaks, and by the leakage of phosphatidylserine from the inner to the outer side of the membrane layer of treated cells. Some bioeffects induced on sonicated HL-60 cells, such as inhibition of cell proliferation, DNA repair, and cell-dependent apoptosis, were found to be similar to those produced by gamma-irradiation. Thus, much of the cell damage induced by therapeutic ULS in leukemia cells surviving ULS exposure appears to occur through an apoptotic mechanism. PMID- 10706119 TI - APRIL/TRDL-1, a tumor necrosis factor-like ligand, stimulates cell death. AB - We have examined the activity of a new member of the tumor necrosis factor (TNF) family identified through Expressed Sequence Tag database searching using TNFalpha protein as the search query. We have termed this protein TNF-related death ligand-la (TRDL-1alpha). Traditional cDNA library screening identified two additional splice variants designated TRDL-1beta and TRDL-1gamma that differed from TRDL-1alpha by the deletion of two small regions within the protein coding region. TRDL-1alpha is identical in sequence to the recently described molecule, APRIL, that may induce cell proliferation. We found, however, that purified, FLAG tagged TRDL-1alpha caused Jurkat cell death with kinetics that paralleled FasL. In vitro binding experiments demonstrated that TRDL-1alpha coprecipitated Fas and HVEM and suggested TRDL-1alpha as an alternate ligand for these receptors. TRDL-1 localized to chromosome 17p13.3 and its expression was widespread in normal tissues. Examination of 48 tumor samples revealed high levels of TRDL-1 expression in several tumors, including those from the gastrointestinal tract. Expression of TRDL-1 in COS-1 cells confirmed membrane association of TRDL-1, typical of TNF family members. PMID- 10706120 TI - Induction of tumor immunity and cytotoxic T lymphocyte responses using dendritic cells transfected with messenger RNA amplified from tumor cells. AB - Unique patient-specific tumor antigens may constitute the dominant antigens in the antitumor immune response. Hence, vaccination with the patient's own repertoire of tumor antigens may offer a superior strategy to elicit protective immunity. We have shown previously that dendritic cells transfected with mRNA isolated from tumor cells stimulate potent CTL responses and engender protective immunity in tumor-bearing mice. In the current study, we demonstrate that tumor mRNA, isolated from murine tumor cell lines or from primary human tumor cells microdissected from frozen tissue sections, can be amplified without loss of function. This study provides the foundations for an effective and broadly applicable treatment that does not require the characterization of the relevant antigenic profile in each patient and will not be limited by tumor tissue availability for antigen preparation. PMID- 10706121 TI - Enhancement of DNA vaccine potency by linkage of antigen gene to an HSP70 gene. AB - Nucleic acid vaccines represent an attractive approach to generating antigen specific immunity because of their stability and simplicity of delivery. However, there is still a need to increase the potency of DNA vaccines. Using human papillomavirus type 16 E7 as a model antigen, we evaluated the effect of linkage to Mycobacterium tuberculosis heat shock protein 70 (HSP70) on the potency of antigen-specific immunity generated by naked DNA vaccines. We found that vaccines containing E7-HSP70 fusion genes increased the frequency of E7-specific CD8+ T cells by at least 30-fold relative to vaccines containing the wild-type E7 gene. More importantly, this fusion converted a less effective vaccine into one with significant potency against established E7-expressing tumors. Surprisingly, E7 HSP70 fusion vaccines exclusively targeted CD8+ T cells; immunological and antitumor effects were completely CD4-independent. These results indicate that fusion of HSP70 to an antigen gene may greatly enhance the potency of DNA vaccines via CD8-dependent pathways. PMID- 10706122 TI - Increasing methylation of the CDKN2A gene is associated with the progression of adult T-cell leukemia. AB - In this study, we examined the methylation status of the CDKN2A gene in patients with different forms of adult T-cell leukemia (ATL) using Southern blot analysis, methylation-specific PCR (MSPCR), and nucleotide sequencing. We found that the CDKN2A gene was more frequently methylated in fresh tumor cells isolated from patients with acute ATL (47%) or lymphoma-type ATL (73%) than in those with less malignant chronic (17%) and smoldering (17%) ATL. In addition, deletions of the CDKN2A gene were found in 24% of acute ATL patients; thus, abnormalities of the CDKN2A gene totaled 71% in acute ATL patients. In contrast, no CDKN2A gene methylation was found in asymptomatic carriers or uninfected individuals. Methylation of the p15 gene was not found in any samples from 36 ATL patients. Direct sequencing of the CDKN2A gene after sodium bisulfite treatment of genomic DNA revealed that the methylation of CpG sites had occurred in 24 of 32 ATL cases (75%) including chronic and smoldering ATL, even when MSPCR and the Southern blot had failed to detect CDKN2A gene methylation. Among fresh ATL samples with methylation, methylation was detected in the promoter region and exon in 17 of 24 cases, and methylation in the exon without promoter region was detected in 7 of 24 cases. In one case, the pattern of methylation proved to be different between peripheral blood cells and lymph node cells, suggesting the presence of multiple subclones with regard to methylation patterns, despite the same HTLV-I integration site. Quantitative PCR showed a marked decrease in CDKN2A mRNA expression in the cells with a methylated CDKN2A gene, especially if the promoter region was methylated. These findings suggest that CpG methylation decreases CDKN2A expression and represents a critical factor in the disease progression of ATL. PMID- 10706123 TI - Alterations of the FHIT gene in human hepatocellular carcinoma. AB - FHIT (fragile histidine triad), a candidate tumor suppressor gene, encompasses FRA3B, a region with the highest fragility in the human genome, and is altered in a large number of human cancers, particularly those of epithelial cell origin and associated with known carcinogenic agents. Human hepatocellular carcinoma (HCC), a major cancer worldwide, is closely related to carcinogenic agents such as hepatitis B and C virus infections, dietary aflatoxin, alcohol consumption, and exposure to chemical carcinogens. To assess the extent and the nature of the FHIT gene alterations and their implications in the development of HCC, several cell lines and primary tumors were cytologically and molecularly examined. The FHIT gene is expressed in normal hepatic cells and is not expressed or is abnormally expressed in cultured tumor cells derived from HCC. Down-regulation of the FHIT gene was detected by Northern blot analysis in 9 of 14 cell lines However, neither abnormal FHIT transcripts nor point mutations in DNA sequences of reverse transcription-PCR products (exons 2-9) were identified. Expression of FHIT protein was not detected by immunostaining in 5 of 10 primary tumors. Four cell lines showing mRNA down-regulation did not express FHIT protein as demonstrated by Western blot analysis. Allelic loss of intron 5 of the FHIT gene was detected in 10 of 34 informative samples from primary tumors. Structural alterations of chromosome 3p were identified in 8 of 13 HCC cell lines. Deletions or translocations involving region 3p14.2 were identified by fluorescence in situ hybridization with a YAC850A6 probe spanning the FHIT locus on chromosomes derived from cell lines with an abnormal FHIT gene expression. These combined results indicate that the FHIT gene is a frequent target and may be implicated in a subset of liver cancers. PMID- 10706124 TI - Mage-b4, a novel melanoma antigen (MAGE) gene specifically expressed during germ cell differentiation. AB - The MAGE genes were initially isolated from different kinds of tumors, and based on their virtually exclusive tumor-specific expression in adult tissues, they have been used as targets for cancer immunotherapy. However, although a large number of MAGE genes have now been identified and extensively studied in tumors of various origin, their functions in normal cells remain unknown. Here we describe the isolation and characterization of a novel murine MAGE homologue, Mage-b4. mRNA expression studies in a wide variety of adult and embryonic tissues revealed that Mage-b4 is specifically expressed in fetal and adult gonads. An antibody specific to Mage-b4 was developed, and using this antibody, we found that the Mage-b4 protein was confined to the cytoplasm of germ cells. Double labeling experiments using antibodies against the meiosis-specific SCP3 protein and the Mage-b4 protein showed that Mage-b4 is down-regulated as the germ cells enter meiosis in adult testis. In contrast, Mage-b4 was expressed in female germ cells throughout meiosis, and the protein was also found in dormant primary oocytes. PMID- 10706125 TI - Elevated frequency and functional activity of a specific germ-line p53 intron mutation in familial breast cancer. AB - Previous studies have determined that the frequency of germ-line p53 mutations in familial breast cancer patients is 1% or less, but these reports have not investigated the importance of polymorphic intron base changes in the p53 gene. Therefore, we investigated the frequency of both exon and intron germ-line p53 base changes in 42 breast cancer patients with a strong family history of breast cancer. The mean age of presentation of these patients was 44.0 years (range, 29 69), and 12 of 42 (29%) were of known Ashkenazi ancestry. Purified DNA obtained from the 42 index cases was screened for germ-line p53 mutations in exons 2-11 and surrounding introns using a combination of intron based primers for PCR single strand conformation polymorphism analysis, direct sequencing, and microarray sequencing using the Affymetrix p53 gene chip methodology. Morphological analysis of apoptosis and cell survival determination were performed on EBV-immortalized lymphoblastoid cell lines from two patients with the p53 intron 6 mutation. A germ-line mutation in the p53 gene at nucleotide 13964 with a G to C base change (13964GC) was identified in 3 of 42 (7.1%) hereditary breast cancer patients. Two patients were heterozygous for this mutation, and one patient had a homozygous mutation. In comparison, 0 of 171 (0%) of sporadic breast cancer patients had the p53 13964GC mutation (P = 0.0003). We found that 0 of 42 (0%) of these hereditary breast cancer patients had other germ line p53 mutation in exons 2-11. However, pedigree analysis demonstrated that all three patients had strong family histories of multiple types of cancers consistent with Li-Fraumeni syndrome but with late age of onset. Comprehensive BRCA1 and BRCA2 nucleotide analysis from patients with the p53 13964GC mutation revealed no concomitant deleterious BRCA1 or BRCA2 mutations, although they were found in the other hereditary breast cancer patients. Functional analysis of two immortalized lymphoblastoid cell lines derived from patients with the p53 13964GC mutation demonstrated prolonged in vitro survival in response to cisplatinum treatment and showed decreased chemotherapy-induced apoptosis. Immunohistochemical analysis of breast tumors from these patients revealed high levels of mutant p53 protein, suggesting a functional mutation in the p53 gene. In summary, we have identified a single p53 intron mutation in familial breast cancer patients that is present at elevated frequency and has functional activity. PMID- 10706126 TI - Insulin-like growth factor II supply modifies growth of intestinal adenoma in Apc(Min/+) mice. AB - Insulin-like growth factor-II (IGF-II) is an embryonic growth promoter and cell survival factor. IGF-II supply is normally limited by gene expression because transcription occurs predominantly from the paternal allele in mouse and man (maternal imprinting). Excess IGF-II has detrimental systemic and local effects in vivo, promoting somatic overgrowth and an increased frequency of tumors. IGF2 mRNA is overexpressed in colorectal and many other human cancers. In this paper, we show that altered IGF-II supply modifies intestinal tumor growth. Mice genetically altered in the IGF-II system were combined in crosses with ApcMin/+, a murine model of human familial adenomatous polyposis. Depending on genetic background, ApcMin/+ acquires multiple small intestinal adenoma before becoming moribund with anemia. Mice that express excess IGF-II delivered using a bovine keratin 10 promoter (k10Igf2/+) develop a disproportionate overgrowth of colon, uterus, and skin. Combination with ApcMin/+ leads to a 10-fold increase in the number and the diameter of colon adenoma (P<0.0001) compared to ApcMin/+ littermate controls (postnatal day 80), an increased susceptibility to rectal prolapse (41%), and a histological progression to carcinoma. Mice with reduced IGF-II supply, secondary to the disruption of the paternal Igf2 allele (Igf2+m/ p), are 60% the weight of wild-type littermates. Combination with ApcMin/+ leads to a 3-fold reduction in small intestinal adenoma number (P<0.0001) compared to ApcMin/+ littermate controls (postnatal day 150), and a significant decrease in adenoma diameter (P<0.001). With in situ hybridization, we show that Igf2 was expressed in all adenoma irrespective of IGF-II supply. This suggests that there is an increased maternal allele expression of Igf2 (loss of imprinting) in adenoma which form, despite paternal Igf2 allele disruption. We conclude that IGF II supply is a modifier of intestinal adenoma growth, and we provide genetic evidence for its functional role in colorectal cancer progression. PMID- 10706127 TI - Relating genotype and phenotype in breast cancer: an analysis of the prognostic significance of amplification at eight different genes or loci and of p53 mutations. AB - Breast cancer heterogeneity can be related directly to its variability at the genetic level. Thus, tumor genotyping could be a valuable approach to define breast tumor subtypes. It has been shown that it is possible to delineate subgroups of breast tumors according to specific sets of DNA amplifications. The aim of the present work was to study the prognostic significance of these DNA amplifications. We studied DNA amplification at eight genes or loci (AIB1, CCND1, EMS1, ERBB2, FGFR1, MDM2, MYC, and RMC20C001) as well as p53 mutations in a series of 640 breast cancer patients who had not received presurgical therapy and analyzed the correlations with survival DNA amplification was assessed by Southern blotting and was scored positive when exceeding three to five copies. Mutations in the p53 gene were searched by four-color fluorescent single. strand conformational polymorphism, using an automated sequencer. Of the nine genetic alterations tested, four (CCND1, EMS1, FGFR1, and p53 mutations) showed a significant association with reduced disease-free (DFS) and/or overall survival (OVS) in the unselected set of patients by univariate test. Correlations for p53 were found only when selecting mutations in exons 5 or 7. Analysis of node negative and -positive subgroups of patients showed that MDM2 amplification and p53 mutations bore prognostic significance in node-negative patients, whereas amplification of CCND1, EMS1, and FGFR1 correlated with poor outcome in node positive patients. Multivariate analysis on an unselected set of patients retained significance for the amplification of EMS1, FGFR1, and MDM2 with DFS, of CCND1 with OVS, and of RMC20C001 with both DFS and OVS. Interestingly, stratified analysis according to nodal status confirmed results obtained in the univariate tests: significance of MDM2 amplification and p53 mutations in node-negative and that of CCND1, EMS1, and FGFR1 in node-positive patients. We also observed an association between the number of genetic alterations observed in a tumor and poor prognosis. Patients with two or more amplified loci had a worsened outcome. Strongly correlating coamplifications such as CCND1 and FGFR1, as well as ERBB2 and MYC, were associated with a significant reduction of patient survival, thus indicating cooperative effects. Our data support the idea that genetic alterations in breast cancer are not only helpful for phenotyping purposes, but can also represent powerful prognostic indicators in the clinical practice. PMID- 10706128 TI - Transcriptional regulation of cyclooxygenase-2 gene expression: novel effects of nonsteroidal anti-inflammatory drugs. AB - Cyclooxygenase-2 (COX-2) gene overexpression is suggested to play important roles in colorectal tumorigenesis. Epidemiological studies revealed that nonsteroidal anti-inflammatory drugs (NSAIDs), such as aspirin and sulindac, which inhibit COX activity, reduce colorectal cancer mortality. Current investigations have focused on delineating the molecular mechanisms that regulate COX-2 gene expression and the roles of NSAIDs in cancer chemoprevention. COX-2 catalyzes the production of prostaglandins (PGs) from arachidonic acid (AA), generated by phospholipases A2 (PLA2s), a family of acyl esterases that cause the release of AA from cellular phospholipids. Pancreatic secretory PLA2 (sPLA2), via its receptor (sPLA2R), transcriptionally activates COX-2 gene expression in several cell types, although a specific transcription factor mediating COX-2 expression has not yet been identified. Here, we report that a transcription factor, CCAAT/enhancer-binding protein beta(C/EBPbeta), plays a critical role in sPLA2IB-induced, receptor mediated COX-2 gene expression in MC3T3E1 and NIH3T3 cells. Furthermore, treatment of these cells with NSAIDs in the presence of sPLA2IB appears to potentiate the stimulatory effects on COX-2 mRNA and COX-2 protein expression and a concomitant elevation in PG production. Most significantly, NSAID treatment appears to drastically suppress the production of cytosolic PLA2 (cPLA2) mRNA. The lack of sPLA2IB, sPLA2IIA, and sPLA2V mRNA expression in both NIH3T3 and MC3T3E1 cells suggests that cPLA2 is the most likely enzyme that catalyzes the release of AA, the rate-limiting substrate of COX for the production of PGs. Our results suggest that: (a) sPLA2IB receptor-mediated COX-2 expression is mediated via C/EBPbeta; (b) NSAIDs in the presence of sPLA2IB potentiate the stimulatory effects of sPLA2IB on COX-2 mRNA expression; and (c) despite the apparent stimulation of COX-2 expression by NSAIDs, they strikingly deprive COX-2 of its substrate, AA, by suppressing cPLA2 mRNA expression. Both AA and PGs regulate many vital biological functions (e.g., motility and invasiveness) that are dysregulated in most cancer cells, and they have profound effects on cellular differentiation. Our results raise the possibility that deprivation of COX-2 of its substrate by the suppression of cPLA2 mRNA expression is an additional mechanism used by NSAIDs to inhibit tumorigenesis. PMID- 10706129 TI - Chromosomal mapping of genes controlling development, histological grade, depth of invasion, and size of rat stomach carcinomas. AB - Rat stomach cancers induced by N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) are widely used as a model of differentiated-type human stomach cancers. ACI/N (ACT) rats are susceptible and BUF/Nac (BUF) rats are resistant to MNNG-induced stomach carcinogenesis, and the presence of an autosomal gene with a dominant BUF allele has been suggested. In this study, we performed a carcinogenicity test by giving MNNG in drinking water to 117 male ACI x (ACIxBUF)F1 backcross rats. Each of 100 effective rats was diagnosed for its "carcinoma development" and when it was bearing stomach carcinoma(s), for histological grade, depth of invasion, and size and number of tumors. Carcinoma development was diagnosed based both on the age of the rat and on the presence of stomach carcinoma(s). Linkage analysis was performed with the genotypes of 161 loci, covering 1637 cM of the rat genome. Contrary to our original expectations, the most influential gene was the one on chromosome (chr.) 15, Gastric cancer susceptibility gene 1 (Gcs1), which confers susceptibility to stomach carcinogenesis (LOD, 3.8) with a dominant BUF allele by promoting conversion from adenomas to carcinomas. Two resistance genes on chr. 4 and chr. 3, Gastric cancer resistance gene 1 (Gcr1) and Gcr2, were shown to confer dominant resistance (LOD, 2.7 and 2.6, respectively). Gcs1, Gcr1, and Gcr2 exerted additive effects on the development of stomach carcinomas. A gene on chr. 16, Gcr3, was indicated to reduce the depth of invasion (LOD, 2.2) and sizes of tumors (LOD, 1.9). No linkage was obtained using the number of tumors. These findings show that the coordinate effect of a susceptibility gene, Gcs1, and two resistance genes, Gcr1 and Gcr2, is responsible for the development of MNNG induced stomach carcinomas and that Gcr3 is responsible for the growth of a stomach carcinoma, reflected in the depth of invasion and in the tumor size. PMID- 10706130 TI - The c-Fes protein-tyrosine kinase suppresses cytokine-independent outgrowth of myeloid leukemia cells induced by Bcr-Abl. AB - The c-Fes protein-tyrosine kinase exhibits strong expression in myeloid hematopoietic cells. Previous studies have shown that Fes induces differentiation in the chronic myelogenous leukemia-derived cell line K-562, suggesting that the Fes signal for differentiation is dominant to the Bcr-Abl signal for transformation in these cells. In addition, Fes has been shown to associate with and phosphorylate Bcr on NH2-terminal sequences retained within Bcr-Abl. To determine whether Fes interacts directly with Bcr-Abl, kinase-inactive Bcr-Abl was coexpressed with Fes in 293T cells, and phosphorylation was assessed by anti phosphotyrosine immunoblotting. Bcr-Abl was strongly phosphorylated by Fes under these conditions, suggestive of direct interaction. Similarly, tyrosine phosphorylation of kinase-inactive Fes was observed after coexpression with active Bcr-Abl. To test for the interaction of Fes with Bcr-Abl under physiological conditions, wild-type and kinase-defective Fes were stably expressed in the cytokine-dependent myeloid leukemia cell line, DAGM. Expression of either form of Fes alone did not affect the proliferation or interleukin 3 dependence of these cells. The DAGM/Fes cells were then infected with Bcr-Abl retroviruses, and their rates of cytokine-independent outgrowth were compared. Fes dramatically suppressed Bcr-Abl-induced DAGM cell outgrowth relative to a cell line expressing beta-galactosidase as a negative control. This effect required Fes tyrosine kinase activity, because the kinase-inactive form of Fes did not affect Bcr-Abl-induced cell outgrowth. The phosphotyrosine content of both wild-type and kinase-inactive Fes was strongly enhanced after coexpression with Bcr-Abl in DAGM cells, similar to the 293T result. Phosphorylation of wild type Fes correlated with stimulation of Fes tyrosine kinase activity in the presence of Bcr-Abl. These results show that Fes and Bcr-Abl interact in myeloid cells, leading to Fes activation and suppression of Bcr-Abl-induced conversion to cytokine independence. PMID- 10706131 TI - The Mr 193,000 vault protein is up-regulated in multidrug-resistant cancer cell lines. AB - Vaults are 13 megadalton ribonucleoprotein particles composed largely of the major vault protein (MVP) and two high molecular weight proteins, p240 and p193, and a small vault RNA (vRNA). Increased levels of MVP expression, vault associated vRNA, and vaults have been linked directly to multidrug resistance (MDR). To further define the putative role of vaults in MDR, we produced monoclonal antibodies against the Mr 193,000 vault protein and studied its expression levels in various multidrug-resistant cell lines. We find that, like MVP, p193 mRNA and protein levels are increased in various multidrug-resistant cell lines. Subcellular fractionation of vault particles revealed that vault associated p193 levels are increased in multidrug-resistant cells as compared with the parental, drug-sensitive cells. Furthermore, protein analysis of postnuclear supernatants and co-immunoprecipitation studies show that drug sensitive MVP-transfected tumor cells lack this up-regulation in vault-associated p193. Our observations indicate that vault formation is limited not only by the expression of the MVP but also by the expression or assembly of at least one of the other vault proteins. PMID- 10706132 TI - Direct in vitro evidence and in vivo analysis of the antiangiogenesis effects of interleukin 12. AB - As an antitumor agent, interleukin-12 (IL-12) has been revealed to be a key regulator of the immune response, particularly that involving CTL and natural killer (NK) cells. We report herein the antiangiogenesis effect of IL-12 on human as well as murine tumors in NK-depleted severe-combined immunodeficient mice using fibroblasts genetically engineered to secrete this cytokine. Although the in vitro growth of tumor cells was not affected by the presence of IL-12, coinoculation of IL-12-secreting fibroblasts strongly inhibited tumor growth in immunodeficient mice. The neovascularization surrounding the tumor was remarkably inhibited in the area in which the IL-12-secreting fibroblasts were implanted, resulting in the suppression of tumor growth. Lectin staining in tumor sample sections also showed a significant reduction in the number of vessels. The RNA expression of IFN-gamma and its inducible antiangiogenic chemokine IFN gamma inducible protein 10 was stimulated in endothelial cells cultured with IL-12. It was also found that IL-12 down-regulated the expression of the endothelial cell mitogens vascular endothelial growth factor and basic fibroblast growth factor. The antitumor effects of IL-12 were accompanied by interesting histological changes consisting of a high degree of keratinization and apoptosis and a decrease in the proliferation rate of human tumors and extensive necrosis in the murine ones. PMID- 10706133 TI - Nuclear matrix of calreticulin in hepatocellular carcinoma. AB - Nuclear matrix protein profiles of malignant cells vary from their normal counterparts. By two-dimensional gel electrophoresis, we analyzed nuclear matrix proteins in 11 hepatocellular carcinomas and compared them with corresponding non neoplastic liver tissue. Although the compositions were mostly similar, several peptides were noted predominantly in the former. The most prominent one was an acidic protein of apparent Mr 62,000, which was identified to be calreticulin upon NH2-terminal amino acid sequencing. By immunoblotting, calreticulin was confirmed to be present abundantly in the nuclear matrix fraction of carcinomas but not in that of the nonmalignant liver tissue. Interestingly, the total content of calreticulin was similar between them. By immunofluorescence microscopy, evident nuclear immunostaining was detected in carcinomas. Calreticulin was also found to be in the nuclear matrices of various carcinoma cell lines. We conclude that calreticulin is a component of the nuclear matrix. The formation and/or expansion of the calreticulin-nuclear matrix may be related to the activated cell growth. PMID- 10706134 TI - Epidermal growth factor-like ligands differentially up-regulate matrix metalloproteinase 9 in head and neck squamous carcinoma cells. AB - Head and neck squamous cell carcinomas (HNSCCs) are characterized by a marked propensity for local invasion and dissemination to cervical lymph nodes, with distant metastases developing in 30-40% of cases. Overexpression of the epidermal growth factor receptor (EGFR/c-erbB-1) and/or its ligands and high levels of certain matrix metalloproteinases (MMPs) have been associated with poor prognosis. The aim of this study was to examine the effects of EGFR ligands on gelatinase expression and invasion in HNSCC cell lines. We tested epidermal growth factor (EGF), transforming growth factor alpha, betacellulin, heparin binding EGF, and amphiregulin and measured expression of gelatinases MMP-9 and MMP-2 in an established squamous carcinoma cell line (Detroit-562) and in two cell lines newly derived from patients with head and neck cancers (SIHN-005A and SIHN-006). Incubation of the cell lines with EGF-like ligands up-regulated MMP-9 (but not MMP-2) expression as measured by semiquantitative reverse transcription PCR in a dose-dependent manner, with the effects being most marked in cells with high EGFR levels and undetectable in cells with low levels. Maximum stimulation was obtained in a concentration range of 10-100 nM. In addition, we confirmed by zymography that gelatinolytic activity consistent with MMP-9 (Mr 92,000) was up regulated in parallel with increases in gene expression. Betacellulin (which binds both to EGFR and c-erbB-4 receptors) consistently increased MMP-9 expression and activation to a significantly greater degree than the other four ligands when tested at equimolar concentrations. In parallel with MMP-9 up regulation, all EGF-like ligands increased tumor cell invasion through Matrigel in in vitro Transwell assays. These activities were independent of ligand effects on cell proliferation. Antagonist (ICR62) or agonist (ICR9) anti-EGFR monoclonal antibodies, respectively, inhibited or potentiated MMP-9 activity and tumor cell invasion induced by all ligands. Furthermore, a monoclonal antibody that neutralizes MMP-9 activity (Abl) also inhibited ligand-induced invasion of HNSCC. We confirmed that tumor cell lines used in these studies (and a larger series not reported here) generally expressed multiple c-erbB receptors and ligands. These results indicate that autocrine or paracrine signaling through EGFR potentiates the invasive potential of HNSCC via the selective up-regulation and activation of MMP-9. Furthermore, ligands such as betacellulin (which is commonly expressed in HNSCC), which can bind to and activate other c-erbB receptors, may be especially potent in this regard. PMID- 10706135 TI - Induction of differentiation and apoptosis by ligands of peroxisome proliferator activated receptor gamma in non-small cell lung cancer. AB - The peroxisome proliferator-activated receptor gamma (PPARgamma) is a ligand activated transcription factor belonging to the steroid receptor superfamily. It is a key regulator of adipogenic differentiation, the ligands of which have also been demonstrated to induce differentiation in human breast and colon cancer cell lines. This study examined PPARgamma, in non-small cell lung cancer (NSCLC). PPARgamma mRNA and protein were expressed in NSCLC cell lines, with highest levels in adenocarcinomas. PPARgamma protein was also expressed in 50% of primary lung cancers by immunohistochemistry. Treatment of multiple cell lines with two distinct PPARgamma ligands in the presence of serum resulted in growth arrest, irreversible loss of capacity for anchorage-independent growth, decreased activity and expression of matrix metalloproteinase 2, and modulation of multiple markers in a manner consistent with differentiation. Specifically, there was up regulation of general markers of the differentiated state such as gelsolin, Mad, and p21. Down-regulation of specific markers of progenitor lineages for the peripheral lung, i.e., the type II pneumocyte lineage markers MUC1 and surfactant protein-A and the Clara cell lineage marker CC10, also occurred. In addition, HTI56, a marker of terminally differentiated type I pneumocytes, was also induced. Consistent with a more mature, less malignant phenotype, ligand treatment also inhibited the expression of cyclin D1 and led to hypophosphorylation of the retinoblastoma protein. In contrast, in the absence of serum, ligand treatment rapidly resulted in apoptosis and substantially earlier onset of differentiation. Taken together, these results show that depending on the growth milieu, ligands of PPARgamma induce differentiation and apoptosis in NSCLC, suggesting clinical utility for these agents. PMID- 10706136 TI - SN-1, a novel leukemic cell line with t(11;16)(q23;p13): myeloid characteristics and resistance to retinoids and vitamin D3. AB - The MLL gene is fused with the cAMP-responsive element binding protein-binding protein (CBP) gene in t(11;16)(q23;p13), which has been reported to be associated with therapy-related acute leukemia. We established a novel myeloid cell line, SN 1, from a patient with T-cell acute lymphoblastic leukemia with t(11;16)(q23;p13) having in-frame MLL-CBP fusion transcripts. The majority of the SN-1 cells were positive for myeloperoxidase when examined using an electron microscope and expressed CD13, CD33, CD56, and HLA-DR antigens, but not CD7, CD10, CD19, CD34, or CD41 antigens, suggesting that these cells are of myeloid origin. SN-1 cells underwent functional and morphological differentiation when treated with actinomycin D or sodium butyrate, but not with all-trans-retinoic acid (ATRA) or 1alpha,25-dihydroxyvitamin D3 (VD3). Exposure of SN-1 cells to ATRA hardly affected cell growth and differentiation, whereas the growth of HL-60 and NB4 cells treated with ATRA was effectively inhibited, and differentiation into mature granulocytes was induced. SN-1 cells were relatively insensitive to VD3 with respect to inhibiting the cell growth and inducing the ability to reduce nitroblue tetrazolium, lysozyme activity, and morphological differentiation, although the expression of CD11b was slightly induced by VD3. These results suggest that the cell line was impaired in the signal transduction systems of ATRA and VD3. This cell line should be useful for the study of the role of CBP as a transcriptional regulator in leukemia differentiation and for the functional analysis of the MLL-CBP fusion gene, which will provide new insights into leukemogenesis caused by 11q23 translocations. PMID- 10706137 TI - Correspondence re: S. Yamamoto et al., Specific p53 gene mutations in urinary bladder epithelium after the Chernobyl accident. Cancer Res., 59: 3606-3609, 1999. PMID- 10706138 TI - Parallels of craniofacial maldevelopment in Down syndrome and Ts65Dn mice. AB - Mouse genetic models can be used to dissect molecular mechanisms that result in human disease. This approach requires detection and demonstration of compelling parallels between phenotypes in mouse and human. Ts65Dn mice are at dosage imbalance for many of the same genes duplicated in trisomy 21 or Down syndrome (DS), the most common live-born human aneuploidy. Analysis of the craniofacial skeleton of Ts65Dn mice using three-dimensional morphometric methods demonstrates an absolute correspondence between Ts65Dn and DS craniofacial dysmorphology, a distinctive and completely penetrant DS phenotype. The genes at dosage imbalance in Ts65Dn are localized to a small region of mouse chromosome 16 and, by comparative mapping, to the corresponding region of human Chromosome 21, providing independent experimental data supporting the contribution of genes in this region to this characteristic DS phenotype. This analysis establishes precise parallels in human and mouse skull phenotypes resulting from dosage imbalance for the same genes, revealing strong conservation of the evolved developmental genetic program that underlies mammalian skull morphology and validating the use of this mouse model in the analysis of this important DS phenotype. This evolutionary conservation further establishes the mouse as a valid model for a wide range of syndromes producing craniofacial maldevelopment. PMID- 10706139 TI - BMP-4 affects the differentiation of metanephric mesenchyme and reveals an early anterior-posterior axis of the embryonic kidney. AB - Bone morphogenetic protein-4 (BMP4), a member of the transforming growth factor beta (TGF-beta) family, regulates several developmental processes during animal development. We have now studied the effects of BMP-4 in the metanephric kidney differentiation by using organ culture technique. Human recombinant BMP-4 diminishes the number of ureteric branches and changes the branching pattern. Our data suggest that BMP-4 affects the ureteric branching indirectly via interfering with the differentiation of the nephrogenic mesenchyme. The clear positional preference of the defects to posterior mesenchyme might reflect an early anterior posterior patterning of the metanephric mesenchyme. The smooth muscle alpha-actin expressing cell population around the ureteric stalk, highly expressing Bmp-4 mRNA, is also expanded in kidneys treated with BMP-4. Thus, BMP-4 may be a physiological regulator of the development of the periureteric smooth muscle layer and ureteric elongation. PMID- 10706140 TI - Tissue interactions mediate early events in pulmonary vasculogenesis. AB - Extensive study has provided considerable insight into the mechanisms governing branching morphogenesis and developmental maturation of the pulmonary epithelium. The process by which the vascular tree arises in the mesodermal mesenchyme of the developing lung, however, is not known. Because normal epithelial branching and differentiation have been shown to be dependent on interactions with the lung mesenchyme, we hypothesized that the developing pulmonary vasculature is dependent on a reciprocal interaction with pulmonary epithelium. In this study we have defined the temporal and spatial expression of flk-1 mRNA, which encodes an endothelial cell-specific vascular endothelial growth factor (VEGF) receptor, in fetal and neonatal rat lung. Flk-1-positive cells were observed in the lung at every prenatal stage from fetal day 11 through birth, demonstrating that vascularization has been initiated as soon as the lung evaginates from the foregut epithelium. The spatial distribution of vascular precursors was distinct and consistent in early lung (fetal days 11-16): clusters of flk-1-positive cells were localized in the mesenchyme closely apposed to the developing epithelium. This spatial relationship between vascular precursors and the developing epithelium suggested that vascular development in the lung may be dependent on interactions between the two tissue types. To investigate this possibility, day 13 distal lung mesenchyme was cultured in the presence and absence of lung epithelium. Lung mesenchyme cultured in the absence of epithelium degenerated significantly, and few flk-1-positive cells were maintained. In contrast, lung mesenchyme recombined with lung epithelium contained abundant flk-1-positive cells, and their spatial distribution mimicked that observed in vivo. These studies provide the first detailed information regarding the temporal and spatial pattern of pulmonary vascularization in early development and suggest that tissue interactions play an important role in growth and maintenance of the developing lung vasculature. PMID- 10706141 TI - Chemotactic migration of mesencephalic neural crest cells in the mouse. AB - We examined the roles of fibroblast growth factor (FGF)-2 and FGF-8 in the migration of mesencephalic mouse neural crest cells. Our in vitro migration assay has shown that FGF-2 (basic FGF) and FGF-8 have chemotactic activity for these cells. Chemotaxis was inhibited by anti-FGF-2 and anti-FGF-8 neutralizing antibodies. In addition, anti-FGF-2 blocked neural crest cell migration in cranial organ cultures. This observation suggests that FGF-2 functions as a chemoattractant in migration of mesencephalic neural crest cells in vivo. In organ culture, the antagonist of FGF binding to a low-affinity fibroblast growth factor receptor (FGFR) heparan sulfate, inositolhexakisphosphate (InsP6), inhibited migration as well. Mesencephalic neural crest cells had high-affinity FGFRs, in particular FGFR-1 and FGFR-3. Thus, the chemotactic activities of FGF-2 can be mediated by the low-affinity FGFR alone or by a combination of low- and high-affinity FGFRs (FGFR-1, FGFR-3, or both). Moreover, differential localization of FGF-2 was found at the mesencephalic axial level of intact embryos during neural crest cell migration. FGF-2 protein expression was predominant in the target regions, in particular the mandibular mesenchyme, that are colonized by mesencephalic neural crest cells. This characteristic distribution supports the notion that FGF-2 acts as a chemoattractant in the mouse embryo that directs mesencephalic neural crest cell migration. Whereas FGF 8 showed chemotactic activity in vitro, neural crest cell dispersion was observed in explants that had been treated with anti-FGF-8 neutralizing antibodies. This result suggests that FGF-8 may not be a chemoattractant in vivo. However, the distribution of neural crest cells in explants treated with anti-FGF-8 differed from that in control explants or in intact embryos. Extreme FGF-2 distribution was observed in the mandibular arch and FGF-8 is expressed in the epithelium. FGF 8 may play a role in mesencephalic neural crest cell migration, and its role may be concerned with the differential localization of FGF-2. To establish this notion, we performed immunohistochemical examination of FGF-2 distribution in explants treated with FGF-8 and analysis of FGF-2 gene expression levels by reverse transcriptase-polymerase chain reaction by using RNA from explants. The data indicate that FGF-2 is distributed throughout the mesenchyme in FGF-8 treated explants and that expression of FGF-2 is promoted by FGF-8. Therefore, we conclude that the expression of FGF-8 in the mandibular arch epithelium is a prerequisite for the differential localization of FGF-2 and that the FGF-2 distribution pattern is essential for chemotaxis of mesencephalic neural crest cell migration. PMID- 10706142 TI - Inhibition of distal lung morphogenesis in Nkx2.1(-/-) embryos. AB - In vitro and in vivo results are consistent with a critical role for NKX2.1, an epithelial homeodomain transcription factor in lung morphogenesis. Nkx2.1 null mutant embryos die at birth due to respiratory insufficiency caused by profoundly abnormal lungs. However, the precise role of NKX2.1 in the multistep process of lung structural morphogenesis and differentiation of various pulmonary cell types remains unknown. In the current study, we tested the hypothesis that the mutant lungs do not undergo branching morphogenesis beyond the formation of the mainstem bronchi and therefore consist solely of dilated tracheobronchial structures. To test this hypothesis, we determined the spatial and temporal expression pattern of a number of extracellular matrix (ECM) proteins and their cellular receptors, including alpha-integrins, laminin, and collagen type IV. Although laminin is expressed in the mutant Nkx2.1(-/-) lungs, expression of alpha-integrins and collagen type IV is significantly reduced or absent. In addition, examination of regionally specific expression of differentially spliced Vegf (vascular endothelial growth factor) transcripts, clearly indicates that the epithelial phenotype of the Nkx2.1(-/-) lungs is similar to the tracheobronchial epithelium. In contrast to wild-type lungs in which both Vegf1 and Vegf3 are developmentally expressed, Nkx2.1(-/-) lungs are characterized by predominant expression of Vegf1 and reduced or absent Vegf3. A similar pattern of Vegf expression is also observed in isolated tracheo-bronchial tissue. The sum of these findings suggest that at least two separate pathways may exist in embryonic lung morphogenesis: proximal lung morphogenesis is Nkx2.1 independent, while distal lung morphogenesis appears to be strictly dependent on the wild-type activity of Nkx2.1. PMID- 10706143 TI - Distribution of different regions of cardiac neural crest in the extrinsic and the intrinsic cardiac nervous system. AB - In this study we focused upon whether different levels of postotic neural crest as well as the right and left cardiac neural crest show a segmented or mixed distribution in the extrinsic and intrinsic cardiac nervous system. Different parts of the postotic neural crest were labeled by heterospecific replacement of chick neural tube by its quail counterpart. Quail-chick chimeras (n = 21) were immunohistochemically evaluated at stage HH28+, HH29+, and between HH34-37. In another set of embryos, different regions of cardiac neural crest were tagged with a retrovirus containing the LacZ reporter gene and evaluated between HH35-37 (n = 13). The results show a difference in distribution between the right- and left-sided cardiac neural crest cells at the arterial pole and ventral cardiac plexus. In the dorsal cardiac plexus, the right and left cardiac neural crest cells mix. In general, the extrinsic and intrinsic cardiac nerves receive a lower contribution from the right cardiac neural crest compared with the left cardiac neural crest. The right-sided neural crest from the level of somite 1 seeds only the cranial part of the vagal nerve and the ventral cardiac plexus. Furthermore, the results show a nonsegmented overlapping contribution of neural crest originating from S1 to S3 to the Schwann cells of the cranial and recurrent nerves and the intrinsic cardiac plexus. Also the Schwann cells along the distal intestinal part of the vagal nerve are derived exclusively from the cardiac neural crest region. These findings and the smaller contribution of the more cranially emanating cardiac neural crest to the dorsal cardiac plexus compared with more caudal cardiac neural crest levels, suggests an initial segmented distribution of cardiac neural crest cells in the circumpharyngeal region, followed by longitudinal migration along the vagal nerve during later stages. PMID- 10706144 TI - Characterization of two amphioxus Wnt genes (AmphiWnt4 and AmphiWnt7b) with early expression in the developing central nervous system. AB - Full-length sequences and developmental expression patterns of two amphioxus Wnt genes (AmphiWnt4 and AmphiWnt7b) are described for the first time. The dynamic expression pattern of AmphiWnt4 suggests roles in the development of the posterior mesoderm, central nervous system, muscular somites, heart, and endostyle (a homolog of the vertebrate thyroid). The less diverse expression domains of AmphiWnt7b indicate that this gene may be involved only in the development of the central nervous system and the endostyle. In contrast to amphioxus, vertebrate embryos do not express Wnt4 homologues in the posterior mesoderm, somites, or heart; instead, Wnt genes of other subfamilies are expressed in these developing vertebrate organs. Because the developmental genetic programs of amphioxus may approximate those in the invertebrate chordate ancestor of the vertebrates, it is possible that some developmental functions of an ancestral Wnt4 gene may have been assumed by genes of other Wnt subfamilies during vertebrate evolution, possibly as a result of functional redundancy among Wnt subfamilies. PMID- 10706145 TI - Apical epithelial cap morphology and fibronectin gene expression in regenerating axolotl limbs. AB - Urodele amphibians (salamanders) are unique among adult vertebrates in their ability to regenerate limbs. The regenerated structure is often indistinguishable from the developmentally produced original. Thus, the two processes by which the limb is produced - development and regeneration - are likely to use many conserved biochemical and developmental pathways. Some of these limb features are also likely to be conserved across vertebrate families. The apical ectodermal ridge (AER) of the developing amniote limb and the larger apical epithelial cap (AEC) of the regenerating urodele limb are both found at the limb's distalmost tip and have been suggested to be functionally similar even though their morphology is quite different. Both structures are necessary for limb outgrowth. However, the AEC is uniformly smooth and thickly covers the entire limb-tip, unlike the AER, which is a protruding ridge covering only the dorsoventral boundary. Previous data from our laboratory suggest the multilayered AEC may be subdivided into separate functional compartments. We used hematoxylin and eosin (H+E) staining as well as in situ hybridization to examine the basal layer of the AEC, the layer that lies immediately over the distal limb mesenchyme. In late stage regenerates, this basal layer expresses fibronectin (FN) message very strongly in a stripe of cells along the dorso-ventral boundary. H+E staining also reveals the unique shape of basal cells in this area. The stripe of cells in the basal AEC also contains the notch/groove structure previously seen in avian and reptilian AERs. In addition, AEC expression of FN message in the cells around the groove correlates with previous amniote AER localization of FN protein inside the groove. The structural and biochemical analyses presented here suggest that there is a specialized ridge-like compartment in the basal AEC in late-stage regenerates. The data also suggest that this compartment may be homologous to the AER of the developing amniote limb. Thus, the external differences between amniote limb development and urodele limb regeneration may be outweighed by internal similarities, which enable both processes to produce morphologically complete limbs. In addition, we propose that this basal layer of the AEC is uniquely responsible for AEC functions in regeneration, such as secreting molecules to promote mesenchymal cell cycling and dictating the direction of limb outgrowth. Finally, we include here a clarification of existing nomenclature to facilitate further discussion of the AEC and its basal layer. PMID- 10706146 TI - YAC transgene-mediated olfactory receptor gene choice. AB - In the mouse, individual olfactory neurons express one of a thousand distinct olfactory receptor genes. Furthermore, only one allele of the expressed gene is transcribed. This phenomenon, random allelic inactivation, along with the observation that the olfactory receptor genes reside in large chromosomal arrays, suggests a role for long-range gene regulation in olfactory receptor gene choice. We have constructed a 300-kb yeast artificial chromosome (YAC) transgene in which a single receptor gene is marked while maintaining its coding region. This 300-kb piece of DNA functions as an independent olfactory receptor gene locus in directing olfactory receptor gene choice in both the olfactory system and the accessory olfactory system (vomeronasal organ, VNO). Furthermore, the transgene, like endogenous olfactory receptor loci, is subject to allelic inactivation. PMID- 10706147 TI - NIAID renews funding for the adult AIDS clinical trials group. PMID- 10706148 TI - Health assessment of the geriatric patient. PMID- 10706149 TI - Genetic testing. PMID- 10706150 TI - Recommended air travel delay in patients with otitis media. PMID- 10706151 TI - Sun protection for children. PMID- 10706152 TI - Osteoporosis and fractures. PMID- 10706153 TI - The impact of genetic testing on primary care: where's the beef? PMID- 10706154 TI - Evaluation of the acutely limping child. AB - A limp may be defined as any asymmetric deviation from a normal gait pattern. The differential diagnosis of a limp includes trauma, infection, neoplasia and inflammatory, congenital, neuromuscular or developmental disorders. Initially, a broad differential diagnosis should be considered to avoid overlooking less common conditions such as diskitis or psoas abscess. In any patient with a complaint of knee or thigh pain, an underlying hip condition should be considered. The patient's age can further narrow the differential diagnosis, because certain disease entities are age-specific. Vigilance is warranted in conditions requiring emergent treatment such as septic hip. The challenge to the family physician is to identify the cause of the limp and determine if further observation or immediate diagnostic work-up is indicated. PMID- 10706155 TI - A primary care approach to the patient with claudication. AB - Peripheral arterial occlusive disease occurs in about 18 percent of persons over 70 years of age. Usually, patients who have this disease present with intermittent claudication with pain in the calf, thigh or buttock that is elicited by exertion and relieved with a few minutes of rest. The disease may also present in a subacute or acute fashion. Symptoms of ischemic rest pain, ulceration or gangrene may be present at the most advanced stage of the disease. In most cases, the underlying etiology is atherosclerotic disease of the arteries. In caring for these patients, the primary care physician should focus on evaluation, risk factor modification and exercise. The physician should consider referral to a vascular subspecialist when symptoms progress or are severe. While the prognosis for the affected limb is quite good, patients with peripheral arterial occlusive disease are at increased risk of myocardial infarction and stroke. Therefore, treatment measures should address overall vascular health. PMID- 10706156 TI - Meckel's diverticulum. AB - Although Meckel's diverticulum is the most prevalent congenital abnormality of the gastrointestinal tract, it is often difficult to diagnose. It may remain completely asymptomatic, or it may mimic such disorders as Crohn's disease, appendicitis and peptic ulcer disease. Ectopic tissue, found in approximately 50 percent of cases, consists of gastric tissue in 60 to 85 percent of cases and pancreatic tissue in 5 to 16 percent. The diagnosis of Meckel's diverticulum should be considered in patients with unexplained abdominal pain, nausea and vomiting, or intestinal bleeding. Major complications include bleeding, obstruction, intussusception, diverticulitis and perforation. The most useful method of diagnosis is with a technetium-99m pertechnetate scan, which is dependent on uptake of the isotope in heterotopic tissue. Management is by surgical resection. PMID- 10706157 TI - Thyroiditis: differential diagnosis and management. AB - Thyroiditis is a group of inflammatory thyroid disorders. Patients with chronic lymphocytic thyroiditis (also referred to as Hashimoto's thyroiditis) present with hypothyroidism, goiter, or both. Measurement of serum thyroid autoantibodies and thyroglobulin confirms the diagnosis. Subacute granulomatous thyroiditis (sometimes referred to as de Quervain's disease) is a self-limited but painful disorder of the thyroid. Physical examination, elevated erythrocyte sedimentation rate, elevated thyroglobulin level and depressed radioactive iodine uptake (RAIU) confirm the diagnosis. Subacute lymphocytic thyroiditis (silent thyroiditis) is considered autoimmune in origin and commonly occurs in the postpartum period. Symptoms of hyperthyroidism and depressed RAIU predominate. Acute (suppurative) thyroiditis is a rare, infectious thyroid disorder caused by bacteria and other microbes. The rare, invasive fibrous thyroiditis (Riedel's thyroiditis) presents with a slowly enlarging anterior neck mass that is sometimes confused with a malignancy. PMID- 10706158 TI - Identification and evaluation of mental retardation. AB - Mental retardation in young children is often missed by clinicians. The condition is present in 2 to 3 percent of the population, either as an isolated finding or as part of a syndrome or broader disorder. Causes of mental retardation are numerous and include genetic and environmental factors. In at least 30 to 50 percent of cases, physicians are unable to determine etiology despite thorough evaluation. Diagnosis is highly dependent on a comprehensive personal and family medical history, a complete physical examination and a careful developmental assessment of the child. These will guide appropriate evaluations and referrals to provide genetic counseling, resources for the family and early intervention programs for the child. The family physician is encouraged to continue regular follow-up visits with the child to facilitate a smooth transition to adolescence and young adulthood. PMID- 10706159 TI - Somatizing patients: Part I. Practical diagnosis. AB - The phenomenon of somatization, which results in unexplained physical complaints, is ubiquitous in primary care settings although it often goes unrecognized. Medical training emphasizes the identification and treatment of organic problems and may leave physicians unprepared to recognize and address somatoform complaints. As a process, somatization ranges from mild stress-related symptoms to severe debilitation. Patients at the low end of the spectrum often respond to simple reassurance, but patients who are more impaired require interventions specifically designed to avoid unnecessary exposure to dangerous, costly and frustrating diagnostic procedures and treatments. PMID- 10706160 TI - Ectopic pregnancy. AB - Ectopic pregnancy occurs at a rate of 19.7 cases per 1,000 pregnancies in North America and is a leading cause of maternal mortality in the first trimester. Greater awareness of risk factors and improved technology (biochemical markers and ultrasonography) allow ectopic pregnancy to be identified before the development of life-threatening events. The evaluation may include a combination of determination of urine and serum human chorionic gonadotropin (hCG) levels, serum progesterone levels, ultrasonography, culdocentesis and laparoscopy. Key to the diagnosis is determination of the presence or absence of an intrauterine gestational sac correlated with quantitative serum beta-subunit hCG (beta-hCG) levels. An ectopic pregnancy should be suspected if transvaginal ultrasonography shows no intrauterine gestational sac when the beta-hCG level is higher than 1,500 mlU per mL (1,500 IU per L). If the beta-hCG level plateaus or fails to double in 48 hours and the ultrasound examination fails to identify an intrauterine gestational sac, uterine curettage may determine the presence or absence of chorionic villi. Although past treatment consisted of an open laparotomy and salpingectomy, current laparoscopic techniques for unruptured ectopic pregnancy emphasize tubal preservation. Other treatment options include the use of methotrexate therapy for small, unruptured ectopic pregnancies in hemodynamically stable patients. Expectant management may have a role when beta hCG levels are low and declining. PMID- 10706161 TI - The geriatric patient: a systematic approach to maintaining health. AB - The number of persons 65 years of age and older continues to increase dramatically in the United States. Comprehensive health maintenance screening of this population is becoming an important task for primary care physicians. As outlined by the U.S. Preventive Services Task Force, assessment categories unique to elderly patients include sensory perception and injury prevention. Geriatric patients are at higher risk of falling for a number of reasons, including postural hypotension, balance or gait impairment, polypharmacy (more than three prescription medications) and use of sedative-hypnotic medications. Interventional areas that are common to other age groups but have special implications for older patients include immunizations, diet and exercise, and sexuality. Cognitive ability and mental health issues should also be evaluated within the context of the individual patient's social situation-not by screening all patients but by being alert to the occurrence of any change in mental function. Using an organized approach to the varied aspects of geriatric health, primary care physicians can improve the care that they provide for their older patients. PMID- 10706162 TI - HIV transmission in children. PMID- 10706163 TI - Adolescent suicide and household access to firearms in Colorado: results of a case-control study. AB - PURPOSE: To determine whether, compared with age- and sex-matched controls who did not commit suicide, adolescents who committed suicide by firearms were more likely to have had household access to firearms (after adjusting for significant risk factors for adolescent suicide). METHODS: A case-control study design was used; case subjects were Colorado adolescents who committed suicide between 1991 and 1993; controls were sex- and age-matched adolescents who were randomly selected from the same school the subjects had attended. Interviews were conducted with the parent or guardian of cases and controls. RESULTS: Of the 36 case subjects in this study, 67% committed suicide using a gun obtained from their home. Adolescent suicide victims who committed suicide by firearms were significantly more likely to have a firearm in their home (72%) than age- and sex matched community controls (50%), after adjusting for significant risk factors. Conduct disorder and previous mental health treatment were also found to be independent risk factors for adolescent firearm suicide. CONCLUSIONS: Two types of public health interventions to prevent adolescent firearm suicides are likely to be successful: (a) limiting household access to firearms, and (b) identifying adolescents at high risk of firearm suicide. PMID- 10706164 TI - Common themes from the extremes: using two methodologies to examine adolescents' perceptions of anti-violence public service announcements. AB - PURPOSE: To determine in what ways adolescents perceive public service announcements (PSAs) in general and, more specifically, anti-violence health messages. METHODS: Seventy-nine adolescents who were involved with the issue of violence (39 pro-social, 40 incarcerated) participated. These youth were from four sites (Philadelphia, Pennsylvania, Detroit, Michigan, Albuquerque, New Mexico, and Portland, Oregon) and were chosen at random from a pool recommended by community leaders. First, adolescents were questioned on their demographics and knowledge of, attitudes about, and experience with, violence. Then, adolescents rated eight PSAs on levels of interest, understanding, believability, and perceived effect. In semistructured individual interviews, the adolescents discussed each of the PSAs as well as how health messages can effectively reach young people. We used quantitative and qualitative methodologies to analyze the data. RESULTS: Adolescents had similar opinions about the presented messages and using PSAs. Across both the pro-social and incarcerated groups, adolescents (a) opposed celebrity spokespeople, (b) preferred authentic-looking characters and realistic situations, (c) dismissed messages directed at either younger or older audiences, (d) confused abstractions, (e) focused on visuals, and (f) suggested using graphic images. CONCLUSIONS: The similarities observed between the pro social and incarcerated adolescents may arise from the fact that, although the nature of their experience varied, both groups had high levels of issue involvement. From this study, we can make three recommendations for creating messages: (a) use authentic-looking characters in realistic situations; (b) employ simple, visual, and graphic messages; and (c) do formative evaluations with target audiences. PMID- 10706165 TI - Developmental risk factors for youth violence. AB - PURPOSE: To replicate earlier research findings on risk factors for youth violence and to explore the effects on violent behavior of constructs shown to increase risk for other problem behaviors, within a developmental frame. METHODS: Data were from the Seattle Social Development Project (SSDP), a prospective study involving a panel of youths followed since 1985. Potential risk factors for violence at age 18 years were measured at ages 10, 14, and 16 years. Bivariate relationships involving risk factor constructs in the individual, family, school, peer and community domains and violence were examined at each age to assess changes in their strength of prediction over time. Attention was also given to the additive strength of increasing numbers of risk factors in the prediction of violence at age 18 years. A final set of analyses explored the extent to which youths were correctly classified as having committed a violent act (or not) at age 18 years on the basis of their overall level of risk at ages 10, 14, and 16 years. RESULTS: At each age, risk factors strongly related to later violence were distributed among the five domains. Ten of 15 risk factors constructs measured at age 10 years were significantly predictive of violence at age 18 years. Twenty of 25 constructs measured at age 14 years and 19 of 21 constructs measured at age 16 years were significantly predictive of later violence. Many constructs predicted violence from more than one developmental point. Hyperactivity (parent rating), low academic performance, peer delinquency, and availability of drugs in the neighborhood predicted violence from ages 10, 14, and 16 years. Analyses of the additive effects of risk factors revealed that youths exposed to multiple risks were notably more likely than others to engage in later violence. The odds for violence of youths exposed to more than five risk factors compared to the odds for violence of youths exposed to fewer than two risk factors at each age were seven times greater at age 10 years, 10 times greater at age 14 years, and nearly 11 times greater at age 16 years. However, despite information gained from all significant risk factors, the overall accuracy in predicting youths who would go on to commit violent acts was limited. CONCLUSIONS: Findings from the study have important implications for preventive intervention programs. Prevention efforts must be comprehensive and developmentally sensitive, responding to large groups or populations exposed to multiple risks. PMID- 10706166 TI - Assessment of school-based health centers in a rural state: the West Virginia experience. AB - PURPOSE: To assess the capability of school-based health centers (SBHCs) to provide access to health care for rural youth. METHODS: Review of annual patient records from SBHCs in West Virginia. Ten of 24 SBHCs in West Virginia in operation from July 1994 to June 1997 were selected for the study. Enrollment and utilization rates were generated for each site. A comparison was made between rates of enrollment, utilization, and annual visits among youth with private insurance, those covered by Medicaid, and youth without insurance. Rural and urban SBHCs within West Virginia were compared based on enrollment, utilization, and visit rates. The diagnostic categories were analyzed. Finally, enrollment rates, utilization rates, and insurance status for the West Virginia SBHCs were compared with national norms. RESULTS: Enrollment rates rose steadily during the study period from 27% in Year 1 to 64% by the end of Year 3. The utilization rate was 67% in Year 3. The youth with either Medicaid or no insurance comprised 52% of enrollees, but they accounted for 63% of all visits. West Virginia SBHCs have a higher rate of Medicaid users than the national average for SBHCs, and the annual visit rate for West Virginia youth is higher than the national average for nonmetropolitan adolescents. The enrollment rate of 64% is slightly higher than the national average for SBHCs. Within West Virginia, the enrollment rate in rural schools was significantly higher, 86% compared to 46% (p < .001), and the utilization rate was 70% in rural centers compared to 63% in the urban centers (p < .001). CONCLUSION: When SBHCs are available in rural areas, students use them. In West Virginia, SBHCs have contributed to providing access to health care for rural youth. PMID- 10706167 TI - Longitudinal nutrient intake patterns of US adolescent women: the Penn State Young Women's Health Study. AB - PURPOSE: To use longitudinal nutrient intake data to determine whether dietary patterns remain consistent (or "track") as U.S. females progress from age 12 to 18 years. METHODS: Three-day diet records were collected at regular intervals over 6 years from participants in the Penn State Young Women's Health Study. Eighty-one subjects remained in the cohort during the study period. Tracking in body weight, in dietary intake of fat, sugar, iron, vitamin C, and in a total dietary score (TDS) was assessed using quartile-ranking analysis, year-to-year Pearson correlation analysis, and longitudinal linear analysis. RESULTS: Rank analysis revealed that subjects maintained their relative quartile positions for body weight throughout the study period, and year-to-year correlation coefficients for this variable were .93-.94. In contrast, rank and correlation analyses showed that the subjects did not track strongly with respect to any nutrient variable. Age 12 to 18 years correlation coefficients ranged from r = .04 for fat intake to r = .15 for the TDS. In longitudinal linear models, slopes differed in direction and significance across the original quartiles for nutrient intake, indicating varying dietary trends over time within the study population. CONCLUSIONS: Nutrient intake patterns do not track strongly throughout adolescence among U.S. females. PMID- 10706168 TI - Perceived consequences of teenage childbearing among adolescent girls in an urban sample. AB - PURPOSE: The purpose of this study was to examine the perceived positive consequences of teenage childbearing among female adolescents, and to determine whether perceived consequences of teenage childbearing are associated with other attitudes and sexual risk behaviors. METHODS: The sample consisted of 584 female students attending three urban high schools in Los Angeles, California. The respondents' mean age was 15.8 years, and 72% were Hispanic/Latina. Respondents completed a paper-and-pencil survey assessing their attitudes and risk behaviors relevant to teenage pregnancy. Multiple regression and logistic regression analyses were used to examine the associations between perceived consequences of teenage childbearing and demographic variables, educational variables, parental characteristics, psychosocial variables, attitudes, and sexual behavior. RESULTS: Higher scores on a scale of perceived positive consequences of teenage childbearing were associated with increased risk of sexual intercourse and unprotected sexual intercourse. Higher scores on this scale were found among girls who were Latinas, were non-U.S. natives, had low levels of expected educational attainment, had low parental monitoring, had good communication with parents, and wished to have many children. CONCLUSION: Potential strategies for preventing adolescent pregnancy include educating girls about the difficulties of teenage childbearing, countering their positive illusions about the expected benefits, and teaching them more adaptive ways to meet their emotional needs. PMID- 10706169 TI - Smart teens don't have sex (or kiss much either). AB - PURPOSE: To examine the relationship between an intelligence measure and a wide spectrum of partnered sexual activity ranging from holding hands to sexual intercourse among adolescents. METHOD: Analyses are based on two separate samples of adolescents. The core sample of the National Longitudinal Study of Adolescent Health (Add Health) includes approximately 12,000 adolescents enrolled in the 7th to 12th grades. The Biosocial Factors in Adolescent Development projects followed approximately 100 white males and 200 black and white females over 3- and 2-year periods, respectively. Both studies used the Peabody Picture Vocabulary Test (PPVT) as an intelligence measure, and confidential self-reports of sexual activity. Logistic regression models were used to examine the relationship between PPVT scores and coital status in Add Health data; proportional hazard models were used to examine the timing of initiation of noncoital and coital activities as a function of PPVT scores in the Biosocial Factors sample. RESULTS: Controlling for age, physical maturity, and mother's education, a significant curvilinear relationship between intelligence and coital status was demonstrated; adolescents at the upper and lower ends of the intelligence distribution were less likely to have sex. Higher intelligence was also associated with postponement of the initiation of the full range of partnered sexual activities. An expanded model incorporating a variety of control and mediator variables was tested to identify mechanisms by which the relationship operates. CONCLUSIONS: Higher intelligence operates as a protective factor against early sexual activity during adolescence, and lower intelligence, to a point, is a risk factor. More systematic investigation of the implications of individual differences in cognitive abilities for sexual activities and of the processes that underlie those activities is warranted. PMID- 10706170 TI - Study in sexuality of medical college students in India. AB - PURPOSE: In India, talking about sex is taboo. Little is known about the knowledge, attitude, and sexual behavior of adolescents. This study was carried out with the purpose of examining: (a) the knowledge of medical students about sex, (b) the sources of learning about sex, and (c) the sexual behavior and practices of young adults. METHODS: This study was carried out among the undergraduate students of a medical college in Delhi. A pretested, semiclosed type questionnaire was voluntarily filled out by the students. Confidentiality and secrecy was assured. RESULTS: Of 500 students, 73% participated in the study. Knowledge regarding sexual intercourse, masturbation, contraception, and sexually transmitted diseases was satisfactory among 70%, 74.8%, 83.5%, and 92.6% of the respondents, respectively. Common source of knowledge about sex were friends (74.5%), pornographic films (56.2%), and books and magazines (55.1%). Only one fifth could communicate with teachers, parents, and persons of the other gender about sex. About 417 of the students viewed homosexuality as normal behavior. Sexual intercourse had been experienced by 11.8% of respondents. The mean age of first sexual intercourse was 17.5 years. Eighty-five percent strongly favored introduction of sex education at school level. CONCLUSION: Evidence is provided for the need to improve knowledge about different aspects of sex among a sample of Indian medical students. PMID- 10706171 TI - The process of decision-making on abortion: a grounded theory study of young men in Sweden. AB - PURPOSE: To gain knowledge of the attitudes, questions, and problems in decision making on abortion by young men recently informed about the positive pregnancy tests of their girlfriends. This knowledge will be used as a basis for developing a service for such young men. METHODS: A grounded theory approach was used for interviews at an outpatient clinic for adolescents in Sweden with 18 individuals faced with sharing the decision about abortion. RESULTS: In a suggested model, the decision-making process can be understood by three concepts: reactions (including feelings, apprehensions, and moral conflicts), impact factors (including quality of relationship, consideration for girlfriend, and psychosocial factors), and tools for process (including communication, secrecy/confidentiality, and organized support). CONCLUSIONS: This study suggests the importance of helping the male adolescent with his reactions as a condition for using the equipment constituting impact factors and tools for process on the way to decision about abortion. PMID- 10706172 TI - Intravascular catheter-associated infections. AB - Serious infections associated with intravascular catheters are common. The available data suggests there are likely to be more than 500 000 cases of catheter-associated bloodstream infections occurring annually in Western Europe and the USA. These may be associated with as many as 100 000 deaths. The pathophysiology of this common condition is still not fully elucidated. With catheters that are in place for short periods (a few days), microbial migration down the outer surface of the device to the intravascular tip predominates. For catheters that are in place for longer periods, migration occurs more often via the internal lumen. After being in place for more than 8 days, nearly all central vein catheters will have microorganisms embedded in a biofilm within the catheter lumen. In some catheters, microorganisms will proliferate to sufficient numbers for systemic sepsis to result. The occurrence and rate of this proliferation is dependent on microbial virulence factors, host factors, and characteristics of the catheter. Diagnosis of intravascular device-associated sepsis remains problematic because the pathophysiology of the condition changes with time and because standard culture techniques rarely detect organisms embedded in biofilms. The semiquantitative roll method on blood agar remains in common use because of its simplicity. However, the method only samples the external surface of the catheter. For catheters that have been in place for extended periods of time, methods that better sample the internal lumen, such as sonication and quantitative broth methods, should be developed and used. PMID- 10706174 TI - Possible role of catheters in Saccharomyces boulardii fungemia. AB - Four cases of Saccharomyces boulardii fungemia, a very rare side effect of Saccharomyces boulardii therapy, are reported. The clinical impact of Saccharomyces boulardii infection appeared to be moderate. However, even though organ involvement was never demonstrated, septic shock with no other etiology was observed in one of our patients. All patients had an indwelling vascular catheter. Contamination of the air, environmental surfaces, and hands following the opening of a packet suggests that catheter contamination may have been a source of infection. To prevent catheter contamination it is recommended that packets or capsules of Saccharomyces boulardii be opened with gloves, outside the patient's room. PMID- 10706175 TI - Long-term outcome of acute hepatitis B and C in an outbreak of hepatitis in 1969 72. AB - The objective of this study was to investigate the epidemiology, etiology, and long-term outcome of an extended outbreak of acute hepatitis that occurred in an area of Sweden between 1969 and 1972. The outbreak was analyzed retrospectively by retesting stored frozen serum samples for the presence of hepatitis A, B and C markers. The results were compared with the diagnoses that had been determined during the outbreak. Of 180 patients, 29 (16%) had acute hepatitis A, 126 (70%) had acute hepatitis B, and eight (4.4%) had acute hepatitis C. The Australia antigen test used during the outbreak had failed to identify 21 patients with acute hepatitis B virus infection. Genotyping of the hepatitis B virus strains showed that genotype D was the most prevalent, irrespective of the transmission route. An attempt was made to follow up patients with unresolved hepatitis B virus infection, 25-27 years after the acute infection. None of the 100 patients with acute hepatitis B infection who were traced had become chronic carriers. In ten patients with hepatitis C virus infection, the follow-up showed considerable variation in the outcome, ranging from spontaneous resolution to death through liver cirrhosis. Intravenous drug users had a high prevalence of hepatitis C virus infection, with 52% testing positive for hepatitis C antibodies. PMID- 10706176 TI - New colorimetric microdilution method for in vitro susceptibility testing of Borrelia burgdorferi against antimicrobial substances. AB - A newly developed colorimetric microdilution method was used to analyze the activity of 12 antimicrobial agents against nine Borrelia burgdorferi isolates, including all three genospecies pathogenic for humans. In addition, in vitro antimicrobial resistance patterns of Borrelia valaisiana and Borrelia bissettii tick isolates were investigated. The applied test system is based upon color changes that occur in the presence of phenol red and result from the accumulation of nonvolatile acid produced by actively metabolizing spirochetes. After 72 h of incubation, minimal inhibitory concentrations (MICs) were determined from the decrease of absorbance by software-assisted calculation of growth curves. MIC values were lowest for azlocillin (MIC, < or = 0.125 microg/ml), ceftriaxone (MIC range, < or = 0.015-0.06 microg/ ml), and azithromycin (MIC range, < or = 0.015 0.06 microg/ml). Whereas tobramycin (MIC range, 8-64 microg/ml) exhibited little activity, spectinomycin (MIC range, 0.25-2 microg/ml) showed in vitro antimicrobial activity against Borrelia burgdorferi. The MICs of penicillin G for Borrelia afzelii isolates were ten times higher than those for Borrelia burgdorferi, Borrelia valaisiana, and Borrelia bissettii isolates (P<0.05) and 100 times higher than those for isolates belonging to the genospecies Borrelia garinii (P < 0.05). Further significant differences with respect to the MIC values of the other antimicrobial agents tested were not noted. The colorimetric microdilution method offered the advantages of reliability, reproducibility, and convenience and could handle large numbers of isolates and antibiotics. PMID- 10706177 TI - Comparison of a commercial disk test with vancomycin and colimycin susceptibility testing for identification of bacteria with abnormal gram staining reactions. AB - In an effort to identify bacteria that fail to give the expected Gram reaction, thus leading to misidentification, two nonstaining tests for Gram reaction, vancomycin and colimycin susceptibility testing and the Gram-Sure test (Remel, USA), were employed on 145 strains from 42 gram-negative and gram-positive genera with contradictory Gram stain results. The Gram-Sure test is a commercially available disk that detects the presence of L-alanine-aminopeptidase, an enzyme usually found only in the cell wall of gram-negative bacteria. In this test, aminopeptidase activity is detected using a substrate that can be hydrolyzed to produce a fluorescent compound under long-wave UV light. The commercial disk test and vancomycin plus colimycin susceptibility testing appeared to perform equally well except in the identification of Erysipelothrix and Lactobacillus, for which the commercial disk test was better, and Moraxella, for which vancomycin and colimycin susceptibility testing was more helpful. An advantage of the commercial disk test is that it can be performed in 10 min, whereas vancomycin and colimycin susceptibility testing requires at least 18 h. The commercial disk test is also less expensive than vancomycin and colimycin susceptibility testing. However, since the same results can be obtained with the 5 microg and 30 microg vancomycin disks, it is possible to use only one vancomycin disk, with the cost then being equivalent to that of the commercial disk test. The major inconvenience of the commercial disk test is the requirement of a UV ray. However, this test could be a useful tool for the identification of unusual organisms. PMID- 10706178 TI - Virulence factors of Enterococcus faecalis and Enterococcus faecium blood culture isolates. AB - Known and potential virulence factors of enterococcal blood culture isolates were studied using 89 Enterococcus faecalis and 24 Enterococcus faecium isolates. The prevalence of the respective factors was (Enterococcus faecalis vs. Enterococcus faecium): hemolysin 16% vs. 0%, gelatinase 55% vs. 0%, aggregation substance 63% vs. 13%, lipase 35% vs. 4%, hemagglutinin 97% vs. 0%. Deoxyribonuclease was not detected in any isolate. The study showed that hemagglutinin and lipase may represent additional virulence factors of Enterococcus faecalis but not Enterococcus faecium. The significance of these factors in the pathogenesis of enterococcal infection needs to be elucidated in further studies. PMID- 10706179 TI - Cluster of cases of Mycobacterium chelonae bacteraemia. AB - Mycobacterium chelonae was isolated from the blood of four immunosuppressed patients over a period of 10 weeks. Three patients had intravascular catheters in situ and the other had a biliary stent. All presented with recurrent fever despite treatment with broad-spectrum antibiotics. Blood cultures using standard bacteriological medium yielded a gram-positive bacillus from each patient. Ziehl Neelsen staining of these cultures demonstrated a branching acid-fast bacillus that was subsequently identified as Mycobacterium chelonae in each case. The isolates were sensitive to clarithromycin and, although success of treatment with clarithromycin monotherapy has been variable, this antibiotic combined with removal of the intravascular catheters was used to treat those three patients. The treatment was successful with no recurrence of symptoms after 12 months of follow-up. The patient with the biliary stent died soon after Mycobacterium chelonae was isolated. Pyrolysis mass spectrometry analysis indicated the isolates were of two distinct strains. Radiological insertion of the Hickman lines and biliary stent was implicated epidemiologically as the source of infection. PMID- 10706180 TI - Limits of detection of Mycobacterium tuberculosis in spiked cerebrospinal fluid using the polymerase chain reaction in tuberculous meningitis. AB - The limit of detection of Mycobacterium tuberculosis in spiked cerebrospinal fluid (CSF) using polymerase chain reaction (PCR) was compared to that of a radiometric liquid culture. Serial dilutions of clinical isolates of Mycobacterium tuberculosis were prepared in CSF (n=3) or broth (n=11) with estimated concentrations of 0-550 cfu/ml. Each dilution was examined concurrently by PCR and radiometric culture. PCR and radiometric culture detected Mycobacterium tuberculosis DNA in all dilutions with an estimated 2 cfu/ml in the CSF. At lower concentrations (estimated <2 cfu/ml), PCR and radiometric culture were positive in three of five (60%) and five of five (100%) CSF samples, respectively. In comparison to PCR in broth dilutions, no evidence of inhibition or interference was noted. These results imply that PCR can provide a rapid and reliable diagnosis of tuberculous meningitis, although there is a potential for false-negative results to occur in samples containing very few organisms ( < 2 cfu/ml). PMID- 10706181 TI - Localized Mycobacterium genavense soft tissue infection in an immunodeficient HIV negative patient. AB - An HIV-negative woman with chronic lymphopenia related to past sarcoidosis situated in the bone marrow presented with an inflammatory lesion in the iliac region due to a localized Mycobacterium genavense soft tissue infection. The lesion resolved after 12 months of antibiotic therapy with clarithromycin, ethambutol and ciprofloxacin. The patient had no recurrence of the subcutaneous abscess during a follow-up period of 14 months after the end of the treatment. PMID- 10706182 TI - Clinical significance of Scedosporium apiospermum in patients with cystic fibrosis. AB - The incidence of airway colonization by Scedosporium apiospermum and of related sensitization was investigated prospectively in 128 patients with cystic fibrosis over a 5-year period, and results were compared with clinical data. Scedosporium apio-spermum, recovered from sputum samples in 11 of 128 (8.6%) patients, was the most frequent filamentous fungus after Aspergillus fumigatus. Counterimmuno electrophoresis, used to detect scedosporiosis serologically, was positive in 27 of 128 (21.1%) patients. The discrepancy between the mycological and serological results may be related to immune cross-reactions between Scedosporium apiospermum and Aspergillus fumigatus. However, symptoms of allergic bronchopulmonary disease were observed in two patients chronically colonized by Scedosporium apiospermum. The results clearly demonstrate that the frequency of this fungus is largely underestimated and that it may trigger an inflammatory response, thus suggesting a pathogenic role in patients with cystic fibrosis. PMID- 10706183 TI - Severe cytomegalovirus-triggered autoimmune hemolytic anemia complicating vertically acquired HIV infection. AB - The association of acute cytomegalovirus infection with severe autoimmune hemolysis has not yet been reported in patients with HIV infection. The case is described of a 9-month-old infant with congenital HIV-1 infection who presented with severe autoimmune hemolysis and a high cytomegalovirus viral plasma load. Alternative causes of the hemolysis, such as drugs or other infections, were ruled out. After birth and after successful therapy of hemolysis, cytomegalovirus was not detected in the plasma, strongly suggesting a causal relationship between the hemolysis and cytomegalovirus infection. Severe autoimmune hemolysis should thus be considered as a cytomegalovirus-associated complication in HIV infection. PMID- 10706184 TI - Candida tropicalis vertebral osteomyelitis complicating epidural catheterisation with disease paralleled by elevated D-arabinitol/L-arabinitol ratios. AB - Deep-seated Candida infections are challenging to diagnose by noninvasive means, and new modalities are needed to improve the yield of such investigations. Reported here is a case of Candida tropicalis vertebral osteomyelitis complicating epidural catheterisation in a diabetic patient with complicated abdominal sepsis. The diagnosis was supported by detection of increased D arabinitol/L-arabinitol ratios in urine samples, and failure of medical management was indicated by elevated D-arabinitol/L-arabinitol ratios, which later decreased to baseline with successful surgical debridement and prolonged antifungal therapy. PMID- 10706185 TI - Epidemiology of quinolone resistance of Klebsiella pneumoniae and Klebsiella oxytoca in Europe. AB - The epidemiology of quinolone resistance and the concomitant resistance to other antibiotic classes was investigated in 445 Klebsiella pneumoniae and 238 Klebsiella oxytoca isolates. Decreased susceptibility to ciprofloxacin was found in 7.2% and 3.4% of these two species, respectively. Ciprofloxacin resistance was significantly linked to ceftazidime resistance, the hallmark of extended-spectrum beta-lactamase production, as well as to resistance to all antibiotic classes tested. Using automated ribotyping, seven intrahospital- and interhospital transmitted clones of ciprofloxacin-resistant isolates were found. The newer fluoroquinolones sitafloxacin and clinafloxacin may become increasingly valuable, since they proved to be active also against ciprofloxacin-resistant isolates. PMID- 10706186 TI - Isolation of mycobacteria from clinical samples using the centrifugation-shell vial technique. PMID- 10706187 TI - Chronic factitious illness presenting as Munchausen's gonarthritis. PMID- 10706188 TI - Mycobacterium kansasii abscess of the chest wall after drainage of bacterial empyema. PMID- 10706189 TI - Characterization of rapidly growing mycobacteria using a commercial identification system. PMID- 10706190 TI - Cultural differences: implications on drug therapy and global drug development. AB - INTRODUCTORY REMARKS: Discussing the "inter-ethnicity" of kinetics and actions of drugs is fraught with terminology problems. It is, however, generally accepted that "ethnicity" covers the influences of factors genetically and culturally transmitted. The study of inter-ethnic variability in drug response addresses the problem of distinguishing variability factors that are common to one particular group of individuals from those which are not specifically shared. DIFFERENCES IN DAILY DOSES BETWEEN DIFFERENT GEOGRAPHIC REGIONS: It is well known that for a number of drugs, the daily dose prescribed in Japan is lower than in the US and Europe. Presently, independent surveys strongly indicate that for a majority of drugs dose differences are not the result of pharmacokinetic differences. In addition, they indicate that inter-ethnic differences do not seem to be larger than intra-ethnic variations. The differences observed for daily doses must thus be found elsewhere than in pharmacogenetic traits. DIFFERENCES IN DIAGNOSES: The most important impediments encountered in the evaluation of minority patients include differences with respect to language, communication style, cultural belief. The same problems arise if studies performed in different geographic areas are compared, socio-economic aspects play then an even greater role. Language problems arise differently if minorities are evaluated and compared to a majority of patients living in the same country or if clinical studies are performed in different regions. Communication styles also differ markedly between cultures. As an example, certain gestures may be considered as disrespectful or insulting by some ethnic groups and constitute normal behavior in others. RATING SCALES: Ethnicity clearly plays a role on the cross-cultural use of rating scales. Sophisticated rating scales established and validated in Western culture must undergo culturally sensitive revision and rigorous evaluation before their use in non-Western culture. EFFICACY-SAFETY ASSESSMENT: As an example, the assessment of risk and benefit is different in Japan, Europe and the United States. In Japan, safety is given a greater weighting relative to efficacy than in the two other regions. PLACEBO/NOCEBO EFFECTS: Placebo and nocebo effects are difficult to study, even in the absence of any cultural difference. They are even more so if ethnicity is concerned. PATIENT COMPLIANCE: Clinicians treating cross cultural patients must carefully explore the beliefs held by their patient regarding illness causality and treatment expectations. CONCLUDING REMARKS: There are many unanswered questions in the field of inter-ethnic variability in drug response. The present overview will not pretend to have given specific answers, but it is hoped that it will point to some areas where more research is needed, in particular in the area of methodologies to take inter-ethnicity into account during drug development. PMID- 10706191 TI - Predictability of the effects of race or ethnicity on pharmacokinetics of drugs. AB - The International Conference on Harmonization has put forth a tripartite guideline addressing mechanisms by which regulatory agencies might be able to accept foreign clinical data. A major issue is the effect of ethnicity on the drug's pharmacokinetics, pharmacodynamics and/or safety. The purpose of this review was to determine whether the effects of ethnicity on pharmacokinetics are predictable. We also evaluated whether the premise that only pharmacokinetic processes which are biologically or biochemically mediated are likely to exhibit ethnic differences was supported by the literature. Bioavailability is determined by absorption, gut metabolism/transport and hepatic first pass metabolism. Absorption is usually a passive process and, as would be expected, no examples of ethnic differences in passive absorption were found in the literature. Direct evaluation of ethnic differences in gut metabolism/transport or hepatic first pass metabolism is largely lacking, although some studies suggest such differences exist. These differences would also be expected based on known ethnic differences in hepatic clearance of drugs. Ethnic differences in plasma protein binding to the two major drug-binding proteins, alpha1-acid glycoprotein (AGP) and albumin, have been studied in several populations. Based on these studies, ethnic differences in plasma protein-binding for drugs which bind exclusively to albumin (e.g. acids) are uncommon. Conversely, ethnic differences in plasma protein-binding of drugs to AGP appear to be very common, with the studies consistently showing Caucasians have higher binding (lower plasma free fractions) than other ethnic groups. This difference appears, in all cases, to be explained by racial differences in plasma AGP concentration. Hepatic metabolism is the most common pharmacokinetic parameter for which there are ethnic differences. Differences have been documented in both oxidative and conjugative metabolism. Ethnic differences in hepatic metabolism are unpredictable by race (e.g. one racial group is not consistently higher or lower than another group) and specific enzyme (e.g. studies with different CYP3A4 substrates have yielded different results). Ethnic differences in renal tubular secretion have been documented, but also appear to be unpredictable, while differences in the passive processes of renal elimination, filtration and reabsorption, have not been observed. The literature supports the premise that pharmacokinetic processes which are active (e.g. involve a protein) are the ones with the potential for differences between ethnic groups while passive pharmacokinetic processes do not exhibit such differences. Based on the available literature, the drugs most likely to exhibit ethnic differences in their pharmacokinetics are those that undergo significant gut metabolism/transport and/or hepatic first pass metabolism; are highly bound to plasma proteins (especially AGP); or have hepatic metabolism as a major route of elimination. PMID- 10706192 TI - Interethnic differences of drug-metabolizing enzymes. AB - Polymorphisms exhibited by drug-metabolizing enzymes are well known and have been investigated for many years. Recently, the exploding field of pharmacogenetics has focused not only on the characterization of enzymes responsible for drug biotransformation but also, on describing the sources of variability in enzyme activity. While initial observations and studies focused on populations of Caucasian origin, reports for other populations followed. The incidence of a poor or slow metabolizer phenotype for a given enzyme caused by allelic variants may vary significantly between populations. The question arises as to whether a prediction of the phenotype (i.e. distribution and/or enzyme activity) can be accurately ascertained from genotype information gathered in a related population. This is exemplified by NAD(P):quinone oxidoreductase (NQO1) investigated in Canadian Native Indian (CNI), Inuit and Chinese populations and the cytochromes P4502C19 and 2D6. While the two North American Native populations are genetically distinct, they are both descendants from northern Asia. Consequently, one might suspect that on a pharmacogenetic basis, CNI and Inuit would be more comparable to Chinese as opposed to Caucasian populations. This is certainly not the case as demonstrated for all three enzymes. Also, for a reliable phenotype prediction, one needs to pay attention to ethnic "mixing" which occurs between certain populations. Ethnic diversity constitutes both a challenge and an opportunity to prudently apply pharmacogenetics so that variability in both drug disposition and effect may be better understood. PMID- 10706193 TI - P-glycoprotein: a defense mechanism limiting oral bioavailability and CNS accumulation of drugs. AB - Transport by ATP-dependent efflux pumps such as P-glycoprotein is an increasingly recognized determinant of drug disposition. P-glycoprotein does not only contribute to multidrug resistance (MDR) in tumor cells, it is also expressed in normal tissues with excretory function such as liver, kidney and intestine. Apical expression of P-glycoprotein in such tissues results in reduced drug absorption from the gastrointestinal tract and enhanced drug elimination into bile and urine. Moreover, expression of P-glycoprotein in the endothelial cells of the blood-brain barrier prevents entry of certain drugs into the central nervous system. Human P-glycoprotein has been shown to transport a wide range of structurally unrelated drugs such as digoxin, quinidine, cyclosporine and HIV-1 protease inhibitors. Drug administration to P-glycoprotein knock-out and control mice provided data on the importance of P-glycoprotein for absorption after oral administration and penetration through the blood-brain barrier. Moreover, P glycoprotein knock-out mice were used to identify inhibition of P-glycoprotein mediated transport as a mechanism for drug interactions such as the digoxin quinidine interaction. Studies in humans indicate a particular importance of intestinal P-glycoprotein for bioavailability of the immunosuppressant cyclosporine. Moreover, induction of intestinal P-glycoprotein by rifampin has now been identified as the major underlying mechanism of reduced digoxin plasma concentrations during concomitant rifampin therapy. In summary, P-glycoprotein functions as a defense mechanism, which determines bioavailability and CNS concentrations of drugs. Modification of P-glycoprotein function is an important underlying mechanism of drug interactions in humans. However, disposition of a drug and its metabolites frequently is not only determined by P-glycoprotein, but also by drug-metabolizing enzymes and possibly by drug transporters other than P glycoprotein [e.g. members of the MRP family (MRP = multidrug resistance associated proteins)]. PMID- 10706194 TI - Effect of race on hypertension and antihypertensive therapy. AB - The presence of hypertension in individual patients confers significant risk in terms of coronary artery disease, myocardial infarction, stroke and congestive heart failure. However, it is also a modifiable risk factor, as risk may be decreased through either lifestyle changes or pharmacotherapy to reduce the elevated blood pressure. Over the past 3 decades, there has been strenuous debate among clinical scientists regarding the role played by racial background in both the pathogenesis and response to pharmacotherapy. A number of studies, such as the third National Health and Nutrition Examination Survey (NHANES III) have demonstrated a higher prevalence of hypertension in black populations. The Hispanic Health and Nutrition Examination Survey (HHANES) suggested that the prevalence of hypertension in Hispanics of Caribbean descent was similar to that of African Americans, while Mexican Americans had lower rates of the disease. It appears that the pathophysiological consequences of elevated blood pressure may also be more severe in black patients. Thus, these patients will have a worse prognosis than their white counterparts at any given blood pressure level. The incidence of end-stage renal disease has been reported to be as much as 17 times more common in African American patients. A number of individual factors have been postulated for these differences including increased sodium intake, differences in sodium handling, decreased potassium intake, decreased calcium intake, elevated fasting insulin levels, lower levels of plasma renin activity and urinary kallikrein excretion. These differences in prevalence and pathophysiology have resulted in recommendations for differential therapeutic approaches in the treatment of hypertension. A major trial conducted in the Veteran Affairs Medical Centers in the USA noted that African Americans are generally more responsive to diuretics and calcium channel blockers than to ACE inhibitors or beta-blockers. However, it has been reported that this resistance may be overcome by increasing the dose of these agents. It has been postulated that these differences may be related to lower plasma renin activity noted in the black population, since diuretics and calcium channel blockers appear to be better suited to this population. These differential therapeutic recommendations will be reviewed in light of our current knowledge of the disease. PMID- 10706195 TI - Necessity and requirements of bridging studies and their present status in Japan. AB - The mainstays of the harmonized final document about ethnic factors in the acceptability of foreign clinical data include a complete clinical data package and a bridging study (for efficacy and/or for safety). A clinical data package that meets all of the regional regulatory requirements is defined as a complete clinical data package for submission and potential approval, irrespective of its geographic origin. The acceptability of the foreign clinical data component of the complete data package depends upon whether it can be extrapolated or employed as a bridge to the population of the new region. Ethnic factors can be defined as the intrinsic characteristics of recipients of a medicine, and extrinsic characteristics associated with the environment and culture in which the subjects reside. Based on retrospective studies, inter-ethnic differences in ADME seem to be not greater than intra-ethnic variations for most medicines, and extrinsic factors appear to be more important than intrinsic factors for the assessment of efficacy and safety of the drug across ethnicity. Medical practice may represent one of the biggest differences and may perhaps prove to be very difficult to provide a harmonization for these extrinsic factors. The bridging study concept has primarily been brought up to overcome the difficulties inherent to extrinsic factors caused by different ethnicity. In Japan, the clinical trials advice division of the organization for pharmaceutical safety and research (OPSR) has been dealing with consultations about bridging studies since February 1998. The contents of consultations can be classified into 5 types. The most common type involves the desire by industries to bridge or extrapolate the results of foreign, phase III, clinical studies by conducting the dose-response studies domestically in the form of bridging studies. Until we have more information and have collected experiences in the variations caused by the regional differences as a result of extrinsic as well as intrinsic ethnic factors, we hope and believe that this document will serve to help provide great advancements in the acceptance of foreign clinical data. PMID- 10706196 TI - East-West development: understanding the usability and acceptance of foreign data in Japan. AB - The introduction in Japan of New GCP regulations, as well as the internationalization of clinical trials under the ICH process (International Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use), has meant that the system is now faced with a number of serious challenges. This is shown by the dramatic decrease in registration of new drugs. Identifying the actors in the new context is essential. We shall first define precisely what is the Regulatory Authority in Japan, and which are the Regulatory Requirements to be fulfilled. We shall then describe the new process for evaluation of foreign clinical data. One of the major issues will be to determine in which cases a complete clinical data package will be sufficient to support regulatory submission, and in which cases additional clinical studies or bridging studies, and what kind of bridging studies will be required. PMID- 10706197 TI - Reconstruction of the scalp and cranium using multiple free-tissue transfers following recurrent basal cell carcinoma. AB - It is well-recognised that recurrent disease can occur following surgery for malignancy in the head and neck region. This is particularly true of basal cell carcinoma in which recurrences may occur over many years and despite the use of different treatment modalities. Reconstruction of large defects may become increasingly difficult and can be optimally managed by free tissue transfer. The authors report a case of basal cell carcinoma that has required treatment for over 20 years, unique in that on five different occasions, free flaps have been used for reconstruction. PMID- 10706198 TI - Incidence of neurapraxia in digital nerve injuries. AB - Hand surgeons presented with a hand or finger laceration and an abnormal static two-point discrimination (> or =10 mm) must determine which patients require surgical exploration. There appears to be a paucity of data in the literature defining the incidence and nature of neurapraxia in this setting. A study was conducted in a busy urban practice to better understand this problem. To determine the incidence of intact nerves (i.e., neurapraxia) in digital nerve injury patients, the authors reviewed experience with 152 patients who presented with isolated digital nerve injuries over a 33-month period. Preoperative return of sensation and negative exploration cases were combined and referred to as the trauma-induced neurapraxia (TIN) group. There were 18 non-repair cases among the 152 isolated digital nerve injuries, giving a 12 percent incidence of TIN. Sensory recovery among the TIN patients ranged from 12 days to 6 months. In this study, the authors defined a group of patients who did not require surgical repair for isolated digital nerve injury as TIN cases. The TIN group represented 12 percent of the digital nerve trauma patients and required long follow-up for sensory recovery. This information is an important part of patient education and informed consent. PMID- 10706199 TI - Monitoring of free TRAM flaps with microdialysis. AB - The aim of this investigation was to follow the metabolism of free TRAM flaps using microdialysis. Microdialysis is a new sampling technique that provide opportunities to follow the biochemistry in specific organs or tissues. A double lumen microdialysis catheter or probe, with a dialysis membrane at the end, is introduced into the specific tissue. Perfusion fluid is slowly pumped through the catheter and equilibrates across the membrane with surrounding extracellular concentrations of low molecular weight substances. The dialysate is collected in microvials and analyzed by an instrument using very small volumes. Glucose, glycerol, and lactate concentrations were measured in the flaps and compared with those in a reference catheter that was placed subcutaneously in the femur. The investigation continued 72 hr postoperatively. The study group consisted of 14 women who underwent reconstruction with a free TRAM flap, and one woman with a double TRAM flap. During flap ischemia, the concentration of glucose was reduced, while the lactate and glycerol levels increased. The differences between the flaps and controls were statistically highly significant. After reperfusion of the flaps, the concentrations of glucose, lactate, and glycerol approached normal. One flap failed because of an arterial anastomosis thrombosis. This was clearly demonstrated by the samples from the microdialysis: the concentration of glucose fell to an unmeasurable level; the concentration of lactate increased for a period before it stopped due to lack of glucose; and the concentration of glycerol increased to a very high level, probably because ischemia caused damage to the cell membranes of which glycerol is an important part. The authors concluded that microdialysis can detect ischemia in free flaps at an early stage, making early surgical intervention possible. PMID- 10706200 TI - Free fibula donor-site morbidity: the Mayo experience with 100 consecutive harvests. AB - A retrospective analysis of 100 consecutive patients undergoing free fibula harvest at the Mayo Clinic is presented. Every patient was analyzed by reviewing postoperative physical examination data. All patients were evaluated and followed in the early postoperative course by the physical medicine and rehabilitation services. Patients were followed from 3 to 60 months, with an average follow up of 17.42 months. In the patient group, 72 flaps were osseous and 28 osteocutaneous. Thirty-six complications at the donor site were observed in 30 patients. An additional 19 patients required prolonged pharmacologic pain control beyond the first 6 postoperative weeks, with no donor-site complications clinically detectable. Hammertoe was observed in six patients and wound dehiscence in seven patients. Tendon exposure was observed in five patients; partial split-thickness skin graft loss was observed in eight. Numbness of the foot was reported in 10 patients. Fifteen patient had limited maximum ambulatory distance to less than 1000 m. An additional six patients reported difficulty walking stairs. Attention to details and meticulous wound care are required to further reduce wound-healing complications. Immediate postoperative involvement of the physical medicine and rehabilitation services was beneficial in early patient mobilization and achievement of preoperative ambulation levels. After a short rehabilitation period, the majority of patients were able to engage in all daily activities. PMID- 10706201 TI - In vivo visualization of the neuromuscular junction before and after autologous nerve transfer in the rat. AB - In vivo visualization of the neuromuscular junction with epifluorescence imaging techniques has become a successful method of observing the ongoing process of re occupation by regenerating motor axons of former post-synaptic sites after nerve injury. By using a light-integrating video camera for digital documentation, all parts of the neuromuscular junction can be visualized, as detailed as when documented with high-speed film, but with a minimum light intensity to prevent damage of neural or muscular structures. Results from comparisons of pre- and post-synaptic staining indicate a non-reoccupation rate up to 37 percent at a 55 day interval after nerve transfer, and up to 34 percent at a 66-day postoperative interval. Morphologic findings suggest that these high non-reoccupation rates are caused jointly by intramuscular missprouting, an insufficient intramuscular guidance apparatus, and intramuscular microneuroma formation at the insufficient neuromuscular junction. PMID- 10706202 TI - Experimental study of vascularized bone grafts: hypertrophy of the grafted bone. AB - The mechanism underlying hypertrophy of experimentally vascularized bone grafts was studied in 15-week-old rats. The segmental ulna was grafted to the tibial defect with an external fixator. In experiment 1, 24 rats were classified into four groups to evaluate conventional (non-vascularized), cuff (periosteum encased, non-vascularized), and vascularized bone grafts, and vascularized segmental grafts with fracture. In experiment 2, 12 rats were classified into two groups according to the presence of mechanical loading. This involved vascularized bone grafts with external fixators, and vascularized bone grafts with external fixators removed after bone union. The bone dynamics of the grafts were investigated by several methods, including roentgenographic analysis, histologic studies, and fluorochrome labeling. In experiment 1, a slight bone formation was recognized in the conventional bone graft, while irregular bone formation with creeping substitution was observed in the cuff graft. The vascularized bone graft showed significant hypertrophy; hypertrophy of the vascularized bone with fracture was greater than that without fracture. In experiment 2, markedly circumferential bone formation was observed after removal of the external fixator, while slight new bone formation was observed during the late postoperative period in bone with an external fixator. These results suggest that hypertrophy can be promoted by artificial fracture of the grafted bone, and that mechanical loading is an important factor for remodeling of grafted bone. PMID- 10706203 TI - Prediction of border necrosis in skin flaps of pigs with microdialysis. AB - Metabolic changes were studied in newly-raised pedicled skin flaps of pigs. These flaps were constructed so that their distal parts were predestined to necrotize. The objective was to find new ways of making early postoperative prognostications about future flap viability. For that purpose, the fluorescein penetration technique was compared with microdialysis monitoring of interstitial tissue concentrations of glucose, lactate, and glycerol. These parameters were measured 6 to 24 hr postoperatively, and collected at five different sites, ranging from base to end of the flap. The fluorescein penetration border appeared closer to the flap base than a subsequent necrotic border, thereby confirming that this technique--when applied early in the postoperative period--underestimates flap viability. The authors also observed that glucose concentration in the flap declined at an early stage close to the border of fluorescein penetration. No further change was seen in more distal parts of the flap. Consequently, glucose concentration underestimates the viable flap area just as fluorescein does. In contrast, both glycerol and lactate concentrations began to increase closer to the necrotic border. Both increased significantly on passing the border between viable tissue and tissue which later on would become necrotic, and reached levels in the necrotic portion which were never seen in the viable parts. These results were obtained within the first postoperative hours. They suggest that microdialysis monitoring of lactate and glycerol concentrations in skin flaps of pigs can be used to estimate at an early stage where the necrotic border will appear later on. If these results hold true in humans, they may have important clinical applications. PMID- 10706204 TI - Direct muscle neurotization in rat soleus muscle. AB - The authors carried out an experimental study using rat soleus muscles to evaluate reinnervation of the denervated muscles by direct muscle neurotization. They also attempted to determine whether there is any difference in the regeneration process between original (tibial) and foreign (peroneal) nerve neurotization. For functional evaluation, they did an electrophysiologic study by analyzing pattern, latency, amplitude, and duration of compound motor action potential. For histochemical studies, both hematoxyline-eosin stain and nicotineamide adenin dehydrogenase stainings were used to identify the morphology and the change of the type of muscle fiber. Combined silver-acetylcholinesterase stain was performed to identify reinnervated motor endplate and axonal sprouting. They confirmed evidence of regeneration of the denervated muscle by direct muscle neurotization. Regenerating muscles showed type groupings of muscle fibers. The newly-formed ectopic motor endplate was connected with axonal sprouting. The giant motor endplate composed of mature axon sprouting and several new ectopic motor endplates, appeared in the neurotization group. There was no significant difference in the regeneration process between original and foreign nerve neurotization. PMID- 10706205 TI - Composite tissue allotransplantation: a comprehensive review of the literature- Part II. PMID- 10706206 TI - Attitudes and perceptions of fitness professionals regarding obesity. AB - The purpose of this study was to assess perceptions of exercise professionals regarding obesity. A three page, 25-item survey was mailed to a random sample of 500 certified Health Fitness Instructors, Exercise Test Technologists, and Exercise Specialists from a certification list provided by the American (College of Sports Medicine (ACSM). The return rate was 66%. Most exercise professionals (74%) supervised or worked directly with obese clients in an exercise/fitness setting. The majority of exercise professionals believed that normal weight is very important to a person's health, that physical activity is very important in the treatment of obesity, that they should be role models by maintaining normal weight, that they are obligated to counsel obese persons concerning the health risks of obesity, that they are very competent to prescribe exercise programs for weight loss, and that counseling obese persons on exercise for weight loss is professionally gratifying. The majority of exercise professionals also believed sedentary lifestyles, poor eating behavior, excessive calorie consumption, and psychological problems play a major role in most obesity. Exercise professionals reported that they received most of their information on weight control from textbooks, college classes, scientific journals, workshops/seminars, and past experience. PMID- 10706207 TI - Predicting cost-benefits before programs are started: looking at conjugate vaccine for invasive pneumococcal infections. AB - This analysis uses existing data to examine how an analysis to predict the net financial impact for an emerging medical program, namely a conjugate vaccine against Streptococcus pneumoniae, and to identify which key variables will have the greatest impact on the program's costs and benefits. Using data available on the prevalence and case fatality rates for invasive diseases caused by S pneumoniae, we examined the theoretical economic impact of vaccinating all newborns versus not vaccinating. Effectiveness estimates for conjugated pneumococcal vaccines and disease incidence and fatality rates were obtained from published sources. Because of scanty or inconclusive data for otitis media and pneumonia, the analysis was limited to cases of meningitis and bacteremia due to S pneumoniae. Based on these two diseases alone, immunization with conjugate pneumococcal vaccine could save an estimated 222 lives per million children vaccinated per year. Analysis of direct costs (projected immunization costs minus savings from reduced illness) show that a pneumococcal vaccine program will result in net direct costs between $0.08 and $2.42 per child. When indirect costs are included in the analysis, the vaccine is cost savings for all cases except when the two year incidence of disease and death rates are lowest and the cost of the vaccine series is $150. Further research should focus on these key issues as the vaccine is introduced into use, as expected in the next few years. PMID- 10706208 TI - Utilization of health care services among adults attending a health fair in South Los Angeles County. AB - A bilingual survey was developed to collect information regarding socio demographics, access to medical and dental care, health insurance coverage, perceived health status, and use of folk medicine providers from 70 adults presenting to a health fair in South Los Angeles County. Ninety-seven percent of respondents were foreign-born. Seventy-nine percent reported having no health insurance during the year prior to survey. Of the uninsured, 61 percent lacked a doctor visit and 76 percent lacked a dental visit during the previous year. The high cost of care was the most frequently cited barrier to seeking medical (58 percent) and dental (67 percent) care even when respondents felt it was necessary. Respondents who felt they needed medical attention but did not seek it had a lower perceived health status (7.0 +/- 2.2) than those who did (8.0 +/ 2.0). Among respondents perceiving themselves in poor health, only 17 percent were insured. Relatively few respondents (7.2 percent) reported seeing a folk healer during the past year. Our results support the argument that the medically indigent in some localities face serious financial, as well as less salient, barriers to access. These local conditions reflect inadequate enforcement by local governments in correcting the difficult problems indigent populations face in accessing medical and dental care. PMID- 10706209 TI - Common perceptions about strokes. AB - The frequency with which strokes occur and the devastating effects they can have make provision of general stroke information an essential element of public health education. This survey study sought to explore the general public's knowledge about various aspects of strokes. A questionnaire consisting of 4 open ended, free-recall questions and 27 yes/no questions about the physiological processes, risk factors, warning signs, and functional consequences of strokes was administered orally to 190 individuals at a regional shopping mall. Additional items provided information about respondents' ages, ethnicity, educational status, and personal experiences with strokes. Free recall results indicated that approximately two-thirds of the survey respondents could provide correct or partially correct explanations of stroke physiology and could name at least one stroke warning sign; over 90% of respondents could name at least one stroke risk factor and one functional consequence of stroke. Most respondents reported acquiring information about strokes through personal acquaintances, popular media, or general life experiences rather than from professionals or as part of their formal schooling. Suggestions about needed content in general educational programs came from respondents' misconceptions about physiological processes, risk factors, warning signs, and functional consequences of strokes. PMID- 10706210 TI - Knowledge of human papillomavirus infection among young adult men and women: implications for health education and research. AB - Human papillomavirus (HPV) infection of the genital tract is one of the most common sexually transmitted diseases (STDs), and a subset of genital tract HPVs are etiologically associated with cervical cancer. The prevalence of HPV infection is highest among adolescents and young adults. This study was undertaken to explore first year college students' knowledge about HPVs and to determine whether there were gender differences in this knowledge. An anonymous survey was distributed to all first year students at a private university. The results were analyzed by gender. We found that 96.2% of males and 95.4% of females had heard of genital warts, although only 4.2% of males and 11.6% of females knew that HPV caused genital warts. Although there was a greater awareness of genital warts than HPV in this population, students were uncertain about modes of transmission of both genital warts and HPVs, and unclear about the importance of HPV infection relative to other STDs. For both men and women (87% and 87.4%, respectively), health education classes were the major source of information about STDs. We conclude that health education should be reconceptualized to incorporate a better understanding of STDs, including HPV infection, by engaging adolescents and young adults in exploring the biological and social context of STDs, their public health importance, strategies for prevention, and the uncertainty of our scientific knowledge. PMID- 10706211 TI - The relationship between the geographic density of alcohol outlets and alcohol related hospital admissions in San Diego County. AB - Increasing concerns regarding the cost of medical care have led to research that has found a relationship between alcohol abuse, increased medical problems, longer hospital stays, and higher medical costs. Research has also found a positive relationship between alcohol availability and crime, car accidents, and liver cirrhosis deaths. One area of interest is how alcohol availability, as measured by the number of alcohol outlets, is related to medical care needs. The purpose of this study was to examine the relationship between the geographic density of alcohol outlets and the number of alcohol-related hospital admissions. Alcohol-related ICD-9 codes were selected based on epidemiologic research in the literature to determine alcohol-related morbidity from the California Discharge Data System, which collects information on all hospital admissions and discharges in California. In San Diego County, in 1996, 3,759 admissions were alcohol related. Alcohol-related admissions for each zip code were compared to the number of liquor licenses that were held by each zip code through a multiple regression analysis. The regression model demonstrated that the number of liquor outlets was a significant predictor of alcohol-related hospital admissions, net of other predictors. Implications are discussed, including regulation of alcohol availability, which may have a beneficial impact on alcohol morbidity. PMID- 10706212 TI - A model of hippocampally dependent navigation, using the temporal difference learning rule. AB - This paper presents a model of how hippocampal place cells might be used for spatial navigation in two watermaze tasks: the standard reference memory task and a delayed matching-to-place task. In the reference memory task, the escape platform occupies a single location and rats gradually learn relatively direct paths to the goal over the course of days, in each of which they perform a fixed number of trials. In the delayed matching-to-place task, the escape platform occupies a novel location on each day, and rats gradually acquire one-trial learning, i.e., direct paths on the second trial of each day. The model uses a local, incremental, and statistically efficient connectionist algorithm called temporal difference learning in two distinct components. The first is a reinforcement-based "actor-critic" network that is a general model of classical and instrumental conditioning. In this case, it is applied to navigation, using place cells to provide information about state. By itself, the actor-critic can learn the reference memory task, but this learning is inflexible to changes to the platform location. We argue that one-trial learning in the delayed matching to-place task demands a goal-independent representation of space. This is provided by the second component of the model: a network that uses temporal difference learning and self-motion information to acquire consistent spatial coordinates in the environment. Each component of the model is necessary at a different stage of the task; the actor-critic provides a way of transferring control to the component that performs best. The model successfully captures gradual acquisition in both tasks, and, in particular, the ultimate development of one-trial learning in the delayed matching-to-place task. Place cells report a form of stable, allocentric information that is well-suited to the various kinds of learning in the model. PMID- 10706213 TI - Hippocampal mossy fibers and swimming navigation learning in two vole species occupying different habitats. AB - We showed previously for mice that size differences of the infrapyramidal hippocampal mossy fiber projection (IIP-MF) correlate with spatial learning abilities. In order to clarify the role of the IIP-MF in a natural environment, we studied the bank vole (Clethrionomys glareolus), adapted to a wide range of different habitats, and the root vole (Microtus oeconomus), living in homogenous grassland habitats with small home ranges. Morphometry on Timm-stained horizontal brain sections of six C. glareolus and six M. oeconomus revealed that the size of the entire mossy fiber projection was 42% larger in C. glareolus than M. oeconomus. C. glareolus had also an IIP-MF projection about 230% larger than that of the root vole. A sample of captured animals was then transferred to the laboratory (C. glareolus, n = 23; M. oeconomus, n = 15) and underwent testing for swimming navigation according to a standardized protocol used to assess water maze learning in about 2,000 normal and transgenic mice. Both species learned faster than laboratory mice. Overall escape times showed no differences, but path length was significantly reduced in C. glareolus, which also showed superior performance in a variety of scores assessing spatial search patterns. On the other hand, M. oeconomus showed faster swimming speed, and strong thigmotaxis combined with circular swimming. M. oeconomus also scored at chance levels during the probe trial, about as poorly as mutant knockout mice considered to be deficient in spatial memory. These differences probably reflect differential styles of water maze learning rather than spatial memory deficits: C. glareolus appears to be superior in inhibiting behavior interfering with proper spatial search behavior, while M. oeconomus succeeds in escaping by using rapid circular swimming. We assume that size variations of the IIP-MF correspond to a mechanism stabilizing hippocampal processing during spatial learning or complex activities. This corresponds to the ecological lifestyle of the two species and is in line with previous observations on the role of the IIP-MF. PMID- 10706214 TI - Immunocytochemical mapping of Fos protein following seizures in gerbils indicates the activation of hippocampal neurons. AB - The expression of the proto-oncogene, c-fos, and its protein, Fos, has been shown to be a useful marker for elevated levels of neuronal activity generated in the brain following different stimuli, including seizures. Since previous studies indicated hippocampal involvement in seizure activity in gerbils, Fos immunocytochemistry was used to determine whether hippocampal neurons become activated following environmentally induced seizures in this animal. Gerbils with maximal seizures showed many Fos-immunolabeled neurons in the granule cell layer and hilus of the dentate gyrus, as well as in CA3 and CA1 of the hippocampus. These gerbils had significantly greater numbers of Fos-immunolabeled dentate granule cells than gerbils with less severe seizures or no seizures. The number of dentate granule cells and CA3 pyramidal cells with Fos immunolabeling increased in an exponential manner with increased seizure severity. Many Fos immunolabeled neurons were found in several regions of the neo- and paleocortex and in other limbic structures including the piriform cortex, cortical amygdaloid nucleus, and arcuate nucleus of the hypothalamus. These results indicate that hippocampal neurons are activated following seizures in a genetic model, and provide further proof that the hippocampal formation is involved in the circuitry for seizures in gerbils. PMID- 10706215 TI - Serotonergic reinnervation reverses lesion-induced decreases in PSA-NCAM labeling and proliferation of hippocampal cells in adult rats. AB - Serotonin (5-HT) is believed to play a role in structural plasticity in the adult brain, and cell adhesion molecules may be involved in such adaptive processes. The present study sought to determine the effects of 5-HT denervation and reinnervation of the hippocampal formation on the expression of glial and neuronal markers and neurogenesis in adult rats. Injections of 5,7 dihydroxytryptamine (5,7-DHT) in the dorsal and medial raphe nuclei, producing a partial loss of 5-HT neurons, induced rapid and transient increases in glial fibrillary acidic protein immunoreactivity indicative of a reactive gliosis, but no changes in the S100beta or tenascin-C normally secreted by astroglial cells. In contrast, as long as the hippocampal formation was deprived of 5-HT innervation, significant decreases were observed in the number of granule cells expressing the highly polysialylated form of the neural cell adhesion molecule (PSA-NCAM) as well as the PSA-NCAM staining of the hilus in the dentate gyrus. Similar decreases in the number of newly generated granule cells labeled with bromodeoxyuridine were also detected during this time. All these effects were reversed later, when the hippocampal formation was reinnervated by 5-HT fibers. These results indicate that 5-HT is one factor which may regulate the number of granule cells proliferating in the adult dentate gyrus and thereafter expressing PSA-NCAM immunoreactive at the level of cell bodies, dendrites, and axonal paths (mossy fibers). They emphasize the critical role played by 5-HT in the neuronal organization of the hippocampus. PMID- 10706216 TI - Long-term memory underlying hippocampus-dependent social recognition in mice. AB - The ability to learn and remember individuals is critical for the stability of social groups. Social recognition reflects the ability of mice to identify and remember conspecifics. Social recognition is assessed as a decrease in spontaneous investigation behaviors observed in a mouse reexposed to a familiar conspecific. Our results demonstrate that group-housed mice show social memory for a familiar juvenile when tested immediately, 30 min, 24 h, 3 days, and 7 days after a single 2-min-long interaction. Interestingly, chronic social isolation disrupts long-term, but not 30-min, social memory. Even a 24-h period of isolation disrupts long-term social memory, a result that may explain why previous investigators only observed short-term social memory in individually housed rodents. Although it has no obvious configural, relational, or spatial characteristics, here we show that social memory shares characteristics of other hippocampus-dependent memories. Ibotenic acid lesions of the hippocampus disrupt social recognition at 30 min, but not immediately after training. Furthermore, long-term, but not short-term social memory is dependent on protein synthesis and cyclic AMP responsive element binding protein (CREB) function. These results outline behavioral, systems, and molecular determinants of social recognition in mice, and they suggest that it is a powerful paradigm to investigate hippocampal learning and memory. PMID- 10706218 TI - Contribution of multiple sensory information to place field stability in hippocampal place cells. AB - Hippocampal place cells in rats display spatially selective firing in relation to both external and internal cues. In the present study, we assessed the effects of removing visual and/or olfactory cues on place field stability. Place cell activity was recorded as rats searched for randomly scattered food in a cylinder. During an initial recording session, the lights were on and the only available cue was a single white cue card. Following this session, three sessions were run in a row with the cue card removed. In addition, the lights were either turned off or left on and the floor was either cleaned or left unchanged, thus creating four conditions: dark/cleaning, dark/no cleaning, light/cleaning, and light/no cleaning. A fifth session was run with the cue card back on the cylinder wall and the lights turned on. The rat remained in the cylinder during all sessions without being removed at any time. In the dark/cleaning and light/cleaning conditions, most place fields were not stable (i.e., abruptly shifted position). In addition, half of the cells stopped firing in the dark/cleaning condition. In contrast, in the dark/no cleaning and light/no cleaning conditions, most place fields remained stable across sessions. These results suggest that 1) rats are not able to rely on only movement-related information to maintain a stable place representation, 2) visual input is necessary for the firing of a large number of cells, and 3) olfactory information can be used to compensate for the lack of visuospatial information. PMID- 10706217 TI - Sustained enhancement of AMPA receptor- and NMDA receptor-mediated currents induced by dopamine D1/D5 receptor activation in the hippocampus: an essential role of postsynaptic Ca2+. AB - The dopaminergic system in the limbic system, particularly the D1/D5 receptor (D1/D5r), is important for certain forms of learning and memory, such as reinforcement learning, as well as the acute development of behavioral sensitization to psychostimulants such as cocaine and methamphetamine. Here, whole-cell patch-clamp recordings of evoked excitatory postsynaptic currents (EPSCs) mediated by ionotropic glutamate receptors were made from pyramidal neurons in the CA1 area of rat hippocampal slices. Activation of the D1/D5r by a selective D1/D5r agonist (+/-)-6-chloro-PB hydrobromide (SKF 81297; 3-100 microM) concentration-dependently induced a delayed-onset, sustained enhancement of EPSCs. The D1/D5r-induced effect was blocked by a selective D1/D5r antagonist (+)SCH 23390 hydrochloride (5 microM). Furthermore, the D1/D5r-induced effect involved a sustained enhancement of both the pharmacologically isolated alpha amino-3-hydroxy-5-methyl-4-isoxazoleproprionate receptor-(AMPAr-) and N-methyl-D asparate receptor- (NMDAr-) mediated EPSCs, respectively. Such persistently enhanced AMPAr- and NMDAr-mediated EPSCs were also associated with significant increases in the normalized amplitudes of the decay times of the isolated EPSCs. Blockade of NMDAr activation failed to prevent the induction of D1/D5r-induced sustained enhancement, suggesting the independence of the D1/D5r-induced effect on NMDAr activation. A rise of postsynaptic calcium was required for the induction of D1/D5r-induced sustained enhancement, being abolished by loading cells with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N '-tetracetic acid (BAPTA; 10 mM). Studies indicated that the D1/D5r activation induced a sustained enhancement of both the AMPAr-and NMDAr-mediated EPSCs in the hippocampus. Moreover, a rise in postsynaptic Ca2+ was necessary for triggering the D1/D5r-induced effect in the hippocampus, although the NMDAr-dependent mechanisms, such as the calcium entry via the NMDA channels, were not. PMID- 10706219 TI - Sparse colocalization of somatostatin- and GABA-immunoreactivity in the entorhinal cortex of the rat. AB - We studied the regional and laminar distribution of neurons expressing immunoreactivity with antibodies against the neuropeptide somatostatin (SOM) in the entorhinal cortex of colchicine-treated rats. We further determined whether these neurons also express immunoreactivity with antibodies against the neurotransmitter gamma-aminobutyric acid (GABA). Frontally and horizontally cut brain sections were subjected to double immunofluorescence histochemistry and investigated in a two-laser confocal laser scanning fluorescence microscope. The exact position of each single- or double-labeled cell was obtained via the preparation of large-scale digital fluorescence images superimposed on a brightfield digital image obtained postscanning after decoverslipping and staining with cresyl violet. Three types of SOM-positive cells were found: big multipolar cells (10-15% of the SOM-positive cells), oval cells (15-20%), and small spherical cells (majority of SOM-positive cells). Most cells were seen in layer III. In addition, we found immunoreactive cells in the other layers, with the fewest cells in layers I and IV (lamina dissecans). Of the SOM-positive cells, 18% also expressed GABA immunoreactivity; of the GABA-positive cells, 8% were also immunoreactive for SOM. Double-labeled cells were mostly small spherical cells and, infrequently, multipolar. These data indicate that in the entorhinal cortex, a large proportion of the cells belonging to the SOM population do not express GABA. We speculate that there may be several subpopulations of SOM cells, of which the largest may consist of non-GABAergic, excitatory interneurons. PMID- 10706220 TI - Ethanol, memory, and hippocampal function: a review of recent findings. AB - For well over a century, ethanol was believed to exert its effects on cognition and behavior by producing a ubiquitous depression of central nervous system activity. A general disruption in brain function was consistent with the belief that ethanol's effects on cognition and behavior were also quite general. Substantial evidence now indicates that ethanol produces a host of selective effects on neural activity, resulting in regional differences in ethanol's effects in the brain. Consistent with such evidence, recent research suggests that ethanol's effects on cognition and behavior are not as global as previously assumed. The present paper discusses evidence that many of ethanol's effects on learning and memory stem from altered cellular activity in the hippocampus and related structures. Potential mechanisms for ethanol's disruption of hippocampal function are reviewed. Evidence suggests that ethanol disrupts activity in the hippocampus by interacting directly with hippocampal neurons and by interacting with critical hippocampal afferents. PMID- 10706221 TI - Effects of prenatal alcohol exposure on the hippocampus: spatial behavior, electrophysiology, and neuroanatomy. AB - Prenatal exposure to alcohol can result in fetal alcohol syndrome (FAS), characterized by growth retardation, facial dysmorphologies, and a host of neurobehavioral impairments. Neurobehavioral effects in FAS, and in alcohol related neurodevelopmental disorder, include poor learning and memory, attentional deficits, and motor dysfunction. Many of these behavioral deficits can be modeled in rodents. This paper reviews the literature suggesting that many fetal alcohol effects result, at least in part, from teratogenic effects of alcohol on the hippocampus. Neurobehavioral studies show that animals exposed prenatally to alcohol are impaired in many of the same spatial learning and memory tasks sensitive to hippocampal damage, including T-mazes, the Morris water maze, and the radial arm maze. Direct evidence for hippocampal involvement is provided by neuroanatomical studies of the hippocampus documenting reduced numbers of neurons, lower dendritic spine density on pyramidal neurons, and decreased morphological plasticity after environmental enrichment in rats exposed prenatally to alcohol. Electrophysiological studies also demonstrate changes in synaptic activity in in vitro hippocampal brain slices isolated from prenatal alcohol-exposed animals. Considered together, these observations demonstrate that prenatal exposure to alcohol can result in abnormal hippocampal development and function. Such studies provide a better understanding of neurological deficits associated with FAS in humans, and may also contribute to the development of strategies to ameliorate the effects of prenatal alcohol exposure on behavior. PMID- 10706222 TI - Septohippocampal pathway as a site for the memory-impairing effects of ethanol. AB - Ethanol affects behavior by interacting with synaptic sites at many levels of the nervous system. However, it targets most readily and at the lowest concentrations those sites mediating higher cognitive functions such as attention and memory. The memory-impairing effects of ethanol are thought to involve the hippocampus, a structure particularly vulnerable to the effects ethanol at low concentrations and early in the rising phase of the blood ethanol concentration curve. One of the early, low-dose effects of ethanol is an interruption of the normal physiological regulation of the hippocampus by the ascending septohippocampal pathway originating in the medial septal area (MSA). Ethanol enhances GABAergic transmission in the MSA, thereby reducing the regularity and vigor with which rhythmically bursting neurons of the MSA drive the hippocampal theta rhythm. Disruption of septohippocampal activity also has consequences on the response of the hippocampus to cortical inputs. Ethanol produces a loss of hippocampal responsivity that reduces the ability of the hippocampus to encode and retrieve relevant stimulus information necessary for accurate memory. This paper examines the behavioral and neural evidence for hippocampal vulnerability to ethanol and explores the hypothesis that these effects are due to ethanol disrupting septohippocampal modulation of the hippocampus, resulting in impairments of memory. PMID- 10706223 TI - Effects of acute and chronic ethanol exposure on spatial cognitive processing and hippocampal function in the rat. AB - Animals, including rats, have a predisposition to process and use spatial information to organize and guide behavior. The hippocampus and related structures are critically involved in this function, and, consequently, it has been proposed that one function of the hippocampus is to construct "spatial cognitive maps" of environments. Lesions to the hippocampus or its connections produce a pattern of alterations in behavior which include shifts from the use of spatial information to guide behavior to the use of cue- or taxon-based information to guide behavior. Recently it was demonstrated that ethanol interacts with a specific group of neurotransmitter systems, i.e., N-methyl-D aspartate receptors and GABA(A) receptors that exist in high proportions in the hippocampus and related structures. In this review, we seek to summarize the literature demonstrating that one effect of acute and chronic ethanol exposure is to produce behavioral alterations that are strikingly similar to those found following lesions to the hippocampal system. Furthermore, cellular and anatomical alterations resulting from similar ethanol exposure paradigms will be reviewed and offered as possible mechanisms for producing the alterations in behavior. Finally, several unanswered questions concerning the interaction between ethanol and spatial cognitive processing will be identified. PMID- 10706224 TI - Complications after ileal pouch-anal anastomosis. AB - Restorative proctocolectomy with ileal pouch-anal anastomosis (IPAA) is currently the procedure of choice for ulcerative colitis patients who require colectomy. Despite its wide acceptance, a variety of long-term complications of the procedure exist that can be severe and even lead to pouch excision. Pouchitis occurs in up to one half of patients after IPAA, but is usually well controlled with medical therapy. A small percentage of patients develop chronic persistent pouchitis, which often requires long-term medical therapy and may result in pouch failure. Fistulas and strictures can also complicate the pouch procedure. In general, patients with Crohn's disease are not usually offered IPAA, because recurrence of disease, fistulas, abscesses, and strictures may lead to a higher incidence of pouch failure. Some ulcerative colitis patients develop complications after IPAA and are subsequently diagnosed with Crohn's disease. These patients may develop refractory fistulas, strictures, and extraintestinal manifestations of inflammatory bowel disease. Neoplastic transformation of the pelvic pouch has also been reported, particularly in patients with chronic pouchitis. Thorough follow-up and endoscopic surveillance with biopsies of the ileal pouch are therefore recommended. PMID- 10706225 TI - The role of laparoscopy and strictureplasty in the management of inflammatory bowel disease. AB - The surgical management of Crohn's disease has always been a challenging issue for physicians because of concerns of the historical need for repeated surgeries over time, the physiological limitations of a shortened small bowel, and the transmural, fistulizing, and/or skip-lesion nature of the disease. Patients are fearful of the potentially disfiguring results, especially the need for a permanent ostomy. The challenge has been to develop surgical approaches that are bowel sparing and/or minimally invasive. Stricutureplasty has been used with relatively good results as a bowel-sparing procedure for patients with small bowel Crohn's disease, potentially sparing patients of a short-bowel syndrome. Laparoscopic approaches to Crohn's disease have thus far been mostly limited to ileocecal resections in selected patients, but as more expertise is developed, will hopefully be extended to other surgical procedures in patients with Crohn's disease in the future. Patient criteria, success rates, complications, and economic implications are discussed for each procedure. PMID- 10706226 TI - Medical and surgical management of severe colitis. AB - The management of severe ulcerative colitis and Crohn's colitis remains a challenge, despite significant advances in medical and surgical therapy. Optimal management of the patient with severe colitis requires close collaboration between the gastroenterologist and surgeon. All patients with severe colitis should be hospitalized and treated with intravenous corticosteroids. If significant improvement does not occur within 7 to 10 days, then intravenous cyclosporine therapy or surgery is appropriate. Newer medical therapies, including heparin, tacrolimus, and other immunomodulatory agents, show promise for the treatment of severe colitis. When surgery is necessary, a total abdominal colectomy with ileostomy is the appropriate surgical intervention in most cases. In patients presenting with fulminant colitis, toxic megacolon, or perforation, earlier surgical intervention is indicated. The evaluation of and approach to the medical and surgical management of severe colitis will be reviewed. PMID- 10706227 TI - Controversies in the construction of the ileal pouch anal anastomosis. PMID- 10706228 TI - The long-term results of resection and multiple resections in Crohn's disease. AB - Crohn's disease is a panenteric, transmural inflammatory disease of unknown origin. Although primarily managed medically, 70% to 90% of patients will require surgical intervention. Surgery for small bowel Crohn's is usually necessary for unrelenting stenotic complications of the disease. Fistula, abscess, and perforation can also necessitate surgical intervention. Most patients benefit from resection or strictureplasty with an improved quality of life and remission of disease, but recurrence is common and 33% to 82% of patients will need a second operation, and 22% to 33% will require more than two resections. Short bowel syndrome is unavoidable in a small percentage of Crohn's patients because of recurrent resection of affected small bowel and inflammatory destruction of the remaining mucosa. Although previously a lethal and unrelenting disease with death caused by malnutrition, patients with short-bowel syndrome today can lead productive lives with maintenance on total parenteral nutrition (TPN). This lifestyle, however, does not come without a price. Severe TPN-related complications, such as sepsis of indwelling central venous catheters and liver failure, do occur. Future developments will focus on more powerful and effective anti-inflammatory medication specifically targeting the immune mechanisms responsible for Crohn's disease. Successful medical management of the disease will alleviate the need for surgical resection and reduce the frequency of short bowel syndrome. Improving the efficacy of immunosuppression and the understanding of tolerance induction should increase the safety and applicability of small bowel transplant for those with short gut. Tissue engineering offers the potential to avoid immunosuppression altogether and supplement intestinal length using the patient's own tissues. PMID- 10706229 TI - Traditional healers and AIDS in Uganda. PMID- 10706230 TI - International conference on ethnomedicine and drug discovery. PMID- 10706231 TI - The effect of vitamin-mineral supplementation on juvenile delinquency among American schoolchildren: a randomized, double-blind placebo-controlled trial. AB - CONTEXT: Numerous studies conducted in juvenile correctional institutions have reported that violence and serious antisocial behavior have been cut almost in half after implementing nutrient-dense diets that are consistent with the World Health Organization's guidelines for fats, sugar, starches, and protein ratios. Two controlled trials tested whether the cause of the behavioral improvements was psychological or biological in nature by comparing the behavior of offenders who either received placebos or vitamin-mineral supplements designed to provide the micronutrient equivalent of a well-balanced diet. These randomized trials reported that institutionalized offenders, aged 13 to 17 years or 18 to 26 years, when given active tablets produced about 40% less violent and other antisocial behavior than the placebo controls. However, generalization could not be made to typical schoolchildren without a controlled trial examining violence and antisocial behavior in public schools. OBJECTIVES: To determine if schoolchildren, aged 6 to 12 years, who are given low dose vitamin-mineral tablets will produce significantly less violence and antisocial behavior in school than classmates who are given placebos. DESIGN: A stratified randomized, double-blind, placebo-controlled trial with pretest and post-test measures of antisocial behavior on school property. SETTINGS AND SUBJECTS: Two "working class," primarily Hispanic elementary schools in Phoenix, Arizona. Approximately half of the potential schoolchildren participated, i.e., 468 students aged 6 to 12 years. INTERVENTION: Daily vitamin-mineral supplementation at 50% of the U.S. recommended daily allowance (RDA) for 4 months versus placebo. The supplement was designed to raise vitamin-mineral intake up to the levels currently recommended by the National Academy of Sciences for children aged 6 to 11 years. OUTCOME MEASURE: Violent and nonviolent delinquency as measured by official school disciplinary records. RESULTS: Of the 468 students randomly assigned to active or placebo tablets, the 80 who were disciplined at least once between September 1st and May 1st served as the research sample. During intervention, the 40 children who received active tablets were disciplined, on average, 1 time each, a 47% lower mean rate of antisocial behavior than the 1.875 times each for the 40 children who received placebos (95% confidence interval, 29% to 65%, < 5 .020). The children who took active tablets produced lower rates of antisocial behavior in 8 types of recorded infractions: threats/fighting, vandalism, being disrespectful, disorderly conduct, defiance, obscenities, refusal to work or serve, endangering others, and nonspecified offenses. CONCLUSION: Poor nutritional habits in children that lead to low concentrations of water-soluble vitamins in blood, impair brain function and subsequently cause violence and other serious antisocial behavior. Correction of nutrient intake, either through a well-balanced diet or low-dose vitamin-mineral supplementation, corrects the low concentrations of vitamins in blood, improves brain function and subsequently lowers institutional violence and antisocial behavior by almost half. This paper adds to the literature by enabling previous research to be generalized from older incarcerated subjects with a history of antisocial behavior to a normal population of younger children in an educational setting. PMID- 10706232 TI - The effect of vitamin-mineral supplementation on the intelligence of American schoolchildren: a randomized, double-blind placebo-controlled trial. AB - CONTEXT: Many medical, nutrition, and education professionals have long suspected that poor diet impairs the academic performance of Western schoolchildren; academic performance often improves after improved diet. However, others have suggested that such academic gains may be due to psychologic effects rather than nutrition. To resolve this issue, two independent research teams conducted randomized trials in which children were given placebos or low-dose vitamin mineral tablets designed to raise nutrient intake to the equivalent of a well balanced diet. Both teams reported significantly greater gains in nonverbal intelligence among the supplemented groups. The findings were important because of the apparent inadequacy of diet they revealed and the magnitude of the potential for increased intelligence. However, none of the ten subsequent replications, or the two original trials, were without limitations leaving this issue in controversy. OBJECTIVES: To determine if schoolchildren who consume low dose vitamin-mineral tablets will have a significantly larger increase in nonverbal intelligence than children who consume placebos in a study that overcomes the primary criticisms directed at the previous 12 controlled trials. DESIGN: A double-blind, placebo-controlled trial using stratified randomization within each teacher's class based on preintervention nonverbal intelligence. SETTINGS AND SUBJECTS: Two "working class," primarily Hispanic, elementary schools in Phoenix, Arizona, participated in the study. Slightly more than half the teachers in each school distributed the tablets daily to 245 schoolchildren aged 6 to 12 years. INTERVENTION: Daily vitamin-mineral supplementation at 50% of the U.S. daily recommended allowance (RDA) for 3 months versus placebo. OUTCOME MEASURES: Post-test nonverbal IQ, as measured by the Wechsler Intelligence Scale for Children-Revised (WISC-R), while controlling for pretest nonverbal IQ as a covariate. FOUR MAIN RESULTS: First, a significant difference of 2.5 IQ points (95% CI: 1.85-3.15) was found between 125 children given active tablets and 120 children given placebo tablets (p = 0.038). Second, this finding is consistent with the mean 3.2 IQ point net gain found in the 12 similar but less rigorous studies. Third, a significantly higher proportion of children in the active group gained 15 or more IQ points when compared to the placebo group (p < 0.01). Fourth, although 81 matched pairs produced no difference at all in nonverbal IQ gain, the modest 2.5 IQ point net gain for the entire sample can be explained by the remaining 24 children who took active tablets, and had a 16 point higher net gain in IQ than the remaining 19 placebo controls. CONCLUSIONS: This study confirms that vitamin-mineral supplementation modestly raised the nonverbal intelligence of some groups of Western schoolchildren by 2 to 3 points but not that of most Western schoolchildren, presumably because the majority were already adequately nourished. This study also confirms that vitamin-mineral supplementation markedly raises the non-verbal intelligence of a minority of Western schoolchildren, presumably because they were too poorly nourished before supplementation for optimal brain function. Because nonverbal intelligence is closely associated with academic performance, it follows that schools with children who consume substandard diets should find it difficult to produce academic performance equal to those schools with children who consume diets that come closer to providing the nutrients suggested in the U.S. RDA. The parents of schoolchildren whose academic performance is substandard would be well advised to seek a nutritionally oriented physician for assessment of their children's nutritional status as a possible etiology. PMID- 10706233 TI - Commentary on Schoenthaler et al: vitamin and mineral supplements--is the methodology sufficient to support the conclusions? PMID- 10706234 TI - Electrostimulators for acupuncture: safety issues. AB - Three representative electrostimulators were evaluated to determine whether they meet the manufacturers' labeled nominal output parameters and how the measured parameters compare with a safety standard written for implanted peripheral nerve stimulators. The pulsed outputs (pulse width, frequency, and voltage) of three devices were measured with an oscilloscope across a 500-ohm resistance, meant to simulate subdermal tissue stimulated during electroacupuncture. For each device, at least two measured parameters were not within 25% of the manufacturer's claimed values. The measured values were compared with the American National Standard ANSI/AAMI NS15 safety standard for implantable peripheral nerve stimulators. Although for two stimulators the pulse voltage at maximum intensity was above that specified by the standard, short-term clinical use may still be safe because the standard was written for long-term stimulation. Similarly, the net unbalanced DC current, which could lead to tissue damage, electrolysis, and electrolytic degradation of the acupuncture needle, was within the limits of the standard at 30 pulses per second, but not at higher frequencies. The primary conclusions are (1) that the outputs of electrostimulators must be calibrated and (2) that practitioners must be adequately trained to use these electrostimulators safely. PMID- 10706235 TI - A pilot study exploring the effect of kudzu root on the drinking habits of patients with chronic alcoholism. AB - PURPOSE: The objective of this study was to assess if kudzu root extract influences the drinking habits of veterans who entered a substance abuse treatment program. DESIGN: Prospective, randomized double-blind controlled clinical trial. SETTING: A nonacademic Veteran Affairs Medical Center. METHODS: Patients with the diagnosis of alcoholism were randomly assigned to receive either kudzu root extract 1.2 g twice daily or a matching placebo. Patients completed questionnaires that focused on craving for alcohol and sobriety status on a monthly basis. OUTCOME MEASURES: Sobriety level and craving for ethanol were assessed on a visual analogue scale from 0 to 10. RESULTS: Thirty-eight patients completed 1 month of the study; 21 randomly assigned kudzu, 17 to placebo. No statistically significance difference in craving and sobriety scores were noted after 1 month between kudzu and placebo, or at later stages with smaller numbers (15-19) of patients. CONCLUSION: In this small patient population, kudzu root appeared to be no better than placebo in reducing the craving for alcohol or promoting sobriety. PMID- 10706236 TI - Clinical trials of homeopathy and placebo: analysis of a scientific debate. AB - OBJECTIVE: Homeopathy is controversial, primarily because of the use of medicines diluted beyond the Avogadro limit. This article examines the scientific debate on whether homeopathy can have effects greater than placebo in humans. METHODS: Five rigorous English-language clinical studies published in high-impact journals that favored homeopathy were identified. Letters and other articles written in response to these articles were then retrieved and analyzed. RESULTS: Much of the content of responses to positive homeopathic research was rhetorical in nature and antagonistic in tone: homeopathic researchers were accused of bias and of being in the pocket of homeopathic manufacturers; arguments were raised that are demonstrably inaccurate or irrelevant to the interpretation of research; words such as "magic" or "potion" were used to paint homeopathic research as inherently unscientific. Other commentators argued that based on theoretical grounds homeopathy could not possibly be effective, any positive trial result must be due to bias. Surprisingly few responses raised substantive methodologic or statistical issues. Many of these are clearly inaccurate or irrelevant. The possibility of publication bias-selective publication of positive results-was raised by some commentators and remains an important criticism of the apparently positive nature of the clinical trial evidence. The most persuasive argument against accepting differences between homeopathy and placebo on the basis of current evidence is that homeopathy is scientifically implausible and so requires more evidence than normal. CONCLUSIONS: Investigators undertaking clinical research in homeopathy need to be responsive both to the dangers of publication bias and to the requirement for stronger than usual levels of evidence. PMID- 10706237 TI - Commentary on Vickers: humean, all too humean--a circular tale of molecules versus miracles. PMID- 10706238 TI - The evaluation of wound-healing potential of Hypericum hookerianum leaf and stem extracts. AB - OBJECTIVES: Hypericum hookerianum Wight and Arnott of the family Hypericaceae is a well-known plant among the 20 different species of Hypericum found in India. Because of its use as a wound-healing agent in traditional practices and literature references, the present study was undertaken to evaluate the wound healing potential of this plant. DESIGN: Methanol extracts of the leaves (HHLM) and stems (HHSM) of H. hookerianum were studied for their wound-healing properties in the form of ointment, using two types of wound models in 36 rats. Ointments of the dried extract of the leaves and stems of this plant were applied in two different concentrations (5% w/w and 10% w/w ointment of extracts in simple ointment base) in both the wound models used in the present study. The effects were studied on incision (skin-breaking strength) and excision (percent wound contraction and epithelialization time) wound models. SUBJECT: The methanol extract of both leaves and stem of H. hookerianum; 72 white albino rats of either gender containing six groups for each experimental model with six animals in each group. RESULTS AND CONCLUSION: The extract ointments at both concentrations performed significant in both the wound models. The leaf extract in both concentrations showed greater activity than the stem. Ointments of both extracts of H. hookerianum showed significant effects on wound contraction, wound closure time, tensile strength, regeneration of tissues at the wound site, among other effects. All these effects were comparable to those of a standard drug, nitrofurazone ointment (0.2% w/w). This investigation confirms the use of aerial parts of H. hookerianum as a potential wound-healing agent, a property known from folklore medicine. PMID- 10706239 TI - The perceived efficacy of various "future-ologies" and complementary medicine. AB - OBJECTIVES: To examine the relationship between beliefs in ways of telling the future (astrology, graphology, palmistry etc) and beliefs in complementary and alternative medicine (CAM). DESIGN: Participants completed a short questionnaire that requested that they rate the efficacy of 8 CAM therapies along with 12 other ways of predicting the future ranging from the well known and established (astrology) to the less well known (tasseography, oneiromancy). Short descriptions of each were provided. They also answered four attitude statements on science as applied to medicine. SUBJECTS: Two hundred three (130 female, 73 male) adult Britains obtained from a university subject panel served as unpaid volunteer subjects. RESULTS: CAM therapies were judged as modestly effective and most of the other "-ologies" ineffective. Further analysis confirmed two clear factors with the different methods loading on two different factors. Regressions showed females who were less concerned with scientific evaluations but more concerned with treatment believed more in the efficacy of the "future-ologies." Also females, who had heard of fewer "future-ologies" but more CAM practices were more likely to believe in the efficacy of CAM therapies. CONCLUSION: Belief in CAM is unrelated to belief in "future-ologies." Interest in the scientific evaluations of treatment is the best predictor of beliefs about efficacy. PMID- 10706240 TI - M.D. programs in the United States with complementary and alternative medicine education opportunities: an ongoing listing. PMID- 10706241 TI - Recreation, consumption of wild game, risk, and the Department of Energy sites: perceptions of people attending the Lewiston, ID, "Roundup". AB - Several federal agencies are reclaiming land through remediation and restoration, and are considering potential future land uses that are compatible with current land uses and local needs. Understanding potential recreational and wild game consumption patterns and risk perceptions are critical for determining cleanup levels and assessing potential risk associated with certain uses. In this article, recreational rates of people attending the Lewiston "Roundup" rodeo in northwestern Idaho were examined, as well as their perceptions of the safety of consuming fish and game from two Department of Energy (DOE) facilities: the Hanford Site and the Idaho National Engineering and Environmental Laboratory (INEEL). These are two of DOE's largest sites. Lewiston is closer to Hanford, but is in the same state as INEEL. Men engaged in significantly higher hunting and fishing rates than women, but there were no gender differences in camping and hiking rates. Rates of hunting and camping decreased significantly with age, while rates of hiking were lowest for 31- to 45-yr-olds. Level of education generally was not related to rates of recreation. Over 70% of the subjects ate deer, elk, and self-caught fish; 30-50% ate grouse, moose, and waterfowl; and fewer people ate other game species. Overall, subjects were less concerned about eating the fish and game from INEEL than from Hanford, and more people thought Hanford should be cleaned up completely compared to INEEL. Mean rates of fishing, hiking, and camping all exceeded the DOE's maximum recreational exposure assumption of 14 d/yr used in their future use documents. Although at present people are generally not allowed access to DOE lands for recreation, recreation is one future land use being considered for these federal facilities. Given that some people would engage in multiple activities, the potential exists for people living in the general region of Hanford and INEEL to exceed the 14-d exposure assumption. The relative gender differences in recreational rates mean that men are potentially more at risk, particularly since hunting (on both sites) and fishing (on Hanford) are attractive. PMID- 10706242 TI - Lipid peroxidation in workers exposed to hexavalent chromium. AB - The aim of this study was to investigate whether exposure to hexavalent chromium induces lipid peroxidation in human. This study involved 25 chrome-plating factory workers and a reference group of 28 control subjects. The whole-blood and urinary chromium concentrations were determined by graphite furnace atomic absorption spectrophotometry. Malondialdehyde (MDA), the product of lipid peroxidation, was determined by high-performance liquid chromatography, and the activities of protective enzymes were measured by ultraviolet-visible spectrophotometry. In the chrome-plating workers, the mean concentrations of chromium in blood and urine were 5.98 microg/L and 5.25 microg/g creatinine, respectively; the mean concentrations of MDA in blood and urine were 1.7 micromol/L and 2.24 micromol/g creatinine. The concentrations of both chromium and MDA in blood and urine were significantly higher in the chromium-exposed workers. The activities of superoxide dismutase (SOD), glutathione peroxidase (GPX), and catalase (CAT) were not markedly different between control and exposed workers. Data suggest that MDA may be used as a biomarker for occupational chromium exposure. Antioxidant enzymic activities are not a suitable marker for chromium exposure. PMID- 10706243 TI - Protein tyrosine kinase activation by polycyclic aromatic hydrocarbons in human HPB-ALL T cells. AB - It has been well established that certain polycyclic aromatic hydrocarbons (PAHs), such as 7,12-dimethylbenz[a]anthracene (DMBA), 3-methylcholanthrene (3MC), and benzo[a]pyrene (BaP), produce immunotoxicity and cancer in rodents and that these effects are also likely seen in humans. Our laboratory has found that polycyclic aromatic hydrocarbons (PAHs) produce an increase in intracellular Ca2+ in lymphocytes that appears to correlate with their immunotoxicity. Specifically, immunotoxic PAHs, such as DMBA and BaP, have been shown to produce a sustained increase in intracellular Ca2+ in lymphocytes, whereas nonimmunosuppressive PAHs, such as benzo[e]pyrene (BeP) and anthracene, do not. Our studies previously demonstrated that the rapid increase in intracellular Ca2+ produced by DMBA in HPB-ALL T cells was caused by protein tyrosine kinase (PTK) activation in human T cells, leading to tyrosine phosphorylation of phospholipase C (PLCgamma) and IP3 dependent Ca2+ mobilization. However, the specificity of PTK activation by PAHs was not established. In the present studies, we extend our observations of PTK activation by examining a number of PAHs for their effects on total and specific (Fyn and ZAP-70) PTK activity. We show that 10 microM concentrations of PAHs nonspecifically and rapidly (within 5 min) stimulate PTKs in the HPB-ALL human T cell line. BeP and anthracene were found to be nearly as effective at increasing total tyrosine kinase activity as DMBA, 3MC, and BaP, observed 5 min after exposure. We found that only immunotoxic PAHs activated the Fyn and ZAP-70 PTKs at 10 min, but total PTK activity was still increased by nonimmunotoxic PAHs, BeP, or anthracene after 10 min of exposure. These studies demonstrate that immunotoxic PAHs increase total and specific PTK activity in the human HPB-ALL T cell line. Thus the rapid increase in PTK activity produced by PAHs may not correlate with the immunotoxicity of these agents. PMID- 10706244 TI - Brainstem axolemmal protein phosphorylation in vitro in hens dosed with di-1 butyl-2,2-dichlorovinyl phosphate. AB - Neuropathy target esterase (neurotoxic esterase, NTE), a protein thought to be involved in the production of organophosphorus compound-induced delayed neurotoxicity (OPIDN), has been postulated to be a component of endogenous neuronal protein phosphorylation systems. The purpose of this work was to test this hypothesis as well as to investigate further the role of endogenous protein phosphorylation in toxic neuropathies. White Leghorn hens were dosed with the neuropathic compounds di-1-butyl-2,2-dichlorovinyl phosphate (dibutyl dichlorvos, DBDCV), tri-o-cresyl phosphate (TOCP), or acrylamide, and regions from brain were fractionated into axolemmal, synaptosomal, and microsomal preparations. Radiolabeling of NTE or endogenously phosphorylated proteins was carried out by incubation with [14C]-DFP or gamma-[32P]-ATP, respectively. Radiolabeled proteins were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS PAGE) and visualized by autoradiography. Relative amounts of phosphoproteins were quantified by densitometry of the autoradiographs. Changes in endogenous phosphorylation of a protein exhibiting the characteristics of NTE were not observed in these experiments. However, levels of a [32P]-labeled 50-kDa brainstem axolemmal protein were decreased significantly on d 15, but not on d 1, 3, 7, or 10 after dosing with 2.8 mg/kg DBDCV. Clinical signs of ataxia and histopathological findings of axonal degeneration in the spinocerebellar tracts of the brainstem were evident on d 10-15, and hens were unable to perch on a horizontal wooden rod from d 12 after dosing with DBDCV. The decrease in the 50 kDa phosphoprotein was not observed on d 15 after the production of clinically evident neuropathy with either 14 daily doses of 50 mg/kg acrylamide or with a single dose of 500 mg/kg TOCP. These results suggest that NTE is not an endogenously phosphorylated protein under the conditions of these experiments. However, an effect on endogenous phosphorylation limited to a 50-kDa axolemmal protein was selectively produced by treatment with a neuropathic dose of DBDCV that was in evidence only after clinical signs and histopathological findings of axonopathy were apparent. PMID- 10706245 TI - Assessment of the estrogenic effects of zearalenone after treatment with ozone utilizing the mouse uterine weight bioassay. AB - The ability of ozone gas (O3) to detoxify zearalenone (ZEN), a commonly occurring estrogenic mycotoxin, was assessed utilizing the mouse uterine weight bioassay. Solutions containing 12 ppm ZEN in water were ozonated for varying time periods (0, 0.5, and 5 min), then extracted with chloroform and evaporated to dryness. The residue was redissolved in acetonitrile and analyzed for ZEN. High performance liquid chromatography (HPLC) analysis of aliquots indicated a rapid degradation and decline in parent ZEN level with increasing time of ozone treatment. The acetonitrile solution containing the degraded ZEN residue was added to a known volume of corn oil and evaporated under nitrogen to eliminate the acetonitrile in the oil. Eighteen-day-old prepubertal female mice (B6C3F1 strain) were gavaged daily with the test chemicals in 50 microl of corn oil between d 18 and 23. Initial dose-response studies showed that a concentration of 60 microg ZEN/mouse/d produced uterine weights that were significantly higher than the uterine weights of control animals (2.7 times higher than that of the solvent control). Treatment groups for the ozonation study included: DES, 0.1 microg (positive control), untreated ZEN (60 microg), extraction control for ZEN (60 microg), 0.5 min ozone-treated ZEN (60 microg), 5 min ozone-treated ZEN (60 microg), solvent control (50 microl), and absolute control. Results showed the uterine weights of animals receiving the ozone-treated ZEN were not significantly affected. These findings were in agreement with HPLC analyses and suggested that ozone can prevent the estrogenic effects of this important mycotoxin in mice. Importantly, ozone treatment of contaminated whole grains may enable the practical detoxification and control of ZEN. Also, the mouse uterine weight bioassay may be useful in assessing the efficacy of other detoxification strategies for estrogenic chemicals. PMID- 10706246 TI - Hollow apologies should be avoided. PMID- 10706247 TI - Celera in talks to launch private sector human proteome project. PMID- 10706248 TI - Merck blocks 'safer' gene therapy trials. PMID- 10706250 TI - Med school to relax rules on business links? PMID- 10706251 TI - AAAS members fret over links with theological foundation PMID- 10706249 TI - Celera's shotgun approach puts Drosophila in the bag. PMID- 10706252 TI - Japan set to tighten ethics rules for genetic sampling. PMID- 10706253 TI - Germany holds up cultivation of GM maize... PMID- 10706254 TI - Open annotation offers a democratic solution to genome sequencing. PMID- 10706255 TI - Affirmative action won't solve sex discrimination. PMID- 10706256 TI - Why Roche deserves the disputed Taq patent. PMID- 10706257 TI - ECT has proved effective in treating depression... PMID- 10706258 TI - And there's no proof of lasting brain damage. PMID- 10706259 TI - Fighting the wrong battle. PMID- 10706260 TI - Avatars in space. PMID- 10706261 TI - Proteomics for the pore. PMID- 10706262 TI - Crystal structure. As weird as they come PMID- 10706263 TI - Neurobiology. Tepid tastes. PMID- 10706264 TI - Earth science. A shear pathway to the core PMID- 10706265 TI - Plant biology. A social stigma. PMID- 10706266 TI - Galaxy formation in action. PMID- 10706268 TI - Laser physics. Attosecond pulses at last PMID- 10706267 TI - Biodiversity. Extinction by numbers. PMID- 10706269 TI - Neurofibromin progress on the fly. PMID- 10706270 TI - Paul Sigler (1934-2000). PMID- 10706271 TI - The sound of many hands clapping. PMID- 10706272 TI - Leptin and diabetes in lipoatrophic mice. PMID- 10706273 TI - Exploitation of gut bacteria in the locust. PMID- 10706274 TI - Recovery of breeding success in wild birds. PMID- 10706275 TI - Biodiversity hotspots for conservation priorities. AB - Conservationists are far from able to assist all species under threat, if only for lack of funding. This places a premium on priorities: how can we support the most species at the least cost? One way is to identify 'biodiversity hotspots' where exceptional concentrations of endemic species are undergoing exceptional loss of habitat. As many as 44% of all species of vascular plants and 35% of all species in four vertebrate groups are confined to 25 hotspots comprising only 1.4% of the land surface of the Earth. This opens the way for a 'silver bullet' strategy on the part of conservation planners, focusing on these hotspots in proportion to their share of the world's species at risk. PMID- 10706276 TI - Structural basis for recognition and repair of the endogenous mutagen 8 oxoguanine in DNA. AB - Spontaneous oxidation of guanine residues in DNA generates 8-oxoguanine (oxoG). By mispairing with adenine during replication, oxoG gives rise to a G x C --> T x A transversion, a frequent somatic mutation in human cancers. The dedicated repair pathway for oxoG centres on 8-oxoguanine DNA glycosylase (hOGG1), an enzyme that recognizes oxoG x C base pairs, catalysing expulsion of the oxoG and cleavage of the DNA backbone. Here we report the X-ray structure of the catalytic core of hOGG1 bound to oxoG x C-containing DNA at 2.1 A resolution. The structure reveals the mechanistic basis for the recognition and catalytic excision of DNA damage by hOGG1 and by other members of the enzyme superfamily to which it belongs. The structure also provides a rationale for the biochemical effects of inactivating mutations and polymorphisms in hOGG1. One known mutation, R154H, converts hOGG1 to a promutator by relaxing the specificity of the enzyme for the base opposite oxoG. PMID- 10706277 TI - Formation of molecular gas in the tidal debris of violent galaxy-galaxy interactions. AB - In many gravitational interactions between galaxies, gas and stars that have been torn from the precursor galaxies can collect in tidal 'tails'. Star formation begins anew in some of these regions, producing tidal dwarf galaxies. Observations of these new galaxies provides insight into processes relevant to galaxy formation more generally, because the timescale of the interaction is well defined. But tracking the star formation process has hitherto been difficult because the tidal dwarf galaxies with young stars showed no evidence of the molecular gas out of which those young stars formed. Here we report the discovery of molecular hydrogen (traced by carbon monoxide emission) in two tidal dwarf galaxies. In both cases, the concentration of molecular gas peaks at the same location as the maximum in atomic-hydrogen density, unlike the situation in most gas-rich galaxies. We infer from this that the molecular gas formed from the atomic hydrogen, rather than being torn in molecular form from the interacting galaxies. Star formation in the tidal dwarf galaxies therefore appears to mimic the process in normal spiral galaxies like our own. PMID- 10706278 TI - Geometric quantum computation using nuclear magnetic resonance AB - A significant development in computing has been the discovery that the computational power of quantum computers exceeds that of Turing machines. Central to the experimental realization of quantum information processing is the construction of fault-tolerant quantum logic gates. Their operation requires conditional quantum dynamics, in which one sub-system undergoes a coherent evolution that depends on the quantum state of another sub-system; in particular, the evolving sub-system may acquire a conditional phase shift. Although conventionally dynamic in origin, phase shifts can also be geometric. Conditional geometric (or 'Berry') phases depend only on the geometry of the path executed, and are therefore resilient to certain types of errors; this suggests the possibility of an intrinsically fault-tolerant way of performing quantum gate operations. Nuclear magnetic resonance techniques have already been used to demonstrate both simple quantum information processing and geometric phase shifts. Here we combine these ideas by performing a nuclear magnetic resonance experiment in which a conditional Berry phase is implemented, demonstrating a controlled phase shift gate. PMID- 10706279 TI - Reduction in the surface energy of liquid interfaces at short length scales AB - Liquid-vapour interfaces, particularly those involving water, are common in both natural and artificial environments. They were first described as regions of continuous variation of density, caused by density fluctuations within the bulk phases. In contrast, the more recent capillary-wave models assumes a step-like local density profile across the liquid-vapour interface, whose width is the result of the propagation of thermally excited capillary waves. The model has been validated for length scales of tenths of micrometres and larger, but the structure of liquid surfaces on submicrometre length scales--where the capillary theory is expected to break down--remains poorly understood. Here we report grazing-incidence X-ray scattering experiments that allow for a complete determination of the free surface structure and surface energy for water and a range of organic liquids. We observe a large decrease of up to 75% in the surface energy of submicrometre waves that cannot be explained by capillary theory, but is in accord with the effects arising from the non-locality of attractive intermolecule interactions as predicted by a recent density functional theory. Our data, and the results of comparable measurements on liquid solutions, metallic alloys, surfactants, lipids and wetting films should thus provide a stringent test for any new theories that attempt to describe the structure of liquid interfaces with nanometre-scale resolution. PMID- 10706280 TI - Electrically induced structure formation and pattern transfer AB - The wavelength of light represents a fundamental technological barrier to the production of increasingly smaller features on integrated circuits. New technologies that allow the replication of patterns on scales less than 100 nm need to be developed if increases in computing power are to continue at the present rate. Here we report a simple electrostatic technique that creates and replicates lateral structures in polymer films on a submicrometre length scale. Our method is based on the fact that dielectric media experience a force in an electric field gradient. Strong field gradients can produce forces that overcome the surface tension in thin liquid films, inducing an instability that features a characteristic hexagonal order. In our experiments, pattern formation takes place in polymer films at elevated temperatures, and is fixed by cooling the sample to room temperature. The application of a laterally varying electric field causes the instability to be focused in the direction of the highest electric field. This results in the replication of a topographically structured electrode. We report patterns with lateral dimensions of 140 nm, but the extension of the technique to pattern replication on scales smaller than 100 nm seems feasible. PMID- 10706281 TI - Variations of Younger Dryas atmospheric radiocarbon explicable without ocean circulation changes AB - The concentration of radiocarbon, 14C, in the atmosphere depends on its production rate by cosmic rays, and on the intensity of carbon exchange between the atmosphere and other reservoirs, for example the deep oceans. For the Holocene (the past approximately 11,500 years), it has been shown that fluctuations in atmospheric radiocarbon concentrations have been caused mostly by variations in the solar magnetic field. Recent progress in extending the radiocarbon record backwards in time has indicated especially high atmospheric radiocarbon concentrations in the Younger Dryas cold period, between 12,700 and 11,500 years before the present. These high concentrations have been interpreted as a result of a reduced exchange with the deep-ocean reservoir, caused by a drastic weakening of the deep-ocean ventilation. Here we present a high resolution reconstruction of atmospheric radiocarbon concentrations, derived from annually laminated sediments of two Polish lakes, Lake Gosciaz and Lake Perespilno. These records indicate that the maximum in atmospheric radiocarbon concentrations in the early Younger Dryas was smaller than previously believed, and might have been caused by variations in solar activity. If so, there is no indication that the deep-ocean ventilation in the Younger Dryas was significantly different from today's. PMID- 10706282 TI - A thermodynamic explanation for black smoker temperatures AB - There is a remarkable difference between the maximum temperature of black smoker effluent (350 degrees C-400 degrees C) and the temperature of the solidifying magma which heats it (approximately 1,200 degrees C). It has been suspected for some time that the nonlinear thermodynamic properties of water might be responsible for this discrepancy. Here, we translate this hypothesis into a physical model, by examining the internal temperature structure of convection cells in a porous medium. We demonstrate that, at pressures appropriate to seafloor crust, plumes of pure water form naturally at approximately 400 degrees C for any heat source with temperature greater than approximately 500 degrees C. Higher temperatures are confined to a boundary layer at the base of the convection cell, where the flow is horizontal. The phenomenon is explained analytically using the thermodynamic properties of water, and is illustrated by numerical simulations. Our model predicts the existence of the high-temperature 'reaction zone' found in ophiolites and suggests that vent temperatures will remain steady as magma chambers solidify and cool. PMID- 10706283 TI - An interconnected network of core-forming melts produced by shear deformation AB - The formation mechanism of terrestrial planetary cores is still poorly understood, and has been the subject of numerous experimental studies. Several mechanisms have been proposed by which metal--mainly iron with some nickel--could have been extracted from a silicate mantle to form the core. Most recent models involve gravitational sinking of molten metal or metal sulphide through a partially or fully molten mantle that is often referred to as a 'magma ocean'. Alternative models invoke percolation of molten metal along an interconnected network (that is, porous flow) through a solid silicate matrix. But experimental studies performed at high pressures have shown that, under hydrostatic conditions, these melts do not form an interconnected network, leading to the widespread assumption that formation of metallic cores requires a magma ocean. In contrast, here we present experiments which demonstrate that shear deformation to large strains can interconnect a significant fraction of initially isolated pockets of metal and metal sulphide melts in a solid matrix of polycrystalline olivine. Therefore, in a dynamic (non-hydrostatic) environment, percolation remains a viable mechanism for the segregation and migration of core-forming melts in a solid silicate mantle. PMID- 10706284 TI - Rapid evolution of reproductive barriers driven by sexual conflict. AB - A growing amount of experimental data indicates extremely rapid evolution of traits and proteins related to fertilization in many diverging taxa. These data come from studies of sperm or pollen competition between closely related species, and from molecular studies of fertilization proteins. The positive selection for evolutionary novelty that appears to be acting on fertilization systems seems paradoxical because successful reproduction requires the close matching of female and male traits. It has been suggested that perpetual coevolution between the sexes can result from sexual conflict in mating. Sexual conflict occurs when characteristics that enhance the reproductive success of one sex reduce the fitness of the other sex. Numerous examples of sexual conflict resulting from sensory exploitation, polyspermy and the cost of mating have been discussed in detail. The potential for coevolution due to such conflict has been evaluated experimentally. Here I develop a simple mathematical model describing coevolutionary dynamics of male and female traits involved in reproduction. The model shows that continual change in such traits at a constant speed is expected whenever females (or eggs) experience fitness loss from having too many compatible males (or sperms). The plausibility of runaway coevolution increases with increasing population size. Rapid evolution of reproductive barriers driven by sexual conflict may explain increased speciation rates after colonization of new habitats ('adaptive radiation') and high species richness in resource-rich environments. PMID- 10706285 TI - Thermal stimulation of taste. AB - The first electrophysiological recordings from animal and human taste nerves gave clear evidence of thermal sensitivity, and studies have shown that as many as half of the neurons in mammalian taste pathways respond to temperature. Because temperature has never been shown to induce sensations of taste, it has been assumed that thermal stimulation in the gustatory system is somehow nulled. Here we show that heating or cooling small areas of the tongue can in fact cause sensations of taste: warming the anterior edge of the tongue (chorda tympani nerve) from a cold temperature can evoke sweetness, whereas cooling can evoke sourness and/or saltiness. Thermal taste also occurs on the rear of the tongue (glossopharyngeal nerve), but the relationship between temperature and taste is different there than on the front of the tongue. These observations indicate the human gustatory system contains several different types of thermally sensitive neurons that normally contribute to the sensory code for taste. PMID- 10706286 TI - Postsaccadic visual references generate presaccadic compression of space. AB - With every rapid gaze shift (saccade), our eyes experience a different view of the world. Stable perception of visual space requires that points in the new image are associated with corresponding points in the previous image. The brain may use an extraretinal eye position signal to compensate for gaze changes, or, alternatively, exploit the image contents to determine associated locations. Support for a uniform extraretinal signal comes from findings that the apparent position of objects briefly flashed around the time of a saccade is often shifted in the direction of the saccade. This view is challenged, however, by observations that the magnitude and direction of the displacement varies across the visual field. Led by the observation that non-uniform displacements typically occurred in studies conducted in slightly illuminated rooms, here we determine the dependence of perisaccadic mislocalization on the availability of visual spatial references at various times around a saccade. We find that presaccadic compression occurs only if visual references are available immediately after, rather than before or during, the saccade. Our findings indicate that the visual processes of transsaccadic spatial localization use mainly postsaccadic visual information. PMID- 10706287 TI - A neurofibromatosis-1-regulated pathway is required for learning in Drosophila. AB - The tumour-suppressor gene Neurofibromatosis 1 (Nf1) encodes a Ras-specific GTPase activating protein (Ras-GAP). In addition to being involved in tumour formation, NF1 has been reported to cause learning defects in humans and Nf1 knockout mice. However, it remains to be determined whether the observed learning defect is secondary to abnormal development. The Drosophila NF1 protein is highly conserved, showing 60% identity of its 2,803 amino acids with human NF1 (ref. 12). Previous studies have suggested that Drosophila NF1 acts not only as a Ras GAP but also as a possible regulator of the cAMP pathway that involves the rutabaga (rut)-encoded adenylyl cyclase. Because rut was isolated as a learning and short-term memory mutant, we have pursued the hypothesis that NF1 may affect learning through its control of the Rut-adenylyl cyclase/cAMP pathway. Here we show that NF1 affects learning and short-term memory independently of its developmental effects. We show that G-protein-activated adenylyl cyclase activity consists of NF1-independent and NF1-dependent components, and that the mechanism of the NF1-dependent activation of the Rut-adenylyl cyclase pathway is essential for mediating Drosophila learning and memory. PMID- 10706288 TI - Three distinct and sequential steps in the release of sodium ions by the Na+/K+ ATPase. AB - The Na+/K+ pump, a P-type ion-motive ATPase, exports three sodium ions and then imports two potassium ions in each transport cycle. Ions on one side of the membrane bind to sites within the protein and become temporarily occluded (trapped within the protein) before being released to the other side, but details of these occlusion and de-occlusion transitions remain obscure for all P-type ATPases. If it is deprived of potassium ions, the Na+/K+ pump is restricted to sodium translocation steps, at least one involving charge movement through the membrane's electric fields. Changes in membrane potential alter the rate of such electrogenic reactions and so shift the distribution of enzyme conformations. Here we use high-speed voltage jumps to initiate this redistribution and show that the resulting pre-steady-state charge movements relax in three identifiable phases, apparently reflecting de-occlusion and release of the three sodium ions. Reciprocal relationships among the sizes of these three charge components show that the three sodium ions are de-occluded and released to the extracellular solution one at a time, in a strict order. PMID- 10706289 TI - The 21-nucleotide let-7 RNA regulates developmental timing in Caenorhabditis elegans. AB - The C. elegans heterochronic gene pathway consists of a cascade of regulatory genes that are temporally controlled to specify the timing of developmental events. Mutations in heterochronic genes cause temporal transformations in cell fates in which stage-specific events are omitted or reiterated. Here we show that let-7 is a heterochronic switch gene. Loss of let-7 gene activity causes reiteration of larval cell fates during the adult stage, whereas increased let-7 gene dosage causes precocious expression of adult fates during larval stages. let 7 encodes a temporally regulated 21-nucleotide RNA that is complementary to elements in the 3' untranslated regions of the heterochronic genes lin-14, lin 28, lin-41, lin-42 and daf-12, indicating that expression of these genes may be directly controlled by let-7. A reporter gene bearing the lin-41 3' untranslated region is temporally regulated in a let-7-dependent manner. A second regulatory RNA, lin-4, negatively regulates lin-14 and lin-28 through RNA-RNA interactions with their 3' untranslated regions. We propose that the sequential stage-specific expression of the lin-4 and let-7 regulatory RNAs triggers transitions in the complement of heterochronic regulatory proteins to coordinate developmental timing. PMID- 10706290 TI - Pgh1 modulates sensitivity and resistance to multiple antimalarials in Plasmodium falciparum. AB - Throughout the latter half of this century, the development and spread of resistance to most front-line antimalarial compounds used in the prevention and treatment of the most severe form of human malaria has given cause for grave clinical concern. Polymorphisms in pfmdr1, the gene encoding the P-glycoprotein homologue 1 (Pgh1) protein of Plasmodium falciparum, have been linked to chloroquine resistance; Pgh1 has also been implicated in resistance to mefloquine and halofantrine. However, conclusive evidence of a direct causal association between pfmdr1 and resistance to these antimalarials has remained elusive, and a single genetic cross has suggested that Pgh1 is not involved in resistance to chloroquine and mefloquine. Here we provide direct proof that mutations in Pgh1 can confer resistance to mefloquine, quinine and halofantrine. The same mutations influence parasite resistance towards chloroquine in a strain-specific manner and the level of sensitivity to the structurally unrelated compound, artemisinin. This has important implications for the development and efficacy of future antimalarial agents. PMID- 10706291 TI - The LIM homeobox gene Lhx9 is essential for mouse gonad formation. AB - During mammalian embryonic development, the ovaries and testes develop from somatic cells of the urogenital ridges as indifferent gonads, harbouring primordial germ cells that have migrated there. After sex determination of the gonads, the testes produce testosterone and anti-Mullerian hormone which mediate male sexual differentiation, and the female developmental pathway ensues in their absence. Here we show that transcripts of the LIM homeobox gene Lhx9 are present in urogenital ridges of mice at embryonic day 9.5; later they localize to the interstitial region as morphological differentiation occurs. In mice lacking Lhx9 function, germ cells migrate normally, but somatic cells of the genital ridge fail to proliferate and a discrete gonad fails to form. In the absence of testosterone and anti-Mullerian hormone, genetically male mice are phenotypically female. The expression of steroidogenic factor 1 (Sf1), a nuclear receptor essential for gonadogenesis, is reduced to minimal levels in the Lhx9-deficient genital ridge, indicating that Lhx9 may lie upstream of Sf1 in a developmental cascade. Unlike mice lacking other genes that mediate early stages of gonadogenesis, Lhx9 mutants do not exhibit additional major developmental defects. Thus, LHX9 mutations may underlie certain forms of isolated gonadal agenesis in humans. PMID- 10706292 TI - The S receptor kinase determines self-incompatibility in Brassica stigma. AB - The self-incompatibility possessed by Brassica is an intraspecific reproductive barrier by which the stigma rejects self-pollen but accepts non-self-pollen for fertilization. The molecular/biochemical bases of recognition and rejection have been intensively studied. Self-incompatibility in Brassica is sporophytically controlled by the polymorphic S locus. Two tightly linked polymorphic genes at the S locus, S receptor kinase gene (SRK) and S locus glycoprotein gene (SLG), are specifically expressed in the papillar cells of the stigma, and analyses of self-compatible lines of Brassica have suggested that together they control stigma function in self-incompatibility interactions. Here we show, by transforming self-incompatible plants of Brassica rapa with an SRK28 and an SLG28 transgene separately, that expression of SRK28 alone, but not SLG28 alone, conferred the ability to reject self (S28)-pollen on the transgenic plants. We also show that the ability of SRK28 to reject S28 pollen was enhanced by SLG28. We conclude that SRK alone determines S haplotype specificity of the stigma, and that SLG acts to promote a full manifestation of the self-incompatibility response. PMID- 10706293 TI - Structure of the winged-helix protein hRFX1 reveals a new mode of DNA binding. AB - Regulatory factor X (RFX) proteins are transcriptional activators that recognize X-boxes (DNA of the sequence 5'-GTNRCC(0-3N)RGYAAC-3', where N is any nucleotide, R is a purine and Y is a pyrimidine) using a highly conserved 76-residue DNA binding domain (DBD). DNA-binding defects in the protein RFX5 cause bare lymphocyte syndrome or major histocompatibility antigen class II deficiency. RFX1, -2 and -3 regulate expression of other medically important gene products (for example, interleukin-5 receptor alpha chain, IL-5R alpha). Fusions of the ligand-binding domain of the oestrogen receptor with the DBD of RFX4 occur in some human breast tumours. Here we present a 1.5 A-resolution structure of two copies of the DBD of human RFX1 (hRFX1) binding cooperatively to a symmetrical X box. hRFX1 is an unusual member of the winged-helix subfamily of helix-turn-helix proteins because it uses a beta-hairpin (or wing) to recognize DNA instead of the recognition helix typical of helix-turn-helix proteins. A new model for interactions between linker histones and DNA is proposed. PMID- 10706294 TI - Structure of a ligand-binding intermediate in wild-type carbonmonoxy myoglobin. AB - Small molecules such as NO, O2, CO or H2 are important biological ligands that bind to metalloproteins to function crucially in processes such as signal transduction, respiration and catalysis. A key issue for understanding the regulation of reaction mechanisms in these systems is whether ligands gain access to the binding sites through specific channels and docking sites, or by random diffusion through the protein matrix. A model system for studying this issue is myoglobin, a simple haem protein. Myoglobin has been studied extensively by spectroscopy, crystallography, computation and theory. It serves as an aid to oxygen diffusion but also binds carbon monoxide, a byproduct of endogenous haem catabolism. Molecular dynamics simulations, random mutagenesis and flash photolysis studies indicate that ligand migration occurs through a limited number of pathways involving docking sites. Here we report the 1.4 A resolution crystal structure of a ligand-binding intermediate in carbonmonoxy myoglobin that may have far-reaching implications for understanding the dynamics of ligand binding and catalysis. PMID- 10706295 TI - Abdominal insufflation does not cause hematogenous spread of colon cancer. AB - BACKGROUND AND PURPOSE: Previous investigators have suggested that port-site recurrences are possibly a result of abdominal insufflation, forcing viable cancer cells into the circulation to metastasize and thrive in areas of trauma. Using a syngeneic animal cancer model, we tested the hypothesis that pneumoperitoneum increases the incidence of wound metastasis by a blood-borne mechanism. METHODS: Male BD IX rats (N = 150) were injected intraperitoneally with 2 x 10(5) viable syngeneic 1,2-dimethylhydralazine-induced colon cancer cells (DHD-K12). Animals were divided into three groups: A (abdominal insufflation with 3-cm incision on the back into muscle remote from the peritoneum); B (3-cm back incision alone); and C (control group with 3-cm midline abdominal incision). Three weeks after surgery, the animals were euthanized and autopsied. RESULTS: In the two groups with back wounds, the incidence of cancer growth at the incision was zero, as demonstrated grossly and by histologic sample (A: 0/47, B: 0/43). In contrast, the autopsied control group had a 42% incidence of metastasis to the wound (25/59). There seemed to be no difference in the distribution of intra-abdominal disease between those rats that underwent insufflation and those that did not. CONCLUSION: It is unlikely that pneumoperitoneum promotes hematogenous wound implantation of free intraperitoneal cancer cells. PMID- 10706296 TI - Feasibility of 23-hour hospitalization after laparoscopic fundoplication. AB - PURPOSE: In order to reduce the costs of laparoscopic fundoplication, a pilot program for outpatient surgery was instituted in 1995. The risks and benefits of reducing postoperative hospitalization to < or =23 hours were assessed. PATIENTS AND METHODS: Patients in ASA grade I or II (N = 22) with refractory gastroesophageal reflux disease underwent laparoscopic fundoplication over a 21 month period in a hospital-affiliated outpatient facility. The results were compared with those of a similar group of 16 patients whose surgery was performed on an inpatient basis. RESULTS: Seventeen patients (77%) were discharged within 23 hours of surgery. The maximum length of stay was 3 days. There were no deaths. Nineteen patients (86%) reported excellent results. The average facility cost declined from $7,169 for the inpatient group to $4,588 for patients on operated under the outpatient protocol. The decrease resulted from a reduction in the cost of room, operating suite, supplies, and anesthesia. CONCLUSION: Laparoscopic fundoplication can be performed safely in a hospital-affiliated outpatient setting, resulting in a significant reduction in procedure costs. PMID- 10706297 TI - The learning curve of laparoscopic cholecystectomy and changes in indications: one institutions's experience with 2,650 cholecystectomies. AB - PURPOSE: In a prospective series of 2,650 consecutive patients undergoing cholecystectomy, we analyzed the learning curve since the introduction of laparoscopic cholecystectomy (LC) in terms of operating time, conversion rate, morbidity, mortality, and consequent changes in indications for either laparoscopic or open cholecystectomy (OC). PATIENTS AND METHODS: Between July 1990 and June 1997, LC was performed in 1,929 patients (73%), 203 of whom (7.5%) had to be converted to OC, while 518 patients (19.5%) had primary OC. Patients having LC were predominantly female, younger, with less comorbidity and less complicated gallstone disease than patients having OC. RESULTS: Barring a learning curve during the first 6 months of LC, operating time remained constant at an average of 71 minutes while operating on ever more complex pathologies. The conversion rate decreased from 9.4% to 6.7% during the 7-year period. A relatively constant team of surgeons with growing experience as well as constantly improving technical equipment allowed the complication rate to remain low. The total morbidity of LC was 2.5% (0.1% bile duct injury), that of conversions 5%, and that of OC 12.5%. The mortality was 0 for LC, 0.5% for conversions, and 1% for OC. CONCLUSION: The indications for primary OC decreased from 50% to 8.5% and the indications for LC could be broadened over the years. PMID- 10706298 TI - Laparoscopic anterior lumbar fusion. AB - PURPOSE: To assess the feasibility and complications of the laparoscopic approach to anterior lumbar fusion and to evaluate the ability of metoclopramide in conjunction with preoperative bowel preparation and early oral feeding to decrease postoperative ileus and reduce the length of hospital stay. PATIENTS AND METHODS: Laparoscopic anterior lumbar fusion was performed on 30 patients with persistent back pain between September 1997 and March 1999. All patients received metoclopramide 10 mg intravenously preoperatively and every 6 hours postoperatively, then 10 mg orally every 8 hours for 7 days. An experienced laparoscopic surgeon exposed the disc space, and lumbar fusion was performed by a neurosurgeon or an orthopedic surgeon. RESULTS: One procedure in an obese patient was converted to open surgery. The average operating time for the remaining patients was 2 hours 23 minutes. The average estimated blood loss was 75 mL. The only intraoperative complication was a trocar injury to the bladder. The average hospital stay was 2.3 days. CONCLUSION: In properly selected patients, laparoscopic anterior lumbar fusion with metoclopramide, preoperative bowel preparation, and early oral feeding results in a short hospital stay and yields better cosmetic results than traditional surgery. PMID- 10706299 TI - Controlled balloon dilatation for laparoscopic extraperitoneal bladder neck suspension in patients with previous abdominal surgery. AB - PURPOSE: The balloon dilator is used in a variety of minimally invasive preperitoneal and retroperitoneal operations. In this study, we compared the ability to create an extraperitoneal cavity using a balloon spacer in patients with and without previous abdominal surgery undergoing laparoscopic bladder neck suspension. PATIENTS AND METHODS: This prospective study included 38 patients in total, 15 of whom had had previous abdominal wall surgery and 23 who had not. A balloon spacer technique was used to develop the extraperitoneal space. RESULTS: In 80% of the patients with previous surgery, the introduction of the balloon spacer was recorded as simple; in 20%, it was considered difficult. In 78% of the patients without previous surgery, the introduction of the balloon spacer was recorded as simple, in 17% it was difficult, and in 4% it failed. In 80% of the patients with previous surgery, the extraperitoneal view was good or acceptable, in 20% it was poor, and in 13% it failed. In 92% of the patients without previous surgery, the extraperitoneal view was good or acceptable, in 4% it was poor, and in 4% dilatation failed. Morbidity was equally divided between the groups. CONCLUSIONS: Previous abdominal surgery is not a contraindication to laparoscopic extraperitoneal surgery using a balloon spacer. The approach carries low morbidity, similar to that in patients without previous abdominal surgery. PMID- 10706300 TI - Laparoscopic subtotal cholecystectomy: a review of 56 procedures. AB - BACKGROUND AND PURPOSE: The essential surgical steps in laparoscopic cholecystectomy remain similar to those of open cholecystectomy. Positive identification of the biliary anatomy, safe clipping or ligature of the cystic duct and artery, and dissection of the gallbladder from the liver bed form the basis of cholecystectomy. Subtotal cholecystectomy is a definitive and safe operation under certain adverse conditions intraoperatively for dissection of the gallbladder from the liver bed. We reviewed our experience with laparoscopic cholecystectomy over a 2-year period between June 1996 and May 1998, when 1,680 operations were performed. The objective was to analyze the pathology, review surgical procedures, and trace the outcome of laparoscopic subtotal cholecystectomy. PATIENTS AND METHODS: In 56 of 1,680 patients, laparoscopic subtotal cholecystectomy was performed, which constituted 3.33% of the laparoscopic cholecystectomies performed at our institution. Dense fibrosis and adhesions were present in 32 patients; 12 patients had Mirizzi syndrome, 6 patients had a sessile gallbladder, and 6 patients had a gangrenous gallbladder. The Endo-GIA 30 stapler was used in 40 patients, sequential clips were used in 9 patients, and a suture for stump closure was used in 5 patients. A subhepatic drain was inserted in 50 patients. RESULTS: Two conversions to open surgery were needed because of gangrene of the gall bladder wall and one conversion as a result of continued bleeding from the cystic artery after application of the Endo GIA 30 stapler. The mean postoperative stay in hospital was 2.5 days. One patient had a solitary bile duct calculus extracted at endoscopic retrograde cholangiopancreatography 3 months after surgery. Three patients had biliary drainage that lasted for a week, and four patients had epigastric port-site infections that resolved with antibiotics, dressings and postural drainage. CONCLUSION: Laparoscopic subtotal cholecystectomy is safe, feasible, and effective and may help prevent conversion to open surgery in carefully selected patients with difficult cholecystectomies. PMID- 10706301 TI - Fundoplication: a model for immunologic aspects of laparoscopic and conventional surgery. AB - BACKGROUND AND PURPOSE: Immunologic investigations of laparoscopic and conventional procedures have recently been performed during cholecystectomy or colon resection, but the results might have been influenced by the amount of dissection or the presence of malignant tumor. Because fundoplication is characterized by moderate dissection and no resection, we supposed it to be an appropriate procedure for comparing immunologic changes during laparoscopic and conventional surgery. PATIENTS AND METHODS: Immunologic analysis (interleukin [IL]-6, IL-10, leukocytes, HLA-DR monocytes) was carried out on the peripheral blood of 34 patients who underwent elective Nissen fundoplication by the laparoscopic (LAP; N = 26) or conventional (OPEN; N = 8) technique for gastroesophageal reflux disease. Blood samples were obtained before and 1 and 4 hours after the beginning of the operation and on days 1, 2, 4, 7 after the procedure. RESULTS: A very fast and significant (P < 0.01) increase of the proinflammatory cytokines (IL-6, IL-10) and leukocytes and a decrease of cell mediated functions (HLR-DR monocytes) were detected. Most of the analyzed measures had returned to preoperative values by 2 days after the procedure. All of the changes were similar in the two groups with the exception of IL-6. Throughout the post-operative study period, IL-6 concentrations were higher in the OPEN group, being significant 4 hours, 1 day, 2 days, and 4 days after the operation. CONCLUSION: The investigation measures do not give evidence that laparoscopic fundoplication is superior to conventional fundoplication in its immunologic effects. PMID- 10706302 TI - Laparoscopic diagnosis and treatment of nontraumatic acute abdominal pain in women. AB - PURPOSE: We reviewed the records of 77 women treated for nontraumatic acute abdomen by the principal author between June 1991 and June 1996. All patients presented to either the surgeon's office or the emergency room at Northwest Hospital, which is an urban community hospital in North Seattle. Our objectives in the study were to determine the effectiveness of diagnostic laparoscopy for nontraumatic acute abdomen and the percentage of cases managed using laparoscopic technique exclusively. PATIENTS AND METHODS: The mean patient age was 36.5 (range 12-65) years. The majority of these women (92%) were premenopausal. Seventy-two (93.5%) were Caucasian, and the remaining 5 (6.5%) were Asian. Thirty-eight of the women (49%) had undergone at least one prior pelvic or abdominal operation, and 28 (36%) had undergone more than one. The principal author performed preoperative clinical evaluations, then diagnostic laparoscopy for all 77 patients. RESULTS: Laparoscopy provided a definitive diagnosis in 76 of the 77 cases. In 70% of the cases (54 of 77) the preoperative diagnosis was confirmed by diagnostic laparoscopy, and in 29% (22 of 77), the diagnosis was confirmed, yet augmented or clarified, by diagnostic laparoscopy. In the remaining case, diagnostic laparoscopy ruled out any acute etiology. Ninety-five percent of the patients (72 of 76) were treated exclusively by laparoscopy (70 cases) or a laparoscopy-assisted procedure (2 cases). Four patients (5%) required conversion to laparotomy. The remaining patient required no therapeutic surgery. Mortality was 0 and morbidity 4%. CONCLUSION: A high proportion of women presenting with acute abdominal pain can be managed using a laparoscopic technique exclusively. PMID- 10706303 TI - Comparison of efficiencies of three techniques for colon surgery. AB - PURPOSE: To determine the most efficient technique for performing a colectomy, we used the methodology of time-motion analysis. METHODS: The efficiency of five hand-assisted and six regular laparoscopic colectomies and one open colectomy, performed by four surgeons in three different hospitals, was measured. The open colectomy was analyzed as a reference procedure. RESULTS AND CONCLUSIONS: The hand-assisted laparoscopic technique was the most efficient. Hand-assisted laparoscopy was therefore less time consuming than laparoscopic surgery. Open surgery was the fastest technique, because the time for every surgical motion is a factor of three shorter than for the two laparoscopic techniques. PMID- 10706304 TI - Laparoscopic aortorenal bypass: a feasibility study. AB - PURPOSE: To demonstrate the technical feasibility of laparoscopic aortorenal bypass in an acute porcine model. MATERIALS AND METHODS: An aorta-to-left renal artery bypass using an interposition Dacron graft was performed in five pigs. Intracorporeal laparoscopic free-hand suturing and knot-tying were employed exclusively. Renoprotective in-situ regional hypothermia was achieved intracorporeally by infusing ice-cold heparinized saline into the renal artery using a balloon catheter. RESULTS: The mean total surgical time was 325 minutes, and the mean renal ischemia time was 61 minutes. The end-to-side graft-to-aorta and end-to-end graft-to-renal artery anastomosis times were 34 minutes and 40 minutes, respectively. The mean estimated blood loss was 66 mL. On revascularization, prompt reperfusion of the kidney and Doppler-confirmed pulsation of the renal artery was noted. Graft patency was confirmed on autopsy. CONCLUSION: Laparoscopic aortorenal bypass is feasible. This study represents the initial report in the literature. A long-term animal survival study is planned. PMID- 10706305 TI - Retroperitoneoscopic excision of a mesenteric cyst. AB - Mesenteric cysts are rare intra-abdominal lesions. We present a case of a mesenteric cyst that was discovered by abdominal computed tomography (CT) and excised by retroperitoneoscopic surgery. There have been 10 reports of excision of mesenteric cysts by laparoscopy in the literature, but retroperitoneoscopic resection of such cysts has not been reported. This case suggests that when a mesenteric cyst arises from the ascending or descending colon, the retroperitoneal approach has a lower risk of traumatizing the bowel than does the laparoscopic intra-abdominal approach, and it does not have to compress other intra-abdominal organs. PMID- 10706306 TI - Videoscopic argon beam coagulator treatment of large persistent lymphocele. AB - Lymphocele formation is a common complication of axillary and inguinal lymphadenectomy. Treatment of persistent lymph collections by excision, open or percutaneous drainage, and sclerosing agents is often associated with infection, pain, prolonged treatment, and unacceptable rates of recurrence. We report our successful experience with a videoscopic approach to a large, chronic groin lymphocele using the argon beam coagulator. When feasible, the videoscopic use of the argon beam coagulator allows the patient the benefits of a minimally invasive approach. PMID- 10706307 TI - Pet travel scheme: participating companies announced. PMID- 10706308 TI - Trends of antimicrobial resistance in bacterial isolates from a small animal referral hospital. AB - A longitudinal, retrospective investigation was made of antimicrobial resistance in bacterial isolates obtained from clinical cases in a small animal hospital between 1989 and 1997. Isolates of Escherichia coli and Staphylococcus species were used as Gram-negative and Gram-positive indicator organisms, respectively, and the annual prevalence of antimicrobial resistance was calculated for each organism to each of nine (for E coli) and 11 (for Staphylococcus species) appropriate antimicrobials, including enrofloxacin. Using a chi-square test for trend, statistically significant, rising trends were identified in the resistance of E coli to amoxycillin (P=0.04), clavulanate-amoxycillin (P<0.01) and streptomycin (P<0.01), and in the resistance of Staphylococcus species to erythromycin (P<0.01). There was an equivocal, rising trend for the resistance of Staphylococcus species to cephalexin. No significant trends were apparent for any of the other 15 organism/drug interactions. The annual prevalence of multiple drug resistance was calculated for E coli, Proteus species, Pseudomonas species, staphylococci and streptococci, but no statistically significant trends were identified. PMID- 10706309 TI - Ground reaction force analysis of large breed dogs when walking after the amputation of a limb. AB - Force plate analysis was used to measure ground reaction forces (GRF) and contact times, and calculate the centre of gravity at a walk of 10 dogs which had had a limb amputated, and the results were compared with the results from 22 normal dogs of the same weight. The loss of a limb caused significant changes in the GRF, impulses and contact times of the remaining limbs and in the location of the dogs' centre of gravity. The changes were greater in dogs which had lost a forelimb than in dogs which had lost a hindlimb. PMID- 10706310 TI - Detection of DNA restriction fragment polymorphisms in Streptococcus equi. AB - Large-restriction-fragment (LRF) polymorphisms in Streptococcus equi (S equi subspecies equi) were studied by pulsed-field gel electrophoresis. Five or six chromosomal fragments of between 194 and 915 kb were separated by digestion with the restriction endonuclease Notl. All 20 isolates of S equi, including 12 from independent Japanese outbreaks, four from independent American outbreaks, two from a single Irish outbreak, us vaccine strain F43, and type strain NCTC 9682 were successfully typed. Seven distinctive, reproducible and stable types were identified. The 12 Japanese isolates collected between 1992 and 1998 were of LRF type II suggesting that they were derived from the same source. The remaining eight isolates were of six types. The results indicate that LRF typing should be a useful technique for investigating the source and transmission of S equi. PMID- 10706311 TI - Enzootic calcinosis in goats caused by golden oat grass (Trisetum flavescens). PMID- 10706312 TI - Trisomy 26 mosaicism in a sterile Holstein-Friesian heifer. PMID- 10706313 TI - Strategic use of gonadotrophins in first litter sows after weaning. PMID- 10706314 TI - Detection of anti-Brucella antibodies in pinnipeds from the Antarctic territory. PMID- 10706315 TI - Isolation and characterisation of a mycoplasma from a kittiwake (Rissa tridactyla). PMID- 10706316 TI - Identification of Clostridium perfringens enterotoxin in penguins. PMID- 10706317 TI - Insurance and the Pet Travel Scheme PMID- 10706318 TI - Midland Counties Veterinary Association PMID- 10706319 TI - The first report of the Systolic and Pulse Pressure (SYPP) Working Group. PMID- 10706320 TI - Blood pressure levels and measurement of subclinical vascular disease. AB - Increased blood pressure, especially systolic blood pressure (SBP), is linearly associated with an increased risk of cardiovascular disease and stroke. The attributable risk of vascular disease due to elevated blood pressure is greater for normal and high normal blood pressures and stage 1 hypertension than for more advanced hypertension (stages 2 and 3). The development of noninvasive methods for measuring the effects of blood pressure on the vascular system help identify early vascular disease, high risk populations at similar levels of blood pressure, and successful methods of therapy. The future goals of the control of elevated blood pressure should be to prevent: (1) the rise in blood pressure associated with increasing age; (2) the development of subclinical vascular disease associated with even slightly elevated blood pressure; and (3) clinical disease. Measurement of subclinical vascular disease should be a component of both observational epidemiological studies and clinical trials. The subclinical measures should include those primarily associated with the progression of atherosclerosis and the pathophysiology of elevated blood pressure. PMID- 10706321 TI - Pulse pressure and cardiovascular risk. AB - Numerous studies have shown that increases in diastolic (DBP) and systolic (SBP) blood pressure are positively associated with cardiac events. Since DBP typically varies by smaller amounts, it has historically been the blood pressure value used most often to assess the risk of cardiovascular disease. Further studies have indicated, however, that SBP continues to increase proportionally with age, while DBP levels off. New data have emerged which suggest that DBP can no longer be valued as a reliable predictor of cardiovascular events and may even be diagnostically misleading. Normal levels of DBP can be indicative of a combination of an increase in peripheral vascular resistance (which elevates DBP) with an increase in arterial stiffening (which decreases DBP). These two phenomena 'cancel' each other out; the underlying risk factors are disguised by the apparent normalcy of the DBP reading. DBP is even less of an indicator in older adults, where the prevailing form of hypertension is isolated systolic hypertension and the most common cause is large-artery stiffness. Since arterial stiffening causes SBP to increase and DBP to decrease, the gap between the two, pulse pressure (PP), may be the best predictor of cardiac events for all the blood pressure values. Several studies have confirmed that subjects with the widest PP have the greatest risk of mortality. Pulse pressure has also been observed to be a significant and independent indicator of myocardial infarction. Furthermore, compelling evidence has emerged that PP is a strong indicator of cardiovascular risk even among normotensive individuals. PMID- 10706322 TI - A new model of risk: implications of increasing pulse pressure and systolic blood pressure on cardiovascular disease. AB - Diastolic blood pressure (DBP) has historically functioned as the primary indicator of cardiovascular disease (CVD). Many clinical and observational, long term, large population studies, including the Framingham Heart Study and Multiple Risk Factor Intervention Trial (MRFIT), have repeatedly demonstrated a positive correlation between CVD events and DBP. However, unexplained results, often emerged from the data, in the form of a DBP threshold (where lower rates of DBP were not associated with lower disease rates) and a J-shaped relationship (a decline of DBP that correlated with an increased risk of CVD), have complicated the association of blood pressure to events. In the past decade studies have focused on other indicators to explain these irregularities. From these clinical and observational studies, new evidence has become available which strongly indicates that systolic blood pressure (SBP) and pulse pressure (PP) may be more reliable predictors of CVD events than DBP. Indeed, in several studies, a wide baseline PP has been identified as the only measure of blood pressure that is an independent risk factor for cardiac events. Moreover, pretreatment SBP has linearly predicted future CVD events in treated patients, while DBP has not. These studies of treated patients have also led to the identification of pretreatment risk factors that can be applied to stratify risk among patients who go on to have controlled blood pressure on treatment. This new information can more precisely identify patients at increased risk of CVD events and thus make it possible to more precisely tailor therapy to enhance the potential for CVD prevention. PMID- 10706323 TI - Ageing and hypertension: the assessment of blood pressure indices in predicting coronary heart disease. AB - OBJECTIVE: To determine whether pulse pressure (PP), diastolic blood pressure (DBP), systolic blood pressure (SBP), or mean arterial pressure (MAP) is the superior haemodynamic predictor for the risk of coronary heart disease (CHD). METHODS: Age-related changes of blood pressure in normotensive and untreated hypertensive subjects in a population-based cohort from the Framingham Heart Study were characterized. The relationship between these blood pressure indices and risk of CH D over a 20-year follow-up period were then evaluated. RESULTS: There was a parallel linear rise in SBP, DBP and MAP from age 30-49 years, suggesting increased peripheral vascular resistance (PVR) in this age group. After age 50-60 years, DBP declined, PP rose steeply, and MAP levelled off, while SBP continued to show a linear increase, suggesting increasing predominance of large artery stiffness (LAS) in this middle-aged and elderly group. After adjusting for age, sex and other risk factors, MAP and DBP consistently underestimated PVR and risk of CHD. Systolic blood pressure fully represented PVR but frequently underestimated LAS and risk of CHD. Pulse pressure was superior to SBP as a surrogate marker for LAS and predictor of CHD risk in the presence of discordantly low DBP, but PP frequently underestimated PVR. CONCLUSIONS: In middle-aged and older subjects, at any given level of SBP > or = 120 mmHg, the risk of CHD rose with discordantly lower DBP, suggesting that the wider PP was driving the risk for CHD. These results suggest that total haemodynamic load, defined as the sum of pulsatile load (LAS) and steady-state load (PVR), is a major determinant of CHD risk. PMID- 10706324 TI - Epidemiological aspects of pulse pressure and arterial stiffness. AB - Systolic and diastolic blood pressure are considered to be exclusive haemodynamic predictors of cardiovascular risk in humans. If high blood pressure is characterized as a mechanical factor which acts on the arterial wall with deleterious consequences, then the totality of the blood pressure curve should be investigated to determine the risk. The purpose of this review is to show that, in addition to systolic and diastolic blood pressure, other haemodynamic variables should be taken into account; namely, brachial pulse pressure, pulse pressure amplification, aortic pulse wave velocity, and carotid incremental elastic modulus. These new components might modify our conventional views on antihypertensive agents and may serve to identify new potentially active molecules. PMID- 10706325 TI - Vascular wall function as a risk marker for cardiovascular disease. AB - Elevated systolic blood pressure (SBP) and wide pulse pressure (PP) are both important risk markers for cardiovascular disease. They are indicative of a vascular abnormality, most likely located at the endothelial-vascular interface, where the balance between vasodilator and vasoconstriction determines vascular tone, structure and change in blood pressure. This abnormality makes vessel walls vulnerable to degeneration. Reducing blood pressure in patients with endothelial vascular dysfunction has been found to reduce the rates of cardiovascular events. Preliminary data also suggest that drug therapy has successfully improved endothelial dysfunction. Endothelial release of nitric oxide has little effect on the aorta, but affects the elasticity of thinner-walled branch vessels and arterioles beneficially. Nitric oxide is normally released in response to increased blood flow and this process is often impaired by various disease states. Since blood flow can be measured, endothelial function can be assessed. Arterial compliance can be determined clinically by a technique that combines pulse wave recording and computer analysis of diastolic decay. In certain disease states, compliance seems to be reduced more in smaller vessels, a marker of early stage vascular disease. Screening for arterial elasticity with this tool can identify those patients with early stages of arterial wall disease and prevent further damage. However, further research is needed to determine whether the benefit lies in lowering blood pressure or in the actual effect of the drug in restoring the function of the arterial wall. PMID- 10706326 TI - Wave travel and reflection in the arterial system. AB - Blood pressure indices reveal pertinent information regarding the risk factors associated with certain cardiovascular conditions. In the last decade, research has focused on determining which module of measurement is most relevant as well as exploring new investigative techniques to aid in detection and treatment of cardiovascular disease (CVD). The rates of isolated systolic hypertension, arteriosclerosis and other diseases associated with ageing continue to rise as the elderly population grows. The cuff sphygmomanometer only measures blood pressure in the brachial artery. Pulse wave analysis (PWA) is a noninvasive method of generating the ascending aorta pressure wave from the arterial pressure pulse measured in the carotid or radial artery. Pulse wave analysis identifies the effects of increased pulse wave velocity (PWV), an indicator of arterial stiffness, and an independent measure of cardiovascular risk. As there is a discrepancy between pulse readings at different sites, PWA is invaluable in providing information on the left ventricle and on large arteries. This measurement can reveal the unseen beneficial effects of antihypertensive drugs, as ascending aortic systolic and pulse pressure often decrease more than brachial pressure, and can indicate the presence of spurious systolic hypertension. The technique is not difficult to learn and results have been reproduced in several studies. Continuing research into PWA needs to be done, however, to assure the reliability of the measurements and minimize the possibility of incorrect readings. PMID- 10706327 TI - Isolated systolic hypertension in the elderly: lessons from clinical trials and future directions. AB - Systolic blood pressure (SBP) is a more reliable predictor of cardiovascular disease (CVD) events than is diastolic blood pressure (DBP). Perhaps the reduction of SBP should be more of the imperative of treatment than the reduction of DBP. Although two recent guidelines (WHO/ISH and JNC-VI) have recommended treating SBP to goal, there seems to be a reluctance in the medical community to embrace this paradigm shift and revise treatment plans. The deleterious effects of ignoring these findings are especially damaging to those with isolated systolic hypertension (ISH), which affects approximately two-thirds of hypertensive patients between the ages of 65 and 89 years. Two large clinical trials, the Systolic Hypertension in the Elderly Program (SHEP) and the Systolic Hypertension in Europe (Syst-EUR) trial have confirmed that reducing SBP in the elderly with stages 2 and 3 ISH (SBP > or = 160 mmHg with DBP < 90 mmHg in SHEP and SBP > 160 mmHg with DBP < 95 mmHg in Syst-EUR) reduced morbidity and mortality. Two SHEP sub-studies found that lowering SBP in subjects with non insulin-dependent diabetes mellitus (NIDDM) and those with a history of myocardial infarction (MI) reduced the incidence of stroke and heart failure, as well as several other endpoints. The beneficial effects were corroborated in Syst EUR where stroke (fatal and nonfatal), CVD endpoints and mortality were all significantly reduced when SBP was lowered 20 mmHg in subjects > 60 years of age. Despite these findings, however, recent analysis suggests that most hypertension treatment decisions continue to be based on DBP measurements instead of SBP. To combat this treatment gap, we must disseminate this information and motivate physicians and other providers to include reduction of SBP in their treatment plans. We must also encourage the development of antihypertensive drugs that lower SBP more effectively than those that are currently available. PMID- 10706329 TI - Comparison of serological and sequence-based methods for typing feline calcivirus isolates from vaccine failures. AB - Feline calicivirus (FCV) can be typed by exploiting antigenic differences between isolates or, more recently, by the sequence analysis of a hypervariable region of the virus's capsid gene. These two methods were used to characterise FCV isolates from 20 vaccine failures which occurred after the use of a commercial, live attenuated vaccine. Using virus neutralisation, the isolates showed a spectrum of relatedness to the vaccine; depending on the criterion adopted for identity, 10 to 40 per cent of them appeared to be similar to the vaccine virus. Using sequence analysis, the isolates fell into one of two categories; 20 per cent had a similar sequence to the vaccine (0-67 to 2-67 per cent distant), and the remainder had a dissimilar sequence (21-3 to 36-0 per cent distant). Sequence analysis identified one cat that appeared to be infected with two distinct FCVs. The serological and sequence-based typing methods gave the same result in 80 to 95 per cent of individual cases, depending on the criterion adopted for serological identity. It is suggested that molecular typing is a more definitive method for characterising the relatedness of FCV isolates. PMID- 10706328 TI - Systolic blood pressure and pulse pressure: role of 24-h mean values and variability in the determination of organ damage. AB - An increasing amount of data suggests that systolic blood pressure (SBP) and pulse pressure (PP) may more closely relate to and thus favour the atherogenic process than does diastolic blood pressure (DBP). The baseline data from the ongoing European Lacidipine Study on Atherosclerosis (ELSA) recently indicated that carotid artery atherosclerosis in normocholesterolaemic patients with mild or moderate essential hypertension is more closely related to SBP and more so PP than to DBP and lipid variables. Other new data point to the effects of hypertension on arterial compliance, as well as the effects of 24-h blood pressure variability on arterial compliance and distensibility. When viewed in their entirety, these data present a compelling case for the closer monitoring of SBP and PP with respect to arterial compliance, and the need for aggressive blood pressure treatment to control and perhaps reverse the underlying pathological changes in arterial structure and function in hypertensive patients. PMID- 10706330 TI - Risk assessment of organoleptic postmortem inspection procedures for pigs. AB - A systematic quantification of foodborne hazards in abnormal and normal tissues of pig carcases was undertaken to provide a risk-based assessment of the effectiveness of traditional organoleptic meat inspection. A total of 36,059 pigs, representing all major pig-producing areas and systems in Australia, were inspected on a seasonal basis at three abattoirs over 12 months. The prevalence of grossly detectable abnormalities of possible food-borne disease significance was recorded. A subset of the grossly detectable abnormalities, together with tissues classified by inspection as normal (controls) were submitted for the detection of a broad range of food-borne hazards. The potential exposure of consumers to hazards in fresh pork was characterised as the number of carcases per 10,000 containing hazards in selected tissues. The results indicated that the level of exposure of consumers to microbiological hazards in fresh pork is unlikely to be reduced significantly by the detection and removal of gross abnormalities in the tissues examined. On the basis of carcase throughput, the rate of contamination of normal lymph nodes was commonly 100 times higher, and no hazards were isolated from two types of grossly abnormal nodes. While further processing, cooking and handling may alter the exposure characterisation, the study nevertheless identifies the proportional contribution of abnormal and normal tissues to risks to consumers and clearly identifies the need for consideration of 'visual only' inspection in the re-evaluation of traditional inspection procedures. PMID- 10706331 TI - Seroprevalence of coxiellosis in cattle, sheep and people in the east of Turkey. AB - Serum samples collected randomly from 416 cattle in 48 herds, and 411 sheep in 47 flocks, in eight different locations in the east of Turkey between June and December 1998, were examined by indirect fluorescent antibody test (IFAT) to determine the prevalence of Q fever. The age, sex, breed, tick control and abortion history of the animals were also recorded. In addition, 102 serum samples were collected from apparently healthy people who were at risk of contracting the disease, such as farmers, veterinarians, abattoir and laboratory workers, and veterinary students. Seropositivity was observed in 5-8 per cent (24/416) of the cattle in 17 (35-4 per cent) of the herds and in 10-5 per cent (43/411) of the sheep in 21 (44-7 per cent) of the flocks. Eight of the 102 people were seropositive, with the highest prevalence (12-0 per cent) in farmers and abattoir workers. All the seropositive farmers had seropositive animals. None of the laboratory workers or veterinary students was seropositive. PMID- 10706332 TI - Intestinal submucosa repair in two cases of feline ulcerative keratitis. PMID- 10706333 TI - Dura mater laceration associated with acute paraplegia in three dogs. PMID- 10706334 TI - Supraorbital trichomoniasis infection in two saker falcons (Falco cherrug). PMID- 10706336 TI - Ionising radiations regulations 1999. PMID- 10706335 TI - PMWS and porcine dermatitis nephropathy syndrome in Great Britain. PMID- 10706337 TI - Insurance and the pet travel scheme PMID- 10706338 TI - Cattle identification. PMID- 10706339 TI - TB and husbandry. PMID- 10706340 TI - Membership fees for retired MsRCVS PMID- 10706341 TI - Survey of sheep farmers PMID- 10706342 TI - Physician's prayer PMID- 10706343 TI - Reciprocal social behavior in children with and without pervasive developmental disorders. AB - An invariant feature of pervasive developmental disorders (PDDs) is a relative deficit in the capacity for reciprocal social behavior (RSB). The authors acquired teacher reports of RSB in 287 schoolchildren and parent reports of RSB in 158 child psychiatric patients using a new research instrument, the Social Reciprocity Scale. Total scores on this measure of RSB were continuously distributed in all groups of subjects; children with PDDs scored significantly higher for the degree of deficits in RSB than did clinical or nonclinical controls. Latent class analysis and factor analysis failed to demonstrate separate categories of deficiency for core autistic symptomatology and more general impairments in RSB, consistent with the notion of a "broader autism phenotype." Assessments of RSB on a continuous scale may be useful clinically for characterizing the behavior of children whose social deficits fall below the threshold for a full diagnosis of autism. They may also be useful in genetic linkage studies of autistic spectrum disorders. PMID- 10706344 TI - Neurodevelopmental and neuroimaging correlates in nonsyndromal microcephalic children. AB - This descriptive study examined the relationship between head size, developmental functioning, and neuroimaging findings in children with absolute microcephaly. Subjects, aged 1 to 48 months, were assigned to one of two groups based on occipitofrontal head circumference (OFC). Group A included subjects with an OFC of 2 to 2.99 standard deviations below the mean, and Group B included subjects with an OFC of 3 or more standard deviations below the mean. Brain scan findings for 62% of the subjects were abnormal. Findings included cerebral atrophy, cortical dysplasia, myelination delay, and white matter hypoplasia. Mean scores for developmental measures in Groups A and B were less than 70. Mean developmental scores in the normal imaging group were 70 or greater, whereas developmental scores in the abnormal imaging group were 52 or less. Forty-three percent of the subjects in Group A and 80% of those in Group B had abnormal findings from imaging studies (p = .0394). Subjects with one or more brain abnormalities determined on the basis of magnetic resonance images or computed tomographic scans had significantly lower scores in all developmental areas (p < .05). The authors concluded that abnormal brain images seem to be a better reflection of developmental performance than the degree of microcephaly. J Dev Behav Pediatr 21:12-18, 2000. Index terms: microcephaly, neuroimaging, neurodevelopment. PMID- 10706345 TI - Thirty-six-month outcome of prenatal cocaine exposure for term or near-term infants: impact of early case management. AB - Gestational cocaine use is associated with serious pregnancy complications having fetal and neonatal implications. However, many cocaine-abusing women deliver uneventfully at term. The purpose of this study was to assess the neurodevelopmental outcome for term or near-term infants after prenatal cocaine exposure and to determine whether that outcome would be modified by early, intensive family case management. Cocaine-exposed infants identified after delivery at an urban hospital were alternately assigned to receive case management (n = 70) or routine follow-up (n = 48). A matched, non-drug-exposed group of infants was identified for comparison (n = 41). Infants aged up to 36 months were serially evaluated in a multidisciplinary clinic with cognitive, psychomotor, and language testing. Group comparisons were performed using one-way analysis of variance. There were no statistical differences in mean cognitive, psychomotor, or language quotients between cocaine-exposed and non-drug-exposed infant groups aged up to 36 months. At 6 months of age, case-managed cocaine exposed infants had a significantly higher mean Bayley Mental Developmental Index score than those who were routinely managed. However, no differences were present at subsequent assessments. Among cocaine-exposed infants who remained with their mothers at 36 months, verbal scores were significantly higher for case-managed compared with routine-managed infants. The negative effects of urban, low socioeconomic status may overshadow the impact of prenatal cocaine exposure on early childhood outcome for those infants born without prenatal complications. PMID- 10706346 TI - Sleep habits and sleep disturbance in elementary school-aged children. AB - Relatively little is known about sleep habits, sleep disturbances, and the consequences of disordered sleep in school-aged children. This descriptive study examined a variety of common sleep behaviors in a group of 494 elementary school children, grades kindergarten through fourth, using a battery of sleep questionnaires that included parent, teacher, and self-report surveys. The prevalence of parent-defined sleep problems ranged from 3.7% (Sleep-Disordered Breathing) to 15.1% (Bedtime Resistance), with 37% of the overall sample described as having significant sleep problems in at least one sleep domain. Younger children were more likely than older children to have sleep problems noted by parents (particularly bedtime struggles and night wakings), as well as by teacher and self-report. Children tended to identify more sleep problems by self-report, particularly sleep-onset delay and night wakings, than did their parents. Overall, approximately 10% of the sample was identified by all three measures as having significant problems with daytime sleepiness. The results of this study emphasize the importance of screening for sleep disorders in this age group in the clinical setting. The need for consensus regarding the use of sleep screening instruments and the definition of "problem" sleep in school-aged children is also discussed. PMID- 10706347 TI - Parents' ratings of everyday cognitive abilities in very low birth weight children. AB - Parents' ratings of everyday cognitive functioning in very low birth weight (VLBW) children free of sensorineural impairments and normal birth weight (NBW) children were compared with the children's actual performance on psychometric measures of cognitive and motor skills. Subjects included 19 VLBW children identified at age 3 years as "suspect" for developmental problems, 19 VLBW children identified at age 3 years as "developing normally," and 30 NBW full-term peers. Results indicated that parents of the suspect VLBW group rated their children as having significant impairments in memory, language, cognitive, and motor skills, findings which were consistent with the results of concurrent psychometric assessments. When compared with psychometric test results, parents identified more children as displaying difficulties in memory, language, and cognitive skills, but fewer children with coordination difficulties. These findings suggest that the parents' ratings and the psychometric measures may be assessing somewhat different aspects of the children's functioning. PMID- 10706348 TI - Child development and the civil society--does social capital matter? PMID- 10706349 TI - Developing civil society through the promotion of positive youth development. PMID- 10706350 TI - Social capital and developmental health: making the connection. PMID- 10706351 TI - ADHD, divorce, and parental disagreement about the diagnosis and treatment. PMID- 10706352 TI - Impact of recurrent and chronic pain on child and family daily functioning: a critical review of the literature. AB - The author reviewed the current status of research on the impact of recurrent and chronic pain on everyday functioning of children and families and organized the research findings around the specific life contexts (e.g., school, peers) that may be affected by pain. Although findings demonstrate that many different aspects of child and family life are affected by pain, the prevalence and severity of children's functional limitations associated with pain remain unknown. Few treatment studies for pediatric recurrent and chronic pain have focused on enhancing children's functioning. It has been shown, however, that functional outcomes can be improved by cognitive-behavioral interventions. Recommendations for research on functional outcomes and implications for clinical practice are discussed. PMID- 10706353 TI - Radioulnar synostosis, radial ray abnormalities, and severe malformations in the male: a new X-linked dominant multiple congenital anomalies syndrome? AB - We describe a multiple congenital anomalies (MCA) syndrome dominantly transmitted through three generations. Radial ray abnormalities with wide variability of expression were observed in four female patients. Moreover, a 14-week-gestation male fetus had severe radial ray malformation, anencephaly, unilateral renal agenesis, and a common dorsal mesentery. Results of high-resolution karyotyping were normal in the malformed fetus and his affected mother. Furthermore, several spontaneous abortions of male fetuses had occurred in this pedigree. To our knowledge, a similar association has not been described previously. It could represent a new X-linked dominant MCA syndrome, or an autosomal dominant condition with severe expression limited to males. PMID- 10706354 TI - Familial occurrence of isolated right ventricular hypoplasia. AB - Isolated right ventricular hypoplasia is a rare congenital anomaly. This condition is usually associated with a communication between the atria in the form of a patent foramen ovale or secondum atrial septal defect. We describe a familial occurrence of this rare disease. A 1-day-old male child and his 34-year old father were found to have isolated right ventricular hypoplasia with atrial septal defect. An autosomal dominant mode of inheritance is likely for this rare congenital anomaly. PMID- 10706355 TI - Growth and developmental retardation, ocular ptosis, cardiac defect, and anal atresia: confirmation of the ROCA-Wiedemann syndrome. AB - We report on a girl with growth and mental retardation, peculiar face with ptosis, epicanthus, broad nasal bridge, low-set and abnormal ears, cleft uvula, congenital heart defect, and anal atresia. A similar condition was reported previously by Wiedemann et al. [1982: An atlas of characteristic syndromes: a visual aid to diagnosis, 2nd ed. p 114-115]. We confirm the existence of this condition that, although similar to Ohdo syndrome, seems to be an independent clinical entity. We propose that, based on the principal clinical manifestations, this condition should be identified with the acronym ROCA (retardation of growth and development, ocular ptosis, cardiac defect, and anal atresia). PMID- 10706356 TI - Genetic counseling for women with an intermediate family history of breast cancer. AB - Women with a family history of breast cancer often over-estimate their personal risk for cancer and may view themselves as candidates for genetic testing even when the likelihood of an informative test result is low. We report here on genetic counseling of women with an intermediate family history of breast cancer, defined as women who have one or more biological relatives with breast cancer but whose pedigree suggests a low likelihood of autosomal dominant transmission. A genetic counseling protocol based on traditional genetic counseling strategies was developed with additional components added to address the needs of women with moderately increased breast cancer risk. These additional components included information about non-genetic risk factors, comparisons of high and moderate risk pedigrees, and evaluation of personal risk based on both genetic and nongenetic risk factors. Most participants liked the genetic counseling and found it useful. At baseline, participants over-estimated both their personal risk of breast cancer and that of the average woman. After counseling, estimates of personal and average risk of breast cancer were lower, although both remained higher than actual risk. Most participants reported that they felt less worried about breast cancer after receiving their personal-risk estimate. At baseline, most women judged themselves to be candidates for genetic testing and expressed interest in testing. The number who considered themselves candidates for testing was reduced after counseling (60% versus 82%) but still constituted a majority. Similarly, interest in testing was partially reduced by counseling (60% versus 91%). We conclude that genetic counseling can help women with an intermediate family history of breast cancer to develop more accurate views of their risk, reduce their breast cancer worry, and aid some of them in developing a more realistic view of genetic testing. PMID- 10706357 TI - Interstitial tandem duplication of 6p: a case with partial trisomy (6)(p12p21.3). AB - A de novo interstitial tandem duplication of 6p12p21.3 was observed in a 7-month old boy with growth retardation, psychomotor delay and craniofacial, brain, limb, and genital anomalies. Fluorescent in situ hybridization using a chromosome 6 paint probe demonstrated that the extra material belonged to chromosome 6. Although it has been suggested that 6p25 is the critical band involved in the expression of the phenotype of 6p duplication, comparison of the clinical findings of this case with those from the literature cases showed strong similarities. PMID- 10706358 TI - Prenatal exposure to valproic acid during pregnancy and limb deficiencies: a case control study. AB - We conducted a case-control study using data from the Spanish Collaborative Study of Congenital Malformations (ECEMC) on the relationship between prenatal exposure to valproic acid (VPA) and the presence of limb deficiencies in newborn infants. Among a total of 22,294 consecutive malformed infants (once we excluded genetic syndromes) and 21,937 control infants with specified data on antiepileptic drugs during gestation, 57 malformed infants and 10 control infants were exposed to VPA during the first trimester of pregnancy. Of the total of malformed infants exposed to VPA, 36.8% (21/57) presented with congenital limb defects of different types (including overlapping digits, talipes, clubfoot, clinodactyly, arachnodactyly, hip dislocation, pre- and postaxial polydactyly, etc.), three of them having limb deficiencies. The result of the case-control analysis shows a risk for limb deficiencies of odds ratio = 6.17 [confidence interval (CI) 1.28 29.66, P = 0.023], after controlling for potential confounder factors. If we consider that in our population the prevalence at birth of this type of defect is 6.88 per 10,000 livebirths (95% CI 6.43-7.36) we can estimate that the risk for women treated with VPA of having a baby with limb deficiencies would be around 0.42%. The limb deficiencies in the three patients exposed to VPA were the following: the first case was a newborn infant with hypoplasia of the left hand, the second patient was a newborn infant with unilateral forearm defect and hypoplastic first metacarpal bone in the left hand, and the third patient presented with short hands with hypoplastic first metacarpal bone, absent and hypoplastic phalanges, retrognathia, facial asymmetry, hypospadias, teleangiectatic angioma in skull, and hypotonia. PMID- 10706359 TI - Ring chromosome 22 and autism: report and review. AB - Ring chromosome 22 has been described in over 50 cases. A characteristic phenotype has not been fully delineated; however, long face, thick eyebrows, 2-3 toe syndactyly, mental retardation, adequate somatic growth and the absence of major malformations are noted in many cases. An 11-year-old boy with ring chromosome 22 and 46,XY,r(22)(p11.31-q13.31 approximately q13.33) karyotype presented with global developmental delay, autistic disorder, and dolichocephaly, apparently low-set and large ears, midface hypoplasia, and 2-3 toe syndactyly. This is the second report of a ring chromosome 22 with autistic disorder. There appears to be an association between abnormalities of chromosome 22, including r(22), and autistic disorder; however, this occurrence may be a result of the association of autistic disorder with mental retardation rather than specifically due to r(22). The physical findings in this case also suggest that ring chromosome 22 causes a subtle but distinct phenotype which has previously been proposed. PMID- 10706360 TI - Abnormal timing in the prenatal ossification of vertebral column and hand in Crouzon syndrome. AB - We report on a radiographically examined fetus (gestational age 13 weeks) with Crouzon syndrome caused by a mutation in the gene encoding the fibroblast growth factor 2 (FGFR2). We found an approximately 2-week delay in vertebral body and hand ossification with normal vertebral arch ossification, suggesting that regionally delayed skeletal maturation might be a manifestation of FGFR2 mutation syndromes. The findings support other studies indicating that different signaling pathways control skeletal maturation in vertebral bodies and vertebral arches. PMID- 10706361 TI - Manifestations and linkage analysis in X-linked autoimmunity-immunodeficiency syndrome. AB - The clinical findings of a kindred with an X-linked disorder are characterized by autoimmune polyendocrinopathy, enteropathy with villous atrophy, chronic dermatitis, and variable immunodeficiency. Linkage analysis was performed on 20 members of the affected kindred to determine the location of the responsible locus. Informative recombinations limited the region to an approximate 20 cM interval bordered by DXS1055 and DXS1196/DXS1050. Multipoint analysis generated a lod score >3 for the region contained between DXS8024 and DXS8031. The candidate region includes the Wiskott-Aldrich syndrome (WAS) locus. Evaluation of the Wiskott-Aldrich syndrome protein gene by single strand conformational analysis, heteroduplex analysis, and direct sequencing of the 12 exons in an affected male and two carrier females revealed no abnormalities. We conclude that this kindred has an X-linked disorder, distinct from WAS, that results in autoimmunity and variable immunodeficiency. The responsible locus maps to the pericentromeric region Xp11.23 to Xq21.1. PMID- 10706362 TI - Molecular diagnosis of Stickler syndrome: a COL2A1 stop codon mutation screening strategy that is not compromised by mutant mRNA instability. AB - We have developed a novel strategy for screening families with type 1 Stickler syndrome due to COL2A1 nonsense mutations, using a modified RNA-based protein truncation test. To overcome the problem of the unavailability of collagen II producing cartilage cells, reverse transcription polymerase chain reaction (RT PCR) was performed on the illegitimate transcripts of accessible cells (lymphoblasts and fibroblasts), which were pre-incubated with cycloheximide to prevent nonsense-mutation-induced mRNA decay. The five overlapping RT-PCR fragments covering the COL2A1 coding region were then transcribed and translated in vitro to identify smaller truncated protein products which result from a premature stop codon. This method was used to screen a 4-generation Stickler family and a protein truncating mutation was identified, which was present in all affected individuals. Targeted sequencing identified the mutation as a G(+1) to A substitution at the 5' splice donor site of intron 25, which led to the activation of a cryptic splice site 8-bp upstream causing aberrant mRNA splicing and a translational frameshift that introduced a premature stop codon. Mutant mRNA was undetectable without cycloheximide protection, demonstrating that the mutant mRNA was subjected to nonsense-mediated mRNA decay. As well as providing further evidence that type 1 Stickler syndrome results from COL2A1 premature stop codon mutations, this study suggests mutant mRNA instability leading to haploinsufficiency may also be an important, but previously unrecognized, molecular basis of Stickler syndrome. This rapid new test for COL2A1 nonsense mutations is of particular clinical importance to Stickler syndrome families, where the identification of individuals who are at risk of this potentially preventable form of blindness will allow them to undergo regular ophthalmological surveillance and preventative or early ameliorative treatment. PMID- 10706363 TI - Fronto-otopalatodigital osteodysplasia: clinical evidence for a single entity encompassing Melnick-Needles syndrome, otopalatodigital syndrome types 1 and 2, and frontometaphyseal dysplasia. AB - Otopalatodigital syndrome type 2 is an X-linked disorder with minimal expression in carrier females and comprises typical facial anomalies and a generalized bone dysplasia with osteodysplastic changes, brachydactyly, and impaired survival. Recently several other severe malformations were reported in the condition. Melnick-Needles syndrome is an X-linked dominant disorder. Affected males are usually sporadic cases. The exceptional males born to symptomatic women present with a lethal disorder comprising generalized osteodysplasia, deficiency of the first ray, and facial anomalies strikingly similar to those of otopalatodigital syndrome type 2. We report here on three boys with classical, severe, and lethal otopalatodigital type 2 syndrome, and three boys with severe (lethal) Melnick Needles syndrome, born to affected mothers. We suggest that otopalatodigital type 1 and 2, Melnick-Needles syndrome and frontometaphyseal dysplasia, sharing many clinical manifestations and a similar mode of inheritance, are variants of the same condition: fronto-otopalatodigital osteodysplasia. The relationships to similar syndromes (i.e., Saint-Martin-Gardner-Morrisson syndrome, serpentine fibula syndrome, atelosteogenesis type 3, boomerang dysplasia, and Yunis-Varon syndrome) are discussed. PMID- 10706364 TI - Holoprosencephaly, hypertelorism, and ectrodactyly in a boy with an apparently balanced de novo t(2;4) (q14.2;q35). AB - A holoprosencephaly, hypertelorism, and ectrodactyly syndrome (HHES) was described in three previous cases in whom chromosomes were apparently normal. We report on a 3-year-old boy with HHES and a de novo apparently balanced t(2;4)(q14.2;q35) confirmed by fluorescent in situ hybridization. He had severe growth and mental retardation, lobar holoprosencephaly, hypertelorism, microphthalmos, and iris, choroid, and retina colobomata. Less-severe facial involvement correlates with the semilobar type of holoprosencephaly; limb defects consisted of foot ectrodactyly and syndactyly. All previous HHES cases were sporadic and of unknown cause. A cryptic imbalance secondary to the translocation (2;4) in our patient may explain the phenotype. PMID- 10706365 TI - Otocephaly: prenatal diagnosis of a new case and etiopathogenetic considerations. PMID- 10706366 TI - Neuroblastoma in a mother and congenital central hypoventilation in her daughter: variable expression of the same genetic disorder? PMID- 10706367 TI - Neuroimaging and echocardiographic findings in Sotos syndrome. PMID- 10706368 TI - Memory for temporal order of new and familiar spatial location sequences: role of the medial prefrontal cortex. AB - Rats with medial prefrontal cortex or sham control lesions were tested on an eight-arm radial maze task to examine memory for the temporal order of a variable and a constant sequence of spatial locations as a function of temporal distance. During the study phase of each trial, rats were allowed to visit each of eight arms once in an order that was randomly selected or fixed for that trial. The test phase required the rats to choose which of two arms occurred earlier in the sequence of arms visited during the study phase. The arms selected as test arms varied according to temporal distance (0, 2, 4, or 6) or the number of arms that occurred between the two test arms in the study phase. For the variable sequences based on new information, control rats showed an increasing temporal distance function. Relative to control rats, medial prefrontal cortex-lesioned rats displayed a temporal order memory deficit across all distances. For the constant sequence based on familiar information, control rats performed well across all distances. Relative to controls, the medial prefrontal cortex-lesioned rats displayed a performance deficit. The results support the idea that the medial prefrontal cortex contributes to mnemonic operations associated with temporal order for new and familiar spatial location information. PMID- 10706369 TI - Visual space from visual motion: turn integration in tethered flying Drosophila. AB - Organisms navigating by path integration need to continuously measure their forward movement and their angular orientation with respect to an external reference. How they do it is little understood. Tethered flies at the flight simulator "navigate" in an artificial visual landscape without forward movement. They can return to a previously held orientation if the panorama provides a singularity (landmark) as reference. Surprisingly, in a regularly striped drum without singularities, they can use a temporal cue instead. In this experiment the arena is illuminated with only one color that is either green or blue. The arena is virtually divided into four quadrants. Whenever a quadrant boundary moves past an arbitrary point, the color of the arena light changes. When a fly is heated with one color it acquires a preference for the other one. Subsequently, it avoids the borders toward the potentially 'hot' quadrants even without touching them. The only way to achieve this is by turn integration, that is, by adding and subtracting all the turns it performs once it crosses the border. The color switch defining the border crossing resets the turn integrator, using the orientation of the arena at this moment as reference. In contrast, landmarks or, if it were available, the skylight compass enable the fly to establish by pattern learning any orientation as a reference. If the reference orientation coincides with the desired orientation, that is, if the animal stores the pattern while being oriented toward the goal, it can maintain its orientation without recourse to turn integration (which may be error prone). PMID- 10706370 TI - Behavioral manipulation of retrieval in a spatial memory task for Drosophila melanogaster. AB - A paradigm for operant conditioning of freely walking single Drosophila flies has been described previously. A fly can be conditioned to avoid one side of a small test chamber if the chamber is heated whenever the fly enters this side. In a subsequent memory test without heat the fly continues to avoid the previously heat-associated side. In this experimental design one cannot exclude that flies mark the heated side by an odor that they subsequently avoid during the test. As a final proof for associative learning in the present experiment, flies are trained in one chamber and tested for learning in another, similar one. Handling in the transfer experiment interferes with memory display, even if the fly is returned to the old chamber instead of a new one. Memory can be reactivated, however, by subjecting the fly to an additional brief training (priming), which is too short to establish significant learning in naive flies. For efficient priming, heat has to be applied to the same side as during training in the old chamber. Only then the fly subsequently shows a side preference and avoids the side of the new chamber, which in the old one had been associated with heat. The two chambers are similar but not identical The transfer experiment therefore raises the question as to what the flies use as spatial reference during training and test. In the light, they can be shown to orientate according to visual landmarks associated with the chamber. In complete darkness, where training and memory scores do not differ from those in the light, they are assumed to use a combination of tactile and idiothetic information for orienting. PMID- 10706371 TI - Neuroanatomical correlates of implicit and explicit memory for structurally possible and impossible visual objects. AB - Implicit memory refers to nonconscious retrieval of past experience demonstrated by facilitation in test performance on tasks that do not require intentional recollection of previous experiences. Explicit memory, in contrast, refers to the conscious retrieval of prior information, as demonstrated during standard recall and recognition tasks. In this experiment, positron emission tomographic (PET) measurements of regional cerebral blood flow (CBF), a marker of local neuronal activity, were used to identify and contrast brain regions that participate in the perception, implicit memory, and explicit memory for structurally possible and impossible visual objects. Ten CBF images were acquired in 16 normal women as they made possible/impossible and old/new recognition decisions about previously studied (old) and nonstudied (new) structurally possible and impossible objects. As reported previously, object decisions for familiar possible objects were associated with increased CBF in the vicinity of the left inferior temporal and fusiform gyri and recognition memory for familiar possible objects was associated with increased CBF in the vicinity of the right hippocampus. In this report, we provide more extensive analyses of the roles of the inferior temporal cortex, the hippocampus, the parahippocampus, and the pulvinar in encoding and retrieval operations. Additionally, patterns of CBF increases and decreases provide information regarding the neural structures involved in implicit and explicit memory. PMID- 10706372 TI - Reactivation-dependent changes in memory states in the terrestrial slug Limax flavus. AB - The change in memory state in the terrestrial slug Limax flavus was studied using cooling-induced retrograde amnesia. Slugs were first conditioned to avoid carrot odor and then a second conditioning procedure was applied 1, 3, 6, or 7 days after the first conditioning trial. Cooling the slugs to approximately 1 degrees C on day 7 immediately after the presentation of the odor used in the conditioning resulted in retrograde amnesia in the slugs that were subject to a second conditioning on day 6 or 7, but not in slugs that were subject to a second conditioning on day 1 or 3. Next, second-order conditioning was used as the second conditioning procedure to distinguish the memory acquired in the first conditioning from that acquired in the second conditioning and similar results were obtained. These results suggest that the reactivation of memory altered the memory state from a cooling-insensitive state to a cooling-sensitive one. A possible model for memory states is discussed. PMID- 10706373 TI - Characterization of 3-epi-1alpha,25-dihydroxyvitamin D3 involved in 1alpha,25 dihydroxyvitamin D3 metabolic pathway in cultured cell lines. AB - Using six different cultured cell models representing osteoblast, intestine, kidney and keratinocyte, we have demonstrated that 1alpha,25-dihydroxyvitamin D3 (1alpha,25(OH)2D3) is metabolized into 3-epi-1alpha,25(OH)2D3 in vitamin D-target cells. Although differences existed in the amount of 3-epi-1alpha,25(OH)2D3 formed with different cell types, it was apparent that 1alpha,25(OH)2D3 was subjected to metabolism both through the C24-oxidation and 3-epimerization pathways. Time course and dose response studies showed that the production of 3 epi-1alpha,25(OH)2D3 was enzymatic. It is interesting to note that this epimerization proceeded from 3beta towards 3alpha unidirectionally, and this conversion was not inhibited by ketoconazole. These data suggest that cytochrome P450 related enzymes including the 24-hydroxylase would not affect this reaction. The biological activity of 3-epi-1alpha,25(OH)2D3 was found to be lower than the native 1alpha,25(OH)2D3 in suppressing of proliferation of HL-60 cells, while the affinity of 3-epi-1alpha,25(OH)2D3 for vitamin D-binding protein was 2.5-fold higher than that of 1alpha,25(OH)2D3. The results indicate that 3-epimerization may change the pharmacokinetics and catabolism of 1alpha,25(OH)2D3 in vitamin D target cells. PMID- 10706374 TI - Changes of caspase activities involved in apoptosis of a macrophage-like cell line J774.1/JA-4 treated with lipopolysaccharide (LPS) and cycloheximide. AB - The addition of lipopolysaccharide (LPS) together with cycloheximide (CHX) induced apoptosis in a subline of a J774.1 macrophage-like cell line, JA-4, as judged by terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate (dUTP) nick end labeling (TUNEL)-staining and poly(adenosine 5' diphosphate (ADP)-ribose) polymerase (PARP)-cleavage. Caspase activities were examined in these macrophages in vitro using fluorogenic substrates such as acetyl-DEVD-aminomethyl coumarine (Ac-DEVD-AMC, caspase-3-like), acetyl-YVAD aminomethyl coumarine (Ac-YVAD-AMC, caspase-1-like), acetyl-VEID-aminomethyl coumarine (Ac-VEID-AMC, caspase-6-like), and carbobenzoxy-IETD-aminofluoro coumarine (Z-IETD-AFC; caspase-8-like). Kinetic studies revealed these caspase activities with different Km and Vmax values in extracts of apoptotic macrophages. In the course of apoptosis, caspase-3-like activity increased first at 75 min, simultaneously with the appearance of TUNEL staining and prior to PARP cleavage, and then caspase-6 and 8-like activities increased at 90 and 105 min, respectively. However, caspase-1-like activity did not change throughout the experiment. Furthermore, removal of LPS and CHX by extensive washing of the cells for 60 min completely abolished the apoptosis and the subsequent release of lactate dehydrogenase (LDH) during additional incubation until 4 h after LPS addition. However, washing of the cells after 75 min or later resulted in the progress of apoptosis and LDH release, which was coordinated with the elevation of caspase-3-like activity at 60 min and that of caspase-6 or 8-like activity at 90 min, but not with that of caspase-1-like activity. These results suggest that caspase-3-like activity represents the most apical caspase among these caspases in terms of the intiation of apoptosis in macrophages treated with LPS and CHX. In the present study, we also provide evidence on the relatively low specificities of a series of caspase inhibitors other than acetyl-DEVD-aldehyde (Ac-DEVD-CHO) which specifically inhibited the caspase-3-like activity. PMID- 10706376 TI - Competition in the metabolism of glycyrrhizin with glycyrrhetic acid mono glucuronide by mixed Eubacterium sp. GLH and Ruminococcus sp. PO1-3. AB - Eubacterium sp. GLH possessing glycyrrhizin (GL) and glycyrrhetic acid mono glucuronide (GAMG) beta-D-glucuronidases, Ruminococcus sp. PO1-3 possessing GL and GAMG beta-D-glucuronidases and 3beta-hydroxysteroid dehydrogenase and these mixed bacteria were cultured in GAM medium with and without GL, GAMG or both. GL added to Eubacterium sp. GLH accelerated the peaks of enhanced GL beta-D glucuronidase activity and suppressed GAMG beta-D-glucuronidase activity, and GAMG delayed the peaks of the enhanced growth with GL and GAMG beta-D glucuronidase activities. GL added to Ruminococcus sp. PO1-3 enhanced gradually the growth with GL and GAMG beta-D-glucuronidase activities, and GAMG enhanced slowly GL beta-D-glucuronidase activity and rapidly the growth with GAMG beta-D glucuronidase activity. The metabolite glycyrrhetic acid (GA) was produced by Eubacterium sp. GLH and Ruminococcus sp. PO1-3 in larger amounts and faster from GAMG than from GL. GL (1.0 mM) and 1.0 mM GAMG added to these mixed bacteria enhanced the growth with GL and GAMG beta-D-glucuronidase activities and were metabolized almost completely to GA in culture of 2 d and 1 d, respectively. It was found that the metabolism of GAMG was faster than that of GL. GL with GAMG added to mixed Eubacterium sp. GLH and Ruminococcus sp. PO1-3 cultured for 0 and 1 d led to a lower level of these enzyme activities and the consumption of GAMG more quickly, not GL. Low GAMG beta-D-glucuronidase had the ability to hydrolyze GAMG well. PMID- 10706375 TI - Two metallothioneins in the fresh-water fish, crucian carp (Carassius cuvieri): cDNA cloning and assignment of their expression isoforms. AB - Two metallothionein cDNAs (MT-A and MT-B) in the fresh-water fish crucian carp (Carassius cuvieri Temminck et Schlegel) were cloned. Sequence analysis of both cDNAs gave the structure of a coding region corresponding to 60 amino acid residues. The homology of their deduced amino acid sequences was completely conserved at the positions of the cysteine residue, but a significant difference existed in the size of their 3'-untranslated regions (130 base pairs for MT-A and 280 base pairs for MT-B). Direct amino acid sequencing of the MT-II isoform purified by HPLC was accomplished for up to 30 residues and its sequence was identical to that deduced from MT-B cDNA. This is the first case in vertebrates that N-terminal methionine in crucian carp MT-II was not blocked. By northern blot analysis, basal and cadmium chloride- or dexamethasone-induced MT-B (MT-II) mRNAs were detected time dependently after treatment. On the other hand, the expression of MT-A mRNA was extremely low. These results indicate that the MT isoform II in crucian carp is coded by the MT-B gene, and that the MT-B-dominant expression of mRNA in crucian carp may be due to the difference in the 3' untranslated regions of MT mRNAs. PMID- 10706377 TI - Catalytic properties for naphthoquinones and partial primary structure of rabbit heart acetohexamide reductase. AB - The catalytic properties of rabbit heart acetohexamide reductase (RHAR) for naphthoquinones were examined. RHAR efficiently reduced 1,4-naphthoquinone and juglone (5-hydroxy-1,4-naphthoquinone), whereas it had little or no ability to reduce menadione (2-methyl-1,4-naphthoquinone) or plumbagin (5-hydroxy-2-methyl 1,4-naphthoquinone). The structural requirements for these four naphthoquinones and one acetohexamide analog, and the kinetic mechanism for the inhibition of acetohexamide reduction by juglone led us to conclude that the 2-methyl group of menadione and plumbagin prevents access of the substrates to the catalytic site of RHAR. Five of six peptides derived from RHAR showed 30-42% residue identities with regions in the amino acid sequence of mouse lung carbonyl reductase (MLCR) belonging to the short-chain dehydrogenase/reductase (SDR) family. The catalytically important residues (Arg-39, Ser-136, Tyr-149 and Lys-153) of MLCR were found in the peptide sequences of RHAR, despite the low residue identities between the two enzymes. RHAR is probably best classified as a member of the SDR family similar to MLCR. PMID- 10706378 TI - Increased band 3 protein aggregation and anti-band 3 binding of erythrocyte membranes on treatment with sesamol. AB - The influence of sesamol, an antioxidant in processed sesame oil, on oxidative modification of human erythrocyte membrane proteins was investigated. Human erythrocytes were incubated with sesamol at various concentrations up to 10 mM at 37 degrees C for 1 h. The amounts of hemoglobin bound to the membranes and detergent C12E8-insoluble membrane protein aggregates were increased as the concentration of sesamol increased. Western blot analysis indicated that aggregates of band 3 protein were increased by the treatment. Binding of anti band 3 antibody to the erythrocytes was increased by the treatment. Isolated cell membranes were incubated with sesamol similarly. Aggregates of band 3 protein were also increased, indicating that the band 3 protein aggregation was little affected by hemoglobin bound to the membranes. Aggregation of band 3 protein in the treatment of isolated cell membranes was partially prevented when the treatment was conducted under anaerobic conditions, suggesting that augmentation of the protein aggregation by sesamol involved both oxygen-dependent and oxygen independent pathways. Among phenolics, sesamol showed a distinctive feature to increase band 3 protein aggregation in erythrocyte membranes and to enhance anti band 3 binding to erythrocytes. PMID- 10706379 TI - Analysis of promoter activity of 5'-upstream regions of zebrafish olfactory receptor genes. AB - The vertebrate olfactory system receives and discriminates a great variety of odorants. Many lines of evidence suggest that individual olfactory neuron expresses a single type or limited types of the olfactory receptor genes. However, the mechanism of selection of a single gene in the olfactory receptor family remains unclear. In the present study, we utilized zebrafish to identify the promoter element of the olfactory receptor genes in their 5'-upstream regions. First, we isolated a number of zebrafish olfactory receptor genes. These olfactory receptor genes were specifically expressed in the olfactory tissue as visualized by whole mount in situ hybridization analysis. Time of onset of the expression of each receptor clone varied from 24 h to 48 h postfertilization. Then, we injected various constructs containing the 5'-upstream regions of the olfactory receptor genes connected to beta-galactosidase reporter gene into fertilized zebrafish embryos. Constructs from two independent olfactory receptor genes exerted beta-galactosidase (promoter) activity that is specifically upregulated in the olfactory tissue. Use of either longer or deleted constructs of these two genes diminished the promoter activity in the olfactory tissue. From these results we discuss the mechanism of the transcription of the olfactory receptor genes in the olfactory neurons. PMID- 10706380 TI - Effects of YM358, an angiotensin II type 1 (AT1) receptor antagonist, and enalapril on blood pressure and vasoconstriction in two renal hypertension models. AB - The effects of the angiotensin II type 1 receptor antagonist YM358 on blood pressure were compared to those of the angiotensin converting enzyme inhibitor enalapril in one-kidney, one-clip renal hypertensive rats (1K1C RHR), two-kidney, one-clip renal hypertensive rats (2K1C RHR) and normotensive rats (NTR). Additionally, the local drug actions in peripheral tissues were investigated using isolated mesenteric arteries from these rats. In 2K1C RHR, YM358 and enalapril produced a long-lasting hypotensive effect in a dose-dependent manner. In 1K1C RHR, YM358 (30 mg/kg) also produced an antihypertensive effect, whereas enalapril (30 mg/kg) had no effect. Administration of YM358, but not enalapril, to 1K1C RHR, 2K1C RHR and NTR did not affect heart rate. In isolated mesenteric arteries from 1K1C RHR and 2K1C RHR, angiotensin II (Ang II), angiotensin I (Ang I) and tetradecapeptide (TDP), a physiologically active renin substrate, produced concentration-dependent vasoconstriction. YM358 (10(-7) M) inhibited the vasoconstricting responses to Ang II, Ang I and TDP in isolated mesenteric arteries. In contrast, enalaprilat (10(-7) M), an active metabolite of enalapril, did not completely inhibit the response to Ang I and TDP. These results indicate that YM358 has higher efficacy than enalapril for the treatment of hypertension. PMID- 10706381 TI - Pharmacological profile of VP-343, a novel selective vasopressin V2 receptor antagonist, in rats. AB - The pharmacological profile of a novel selective vasopressin V2 receptor antagonist, VP-343(N-[4-[[(2S,3aR)-2-hydroxy-2,3,3a,4-tetrahydropyrrolo[ 1,2 a]quinoxalin-5(1H)-yl]carbonyl]phenyl]-4'-methyl[1,1'-biphenyl ]-2-carboxamide) was characterized in several in vitro and in vivo rat models. The IC50 values of VP-343 for vasopressin V1A and V2 receptors were 110 and 0.77 nM, respectively. VP-343 inhibited dose-dependently the pressor response to exogenous arginine vasopressin (AVP; 30 mU/kg, i.v.) in pithed rats, with an ID50 value of 0.57 mg/kg (i.v.). VP-343 induced strong aquaresis in normal saline-loaded conscious rats. Antidiuretic activities of VP-343 have not been detected in AVP deficient Brattleboro rats, showing its lack AVP V2 agonistic activity. During repeated administration for 21 d (3 mg/kg, p.o.) and after recovery, the aquaretic action of VP-343 still remained. In the aged (17 month) saline-loaded conscious rats study, VP-343 (3 mg/kg, p.o.) exhibited remarkable diuretic action. In a single dose oral toxicity study in mice, VP-343 did not produce any clinical signs and mortality at any of the tested doses. The results indicate that VP-343 is a potent, orally active, selective V2 receptor antagonist, suggesting that it can be expected to be useful as an aquaretic drug. PMID- 10706382 TI - Studies of methyl 2-nitroimidazole-1-acetohydroxamate (KIN-804) 1: effect on free radical scavenging system in mice bearing Ehrlich ascites carcinoma. AB - Methyl 2-nitroimidazole-1-acetohydroxamate (KIN-804) is a 2-nitroimidazole derivative containing a hydroxamate side chain designed to enhance the radiosensitization response of hypoxic cells. The possible sensitization of tumor tissue by KIN-804 can be evaluated through investigation of the levels of the free radical scavengers; namely, glutathione (GSH) and its complex enzyme system including glutathione reductase (GR) and glutathione peroxidase (GSH-Px), as well as glucose-6-phosphate dehydrogenase (G-6-PD). Female albino mice were inoculated with Ehrlich carcinoma in the thigh. Administration of KIN-804 (i.p. 80 mg/kg body weight) was carried out 20 min before localized irradiation of 10 Gy. The data revealed that KIN-804 administration, followed or not by gamma irradiation, resulted in a significant decrease in GSH content in tumor tissues associated with inhibition in GR and G-6-PD activities. Blood GSH-Px was enhanced in tumor inoculated mice and the administration of KIN-804 returned it to the normal value. These changes were more noticeable in tumor bearing mice exposed to both KIN-804 and irradiation. PMID- 10706383 TI - Studies of methyl 2-nitroimidazole-1-acetohydroxamate (KIN-804) 2: effect on certain antioxidant enzyme systems in mice bearing Ehrlich ascites carcinoma. AB - In order to decrease toxicity and/or increase radiosensitizing activity, a new 2 nitroimidazole derivative, methyl 2-nitroimidazole-1-acetohydroxamate (KIN-804), was synthesized to solve the problem of tumor hypoxia. Evaluation of the efficiency of KIN-804 was carried out through studying the antioxidant enzyme system: The superoxide dismutase (SOD), catalase and lipid peroxide levels provide a rough index of the balance between free radical generation and scavenging. Female albino mice were inoculated with Ehrlich ascites carcinoma (EAC) in the thigh. The administration of KIN-804 (i.p. 80 mg/kg body weight) was carried out 20 min before localized irradiation of 10 Gy. In general, the data revealed that KIN-804 administration, followed or not by gamma irradiation, exerted significant inhibition of SOD and catalase activities accompanied by a significant increase in lipid peroxide level in tumor-bearing mice. PMID- 10706384 TI - The inhibition of phenylhydroquinone-induced oxidative DNA cleavage by constituents of Moutan Cortex and Paeoniae Radix. AB - Moutan Cortex (root cortex of Paeonia suffruticosa ANDREWS) and Paeoniae Radix (root of Paeonia lactiflora PALLAS) are crude drugs used in many traditional prescriptions and have constituents in common. We studied the effects of extracts of these crude drugs and their constituents on oxidative DNA damage caused by phenylhydroquinone (PHQ), a major metabolite of o-phenylphenol. Both drugs suppressed the cleavage of pUC18 DNA induced by PHQ, and scavenged the superoxide and hydroxy radical generated by the chemical. They also inhibited the oxidative DNA cleavage by tert-butylhydroquinone (TBHQ), one of the major metabolites of butylated hydroxyanisole. When constituents were examined with the same system, galloylpaeoniflorin and 1,2,3,4,6-penta-O-galloyl-beta-D-glucose (PGG) were found to be the most potent inhibitors of the DNA cleavage. These constituents had oxygen radical scavenging activity. Paeonol also attenuated the DNA cleavage. Paeoniflorin and albiflorin had relatively small inhibitory effects on DNA cleavage. However, catechin enhanced the PHQ-induced DNA cleavage. The suppression of oxidative DNA damage by Moutan Cortex and Paeoniae Radix might be attributable to the additive effects of galloylpaeoniflorin, PGG and other constituents. PMID- 10706385 TI - Evaluation of L-glutamate clearance capacity of cultured rat cortical astrocytes. AB - Astrocytes have a function in the uptake of excitatory neurotransmitter L glutamate via the glutamate transporter. To evaluate the L-glutamate clearance capacity of astrocytes, we developed a colorimetric method for the determination of L-glutamate concentration and measured changes in extracellular L-glutamate concentration in rat cortical astrocyte cultures. When L-glutamate (50-200 microM) was added to astrocyte cultures and incubated for 1-8 h, the extracellular L-glutamate concentration declined with time. When L-glutamate was mixed with astrocyte culture supernatants only, no significant change in L glutamate concentration was observed, ruling out the possibility that L-glutamate is spontaneously or enzymatically degraded in the extracellular space. Alternatively, the time-dependent decline of extracellular L-glutamate concentration was blocked by the presence of glutamate uptake inhibitors, indicating that the glutamate uptake system of astrocytes plays a major role in the clearance of extracellular L-glutamate. PMID- 10706386 TI - Role of prostaglandin E2 and leukotriene B4 in skin reaction induced by transdermal application of propranolol. AB - Dermal application of propranolol (PRL) induced formation of erythema and edema, and pseudoeosinophil infiltration in epidermis and dermis at the application site in guinea pigs. We investigated the production of prostaglandin E2 (PGE2) and leukotriene B4 (LTB4) at the application site of PRL and the role of these inflammatory chemical mediators in the occurrence of the skin reactions. PGE2 was found to be produced at the application site slightly after the accumulation of PRL released from the adhesive bandage in the patch test, and the amount of PGE2 increased continuously, with a peak value obtained at 24 h after application. The time-course changes resembled those of delta a* value, the index of erythema formation determined by colorimetric measurement, and edema formation. The production of PGE2 by dermal application of PRL was suppressed by local pretreatment with dexamethasone or indomethacin. However, no notable production of LTB4 was observed at the application site of PRL. PMID- 10706387 TI - Inhibitory effect of sanguiin H-11 on chemotaxis of neutrophil. AB - Neutrophil is accumulated by chemoattractants at inflammatory sites. In order to find a new substance to regulate the chemotaxis of neutrophil, we examined the effects of purified tannins and related compounds (33 species). Among those studied, sanguiin H-11, purified from the plant Sanguisorba officinalis, was the most potent inhibitor of rat cytokine induced neutrophil chemoattractant-1 (CINC 1, a counterpart of human GRO (melanoma growth-stimulating activity)) dependent chemotaxis in rat neutrophils. Sanguiin H-11 inhibited platelet-activating factor and N-formylmethionyl-leucyl-phenylalanine-dependent chemotaxis of neutrophil in addition to CINC-1-dependent reactions. Sanguiin H-11 also inhibited chemokinesis of neutrophil, but did not drastically affect the increase of intracellular free [Ca2+] level or degranulation monitored by the secretion of elastase, following chemoattractant stimulation. These results indicated that sanguiin H-11 is a potent inhibitor of chemoattractant-dependent and independent neutrophil movement. PMID- 10706388 TI - Clastogenicity of quinoline derivatives tested by micronucleus induction in vivo in the hepatocytes of partially hepatectomized mice. AB - The induction of micronucleated liver cells (MN-liver cell) was examined with halogenated and hydroxylated quinolines using partially hepatectomized mice. Among the chloroquinolines, 8-chloroquinoline demonstrated a significantly higher level of induction than the control. All the fluorinated derivatives examined, except for 6-fluoroquinoline, induced significantly higher levels, and there were no appreciable differences in MN-liver cell induction among the fluorinated quinolines, regardless of their mutagenic potencies in the Ames test. Of the hydroxylated quinolines examined, 2- and 4-isomers, which are not mutagenic, induced MN-liver cells to the same extent as a mutagenic isomer, 8 hydroxyquinoline. It seems that clastogenicity was not satisfactorily correlated with mutagenicity in the Ames test as far as this class of compounds is concerned. PMID- 10706389 TI - Antioxidant properties of N-(4-hydroxyphenyl)retinamide (fenretinide). AB - Fenretinide, N-(4-hydroxyphenyl)retinamide (4-HPR), is a cancer chemopreventive and antiproliferative agent whose mechanism of action is unknown. 4-HPR is a potent inducer of apoptosis in HL60 human leukemia cells which generates intracellular reactive oxygen species. The structural similarity of retinoic acid (RA), 4-HPR, and alpha-tocopherol (vitamin E) led us to investigate whether 4-HPR exhibits antioxidant activity. It was found that 4-HPR scavenged alpha,alpha diphenyl-beta-picrylhydrazyl (DPPH) radicals in a 1:1 ratio in contrast to vitamin E, where a 1:2 ratio relative to DPPH radicals was observed. In addition, linoleic acid peroxidation initiated by hydroxyl radicals was decreased by 4-HPR to the same extent as by vitamin E. Furthermore, lipid peroxidation in rat liver microsomes was reduced by 4-HPR to a greater extent than by vitamin E. Based on these results, 4-HPR appears to be an effective antioxidant that may have clinical utility for diseases treated with vitamin E. PMID- 10706390 TI - Post-translational modification of alphaB-crystallin of normal human lens. AB - The current study reports several post-translational modifications of alphaB crystallin in normal human lenses. The isoforms of post-translational modified alphaB-crystallin were isolated from the normal human lenses of >70-age group by ion exchange chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The isoform modifications were determined in detail by fast atom bombardment mass spectroscopy and amino acid sequence analysis. As the criterion of non-modified alphaB-crystallin, alphaB-crystallin from 1-d-old infant lenses were used. The modifications found in this study involve oxidation of the N terminal methionine-1 residue, phosphorylation of the serine-59 residue, and truncation of four amino acids from the C-terminal position of the crystallin. The oxidation of methionine-1 was found in the early stage of human life in 1-d old lens, although other modification of alphaB-crystallin were usually only found in old lenses (>70-age group). PMID- 10706391 TI - Molecular cloning and characterization of a cDNA for Glycyrrhiza glabra cycloartenol synthase. AB - A cDNA clone (GgCAS1) encoding cycloartenol synthase (CAS) has been isolated from Glycyrrhiza glabra (licorice) by cross-hybridization with that of Pisum sativum CAS as a probe. The deduced amino acid sequence of GgCAS1 exhibits 89%, 83% and 81% identity to those of Pisum sativum, Panax ginseng and Arabidopsis thaliana CASs, respectively. CAS activity has been detected in the homogenate of the yeast transformed with the expression vector containing the open reading frame of GgCAS1. Southern blot analysis suggested that at least two CAS genes exist in the licorice genome. In Northern blot analysis, the strong signal for CAS mRNA is detected in the cultured licorice cells of all growth phases, but no significant increase of CAS mRNA expression was observed in the cells treated with the 3 hydroxy-3-methylglutaryl-CoA reductase inhibitor, pravastatin. PMID- 10706392 TI - A computer program for pharmacokinetics based on maximum likelihood estimation using the gamma distribution with a probability density function: comparison with the normal distribution. AB - A computer program is described for maximum likelihood estimation within the gamma or normal distribution which can be used to estimate pharmacokinetic parameters. Pharmacokinetic analysis using this proposed program was investigated by the Monte Carlo method. The assumed pharmacokinetic models were a one compartment intravenous model and an oral model. The simulated drug concentrations were generated using a 10% S.D. based on the gamma or normal distribution. The gamma or normal distribution was adopted as the probability density function (p.d.f.) to estimate model parameters. The Powell method was used to maximize the logarithmic likelihood. There were no differences in the estimated parameters in terms of statistical and frequency distributions between the gamma and normal distributions using the generated data and the p.d.f. distributions. However, the number of failures to calculate the parameters using the p.d.f. with the normal distribution was more than five times that using the gamma distribution. This result suggests that it may be necessary to evaluate the validity of results computed using the maximum likelihood estimation based on a normal distribution as a data error distribution and p.d.f. PMID- 10706393 TI - Antifibrotic effects of the methanol extract of Polygonum aviculare in fibrotic rats induced by bile duct ligation and scission. AB - The antifibrotic effect of the methanol extract from Polygonum aviculare (PA), Artemisia capillaris (AC) and an aqueous solution of biphenyl dimethyl dicarboxylate (DDB) on liver fibrosis were studied. Liver fibrosis was induced by a bile duct ligation and scission (BDL/S) operation, duration of 4 weeks in rats. In BDL/S rats, the levels of aspartate transaminase (AST), alanine transaminase (ALT), alkaline phosphatase (ALP), total bilirubin in serum and hydroxyproline content in liver were dramatically increased. The PA treatment in BDL/S rats reduced the serum AST, ALT and ALP levels significantly (p<0.01). Liver hydroxyproline content in PA treated BDL/S group was also reduced to 40% that of BDL/S control group (p<0.01). The morphological characteristics of fibrotic liver, which appeared in BDL/S control group were improved in the PA treated BDL/S group. But neither AC nor DDB treatment improved any parameters in fibrotic rats induced by BDL/S. These results indicate that PA has an antifibrotic effect on fibrotic rats induced by BDL/S. PMID- 10706394 TI - Inhibitory effects of fidarestat on aldose reductase and aldehyde reductase activity evaluated by a new method using HPLC with post-column spectrophotometric detection. AB - A new method to assay the activity of aldose reductase (AR) and aldehyde reductase (AHR) by high-performance liquid chromatography is described. The separation of AR and AHR from tissue extracts using an anion-exchange column was followed by chromatographic measurement of the activity in the elute. AR and AHR activity were expressed as the area under the peak obtained by post-column spectrophotometric detection of the decrease of coenzyme (NADPH) in each enzyme reaction. In the enzyme preparation from rat or human tissues obtained by this method, two active peaks were identified as AR and AHR. The correlation coefficient between the injection volume of the enzyme preparation from each tissue and each peak area was 0.998 or greater. In addition, the within-day preservation rate of AR or AHR activity from each tissue was over 95%. In a comparative study of fidarestat with other AR inhibitors using this method, it was confirmed that the inhibitory effect of fidarestat on AR activity from each rat tissue was more potent than that produced by sorbinil and equipotent to that of epalrestat and zenarestat. Fidarestat was also found to inhibit AR activity more potently than AHR activity in human erythrocytes. Therefore, this method is applicable to studies of the selective inhibition of AR or AHR by test compounds. PMID- 10706395 TI - Failure in antitumor activity by overdose of an immunomodulating beta-glucan preparation, sonifilan. AB - Schizophyllan (SPG, Sonifilan) is a soluble (1-->3)-beta-D-glucan, used as a biological response modifier (BRM) with radiation therapy for cancer treatment in Japan. The mechanism of SPG mediated antitumor activity is thought to be via immune stimulation, which includes cytokine production, hematopoietic response, and so on. In this paper, we found that the activity of SPG was quite long-lived and an overdose significantly failed to display the antitumor activity. To demonstrate the mechanism several parameters were examined using a high dose of SPG administration as follows: i) the effect on vascular permeability in vivo, ii) the priming effect on tumor necrosis factor (TNF-alpha) production in vivo, iii) the effect on macrophage adherence to plastic plate in vitro, and iv) anti Sarcoma 180 antibody production in vivo. It was evident that vascular permeability and anti-Sarcoma 180 antibody production remained unchanged, but TNF alpha production and adherence to a plastic plate was significantly reduced by a high dose of SPG. These facts strongly suggested that modulation of the cytokine syntheses and the leukocyte traffic would be the causative mechanisms of the failure of antitumor activity by an overdose of SPG. PMID- 10706396 TI - Biological activity of alkyl 2-(acylthio)benzoates. AB - Of a series of synthetic alkyl 2-(acylthio)benzoate (1-20), all the derivatives except for n-butyl 2-butyrylthiobenzoate (18) and n-butyl 2-n-valerylthiobenzoate (20) showed clear phytogrowth-inhibitory activity. All the compounds tested except for methyl 2-butyrylthiobenzoate (3) exhibited cytotoxic activity on mouse splenic T cells. Strong phytogrowth-inhibition and cytotoxic activity were found with 1, 6, 11 and 16 with an acetylthio group at C-2, suggesting that the acetyl group seems to play an important role in both activities of alkyl 2 (acylthio)benzoates. Among them, methyl 2-acetylthiobenzoate (1) was the strongest inhibitor. On the other hand, potent inhibition of prolyl endopeptidase was exhibited by 2, 7, 12 and 17 with a propionylthio group at C-2. These findings imply that a propionyl group might be useful for increasing the inhibitory activity against on prolyl endopeptidase. PMID- 10706397 TI - Antimicrobial activities of some amino derivatives of 5,7-dibromo-2-methyl-8 hydroxyquinoline. AB - The bromine atoms of the title compound, 5,7-dibromo-2-methyl-8-hydroxyquinoline were replaced by the requisite amino compound to afford 6 amino derivatives viz: bis(diethylamino)-, bis(dibutylamino)-, bis(dicyclohexylamino)-, dipyrolidino-, dipiperidino- and dipiperazino derivatives. The antimicrobial activity of these compounds were investigated against selected gram positive (Staphylococcus aureus and Bacillus subtilis), gram negative bacteria (Escherichia coli and Pseudomonas aeruginosa) and yeast (Candida albicans). All the compounds showed significant activity against the test microorganisms, from 5-30 times compared to the title compound. It was observed that all derivatives were more effective against gram positive bacteria. No correlation has been established between the minimum inhibitory (MIC) concentrations of the derivatives and the structural modifications. PMID- 10706398 TI - Dependence of estrogenic activity on the shape of the 4-alkyl substituent in simple phenols. AB - The ability of certain chemicals to mimic the effects of natural steroid hormones and their potential to disrupt the delicate balance of the endocrine system in animals has attracted much interest in recent years. Alkylphenolic chemicals have been reported to be weakly estrogenic. Estrogen receptor (ER) binding is primarily the result of interaction of the receptor with both a phenolic residue, and a hydrophobic pharmacophore. We have prepared and screened various phenols having a bulky 4-alkyl group, which may interact hydrophobically with the receptor, for estrogenic activity by using a previously described reporter gene assay employing COS-1 cells transfected with rat ER alpha-expression plasmid and an appropriate reporter plasmid. Some of the tested compounds, such as 4-(1 adamantyl)phenol and 4-cycloalkylphenols, exhibited much more potent activity than the typical estrogenic alkylphenol, 4-tert-octylphenol. PMID- 10706399 TI - Cytotoxic alkaloids of Pachysandra terminalis. AB - Four steroidal alkaloids, epipachysamines B (1) and E (2), pachystermine A (3) and pachysamine E (4), were isolated as cytotoxic principles from the MeOH extract of the stems of Pachysandra terminalis SIEB. et ZUCC. (Buxaceae). These alkaloids showed cytotoxic activity against P388 and P388/ADR leukemia cells in vitro. Three of the alkaloids (1-3) were previously isolated from this plant material, and this is the first report of their cytotoxic activity. Pachysamine E (4) is a new alkaloid. PMID- 10706400 TI - Reductive metabolism of stilbene oxide and styrene oxide to the olefins in rats. AB - The present study provides the first evidence that stilbene oxide and styrene oxide are reductively metabolized to the corresponding olefins in rats. When cis- or trans-stilbene oxide was given orally to rats, both cis- and trans-stilbene were isolated from the urine and feces. Styrene was also isolated from the urine and feces of rats given styrene oxide. These metabolites were identified unequivocally by UV and mass spectral comparison with authentic samples, and on the basis of their TLC and HPLC behavior. However, these olefins were not detected in the urine or feces of antibiotics-treated rats dosed with cis- or trans-stilbene oxide. Cecal contents of the untreated rats exhibited olefin oxide reductase activities toward cis- and trans-stilbene oxides under anaerobic conditions. The results suggest that intestinal bacteria play an important role in the reduction of olefin oxides to the corresponding olefins in the animal body. PMID- 10706401 TI - Effect of inhibitors of glycoprotein processing on cytokine secretion and production in anti CD3-stimulated T cells. AB - We have investigated the effect of inhibitors of glycoprotein processing on cytokine secretion and production in anti CD3-stimulated T cells to elucidate the role of carbohydrate in the triggering of T cell function. The inhibitors of glycoprotein processing, especially mannnosidase inhibitors, enhanced the anti CD3-induced production of interleukin-2 (IL-2), which is a cytokine without the linkage sequence of N-linked oligosaccharides. On the other hand, N-methyl-1 deoxynojirimycin (NMdNM, an inhibitor of processing glucosidase 1), 1 deoxynojirimycin (dNM, an inhibitor of processing glucosidase I and II) and bromoconduritol (BCD, an inhibitor of processing glucosidase II) inhibited the secretion of interleukin-4 (IL-4), interferon-gamma (IFN-gamma), or interleukin-5 (IL-5) into culture supernatants of anti CD3-stimulated T cells, which had N linked oligosaccharides. Mannosidase inhibitors, 1-deoxymannojirimycin (dMAN, an inhibitor of processing mannosidase I) and swainsonine (SWN, an inhibitor of processing mannosidase II) did not inhibit the secretion or production of IL-4, IFN-gamma and IL-5. To confirm the inhibition of N-linked oligosaccharide processing in the cytokines by the above inhibitors, the binding of IFN-gamma to lectins with various sugar-binding specificities was investigated. All inhibitors reduced the binding of IFN-gamma to PHA E4, which had a high affinity to bi- or tri-antennary complex type N-linked oligosaccharides with bisecting N acetylglucosamine. Similarly, all inhibitors reduced the binding of IFN-gamma to PHA L4, which had high affinity to tri- or tetra-antennary complex type N-linked oligosaccharides with beta1-6-linked branching. SWN and dMAN increased the binding of IFN-gamma to concanavalin A (ConA), which had a high affinity to bi antennary complex type, hybrid type and high-mannose type N-linked oligosaccharides. These results suggest that the processing inhibitors used here inhibit the N-linked oligosaccharide processing of cytokines, and the inhibition of processing enzyme glucosidases I and II induces a decreased secretion of cytokines with N4-linked oligosaccharides. PMID- 10706402 TI - Hasty effect on the metabolism of glycyrrhizin by Eubacterium sp. GLH with Ruminococcus sp. PO1-3 and Clostridium innocuum ES24-06 of human intestinal bacteria. AB - Eubacterium sp. GLH with Ruminococcus sp. PO1-3 and Clostridium innocuum ES24-06 possessing enzymes involved in the metabolism of glycyrrhizin (GL) was cultured in GAM medium with and without 1.0 mM GL or 1.0 mM glycyrrhetic acid (GA). GL (1.0 mM) enhanced 3alpha-hydroxyglycyrrhetinate (3alpha-hydroxyGA) dehydrogenase activity, GA (1.0 mm) suppressed 3alpha-hydroxyGA dehydrogenase activity, GL beta D-glucuronidase activity and the mixed bacterial growth, and GL and GA showed almost no change in a lower level of 3beta-hydroxysteroid dehydrogenase (3beta HSD) activity during 5 d of culture. GL (1.0 mM) and GA (1.0 mM) were metabolized to a small amount of GA and a negligible amount of 3-oxo-glycyrrhetic acid (3-oxo GA) and 3alpha-hydroxyGA, and to a negligible amount of 3-oxo-GA, respectively, by these mixed bacteria. These amounts coincided with those of metabolites produced from 1.0 mM GL and 1.0 mM GA added to these mixed bacteria after 24 h culture. Whole bacteria and sonicated bacteria derived from the collection of these mixed bacteria reached a maximal stage and metabolized GL to a relatively large amount of GA and 3-oxo-GA, and a negligible amount of 3alpha-hydroxyGA and GA to a small amount of 3-oxo-GA and 3alpha-hydroxyGA within 180 min. GL beta-D glucuronidase with 3beta-HSD and 3alpha-hydroxyGA dehydrogenase partially purified from each bacterium was converted GL to 3alpha-hydroxyGA, producing metabolites of about 60% after 10 min of incubation. These mixed bacteria possessed high enzyme activities could produce the metabolites of GL in under one hour under conditions. PMID- 10706403 TI - Different recognition by peroxisome proliferator structures in rat peroxisomal induction: application of sandwich ELISA using monoclonal antibody against rat peroxisomes. AB - A novel assay for a peroxisomal beta-oxidation enzyme by sandwich ELISA using a monoclonal antibody (RPX-5) against purified rat liver peroxisomes was developed. Immunoblot analysis revealed that RPX-5 recognized a 78 Kd protein, which is a peroxisomal bifunctional enzyme (PBE) in the beta-oxidation pathway. Immunoprecipitation by RPX-5 and the resulting reduction of PBE activity were dependent on RPX-5 concentrations. Sandwich ELISA using RPX-5 could be used to assay PBE in the range of 30 to 2000 ng protein/ml. In rat hepatocyte cultures, the PBE amount by this assay correlated well with PBE activity, with correlation coefficients of 0.965. Studying the mechanisms of peroxisomal induction, patterns of peroxisomal induction were examined by co-treatment of rat hepatocytes with various peroxisome proliferators (PxPs). Treatment with clofibrate and bezafibrate resulted in neither an additive nor synergistic effect on PBE level. On the other hand, co-treatment with either bezafibrate-Wy-14,643 or clofibrate MEHP(mono(2-ethylhexyl)phthalate) both resulted in an additive effect. From these results, it is suggested that PxPs of the fibrate group may exert their functions via a common process, and non-fibrate PxPs via a different process in hepatocytes. The cognition site for peroxisome proliferators, therefore, might not involve a single site for inducing peroxisomal enzymes. PMID- 10706404 TI - Characteristics of post-tetanic contraction induced by naloxone in guinea pig ileum. AB - The characteristics of isolated guinea-pig ileal contractions of basal tension after tetanic stimulation in the presence of a high concentration of naloxone (NLX) [post-tetanic contraction] were investigated. The post-tetanic contraction did not occur in the absence of NLX, but did occur in a concentration-dependent manner in the presence of a high concentration of NLX (5 x 10(-7), 10(-6), 10(-5) and 5 x 10(-5) M), the concentration of which was higher than that required for antagonizing post-tetanic twitch inhibition. The contraction in the presence of 10(-6) M NLX was diminished by washing NLX from the preparation with Krebs bicarbonate solution. The contraction under 10(-6) M NLX occurred in a frequency dependent manner (5, 10 and 20 Hz), but not at 0.1 Hz. Tetanic stimulation (5, 10 and 20 Hz) without NLX did not induce this contraction. The post-tetanic contraction with 10(-6) M NLX had a tendency to be antagonized in the presence of 5 x 10(-6) M atropine. Methysergide (5 x 10(-5) M) had no effect on this contraction. Spantide (10(-5) M) largely inhibited the contraction, and indomethacin (5 x 10(-6) M) and tetrodotoxin (5 x 10(-7) M) completely inhibited this contraction. These results indicate that tetanic stimulation in the presence of a high concentration of NLX induces contraction of the ileal muscle due to the release of endogenous ileal contractile substances (substance P, prostaglandins and acetylcholine), and suggests that these contractions are closely linked to the endogenous opioid system induced by tetanic stimulation in the ileum. PMID- 10706405 TI - Antidiabetic activity of white skinned sweet potato (Ipomoea batatas L.) in obese Zucker fatty rats. AB - Antidiabetic effects of white skinned sweet potato (Ipomoea batatas L.) (WSSP) and troglitazone, an insulin sensitizer, were investigated. Hyperinsulinemia in Zucker fatty rats was reduced by 23%, 26%, 60% and 50%, respectively, 3, 4, 6 and 8 weeks after starting the oral administration of WSSP. Similar results were obtained with troglitazone. In the glucose tolerance test after 7 weeks of treatment, increases in blood glucose levels after glucose loading were inhibited by the administration of WSSP. Glucose tolerance was also improved. Blood triacylglyceride (TG) and free fatty acid (FFA) lactate levels were lowered by the oral administration of WSSP. Similar effects on blood insulin, lipid and lactate levels were observed after the administration of troglitazone. Body weight gain increased in the troglitazone group, but not in the WSSP group, compared to the control group. In histological examinations of the pancreas of Zucker fatty rats, remarkable regranulation of pancreatic islet B-cells was observed in the WSSP and troglitazone groups after 8 weeks of treatment. These results suggest that WSSP shows remarkable antidiabetic activity and improves the abnormality of glucose and lipid metabolism by reducing insulin resistance. PMID- 10706406 TI - Biochemical characterization of 60S acidic ribosomal P proteins from porcine liver and the inhibition of their immunocomplex formation with sera from systemic lupus erythematosus (SLE) patients by glycyrrhizin in vitro. AB - The three casein kinase II (CK-II) phosphate acceptors (p35, p17 and p15) in the Superdex CK-II fraction prepared from a 1.5 M NaCl extract of porcine liver were selectively purified by glycyrrhizin (GL)-affinity column chromatography (HPLC) as a heterocomplex associated with CK-II. Determination of the N-terminal amino acid sequences and immunological tests confirmed that these three CK-II phosphate acceptors belong to the family of 60S acidic ribosomal proteins (P0, P1 and P2). Three polyphenol-containing anti-oxidant compounds [catechin, epigallocatechin gallate (EGCG) and quercetin] inhibited CK-II activity (phosphorylation of these ribosomal P proteins) in a dose-dependent manner in vitro. Quercetin (ID50 = approx. 50 nM) was found to be an effective CK-II inhibitor. In contrast, CK-II activity was significantly stimulated by lower doses (0.3-3 microl) of GL, but was inhibited at high doses above 30 microM. As expected, GL at high doses above 200 microM inhibited the immunocomplex formation of 60S acidic ribosomal P proteins with their specific antibodies in the sera from patients with systemic lupus erythematosus (SLE). These results suggest that (i) a GL-affinity column is useful for effective purification of 60S acidic ribosomal P proteins from various mammalian cells as a heterocomplex associated with CK-II; and (ii) a relative high dose of GL may prevent the immunocomplex formation of 60S acidic ribosomal P proteins with their specific antibodies in the sera of SLE patients. PMID- 10706407 TI - Gender differences in the antidiarrheal effect of zaldaride maleate in rats. AB - The amelioration of secretory diarrhea has been reported after the administration of zaldaride maleate (ZAL), a selective calmodulin inhibitor, to male rodents. In this study, the antidiarrheal effect of ZAL in female rats was compared with that in male rats. In female and male rats, ZAL significantly ameliorated 16,16 dimethyl prostaglandin E2-induced diarrhea at doses of 1 and 3 mg/kg (p.o.), respectively, with ID50 values of 0.7 mg/kg (p.o.) in the females and 10.3 mg/kg (p.o.) in males. In castor oil-induced diarrhea, ZAL also significantly reduced the incidence of diarrhea in female and male rats at doses of 10 and 30 mg/kg (p.o.), respectively. When the same dose of ZAL was given orally to female and male rats, the maximum plasma level of this compound was approximately 3 times higher in female rats than in male rats. In contrast, after intravenous administration of the same dose of ZAL to female and male rats, the total clearance of this compound was similar. In an Ussing chamber experiment, the inhibitory action of ZAL on vasoactive intestinal polypeptide-induced ion secretion in the colon showed no difference between female and male rats. In conclusion, the antidiarrheal effect of ZAL in female rats is more potent than that in males, and could be due to the difference in plasma levels of this compound between female and male rats after oral administration. PMID- 10706408 TI - Characterization of hydrogen peroxide-induced apoptosis in mouse primary cultured hepatocytes. AB - The influence of oxidative stress by hydrogen peroxide (H2O2) was examined in mouse primary cultured hepatocytes. A change in morphology was observed in hepatocytes incubated for 30 min in saline A containing H2O2. The percentage of dead cells, as measured by the fluorescence method, was increased in a dose dependent manner. In addition, a ladder-like DNA fragmentation pattern was detected by agarose gel electrophoresis 1 h after exposure to 3 mM H2O2. This phenomenon was prolonged for 24 h. Hydrogen peroxide-induced cell viability reduction and DNA fragmentation were dose-dependently protected by the addition of antioxidants (N-acetylcysteine, L-ascorbic acid), a metal-chelator (1,10 phenanthroline), iron-chelator (deferoxamine) and intracellular calcium ion chelator (quin 2-AM). No influence, however, was detected by endonuclease inhibitors (zinc, aurintricarboxylic acid) and poly (ADP-ribose) polymerase inhibitors (3-aminobenzamide, theophylline). These results following H2O2-induced cell viability reduction suggested that oxidative stress by H2O2 itself or H2O2 derived changes involved in ferrous or intracellular calcium ions resulted in apoptosis in mouse primary cultured hepatocytes. These phenomena are not likely to be associated with endonuclease or poly (ADP-ribose) polymerase. PMID- 10706409 TI - Characterization of microsomal alcohol oxygenase catalyzing the oxidation of 7 hydroxy-delta 8-tetrahydrocannabinol to 7-oxo-delta 8-tetrahydrocannabinol in rat liver. AB - The formation of 7-oxo-delta8-tetrahydrocannabinol (7-oxo-delta8-THC) from 7beta hydroxy-delta8-THC was found in hepatic microsomes of rats. The activity was stereoselective and about 3-fold higher than that from 7alpha-hydroxy-delta8-THC. The oxidative activity of 7alpha- and 7beta-hydroxy-delta8-THC to 7-oxo-delta8 THC was significantly higher in male than in female, and significantly enhanced by both dexamethasone and phenobarbital, and then inhibited up to about 20% of the control value by antibody against P450GPF-B, presumably a member of the 3A subfamily, a major enzyme responsible for the formation of 7-oxo-delta8-THC in guinea pigs. This antibody also inhibited the formation of 7alpha- and 7beta hydroxy-delta8-THC, and 7-oxo-delta8-THC from delta8-THC by hepatic microsomes of rats. These results indicate that there is a sex-related difference in the oxidation of 7-hydroxy-delta8-THC to 7-oxo-delta8-THC and the reaction is mainly catalyzed by P450 enzyme(s) belonging to the 3A subfamily as major enzyme(s) of microsomal alcohol oxygenase in rats. PMID- 10706410 TI - Inhibition of prostaglandin E2 and leukotriene C4 in mouse peritoneal macrophages and thromboxane B2 production in human platelets by flavonoids from Stachys chrysantha and Stachys candida. AB - Seven flavonoids of Stachys chrysantha and Stachys candida have been isolated. The structures of the compounds were elucidated by spectroscopic methods, particularly highfield NMR spectroscopy. The effects of the methanol extracts of these two endemic Greek Stachys sp. and their main flavonoids were examined on arachidonic acid (AA) metabolism in the cellular system (mouse peritoneal macrophages and human platelets). Their cytotoxicity on cells was also investigated. Most samples assayed did not exhibit any significant effect on prostaglandin E2 (PGE2)-release from calcium ionophore-stimulated mouse peritoneal macrophages. Only chrysoeriol-7-O-beta-(3''-E-p-coumaroyl) glucopyranoside, at the highest non-cytotoxic dose (50 microM), inhibited the release of PGE2, but this effect is not statistically significant. The release of leukotriene C4 (LTC4) by mouse peritoneal macrophages stimulated with calcium ionophore was inhibited by a crude extract of S. chrysantha, with an IC50 value of 34.3 microg/ml. Xanthomicrol (IC50 = 29.2 microM) and chrysoeriol-7-O-beta-D (3''-E-p-coumaroyl)-glucopyranoside (IC50 = 11.1 microM) also inhibited the release of LTC4, although it showed less potency than the reference compound nordihydroguaiaretic acid (NDGA) (IC50 = 2 microM). However, most samples assayed showed a significant effect on thromboxane B2 (TXB2)-release from calcium ionophore-stimulated human platelets, with inhibition percentages slightly lower than the reference drug ibuprofen (IC50 = 7 microM). The IC50 values are: crude extract of S. candida 23.3 microg/ml; crude extract of S. chrysantha 23.1 microg/ml; xanthomicrol 28.8 microM; calcycopterin 2.66 microM and chrysoeriol-7 O-beta-D-(3''-E-p-coumaroyl)-glucopyranoside 8.8 microM. Our results indicate that the selective inhibition of TX-synthase enzyme may be the primary target of action of most of these samples, and one of the mechanisms through which thus exert their antiinflammatory effects. PMID- 10706411 TI - The promoting effect of eucommiol from Eucommiae cortex on collagen synthesis. AB - The effect of a methanol extract of Eucommiae Cortex on collagen synthesis was investigated in false aged model rats. Granuloma formation and collagen synthesis were significantly increased by the administration of the methanol extract of Eucommiae Cortex. The effective component of Eucommiae Cortex was then discussed by fractionating the methanol extract of Eucommiae Cortex. Eucommiol, a main component in the water fraction of the methanol extract, was found to be an effective compound. In our previous paper, we reported the promoting effect of Eucommia ulmoides OLIVER leaf on collagen synthesis, and found geniposidic acid and aucubin were the main effective compounds in the leaf. Based on our data in this paper, we clarified that the main effective components of the Eucommia ulmoides OLIVER leaf and Eucommiae Cortex were different. Geniposidic acid and aucubin were reported to be contained at a high concentration in the fresh cortex of Eucommia ulmoides OLIVER, but during the drying process and storage, most of them were destroyed by enzymes in the cortex and very little remained in the Eucommiae Cortex. Therefore, we investigated the effect of the methanol extract of fresh cortex of Eucommia ulmoides OLIVER. A stronger effect than Eucommiae cortex was shown, and geniposidic acid, aucubin and geniposide were concluded to be the main effective components. Although geniposide was found to be an effective compound, when the dose was higher than 50 mg/kg/d, toxicity was shown. The pharmaceutical effect of eucommiol was reported for the first time. PMID- 10706412 TI - Granuloma maturation in the rat is advanced by the oral administration of Eucommia ulmoides OLIVER leaf. AB - We have reported that collagen metabolism was improved by the administration of Eucommia ulmoides OLIVER leaf. In this paper, we examine the granuloma maturation and deposition of collagen in the granuloma of rats due to the oral administration of this leaf. After 3 weeks of the oral administration, granuloma formation was induced by the formalin soaked filter paper-pellet method. A week later, the developing granuloma was dissected. Granuloma formation was significantly increased due to ingestion of the dried leaf at a dose of 1.8 g/kg of body weight/d. The collagen content in the granuloma was also significantly increased. In the case of the collagen profile, the pepsin-solubilized collagen content and its relative percentage to the total collagen were significantly higher than in the control. Histochemical examination showed that the granuloma tissues were well developed, and displayed many newly synthesized capillary vessels and a greater quantity of fibroblasts and monocytes in the 1.8 g leaf group. High density lipoprotein (HDL)-cholesterol and triglyceride content in the blood plasma were significantly higher than in the control. These results show that granuloma maturation was accelerated and the energy was supplied from fatty acid metabolism. The administration of Eucommia ulmoides OLIVER leaf may be effective at speeding up the wound healing process. PMID- 10706413 TI - In vitro biological activities of a series of 2 beta-substituted analogues of 1 alpha,25-dihydroxyvitamin D3. AB - Biological activities of a series of 2beta-substituted analogues of 1alpha,25 dihydroxyvitamin D3 [1alpha,25(OH)2D3] were evaluated in vitro in terms of their binding affinity with regard to calf thymus cytosolic vitamin D receptor (VDR) and rat plasma vitamin D-binding protein (DBP). Additionally, reporter gene luciferase activities using either a rat 25-hydroxyvitamin D3-24-hydroxylase gene promoter, including two vitamin D-responsive elements (VDREs), in transfected rat osteoblast-like ROS17/2.8 cells, or a human VDR-GAL4 modified two-hybrid system in transfected human epitheloid carcinoma, cervix HeLa cells were examined. Binding affinity for VDR, transactivation potency on the target gene and VDR mediated gene regulation of the hydroxyalkyl and hydroxyalkoxy 2beta-substituted analogues were almost comparable to those of 1alpha,25(OH)2D3, while the alkyl and alkenyl analogues were much less active than 1alpha,25(OH)2D3. This study investigated the biological evaluation of a series of 2beta-substituted analogues at the molecular level, with regard to the structural differences of alkyl, alkenyl, hydroxyalkyl, hydroxyalkoxy, alkoxy, hydroxy and chloro substituents at the 2beta-position of 1alpha,25(OH)2D3. PMID- 10706414 TI - Survey of solasodine-type glycoalkaloids by western blotting and ELISA using anti solamargine monoclonal antibody. AB - Solasodine glycosides were analyzed for the selection of higher-producing natural resources in the fruits of Solanum spp. by western blotting and ELISA using anti solamargine monoclonal antibody (MAb). Western blotting of Solanum khasianum showed a correlation with the analysis by ELISA. The results indicated that the fruits of S. khasianum contained the highest level of solasodine glycosides, compared to other species having 53.43 +/- 3.90 microg/mg dry wt. The high level of solasodine glycosides in S. khasianum suggests that it may be suitable for commercial production. PMID- 10706415 TI - Pharmacological properties of traditional medicines. XXVI. Effects of Sansohnin to on pentobarbital sleep in stressed mice. AB - We investigated the effect of Sansohnin-to (SAT) on changes of duration in sodium pentobarbital (PB)-induced sleeping time caused by five types of stress. SAT reversed shortened PB sleep in repeated cold stress or 45 min-restraint stress tests and the prolonged PB sleep in 120 min-restraint stress. SAT did not reverse the shortened PB sleep in the specific stress state caused by an alternating rhythm in temperature stress or social isolation stress. In addition, SAT influenced both shortened PB sleep in 45 min-restraint stress and prolonged PB sleep in 120 min-restraint stress. SAT had no effect on PB sleep in unstressed control mice. These findings suggest that SAT has unusual activity, different from synthetic narcoleptics such as benzodiazepine. This is because SAT had no effect on PB sleep in unstressed mice, and it reverses stress-induced decrease and/or increase in PB sleep by improving stress-induced functional changes in the central nervous system, rather than by acting like a synthetic hypnotic on the gamma-aminobutyric acidA (GABA(A)) receptor. PMID- 10706416 TI - Effect of tranilast oily gel on carrageenin-induced granulation in rats. AB - Tranilast (TL) oily gels consisting of hydrogenated soybean phospholipid and fatty-acid ester were prepared, and the inhibitory effect of the gels on the growth of granulation tissue were evaluated in a carrageenin-induced rat granulation model. By the application of 0.1 and 0.2% TL oily gel, the weight of granulation tissue was significantly reduced to 64 and 55%, respectively, of control value. Furthermore, these gels reduced their respective hydroxyproline content to 64 and 51% of the control. On the other hand, the inhibitory effect of 10% TL ointments, which are clinically used for the treatment of keloids and hypertrophic scars as hospital preparations, was much lower than that of the oily gels. In addition, the application of 0.1 and 0.2% oily gel led to high concentration (0.1% gel, 168+/-18 microg/g; 0.2% gel, 221+/-16 microg/g) of TL in the dermis as compared with the 10% TL ointments. These results suggest that TL oily gels may be a useful topical formulation for the treatment of keloids and hypertrophic scars. PMID- 10706417 TI - The effects of absorption enhancers on the pulmonary absorption of recombinant human granulocyte colony-stimulating factor (rhG-CSF) in rats. AB - Pulmonary absorption of recombinant human granulocyte colony-stimulating factor (rhG-CSF) with various surfactants and protease inhibitors were examined in rats. The relative bioavailabilities of rhG-CSF with surfactants, such as polyoxyethylene 9-lauryl ether (Laureth-9) and sodium glycocholate (SGC), after intratracheal (i.t.) administration by intravenous (i.v.) and subcutaneous (s.c.) means were 37% (i.v.), 88% (s.c.), 84% (i.v.) and 197% (s.c.), respectively. These values were evaluated from the ratio of the area under the curve (AUC) of the plasma rhG-CSF concentration versus time for 8 h. In the presence of various kinds of protease inhibitors, such as (p-amidinophenyl) methanesulfonyl fluoride x HCl (p-APMSF), aprotinin and bestatin, an increase in the plasma rhG-CSF concentration was observed, and the effect with p-APMSF was maximal. The relative bioavailabilities of rhG-CSF with p-APMSF after i.t. administration by i.v. and s.c. means were increased about 2-fold. To clarify the absorption mechanism of rhG-CSF, rhG-CSF was intratracheally administered with both Laureth-9 and p APMSF. The AUC of rhG-CSF increased with both agents, and was approximately equal to that with SGC, which has both an enhancing effect on membrane permeation and an inhibitory effect on enzymatic degradation after i.t. administration. Consequently, it was considered that permeation and enzymatic degradation were rate-determining steps in the pulmonary absorption of rhG-CSF after i.t. administration. PMID- 10706418 TI - Bioavailability assessment of arginine-vasopressin (AVP) using pharmacokinetic pharmacodynamic (PK-PD) modeling in the rat. AB - A novel method of assessing the extent of oral bioavailability of arginine vasopressin (AVP) from pharmacological data was presented. After intravascular administration (i.v. bolus or short-term infusion) of AVP to rats, the relationship between blood concentrations and its effect on both mean arterial pressure (hemodynamic effect) and urinary sodium concentration (anti-diuretic effect) was described on the basis of an integrated pharmacokinetic pharmacodynamic (PK-PD) model. A direct model was used for the hemodynamic response, while an indirect response model, rather than a hypothetical link model was used for the anti-diuretic response. A sigmoid Emax model was applied to describe the drug-receptor interaction. Pharmacological responses after intravascular administration of AVP were reasonably described by the PK-PD model. However, PD parameters estimated by the PK-PD analysis suggested that apparent receptor affinity rather than efficacy in i.v. bolus study was significantly higher than that in the short-term infusion study. This fact indicated that PK-PD relationship was influenced by the intravascular input rate of AVP. We then investigated the relationship between plasma concentration and amount of AVP bound to the V2 receptors in the kidney. The result indicated that the amount of AVP bound to the receptors after i.v. bolus injection was always greater than that after short-term infusion. Since the PK-PD relationship after oral administration was almost identical with that after short-term infusion, the PK PD model obtained in the short-term infusion study was used to assess the extent of oral bioavailability (EBAPp.o.). The EBAp.o. values, estimated from pharmacological effects (hemodynamic effect and anti-diuretic effect) after oral administration of 5 microg/kg of AVP were 0.68% to 0.93% and were almost identical with the actual EBAPp.o. value (0.81%). From these results, we concluded that oral bioavailability of AVP was reasonably predicted by the PK-PD model, provided that appropriate pharmacological effects and appropriate intravascular dosing rate as a reference formulation are available. The method may be an alternative to methods based on plasma concentrations, when drug concentration cannot be measured and when appropriate pharmacological data are available. PMID- 10706419 TI - Magnetic resonance lymphography of profundus lymph nodes with liposomal gadolinium-diethylenetriamine pentaacetic acid. AB - Lymphography, especially imaging of profundus lymph nodes, is a useful tool for diagnosis of cancer metastases in lymph nodes. However, positive enhancement agents for magnetic resonance lymphography (MRL) have not been available, since the positive imaging agents so far introduced are low-molecular-weight materials that are not trapped in lymph nodes. For the purpose of improved positive enhanced MRL, we employed liposomes as carriers of a positive enhancer, gadolinium-diethylenetriamine pentaacetic acid (Gd-DTPA). Magnetic resonance (MR) imaging was performed after subcutaneous injection of Gd-liposomes into the hind feet of rabbits which had reactive enlarged retroperitoneal lymph nodes. As a result, not only popliteal but also profundus retroperitoneal lymph nodes were positively enhanced by Gd-liposomes, especially after 20 min massage of the injected sites. Gd-Liposomes containing dipalmitoylphosphatidylglycerol were more effective than Gd-liposomes containing palmityl-D-glucuronide, a type of long circulating liposomes, suggesting that liposomal accumulation in lymph node is, at least partly, mediated by the trapping of liposomes by macrophages. These data show that liposomes modified with Gd-DTPA are effective for positive enhancement of both regional and profundus lymph nodes in MR lymphography. PMID- 10706420 TI - Free 4-hydroxyproline content in serum of bedridden aged people is elevated due to fracture. AB - Hydroxyproline (Hyp) content in biological fluids is used as a parameter of collagen catabolism, especially bone resorption. In this paper, we examined the relationship between aging and serum free 4-Hyp and proline (Pro) content using a newly introduced analytical technique which is chemiluminescence determination with electrogenerated tris (2,2'-bipyridine) ruthenium(III). Serum was collected from 50 bedridden aged people and 131 normal subjects living in Yamato-son, Amamioshima, Kagoshima. In the free 4-Hyp content of normal serum, the change in females is more gentle than that males with aging. The free 4-Hyp content of bedridden aged serum is significantly (p<0.05) elevated compared to normal aged serum. Bedridden aged females and males also show levels significantly (p<0.05) elevated in comparison to normal. These changes are due to bone resorption based on bedridden patients' atrophy. In bedridden aged people, free 4-Hyp of those with a fracture was significantly (p<0.05) elevated than in people without a fracture. No relationship between low bone density and high free 4-Hyp in serum was observed. It seems that bone resorption or bedridden atrophy may be enhanced in bedridden aged people by a fracture. In free Pro content in serum, no significant change was observed. These results show that the free 4-Hyp content in serum is useful in the clinical or biological inspection of bone resorption, especially in aged people. PMID- 10706421 TI - Effects of glycyrrhizin and glycyrrhetic acid on the growth, glycyrrhizin beta-D glucuronidase and 3 beta-hydroxysteroid dehydrogenase of human intestinal bacteria. AB - The peak of glycyrrhizin (GL) beta-D-glucuronidase activity for Ruminococcus sp. PO1-3 and Eubacterium sp. GLH changed to 24 h from 12 h of culture and to 12 h from 48 h, respectively, at almost the same level by the addition of 1.0 mM GL. This enzyme activity was about 20-fold higher in Eubacterium sp. GLH than in Ruminococcus sp. PO1-3. GL beta-D-glucuronidase activity of Ruminococcus sp. PO1 3 with Eubacterium sp. GLH and the intestinal flora showed a maximal peak at 12 h of culture in the presence and absence of 1.0 mM GL. This enzyme activity was about 2.5-fold higher in mixed bacteria than in intestinal flora. 3Beta hydrosteroid dehydrogenase activity of Ruminococcus sp. PO1-3 and Ruminococcus sp. PO1-3 with Eubacterium sp. GLH was suppressed greater in the presence of GL than without GL. Also, Ruminococcus sp. PO1-3, Eubacterium sp. GLH, and a mixture of both and intestinal flora, metabolized 1.0 mM GL to glycyrrhetic acid (GA) in yields of about 10, 70, 40 and 100%, respectively, with 24 h culture. From the level of GL beta-D-glucuronidase activity, it is considered that the metabolism of GL by intestinal flora is due to both enzymatic and non-enzymatic reactions. Moreover, GA at a concentration of 1.0 mM suppressed growth of Ruminococcus sp. PO1-3, Eubacterium sp. GLH, and the mixture of both and intestinal flora, which metabolized 1.0 mM GA to a negligible amount of 3-oxo-glycyrrhetic acid, indicating the accumulation of unchanged GA. GL beta-D-glucuronidase activity of intestinal flora was enhanced by GA, which stimulated bacteria possessing particular this characteristic. PMID- 10706422 TI - Biosynthesis of thiamin under anaerobic conditions in Saccharomyces cerevisiae. AB - We studied the biosynthetic route of thiamin in Saccharomyces cerevisiae to see whether the route differed under aerobic and anaerobic conditions. Histidine and pyridoxine are the precursors of the pyrimidine moiety of thiamin under aerobic conditions. Formate is incorporated into the pyrimidine via histidine. The incorporation of [13C]formate and [5'-(2)H2]pyridoxine into the pyrimidine was examined under anaerobic conditions. The labels from [13C]formate and [5' (2)H2]pyridoxine were not incorporated into the pyrimidine under anaerobic conditions, indicating that the biosynthetic pathway of the pyrimidine differed from that under aerobic conditions. On the other hand, [15N]glycine was incorporated into the thiazole under both anaerobic and aerobic conditions. The biosynthetic pathway of the thiazole was therefore unaltered by the O2 concentration. PMID- 10706423 TI - Reversal of acquired resistance to doxorubicin in K562 human leukemia cells by astemizole. AB - This study demonstrates that astemizole, a non-sedating anti-histaminergic drug with low toxicity in vivo, greatly potentiates the growth-inhibitory activity of doxorubicin in doxorubicin-resistant human leukemia cells (K562/DXR). Astemizole synergistically potentiated the cytotoxicity of doxorubicin for K562/DXR cells at a concentration of 0.1-3 microM in a dose-dependent manner, whereas they showed hardly any synthergistic effect in the parental cell line (K562) at the same concentration. Since doxorubicin resistance in these cells is associated with the expression of high levels of P-glycoprotein, we evaluated the effect of astemizole on P-glycoprotein activity in cytofluorographic efflux experiments with doxorubicin. Our results indicate that astemizole inhibits the P glycoprotein pump-efflux activity in a dose-related manner. Moreover, it also inhibits the photolabeling of P-glycoprotein by [3H]azidopine in a dose-dependent manner. These findings provide a biological basis for the potential therapeutic application of astemizole as an anticancer drug either alone or in combination with doxorubicin to multidrug-resistant leukemic cells. PMID- 10706424 TI - Role of endogenous endotoxin on tumor necrosis factor-hypersensitivity caused by D-galactosamine challenge. AB - We examined the role of endotoxin in the mechanism of recombinant human tumor necrosis factor (rhTNF)-hypersensitivity caused by D-galactosamine (GalN). We used polymyxin B, an antibiotic with anti-endotoxin activity, to determine the participation of endogenous endotoxin. The glycogen and blood glucose level of rhTNF (1 x 10(4) units/mouse, i.v.)-injected mice was lower at 7 h post intoxication than that in the control. Administration of rhTNF to GalN (700 mg/kg, i.p.)-treated mice resulted in lower levels of glycogen and blood glucose than those in animals treated with rhTNF alone. In mice pretreated with polymxin B (20 mg/kg, i.p.), the level at 7 h after rhTNF/GalN-injection was markedly increased compared to that in mice treated with rhTNF/GalN alone. The injection of a low endotoxin dose (0.1 mg/kg, i.p.) markedly decreased the rectal temperature in mice treated with rhTNF (5 x 10(3) units/mouse, i.v.) and GalN, and none of these animals survived after treatment for 18 h. These findings suggest that endogenously produced endotoxin may contribute to the extent of rhTNF-hypersensitivity caused by GalN. PMID- 10706425 TI - Effects of the ethyl and benzyl ethers of indenestrols A and B on cytotoxicity in Chinese hamster V79 cells. AB - We recently investigated the relationship between the structures of various indenestrols and their cytotoxicity, and reported that indenestrol A (IA), a metabolite of the synthetic nonsteroidal estrogen diethylstilbestrol, and indenestrol B (IB), an analog of IA, disrupt the microtubule architecture of Chinese hamster V79 cells in vitro. We then synthesized 16 optically active indenestrol derivatives by substituting monoethyl, monobenzyl and diethyl ether groups at the 6- and/or 4'-hydroxyl positions, and examined their cytotoxic activities in Chinese hamster V79 cells. The results indicated that the monoethyl ethers had cytotoxic activities similar to monomethyl ethers. However, the (+)- and (-)-monobenzyl ethers were less cytotoxic than the corresponding monomethyl and monoethyl derivatives. PMID- 10706426 TI - Metabolites of orally administered Perilla frutescens extract in rats and humans. AB - As a part of our search for bioactive substances from the leaves of Perilla frutescens BRITTON var. acuta KUDO (Perillae Herba, Labiatae), the aqueous extract was orally administered to rats and humans, and metabolites in the urine, plasma, and/or bile were analyzed by a high-performance liquid chromatograph (HPLC) equipped with a photodiode array detector. When the extract was administered to rats, 10 metabolites, trans-caffeic acid-4-O-sulfate (1), trans-p coumaric acid-4-O-sulfate (2), trans-ferulic acid-4-O-sulfate (3), trans-m coumaric acid-3-O-sulfate (4), trans-caffeic acid (5), m-hydroxyphenylpropionic acid (6), trans-p-coumaric acid (7), trans-m-coumaric acid (8), luteolin (9), and apigenin (10) were detected in the urine, whereas four metabolites, scutellarein 6, 7-di-O-beta-glucuronide (11), apigenin-4'-O-sulfate-7-O-beta-glucuronide (12), apigenin-7-O-beta-glucuronide (13), and diosmetin-7-O-beta-glucuronide (14) were found in the bile. Compounds 1-8 and 11-14 were also found in the plasma. When the extract was given to humans, however, two metabolites, 1-O-(2,4,5 trimethoxycinnamoyl)-beta-glucuronic acid (15) and apigenin-4'-O-beta-glucuronide (16), were found in the urine and plasma. Thus, a species difference in the metabolism of the extract constituents was observed between rats and humans. Structures 1-16 were identified based on their chemical and spectral data. PMID- 10706427 TI - Identification and clearance involved in the formation of glucuronides of RT 3003, a new peripheral blood flow enhancer, and its metabolite in rats. AB - Glucuronides of RT-3003 and its metabolite (9-OH-RT-3003), which was hydroxylated at the 9 position on the benzene ring, were separated by HPLC and identified by liquid chromatography (LC)/MS/MS and NMR. The conjugation sites of these glucuronides were determined by nuclear Overhauser effects (NOE) irradiation; RT 3003 was conjugated at an alcoholic hydroxyl group of the hydroxymethyl moiety, and 9-OH-RT-3003 at a phenolic hydroxyl group on a benzene ring and at an alcoholic hydroxyl group of a hydroxymethyl moiety. On a reversed-phase HPLC of 9 OH-RT-3003, alcoholic glucuronide was eluted later than phenolic glucuronide, indicating the high hydrophobicity of alcoholic glucuronide. Clearance for the glucuronidation (ClG) of RT-3003 was lower than the summation of ClG for two types of glucuronidation of 9-OH-RT-3003. ClG of 9-OH-RT-3003 was high in phenolic glucuronide. The activity of UDP-glucuronyltransferase (UDPGT) for RT 3003 was 9.63 times that for 9-OH-RT-3003, and the activity ratio of the two types of glucuronidation of 9-OH-RT-3003 was similar to the ratio of the corresponding ClG. The difference between ClG and UDPGT activity is discussed in association with clearance for the hydroxylation and interaction of substrates with UDPGT. PMID- 10706429 TI - Influence of central set on anticipatory and triggered grip-force adjustments. AB - The effects of predictability of load magnitude on anticipatory and triggered grip-force adjustments were studied as nine normal subjects used a precision grip to lift, hold, and replace an instrumented test object. Experience with a predictable stimulus has been shown to enhance magnitude scaling of triggered postural responses to different amplitudes of perturbations. However, this phenomenon, known as a central-set effect, has not been tested systematically for grip-force responses in the hand. In our study, predictability was manipulated by applying load perturbations of different magnitudes to the test object under conditions in which the upcoming load magnitude was presented repeatedly or under conditions in which the load magnitudes were presented randomly, each with two different pre-load grip conditions (unconstrained and constrained). In constrained conditions, initial grip forces were maintained near the minimum level necessary to prevent pre-loaded object slippage, while in unconstrained conditions, no initial grip force restrictions were imposed. The effect of predictable (blocked) and unpredictable (random) load presentations on scaling of anticipatory and triggered grip responses was tested by comparing the slopes of linear regressions between the imposed load and grip response magnitude. Anticipatory and triggered grip force responses were scaled to load magnitude in all conditions. However, regardless of pre-load grip force constraint, the gains (slopes) of grip responses relative to load magnitudes were greater when the magnitude of the upcoming load was predictable than when the load increase was unpredictable. In addition, a central-set effect was evidenced by the fewer number of drop trials in the predictable relative to unpredictable load conditions. Pre-load grip forces showed the greatest set effects. However, grip responses showed larger set effects, based on prediction, when pre-load grip force was constrained to lower levels. These results suggest that anticipatory processes pertaining to load magnitude permit the response gain of both voluntary and triggered rapid grip force adjustments to be set, at least partially, prior to perturbation onset. Comparison of anticipatory set effects for reactive torque and lower extremity EMG postural responses triggered by surface translation perturbations suggests a more general rule governing anticipatory processes. PMID- 10706430 TI - Dimorphic features of the different alpha-containing GABA-A receptor subtypes in the cortico-basal ganglia system of two distantly related mammals (hedgehog and rat) AB - This investigation represents a first study dealing with the dimorphic differences of the main alpha-containing gamma-aminobutyric acid (GABA(A)) receptors in the brain of two distantly related mammals (hedgehog and rat). The labeling of these receptors in the presence of zolpidem (highly selective benzodiazepine agonist) and under the different degree of GABA(A)/benzodiazepine allosteric coupling activities accounted for a heterogeneous colocalization of alpha1-enriched and of alpha2/3-enriched and alpha5-enriched GABA(A) receptors in some areas of the cortico-basal ganglia system (including the important ventrolateral thalamic station) of both mammalian sexes. In the hedgehog, the greatest (P<0.001) GABA-dependent reduction of zolpidem inhibition constants was mostly registered in alpha1- and/or alpha5-enriched areas, such as the frontoparietal cortex lamina III (235%), ventrolateral thalamic nucleus (128%), and substantia nigra pars reticulata (110%) of the male. However, the greatest reductions in the rat were instead detected in the male substantia nigra pars reticulata (192%) and female striatum (120%), areas which are enriched either by the colocalization of alpha1- with alpha2/3-subunits or by all three alpha subunits. These results support the contention that a sex-related alpha containing GABA(A) receptor sensitivity constitutes an important element in the execution of skilled motor activities during the different socio-sexual behaviors of the two mammals. PMID- 10706428 TI - Afferent diversity and the organization of central vestibular pathways. AB - This review considers whether the vestibular system includes separate populations of sensory axons innervating individual organs and giving rise to distinct central pathways. There is a variability in the discharge properties of afferents supplying each organ. Discharge regularity provides a marker for this diversity since fibers which differ in this way also differ in many other properties. Postspike recovery of excitability determines the discharge regularity of an afferent and its sensitivity to depolarizing inputs. Sensitivity is small in regularly discharging afferents and large in irregularly discharging afferents. The enhanced sensitivity of irregular fibers explains their larger responses to sensory inputs, to efferent activation, and to externally applied galvanic currents, but not their distinctive response dynamics. Morphophysiological studies show that regular and irregular afferents innervate overlapping regions of the vestibular nuclei. Intracellular recordings of EPSPs reveal that some secondary vestibular neurons receive a restricted input from regular or irregular afferents, but that most such neurons receive a mixed input from both kinds of afferents. Anodal currents delivered to the labyrinth can result in a selective and reversible silencing of irregular afferents. Such a functional ablation can provide estimates of the relative contributions of regular and irregular inputs to a central neuron's discharge. From such estimates it is concluded that secondary neurons need not resemble their afferent inputs in discharge regularity or response dynamics. Several suggestions are made as to the potentially distinctive contributions made by regular and irregular afferents: (1) Reflecting their response dynamics, regular and irregular afferents could compensate for differences in the dynamic loads of various reflexes or of individual reflexes in different parts of their frequency range; (2) The gating of irregular inputs to secondary VOR neurons could modify the operation of reflexes under varying behavioral circumstances; (3) Two-dimensional sensitivity can arise from the convergence onto secondary neurons of otolith inputs differing in their directional properties and response dynamics; (4) Calyx afferents have relatively low gains when compared with irregular dimorphic afferents. This could serve to expand the stimulus range over which the response of calyx afferents remains linear, while at the same time preserving the other features peculiar to irregular afferents. Among those features are phasic response dynamics and large responses to efferent activation; (5) Because of the convergence of several afferents onto each secondary neuron, information transmission to the latter depends on the gain of individual afferents, but not on their discharge regularity. PMID- 10706431 TI - Light microscopic observations on the relationships between 5-hydroxytryptamine immunoreactive axons and dorsal spinocerebellar tract cells in Clarke's column in the cat. AB - Serotonin (5-HT) exerts a variety of effects on the excitability of motoneurons, interneurons, and ascending tract cells. Spinocerebellar-tract cells in the dorsal horn receive synaptic connections from serotoninergic axons, but little is known about the relationships between serotoninergic axons and dorsal spinocerebellar tract (DSCT) cells in Clarke's column. We studied these relationships by using a combination of immunohistochemical localization of 5-HT immunoreactive boutons and intracellular staining with horseradish peroxidase (HRP) or neurobiotin of identified DSCT cells in vivo. In the adult cat, Clarke's column displayed a lower density of 5-HT-immunoreactive axons and boutons than adjacent regions of the spinal gray matter. Eleven intracellularly stained DSCT cells were analyzed with light microscopy, and six of these cells were entirely reconstructed in three dimensions. A total of 3,739 close appositions (340+/-101 per postsynaptic neuron: mean +/- SD) were observed on the labeled DSCT cells. The majority (97%) of the appositions were formed on dendrites, including proximal and distal branches, with an average density of about 1.4 appositions per 1,000 microm2 of dendritic membrane. These results indicate that the bulbo spinal serotoninergic system(s) provide direct innervation of Clarke's column DSCT cells in the upper lumbar spinal cord and that the inputs are spread widely over all regions of the target neurons' soma-dendritic membrane. PMID- 10706432 TI - Visual feedback about time estimation is related to a right hemisphere activation measured by PET. AB - In previous EEG experiments we have presented a time estimation task to our subjects, who had to press a button with either the left or right index finger 3 s after an auditory warning stimulus (WS). Two seconds later a visual Knowledge of Results (KR) stimulus was presented on a screen in front, informing them about whether the movement had been made in the correct time window (a vertical line), whether it was too early (a minus sign) or too late (a plus sign). The potential distribution underlying the anticipatory attention for the KR stimulus suggested a right hemisphere network in which the prefrontal cortex, the insula Reili and the parietal cortex were involved. In the present positron emission tomography (PET) activation study we aimed to further localize the exact positions of these regions, using the same paradigm. Two conditions were compared in which the WS had to be followed by a button press with the left index finger. In experimental condition A, subjects received true information about their performance, while in condition B false information was given, utilizing the same stimuli, but randomly, thus without any relation to the actual performance. In both conditions identical stimuli were presented and identical movements were made. Therefore we applied statistical parameter mapping (SPM) for comparison of condition A with B in order to identify regional increases in perfusion related to the anticipation and use of the KR. We found in line with our predictions a right hemisphere activation of (1) BA45, (2) the junction of the posterior insula with the temporal transverse gyrus and (3) the posterior part of the parietal cortex. This activation pattern was accompanied by a better performance due to KR. A second, though not predicted, effect was the increase in correct responses during the last two sessions compared to the first two sessions, independent of KR. This learning effect was accompanied by an activation of BA46 and the supplementary motor area (SMA), again in the right hemisphere. Summarizing, two different prefrontal areas in the right hemisphere were activated: a more ventral area, related to the use of external stimuli providing feedback about a past performance, in order to produce movements in time, and another mid-dorsal one, related to temporal programming on the basis of internal cues. PMID- 10706434 TI - Posture-related changes in heteronymous recurrent inhibition from quadriceps to ankle muscles in humans. AB - The possibility was investigated that changes in heteronymous recurrent inhibition (RI) from quadriceps (Q) to soleus (Sol) and tibialis anterior (TA) motoneurons (MNs) occur during postural tasks requiring cocontraction of Q with one of these muscles. Stimulation of the femoral nerve (FN), which elicited a Q H reflex discharge, was used to activate Renshaw cells. The resulting inhibition of TA and Sol MNs was assessed using three test responses: (1) the rectified and averaged ongoing electromyogram (EMG) activity in TA or Sol; (2) the motor-evoked potential (MEP) elicited by cortical stimulation in these muscles; and (3) the Sol H reflex. The characteristics of the depression (appearance and increase with the conditioning reflex discharge, short central delay and long duration) are consistent with a Renshaw origin. In addition, results obtained in control experiments (no change in the EMG suppression after an ischaemic blockade of group-I afferents from the leg, time course of the FN-induced depression of the MEP similar to that of the ongoing EMG) made a significant contribution from other pathways activated by FN stimulation unlikely. Posture-related heteronymous RI from Q was compared in different postural tasks at matched levels of background EMG activity: voluntary co-contraction of Q and of the relevant ankle muscle while sitting (control situation), postural co-contraction of Q and TA (while leaning backwards during stance), or contraction of Sol with (preparation for hopping) and without (standing on tip of toes and leaning forwards during stance) associated contraction of the Q. During stance, heteronymous RI from Q was reduced to TA (but not to Sol) while leaning backwards and to Sol in preparation for hopping, but not in the other situations. Thus, RI from Q to TA or Sol was specifically decreased when a co-contraction of the Q and of the relevant muscle operating at the ankle was required to maintain bipedal stance. It is argued that this control of Renshaw cells is descending in origin and contributes to selection of the appropriate synergism in various postural tasks. PMID- 10706433 TI - Niravoline, a selective kappa-opioid receptor agonist effectively reduces elevated intracranial pressure. AB - To ascertain the effects of niravoline (RU 51599, a selective kappa-opioid receptor agonist) on elevated intracranial pressure with mass lesion, the authors experimentally induced intracranial hypertension in cats by progressive inflation of an extradural balloon with physiological saline at the constant rate of 0.5 ml/h for 2.5 h. After 2.5 h, inflation was discontinued, but the balloon remained inflated for an additional 3 h. Immediately after cessation of balloon inflation and while the balloon remained expanded, the control group (n = 8) received ringer's lactate solution only. In the treatment group (n = 8), each cat was treated with an intravenous administration of niravoline at a dose of 1.0 mg/kg immediately after the cessation of balloon inflation and every hour for 3 h in post-inflation period (three injections total). Changes in intracranial pressure (ICP), mean arterial blood pressure (MAP), cerebral perfusion pressure (CPP), electroencephalogram (EEG), pupil size, blood gasses and pH, plasma osmolality and electrolytes, and brain water content were studied in both groups. Compared with the untreated group, niravoline treatment produced significant decreases in ICP and significant increases in CPP at 1, 2, and 3 h post-inflation in the presence of an extradural mass lesion. Brain water content was significantly reduced both in the compressed and contralateral hemispheres following niravoline treatment. No significant changes were observed in plasma osmolality and systemic arterial blood pressure following niravoline administration. The results from this present study provide further evidence that niravoline is effective in reducing elevated intracranial pressure, brain water content, and maintaining an adequate cerebral perfusion pressure even in the presence of an extradural mass lesion. Niravoline may offer a new therapeutic modality in head-injury patients with an acute intracranial, expanding mass lesion by providing a safer extended time-period until the mass can be surgically evacuated. PMID- 10706435 TI - Impaired direction and extent specification of aimed arm movements in humans with stroke-related brain damage. AB - The role of sensorimotor (S-M) areas in the specification of kinematic parameters for aiming movements was studied by comparing the performance of six subjects with unilateral stroke to that of matched control subjects. Rapid arm movements were made to one of four targets by rotating the forearm in a short (20 degrees) or long (45 degrees) arc of motion. Thus, the four targets represented two directions (flexion or extension) and two extents (short or long). Subjects with stroke used the arm ipsilateral to the side of the lesion. A timed-response paradigm was used to dissociate response initiation and specification. Subjects initiated movements in concert with the last of four regularly timed tones. A visual cue of the designated target was presented during the preparation interval (400-0 ms) before the last tone. Targets were presented in a fixed sequence (predictable condition) or a random sequence (unpredictable condition). No significant differences in performance were found between stroke and control groups in the predictable condition. In the unpredictable condition, subjects with stroke produced more direction errors and were less accurate in extent than the control subjects. As specification time increased to 400 ms, the frequency of direction errors attenuated less for stroke than for control groups, but the reduction in magnitude of extent errors was similar for the two groups. When specification was minimal (i.e., <100 ms), default responses were distributed equally between directions and clustered around the short extent. Further, wrong direction responses did not converge on the designated extent as specification time increased. This pattern of findings is consistent with a view of parameterization of planning and executing movements, in which direction and extent can be specified in parallel. Our results suggest that ipsilateral S-M areas contribute to the specification of an optimal motor program, particularly when imperative programming of unimanual goal-directed aiming movements is required. PMID- 10706436 TI - Is lower leg proprioception essential for triggering human automatic postural responses? AB - It is unknown to what extent automatic postural responses are triggered by lower leg proprioception. This issue was addressed by studying postural control in five carefully selected patients with subtle diabetic polyneuropathy (restricted to the lower legs) and 15 healthy subjects. All patients had bilaterally absent Achilles tendon reflexes and weak or absent patella tendon reflexes, but muscle strength was fully preserved. Subjects were tested while standing on a supporting, movable force-plate. The contribution of lower leg proprioception to automatic postural responses was investigated by randomly exposing the subjects to either a 4 degrees 'toe-up' rotational perturbation ('normal ankle input'), a simultaneous 4-cm rearward translation and 4 degrees toe-up rotation ('enhanced ankle input'), or a simultaneous 4-cm rearward translation and 4 degrees 'toe down' rotation ('nulled ankle input'). We recorded surface EMG (stretch reflexes and balance-correcting responses) from leg and trunk muscles, ankle torque and angular velocities of the upper and lower legs and trunk. We argued that automatic postural responses that have abnormally small amplitudes in patients and are modulated in controls with the velocity of different types of ankle rotations must receive a major input from lower leg proprioception. Conversely, automatic postural responses that are weakly modified in amplitude or onset by different ankle perturbations and are present despite nulled ankle inputs and, finally, are unaffected in patients with distal polyneuropathy must be triggered or modulated by inputs other than from lower leg proprioception. Normal postural synergies and strategies were maintained in patients, although within a given synergy the timing and amplitude of some automatic postural responses were abnormal. A few automatic postural responses appeared to be triggered or modulated by lower leg proprioception. Thus, early stretch reflexes in soleus and medial gastrocnemius were severely diminished in patients, while in controls these stretch reflexes were modulated by different ankle perturbations. Furthermore, balance-correcting responses in tibialis anterior were diminished and delayed in patients, while in controls these balance-correcting responses were modulated by different ankle perturbations. Other automatic postural responses were apparently not triggered or modulated by lower leg proprioception, but likely received a major input from more proximal sensory systems. Thus, in both groups prominent balance-correcting responses were present in several muscles (soleus, gastrocnemius, quadriceps, paraspinals and trapezius) during the 'nulled ankle input' condition, where ankle position was stabilised over the first 250 ms. During the 'enhanced ankle input' condition, where prominent ankle dorsiflexion occurred during the first 200 ms, amplitudes of balance-correcting responses were only marginally weaker in patients than in controls. We analysed body segment displacements to unveil the potential nature of proximal triggers for automatic postural responses. As opposed to the 'inverted pendulum' concept of postural control, early movement occurred in the knees, hips and trunk well before the onset of automatic postural responses. For example, during the 'nulled ankle input' condition, the lower leg moved forward with early knee flexion, followed by knee extension. The trunk extended backwards at 80 ms, which was followed by forward flexion. The absent stretch reflex and weaker balance correcting responses in patients produced changed trunk velocity profiles (mainly a reduced initial backward motion of the trunk), but lower-body segment movements showed no consistent differences between the two groups. Considering these body segment displacements, any automatic postural response with an onset within the first 200 ms could well be triggered by receptors located at the knee, hip or trunk. (ABSTRACT TRUNCATED) PMID- 10706437 TI - Role of limb movement in the modulation of motor unit discharge rate during fatiguing contractions. AB - Motor unit firing rates of the triceps brachii muscle have been shown to decline during sustained isometric contractions, but not if the fatiguing contraction incorporates arm movements. The purpose of this study was to determine the impact of the actual physical displacement of the limb on the maintenance of motor unit discharge rate during dynamic muscle fatigue. An isometric force pulse paradigm was used to recreate the motor unit activity patterns that occur during a dynamic contraction. With this paradigm, the variable force output that would occur during a dynamic contraction remained intact, but the movement of the limb was eliminated. Motor unit firing rates declined in the isometric force pulse protocol. Thus, factors related to the actual movement of the limb appear to enable the maintenance of motor unit discharge rates during fatigue. PMID- 10706438 TI - Transient expression of a functional serotonin transporter in Merkel cells during late gestation and early postnatal rat development. AB - We and others have previously identified serotonin transporter mRNA throughout the trigeminal system in the whisker region, trigeminal ganglion, trigeminal nucleus and thalamic relay stations. In order to further implicate a role for the serotonin transporter in this sensory system, we have now characterized serotonin transporter gene expression and function in primary cultures from the rat snout, at several stages of gestation. In this study, we have demonstrated a transient expression of serotonin transporter mRNA in quinacrine-positive Merkel cells between embryonic day 16 and postnatal day 5. Peak levels of mRNA occurred at embryonic day 20 and postnatal day 1. Merkel cells in culture exhibited a transient, antidepressant-sensitive [3H]-serotonin uptake, which was maximal at a time in culture corresponding to embryonic day 22 (day of birth). This transient uptake of serotonin suggests a role for this monoamine during a critical time period of the developing trigeminal sensory system. Regulation of extracellular serotonin levels by transporter activity may reflect the specific formation of the merkel cell-sensory neuron complex in an analogous mechanism by which serotonin modulates synaptogenesis in the central nervous system. PMID- 10706439 TI - Binaural interaction component and white-noise enhancement in middle latency responses: differential effects of anaesthesia in guinea pigs. AB - It is known that the response to binaural clicks is smaller in amplitude than the sum of two monaural responses. The difference is called the binaural interaction component (BIC). Also, an amplitude enlargement occurs in guinea pig middle latency responses (MLRs) to monaural clicks when white noise is applied to the other ear. This study was conducted to find out whether these two signs of binaural interaction result from the same mechanism. White noise enhancement (WNE) and BIC were computed from the evoked potential data simultaneously recorded in different phases of anaesthesia. WNE gradually decreased and disappeared with anaesthesia, but the relative amplitude of the BIC remained unchanged. This differential effect showed that different neural mechanisms must be responsible for BIC and WNE in guinea pig MLR. PMID- 10706440 TI - Outbreaks of Salmonella serotype enteritidis infection associated with eating raw or undercooked shell eggs--United States, 1996-1998. AB - During the 1980s and 1990s, Salmonella serotype Enteritidis (SE) emerged as an important cause of human illness in the United States. The rate of SE isolates reported to CDC increased from 0.6 per 100,000 population in 1976 to 3.6 per 100,000 in 1996 (Figure 1). Case-control studies of sporadic infections and outbreak investigations found that this increase was associated with eating raw or undercooked shell eggs (1). From 1996 to 1998, the rate of culture-confirmed SE cases reported to CDC declined to 2.2 per 100,000; however, outbreaks of illness caused by SE continue to occur. This report describes four SE outbreaks during 1996-1998 associated with eating raw or undercooked shell eggs and discusses measures that may be contributing to the decline in culture-confirmed SE cases. PMID- 10706441 TI - Prevalence of selected risk factors for chronic disease and injury among American Indians and Alaska Natives--United States, 1995-1998. AB - Since the 1950s, morbidity and mortality attributable to infectious diseases among American Indians and Alaska Natives (AIs/ANs) have declined and chronic diseases, especially diabetes, and injury have remained important determinants of poor health (1). Knowledge of the prevalence of behavioral risk factors for chronic disease and injury can be used to form policies and programs to improve the health of AIs/ANs. Based on data obtained from the Behavioral Risk Factor Surveillance System (BRFSS) from 1993 through 1996, CDC published regional estimates of the prevalence of 10 behavioral risk factors for AIs/ANs (2). This report updates data from the earlier report and focuses on three of the 10 risk factors for chronic disease and injury among AIs/ANs. PMID- 10706442 TI - Gene-delivery systems using cationic polymers. AB - Gene therapy will benefit a range of diseases from single-gene defects, to chronic diseases such as cancer, to vaccination. Initially, gene therapy used viral vectors, but the advantages of nonviral systems are now being fully appreciated. This review focuses on cationic polymers as a delivery system for DNA. The physicochemical characterization of DNA polycation complexes that condense and protect DNA from nuclease digestion are considered, together with further factors such as ligand targeting, endosomal escape, and nuclear localization. Where possible, the relative efficacy of different cationic polymer delivery systems is compared. PMID- 10706443 TI - Targeted delivery of radiolabeled imaging and therapeutic agents: bifunctional radiopharmaceuticals. AB - Clinical application of radioactive diagnostic and therapeutic agents constitutes one of the great advances in noninvasive medicine, nuclear medicine. The radioactive agents used in the nuclear medical field are called "radiopharmaceuticals," and are required to exhibit high and specific localization of radioactivity into target tissue. Among radionuclides used in radiopharmaceuticals, radiometals such as 99mTc and (111)In have received much attention because of their nuclear physical characteristics and widespread availability. However, since these metallic elements are not constituents of bioactive molecules, they cannot simply replace common constituent atoms in biologically interesting compounds. Thus, demand for biospecific radiopharmaceuticals constitutes a great challenge in rational design of biologically active molecules labeled with metallic radionuclides, and evolves into a generation of bifunctional radiopharmaceuticals. Molecules contain both a biologically active site and a chelating group for binding the metallic radionuclide in which attachment of a chelating group does not affect the inherent biospecificity of the mother compound. This paper describes recent progress in research of macro- and small-molecular bifunctional radiopharmaceuticals for targeted diagnosis and therapy. PMID- 10706444 TI - Plasma cell generation from B-lymphocytes via CD27/CD70 interaction. AB - To produce antibodies, the differentiation of B cells into antibody-secreting cells, plasma cells, is required. We describe that ligation of CD27, which belongs to the tumor necrosis factor receptor (TNFR) family and is a memory marker of B cells, yields crucial signals that positively control the entry of B cells into the pathway to plasma cells. The triggering via CD27 by CD27 ligand (CD70) on purified peripheral blood B cells yielded an increase in the number of plasma cells in the presence of interleukin-10 (IL-10). The differentiation into plasma cells by a combination of IL-10 and CD70-transfectants occurred in CD27+ B cells, but not in CD27- B cells. Moreover, the addition of IL-2 to the IL-10 and CD70-transfectants greatly induced the differentiation into plasma cells. In the presence of only IL-2, IL-4 or IL-6, CD70-transfectants did not promote the differentiation into plasma cells. On the other hand, CD40 signaling increased the expansion of a B cell pool from peripheral blood B cells primarily activated by IL-2, IL-10 and anti-CD40 mAb. These data demonstrate that CD27 ligand (CD70) is a key molecule to direct the differentiation of CD27+ memory B cells toward plasma cells in cooperation with IL-10. PMID- 10706445 TI - Immunophenotype of bone marrow mast cells in indolent systemic mast cell disease in adults. AB - One of the major advances in the histological diagnosis of bone marrow (BM) involvement in mastocytosis has been the specific immunohistochemical detection of tryptase on most cells (MC), which has shown to be of great diagnostic value, especially in cases of malignant mastocytosis. On the other hand, recent studies have clearly shown that bone marrow mast cells can be specifically identified and accurately enumerated using multiparametric flow cytometry, which allow a systematic analysis of the immunophenotypic characteristics of bone marrow mast cells. Once this flow cytometric approach was applied for the analysis of BMMC from mastocytosis patients clear immunophenotypical differences were found between BMMC from normal individuals and adults with mastocytosis. The most characteristic immunophenotypic feature, both in malignant and adult indolent systemic mast cell disease, being the coexpression of CD2 and CD25 antigens, never present in normal bone marrow mast cells and, which constitute an aberrant hallmark of bone marrow mast cells in adult mastocytosis. Furthermore, bone mast cells from mastocytosis display a higher reactivity for CD35, CD63, and CD69 activation-associated antigens. Based on these results it could be concluded that the use of multiparametric flow cytometric immunophenotyping of BMMC in adult patients suffering from cutaneous mastocytosis can be of great utility for the diagnosis of BM involvement; additionally, this might also help to establish the real incidence of BM involvement in cutaneous mastocytosis. PMID- 10706446 TI - CD148, a new membrane tyrosine phosphatase involved in leukocyte function. AB - Protein tyrosine phosphatases play an essential role in the control of leucocyte cell growth an differentiation. Recently a new receptor type membrane tyrosine phosphatase named CD148 has been identified. This molecule is present on the membrane of all the hematopoietic lineages as well as on several other cell types, mainly epithelial cells and its expression increases after cell activation. This molecule is able to act as a transducing molecule. Moreover, CD148 is able to modulate the signal transduction through the TCR/CD3 complex, in a manner similar to CD45. It has also been suggested that CD148 could be involved in mechanisms of differentiation and inhibition of cell growth. In addition, CD148 seems to be associated with a serine/threonine kinase in certain epithelial cell lines and leucocytes. Here, we review recent data on the expression and function of CD148 in both human, mouse and rat. PMID- 10706447 TI - Dysregulation of the protein tyrosine kinase LCK in lymphoproliferative disorders and in other neoplasias. AB - Initially identified as a T-cell specific member of the Src family of protein tyrosine kinases, Lck has become the object of intensive investigations which have revealed a key role for this kinase in the central processes controlling T cell development, activation, proliferation and survival. Experimental evidence of the oncogenic potential of Lck, together with the identification of defects in the regulation of Lck expression or activity in T-cell leukemias, suggests that dysregulation of Lck might play a role in neoplastic transformation. Here we review the data documenting a potential role for this kinase in the initiation and maintenance of the transformed state in human cancers. PMID- 10706448 TI - Hepatitis B and hepatitis C virus infections in stem cell transplantation. PMID- 10706450 TI - The benzene metabolites hydroquinone and catechol act in synergy to induce dose dependent hypoploidy and -5q31 in a human cell line. AB - Chronic exposure to high concentrations of benzene is associated with an increased incidence of myelodysplastic syndrome (MDS) and acute myelogenous leukemia (AML). Studies of patients occupationally exposed to benzene show a pattern of cytogenetic aberrations involving loss of all or part of chromosomes 5 and/or 7 as well as trisomy 8 and we have previously reported that hydroquinone (HQ) induces deletions of 5, 7 and 8. Benzene metabolism is a requirement for bone marrow toxicity and the phenolic metabolites, HQ and catechol (CAT), have been implicated in benzene hematotoxicity. A research project was designed to determine whether CAT by itself and in conjunction with HQ could directly induce loss of chromosome 5 and/or 7 and gain of chromosome 8. Using fluorescence in situ hybridization with chromosome-specific 5, 7, and 8 probes we demonstrate that 5 to 150 uM CAT does not produce chromosomal aberrations, however CAT and 25 uM HQ can act in synergy to induce dose dependent loss of these chromosomes. In addition HQ/CAT selectively induces -5q which is not observed for HQ only. These results demonstrate for the first time that CAT/HQ act in synergy to induce specific chromosome loss found in secondary MDS/AML. PMID- 10706449 TI - Inhibition of myeloma cell growth by all-trans retinoic acid is associated with upregulation of p21WAF1 and dephosphorylation of the retinoblastoma protein. AB - Retinoic acid and dexamethasone, in combination, inhibit the growth of human myeloma cell lines in a synergistic manner. Previously, we observed that all trans retinoic acid (ATRA) caused G1 arrest and inhibited clonogenic growth of the OPM-2 human myeloma cell line. This was associated with downregulation of the IL-6 receptor (IL-6R) gp80 protein, while autocrine IL-6 production and gp130 were not affected. Growth inhibition was not reversed by the addition of exogenous IL-6 or forced, constitutive expression of the IL-6 receptor gp80 protein, suggesting that the mechanism of action of ATRA may be due to effects on the post-receptor pathway. Therefore, in this study we have investigated whether growth arrest was associated with changes in the level of phosphorylation of the RB protein. ATRA decreased the level of phosphorylation of the RB protein at doses > 5 x 10(-9) M and also induced a five fold increase in p21WAF1, while levels of p27KIP1 and CDK2 were unchanged. The ATRA-mediated increase in p21 preceded the change in RB phosphorylation and G1 arrest and was not reversed by the addition of exogenous IL-6. The levels of CDK2 activity were inhibited approximately 60% in ATRA-treated cells, suggesting that the increased p21 levels were sufficient to inhibit CDK activity and cause RB hypophosphorylation. Increased levels of p21 have recently been observed in human myeloma cells exposed to dexamethasone, and we suggest that the common ability of these two agents to inhibit myeloma cell growth depends on their induction of p21. PMID- 10706451 TI - Altered expression of retinoblastoma (RB) protein in acute myelogenous leukemia does not result from methylation of the Rb promotor. AB - Prior studies demonstrated that expression of the retinoblastoma (RB) protein in acute myelogenous leukemia (AML) is heterogeneous with low expression conferring a poor prognosis. The molecular change(s) responsible for low RB expression in AML are unknown. Since methylation of the RB promoter has been shown to result in decreased expression we hypothesized that this might explain some cases of low RB expression in AML. To investigate this hypothesis Southern blotting and PCR sequencing after bisulfite conversion were used to study the methylation status of the RB gene promoter. DNA and protein lysates were prepared from the mononuclear cell fraction from peripheral blood or bone marrow samples from 46 patients with newly diagnosed AML. By Western blot 16, 22 and 8 patients had low, elevated and hyperphosphorylated patterns of RB expression respectively using previously defined criteria. The SacI endonuclease cuts a 5.7-kb or 6.8 -kb fragment, depending on polymorphism, containing the RB promoter, detected by the probe p123M1.8 that covers the RB promoter region and exon 1. The methylation sensitive endonuclease SacII cuts twice within a key hairpin loop structure in the RB promoter that contains binding sites for AP1, Sp1 and RBF1. Others have demonstrated that methylation within this hairpin loop can decrease RB mRNA transcription by up to 92%. Comparison of the SacI and SacI + SacII digestion fragments showed no evidence of methylation in the promoter region of RB in any of the patients studied. DNA from the promoter region of 11 patients with no/low RB expression was subjected to bisulfite conversion and PCR sequencing. No evidence of methylation was seen by this method either. These results suggests that hypermethylation of the RB promoter region is at best an infrequent event in AML and that RB promoter hypermethylation is not the predominant cause of the low levels of RB expression observed in 20% of AML patients. PMID- 10706452 TI - De novo and secondary acute myeloid leukemia in patients over the age of 65: a review of fifty-six successive and unselected cases from a general hospital. AB - Fifty-six patients older than 65 (median age: 77, range: 65-91) were treated in our general hospital, between January 1991 and November 1995 with the diagnosis of AML. Twenty-five were de novo AML (45%). The other cases which were considered as secondary AML (sAML) occurred after: myelodysplastic syndromes (19 cases: 34%), myeloproliferative disorders (7: 13%) or therapy of malignancies (5: 9%). Distribution of FAB subtypes was: M0: 4, M1: 8, M2: 14, M4: 10, M4eo: 1, M5: 10, M6: 1, unclassified: 8. Twenty-seven patients (48%) (de novo: 18, sAML: 9) received conventional "3 + 5" or "3 +7" induction chemotherapy +/- consolidation and maintenance (group 1). Low-dose Ara-C was given in eighteen cases (32%) (group 2), miscellaneous single agents were prescribed in seven cases and two patients received only supportive care. Sixteen early deaths (ED) (<1 month) occurred (29%). Distribution of age, WHO-PS, hyperleukocytosis, ED and median survival (MS) (3 months) was equivalent in de novo and sAML. Fifteen CR were achieved including twelve patients from group 1 (no difference between age <75 versus >75 years) and three from group 2. The MS of CR patients was 13 months. Eleven patients survived more than one year (de novo: 8). M5 subtype, fever >38 degrees C, high LDH level, WHO-PS>2, CR non achievement were predictive of reduced survival. Besides conventional induction, investigational therapies currently remain the best antileukemic modalities. Age per se should not be an exclusion criteria for treatment. Our data underline the high frequency of sAML in the community. Of note, this common type AML is largely excluded from many trials. PMID- 10706453 TI - Marrow cytokine transcripts and the secondary hematologic disorders. AB - A comparison was made of the cytokine transcripts in normal, monoclonal, MDS, and AML marrow aspirates. While both normal and monoclonal marrow aspirates contain transcripts for SCF, few MDS or AML marrow aspirates contain these transcripts. Similarly, IL1ra transcripts are found with reduced frequency in MDS and AML marrow aspirates. The fall in SCF transcripts between monoclonal and MDS marrow aspirates parallels the appearance of apoptosis and the reduced in vitro proliferative ability which are characteristics of MDS marrow aspirate cells. The frequent IL1beta production by MDS and AML marrow aspirate cells, with few marrow aspirates producing IL1ra transcripts, suggests that unbalanced IL1beta effects may contribute to the proliferative advantage of MDS and AML cells over their normal counterparts. PMID- 10706454 TI - Granulocyte-colony stimulating factor (G-CSF)-primed, delayed marrow harvests as a source of hematopoietic stem and progenitor cells for allogeneic transplantation. AB - We evaluated the ability of G-CSF to increase the number of hematopoietic stem cells obtained by "delayed" BM harvest for allogeneic transplantation. Five normal donors received G-CSF @ 10 mcg/kg/day x 5 followed by repeat PB and BM assays at day 6 and 16, and BM harvest at day 16. Stem cells were not increased in the BM at day 16. Five patients underwent BMT and engrafted at +10 to +19 days. While the tested strategy offers no intrinsic advantages, its potential cannot be evaluated fully without alternative timing and/or additional, "early acting" growth factors. PMID- 10706455 TI - Comparative genomic hybridization and conventional cytogenetic analyses in childhood acute myeloid leukemia. AB - Comparative genomic hybridization (CGH) analysis was performed on bone marrow specimens from 19 children with acute myeloid leukemia (AML) at diagnosis. The results of CGH were compared to those of conventional cytogenetic analysis. The most common CGH aberrations were gains of whole chromosomes 6 and 8, both of which appeared three times. Two losses were seen twice; losses of whole chromosomes 7 and X. The CGH findings were concordant with the results of conventional karyotyping. CGH did not add new information to the karyotypes. Since no high-level amplification was found among the samples and standard karyotyping was highly successful, we do not advocate routine use of CGH in the diagnostic evaluation of childhood AML. PMID- 10706456 TI - Intensive dose ifosfamide and etoposide with G-CSF for stem cell mobilization in patients with non-Hodgkin's lymphoma. AB - We studied 36 patients with non-Hodgkin's lymphoma to evaluate the stem cell yield following recovery from intensive dose ifosfamide and etoposide given as mobilization chemotherapy. We also assessed the toxicity of the regimen and engraftment kinetics. All patients had intermediate grade lymphoma and had either failed to achieve a complete remission to induction chemotherapy or had relapsed. Patients received ifosfamide 10 g/m2 IV total dose given over 72 hours, etoposide 150 mg/m2 IV every 12 hours for 6 doses and G-CSF 10 microg/kg/d. Thirty-four patients went on to receive high-dose chemotherapy with BEAM or with CVP and BEAM. A median of 2 (1-10) apheresis was required to reach the target CD34+ count of >4 x 10(6)/kg. A median of 13.1 CD34+ cells/kg (4.1-148) was obtained. Toxicity was limited to mucositis in 3 patients, transient confusion and transient rise in liver function tests in 3 and 2 patients respectively. The median time to engraftment was 10 days (8-17) for all the patients undergoing high-dose chemotherapy. The regimen of intensive dose ifosfamide and etoposide along with G-CSF is well tolerated and in this group of patients has lead to successful stem cell harvests and sustained engraftment. PMID- 10706457 TI - Cytogenetic analysis of non Hodgkin's lymphomas by ratio-painting and comparative genomic hybridization reveals unsuspected chromosomal abnormalities. AB - Cytogenetic analysis of cancer cells has proven to be a powerful tool in understanding malignant evolution and in providing clinically useful markers. In recent years the advent of new fluorescence in-situ hybridization (FISH) methods such as ratio-painting and comparative genomic hybridization (CGH) have enabled much more accurate karyotypes of malignant cells to be detected. In this study, we have examined the chromosomes present in malignant cells from a series of 6 low grade follicular centre and 2 high grade diffuse large cell non-Hodgkin's lymphomas (NHL) using conventional G-banding. In all cases chromosome abnormalities were observed, including the presence of marker chromosomes in six cases. The NHL cells were then subjected to the FISH method of ratio-painting. This provided a more accurate understanding of the origins of derivative chromosomes and identified the origins of all of the marker chromosomes. It also revealed hitherto unsuspected abnormalities. For example, in one case four abnormal chromosomes were demonstrated to contain material from chromosome 8, which had not been previously suspected from G-banding. Regions of amplification and deletion on the chromosomes were also investigated by CGH, which identified further unsuspected chromosomal abnormalities. For example, in case L124, trisomy of chromosome 7 was confirmed by CGH, but an unsuspected amplification of 3(p12) was also revealed. These approaches demonstrate the power of FISH technology in providing a more precise analysis of malignant cell chromosomes, and in doing so have produced comprehensive karyotypes of the NHL under study. PMID- 10706458 TI - Epic as an effective, low toxicity salvage therapy for patients with poor risk lymphoma prior to beam high dose chemotherapy and peripheral blood progenitor cell transplantation. AB - We treated 33 patients with relapsed or refractory non-Hodgkin's lymphoma (NHL) or Hodgkin's disease (HD) with a combination of etoposide, prednisolone, ifosfamide and carboplatin (EPIC). After a median of two courses (range 1-5) complete response was achieved in 7 (22%) patients and partial response in 12 (37%) patients, an overall response rate of 59%. The regimen was well tolerated with myelosuppression being the most common toxicity. There were no toxic deaths. 25 (78%) patients were able to proceed to high dose therapy (BEAM) with peripheral blood progenitor cell transplantation either immediately post EPIC or following further salvage therapy. Most patients mobilised peripheral blood progenitor cells well and 24 out of 25 patients subsequently undergoing autologous transplantation had rapid regeneration of counts. EPIC is an effective salvage therapy in the majority of patients with relapsed or refractory lymphoma and does not appear to be toxic to stem cells. Although severe, myelosuppression is of short duration and the generally low toxicity enables patients to proceed to successful peripheral blood stem cell harvest and transplantation. PMID- 10706459 TI - Treatment of hairy cell leukemia-variant with cladribine. AB - Hairy cell leukemia-variant (HCL-V) is an extremely rare chronic B-cell lymphoproliferative disorder clinically and morphologically distinct from classic hairy cell leukemia (HCL). HCL-V is thought to represent a hybrid between prolymphocytic leukemia and HCL, the nucleus more closely resembling a prolymphocyte and the cytoplasm a hairy cell. The clinical course of HCL-V is aggressive with short survivals. Since single courses of cladribine have profound activity in HCL, inducing durable complete responses in 91% of patients, we administered cladribine to 4 patients with HCL-V over a 7-year period. During this time interval 357 patients with classic HCL received cladribine at Scripps Clinic. Each patient received cladribine at 0.1 mg/kg per day by continuous intravenous infusion for 7 days, repeated at 28-day intervals depending on response status. The 4 patients ranged in age from 28 to 70. Two presented with B symptoms, 1 had peripheral adenopathy, and all 4 displayed massive splenomegaly. Peripheral blood counts were notable for lymphocytosis associated with mild anemia and thrombocytopenia. Only 1 of the 4 patients had received prior treatment. Peripheral blood immunophenotypic analysis revealed monoclonal B cells with expression of CD11c in 3 patients, lack of CD25 expression in 3 patients and expression of CD103 in all but 1 patient. The number of cladribine courses administered ranged from two to five. Of these 4 patients, 1 (25%) achieved a complete response and 2 (50%) partial responses, for an overall response rate of 75%. Three patients underwent splenectomy after cladribine. Cladribine is an active agent in HCL-V albeit with a lower response rate than in classic HCL. The role of other treatment modalities, such as splenectomy, interferon-alpha, and 2' deoxycoformycin, alone or in combination with cladribine awaits further evaluation. PMID- 10706460 TI - Influence of low dose rIL-2 treatment on endogenous cytokine production, expression of surface IL-2R and the level of soluble IL-2R in patients with minimal residual disease. AB - This study was designed to investigate the immunomodulatory effect of low-dose IL 2 therapy (100 microg/day for 3 weeks) on interferon (IFN), tumor necrosis factor (TNF) production in vivo and in vitro and on the expression of IL-2Ralpha/beta and soluble form of IL-2Ralpha. Patients enrolled in the study suffered from multiple myeloma (MM), Hodgkin's disease (HD) and non-Hodgkin's lymphoma (NHL) All of them were in remission after chemotherapy or radiotherapy. Our results indicated that IL-2 given subcutaneously at a low dose of 100 microg/day for 3 weeks induced IFN-gamma and TNF-alpha in plasma (measured 24 hrs after the last dose of IL-2) and affected the ability of blood leukocytes to produce cytokines. Production of IFN-gamma induced in vitro with PHA was enhanced, but TNF-alpha production induced by lipopolysaccharide (LPS) and virus (Newcastle Disease Virus) was depressed. The expression of both: surface IL-2R, especially beta subunit on total population of lymphocytes and NK cells, and soluble form of IL 2R, of chain were significantly enhanced after low-dose IL-2 therapy. Low dose IL 2 therapy was well tolerated by all patients, and side effects not exceeding II grade of toxicity according to WHO scale were observed. Five patients with MM relapsed 3-10 month after cessation of IL-2 therapy, but all patients with Hodgkin's and non-Hodgkin's lymphomas are still in remission (20 months of observation). PMID- 10706461 TI - Dihydropteridine reductase activity and neopterin levels in leukemias and lymphomas: is there any correlation between these two parameters? AB - Urinary neopterin levels, blood dihydropteridine reductase activity as well as other frequently used clinical parameters were evaluated in 110 patients suffering from various types of lymphomas and leukemias. Among them neopterin was detected as the most sensitive marker representing the severity of malignancy (p<0.00001). All patients with active diseases had significantly raised urinary neopterin levels compared to those in remission and healthy controls. Of 69 patients with active disease 66 (96%) were above the upper limit seen in healthy subjects. In addition, the highest neopterin excretion was found in patients with active chronic myeloid leukemia (1469+/-479 micromol/mol creatinine n=16). In contrast, only 1 of 41 patients in stable responsive disease and remission (2.4%) had increased urinary neopterin levels above the upper limit. Dihydropteridine reductase (DHPR) activities were also detected in all patients and control groups. In active disease slightly reduced (DHPR) activities were evident (3.42+/ 0.37 for controls, 2.92+/-0.39 in active disease and 3.28+/-0.42 nmol red cytochrome C/min/5 mm diameter disc in remission patients). However in patients under medication this was strengthened. This data also suggest that DHPR activity can be effected by chemotherapy. The results of the present study support the fact that urinary neopterin levels may be an useful and reliable early prognostic marker for neoplasia when used together with other prognostic indicators. Our data also suggest that reductions in DHPR activities may also be an underlying cause for the neurological disorders that are commonly seen in patients with haematological malignancies. PMID- 10706462 TI - Evaluation of mitoguazone in patients with refractory chronic lymphocytic leukemia: a phase II study (P-H482) of the Eastern Cooperative Oncology Group. AB - Mitoguazone is a unique antitumor agent that interferes with polyamine synthesis that has been reported to have activity against AIDS-related malignant lymphoma. We, therefore, tested this agent for activity against chronic lymphocytic leukemia (CLL) in this phase II study. Mitoguazone, 500 mg/M2 was given intravenously weekly to 13 patients with relapsed or refractory, previously treated Rai stages 2-4 CLL. There were no complete or partial responses as judged by standard criteria. Toxicity was acceptable. Mitoguazone in the dose and schedule employed in this study has no significant activity as a single agent in patients with relapsed or refractory CLL. PMID- 10706463 TI - Human herpesvirus 8 and Epstein Barr-virus in a cutaneous B-cell lymphoma and a malignant cell line established from the blood of an AIDS patient. AB - Human Herpesvirus 8 (HHV-8) has been consistently associated with Primary Effusion Lymphoma (PEL or body-cavity-based lymphoma) but not with other lymphomas. This paper reports on an AIDS patient without obvious malignant effusion in body cavities but with a cutaneous lymphoma where HHV-8 and Epstein Barr virus (EBV) were detected by PCR and electron microscopy. Both viruses were also detected in all the cells of a malignant cell line (BBG1) established from the patient's peripheral blood mononuclear cells. As in PEL and PEL-derived cell lines, both the tumor and the lines lacked B-antigen expression in immunological studies but were of the same B origin as shown by clonal immunoglobulin gene rearrangements. In contrast to other co-infected cell lines, BBG1 and subclones spontaneously expressed the HHV-8 lytic antigens p40, p27, p60 and the EBV transforming latent antigen EBNA2. These data suggest that the clinical and biological features of HHV-8-and EBV-associated lymphomas could be wider than has been described to date in PEL particularly with the in vivo presence of circulating malignant dually-infected cells engaged in a spontaneous HHV-8 lytic infection. PMID- 10706464 TI - Successful treatment of immunoblastic lymphadenopathy-like T-cell lymphoma with cyclosporin A. AB - Immunoblastic lymphadenopathy (IBL)-like T-cell lymphoma is considered to belong to peripheral T-cell lymphoma. Its prognosis is grave and effective treatments have not been established. Recently, we gave oral cyclosporin A (CsA) to a patient with IBL-like T-cell lymphoma, and succeeded in achieving dramatic remission. In this case, serum levels of interleukin-12 (IL-12) and tumor necrosis factor-alpha (TNF alpha) were elevated and decreased or returned to normal after achieving remission. Since CsA is a potent suppressor of the immune system and most notably T-cells, the immunosuppression of T-cell function might have played an important role in achieving remission in this case, although the precise mechanism still remains to be elucidated. The present case indicates that administration of CsA may be a very effective and safe selection of therapy for IBL-like T-cell lymphoma, as well as analogous disorders such as IBL and angioimmunoblastic lymphadenopathy with dysproteinemia (AILD), thereby will contribute to improving the prognosis of patients with these diseases. PMID- 10706465 TI - Gastric-Mucocutaneous gammadelta T cell lymphoma: possible association with Epstein-Barr virus? AB - Gammadelta T cell lymphoma is usually either subcutaneous or hepato-splenic and involvement of other extranodal sites is rare. Here we report an unusual case of gammadelta T cell lymphoma involving the subcutaneous tissue, vocal cords, gastric mucosa and the central nervous system with a rapidly progressive clinical course and fatal outcome. Epstein-Barr virus (EBV) was shown to be present in the tumor cells, and is thought to play a role in the pathophysiology of this particular case of lymphoma. PMID- 10706466 TI - Primary bilateral adrenal lymphoma associated with idiopathic thrombocytopenic purpura. AB - Autoimmune disorders are occasionally associated with malignant lymphoma. To date only one case of primary adrenal lymphoma associated with idiopathic thrombocytopenic purpura (ITP) has ever been reported. This paper reports the case of a 63-year-old man with bilateral adrenal masses whose laboratory data showed decreased platelet count. Despite normal blood pressure, the adrenal tumors endocrinologically appeared to be pheochromocytoma. Core needle biopsy was not done due to thrombocytopenia attributed to concurrent ITP. After intravenous immunoglobulin treatment, splenectomy and bilateral adrenalectomy were performed since the first pathological findings of the frozen specimen suggested the possibility of a poorly-differentiated carcinoma. Immunohistochemical study finally showed the tumors to be diffuse large B-cell lymphoma. The patient underwent a subsequent course of combination chemotherapy and survived 6 years recurrence-free without any need for further treatment other than steroid replacement. The coincidence of adrenal lymphoma and ITP should be considered even if another kind of tumor is suspected, and core needle biopsy should be performed prior to operation, since the specific kind of tumor found alters the therapeutical strategy adopted. PMID- 10706467 TI - Chronic myelogenous leukemia in chronic phase for 16 years: ongoing hematological remission and late minor cytogenetic response under minimal interferon maintenance therapy. AB - Philadelphia chromosome-positive chronic myelogenous leukemia was diagnosed in a now 37-year old woman 16 years ago. Induction therapy with hydroxyurea and busulphan led to hematological remission lasting for about 4 months without treatment. Then, intermittent busulphan over a 7 years' period, and subsequently, alpha-interferon was given, of which ever decreasing doses (currently 3.5 megaunits interferon-alpha-2c once every 14 days) have been required to keep leukocyte counts in the target range. Although no major cytogenetic response was achieved by maintenance therapy, the patient has now been in an ongoing chronic phase of disease for 16 years. This is a rare case of indolent chronic myelogenous leukemia, in which, for undefined reasons, the leukemic cells have not acquired the capacity to transform leading to disease acceleration, which usually is imminent after a few years. PMID- 10706468 TI - Trisomy 13 is associated with poor prognosis in idiopathic myelofibrosis with myeloid metaplasia. AB - We report a case of idiopathic myelofibrosis with trisomy 13 as the sole clonal aberration, as demonstrated by metaphase cytogenetics. The clinical course was especially poor in this case, with death in blast crisis occurring within two weeks from diagnosis. The dismal outcome bears striking similarity to two previous cases of idiopathic myelofibrosis and trisomy 13 reported in the literature. Therefore trisomy 13 may be a predictor of a rapidly fatal outcome in this otherwise indolent disease. Fluorescence in situ hybridisation (FISH) with a chromosome 13 specific probe may enhance the detection of this aberration, since only 50% of cases of idiopathic myelofibrosis are karyotyped successfully using conventional techniques. PMID- 10706469 TI - Metastatic extramedullary plasmacytoma of the lung. AB - Extramedullary plasmacytomas (EMP) comprise 4% of all plasma cell neoplasms and commonly in the upper airway or digestive tract but rarely develop in the lungs. We present a case of primary pulmonary plasmacytoma in an 89 year old man, presented as a hilar mass with associated intrathoracic and extrathoracic lymph node metastases, but without evidence of myeloma. Treatment options for EMP include surgery, surgery and radiotherapy, surgery and chemotherapy or chemotherapy alone. Local recurrence rate is reported as 10-30%, with 17-48% progressing to multiple myeloma and median survival being 63-101 months. In view of the advanced age of this patient, who was initially treated unsuccessfully with intravenous cyclophosphamide and subsequently with two cycles of VAMP chemotherapy with good resolution of his disease, he is undergoing regular follow up only. PMID- 10706470 TI - Impact of treatment provision on the epidemiological recording of root caries. AB - The estimation of root caries prevalence and the identification of risk factors for decay depend upon the successful identification of carious lesions in epidemiological studies. Root surface restorations can either be placed to manage decay or cervical wear/sensitivity. The handling of data for restorations during epidemiological surveys is critical to the accurate assessment of caries prevalence. The objective of this study was to determine the relative frequency of dentists' placing root surface restorations according to their reason for placement. Data for 696 restorations were recorded from 35 dentists. Forty-five % of restorations were placed because of decay compared with 55% for cervical wear/sensitivity. There were no significant differences in proportion of placement of restoration with age of the patient or between regular and irregular attenders of different ages. Using these data a correction factor was developed for inclusion in the Root Caries Index (RCI) to make allowance for the proportion of restorations placed because of wear/sensitivity. When this correction factor was introduced into an existing data-set for root caries, the RCI was reduced for all groups. This reduction was greatest in regular attenders. When these data were analysed without making allowance for treatment effects, there was a significant difference in RCI between regular and irregular attenders. When the correction factor was applied to these data, this difference was eliminated. PMID- 10706471 TI - Smallest detectable difference of maximal mouth opening in patients with painfully restricted temporomandibular joint function. AB - Changes in maximal mouth opening reflect the impact of temporomandibular disorders and the effect of a therapeutic intervention. No information about the amount of change in maximal mouth opening with regard to reasoned decision-making is available. The smallest detectable difference, as a measure of reliability assessment, provides this information and is expressed in the unit of the measurement instrument. Twenty-five consecutive patients (5 males, 20 females) with a painfully restricted temporomandibular joint participated in this study. Measurements of maximal mouth opening were performed by two well-trained observers on two separate measurement days, one week apart. The maximal mouth opening measurements were repeated three times. Inter-observer, intra-observer, and test-retest reliability varied between 0.90 and 0.96. Inconsistency in measurement results analyzed in terms of absolute error variance, i.e. the measurement facets plus all the interactions, represented 11% of total variance. The smallest detectable difference of maximal mouth opening varied from 9 to 6 mm. For being successful in painfully restricted temporomandibular joint patients, statistically as well as clinically, the clinician has to measure at least 9 mm of improvement in maximal mouth opening. To reduce the smallest detectable difference from 9 to 6 mm, repeated measurement is necessary. PMID- 10706472 TI - Changes in function and in pain-related and cognitive-behavioral variables after arthroscopy of temporomandibular joints. AB - The purpose was to prospectively evaluate changes in clinical, pain, and cognitive-behavioral variables in a structurally homogenous group of patients with painful temporomandibular joints (TMJ), who had undergone an identical intervention, arthroscopy. Twenty-six consecutive patients who had previously undergone unsuccessful conservative treatment participated. They were evaluated with the Craniomandibular Index (CMI), pain-related measures on visual analogue scales (VASs) for 1 wk, questionnaires, and the Multidimensional Pain Inventory (MPI). The mean CMI decreased significantly, from 0.28 to 0.18, 3 months after surgery. Pain measures also decreased significantly as rated on questionnaires, and "at worst" and "most of the time" on VASs. Intrapsychic variables related to pain also decreased significantly, while interpersonal and activity measures remained unchanged. An overall MPI dysfunctional variable correlated significantly with pain. Few further changes were observed at 12 months. Lysis and lavage of the upper TMJ compartment appears to effectively alleviate persisting functional and pain-related symptoms with low morbidity, in line with previous findings. Recoveries seem to be accompanied by changes in certain pain related cognitive-behavioral variables within a limited sphere. Biological and intrapsychic features may interact with interpersonal factors in a complicated way in patients with orofacial pain. PMID- 10706473 TI - Water characteristics associated with the occurrence of Legionella pneumophila in dental units. AB - This study evaluated the incidence of Legionella pneumophila in dental unit water samples and investigated how the occurrence of these bacteria may be related to some physical, chemical and bacteriological characteristics of the water. The samples were taken from the incoming tap water, oral rinsing cup, air-water syringe, ultrasonic scaler, and the turbine of 23 dental units of private and public institutions. Apart from L. pneumophila (serogroup 1 and 3) isolated in 22 out of the 101 (21.8%) water samples tested, two other species were found: L. bozemanii and L. dumoffii. The highest densities and frequency of L. pneumophila were observed in the water coming into the units and in the dental units of public institutions. A negative association between L. pneumophila and 36 degrees C and 22 degrees C heterotrophic total plate counts and other gram-negative bacteria was found. An inverse association between the concentration of L. pneumophila and water temperature was also observed. The values of pH and total hardness did not show any significant difference in the L. pneumophila-positive and -negative dental unit waters. Finally, the chemical oxygen demand (COD) and residual chlorine were found to correlate positively with L. pneumophila. PMID- 10706474 TI - Inhibitory effect of low-level laser irradiation on LPS-stimulated prostaglandin E2 production and cyclooxygenase-2 in human gingival fibroblasts. AB - It has been reported that lipopolysaccharide (LPS) from periodontal pathogens can penetrate gingival tissues and stimulate the production of prostaglandin E2 (PGE2), which is known as a potent stimulator of inflammation and bone resorption. Although biostimulatory effects of low-level laser irradiation such as anti-inflammatory results have been reported, the physiological mechanism is not yet clarified. The purpose of the present study was to determine the effect of laser irradiation on PGE2 production and cyclooxygenase (COX)-1 and COX-2 gene expression in LPS-challenged human gingival fibroblast (hGF) cells in vitro. hGF cells were prepared from healthy gingival tissues and challenged with LPS, and Ga Al-As diode laser was irradiated to the hGF cells. The amount of PGE2 released in the culture medium was measured by radioimmunoassay, and mRNA levels were analyzed by reverse transcriptase-polymerase chain reaction (RT-PCR). Irradiation with Ga-Al-As diode low-level laser significantly inhibited PGE2 production in a dose-dependent manner, which led to a reduction of COX-2 mRNA levels. In conclusion, low-level laser irradiation inhibited PGE2 by LPS in hGF cells through a reduction of COX-2 mRNA level. The findings suggest that low-level laser irradiation may be of therapeutic benefit against the aggravation of gingivitis and periodontitis by bacterial infection. PMID- 10706475 TI - Molecular cloning of a human dentin sialophosphoprotein gene. AB - Dentin sialoprotein (DSP) and dentin phosphoprotein (DPP; phosphophoryn) are two principal dentin-specific non-collagenous proteins. DPP is extremely acidic and is rich in aspartic acid and serine. By virtue of this structure, DPP may bind large amounts of calcium and may facilitate initial mineralization of dentin matrix collagen as well as regulate the size and shape of the crystals. The function of DSP is not known. DSP and DPP are encoded by a single gene in both rat and mouse, and are uniquely expressed in odontoblasts and transiently in pre ameloblasts. Because DSP and DPP are isolated from dentin as distinct proteins and appear to be present in different amounts, the nascent dentin sialophosphoprotein (DSPP) is likely cleaved to yield DSP and DPP. However, when, where and how the DSPP is cleaved into DSP and DPP is not clear. To further elucidate the structure and function of human DSP and DPP, we have cloned DPP and DSP cDNA by reverse transcriptase-polymerase chain reaction (RT-PCR) strategies, and then cloned and initiated characterization of a human dentin sialophosphoprotein gene. The genomic organization of human DSPP is very similar to that of mouse, containing five exons and four introns, suggesting it is a homologue of mouse dentin sialophosphoprotein (DSPP). Exons 1-4 encode for DSP, while exon 5 encodes for the C-terminus of DSP and the whole DPP. A 4.6-kb RNA transcript was detected on Northern blot analyses of total RNA extracted from immature (open root apices) human teeth using either a human DPP or DSP probe. PMID- 10706476 TI - Expression of cytokeratins in human enamel organ. AB - Cytokeratin (CK) is a filament which plays a central role in epithelial tissue and, like the polypeptides of intermediate filaments in general, shows a high degree of tissue specificity. The CK expression patterns of odontogenic epithelia are still poorly described. We studied the distribution of individual CK polypeptides in the human enamel organ at bell stage and in remnants of the dental lamina. Our immunohistochemical study showed that epithelial cells stained for CKs 7, 13, 14 and 19 with slight changes in their pattern during the differentiation phase of odontogenesis. There was negative staining for all other CK polypeptides tested (CKs 8, 10, 16, 17 and 18). Most of the CKs in the enamel organ epithelia did not show differences related to the stage-specific state of differentiation, except for CKs 14 and 19 at the inner enamel epithelium. A strong label for CK 14 was present at the inner dental epithelium at early bell stage, and this was substituted by CK 19 at the late bell stage when the ameloblasts were fully differentiated. PMID- 10706477 TI - Fluoride effect on the activity of enamel matrix proteinases in vitro. AB - Dental fluorosis is common in individuals exposed to different sources of fluoride during tooth development. The mechanism causing this enamel defect is still unknown. Enamel matrix proteinases play a central role in the maturation of dental enamel, and inhibition of these enzymes by fluoride has been one explanation for dental fluorosis. We have investigated the effect of fluoride on the activity of enamel matrix proteinases using a colorimetric assay, casein zymography, and an enamel protein degradation assay. Fluoride (625 microM to 10 mM) inhibited neither the enzymatic activity of the crude matrix extract nor the activity of individual enamel enzymes separated by SDS-PAGE. The proposition that fluoride could directly inhibit enzymes was not confirmed in this study. PMID- 10706478 TI - Cultured tumor cells of murine submandibular gland origin: a model to investigate pHi regulation of salivary cells. AB - Intracellular pH (pHi) and several ion transport mechanisms in cultured murine salivary gland cells (SCA-9) were studied using a videomicrofluorometric method and the H+-specific probe C-SNARF-1. The aim of this study was to test SCA-9 cells' pHi regulation mechanisms and evaluate if this cell line is representative of submandibular gland cells. Resting pHi in unstimulated cells was estimated to be 7.17+/-0.07. To investigate the presence of Na+/H+ and Cl-/HCO3- antiports as well as Na+/K+/2Cl- symports in SCA-9 cells, we used different specific blockers, dimethyl-amiloride, disulfonic stilbene, bumetanide and furosemide. In order to study SCA-9 cell capacity to regulate their pHi in response to alkaline and acid loads, we applied the NH4Cl prepulse method to all these blockers. The results showed that SCA-9 cells possess both antiports and symports involved in pHi regulation, and that this cell line can be used as a convenient model to study pHi regulatory mechanisms in salivary cells. PMID- 10706479 TI - Patterns of cell death induced by eluates from denture base acrylic resins in U 937 human monoblastoid cells. AB - The purpose of this study was to investigate in vitro the apoptosis- and necrosis inducing potential of eluates from three heat-polymerized and four autopolymerized poly(methyl methacrylate)-based denture base resins. Our hypothesis was that the rate of cell death by apoptosis and/or necrosis induced by such denture base resins could be an important indicator of their cytotoxicity degree. U-937 human monoblastoid cells were exposed for 24 h and 48 h to eluates of 0.1 g/ml, 0.2 g/ml, 0.4 g/ml, and 0.8 g/ml extracted for 24 h and 48 h. The characteristics of apoptosis and necrosis were evaluated by flow cytometry and light and electron microscopy. Eluates from all resins enhanced cell death by apoptosis and necrosis in U-937 cells in a dose- and time-dependent fashion. Eluates from autopolymerized resins yielded higher percentages of apoptosis and necrosis than the heat-polymerized ones. The results support our hypothesis that eluates of poly(methyl methacrylate)-based denture base acrylic resins activate death-signaling pathways, and that the extent of this process reflects their biocompatibility degree. PMID- 10706480 TI - Accuracy of radiographic assessment of interproximal bone loss in intrabony defects using linear measurements. AB - The aim of the present study was to examine the accuracy of linear measurements on radiographs of interproximal bone loss in intrabony defects utilizing the gold standard of surgical measurements. In 22 patients with untreated advanced periodontal disease, 33 standardized radiographs were taken presurgically. The horizontal and vertical angulation difference of the central beam from the orthoradial projection and radiographic magnification was calculated for each radiograph. At the time of surgery, for 34 interproximal intrabony defects, the distances from the cemento-enamel junction (CEJ) to the bottom of the bony defect (BD) and alveolar crest (AC) to BD, and the height of the one-, two- and three walled section of the intrabony defects were measured. In all radiographs, the linear distance CEJ to BD was assessed. Radiographic and surgical assessments were compared. A stepwise multiple linear regression analysis was used to evaluate factors (angulation difference, patient, surgical measurements) that influenced the discrepancy between radiographic and surgical measurements. The radiographic assessments underestimated bone loss as compared to surgical measurements (1.41+/-2.58 mm). PMID- 10706481 TI - Ultrastructural preservation of rat embryonic dental tissues after rapid fixation and dehydration under microwave irradiation. AB - Adequate preservation of the cells and matrix of mineralising tissues remains difficult, as organic components and initial mineral deposits may be lost during conventional processing for electron microscopy. In this study, we have reduced significantly the processing time using microwave irradiation. Rat molar tooth germs were fixed in 4% glutaraldehyde + 4% formaldehyde with 0.1 M sodium cacodylate in a laboratory microwave oven for two periods of 20 s with a maximal temperature of 37 degrees C. After conventional washing and post-fixation, specimens were dehydrated in graded ethanols under microwave irradiation for a total of 7 min 20 s. For comparison, some specimens were processed by conventional methods. After embedding, ultrathin sections were examined by electron microscopy. In differentiating ameloblasts and odontoblasts, plasma membranes, mitochondria, rough endoplasmic reticulum, the Golgi complex, together with all other cytoplasmic organelles exhibited excellent preservation. Microtubules, microfilaments and coated vesicles were particularly evident. Crystal-like mineral deposits were conspicuously present in relation to dentine matrix vesicles and collagen fibrils as well as in enamel matrix. The matrix of forming enamel had a globular electron-lucent appearance. It is concluded that this is a rapid method which provides a preserved or even improved morphology. PMID- 10706482 TI - The Franklin expedition and lead poisoning. PMID- 10706483 TI - Delivering effective asthma care--how do we implement asthma guidelines? PMID- 10706484 TI - Is lung function really a good parameter in evaluating the long-term effects of inhaled corticosteroids in COPD? PMID- 10706485 TI - Possible future of induced sputum in interstitial lung disease. PMID- 10706486 TI - Pituitary adenylate cyclase-activating peptide 38 a potent endogenously produced dilator of human airways. AB - Pituitary adenylate cyclase-activating peptide (PACAP) 38 displays several biological activities relevant to obstructive airway disease. In this study, the occurrence of PACAP 38 in human small bronchi and corresponding pulmonary arteries was analysed immunocytochemically. The dilatory effects of this peptide on the same structures were also studied in vitro. A moderate number of PACAP like immunoreactive nerve fibres was seen in association with bronchial and vascular smooth muscle and around seromucous glands. PACAP 38 caused a concentration-dependent relaxation of precontracted bronchial and pulmonary arterial segments. The maximal relaxation was more pronounced in the airways than in the arteries, whereas the potency in both was identical. PACAP 38 caused relaxation of all segments tested (nine patients), whereas vasoactive intestinal polypeptide (VIP) failed to cause relaxation of bronchial segments from six of nine patients. Both PACAP and VIP dilated all pulmonary arterial segments tested. In conclusion, pituitary adenylate cyclase-activating peptide 38 is a potent dilator of human bronchi and is present in the human lung. Pituitary adenylate cyclase-activating peptide 38 may, therefore, play a role in the endogenous regulation of airway tone. The inhibitory effects of pituitary adenylate cyclase activating peptide 38 are more consistent than those of the related neuropeptide vasoactive intestinal polypeptide, perhaps reflecting a difference in susceptibility to degrading enzymes. PMID- 10706487 TI - Baseline airway hyperresponsiveness and its reversible component: role of airway inflammation and airway calibre. AB - Airway hyperresponsiveness (AHR), in which airway inflammation has been reported to be a key factor, is an important component of asthma. However the precise role of inflammation in AHR is still unclear. In this report, airway inflammatory changes were assessed using hypertonic saline-induced sputum examination and exhaled nitric oxide analysis, and the relation between AHR to methacholine, airway calibre forced expiratory volume in one second (FEV1) and airway inflammatory indices examined. Furthermore, the changes in these variables were also examined by means of 8 weeks' open uncontrolled inhaled steroid administration (800 microg x beclomethasone x day(-1)). Asthmatic subjects had higher eosinophil counts and bradykinin concentration in induced sputum and higher exhaled NO levels, and showed AHR to methacholine. Baseline AHR significantly correlated with FEV1 but not with indices of inflammation in sputum or exhaled air. Steroid inhalation therapy was associated with a reduction in eosinophil and bradykinin concentration in sputum and NO levels in exhaled air and an improvement in FEV1 and AHR. The changes in FEV1 and AHR were significantly related to changes in markers in sputum and exhaled air (p<0.01 for each). These results suggest that baseline airway hyperresponsiveness can be predicted from the airway calibre but not from inflammatory parameters in sputum or exhaled air. In contrast, the reversible component of airway hyperresponsiveness appeared to be associated with the reduction in airway inflammation. PMID- 10706488 TI - Serological evidence of infection with Chlamydia pneumoniae is related to the severity of asthma. AB - There is evidence that infection with Chlamydia pneumoniae is associated with asthma of recent onset and that it can influence the severity of asthma. This has led to the suggestion that macrolide antibiotics may be useful in the treatment of asthma in subjects infected with C. pneumoniae. This study examined the association between immunoglobulin (Ig)G and IgA titres to C. pneumoniae and the severity of asthma. IgG and IgA antibodies to C. pneumoniae were measured in 619 subjects with asthma (18-60 yrs), using the microimmunofluoresence method. Subjects were asked about their use of asthma medicines, symptoms, previous hospitalization for asthma, smoking status and age of onset of asthma. In subjects with IgG titres of > or =1:64 and/or IgA titres > or =1:16 (n=212), spirometry was performed and peak expiratory flow rate (PEFR) and symptoms were recorded twice daily for 4 weeks on a diary card. The use of high dose inhaled steroids was associated with an increase of 74.1% in the titre of IgG antibodies (p=0.04) and an increase of 70.6% in the titre of IgA antibodies (p=0.0001) when compared with the use of low dose inhaled steroids. There was an inverse association between IgG antibodies and forced expiratory volume in one second (FEV1) as a percentage of predicted in those subjects with elevated IgG and/or IgA (p=0.04). In this group IgA antibodies were also associated with a higher daytime symptom score (p=0.04). Higher titres of antibodies to Chlamydia pneumoniae appears to be associated with markers of asthma severity. This raises the possibility that chronic infection with Chlamydia pneumoniae leads to an increase in the severity of asthma. Studies aimed at eradicating chronic infection with Chlamydia pneumoniae are necessary to determine whether or not this is the case. PMID- 10706489 TI - Differences in airway responsiveness to acetaldehyde and methacholine in asthma and chronic bronchitis. AB - Inhaled acetaldehyde may induce bronchoconstriction in asthmatic subjects and provides a new method to investigate airway responsiveness. The objective of the study was to determine whether acetaldehyde was a more specific stimulus than methacholine in differentiating asthma from chronic bronchitis with or without airflow limitation. Bronchial provocation challenges with methacholine and acetaldehyde were performed in 62 asthmatics and in 59 smokers with chronic bronchitis (32 with chronic bronchitis alone and 27 with chronic bronchitis and coexisting chronic obstructive pulmonary disease (COPD)). The response to both bronchoconstrictor agents was measured by the provocative concentration required to produce a 20% fall in forced expiratory volume in one second (FEV1; PC20). The two types of challenge yielded a similarly high level of sensitivity (100% for methacholine and 92% for acetaldehyde) in revealing airway hyperresponsiveness in asthma. However, bronchoprovocation with acetaldehyde yielded considerably greater specificity (95%) than bronchoprovocation with methacholine (24%) in separating asthma from chronic bronchitis. In subjects with asthma, methacholine and acetaldehyde responsiveness were weakly but significantly correlated (r=0.42, p=0.001) but no correlation was found between airway responsiveness to acetaldehyde and baseline FEV1 (r=0.13, p=0.33). These findings suggest that the demonstration of bronchoconstriction in response to acetaldehyde may be a more specific test than methacholine in the differentiation of asthma from chronic bronchitis. Furthermore, methacholine and acetaldehyde hyperresponsiveness are not reflecting the same pathophysiological process in the airways. PMID- 10706490 TI - Sudden-onset asthma exacerbations: clinical features, response to therapy, and 2 week follow-up. Multicenter Airway Research Collaboration (MARC) investigators. AB - Sudden-onset asthma exacerbations may have different triggers and responses to treatment than slower-onset exacerbations. The authors studied this hypothesis among patients with severe asthma exacerbations. The Multicenter Airway Research Collaboration prospectively enrolled patients presenting to 64 North American emergency departments with asthma exacerbations. Of 1,847 patients aged 18-54 yrs, 900 had severe exacerbations (peak expiratory flow rate (PEFR) <50% predicted or hospitalized without PEFR). These patients were divided into sudden onset (< or =3 h of symptoms) and slower-onset (>3 h of symptoms) groups. Fourteen per cent (95% confidence interval, 11-16%) of patients with severe asthma exacerbations had sudden-onset exacerbations. Sudden-onset patients were similar to slower-onset patients, except triggers of their exacerbations were more often respiratory allergens, exercise or psychosocial stress and less often respiratory infections. Sudden-onset patients were more likely to have used oral beta-agonists and salmeterol in the preceding 4 weeks. Although initial PEFRs and management were similar, sudden-onset patients had a greater improvement in PEFR (35 versus 28% p<0.001). Sudden-onset patients were less often discharged on systemic corticosteroids, but had similar 2-week relapse rates compared with slower-onset patients. Among patients presenting with severe asthma exacerbations, sudden-onset exacerbations had a different pattern of triggers and greater improvement with treatment than slower-onset exacerbations. PMID- 10706491 TI - Bronchial inflammation in acute bacterial exacerbations of chronic bronchitis: the role of leukotriene B4. AB - Neutrophils recruited to the airways in chronic obstructive pulmonary disease (COPD) are thought to mediate tissue destruction. Neutrophil recruitment is increased during bacterial exacerbations. The inflammatory process was studied in patients with an acute exacerbation of COPD in order to ascertain the role of leukotriene B4 (LTB4). The sputum of eight subjects with a bacterial exacerbation of COPD was analysed for neutrophil products (myeloperoxidase, elastase) and chemoattractants (interleukin-8 (IL-8) and LTB4). The contribution of LTB4 to the chemotactic activity of the sputum sol phase was determined using the LTB4 receptor antagonist LY293111. The concentrations of the serum acute phase proteins alpha1-proteinase inhibitor, alpha1-antichymotrypsin and C-reactive protein were measured. All patients received appropriate broad-spectrum antibiotic treatment for 7-14 days. Initially, the sputum myeloperoxidase activity was high, indicating neutrophil influx; this was associated with high levels of IL-8 and LTB4. All these concentrations fell with treatment (p<0.01). The chemotactic activity of the sputum was raised on presentation and fell with treatment (p<0.01). LTB4 contributed approximately 30% of the total chemotactic activity on presentation; this diminished with therapy. All acute phase proteins were raised on presentation and fell with therapy (p<0.01). These findings suggest that leukotriene B4 contributes to neutrophil influx into the airway in chronic obstructive pulmonary disease and may influence disease progression. PMID- 10706492 TI - Tumour necrosis factor-alpha gene promoter polymorphism in chronic obstructive pulmonary disease. AB - Tumour necrosis factor(TNF)-alpha levels are elevated in airways of patients with chronic obstructive pulmonary disease (COPD) and may contribute to its pathogenesis. A guanine to adenine substitution at position -308 of the TNF-alpha gene promoter (TNF1/2) has been associated with chronic bronchitis of various aetiologies in a Taiwanese population. The authors performed a study investigating association of the polymorphism with smoking-related COPD in Caucasians. Frequencies of TNF1/2 alleles in 86 Caucasians (52 males) with COPD were compared with 63 (52 males) asymptomatic smoker/exsmoker control subjects and a population control of 199 (99 males) blood donors. Genotyping was performed by the polymerase chain reaction-restriction fragment length polymorphism technique on genomic deoxyribonucleic acid (DNA) obtained from peripheral blood. There were no significant differences in TNF1/2 allele frequencies between groups: 0.85/0.15 in COPD, 0.85/0.15 in smoker control subjects, 0.83/0.17 in population control subjects. Within the COPD group there was no association of TNF1/2 alleles with indices of airflow obstruction (% predicted forced expiratory volume in one second (FEV1) and % predicted FEV1/vital capacity ratio) nor gas transfer (% predicted carbon monoxide transfer coefficient and % predicted carbon monoxide diffusing capacity of the lung). It is concluded that: 1) the tumour necrosis factor gene promoter allele does not influence the risk of developing chronic obstructive pulmonary disease in a Caucasian population of smokers; and 2) there is no association of the tumour necrosis factor gene promoter genotype with severity of airflow obstruction nor degree of emphysema in chronic obstructive pulmonary disease. PMID- 10706493 TI - Bronchoalveolar lavage fluid urea as a measure of pulmonary permeability in healthy smokers. AB - The effects of cigarette smoking on blood to airway pulmonary permeability to the low-molecular-weight solute urea were investigated, in an attempt to evaluate its use as a dilution marker for bronchoalveolar lavage (BAL) studies. Five healthy normal smokers who smoked a cigarette 10 min prior to undergoing a 3 x 60 mL bronchoalveolar lavage (BAL), and five nonsmokers who also underwent BAL but without cigarette smoke exposure were studied. Five minutes before bronchoscopy, 4 MBq 3H-water and 1 MBq 14C-urea were injected intravenously and biochemical urea assays and an indirect radiotracer method were used to evaluate permeability. It was shown that the smoking group had less urea in their BAL supernatants compared to nonsmokers the results using the radiotracer method being significant (p<0.005). Using both methods, it was shown that levels of urea increased in sequentially aspirated aliquots in both groups. The median directly assayed levels of urea in the smokers rose as follows: aliquot 1 0.05 micromol x mL(-1), (range 0.03-0.14), aliquot 2 0.10 micromol x mL(-1) (0.07-0.17), aliquot 3 0.12 micromol x mL(-1) (0.06-0.23) (p<0.05). This led to significantly increased calculated levels of epithelial lining fluid in the sequential aliquots (p<0.05). In addition, there were large but variable amounts of labelled water detected in both subject groups indicating a complex interaction between the BAL procedure and the circulation. Changing urea measurements during the bronchoalveolar lavage procedure confound the use of the urea (epithelial lining fluid) method for normalizing dilution factors. The use of epithelial lining fluid determinations in smokers ignores the additional and probably complex permeability changes. The present data suggest that acute exposure to cigarette smoke in smokers may decrease blood to airway permeability. PMID- 10706494 TI - Risk factors for pulmonary disease due to culture-positive M. tuberculosis or nontuberculous mycobacteria in South African gold miners. AB - The aim of this study was to determine risk factors for disease due to nontuberculous mycobacteria (NTM) compared to those due to Mycobacterium tuberculosis in South African gold miners with pulmonary mycobacterial disease. A case/control study comparing tuberculosis and NTM cases amongst all patients with a positive sputum mycobacterial culture in 1995 was carried out. The 51 cases of disease due to NTM and 425 tuberculosis cases were similar with regard to age, education, home region, smoking habits and percentage of CD4 cells. After adjustment for confounders, those with NTM were more likely to have had previous tuberculosis treatment (odds ratio (OR) 3.61; 95% confidence interval (CI) 1.9 6.9), have worked longer underground (p-value for trend=0.05) or have evidence of silicosis (OR 12.6; 95% CI 2.2-71) and were less likely to drink regularly (OR 0.12; 95% CI 0.02-0.93) than patients with tuberculosis. In patients with disease due to NTM, 35.3% were human immunodeficiency virus-positive compared with 48.8% of tuberculosis patients (p=0.2) and an estimated 21% overall in the mines at the time of the study. Previous tuberculosis treatment, silicosis and duration of underground work are even more strongly associated with disease due to nontuberculous mycobacteria than with tuberculosis. Attempts to reduce the incidence of all pulmonary mycobacterial disease in this community should address recognized risk factors and ensure that those with tuberculosis are diagnosed, treated and cured. PMID- 10706495 TI - Increased calcium influx in a monocytic cell line on exposure to ultrafine carbon black. AB - Ultrafine particles have been shown to induce pro-inflammatory effects both in vivo and in vitro. Increased expression of pro-inflammatory genes probably requires the activation of specific transcription factors such as nuclear factor kappa B (NF-kappaB) via a number of possible pathways including Ca2+ and reactive oxygen species. The fluorescent dye fura 2, was used to measure cytosolic Ca2+ in the human monocytic cell line, Monomac 6 on exposure to 66 microg x mL(-1) of either ultrafine carbon black (ufCB; diameter 14 nm), carbon black (CB; diameter 260 nm), quartz (diameter 1.45 microm), or medium alone. UfCB but not fine CB induced a 1.6-fold increase (p<0.01) in the resting cytosolic Ca2+ concentration of Monomac 6 cells. In addition ufCB induced a 2.6-fold increase (p<0.001) in the response to the endoplasmic reticulum Ca2+- adenosine triphosphatase (ATPase) inhibitor, thapsigargin, suggesting the Ca2+ release-activated Ca2+ current across the plasma membrane was enhanced. This response was inhibited by the removal of extracellular Ca2+ and by the Ca2+ channel blocker, verapamil. In addition, ufCB stimulated the entry of extracellular Mn2+. Finally, the antioxidants mannitol and nacystelin both inhibited the effects of ufCB on the response to thapsigargin. These data suggest that ultrafine carbon black particles stimulated an increase in cytosolic Ca2+, possibly through the entry of extracellular Ca2+ via Ca2+ channels in the plasma membrane. The particles may in part activate the opening of Ca2+ channels via a mechanism involving reactive oxygen species. PMID- 10706496 TI - Tuberculosis among health care workers during three recent decades. AB - Some studies have found that health care workers have an increased risk of tuberculosis, whereas other studies have reported the opposite. This study examined data of Finnish health care workers (HCWs) for the incidence of tuberculosis disease. Cases of occupational tuberculosis in Finland were analysed over a period of 30 yrs (1966-1995). Control subjects were all other incident tuberculosis cases at working age. The material thus obtained included 658 people with notified occupational tuberculosis, and 56,146 control cases. The authors studied incidence, and age specific rate, as well as their trends. The incidence of tuberculosis among health care workers decreased from 57.9 to 6.1 per 100,000 and the corresponding figures among control subjects decreased from 156.8 to 9.1 per 100,000. The overall risk in health care workers was lower than in the general population throughout the study period. Analysis of age specific rates revealed no age group at increased risk. The reasons are multifactorial, among them the successful tuberculosis programme, and the lack of impact of such risk factors as immigration, human immunodeficiency virus infection and drug resistance; high coverage of bacille Calmette-Guerin vaccination may also be a factor, although it is difficult to assess. PMID- 10706497 TI - Isotonic mechanics of a pharyngeal dilator muscle and diaphragm in the rat before and after fatigue. AB - Pharyngeal and diaphragm muscles contract and relax in synergy, which is why it was decided to compare their mechanical performance throughout the overall load continuum. The effects of fatigue were also studied. The isotonic mechanics of rat sternohyoid (SH; n=10) and diaphragm (D; n=10) were investigated in vitro. Force and length were measured in muscles contracting from zero load up to isometry. Maximum isometric tension (Pmax), peak mechanical work (Wmax), maximum unloaded shortening velocity (vzL) and mechanical efficiency (eff(max)) were recorded. Data were obtained both at baseline and after fatigue. SH muscles had a lower Pmax (96.0+/-13.7 versus 119.5+/-22.7 mN x mm(-2); p<0.05), a lower Wmax (5.5+/-1.2 versus 8.0+/-2.1 mJ x g(-1); p<0.01), a lower eff(max) (56.0+/-6.9 versus 62.6+/-5.8%; p<0.05) and a higher vzL (4.8+/-0.4 versus 3.4+/-0.4 initial length (L0) x s(-1); p<0.001) than D muscles. Wmax occurred at a higher relative load in SH (40% Pmax) than in D (30% Pmax). Fatigue did not modify eff(max) in SH muscles, whereas it significantly improved eff(max) in D muscles. These findings suggest that under control conditions, economy of force generation was less efficient in sternohyoid than in diaphragm muscles. Fatigue in sternohyoid muscles induced unfavourable mechanical behaviour. This may partly explain pharyngeal dilator muscle failure in the presence of increased loads. Whether these findings are relevant to human sleep apnoea syndrome has yet to be determined. PMID- 10706498 TI - Noninvasive assessment of respiratory resistance in severe chronic respiratory patients with nasal CPAP. AB - Noninvasive measurement of respiratory resistance during nasal ventilatory support could be useful to assess the mechanical status of the patient and to optimize the ventilator settings. The aim was to investigate whether the forced oscillation technique (FOT) applied through a nasal mask allows reliable noninvasive estimation of respiratory resistance (Rrs) in patients with severe chronic respiratory disease. FOT Rrs (5 Hz) and lung resistance (R(L)) measured simultaneously from spontaneous breathing signals by an oesophageal balloon were compared in eight patients with chronic obstructive pulmonary disease and in six patients with a restrictive ventilatory defect due to chest wall disease. Measurements were performed in sitting and supine postures during application of nasal continuous positive airway pressure (CPAP): 4, 8 and 12 cmH2O in obstructive patients and 4 cmH2O in restrictive patients. In the restrictive patients Rrs and R(L) (in cmH2O x s x L(-1)) were virtually coincident: mean+/ SD, 12.6+/-6.1 and 11.6+/-6.6 (r=0.96) in sitting and 9.7+/-3.1 and 10.2+/-3.3 (r=0.92) in supine posture, respectively. In the obstructive patients (CPAP = 4 cmH2O), Rrs slightly underestimated R(L): mean+/-SD, 11.5+/-5.9 and 14.4+/-16.8 (r=0.92) in sitting and 15.0+/-9.8 and 21.1+/-12.6 (r=0.96) in supine posture, respectively. Similar results were found at CPAP = 8 and 12 cmH2O. The results obtained in patients with resistance values in the range typically found in nasal ventilatory support suggest that forced oscillation technique could be valuable to noninvasively estimate a patient's respiratory mechanical resistance. PMID- 10706499 TI - Endothelin-1 plasma levels are not elevated in patients with obstructive sleep apnoea. AB - Endothelin-1 (ET-1), a potent vasoconstrictor, is released mainly by vascular endothelial cells under the influence of hypoxia and other stimuli. ET-1 is related to endothelial dysfunction, as well as arterial and pulmonary hypertension, all of which are thought to be associated with obstructive sleep apnoea (OSA). This study evaluated venous plasma concentrations of ET-1 and noradrenaline and 24-h systemic blood pressure in 29 patients with OSA (age=56.9+/-1.6 yrs; body mass index=29.5+/-0.7 kg x m2 (mean+/-SEM)). Blood samples were taken in the morning, evening and during sleep. In the same way, the patients were assessed during a night of continuous positive airway pressure (CPAP) and after 13.9+/-1.4 months while still on CPAP. ET-1 levels were compared to those of control subjects, who were selected from in- and outpatients and were matched to patients for age, sex, presence of arterial hypertension and coronary artery disease. ET-1 plasma levels were not elevated in the patients compared to the controls (41.6+/-2.2 and 44.9+/-1.3 pg x mL(-1), respectively, p=0.20). The ET-1 concentration did not change significantly, neither during sleep nor in the first night on CPAP therapy, nor under long-term treatment with CPAP. ET-1 neither correlated to the severity of OSA nor to that of systemic hypertension. The results suggest that endothelin-1 does not play a crucial role in the pathophysiology of obstructive sleep apnoea. PMID- 10706500 TI - Mortality of sleep apnoea patients treated by nasal continuous positive airway pressure registered in the ANTADIR observatory. Association Nationale pour le Traitement A Domicile de l'Insuffisance Respiratoire chronique. AB - The aim of this study was to examine risk factors for and causes of mortality in patients with obstructive sleep apnoea syndrome (OSAS) treated by nasal continuous positive airway pressure (CPAP). Univariate and multivariate analyses of the data on patients registered in the Association Nationale pour le Traitement A Domicile de l'Insuffisance Respiratoire chronique (ANTADIR) Observatory between January 1, 1985 and December 31, 1993 and followed to January 1, 1996. Survival ratios were compared to those of the French population. A case control study compared patients who died with patients of the same age and sex, in the same Regional Association, who were equipped with CPAP at the same time. Five-thousand-six-hundred-and-sixty-nine patients had CPAP treatment. Two-hundred and-seventy-six had died. One-hundred-and-twenty-four deaths were examined and compared to 123 control subjects. Overall mortality was the same as the general French population. Independent risk factors for death were age, oxygen tension in arterial blood (Pa,O2) and forced expiratory volume in one second (FEV1) (per cent predicted). In the case-control study independent risk factors for death in the past history were cardiac arrhythmia with an odds ratio (OR) of 2.8 (95% confidence interval (CI) 1.1-7.2), respiratory disorders (OR 2.8; CI 1.6-4.9) ischaemic events (OR 2.2; CI 1.2-4.2), neurological and psychiatric disorders (OR 2.4; CI 1.1-5.4). A significant excess of cardiovascular deaths and an excess of deaths from accidents and poisonings was found. In conclusion, patients die on therapy predominantly from cardiovascular causes but many have a past history of cardiovascular conditions. Compliance with treatment may be important for survival. Continuous positive airway pressure is an effective therapy for obstructive sleep apnoea syndrome but older patients with reduced spirometry and hypoxaemia may need more attention paid to these aspects of their condition. PMID- 10706501 TI - Diaphragmatic dysfunction and dyspnoea in amyotrophic lateral sclerosis. AB - Amyotrophic lateral sclerosis (ALS) is a progressive disorder of unknown origin. Respiratory involvement is the principal cause of death, and dyspnoea is a major source of discomfort. In this study, diaphragm function is described and its relationship with dyspnoea examined in 48 ALS patients (32 male, age 26-80 yrs). The detailed neurological and respiratory evaluation (clinical examination, pulmonary function tests, static pressures, mouth twitch pressures (Pm,t), electromyographic responses to phrenic nerve stimulation and cortical magnetic stimulation were analysed after stratification according to dyspnoea. Dyspnoeic (group I) and nondyspnoeic (group II) patients were similar, bulbar signs being more frequent in group I. Vital capacity was lower in group I (mean+/-SD 67.9+/ 22.7 versus 87.9+/-15.6% of the predicted value, p=0.0028), as were maximal static inspiratory pressure (41+/-24 versus 60+/-27% pred, p=0.0242) maximal static inspiratory pressure (18+/-11 versus 32+/-14% pred, p=0.0042), and Pm,t (3.71+/-2.5 versus 7.26+/-3.45 cmH2O, p=0.0011). Abdominal (Abd) paradox and respiratory pulse were frequent in group I (15 of 25 and 14 of 25) but absent or rare in group II (0 of 23 and four of 23) (p<0.05). The electromyographic responses to phrenic and cortical stimulation were generally abnormal in group I but subnormal in group II. Multivariate analysis selected only signs of diaphragm dysfunction (namely, Abd paradox and abnormal electromyographic responses) as significant predictors of dyspnoea. It is concluded that dyspnoea in amyotrophic lateral sclerosis patients should prompt diaphragm function tests. PMID- 10706502 TI - Inaccuracy of tidal volume delivered by home mechanical ventilators. AB - Ideally, the inspired (tidal) volume (V(T)) provided by a volume-controlled ventilation device should not change when the pressure imposed on the ventilator varies. A bench study evaluation of V(T) versus pressure was performed on 10 commercially available devices. The difference between the desired V(T) and the observed V(T) reached 100 mL for some devices when inspiratory resistance was at its lowest, rising to 150 mL when inspiratory resistance was increased to obtain peak airway pressure of 60 cmH2O. The present data indicate that some home ventilators are inaccurate in delivering the preset tidal volume when the pressure imposed on the ventilator is increased to simulate high airway resistance. PMID- 10706503 TI - Phosphodiesterase and cyclic adenosine monophosphate-dependent inhibition of T lymphocyte chemotaxis. AB - There is abundant evidence for T-lymphocyte recruitment into the airways in allergic inflammatory responses. This study has tested the hypothesis that T-cell chemotaxis induced by platelet-activating factor (PAF) and human recombinant interleukin-8 (hrIL-8) can be attenuated by inhibition of phosphodiesterase activity and raised intracellular 3',5'-cyclic adenosine monophosphate (cAMP) levels. This study used theophylline, a nonselective phosphodiesterase (PDE) inhibitor, and rolipram, a selective PDE4 inhibitor, to study the effect of PDE inhibition on T-cell chemotaxis. The beta2-adrenoceptor agonist, salbutamol, the adenylyl cyclase activator, forskolin, and the cAMP analogue, dibutyryl cAMP (db cAMP), were used to demonstrate a role for raised cAMP levels. T-cells were obtained from 10 atopic asthmatics, and the phenotype of migrating cells was examined by flow cytometry. Theophylline caused an inhibition of both PAF-and hrIL-8-induced chemotaxis (mean+/-SEM maximum inhibition at 1 mM: 73+/-4% and 48+/-8% for hrIL-8 and PAF, respectively) that was not specific for the CD4+, CD8+, CD45RO+ or CD45RA+ T-cell subsets. T-cell chemotaxis was more sensitive to treatment with rolipram whose effect was already significant from 0.1 microM on hrIL-8-induced chemotaxis. Both a low concentration of salbutamol (0.1 mM) and forskolin (10 microM) potentiated the inhibitory effect of a low concentration of theophylline (25 microM) on responses to PAF but not to hrIL-8. Finally, T-cell chemotaxis was also inhibited by db-cAMP. It is concluded that attenuation of T cell chemotaxis to two chemoattractants of relevance to asthma pathogenesis can be achieved via phosphodiesterase inhibition and increased intracellular 3', 5' cyclic monophosphate using drugs active on cyclic nucleotide phosphodiesterase. This action may explain the anti-inflammatory effects of theophylline and related drugs in asthma. PMID- 10706504 TI - GM-CSF and GM-CSF beta c receptor in adult patients with pulmonary alveolar proteinosis. AB - Pulmonary alveolar proteinosis (PAP) is a rare disorder of unknown origin characterized by alveolar fillings with periodic acid-Schiff (PAS)-positive material mainly consisting of phospholipids. Mice defective in the granulocyte macrophage colony-stimulating factor (GM-CSF) gene or the GM-CSF/interleukin (IL) 3/IL-5-receptor common beta chain (beta c) demonstrate a pathology resembling PAP. A recent study revealed defects in the beta c chain of the GM-CSF receptor in four out of eight paediatric patients. This study investigates the role of the GM-CSF coding region and components of the GM-CSF receptor in adult patients. Four adult patients with proven PAP were analysed for GM-CSF and GM-CSF-beta c receptor in regard to protein level, messenger ribonucleic acid (mRNA) expression and sequence composition. None of the adult patients displayed the mutation at position 1,835 of the beta c-receptor previously described in paediatric patients. Expression of the beta c receptor was found to be normal on the surface of peripheral blood cells. In three out of four patients GM-CSF release from blood cells failed to respond adequately to lipopolysaccharide (LPS). In one of these patients a heterozygous mutation was found in the GM-CSF complementary deoxyribonucleic acid (cDNA) from thymine (T) to cytosine (C) at position 382 of the published sequence putatively causing a change in the protein from isoleucine to threonine at position 117. This study indicates that the mutation of the beta chain receptors found in some of the paediatric patients suffering from pulmonary alveolar proteinosis is not a common problem in adult patients. PMID- 10706505 TI - Adhesion molecule expression on epithelial cells infected with respiratory syncytial virus. AB - Respiratory epithelium is both a target and an effector of airway inflammation. Adhesion molecules on epithelium play an important role in a variety of airway diseases. Respiratory syncytial virus (RSV) is the most important pathogen for airway diseases in infants. The expression of adhesion molecules on epithelium in RSV infection, however, is unclear. The expression of selected adhesion molecules and major histocompatibility complex (MHC) class I and II antigens on a human alveolar type II epithelial cell line (A549) infected with RSV was investigated by means of flow cytometry and immunocytochemistry. The results showed that intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) were expressed on A549 cells at a low level. E-cadherin and MHC class I antigen were constitutively expressed on the cells. RSV infection of A549 cells significantly upregulated the expression of ICAM-1, VCAM-1 and MHC class I and II antigens on these cells. RSV infection also altered the expression of E-cadherin on A549 cells. Immunostaining showed that E-cadherin was mainly upregulated around or in RSV-induced giant cells. These data suggest that respiratory syncytial virus infection of respiratory epithelial cells enhances the expression of adhesion molecules and major histocompatibility complex antigens. These changes may play an important role in the pathophysiology of respiratory syncytial virus disease. PMID- 10706506 TI - Inflammatory cells as well as epithelial cells in nasal polyps express vascular endothelial growth factor. AB - In nasal polyps (NPs), locally secreted growth factors are involved in the remodelling of the epithelium and extracellular matrix but little is known concerning vessel remodelling. The in situ expression of vascular endothelial growth factor (VEGF) in NPs and control nasal mucosa (CM) were evaluated and in vitro secretion of VEGF from primary human cultures of nasal epithelial cells (HNECs) was quantified. VEGF expression was evaluated in NP (n=14) and CM (n=6) after immunolabelling. In supernatants from HNECs cultured at air/liquid interface, VEGF was quantified by immunoassay, under baseline conditions and after transforming growth factor-beta1 (TGF-beta1) stimulation. In HNEC lysates, VEGF and VEGF messenger ribonucleic acid (mRNA) were detected using Western blot analysis and reverse transcriptase polymerase chain reaction respectively. VEGF positivity was more frequent in inflammatory cells in NPs (14 of 14) than in CM (three of six) (p<0.05) and in the epithelium in NPs (six of 14) than in CM (two of six) (nonsignificant). Under baseline conditions, the VEGF concentration in HNEC culture medium increased from day 2 to 4, then decreased and became undetectable. VEGF concentrations increased significantly after TGF-beta1 stimulation. In HNEC lysates, VEGF and VEGF mRNA were detected on days 4 and 14 of culture. It was concluded that vascular endothelial growth factor is intensely expressed in situ in nasal polyps, mainly in inflammatory cells but also in epithelial cells. Human nasal epithelial cells are able to secrete in vitro vascular endothelial growth factor. Transforming growth factor-beta1 upregulates this secretion. This suggests that vascular endothelial growth factor, inducing oedema and angiogenesis, could be involved in the pathogenesis of nasal polyps. PMID- 10706507 TI - Methotrexate pneumonitis: review of the literature and histopathological findings in nine patients. AB - Pneumonitis is a serious and unpredictable side-effect of treatment with methotrexate (MTX) that may become life-threatening. The clinical and histological features of nine cases of MTX pneumonitis are reported and the literature reviewed. The typical clinical symptoms include progressive shortness of breath and cough, often associated with fever. Hypoxaemia and tachypnoea are always present and crackles are frequently audible. Chest radiography reveals a diffuse interstitial or mixed interstitial and alveolar infiltrate, with a predilection for the lower lung fields. Pulmonary function tests show a restrictive pattern with diminished diffusion capacity. Lung biopsy reveals cellular interstitial infiltrates, granulomas or a diffuse alveolar damage pattern accompanied by perivascular inflammation. These clinical and pathological findings are not specific to MTX pneumonitis and can be seen with other drug induced lung toxicities. It is important that all patients receiving methotrexate be educated concerning this potential adverse reaction and instructed to contact their physicians should significant new pulmonary symptoms develop while undergoing therapy. If methotrexate pneumonitis is suspected, methotrexate should be discontinued, supportive measures instituted and careful examination for different causes of respiratory distress conducted. PMID- 10706508 TI - Development of tuberculin reactivity and sensitization to M. scrofulaceum and M. fortuitum in children BCG-vaccinated at birth. AB - Since the incidence of tuberculosis is steadily declining in Finland and infections by environmental mycobacteria may be increasing, the aim of the present study was to evaluate the development of tuberculin reactivity and sensitization to environmental mycobacteria. Healthy Finnish schoolchildren aged 10.4-12.4 yrs (n=201) were tested with tuberculin purified protein derivative RT23, Mycobacterium scrofulaceum RS95 and M. fortuitum RS20 sensitins. The same children had been previously tested with the same antigens and methods at the age of 4-6 yrs in 1989. Rapid waning of tuberculin reactivity and decrease in sensitization to environmental mycobacteria were observed between 4-6 yrs. Both tuberculin and sensitin skin reaction sizes decreased significantly over the 6 yrs period. The mean tuberculin skin reaction size was 3.2 mm in diameter, which was significantly (p<0.001) smaller than the mean induration size (4.8 mm) at the age of 4-6 yrs. Similarly, the mean skin reaction sizes to M. scrofulaceum and M. fortuitum sensitins were 3.4 and 1.7 mm, respectively, which were significantly (p<0.001) smaller than 6 yrs earlier (mean 4.5 and 3.1 mm). The number of zero reactions to all antigens increased significantly during the follow-up period. Contacts with pets or farm animals were associated with larger reactions. In contrast, children suffering from allergic symptoms had smaller reactions. Contacts with mycobacteria, either with Mycobacterium tuberculosis or environmental mycobacteria, seem to be too rare to maintain tuberculin responsiveness and a high sensitivity to other mycobacteria. Different bacille Calmette-Guerin vaccine products and dosages used, the declining incidence of tuberculosis and geographical factors, which can influence environmental mycobacterial exposure, may explain the disparity between the present and previous Finnish studies. PMID- 10706509 TI - No objective benefit from steroids inhaled via a spacer in infants recovering from bronchiolitis. AB - A double-blind randomized placebo-controlled trial was conducted to investigate the efficacy of 3 months' inhaled steroids delivered via a spacer device with face mask attachment to infants recovering from bronchiolitis. Forty-eight previously healthy infants recovering from their first documented episode of acute bronchiolitis were randomized to receive 150 microg fluticasone propionate (FP) b.i.d. or placebo delivered via the Babyhaler spacer. Longitudinal assessments were performed on seven occasions over 1 yr based on symptom diaries and health records, clinical examinations, overnight cough recordings and oxygen saturation readings. Lung function was measured 6 months after hospital discharge. Forty-three infants completed the trial (FP 21, placebo 22). There were no significant differences in the three objective end-points measured, recorded night cough, oxygen saturation and lung function test results. Symptom scores were low in both the FP and placebo groups with the absence of (0) or mild (1) symptoms > or =90% of the trial days. No statistical differences in symptom frequency, use of rescue respiratory medications or hospital admissions between treatment groups were found throughout the trial or follow-up periods. In conclusion, the use of inhaled fluticasone propionate in infants recovering from acute bronchiolitis cannot be recommended. PMID- 10706510 TI - Thrombotic risk factors in pulmonary hypertension. AB - Thrombotic lesions are consistently observed in chronic thromboembolic pulmonary hypertension (CTEPH) and frequently found in primary pulmonary hypertension (PPH). It remains unknown, however, whether thrombosis is related to defects of the antithrombotic pathway or to previous vascular injury. This study therefore analysed the frequency of both hereditary and acquired thrombotic risk factors in CTEPH and PPH. One hundred and forty-seven consecutive patients with CTEPH investigated in the author's institution were compared to 99 consecutive patients with PPH. In 116 CTEPH patients and 83 PPH patients, phospholipid-dependent antibodies (antiphospholipid antibodies and lupus anticoagulant) were analysed by both immunological and clotting assays. In patients enrolled since 1994 (46 CTEPH and 64 PPH), hereditary thrombotic risk factors were also determined. Antithrombin, protein C and protein S activities were measured by functional assays. Mutations of factor V and factor II were identified by polymerase chain reaction. The prevalence of hereditary thrombotic risk factors was not increased in patients with either PPH or CTEPH. In contrast, a high frequency of phospholipid-dependent antibodies was observed in PPH (10%) and more notably in CTEPH (20%). Moreover, in PPH, antibodies were present only in low titre whereas in CTEPH, half of the patients with antiphospholipid antibodies had high titres. In addition, in CTEPH all but one of the patients with lupus anticoagulant also had antiphospholipid antibodies. The most striking finding of this study was the high prevalence of phospholipid-dependent antibodies but their clinical relevance appears to be different in primary pulmonary hypertension and chronic thromboembolic pulmonary hypertension. In primary pulmonary hypertension, these antibodies in low titre probably reflect endothelial dysfunction. In contrast, in chronic thromboembolic pulmonary hypertension the presence of antibodies in high titre associated with lupus anticoagulant, underlines the role of thrombosis in the pathogenesis of this condition. PMID- 10706511 TI - Modulated vasodilator responses to natriuretic peptides in rats exposed to chronic hypoxia. AB - Natriuretic peptides (NPs), such as atrial natriuretic peptide (ANP), C-type natriuretic peptide (CNP), and adrenomedullin (ADM), are endogenous vasodilators acting via specific receptors. This study addressed the question of how pulmonary artery (PA) responses to these peptides and the gene expression of their receptors are modulated in pulmonary hypertension rat models exposed to chronic hypoxia. In this study, isometric tension was measured in PA rings exposed to these NPs and 8-bromoguanosine 3', 5'-cyclic monophosphate (8-bromo-cGMP). It was compared with messenger ribonucleic acid (mRNA) levels of NP-A and -B receptors, which bind to ANP and CNP, respectively, as determined by ribonuclease (RNase) protection assay. Chronic hypoxia increased the maximal relaxation elicited by ANP, but the responses to CNP and 8-bromo-cGMP were unchanged. Chronic hypoxia did not change NP-A and -B receptor mRNA levels. The results showed that pulmonary artery response to atrial natriuretic peptide is selectively enhanced, possibly via a post-transcriptional modulation of its receptor in chronically hypoxia rats. These pharmacological characteristics of atrial natriuretic peptide are consistent with the hypothesis that the atrial natriuretic peptide system is protective against the progression of pulmonary hypertension. PMID- 10706512 TI - Diagnosis of pulmonary aspergillosis using optical brighteners. AB - Invasive pulmonary aspergillosis (IPA) is increasingly recognized especially in immunocompromised patients, but early diagnosis remains a problem. Fungal elements in clinical specimens can be directly stained with an optical brightener. The high intensity of the elicited fluorescence allows for rapid and reliable microscopic screening. In the present study the authors aimed to validate this method. All specimens from bronchial secretions or pleural fluid suspected of mycosis (n=94) and all bronchoalveolar lavages (n=439) were prospectively evaluated by culture and staining with the optical brightener Blankophor-P-Flussig. IPA was diagnosed for 17 specimens from 13 patients, using a combination of clinical, culture and radiological data, and by biopsy (n=3) or autopsy (n=3). The overall incidence of IPA was 3.3%. Nine of the 13 patients with IPA died (mortality=69%). Staining with the optical brightener and consecutive microscopic screening took 9+/-3 min. For the diagnosis of invasive aspergillosis, sensitivity was 0.88 and specificity was 0.99. Using culture, sensitivity was 0.76 and specificity was 0.99. In conclusion, direct examination of clinical specimens using the optical brightener has a high diagnostic potential for the diagnosis of invasive pulmonary aspergillosis. The reliability, simplicity and speed of the method render it suitable for routine diagnostic work. PMID- 10706513 TI - Acute interstitial pneumonia. AB - The term "acute interstitial pneumonia" (AIP) describes an idiopathic clinicopathological condition, characterized clinically by an interstitial lung disease causing rapid onset of respiratory failure, which is distinguishable from the other more chronic forms of interstitial pneumonia. It is synonymous with Hamman-Rich syndrome, occurring in patients without pre-existing lung disease. The histopathological findings are those of diffuse alveolar damage. AIP radiologically and physiologically resembles acute respiratory distress syndrome (ARDS) and is considered to represent the small subset of patients with idiopathic ARDS. It is frequently confused with other clinical entities characterized by rapidly progressive interstitial pneumonia, especially secondary acute interstitial pneumonia, acute exacerbations and accelerated forms of cryptogenic fibrosing alveolitis . Furthermore, many authors use the above terms, both erroneously and interchangeably. It has a grave prognosis with >70% mortality in 3 months, despite mechanical ventilation. This review aims to clarify the relative clinical and pathological issues and terminology. PMID- 10706514 TI - Opacity over the left hemithorax after abdominal aortic aneurysm repair. PMID- 10706515 TI - Pulmonary veno-occlusive disease in pulmonary Langerhans' cell granulomatosis. AB - This report describes unusual clinical and pathological findings in a 29-yr-old female with pulmonary Langerhans' cell granulomatosis (LCG). During a 7-yr clinical course her condition deteriorated despite corticosteroid therapy, and she died of respiratory failure and pulmonary hypertension. At autopsy, there were widespread pulmonary veno-occlusive disease (PVOD) lesions as well as abundant advanced and healed lesions of pulmonary LCG composed of multiple cysts and stellate fibrosis. The present case demonstrates that pulmonary Langerhans' cell granulomatosis should be considered as a possible cause of pulmonary veno occlusive disease. PMID- 10706516 TI - A method for bronchoscopic evaluation of salivary aspiration in a disabled child. AB - Chronic aspiration is a cause of life-threatening respiratory complications and repeated hospital admissions, particularly in children with neurological disabilities. Determining the source of aspiration is important for optimizing treatment. This report describes a simple technique to demonstrate salivary aspiration during fibreoptic bronchoscopy. A child with a history of recurrent pneumonia was given methylene blue orally 2 h prior to fibreoptic bronchoscopy. Bronchoscopy was carried out through a laryngeal mask airway under inhalational anaesthesia. The stained saliva was seen to be pooling in the valleculae and then running down the trachea into the bronchi, confirming salivary aspiration. PMID- 10706517 TI - Pulmonary alveolar proteinosis: two contrasting cases. AB - Pulmonary alveolar proteinosis is a rare condition characterized by the abnormal accumulation of surfactant-like material within the alveolar spaces and distal bronchioles. Two cases with contrasting modes of presentation, course, and response to therapeutic whole lung lavage are described. Both cases were in hypoxaemic respiratory failure at the time the definitive diagnosis was made, and in both cases the diagnosis was made by segmental bronchoalveolar lavage following negative open lung biopsy. In neither was an underlying causative organism or agent identified. In one case the alveolar proteinosis developed in late pregnancy, a presentation that is previously unreported. Clinical improvement in this case required repeated whole lung lavages and was accompanied by a trend towards normalization of the ratios of surfactant protein-A and surfactant protein-B to disaturated phospholipid, ratios which may be useful as prognostic indicators. The response to therapeutic lavage was markedly different in the two cases, and it is postulated that this may relate to the fact that alveolar proteinosis is a heterogeneous disease and that the course and response to treatment may relate in part to the specific composition of the abnormal proteinaceous fluid. PMID- 10706518 TI - Haemopneumothorax from congenital cystic adenomatoid malformation in a cryptorchidism patient. AB - Congenital cystic adenomatoid malformation (CCAM) of the lung is an uncommon congenital anomaly, especially in young adults. This study reports an 18-yr-old male with CCAM involving the right upper lobe, who presented with a moderate spontaneous haemopneumothorax initially. The patient also had bilateral abdominal cryptorchidism which required surgical treatment earlier in childhood. The chest radiographs and contrast-enhanced computed tomographic scan of the chest showed a multicystic lesion with air-fluid levels in the right upper lung. The right upper lobe was resected through a posterolateral thoracotomy. Histological examination confirmed the diagnosis of CCAM. To the authors' knowledge, congenital cystic adenomatoid malformation presenting with spontaneous haemopneumothorax and haemoptysis has never been described in the literature. PMID- 10706519 TI - Hyperbaric oxygen treatment. PMID- 10706520 TI - Drug-polymer conjugates containing acid-cleavable bonds. AB - Drug-polymer conjugates are potential candidates for the selective delivery of anticancer agents to tumor tissue. Incorporating acid-sensitive bonds between the drug and the polymer is an attractive approach because it ensures effective release of the polymer-bound drug at the tumor site. This release is either extracellular, resulting from the slightly acidic pH in tumor tissue, or intracellular, in acidic endosomes or lysosomes after cellular uptake of the drug polymer conjugate. This paper reviews acid-sensitive drug-polymer conjugates developed during the past 20 years and outlines aspects for further development in this research field. PMID- 10706521 TI - Polyrotaxanes: synthesis, structure, and potential in drug delivery. AB - This article reviews the potential of polyrotaxanes in drug delivery with the historical background of polyrotaxane syntheses. Pseudopolyrotaxanes and polyrotaxanes, including classifications, synthetic methods, structures and physical properties are discussed in the first section. The second section provides our concept of drug carriers using drug-polyrotaxane conjugates in comparison with conventional drug-polymer conjugates. The third and fourth sections describe the synthetic method for biodegradable polyrotaxanes, the conjugation with drugs, and their association under physiological conditions. The fifth section discusses other possibilities for the polyrotaxanes such as drug penetration enhancers. These studies suggest the potential of polyrotaxanes in pharmaceutical applications. PMID- 10706522 TI - Personal exposure to JP-8 jet fuel vapors and exhaust at air force bases. AB - JP-8 jet fuel (similar to commercial/international jet A-1 fuel) is the standard military fuel for all types of vehicles, including the U.S. Air Force aircraft inventory. As such, JP-8 presents the most common chemical exposure in the Air Force, particularly for flight and ground crew personnel during preflight operations and for maintenance personnel performing routine tasks. Personal exposure at an Air Force base occurs through occupational exposure for personnel involved with fuel and aircraft handling and/or through incidental exposure, primarily through inhalation of ambient fuel vapors. Because JP-8 is less volatile than its predecessor fuel (JP-4), contact with liquid fuel on skin and clothing may result in prolonged exposure. The slowly evaporating JP-8 fuel tends to linger on exposed personnel during their interaction with their previously unexposed colleagues. To begin to assess the relative exposures, we made ambient air measurements and used recently developed methods for collecting exhaled breath in special containers. We then analyzed for certain volatile marker compounds for JP-8, as well as for some aromatic hydrocarbons (especially benzene) that are related to long-term health risks. Ambient samples were collected by using compact, battery-operated, personal whole-air samplers that have recently been developed as commercial products; breath samples were collected using our single-breath canister method that uses 1-L canisters fitted with valves and small disposable breathing tubes. We collected breath samples from various groups of Air Force personnel and found a demonstrable JP-8 exposure for all subjects, ranging from slight elevations as compared to a control cohort to > 100 [mutilpe] the control values. This work suggests that further studies should be performed on specific issues to obtain pertinent exposure data. The data can be applied to assessments of health outcomes and to recommendations for changes in the use of personal protective equipment that optimize risk reduction without undue impact on a mission. PMID- 10706523 TI - Economic activity and congenital anomalies: an ecologic study in Argentina. ECLAMC ECOTERAT Group. AB - In this study, we analyze the association between industrial activity and the occurrence of 34 congenital anomalies. We selected 21 counties in Argentina during 1982-1994 and examined a total of 614,796 births in these counties in consecutive series. We used the International Standard Industrial Classification of All Economic Activities (United Nations, 1968) as an indicator of exposure to 80 specific industrial activities. Incidence rate ratios for each congenital anomaly were adjusted by the socioeconomic level of the county according to a census index of social deprivation. For a given exposure/anomaly association to be considered as significant and relevant, the exposure had to be a statistically significant risk for the occurrence of the anomaly and an increase in the birth prevalence rate of the congenital anomaly type involved had to be observed in those counties where the putative causal activity was being performed. Significant associations (p < 0.01) were identified between textile industry and anencephaly, and between the manufacture of engines and turbines and microcephaly. These observations are consistent with previous reports on occupational exposure, and their further investigation by means of case-control studies is recommended. PMID- 10706524 TI - Associations of tibial lead levels with BsmI polymorphisms in the vitamin D receptor in former organolead manufacturing workers. AB - We evaluated associations of tibial lead levels with polymorphisms in the vitamin D receptor (VDR) in 504 former organolead manufacturing workers with past exposure to lead. In this cross-sectional study, we measured tibial lead by (109)Cd K-shell X-ray fluorescence. Tibial lead was evaluated in subjects with different VDR genotypes defined using the BsmI restriction enzyme, adjusting for confounding variables. Study participants had a mean age +/- SD of 57.4 +/- 7.6 years. A total of 169 (33.5%) subjects were homozygous for the BsmI restriction site (designated bb), 251 (49.8%) were heterozygous (Bb), and 84 (16.7%) were homozygous for the absence of the restriction site (BB). Among all of the study subjects, tibial lead concentrations were low, with a mean +/- SD of 14.4 +/- 9.3 microg Pb/g bone mineral. There were only small differences in tibial lead concentrations by VDR genotype, with mean +/- SD tibial lead concentrations of 13.9 +/- 7.9, 14.3 +/- 9.5, and 15.5 +/- 11.1 in subjects with bb, Bb, and BB, respectively. In a multiple linear regression model of tibial lead concentrations, the VDR genotype modified the relation between age and tibial lead concentrations; subjects with the B allele had larger increases in tibial lead concentrations with increasing age (0.37, 0.48, and 0.67 microg/g per year of age in subjects with bb, Bb, and BB, respectively; the adjusted p-value for trend in slopes = 0.04). The VDR genotype also modified the relation between years since last exposure to lead and tibial lead concentrations. Subjects with bb evidenced an average decline in tibial lead concentrations of 0.10 microg/g per year since their last exposure to lead, whereas subjects with Bb and BB evidenced average increases of 0.03 and 0.11 microg/g per year, respectively (the adjusted p-value for trend in slopes = 0.01). Polymorphisms in the vitamin D receptor modified the relations of age and years since the last exposure to lead with tibial lead concentrations. Although controversy remains on the influence of the VDR genotype on bone mineral density, the data suggest that variant VDR alleles modify lead concentrations in bone, either by influencing lead content or calcium content or both. PMID- 10706525 TI - Susceptibility to infections and immune status in Inuit infants exposed to organochlorines. AB - We investigated whether organochlorine exposure is associated with the incidence of infectious diseases in Inuit infants from Nunavik (Arctic Quebec, Canada). We compiled the number of infectious disease episodes during the first year of life for 98 breast-fed and 73 bottle-fed infants. Concentrations of organochlorines were measured in early breast milk samples and used as surrogates to prenatal exposure levels. Immune system parameters were determined in venous blood samples collected from infants at 3, 7, and 12 months of age. Otitis media was the most frequent disease, with 80. 0% of breast-fed and 81.3% of bottle-fed infants experiencing at least one episode during the first year of life. During the second follow-up period, the risk of otitis media increased with prenatal exposure to p,p'-DDE, hexachlorobenzene, and dieldrin. The relative risk (RR) for 4- to 7-month-old infants in the highest tertile of p, p'-DDE exposure as compared to infants in the lowest tertile was 1. 87 [95% confidence interval (CI), 1.07-3.26]. The RR of otitis media over the entire first year of life also increased with prenatal exposure to p,p'-DDE (RR, 1.52; CI, 1.05-2.22) and hexachlorobenzene (RR, 1.49; CI, 1.10-2.03). Furthermore, the RR of recurrent otitis media ( [Greater/equal to] 3 episodes) increased with prenatal exposure to these compounds. No clinically relevant differences were noted between breast-fed and bottle-fed infants with regard to immunologic parameters, and prenatal organochlorine exposure was not associated with immunologic parameters. We conclude that prenatal organochlorine exposure could be a risk factor for acute otitis media in Inuit infants. PMID- 10706526 TI - Airborne concentrations of PM(2.5) and diesel exhaust particles on Harlem sidewalks: a community-based pilot study. AB - Residents of the dense urban core neighborhoods of New York City (NYC) have expressed increasing concern about the potential human health impacts of diesel vehicle emissions. We measured concentrations of particulate matter [less than/equal to] 2.5 micro in aerodynamic diameter (PM(2.5)) and diesel exhaust particles (DEP) on sidewalks in Harlem, NYC, and tested whether spatial variations in concentrations were related to local diesel traffic density. Eight hour (1000-1800 hr) air samples for PM(2.5 )and elemental carbon (EC) were collected for 5 days in July 1996 on sidewalks adjacent to four geographically distinct Harlem intersections. Samples were taken using portable monitors worn by study staff. Simultaneous traffic counts for diesel trucks, buses, cars, and pedestrians were carried out at each intersection on [Greater/equal to] 2 of the 5 sampling days. Eight-hour diesel vehicle counts ranged from 61 to 2,467 across the four sites. Mean concentrations of PM(2.5) exhibited only modest site-to-site variation (37-47 microg/m(3)), reflecting the importance of broader regional sources of PM(2.5). In contrast, EC concentrations varied 4-fold across sites (from 1.5 to 6 microg/m(3)), and were associated with bus and truck counts on adjacent streets and, at one site, with the presence of a bus depot. A high correlation (r = 0.95) was observed between EC concentrations measured analytically and a blackness measurement based on PM(2.5) filter reflectance, suggesting the utility of the latter as a surrogate measure of DEP in future community-based studies. These results show that local diesel sources in Harlem create spatial variations in sidewalk concentrations of DEP. The study also demonstrates the feasibility of a new paradigm for community-based research involving full and active partnership between academic scientists and community based organizations. PMID- 10706527 TI - Permanent and functional male-to-female sex reversal in d-rR strain medaka (Oryzias latipes) following egg microinjection of o,p'-DDT. AB - Complete sex reversal of fish is accomplished routinely in aquaculture practices by exposing fish to exogenous sex steroids during gonadal differentiation. A variety of environmental chemicals are also active at sex steroid receptors and theoretically possess the potential to alter normal sexual differentiation in fish. However, in controlled environmental chemical exposures to date, only partial alterations of fish sexual phenotype have been observed. Here we report complete, permanent, and functional male-to-female sex reversal in the Japanese medaka (Oryzias latipes, d-rR strain) after a onetime embryonic exposure to the xenoestrogen o, p'-DDT. d-rR strain medaka are strict gonochorists that possesses both sex-linked pigmentation, which distinguishes genotypic sex, and sexually dimorphic external secondary sexual characteristics, which distinguish phenotypic sex. We directly microinjected the xenoestrogen o, p'-DDT into the egg yolks of medaka at fertilization to parallel the maternal transfer of lipophilic contaminants to the embryo. At 10 weeks of age, microinjected medaka were examined for mortality and sex reversal. A calculated embryonic dose of 511 +/- 22 ng/egg o, p'-DDT (mean +/- standard error) resulted in 50% mortality. An embryonic exposure of 227 +/- 22 ng/egg o, p'-DDT resulted in 86% (6 of 7) sex reversal of genetic males to a female phenotype (XY females). XY females were distinguished by sex-linked male pigmentation accompanying female secondary sexual characteristics. Histologic examination of the gonads confirmed active ovaries in 100% of the XY females. In 10-day breeding trials in which XY females were paired with normal XY males, 50% of the XY females produced fertilized embryos; this represents a comparable breeding success rate to normal XX females. Fertilized eggs produced from XY females hatched to viable larvae. These results clearly indicate that a weakly estrogenic pesticide, o, p'-DDT, when presented during the critical period of gonadal development, can profoundly alter sexual differentiation. PMID- 10706528 TI - Environmental injustice in North Carolina's hog industry. AB - Rapid growth and the concentration of hog production in North Carolina have raised concerns of a disproportionate impact of pollution and offensive odors on poor and nonwhite communities. We analyzed the location and characteristics of 2,514 intensive hog operations in relation to racial, economic, and water source characteristics of census block groups, neighborhoods with an average of approximately 500 households each. We used Poisson regression to evaluate the extent to which relationships between environmental justice variables and the number of hog operations persisted after consideration of population density. There are 18.9 times as many hog operations in the highest quintile of poverty as compared to the lowest; however, adjustment for population density reduces the excess to 7.2. Hog operations are approximately 5 times as common in the highest three quintiles of the percentage nonwhite population as compared to the lowest, adjusted for population density. The excess of hog operations is greatest in areas with both high poverty and high percentage nonwhites. Operations run by corporate integrators are more concentrated in poor and nonwhite areas than are operations run by independent growers. Most hog operations, which use waste pits that can contaminate groundwater, are located in areas with high dependence on well water for drinking. Disproportionate impacts of intensive hog production on people of color and on the poor may impede improvements in economic and environmental conditions that are needed to address public health in areas which have high disease rates and low access to medical care as compared to other areas of the state. PMID- 10706529 TI - Intensive livestock operations, health, and quality of life among eastern North Carolina residents. AB - People who live near industrial swine operations have reported decreased health and quality of life. To investigate these issues, we surveyed residents of three rural communities, one in the vicinity of an approximately 6,000-head hog operation, one in the vicinity of two intensive cattle operations, and a third rural agricultural area without livestock operations that use liquid waste management systems. Trained interviewers obtained information about health symptoms and reduced quality of life during the previous 6 months. We completed 155 interviews, with a refusal rate of 14%. Community differences in the mean number of episodes were compared with adjustment for age, sex, smoking, and employment status. The average number of episodes of many symptoms was similar in the three communities; however, certain respiratory and gastrointestinal problems and mucous membrane irritation were elevated among residents in the vicinity of the hog operation. Residents in the vicinity of the hog operation reported increased occurrences of headaches, runny nose, sore throat, excessive coughing, diarrhea, and burning eyes as compared to residents of the community with no intensive livestock operations. Quality of life, as indicated by the number of times residents could not open their windows or go outside even in nice weather, was similar in the control and the community in the vicinity of the cattle operation but greatly reduced among residents near the hog operation. Respiratory and mucous membrane effects were consistent with the results of studies of occupational exposures among swine confinement-house workers and previous findings for neighbors of intensive swine operations. Long-term physical and mental health impacts could not be investigated in this study. PMID- 10706530 TI - Measurement variability associated with KXRF bone lead measurement in young adults. AB - In vivo bone lead measurement using K X-ray fluorescence (KXRF) has been used to estimate long-term lead exposure, especially in adults. Relatively few studies have been conducted on young subjects with this technique. To explore the measurement variability of KXRF bone lead measurements in young subjects, the tibiae of two male cadavers from Boston, Massachusetts, 17 and 20 years of age, were obtained for repeated bone lead measurements. Bone lead concentrations were measured using a grid of nine locations, 1 cm apart, centered at the midpoint of the tibia. Each location was sampled using five 60-min measurements. Measured concentrations ranged from < 0 to 11.8 microg Pb/g bone mineral across a tibia with mean concentrations for the midpoint locations of 0.8 microg Pb/g bone mineral SD = 2.5 and 2.0 microg Pb/g bone mineral (SD = 1.9) for the left and right legs of the younger subject and 3.6 microg Pb/g bone mineral (SD = 2.6) and 6.0 microg Pb/g bone mineral (SD = 3.3) for the left and right legs of the older subject. Although bone lead concentrations did not vary significantly by measurement location in an individual leg, reported measurement uncertainty increased significantly at locations that were 1 cm from the center of the tibia horizontally (p < 0.0001). Symmetry in bone lead concentration between legs was observed for the 17-year-old subject. Potential asymmetry between the left and right legs was suggested for the 20-year-old subject (p = 0.06). These data describe the degree of variability that may be associated with bone lead measurements of young subjects with low bone lead concentrations using a standard spot-source KXRF instrument. Because of the importance of conducting additional research on adolescent lead toxicity, further improvements to the precision of KXRF measurement are needed. PMID- 10706531 TI - Strain differences in vaginal responses to the xenoestrogen bisphenol A. AB - Bisphenol A (BPA) is the monomer component of polycarbonate plastics and epoxy resins; human exposure derives from leachate in foodstuffs packaged in certain plastics or from epoxy-based dental appliances. BPA stimulates prolactin secretion in Fischer 344 (F344) rats but not in Sprague-Dawley (S-D) rats. The present studies were performed to determine if another classic estrogen target tissue, the rat vagina, responds to BPA in a strain-specific manner. In F344 rats BPA increased DNA synthesis in vaginal epithelium with a median effective dose (ED(50)) of 37.5 mg/kg body weight; DNA synthesis was not stimulated in S-D rats by any dose tested. Clearance of (3)H-BPA from blood followed the same time course in both strains of rats, with a half-life of 90 min. Scatchard analysis of [(3)H]estradiol binding showed no strain differences in concentration or affinity of the vaginal estrogen receptor. BPA increased the level of mRNA for the immediate early gene, c-fos, with similar dose-response curves in both rat strains. Thus, F344 and S-D rats exhibit differences in sensitivity to BPA at the level of cell proliferation in the vaginal epithelium. However, metabolic clearance of BPA and the early events that lead to the proliferative response, receptor-ligand interaction and induction of immediate early genes, show no strain differences. These observations suggest that differences in intermediate effects must account for the difference in sensitivity of the proliferative response to the xenoestrogen. Furthermore, these results point to the need for caution in choosing a suitable end point and animal model when seeking to test the estrogenic effects of xenobiotics. PMID- 10706532 TI - Salmonid sexual development is not consistently altered by embryonic exposure to endocrine-active chemicals. AB - Fish sexual development is sensitive to exogenous hormone manipulation, and salmonids have been used extensively as environmental sentinels and models for biomedical research. We simulated maternal transfer of contaminants by microinjecting rainbow trout (Oncorhynchus mykiss) and chinook salmon (Oncorhynchus tshawytscha) embryos. Fish were reared for 6 months and sexed, and gonads were removed for histology and measurement of in vitro steroid production. Analysis of fat samples showed that dichlorodiphenylethylene (DDE) levels, o, p'M DDE and p,o, p'-DDE isomers, were elevated 6 months after treatment. A preliminary study showed an increased ratio of males to females after treatment with 80 mg/kg and 160 mg/kg of the xenoestrogen o,o, p'-DDE. One fish treated with 160 mg/kg o,o, p'-DDE had gonads with cells typical of both males and females. A follow-up study, using more fish and excluding the highly toxic 160 mg/kg o,o, p'-DDE dose, showed no effect on sex ratio or gonadal histology. Embryonic exposure of monosex male trout, monosex female trout, and mixed sex salmon to o, o, p'-DDE, p,o, p'-DDE, mixtures of DDE isomers, and octylphenol failed to alter sexual development. We observed no treatment-dependent changes in in vitro gonadal steroid production in any experiments. Trout exposed in ovo and reared to maturity spawned successfully. These results suggest that mortality attributable to the xenoestrogens o,o, p'-DDE, chlordecone, and octylphenol, and the antiandrogen p,o, p'-DDE, is likely to occur before the appearance of subtle changes in sexual development. Because trout appeared to be sensitive to endocrine disruption, we cannot dismiss the threat of heavily contaminated sites or complex mixtures to normal sexual development of salmonids or other aquatic organisms. PMID- 10706533 TI - Benchmark concentrations for methylmercury obtained from the Seychelles Child Development Study. AB - Methylmercury is a neurotoxin at high exposures, and the developing fetus is particularly susceptible. Because exposure to methylmercury is primarily through fish, concern has been expressed that the consumption of fish by pregnant women could adversely affect their fetuses. The reference dose for methylmercury established by the U.S. Environmental Protection Agency was based on a benchmark analysis of data from a poisoning episode in Iraq in which mothers consumed seed grain treated with methylmercury during pregnancy. However, exposures in this study were short term and at much higher levels than those that result from fish consumption. In contrast, the Agency for Toxic Substances and Disease Registry (ATSDR) based its proposed minimal risk level on a no-observed-adverse-effect level (NOAEL) derived from neurologic testing of children in the Seychelles Islands, where fish is an important dietary staple. Because no adverse effects from mercury were seen in the Seychelles study, the ATSDR considered the mean exposure in the study to be a NOAEL. However, a mean exposure may not be a good indicator of a no-effect exposure level. To provide an alternative basis for deriving an appropriate human exposure level from the Seychelles study, we conducted a benchmark analysis on these data. Our analysis included responses from batteries of neurologic tests applied to children at 6, 19, 29, and 66 months of age. We also analyzed developmental milestones (age first walked and first talked). We explored a number of dose-response models, sets of covariates to include in the models, and definitions of background response. Our analysis also involved modeling responses expressed as both continuous and quantal data. The most reliable analyses were considered to be represented by 144 calculated lower statistical bounds on the benchmark dose (BMDLs; the lower statistical bound on maternal mercury hair level corresponding to an increase of 0.1 in the probability of an adverse response) derived from the modeling of continuous responses. The average value of the BMDL in these 144 analyses was 25 ppm mercury in maternal hair, with a range of 19 to 30 ppm. PMID- 10706535 TI - Genomic imprinting and environmental disease susceptibility. AB - Genomic imprinting is one of the most intriguing subtleties of modern genetics. The term "imprinting" refers to parent-of-origin-dependent gene expression. The presence of imprinted genes can cause cells with a full parental complement of functional autosomal genes to specifically express one allele but not the other, resulting in monoallelic expression of the imprinted loci. Genomic imprinting plays a critical role in fetal growth and behavioral development, and it is regulated by DNA methylation and chromatin structure. This paper summarizes the Genomic Imprinting and Environmental Disease Susceptibility Conference held 8-10 October 1998 at Duke University, Durham, North Carolina. The conference focused on the importance of genomic imprinting in determining susceptibility to environmentally induced diseases. Conference topics included rationales for imprinting: parental antagonism and speciation; methods for imprinted gene identification: allelic message display and monochromosomal mouse/human hybrids; properties of the imprinted gene cluster human 11p15.5 and mouse distal 7; the epigenetics of X-chromosome inactivation; variability in imprinting: imprint erasure, non-Mendelian inheritance ratios, and polymorphic imprinting; imprinting and behavior: genetics of bipolar disorder, imprinting in Turner syndrome, and imprinting in brain development and social behavior; and aberrant methylation: methylation and chromatin structure, methylation and estrogen exposure, methylation of tumor-suppressor genes, and cancer susceptibility. Environmental factors are capable of causing epigenetic changes in DNA that can potentially alter imprint gene expression and that can result in genetic diseases including cancer and behavioral disorders. Understanding the contribution of imprinting to the regulation of gene expression will be an important step in evaluating environmental influences on human health and disease. PMID- 10706534 TI - The Harvard Southern California Chronic Ozone Exposure Study: assessing ozone exposure of grade-school-age children in two Southern California communities. AB - The Harvard Southern California Chronic Ozone Exposure Study measured personal exposure to, and indoor and outdoor ozone concentrations of, approximately 200 elementary school children 6-12 years of age for 12 months (June 1995-May 1996). We selected two Southern California communities, Upland and several towns located in the San Bernardino mountains, because certain characteristics of those communities were believed to affect personal exposures. On 6 consecutive days during each study month, participant homes were monitored for indoor and outdoor ozone concentrations, and participating children wore a small passive ozone sampler to measure personal exposure. During each sampling period, the children recorded time-location-activity information in a diary. Ambient ozone concentration data were obtained from air quality monitoring stations in the study areas. We present ozone concentration data for the ozone season (June September 1995 and May 1996) and the nonozone season (October 1995-April 1996). During the ozone season, outdoor and indoor concentrations and personal exposure averaged 48.2, 11.8, and 18.8 ppb in Upland and 60.1, 21.4, and 25.4 ppb in the mountain towns, respectively. During the nonozone season, outdoor and indoor concentrations and personal exposure averaged 21.1, 3.2, and 6.2 ppb in Upland, and 35.7, 2.8, and 5.7 ppb in the mountain towns, respectively. Personal exposure differed by community and sex, but not by age group. PMID- 10706536 TI - Environmental ethics. AB - The U.S. Environmental Protection Agency (EPA) held the first meeting on environmental ethics sponsored by the Scientific Advisory Panel and Board on 10 11 December 1998 in Arlington, Virginia (1). The report from the meeting will more completely inform scientists and the community of current issues. This editorial should serve as an initial brief of this meeting [which was held on the fiftieth anniversary of the Declaration of Human Rights (adopted by the United Nations on 10 December 1948)]. PMID- 10706537 TI - Carbon disulfide. PMID- 10706538 TI - Alternative tests make the grade. AB - Toxicity testing is absolutely necessary for assessing the safety of substances in food, air, and water, in the workplace and at home. Although there are several tried-and-true toxicity assays, the search is always on for methods that can even better predict toxic effects. As scientific understanding of the effects of environmental toxicants grows, new tests are needed to evaluate previously unexamined end points and to take advantage of advances in biotechnology and the growing knowledge of how toxicants work at the molecular and cellular levels. Another issue is how to develop tests that can reliably and accurately assess toxicity using less time, money, and materials, and with greater regard for animal welfare. The Interagency Coordinating Committee on the Validation of Alternative Methods (ICCVAM) was established in 1997 to address these needs through the development, validation, acceptance, and harmonization of new and revised toxicological test methods throughout the federal government. PMID- 10706539 TI - The dead zones: oxygen-starved coastal waters. AB - After the great Mississippi River flood of 1993, the hypoxic (or low-oxygen) "dead zone" in the Gulf of Mexico more than doubled its size, reaching an all time high of over 7,700 square miles in July of 1999. Scientists attribute the Gulf of Mexico dead zone largely to nutrient runoff from agriculture in the Mississippi River basin. During the warm months, these nutrients fuel eutrophication, or high organic production, causing large algal blooms. When the algae decay, the result is hypoxia. Reports of such hypoxic events around the world have been increasing since the mid 1960s. Eutrophication and hypoxia have resulted in mortality of bottom-dwelling life in dozens of marine ecosystems and have stressed fisheries worldwide. Some algal blooms can alter the function of coastal ecosystems or, potentially, threaten human health. Anthropogenic nutrient loading from sources such as agriculture, fossil fuel emissions, and climate events is believed to be related to the global increase in frequency, size, and duration of certain algal blooms. PMID- 10706540 TI - The next target of bioterrorism: your food. AB - One of the many forms that biological warfare may take is the targeting of major food crops. In a poor country where millions of citizens depend on staple crops such as rice, an act of bioterrorism that destroys the crop would create a famine, resulting not only in malnutrition and starvation but also in reduced immune resistance to a range of common illnesses. To reduce the potential of deliberate introductions of crop pathogens as acts of terrorism, researchers must be able to "fingerprint" pathogens at the molecular level and discriminate between naturally occurring and deliberately introduced outbreaks. Several domestic and international surveillance, tracking, and reporting efforts are under way. PMID- 10706541 TI - Farming from a new perspective: remote sensing comes down to earth. AB - Farmers strive to increase the yield of their fields by adding nutrients and water to the land, and using pesticides to control insects and disease. In addition to bountiful harvests, the results of their endeavors may include elevated amounts of fertilizers in surface waters and aquifers and potential risk to themselves and their neighbors from exposure to pesticides. Precision agriculture is the use of modern information technologies such as geographic information systems, the global positioning system, and remote sensing from the air to reduce the environmental effects of these chemicals while enhancing the productivity of farming. By combining crop yield maps with soil survey maps and remote sensing output, farmers can identify areas that need more or less fertilizer, water, or pesticide. PMID- 10706542 TI - Functional heterogeneity of the intrahepatic biliary epithelium. PMID- 10706543 TI - Sodium, hydrogen exchange type 1 and bile ductular secretory activity in the guinea pig. AB - Biliary epithelial cells (BECs) express different Na(+), H(+) exchange (NHE) isoforms. In this study, the potential role of NHE in ductular bile secretion is assessed. Experiments were performed in guinea pig perfused livers and isolated BECs. Inhibition of NHE was achieved by hypotonic stress and by using the unspecific NHE inhibitor, amiloride, or the specific NHE 1 inhibitor, cariporide (HOE 642). Hypotonic stress inhibited basal bile flow by 46% and prevented secretin stimulation of bile flow by reducing biliary bicarbonate output by 50%. Secretin increased bile flow from 3.7 +/- 0.8 microL/min/g to 4.78 microL/min/g (P <.01); subsequent exposure to hypotonic stress decreased secretin-stimulated bile flow by 35% and biliary bicarbonate secretion by approximately 50%. Inhibition of NHE by amiloride or cariporide resulted in a similar reduction of secretin-stimulated bile flow and bicarbonate secretion. Basal bile flow was unaffected by the NHE inhibitors. In isolated guinea pig BECs, regulatory volume decrease and inhibition of NHE was demonstrated after hypotonic stress under basal and secretin-stimulated conditions. In contrast, hypotonic exposure inhibited Cl(-), HCO(3)(-) exchange activity in isolated BECs only during basal conditions but incompletely after secretin stimulation. Our study shows that hypotonic stress inhibits basal bile flow in the guinea pig by inhibition of Cl( ), HCO(3)(-) exchange. NHE1 is not involved in basal bile formation. Increased choleresis after ductular stimulation by secretin depends on intact NHE1 activity. These data indicate that BEC volume changes have profound effects on biliary secretory function. PMID- 10706544 TI - Local arterial vasoconstriction induced by octreotide in patients with cirrhosis. AB - Peripheral vasodilation initiates the hyperdynamic circulation in cirrhosis. Somatostatin and its analogues, such as octreotide, have a vasoconstrictive effect in cirrhotic patients and experimental animals with portal hypertension. The exact mechanism of octreotide-induced vasoconstriction remains unknown. To investigate whether octreotide produces vasoconstriction through suppression of vasodilatory peptides, such as glucagon, or through a local effect, we evaluated the effect of an intra-arterial dose on forearm blood flow (FBF), while measuring systemic glucagon levels. FBF was measured in 10 cirrhotic patients by venous occlusion plethysmography. The brachial artery of the nondominant arm was catheterized, and vasoactive drugs were administered: methacholine 4 microg/min; octreotide 20 microg/h, and octreotide 20 microg/h + methacholine 4 microg/min. Each infusion, lasting 5 minutes, was followed by saline for washout. FBF was measured in both arms during the last minute of each infusion and at the end of washout, with the uninfused arm acting as the control. Nitrates and nitrites, octreotide, and glucagon blood levels were determined at baseline and after each infusion. Percent change in flow (%triangle up) was obtained by comparing the flow during drug administration to that during the preceding saline infusion. Saline infusion did not alter FBF, but octreotide infusion resulted in a 34% +/- 7.7 (P <.005) reduction in FBF in the infused arm. FBF in the control arm was unchanged despite a significant decrease in systemic glucagon levels. Methacholine infusion increased FBF around 300%, which was not altered by the concomitant infusion of octreotide. Octreotide has a local vasoconstrictive effect that seems nitric oxide (NO)-independent. Octreotide probably has a facilitating effect over vasoconstrictors increased in chronic liver diseases. PMID- 10706545 TI - Influence of donor cardiopulmonary arrest in human liver transplantation: possible role of ischemic preconditioning. AB - Hepatic allografts from donors who have suffered a brief cardiopulmonary arrest may sustain ischemic damage before organ procurement. However, there is no reported correlation between donor cardiopulmonary arrest and hepatic allograft dysfunction. On the other hand, brief ischemia-reperfusion injury has been shown experimentally to result in protection in several organ models. Induction of ischemic tolerance has been called ischemic preconditioning. Our objective was to study the influence of brief donor cardiopulmonary arrest on hepatic allograft outcome in human liver transplantation. Between May 1997 and July 1998, 181 consecutive orthotopic liver transplant (OLT) cases were divided into 2 groups based on the occurrence of donor cardiopulmonary arrest. Group A consisted of 37 donors who suffered a cardiopulmonary arrest. Group B consisted of the remaining 144 patients. Liver graft survival within 90 days of OLT and early postoperative graft function were analyzed. Although there was significant liver damage resulting from circulatory failure during cardiopulmonary arrest before organ procurement in group A, graft survival was not affected. After OLT, the mean peak aspartate transaminase and alanine transaminase concentrations in group A (1, 444.1 and 718.2 U/L) were significantly lower than those in group B (2,382.8 and 1,507.3 U/L) (P <.05). Experiences of brief cardiopulmonary arrest in organ donors did not affect post-OLT hepatic allograft survival and function. Although the liver function tests are elevated in an organ donor, the hepatic allograft is suitable for OLT if the liver damage is induced by brief donor cardiopulmonary arrest. PMID- 10706546 TI - The role of portal pressure in the severity of bleeding in portal hypertensive rats. AB - The aim of this study was to investigate the role of portal hypertension determining the severity of bleeding in portal hypertensive rats. The effects of section of branches of the ileocolic vein were studied in sham-operated (SO), partial portal vein-ligated (PPVL), and common bile duct-ligated (CBDL) rats. The ensuing hemorrhage was compared with that caused by section of femoral vein, where the portal hypertensive factor is excluded. In PPVL rats, section of branches of increasing size (divided into fourth, third, second, and first order) resulted in increasingly severe bleeding (arterial pressure: / +/- 4%, / 6 +/- 12%, / /15 +/- 8%, and / 28 +/- 13%; P <.005; hematocrit / 4 +/- 2%, / 6 +/- 1%, / 7 +/- 2%, and / 10 +/- 4%; P <.005). Bleeding from first-order branches was mild in SO, moderate in PPVL, and severe in CBDL rats, as shown by increasing changes in arterial pressure (/ 3 +/- 3%, / 12 +/- 16% and, / 43 +/- 23%; P <.01), hematocrit (/ 4 +/- 1%, / 12 +/- 2%, and / 32 +/- 19%; P <.01), and mortality (0%, 0%, and 56%; P <.001). Greater blood loss in CBDL rats was associated with higher portal pressure (16.6 +/- 2.7 vs. 13. 1 +/- 1.1 mm Hg in PPVL; P <.01) and more prolonged bleeding time (70 +/- 4 vs. 35 +/- 3 seconds in PPVL; P <.001). Vessels were similarly dilated in CBDL and PPVL (0.7 +/- 0.2 and 0.7 +/- 0.1 vs. 0.4 +/- 0.1 mm in SO; P <.05). Section of femoral vein caused equal blood loss in SO, PPVL, and CBDL rats, assessed by falls in hematocrit (/ 8 +/- 2%, / 7 +/- 1%, / 8 +/- 1%, respectively; NS) and by the blood loss (3.6 +/- 0.7, 3.5 +/- 0.9, and 3.8 +/- 0.7 g; NS). The study shows that the degree of portal pressure elevation is a major determinant of the severity of portal hypertension-related bleeding in PPVL and CBDL rats. PMID- 10706547 TI - Cause of portal or hepatic venous thrombosis in adults: the role of multiple concurrent factors. AB - According to a recent hypothesis, venous thrombosis results from the concurrence of several factors. This hypothesis was assessed in patients with portal or hepatic venous thrombosis by simultaneously investigating most of the currently identified prothrombotic disorders, local precipitating factors, and other risk factors such as oral contraceptive use. Patients with a tumorous obstruction and patients with cirrhosis with portal vein thrombosis were excluded. The prothrombotic disorders that were investigated included classical and occult myeloproliferative disorders; antiphospholipid syndrome; protein C; protein S and antithrombin deficiency; factor V Leiden; factor II; and methylene tetrahydrofolate-reductase gene mutations. We found 1 or several prothrombotic disorders and a local precipitating factor in 26 and 10 of the 36 patients with portal vein thrombosis, respectively; and in 28 and none of the 32 patients with hepatic vein thrombosis, respectively. We found a combination of prothrombotic disorders in 5 and 9 patients with portal and hepatic vein thrombosis, respectively, whereas such a combination is expected in less than 1% of asymptomatic subjects. Of the 10 patients with a local precipitating factor, 8 had a prothrombotic disorder. Of the 13 patients who use oral contraceptives, 10 had a prothrombotic disorder. We conclude that portal or hepatic venous thrombosis should be regarded as an index for 1 or several prothrombotic disorders, whether or not local precipitating factors or oral contraceptive use are found. Concurrence of prothrombotic disorders is more common than expected. Extensive investigation of prothrombotic disorders and anticoagulation should be considered in patients with portal or hepatic venous thrombosis. PMID- 10706548 TI - Insulin-like growth factor-I reverts testicular atrophy in rats with advanced cirrhosis. AB - The pathogenesis of hypogonadism in cirrhosis is not completely understood. The levels of insulin-like growth factor-I (IGF-I), an anabolic factor with trophic actions on testes, are reduced in cirrhosis. This study was undertaken to evaluate whether rats with advanced cirrhosis develop hypogonadism and whether the administration of IGF-I exerts beneficial effects on testicular structure and function. Wistar rats with ascitic cirrhosis induced with CCl(4) were allocated into 2 groups (n = 10, each) to receive recombinant IGF-I (20 microg x kg(-1) x d(-1), subcutaneously) or vehicle for 3 weeks. Healthy rats receiving vehicle were used as the control group (n = 10). At baseline, both cirrhotic groups showed similar deterioration of liver function tests. Compared with controls, nontreated cirrhotic rats showed decreased serum levels of IGF-I (P <.05), reduced testicular size and weight (P <.001), and intense histopathological testicular abnormalities, including reduced tubular diameters (P <.001), loss of the germinal line (P <. 001), and diminutions in cellular proliferation, spermatogenesis (P <.001), and testicular transferrin expression (P <.001). In addition, low serum testosterone (P <.01) and high serum LH (P <.01) were present in untreated cirrhotic animals. Cirrhotic rats that received IGF-I showed full recovery of testicular size and weight and of all histopathological abnormalities (P <.001 to <.01 vs. nontreated cirrhotic rats; P = ns vs. controls). Serum levels of sex hormones tended to normalize. In conclusion, IGF-I deficiency may play a pathogenetic role in hypogonadism of cirrhosis. Low doses of IGF-I for a short period of time revert testicular atrophy and appear to improve hypogonadism in advanced experimental cirrhosis. PMID- 10706549 TI - p73 is up-regulated in a subset of hepatocellular carcinomas. AB - Loss of heterozygosity (LOH) at 1p36 occurs in a number of solid tumors including hepatocellular carcinoma (HCC). Recently, a novel gene, p73, has been identified at 1p36.33. p73 is structurally and functionally related to p53 located at 17p13.1, which is a target for inactivation in HCCs. p73 produces at least two splicing variants, p73alpha and beta, and a polymorphism in exon 2 results in two alleles, GC or AT. Initially, only the AT allele and p73alpha transcripts were identified in malignant cell lines, suggesting a role for these in the malignant phenotype. The aims of this study were to determine the extent of LOH at 1p36 and 17p13.1 in HCCs from Australia and South Africa, and to identify patterns of p73 mRNA and p73 and p53 protein expression. LOH at 1p36 was found in 8 of 25 Australian and 6 of 10 South African cases. p73 mRNA expression occurred in 8 HCCs, but not in nonmalignant liver tissue. Two of these 8 HCCs had LOH of 1p36. Both alpha and beta transcripts were observed in GC/GC homozygotes and GC/AT heterozygotes. No p73 protein expression was observed by immunohistochemistry in nonmalignant liver tissue or in HCC. p53 inactivation appeared to be associated with up-regulation of p73 expression, suggesting a compensatory role for p73 in this situation. The LOH at 1p36 implies a liver-specific tumor suppressor gene is in this region. However, the up-regulation of p73 mRNA suggests p73 is not the target of this loss. PMID- 10706550 TI - Gene therapy of hepatocellular carcinoma in vitro and in vivo in nude mice by adenoviral transfer of the Escherichia coli purine nucleoside phosphorylase gene. AB - Expression of viral or bacterial enzymes in tumor cells to convert nontoxic prodrugs into highly toxic metabolites is an attractive gene-therapeutic approach for the treatment of hepatocellular carcinoma (HCC). The Escherichia coli purine nucleoside phosphorylase (PNP) converts purine analogs into freely diffusible metabolites, which are highly toxic to dividing and nondividing cells. We investigated the antitumor effects of PNP in the human HCC cell lines, HepG2, Hep3B, and HuH-7, and performed a comparison with herpes simplex thymidine kinase (TK). The genes for PNP, TK, and enhanced green fluorescent protein (EGFP) were delivered to HCC cells by identical adenoviral vectors. Fludarabine and ganciclovir (GCV) served as prodrugs for PNP and TK, respectively. Expression of PNP highly sensitized HCC cells to fludarabine treatment. Fludarabine concentrations between 0.5 and 1 microg/mL killed 100% of the cells expressing PNP with no detectable toxicity in control cells expressing EGFP. Expression of PNP in as few as 10% of HCC cells induced efficient killing of most bystander cells. Expression of TK followed by GCV treatment produced a potent growth inhibition but failed to kill all TK-expressing HCC cells. More importantly, the TK system exhibited a lower degree of bystander effect. Adenoviral delivery of PNP followed by fludarabine administration prevented subcutaneous and intrahepatic tumor formation in nude mice and was also effective for the treatment of established tumors. These results demonstrate the potential of the PNP/fludarabine system for the treatment of HCC. PMID- 10706551 TI - Clonal expansion in evolution of chronic hepatitis to hepatocellular carcinoma as seen at an X-chromosome locus. AB - Clonal analysis has shown that hepatocellular carcinoma arises from a single cell. However, the clonality of precancerous lesions and adjacent nonneoplastic tissues is not clear. We analyzed a human androgen receptor locus to elucidate the clonal state of liver tissues including post-hepatitic lesions associated with hepatocarcinogenesis. The analysis was based on a restriction fragment length polymorphism involving an androgen receptor locus on the X chromosome, taking advantage of physiologic random inactivation by methylation of 1 of 2 X chromosomes in females during embryogenesis. Clonality was assessed in 79 randomly located tissue samples microdissected from noncirrhotic liver, including a total of 40 morphologically normal sites in 4 normal livers and 39 sites from a single HCV-infected liver. In addition, 51 regenerative nodules, 4 areas of adenomatous hyperplasia, and 18 hepatocellular carcinomas were sampled. All samples were obtained from livers involved by various neoplasms. Eight of forty samples (20.0%) from the four normal livers and 20 of the 39 samples (51.3%) from the single HCV-infected liver showed a monoclonal pattern. Moreover, 30 of 51 regenerative nodules (58.9%) showed a monoclonal pattern. No histologic differences were evident between mono- and polyclonal nodules. On the other hand, the 18 carcinomas and 4 areas of adenomatous hyperplasia all were monoclonal. Mean calculated monoclonal areas of normal liver and liver with chronic hepatitis were 1.1 and 3.3 mm(2). Our results suggest that areas representing a single clone of hepatocytes are present in normal liver, and these progressively expand as changes advance from chronic hepatitis to hepatocellular carcinoma. PMID- 10706552 TI - In situ determination by surface chemiluminescence of temporal relationships between evolving warm ischemia-reperfusion injury in rat liver and phagocyte activation and recruitment. AB - Liver ischemia-reperfusion is characterized by an increased oxygen-dependent free radical chain-reaction rate and an increased steady-state concentration of reactive oxygen species. The aim of this study was to evaluate the in situ generation of reactive oxygen species and its relationship with phagocyte activation and recruitment in reperfused rat liver. Rat livers were subjected to 2 hours of selective lobular ischemia and reperfusion for up to 12 hours. The following parameters were determined: in situ liver chemiluminescence, understood to reflect the tissue steady-state concentration of singlet oxygen ((1)O(2)); myeloperoxidase tissue activity; the number of neutrophils; and the degree of necrosis. An early chemiluminescence burst was measured after 30 minutes of blood reflow (early phase of oxidative stress), followed by a relapse and a further increase after 4 to 12 hours of reperfusion (late phase of oxidative stress). Both early and late phases were modified by pretreatment with gadolinium chloride (GdCl(3)), pointing to a key role of the Kupffer cells. Neutrophils infiltrated into the liver, myeloperoxidase activity, in situ chemiluminescence, and necrosis were found to be strongly correlated over the 4- to 12-hour reperfusion period (r =.960; average of the 4 correlation coefficients). Together with resident phagocytes, neutrophil recruitment and activation appear to provide a major contribution to the increase of oxygen-dependent free-radical reactions and amplification of liver reperfusion damage. Surface chemiluminescence appears to properly describe the in situ and in vivo progressive organization of the acute inflammatory response with phagocyte-mediated liver injury. PMID- 10706553 TI - Altered hepatic lymphocyte subpopulations in obesity-related murine fatty livers: potential mechanism for sensitization to liver damage. AB - Although obesity-related fatty livers are vulnerable to damage from endotoxin, the mechanisms involved remain obscure. The purpose of this study was to determine if immunologic priming might be involved by determining if fatty livers resemble normal livers that have been sensitized to endotoxin damage by Propionibacterium acnes infection. The latter induces interleukin (IL)-12 and 18, causing a selective reduction of CD4+NK T cells, diminished IL-4 production, deficient production of T-helper type 2 (Th-2) cytokines (e.g., IL-10), and excessive production of Th-1 cytokines (e.g., interferon gamma [IFN-gamma]). Liver and spleen lymphocyte populations and hepatic cytokine production were compared in genetically obese, ob/ob mice (a model for obesity-related fatty liver) and lean mice. Obese mice have a selective reduction of hepatic CD4+NK T cells. Serum IL-18 is also increased basally, and the hepatic mRNA levels of IL 18 and -12 are greater after endotoxin challenge. Thus, up-regulation of IL-18 and IL-12 in fatty livers may reduce hepatic CD4+NK T cells. In addition, mononuclear cells from fatty livers have decreased expression of the adhesion molecule, leukocyte factor antigen-1 (LFA-1), which is necessary for the hepatic accumulation of CD4+NK T cells. Consistent with reduced numbers of hepatic CD4+NK T cells, mononuclear cells from fatty livers produce less IL-4. Furthermore, after endotoxin treatment, hepatic induction of IL-10 is inhibited, while that of IFN-gamma is enhanced. Thus, fatty livers have inherent immunologic alterations that may predispose them to damage from endotoxin and other insults that induce a proinflammatory cytokine response. PMID- 10706554 TI - Oral tolerization ameliorates liver disorders associated with chronic graft versus host disease in mice. AB - In chronic graft versus host disease (cGVHD), an immune attack by transplanted donor lymphocytes results in damage of host target organs. A disbalance between proinflammatory (Th1) and anti-inflammatory cytokines (Th2) plays an important role in the pathogenesis. Immune hyporesponsiveness induced by oral antigen administration has been shown to suppress autoimmunity. We evaluated the efficacy of oral tolerization in preventing cGVHD in a mouse model. cGVHD was generated by infusing 2.5 x 10(7) splenocytes from B10.D2 donor mice, to sublethally irradiated (6 Gy) BALB/c recipient mice, which differ in minor histocompatibility antigens. The transplantation resulted in cGVHD, with characteristic hepatic and small bowel inflammation, and increased skin collagen content and fibrosis. Oral tolerance was induced by feeding donor B10.D2 mice with proteins extracted from BALB/c splenocytes at 50 microg/d per mouse for 11 days before transplantation. Tolerization was evidenced by reduction in mixed lymphocyte response of effector splenocytes from tolerized B10.D2 mice against BALB/c target splenocytes. Liver and small bowel biopsy specimens revealed much less inflammation. Oral tolerization prevented weight and subcutaneous fat loss, reduced thickening, and skin collagen deposits. Reduction of collagen alpha1 (I) gene expression was shown by in situ hybridization. Serum interleukin 10 (IL-10) levels measured significantly higher in tolerized mice than in controls, whereas interferon gamma (IFN-gamma), IL-2, and tumor necrosis factor alpha (TNF-alpha) were reduced significantly. Oral tolerization of splenocyte donors towards recipient-strain splenocytes ameliorated cGVHD of the liver, small intestine, and skin. A cytokine shift from a proinflammatory to an anti-inflammatory pattern may play a role in down-regulation of the immune-mediated target organ damage. PMID- 10706555 TI - Selective blockade of mGlu5 metabotropic glutamate receptors protects rat hepatocytes against hypoxic damage. AB - Western blot analysis of protein extracts from rat liver revealed the presence of the mGlu5 receptor, one of the G-protein-coupled receptors activated by glutamate (named "metabotropic glutamate receptors" or mGlu receptors). mGlu5 expression was particularly high in extracts from isolated hepatocytes, where levels were comparable with those seen in the rat cerebral cortex. The presence of mGlu5 receptors in hepatocytes was confirmed by reverse-transcription polymerase chain reaction (RT-PCR) analysis, immunohistochemistry in neonate or adult rat liver, as well as by immunocytochemical analysis in HepG2 hepatoma cells, where the receptor appeared to be preferentially distributed in cell membranes. Interestingly, mGlu1 receptors (which are structurally and functionally homologous to mGlu5 receptors) were never found in rat liver or hepatocytes. In hepatocytes exposed to anoxic conditions for 90 minutes, glutamate, (1S,3R)-1 aminocyclopentane-1, 3-dicarboxylic acid (1S,3R-ACPD) and quisqualate, which all activate mGlu5 receptors, accelerated the onset and increased the extent of cell damage, while 4-carboxy-3-hydroxyphenylglycine (4C3HPG), an agonist of mGlu2/3 receptors, was inactive. 2-methyl-6-(2-phenyl-1-ethynyl)-pyridine (MPEP), a novel, noncompetitive, highly selective mGlu5 receptor antagonist, not only abolished the toxic effect of 1S,3R-ACPD, but, unexpectedly, was protective by itself against anoxic damage. This suggests that hepatocytes express mGlu5 receptors and that activation of these receptors by endogenous glutamate facilitates the development of anoxic damage in hepatocytes. PMID- 10706556 TI - Activation of nuclear factor kappaB in hepatitis C virus infection: implications for pathogenesis and hepatocarcinogenesis. AB - The hepatitis C virus (HCV) core protein is a multifunctional protein. It may bind to the death domain of tumor necrosis factor receptor 1 (TNFR1) and to the cytoplasmic tail of lymphotoxin-beta receptor, implying that it may be involved in the apoptosis and anti-apoptosis signaling pathways. In vitro studies have been inconclusive regarding its ability to inhibit or enhance TNF-alpha-induced apoptosis. To address this issue, electrophoretic mobility shift assay (EMSA) and immunohistochemical studies were used to show the activation of nuclear factor kappaB (NF-kappaB) in HCV-infected liver tissues and in HCV core-transfected cells. The activation of NF-kappaB was correlated with the apoptosis assays. The results showed that NF-kappaB activation could be shown in HCV-infected livers and HCV core-transfected cells. The data of EMSA correlated with those of immunohistochemical studies, which revealed a higher frequency of NF-kappaB nuclear staining in HCV-infected than in normal livers. NF-kappaB activation conferred resistance to TNF-alpha-induced apoptosis in HCV core-transfected cells. Inhibition of NF-kappaB activation by pyrrolidine dithiocarbamate sensitized them to TNF-alpha-induced apoptosis. These findings suggest that HCV infection may cause anti-apoptosis by activation of NF-kappaB and implicate a mechanism by which HCV may evade the host's immune surveillance leading to viral persistence and possibly to hepatocarcinogenesis. PMID- 10706557 TI - Fas- and tumor necrosis factor receptor 1-dependent but not perforin-dependent pathways cause injury in livers infected with an adenovirus construct in mice. AB - Intravenous injection of type 5 adenovirus, deleted in the E1 and E3 regions, is shown to result in expression of viral antigens in the liver, initiating lymphocyte infiltration and liver injury. Following this infection, induction of Fas ligand (FasL), tumor necrosis factor alpha (TNF-alpha), and perforin mRNA are all demonstrable in the liver, pointing to a role of respective pathways in liver injury. Making use of mice in which the genes coding for Fas, FasL, TNF receptors (TNFRs), and perforin are inactivated, as well as recombinant proteins that inhibit Fas- and TNF-alpha-mediated apoptosis, it is shown that a functional perforin-mediated mechanism is not obligatory for cellular infiltration and progression of liver injury. In contrast functional Fas- and TNF-alpha-mediated mechanisms were found to be essential for liver injury to occur. Results are presented demonstrating that signaling through TNFR1, but not TNFR2, is involved in TNF-alpha-mediated liver damage. The conclusion is drawn that although perforin mRNA is induced in the virus-infected liver, Fas- and TNF-alpha-mediated mechanisms constitute the principal pathways by which the cell-mediated immune system causes acute liver injury. PMID- 10706558 TI - Opening of the mitochondrial permeability transition pore causes matrix expansion and outer membrane rupture in Fas-mediated hepatic apoptosis in mice. AB - Although Fas stimulation has been reported to cause outer mitochondrial membrane rupture in Jurkat cells, the mechanism of this effect is debated, and it is not known if outer membrane rupture also occurs in hepatocyte mitochondria. We studied the in vivo effects of Fas stimulation on ultrastructural lesions and mitochondrial function in mice. Four hours after administration of an agonistic anti-Fas antibody (8 microg/animal), caspase activity increased 5.4-fold. Nuclear DNA showed internucleosomal fragmentation, whereas supercoiled mitochondrial DNA was replaced by circular and linear forms. Mitochondrial cytochrome c was partly released into the cytosol. Ultrastructurally, mitochondrial lesions were observed in both apoptotic hepatocytes (with nuclear chromatin condensation/fragmentation) and nonapoptotic hepatocytes (without nuclear changes). In nonapoptotic cells, outer mitochondrial membrane rupture allowed herniation of the inner membrane and matrix through the outer membrane gap. In apoptotic hepatocytes, the matrix became electron-lucent and no longer protruded through the outer membrane gap. Mitochondria clustered around the nucleus, whereas rough endoplasmic reticulum cisternae became peripheral. In liver mitochondria isolated after Fas stimulation, the membrane potential decreased, whereas basal respiration increased. Pretreatment with either z-VAD-fmk (an inhibitor of caspases) or cyclosporin A (a permeability transition inhibitor) totally or mostly prevented mitochondrial outer membrane rupture, membrane potential decrease, cytochrome c release, and apoptosis. In conclusion, in vivo Fas stimulation causes caspase activation, mitochondrial permeability transition (decreasing the membrane potential and increasing basal respiration), mitochondrial matrix expansion (as shown by matrix herniation), outer mitochondrial membrane rupture, and cytochrome c release. PMID- 10706559 TI - Zonal down-regulation and redistribution of the multidrug resistance protein 2 during bile duct ligation in rat liver. AB - We have studied regulation of the multidrug resistance protein 2 (mrp2) during bile duct ligation (BDL) in the rat. In hepatocytes isolated after 16, 48, and 72 hours of BDL, mrp2-mediated dinitrophenyl-glutathione (DNP-GS) transport was decreased to 65%, 33%, and 33% of control values, respectively. The impaired mrp2 mediated transport coincided with strongly decreased mrp2 protein levels, without any significant changes in mrp2 RNA levels. Restoration of bile flow after a 48 hour BDL period resulted in a slow recovery of mrp2-mediated transport and protein levels. Immunohistochemical detection of the protein in livers of rats undergoing BDL showed strongly reduced mrp2 staining after 48 hours, which was initiated in the periportal areas of the liver lobule and progressed toward the pericentral areas after 96 hours. Immunofluorescent detection of mrp2 in livers of rats undergoing 48 hours of BDL revealed decreased staining accompanied by intracellular localization of the protein in pericanalicular vesicular structures. Within this intracellular compartment, mrp2 colocalized with the bile salt transporter (bsep) and was still active as shown by vesicular accumulation of the fluorescent organic anion glutathione-bimane (GS-B). We conclude that down regulation of mrp2 during BDL-induced obstructive cholestasis is mainly posttranscriptionally regulated. We propose that this down-regulation is caused by endocytosis of apical transporters followed up by increased breakdown of mrp2, probably in lysosomes. This breakdown of mrp2 is more severe in the periportal areas of the liver lobule. PMID- 10706560 TI - Contribution of reduced insulin sensitivity and secretion to the pathogenesis of hepatogenous diabetes: effect of liver transplantation. AB - Diabetes mellitus frequently complicates cirrhosis but the pathogenic mechanisms are unknown. To assess the contribution of reduced insulin action and secretion, 24 cirrhotic-diabetic patients waiting for liver transplant because of an unresectable hepatocarcinoma underwent an oral glucose tolerance test (OGTT) to assess the beta-cell function and an insulin clamp combined with [3-(3)H]glucose infusion to measure whole body glucose metabolism before and 2 years after the transplant. Seven cirrhotic nondiabetic patients, 11 patients with chronic uveitis on similar immunosuppressive therapy, and 7 healthy subjects served as control groups. Cirrhotic patients showed a profound insulin resistance, and diabetics in addition also showed increased endogenous glucose production (P <.05) and insulin deficiency during the OGTT (P <.05). Liver transplantation normalized endogenous glucose production and insulin sensitivity but failed to cure diabetes in 8 of the 24 patients because a markedly low insulin response during the OGTT. Age, body mass index, family history of diabetes, immunosuppressive drugs, and pathogenesis of cirrhosis did not predict in whom liver transplant was going to cure diabetes. On the contrary, a reduced secretory response characterized the patients in whom the transplant would not be curative. In summary, insulin resistance was a primary event complicating cirrhosis but additional beta-cell secretory defects were crucial for development of diabetes. Liver transplantation, lessening insulin resistance, cured hepatogenous diabetes in 67% of cirrhotic-diabetic patients; nevertheless 33% were still diabetics because the persistence of a reduced beta-cell function, which makes these patients eventually eligible for combined islet transplantation. PMID- 10706561 TI - Novel molecular defects of the delta-aminolevulinate dehydratase gene in a patient with inherited acute hepatic porphyria. AB - Cloning and expression of the defective gene for delta-aminolevulinate dehydratase (ALAD) from the second of 2 German patients with ALAD deficiency porphyria (ADP), who had been originally reported by Doss et al. in 1979, were performed. Cloning of cDNAs for the defective ALAD were performed using Epstein Barr virus (EBV)-transformed lymphoblastoid cells of the proband, and nucleotide sequences of cloned cDNA were determined. Two separate mutations of ALAD cDNA were identified in each ALAD allele. One was G457A, termed "H1," resulting in V153M substitution, while the other was a deletion of 2 sequential bases at T(818) and C(819), termed "H2," resulting in a frame shift with a premature stop codon at the amino acid position of 294. Using allele-specific oligonucleotide hybridization, the mother of the proband was shown to have an H1 defect, while using genomic DNA analysis, the father was shown to have an H2 defect. Expression of H1 cDNA in Chinese hamster ovary cells produced an ALAD protein with only a partial activity (10.65% +/- 1.80% of the normal), while H2 cDNA encoded no significant protein. These data thus demonstrate that the proband was associated with 2 novel molecular defects of the ALAD gene, 1 in each allele, and account for the extremely low ALAD activity in his erythrocytes ( approximately 1% of normal). PMID- 10706562 TI - Activation of N-methyl-D-aspartate receptors in rat brain in vivo following acute ammonia intoxication: characterization by in vivo brain microdialysis. AB - Ammonia is considered the main agent responsible for the neurological alterations in hepatic encephalopathy. It was suggested that ammonia toxicity is mediated by activation of N-methyl-D-aspartate (NMDA) receptors. The aim of this work was to assess, by in vivo brain microdialysis in freely moving rats, whether acute ammonia intoxication leads to activation of NMDA receptors in the cerebellum of the rat in vivo. We measured the effects of ammonia intoxication on the neuronal glutamate-nitric oxide-cyclic guanosine monophosphate (cGMP) pathway, by measuring the ammonia-induced increase of extracellular cGMP. Ammonia intoxication increases extracellular cGMP, and this increase is prevented by (5R,10S)-5-methyl-10,11-dihydro-5H-dibenzo[a, d]cyclohepten-5,10-imine hydrogen maleate (MK-801). There is a good correlation between the increase in cGMP and the seriousness of the neurological symptoms elicited by different doses of ammonia. Ammonia doses inducing coma did not affect extracellular glutamate, while doses leading to death increased it by 349%. The time courses of ammonia induced increases in extracellular ammonia, cGMP, and glutamate indicate that NMDA receptor activation occurs before the increase in extracellular glutamate. Ammonia-induced increase in glutamate is prevented by MK-801. These results indicate that ammonia intoxication leads to activation of NMDA receptors in the animal in vivo, and that this activation is not caused by increased extracellular glutamate. The possible underlying mechanism is discussed. PMID- 10706563 TI - Mutations of the core promoter and response to interferon treatment in chronic replicative hepatitis B. AB - In chronic replicative hepatitis B the significance of mutations in the basic core promoter (BCP), core upstream regulatory sequences (CURS) and negative regulatory element (NRE) for response to interferon (IFN) is unknown. A sequence analysis of the NRE, CURS, BCP, and precore region was performed from sera of 96 patients with chronic replicative hepatitis B (64 hepatitis B e antigen [HBeAg] positive patients and 32 HBeAg-negative patients) treated with alfa-IFN (IFN alpha). The overall sustained response (SR) rate to IFN was 30% with no significant difference between HBeAg-positive and HBeAg-negative patients. IFN responsiveness correlated to hepatitis B virus (HBV)-DNA levels, hepatitis B surface antigen (HBsAg) levels, the number of mutations in the complete BCP, especially nucleotide (nt) region 1753 to 1766 and mutations at nt 1762 and 1764. In HBeAg-positive hepatitis, SR to IFN was associated with a high number of mutations in the BCP (P <.04) and nucleotide region 1753 to 1766 (P <.015) as well as mutations at nucleotide 1764 (P <.007). In HBeAg-negative hepatitis, SR to IFN correlated with a low number of mutations in the BCP (P <.04) and nucleotide region 1753 to 1766 (P <.02) and a wild-type sequence at nt 1764 (P <.003). Prediction of IFN response was possible on the basis of nt 1764 in 77% of HBeAg-positive patients and 78% of HBeAg-negative patients. IFN response did not correlate with the occurrence of the 1896 mutation, mutations in the CURS or NRE, disease duration, ethnic origin of the patient, alanine transaminase (ALT) levels and HBV genotype. Our data suggest that HBV genome mutations located within the BCP are determinants of a response to IFN therapy. PMID- 10706564 TI - A new step toward standardization of serum hepatitis C virus-RNA quantification in patients with chronic hepatitis C. AB - The need to improve efficacy of antiviral therapy for chronic hepatitis C has prompted the development of quantitative assays, which allows the assessment of viral load before therapy. The aim of our study was to evaluate the clinical relevance of 3 serum HCV-RNA quantitative assays in 87 patients with chronic hepatitis C, the noncommercially available SuperQuant assay (National Genetic Institute), recently used in large international controlled trials, the most early and widely used Quantiplex HCV RNA v2.0 assay (branched DNA [bDNA] v2.0; Bayer Diagnostics, Puteaux, France), and the new generation Cobas Amplicor HCV Monitor assay (COBAS v2.0; Roche Diagnostics Systems, Meylan, France), which is a semiautomated reverse transcription-polymerase chain reaction (RT-PCR) assay. The level and range of quantification were similar between all assays and a strong correlation was observed over all HCV genotypes among the assays. Recent publications have suggested that the baseline cut-off level of 2 x 10(6) copies/mL, as determined by the SuperQuant assay, is able to discriminate between patients with low viral load from those with high viral load and can be used to predict responses to therapy. Because all 3 assays use different testing technologies we examined how many of our patients fell above this defined cut-off level when tested by the other assays; of 22 patients measured below 2 x 10(6) copies/mL with the SuperQuant assay, 17 of 22 and 19 of 22 patients were eligible with the bDNA v2. 0 and the COBAS v2.0 assays, respectively (P >.05). Our results indicate that the 2 commercial assays can be used to determine treatment schedules in patients with chronic hepatitis C, providing a flexibility in multicenter controlled trials by offering better accessibility of test results. PMID- 10706565 TI - Iron reduction before and during interferon therapy of chronic hepatitis C: results of a multicenter, randomized, controlled trial. AB - Patients with chronic hepatitis C and low serum and hepatic iron stores may have an improved response to interferon (IFN). We tested whether iron reduction before and during IFN therapy would lead to an improved sustained biochemical and virological response compared with IFN alone. Eighty-two previously untreated patients with chronic hepatitis C were randomized to either: group A IFN-alpha2b 3 MU 3 times per week for 6 months, or group B iron reduction before and during IFN-alpha2b 3 MU 3 times per week for 6 months. Group B patients had lower mean serum alanine transaminase (ALT) levels than group A patients during treatment and follow-up. Group B patients had significantly lower mean hepatitis C virus (HCV)-RNA levels at treatment weeks 4 and 12 (P <.05). Serum HCV RNA was undetectable at the end of treatment in 15 group B patients compared with 7 group A patients (P =.03); 7 group B patients and 3 group A patients had persistently undetectable serum HCV RNA 24 weeks after the end of therapy (P =.20). Paired pre and posttreatment liver biopsies in 18 group B patients demonstrated significant improvements in 2 of the 3 inflammation scores of the Knodell histological activity index (P <. 05). No changes occurred in the paired biopsies from 15 group A patients. We conclude that iron reduction via therapeutic phlebotomy improves the end-of-treatment virological and histological response to short-term IFN therapy. Additional studies are needed to determine if iron reduction in combination with higher doses or longer duration of IFN may be of benefit. PMID- 10706566 TI - Hepatitis C virus (HCV) infection and cryoglobulinemia: analysis of whole blood and plasma HCV-RNA concentrations and correlation with liver histology. AB - The influence of cryoprecipitate (CP) on liver histology and peripheral titers of hepatitis C virus (HCV) RNA was evaluated for 115 patients with chronic hepatitis. Fifty-four patients had measurable CP levels whereas 61 did not. Assessment of liver biopsies for grade of fibrosis revealed that patients with CP had increased fibrosis (P <.001) and incidence of cirrhosis (P =.001) compared with those without CP. In contrast, there was not a significant difference in the inflammatory activity score between the 2 groups. HCV RNA in whole blood (WB) and plasma (Pl) was evaluated in patients with or without CP by end-point-limiting dilution titer. Among patients with CP, WB titers were significantly higher than Pl titers (P <.001); however, there was no difference in WB or Pl titers in patients without CP (P =.068). Histological activity and fibrosis scores of patients from either group were compared with peripheral viral titers of WB and Pl, percentage of CP, rheumatoid factor (RF) titer, and serum alanine transaminase (ALT). There were significant correlations between the amount of fibrosis and the percentage of CP and rheumatoid factor titer, yet neither of the latter parameters was correlated with inflammatory activity. These data suggest that patients with CP and chronic hepatitis owing to HCV are more likely to have progressive disease than patients without CP. Furthermore, the presence of CP in patients infected with HCV appears to influence the amount of virus detected in patient Pl, suggesting that WB assays may be more reliable for HCV-RNA quantitation in patients with CP. PMID- 10706567 TI - Characteristics of hepatitis C viral genome associated with disease progression. AB - The clinical presentations of chronic hepatitis C are not uniform. Some patients show persistently high serum alanine transaminase (ALT) values and develop liver cirrhosis and hepatocellular carcinoma (HCC), whereas serum ALT values stay normal in other patients. The mechanism causing this diversity remains to be elucidated. The aim of this study was to identify genomic characteristics of hepatitis C virus (HCV) genotype 1b associated with disease progression. Full length sequences of HCV were determined in 14 patients who showed persistently normal serum ALT values (normal ALT group) and 13 cirrhotics with HCC (HCC group). Residues in which amino acid usage was different between these 2 groups were extracted, and Progression score was defined as the total number of residues with 7 amino acids, more frequently present in the HCC group than in the normal ALT group. In the validation of this Progression score in 9 patients with normal ALT and 25 with HCC, the score was significantly higher in the HCC group (3.1 +/- 1.1 vs. 2.0 +/- 0.9, P =.019). Finally, the correlation between the score and clinical markers related to disease progression was analyzed. In a total of 107 patients with chronic HCV infection, the Progression score was correlated significantly with platelet counts (r = -0.31, P =.0024) by multivariate analysis. In conclusion, high Progression scores were associated with the presence of HCC and low platelet counts. Sequences of the HCV-1b genome may be related to the progression of chronic hepatitis C. PMID- 10706568 TI - Prevalence and clinical course of chronic hepatitis C virus (HCV) infection and rate of HCV vertical transmission in a cohort of 15,250 pregnant women. AB - The prevalence and natural course of chronic hepatitis C virus (HCV) infection was evaluated in 15,250 consecutive pregnant women. The rate of HCV vertical and perinatal transmission was also assessed. The presence of anti-HCV was tested by means of EIA III and confirmed by recombinant immunoblot assay III. Alanine transaminase (ALT), anti-human immunodeficiency virus (HIV), and HCV-RNA were tested during the first month and third trimester of pregnancy, and 6 months after delivery; the same tests were made in all of the newborns of anti-HCV positive mothers at birth (on cord blood samples) and then at 4-month intervals. Anti-HCV positivity was found in 370 cases (2.4%), 72% of whom were also HCV-RNA positive. The proportion of women with hypertransaminases decreased from 56.4% at the first examination during the first month of pregnancy to 7.4% in the last trimester, and then increased again after delivery (54. 5%), without any concomitant changes in the proportion of those with viremia. The proportion of anti-HCV- and HCV-RNA-positive newborns was 5.1% after 1 year (8 of 155), all of whom had the same genotype as their mother. The rate of HCV transmission was not affected by the type of delivery or feeding, or the HIV status of the mother. The results of this large-scale study confirm previous data in smaller series concerning the prevalence of HCV infection in pregnant women, and strongly support the hypothesis of a favorable (possibly immunomediated) effect of pregnancy on liver cell necrosis in anti-HCV-positive women. PMID- 10706569 TI - Risk factors for hepatitis C virus infection in United States blood donors. NHLBI Retrovirus Epidemiology Donor Study (REDS) AB - Injection drug use (IDU) is a known risk factor for hepatitis C virus (HCV) infection, but the strength of other parenteral and sexual risk factors is unclear. In 1997, we performed a case-control study of 2,316 HCV-seropositive blood donors and 2,316 seronegative donors matched on age, sex, race/ethnicity, blood center, and first-time versus repeat-donor status. Odds ratios (OR) and 95% confidence intervals (CIs) were calculated using conditional logistic regression. Questionnaires were returned by 758 (33%) HCV(+) and 1,039 (45%) control subjects (P =.001). The final multivariate model included only the following independent HCV risk factors: IDU (OR = 49.6; 95% CI: 20.3-121.1), blood transfusion in non IDU (OR = 10.9; 95% CI: 6.5-18.2), sex with an IDU (OR = 6.3; 95% CI: 3.3-12.0), having been in jail more than 3 days (OR = 2.9; 95% CI: 1.3-6.6), religious scarification (OR = 2.8; 95% CI: 1.2-7. 0), having been stuck or cut with a bloody object (OR = 2.1; 95% CI: 1.1-4.1), pierced ears or body parts (OR = 2.0; 95% CI: 1.1-3.7), and immunoglobulin injection (OR = 1.6; 95% CI: 1.0-2.6). Although drug inhalation and a high number of lifetime sex partners were significantly more common among HCV seropositives, they were not associated with HCV after controlling for IDU and other risk factors. IDU, blood transfusion among non-IDU, and sex with an IDU are strong risk factors for HCV among United States blood donors. Weaker associations with incarceration, religious scarification, being stuck or cut with a bloody object, pierced ears or body parts, and immunoglobulin injection must be interpreted with caution. PMID- 10706570 TI - Hepatitis B e antigen-negative chronic hepatitis B in Hong Kong. AB - Hepatitis B e antigen-negative chronic hepatitis B (e-CHB) has been reported in Asia but its prevalence and clinical significance have not been determined. The aims of this study were to determine the prevalence of e-CHB in Hong Kong and the frequency of precore and core promoter mutations in these patients. A cross sectional study was performed in 350 consecutive Chinese patients (230 men and 120 women; mean age +/-SD, 42 +/- 13 years) with chronic hepatitis B virus infection. A total of 243 (69%) patients were hepatitis B e antigen (HBeAg) negative of whom 15% had clinical cirrhosis. In the remaining 85% of patients, 63% had normal and 22% had elevated transaminases. Serum hepatitis B virus (HBV) DNA was detectable using branched DNA assay in 46% of HBeAg-negative patients with clinical cirrhosis/elevated transaminases. Forty-five percent of the patients with e-CHB had the precore stop codon mutation, and an additional 41% had core promoter changes. There was no correlation between the presence of precore/core promoter mutations and liver disease or HBV-DNA levels. Overall, 17% of HBeAg-negative patients were viremic and had evidence of chronic liver disease (e-CHB) with mean HBV-DNA levels comparable with that in HBeAg-positive patients. In summary, we found that e-CHB may be present in up to 17% of HBeAg-negative patients seen in a tertiary referral center in Hong Kong. e-CHB may be a heterogeneous condition and is not invariably associated with the precore HBV mutant. Population studies are needed to determine the true prevalence of e-CHB in Asia and to assess its natural course and response to treatment. PMID- 10706571 TI - Inhibition of hepatitis C virus (HCV)-RNA-dependent translation and replication of a chimeric HCV poliovirus using synthetic stabilized ribozymes. AB - Ribozymes are catalytic RNA molecules that can be designed to cleave specific RNA sequences. To investigate the potential use of synthetic stabilized ribozymes for the treatment of chronic hepatitis C virus (HCV) infection, we designed and synthesized hammerhead ribozymes targeting 15 conserved sites in the 5' untranslated region (UTR) of HCV RNA. This region forms an internal ribosome entry site that allows for efficient translation of the HCV polyprotein. The 15 synthetic ribozymes contained modified nucleotides and linkages that stabilize the molecules against nuclease degradation. All 15 ribozymes were tested for their ability to reduce expression in an HCV 5' UTR/luciferase reporter system and for their ability to inhibit replication of an HCV-poliovirus (HCV-PV) chimera. Treatment with several ribozymes resulted in significant down-regulation of HCV 5' UTR/luciferase reporter expression (range 40% to 80% inhibition, P <.05). Moreover, several ribozymes showed significant inhibition (>90%, P <.001) of chimeric HCV-PV replication. We further show that the inhibitory activity of ribozymes targeting site 195 of HCV RNA exhibits a sequence-specific dose response, requires an active catalytic ribozyme core, and is dependent on the presence of the HCV 5' UTR. Treatment with synthetic stabilized anti-HCV ribozymes has the potential to aid patients who are infected with HCV by reducing the viral burden through specific targeting and cleavage of the viral genome. PMID- 10706572 TI - The past incidence of hepatitis C virus infection: implications for the future burden of chronic liver disease in the United States. AB - Because chronic liver disease may develop many years after acute hepatitis C virus (HCV) infection, the past incidence of acute infections is a major determinant of the future burden of HCV-associated complications. We estimated past incidence of acute HCV infection using national seroprevalence data and relative age-specific incidence data from a sentinel counties surveillance system. Projections of the future prevalence of HCV-infected patients were derived from models that included an 85% drop in HCV infection incidence as observed for reported cases in the early 1990s. The models showed a large increase in the incidence of HCV infections from the late 1960s to the early 1980s. The degree of increase was dependent on the assumed rate of antibody loss; a model with 2.5% annual antibody loss showed annual incidence increasing from 45,000 infections (95% confidence interval [95% CI]: 0-110,000) in the early 1960s to 380,000 infections (95% CI: 250,000 to 500, 000) in the 1980s. Projections showed that although the prevalence of HCV infection may be declining currently because of the decline in incidence in the 1990s, the number of persons infected for >/=20 years could increase substantially before peaking in 2015. If the incidence of new HCV infections does not increase in the future, persons born between 1940 and 1965 will be at highest lifetime risk of acquiring the infection. PMID- 10706573 TI - Insulin growth factor I and hypogonadism in cirrhosis. PMID- 10706574 TI - Nuclear factor kappaB and hepatitis C--Is there a connection? PMID- 10706575 TI - Standardization of hepatitis C virus--RNA quantification: advances and unfinished business. PMID- 10706576 TI - Hepatitis C epidemiology: injecting new tools in the field. PMID- 10706577 TI - Update of the International Banff Schema for Liver Allograft Rejection: working recommendations for the histopathologic staging and reporting of chronic rejection. An International Panel. PMID- 10706578 TI - The 20th United States-Japan Joint Hepatitis Panel Meeting. PMID- 10706579 TI - Differential response to low-fat diet between low and normal HDL-cholesterol subjects. AB - Heart attacks frequently occur in normolipidemic subjects with low concentration of high density lipoproteins (35 mg/dL). We hypothesized that as subjects with low HDL-C already have low HDL concentrations, the major decrease of HDL-C will occur in subjects with normal HDL-C when a low-fat diet is consumed. Normolipidemic male subjects consumed three diets differing in total fat and saturated fat composition (AAD: 37%, Step-1: 28%, Step-2: 24% total fat) for 6 weeks in a three-period double-blind randomized crossover design. Plasma lipids and apolipoproteins were determined and changes in distribution of HDL subpopulations were evaluated. As a result of a low-fat diet, low HDL-C individuals slightly decreased their HDL-C, but substantially decreased their LDL C resulting in a significant improvement in the LDL-C/HDL-C ratio. However, subjects with normal HDL-C levels decreased both their LDL-C and HDL-C resulting in an unchanged LDL-C/HDL-C ratio. We also observed significant differences in response to low-fat diets in HDL-C and alpha(1) concentrations between low and normal HDL-C subjects. In the normal HDL-C group, consumption of a low-fat diet also resulted in redistribution of apoA-I-containing HDL subpopulations, indicated by a decrease in the large apoA-I-only alpha(1) subpopulation. These data demonstrate that male subjects with low HDL-C respond to a low-fat diet differently than individuals with normal HDL-C. PMID- 10706580 TI - Increased chylomicron triglyceride hydrolysis by connective tissue flow in perfused rat hindlimb. Implications for lipid storage. AB - Skeletal muscle has two circulatory routes, nutritive (in contact with muscle) and non-nutritive (part of which is located in the connective tissue), and the balance of flow between the two is controlled by neural input and circulating vasomodulators. The purpose of this study was to assess muscle triglyceride hydrolysis given that the two circuits may have a differing vascular distribution of hydrolytic activity. The isolated rat hindlimb was perfused with 6% Ficoll((R)) and a radiolabeled chylomicron;-lipid emulsion containing apolipoprotein C-II. Serotonin (0.5-1 micrometer), a model vasoconstrictor previously shown to preferentially increase connective tissue flow, inhibited hindlimb oxygen uptake (from 16.7 +/- 0.6 to 10.2 +/- 1.0, mean +/- SE, n = 7 (P <0.001)) and stimulated [(14)C]-labeled fatty acid uptake into muscles (from 184 +/- 28 to 602 +/- 132, mean +/- SE, n = 7 (P = 0.009)). These effects were reversed by the vasodilator carbamyl choline. Vasopressin resulted in increased oxygen consumption but no change in triglyceride hydrolysis. Cholesteryl oleate uptake (an indicator of endocytosis of the chylomicron or remnant particle) was unaltered by serotonin. It is concluded that chylomicron triglyceride hydrolysis is enhanced by vasoconstrictors that increase connective tissue flow in the perfused rat hindlimb. Increased hydrolysis appears to be primarily due to an increased access of triglyceride to hydrolytic enzymes, presumably lipoprotein lipase associated with the fat cells commonly observed interlaced amongst bundles of muscle fibers. PMID- 10706581 TI - Peroxisomal fatty acid oxidation disorders and 58 kDa sterol carrier protein X (SCPx). Activity measurements in liver and fibroblasts using a newly developed method. AB - Sterol carrier protein X (SCPx) plays a crucial role in the peroxisomal oxidation of branched-chain fatty acids. To investigate whether patients with an unresolved defect in peroxisomal beta-oxidation are deficient for SCPx, we developed a novel and specific assay to measure the activity of SCPx in both liver and fibroblast homogenates. The substrate used in the assay, 3alpha, 7alpha,12alpha-trihydroxy 24-keto-5beta-cholestanoy l-CoA (24-keto-THC-CoA), is produced by preincubating the enoyl-CoA of the bile acid intermediate THCA with a lysate from the yeast Saccharomyces cerevisiae expressing human D-bifunctional protein. After the preincubation period, liver or fibroblast homogenate is added plus CoASH, and the production of choloyl-CoA is determined by HPLC. The specificity of the assay was demonstrated by the finding of a full deficiency in fibroblasts from an SCPx knock-out mouse. In addition to SCPx activity measurements in fibroblasts from patients with a defect in peroxisomal beta-oxidation of unresolved etiology, we studied the stability and activity of SCPx in fibroblasts from patients with Zellweger syndrome, which lack functional peroxisomes. We found that SCPx is not only stable in the cytosol, but displays a higher activity in fibroblasts from patients with Zellweger syndrome than in control fibroblasts. Furthermore, in all patients studied with a defect in peroxisomal beta-oxidation of unknown origin, SCPx was found to be normally active, indicating that human SCPx deficiency remains to be identified. PMID- 10706582 TI - Expression and microvillar localization of scavenger receptor class B, type I (SR BI) and selective cholesteryl ester uptake in Leydig cells from rat testis. AB - This study investigates the relationship between the high density lipoprotein (HDL) receptor (scavenger receptors, SR-BI and SR-BII), selective lipoprotein cholesteryl ester uptake, and testosterone production in Leydig cells of control, hypocholesterolemic and gonadotrophic hormone (hCG) treated rats. Leydig cells from mature control rats show poor efficiency in incorporation of labeled HDL cholesteryl esters into testosterone, poor selective uptake of lipoprotein lipids overall, and a dramatic reduction of circulating levels of lipoproteins has no apparent effect on testosterone production or expression of intracellular enzymes synthesizing cholesterol. Leydig cells from control rats show minimal levels of SR-BI and SR-BII. However, similarly aged rats treated with hCG for several days undergo changes consistent with hormone-desensitization. Despite the resulting low levels of testosterone production, SR-BI levels are dramatically increased, Leydig cells now efficiently internalize HDL-supplied cholesteryl esters by the selective cholesterol uptake process, and various other cholesterol-sensitive genes of the cells are up-regulated. Only SR-BII expression remains negligible and unchanged throughout this period. It is of interest that Leydig cell SR-BI of hCG-treated rats is localized in surface microvilli, but is present also in an elaborate and complex channel system within the cytoplasm of the cells. In summary, Leydig cells differ from other rat steroidogenic cells in not depending on exogenous lipoprotein-cholesterol during periods of normal steroid hormone production. However, trophic hormone desensitization is accompanied by increased Leydig cell SR-BI expression and increased selective HDL-cholesteryl ester uptake, presumably in preparation for renewed testosterone production. PMID- 10706583 TI - Studies of the metabolism of alpha-tocopherol stereoisomers in rats using [5 methyl-(14)C]SRR- and RRR-alpha-tocopherol. AB - We investigated the distribution and metabolism of SRR-alpha-tocopherol (SRR alpha-Toc), synthetic alpha-Toc compared with RRR-alpha-Toc, in rats after a single oral administration of 2 mg (20 microCi) SRR- and RRR-alpha-[5-methyl (14)C]Toc. In the liver, there was no difference in the recovery of radioactivity until 12 h after administration, and it reached a maximum of 4.4% of the dose after 12 h, but in other tissues, radioactivity derived from RRR-alpha-Toc was clearly higher than that derived from SRR-alpha-Toc after 12 h. For 96 h after administration, urinary excretions of SRR-alpha-Toc were 7.8% of the dose and significantly greater than that of RRR-alpha-Toc, which was 1.3% of the dose. On the other hand, total fecal excretions of SRR- and RRR-alpha-Toc were 87.6% and 83.0%, respectively. Therefore, radioactivity in the urine was assumed to have transferred out of the liver. Furthermore, the urine samples were hydrolyzed with 3 N methanolic HCl and analyzed by high performance liquid chromatography (HPLC) and liquid chromatography/mass spectrometry. The results showed that about 73% of the total radioactivity injected into HPLC was found to be 2,5,7, 8-tetramethyl-2 (2'-carboxyethyl)-6-hydroxy chroman (alpha-CEHC), as well as RRR-alpha-Toc. Thus, there is no difference between SRR-alpha-Toc and RRR-alpha-Toc in metabolic pathways, and it is suggested that SRR-alpha-Toc discriminated in the liver is rapidly metabolized by the liver and excreted as the conjugate of alpha-CEHC in the urine. PMID- 10706584 TI - Effects of continuous conjugated estrogen and micronized progesterone therapy upon lipoprotein metabolism in postmenopausal women. AB - The effects of continuously administering both conjugated equine estrogens (CEE) and micronized progesterone (MP) on the concentration, composition, production and catabolism of very low density (VLDL) and low density lipoproteins (LDL) have not previously been reported. The mechanism of the hormonally induced reductions of plasma LDL cholesterol of S(f) 0;-20 (mean 16%, P < 0.005) and LDL apoB (mean 6%, P < 0.025) were investigated by studying the kinetics of VLDL and LDL apolipoprotein (apo) B turnover after injecting autologous (131)I-labeled VLDL and (125)I-labeled LDL into each of the 6 moderately hypercholesterolemic postmenopausal subjects under control conditions and again in the fourth week of a 7-week course of therapy (0.625 mg/d of CEE + 200 mg/d of MP). The combined hormones significantly lowered plasma LDL apoB by increasing the mean fractional catabolic rate of LDL apoB by 20% (0. 32 vs. 0.27 pools/d, P < 0.03). Treatment also induced a significant increase in IDL production (6.3 vs. 3.7 mg/kg/d, P = 0.028). However, this did not result in an increase in LDL production because of an increase in IDL apoB direct catabolism (mean 102%, P = 0.033). VLDL kinetic parameters were unchanged and the concentrations of plasma total triglycerides (TG), VLDL-TG, VLDL-apoB did not rise as often seen with estrogen alone. Plasma HDL-cholesterol rose significantly (P < 0.02). Our major conclusion is that increased fractional catabolism of LDL underlies the LDL-lowering effect of the combined hormones. PMID- 10706585 TI - Interleukin-4 augments acetylated LDL-induced cholesterol esterification in macrophages. AB - Activated subpopulations of lymphocytes and mast cells have been detected in atherosclerotic lesions. Interleukin-4 (IL-4) is a prominent cytokine released during activation of both cell types and its transcripts have been detected in both human and mouse atherosclerotic lesions. To define whether this local release of IL-4 influences macrophage lipid metabolism, we examined the effects of this cytokine on intracellular cholesterol esterification during incubation with modified low density lipoprotein (LDL). IL-4 greatly augmented cholesterol esterification induced by acetylated LDL (AcLDL) in both mouse peritoneal macrophages and the murine macrophage cell line, J774. This augmentation was maximal at a concentration of 1 ng/ml after incubation for 48 h. This was not a generalized effect on lipoprotein metabolism as IL-4 had no effect on cholesterol esterification in the presence of either LDL or beta-VLDL. Determination of binding isotherms demonstrated that IL-4 increased the number of cell surface binding sites for AcLDL. The IL-4-augmented AcLDL-induced cholesterol esterification was attenuated by the scavenger receptor class A (SR-A) antagonist, fucoidan, and the anti-mouse SR-A monoclonal antibody, 2F8. These data, combined with the known receptor specificity of AcLDL interactions, imply a role of SR-A in the IL-4 induced responses. Two cytokines that have been demonstrated previously to down-regulate SR-A, TNF-alpha and TGF-beta, antagonized the IL-4-induced augmentation of cholesterol esterification. Therefore, local release of IL-4 within atherosclerotic lesions could have a profound effect on macrophage lipid metabolism and the subsequent atherogenic process. PMID- 10706586 TI - Determination of the tissue sites responsible for the catabolism of large high density lipoprotein in the African green monkey. AB - In vivo multicompartmental modeling of the turnover of HDL subfractions has suggested that HDL containing four molecules of apoA-I per particle and no other apolipoproteins (large LpA-I) are terminal particles in plasma. We hypothesized that these terminal particles were the end product of HDL metabolism and, as such, would be cleared preferentially by the liver. Thus, the purpose of this study was to determine: 1) the tissue sites of catabolism of large LpA-I in African green monkeys, and 2) whether saturated versus n;-6 polyunsaturated dietary fat affected tissue accumulation. Large LpA-I were isolated, without ultracentrifugation, by size exclusion and immunoaffinity chromatography and radiolabeled with either the residualizing compound, (125)I-labeled tyramine cellobiose (TC), or with (131)I. After injection into recipient animals, the plasma die-away of the radiolabels was followed for 12 or 24 h, after which the animals were killed and tissues were collected for determining radiolabel sites of catabolism. The plasma die-away of the (125)I-labeled TC-LpA-I and (131)I labeled LpA-I doses was similar suggesting that the TC radiolabeling did not modify the metabolism of the large LpA-I dose. The liver, adrenal, kidney, and spleen had the greatest accumulation of large LpA-I degradation products on a per gram tissue basis. On a whole organ basis, the liver was the major site of large LpA-I degradation in both the 12-h (15.4 +/- 0.3% of injected dose) and 24-h (9.1 +/- 0.6% of injected dose) catabolic studies. The kidney, compared to the liver, had less uptake of large LpA-I radioactivity in either study (1.3 +/- 0.4% and 1.2 +/- 0.3% of injected dose). There was no apparent influence of dietary fat type on the tissue accumulation of large LpA-I. We conclude that the liver is the primary site of catabolism of large LpA-I in the African green monkey. PMID- 10706587 TI - Head group-independent interaction of phospholipids with bile salts. A fluorescence and EPR study. AB - Bile salts are essential for phospholipid secretion into the bile. To study the relevance of the structure of phospholipids for their interaction with bile salts, we used spin-labeled or fluorescent phospholipid analogues of different head groups and acyl chain length. Those analogues form micelles in aqueous suspension. Their solubilization by bile salts resulting in the formation of mixed micelles was followed by the decrease of spin-spin interaction of spin labeled analogues or by the relief of fluorescence self-quenching of (7-nitro-2 1,3-benzooxadiazol (NBD))-labeled analogues. Solubilization of analogue micelles occurred at and above the critical micellar concentration (CMC) of the bile salts. As revealed by stopped-flow technique, solubilization of NBD-analogues was very rapid with half times as low as 0.1 sec above the CMC of taurocholate. Both kinetics and extent of solubilization were independent of the phospholipid head group, but were significantly affected by the fatty acid chain length. Furthermore, using vesicles with varying phospholipid composition and different types of analogues in self-quenching concentrations, we could show that bile salt mediated vesicle solubilization depended on the fatty acid chain length of phospholipids. In contrast, neither for phospholipids nor for analogues could an influence of the lipid head group on the solubilization process be observed. These findings support a head group-independent mechanism of bile salt-mediated enrichment of specific phospholipids in the bile fluid. PMID- 10706588 TI - Differential effects of interferon-gamma on metabolism of lipoprotein immune complexes mediated by specific human macrophage Fcgamma receptors. AB - The objectives were to determine whether there are differences in the mechanisms of lipoprotein metabolism associated with different FcgammaRs and how metabolism associated with FcgammaRs compares to that mediated by scavenger receptors (SRA). To analyze lipoprotein metabolism in a receptor-specific manner, bispecific antibodies were used to target low density lipoproteins (LDL) labeled with (125)I or [(3)H]cholesterol linoleate to FcgammaRI or FcgammaRIIA in human macrophages. Interferon-gamma (IFN-gamma), which stimulates expression of FcgammaRI while inhibiting expression of SRA, was used to help delineate differences in metabolism between each receptor. For each receptor, the total amount of lipoprotein degradation paralleled changes in receptor expression induced by IFN gamma. In particular, while SRA-mediated degradation typically exceeded degradation mediated by FcgammaRI, in IFN-gamma-treated cells degradation associated with FcgammaRI and SRA was similar. Assay of [(3)H]cholesterol linoleate-labeled lipoproteins indicated that total uptake and hydrolysis of [(3)H]cholesterol linoleate was similar for each class of receptor, and inhibited by IFN-gamma. For FcgammaRI versus FcgammaRIIA, in the presence or absence of IFN gamma, the [(3)H]cholesterol derived from FcgammaRIIA-mediated uptake was preferentially targeted for re-esterification to [(3)H]cholesterol oleate, in comparison to that resulting from hydrolysis of [(3)H]cholesterol linoleate incorporated by selective uptake. For SRA, the formation of [(3)H]cholesterol oleate, which was substantial in control cells, was significantly inhibited in the presence of IFN-gamma. We conclude that there may be differences in cholesterol trafficking with respect to lipoprotein immune complex metabolism mediated by different classes of FcgammaRs. PMID- 10706589 TI - Interaction of locust apolipophorin III with lipoproteins and phospholipid vesicles: effect of glycosylation. AB - Apolipophorin III (apoLp-III) from Locusta migratoria is an exchangeable apolipoprotein that binds reversibly to lipoprotein surfaces. The native protein is glycosylated at Asn-18 and Asn-85. Variable attachment of five distinct oligosaccharide moieties at the two glycosylation sites results in molecular weight heterogeneity, as seen by mass spectrometry. The main mass peak of 20,488 Da decreases to 17,583 Da after removal of carbohydrate, indicating that apoLp III carbohydrate mass is approximately 14% by weight. Deglycosylated apoLp-III induced clearance of dimyristoylphosphatidylcholine and dimyristoylphosphatidylglycerol vesicles at a faster rate than glycosylated apoLp III. However, in lipoprotein binding assays, in which apoLp-III interacts with surface-localized diacylglycerol, only minor differences in binding were observed. The fluorescence properties of 1-anilinonaphthalene-8-sulfonate were unaffected by the glycosylation state of apoLp-III, indicating that no changes in the relative amount of exposed hydrophobic surface occurred as a result of carbohydrate removal. We propose that glycosyl moieties affect the ability of apoLp-III to transform phospholipid bilayer vesicles into disc-like complexes by steric hindrance. This is due to the requirement that apoLp-III penetrate the bilayer substrate prior to conformational opening of the helix bundle. On the other hand, the glycosyl moieties do not affect lipoprotein binding interactions as it does not involve deep protein penetration into the lipid milieu. Rather, lipoprotein binding is based on oriented protein contact with the lipid surface followed by opening of the helix bundle, which allows formation of a stable interaction with surface exposed hydrophobic sites. PMID- 10706590 TI - Maternal high density lipoproteins affect fetal mass and extra-embryonic fetal tissue sterol metabolism in the mouse. AB - Previous studies have shown that antibodies to cubulin, a receptor on the yolk sac that binds high density lipoproteins (HDL) and cobalamin, induce fetal abnormalities. Mice with markedly low concentrations of plasma HDL-cholesterol (HDL-C) give birth to healthy pups, however. To establish whether maternal HDL-C has a role in fetal development, sterol metabolism was studied in the fetus and extra-embryonic fetal tissues in wild-type and apolipoprotein A-I-deficient mice (apoAI-/-). Maternal HDL-C content was markedly greater in apoAI+/+ mice prior to pregnancy and at 13 days into gestation. By 17 days into gestation, HDL-C content was similar between both types of mice. Fetuses from apoAI (-/- x -/-) matings were 16;-25% smaller than control mice at 13 and 17 days of gestation and contained less cholesterol. The differences in size and cholesterol content were not due to a lack of cholesterol synthesis or apoA-I in the fetus. In the yolk sac and placenta, sterol synthesis rates were approximately 50% greater in the 13 day-old apoAI-/- mice as compared to the apoAI+/+ mice. Even though synthesis rates were greater, cholesterol concentrations were 22% lower in the yolk sac and similar in the placenta of apoAI-/- mice as compared to tissues of wild-type mice. These data suggest that a difference in maternal HDL-C concentration or composition can affect the size of the fetus and sterol metabolism of the yolk sac and placenta in the mouse. PMID- 10706591 TI - Novel mutations in the gene encoding ATP-binding cassette 1 in four tangier disease kindreds. AB - Tangier disease (TD) is an autosomal co-dominant disorder in which homozygotes have a marked deficiency of high density lipoprotein (HDL) cholesterol and, in some cases, peripheral neuropathy and premature coronary heart disease (CHD). Homozygotes are further characterized by cholesteryl ester deposition in various tissues throughout the body, most notably in those of the reticuloendothelial system. Several studies have demonstrated that the excess lipid deposition in TD is due to defective apolipoprotein-mediated efflux of cellular cholesterol and phospholipids. Although much progress has been made in our understanding of the metabolic basis of TD, the precise molecular defect had remained elusive until very recently. By positional cloning methods, we: 1) confirm the assignment of TD to chromosome 9q31, 2) provide evidence that human ATP-binding cassette-1 (hABC 1) maps to a 250 kb region on 9q31, and 3) describe novel deletion, insertion, and missense mutations in the gene encoding hABC-1 in four unrelated TD kindreds. These results establish a causal role for mutations in hABC-1 in TD and indicate that this transporter has a critical function in the regulation of intracellular lipid trafficking that dramatically affects plasma HDL cholesterol levels. PMID- 10706593 TI - Structure and characterization of the genes for murine choline/ethanolamine kinase isozymes alpha and beta. AB - Choline/ethanolamine kinase (CK/EK) is the first enzyme in phosphatidylcholine/phosphatidylethanolamine biosynthesis in all animal cells. The highly purified CKs from mammalian sources and their recombinant gene products so far were all shown to have EK activity also, indicating that both activities reside on the same protein. CK/EK in most animal cells exists as several isoforms, for two of which (alpha and beta) their cDNAs have been cloned from both the rat and mouse, and they are found to be separate gene products. The physiological significance for the existence of more than one CK/EK enzyme, however, remains to be clarified. In this study, we isolated mouse genes encoding both types of CK/EK isozyme and determined their entire structure. The 5' flanking promoter regions were found to have quite different features from each other, indicating that their expression could be under distinct control. Comparison of the nucleotide sequence between the corresponding coding exons showed the best homology (75%) residing on exon VIII. A search of the database resulted in the possible existence of 17 different origins of eukaryotic CK and/or EK, each of which presumably contained the entire amino acid sequence. Multialignment of their putative amino acid sequences led to an identification of the novel consensus sequence possibly required for the expression of either CK or EK activity, which corresponded to the sequence within exons VII and VIII of CK/EK-alpha and -beta genes from the mouse. This sequence was localized in close proximity to the C-terminal region of the general (Brenner's) phosphotransferase concensus sequence which was also completely conserved in all of the putative eukaryotic CK/EK proteins. The results demonstrated that, while both CK/EK-alpha and -beta genes were composed of 11 major exons, the size of their genes was quite different: 40 kb for CK/EK-alpha, whereas it was only 3.5 kb for CK/EK beta. PMID- 10706592 TI - Regulation of 25- and 27-hydroxylation side chain cleavage pathways for cholic acid biosynthesis in humans, rabbits, and mice. Assay of enzyme activities by high-resolution gas chromatography;-mass spectrometry. AB - In classic cholic acid biosynthesis, a series of ring modifications of cholesterol precede side chain cleavage and yield 5beta-cholestane-3alpha, 7alpha, 12alpha-triol. Side chain reactions of the triol then proceed either by the mitochondrial 27-hydroxylation pathway or by the microsomal 25-hydroxylation pathway. We have developed specific and precise assay methods to measure the activities of key enzymes in both pathways, 5beta-cholestane-3alpha, 7alpha, 12alpha-triol 25- and 27-hydroxylases and 5beta-cholestane-3alpha, 7alpha, 12alpha, 25-tetrol 23R-, 24R-, 24S- and 27-hydroxylases. The extracts from either the mitochondrial or microsomal incubation mixtures were purified by means of a disposable silica cartridge column, derivatized into trimethylsilyl ethers, and quantified by gas chromatography;-mass spectrometry with selected-ion monitoring in a high resolution mode. Compared with the addition of substrates in acetone, those in 2-hydroxypropyl-beta-cyclodextrin increased mitochondrial triol 27 hydroxylase activity 132% but decreased activities of the enzymes in microsomal 25-hydroxylation pathway (triol 25-hydroxylase and 5beta-cholestane-3alpha, 7alpha, 12alpha, 25-tetrol 23R-, 24R-, 24S- and 27-hydroxylases) 13;-60% in human liver. The enzyme activities in both pathways were generally 2- to 4-times higher in mouse and rabbit livers compared with human liver. In all species, microsomal triol 25-hydroxylase activities were 4- to 11-times larger than mitochondrial triol 27-hydroxylase activities but the activities of tetrol 24S-hydroxylase were similar to triol 27-hydroxylase activities in our assay conditions. The regulation of both pathways in rabbit liver was studied after bile acid synthesis was perturbed. Cholesterol feeding up-regulated enzyme activities involved in both 25- (64;-142%) and 27- (77%) hydroxylation pathways, while bile drainage up regulated only the enzymes in the 25-hydroxylation pathway (178;-371%). Using these new assays, we demonstrated that the 25- and 27-hydroxylation pathways for cholic acid biosynthesis are more active in mouse and rabbit than human livers and are separately regulated in rabbit liver. PMID- 10706594 TI - Specific phospholipid fatty acid composition of brain regions in mice. Effects of n-3 polyunsaturated fatty acid deficiency and phospholipid supplementation. AB - This study examined the effects of dietary alpha-linolenic acid deficiency followed or not by supplementation with phospholipids rich in n;-3 polyunsaturated fatty acid (PUFA) on the fatty acid composition of total phospholipids in 11 brain regions. Three weeks before mating, mice were fed a semisynthetic diet containing both linoleic and alpha-linolenic acid or deficient in alpha-linolenic acid. Pups were fed the same diet as their dams. At the age of 7 weeks, a part of the deficient group were supplemented with n;-3 polyunsaturated fatty acids (PUFA) from either egg yolk or pig brain phospholipids for 2 months. Saturated and monounsaturated fatty acid levels varied among brain regions and were not significantly affected by the diet. In control mice, the level of 22:6 n-3 was significantly higher in the frontal cortex compared to all regions. alpha-Linolenic acid deficiency decreased the level of 22:6 n-3 and was compensated by an increase in 22:5 n-6 in all regions. However, the brain regions were affected differently. After the pituitary gland, the frontal cortex, and the striatum were the most markedly affected with 40% reduction of 22:6 n-3. Supplementation with egg yolk or cerebral phospholipids in deficient mice restored a normal fatty acid composition in brain regions except for the frontal cortex. There was a regional distribution of the fatty acids in the brain and the impact of deficiency in alpha-linolenic acid was region specific. Dietary egg yolk or cerebral phospholipids are an effective source of n 3 PUFA for the recovery of altered fatty acid composition induced by a diet deficient in n-3 PUFA. PMID- 10706595 TI - Phospholipid supplementation reverses behavioral and biochemical alterations induced by n-3 polyunsaturated fatty acid deficiency in mice. AB - This study investigated the effects of a diet deficient in alpha-linolenic acid followed or not by supplementation with phospholipids rich in n-3 polyunsaturated fatty acids (PUFA) on behavior and phospholipid fatty acid composition in selected brain regions. Three weeks before mating, two groups of mice were fed a semisynthetic diet containing both linoleic and alpha-linolenic acid or a diet deficient in alpha-linolenic acid. Pups were fed the same diet as their dams. At the age of 7 weeks, a part of the deficient group was supplemented with n-3 PUFA from either egg yolk or pig brain phospholipids for 2 months. In the open field, rearing activity was significantly reduced in the deficient group. In the elevated plus maze (anxiety protocol), the time spent on open arms was significantly smaller in deficient mice than in controls. Using the learning protocol with the same task, the alpha-linolenic acid deficiency induced a learning deficit. Rearing activity and learning deficits were completely restored by supplementation with egg yolk or cerebral phospholipids, though the level of anxiety remained significantly higher than that of controls. There were no differences among the 4 diet groups for either the Morris water maze or passive avoidance. In control mice, the level of 22:6 n-3 was significantly higher in the frontal cortex compared to all other regions analysed. The frontal cortex and the striatum were the most markedly affected by the deficiency. Supplementation with phospholipids restored normal fatty acid composition in brain regions except for frontal cortex. Egg yolk or cerebral phospholipids are an effective source of n-3 PUFA for reversing behavioral changes and altered fatty acid composition induced by a diet deficient in n-3 PUFA. PMID- 10706596 TI - Genotypic associations of the hepatic secretion of VLDL apolipoprotein B-100 in obesity. AB - We examined the effect of genetic polymorphisms of proteins regulating intrahepatic processing of apolipoprotein B-100 (apoB) and the supply of neutral lipids to the liver on the hepatic secretion of very low density lipoprotein (VLDL) apoB in obesity. Hepatic secretion of very low density apolipoprotein B 100 (VLDL apoB) was measured using an infusion of [1-(13)C]leucine in 29 obese men. Isotopic enrichment and turnover of VLDL apoB was determined using gas chromatography-mass spectrometry and multi-compartmental modelling, respectively. Visceral fat was measured by magnetic resonance imaging. Genotypes for the apoB signal peptide (SP27/SP24 alleles), microsomal triglyceride transfer protein promoter (MTP, -493 G/T alleles), apoE (E2, E3, E4 alleles), hepatic lipase promoter (-514 C/T alleles), and cholesteryl ester transfer protein (CETP, Taq1B B1/B2 alleles) were determined using polymerase chain reaction. Statistically significant associations were found between hepatic secretion of apoB and allelic combinations of i) apoB SP with apoE (P = 0.02), hepatic lipase (P = 0.02), and CETP (P = 0. 006) genes, ii) MTP promoter with CETP genes (P = 0.03); the association with apoBSP/MTP promoter allelic combinations just failed to reach significance (P = 0.06), however. The CETP/apoBSP allelic combination was the most significant predictor of apoB secretion, and this was independent of visceral fat, plasma lathosterol and insulin levels, and dietary fat. SP24 carriers who were homozygous for CETP B1 had 60% lower apoB secretion than B2 heterozygotes who were non-carriers of SP24 (10.5 +/- 1.74 mg/kg fat free mass/day, n = 7 vs. 26.1 +/- 3.16, n = 22). The data suggest that variation in both the apoB and CETP genes may be a major genetic determinant of the hepatic secretion of apoB in men with visceral obesity. PMID- 10706597 TI - In vitro knockout of human p47phox blocks superoxide anion production and LDL oxidation by activated human monocytes. AB - We previously reported that superoxide dismutase (SOD) blocked human monocyte oxidation of LDL and therefore concluded that superoxide anion (O(2)(.-)) was required for oxidation. Others, however, have suggested that SOD may inhibit by mechanisms alternative to the dismutation of O(2)(.-). This study definitively addresses the involvement of O(2)(.-) in monocyte oxidation of LDL. Using an antisense ODN designed to target p47phox mRNA, we found that treatment of monocytes with antisense ODN caused a substantial and selective decrease in expression of p47phox protein, whereas sense ODN was without effect. Corresponding functional assays demonstrated that antisense ODN inhibited production of O(2)(.-). As sense ODN caused no inhibition of O(2)(.-) production, these results suggested that inhibition of p47phox expression caused reduction in O(2)(.-) production. Evaluation of the contribution of O(2)(.-) production to monocyte-mediated oxidation of LDL lipids confirmed that O(2)(.-) production is required for LDL lipid oxidation as antisense ODN treatment significantly inhibited LDL oxidation whereas sense ODN treatment caused no inhibition. This is the first report of the reduction of NADPH oxidase activity in intact human monocytes by directly targeting the mRNA of a significant member of this enzyme complex. Our results provide convincing data that O(2)(.-) is indeed required for monocyte-mediated LDL oxidation. PMID- 10706598 TI - God's organism? The chick as a model system for memory studies. AB - The young chick is a powerful model system in which to study the biochemical and morphological processes underlying memory formation. Training chicks on a one trial passive avoidance task results in a molecular cascade in a specific brain region, the intermediate medial hyperstriatum ventrale. This cascade is initiated by glutamate release and engages a series of synaptic transients including increased calcium flux, up-regulation of NMDA-glutamate receptors, membrane protein phosphorylations, and the retrograde messenger NO. Expression of immediate early genes c-fos and c-jun precedes the synthesis, glycosylation, and redistribution, >4 hr downstream, of a number of synaptic membrane proteins, notably NCAM and L1. Other membrane proteins required in the early phase of memory formation include the amyloid precursor protein (APP) and apolipoprotein E. There are concomitant increases in dendritic spine number and changes in synaptic structure. Nonsynaptic factors, including corticosterone and BDNF, can modulate retention of the avoidance response, enhancing the salience of otherwise weakly retained memory. These results are discussed in relation to general concepts of memory formation and the spatio-temporal distribution of the putative memory trace. PMID- 10706599 TI - Tissue-specific expression of a type I adenylyl cyclase rescues the rutabaga mutant memory defect: in search of the engram. AB - Most attempts to localize physical correlates of memory in the central nervous system (CNS) rely on ablation techniques. This approach has the limitation of defining just one of an unknown number of structures necessary for memory formation. We have used the Drosophila rutabaga type I Ca(2+)/CaM-dependent adenylyl cyclase (AC) gene to determine in which CNS region AC expression is sufficient for memory formation. Using pan-neural and restricted CNS expression with the GAL4 binary transcription activation system, we have rescued the memory defect of the rutabaga mutant in a fast robust spatial learning paradigm. The ventral ganglion, antennal lobes, and median bundle are likely the CNS structures sufficient for rutabaga AC- dependent spatial learning. PMID- 10706600 TI - Visual input regulates circuit configuration in courtship conditioning of Drosophila melanogaster. AB - Courtship and courtship conditioning are behaviors that are regulated by multiple sensory inputs, including chemosensation and vision. Globally inhibiting CaMKII activity in Drosophila disrupts courtship plasticity while leaving visual and chemosensory perception intact. Light has been shown to modulate CaMKII-dependent memory formation in this paradigm and the circuitry for the nonvisual version of this behavior has been investigated. In this paradigm, volatile and tactile pheromones provide the primary driving force for courtship, and memory formation is dependent upon intact mushroom bodies and parts of the central complex. In the present study, we use the GAL4/UAS binary expression system to define areas of the brain that require CaMKII for modulation of courtship conditioning in the presence of visual, as well as chemosensory, information. Visual input suppressed the ability of mushroom body- and central complex-specific CaMKII inhibition to disrupt memory formation, indicating that the cellular circuitry underlying this behavior can be remodeled by changing the driving sensory modality. These findings suggest that the potential for plasticity in courtship behavior is distributed among multiple biochemically and anatomically distinct cellular circuits. PMID- 10706601 TI - Dishabituation of the Tritonia escape swim. AB - When repeatedly elicited, the oscillatory escape swim of the marine mollusc Tritonia diomedea undergoes habituation of the number of cycles per swim. Although similar in most respects to habituation observed in vertebrates and other invertebrates, one key feature, dishabituation, has been surprisingly difficult to demonstrate. Here we evaluate the hypothesis that this is due to interference from short-term sensitization, which is manifested as a reduction in swim onset latency, that occurs simultaneously during habituation training. Robust dishabituation was obtained using a multisession habituation protocol designed to allow this sensitization to dissipate before the dishabituatory stimulus was applied. These results extend the similarity of habituation in Tritonia to that described in other species, strengthening the usefulness of this preparation as a model system for studies of the cellular basis of habituation. PMID- 10706602 TI - Primacy versus recency in a quantitative model: activity is the critical distinction. AB - Behavioral and neurobiological evidence shows that primacy and recency are subserved by memory systems for intermediate- and short-term memory, respectively. A widely accepted explanation of recency is that in short-term memory, new learning overwrites old learning. Primacy is not as well understood, but many hypotheses contend that initial items are better encoded into long-term memory because they have had more opportunity to be rehearsed. A simple, biologically motivated neural network model supports an alternative hypothesis of the distinct processing requirements for primacy and recency given single-trial learning without rehearsal. Simulations of the model exhibit either primacy or recency, but not both simultaneously. The incompatibility of primacy and recency clarifies possible reasons for two neurologically distinct systems. Inhibition, and its control of activity, determines those list items that are acquired and retained. Activity levels that are too low do not provide sufficient connections for learning to occur, while higher activity diminishes capacity. High recurrent inhibition, and progressively diminishing activity, allows acquisition and retention of early items, while later items are never acquired. Conversely, low recurrent inhibition, and the resulting high activity, allows continuous acquisition such that acquisition of later items eventually interferes with the retention of early items. PMID- 10706603 TI - Computer-assisted behavioral assessment of Pavlovian fear conditioning in mice. AB - In Pavlovian fear conditioning, a conditional stimulus (CS, usually a tone) is paired with an aversive unconditional stimulus (US, usually a foot shock) in a novel context. After even a single pairing, the animal comes to exhibit a long lasting fear to the CS and the conditioning context, which can be measured as freezing, an adaptive defense reaction in mice. Both context and tone conditioning depend on the integrity of the amygdala, and context conditioning further depends on the hippocampus. The reliability and efficiency of the fear conditioning assay makes it an excellent candidate for the screening of learning and memory deficits in mutant mice. One obstacle is that freezing in mice has been accurately quantified only by human observers, using a tedious method that can be subject to bias. In the present study we generated a simple, high-speed, and highly accurate algorithm that scores freezing of four mice simultaneously using NIH Image on an ordinary Macintosh computer. The algorithm yielded a high correlation and excellent linear fit between computer and human scores across a broad range of conditions. This included the ability to score low pretraining baseline scores and accurately mimic the effects of two independent variables (shock intensity and test modality) on fear. Because we used a computer and digital video, we were able to acquire a secondary index of fear, activity suppression, as well as baseline activity scores. Moreover, we measured the unconditional response to shock. These additional measures can enhance the sensitivity of the assay to detect interesting memory phenotypes and control for possible confounds. Thus, this computer-assisted system for measuring behavior during fear conditioning allows for the standardized and carefully controlled assessment of multiple aspects of the fear conditioning experience. PMID- 10706604 TI - Pleiotrophin signals increased tyrosine phosphorylation of beta beta-catenin through inactivation of the intrinsic catalytic activity of the receptor-type protein tyrosine phosphatase beta/zeta. AB - Pleiotrophin (PTN) is a platelet-derived growth factor-inducible, 18-kDa heparin binding cytokine that signals diverse phenotypes in normal and deregulated cellular growth and differentiation. To seek the mechanisms of PTN signaling, we studied the interactions of PTN with the receptor protein tyrosine phosphatase (RPTP) beta/zeta in U373-MG cells. Our results suggest that PTN is a natural ligand for RPTP beta/zeta. PTN signals through "ligand-dependent receptor inactivation" of RPTP beta/zeta and disrupts its normal roles in the regulation of steady-state tyrosine phosphorylation of downstream signaling molecules. We have found that PTN binds to and functionally inactivates the catalytic activity of RPTP beta/zeta. We also have found that an active site-containing domain of RPTP beta/zeta both binds beta-catenin and functionally reduces its levels of tyrosine phosphorylation when added to lysates of pervanidate-treated cells. In contrast, an (inactivating) active-site mutant of RPTP beta/zeta also binds beta catenin but fails to reduce tyrosine phosphorylation of beta-catenin. Finally, in parallel to its ability to inactivate endogenous RPTP beta/zeta, PTN sharply increases tyrosine phosphorylation of beta-catenin in PTN-treated cells. The results suggest that in unstimulated cells, RPTP beta/zeta is intrinsically active and functions as an important regulator in the reciprocal control of the steady-state tyrosine phosphorylation levels of beta-catenin by tyrosine kinases and phosphatases. The results also suggest that RPTP beta/zeta is a functional receptor for PTN; PTN signals through ligand-dependent receptor inactivation of RPTP beta/zeta to increase levels of tyrosine phosphorylation of beta-catenin to initiate downstream signaling. PTN is the first natural ligand identified for any of the RPTP family; its identification provides a unique tool to pursue the novel signaling pathway activated by PTN and the relationship of PTN signaling with other pathways regulating beta-catenin. PMID- 10706605 TI - Erythroid Kruppel-like factor is recruited to the CACCC box in the beta-globin promoter but not to the CACCC box in the gamma-globin promoter: the role of the neighboring promoter elements. AB - The programmed expression of the five beta-like globin genes (epsilon, (A)gamma, (G)gamma, delta, and beta) is characterized by a series of switches that are developmentally regulated. The (A)gamma- and (G)gamma- (fetus) to beta-globin (adult) switch depends on transcription factor erythroid Kruppel-like factor (EKLF), which, like Sp1, binds to CACCC boxes. EKLF is essential for the expression of the beta-globin but not the gamma-globin gene. Because both gamma globin and beta-globin promoters contain the CACCC box, and their promoter elements are similar, it is not known why the two promoters behave so differently. In this report, we searched for the functional differences between the two promoters by studying their ability to recruit EKLF. We used the in vivo PIN*POINT assay to show that EKLF is recruited to the beta-globin promoter but not to the gamma-globin promoter. We show that this selectivity is a result of differences in surrounding promoter elements and not CACCC box alone. One of the differences between the two promoters with a functional consequence is the CCTTG repeat that is present in the gamma-globin promoter but not in the beta-globin promoter. The repeat, when inserted in the beta-globin promoter, decreases EKLF recruitment to and activity of the beta-globin promoter, suggesting that the repeat functions as a suppressor element. The CCTTG repeat can also suppress the SV40 promoter in cis, and the suppressor factor binding to the repeat can be squelched with a plasmid containing a high copy number of the repeat. These findings may have implications in designing drug targets for treatment of beta globin disorders. PMID- 10706606 TI - The missing organic molecules on Mars. AB - GC-MS on the Viking 1976 Mars missions did not detect organic molecules on the Martian surface, even those expected from meteorite bombardment. This result suggested that the Martian regolith might hold a potent oxidant that converts all organic molecules to carbon dioxide rapidly relative to the rate at which they arrive. This conclusion is influencing the design of Mars missions. We reexamine this conclusion in light of what is known about the oxidation of organic compounds generally and the nature of organics likely to come to Mars via meteorite. We conclude that nonvolatile salts of benzenecarboxylic acids, and perhaps oxalic and acetic acid, should be metastable intermediates of meteoritic organics under oxidizing conditions. Salts of these organic acids would have been largely invisible to GC-MS. Experiments show that one of these, benzenehexacarboxylic acid (mellitic acid), is generated by oxidation of organic matter known to come to Mars, is rather stable to further oxidation, and would not have been easily detected by the Viking experiments. Approximately 2 kg of meteorite-derived mellitic acid may have been generated per m(2) of Martian surface over 3 billion years. How much remains depends on decomposition rates under Martian conditions. As available data do not require that the surface of Mars be very strongly oxidizing, some organic molecules might be found near the surface of Mars, perhaps in amounts sufficient to be a resource. Missions should seek these and recognize that these complicate the search for organics from entirely hypothetical Martian life. PMID- 10706607 TI - An advanced glycation endproduct cross-link breaker can reverse age-related increases in myocardial stiffness. AB - Decreased elasticity of the cardiovascular system is one of the hallmarks of the normal aging process of mammals. A potential explanation for this decreased elasticity is that glucose can react nonenzymatically with long-lived proteins, such as collagen and lens crystallin, and link them together, producing advanced glycation endproducts (AGEs). Previous studies have shown that aminoguanidine, an AGE inhibitor, can prevent glucose cross-linking of proteins and the loss of elasticity associated with aging and diabetes. Recently, an AGE cross-link breaker (ALT-711) has been described, which we have evaluated in aged dogs. After 1 month of administration of ALT-711, a significant reduction ( approximately 40%) in age-related left ventricular stiffness was observed [(57.1 +/- 6.8 mmHg x m(2)/ml pretreatment and 33.1 +/- 4.6 mmHg x m(2)/ml posttreatment (1 mmHg = 133 Pa)]. This decrease was accompanied by improvement in cardiac function. PMID- 10706608 TI - On the interpretation of quantitative structure-function activity relationship data for lactate oxidase. AB - The native flavin, FMN, has been removed from the l-lactate oxidase of Aerococcus viridans, and the apoprotein reconstituted with 12 FMN derivatives with various substituents at the flavin 6- and 8-positions. Impressive linear relationships are exhibited between the sum of the Hammett final sigma(para) and final sigma(ortho) parameters and the redox potentials of the free flavins, and between the redox potentials of the free and enzyme-bound flavins. Rapid reaction kinetics studies of the reconstituted enzymes with the substrates l-lactate and l mandelate show an increase in the reduction rate constant with increasing redox potential, except that, with lactate, a limiting rate constant of approximately 700 s(-1) is obtained with flavins of high potential. Similar breakpoints are found in plots of the rate constants for flavin N5-sulfite adduct formation and for the reaction of the reduced enzymes with molecular oxygen. These results are interpreted in terms of a two-step equilibrium preceding the chemical reaction step, in which the second equilibrium step provides an upper limit to the rate with which the particular substrate or ligand is positioned with the flavin in the correct fashion for the observed chemical reaction to occur. The relationship of rate constants for flavin reduction and N5-sulfite adduct formation with flavin redox potential below the observed breakpoint indicate development of significant negative charge in the transition states of the reactions. In the case of reduction by substrate, the results are consistent either with a hydride transfer mechanism or with the so called "carbanion" mechanism, in which the substrate alpha-proton is abstracted by an enzyme base protected from exchange with solvent. These conclusions are supported by substrate alpha-deuterium isotope effects and by solvent viscosity effects on sulfite binding. PMID- 10706609 TI - Dominant negative down-regulation of endotoxin-induced tumor necrosis factor alpha production by Lps(d)/Ran. AB - We recently showed that adenoviral transfer and expression of the Lps(d)/Ran gene isolated from endotoxin-resistant C3H/HeJ mice could protect endotoxin-sensitive mice from endotoxic shock. Elevation of proinflammatory cytokines, such as tumor necrosis factor alpha (TNF-alpha), is thought to be essential for the development of septic shock. To investigate the extent to which Lps(d)/Ran affects TNF-alpha production, we transduced primary macrophages from endotoxin-sensitive and resistant mice with adenoviral vectors expressing the wild-type and the mutant Lps/Ran cDNAs and other control genes, and compared the amount of TNF-alpha produced by these various transduced macrophages. Successful transfer and expression of Lps(d)/Ran cDNA in endotoxin-sensitive C3H/HeOuJ macrophages reduced TNF-alpha production upon lipopolysaccharide (LPS) stimulation, as compared with macrophages transduced with vectors expressing the wild-type Lps(n)/Ran cDNA, the green fluorescent protein gene, or the lacZ gene. On the other hand, successful transfer and expression of the wild-type Lps(n)/Ran cDNA in primary macrophages from endotoxin-resistant C3H/HeJ mice failed to induce TNF alpha production to any significant extent unless a very high LPS concentration was used. Given our previous demonstration that Lps(n)/Ran functions effectively in restoring LPS responsiveness in B cells from C3H/HeJ mice, we conclude that Lps/Ran is involved in a CD14-independent signal transduction pathway. This dominant negative down-regulation by Lps(d)/Ran on TNF-alpha production by macrophages and probably other innate immune responses may be key to the development of an effective gene therapy for endotoxic or septic shock. PMID- 10706610 TI - Concurrent MRI and diffuse optical tomography of breast after indocyanine green enhancement. AB - We present quantitative optical images of human breast in vivo. The images were obtained by using near-infrared diffuse optical tomography (DOT) after the administration of indocyanine green (ICG) for contrast enhancement. The optical examination was performed concurrently with a magnetic resonance imaging (MRI) exam on patients scheduled for excisional biopsy or surgery so that accurate image coregistration and histopathological information of the suspicious lesions was available. The ICG-enhanced optical images coregistered accurately with Gadolinium-enhanced magnetic resonance images validating the ability of DOT to image breast tissue. In contrast to simple transillumination, we found that DOT provides for localization and quantification of exogenous tissue chromophore concentrations. Additionally our use of ICG, an albumin bound absorbing dye in plasma, demonstrates the potential to differentiate disease based on the quantified enhancement of suspicious lesions. PMID- 10706611 TI - Environment-dependent residue contact energies for proteins. AB - We examine the interactions between amino acid residues in the context of their secondary structural environments (helix, strand, and coil) in proteins. Effective contact energies for an expanded 60-residue alphabet (20 aa x three secondary structural states) are estimated from the residue-residue contacts observed in known protein structures. Similar to the prototypical contact energies for 20 aa, the newly derived energy parameters reflect mainly the hydrophobic interactions; however, the relative strength of such interactions shows a strong dependence on the secondary structural environment, with nonlocal interactions in beta-sheet structures and alpha-helical structures dominating the energy table. Environment-dependent residue contact energies outperform existing residue pair potentials in both threading and three-dimensional contact prediction tests and should be generally applicable to protein structure prediction. PMID- 10706612 TI - Isotope-labeled immunoassays without radiation waste. AB - The practice of immunoassay has experienced a widespread transition from radioisotopic labeling to nonisotopic labeling over the last two decades. Radioisotope labels have drawbacks that hamper their applications: (i) perceived radiation hazards of reagents, (ii) regulatory requirements and disposal problems of working with radioactive materials, and (iii) short shelf-life of the labeled reagents. The advantage of isotopic labeling is the incorporation into analytes without altering structure or reactivity, as is often the case with ELISA or fluorescent detection systems. We developed a format for isotope label immunoassay with the long-life isotope (14)C as the label and accelerator mass spectrometer (AMS) as the detection system. AMS quantifies attomole levels of several isotopes, including (14)C. With this exquisite sensitivity, the sensitivity of an immunoassay is limited by the K(d) of the antibody and not the detection system. The detection limit of the assays for atrazine and 2,3,7,8 tetrachlorodibenzo-p-dioxin was 2.0 x 10(-10) M and 2.0 x 10(-11) M, respectively, approximately an order of magnitude below the standard enzyme immunoassay. Notably, <1 dpm (0.45 pCi) of (14)C-labeled compound was used in each assay, which is well below the limit of disposal (50 nCi per g) as nonradioactive waste. Thus, endogenous reporter ligands quantified by AMS provide the advantages of an RIA without the associated problems of radioactive waste. PMID- 10706613 TI - Notch signaling directly controls cell proliferation in the Drosophila wing disc. AB - Notch signaling is involved in cell differentiation and patterning during morphogenesis. In the Drosophila wing, Notch activity regulates the expression of several genes at the dorsal/ventral boundary, and this is thought to elicit wing cell proliferation. In this work, we show the effect of clones of cells expressing different forms of several members of the Notch signaling pathway, which result in an alteration of Notch activity. The ectopic expression in clones of activated forms of Notch or of its ligands (Delta or Serrate) in the wing causes outgrowths associated with the appearance of ectopic wing margins. These outgrowths consist of mutant territories and of surrounding wild-type cells. However, the ectopic expression of Delta, at low levels in ventral clones, causes large outgrowths that are associated neither with the generation of wing margin structures nor with the expression of genes characteristic of the dorsal/ventral boundary. These results suggest that Notch activity is directly involved in cell proliferation, independently of its role in the formation of the dorsal/ventral boundary. We propose that the nonautonomous effects (induction of extraproliferation and vein differentiation in the surrounding wild-type cells) result from pattern accommodation to positional values caused by the ectopic expression of Notch. PMID- 10706614 TI - Hyperphosphorylated tau and neurofilament and cytoskeletal disruptions in mice overexpressing human p25, an activator of cdk5. AB - Hyperphosphorylation of microtubule-associated proteins such as tau and neurofilament may underlie the cytoskeletal abnormalities and neuronal death seen in several neurodegenerative diseases including Alzheimer's disease. One potential mechanism of microtubule-associated protein hyperphosphorylation is augmented activity of protein kinases known to associate with microtubules, such as cdk5 or GSK3beta. Here we show that tau and neurofilament are hyperphosphorylated in transgenic mice that overexpress human p25, an activator of cdk5. The p25 transgenic mice display silver-positive neurons using the Bielschowsky stain. Disturbances in neuronal cytoskeletal organization are apparent at the ultrastructural level. These changes are localized predominantly to the amygdala, thalamus/hypothalamus, and cortex. The p25 transgenic mice display increased spontaneous locomotor activity and differences from control in the elevated plus-maze test. The overexpression of an activator of cdk5 in transgenic mice results in increased cdk5 activity that is sufficient to produce hyperphosphorylation of tau and neurofilament as well as cytoskeletal disruptions reminiscent of Alzheimer's disease and other neurodegenerative diseases. PMID- 10706615 TI - The olfactory receptor gene repertoire in primates and mouse: evidence for reduction of the functional fraction in primates. AB - Olfactory receptors (ORs) located in the cell membrane of olfactory sensory neurons of the nasal epithelium are responsible for odor detection by binding specific odorant ligands. Primates are thought to have a reduced sense of smell (microsmatic) with respect to other mammals such as dogs or rodents. We have previously demonstrated that over 70% of the human OR genes have become nonfunctional pseudogenes, leading us to hypothesize that the reduced sense of smell could correlate with the loss of functional genes. To extend these results, we sampled the OR gene repertoire of 10 primate species, from prosimian lemur to human, in addition to mouse. About 221 previously unidentified primate sequences and 33 mouse sequences were analyzed. These sequences encode ORs distributed in seven families and 56 subfamilies. Analysis showed a high fraction ( approximately 50% on average) of pseudogenes in hominoids. In contrast, only approximately 27% of OR genes are pseudogenes in Old World monkeys, and New World monkeys are almost free of pseudogenes. The prosimian branch seems to have evolved differently from the other primates and has approximately 37% pseudogene content. No pseudogenes were found in mouse. With the exception of New World monkeys, we demonstrate that primates have a high fraction of OR pseudogenes compared with mouse. We hypothesize that under relaxed selective constraints, primates would have progressively accumulated pseudogenes with the highest level seen in hominoids. The fraction of pseudogenes in the OR gene repertoire could parallel the evolution of the olfactory sensory function. PMID- 10706616 TI - Functional overlap of sequences that activate transcription and signal ubiquitin mediated proteolysis. AB - Many transcription factors, particularly those involved in the control of cell growth, are unstable proteins destroyed by ubiquitin-mediated proteolysis. In a previous study of sequences targeting the transcription factor Myc for destruction, we observed that the region in Myc signaling ubiquitin-mediated proteolysis overlaps closely with the region in Myc that activates transcription. Here, we present evidence that the overlap of these two activities is not unique to Myc, but reflects a more general phenomenon. We show that a similar overlap of activation domains and destruction elements occurs in other unstable transcription factors and report a close correlation between the ability of an acidic activation domain to activate transcription and to signal proteolysis. We also show that destruction elements from yeast cyclins, when tethered to a DNA binding domain, activate transcription. The intimate overlap of activation domains and destruction elements reveals an unexpected convergence of two very different processes and suggests that transcription factors may be destroyed because of their ability to activate transcription. PMID- 10706617 TI - Visualization of gas flow and diffusion in porous media. AB - The transport of gases in porous materials is a crucial component of many important processes in science and technology. In the present work, we demonstrate how magnetic resonance microscopy with continuous flow laser polarized noble gases makes it possible to "light up" and thereby visualize, with unprecedented sensitivity and resolution, the dynamics of gases in samples of silica aerogels and zeolite molecular sieve particles. The "polarization weighted" images of gas transport in aerogel fragments are correlated to the diffusion coefficient of xenon obtained from NMR pulsed-field gradient experiments. The technique provides a unique means of studying the combined effects of flow and diffusion in systems with macroscopic dimensions and microscopic internal pore structure. PMID- 10706618 TI - The retinoblastoma-interacting zinc-finger protein RIZ is a downstream effector of estrogen action. AB - Co-immunoprecipitation experiments in cell extract from cultured cells or target tissues indicated that estrogen receptor was complexed with the retinoblastoma binding protein RIZ in a ligand-dependent manner. Mapping of interaction sites indicated that in both proteins the same regions and motifs responsible for the interaction of transcriptional co-activator and nuclear receptors were involved. In cultured cells, estradiol induced a redistribution of RIZ protein within the nucleus and in the cytoplasm. A similar effect was produced in vivo, in prepuberal rat endometrium, by administration of a physiological dose of estradiol. Therefore, RIZ protein could be a specific effector of estrogen action downstream of the hormone-receptor interaction, presumably involved in proliferation control. PMID- 10706619 TI - Detection of leukemia-associated MLL-GAS7 translocation early during chemotherapy with DNA topoisomerase II inhibitors. AB - Leukemias with MLL gene translocations are a complication of primary cancer treatment with DNA topoisomerase II inhibitors. How early translocations appear during primary cancer treatment has not been investigated. We tracked the leukemic clone with an MLL gene translocation during neuroblastoma therapy in a child who developed acute myeloid leukemia. The karyotype of the leukemic clone showed del(11)(q23). We used panhandle PCR-based methods to isolate the breakpoint junction involving MLL and an unknown partner gene. Marrow DNA from neuroblastoma diagnosis and DNA and RNA from serial preleukemic marrows were examined for the translocation. The karyotypic del(11)(q23) was a cryptic t(11;17). GAS7, a growth arrest-specific gene at chromosome band 17p13, was the partner gene of MLL. Two different MLL-GAS7 fusion transcripts were expressed. The translocation was already detectable by 1.5 months after the start of neuroblastoma treatment. The translocation was not detectable in the marrow at neuroblastoma diagnosis or in peripheral blood lymphocyte DNAs of six normal subjects. GAS7 is a new partner gene of MLL in treatment-related acute myeloid leukemia. MLL gene translocations can be present early during anticancer treatment at low cumulative doses of DNA topoisomerase II inhibitors. Although MLL has many partner genes and most have not been characterized, panhandle PCR strategies afford new means for detecting MLL gene translocations early during therapy when the partner gene is unknown. PMID- 10706621 TI - Learning improves growth rate in grasshoppers. AB - To quantify the adaptive significance of insect learning, we documented the behavior and growth rate of grasshoppers (Schistocerca americana) in an environment containing two artificial food types, one providing a balanced diet of protein and carbohydrate, which maximizes growth, and the other being carbohydrate-deficient, which is unsuitable for growth. Grasshoppers in the Learning treatment experienced a predictable environment, where the spatial location, taste, and color of each food source remained constant throughout the experiment. In contrast, grasshoppers of the Random treatment developed in a temporally varying environment, where the spatial location, taste, and color of the balanced and deficient food types randomly alternated twice each day. Our results show that the grasshoppers that could employ associative learning for diet choice experienced higher growth rates than individuals of the Random treatment, demonstrating the adaptive significance of learning in a small short lived insect. PMID- 10706620 TI - Mantle cell lymphoma is characterized by inactivation of the ATM gene. AB - In mantle cell lymphoma (MCL), the translocation t(11;14) is considered the cytogenetic hallmark of the disease. Recently, however, deletion of the chromosomal region 11q22-q23 has been identified as a frequent event in this type of cancer, indicating the existence of a pathogenically relevant tumor suppressor gene in this region. The deleted segment contains the ATM (ataxia telangiectasia mutated) gene. ATM is an interesting candidate as a tumor suppressor gene because constitutive inactivation of the gene predisposes ataxia telangiectasia patients to lymphoid malignancies. To assess the potential involvement of the gene in MCL lymphomagenesis, we performed mutation analysis of ATM in 12 sporadic cases of MCL, 7 of them with a deletion of one ATM gene copy, by using single-strand conformation polymorphism analysis of reverse transcription-PCR-amplified mRNA and subsequent DNA sequencing. In all seven cases containing a deletion of one ATM allele, a point mutation in the remaining allele was detected, which resulted in aberrant transcript splicing, truncation, or alteration of the protein. In addition, biallelic ATM mutations were identified in two MCLs that did not contain 11q deletions. Interestingly, in three cases analyzed, the ATM mutations detected in the tumor cells were not present in nonmalignant cells, demonstrating their somatic rather than germ-line origin. The inactivation of both alleles of the ATM gene by deletion and deleterious point mutation in the majority of cases analyzed indicates that ATM plays a role in the initiation and/or progression of MCL. PMID- 10706622 TI - Thyroid hormone-dependent metamorphosis in a direct developing frog. AB - The direct developing anuran, Eleutherodactylus coqui, lacks a tadpole, hatching as a tiny frog. We investigated the role of the metamorphic trigger, thyroid hormone (TH), in this unusual ontogeny. Expression patterns of the thyroid hormone receptors, TRalpha and TRbeta, were similar to those of indirect developers. TRbeta mRNA levels increased dramatically around the time of thyroid maturation, when remodeling events reminiscent of metamorphosis occur. Treatment with the goitrogen methimazole inhibited this remodeling, which was reinitiated on cotreatment with TH. Despite their radically altered ontogeny, direct developers still undergo a TH-dependent metamorphosis, which occurs before hatching. We propose a new model for the evolution of anuran direct development. PMID- 10706623 TI - Alphabeta protomers of Na+,K+-ATPase from microsomes of duck salt gland are mostly monomeric: formation of higher oligomers does not modify molecular activity. AB - The distance that separates alphabeta protomers of the Na(+), K(+)-ATPase in microsomes and in purified membranes prepared from duck nasal salt glands was estimated by measuring fluorescence resonance energy transfer between anthroylouabain bound to a population of alphabeta protomers and either N-[7 nitrobenz-2-oxa-1, 3-diazol-4-yl]-6-aminohexyl ouabain or 5-(and-6) carboxyfluorescein-6-aminohexyl ouabain bound to the rest. Energy transfer between probes bound in the microsomal preparation was less than in the purified membranes. The efficiency of energy transfer between anthroylouabain and N-[7 nitrobenz-2-oxa-1, 3-diazol-4-yl]-6-aminohexyl ouabain was 29.2% in the microsomes compared with 62.6% in the purified preparation. Similar results were obtained with 5-(and-6)-carboxyfluorescein-6-aminohexyl ouabain as acceptor. We calculate that either the protomer bound probes were on the average 13 A farther apart in the microsomes than in the purified membranes, or that 53% of the protomers are monomeric in the microsome preparation. Microsomes prepared in the presence of phalloidin (a toxin that binds to F actin and stabilizes the actin based cytoskeleton) showed less quench than those prepared in its absence. The data support the hypothesis that protomers are kept apart by their association with the cytoskeleton. The turnover rate while hydrolyzing ATP is the same in the microsomal and purified preparations; higher oligomer formation has no significant effect on the enzyme reaction mechanism. PMID- 10706624 TI - Genetic analysis of speciation by means of introgression into Drosophila melanogaster. AB - In the last decade, the genetic basis of reproductive isolation has been shown to be surprisingly polygenic, and yet even the most efficient system currently in use could lend itself to molecular analysis only in highly selected cases. By extending the recent discovery of fertility rescue between Drosophila melanogaster and Drosophila simulans, we show that this hybridization can permit systematic and precise delineation of the genetic and molecular basis of speciation. In a region of 5% of the D. simulans genome introgressed into D. melanogaster, we discover at least six genes of hybrid male sterility and none for female sterility by deficiency mapping. A single case of hybrid inviability has been tracked down to a 3-Kb element that was inserted into the Cyclin E locus during species hybridization. The extent of interspecific genetic divergence underlying hybrid male sterility, especially in contrast with the low degree of inviability and female sterility, is far greater than expected from previous studies. PMID- 10706625 TI - The orphan nuclear receptor Tlx regulates Pax2 and is essential for vision. AB - Although the development of the vertebrate eye is well described, the number of transcription factors known to be key to this process is still limited. The localized expression of the orphan nuclear receptor Tlx in the optic cup and discrete parts of the central nervous system suggested the possible role of Tlx in the formation or function of these structures. Analyses of Tlx targeted mice revealed that, in addition to the central nervous system cortical defects, lack of Tlx function results in progressive retinal and optic nerve degeneration with associated blindness. An extensive screen of Tlx-positive and Tlx-negative P19 neural precursors identified Pax2 as a candidate target gene. This identification is significant, because Pax2 is known to be involved in retinal development in both the human and the mouse eye. We find that Pax2 is a direct target and that the Tlx binding site in its promoter is conserved between mouse and human. These studies show that Tlx is a key component of retinal development and vision and an upstream regulator of the Pax2 signaling cascade. PMID- 10706627 TI - A Bacillus subtilis operon containing genes of unknown function senses tRNATrp charging and regulates expression of the genes of tryptophan biosynthesis. AB - Strains of Bacillus subtilis containing a temperature-sensitive tryptophanyl-tRNA synthetase produce elevated levels of the tryptophan pathway enzymes, when grown at high temperatures in the presence of excess tryptophan. This increase is because of reduced availability of the tryptophan-activated trp RNA-binding attenuation protein (TRAP). To test the hypothesis that this elevated trp gene expression was caused by the overproduction of a transcript capable of binding and sequestering TRAP, a computer program was designed to search the B. subtilis genome sequence for additional potential TRAP binding sites. A region containing a stretch of (G/A)AG trinucleotide repeats, characteristic of a TRAP binding site, was identified in the yczA-ycbK operon. We show that transcriptional regulation of the yczA-ycbK operon is controlled by the T-box antitermination mechanism in response to the level of uncharged tRNA(Trp), and that the presence of a trpS1 mutant allele increases production of the yczA-ycbK transcript. Elevated yczA-ycbK expression was shown to activate transcription of the trp operon. Deletion of the yczA-ycbK operon abolishes the trpS1 effect on trp gene expression. The purpose of increasing expression of the genes of tryptophan biosynthesis in the trpS mutant would be to provide additional tryptophan to overcome the charged tRNA(Trp) deficiency. Therefore, in B. subtilis, as in Escherichia coli, transcription of the tryptophan biosynthetic genes is regulated in response to changes in the extent of charging of tRNA(Trp) as well as the availability of tryptophan. PMID- 10706626 TI - Stress-induced enhancement of skin immune function: A role for gamma interferon. AB - Contrary to the widespread belief that stress is necessarily immunosuppressive, recent studies have shown that, under certain conditions, stress can induce a significant enhancement of a skin cell-mediated immune response [delayed-type hypersensitivity (DTH) or contact hypersensitivity]. Adrenal stress hormones and a stress-induced trafficking of leukocytes from the blood to the skin have been identified as systemic mediators of this immunoenhancement. Because gamma interferon (IFNgamma) is an important cytokine mediator of DTH, the studies described here were designed to examine its role as a local mediator of the stress-induced enhancement of skin DTH. The effect of acute stress on skin DTH was examined in wild-type and IFNgamma receptor-deficient (IFNgammaR-/-) mice that had previously been sensitized with 2,4-dinitro-1-fluorobenzene. Acutely stressed wild-type mice showed a significantly larger DTH response than nonstressed mice. In contrast, IFNgammaR-/- mice failed to show a stress-induced enhancement of skin DTH. Immunoneutralization of IFNgamma in wild-type mice significantly reduced the stress-induced enhancement of skin DTH. In addition, an inflammatory response induced by direct IFNgamma administration to the skin was significantly enhanced by acute stress. Our results suggest that IFNgamma is an important local mediator of a stress-induced enhancement of skin DTH. These studies are clinically relevant because, depending on the nature of the antigen, DTH reactions mediate numerous protective (e.g., resistance to viral, bacterial, parasitic, and fungal infections) or pathological (e.g., autoimmune reactions and contact sensitivity reactions such as that to poison ivy) immune responses. PMID- 10706628 TI - Dendritic cells purified from myeloma are primed with tumor-specific antigen (idiotype) and activate CD4+ T cells. AB - Multiple myelomas produce tumor-specific antigen (TSA) in the form of idiotype (Id) on monoclonal Ig. CD4(+) T cells can recognize Id-peptide on MHC class II molecules and protect against challenges with MOPC315 cells, which are, as common for myelomas, class II-negative. The present study explains these previous results by demonstrating that Id can be transferred from myeloma cells to antigen presenting cells (APC), which present processed Id-peptide on their class II molecules to Id-specific T cell receptor-transgenic (TCR-TG) CD4(+) T cells. Id primed tumor APC were heterogeneous, the majority being dendritic cells with class II(+), CD11b(+) CD11c(+) CD40(+) CD80(+) CD86(+) markers. The APC were localized beneath CD31(+) endothelial cells of tumor microvessels, and their frequency declined with tumor progression. The APC could stimulate Id-specific naive TCR-TG, short-term polarized TCR-TG, and cloned CD4(+) T cells to proliferate and produce cytokines in vitro. Furthermore, small MOPC315 tumors established in Id-specific TCR-TG mice contained clusters of activated (CD69(+)CD25(+)) and proliferating (BrdUrd(+)) Id-specific transgenic CD4(+) blasts. The activated Id-specific T cells were located adjacent to Id-primed dendritic cells in the tumor. Thus, a TSA can be transferred in vivo from myeloma, and possibly other types of cancer cells to APC for MHC class II presentation to CD4(+) T cells. PMID- 10706630 TI - Generation and rescue of a murine model of platelet dysfunction: the Bernard Soulier syndrome. AB - The human Bernard-Soulier syndrome is an autosomal recessive disorder of platelet dysfunction presenting with mild thrombocytopenia, circulating "giant" platelets and a bleeding phenotype. The bleeding in patients with the Bernard-Soulier syndrome is disproportionately more severe than suggested by the reduced platelet count and is explained by a defect in primary hemostasis owing to the absence of the platelet glycoprotein (GP) Ib-IX-V membrane receptor. However, the molecular basis for the giant platelet phenotype and thrombocytopenia have remained unresolved but assumed to be linked to an absent receptor complex. We have disrupted the gene encoding the alpha-subunit of mouse GP Ib-IX-V (GP Ibalpha) and describe a murine model recapitulating the hallmark characteristics of the human Bernard-Soulier syndrome. The results demonstrate a direct link between expression of a GP Ib-IX-V complex and normal megakaryocytopoiesis and platelet morphogenesis. Moreover, using transgenic technology the murine Bernard-Soulier phenotype was rescued by expression of a human GP Ibalpha subunit on the surface of circulating mouse platelets. Thus, an in vivo model is defined for analysis of the human GP Ib-IX-V receptor and its role in the processes performed exclusively by megakaryocytes and platelets. PMID- 10706629 TI - Specific association of estrogen receptor beta with the cell cycle spindle assembly checkpoint protein, MAD2. AB - Estrogen receptors (ERs) are ligand-activated transcription factors that regulate gene expression and cell growth. Two ERs now have been identified: ERalpha and the more recently discovered ERbeta. The physiological function of ERbeta remains unclear, but evidence from vascular injury studies and from ERbeta knockout mice suggests that ERbeta may be involved in the regulation of cellular proliferation. Here we show a direct and specific interaction between ERbeta and the cell cycle mitotic spindle assembly checkpoint protein, MAD2 (mitosis arrest-deficient 2). The ERbeta-MAD2 interaction was identified by screening of a yeast two-hybrid system vascular endothelial cell library with ERbeta and confirmed with glutathione S-transferase-fusion protein interaction studies. In contrast, ERalpha did not interact with MAD2 in either the two-hybrid system or in the protein-protein interaction experiments. Amino acids 173-208 in the hinge region of ERbeta were sufficient to mediate the interaction with MAD2 in the two-hybrid system and in glutathione S-transferase-fusion protein studies. These data identify a link between ERbeta and MAD2 of potential importance to regulation of the cell cycle and support a function of ERbeta distinct from the established role of ERs as transcription factors. PMID- 10706631 TI - Isolation of a vascular cell wall-specific monoclonal antibody recognizing a cell polarity by using a phage display subtraction method. AB - Using a strategy consisting of (i) the isolation of cell walls from synchronously differentiating cells of Zinnia, (ii) the generation of mAbs with an antibody phage display method, and (iii) screening with a subtraction method, we isolated mAbs recognizing vascular development-specific cell wall components without prior antigen identification. One of the isolated mAbs, designated CN 8, recognized a cell wall component contained in the hemicellulosic fraction. Immunohistochemical analyses showed that the CN 8 epitope was localized to the cell wall of immature tracheary elements and xylem parenchyma cells. In immature tracheary elements, the CN 8 epitope had a polarized localization pattern regardless of whether the cells are formed as parts of vessels in situ or as single tracheary elements in vitro, suggesting that cell polarity autonomously formed on the cell wall may function in tracheary element differentiation. PMID- 10706632 TI - CD38 expressed on human monocytes: a coaccessory molecule in the superantigen induced proliferation. AB - This work analyzes the hypothesis that human CD38 may cooperate with MHC Class II by acting as coreceptor in a superantigen-induced activation. The initial evidence is that CD38 ligation by specific monoclonal antibodies inhibits superantigen-induced T lymphocyte proliferation. Inhibitory effects become apparent after engagement of CD38 expressed by monocytes, whereas ligation of CD38 expressed by T lymphocytes does not apparently affect activation. The inhibition requires a cell-to-cell interaction, followed by the relevant transmembrane signaling that is reproduced by CD38 ligation. Indeed, CD38 ligation on monocytes induces tyrosine phosphorylation of several intracellular proteins including the protooncogene product c-cbl and the fgr and hck tyrosine kinases. The receptorial nature of the CD38-mediated events is confirmed by the observation of an intracellular calcium flux in monocytes secondary to CD38 ligation. These effects are additive with the similar events elicited by MHC Class II ligation, a likely indication that CD38 and MHC Class II share a common activation pathway. This conclusion is strengthened by results of comodulation experiments, indicating that CD38 and MHC Class II display lateral associations on monocytes. These results attribute to CD38 expressed by monocytes a role in the transduction of signal(s) involved in superantigen-induced activation, operating in synergy with MHC Class II. PMID- 10706633 TI - Decreases in rat extracellular hippocampal glucose concentration associated with cognitive demand during a spatial task. AB - Using in vivo microdialysis, we measured hippocampal extracellular glucose concentrations in rats while they performed spontaneous alternation tests of spatial working memory in one of two mazes. Extracellular glucose levels in the hippocampus decreased by 32% below baseline during the test period on the more complex maze, but by a maximum of 11% on the less complex maze. Comparable decreases were not observed in samples taken from rats tested on the more complex maze but with probes located near but outside of the hippocampus. Systemic glucose fully blocked any decrease in extracellular glucose and enhanced alternation on the more complex maze. These findings suggest that cognitive activity can deplete extracellular glucose in the hippocampus and that exogenous glucose administration reverses the depletion while enhancing task performance. PMID- 10706634 TI - Processive movement of single 22S dynein molecules occurs only at low ATP concentrations. AB - We have analyzed the movement of single 22S dynein molecules from Tetrahymena cilia by using a nanometer measuring system equipped with optical tweezers. Statistical analysis proved that a single molecule of 22S dynein can move processively and develop force at low concentrations of ATP (<20 microM). The maximum force was approximately 4.7 pN, and the force-velocity curve was convex down. During force development, dynein molecules showed stepwise displacement of approximately 8 nm and frequently exhibited backward steps of approximately 8 nm. At higher concentrations of ATP (>/=20 microM) single molecules of 22S dynein were not observed to move processively. Twenty-two S dynein seems to switch over from a processive mode to a nonprocessive mode, sensing a subtle change of ATP concentrations. These observations indicate that the processivity, maximum force, and step size of dynein are similar to those of kinesin, but the ATP concentration-dependence, force-velocity relationship, and backward steps are clearly distinct from kinesin. PMID- 10706635 TI - Structures of recombinant human and mouse NAD(P)H:quinone oxidoreductases: species comparison and structural changes with substrate binding and release. AB - NAD(P)H/quinone acceptor oxidoreductase (QR1, NQO1, formerly DT-diaphorase; EC ) protects animal cells from the deleterious and carcinogenic effects of quinones and other electrophiles. In this paper we report the apoenzyme structures of human (at 1.7-A resolution) and mouse (2.8 A) QR1 and the complex of the human enzyme with the substrate duroquinone (2.5 A) (2,3,5, 6-tetramethyl-p benzoquinone). In addition to providing a description and rationale of the structural and catalytic differences among several species, these structures reveal the changes that accompany substrate or cofactor (NAD) binding and release. Tyrosine-128 and the loop spanning residues 232-236 close the binding site, partially occupying the space left vacant by the departing molecule (substrate or cofactor). These changes highlight the exquisite control of access to the catalytic site that is required by the ping-pong mechanism in which, after reducing the flavin, NAD(P)(+) leaves the catalytic site and allows substrate to bind at the vacated position. In the human QR1-duroquinone structure one ring carbon is significantly closer to the flavin N5, suggesting a direct hydride transfer to this atom. PMID- 10706636 TI - Linking development with generation of novelty in mammalian teeth. AB - The evolution of mammalian teeth is characterized by the frequent and convergent evolution of new cusps. The evolution of new cusps can be linked to tooth development via population-level variation. This allows testing whether development increases the capacity to evolve, or evolvability, by facilitating and even directing morphological change. In a population sample of living seals, variation in cusp number of individual teeth is from three to five cusps, the variably present cusps being the shortest ones that also develop last. By factoring in recent evidence on development, I show that the variation in cusp number can be explained by a patterning cascade mode of cusp development that cumulatively increases and directs height variation in short cusps. The biased variation in seal tooth cusps supports the recognition of teeth as highly evolvable because only small developmental changes are needed to produce large changes in size and number of small cusps. This evolvability of tooth cusps may have facilitated the fast and independent acquisition of new cusps in mammalian evolution. In phylogenetic studies, small cusps may be unreliable as phylogenetic signals. Population level variation can be a powerful tool in testing and generating hypotheses in developmental evolution studies. PMID- 10706637 TI - Human cytomegalovirus UL69 protein is required for efficient accumulation of infected cells in the G1 phase of the cell cycle. AB - Human cytomegalovirus blocks cell-cycle progression in the G(1) compartment upon infection of primary human fibroblasts. The virus-coded UL69 protein can institute a G(1) block when expressed in cells in the absence of virus infection. We have constructed a cytomegalovirus mutant, TNsubUL69, that lacks the UL69 coding region. This virus grows slowly in fibroblasts, but produces a wild-type yield after an extended delay. It grows with normal kinetics in cells coinfected with a recombinant retrovirus, retroUL69, which expresses UL69 protein, demonstrating that its growth defect results from the mutation in the UL69 gene. UL69 protein is packaged within virus particles, and it was possible for us to produce two types of virus stocks. TNsubUL69(+pUL69) lacks the UL69 gene but contains UL69 protein in virus particles. It is produced by growth in fibroblasts that are coinfected with retroUL69. TNsubUL69(-pUL69) lacks the UL69 gene and protein. It is produced by growth in fibroblasts that do not contain UL69 protein. The mutant virions lacking both the UL69 gene and protein fail to induce a cell-cycle block with normal efficiency, whereas the mutant particles lacking the gene but containing the protein can institute the block. These results are consistent with the view that the UL69 protein contributes to the cytomegalovirus induced cell-cycle block, and they suggest that UL69 protein delivered to cells within virions can induce the block without the synthesis of additional UL69 protein encoded by the infecting viral genome. PMID- 10706638 TI - A nonpeptide integrin antagonist can inhibit epithelial cell ingestion of Streptococcus pyogenes by blocking formation of integrin alpha 5beta 1 fibronectin-M1 protein complexes. AB - Streptococcus pyogenes can be efficiently internalized by a variety of human epithelial cells. beta-lactam antibiotics, commonly used to treat S. pyogenes infections, do not readily permeate mammalian cells. There is growing evidence that the ability of streptococci to enter host cells contributes to the frequent failure of antibiotics to eradicate the organism from infected individuals. Recent studies have suggested that host cell entry requires the formation of a complex of a bacterial fibronectin (Fn) binding protein (e.g., M1 protein or protein F1/SfbI), human Fn, and the epithelial cell Fn receptor, integrin alpha5beta1. We report here that a low molecular weight, nonpeptide antagonist of integrin alpha5beta1, SJ755, can inhibit internalization of streptococci by primary human tonsillar epithelial cells and immortalized human epithelial (A549) cells, thus increasing the extent of bacterial killing by antibiotics. SJ755 blocked Fn binding by human tonsillar epithelial and A549 cells, suggesting that integrin alpha5beta1 is the major Fn receptor expressed by both cell types. SJ755 did not affect Fn binding by purified M1 protein or M1(+) bacteria. Purified M1 protein failed to associate with integrin alpha5beta1 unless the integrin had been prebound by Fn. Also, SJ755 blocked formation of alpha5beta1-Fn-M1 complexes in vitro. These results support the previous proposal that Fn functions as a molecular bridge between M1 protein and integrin alpha5beta1. Furthermore, these results suggest that integrin antagonists may enhance the efficacy of antibiotics in treatment of S. pyogenes infections. PMID- 10706639 TI - Clustering of connexin 43-enhanced green fluorescent protein gap junction channels and functional coupling in living cells. AB - Communication-incompetent cell lines were transfected with connexin (Cx) 43 fused with enhanced green fluorescent protein (EGFP) to examine the relation between Cx distribution determined by fluorescence microscopy and electrical coupling measured at single-channel resolution in living cell pairs. Cx43-EGFP channel properties were like those of wild-type Cx43 except for reduced sensitivity to transjunctional voltage. Cx43-EGFP clustered into plaques at locations of cell cell contact. Coupling was always absent in the absence of plaques and even in the presence of small plaques. Plaques exceeding several hundred channels always conferred coupling, but only a small fraction of channels were functional. These data indicate that clustering may be a requirement for opening of gap junction channels. PMID- 10706640 TI - Recombinant rabies virus as potential live-viral vaccines for HIV-1. AB - Recombinant, replication-competent rabies virus (RV) vaccine strain-based vectors were developed expressing HIV type I (HIV-1) envelope glycoprotein (gp160) from both a laboratory-adapted (CXCR4-tropic) and a primary (dual-tropic) HIV-1 isolate. An additional transcription stop/start unit within the RV genome was used to express HIV-1 gp160 in addition to the other RV proteins. The HIV-1 gp160 protein was stably and functionally expressed, as indicated by fusion of human T cell lines after infection with the recombinant RVs. Inoculation of mice with the recombinant RVs expressing HIV-1 gp160 induced a strong humoral response directed against the HIV-1 envelope protein after a single boost with recombinant HIV-1 gp120 protein. Moreover, high neutralization titers up to 1:800 against HIV-1 could be detected in the mouse sera. These data indicate that a live recombinant RV, a rhabdovirus, expressing HIV-1 gp160 may serve as an effective vector for an HIV-1 vaccine. PMID- 10706641 TI - Selecting protein targets for structural genomics of Pyrobaculum aerophilum: validating automated fold assignment methods by using binary hypothesis testing. AB - Three-dimensional protein folds were assigned to all ORFs of the recently sequenced genome of the hyperthermophilic archaeon Pyrobaculum aerophilum. Binary hypothesis testing was used to estimate a confidence level for each assignment. A separate test was conducted to assign a probability for whether each sequence has a novel fold-i.e., one that is not yet represented in the experimental database of known structures. Of the 2,130 predicted nontransmembrane proteins in this organism, 916 matched a fold at a cumulative 90% confidence level, and 245 could be assigned at a 99% confidence level. Likewise, 286 proteins were predicted to have a previously unobserved fold with a 90% confidence level, and 14 at a 99% confidence level. These statistically based tools are combined with homology searches against the Online Mendelian Inheritance in Man (OMIM) human genetics database and other protein databases for the selection of attractive targets for crystallographic or NMR structure determination. Results of these studies have been collated and placed at http://www.doe-mbi.ucla.edu/people/parag/P A_HOME/, the University of California, Los Angeles-Department of Energy Pyrobaculum aerophilum web site. PMID- 10706642 TI - Mating experience and juvenile hormone enhance sexual signaling and mating in male Caribbean fruit flies. AB - Young mated male Caribbean fruit flies [Anastrepha suspensa (Loew)] have greater sexual prowess than their virgin counterparts. After mating for the first time, 6 to 7-day-old males released twice as much sex pheromone and acquired another mate in less than half the time required by virgin males of the same age. Mass spectroscopic analysis of extracts of hemolymph from mated and virgin 7-day-old males resulted in identification of juvenile hormone III bisepoxide and juvenile hormone III in a ratio of 2.5:1. Extracts from mated males contained 3-fold more juvenile hormone than did extracts from virgins. Enhancement of sexual signaling, pheromone release, and mating was induced by topical application of juvenile hormone, methoprene, or fenoxycarb. Newly eclosed adult males treated with juvenoids engaged in sexual signaling, released pheromone, and mated at significantly earlier ages than control males. We conclude that juvenile hormone mediated a positive feedback system that imparted a competitive advantage, guaranteeing that males who mated at an early age would out-compete virgins of the same age for mating opportunities. Additionally, the results support the hypothesis that juvenile hormone is a pivotal hormone coordinating the development of sexual signaling and reproductive maturity in these flies. PMID- 10706643 TI - High-dose chemotherapy and peripheral blood progenitor cell transplantation in the treatment of breast cancer. AB - Each year in the USA, 180,000 new cases of breast cancer are diagnosed and about 44,000 women die of the disease. Current primary treatment consists of adjuvant chemotherapy and hormone therapy, and statistics show that combination chemotherapy favorably influences the outcomes in both node-negative and node positive primary disease. However, a significant number of breast cancer patients succumb to the disease, and nearly every patient diagnosed with metastatic breast cancer will be dead within five years. High-dose chemotherapy (HDC) and peripheral blood progenitor cell transplantation (PBPCT) are based upon laboratory and clinical observations of the ability to modify growth properties of quiescent and replicating cancer cells. A large number of HDC and PBPCT regimens have been evaluated for treatment of metastatic breast cancer, and recent autologous bone marrow transplantation data indicate that three HDC regimens (CPB, CTCb and cytoxan and thiotepa) predominate. Unfortunately, negative media coverage surrounding and subsequent to the presentation of preliminary findings reported at the May 1999 American Society of Clinical Oncologists, that were not allowed adequate follow-up time for full analysis of treatment results, has had a detrimental effect on the ability to conduct trials in this area. Several randomized trials have been conducted in both the metastatic and high risk primary disease settings. Thorough analysis of these studies indicates an evaluable improvement in favor of HDC and PBPCT in three of the four randomized studies performed in metastatic breast cancer and two of the four high risk primary studies. Also, initial evaluations found that quality of life appeared comparable in patients receiving either HDC or not. Each randomized trial studied asks a different question and, depending on the intensity of HDC regimen, the intensity and duration of the standard dose chemotherapy control and the schedule of events in relation to induction chemotherapy, the outcomes may be quite variable. Still, certain general trends are indentifiable. HDC alone will not completely cure breast cancer and should be considered as part of an overall therapeutic plan. In some of these studies, significantly longer follow-up is required before definitive analysis can be completed. PMID- 10706645 TI - Editor's Note. PMID- 10706644 TI - High dose chemotherapy for breast cancer: taking stock. PMID- 10706646 TI - The role of intraoperative radiation therapy (IORT) in the treatment of locally advanced gynecologic malignancies. AB - The prognosis in women with locally advanced primary or recurrent gynecologic malignancies is rather poor. Doses of external beam radiation necessary to treat gross or microscopic recurrence among patients surgically treated or previously irradiated exceed what is tolerated by normal structures. In this group of patients, intraoperative radiation therapy (IORT) can be utilized to maximize local tumor control, minimizing the radiation exposure of dose-limiting surrounding structures. Review of the available literature indicates that IORT may improve long-term local control and overall survival in women with pelvic sidewall and/or para-aortic nodal recurrence. The most encouraging results have been reported in the cases of microscopic residual disease, following surgical debulking. PMID- 10706647 TI - Second-line treatment of ovarian cancer. AB - Patients with epithelial ovarian cancer are often diagnosed with advanced-stage disease. Although clinical complete remissions are obtained in the majority of patients through a combination of cytoreductive surgery and chemotherapy, relapse is common. A number of agents with diverse biologic mechanisms have been identified with activity in the setting of recurrent disease. Strategies for management of patients with recurrent disease, including classification, treatment goals, and therapeutic options will be reviewed. PMID- 10706648 TI - Management of advanced prostate cancer. AB - Most cases of advanced carcinoma of the prostate are hormonosensitive. The use of combined androgen blockade (CAB) seems to improve survival and quality of life, but only when combined with chemical castration by luteinizing-hormone-releasing hormone analog and without the use of steroidal antiandrogens. After CAB, further hormonal treatments remain efficacious, such as antiandrogen withdrawal followed by estrogens, aromatase inhibitors, and hormone-refractory prostate cancer multiple cytotoxic agents. For painful bone lesions, external beam radiotherapy, biphosphonates, and strontium 89 or samarium 153 provide pain relief. The use of new methods for the evaluation of response and quality of life will allow the rapid identification of effective treatments and permit powered phase III trials. PMID- 10706649 TI - Intermittent androgen deprivation in prostate cancer patients: factors predictive of prolonged time off therapy. AB - OBJECTIVES: We hypothesize that prostate cancer (PC) patients who achieve and maintain an undetectable prostate-specific antigen (UD-PSA) on androgen deprivation therapy (ADT) have a predominantly androgen-dependent cancer cell population sensitive to apoptosis that allows for a prolonged time off ADT. This study summarizes patient- and treatment-related factors associated with a prolonged time off ADT in patients electing intermittent androgen deprivation (IAD). METHODS: Hormone-naive patients with PC were treated with ADT using an antiandrogen and a luteinizing-hormone-releasing hormone-agonist. Of 255 consecutive patients, 216 (85%) achieved a UD-PSA (< 0.05 ng/ml). Ninety-three (43%) of 216 elected to stop ADT after maintaining a UD-PSA for a median of one year. Patients were followed off therapy and advised to restart ADT if the PSA level reached > or = 5.0 ng/ml. Forty-one patients received finasteride as part of IAD induction and as maintenance off therapy; these patients are excluded from the current study and are the focus of another publication. The remaining 52 patients are assessable for response being either in the off-phase of IAD > or = 1 year or having restarted IAD. RESULTS: In the first IAD cycle, the median duration of the on-phase of IAD was 16 months (mean 19.0 months, range 3.6-71 months), and the median off-phase duration was 15.5 months (mean 24.1 months, range 3.2-87+ months). In 28 patients who maintained a UD-PSA for > or = 1 year, their median off-phase duration was 29 months (mean 35.8 months, range 7.8-87+ months), with nine (32%) still off IAD after a median follow-up of 62 months. Significant (p < 0.05) independent factors associated with prolonged off-phase duration by multivariate analysis included UD-PSA on ADT > or = 1 year (p = 0.010), PSA-only recurrence after local therapy (p = 0.039), and reaching a testosterone level > or = 150 ng/dl in > or = 4 months off ADT (p = 0.041). After a median of 66 months of follow-up, only one (2%) patient developed androgen independent PC. CONCLUSIONS: Hormone-naive patients who achieve and maintain a UD PSA for at least one year during ADT may initiate IAD and anticipate a prolonged off-phase duration. Attainment of a UD-PSA on ADT may serve as an in vivo sensitivity test of a patient's tumor cell population, and allow for better selection of those best suited for IAD. PMID- 10706650 TI - Sedation for intractable distress of a dying patient: acute palliative care and the principle of double effect. AB - Shortly before his death in 1995, Kenneth B. Schwartz, a cancer patient at Massachusetts General Hospital (MGH), founded the Kenneth B. Schwartz Center at MGH. The Schwartz Center is a nonprofit organization dedicated to supporting and advancing compassionate health care delivery, which provides hope to the patient, support to caregivers, and encourages the healing process. The Center sponsors the Schwartz Center Rounds, a monthly multidisciplinary forum where caregivers reflect on important psychosocial issues faced by patients, their families, and their caregivers, and gain insight and support from fellow staff members. The case presented is of a young man dying of recurrent epithelioid hemangioendothelioma, distressed with stridor and severe pain, whose poorly controlled symptoms were successfully treated with an infusion of propofol, titrated to provide effective comfort in the last few hours of the patient's life. The tenet of double effect, which allows aggressive treatment of suffering in spite of foreseeable but unintended consequences, is reviewed. The patient's parents were invited and contributed to the Rounds, providing compelling testimony to the power of the presence of clinicians at the time of death and the importance of open communication about difficult ethical issues. PMID- 10706651 TI - Waldenstrom's macroglobulinemia. AB - Waldenstrom's macroglobulinemia is a low-grade lymphoplasmacytic lymphoma. It has an overall incidence of 2.5/million/year. The median age at diagnosis is 63 years. The clinical manifestations are hepatomegaly (20%), splenomegaly (15%), and lymphadenopathy (15%). The most common symptom is fatigue related to a normochromic, normocytic anemia, and the median hemoglobin value at diagnosis is 10 gm/dl. All patients with Waldenstrom's macroglobulinemia have a circulating tumor marker, the monoclonal IgM protein. Occasionally high levels of the IgM monoclonal protein can produce a hyperviscosity syndrome manifested by oronasal bleeding. Occasionally retinal hemorrhage or serious neurologic complications, such as somnolence or coma, may occur. The most important prognostic factors are hemoglobin, age, weight loss, and a cryoglobulin. Therapy has included alkylating agents, particularly chlorambucil, purine nucleoside analogs such as fludarabine or cladribine, and most recently the use of rituximab. The median survival of symptomatic patients is 65 months. Patients without symptoms should not be treated. PMID- 10706652 TI - Cellular suicide therapy of malignant disease. AB - Adoptive cellular therapy is developing as a supplement or alternative to chemotherapy and/or radiation for malignant disease. Our focus is two ongoing clinical studies with transgeneic (genetically altered) cellular therapy; one uses allogeneic (from another person) lymphocytes to treat leukemia, and the second uses xenogeneic (from another species) fibroblast cells genetically altered to contain a toxin-producing suicide gene to treat ovarian cancer. Allogeneic donor lymphocyte infusions (DLI) are known to induce remission of hematologic malignancies. However, the toxicity associated with DLI is related to graft-versus-host-disease, which is due to donor lymphocytes attacking normal tissue in the recipient. Therefore, we have taken the approach of infusing DLI that have been modified to contain a latent suicide gene to treat leukemia. To treat ovarian cancer, we used xenogeneic nonimmune fibroblast-derived cells to deliver a tumor-directed cytotoxic gene to carcinoma cells. These cells release HStk transgene retroviruses that in turn transduce replicating tumor cells but not quiescent epithelium, rendering the tumor selectively susceptible to ganciclovir-mediated killing. These initial trials summarize the early stage of allogeneic/xenogeneic adoptive cellular therapy for cancer, and although the data are limited, it is encouraging to see some patients with evidence of antitumor responses. Advances in our understanding of the basic science of these treatments, together with improvements in the technology of vector design, will be required to stream-line these methodologies into broader application. PMID- 10706653 TI - Chemokines induce moesin interaction with ICAM-3. PMID- 10706654 TI - Viral cytokines. PMID- 10706655 TI - The molecular perspective: DNA. PMID- 10706656 TI - The Oncologist News Bulletin. PMID- 10706657 TI - A group I intron in the nuclear small subunit ribosomal DNA of Gaeumannomyces graminis. AB - The length of the small subunit ribosomal DNA (SSU rDNA) differs among isolates of species and varieties of Gaeumannomyces. The sequence of the 3' region of the SSU rDNA revealed 340-, 365-, and 520-bp insertions for G. graminis varieties avenae, tritici, and graminis, respectively. The intron sequences from varities tritici and avenae were similar, except there was an insert of 23 nucleotides at base 328 from the 5' end of the G. g. var. tritici intron. The G. g. var. graminis intron sequences had 92.4% homology compared with the intron sequences of varieties tritici and avenae. In addition, the intron sequence of variety graminis is larger, having an insert of 155 nucleotides at base 365 of the 5' end of the intron. Little variation in the DNA sequences flanking the introns has been detected among the isolates of Gaeumannomyces that either have or lack an intron. Reverse transcriptase PCR (RT-PCR) indicated the absence of the intron in the mature rRNA. The intron sequence had both a conserved sequence and secondary structural elements classifying it as a group I intron. PMID- 10706658 TI - Spectral light quality affects protein profile of Synechococcus sp. PCC 7942: a comparative 2-dimensional gel electrophoresis (2-DGE) analysis. AB - We report here a comparative analysis of the effect of blue (450 nm), red (660 nm), and white light (400-700 nm) on the protein profile of cyanobacteria Synechococcus sp. PCC 7942. In vivo labeling of cells with [(35)S] methionine and their subsequent analysis by two-dimensional gel electrophoresis (2-DGE) showed that eight polypeptides were unique to dark adapted cells, ten were blue light specific, and four were specifically induced in red light. The results show that Synechococcus sp. respond to various light treatments rapidly and synthesize new polypeptides in dark and blue/red light. PMID- 10706659 TI - Development of a plasmid vector for easy selection of strong promoters. AB - A promoter vector pACPR33 for Escherichia coli based on the promotorless ampicillin-resistance gene from pBR322 has been constructed. The promoter of the ampicillin-resistance gene was deleted and replaced by a suitable multiple cloning site. Molecular cloning of promoters into the polylinker resulted in activation of the ampicillin resistance in E. coli. The plasmid contains a functional origin of DNA replication and a tetracycline resistance gene for E. coli, and a chloramphenicol resistance gene for S. aureus. The vector permitted direct detection of promoter activity, especially strong promoters, by easy iodometric determination of beta-lactamase activity in liquid or solid media. PMID- 10706660 TI - Lipase activity from strains of Pasteurella multocida. AB - Thirteen clinical isolates of Pasteurella multocida from a variety of different animals and humans were examined for their ability to produce lipase. Lipase substrates used included Tween 20, Tween 40, Tween 80, and Tween 85. Lipase activity was detected in the filtrates of organisms grown to the exponential phase in Roswell Park Memorial Institute-1640 defined media (RPMI-1640), but activity increased in the filtrates when the cultures were allowed to proceed to the stationary phase. All strains examined (except for serotype 2) showed lipase activity against at least one of the Tweens. Tween 40 was the best substrate to demonstrate lipase activity. Pasteurella multocida serotype 8 produced the most active lipase against Tween 40 (3,561.7 units of activity/microgram of protein). This activity continued to increase after P. multocida entered a stationary growth phase. P. multocida lipase activity was optimal at pH 8.0. Lipase activity of P. multocida serotype 8 was eluted from a Sepharose 2B column at several points, indicating that several lipases may be produced in vitro by this organism. These data demonstrate that clinical isolates of P. multocida produce lipase; therefore, this enzyme should be considered a potential virulence factors for this organism. PMID- 10706661 TI - Influence of water potential on growth, enzyme secretion and in vitro enzyme activities of Trichoderma harzianum at different temperatures. AB - The influence of water potential on linear mycelial growth, secretion, and the in vitro activities of enzymes beta-glucosidase, cellobiohydrolase, beta-xylosidase, exochitinase, and chymotrypsin of Trichoderma harzianum strain T66 was studied at different temperatures. Nearly linear correlation was found between water potential and colony growth rate at both 25 degrees C and 10 degrees C, with higher growth rates at the higher temperature and higher water potentials. The amounts of enzyme secretion depended on the water potential and not on the quality of salt (NaCl or KCl) used as osmoticum. Enzyme activities were significantly affected by water potential. Significant enzyme activities were measured for most of the enzymes even at -14.800 megapascal (MPa), which is below the water potential where mycelial growth ceased. These results suggest the possibility of using mutants with improved xerotolerance for biocontrol purposes in soils with lower water potentials. PMID- 10706662 TI - Transcriptional analysis of hydrogenase genes in the Cyanobacteria Anacystis nidulans and Anabaena variabilis monitored by RT-PCR. AB - Diverse cyanobacteria express an uptake hydrogenase, encoded by the genes hupSL, and a bidirectional, NAD(P)(+)-reducing hydrogenase with the genes hox(E)FUYH. In the unicellular Anacystis nidulans, the hox genes are organized on two separate loci, whereas they are contiguous in one cluster, though interspersed with two unidentified reading frames, ORF 3 and 8, in the heterocystous Anabaena variabilis. The hox gene clusters of these two cyanobacteria have now been transcriptionally analyzed by RT-PCR. A polycistronic transcript was identified in both cyanobacteria. In A. nidulans, one message for each locus has been detected, the dicistronic hoxEF unit, and the polycistronic hoxUYHWhypAB one. In A. variabilis, the transcript consists of the hox genes hoxFUYH as well as the unidentified ORFs. Previous enzyme determinations on the occurrence of the uptake hydrogenase in vegetative cells and thus outside of heterocysts gave ambiguous results. Therefore, transcription of both hup and hox genes has been analyzed in both heterocysts and vegetative cells of A. variabilis. A hupL transcript is detectable in heterocysts and also, though less extensive but clearly discernible, in vegetative cells of NH(4)(+)-grown A. variabilis. PMID- 10706663 TI - Coexpression of cyt1Aa of Bacillus thuringiensis subsp. israelensis with Bacillus sphaericus binary toxin gene in acrystalliferous strain of B. thuringiensis. AB - The cyt1Aa gene of Bacillus thuringiensis subsp. israelensis and binary toxin gene of Bacillus sphaericus C3-41 were introduced into an acrystalliferous strain of B. thuringiensis independently and in combination by using shuttle vector pBU4. SDS-PAGE and Western blot analysis proved that cyt1Aa and binary toxin genes coexpressed during the sporulation of the recombinant. Transformant strain expressing the Cyt1Aa and binary toxin proteins in combination was more toxic to susceptible and resistant Culex pipiens quinquefasciatus than the transformants expressing Cyt1Aa protein or binary toxin proteins independently. It was suggested that large amount of production of Cyt1Aa protein and binary toxin proteins possibly interacted synergistically, thereby increasing its mosquitocidal toxicity significantly. PMID- 10706664 TI - Isolation and identification of ruminal methanogens from grazing cattle. AB - To obtain information on the diversity of ruminal methanogens in grazing animals, three ruminal methanogens from grazing cattle were characterized and identified. Two of the isolates were rod-shaped, with one staining Gram-positive and being non-motile (BRM9), and the other (BRM16) staining Gram-negative and being motile. These isolates grew only on H(2)/CO(2) and formate, and optimally at 38 degrees C and pH 6.5-7.0. The third isolate (CM1) was non-motile, pseudosarcina-shaped, and grew on H(2)/CO(2), acetate, and methyl-containing compounds, with optimal growth at 40 degrees C and pH 6.5. DNA was prepared from the three isolates, and their 16S rRNA genes were sequenced. Phenotypic data and comparisons of nearly complete 16S rDNA sequences showed that BRM9, BRM16, and CM1 are strains of Methanobacterium formicicum, Methanomicrobium mobile, and Methanosarcina barkeri respectively. To the best of our knowledge, this is the first information on ruminal methanogens in cattle maintained under grazing management. PMID- 10706665 TI - Multidomain and multifunctional glycosyl hydrolases from the extreme thermophile Caldicellulosiruptor isolate Tok7B.1. AB - DNA sequencing techniques have revealed widespread molecular diversity of the genomic organization of apparently closely related bacteria (as judged from SSU rDNA sequence similarity). We have previously described the extreme thermophile Caldicellulosiruptor saccharolyticus, which is unusual in possessing multi catalytic, multidomain arrangements for the majority of its glycosyl hydrolases. We report here the sequencing of three gene clusters of glycosyl hydrolases from Caldicellulosiruptor sp. strain Tok7B.1. These clusters are not closely linked, and each is different in its organization from any described for Cs. saccharolyticus. The catalytic domains of the enzymes belong to glycosyl hydrolase families 5, 9, 10, 43, 44, and 48. The cellulose binding domains (CBDs) of these enzymes from Caldicellulosiruptor sp. Tok7B.1 are types IIIb, IIIc, or VI. A number of individual catalytic and binding domains have been expressed in Escherichia coli, and biochemical data are reported on the purified enzymes for cellulose degradation encoded by engineered derivatives of celB and celE. PMID- 10706666 TI - A new Mesorhizobium loti HAMBI 1129 phage isolated from Polish soil. AB - Phage A1 isolated from the rhizosphere of Lotus corniculatus was studied. It had a very narrow host range, as it was active only against Mesorhizobium loti HAMBI 1129. Phage A1 was classified as belonging to C Bradley's group bacteriophages. The latent period of A1 was 120-130 min and a burst size 13-17 particles per cell. The nature of the phage receptor was examined. Lipopolysaccharide from the phage-sensitive strain inactivated phage A1 in contrast to LPS from the phage resistant bacteria. Purified LPS obtained from M. loti HAMBI 1129 had a high receptor activity with PhI(50) value of 0. 025 microgram/ml. PMID- 10706667 TI - Dissimilatory Fe(III) oxide reduction by Shewanella alga BrY requires adhesion. AB - The Derjaguin-Landau-Verwey-Overbeek (DLVO) theory was used to examine the relationship between adhesion and dissimilatory Fe(III) oxide reduction. Adhesion of Shewanella alga BrY to hydrous ferric oxide (HFO) was correlated with ionic strength and thus was accurately described by the DLVO theory. Reduction of insoluble HFO was also correlated with KCl concentration. In contrast, there was no correlation between soluble Fe(III) reduction and ionic strength. A correlation between HFO reduction rate and adhesion to HFO was observed. These results provide direct evidence that adhesion is requisite for Fe(III) oxide reduction in the absence of soluble electron shuttles. PMID- 10706668 TI - Cutting edge: identification of a novel chemokine receptor that binds dendritic cell- and T cell-active chemokines including ELC, SLC, and TECK. AB - Searching for new receptors of dendritic cell- and T cell-active chemokines, we used a combination of techniques to interrogate orphan chemokine receptors. We report here on human CCX CKR, previously represented only by noncontiguous expressed sequence tags homologous to bovine PPR1, a putative gustatory receptor. We employed a two-tiered process of ligand assignment, where immobilized chemokines constructed on stalks (stalkokines) were used as bait for adhesion of cells expressing CCX CKR. These cells adhered to stalkokines representing ELC, a chemokine previously thought to bind only CCR7. Adhesion was abolished in the presence of soluble ELC, SLC (CCR7 ligands), and TECK (a CCR9 ligand). Complete ligand profiles were further determined by radiolabeled ligand binding and competition with >80 chemokines. ELC, SLC, and TECK comprised high affinity ligands (IC50 <15 nM); lower affinity ligands include BLC and vMIP-II (IC50 <150 nM). With its high affinity for CC chemokines and homology to CC receptors, we provisionally designate this new receptor CCR10. PMID- 10706669 TI - Cutting edge: the Ets1 transcription factor is required for the development of NK T cells in mice. AB - Ets1-deficient mice develop B and T cells but display a severe defect in the development of the NK cell lineage. In this report, we demonstrate that Ets1 is also required for the development of NK1.1+ T (NK T) cells. We observed significantly decreased numbers of NK T cells in the thymus, spleen, and liver of Ets1-deficient mice. These organs also contained markedly decreased levels of the canonical Valpha14-Jalpha281 TCRalpha transcript seen in NK T cells. Unlike wild type NK T cells, Ets1-deficient thymocytes failed to produce detectable levels of IL-4 following anti-CD3 stimulation. The absence of NK T cells in the Ets1 deficient mice was not associated with defective expression of CD1, an MHC class I molecule required for NK T cell development. We conclude that Ets1 defines a novel transcriptional regulatory pathway that is required for the development of both the NK and NK T cell lineages. PMID- 10706670 TI - Cutting edge: ectopic expression of the IL-12 receptor-beta 2 in developing and committed Th2 cells does not affect the production of IL-4 or induce the production of IFN-gamma. AB - The IL-12 receptor-beta 2 (IL-12R beta 2) chain is expressed on Th1 cells and lost upon differentiation to the Th2 phenotype. This has been suggested as the basis for commitment of Th1 cells, because early differentiated Th2 cells do not reverse their phenotype and do not produce IFN-gamma on restimulation in the presence of IL-12. In this study, we ectopically expressed the IL-12 receptor beta 2 (IL-12R beta 2) bicistronically with enhanced green fluorescent protein by retroviral infection in developing and committed Th2 cells. Restimulation of Th2 cells expressing this ectopic IL-12R beta 2 in the presence of IL-12 led to levels of IL-4 production similar to those in control Th2 cells. The expression of IL-12R beta 2 in Th2 cells did not lead to significant levels of IFN-gamma production, although IL-12-mediated STAT signaling and proliferation were restored. Thus, although the IL-12R beta 2 and IL-12-dependent STAT4 activation are required for Th1 responses, activation of this pathway is not sufficient to restore a Th1 phenotype in developing or committed Th2 cells. PMID- 10706671 TI - Cutting edge: WIP, a binding partner for Wiskott-Aldrich syndrome protein, cooperates with Vav in the regulation of T cell activation. AB - Wiskott-Aldrich syndrome protein (WASP)-interacting protein (WIP), specifically binds to a region of WASp that is frequently mutated in Wiskott-Aldrich syndrome. Due to the similar phenotypes of WASp- and Vav-deficient T cells, and the putative importance of the WIP/WASp complex in mediating normal signals from the TCR, we investigated the role of WIP in regulating NF-AT/AP-1-mediated gene transcription. We show that WIP has the ability to enhance Vav-mediated activation of NF-AT/AP-1 gene transcription. In addition, we provide evidence that the interaction of WIP with WASp is necessary, but not sufficient for the ability of WIP to regulate NF-AT/AP-1 activity. Finally, we have identified a region in WIP required for its regulation of NF-AT/AP-1 activity. Our data suggests that the WIP-WASp interaction is important for NF-AT/AP-1-mediated gene transcription, and that defects seen in the activation of T cells from WAS patients may be due to the inability of these cells to form a functional WIP/WASp signaling complex. PMID- 10706672 TI - A transcriptional defect underlies B lymphocyte dysfunction in a patient diagnosed with non-X-linked hyper-IgM syndrome. AB - To establish the underlying cause of hyper-IgM syndrome in one female patient, B cell function was examined in response to CD40- and IL-4-mediated pathways. When CD40-induced functional responses were measured in unfractionated B cells, CD80 up-regulation, de novo Cmu-Cgamma recombination, and Igamma transcription were all found to be relatively unaffected. However, CD40- and IL-4-mediated CD23 up regulation and VDJ-Cgamma transcription were clearly diminished compared to control cells. IL-4-induced CD23 expression was measurably reduced in the CD20- population as well. These results suggested that the patient's defect is positioned downstream of CD40 contact and affects both CD40- and IL-4 signal transduction pathways. Further analysis of B cell function in CD19+ B cells revealed a clear B cell defect with respect to Igamma and mature VDJ-Cgamma transcription and IgG expression. However, under the same conditions Iepsilon transcription was relatively normal. Partial restoration of B cell function occurred if PBMC or CD19+ B cells were cultured in vitro in the presence of CD154 plus IL-4. Because addition of IL-4 to cocultures containing activated T cells failed to induce B cells to undergo differentiation, the ability of the patient's B cells to acquire a responsive phenotype correlated with receiving a sustained signal through CD40. These findings support a model in which the patient expresses an intrinsic defect that is manifested in the failure of specific genes to become transcriptionally active in response to either CD154 or IL-4 and results in a functionally unresponsive B cell phenotype. PMID- 10706673 TI - A population of in vivo anergized T cells with a lower activation threshold for the induction of CD25 exhibit differential requirements in mobilization of intracellular calcium and mitogen-activated protein kinase activation. AB - Chronic exposure of mature T cells with specificity for self-Ags can lead to the induction of a nonfunctional state which is referred to as T cell anergy. It is unclear whether anergic T cells are destined for cell death and thereby harmless or whether they can contribute to the induction of autoimmunity and/or regulation of anti-self reactivity. We have begun to address this issue. In a recent study, we showed that a population of mature CD4-CD8- T cells that express a transgenic TCR specific for the Ld MHC class I molecule are rendered anergic in Ld expressing mice. In this study, we show that this population of anergic T cells possess a lower activation threshold for the induction of CD25 and CD69 in response to stimulation by antigenic ligands. Furthermore, these anergic T cells undergo extensive proliferation when stimulated with a low-affinity ligand in the presence of an exogenous source of IL-2. Biochemical analysis of the early intracellular signaling events of these in vivo anergized T cells showed that they have a signaling defect at the level of ZAP-70 and linker for the activation of T cell (LAT) phosphorylation. They also exhibit a defect in mobilization of intracellular calcium in response to TCR signaling. However, these anergic T cells demonstrate no defect in SLP-76 phosphorylation and extracellular signal regulated kinase 1/2 activation. These biochemical characteristics of the anergic T cells were associated with an elevated level of Fyn, but not Lck expression. The potential contributions of these anergic T cells in the induction and/or regulation of autoimmune responses are discussed. PMID- 10706674 TI - A bone marrow-derived APC in the gut-associated lymphoid tissue captures oral antigens and presents them to both CD4+ and CD8+ T cells. AB - We have previously reported that feeding OVA to C57BL/6 mice can lead to a weak CTL response that is dependent on CD4+ T cell help and is capable of causing autoimmunity. In this study, we investigated the basis of the class I and class II-restricted Ag presentation required for such CTL induction. Two days after feeding OVA, Ag-specific CD4+ and CD8+ T cells were seen to proliferate in the Peyer's patches and mesenteric lymph nodes. Little proliferation was evident in other lymphoid tissues, except at high Ags doses, in which case some dividing CD4+ T cells were observed in the spleen and peripheral lymph nodes. Using chimeric mice, the APC responsible for presenting orally derived Ags was shown to be derived from the bone marrow. Examination of the Ag dose required to activate either CD4+ or CD8+ T cells indicated that a single dose of 6 mg OVA was the minimum dose that consistently stimulated either T cell subset. These data indicate that oral Ags can be transported from the gut into the gut-associated lymphoid tissue, where they are captured by a bone marrow-derived APC and presented to both CD4+ and CD8+ T cells. PMID- 10706675 TI - TRAIL (Apo2 ligand) and TWEAK (Apo3 ligand) mediate CD4+ T cell killing of antigen-presenting macrophages. AB - The human marrow produces approximately 1010 monocytes daily, and this production must be balanced by a similar rate of destruction. Monocytes/macrophages can undergo apoptosis after activating CD4+ T cells, suggesting one mechanism that may contribute to macrophage homeostasis. Previous reports indicate that Fas-Fas ligand interactions are the principle molecules mediating this response. However, D10, an Iak-restricted cloned Th2 line, will similarly induce apoptosis in Ag presenting macrophages, and D10 cells lack Fas ligand. To confirm that D10 cells kill macrophages through Fas-independent pathways, D10 cells were shown to kill MRL lpr/lpr (Iak) macrophages in an Ag-dependent fashion, indicating additional mechanisms. Recent reports demonstrate that TNF-related apoptosis-inducing ligand (TRAIL), interacting with Apo2, and TNF-like weak inducer of apoptosis (TWEAK), interacting with Apo3, will induce apoptosis in some cells. Using Abs to TRAIL and an Apo3-IgG Fc fusion protein, we demonstrated that D10 cells express both TRAIL and TWEAK. The Apo3 fusion protein, but not human IgG, inhibited D10 induced macrophage apoptosis, as did anti-TRAIL. Further studies demonstrated that AE7, a cloned Th1 line, and splenic T cells express TWEAK, TRAIL, and Fas ligand, and inhibiting these molecules also inhibited macrophage killing. These results indicate that D10 cells induce macrophage apoptosis through TRAIL- and TWEAK-dependent pathways. Because normal T cells also express these molecules, these results support the concept that T cells have multiple pathways by which to induce macrophage apoptosis. These pathways may be important in immune processes such as macrophage homeostasis as well as in down-regulation of immune responses and elimination of macrophages infected with intracellular organisms. PMID- 10706676 TI - Normal lymphoid homeostasis and lack of lethal autoimmunity in mice containing mature T cells with severely impaired IL-2 receptors. AB - The importance of IL-2Rbeta function for immune regulation is highlighted by the severe impairment in lymphoid cell function in IL-2Rbeta-deficient mice. It has been speculated that failed IL-2/IL-2R signaling in peripheral T cells causes the associated autoimmunity, imbalanced peripheral lymphoid homeostasis, and defective T cell function. This study explored the requirement for IL-2Rbeta function in mature T lymphocytes. We show that transgenic thymic expression of the IL-2R beta-chain in IL-2Rbeta-deficient mice prevents lethal autoimmunity, restores normal production of B lymphocytes, and results in a peripheral T cell compartment that is responsive to triggering through the TCR, but not the IL-2R. The dysfunction of the IL-2R is illustrated by the near complete failure of mature T cells to proliferate to IL-2 in vitro and in vivo, to differentiate into CTL, and to up-regulate IL-2Ralpha expression. These data indicate that lymphoid homeostasis is largely maintained despite a nonfunctional IL-2R in mature T lymphocytes and suggest that IL-2Rbeta provides an essential signal during thymic development to regulate self-reactivity. PMID- 10706677 TI - Effects of geldanamycin, a heat-shock protein 90-binding agent, on T cell function and T cell nonreceptor protein tyrosine kinases. AB - The benzoquinoid ansamycins geldanamycin (GA), herbimycin, and their derivatives are emerging as novel therapeutic agents that act by inhibiting the 90-kDa heat shock protein hsp90. We report that GA inhibits the proliferation of mitogen activated T cells. GA is actively toxic to both resting and activated T cells; activated T cells appear to be especially vulnerable. The mechanism by which GA acts is reflected by its effects on an essential hsp90-dependent protein, the T cell-specific nonreceptor tyrosine kinase lck. GA treatment depletes lck levels in cultured T cells by a kinetically slow dose-dependent process. Pulse-chase analyses indicate that GA induces the very rapid degradation of newly synthesized lck molecules. GA also induces a slower degradation of mature lck populations. These results correlate with global losses in protein tyrosine kinase activity and an inability to respond to TCR stimuli, but the activity of mature lck is not immediately compromised. Although the specific proteasome inhibitor lactacystin provides marginal protection against GA-induced lck depletion, proteasome inhibition also induces changes in lck detergent solubility independent of GA application. There is no other evidence for the involvement of the proteosome. Lysosome inhibition provides quantitatively superior protection against degradation. These results indicate that pharmacologic inhibition of hsp90 chaperone function may represent a novel immunosuppressant strategy, and elaborate on the appropriate context in which to interpret losses of lck as a reporter for the pharmacology of GA in whole organisms. PMID- 10706678 TI - Identification of a precursor to phosphatidyl choline-specific B-1 cells suggesting that B-1 cells differentiate from splenic conventional B cells in vivo: cyclosporin A blocks differentiation to B-1. AB - The origin of B-1 cells is controversial. The initial paradigm posited that B-1 and B-2 cells derive from separate lineages. More recently it has been argued that B-1 cells derive from conventional B cells as a result of T-independent Ag activation. To understand B-1 cell differentiation, we have generated Ig transgenic (Tg) mice using the H and L chain genes (VH12 and Vkappa4) of anti phosphatidyl choline (anti-PtC) B cells. In normal mice anti-PtC B cells segregate to B-1. Segregation is intact in VH12 (6-1) and VH12/Vkappa4 (double) Tg mice that develop large numbers of PtC-specific B cells. However, if B-1 cell differentiation is blocked, anti-PtC B cells in these Tg mice are B-2-like in phenotype, suggesting the existence of an Ag-driven differentiative pathway from B-2 to B-1. In this study, we show that double Tg mice have a population of anti PtC B cells that have the phenotypic characteristics of both B-2 and B-1 cells and that have the potential to differentiate to B-1 (B-1a and B-1b). Cyclosporin A blocks this differentiation and induces a more B-2-like phenotype in these cells. These findings indicate that these cells are intermediate between B-2 and B-1, further evidence of a B-2 to B-1 differentiative pathway. PMID- 10706679 TI - Inhibition of autoimmune diabetes by Fas ligand: the paradox is solved. AB - Previous reports that diabetogenic lymphocytes did not induce diabetes in nonobese diabetic (NOD)-lpr mice suggested the critical role of Fas-Fas ligand (FasL) interaction in pancreatic beta cell apoptosis. However, recent works demonstrated that FasL is not an effector molecule in islet beta cell death. We addressed why diabetes cannot be transferred to NOD-lpr mice despite the nonessential role of Fas in beta cell apoptosis. Lymphocytes from NOD-lpr mice were constitutively expressing FasL. A decrease in the number of FasL+ lymphocytes by neonatal thymectomy facilitated the development of insulitis. Cotransfer of FasL+ lymphocytes from NOD-lpr mice completely abrogated diabetes after adoptive transfer of lymphocytes from diabetic NOD mice. The inhibition of diabetes by cotransferred lymphocytes was reversed by anti-FasL Ab, indicating that FasL on abnormal lymphocytes from NOD-lpr mice was responsible for the inhibition of diabetes transfer. Pretreatment of lymphocytes with soluble FasL (sFasL) also inhibited diabetes transfer. sFasL treatment decreased the number of CD4+CD45RBlow cells and increased the number of propidium iodide-stained cells among CD4+CD45RBlow cells, suggesting that sFasL induces apoptosis on CD4+CD45RBlow "memory" cells. These results resolve the paradox between previous findings and suggest a new role for FasL in the treatment of autoimmune disorders. Our data also suggest that sFasL is involved in the deletion of potentially hazardous peripheral "memory" cells, contrary to previous reports that Fas on unmanipulated peripheral lymphocytes is nonfunctional. PMID- 10706680 TI - Induction of rapid T cell activation, division, and recirculation by intratracheal injection of dendritic cells in a TCR transgenic model. AB - Dendritic cells (DCs) are thought to be responsible for sensitization to inhaled Ag and induction of adaptive immunity in the lung. The characteristics of T cell activation in the lung were studied after transfer of Ag-pulsed bone marrow derived DCs into the airways of naive mice. Cell division of Ag-specific T cells in vivo was followed in a carboxyfluorescein diacetate succinimidyl ester-labeled cohort of naive moth cytochrome c-reactive TCR transgenic T cells. Our adoptive transfer system was such that transferred DCs were the only cells expressing the MHC molecule required for presentation of cytochrome c to transgenic T cells. Ag specific T cell activation and proliferation occurred rapidly in the draining lymph nodes of the lung, but not in nondraining lymph nodes or spleen. No bystander activation of non-Ag-specific T cells was induced. Division of Ag specific T cells was accompanied by transient expression of CD69, while up regulation of CD44 increased with each cell division. Divided cells had recirculated to nondraining lymph nodes and spleen by day 4 of the response. In vitro restimulation with specific Ag revealed that T cells were primed to proliferate more strongly and to produce higher amounts of cytokines per cell. These data are consistent with the notion that DCs in the lung are extremely efficient in selecting Ag-reactive T cells from a diverse repertoire. The response is initially localized in the mediastinal lymph nodes, but subsequently spreads systemically. This system should allow us to study the early events leading to sensitization to inhaled Ag. PMID- 10706681 TI - Central role of thrombospondin-1 in the activation and clonal expansion of inflammatory T cells. AB - Thrombospondin-1 (TSP) is a transiently expressed matricellular protein known to promote chemotaxis of leukocytes to inflammatory sites. However, TSP and its receptor CD36 are abundantly expressed in chronically inflamed tissues such as the rheumatoid synovium. Here, we show that TSP provides the costimulatory signal that is necessary for the activation of autoreactive T cells. Data presented reveal that TSP-mediated costimulation is achieved through its independent interaction with CD36 on APCs and with CD47 on T cells. We propose that a CD47 TSP-CD36 trimolecular complex is a novel costimulatory pathway that significantly decreases the threshold of T cell activation. Consistent with the paradigm that lesions in rheumatoid synovitis are sites of antigenic recognition, the characteristic focal expression of TSP on APCs such as macrophages and fibroblast like synoviocytes suggest a central role of TSP in the expansion of tissue infiltrating T cells. PMID- 10706682 TI - CD28, Ox-40, LFA-1, and CD4 modulation of Th1/Th2 differentiation is directly dependent on the dose of antigen. AB - The involvement of specific accessory/costimulatory molecules in differentiation to Th1 and Th2 phenotypes is controversial. Reports suggest that molecules such as CD4, CD28, and Ox-40 support Th2 differentiation and suppress Th1 differentiation, whereas others such as LFA-1 support Th1 responses and suppress Th2 responses. We have previously defined an in vitro model of differentiation that is absolutely dependent on the initial dose and affinity of peptide presented to a naive CD4 cell. The dose and affinity of Ag regulate autocrine production of IL-2, IL-4, and IFN-gamma, which in turn govern differentiation to Th1 and Th2 phenotypes. We have used this system to confirm that CD4, CD28, and Ox-40 interactions can promote, and LFA-1 interactions can suppress, differentiation of cells secreting the Th2 cytokines IL-5 and IL-13. However, for CD4 and LFA-1, this is only seen over a certain range of peptide doses. In addition, CD28 and Ox-40 interactions also promote Th1 differentiation. In general, agonist Abs to accessory molecules shifted the response curves for IFN gamma, IL-5, and IL-13 to lower doses, whereas antagonist reagents resulted in similar curves shifted toward the higher doses. We conclude that ligation of cell surface accessory receptors enables low doses of Ag to promote responses normally induced only by higher doses. Individual receptors do not intrinsically regulate one cytokine phenotype or another, suggesting that differentiation is controlled by the level of expression of multiple accessory molecule pairs integrated with the number and affinity of peptide/MHC complexes. PMID- 10706683 TI - Importance of histamine in the cytokine network in the lung through H2 and H3 receptors: stimulation of IL-10 production. AB - Histamine, a well-known inflammatory mediator, has been implicated in various immunoregulatory effects that are poorly understood. Thus, we tested the hypothesis that histamine inhibits the release of a proinflammatory cytokine, namely TNF, by stimulating the release of an anti-inflammatory cytokine, IL-10. Alveolar macrophages (AMs) from humans, Sprague Dawley rats, and the AM cell line, NR8383, were treated with different concentrations of histamine (10-5-10-7 M) for 2 h prior to their stimulation with suboptimal concentration of LPS (1 ng/ml) for 4 h. Histamine inhibited TNF release in a dose-dependent manner. This inhibition was mimicked by H2 and H3 receptor agonists, but not by H1 receptor agonist. Furthermore, we demonstrated the expression of H3 receptor mRNA in human AMs. Interestingly, treatment of AMs with anti-IL-10, anti-PGE2, or a NO synthase inhibitor (Nomega-nitro-l -arginine methyl ester) before the addition of histamine abrogated the inhibitory effect of the latter on TNF release. Histamine treatment (10-5 M) increased the release of IL-10 from unstimulated (2.2-fold) and LPS-stimulated (1. 7-fold) AMs. Unstimulated AMs, NR8383, express few copies of IL-10 mRNA, as tested by quantitative PCR, but expression of IL-10 was increased by 1.5-fold with histamine treatment. Moreover, the stimulation of IL 10 release by histamine was abrogated by pretreatment with anti-PGE2 or the NO synthase inhibitor, Nomega-nitro-l -arginine methyl ester. Thus, histamine increases the synthesis and release of IL-10 from AMs through PGE2 and NO production. These results suggest that histamine may play an important role in the modulation of the cytokine network. PMID- 10706684 TI - Development and analysis of various clonal alloantigen-dependent cytotoxic cell lines from channel catfish. AB - To determine the phenotypes of cytotoxic cells in channel catfish, clonal alloantigen-dependent leukocyte lines were established from mixed leukocyte cultures. Each clone was analyzed for expression of TCR alpha and beta genes by RT-PCR and for target cell specificity by 51Cr-release assay. Based on the above criteria, the following five different cell types were identified among the 19 clones analyzed: 1) TCR alphabeta+ allospecific cytotoxic cells, 2) TCR alphabeta+ nonspecific cytotoxic cells, 3) allospecific TCR alphabeta+ noncytotoxic cells, 4) TCR alphabeta- nonspecific cytotoxic cells, and 5) TCR alphabeta- allospecific cytotoxic cells. The demonstration of cloned, TCR alphabeta+, allospecific cytotoxic effectors provides the strongest evidence to date for the existence of cytotoxic T cells in fish. PMID- 10706685 TI - CD4 and CD8 expression by dendritic cell subtypes in mouse thymus and spleen. AB - The dendritic cells (DC) of mouse spleen and thymus were examined for expression of CD4 and CD8. Provided care was taken to avoid selective extraction or selective depletion of DC subpopulations, three main types of DC were detected in mouse spleen: a major new population of CD4+8- DEC-205low CD11bhigh DC, together with the previously described CD4-8- DEC-205low CD11bhigh DC and CD4-8alphaalpha+ DEC-205high CD11blow DC. The CD4 on the surface of the CD4+ splenic DC subpopulation was produced by the DC themselves, and CD4 RNA transcripts were present. Likewise, the CD8alpha on the surface of the splenic CD8+ DC was shown to be a product of the DC themselves, in agreement with earlier evidence. All three spleen DC types would be considered as mature, based on expression of CD80, CD86, and CD40 as well as on T cell stimulating function. Mouse thymuses appeared to contain two DC types; both were DEC-205highCD11blow, but they differed in the level of CD8alphaalpha expression. However, as well as this authenticated marker expression, immunofluorescent staining was also found to reflect a series of artifacts, due to the autofluorescence of contaminating cells and due to pickup of CD4 and CD8alphabeta. By constructing mice chimeric for the hemopoietic lineages using mixtures of wild-type bone marrow with CD4null or CD8alphanull bone marrow, a marked pickup by thymic DC of Ags derived from thymocytes was demonstrated. PMID- 10706686 TI - Induction of T cell anergy in the absence of CTLA-4/B7 interaction. AB - Immunologic tolerance in T lymphocytes is maintained through both thymic and peripheral contributions. One peripheral tolerance mechanism is the induction of T cell anergy, a form of nonresponsiveness resulting from incomplete T cell activation, such as stimulation through the TCR in the absence of costimulation. Recent reports have suggested that engagement of the inhibitory receptor CTLA-4 by its B7 ligand is critical for the initiation of anergy. We tested the importance of CTLA-4 in anergy induction in primary T cells with an in vitro anergy system. Using both CTLA-4/B7-blocking agents and CTLA-4-deficient T cells, we found that T cell anergy can be established in the absence of CTLA-4 expression and/or function. Even in the absence of CTLA-4 signal transduction, T cells activated solely through TCR ligation lose the ability to proliferate as a result of autocrine IL-2 production upon subsequent receptor engagement. Thus, CTLA-4 signaling is not required for the development of T cell anergy. PMID- 10706687 TI - Staphylococcal enterotoxin B stimulates expansion of autoreactive T cells that induce apoptosis in intestinal epithelial cells: regulation of autoreactive responses by IL-10. AB - T cell responses to self Ags and normal microbial flora are carefully regulated to prevent autoreactivity. Because IL-10-deficient mice develop colitis, and this response is triggered by luminal flora, we investigated whether IL-10 regulates the ability of microbial Ags to induce autoreactive T cells that could contribute to intestinal inflammation. T cells from wild-type mice were primed with staphylococcal enterotoxin B (SEB) in vitro, which induced an autoreactive proliferative response to syngeneic feeder cells. The cells were predominately CD3+ and CD4+. T cells from IL-10-deficient mice were constitutively autoreactive, and SEB priming enhanced this further. The autoreactive, proliferative response of T cells from wild-type mice was suppressed by IL-10 in the primary or secondary culture, and this effect was inhibited by neutralizing Abs to the IL-10R. To confirm that an autoreactive repertoire was expanded after SEB priming, we used CBA/J mice (Mls-1a) in which autoreactive T cells recognizing the endogenous viral superantigen are depleted (Vbeta6, 7, 8.1 TCR bearing cells). However, SEB rescued these autoreactive T cell repertoires. Adding anti-MHC class II Ab blocked the autoreactive response. SEB-primed splenic or colonic T cells also induced apoptosis in syngeneic intestinal epithelial cells that was blocked significantly by IL-10. Thus, microbial Ags have the potential to abrogate self tolerance by stimulating autoreactive T cells that become cytolytic to target cells. IL-10 plays a protective role in maintaining self tolerance after microbial stimulation by preventing the activation of T cells that contribute to epithelial cell damage. PMID- 10706688 TI - Role of activator protein-1 in TCR-mediated regulation of the murine fasl promoter. AB - The present study demonstrates that transcription factor interactions are important in regulating the murine fasl promoter following TCR-mediated activation. We used DNase I-footprinting, EMSAs, and transient transfection assays to identify the minimal TCR signal-responsive region within the fasl promoter. This region contains the previously identified binding sites for NF kappaB and Egr and the AP-1 site identified in this study. We found that TCR signaling induces AP-1 binding to this site and regulates the fasl promoter function in a fashion dependent on NF-kappaB binding. However, mutation in the AP 1 site alone did not show a significant effect on the promoter function. The data suggest that the minimal promoter required at least two transcription factors to function. PMID- 10706689 TI - Selective suppression of IL-12 production by chemoattractants. AB - We investigated the ability of chemoattractants to affect IL-12 production by human monocytes and dendritic cells. We found that pretreatment of monocytes with macrophage chemoattractant proteins (MCP-1 to -4), or C5a, but not stromal derived factor-1, macrophage inflammatory protein-1alpha, RANTES, or eotaxin, inhibited IL-12 p70 production in response to stimulation with Staphylococcus aureus, Cowan strain 1 (SAC), and IFN-gamma. The production of TNF-alpha and IL 10, however, was minimally affected by any of the chemoattractants. The degree of inhibition of IL-12 p70 production by MCP-1 to -4 was donor dependent and was affected by the autocrine inhibitory effects of IL-10. In contrast, C5a profoundly suppressed IL-12 production in an IL-10-independent fashion. Neither TGF-beta1 nor PGE2 was important for the suppression of IL-12 by any of the chemoattractants tested. The accumulation of mRNA for both IL-12 p35 and p40 genes was inhibited by chemokine pretreatment. Interestingly, MCP-1 to -4 and C5a did not suppress IL-12 production by monocyte-derived dendritic cells (DC) stimulated with CD40 ligand and IFN-gamma or by SAC and IFN-gamma, suggesting that these factors may act at the site of inflammation to suppress IL-12 and IFN gamma production rather than in the lymph node to affect T cell priming. Despite the inability of C5a to inhibit IL-12 production by DCs, the receptor for C5a (CD88) was expressed by these cells, and recombinant C5a induced a Ca2+ flux. Taken together, these results define a range of chemoattractant molecules with the ability to suppress IL-12 production by human monocytes and have broad implications for the regulation of immune responses in vivo. PMID- 10706690 TI - Role of mitogen-activated protein kinase cascades in mediating lipopolysaccharide stimulated induction of cyclooxygenase-2 and IL-1 beta in RAW264 macrophages. AB - LPS stimulation of RAW264 macrophages triggered the activation of mitogen- and stress-activated protein kinases-1 and -2 (MSK1, MSK2) and their putative substrates, the transcription factors cyclic AMP response element-binding protein (CREB) and activating transcription factor-1 (ATF1). The activation of MSK1/MSK2 was prevented by preincubating the cells with a combination of two drugs that suppress activation of the classical mitogen-activated protein kinase cascade and stress-activated protein kinase/p38, respectively, but inhibition was only partial in the presence of either inhibitor. The LPS-stimulated activation of CREB and ATF1, the transcription of the cyclooxygenase-2 (COX-2) and IL-1 beta genes (the promoters of which contain a cyclic AMP response element), and the induction of the COX-2 protein were prevented by the same drug combination, as well as by Ro 318220 or H89, potent inhibitors of MSK1/MSK2. Two other transcription factors, C/EBP beta and NF-kappa B, have been implicated in the transcription of the COX-2 gene. However, PD 98059 and/or SB 203580 did not prevent the LPS-induced increase in the level of the transcription factor C/EBP beta, and none of the four inhibitors used in this study prevented the activation of NF-kappa B. Our results demonstrate that two different mitogen-activated protein kinase cascades are rate limiting for the LPS-induced activation of CREB/ATF1 and the transcription of the COX-2 and IL-1 beta genes. They also suggest that MSK1 and MSK2 may play a role in these processes and hence are potential targets for the development of novel antiinflammatory drugs. PMID- 10706691 TI - Dual phase priming by IL-3 for leukotriene C4 generation in human basophils: difference in characteristics between acute and late priming effects. AB - Previous studies have suggested that enhancement of mediator release from human basophils by IL-3 occurs in at least two phases, and the current studies further characterize the signaling changes that accompany these two phases of the basophil in response to IL-3. The test stimulus for these studies was anaphylatoxin split product of C component (C5a), which does not induce leukotriene C4 release without prior IL-3 treatment. Functionally, IL-3 priming occurs after 5 min, disappears by 2 h, and returns by 18 h. In contrast, the kinetics of cytosolic phospholipase A2 (cPLA2) and extracellular signal-regulated kinase (ERK1/2) phosphorylation, induced by IL-3, do not show the second rise by 18 h. The kinetics of cPLA2 and ERK1/2 phosphorylation following stimulation with C5a are the same for cells that were not treated with IL-3 as for those treated for 18 h, i.e., a lag in phosphorylation of cPLA2 and ERK1/2 lasting 30 s before its eventual rise. Previous studies showed that a 5-min treatment with IL-3 induced little change in the C5a-induced cytosolic calcium response, while 24 h of treatment resulted in a marked and sustained cytosolic calcium elevation during the C5a-induced response. The first phase of the IL-3 priming effect (5-15 min of treatment) was unaffected by cycloheximide, while the second phase (18 h) was inhibited. These data suggest that early IL-3 priming results from preconditioning cPLA2, i.e., causing its phosphorylation, while late priming results from a qualitative change in the cytosolic calcium response. PMID- 10706692 TI - In vivo survival of autoreactive B cells: characterization of long-lived B cells. AB - To determine the effects of chronic Ag stimulation on B cell survival and phenotype, we compared survival and surface markers of hen egg lysozyme (HEL) specific B cells in Ig transgenic (Tgn) mice, which lack HEL, and in HEL-Ig transgenic mice, which express soluble HEL. Serum HEL levels were maximized in HEL-Ig Tgn mice by feeding them zinc, which activates the metallothionein promoter that regulates HEL expression. B cell age was characterized by expression of heat-stable Ag, and B220 and B cell survival was studied by evaluating changes in B cell number when lymphopoiesis was suppressed with anti IL-7 mAb and by identifying newly generated B cells through 5-bromo-2' deoxyuridine incorporation. Our observations show that the mean B cell life span is considerably reduced in HEL-Ig Tgn compared with Ig Tgn mice, but also demonstrate that some HEL-Ig Tgn B cells survive to maturity. Some of these surviving B cells have undergone receptor editing (substitution of an endogenous Ig light chain for the transgenic Ig light chain), so that their ability to bind HEL is decreased or absent. Surviving HEL-Ig Tgn B cells that retain HEL specificity express decreased mIgD and little or no mIgM. mIgD expression progressively decreases with increasing HEL-Ig Tgn B cell age. These observations suggest that self Ag-specific B cells can survive in the presence of soluble self Ag by down-regulating mIg expression, which should limit B cell signaling by Ag that might otherwise cause deletion of these cells. PMID- 10706693 TI - Single cell analysis reveals that IL-4 receptor/Stat6 signaling is not required for the in vivo or in vitro development of CD4+ lymphocytes with a Th2 cytokine profile. AB - The concept that IL-4 is the primary signal for Th2 lymphocyte differentiation has recently been put in doubt by studies in which the production of Th2 associated cytokines was detected in mice deficient in IL-4 synthesis or IL-4R triggering. In this study, we formally demonstrate by single cell analysis that CD4+ lymphocytes with a classical Th2 phenotype (IL-4+, IL-5+, IFN-gamma-, IL-2-) develop in significant numbers in helminth-infected mice deficient in either IL 4R alpha-chain or Stat6. While an expanded population of Th1 (IL-4-, IL-5-, IFN gamma+, IL-2+) lymphocytes was observed in the same animals, surprisingly, cells with a mixed Th0 cytokine pattern were rare. The cytokine production phenotypes of the Th1 and Th2 subpopulations generated in infected Stat6-deficient mice were unaffected by in vitro neutralization of endogenous IL-4 or IFN-gamma. Nevertheless, while addition of exogenous rIL-12 resulted in transitory IFN-gamma production by Th2 lymphocytes from both wild-type and Stat6-deficient mice, IL-4 synthesis was preserved in the former, but temporarily ablated in the latter cells. Importantly, IL-4+ IFN-gamma- and IL-4- IFN-gamma+ populations similar to those arising in helminth-infected Stat6-deficient mice could also be generated in vitro by repetitive polyclonal stimulation of CD4+CD62Lhigh lymphocytes from uninfected mice of the same strain. Together, the results of these single cell analysis experiments demonstrate that IL-4R/Stat6 signaling, while influencing the final frequency of Th2 lymphocytes, is not essential for Th2 cell development, and suggest that this pathway has a previously unrecognized function in stabilizing Th2 populations once they have emerged. PMID- 10706694 TI - Dominance of IL-12 over IL-4 in gamma delta T cell differentiation leads to default production of IFN-gamma: failure to down-regulate IL-12 receptor beta 2 chain expression. AB - Gamma delta T cells secrete Th1- and Th2-like cytokines that help mediate innate and acquired immunity. We have addressed the mechanism whereby murine gamma delta T cells acquire the capacity to differentially produce such cytokines. Splenic gamma delta T cells could be polarized into IFN-gamma- or IL-4-secreting cells in vitro; however, in contrast to CD4+ alpha beta T cells, gamma delta T cells predominantly produced IFN-gamma, even in the presence of IL-4, a finding independent of genetic background. Like CD4+ Th1 cells, IFN-gamma-producing cells expressed the IL-12 receptor beta 2-chain after activation in the presence of IL 12; however, unlike Th2 cells, IL-4-primed gamma delta T cells also expressed this receptor, even in the absence of IFN-gamma and despite the presence of the transcription factor GATA-3. IL-12 also induced IL-4-primed gamma delta T cells to proliferate and to translocate Stat3/Stat4, indicating signaling through the IL-12 receptor. These molecular events can account for the predominant production of IFN-gamma by gamma delta T cells in the presence of IL-12, despite the availability of IL-4. Early and predominant production of IFN-gamma by gamma delta T cells likely is critical for the roles that these cells play in protection against intracellular pathogens and in tumor immunity. PMID- 10706695 TI - Vigorous allograft rejection in the absence of danger. AB - Tolerance to self is a necessary attribute of the immune system. It is thought that most autoreactive T cells are deleted in the thymus during the process of negative selection. However, peripheral tolerance mechanisms also exist to prevent development of autoimmune diseases against peripheral self-Ags. It has been proposed that T cells develop tolerance to peripheral self-Ags encountered in the absence of inflammation or "danger" signals. We have used immunodeficient Rag 1-/- mice to study the response of T cells to neo-self peripheral Ags in the form of well-healed skin and vascularized cardiac allografts. In this paper we report that skin and cardiac allografts without evidence of inflammation are vigorously rejected by transferred T cells or when recipients are reconstituted with T cells at a physiologic rate by nude bone graft transplantation. These results provide new insights into the role of inflammation or "danger" in the initiation of T cell-dependent immune responses. These findings also have profound implications in organ transplantation and suggest that in the absence of central deletional tolerance, peripheral tolerance mechanisms are not sufficient to inhibit alloimmune responses even in the absence of inflammation or danger. PMID- 10706696 TI - Induction of diabetes in nonobese diabetic mice by Th2 T cell clones from a TCR transgenic mouse. AB - We have produced a panel of cloned T cell lines from the BDC-2.5 TCR transgenic (Tg) mouse that exhibit a Th2 cytokine phenotype in vitro but are highly diabetogenic in vivo. Unlike an earlier report in which T cells obtained from the Tg mouse were cultured for 1 wk under Th2-promoting conditions and were found to induce disease only in NOD.scid recipients, we found that long-term T cell clones with a fixed Th2 cytokine profile can transfer disease only to young nonobese diabetic (NOD) mice and never to NOD.scid recipients. Furthermore, the mechanism by which diabetes is transferred by a Tg Th2 T cell clone differs from that of the original CD4+ Th1 BDC-2.5 T cell clone made in this laboratory. Whereas the BDC-2.5 clone rapidly causes disease in NOD.scid recipients less than 2 wk old, the Tg Th2 T cell clones can do so only when cotransferred with other diabetogenic T cells, suggesting that the Th2 T cell requires the presence of host T cells for initiation of disease. PMID- 10706697 TI - Thymic transplantation in miniature swine. II. Induction of tolerance by transplantation of composite thymokidneys to thymectomized recipients. AB - Previous studies in our laboratory have demonstrated that the presence of the thymus is essential for rapid and stable tolerance induction in allotransplant models. We now report an attempt to induce tolerance to kidney allografts by transplanting donor thymic grafts simultaneously with the kidney in thymectomized recipients. Recipients were thymectomized 3 wk before receiving an organ and/or tissues from a class I-mismatched donor. Recipients received 1) a kidney allograft alone, 2) a composite allogeneic thymokidney (kidney with vascularized autologous thymic tissue under its capsule), or 3) separate kidney and thymic grafts from the same donor. All recipients received a 12-day course of cyclosporine. Thymectomized animals receiving a kidney allograft alone or receiving separate thymic and kidney grafts had unstable renal function due to severe rejection with the persistence of anti-donor cytotoxic T cell reactivity. In contrast, recipients of composite thymokidney grafts had stable renal function with no evidence of rejection histologically and donor-specific unresponsiveness. By postoperative day 14, the thymic tissue in the thymokidney contained recipient type dendritic cells. By postoperative day 60, recipient-type class I positive thymocytes appeared in the thymic medulla, indicating thymopoiesis. T cells were both recipient and donor MHC-restricted. These data demonstrate that the presence of vascularized-donor thymic tissue induces rapid and stable tolerance to class I disparate kidney allografts in thymectomized recipients. To our knowledge, this is the first evidence of functional vascularized thymic grafts permitting transplantation tolerance to be induced in a large animal model. PMID- 10706698 TI - Mature CD4+ T cells perceive a positively selecting class II MHC/peptide complex in the periphery. AB - A repertoire of TCRs is selected in the thymus by interactions with MHC bound to self-derived peptides. Whether self peptides bound to MHC influence the survival of mature T cells in the periphery remains enigmatic. In this study, we show that the number of naive CD4+ T cells that developed in mice with class II MHC bound with endogenous peptides (Abwt) diminished when transferred into mice with Ab covalently bound with a single peptide (AbEp). Moreover, transfer of a mixture of naive CD4+ T cells derived from Abwt and from AbEp mice into AbEp mice resulted in the expansion of the latter and decline of the former. In contrast, when wild type activated CD4+ T cells were transferred into AbEp or Abwt mice, these cells survived in both recipients for more than 4 wk, but further expanded in the Abwt host. We conclude that to survive, naive CD4+ T cells favor peripheral expression of the class II MHC/peptide complex(es) involved in their thymic selection, whereas some of activated CD4+ T cells may require them only for expansion. PMID- 10706699 TI - CD8+ T cell-dependent elimination of dendritic cells in vivo limits the induction of antitumor immunity. AB - The fate of dendritic cells (DC) after they have initiated a T cell immune response is still undefined. We have monitored the migration of DC labeled with a fluorescent tracer and injected s.c. into naive mice or into mice with an ongoing immune response. DC not loaded with Ag were detected in the draining lymph node in excess of 7 days after injection with maximum numbers detectable approximately 40 h after transfer. In contrast, DC that had been loaded with an MHC class I binding peptide disappeared from the lymph node with kinetics that parallel the known kinetics of activation of CD8+ T cells to effector function. In the presence of high numbers of specific CTL precursors, as in TCR transgenic mice, DC numbers were significantly decreased by 72 h after injection. The rate of DC disappearance was extremely rapid and efficient in recently immunized mice and was slower in "memory" mice in which memory CD8+ cells needed to reacquire effector function before mediating DC elimination. We also show that CTL-mediated clearance of Ag-loaded DC has a notable effect on immune responses in vivo. Ag specific CD8+ T cells failed to divide in response to Ag presented on a DC if the DC were targets of a pre-existing CTL response. The induction of antitumor immunity by tumor Ag-loaded DC was also impaired. Therefore, CTL-mediated clearance of Ag-loaded DC may serve as a negative feedback mechanism to limit the activity of DC within the lymph node. PMID- 10706700 TI - The timing of GM-CSF expression plasmid administration influences the Th1/Th2 response induced by an HIV-1-specific DNA vaccine. AB - The mechanism of immune activation induced by a plasmid-encoding GM-CSF (pGM CSF), administered in combination with a DNA vaccine encoding the envelope of HIV, was studied. Injecting pGM-CSF i.m. into mice 3 days before DNA vaccination primarily induced a Th2 response. Simultaneous administration of the DNA vaccine plus pGM-CSF activated both a Th1 and a Th2 response. When the plasmid was injected 3 days after DNA vaccination, enhancement of Th1 immunity predominated. These results suggest that the timing of cytokine expression determines the phenotype of the resultant Th response. After 3 days of pGM-CSF injection, the increased percentages of CD11c+, CD8+ cells were observed in the regional lymph nodes. In addition, many infiltrated cells, including S-100 protein-positive cells, were found in the pGM-CSF-injected tissue. The importance of these S-100+ cells or both CD8+ and CD11c+ cells, especially that of dendritic cells (DCs), was also studied. DCs derived from bone marrow and cultured in RPMI 1640 medium containing IL-4 and GM-CSF were incubated with DNA vaccine and then transferred into naive mice. Mice receiving DCs showed strong HIV-1-specific Th2 immune responses. Our results suggest that DCs play important roles in the activation or modification of the Th2-type immune response induced by DNA vaccination. PMID- 10706701 TI - Intratumoral coinjection of two adenoviruses, one encoding the chemokine IFN gamma-inducible protein-10 and another encoding IL-12, results in marked antitumoral synergy. AB - We have constructed a recombinant defective adenovirus that expresses functional murine IFN-gamma-inducible protein-10 (IP-10) chemokine (AdCMVIP-10). Injection of AdCMVIP-10 into s.c. tumor nodules derived from the CT26 murine colorectal adenocarcinoma cell line displayed some antitumor activity but it was not curative in most cases. Previous studies have shown that injection of similar s. c. CT26 tumor nodules with adenovirus-encoding IL-12 (AdCMVIL-12) induces tumor regression in nearly 70% of cases in association with generation of antitumor CTL activity. AdCMVIP-10 synergizes with the antitumor effect of suboptimal doses of AdCMVIL-12, reaching 100% of tumor eradication not only against injected, but also against distant noninjected tumor nodules. Colocalization of both adenoviruses at the same tumor nodule was required for the local and distant therapeutic effects. Importantly, intratumoral gene transfer with IL-12 and IP-10 generated a powerful tumor-specific CTL response in a synergistic fashion, while both CD4 and CD8 T cells appeared in the infiltrate of regressing tumors. Moreover, the antitumor activity of IP-10 plus IL-12 combined gene therapy was greatly diminished by simultaneous in vivo depletion of CD4+ and CD8+ T cells but was largely unaffected by single depletion of each T cell subset. An important role for NK cells was also suggested by asialo GM1 depletion experiments. From a clinical point of view, the effects of IP-10 permit one to lower the required gene transfer level of IL-12, thus preventing dose-dependent IL-12-mediated toxicity while improving the therapeutic efficacy of the elicited antitumor response. PMID- 10706702 TI - Systematic analysis of the role of CD19 cytoplasmic tyrosines in enhancement of activation in Daudi human B cells: clustering of phospholipase C and Vav and of Grb2 and Sos with different CD19 tyrosines. AB - CD19 is a coreceptor on B cells that enhances the increase in cytoplasmic calcium and ERK2 activation when coligated with the B cell Ag receptor. Constructs containing point mutations and truncations were expressed in Daudi human B lymphoblastoid cells to systematically determine the requirement for individual CD19 cytoplasmic tyrosines in these responses. Evidence for activity was found for Y330, Y360, and Y421 as well as that previously published for Y391. Precipitates formed with phosphopeptides consisting of CD19 sequences flanking these residues were used to screen for cytoplasmic proteins that mediate signaling. Phosphopeptide Y330 precipitated Grb2 and Sos, whereas phosphopeptides Y391 and Y421 both precipitated Vav and phospholipase C-gamma2. These molecules also were found associated with native CD19. In mapping studies with altered constructs, CD19 Y330 and/or Y360 were necessary for binding Grb2 and Sos. Vav associated with CD19 constitutively in unstimulated cells by a tyrosine independent mechanism requiring the portion of CD19 encoded by exons 9-12. After B cell Ag receptor stimulation, Vav association was tyrosine-dependent, but binding was influenced by multiple residues. However, when maximally phosphorylated by pervanadate, Y391 and, to a lesser extent, Y421 were sufficient. CD19 Y391 was also both necessary and sufficient for binding phospholipase C-gamma2. Thus, different tyrosines along the CD19 cytoplasmic domain provide scaffolding for the formation of complexes of different signaling molecules. PMID- 10706703 TI - Description of an ectothermic TCR coreceptor, CD8 alpha, in rainbow trout. AB - We have cloned the first CD8 alpha gene from an ectothermic source using a degenerate primer for Ig superfamily V domains. Similar to homologues in higher vertebrates, the rainbow trout CD8 alpha gene encodes a 204-aa mature protein composed of two extracellular domains including an Ig superfamily V domain and hinge region. Differing from mammalian CD8 alpha V domains, lower vertebrate (trout and chicken) sequences do not contain the extra cysteine residue (C strand) involved in the abnormal intrachain disulfide bridging within the CD8 alpha V domain of mice and rats. The trout membrane proximal hinge region contains the two essential cysteine residues involved in CD8 dimerization (alpha alpha or alpha beta) and threonine, serine, and proline residues which may be involved in multiple O-linked glycosylation events. Although the transmembrane region is well conserved in all CD8 alpha sequences analyzed to date, the putative trout cytoplasmic region differs and, in fact, lacks the consensus p56lck motif common to other CD8 alpha sequences. We then determined that the trout CD8 alpha genomic structure is similar to that of humans (six exons) but differs from that of mice (five exons). Additionally, Northern blotting and RT PCR demonstrate that trout CD8 alpha is expressed at high levels within the thymus and at weaker levels in the spleen, kidney, intestine, and peripheral blood leukocytes. Finally, we show that trout CD8 alpha can be expressed on the surface of cells via transfection. Together, our results demonstrate that the basic structure and expression of CD8 alpha has been maintained for more than 400 million years of evolution. PMID- 10706704 TI - NKT cells in the rat: organ-specific distribution of NK T cells expressing distinct V alpha 14 chains. AB - Rat invariant TCR alpha-chains and NKT cells were investigated to clarify whether CD1d-mediated recognition by NKT cells is conserved further in evolution. Rats had multiple-copies of TRAV14 genes, which can be categorized into two types according to the diversity accumulated in the CDR2 region. Rats retained invariant TCR alpha forms with the homogeneous junctional region similar to mouse invariant TRAV14-J281. The proportion of invariant TCR among V alpha 14+ clones was 12.9% in the thymus and increased in the periphery, 31% in the spleen and 95% in hepatic sinusoidal cells. The invariant TRAV14-J281 was expressed by liver sinusoidal and splenic NKT cells with CD8, CD44high, and TCR V beta 8. Type 1 invariant TCR alpha was expressed more frequently in hepatic lymphocytes, while type 2 invariant TCR alpha was expressed predominantly in the spleen. Both types of cells cytolyzed to and were stimulated to proliferate by CD1d-expressing cells in a CD1d-restricted manner. These results suggested that rat NKT cells bearing distinct V alpha 14 chains are distributed in a tissue-specific pattern. NKT cell populations in rats were more variable than those in mice, indicating that they play novel roles in nature. The implication of the molecular interaction between the structurally diverse invariant TCR alpha and CD1d/ligand complex in different organs is discussed. PMID- 10706705 TI - Salmonella-type heptaacylated lipid A is inactive and acts as an antagonist of lipopolysaccharide action on human line cells. AB - The stimulation of both THP-1 and U937 human-derived cells by Salmonella lipid A preparations from various strains, as assessed by TNF-alpha induction and NF kappaB activation, was found to be very low (almost inactive) compared with Escherichia coli lipid A, but all of the lipid As exerted strong activity on mouse cells and on Limulus gelation activity. Experiments using chemically synthesized E. coli-type hexaacylated lipid A (506) and Salmonella-type heptaacylated lipid A (516) yielded clearer results. Both lipid A preparations strongly induced TNF-alpha release and activated NF-kappaB in mouse peritoneal macrophages and mouse macrophage-like cell line J774-1 and induced Limulus gelation activity, although the activity of the latter was slightly weaker than that of the former. However, 516 was completely inactive on both THP-1 and U937 cells in terms of both induction of TNF-alpha and NF-kappaB activation, whereas 506 displayed strong activity on both cells, the same as natural E. coli LPS. In contrast to the action of the lipid A preparations, all the Salmonella LPSs also exhibited full activity on human cells. However, the polysaccharide portion of the LPS neither exhibited TNF-alpha induction activity on the cells when administered alone or together with lipid A nor inhibited the activity of the LPS. These results suggest that the mechanism of activation by LPS or the recognition of lipid A structure by human and mouse cells may differ. In addition, both 516 and lipid A from Salmonella were found to antagonize the 506 and E. coli LPS action that induced TNF-alpha release and NF-kappaB activation in THP-1 cells. PMID- 10706706 TI - The role of high-mobility group I(Y) proteins in expression of IL-2 and T cell proliferation. AB - The high-mobility group I(Y) (HMGI(Y)) family of proteins plays an important architectural role in chromatin and have been implicated in the control of inducible gene expression. We have previously shown that expression of HMGI antisense RNA in Jurkat T cells inhibits the activity of the IL-2 promoter. Here we have investigated the role of HMGI(Y) in controlling IL-2 promoter-reporter constructs as well as the endogenous IL-2 gene in both Jurkat T cells and human PBL. We found that the IL-2 promoter has numerous binding sites for HMGI(Y), which overlap or are adjacent to the known transcription factor binding sites. HMGI(Y) modulates binding to the IL-2 promoter of at least three transcription factor families, AP-1, NF-AT and NF-kappaB. By using a mutant HMGI that cannot bind to DNA but can still interact with the transcription factors, we found that DNA binding by HMGI was not essential for the promotion of transcription factor binding. However, the non-DNA binding mutant acts as a dominant negative protein in transfection assays, suggesting that the formation of functional HMGI(Y) containing complexes requires DNA binding as well as protein:protein interactions. The alteration of HMGI(Y) levels affects IL-2 promoter activity not only in Jurkat T cells but also in PBL. Importantly, we also show here that expression of the endogenous IL-2 gene as well as proliferation of PBL are affected by changes in HMGI(Y) levels. These results demonstrate a major role for HMGI(Y) in IL-2 expression and hence T cell proliferation. PMID- 10706707 TI - LST1: a gene with extensive alternative splicing and immunomodulatory function. AB - The gene of the leukocyte-specific transcript (LST1) is encoded within the TNF region of the human MHC. The LST1 gene is constitutively expressed in leukocytes and dendritic cells, and it is characterized by extensive alternative splicing. We identified 7 different LST1 splice variants in PBMC; thus, 14 LST1 splice variants (LST1/A-LST1/N) have been detected in various cell types. These isoforms code for transmembrane as well as soluble LST1 proteins characterized by two alternative open reading frames at their 3' end. We demonstrate the presence of the transmembrane variant LST1/C on the cell surface of the monocytic cell lines U937 and THP1. Recombinant expression of LST1/C permitted its profound inhibitory effect on lymphocyte proliferation to be observed. In contrast, the alternative transmembrane variant LST1/A, the extracellular domain of which shows no amino acid sequence homology to LST1/C exerted a weaker but similar inhibitory effect on PBMC. These data demonstrate the protein expression of LST1 on the cell surface of mononuclear cells, and they show an inhibitory effect on lymphocyte proliferation of two LST1 proteins although they have only a very short amino acid homology. PMID- 10706708 TI - HLA-DR restricted peptide candidates for bee venom immunotherapy. AB - T cell epitopes containing peptides have been recently proposed as an alternative to conventional immunotherapy of allergic diseases because they are expected to be better tolerated than allergen extracts. A principal limitation to their clinical use is that they present an important diversity, which primarily results from the polymorphism of HLA class II molecules. In Caucasian populations, however, seven alleles of the most expressed molecules (namely DRB1*0101, DRB1*0301, DRB1*0401, DRB1*0701, DRB1*1101, DRB1*1301, and DRB1*1501) predominate. Peptides from allergens that would efficiently bind to them should be potential candidates for specific immunotherapy. In this paper, we have determined the peptides present in the major bee venom allergen by investigating the capacity of synthetic peptides that encompass its whole sequence to bind to each allele. Several efficient binders have been identified and are either allele specific or common to several HLA-DR molecules. Interestingly enough, the 81-97 sequence is universal in the sense that it binds to all studied molecules. This sequence is surrounded by several active regions, which make the 76-106 sequence particularly rich of binding determinants and a good candidate for specific immunotherapy. Statistical analyses of the binding data also provide an overview of the preponderant HLA-DR alleles specificity. PMID- 10706709 TI - A putative 12 transmembrane domain cotransporter expressed in thymic cortical epithelial cells. AB - We have isolated a full-length cDNA clone (thymic stromal origin (TSO)-1C12) from a SCID thymus library using a probe from a PCR-based subtractive library enriched for sequences from fetal thymic stromal cells. TSO-1C12 mRNA is expressed mainly in the thymic cortex and is highly enriched in SCID thymus. Expression per cell is highest during fetal thymus development and decreases after day 16. Antipeptide Abs immunoprecipitated a hydrophobic, plasma membrane glycoprotein (thymic stromal cotransporter, TSCOT) whose translated sequence has weak homology to bacterial antiporters and mammalian cation cotransporters with 12 transmembrane domains. TSCOT represents a new member of this superfamily that is highly expressed in thymic cortical epithelial cells. PMID- 10706710 TI - Unprecedented polymorphism of Mhc-DRB region configurations in rhesus macaques. AB - The rhesus macaque is an important model in preclinical transplantation research and for the study of chronic and infectious diseases, and so extensive knowledge of its MHC (MhcMamu) is needed. Nucleotide sequencing of exon 2 allowed the detection of 68 Mamu-DRB alleles. Although most alleles belong to loci/lineages that have human equivalents, identical Mhc-DRB alleles are not shared between humans and rhesus macaques. The number of -DRB genes present per haplotype can vary from two to seven in the rhesus macaque, whereas it ranges from one to four in humans. Within a panel of 210 rhesus macaques, 24 Mamu-DRB region configurations can be distinguished differing in the number and composition of loci. None of the Mamu-DRB region configurations has been described for any other species, and only one of them displays major allelic variation giving rise to a total of 33 Mamu-DRB haplotypes. In the human population, only five HLA-DRB region configurations were defined, which in contrast to the rhesus macaque exhibit extensive allelic polymorphism. In comparison with humans, the unprecedented polymorphism of the Mamu-DRB region configurations may reflect an alternative strategy of this primate species to cope with pathogens. Because of the Mamu-DRB diversity, nonhuman primate colonies used for immunological research should be thoroughly typed to facilitate proper interpretation of results. This approach will minimize as well the number of animals necessary to conduct experiments. PMID- 10706711 TI - Tumor rejection and immune memory elicited by locally released LEC chemokine are associated with an impressive recruitment of APCs, lymphocytes, and granulocytes. AB - The human beta chemokine known as LEC (also called NCC-4, HCC-4, or LMC) displays chemotactic activity for monocytes and dendritic cells. The possibility that its local presence increases tumor immunogenicity is addressed in this paper. TSA parental cells (TSA-pc) are poorly immunogenic adenocarcinoma cells that grow progressively, kill both nu/nu and syngeneic BALB/c mice, and give rise to lung metastases. TSA cells engineered to release LEC (TSA-LEC) are still able to grow in nu/nu mice, but are promptly rejected and display a marginal metastatic phenotype in BALB/c mice. Rejection is associated with a marked T lymphocyte and granulocyte infiltration, along with extensive macrophage and dendritic cell recruitment. NK cells and CD4+ T lymphocytes are uninfluential in TSA-LEC cell rejection, whereas both CD8+ lymphocytes and polymorphonuclear leukocytes play a major role. An antitumor immune memory is established very quickly after rejection, since 6 days later 75% of BALB/c mice were already resistant to a TSA pc challenge. Spleen cells from rejecting mice display specific cytotoxic activity against TSA-pc and secrete IFN-gamma and IL-2 when restimulated by TSA pc. The ability of LEC to markedly improve recognition of poorly immunogenic cells by promoting APC-T cell cross-talk suggests that it could be an effective component of antitumor vaccines. PMID- 10706712 TI - TGF-beta 2 reduces demyelination, virus antigen expression, and macrophage recruitment in a viral model of multiple sclerosis. AB - TGF-beta 2 is a potent immunoregulatory mediator that influences B cell, T cell, and macrophage function. To test whether this cytokine alters pathology in a model of virus-induced demyelinating disease, we treated SJL/J mice with TGF-beta 2 either before or after infection with Theiler's murine encephalomyelitis virus. Treatment continued three times weekly through day 35 postinfection. TGF-beta 2 administration resulted in significantly smaller lesions and fewer virus Ag positive cells in the spinal cords of infected SJL/J mice. Mice treated with TGF beta 2 had similar levels of virus-specific IgG as infected, control-treated mice. TGF-beta 2 administration significantly increased the level of non-virus specific activated CTLs, but had no effect on virus-specific CTLs. TUNEL revealed a decrease in the number of apoptotic nuclei in the spinal cord white matter of mice treated in vivo with TGF-beta 2. Immunostaining with an Ab to F4/80 revealed that TGF-beta 2-treated mice had significantly fewer F4/80-positive cells in the white matter of the spinal cord as compared with infected control-treated mice. These data suggest that TGF-beta 2 may control virus-induced demyelination via an immunomodulatory mechanism that reduces macrophage infiltration. PMID- 10706713 TI - Blockade of CD14 increases Shigella-mediated invasion and tissue destruction. AB - Shigella is a diarrheal pathogen that causes disease through invasion of the large intestinal mucosa. The endotoxin of the invading bacterium may play a key role in the disease process by causing inflammation and tissue injury during infection. Earlier studies have shown that various animal species lacking functional CD14 were protected against endotoxin-mediated shock. Rabbits experimentally infected with Shigella were used to test the hypothesis that blockade of endotoxin-induced cell activation with anti-CD14 mAb would diminish inflammation and thus disease severity. Unexpectedly, we observed that the intestinal mucosa of anti-CD14-treated animals exhibited a 50-fold increase in bacterial invasion and more severe tissue injury compared with controls. Despite higher bacterial loads in treated animals, the numbers of polymorphonuclear leukocytes that were recruited to the infection site were similar to those in controls. Furthermore, the phagocytic cells of CD14-blocked animals produced IL-1 and TNF-alpha. Moreover, in vitro blockade of CD14 did not impede bactericidal activity. Thus, anti-CD14 treatment interfered with host defense mechanisms involved with removal/eradication of Shigella. PMID- 10706714 TI - p38 mitogen-activated protein kinase and c-jun-NH2-terminal kinase regulate RANTES production by influenza virus-infected human bronchial epithelial cells. AB - Airway epithelial cells which are the initial site of influenza virus (IV) infection are suggested to participate in airway inflammatory response by expressing various cytokines including RANTES; however, the intracellular signal that regulates RANTES expression has not been determined. In the present study, we examined the role of p38 mitogen-activated protein (MAP) kinase, extracellular signal-regulated kinase (Erk), and c-Jun-NH2-terminal kinase (JNK) in RANTES production by IV-infected human bronchial epithelial cells. The results showed that IV infection induced increases in p38 MAP kinase, and Erk and JNK phosphorylation and activity. SB 203580, PD 98059, and CEP-1347 attenuated IV infection induced p38 MAP kinase activity, Erk activity, and JNK activity, respectively. SB 203580 and CEP-1347 attenuated RANTES production by 45.3% and 45.2%, respectively, but a combination of these inhibitors additively attenuated by 69.1%. In contrast, PD 98059 did not attenuate. Anti-IL-1alpha mAb, anti-IL 1beta mAb, anti-TNF-alpha mAb, anti-IL-8 mAb, anti-IFN-beta mAb, anti-RANTES mAb, and a combination of these mAbs did not affect IV infection-induced increases in p38 MAP kinase, Erk, and JNK phosphorylation, indicating that each cytokine neutralized by corresponding Ab was not involved in IV infection-induced phosphorylation of MAP kinases. N-acetylcysteine (NAC) did not affect IV infection-induced increases in MAP kinase phosphorylation, whereas NAC attenuated RANTES production by 18.2%, indicating that reactive oxygen species may act as a second messenger leading to RANTES production via p38 MAP kinase- and JNK independent pathway. These results indicate that p38 MAP kinase and JNK, at least in part, regulate RANTES production by bronchial epithelial cells. PMID- 10706715 TI - Tumor immunity in perforin-deficient mice: a role for CD95 (Fas/APO-1). AB - CTL and NK cells use two distinct cytocidal pathways: 1) perforin and granzyme based and 2) CD95L/CD95 mediated. The former requires perforin expression by the effectors (CTL or NK), whereas the latter requires CD95 (Fas/APO-1) expression by the target. We have investigated how these two factors contribute to tumor immune surveillance by studying the immunity of perforin-deficient mice against the progressor C57BL/6 Lewis lung carcinoma 3LL, which expresses no CD95 when cultured in vitro. Unexpectedly, the results indicated that the perforin independent CD95L/CD95 pathway of CTL/NK plays a role in acting against D122 and Kb39.5 (39.5) high and low metastatic sublines, respectively, derived from the 3LL tumor. Although no membrane-bound CD95 was detected on cultured D122 and 39. 5 cells, surface CD95 expression on both D122 and 39.5 was considerably up regulated when the tumors were grown in vivo. A similarly enhanced expression of CD95 was observed with three additional tumors; LF-, BW, and P815, injected into syngeneic and allogeneic mice. The finding of up-regulated CD95 expression on tumor cells placed in vivo suggests that a CD95-based mechanism plays a role in tumor immunity at early stages of tumor growth. Consequently, the progressive down-regulation of CD95 expression during tumor progression may indeed be an escape mechanism as previously reported. Together, these results suggest a role for CD95-dependent, perforin-independent immunity against certain tumors. PMID- 10706716 TI - Superfibronectin, a multimeric form of fibronectin, increases HIV infection of primary CD4+ T lymphocytes. AB - The ability of viruses and bacteria to interact with the extracellular matrix plays an important role in their infectivity and pathogenicity. Fibronectin is a major component of the extracellular matrix in lymph node tissue, the main site of HIV deposition and replication during the chronic phase of infection. Therefore, we asked whether matrix fibronectin (FN) could affect the ability of HIV to infect lymphocytes. To study the role of matrix FN on HIV infection, we used superfibronectin (sFN), a multimeric form of FN that closely resembles in vivo matrix FN. In this study we show that HIV-1IIIB efficiently binds to multimeric fibronectin (sFN) and that HIV infection of primary CD4+ lymphocytes is enhanced by >1 order of magnitude in the presence of sFN. This increase appears to be due to increased adhesion of viral particles to the cell surface in the presence of sFN, followed by internalization of virus. Enzymatic removal of cell surface proteoglycans inhibited the adhesion of HIV-1IIIB/sFN complexes to lymphocytes. In contrast, Abs to integrins had no effect on binding of HIV 1IIIB/sFN complexes to lymphocytes. The III1-C peptide alone also bound HIV-1IIIB efficiently and enhanced HIV infection, although not as effectively as sFN. HIV 1IIIB gp120 envelope protein binds to the III1-C region of sFN and may be important in the interaction of virus with matrix FN. We conclude that HIV-1IIIB specifically interacts with the III1-C region within matrix FN, and that this interaction may play a role in facilitating HIV infection in vivo, particularly in lymph node tissue. PMID- 10706717 TI - A poxvirus protein that binds to and inactivates IL-18, and inhibits NK cell response. AB - IL-18 induces IFN-gamma and NK cell cytotoxicity, making it a logical target for viral antagonism of host defense. We demonstrate that the ectromelia poxvirus p13 protein, bearing homology to the mammalian IL-18 binding protein, binds IL-18, and inhibits its activity in vitro. Binding of IL-18 to the viral p13 protein was compared with binding to the cellular IL-18R. The dissociation constant of p13 for murine IL-18 is 5 nM, compared with 0.2 nM for the cellular receptor heterodimer. Mice infected with a p13 deletion mutant of ectromelia virus had elevated cytotoxicity for YAC-1 tumor cell targets compared with control animals. Additionally, the p13 deletion mutant virus exhibited decreased levels of infectivity. Our data suggest that inactivation of IL-18, and subsequent impairment of NK cell cytotoxicity, may be one mechanism by which ectromelia evades the host immune response. PMID- 10706718 TI - CD14 employs hydrophilic regions to "capture" lipopolysaccharides. AB - CD14 participates in the host innate inflammatory response to bacterial LPS obtained from Escherichia coli and other Gram-negative bacteria. Evidence from several laboratories suggests that different regions of the amino-terminal portion of the molecule may be involved in LPS binding. In this report a series of single-residue serine replacement and charge reversal mutations were generated to further elucidate the mechanism by which this protein may bind a multitude of different LPS ligands. Single-residue CD14 mutation proteins were examined for their ability to bind LPS obtained from E. coli, Porphyromonas gingivalis, and Helicobacter pylori and facilitate the activation of E-selectin from human endothelial cells. In addition, the single-residue CD14 mutation proteins were employed to perform monoclonal epitope-mapping studies with three LPS-blocking Abs that bound tertiary epitopes. Evidence that several different hydrophilic regions of the amino-terminal region of CD14 are involved in LPS binding was obtained. Epitope-mapping studies revealed that these hydrophilic regions are located on one side of the protein surface. These studies suggest that CD14 employs a charged surface in a manor similar to the macrophage scavenger receptor to "capture" LPS ligands and "present" them to other components of the innate host defense system. PMID- 10706719 TI - Role of CCAAT/enhancer-binding protein site in transcription of human neutrophil peptide-1 and -3 defensin genes. AB - The human neutrophil defensins (human neutrophil peptides (HNPs)), major components of azurophilic granules, contribute to innate and acquired host immunities through their potent antimicrobial activities and ability to activate T cells. Despite being encoded by nearly identical genes, HNP-1 is more abundant in the granules than HNP-3. We investigated the regulation of HNP-1 and HNP-3 expression at the transcriptional level using a promyelocytic HL-60 cell line. Luciferase analysis showed that transcriptional levels of HNP-1 and HNP-3 promoters were equivalent and that an approximately 200-bp region identical between promoters was sufficient for transcriptional activity. Furthermore, overlapping CCAAT/enhancer-binding protein (C/EBP) and c-Myb sites in the region were found to be required for efficient transcription. Gel mobility shift assay demonstrated that C/EBPalpha predominantly bound to the C/EBP/c-Myb sites using HL-60 nuclear extracts. No specific binding to C/EBP/c-Myb sites was observed in nuclear extracts from mature neutrophils, which expressed neither C/EBPalpha protein nor HNP mRNAs. Taken together, these findings suggest that the difference in the amounts of HNP-1 and HNP-3 peptides in neutrophils is caused by posttranscriptional regulation and that C/EBPalpha plays an important role in the transcription of HNP genes in immature myeloid cells. PMID- 10706720 TI - CD4+ T cell priming accelerates the clearance of Sendai virus in mice, but has a negative effect on CD8+ T cell memory. AB - Current vaccines designed to promote humoral immunity to respiratory virus infections also induce potent CD4+ T cell memory. However, little is known about the impact of primed CD4+ T cells on the immune response to heterologous viruses that are serologically distinct, but that share CD4+ T cell epitopes. In addition, the protective capacity of primed CD4+ T cells has not been fully evaluated. In the present study, we addressed these two issues using a murine Sendai virus model. Mice were primed with an HN421-436 peptide that represents the dominant CD4+ T cell epitope on the hemagglutinin-neuraminidase (HN) of Sendai virus. This vaccination strategy induced strong CD4+ T cell memory to the peptide, but did not induce Abs specific for the Sendai virus virion. Subsequent Sendai virus infection of primed mice resulted in 1) a substantially accelerated virus-specific CD4+ T cell response in the pneumonic lung; 2) enhanced primary antiviral Ab-forming cell response in the mediastinal lymph nodes; and 3) accelerated viral clearance. Interestingly, the virus-specific CD8+ T cell response in the lung and the development of long-term memory CD8+ T cells in the spleen were significantly reduced. Taken together, our data demonstrate that primed CD4+ T cells, in the absence of pre-existing Ab, can have a significant effect on the subsequent immune responses to a respiratory virus infection. PMID- 10706721 TI - The tryptophan catabolite picolinic acid selectively induces the chemokines macrophage inflammatory protein-1 alpha and -1 beta in macrophages. AB - We previously found that the tryptophan catabolite picolinic acid (PA) is a costimulus for the activation of macrophage effector functions. In this study, we have investigated the ability of PA to modulate the expression of chemokines in macrophages. We demonstrate that PA is a potent activator of the inflammatory chemokines MIP (macrophage inflammatory protein)-1 alpha and MIP-1 beta (MIPs) mRNA expression in mouse macrophages in a dose- and time-dependent fashion and through a de novo protein synthesis-dependent process. The induction by PA occurred within 3 h of treatment and reached a peak in 12 h. The stimulatory effects of PA were selective for MIPs because other chemokines, including monocyte chemoattractant protein-1, RANTES, IFN-gamma-inducible protein-10, MIP 2, and macrophage-derived chemokine, were not induced under the same experimental conditions and were not an epiphenomenon of macrophage activation because IFN gamma did not affect MIPs expression. Induction of both MIP-1 alpha and MIP-1 beta by PA was associated with transcriptional activation and mRNA stabilization, suggesting a dual molecular mechanism of control. Iron chelation could be involved in MIPs induction by PA because iron sulfate inhibited the process and the iron-chelating agent, desferrioxamine, induced MIPs expression. We propose the existence of a new pathway leading to inflammation initiated by tryptophan catabolism that can communicate with the immune system through the production of PA, followed by secretion of chemokines by macrophages. These results establish the importance of PA as an activator of macrophage proinflammatory functions, providing the first evidence that this molecule can be biologically active without the need for a costimulatory agent. PMID- 10706722 TI - Chemokines fail to up-regulate beta 1 integrin-dependent adhesion in human Th2 T lymphocytes. AB - Th1 and Th2 cells are functionally distinct subsets of CD4+ T lymphocytes whose tissue-specific homing to sites of inflammation is regulated in part by the differential expression of P- and E-selectin ligands and selected chemokine receptors. Here we investigated the expression and function of beta 1 integrins in Th1 and Th2 cells polarized in vitro. Th1 lymphocytes adhere transiently to the extracellular matrix ligands laminin 1 and fibronectin in response to chemokines such as RANTES and stromal cell-derived factor-1, and this process is paralleled by the activation of the Rac1 GTPase and by a rapid burst of actin polymerization. Selective inhibitors of phosphoinositide-3 kinase prevent efficiently all of the above processes, whereas the protein kinase C inhibitor bisindolylmaleimide prevents chemokine-induced adhesion without affecting Rac1 activation and actin polymerization. Notably, chemokine-induced adhesion to beta 1 integrin ligands is markedly reduced in Th2 cells. Such a defect cannot be explained by a reduced sensitivity to chemokine stimulation in this T cell subset, nor by a defective activation of the signaling cascade involving phosphoinositide-3 kinase, Rac1, and actin turnover, as all these processes are activated at comparable levels by chemokines in the two subsets. We propose that reduced beta 1 integrin-mediated adhesion in Th2 cells may restrain their ability to invade and/or reside in sites of chronic inflammation, which are characterized by thickening of basement membranes and extensive fibrosis, requiring efficient interaction with organized extracellular matrices. PMID- 10706723 TI - Leukocyte arrest during cytokine-dependent inflammation in vivo. AB - Leukocyte rolling along the walls of inflamed venules precedes their adhesion during inflammation. Rolling leukocytes are thought to arrest by engaging beta2 integrins following cellular activation. In vitro studies suggest that chemoattractants may instantaneously activate and arrest rolling leukocytes. However, how leukocytes stop rolling and become adherent in inflamed venules in vivo has remained rather mysterious. In this paper we use a novel method of tracking individual leukocytes through the microcirculation to show that rolling neutrophils become progressively activated while rolling down the venular tree. On average, leukocytes in wild-type mice roll for 86 s (and cover 270 microm) before becoming adherent with an efficiency around 90%. These rolling leukocytes exhibit a gradual beta2 integrin-dependent decrease in rolling velocity that correlates with an increase in intracellular free calcium concentration before arrest. Similar tracking analyses in gene-targeted mice demonstrate that the arrest of rolling leukocytes is very rare when beta2 integrins are absent or blocked by a mAb. Arrest is approximately 50% less efficient in the absence of E selectin. These data suggest a model of leukocyte recruitment in which beta2 integrins play a critical role in stabilizing leukocyte rolling during a protracted cellular activation period before arrest and firm adhesion. PMID- 10706724 TI - TNF-alpha, IL-4, and IFN-gamma regulate differential expression of P- and E selectin expression by porcine aortic endothelial cells. AB - P- and E-selectin are surface glycoproteins that mediate leukocyte rolling on the surface of endothelium in inflammation. We have cloned porcine P-selectin cDNA and generated a mAb, 12C5, with which to examine P-selectin expression by porcine aortic endothelial cells (PAEC) in comparison with that of E-selectin. Basal expression by PAEC of P-selectin was greater than that of E-selectin, whereas E selectin expression was more prominently enhanced than that of P-selectin by stimulation with TNF-alpha or IL-1alpha. Both human or porcine IL-4 led to an increase in P-selectin expression, with kinetics that were delayed compared with those seen following stimulation with TNF-alpha or IL-1alpha, but IL-4 did not stimulate expression of E-selectin. When cells were stimulated with TNF-alpha in the presence of IL-4, we observed enhanced P-selectin expression with a parallel reduction in E-selectin expression. Finally, the increase in P-selectin expression due to human IL-4 was reduced in the presence of porcine but not human IFN-gamma. These observations show that E-selectin and P-selectin expression are differentially regulated in PAEC, and that IL-4 leads to a shift in the relative surface density of the two molecules toward P-selectin. The ability of porcine IFN-gamma to inhibit IL-4-induced P-selectin expression suggests that the balance between Th1 and Th2 cytokine production may determine the relative densities of the two selectins in chronic immune-mediated inflammation. Because the increased expression of P-selectin induced by human IL-4 was not inhibited by human IFN gamma, this balance may be shifted toward P-selectin expression in porcine xenografts infiltrated by human lymphocytes. PMID- 10706725 TI - A functional role of I kappa B-epsilon in endothelial cell activation. AB - The NF-kappa B inhibitor I kappa B-epsilon is a new member of the I kappa B protein family, but its functional role in regulating NF-kappa B-mediated induction of adhesion molecule expression is unknown. In vascular endothelial cells, I kappa B-epsilon associates predominantly with the NF-kappa B subunit Rel A and to a lesser extent with c-Rel, whereas I kappa B-alpha and I kappa B-beta associate with Rel A only. Following stimulation with TNF-alpha, pyrrolidine dithiocarbamate (PDTC), N-acetylcysteine, and dexamethasone prevented I kappa B kinase-induced I kappa B-alpha, but not I kappa B-beta or I kappa B-epsilon phosphorylation and degradation. Since the activation of NF-kappa B is required for the induction of adhesion molecule expression, we examined the role of I kappa B-epsilon in the transactivation of promoters from VCAM-1, ICAM-1, and E selectin. Using reporter gene constructs of adhesion molecule promoters, PDTC inhibited VCAM-1 and E-selectin, but to a lesser extent, ICAM-1 promoter activity. Subcloning of kappa B cis-acting elements of VCAM-1, E-selectin, and ICAM-1 into a heterologous promoter construct revealed that PDTC inhibited VCAM-1 and E-selectin, but to a lesser extent, ICAM-1 kappa B promoter activity. By electrophoretic mobility shift assay, NF-kappa B heterodimers containing c-Rel specifically bind to the kappa B motif in the ICAM-1, but not VCAM-1 or E selectin promoter. Indeed, overexpression of c-Rel induced ICAM-1 kappa B promoter activity to a greater extent than that of E-selectin and overexpression of I kappa B-epsilon inhibited ICAM-1 and VCAM-1 promoter activity in endothelial cells. These findings indicate that c-Rel-associated I kappa B-epsilon is involved in the induction of ICAM-1 expression. PMID- 10706726 TI - Distinct T cell/renal tubular epithelial cell interactions define differential chemokine production: implications for tubulointerstitial injury in chronic glomerulonephritides. AB - Chemokines can promote interstitial fibrosis that is, in turn, a strong predictor of renal failure in chronic glomerulonephritides (GN). Resident renal cells, including renal tubular epithelial cells (RTEC), represent a prominent source of chemokine expression. Evaluating those factors responsible for sustained chemokine production by RTEC during GN is therefore crucial. The contribution of interstitial T cells to such expression, and in particular the precise nature of their interactions with RTEC, are poorly understood. Activated T cell/RTEC coculture induced production of high levels of monocyte chemoattractant protein-1 (MCP-1), RANTES, and IFN-inducible protein-10 from RTEC. Using double-chamber cultures and activated T cell plasma membrane preparations we demonstrated that both cell contact and soluble factors contributed to RTEC chemokine production. Importantly, different chemokines exhibited distinct activation requirements. Thus, for RANTES cell contact was essential, but not sufficient. In contrast, either soluble factors or cell contact induced MCP-1 and IFN-inducible protein-10 production, although both pathways were required for a maximal response. Neutralization experiments identified critical roles in this process for proinflammatory cytokines such as TNF-alpha, IL-1beta, and IFN-gamma as well as membrane molecules such as LFA-1, CD40 ligand, and membrane bound TNF-alpha. Finally, chemotactic bioassays of T cell/RTEC coculture supernatants demonstrated 80% reduction of monocyte migration following MCP-1 neutralization, indicating a dominant role for this chemokine. In summary, activation of renal tubular cells by infiltrating T cells can amplify and perpetuate local inflammatory responses through chemokine production differentially mediated by soluble and cell contact dependent factors. Recognition of this regulatory diversity has important implications in the choice of potential therapeutic targets in GN. PMID- 10706727 TI - IFN-gamma-dependent and -independent mechanisms in adverse effects caused by concomitant administration of IL-18 and IL-12. AB - IL-18 is a new type of inflammatory cytokine similar to but distinct from IL-12 and IL-1beta. One intriguing property of IL-18 is synergism with IL-12 in many respects. In this study we examined the in vivo synergistic effects of IL-18/IL 12 in mice and found lethal toxicity accompanying an elevated IFN-gamma level in the serum. Since treatment with IL-18 alone did not have any apparent toxicity, and treatment with IL-12 alone showed only limited toxicity in our system, the synergy between the two cytokines was all the more remarkable. The major symptoms of the toxicity were weight loss, diarrhea, hemorrhagic colitis, splenomegaly, fatty liver, and atrophic thymus, most of which are similarly found in endotoxin induced septic shock. However, in contrast to septic shock, TNF-alpha was not induced. The involvement of IFN-gamma in the toxicity was further studied in detail. Treatment of athymic nude mice with anti-asialo-GM1 did not reduce the toxicity, whereas anti-IFN-gamma treatment of wild-type mice alleviated it. When IFN-gamma-deficient mice were treated with IL-18/IL-12, the majority of them showed mortality and toxicity with severe pulmonary edema. These results indicate that IL-18/IL-12 treatment induces severe adverse effects through not only IFN gamma-dependent mechanisms but also IFN-gamma-independent processes. PMID- 10706728 TI - Alpha 4 integrin-dependent leukocyte recruitment does not require VCAM-1 in a chronic model of inflammation. AB - Rats immunized with Mycobacterium butyricum in Freund's adjuvant develop a chronic vasculitis, with large increases in leukocyte rolling and adhesion in mesenteric postcapillary venules that are significantly inhibited with an alpha 4 integrin Ab. Using intravital microscopy to visualize chronically inflamed microvessels, we demonstrated that alpha 4 integrin-dependent leukocyte rolling and adhesion was inhibited with a beta 1 integrin, but not a beta 7 integrin Ab. To date, VCAM-1 has been presumed to be the primary ligand for alpha 4 beta 1 integrin in the vasculature. However, alpha 4 beta 1 integrin-dependent interactions were not reduced by monoclonal or polyclonal VCAM-1 Abs or a VCAM-1 antisense oligonucleotide despite increased VCAM-1 expression in the mesenteric vasculature. To ensure that the VCAM-1 Abs were functional and used at saturating concentrations, blood from Ab-treated rats was perfused over monolayers of CHO cells transfected with rat VCAM-1. Sufficient alpha 4 integrin or VCAM-1 Ab was present to inhibit leukocyte interactions with rat VCAM-1 by 95-100%. Under in vitro flow conditions, only mononuclear leukocytes were recruited from blood of control rats onto purified VCAM-1. However, neutrophils were also recruited onto VCAM-1 from whole blood of adjuvant-immunized animals via alpha 4 integrin. Another ligand for alpha 4 beta 1 integrin is the connecting segment-1 (CS-1) region of fibronectin. An Ab to the CS-1 portion of fibronectin, which did not reduce rolling and adhesion in adjuvant arthritis animals, completely inhibited leukocyte adhesion to CS-1 under static conditions. These findings provide the first evidence that alpha 4 beta 1 integrin-dependent leukocyte rolling and adhesion can occur in vivo via a mechanism other than VCAM-1. PMID- 10706729 TI - The coupling of 5-oxo-eicosanoid receptors to heterotrimeric G proteins. AB - 5-Oxo-eicosatetraenoic acid (5-oxoETE) stimulated human neutrophil (PMN) and eosinophil chemotaxis, PMN hexose uptake, and PMN membrane GTP/GDP exchange. Pertussis toxin (PT), a blocker of heterotrimeric G proteins (GP), completely inhibited these responses, but proved far less effective on the same responses when elicited by leukotriene B4, C5a, FMLP, platelet-activating factor, IL-8, or RANTES chemotactic factors. 5-OxoETE also specifically bound to the membrane preparations that conducted GTP/GDP exchange. This binding was down-regulated by GTPgammaS, but not ADPgammaS, and displaced by 5-oxoETE analogues, but not by leukotriene B4, lipoxin A4, or lipoxin B4. Finally, PMN expressed PT-sensitive GP alphaiota2 and PT-resistant GP alphaq/11- and alpha13-chains; eosinophils expressed only alphai2 and alphaq/11. We conclude that 5-oxoETE activates granulocytes through a unique receptor that couples preferentially to PT sensitive GP. The strict dependency of this putative receptor on PT-sensitive GP may underlie the limited actions of 5-oxoETE, compared with other CF, and help clarify the complex relations between receptors, GP, cell signals, and cell responses. PMID- 10706730 TI - Expression and activation of NF-kappa B in the antrum of the human stomach. AB - Both in vitro studies and experiments in mice suggest a key role for transcription factor NF-kappa B as a mediator of mucosal inflammation. Experiments in vitro show that NF-kappa B activation may be a critical event in the production of proinflammatory molecules in Helicobacter pylori-associated gastritis. This study examines the expression and activity of NF-kappa B in situ in antral biopsies of 69 consecutive patients with immunohistochemical techniques. In the uninflamed stomach, NF-kappa B was highly expressed and active in a subset of epithelial cells, which were identified as predominantly G cells. In accordance with this activity, G cells were shown to express high levels of the NF-kappa B target cytokine TNF-alpha, a well-documented stimulator of gastrin production. In patients with H. pylori-associated gastritis, NF-kappa B activity was markedly enhanced. Activation occurred preferentially in the epithelial cells. The number of cells showing activated NF-kappa B correlated with the activity of gastritis, a measure of neutrophil influx, whereas no correlation was found with the chronicity of inflammation, a measure of the presence of mononuclear inflammatory cells. This correlation is direct evidence of the importance of NF-kappa B-dependent signal transduction for neutrophil influx in H. pylori-associated gastritis. PMID- 10706731 TI - Recombinant human platelet-activating factor-acetylhydrolase inhibits airway inflammation and hyperreactivity in mouse asthma model. AB - Numerous in vitro and in vivo studies in both animal models and human asthmatics have implicated platelet-activating factor (PAF) as an important inflammatory mediator in asthma. In a murine asthma model, we examined the anti-inflammatory activities of recombinant human PAF-acetylhydrolase (rPAF-AH), which converts PAF to biologically inactive lyso-PAF. In this model, mice sensitized to OVA by i.p. and intranasal (i.n.) routes are challenged with the allergen by i.n. administration. The OVA challenge elicits an eosinophil infiltration into the lungs with widespread mucus occlusion of the airways and results in bronchial hyperreactivity. The administration of rPAF-AH had a marked effect on late-phase pulmonary inflammation, which included a significant reduction in airway eosinophil infiltration, mucus hypersecretion, and airway hyperreactivity in response to methacholine challenge. These studies demonstrate that elevating plasma levels of PAF-AH through the administration of rPAF-AH is effective in blocking the late-phase pulmonary inflammation that occurs in this murine allergen-challenge asthma model. These results suggest that rPAF-AH may have therapeutic effects in patients with allergic airway inflammation. PMID- 10706733 TI - High susceptibility for cystic fibrosis human airway gland cells to produce IL-8 through the I kappa B kinase alpha pathway in response to extracellular NaCl content. AB - Increasing evidence suggests that in airways from cystic fibrosis (CF) patients, inflammation may precede bacterial infection and be related to an endogenous dysregulation of proinflammatory cytokines in airway epithelial cells. Several investigators have reported that, in CF airway fluids, elevated NaCl concentrations may also contribute to the diseased state by inhibiting the bactericidal properties of airway fluid. Because many proinflammatory cytokines are transcriptionally regulated by the NF-kappa B, we investigated whether an elevated extracellular NaCl content in airway fluids significantly impaired the regulation of the NF-kappa B/I kappa B alpha complex and the chemokine IL-8 production in primary non-CF and CF human bronchial gland epithelial cells. Exposure of non-CF gland cells to hypotonic (85 mM) NaCl solution, compared with isotonic (115 mM) NaCl and hypertonic (170 mM) NaCl solutions, resulted in a significant decrease in IL-8 production that was paralleled by a strong inhibition of activated NF-kappa B associated with an increased cytosolic expression of I kappa B alpha and a decrease in the I kappa B kinase alpha protein level. In CF gland cells, we demonstrated that, compared with the high IL 8 in an hypertonic solution, the release of IL-8 was significantly reduced 2-fold in an isotonic solution and 5-fold in a hypotonic solution. Strikingly, exposure of CF bronchial gland cells to either hypotonic or isotonic milieu did not result in a marked inhibition of the activated NF-kappa B/I kappa B alpha system. This is the first demonstration that primary human CF bronchial gland cells exhibit abnormally high IL-8 production through constitutively activated NF-kappa B and high I kappa B kinase alpha level, whatever the hypo-, iso-, and hypertonic NaCl milieu. PMID- 10706732 TI - Fractalkine is an epithelial and endothelial cell-derived chemoattractant for intraepithelial lymphocytes in the small intestinal mucosa. AB - Fractalkine is a unique chemokine that combines properties of both chemoattractants and adhesion molecules. Fractalkine mRNA expression has been observed in the intestine. However, the role of fractalkine in the healthy intestine and during inflammatory mucosal responses is not known. Studies were undertaken to determine the expression and function of fractalkine and the fractalkine receptor CX3CR1 in the human small intestinal mucosa. We identified intestinal epithelial cells as a novel source of fractalkine. The basal expression of fractalkine mRNA and protein in the intestinal epithelial cell line T-84 was under the control of the inflammatory mediator IL-1beta. Fractalkine was shed from intestinal epithelial cell surface upon stimulation with IL-1beta. Fractalkine localized with caveolin-1 in detergent-insoluble glycolipid-enriched membrane microdomains in T-84 cells. Cellular distribution of fractalkine was regulated during polarization of T-84 cells. A subpopulation of isolated human intestinal intraepithelial lymphocytes expressed the fractalkine receptor CX3CR1 and migrated specifically along fractalkine gradients after activation with IL-2. Immunohistochemistry demonstrated fractalkine expression in intestinal epithelial cells and endothelial cells in normal small intestine and in active Crohn's disease mucosa. Furthermore, fractalkine mRNA expression was significantly up regulated in the intestine during active Crohn's disease. This study demonstrates that fractalkine-CX3CR1-mediated mechanism may direct lymphocyte chemoattraction and adhesion within the healthy and diseased human small intestinal mucosa. PMID- 10706734 TI - CD4+ T cell and eosinophil adhesion is mediated by specific ICAM-3 ligation and results in eosinophil activation. AB - T cells and eosinophils, which are found in close proximity in asthmatic lungs, express many surface receptors that are counterligands. These data suggest that direct interactions between these cell types could play an important role in regulating airway inflammation in asthma. We examined the effect of selective adhesion between counterligands on human eosinophils and CD4+ T cells to determine 1) the existence of specific adhesive interactions and 2) if augmented specific adhesion to CD4+ T cells also caused augmented secretion of leukotriene C4 (LTC4) from eosinophils. A new method for binding of human CD4+ T cells to microwell plates was developed, which allowed for specific quantitative assessment of eosinophil adhesion to individual CD4+ T cells in culture. Adhesion of CD4+ T cells to eosinophils was minimal in unstimulated cells but increased after activation of T cells by PMA. Augmented adhesion was regulated substantially through binding of ICAM-3 and only minimally by ICAM-1. We further evaluated whether this specific adhesion up-regulated stimulated secretion of LTC4 from eosinophils. Adhesion with CD4+ T cells augmented eosinophil secretion of LTC4 caused by FMLP plus cytochalasin. Blockade of ICAM-3, as well as ICAM-1, inhibited completely the augmented secretion of eosinophil LTC4. We demonstrate that eosinophils and CD4+ T cells are capable of ligand-specific adhesion that is mediated predominantly by ICAM-3 ligation and that this binding causes augmented eosinophil secretion. PMID- 10706735 TI - Cutaneous injection of human subjects with macrophage inflammatory protein-1 alpha induces significant recruitment of neutrophils and monocytes. AB - Macrophage inflammatory protein (MIP-1 alpha), a member of the CC chemokine subfamily, has been shown to attract T cells and monocytes in vitro and to be expressed at sites of inflammation. Although the in vitro activities of MIP-1 alpha have been well documented, the in vivo biological activities of MIP-1 alpha in humans have not been studied. To address this, we challenged human subjects by intradermal injection with up to 1000 pmol of MIP-1 alpha and performed biopsies 2, 10, and 24 h later. Although no acute cutaneous or systemic reactions were noted, endothelial cell activation, as indicated by the expression of E-selectin, was observed. In agreement with its in vitro activity, monocyte, lymphocyte, and, to a lesser degree, eosinophil infiltration was observed, peaking at 10-24 h. Surprisingly, in contrast to its reported lack of in vitro neutrophil-stimulating activity, a rapid infiltration of neutrophils was observed in vivo. This neutrophil infiltration occurred as early as 2 h, preceding the appearance of other cells, and peaked at 10 h. Interestingly, we found that neutrophils in whole blood, but not after isolation, expressed CCR1 on their cell surface. This CCR1 was thought to be functional as assessed by neutrophil CD11b up-regulation following whole-blood MIP-1 alpha stimulation. These studies substantiate the biological effects of MIP-1 alpha on monocytes and lymphocytes and uncover the previously unrecognized activity of MIP-1 alpha to induce neutrophil infiltration and endothelial cell activation, underscoring the need to evaluate chemokines in vivo in humans. PMID- 10706736 TI - In vitro priming with adenovirus/gp100 antigen-transduced dendritic cells reveals the epitope specificity of HLA-A*0201-restricted CD8+ T cells in patients with melanoma. AB - Replication-deficient recombinant adenovirus (Ad) encoding human gp100 or MART-1 melanoma Ag was used to transduce human dendritic cells (DC) ex vivo as a model system for cancer vaccine therapy. A second generation E1/E4 region deleted Ad which harbors the CMV immediate-early promoter/enhancer and a unique E4-ORF6/pIX chimeric gene was employed as the backbone vector. We demonstrate that human monocyte-derived DC are permissive to Ad infection at multiplicity of infection between 100 and 500 and occurs independent of the coxsackie Ad receptor. Fluorescent-labeled Ad was used to assess the kinetics and distribution of viral vector within DC. Ad-transduced DC show peak transgene expression at 24-48 h and expression remains detectable for at least 7 days. DC transduced with replication deficient Ad do not exhibit any unusual phenotypic characteristics or cytopathic effects. DC transduced with Ad2/gp100v2 can elicit tumor-specific CTL in vitro from patients bearing gp100+ metastatic melanoma. Using a panel of gp100-derived synthetic peptides, we show that Ad2/gp100v2-transduced DC elicit Ag-specific CTL that recognize only the G209 and G280 epitopes, both of which display relatively short half-lives ( approximately 7-8 h) on the surface of HLA-A*0201+ cells. Thus, patients with metastatic melanoma are not tolerant to gp100 Ag based on the detection of CD8+ T cells specific for multiple HLA-A*0201-restricted, gp100 derived epitopes. PMID- 10706737 TI - Prediction of the immunodominant epitope of the pyruvate dehydrogenase complex E2 in primary biliary cirrhosis using phage display. AB - Primary biliary cirrhosis (PBC) is an autoimmune liver disease characterized by autoantibodies reactive with the pyruvate dehydrogenase complex. A conformational epitope has been mapped to aa 91-227 within the inner lipoyl domain of the E2 subunit (pyruvate dehydrogenase complex E2 (PDC-E2)). We have used phage display to further localize this epitope. A random heptapeptide library was screened using IgG from two patients with PBC, with negative selection using pooled normal IgG. Phage that contained peptide inserts (phagotopes) selected using PBC sera differed from those selected using IgG from patients with RA or polychondritis. Two motifs occurred only among the PBC-selected phagotopes; these were MH (13 sequences, 16 phagotopes) and FV (FVEHTRW, FVEIYSP, FVLPWRI). The phagotopes selected were tested for reactivity with anti-PDC-E2 affinity purified from four patients with PBC. Phagotopes that contained 1 of 15 different peptide sequences were reactive with one or more of these four anti-PDC-E2 preparations, whereas phagotopes that contained 1of the remaining 28 sequences were negative. The peptides (FVLPWRI, MHLNTPP, MHLTQSP) encoded by three phagotopes that were strongly reactive with all four preparations of anti-PDC-E2 were synthesized. Each of the selected peptides, but not an irrelevant peptide, inhibited the reactivity by ELISA of PBC serum with recombinant PDC-E2 and reduced the inhibition of the enzyme activity of PDC by a PBC serum. The peptide sequences, along with the known NMR structure of the inner lipoyl domain of PDC-E2, allow the prediction of nonsequential residues 131HM132 and 178FEV180 that contribute to a conformational epitope. PMID- 10706738 TI - Genetic analysis of the influence of pertussis toxin on experimental allergic encephalomyelitis susceptibility: an environmental agent can override genetic checkpoints. AB - Pertussis toxin (PTX) is a potent ancillary adjuvant used to elicit several different autoimmune diseases, including experimental allergic encephalomyelitis (EAE). To delineate the genetics of PTX effect in EAE, we mapped EAE-modifying (eae-m) loci in cohorts of backcross mice immunized with and without PTX. In this study, we analyzed the genetic basis of EAE susceptibility and severity and the intermediate phenotypes of mononuclear cell infiltration, suppuration, and demyelination. In animals immunized with PTX, one major locus, eae9, controls disease susceptibility and severity. Eae9 also regulates the extent of mononuclear cell infiltration of the spinal cord in male mice. Without PTX, five eae-m loci were noted, including three new loci in intervals on chromosomes 8 (eae14), 10 (eae17), and 18 (eae18). Taken together, these results suggest that eae9 controls the effects of PTX in EAE susceptibility, and is capable of overriding the other genetic checkpoints in the pathogenesis of this disease. PMID- 10706741 TI - Corrections PMID- 10706739 TI - CCR5-reactive antibodies in seronegative partners of HIV-seropositive individuals down-modulate surface CCR5 in vivo and neutralize the infectivity of R5 strains of HIV-1 In vitro. AB - Exposure to HIV does not necessarily result in infection. Because primary HIV infection is associated with CCR5-tropic HIV variants (R5), CCR5-specific Abs in the sera of HIV-seronegative, HIV-exposed individuals (ESN) might be associated with protection against infection. We analyzed sera from ESN, their HIV-infected sexual partners (HIV+), and healthy controls (USN) searching for CCR5-specific Abs, studying whether incubation of PBMC with sera could prevent macrophage inflammatory protein 1 beta (Mip1 beta) (natural ligand of CCR5) binding to CCR5. Results showed that Mip1 beta binding to CCR5 was not modified by sera of either 40 HIV+ or 45 USN but was greatly reduced by sera of 6/48 ESN. Binding inhibition was due to Abs reactive with CCR5. The CCR5-specific Abs neutralized the infectivity of primary HIV isolates obtained from the corresponding HIV+ partners and of R5-primary HIV strains, but not that of CXCR4-tropic or amphitropic HIV strains. Immunoadsorption on CCR5-transfected, but not on CXCR4-transfected, cells removed CCR5-specific and virus-neutralizing Abs. Epitope mapping on purified CCR5-specific Abs showed that these Abs recognize a conformational epitope in the first cysteine loop of CCR5 (aa 89-102). Affinity-purified anti CCR5-peptide neutralized the infectivity of R5 strains of HIV-1. Anti-CCR5 Abs inhibited Mip1beta-induced chemotaxis of PBMC from healthy donors. PBMC from two ESN (with anti-CCR5 Abs) were CCR5-negative and could not be stimulated by Mip1beta in chemotaxis assays. These results contribute to clarifying the phenomenon of immunologic resistance to HIV and may have implications for the development of a protective vaccine. PMID- 10706740 TI - Blockade of CD28-B7, but not CD40-CD154, prevents costimulation of allogeneic porcine and xenogeneic human anti-porcine T cell responses. AB - Despite increasing use of swine in transplantation research, the ability to block costimulation of allogeneic T cell responses has not been demonstrated in swine, and the effects of costimulatory blockade on xenogeneic human anti-porcine T cell responses are also not clear. We have compared the in vitro effects of anti-human CD154 mAb and human CTLA4IgG4 on allogeneic pig T cell responses and xenogeneic human anti-pig T cell responses. Both anti-CD154 mAb and CTLA4IgG4 cross-reacted on pig cells. While anti-CD154 mAb and CTLA4IgG4 both inhibited the primary allogeneic pig MLRs, CTLA4IgG4 (7.88 microg/ml) was considerably more inhibitory than anti-CD154 mAb (100 microg/ml) at optimal doses. Anti-CD154 mAb inhibited the production of IFN-gamma by 75%, but did not inhibit IL-10 production, while CTLA4IgG4 completely inhibited the production of both IFN-gamma and IL-10. In secondary allogeneic pig MLRs, CTLA4IgG4, but not anti-CD154 mAb, induced Ag specific T cell anergy. CTLAIgG4 completely blocked the indirect pathway of allorecognition, while anti-CD154 mAb blocked the indirect response by approximately 50%. The generation of porcine CTLs was inhibited by CTLA4IgG4, but not by anti-CD154 mAb. Human anti-porcine xenogeneic MLRs were blocked by CTLA4IgG4, but only minimally by anti-CD154 mAb. Finally, CTLA4IgG4 prevented secondary xenogeneic human anti-porcine T cell responses. These data indicate that blockade of the B7-CD28 pathway was more effective than blockade of the CD40 CD154 pathway in inhibiting allogeneic pig T cell responses and xenogeneic human anti-pig T cell responses in vitro. These findings have implications for inhibiting cell-mediated immune responses in pig-to-human xenotransplantation. PMID- 10706742 TI - [Active francophone medicine. Speech of Pr Pierre Farah at the Cambodia Health 2000 Congress, Phnom Penh, Cambodia, 15-19 November 1999]. PMID- 10706743 TI - Pulmonary mineral fibers after occupational and environmental exposure to asbestos in the Russian chrysotile industry. AB - BACKGROUND: As an indicator of occupational, domestic, and environmental exposure, the level and type of asbestos fibers were determined from lung tissue samples of workers and residents who resided in the area of the world's largest asbestos mine at Asbest, Russia. METHODS: Electron microscopy was used to analyze and measure the concentration of asbestos fibers in a series of 47 autopsies at the Asbest Town Hospital. Work histories were obtained from pathology reports and employment records. RESULTS: In 24 chrysotile miners, millers, and product manufacturers, the pulmonary concentrations of retained fibers (over 1 microm in length) were 0. 8-50.6 million f/g for chrysotile, and < 0.1-1.9 million f/g for amphiboles (tremolite and anthophyllite). The concentrations were lower in 23 persons without any known occupational contact with asbestos; 0.1-14.6 million f/g for chrysotile, and < 0.1-0.7 million f/g for amphiboles. On average, 90% of all inorganic fibers were chrysotile, and 5% tremolite/anthophyllite. No amosite or crocidolite fibers were detected in any of the samples. CONCLUSIONS: The mean and range of pulmonary chrysotile concentrations were about the same as reported previously from the Canadian mining and milling industry. In the Russian samples, the mean concentration of tremolite fibers were less by at least one order of magnitude. Occupational contact was the most important source of asbestos exposure. PMID- 10706744 TI - Comparison of NIOSH noise criteria and OSHA hearing conservation criteria. AB - BACKGROUND: This study was conducted to compare noise exposure measurements based on the recently revised noise exposure criteria recommended by the U.S. National Institute for Occupational Safety and Health (NIOSH) and the current U.S. Occupational Safety and Health Administration (OSHA) Hearing Conservation Amendment to the occupational noise standard. METHODS: Daily 8-hour time-weighted average (TWA) personal noise exposures were obtained for 61 workers using dosimeters set simultaneously to the NIOSH and OSHA Hearing Conservation Amendment (OSHA-HCA) criteria. A variety of work groups with the potential for noise exposure were evaluated as a part of this investigation. RESULTS: Noise dose based on the NIOSH criteria was higher than the corresponding OSHA-HCA noise dose with differences in noise exposures measured under the two criteria equal to 6.6 dBA. Should the new NIOSH recommendation on noise measurement be adopted as standard, the number of workers to be enrolled in a hearing loss prevention program was estimated to increase by 2. 7-fold from 23% to 75% of the study population. CONCLUSIONS: The results of this study indicate that if the NIOSH criteria are to be adopted as an OSHA standard, there is likely to be a substantial increase in the number of workers in hearing conservation programs. PMID- 10706746 TI - Elevated risk for male breast cancer after occupational exposure to gasoline and vehicular combustion products. AB - BACKGROUND: Automotive gasoline contains benzene, 1,3-butadiene, 1, 2 dibromoethane and 1,2-dichloroethane, and the combustion products include certain polycyclic aromatic hydrocarbons, all of which have shown mammary gland carcinogenicity in long-term bioassays. It is the aim of this paper to investigate whether men occupationally exposed to gasoline and its combustion products have an elevated risk of breast cancer. METHODS: A nationwide register based case control study on male breast cancer morbidity was established among members of a pension fund, compulsory for all employees. Employment histories were reconstructed for each of 230 cases and 12,880 control subjects based on computerized records. The odds ratios, adjusted for socioeconomic status, were estimated by conditional logistic regression analysis. RESULTS: When a lag time of at least 10 years was included, the odds ratio for breast cancer among men with over three months of employment in trades with potential exposure to gasoline and combustion products was 2.5 (95% confidence interval: 1.3-4.5). Among men younger than 40 years at the time of first employment, the odds ratio was 5.4 (2.4-11.9). CONCLUSIONS: This study supports the hypothesis that occupational exposure to gasoline vapors and combustion products may play a role in the causation of male breast cancer. This hypothesis warrants further evaluation particularly in women. PMID- 10706745 TI - Mortality among Catholic nuns certified as radiologic technologists. AB - BACKGROUND: Several studies have shown that Catholic nuns have a different mortality experience than women of similar age in the general population. We had a unique opportunity to evaluate mortality patterns of nuns identified in an occupational study of nearly 145,000 radiologic technologists (73% female). METHODS: A total of 1,103 women were classified as nuns based on their titles of "Sister" or "SR". Their mortality experience was compared to other female radiologic technologists and to U.S. white females. RESULTS: Five hundred eighty three nuns (53%) were deceased as of January 1, 1995. Compared to other technologists, nuns were at significantly increased risk of dying from all causes (Standardized mortality ratio (SMR)=1.1; 95% Confidence interval (CI)=1.0-1.2, stomach cancer (SMR=2.7; 95% CI=1.2-5.4), diabetes (SMR=2.2; 95% CI=1.0-4.1), ischemic heart disease (SMR=1.2; 95% CI=1.1-1.4), all digestive diseases (SMR=2.0; 95% CI=1.3-3.0), and gastric and duodenal ulcers (SMR=8.3; 95% CI=2.3 21.3). In contrast, we observed a significant deficit in lung cancer (SMR=0.5; 95% CI=0.2-0.9), no deaths from cervical cancer, and a breast cancer risk 10% lower than expected (SMR=0.9; 95% CI=0.6-1.3). When compared to U.S. females, nuns experienced significantly reduced mortality from all causes (SMR=0.8; 95% CI=0.7-0.9), cervical cancer (SMR=0.0; 95% CI=0.0-0.7), all endocrine, metabolic and nutritional diseases (SMR=0.5; 95% CI=0.3-0.9), all circulatory diseases (SMR=0.7; 95% CI=0.7-0.8) including ischemic heart disease and cerebrovascular disease, and all respiratory diseases (SMR=0.5; 95% CI=0.3-0.8), and a nearly significant deficit of diabetes (SMR=0.6; 95% CI=0.3-1.0). In contrast, nuns had an almost 3-fold greater risk of tuberculosis (SMR=2.9; 95% CI=1.4-5.3) and a 20% excess of breast cancer (SMR=1. 2; 95% CI=0.8-1.7). The breast cancer excess was concentrated among nuns first certified before 1940 (SMR=2.0; CI=1.3-3.0), when radiation doses were possibly the highest, but the risk did not increase with increasing length of certification. CONCLUSIONS: Compared with the general population, the mortality experience of nuns was favorable and reflected the "healthy worker effect" commonly seen in occupational studies. Patterns observed for breast and cervical cancer possibly indicate differences in reproductive and sexual activities associated with belonging to a religious order. The possibility of a radiation-related excess for breast cancer among nuns certified before 1940 cannot be completely discounted, although there was no dose-response relationship with a surrogate measure of exposure (number of years certified). When their mortality experience was compared with other radiologic technologists, the influence of lifestyle factors was not apparent. Am. J. Ind. Med. 37:339-348, 2000. Published 2000 Wiley-Liss, Inc. dagger PMID- 10706747 TI - Acute respiratory symptoms in workers exposed to vanadium-rich fuel-oil ash. AB - BACKGROUND: Occupational exposure to fuel-oil ash, with its high vanadium content, may cause respiratory illness. It is unclear, however, what early acute health effects may occur on the pathway from normal to compromised respiratory function. METHODS: Using a repeated measures design, we studied prospectively 18 boilermakers overhauling an oil-fired boiler and 11 utility worker controls. Subjects completed a respiratory symptom diary five times per day by using a 0-3 scale where 0=symptom not present, 1=mild symptom, 2=moderate symptom, and 3=severe symptom. Daily symptom severity was calculated by using the highest reported score each day for upper and lower respiratory symptoms. Daily symptom frequency was calculated by summing all upper or lower airway symptom reports, then dividing by number of reporting times. Respiratory symptom frequency and severity were analyzed for dose-response relationships with estimated vanadium and PM(10) doses to the lung and upper airway by using robust regression. RESULTS: During the overhaul, 72% of boilermakers reported lower airway symptoms, and 67% reported upper airway symptoms. These percentages were 27 and 36 for controls. Boilermakers had more frequent and more severe upper and lower respiratory symptoms compared to utility workers, and this difference was greatest during interior boiler work. A statistically significant dose-response pattern for frequency and severity of both upper and lower respiratory symptoms was seen with vanadium and PM(10) in the three lower exposure quartiles. However, there was a reversal in the dose-response trend in the highest exposure quartile, reflecting a possible healthy worker effect. CONCLUSIONS: Boilermakers experience more frequent and more severe respiratory symptoms than utility workers. This is most statistically significant during boiler work and is associated with increasing dose estimates of lung and nasal vanadium and PM(10). PMID- 10706748 TI - Expanded analysis of injury mortality among unionized construction workers. AB - BACKGROUND: To evaluate the utility of expanding the number and precision of injury categories used in previous occupational mortality studies, this study reanalyzed data from four previous studies of unionized construction workers (construction laborers, ironworkers, sheet metal workers, and operating engineers), by expanding the number of injury categories from 6 to 33. METHODS: Proportionate mortality ratios (PMRs) were computed using the distribution of deaths from the National Occupational Mortality Surveillance System, a mortality surveillance system from 28 states, as a comparison. A blue collar comparison group was also used in additional analyses to adjust for socioeconomic and other factors. RESULTS: This reanalysis identified significantly elevated PMRs in at least one of the four worker groups for falls, motor vehicle crashes, machinery incidents, electrocutions, being struck by falling objects, being struck by flying objects, explosions, suffocation, and water transport incidents. Limiting the comparison population to deaths among blue collar workers did not change the results substantially. CONCLUSIONS: This study demonstrates that increasing the precision of categories of death from injury routinely used in mortality studies will provide improved information to guide prevention. Am. J. Ind. Med. 37:364 373, 2000. Published 2000 Wiley-Liss, Inc. PMID- 10706749 TI - The impact of agricultural injury on farm owners and workers in Alabama and Mississippi. AB - BACKGROUND: Few studies have assessed the consequences of agricultural injury and none have done so comparing Caucasian and African-American farm owners and workers. METHODS: 1244 farmers were enrolled and prospectively observed between 1994-1996 for farming-related injuries. Injured farmers provided information on the consequences of injuries. RESULTS: One-hundred and thirty-one subjects reported a total of 140 injuries. The majority of injuries were classified as minor or moderate and required medical attention. African-American farm workers tended to have more severe injuries. Nearly all injured subjects experienced acute residual effects (e.g., pain when moving), while persistent effects occurred in about half of the injured subjects, the latter being more common among African-American workers. Lost work was a frequent nonmedical effect of the injury. African-American workers tended to be more likely to lose work and/or be hurt financially. CONCLUSIONS: The impact of agricultural injury is nontrivial, particularly for African-American farm workers. The provision of better medical care facilities for African-American farm workers may be a positive approach for reducing the impact of agricultural injuries in this population. PMID- 10706750 TI - The prevalence of depressive symptoms and risk factors among Iowa and Colorado farmers. AB - BACKGROUND: Farmers have been previously reported to have higher rates of depression and suicide compared to other occupations. Comparisons of depressive symptoms and risk factors for states should increase understanding of depression in farmers. METHODS: Representative samples of 385 Iowa and 470 Colorado male principal farm operators in the respective state Farm Family Health and Hazard Surveillance projects were evaluated for depressive symptoms by using the CES-D scale. Risk factors were determined by using weighted multiple logistic regression analyses. RESULTS: Iowa farmers were 1.74 times (P < 0.05) more likely to have had depressive symptoms than Colorado farmers. Being unmarried (odds ratio=3.46), having negative life events within the past year [legal problems (4.67), substantial income decline (2.71), loss of something of sentimental value (3.20)], and lower perceived general health status were risk factors (P < 0.05) for depressive symptoms for male Iowa and Colorado farmers. Higher levels of most risk factors for Iowa farmers, almost twice the frequency of substantial income decline, accounted for the majority of the difference in depressive symptoms between Iowa (12.2%) and Colorado (7.4%). CONCLUSIONS: Although Iowa farmers had 1.74 times higher level of depression symptoms than Colorado farmers, this difference was not significant after adjusting for the higher levels of most risk factors for Iowa farmers. PMID- 10706751 TI - Analysis of construction injury burden by type of work. AB - BACKGROUND: To lay groundwork for identifying patterns of injury etiology, we sought to describe injury experience associated with types of work performed at construction sites by examining workers' compensation (WC) claims for the 32,081 construction workers who built Denver International Airport (DIA). METHODS: Injury rates and WC payment rates were calculated for 25 types of work based on claims and payroll data reported to DIA's owner-controlled insurance program according to National Council on Compensation Insurance job classifications. By linking DIA claims with corresponding lost-work-time (LWT) claims filed with Colorado's Workers' Compensation Division, we were also able to obtain and examine both total and median lost days for each type of work. RESULTS: Injury experience varied widely among the types of construction work. Workers building elevators and conduits and installing glass, metal, or steel were at particularly high risk of both LWT and non-LWT injury. Median days lost by injured workers was highest (202 days) for driving/trucking. Median days lost for most types of work was much greater than previously reported for construction: 40 days or more for 18 of the 25 types of work analyzed. WC payment rates reflect both number and severity of injuries and were generally not significantly different from expected losses. They were, however, significantly higher than expected for driving/trucking, metal/steel installation, inspection/analysis, and elevator construction. CONCLUSIONS: Analysis of injury data by type of work allows targeting of safety resources to high risk construction work and would be useful in prospective surveillance at large construction sites with centrally administered workers' compensation plans. PMID- 10706752 TI - Outcomes in work-related upper extremity and low back injuries: results of a retrospective study. AB - BACKGROUND: The outcomes of treatment for work-related injuries and illnesses are multidimensional and complex, but have rarely been explored in detail. This study was intended to provide information on a sample of workers representing a range of jobs and employers typical of the workers compensation system. METHODS: A mailed, self-report survey measuring multiple dimensions was conducted. Identified through the New Hampshire Division of Workers' Compensation First Report of Injury database, a sample of workers with injuries to their lower back (60%) or upper extremities (40%) a year prior to the study were surveyed. Response rate was 80% (N=169; upper extremity cases=70; low back cases=99). RESULTS: Most (82.8%) were working one year post-injury. Over half reported residual effects of the injury on work or activities of daily living. Many working subjects reported persistent injury-related anxiety and pain at the end of the work day, worse in those with low back pain compared to those with upper extremity injuries. Almost 40% of those who returned to work suffered a reinjury. Forty-four percent of respondents suffered significant injury-related financial problems, which were worse in those who had been out of work for longer periods. CONCLUSIONS: Occupational musculoskeletal injuries do result in significant, long term adverse physical, economic, and psychological consequences, as demonstrated in self-reported surveys. PMID- 10706753 TI - Occupational disability related to back pain: application of a theoretical model of work disability using prospective cohorts of manual workers. AB - BACKGROUND: A new model of work disability was developed based on the assumption that four different groups of workers are present at the beginning of a prospective epidemiologic study: one group of workers without back pain, and three groups of workers with back pain and a gradient of work disability. The goal of this research was to verify if these groups comprise workers at different levels of risk of occurrence of complete work disability related to back injury. METHODS: Prospective cohorts of manual workers (n=578) were followed for 1 year to document the risk of occurrence of complete disability related to back injury. RESULTS: The results showed that the workers who presented with back pain without work disability at the beginning of the study were at less risk compared to all the other workers in the cohort. Moreover, an effect modification was found between the workers who initially presented with back pain without work disability and a past history of compensation for back injury, adding credence to the non-similarity of these workers to the others. CONCLUSIONS: Based on these results, further studies should focus on improving the knowledge of the characteristics of these workers leading to a better understanding of how to prevent occupational low-back pain. PMID- 10706754 TI - Reliability of physical examination of the upper extremity among keyboard operators. AB - BACKGROUND: Physical examination is a traditional outcome measure in epidemiological research. Its value as a reliable measure depends, in part, on the prevalence of positive findings. The purpose of this paper is to determine the empirical reliability of physical examination and anthropometry in a field study of upper extremity disorders among keyboard operators. METHODS: Two experienced examiners independently performed common provocative tests and procedures in physical examinations of the neck and upper extremity among 160 keyboard operators. Two additional examiners conducted anthropometric surveys among 137 workers. Inter-examiner reliability was assessed with observed agreement, kappa statistics, and intra-class correlations (ICC). RESULTS: Observed agreement was between 96% and 100% for neck and upper extremity signs, muscle stretch reflexes, and muscle strength, however, with the exception of provocative tests, reliability statistics were unstable. Among the provocative tests, Phalen and Tinel tests had modest agreement after adjusting for chance (kappa range: 0.20-0.43). The carpal compression test had the best reliability (kappa=0.60 and kappa=0.67, left and right side, respectively). The ICCs for anthropometry ranged from 0.36-0.91. CONCLUSIONS: Results from the study showed that statistically, except for the carpal compression test, physical examination contributed minimal reliable information. This was attributed mainly to the low prevalence of positive findings, and generally mild nature of upper extremity disorders in this population. The results are the best estimate of what would be found in a field study with experienced examiners. While it may reduce bias, separating physical examination from medical history may contribute to the poor reliability of findings. With a shift toward reliable measures, resources can be allocated to more effective tools, like questionnaires, in epidemiological research of upper extremity disorders among keyboard operators. PMID- 10706755 TI - Too hot to handle: an unusual exposure of HDI in specialty painters. AB - BACKGROUND: Hexamethylene Diisocyanate (HDI) is a color stable aliphatic isocyanate that is used in specialty paints as a hardener. Due to the lower vapor pressure of its commercial biuret form, it is considered a relatively "safe" isocyanate from an exposure standpoint. This case series reports on an unusual toxic exposure to HDI. Between November 1993 and May 1994, seven specialty painters and one boiler maker who were working at three different power plants were examined at the Institute of Occupational and Environmental Health at West Virginia University. At their respective work sites, HDI was applied to the hot surfaces of boilers that were not shut down, and allowed sufficient time to cool. Consequently, these workers were exposed to volatile HDI and its thermal decomposition products. METHODS: All of these workers underwent a complete physical examination, spirometry, and methacholine challenge testing. RESULTS: All 8 workers complained of dyspnea, while 4 of the 8 also complained of rash. On examination 3 workers were methacholine challenge positive and 2 had persistent rash. At follow-up 4 years later, 5 workers still had to use inhalation medication and one had progressive asthma and dermatitis. All 8 workers, by the time of the follow-up, had gone through economic and occupational changes. CONCLUSIONS: This case series reports on an unusual exposure to HDI. It is unusual in that: 1) There were two simultaneous sentinel cases with two different Material Safety Data Sheets (MSDS) for the same product, 2) Exposure was to volatile HDI and its decomposition products and 3) Hazardous conditions of exposure occurred at three different sites. PMID- 10706756 TI - Two year follow-up of a garbage collector with allergic bronchopulmonary aspergillosis (ABPA). AB - BACKGROUND: Separate collection of biodegradable garbage and recyclable waste is expected to become mandatory in some western countries. A growing number of persons engaged in garbage collection and separation might become endangered by high loads of bacteria and fungi. Case history and examination A 29 year old garbage collector involved in emptying so-called biological garbage complained of dyspnea, fever, and flu-like symptoms during work beginning in the summer of 1992. Chest x-ray showed streaky shadows near both hili reaching into the upper regions. IgE- and IgG-antibodies (CAP, Pharmacia, Sweden) were strongly positive for Aspergillus fumigatus with 90.5 kU/L and 186%, respectively. Total-IgE was also strongly elevated with 5430 kU/L. Bronchial challenge testing with commercially available Aspergillus fumigatus extract resulted in an immediate type asthmatic reaction. Two years later he was still symptomatic and antibodies persisted at lower levels. CONCLUSIONS: Our diagnosis was allergic bronchopulmonary aspergillosis (ABPA) including asthmatic responses as well as hypersensitivity pneumonitis (extrinsic allergic alveolitis) due to exposure to moldy household waste. A growing number of persons engaged in garbage collection and handling are exposed and at risk to develop sensitization to fungi due to exposure to dust of biodegradable waste. Further studies are necessary to show if separate collection of biodegradable waste increases the health risks due to exposure to bacteria and fungi in comparison to waste collection without separation. PMID- 10706757 TI - Occipital lobe meningioma in a patient with multiple chemical sensitivities. AB - BACKGROUND: The concurrent diagnosis of meningioma with increased intracranial pressure has not been reported previously in a patient who meets diagnostic criteria for multiple chemical sensitivities (MCS). METHODS: A patient who had been evaluated in an occupational medicine practice, and by several other physicians for sensitivity to chemical odors was found to have papilledema and a visual field deficit. The patient met the clinical criteria set forth by Cullen in 1987 for MCS. A magnetic resonance imaging (MRI) scan was performed. RESULTS: The MRI revealed a large occipital lobe meningioma, which was surgically resected. Removal of the meningioma had little effect on the patient's symptoms. She has been unable to return to her job as a custodian. DISCUSSION: The etiology of MCS has been disputed and is currently unresolved. Those who evaluate patients with MCS are reminded that meningiomas and other intracranial mass lesions can affect olfaction, and that patients with MCS can have treatable intracranial abnormalities. PMID- 10706758 TI - A novel mutation that leads to a congenital factor XI deficiency in a Japanese family. AB - We have identified a novel mutation leading to a congenital deficiency of the coagulation factor XI (FXI) in a Japanese family. A propositus was a 42-year-old female patient without bleeding tendency. Coagulant activity and the antigen level of FXI in her plasma were below the detectable range. The nucleotide sequences of the FXI gene of this patient were determined by a direct sequence method established in this study. A novel nonsense mutation (CAA; Gly263 --> TAA; stop) was identified in exon 8 of the FXI gene. Her parents are first cousins, and a polymerase chain reaction-restriction-fragment length polymorphism analysis revealed that her parents were heterozygous at this nucleotide position. This patient inherited mutant alleles from her parents and is homozygous at this nucleotide position. The nonsense mutation in the FXI gene is responsible for her deficiency of FXI. PMID- 10706759 TI - Mutation analysis of PTEN/MMAC1 in acute myeloid leukemia. AB - Recently, a putative tumor suppressor gene, PTEN/MMAC1, has been identified at chromosome 10q23.3, which encodes a 403 amino acid dual-specificity phosphatase containing a region of homology to tensin and auxillin. Somatic mutations of the PTEN/MMAC1 gene have been identified in a number of cancer cell lines and primary cancers. Mutations in PTEN/MMAC1 are most frequently found in advanced cancers. To evaluate the role of the PTEN/MMAC1 gene in leukemia, bone marrow and/or peripheral blood from 62 acute myeloid leukemia (AML) patients, 5 hemopoietic cell lines (HL60, U937, Raji, KG-1, K562), and 30 normal controls were analyzed. The results showed aberrant PTEN/MMAC1 transcripts in 15 of the 62 (24%) AML patients, 4 of the 5 cell lines (80%), and 4 of the 30 (13%) normal controls. As in our previous study of TSG101, the abnormal transcripts may result from aberrant RNA splicing as evidenced by the presence of both these aberrant transcripts and normal full length transcripts in all specimens examined. Loss of heterozygosity (LOH) analysis and PCR-SSCP of the entire coding region showed that none of the AML cases had LOH or mutation. Only one frameshift mutation at codon 130 (insertion of CCCG) with premature termination of coding sequence was observed in the U937 cell line. Our results indicate that the PTEN/MMAC1 gene may play a role in a small percentage of AML, but its significance needs to be further evaluated. PMID- 10706760 TI - Enhancement of hemoglobin and F-cell production by targeting growth inhibition and differentiation of K562 cells with ribonucleotide reductase inhibitors (didox and trimidox) in combination with streptozotocin. AB - Upon appropriate drug treatment, the human erythroleukemic K562 cells have been shown to produce hemoglobin and F-cells. Fetal hemoglobin (Hb F) inhibits the polymerization events of sickle hemoglobin (Hb S), thereby ameliorating the clinical symptoms of sickle cell disease. Ribonucleotide reductase inhibitors (RRIs) have been shown to inhibit the growth of myeloid leukemia cells leading to the production of Hb F upon differentiation. Of the RRIs currently in use, hydroxyurea is the most effective agent for Hb F induction. We have examined the capacity of two novel RRIs, didox (DI) and trimidox (TRI), in combination with streptozotocin (STZ), to induce hemoglobin and F-cell production. The K562 cells were cultured with different concentrations of didox-STZ or trimidox-STZ at a fixed molar ratio of 3:1 and 1:5 for 96 hr, respectively. At pre-determined time intervals, aliquots of cells were obtained and total hemoglobin (benzidine positive) levels, number of F-cells, and Hb F were determined by the differential staining technique, fetal hemoglobin assay kit, and fluorescence cytometry respectively. The effect of combined drug treatment on the growth of K562 cells was examined by isobologram analysis. Our results indicate that a synergistic growth-inhibitory differentiation effect occurred when didox or trimidox was used in combination with STZ on K562 cells. There was an increase in the number of both benzidine-positive normoblasts and F-cells, accompanied by morphologic appearances typical of erythroid maturation. On day 4, the number of benzidine positive cells showed a 6-9-fold increase and the number of F-cells was between 2.5- and 5.7-fold higher than the respective controls. Based upon these results, treatment with a ribonucleotide reductase inhibitor, such as didox or trimidox, in combination with STZ, might offer an additional promising option in sickle cell disease therapy. PMID- 10706761 TI - A polymorphism of the X-linked gene IDS increases the number of females informative for transcriptional clonality assays. AB - Studies of clonality have been essential for understanding the hierarchy of hematopoiesis and the biology of malignancies. Most clonality assays are based on the X chromosome inactivation phenomenon in females; these assays detect protein polymorphisms, differences in DNA methylation, or transcripts of the active X chromosome. Assays based on protein polymorphisms or DNA methylation have significant shortcomings. The major disadvantage of transcriptional assays is their limited applicability since only approximately half of females are informative for these studies. We have developed a new transcriptional assay based on an exonic polymorphism of the X-chromosome gene IDS. This gene is located in the same X-chromosome region (Xq28) as G6PD and p55, two genes with exonic polymorphisms for which we previously developed transcriptional assays. We developed non-radioactive PCR-based assays for rapid screening of genotype and determination of clonality. We also report reaction conditions for a quantitative ligase detection assay of IDS allelic transcripts. The frequency of the IDS polymorphism is 46% in Caucasian females and 39% in African-American females; in combination with G6PD and p55, 76% of Caucasian females and 62% of African American females are informative for these assays. While this gene is highly polymorphic in Caucasian and African-American females, it is not informative in Oriental females. We established that the IDS gene is in linkage equilibrium with G6PD and p55. Unlike methylation-based assays, this assay is suitable for studying clonality in non-nucleated cells such as platelets and reticulocytes. With the discovery of exonic polymorphisms of other X-chromosome genes, all females should eventually be suitable for X-chromosome transcriptional clonality analysis. PMID- 10706762 TI - Bleeding risk factors in chronic oral anticoagulation with acenocoumarol. AB - We studied major bleeding complications, death related to hemorrhage, and tried to identify predisposing factors for bleeding in outpatients treated with acenocoumarol. We evaluated 811 outpatients attending a specialized anticoagulant therapy unit. The intended INR range was 3.5-4.5 for mechanical heart valve replacement (N= 384) and 2.0-3.0 for other indications (N= 427). The variability of INR for the total follow-up and the 2 months before the hemorrhage was calculated. The total follow-up was 1,963.26 years with 27,321 control tests. We observed 47 major bleeding episodes, including 2 fatal (central nervous system hemorrhages), in 37 patients. 49.5% of the patients had underlying diseases. The rate of major and fatal hemorrhage was 2.39 and 0.10 episodes per 100 patients year, respectively. Hemorrhagic complications were more frequently observed in patients with a more intense intended range (8.2% in the INR 3.5-4.5 group vs. 1.5% in the 2.0-3.0 INR group). The risk of major bleeding increased in patients with an achieved INR higher than 6 and in those with higher INR variability during follow-up. The estimated probability of bleeding also increased with time: it was 0.102% at 78 months, and at the beginning of therapy it was 0.006% and 0.007% at 1 and 4 months, respectively. The intensity of anticoagulation and the deviation of the INR from the target are the most important risk factors for bleeding in patients taking acenocoumarol. Monitoring the variability of INR can help identifying patients predisposed to bleeding. However, the screening for underlying disease should always be performed. PMID- 10706763 TI - Type 2B von Willebrand's disease in thirteen individuals from five unrelated Australian families: phenotype and genotype correlations. AB - Type 2B von Willebrand's disease (VWD) is due to a qualitative defect in von Willebrand factor (VWF) in which there is an increased affinity for the platelet glycoprotein Ib-IX-V receptor complex. Spontaneous binding of type 2B VWF to platelets and subsequent clearance from the plasma is thought to account for the characteristic phenotype of type 2B VWD. These gain-of-function mutations are due to single amino substitutions that are clustered within the functionally important A1 domain of VWF. We describe 13 individuals from five unrelated families in Australia with type 2B VWD, report their phenotypic abnormalities, and delineate their causative mutations. We confirm that the mutation Arg543Trp is also particularly common among families with type 2B VWD in Australia. PMID- 10706764 TI - Nitric oxide attenuates normal and sickle red blood cell adherence to pulmonary endothelium. AB - Increased adherence of sickle red blood cells (RBC) to endothelium is implicated as an initiating event of vaso-occlusion in sickle cell disease. Although much is known about the humoral influences of this interaction, there has been little investigation regarding endothelial contributions. Endothelial derived nitric oxide (NO) inhibits adhesion of platelets and leukocytes to endothelium and decreases expression of VCAM-1, an endothelial adhesion site implicated in sickle RBC/endothelial adherence. However, whether NO inhibits RBC adherence to endothelium is unexplored. We tested this hypothesis with endothelial monolayers exposed to RBC from normal (Hb AA) and sickle cell (Hb SS) volunteers in a parallel plate flow chamber. To decrease NO production, endothelial monolayers were exposed to 100 microM nitro-L-arginine (NLA), an inhibitor of nitric oxide synthase, resulting in an 87% increase in normal RBC adherence (P = 0.002). Because adherence of normal RBC to endothelium was low, the effect of DETA-NO, an NO donor, was tested after activation of endothelium with TNF-alpha increased adherence by 130% (P < 0.001). Subsequent addition of 2 mM DETA-NO produced a 75% decrease in adherence of normal RBC to endothelium (P = 0.03). At baseline, sickle RBC were significantly more adherent than normal RBC (P < 0.001) and DETA NO decreased sickle RBC adherence by 54% (P = 0.04). Thus, NO inhibits both normal and sickle RBC adherence to endothelium. Strategies that enhance NO activity may be therapeutic in sickle cell disease. PMID- 10706765 TI - Renal abnormalities in sickle cell disease. AB - Sickle cell anemia and the related hemoglobinopathies are associated with a large spectrum of renal abnormalities. The patients have impaired urinary concentrating ability, defects in urinary acidification and potassium excretion, and supranormal proximal tubular function. The latter is manifest by increased secretion of creatinine and by reabsorption of phosphorus and beta(2) microglobulin. Young patients with sickle cell disease (SCD) have supranormal renal hemodynamics with elevations in both effective renal plasma flow (ERPF) and glomerular filtration rate (GFR). These parameters decrease with age as well as following the administration of prostaglandin inhibitors. Proteinuria, a common finding in adults with sickle cell disease, may progress to the nephrotic syndrome. Proteinuria, hypertension, and increasing anemia predict end-stage renal disease (ESRD). While ESRD can be managed by dialysis and/or renal transplantation, there may be an increased rate of complications in renal transplant recipients with SCD. Hematuria is seen in individuals with all of the SCDs as well as with sickle cell trait. In most cases the etiology of the hematuria turns out to be benign. However, there does appear to be an increased association between SCD and renal medullary carcinoma. Therefore, those SCD patients who present with hematuria should initially undergo a thorough evaluation in order to exclude this aggressive neoplasm. Papillary necrosis may occur due to medullary ischemia and infarction. Erythropoietin levels are usually lower than expected for their degree of anemia and decrease further as renal function deteriorates. An abnormal balance of renal prostaglandins may be responsible for some of the changes in sickle cell nephropathy. Acute renal failure is a component of the acute multiorgan failure syndrome (MOFS). Finally, progression of sickle cell nephropathy to ESRD may be slowed by adequate control of hypertension and proteinuria. However, the prevention of the renal complications of SCD will require a cure for this genetic disorder. PMID- 10706766 TI - Myelodysplastic syndromes in children. A critical review of the clinical manifestations and management. AB - The FAB group has defined myelodysplasia in adults but direct application of this categorization to children has been controversial. Consequently, to outline the natural history of the disease better we have retrospectively analysed case reports and series published in English between 1982 and 1996. This study also included children with juvenile chronic myelomonocytic leukaemia (JCML) and monosomy 7 (Mo7). 340 patients were described in 27 publications. The mean presentation age was 5.91 (SD 5.04) years, and 34.9% were female. Constitutional alterations were described in 68 (20%) where refractory anemia (RA) and RA with excess of blasts (RAEB) predominated and were associated with a significantly longer survival. Among all patients progression to higher forms of MDS was noted in 61 (18%). Cytogenetic anomalies were detected in 59% of 227 children, and in 67 it was to Mo7. Amid those with Mo7, the clinical and laboratory characteristics as well as survival, closely followed their FAB type. Of the treatment options described, survival was significantly higher in those who underwent bone marrow transplant (BMT) (46.9%; P = 0.00021). Among children with JMML (CMML/JCML) not receiving a BMT, time to death was shortest in those best described as JCML (absence of constitutional and karyotypic derangement, thrombocytopenia and elevated Hb F). We conclude that children with constitutional abnormalities survive longer, Mo7 disorders are clinically and morphologically heterogeneous and should not be grouped into a single entity and that CMML and JCML may have biological differences. Finally, BMT remains the treatment of choice for those with primary MDS, as intensive chemotherapy is no better than supportive measures. PMID- 10706767 TI - A rare mutation [IVS-I-130 (G-A)] in a Turkish beta-thalassemia major patient. AB - Here we describe the identification of the rare beta-thalassemia mutation IVS-I 130 (G-A) for the first time in Turkey. The hematological evaluation of the patient showed classical signs of beta-thalassemia major requiring regular blood transfusions every 30-35 days. DNA analysis was carried out using reverse dot blot hybridization and restriction endonuclease digestion, as well as genomic sequencing. The patient was found to be heterozygous for the IVS-I-6 (T-C) and IVS-I-130 (G-A) mutations. In order to deduce a possible origin for the IVS-I-130 (G-A) mutation, the sequence polymorphisms in the DNA of the patient and her family were characterized. The method included the analysis of nine polymorphic nucleotides and the hypervariable microsatellite of composite sequence (AT)(x)T(y) 5' to the beta-globin gene by DNA sequencing. The sequence haplotype (HT4) carrying the IVS-I-130 (G-A) mutation is also observed in Algeria. This favors a Northeastern African origin for this allele. The observed results agree well with a recent introduction of this mutation to Turkey from Egypt toward the end of the 19th century. PMID- 10706768 TI - Expression of high amounts of the CD117 molecule in a case of B-cell non Hodgkin's lymphoma carrying the t(14:18) translocation. AB - The c-kit proto-oncogen (CD117) has been described to be present in normal and neoplastic hemopoietic cells including both myeloid and lymphoid lineages. Among the normal lymphoid cells CD117 expression would be restricted to a small subset of NK-cells, and to early T-cell precursors and it is not expressed by normal B cells. Regarding chronic lymphoproliferative disorders the only data provided up to now suggests that CD117 expression is restricted to cases of Hodgkin's disease and anaplastic large-cell lymphoma. In the present paper we describe a case of a B-cell chronic lymphoproliferative disorder carrying the t(14:18) translocation as demonstrated by molecular studies, in which the flow cytometric immunophenotypic analysis of both peripheral blood and bone marrow samples revealed the expression of high amounts of the CD117 antigen in the surface of the clonal B-cell population. Further studies are necessary to explore both the functional role of c-kit expression in the neoplastic B-cells from this patient and its potential utility for the diagnosis and follow-up of patients with B-cell non-Hodgkin's lymphoma. PMID- 10706769 TI - beta-thalassemia trait might increase the severity of hemochromatosis in subjects with the C282Y mutation in the HFE gene. PMID- 10706770 TI - Successful autologous stem cell transplantation in aggressive prolymphocytic leukemia. PMID- 10706771 TI - Diffuse large-cell B-cell lymphoma in a pituitary adenoma: an unusual cause of pituitary apoplexy. PMID- 10706772 TI - Sarcoidosis presenting as massive splenomegaly and bicytopenia. PMID- 10706773 TI - Man to trypanosome: the tubulin tyrosination/detyrosination cycle revisited. PMID- 10706774 TI - Role of lysophosphatidic acid and rho in glioma cell motility. AB - We have studied the effects of the bioactive phospholipid lysophosphatidic acid (LPA) on cell lines derived from highly invasive human glioblastoma multiforme (GBM). Using transwell migration assays, we show that LPA stimulates both chemokinetic and chemotactic migration of glioma cells. Blood brain barrier breakdown and leakage of serum components that most likely include LPA are common features of GBM. Therefore, the effects of LPA on glioma cell motility are intriguing given the fact that, in vivo, GBM cells often migrate great distances from the main tumor, rendering successful therapy extremely difficult. We show here that LPA initiates a variety of signaling cascades in glioma cells. LPA enhanced transwell migration was sensitive to pertussis toxin (PTX) treatment suggesting an important role for G(i) subtype of G proteins. LPA also stimulated Ca(2+) fluctuations and activation of extracellular signal-regulated kinases (ERKS) 1 and 2, although blocking either pathway had little effect on glioma cell migration. Exposure of glioma cells to LPA resulted in phosphorylation of the regulatory light chain (RLC) of myosin II and the formation of stress fibers and focal adhesions. These effects were blocked by Y-27632, an inhibitor of Rho activated ROCK kinases. Time-lapse video microscopy revealed that Y-27632 treatment caused cells to assume long thin morphologies that suggested deficiencies in the contractile apparatus. Furthermore, many cells exhibited a conspicuous extension of processes when Rho/ROCK kinase cascades were inhibited. The above results suggest that LPA/Rho signaling cascades play important roles in glioma cell motility and that exposure of tumor cells to LPA in vivo may contribute to their invasive phenotype. PMID- 10706775 TI - Identification of a 195 kDa protein in the striated rootlet: its expression in ciliated and ciliogenic cells. AB - Little is known about the molecular composition of the ciliary rootlet. We raised monoclonal antibodies to a crude preparation of striated rootlets isolated from the human oviduct, and obtained a clone (R4109) that specifically labeled the ciliary rootlets. Rootlets associated with the solitary cilium in secretory cells and fibroblasts were also labeled. R4109 identified a 195-kDa protein by immunoblotting. Ciliogenic cells in the oviduct epithelium of young mice were labeled in the globular and/or granular pattern by R4109 by immunofluorescence microscopy. Immunoelectron microscopy showed that they corresponded to fibrogranular complex and dense granule, respectively. The result demonstrated that the 195-kDa protein is a component common to the striated rootlet and dense granule, and thus suggested that dense granules are involved in the rootlet formation. PMID- 10706776 TI - Terminal regions of mouse nebulin: sequence analysis and complementary localization with N-RAP. AB - The regions of mouse nebulin extending from the ends of the super repeats to the C-terminus and N-terminus were cloned and sequenced. Comparison of the mouse sequence with the previously published human sequence shows that the terminal regions of nebulin are highly conserved. The four phosphorylation motifs and SH3 domain found at the C-terminus of mouse nebulin are identical to those found in human nebulin, with the exception of four conservative substitutions. The modules linking this C-terminal region to the super repeats have deletions relative to both fetal and adult human nebulins that correspond to integral numbers of modules, making the mouse C-terminal simple repeat region among the shortest observed to date. The N-terminal region and the C-terminal modules were expressed in Escherichia coli and used for antibody production. Immunofluorescent labeling of these regions of nebulin in isolated myofibrils demonstrates that they are located near the center of the sarcomere and near the Z-line, respectively. Immunogold labeling with antibodies raised against the N-terminal nebulin sequence localizes this region in the A-band near the tips of the thin filaments. Nebulin localization is complementary to that of N-RAP, another muscle-specific protein containing nebulin-like super repeats; nebulin is exclusively found in the sarcomeres, while N-RAP is confined to the terminal bundles of actin filaments at the myotendinous junction. Cell Motil. Cytoskeleton 3:211-222, 2000 Published 2000 Wiley-Liss, Inc. PMID- 10706777 TI - Disruption of microtubules in living cells by tyrphostin AG-1714. AB - Tyrphostin AG-1714 and several related molecules with the general structure of nitro-benzene malononitrile (BMN) disrupt microtubules in a large variety of cultured cells. This process can be inhibited by the stabilization of microtubules with taxol or by pretreatment of the cells with pervanadate, which inhibits tyrosine phosphatases and increases the overall levels of phosphotyrosine in cells. Unlike other microtubule-disrupting drugs such as nocodazole or colchicine, tyrphostin AG-1714 does not interfere with microtubule polymerization or stability in vitro, suggesting that the effect of this tyrphostin on microtubules is indirect. These results imply an involvement of protein tyrosine phosphorylation in the regulation of overall microtubule dynamics. Tyrphostins of AG-1714 type could thus be powerful tools for the identification of such microtubule regulatory pathways. PMID- 10706778 TI - Rpg1p, the subunit of the Saccharomyces cerevisiae eIF3 core complex, is a microtubule-interacting protein. AB - The essential gene RPG1/TIF32 of Saccharomyces cerevisiae encodes the 110-kDa subunit of the translation initiation factor 3 (eIF3) core complex. In this study, the Rpg1p-specific monoclonal antibody PK1/1 was used to analyse the cellular distribution of Rpg1p by epifluorescence and confocal laser scanning microscopy (CLSM). In budded cells, a portion of Rpg1p was obviously co-localised with microtubules. In addition, CLSM revealed an accumulation of Rpg1p in a patch at the very end of cytoplasmic microtubules reaching the bud tip. A punctate fluorescence pattern was typical for separated unbudded cells. Distribution of Rpg1p was confirmed using a strain expressing exclusively a hemaglutinin-tagged version of Rpg1p. In nocodazole-treated cells, the pattern of the PK1/1 staining was disturbed. No staining was observed in Rpg1p-depleted cells. In vitro experiments revealed that Rpg1p was specifically co-immunoprecipitated with alpha tubulin from the yeast cell free extract and this observation was further supported by showing that Rpg1p co-sedimented with hog brain microtubules. We conclude that Rpg1p is a microtubule-interacting protein that indicates an interesting connection between the translation initiation machinery and cytoskeleton in yeast Saccharomyces cerevisiae. PMID- 10706779 TI - Erratum: volume 45, number 1, january 2000 AB - The legend to the issue's cover figure was incorrectly printed. It should have read: Cover micrograph: Actin-comet tails induced by the bacterial pathogen Listeria monocytogenes. Vero cells were infected by Listeria monoctyogenes. After fixation, cells were labeled with FITC-phalloidin (green) and anti-listeria antibodies (red). Photo courtesy of Inigo Lasa and Pascale Cossart. PMID- 10706780 TI - Determination of harmane and harmine in human blood using reversed-phased high performance liquid chromatography and fluorescence detection. AB - A number of tremorogenic beta-carboline alkaloids have been found in common plant derived foodstuffs, beverages, and inhaled substances. Because of their natural presence in the food chain, there is a growing concern regarding the potential risks of certain essential tremors associated with the long-term, low-level dietary exposure to these alkaloids. The purpose of this study was to develop an effective analytical method to determine blood levels of two major beta-carboline derivatives, harmane and harmine. Human blood was extracted with ethyl acetate and methyl-t-butyl ether (2:98) under an alkaline condition. After evaporation of organic solvent, the samples were reconstructed in methanol. The samples were fractionated on a 250 x 4.6-mm C18 reversed-phase column with an isocratic mobile system consisting of 17.5 mM potassium phosphate buffer (ph 6.5) and methanol (30:70), followed by an on-line fluorescence detection. The method had the detection limit to determine 206 and 81 pg/ml of harmane and harmine, respectively, in 10 ml of human blood. The intraday precision (C.V.) at 25 ng/ml was less than 6.7 and 3.4% for harmane and harmine, respectively. The interday precision was 7.3% for harmane and 5.4% for harmine. The method has proven sensitive, reproducible, and thus useful for both laboratory and clinical studies of beta-carboline toxicities. PMID- 10706781 TI - Measurement of plasma unbound unconjugated bilirubin. AB - A method is described for measuring the unconjugated fraction of the unbound bilirubin concentration in plasma by combining the peroxidase method for determining unbound bilirubin with a diazo method for measuring conjugated and unconjugated bilirubin. The accuracy of the unbound bilirubin determination is improved by decreasing sample dilution, eliminating interference by conjugated bilirubin, monitoring changes in bilirubin concentration using diazo derivatives, and correcting for rate-limiting dissociation of bilirubin from albumin. The unbound unconjugated bilirubin concentration by the combined method in plasma from 20 jaundiced newborns was significantly greater than and poorly correlated with the unbound bilirubin determined by the existing peroxidase method (r = 0.7), possibly due to differences in sample dilution between the methods. The unbound unconjugated bilirubin was an unpredictable fraction of the unbound bilirubin in plasma samples from patients with similar total bilirubin concentrations but varying levels of conjugated bilirubin. A bilirubin-binding competitor was readily detected at a sample dilution typically used for the combined test but not at the dilution used for the existing peroxidase method. The combined method is ideally suited to measuring unbound unconjugated bilirubin in jaundiced human newborns or animal models of kernicterus. PMID- 10706782 TI - An unambiguous microassay of galactofuranose residues in glycoconjugates using mild methanolysis and high pH anion-exchange chromatography. AB - An original, unambiguous microassay of galactofuranose (Galf) residues in glycoconjugates is described. The method involves mild acid methanolysis (5 mM HCl) for 3 h at 84 degrees C followed by high pH anion-exchange chromatography using a routine monosaccharide system. The methanolysis products Mealpha-Galf and Mebeta-Galf were characterized chromatographically by comparison with the authentic compounds and by their response to treatment with mild acid and with beta-galactofuranosidase. Testing against p-nitrophenyl-beta-Galf and UDPalpha Galf showed the method to be applicable to both alpha- and beta- galactofuranosides over the range 10-200 pmol. The results of partial mild methanolysis over shorter periods were consistent with initial inversion of anomeric configuration at methylation followed by anomerization to an equilibrium mixture of alpha- and beta-forms. When applied to a sample of invertase from Aspergillus nidulans, the method indicated that all of the mild acid-labile galactose (78% of the total galactose present) was in the form of a galactofuranoside and that much of this was in the beta-configuration. As expected, when applied to asialofetuin (known to contain galactose only in the pyranoside form, Galp), NPalpha-Galp, NPbeta-Galp, or UDPalpha-Galp, mild acid methanolysis failed to produce any galactofuranoside. PMID- 10706783 TI - Fluorescence properties of rhodamine 800 in whole blood and plasma. AB - We have characterized the fluorescence spectral properties of rhodamine 800 (Rh800) in plasma and blood in order to test the possibility of making clinical fluorescence measurements in whole blood without separation steps. Rh800 was used because of its absorption at red/near-infrared wavelengths away from the absorption bands of hemoglobin. We utilized the front-face illumination and detection to minimize the effects of absorption and/or scatter during measurements. The presence of Rh800 was detected in plasma and blood using steady state fluorescence measurements. Absorption due to hemoglobin reduced the Rh800 intensity from the blood. Fluorescence lifetime measurements in plasma and blood showed that it is possible to recover lifetime parameters of Rh800 in these media. We obtained mean lifetimes of 1.90 and 1.86 ns for Rh800 in plasma and blood, respectively. Using the recently described modulation sensing method, we quantified the concentrations of Rh800 in plasma and blood. Rh800 was detected at a concentration of as low as 2 microM in both media. High anisotropy values were obtained for Rh800 in plasma and blood using steady-state and anisotropy decay measurements, implying the tight binding of this probe to the contents of these media. This binding can be exploited to monitor the concentrations of different blood components using already existing or new red-emitting probes that will be specially designed to bind to these components with high specificity. To test this possibility of direct measurements in blood, we used Rh800 to monitor albumin in the presence of red blood cells. Increase in the polarization of Rh800 as the concentration of albumin was increased in the presence of the red cells showed the feasibility of such measurements. PMID- 10706784 TI - A method for directly fitting the time derivative of sedimentation velocity data and an alternative algorithm for calculating sedimentation coefficient distribution functions. AB - The time-derivative method for deriving the sedimentation coefficient distribution, g(s*), from sedimentation velocity data that was developed by Walter Stafford has many advantages and is now widely used. By fitting Gaussian functions to the g(s*) distribution both sedimentation and diffusion coefficients (and therefore molecular masses) for individual species can be obtained. However, some of the approximations used in these procedures limit the accuracy of the results. An alternative approach is proposed in which the dc/dt data are fitted rather than g(s*). This new approach gives improved accuracy, extends the range to sedimentation coefficients below 1 S, and enhances resolution of multiple species. For both approaches the peaks from individual species are broadened when the data cover too wide a time span, and this effect is explored and quantified. An alternative algorithm for calculating g(s*) from the dc/dt curves is presented and discussed. Rather than first averaging the dc/dt data for individual scan pairs and then calculating g(s*) from that average, the g(s*) distributions are calculated for every scan pair and then subsequently averaged. This alternative procedure yields smaller error bars for g(s*) and somewhat greater accuracy for fitted hydrodynamic properties when the time span becomes large. PMID- 10706785 TI - An improved method for the determination of green and black tea polyphenols in biomatrices by high-performance liquid chromatography with coulometric array detection. AB - Tea polyphenols are strong antioxidants and are believed to have beneficial health effects. However, the blood and tissue levels of these compounds are not well characterized because of a lack of suitable analytical methods for the biological resolution of these compounds. Previously, we developed methods for the analysis of three green tea catechins. Now we report an improved method for the measurement of the levels of the different catechins and theaflavins in biological fluids and tissues. The method includes digestion of the plasma, urine, or tissue samples with beta-d-glucuronidase and sulfatase, followed by extraction with ethyl acetate and subsequent separation by reversed-phase high performance liquid chromatography (HPLC). The polyphenols are identified on the basis of their retention times, spectral analysis, and electrochemical behavior across an array of electrodes. In a single HPLC run, it is possible to determine the major catechins and theaflavins as well as some of the catechin metabolites. The detection limits for catechins and theaflavins are from 5 to 10 ng/ml of saliva, plasma, or urine. PMID- 10706786 TI - A BODIPY fluorescent microplate assay for measuring activity of calpains and other proteases. AB - The use of 4,4-difluoro-5,7-dimethyl-4-bora-3a, 4a-diaza-s-indacene-3-propionic acid (BODIPY-FL) labeled casein in autoquenching assays of proteolytic activity has been recently described, and we have adapted this assay to measurement of calpain activity. BODIPY-FL coupled to casein at a ratio of 8 mol of BODIPY FL/mol of casein or higher produces a BODIPY-FL-casein substrate that can be used in an autoquenching assay of calpain proteolytic activity. This assay has a number of advantages for measuring calpain activity. (1) The procedure does not require precipitation and removal of undegraded protein, so it is much faster than other procedures that require a precipitation step, and it can be used directly in kinetic assays of proteolytic activity. (2) The BODIPY-FL-casein assay is easily adapted to a microtiter plate format, so it can be used to screen large numbers of samples. (3) Casein is an inexpensive and readily available protein substrate that more closely mimics the natural substrates of endoproteinases, such as the calpains, than synthetic peptide substrates do. Casein has K(m) values for micro- and m-calpain that are similar to those of other substrates such as fodrin or MAP2 that may be "natural" substrates for the calpains, and there is no reason to believe that calpain hydrolysis of casein is inherently different from hydrolysis of fodrin or MAP2, which are much less accessible as substrates for protease assays. (4) The BODIPY-FL-casein assay is capable of detecting 10 ng ( approximately 5 nM) of calpain and is nearly as sensitive as the most sensitive calpain assay reported thus far. (5) The BODIPY FL-casein assay is as reproducible as the FITC-casein assay, whose reproducibility is comparable to or better than the reproducibility of other methods used to assay calpain activity. The BODIPY-FL-casein assay is a general assay for proteolytic activity and can be used with any protease that cleaves casein. PMID- 10706787 TI - Electrospray/tandem mass spectrometry for quantitative analysis of lipid remodeling in essential fatty acid deficient mice. AB - A method utilizing electrospray ionization coupled with tandem mass spectrometry was developed as a facile and rapid method to identify and quantify lipid remodeling in vivo. Electrospray/tandem mass spectrometric analyses were performed on lipids isolated from liver tissue and resident peritoneal cells from essential fatty acid sufficient and deficient mice. Essential fatty acid deficiency was chosen as the paradigm to evaluate the methodology because it epitomizes the most extreme dietary means of altering fatty acid composition of virtually all cellular lipid species. Qualitative and quantitative changes were measured in the phospholipid and cholesterol ester species directly in the chloroform/methanol lipid extract without any prior chromatographic separation. Lipid remodeling in liver and peritoneal cells from essential fatty acid deficient mice was qualitatively similar in cholesterol ester, phosphatidylcholine, and phosphatidylethanolamine. The monoenoic fatty acids palmitoleic acid (16:1 n-7) and oleic acid (18:1 n-9) were increased markedly, whereas all n-6 and n-3 polyunsaturated fatty acids were nearly depleted in phospholipid and cholesterol ester species. The n-9 polyunsaturated fatty acid surrogate, Mead acid (20:3 n-9), substituted for arachidonic acid (20:4 n-6) and docosahexaenoic acid (22:6 n-3) in phospholipid, but not in cholesterol ester, species. Another notable difference was that adrenic acid (22:4 n-6) and docosapentaenoic acid (22:5 n-6), both metabolites of arachidonic acid, accumulated in phospholipid and cholesterol ester species of peritoneal cells, but not in liver cells, of essential fatty acid sufficient mice. The overall body of data presented illustrates the implementation of electrospray/tandem mass spectrometry as a method for facile and direct quantification of changes in lipid species during lipid metabolic studies. PMID- 10706788 TI - Soft- and hard-modeling approaches for the determination of stability constants of metal-peptide systems by voltammetry. AB - The Zn(2+)-glutathione system is studied as a model for metal-peptide systems where some critical factors must be considered when using voltammetric techniques for the determination of stability constants. These factors are the presence of side reactions (in this case, both the protonation of glutathione and the hydrolysis of Zn(2+)), the association-dissociation rates of the complexes compared with the time scales of the measurements (which makes the complexes electrochemically labile or inert), and the electron transfer kinetics on the electrode surface (which makes the metal ion reduction reversible or irreversible). For the study of these factors, three data treatment approaches have been applied: (i) the electrochemical hard-modeling approach (modelization of both chemical equilibrium and electrochemical processes), (ii) a chemical hard modeling approach (modelization of chemical equilibria only, based on the least squares curve-fitting program SQUAD), and (iii) a previously developed model-free soft-modeling approach based on multivariate curve resolution with a constrained alternating least-squares optimization. By analyzing differential pulse polarographic data obtained under different experimental conditions, the influence of the mentioned factors on every approach is discussed and, if possible, the corresponding stability constants are computed. The results of this study showed the potential usefulness of voltammetry in combination with hard- and soft-modeling data analysis for the study of peptide complexation equilibria of metal ions such as Zn which have neither relevant spectroscopic properties nor proper isotopes for NMR measurements. PMID- 10706789 TI - Analysis of nitrite/nitrate in biological fluids: denitrification of 2 nitropropane in F344 rats. AB - 2-Nitropropane (2-NP), a rat hepatocarcinogen, is denitrified to nitrite and acetone by rat liver microsomes; the denitrification rate is increased using microsomes from phenobarbital (PB)-pretreated rats. To obtain evidence that denitrification of 2-NP also occurs in vivo, we attempted to determine nitrite and nitrate levels in blood sera and urines of 2-NP-treated (1.5 mmol/kg, ip, once) rats with and without PB pretreatment (80 mg/kg, ip, once daily, 3 days), using enzymatic reduction followed by the standard Griess reaction. However, due to various interfering factors, including pigment from methemoglobinemia, we found the assay had to be modified as follows: (a) reduction of nitrate to nitrite was accomplished using NADPH and nitrate reductase, (b) excess NADPH, proteins, and interfering pigments were precipitated using zinc acetate and Na(2)CO(3), and (c) the Griess reagents were prepared in 3 N HCl rather than 5% H(3)PO(4). With these modifications it became possible to show that 2-NP is indeed metabolized to nitrite in vivo and that the metabolism is increased by PB pretreatment. Two hours after 2-NP administration, rat blood serum nitrate plus nitrite levels were approximately 1600 microM (PB-pretreated) and 940 microM (vehicle-pretreated controls). The PB-pretreated and control rats, respectively, excreted 250 and 120 micromol nitrate/nitrite in the 24-h urine post 2-NP treatment. The modifications described make the method more specific, reproducible, and more widely applicable. PMID- 10706790 TI - Temperature dependence of DNA affinity chromatography of transcription factors. AB - Oligonucleotides bound by the CAAT enhancer binding protein (C/EBP), the lactose repressor, and Gal4 were chemically coupled to cyanogen bromide-activated Sepharose and the temperature dependence of transcription factor chromatography was characterized. Each transcription factor was applied to the appropriate column and eluted using a salt gradient at several temperatures. Each transcription factor showed a unique behavior. As temperature was increases, less salt was required to elute C/EBP, more salt was required to elute lac repressor, while Gal4 showed a biphasic dependency with the amount of salt first decreasing between 4 and 19 degrees C and then increasing above 19 degrees C. This temperature dependence is not due to protein or DNA unfolding but rather is a property of complex formation. By loading a column, washing it at a permissive temperature, and then rapidly changing the column temperature, highly selective elution can be obtained. The thermodynamics of this temperature effect are different for the binding of specific and nonspecific DNA sequences, making chromatography at different temperatures a potentially important way of purifying transcription factors. PMID- 10706791 TI - Ultrasensitive fluorescence-based detection of nascent proteins in gels. AB - The most common method of analysis of proteins synthesized in a cell-free translation system (e.g., nascent proteins) involves the use of radioactive amino acids such as [(35)S]methionine or [(14)C]leucine. We report a sensitive, nonisotopic, fluorescence-based method for the detection of nascent proteins directly in polyacrylamide gels. A fluorescent reporter group is incorporated at the N-terminus of nascent proteins using an Escherichia coli initiator tRNA(fmet) misaminoacylated with methionine modified at the alpha-amino group. In addition to the normal formyl group, we find that the protein translational machinery accepts BODIPY-FL, a relatively small fluorophore with a high fluorescent quantum yield, as an N-terminal modification. Under the optimal conditions, fluorescent bands from nanogram levels of in vitro-produced proteins could be detected directly in gels using a conventional UV-transilluminator. Higher sensitivity ( approximately 100-fold) could be obtained using a laser-based fluorescent gel scanner. The major advantages of this approach include elimination of radioactivity and the rapid detection of the protein bands immediately after electrophoresis without any downstream processing. The ability to rapidly synthesize nascent proteins containing an N-terminal tag facilitates many biotechnological applications including functional analysis of gene products, drug discovery, and mutation screening. PMID- 10706792 TI - Development and validation of an NMR-based identity assay for bacterial polysaccharides. AB - A method utilizing NMR spectroscopy has been developed to confirm the identity of bacterial polysaccharides used to formulate a polyvalent pneumococcal polysaccharide vaccine. The method is based on 600 MHz proton NMR spectra of individual serotype-specific polysaccharides. A portion of the anomeric region of each spectrum (5.89 to 4.64 ppm) is compared to spectra generated for designated reference samples for each polysaccharide of interest. The selected region offers a spectral window that is unique to a given polysaccharide and is sensitive to any structural alteration of the repeating units. The similarity of any two spectral profiles is evaluated using a correlation coefficient (rho), where rho >/= 0.95 between a sample and reference profile indicates a positive identification of the sample polysaccharide. This method has been shown to be extremely selective in its ability to discriminate between serotype-specific polysaccharides, some of which differ by no more than a single glycosidic linkage. Furthermore, the method is rapid and does not require extensive sample manipulations or pretreatments. The method was validated as a qualitative identity assay and will be incorporated into routine quality control testing of polysaccharide powders to be used in preparation of the polyvalent pneumococcal vaccine PNEUMOVAX 23. The specificity and reproducibility of the NMR-based identity assay is superior to the currently used colorimetric assays and can be readily adapted for use with other bacterial polysaccharide preparations as well. PMID- 10706793 TI - Staining and quantification of proteins transferred to polyvinylidene fluoride membranes. PMID- 10706794 TI - Native extraction of phosphotyrosine-containing proteins: requirement of tyrosine kinase inhibitors to obtain specific phosphorylation signals. PMID- 10706795 TI - Purification of free sphingoid bases by solid-phase extraction on weak cation exchanger cartridges. PMID- 10706796 TI - A sensitive assay for phosphoinositide phosphatases. PMID- 10706797 TI - The management and control of hospital-acquired infection in acute NHS trusts in England: a report by the Comptroller and Auditor General--the who, how and what. PMID- 10706798 TI - Methicillin-resistant Staphylococcus aureus typing methods: which should be the international standard? AB - Methicillin-resistant Staphylococcus aureus (MRSA) has spread to all parts of the world. Effective control measures are dependent on a thorough knowledge of the organism's epidemiology which requires a typing technique that can be universally applied. Many typing methods have been developed for MRSA but none has been adopted as the internationally recognized standard. This review summarizes the information available on each in order to assess their suitability as a reference procedure. The majority of phenotypic and genotypic techniques are not sufficiently discriminatory, reproducible, stable or useful in an outbreak to be acceptable. The methods which do fulfil these requirements and have a potential for standardization, such as pulsed-field gel electrophoresis, binary typing or a combination of more rapid techniques, require further systematic evaluation. PMID- 10706799 TI - Epidemiological investigation of Ochrobactrum anthropi strains isolated from a haematology unit. AB - Ochrobactrum anthropi is an oxidase-producing gram-negative bacillus preferring aqueous environments. It is an opportunist of low pathogenicity with a wide and unpredictable antibiotic resistance. We observed bacteraemia caused by this organism in two immunocompromized patients hospitalized in the same haematology unit and catheter-associated sepsis was recognized within two days. Another isolate was obtained from the stools of a third patient of the same unit. Environmental investigations recovered an isolate from a tap-water sample of the unit. Pulsed-field gel electrophoresis analysis of these four isolates and two others isolates previously found in the same ward, showed identical restriction patterns for the two blood isolates and confirmed that the two bacteraemia were epidemiologically related. PMID- 10706800 TI - Iatrogenic GB virus C/hepatitis G virus infection in an area endemic for hepatitis C virus. AB - GB virus C/hepatitis G virus (GBV-C/HGV) is reported to be transmitted by blood products. This study reports infection with GBV-C/HGV from Area-O of town T, an area of high prevalence of antibody to hepatitis C virus (anti-HCV). Four hundred and thirty-five inhabitants of Area-O in town T were examined. Three hundred and forty-three inhabitants of Area-H in town T (where differences of age or sex are not markedly different to Area-O) were studied as controls. We investigated the virus markers and conducted a survey of life history in both areas. The seroprevalence of anti-HCV and GBV-C/HGV markers in Area-O was 17.7% and 11.7%, significantly higher than in Area-H (1.5% and 4.4%). The prevalence of GBV-C/HGV markers was significantly higher in the anti-HCV-positive group than in the sero negative group. Anti-HCV- or GBV-C/HGV positive subjects tended to have a history of intravenous medications at hospital C in town T, suggesting iatrogenic infection through insufficient sterilization of needles and/or syringes. PMID- 10706801 TI - Morbidity associated with central venous catheter-use in a cohort of 212 hospitalized subjects with HIV infection. AB - Technical complications and nosocomial bloodstream infections associated with short-term central venous catheterization remain a heavy burden in terms of morbidity, mortality and cost in HIV-positive subjects. Between 1994 and 1997, 327 central venous catheters (CVCs) inserted in 212 patients for a total of 5005 catheter days were investigated. Forty-two technical complications (13%) occurred in 40 patients. Logistic regression analysis revealed that a high APACHE III score was associated with development of CVC-related complications (P = 0.01). One hundred and eight of 327 CVCs (33%) were suspected as being infected. However only 61 episodes (61/327, 19%) were finally diagnosed as CVC-related sepsis. Three variables affecting the rate of CVC-related sepsis were identified: 1) administration of TPN (P = 0.01); 2) low number of circulating CD4+ cells (P = 0.04); 3) high APACHE III score (P = 0. 04). Doctors responsible for AIDS patients should carefully consider the relative risks and benefits of CVC insertion in an individual patient. PMID- 10706802 TI - Infection control training: evaluation of a computer-assisted learning package. AB - An evaluation of the training module of an interactive infection control computer assisted learning (CAL) software program was carried out with ward-based nurses, third-year medical students and infection control personnel. All nursing staff, 87% of the medical students and all infection control staff found the programme easy and enjoyable to use. The module was accessed 3101 times on the hospital network in 18 months with usage settling to between 100-150 times per month. There was a higher level of use by night-duty and weekend staff. Medical students gained as much infection control knowledge from using the CAL package (increase in correct responses from 63.5% to 83.4%;P<0.0001) as they did from a formal lecture (increase in correct responses from 62.1% to 79.5%;P<0.0001). We conclude the training module which is accessible on the hospital wards and across the academic network, is a convenient and effective way for staff and students to gain a basic understanding in evidence-based infection control practices, at locations and times suitable for them. PMID- 10706803 TI - Device-associated, device-day infection rates in an Israeli adult general intensive care unit. AB - Surveillance is an essential element of hospital infection control programs. Previous studies have shown that interhospital comparison of intensive care unit (ICU) nosocomial infections (NI) may be best made by comparing ICU-type-specific, device-associated infection rates and that these adjusted rates vary by ICU type. The aim of this study was to evaluate whether significant structural improvements introduced in an adult general ICU were associated with changes in the NI rates in this unit. In addition, we compared these rates with those of ICUs reported by the National Nosocomial Infections Surveillance (NNIS) System of the Centers for Diseases Control and Prevention. During a 12-month period 337 patients were surveyed. There were 20 ventilator-associated pneumonias (VAP)/1000 ventilator (VEN)-days, 12 bloodstream infections (BSI)/1000 central vascular catheter (CVC) days and 14 urinary tract infection (UTI)/1000 indwelling urinary catheter (IUC) days. Structural changes and reduction in device utilization ratios were not followed by change in NI rates in this unit. VAP and BSI rates were comparable to those reported for neurosurgical and burn ICUs, respectively, in the NNIS System, despite a much higher device utilization ratios. The present study provides specific surveillance data for further interhospital comparison with similar types of ICUs. PMID- 10706804 TI - A one-year prospective study of nosocomial bacteraemia in ICU and non-ICU patients and its impact on patient outcome. AB - A one-year, prospective, two-observational cohort study was performed to evaluate the incidence and outcome in hospitalized patients (ICU and non-ICU) of nosocomial bacteraemia, and to assess its prognostic value in the ICU group. A group of 18 098 hospitalized patients and a group of 291 consecutive ICU patients were followed. Prognostic factors were determined using single and multivariable analyses. 109 (90 non-ICU and 19 ICU) patients developed 118 nosocomial bacteraemic episodes. The incidence of nosocomial bacteraemia was 6.0 per 1000 admissions (95% confidence interval (CI): 5-7%) and 65 per 1000 admissions in ICU patients (95% CI: 4.5-8.5%). Gram-positive and Gram-negative bacteria were 63/133 (47%) and 70/133 (53%) of the isolated micro-organisms respectively. Crude mortality rates were 41/109 (38%) with adverse outcome associated with mechanical ventilation (OR: 3.6; 95% CI: 1.4-9.2, P =0.01), neutropenia (OR: 7.7; 95% CI: 0.8-73.1;P =0.07) while gastro-intestinal surgery was associated with an improved outcome (OR: 0.4; 95% CI: 0.16-0.96;P =0.04). Of the 291 ICU patients, 19 acquired 22 episodes of nosocomial bacteraemia, and 18 were referred from the wards with documented nosocomial bacteraemia. Of these 37 bacteraemic patients, 17 (46%) died. When adjusting for predictors of death (SAPS II>/=40, cardiac and neurological failure), nosocomial bacteraemia markedly influence the outcome in ICU patients (OR: 3.4; 95% CI: 1.3-8.7;P =0.010). This study suggests that the outcome of nosocomial bacteraemia in hospitalized patients is poor in ventilated and neutropenic patients and that nosocomial bacteraemia per se influenced outcome in ICU patients. PMID- 10706805 TI - A three-year survey of nosocomial and community-acquired infections, antibiotic treatment and re-hospitalization in a Norwegian health region. AB - In Norway, hospital-acquired infections (HAI) were analysed by repeated point prevalence studies (four each year) performed simultaneously at 14 hospitals in a health region (860,000 inhabitants) during the period 1996-1998. The study included 3200 beds and 121,000 discharged patients each year, and was initiated by and co-ordinated from the regional university hospital; Ulleval University Hospital (UHH). An overall prevalence rate of HAI of 6.5% (interhospital variation 1.4-11.7%) was found for the 32,248 patients studied. The rate of HAI was reduced from 7.7% in 1996 to 5. 9% in 1998. Smaller hospitals (<200 beds) generally had lower rates of HAI, community acquired infections (CAI), postoperative infections and use of antibacterial agents, than the large regional hospital (1200 beds). HAI was reduced in non-operated patients from 5.8% in 1996 to 4.4% in 1998 and in operated patients from 13.2% in 1996 to 10.5% in 1998. The risk of developing HAI was twice as high after surgery. From 1996 to 1998 there was a reduction in: urinary tract infections from 2.4% to 1.7%, lower respiratory tract infections from 1.5% to 0.8% and postoperative wound infections from 5.7% to 4.3%, while septicaemia (from 0.5% to 0.4%) remained unchanged. Re hospitalization because of HAI was registered in 0.6% (interhospital variation 0.3-1.1%) of patients. The CAI rate in hospitals increased from 8.3% in 1996 to 10.8% in 1998. Approximately 16% (variation:14.4-20.6%) of the patients had an infection. The total use of antibacterial agents was 19.2% in 1996, 16.6% in 1997 and 17.8% in 1998 (variation: 14.9-23%). PMID- 10706806 TI - Variation in risk for nosocomial chickenpox after inadvertent exposure. AB - Inadvertent exposure to chickenpox in the healthcare setting results in time consuming and expensive infection control management strategies. In households, secondary cases occur in up to 96% of susceptibles, but transmission risk after exposure in an occupational setting is less well defined. In this prospective cohort study of inadvertent exposures in a 180-bed paediatric hospital, the secondary transmission rate was 4.5% (4/89; 95% confidence interval 1.2, 11.1%). Fourteen index cases exposed 158 patients and 93 healthcare workers over a 36 month study period. Exposures occurred in inpatient and ambulatory settings, with patients, staff and siblings serving as index cases. Transmission only occurred when the index case and contacts were in the same room and not in a multi-room setting (12% v. 0%, Fisher exact test, P = 0.02). Occupational exposures present a lesser transmission risk than those in households. Definition of those exposure variables that increase risk of transmission in the occupational setting should be explored in future studies. PMID- 10706807 TI - The population impact of MRSA in a country: the national survey of MRSA in Wales, 1997. AB - Continuous data collection on all new isolates of MRSA via CoSurv has taken place in Wales since January 1996. In order to audit this data collection, and to address some of the issues that it does not include, a survey of MRSA was carried out. Questionnaires were completed by infection control teams. Rates were calculated using hospital throughput denominators. Results from the one-day prevalence survey, the two-week incidence survey, and the follow-up survey carried out on new MRSA patients identified in the incidence survey, are presented. Results were found to be broadly similar to those collected via routine surveillance. MRSA was found frequently and disproportionately in the elderly, with higher rates in male than female patients. The highest incidence of total and invasive MRSA was in males aged 75 and over (total: 12.5/1000 finished consultant episodes; invasive: 2.8/1000). Although there was a large community reservoir of MRSA, most appeared to have been acquired in hospital, since most patients had a history of hospitalization, often with multiple hospital admissions. Community-based isolates from cases with no hospital history tended to have been from ulcers. Prevalence and incidence of MRSA was relatively low compared with hospital throughput (mean prevalence: 2.4/100 occupied beds; mean incidence: 3.6/1000 finished consultant episodes), there was also quite large variation between sites, even when screening samples were removed. Patients with MRSA had strikingly long stays before isolation of the organism (prevalence survey: 39 days; incidence survey: 31 days) and highest incidence occurred in elderly care wards. The outcome survey showed that approximately half of the patients were treated with some type of antimicrobial therapy for MRSA. Decontamination therapy was associated with clearance of MRSA only when controlling for sex of the patient. The majority of patients were discharged still with MRSA, mostly to their own homes. The survey emphasizes the need to continue surveillance to detect any changes, to allow guidelines based on evidence to be developed and to monitor the effectiveness of such guidelines. PMID- 10706808 TI - Needlestick injuries amongst medical students in Birmingham, UK. PMID- 10706809 TI - Evidence for a self-fulfilling hypothesis: chlorhexidine dressing for reduction of microbial colonization of the skin with central venous catheters. PMID- 10706810 TI - This letter was shown to Dr hanazaki and his reply follows PMID- 10706812 TI - Working Together but in Opposition: An Examination of the "Good-Cop/Bad-Cop" Negotiating Team Tactic. AB - Unlike solo negotiators, members of negotiating teams may for strategic reasons choose to play different roles; the familiar "good cop/bad cop" distributive bargaining tactic is one example of role differentiation designed to enhance a team's success at the bargaining table. In two empirical studies about a hypothetical three-person work group, we examined the cognitive processes underlying this tactic using a social-cognitive decision model (Brodt & Duncan, 1998) that conceptualizes the negotiators' decision tasks and persuasion processes. Results generally supported the model except for an intriguing asymmetry depending on a person's initial inclination (accepting, rejecting). This research extends findings on the tactic and on contrast effects (Cialdini, 1984) and supports the model's usefulness as an approximate representation of negotiator cognition. Copyright 2000 Academic Press. PMID- 10706811 TI - Implementation of HTM2030 for washer-disinfectors. PMID- 10706813 TI - Deciding Whether to Complete or Terminate an Unfinished Project: A Strong Test of the Project Completion Hypothesis. AB - The primary purpose of this research was to subject Conlon and Garland's (1993; Garland & Conlon, 1998) project completion hypothesis to a stronger test than it has faced in the past. We conducted an experiment in which we manipulated the degree of completion, sunk-cost amount, and anticipated sales price for a hypothetical real estate development project. The sales price information allowed participants to calculate the economic value of the project. Participants were also held "accountable" in the sense that they had to explain the reasoning behind their decisions to the experimenter. Together, the sales price information and accountability stipulation created an inducement to engage in an economically rational decision process. In spite of this inducement, participants were unduly affected by the project's closeness to completion. In fact, when the project was close to being finished, they often recommended completing the project even when it was economically unwise to do so. While the effects of the project completion manipulation were surprisingly strong, the sunk-cost manipulation had virtually no effect. Implications of these results and directions for future research are discussed. Copyright 2000 Academic Press. PMID- 10706814 TI - The Binary Additivity of Subjective Probability Does not Indicate the Binary Complementarity of Perceived Certainty. AB - People's numeric probability estimates for 2 mutually exclusive and exhaustive events commonly sum to 1.0, which seems to indicate the full complementarity of subjective certainty in the 2 events (i.e., increases in certainty for one event are accompanied by decreases in certainty for the other). In this article, however, a distinction is made between the additivity of probability estimates and the complementarity of internal perceptions of certainty. In Experiment 1, responses on a verbal measure of certainty provide evidence of binary noncomplementarity in the perceived likelihoods of possible scenario outcomes, and a comparison of verbal and numeric certainty estimates suggests that numeric probabilities overestimated the complementarity of people's certainty. Experiment 2 used a choice task to detect binary noncomplementarity. Soliciting numeric probability estimates prior to the choice task changed the participants' choices in a direction consistent with complementarity. Possible mechanisms yielding (non)complementarity are discussed. Copyright 2000 Academic Press. PMID- 10706815 TI - Choosing Work Group Members: Balancing Similarity, Competence, and Familiarity. AB - This study explores one of the contributors to group composition-the basis on which people choose others with whom they want to work. We use a combined model to explore individual attributes, relational attributes, and previous structural ties as determinants of work partner choice. Four years of data from participants in 33 small project groups were collected, some of which reflects individual participant characteristics and some of which is social network data measuring the previous relationship between two participants. Our results suggest that when selecting future group members people are biased toward others of the same race, others who have a reputation for being competent and hard working, and others with whom they have developed strong working relationships in the past. These results suggest that people strive for predictability when choosing future work group members. Copyright 2000 Academic Press. PMID- 10706816 TI - Using Advice and Assessing Its Quality. AB - People received advice from four sources and used it to produce a judgment. They also assessed the quality of advice by estimating the probability that it would be correct. They were better at assessing than at using advice: combinations of advice based on their assessments were superior to their judgments. Order of assessing and using advice, superficial differences between advisors, and using other methods of advice assessment had no significant effects on this superiority of advice assessment over advice use. However, use but not assessment was improved when some advisors exhibited biases opposite to those that people typically show. It appears that using advice imposes a heavier processing load than assessing its quality and that this load can be lightened by including advisors who exhibit unusual behavior. Their salience may help people working under a heavy processing load make appropriate pairings between advisor weights and advice. Copyright 2000 Academic Press. PMID- 10706817 TI - Coping with Unfavorable Attribute Values in Choice. AB - This paper examines how decision makers cope when faced with trade-offs between a higher quality alternative and a lower price alternative in situations where both alternatives involve relatively unfavorable versus relatively favorable values for quality. We hypothesize that choices between alternatives defined by unfavorable quality values will generate negative emotion, resulting in emotion focused coping behavior. Choosing the higher quality alternative (i.e., maximizing the quality attribute in choice) appears to function as a coping mechanism in these situations. These apparently coping-motivated choice effects are found even after methods are implemented to control for more cognitive factors associated with manipulations of quality-attribute value, such as the possibility that unfavorable attribute values are associated with increased attribute ranges and therefore increased relative importance for quality. Copyright 2000 Academic Press. PMID- 10706818 TI - Examining Models of Nondominated Decoy Effects across Judgment and Choice. AB - Three experiments explored cognitive models of inferior, compromise, and phantom decoy effects in both judgment and choice. Participants made judgments of attractiveness, justifiability, and evaluation anxiety associated with each alternative in the set, along with judgments of the attractiveness of each alternative's dimensional values. In another session, they also chose the alternative they most preferred. Results were analyzed in terms of the degree to which decoy effects reflected shifts in dimensional values or reflected emergent values based on relationships with other alternatives in the set. Both emergent value and value-shift models of inferior decoy effects were supported, but only the emergent-value model of compromise decoy effects was supported. Results for the phantom decoy indicated that this effect was choice-based and did not occur in judgment. Thus, although decoy effects were largely similar in choice and judgment, they also differed in important ways. Copyright 2000 Academic Press. PMID- 10706819 TI - The Impact of Conflict Issues on Fixed-Pie Perceptions, Problem Solving, and Integrative Outcomes in Negotiation. AB - It is argued that a negotiator's fixed-pie perception, cooperative motivation, problem-solving behavior, and integrative outcomes are influenced by the content of the negotiation-the conflict issue. Negotiation involves conflicting interests, conflicting ideas about intellective problems, or conflicting ideas about evaluative problems. Study 1 showed that individuals in a negotiation about interests have a stronger fixed-pie perception and have a lower cooperative motivation than individuals in an evaluative negotiation, with intellective negotiations taking an intermediate position. Study 2 showed that individuals in a negotiation about interests made more trade-offs and reached higher joint outcomes than individuals in an intellective or evaluative negotiation. Study 3 replicated this finding in a field study. The studies bridge insights from negotiation research and decision-making research and show that the conflict issue has important effects on the negotiation process. Copyright 2000 Academic Press. PMID- 10706820 TI - Introduction. PMID- 10706821 TI - Dictyostelium development-socializing through cAMP. AB - Starving Dictyostelium amoebae use cAMP as a chemoattractant to gather into aggregates, as a hormone-like signal to induce cell differentiation, and as an intracellular messenger to control stalk- and spore cell maturation and germination of spores. In this chapter we describe the respective roles of the three adenylyl cyclases ACA, ACB and ACG in controlling cAMP signaling during development and we discuss how cAMP signals are processed by the cells to trigger the large repertoire of gene regulatory events that is under control of this signal molecule. PMID- 10706822 TI - DIF signalling and cell fate. AB - The DIFs are a family of secreted chlorinated molecules that control cell fate during development of Dictyostelium cells in culture and probably during normal development too. They induce stalk cell differentiation and suppress spore cell formation. The biosynthetic and inactivation pathways of DIF-1 (the major bioactivity) have been worked out. DIF-1 is probably synthesised in prespore cells and inactivated in prestalk cells, by dechlorination. Thus, each cell type tends to alter DIF-1 level so as to favour differentiation of the other cell type. This relationship leads to a model for cell-type proportioning during normal development. PMID- 10706823 TI - Signalling pathways that direct prestalk and stalk cell differentiation in Dictyostelium. AB - Prestalk cell differentiation in Dictyostelium is induced by DIF and two DIF induced genes, ecmA and ecmB, have revealed the existence of multiple prestalk and stalk cell sub-types. These different sub-types are defined by the pattern of expression of subfragments derived from the ecmA and ecmB promoters. These markers have been utilised in three ways; for fate mapping in vivo, to investigate the molecular mechanisms underlying DIF signalling and to explore the relative requirement for DIF and other signalling molecules for prestalk and stalk cell differentiation in vitro. The heterogeneity of the prestalk and stalk populations seems to be reflected in differences in the cell signalling pathways that they utilise. PMID- 10706824 TI - Control of spatial patterning and cell-type proportioning in Dictyostelium. AB - The spatial patterning of prestalk and prespore cells in the slug arises from the differential sorting of newly differentiated cell types as the mound forms. This pattern is highly organized along an anterior-posterior axis and is constant irrespective of the size of the organism. Cell-type differentiation is plastic until late in development. A change in the ratio of cell types resulting from removal of part of the slug leads to a rapid restoration of the original ratio of the cell types through a pathway involving dedifferentiation, redifferentiation, and sorting of the existing cells. This review provides insight into various molecules, morphogens, and pathways regulating spatial patterning and cell-type proportioning. PMID- 10706825 TI - Morphogenetic cell movement in Dictyostelium. AB - Dictyostelium morphogenesis starts with the chemotactic aggregation of starving individual cells. The cells move in response to propagating waves of the chemoattractant cyclic AMP initiated by cells in the aggregation centre. During aggregation the cells begin to differentiate into several types with different signalling and chemotactic properties. These cell types sort out from each other to form an axial pattern in the slug. There is now good evidence that periodic chemotactic signals not only control aggregation, but also later stages of morphogenesis. These signals take the form of target patterns, spirals, multi armed spirals and scroll waves. I will discuss their role in the control of cell movement during mound and slug formation and in the formation of the fruiting body. PMID- 10706827 TI - How will We improve our journal? PMID- 10706826 TI - European journal of vascular and endovascular surgery: meeting the new millennium with optimism and new editors PMID- 10706828 TI - Endovascular healing is inadequate for fixation of Dacron stent-grafts in human aortoiliac vessels. AB - BACKGROUND: migration and kinking of stent-grafts can occur late after endovascular aneurysm repair. It is unknown if endovascular grafts incorporate enough to be permanently anchored. In this report, healing of aortic stent-grafts was assessed in humans. PATIENTS AND METHODS: we retrieved 23 Dacron stent-grafts from patients treated for an aortic aneurysm since 1993. Twelve stent-grafts were explanted at late conversion to open repair and 11 at autopsy. The deaths were unrelated to graft fixation. The median age of the patients was 74 years (IQR 55 84 years) and the grafts were explanted 9 months (1-31 months) after insertion. Microscopic slides were prepared by conventional techniques or by cutting and grinding arterial specimens embedded in plastic with the stent-grafts in situ. RESULTS: the stent-grafts detached readily from the native arteries at surgery or autopsy, except when the stents had hooks or barbs which engaged the vessel wall. A space filled with poorly organised blood components persisted between the graft and the aortic wall 2.5 years after implantation. No firm incorporation of the grafts was observed proximally in the aneurysm neck or distally in the iliac segment. A friable neo-intimal layer covered parts of the luminal aspect of the grafts. CONCLUSIONS: endovascular healing provides poor fixation of Dacron stent grafts in humans. At present, fixation relies on the mechanical properties of the stent-grafts. PMID- 10706829 TI - Effects of hypertonic saline-dextran solution on regional blood flow and thrombogenicity in PTFE grafts in the vena cava of the rabbit. AB - OBJECTIVES: to study the effects of hypervolaemic haemodilution with hypertonic saline-dextran solution (HSD) on regional blood flow and thrombogenicity of small diameter polytetrafluoroethylene (PTFE) grafts. DESIGN: blood flow in rabbit aorta, vena cava and femoral, renal and ear arteries was determined in five groups: controls, isovolaemic haemodilution with dextran-70 (10 ml/kg body weight (b.w. )), hypervolaemic haemodilution (10 ml/kg b.w.) with either dextran-70, 7.5% NaCl or a combination of dextran and NaCl (HSD). In a second series PTFE grafts were inserted into the vena cava of rabbits treated with hypervolaemic haemodilution with dextran, hypertonic saline or HSD and examined after two days. RESULTS: blood flow increased in aorta, vena cava and femoral artery after haemodilution. The increase was transient in animals treated with hypertonic NaCl alone but sustained in the dextran-70 groups. The grafts from animals treated with hypertonic saline alone had a lower thrombus mass and higher blood flow compared to those from rabbits haemodiluted with dextran-70 only, indicating that both dextran and NaCl have antithrombotic effects. Superior results were obtained with HSD solution. CONCLUSIONS: HSD solution has a strong flow-promoting action in several vascular beds and beneficial effects on the patency of small diameter vessel grafts. PMID- 10706830 TI - Abdominal aortic laparoscopic surgery: retroperitoneal or transperitoneal approach? AB - OBJECTIVE: to define the respective advantages and pitfalls of the trans- or retroperitoneal approaches in laparoscopic abdominal aortic reconstruction (LAOR). DESIGN: prospective study. MATERIAL: ten patients (8 males; average age 58) underwent an aortouni- (n=2) or bifemoral bypass (n=8) to treat aortoiliac occlusive disease (n=8) or an aortic aneurysm (n=2). METHODS: a retroperitoneal approach (the "apron" technique) was used in the first 5 cases (Group I) and a transperitoneal approach in the last 5 cases (Group II). RESULTS: no early or late death occurred, and all bypasses remain patent after a mean follow-up of 5.7 months. Mean surgical and clamping times are similar in both groups (370 and 126 min in Group I; 324 and 137 min in Group II). One intraoperative conversion to open surgery and two postoperative surgical complications occurred in Group I. Four minilaparotomies of 8-10 cm were necessary in Group II. Two patients were discharged on postoperative day 6 in Group I and five in Group II. CONCLUSION: this preliminary study shows the feasibility of LAOR through both approaches. In Group II, a better exposure of the right aortic wall and of the right iliac axis was noted and division of the inferior mesenteric artery was not always necessary. PMID- 10706831 TI - Endovascular femoropopliteal bypass combined with remote endarterectomy in SFA occlusive disease: initial experience. AB - OBJECTIVES: to evaluate the feasibility of endovascular femoropopliteal bypass in combination with remote endarterectomy of the superficial femoral artery (SFA). DESIGN: prospective, open study. MATERIALS: thirteen patients with chronic lower leg ischaemia due to femoropopliteal occlusive disease underwent 14 SFA remote endarterectomy procedures followed by endovascular ePTFE femoropopliteal bypass. Primary endografting was performed in seven cases. The indication for endograft insertion was vessel-wall perforation during endarterectomy in the remaining seven cases. METHODS: pre- and postoperative clinicl and haemodynamic data were collected and compared. Technical problems and procedure-related complications were noted. RESULTS: initial technical success was achieved in all 14 limbs. However, four early reocclusions occurred after 1, 4, 6 and 10 weeks postoperatively. Two late reocclusions were detected after 16 and 22 months without any preceding symptoms or haemodynamic changes. Primary and secondary patency rates were 61% and 70% at two years, probably due to graft-related factors, such as lack of radial force, graft folding or kinking, and possibly altered mechanical or thrombogenetic properties after dilatation of the ePTFE graft. CONCLUSIONS: endovascular femoropopliteal endo-bypass after SFA remote endarterectomy is a feasible procedure. Further technical improvements are necessary to avoid procedure- and graft-related early failures. PMID- 10706832 TI - The importance of revision of early restenosis after endovascular remote endarterectomy in SFA occlusive disease. AB - OBJECTIVES: to investigate the results of revision of recurrent stenoses after superficial femoral artery (SFA) remote endarterectomy. DESIGN: prospective, non open, study. MATERIALS: eighty-eight consecutive patients with long segmental SFA occlusive disease underwent 101 remote end-arterectomy procedures. All patients had chronic lower extremity ischaemia necessitating surgical intervention. METHODS: clinical, haemodynamic, and duplex examinations were performed postoperatively at regular intervals, identifying 46 recurrent stenosed (PSV ratio >2.5) limbs, which formed the cohort for this study. The median follow-up was 25 months. Secondary revision was performed in 23 limbs, based on recurrent symptoms and individual preference of the attending vascular surgeon. Cumulative primary and primary assisted-patency rates were compared using the log-rank test of significance. RESULTS: univariate analysis did not show any significant differences for other demographic and lesion characteristics apart from recurrent symptoms (all revised). Multivariate analysis revealed that revision "adjusted for time-of-onset" predicted reocclusion (p=0.007; HR 0. 21; 95% CI 0.06, 0.66). Among subjects in whom restenoses developed within 1 year, revision of recurrent stenoses improved primary patency rates from 47% to 77% at 30 months. CONCLUSIONS: revision of early (<1 year) recurrent stenoses improves the mid-term patency rates of SFA remote endarterectomy. PMID- 10706833 TI - Management of co-existing intra-abdominal disease in aortic surgery. AB - OBJECTIVES: the treatment of abdominal aortic aneurysms more than 5 cm in diameter is well accepted, but controversy surrounds the management of concomitant serious intra-abdominal lesions diagnosed in the perioperative period. This study was undertaken to demonstrate that synchronous surgery is feasible and safe in this group of patients. DESIGN: in 1978 a decision was made to undertake combined operations on all patients with an aortic aneurysm of 5 cm or more in diameter and a significant non-vascular intra-abdominal lesion requiring surgery. METHODS: the case records of 676 patients who had aortic grafting for aneurysmal disease or the urgent management of occlusive disease between 1978 and 1998 were analysed retrospectively. SETTING: district general hospital. RESULTS: fifty-six (8%) patients had co-existing intra-abdominal disease treated at the time of aortic graft surgery. There were three (5%) hospital deaths and seven patients required early reoperation. One patient developed a subphrenic abscess and there were three superficial wound infections. There has been no clinical evidence of aortic graft infection in this series. CONCLUSION: this single centre experience with synchronous surgery demonstrates that it is safe and does not appear to predispose to an increased risk of graft infection. PMID- 10706835 TI - The long-term outcome after axillo-axillary bypass grafting for proximal subclavian artery disease. AB - OBJECTIVES: to investigate the outcome of patients undergoing axillo-axillary bypass grafting for symptomatic subclavian artery stenoses or occlusions. DESIGN: retrospective case-note review and prospective review of patients available for follow-up. PATIENTS AND METHODS: sixteen patients had axillo-axillary grafts in a 17-year period. Ten patients were available for review and assessed clinically, by measurement of arm blood pressures, and by duplex scanning of their grafts. RESULTS: one patient died and three grafts occluded within 30 days of operation. Nine out of 10 grafts scanned were patent, with three further grafts clinically patent at death. Overall secondary patency was 75% at a combined median follow-up of 56 months (range 12-204 months). Recurrent symptoms occurred in two patients, one with an occluded graft and one with a patent graft. CONCLUSION: axillo axillary bypass grafts give good long-term symptom-free results. PMID- 10706834 TI - Pulmonary nitric oxide metabolism following infrarenal aortic cross-clamp-induced ischaemia-reperfusion injury. AB - OBJECTIVES: to investigate endogenous pulmonary nitric oxide metabolism following infrarenal aortic cross-clamp-induced ischaemia-reperfusion injury. METHODS: groups of male Wistar rats (n=6) were subjected to 60 minutes of infrarenal aortic cross-clamping under general anaesthesia. Rats were culled after 0, 60 and 120 minutes>> reperfusion, following release of the aortic clamp. A sham-operated control group was also studied. Acute lung injury (ALI) was quantified by measuring the protein concentration in lung bronchoalveolar lavage (BAL) fluid. Pulmonary myeloperoxidase activity (MPO) was measured as an index of neutrophil infiltration and degranulation in the lung. Plasma tumour-necrosis factor-alpha (TNF-alpha) was measured as an index of the pro-inflammatory cytokine response and pulmonary nitric oxide synthase (NOS) activity was determined by measuring conversion of(3)H L-arginine to(3)H L-citrulline in tissue homogenates. RESULTS: these data show significant ALI with increased pulmonary microvascular permeability and MPO activity in animals subject to 60 minutes>> ischaemia and 60 minutes or 120 minutes of reperfusion compared to control animals (p<0.01). Plasma TNF-alpha levels were significantly increased following 60 minutes of ischaemia compared to controls (p<0.01) and remained significantly increased in animals subject to reperfusion (p<0.01). Pulmonary NOS activity was significantly increased in animals subject to reperfusion (p<0.01). CONCLUSIONS: the reperfusion phase of infrarenal aortic cross-clamping provokes a significant increase in pulmonary NOS metabolism. The increase in plasma TNF-alpha and MPO activity suggests that this response may be secondary to inducible NOS expression. Manipulation of this response may benefit patients at risk of acute injury following infrarenal aortic reconstruction. PMID- 10706836 TI - Transcranial Doppler-directed Dextran-40 therapy is a cost-effective method of preventing carotid thrombosis after carotid endarterectomy. AB - OBJECTIVES: perioperative stroke reduces the clinical effectiveness of carotid endarterectomy (CEA). Postoperative thrombotic stroke may be reduced in incidence by the use of transcranial Doppler-directed Dextran-40 therapy. This programme requires the purchase of additional equipment and employment of more staff. This study examined whether this additional financial outlay was cost-effective in terms of saving expenditure by preventing postoperative thrombotic stroke. MATERIALS AND METHODS: data was collected prospectively on a series of 600 consecutive CEAs. The costs of the monitoring programme were analysed over 1- and 5-year periods. Formulae were derived allowing other units to calculate whether this technique will be cost-effective for them. RESULTS: after the introduction of TCD monitoring the postoperative thrombotic stroke rate fell from 2.7% to 0% (8 strokes prevented). Our local unit cost for the treatment of stroke was 25,702 pounds. After allowing for the additional costs of the monitoring programme, we calculate that postoperative TCD has saved 171,393 pounds. CONCLUSIONS: postoperative TCD monitoring is a clinically effective and also cost-effective method of reducing the stroke rate associated with CEA. For units performing more than 50 CEAs per year who experience occasional postoperative carotid thrombosis, its introduction should be considered. PMID- 10706837 TI - The clinical management and outcome of venous ulcers in legs with deep-venous obstruction. AB - OBJECTIVE: as a result of a serious complication from compression bandaging in a patient with venous ulceration and deep-vein obstruction, a policy of incremental compression in such limbs has been developed. The purpose of this retrospective study is to review the outcome of this policy. DESIGN: limbs with deep-venous obstruction (stenosis or occlusion) were treated initially with 3-layer compression bandaging and reviewed 24 h later. If 3-layer bandaging was tolerated, it was re-applied for a further 48 h. If there were no problems, then 4-layer bandaging was applied and the patient reviewed at 24 and 72 h. If 4-layer bandaging could not be tolerated, the limb was returned to 3-layer bandaging. RESULTS: of 325 limbs seen in a venous-ulcer clinic, 22 (7%) had deep-vein obstruction. Fifteen (68%) limbs were able to tolerate 4-layer bandaging, five (23%) could tolerate 3-layer bandaging and two limbs (9%) could only tolerate class 2 compression hosiery. The overall 1-year healing rate was 55%. There were no serious complications from bandaging. CONCLUSIONS: a protocol of incremental compression bandaging is safe in ulcerated legs with deep-vein obstruction and produces healing in up to 55% of cases. PMID- 10706838 TI - Management of penetrating cervicomediastinal venous trauma. AB - OBJECTIVES: to evaluate the results of management of penetrating cervicomediastinal venous trauma. DESIGN: retrospective study. Materials forty nine consecutive patients with cervical and thoracic venous injuries treated at a tertiary hospital between 1991 and 1997. Method patients identified from a computerised database and data extracted from case records. RESULTS: forty-five patients were male and the mean age was 25.3 years. Forty injuries were due to stabs and 9 to gunshots. 22 patients were shocked, 25 actively bleeding and 31 were anaemic. Veins injured were internal jugular in 25, subclavian in 15, brachiocephalic in 6, and superior vena cava in 3. Injured veins were ligated in 25 cases and repaired by lateral suture in 22. No complex repairs were performed. There were 8 perioperative deaths and 5 cases of transient postoperative oedema. Venous ligation was not associated with increased risk of postoperative oedema. CONCLUSIONS: ligation is an acceptable form of treatment of cervicomediastinal venous injuries in the presence of haemodynamic instability, or where complex methods of repair would otherwise be necessary. PMID- 10706839 TI - The snuffbox arteriovenous fistula for vascular access. AB - OBJECTIVES: to determine the applicability, patency rates and factors influencing patency of snuffbox arteriovenous fistulae for haemodialysis access. DESIGN: retrospective non-randomised study. MATERIALS AND METHODS: patency was determined by reference to an ongoing database and dialysis records of 645 vascular access procedures between 1985 and 1997, including 210 snuffbox fistulae in 201 patients. RESULTS: snuffbox fistulae comprised 189/376 (50%) primary procedures. Records of 208 snuffbox fistulae were available for patency analysis by the life table method. Twenty-two (11%) thrombosed within 24 hours of operation. After six weeks 80% were used for dialysis. Cumulative patency was 65% at 1 year and 45% at 5 years. After thrombosis of snuffbox fistulae, ipsilateral wrist fistulae could be constructed in 45%. Fistula patency was significantly better in men than women (p<0.001) and for left- than right-sided fistulae (p<0.001). Diabetes, age >70 years, and the prior commencement of haemodialysis did not significantly affect fistula survival. CONCLUSIONS: the snuffbox AV fistula gives a long segment of arterialised vein for needling and preserves proximal vessels. It is feasible in 50% of patients requiring primary access and has good long-term patency, especially in men. A more proximal fistula may be preferable in women with smaller vessels. PMID- 10706840 TI - Angiographic embolisation in arterial trauma. AB - OBJECTIVES: to evaluate the use of endovascular occlusion in the treatment of arterial trauma. METHODS: records of patients with penetrating arterial injuries treated by endovascular occlusive techniques were culled from the computerised database of the vascular service. RESULTS: the study period spanned 7 years. Forty-two patients were studied with injuries to the cervicofacial vessels (24), lower limb (16) and upper limb (1). 13 had an arteriovenous fistula. There were 4 failures. In 2 cannulation was not achieved and in 2 with A-VF distal vessel occlusion was impossible. Two patients developed complications. In the remainder, treatment was effective and durable. CONCLUSION: this treatment modality is effective and safe in the treatment of penetrating trauma in selected patients. PMID- 10706841 TI - Endovascular repair of aortic pseudoaneurysms. PMID- 10706842 TI - Vertebral artery embolism post subclavian artery injury with occipital lobe infarction. PMID- 10706843 TI - Buttock claudication secondary to isolated internal iliac artery stenosis. PMID- 10706844 TI - Focal myositis - a new presentation. AB - This article is a case report and review of literature of a very rare condition, not previously written in general surgical literature, including a new presentation PMID- 10706845 TI - A long distal prosthetic bypass grafting from descending aorta to distal peroneal artery. Sometimes it works! PMID- 10706846 TI - Consumptive coagulopathy following endovascular stent repair of abdominal aortic aneurysm. PMID- 10706847 TI - Ergotism precipitated by erythromycin: a rare case of vasospasm. PMID- 10706848 TI - An aneurysm at the back of a thigh: a rare presentation of a congenitally persistent sciatic artery. PMID- 10706849 TI - Endovascular treatment of a ruptured thoracic aortic aneurysm. PMID- 10706851 TI - Prognostic implications of differences in telomere length between normal and malignant cells from patients with chronic myeloid leukemia measured by flow cytometry. AB - Chronic myeloid leukemia (CML) is a clonal, multilineage myeloproliferative disorder characterized by the Philadelphia chromosome (Ph) and a marked expansion of myeloid cells. Previous studies have indicated that the telomere length in blood cells may indicate their replicative history. However, the large variation in telomere length between individuals complicates the use of this parameter in CML and other hematologic disorders. To circumvent this problem, we compared the telomere length in peripheral blood or bone marrow cells with purified normal (Ph(-)) T lymphocytes from the same CML patient using fluorescence in situ hybridization and flow cytometry. Overall telomere fluorescence was significantly reduced in Ph(+) cells from patients with CML compared to blood leukocytes from normal individuals (P < 0.001) or normal (Ph(-)) T lymphocytes from the same individuals (n = 51, P < 0.001). Cells from patients in accelerated phase or blast phase (AP/BP) showed significantly shorter average telomere length than cells from patients in chronic phase (CP, P = 0.02) or cytogenetic remission (CR, P = 0.03). Patients in CP who subsequently developed BP within 2 years had significantly shorter telomeres than those who did not develop BP for at least 2 years (P < 0.05). Accelerated replication-dependent telomere shortening in Ph(+ )versus Ph(-) leukocytes supports previous evidence that Ph(+) stem cells cycle more actively than their counterparts in normal individuals. Our data further suggest that telomere shortening may serve as a surrogate marker of disease progression in patients with CP CML, supporting a mechanistic link between CML stem cell turnover, genetic instability, and malignant evolution in this disease. (Blood. 2000;95:1883-1890) (Blood. 2000;95:1883-1890) PMID- 10706852 TI - Evidence that tristetraprolin is a physiological regulator of granulocyte macrophage colony-stimulating factor messenger RNA deadenylation and stability. AB - Deficiency of tristetraprolin (TTP), the prototype of the CCCH zinc finger proteins, results in a complex inflammatory syndrome in mice. Most aspects of the syndrome are secondary to excess circulating tumor necrosis factor (TNF)-alpha, a consequence of increased stability of TNF-alpha messenger RNA (mRNA) in TTP deficient macrophages. TTP can bind directly to the AU-rich element in TNF-alpha mRNA, increasing its lability. Here we show that TTP deficiency also results in increased cellular production of granulocyte-macrophage colony-stimulating factor (GM-CSF) and increased stability of its mRNA, apparently secondary to decreased deadenylation. Similar findings were observed in mice also lacking both types of TNF-alpha receptors, excluding excess TNF-alpha production as a cause of the increased GM-CSF mRNA levels and stability. TTP appears to be a physiological regulator of GM-CSF mRNA deadenylation and stability. (Blood. 2000;95:1891-1899) PMID- 10706853 TI - Cell adhesion receptors in lymphoma dissemination. AB - Regulated lymphocyte trafficking is essential for the control and integration of systemic immune responses. This homing process disperses the immunologic repertoire, guides lymphocyte subsets to the specialized microenvironments that control their differentiation and survival, and targets immune effector cells to sites of antigenic insult. This review discusses data indicating that the adhesion receptors regulating the trafficking of normal lymphocytes are also expressed and functionally active in their malignant counterparts, the non Hodgkin lymphomas. These "homing receptors" appear to mediate the highly tissue specific dissemination of specific lymphoma subtypes, such as lymphomas of the mucosa-associated lymphoid tissues and lymphomas of the skin. Furthermore, as a result of their capability to enhance lymphoma dissemination and to transduce signals into the cell, promoting cell growth and survival, adhesion receptors may contribute to lymphoma aggressiveness. Taken together, the data offer a framework for understanding the dissemination routes of non-Hodgkin lymphomas and suggest that adhesion receptors, specifically those of the CD44 family, may present useful tools to predict prognosis in patients with lymphomas. (Blood. 2000;95:1900-1910) PMID- 10706854 TI - Eotaxin induces degranulation and chemotaxis of eosinophils through the activation of ERK2 and p38 mitogen-activated protein kinases. AB - Eotaxin and other CC chemokines acting via CC chemokine receptor-3 (CCR3) are believed to play an integral role in the development of eosinophilic inflammation in asthma and allergic inflammatory diseases. However, little is known about the intracellular events following agonist binding to CCR3 and the relationship of these events to the functional response of the cell. The objectives of this study were to investigate CCR3-mediated activation of the mitogen-activated protein (MAP) kinases extracellular signal-regulated kinase-2 (ERK2), p38, and c-jun N terminal kinase (JNK) in eosinophils and to assess the requirement for MAP kinases in eotaxin-induced eosinophil cationic protein (ECP) release and chemotaxis. MAP kinase activation was studied in eotaxin-stimulated eosinophils (more than 97% purity) by Western blotting and immune-complex kinase assays. ECP release was measured by radioimmunoassay. Chemotaxis was assessed using Boyden microchambers. Eotaxin (10(-11) to 10(-7) mol/L) induced concentration-dependent phosphorylation of ERK2 and p38. Phosphorylation was detectable after 30 seconds, peaked at about 1 minute, and returned to baseline after 2 to 5 minutes. Phosphorylation of JNK above baseline could not be detected. The kinase activity of ERK2 and p38 paralleled phosphorylation. PD980 59, an inhibitor of the ERK2 activating enzyme MEK (MAP ERK kinase), blocked phosphorylation of ERK2 in a concentration-dependent manner. The functional relevance of ERK2 and p38 was studied using PD98 059 and the p38 inhibitor SB202 190. PD98 059 and SB202 190 both caused inhibition of eotaxin-induced ECP release and chemotaxis. We conclude that eotaxin induces a rapid concentration-dependent activation of ERK2 and p38 in eosinophils and that the activation of these MAP kinases is required for eotaxin-stimulated degranulation and directed locomotion. (Blood. 2000;95:1911 1917) PMID- 10706855 TI - Impact of bone marrow transplantation for symptomatic sickle cell disease: an interim report. Multicenter investigation of bone marrow transplantation for sickle cell disease. AB - Fifty children who had symptomatic sickle cell disease received matched sibling marrow allografts between September 1991 and March 1999, with Kaplan-Meier probabilities of survival and event-free survival of 94% and 84%, respectively. Twenty-six patients (16 male, 10 female) had at least 2 years of follow-up after transplantation and were evaluated for late effects of transplantation and for its impact on sickle cell-related central nervous system (CNS) and pulmonary disease. Patients ranged between 3.3 and 14.0 (median, 9. 4) years of age and had a median follow-up of 57.9 (range 38-95) months after transplantation. Among 22 of 26 patients who had stable donor engraftment, complications related to sickle cell disease resolved, and none experienced further episodes of pain, stroke, or acute chest syndrome. All 10 engrafted patients with a prior history of stroke had stable or improved cerebral magnetic resonance imaging results. Pulmonary function tests were stable in 22 of the 26 patients, worse in two, and not studied in two. Seven of eight patients transplanted for recurrent acute chest syndrome had stable pulmonary function. Linear growth measured by median height standard deviation score improved from -0.7 before transplantation to -0.2 after transplantation. An adverse effect of busulfan conditioning on ovarian function was demonstrated in five of seven evaluable females who are currently at least 13 years of age. None of the four males tested had elevated serum gonadotropin levels. These data confirm that allogenic bone marrow transplantation establishes normal erythropoiesis and is associated with improved growth and stable CNS imaging and pulmonary function in most patients. (Blood. 2000;95:1918-1924) PMID- 10706856 TI - Deletion of 13q14 remains an independent adverse prognostic variable in multiple myeloma despite its frequent detection by interphase fluorescence in situ hybridization. AB - Interphase fluorescence in situ hybridization (FISH) studies of chromosomal region 13q14 were performed to investigate the incidence and clinical importance of deletions in multiple myeloma (MM). Monoallelic deletions of the retinoblastoma-1 (rb-1) gene and the D13S319 locus were observed in 48 of 104 patients (46.2%) and in 28 of 72 (38.9%) patients, respectively, with newly diagnosed MM. FISH studies found that 13q14 was deleted in all 17 patients with karyotypic evidence of monosomy 13 or deletion of 13q but also in 9 of 19 patients with apparently normal karyotypes. Patients with a 13q14 deletion were more likely to have stage III disease (P =.022), higher serum levels of beta(2) microglobulin (P =.059), and a higher percentage of bone marrow plasma cells (P =.085) than patients with a normal 13q14 status on FISH analysis. In patients with a deletion of 13q14, myeloma cell proliferation (Ki-67) was markedly increased (22.0% +/- 6.9% compared with 15.6% +/- 8.2% in patients without the deletion; P =.0008). Evaluation of bromodeoxyuridine incorporation in 5 patients revealed that both rb-1-deleted and rb-1-normal MM subpopulations were proliferative. The presence of a 13q14 deletion on FISH analysis was associated with a significantly lower rate of response to conventional-dose chemotherapy (40.8% compared with 78. 6%; P =.009) and a shorter overall survival (24.2 months compared with > 60 months; P <.005) than in patients without the deletion. Multivariate analysis of prognostic factors confirmed the independent predictive value of 13q14 deletions for shortened survival. In conclusion, deletions of 13q14 are frequently detected by interphase FISH in patients with newly diagnosed MM, correlate with increased proliferative activity, and represent an independent adverse prognostic feature in MM. (Blood. 2000;95:1925-1930) PMID- 10706857 TI - Antilymphocyte globulin, cyclosporine, prednisolone, and granulocyte colony stimulating factor for severe aplastic anemia: an update of the GITMO/EBMT study on 100 patients. European Group for Blood and Marrow Transplantation (EBMT) Working Party on Severe Aplastic Anemia and the Gruppo Italiano Trapianti di Midolio Osseo (GITMO). AB - One hundred consecutive patients with severe aplastic anemia (SAA) received horse antilymphocyte globulin (ALG), cyclosporin A (CyA), 6-methylprednisolone (6Mpred), and granulocyte colony-stimulating factor (G-CSF) as first-line therapy. The median age was 16 years (range, 1-72 years) and median neutrophil count was 0.2 x 10(9)/L (range, 0-0.5 x 10(9)/L). Trilineage hematologic recovery (at a median interval of 96 days from treatment) was seen in 77 patients (48 complete, 29 partial) after 1 (n = 50) or more courses of ALG (n = 27). Of the 23 nonresponders, 11 patients died at a median interval of 83 days (range, 16-1132 days), 6 were considered treatment failures and underwent transplantation, and 6 were pancytopenic. Cytogenetic abnormalities were seen in 11% of patients, clonal hematologic disease in 8%, and relapse of marrow aplasia in 9%. The actuarial survival at 5 years was 87% (median follow-up 1424 days): 76% versus 98% for patients with neutrophil counts less than versus greater than 0.2 x 10(9)/L (P =.001) and 88% versus 87% for patients aged less than versus more than 16 years (P =.8). The actuarial probability of discontinuing CyA was 38%. Patients who did not achieve a white blood cell (WBC) count of 5 x 10(9)/L during G-CSF treatment have a low probability of responding (37%) and a high mortality rate (42%). This update confirms a high probability for SAA patients of becoming transfusion independent and of surviving after treatment with ALG, CyA, 6Mpred, and G-CSF, with a significant effect of neutrophil counts on outcome. Problems still remain, such as absent or incomplete responses, clonal evolution, relapse of the original disease, and cyclosporine dependence. Early transplantation, also from alternative donors, may be warranted in patients with poor WBC response to G-CSF. (Blood. 2000;95:1931-1934) PMID- 10706858 TI - Genetic analysis, phenotypic diagnosis, and risk of venous thrombosis in families with inherited deficiencies of protein S. AB - Protein S deficiency is a recognized risk factor for venous thrombosis. Of all the inherited thrombophilic conditions, it remains the most difficult to diagnose because of phenotypic variability, which can lead to inconclusive results. We have overcome this problem by studying a cohort of patients from a single center where the diagnosis was confirmed at the genetic level. Twenty-eight index patients with protein S deficiency and a PROS1 gene defect were studied, together with 109 first-degree relatives. To avoid selection bias, we confined analysis of total and free protein S levels and thrombotic risk to the patients' relatives. In this group of relatives, a low free protein S level was the most reliable predictor of a PROS1 gene defect (sensitivity 97.7%, specificity 100%). First degree relatives with a PROS1 gene defect had a 5.0-fold higher risk of thrombosis (95% confidence interval, 1. 5-16.8) than those with a normal PROS1 gene and no other recognized thrombophilic defect. Although pregnancy/puerperium and immobility/trauma were important precipitating factors for thrombosis, almost half of the events were spontaneous. Relatives with splice-site or major structural defects in the PROS1 gene were more likely to have had a thrombotic event and had significantly lower total and free protein S levels than those relatives having missense mutations. We conclude that persons with PROS1 gene defects and protein S deficiency are at increased risk of thrombosis and that free protein S estimation offers the most reliable way of diagnosing the deficiency. (Blood. 2000;95:1935-1941) PMID- 10706859 TI - Aberrant p15 promoter methylation in adult and childhood acute leukemias of nearly all morphologic subtypes: potential prognostic implications. AB - We prospectively analyzed p15 and p16 promoter methylation patterns using methylation-specific polymerase chain reaction (PCR) in patients with adult and childhood acute leukemias and studied the association of methylation patterns with chromosomal abnormalities and prognostic variables. In nearly all French American-British leukemia subtypes, we found p15 methylation in bone marrow or peripheral blood cells from 58% (46/79) of patients with acute myeloid leukemia (AML), acute lymphoblastic leukemia (ALL), or acute biphenotypic leukemia (ABL). An identical alteration was detected in blood plasma from 11 of 12 of these patients (92%). We also demonstrated for the first time concomitant p16 and p15 methylation in 22% (8/37) of adults with AML or ALL, exclusively in those with M2, M4, or L2 subtypes. According to cytogenetic data from 35 patients with ALL, AML, or ABL, 82% (14/17) of those with unmethylated p15 alleles had normal karyotypes or hyperdiploidies associated with a favorable prognosis. Conversely, 44% (8/18) of patients with p15 methylation had chromosomal translocations, inversions, or deletions, suggesting an interplay of these abnormalities with p15 methylation. As a prognostic marker for disease monitoring, p15 methylation appears to be more widely applicable than BCR-ABL, AF4-MLL, and AML1-ETO transcripts, which were detectable in only 8% (4/48) of patients by reverse transcriptase-PCR. Thirty-nine of 43 blood samples (91%) sequentially collected from 12 patients with AML, ALL, or ABL showed p15 methylation status in excellent concordance with morphologic disease stage. Early detection of p15 methylation at apparent remission or its acquisition during follow-up may prove valuable for predicting relapse. Overall survival of patients with p15 methylation was notably shortened among 38 adults with AML and 12 adults with ALL. Aberrant p15 methylation may have important prognostic implications for clinical monitoring and risk assessment. (Blood. 2000;95:1942-1949) PMID- 10706860 TI - Non-MALT marginal zone B-cell lymphomas: a description of clinical presentation and outcome in 124 patients. AB - Marginal zone B-cell lymphoma (MZL) is a recently individualized lymphoma that encompasses mucosa-associated lymphoid tissue (MALT) lymphoma, splenic lymphoma with or without villous lymphocytes, and nodal lymphoma with or without monocytoid B-cells. If the clinical description and outcome of MALT lymphoma is well known, this is not the case for the other subtypes. We reviewed 124 patients presenting non-MALT MZL treated in our department to describe the morphologic and clinical presentation and the outcome of these lymphomas. Four clinical subtypes were observed: splenic, 59 patients; nodal, 37 patients; disseminated (splenic and nodal), 20 patients; and leukemic (not splenic nor nodal), 8 patients. These lymphomas were usually CD5-, CD10-, CD23-, and CD43-, but the detection of one or, rarely, two of these antigens may be observed. Bone marrow and blood infiltrations were frequent, except in the nodal subtype, but these locations were not associated with a poorer outcome. Splenic and leukemic subtypes were associated with a median time to progression (TTP) longer than 5 years, even in the absence of treatment or of complete response to therapy. Nodal and disseminated subtypes were associated with a median TTP of 1 year. However, in all these subtypes, survival was good with a median survival of 9 years, allowing these lymphomas to be classified as indolent. Because of the retrospective nature of this analysis, no conclusion may be drawn on the therapeutic aspects, but conservative treatments seem recommended for leukemic and splenic subtypes. (Blood. 2000;95:1950-1956) PMID- 10706862 TI - Circulating hematopoietic progenitor cells in first trimester fetal blood. AB - The yolk sac and aorto-gonad-mesonephros region are well recognized as the principal sites of hematopoiesis in the developing embryo, and the liver is the principal site of hematopoiesis in the fetus. However, little is known about circulating hematopoietic stem and progenitor cells in early fetal life. We investigated the number and characteristics of circulating progenitors in first trimester blood of 64 human fetuses (median gestational age, 10(+4) weeks; range, 7(+6)-13(+6) weeks). CD34+ cells accounted for 5.1 +/- 1.0% of CD45+ cells in first trimester blood, which is significantly more than in term cord blood (0.4 +/- 0.03%; P =.0015). However, the concentration of CD34+ cells (6.6 +/- 2.4 x 10(4)/mL) was similar to that in term cord blood (5.6 +/- 3.9 x 10(4)/mL). The total number of progenitors cultured from unsorted mononuclear cells (MNCs) in first trimester blood was 19.2 +/- 2.1 x 10(3)/mL, which is similar to that in term cord blood (26.4 +/- 5.6 x 10(3)/mL). All lineages were seen: colony-forming unit-GEMM (CFU-GEMM), CFU-GM, BFU-e, BFU-MK, and CFU-MK. Clonogenic assays of CD34+ cells purified from first trimester samples produced mainly two lineages: BFU-e (39.0 +/- 9.6 x 10(3)/mL CD34+ cells) and CFU-GEMM (22.6 +/- 4.7 x 10(3)/mL CD34+ cells). Short-term liquid culture of first trimester blood MNCs in SCF + IL 3 + Flt-3 (stem cell factor + interleukin-3 + Flt-3) increased, by 7-fold, the numbers of CFU-GEMM and induced a dramatic increase in BFU-e (65.6 +/- 12.1 fold). These data show that significant numbers of committed and multipotent progenitors with capacity for expansion circulate in first trimester fetal blood and can be CD34 selected. These cells should be suitable targets for gene transfer and stem cell transplantation and, because fetal hematopoietic progenitors have been demonstrated in the maternal circulation from early gestation, may also be manipulated for noninvasive prenatal diagnosis of major genetic disorders. (Blood. 2000;95:1967-1972) PMID- 10706861 TI - A secreted and LIF-mediated stromal cell-derived activity that promotes ex vivo expansion of human hematopoietic stem cells. AB - The development of culture systems that facilitate ex vivo maintenance and expansion of transplantable hematopoietic stem cells (HSCs) is vital to stem cell research. Establishment of such culture systems will have significant impact on ex vivo manipulation and expansion of transplantable stem cells in clinical applications such as gene therapy, tumor cell purging, and stem cell transplantation. We have recently developed a stromal-based culture system that facilitates ex vivo expansion of transplantable human HSCs. In this stromal-based culture system, 2 major contributors to the ex vivo stem cell expansion are the addition of leukemia inhibitory factor (LIF) and the AC6.21 stromal cells. Because the action of LIF is indirect and mediated by stromal cells, we hypothesized that LIF binds to the LIF receptor on AC6.21 stromal cells, leading to up-regulated production of stem cell expansion promoting factor (SCEPF) and/or down-regulated production of stem cell expansion inhibitory factor (SCEIF). Here we demonstrate a secreted SCEPF activity in the conditioned media of LIF-treated AC6.21 stromal cell cultures (SCM-LIF). The magnitude of ex vivo stem cell expansion depends on the concentration of the secreted SCEPF activity in the SCM LIF. Furthermore, we have ruled out the contribution of 6 known early-acting cytokines, including interleukin-3, interleukin-6, granulocyte macrophage colony stimulating factor, stem cell factor, flt3 ligand, and thrombopoietin, to this SCEPF activity. Although further studies are required to characterize this secreted SCEPF activity and to determine whether this secreted SCEPF activity is mediated by a single factor or by multiple growth factors, our results demonstrate that stromal cells are not required for this secreted SCEPF activity to facilitate ex vivo stem cell expansion. (Blood. 2000;95:1957-1966) PMID- 10706863 TI - Recombinant full-length tissue factor pathway inhibitor fails to bind to the cell surface: implications for catabolism in vitro and in vivo. AB - Tissue factor pathway inhibitor (TFPI) plays a key role in the regulation of tissue factor-initiated blood coagulation secondary to loss of the integrity of the blood vessel wall. TFPI is a naturally occurring Kunitz-type protease inhibitor that inhibits coagulation factor Xa and, in a factor Xa-dependent manner, mediates feedback inhibition of the factor VIIa/tissue factor catalytic complex. In vivo full-length TFPI is thought to be primarily bound to the vascular endothelium and the high affinity binding requires an intact carboxy terminus. Here we describe a full-length TFPI molecule, expressed in mouse C127 cells (TFPI(C127)), which exhibits virtually no cellular binding yet contains the intact carboxy terminus. This TFPI (TFPI(C127)) is neither internalized nor degraded via the TFPI endocytic receptor, LDL-receptor-related protein. Pharmacokinetic studies of TFPI(C127 )in vivo demonstrate a 10-fold prolongation in the plasma half-life, compared with that of bacterial recombinant TFPI. (Blood. 2000;95:1973-1978) PMID- 10706864 TI - Characterization of the vasculogenic block in the absence of vascular endothelial growth factor-A. AB - Vascular endothelial growth factor (VEGF) signaling is required for both differentiation and proliferation of vascular endothelium. Analysis of differentiated embryonic stem cells with one or both VEGF-A alleles deleted showed that both the differentiation and the expansion of endothelial cells are blocked during vasculogenesis. Blood island formation was reduced by half in hemizygous mutant VEGF cultures and by 10-fold in homozygous mutant VEGF cultures. Homozygous mutant cultures could be partially rescued by the addition of exogenous VEGF. RNA levels for the endothelial adhesion receptors ICAM-2 and PECAM were reduced in homozygous mutant cultures, but ICAM-2 RNA levels decreased substantially, whereas PECAM RNA levels remained at hemizygous levels. The quantitative data correlated with the antibody staining patterns because cells that were not organized into vessels expressed PECAM but not ICAM-2. These PECAM+ cell clumps accumulated in mutant cultures as vessel density decreased, suggesting that they were endothelial cell precursors blocked from maturation. A subset of PECAM+ cells in clumps expressed stage-specific embryonic antigen-1 (SSEA-1), and all were ICAM-2(-) and CD34(-), whereas vascular endothelial cells incorporated into vessels were PECAM(+), ICAM-2(+), CD34(+), and SSEA-1(-). Analysis of flk-1 expression indicated that a subset of vascular precursor cells coexpressed PECAM and flk-1. These data suggest that VEGF signaling acts in a dose-dependent manner to affect both a specific differentiation step and the subsequent expansion of endothelial cells. (Blood. 2000;95:1979-1987) PMID- 10706865 TI - Rifampicin-dependent antibodies bind a similar or identical epitope to glycoprotein IX-specific quinine-dependent antibodies. AB - The drug-dependent antibody of a patient with rifampicin-induced thrombocytopenia was characterized using the antigen-capture enzyme-linked immunosorbent assay (MAIPA assay), flow cytometry, and immunoprecipitation. The antibody was found to bind glycoprotein (GP) Ib-IX but not GPIIb-IIIa because (1) it immunoprecipitated drug-dependently the former but not the latter glycoprotein complex and (2) the MAIPA assay showed strong rifampicin-dependent antibody binding when anti-GPIb-IX monoclonal antibodies (mAbs) (AK2 and FMC25) but not anti-GPIIb-IIIa mAbs (AP2, SZ21, and SZ22) were used to capture the antigen. The antibody binding site was further localized to the GPIX subunit of the GPIb-IX complex because flow cytometric analysis revealed drug-dependent antibody binding to L cells transfected with human GPIbbeta and GPIX complementary DNA (L betaIX cells) but not with human GPIbalpha and GPIbbeta complementary DNA (L alphabeta cells). Finally, in the MAIPA assay, the rifampicin-dependent antibody almost completely cross-blocked the binding of the anti-GPIX mAb (SZ1) to platelets. Similar cross blocking of SZ1binding to platelets by the quinine-dependent antibodies was also observed. This finding not only confirms that the epitope of the rifampicin dependent antibody is on GPIX but it is also identical to or located in close proximity to that of the quinine-dependent antibody and SZ1. Further characterization of the epitopes of these antibodies may have important implications for a general understanding of the mechanism of drug-induced thrombocytopenia. (Blood. 2000;95:1988-1992) PMID- 10706866 TI - Identification of a family of alternatively spliced mRNA species of angiopoietin 1. AB - Angiopoietin-1 (Ang-1) is required for developing vessels, and its absence leads to defects in vessel remodeling. Ang-1 has been identified as the ligand for the tyrosine kinase receptor Tie-2, which is expressed specifically on endothelial cells and early hematopoietic cells. In studying the role of Tie-2 and Ang-1 in megakaryocytopoiesis, 3 alternatively spliced species of Ang-1 mRNA (Ang-1.3 kb, Ang-0.9 kb, and Ang-0.7 kb) were identified in addition to the full-length Ang-1 (Ang-1.5 kb), in the megakaryocyte cell line CHRF by reverse transcription polymerase chain reaction (RT-PCR), and then cloned and sequenced. The expression of 3 alternatively spliced isoforms of Ang-1 was confirmed by RT-PCR using specific primer pairs derived from junction sites and the 3' end of Ang-1 cDNA, and it was further demonstrated by nuclease protection assay, Northern blotting, and immunoblotting in CHRF cells. Expression of the Ang-1.3 kb isoform was also detected in human primary fibroblast cell line FS4, breast cancer cell line MDAMB 468, and CD34(+)CD41(+) cells of fetal liver and platelets. The function of the 1.5-kb, 1.3-kb, and 0.9-kb isoforms was examined. Recombinant proteins Ang-1.5 and 0.9 kb bind strongly to the recombinant Tie-2 receptor (Tie-2-Fc), whereas the 1.3-kb isoform does not. The Ang-1.3 kb isoform binds to the 1.5-kb isoform. Ang-1. 5 kb, but not the 1.3-kb and 0.9-kb isoforms, induces tyrosine phosphorylation of Tie-2 in human umbilical vein endothelial cells. These data suggest that isoforms 1.3 kb and 0.9 kb could serve as dominant negative molecules for the full-length Ang-1. The possible involvement of the newly identified Ang-1 isoforms in angiogenesis and in growth and differentiation of hematopoietic progenitor cells provides a greater complexity to these processes. (Blood. 2000;95:1993-1999) PMID- 10706867 TI - Two novel type 2N von Willebrand disease-causing mutations that result in defective factor VIII binding, multimerization, and secretion of von Willebrand factor. AB - Two novel mutations, a T-to-C transition at nucleotide 2612 and a T-to-G transversion at nucleotide 3923 of the von Willebrand factor (vWF) complementary DNA, were detected by analysis of the vWF gene in DNA from members of 2 families with atypical von Willebrand disease. The T2612C transition predicts substitution of cysteine by arginine at amino acid position 788 (C788R), and the T3923G transversion predicts substitution of cysteine by glycine at position 1225 (C1225G) of pre-pro-vWF. The patients homozygous for the C788R and C1225G mutations both had a partial vWF deficiency (0. 18 IU/mL and 0.07 IU/mL vWF antigen, respectively); vWF in plasma from patients homozygous for either the C788R or the C1225G mutation failed to bind factor VIII and lacked high molecular weight multimers. Recombinant (r) vWF molecules having the C788R or C1225G mutation were expressed in COS-7 cells. Both rvWF C788R and rvWF C1225G exhibited significantly impaired secretion and failed to bind factor VIII. Recombinant vWF C788R in COS-7 culture medium showed a severe reduction in high molecular weight multimers, whereas rvWF C1225G showed a very mild reduction in high molecular weight multimers when compared with wild-type rvWF. (Blood. 2000;95:2000-2007) PMID- 10706868 TI - The anticoagulant factor, protein S, is produced by cultured human vascular smooth muscle cells and its expression is up-regulated by thrombin. AB - The anticoagulant factor protein S is a secreted vitamin K-dependent gamma carboxylated protein that is mainly made in the liver. Protein S is homologous to the growth arrest specific protein, Gas6, the expression of which is up-regulated in cultured fibroblasts upon serum withdrawal. We report here the synthesis and secretion of protein S by cultured human vascular smooth muscle cells (HVSMCs). Western blot analysis revealed that similar amounts of protein S are secreted by both growing and growth-arrested HVSMCs. HVSMC-derived protein S was found to be gamma-carboxylated as it was precipitated by barium citrate and was shown to possess protein C cofactor activity. Treatment with the vitamin K antagonist warfarin led to the accumulation of intracellular undercarboxylated protein S forms that were rapidly secreted upon the reintroduction of vitamin K. Northern blotting analysis showed that cultured HVSMCs express a protein S transcript. The expression of protein S messenger RNA was unaffected by either warfarin, growth arrest, or various VSMC mitogens, such as platelet-derived growth factor-BB, basic fibroblast growth factor, transforming growth factor-beta, or hepatocyte growth factor. Thrombin, however, induced an up-regulation of protein S expression at both messenger RNA and protein levels. The evidence we provide for protein S secretion by cultured HVSMCs and its up-regulation by thrombin, together with earlier reports showing that protein S acts as a mitogen for these cells, suggests that, in addition to its known role in regulating blood clotting, protein S may also be an important autocrine factor in the pathophysiology of the vasculature. (Blood. 2000;95:2008-2014) PMID- 10706869 TI - Tumor-induced apoptosis of T lymphocytes: elucidation of intracellular apoptotic events. AB - Our recent studies suggest that human squamous cell carcinoma of the head and neck (SCCHN) is capable of activating an intrinsic mechanism of programmed-cell death in interacting lymphocytes in situ and in vitro. The current study used Jurkat T-cell line as a model to investigate intracellular apoptotic events in T cells interacting with SCCHN. Apoptosis induced in T lymphocytes by tumor cells was in part Fas-mediated, since it was partially, but significantly, inhibited in the presence of anti-Fas ligand Ab or in Fas-resistant Jurkat cells. The synthetic caspase inhibitors, N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone (Z-VAD-FMK) and N-benzyloxycarbonyl-Asp-glu-Val-Asp-fluoromethyl ketone (Z-DEVD FMK), effectively blocked apoptosis of Jurkat cells co-incubated with SCCHN cell lines, suggesting the involvement of caspases in tumor-induced apoptosis of lymphocytes. Overexpression of CrmA, an inhibitor of caspase-1 and caspase-8, partially inhibited tumor-induced T-cell death. Caspase-8 and caspase-3 were identified as effector molecules in the execution of tumor-induced T-cell death, since the proform enzymes were processed into active subunits during co incubation of T cells with tumor cells. Furthermore, co-incubation with tumor cells resulted in cleavage of poly(ADP-ribose) polymerase (PARP), a common caspase-3 substrate, and in cleavage of TcR-zeta chain, shown by us to be a T cell specific caspase-3 substrate. Overexpression of Bcl-2 did not provide protection of T cells from SCCHN-induced DNA degradation. Instead, the Bcl-2 protein was cleaved in the target T cells during their co-incubation with tumor cells. These findings demonstrate that tumor cells can trigger in T lymphocytes caspase-dependent apoptotic cascades, which are not effectively protected by Bcl 2. (Blood. 2000;95:2015-2023) PMID- 10706870 TI - Cyclophosphamide induces type I interferon and augments the number of CD44(hi) T lymphocytes in mice: implications for strategies of chemoimmunotherapy of cancer. AB - In a previous study, we reported that a single injection of cyclophosphamide (CTX) in tumor-bearing mice resulted in tumor eradication when the animals were subsequently injected with tumor-sensitized lymphocytes. Notably, CTX acted by inducing bystander effects on T cells, and the response to the combined CTX/adoptive immunotherapy regimen was inhibited in mice treated with antibodies to mouse interferon (IFN)-alpha/beta. In the present study, we have investigated whether CTX induced the expression of type I IFN, and we have characterized the CTX effects on the phenotype of T cells in normal mice. CTX injection resulted in an accumulation of type I IFN messenger RNA in the spleen of inoculated mice, at 24 to 48 hours, that was associated with IFN detection in the majority of the animals. CTX also enhanced the expression of the Ly-6C on spleen lymphocytes. This enhancement was inhibited in mice treated with anti-type I IFN antibodies. Moreover, CTX induced a long-lasting increase in in vivo lymphocyte proliferation and in the percentage of CD44(hi)CD4(+) and CD44(hi)CD8(+ )T lymphocytes. These results demonstrate that CTX is an inducer of type I IFN in vivo and enhances the number of T cells exhibiting the CD44(hi) memory phenotype. Since type I IFN has been recently recognized as the important cytokine for the in vivo expansion and long-term survival of memory T cells, we suggest that induction of this cytokine may explain at least part of the immunomodulatory effects observed after CTX treatment. Finally, these findings provide a new rationale for combined treatments with CTX and adoptive immunotherapy in cancer patients. (Blood. 2000;95:2024-2030) PMID- 10706871 TI - The opioid antagonist naloxone induces a shift from type 2 to type 1 cytokine pattern in BALB/cJ mice. AB - Opioid peptides affect different immune functions. We present evidence that these effects could be mediated by the modulation of T(H)1/T(H)2 cytokine production. BALB/cJ mice were immunized with 50 or 100 microg of the protein antigen keyhole limpet hemocyanin (KLH), and treated acutely or chronically with the opioid antagonist naloxone. One and 2 weeks after immunization, the production of cytokines by splenocytes was evaluated by in vitro restimulation with KLH. The acute and chronic treatment with the opioid receptor antagonist naloxone decreased the production of interleukin (IL)-4 by splenocytes of BALB/cJ mice. In contrast, IL-2 and interferon-gamma levels increased after naloxone treatment. Finally, the opioid antagonist diminished the serum immunoglobulin G anti-KLH antibody titers. These results suggest that naloxone increases T(H)1 and decreases T(H)2 cytokine production. The effect of naloxone could be ascribed to the removal of the regulatory effects exerted by endogenous opioid peptides, which could therefore activate T(H)2 and suppress T(H)1 cytokines. (Blood. 2000;95:2031-2036) PMID- 10706872 TI - A critical role for PI 3-kinase in cytokine-induced Fcalpha-receptor activation. AB - Fc-receptors, such as FcalphaR and FcgammaRII, play an important role in leukocyte activation, and rapid modulation of ligand binding ("activation") is critical for receptor regulation. We have previously demonstrated that ligand binding to Fc-receptors on human eosinophils is dependent on cytokine stimulation. Utilization of pharmacological inhibitors provided evidence that the phenomenon of interleukin (IL)-5 induced immunoglobulin A (IgA) binding to human eosinophils requires activation of phosphatidylinositol 3-kinase (PI3K). However, eosinophils are refractory to manipulation by molecular techniques such as DNA transfection or viral infection. Here we utilize an IL-3 dependent pre-B cell line to investigate the molecular mechanism of cytokine-mediated ligand binding to FcalphaR. In this system, IgA binding is dependent on IL-3, similarly to the requirement for IL-5 of eosinophils. We show that IL-3-mediated activation of FcalphaR (CD89) requires the activation of PI3K, independent of p21ras activation. Co-expression of dominant negative (triangle upp85) and active (p110_K227E) forms of PI3K demonstrate that the affinity switch regulating FcalphaR activation requires PI3K. Moreover, overexpression of PI3K is both necessary and sufficient for activation of FcalphaR. Furthermore, we show that IL 3/IL-5/GM-CSF induced inside-out signaling pathways activating FcalphaR require the involvement of protein kinase C downstream of PI3K. Finally, we show that these inside-out signaling pathways responsible for Fcalpha-receptor modulation require CD89, independent of its association with the FcRgamma chain. (Blood. 2000;95:2037-2043) PMID- 10706873 TI - Engagement of the alpha2beta1 integrin inhibits Fas ligand expression and activation-induced cell death in T cells in a focal adhesion kinase-dependent manner. AB - T-cell receptor (TCR)-mediated apoptosis, also known as activation-induced cell death (AICD), plays an important role in the control of immune response and in the development of T-cell repertoire. Mechanistically, AICD has been largely attributed to the interaction of Fas ligand (Fas-L) with its cell surface receptor Fas in activated T cells. Signal transduction mediated by the integrin family of cell adhesion receptors has been previously shown to modulate apoptosis in a number of different cell types; in T cells, integrin signaling is known to be important in cellular response to antigenic challenge by providing a co stimulatory signal for TCR. In this study we demonstrate that signaling via the collagen receptor alpha2beta1 integrin specifically inhibits AICD by inhibiting Fas-L expression in activated Jurkat T cells. Engagement of the alpha2beta1 integrin with monoclonal antibodies or with type I collagen, a cognate ligand for alpha2beta1, reduced anti-CD3 and PMA/ionomycin-induced cell death by 30% and 40%, respectively, and the expression of Fas-L mRNA by 50%. Further studies indicated that the alpha2beta1-mediated inhibition of AICD and Fas-L expression required the focal adhesion kinase FAK, a known component in the integrin signaling pathways. These results suggest a role for the alpha2beta1 integrin in the control of homeostasis of immune response and T-cell development. (Blood. 2000;95:2044-2051) PMID- 10706874 TI - Nerve growth factor functions as a chemoattractant for mast cells through both mitogen-activated protein kinase and phosphatidylinositol 3-kinase signaling pathways. AB - Despite being a well-characterized neurotrophic factor, nerve growth factor (NGF) influences survival, differentiation, and functions of mast cells. We investigated whether NGF was able to induce directional migration of rat peritoneal mast cells (PMCs). NGF clearly induced chemotactic movement of PMCs in a dose-dependent manner with the drastic morphological change and distribution of F-actin, which was completely blocked by pretreatment with Clostridium botulinum C(2) toxin, an actin-polymerization inhibitor. Because PMCs constitutively express the NGF high-affinity receptor (TrkA) with a tyrosine kinase domain, we focused on downstream effectors in signaling cascades following the TrkA. NGF rapidly activated both mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K), and the addition of inhibitors specific for MAPK kinase and PI3K suppressed cell migration and these signals. In the coculture system with PMCs and fibroblasts, which produce biologically active NGF, directional migration of PMCs to fibroblasts was observed, and the addition of anti-NGF polyclonal antibodies significantly suppressed the migration of PMCs. These findings suggested that NGF initiated chemotactic movement of PMCs through both MAPK and PI3K signaling pathways following TrkA activation. Thus, locally produced NGF may play an important role in mast cell accumulation in allergic and nonallergic inflammatory conditions. (Blood. 2000;95:2052-2058) PMID- 10706875 TI - Estrogen influences the differentiation, proliferation, and survival of early B lineage precursors. AB - B lymphocyte production in murine bone marrow is negatively regulated by sex steroids and the aim of this study was to identify early hormone sensitive checkpoints. Estrogen (E2) treatment reduced cmu(+) pre-B cells, a change that occurred concomitantly with decreased Ig gene rearrangements and rag-1 transcripts. Estrogen decreased B lineage precursors in Ig transgenic mice, demonstrating that hormonal regulation is independent of the recombination process. B lineage precursors in Bcl-2 transgenic mice were resistant to estrogen treatment, suggesting that life/death decisions are involved in hormonal regulation. A previously uncharacterized population of CD43(-)cmu(-) B lineage precursors was identified in normal, Ig transgenic, and RAG(-/-) mice after estrogen treatment, revealing that down-regulation of CD43 can occur independent of Ig heavy chain expression. These cells expressed transcripts for both tdt and bcl-2, characteristics of early B-cell precursors. BrdU incorporation analysis revealed that the mitotic activity of early B-lineage cells is reduced in hormone treated mice. We conclude that sex steroids modulate the production of B-lineage cells by influencing the differentiation, proliferation, and survival of early B cell precursors. These findings are informative about mechanisms of hormonal regulation, as well as the significance of some differentiation-related events. (Blood. 2000;95:2059-2067) PMID- 10706876 TI - Inducible loss of NF-kappaB activity is associated with apoptosis and Bcl-2 down regulation in Epstein-Barr virus-transformed B lymphocytes. AB - The Epstein-Barr virus (EBV)-encoded latent membrane protein-1 induces NF-kappaB activity by targeting IkappaBalpha. To understand the role of NF-kappaB activation in EBV-related oncogenesis, we have subcloned mutated IkappaBalpha(32/36A) cDNA into a pHEBo vector containing doxycycline regulatory sequences and stably transfected this construct into a lymphoblastoid cell line. Two tightly regulated clones were obtained in which IkappaBalpha(32/36A) was inducible in a doxycycline dose-dependent manner. Levels of inducible IkappaBalpha(32/36A) peaked at day 2. Inhibition of NF-kappaB activity was closely correlated with levels of inducible IkappaBalpha(32/36A). Levels of 3 well-known NF-kappaB-dependent genes, CD54, p105, and endogenous IkappaBalpha, were decreased when IkappaBalpha(32/36A) was induced, and the growth of IkappaBalpha(32/36A)-induced EBV-infected cells was slightly reduced. Loss of NF kappaB activity was associated with decreased Bcl-2 protein levels. Finally, the induction of apoptosis was strongly increased in IkappaBalpha(32/36A) overexpressing cells. Together these results show that it is possible to control IkappaBalpha(32/36A) levels, ie, NF-kappaB activity, in EBV-infected B lymphocytes using a doxycycline-inducible vector. Moreover, our results indicate that NF-kappaB can protect EBV-infected cells from apoptosis by Bcl-2. Finally, our results suggest that a cellular model with doxycycline-inducible IkappaBalpha(32/36A) may be useful in the identification of genuine NF-kappaB target genes in EBV-infected B cells. (Blood. 2000;95:2068-2075) PMID- 10706877 TI - Transforming properties of chimeric TEL-JAK proteins in Ba/F3 cells. AB - The involvement of the cytokine signaling pathway in oncogenesis has long been postulated. Recently, rearrangements of the gene encoding the tyrosine Janus kinase 2 (JAK2) have been reported in human leukemias indicating a direct JAK signal transduction and activator of transcription (STAT)-mediated leukemic process. The leukemia-associated TEL-JAK2 fusion protein is formed by the oligomerization domain of the translocated ets leukemia (TEL) protein fused to the catalytic domain of JAK2. TEL-mediated oligomerization results in a constitutive tyrosine kinase activity that, in turn, is able to confer growth factor independence to the murine hematopoietic interleukin-3 (IL-3)-dependent Ba/F3 cell line. Results of the present study indicate that fusion proteins containing the oligomerization domain of TEL and the tyrosine kinase domains of Jak1, Jak2, JAK3, or TYK2 share similar properties and are able to efficiently substitute for the survival and mitogenic signals controlled by IL-3, without concomitant activation of the IL-3 receptor. Electrophoretic mobility shift assays demonstrated Stat5 as the only activated Stat factor in TEL-Jak2- and TEL Jak1-expressing cells, whereas other Stats, namely Stat1 and Stat3, could be detected in TEL-JAK3-, TEL-TYK2-, and also in TEL-ABL-expressing Ba/F3 cells. High levels of expression of the Stat5-target genes pim-1, osm, and Cis were observed in all the cytokine-independent cell lines. Furthermore, the expression of a dominant negative form of Stat5A markedly interfered with the growth factor independence process mediated by TEL-Jak2 in Ba/F3 cells. Because the BCR-ABL and TEL-PDGFbetaR oncoproteins also activate Stat5, activation of this factor should be a crucial step in activated tyrosine kinase-mediated leukemogenesis. (Blood. 2000;95:2076-2083) PMID- 10706878 TI - A monoclonal antibody (MUM1p) detects expression of the MUM1/IRF4 protein in a subset of germinal center B cells, plasma cells, and activated T cells. AB - A new monoclonal antibody (MUM1p) was used to study the cell/tissue expression of human MUM1/IRF4 protein, the product of the homologous gene involved in the myeloma-associated t(6;14) (p25;q32). MUM1 was expressed in the nuclei and cytoplasm of plasma cells and a small percentage of germinal center (GC) B cells mainly located in the "light zone." Its morphologic spectrum ranged from that of centrocyte to that of a plasmablast/plasma cell, and it displayed a phenotype (MUM1(+)/Bcl-6(-)/Ki67(-)) different from that of most GC B cells (MUM1(-)/Bcl 6(+)/Ki67(+)) and mantle B cells (MUM1(-)/Bcl-6(-)/Ki67(-)). Polymerase chain reaction (PCR) analysis of single MUM1(+ )cells isolated from GCs showed that they contained rearranged Ig heavy chain genes with a varying number of V(H) somatic mutations. These findings suggest that these cells may represent surviving centrocytes and their progeny committed to exit GC and to differentiate into plasma cells. MUM1 was strongly expressed in lymphoplasmacytoid lymphoma, multiple myeloma, and approximately 75% of diffuse large B-cell lymphomas (DLCL B). Unlike normal GC B cells, in which the expression of MUM1 and Bcl-6 were mutually exclusive, tumor cells in approximately 50% of MUM1(+) DLCL-B coexpressed MUM1 and Bcl-6, suggesting that expression of these proteins may be deregulated. In keeping with their proposed origin from GC B cells, Hodgkin and Reed-Sternberg cells of Hodgkin's disease consistently expressed MUM1. MUM1 was detected in normal and neoplastic activated T cells, and its expression usually paralleled that of CD30. These results suggest that MUM1 is involved in the late stages of B-cell differentiation and in T-cell activation and is deregulated in DLCL-B. (Blood. 2000;95:2084-2092) PMID- 10706879 TI - Cytogenetic abnormalities in the myelodysplastic syndromes and occupational or environmental exposure. AB - Patients with myelodysplastic syndromes (MDS) have high frequencies of cytogenetic abnormalities and evidence is accumulating of associations between exposure history and primary MDS. The objective of this article is to examine the relationship between histories of occupational or environmental exposure and presence of cytogenetic abnormalities. A case control study of MDS patients estimated lifetime exposure to more than 90 potential hazards in 400 age, sex, and area of residence matched patient and control pairs. A parallel cytogenetics study undertaken at time of diagnosis, independently of any knowledge of exposure history, identified 75 cytogenetically abnormal and 139 normal (186 not studied). Odds ratios of MDS patients and their matched controls were compared for 3 groups: cytogenetically abnormal, normal, and not known. The odds ratios for all exposures combined were possibly higher among cytogenetically abnormal 2.0 (95% confidence interval 0.8-5.9) than among normal 1.0 (0.6-1.8). This pattern was observed for exposure to semimetals, abnormal 4.0 (0.4-195.1) and normal 0.5 (0.1 1.0) and inorganic dusts, 1.6 (0. 6-3.8) and 0.4 (0.1-1.4) respectively. The pattern was principally in abnormalities in chromosomes 5 and 7. For organic chemicals and radiation, the odds ratios for both cytogenetically abnormal and normal were marginally raised: organic 1.8 (0.6-6.0) and 1.3 (0.6-2.9), respectively, and radiation 1.7 (0.5-5.6) and 1.3 (0.4-4.7) respectively. For radiation, abnormalities were mostly in chromosome 8. This study of association between exposures and cytogenetics in primary MDS complements those previously reported in secondary MDS and may provide some insight into pathogenetic mechanisms that lead to development of MDS. (Blood. 2000;95:2093-2097) PMID- 10706880 TI - A recombinant bispecific single-chain antibody, CD19 x CD3, induces rapid and high lymphoma-directed cytotoxicity by unstimulated T lymphocytes. AB - Although bispecific antibodies directed against malignant lymphoma have been shown to be effective in vitro and in vivo, extended clinical trials so far have been hampered by the fact that conventional approaches to produce these antibodies suffer from low yields, ill-defined byproducts, or laborious purification procedures. To overcome this problem, we have generated a small, recombinant, lymphoma-directed, bispecific single-chain (bsc) antibody according to a novel technique recently described. The antibody consists of 2 different single-chain Fv fragments joined by a glycine-serine linker. One specificity is directed against the CD3 antigen of human T cells, and the other antigen-binding site engages the pan-B-cell marker CD19, uniformly expressed on the vast majority of B-cell malignancies. The construct was expressed in Chinese hamster ovary cells and purified by its C-terminal histioline tag. Specific binding to CD19 and CD3 was demonstrated by fluorescence-activated cell sorter analysis. By redirecting unstimulated primary human T cells derived from the peripheral blood against CD19-positive lymphoma cells, the bscCD19 x CD3 antibody showed significant cytotoxic activity at very low concentrations of 10 to 100 pg/mL and at effector to target cell ratios as low as 2:1. Moreover, strong lymphoma directed cytotoxicity at low antibody concentrations was rapidly induced during 4 hours even in experiments without any T-cell prestimulation. Thus, this particular antibody proves to be much more efficacious than the bispecific antibodies described until now. Therefore, the described bscCD19 x CD3 molecule should be a suitable candidate to prove the therapeutic benefit of bispecific antibodies in the treatment of non-Hodgkin lymphoma. (Blood. 2000;95:2098-2103) PMID- 10706881 TI - c-Myc hot spot mutations in lymphomas result in inefficient ubiquitination and decreased proteasome-mediated turnover. AB - The c-myc proto-oncogene encodes a short-lived transcription factor that plays an important role in cell cycle regulation, differentiation and apoptosis. c-myc is often rearranged in tumors resulting in deregulated expression. In addition, mutations in the coding region of c-myc are frequently found in human lymphomas, a hot spot being the Thr58 phosphorylation site, a mutation shown to enhance the transforming capacity of c-Myc. It is, however, still unclear in what way this mutation affects c-Myc activity. Our results show that proteasome-mediated turnover of c-Myc is substantially impaired in Burkitt's lymphoma cells with mutated Thr58 or other mutations that abolish Thr58 phosphorylation, whereas endogenous or ectopically expressed wild type c-Myc proteins turn over at normal rates in these cells. Myc Thr58 mutants expressed ectopically in other cell types also exhibit reduced proteasome-mediated degradation, which correlates with a substantial decrease in their ubiquitination. These results suggest that ubiquitin/proteasome-mediated degradation of c-Myc is triggered by Thr58 phosphorylation revealing a new important level of control of c-Myc activity. Mutation of Thr58 in lymphoma thus escapes this regulation resulting in accumulation of c-Myc protein, likely as part of the tumor progression. (Blood. 2000;95:2104-2110) PMID- 10706882 TI - Growth of FasL-bearing tumor cells in syngeneic murine host induces apoptosis and toxicity in Fas(+) organs. AB - In the current study, we investigated whether the growth of FasL-bearing tumor cells would induce apoptosis and toxicity in organs that express high level of Fas. Sera from C57BL/6 +/+ (wild-type) mice injected with syngeneic FasL(+) tumors, LSA, or EL-4, showed significantly higher levels of soluble FasL than that from the nontumor-bearing mice. Furthermore, the soluble FasL was functional inasmuch as the sera from tumor-bearing mice were able to induce apoptosis in Fas(+) but not Fas(-) targets. Histopathologic studies and in situ TUNEL assay to detect apoptosis were carried out in C57BL/6 +/+ (Fas(+)) or C57BL/6 lpr/lpr (Fas(-)) mice injected with syngeneic LSA and EL-4 tumor cells. The morphology of the liver and thymus from tumor bearing C57BL/6 +/+ mice showed marked damage and tissue destruction. In contrast, the liver and thymus from tumor-bearing C57BL/6 lpr/lpr mice showed minimal damage. Furthermore, the tumor-bearing C57BL/6 +/+, but not the C57BL/6 lpr/lpr, mice exhibited significant apoptosis in the liver and thymus. The FasL responsible for toxicity was tumor derived rather than host derived; tumor-bearing C57BL/6 gld/gld (FasL-defective) mice also exhibited significant apoptosis in the liver and thymus. Together, these data suggested that the in vivo growth of FasL-bearing tumor cells can induce significant apoptosis and toxicity in Fas(+) tissues of the host. Such toxicity may be mediated by the soluble FasL produced by tumor cells. (Blood. 2000;95:2111-2117) PMID- 10706883 TI - STAT5 activation contributes to growth and viability in Bcr/Abl-transformed cells. AB - The transcription factor STAT5 is constitutively tyrosine phosphorylated and activated after transformation of hematopoietic cells by p210Bcr/Abl. A truncated form of STAT5B (triangle upSTAT5; aa, 1-683) that lacks tyrosine 699 and the transcriptional activation domain was introduced into Ba/F3p210 cells under the control of a tetracycline-inducible promoter. Treatment of these cells with doxycycline, a tetracycline analogue, induced expression of triangle upSTAT5 and inhibited STAT5-dependent transcription. triangle upSTAT5 coprecipitated with STAT5 and decreased Bcr/Abl-dependent tyrosine phosphorylation of endogenous STAT5. Induction of triangle upSTAT5 inhibited growth of Ba/F3p210 cells (26%-52% of control levels at 4 days) but did not cause cell-cycle arrest. triangle upSTAT5 reduced viability of Ba/F3p210 cells and increased sensitivity of the cells to the cytotoxic drugs hydroxyurea and cytarabine. These results indicate that high-level expression of triangle upSTAT5, as achieved here by using a tetracycline-inducible promoter, inhibits STAT5 activity, reduces the growth rate of Ba/F3p210 cells by inhibiting viability, and results in increased sensitivity to chemotherapeutic drugs. It is therefore likely that STAT5 activation plays a role in the transformation of hematopoietic cell lines by p210Bcr/Abl. (Blood. 2000;95:2118-2125) PMID- 10706884 TI - A new ETV6/TEL partner gene, ARG (ABL-related gene or ABL2), identified in an AML M3 cell line with a t(1;12)(q25;p13) translocation. AB - The ETV6/TEL gene has been reported to fuse to PDGFRbetab MDS1/EVI1, BTL, ACS2, STL, JAK2, ABL, CDX2, TRKC, AML1, and MN1. Among them, PDGFRbeta, ABL, JAK2, and TRKC are tyrosine kinases (TK). We identified a novel ETV6 partner gene, ARG (ABL related gene or ABL2), another TK gene in a cell line established from a patient with acute myelogenous leukemia (AML-M3) with a t(15;17)(q22;q11.2) and a t(1;12)(q25;p13), which has the remarkable feature to differentiate to mature eosinophils in culture with all-trans retinoic acid and cytokines. The ETV6/ARG transcripts consisted of exon 1 to 5 of ETV6 and the 3' portion of ARG starting from exon 1B or exon 2, resulting in an open reading frame for a fusion protein consisting of the entire PNT oligomerization domain of ETV6 and all of the functional domains of ARG including the TK domain. This is the same protein structure as identified in the other ETV6 TK fusion proteins. The reciprocal ARG/ETV6 transcript was not expressed, and the normal ETV6 allele was not deleted or rearranged. Although the ABL is known to be involved in various human malignancies, ARG has not been involved in human malignancies despite its high homology to ABL. Thus, this is the first report showing involvement of ARG in human leukemia. The ETV6/ARG protein may be involved in the unique differentiation capacity of this cell line. (Blood. 2000;95:2126-2131) PMID- 10706885 TI - Granulocyte colony-stimulating factor receptor mutations in severe congenital neutropenia transforming to acute myelogenous leukemia confer resistance to apoptosis and enhance cell survival. AB - Patients with severe congenital neutropenia (SCN) are at increased risk for the development of acute myelogenous leukemia (AML). In the subset of patients with SCN that progresses to AML, acquired mutations in the receptor for granulocyte colony-stimulating factor (G-CSF) have been detected that result in the expression of truncated forms of the G-CSF receptor (G-CSFR) protein. G-CSFR truncation mutants from these patients transduce hyperproliferative growth responses. In this paper, we show that the most frequently isolated mutant G-CSFR form from patients with SCN/AML (delta716) confers resistance to apoptosis and prolongs cell survival through a mechanism involving Akt, a downstream target of PI3-kinase. G-CSF stimulation of cells expressing the G-CSFR truncation mutant induces sustained activation of Akt and prolonged phosphorylation of the pro apoptotic protein Bad, resulting in enhanced cell survival. Extension of cell survival allowing for sufficient time for the acquisition of additional oncogenic events may represent an important mechanism by which G-CSFR mutations contribute to leukemogenesis. These data provide further insight into the pathophysiologic contribution of G-CSFR mutations to AML. (Blood. 2000;95:2132-2137) PMID- 10706886 TI - Deletions of chromosome 5q13.3 and 17p loci cooperate in myeloid neoplasms. AB - Nonrandom interstitial deletions and monosomy of chromosomes 5, 7, and 17 in refractory myelodysplasia (MDS) and acute myelogenous leukemia (AML) suggest a multistep pathway that culminates in aggressive clinical course. Because cytogenetic studies frequently identify chromosome 5 and 17 deletions within a single clone, we searched for allele loss for 5q loci and TP53 gene mutations in the same leukemic samples. Cosegregating deletions of chromosomes 5 and 17 were found to specifically include the 5q13.3 interval between the loci D5S672 and D5S620/D5S626, a locus hypothesized to harbor a tumor suppressor gene(1) and the TP53 gene on 17p. A rare patient with secondary refractory MDS and an unbalanced translocation [der(5;17)], which resulted in deletions of the 5q13.3-qter and 17p loci, provided clues on the sequence of genetic alterations. Serial molecular analysis of this patient revealed a dysplastic clone with der(5;17), which gave rise to a leukemic clone on acquiring an inactivating mutation of TP53. Our findings are consistent with functional cooperation between a putative tumor suppressor gene at 5q13.3 that contributes toward the progression of early stages of MDS, and the TP53 gene when mutated, causes transformation to AML. (Blood. 2000;95:2138-2143) PMID- 10706887 TI - Inv(2)(p23q35) in anaplastic large-cell lymphoma induces constitutive anaplastic lymphoma kinase (ALK) tyrosine kinase activation by fusion to ATIC, an enzyme involved in purine nucleotide biosynthesis. AB - The non-Hodgkin lymphoma (NHL) subtype anaplastic large-cell lymphoma (ALCL) is frequently associated with a t(2;5)(p23;q35) that results in the fusion of the ubiquitously expressed nucleophosmin (NPM) gene at 5q35 to the anaplastic lymphoma kinase (ALK) gene at 2p23, which is not normally expressed in hematopoietic tissues. Approximately 20% of ALCLs that express ALK do not contain the t(2;5), suggesting that other genetic abnormalities can result in aberrant ALK expression. Here we report the molecular characterization of an alternative genetic means of ALK activation, the inv(2)(p23q35). This recurrent abnormality produces a fusion of the amino-terminus of 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase/IMP cyclohydrolase (ATIC), a bifunctional homodimeric enzyme that catalyzes the penultimate and final steps of de novo purine nucleotide biosynthesis, with the intracellular portion of the ALK receptor tyrosine kinase. RT-PCR analysis of 5 ALCL tumors that contained the inv(2) revealed identical ATIC-ALK fusion cDNA junctions in all of the cases. Transient expression studies show that the ATIC-ALK fusion transcript directs the synthesis of an approximately 87-kd chimeric protein that is localized to the cytoplasm, in contrast to NPM-ALK, which typically exhibits a cytoplasmic and nuclear subcellular distribution. ATIC-ALK was constitutively tyrosine phosphorylated and could convert the IL-3-dependent murine hematopoietic cell line BaF3 to cytokine-independent growth. Our studies demonstrate an alternative mechanism for ALK involvement in the genesis of NHL and suggest that ATIC-ALK activation results from ATIC-mediated homodimerization. In addition, expected decreases in ATIC enzymatic function in ATIC-ALK-containing lymphomas may render these tumors more sensitive to antifolate drugs such as methotrexate. (Blood. 2000;95:2144-2149) PMID- 10706888 TI - Recombination events between the p47-phox gene and its highly homologous pseudogenes are the main cause of autosomal recessive chronic granulomatous disease. AB - Chronic granulomatous disease (CGD) is an inherited disease caused by defects in the superoxide-generating nicotinamide adenine dinucleotide phosphate (NADPH) oxidase of phagocytes. Genetic lesions in any of 4 components of this antimicrobial enzyme have been detected. Family-specific mutations are found in 3 of 4 forms of CGD due to deficiencies of the gp91-phox, p22-phox, and p67-phox genes. In p47-phox-deficient CGD (autosomal recessive form A47 degrees ) patients, a GT deletion (triangle upGT) at the beginning of exon 2 of the p47 phox gene has been reported in 19 of 20 alleles. This GT deletion is also characteristic for the recently identified p47-phox pseudogenes. To explore a possible link between these findings, a sequence analysis of 28 unrelated, racially diverse A47 degrees CGD patients and 37 healthy individuals was performed. The GT deletion in exon 2 was present on all alleles in 25 patients. Only 3 patients but all healthy individuals contained the GTGT and triangle upGT sequences. A total of 22 patients carried additional pseudogene-specific intronic sequences on all alleles, either only in intron 1 or in intron 1 and intron 2, which lead to different types of chimeric DNA strands. It is concluded that recombination events between the p47-phox gene and its highly homologous pseudogenes result in the incorporation of triangle upGT into the p47-phox gene, thereby leading to the high frequency of GT deletion in A47 degrees CGD patients. (Blood. 2000;95:2150-2156) PMID- 10706889 TI - Scavenger receptors on liver Kupffer cells mediate the in vivo uptake of oxidatively damaged red blood cells in mice. AB - In vitro studies have shown that damaged red cells and apoptotic cells are efficiently phagocytosed by scavenger receptors from macrophages, even under non opsonizing conditions. Damaged red blood cells are in vivo effectively removed from the blood circulation, but the responsible receptor systems are largely unknown. We used a murine model in which (51)Cr-labeled oxidized red blood cells were injected intravenously, and the cellular uptake sites and the potential involvement of scavenger receptors were analyzed. The decay of damaged red cells was rapid (more than 50% removed within 10 minutes after injection), whereas native red cells were not cleared. The main site of uptake of damaged red cells was the liver Kupffer cells, which contained 24% of the injected dose at 10 minutes after injection. The blood decay and liver uptake were inhibited by typical ligands for scavenger receptors, such as polyinosinic acid, liposomes containing phosphatidylserine, oxidized low-density lipoprotein, and fucoidan, but not by polyadenosinic acid or liposomes without phosphatidylserine. Mice lacking scavenger receptors class A type I and II showed no significant decrease in the ability to take up damaged red cells from the circulation. We conclude that Kupffer cells are mainly responsible for the removal of damaged red cells from the blood circulation, a process mediated by polyinosinic acid- and phosphatidylserine-sensitive scavenger receptors, different from scavenger receptor class A type I and II. Our data indicate that scavenger receptors, as pattern-recognizing receptors, play an important role in vivo in the removal of apoptotic, damaged, or other unwanted cells from the blood circulation. (Blood. 2000;95:2157-2163) PMID- 10706890 TI - Dehydration of mature and immature sickle red blood cells during fast oxygenation/deoxygenation cycles: role of KCl cotransport and extracellular calcium. AB - Sickle red blood cells (RBC) become dehydrated as a consequence of potassium loss. This process depends at least partly on deoxygenation and may be influenced by the presence of oxygenation/deoxygenation cycles and the frequency of cycling. In this study, sickle RBC were subjected to approximately 180 oxygenation/deoxygenation cycles during 4 hours to evaluate RBC dehydration with cycle periods more similar to in vivo cycles than those in previous studies. A continuous-flow, steady-state apparatus circulated a dilute RBC suspension through gas-permeable silicone tubing with segments that were exposed to either nitrogen or ambient oxygen. The percentage of sickling and partial pressure of oxygen were measured by means of sampling ports in the deoxygenation and oxygenation regions. The density increase (dehydration) of young (transferrin receptor-positive) and mature (transferrin receptor-negative) RBC and the requirements for calcium and chloride were evaluated. Density increase correlated with the percentage of sickled cells at the deoxygenation sampling port and was observed only in the presence of calcium, thereby implicating the calcium dependent potassium channel (Gardos pathway). Density increase was not dependent on the presence of chloride, making it unlikely that KCl cotransport was an important pathway under these conditions. (Blood. 2000;95:2164-2168) PMID- 10706891 TI - Autologous transplantation of ex vivo expanded bone marrow cells grown from small aliquots after high-dose chemotherapy for breast cancer. AB - The collection of small aliquots of bone marrow (BM), followed by ex vivo expansion for autologous transplantation may be less morbid, and more cost effective, than typical BM or blood stem cell harvesting. Passive elimination of contaminating tumor cells during expansion could reduce reinoculation risks. Nineteen breast cancer patients underwent autotransplants exclusively using ex vivo expanded small aliquot BM cells (900-1200 x 10(6)). BM was expanded in media containing recombinant flt3 ligand, erythropoietin, and PIXY321, using stromal based perfusion bioreactors for 12 days, and infused after high-dose chemotherapy. Correlations between cell dose and engraftment times were determined, and immunocytochemical tumor cell assays were performed before and after expansion. The median volume of BM expanded was 36.7 mL (range 15.8-87.0). Engraftment of neutrophils greater than 500/microL and platelets greater than 20,000/microL were 16 (13-24) and 24 (19-45) days, respectively; 1 patient had delayed platelet engraftment, even after infusion of back-up BM. Hematopoiesis is maintained at 24 months, despite posttransplant radiotherapy in 18 of the 19 patients. Transplanted CD34(+)/Lin(-) (lineage negative) cell dose correlated with neutrophil and platelet engraftment, with patients receiving greater than 2.0 x 10(5) CD34(+)/Lin(-) cells per kilogram, engrafting by day 28. Tumor cells were observed in 1 of the 19 patients before expansion, and in none of the 19 patients after expansion. It is feasible to perform autotransplants solely with BM cells grown ex vivo in perfusion bioreactors from a small aliquot. Engraftment times are similar to those of a typical 1000 to 1500 mL BM autotransplant. If verified, this procedure could reduce the risk of tumor cell reinoculation with autotransplants and may be valuable in settings in which small stem cell doses are available, eg, cord blood transplants. (Blood. 2000;95:2169-2174) PMID- 10706892 TI - Allogeneic hematopoietic chimerism in mice treated with sublethal myeloablation and anti-CD154 antibody: absence of graft-versus-host disease, induction of skin allograft tolerance, and prevention of recurrent autoimmunity in islet allografted NOD/Lt mice. AB - We describe a tolerance-based stem cell transplantation protocol that combines sublethal radiation with transient blockade of the CD40-CD154 costimulatory pathway using an anti-CD154 antibody. With this protocol, we established hematopoietic chimerism in BALB/c mice transplanted with fully allogeneic C57BL/6 bone marrow. The percentage of donor-origin mononuclear cells in recipients was more than 99%. In addition, all chimeric mice treated with anti-CD154 antibody remained free of graft-versus-host disease (GVHD) and accepted donor-origin but not third-party skin allografts. It was similarly possible to create allogeneic hematopoietic chimerism in NOD/Lt mice with spontaneous autoimmune diabetes. Pancreatic islet allografts transplanted into chimeric NOD/Lt mice were resistant not only to allorejection but also to recurrence of autoimmunity. We conclude that it is possible to establish robust allogeneic hematopoietic chimerism in sublethally irradiated mice without subsequent GVHD by blocking the CD40-CD154 costimulatory pathway using as few as 2 injections of anti-CD154 antibody. We also conclude that chimerism created in this way generates donor-specific allograft tolerance and reverses the predisposition to recurrent autoimmune diabetes in NOD/Lt mice, enabling them to accept curative islet allografts. (Blood. 2000;95:2175-2182) PMID- 10706893 TI - Localization of biologically uncommon D-beta-aspartate-containing alphaA crystallin in human eye lens. AB - PURPOSE: Previous studies demonstrated that the Asp-151 residue of alphaA crystallin from human eye lens is stereoinverted to the biologically uncommon D isomer and isomerized to the beta-aspartyl residue (isoaspartate) with age. To detect the locality of the D-beta-Asp-containing peptide in aged human lens, we prepared a highly specific antibody against peptide Gly-Leu-D-beta-Asp-Ala-Thr which corresponds to positions 149-153 of human alphaA-crystallin using peptide Gly-Leu-D-beta-Asp-Ala-Thr-Gly-Leu-D-beta-Asp-Ala-Thr-Gly-Leu-D-beta- Asp-Ala-Thr (designated peptide 3R) as an immunogen. METHODS: Peptide 3R was synthesized with F-moc (9-fluorenylmethoxycarbonyl) solid phase chemistry and then the peptide was immunized in rabbits to generate antibody against peptide 3R. The antibody in rabbit serum was purified by affinity chromatography using peptide 3R and bovine alphaA-crystallin as ligands. The specificity and titer of antibody were checked by ELISA assay. We synthesized four kinds of peptide T18 (IQTGLDATHAER; corresponding to the amino acid sequences 146-157 in human alphaA-crystallin) in which Asp-151 residues were normal L-alpha-Asp, abnormal D-alpha-Asp, L-beta-Asp, and D-beta-Asp, respectively. The specificity of antibody was confirmed by ELISA using these peptides and utilized in immunohistochemistry. RESULTS: The antibody we prepared crossreacted specifically to D-beta-Asp-151-containing alphaA crystallin. Immunohistochemical staining of human lens with the antibody demonstrated that D-beta-Asp-151-containing alphaA-crystallin was predominantly localized in the core of aged human lens. CONCLUSIONS: The peptide 3R antibody clearly recognized the presence of racemized and isomerized Asp-151 in both protein solution and lens tissue obtained from aged human lens. PMID- 10706894 TI - Evaluation of human diacylglycerol kinase(iota), DGKI, a homolog of Drosophila rdgA, in inherited retinopathy mapping to 7q. AB - PURPOSE: To determine the genomic organization of diacylglycerol kinase(iota) and to test whether defects in this gene are present in individuals affected with autosomal dominant retinitis pigmentosa (adRP). Diacylglycerol kinase(iota) has been mapped to the RP10 locus on 7q and shows 49% sequence similarity to the Drosophila DGK2 rdgA gene. Since mutations in the DGK2 rdgA gene cause photoreceptor degeneration in Drosophila, it is possible that mutations in diacylglycerol kinase(iota) could be responsible for human retinal degeneration. METHODS: DNA sequence from genomic clones containing diacylglycerol kinase(iota) was compared with the cDNA sequence to identify intron/exon boundaries. Single strand conformational analysis and PCR product sequencing were used to screen members of one family previously mapped to the RP10 locus and 47 small unmapped families with autosomal dominant retinitis pigmentosa. RESULTS: Diacylglycerol kinase(iota) is divided into 35 exons with the initiation codon being present in exon 2. Mutational analysis found a missense change (Lys153Phe) in three adRP families; however, it did not segregate with disease in one of the families. Silent substitutions were seen in codons 865 and 875. Intronic variation was detected in the amplifications of exons 3,5,18, 28, and 32; these do not affect splice site consensus sequences. Typing of a polymorphic variant detected in intron 31 in members of the RP10 family gave a LOD score of -4.2 at 0% recombination. CONCLUSIONS: No evidence of disease-associated mutations was found in any of the samples tested. Based on the linkage data and mutation screening, diacylglycerol kinase(iota) is excluded as a candidate for the RP10 form of adRP and cannot be a frequent cause of other forms of adRP. Mutations in diacylglycerol kinase(iota) may yet be the cause of recessive forms of retinal degeneration in humans, either in homozygous or compound heterozygous forms. The data provided here will permit testing of this hypothesis. PMID- 10706895 TI - Heat-induced conformational change of human lens recombinant alphaA- and alphaB crystallins. AB - PURPOSE: To determine which component of lens alpha-crystallin is responsible for heat-induced transition, conformational change and high molecular weight (HMW) aggregation. METHODS: Recombinant alphaA- and alphaB-crystallins were used. Temperature dependent changes were probed by Trp fluorescence and circular dichroism (CD) measurements. HMW aggregates were induced by heating at 62 degrees C for 1-2 h and then cooling to room temperature. The nature of HMW aggregation was studied with fluorescent probes, 4,4'-dianilino-1, 1'-binaphthalene-5,5' disulfonic acid (bis-ANS) and thioflavin T (ThT). RESULTS: CD and Trp fluorescence revealed that alphaB-crystallin was more susceptible than alphaA crystallin to heat-induced conformational change and aggregation. At temperatures greater than 70 degrees C, alphaB-crystallin precipitated but alphaA-crystallin remained soluble. Both bis-ANS and ThT probes displayed increased fluorescence intensity with HMW aggregation, but the increase for bis-ANS was greater with alphaB-crystallin than with alphaA-crystallin, while the reverse was true for ThT. CONCLUSIONS: These results indicate that alphaB-crystallin is more susceptible than alphaA-crystallin to heat-induced conformational change and aggregation and are consistent with the notion that alphaA- and alphaB crystallins have different biochemical and biophysical properties in spite of their high degree of homology. PMID- 10706896 TI - Clinical and genetic features of Ehlers-Danlos syndrome type IV, the vascular type. AB - BACKGROUND: Ehlers-Danlos syndrome type IV, the vascular type, results from mutations in the gene for type III procollagen (COL3A1). Affected patients are at risk for arterial, bowel, and uterine rupture, but the timing of these events, their frequency, and the course of the disease are not well documented. METHODS: We reviewed the clinical and family histories of and medical and surgical complications in 220 index patients with biochemically confirmed Ehlers-Danlos syndrome type IV and 199 of their affected relatives. We identified the underlying COL3A1 mutation in 135 index patients. RESULTS: Complications were rare in childhood; 25 percent of the index patients had a first complication by the age of 20 years, and more than 80 percent had had at least one complication by the age of 40. The calculated median survival of the entire cohort was 48 years. Most deaths resulted from arterial rupture. Bowel rupture, which often involved the sigmoid colon, accounted for about a quarter of complications but rarely led to death. Complications of pregnancy led to death in 12 of the 81 women who became pregnant. The types of complications were not associated with specific mutations in COL3A1. CONCLUSIONS: Although most affected patients survive the first and second major complications, Ehlers-Danlos syndrome type IV results in premature death. The diagnosis should be considered in young people who come to medical attention because of uterine rupture during pregnancy or arterial or visceral rupture. PMID- 10706897 TI - Cigarette smoking and invasive pneumococcal disease. Active Bacterial Core Surveillance Team. AB - BACKGROUND: Approximately half of otherwise healthy adults with invasive pneumococcal disease are cigarette smokers. We conducted a population-based case control study to assess the importance of cigarette smoking and other factors as risk factors for pneumococcal infections. METHODS: We identified immunocompetent patients who were 18 to 64 years old and who had invasive pneumococcal disease (as defined by the isolation of Streptococcus pneumoniae from a normally sterile site) by active surveillance of laboratories in metropolitan Atlanta, Baltimore, and Toronto. Telephone interviews were conducted with 228 patients and 301 control subjects who were reached by random-digit dialing. RESULTS: Fifty-eight percent of the patients and 24 percent of the control subjects were current smokers. Invasive pneumococcal disease was associated with cigarette smoking (odds ratio, 4.1; 95 percent confidence interval, 2.4 to 7.3) and with passive smoking among nonsmokers (odds ratio, 2.5; 95 percent confidence interval, 1.2 to 5.1) after adjustment by logistic-regression analysis for age, study site, and independent risk factors such as male sex, black race, chronic illness, low level of education, and living with young children who were in day care. There were dose-response relations for the current number of cigarettes smoked per day, pack years of smoking, and time since quitting. The adjusted population attributable risk was 51 percent for cigarette smoking, 17 percent for passive smoking, and 14 percent for chronic illness. CONCLUSIONS: Cigarette smoking is the strongest independent risk factor for invasive pneumococcal disease among immunocompetent, nonelderly adults. Because of the high prevalence of smoking and the large population attributable risk, programs to reduce both smoking and exposure to environmental tobacco smoke have the potential to reduce the incidence of pneumococcal disease. PMID- 10706898 TI - Long-term outcome of fulminant myocarditis as compared with acute (nonfulminant) myocarditis. AB - BACKGROUND: Lymphocytic myocarditis causes left ventricular dysfunction that may be persistent or reversible. There are no clinical criteria that predict which patients will recover ventricular function and which cases will progress to dilated cardiomyopathy. We hypothesized that patients with fulminant myocarditis may have a better long-term prognosis than those with acute (nonfulminant) myocarditis. METHODS: We identified 147 patients considered to have myocarditis according to the findings on endomyocardial biopsy and the Dallas histopathological criteria. Fulminant myocarditis was diagnosed on the basis of clinical features at presentation, including the presence of severe hemodynamic compromise, rapid onset of symptoms, and fever. Patients with acute myocarditis did not have these features. The incidence of the end point of this study, death or heart transplantation, was ascertained by contact with the patient or the patient's family or by a search of the National Death Index. The average period of follow-up was 5.6 years. RESULTS: A total of 15 patients met the criteria for fulminant myocarditis, and 132 met the criteria for acute myocarditis. Among the patients with fulminant myocarditis, 93 percent were alive without having received a heart transplant 11 years after biopsy (95 percent confidence interval, 59 to 99 percent), as compared with only 45 percent of those with acute myocarditis (95 percent confidence interval, 30 to 58 percent; P=0.05 by the log rank test). Fulminant myocarditis was an independent predictor of survival after adjustments were made for age, histopathological findings, and hemodynamic variables. The rate of transplantation-free survival did not differ significantly between the patients considered to have borderline myocarditis and those considered to have active myocarditis according to the Dallas histopathological criteria. CONCLUSIONS: Fulminant myocarditis is a distinct clinical entity with an excellent long-term prognosis. Aggressive hemodynamic support is warranted for patients with this condition. PMID- 10706899 TI - High levels of coagulation factor XI as a risk factor for venous thrombosis. AB - BACKGROUND: Factor XI, a component of the intrinsic pathway of coagulation, contributes to the generation of thrombin, which is involved in both the formation of fibrin and protection against fibrinolysis. A deficiency of factor XI is associated with bleeding, but a role of high factor XI levels in thrombosis has not been investigated. METHODS: We determined factor XI antigen levels in the patients enrolled in the Leiden Thrombophilia Study, a large population-based, case-control study (with a total of 474 patients and 474 controls) designed to estimate the contributions of genetic and acquired factors to the risk of deep venous thrombosis. Odds ratios were calculated as a measure of relative risk. RESULTS: The age- and sex-adjusted odds ratio for deep venous thrombosis in subjects who had factor XI levels above the 90th percentile, as compared with those who had factor XI levels at or below that value, was 2.2 (95 percent confidence interval, 1.5 to 3.2). There was a dose-response relation between the factor XI level and the risk of venous thrombosis. Adjustment of the odds ratios for other risk factors such as oral-contraceptive use, homocysteine, fibrinogen, factor VIII, female sex, and older age did not alter the result. Also, when we excluded subjects who had known genetic risk factors for thrombosis (e.g., protein C or S deficiency, antithrombin deficiency, the factor V Leiden mutation, or the prothrombin G20210A mutation), the odds ratio remained the same, indicating that the risk of venous thrombosis associated with elevated levels of factor XI was not the result of one of the known risk factors for thrombosis. CONCLUSIONS: High levels of factor XI are a risk factor for deep venous thrombosis, with a doubling of the risk at levels that are present in 10 percent of the population. PMID- 10706900 TI - Images in clinical medicine. Retroperitoneal hemorrhage. PMID- 10706901 TI - The evaluation and management of bradycardia. PMID- 10706902 TI - Vancomycin-resistant enterococcal infections. PMID- 10706903 TI - Case records of the Massachusetts General Hospital. Weekly clinicopathological exercises. Case 7-2000. A 23-year-old man with hemolytic anemia and bloody diarrhea. PMID- 10706904 TI - Ehlers-Danlos syndrome. PMID- 10706905 TI - Smoking and pneumococcal infection. PMID- 10706906 TI - Fulminant myocarditis. PMID- 10706907 TI - Correction: Case Records of the Massachusetts General Hospital (Case 4-2000). PMID- 10706908 TI - Correction: Effects of An Angiotensin-Converting-Enzyme Inhibitor, Ramipril, on Cardiovascular Events in High-Risk Patients. PMID- 10706909 TI - Smooth muscle tumors of the gastrointestinal tract: introduction. PMID- 10706910 TI - Musculature of the alimentary tract. AB - Leiomyomas and leiomyosarcomas are derived from smooth muscle tissues. Smooth muscle cells also surround the blood vessels that supply the alimentary tract. These cells have at times been said to contribute to the formation of smooth muscle tumors in the alimentary canal. With such an abundance of smooth muscle, there is little reason at present to implicate the smooth muscle elements of blood vessel walls. PMID- 10706912 TI - Tumors of the esophagus. AB - This collective review includes all available case reports of smooth muscle (stromal) tumors of the esophagus in the world literature. Compiling this review, we endeavored to examine cumulative and recently collected data of both benign and malignant esophageal smooth muscle tumors found in the literature spanning the period from 1875 to 1996, which totaled 1679 leiomyomas (LMs) and 165 leiomyosarcomas (LMSs). The peak age of occurrence of benign smooth muscle tumors in the esophagus was found to be between the ages of 30 and 59, whereas the highest frequency of malignant tumors was seen later in life, during the decade from age 60 to 69. The most common location of both LMs and LMSs was the lower third of the esophagus. Their patterns of growth differed; LMs were more likely to grow intramurally, and LMSs were predominantly intraluminal. Most patients with LMs presented with dysphagia and pain or discomfort; patients with LMSs additionally commonly complained of weight loss. As with smooth muscle tumors of other areas of the gastrointestinal tract, the duration of symptoms averaged 1 month to 1 year, and malignant tumors grew to larger sizes than benign neoplasms. Approximately one-third of LMSs had metastasized at diagnosis, and there was a 5 year survival rate of approximately 20%. PMID- 10706911 TI - Tumors of the oral cavity and pharynx. AB - We have compiled all the reported cases of smooth muscle (stromal) tumors of the oral cavity and pharynx from 1884 through 1996. Our collective data included 139 leiomyomas (LM) and 68 leiomyosarcomas (LMS); but because we did not have sufficient information for 13 cases of LM, we report on only 126; and we report on only 66 of 68 cases of LMS. The peak age of incidence was 40 to 49 years for benign tumors and 50 to 59 years for malignant lesions, with the incidence in men slightly predominating over that in women. The most common sites of LM of the oral cavity and pharynx were the lips, tongue, and hard and soft palate. The most common sites of LMS included the maxilla and mandible. More than 40% of LMs presented as an intraoral mass, and more than half were known to be present for longer than 1 year. About 10% presented with pain, difficulty chewing or swallowing, swelling, toothache or loose teeth, or a combination of these symptoms. Patients with LMS were much more likely to have obvious symptoms of shorter duration, and one-third presented with pain or swelling (or both). Other relatively common symptoms of LMS included tenderness, interference with dentures, or an intraoral mass. In this review, there were almost twice as many LMs as LMSs, which was consistent with smooth muscle tumors found in other areas of the gastrointestinal tract. PMID- 10706913 TI - Tumors of the stomach. AB - This collective review includes all available case reports of smooth muscle (stromal) tumors of the stomach in the world literature from 1762 to 1996. It updates our previous review from 1767 to 1959. Overall, we identified 2189 patients with leiomyoma (LM) and 1594 with leiomyosarcoma (LMS). The peak age of incidence of LM was 50 to 59 years, while LMS was most frequently seen between ages 60 and 69. Women were more likely to develop LM, and men more commonly presented with malignant smooth muscle tumors of the stomach. Concerning the patterns of growth, LMs were more likely to grow intraluminally (endogastric), whereas LMSs were predominantly exogastric. The most common site of LMs was on the anterior or posterior wall of the body of the stomach; LMSs were most likely found along the greater curve. The presenting symptoms of both types of smooth muscle tumors were similar; in decreasing order of frequency they were bleeding, pain, palpable mass, and weight loss. Interestingly, there was no correlation between the size of the tumor and signs or symptoms of bleeding, pain, weight loss, or ulceration, although patients with LMSs were more likely to report weight loss than patients with benign tumors. For LMS, there seemed to be no correlation between tumor size or location and rate of metastasis, although the tumors that grew in a dumbbell shape (i.e., both intraluminally and extraluminally) had a higher frequency of metastasis than other growth patterns. Overall, the rate of metastasis at diagnosis was 35.4%, with the liver, spleen, and regional lymph nodes the most common sites. PMID- 10706914 TI - Tumors of the small intestine. AB - This collective review includes all available case reports and series of smooth muscle (stromal) tumors of the small intestine in the world literature from 1881 to 1996. We identified 1074 patients with leiomyoma (LM) and 1689 with leiomyosarcoma (LMS). Our purpose was to update our previous review, which encompassed case reports and series from 1881 to 1959, which included 350 LMs and 257 LMSs. The peak incidence of smooth muscle tumors in the small intestine in both male and female patients was between the ages of 50 and 59. Most commonly, the presenting complaint was gastrointestinal bleeding. Computed tomography was found to detect LM and LMS most successfully and had the additional advantage of locating metastatic disease. The jejunum contained the highest numbers of smooth muscle tumors, followed by the ileum and then the duodenum, with malignant lesions in all locations typically attaining larger diameters than benign tumors. The overall rate of metastatic spread of LMS ranged from 24% to 50%, with the liver being most commonly involved. Unlike other sarcomas, both hematogenous and lymphatic spread were common. The 5-year survival of 705 patients with LMS from 22 series was 27. 8%. For both benign and malignant smooth muscle tumors of the small intestine, surgery remains the treatment of choice, with little efficacy reported for irradiation, chemotherapy, or both. PMID- 10706915 TI - Tumors of the appendix and colon. AB - This collective review includes all available case reports of smooth muscle (stromal) tumors of the appendix and large intestine in the world literature. When compiling this review, we endeavored to examine cumulative as well as recently collected data on both benign and malignant smooth muscle tumors spanning the period 1875 to 1996. In total, there were reports of 331 leiomyomas (LMs) and 263 leiomyosarcomas (LMSs). The peak age of incidence of LM was 30 to 39 years, and the peak age of incidence of LMSs was 50 to 59 years. The female/male ratio was slightly higher for LM, and the male/female ratio was higher for LMS. The descending colon and sigmoid colon were the most common sites of both benign and malignant smooth muscle tumors. The growth of LMs most often occurred extraluminally, whereas LMSs tended to grow within the lumen of the colon. With both tumor types pain was the most frequent presenting complaint, followed less commonly by complaints of a palpable mass or gastrointestinal bleeding. LMSs tended to be larger at diagnosis than LMs, though the duration of symptoms for both types of tumor was most often reported to be between 1 month and 1 year. Finally, LMSs were found to metastasize mo PMID- 10706916 TI - Tumors of the rectum and anal canal. AB - This collective review includes all available case reports of smooth muscle (stromal) tumors of the rectum and anal canal in the world literature. When compiling this review we endeavored to present cumulative and recently collected data of both benign and malignant smooth muscle tumors found in the literature spanning the period from 1881 to 1996, which totaled 432 leiomyomas (LMs) and 480 leiomyosarcomas (LMSs) of the anus and rectum. The peak age of frequency for LMs was 40 to 59 years and 50 to 69 years for LMSs; men were slightly more likely to develop both benign and malignant anorectal smooth muscle tumors than women. Intraluminal growth of both LMs and LMSs was more frequently seen than extraluminal or intramural patterns, and tumors were more likely to be found in the rectum than in the anus. Patients with LMs presented most commonly with gastrointestinal (GI) bleeding, a palpable mass, or anorectal pain. As with smooth muscle tumors in other areas of the alimentary tract, symptoms likely persisted for less than 1 year prior to diagnosis. As was also the case for these neoplasms in other GI locations, LMSs tended to be larger than LMs. Approximately 20% of LMSs reported from 1881 to 1996 had metastasized at diagnoses. The local recurrence rate for resectable tumors was more than 80%, exceeding the propensity of LMSs in other areas of the GI tract to recur. PMID- 10706917 TI - Influence of two-dimensional and three-dimensional imaging on endoscopic bowel suturing. AB - Several three-dimensional (3-D) video-endoscopic systems have been introduced in surgical practice to enhance depth perception during minimal access surgery (MAS), but the facilitation of endoscopic manipulations by the current 3-D systems remains unproved. The aim of the study was to investigate the influence of 2-D and 3-D imaging modalities on intracorporeal suturing. The standard task consisted of suture closure of 60 mm enterotomies made in porcine small bowel with continuous seromuscular 3/0 Polysorb. Ten experienced surgeons participated in the study. The imaging systems were Storz (2-D), Welch Allyn (3-D), and Zeiss (as both 2-D and 3-D). Each surgeon performed two tasks with each modality in a random sequence. The outcome measures were execution time, suture line leakage pressure, and suture placement score. In addition, the participating surgeons assigned subjective scores on the image quality and the adverse effects of the imaging systems. There was no significant difference in the execution time, leakage pressure, and suture placement score among the various imaging modalities. Depth perception was rated as similar with 2-D and 3-D imaging. Surgeons experienced visual strain with the three systems, but it was rated higher with 3-D imaging. With the current technology, we have not documented any significant difference in task efficiency and quality of endoscopic bowel suturing by trained surgeons between 2-D and 3-D imaging systems. PMID- 10706918 TI - Free radical scavenging activity of fullerenol on the ischemia-reperfusion intestine in dogs. AB - Fullerenol, a water-soluble C(60)-fullerene derivative, has been demonstrated to have the capability to scavenge free radicals in vitro and in vivo. The purpose of this study was to investigate whether fullerenol can scavenge the free radicals that are massively induced during ischemia-reperfusion (I/R) injury of the small intestine, either preventively or therapeutically. Clamping the superior mesenteric artery and vein for 60 minutes to induce I/R injury was performed on male mongrel dogs. Thirty dogs were divided into three groups (10 in each): The control (C) group received no medication; the preventive (P) group received fullerenol (1 mg/kg) intravenously 30 minutes before ischemia; the therapeutic (T) group received the same dose of fullerenol immediately after reperfusion. This study was an experimental randomized trial. Intestinal segments were obtained 10, 20, 30, and 60 minutes after reperfusion; and blood samples and specimens of major organs were taken 60 minutes after reperfusion. Concentrations of lipid peroxidation products, including conjugated diene (CD) and malondialdehyde (MDA), and the level of glutathione (GSH) in intestinal tissue were determined. Serum indicators of liver and renal function were measured. Histologic examination of the small intestine and major organs were also performed. A significant increase in intestinal MDA and CD contents was detected at 30 and 60 minutes after reperfusion. The tissue GSH content, in contrast, was decreased 60 minutes after reperfusion. Administration of fullerenol diminished these changes both preventively and therapeutically. Liver and renal functions were within normal limits in all groups. Moreover no obvious histopathologic additional damage could be found in either the P or the T group. It is suggested that fullerenol can be considered a powerful scavenger for the free radicals induced by I/R injury of the small intestine. PMID- 10706919 TI - Significance of plasma thymosin alpha1 measurements in gastric cancer patients. AB - Thymosin alpha1 is a cleavage product of prothymosin alpha. Expression of prothymosin alpha increases during cell proliferation. Thymosin alpha1, measurable in plasma, may be an indicator of cell proliferation especially if a cancer is present. In this report we investigated the relation between the clinical behavior of gastric cancer and the plasma thymosin alpha1 level. Plasma thymosin alpha1 was measured in 52 gastric cancer patients using a newly developed radioimmunoassay. Twenty-one tumors and lymph nodes were examined for thymosin alpha1 using immunohistochemistry. The plasma thymosin alpha1 level was higher in gastric cancer patients than in normal volunteers; and it was higher in patients with lymph node involvement than in those with negative nodes. Immunohistochemical study of thymosin alpha1 showed positivity in 52% of gastric cancers and 100% of lymph node metastasis. We concluded that a high level of plasma thymosin alpha1 suggests aggressive behavior of a gastric cancer, such as lymph node involvement. PMID- 10706920 TI - Surgery: independent prognostic factor in curable and far advanced gastric cancer. AB - The hospital records of 639 patients affected by primary gastric cancer who were consecutively admitted to our unit during the period 1981-1995 were reviewed. Overall 220 underwent total gastrectomy (38 palliative), 12 had resection of the gastric stump, 195 had distal subtotal gastrectomy (55 palliative), 78 had bypass procedures, 72 had explorative laparotomy, and 62 had no operation. Univariate and multivariate analyses were used to evaluate 5-year survival with respect to the main clinical, pathologic, and treatment variables after both curative and palliative treatments. Overall the 5-year survival after curative treatment (320 patients-operative mortality excluded) was 55.5%: 91.1% for stage IA, 71.5% IB, 62.4% II, 37.5% IIIA, 31.5% IIIB. Among patients who underwent palliative treatment 5-year survival was 13.1% after gastric resection (total or distal subtotal), 4.9% after the bypass procedures, 0 after explorative laparotomy, and 0 after no operation. Univariate and multivariate survival analyses showed that variables independently associated with poor survival were advanced stage, upper location and D1 lymphadenectomy after curative treatment, tumor spread to distant sites, and nonresectional surgery after palliative treatment. Multivariate analysis showed that even though survival with gastric cancer depends on predetermined factors, the type of surgery can have a significant effect on prognosis after both curative and palliative treatment. PMID- 10706921 TI - Surgical mortality, survival, and quality of life after resection for gastric cancer in the elderly. AB - Although there were some studies on clinicopathologic characteristics, operative morbidity, and mortality in elderly patients with gastric cancer, no reports have specifically focused on survival and quality of life after resection. A total of 433 patients aged >/= 65 years (1987-1994) who underwent gastric resection for gastric adenocarcinoma were studied. Two groups were considered: patients aged 65 to 74 years and those > 74 years. Most of the patients (78.1%) had advanced diseases, and nearly half (41. 3%) had associated chronic disease(s). Resections with curative intention were performed in 362 patients (83.6%). The overall operative morbidity rate was 21.7% and mortality rate 5.1%. Although operative procedures were similar in both groups, patients aged >74 years had a higher mortality rate than those aged 65 to 74 years (10. 1% vs. 3.5%; p = 0.034). Age and extent of gastric resection were two independent factors negatively affecting mortality. The cumulative survival rates for patients who underwent curative resection were 86.2%, 72.4%, 67.2%, 62.9%, and 60.0% at 1, 2, 3, 4, and 5 years, respectively. Nearly all patients (96%) after surgery had normal work and daily activities. Some patients appeared to lack energy (16%) or experienced a period of anxiety or depression. There was no statistical difference in survival and quality of life assessed by the Spitzer index after curative resection between the two groups. Therefore resection with curative intention can be performed for the elderly with acceptable morbidity and mortality rates, possible long-term survival, and good quality of life, but a limited operation should be considered in the very elderly patients. PMID- 10706922 TI - Carcinoma of the gastric cardia: surgical practices and short-term operative results in a defined Swedish population. AB - Adenocarcinoma of the gastric cardia has one of the most rapidly increasing incidence rates of all tumors in Western countries. The aim of this population based investigation was to study surgical practices and postoperative morbidity and mortality during routine hospital care. The study comprised 176 patients given a new diagnosis of adenocarcinoma of the gastric cardia from February 1, 1989 to the January 31, 1995 in five Swedish counties. The tumor was resectable in 100 (57%) patients (in 36% of the women and 64% of the men), but only 46% of all patients could be offered a potentially curative operation. A total gastrectomy was performed in 54 patients and a proximal gastric resection in 44. Postoperative complications occurred in 39%: in 20% of the patients under age 60 years and in 47% of those aged 60 and over (p = 0.006). Seventeen operated patients (13%) died before discharge. The hospital mortality increased from 3% among those < 60 years of age to 18% among those > 69 years (p = 0.041). Surgical treatment of carcinoma of the gastric cardia carries substantial morbidity and mortality. No important progress seems to have taken place since the 1960s. PMID- 10706923 TI - Repeated clinical and laboratory examinations in patients with an equivocal diagnosis of appendicitis. AB - In-hospital observation with repeated clinical examinations is commonly used in patients with an equivocal diagnosis of appendicitis. It is not known if repeated measurements of temperature and laboratory examinations have any diagnostic importance in this situation. The importance of repeated measurements of the body temperature, white blood cell (WBC) and differential cell counts, C-reactive protein concentration (CRP) and of the surgeon's repeated assessments was prospectively analyzed in 420 patients with an equivocal diagnosis of appendicitis at admission who were reexamined after a median of 6 hours of observation. The final diagnosis was appendicitis in 137 patients. After observation the inflammatory response was increasing among patients with appendicitis and decreasing among patients without appendicitis. The variables discriminating power for appendicitis consequently increased, from an area under the receiver operating characteristic (ROC) curve of 0.56 to 0.77 at admission, to 0.75 to 0.85 after observation. The ROC area of the surgeons' clinical assessment increased from 0.69 to 0.89. The WBC and differential cell counts were the best discriminators at the repeat examination. The change in the variables between the observations had weak discriminating power and had no additional importance in addition to the actual level at the repeat examination. To conclude, the diagnostic information of the temperature and laboratory examinations increased after observation. Repeated controls of the body temperature and laboratory examinations are therefore useful in the management of patients with equivocal signs of appendicitis, but the result of the examinations must be integrated with the clinical assessment. PMID- 10706924 TI - Resection of local recurrence of rectal cancer: results. AB - Locally recurrent rectal cancer is a difficult clinical problem, and surgical resection can be done only in selected patients. The aims of this study were to evaluate the results of resecting the local recurrence of rectal cancer and to analyze factors that might predict curative re-resection and those that affect survival. Forty-seven patients who underwent resection for locally recurrent rectal cancer formed the basis of the study. Twenty-four were curative in nature, and the others were palliative. There was no operative mortality, and the complication rate was 38%. The median survival of the whole group was 16.5 months. The ability to perform curative resection was found to be the only independent factor associated with improved survival. Female gender is a significant factor associated with curative resection of local recurrence. In patients with curative reresection, local control is up to 87%. It was concluded that resection of local recurrent rectal cancer can achieve good local control and can improve survival in selected patients. The ability to perform curative resection is associated with survival benefit, and female gender is associated with the increased possibility of carrying out curative resection. PMID- 10706925 TI - Standard and nonstandard applications of sentinel node-guided melanoma surgery. AB - Identification and histologic study of the sentinel node (SN) is an acceptable, yet not firmly established, guide for treating intermediate-thickness melanoma. This study widens the range of applications of this technique. We included 97 patients with intermediate-thickness melanoma lesions or lesions for which there is no standard treatment. Fifty-six underwent preoperative lymphoscintigraphy, and all underwent intraoperative lymphatic mapping (IOLM) using blue dye, followed by frozen section study and total node processing by serial sections. Elective lymph node dissection was performed in cases of metastasis to the sentinel node or technical failures with high risk. Four categories were defined: (A) intermediate-thickness lesions (mean 2.27 mm) (n = 45); (B) thin lesions (mean 1.14 mm) with risk factors of regional failure (n = 27); (C) lesion thickness close to but more than 4 mm (n = 10); and (D) lesions of undetermined thickness (n = 15). Median follow-up was 30 months (range 13-51 months). Intraoperative lymphatic mapping successfully identified the sentinel node (SN) in 93% of basins explored. Metastases were detected in 11 SNs. There were three lymph basin recurrences in patients with previously negative SNs, all salvaged by therapeutic lymph basin dissection and are NED (no evidence of disease). Two SN(+) patients had systemic recurrences; one died of his disease, and the other is alive with disease. One SN(-) patient died NED owing to other cause. This technique spared 83% of category A patients from lymph node dissection. It allowed better staging and better decision making for treatment in categories B and D; and it prevented early regional recurrences in category C patients. Intraoperative lymphatic mapping with SN guidance is a novel, lo PMID- 10706926 TI - The role of ultrasonography in acute appendicitis. PMID- 10706927 TI - Quality of life following radical surgical treatment of gastric carcinoma. PMID- 10706928 TI - Common C677T polymorphism in the methylenetetrahydrofolate reductase gene increases the risk for deep vein thrombosis in patients with predisposition of thrombophilia. AB - The alanine/valine (A/V) gene polymorphism of 5, 10-methylenetetrahydrofolate reductase (MTHFR), one of the key enzymes catalyzing remethylation of homocysteine, has been reported and the VV genotype is associated with increased plasma homocysteine levels as a result of the reduced activity and increased thermolability of this enzyme. Although previous studies have suggested that the VV genotype is a risk factor for arterial occlusive disease, whether the VV genotype is a risk factor for venous thrombosis is still controversial. Here we screened 72 Japanese patients with deep venous thrombosis (DVT) and 85 controls for this mutation, and we measured plasma levels of homocysteine to determine whether the thermolabile variant with the VV genotype is a risk factor for DVT in a Japanese population. Of the 72 patients with DVT, 10 (13.9%) were found to be homozygous for the VV genotype, and in 6 (7.0%) of 85, control individuals and the difference was not significant (odds ratio=2.12, 95% CI=0.73-6.16, p=0.19). When we divided the DVT patients into subgroups, with and without predisposition of thrombophilia, including deficiencies of proteins C and S, plasminogen, and lupus anticoagulant, the prevalence of the VV genotype in DVT patients with predisposition was significantly higher than that of the normal controls (odds ratio=5.99, 95% CI=1. 56-22.96, p=0.01). However, the prevalence of the VV genotype in DVT patients without predisposition was not significantly different from that of the normal controls (odds ratio=1.20, 95% CI=0.32-4.47, p=0. 75). The plasma homocysteine levels in patients with DVT (11.6+/-5.2 nmol/ml) was not significantly different from that of the control subjects (11.6+/-3.7 nmol/ml). Individuals with the VV genotype showed higher plasma homocysteine levels (15.4+/ 6.9 nmol/ml) than did individuals with the AV genotype (11.2+/-3.7 nmol/ml, p=0.009) or in individuals with the AA genotype (11.1+/-4.2 nmol/ml, p=0.004). Serum folate and vitamin B12 levels were not correlated with the plasma homocysteine levels. In conclusion, even though homozygosity for the VV genotype of the MTHFR gene was associated with higher plasma homocysteine levels, we found no association between plasma levels of homocysteine and DVT or between the genotype of the MTHFR gene and the DVT incidence. However, we found that the VV genotype of the MTHFR gene is a risk factor for DVT only when combined with the predisposition of thrombophilia. PMID- 10706929 TI - The decanucleotide insertion/deletion polymorphism in the promoter region of the coagulation factor VII gene and the risk of familial myocardial infarction. AB - Recently, an association has been found between factor VII polymorphisms and the risk of familial myocardial infarction. To obtain a thorough evaluation of the influence of factor VII gene on the risk of myocardial infarction, we extended our analysis to the role of a decanucleotide insertion/deletion functional polymorphism (-323 0/10-bp) in the promoter region of factor VII and to possible interactions with the HVR4 intron polymorphism. We performed a case-control study of 176 patients with myocardial infarction, over 45 years, who had a familial history of arterial thrombosis and 227 control subjects without a personal or family history of cardiovascular disease. The frequency of the rare allele of 10 bp was lower in cases (0.14 95% CI, 0.10-0.17) than in controls (0.19 95% CI, 0.16-0.23; chi(2)=4.7, p=0.03). Allowing for Hardy-Weinberg equilibrium in controls and testing for association under restricted maximisation, there was a significant difference in genotype frequency between cases and controls (p=0.02). Carriers of the 10-bp allele had an odds ratio for myocardial infarction of 0.65 (95% CI, 0.37-1.12), in multivariate logistic regression analysis. Combination analysis of -323 0/10-bp and HVR4 polymorphisms shows half reduction in the risk of myocardial infarction in comparison with the reference group for all the other groups, suggesting that there was no additivity between the effect of the 10-bp and the H7 alleles. Our findings suggest that the promoter polymorphism of factor VII gene may influence the risk of familial myocardial infarction. PMID- 10706930 TI - The effects of hormone replacement therapy on hemostatic variables in women with angiographically verified coronary artery disease: results from the estrogen in women with atherosclerosis study. AB - Data on the effect of hormone replacement therapy on hemostasis are inconsistent, and there are few data in women with coronary artery disease. In a single-center, open, randomized study, 118 postmenopausal women with angiographically verified coronary artery disease were randomized to hormone replacement therapy, given as long-cycle transdermal 17-beta-estradiol (50 microg/24 hour) for 3 months with sequential medroxy-progesterone acetate for 14 days, or to a control group receiving no therapy. Hemostatic parameters were measured at baseline and after 3 and 12 months of therapy. The coagulation inhibitors antithrombin, protein C, and protein S, but not tissue factor pathway inhibitor, decreased significantly from baseline in the hormone replacement therapy group at both 3 and 12 months as compared with the control group. The absolute decreases within the hormone replacement therapy group were 3 to 10%. No significant differences between the two treatment groups were observed for the coagulation products prothrombin fragment 1+2 or thrombin-antithrombin complex or for D-dimer, although there were significant decreases in the levels within the hormone replacement therapy group. Levels of fibrinogen, activated factor VII, and factor VII antigen were not significantly influenced by hormone replacement therapy treatment. Similarly, nonsignificant changes were detected for the fibrinolytic parameters tissue plasminogen activator activity, tissue plasminogen activator antigen, and global fibrinolytic activity, but plasminogen activator inhibitor type 1 was significantly lower in the hormone replacement therapy group due to a questionable increase in the levels in the control group. In conclusion, treatment with transdermal estradiol combined with long-cycle progestins was associated with no net activation of coagulation despite reduced levels of coagulation inhibitors. PMID- 10706931 TI - Thrombolytic properties of leukocytes from peripheral blood in healthy subjects and in patients with acute cerebral ischemia. AB - Polymorphonuclear leukocytes are activated in acute ischemic stroke. Activated polymorphonuclear leukocytes may contribute to thrombolysis by proteolytic degradation of fibrin and by modification of the plasminogen system. We used an in vitro thrombolysis model to investigate (1) thrombolytic properties of leukocytes in young and healthy subjects, (2) to test the hypothesis of increased polymorphonuclear leukocyte-associated thrombolysis in patients with acute cerebral ischemia, and (3) to assess plasminogen-dependent and -independent thrombolytic properties of polymorphonuclear leukocyte elastase. Coincubation of polymorphonuclear leukocytes with fibrin clots led to increased thrombolysis, a process reaching statistical significance after 8 hours [1x10(7) polymorphonuclear leukocytes/mL; 12.8+/-1.9% (mean+/-SEM), spontaneous clot lysis: 7.3+/-0.7%]. Polymorphonuclear leukocytes inside clots caused more efficient thrombolysis than polymorphonuclear leukocytes in the incubation medium. Spontaneous and polymorphonuclear leukocyte-associated lysis tended to be lower in patients with acute cerebral ischemia (n=9, 24 hours, 9.5+/-1.8% and 12.9+/-2.2%) than in age- and sex-matched control subjects (n=8; 12.2+/-2.0% and 17.4+/-1.9%). In the presence of alpha(2)-antiplasmin, thrombolysis tended to be faster with elastase-digested plasminogen (miniplasminogen) than with native plasminogen. Purified polymorphonuclear leukocyte elastase itself had no thrombolytic effect. We conclude that the thrombolytic capacity of polymorphonuclear leukocytes from peripheral blood is small and slow and may have been overestimated in previous reports. Polymorphonuclear leukocyte thrombolytic activity may not be increased in acute cerebral ischemia. Miniplasminogen may be an interesting adjunct to plasminogen activators in acute stroke models. PMID- 10706932 TI - Haemostatic function in patients undergoing coronary artery bypass grafting: peroperative perturbations and relations to saphenous vein graft closure. AB - Vein graft failure remains a major problem after coronary artery bypass grafting. Occlusion in the first weeks usually is caused by thrombosis, whereas intimal hyperplasia and eventually atherosclerotic changes with superimposed thrombus formation underlie subsequent closure. The present investigation was conducted as a pilot study to examine whether perturbations of haemostatic function predispose to early saphenous vein graft occlusion after coronary artery bypass grafting. Pre- and postoperative determinations (performed on the first, third, and sixth postoperative days) of haemostatic factors and inhibitors were related to the presence of graft occlusion assessed by angiography at 3 months after surgery in 100 men undergoing elective coronary artery bypass grafting for stable angina pectoris. Occlusion of one or more vein grafts within three months of surgery occurred in 23 of the 100 patients examined. The percentage increase in plasma plasminogen activator inhibitor-1 activity on the first postoperative day was significantly higher in patients who subsequently were found to have vein graft occlusion (p<0.05). Otherwise no postoperative haemostatic measurements were found to predict early vein graft closure. A perturbed plasma plasminogen activator inhibitor-1 response to coronary artery bypass grafting tentatively could be added to the vessel-specific factors that remain the main determinants of early vein graft closure. PMID- 10706933 TI - Inhibition of human platelet aggregation by gangliosides. AB - The content and composition of gangliosides is modified upon platelet stimulation, suggesting that these lipids may play functional roles in platelet physiology. Therefore, the effect of exogenously added gangliosides on human platelet aggregation was evaluated. The pretreatment of platelets with a mixture of total gangliosides from bovine brain and a series of purified mono-, di- and tri-sialogangliosides partially inhibit the collagen-induced aggregation process and ATP release and completely block the generation of the second aggregation wave when ADP is used as agonist. The inhibition was exerted at around 100 microM by G(TOT) as well as purified G(M1), G(M3), G(D1a), and G(T1b) gangliosides, whereas asialoG(M1) and sulphatide did not show a significant influence on platelet aggregation. Thrombin, Ca(2+) ionophores (A23187 and Ionomycin), arachidonic acid, and U46619 were unable to bypass the inhibitory effect exerted by gangliosides, suggesting that gangliosides inhibit platelet aggregation by inhibiting the synthesis or action of prostaglandins. Gangliosides inhibited U46619-induced aggregation, thus suggesting that they block the action of thromboxane A(2). Epinephrine induces a partial aggregation on gangliosides treated platelets, similar to fluoroaluminate and phorbol myristate acetate, indicating that these platelets are still functional. To summarize, these results indicate that the major pathway(s), but not all, driving to the aggregation process following the interaction of ligand-receptor may be blocked by pretreatment of human platelets with gangliosides. PMID- 10706934 TI - On the mechanism of the spermine-exerted inhibition on alpha-thrombin-induced platelet activation. AB - Previous reports have shown that various amines inhibited platelet activation, but no definitive conclusions on their action mechanism were drawn. We have further investigated the action of spermine on platelet responses evoked by alpha thrombin and other agonists. Spermine inhibited in a concentration-dependent manner (1-10 mM), and more efficiently than spermidine and putrescine, the alpha thrombin-induced (1.5 nM) platelet activation. Spermine added at a concentration that inhibited completely aggregation only partially affected the thrombin induced increase in cytosolic Ca(2+) concentration, protein phosphorylation, and ATP secretion. The polyamine had little effect on the morphology of resting platelets, as measured by electron microscopy, thrombin hydrolytic activity, and fibrinogen clotting capacity but decreased the thrombin binding to platelets and isolated glycocalicin. Spermine partially inhibited the aggregation elicited by ADP, vasopressin, platelet-activating factor, thrombin receptor-activating peptide, fluoroaluminate, ionomycin, and dioctanoylglycerol but did not affect the cytosolic Ca(2+) increase induced by these agonists. The polyamine bound to both glycocalicin and platelets, and it inhibited the fibrinogen binding to stimulated platelets. The amount of 14C-spermine bound to resting cells decreased in the presence of the glycoprotein GPIb-antibody LJIB1, whereas the polyamine bound to activated platelets, which was higher than that tied to resting cells, was markedly reduced by LJCP8 or decorsin, a GPIIb/IIIa antibody and antagonist peptide, respectively. These results indicate that spermine specifically inhibits the thrombin binding to GPIb of resting platelets and the fibrinogen binding to GPIIb/IIIa (integrin alpha(IIb)beta(3)) of activated platelets. PMID- 10706935 TI - Epitope mapping of monoclonal antibodies directed to PAI-1 using PAI-1/PAI-2 chimera and PAI-1-derived synthetic peptides. AB - Plasminogen activator inhibitor type-1 is a key regulatory protein of the fibrinolytic system that is involved in a variety of physiological and pathophysiological processes. A panel of 14 monoclonal antibodies directed against plasminogen activator inhibitor type-1 was analyzed regarding epitope specificity on plasminogen activator inhibitor type-1. For this purpose, chimera consisting of plasminogen activator inhibitor type-1 and another plasminogen activator inhibitor, plasminogen activator inhibitor type-2, with different portions of the respective wild-type proteins, were generated and plasminogen activator inhibitor type-1-derived 20-mer and 10-mer linear peptides were synthesized. Nine of the 14 monoclonal antibodies recognized an epitope located in the region between amino acid 76-188 of plasminogen activator inhibitor type 1, which encompasses the binding sites for vitronectin, heparin, and part of the fibrin binding region. Of these nine monoclonal antibodies, six reacted with a quadruple plasminogen activator inhibitor type-1 mutant (N152H, K156T, Q321L, M356I), and seven detected a plasminogen activator inhibitor type-1 deletion mutant (DeltaF111-H114). Two of the remaining five monoclonal antibodies recognized epitopes located between amino acid 209-227 and amino acid 352-371, respectively, while the other three antibodies reacted with wild-type plasminogen activator inhibitor type-1, only. Additional experiments revealed that two of the 14 mAbs neutralized and one monoclonal antibodies increased plasminogen activator inhibitor type-1 activity toward urokinase-type plasminogen activator, one of its target proteases. PMID- 10706936 TI - Antithrombotic properties of RWJ-50353, a potent and novel thrombin inhibitor. AB - The antithrombotic, anticoagulant, and kinetic properties of RWJ-50353, a novel, reversible, active-site-directed thrombin inhibitor, were evaluated. RWJ-50353 inhibited the catalytic activity of human alpha-thrombin with a K(i) of 0.19+/ 0.02 nM. It showed a 16-fold selectivity relative to inhibition of trypsin and at least 330-fold selectivity relative to inhibition of other biologically important serine proteases. In a gel-filtered platelet preparation, RWJ-50353 inhibited alpha-thrombin-induced aggregation with an IC(50) of 32+/-6 nM. In a canine arteriovenous shunt antithrombotic model, RWJ-50353 demonstrated a significant dose-related (0.1-1.0 mg/kg, i.v.) reduction in thrombus formation with 50% inhibition (ID(50)) obtained at 0.46+/-0.1 mg/kg. In a rabbit deep vein thrombosis model, RWJ-50353 dose-dependently (0.1-1. 0 mg/kg, i.v.) reduced thrombus formation with an ID(50) of 0.25+/-0. 03 mg/kg. The antithrombotic activity in both of these models was associated with only mild prolongations in bleeding time and coagulation parameters. These results demonstrate that RWJ 50353 is a potent, selective thrombin inhibitor that is an effective antithrombotic agent after intravenous administration in models of arterial and venous thrombosis and may be useful in the management of various thrombotic disorders. PMID- 10706938 TI - The action of Lonomia achelous caterpillars venom on human factor V. AB - The bleeding syndrome produced by contact with the Lonomia achelous caterpillars is characterized by a decrease of fibrinogen, factor XIII, plasminogen, and factor V with normal platelets. In this study, we report the effect of crude hemolymph and some semipurified chromatographic fractions on human factor V. Incubation of factor V with crude hemolymph resulted in an increase in procoagulant activity, followed by a subsequent decline in factor V activity. Identical results were obtained with fraction I, whereas with fraction II there was only a decrease in activity reaching its minimum at 30 minutes. fraction III did not modify the activity of factor V. All concentrations of fraction I tested produced an initial rise and subsequent fall in activity. However, at lower relative concentrations of fraction I, more sustained increases in activity were observed. The activator and inactivator activities present in fraction I show differences in temperature and pH stability, susceptibility to different inhibitors, and in SDS/PAGE pattern. The factor V activator is a thermostable protein, with maximum activity at acid pH and is inhibited by o-phenantroline, EDTA, and EGTA, while the factor V inactivator is thermolabile, presents maximum activity at basic pH, precipitates at pH 5.0, and is completely inhibited by iodoacetic acid and TLCK. It is partially blocked by diisopropyl fluorophosphate, phenylmethylsulfonyl fluoride, and p-chloromercuribenzoic acid. These results suggest that the activator should be a metallo-proteinase, while the inactivator is a serine or cysteine proteinase with a serine, histidine, or cysteine residue in the active site. PMID- 10706937 TI - Heparin cofactor II inhibits thrombus formation in a rat thrombosis model. AB - Heparin cofactor II is postulated to be an extravascular thrombin inhibitor that is physiologically stimulated by dermatan sulfate. However, the role of heparin cofactor II has not yet been clearly demonstrated in vivo. In this study, we estimated the antithrombotic effect of heparin cofactor II administered exogenously in a rat model of thrombosis. Thrombus was induced in the rat femoral artery by endothelial damage due to the photochemical reaction between systemically injected rose bengal and transillumination with green light. Pretreatment with heparin cofactor II significantly prolonged the time required to occlude the femoral artery (occlusion time) in a dose-dependent manner. At an effective dose in this thrombosis model, heparin cofactor II did not prolong the activated partial thromboplastin time and the prothrombin time in normal rats. Argatroban, a selective synthetic thrombin inhibitor, significantly prolonged the occlusion time. However, argatroban also prolonged the activated partial thromboplastin time and prothrombin time at an effective dose. These results suggest that the administration of heparin cofactor II in vivo effectively inhibited thrombus formation on the vessel walls whose endothelium is damaged without a prolongation of the coagulation time while heparin cofactor II may also inhibit the thrombin activity in the subendothelial tissue in vivo. PMID- 10706939 TI - High lactoferrin levels in disseminated intravascular coagulation and its possible negative role in coagulation. PMID- 10706940 TI - Expression of Fas-ligand in first trimester and term human placental villi. AB - The expression of Fas-ligand (FasL) on trophoblast cells is thought to play a role in immune regulation during human pregnancy. However, there are some discrepancies in the published data concerning the cell types expressing FasL in the placental villi. Therefore, we examined the expression of FasL on cryosections of first trimester and term placental tissue with three different anti-sera against FasL, which are in common use. By immunohistochemistry, all three anti-sera principally gave the same staining result. In the first trimester of pregnancy, villous cytotrophoblast cells underlying the syncytium, as well as all extravillous trophoblast cells of cell columns and cell islands, gave a clear, mainly membrane-located staining, whereas the syncytiotrophoblast, which forms the borderline to the maternal blood flow, only gave a spot-like reaction in distinct areas. The same result was obtained with term placental villi; however, in this tissue, the staining of the villous cytotrophoblast cells was less pronounced. From our results, we suggest that in placental villi, an important role of FasL in immune regulation is not very conclusive because this molecule is mainly expressed on trophoblast with no access to maternal blood or tissue. This is in contrast to the uterine part of the placenta, where FasL expressing trophoblast cells are in close contact with apoptotic maternal leukocytes. PMID- 10706941 TI - Expression of antigens involved in the presentation of lipid antigens and induction of clonal anergy in the female reproductive tract. AB - The molecular backgrounds of the anti-phospholipid syndrome and immunisation against the Rhesus proteolipid antigens are still largely unknown. In the present study, expression of (1) CD1, a major histocompatibility complex class l-like lipid antigen presenting molecule, (2) IL-10, a cytokine promoting induction of clonal anergy, and (3) CD80 and CD86, two T-cell costimulators preventing induction of clonal anergy when present, was investigated in frozen sections of cervix, corpus and the fallopian tube (FT) of 25-day-old BALB/c mice injected with FSH, progesterone or medium and of pregnant mice from each trimester (days 7, 14 and 19). CD1 was expressed by all endometrial epithelial cells. Enhanced immunostaining of the endometrial epithelial cells was observed after FSH treatment, and cervix and FT expressed generally more than the corpus of the uterus. After treatment with medroxy progesterone acetate (MPA), expression of CD1 by the endometrial epithelia was weak. During pregnancy CD1 was absent from the endometrium adjacent to the foetus, but was unaltered in the cervix and FT. IL-10 was expressed by the endometrial glands and also by the endometrial surface epithelium. MPA treatment increased the intensity of the IL-10 immunofluorescence. There were also chains of positive cells between the muscle bundles within the pregnant myometrium. CD80 and CD86 were usually absent from the female reproductive tract, but were occasionally found in the cervix during pregnancy. The present study demonstrates definite differences in the expression of both CD1 and IL-10 between the FSH and MPA treated mice, suggesting differences during the oestrous cycle. As IL-10 is expressed more in the secretory phase, it is probably involved in making the endometrium more acceptable for implantation by inducing clonal anergy. This is supported by the absence of CD80 and CD86. These results also suggest that abnormal expression of CD1d during pregnancy may predispose the mother for immunisation against lipid antigens such as membrane phospholipids and Rhesus-antigens. PMID- 10706942 TI - Possible mechanisms of mammalian immunocontraception. AB - Ecological and conservation programs in ecosystems around the world have experienced varied success in population management. One of the greatest problems is that human expansion has led to the shrinking of wildlife habitat and, as a result, the overpopulation of many different species has occurred. The pressures exerted by the increased number of animals has caused environmental damage. The humane and practical control of these populations has solicited the scientific community to arrive at a safe, effective, and cost-efficient means of population control. Immunocontraception using zona pellucida antigens, specifically porcine zona pellucida (pZP), has become one of the most promising population control tools in the world today, with notable successes in horses and elephants. A conundrum has risen where pZP, a single vaccine, successfully induces an immunocontraceptive effect in multiple species of mammals. This review describes the most current data pertaining to the mammalian zona pellucida and immunocontraception, and from these studies, we suggest several potential mechanisms of immunocontraception. PMID- 10706943 TI - Prolonged gestation does not extend survival of uterine natural killer lymphocytes in mice deleted in the receptor for prostaglandin F2alpha. AB - During decidualization in mice and women, expansion of the Natural Killer (NK) cell lineage occurs within the uterus. In rodents, peak numbers of uterine (u)NK cells are reached at mid-gestation. The population then declines and residual cells are shed with the placenta. Decidualization, but not a fetus, is required to induce division and maturation of uNK cells. Mechanisms regulating the decline in uNK cells are unknown. To determine if the conceptus or its products have regulatory roles on uNK cell survival during normal gestation, a histological time course study was undertaken of implantation sites in mice ablated in the gene for the Prostaglandin F2alpha receptor (PGF2alphaR). These females experience normal gestation but fail to initiate labour and delivery. Their pregnancies extend a further 4-7 days before onset of maternal compromise. Large numbers of uNK cells were present in PGF2alphaR null mice by gestational day (gd) 10 and numbers had begun to decline at gd 14. By gd 18, very few uNK cells remained and no uNK cells were found at day 22 of extended gestation. Thus, the population history of uNK cells in PGF2alphaR null mice resembles that of uNK cells in normal mice, suggesting that the placenta, its products, the fetus and PGF2alpha are not factors that influence the rate of uNK cell decline in late gestation. PMID- 10706944 TI - Granulocyte-macrophage colony-stimulating factor (GM-CSF) targets myeloid leukocytes in the uterus during the post-mating inflammatory response in mice. AB - Factors in seminal plasma elicit a surge of GM-CSF expression in uterine epithelial cells after mating in mice. This study investigates the nature of the endometrial cell populations targeted by epithelial GM-CSF. In quantitative RT PCR studies, expression of the alpha-subunit of the GM-CSF receptor (GM-CSF-R) parallelled GM-CSF expression, being maximal during the 48 h period after mating and declining thereafter. Expression of mRNA encoding beta-common chain (AIC2B) also increased after mating and remained high until the time of embryo implantation on day 4 of pregnancy. Cells expressing GM-CSF receptors were identified in sections of uterus on the day after mating using 125I-GM-CSF, and were located predominantly in the endometrial stroma subjacent to the luminal epithelium, co-localising with abundant populations of myeloid leukocytes. Cells expressing GM-CSF receptor were identified as macrophages, granulocytes and putative dendritic cells by flow cytometric analysis using lineage and receptor subunit specific antibodies. Recombinant GM-CSF injected into the uterine lumen of ovariectomised mice was found to elicit a dose-dependant accumulation of macrophages and granulocytes in the endometrium, in a pattern of distribution comparable to that seen in uteri after natural mating. Together, these data indicate a role for epithelial cell-derived GM-CSF in mediating the recruitment and potentially in modifying the behaviour of uterine leukocytes during the post mating inflammatory response in mice. PMID- 10706945 TI - Correlation between oral sex and a low incidence of preeclampsia: a role for soluble HLA in seminal fluid? AB - The involvement of immune mechanisms in the aetiology of preeclampsia is often suggested. Normal pregnancy is thought to be associated with a state of tolerance to the foreign antigens of the fetus, whereas in preeclamptic women this immunological tolerance might be hampered. The present study shows that oral sex and swallowing sperm is correlated with a diminished occurrence of preeclampsia which fits in the existing idea that a paternal factor is involved in the occurrence of preeclampsia. Because pregnancy has many similarities with transplantation, we hypothesize that induction of allogeneic tolerance to the paternal HLA molecules of the fetus may be crucial. Recent data suggest that exposure, and especially oral exposure to soluble HLA (sHLA) or HLA derived peptides can lead to transplantation tolerance. Similarly, sHLA antigens, that are present in the seminal plasma, might cause tolerance in the mother to paternal antigens. In order to test whether this indeed may be the case, we investigated whether sHLA antigens are present in seminal plasma. Using a specific ELISA we detected sHLA class I molecules in seminal plasma. The level varied between individuals and was related to the level in plasma. Further studies showed that these sHLA class I molecules included classical HLA class I alleles, such as sHLA-A2, -B7, -B51, -B35 and sHLA-A9. Preliminary data show lower levels of sHLA in seminal plasma in the preeclampsia group, although not significantly different from the control group. An extension of the present study is necessary to verify this hypothesis. PMID- 10706946 TI - Pronounced substance P innervation in irradiation-induced enteropathy--a study on human colon. AB - The immunohistochemical expression of various neuropeptides, including substance P (SP), and the substance P receptor (SPR), was examined in irradiation-induced enteropathy in man. Samples from irradiated and non-irradiated patients operated on for rectal carcinoma were examined. The samples were from the sigmoid and corresponded macroscopically to non-cancerous sigmoid colon. There was a marked atrophy, ulcerations and inflammatory reactions in the irradiation-influenced mucosa. In this mucosa, there was a very pronounced innervation of varicose nerve fibers showing SP-like immunoreactivity (LI). The degree of SP-LI in the ganglionic cells of the submucous plexus was increased as compared to non irradiated patients. There were only few or no nerve fibers showing immunoreaction for other neuropeptides examined (CGRP, enkephalin, NPY) in the irradiation-influenced mucosa. A marked SPR immunoreaction was detected in cells in the lamina propria which were interpreted as representing polymorphonuclear leukocytes. The marked expression of SP in the irradiation-damaged mucosa and the presence of SPR immunoreactive leukocytes suggest that SP is highly involved in the inflammatory reactions that occur in response to radiotherapy. The observations also suggest that SP, but not NPY, CGRP and enkephalin, has an important role in the reorganisation processes that take place in the mucosa in irradiation-induced enteropathy. PMID- 10706947 TI - Neuroendocrine peptide levels in the gastrointestinal tract of mice after unilateral cervical vagotomy. AB - The effects of left and right unilateral cervical vagotomy on the content of several neuroendocrine peptides were studied in different parts of the murine gastrointestinal tract, known to receive vagal innervation. The neuroendocrine peptides investigated were secretin, gastric inhibitory peptide (GIP), gastrin, motilin, peptide YY (PYY), somatostatin, substance P, VIP, neurotensin, neuropeptide Y (NPY), and galanin. The neuroendocrine peptide concentration was affected after both left and right vagotomy, and that the changes in the concentrations of the neuroendocrine peptide levels occurred in all the gastrointestinal segments investigated, namely antrum, small and large intestine. However, these changes varied, depending on which side was vagotomized and the interval after vagotomy. It is concluded that the vagus nerve had an important impact on the neuroendocrine system in the murine gut. It is suggested, furthermore that the contradictory results obtained earlier on the effect of vagotomy on the gastrointestinal peptides may depend on differences in the vagotomy methods used and on differences in observation time after vagotomy. PMID- 10706948 TI - Calcitonin modifies ligand binding to muscarinic receptor in CNS membranes. AB - Calcitonin (CT) is a peptide produced by the thyroid gland, whose best described role is to prevent bone reabsorption, though it also participates in other biological functions through both central and peripheral mechanisms. CT is able to inhibit brain Na(+), K(+)-ATPase activity (Rodriguez de Lores Arnaiz, Lopez Ordieres, Peptides 1997;18:613-5) and a relationship between such enzyme activity and cholinergic function has been suggested. Accordingly, we tested CT effect on [(3)H]-quinuclidinyl benzilate ([(3)H]-QNB) binding to rat CNS membranes to determine whether the peptide is able to modify the cholinergic muscarinic receptor as well. It was found that 1x10(-7)-1x10(-5) M CT decreased 20-70% ligand binding to hippocampal, cerebellar, cortical and striatal membranes. Scatchard analysis of saturation curves showed that 5x10(-6) M CT significantly modified binding kinetic constants, thus it increased roughly 220% K(d) values and decreased 20-36% B(max) values in cerebral cortical and cerebellar membranes. Since the peptide decreases affinity ligand binding and reduces the number of binding sites, CT may well be acting as a cholinergic modulator through a decrease in muscarinic receptor functionality. PMID- 10706949 TI - Improved conditioned avoidance learning by oxytocin administration in high emotional male Sprague-Dawley rats. AB - OBJECTIVE: To examine anti-stress-like properties of oxytocin as a means to improve conditioned avoidance learning in a low-performing, high-emotional, stock of Sprague-Dawley male rats. METHODS: Adult male rats of two stocks of the Sprague-Dawley strain, designated Stock A and Stock B, were treated daily with oxytocin (1 mg kg(-1) s. c.) for 5 days preceding four daily conditioned avoidance acquisition sessions (approximately 20 trials per 15 min session). The Stock B animals were previously characterized as high-emotional based on [1] elevated plasma corticosterone, and lowered plasma oxytocin, levels and [2] decreased reaction time and an increased startle amplitude to an acoustic stimulation. Finally, [3] these animals were unable to acquire a conditioned avoidance response within 5 days of training. RESULTS: The Stock A animals rapidly and statistically significantly acquired the avoidance behaviour within 4 days of daily training, whereas Stock B animals did not improve over this time period. The avoidance performance of Stock B animals was markedly and statistically significantly improved by the oxytocin pre-treatment, whereas the performance of Stock A animals was not affected by the same oxytocin treatment. CONCLUSIONS: Pre-treatment with oxytocin markedly improved avoidance learning in the Stock B high-emotional animals. It is suggested that the improvement is due to previously demonstrated anti-stress-like properties of oxytocin, rendering the animals able to successfully cope with the demands of the conditioned avoidance situation. PMID- 10706950 TI - Influence of bombesin on neuronal hypothalamic thermosensitivity during the early postnatal period in the Muscovy duck (Cairina moschata). AB - The influence of bombesin (1 microg/0.1 ml artificial cerebrospinal fluid) on neuronal thermosensitivity of the preoptic area of the anterior hypothalamus in brain slices of 5- (n = 7 neurons) and 10-day-old (n = 36 neurons) Muscovy ducks (Cairina moschata) was investigated. Similar to adult mammals, most of the neurons investigated increased the firing rate (FR) after bombesin application. Changes in FR were not related to changes in thermal coefficient (TC). The neurons react to bombesin also under synaptic blockade. The bombesin-induced effect on TC (increase or decrease in nearly the same number of neurons, e.g. nine neurons increased and ten decreased TC in 10-day-old ducklings) in the postnatal bird neurons investigated was different from the results described in adult mammals, where the main reaction to bombesin was an increase of TC in warm sensitive and temperature-insensitive-neurons and a transformation of temperature insensitive-neurons into warm-sensitive ones. This may be related to the assumption that during early ontogeny, body functions react to exogenous and endogenous factors nonspecifically. It is to speculate, that later, probably at the end of embryonic development or during the early postnatal period, the reactivity of these functions changes qualitatively, so that the reaction of an individual function to different factors becomes specific (ultimately adaptive). PMID- 10706951 TI - Endothelin-1 increases isoprenaline-enhanced cyclic AMP levels in cerebral cortex. AB - We examined the effect of ET-1 on cyclic AMP levels in rat cerebral cortex. The peptide caused a concentration-dependent increase of [(3)H]cyclic AMP accumulation after 10 min of treatment. This effect was due to adenosine accumulation since it was inhibited by the treatment with adenosine deaminase. ET 1, apart from being able to increase cyclic AMP, also potentiated the cyclic AMP generated by isoprenaline in the presence of adenosine deaminase. Experiments performed in the presence of BQ-123 or BQ-788, specific ET(A) or ET(B) receptor antagonists respectively indicated that ET(B) was the receptor involved. This effect was dependent on extracellular and intracellular calcium concentration. These findings suggest that ET-1 plays a modulatory role in cyclic AMP generation systems in cerebral cortex. PMID- 10706952 TI - Role of endogenous CCK in the inhibition of gastric emptying by peptone and Intralipid in rats. AB - To assess the role of endogenous cholecystokinin in the control of gastric emptying of peptone solutions and Intralipid suspensions, we examined the ability of a dose range of the CCK-A antagonist, devazepide to accelerate the gastric emptying of various caloric concentrations of peptone and Intralipid in rats. In the absence of devazepide, both peptone and Intralipid emptying slowed with increasing concentration. Devazepide's effect on peptone gastric emptying diminished with increasing peptone concentration. The threshold dose for accelerating the emptying of 0.2 kcal/ml peptone was lower than the threshold dose for affecting 0.5 kcal/ml peptone and devazepide had no effect on the gastric emptying of 1.0 kcal/ml peptone. In contrast, devazepide affected Intralipid gastric emptying at all three Intralipid concentrations and the threshold dose decreased with increasing Intralipid concentration. However, the magnitude of the effect of devazepide on peptone or Intralipid gastric emptying was partial and did not increase as a function of concentration. These data demonstrate a role for endogenous CCK in the emptying of peptone and Intralipid but suggest that endogenous CCK does not account for the increased slowing of gastric emptying evident with increased caloric concentration. PMID- 10706953 TI - Functional CCK-A and Y2 receptors in guinea pig esophagus. AB - Effects of cholecystokinin octapeptide (CCK-8), peptide YY (PPY), neuropeptide Y (NPY) and their analogs on muscle contractions of esophageal strips were investigated. CCK-8 induced a tetrodotoxin and atropine-sensitive contraction. The relative potencies for CCK related peptides to induce contractions were CCK-8 > desulfated CCK-8 > gastrin-17-I. The CCK-A receptor antagonist L-364,718 was 300-fold more potent than the CCK-B receptor antagonist L-365,260 at inhibiting CCK-8-induced contraction. These indicate that neural CCK-A receptors mediate this contraction. PYY or NPY did not cause muscle contraction or inhibit muscle contraction induced by carbachol, endothelin-1 or KCl. However, both PYY and NPY concentration-dependently inhibited contraction induced by CCK-8. This inhibition was not affected by nitric oxide (NO) synthase inhibitors L-NMMA or L-NAME. The relative potencies of PYY related peptides to inhibit CCK-8 induced contraction were PYY > NPY > NPY13-36 > [Leu(31), Pro(34)]NPY > pancreatic polypeptide (PP). We conclude that CCK interacts with neural CCK-A receptors to cause esophageal muscle contraction. PYY and NPY interact with Y2 receptors to inhibit this CCK induced muscle contraction by an effect not related to NO. PMID- 10706954 TI - Somatostatin receptor gene expression in neuroblastoma. AB - Somatostatin receptor expression is a favorable prognostic factor in human neuroblastoma. Somatostatin receptors have been demonstrated in vitro by pharmacologic analysis of tumor tissue and in vivo by diagnostic radioreceptor scintigraphy. However, which receptor subtypes (sst(1), sst(2), sst(3), sst(4), and sst(5)) are expressed in these tumors has not yet been delineated. We used RT PCR to analyze expression of the five somatostatin receptor genes in 32 neuroblastoma tumor specimens. All 32 tumor specimens expressed mRNA for c-abl and sst(1); sst(2) mRNA was detected in 27/32 samples and somatostatin mRNA was detected in 30/32 tumor specimens. The remaining receptor subtypes, sst(3), sst(4), and sst(5) were variably expressed. Receptor protein for sst(1) and sst(2) was visualized in tumor neuroblasts as well as in endothelial cells of tumor vessels using immunostaining with specific anti-receptor antibodies. The effect of high expression of somatostatin receptors on cell proliferation was examined in SKNSH neuroblastoma cells transfected with sst(1) and sst(2). SS(14) binding to wild-type SKNSH cells was undetectable; but the native peptide bound with high affinity to the SKNSH/sst(1) and SKNSH/sst(2) neuroblastoma cell lines. Pharmacologic analysis of binding with two long-acting analogues, CH275 and octreotide, confirmed selective expression of sst(1) and sst(2) in stably transfected SKNSH cells. Formation of neuroblastoma xenograft tumors in nude mice was significantly delayed for both SKNSH/sst(1) (P<0.001) and SKNSH/sst(2) (P<0.05) cells compared to wild-type SKNSH. We conclude that: (1) Somatostatin receptors, sst(1) and sst(2), are expressed in the majority of neuroblastomas at diagnosis; and (2) upregulation of functional sst(1) or sst(2) in neuroblastoma cell lines suppresses tumorigenicity in a xenograft model. These observations suggest that somatostatin receptors may be a useful therapeutic target in neuroblastoma. PMID- 10706955 TI - CCK-A receptor activation induces fos expression in myenteric neurons of rat small intestine. AB - Cholecystokinin (CCK), a hormone secreted from endocrine cells of the small intestine, participates in the control of motility and secretion in the gastrointestinal tract, and in the control of food intake. At least some of the effects of CCK on intestinal function appear to be mediated via activation of intrinsic neurons in the myenteric plexus. However, the distribution of CCK responsive enteric neurons within the rat small intestine is not known. Neither has the role of CCK-A receptors in the activation of rat myenteric neurons been investigated. Therefore, to determine the distribution of CCK-responsive neurons in the small intestinal myenteric plexus we utilized immunohistochemical detection of Fos, the protein product of the immediate early gene c-fos, to identify neurons that were activated by exogenous CCK. We also monitored Fos expression in the dorsal hindbrain, and examined CCK-induced Fos expression in the presence or absence of a receptor antagonist for the type-A CCK receptor. We found that CCK significantly increased Fos expression in the hindbrain and in myenteric neurons of the duodenum and jejunum, but not the ileum. Neuronal Fos responsiveness in both brain and myenteric neurons was mediated by CCK-A receptors, as CCK-induced Fos expression was eliminated in rats pretreated with a CCK-A receptor antagonist. We conclude that CCK activates small intestinal myenteric neurons, via CCK-A receptors. Activation of these intrinsic intestinal neurons may participate in reflexes and behaviors that are mediated by CCK. PMID- 10706956 TI - Involvement of ryanodine receptors in EPYLRFamide-mediated reduction of acetylcholine-induced inward currents in helix lucorum identified neurones. AB - The effects of several modulators of ryanodine receptors (RYRs) on the reduction of acetylcholine induced inward current (ACh-current) evoked by EPYLRFamide (5 microM, bath application), the potent N-terminally modified analogue of the endogenous Helix heptapeptide SEPYLRFamide, were investigated. These modulators were applied intracellularly. Inward currents were recorded from identified Helix lucorum LPa2, LPa3, RPa3, RPa2 neurones in ganglia preparations using the two electrode voltage clamp technique. ACh was applied ionophoretically. BAPTA (0.1 mM), chelator of intracellular Ca(2+), ryanodine (0.1 mM), agonist/antagonist of RYRs and dantrolene (0.1 mM), antagonist of RYRs decrease the effect of EPYLRFamide. Adenosine (1 mM), alpha,beta-methylene ATP (0.1 mM), the nonhydrolisable ATP analogue and cyclic adenosine diphosphate ribose (0.1 mM) (agonists of RYRs) potentiate the modulatory effect of EPYLRFamide. Ruthenium red (1 mM), antagonist of RYRs and caffeine (1 mM), agonist of RYRs do not change the modulatory effect of EPYLRFamide. These data suggest that intracellular Ca(2+) and RYRs are involved in the modulatory effect of EPYLRFamide on ACh-currents. It was concluded that EPYLRFamide decreases ACh-current through elevation of basal intracellular level of a putative endogenous agonist of RYRs which activates RYR dependent mobilization of Ca(2+) by binding to the adenine nucleotide site of the ryanodine receptor-channel complex and does not bind the site activated by caffeine. PMID- 10706957 TI - Immune responses to pertussis antigens eight years after booster immunization with acellular vaccines in adults. AB - Pertussis-specific antibody and cell-mediated immune (CMI) responses were studied in adults 8 years after booster immunization with either a bicomponent (pertussis toxin and filamentous hemagglutinin) or a monocomponent (pertactin) acellular vaccine and in age-matched healthy controls. The levels of vaccine-induced antibodies were also compared between the serum samples collected before, 1 month, 4 years, and 8 years after immunization. Over the follow-up period, geometric mean values (GMV) of antibodies to the vaccine antigens decreased in both groups of vaccinees. However, the 8-year postimmunization GMV were 3-20 times higher than preimmunization GMV (all P values <0.01). Moreover, both antibody and CMI responses to the vaccine antigens were significantly higher in the vaccinees than in the controls (all P<0.01 for antibody; all P<0.001 for CMI responses). The results show that antibody and CMI responses induced by acellular pertussis vaccines can persist for up to 8 years after booster immunization of adults primed with whole-cell vaccine. PMID- 10706958 TI - A gE-negative bovine herpesvirus 1 vaccine strain is not re-excreted nor transmitted in an experimental cattle population after corticosteroid treatments. AB - To study possible reactivation and to quantify subsequent transmission of a live gE-negative bovine herpesvirus 1 (BHV1) vaccine strain in cattle populations, four experiments were performed. Two groups of cattle were each tested twice for the possibility of reactivation. Inoculation with a gE-negative BHV1 vaccine was done either intramuscularly or intranasally and treatment with corticosteroids in an attempt to reactivate vaccine virus, was done after 6 or 11 weeks, and again after 6 months. To quantify transmission of vaccine virus following possible reactivation, each cattle was housed together with one susceptible contact cattle. Contact-infections were monitored using virus shedding and antibody responses. After corticosteroid treatments, re-excretion of virus was never detected in cattle that had been inoculated with the gE-negative BHV1 vaccine strain. Contact cattle did not shed gE-negative BHV1, nor mounted any antibody response against BHV1. In contrast, positive control cattle, inoculated intranasally with wild-type BHV1, re-excreted virus in high titers in nasal fluids and transmitted the virus to contact cattle. Based on these results, the transmission ratio R(0) of the vaccine strain was zero. We concluded that it is highly unlikely that the live gE-negative BHV1 vaccine strain will be re-excreted after possible reactivation, and consequently, it is even less likely that reactivated vaccine virus will spread in the cattle population. PMID- 10706959 TI - Quantitative determination of saccharide in Haemophilus influenzae type b glycoconjugate vaccines, alone and in combination with DPT, by use of high performance anion-exchange chromatography with pulsed amperometric detection. AB - The stability and integrity of glycoconjugate vaccines requires determination of the total saccharide and quantification of the unbound or free saccharide present. The traditional assay for Hib conjugates, based on colorimetric determination of ribose, has been much improved by the use of base hydrolysis and analysis of the Hib subunit generated using high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD). The production of this subunit was confirmed by NMR analysis. However, quantification of free Hib saccharide using this method was not possible in the combination vaccines evaluated due to interferences emanating from DPT. Thus a method based on TFA hydrolysis followed by the chromatographic separation and quantification of ribitol on a CarboPac MA1 column was developed. The method is selective, and with the use of ED40 electrode, requires only nanomole amounts for the chromatographic step, thereby ensuring that free saccharide can be monitored accurately in the formulated Hib-CRM vaccine alone and when in combination with other vaccines. PMID- 10706960 TI - Is there a causal link between hepatitis B vaccination and multiple sclerosis? AB - After the publication of case reports of hepatitis B vaccinees with onset or relapse of multiple sclerosis (MS), followed by a media-driven scare campaign in France, the perception that hepatitis B vaccine causes MS has developed. This has led to a fall in the acceptance of hepatitis B vaccination particularly in French speaking communities which was accelerated by court decisions in favour of vaccination "victims" and the suspension of routine vaccination of pre adolescents in French schools as a "precautionary measure". This situation has arisen in spite of the absence of scientific data to support a causal link between vaccination and multiple sclerosis. In this article, initially written to inform and reassure employees of one of the vaccine manufacturers, the epidemiological importance of hepatitis B and current knowledge on the aetiology of MS are described. All available data that may throw light on the hypothesis that hepatitis B vaccination is causally linked to MS was reviewed. The conclusion reached on the basis of available data is that the most plausible explanation for the observed temporal association between vaccination and MS is that it is a coincidental association. It is now important to rebuild public confidence in hepatitis B vaccine as well as in vaccination in general. PMID- 10706961 TI - IL-18 potentiates the adjuvant properties of IL-12 in the induction of a strong Th1 type immune response against a recombinant antigen. AB - Due to the synergistic effects of IL-12 and IL-18, and to their importance in establishing a Th1 type immune response, we investigated the potential of a combined administration of both cytokines as an adjuvant for recombinant antigens. As a model system, we used a schistosome T cell antigen recently identified in our group. By co-adsorption of this antigen on alum in the presence of IL-12 and IL-18, we demonstrate that IL-18 enhances the effects of IL-12 in inducing an antigen-specific Th1 type CD4(+) T cell response as well as high titres of IgG1, IgG2a, and IgG2b antibodies. PMID- 10706962 TI - Study of the safety, immunogenicity and seroconversion of a hepatitis-B vaccine in malnourished children of India. AB - Sixty rural children who were seronegative for HBV markers received three doses of 10 microgram of a new Hepatitis-B vaccine, Revac-B (1 ml of vaccine contains 20 microgram recombinant surface antigen) that was formulated from hepatitis-B surface antigen expressed in a recombinant strain of Saccharomyces cerevisiae. Vaccines were administered on a 0, 30 and 60-day schedule. Levels of anti-HBs titres were determined on the 30th, 60th and 90th days following the initial injection. None of the participants in the trial had serious adverse reactions and the frequencies of minor side effects were minimal. No clinically important adverse effects which could be considered as directly related to the vaccination were recorded. The volunteers showed a very good immune response and were seroprotected on the 30th day after the first dose of vaccination. The present study revealed that the new vaccine, Revac-B is highly immunogenic and is well tolerated. PMID- 10706964 TI - Immunogenicity of porcine transmissible gastroenteritis virus spike protein expressed in plants. AB - Transgenic plants expressing recombinant proteins from pathogenic microorganisms provide an inexpensive edible vaccine for induction of local immunity. Three transgenic plant lines were generated expressing the spike (S) protein of transmissible gastroenteritis virus (TGEV), a protein crucial for establishing mucosal immunity. All three of them were driven by a strong plant promoter. One construct contained the 3.7 kb 5' end of the native S gene sequence. In the second construct part of the S gene, from nucleotide 49 to 1785, was modified for optimal plant recognition and was fused to a plant signal peptide coding sequence. The third construct contained the D epitope-coding region of the S gene, from nucleotide 1201 to 1591, which was fused to the alfalfa beta-amylase gene. The S gene products were detected by enzyme-linked immunosorbent assay (ELISA) and Western blotting. Antigens from all three transgenic plant lines induced TGEV-specific immune responses in pigs as determined by virus neutralization and ELISA, and the resultant antibody titers for all three constructs were similar. PMID- 10706963 TI - Boosting with recombinant vaccinia increases HPV-16 E7-specific T cell precursor frequencies of HPV-16 E7-expressing DNA vaccines. AB - We have previously linked the sorting signals of the lysosome-associated membrane protein-1 (LAMP-1) to HPV-16 E7 antigen, creating a chimera, Sig/E7/LAMP-1. We found that both Sig/E7/LAMP-1-containing recombinant vaccinia virus (Vac Sig/E7/LAMP-1) and Sig/E7/LAMP-1 DNA can generate strong antitumor immunity. To determine whether combination of Sig/E7/LAMP-1 DNA and Vac-Sig/E7/LAMP-1 can further enhance immune responses, sequential vaccination with Sig/E7/LAMP-1 DNA and Vac-Sig/E7/LAMP-1 was given. We found that priming with Sig/E7/LAMP-1 DNA and boosting with Vac-Sig/E7/LAMP-1 generated the strongest E7-specific CD8(+) T cell responses. Our results encourage the use of the DNA prime/vaccinia booster regimen in future clinical trials. PMID- 10706965 TI - Immunogenicity of a combined hepatitis A and B vaccine in healthy young adults. AB - Combining several vaccines in a single formulation can change the potency of the vaccine antigens. Previous studies suggested a higher immunogenicity of a new combined hepatitis A and B vaccine compared with the monovalent hepatitis B vaccine. We investigated the immune response to hepatitis B surface antigen 1 month after the third vaccine dose in 282 healthy adults who had received either a monovalent hepatitis B vaccine (n=148) or the combined hepatitis A/B vaccine (n=134). A slight trend towards higher geometric mean titres of anti HBs was found at this point in time in the group immunised with the combined vaccine, especially in the few vaccinees with preexisting antibodies against hepatitis A virus. However none of these differences was statistically significant, arguing against an advantage of the combined vaccine regarding hepatitis B immunisation. PMID- 10706966 TI - Induction of a protection against S. mansoni with a MAP containing epitopes of Sm37-GAPDH and Sm10-DLC. Effect of coadsorption with GM-CSF on alum. AB - Studies of anti-S. mansoni immunological responses in individuals living in endemic areas identified immunogens (Sm37-GAPDH and Sm10-DLC) with vaccine candidate properties. Analysis of the epitopes of these immunogens indicated that: (i) Sm37-5 is a major B-cell epitope of Sm37-GAPDH and the IgG antibody reactivity toward this determinant is associated with resistance to reinfection; (ii) Sm10-T is a T-cell epitope of the major T-cell immunogen Sm10-DLC. This led us to test a multiple antigen peptide (MAP) containing Sm37-5 and Sm10-T as an anti-schistosome vaccine. This MAP induced a significant protective immune response in mice when injected in Freund's adjuvant or coadsorbed with GM-CSF on aluminium hydroxide. In the latter case the physical link between the cytokine and the antigen via the coadsorption on alum was necessary to obtain a protective response. Results of the antibody response indicated that when the MAP and GM-CSF were coadsorbed on alum, the antibody response against the Sm10-T epitope located in the NH(2)-terminal position was significantly amplified up to 30% of the anti Sm37-5 response. PMID- 10706967 TI - Identification of a candidate vaccine peptide on the 37 kDa Schistosoma mansoni GAPDH. AB - A previous study performed in adolescents living in an area endemic for Schistosoma mansoni in Brazil has shown that a 37 kDa schistosome surface antigen is a selective target for antibodies in sera from those who were resistant to reinfection. This antigen was shown by molecular cloning to be the schistosome GAPDH. The aim of the present work was to assess whether peptides corresponding to GAPDH antigenic determinants could be used in a subunit vaccine. Five B cell and two T cell epitopic regions were identified on Sm37-GAPDH. One of the B cell determinants (Sm37-5, aa 268-289) is highly antigenic in human infections and antibody reactivity toward this determinant is associated with resistance to reinfection. Mice and rats immunized with Sm37-5 were partially protected against a challenge infection, indicating that this peptide can induce protective immunity. Analysis of Sm37-5 amino acid sequence indicated that this antigenic determinant is likely conserved among other pathogenic strains of schistosome (S. haematobium, S. intercalatum and S. japonicum), although it shows major amino acid differences with the corresponding human GAPDH sequence. All together these results indicate that Sm37-5 should be considered as a candidate component for an anti-schistosome subunit vaccine. PMID- 10706968 TI - Humoral immunoresponse to varicella-zoster virus pernasally coadministered with Escherichia coli enterotoxin in mice. AB - It is evaluated whether Escherichia coli enterotoxin is useful for induction of immunity to varicella-zoster virus (VZV) as a mucosal adjuvant in mice. When a commercially available live varicella vaccine (Oka strain) and toxin were administered simultaneously via a nasal route three times at 2 or 6 month intervals, an antibody neutralizing VZV was detected in half or all of the mice vaccinated, respectively. The antibody specific to the vaccine strain of VZV reacted to five proteins, molecular weights of which were 110 K, 100 K, 62 K, 54 K and 46 K. These proteins were composed of glycosylated products of all kinds of glycoproteins. These results suggest that although a nasal administration of the vaccine without the adjuvant has little immunogenicity in mice, the simultaneous administration of the live vaccine and the toxin over a long period induces a specific humoral immunity to VZV. PMID- 10706969 TI - IgE sensitization to gelatin: the probable role of gelatin-containing diphtheria tetanus-acellular pertussis (DTaP) vaccines. AB - We recently found that most events of anaphylaxis to live attenuated viral vaccines containing gelatin as a stabilizer might be caused by the gelatin. However, the mechanism that the children were sensitized to gelatin was unclear. In Japan, both diphtheria-tetanus-acellular pertussis (DTaP) vaccines with and without gelatin are available. We explored the possibility that gelatin containing DTaP vaccines before live viral vaccines sensitize children to gelatin. We received the serum samples of 87 children who had systemic immediate type reactions including anaphylaxis to the vaccines from both physicians and vaccine manufacturers throughout Japan. We then surveyed the DTaP vaccination histories of the children who demonstrated anti-gelatin IgE. Of the above 87 children, 79 (91%) had anti-gelatin IgE. We successfully collected DTaP vaccination histories including the manufacturers' names and numbers of doses on 55 children. Only one child had not received any DTaP vaccine, the other 54 had received gelatin-containing DTaP vaccines and none received gelatin-free DTaP vaccines. We concluded that there was a strong causal relationship between gelatin-containing DTaP vaccination, anti-gelatin IgE production, and risk of anaphylaxis following subsequent immunization with live viral vaccines which contain a larger amount of gelatin. PMID- 10706970 TI - Definition of MHC-restricted CTL epitopes from non-variable number of tandem repeat sequence of MUC1. AB - Mucin1 (MUC1) is expressed ubiquitously on breast cancer cells and is a potential target for the generation of cytotoxic T cells for vaccination against breast cancer. Thus far studies of the immunogenicity of MUC1 have used peptides from the variable number of tandem repeat (VNTR); mice so immunised can generate strong cellular and antibody responses to the VNTR of human MUC1. We now demonstrate that significant CTL and CTLp can be induced to other regions of MUC1. Using the whole native MUC1 molecule, the human milk fat globule membrane antigen (HMFG) linked to mannan, cytotoxic T cell precursors (CTLp) can be generated in BALB/c, C57BL/6, transgenic HLA-A*0201/K(b) and double transgenic HLA-A*0201/K(b)xhuman MUC1 (A2 K(b)MUC1) mice. By immunising with HMFG and testing selectively on (a) extracellular (non-VNTR); (b) VNTR and (c) intracellular peptides, it was shown that all three regions generated effective CTL. Further, the CTL responses to non-VNTR peptides were as strong as those generated to the VNTR. Epitope prediction algorithms were not particularly helpful to describe CTL epitopes: overlapping peptides had to be synthesised and tested to find the epitopes. Thus, for CTL generation, the whole HMFG molecule is a powerful immunogen when linked to mannan, especially as multiple peptide epitopes for presentation by many Class I molecules are contained within the one molecule. Furthermore, Class I restricted MUC1 CTL were generated in double transgenic A2 K(b)MUC1 mice by immunising with mannan-native mucin (HMFG), suggesting that tolerance to MUC1 can be overcome with mannan-HMFG. PMID- 10706971 TI - In search of a new pharmacological treatment for drug and alcohol addiction: N methyl-D-aspartate (NMDA) antagonists. AB - The most challenging aspect of treating alcohol and drug addiction is the relapsing course of these disorders. Although substitution therapies for nicotine and opioid dependence have proven to be relatively effective, there is a need for new pharmacotherapies designed to decrease the frequency and severity of relapse. The aim of this paper is to provide an overview of the potential utility of N methyl-D-aspartate (NMDA) receptor antagonists as treatments for substance abuse as shown in preclinical models and preliminary clinical trials. It is hypothesized that NMDA receptors mediate the common adaptive processes that are involved the development, maintenance, and expression of drug and alcohol addiction. Modulation of glutamatergic neurotransmission with NMDA receptor antagonists offers a novel treatment approach. It is proposed that NMDA antagonists may have multiple functions in treating addictions, including an attenuation of withdrawal effects, normalization of the affective changes following initiation of abstinence which arise from neurochemical changes resulting from chronic addiction, and an attenuation of conditioned responses arising from drug-related stimuli. PMID- 10706972 TI - Drug abuse treatment as an HIV prevention strategy: a review. AB - We review drug abuse treatment as a means of preventing infection with HIV. Thirty-three studies, with an aggregate of over seventeen thousand subjects, were published in peer-reviewed journals from 1988-1998. Research on the utility of drug abuse treatment as an HIV prevention strategy has focused primarily on methadone maintenance treatment (MMT) rather than other modalities such as residential or outpatient drug-free treatment. Recent research provides clear evidence that MMT reduces HIV risk behaviors, particularly needle-use, and strong evidence that MMT prevents HIV infection. There is less definitive evidence that MMT reduces needle-sharing and unsafe sexual behavior, or that other treatment modalities prevent HIV infection. Future research should take into account patient self-selection processes and investigate other treatment modalities for heroin and stimulant abuse to determine their effects on HIV risk behaviors and HIV infection. PMID- 10706974 TI - Comparison of patient self-reports and urinalysis results obtained under naturalistic methadone treatment conditions. AB - This study examined under naturalistic assessment conditions the validity of self reported opiate and cocaine use among 175 veterans enrolled in methadone treatment, and factors related to self-report validity, such as stage in treatment and drug of abuse. Veterans were interviewed by clinical staff about past 30-day drug use with the addiction severity index (ASI), and urinalysis results were obtained for the same 30-day interval assessed with the ASI. Analysis revealed that urinalysis generally produced higher rates of substance use than patient self-report, and with the exception of reported opiate use among new patients presenting for treatment, validity of patient self-reported drug use generally was poor with patients under-reporting both opiate and cocaine use. The findings are in marked contrast to those obtained in other studies in which participants are ensured confidentiality regarding their self-reports. Further, the results raise questions about the utility of self-report measures of substance use to assess patient progress or methadone program performance. PMID- 10706973 TI - Mood state and recent cocaine use are not associated with levels of cocaine cue reactivity. AB - Eighty-one cocaine-dependent outpatients were assessed for their reactions to cocaine-related cues in a laboratory setting. All subjects contributed a urine sample prior to the session. Compared with non-drug control cues, the cocaine stimuli produced increases in physiological arousal, self-reports of high, craving, and withdrawal, and self-reports of negative mood. Subjects who tested cocaine-positive on the day of testing differed only in skin resistance responding from those who tested cocaine-negative. Changes in cue-induced physiological and self-report measures were also not associated with between subject variations in mood as measured by the Profile of Mood States (POMS) questionnaire administered prior to cue assessment. Thus, variations in baseline mood and recent cocaine use history do not introduce an additional source of variability in cue reactivity measurements. However, negative mood states at the start of a session were associated with higher levels of self-reported craving, high, and withdrawal both before and after cue exposure. PMID- 10706975 TI - Cocaine self-administration in monkeys: effects on the acquisition and performance of response sequences. AB - A three-component multiple schedule of intravenous cocaine self-administration (0.01-0.3 mg/kg), repeated acquisition and performance was used to examine the effects of self-administered cocaine on learning in rhesus monkeys. A 0.03 mg/kg infusion of cocaine maintained reliable self-administration without markedly decreasing overall response rate or increasing the percentage of errors in the acquisition and performance components in which food was presented. When saline was substituted for 0.03 mg/kg of cocaine, there was little or no effect on responding in the acquisition or performance components while the number of infusions and response rate in the self-administration component decreased. These effects occurred to a greater extent under a FR 90 schedule (Experiment 2) as compared to a FR 30 schedule (Experiment 1) of cocaine self administration. Substitution of higher infusion doses of cocaine also decreased response rate and the number of infusions in the self-administration components, and substantially decreased responding in the acquisition components; decreases in overall accuracy of responding were evident when responding in this schedule component occurred. Taken together, these data indicate that learning is generally more sensitive than performance to the disruptive effects of self-administered cocaine. PMID- 10706976 TI - Substance Dependence Severity Scale (SDSS): reliability and validity of a clinician-administered interview for DSM-IV substance use disorders. AB - No existing diagnostic interview assesses severity of dependence based on DSM-IV criteria across a range of substances. The Substance Dependence Severity Scale (SDSS) was designed to serve this purpose, consisting of substance-specific scales of both severity and frequency of DSM-IV criteria. This study investigated the reliability and validity of the SDSS. The test-retest reliability of the SDSS in 175 (112 male and 63 female) treated substance users ranged from good to excellent for alcohol, cocaine, heroin and sedatives (interclass correlation coefficients (ICCs)=0.75-0.88 for severity, 0.67-0.85 for frequency). Results for cannabis were lower, ranging from fair to good (ICCs=0.50-0.62). Results for joint rating and internal consistency reliability were comparable to test-retest findings. In addition to indicators of concurrent validity, scale applications are presented and discussed. PMID- 10706978 TI - The suspected association between methamphetamine ('ice') smoking and frequent episodes of alcohol intoxication: data from the 1993 National Household Survey on Drug Abuse. AB - This study estimates the strength of association between frequent episodes of alcohol intoxication and recent smoking of methamphetamine ('ice'). Drawn from the 1993 National Household Survey on Drug Abuse, a total of 101 ice smokers were matched on neighborhood of residence to 816 non-smokers. Based upon conditional logistic regression analyses, persons with daily episodes of alcohol intoxication were an estimated five times more likely to have smoked ice, as compared with non drinkers or drinkers with little or no history of alcohol intoxication. This estimate includes statistical adjustment for potential confounders (e.g. age, sex) and was statistically significant (P=0.01). The association between frequent alcohol intoxication and 'ice smoking' offers an intriguing lead for a broad range of new research. PMID- 10706977 TI - Concurrent and predictive validity of the Substance Dependence Severity Scale (SDSS). AB - This study investigated the concurrent and predictive validity of the Substance Dependence Severity Scale (SDSS), a clinician-administered interview designed to assess the severity and frequency of DSM-IV dependence symptoms for a range of substances. A total of 172 (107 males and 66 females) treated substance users participated in the study. Of those, 89% (n=153) received at least one follow-up interview within 1-6 months of an initial assessment. For alcohol, cocaine and heroin, convergent and discriminant validity was supported by significant relationships between SDSS scores at baseline and other baseline measures of substance use consequences, such as the Addiction Severity Index (ASI), as well as significant relationships between SDSS change scores from baseline to follow up and change scores of other measures of consequences. SDSS scores were significantly associated with time to first post treatment use of alcohol, cocaine and heroin, although the nature of the associations was complex. Scale applications and areas for further study are discussed. PMID- 10706979 TI - Convulsive status epilepticus following abrupt high-dose benzodiazepine discontinuation. AB - The misuse of benzodiazepines (BNZ)s may result in serious side effects. Three cases of convulsive status epilepticus (CSE) following abrupt discontinuation of long-term use of 25 mg of lorazepam in one patient and more than 20 mg of flunitrazepam in two patients are presented; they were non-epileptics and free of other high-risk factors for seizures. A favorable outcome for all three cases was noted. They remain free of seizures without antiepileptic treatment. Nevertheless, because of the extensive use of benzodiazepines, such rare high risk side effects must be emphasized. PMID- 10706980 TI - If patients with schizophrenia have small brains, why don't they have small heads? AB - Although patients with schizophrenia have reduced brain size, there is no conclusive evidence that they have reduced head size. This begs the question: 'What is the precise relationship between head size and brain size?' We used a unique osteological collection to explore the relationship between external head measures and cranial capacity. The external measures accounted for, at most, 60% of the variance in cranial capacity - a value low enough to question the oft assumed tight relationship between head measures and brain size. Obviously, various tissues and spaces [skull, sinus, muscle (frontalis, temporalis and occipitalis), subcutaneous fat and epidermal layers] contribute to head size without contributing to brain volume. The contribution of these other tissues and spaces tends to decrease the signal and increase the noise in the estimation of brain volume. Thus, it is understandable that patients with schizophrenia can have reduced cranial capacity and not reduced head size. PMID- 10706981 TI - Membrane phospholipid abnormalities in postmortem brains from schizophrenic patients. AB - Previous studies in schizophrenia have shown alterations in membrane phospholipids and polyunsaturated fatty acids. However, these studies have primarily examined peripheral (non-neuronal) cell types. The purpose of the present study was to examine whether the membrane deficits seen in peripheral tissues are also observed in the brain. The caudate was the primary region of interest for this study. Using high-pressure liquid chromatography in conjunction with an evaporative light-scattering detector, we first measured the level of various membrane phospholipids (PL) in schizophrenic (n=11) and control groups with (n=7) and without (n=14) other mental disorders. Polyunsaturated fatty acids (PUFAs) were then determined by capillary gas chromatography. Within groups, there are no significant correlations between membrane PL levels and other collection and demographic parameters including age, postmortem interval, storage time and brain weight. Significantly lower amounts of phosphatidylcholine and phosphatidylethanolamine were found in postmortem brain tissue from schizophrenic patients than in those from control groups, even after accounting for potential confounds. In addition, strong reductions of total PUFAs and saturated fatty acids were found in schizophrenic brains, relative to control brains. Specifically, the reduced PUFAs were largely attributable to decreases in arachidonic acid (AA) and, to a lesser extent, its precursors, linoleic and eicosadienoic acids. There are no significant differences between the control groups with and without other mental disorders. The present findings suggest that deficits identified in peripheral membranes may also be present in the brain from schizophrenic patients. Such a deficit in membrane AA may contribute to the many biological, physiological, and clinical phenomena observed in schizophrenia. PMID- 10706982 TI - Cerebellar vermis area in schizophrenic patients - a post-mortem study. AB - Neuroimaging studies of cerebellar atrophy in schizophrenia have yielded contradictory results. In computer-tomography (CT) studies, cerebellar atrophy was found in up to 40% of schizophrenic patients. However, several recent magnetic resonance imaging (MRI) studies could not replicate these early findings; in addition, contradictory observations of enlargement of vermal structures were reported. In contrast to the number of CT and MRI studies, there are only a few neuropathological reports on this subject. In a post-mortem study we analyzed the midsagittal vermal area of formaldehyde-fixed cerebella of 12 deceased schizophrenic patients and 12 age- and gender-matched control subjects by using morphometrical methods. Statistical analysis using ANOVA revealed no significant group effects, but there were interactions with gender and cerebellar brain weight. In view of the present results, the common concept of cerebellar atrophy in schizophrenic patients appears premature. Gender effects and secondary processes (e.g., relevant alcohol or drug abuse) cannot be excluded as possible factors causing decrease of vermal areas in schizophrenic patients. PMID- 10706983 TI - Regional cerebral blood flow in male schizophrenic patients performing an auditory discrimination task. AB - Regional cerebral blood flow (rCBF) was measured in 11 schizophrenic patients amid 10 normal controls, both at rest and while performing an auditory discrimination task. Single photon emission computed tomography with technetium 99m hexamethylpropylene amine oxime was used for quantitative evaluation of rCBF. The schizophrenic patients showed greater rCBF in the temporal and parietal regions at rest than the controls, but no abnormalities were found in frontal perfusion. During task performance. on the other hand, the patients showed a reduced frontal rCBF. whereas there was no group difference in rCBF in the temporal and parietal regions. In addition, the left> right hemisphere asymmetries of rCBF observed in the controls during task performance were not present in the patients. although there was no group difference in hemisphere laterality in rCBF at rest. These findings suggest that the employment of a cognitive task for neuroimaging studies is useful for detecting abnormalities of brain activation. such as hypofrontality and altered hemisphere laterality. in patients with schizophrenia. PMID- 10706984 TI - Tardive dyskinesia is associated with impaired retrieval from long-term memory: the Curacao Extrapyramidal Syndromes Study: IV. AB - Tardive dyskinesia may be associated with cognitive dysfunction. It is not clear whether this dysfunction occurs in the form of a global or specific cognitive deficit. A cross-sectional study was conducted in a well-defined catchment area (Curacao, The Netherlands Antilles). All schizophrenic inpatients who had been taking neuroleptic medication for at least 3months and who were younger than 65years were included (n=53). Tardive dyskinesia was assessed with the Abnormal Involuntary Movement Scale. The neuropsychological assessment comprised tests of memory, executive function, and speed of information processing. Of the six cognitive measures, only delayed recall was significantly associated with orofacial dyskinesia. Limb-truncal dyskinesia was not associated with any of the cognitive measures. The pattern of memory impairment is consistent with there being a frontal-subcortical disturbance in orofacial dyskinesia. The results underscore the importance of using specific cognitive test procedures in the search for the cognitive correlates of dyskinesia. PMID- 10706985 TI - The longitudinal relationship of clinical symptoms, cognitive functioning, and adaptive life in geriatric schizophrenia. AB - Cognitive dysfunction is increasingly being recognized as a major contributor to the adaptive impairment seen in most patients with schizophrenia. Reported here is a prospective longitudinal evaluation of the relationship between cognitive and adaptive functioning in elderly patients with schizophrenia. It was hypothesized that baseline cognitive and negative, but not positive symptoms, would be predictive of cross-sectional impairment and longitudinal outcome. Subjects were 168 elderly patients with schizophrenia, free of major neurological disorders, who were residents of a long-term psychiatric facility. Subjects were assessed at baseline and again an average of 15months later. The PANSS was used to assess the severity of symptoms of schizophrenia. Cognitive symptoms were assessed using the components of the CERAD cognitive battery. Social and adaptive functioning was assessed using the SAFE scale. Spearman correlations were determined among clinical variables, and the rank ordering of prediction of SAFE scale scores at follow-up was determined using a stepwise regression procedure. At follow-up, adaptive life skills correlated with cognitive performance and negative symptoms (Spearman rho values 0. 41-0.57, all p values <0.0001), but not positive symptoms (r=0.09, n. s.). Among cognitive tasks, verbal learning and memory were most highly correlated with adaptive skills at follow-up. These results confirm and extend previous studies that indicate that cognitive impairments are predictive, both cross-sectionally and longitudinally, of adaptive life skills in persons with schizophrenia. Negative symptoms, but not positive symptoms, were correlated with impaired adaptive skills. Taken together, these results underscore the need to develop more effective treatments for cognitive and negative symptoms in schizophrenia. PMID- 10706986 TI - Emotional processing in schizophrenia across cultures: standardized measures of discrimination and experience. AB - Schizophrenia appears quite similar across a range of cultures. However, variability has been noted, and understanding the variant and invariant features of the disorder is necessary for elucidating its biological and environmental basis. Evidence of prominent emotion processing deficits in schizophrenia, including perceptual and experiential aspects, led us to extend the paradigm of standardized measures cross-culturally. We assessed performance of American, German, and Indian patients with schizophrenia and healthy controls on standardized emotion discrimination and experience (mood induction) procedures using happy, sad, and neutral facial expressions of Caucasian actors. Participants were 80 Americans (40 patients; 40 controls), 48 Germans (24 patients; 24 controls), and 58 Indians (29 patients; 29 controls). Face discrimination performance was impaired across patient groups, but was most impaired in those of Indian origin. Lower performance was also found in Indian controls, relative to their American and German counterparts. Mood induction produced weaker effects in all patient groups relative to their respective controls. The results supported the feasibility of cross-cultural comparisons and also emphasized the importance of poser ethnic background for facial affect identification, while poser ethnicity was less consequential for mood induction effects. Emotion processing deficits in schizophrenia may add to the clinical burden, and merit further examination. PMID- 10706987 TI - Reliability of the life chart schedule for assessment of the long-term course of schizophrenia. AB - We report on the inter-rater reliability of the Life Chart Schedule (LCS). The LCS is designed to assess the long-term course of schizophrenia in four key domains (symptoms, treatment, residence, and work) over two time periods (past two years, entire period of illness). The subjects were 27 consecutive admissions to a schizophrenia research unit. The LCS was filled out by pairs of raters, blinded to each others' ratings, using the same data (interview with subject and chart). Reliability was examined for 45 LCS ratings selected from all four domains and both time periods. Selected ratings pertained to the duration of specified experiences, the quality of these experiences, and the long-term time trend. The kappa statistic and the intra-class correlation coefficient (ICC) were used to determine inter-rater reliability for continuous and categorical ratings, respectively. LCS ratings proved reliable in all four key domains and both time periods. The reliability was fair to excellent for ratings of duration of experience (ICC ranged from 0.53 to 0.99), quality of experience (kappa ranged from 0.46 to 0. 92) and long-term time trends (kappa ranged from 0.66 to 0.94). The LCS can be used to obtain reliable ratings of the long-term course of schizophrenia in multiple domains. PMID- 10706988 TI - Type of feeding during infancy and later development of schizophrenia. AB - Nutrition may be critical for neurodevelopment and can affect the later development of schizophrenia. Recently, a marked reduction in breast-feeding was reported in infants that developed schizophrenia in later life (McCreadie, R.G., 1997. The Nithsdale Schizophrenia Surveys. 16. Breast-feeding and schizophrenia: preliminary results and hypothesis. Br. J. Psychiatr. 170, 334-337). In the present study, we investigated feeding patterns during the infancy of 100 schizophrenia patients, 37 of their siblings and 200 age-matched healthy controls using a structured written questionnaire. Having been breast-fed was not negatively associated with schizophrenia. PMID- 10706989 TI - 5-HT(3) receptor antagonists and anxiety; a preclinical and clinical review. AB - The present paper reviews the evidence for anxiolytic activity of 5-HT(3) receptor antagonists in animal models of anxiety and in clinical trials in humans. Compared to the established anxiolytics (benzodiazepine receptor agonists and, to a lesser extent, 5-HT(1A) receptor agonists) 5-HT(3) receptor antagonists display a different anxiolytic profile. They are anxiolytic in a limited number of animal anxiety models. If active, they often are very potent and display bell shaped dose response curves, whereas the ratio between therapeutic activity and side effects appears remarkably large. 5-HT(3) receptor antagonists remain active after chronic dosing and no indications for tolerance, dependence or rebound effects were found, which seems to make these drugs an attractive alternative to the benzodiazepines. However, the large body of animal data indicating a complete lack of psychotropic activity of 5-HT(3) receptor antagonists weakens the prediction of anxiolytic activity in these drugs. Human data are also controversial; some investigators have reported positive effects in anxiety disorders (panic disorder, GAD), others did not. It can be concluded that 5-HT(3) receptor antagonists do not represent a breakthrough in the treatment of various anxiety disorders, as initially suggested. PMID- 10706990 TI - Intracellular pH modulates spontaneous and epileptiform bioelectric activity of hippocampal CA3-neurones. AB - A growing body of evidence hints at intracellular free protons to be involved in the modulation of electric activity of cortical neurones. In this study we demonstrate that application of the weak acid propionate (2.5-20 mM) transiently lowers intracellular pH (pH(i)) of BCECF-AM loaded CA3-neurones in hippocampal slices. The predictability of this acidification prompted us to use propionate as a tool to investigate effects of pH(i) on spontaneous bioelectric activity (SBA) and epileptiform activity (EA, induced by bicuculline, caffeine or low magnesium) of CA3 neurones: SBA and EA were transiently suppressed by 2-20 mM propionate - coinciding with the transient neuronal acidification. As activation of Na(+)/H(+) exchangers (NHE) is involved in the recovery from neuronal acidosis and NHE inhibition alone is known to increase the activity of intracellular free protons of hippocampal neurones, we tested the effect of the NHE-blockers amiloride (0.5 1 mM) or HOE642 (200 microM) on SBA and EA of CA3-neurones. Long-term application of NHE-inhibitors alone continuously suppressed SBA and EA, which recovered during additional exposure to the weak base trimethylamine (5-10 mM). Simultaneous administration of propionate and NHE-blockers intensified the inhibition of neuronal activity. Together, these results indicate that intracellular acidification inhibits bioelectric activity of hippocampal CA3 neurones. This supports the hypothesis that pH(i) contributes to the control of cortical excitability. PMID- 10706991 TI - Increased adenylyl cyclase type 1 mRNA, but not adenylyl cyclase type 2 in the rat hippocampus following antidepressant treatment. AB - The adenylyl cyclase (AC) system is affected by several types of antidepressant treatments, and increased activity in this system is linked to the therapeutic action of antidepressants. The present study was undertaken to compare the effects of single-dose and long-term treatment with the selective serotonin reuptake inhibitor, citalopram (10 mg/kg, i.p.), on the AC system in the male rat brain of Wistar rats. Furthermore, we compared the effects of long-term citalopram and lithium treatments on the AC system. Long-term citalopram, but not single-dose administration, increased the AC type 1 mRNA in the hippocampus, whereas type 2 mRNA was unaffected. Long-term lithium treatment also increased AC1 in the hippocampus. However, long-term citalopram treatment did not increase AC type 1 protein, basal or forskolin-stimulated AC activity, but GTP increased AC activity in the hippocampus. This may indicate enhanced AC/G protein coupling. Thus, citalopram may increase cAMP signalling by acting on components of the AC system without affecting the protein level of the AC type 1. PMID- 10706992 TI - Decreased platelet monoamine oxidase activity in female bulimia nervosa. AB - The involvement of brain serotonin systems in the pathophysiology of eating disorders has been repeatedly demonstrated in recent studies. Platelet MAO activity is an index of brain serotonin activity and lowered platelet MAO levels have been found in association with impulsive behaviors. In addition, some preliminary reports indicate that platelet MAO could be lowered in eating disorder patients. METHODS: 47 patients with DSM-IV eating disorders were studied, including 30 with bulimia nervosa and 17 with anorexia nervosa binge eating-purging type. Platelet MAO activity was measured by isotopic methods using C-14 benzylamine and compared with a control group of 30 healthy subjects. Impulsive personality features were studied with specific rating scales. RESULTS: Platelet MAO activity was significantly lower (4.4+/-2.4 nmol/h/10(8) platelets) in the bulimic patients than in the control group (6.9+/-2.5) (p<0.001). No significant differences were found between pure bulimics and binge eating-purging anorectics. Platelet MAO was inversely and significantly correlated with scores on impulsivity scales and with borderline personality disorder characteristics. CONCLUSIONS: Platelet MAO activity is lowered in patients with bulimia, which may reflect dysfunction in impulse control mechanisms. Since platelet MAO has a predominant genetic component, there is need for studies on the association of low platelet MAO and higher risk for developing eating disorders. PMID- 10706994 TI - MAO-A and MAO-B activities in rat striatum, frontal cortex and liver are unaltered after long-term treatment with fluvoxamine and desipramine. AB - In the course of investigating the mechanisms underlying the beneficial effect of fluvoxamine augmentation on negative symptoms of schizophrenia, the authors found a reduction in human platelet monoamine oxidase-B activity after 5 weeks of treatment. This unexpected finding raised the possibility that MAO activity may be one of the factors altered by chronic tricyclic or SSRI antidepressant treatment. The current study examined the effect of long-term administration, up to 6 weeks, of fluvoxamine, desipramine or saline on MAO-A and MAO-B activities in rat striatum, frontal cortex and liver. No differences were noted between drug treated groups and their saline-treated controls. The hypothesis that long-term treatment with tricyclic and SSRI antidepressants alters MAO activity was not supported. MAO is not among proteins whose activity may be altered by chronic tricyclic or SSRI antidepressant treatment. PMID- 10706993 TI - Effects of atypical antipsychotics on the inflammatory response system in schizophrenic patients resistant to treatment with typical neuroleptics. AB - There is now some evidence that schizophrenia may be accompanied by an activation of the inflammatory response system (IRS) and that typical antipsychotics may suppress some signs of IRS activation in that illness. This study was carried out to examine (i) the serum concentrations of interleukin-6 (IL-6), IL-6 receptor (IL-6R), IL-1R antagonist (IL-1RA) and Clara Cell protein (CC16), an endogenous anticytokine, in nonresponders to treatment with typical neuroleptics and (ii) the effects of atypical antipsychotics on the above IRS variables. The above parameters were determined in 17 patients with treatment-resistant schizophrenia (TRS) to treatment with neuroleptics and in seven normal volunteers and 14 schizophrenic patients who had a good response to treatment with antipsychotic agents. Patients with TRS had repeated measurements of the IRS variables before and 2 and 4 months after treatment with atypical antipsychotics. Serum IL-6 was significantly higher in schizophrenic patients, irrespective of their response to typical antipsychotics, than in normal controls. Serum IL-1RA was significantly higher in the TRS patients than in controls, whereas responders took up an intermediate position. The serum concentrations of CC16 were significantly lower after treatment with atypical antipsychotics during 4 months than before treatment. It is concluded that (i) schizophrenia and, in particular, TRS is characterized by an activation of the monocytic arm of cell-mediated immunity and (ii) atypical antipsychotics may decrease the anti-inflammatory capacity of the serum in TRS patients. PMID- 10706995 TI - Effect of single intracerebroventricular injection of alpha-interferon on monoamine concentrations in the rat brain. AB - The effect of a single intracerebroventricular (i.c.v.) injection of alpha-IFN on levels of central monoamines and their metabolites in six brain regions (frontal cortex, striatum, hypothalamus, hippocampus, mid brain and medulla) of the rat was investigated. Wistar rats (n=10) were decapitated 2 h after i.c.v. injection of alpha-IFN. The brain tissues were homogenized, and monoamine concentrations were measured by high-performance liquid chromatography with an electrochemical detector. The levels of 5-hydroxytryptamine (5-HT) were significantly reduced in the frontal cortex in a dose-dependent manner, and the levels of both 5-HT and 5 hydroxyindoleacetic acid (5-HIAA) were reduced in the mid brain and the striatum. The levels of noradrenaline (NA) were also significantly reduced in a dose dependent manner in the frontal cortex. Some neurophysiological changes that affect activity of the noradrenergic or/and the serotonergic neuron system may occur during IFN therapy. PMID- 10706996 TI - Suicide by antidepressant intoxication identified at autopsy in Vienna from 1991 1997: the favourable consequences of the increasing use of SSRIs. AB - In the area of Vienna, any person dying under questionable circumstances is examined at the Institute of Forensic Medicine, where the cause of death is determined by means of autopsy and chemical analysis. Our study on fatal intoxications was performed in the period between 1991 and 1997, when selective serotonin reuptake inhibitors (SSRIs) were establishing themselves on the market, reaching the top of prescription statistics. Tricyclic antidepressants (TCAs) were involved in 30 single- and 127 multiple-substance intoxications, with amitriptyline and doxepin being the most frequently used drugs. SSRIs were involved in five multiple-substance intoxications. The f-value, which refers to the number of deaths per million defined daily doses prescribed, was found to be significantly (PBL) direction in the accelerated model were approximately twice those in the traditional model. As observed with microscopy and transepithelial electrical resistance measurements, this difference was attributed to less confluent and differentiated Caco-2 cell monolayers in the accelerated model. However, there were no significant differences in rank ordering of the compounds. The expression of P-glycoprotein in the accelerated model was shown to be significantly less than that in the traditional model. This resulted in lower permeability directional ratios defined as the ratio between permeability coefficients from BL-->AP and from AP-->BL for compounds that were cellular efflux pump substrates. The accelerated model may not be suitable for studying cellular efflux pumps such as P-glycoproteins. However, it is a feasible alternative to the traditional model for rank ordering of compounds in the process of drug discovery and development by significantly improving the turnover time and labor efficiency. This makes it an excellent Caco-2 cell permeability model for high throughput screening. PMID- 10707015 TI - Disorder and twinning in molecular crystals: impurity-induced effects in adipic acid. AB - The variation in physical properties of crystals grown in the presence of additives or impurities have previously been attributed to lattice disorder developed during crystallization. Adipic acid crystallized in the presence of a variety of stereochemically related impurities typifies such behavior with disorder manifest in variations of dissolution rates and enthalpies of solution and fusion. In this case the most extreme habit, produced by the presence of added monoalkanoic acids, is a rounded dumbbell that was suggested previously to be a twinned crystal. In this contribution such crystals are fully characterized both through their external morphology and by means of single crystal X-ray diffraction. These techniques show that these particles are not twinned but rather are disordered single crystals comprising a small number of slightly misaligned domains. The interaction between additive and substrate is modeled and new additives selected that induce the formation of true mechanical twins in adipic acid. PMID- 10707016 TI - Lyophilization of cationic lipid-protamine-DNA (LPD) complexes. AB - Cationic lipid-based gene delivery systems have shown promise in transfecting cells both in vitro and in vivo. However, these systems tend to form aggregates in liquid formulation during storage, which has limited their clinical applications. As a result, lyophilization of these systems has recently become a subject of increasing interest. In this paper, lyophilization of LPD, a novel cationic lipid-based gene delivery system, was studied. Both particle size and transfection efficiency could be preserved in the presence of sufficient amount of appropriate lyoprotectant. A series of monosaccharides and disaccharides, including dextrose, galactose, mannose, lactose, maltose, sucrose and trehalose, were evaluated for their lyoprotective effect and disaccharides showed more superior protection to monosaccharides. The effect of different freezing protocols for lyophilization was also evaluated and no significant difference was found. However, for freeze-thawing, fast freezing caused less aggregation. Finally, nonlyophilized LPD and LPD lyophilized with 10% sucrose were stored at different temperatures and their stability was followed for eight weeks. Lyophilized LPD could be stored at room temperature without significant change in particle size or loss of transfection efficiency. PMID- 10707017 TI - Correlation between drug release kinetics from proteineous matrix and matrix structure: EPR and NMR study. AB - The present study was conducted in order to probe the microstructure, microviscosity, and hydration properties of matrices containing two model drugs, naproxen sodium (NS) and naproxen (N), and egg albumin (EA) as matrix carrier. The results suggested that N release from EA matrix was controlled by a bulk erosion mechanism in combination with additional processes (crystal dissolution/crystallization rate) compared with NS matrix, which behaved as a non erodible matrix and drug release occurred by diffusion through the gel. Using EPR technique it has been shown that incorporating NS into EA matrix strongly influences the microstructure of the protein gel, and hence the transport of the penetrant within the matrix, compared with matrices containing N. The presence of NS increased the protein chain mobility and hydration which supports our previous results showing that NS cause unfolding of EA. In contrast, N caused only marginal effect on EA chain mobility. The gel formed in EA/NS matrices was more porous compared with EA/N matrices as revealed by the lower rotational correlation time of PCA (lower microviscosity) in EA/NS matrices compared with EA/N. However, EA/N gelled matrices were more heterogeneous, i.e., containing a higher number of components having different mobility. The T(1) and T(2) relaxation studies by NMR provided an additional support for the higher chain hydration in EA/NS matrices compared with EA/N as indicated by the higher relaxation rates in the gelled matrices. Internal pH measurements by EPR revealed that the micro-pH inside 100% EA and 50/50 EA/N matrices were lower than 50/50 EA/NS matrices and in all cases lower than the penetrating buffer pH. The lower pH compartment formed in N matrices affected N solubility and crystal dissolution rate, which can explain its lower release rate compared with EA, from the same formulation. The EPR and NMR data supports our findings that NS caused unfolding of the protein, affected matrix structure, and converted it to a hydrophobic non erodible matrix compared with EA/N matrix in which the native properties of EA were mainly retained. PMID- 10707018 TI - Wetting properties of bile salt solutions and dissolution media. AB - The objective of this study was to determine the extent to which specific bile salt solutions and compendial dissolution media differ in their ability to wet a model surface. Solutions were examined in the concentration range of bile salts found in the gastrointestinal tract and at pH values approximating those of the stomach and small intestine. Wetting was evaluated from measurement of the surface tension of the solutions and contact angles of sessile drops on poly(methyl methacrylate). Compendial dissolution media had higher surface tensions and contact angles than bile salt solutions at 10 mM. Individual bile salts at 10 mM varied in surface tension lowering and contact angles. The contact angle-concentration profiles achieved plateau values at 2.5 mM. Dewetting was observed at low bile salt concentrations at pH 7.5. Calculated adhesion tension and interfacial tension were consistent with this behavior. The effect was attributed to the influence of the substrate surface charge on the orientation of the adsorbed bile salt molecule. Adhesion tension profiles showed that from low (<0.5 mM) to moderate (2 mM) concentrations preferential bile salt adsorption to the liquid-vapor interface occurred, but at higher values (>2 mM) the preference shifted toward the solid-liquid interface. These results have implications in the design of physiologically based dissolution media. PMID- 10707019 TI - Validation of excised bovine nasal mucosa as in vitro model to study drug transport and metabolic pathways in nasal epithelium. AB - The present work aims at the validation of excised bovine nasal mucosa as an in vitro model to address transport and metabolism pathways relative to the nasal mucosal uptake of therapeutic peptides. Preservation of the viability of the excised tissue in the course of in vitro studies of up to 3 h was demonstrated by (i) positive viability staining, (ii) constant transepithelial electrical resistance (42 +/- 12 Omega cm(2)), (iii) constant rates of metabolic turnover, and (iv) linear permeation profiles of therapeutic peptides and (3)H-mannitol. Using 1-leucine-4-methoxy-2-naphthylamide as a model substrate, we observed no difference between bovine and human nasal aminopeptidase activity. By a series of therapeutic peptides, no direct correlation was found between their effective permeability coefficients (from 0. 1 x 10(-5) to 5 x 10(-5) cm s(-1)) and their respective molecular masses (from 417 to 3,432 Da), indicating that other factors dominate nasal permeability. For instance, the permeabilities of metabolically labile peptides were concentration dependent and saturable, as demonstrated for two short thymopoietin fragments, Arg-Lys-Asp (TP3) and Arg-Lys-Asp-Val (TP4). By permeation studies using gonadorelin and two gonadorelin derivatives, buserelin and Hoe 013, without and in the presence of the chemical enhancer bacitracin, we also verified the ability of the model to assess chemical enhancer effects and their reversibility. In conclusion, our work demonstrates the potential of the investigated in vitro model, excised bovine nasal mucosa, to explore mechanistic aspects of nasal transport and metabolism of therapeutic peptides. PMID- 10707020 TI - Relationship between the crystallization rates of amorphous nifedipine, phenobarbital, and flopropione, and their molecular mobility as measured by their enthalpy relaxation and (1)H NMR relaxation times. AB - Isothermal crystallization of amorphous nifedipine, phenobarbital, and flopropione was studied at temperatures above and below their glass transition temperatures (T(g)). A sharp decrease in the crystallization rate with decreasing temperature was observed for phenobarbital and flopropione, such that no crystallization was observed at temperatures 20-30 degrees C lower than their T(g) within ordinary experimental time periods. In contrast, the crystallization rate of nifedipine decreased moderately with decreasing temperature, and considerable crystallization was observed at 40 degrees C below its T(g) within 4 months. The molecular mobility of these amorphous drugs was assessed by enthalpy relaxation and (1)H-NMR relaxation measurements. The enthalpy relaxation time of nifedipine was smaller than that of phenobarbital or flopropinone at the same T - T(g) values, suggesting higher molecular mobility of nifedipine. The spin-lattice relaxation time in the rotating frame (T(1rho)) decreased markedly at temperature above T(g). The slope of the Arrhenius type plot of the T(1rho) for nifedipine protons changed at about 10 degrees C below the T(g), whereas the slope for phenobarbital protons became discontinuous at about 10 degrees C above the T(g). Even at temperatures below its T(g), the spin-spin relaxation process of nifedipine could be described by the sum of its Gaussian relaxation, which is characteristic of solid protons, and its Lorentzian relaxation, which is characteristic of protons with higher mobility. In contrast, no Lorentzian relaxation was observed for phenobarbital or flopropione at temperatures below their T(g). These results also suggest that nifedipine has higher molecular mobility than phenobarbital and flopropione at temperatures below T(g). The faster crystallization of nifedipine than that of phenobarbital or flopropione observed at temperatures below its T(g) may be partly ascribed to its higher molecular mobility at these temperatures. PMID- 10707021 TI - Interpretation of relaxation time constants for amorphous pharmaceutical systems. AB - The molecular mobility of amorphous pharmaceutical materials is known to be a key factor in determining their stability, reactivity, and physicochemical properties. Usually such molecular mobility is quantified using relaxation time constants. Typically relaxation processes in amorphous systems are non exponential and relaxation time constants are usually obtained from experimental data using a curve fitting procedure involving the empirical Kohlrausch-Williams Watts (KWW) equation. In this article we explore the possible relationship between the KWW curve fitting parameters (tau(KWW), beta(KWW)) and common statistical measures of the average and the distribution (e.g., median, standard deviation) of the relaxation time values. This analysis is performed for several common statistical distributions (e.g., normal, lognormal, and Lorentzian), and the results are compared and analyzed in the context of pharmaceutical product stability predictions. The KWW function is able to describe relaxation processes stemming from several different statistical distribution functions. Under some circumstances the "average" relaxation time constant of the KWW equation (tau(KWW)) is significantly different from common statistical measures of the central value of a distribution (e.g., median). Simply knowing the relaxation time constants from the fit of the KWW equation is not sufficient to completely characterize and quantify the molecular mobility of amorphous pharmaceutical materials. An appreciation of the distribution of relaxation times and the resulting effects upon the KWW constants should be considered to be essential when working with amorphous pharmaceutical materials, especially when attempting to use relaxation time constants for predicting their physical or chemical stability. PMID- 10707022 TI - The thermophilic esterase from Archaeoglobus fulgidus: structure and conformational dynamics at high temperature. AB - The esterase from the hyperthermophilic archaeon Archaeoglobus fulgidus is a monomeric protein with a molecular weight of about 35.5 kDa. The enzyme is barely active at room temperature, displaying the maximal enzyme activity at about 80 degrees C. We have investigated the effect of the temperature on the protein structure by Fourier-transform infrared spectroscopy. The data show that between 20 degrees C and 60 degrees C a small but significant decrease of the beta-sheet bands occurred, indicating a partial loss of beta-sheets. This finding may be surprising for a thermophilic protein and suggests the presence of a temperature sensitive beta-sheet. The increase in temperature from 60 degrees C to 98 degrees C induced a decrease of alpha-helix and beta-sheet bands which, however, are still easily detected at 98 degrees C indicating that at this temperature some secondary structure elements of the protein remain intact. The conformational dynamics of the esterase were investigated by frequency-domain fluorometry and anisotropy decays. The fluorescence studies showed that the intrinsic tryptophanyl fluorescence of the protein was well represented by the three exponential model, and that the temperature affected the protein conformational dynamics. Remarkably, the tryptophanyl fluorescence emission reveals that the indolic residues remained shielded from the solvent up to 80 degrees C, as shown from the emission spectra and by acrylamide quenching experiments. The relationship between enzyme activity and protein structure is discussed. PMID- 10707023 TI - Receptor-binding conformation of the "ELR" motif of IL-8: X-ray structure of the L5C/H33C variant at 2.35 A resolution. AB - The "ELR" (Glu-Leu-Arg) tripeptide sequence near the N-terminus of interleukin-8 (IL-8) contributes a large part of the receptor binding free energy. Prior X-ray and nuclear magnetic resonance (NMR) structures of IL-8 have shown this region of the molecule to be highly mobile. We reasoned that a hydrophobic interaction between the leucine and the neighboring beta-turn might exist in the receptor binding conformation of the N-terminus. To test this hypothesis, we mutated two residues to cysteine and connected the N-terminus to the beta-turn. The mutant retains receptor binding affinity reasonably close to wild type and allows the characterization of a high-affinity conformation that may be useful in the design of small IL-8 mimics. The L5C/H33C mutant is refined to R-values of R = 20.6% and Rfree = 27.7% at 2.35 A resolution. Other receptor binding determinants reside in the "N-loop" found after "ELR" and preceding the first beta-strand. All available structures of IL-8 have been found with one of two distinct N-loop conformations. One of these is relevant for receptor binding, based on NMR results with receptor peptides. The other conformation obscures the receptor-peptide binding surface and may have an undetermined but necessarily different function. PMID- 10707024 TI - Electrostatic strengths of salt bridges in thermophilic and mesophilic glutamate dehydrogenase monomers. AB - Here we seek to understand the higher frequency of occurrence of salt bridges in proteins from thermophiles as compared to their mesophile homologs. We focus on glutamate dehydrogenase, owing to the availability of high resolution thermophilic (from Pyrococcus furiosus) and mesophilic (from Clostridium symbiosum) protein structures, the large protein size and the large difference in melting temperatures. We investigate the location, statistics and electrostatic strengths of salt bridges and of their networks within corresponding monomers of the thermophilic and mesophilic enzymes. We find that many of the extra salt bridges which are present in the thermophilic glutamate dehydrogenase monomer but absent in the mesophilic enzyme, form around the active site of the protein. Furthermore, salt bridges in the thermostable glutamate dehydrogenase cluster within the hydrophobic folding units of the monomer, rather than between them. Computation of the electrostatic contribution of salt bridge energies by solving the Poisson equation in a continuum solvent medium, shows that the salt bridges in Pyrococcus furiosus glutamate dehydrogenase are highly stabilizing. In contrast, the salt bridges in the mesophilic Clostridium symbiosum glutamate dehydrogenase are only marginally stabilizing. This is largely the outcome of the difference in the protein environment around the salt bridges in the two proteins. The presence of a larger number of charges, and hence, of salt bridges contributes to an electrostatically more favorable protein energy term. Our results indicate that salt bridges and their networks may have an important role in resisting deformation/unfolding of the protein structure at high temperatures, particularly in critical regions such as around the active site. PMID- 10707025 TI - Modeling of ion permeation in calcium and sodium channel selectivity filters. AB - Structure-function studies have shown that it is possible to convert a sodium channel to a calcium-selective channel by a single amino acid substitution in the selectivity filter locus. Ion permeation through the "model selectivity filter" was modeled with a reduced set of functional groups representative of the constituent amino acid side chains. Force-field minimizations were conducted to obtain the energy profile of the cations as they get desolvated and bind to the "model selectivity filter." The calculations suggest that the ion selectivity in the calcium channel is due to preferential binding, whereas in the sodium channel it is due to exclusion. Energetics of displacement of a bound cation from the calcium "model selectivity filter" by another cation suggest that "multi-ion mechanism" reduces the activation barrier for ion permeation. Thus, the simple model captures qualitatively most of the conduction characteristics of sodium and calcium channels. However, the computed barriers for permeation are fairly large, suggesting that ion interaction with additional residues along the transport path may be essential to effect desolvation. PMID- 10707026 TI - Transcriptional repressor CopR: structure model-based localization of the deoxyribonucleic acid binding motif. AB - The plasmid pIP501 encoded transcriptional repressor CopR is one of the two regulators of plasmid copy number. CopR binds as a dimer to a nearly palindromic operator with the consensus sequence 5'-CGTG. Intermediate sequence searches revealed a significant structural relationship between CopR and the bacteriophage P22 c2 and the 434 c1 repressors. In this report we describe the experimental verification of a CopR homology model, which is based on a fairly low-sequence identity of 13.8% to P22 c2 repressor. A model for the complex of CopR with the deoxyribonucleic acid (DNA) target was built on the basis of experimental footprinting data, the above-mentioned CopR homology model, and the crystal structure of the 434 c1 repressor-DNA complex. Site-directed mutagenesis was used to test the function of amino acids involved in sequence and nonsequence-specific DNA recognition and amino acids important for correct protein folding. CD measurements were performed to detect structural changes caused by the mutations. Exchanges of residues responsible for sequence-specific DNA recognition reduced binding to a nonspecific level. Mutations of amino acids involved in nonspecific DNA binding lead to decreased binding affinity while maintaining selectivity. Substitution of amino acids necessary for proper folding caused dramatic structural changes. The experimental data support the model of CopR as a helix turn-helix protein belonging to the lambda repressor superfamily. PMID- 10707027 TI - Cold denaturation of alpha-lactalbumin. AB - We have investigated the thermal unfolding of bovine alpha-lactalbumin by means of circular dichroism spectroscopy in the far- and near-ultraviolet regions, and shown that the native alpha-lactalbumin undergoes heat and cold denaturation. The guanidine hydrochloride-induced unfolding of alpha-lactalbumin was also investigated by circular dichroism spectroscopy at various temperatures from 261 to 318 K. It is shown that the population of the molten globule state is strongly dependent on temperature and that the molten globule state does not accumulate during the guanidine hydrochloride-induced unfolding transition at 261 K. Our results indicate that the molten globule state of alpha-lactalbumin undergoes cold denaturation as the native alpha-lactalbumin does, and that the heat capacity change of unfolding from the molten globule to the unfolded state is positive and significant. The present results further support the idea that the molten globule and the unfolded states do not belong to the same thermodynamic state, and that the native, molten globule and unfolded states are sufficient for interpreting the guanidine hydrochloride-induced unfolding behavior of alpha lactalbumin. PMID- 10707028 TI - Monte Carlo-minimized energy profile of estradiol in the ligand-binding tunnel of 17 beta-hydroxysteroid dehydrogenase: atomic mechanisms of steroid recognition. AB - 17 beta-Estradiol (E2) is a potent stimulator of certain forms of breast cancer. The final step of E2 biosynthesis is catalyzed by the estrogenic 17 beta hydroxysteroid dehydrogenase (17 beta-HSD1), which is an important target for anti-cancer drugs. X-ray crystallography indicated that the binding site for the steroids has a tunnel-like shape. We have used a Monte Carlo-Minimization (MCM) protocol to explore possibilities of interactions of E2 with the binding site tunnel of 17 beta-HSD1. The enzyme was represented by flexible residues having at least one atom within 6 A from either E2 or NADP (as seen in a crystal ternary complex) and by rigid residues having at least one atom within 10 A from E2 or NADP. Special constraints were used to pull the substrate 10 A along the tunnel with 1 A step; the complex was MCM-optimized at each position of the steroid. The optimal binding mode of E2 in 17 beta-HSD agrees with the crystallographic data; however, wide and flat minima of the MCM profile suggest alternative modes of the steroid binding. The advance of the steroid along the tunnel is accompanied by essential conformational rearrangements of the enzyme side chains, noticeable rotation of the substrate along its longitudinal axis, and certain conformational deformations of the substrate. The contributions of the enzyme residues and of the steroid atoms to the intermolecular energy were estimated. PMID- 10707029 TI - Protein structure alignment using a genetic algorithm. AB - We have developed a novel, fully automatic method for aligning the three dimensional structures of two proteins. The basic approach is to first align the proteins' secondary structure elements and then extend the alignment to include any equivalent residues found in loops or turns. The initial secondary structure element alignment is determined by a genetic algorithm. After refinement of the secondary structure element alignment, the protein backbones are superposed and a search is performed to identify any additional equivalent residues in a convergent process. Alignments are evaluated using intramolecular distance matrices. Alignments can be performed with or without sequential connectivity constraints. We have applied the method to proteins from several well-studied families: globins, immunoglobulins, serine proteases, dihydrofolate reductases, and DNA methyltransferases. Agreement with manually curated alignments is excellent. A web-based server and additional supporting information are available at http://engpub1.bu.edu/-josephs. PMID- 10707030 TI - Structural and functional differences of two toxins from the scorpion Pandinus imperator. AB - The Pandinotoxins, PiTX-K alpha and PiTX-K beta, are members of the Charybdotoxin family of scorpion toxins that can be used to characterize K+ channels. PiTX-K alpha differs from PiTX-K beta, another peptide from Pandinus imperator, by one residue (P10E). When the two toxins are compared in a physiological assay, the affinity of PiTX-K beta for voltage-gated, rapidly inactivating K+ channels in dorsal root ganglia (DRG) neurons is 800-fold lower than that of PiTX-K alpha (K alpha-IC50 = 8.0 nM versus K beta-IC50 = 6,500 nM). To understand this difference, the three-dimensional structure of PiTX-K beta was determined by nuclear magnetic resonance (NMR) spectroscopy and compared to that of PiTX-K alpha. This comparison shows that structural differences between the two toxins occur at a residue that is critical for blocking K+ channels (K27) as well as at the site of the natural mutation (P10E). In PiTX-K beta, the negatively charged carboxylate oxygen of E10 can approach the positive charge of K27 and presumably reduces the net positive charge in this region of the toxin. This is likely the reason why PiTX-K beta binds K+ channels from DRG neurons with a much lower affinity than does PiTX-K alpha. PMID- 10707031 TI - In this issue PMID- 10707032 TI - Treatment with metformin of non-diabetic men with hypertension, hypertriglyceridaemia and central fat distribution: the BIGPRO 1.2 trial. AB - BACKGROUND: In the BIGPRO 1 trial, one year of treatment with metformin in non diabetic obese subjects with a central fat distribution had no significant effect on fasting plasma triglyceride concentration or on blood pressure despite a decrease in weight, fasting plasma insulin and glucose concentrations. To re evaluate the effect of metformin on fasting triglyceride concentration and on blood pressure, the BIGPRO 1.2 trial included non-diabetic men (n=168) with a fasting plasma triglyceride concentration > or =1.7 and < or =6.5 mmol/l, high blood pressure (systolic > or =140 and < or =180 and/or diastolic > or =90 and < or =105 mmHg, or treatment for hypertension) and a waist-to-hip ratio > or =0.95. METHODS: A randomised double-blind trial comparing metformin treatment (850 mg bid) with placebo. RESULTS: Metformin had no significant effect either on blood pressure or plasma triglyceride concentration. In comparison with the placebo group, fasting plasma insulin (p<0.04), total cholesterol (p<0.05) and Apo B (p<0.008) concentrations decreased more in the metformin group in the BIGPRO 1. 2 trial, confirming most of the previous results of the BIGPRO 1 trial. Tissue plasminogen activator antigen concentration decreased significantly (p<0.01) only in the metformin group, but this was not significantly different from the placebo group (p<0.12); further, there were no significant differences in the change in plasminogen activator inhibitor 1. CONCLUSIONS: The consistency of the two BIGPRO trials supports the conclusion that metformin affects several cardiovascular risk factors favourably in non-diabetic subjects with a central fat distribution. PMID- 10707033 TI - Autoantibody negative new onset type 1 diabetic patients lacking high risk HLA alleles in a caucasian population: are these type 1b diabetes cases? AB - BACKGROUND: In Caucasians, a small number of Type 1 diabetic patients do not show evidence of humoral islet autoimmunity at disease onset, at least with common screening procedures. In African- and Hispanic-American diabetic children at time of diagnosis, many show no evidence of autoimmunity but have an atypical clinical form of the disease. According to the recent American Diabetes Association classification, this subgroup of autoantibody negative patients is referred to as Type 1b diabetic subjects. In the present study, a homogeneous Caucasian Type 1 diabetic clinic-based cohort has been evaluated at diagnosis using a large panel of diabetes-related antibodies and then characterized for various genetic features in order to identify newly diagnosed Type 1 diabetics who are potentially autoantibody negative, i.e. possibly referrable to as idiopathic Type 1b diabetes. METHODS: Newly diagnosed Type 1 diabetic patients of Italian origin (n=141, mean age 12.0+/-7.6 years) were tested for anti-islet cell, anti-insulin, anti-65 kDa isoform of glutamic acid decarboxylase and anti-amino acid residues 256-979 of the tyrosine-phosphatase IA-2 molecule autoantibodies (Step 1). Only those patients found to be autoantibody negative were tested for anti-disialo ganglioside GD3, anti-thyroid peroxidase, anti-thyroglobulin, anti-21-OH hydroxylase, anti-gastric parietal cell and anti-transglutaminase antibodies (Step 2). Sera negative for the presence of these six autoantibodies as well were characterized in terms of HLA DRB1, DQB1 and CTLA-4. RESULTS: Six out of 141 subjects (3.5%) were autoantibody negative in the first step of the study and five out of six in the second. These five autoantibody negative patients underwent genetic analysis. Three of them had at least one Type 1 diabetes related high risk HLA haplotype (3/141, 2.1%) while the remaining two cases showed neutral (DR5-DQB1*0301/DR5-DQB1*0301) or strongly protective (DR2 DQB1*0602/DR2-DQB1*0602) HLA genotypes, respectively (2/141, 1. 4%). CONCLUSIONS: Clinically defined Type 1 diabetic patients with no sign of autoimmunity do exist in a Caucasian population. These patients (2 out of 141) that cannot be classified as Type 1a diabetic patients lack clinical characteristics of Type 1b diabetes and have to be reconsidered for a more appropriate ADA classification. These data suggest the need of further large population-based studies to understand if Type 1b diabetes really occurs in a Caucasian population. The patient with a strongly protective HLA genotype is particularly interesting considering that among Caucasians only a few sporadic cases with Type 1 diabetes and DQB1*0602, have been reported, none of whom was homozygous at DQB1 locus. PMID- 10707034 TI - Plasma levels of lipophilic antioxidants in very old patients with type 2 diabetes. AB - BACKGROUND: Experimental research indicates that oxidative stress is implicated in aging and in the pathogenesis of diabetes and its complications. This evidence is limited in elderly patients with non-insulin dependent diabetes, in which age- and disease-related production of reactive oxygen species might exert synergistic damaging effects on tissues and organs. METHODS: Plasma levels of lipid-soluble compounds with antioxidant properties including vitamin A, vitamin E and carotenoids (lutein, zeaxanthin, beta-cryptoxanthin, lycopene, alpha- and beta carotene) were measured by HPLC in 72 elderly patients with non-insulin dependent diabetes (75.7+/-0.8 years, 40 F, 32 M) and in 75 age-matched controls (77.2+/ 1.2 years, 48 F, 27 M). RESULTS: All compounds measured were significantly lower in plasma from diabetic patients as compared to controls (p<0.0001). Plasma levels of vitamins A and E and of carotenoids did not significantly correlate with dietary intake and lipid profile in both groups. In patients, significant inverse correlations were found between age and levels of vitamin E, beta cryptoxanthin, lycopene and beta-carotene. CONCLUSIONS: We conclude that patients of very old age with Type 2 diabetes show a poor plasma status of vitamins A and E and carotenoids, which negatively correlates with age. Further studies are needed to explore the possible therapeutic role of lipid-soluble vitamin supplements in elderly diabetic subjects. PMID- 10707035 TI - Hormone replacement therapy and glucose tolerance in EPIC-Norfolk: a population based study. AB - BACKGROUND: Hormone replacement therapy (HRT) can affect glucose homeostasis in postmenopausal women but it is unclear whether long-term use is associated with changes in glucose tolerance. The objective was to examine the relationship of glycated haemoglobin (HbA(1C)) concentration with HRT use in non-diabetic postmenopausal women. METHODS: A cross-sectional analysis of baseline data on 2753 postmenopausal women, aged 45-74 years who were recruited to the EPIC Norfolk study between 1995 and 1998 was performed. Women completed a health and lifestyle questionnaire from which information on HRT use was obtained and gave blood for HbA(1C) assay. RESULTS: Of the women, 23% were current HRT users and 14% were former users. Mean HbA(1C) was significantly lower in current users compared to former and never users. This difference was independent of age, body mass index (BMI), waist-to-hip ratio (WHR), family history of diabetes, educational status, employment status, smoking history, history of alcohol consumption, parity, known illness and hysterectomy status. Compared to never users of HRT, the unadjusted odds ratio (OR) for being in the highest quintile of HbA(1C) distribution as opposed to the lowest was 0.28 (95% CI 0.20-0.39) for current users and 0.41 (0.32-0.53) for ever users. After adjustment for confounders, the OR were 0.52 (0.34-0.79) and 0.72 (0. 51-1.03) for current and ever users, respectively. CONCLUSION: Women currently using HRT have lower HbA(1C) levels not explained by known confounders though we cannot completely exclude a healthy user effect. Nevertheless, this population study suggest that current use of HRT was not associated with impairment of glucose tolerance in postmenopausal women. PMID- 10707037 TI - Pyridine nucleotides in glucose metabolism and diabetes: a review. AB - Nicotinamide adenine dinucleotide (NAD) and its derivatives NADH, NADP and NADPH have regulatory functions in the generation of triose phosphates and pyruvate from glucose. In many studies of the influence of the diabetic state on relationships between pyridine nucleotide and glucose metabolism, the focus has been on the sorbitol pathway. Less attention has been paid to other aspects of the role of pyridine nucleotides in pyruvate formation from glucose, in particular the effects of the NAD precursors nicotinamide and nicotinic acid on glucose metabolism. This paper reviews current knowledge of the involvement of pyridine nucleotides and their precursors in glucose catabolism in the normal and diabetic state. Reference is also made to the following three current hypotheses for mechanisms underlying diabetic microangiopathy: 1. Chronic glucose overutilization, caused by hyperglycemia, in tissues which lack insulin receptors and therefore are freely permeable to glucose. 2. Enhancement of sorbitol pathway activity with an ensuing decrease in the ratio of NAD/NADH. 3. Enhanced utilization of both glucose and pyridine nucleotides in formation of triose phosphates and pyruvate. Therapy with NAD precursors like nicotinamide might have corrective effects on these proposed biochemical aberrations, thereby retarding progression of microangiopathy. PMID- 10707036 TI - High insulin levels do not influence PC-1 gene expression and protein content in human muscle tissue and hepatoma cells. AB - BACKGROUND: To verify whether insulin levels influence PC-1 tissue content, we studied PC-1 gene expression and protein content in skeletal muscle of patients with insulinoma, a model of primary hyperinsulinemia. Data were compared with those obtained in matched insulin sensitive or resistant healthy subjects. In addition, the effect of high insulin concentration on PC-1 protein content was studied in HepG2 cells. METHODS: The following measurements were performed: insulin sensitivity by euglycemic clamp; PC-1 protein content and insulin receptor autophosphorylation by specific ELISAs; PC-1 gene expression by competitive polymerase chain reaction (PCR); phosphatidyl-inositol-3 kinase by immunoprecipitation and thin layer chromatography; glycogen synthesis by (14)C glucose incorporation. RESULTS: Muscle PC-1 content was similar in the insulinoma patients and in insulin sensitive controls but higher (p<0.01) in insulin resistant controls (21.9+/-4.6 ng/mg protein, 23.8+/-3.9, 48.0+/-8.7, respectively). PC-1 protein content was inversely correlated with insulin sensitivity (r=-0.5, p<0.015) but with neither plasma insulin nor glucose levels. PC-1 protein content was correlated with PC-1 gene expression (r=0.53, p<0.05, n=14). Exposure to high insulin (100 nmol/l for 16 h) caused a significant (p<0.05-0.01) impairment of insulin receptor autophosphorylation, phosphatidyl inositol-3 kinase activity and glycogen synthesis, but not of PC-1 protein content (114+/-3 vs 102+/-14 ng/mg protein) in HepG2 cells. CONCLUSION: These findings suggest that chronic high insulin levels do not influence PC-1 expression. PMID- 10707038 TI - An overview of Argentine contributions to diabetes research in the decade of the 1990s. AB - Argentina has a longstanding tradition of diabetes research, beginning with the seminal work of Prof. Bernardo A. Houssay, who was awarded the first Nobel Prize in Medical Sciences for his studies on the relationship between diabetes and pituitary function. Prof. Luis F. Leloir, who was also awarded the Nobel Prize for his work in carbohydrate metabolism, also inspired younger generations of biologists to work in the field of diabetes research. The aim of this paper is to provide a review of the contributions of Argentine researchers during the 1990s. This manuscript includes only reports of Argentine researchers working on diabetes in local laboratories and quoted in Medline. Thus, important contributions not reported in journals included in Medline or produced by Argentine researchers working abroad may have been omitted. The material consists of a brief description of clinical research (epidemiology and costs, metabolic control, associated risk factors, immunological aspects, and other clinical studies) and basic research (animal model with spontaneous diabetes, islet morphology and function in normal and pathological conditions, insulin action, metabolic disorders related to diabetes, and some miscellaneous effects related to drug-induced diabetes). Altogether, a broad idea of the continuous contribution of our national research to the international field of diabetes is provided, as well as a list of Argentine researchers and research centers devoted to the study of diabetes. PMID- 10707039 TI - Current awareness AB - In order to keep subscribers up-to-date with the latest developments in their field, John Wiley & Sons are providing a current awareness service in each issue of the journal. The bibliography contains newly published material in the field of diabetes/metabolism. Each bibliography is divided into 17 sections: 1 Books, Reviews & Symposia; 2 General; 3 Genetics; 4 Epidemiology; 5 Immunology; 6 Prediction; 7 Prevention; 8 Intervention: a) General; b) Pharmacology; 9 Pathology: a) General; b) Cardiovascular; c) Neurological; d) Renal; 10 Endocrinology & Metabolism; 11 Nutrition; 12 Animal Studies; 13 Techniques. Within each section, articles are listed in alphabetical order with respect to author (9 Weeks journals - Search completed at 27th Oct. 1999) PMID- 10707041 TI - BioArtificial Muscle implants as drug delivery systems. PMID- 10707040 TI - Medicines for the 21st century? PMID- 10707042 TI - New liposomal formulation of vincristine for non-Hodgkin's lymphoma. PMID- 10707043 TI - The importance of drug delivery systems in tissue engineering. AB - Tissue engineering is designed to regenerate natural tissues or to create biological substitutes for defective or lost tissues and organs through the use of cells. In addition to cells and their scaffolds, growth factors are required to promote tissue regeneration. Indeed, growth factor-induced vascularization is effective in supplying the oxygen and nutrients necessary for the survival of transplanted cells in organ substitution. However, growth factors have poor in vivo stability and so the biological effects are often unpredictable unless the delivery system is contrived. This review provides several examples to emphasize the importance of drug delivery systems in tissue engineering. PMID- 10707044 TI - Advances in the use of monoclonal antibodies in cancer radiotherapy. AB - The use of monoclonal antibodies (MAbs) as radiation carriers in argeted radiotherapy of cancers has produced striking clinical responses in hematologic diseases, such as non-Hodgkin's lymphoma. Novel strategies are currently being examined in an effort to improve efficacy in solid tumor therapies. Two of these strategies involve minimizing the systemic toxicity of a circulating radionuclide via 'pretargeting', and the sensitization of tumors to radiation by combination therapy with radiosensitizing drugs. Advances made in radiolabeling chemistries and in the use of alpha-particle emitters can also improve utility. Clinical evidence suggests that radioimmunotherapy may be best applied in minimal-disease and adjuvant settings in combination with other cancer therapy modalities. PMID- 10707045 TI - Recombinant in vitro tools to predict drug metabolism and safety. AB - Drug metabolism determines several pharmacological and toxicological properties of pharmaceuticals and is catalysed by drug metabolizing enzymes. Prediction of drug metabolism in humans based on animal experiments is complicated by species differences in the catalytic properties of these enzymes. This review describes and evaluates the use of recombinant models that contain human drug metabolizing enzymes to facilitate the prediction of pharmacokinetic properties of candidate drugs in humans. PMID- 10707046 TI - Monitor: progress and profiles. AB - Monitor provides an insight into the latest developments in pharmaceutical science and technology through brief synopses of recent presentations, publications and patents, and expert commentaries on the latest technologies. There are two sections: Progress summarizes the latest developments in pharmaceutical process technology, formulation, analytical technology, sterilization, controlled drug delivery systems and regulatory issues; Profiles offers expert commentary on emerging technologies, novel processes and strategic, organizational and logistic issues underlying pharmaceutical R&D. PMID- 10707047 TI - Glucocorticoid-induced osteoporosis. AB - Endogenous cortisol excess and glucocorticoid (GC) treatment have a profound effect on bone metabolism, acting at many sites. The mechanism of GC action on bone turnover is complex and has not been elucidated completely. GCs increase bone resorption, inhibit bone formation and have an indirect action on bone by decreasing intestinal Ca2+ absorption, modifying vitamin D metabolism, and sustaining a marked hypercalciuria, with variable changes in plasma PTH levels; finally, GCs inhibit the gonadotropic and somatotropic axis. GC-induced osteoporosis is preventable, treatable and potentially reversible. The prevention and treatment of GC-induced osteoporosis include some general measures (as well as the use of the minimal effective dose of GC), Ca2+ and vitamin D supplementation and treatment with bone anabolic and antiresorptive agents. Recent trials suggest that bisphosphonates are an effective therapeutic tool in the treatment of GC-induced bone damage. Recent data on GC receptor-selective modulators indicate that these new molecules might induce only minimal bone loss while maintaining the typical anti-inflammatory properties of GC. Another new line of study for the prevention of GC-induced osteoporosis is the characterization of the individual's susceptibility to GC-induced bone damage. PMID- 10707048 TI - Do glucocorticoids cause spinal epidural lipomatosis? When endocrinology and spinal surgery meet. AB - Here, we report pathogenetic aspects of spinal epidural lipomatosis (SEL) based on a literature review. SEL is a rare entity but can cause significant morbidity. Its symptoms can be identical to those of more common disorders such as vertebral and disc disease, and cord lesions (for example, transverse myelitis, multiple sclerosis and syringomyelia). Therefore, it often goes undiagnosed. In addition, SEL occurs in patients on glucocorticoid therapy, which can lead to myopathy, thereby mimicking the motor symptoms of SEL. Glucocorticoids seem to play a major role in the development of SEL, although idiopathic SEL has also been reported. The latter occurs almost exclusively in obese individuals who may have concurrent hypercortisolism. Once clinically suspected, SEL is best diagnosed by magnetic resonance imaging (MRI). Treatment of SEL is directed at reducing body weight in patients with idiopathic SEL, and at decreasing glucocorticoid excess in patients with endogenous or exogenous hypercortisolism. In severe cases, decompressive laminectomy might become necessary to alleviate the neurological symptoms caused by spinal cord compression. PMID- 10707049 TI - Activation of MAPK cascades by G-protein-coupled receptors: the case of gonadotropin-releasing hormone receptor. AB - G-protein-coupled receptors (GPCRs) are a large group of integral membrane receptors that transmit signals from a diverse array of external stimuli, including neurotransmitters, hormones, phospholipids, photons, odorants and taste ligands. In response to ligand binding, the GPCRs initiate diverse downstream signaling pathways through four groups of G proteins and other interacting proteins. Key components in GPCR-induced intracellular signaling are four groups of mitogen-activated protein kinase (MAPK) cascades: extracellular signal-related kinase (ERK), Jun N-terminal kinase (JNK), p38MAPK and big MAPK (BMK). The hallmark of MAPK signaling is the stimulation-dependent nuclear translocation of the involved kinases, which regulate gene expression and the cytoplasmic acute response to mitogenic, stress-related, apoptotic and survival stimuli. A special type of GPCR is the gonadotropin-releasing hormone (GnRH) receptor, which uses primarily the Gq protein for its downstream signaling. GnRH activates all four MAPK cascades by a PKC-dependent mechanism. Common signaling molecules, including the tyrosine kinase c-SRC and the small GTPases CDC42, RAC and RAS, are implicated in various aspects of the GnRH-MAPK pathways. Thus, the activation of MAPK cascades by GnRH opens a new vista in the understanding of the transcriptional regulation of genes encoding gonadotropins. However, additional studies on cell lines and whole animals are required to understand GnRH signaling in the context of other hormones during the reproductive cycle of mouse and human. PMID- 10707050 TI - Molecular study of the sodium-iodide symporter (NIS): a new field in thyroidology. AB - The active transport of iodide into the thyroid is mediated by the Na(+)-I- symporter (NIS), an intrinsic membrane protein. NIS plays key roles in thyroid pathophysiology as the route by which I- reaches the gland for thyroid hormone biosynthesis, and as a means for diagnostic scintigraphic imaging and for radioiodide therapy in thyroid cancer. The molecular characterization of NIS started with the isolation in 1996 of a cDNA encoding rat NIS, and has subsequently led to a virtually new field in thyroidology. The research reviewed in this article clearly has far-reaching implications in the areas of structure/function of transport proteins, thyroid pathophysiology, hormone action mechanisms, cell differentiation and cancer. PMID- 10707052 TI - Plasticity of the transmembrane beta-barrel. PMID- 10707051 TI - Genetic defects in postsqualene cholesterol biosynthesis. AB - In humans and mice, four different genetic defects in the nine biosynthetic steps from lanosterol to cholesterol have been identified. They impair the activity of a putative C3-sterol dehydrogenase (Nshdl, X-linked dominant bare patches/striated mutation in mice), the sterol delta 8-delta 7 isomerase/EBP (Ebp, X-linked dominant tattered mutation in mice; chondrodysplasia punctata (CDPX2) in humans), the delta 24-sterol reductase (autosomal recessive desmosterolosis) and the delta 7-sterol reductase (DHCR7 gene, autosomal recessive Smith-Lemli-Opitz syndrome in humans). These inborn errors in postsqualene cholesterol metabolism result in dysmorphogenetic syndromes of variable severity. The X-linked dominant mutations result in mosaicism in females, as a result of X-inactivation, and midgestational lethality in males. The mechanisms by which the depletion of cholesterol or the accumulation of intermediates impair morphogenetic programs are unclear. So far, no cellular processes that require an intact cholesterol biosynthetic pathway have been identified, although the morphogenetic hedgehog-patched signaling cascade is a candidate. PMID- 10707053 TI - 'Deja vu all over again': the similar structures of bacteriophage PRD1 and adenovirus. PMID- 10707055 TI - Analysis of a malaria sporozoite protein family required for gliding motility and cell invasion: response PMID- 10707054 TI - Analysis of a malaria sporozoite protein family required for gliding motility and cell invasion. PMID- 10707056 TI - Identifying Mycobacterium tuberculosis virulence determinants - new technologies for a difficult problem. PMID- 10707057 TI - Identifying mycobacterium tuberculosis virulence determinants - new technologies for a difficult problem: response PMID- 10707058 TI - New signal molecules on the quorum-sensing block. PMID- 10707059 TI - New signal molecules on the quorum-sensing block: response PMID- 10707060 TI - Understanding microbial MIP channels. PMID- 10707061 TI - Photosynthesis in Rhodobacter sphaeroides. PMID- 10707062 TI - Response from Vermeglio and joliot PMID- 10707063 TI - Response from crofts PMID- 10707064 TI - Exploitation of cellular signaling and regulatory pathways by human cytomegalovirus. AB - Human cytomegalovirus is a ubiquitous human pathogen that is the leading viral cause of birth defects. It also causes significant morbidity and mortality in both chemically and virally immunosuppressed individuals. Recent studies have begun to elucidate the interplay between this virus and its host cell on a molecular level. The interactions begin upon contact with the cell membrane, involve multiple processes including cell signaling, cell-cycle control and immune response mechanisms, and culminate in a productive infection. PMID- 10707065 TI - Phages will out: strategies of host cell lysis. AB - Most phages accomplish host lysis using a muralytic enzyme, or endolysin, and a holin, which permeabilizes the membrane at a programmed time and thus controls the length of the vegetative cycle. By contrast, lytic single-stranded RNA and DNA phages accomplish lysis by producing a single lysis protein without muralytic activity. PMID- 10707066 TI - Horizontal gene transfer and the origin of species: lessons from bacteria. AB - In bacteria, horizontal gene transfer (HGT) is widely recognized as the mechanism responsible for the widespread distribution of antibiotic resistance genes, gene clusters encoding biodegradative pathways and pathogenicity determinants. We propose that HGT is also responsible for speciation and sub-speciation in bacteria, and that HGT mechanisms exist in eukaryotes. PMID- 10707067 TI - cAMP signalling in pathogenic fungi: control of dimorphic switching and pathogenicity. AB - Morphological changes in pathogenic fungi often underlie the development of virulence and infection by these organisms. Our knowledge of the components of the cell signalling pathways controlling morphological switching has, to a large extent, come from studies of pseudohyphal growth of the model organism Saccharomyces cerevisiae, in which control is exerted via changes in the intracellular cAMP and mitogen-activated protein kinase cascades. There is evidence that pathogenic fungi also utilize these pathways to control dimorphic switching between saprobic and pathogenic forms and, as such, the elements of these pathways have potential as drug targets. PMID- 10707068 TI - Caenorhabditis elegans for the study of host-pathogen interactions. AB - The nematode worm Caenorhabditis elegans, for which the complete genome sequence is available, has several other advantages as an experimental system, and has already been widely used as a model for the study of vertebrate biology. Recent investigations have revealed that C. elegans could also be an extremely useful model system in the study of bacterial pathogenesis and have reinforced the notion that common virulence and host defence mechanisms exist. PMID- 10707069 TI - Moving on up: auxin-induced K+ channel expression regulates gravitropism. PMID- 10707070 TI - . . . response: living with gravity. PMID- 10707072 TI - Expanding the club: engineering plants to talk to bacteria. PMID- 10707071 TI - Do phytochromes interact with diverse partners? PMID- 10707073 TI - Grafting to save the forests. PMID- 10707074 TI - All fatty acids are not equal: discrimination in plant membrane lipids. AB - Plant membrane lipids are primarily composed of 16-carbon and 18-carbon fatty acids containing up to three double bonds. By contrast, the seed oils of many plant species contain fatty acids with significantly different structures. These unusual fatty acids sometimes accumulate to >90% of the total fatty acid content in the seed triacylglycerols, but are generally excluded from the membrane lipids of the plant, including those of the seed. The reasons for their exclusion and the mechanisms by which this is achieved are not completely understood. Here we discuss recent research that has given new insights into how plants prevent the accumulation of unusual fatty acids in membrane lipids, and how strict this censorship of membrane composition is. We also describe a transgenic experiment that resulted in an excessive buildup of unusual fatty acids in cellular membranes, and clearly illustrated that the control of membrane lipid composition is essential for normal plant growth and development. PMID- 10707075 TI - Gibberellin and abscisic acid signalling in aleurone. AB - The plant hormones gibberellin and abscisic acid regulate gene expression, secretion and cell death in aleurone. The emerging picture is of gibberellin perception at the plasma membrane whereas abscisic acid acts at both the plasma membrane and in the cytoplasm - although gibberellin and abscisic acid receptors have yet to be identified. A range of downstream-signalling components and events has been implicated in gibberellin and abscisic acid signalling in aleurone. These include the Galpha subunit of a heterotrimeric G protein, a transient elevation in cGMP, Ca2+-dependent and Ca2+-independent events in the cytoplasm, reversible protein phosphory-lation, and several promoter cis-elements and transcription factors, including GAMYB. In parallel, molecular genetic studies on mutants of Arabidopsis that show defects in responses to these hormones have identified components of gibberellin and abscisic acid signalling. These two approaches are yielding results that raise the possibility that specific gibberellin and abscisic acid signalling components perform similar functions in aleurone and other tissues. PMID- 10707076 TI - Formation and maintenance of the shoot apical meristem. AB - Development in higher plants is characterized by the reiterative formation of lateral organs from the flanks of shoot apical meristems. Because organs are produced continuously throughout the life cycle, the shoot apical meristem must maintain a pluripotent stem cell population. These two tasks are accomplished within separate functional domains of the apical meristem. These functional domains develop gradually during embryogenesis. Subsequently, communication among cells within the shoot apical meristem and between the shoot apical meristem and the incipient lateral organs is needed to maintain the functional domains within the shoot apical meristem. PMID- 10707077 TI - Cytochromes P450 for engineering herbicide tolerance. AB - In recent years, genome sequencing has revealed that cytochromes P450 (P450s) constitute the largest family of enzymatic proteins in higher plants. P450s are mono-oxygenases that insert one atom of oxygen into inert hydrophobic molecules to make them more reactive and hydrosoluble. Besides their physiological functions in the biosynthesis of hormones, lipids and secondary metabolites, P450s help plants to cope with harmful exogenous chemicals including pesticides and industrial pollutants, making them less phytotoxic. The recovery of an increasing number of plant P450 genes in recombinant form has enabled their use in experimentation, which has revealed their extraordinary potential for engineering herbicide tolerance, biosafening, bioremediation and green chemistry. PMID- 10707078 TI - Programmed cell death and aerenchyma formation in roots. AB - Lysigenous aerenchyma contributes to the ability of plants to tolerate low-oxygen soil environments, by providing an internal aeration system for the transfer of oxygen from the shoot. However, aerenchyma formation requires the death of cells in the root cortex. In maize, hypoxia stimulates ethylene production, which in turn activates a signal transduction pathway involving phosphoinositides and Ca2+. Death occurs in a predictable pattern, is regulated by a hormone (ethylene) and provides an example of programmed cell death. PMID- 10707079 TI - The role of root border cells in plant defense. AB - The survival of a plant depends upon the capacity of root tips to sense and move towards water and other nutrients in the soil. Perhaps because of the root tip's vital role in plant health, it is ensheathed by large populations of detached somatic cells - root 'border' cells - which have the ability to engineer the chemical and physical properties of the external environment. Of particular significance, is the production by border cells of specific chemicals that can dramatically alter the behavior of populations of soilborne microflora. Molecular approaches are being used to identify and manipulate the expression of plant genes that control the production and the specialized properties of border cells in transgenic plants. Such plants can be used to test the hypothesis that these unusual cells act as a phalanx of biological 'goalies', which neutralize dangers to newly generated root tissue as the root tip makes its way through soil. PMID- 10707082 TI - CED-2/CrkII and CED-10/Rac control phagocytosis and cell migration in Caenorhabditis elegans. AB - Engulfment of apoptotic cells in Caenorhabditis elegans is controlled by two partially redundant pathways. Mutations in genes in one of these pathways, defined by the genes ced-2, ced-5 and ced-10, result in defects both in the engulfment of dying cells and in the migrations of the two distal tip cells of the developing gonad. Here we find that ced-2 and ced-10 encode proteins similar to the human adaptor protein CrkII and the human GTPase Rac, respectively. Together with the previous observation that ced-5 encodes a protein similar to human DOCK180, our findings define a signalling pathway that controls phagocytosis and cell migration. We provide evidence that CED-2 and CED-10 function in engulfing rather than dying cells to control the phagocytosis of cell corpses, that CED-2 and CED-5 physically interact, and that ced-10 probably functions downstream of ced-2 and ced-5. We propose that CED-2/CrkII and CED 5/DOCK180 function to activate CED-10/Rac in a GTPase signalling pathway that controls the polarized extension of cell surfaces. PMID- 10707083 TI - Rapid movement of axonal neurofilaments interrupted by prolonged pauses. AB - Axonal cytoskeletal and cytosolic proteins are synthesized in the neuronal cell body and transported along axons by slow axonal transport, but attempts to observe this movement directly in living cells have yielded conflicting results. Here we report the direct observation of the axonal transport of neurofilament protein tagged with green fluorescent protein in cultured nerve cells. Live-cell imaging of naturally occurring gaps in the axonal neurofilament array reveals rapid, intermittent and highly asynchronous movement of fluorescent neurofilaments. The movement is bidirectional, but predominantly anterograde. Our data indicate that the slow rate of slow axonal transport may be the result of rapid movements interrupted by prolonged pauses. PMID- 10707084 TI - Ras is involved in nerve-activity-dependent regulation of muscle genes. AB - Gene expression in skeletal muscle is regulated by the firing pattern of motor neurons, but the signalling systems involved in excitation-transcription coupling are unknown. Here, using in vivo transfection in regenerating muscle, we show that constitutively active Ras and a Ras mutant that selectively activates the MAPK(ERK) pathway are able to mimic the effects of slow motor neurons on expression of myosin genes. Conversely, the effect of slow motor neurons is inhibited by a dominant-negative Ras mutant. MAPK(ERK) activity is increased by innervation and by low-frequency electrical stimulation. These results indicate that Ras-MAPK signalling is involved in promoting nerve-activity-dependent differentiation of slow muscle fibres in vivo. PMID- 10707085 TI - p16INK4A and p19ARF act in overlapping pathways in cellular immortalization. AB - The INK4A locus encodes two independent but overlapping genes, p16INK4A and p19ARF, and is frequently inactivated in human cancers. The unusual structure of this locus has lead to ambiguity regarding the biological role of each gene. Here we express, in primary mouse embryonic fibroblasts (MEFs), antisense RNA constructs directed specifically towards either p16INK4A or p19 ARF. Such constructs induce extended lifespan in primary MEFs; this lifespan extension is reversed upon subsequent elimination of the p16INK4A or p19ARF antisense constructs. In immortal derivatives of cell lines expressing antisense p16INK4A or p19ARF RNA, growth arrest induced by recovery of p16INK4A expression is bypassed by compromising the function of the retinoblastoma protein (Rb), whereas growth arrest induced by re-expression of p19ARF is overcome only by simultaneous inactivation of both the Rb and the p53 pathways. Thus, the physically overlapping p16INK4A and p19ARF genes act in partly overlapping pathways. PMID- 10707086 TI - The coordinate release of cytochrome c during apoptosis is rapid, complete and kinetically invariant. AB - Release of cytochrome c from mitochondria triggers activation of caspase proteases and death of a cell by apoptosis. However, the mechanism and kinetics of cytochrome c release remain unknown. Here we study this event by using green fluorescent protein (GFP)-tagged cytochrome c, and find that the release of cytochrome-c-GFP always precedes exposure of phosphatidylserine and the loss of plasma-membrane integrity - characteristics of apoptotic cells. Once initiated, the release of cytochrome- c-GFP continues until all of the protein is released from all mitochondria in individual cells, within about 5 minutes, regardless of the type or strength of stimulus or the time elapsed since the stimulus was applied. Temperatures ranging from 24 degrees C to 37 degrees C do not change the duration of release, and nor does the addition of caspase inhibitors. Further, we find that the electron-transport chain can maintain the mitochondrial transmembrane potential even after cytochrome c has been released. PMID- 10707087 TI - HIV-1 Nef protein binds to the cellular protein PACS-1 to downregulate class I major histocompatibility complexes. AB - Major-histocompatibility-complex (MHC) proteins are used to display, on the surface of a cell, peptides derived from foreign material - such as a virus - that is infecting that cell. Cytotoxic T lymphocytes then recognize and kill the infected cell. The HIV-1 Nef protein downregulates the cell-surface expression of class I MHC proteins, and probably thereby promotes immune evasion by HIV-1. In the presence of Nef, class I MHC molecules are relocalized from the cell surface to the trans-Golgi network (TGN) through as-yet-unknown mechanisms. Here we show that Nef-induced downregulation of MHC-I expression and MHC-I targeting to the TGN require the binding of Nef to PACS-1, a molecule that controls the TGN localization of the cellular protein furin. This interaction is dependent on Nef's cluster of acidic amino acids. A chimaeric integral membrane protein containing Nef as its cytoplasmic domain localizes to the TGN after internalization, in an acidic-cluster- and PACS-1-dependent manner. These results support a model in which Nef relocalizes MHC-I by acting as a connector between MHC-I's cytoplasmic tail and the PACS-1-dependent protein-sorting pathway. PMID- 10707088 TI - Single-molecule imaging of EGFR signalling on the surface of living cells. AB - The early events in signal transduction from the epidermal growth factor (EGF) receptor (EGFR) are dimerization and autophosphorylation of the receptor, induced by binding of EGF. Here we observe these events in living cells by visualizing single molecules of fluorescent-dye-labelled EGF in the plasma membrane of A431 carcinoma cells. Single-molecule tracking reveals that the predominant mechanism of dimerization involves the formation of a cell-surface complex of one EGF molecule and an EGFR dimer, followed by the direct arrest of a second EGF molecule, indicating that the EGFR dimers were probably preformed before the binding of the second EGF molecule. Single-molecule fluorescence-resonance energy transfer shows that EGF-EGFR complexes indeed form dimers at the molecular level. Use of a monoclonal antibody specific to the phosphorylated (activated) EGFR reveals that the EGFR becomes phosphorylated after dimerization. PMID- 10707089 TI - Calmodulin kinase determines calcium-dependent facilitation of L-type calcium channels. AB - A dynamic positive feedback mechanism, known as 'facilitation', augments L-type calcium-ion currents (ICa) in response to increased intracellular Ca2+ concentrations. The Ca2+-binding protein calmodulin (CaM) has been implicated in facilitation, but the single-channel signature and the signalling events underlying Ca2+/CaM-dependent facilitation are unknown. Here we show that the Ca2+/CaM-dependent protein kinase II (CaMK) is necessary and possibly sufficient for ICa facilitation. CaMK induces a channel-gating mode that is characterized by frequent, long openings of L-type Ca2+ channels. We conclude that CaMK-mediated phosphorylation is an essential signalling event in triggering Ca2+/CaM-dependent ICa facilitation. PMID- 10707090 TI - Identification of a cryptic nucleolar-localization signal in MDM2. PMID- 10707091 TI - Mitotic stability of an episomal vector containing a human scaffold/matrix attached region is provided by association with nuclear matrix. PMID- 10707092 TI - The importance of technological advances. PMID- 10707093 TI - The ins and outs of cell-polarity decisions. AB - The guanine-nucleotide-exchange factor Cdc24 is a critical regulator of cell polarity. Far1 is a key player in Cdc24 regulation, controlling Cdc24 activity in budding yeast by regulating its subcellular localization in response to two very different signals - one external and one internal. PMID- 10707094 TI - Wingless and naked. PMID- 10707095 TI - Cytochrome c release from mitochondria: all or nothing. AB - During the process of apoptosis, cytochrome c is released from mitochondria into the cytosol where it helps to activate the caspases, a family of killer proteases. The release of cytochrome c is a rapid, complete and kinetically invariant event. PMID- 10707096 TI - Neurofilaments run sprints not marathons. AB - In neurons, cytoskeletal proteins are transported from where they are made - the cell body - along the axons, but it has long been disputed whether they are transported as subunits or polymers. A new analysis of neurofilament movement may help to resolve the controversy. PMID- 10707097 TI - Zipping up adhesion. PMID- 10707098 TI - Naked antigen-presenting molecules on dendritic cells. AB - Antigen-presenting cells work to present peptides derived from exogenous and endogenous antigens to circulating T cells, sparking off an immune response. Dendritic cells are unique amongst antigen-presenting cells, not least for their newly described ability to circumvent the need to internalize exogenous antigens before presenting them. PMID- 10707099 TI - p53: only ARF the story. AB - Two tumour-suppressor proteins - p16INK4A and p19 ARF - encoded by the same genetic locus both have a role in arresting the cell-division cycle. New results have revealed further complexities in the pathways involved. PMID- 10707100 TI - Malaria-treatment policies: when and how should they be changed? AB - There appears to be a large a gap in the literature between primary work on malaria control and policy on the one hand and the interpretation of such work in making real policy decisions on the other. The focus of the present review is policy formulation for treatment of uncomplicated falciparum malaria, rather than prophylaxis in travellers or the treatment of severe disease. The World Health Organization has formulated guidelines addressing the issue of changing from one recommended drug for treating malaria to another, but there does not appear to have been a comprehensive attempt to describe how and when such a decision on drug policy should be made. Issues of drug availability, both to countries and to communities within them, are discussed, as well as the acceptability of drug regimens and compliance with them. It emerges that the cost of treatment has a disproportionate influence on the decision-making process, and that the indirect costs of drug failure are often not considered properly. Brief mention is made of the indicators of overall disease burden. There is some discussion about the usefulness of one recently introduced economic indicator: the disability-adjusted life-year (DALY). Also examined are the difficulties that arise within the context of drug-policy changes, such as a regimen's appropriateness to all target groups, and the strong influence of the private sector on decision-making that affects its own financing. The consensus seems to be that a policy change is urgent when high-level resistance occurs in 40% or more of treated cases, when parasitological response is poor, and when the costs of treatment failures are higher than those of treatments with a newer drug. It also emerges that much remains to be done regarding co-operation between public and private sectors; considering the importance of private-sector provision of health care, this needs to be addressed. PMID- 10707101 TI - Surprisingly little polymorphism in the merozoite-surface-protein-2 (MSP-2) gene of Indian Plasmodium falciparum. AB - The polymorphism in the merozoite-surface-protein-2 (MSP-2) gene of six Indian Plasmodium falciparum isolates was studied by PCR amplification, cloning and sequencing. One of the isolates showed a deletion of 63 bp and all showed point mutations, although some of these mutations were silent. All the isolates also exhibited 5' and 3' conserved regions, with the two 32-mer amino-acid repeats characteristic of the FC27 family, and none belonged to the IC-1/3D7 family. Although the MSP-2 genes of these isolates represent new allelic sequences, they belong to the FC27 family and show remarkably little variation. PMID- 10707102 TI - Response to chloroquine of Plasmodium vivax among South Korean soldiers. AB - The response to standard chloroquine treatment was evaluated, by microscopical examination of blood-smears, among 81 soldiers diagnosed with Plasmodium vivax malaria in South Korea in 1996. The smears were prepared pre-treatment and 3, 14 and 28 days after starting chemotherapy. Parasitaemias were determined after staining the smears with Giemsa's stain. Blood samples from the patients who were not smear-negative by day 3 were carefully checked for parasites, by staining smears with Acridine Orange and by a PCR-based assay. Only two of the patients appeared to be parasitaemic on day 14 and were therefore considered treatment failures. Although both were apparently cured after additional therapy with the same regimen, one had a recurrence 8 months later. Most cases of recent, resurgent malaria in South Korea therefore appear to sensitive to chloroquine. PMID- 10707103 TI - The chemotherapy of rodent malaria. LVII. Drug combinations to impede the selection of drug resistance, Part 1: Which model is appropriate? AB - The principle has finally been accepted that, whenever possible, antimalarial drugs should be deployed in appropriate combinations in endemic areas, in order to minimize the inevitability that monotherapy will, probably sooner than later, select populations of drug-resistant parasites. Which laboratory models can predict the combinations of old or novel compounds that are likely to be of practical value in minimizing this risk? Very few relevant data on the use of Plasmodium falciparum in vitro have been published. Most research has been carried out with one or other strain of chloroquine-sensitive P. berghei or with chloroquine-resistant P. yoelii ssp. NS in mice. The two most widely used procedures to select for resistance are the 'serial technique' (ST), in which drug selection pressure in vivo is gradually increased, and the '2% relapse technique' (2%RT), in which a single, large drug dose is applied at the time of each passage. Both procedures have been used to demonstrate the ability of pairs of drugs (e.g. sulfadoxine with pyrimethamine, cycloguanil with dapsone, pyrimethamine or sulphonamides with chloroquine, mepacrine or mefloquine) or triple combinations (e.g. sulfadoxine-pyrimethamine with chloroquine, mefloquine or pyronaridine) to delay the development of resistance. The relative merits of the ST and 2%RT are discussed and the data obtained by these procedures are compared with the results of deploying drug combinations in man, especially against multidrug-resistant P. falciparum. It is concluded that, even though the rodent malaria models are imperfect, no better alternatives are available at present with which to predict the value of antimalarial combinations for the protection of the individual components. The 2%RT is considered to be the procedure of first choice. PMID- 10707104 TI - Oral treatment of visceral leishmaniasis with miltefosine. AB - In a pilot trial, 28 days of oral treatment with 100-200 mg miltefosine (hexadecylphosphocholine) per day cured 14 of 15 patients with Indian visceral leishmaniasis (VL). To extend the testing of this regimen, 45 additional subjects with VL, of whom 17 had failed previous antimony therapy, were treated with 100 (N = 17), 150 (N = 18) or 200 (N = 10) mg/day. Enrollment at 200 mg/day was stopped after three subjects in this treatment arm developed reversible but serious (grade-3) adverse reactions. The overall clinical and parasitological responses to miltefosine were rapid, with 40 [89%; 95% confidence interval (CI) = 76%-96%] and 44 (98%; CI = 88%-100%) of the patients apparently cured on days 14 and 28, respectively. The one 'treatment failure' recorded on day 28 (and at 6 months) was a subject lost to follow-up. Those apparently cured by day 28 included six patients (one on 100 mg, two on 150 mg and three on 200 mg/day) removed from treatment on days 7-17 because of grade-3 diarrhoea (two cases), vomiting (two cases), diarrhoea and hepatotoxicity (one case) or nephrotoxicity (one case). Transient, mild-moderate vomiting and/or diarrhoea were common during weeks 1-2 and about 25% of the patients also developed primarily mild, self limited increases in concentrations of aspartate aminotransferase and creatinine and/or blood urea nitrogen. At a 6-month follow-up, all 44 patients apparently cured at day 28 were considered complete responders (definitive cures), including the six treated for only 7-17 days. These results indicate that 100 mg miltefosine/day for 28 days is a promising oral-treatment regimen for VL cases, including those with antimony-unresponsive infections. PMID- 10707105 TI - Leishmania major: effect of repeated passages through sandfly vectors or murine hosts. AB - Virulence for BALB/c mice, infectivity for Phlebotomus papatasi, haemagglutination activity and expression of metacyclic lipophosphoglycan (LPG) were studied in four strains of Leishmania major (LV561, FV1, L119 and Neal) and various lines of the LV561 strain. Attenuated line LV561/AV was passaged five times through sandflies or mice and the resulting lines (AVS5 and AVM5, respectively) and two of the earlier sandfly passages (AVS1 and AVS2) were used for further study. The highly virulent line LV561/V served as a control. Virulence for mice was not regained during passaging of LV561/AV in sandflies or mice (none of the mice infected with AVM5, AVS1, AVS2 or AVS5 displayed overt lesions) and the success rate in cultivating parasites, from lymph-node samples from inoculated mice, was not significantly higher for any of these lines than for the original line (LV561/AV). However, AVM5 and AVS5 developed better than LV561/AV in P. papatasi and the intensity and localisation of their infections were similar to those of the virulent control. In smears of the infected guts of P. papatasi, the AVS5 parasites resembled the virulent line (LV561/V) morphologically whereas the AVM5 parasites were similar to the avirulent LV561/AV. Haemagglutination activity increased as a result of passaging, the most pronounced difference being observed in AVM5, which had 60-fold higher activity than LV561/AV. Expression of metacyclic LPG was not increased by passaging. The proportion of forms reacting positively with 3F12 antibodies was high (about 17%) in the virulent LV561/V but low (2%-3%) in the avirulent lines LV561/AV, AVS5 and AVM5. A defect in LPG is not, however, likely to be the only reason for the avirulence observed, as the avirulent lines of LV561 still produced about 10 times as many metacyclic promastigotes as the strain L119, which caused delayed lesions in mice. PMID- 10707106 TI - Use of urine samples from healthy humans, nephritis patients or other animals as an alternative to foetal calf serum in the culture of Leishmania (L.) donovani in vitro. AB - The effect of supplementing in-vitro cultures of Leishmania donovani with urine was investigated. The parasites were isolated from Bangladeshi patients with visceral leishmaniasis. The urine samples used were collected from healthy human donors, patients with nephrotic syndrome, diabetic nephritis (DN) or diabetes mellitus, a dog and a cow. Promastigotes from blood-agar cultures were inoculated into RPMI-1640 basal medium with 10% heat-inactivated foetal calf serum (FCS) and/or 1%-20% urine. The parasites were then counted in a haemocytometer, on days 2, 4, 5, 6, 7, 8, 10, 12 and 14 post-inoculation. From day 4, the numbers of parasites/ml in cultures containing 5% healthy-human urine but no FCS were at least as high as those in cultures containing 10% FCS but no urine (P = 0.191). The wet weights of parasites harvested from mass cultures of the parasites in RPMI-1640 plus 5% healthy-human urine and in RPMI-1640 plus 10% FCS were practically the same. Multiplication of the parasites in the presence of 5% urine from a DN patient was significantly greater (P < 0.01) than that seen with other urine samples at the same concentration or with 10% FCS. The multiplication seen with 8% canine urine was almost the same as with 5% healthy-human urine. Parasites could be maintained in RPMI-1640 plus 5% healthy-human urine for at least 40 days, sub-culturing every 4 days. Urine may be a better and much cheaper stimulant of Leishmania multiplication in vitro than FCS. PMID- 10707107 TI - The possible role of the age of the human host in determining the localization of hydatid cysts. AB - Cystic hydatid disease or cystic echinococcosis (CE) is a cyclozoonotic infection distributed world-wide. The morbidity attributable to the infection depends on the size of the cyst(s) and the organ(s) involved. The cysts are most commonly found in the liver and lungs but certain locations have been reported to be more prevalent in children and/or young adults than in older subjects. In order to identify the relationship, if any, between the age of the patient and the site of involvement, the age and cyst distribution of 92 cases of CE were analysed. Lung, brain, spinal and orbital hydatid cysts were more commonly seen in younger patients whereas other sites were preferentially involved in older patients. The factors that determine the final localization of the cysts are discussed. It is concluded that age somehow alters the host-parasite relationship and thus affects the organ distribution of the cysts. PMID- 10707108 TI - A 2-year prospective study in China provides epidemiological evidence for resistance in humans to re-infection with Schistosoma japonicum. AB - In 1996, 250 people living in the Dongting-Lake region of China were selected for a 2-year study. All had been or were infected with Schistosoma japonicum. All were treated with praziquantel, although eggs of S. japonicum were only detected in stool samples from 75 of the subjects. In 1998, 213 (85.5%) of these subjects, then with a mean (S.D.) age of 40.2 (14.2) years, provided stool samples for final assessment. Forty-nine (23%) of the 213 were found to be re-infected in 1998, with a geometric mean intensity of infection among the infected of 64.5 eggs/g faeces. The rate of re-infection was highest among those aged < 10 years, declining with increasing age, and higher in males than females (27.3% v. 8.3%; P < 0.005). The mean intensity of infection among the infected males was also higher than that among the infected females [72.4 (4.8) v. 17.8 (2.5) eggs/g; P < 0.005]. Water contact by the subjects was estimated from activity diaries, for 141 days over the 2-year study period, and expressed as skin exposure, in m2 min/day. The mean exposure of a group of subjects was calculated by detransforming the mean of the fourth-root-transformed (i.e. normalized) values for the exposures of each subject within the group. Overall, the 213 individuals had a mean exposure of 6.2 m2-min/day. Differences in occupation led to males having much higher mean water exposures than females (9.2 v. 1.1 m2-min/day). As there was an inverse association between age-specific exposure and age-specific re-infection intensity, the marked reduction seen in intensity of re-infection with increasing age is not attributable to decreasing exposure to water. Instead, the results of this 2-year cohort study provide evidence for age-dependent resistance to re-infection with S. japonicum. The 213 subjects who were followed up were classified, according to epidemiological outcome at the end of the study and the data on water contact, as 'susceptible' (N = 49; 23%), 'insusceptible' (N = 29; 13.6%) or of 'uncertain status' (N = 135; 63.4%). Thus, 78 subjects who are potentially informative in terms of immunogenetics were identified. Further investigation of these individuals should help to shed some light on the role of immunogenetic status in human immunity to Schistosoma japonicum. PMID- 10707109 TI - Treatment of the microfilaraemia of asymptomatic brugian filariasis with single doses of ivermectin, diethylcarbamazine or albendazole, in various combinations. AB - Several new chemotherapeutic tools are now available for the control of lymphatic filariasis. Combinations of single doses of antifilarial drugs are generally superior to single drugs. The efficacy and safety of albendazole in combination with diethylcarbamazine (DEC) or ivermectin, for the treatment of Brugia malayi infection, were investigated, for the first time, in an open, hospital-based study. Fifty-one asymptomatic microfilaraemics (with 108-4034 microfilariae/ml; median = 531) of both sexes and aged 14-70 years were randomly allocated to receive single-dose treatments of ivermectin (200 micrograms/kg) with diethylcarbamazine (DEC; 6 mg/kg), ivermectin (200 micrograms/kg) with albendazole (400 mg), DEC (6 mg/kg) with albendazole (400 mg), or albendazole (400 mg) alone. Albendazole alone had no effect on the microfilarial levels at the 1-year follow-up but both groups given DEC had significantly lower microfilaraemias (P < 0.015 and P < 0.02) than that given ivermectin with albendazole. Overall, 47%-64% of those given DEC but only 14% of those given ivermectin with albendazole appeared to be amicrofilaraemic 1 year post treatment. The adverse reactions seen in the study were mild, transient and qualitatively similar to those seen earlier with ivermectin and DEC. The combination of DEC and albendazole, both well tested drugs, offers a new option for countries such as India where there is no onchocerciasis or loiasis and where ivermectin may not be immediately available. The direct and indirect effects of albendazole on intestinal helminths would be additional benefits. PMID- 10707110 TI - Community-directed, ivermectin-treatment programmes for onchocerciasis control in Uganda: the selection and validation of indicators for monitoring sustainability at the district level. AB - The selection and validation of indicators for predicting and monitoring sustainment in community-directed, ivermectin-treatment programmes (CDITP) at the community level in the Kabale, Kisoro and Rukungiri districts of Uganda have already been reported. The aim of the present study was to select and validate similar indicators at the district level, in the same districts over the same 4 year period. Three dependent-variable scales of programme sustainability (PS), PS1, PS2 and PS3, were compared by district. As at the community level, Rukungiri district clearly performed better than Kabale or Kisoro. Cost variables compiled at the district level and the ratios of numbers of community-based distributors (CBD) to community members were used as input variables in regression and correlation models, with PS1, PS2, and PS3 as outcome variables. In the regression model, cost of training CBD was found to be statistically significant (P = 0.0186). This variable also scored 100% on a scale for programme-indicator sensitivity and hence was selected as a helpful indicator. In the correlation model, cost of health education of community members had a weak relationship with PS1 (P = 0.0662). Cost of training CBD had a significant negative correlation with PS2 (P = 0.0186), indicating that reducing the cost of training would facilitate sustainability. PS3 showed weak negative correlations with cost of health education of community members (P = 0.0586) and cost/person treated in the district (P = 0.0584). Sustainment of CDITP might be better, therefore, if the costs per person could be reduced. As correlation relationships may not be linear, however, they were not considered particularly useful in the selection of helpful indicators. PMID- 10707111 TI - Infection rates in, and the number of Plasmodium falciparum genotypes carried by Anopheles mosquitoes in Tanzania. AB - Naturally fed mosquitoes were collected from houses in a rural village in northern Tanzania. The number of Plasmodium falciparum oocysts which developed in each was counted, and the number of parasite genotypes carried by a subset was determined by PCR-based techniques. A higher proportion of Anopheles gambiae s.l. mosquitoes developed oocysts than of An. funestus but, on average, both species carried similar numbers of parasite genotypes. Overall, 68% of the sampled mosquitoes carried more than one genotype. PMID- 10707112 TI - Cytochrome P450 substrate specificities, substrate structural templates and enzyme active site geometries. AB - The structural characteristics of human cytochrome P450 substrates are outlined in the light of extensive studies on P450 substrate specificity. Templates of superimposed substrates for individual P450 isozymes are shown to fit the corresponding enzyme active sites, where contacts with specific amino acid residues appear to be involved in the interaction with each structural template. Procedures leading to the evaluation of likely P450 specificity, binding affinity and rate of metabolism are described in the context of key examples in which molecular modelling appears to rationalize experimentally observed findings. PMID- 10707113 TI - Pharmacokinetic and pharmacodynamic interactions between dehydroepiandrosterone and prednisolone in the rat. AB - The effects of multiple-dosing with dehydroepiandrosterone sulfate (DHEA-SO4) on the pharmacokinetics and pharmacodynamics of prednisolone were examined. Prednisolone (25 mg/kg i.v.) was administered to male and female Sprague-Dawley rats (250-350 g) alone and following DHEA-SO4 (4 mg/kg i.v., every 8 h for 4 days). Male control rats cleared prednisolone faster [3.68 +/- 1.30 (males) vs 1.01 +/- 0.7 l/h/kg; p < 0.05] and had larger Vss (1.38 +/- 0.459 vs 0.394 +/- 0.500 l/kg; p < 0.05) than females both due largely to lesser plasma protein binding. Prednisolone clearance and Vss were not altered by DHEA-SO4 in males or females. The net effect of prednisolone on basophils and plasma corticosterone did not differ with gender. DHEA-SO4 had no effect on plasma corticosterone and did not alter prednisolone action. DHEA-SO4 inhibited basophil trafficking in males, but to a lesser extent than prednisolone, and antagonized the effect of prednisolone on basophil trafficking in both sexes. The steroid-sparing effect observed with DHEA clinically may not be due to an alteration of corticosteroid pharmacokinetics but partly to its ability to affect immune functions. PMID- 10707114 TI - Time dependent influence of diazepam on the pharmacokinetics of ibuprofen in man. AB - Circadian variation in the disease activity of rheumatoid arthritis has been established. Several nonsteroidal anti-inflammatory drugs have been studied for their chronokinetic behaviour. Time dependent influence of diazepam on the pharmacokinetics of diclofenac and naproxen has been reported. We report the time dependent influence of diazepam on the pharmacokinetics of ibuprofen in healthy subjects. Either ibuprofen or ibuprofen with diazepam was administered at 10.00 or 22.00 hours to eight healthy volunteers in a randomized crossover study. Serum ibuprofen levels were estimated by high performance liquid chromatography. There was a significant (p < 0.05) increase in mean elimination half life (2.39 +/- 0.42 to 3.59 +/- 0.35 h) following ibuprofen and diazepam administration compared to ibuprofen alone administered at 22.00 hours. The mean clearance of ibuprofen was therefore lowered from 62.7 +/- 8.9 to 41.7 +/- 2.6 ml/h/kg under the influence of diazepam during the night. Such a time dependent influence of diazepam on the pharmacokinetics of ibuprofen may be due to circadian variation in the pattern of protein production in the liver and/or competitive protein binding of the two drugs during the dark period. PMID- 10707115 TI - Dose independent pharmacokinetics of caffeine after intravenous administration under a chronic food-limited regimen. AB - Several studies have shown that caffeine follows non-linear pharmacokinetics in both rats and humans. Recent data have demonstrated that caffeine may following linear pharmacokinetics when administered orally and intraperitoneally to food limited rats. In this study the pharmacokinetics of caffeine was analyzed following intravenous (i.v.) administration to rats under a food-limited regimen. Four rats were administered four doses of caffeine and a standard dose of the caffeine metabolites, paraxanthine, theobromine, and theophylline. Caffeine pharmacokinetic parameters were dose independent following intravenous doses ranging from 1 to 20 mg/kg. Furthermore, the caffeine area under the curve (AUC) increased linearly as a function of dose. The mean fraction of caffeine converted to paraxanthine, theobromine, and theophylline was 16%, 16%, and 7%, respectively. The linear pharmacokinetics demonstrated in the present study may be attributed to the induction of hepatic metabolism under a chronic food-limited regimen. PMID- 10707116 TI - In vitro metabolism of (S)-(-)-[2'-14C]nicotine, using various tissue preparations of marmoset. AB - The nicotine metabolite profile produced by marmoset liver, lung and kidney preparations was investigated after 30 minutes incubation of (S)-(-)-[2' 14C]nicotine. Cation-exchange high performance liquid radiochromatography was employed to separate and quantify nicotine and its metabolites. Cotinine-N-oxide (CNO, 0.7%), 3'-hydroxy-cotinine (3'-OH-C, 0.2%), norcotinine (NORC, 0.9%) and nornicotine (NORN, 0.4%) were formed in the incubates of marmoset lung homogenates; when marmoset kidney homogenates were used, CNO, 0.4%; 3'-OH-C, 0.2%; NORC, 0.7%; NORN, 0.7%; and cotinine (COT, 0.4%) were detected in the incubates. These nicotine metabolites constituted only approximately 2.2% and 2.4% of the original nicotine substrate used by lung and kidney homogenates respectively. When marmoset hepatic homogenates and microsomes were used, both COT and NORN were detected as the major nicotine metabolites. In addition, traces of CNO and 3'-OH-C were also detected in both incubates. The amounts of COT (6.4%) and NORN (1.8%) in the hepatic homogenates were approximately twice that of those formed by hepatic microsomes (3.8% and 0.9%, respectively). Nicotine-1' N-oxide (NNO, 1.1%) was only detected in the latter preparation. Under the experimental conditions, these nicotine metabolites constituted only 8.2% and 5.8% of the substrate nicotine used in the respective incubates. The present results showed that both primary C-oxidation pathways, i.e. cotinine formation and N-demethylation of nicotine, occurred in the lung, kidney and liver of marmoset in vitro. However, N-oxidation of nicotine was only observed when a marmoset hepatic microsomal preparation was used. PMID- 10707117 TI - The in vitro hepatic microsomal metabolism of 3,5-dimethyl-4-(phenylazo)-(1H) pyrazole in rats. AB - The in vitro hepatic microsomal metabolism of 3,5-dimethyl-4-(phenylazo)-(1H) pyrazole (DMPAP) was studied using washed rat hepatic microsomal preparations fortified with NADPH. The substrate, DMPAP, and its potential metabolites, i.e. the corresponding reduction product, 3,5-dimethyl-4-amino-(1H)-pyrazole (DMAP), and the oxidation product, 3,5-dimethyl-4-(phenylazoxy)-(1H)-pyrazole (DMAPO), were synthesized and their structures elucidated by use of their spectral characteristics. DMPAP and its potential metabolites were then separated using a reverse phase HPLC system consisting of a C18 column and a mobile phase of acetonitrile:water (50:50) at a flow rate of 1 ml/min with UV detection at 254 nm. DMPAP was incubated with rat microsomal preparations, extracted into DCM in the presence of NaCl, and finally evaporated under a stream of nitrogen. The results from HPLC studies showed that DMPAP was metabolised to the corresponding reduction and oxidation products in the presence of NADPH. PMID- 10707118 TI - The in vitro hepatic microsomal metabolism of N-benzyladamantanamine in rats. AB - The metabolism of N-benzyladamantanamine (NBAD) was studied in vitro using rat hepatic microsomal preparations. The substrate and proposed metabolites were synthesized and characterized using spectroscopic techniques and separated using a reverse phase HPLC system. NBAD was incubated with rat microsomal preparations, extracted into DCM in the presence of NaCl and evaporated under a stream of nitrogen. The results from HPLC studies showed that NBAD produced the corresponding nitrone and hydroxylamine. This experiment also revealed that dealkylation occurred. No metabolites were observed which corresponded to authentic amide or oxaziridine. The reactions required a microsomal enzyme source and NADPH as a cofactor. The results indicate that the nitrone observed as a metabolite of NBAD is not an intermediate leading to the formation of an oxaziridine and hence an amide, under careful experimental conditions excluding light. PMID- 10707119 TI - The in vivo metabolism of 5-(4-nitrophenyl)-4-phenyl-2,4-dihydro-3H-1,2,4 triazole-3-thione in rats. AB - It is known that substituted 1,2,4-triazole-3-thione derivatives have several biological activities, such as antimicrobial, diuretic and antidepressant activities. In our previous studies, the antifungal activity of 5-(4-aminophenyl) 4-phenyl-2,4-dihydro-3H-1,2,4-triazole-3-thione was found to be active against Candida tropicalis K1022. The aim of this study was to investigate the in vivo metabolic pathway of 5-(4-nitrophenyl)-4-phenyl-2,4-dihydro-3H-1,2,4-triazole-3 thione which was selected as a model compound for this study. The substrate and its potential metabolites, i.e. the acetylation and nitro reductive products, were synthesized and then separated using HPLC on a reverse phase system. In the in vivo metabolism study, a 4 mg dose was administered i.p. to male Wistar rats. Blood samples were collected at 0, 2, 4, 8, 12, 24 and 56 hours after administration and were passed through a Sep-Pak cartridge. The acetylated metabolite [5-(4-acetylaminophenyl)-4-phenyl-2,4-dihydro-3H-1,2,4-triazole-3 thione ], the amine metabolite [5-(4-aminophenyl)-4-phenyl-2,4-dihydro-3H-1,2,4 triazole-3-thione] and an unknown metabolite were detected. PMID- 10707120 TI - Induction and inhibition of the in vitro N1-oxidation of 9-benzyladenine and isomeric 9-(nitrobenzyl)adenines. AB - The present study investigated some aspects of the enzymology of the in vitro N1 oxidation of 9-benzyladenine (BA) and isomeric 9-(nitrobenzyl)adenines (NBAs) using various potential inducers and inhibitors of cytochrome P-450 (CYP). When incubated with phenobarbital-induced rabbit hepatic microsomes, the N1-oxidation rates of BA and 9-(4-nitrobenzyl)adenine were about 6- and 2-fold higher than that of the control, respectively; while the N1-oxidation of 9-(2 nitrobenzyl)adenine and 9-(3-nitrobenzyl)adenine was not markedly affected. In contrast, beta-naphthoflavone and Arochlor 1254 showed no inductive effects towards the N1-oxidation of any of these substrates. Using 12 typical CYP inhibitors, it was found that nifedipine (CYP3A inhibitor) and haloperidol (CYP2D inhibitor) showed significant inhibition towards the N1-oxidation of BA and NBAs. Therefore, the N1-oxidation of BA and NBAs is probably catalysed by CYP3A and CYP2D subfamilies. Furthermore, when 9-(4-nitrobenzyl)adenine was incubated with compounds which possessed a certain chemical similarity to the adenine substrate, various degrees of inhibition of N1-oxidation of 9-(4-nitrobenzyl)adenine were observed. These observations allowed a preliminary indication as to the structure metabolism relationship of 9-substituted adenine derivatives. PMID- 10707121 TI - Comparison of metabolic rates of some 9-aralkyladenines obtained using hamster hepatic microsomes. AB - Previous investigations have revealed that N1-oxidation is a major metabolic pathway in vitro for some 9-aralkyladenines (AAs) such as 9-benzyladenine (BA). However, dealkylation and other metabolic pathways are also involved. In addition, various substituents on the benzyl moiety of BA seem to have a marked effect on the metabolic rate. In order to establish the potential structure metabolism relationship of this class of compounds, the enzyme kinetics of the substrates, which possess 2'-nitro (2NBA), 3'-nitro (3NBA), 4'nitro (4NBA), 2' chloro (2CBA), 2'-methyl (2MBA), or 2-methoxy (2MOBA) substitution of the benzyl moiety of BA, were compared using hamster hepatic microsomes. The results show that the formation rates of the N1-oxides are in the order 2NBA > 2CBA > BA > 3NBA and 4NBA > 2MBA and 2MOBA; the formation rates of the total metabolites except N1-oxides are in the order 2MOBA and 2MBA > 2CBA > BA > 4NBA > 3NBA > 2NBA; however, the total biotransformation rates of the substrates are in the order 2MBA and 2MOBA > 2CBA > BA and 2NBA > 4NBA > 3NBA. The results strongly imply that the electronic, steric and other physicochemical properties are potential controlling factors for AA metabolism. PMID- 10707122 TI - The in vitro hepatic microsomal metabolism of methyl 2-(2(3H)-benzoxazolone-3 yl)acetate in rats. AB - The in vitro hepatic microsomal metabolism of methyl 2-(2(3H)-benzoxazolone-3 yl)acetate (I) was studied using hepatic washed rat microsomal preparations fortified with NADPH. The substrate (I) and its potential hydrolytic metabolite 2 (2(3H)-benzoxazolone-3-yl)acetic acid (II) and 2(3H)-benzoxazolone (III), a potential dealkylation metabolite, were separated using a reverse phase HPLC system which consisted of a C18 column and a mobile phase of acetonitrile: 0.02 M phosphate buffer (30:70, final pH 7) at a flow rate of 1 ml/min with UV detection at 254 nm. The substrate (I) was incubated with rat microsomal preparations, extracted into DCM, and finally evaporated under nitrogen. The results from HPLC studies showed that (I) was metabolised to (II) and (III) by rat microsomes in the presence of NADPH. PMID- 10707123 TI - Beta-lactamase activity of anaerobic bacteroides strains isolated from clinical samples and their susceptibility to antimicrobial agents. AB - Beta-lactamase production and susceptibility to 13 antimicrobial agents (penicillin-G, amoxycillin, amoxycillin/clavulanic acid, cefoxitin, imipenem, clindamycin, metronidazole, piperacillin, ticarcillin, rifampicin, chloramphenicol, tetracycline and erythromycin) of 32 isolated Bacteroides strains were determined. The strains included 23 isolates of B. frugilis, 2 B. thetaioatomicron, 2 B. ovatus, 1 B. distasonis, 1 B. capillosus, 1 B. uniformis, 1 B. ureolyticus and 1 B. merdae. beta-Lactamase production was detected in 65% of the Bacteroides using the nitrocefin test, All the antibiotic agents showed excellent activity against beta-lactamase negative strains (for tetracycline, ticarcillin and clindamycin, 90% were susceptible, whereas for the other drugs, 100% were susceptible). beta-Lactamase-positive Bacteroides strains showed 95% susceptibility to metronidazole and rifampicin; 90% susceptibility to piperacillin and cefoxitin; 85% susceptibility to tetracycline and erythromycin; 80% susceptibility to clindamycin and amoxycillin/clavulanic acid, and 76% susceptibility to ticarcillin. All beta-lactamase-positive strains were found to be susceptible to imipenem and chloramphenicol. PMID- 10707124 TI - Synthesis and antibacterial activity of 1-(3-hydroxy-2-naphthoyl)-4-substituted thiosemicarbazides. AB - 1-(3-Hydroxy-2-naphthoyl)-4-substituted thiosemicarbazides were obtained by the addition of 3-hydroxy-2-naphthoic acid hydrazide to various isothiocyanates. The structures of the synthesized compounds were confirmed using UV and 1H-NMR spectral methods together with elemental analysis. PMID- 10707125 TI - Simultaneous determination of acetaminophen, acetylsalicylic acid and ascorbic acid in tablet form using HPLC. AB - The purpose of the present study was to develop a simple and accurate HPLC method to measure the amount of each agent in a multidrug pharmaceutical formulation. Three drugs, acetaminophen, acetylsalicylic acid and ascorbic acid, were analyzed simultaneously. A commercial pharmaceutical effervescent tablet was examined and the amount of each of these agents successfully determined. The present method appears to be more convenient than the current procedures described in American and British Pharmacopoeias in which each drug is assayed separately. PMID- 10707126 TI - Effect of tenoxicam on biochemical serum parameters of rats. AB - Tenoxicam is a nonsteroidal analgesic of the oxicam group, which possesses both antipyretic and anti-inflammatory characteristics. The use of tenoxicam has recently increased and it is reported in the literature that treatments lasting between a few weeks to three months caused increases in serum alanine transferase (ALT), aspartate transferase (AST), gamma glutamyl transferase (GGT) and bilirubin in humans. Toxic dose treatments to rats caused alterations in renal parameters. To verify these observations, various biochemical parameters were examined following administration of nontoxic doses of tenoxicam to rats. Rats were divided into three groups. One group received tenoxicam 0.6 mg/kg/day; the second group received 1.2 mg/kg/day i.p. The control group received normal saline i.p. At the end of 15 days, blood samples from the animals' hearts were taken for routine biochemical tests. No statistically significant changes were observed in serum urea, uric acid, creatinine, electrolytes, ALT, AST, total protein, bilirubin or glucose levels between the treatment groups and control groups. Increases in GGT levels were found to be statistically significant in both of the treatment groups compared with the control group. PMID- 10707127 TI - The in vivo effect of a Brassica oleracea var. capitata extract on Ehrlich ascites tumors of MUS musculus BALB/C mice. AB - An extract of Brassica oleracea var. capitata juice was prepared using petroleum ether, ether, ethanol and an Al2O3 column. The healing and tumor protecting effects of this extract were tested on Ehrlich ascites (EA) solid tumors of Mus musculus BALB/C mice. Complete disappearance of the tumors was observed in 54.5% of the animals in the experimental group (n = 22) which received 20 mg/day of the extract i.p. for 28 days. Regression of the tumors (27%), fixation of tumor size (4%) and an increase in tumor size (18%) were also recorded. Neither tumor size fixation nor regression was recorded in the control group which received physiological serum (0.5 ml/day). The healing effect was found to be related to the starting tumor size. The healed animals in the experimental group were followed for 6-7 months and no tumor recurrence was recorded. The protective effect of this extract on tumor formation was also tested. Experimental animals (n = 35) received 20 mg/day of the extract i.p. for 20 days. Physiological serum was administered to a control group (n = 30). Transplantation of solid tumors was performed on the 20th day and extract administration was discontinued. Transplantation success was recorded 20 days after transplantation. In the experimental group, only three out of 35 mice showed tumor development, whereas in the control group the number was 23 out of 35 mice. It was also observed that the extract prevented the development of liquid EA tumors. This extract was also found to be nontoxic. Brassica oleracea var. capitata had a healing effect as well as a protective effect on EA solid tumors of mice. These results are in agreement with our previous results obtained from a liquid Brassica oleracea var. acephala juice extract. PMID- 10707128 TI - Cataract surgery: who and how much? PMID- 10707129 TI - Pneumatic retinopexy: new breaks for old? PMID- 10707130 TI - Impact of cataract surgery on visual acuity and subjective functional outcomes: a population-based study in Sweden. AB - PURPOSE: First, to determine the effects of cataract surgery on subjectively experienced visual function and visual acuity in a defined population, at a specific frequency of surgery. Secondly, to validate questionnaire data regarding the visual function of cataract patients. METHODS: A prospective population-based investigation of the subjective visual functional and visual acuity outcomes of cataract surgery over a 1 year time interval at one institution was conducted. All operated cases (n = 459) were grouped into three levels of visual impairment, according to the preoperative visual acuities of their better eyes. Subjective reading, TV watching, distance estimation and ability to orientate in unfamiliar surroundings, before and after surgery, were assessed using self-administered questionnaires. The subjective outcomes were related to the subjects' post operative visual acuities. The statistical evaluations comprised analyses of variance, Yates'-corrected chi-squared tests, weighted kappa and correlation statistics. RESULTS: The pre-operative subjective visual disabilities of the patients were significantly correlated with the pre-operative visual acuities of the patients' better eyes. There was an improvement in subjective reading ability, distance estimation and ability to orientate in unfamiliar surroundings for most patients at all three pre-operative visual acuity levels. After surgery there was a stronger correlation between the subjective functional improvement and the increase in visual acuity for the operated eye than for the better eye. CONCLUSIONS: An incidence of cataract surgery of 3.3 per 1000 population for the year the present study was conducted seems not to be an over-utilisation of resources. Irrespective of the visual acuity level before cataract surgery, the vast majority of patients gain better subjective visual function and better acuity after surgery. It is possible to gain valid information from cataract surgery patients using a short questionnaire. PMID- 10707131 TI - The incidence of systemic side-effects following subconjunctival Mydricaine no. 1 injection. AB - PURPOSE: To investigate the cardiovascular response to the subconjunctival injection of 0.25 ml of Mydricaine No. 1 during vitrectomy surgery. METHODS: Pulse and blood pressure were recorded at 5 min intervals before and following the subconjunctival injection of Mydricaine No. 1 in a group of 49 sequential patients undergoing vitrectomy surgery under general anaesthetic during a 6 month period. These responses were compared with a sequential and similar group of 35 patients during the following 6 months. RESULTS: Ten patients in the group administered Mydricaine, but no patients in the control group, developed a sinus tachycardia of > 100 beats/min for more than 10 min which was attributable to the mydriatic regime used. The occurrence of this response was not predictable based on the patients' age, weight or the presence of conjunctival erythema. The magnitude and temporal course of the tachycardia observed were variable. Blood pressure recordings showed no clinically significant changes during the tachycardias. CONCLUSION: Twenty per cent of patients administered 0.25 ml of Mydricaine No. 1 subconjunctivally develop a significant sinus tachycardia following injection. This response is unpredictable and all patients given Mydricaine should be monitored carefully after injection. PMID- 10707132 TI - Surgically induced scleral necrosis. AB - PURPOSE: To present a group of patients with surgically induced scleral necrosis characterised by conjunctival retraction. METHODS: Three case reports are presented. RESULTS: The scleral melt responded to conjunctival covering of the scleral defect. CONCLUSION: Surgically induced scleral necrosis associated with conjunctival retraction is best managed by covering the exposed sclera either by stretching the retracted conjunctiva in the early post-operative period or by a conjunctival transplant. PMID- 10707133 TI - Pneumatic retinopexy in the treatment of primary rhegmatogenous retinal detachment. AB - PURPOSE: To review the management by pneumatic retinopexy of 31 primary rhegmatogenous retinal detachments performed between August 1994 and December 1997. METHODS: Ocular indications included superior retinal breaks, no evidence of proliferative vitreoretinopathy (PVR) and ability to posture. Patients with inferior breaks and/or areas of vitreoretinal degeneration were excluded. Surgery was performed under local anaesthetic using sulphur hexafluoride (SF6) or perfluoropropane (C3F8) gas injection. Transconjunctival cryotherapy or laser retinopexy was used to create permanent retinal adhesion. The mean length of patient follow-up was 11 months (range 5-24 months). RESULTS: Thirty-one patients (20 men, 11 women) with a mean age of 63.4 years (range 29-81 years) underwent pneumatic retinopexy which resulted in initial retinal reattachment in 22 patients. Two detachments recurred in the first month and a third at 4 months post-operatively, giving an anatomical reattachment rate with one procedure in 19 out of 31 eyes (61%). Of the 12 failures, 7 were reattached with one additional operation and one case reattached after multiple procedures, giving an overall reattachment rate of 87%. Post-operatively, new or missed breaks were present in 7 patients (22%) and PVR developed in 4 patients (13%). There was no difference in age, gender or extent of detachment between the failed and reattached groups and pseudophakia did not appear to be a poor prognostic factor. CONCLUSION: Pneumatic retinopexy can be a useful alternative to conventional rhegmatogenous retinal detachment surgery in carefully selected cases. A larger study addressing the influence of non-ocular factors is warranted. PMID- 10707134 TI - Ocular toxicity in low-dose tamoxifen: a prospective study. AB - PURPOSE: To look at the incidence, symptomatology, course and reversibility of low-dose tamoxifen ocular toxicity. METHODS: Sixty-five women with breast cancer, on tamoxifen oral therapy (20 mg/day), and a totally normal eye examination, were prospectively followed up. A full ophthalmic evaluation was done every 6 months, for a median of 30 months (range 4-79 months). Any sign of toxicity in the cornea, lens, retina or optic nerve was looked for, whether associated with a change in visual acuity or not. RESULTS: Ocular toxicity was documented in 8 patients, giving an incidence of 12%. Seven patients had keratopathy in the form of subepithelial deposits, whorls and linear opacities. Three of these patients had a concurrent symptomatic bilateral pigmentary retinopathy that warranted discontinuation of therapy. One patient developed bilateral optic neuritis that left her with optic nerve pallor and a decrease in vision. The patients who had the toxicity had a significantly higher tamoxifen cumulative dose (p = 0.03), and were longer on treatment (p = 0.04), than the non-affected ones. The keratopathy changes were reversible upon discontinuation of the drug. CONCLUSION: Prompt reporting of symptoms and yearly ophthalmic examinations are mandatory in patients on tamoxifen to detect toxic effects while these are still reversible. PMID- 10707135 TI - The effect of anthocyanosides in a multiple oral dose on night vision. AB - PURPOSE: In view of research demonstrating the ability of anthocyanosides in a multiple oral dose to improve night vision in normal individuals, we assessed their effect on three night vision tests: full-field absolute scotopic retinal threshold (SRT), dark adaptation rate (DAR) and mesopic contrast sensitivity (MCS). METHODS: In a double-masked, placebo-controlled, cross-over study, 18 young normal volunteers were randomly assigned to one of three different regimens of multiple oral administrations of 12 and 24 mg anthocyanosides, and a placebo, given twice daily for 4 days. A 2 week washout period was allowed between each 4 day treatment period. SRT, DAR and MCS was tested 1 day before and at days 1, 2, 3 and 4 during the treatment period. RESULTS: No significant effect was found on any of the three above-mentioned night vision tests. The study had a power of 0.95 to detect a 0.1 log unit improvement in SRT and 0.5 log unit improvement in MCS. CONCLUSIONS: Multiple oral administrations of 12 and 24 mg anthocyanosides twice a day appear to lack significant effect on night vision tests. PMID- 10707136 TI - Corneal guttata: a comparative clinical and specular micrographic study. AB - BACKGROUND: The relationship between corneal guttata and Fuchs' endothelial dystrophy is unclear, with the result that the clinical differentiation of the two conditions is often made on the basis of the presence or absence of symptoms. METHODS: In this study the authors compare the clinical and specular micrographic findings, as recorded from 20 patients noted to have biomicroscopic clinical findings consistent with corneal guttata or early Fuchs' endothelial dystrophy. RESULTS: Results confirm the increased prevalence of corneal findings in elderly women. The topographical distribution of guttata, across the posterior corneal surface, as observed clinically was confirmed specular micrographically (rS = 0.55, p = 0.01). Increased numbers of guttata correlated with a statistically significant reduction in the endothelial cell counts recorded from the midperipheral cornea (rS = 0.49, p = 0.02). The relationship between the presence of guttata and a reduction in cellular hexagonality or an increase in polymegethism failed to reach a statistically significant level. Pigment deposits adherent to the posterior endothelial surface were also noted in all but one of the eyes examined. CONCLUSIONS: The authors advocate the use of a grading scale, developed from specular micrographs taken during the course of the study, to assist in the clinical classification of corneal guttata and pre-clinical Fuchs' dystrophy. The authors also recommend specular microscopy as a tool capable of differentiating corneal guttata from pigment deposits in even the most severely affected cases. PMID- 10707137 TI - Comparison of topical 0.3% ofloxacin with fortified tobramycin plus cefazolin in the treatment of bacterial keratitis. AB - PURPOSE: Ofloxacin is a broad spectrum fluoroquinolone antibiotic with good ocular penetration. We compared ofloxacin 3% solution with a combination of fortified tobramycin sulphate and cefazolin sodium solutions in the treatment of culture-proven bacterial keratitis. METHODS: Thirty eyes with culture-proven bacterial corneal ulcers were enrolled in a prospective randomised, controlled, double-masked study for comparison. The ofloxacin drop and saline were decanted into two identical-looking bottles to the tobramycin and cefazolin. The cases were randomly allocated into treatment with 0.3% ofloxacin solution or a combination of fortified antibiotics (1.5% tobramycin and 10% cefazolin solutions; control group) along with supportive cycloplegic, vitamins and anti glaucoma therapy. Student's t-test was used to compare the results. RESULTS: Staphylococcus aureus and coagulase-negative staphylococci were the two most common organisms isolated. Resolution of the ulcer was achieved in 93% and 87% of cases in the ofloxacin and control groups respectively. The mean time required for symptomatic relief was 7.8 +/- 1.54 days and for epithelial healing 15.0 +/- 3.86 days in ofloxacin group, compared with 8.33 +/- 1.54 days for symptomatic relief and 15.46 +/- 3.86 days for epithelial healing in the control group. Post resolution best corrected visual acuity of 20/200 or better was achieved in all but one eye in both groups. CONCLUSIONS: Ofloxacin and combined fortified tobramycin and cefazolin topical drops were comparable for treating cases of bacterial corneal ulcer. However, considering its easy availability and cost effectiveness, monotherapy with ofloxacin is preferred over the combined fortified tobramycin and cefazolin therapy. PMID- 10707138 TI - The ocular and systemic prognosis of patients presenting with sarcoid uveitis. AB - PURPOSE: To describe the visual and systemic outcomes in patients presenting with sarcoid uveitis. METHODS: Seventy-five patients with definite or presumed sarcoid uveitis were followed up for a median of 4 years. The patients came from a primary ophthalmic referral centre and a specialist uveitis centre. The prognostic value of demographic and clinical features at the onset of disease were studied. Baseline and outcome variables were analysed by survival analysis. RESULTS: After 10 years, 54% of patients retained normal visual acuity and 4.6% had lost vision to less than 6/36 in both eyes. Fifty-one per cent required oral steroids for uveitis and a further 11% needed additional immunosuppressants. Twenty-one per cent of patients had undergone a surgical procedure. At the onset of uveitis the lung was the most common organ involved (35%). After 10 years follow-up disease spread to other organs in 13 patients (17%); in 8 of 13 patients this was the central nervous system. The only outcome associated with baseline variables was bilateral visual loss, which was more likely in those over 40 years at presentation (p = 0.004). CONCLUSIONS: The ocular prognosis of sarcoid uveitis is unrelated to the extent of disease at onset. Patients with extraocular disease fared no differently from those with isolated ocular disease. Patients with sarcoid uveitis are at risk of neurological involvement for at least 15 years. PMID- 10707139 TI - Comparison of the erythrocyte sedimentation rate measured in the eye casualty department by the Seditainer method with an automated system. AB - PURPOSE: To compare a new automated system for the measurement of erythrocyte sedimentation rate (ESR) with the established manual Seditainer method. METHODS: Two hundred and twelve patients undergoing investigation for giant cell arteritis or other systemic vasculitides had ESR measurements by both the established manual Seditainer and the new laboratory-based automated system. The results were compared by correlation coefficient and mean difference. The limits of agreement with confidence intervals were also calculated. RESULTS: Across the range of results from 1 to 120 mm/h, the correlation coefficient was 0.844. The automated method had a mean negative bias of -9.8 mm/h (95% confidence interval: -12.2 to 7.4 mm/h). The wide scatter of results produced limits of agreement (+/- 2 standard deviations) between the two methods of -45 to 26 mm/h. There were seven results that were underestimated by the automated system which were clinically significant. CONCLUSIONS: There is a wide degree of scatter between the two sets of results. The automated system has a negative bias when compared with the manual method. There is a propensity for the automated system to sporadically underestimate the true result, sometimes to a degree that is clinically significant. The authors therefore cannot recommend replacement of the manual Seditainer system at the present time. PMID- 10707140 TI - The lack of efficacy of topical beta-blockers, timolol and betaxolol on intraocular pressure in Nigerian healthy volunteers. AB - PURPOSE: The beta-adrenoceptor antagonists are commonly used drugs in ophthalmic and medical practice. While beta-blockers may show reduced antihypertensive efficacy in African patients, the effect of beta-blockers on intraocular pressure (IOP) in African healthy volunteers is less well known. METHODS: This single masked, placebo-controlled, randomised study was conducted to investigate the response of healthy Nigerian volunteers to a single drop of the beta-adrenoceptor antagonists timolol and betaxolol. Twenty-five volunteers participated in the study; however, only 19 were able to complete the study. The concentrations of the beta-blocker used were 0.0625%, 0.125%, 0.25% and 0.5%. One eye of the volunteers was used while the other eye served as control. The baseline IOP was documented and IOP measured hourly over 6 h. Pupillary size, corneal sensitivity and visual acuity were also assessed. Cardiovascular parameters were also documented hourly (blood pressure, heart rate, pulse rate). RESULTS: Only 0.5% concentrations of both beta-adrenoceptor antagonists caused any significant IOP reduction in normal volunteers (p < 0.05). The maximal falls were -2.33 +/- 2.2 mmHg and -1.23 +/- 0.6 mmHg with timolol and betaxolol, respectively. The IOP reduction produced lasted for only 4 h, after which the IOP returned to baseline. There was also an overshoot of the IOP above the baseline values in all the volunteers (+2.0 to +2.3 mmHg). There was no significant change in the cardiovascular parameters. There was no effect on the pupillary size, visual acuity or corneal sensitivity. There was no significant change in IOP and cardiovascular parameters in the placebo group. CONCLUSIONS: Both beta adrenoceptor antagonists caused an attenuated IOP reduction in African normal volunteers, compared with values reported in Caucasians. There was a rebound intraocular hypertensive effect demonstrated with both beta-adrenergic antagonists in blacks. PMID- 10707141 TI - Aneurysmal bone cyst of the orbit: a case report and review of literature. AB - Aneurysmal bone cyst is a benign fibroosseous lesion which rarely occurs in the orbit. We report on a 7-year-old girl with aneurysmal bone cyst of the orbit who presented with painless proptosis and diplopia. Optic nerve compression resulted in field loss and delayed visual evoked potentials. Radiological and histological features are discussed. The lesion was excised via a frontal craniotomy and the orbital roof reconstructed with a prefabricated titanium plate. Post-operatively a rapid resolution of the proptosis and diplopia followed. Previous reported cases of this rare entity in the orbit are also reviewed. PMID- 10707142 TI - Evaluation of bupivacaine-induced muscle regeneration in the treatment of ptosis in patients with chronic progressive external ophthalmoplegia and Kearns-Sayre syndrome. AB - PURPOSE: Ptosis is common in patients with mitochondrial disease. Whilst surgical shortening of the levator muscle can mechanically elevate the lid, this procedure does not restore normal movement and leaves patients at risk of corneal exposure due to concomitant ophthalmoparesis. Recent studies have shown that bupivacaine induced muscle regeneration is capable of reversing the molecular genetic and biochemical defect in patients with mitochondrial myopathies. This study was undertaken to assess the potential of this approach in restoring levator muscle function in patients with mitochondrial disease and ptosis. METHODS: The levator muscle of one eye in five patients with molecularly genetically confirmed mitochondrial DNA disease and ptosis was directly injected with 3 ml of bupivacaine hydrochloride (0.75%). Levator function was compared before and 3 months after the injection. RESULTS: No objective clinical improvement in levator function was detected following bupivacaine administration. DISCUSSION: The lack of functional recovery seen in our patients is most likely to result from a failure of bupivacaine to induce sufficient regeneration necessary to improve levator muscle function. This result indicates that consideration now needs to be given to the use of alternative and more potent myotoxic agents capable of inducing a more widespread regenerative response from the endogenous muscle satellite cells which contain low or undetectable amounts of mutant mitochondrial DNA. PMID- 10707143 TI - Weill-Marchesani syndrome in three generations. AB - BACKGROUND: Weill-Marchesani syndrome is a rare systemic connective tissue disorder consisting of brachymorphy, brachydactyly, ectopia lentis, spherophakia and glaucoma. METHODS: We report 6 patients with Weill-Marchesani syndrome (with or without ocular involvement) in three generations, identified by screening 26 members of two families. This is the largest family in the literature showing an autosomal dominant pattern of inheritance. RESULTS: Presenile vitreous liquefaction was present in all the younger cases. Weill-Marchesani syndrome was full-blown in two cases in the third generation, in which asymmetrical axial length and glaucomatous damage were present. To our knowledge this is the first report regarding asymmetrical axial length and glaucomatous damage, and presenile vitreous liquefaction in Weill-Marchesani syndrome with or without ocular involvement. CONCLUSIONS: The longer axial length might be the precursor of impending severe glaucomatous damage. Presenile vitreous liquefaction in subtle young cases should alert the physician to the diagnosis of Weill-Marchesani syndrome on screening of the family members. PMID- 10707144 TI - Congenital unilateral buphthalmos in Walker-Warburg syndrome: a clinicopathological study. AB - BACKGROUND AND PURPOSE: Walker-Warburg syndrome is a congenital autosomal recessive oculocerebral disorder characterised by hydrocephalus, brain agyria, microphthalmos and retinal dysplasia with or without meningoencephalocele. We describe an unusual finding of congenital unilateral glaucoma and buphthalmos in one eye and microphthalmos in the fellow eye of two neonates with Walker-Warburg syndrome. PATIENTS: Two neonates with Walker-Warburg syndrome and unusual findings of buphthalmos in one eye and a microphthalmic fellow eye are presented. RESULTS: Histological examination of the buphthalmic eyes revealed the presence of mesenchymal tissue in the anterior angle covered by endothelium. No anterior chamber angle was identified in the microphthalmic fellow eye and the iris was adherent to the corneal periphery. CONCLUSIONS: Congenital buphthalmos may also appear in Walker-Warburg syndrome. The buphthalmos may result from later embryonal ocular developmental arrest than that of the microphthalmic eye. PMID- 10707145 TI - Ocular, cerebral and systemic interrelationships of cytomegalovirus infection in a post-mortem study of AIDS patients. AB - PURPOSE: Eighty-six post-mortems of AIDS patients were reviewed microscopically and the presence of cytomegalovirus (CMV) infection in the viscera, brain and eye was recorded. METHODS: Immunohistochemical stains and in situ hybridisation with a CMV probe were performed. RESULTS AND CONCLUSION: CMV infection was observed in 63% of the cases. Visceral, cerebral and ocular involvement were overall 49%, 33% and 29%, respectively. The visceral form with no concomitant ocular and/or cerebral infection was the main cause of death (31%) in the 54 CMV-infected patients. Although CMV retinitis occurred mostly (20%) as a component of systemic disease, in 13% of the CMV-infected patients the eyes only were involved, while there were no cases with CMV limited to the brain. In the absence of systemic involvement, 9% of the cases showed concomitant ocular and cerebral infection, but because we failed to observe CMV optic neuritis without ocular involvement, retrograde viral spread from the brain through the optic nerve appears to be an infrequent mechanism of CMV retinitis. PMID- 10707146 TI - Neovascular glaucoma after bypass surgery in Takayasu's disease. PMID- 10707147 TI - Conjunctival haemangioma in an elderly patient. PMID- 10707148 TI - Primary surgical management in a case of subtotal iridodialysis. PMID- 10707149 TI - Treatment of preretinal Valsalva haemorrhages with neodymium:YAG laser. PMID- 10707150 TI - Spontaneous acute scleritis and scleral necrosis in choroidal malignant melanoma. PMID- 10707151 TI - Choroidal ischaemia and serous retinal detachment in toxaemia of pregnancy. PMID- 10707152 TI - Severe systemic toxicity caused by brimonidine drops in an infant with presumed juvenile xanthogranuloma. PMID- 10707153 TI - Hormone replacement therapy and retinal vein occlusion. PMID- 10707154 TI - Ischaemic retinal vasculitis in biopsy-proven sarcoidosis. PMID- 10707155 TI - Lens notching in association with presumed Marfan's syndrome. PMID- 10707156 TI - CSR-like presentation in epidemic dropsy. PMID- 10707157 TI - One-and-a-half syndrome: a different type. PMID- 10707158 TI - Late clouding of an acrylic intraocular lens following routine phacoemulsification. PMID- 10707159 TI - Familial intermediate uveitis: a case report of two brothers. PMID- 10707160 TI - The National Survey of Local Anaesthesia for Ocular Surgery. PMID- 10707161 TI - Was the Spitfire the catalyst for the plastic multi-focal lens? PMID- 10707162 TI - Tono-Pen tonometer and corneal thickness. PMID- 10707163 TI - The role of ophthalmic triage and the nurse practitioner in an eye-dedicated casualty department. PMID- 10707165 TI - Quality assurance and locum doctors in NHS trusts. PMID- 10707164 TI - Measuring surgical outcome. PMID- 10707166 TI - Early prevention of fracture in osteoporosis: new developments. PMID- 10707167 TI - Evidence-based management of reproductive problems. AB - Evidence-based medicine has become the catch-cry phrase of the 1990s. It is about using evidence from well-designed research and applying it to everyday practice. It would be a difficult task to appraise all the evidence from research without the help of well-prepared systematic reviews. This article examines some aspects of this approach. PMID- 10707168 TI - External cephalic version. AB - The management of breech presentation at term is variable. This article presents the options, and reviews the current evidence for, and success rate of, external cephalic version. The work of the breech clinic at the Liverpool Women's Hospital is also discussed. PMID- 10707169 TI - Reproductive health in women with cystic fibrosis. AB - The life expectancy of women with cystic fibrosis has doubled in the last 20 years. A major implication of this is the advent of previously unseen reproductive health problems. We review the management problems presented by these women throughout their reproductive lives, including pregnancy. PMID- 10707170 TI - Postpartum haemorrhage. AB - Postpartum haemorrhage can be either primary (within 24 hours of delivery) or secondary (within the following weeks). This article reviews the factors that may help anticipation of postpartum haemorrhage, and looks at issues involved in the management and treatment of women with this condition. PMID- 10707171 TI - Palivizumab: an overview. AB - Respiratory syncytial virus (RSV) affects almost all children in their first 2 years of life and can cause severe or even life-threatening disease in some at risk infants. Treatment is limited and there is currently no safe or effective vaccine. However, a new monoclonal antibody, palivizumab, reduces RSV hospitalization by 55% in at-risk groups if given prophylactically throughout the RSV season. PMID- 10707172 TI - Radionuclide imaging of the heart. AB - Nuclear cardiology is an established part of diagnosis and assessment of patients with possible heart disease, the two most common tests being myocardial perfusion imaging and radionuclide ventriculography. Myocardial perfusion imaging comprises approximately 75% of nuclear cardiology studies in the UK, and is used in diagnosis and management of coronary artery disease. PMID- 10707173 TI - Haemochromatosis: a time for guidelines? AB - Hereditary haemochromatosis is an autosomal recessive metabolic abnormality which causes excessive absorption of dietary iron. Iron accumulation leads to potentially fatal damage to organs such as the heart, liver and pancreas. Normal life expectancy can be restored simply by therapeutic venesection. Discovery of the gene, HFE, has rekindled interest in pathogenesis, management and screening strategies. PMID- 10707174 TI - Clinical governance and revalidation. AB - The quality agenda in the NHS has many parts. For the medical profession the means of enhancing standards and consistency will be revalidation. This article outlines progress to date in defining how doctors will demonstrate they are 'up to date' and 'fit to practice'. PMID- 10707175 TI - Bias in surveys. AB - The simple survey is a regular tool in health services research. However, like any research method, surveys can be flawed in design, execution, analysis or interpretation. This short article outlines the basis of good survey design and advises how bias in published studies can be assessed. PMID- 10707176 TI - Medical workforce and the gender shift. AB - The future medical staffing for the NHS needs to take account of the demands of doctors for a humane lifestyle and the service demands of all groups of patients. The increasing numbers of female practitioners and the reduction in hours required by new legislation will be important influences. Suggestions are made about possible future working patterns. PMID- 10707177 TI - A pilot Internet teaching project to support specialist medical training. AB - Regional training programmes involving specialist medical trainees at geographically separate sites lend themselves to distance learning methods. This paper describes the setting up and early evaluation of an internet-based project designed to support regional study days across North East Thames for respiratory medicine. PMID- 10707178 TI - Acute adrenocortical crisis and an abnormal electrocardiogram. PMID- 10707179 TI - Squamous cell carcinoma. PMID- 10707180 TI - Spontaneous rupture of the spleen as a result of infectious mononucleosis in two siblings. PMID- 10707181 TI - The role of the surgeon in the ICU. PMID- 10707182 TI - Effective anticoagulation. PMID- 10707183 TI - Percutaneous central venous catheter placement. PMID- 10707184 TI - Anaesthesia for caesarean section: the debate continues. PMID- 10707185 TI - The artificial heart: current concepts. PMID- 10707186 TI - Breastfeeding and HIV infection: a global perspective. PMID- 10707187 TI - Interpretation of the neonatal chest X-ray. AB - Most neonatal X-rays are seen initially by a paediatrician without formal training in interpretation of chest X-rays. This article aims to help improve the information obtained from these X-rays which are often complex. Many factors affect accurate interpretation of the neonatal chest X-ray, including good quality radiographs, appropriate viewing conditions and thorough education. PMID- 10707188 TI - Diagnosis of hip pain in children. AB - Hip pain is uncommon in children and imaging plays a central role in diagnosis and follow-up. The commonest cause of hip pain is acute transient synovitis. Other causes, which vary according to the age group of the child, include Perthes' disease and slipped capital femoral epiphyses. This review discusses the differential diagnosis and imaging pathways required. PMID- 10707189 TI - Radiological features of non-accidental injury. AB - Non-accidental injury is not uncommonly met in clinical practice. One should be aware of its presentation, the radiographic signs suspicious of abuse and the appropriate further imaging assessment to confirm or refute the diagnosis. Erroneous diagnosis can have grave consequences. PMID- 10707190 TI - Insulin lispro for the treatment of type 2 diabetes. AB - Tight control can prevent complications in type 2 diabetes, and many patients will require insulin therapy to achieve this. Newer insulin formulations may offer some advantages with regard to patient convenience and a reduction in hypoglycaemia. PMID- 10707191 TI - Current practice in thoracic sympathectomy. AB - Thoracic sympathectomy has been performed for many years. With the recent development of video assisted thoracic surgical techniques the indications for surgery have increased, and the outcome is much better. PMID- 10707192 TI - Catatonia. 2: Diagnosis, management and prognosis. AB - This is the second of two articles reviewing catatonia. In the first, catatonia was described as an under-recognized syndrome with a potentially fatal outcome. It was suggested that treatment with neuroleptics may exacerbate the syndrome. The differential diagnosis, management and prognosis of catatonia are reviewed below. PMID- 10707193 TI - Use of apomorphine in Parkinson's disease. AB - Apomorphine is a dopamine agonist administered subcutaneously as intermittent injections or in a continuous infusion. It is useful in managing advanced Parkinson's disease, and provides an alternative to neurosurgical procedures. This review summarizes indications and practical guidelines for its use. PMID- 10707194 TI - Clinical governance in primary care. AB - The government's stated top priority for the NHS is the quality of care of patients and there now exists a new duty of quality on all those providing care, accompanied by a drive for greater accountability to the public of clinical decision making. The process which governs these initiatives is termed clinical governance. PMID- 10707195 TI - Staff grade doctors and the consultant ladder: falling off or stepping off? AB - The majority of the doctors in staff grade posts surveyed had qualified overseas and a substantial proportion were dissatisfied with their posts. Very few doctors had received adequate careers counselling before starting their post and many were concerned at the lack of opportunities for career development. PMID- 10707196 TI - Can women have careers as staff grade doctors? PMID- 10707197 TI - Addressing issues: status and career development. PMID- 10707198 TI - Representing staff grade doctors. PMID- 10707199 TI - Postgraduate education and training: challenges for the future. AB - In spite of much attention to the needs of doctors in the training grades over the last decade, the current agenda for change in the NHS is very large and much needs to be done to fit these trainees for the career grade doctors of the future. This article looks briefly at some of the issues facing training grade doctors as they train for a career in the NHS in the first part of the twenty first century. PMID- 10707200 TI - Folie a deux in a Seychellois mother and adult son. PMID- 10707201 TI - Hip pain in the third trimester of pregnancy. PMID- 10707202 TI - It's not over 'til the fat lady sings. PMID- 10707203 TI - Evidence-based stroke management. PMID- 10707204 TI - Guillain-Barre syndrome and osteoporosis. PMID- 10707205 TI - Management of severe upper airway trauma. PMID- 10707206 TI - Open tibial fractures: faster union after unreamed nailing than external fixation. AB - Unreamed intramedullary nailing is an alternative to external fixation in the treatment of open tibial fractures. We compared a prospective series of thirty one patients managed with a solid nail with static interlocking without intramedullary reaming, with a retrospective series of thirty-one patients managed by external fixation. The protocol for soft tissue treatment was the same throughout the study period. Most fractures were caused by high energy trauma and included Grade I to III B injuries. The fracture wound infection rate was equal in both groups; there were two deep and three superficial infections in the nail group and three deep and two superficial infections in the external fixation group. In addition, eleven patients in the external fixation group had severe pin track infections. The mean time to union was five months in the nail group and eight months in the external fixation group. The incidence of delayed union was twice as high in the external fixation group as in the nail group. The number of surgical procedures performed to promote union was three times higher in the external fixation group. The malunion rate did not differ between the groups. Although the treatment groups are not fully comparable, the results indicate that intramedullary nailing is superior to external fixation in the treatment of most open tibial fractures. PMID- 10707207 TI - Bone transport over an intramedullary nail. A case report with histologic examination of the regenerated segment. PMID- 10707208 TI - Post traumatic pseudoaneurysm of an intrasplenic segmental artery; a mechanism of delayed splenic rupture. PMID- 10707209 TI - Unusual treatment of slaughterer's gun injury. PMID- 10707210 TI - The use of hinged Kirschner wires for fixation of patellar tendon rupture. PMID- 10707211 TI - Plate fixation of proximal humeral fractures with indirect reduction: surgical technique and results utilizing three shoulder scores. AB - To determine the outcome after indirect reduction and buttress plate fixation of displaced and unstable proximal humeral fractures, we retrospectively evaluated 98 patients, an average of 34 months (range 24-72 months) after fracture fixation. The patients were reviewed and results were evaluated clinically according to the Neer, UCLA and Constant score. A radiographic evaluation of fracture healing, avascular necrosis and degenerative changes of the shoulder joint was performed in all patients. Any complications of treatment were assessed. Results were, according to the UCLA-rating system, good to excellent in 76% of fractures. According to the Constant-score and the Neer score, good to excellent results were obtained in 69 and 59% of fractures, respectively. Poor results were mainly due to secondary malunion. The avascular necrosis rate was 4%. Non-union was seen in one case. Secondary varus deformity and retroversion of the humeral head as a result of lack of rotational and angular stability of the plate developed in twelve (12%) and eight (8%) cases, respectively. Plate fixation is an adequate procedure for treating unstable and displaced two- to four-part fractures of the proximal humerus, enabling early functional after treatment. The incidence of avascular necrosis and nonunion are low, when fracture reduction is performed indirectly. Poor rotational and angular instability can lead to a loss of reduction. PMID- 10707212 TI - Emergent management of multisystem injuries by a general trauma surgeon. AB - OBJECTIVE: To determine the requisite education and scope of practice for a general surgeon trained to deliver emergent trauma care without the need for specialty consultation. DESIGN: Retrospective case review. SETTING: Private Level I trauma center. PATIENTS: 4097 trauma patients admitted between 1/1/92 and 30/6/97. MAIN OUTCOME MEASURES: Mechanisms of injury; operations (total within the first 24 h) by mechanism and by surgical specialty. RESULTS: Of 4097 trauma patients, 1086 (27%) underwent 1772 operative procedures within 24 h of admission, and 246 (6%) underwent 484 later operations. Orthopaedic and general surgical procedures were most common (51% of early operations). Early operations were most commonly orthopaedic for blunt trauma and general surgical for penetrating trauma. Although 685 patients (16.7%) received neurosurgical evaluation, only 150 early operations were performed (8% of the total early operations). At least 1244 procedures (55%) fell within the scope of current trauma general surgical practice advocated by some authorities. CONCLUSIONS: The contribution of the various specialties to early operative trauma management is a function of the injury mechanism. As orthopaedics and neurosurgery together comprise 34% of emergency practice, a fully credentialed general surgeon with additional training in these disciplines could perform up to 90% of early operations. PMID- 10707213 TI - CT assessment of torsion following locked intramedullary nailing of tibial fractures. AB - The aim of our study is to determine the incidence and extent of torsional malalignment in patients with tibial diaphyseal fractures treated with closed antegrade intramedullary (IM) nailing. We measured torsion using CT scanning in 22 patients. A difference of 8 degrees or more was found in 8 cases (36%) as compared with the uninjured side. Only 2 of these cases could be clinically detected and only one patient noted the problem. We also describe a method of directly calculating torsion from CT images captured on a single film that is easy and does not require any special equipment. PMID- 10707215 TI - The supracondylar intramedullary nail in elderly patients with distal femoral fractures. AB - From February 1994 until July 1997, a prospective study of all elderly patients with a displaced distal femoral fracture, who were treated with an intramedullary supracondylar nail, was made. The outcome of 31 fractures in 30 elderly patients was studied. The average age was 82 years (55-98). Two-thirds of the patients had had previous ipsilateral femoral pathology. Average acute hospital stay was 17 days. All patients were reviewed at 6 months and all cases have been followed for over 1 year. More than 90% of surviving fractures had healed within 6 months of the operation. Outcome scores were; 22 (85%) excellent or satisfactory, 2 (7.5%) unsatisfactory and 2 (7.5%) failures. The mortality rate was 17% at 6 months and 30% at 1 year, which is similar to patients with a fractured neck of femur. This nail is recommended for its versatility and favourable outcome scores in this age group. PMID- 10707214 TI - Abdominal injuries associated with lumbar spine fractures in blunt trauma. AB - BACKGROUND: Specific analysis of the relationship between abdominal injuries and lumbar spine fractures has not yet been reported. METHODS: A retrospective review of 258 blunt trauma patients with lumbar spine fractures treated between 1991 and 1996. RESULTS: 26 patients sustained concomitant lumbar spine fractures and abdominal injuries. The mechanism of injury was motor vehicle collision (73%), pedestrian-struck (11%), fall (8%) and assault (8%) resulting in ISS, RTS and mortality of 27 +/- 4, 6.5 +/- 0.4 and 8%, respectively. Forty-four lumbar spine fractures were identified (1.7/pt) in association with splenic (54%), renal (41%), hepatic (32%) and small bowel (23%) injuries and no retroperitoneal involvement. Multilevel lumbar spine fractures were associated with a higher organ injury/fracture ratio compared with single level fractures (p < 0.01) including a twofold higher incidence of solid organ (spleen, liver and kidney) injury (p < 0.01). The level and type of fracture did not affect the incidence of total and individual organ injury. Patients with abdominal injuries were more severely injured mainly due to increased incidence of associated thoracic injuries although no significant difference in mortality was observed. CONCLUSION: Abdominal injuries occurred only in the minority of blunt trauma patients with lumbar spine fractures. These injuries, which followed a similar distribution pattern as in blunt trauma in general, occurred most commonly due to motor vehicle collisions and in association with multilevel vertebral fractures. No correlation with fracture type or level was identified. PMID- 10707216 TI - Lateral canthotomy and inferior cantholysis: an effective method of urgent orbital decompression for sight threatening acute retrobulbar haemorrhage. AB - Retrobulbar haemorrhage (RBH) occurs in a variety of situations. It can complicate facial fractures, orbital surgery and retrobulbar injections and can occur spontaneously. It is relatively uncommon and sight-threatening RBH is even less common. If not detected early enough it can lead to devastating loss of vision. We have collected five cases of acute RBH, following trauma, associated with a profound reduction in vision. In each case a permanent loss of vision was avoided using a lateral canthotomy and inferior cantholysis approach to obtain urgent orbital decompression. PMID- 10707218 TI - Plate fixation of fresh displaced midshaft clavicle fractures. AB - From 1992-1994, we operated on 251 fresh completely displaced mid-third clavicle fractures in adults; 232 were followed up. The fractures were plated with a Mizuho C-type plate or an AO/ASIF 3.5 mm reconstruction plate. Comminuted fragments were reduced and wired (133 cases). There were 150 men and 82 women; the median age was 37.3 years (range 18-79). The mean follow-up was 4.4 years (range 3.0-5.9). The mean time to radiographic union was 10 weeks. Seven patients (3%) developed nonunion. Healing with angulation occurred in 14 patients. Deep infection developed in one patient, and superficial infection in four cases; 21 patients reported soreness with changes in the weather and activity; 28 patients had residual skin numbness caudal to the incision. No patient had shoulder droop, and none had impairment of range of motion or shoulder strength. None developed new or late neurovascular impairment; 171 patients eventually had the hardware removed at an average 401 days post operatively. Overall, 94% were satisfied with the procedure. For completely displaced clavicle fractures in adults, plating is a reliable procedure. PMID- 10707217 TI - The epidemiology of skiing injuries in Antarctica. AB - A retrospective analysis of all skiing injuries experienced by members of the British Antarctic Survey between 1989 and 1995 was undertaken to test the hypothesis that skiing was responsible for a disproportionate number and severity of injuries compared with other activities. Fifty-nine new consultations for skiing injuries were recorded. This represented 3.2% of all consultations (annual range 1.3-6.7%), or 9.7% of all consultations due to trauma. The mean incidence was 84.3/1000 population/year. The annual proportion and rate of consultation fluctuated but no overall trends were noted. The lower limb was the commonest site of injury (76.3%), with the ratio of lower limb: upper limb injuries being 6.4:1. The commonest single injury was an isolated medial collateral ligament knee sprain (23.7% of all consultations). Head injuries comprised 8.5% and ulnar collateral ligament thumb sprains 5.1%. Assessment of injury by the Injury Severity Score (ISS) showed that skiing injuries were significantly more likely to be non-trivial (ISS > 2) than work-related injuries [chi 2(1, N = 56) = 55.6, p < 0.001] or injuries of all causes [chi 2(1, N = 56) = 65.0, p < 0.001]. They were significantly more likely to need radiological investigation than all injuries [chi 2(1, N = 59) = 22.0, p < 0.001]. The most severe (ISS 13), survivable injury seen during the study period resulted from a skiing accident. This excess of non-trivial injury raises important management issues, particularly as the majority (81%) were recreational. PMID- 10707219 TI - Case of severe injury of lower limb treated with new Ljubljana traction method. PMID- 10707220 TI - Spinal epidural abscess: adding insult to injury? PMID- 10707221 TI - Toxic shock syndrome due to percutaneous Kirschner wires. PMID- 10707223 TI - The effect of subtle changes in self-tapping screw design. PMID- 10707222 TI - Idiopathic pneumoperitoneum following blunt chest trauma: a case report. PMID- 10707224 TI - Refractures of the radius and ulna in children. AB - Seven hundred and sixty-eight children with displaced forearm fractures requiring reduction were studied retrospectively. Of 38 refractures (incidence 4.9%), 34 occurred within nine months at the original fracture site. The median time to refracture was eight weeks after discontinuing cast immobilisation. Diaphyseal fractures were eight times more likely to refracture than metaphyseal fractures. The risk of refracture was inversely proportional to the duration of cast immobilisation. Cast immobilisation for a minimum of six weeks reduces the risk of refracture by a factor of between four and six. Midshaft forearm fractures are at risk of refracture for sixteen weeks from cast removal. PMID- 10707225 TI - Local plate infections in a rabbit model. AB - We used 30 New Zealand white rabbits to compare the susceptibility to bacterial challenge of two different orthopaedic implants: a standard-design stainless steel plate, or a PC-FIX titanium plate applied on the right tibia were compared with sham operated animals. Directly after surgery Staphylococcus aureus (10(8) 10(9) colony forming units) were inoculated close to the plate. The infection rate in the group of plated animals was 11/20 (stainless steel plates 6/10, PC FIX titanium plates 5/10) and in sham operated animals only 1/10. Thus, a foreign body increased the risk for infection (p = 0.02). However, the implant type did not appear to be of major importance when the bacteria were inoculated locally. In experimental haematogenous infections, by contrast, implant design and material are considered to be important. This may either indicate different pathogenic mechanisms in locally and haematogenously induced infections, or simply that the large number of microorganisms in local inoculation 'overload' the normal defence systems. PMID- 10707226 TI - The influence of screw omission on construction stiffness and bone surface strain in the application of bone plates to cadaveric bone. AB - Biological fracture repair is becoming an increasingly popular means of fracture fixation. This technique involves a reduction in the amount of surgical trauma thereby preserving vascular supply and soft-tissue integrity combined with the implantation of less hardware. The aim is the stimulation of callus formation as the means by which fracture union occurs. This paper describes the mechanical effect of the symmetrical omission of screws on construction stiffness and bone surface strain following the application of bone plates to cadaveric bone. The influence of the pattern of screw omission was evaluated in both intact and osteotomized bone specimens. The application of bone plates using certain patterns of screw omission did not significantly effect the stiffness of the final construction whilst inherent bone surface strain was increased. It was concluded that the omission of screws in certain defined patterns for a given plate-bone construction may meet the criteria for 'biological fixation'. There was no apparent deleterious effect on structural stiffness following the omission of 40% of the total screw complement from a plate-bone construction. PMID- 10707227 TI - The efficacy of serial physical examination in penetrating abdominal trauma. AB - Over a 10-year period we reviewed the records of 370 consecutive patients with potentially penetrating abdominal wounds (48 gunshot and 322 stab wounds). Selective non-operative management for abdominal stab wounds was introduced, guided by serial physical examination. In our study the terms therapeutic and non therapeutic laparotomies were used, the latter comprising negative as well as insignificant findings. Initially diagnostic peritoneal lavage and local wound exploration were used, but these methods were later abandoned. Mortality was 10.2% and morbidity 25% in the group of gunshot wounds and 1.2% and 8.6% in the group of stab wounds. During this period the rate of laparotomies for stab wounds decreased from 55% to 30%, while the rate of non-therapeutic laparotomies decreased from 24% to 0% in the last year. Delayed laparotomy did not cause death or increase morbidity. Our results support the concept of selective non-operative management of abdominal stab wounds using repeated physical examination. Peritoneal perforation and haemoperitoneum should not be an indication for routine laparotomy. PMID- 10707228 TI - The preoperative prevalence of deep vein thrombosis in patients with femoral neck fractures and delayed operation. AB - Out of 61 consecutive patients admitted for femoral neck fracture 21 patients had a delay to operation for more than 48 h from the time of injury. We studied these patients prospectively for the presence of deep-vein thrombosis (DVT). 13 (62%) had venographic evidence of thrombosis. All occurred in the broken limb. Five patients had bilateral thrombosis. The delay alone seems to be the major risk factor for thrombosis irrespective of age, fracture type, premorbid mobility and coexisting illness. The prevalence of preoperative DVT 48 h after injury approaches the reported postoperative incidence of DVT, which suggests that DVT will occur in a high proportion of patients regardless of treatment and prophylaxis. We recommend that those patients, in whom operation is delayed, should be routinely investigated for the presence of thrombosis preoperatively and a prophylactic vena cava filter should be considered when major deep vein thrombosis occurred. PMID- 10707229 TI - Reconstructive operations for the paralyzed shoulder in brachial plexus palsy: concept of treatment. AB - Sixty-three patients with persistent brachial plexus palsy underwent a transfer of the trapezius muscle and 14 patients a shoulder arthrodesis. Thirteen female and 64 male patients were treated with a mean age of 31 yr (17-69 yr). The average follow-up period was 14 months (6-50 months). In all cases, the trapezius transfer resulted in increased abduction from 6.1 degrees to an average of 36.4 degrees (20-80 degrees) and forward flexion from 13.8 degrees to an average of 31.9 degrees (10-90 degrees). The multidirectional shoulder instability was improved in 60 patients. Strength and functional improvement was, on average, greater following shoulder arthrodesis (abduction from 9.6 to 59.3 degrees (40-90 degrees), forward flexion from 11.4 to 50.7 degrees (30-90 degrees)). In patients with brachial plexus palsy, trapezius transfer resulted in an improvement of shoulder function and stability as well as subjectively. The increase in function was, however, less pronounced in comparison with shoulder arthrodesis. The advantages of the transfer are the regaining of normal passive function and the shorter duration of surgery. Shoulder fusion is more suitable for those patients who require the best possible extent of function and strength in the shoulder. PMID- 10707230 TI - Massive blood transfusion exceeding 50 units of plasma poor red cells or whole blood: the survival rate and the occurrence of leukopenia and acidosis. AB - The survival rate after bleeding requiring massive blood transfusions exceeding 50 units has been reported to be low or zero. There seems to be no reports of leukopenia in connection with massive blood transfusion. This retrospective study was carried out to investigate the survival rate and the occurrence of leukopenia and acidosis in patients who were transfused with more than 50 units of plasma poor red cells or whole blood. The survival rate was 16 of 23. Three of the five patients with a blood transfusion of over 100 units survived. Pure component therapy was used on 18 occasions. All patients had a leukopenia, which lasted up to five days. All patients had an acidosis. The range of the lowest pH values in patients who did not survive was from 6.77 to 7.27 and in survivors from 6.87 to 7.28. The survival rate was considerably higher than reported in previous studies. Pure component therapy appeared to be particularly suited to massive transfusion. Leukopenia was a regular phenomenon. Severe acidosis did not predict a poor outcome. PMID- 10707231 TI - Complete transposition of the radial nerve complicating an open fracture of the shaft of the humerus. PMID- 10707232 TI - Quadriplegia following grand mal seizures. PMID- 10707233 TI - The 'pin and plaster technique' for maintaining reduction after traumatic knee dislocations. PMID- 10707234 TI - A new technique of fixation of displaced proximal radial physeal fracture. PMID- 10707235 TI - Trans iliac-sacral-iliac bar stabilization to treat bilateral sacro-iliac joint disruptions. PMID- 10707236 TI - Flail penis: a complication of 'open-book' pelvic fracture. PMID- 10707237 TI - Horizontal fingertrap traction in distal radial fractures. PMID- 10707238 TI - Material and design in haematogenous implant-associated infections in a rabbit model. AB - We used 111 rabbits to study the susceptibility to intravenously injected bacteria of conventional stainless steel plates, and titanium plates of either traditional design or of the PC-FIX concept, that is less traumatic to bone. After plating, the animals were given between 1 x 10(8) and 2 x 10(9) colony forming units of Staphylococcus aureus Wood 46 intravenously. Significant differences in infection rates (positive cultures) were found between conventional stainless steel plates (36-67% infected, depending on inoculum size) and titanium PC-FIX plates (6-7% infected). In fact, the infection rate at the PC FIX plate did not differ from sham operated animals. Since conventionally designed titanium plates had an intermediate infection rate, it appears that design and material both are important for the risk of infection. PMID- 10707239 TI - A stab in the dark! Are you ready to perform needle cricothyroidotomy? AB - The objective of this study was to assess the availability of pre-prepared equipment for needle cricothyroidotomy, and the knowledge of staff in its use in Accident and Emergency (A&E) departments in Great Britain. A telephone survey was undertaken of all A&E departments seeing more than 30,000 new patients per year. 184 hospitals were contacted. 98% of the doctors agreed to be interviewed. 47% of the departments had made provision for immediate use of needle cricothyroidotomy. 45% of the doctors interviewed were fully conversant in the use of needle cricothyroidotomy. Provision of equipment for immediate use of needle cricothyroidotomy in A&E departments is generally inadequate. All departments should ensure that such equipment is immediately accessible, and that the staff is regularly trained in its use. PMID- 10707240 TI - Does Kapandji wiring help in older patients? A retrospective comparative review of displaced intra-articular distal radial fractures in patients over 55 years. AB - Forty-six patients aged 55-90 with intra-articular displaced fractures of the distal radius were reviewed retrospectively. All patients were treated with either manipulation and plaster of Paris or Kapandji wiring. Radiographic and functional review was performed by an independent observer a mean of 17 months after the fracture. The results showed superior anatomical and functional results in the group treated with Kapandji wiring. The mean dorsal angle was significantly better in the wired group, and the improvement in dorsal angle, radial angle and radial length from presentation to final result was also significantly better. Functional results were excellent or good in 19/23 of the wired group, compared with 12/23 of the plaster group. There was a strong correlation between functional outcome and both dorsal angle and radial length at union. These results support the use of this method of wire fixation in older patients, as the technique is simple and complications were few. PMID- 10707241 TI - Cemented versus uncemented Thompson's prostheses: a functional outcome study. AB - One hundred and one elderly patients (90 female and 11 male) with a mean age of 83.5 years (range 69-100) were treated for intracapsular femoral neck fracture by Thompson's prosthesis. The prosthesis was fixed in the femoral shaft using Palacos cement in 23% and was inserted uncemented in 77%. We compared the following pre and postoperative variables in each group; mobility of the patient, their activity, walking aids and postoperative thigh pain. We also studied the pre and postoperative hip X-rays. Our patients did well in both groups. We conclude that there is no statistically significant difference between the variables in the two groups. Thompson's prosthesis can be inserted uncemented. Patients with radiological loosening of the prosthesis are not necessarily symptomatic. Intra and postoperative complications were similar to other published series of hip prostheses. The femoral neck osteotomy for Thompson's prosthesis can be performed safely well above the calcar femoris without detrimental complications. The original inter-trochanteric cut Thompson described is not necessary for acute fractures. This consequently makes revision of failed Thompson's prosthesis easier. PMID- 10707242 TI - Identification of patients to include in trauma audit: a modification of the pre chart. AB - Trauma audit is commonly focused using a Pre-Chart to illustrate calculations made using the TRISS model. A line is drawn at Ps = 0.5 to divide expected survivors and nonsurvivors. The use of this cut-off in a severely injured population was examined. The 'M statistic' for a group of injured patients selectively triaged to a Trauma Centre was calculated. The ideal cut-off point between predicted outcomes when using the TRISS model to focus trauma audit in this population was determined using a Receiver Operating Characteristic (ROC) curve. For this population the TRISS 'M statistic' was 0.71 (indicating a significantly different case mix from the reference database) and the best cut off point was at Ps = 0.76. Trauma audit in populations with a case mix of injury severity different from the reference database should use a different Ps line to define unexpected outcomes. PMID- 10707243 TI - Semi-quantitative assessment of tibial blood flow and distribution in response to surgical intervention using first pass radionuclide angiography and intravascular vital dye. AB - The acute vascular response in bone to surgical trauma was investigated utilizing a sheep model. Blood flow and distribution were determined using two methods; perfusion of the vasculature with an intravascular vital dye (Disulphine blue) prior to euthanasia and by radionuclide angiography (RNA) before and after each surgical intervention. The pattern of Disulphine blue distribution provided a good indication of local perfusion and response to surgical trauma (drilling holes). Radionuclide angiography provided a dynamic image of the vascular response to surgical trauma. The generation of time activity curves of the first pass of radionuclide bolus enabled calculation of the relative blood flow through selected regions. For both techniques areas of ischaemia were apparent which were directly related to the location of screw holes. We conclude that factors other than bone plate contact influence the ischaemia that develops in bone subsequent to the application of bone plates. PMID- 10707244 TI - Peri operative ultrasound guided needle localisation of amputation stump neuroma. AB - Ten patients with clinically suspected neuromas following amputation were submitted for ultrasound examination. Neuromas in seven of the ten patients were identified as the cause of pain with ultrasound guided infiltration of local anaesthetic. Pre operative localisation, using breast localisation wires allowed the neuromas to be surgically excised with reduced dissection. This small study demonstrates that ultrasound is an effective method for identifying and localising neuromas in amputation stumps. This leads to reduced dissection and may lead to a better outcome in these patients. PMID- 10707245 TI - A safe surgical technique for the partial resection of the ruptured spleen. A clinical report. AB - A consecutive series of 11 patients with an acute blunt splenic injury were treated with a 'safe resection' technique. 57% of the injured spleens (range 35 100%) were saved. None of the patients had any signs of secondary bleeding in control CT scan and the mortality was zero. No second-look laparotomies were performed. Follow-up time was at least two months (range 2 month-6 yr). Operation time was in average 120 min. Total mean peroperative bleeding was 1400 ml. Partial resection may offer patient a change for normal function of the injured spleen. However, it is not yet known what is the critical mass of spleen tissue needed for humans. The follow-up time of the present study is still too short to estimate this fact, but further studies may show the benefit of the present method in avoiding serious long term immunological complications of splenectomy. This present study introduces a novel technique for partial resection of injured spleen. Operation can be performed safely and quickly with a complication risk comparable to splenectomy. Resection is applicable even for multi-trauma patients. PMID- 10707246 TI - Routine radiography following ankle fracture fixation: a case for limiting its use. AB - The use of fluoroscopic screening in the orthopaedic theatre is a necessary operative aid in many procedures. Modern systems give good image resolution and allow the production of per-operative hard-copy prints. This study was performed to compare these prints with postoperative radiographs in 41 patients who underwent internal fixation for an ankle fracture in a 6-month period. The hard copy prints and the postoperative X-ray films were independently assessed for several features, which included status of the tibiofibular syndesmosis, fibular length, talo-crural angle, talar tilt, presence and size of a posterior malleolar fracture, and abnormality of the medial clear space. Information was also recorded as to whether there had been a change in postoperative management plan after review of the check X-ray. Of the 41 cases, 30 were suitable for full assessment. In 25 of these cases there was no difference in the information provided by the hard-copy prints from fluoroscopic images and the postoperative check X-rays. In the other five cases, the differences were not significant. In none of the cases did the check X-ray effect a change in postoperative management. We therefore suggest that if per-operative hard-copy prints are obtained from the fluoroscopic images, postoperative radiographs of the ankle are only necessary in exceptional circumstances. PMID- 10707247 TI - The outcome following major trauma in the elderly. Predictors of survival. AB - OBJECTIVES: To assess the reliability of the predicted probability of survival calculated using TRISS methodology by the UK Trauma Network for elderly patients. METHOD: Analysis of 100 consecutive trauma patients 65 years and over, prospectively entered into the UK Trauma Network database from a single centre. The probability of survival (Ps) was calculated from the UK Trauma database and retrospectively related to survival, premorbid medical condition and mobility. RESULTS: Of 100 patients, 16 died and 84 survived. Eleven of the 16 who died and 12 of the survivors had pre-existing medical disease (ASA grade III-V) and social dependency suggesting a poor outcome, these factors being significantly associated with mortality (P < 0.005). The mean Ps for the 11 with severe medical disease who died was 0.85 (+/- 0.07) with a mean age 85 (+/- 3.5). The remaining five patients who died suffered high energy injuries, had a mean age of 70 (+/- 4.8) and a low probability of survival (Ps 0.40 +/- 0.24). The median pre-injury mobility score was 8 in patients who survived and 4.5 in those who died. Mobility score < 5 was associated with an increased mortality following admission from Trauma (P < 0.05). CONCLUSIONS: There is a significant association between severe preexisting medical disease (ASA III-V) and death during admission for trauma. The Ps score is unrealistically high in this group of patients. A simple mobility score correlates well with outcome in this group. PMID- 10707248 TI - Complex pelvic trauma with extensive gluteal skin loss treated by suspension traction technique. PMID- 10707249 TI - Extensive bone loss in an open tibial shaft fracture (immediate bone boiling reimplantation). PMID- 10707250 TI - Disabling leg length inequality following overhead extension in an infant. PMID- 10707251 TI - Olecranon traction using a recycled plate: a new technique for supracondylar humeral fractures. PMID- 10707252 TI - Axial dynamisation cannot be a static process. PMID- 10707254 TI - Dynamic axial fixation is a cyclic process of loading and unloading PMID- 10707253 TI - A response to "Axial dynamisation cannot be a static process" by Mr John C.R. Scott FRCS, Orthofix Group Medical Advisor PMID- 10707255 TI - Multiplex PCR assay to aid in the identification of the highly transmissible Mycobacterium tuberculosis strain CDC1551. AB - SETTING: Mycobacterium tuberculosis strain CDC1551 outbreak area in Tennessee and Kentucky and selected locations in the USA. OBJECTIVE: Develop a PCR assay to distinguish the highly transmissible CDC1551 from strains which have similar 4 band IS6110 fingerprints. DESIGN: Compare the IS6110 insertion sites in CDC1551 with those in 10 isolates which have similar 4-band IS6110 fingerprints. Utilize unique characteristics of insertion sites in CDC1551 to design a multiplex PCR to identify this strain. RESULTS: A multiplex PCR was developed which targets an IS6110 insertion conserved in most IS6110 low copy number strains and a deletion within the direct repeat region adjacent to an IS6110 insertion. Of 139 isolates with similar 4-band fingerprints, the CDC1551 PCR pattern was generated by only the 14 outbreak associated isolates. Of 154 isolates with different fingerprints, only four generated the CDC1551 pattern and these could be distinguished from CDC1551 by their IS6110 fingerprint. CONCLUSIONS: The multiplex PCR used in conjunction with the IS6110 fingerprint should be a useful tool to aid in the continued surveillance of the outbreak area and follow the spread of this highly transmissible strain of M. tuberculosis. PMID- 10707256 TI - Increased TNF-alpha, IL-1 beta and IL-6 levels in the bronchoalveolar lavage fluid with the upregulation of their mRNA in macrophages lavaged from patients with active pulmonary tuberculosis. AB - SETTING: We hypothesized that patients with active pulmonary tuberculosis (TB) have tubercular pneumonitis and that alveolar macrophages at these sites release proinflammatory cytokines, resulting in high levels of cytokines in alveolar epithelial lining fluid. OBJECTIVE: To measure cytokine levels in bronchoalveolar lavage fluid (BALF) and to confirm the source of any cytokines by examination of alveolar macrophage cytokine mRNA. DESIGN: Seventeen active pulmonary TB patients and 15 healthy controls were prospectively studied. Bronchoalveolar lavage (BAL) was performed, proinflammatory cytokine levels were determined and alveolar macrophages isolated from BALF were prepared for RNA extraction and Northern blot analysis. RESULTS: Compared with healthy controls, TNF-alpha, IL-1 beta and IL-6 in BALF were all significantly higher in patients with active pulmonary TB, 298.7 +/- 85.9 vs. 8.9 +/- 2.7 (P = 0.0001); 164.4 +/- 67.5 vs. 8.9 +/- 2.7 (P = 0.003); 969.2 +/- 214.2 vs. 86.4 +/- 17.0 (mean +/- SE pg/ml) (P = 0.0001), respectively. Only TNF-alpha and IL-6 levels were significantly higher in sera of active pulmonary TB patients, 92.3 +/- 28.7 vs. 3.5 +/- 1.2; 15.2 +/- 5.4 vs. 2.1 +/- 2.1, respectively. Northern blot analysis revealed increased gene expression of these alveolar macrophage cytokines in patients with active pulmonary TB compared healthy controls. CONCLUSION: Significantly higher levels of TNF-alpha, IL-1 beta and IL-6 were found in BALF from patients with active pulmonary TB, and were released by alveolar macrophages in the TB lesions. PMID- 10707257 TI - Identification and characterization of two divergently transcribed iron regulated genes in Mycobacterium tuberculosis. AB - SETTING: Low iron availability in the host induces the expression of iron acquisition systems and virulence genes in many pathogens. IdeR is a mycobacterial iron dependent regulator that controls the iron starvation and oxidative stress responses in Mycobacterium smegmatis. It is important to determine the role of IdeR and its regulon in M. tuberculosis, as identification of iron regulated genes can aid in the design of new drugs and generation of attenuated strains. OBJECTIVE: A potential IdeR binding site was found in the M. tuberculosis genome flanked by two divergently oriented open reading frames, irg1 and irg2. The aim of this study was to determine whether irg1 and irg2 were iron and IdeR regulated genes. DESIGN: Interaction of IdeR with the putative binding sequence was examined by gel shift and footprinting assays. Transcriptional fusions of irg1 and irg2 to IacZ were used to study the effect of iron levels on the expression of these genes. RESULTS: IdeR binds to the predicted binding site, which overlaps with the irg1 promoter. irg1 and irg2 expression was decreased by iron in M. tuberculosis and in wild type M. smegmatis, but not in a M. smegmatis ideR mutant. CONCLUSION: Two M. tuberculosis iron/IdeR regulated genes were identified. irg1 is predicted to be the M. tuberculosis hisE gene, which is involved in histidine biosynthesis. It is directly upstream of the M. tuberculosis hisG. irg2 encodes a putative membrane protein that is a member of the PPE family. PMID- 10707258 TI - Characterization of a Mycobacterium tuberculosis homologue of the Streptomyces coelicolor whiB gene. AB - SETTING: Molecular Research Laboratory, Department of Medical Microbiology, University of Cape Town and Groote Schuur Hospital. OBJECTIVE: Characterize Mycobacterium tuberculosis homologue of the Streptomyces coelicolor, sporulation specific, whiB regulatory gene. DESIGN: The M. tuberculosis whiB3 gene was isolated by enriched cloning of a 2.8 kb BamHl fragment to which the S. coelicolor whiB gene hybridized. Expression of the gene was analysed by S1 nuclease analysis and promoter studies. RESULTS: An open reading frame within the 2.8 kb BamHl fragment was identified as the M. tuberculosis whiB3 gene, one of four whiB homologues in the M. tuberculosis genome. The deduced amino acid sequence has a 92% identity with a M. leprae protein, and 32% identity with the S. coelicolor WhiB protein. S1 nuclease analysis showed that the M. tuberculosis whiB3 gene is constitutively expressed by the cells in liquid culture. Primer extension analysis revealed three transcriptional start sites. Expression from the three potential promoters is growth phase-dependent. CONCLUSION: The M. tuberculosis whiB3 gene is expressed throughout growth, but expression from the individual promoters is growth phase dependent. PMID- 10707259 TI - Associations of HLA-DRB1, DQB1 and DPB1 alleles with pulmonary tuberculosis in south India. AB - SETTING: Tuberculosis is endemic in south India: sputum positive pulmonary tuberculosis is predisposed by HLA-DR2 in south India and few other populations of the world. OBJECTIVE: To study HLA-DRB1, DQB1, DQA1 and DPB1 allelic polymorphism in pulmonary tuberculosis patients and endemic controls from south India. DESIGN: One hundred and twenty-six, sputum positive pulmonary tuberculosis patients and 87, endemic controls, from Madurai were studied for MHC class II allelic polymorphism by PCR-SSOP method. XI IHWC primers and probes and non radioactive probing methods were employed. RESULTS: HLA DRB1*1501 and DQB1*0601 predisposed for pulmonary tuberculosis (DRB1*1501: odds ratio (OR) = 2.68, 95% confidence interval (CI) = 1.30-5.89, P value (P) = 0.013, aetiological fraction (EF) = 0.17; DQB1*0601: OR = 2.32, CI = 1.29-4.27, P = 0.008, EF = 0.26). Haplotype DRB1*1501-DQB1*0601 was higher in patients (1324 per 10,000, X2 = 27.07) than controls (F = 404/10,000, X2 = 8.84). In a subset of 63 caste matched samples, DPB1*04 was preventive (OR = 0.45, CI = 0.21-0.95, P = 0.036, PF = 0.26): the distributions of DRB1*1501-DQB1*0601-DPB1*04 phenotypes were different between patients and controls (P = 0.0092). These alleles were predominant in patients and controls of T5SU caste. CONCLUSION: HLA-DRB1*1501 and DQB1*0601 predisposed to sputum positive pulmonary tuberculosis, and DPB1*04 was preventive and epistatic to this risk. Caste T5SU is an ideal model to study immunology of tuberculosis. PMID- 10707260 TI - Antituberculosis activity of certain antifungal and antihelmintic drugs. AB - A panel of antifungal and antihelmintic drugs was tested for activity against Mycobacterium tuberculosis in vitro. Antifungal drugs, miconazole, 2 nitroimidazole, clotrimazole, and the antihelmintic drug niclosamide were found to have significant antituberculosis activity, with minimal inhibitory concentrations (MICs) between 1 and 10 micrograms/ml. Niclosamide and 2 nitroimidazole also had activity against stationary phase tubercle bacilli. Further testing of these drugs and their derivatives in vitro and in vivo appears to be warranted. PMID- 10707262 TI - Caring with the simplest acts. PMID- 10707263 TI - Affective competencies. PMID- 10707261 TI - Effects of iron deprivation on Mycobacterium avium growth. AB - SETTING: Acquired immune deficiency syndrome (AIDS) patients have increased iron deposition in different tissues which may favour the growth of Mycobacterium avium, a common bacterial opportunist in these patients. OBJECTIVE: To test whether reducing the iron loads in macrophages in vitro and in vivo reduces M. avium proliferation. DESIGN: Mycobacterial proliferation was evaluated in vitro either in axenic media or cultured macrophages and in vivo in mice after manipulation of the iron status. RESULTS: Three different compounds- desferrioxamine (DFO), N,N'bis(2-hydroxybenzyl)ethylenediamine-N,N'-diacetic acid (HBED) and a 1-amino-3-(2-bipyridyl)isoquinoline derivative (VUF8514)--were found to inhibit the growth of M. avium in axenic medium. DFO and HBED were also active in inhibiting the intramacrophagic growth of M. avium, while the use of VUF8514 was prevented by its toxicity towards the host cell. Both DFO and HBED enhanced the mycobacteriostatic effect induced in bone marrow derived macrophages by interferon gamma. In vivo, an iron poor diet led to reduced M. avium proliferation whereas the intraperitoneal administration of either DFO or HBED had small effects as they impacted little on the iron status of mice. CONCLUSION: These results confirm that iron withholding is a means of inhibiting the growth of M. avium. In vitro data suggest that iron chelating compounds may be useful as adjunct therapy against M. avium, once their in vivo activity is optimized. PMID- 10707264 TI - Preparing children for surgery--an integrative research review. AB - The purpose of this integrative research review was to determine the methodological and substantive characteristics of the research pertaining to preparation of children for surgery, examine pertinent research issues, and suggest recommendations for future research and practice. A total of 400 articles were reviewed using inclusion and exclusion criteria. Twenty-two studies, published between 1974 and 1995, met the criteria and, therefore, constituted the study corpus. The total sample comprised 1,263 subjects. Findings indicate that further research is needed to examine the efficacy of preparation strategies, address diverse populations, and include patient and parent involvement in the preparation process. PMID- 10707265 TI - Perioperative data elements: interventions and outcomes. AB - In 1999, the American Nurses Association's committee on nursing practice recognized AORN's Perioperative Nursing Data Set (PNDS) as a data set useful in the practice of nursing. The PNDS is a standardized nursing language specific to perioperative settings that is both clinically relevant and empirically validated. This data set is the culmination of six years of effort by AORN and AORN staff members to develop a standardized language that is clearly defined, common to all cases, and consistent across time. PMID- 10707266 TI - TRAM flap breast reconstruction with expanders and implants. AB - Breast reconstruction has become an essential part of breast cancer management. The transverse rectus abdominis myocutaneous (TRAM) flap has replaced the prosthetic implant as the first choice for breast reconstruction. There are several incidences in which the volume of autogenous tissue cannot fulfill the requirements for symmetric reconstruction, especially in those patients with limited abdominal tissue and large ptotic breasts. The TRAM flap can be combined with tissue expanders and implants to obtain symmetry in these difficult reconstructive situations. PMID- 10707267 TI - Body contouring with ultrasound-assisted lipoplasty. AB - Lipoplasty (i.e., liposuction) is the most commonly performed cosmetic surgical procedure in the United States. During the past 20 years, there have been dramatic technological advancements in both surgical technique and equipment used to improve postoperative outcomes. As a result, perioperative nurses are faced with the challenge of providing quality care for this patient population in a variety of ambulatory surgical settings with highly specialized equipment. Perioperative team members play significant roles in the coordination of individualized patient-focused care, which includes detailed instructions and education, appropriate use and care of equipment, and postoperative follow-up. The efforts of the team members contribute to positive surgical outcomes, as well as to overall patient satisfaction. PMID- 10707268 TI - Internet resources for reconstructive breast surgery. PMID- 10707269 TI - Clinical expertise and research findings--understanding the fit. AB - Clinical expertise is as valuable as the information that supports decisions. The worth of that information must be evaluated carefully and constantly questioned. Clinicians must work together to carefully and systematically analyze what they bring to the decision-making process. Rules that serve as the foundation of one's decision need to be evaluated continuously. As well, every clinician must be aware of inherent and subtle biases that influence decisions. Relying on clinical expertise can be a helpful strategy, but clinicians must proceed with caution. As with research, nothing is correct 100% of the time--even experts make mistakes. Use your expertise to determine what strategy is correct when facing a clinical problem. If your clinical expertise will serve the patient best, do not be afraid to use this hard-earned knowledge for the optimum patient outcome. PMID- 10707270 TI - AORN set for certified RN first assistant Medicare reimbursement in the new year. AB - As a provider of health care, the CRNFA is a viable solution for controlling rising health care costs. Working in collaborative practice with surgeons, CRNFAs are cost-effective for the patient and for the health care delivery system. The AORN proposal would extend Medicare coverage eligibility to CRNFAs for their surgical first assisting services. Use of CRNFAs could well decrease the frequency and length of hospital stays because CRNFAs would be reimbursed under Medicare at a lower rate than physicians who first assist and because CRNFAs routinely provide much-needed patient education and counseling. PMID- 10707271 TI - [The 275th anniversary of Russian Academy of Sciences and history of biophysics]. AB - Within the framework of scientific arrangements devoted to the 275th Anniversary of the Russian Academy of Sciences, the Second Congress of Biophysicists of Russia was held in Moscow from 23 to 27 August 1999 on the basis of the Lomonosov State University. More than 600 scientists from Russia and the Commonwealth of Independent States took part in the Congress. Below we present abridged a report of the Corresponding Member of the Russian Academy of Sciences Prof. G.R. Ivanitskii. This report opened the Congress. PMID- 10707272 TI - [When and how can homologs overcome errors in the energy estimates and make the 3D structure prediction possible]. AB - One still cannot predict the 3D fold of a protein from its amino acid sequence, mainly because of errors in the energy estimates underlying the prediction. However, a recently developed theory [1] shows that having a set of homologs (i.e., the chains with equal, in despite of numerous mutations, 3D folds) one can average the potential of each interaction over the homologs and thus predict the common 3D fold of protein family even when a correct fold prediction for an individual sequence is impossible because the energies are known only approximately. This theoretical conclusion has been verified by simulation of the energy spectra of simplified models of protein chains [2], and the further investigation of these simplified models shows that their true "native" fold can be found by folding of the chain where each interaction potential is averaged over the homologs. In conclusion, the applicability of the "homolog-averaging" approach is tested by recognition of real protein 3D structures. Both the gapless threading of sequences onto the known protein folds [3] and the more practically important gapped threading (which allows to consider not only the known 3D structures, but the more or less similar to them folds as well) shows a significant increase in selectivity of the native chain fold recognition. PMID- 10707273 TI - [Adsorption of dimethylsulfoxide on proteins]. AB - A method for measuring the adsorption of dimethyl sulfoxide on native and denatured trypsin and albumin was developed. On native proteins, no positive adsorption was registered, and a slight negative adsorption within the limits of experimental error was observed. It was shown that the properties of denatured proteins depend on the mode and conditions of denaturation. On one of denatured trypsin specimens, positive adsorption of dimethyl sulfoxide was registered, on other specimens no adsorption was observed. The reason for this behavior lies in the hydrophobic nature of adsorption of dimethyl sulfoxide at the interface, while the surface of native protein globules and, probably, most denatured protein specimens is hydrophilic. PMID- 10707274 TI - [Analysis of low temperature magnetic circular dichroism of high spin ferrihemoproteids in the near UV region]. AB - Magnetic circular dichroism spectra of fluoride complexes of metmyoglobin, methemoglobin, and horseradish peroxidase in the region of 300-450 nm at temperatures from 300 to 2.1 K were measured and analyzed. The temperature dependence of magnetic circular dichroism in the Soret region was found to be different from that of other paramagnetic forms and from the theoretically predicted dependence. The difference is explained by the superposition of the pi- >pi*-transition of porphyrin with one (peroxidase) or two charge transfer transitions and by substantially different temperature dependences of magnetic circular dichroism for the transitions of the two types. By minimization of differences between the expected and observed temperature dependences of magnetic circular dichroism, the parameters of its temperature dependence for charge transfer transitions and the parameter D of the zero-field splitting of the electronic ground state of the heme were found. The values of D for the fluoride complexes of metmyoglobin (5.8 cm-1) and methemoglobin (6.1 cm-1) agree well with those obtained by other methods. The D value for the fluoride complex of horseradish peroxidase (8.8 cm-1) was determined for the first time. PMID- 10707275 TI - [Phosphorescent analysis of the intramolecular dynamics of the muscle glycogen phosphorylase b]. AB - Large-scale functionally significant changes in the intramolecular dynamics of muscle glycogen phosphorylase b (EC 2.4.1.1) in solution upon ligand binding, transition from dimeric to tetrameric form, temperature denaturation and aggregation were registered at room temperature using the tryptophan phosphorescence technique. It was shown that binding of glucose-1-phosphate (substrate, 0.25-4 mM) and glucose (competitive inhibitor, 0.5-8 mM) to the active site and temperature-induced protein aggregation decrease large-scale structural fluctuations of the protein matrix at the level of domains and subunits; whereas the transition of glycogen phosphorylase b to tetrameric form (R-conformation) leads to a dramatic increase in the structural flexibility of the peripheral parts of the protein globule. PMID- 10707276 TI - [Analysis of major antigenic determinants in Mistletoe lectin I]. AB - Antigenic determinants of Mistletoe Lectin I, a toxin from Viscum album were predicted on the basis of the primary amino acid sequence of the protein. Based on the results of analysis, the peptide FPGGSTRTQARS, which corresponds to the 144-155 segment of the viscumin A-chain, was synthesized. The peptide was tested in enzyme-linked immunosorbent assay with monoclonal antibodies against the viscumin A-chain obtained previously. The peptide reacted with antibodies with a low affinity and did not inhibit the binding of viscumin molecule to any of the antibodies. Analysis of the peptide by 1H-NMR spectroscopy in aqueous solution was performed. The three-dimensional structure of the 144-155 segment in the native protein globule was shown. PMID- 10707277 TI - [Probabilistic description of the system "ligand-receptor"]. AB - The theory of probabilities was used to describe the ligand-receptor interaction. Mean and variance of number ligand-receptor complexes are calculated. It is shown that the mean number of ligand-receptor complexes coincides with that obtained from the law of conservation masses. Proceeding from a ratio of mean and expectation it is shown that the variance of the number of ligand-receptor complexes should be taken into account with concentration of ligand-receptor complexes component less than 1 fmol. PMID- 10707278 TI - [Luminescence of thymine aqueous solutions at the room temperature]. AB - The distinguishing features of luminescence of aqueous thymine solutions at room temperature (a broadening of luminescence spectra as compared with low temperature spectra and differences between excitation and absorption spectra) are discussed. It is shown that these features are due to bimolecular photochemical reactions that lead to the formation of photoproducts (at the first stages, photoadducts) with a comparatively high luminescence ability. PMID- 10707279 TI - [Changes in the model membrane structure induced by ribonuclease and lysozyme studied by the fluorescent probe technique]. AB - Using fluorescent probes DSM and DSP-12, the effect of ribonuclease and lysozyme on the structural state of liposomes composed of phosphatidylcholine and diphosphatidylglycerol was studied. A correlation between the changes in probe quantum yield and the amount of protein-bound lipids was established. It is assumed that the formation of protein-lipid complexes increases the packing density of lipids and restricts their mobility. As the content of diphosphatidylglycerol in the lipid bilayer increases, the condensing effect of proteins becomes more pronounced. PMID- 10707280 TI - [Effect of mitochondria on the redox reaction between oxyhemoglobin and ferricytochrome c]. AB - The effect of mitochondria on the redox reaction between oxymyoglobin (oxy-Mb) and ferricytochrome c was studied. The parameters of this reaction in the absence of mitochondria have been investigated earlier. It is shown that the course of oxidation of oxymyoglobin by cytochrome c in the presence of mitochondria differs from that without mitochondria: no reduced cytochrome c is observed; in addition, the order of this redox reaction and its dependence on pH and ionic strength change. The factors influencing the state of mitochondrial membrane and uncouples enhance markedly the reaction rate. The conclusion was drawn that mitochondria directly participate in the oxymyoglobin-cytochrome c redox reaction. The possibility of this reaction in vivo under extreme conditions and during pathological processes is discussed. PMID- 10707281 TI - [Determination of the functional state of cell population: lymphocytes]. AB - A mathematical model is proposed, which describes the relationship between the spectrum of cell sizes in the population and its functional state. The size spectra for lymphocytes both before and after the influence of gene modulator on these cells were quantitatively described. PMID- 10707282 TI - [Damage to erythrocytes caused by the interaction of nitrite-ions with hemoglobin]. AB - The formation of two hemoglobin forms (methemoglobin and nitrite methemoglobin) in native human erythrocytes in the presence of sodium nitrite in suspension was shown. In normal erythrocytes, the interaction of intracellular oxyhemoglobin with nitrite ions results in the formation of methemoglobin, whereas in metabolically exhausted erythrocytes, this leads predominantly to the formation of nitrite methemoglobin. The nitrite methemoglobin reacts with hydrogen peroxide to form reactive intermediates (e.g. peroxynitrous acid) and the products of hemoglobin destruction. During the storage of erythrocyte suspensions containing methemoglobin and modified nitrite methemoglobin, differences in the forms of erythrocytes and the degree of their hemolysis were revealed. It is assumed that the formation of methemoglobin leads to the destruction of erythrocytes. PMID- 10707283 TI - [Resonance interactions of surface charged lipid vesicles with the microwave electromagnetic field]. AB - The occurrence of collective excitations in an ionic medium on the surface of lipid membranes was shown. The excitations are due to a fast lateral mobility of ions and the excitation of high-frequency displacement currents in the Stern's layer at the charged surface of the membrane. These effects determine the mechanism of induction of resonant dipole moments on lipid vesicles which can underlie the effect of "recognition" and the autooscillation mode of aggegation of vesicles in a colloidal solution. PMID- 10707284 TI - [Comparative study of the crystallin supramolecular structure in the carp, frog, and rat lenses by small-angle roentgen ray scattering]. AB - The supramolecular structure of crystallins in intact ocular lenses of carp, frog and rat as well as in the interior (nuclear) and outer (cortical) parts of these lenses was studied by the small-angle X-ray scattering method. The results show that the supramolecular structure of crystallins substantially varies both in lenses of different vertebrate species and in various parts of the same lens. In carp lens and in the cortical part of rat lens, crystallins have an ordered supramolecular structure, as indicated by a small-angle X-ray diffraction maximum in the region of Bragg distances 15-20 nm, whereas in frog lens and in the nuclear part of rat lens, the supramolecular structure of these proteins is disordered. The power-law X-ray scattering by rat lens nucleus may be evidence of fractal structures in the lens. A comparison of these results with literary data indicates that there is no obvious correlation between the type of supramolecular structure of crystallins and their polypeptide composition in lenses of different vertebrate species. The results suggest that the supramolecular ordering (short range order) of crystallins is not a necessary condition for lens transparency. PMID- 10707285 TI - [Activity of the climbing fiber innervating various Purkinje cells]. AB - Low-amplitude potentials (10-130 microV) related to the action of a distant branch of the climbing fiber, which elicits complex spikes of the reference Purkinje cell were revealed by means of potential averaging synchronously with complex spikes of Purkinje cells in 10 out of 255 paired records of cerebellar Purkinje cells activity and extracellular field potentials at interelectrode distances of 200-1500 microns. These potential waves had a stable form in independent sets of data. In 3 out of 10 cases, the low-amplitude potentials included a slow (about 100 ms in duration) component. In one case, both test and reference electrodes recorded both simple and complex spikes of different Purkinje cells so that complex spikes of both cells were practically synchronous (conditional probability of complex spikes p = 0.97, onset time difference 0.54 ms). Thus for the first time in cerebellar physiology both simple and complex spikes activity of two Purkinje cells controlled by the same climbing fiber was recorded. PMID- 10707286 TI - [Sound reception in marine mammals depending on sound parameters and conduction pathways]. AB - The interaction of complex sounds with the body tissues of Black Sea dolphin (Tursiops truncatus) was studied by the method of instrumental conditioned reflexes with food reinforcement. The thresholds of detecting underwater acoustic signals of different frequencies for dolphin and northern fur seal (Callorhinus ursinus) were measured as a function of pulse duration under conditions of full and partial (head above water) submergence of animals into water. It was found that sound conduction through dolphin tissues was more effective than that in a northern fur seal in a wide frequency range. Presumably, the process of sound propagation in dolphin is accompanied by changes in the amplitude-frequency structure of broad-band sounds. The temporal summation in dolphin hearing was observed at all frequencies under conditions of full and partial submergence, whereas in northern fur seal it was nearly absent at a frequency of 5 kHz under the conditions of head lifting above water. PMID- 10707287 TI - [Fractal diagnostics of disturbances in the alpha-rhythm dynamics in patients with epilepsy]. AB - A new method for analyzing the chaotic component of EEG is proposed. The method is based on estimating the fractal dimension of fluctuations of alpha-rhythm power (the square of amplitude). It is shown that the dimensions of the background EEG fragments for epilepsy patients is significantly higher than that in norm, indicating a disbalance of cerebral mechanisms that control the alpha activity in this disease. A tendency toward the disturbance of the normal fractal structure of EEG in a group of patients with initial signs of epilepsy was revealed. This suggests that the method is of considerable promise for setting the individual long-term prognosis of the development of the epileptic syndrome. PMID- 10707288 TI - [Electrotonic conduction of excitation and inhibition along the dendritic fiber with the uniformly distributed synaptic conductance]. AB - The existence of an optimum radius of fibre is shown, when the physical parameters of the intracellular medium and the nonexcitable dendritic membrane are known and the length and load resistance are fixed. This provides the maximum potential for the proximal end of the fibre if synaptic conductance is distributed uniformly along the fibre. A formula for calculating the soma potential of the whole neuron is proposed. The optimal ratio l/lambda is 0.9193 ... if the volume of the fibre and synaptic conductivity are fixed. PMID- 10707289 TI - [Interrelation between arterial pressure and speed of the pulse wave spreading]. AB - To describe the dependence of arterial pressure on the speed of spreading of pulse wave, an curvilinear regression equation with two constants was proposed. The causes of the discrepancy in the dependences reported in literature are discussed. PMID- 10707290 TI - [An elementary bioelectrical generator of the anisotropic homogeneous myocardium and its extracellular field]. AB - On the basis of the bidomain model that takes into account the electric anisotropy of body tissues, analytical relationships were developed for calculating the characteristics of electric and magnetic fields produced by an elementary (dipole) bioelectric generator that arises in the electrogenic excitable tissue of the myocardium. The errors in the identification of intensity and location of the bioelectric generator in the myocardium were estimated from the measurements of its external fields (noninvasive identification of the excited region) using approximate methods based on isotropic models of the physical medium. PMID- 10707291 TI - [Atrial fibrillation as the nonstationary process: analysis of chaotic dynamics]. AB - A method for the mathematical analysis of recordings of the myocardium electrical activity during ventricular fibrillation is proposed. The method is based on evaluating the indices of the orderliness of the process. The entropy was one of the indices used. The analysis of EGG during ventricular fibrillation (an acute experiment on rats) revealed certain regularities in the apparently chaotic process of fibrillar oscillations. The entropy increased at the first stages of the fibrillation and then reached its maximal level. A gradual decrease in the entropy level preceded the spontaneous termination of fibrillation. This indicates that different stages of the process the degree of its disorganization changes. Upon spontaneous termination of fibrillation, quantitative changes transfer to qualitative changes. This reflects the transition of the myocardial system to another level of structural and functional organization. PMID- 10707292 TI - [Growth of human epidermal keratinocytes on the collagen gel]. PMID- 10707293 TI - [Changes in the structural and functional characteristics of the cellular membranes during impulse acoustic exposure]. PMID- 10707294 TI - [The role of thymic peptides in the regulation of the lymphoid tissue growth in rats of various ages]. PMID- 10707295 TI - [Genome heterozygosity, metabolism rate, and life span]. PMID- 10707296 TI - The sustainability of welfare states into the twenty-first century. AB - Few would deny that the advanced welfare state faces a set of severe challenges. There is less agreement on what the challenges are and how important they are. The three most cited sources of crisis are population aging, family instability, and the labor market consequences of globalization and technological change. It is, however, questionable whether these affect the three dominant types of welfare systems similarly. The past decade bears witness to numerous attempts at reform and readaptation but such efforts have, so far, perpetuated or even strengthened underlying disequilibria. The result is mounting diswelfare, especially among younger households. The author examines various reform strategies (privatization, decentralization, and familialization) and concludes that these imply suboptimality. A "win-win" policy can be identified to the extent that it simultaneously maximizes fertility and women's employment and minimizes poverty risks. Greater earnings and income inequalities probably cannot be escaped, but their impact can be neutralized through a shift toward guaranteed life chances through education and skills. PMID- 10707297 TI - The widening gap in death rates among income groups in the United States from 1967 to 1986. AB - Death rates in the United States have fallen since the 1960s, but improvements have not been shared equally by all groups. This study investigates the change in inequality in mortality by income level from 1967 to 1986. Comparable death rates are constructed for 1967 and 1986 using National Mortality Followback Surveys as numerators and National Health Interview Surveys as denominators. Direct age adjusted death rates are calculated for income levels for the U.S. noninstitutionalized civilian population 35 to 64 years old. A summary measure of inequality in mortality adjusts for differences in the size and definition of income groups in the two years. In both 1967 and 1986, mortality decreased with each rise in income level. Measured in relative terms, this inverse relationship was greater in 1986 then in 1967 for men and women, blacks and whites. Between 1967 and 1986, death rates for those with maximal income declined between two and three times more rapidly than did rates for the middle and low income groups. The greatest increase in relative inequality was seen among white males. PMID- 10707298 TI - Understanding income inequalities in health among men and women in Britain and Finland. AB - The aims of this study were to investigate whether the relationship between income and self-perceived health is similar for men and women in two contrasting welfare states, Britain and Finland; whether the relationship between income and health is accounted for by employment status, education, and occupational social class; and whether the association differs when using alternative ways of measuring income: gross individual and net household equivalent income. Among British and Finnish men, low household and low individual income were related to poor health, even after adjusting for employment status, education, and social class. The adjusted relationship between individual income and health was stronger for British than Finnish men. Among British and Finnish women, net household equivalent income was strongly related to health, but after adjusting for employment status, education, and social class this relationship became weaker for British women and practically disappeared for Finnish women. For British women the association between income and health differed strongly depending on the income measure used; gross individual income had almost no effect on health. These results indicate that the association between health and income has no threshold in the sense that only people in poverty have poorer health than others. In further studies of income and health, household equivalent income should be used as the principal measure of income with adjustments for employment status, and men and women should be studied separately. PMID- 10707299 TI - Upstream healthy public policy: lessons from the battle of tobacco. AB - Many consider public health and politics to be entirely separate worlds. Public health activities are generally well-motivated by public interest, perceived as value-free, scientific, and devoid of partisan preference. Politics, in contrast, can be viewed as a distasteful activity involving self-interested pressure groups, misuse of state power, and influence of money on national decisions. Public health and politics are inappropriate bedfellows if politics is reduced to party politics. Politics, of course, involves more than just party activities; it concerns the structure, distribution, and effects of power in society. Which groups pattern the social order? What are their sources of influence? How do they retain privileged status? What social effects result from the policies these groups shape? Viewed in this broader sense, politics is essential for effective public health and thus is the inescapable context of public health interventions. To disregard sociopolitical determinants of health is to relegate public health to prevention and promotion of individual risk behaviors. If public health is to be more successful in the 21st century, it must comprehend the magnitude of the forces against it and the strategies used to engineer its defeat. Public health interventions in the new millennium must be appropriate to their sociocultural context. PMID- 10707300 TI - Blaming the victim: aspects of the Brazilian case. AB - This report describes the consequences and some aspects of the origin and development of victim blaming in accident analysis, and some methods for investigating such events, with particular emphasis on the situation in Brazil. In Brazil, the spread of this practice seems to have been helped by several factors. (1) The idea that occupational accidents are simple phenomena with a limited number of causal factors linked to unsafe actions and/or conditions. In the past, the theory of accident proneness had less influence than in other countries. (2) Government regulations that stipulate the hiring of health and safety officers, production of "educational" material, and "preventive" campaigns that emphasize the role of the victim's "faulty" behavior in the origin of an accident. (3) Mandatory implementation of standardized models for accident investigation directed toward searching for a single "cause." Usually one conclusion, expressed in terms of unsafe acts or conditions, is formulated so that whoever performs an unsafe act is responsible for the accident. (4) Lack of knowledge, as shown in Brazilian publications on occupational accidents and in the evolution of studies on the nature of accident phenomena and of strategies adopted for their prevention. PMID- 10707301 TI - International actors and population policies in India, with special reference to contraceptive policies. AB - The international population policy agenda has traditionally been dominated by demographically driven population control policies. However, in the population policy development that preceded the International Conference on Population and Development in 1994, people's reproductive needs and rights received more emphasis. The aim of this study was to analyze how the new emphasis in population policies has been interpreted at the country level. In analyzing population policy rhetoric and its practical interpretations in India in 1994, the authors found that the rhetoric was broadening to encompass women's empowerment and reproductive health and that the use of direct method-specific monetary incentives and disincentives for accepting family planning methods was disapproved. However, population policy options were still considered mainly in terms of their ability to reduce fertility. Furthermore, the increased emphasis on the general market agenda was more important than that on reproductive needs and rights in molding population policies, as was evident in the greater stress on cost-recovery systems and nongovernmental actors. The findings suggest that the broader agenda for population policies and reproductive rights has been interpreted so that it can serve the aims of population-growth control and be implemented in the context of more market-oriented social policies and trade liberalization. PMID- 10707302 TI - To decentralize or not to decentralize, is that the question? Nicaraguan health policy under structural adjustment in the 1990s. AB - Since 1990, health services decentralization in Nicaragua has been accompanied by structural adjustment, resulting in reduced equity and accountability. Sandinista efforts in the 1980s to extend access to primary care and reduce class and regional disparities in the delivery of health services were accompanied by modest attempts to increase local-level accountability and responsiveness. The escalation of war in the late 1980s transformed this effort into greater de facto decentralization. Over the past decade, Nicaragua has used decentralization policy to restructure the health system through health spending cuts and the favoring of curative over preventive services; privatization and the promotion of user fees; and confusion of lines of accountability. The authors analyze the 1990s' health policies in Nicaragua, paying particular attention to the blending of decentralization policy with the fiscal and administrative reforms advanced by the International Monetary Fund, World Bank, and other international agencies. They conclude that analyzing decentralization as a sector-specific reform that can be ameliorated through technocratic modifications is insufficient. A full understanding of the problems and possibilities of decentralization requires an analysis of the political and economic context that conditions these policies. PMID- 10707303 TI - Health sector reform in Brazil: a case study of inequity. AB - Health sector reform in Brazil built the Unified Health System according to a dense body of administrative instruments for organizing decentralized service networks and institutionalizing a complex decision-making arena. This article focuses on the equity in health care services. Equity is defined as a principle governing distributive functions designed to reduce or offset socially unjust inequalities, and it is applied to evaluate the distribution of financial resources and the use of health services. Even though in the Constitution the term "equity" refers to equal opportunity of access for equal needs, the implemented policies have not guaranteed these rights. Underfunding, fiscal stress, and lack of priorities for the sector have contributed to a progressive deterioration of health care services, with continuing regressive tax collection and unequal distribution of financial resources among regions. The data suggest that despite regulatory measures to increase efficiency and reduce inequalities, delivery of health care services remains extremely unequal across the country. People in lower income groups experience more difficulties in getting access to health services. Utilization rates vary greatly by type of service among income groups, positions in the labor market, and levels of education. PMID- 10707304 TI - International dimensions of Colombian violence. AB - Violence is the main public health problem in Colombia, as demonstrated by current homicide rates (the highest in the world), the strong effect of violence on the health care sector, and the forceful displacement of citizens, among other factors. This violence has international dimensions and consequences: inequity resulting from the political-economic system and increased by the neoliberal adjustments; the international traffic of drugs and weapons; exile and forceful displacement of persons into bordering countries; and frequent violations of human rights and humanitarian international rights. The international community can play a role in the search for a solution to this problem, as the author discusses in terms of contributions made in the past and those that could be made in the near future. Colombian violence can be viewed as a problem, a message, and a challenge for international public health, and the author suggests specific areas in which international public health could contribute to the study and solution of this complex situation. PMID- 10707305 TI - Globalization and international trade in the twenty-first century: opportunities for and threats to the health sector in the south. AB - Globalization and international trade are important forces at the turn of the century. This article explores how freer international trade will affect developing countries that are net importers of health care goods and services. Four commodities are used as special cases for discussion: pharmaceuticals, health care technologies, pesticides, and tobacco and its related products. The authors discuss the role of international specialized agencies, such as the World Trade Organization, World Health Organization, and World Bank, that are concerned with international trade and its health and health care consequences, and argue that closer collaboration is required among these agencies if the negative effects of trade liberalization on developing countries are to be mitigated. The authors pose a number of research questions that could help in developing proactive policies for the South on the trade of goods and services with harmful effects on health as well as those with potential health and economic benefits. PMID- 10707306 TI - Refiguring "race": epidemiology, racialized biology, and biological expressions of race relations. AB - Given growing appreciation of how race/ethnicity is a social, not biological, construct, some epidemiologists are proposing that studies omit data on "race" and instead collect better socioeconomic data. This suggestion, however, ignores a growing body of evidence on how noneconomic as well as economic aspects of racial discrimination are embodied and harm health across the lifecourse. Developing a critical epidemiology of social inequalities in health will, at the very least, require incorporating thoughtful measures of race/ethnicity and social class in epidemiological studies and public health surveillance systems. PMID- 10707307 TI - On the study of race, racism, and health: a shift from description to explanation. AB - While no credible scientist believes that race has any biological or genetic basis, it does have profound social meaning, rooted in history but with contemporary consequences. Racial status is a risk marker for exposure to racism, which may be a primary etiological factor in race differences in morbidity and mortality. The study of racism and health is highly complex and still in its infancy. What we need is not just more research on race, but better research. PMID- 10707308 TI - Reply to commentaries by Drs. Krieger and LaVeist on "race in epidemiology". AB - Racial categorizations are a dangerous anachronism with a disastrous history and fatal consequences for the human species. They play into the hands of racists and undermine the solidarity of some impoverished and neglected segments of our societies. Race is no longer a constructive tool of health or social research and should be abandoned as a category in research and programs of economic, social, and health amelioration. PMID- 10707309 TI - Multiple motor patterns in the stomatogastric ganglion of the shrimp Penaeus japonicus. AB - Motor patterns of the cardiac sac, the gastric and the pyloric network in the stomatogastric nervous system of the shrimp Penaeus japonicus, the most primitive decapod species, were studied. Single neurons can switch from the gastric or the pyloric pattern to the cardiac sac pattern. Some of the pyloric neurons fire with the gastric pattern. All of the gastric neurons fire with the pyloric pattern, unlike those in reptantians. Proctolin activates and modulates the cardiac sac and the pyloric rhythm, and promotes reconfiguration of the networks. Neurons of the three networks have so many interconnections that they construct a multifunctional neural network like those in Cancer. This network may function in different configurations under the appropriate conditions. Several modes of interactions between the networks found in different reptantian species can apply to the penaeidean shrimp. Such interactions are general features of the stomatogastric nervous system in decapods. Phylogenetic differences among the decapod infraorders are seen in the number and orientation of muscles and the innervation pattern of muscles. The multifunctional networks have existed in the most primitive decapod species, and types of configurations of the networks would have evolved to produce a wide range of motor patterns as the foregut structure has become complex. PMID- 10707310 TI - Photoreceptor visual fields, ommatidial array, and receptor axon projections in the polarisation-sensitive dorsal rim area of the cricket compound eye. AB - We made intracellular recordings from the photoreceptors of the polarisation sensitive dorsal rim area of the cricket compound eye combined with dye marking. By measuring visual field sizes and optical axes in different parts of the dorsal rim area, we assessed the optical properties of the ommatidia. Due to the large angular sensitivities (median about 20 degrees) and the high sampling frequency (about 1 per degree), the visual fields overlap extensively, such that a given portion of the sky is viewed simultaneously by a large number of ommatidia. By comparing the dye markings in the retina and in the optic lobe, the axon projections of the retinula cells were examined. Receptors R1, R2, R5 and R6 project to the lamina, whereas R7 projects to the medulla. The microvilli orientation of the two projection types differ by 90 degrees indicating the two analyser channels that give antagonistic input to polarisation-sensitive interneurons. Using the retinal marking pattern as an indicator for the quality of the intracellular recordings, the polarisation sensitivity of the photoreceptors was re-examined. The polarisation sensitivity of recordings from dye-coupled cells was much lower (median: 4.5) than that of recordings in which only one cell was marked (median: 9.8), indicating that artefactual electrical coupling between photoreceptors can significantly deteriorate polarisation sensitivity. The physiological value of polarisation sensitivity in the cricket dorsal rim area is thus typically about 10. PMID- 10707311 TI - The sounds of silence: cessation of singing and song pausing are ultrasound induced acoustic startle behaviors in the katydid Neoconocephalus ensiger (Orthoptera; Tettigoniidae). AB - Previous studies of acoustic startle in insects have dealt with behavioral and/or neural mechanisms employed in evading aerially hawking, echolocating bats; however, insects also face terrestrial predators. Here we describe an acoustic startle response of the nocturnal katydid, Neoconocephalus ensiger. Stridulating males disturbed in the field perform obvious antipredatory behaviors--cessation of singing, freezing, jumping, and evasive flight. Under controlled laboratory conditions we found that cessation of singing and song pausing are ultrasound specific behaviors: when stimulated with pulsed ultrasound (20-100 kHz), but not audio-sound (< 20 kHz), males cease mate calling or insert pauses in their song. A second factor influencing acoustic startle is the phase of stimulation: an acoustic startle response occurs only when the pulse of ultrasound arrives during the window of silence between stridulatory syllables. The average startle threshold and response latency was 70 +/- 5 dB SPL and 34.2 +/- 6.0 ms, respectively. N. ensiger is particularly useful for examining acoustic startle responses of nonflying insects because (1) its calling song is broadband and contains ultrasound, thus the possibility exists of confusion over the biological meaning of ultrasound, and (2) this species shows the classic bat-avoidance response while flying, so a direct comparison between two types of acoustic startle is possible within the same species. PMID- 10707312 TI - Sulphur-containing "perfumes" attract flower-visiting bats. AB - We tested the attractiveness of individual scent compounds of bat-pollinated flowers to their pollinators, small flower-visiting bats of the genus Glossophaga (Phyllostomidae). Twenty compounds belonging to four different chemical substance classes were tested, both in the laboratory and in the field. In the laboratory, the bats (Glossophaga soricina) approached odour sources spontaneously and without preceding experience. Without ever receiving any reward they preferred the scent of a sulphur-containing compound, dimethyl disulphide, to several other odour components emitted by bat-pollinated flowers, and to scentless controls. In the field, at La Selva station in the tropical lowland rain forest of Costa Rica, G. commissarisi were attracted by two sulphur-containing compounds, dimethyl disulphide and 2,4-dithiapentane, to visit artificial flowers filled with sugar water. Thus, in nectarivorous bats the sense of smell obviously plays an important role in searching for and localising food sources, and even single components of the scent bouquets of bat-pollinated flowers are attractive. The preference for sulphur-containing odours seems to be innate. PMID- 10707313 TI - Prey pursuit and interception in dragonflies. AB - Perching dragonflies (Libellulidae; Odonata) are sit-and-wait predators, which take off and pursue small flying insects. To investigate their prey pursuit strategy, we videotaped 36 prey-capture flights of male dragonflies, Erythemis simplicicollis and Leucorrhinia intacta, for frame-by-frame analysis. We found that dragonflies fly directly toward the point of prey interception by steering to minimize the movement of the prey's image on the retina. This behavior could be guided by target-selective descending interneurons which show directionally selective visual responses to small-object movement. We investigated how dragonflies discriminate distance of potential prey. We found a peak in angular velocity of the prey shortly before take-off which might cue the dragonfly to nearby flying targets. Parallax information from head movements was not required for successful prey pursuit. PMID- 10707314 TI - Spherical sound radiation patterns of singing grass cicadas, Tympanistalna gastrica. AB - The spatial pattern of sound radiation of grass cicadas emitting normally patterned calling songs was measured in the acoustic far field with an array of eight microphones at a distance of 15 cm. The array could be rotated to cover the sphere around the cicada. The sound was analysed in one-third-octave bands with centre frequencies from 3.15 kHz to 16 kHz, the frequency range of the calling song. The seven cicadas studied had very similar spatial radiation patterns, but somewhat different emitted sound powers (range 190-440 nW, mean 280 nW, at 22 degrees C). At low frequencies, the pattern of sound radiation was close to spherical. At higher frequencies, systematic deviations from a spherical pattern were evident. The deviations were of the order of magnitude expected for monopolar sound sources located on sound-shielding bodies. We conclude that, although the singing cicada produces a quite complex acoustic near field, it behaves as a monopole in the far field. These findings are compared with data from a singing grasshopper of similar size, which in the far field behaves as a multipole. PMID- 10707315 TI - A circadian pacemaker in free-living chipmunks: essential for survival? AB - The importance of circadian timing was evaluated for 18 months from late-April 1997 through October 1998 in a high-density population of free-living eastern chipmunks, Tamias striatus, at a 4-ha forest site in the Allegheny Mountains. Included in the radiocollared field group were 30 chipmunks with supra-chiasmatic nucleus-targeted lesions, 24 surgical controls, and 20 intact controls. An additional 17 chipmunks were used in a laboratory study as lesion-calibration controls to correlate degree of circadian arrhythmicity with extent of supra chiasmatic nucleus deletion. Survival was documented in the field by daily radio tracking and by regular trapping censuses except during winter hibernation. A significantly higher proportion of supra-chiasmatic nucleus-lesioned than surgical control chipmunks or intact controls were killed by weasel predation during the first 80 days after repatriation. A 28-h continuous census found no surface activity of any chipmunks during hours of darkness. However, episodes of nocturnal movement were detected within the permanent dens by radio telemetric data logging, especially in supra-chiasmatic nucleus-lesioned animals. Excavation and mapping of six chipmunk burrow systems aided in the interpretation of the telemetric activity data. Nighttime restlessness of supra-chiasmatic nucleus lesioned animals may have acted as a clue to the predator for locating its prey. PMID- 10707316 TI - A gain-control mechanism for processing of chorus sounds in the afferent auditory pathway of the bushcricket Tettigonia viridissima (Orthoptera; Tettigoniidae). AB - The representation of alternative conspecific acoustic signals in the responses of a pair of local interneurons of the bushcricket Tettigonia viridissima was studied with variation in intensity and the direction of sound signals. The results suggest that the auditory world of the bushcricket is rather sharply divided into two azimuthal hemispheres, with signals arriving from any direction within one hemisphere being predominantly represented in the discharge of neurons of this side of the auditory pathway. In addition, each pathway also selects for the most intense of several alternative sounds. A low-intensity signal at 45 dB sound pressure level is quite effective when presented alone, but completely suppressed when given simultaneously with another signal at 60 dB sound pressure level. In a series of intracellular experiments the synaptic nature of the intensity-dependent suppression of competitive signals was investigated in a number of interneurons. The underlying synaptic mechanism is based on a membrane hyperpolarization with a time-constant in the order of 5-10 s. The significance of this mechanism for hearing in choruses, and for the evolution of acoustic signals and signalling behaviour is discussed. PMID- 10707317 TI - Chemosensory control of pollen ingestion in the hoverfly Eristalis tenax by labellar taste hairs. AB - The labellar gustatory system of the dronefly Eristalis tenax L. (Syrphidae; Diptera) that enables the fly to discriminate between pollen and nectar is investigated, and the triggering of pollen ingestion is examined. In behavioural preference tests, exhaustively extracted pollen of the sunflower Helianthus annuus is consumed in smaller amounts than untreated pollen, indicating that water-soluble substances are important for acceptance. Dry pollen is preferred over moist pollen in which the grains stick together, suggesting that mechanical properties of the pollen also play a role in its sensory assessment. Electrophysiological studies of the labellar taste hairs reveal that aqueous extracts of pollen (2% w/v) stimulate the salt receptor cell, but not the sugar receptor cell. The response of the water receptor cell remains the same as to pure water (or standard electrolyte, 10 mmol.l-1 KCl). Of the 20 amino acids tested, the salt cell is sensitive only to proline in a submillimolar range. Behavioural experiments support the electrophysiological findings. When KCl is applied at concentrations eliciting salt-cell spike frequencies equal to those produced by pollen extract (which is often accepted), the water receptor cell is inhibited and a pronounced rejection behaviour occurs. This rejection of concentrated salt solution in Eristalis is therefore mainly mediated by the inhibition of the water cell. PMID- 10707318 TI - Circadian rhythms of melatonin in European starlings exposed to different lighting conditions: relationship with locomotor and feeding rhythms. AB - In passerine birds, the periodic secretion of melatonin by the pineal organ represents an important component of the pacemaker that controls overt circadian functions. The daily phase of low melatonin secretion generally coincides with the phase of intense activity, but the precise relationship between the melatonin and the behavioral rhythms has not been studied. Therefore, we investigated in European starlings (Sturnus vulgaris) (1) the temporal relationship between the circadian plasma melatonin rhythm and the rhythms in locomotor activity and feeding; (2) the persistence of the melatonin rhythm in constant conditions; and (3) the effects of light intensity on synchronized and free-running melatonin and behavioral rhythms. There was a marked rhythm in plasma melatonin with high levels at night and/or the inactive phase of the behavioral cycles in almost all birds. Like the behavioral rhythms, the melatonin rhythm persisted for at least 50 days in constant dim light. In the synchronized state, higher day-time light intensity resulted in more tightly synchronized rhythms and a delayed melatonin peak. While all three rhythms usually assumed a rather constant phase relationship to each other, in one bird the two behavioral rhythms dissociated from each other. In this case, the melatonin rhythm retained the appropriate phase relationship with the feeding rhythm. PMID- 10707319 TI - Gas exchange and heart rate in the harbour porpoise, Phocoena phocoena. AB - The respiratory physiology, heart rates and metabolic rates of two captive juvenile male harbour porpoises (both 28 kg) were measured using a rapid-response respiratory gas analysis system in the laboratory. Breath-hold durations in the laboratory (12 +/- 0.3 s, mean +/- SEM) were shorter than field observations, although a few breath-holds of over 40 s were recorded. The mean percentage time spent submerged was 89 +/- 0.4%. Relative to similarly-sized terrestrial mammals, the respiratory frequency was low (4.9 +/- 0.19 breaths.min-1) but with high tidal volumes (1.1 +/- 0.011), enabling a comparatively high minute rate of gas exchange. Oxygen consumption under these experimental conditions (247 +/- 13.8 ml O2.min-1) was 1.9-fold higher than predicted by standard scaling relations. These data together with an estimate of the total oxygen stores predicted an aerobic dive limit of 5.4 min. The peak end-tidal O2 values were related to the length of the previous breath-hold, demonstrating the increased oxygen uptake from the lung for the longer dives. Blood oxygen capacity was 23.5 +/- 1.0 ml.100 ml-1, and the oxygen affinity was high, enabling rapid oxygen loading during ventilation. PMID- 10707320 TI - Ionic currents in cardiac myocytes of squid, Alloteuthis subulata. AB - This paper provides the first study of voltage-sensitive membrane currents present in heart myocytes from cephalopods. Whole cell patch clamp recordings have revealed six different ionic currents in myocytes freshly dissociated from squid cardiac tissues (branchial and systemic hearts). Three types of outward potassium currents were identified: first, a transient outward voltage-activated A-current (IA), blocked by 4-aminopyridine, and inactivated by holding the cells at a potential of -40 mV; second, an outward, voltage-activated, delayed rectifier current with a sustained time course (IK); and third, an outward, calcium-dependent, potassium current (IK(Ca)) sensitive to Co2+ and apamin, and with the characteristic N-shaped current voltage relationship. Three inward voltage-activated currents were also identified. First, a rapidly activating and inactivating, sodium current (INa), blocked by tetrodotoxin, inactivated at holding potentials more positive than -40 mV, and abolished when external sodium was replaced by choline. Second, an L-type calcium current (ICa,L) with a sustained time course, suppressed by nifedipine or Co2+, and enhanced by substituting Ca2+ for Ba2+ in the external medium. The third inward current was also carried by calcium ions, but could be distinguished from the L-type current by differences in its voltage dependence. It also had a more transient time course, was activated at more negative potentials, and resembled the previously described low-voltage-activated, T-type calcium current. PMID- 10707321 TI - Amylase mRNA expression in Crassostrea gigas during feeding cycles. AB - A Crassostrea gigas digestive gland copy DNA (cDNA) library constructed in the lambda phage ZapII (Stratagene, La Jola, USA) was screened with an amylase heterologous proble. To get access to the complete cDNA, a polymerase chain reaction extension was conducted using DNA extracted from the phages. The complete cDNA sequence is 1688 base pairs (EMBL = Y08370). The deduced protein sequence is 519 aminoacids long with a 19 aminoacid signal peptide. Similarity with Pecten maximus amylase is 72%. A 3-day nutrition experiment with a cyclic algal food supply was carried out. Amylase enzyme activities and mRNAs were individually measured on five animals, nine times a day. Messenger RNAs were quantified by dot hybridization using the previously characterized cDNA as probe. Variation of amylase mRNA was observed, in relation with the level of activity of the enzyme. Coordinated changes in RNA and enzyme levels suggested a possible transcriptional regulation of amylase in C. gigas as in vertebrates. PMID- 10707322 TI - Control of salt gland activity in the hatchling green sea turtle, Chelonia mydas. AB - We studied the control of salt gland secretion in hatchling Chelonia mydas. The threshold salt load to activate salt secretion was between 400 mumol NaCl 100 g bodymass (BM)-1 and 600 mumol NaCl 100 g BM-1, which caused an increase in plasma sodium concentration of 13% to 19%. Following a salt load of 2700 mumol NaCl 100 g BM-1, salt gland secretion commenced in 12 +/- 1.3 min and reached maximal secretory concentration within 2-7 min. Maximal secretory rate of a single gland averaged 415 mumol Na 100 g BM-1 h-1. Plasma sodium concentration and total osmotic concentration after salt loading were significantly higher than pretreatment values within 2 min. Adrenalin (25 micrograms kg BM-1) and the cholinergic agonist methacholine (1 mg kg BM-1) inhibited salt gland activity. Atropine (10 mg kg BM-1) reversed methacholine inhibition and stimulated salt gland secretion when administered with a subthreshold salt load. Arginine vasotocin produced a transient reduction in sodium secretion by the active gland, while atrial natriuretic factor, vasoactive intestinal peptide and neuropeptide Y had no measurable effect on any aspect of salt gland secretion. Our results demonstrated that secretion of the salt gland in C. mydas can be modified by neural and hormonal chemicals in vivo and that the cholinergic and adrenergic stimulation of an exocrine gland do not appear to have the typical, antagonist actions on the chelonian salt gland. PMID- 10707323 TI - Modulation of leptin sensitivity by short photoperiod acclimation in the Djungarian hamster, Phodopus sungorus. AB - During seasonal acclimation, Djungarian hamsters spontaneously exhibit a reduction in food intake, body mass and body fat stores, which is externally cued by shortening of day length in autumn and controlled by a sliding set-point. We investigated the function of the leptin adipostatic feedback system in the photoperiodic control of seasonal acclimation. In response to mouse recombinant leptin injections for 10 days, long day photoperiod (LD) and short day photoperiod (SD)-acclimated hamsters decreased food intake and body mass. The reduction of body mass was due to the depletion of body fat, as revealed by carcass composition analysis. In SD hamsters, leptin caused a larger reduction of body fat mass than observed under LD conditions, whereas the anorectic effect was similar in both photoperiods. The serum leptin concentration was 9.3 +/- 1.2 ng/ml in LD-acclimated hamsters and decreased significantly to 4.2 +/- 0.8 ng/ml and 2.1 +/- 0.6 ng/ml in hamsters exposed to SD for 66 days and 116 days, respectively (P < 0.001). A strong positive correlation between total body fat mass and serum leptin concentration was found (rS = 0.935, P < 0.0001, n = 70). Despite the anorectic action of exogenous leptin, higher endogenous leptin levels in LD hamsters were paralleled by higher food intake in LD hamsters as compared to SD hamsters. This paradoxical finding further supports the increased leptin sensitivity in SD hamsters as judged from leptin treatment experiments. We tested the functional significance of leptin for the controlled down-regulation of food intake and body mass induced by short photoperiod. Food restriction for 10 days during the transition phase decreased body mass below the desired sliding set point, which was recovered in control hamsters following ad libitum refeeding. Treatment with mouse recombinant leptin during ad libitum refeeding inhibited the recovery of body mass and blunted the increase of food intake observed in controls, indicating that the sliding set-point utilizes leptin as a signal for the adjustment of the appropriate body mass level. PMID- 10707324 TI - Accumulation of lactate by supercooled hatchlings of the painted turtle (Chrysemys picta): implications for overwinter survival. AB - Hatchlings of the North American painted turtle (Chrysemys picta) typically spend their first winter of life inside the shallow, subterranean nest where they completed embryogenesis the preceding summer. Neonates at northern localities consequently may be exposed during winter to subzero temperatures and frozen soil. Hatchlings apparently survive exposure to such conditions by supercooling, but the physiological consequences of this adaptive strategy have not been examined. We measured lactate in hatchling painted turtles after exposure to each of three temperatures (0 degree C, -4 degrees C, and -8 degrees C) for three time periods (5 days, 15 days, and 25 days) to determine the extent to which overwintering hatchlings might rely on anaerobic metabolism to regenerate ATP. Whole-body lactate increased with increasing duration of exposure and decreasing temperature, and the highest levels were associated with the group that experienced the highest mortality. These results indicate that animals may develop a considerable lactic acidosis during a winter in which temperatures fall below 0 degree C for weeks or months and that accumulation of lactate may contribute to mortality of overwintering animals. PMID- 10707325 TI - Influence of flight on protein catabolism, especially myofilament breakdown, in homing pigeons. AB - In order to study protein degradation during flight in homing, a high-performance liquid chromatography technique was developed for the quantitative analysis of N tau-methylhistidine. Secondly, it was necessary to confirm that the excretion of N tau-methylhistidine correlates with myofilament breakdown in homing pigeons. In these experiments, ten birds were subcutaneously injected with N tau [14C]methylhistidine and the excreta were quantitatively collected for 1 week. Of the 94.5% radioactivity recovered, 87.1% was associated with N tau [14C]methylhistidine and 6.1% with N-acetyl-N tau-[14C]methylhistidine. This rapid excretion of unmetabolized N tau-[14C]methylhistidine validates the assumption that the amount of N tau-methylhistidine excreted is a measure of myofilament catabolism in homing pigeons. The influence of endurance flight on protein breakdown was determined after flights from release sites 368-646 km away. Immediately after return, plasma urea and uric acid levels were increased, whereas plasma concentration of N tau-methylhistidine remained unchanged compared to unflown control birds. Flown pigeons excreted significantly more urea and N tau-methylhistidine within 24 h and significantly more urea and uric acid within 96 h after flight than unflown controls. Our findings support the hypothesis that in homing pigeons protein catabolism is increased during endurance flight. Elevated N tau-methylhistidine excretion probably results from repair processes in damaged muscle fibers, including breakdown of myofilaments. PMID- 10707326 TI - Metabolism and temperature regulation during daily torpor in the smallest primate, the pygmy mouse lemur (Microcebus myoxinus) in Madagascar. AB - Thermoregulation, energetics and patterns of torpor in the pygmy mouse lemur, Microcebus myoxinus, were investigated under natural conditions of photoperiod and temperature in the Kirindy/CFPF Forest in western Madagascar. M. myoxinus entered torpor spontaneously during the cool dry season. Torpor only occurred on a daily basis and torpor bout duration was on average 9.6 h, and ranged from 4.6 h to 19.2 h. Metabolic rates during torpor were reduced to about 86% of the normothermic value. Minimum body temperature during daily torpor was 6.8 degrees C at an ambient temperature of 6.3 degrees C. Entry into torpor occurred randomly between 2000 and 0620 hours, whereas arousals from torpor were clustered around 1300 hours within a narrow time window of less than 4 h. Arousal from torpor was a two-step process with a first passive climb of body temperature to a mean of 27 degrees C, carried by the daily increase of ambient temperature when oxygen consumption remained more or less constant, followed by a second active increase of oxygen consumption to further raise the body temperature to normothermic values. In conclusion, daily body temperature rhythms in M. myoxinus further reduce the energetic costs of daily torpor seen in other species: they extend to unusually low body temperatures and consequently low metabolic rates in torpor, and they employ passive warming to reduce the energetic costs of arousal. Thus, these energy-conserving adaptations may represent an important energetic aid to the pygmy mouse lemur and help to promote their individual fitness. PMID- 10707327 TI - Availability of water affects organ growth in prenatal and neonatal snapping turtles (Chelydra serpentina). AB - We manipulated the amount of water that was available to prenatal and neonatal snapping turtles (Chelydra serpentina) in order to assess the impact of water on growth by different organs in these animals. Three treatments were used: (1) turtles that completed their incubation on a wet substrate, (2) turtles that completed their incubation on a dry substrate, and (3) turtles that spent a few days in water after completing incubation on a dry substrate. Turtles hatching on a dry substrate (treatment 2) were smaller than animals in the other two treatments (which did not differ in size), so data for mass of different organs were adjusted by ANCOVA to remove effects of body size. Scaled masses of liver, stomach, lungs, kidneys, and small intestine did not differ between turtles emerging in wet environments and those hatching in dry environments, but hearts of turtles hatching in dry settings were substantially larger than those of animals hatching in wet ones. Thus, the mass of most organs in turtles developing in wet and dry environments scaled to body size, whereas the heart was hypertrophied in embryos developing in dry environments. Turtles that spent a few days in water after hatching from eggs in dry environments grew rapidly in size, and the increase in body size was accompanied by disproportionately rapid growth in the liver, stomach, lungs, kidneys, and small intestine. The heart did not increase in size during this period, despite the substantial increase in body mass over that at hatching. The enlarged heart of turtles hatching on dry substrates may have been caused by a circulatory hypovolemia late in incubation; the rapid growth of organs other than the heart when these animals were placed in water may reflect a release from constraints on growth once circulatory volume was restored. PMID- 10707328 TI - The role of ion regulation in the control of the distribution of Gammarus tigrinus (Sexton) in salt-polluted rivers. AB - Fluctuating salinities at different sites on the German salt-polluted rivers Werra and Weser were compared with extracellular ion levels of specimens of Gammarus tigrinus (Sexton; Amphipoda, Crustacea), collected at the same sites. G. tigrinus regulated haemolymph concentrations of inorganic anions (Cl-, SO(4)2-, PO(4)3-) and cations (Na+, K+, Mg2+, Ca2+) during fluctuations of salt pollution in the upper Weser. This capacity to regulate varying levels of salt pollution in the upper Weser, correlated well with the distribution of the brackish amphipods in this river ecosystem. G. tigrinus tolerated periods of Na+ and Cl- stress (> 380 mmol l-1) without compensating these maxima by regulating extracellular Na+ and Cl-. However, during such bursts of Na+ and Cl- stress in Werra and Weser, the ability to regulate extracellular [K+] at river water K+ stress of > or = 6.0 mmol l-1 may explain why this brackish species has been more successful in these rivers than its competitors like Gammarus pulex. The present investigation demonstrates that the water salinity affects the [NO3-] in the haemolymph of G. tigrinus. With increasing hypo-osmotic stress the animals accumulate increasing amounts of NO3-. A simultaneous increase in stream water [NO3-] causes an additional accumulation of NO3- in the haemolymph. The high extent of accumulation indicates that active ion transport systems may be involved. The accumulation of NO3- in the haemolymph has low physiological consequences to G. tigrinus, but when hypo-osmotically stressed under anoxic conditions, nitrite formed by the reduction of nitrate may have an adverse affect on the metabolism of G. tigrinus. PMID- 10707329 TI - On the costs of accessible attitudes: detecting that the attitude object has changed. AB - The present research examined whether individuals with more accessible attitudes have more difficulty detecting that the attitude object has changed. While being repeatedly exposed to photographs of undergraduates, participants either rehearsed their attitudes toward each photo or performed a control task. They then saw these original photos and computer-generated morphs representing varying degrees of change in an original. Participants in the attitude rehearsal condition required more time to correctly identify morphs that were similar to the original as "different" (Experiment 1) and made more errors in response to such morphs (Experiment 2). Experiment 3 revealed that participants with accessible attitudes perceived relatively less change; they were less likely to view a morph as a photo of a novel person and more likely to view it as a different photo of a person seen before. The costs and benefits of accessible attitudes are discussed. PMID- 10707330 TI - The spotlight effect in social judgment: an egocentric bias in estimates of the salience of one's own actions and appearance. AB - This research provides evidence that people overestimate the extent to which their actions and appearance are noted by others, a phenomenon dubbed the spotlight effect. In Studies 1 and 2, participants who were asked to don a T shirt depicting either a flattering or potentially embarrassing image overestimated the number of observers who would be able to recall what was pictured on the shirt. In Study 3, participants in a group discussion overestimated how prominent their positive and negative utterances were to their fellow discussants. Studies 4 and 5 provide evidence supporting an anchoring-and adjustment interpretation of the spotlight effect. In particular, people appear to anchor on their own rich phenomenological experience and then adjust- insufficiently--to take into account the perspective of others. The discussion focuses on the manifestations and implications of the spotlight effect across a host of everyday social phenomena. PMID- 10707331 TI - Varieties of groups and the perception of group entitativity. AB - Three studies examined perceptions of the entitativity of groups. In Study 1 (U.S.) and Study 2 (Poland), participants rated a sample of 40 groups on 8 properties of groups (e.g., size, duration, group member similarity) and perceived entitativity. Participants also completed a sorting task in which they sorted the groups according to their subjective perceptions of group similarity. Correlational and regression analyses were used to determine the group properties most strongly related to entitativity. Clustering and multidimensional scaling analyses in both studies identified 4 general types of groups (intimacy groups, task groups, social categories, and loose associations). In Study 3, participants rated the properties of groups to which they personally belonged. Study 3 replicated the results of Studies 1 and 2 and demonstrated that participants most strongly valued membership in groups that were perceived as high in entitativity. PMID- 10707332 TI - Motivated cultural cognition: the impact of implicit cultural theories on dispositional attribution varies as a function of need for closure. AB - The authors propose that need for closure (NFC) leads attributors to respond to an ambiguous social event by increasing reliance on implicit theories received from acculturation. Hence, the influence of NFC should be shaped by chronically accessible knowledge structures in a culture, and, likewise, the influence of culture should be moderated by epistemic motives such as NFC. The specific hypotheses drew on past findings that North American and Chinese attributors possess differing implicit social theories, North Americans conceiving of individuals as autonomous agents and Chinese conceiving of groups as autonomous. The present studies found the predicted pattern that among North American participants, NFC increased attributions to personal but not group dispositions. Among Chinese participants, NFC increased attributions to group but not personal dispositions. The findings are discussed in light of an emerging dynamic account of culture and cognition. PMID- 10707333 TI - On inferring one's beliefs from one's attempt and consequences for subsequent compliance. AB - D. J. Bem (1967, 1972) has suggested that a person may infer his or her beliefs from his or her actions. With his information-processing viewpoint, D. J. Bem proposed that individuals, by observing their past behaviors, may draw information for assessing their beliefs about themselves. There is a question, however, about the mechanism of self-perception when there is inconsistency between one's attempt to realize an intended goal and the outcome of the action. In a series of field studies, participants who had unsuccessfully tried to help a stranger were more willing to comply with a relatively large request made later. Implications for self-perception theory as well as for enhancing susceptibility to influence techniques are discussed. PMID- 10707334 TI - Couples' shared participation in novel and arousing activities and experienced relationship quality. AB - Using a newspaper questionnaire, a door-to-door survey, and 3 laboratory experiments, the authors examined a proposed effect of shared participation in novel and arousing activities on experienced relationship quality. The questionnaire and survey studies found predicted correlations of reported shared "exciting" activities and relationship satisfaction plus their predicted mediation by relationship boredom. In all 3 experiments, the authors found predicted greater increases in experienced relationship quality from before to after participating together in a 7-min novel and arousing (vs. a more mundane) task. Comparison with a no-activity control showed the effect was due to the novel-arousing task. The same effect was found on ratings of videotaped discussions before and after the experimental task. Finally, all results remained after controlling for relationship social desirability. Results bear on general issues of boredom and excitement in relationships and the role of such processes in understanding the typical early decline of relationship quality after the honeymoon period. PMID- 10707335 TI - Understanding attachment security in family context. AB - Attachment theory (J. Bowlby, 1969) is not just about how internalized models of relationships affect interpersonal outcomes; it is primarily a theory about how interpersonal processes affect social and cognitive development. This study tested 3 hypotheses about the interpersonal sources of adult attachment security: (a) attachment security is relationship specific, (b) characteristics of partners affect attachment security, and (c) security of attachment is reciprocated. Measures of attachment security were obtained from 2 parents and 2 children (adolescent or older) in 208 middle-class families. Results of social relations model analysis (D. A. Kenny & L. La Voie, 1984) supported all 3 hypotheses. The author concludes that internal working models of relationships may not be so "internal" after all and that greater emphasis on the interpersonal sources of adult attachment security is warranted. PMID- 10707336 TI - Attributions in marriage: state or trait? A growth curve analysis. AB - Research on attributions in marriage rests on 2 assumptions. First, the attributions spouses make for their partners' behaviors have been treated as a style or a trait, reflecting enduring aspects of the perceiver. Second, attributions have been described as a causal factor in the development of the marriage over time. To evaluate the evidence for these assumptions, the authors analyzed 8 waves of longitudinal data from a sample of newlywed couples. Results offered no support for the idea of an enduring attributional style; attributions changed linearly, and changes in attributions were strongly associated with changes in marital satisfaction within each spouse. Nevertheless, controlling for these associations, initial levels of attributions predicted changes in marital satisfaction more than initial satisfaction predicted changes in attributions. Effects of neuroticism and effects on marital dissolution were also examined. PMID- 10707337 TI - Linking childhood personality with adaptation: evidence for continuity and change across time into late adolescence. AB - Four personality traits--Mastery Motivation, Academic Conscientiousness, Surgent Engagement, and Agreeableness--were measured in a community sample of 205 children (ages 8-12), who were followed up 10 years later. Childhood personality was examined in relation to concurrent and longitudinal adaptation in 3 domains- academic achievement, rule-abiding versus antisocial conduct, and peer social competence. The childhood personality traits evidenced robust, conceptually coherent relationships with adaptation, both concurrently and across time. Childhood personality added to the prediction of later adaptation, beyond childhood IQ and earlier adaptation in the same domain. Few sex differences were obtained in the relationships between childhood personality and adaptation. The present results document both continuity and discontinuity in the links between childhood personality and adaptation across childhood into late adolescence. PMID- 10707338 TI - How does personality matter in marriage? An examination of trait anxiety, interpersonal negativity, and marital satisfaction. AB - Although trait anxiety and its aliases (negative affectivity, neuroticism) have frequently been found to be associated with marital dissatisfaction, few efforts have been made to identify the processes through which trait anxiety exerts its influence. This study reports findings from a 13-year, 4-phase longitudinal study in which trait anxiety, measured when spouses were newlyweds, consistently predicted marital negativity which, in turn, was associated with partner's marital dissatisfaction. Some support was also found for effects of trait anxiety on partner's marital satisfaction, independent of marital negativity, as well as for the idea that trait anxiety is directly related to spouses' own marital satisfaction. Trait anxiety did not distinguish couples who divorced from those who remained married, and it generally did not predict declines in marital satisfaction. The disagreeable impact of trait anxiety on marriage was evident at the outset of marriage and was stable over time. PMID- 10707339 TI - Longitudinal multilevel models of the big-fish-little-pond effect on academic self-concept: counterbalancing contrast and reflected-glory effects in Hong Kong schools. AB - Longitudinal multilevel path models (7,997 students, 44 high schools, 4 years) evaluated effects of school-average achievement and perceived school status on academic self-concept in Hong Kong, which has a collectivist culture with a highly achievement-segregated high school system. Consistent with a priori predictions based on the big-fish-little-pond effect (BFLPE), higher school average achievements led to lower academic self-concepts (contrast effect), whereas higher perceived school status had a counterbalancing positive effect on self-concept (reflected-glory, assimilation effect). The negative BFLPE is the net effect of counterbalancing influences, stronger negative contrast effects, and weaker positive assimilation effects so that controlling perceived school status led to purer--and even more negative--contrast effects. Attending a school where school-average achievement is high simultaneously resulted in a more demanding basis of comparison for one's own accomplishments (the stronger negative contrast effect) and a source of pride (the weaker positive assimilation effect). PMID- 10707340 TI - An item response theory analysis of self-report measures of adult attachment. AB - Self-report measures of adult attachment are typically scored in ways (e.g., averaging or summing items) that can lead to erroneous inferences about important theoretical issues, such as the degree of continuity in attachment security and the differential stability of insecure attachment patterns. To determine whether existing attachment scales suffer from scaling problems, the authors conducted an item response theory (IRT) analysis of 4 commonly used self-report inventories: Experiences in Close Relationships scales (K. A. Brennan, C. L. Clark, & P. R. Shaver, 1998), Adult Attachment Scales (N. L. Collins & S. J. Read, 1990), Relationship Styles Questionnaire (D. W. Griffin & K. Bartholomew, 1994) and J. Simpson's (1990) attachment scales. Data from 1,085 individuals were analyzed using F. Samejima's (1969) graded response model. The authors' findings indicate that commonly used attachment scales can be improved in a number of important ways. Accordingly, the authors show how IRT techniques can be used to develop new attachment scales with desirable psychometric properties. PMID- 10707341 TI - Isms and the structure of social attitudes. AB - Social attitude measurement has been limited by inadequate structural models. In this study, broad, basic dimensions were sought, with the assumption that crucial variables are represented as terms ending in -ism (isms). 266 isms were extracted from a dictionary, and items were built from their definitions. In a sample of 500 college students, the most replicable item structure had 3 factors; one of these 3 factors split into 2 factors in the 4-factor structure. A replication study confirmed that Conservatism and Authoritarianism are subcomponents of the largest factor. The other factors, though highly meaningful, seem more sparsely represented in previous research. No factor was highly related to personality traits other than Openness to Experience. The factors can serve as content validity benchmarks for developing improved measurement models in this consequential, discrete domain of individual differences. PMID- 10707342 TI - Recent developments in selenium metabolism and chemical speciation: a review. AB - The biological roles of selenium and its mode of action have only recently begun to be revealed. To date, the major functions of selenium can be attributed to its antioxidative properties and its role in the regulation of thyroid hormone metabolism, cell growth and eicosanoid biosynthesis. The unusual feature of selenoprotein synthesis is that selenocysteine insertion is specified by the stop UGA codon. A number of selenocysteine-specific gene products and a stem-loop structure in the 3' untranslated region are required for selenocysteine biosynthesis and the decoding of UGA codons in the open reading frame of the mRNA. The major biological functions of selenium are achieved through its redox activity when present as selenocysteine at the active sites of selenoproteins and these proteins are selenium-dependent since replacement with the sulphur analogue cysteine causes loss of enzyme activity. Both organic and inorganic forms of selenium may be utilised by the body, with the selenoamino acids showing greatest bioavailability. Knowledge of the biochemistry of the element coupled with appropriate techniques for the study of the distribution of selenium species in health and disease could help to identify sensitive markers of selenium status. PMID- 10707343 TI - Comparative performance of pearl millet- and sorghum- based diets vs. wheat- and rice-based diets for trace metal bioavailability. AB - Pearl millet and sorghum offer a cheap source of energy compared to wheat and rice and are widely consumed by rural communities in many parts of the world. Due to the low consumption of vegetables and animal foods, millets also are the major suppliers of micronutrients especially for low-income groups. It is of prime importance to study how millets perform in terms of bioavailable contents of trace metals. Investigations were carried out using weanling mice which offer a model for the initial testing of bioavailability of trace metals before human trials. Four isocaloric diets differing only in the type of cereal, i.e. pearl millet, sorghum, wheat and rice, were prepared representing habitual dietary patterns observed by National Nutrition Monitoring Bureau (NNMB) of India. Mice were allocated randomly to 4 groups of 8 mice each, and housed individually in metal free metabolic cages. A fifth group of 8 mice fed a balanced synthetic diet served as control. All the groups were fed ad libitum. The absorption of zinc and iron averaged for 3 periods of 5 days each was significantly higher for the wheat and pearl millet group than for the other 2 experimental groups (p < 0.05), as were also the levels of liver zinc and iron. The weight gain was also highest (6.9 +/- 1.2 g) in the pearl millet group as compared to sorghum (1.58 +/- 0.59 g), wheat (1.66 +/- 1.27 g) and rice (-0.72 +/- 0.62 g) groups. The levels of liver copper were comparable in all the 5 groups. These results further confirm our earlier in vitro results indicating the superiority of pearl millet but not sorghum in bioavailability of zinc and iron. PMID- 10707344 TI - Epidemiological study of blood lead levels of children and adolescents living in Campania, Italy. AB - The aim of the present study is to evaluate blood levels (PbB) in a group of 500 (245 M, 255 F) children and adolescents of Campania (Italy) aged from 0.197 to 16.915 years, 269 (136 M, 133 F) of whom lived in urban zones and 231 (109 M, 122 F) in rural zones. PbB was assayed by electrothermal atomic absorption spectroscopy. The parents of the examined subjects children were interviewed about common risk factors for lead exposure using a standardized questionnaire. The PbB of children living in urban zones were significantly higher than the PbB of those living in rural zones (60.0 +/- 3.0 mg/L vs. 40.0 +/- 2.0 mg/L, p < 0.001). A PbB higher than 100 mg/L was found in 27 children (5.4%). We observed a significant correlation between age and PbB (p < 0.001, r = 0.529). Our data regarding children and adolescents demonstrate that the prevalence of PbB higher than 100 mg/L is greater in children living in urban areas (6.89%) than in subjects living in rural areas (3.89%). The findings can be explained by the higher presence of risk factors of Pb exposure in urban areas. Our data, if compared with those of previous studies concerning children of Campania, show a clear decrease of PbB. The correlation that we found between age and PbB indicates that long-term exposure at low doses more than a more intensive but short-term exposure seems to be important for the increase of blood lead levels. PMID- 10707345 TI - Altered dopamine turnover in murine hypothalamus after low-dose continuous oral administration of aluminum. AB - Aluminum, a known neurotoxic substance, has been suggested as a possible contributing factor in the pathogenesis of Alzheimer's disease. Ground-water pollution by aluminum has been recently reported. In the current study groups of 5 male BALB/c mice were administered aluminum ammonium sulfate in drinking water ad libitum at 0, 5, 25, and 125 mg/L aluminum for 4 weeks. At the termination of aluminum exposure, their brains were removed and dissected into cerebrum, cerebellum, medulla oblongata, midbrain, corpus striatum, and hypothalamus. The concentration of norepinephrine (NE), dopamine (DA), dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), serotonin (5-HT), and 5-hydroxyindoleacetic acid (5-HIAA), were determined in each brain area. DA, DOPAC, and HVA levels were lower in the hypothalamus of aluminum-treated mice, most notably in the low-dose group, as compared with control. No marked alterations in NE, 5-HT, and 5-HIAA levels were detected in any brain region. Changes in the concentration of DA and its metabolites measured in the hypothalamus suggest an inhibition of DA synthesis by aluminum. PMID- 10707346 TI - Bone-fracture incidence rate in two Italian regions with different fluoride concentration levels in drinking water. AB - The effect of the fluoride concentration in drinking water on the prevention of fractures related to osteoporosis has been questioned or contradicted in several recent studies. These studies have been mostly performed using water with artificially added fluoride, at the optimum level of about 1 mg/l. In the present study authors have investigated the effect of equal or greater fluoride concentrations (mean 1.45 mg/l) naturally present in waters supplied for human consumption to a population of 72.756 (Bracciano county), in comparison with a population of 126.189 (Avezzano county), supplied with low fluoride concentration water (mean 0.05 mg/l). The incidence of fractures in the years 1990 and 1991 was evaluated in the two areas (Bracciano and Avezzano), which are located in central Italy and where population have a similar life style, economic and social level and employment structure. The incidence data were obtained from the registers of the public hospital taken as a reference in each district. The authors noticed a significantly greater rate of fracture incidence at several parts of the body, in particular femur fractures (relative risks for males 4.28 and for females 2.64), in the population of the district of Avezzano than in the population of Bracciano. The greater concentration of fluoride in waters distributed for human consumption in Bracciano district seems to have the effect of protecting its inhabitants against fractures. PMID- 10707347 TI - Oral selenium supplementation in rats does not protect isolated rings of aorta against exogenous hydrogen peroxide. PMID- 10707348 TI - Determination of selenium in blood serum of children with acute leukemia and effect of chemotherapy on serum selenium level. AB - The concentration of selenium in serum was measured by the neutron activation method in three groups of children: 30 healthy children, 20 children with Acute Myeloblastic Leukemia (AML) and 40 with Acute Lymphoblastic Leukemia (ALL) (L1; n = 20, L2; n = 20). The samples were taken before and after induction chemotherapy. Age, sex, FAB, initial WBC, BUN, creatinine and urinary analysis did not show a significant change in the amount of selenium in serum. Selenium concentration in serum samples of ALL children before chemotherapy showed no significant differences as compared with that of normal individuals, but there were significant differences between children with AML and normal individuals (76.46 +/- 24.59 micrograms/L vs 102.38 +/- 19.25 micrograms/L, with p < 0.02). In conclusion, the question of whether these deficiencies are responsible for the disease or are the result of a secondary effect of the cancer remain to be answered. Immediately after induction chemotherapy, the selenium concentration in the serum of ALL children decreased significantly (80.14 +/- 15.48 micrograms/L vs 110.72 +/- 28.3 micrograms/L, p < 0.001), but this was not the case for AML children. These findings may be due to the difference in the drugs administered in induction chemotherapy of ALL and AML children. PMID- 10707349 TI - Review of publications. PMID- 10707350 TI - Virus recovery is induced in Brome mosaic virus p2 transgenic plants showing synchronous complementation and RNA-2-specific silencing. AB - Nicotiana benthamiana plants expressing Brome mosaic virus (BMV) p2 protein complemented replication of RNAs1 + 3 but, surprisingly, supported little or no replication of RNA-2. Despite this, the p2 transgenic plants were able to support systemic migration of RNAs-1 and -3. Kinetic analyses showed identical degradation rates for RNAs-2 and -3, greatly detracting from the concept of an induction of an RNA-2-specific degradation system. Deletion analysis identified a 200-nucleotide sequence that may contribute to silencing in a context-specific manner. When R1 progeny of a severely silencing p2 transgenic line were tested for virus resistance, three different classes of reactions were observed. In class 1 and class 3 plants, the virus moved systemically and showed various extents of RNA-2 silencing. However, in class 2 plants, there was a stochastic onset of post-transcriptional silencing in the systemic leaves that was reminiscent of virus recovery. Plants showing recovery tended to have a greater number of transgene loci than did those exhibiting component-specific silencing. The induction of silencing did not appear to be dependent solely on the combined steady state levels of the transgene and viral RNA. Some plants transformed with a p2 frameshift construct showed a complete silencing phenotype, but none showed RNA-2-specific silencing. While the relationship between the two types of silencing remains unclear, we speculate that our observations reflect early events in the induction of virus recovery. PMID- 10707351 TI - The hrpB and hrpG regulatory genes of Ralstonia solanacearum are required for different stages of the tomato root infection process. AB - hrp genes, encoding type III secretion machinery, have been shown to be key determinants for pathogenicity in the vascular phytopathogenic bacterium Ralstonia solanacearum GMI1000. Here, we show phenotypes of R. solanacearum mutant strains disrupted in the prhJ, hrpG, or hrpB regulatory genes with respect to root infection and vascular colonization in tomato plants. Tests of bacterial colonization and enumeration in tomato plants, together with microscopic observations of tomato root sections, revealed that these strains display different phenotypes in planta. The phenotype of a prhJ mutant resembles that of the wild-type strain. An hrpB mutant shows reduced infection, colonization, and multiplication ability in planta, and induces a defense reaction similar to a vascular hypersensitive response at one protoxylem pole of invaded plants. In contrast, the hrpG mutant exhibited a wild-type level of infection at secondary root axils, but the ability of the infecting bacteria to penetrate into the vascular cylinder was significantly impaired. This indicates that bacterial multiplication at root infection sites and transit through the endodermis constitute critical stages in the infection process, in which hrpB and hrpG genes are involved. Moreover, our results suggest that the hrpG gene might control, in addition to hrp genes, other functions required for vascular colonization. PMID- 10707352 TI - Heterologous rhizobial lipochitin oligosaccharides and chitin oligomers induce cortical cell divisions in red clover roots, transformed with the pea lectin gene. AB - Division of cortical cells in roots of leguminous plants is triggered by lipochitin oligosaccharides (LCOs) secreted by the rhizobial microsymbiont. Previously, we have shown that presence of pea lectin in transgenic white clover hairy roots renders these roots susceptible to induction of root nodule formation by pea-specific rhizobia (C. L. Diaz, L. S. Melchers, P. J. J. Hooykaas, B. J. J. Lugtenberg, and J. W. Kijne, Nature 338:579-581, 1989). Here, we report that pea lectin-transformed red clover hairy roots form nodule primordium-like structures after inoculation with pea-, alfalfa-, and Lotus-specific rhizobia, which normally do not nodulate red clover. External application of a broad range of purified LCOs showed all of them to be active in induction of cortical cell divisions and cell expansion in a radial direction, resulting in formation of structures that resemble nodule primordia induced by clover-specific rhizobia. This activity was obvious in about 50% of the red clover plants carrying hairy roots transformed with the pea lectin gene. Also, chitopentaose, chitotetraose, chitotriose, and chitobiose were able to induce cortical cell divisions and cell expansion in a radial direction in transgenic roots, but not in control roots. Sugar-binding activity of pea lectin was essential for its effect. These results show that transformation of red clover roots with pea lectin results in a broadened response of legume root cortical cells to externally applied potentially mitogenic oligochitin signals. PMID- 10707353 TI - Identification of Arabidopsis mutants exhibiting an altered hypersensitive response in gene-for-gene disease resistance. AB - A mutational study was carried out to isolate Arabidopsis thaliana plants that exhibit full or partial disruption of the RPS2-mediated hypersensitive response (HR) to Pseudomonas syringae that express avrRpt2. Five classes of mutants were identified including mutations at RPS2, dnd mutations causing a "defense, no death" loss-of-HR phenotype, a lesion-mimic mutant that also exhibited an HR- phenotype, and a number of intermediate or partial-loss-of-HR mutants. Surprisingly, many of these mutants displayed elevated resistance to virulent P. syringae and, in some cases, to Peronospora parasitica. Constitutively elevated levels of pathogenesis-related (PR) gene expression and salicylic acid were also observed. In the lesion-mimic mutant, appearance of elevated resistance was temporally correlated with appearance of lesions. For one of the intermediate lines, resistance was shown to be dependent on elevated levels of salicylic acid. A new locus was identified and named IHR1, after the mutant phenotype of "intermediate HR." Genetic analysis of the intermediate-HR plant lines was difficult due to uncertainties in distinguishing the partial/intermediate mutant phenotypes from wild type. Despite this difficulty, the intermediate-HR mutants remain of interest because they reveal potential new defense-related loci and because many of these lines exhibit partially elevated disease resistance without dwarfing or other apparent growth defects. PMID- 10707354 TI - Inheritance of partial resistance against Colletotrichum lindemuthianum in Phaseolus vulgaris and co-localization of quantitative trait loci with genes involved in specific resistance. AB - Anthracnose, one of the most important diseases of common bean (Phaseolus vulgaris), is caused by the fungus Colletotrichum lindemuthianum. A "candidate gene" approach was used to map anthracnose resistance quantitative trait loci (QTL). Candidate genes included genes for both pathogen recognition (resistance genes and resistance gene analogs [RGAs]) and general plant defense (defense response genes). Two strains of C. lindemuthianum, identified in a world collection of 177 strains, displayed a reproducible and differential aggressiveness toward BAT93 and JaloEEP558, two parental lines of P. vulgaris representing the two major gene pools of this crop. A reliable test was developed to score partial resistance in aerial organs of the plant (stem, leaf, petiole) under controlled growth chamber conditions. BAT93 was more resistant than JaloEEP558 regardless of the organ or strain tested. With a recombinant inbred line (RIL) population derived from a cross between these two parental lines, 10 QTL were located on a genetic map harboring 143 markers, including known defense response genes, anthracnose-specific resistance genes, and RGAs. Eight of the QTL displayed isolate specificity. Two were co-localized with known defense genes (phenylalanine ammonia-lyase and hydroxyproline-rich glycoprotein) and three with anthracnose-specific resistance genes and/or RGAs. Interestingly, two QTL, with different allelic contribution, mapped on linkage group B4 in a 5.0 cM interval containing Andean and Mesoamerican specific resistance genes against C. lindemuthianum and 11 polymorphic fragments revealed with a RGA probe. The possible relationship between genes underlying specific and partial resistance is discussed. PMID- 10707355 TI - Transgenic plants expressing geminivirus movement proteins: abnormal phenotypes and delayed infection by Tomato mottle virus in transgenic tomatoes expressing the Bean dwarf mosaic virus BV1 or BC1 proteins. AB - Transgenic tomato plants expressing wild-type or mutated BV1 or BC1 movement proteins from Bean dwarf mosaic virus (BDMV) were generated and examined for phenotypic effects and resistance to Tomato mottle virus (ToMoV). Fewer transgenic plants were recovered with the wild-type or mutated BC1 genes, compared with the wild-type or mutated BV1 genes. Transgenic tomato plants expressing the wild-type or mutated BV1 proteins appeared normal. Interestingly, although BDMV induces only a symptomless infection in tomato (i.e., BDMV is not well adapted to tomato), transgenic tomato plants expressing the BDMV BC1 protein showed a viral disease-like phenotype (i.e., stunted growth, and leaf mottling, curling, and distortion). This suggests that the symptomless phenotype of BDMV in tomato is not due to a host-specific defect in the BC1 protein. One transgenic line expressing the BC1 gene did not show the viral disease-like phenotype. This was associated with a deletion in the 3' region of the gene, which resulted in expression of a truncated BC1 protein. Several R0 plants, expressing either wild type or mutated BV1 or BC1 proteins, showed a significant delay in ToMoV infection, compared with non-transformed plants. R1 progeny plants also showed a significant delay in ToMoV infection, but this delay was less than that in the R0 parents. These results also demonstrate that expression of viral movement proteins, in transgenic plants, can have deleterious effects on various aspects of plant development. PMID- 10707356 TI - Changes in mRNA abundance within Heterodera schachtii-infected roots of Arabidopsis thaliana. AB - Gene expression changes in plant roots infected by plant-parasitic cyst nematodes are involved in the formation of nematode feeding sites. We analyzed mRNA abundance changes within roots of Arabidopsis thaliana during the early compatible interaction with Heterodera schachtii, the sugarbeet cyst nematode. Approximately 1,600 root sections, each containing a single parasitic nematode and its feeding site, and 1,600 adjacent, nematode-free root sections were excised from aseptic A. thaliana cultures 3 to 4 days after inoculation with H. schachtii. These tissue samples were termed infected and uninfected, respectively. Preparasitic nematodes were added to the uninfected tissue sample to maintain the nematode to plant tissue proportion. mRNA extracted from these two tissue samples was subjected to differential display analysis. Thirty-six cDNA clones corresponding to mRNA species with different abundance between both tissue samples were isolated. Of these clones, 24 were of A. thaliana origin and 12 were from H. schachtii. Differential display data predicted that the A. thaliana cDNA clones corresponded to 13 transcripts that were more abundant in the infected root sections and 11 transcripts that were more abundant in the uninfected root sections. H. schachtii cDNA clones were predicted to correspond to four transcripts that were more abundant in parasitic nematodes and to eight transcripts that were more abundant in preparasitic nematodes. In situ hybridization experiments confirmed the mRNA abundance changes in A. thaliana roots predicted by the differential display analyses for two A. thaliana clones. PMID- 10707357 TI - Effects of green fluorescent protein or beta-glucuronidase tagging on the accumulation and pathogenicity of a resistance-breaking Lettuce mosaic virus isolate in susceptible and resistant lettuce cultivars. AB - The RNA genome of a resistance-breaking isolate of Lettuce mosaic virus (LMV-E) was engineered to express the jellyfish green fluorescent protein (GFP) or beta glucuronidase (GUS) fused to the helper-component proteinase (HC-Pro) to study LMV invasion and spread in susceptible and resistant lettuce cultivars. Virus accumulation and movement were monitored by either histochemical GUS assays or detection of GFP fluorescence under UV light. The GFP- and GUS-tagged viruses spread systemically in the susceptible lettuce cultivars Trocadero and Vanguard, where they induced attenuated symptoms, compared with the wild-type virus. Accumulation of the GFP-tagged virus was reduced but less affected than in the case of the GUS-tagged virus. Systemic movement of both recombinant viruses was very severely affected in Vanguard 75, a lettuce cultivar nearly isogenic to Vanguard but carrying the resistance gene mo1(2). Accumulation of the recombinant viruses in systemically infected leaves was either undetectable (GUS-tag) or erratic, strongly delayed, and inhibited by as much as 90% (GFP-tag). As a consequence, and contrary to the parental virus, the recombinant viruses were not able to overcome the protection afforded by the mo1(2) gene. Taken together, these results indicate that GUS or GFP tagging of the HC-Pro of LMV has significant negative effects on the biology of the virus, abolishing its resistance-breaking properties and reducing its pathogenicity in susceptible cultivars. PMID- 10707358 TI - Identification with proteomics of novel proteins associated with the peribacteroid membrane of soybean root nodules. AB - Soybean peribacteroid membrane (PBM) proteins were isolated from nitrogen-fixing root nodules and subjected to N-terminal sequencing. Sequence data from 17 putative PBM proteins were obtained. Six of these proteins are homologous to proteins of known function. These include three chaperones (HSP60, BiP [HSP70], and PDI) and two proteases (a serine and a thiol protease), all of which are involved in some aspect of protein processing in plants. The PBM homologs of these proteins may play roles in protein translocation, folding, maturation, or degradation in symbiosomes. Two proteins are homologous to known, nodule-specific proteins from soybean, nodulin 53b and nodulin 26B. Although the function of these nodulins is unknown, nodulin 53b has independently been shown to be associated with the PBM. All of the eight proteins with identifiable homologs are likely to be peripheral rather than integral membrane proteins. Possible reasons for this apparent bias are discussed. The identification of homologs of HSP70 and HSP60 associated with the PBM is the first evidence that the molecular machinery for co- or post-translational import of cytoplasmic proteins is present in symbiosomes. This has important implications for the biogenesis of this unique, nitrogen-fixing organelle. PMID- 10707359 TI - Resistance gene analogs within an introgressed chromosomal segment derived from Triticum ventricosum that confers resistance to nematode and rust pathogens in wheat. AB - A resistance (R) gene-rich 2S chromosomal segment from Triticum ventricosum contains a cereal cyst nematode (CCN; Heterodera avenae) R gene locus CreX and a closely linked group of genes (Sr38, Yr17, and Lr37) that confer resistance to stem rust (Puccinia graminis f. sp. tritici), stripe rust (P. striiformis f. sp. tritici), and leaf rust (P. recondita f. sp. tritici) when introgressed into wheat. The 2S chromosomal segment from T. ventricosum is further delineated in translocations onto chromosome 2A of bread wheat, where the rust genes are retained but not the CreX gene. Using these critical genetic stocks, we have isolated family members of R gene analogs that are associated with either the 2S segment from T. ventricosum carrying the CreX locus or the rust genes. Derivatives of the Cre3 candidate R gene sequence and a rice (Oryza sativa) R gene analog that mapped to the 2S homologous chromosome groups in wheat were used to isolate related gene sequences from T. ventricosum that contain a nucleotide binding site-leucine rich repeat domain. The potential of these gene sequences as entry points for isolating candidate genes or gene family members of the CreX or rust genes and their further applications to plant breeding are discussed. PMID- 10707360 TI - Syringolin-mediated activation of the Pir7b esterase gene in rice cells is suppressed by phosphatase inhibitors. AB - Inoculation of rice plants (Oryza sativa) with the nonhost pathogen Pseudomonas syringae pv. syringae leads to the activation of defense-related genes and ultimately to induced resistance against the rice blast fungus Pyricularia oryzae. One of the molecular determinants of P. syringae pv. syringae that is recognized by the plant cells and evokes these defense responses is syringolin A, an elicitor that is secreted by the bacteria under appropriate conditions. In order to investigate signal transduction events elicited by syringolin A, the response of cultured rice cells to syringolin A application was analyzed. Cultured rice cells were able to sense syringolin A at concentrations in the nanomolar range as observed by the transient accumulation of Pir7b esterase transcripts. Syringolin A-mediated Pir7b transcript accumulation was inhibited by cycloheximide, indicating that de novo protein synthesis was required. Calyculin and okadaic acid, two protein phosphatase inhibitors, blocked Pir7b gene induction, whereas the serine/threonine protein kinase inhibitors staurosporine and K-252a had no effect on Pir7b transcript levels. Actin transcript levels were essentially not affected by inhibitor treatments over the experimental time span. These results imply that dephosphorylation of a phosphoprotein is an important step in the syringolin A-triggered signal transduction pathway. PMID- 10707361 TI - Differential induction of tobacco MAP kinases by the defense signals nitric oxide, salicylic acid, ethylene, and jasmonic acid. AB - In tobacco, two mitogen-activated protein (MAP) kinases, designated salicylic acid (SA)-induced protein kinase (SIPK) and wounding-induced protein kinase (WIPK) are activated in a disease resistance-specific manner following pathogen infection or elicitor treatment. To investigate whether nitric oxide (NO), SA, ethylene, or jasmonic acid (JA) are involved in this phenomenon, the ability of these defense signals to activate these kinases was assessed. Both NO and SA activated SIPK; however, they did not activate WIPK. Additional analyses with transgenic NahG tobacco revealed that SA is required for the NO-mediated induction of SIPK. Neither JA nor ethylene activated SIPK or WIPK. Thus, SIPK may function downstream of SA in the NO signaling pathway for defense responses, while the signals responsible for resistance-associated WIPK activation have yet to be determined. PMID- 10707362 TI - Pulmonary aspergillosis in an unselected autopsy series. AB - The present study was performed to analyze the relationship between underlying diseases and the morphologic form of aspergillosis. This retrospective analysis of 3284 autopsies yielded 18 cases of aspergillosis. The specific diagnosis of aspergillosis was rendered by a monoclonal antibody versus Aspergillus spp. Patients with hematological disorders, such as acute leukemia and aplastic anemia, made up about 35% of all patients dying of invasive aspergillosis. Diseases of the airways and the pulmonary parenchyma constituted the second most pathogenetic factor for the development of aspergillosis. The morphologic form of aspergillosis was closely related to the underlying diseases. Non- and semi invasive forms of aspergillosis--saprophytic infection and chronic necrotizing aspergillosis--were observed only in patients with an isolated underlying pulmonary disease devoid of any other precipitating factor. In contrast, seven patients, five of whom suffered from hematological diseases, had no underlying lung disease and developed aspergillus pneumonia. The remaining 5 patients with aspergillus pneumonia showed a combination of underlying extrapulmonary disease and pulmonary alterations that preceded aspergillosis. The local distribution of fungal infection showed a characteristic distribution pattern with a predominance of the upper lung lobes. Hematogeneous spread beyond the lungs occurred exclusively in cases with aspergillus pneumonia. We conclude that the different forms of aspergillosis are closely related to the nature of the underlying disease. PMID- 10707363 TI - Association of concentration of asbestos and asbestos-like fibers with the patient's survival and the binding capacity of lung parenchyma to galectin-1 and natural alpha-galactoside- and alpha-mannoside-binding immunoglobulin G subfractions from human serum. AB - Our aim in this study was to search for lung parenchyma alterations associated with asbestos and/or asbestos-like fiber concentration. This was done by means of immuno- or glycohistochemistry. The hot-ashing technique determined the asbestos and asbestos-like fiber concentrations in the lung tissues of 100 patients of whom 52 were treated for primary lung and 25 for secondary lung tumors; fiber concentration was also measured for 23 patients whose disease was benign. The results were correlated to smoking habits, survival of the patients, and expression of binding capacities for endogenous lectins, natural carbohydrate binding and lectin-specific antibodies. The cohort with proven asbestos exposure revealed a mean fiber concentration 114 f/g compared to 95 f/g in the non-exposed group. An increased asbestos fiber concentration was correlated to galectin-1 binding and the presence of epitopes for natural immunoglobulin G subfractions with selectivity to alpha-galactosides and alpha-mannosides. The survival of patients with primary and secondary lung tumors was negatively associated with the fiber concentration. The data indicate that increased presence of asbestos is correlated with an alteration of defined glycohistochemical features of alveolar lining cells. PMID- 10707364 TI - Histological features of resolving acute, non-complicated phlegmonous appendicitis. AB - The histological features of resolving acute appendicitis are described. Formalin fixed, paraffin-embedded appendices of 200 cases with acute, non-complicated phlegmonous appendicitis were reviewed. In 80 out of 200 cases, a histological picture characterized by a predominantly lymphocytic infiltrate of the subserosa and muscularis propria or the subserosa alone was found. In the affected muscularis propria, eosinophils were admixed with lymphocytes, and the cellular infiltrate showed a lesser degree than that of the classic phlegmonous appendicitis. A multifocal, rather than a diffuse pattern of infiltration was observed. Cases were divided into three groups. Group 1: appendices with the typical features of phlegmonous appendicitis: 120 cases, 60%. Group 2: appendices with a predominantly lymphocytic infiltrate in the muscularis propria, subserosa, or both, and no granulation tissue: 65 cases, 32.5%. Group 3: appendices with granulation tissue: 15 cases, 7.5%. Complicated appendicitis was excluded. Data on the duration of the clinical symptoms were derived from the clinical history. The differences between the mean duration time of groups 1 and 2, and of groups 2 and 3 were statistically significant. The findings support the contention that a mixed infiltrate of lymphocytes and eosinophils represents a regression phase of acute appendicitis. PMID- 10707365 TI - Clinicopathologic study and classification of vocal cord cysts. AB - Cysts of the true vocal cords are less common than other laryngeal cysts. They are usually easily recognized and managed. Patients present with complaints of hoarseness and/or dyspnea. We report our experience with 41 cases of cysts located in the true vocal cords. Clinical and histological aspects are reviewed and discussed. A new histological classification is proposed: A: cysts lined by columnar epithelium with mucous content; B: lined by columnar epithelium with cilia; C: lined b squamous epithelium without keratinization; D: lined by squamous epithelium with keratinization. PMID- 10707366 TI - Angiolymphoid hyperplasia with eosinophilia: report of a lesion mimicking soft tissue tumor of extremely long duration. AB - We present a large sized lesion of the right upper arm in which characteristics of the angiolymphoid hyperplasia with eosinophilia (ALHE) intermingled with those of Kimura's disease (KD). The laboratory findings, the prominent vascular proliferation and the features of endothelial cells were suggestive of ALHE. However, the long duration of the disease, the site of involvement, the abundant lymphoid component forming lymph follicles with germinal centers and the fibrosis are features of KD. In agreement with other reports, our case shows that clinicopathologically there is an overlap between ALHE and KD. PMID- 10707367 TI - Overexpression of Ets-1 transcription factor in angiosarcoma of the skin. AB - Angiosarcoma of the skin is a rare malignant tumor which is slow-growing but highly aggressive and often recurs following surgery and/or radiation therapy, finally metastasizing to the regional lymph nodes. The ets-1 protooncogene is shown to be transcribed in endothelial cells during angiogenesis in granulation tissue and in malignant cells during tumor invasion. Furthermore, it can regulate the expression of metalloproteinase genes such as collagenase-1 (MMP-1), stromelysin (MMP-3) and urokinase-type plasminogen activator (uPA). In this study we investigated the ets-1 and MMP-1 expression in 7 angiosarcomas of the skin, compared with 7 hemangiomas and 7 granuloma pyogenicums of the skin, which are well known as benign vascular diseases. The ets-1 and MMP-1 mRNAs and their proteins were overexpressed in all angiosarcomas tested, and the localization of MMP-1 expression corresponded to that of ets-1. On the other hand, they were weakly or not at all expressed in hemangiomas and granuloma pyogenicums. These results suggest that the constitutive overexpression of ets-1 might be closely related with the malignant progression of angiosarcoma, possibly through the up regulation of the transcription of MMP-1. PMID- 10707368 TI - Comparison of two stereological methods for quantitative renal morphology: a modified fractionator and modified Weibel-Gomez method. AB - To determine the best method for quantitative analysis of renal structure, we compared two stereological techniques: the unbiased fractionator method and the model-based method described by Weibel-Gomez. Kidneys of 20 week old pregnant female Sprague Dawley rats were investigated. Using these two stereological techniques, the mean glomerular volume and the total glomerular number per kidney were determined in 16 kidneys of 8 rats. Both methods were used in a modified way to correct for variations in section thickness (fractionator) and to reduce the length of the measuring process (Weibel-Gomez), respectively. The mean glomerular volume was comparable in both methods (Fractionator: 5.49 +/- 0.56 x 10(4) mm3, Weibel-Gomez: 5.35 +/- 0.34 x 10(4) mm3). In contrast, using the Weibel-Gomez method (43,774 +/- 2338), the total number of glomeruli per kidney was significantly higher than that obtained by the fractionator technique (39,359 +/- 4250). The results as well as the time necessary for each method and the practicability of the techniques were compared. In our hands, apart from the respective advantages and disadvantages, the Weibel-Gomez technique is easier to perform and much more efficient than the probably more elegant fractionator method. The bias problem of the Weibel-Gomez method does not play an important role with respect to the most common biological problems. PMID- 10707369 TI - Immunocytochemical investigations of heat shock proteins expression during thymic apoptosis induced by glucocorticoids. AB - The aim of our study was to investigate a possible expression of different HSPs in rat's thymuses after hydrocortisone administration. The thymuses of 41 young rats (25 to 45 days age old) were studied immunocytochemically: 12 rats were not injected, 8 received an injection of physiological serum, and 21 received HC (125 mg/kg). HSP27, 70 and 110 expression was investigated following the PAP method. HSPs27 were expressed neither in normal thymic lobules nor in the cortical thymic cells after HC injection. HSPs70 were objectivated only in 1 control animal, but were frankly expressed in cortical thymic cells 1 to 48 hours after HC injection and remained significantly expressed until the 7th day after HC injection. HSPs 110 were present in only 1 control animal and appeared to be distinctly expressed 48 hours after HC injection. HSPs 70 and 110 were never expressed in the regenerated thymuses 14 and 21 days after HC injection. This report objectivates for the first time 70 and 110 kDa "stress proteins" expression during the thymic apoptosis induced by glucocorticoids. PMID- 10707370 TI - Superficial gastric carcinoma developed on localized hypertrophic lymphocytic gastritis: a variant of localized Menetrier's disease? AB - Menetrier's disease is a rare premalignant condition that usually involves the entire stomach. Only few cases of localized disease have been reported, rarely with cancer. Lymphocytic gastritis is a newly described entity that may share a common pathogenesis with Menetrier's disease. The authors report the case of a 62 year old woman with known liver cirrhosis in whom endoscopic examination of the stomach showed an antral tumor. Examination of the surgical specimen showed a superficial gastric adenocarcinoma developed on an hypertrophic gastropathy with both Menetrier's disease and lymphocytic gastritis features. This observation strengthens the hypothesis of a common mechanism between Menetrier's disease and lymphocytic gastritis, which may be part of the same disease spectrum. This disease could also correspond to the "hypertrophic lymphocytic gastritis" recently described. PMID- 10707371 TI - Detection of pulmonary metastasis of low-grade endometrial stromal sarcoma 25 years after hysterectomy. AB - Endometrial stromal sarcoma (ESS) is a rare uterine sarcoma. Low-grade ESS occasionally recurs or metastasizes after long disease-free periods, a fact that may complicate the diagnosis. Here we report a case of multiple lung metastases in a 68-year-old woman who had been disease-free for 25 years after hysterectomy for a uterine tumor. Biopsy revealed that the tumor was composed of oval cells with slight nuclear atypism but without mitotic figures, suggesting a low-grade neoplasm. Immunostaining for intermediate filaments revealed strong positivity for vimentin and weak positivity for alpha-smooth muscle actin. In addition, immunostaining for estrogen and progesterone receptors, performed under suspicion of low-grade ESS, was positive. The uterine tumor resected many years before had shown a similar morphology. Thus, it was demonstrated that the lung neoplasm was a metastatic low-grade ESS that had appeared after many disease-free years. A review of the literature revealed that this case had the longest recorded interval between the occurrence of the initial ESS and the development of distant metastases. When low-grade sarcoma appears in the lungs of female patients, it is important to consider the possibility of low-grade ESS. Detailed information on the past clinical history, together with immunostaining for estrogen and progesterone receptors, are important diagnostic keys. PMID- 10707372 TI - Adult mesoblastic nephroma. AB - We report a case of asymptomatic mesoblastic nephroma in a 54-year-old woman. The tumor showed immunohistochemical reactions similar to developing nephrons. Electron microscopy showed immature tubules with numerous intracytoplasmic intermediate filaments. Recent studies support the concept of pathogenesis of the mesoblastic nephroma originating from collecting ducts. However, this case exhibited a complex pattern of antigenic expression not restricted to the collecting ducts, but including the glycoprotein CD24 and the neural cell adhesion molecule (NCAM). The following differential diagnoses will be discussed: benign mixed epithelial and stromal tumor, metanephric adenoma, and nephrogenic adenofibroma. PMID- 10707373 TI - Quality through metrics. AB - The Quality Assurance Unit analyzed 18 months of departmental data regarding the report-audit cycle. Process mapping was utilized to identify milestones in the cycle for measurement. Five milestones were identified in the audit cycle, as follows: (1) time from report receipt in quality assurance to start of audit, (2) total calendar days to audit a report, (3) actual person-hours to perform a report audit, (4) time from completion of audit to issuance of report, and (5) total time a report is in quality assurance. An interrelationship diagraph is a quality tool that is used to identify what activities impact the overall report auditing process. Once the data collection procedure is defined, a spreadsheet is constructed that captures the data. The resulting information is presented in time charts and bar graphs to visually aid in interpretation and analysis. Using these quality tools and statistical analyses, the Quality Assurance Unit identified areas needing improvement and confirmed or dispelled previous assumptions regarding the report-auditing process. By mapping, measuring, analyzing, and displaying the data, the overall process was examined critically. This resulted in the identification of areas needing improvement and a greater understanding of the report-audit cycle. A further benefit from our increased knowledge was the ability to explain our findings objectively to our client groups. This sharing of information gave impetus to our clients to examine their report-generation process and to make improvements. PMID- 10707374 TI - Quality beyond compliance. AB - The service sector within the biopharmaceutical industry has experienced phenomenal growth over the past decade. In the highly regulated Good Laboratory Practices environment, the need for timely, high-quality service, accurate results, and on-time deliverables becomes paramount for the success and profitability of biopharmaceutical companies. The quality assurance process is a vital component of this drug product-development cycle and ensures compliance to the highest domestic and international regulatory standards. Quality-assurance professionals historically have held the role of independent auditors of the processes, who certify that results meet current standards of practice. Covance, a contract research organization that includes Good Laboratory Practices laboratories, reorganized and expanded the functional responsibilities of its quality assurance team in 1997. Auditors and quality assurance professionals have assumed roles beyond traditional compliance auditing and are forging new leadership and mentoring roles as process-improvement specialists. The results have been tangible, measurable benefits for clients and the Covance organization. This article provides an overview of this cultural change and the processes put in place to improve efficiency, productivity, and customer and employee satisfaction. PMID- 10707375 TI - Teaching fractional factorial experiments via course delegate designed experiments. AB - Industrial experiments are fundamental in enhancing the understanding and knowledge of a process and product behavior. Designed industrial experiments assist people in understanding, investigating, and improving their processes. The purpose of a designed experiment is to understand which factors might influence the process output and then to determine those factor settings that optimize the process output. Teaching "design of experiments" using textbook examples does not fully shed light on how to identify and formulate the problem, identify factors, and determine the performance of the physical experiment. Presented here is an example of how to teach fractional factorial experiments in a course on designed experiments. Also presented is a practical, hands-on experiment that has been found to be extremely successful in instilling confidence and motivation in course delegates. The experiment provides a great stimulus to the delegates for the application of experimental design in their own work environment. PMID- 10707376 TI - A review of the FDA draft guidance document for software validation: guidance for industry. AB - A Draft Guidance Document (Version 1.1) was issued by the United States Food and Drug Administration (FDA) to address the software validation requirement of the Quality System Regulation, 21 CFR Part 820, effective June 1, 1997. The guidance document outlines validation considerations that the FDA regards as applicable to both medical device software and software used to "design, develop or manufacture" medical devices. The Draft Guidance is available at the FDA web site http:@www.fda.gov/cdrh/comps/swareval++ +.html. Presented here is a review of the main features of the FDA document for Quality System Regulation (QSR), and some guidance for its implementation in industry. PMID- 10707377 TI - History of the Korean GLPs and the activities and perspectives of the Korean Society of GLP. AB - The Korean Society of Good Laboratory Practice (KSGLP) was established Dec. 10, 1998. The objectives of the KSGLP are to enhance the quality of domestic studies and the level of GLP compliance, in safety testing, and to promote information exchange among its members. The activities of KSGLP include: offering workshops and symposiums, linking with related governmental organizations, collecting GLP related information and providing the information to the related organizations, building international networks to collect information and to establish relationship, developing training materials and publishing periodicals, and other business necessary to achieve the objectives of the KSGLP. The KSGLP achieved its goals within a short period of time by offering workshops and symposia, and by providing important GLP related information in newspapers or via the KSGLP's internet homepage (www.ksglp.or.kr). The main role of the KSGLP will be to disseminate GLP technology nationwide. The KSGLP would like to help many labs that are preparing their facilities for GLP compliance. Further, the KSGLP is hoping to share GLP experiences with other members. PMID- 10707379 TI - [Analyses of work-relatedness of health problems among truck drivers by questionnaire survey]. AB - In order to estimate occupational risk factors for health problems among truck drivers, a questionnaire survey of working conditions, job content in truck transportation, subjective symptoms and present illnesses was carried out among 541 truck transportation workers in 1997. The valid response rate was 85.7%, and 134 local truck drivers, 199 long-distance truck drivers and 71 clerical workers were analyzed. First, to examine occupational risk factors and health problems among the three groups, the authors analyzed working conditions, job content in truck transportation, subjective symptoms and present illnesses. Second, to estimate the work-relatedness of health problems among local truck drivers and long-distance truck drivers, logistic regression analyses were conducted, and odds ratios and 95% confidence intervals were computed. The prevalence rates of working factors affecting health problems of truck drivers were significantly higher than those of clerical workers in the items on irregular shift work, working environment, working posture, handling heavy materials, job stress due to overloading and long working time and limited time off. The prevalence rates for subjective symptoms (ringing in the ears, neck pain and low back pain) and present illnesses (hypertension, ulcers in the digestive tract, back injuries, whiplash injuries and hemorrhoids) among truck drivers were significantly higher than those of clerical workers. In logistic regression analyses, many work related items except age, BMI and smoking habit showed significantly higher odds ratios for subjective symptoms and present illnesses of truck drivers. Odds ratios for hypertension, heart diseases and related subjective symptoms among local truck drivers were significantly increased by job career, twisting posture, vibration and driving stress. Odds ratios for gastro-duodenal diseases and related subjective symptoms were significantly increased by narrow working space, sleeping in the truck, driving distance, squatting posture and driving stress. Odds ratios for ringing in the ears among local truck drivers were significantly increased by job career, long working time, narrow working space, sleeping in the truck and driving stress. Odds ratios for musculo-skeletal diseases and related subjective symptoms were significantly increased by overwork, vibration, narrow working space, sitting posture and shortage of recess. Odds ratios for fatigue symptoms were significantly increased by the shortage of recess, vibration and driving stress. In order to cope with the health problems of truck drivers, it is recommended that working conditions and work loads for among truck drivers as described above be improved. PMID- 10707378 TI - [The effective smoking corner in an office]. AB - We installed an effective, practical and low-cost smoking corner in an office to protect against passive smoking. The smoking corner was separated from the non smoking area with transparent, nonflammable screens. Four exhaust fans were installed in the smoking corner so that there was no leakage of environmental tobacco smoke. The required exhaust air rate in the smoking corner was pre calculated from the volume of the smoking corner and the rate of consumption of cigarettes. The suspended airborne particle concentration definitely decreased in the non-smoking area after installation of the smoking corner. The result of a questionnaire survey also revealed the improvement in the air quality in this office. PMID- 10707380 TI - [The workload of computer system engineers and mental health]. AB - The nature of labor in Japan is changing from work which is physically demanding to work in which mental activity requiring intellectual and psychological ability results in production. The author studied and analyzed the type of workload experienced by system engineers in informatics. The results of a health study of 2,396 system design engineers in informatics at our company were analyzed (only data on 2,169 males were analyzed). Workload related to the 'quality', or type of work done ('Workload of quality') was felt by 63.5%, with 36.0% feeling workload related to the amount of work ('Workload of quantity'). A very large number of workers felt workload related to the quality, or type of work rather than to the amount of work. The relationship between emotional health indices and workload showed insomnia in 25.5% of workers who felt a 'Workload of quality', and in 11.1% of those who did not feel such a workload. Similarly, depressive mood was seen in 40.3% of workers who felt a 'Workload of quality', and in 18.7% of workers who did not. Insomnia was seen in 42.8% of workers with a feeling of a 'Workload of quality', and in 26.3% of those without. The relationship between workload, related to the quality and quantity of work was also strong for other mental health indices. Workload related to the quality of work was particularly strong. Workload related to the quality or type or work comes from a myriad of factors including the variety of work required from workers in informatics, the still developing nature of the production process, and the rapid pace of technical progress with each type of work carrying a different set of factors. The ability and personality of each individual worker also plays an important role. The results of the survey showed the importance of carrying out occupational healthcare activities in health management from the standpoint of work related health hazards or "quality of work"; in other words, from the standpoint of work type, system operation and the individuality of the worker. PMID- 10707381 TI - [Relationship between fatty liver and coronary risk factors]. AB - Fatty liver is a common finding in abdominal ultrasonographic examination in health check-ups, but the relationship between fatty liver and so-called coronary risk factors has rarely been investigated from the viewpoint of prevention of coronary heart disease. The purpose of the present study was to elucidate such a relationship by comparing the coronary risk factors with and without fatty liver by using data from health check-ups for the mid-management and management staff of a manufacturing company. The majority (77.1%) of those with fatty liver in the present study were categorized as "normal" or "marginally obese" and only a small portion (22.9%) were categorized as "obese" according to the classification of the body mass index. The group of subjects with fatty liver had significantly lower mean HDL-cholesterol and higher levels of fasting blood sugar, HDL/total cholesterol ratio, triglyceride, uric acid and transaminases, than those parameters in subjects without fatty liver, even after adjustment for age and body-mass index. The blood pressure (both systolic and diastolic) and total cholesterol level did not show any significant difference after controlling the covariates. Our results indicated that fatty liver has a close correlation with the majority of coronary risk factors causing atheroscleotic diseases, and most of these relationships are independent of total body mass. Our results regarding fatty liver are a help to occupational health personnel when advising workers to reduce their own risk of atherogeic diseases. PMID- 10707382 TI - Historical perspectives 2000. PMID- 10707383 TI - A novel spinal implant infection model in rabbits. AB - STUDY DESIGN: A new spinal implant model was designed to study device-centered infection with methicillin-resistant Staphylococcus aureus in multiple noncontiguous surgical sites in the lumbar spine region of a rabbit. OBJECTIVE: To develop a multiple-site spinal implant device-centered infection model in rabbits. SUMMARY OF BACKGROUND DATA: Results in many recent studies show that postoperative wound infection after spinal implant surgery and the increase in antibiotic-resistant bacteria are a concern. Anti-infection strategies must be tested in relevant animal models that will lead to appropriate clinical studies. METHODS: Eight anesthetized New Zealand White rabbits underwent completely isolated partial laminectomy and subsequent stainless steel Kirschner wire implantation directly into the transverse processes of vertebrae T13, L3, and L6. The middle sites (L3) were used as sterile control sites, and the outer sites (T13, L6) were challenged with different amounts of methicillin-resistant Staphylococcus aureus. Rabbits were killed after 7 days, and biopsies were performed to provide evidence for device-centered infection. Bacterial growth on the implant surfaces and in surrounding tissues and bone was assayed. RESULTS: Overall device-centered infection was established after 7 days in 100% of the sites challenged with 10(3) colony-forming units methicillin-resistant Staphylococcus aureus or higher. No infection was seen in any of the control sites located between infected vertebrae. Multiple blood and liver samples showed that the separate localized infections did not become systemic after 7 days. CONCLUSIONS: This new animal model demonstrates that multiple biomaterial implants can be evaluated in the same animal and provides a technique for investigating postoperative device-centered infection of the spine. Infection was demonstrated in noncontiguous lumbar sites of the spine, whereas adjacent control sites remained sterile. Because there was no cross contamination or systemic spread of the infection, multiple anti-infection strategies or implant materials can now be tested for efficacy in a single animal to combat dramatic and costly postoperative implant infections. PMID- 10707384 TI - Electrical stimulation of the rat lumbar spine induces reflex action potentials in the nerves to the lower abdomen. AB - STUDY DESIGN: The distribution of the nerve action potentials reflexively elicited by electrical stimulation of the lumbar spine was investigated in rats. OBJECTIVES: To elucidate the relation between the lumbar spine and other body regions that compose the spinal reflex. SUMMARY OF BACKGROUND DATA: The hypothesis was that the ventral portion of the L5-L6 disc spatially corresponds to the groin. METHODS: In Experiments 1 and 2, wire electrodes were placed 1) in the ventral and dorsal portions of the disc, facet joint, and muscle fascia at L5 L6, and 2) in the ventral portions of L3-L4, L4-L5, L5-L6, and L6-S discs. A needle electrode was inserted in the L5-L6 disc by 0.4-mm increments, and action potentials were serially recorded from the genitofemoral nerve. RESULTS: Experiments 1 and 2: Reflex action potentials were elicited in the iliohypogastric (T13 and L1), ilioinguinal (L1), and genitofemoral (L2) nerves. Experiment 1: Stimulation of the disc induced reflex discharges significantly more frequently than stimulation of the facet joint and muscle fascia. Experiment 2: The more cranial the disc stimulated, the more frequently the reflex discharge was induced in the iliohypogastric nerve. Experiment 3: The depth of stimulation did not influence the size of the reflex action potential. CONCLUSIONS: Electrical stimulation of the lumbar disc and facet joint induced reflex discharges in the nerves to the lower abdominal regions. It was postulated that the reflex discharges are related to muscle contraction resulting in referred pain in the loin and groin. PMID- 10707385 TI - An experimental study of the effects of nerve root retraction on the posterior ramus. AB - STUDY DESIGN: The histologic and ultrastructural changes in the posterior ramus after posterior lumbar surgery were studied in rabbits. OBJECTIVE: To investigate the structural changes in the posterior ramus after posterior lumbar surgery that may cause injury to the posterior ramus after the procedure. SUMMARY OF BACKGROUND DATA: Investigators in previous studies have pointed out that low back discomfort after lumbar discectomy relates to neurogenic changes and/or myogenic changes of paravertebral muscle. However, no previous study has demonstrated the effects of excessive nerve root retraction on spinal posterior rami. METHODS: Eighteen male Japanese White rabbits were used. The posterior ramus arising from the S1 nerve root was examined after exposure of the lamina only, fenestration, or retraction of the S1 nerve root, with light microscopy and transmission electron microscopy at 2, 4, and 6 weeks after the procedure. Results were compared with a those in control specimens that did not undergo the procedure. RESULTS: In the exposed group, no distinct difference was found compared with the control specimen. In the fenestration group, especially at 6 weeks, some attenuation and splitting of myelin sheaths was observed. In the retraction group, the structural alteration was most severe. Even at 2 weeks, fragmentation of many myelin sheaths was detected. Examination of specimens by electron microscopy indicated phagocytosis of myelinated fibers at 4 and 6 weeks. CONCLUSIONS: Findings showed that posterior lumbar procedures, including retraction of paravertebral muscle, fenestration of the lamina, and retraction of the nerve root affect the posterior ramus. Excessive retraction of the nerve root has an especially disastrous effect on the posterior ramus. Such a violent maneuver within the spinal canal must be avoided. PMID- 10707386 TI - Posterior lumbar interbody fusion using posterolateral placement of a single cylindrical threaded cage. AB - STUDY DESIGN: An in vitro biomechanical study of posterior lumbar interbody fusion (PLIF) with threaded cages was performed on 18 bovine lumbar functional spinal units. OBJECTIVES: To compare the segmental stiffness among PLIF with a single long posterolateral cage, PLIF with a single long posterolateral cage and simultaneous facet joint fixation, and PLIF with two posterior cages. SUMMARY OF BACKGROUND DATA: In most cases, PLIF with threaded cage techniques needs bilateral facetectomy, extensive exposure, and retraction of the cauda equina. Posterior element deficiency is detrimental to postoperative segmental stiffness. METHODS: All specimens were tested intact and with cage insertion. Group 1 (n = 12) had a long threaded cage (15 x 36 mm) inserted posterolaterally and oriented counter anterolaterally on the left side by posterior approach with left unilateral facetectomy. Group 2 (n = 6) had two regular-length cages (15 x 24 mm) inserted posteriorly with bilateral facetectomy. Six specimens from Group 1 were then retested after unilateral facet joint screw fixation in neutral (Group 3). Similarly, the other six specimens from Group 1 were retested after fixation with a facet joint screw in an extended position (Group 4). Nondestructive tests were performed in pure compression, flexion, extension, lateral bending, and torsion. RESULTS: The PLIF procedure involving a single cage (Group 1) had a significantly higher stiffness than PLIF with two cages (Group 2) in left and right torsion (P < 0.05). Group 1 had higher stiffness values than Group 2 in pure compression, flexion, and left and right bending, but differences were not significant. Group 3 had a significant increase in stiffness in comparison with Group 1 for pure compression, extension, left bending, and right torsion (P < 0.05). For Group 4, the stiffness significantly increased in comparison with Group 1 for extension, flexion, and right torsion (P < 0.05). Although there was no significant difference between Groups 3 and 4, Group 4 had increased stiffness in extension, flexion, right bending, and torsion. CONCLUSIONS: Posterior lumbar interbody fusion with a single posterolateral long threaded cage with unilateral facetectomy enabled sufficient decompression while maintaining most of the posterior elements. In combination with a facet joint screw, adequate postoperative stability was achieved. PMID- 10707387 TI - Effects of backward bending on lumbar intervertebral discs. Relevance to physical therapy treatments for low back pain. AB - STUDY DESIGN: Mechanical testing of cadaveric motion segments. OBJECTIVES: To test the hypothesis that backward bending of the lumbar spine can reduce compressive stresses within lumbar intervertebral discs. SUMMARY OF BACKGROUND DATA: Lumbar extension affects the distribution of compressive stress inside normal cadaveric discs, but little is known about its effect on mechanically disrupted and degenerated discs. METHODS: Nineteen lumbar motion segments (mean donor age, 48 years) were subjected to complex mechanical loading to simulate the following postures: moderate lumbar flexion, 2 degrees of extension, 4 degrees of extension, and the neutral position (no bending). The distribution of compressive stress within the disc matrix was measured in each posture by pulling a miniature pressure transducer along the midsagittal diameter of the disc. Stress profiles were repeated after a mechanical treatment that was intended to simulate severe disc degeneration in vivo. RESULTS: The "degeneration" treatment reduced pressure in the nucleus pulposus and generated stress concentrations within the anulus, in a manner similar to that found in severely degenerated discs in vivo. When all discs were considered together, 2 degrees of extension increased the maximum compressive stress within the posterior anulus by an average of 16%, compared with the neutral posture. The size of localized stress peaks within the posterior anulus was increased by 43% (P = 0.02). In 4 degrees of extension, changes observed between 0 degree and 2 degrees were usually exaggerated. In contrast, moderate flexion tended to equalize the distribution of compressive stress. In 7 of the 19 discs, 2 degrees of lumbar extension decreased maximum compressive stress in the posterior anulus relative to the neutral posture by up to 40%. Linear regression showed that lumbar extension tended to reduce stresses in the posterior anulus in those discs that exhibited the lowest compressive stresses in the neutral posture (P = 0.003; R2 = 41%). CONCLUSIONS: The posterior anulus can be stress shielded by the neural arch in extended postures, but the effect is variable. This may explain why extension exercises can relieve low back pain in some patients. PMID- 10707388 TI - The clinical presentation of uppermost cervical disc protrusion. AB - OBJECT: The purpose of this study is to clarify the clinical presentation of the C2-C3 cervical herniation disc. SUMMARY OF BACKGROUND DATA: Uppermost cervical disc protrusion is an uncommon condition. The pattern of large central fragments of nucleus impinging on the highest cervical disc region is often poorly localized according to its clinical presentation. METHODS: Eight patients treated with anterior cervical discectomy with fusion for C2-C3 disc herniation participated in a detailed clinical and radiologic review to determine early detection and clarify potential hazards. Each patient's neurologic function was tested and recorded successively by a team of physicians and qualified physiotherapists. RESULTS: Reviewing the symptomatology, most patients presented ascending radicular symptoms secondary to trivial trauma, characterized by suboccipital pain, loss of hand dexterity, and paresthesia over face and unilateral lateral arm. Six (75%) patients had remarkable improvement postoperatively in neurologic function, except for some residual sensory embarrassment in at least 6 months follow-up. CONCLUSIONS: Clinical neurologic examination provides a less precise anatomic basis, to point to a particular upper cervical disc protrusion. Nonspecific neck and shoulder pain, a variety of cervical radiculopathy, and myelopathy may present. However, this rare spondylotic pattern is usually characterized by impairment of motor and sensory function more in the upper extremities than lower extremities and mostly starting following trauma. Radiculopathy generally outweighs the cord sign. Cruciate paralysis associated with vague diffuse and patch regions of hypesthesia over perioral distribution may help to localize this upper cervical lesion. The present study demonstrates that early detection and adequate anterior decompression may provide excellent outcome. PMID- 10707389 TI - Noninvasive three-dimensional analysis of cervical spine motion in normal subjects in relation to age and sex. An experimental examination. AB - STUDY DESIGN: Experimental examination in vivo. OBJECTIVES: To determine the precision of the ultrasound-based Coordinate Measuring System (CMS 50; Zebris Medizintechnik GmbH, D-88316, Isny, Germany) and then to establish a reference range for the active range of motion of the cervical spine in normal test subjects grouped according to age and sex. SUMMARY OF BACKGROUND DATA: Many different devices such as inclinometers, goniometers, potentiometers, computer aided devices, and radiographic procedures have been developed to examine the range of motion of the cervical spine. All of them have more or less inherent limitations. METHODS: To assess the precision of this examination method, preliminary experiments were performed including intraobserver retest and two observer repeatability, intraindividual variability, a daily profile, and a comparison between active and passive motions. In the subsequent main experiment 157 persons (86 women and 71 men) were examined during active motion. The sex groups were further subdivided into age groups of 10 years each. A comparison of weight and athletic activity was also performed. RESULTS: The range of motion decreased with increasing age, increasing body weight, and decreasing athletic activity. The rotation in the upper cervical spine increases with age to compensate for the reduced range of motion in the lower levels. Women showed significantly better mobility than men of the same age, only above the age of 70. CONCLUSIONS: The CMS 50 device provides precise reproducible measurements of the active range of motion of the cervical spine in all three planes. Criteria such as age, sex, body weight, and athletic activity influence the range of motion of the cervical spine. PMID- 10707390 TI - Clinical and radiographic evaluation of disc excision for lumbar disc herniation with and without posterolateral fusion. AB - STUDY DESIGN: A prospective study evaluating the clinical and radiographic results in 95 patients with lumbar disc herniation. OBJECTIVES: To evaluate the results of disc excision, with and without posterolateral fusion. SUMMARY OF BACKGROUND DATA: The effect of posterolateral fusion on the outcomes and radiologic changes in patients with lumbar disc herniation has rarely been reported. METHODS: Forty-four patients underwent disc excision, and 51 patients underwent disc excision and fusion. Clinical symptoms were evaluated using the Japanese Orthopaedic Association Back scores. All medical and surgical records were examined with regard to intraoperative blood loss, operation time, and other data. Preoperative and follow-up radiographs were analyzed to determine the spinal motion and disc height. RESULTS: Clinical outcome was excellent or good in 73% of the nonfusion group and in 82% of the fusion group (P = 0.31). The reduction in lower back pain after surgery was greater in the fusion group. The rate of recurrent disc herniation at the surgical level in the nonfusion group increased, but intraoperative blood loss, operation time, length of hospital stay, and total cost of procedure were all significantly less in the patients undergoing disc excision alone than in the fusion group. The radiologic analysis provided evidence that the disc height at the level of disc excision and posterolateral fusion in the fusion group decreased with time, as in the nonfusion group. The changes in disc height and spinal motion were not related to the clinical results. CONCLUSIONS: Although there is still controversy regarding the pros and cons of fusion in association with disc excision, there is seldom an indication for primary fusion for lumbar disc herniation. PMID- 10707391 TI - Quantifying changes in standing body segment alignment following spinal instrumentation and fusion in idiopathic scoliosis using an optoelectronic measurement system. AB - STUDY DESIGN: Longitudinal case studies before and after posterior spinal instrumentation and fusion (PSIF) in idiopathic scoliosis (IS). OBJECTIVES: To quantitate the changes in body segment alignment following PSIF using standard radiographic techniques and an optoelectronic measurement system. SUMMARY OF BACKGROUND DATA: Evaluation of surgical correction following PSIF is traditionally performed radiographically. Radiographic film cannot reliably document transverse and coronal plane pelvic, torso, and shoulder orientation resulting from the global effect of vertebral malalignment. METHODS: Thirty-three subjects with IS were evaluated radiographically and with an optoelectronic measurement system before and 13 months after PSIF. All subjects had a primary right thoracic curve pattern. Thirteen subjects without scoliosis were evaluated as controls. Symmetry measures and transverse and frontal plane orientation relationships were measured and compared preop, postop, and with controls. RESULTS: The major curve decreased from 60 degrees to 24 degrees after surgery. The magnitude of C7 decompensation from the sacrum decreased following PSIF. Control subjects demonstrated neutral coronal and transverse plane alignment. Preoperatively, patients had an elevated and protracted right shoulder. Postoperative data showed correction in both the coronal and transverse planes. Ten out of 33 subjects were within 5 degrees of neutral shoulder protraction before surgery. Twenty-six out of 33 were within 5 degrees of neutral following surgery. CONCLUSIONS: Optoelectronic measurement of body segment alignment may be a useful noninvasive technique in the evaluation of scoliotic deformity. This is a new method of evaluating the global effect of vertebral malalignment on body segment alignment and can be used to quantify changes following PSIF. PMID- 10707392 TI - Reoperation after primary posterior instrumentation and fusion for idiopathic scoliosis. Toward defining late operative site pain of unknown cause. AB - STUDY DESIGN: Retrospective case series. OBJECTIVES: To determine the frequency and categorize the indications for reoperation in three implant systems (Harrington, Cotrel-Dubousset [C-D], and Isola). To define late operative site pain (LOSP) of no apparent cause as an indication for implant removal and determine the success of implant removal in relieving LOSP. SUMMARY OF BACKGROUND DATA: Late operative site pain of no apparent cause has been discussed briefly in the literature but has not been investigated as a major indication for implant removal. METHODS: One hundred eighty-two of 190 consecutive patients with idiopathic scoliosis (96%) who underwent primary surgery between January 1, 1981, and December 31, 1992, by one surgeon in one hospital, with use of Harrington, C D, or Isola instrumentation were studied an average of 9 years after surgery to determine the indications for and frequency of reoperation. RESULTS: The overall frequency of reoperation for all instrumentation types combined was 19%: Harrington, 19%; C-D, 24%; and Isola, 14%. By 6 years' follow-up the cumulative risk of reoperation by Kaplan-Meier analysis was Harrington, 14%; C-D, 21%; and Isola, 14%. (statistically nonsignificant difference). The most frequent indication for reoperation was LOSP of no apparent cause: 8% (14 patients) for all instrumentation types combined. The average interval between the initial operation and reoperation for LOSP was 46 months (range, 20-97 months). The frequency of each implant type was Harrington, 6%; C-D, 12%; and Isola, 6%. By 6 years' follow-up, the cumulative Kaplan-Meier risk for reoperation due to LOSP was Harrington, 5%; C-D, 13%; and Isola, 8% (statistically non-significant difference). Of the 14 patients who had instrumentation removal for LOSP, 10 (71%) had successful relief of pain after implant removal. CONCLUSIONS: Occurring regardless of implant type, LOSP of no apparent cause after posterior instrumentation of scoliosis is a distinct clinical entity and is relieved by implant removal in most patients. PMID- 10707393 TI - Vertebral decancellation for severe scoliosis. AB - STUDY DESIGN: The results of staged surgery including vertebral decancellation were reviewed retrospectively for 21 patients with severe scoliosis. OBJECTIVES: To evaluate the benefits and limitations of vertebral decancellation as new anterior surgical procedure. SUMMARY OF BACKGROUND DATA: The curvatures of severe scoliosis are often very rigid, and surgical correction using the anterior or posterior approach may not achieve the desired correction. Some studies reported neurologic complications might appear due to the aggressive approach or excessive correction force. METHODS: Twenty-one patients (average age, 17.0 years) with severe scoliosis, in whom Cobb angle was over 80 degrees (average angle, 107 degrees), underwent staged anterior and posterior spinal reconstruction. Vertebral decancellation was performed as anterior procedure, and until posterior instrumentation, halo traction was carried out. The transition of curvatures in coronal and sagittal planes was assessed in this series. RESULTS: The average correction rate of lateral curvature at the final follow-up was 46%. The average loss of correction was 2.5 degrees. Kyphosis, measured between T5 and T12, changed from 41 degrees to 36 degrees. Lordosis, measured between L1 and S1, changed from 56 degrees to 45 degrees. Transient neurologic deficit was seen in one case after vertebral decancellation. CONCLUSIONS: Staged surgery including vertebral decancellation is an effective surgical method for patients with severe scoliosis, where an inflexible rigid curve or the risk of occurrence of neurologic complications due to temporary correction may exist. PMID- 10707394 TI - Effect of the transligamentous extension of lumbar disc herniations on their regression and the clinical outcome of sciatica. AB - STUDY DESIGN: Magnetic resonance imaging of symptomatic herniated lumbar discs was investigated longitudinally and prospectively for the presence of tear in the posterior longitudinal ligament (PLL). OBJECTIVES: To clarify the effect of transligamentous extension through the PLL of herniated disc on its regression and to determine the factors contributing to a successful clinical outcome. SUMMARY OF BACKGROUND DATA: Greater regression of the herniated fragment has been noted with larger initial disc herniations. The exposure of herniated disc materials to the epidural vascular supply through the ruptured PLL has been suspected to play a part in the mechanism of disappearance of the herniated nucleus pulposus. However, it had not been shown clinically. METHODS: Clinical outcomes and magnetic resonance images of 36 patients with symptomatic lumbar disc herniations, treated conservatively, were analyzed. Patients were divided into three groups: subligamentous, transligamentous, and sequestered herniations. The size of the herniated disc was measured by herniation ratio, which is defined as the ratio of the area of herniated disc to that of the thecal sac on the axial view. Factors associated with the natural regression of herniated disc and the successful clinical outcome were explored. RESULTS: Of the 36 herniated discs, 25 decreased in size. Ten (56%) of 18 subligamentous herniations, 11 (79%) of 14 transligamentous herniations, and all 4 (100%) sequestered herniations were reduced in size. The average decreases in herniation ratio of the subligamentous, transligamentous, and sequestered disc groups were 17%, 48%, and 82% respectively. The decrease in herniation ratio was related to the presence of transligamentous extension but was not related to the initial size of herniation. Successful outcome correlated with a decrease in herniation of more than 20%. CONCLUSION: Transligamentous extension of herniated disc materials through the ruptured PLL is more important to its reduction in size than is the initial size of the herniated disc. Decrease in herniation ratio of more than 20% seems to correspond to successful clinical outcome. PMID- 10707395 TI - Incidence of intravascular uptake in lumbar spinal injection procedures. AB - STUDY DESIGN: Multicenter, prospective, observational study. OBJECTIVES: To document the incidence of and factors associated with intravascular uptake during lumbar spinal injection procedures. SUMMARY OF BACKGROUND DATA: In prior reports, the incidence of inadvertent intravascular needle placement during contrast enhanced, fluoroscopically guided lumbar spinal injection procedures has been incidentally noted to range from 6.4% to 9.2%. We present the first systematic prospective documentation of intravascular uptake of contrast dye during different types of lumbar injection procedures. METHODS: Fifteen interventional spine physicians in seven centers recorded data regarding intravascular uptake during 1219 contrast-enhanced, fluoroscopically guided lumbar spinal injection procedures. RESULTS: The overall incidence of intravascular uptake during lumbar spinal injection procedures as determined by contrast enhanced fluoroscopic observation is 8.5%. Caudal and transforaminal routes have the highest rates at 10.9% and 10.8%, respectively, followed by zygapophyseal joint (6.1%), sacroiliac joint (5.3%), and translaminar (1.9%) injections. Intravascular uptake is twice as likely to occur in those patients over rather than under 50 years of age. Preinjection aspiration failed to produce a flashback of blood in 74% of cases that proved to be intravascular upon injection of contrast dye. CONCLUSION: The incidence of intravascular uptake during lumbar spinal injection procedures is approximately 8.5%. The route of injection and the age of the patient greatly affect this rate. Absence of flashback of blood upon preinjection aspiration does not predict extravascular needle placement. Contrast-enhanced, fluoroscopic guidance is recommended when doing lumbar spinal injection procedures to prevent inadvertent intravascular uptake of injectate. PMID- 10707396 TI - Low back pain in relation to lumbar disc degeneration. AB - STUDY DESIGN: Cross-sectional magnetic resonance imaging (MRI) study. OBJECTIVES: To study the relation of low back pain (LBP) to disc degeneration in the lumbar spine. BACKGROUND DATA: Controversy still prevails about the relationship between disc degeneration and LBP. Classification of disc degeneration and symptoms varies, hampering comparison of study results. METHODS: Subjects comprised 164 men aged 40-45 years-53 machine drivers, 51 construction carpenters, and 60 office workers. The data of different types of LBP, individual characteristics, and lifestyle factors were obtained from a questionnaire and a structured interview. Degeneration of discs L2/L3-L5/S1 (dark nucleus pulposus and posterior and anterior bulge) was assessed with MRI. RESULTS: An increased risk of LBP (including all types) was found in relation to all signs of disc degeneration. An increased risk of sciatic pain was found in relation to posterior bulges, but local LBP was not related to disc degeneration. The risks of LBP and sciatic pain were strongly affected by occupation. CONCLUSIONS: Low back pain is associated with signs of disc degeneration and sciatic pain with posterior disc bulges. Low back pain is strongly associated with occupation. PMID- 10707397 TI - To what extent do current and past physical and psychosocial occupational factors explain care-seeking for low back pain in a working population? Results from the Musculoskeletal Intervention Center-Norrtalje Study. AB - STUDY DESIGN: A population-based case referent study. OBJECTIVES: To determine whether current and past physical and psychosocial occupational factors are associated with care-seeking for low back pain in working men and women. SUMMARY OF BACKGROUND DATA: The importance of physical and psychosocial workloads as causal factors of low back pain has mostly been investigated in special occupational groups and with a cross-sectional design, which makes generalizability and interpretations more difficult. METHODS: The study comprised 2118 working men and women 20 to 59 years old (695 cases, and 1423 referents). Cases were defined as persons seeking care by any caregiver for low back pain. The exposure assessments were made through questionnaires and interviews about current and past physical and psychosocial loads during work and leisure time. RESULTS: In a logistic regression analysis, physical load from forward bending in men (RR = 1.8) and high physical load, in general, in women (RR = 2.0) showed increased relative risks. Psychosocial factors alone seemed to be of less importance in women, but "poor job satisfaction" and "mostly routine work without possibilities of learning" increased the risk in men. Combined current and past exposures further increased the risks. A combination of high physical and psychosocial loads increased the risk substantially, but few were exposed to such loads. Adjustment for lifestyle and other loads outside work did not change the results. CONCLUSION: Current and past physical and psychosocial occupational factors, both separately and combined, seem to be gender-specific, and to have a moderate impact on care-seeking for low back pain in a general working population. PMID- 10707398 TI - Clinical experience with functional electrical stimulation-assisted gait with Parastep in spinal cord-injured patients. AB - STUDY DESIGN: Clinical evaluation of the Parastep method, a six-channel transcutaneous functional electrical stimulation device, in spinal cord-injured patients. OBJECTIVES: To investigate the motor performances of this new technique regarding energy expenditure and to evaluate its advantages and limitations, especially in social activities involving ambulation. METHODS: This study was conducted in 15 thoracic spine-injured patients. The lesion was complete except in two patients. The gait ability and the functional use were judged clinically. Energy cost was evaluated from heart rate, peak oxygen uptake, and lactatemia. RESULTS: Thirteen patients completed the training (mean: 20 sessions) and achieved independent ambulation with a walker. The mean walking distance, without rest, was 52.8 +/- 69 m, and the mean speed was 0.15 +/- 0.14 m/sec. One patient with incomplete lesion, who had been nonambulatory for 8 months after the injury, became able to walk without functional electrical stimulation after five sessions. The follow-up was 40 +/- 11 months. Five patients pursued using functional electrical stimulation-assisted gait as a means of physical exercise but not for ambulation in social activities. The patients experienced marked psychological benefits, with positive changes in their way of life. In three subjects, a comparison of physiologic responses to exercise between a progressive arm ergometer test and a walking test with the Parastep (Sigmedics, Inc., Northfield, IL) at a speed of 0.1 m/sec was performed, showing that the heart rate, the peak oxygen uptake, and lactatemia during gait were close to those obtained at the end of the maximal test on the ergometer. CONCLUSIONS: In spite of its ease of operation and good cosmetic acceptance, the Parastep approach has very limited applications for mobility in daily life, because of its modest performance associated with high metabolic cost and cardiovascular strain. However, it can be proposed as a resource to keep physical and psychological fitness in patients with spinal cord injury. PMID- 10707399 TI - Transperitoneal laparoscopic exposure for lumbar interbody fusion. AB - STUDY DESIGN: A prospective clinical trial of the transperitoneal laparoscopic approach to the lumbar spine in a consecutive series of patients undergoing anterior lumbar interbody fusion. OBJECTIVES: To determine safety and effectiveness, and to document technique and perioperative complications of a laparoscopic exposure for lumbar interbody fusion. SUMMARY OF BACKGROUND DATA: With the widespread adoption of laparoscopic techniques, the benefits of minimal access surgery are now well recognized--in general, gynecologic and urologic surgery. Only recently have minimal access techniques been applied to spinal procedures. METHODS: Forty-seven patients with symptomatic degenerative disc disease underwent transperitoneal laparoscopic exposure of the lumbar spine to facilitate implantation of cylindrical threaded interbody fusion cages. These patients were prospectively followed and all perioperative considerations and complications were documented and analyzed. The surgical technique of laparoscopic exposure will be described. RESULTS: The laparoscopic approach was attempted in 47 consecutive patients. Forty-four were completed laparoscopically- 36 single level fusions, seven two level fusions, and one three level fusion. Early in the series, conversion to open surgery was required in one patient (case #3) because of bleeding from the presacral veins which hindered the view. In one case, mobilization of the great vessels proved to be difficult, and in one other case the patient could not tolerate abdominal insufflation. The mean blood loss for the entire group was 105 mls. Complications related to the endoscopic exposure were few. There were no injuries to major vascular structures or to bowel, and no mortalities. In two patients, the cages were malpositioned necessitating repeat endoscopic exposure for cage realignment. One patient required a laparotomy for a postoperative small bowel obstruction. The median postoperative stay was 4 days. CONCLUSIONS: Transperitoneal laparoscopic exposure for single or multiple level, anterior lumbar interbody fusion can be performed with low risk. Experience in open anterior spinal surgery and laparoscopic general surgery is vital in minimizing the risks. PMID- 10707400 TI - Use of epidural steroids after discectomy may predispose to infection. AB - STUDY DESIGN: This is a report of three cases of epidural abscess occurring after use of intraoperative epidural methylprednisolone in 31 patients who had undergone lumbar microdiscectomy. The possible role of epidural steroids in the cause of these abscesses is discussed, and a review the literature concerning its value is provided. OBJECTIVE: To evaluate experiences with the efficacy and safety of perioperative methylprednisolone. SUMMARY OF BACKGROUND DATA: No previous study has described a high infection rate with the use of epidural methylprednisolone. The literature supporting epidural steroids is equivocal, and reports supporting their perioperative use are scant. METHODS: In an 8-month period, 31 patients received 1 mL (40 mg) epidural methylprednisolone at the conclusion of microdiscectomy. Therapy was discontinued after an increased postoperative deep infection rate was noted. Results in these patients were compared with those in more than 400 others who did not receive intraoperative steroids during a 7-year period. RESULTS: In the steroid group, three epidural abscesses were encountered. There were no deep infections in the nonsteroid group. CONCLUSION: The use of perioperative epidural methylprednisolone in the currently reported cases was associated with three incidences of infection. A prospective study is needed to examine its use. PMID- 10707401 TI - Primary solitary Echinococcosis in cervical spine. Postsurgical successful outcome after long-term albendazole treatment. AB - STUDY DESIGN: A case report of a young man with isolated cervical hydatidosis treated postoperatively with sustained cyclical albendazole therapy for 9 years of follow-up. OBJECTIVES: To communicate the efficacy and safety of prolonged albendazole treatment in the postoperative management of spinal hydatid disease, and recommend therapeutic regimes for preventing its recurrence. SUMMARY AND BACKGROUND DATA: Bone involvement in hydatid disease is uncommon and the cervical region of the spine is rarely affected. Surgical excision remains the treatment of choice but high rates of postoperative recurrence have highlighted the importance of adjuvant anthelmintic therapy. The selection of the drug(s) and the duration of the medical treatment is still controversial. METHODS: The patient described herein presented with isolated bone lesions, in an unusual cervical location, and without coincidental visceral involvement. Therefore, diagnosis was delayed and surgical debridement was carried out without any preoperative anthelmintic therapy. To prevent late recurrences, therapy with intermittent courses of albendazole has been maintained for nine years and is still ongoing. Response and toxicity related to therapy has been closely monitored by clinical, biochemical and radiological follow up. RESULTS: After surgery the patient has remained asymptomatic without sequelae or evidence of relapses. No clinically relevant side effects has been observed. CONCLUSION: Prolonged albendazole treatment appears to be safe and effective in the prevention of late recurrences after spine hydatidosis surgery. Long-term chemotherapeutic schedules should be considered after surgical excision of spine or bone lesions. PMID- 10707402 TI - Atlantoaxial rotatory subluxation in patients with Marfan syndrome. A report of three cases. AB - STUDY DESIGN: This is a retrospective case series of three patients, ages 9 1/2, 13, and 20 years old, with Marfan syndrome treated for atlantoaxial rotatory subluxation. In the first two cases, acute torticollis was noted postoperatively, following pectus excavatum repair. The diagnosis was made in the third patient after she presented to the emergency room with a week-long history of unresolved neck pain following minor trauma. OBJECTIVE: To report and discuss the courses and clinical sequelae of atlantoaxial subluxation in patients with Marfan syndrome. SUMMARY OF BACKGROUND DATA: Radiographic analysis of patients with Marfan syndrome has shown that increased atlantoaxial translation, larger odontoid height, and basilar impression are more prevalent in this population compared to age-matched controls. Despite these findings, there are sparse data on injuries secondary to cervical spine instability or abnormalities in this population. To the authors' best knowledge, no report of atlantoaxial rotatory subluxation in patients with Marfan syndrome exists in the literature. METHODS: Case records of rotatory instability of the atlanto-axial level were reviewed and are presented in the following report. RESULTS: The first two patients described in this report were noted to have "cock robin" posturing of their necks following pectus excavatum repairs. The first patient's subluxation was partially reduced with halo traction, and he subsequently underwent posterior spinal fusion of C1 C2 with internal fixation. The patient was well aligned postoperatively, and had no neurologic deficits. The second patient's subluxation reduced after 20 days of halter and traction; he was immobilized in a collar following discharge and reduction was maintained. The third patient's subluxation failed to reduce with halo traction; further imaging studies revealed odontoid prominence in the foramen magnum. She underwent posterior spinal fusion, occiput to C3, with satisfactory result. CONCLUSIONS: The cervical bony and ligamentous abnormalities seen in patients with Marfan syndrome may slightly increase their risk for atlantoaxial rotatory instability. Special attention to intubation and positioning, both intraoperatively and postoperatively, may be necessary in patients with Marfan syndrome. Additionally, rotatory subluxation should be included in the differential diagnosis for Marfan patients with neck pain after injury. PMID- 10707403 TI - Aneurysmal bone cyst of the first cervical vertebrae in a child treated with percutaneous intralesional injection of calcitonin and methylprednisolone. A case report. AB - STUDY DESIGN: First published report of a child with an aneurysmal bone cyst (ABC) of the first cervical vertebrae treated successfully with intralesional injection of calcitonin and methylprednisolone. OBJECTIVES: To describe a safe and effective nonsurgical treatment method for an ABC of the first cervical vertebrae. SUMMARY OF BACKGROUND DATA: Aneurysmal bone cysts of the spine comprise from 3% to 20% of all such lesions. Upper cervical spine involvement is rare and these lesions are difficult to treat. Standard treatment with curettage and bone grafting or other alternatives such as radiation therapy or embolization may not be possible in this location. Percutaneous injection with a variety of agents has also been described. Methylprednisolone and calcitonin were selected in this case in an effort to combine the proposed angiostatic and fibroblastic inhibitory effects of steroid with the proposed osteoclastic inhibitory and promotion of new bony trabeculae formation effects of calcitonin. METHOD: This case was described, and pertinent literature reviewed. RESULTS: Sclerosis and shrinkage of the lesion with concomitant symptom resolution occurred after two injections with calcitonin and methylprednisolone via computed tomography (CT) guidance. No complications occurred. The lesion remained quiescent at a 2-year 7 month follow-up. CONCLUSIONS: Percutaneous intralesional injection of an ABC of the first cervical vertebrae with calcitonin and methylprednisolone in a child via CT guidance was a safe and effective treatment. This is a promising treatment for surgically inaccessible aneurysmal bone cysts. PMID- 10707404 TI - The role of activity in the therapeutic management of back pain. Report of the International Paris Task Force on Back Pain. PMID- 10707405 TI - [The coupled variability of quantitative and qualitative traits: a single-locus 3 allele model]. AB - Statistics for estimation of additive and non-additive effects of marker gene on quantitative trait are developed from the mad-model of a quantitative trait for three-allelic codominant marker locus. All they may be obtained directly from population data, without any hybridological experiments. PMID- 10707406 TI - [The luminescence of blood leukocytes, stained with the fluorochrome acridine orange, from surgical patients in the dynamics of emotional, anesthesiological and surgical stress]. AB - Luminescence of blood leukocytes of surgical patients during treatment was studied using orange acridine. Luminescence intensity at 640 nm wave length (I640) was the most informative. It characterized amount of single-stranded nucleic acids, synthesized in cell due to previous activation of protein synthesizing system and hence cell functional activity, while in lymphocytes these changes exceeded by duration and amplitude those in neutrophils. During premedication (emotional stress) I640 was minimal, during anesthetization it developed, and sharply raised during traumatic period of surgery and after it, as reflection of immune system participation in reparative regeneration. PMID- 10707407 TI - [The antimutagenic activity of biomass extracts from the cultured cells of medicinal plants in the Ames test]. AB - Antimutagenic activity of 20 and 40% ethanol extracts from the biomass of Rhodiola rosea, Polyscias filicifolia, Panax ginseng and Ungernia victoris cultured cells have been studied. DDDTDP, ethidium bromide, benz(a)pyrene, benzidine served as model mutagens for Salmonella typhimurium TA 98 strain (the latter two were tested in presence of metabolic activation system); for S. typhimurium TA 100 strain these were tio-tefa, bichromate potassium and sodium azide and heavy metal compounds (chlorides of manganese, zinc, cadmium, lead acetate) for both strains. Higher capacity of the extracts from the biomass of R. rosea and P. filicifolia to counteract gene mutations induced by various mutagens was demonstrated (ca. 90% inhibition in isolated cases). In the experiment with the metabolic activation most effective proved to be the extracts from the P. ginseng biomass (up to 34% and 47% mutagenicity inhibition). PMID- 10707408 TI - [The polymorphism of proteins, RAPD-PCR and ISSR-PCR markers in European and American bison and cattle]. AB - The comparative analysis of genetic differentiations between three species of Bovinae--Bos taurus, Bison bonasus, Bison bison with the use of different types of molecular-genetic markers--genetical-biochemical (35 loci) and DNA markers (RAPD-PCR, ISSR-PCR) was carried out. It was shown, that the evaluation of interspecies genetic interrelations was connected more with the determined molecular-genetic markers (loci), included in analysis, than with the marker's belonging to certain type (protein polymorphism, variability of DNA repeat distributions). PMID- 10707409 TI - [The cytogenetic effect in a group of settlers from a 30-kilometer area of right of way]. AB - One of the critical human group of potential risk is the group of population from 30-km zone around the Chernobyl nuclear power plant. The cytogenetical examination of 12 persons (8 man, 4 woman, 30-60 years old) from 5 villages situated in north-west and south-east directions from the ChNPP in the frame of this zone was conducted. The data of the conventional analysis showed relatively low cytogenetic effects comparable with the effects in exposed people out of the zone (the mean level of unstable cytogenetic markers in different villages was 0.21-0.35 per 100 cells). The level of stable translocations in one case (woman, 59 years old) determined using FISH (16.62 per 100 cells for the whole genome) exceeded their spontaneous aged frequency and confirmed the high sensitivity of this method for the evaluation of real radiation exposure. PMID- 10707411 TI - Interpretation of basic gross pathologic changes of the digestive tract. AB - The necropsy is a valuable diagnostic tool. When presented with a dead animal, it is not uncommon for the necropsy to be the springboard for the entire diagnostic evaluation. Not only is important information gained from gross examination of the organs but during necropsy, tissue and fluid samples for supportive tests- bacterial culture, antibiotic sensitivity, virus isolation, serology, parasite burden, and toxicologic and histopathologic studies--are collected. It is not essential for a veterinarian to be a pathologist to get good information from a necropsy. This article attempted to identify a number of basic lesions that occur with the most common diseases of the digestive tract of food animals. Additionally, associations between lesions and certain etiologies as well as diseases have been made so that when one identifies a particular lesion (in a live or dead animal), a prioritized list of possible differential diagnoses comes to mind. The necropsy does not stand alone or above the other sources of diagnostic information. The information gained from a necropsy must be correlated with the other information to arrive at either a specific diagnosis or a short list of possible diagnoses. The veterinarian must seek further input, in the latter situation, from additional clinical examinations, laboratory tests, or interviews with the client to arrive at the diagnosis. PMID- 10707410 TI - [Tumor-associated changes in the buccal epithelium in human breast and thyroid pathology]. AB - The estimation of the malignancy-associated changes in oral mucosa of patients with benign and malignant processes in mammary and thyroid gland is carried out on the basis of textural and densitometric indices of the interphase nuclei of the epitheliocytes with the help of the methods of mathematical statistics. PMID- 10707412 TI - Sampling techniques for the diagnosis of digestive disease. AB - This article covers sample selection, testing methods performed for specific agents, and pertinent clinical history information required for the diagnosis of food animal digestive disease. Sampling techniques for diagnostic testing and packaging and shipping information are also presented. PMID- 10707413 TI - Differentiation of gastrointestinal diseases of calves. AB - With a complete history, careful physical examination, and targeted diagnostics, the practitioner can differentiate the causes of gastrointestinal diseases in calves. The authors hope that this article helps the practitioner in this regard. Armed with a diagnosis, he or she can then proceed with proper treatment and prevention, which is the ultimate goal. PMID- 10707414 TI - Differentiation of gastrointestinal diseases in adult cattle. AB - Many of the common gastrointestinal disorders of adult cattle may be diagnosed by a careful physical examination, whereas other disturbances require the use of diagnostic testing. It is important to differentiate the causes of gastrointestinal disturbances to make better treatment decisions and have a clearer prognosis for the specific animal or herd of cattle. PMID- 10707415 TI - Diagnosis of enteric disease in small ruminants. AB - Diagnosis of gastrointestinal disease in small ruminants requires integration of information obtained in the signalment, history, physical or necropsy examination, and ancillary diagnostic tests. The purpose of this article is to provide the practitioner with a review of the clinical features of several common gastrointestinal diseases of sheep and goats. Rumen acidosis, enterotoxemia, gastrointestinal parasitism, neonatal diarrhea, and salmonellosis are discussed, and where appropriate, reviews of the pathophysiology, prevention, and control of these diseases are cited for further reading. PMID- 10707416 TI - Diagnosis of neonatal pig diarrhea. AB - To be effective, swine practitioners should develop a unit health program. Development should involve unit managers, owners, and employees involved in day to-day operations. Emphasis on training personnel and management to reduce disease and collection of accurate records is necessary. Routine diagnostics are needed to solve disease problems. Communication with laboratory personnel to ascertain what samples are needed for diagnosis of particular problems cannot be overemphasized. General diagnosis of disease problems outlined by Vinson can be similarly followed within the specifics of diarrheal problems within units. 1. Observe symptoms exhibited by pigs, i.e., huddling, fecal material around perineum, extreme thirst, etc. 2. Evaluate the degree of morbidity and potential production losses. 3. Analyze possible specific causes of symptoms, i.e., environmental cleanliness, affected litter distribution, age of affected neonates, and other populations affected. 4. Examine live animals, i.e., obtain serum samples from a random population, take rectal temperatures of affected neonates, and evaluate fecal pH. 5. Necropsy dead or dying pigs [that] appear to represent the problem. 6. Submit live pigs or appropriate tissues from necropsied pigs to a diagnostic laboratory. 7. Re-evaluate environmental conditions that may be contributing to the problem (remember, unit employees are a part of the pigs' environment). 8. Evaluate management procedures contributing to the disease problem, i.e., lack of adherence to all-in all-out, rapid turn-around decreasing cleaning time etc. Following this format and communicating with diagnosticians should provide for positive results for the producers both entities serve. PMID- 10707417 TI - Diarrhea in growing-finishing swine. AB - Regardless of the etiology of an enteric disease in nursery age to finisher swine, making a prompt and accurate diagnosis is crucial. Eliciting a complete history, assessing clinical signs and pathology, and selecting and interpreting laboratory tests are essential components in achieving this. Early detection and diagnosis of enteric disease is particularly critical in the nursery through finisher phase because of economic impacts. Recurrent topics when discussing control and prevention of enteric diseases are reducing stress and improving pig comfort and reducing or eliminating exposure through sanitation and biosecurity. These are not new concepts; in fact, prior to the advent of antimicrobials, they were the mainstay of treatment of enteric diseases. With concern over the use of antimicrobials in food animal production increasing, exploiting disease ecology to control enteric diseases is increasing in importance. New vaccines and bacterins for postweaning swine enteric diseases are needed tools to exploit the pig's immune system. Recent advances in diagnostic capabilities allow an increase in understanding and exploitation of disease ecology. PMID- 10707418 TI - Porcine gastric ulcer. AB - Ulceration of the pars esophagea in swine develops from a complex interaction of dietary particle size, gastric fluidity, dietary carbohydrate content, and presence of certain species of commensal gastric organisms capable of fermenting dietary carbohydrates. Unlike in humans, the significance of the role of Helicobacter sp. in development of porcine gastric ulcers is yet undefined. Management practices that limit the incidence and severity of gastric ulceration without interfering with growth performance appear to be the best option for control. PMID- 10707419 TI - Edema disease. AB - Edema disease is a common cause of illness and death loss in pigs during the first 2 weeks after weaning. The disease is an enterotoxemia caused by strains of E. coli that colonize the small intestine and produce Stx2e. Bacterial colonization is mediated by F18ab fimbriae. Susceptibility to disease is determined by presence of receptors for these fimbriae on small intestinal epithelial cells and is inherited as a dominant trait. Clinical signs and lesions are largely the result of Stx2e, which causes necrosis of endothelial and smooth muscle cells in small arteries and arterioles. Vascular damage in the brain stem with resultant infarction and malacia is the main cause of death in affected pigs. Studies conducted by veterinary researchers in the 1950s and 1960s identified the cause of the disease and provided future scientists with hypotheses to test regarding the pathogenesis. In the last two decades, studies using molecular-based techniques have allowed for the definitive identification of bacterial virulence factors that mediate intestinal colonization and vascular damage, that is, F18ab fimbriae and Stx2e. Identification of these virulence factors has provided a basis for current and future development of effective preventative measures, for example, vaccines. PMID- 10707420 TI - Toxicologic disease of the digestive tract. AB - There is a diverse and long list of toxicants that can affect the digestive system of food-producing animals. The plants and other natural toxicants discussed in this article are those primarily affecting the GI system. A number of other plants may also affect the digestive tract, but the effects from these are considered secondary and less pronounced. Often, plant poisonings affecting the digestive tract present with similar clinical signs, and a good thorough history is necessary to help differentiate between them. Moreover, a careful walk through the pasture with a keen eye to note plants that have been browsed or grazed may greatly assist the history. In cases where toxins are suspected as the cause of a GI disorder, consultation with a veterinary toxicologist at a diagnostic laboratory may be indicated. These professionals are knowledgeable about a wide variety of natural and other toxicants that may be present in your area. They can help with developing a differential diagnosis and the selection of appropriate samples to confirm the diagnosis. PMID- 10707421 TI - [Key responses of the visual cycle of vertebrates and invertebrates: activation and regeneration of rhodopsin]. PMID- 10707422 TI - [Species specific activity of digestive enzymes in insectivorous Chiroptera]. PMID- 10707423 TI - [Effects of alpha-tocopherol, ionol, triiodothyronine, and their combinations on the total plasma calcium level of mature white rats]. PMID- 10707424 TI - [Species specificity of the brain monoamine oxidase level in minks and rats]. PMID- 10707425 TI - [Responses of the lateral clivus neurons to the sound with the spectral cut-out in the mouse Mus musculus]. PMID- 10707426 TI - [Thermoregulation and wakefulness-sleep cycle in the ground squirrel subjected changes in the temperature regime of the thermosensitive region of the frontal hypothalamus]. PMID- 10707427 TI - [TRP protein in the Drosophila photoreceptors: immunolocalization of the hypothetical light-dependent calcium channel]. PMID- 10707428 TI - [The role of the left and right brain hemispheres in face recognition]. PMID- 10707429 TI - Drinking after a heart attack. PMID- 10707430 TI - Mechanisms of fatal opioid overdose. AB - There has been increasing recognition of the problem of fatal opioid overdose. This review examines the pharmacological basis of respiratory depression following opioid administration. Respiration is controlled principally through medullary respiratory centres with peripheral input from chemoreceptors and other sources. Opioids produce inhibition at the chemoreceptors via mu opioid receptors and in the medulla via mu and delta receptors. While there are a number of neurotransmitters mediating the control of respiration, glutamate and GABA are the major excitatory and inhibitory neurotransmitters, respectively. This explains the potential for interaction of opioids with benzodiazepines and alcohol: both benzodiazepines and alcohol facilitate the inhibitory effect of GABA at the GABAA receptor, while alcohol also decreases the excitatory effect of glutamate at NMDA receptors. Heroin and methadone are the major opioids implicated in fatal overdose. Heroin has three metabolites with opioid activity. Variation in the formation of these metabolites due to genetic factors and the use of other drugs could explain differential sensitivity to overdose. Metabolites of methadone contribute little to its action. However, variation in rate of metabolism due to genetic factors and other drugs used can modify methadone concentration and hence overdose risk. The degree of tolerance also determines risk. Tolerance to respiratory depression is less than complete, and may be slower than tolerance to euphoric and other effects. One consequence of this may be a relatively high risk of overdose among experienced opioid users. While agonist administration modifies receptor function, changes (usually in the opposite direction) also result from use of antagonists. The potential for supersensitivity to opioids following a period of administration of antagonists such as naltrexone warrants further investigation. While our understanding of the pharmacological basis of opioid-related respiratory depression has advanced, better understanding of the role of heroin metabolites, the metabolism of methadone, drug interactions and tolerance would all be of considerable value in knowing how best to respond to this problem. PMID- 10707431 TI - Heroin overdose is often the truer description. PMID- 10707432 TI - Fatal opioid toxicity--a clinical perspective. PMID- 10707433 TI - Heroin overdose: new directions for research. PMID- 10707434 TI - More on opioid overdose. PMID- 10707435 TI - The answer lies within selective ligands and pharmacogenomics. PMID- 10707436 TI - Future directions in opioid overdose. PMID- 10707437 TI - Lifetime tobacco, alcohol and other substance use in adolescent Minnesota twins: univariate and multivariate behavioral genetic analyses. AB - AIMS: We sought to estimate the contribution of genetic and environmental factors to adolescent tobacco, alcohol and other substance use. DESIGN, SETTING AND PARTICIPANTS: The sample consisted of 327 monozygotic and 174 like-sex dizygotic twin pairs born in Minnesota and aged 17-18 years at time of assessment. Biometrical methods were used to estimate the contribution of additive genetic, shared and non-shared environmental factors to adolescent substance use. MEASUREMENTS: As part of a day-long psychological assessment, adolescent twins completed a computerized substance use interview to determine whether they had ever used tobacco, alcohol or other illicit drugs. FINDINGS: The heritability for the liabilities to tobacco, alcohol and other drug use was estimated to be 59%, 60% and 33% among males, and 11%, 10% and 11% among females. However, the gender difference was not statistically significant. Estimates of shared environmental effect were substantial and insignificantly higher among females (71%, 68% and 36%, respectively) than among males (18%, 23% and 23%, respectively). The covariation among the three substance use phenotypes could be accounted for by a common underlying substance use factor. Estimates of the contributions of genetic, shared environmental and non-shared environmental factors to variance in this factor were 23% 63% and 14%, respectively. CONCLUSIONS: These findings add to the growing behavioral genetic literature indicating that adolescent initiation of substance use, a powerful predictor of adult substance use diagnosis, is influenced primarily by environmental rather than genetic factors. PMID- 10707438 TI - Predicting tobacco use to age 18: a synthesis of longitudinal research. AB - AIMS: To synthesize the available evidence on predictors of adolescent tobacco use. DESIGN: Meta-analysis was conducted on the empirical findings of published and unpublished studies of the natural development of tobacco use that used prospective multi-wave panel designs. PARTICIPANTS: The research literature that was analyzed included 106 reports on 64 studies representing data from a total of 145,750 study subjects; 1261 prospective and cross-sectional effect sizes were computed from these studies and used in the meta-analysis. MEASUREMENTS: Product moment correlations were analyzed examining the strength of the relationships between predictor variables and current and later tobacco use. In addition, findings reported as 2 x 2 contingency tables were analyzed to examine conditional relations and estimate the positive predicted values (PPV) and sensitivity indices for the predictive relationships. FINDINGS: The mean correlations for 17 different categories of predictors and current or later tobacco use ranged from -0.08 for race to 0.52 for prior tobacco use and were significant and positive except for race. Analysis of the conditional relationships showed that PPV for tobacco use ranged from a mean of 0.18 for predictors related to personal skills and knowledge (i.e. 82% of those 'at risk' on this construct did not use tobacco) to 0.70 for use of tobacco or other substances by parents. CONCLUSIONS: Current use of tobacco and other substances by youths, and use among their peers, showed stronger relationships with later tobacco use than other examined predictors. Combined with other predictive risk factors, these relationships are sufficiently strong to be useful in identifying for intervention those children most likely to become habitual tobacco users. PMID- 10707439 TI - The effects of fluoxetine combined with nicotine inhalers in smoking cessation--a randomized trial. AB - AIMS: Nicotine replacement therapy (NRT) is an established aid in stopping smoking, while the role of antidepressants remains uncertain. Antidepressants added to NRT might improve abstinence rates. Our aim was to determine the efficacy of nicotine inhaler and fluoxetine vs. nicotine inhaler and placebo in attempts to quit smoking. DESIGN: A randomized, double-blind, placebo-controlled trial. SETTING: A smoker's cessation clinic. PARTICIPANTS: One hundred volunteers smoking 10 cigarettes/day or more. INTERVENTIONS: Subjects were instructed to start taking a daily dose of 10 mg of fluoxetine or placebo 16 days before stopping smoking, then 20 mg 10 days before quitting, continuing for up to at least 3 months. Subjects were instructed to use 6-12 units per day of nicotine inhalers after stopping smoking for up to 6 months. MEASUREMENTS: Continuous abstinence rates recorded at various time points up to 12 months from the quit date. FINDINGS: The sustained abstinence rate for the inhaler-fluoxetine group was 54%, 40%, 29% and 21% after 1.5, 3, 6 and 12 months, respectively, compared to 48%, 40%, 32% and 23% for the inhaler-placebo group. The differences were not significant at any time point. Abstinence up to 3 months was more likely in older smokers, those with a lower Beck Depression Inventory Score (BDI), lower Fagerstrom Test of Nicotine Dependence (FTND) score and no history of alcoholism. Fluoxetine appeared to increase abstinence rates among high BDI smokers compared to high BDI smokers assigned placebo. Serum levels of nicotine during treatment in the inhaler-fluoxetine group were lower than in the inhaler-placebo group so that fluoxetine may have reduced inhaler use through a common site of action. CONCLUSIONS: We found no evidence that fluoxetine treatment when used as an adjunct to NRT in unselected smokers is effective, but there may be an advantage to using it in depressed smokers. PMID- 10707440 TI - Young, wet & wild? Associations between alcohol intoxication and violent behaviour in adolescence. AB - AIMS: To assess gender- and age-specific associations between alcohol intoxication and engagement in violent behaviours in young people. DESIGN, PARTICIPANTS AND MEASUREMENT: Cross-sectional study comprising a national sample of 12,000 Norwegian adolescents aged 12-20 years. Data on violent behaviour, alcohol intoxication and various confounders were obtained by self-administered questionnaires in school. FINDINGS: 2.8% had been in fight with a weapon and 32.6% had been beating or threatening to beat someone during the past year. Violent behaviours were more often reported among boys, in the younger age groups, with increasing frequency of alcohol intoxication, among users of other drugs, among those engaged in criminal activities and among those in wet environments (friends drinking regularly and parents often being intoxicated). The impact of intoxication frequency on number of times engaged in violent behaviours was of modest magnitude. It was greater in the youngest age group compared to those in the middle and late teens and greater for boys than for girls. However, when criminal activities were controlled for, the adjusted effect of intoxication on violent behaviour was significantly reduced, the effect was then of the same magnitude for both genders, whereas there was no longer any significant effect in the youngest age group. Controlling also for parents' and friends' drinking and parental monitoring did not alter these findings. CONCLUSIONS: A small direct effect of alcohol intoxication on violent behaviour appears to remain after controlling for various relevant confounders in middle and late teens. However, possible indirect effects of alcohol intoxication, mediated by own deviant life-style and wetness of environment, should also be taken into consideration. PMID- 10707441 TI - Activation of alcohol-related associative networks by recent alcohol consumption and alcohol-related cues. AB - AIMS: To investigate the influence of recent alcohol consumption and alcohol related cues on performance in a sentence generation task. DESIGN: Two experiments were carried out. In the first, the performances of light, moderate and heavy drinkers were compared. In the second, subjects were randomly assigned to one of three experimental treatments (alcohol-priming, non-alcohol priming, and control) and classified as light, moderate or heavy drinkers. The effect of experimental treatment, drinking status, gender and the interaction between these factors was studied. SETTING: The experiments were carried out in quiet research rooms in psychology departments. PARTICIPANTS: Volunteers recruited from university campuses. MEASUREMENTS: Questionnaires were used to ascertain recent drinking histories. Subjects generated sentences incorporating ambiguous alcohol related words which were provided by the experimenter. The sentences were then classified as alcohol-related or not, the dependent measure was the number of alcohol-related sentences produced. FINDINGS: In both experiments heavier drinkers produced more alcohol-related sentences and males produced more alcohol related sentences than females. In the second experiment more alcohol-related sentences were produced after subjects were exposed the alcohol priming condition. CONCLUSIONS: The alcohol-related meaning of ambiguous words is more likely to be accessed by males and by heavier drinkers and after exposure to other alcohol cues. PMID- 10707442 TI - Substance use among young people: the relationship between perceived functions and intentions. AB - AIMS: To explore the relationship between young people's use of psychoactive substances, perceived functions for using, the experience of negative effects, and the influences of these variables on their intention to use substances again. DESIGN: Cross-sectional survey in which respondents were purposively recruited using snowballing techniques. SETTING: Interviews were conducted in informal community settings. PARTICIPANTS: One hundred young drug and alcohol users (45 females) aged between 16 and 21 years. MEASUREMENTS: Life-time prevalence, current frequency and intensity of substance use and intentions to use again were assessed for four target substances (alcohol, cannabis, amphetamines and ecstasy) together with measures of the perceived functions for their use and peer substance involvement. FINDINGS: The life-time experience of negative effects from using the assessed substances was not found to correlate with current consumption patterns. Statistically significant associations were observed between the reported frequency of taking substances and the perceived social/contextual and/or mood altering functions cited for their consumption. The substance use function measures together with the reported extent of peer use were significant predictors of intentions to use again. CONCLUSIONS: If these findings are confirmed in larger studies, educational and preventative efforts may need to acknowledge the positive personal and social functions which different substances serve for young people. The results also call into question the extent to which the experience of negative effects influences future patterns of use. PMID- 10707443 TI - Continuing injecting risk behaviour: results from the Amsterdam Cohort Study of drug users. AB - AIMS: To give a detailed description of injection-related risk behaviours, and to estimate the relative importance of these behaviours with regard to HIV transmission. DESIGN: The present study was part of the Amsterdam Cohort Study of drug users. SETTING: In Amsterdam, a city with extensive preventive measures, large HIV-risk reductions have taken place, but no further decreases have occurred since 1991. PARTICIPANTS AND MEASUREMENTS: A detailed questionnaire on injecting risk behaviour was completed by a cross-section of participants in 1992/93 (n = 168). Among 48 HIV-seroconverters, a questionnaire was completed concerning possible HIV-transmission route. FINDINGS: Of 96 HIV-negative participants, 23% deliberately borrowed a used syringe, 18% reported possible "accidental" borrowing, 9% front/backloading, 4% simultaneous injection, and 32% possible sharing of ancillary equipment. Of deliberate borrowers, 64% borrowed from a person with unknown or positive HIV serostatus, and 81% did not appropriately clean the equipment; 79% borrowed in the absence of serious withdrawal symptoms. Risk factors differed for deliberate and 'accidental' borrowing. Among the HIV seroconverters, the most likely transmission route was borrowing in 29% of cases, front/backloading in 8%, borrowing or front/backloading in 21%, unprotected sexual contact in 23% (mainly with regular partner) and either injecting or sexual risk in 13%. Women were much more likely to report sexual transmission (p = 0.016). Borrowing was admitted by 43% before, and 64% after awareness of HIV-seroconverion. CONCLUSIONS: As the injecting risk is high, usually deliberate, and often in the absence of withdrawal symptoms, further prevention seems difficult. Although deliberate borrowing is the main risk for HIV seroconversion, unprotected sexual contacts and front- and backloading may be more important than previously thought in Amsterdam. Under reporting of borrowing is probably substantial, but does not alter the above conclusions. PMID- 10707444 TI - HIV and HCV infection among drug users in Japan. AB - AIMS: To assess seroprevalence of human immunodeficiency virus (HIV), hepatitis C virus, injecting drug use, unsafe sexual behaviours, self-mutilation and tattoos in patients attending a drug and alcohol treatment centre in Japan. DESIGN: Cross sectional survey. SETTING: The work was carried out at the National Sanitarium of Shimousa, Chiba, Japan, a 32-bed inpatient centre specializing in drug and alcohol treatment. MEASUREMENTS: Laboratory analyses for HIV antibody, hepatitis C antibody, hepatitis B antigen and antibody; questionnaires for history of sexual activities, needle and syringe use; physical examination with assessment of self-amputated finger joints, tattoos, scars from lacerations and cigarette burns. FINDINGS: No patients tested positive for anti-HIV. The seroprevalence of anti-HCV positives was 53.8% of methamphetamine-dependent patients, 18.4% of solvent-dependent patients and 5.6% of alcohol-dependent patients. Past needle sharing was reported by 82.1% of methamphetamine-dependent patients, 18.4% of solvent-dependent patients and 5.6% of alcohol-dependent patients. A history of syringe sharing was reported by 87.2% of methamphetamine-dependent patients. More than two-thirds of all patients reported contact with commercial sex workers. Casual sexual contacts were more common among solvent and methamphetamine dependent patients than those dependent on alcohol. Tattoos and cigarette burns were more common among methamphetamine and solvent-dependent patients than among alcohol-dependent patients. Among the methamphetamine-dependent patients, 20.5% reported self-amputated finger joints compared with none in the other patient groups. CONCLUSIONS: HCV prevalence is a significant problem among methamphetamine users in Japan, probably because of a high rate of needle and/or syringe sharing. Although HIV infection is currently negligible, the very high rate of needle and syringe sharing could give rise to a significant increase in the HIV rate among drug users in the future. PMID- 10707445 TI - Tuberculosis chemoprophylaxis using a liquid isoniazid-methadone admixture for drug users in methadone maintenance. AB - BACKGROUND: Tuberculosis is common in drug users, although compliance with therapy may be difficult in this population. OBJECTIVE: To evaluate an approach to enhancing compliance with tuberculosis chemoprophylaxis in drug users enrolled on methadone maintenance utilizing an isoniazid (INH)-methadone admixture. DESIGN: A prospective cohort study. SETTING: A drug treatment program in New Haven, Connecticut, USA. PATIENTS: Opioid-dependent drug users enrolled in methadone maintenance. INTERVENTION: Liquid isoniazid was mixed into subjects' daily dose of methadone. Vitamin B6 was given to subjects for self administration. MEASUREMENTS AND MAIN RESULTS: Number of eligible subjects, reasons for not starting therapy, number starting therapy, proportion completing therapy and median duration of INH therapy were calculated. Thirty-nine subjects were eligible for INH chemoprophylaxis: 34 (87%) received INH mixed directly in their methadone and five (13%) had their INH consumption supervised by a nurse. Among these subjects, 72% (28/39) completed therapy. Among the 11 subjects who discontinued INH, discharge from the methadone maintenance program was the most common reason--73% (8/11). Thus, among the 31 subjects who were not discharged from methadone maintenance, 90% (28/31) successfully completed INH prophylaxis. The median duration of therapy was 182 days. CONCLUSIONS: Tuberculosis chemoprophylaxis using a liquid isoniazid-methadone admixture appears to be an effective approach to enhancing compliance with this therapy in methadone maintained drug users. PMID- 10707446 TI - Drugs in the peri-operative period: 4--Cardiovascular drugs. AB - Many patients undergoing elective surgery will be taking medicines for cardiovascular disorders. Here, in the fourth and final article in our series on drug therapy in the peri-operative period, we review the management of patients taking certain antithrombotic, antihypertensive, anti-anginal or anti-arrhythmic drugs. PMID- 10707447 TI - Helping undernourished adults in the community. AB - People who are undernourished, whatever the cause, feel unwell, function badly and, compared with others, are more likely to consult their GPs, to need medication and be admitted to hospital. Once in hospital, they have a higher rate of complications, such as infections and pressure sores, and they take longer to recover and to return home. In an article 3 years ago, we discussed the detection and management of undernourished adults in hospital. Here, we consider undernourishment in those living in the community. As in the previous article, we concentrate on general undernourishment (protein and energy deficiency), rather than specific deficiencies of micronutrients (such as vitamins and minerals). PMID- 10707448 TI - Self regulation and revalidation--probity or quality? PMID- 10707449 TI - Lessons from liaison (or a submariner's jolly!). PMID- 10707450 TI - Role of general duty medical officers in humanitarian operations: child health, advocacy and protection? PMID- 10707451 TI - Heat strain in Royal Navy helicopter aircrew. AB - A review of the literature on heat strain and aircrew and a questionnaire survey of Royal Navy aircrew have been completed. Aircrew appreciate, some 50% from first hand experience, that heat strain can reduce their operational endurance and performance. They are at greatest risk of developing it in the pre-flight period, especially when wearing Nuclear, Biological, or Chemical (NBC) protective equipment. Several techniques they use to reduce this risk are described. Some may be of particular assistance in the field should air conditioned facilities be unavailable. However, opportunities to improve the thermal environment within the aircraft on the ground and in flight are limited as the heat generated within it and high levels of solar radiation impinging on it severely challenge air conditioning units, themselves constrained by weight and size. Other demands placed on protective clothing offer little potential to increase the rate at which aircrew can lose accumulated heat. It is concluded that an appropriate micro-climate cooling system worn next to the skin may be required to achieve truly significant reductions in heat strain. Research at the Institute of Naval Medicine has identified liquid cooling techniques which may be suitable for aircrew in all but the smallest helicopters. Any views expressed are those of the author and do not necessarily represent those of the Department. PMID- 10707452 TI - Handsearching the Journal of the Royal Naval Medical Service for Trials. AB - As part of the Cochrane Collaboration's international research endeavour, the authors carried out a handsearch of the Journal of the Royal Naval Medical Service from 1948 to 1998, searching for randomised controlled trials (RCTs) and controlled clinical trials (CCTs). Five trials were identified, of which three were RCTs and two were CCTs. The first trial was published in 1960. The identified trials were in the fields of dentistry (two trials), gastroenterology, occupational medicine and orthopaedic surgery. Of the five trials, only two had been located previously through a rigorous interrogation of Medline. The three newly identified trials were reported to the UK Cochrane Centre, and details of these three trials were entered into Medline for use by clinicians and investigators in the future. PMID- 10707453 TI - Royal Navy surgeons and the transmission of brucellosis by goats' milk. PMID- 10707454 TI - Royal British Legion war grave pilgrimages: a medical escort's perspective. AB - The Royal British Legion organizes pilgrimages to nearly all parts of the world where British servicemen and service-women and their allies fought and died. The Pilgrimage Department has taken thousands of widows, other relatives, veterans and friends to visit the grave of a loved one or comrade buried overseas. The parties of pilgrims are escorted by Service medical officers and nurses of the Regular and Reserve Armed Forces. The role of the medical escort is described. PMID- 10707455 TI - Submarine escape and rescue in today's Royal Navy. PMID- 10707456 TI - Defibrillation in hyperbaric chambers: a review. AB - Defibrillation plays a crucial role in the resuscitation of patients from acute life-threatening cardiac dysrhythmias causing cardiac arrest. Concerns over safety and function of defibrillators under pressure have so far prevented their routine use in clinical hyperbaric chambers. Increasing numbers of unstable and critically ill patients are being treated in such facilities for both diving and non-diving indications. This report reviews the literature relating to hyperbaric defibrillation and examines the indications, contraindications and therapeutic alternatives to this procedure. PMID- 10707457 TI - French emergency care systems. PMID- 10707458 TI - An evaluation of self-administration of prescribed medicine protocol for Royal Marines under training. PMID- 10707459 TI - Pelvic synovial cyst formation secondary to perforation of the quadrilateral plate: a case report. AB - A seventy seven-year-old female presented to her gynaecologist with right-sided pelvic pain and irregular per vaginal bleeding. Ten years earlier she had undergone a right cementless total hip arthroplasty. The press fit acetabular component had been augmented by screws. Radiological investigation requested by her gynaecologist identified a right iliac fossa mass. This communicated with the hip joint via a screw track formed by a fixation screw that had penetrated the medial wall of the acetabulum at total hip arthroplasty. Following aspiration of the fluid within the cyst, microscopy demonstrated it to be of synovial origin. Her symptoms subsequently resolved. This condition has not been reported previously. PMID- 10707460 TI - Middle-term results of a cementless threaded self-tapping acetabular cup. AB - Various designs of acetabular cup are available for cementless fixation in patients undergoing total hip arthroplasty. Conflicting results have been reported in the literature about the middle to long-term outcomes with the use of these cups. We present our experience of a design of self-tapping threaded acetabular cup with metal backing (the ACSYS acetabular cup). This is a study of 41 hips with average follow up of 6.43 years. Functional grading of the hips was very satisfactory and none of the patients needed revision for aseptic loosening. No case of significant cup migration or osteolysis was noted. The paper also reviews the literature. PMID- 10707461 TI - Subsmash in Liverpool Bay--a medical and naval disaster? PMID- 10707462 TI - [Antithrombotic therapy in atrial fibrillation]. PMID- 10707464 TI - [The use of peripheral blood progenitor cells as an autologous hematopoietic support in high-dose chemotherapy. I. The rationale and results]. AB - We review the rationale for PBPC transplantation and the results reported in the literature. In order to prolong complete remissions and increase cure rates, high dose chemotherapy is frequently used in the treatment of selected neoplasias. Hematological toxicity can be overcome by the infusion of autologous hemopoietic progenitors. Recently, peripheral blood is being used as the preferred source for hemopoietic progenitors, since it allows faster hematopoietic recoveries when compared to progenitors harvested from bone marrow. An adequate graft is defined by its content in clonogenic progenitors (mainly CFU-GM) and CD34 positive cells; these two parameters need to be accurately determined by specific laboratory methods. PBPC grafts are harvested using cell separators during leukaphereses; to increase efficiency, hemopoietic progenitors are first mobilized into the circulation with growth factors and or chemotherapy. PBSC transplantation may have procedure-associated toxicity related to the mobilization, harvest or reinfusion of the graft. PMID- 10707465 TI - [The use of peripheral blood progenitor cells as an autologous hematopoietic support in high-dose chemotherapy. II. The experience of the Hematological Intensive Care Unit of the IPOFG. Franciso Gentil Portuguese Institute of Oncology]. AB - We report the results of PBPC collection by large-volume leukaphereses and the hematologic recovery after high-dose chemotherapy supported by autologous PBPC reinfusion in a series of cancer patients treated at the Hematological Intensive Care Unit (UCHI) (Portuguese Institute of Oncology, Lisbon). Large volume leukaphereses were used to increase the efficacy of the PBPC collection. This modification of the standard apheresis technique allowed the harvesting, in only one session, of enough progenitors to proceed to transplantation in nearly 2/3 of patients and without significant toxicity. From December 1993 until September 1997, 95 autologous PBSC transplants were performed at the UCHI; 45% were performed in solid tumor patients and 55% in patients with hematologic malignancies. Hematologic recovery was similar to that published in the literature and related to the number of CD34+ cells infused. Patients supported with bone marrow in addition to PBPC showed delayed hematopoietic recovery, probably because the bone marrow harvest was only performed when an insufficient number of PBPC had been collected (2 x 10(6) CD34+ cells/Kg). The speed of hematological recovery differed per diagnosis, being higher in multiple myeloma and solid tumor patients and lower in Hodgkin's disease patients. PMID- 10707463 TI - [Donor leukocyte infusion after allogeneic stem cell transplantation]. AB - Adoptive cellular immunotherapy with donor leukocytes of patients submitted to allogenic stem cell transplantation has had significant success in the past few years, especially in the treatment of primary disease relapse and in the prevention and treatment of some post-transplant infectious complications. Most patients treated with donor leukocytes had a relapse of chronic myelogenous leukemia, which was successfully re-induced into remission. The most significant toxicities of this treatment are the development of graft versus host disease and marrow aplasia. Three strategies were developed to limit the former: the infusion of graded doses of donor leukocytes, the depletion of CD8+ cells and the transfer of donor leukocytes transvected with a timidine kinase gene, which renders these cells sensitive to gancyclovir. The post-transplant infectious complications treated successfully with donor leukocytes were Epstein-Barr virus-induced lymphoproliferative disorders and cytomegalovirus infection. The former, arising most frequently in recipients of unrelated and/or mismatched T-cell depleted grafts, were treated with donor unseparated leukocytes or Epstein-Barr virus specific T-cells. Cytomegalovirus infection in the early post-transplant period was largely prevented by the infusion of virus-specific T-cell clones, which restored donor-specific immunity to cytomegalovirus in the recipient. PMID- 10707466 TI - [Glucose-6-phosphate dehydrogenase deficiency in 2 girls]. AB - Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common erythrocyte enzymopathy, affecting over 400 million people worldwide. Portugal is a low prevalence country, but immigrants from endemic regions are common, particularly in the south of the country. In the present study, we report the laboratory findings observed in two black proband children with low G6PD enzyme activity (23 and 18%). The study also included their first-degree relatives. Both biochemical parameters (enzyme activity, electrophoretic mobility and cytochemical test) and genetic determinations (mutation and haplotype characterisation) were performed. PMID- 10707467 TI - [Acute lymphoblastic leukemia in childhood. A 10-year experience with the DFCI 81 01 protocol]. AB - Acute lymphoblastic leukemia is the most frequently encountered pediatric cancer. Approximately 70% of cases can be cured of the disease. In this article, we describe the experience of our Center in the last ten years with a treatment protocol adapted from protocol DFCI 81-01 (from the Dana Farber Cancer Institute, Boston). We conclude that it is easily accomplished, well tolerated and that it allowed us to significantly improve the outcome of our patients (80% 5 year survival). Nevertheless, we are currently in the process of changing to a new protocol--one that will hopefully achieve a comparable cure rate with less long term toxicity. PMID- 10707468 TI - [The molecular basis of dominantly inherited beta-thalassemia]. AB - In this study, we sought to clarity the molecular basis of a dominant inherited beta-thalassemia, found in heterozygosity in a northern Portuguese family with thalassemia intermedia. We characterized: i) the alpha-globin gene cluster structure; ii) the beta-globin gene cluster haplotype; and iii) the beta thalassemia mutation. The alpha-globin gene cluster was structurally normal. The G-->T transversion at codon 121 of the beta-globin gene was found in the affected individuals in association with Orkin's haplotype V. This is an uncommon, though ubiquitous, mutation. Which has also been found, in association with different haplotypes, in several distant populations. It has only been observed in this three-generation family, in the Portuguese population. We suggest a mechanism to explain the genotype/phenotype correlation. PMID- 10707469 TI - [Clinical trials. Some general ethical questions]. AB - This article provides an overview of the main problems that ethics committees deal with when analysing clinical trials. Some characteristics of the different phases are discussed as well as some particular problems of the Portuguese law. PMID- 10707470 TI - [Infectious diseases: 2 decades of challenge]. PMID- 10707471 TI - [Prediction, prevention and early intervention of allergic disease]. PMID- 10707472 TI - [Concept of sick house syndrome, and the strategy for the management and the prevention]. PMID- 10707473 TI - [Primary immunodeficiency disorders--new concept of their pathogenesis and clinical entity]. PMID- 10707474 TI - [A study of free radical defense and oxidative stress in the sera of patients with neuroimmunological disorders]. AB - Free radicals are molecules that contain at least one unpaired electron and by nature are highly reactive and potentially destructive. Free radical damage can play an important role of demyelination. Glutathione peroxidase, which plays a role in free radical defenses, and myeloperoxidase and lactoferrin, which are considered to reflect the strength of oxidative stress, were examined by monoclonal antibody-based enzyme immunoassay on serum samples taken from patients with neuroimmunological disorders, namely, 35 multiple sclerosis(MS), and 2 Balo disease, 10 Guillain-Barre syndrome(GBS), and 25 human T-lymphotropic virus type 1 associated myelopathy (HAM). The levels of glutathione peroxidase in active phase of MS (8.37 +/- 5.59 micrograms/ml: p < 0.05) were increased rather than in inactive phase (5.05 +/- 2.44 micrograms/ml) and control (5.41 +/- 1.40 micrograms/ml), the levels of myeloperoxidase in HAM (95.5 +/- 89.1 ng/ml: p < 0.05) were increased rather than in controls (21.5 +/- 4.1 ng/ml), and the levels of lactoferrin were not significantly increased than in other disease and control. Moreover the levels of myeloperoxidase and lactoferrin are increased in Balo disease (myeloperoxidase 487, 762 ng/ml; not significant, lactoferrin 2.58, 2.77 ng/ml; not significant) than in control (myeloperoxidase 21.5 +/- 4.1 ng/ml, lactoferrin 0.69 +/- 0.32 ng/ml). In conclusion, we have here first demonstrated that the levels of these enzyme were not paralleled in MS and Balo diseases. In GBS the levels of all these enzyme were not increased. Thus, these findings suggest that these enzyme may play an important role of the disease activity of Balo, and may reflect the activity of the defense of MS. PMID- 10707475 TI - [Hospitalization reduction by an asthma tele-medicine system]. AB - We examined an effectiveness of a new asthma telemedicine system in reducing hospitalizations using a multi-site randomized control study. In this program, a nurse under physician supervision monitors the patient's airway status at home and provides instructions to individuals via the telephone, helping them manage exacerbations as well as reinforcing proper use of a zone-controlled management plan. Patients with a high risk for hospitalization were screened based on the numbers of emergency room visits and hospitalizations found in a previous study and randomly assigned to either the telemedicine or control group. After a six month study period, an 83% reduction in hospitalization was demonstrated in the telemedicine group versus the control group, with a P value of 0.01. Improvement of peak expiratory flow and symptoms were also shown in the study group. We conclude that the key success factors in home asthma management for poorly controlled asthma patients are early detection of exacerbations through daily peak flow monitoring, compliance with prescribed daily prophylactic anti inflammatory steroid medications, and immediate action as specified by a zone controlled action plan upon the first signs of deterioration. PMID- 10707476 TI - [Comparison between repeated antigen and acetylcholine-induced bronchoconstriction in development of airway wall thickness in a chronic guinea pig model of asthma]. AB - Asthma is associated with a chronic remodeling of the airways, but it remains to be elucidated whether repeated bronchoconstriction (BC) influences any structural attributes of the lungs. To investigate the role of repeated BC on the morphology of the airways, we chronically exposed guinea pigs to an aerosol of acetylcholine (ACH) over a period of 6 months. The guinea pigs were challenged with either ACH or saline daily for 10 days and thereafter were challenged once a week. To compare repeated BC with a chronic model of allergic inflammation, an additional group of animals were sensitized to aerosolized ovalbumin (OA) and subsequently exposed with OA. The airway wall area and basement membrane (BM) thickness were measured by standard morphometric techniques. Both airway wall area and BM thickness were significantly increased in OA exposed guinea pigs compared with both saline control and AHC-exposed guinea pigs. Our results suggest that persistent bronchoconstriction and/or allergic airway inflammation in asthmatics may contribute to the thickness of the airway wall observed in this disorder. However, repeated acute bronchoconstriction events may not contribute to the increase in the airway wall or BM thickness. PMID- 10707477 TI - [The relationships of mononuclear leukocyte beta-adrenergic receptors to aerobic capacity and exercise-induced asthma in asthmatic children]. AB - beta-adrenaline receptors exist on peripheral mononuclear leukocytes as well as in lung tissue. We assessed the relationships of plasma catecholamine release by exercise to aerobic capacity and to exercise-induced asthma (EIA) in asthmatic children (Study 1). We then measured mononuclear leukocyte beta-adrenergic receptor densities at rest and assessed the relationships of the number of receptors to aerobic capacity, EIA, and bronchial responsiveness to acetylcholine (Study 2). Study 1: Eleven children (9 males, 2 females; 11-16 years old) with bronchial asthma participated in this study. The subjects underwent an incremental aerobic exercise test on a cycle ergometer to determine their aerobic capacity at the lactic threshold (LT) and VO2max. Each subject underwent an EIA test of which the intensity was 175% of LT, and plasma catecholamine concentrations were measured before and after exercise. A significant negative relationship was found between the degree of EIA and % change of plasma adrenaline concentrations to rest level (p < 0.05), and a significant positive relationship was found between VO2 max/wt and % change of plasma adrenaline concentrations (p < 0.05). Study 2: Twelve asthmatic children (10 males, 2 females; 11-16 years old) participated in this study. Aerobic capacity, and degree of EIA were also measured in each subject by the same method as that used in Study. 1. The number of mononuclear leukocyte beta-adrenergic receptors at rest was determined by (-) [125I]-iodocyanopindolol binding in each subject. A significant negative relationship was found between the number of adrenergic receptors and Max. % fall in FEV1.0 (p < 0.01), and a significant positive relationship was found between the number of adrenergic receptors and VO2max/kg (p < 0.001). These results suggested that a reduced adrenaline production and a reduced number of beta-receptors contributed to the pathogenesis of EIA. PMID- 10707478 TI - [The genome project in its final phase: It will soon be possible to explain many of the brain diseases]. PMID- 10707479 TI - [Chromosome 22 surveyed. The big challenge now is to understand the function of proteins]. PMID- 10707480 TI - [Mental disease a heritage. New genetic knowledge can reveal "public diseases" such as autism, dyslexia, alcoholism, anorexia, schizophrenia]. AB - Family and adoption studies indicate that genetic factors play a role in the development of many psychiatric disorders. A variable number of possible interacting genes predisposing to the diseases is likely. The genetic dissection has been hampered by genetic complexity as well as by difficulties in defining the phenotypes. Genetic mapping efforts using sib pairs, twins and individual large families has revealed preliminary or tentative evidence for susceptibility loci for a number of psychiatric disorders. Illnesses include the prion disease familial fatal insomnia (FFI), alcoholism, anorexia nervosa, autism, bipolar affective disorder, dyslexia, enuresis nocturnal, epilepsia, obsessive-compulsive disorders (OCD), schizophrenia, as well as the dementias, Alzheimer's disease and frontal lobe dementia, and mental retardation. The genes and proteins related to the newly discovered transmitter in the central nervous system, nitric oxide (NO), and its genes and proteins are also reviewed. The number of mapped human genes now exceeds 30,000 of the estimated total number of 60,000 to 100,000 genes. This rapid development will facilitate gene mapping, as well as efforts to isolate and identify the genes responsible for symptom susceptibility in many of the etiologically unclear psychiatric diseases with complex genetic origin. PMID- 10707481 TI - [Evidence-based treatment of hypertension in type 2-diabetes. New studies support active blood pressure reduction]. AB - Tre treatment of hypertension in type 2 diabetes in currently much in focus. The aim of this treatment is to prevent cardiovascular disease by reducing the mean blood pressure level to less than 140/85 mm Hg in all patients who do not experience orthostatic reactions. In most patients it seems as if the strict blood pressure control is more important for the prognosis than the choice made of specific drug therapy. Nevertheless, specific pharmacological properties may be of special importance for some patients. During the last few years several new randomised trials have increased our knowledge for an approach based on evidence based medicine towards hypertension in type 2 diabetes. These studies are summarized and discussed in this paper. Of particular importance is that normally a combination-therapy is needed in the vast majority of patients. Synergistic drugs should therefore be combined to achieve an optimal blood pressure control with a minimum of side effects. PMID- 10707482 TI - [Tough working conditions for urologists]. AB - A questionnaire sent by the Swedish Association of Urology to all active Swedish urologists (n = 249) was returned by 89%. Questions concerned actual urological work, emergency duties, education, development facilities and research access, but also job satisfaction, psychological fatigue and emotional stress, as well as each urologist's plans for the future. Answers indicate that Swedish urologists carry a heavy work load and experience considerable demands from patients, relatives and colleagues. Physical and psychological exhaustion are common, and many hope to find work outside hospitals or abroad. The tightening of health-care purses presumably augments the work load, but local factors are also involved. For the future, more and improved educational programs are planned, together with greater participation on the part of junior doctors in organizational and structural processes. Compared to other physicians, urologists show greater reserves of psychic energy, but also signs of increasing intellectual exhaustion. PMID- 10707483 TI - [Forensic psychiatric risk assessment is ethically defensible. The dangerousness and risk of relapse are possible to judge with more precision than random assessment]. PMID- 10707484 TI - [Asia on the verge of a HIV disaster]. PMID- 10707485 TI - [Alarming development of HIV and AIDS in Africa. Political leaders must dare to take more responsibility in the fight against HIV]. PMID- 10707486 TI - [Swimming lice in child day care center--ABC on the treatment of lice infestations. An adequate therapy and contact tracing are successful factors]. PMID- 10707487 TI - [The skeptic and reality. Continuous connection between good and bad theories]. PMID- 10707488 TI - [A reply on MedAnalys: naive bidding]. PMID- 10707489 TI - [Views on Swedish medical education after five years in South Africa: patient centered work can reduce physician's stress]. PMID- 10707490 TI - [Ombudsman of the disabled: the excesses in Vipeholm are not worthy the result]. PMID- 10707491 TI - [Restore the honour of school health services!]. PMID- 10707492 TI - [Consequences of the "seven crowns' reform"--physician--from a free professional to an employee with regulated working hours]. PMID- 10707493 TI - ["The good aging" is a process of adaptation. Not how one has it, but how one takes it!]. PMID- 10707494 TI - [The brain is shaped by stimulation and challenge]. AB - The healthy adult brain retains a certain capacity for plasticity and functional reorganization throughout the life span. Morphologic, neuropsychological and neuroimaging studies have demonstrated that neuronal connections and cortical maps can be remodeled by our experience and activities. Activity-induced increase in neuronal connections may to some extent compensate for neuronal loss during aging. Increased knowledge of the potential capability of the adult brain to compensate for brain lesions is likely to improve rehabilitation strategies. PMID- 10707495 TI - [New discoveries explain how smoking accelerates atherosclerosis]. AB - Although it is well known that cigarette smoking is a risk factor for the development of atherosclerosis, facts have been scarce concerning mechanisms for the effects of smoke on the blood vessel wall. New findings show that endothelial cells are activated by substances found in smoke, so that leukocytes adhere to and interact with them. Among such substances platelet activating factor and nitric oxide have attracted recent attention. This paper reviews hypotheses concerning these reactions. PMID- 10707496 TI - [Boy or girl? Molecular mechanisms in sex differentiation]. AB - A growing number of factors have been recognized as crucial in sexual development, and many clinical conditions have become understandable as mutations in relevant genes have been identified. In some cases this has led to the development of DNA diagnostics, which can be of some aid in the workup of this type of patient. So far it is mainly more serious disorders which have been elucidated, thanks to the identification of extensive mutations in the key genes for sexual development. Exactly to what extent less severe damage in these genes underlies more subtle disorders of sexual development, such as disturbances in pubertal development and diminished fertility, is with few exceptions unknown. We are still aware of only a fraction of the information hidden in our genetic heritage. Developments in this field are nonetheless rapid, and molecular analyses will probably become more and more part and parcel of the workup of patients with a wide variety of disturbances in sexual development. PMID- 10707498 TI - [Marfan syndrome--diagnosis within many specialties]. PMID- 10707497 TI - [Bone marrow depression after azathioprine. New discoveries on an old drug]. AB - Azathioprine, a cytostatic and immunosuppressive drug in use for some 30 years, can give rise to life-threatening neutropenia and thrombocytopenia. This may be caused by unexpectedly high concentrations of cytotoxic metabolites due to abnormally slow inactivation of 6-mercaptopurine (6-MP) by thiopurine S methyltransferase (TPMT) and/or xanthine oxidase. Low TPMT activity may be due to genetic polymorphism or interaction with drugs such as salicylic acid derivatives, while xanthine oxidase may be inhibited by allopurinol. High TPMT activity, on the other hand, may hamper cytostatic treatment. Safer and more effective treatment with azathioprine and its metabolite 6-MP becomes possible with new laboratory methods for pharmacotherapy monitoring. PMID- 10707499 TI - [A life-threatening disease among children often diagnosed too late. Normal sodium levels do not exclude Addison's disease]. PMID- 10707500 TI - [Five breakthroughs for hematology]. PMID- 10707501 TI - [Balint group work against burnout in somatic care. Psychoanalytic thinking in connection with severe cases]. PMID- 10707502 TI - [Norman Bethune, art collector and leading Canadian thoracic surgeon, became people's hero in China]. PMID- 10707503 TI - [Medical consequences of incorrect or delayed results of measurements. What can be learned by the mistakes?]. PMID- 10707504 TI - [A reply on MedAnalys: Illumination of the role of health care authorities would be valuable]. PMID- 10707505 TI - [Evidence-based medicine in everyday work. High priests showing us the right, evidence-based way?]. PMID- 10707506 TI - [Apropos physicians and the spirit of the time 1930-1950: the German influence was normative for our culture]. PMID- 10707508 TI - [Vitamin B12 and chronic fatigue]. PMID- 10707507 TI - [Vitamin B12--which dose and how?]. PMID- 10707509 TI - [What to do against the negative attitude toward vaccinations?]. PMID- 10707510 TI - [Physicians and the spirit of the time. The situation on the labor market is not an excuse]. PMID- 10707511 TI - [Why against the voluntary specialists' examination?]. PMID- 10707512 TI - [On history writing, comment I: History of transplantation--incompletely narrated]. PMID- 10707513 TI - [Centennial for the Meyer-Overton rule: anesthetics and receptors]. AB - As a backlash to the dominance of lipid theories of anesthesia for almost a century, protein theories are prevalent at present. Lipid theories assume nonspecific interaction with membranes. Protein theories assume specific interaction with specific receptors in specific proteins. The Meyer-Overton rule does not specify the anesthetic action site to lipid membranes. The correlation between the olive oil solubility to the anesthetic potency means that the action sites have similar physical properties to olive oil. It does not discriminate between lipids and proteins. Olive oil is homogeneous (isotropic) liquid whereas membranes and proteins are structured (anisotropic). The physical properties of proteins and membranes are not uniform throughout the structure. The rule shows that the anesthetics bind multiple areas in nonspecific proteins and membranes. The diversity of anesthetic structures is difficult to reconcile with the idea that there is a specific receptor on specific proteins. PMID- 10707514 TI - [Perioperative stress response in elderly patients for elective gastrectomy--the comparison between isoflurane anesthesia and sevoflurane anesthesia both combined with epidural anesthesia]. AB - The difference in stress responses between isoflurane anesthesia (I group) and sevoflurane anesthesia (S group) was studied. Twelve patients for elective gastrectomy were divided into two groups: S group, 7 patients, 78 +/- 4.3 years of age, and I group, 5 patients, 77.4 +/- 6.9 years of age. Anesthesia was induced by fentanyl, midazolam and sevoflurane or isoflurane with 100% oxygen. After laryngeal mask air way was inserted under spontaneous ventilation, anesthesia was maintained with air (3 l.min-1), oxygen (2 l.min-1), sevoflurane or isoflurane and epidural block. Vecuronium bromide was given during surgery when needed. The demographic data were not different between the two groups. During operation, it was confirmed that the responses of sympathetic nervous system (epinephrine, norepinephrine) and pituitary-adrenocortical system (ACTH, cortisol) were maintained in both groups. After operation plasma norepinephrine levels increased in both groups. Although the responses of I group tended to be stronger than that of S group, there was no significant difference between the two groups. PMID- 10707515 TI - [Depressive effects of propofol on apoptotic injury and delayed neuronal death after forebrain ischemia in the rat--comparison with nitrous oxide-oxygen isoflurane]. AB - We investigated the brain protection effects of propofol anesthesia and nitrous oxide-oxygen-isoflurane anesthesia (GOI) using forebrain ischemic model of male Sprague-Dawley rats. Propofol group (P, n = 15) was anesthetized with propofol, oxygen and nitrogen (FIO2 = 0.33), and isoflurane group (GOI, n = 15) with 66% nitrous oxide, 33% oxygen and 1.2% isoflurane under mechanical ventilation. The anesthesia was deepened until electroencephalographic burst suppression appeared in each group. The bilateral common carotid arteries were, then, occluded for 10 minutes while the blood pressure was maintained at about 40 mmHg by venesection. The venesected blood was returned at the end of ischemic period. The animals were kept and fed in cage after emergence. On the day 2, 4, and 7, five animals of each group were sacrificed and the microscopic samples were obtained. The CA-1 cells of hippocampus were then stained with hematoxylin and eosin for the delayed neuronal death (DND) and with TUNEL method for the apoptosis. Propofol reduced the apoptosis, i.e., reduced the TUNEL positive cell count (GOI = 121.2 +/- 25.2.mm-1; P = 53.8 +/- 11.4.mm-1; P < 0.01; mean +/- SD) on the day 2 after ischemia, and also reduced the delayed neuronal death (alive CA-1 cell count; GOI = 18.1 +/- 8.9.mm-1; P = 33.1 +/- 12.8.mm-1; P < 0.01) on the day 7 after ischemia. It is important to determine the recovery interval after brain ischemia in detection of DND and apoptosis. We conclude that propofol inhibits neuronal apoptosis after brain ischemia and consequently reduces the delayed neuronal death in the CA-1 pyramidal cell layer of the hippocampus. PMID- 10707516 TI - [Halothane anesthesia decreases the level of interstitial striatal dopamine of awake freely moving rats in an in vivo microdialysis study]. AB - We investigated the effect of halothane on the level of interstitial dopamine of in vivo awake, free moving rats brain striatum using microdialysis techniques. Rats were implanted a microdialysis probe to right striatum of the brain and administered 1.5% of halothane (approximately 1.2 MAC) for 1 or 2 hours, and dialysates from the probe were determined every 20 minutes. Halothane anesthesia reduced the amount of dopamine derived from dialysate, and after discontinuation of halothane and at emergence from anesthesia, the level of dopamine was increased. The levels of metabolites of dopamine during anesthesia were increased lineally in a time dependent manner. We hypothesized that halothane might increase the rate of re-uptake of dopamine at nerve endings and decreased level of interstitial dopamine is compensated by dopamine releases during anesthesia. PMID- 10707517 TI - [Myocardial ischemia score as a preoperative screening method for intraoperative myocardial ischemia]. AB - Three hundred patients for abdominal surgery with risk factors of ischemic heart disease (IHD), such as hypertension, diabetes mellitus, hyperlipidemia, smoking, old age, obesity, familial history, electrocardiographic abnormality, other perivascular diseases and male, were included in this study. Patients older than forty years were included in the study. ST segment in lead II and V5 was recorded by ST trend monitor from the time of entering the operating room to the time of exit and intraoperative myocardial ischemia occurred in 58 patients (19.3%). Correlation coefficients between each risk factor of IHD and intraoperative myocardial ischemia were calculated by multiple regression analysis and myocardial ischemia score (MIS) was determined. Intraoperative myocardial ischemia increased in patients with more than 10 points of MIS by discriminant analysis and hitting ratio of MIS was 86.3%. PMID- 10707518 TI - [Factors influencing the level of spinal anesthesia: (II). Patient characteristics and technique of injection]. AB - The extent of sensory block is determined by the cephalad distribution of the local anesthetic in the cerebrospinal fluid and uptake by neuronal tissue in sufficient amounts to produce the block. Out of many factors that have been considered to affect the distribution, this paper discusses factors involved in patient characteristics and technique of injection; the age of the patient, the volume of cerebrospinal fluid, the rate of injection, the site of injection, and the position of the patient. Each of the factors does not seem to have a major impact on the level of sensory block as compared with characteristics of anesthetic solutions discussed in the previous review, but this has not been fully understood. Recent studies have shown that the management of posture to control the level of spinal anesthesia is not so easy as previously thought. It is my conclusion that knowledge of these factors is essential in the performance of reliable and safe spinal anesthesia. PMID- 10707519 TI - [Evaluation with blink reflex of bilateral facial palsy]. AB - We examined the accuracy of electroneuronography (ENoG) and blink reflex as prognostic indicators in four patients with bilateral facial palsy. Electrophysiological investigation consisted of the recording of ENoG amplitude and R1 amplitude of blink reflex elicited by the peripheral nerve stimulation. Examinations were performed at the first visit, after 10 days, 2, 4 and 6 weeks. ENoG amplitude was less reliable for the neurological evaluation of bilateral facial palsy, because in three cases amplitude ratio of affected side to the other side was more than 100%. R1 amplitude of blink reflex recovered satisfactorily in two cases of good outcome. In other two cases of poor outcome, R1 amplitude was absent during the examination period. These findings suggest that analysis of blink reflex provides valuable information for evaluation of the prognosis of patients with bilateral facial palsy. PMID- 10707520 TI - [Supplement of ulinastatin on renal function after cardiopulmonary bypass]. AB - The effects of Miraclid (ulinastatin) on renal tubular function after open thorax surgery under cardiopulmonary bypass were investigated. On the 3rd day after open thorax surgery, which had lasted more than 127 min under cardiopulmonary bypass, the levels of urinary ulinastatin in the Miraclid group and control (without Miraclid) were 170 IU.mg Cr-1 and 95 IU.mg Cr-1, respectively. In the Miraclid group, 300,000 units.day-1 of Miraclid was administrated for three postoperative days. N-acetyl-beta-d-glucosaminidase in urine as a marker of tubular function rose significantly on the seventh postoperative day in the control group but not in patients with Miraclid group. These data suggested that Miraclid 300,000 units.day-1 was needed to protect renal tubular function and more than that dose was needed to prevent the deterioration of renal function after open thorax surgery after cardiopulmonary bypass lasting more than 127 min. PMID- 10707521 TI - [Perioperative management for total en bloc spondylectomy--the effects of preoperative embolization and hypotensive anesthesia]. AB - We retrospectively evaluated the effects of preoperative embolization and hypotensive anesthesia on total en bloc spondylectomy (TES) for solitary spinal metastases. In ten patients (treatment group), feeding arteries of spinal metastases were embolized preoperatively and controlled hypotensive anesthesia was induced during operation. In other ten patients (control group), these treatments were not applied. Intraoperative blood loss as well as the amount of blood transfused in the treatment group were significantly lower than those in the control group. Moreover, postoperative platelet counts in the treatment group were significantly higher than those in the control group. These findings indicate that embolization of feeding arteries of metastases and hypotensive anesthesia decrease intraoperative blood loss and may prevent postoperative complications in TES. PMID- 10707522 TI - [Optimal dose of indocyanine-green injected from the peripheral vein in cardiac output measurement by pulse dye-densitometry]. AB - Cardiac output is measured by pulse dye-densitometry using indocyanine-green (ICG). This cardiac output is estimated by correlating with the cardiac output measured by pulmonary artery catheter using thermodilution method. Twenty-four patients scheduled for elective cardiovascular surgeries under general anesthesia were studied. The pulse dye-densitometry monitoring system used was DDG-2001 (Nihon Kohden, Japan). In group A (13 patients: 19 times), ICG was administered from the peripheral vein as bolus doses of 5, 10 or 20 mg (5 mg.ml-1 water solution). In group B (11 patients: 12 times), ICG was administered from the peripheral vein as bolus doses of 20, 10 or 5 mg (5 mg.ml-1 water solution). The correlation (Pearson's correlation coefficient) and precision (the method proposed by Bland and Altman) compared with cardiac output measured by pulmonary artery catheter were examined. Better correlation and precision were recognized after 20 mg ICG injection than 5 or 10 mg ICG injection. In conclusion, the measurement of cardiac output by pulse dye-densitometry with peripheral vein ICG injection was useful using a bolus dose of ICG 20 mg. PMID- 10707523 TI - [Clinical evaluation of accuracy of a new disposable pump (Syrinjector) for continuous epidural infusion]. AB - We evaluated the accuracy of a new disposable pump (Coopdech Syrinjector), which employed the negative pressure made by injecting fluid into the pump for continuous epidural infusion. Thoracic epidural catheter was placed at the T6-T8 level prior to the lung surgery. The continuous epidural infusion of 0.25% bupivacaine at 2 or 3 ml.h-1 was started immediately after the surgery. The residual volume of bupivacaine was recorded. From our results, the injection speed of Syrinjector was satisfactory for clinical use of continuous epidural infusion of bupivacaine. However, when this pump is reused or the volume of the residual local anesthetics is less than 10 ml, the infusion accuracy will be potentially imprecise. PMID- 10707524 TI - [A case of free rupture of abdominal aortic aneurysm into the peritoneal cavity during posture change after induction of anesthesia]. AB - We report a case in which posture change for radiography after induction of anesthesia caused free rupture of the abdominal aortic aneurysm (AAA) into the peritoneal cavity, resulting in shock, although in the patient an AAA had ruptured into only the retroperitoneal space and hemodynamics had been stable preoperatively. The massive bleeding was controlled with autotransfusion using a washing salvaging autotransfusion device and a roller pump for hemodialysis. In addition, international mild hypothermia was effective for protection of the brain from suspected ischemia during shock. Meticulous attention should be paid for anesthetic management of patients with ruptured AAA even if their hemodynamic status is stable. PMID- 10707525 TI - [Anesthetic management of patients with tracheal stenosis for endoscopic treatment: usefulness of laryngeal mask airway]. AB - We report two cases of tracheal stenosis for endoscopic treatment under general anesthesia with laryngeal mask airway. The tracheal stenosis of the two patients was so close to the glottis that endotracheal tube could not be inserted, and laryngeal mask airway was beneficial for maintaining airway and obtaining operating field. During the procedure, patients breathed spontaneously and we could support their ventilation easily and sufficiently. Endoscopic treatment of the airway obstruction by Nd-YAG laser associated with balloon dilatation and stent is an effective method of relieving the distressing symptom of asphylaxia, and laryngeal mask airway is considered to be useful for performing successful endoscopic procedure. PMID- 10707526 TI - [A case of acute exacerbation of idiopathic interstitial pneumonia after pneumonectomy]. AB - A 52-year-old male was scheduled for right upper lobectomy due to lung cancer. The patient was diagnosed as having stable idiopathic interstitial pneumonia (IIP) based on preoperative lung biopsy, physical examination and laboratory data. He received prophylactic steroids to prevent an acute exacerbation of IIP. However, he fell into severe hypoxia 5 days after operation, and died of respiratory failure 43 days after operation. It must be born in mind that the patients with IIP may suffer exacerbation even if their lungs have been diagnosed as in stable stage. Prophylactic use of steroids may cause a deterioration in IIP due to delayed healing and infection. PMID- 10707527 TI - [Anesthetic management of a patient with amyotrophic lateral sclerosis]. AB - A 49-year-old male with amyotrophic lateral sclerosis (ALS) was scheduled for gastrectomy. Anesthetic management was performed under general anesthesia with sevoflurane and epidural anesthesia with lidocaine. He showed increased response to vecuronium under monitoring of neuromuscular block. But he responded favorably to anticholineesterase. He had little pain and showed no progress in neurological symptoms in the postoperative period. Neuromuscular monitoring is essential in administrating non-depolarizing neuromuscular blocking agents to patients with ALS, and epidural anesthesia may be useful for perioperative management of patients with ALS. PMID- 10707528 TI - [Anesthesia, department of anesthesiology and anesthesiology--why has anesthesiology not been accepted socially in Japan?]. AB - An incorrect Japanese terminology of "Masuigaku [symbol: see text]" has been used widely to express "anesthesiology" or "anaesthetics" [symbol: see text] since the first Department of Anesthesiology was established in Tokyo University in 1952. The reason why the nomenclature "Masui-gaku" is wrong is as follows: Japanese nomenclatures for clinical medical sciences should include a Chinese character "Ka [symbol: see text]" such as "nai-ka-gaku" for internal medicine, "ge-ka-gaku" for surgery and "gan-ka-gaku" for ophthalmology. Accordingly the name "Masui gaku" is erroneous to mean "Anesthesiology" and it should be "Masui-ka-gaku" [symbol: see text]. Thus a big confusion has occurred among lay people as well as many physicians in medical field. "Ma-sui" is etymologically a Japanese word which Dr Seikei Sugita coined when he translated a Dutch edition of J. Schlesinger's monograph on ether anesthesia in 1850. "Ma [symbol: see text]" means analgesia or loss of regional sensation and "Sui [symbol: see text]" means loss of consciousness. Most people consider that "Ma [symbol: see text]" is originated from "[symbol: see text] (Hemp, Asa)" or "[symbol: see text] (Marihuana, Taima)", however, this is definitely incorrect and "Ma [symbol: see text]" of "Ma-sui" has no direct relation with the pharmacological effect of hemp. Thus the misuse of "Masui-ga-ku" might have caused serious academic and social confusions, such as misunderstanding of anesthesiologists as comedical technicians, leading to a poor social acceptance of anesthesiology and anesthesiologists for these fifty years in Japan. To correct this confused situation I would like to ask our colleagues to use correctly these nomenclatures. PMID- 10707529 TI - [A questionnaire survey of the panel discussion at the annual meeting of the Japan Society of Anesthesiologists]. AB - Two hundred questionnaire sheets were distributed to the audience of the panel discussion session titled "Progresses of balanced anesthesia" at the annual meeting of the Japan Society of Anesthesiologists. The audience was requested to rate their interest to the program and also rate the usefulness of the presentations by each panel. One hundred and seventeen questionnaire sheets (58%) were returned. Eighty five % of the returned survey expressed interests in the program. Forty three % expressed their interests in altering their anesthesia techniques along the line of the recommendation by the panels. These data suggest that the program met the educational need of the audience. Questionnaire survey may be useful in identifying the need of the members, and thus helpful in planning future meetings of the society. PMID- 10707530 TI - [The toxicity and interferon-inducing activity of the lipopolysaccharides of Cytophaga sp. (strains 81 and 92)]. AB - The experiments on mice and cell culture of a kidney of green monkey Vero have shown that lipopolysaccharides (LPS) of the studied strains 81 and 92 of Cytophaga sp. were nontoxic. They were less active as compared to LPS of other Gram-negative bacteria as to interferon-inducing activity. A comparative study of initial and dephosphorylated LPS has shown that phosphate groups were not responsible for the interferon-inducing activity of LPS of Cytophaga genus representatives. PMID- 10707532 TI - [The bacteriocinogenic and lysozyme-synthesizing activity of lactobacilli]. AB - It was shown that lactic acid bacteria isolated from different sources produced along with lactic acid the bacteriocine substances. Strains producing bacteriocines were mainly isolated from digestive tract of people and calves. Bacteriogenic properties have been shown more frequently in the case of lactobacilli (7.8%) more rarely--in coccoid forms of lactic acid bacteria (4.5%). At the same time bacteriocine activity was higher in the case of Enterococcus and depended both on the strain type and cultivation conditions. Synthesis of bacteriocines started from the hours of phase of the culture growth and reached maximum at the beginning of stationary phase. The bacteriocine accumulation rate depended on composition of the growth medium. PMID- 10707531 TI - [The role of the structural components of bacterial lipopolysaccharide in its inductive immunosuppressive activity]. AB - The inductive immunosuppressive activity of lipopolysaccharides (LPS) and structural parts (O-chains,cor,lipid A) obtained from bacteria of the genus Shigella and Escherichia coli was studied. LPS preparations were extracted by phenol-water method. Its structural components were obtained by the acetic acid hydrolysis and gel filtration. It has been shown that all these preparations did not change the level of delayed type hypersensitivity (DTH) to test-antigen in mice in the case of intraperitoneal injection. They did not also induce immunosuppressive activity in avirulent bacterial strains. After the redox treatment all LPS preparations acquired the ability to induce immunosuppressive property in avirulent bacteria. As a result of redox treatment the immunosuppressive activity appeared in lipid A preparations. O-chains and cor preparations were not active. Immunosuppressive activity was lost after treatment with phenol. It is supposed that active chemical groups are located in lipid A. PMID- 10707533 TI - [The effectiveness of thin-section computed tomography in diagnosing bullous lesions in patients with spontaneous pneumothorax]. AB - We investigated the effectiveness of thin-section computed tomography (CT) for the diagnosis of bullous lesions in patients with spontaneous pneumothorax. The study group consisted of 74 patients. Apical regions of the lung were scanned for lesions by thin-section CT prior to operation. The presence, number, and locations of bullous lesions were assessed. Bullous lesions were also classified into 2 groups according to their shape as demonstrated by CT findings. Operative findings confirmed that 73 of the 74 patients had bullous lesions. Of these, 33 had a single bulla and 40 had multiple bulla. Thin-section CT accurately detected the presence and location of the bullous lesions, accurately identified the number in all but 6 patients with multiple bulla (accuracy: 91.8%), and also classified them accurately by type. Precise evaluations of bullous lesions are crucial to the treatment of patients with spontaneous pneumothorax. The presence of bullous lesions, as well as their location and shape, are important factors in determining whether to operate or not. Such information also allows for a better understanding of the surgical options available. Our study demonstrated that thin section CT of apical regions of the lung is an effective diagnostic procedure for patients with spontaneous pneumothorax. PMID- 10707534 TI - [Clinical investigation of pulmonary tuberculomas diagnosed by video-assisted thoracoscopic biopsy]. AB - We investigated the clinical characteristics of pulmonary tuberculomas and discussed strategies for their diagnosis. We compared a group of patients with lesions that had been diagnosed by bronchoscopy (the BF group: 15 patients, 17 lesions) with a group whose lesions had been diagnosed by thoracoscopy (the VATS group: 19 patients, 20 lesions). The VATS group included patients with lesions that could not be diagnosed by bronchoscopy alone. The 2 groups did not differ significantly in terms of patient age, gender, lesion size, number of bronchoscopic examinations, or tuberculin test findings. In both groups, the lesions always existed just below the pleural surface and were mostly located in the upper lobes of the lungs. Lesion locations in the middle lobe and lingular segment were observed only in the BF group, whereas locations in the lower lobes were observed only in the VATS group. Tuberculomas in 2 BF group patients (11%) and 10 VATS group patients (50%) were diagnosed on the basis of histopathologic findings in the absence of bacteriological evidence. No postoperative complications were observed in the VATS group, and all patients in this group were discharged from the hospital within 1 week. No relapse of tuberculosis was observed in either group following anti-tuberculous therapy. Video-assisted thoracoscopic biopsy is a reliable and minimally invasive diagnostic procedure, and should be performed on any solitary pulmonary lesion that cannot be diagnosed by bronchoscopy. It is important that tuberculomas be quickly and accurately diagnosed not only in order to distinguish them from lung cancer, but also to treat the patient's tuberculosis at an early stage and prevent it from spreading. PMID- 10707535 TI - [Relationship between p 53 gene mutation and MDR 1 gene expression in surgically resected non-small cell lung cancer]. AB - The resistance of tumor cells to chemotherapeutic drugs is a major obstacle to successful cancer chemotherapy. Expression of the MDR 1 gene, which encodes for a transmembrane efflux pump (P-glycoprotein), leads to decreased intracellular accumulation and resistance to a variety of anticancer drugs. Recently, one mutant p 53 form was shown to stimulate the MDR 1 gene promoter in vitro, whereas wild-type p 53 repressed this activity. We examined the relationship between p 53 gene mutation and MDR 1 gene expression in specimens from non-small cell lung cancer patients. Tumor samples were obtained from 21 patients during surgery. Mutations of exon 5 through exon 8 of the p 53 gene were detected by the polymerase chain reaction single strand conformation polymorphism method. MDR 1 expression was semi-quantified by the reverse transcriptase polymerase chain reaction method. We identified p 53 gene mutation in samples from 7 patients. MDR 1 gene expression was observed in samples from 20 patients. The expressivity of the MDR 1 gene tended to be higher in patients with adenocarcinoma. No significant relationship between p 53 mutation and MDR 1 expressivity was observed in our study. PMID- 10707536 TI - [ADOC regimen for unresectable advanced thymic cancer]. AB - Between 1996 and 1998, we treated 6 patients with unresectable and advanced thymic cancer (stages IVa and IVb). All received 50 mg/m2 of cisplatin and 40 mg/m2 of doxorubicin intravenously (i.v.) on day 1,0.6 mg/m2 of vincristine i.v. on day 3, and 700 mg/m2 of cyclophosphamide i.v. on day 4; ADOC regimen, respectively at 3-4 week intervals. Four patients obtained a partial response (PR) after ADOC chemotherapy and the overall clinical response rate was 67%. No life-threatening side effects were noted. In 2 patients, cisplatin plus VP-16 chemotherapy failed to demonstrate any benefits prior to the ADOC regimen. Radiotherapy was initiated after the achievement of PR in the other 2 patients. ADOC chemotherapy appears to be an effective treatment for thymic cancer. PMID- 10707537 TI - [Pulmonary infarction diagnosed by transbronchial lung biopsy]. AB - A 43-year-old man complained of chest pain, fever, and hemosputum in February 1997. Chest X-ray films revealed small opacity in the right lower lung field. The patient received therapy for pneumonia and his condition gradually improved. However, on March 21 he was admitted to our hospital with worsened chest pain and radiographic findings. A transbronchial lung biopsy revealed thrombus and necrosis, thus yielding a diagnosis of pulmonary infarction. The patient did not exhibit any underlying disease or coagulation abnormalities. Treatment with ticlopidine resulted in a favorable course. This was a rare case of pulmonary infarction in which TBLB findings led to the diagnosis. PMID- 10707539 TI - [Acute respiratory failure caused by inhalation of waterproofing spray fumes]. AB - A 25-year-old woman was admitted to our hospital because of dry cough, slight fever, and severe dyspnea 5 hrs after inhalation of waterproofing spray. She had used the spray indoors near an oil heater, and then had smoked with spray contaminated fingers. Chest roentgenograms revealed diffuse interstitial shadows mixed with patchy alveolar infiltration, and computed tomographic (CT) scans confirmed diffuse infiltration in both lungs. Marked leukocytosis and severe hypoxemia were noted. A transbronchial lung biopsy performed 4 days later demonstrated extensive alveolitis characterized by edema in alveolar septa and marked neutrophil migration into alveoli as well as alveolar septa. Oxygen therapy and the administration of methylprednisolone (1 g/day) achieved a complete recovery in about 1 week. We speculated that acute lung injury in this patient may have been induced by direct inhalation not only of the waterproofing spray itself, but also of spray by-products resulting from heat-decomposition. When using waterproofing spray, precautions should be taken to avoid inhaling the spray fumes or the more toxic by-products of thermal degradation. PMID- 10707538 TI - [Pulmonary radiation injury manifested by signs of bronchiolitis obliterans with organizing pneumonia after postoperative breast cancer radiotherapy]. AB - A 67-year-old woman underwent surgery for cancer of both breasts (right: mastectomy, left: conserving surgery), and received 60 Gy radiation to the left postoperative breast. Three months later, cough and fever developed. A chest radiograph demonstrated infiltrative shadows in the left lung field. Transbronchial lung biopsy specimens disclosed organizing exudates in the alveolar spaces and bronchioles. After treatment with prednisolone, the clinical symptoms and radiographic infiltrates disappeared. This was a case of pulmonary radiation injury pathologically manifested by signs of bronchiolitis obliterans with organizing pneumonia. PMID- 10707540 TI - [Changes in tracheal and lung sounds before and after resection of adenoid cystic carcinoma of the trachea]. AB - A 79-year-old woman was admitted to our hospital with complaints of wheezing and dyspnea. Adenoid cystic carcinoma was diagnosed from the findings on biopsy specimens obtained by fiberoptic bronchoscopy. The tumor was resected and end-to end anastomosis was performed. Precision analyses of lung sounds were conducted before and after the operation. During eupnea the tracheal sounds prior to operation contained an accentuated, high-frequency component at about 1,500 Hz. The tracheal sounds included a single monophonic wheeze during deep inspiration and 5 wheezes during forced expiration. By contrast, lung sounds in the chest wall were almost normal both before and after the operation. We concluded that increases in high-frequency components in tracheal sounds and the manifestation of various wheezes can be useful in diagnosing tracheal stenosis and/or tumors. PMID- 10707541 TI - [Small cell lung cancer associated with paraneoplastic cerebellar degeneration and Lambert-Eaton myasthenic syndrome]. AB - A 57-year-old man was admitted to our hospital in November 1997 because of dysarthria, progressive ataxia, generalized weakness, and incoordination in both hands. He had been aware of the dysarthria 6 months earlier. Chest roentgenograms and computed tomographic films disclosed a 5 cm x 6 cm mass in the left S3b. The patient was given a diagnosis of small cell lung cancer (T3N2M0, stage IIIA) associated with paraneoplastic cerebellar degeneration (PCD). Three courses of chemotherapy (carboplatin and etoposide) eliminated the tumor and slightly alleviated the PCD symptoms. In March 1998, electromyograms revealed a fall in the single-stimulated M wave and a waxing phenomenon that had not been observed on admission. Anti-P/Q type voltage gated calcium channel antibody was detected in serum samples obtained on admission and after chemotherapy. These findings confirmed an association with Lambert-Eaton myasthenic syndrome. No relapse of the tumor has been observed 15 months after the last course of chemotherapy. PMID- 10707542 TI - [Nitric oxide inhalation as an effective therapy for acute respiratory distress syndrome due to near-drowning: a case report]. AB - A 16-year-old boy with acute respiratory distress syndrome (ARDS) due to near drowning was admitted to our hospital. ARDS was treated with low-level nitric oxide (NO) inhalation (ranging from 4 ppm to 1 ppm) for 24 days. Oxygenation was improved and pulmonary hypertension was reduced after NO inhalation, but systemic blood pressure, heart rate, and cardiac output were not affected. PaO2 improved from 153 Torr to 354 Torr under identical ventilating conditions (F1O2 1.0), and mean pulmonary arterial pressure fell from 40 mm Hg to 27 mmHg. It has been reported that NO inhalation alleviates ventilation-flow mismatch and pulmonary hypertension. It is unclear, however, whether this therapy improves the prognosis for ARDS. In our patient, NO inhalation was effective in alleviating the oxygenation impairment and pulmonary hypertension associated with ARDS. PMID- 10707543 TI - [Four cases of cancer of unknown primary site that manifested as mediastinal metastatic lesions]. AB - We encountered 4 patients with cancers of unknown primary sites that were manifested by mediastinal lesions. Patient 1 was a 58-year-old man with enlarged superior mediastinal lymph nodes. An exploratory thoracotomy yielded a diagnosis of lymph node metastasis of poorly differentiated adenocarcinoma. The patient was treated with chemotherapy and radiation therapy. Patient 2 was a 68-year-old man with a tumor in the right superior mediastinum. A total resection of the tumor was performed through a thoracotomy. The diagnosis was lymph node metastasis of squamous cell carcinoma, and treatment consisted of irradiation. A tumor shadow in the right upper lobe appeared 14 months after the thoracotomy, and was considered to be a primary lesion requiring a right pneumonectomy. The patient died of hepatic metastasis 6 months after the second operation. Patient 3 was a 59-year-old man with mediastinal and hilar lymph node swelling. Mediastinoscopic findings resulted in a diagnosis of squamous cell carcinoma. Because of the patient's insistence, only radiation therapy was performed. Patient 4 was a 65 year-old woman with a tumor in the right superior mediastinum who underwent a median sternotomy for total resection of the tumor. The pathological findings were strongly suggestive of metastasis of clear cell carcinoma. Patients 1, 3, and 4 were alive 33, 24, and 51 months, respectively, after their initial operation, without detectable primary sites. Patient 2 was considered to have had T 0 N 2 lung cancer. PMID- 10707544 TI - [Two cases of Kimura's disease associated with bronchial asthma]. AB - We encountered two rare cases of Kimura's disease associated with bronchial asthma presenting eosinophilia and hyperimmunoglobulinemia E. Patient 1 was a 26 year-old man who had been admitted to our hospital with recurrent increase in left parotid mass in May 1997. He had previously undergone surgery for local excision at another hospital in September 1987; the excised specimens were re evaluated and the diagnosis of Kimura's disease was confirmed. Because the patient was suffering from an acute asthma attack on admission, prednisolone (PSL) 30 mg/day was administered orally. PSL reduced the parotid mass and improved control of the asthma. Patient 2 was an 18 year-old man who had been given a diagnosis of Kimura's disease on the basis of histologic findings from a biopsy specimen of a subcutaneous tumor in the left cheek in 1988. Following the diagnosis, the patient was treated with methotrexate for the first several months, and then with loxioprofen for 9 years, but the size of the mass remained unchanged. Bronchial asthma developed in this patient in 1995 and had been treated with theophylline. However, because this therapy caused a deterioration of asthma control, the patient was admitted to our hospital in October 1997 for the treatment of bronchial asthma. Inhaled corticosteroids (beclometasone 0.8 mg/day) in addition to theophylline alleviated the patient's asthma symptoms and yielded improved lung function. Because few cases of Kimura's disease associated with bronchial asthma have been reported, patients with eosinophilia and hyperimmunoglobulinemia E were not necessarily considered at high risk for the onset of bronchial asthma. PMID- 10707545 TI - [An autopsy case of interstitial pneumonia probably induced by Sho-saiko-to]. AB - A 66-year-old woman had been treated for 3 years by her local physician with Sho saiko-to for chronic hepatitis C virus (HCV) infection and liver cirrhosis. She was admitted to our hospital because of cough, fever, and infiltrative shadows on chest x-ray films. Sho-saiko-to-induced pneumonitis was diagnosed and steroid therapy started. Though a temporary improvement was observed, interstitial pneumonitis relapsed and the patient died of respiratory failure and liver dysfunction. Autopsy findings showed diffuse alveolar damage and honeycombing. Furthermore, reverse-transcriptase polymerase chain reaction techniques detected HCV-RNA in specimens of fibrotic lung tissue. For comparison, HCV-RNA was not histologically detected in lung tissue specimens from 4 control subjects who were positive for HCV antibodies but who did not have interstitial lung disease. It was speculated that the progression of interstitial pneumonia in the present case may have been caused by HCV in combination with Sho-saiko-to-induced lung injury. PMID- 10707546 TI - [A case of preoperatively diagnosed primary pulmonary leiomyosarcoma]. AB - A 29-year-old woman had been suffering from right back pain for 3 months. Chronic pulmonary thromboembolism was suspected and she was referred to our hospital. She presented with no risk factors for thromboembolism, and during the previous 6 months had lost 4 kg in body weight. Chest radiography showed nodular shadows in the lower field of the right lung. Contrast-enhanced computed tomography demonstrated a filling defect in the right pulmonary artery and nodular lesions in the lower field of the right lung, which were considered to be signs of pulmonary infarction. Absence of perfusion into the right lung was demonstrated by a perfusion scan. Right heart catheterization showed normal pressure in the pulmonary arteries, and pulmonary angiography showed an abrupt cutoff of the right pulmonary artery, which was similar to the finding of pulmonary thromboembolism. A transvenous catheter suction biopsy was performed in the right pulmonary artery and the histopathologic findings yielded a diagnosis of leiomyosarcoma. The patient underwent surgical resection under total cardiopulmonary bypass. A large tumor completely filled the right main pulmonary artery and invaded the posterior wall of the pulmonary trunk close to the left main pulmonary artery. Primary pulmonary leiomyosarcoma is a rare tumor and its prognosis is very poor. Radical surgical resection is the only effective treatment, but early diagnosis is very difficult. Transvenous catheter suction biopsy is a useful procedure for the early diagnosis of pulmonary artery sarcoma. PMID- 10707547 TI - [Relative importance of insulin actin in insulin's target tissues]. AB - Insulin is a hormone with various metabolic effects such as glucose uptake and glycogen synthesis in insulin's target tissues. Skeletal muscle has been considered to be a principal site of insulin resistance in type 2 diabetes. Studies of mice with muscle specific-knockout of the insulin receptor demonstrated that although skeletal muscle was insulin resistant, glucose tolerance was near normal at the whole body, indicating the possible importance of liver. beta cell specific disruption of the insulin receptor indicated that primary insulin resistance at the beta cells resulted in a defect of insulin secretion and impaired glucose tolerance. These animal models suggest that a combination of insulin resistance in muscle, liver, adipose cells, and beta cells appears to create the phenotype of type 2 diabetes. PMID- 10707548 TI - [Insulin signalling system and mechanism of insulin resistance]. AB - Insulin exerts wide variety of biological effects through interaction with its specific receptor, which belongs to a large family of receptor tyrosine kinases. The activated insulin receptor phosphorylates the intracellular substrate IRS protains, which then bind various signalling molecules that contain Src homology 2 domains. The first downstram molecule that was shown to associate with IRS protains is PI3-kinase. PI3-kinase contributes to a wide variety of biological actions. Both Akt(PKB), a serine-threonine kinase with a PH domain, and atypical PKC(PKC zeta, PKC lambda) have been implicated as downstream effectors of PI3 kinase. Insulin resistance contributes to the pathogenesis of NIDDM. Both primary, genetically, and secondary, environmentally factors are important for insulin resistance. The secondary factors include hyperglycemia, hyperlipidemia, obesity, TNF alpha, FFA(free fatty acid). PMID- 10707549 TI - [Impairments of insulin receptor function in insulin resistant states]. AB - Type 2 diabetes is characterized by insulin resistance in skeletal muscle. Since the molecular mechanism of insulin resistance is still unknown, insulin receptor dysfunction including abnormal IRS-1 phosphorylation is considered to be responsible for insulin resistance in some pathological states. Obesity is one of major factors to induce insulin receptor dysfunction. Regarding the mechanism of insulin resistance related obesity, the increased expression of Tumor necrosis factor alpha and abnormality in PTPase in skeletal muscle are postulated. As well as obesity, prolonged hyperglycemia, dyslipidemia and hypertension also induce the impairment of insulin receptor function. Therefore, the enhancement of insulin sensitivity by modulating these factors is a possible treatment modality in insulin resistant states. PMID- 10707550 TI - [Insulin resistance and glucose transporter]. AB - Insulin stimulates glucose transport in muscle and adipose tissue by promoting the appearance of GLUT4, the main glucose transporter isoform in these tissues, on the cell surface. Insulin resistance is instrumental in pathogenesis of type 2 diabetes mellitus and involves decreased glucose transport activity in these tissues. No significant differences are observed between the diabetic and non diabetic subjects in muscle GLUT4 levels. Polymorphism in the GLUT4 gene, which is very rare, has the same prevalence between subjects with type 2 diabetes mellitus and the non-diabetic subjects. The most likely explanation for the insulin resistance is a defect in insulin signaling pathways or GLUT4 intracellular trafficking pathways. PMID- 10707551 TI - [Genetic factors and insulin resistance]. AB - Diabetes is characterized by impaired insulin secretion from pancreatic beta-cell and/or insulin resistance in peripheral tissues. Genetic factors affecting the insulin sensitivities have been described, including the mutations of genes of insulin receptor, insulin receptor substrate-1, glycogen synthase, uncoupling proteins, beta3-adrenergic receptor, and peroxisome proliferator-activated receptor gamma. Mutant insulin receptors might act as a monogenic factor, but most of the others act as one of the genetic factors of a polygenetic disease. PMID- 10707552 TI - [Insulin resistance and cytokine, cytokine receptor]. AB - Insulin resistance is associated with many common diseases including diabetes mellitus, hyperlipidemia and hypertension, and plays an important role for determining their clinical courses. Obesity is a multifactorial syndrome characterized by an excessive adipose tissue accumulation, and is associated with acquired insulin resistance. Adipose tissue, acting as one of the endocrine organs, has been revealed to produce and secrete some bioactive molecules, "adipocytokines", which regulate cell growth and/or metabolic pathways. Tumor necrosis factor(TNF)-alpha is also synthesized by adipocytes, and is involved in the expression of peripheral insulin resistance. This review deals with molecular mechanisms of the TNF/TNF receptor system promoting insulin resistance, and its prevention by the insulin-sensitizing drugs, thiazolidinediones. PMID- 10707553 TI - [Insulin resistance, role of leptin and leptin receptor]. AB - Leptin, the obese gene product, is an adipocyte-derived satiety factor which is involved in the regulation of food intake and energy expenditure. Obesity often accompanies insulin resistance and high levels of leptin. In in vitro studies, leptin has been reported to increase fatty acid oxidation and decrease fatty acid synthesis in adipocytes and hepatocytes. The direct effects of leptin on glucose metabolism and insulin signaling have not been clarified yet. In in vivo studies, however, leptin has been reported to improve insulin sensitivity and glucose metabolism in normal and obese rodents acting mainly through hypothalamus. Moreover leptin has been reported to have antidiabetic effects in insulin deficient diabetes rats and lipoatrophic diabetes mice. It is suggested that leptin modulates insulin sensitivity and glucose disposal and that leptin may have a pathophysiological and therapeutic implications in diabetes. PMID- 10707554 TI - [Insulin resistance and beta 3-adrenergic receptor function]. AB - Beta 3-adrenergic receptors are expressed predominantly on white and brown adipocytes. Activation of the receptor stimulates lipolysis in adipose tissues and increases energy expenditure by thermogenesis in brown adipocytes. It is proposed that dysfunction of beta 3-adrenergic receptor result to obesity and insulin resistance. Trp64Arg mutation of human beta 3-adrenergic receptor gene is found frequently in Pima Indians and Japanese, and rare in Caucasians. Although the mutation have little, if any, functional disturbance especially in acute phase in vivo, it is reported to associate to obesity and earlier onset of type 2 diabetes. The role of beta 3-adrenergic receptor in insulin resistance is still unknown in detail. Further studies and clinical applications are expected. PMID- 10707555 TI - [Adipocyte function and insulin resistance]. AB - The importance of free fatty acids (FFA) and adipocytokines released from adipocytes in the development of insulin resistance is discussed in this review article. FFA may cause insulin resistance through so-called Rundle cycle and may also inhibit glucose uptake by skeletal muscles. Adipocytokines, bioactive substances secreted from adipose tissue may have important roles in occurrence of insulin resistance. For example, TNF-alpha from adipocytes reduces tyrosine kinase activity of the insulin receptor in obesity. A novel collagen-like protein, adiponectin inhibits TNF-alpha induced cell phenomena and its reduction at obesity may be one of molecular mechanism of insulin resistance. PMID- 10707556 TI - [Insulin resistance and vascular function]. AB - There is a very close interrelationship between the metabolic disorders such as obesity and diabetes mellitus and cardiovascular diseases such as hypertension and atherosclerosis, with insulin resistance and endothelial dysfunction as common features. Insulin has vasculoprotective effects through production of nitric oxide in the endothelial cells, while it produces atherogenic effects by stimulating proliferation and migration of vascular smooth muscle cells(VSMC). The insulin-activated pathway is the phosphatidylinositol 3-kinase pathway in the endothelial cells and MAP kinase pathway in the VSMC. Insulin resistance and hyperinsulinemia may result in the attenuation of the endothelium-mediated action and stimulation of the VSMC-mediated action. Insulin resistance and endothelial dysfunction are related to each other and may cause vicious cycle, leading to the metabolic and cardiovascular diseases. PMID- 10707557 TI - [Insulin resistance and liver]. AB - The liver plays an important role in glucose homeostasis. In the postabsorptive state, plasma glucose is regulated by both hepatic glucose output(HGO) and peripheral glucose utilization(PGU). Insulin and glucagon regulate both HGO and PGU. Increased HGO and decreased PGU, suggesting insulin resistances bring increased fasting blood glucose. While glucose homeostasis after glucose ingestion is regulated by several factors. The regulation of hepatic glucose uptake(HGU) occurs by way of the hormonal milieu(insulin and glucagon), the glucose level, and the rote of glucose delivery. The presence of coordinated changes in insulin, glucagon and the glucose level in combination with the portal signal ensures adequate HGU in response to liver. Hepatic insulin resistance(increased HGO and decreased HGU) and peripheral insulin resistance(decreased PGU) are the characters of glucose intolerance. PMID- 10707558 TI - [Peroxisome proliferator-activated receptor(PPAR)--structure, function, tissue distribution, gene expression]. AB - Peroxisome proliferator-activated receptors(PPARs, alpha, beta/delta, and gamma) are members of the nuclear receptor superfamily of ligand-activated transcription factors. PPAR has been shown to be regulated the expression of genes involved in lipid metabolism by various compounds such as fibrates, thiazolidinediones, prostaglandins, and fatty acids that is important in adipocyte differentiation and glucose homeostasis. PPARs form a heterodimer with the retinoid X receptor(RXR) and bind to specific DNA sequences located upstream of peroxisome proliferator responsive genes and interact with co-activators, including SRC-1 family, CBP/p300 or TRAP/DRIP complex by ligand dependent way. It is suggested that the selective interactions of PPAR with coactivators induced by PPAR's compounds specify the biological activities of the its compounds. Such differential combinations of transcription factors/coactivators are believed to activate only particular sets of target gene promoters. PMID- 10707559 TI - [The development of thiazolidinedione drugs as anti-diabetic agents]. AB - Thiazolidinedione drugs which increase insulin sensitivity are attracting attention of diabetologists. The first drug, ciglitazone ameliorated hyperglycemia in animal models of obese type 2 diabetes such as KKAy mice, but the effect was too weak for clinical application. The first clinical drug troglitazone was approved and marketed in 1997, and the second drug pioglitazone in 1999. Troglitazone was designed to combine tocopherol, anti-peroxidant, and thiazolidinedione. Plasma glucose is lowered in type 2 diabetic patients by troglitazone or pioglitazone alone or in combination with sulfonylureas. The decrease in glycemia is accompanied with the decrease in plasma insulin, suggesting that the effect is mediated by the improvement of insulin sensitivity. Other new thiazolidinedione drugs such as rosiglitazone are in development. Rare but severe hepatic injury occasionally leading to death has been noticed after several months of clinical application of troglitazone. Monthly examination of liver enzymes reduced the number of severe hepatotoxicity. The ultimate evaluation of thiazolidinediones awaits more clinical experiences. PMID- 10707560 TI - [Mechanisms of thiazolidinedione derivatives for hypoglycemic and insulin sensitizing effects]. AB - Ciglitazone was the first insulin sensitizer with a thiazolidinedione structure to reduce insulin resistance and hyperglycemia and many thiazolidinedione derivatives (TZDs) have since been reported as insulin sensitizers. Pioglitazone is reported to lower blood glucose through reductions in both hepatic and peripheral insulin resistance. Regarding the molecular mechanism, pioglitazone was first reported to increase the tyrosine kinase activity of the insulin receptor and then TZDs were reported to increase the insulin signal transduction. TNF-alpha causes insulin resistance through inhibition of the insulin signal transduction and TZDs have been shown to reduce insulin resistance by the suppression of the TNF-alpha gene expression. In addition, TZDs are specific agonists of PPAR gamma and the hypoglycemic effect of TZDs has been shown to involve PPAR gamma among the PPARs. Based on these observations, the molecular mechanisms of TZDs are discussed. PMID- 10707561 TI - [Clinical effect and side effect of troglitazone]. AB - Troglitazone, a PPAR-gamma agonist, is a new drug for type 2 diabetes. The drug decreases blood glucose via enhancing insulin action. Recently Sankyo pharmaceutical company is warning severe hepatotoxicity by troglitazone. It recommends to examine liver function every month in diabetic patients treated with the drug in order early to find drug-induced hepatitis. In Japan 153 diabetic patients treated with the drug developed severe hepatitis and 8 of them died of drug-side effects. Quinone metabolite of troglitazone predominantly in the liver to a sulfate conjugate and activation of PPAR gamma and PXR(pregnane X receptor) by troglitazone are supposed to be factors of hepatotoxic mechanism. PMID- 10707562 TI - [Proper usage of thiazolidinediones]. AB - For the proper usage of thiazolidinedione, 5 preparations including troglitazone, pioglitazone and rosiglitazone in Japan and USA, the package insert of preparation were carefully reviewed from the point of view on indication of pathophysiology of Type 2 Diabetes Mellitus, mono or combined therapy and safety including hepatotoxicity, fluid accumulation. Careful reading of the package insert and patient leaflet, if any, is essential for the proper and safe usage of this important and prospective medicine. True usefulness waits for many long-term multi-center mega-size clinical trials. PMID- 10707563 TI - [Troglitazone]. AB - There are good responders to troglitazone in whom sulfonylurea had failed to improve glycemia. The anti-diabetic effect of this agent seems to be partially composed of prolonged action. The action on different adipose tissue depots of troglitazone may be involved in this slow action. Significant decrease in fasting plasma glucose occurred. The effect may be related to reducing nocturnal hepatic glucose output by the drug. The efficacy of troglitazone is dependent on the levels of circulating insulin. Recent reports demonstrate the good therapeutic power of troglitazone in combination with a sulfonylurea or metformin, or insulin. Thus, we got a new approach of oral therapy to patients with type 2 diabetes in sulfonylurea failure before entering the insulin treatment. Because of low frequency of hypoglycemia, we can safely continue to use this drug in patients with euglycemia by the therapy for preventing return to hyperglycemia. PMID- 10707564 TI - [Clinical evaluation of pioglitazone]. AB - Pioglitazone is the second thiazolidinedione derivative to be clinically used for type 2 diabetes in Japan. It is ten times more potent than troglitazone in glucose-lowering effect. Favourable effects against abnormal lipid levels including decreasing blood triglyceride levels, free fatty acid levels and increasing HDL-cholesterol is advantage to treat obese diabetic patient who may develop atherosclerosis. Up to now, there has been no report of severe hepatic dysfunction due to pioglitazone treatment. Pioglitazone should be carefully monitored in its clinical treatment regarding possible its side effect of hepatic dysfunction. PMID- 10707565 TI - [Rosiglitazone (BRL-49653)]. AB - Rosiglitazone, a thiazolidinedion antidiabetic agent, improves insulin resistance in patients with type 2 diabetes mellitus. Rosiglitazone binds to PPAR-gamma with high affinity and the in vivo antidiabetic potency of rosiglitazone is correlated with its high biding affinity. In animal models of insulin resistence, rosiglitazone decreased plasma glucose, triglyceride and insulin levels and also prevented diabetic nephropathy and pancreatic islet cell degeneration. In clinical trials in patients with type 2 diabetes mellitus, rosiglitazone, 2 to 12 mg/day (as a single daily dose or 2 divided daily doses), improved glycemic control as demonstrated, by decreases in fasting plasma glucose and HbA1C levels. Rosiglitazone did not appear to increase the risk of hypoglycemia and there was no evidence of hepatotoxicity in pre-clinical trials. PMID- 10707566 TI - [Diagnostic criteria of insulin resistance and multiple risk factor syndrome]. AB - Multiple risk factor syndrome (MRFS) is a clustering of cardiovascular risk factors, which describes the epidemiological association of glucose intolerance, central obesity, hypertension, increased triglyceride level and decreased HDL cholesterol, leading to the atherosclerosis. Insulin resistance is diagnosed clinically by fasting hyperinsulinemia, steady state plasma glucose (SSPG) method, insulin tolerance test and glucose clamp study. Visceral fat accumulation is supposed to play a central role in pathogenesis of MRFS and induces risk factors for cardiovascular disease through the increased TNF-alpha expression in adipose tissue and serum FFA level, which cause insulin resistance state. These risk factors should be prevented at early stage by the intervention in obesity, especially visceral fat accumulation. PMID- 10707567 TI - [Obesity, insulin resistance and the implication of thiazolidinediones]. AB - The mechanism of insulin resistance in obesity is not fully understood. In muscle cells, the number of insulin receptor, the function of glucose transporter 4 and the activity of tyrosine kinase decrease. The rink of body fat accumulation and insulin resistance in muscle is thought through free fatty acid and tumor necrosis factor alpha secreted in adipose tissue. Thiazolidinediones (TZDs) are useful to reduce insulin resistance especially in obesity. TZDs seem to cause small weight gain, but to reduce visceral fat in 12-24 weeks. In longer period, it hasn't been studied very much. There are some unsolved problems. So now, targets of TZDs are obese diabetes failed in other medicines. When using TZDs, be cautious of excess eating and physical inactivity. PMID- 10707568 TI - [Insulin sensitizer and hypertension]. AB - Insulin resistance syndrome is the theory that glucose intolerance, hyperinsulinemia, increased very low density lipoprotein triglyceride level, decreased high density lipoprotein cholesterol level, and hypertension are proposed consequences of insulin resistance. In this theory, insulin resistance and the resultant hyperinsulinemia are considered to raise blood pressure through 1) sympathetic nervous system activation, 2) renal sodium retention, 3) renin angiotensin system stimulation, and 4) intracellular calcium accumulation in vascular smooth muscle. Indeed, metabolic disturbance and insulin resistance have been pointed out in essential hypertensives. The effect of insulin sensitizer on blood pressure have been investigated in human and animal experiments. It has been clarified that troglitazone reduces blood pressure in essential hypertensives and fructose fed rats. However, it has also been reported that troglitazone did not lower blood pressure in SHR and fructose fed rats. Further study will be necessary to clarify the effects of insulin sensitizer on blood pressure. PMID- 10707569 TI - [Diabetes mellitus]. AB - The thiazolidinedione compound is known as an insulin-sensitizing agent and is considered a very promising drug, because insulin resistance is a major component of type 2 diabetes. Many reports have demonstrated that the thiazolidinedione has the good hypoglycemic effect in monotherapy and in combination therapy with sulfonylureas, metformin, alpha-glucosidase inhibitors, or insulin. Furthermore, the thiazolidinedione has additional beneficial effects on serum lipids, arterial blood pressure, coagulation-fibrinolysis system, arterial wall cell function. These favorable effects suggest the thiazolidinedione may have inhibitory actions on both macroangiopathy and microangiopathy in patients with diabetes mellitus. However, some patients treated with troglitazone, the first available thiazolidinedione, have suffered from liver dysfunction; in some cases, patients have progressed to hepatic failure and death unfortunately. As the thiazolidinediones are clearly excellent drugs to diabetics with insulin resistance, we should check hepatic function and monitor symptomes of hepatic damage. PMID- 10707570 TI - [Dyslipidemia in insulin resistance and its improvement by troglitazone]. AB - Dyslipideia in insulin resistant state are characterized by 3 major components; increased triglyceride levels, decreased high-density lipoprotein (HDL) cholesterol, and change in the composition of low-density lipoprotein (LDL) cholesterol particle which results in small-dense LDL. Insulin resistance is thought to lead to overproduction of very low-density lipoprotein (VLDL) cholesterol through the decreased peripheral lipoprotein lipase (LPL) activity, increased production of apolipoprotein B-100 and decreased clearance of remnant particles. Troglitazone, an insulin action enhancer, decreases the triglyceride level, increases HDL cholesterol level and decreases th proportion of small-dense LDL through the direct effect on lipoprotein metabolism. However, activation of PPAR-gamma is considered to possese potential atherogeneity and more closer examination is needed. PMID- 10707571 TI - [Insulin sensitizer and urate metabolism]. AB - Gout patients often have various characteristics of insulin resistance (IR) syndrome such as glucose intolerance, hyperlipidemia, hypertension and obesity. In addition, epidemiological data suggest that hyperuricemia is associated with higher rates of death due to cardiovascular and cerebrovascular disorders. However, it has not conclusively been shown whether the association between hyperuricemia and increased death rate is secondary to the association between IR and death or hyperuricemia itself is an independent risk of death. It is of interest to examine the effects of insulin sensitizer which was developed recently on serum urate concentration because it may provide a new idea as to the mechanism of the association between IR, hyperuricemia and vascular disorders. In the present paper, we discuss the relevance of IR to hyperuricemia and gout, and show the data of urate and glucose metabolism obtained from control subjects or the patients with hyperuricemia, gout or type 2 diabetes. PMID- 10707572 TI - [Cardiovascular effects of the thiazolidinedione troglitazone]. AB - Troglitazone, a newly introduced insulin sensitizer, has been implicated in prevention and treatment of atherosclerotic cardiovascular disease especially associated with type 2 diabetes mellitus and insulin resistance. Beneficial effects of troglitazone on multiple risk factor syndrome have been reported in terms of blood pressure lowering effect and ameliorations of dyslipidemia and hyperinsulinemia. Moreover, in vitro and in vivo studies have shown vasodilating and antiatherogenic effects as well as cardioprotective action of this compound. Thus, troglitazone may have potential to prevent and delay diabetic heart disease and large vessel complications. PMID- 10707573 TI - [Kidney disease and insulin resistance--clinical impact of thiazolidinedione compounds for kidney disease]. AB - The thiazolidinedione compounds are well known hypoglycemic agents via increasing insulin-sensitivity. Herein, we provide the possibility that thiazolidinedione compounds could be useful for renal dysfunction through mechanism dependent or independent of its insulin-sensitizing action. In type 2 diabetes, troglitazone could reduce urinary albumin-creatinine ratio compared to metformin. Furthermore, we have shown that troglitazone was able to prevent diabetic glomerular dysfunction through inhibition of diacylglycerol-protein kinase C-extracellular signal-regulated kinase pathway in type 1 diabetic rats. Thus, thiazolidinediones might be effective agents for treating insulin-resistant diabetes as well as diabetes-induced kidney disease. PMID- 10707574 TI - [Chronic liver diseases]. AB - Insulin resistance (reduced insulin action to stimulate glycogen synthesis in muscle and occasionally to inhibit glucose output in liver) is well known to be frequently present in patients with chronic liver diseases, especially cirrhosis. It has been documented that 60% to 80% of patients with cirrhosis are glucose intolerant, and 10% to 30% eventually develop overt diabetes mellitus when their impaired insulin secretion cannot meet the increased demand for insulin due to insulin resistance. Therefore, it seems rational to treat cirrhotic patients with such an insulin sensitizer as thiazolidinediones. However, it must be forbidden to use thiazolidinediones in cirrhotic patients whose liver function capacity is limited until the possible liver toxicity is clarified. PMID- 10707575 TI - [Insulin resistance in pituitary, thyroid, and adrenal diseases]. AB - Patients with acromegaly, Cushing's syndrome, and Graves' disease often have impaired glucose tolerance with high insulin secretion. Persistent elevation of serum GH levels has insulin-antagonistic effects on peripheral tissues. The number of insulin receptors on monocytes is decreased in patients with acromegaly. The expression of GLUT-1 glucose transporters is also decreased in GH treated adipocytes in vitro. Insulin resistance in patients with Graves' disease may be due to antagonism between the effect of insulin and hyperthyroidism at the hepatic level. Glucocorticoids also have insulin-antagonistic effects on both hepatic and extrahepatic tissues, although the precise mechanism still remains to be elucidated. Insulin resistance in these patients can be improved after successful treatment of their endocrine diseases. PMID- 10707576 TI - [Skeletal effects of thiazolidinediones]. AB - Thiazolidinedione (TZD), a new class of anti-diabetic agents, is known to promote adipocytic differentiation by activating peroxisome proliferator-activated receptor-gamma (PPAR gamma). In the bone marrow, osteoblasts and adipocytes are derived from common mesenchymal stem cells or stromal cells, which also play crucial roles in the generation of osteoclasts from their progenitor hematopoietic cells. Recent in vitro studies demonstrated that TZDs promote adipocytic differention of the stromal cells. However, whether or not this affects osteoblastic differentiation is unclear. On the other hand, TZDs clearly inhibit osteoclast-like cell formation and bone resorption in vitro. These results indicate that TZDs have direct effects on bone cells. However, little is known about their in vivo effects on bone. Our recent study demonstrated that troglitazone, a TZD, decreased bone resorption markers before it affected glycemic indices in type2 diabetics, suggesting TZDs affects bone in vivo and may be beneficial for bone health in type2 diabetics. PMID- 10707577 TI - [PPARs, cancer cells and inflammation]. AB - The peroxisome proliferator-activated receptor-gamma (PPAR gamma) is a ligand dependent nuclear receptor that has been implicated in the modulation of aspects of development and homeostasis, including adipocyte differentiation and glucose metabolism. Recent reports indicate that PPAR gamma is also expressed at significant level in several other tissues, including macrophages, monocytes and aortic smooth muscle and cancer cells. In this point of view, PPARs are susceptible to mediated the modulation of immune cell activation and tumor cell differentiation. If a major role for PPARs in tumor cell differentiation is manifested, PPARs may also provide a link between nutrition and certain types of cancers. PMID- 10707578 TI - [Troglitazone for treatment of polycystic ovary syndrome]. AB - Polycystic ovary syndrome(PCOS) is characterized by clinical symptoms such as menstrual dysfunction, unovulatory infertility, masculinization, obesity, polycystic ovary by ultrasound, and endocrine abnormalities such as hyperandrogenism, and elevated LH to FSH ratio. Recent reports suggest that insulin resistance plays an important role in the pathogenesis of PCOS, and several insulin sensitizing agents have been used for the treatment of PCOS. Troglitazone, one of the thiazolidinediones, improves not only insulin sensitivity but also hyperandrogenism and ovulatory function. Troglitazone appears to be useful in treating women with PCOS. Further investigations are needed to assess the effectiveness and safety. PMID- 10707579 TI - [Insulin resistance induced by drugs or agents]. AB - Glucocorticoids, oral contraceptives and nicotinic acid are associated with glucose intolerance, and the main cause of the glucose intolerance induced by these drugs is insulin resistance. Particularly glucocorticoids are difficult to substitute for another drugs, and frequency cause glucose intolerance. Now thiazolidinediones is available for reducing insulin resistance in clinical. And there are several evidences that thiazolidinediones can reduce insulin resistance induced by glucocorticoids in rats. But the clinical evidences are little. In future, it is necessary to investigate whether thiazolidinediones would be effective on reducing glucocorticoid-induced insulin resistance in clinical. PMID- 10707580 TI - [Genetic diagnosis and cardiovascular diseases]. AB - Many of cardiovascular diseases are multifactorial and both environmental and genetic factors are associated with their onset and/or progression. Recently, the development of genetic analysis has enabled us to understand genetic backgrounds regulating the susceptibility to the specific environmental factors. These progress may bring us new strategies for diagnosis and/or prevention of diseases. In this review, we mainly discuss the current knowledge of these genetic analysis and perspectives in cardiovascular diseases. We also present briefly genetic mutations in cardiomyopathy, long QT syndrome, congenital cardiovascular diseases and so on. Additionally, we describe considerable problems about the interpretation and notification of the results of genetic diagnosis. PMID- 10707581 TI - [Regulation of ER(estrogen receptor) gene expression in the genesis and development of breast cancer]. AB - We studied the regulation of ER alpha gene expression in terms of transcriptional regulation and DNA-methylation in human breast cancer. In cancer, levels of ER alpha mRNA transcribed from a distal promoter (promoter B) remarkably correlated to the protein levels. The binding protein(ERBF-1) of an important cis-element, which we identified, was exclusively expressed in the cells expressing ER alpha, indicating its important role in the enhanced expression of ER alpha in breast cancer. Furthermore, loss of ER alpha expression is the most important event in development of breast cancer. We found that methylation of two promoters of ER alpha was inversely associated with the gene expression in human breast cancer. Specifically, methylation of promoter B suppressed ER alpha expression, and demethylating treatment recovered it. Both methylation of ER alpha gene and modification of transcription by certain trans-acting factors are important for the regulation of ER alpha expression during breast carcinogenesis. PMID- 10707582 TI - [Medical treatment of Crohn's disease]. PMID- 10707583 TI - [Current status of surgical treatment for pancreatic cancer]. PMID- 10707584 TI - [Measurement of fecal proteins in inflammatory bowel disease--usefulness as an activity index]. AB - Fecal alpha 1-antitrypsin (alpha 1-AT), alpha 2-macroglobulin (alpha 2-M), lysozyme (Lz), and lactoferrin (Lf) levels were measured in 73 samples from 32 patients with ulcerative colitis (UC), 52 samples from 21 patients with Crohn's disease (CD), and 41 samples from 21 healthy volunteers. According to three degree of bowel activity, the UC patients were divided into 4 groups and the CD patients were divided into 2 groups. Fecal alpha 1-AT levels were measured by latex agglutination and the other protein parameters by ELISA. All protein levels, except alpha 1-AT, rose as the degree of activity increased. The fecal protein markers alpha 2-M, LZ, and Lf had significantly higher positive rates than the serum inflammatory markers and activity index in the moderate and severe UC groups, and alpha 2-M and Lf had significantly higher rates in the CD (+) group. Based on these findings measurement of fecal levels on alpha 2-M, LZ, and Lf appeared to be useful activity markers for UC, and alpha 2-M and Lf for CD. PMID- 10707585 TI - [A case of esophageal and duodenal varices treated with Hassab's procedure and ligation of the duodenal varix]. PMID- 10707586 TI - [Recurrent gastric cancer with hypercalcemia that represented elevated serum level of parathyroid hormone related protein (PTHrP) and rapid clinical course]. PMID- 10707587 TI - [A case of perforated diverticulum of the transverse colon]. PMID- 10707588 TI - [A case of malignant fibrous histiocytoma of mesentery of the ileum]. PMID- 10707589 TI - [A case of ischemic enteritis showing a tubular stenosis]. PMID- 10707590 TI - [A case of rectal lues]. PMID- 10707591 TI - [A case of chronic idiopathic intestinal pseudo-obstruction and pneumatosis cystoides intestinalis with pneumatoperitoneum, improved by the hyperbaric oxygen therapy]. PMID- 10707592 TI - [A case of signet ring cell carcinoma of the gallbladder with anomalous pancreaticobiliary ductal union]. PMID- 10707593 TI - [A huge polypoid early gallbladder carcinoma with wide mucosal spreading]. PMID- 10707594 TI - [Spontaneous internal drainage of pancreatic pseudocyst with fistula to the common bile duct]. PMID- 10707595 TI - [Surgery in the elderly]. PMID- 10707596 TI - [Celiac disease in adults]. AB - Studies from western countries have shown that coeliac disease (CD) is common with prevalence figures about 1:300. The clinical spectrum varies greatly, steatorrhoea and weight loss affecting less than half of the patients. CD should be suspected in case of positive gliadin (IgA and IgG) and endomysial (IgA) antibodies. The diagnosis is based upon histological examination of duodenal biopsies taken during upper gastrointestinal endoscopy. Most patients respond quickly and satisfactorily to treatment with a gluten-free diet. This treatment also eliminates the excess risk of small bowel malignancy. Screening among first degree relatives and patients with insulin-dependent diabetes mellitus should be considered. PMID- 10707597 TI - [Communication between general practitioners and hospitals]. AB - The study was partly based on a retrospective analysis of 408 hospital referrals and 261 discharge summary letters and partly on interviews with chief physicians/surgeons and general practitioners. The level of information in hospital referrals: patient history 87%, objective findings 94%, social medicine 31%, plan/expectations 21%. The diagnostic applicability of patient history/objective findings was 95% and social medicine 70%. The discharge summary letter was received 2-3 days after hospital discharge in 17% cases. In the discharge letters information about medication was described in 41%, information given to patient/relatives in 9%. When discharge summary letters from departments of internal medicine and surgery were compared the level of informations from departments of internal medicine to departments of surgery was superior e.g. descriptions of medication (62% against 26%), date of control (34% against 24%) and information to patient/relatives (12% against 5%). The conclusion was that the level of information and the diagnostic applicability showed variation with regard to quality. General practitioners and hospitals must develop guidelines for hospital referrals and discharge summary letters in order to improve the patient's course. PMID- 10707598 TI - [How do patients evaluate their general practitioners? Danish results from a European study]. AB - This study aims to assess how patients evaluate the quality of general practice care. With a questionnaire answered by a sample of 1309 (90.9% response) from 36 practices. Patients answered 25 questions relating to their general practice care. Patients were, generally seen, very positive about their general practitioners. "Keeping your records and data confidential" and "providing quick services for urgent health problems" received most positive evaluations, while "waiting time in the waiting room", "being able to speak to the general practitioner on the telephone" and "getting through to the practice on the phone" were assessed most negatively. PMID- 10707599 TI - [Parent groups. Crisis intervention for parents of extremely premature infants during hospitalization]. AB - The aim of the study was to evaluate the benefit from participation in parent groups for parents of extremely premature new-born babies. The participation in the parent groups led by the psychologist took place during the hospitalization of the child on the neonatal ward. All parents of extremely premature new-borns admitted to the neonatal department, Hvidovre Hospital, Copenhagen, Denmark in the period from 1 January 1992 to 30 June 1994, were asked to fill in questionnaires. The study population comprised the parents of 58 children. The parents of 14 children did not want to participate. Of the remaining 44 children, 36 were alive and eight had died at the time of the study. Most parents participated in the parent group and the majority stated they had benefited from the participation. It is therefore considered a relevant psychological task to establish parent groups for parents of extremely premature new-borns as a mean of crisis intervention in the neonatal department. PMID- 10707600 TI - [The young clientele of a sexual disorders clinic]. AB - This study is based on the charts of all the adolescents (age 11-22 incl., total number 234) treated in a Danish sexological clinic 1986-1995. They account for rather a small part of the total sexological clientele. Among the youngest there was a majority of boys, some with congenital, genital deformities. From the age of 17 the number of patients increased abruptly, now with a majority of women. In the age group of 17-20 year-old there were many adolescents with sexual identity disorder, in the oldest age groups many women with sexual dysfunctions. For half of the adolescents the duration of therapy was 1-12 months. The study illustrates the distinctive features of the young group including the culmination of conflicts of sexual identity in mid-adolescence. PMID- 10707601 TI - [Atypical endometrial hyperplasia. Prognosis and course]. AB - The treatment of patients with the diagnosis atypical endometrial hyperplasia has been disputed during the last decades. The aim of the study was to evaluate the treatment of these patients and analyse the progression rate to invasive carcinoma of the endometrium. Fifty-seven patients with atypical hyperplasia were examined and treated from 1976 through 1991. The medical records were examined retrospectively and the pathology slides were revised by one pathologist in accordance with the 1975 WHO recommendations. Thirty-one (54%) patients were on oestrogen treatment as monotherapy at the time of diagnosis. Forty-two patients had a hysterectomy performed within five months, and five patients had a hysterectomy performed 10 to 61 months after diagnosis. A total of 18 out of 57 patients (31.6%) had or developed endometrial carcinoma all with myometrial invasion: 14 stage I with < or = 50% myometrial invasion, three stage I with > 50% myometrial invasion, and one stage IV. There was no significant difference in age, body mass index, parity or hormone replacement treatment between the group with endometrial carcinoma and the group without endometrial carcinoma. We conclude that unopposed oestrogen treatment and nulliparity are the main risk factors for atypical hyperplasia and that hysterectomy is the appropriate treatment for patients with atypical hyperplasia of the endometrium. PMID- 10707602 TI - [Overlooked celiac disease in a 59-year old woman with severe osteomalacia]. AB - Coeliac disease in adults is an overlooked and undiagnosed illness. We present a case where a woman was diagnosed at the age of 59 despite symptoms suggestive of coeliac disease since childhood. At the time of diagnosis the patient had severe osteomalacia with multiple vertebral fractures of the thoracic and lumbar spine. PMID- 10707603 TI - [Rett syndrome: congenital defect of a transcription inhibitory protein]. PMID- 10707604 TI - [Hearing tests in specialists' practice one more time]. PMID- 10707605 TI - [Obesity as a risk factor of ischemic heart disease]. PMID- 10707606 TI - Evaluation of cryopreserved murine and human hematopoietic stem and progenitor cells designated for transplantation. AB - The influence of five different cryopreservation protocols on the quality and/or quantity of frozen cells was investigated on mouse bone marrow cells and human peripheral blood mononuclear cells (MNC). The efficiency of the protocols was evaluated on the basis of the recovery of very primitive pluripotent hematopoietic stem cells (MRA), pluripotent progenitors (CFU-Sd12), committed granulocyte-monocyte progenitors (CFU-GM) of mouse cells after thawing. The recovery of MRA, CFU-Sd12 and CFU-GM varied depending on the type of freezing procedure and cryoprotector (DMSO) concentrations used. It was shown that the controlled-rate protocol was more efficient, enabling better recovery of all categories of progenitor cells in frozen samples. The most efficient was the controlled-rate protocol of the cryopreservation designed to compensate for the release of fusion heat, which enabled the best recovery of CFU-GM (73.0 +/- 8.8%) and CFU-Sd12 (90.0 +/- 15.9%) when combined with 5% DMSO concentration (protocol 4). On the contrary, a better recovery (79.8 +/- 13.5%) of very primitive stem cells (MRA) was achieved only when the higher concentration (10%) DMSO was used in combination with a five-step protocol of cryopreservation (protocol 1). These results pointed out the adequately used controlled-rate freezing to be essential for a highly efficient cryopreservation of some of the categories of hematopoietic stem and progenitor cells. At the same time, it was obvious that a higher DMSO concentration was necessary for the cryopreservation of MRA, but not for more mature progenitor cells (CFU-S, CFU-GM). These results imply the existence of a mechanism that decreases the intracellular concentration of DMSO in MRA cells, which is not the case in less primitive progenitors. For human MNC, the recovery and viability of the cells, as well as the engraftment potential of cryopreserved cells after thawing were investigated. Cryopreservation protocol 1 resulted in better MNC recovery (82.7 +/- 10.4%) than protocol 3 (49.9 +/- 15.1%). The mean recovery of MNCs (collected from patients for autologous transplantation) was 78.5 +/- 7.3% (protocol 1) and 53.1 +/- 26.2% (protocol 3). The obtained favorable recovery of thawed cells and rapid reconstitution of hematopoiesis (on the day 11th following the transplantation) in patients confirmed that the controlled-rate freezing in combination with optimal DMSO concentration was able to obtain sufficient progenitor cryoprotection. PMID- 10707607 TI - Professional exposure of drivers to carbon monoxide as a possible risk factor for the occurrence of traffic accidents in the road traffic. AB - The aim of this study was to examine the effects of increased carbon monoxide concentrations in the air of car cabin on the health status of exposed drivers and the occurrence of traffic accidents. Exposed group was comprised of 250 drivers professionally exposed to increased carbon monoxide concentrations in the car cabins. Control group was comprised of 120 professional drivers who were not exposed to increased concentrations of carbon monoxide in the air of the car cabins. Average carbon monoxide concentration in the air of the car cabins of the drivers from exposed group was 71.2 +/- 8.1 ppm, which was significantly higher compared to the controls (5.4 +/- 1.2 ppm). The drivers from the exposed group more frequently suffered from headaches, irritability, vertigo and palpitation than the drivers from the control group. The prevalence of diabetes mellitus, arrhythmia, atherosclerotic peripheral artery diseases and coronary heart diseases was more frequent in the exposed than in the control group. Drivers from the exposed had longer reaction time on acoustic and visual stimulation compared to the control group. The examined drivers statistically caused more traffic accidents than the drivers from the control group. PMID- 10707608 TI - [Role of nimodipine in the therapy of subarachnoid and intracerebral hemorrhage]. AB - Syndromes of intracranial hemorrhage, and particularly subarachnoidal, i.e., intracerebral hemorrhage (SAH and IH) present clinical entities that are the most severe conditions in neurology. Timely recognition, diagnosis and adequate therapy are imperative. The most important factor that aggravates an already difficult prognosis of those entities is cerebral vasospasm. Upon the presented facts, the aim of this investigation was to establish the value and role of administration of selective calcium channel blocker--nimodipine in patients with SAH and IH compared to the degree of neurological and functional impairment, as well as the recovery of the function of consciousness compared to the patients with those syndromes from an earlier period, who were not treated with this medicament. Investigation comprised 30 patients who received nimodipine and 20 patients without this agent in therapeutic program. Results of the investigation confirmed significant difference concerning the neurological recovery, improvement of functional capability and recovery or consciousness disturbances, respectively, in patients who received nimodipine compared to the group without this agent. It can be concluded that nimodipine as calcium channel blocker with multitopic pharmacological effects on mechanism of SAH or IH development, respectively, as well as on the development of complications of those syndromes, particularly to the development of vasospasm and reactive ischemia, with the improvement of hemorrheologic disorders deserves to be included as the unavoidable segment of therapeutic program of SAH and IH syndrome immediately after clinical phenomenology is revealed. PMID- 10707609 TI - [Use of Bernard's diadynamic current in evaluating recovery of pain sensitivity in transplanted microvascular flaps]. AB - The application of microvascular transfer of tissues provides a single act transplantation of free flaps to any part of the body. Following the operation, free flaps are nerveless. By spreading out of nerves from the recipient region surrounding into the flap tissue, transplanted free flaps become reinervated. The sensitivity of the transferred free flaps is very important in feet, palms, breasts, genitals, face etc. The aim of our research was to use Bernard's diadynamic currents in establishing the time needed for recovery of the transferred flap's pain sensitivity in comparison with the recipient region's surrounding. At the Clinic for Plastic Surgery and Burns and Clinic for Physical Medicine and Rehabilitation of the Military Medical Academy, we used Bernard's diadynamic current to analyze the recovery of the pain sensitivity in 33 patients with transferred free flaps. The first tests were performed six moths after the free flap transfer and were repeated in the intervals of 6 to 36 months. The pain recovery was tested in 5 cutaneous, 18 myocutaneous and 10 osteocutaneous flaps transplanted to foot-10, lower leg-13 and head-10. Pain sensitivity in the transferred flaps was detected 6 months after the free flaps transfer and was definitely ascertained 12 months after the operation. Current of 10.11 mA was required to reach the pain sensitivity threshold after the first six months and 8.52 mA after 36 months. It is obvious that the pain sensitivity threshold is significantly higher in the transferred free flaps (p < 0.001) than in the donor or recipient regions' surroundings. PMID- 10707610 TI - [Significance of determination of certain clinical and laboratory parameters in the evaluation of severity and outcome in sepsis]. AB - Sepsis and its complications are severe clinical syndrome that is caused by systemic inflammatory response of the host to infection. Despite the use of common and numerous new therapeutic protocols, mortality from this severe disease is still very high. In the study are presented 155 patients (111 males, 44 females) of average age 49.6 years with mean septic score 12.9 (2-40). Mortality in our patients was 20.6%, septic shock developed in 31.6%, ARF in 20.0%, DIC in 12.9%, and MODS in 25.8% of patients. Positive correlation existed between initial sepsis score and mortality. Older age and the presence of primary diseases (34.2% of patients) were associated with significantly higher septic score and were good prognostic factor for the poor outcome of sepsis. Between mean arterial pressure in the first 24 h after the admission and mortality existed negative correlation (p < 0.05). Positive hemocultures were found in 69.7%, and bacterial infection in 78.7% of patients. GP bacteremia was found in 55.6% of patients and GN in 45.4% of all positive hemocultures. Confirmed bacteremia and bacteremia caused by GPB were associated with the higher mortality rate compared to the patients with negative hemocultures and GN bacteremia (p < 0.05). Concentrations of fibrinogen and urea in the blood at the admission in the patients with sepsis were very good prognostic factors of the disease outcome, and leukopenia, leukocytosis and neutropenia were associated with the increased mortality. Negative correlation existed between fibrinogen concentration and mortality (p < 0.001), while positive correlation (p < 0.001) existed between urea concentration and mortality. In the absence of more efficacious therapeutic protocols, fast recognition of the sepsis, evaluation of its severity, knowledge of the risk factors for its poor outcome and aggressive use of antibiotic and existing supportive therapy can significantly decrease high mortality of this too severe clinical syndrome. PMID- 10707611 TI - [Serum levels of beta 2-microglobulin in various types of hemodialysis]. AB - beta 2-microglobulin (beta 2 m) is the major constituent of amyloid fibrils in dialysis-related amyloidosis (DRA), which is considered to be one of the most severe adverse effect of long-term dialysis. In this study we evaluated the efficiency of beta 2 m removal during different dialysis procedures. A total of 45 patients undergoing hemodialysis were divided in five groups: cuprophane dialysis (n = 10), high-flux polysulphone dialysis (n = 10), postdilutional hemodiafiltration (n = 10), conventional postdilutional hemofiltration (n = 10) and predilutional on-line hemofiltration (n = 5). Serum level of beta 2 m was determined before and after different procedures using ELISA. In the group of patients on cuprophane dialysis was registered an elevation of beta 2 m and of 16.8 +/- 11.4% on the average. Serum level of beta 2 m was decreased following all other procedures on the average of 40.7 +/- 16.4% after high-flux polysulphone dialysis, 42.0 +/- 13.7% after conventional hemofiltration, 64.7 +/- 9% after hemodiafiltration and 67.9 +/- 10.1% after predilutional hemofiltration. The best removal of serum beta 2 m was realized by predilutional hemofiltration. Also, we have noticed that patients treated with high-flux synthetic membranes in the longer time-period have lower predyalisis value of beta 2 m compared to patients treated with cuprophane membrane. Further long-term studies will be necessary to conclude whether these procedures could be successful prophylactic and/or therapeutic regimen for dialysis-related amyloidosis. PMID- 10707612 TI - [Surveillance of communicable diseases using a computer database of reported cases]. AB - Epidemiology services during the surveillance of communicable diseases collects of different sorts of data, which are used for an analysis of epidemiologic situation. Those data are the starting point for timeline control and preventive activities. Data processing of notified communicable diseases cases provides information on types of diseases, number of cases, time and place of their occurrence. Manual data processing, used till 1993, was slow, unreliable and considerably decreased the efficiency of epidemiology service activities. In this paper we have set the hypothesis that is possible to form a computerized database with the following aims: to form user friendly computerized database model for those without knowledge in using computers: to get output spread sheets with information needed for epidemiologic situation analyses at any time. Database was developed in 1993 and has been used as source of the information in epidemiologic diagnosis process. The significant accuracy, reliability, timelines, and shortening of the time of data processing was achieved. The database can also serve as the initial component for designing an epidemiologic services information network in Belgrade county. In designing such a network it is necessary to form the additional databases of isolated infectious agents and their drug resistance, database of health status of persons under surveillance and database of environmental and sanitary condition in children and youth facilities. PMID- 10707613 TI - [Are corticosteroids being used again in the treatment of patients with sepsis and septic shock?]. PMID- 10707614 TI - [Adhesion molecules and lymphocyte circulation in inflammatory diseases of the liver]. PMID- 10707615 TI - [Importance of echocardiography in the diagnosis of causes of ischemic brain disease]. PMID- 10707616 TI - [Transplantation of the small intestine]. PMID- 10707617 TI - [Stress--an etiologic factor in the development of anxiety and depressive disorders]. PMID- 10707619 TI - [Diagnostic dilemmas in surgical pathology]. AB - In the majority of presented cases persisted certain diagnostic perplexities even after the careful investigations were performed, which required surgical exploration. For example, carcinomatous pericarditis as the sequel of metastasis from occult thyroid carcinoma and two cases of double malignity (papillary carcinoma and Hodgkin's lymphoma of the thyroid), retroperitoneal leiomyosarcoma and the clear cells renal carcinoma. All the cases were not successfully solved until the application of highly sophisticated and precise methods of immunohistochemical staining, which without a mistake confirmed our assumed histopathologic diagnosis. Metastatic lesion of folicular thyroid carcinoma in the adrenal gland should be mentioned because of the long remission period of 15 years following total thyroidectomy and neck irradiation during the primary therapy of thyroid carcinoma. It is very significant to point out that pathologic disorders such as undifferentiated neoplasm, strange inflammatory conditions, endocrine proliferative lesions deserve special attention because in certain cases consultative approach is necessary for obtaining the definite finding that can in some degree histoprognostically predict the outcome of pathologic process. PMID- 10707618 TI - [Contusion injury of the brachial artery]. AB - We present a case of a successful repair of brachial artery after blunt trauma, almost five days after the injury. Autovenous graft (great saphenous vein) was used for the repair. The diagnosis was confirmed by angiography, and this method was indisposable in the postoperative course. Precise surgical technique is compulsory in obtaining optimal surgical revascularization. PMID- 10707621 TI - [Development and differentiation of the nervous system: overview] [In Process Citation] PMID- 10707620 TI - [Russian physician emigrants in the Yugoslav military medical service between the two wars (1920-1940)]. PMID- 10707622 TI - [Notch pathway in neural development]. PMID- 10707623 TI - [Regulation of neural development mediated by neural RNA binding proteins]. PMID- 10707624 TI - [Islet-1 family and neural differentiation: study using zebrafish embryos]. PMID- 10707625 TI - [Molecular mechanisms of neuronal alignment in the cerebral cortex and cerebellar cortex]. PMID- 10707626 TI - [Development expression of voltage-gated potassium channels in mouse cerebellar granule cells]. PMID- 10707627 TI - [Actin cytoskeleton in dendrite formation of CNS neuron]. PMID- 10707629 TI - [Pathfinding and target recognition by growth cones: overview] [In Process Citation] PMID- 10707628 TI - [Postnatal development of the cerebellar circuit and gene expression]. PMID- 10707631 TI - [Molecular mechanisms of the pathway formation in the fetal rat cerebral neocortex]. PMID- 10707630 TI - [Mechanism of formation of crossed projection in the brain]. PMID- 10707632 TI - [Genetic analysis of neuromuscular specificity in Drosophila melanogaster]. PMID- 10707633 TI - [Mechanism for formation and maintenance of synapses by biogenic amines]. PMID- 10707634 TI - [The cadherin gene family that regulate neural circuits formation and higher order brain functions]. PMID- 10707636 TI - [Morphogenesis of the optic tectum and retinotectal map formation]. PMID- 10707638 TI - [Brain function and gene expression: overview] [In Process Citation] PMID- 10707635 TI - [Molecular mechanisms underlying the formation of the topographic retinotectal map]. PMID- 10707637 TI - [Molecular mechanisms that regulate neuron network formation in PNS]. PMID- 10707639 TI - [Gene regulation and synapse-specific expression of macromolecules in neuronal plasticity]. PMID- 10707640 TI - [Molecular bases for induction of maternal behavior and stress tolerance by prolactin]. PMID- 10707641 TI - [Drug dependence and gene expression]. PMID- 10707642 TI - [Gene expression of glutamic acid decarboxylase during neural development and activity]. PMID- 10707643 TI - [Associate learning and mechanism of synaptic reinforcement: overview: relationship between associative learning and LTP]. PMID- 10707644 TI - [Activity dependent signaling in neuron]. PMID- 10707645 TI - [Odor-taste associative learning in land slug]. PMID- 10707646 TI - [Application of organotypic cultures to researches on functional synapses]. PMID- 10707647 TI - [Synaptogenesis of the isolated brain in Hermissenda]. PMID- 10707648 TI - [Synapse formation and the regulation of synaptic functions]. PMID- 10707649 TI - [Plasticity in the transmitter release: overview: recent advance of exocitosis research]. PMID- 10707650 TI - [Studies of synaptic plasticity using cultured neuronal cells]. PMID- 10707651 TI - [Functional plasticity of the presynaptic terminal]. PMID- 10707653 TI - [Role of proline-directed protein kinases in synapse formation and in neurotransmitter release]. PMID- 10707652 TI - [Molecular processes of the conversion from the growth cone to the presynaptic terminal]. PMID- 10707654 TI - [Regulatory mechanism of exocytosis at the central synapse]. PMID- 10707655 TI - [Synaphins/complexins, cytosolic proteins associated for neurotransmitter release]. PMID- 10707656 TI - [Intrinsic glutaminergic systems and its negative regulation of melatonin synthesis in mammalian pineal glands]. PMID- 10707657 TI - [Functional significance and mechanism of long-term potentiation: overview]. PMID- 10707658 TI - [LTP as a cellular basis for learning and memory]. PMID- 10707659 TI - [Regulation of synaptic efficacy by neural activity in the hippocampus]. PMID- 10707660 TI - [Plasticity of developing visual cortex and neurotrophins]. PMID- 10707661 TI - [Induction mechanism of long-term potentiation at visual cortical inhibitory synapses]. PMID- 10707662 TI - [Intracellular Ca2+ release and synaptic plasticity]. PMID- 10707663 TI - [Regulation of neural network by changes in receptors: overview]. PMID- 10707664 TI - [Changes in glutamate receptors after brain ischemia]. PMID- 10707665 TI - [Molecular diversity of AMPA-type glutamate receptor channels and its functional implication]. PMID- 10707666 TI - [Molecular organization and function of postsynaptic density (PSD)]. PMID- 10707667 TI - [Changes in apoptosis-regulating proteins in Alzheimer's disease: comparison with development and aging]. PMID- 10707668 TI - [Neuronal tracing method with a recombinant adenovirus]. PMID- 10707669 TI - American Roentgen Ray Society 100th annual meeting. Washington, DC, USA. May 7 12, 2000. Abstracts. PMID- 10707670 TI - 2000 Annual meeting of the American Society for Clinical Pharmacology and Therapeutics, Los Angeles, California, USA. March 14-18, 2000. PMID- 10707671 TI - American National Standard--Specifications for the bottle manikin absorption phantom. PMID- 10707672 TI - Brucella melitensis infection discovered in cattle for first time, goats also infected. PMID- 10707673 TI - Sit up, roll over, and heal. PMID- 10707674 TI - Florida not tickled by threat of infestation. PMID- 10707675 TI - Favors early spay/neuter. PMID- 10707676 TI - Feels in-house laboratory testing is not enough. PMID- 10707677 TI - The underlying interrelated issues of biosecurity. PMID- 10707678 TI - What is your diagnosis? Retropharyngeal soft tissue mass in a ferret. PMID- 10707679 TI - ECG of the month. Ventricular preexcitation in a cat. PMID- 10707680 TI - Use of peripheral parenteral nutritional support in dogs and cats. PMID- 10707681 TI - Horse sales and the veterinarian. PMID- 10707682 TI - Analysis of the 1998 outbreak of leptospirosis in Missouri in humans exposed to infected swine. AB - OBJECTIVE: To determine the extent of leptospirosis in persons exposed to infected swine, confirm the source of disease, define risk factors for infection, and identify means for preventing additional infections during an outbreak in Missouri in 1998. DESIGN: Cross-sectional study. SAMPLE POPULATION: 240 people and 1,700 pigs. PROCEDURE: An epidemiologic investigation was conducted of people exposed to infected pigs from the University of Missouri-Columbia swine herd. The investigation included review of health of the pigs, a cross-sectional study of the people handling the pigs, serologic testing of human and porcine sera, and risk-factor analysis for leptospirosis within the human population. RESULTS: Serologic testing of samples collected at the time of the investigation indicated that 59% of the pigs had titers to leptospires, denoting exposure. Of the 240 people in the exposed study population, 163 (68%) were interviewed, and of these, 110 (67%) submitted a blood sample. Nine (8%) cases of leptospirosis were confirmed by serologic testing. Risk factors associated with leptospirosis included smoking (odds ratio [OR], 14.4; 95% confidence interval [CI], 1.39 to 137.74) and drinking beverages (OR, 5.1; 95% CI, 1.04 to 24.30) while working with infected pigs. Washing hands after work was protective (OR, 0.2; 95% CI, 0.03 to 0.81). CONCLUSIONS AND CLINICAL RELEVANCE: Leptospirosis is a risk for swine producers and slaughterhouse workers, and may be prevented through appropriate hygiene, sanitation, and animal husbandry. It is essential to educate people working with animals or animal tissues about measures for reducing the risk of exposure to zoonotic pathogens. PMID- 10707683 TI - Prevalence of enteric zoonotic organisms in cats. AB - OBJECTIVE: To determine prevalence of enteric zoonotic organisms in cats in north central Colorado. DESIGN: Prospective study. SAMPLE POPULATION: Serum and fecal samples from 87 cats with diarrhea, 106 cats without diarrhea, and 12 cats for which fecal consistency was unknown. PROCEDURES: Samples were obtained from client-owned cats and cats at a humane society shelter. Serum was assayed for feline leukemia virus antigen and antibodies against feline immunodeficiency virus, IgM antibodies against Toxoplasma gondii, and IgG antibodies against T gondii and Cryptosporidium parvum. Microscopic examination of unstained feces was performed after centrifugation in a zinc sulfate solution, thin fecal smears were stained with acid fast stain and examined for C parvum, and bacteriologic culture of feces was used to detect aerobic and anaerobic bacteria. RESULTS: Enteric zoonotic organisms were detected in feces from 27 of 206 (13.1%) cats and included C parvum (5.4%), Giardia spp (2.4%). Toxocara cati (3.9%), Salmonella enterica serotype Typhimurium (1.0%), and Campylobacter jejuni (1.0%); each organism was detected in samples from cats with and without diarrhea. Although differences between groups were not significant, a higher proportion of shelter cats (18.2%) had enteric zoonotic organisms than client-owned cats (10.1%). CONCLUSIONS AND CLINICAL RELEVANCE: Enteric zoonotic organisms were detected in feces of 13.1% of cats, suggesting that cats, particularly those in homes of immunocompromised humans, should be evaluated for enteric zoonotic organisms. PMID- 10707684 TI - Performance of serologic tests used to detect heartworm infection in cats. AB - OBJECTIVE: To compare heartworm serum antibody (Ab) and antigen (Ag) test results, using commercial laboratories and in-house heartworm test kits, with necropsy findings in a population of shelter cats. DESIGN: Prospective study. ANIMALS: 330 cats at an animal shelter. PROCEDURE: Between March and June 1998, 30 ml of blood was collected from the cranial and caudal venae cavae of 330 cats that were euthanatized at a local animal shelter. Results of heartworm Ab and Ag serologic tests for heartworm infection were compared with necropsy findings in this population of cats, using commercial laboratories and in-house test kits to measure serum Ab and Ag concentrations. RESULTS: On necropsy, adult Dirofilaria immitis were found in 19 of 330 (5.8%) cats. Combining results from serum Ab and Ag tests achieved higher sensitivities than using serum Ab and Ag test results alone (i.e., maximum sensitivities of 100% vs 89.5%, respectively, whereas use of serum Ag and Ab test results alone achieved higher specificities compared with the use of a combination of serum Ab and Ag results (i.e., maximum specificities of 99.4% vs 92.9%, respectively). CONCLUSIONS AND CLINICAL RELEVANCE: On the basis of our findings, if a cat has clinical signs that suggest heartworm disease despite a negative heartworm serum Ab test result, an alternative heartworm Ab test, a heartworm Ag test, thoracic radiography, or two-dimensional echocardiography should be performed. PMID- 10707685 TI - Influence of tumor cell proliferation and sex-hormone receptors on effectiveness of radiation therapy for dogs with incompletely resected meningiomas. AB - OBJECTIVE: To assess the influence of tumor cell proliferation and sex-hormone receptors on the efficacy of megavoltage irradiation for dogs with incompletely resected meningiomas. DESIGN: Longitudinal clinical trial. ANIMALS: 20 dogs with incompletely resected intracranial meningiomas. PROCEDURE: Dogs were treated with 48 Gy of radiation administered 3 times per week on an alternate-day schedule of 4 Gy/fraction for 4 weeks, using bilateral parallel-opposed fields. RESULTS: Tumor proliferative fraction measured by immunohistochemical detection of proliferating cell nuclear antigen (PFPCNA index) ranged from 10 to 42% (median, 24%). Progesterone receptor immunoreactivity was detected in 70% of tumors. Estrogen receptor immunoreactivity was not detected. An inverse correlation was found between detection of progesterone receptors and the PFPCNA index. The overall 2-year progression-free survival (PFS) rate was 68%. The only prognostic factor that significantly affected PFS rate was the PFPCNA index. The 2-year PFS was 42% for tumors with a high PFPCNA index (value > or = 24%) and 91% for tumors with a low PFPCNA index (value < 24%). Tumors with a high PFPCNA index were 9.1 times as likely to recur as were tumors with a low PFPCNA index. CONCLUSIONS AND CLINICAL RELEVANCE: This study confirms the value of irradiation for dogs with incompletely resected meningiomas. Prognostic value of the PFPCNA index suggests that duration of treatment and interval from surgery to start of irradiation may affect outcome. Loss of progesterone receptors in some tumors may be responsible for an increase in PFPCNA index and may indirectly affect prognosis after radiation therapy. PMID- 10707686 TI - Disseminated infection with Phialemonium obovatum in a German shepherd dog. AB - A 4-year-old spayed female German Shepherd Dog was evaluated because of left forelimb lameness. A fungal granuloma on the distal portion of the radius was determined to be the cause of the lameness; the infecting organism was identified as Phialemonium obovatum. Despite aggressive treatment with amphotericin B, itraconazole, and ketoconazole and curettage of the local area, the dog developed systemic disease and was euthanatized 5 months after initial evaluation. Immune dysfunction may have played a role in development of disseminated disease, because although serum concentrations of total IgG, IgA, and IgM were within or greater than reference ranges, results of lymphocyte proliferation assays were abnormal, which indicated cellular immune dysfunction. Infection with Phialemonium obovatum should be considered as a differential diagnosis when branching fungal organisms are detected during histologic, cytologic, or microbiologic evaluation of tissue specimens. PMID- 10707687 TI - Intravenous administration of levothyroxine for treatment of suspected myxedema coma complicated by severe hypothermia in a dog. AB - A 7-year-old male English Coonhound with suspected myxedema coma complicated by severe hypothermia and metabolic abnormalities was treated with a combination of active external and core rewarming techniques, i.v. and oral administration of levothyroxine, supplemental oxygen, and administration of fluids (0.9% NaCl solution). Myxedema coma develops as a consequence of severe hypothyroidism and is characterized by a hypometabolic, stuporous state. Myxedema coma is associated with a high mortality rate, and most reported cases have involved Doberman Pinschers. Intravenous administration of levothyroxine can be used successfully in combination with oral administration to restore normal metabolic function and assist in warming and thermoregulation, although dosages should be conservative to avoid adverse cardiovascular effects. PMID- 10707688 TI - Hepatoblastoma with erythrocytosis in a young female horse. AB - A 2.5-year-old female Thoroughbred was examined because of lethargy, anorexia, and weight loss. Analysis of a CBC revealed erythrocytosis and an increase in PCV. Serum biochemical analysis revealed increases in activities of several hepatic enzymes. Ultrasonography revealed hepatomegaly and a heterogeneous appearance of the hepatic parenchyma. The horse did not improve despite supportive care, and it was euthanatized. Necropsy revealed numerous raised white to gray foci in the liver. Histologically, these foci consisted of neoplastic cells that resembled fetal hepatocytes, embryonal-type cells, and cells with features intermediate between those 2 cell types. Immunohistochemical staining revealed that hepatocytes stained strongly with anti-alpha-fetoprotein. On the basis of these results, hepatoblastoma was diagnosed. Diagnosis of hepatoblastoma is difficult, because it can appear histologically similar to other hepatic tumors, such as hepatocellular carcinomas. Definitive diagnosis requires histologic evaluation of tumor architecture and cell morphology. Immunohistochemical staining for alpha-fetoprotein in tumor cells may serve as a tumor marker but is not pathognomonic of hepatoblastoma. Paraneoplastic syndromes, such as erythrocytosis, can accompany hepatoblastoma. The prognosis for horses with hepatoblastoma is grave. PMID- 10707689 TI - Dynamic tracheal collapse as a cause of exercise intolerance in a thoroughbred. AB - A 2-year-old Thoroughbred filly was admitted to the hospital for evaluation of exercise intolerance. Resting videoendoscopic evaluation (i.e., while the horse was standing) of the nasopharynx and trachea revealed right arytenoid paresis and a tracheal defect that was 100 cm distal to the external nares. Surgery, consisting of a right prosthetic laryngoplasty, was performed. However, postoperative videoendoscopic evaluation revealed minimal abduction of the affected arytenoid cartilage. Dynamic videoendoscopic evaluation (i.e., while the horse was exercising) revealed the right arytenoid to be fixed in a submaximal position with no evidence of collapse into the airway. When the endoscope was positioned in the midcervical tracheal region, marked tracheal collapse was identified during exercise. Tracheal collapse can critically limit athletic function. Treatment of tracheal collapse depends on causative factors, the length of the trachea involved, and accessibility of the affected tracheal segment. The use of dynamic tracheal videoendoscopy should be considered in athletic horses with exercise intolerance in which the cause cannot be determined from resting or dynamic videoendoscopic evaluations of the nasopharynx. PMID- 10707691 TI - Abomasal displacement and volvulus in beef cattle: 19 cases (1988-1998). AB - OBJECTIVE: To assess signalment, history, results of clinical and laboratory testing, and outcome for beef cattle with a left displaced abomasum (LDA), right displaced abomasum (RDA), or abomasal volvulus (AV). DESIGN: Retrospective study. ANIMALS: 19 beef cattle with an AV, LDA, or RDA. PROCEDURE: Signalment; history; results of physical examination, diagnostic testing, and surgical exploration; and condition of the animal at discharge were obtained from medical records. RESULTS: Fourteen cattle had an AV, 4 had an RDA, and 1 had an LDA. Duration of clinical signs ranged from 1 to 21 days. Eighteen cattle had an AV or RDA; 7 were Brahmans, 12 were males, and median age was 10 months. Abdominal distention was observed in 11 cattle, heart rate of > or = 100 beats/minute was detected in 14, and the abomasum was palpable per rectum in all cattle in which per rectal examination was performed. Leukocytosis, neutrophilia, hyperglycemia, azotemia, hypochloremia, and hypokalemia were common laboratory findings. At surgery, 3 cattle with an AV or RDA had a ruptured abomasum. Of the remaining 15 cattle, 12 survived. CONCLUSIONS: Clinical course in beef cattle with an AV or RDA was more protracted than that typically associated with these conditions in dairy cattle, but survival rate in beef cattle that did not have rupture of the abomasum was similar to that of dairy cattle. CLINICAL RELEVANCE: Abomasal displacement should be considered for beef cattle with abdominal distention. Prognostic indicators recommended for use in dairy cattle may not be useful for beef cattle. PMID- 10707690 TI - Clinical and pathologic findings in donkeys with hypothermia: 10 cases (1988 1998). AB - OBJECTIVE: To describe clinical signs and clinicopathologic findings in donkeys with hypothermia. DESIGN: Retrospective study. ANIMALS: 10 hypothermic donkeys. PROCEDURE: Information on signalment, history, physical examination findings, results of diagnostic tests, treatments, and necropsy findings was extracted from medical records of all donkeys with hypothermia between 1988 and 1998 and compared with information from medical records of all normothermic donkeys and hypothermic horses admitted to the hospital during the same period. RESULTS: Donkeys were more likely to be hypothermic than horses. The mean age of hypothermic donkeys was 6 years (range, 7 months to 11 years), compared with 4.2 years (range, < 1 month to 15 years) for normothermic donkeys; this difference was not significant. Ten of 12 horses with hypothermia were neonates; there were no hypothermic neonatal donkeys. At admission, 7 of 8 hypothermic donkeys were in good body condition and all hypothermic donkeys were weak. Six hypothermic donkeys were able to maintain sternal recumbency, 1 remained in lateral recumbency, and 3 were able to stand. Of the 10 hypothermic donkeys, 2 survived, 1 died, and 7 were euthanatized. Histologically, the thyroid glands from 4 of 5 hypothermic donkeys appeared abnormal and were similar to those of foals with hypothyroidism. During the months that hypothermic donkeys were admitted, there was not a significant difference in environmental temperatures on days of admission between hypothermic and normothermic donkeys. CONCLUSIONS AND CLINICAL RELEVANCE: Hypothermia is a problem in donkeys during cold winter months, and may not be secondary to other disease or related to diet or management. PMID- 10707692 TI - Prevention, politics, and "having sex". PMID- 10707693 TI - Exploring the complexity of sleep disturbances in persons with HIV/AIDS. AB - Sleep disturbances are influenced by physiological, psychological, and environmental factors. Many persons living with HIV/AIDS manifest a variety of sleep disorders. Although completion of a brief sleep assessment can be helpful to determine the presence of a sleep problem, more in-depth investigation is necessary when sleep problems are reported. This pilot work used an open-ended interview guide developed from elements of Hauri's comprehensive sleep history. A convenience sample of six HIV-positive clients was interviewed during January 1997. In addition to answering open-ended questions about sleep-wake patterns, participants completed the Pittsburgh Sleep Quality Index and a personal characteristics form. HIV-severity information was obtained from the medical record. A complete sleep evaluation includes a thorough history and physical assessment, with a special focus on underlying pathology and medication use such as over-the-counter drugs, vitamins and herbal products, and mood-altering substance use. Treatment plans can include instruction about a variety of cognitive behavioral strategies. The nurse plays a major role in assessing, evaluating, teaching, and referring, which can contribute to identification of strategies that can improve quality of life. PMID- 10707694 TI - Nurse practitioners in rural California and AIDS. AB - A random sample of nurse practitioners located in the 26 rural counties of California were surveyed to assess their experience with HIV-positive patients. Of those selected, 70.5% participated; of those interviewed, 65.5% had seen a patient with HIV infection in the past 12 months. Of those providing primary care, 85% had seen HIV-positive patients. Their specific practice activities ranged from simply assessing to making all critical decisions with regard to therapeutic regimes in consultation with AIDS experts. Their most frequent activities involved education and/or counseling and the provision of psychosocial support. PMID- 10707695 TI - Treatment of fungal infections with ABLC in the home-care setting. AB - Invasive fungal infections are an important cause of morbidity and mortality in HIV-infected individuals. The management of opportunistic infections in a home care setting offers many psychological and economic advantages over hospitalization. Amphotericin B, the gold standard treatment for invasive fungal infections, is associated with significant adverse reactions, particularly nephrotoxicity, that make it difficult to administer as home infusion therapy. Lipid formulations of amphotericin B offer therapeutic alternatives to the parent compound with comparable efficacy, significantly lower rates of nephrotoxicity, and decreased infusion times versus the conventional form. Clinical experience with amphotericin B lipid complex injection in the treatment of fungal infections demonstrates its usefulness as an effective alternative to conventional amphotericin B in home infusion therapy. PMID- 10707697 TI - The relationship between social support and health in gay men with HIV/AIDS: an integrative review. AB - The purpose of this article is to analyze the existing research on the relationship between social support and health in gay men with HIV/AIDS. After literature review, 24 studies that met inclusion criteria were analyzed. It was found that the research could be divided into three distinct groups: (a) those studies that focused on the social networks of gay men and the role of significant others in regard to social support, (b) those studies that focused on coping and its relationship to social support, and (c) those studies that attempted to relate social support to other concepts such as hardiness, hope, depression, and illness concerns. A discussion of findings and conclusions are presented along with recommendations for future nursing knowledge development and practice. PMID- 10707696 TI - Most frequently used alternative and complementary therapies and activities by participants in the AMCOA study. AB - This literature review examines the current state of the scientific evidence published in peer-reviewed journals indexed in MedLine for the 10 most commonly noted alternative activities reported by the first 1,016 eligible participants in the Alternative Medical Care Outcomes in AIDS study. The most frequently used activities are aerobic exercise (64%), prayer (56%), massage (54%), needle acupuncture (48%), mediation (46%), support groups (42%), visualization and imagery (34%), breathing exercises (33%), spiritual activities (33%), and other exercise (33%). Despite frequency of usage, clinical research is not reported on MedLine to support the use of most of these activities for HIV/AIDS. The limitations of using MedLine as the sole source for this review are discussed. PMID- 10707698 TI - Hemophilia: a story of success--disaster and the perseverance of the human spirit. Part 1: Background and overview. PMID- 10707699 TI - Science is objective; isn't it? PMID- 10707700 TI - Post-exposure prophylaxis of occupational exposure to HIV infection. PMID- 10707701 TI - Support for vaccine development in primary HIV prevention. PMID- 10707702 TI - The impact of religion on adherence with antiretrovirals. PMID- 10707703 TI - Antimicrobial resistance--a strident alarm. PMID- 10707704 TI - A case for partial patellectomy. AB - The treatment of fractures of the the patella is a subject of controversy. Partial patellectomy with retention of a major fragment and suture of the quadriceps to it, seems reasonable. 18 cases of patella fracture underwent such a procedure. The average age of the patients was 47 years. Maximum recovery took an average of 5 months. There were 6 excellent results, 9 good, 3 fair. Results were assessed on the basis of pain, muscle wasting, quadriceps power, and range of knee motion. Total patellectomy and patella fixation as alternative modes of treatment are discussed. Partial patellectomy, whenever possible, is a good choice. PMID- 10707705 TI - Effect of Blalock Taussig shunt on clinical parameters, left ventricular function and pulmonary arteries. AB - Twenty children (mean age 3.25 years) with congenital cyanotic heart disease undergoing modified left Blalock-Taussig (BT) shunt were studied. The mean follow up period was 9.5 months (range 6 months to 1 year). The shunt was performed for cyanotic spells in 15 (75%) and hypoplastic pulmonary arteries in 5 (25%) patients. There were no immediate or late complications. None had cyanotic spell after the shunt. The mean arterial oxygen saturation improved from 66.47 +/- 11.9 to 76.97 +/- 8.16% (p = 0.0003) and mean hematocrit decreased from 51.55 +/- 9.5 to 46.5 +/- 9.7 (p = 0.002) after the shunt. The left atrial systolic volume and left ventricular diastolic volume also increased significantly following the shunt (from 15.82 +/- 6.37 to 20.83 +/- 8.91 ml p = 0.006 and from 36.13 +/- 16.08 to 41.08 +/- 20.07 ml (p = 0.01) respectively. There was significant growth of main, right and left pulmonary arteries and pulmonary valve annulus after the procedure. PMID- 10707706 TI - Family planning: views of female non-acceptors in rural India. AB - A study conducted on eligible rural women who were unwilling to accept family planning methods revealed that many women were concerned about child survival and viewed children as a source of support in old age. Family size was usually decided by in-laws. Pressure from in-laws to have more children was significantly higher in families where the women were less educated or illiterate. PMID- 10707707 TI - Disseminated mucormycosis in healthy adults. AB - Three patients of disseminated mucormycosis are described. None had predisposing factors. Two of them presented with nonspecific symptoms along with acute renal failure and peritonitis. Third patient had fulminating primary cutaneous mucormycosis which disseminated later. Development of acute renal failure with smooth enlargement of both kidneys in an apparently healthy individual or appearance of mould in a wound should raise the suspicion of mucormycosis. The hallmark of the infection was vascular invasion and thrombosis. Antemortem diagnosis could be made in one patient only. All patients had progressive downhill course despite supportive treatment, antibiotic and amphotericin in-B in one patient. PMID- 10707708 TI - Isolated jejunal varices. AB - Isolated jejunal varices are an uncommon manifestation of portal hypertension. A one and a half year old boy presented with recurrent, massive gastrointestinal bleeding from jejunal varices. The bleeding site was identified at exploratory laparotomy. Jejunal resection and anastomosis resulted in complete resolution of the bleeding and there has been no recurrent bleeding over an eight month follow up period. PMID- 10707709 TI - Situs inversus with cholelithiasis. AB - Situs inversus totalis is a form of heterotaxia which is usually detected accidentally while investigating for any associated condition. If undetected, this condition can create a diagnostic puzzle. We report one such case in which situs inversus was associated with cholelithiasis. PMID- 10707710 TI - Anterior urethral valve in an adolescent boy. AB - A 14 year old boy with a relatively uncommon anterior urethral valve is described herein. PMID- 10707711 TI - Fracture sacrum. AB - An extremely rare case of combined transverse and vertical fracture of sacrum with neurological deficit is reported here with a six month follow-up. The patient also had an L1 compression fracture. The patient has recovered significantly with conservative management. PMID- 10707712 TI - Rectal duplication. AB - Duplications of the alimentary tract are of a great rarity, particularly so in the rectum. Because of its rarity, the difficulty of making a correct diagnosis and of selection of proper approach for treatment, this entity bears a special significance. The present case report deals with a female newborn who presented with imperforate anus and a rectovestibular fistula and a mass prolapsing at the introitus. Complete excision of the mass was carried out through the perineal approach and the child then underwent, a PSARP for the correction of the rectal anomaly. Histology confirmed the mass to be a rectal duplication. PMID- 10707714 TI - Fracture of the entire posterior process of the talus. AB - A 25 year old, who had sustained a fracture of the entire posterior process of the talus, is presented. THe fracture was successfully managed conservatively. PMID- 10707713 TI - Thyroid abscess. AB - Thyroid abscess arising from Acute Suppurative Thyroiditis (AST) is a rare clinical disorder. The ability of the thyroid gland to resist infection is well known and infection in the thyroid gland is rare, particularly so with the advent of widespread usage of antibiotics. An internal pharyngeal fistula (Pyriform sinus fistula) is the most common underlying abnormality in patients with AST. We report a case of an adult male who presented with a thyroid abscess. The causal organism was found to be Staphylococcus aureus. Intravenous antibiotics and, incision and drainage of the abscess led to an uncomplicated recovery. PMID- 10707715 TI - Evaluation of solitary pulmonary nodule. PMID- 10707716 TI - Anterior seromyotomy with posterior truncal vagotomy in uncomplicated chronic duodenal ulcer. AB - Thirty cases of uncomplicated duodenal ulcer treated by anterior superficial lesser curvature seromyotomy and posterior truncal vagotomy were studied to evaluate the efficacy of this procedure. There was completeness of vagotomy in all the cases as shown by endoscopic Congo Red test. Twenty-seven cases were asymptomatic at 1-48 months (Mean 22.3) follow up, while 3 patients had controllable side effects such as dumping and diarrhoea. There was no mortality. This procedure is safe, effective and is a favourable alternative to highly selective vagotomy. PMID- 10707717 TI - Roxatidine in duodenal ulcer. AB - Roxatidine acetate is a new H2-receptor antagonist. A randomized double-blind clinical trial in fifty-three patients with endoscopically proven duodenal ulcers > 5 mm in diameter was undertaken to compare safety and efficacy of roxatidine with that of ranitidine. Twenty-six patients received roxatidine (75 mg bid) while 27 patients received ranitidine (150 mg bid) for 4 weeks. One patient in each group did not come for follow up. Roxatidine and ranitidine had comparable ulcer healing rates (22/25 vs 22/26); roxatidine, however, resulted in greater reduction in the number and severity of night time pain episodes (p < 0.05). No adverse event was reported during 4 weeks of treatment with roxatidine. Thus roxatidine achieves the primary therapeutic goal of relief of pain better than ranitidine. PMID- 10707718 TI - Serum immunoglobulin profile in normal Kashmiri adults. AB - Serum levels of the immunoglobulins IgG, IgA and IgM were estimated in 102 apparently healthy Kashmiri adults in the age group of 16-60 years, using single radial immunodiffusion method of Mancini et al. The mean serum levels of IgG, IgA and IgM were observed to be 1289.19 +/- 234.9, 216.18 +/- 50.70 and 118.97 +/- 41.88 respectively. No significant difference in the mean serum levels was observed between the two sexes as such, but IgM showed a significant increase in females in the age group of 16-30 years. IgA showed a significant increase with age, with no such increase in case of IgG and IgM. PMID- 10707719 TI - Septal splint with wax plates. AB - To pack or not to pack, has always been a debate, especially after septal and functional endoscopic sinus surgery. The authors have studied the symptoms of packing versus not packing in their series of 100 patients having undergone nasal surgery. They advocate the use of dental wax for the fashioning of septal splints, since they are easy to introduce, cheap and malleable. The patients postoperative comfort is greatly enhanced with the use of dental wax plate splints instead of nasal packing. PMID- 10707720 TI - Nephroblastomatosis--pathologic and imaging characteristics. PMID- 10707721 TI - Acanthamoeba keratitis. AB - Acanthamoeba keratitis, common in soft lens wearers, is not commonly isolated. The reports of Acanthamoeba keratitis in Indian literature are few. We report here a case of Acanthamoeba Keratitis in a medical student using soft contact lenses, initially diagnosed and treated as a bacterial and later as a viral corneal ulcer, who responded extremely well to medical line of therapy. PMID- 10707722 TI - Distal fibular giant cell tumour. AB - A patient who reported with a slowly growing swelling ovr thee lateral aspect of the left ankle, was investigated and diagnosed to have a giant cell tumour which was confirmed on FNAC. The tumour was managed with excision biopsy and reconstruction. The case is being reported for its rare site of occurrence. PMID- 10707723 TI - Extradural lipomatosis presenting with paraplegia. AB - An unusual case with spinal extradural lipomatosis in a non-obese and otherwise healthy man is reported. The patient presented with a history of weakness of legs which progressed to paraplegia over a 40 day period. PMID- 10707724 TI - Sebaceous adenoma in the region of the medial canthus causing proptosis. AB - A case of sebaceous adenoma in the region of the medial canthus causing proptosis is presented along with a review of the medical literature. The clinicopathological aspects of the tumour are discussed. The mode of treatment was surgical excision. A six month follow-up showed a reduction in the proptosis with no recurrence. PMID- 10707725 TI - Lymphangioma of the chest wall. AB - A lymphangioma of the chest wall, hitherto unreported is described here. PMID- 10707726 TI - Isolated involvement of the mandible by non-Hodgkin's lymphoma. AB - A case report of isolated involvement of the mandible by non-Hodgkin lymphoma is presented. The patient presented with a non-healing ulcer following a tooth extraction. Biopsy revealed an undifferentiated cancer. Investigations failed to reveal any involvement of the organs. A hemimandibulectomy was performed followed by radiotherapy on receipt of the histopathological diagnosis of non-Hodgkin lymphoma. The patient is asymptomatic two and a half years after treatment. PMID- 10707727 TI - Symphysis fundal height curve--a simple method for foetal growth assessment. AB - In this prospective study, symphisis fundal height (SFH) was measured in centimeters at different weeks of gestation from 20th week onwards in 100 healthy women with uncomplicated pregnancies. A curve was plotted based on the mean SFH measurements with standard deviation. Readings were also arranged on the basis of 10th, 50th and 90th percentiles. Percentile curve was similar to the curve based on mean with standard deviation. The rate of growth was 1 cm per week between 20 32 weeks. Thereafter, there was a slight fall in the rate of growth. SFH measurement is a simple method of foetal growth assessment which can be utilized even by paramedical workers to screen for small for gestational age babies. It is better to have a standard curve derived from the population as there is regional variation. PMID- 10707728 TI - The impact of cisplat based chemotherapy on advanced ovarian cancer. AB - Cisplatinum based chemotherapy has become the standard treatment for ovarian cancers due to its proved superiority over non-cisplat based regimes. However, the therapeutic impact of cisplat based regimes compared to cheaper non cisplatinum based regimes is questionable when multiple variables such as residual disease, histologic type, grade are introduced. This report is a study of 110 Stage III ovarian cancer patients from 1985-89, with cisplat (n = 69) and non cisplat (n = 41) based chemotherapy. The results of both regimes with reference to the multiple variable factors are presented. We conclude that cisplat based regimes appear to be superior to non-cisplat based regimes except probably in poorly differentiated ovarian tumors where the results were similar with either regimen. PMID- 10707729 TI - Alpha blocker prazosin for the treatment of benign prostatic hypertrophy (BPH). AB - In this study medical treatment with alpha blocker-prazosin is compared with transurethral resection of prostate (TURP) in 62 patients suffering from benign enlargement of prostate with a gland size of less than 20 gms. After thorough interrogation patients were offered either TURP or prazosin therapy. Symptom scoring, residual volume of urine and urinary flow rates were estimated in both the groups before and 3 months after the therapy. 23.5% patients in prazosin group while 90% of patients in TURP group had significant improvement. This distinctly brings out the superiority of TURP for benign enlargement of prostate. PMID- 10707730 TI - Is McMurray's osteotomy obsolete? AB - A review of the method of performing, advantages, disadvantages of McMurray's displacement osteotomy with regard to treatment of nonunion of transcervical fracture neck femur with viable femoral head was carried out in this study of ten cases, in view of the abandonment of the procedure in favour of angulation osteotomy. Good results obtained in the series attest to the usefulness of McMurray's osteotomy in the difficult problem of nonunion of transcervical fracture neck femur in well selected cases with certain advantages over the angulation osteotomy due to the 'Armchair effect'. PMID- 10707731 TI - Serogroup prevalence of Shigellae in Bombay. AB - Prevalence of Shigellae serotypes in Bombay was studied from June 1988 to May 1991. A total of 2758 faecal specimens were collected from paediatric patients (< 12 yrs) with acute gastroenteritis. A total of 90 Shigella were isolated giving the isolation rate of 3.2%. Shigella flexneri was the predominant serogroup (73.3%) followed by Shigella dysenteriae (16.6%). All the isolates were sensitive to nalidixic acid. Eighty percent of the Shigellae were multidrug resistant. Present data were compared with the study carried out during the period of 1983 87 from the same institute. A change in the serogroup prevalence was noted wherein Shigella flexneri dominated over Shigella dysenteriae since 1985. Increase in resistance to ampicillin and cotrimoxazole was seen in Shigella flexneri strains as compared to previous years. PMID- 10707732 TI - Short colon variant. AB - A one day old neonate with a short colon, associated exomphalos minor; bifid scrotum and ileovesical fistula is reported. PMID- 10707733 TI - Typhoid fever in a 7 month old infant. AB - The clinical profile of typhoid fever in an infant is variable and non-specific. A rare case of typhoid fever in a 7 month old infant is reported. The child presented with only a day's history of fever and loose motions which resulted in severe dehydration, acute tubular necrosis and death. The diagnosis of typhoid fever was made only on post-mortem study. The problem in diagnosing typhoid fever in a young infant is highlighted with a brief literature review on the subject. PMID- 10707734 TI - Tuberculous arthritis of the knee--an unusual presentation. AB - A 54 year old male who had an unusual clinical manifestation and radiological features proven to have tuberculosis arthritis of the knee on synovial biopsy is presented here. PMID- 10707735 TI - Waardenburg syndrome with anisocoria and exotropia. AB - A case of Waardenburg syndrome with unusual features such as anisocoria, exotropia is reported. PMID- 10707736 TI - Solitary synovial osteochondroma of the knee. AB - A very rare case of solitary osteochondroma of the knee is reported. The patient presented with a slowly growing retropatellar bony tumour of 4 years duration following a minor trauma. An excisional biopsy with a total patellectomy was performed as the patellar articular surface was unsalvageable. A 20 month follow up revealed no recurrence and a functional knee. A brief review of literature is also presented. PMID- 10707737 TI - Total hip arthroplasty in healed tuberculous hip. AB - Total hip arthroplasty was performed in a patient who had tuberculous hip, quiescent for the last 15 years, without pre or postoperative anti-tuberculous chemotherapy. At a 27 month follow up, there has been no evidence of reactivation. A brief review of relevant literature is also presented. PMID- 10707738 TI - [Acute abdomen: consider also the thorax]. AB - Three patients, 2 men aged 22 and 62 years en 1 woman aged 49, presented with symptoms of an acute abdomen. While infiltrative signs were described on radiodiagnostic images two patients underwent laparotomies. In all three subsequently the diagnosis of pneumonia was established and the patients made full recovery after antibiotic therapy. When a patient presents with symptoms of an acute abdomen, the possibility of an existing pneumonia should always be borne in mind. It is therefore recommended to make a chest radiograph with frontal and lateral view. In the presence of infiltrative signs the existence of pneumonia as the cause of abdominal symptoms should be considered in order to avoid unnecessary laparotomy. PMID- 10707739 TI - [Additional effects of statins independent of the cholesterol-lowering as yet not shown to be clinically relevant]. AB - The reduction in cardiovascular morbidity and mortality during the use of statins is due to their cholesterol lowering effect. In addition, the statins are assumed to exert lipid lowering independent actions, which may contribute to the beneficial effect. However, the evidence for these effects is based on less solid sub- and post hoc analyses of clinical trials and in vitro and animal experiments in which unrealistic doses of statins are used. On closer examination, most of these so-called lipid independent effects are in fact based on either direct or indirect cholesterol lowering. The clinical relevance has to be shown in prospective clinical trials. PMID- 10707740 TI - [Prenatal ultrasonic detection of asymptomatic urogenital abnormalities: advantages and disadvantages]. AB - Prenatal ultrasound studies have significantly contributed to the understanding of congenital abnormalities of the urinary tract and the development of pediatric urology over the past decade. Besides providing more insight into pathophysiology of the developing urinary tract, it has promoted the possibilities of early postnatal intervention and of studying the natural history of developmental abnormalities such as pelviureteral junction obstruction and vesico-ureteric reflux. The downside of this development has been and probably still is a considerable amount of diagnostic and therapeutic interventions for so-called abnormalities which, left alone, will disappear with time and without significant sequelae. On the other hand, many children with severe obstructive uropathy are alive today because of early intervention. Longterm follow-up studies are needed to prove the efficacy of prenatal screening and to ascertain the contribution of these technical possibilities to quality of life. PMID- 10707741 TI - [Asymptomatic dilatation of the pyelocaliceal system in young children detected by echography: altered insights and current management]. AB - Since the introduction of antenatal ultrasound, the age of patients presenting with pyelocaliceal dilatation has continued to decrease: before the era of ultrasound most patients with this diagnosis were symptomatic; today most patients are diagnosed before birth and most of them are without symptoms. The current diagnostic tools are diuretic renography and ultrasound, after vesicoureteral reflux has been excluded. The realization that obstructive renographic drainage curves were not reliable indicators of obstruction in this patient group eventually altered the therapeutic approach from early operative intervention to a more conservative management in patients with a good relative kidney function. Ultrasound has significant value in detection of patients at risk of loss of renal function. With this management only a quarter appear to have a significant ureteropelvic junction stenosis needing surgery. PMID- 10707742 TI - [Statins: possibly more than just lowering of the lipid level]. AB - Clinical trials have demonstrated that treatment of hypercholesterolemia with HMG CoA reductase inhibitors (statins) is beneficial in primary and secondary prevention of vascular diseases. The observed reduction in cardiovascular morbidity and mortality cannot only be explained by lipid-lowering only. Apart from lowering cholesterol, statins conceivably also exert effects on the vascular wall that may directly contribute to decrease of vascular incidents: (a) a favourable influence on endothelial dysfunction through stimulation of nitrous oxide synthetase: (b) stabilization of plaques by reducing influx of macrophages into the vascular wall and decreasing the production of matrix metalloproteinases, that may affect the connective tissue cover of the plaque: (c) inhibition of the initiation and progression of atherosclerosis by reducing adhesion of leukocytes to the vascular wall: (d) reducing the haemorrhagic diathesis by increasing the fibrinolytic capacity and inhibiting tissue factor expression on macrophages. All these effects of statins independent of the lowering of the cholesterol level might contribute to primary and secondary prevention of vascular incidents. While most nonlipid mechanisms of statins are being studied in vitro and in animals, the clinical relevance is still to be determined. PMID- 10707743 TI - [Roaming through methodology. XVII. The placebo effect]. AB - The placebo effect is well known, but there are many misconceptions. One of these misconceptions is that one-third of patients respond to placebos. This misunderstanding is probably due to methodologically poor research conducted in the 1950s. Another error is that the effect in the placebo arm of a clinical trial is often confused with the placebo effect. The belief in the placebo effect is enormous, but the quantity and quality of data to substantiate this belief are very limited. Investigating the placebo effect is methodologically difficult, not easy to get financed and considered unrewarding. PMID- 10707744 TI - [Percutaneous endoscopic gastrostomy in children with psychomotor retardation; less complaints and not as stressful]. AB - OBJECTIVE: To evaluate the effects of percutaneous endoscopic gastrostomy (PEG) in children with psychomotor retardation. DESIGN: Prospective. METHOD: Data on symptoms, pros and cons and complications were collected by means of questionnaires from the parents of children with psychomotor retardation and severe nutritional problems in whom a PEG tube had been introduced between August 1995 and March 1998 in the department of Children's Gastroenterology and Nutrition of the Emma Children's Hospital/Academic Medical Centre in Amsterdam, the Netherlands, comparing the situations before the introduction and 6 and 18 months afterward. RESULTS: The patient group consisted of 17 boys and 23 girls with a mean age of 6 years and 3 months (range 8 months-10 years and 1 month). The frequency of vomiting and of airway infections decreased and the general nutritional condition improved. Restlessness and pain occurred less often in over half the children. The disadvantages most often reported were the logistics concerning the feeding (n = 11) and the reduced contact with the child (n = 10). Thirteen children displayed mild side effects such as skin irritation and proud flesh. Technical problems consisted of leakage (n = 11) and obstruction of the tube (n = 2). In one child, introduction of the tube was followed by a major complication. CONCLUSION: The PEG tube in this patient group reduced the frequency of complaints about nutrition and constituted a patient-friendly alternative to the nasal tube. PMID- 10707745 TI - [Same-day diathermic hemorrhoidectomy in 48 patients; few complications and tolerable pain]. AB - OBJECT: To measure pain characteristics and to ascertain patient satisfaction and level of complications after day-care haemorrhoidectomy. DESIGN: Prospective. METHOD: Diathermic haemorrhoidectomy was performed in daytime care in 40 successive patients in hospital De Heel, Zaandam in 1997-1998. The operation was part of a package of measures, such as extensive counseling, preoperative bulking agents, surgery performed by colorectal surgeon, dedicated anaesthesiological techniques, adequate pain medication, and frequent outpatient clinic visits. RESULTS: The 40 patients were 21 men and 19 women with a mean age of 43 years (range: 27-67). One male patient was admitted for 24 hours because of urinary retention and 1 female patient developed a wound infection. No other serious complications were seen. During the first 5 postoperative bowel movements 75% of the total pain score was obtained. Postdefaecatory pain lasted on average 81 min on day 1 to 8 min on day 7. Time away from work was on average 6.4 days (range: 0 12). In between clinic visits 5 patients consulted their general practitioners. After 6 weeks 95% of the patients would again have consented to day-care haemorrhoidectomy. CONCLUSION: Diathermic haemorrhoidectomy has a low complication rate and tolerable pain and can be performed in day care. PMID- 10707746 TI - [The miraculous growth of "quackery funds"]. AB - In 1998 and 1999 three new charity foundations were established in the Netherlands, covering the same areas as the Dutch Heart Foundation, the Netherlands Cancer Foundation and the National Rheuma Foundation respectively. Analysis of the set-up of the three new foundations, of their philosophy, their constitutions and the background of the persons involved shows a remarkably uniform pattern, making it very likely that the set-up of these alternative foundations is a part of the marketing strategy of some Dutch wholesalers in orthomolecular drugs. More legal regulation of charity foundations is seriously needed. PMID- 10707748 TI - [In Process Citation] PMID- 10707747 TI - [Electronic medical record, confidentiality and safeguarding of privacy]. AB - An electronic patient file is expected to contribute to individual health care, as well as to policy goals. For acceptance by doctors and patients, some conditions have to be fulfilled. Among them is the appropriate specification of the right of the patient to medical secrecy and privacy. Reference is made to new legal requirements (Wet Bescherming Persoonsgegevens (Act on the protection of personal data)), which e.g. regulate when access to personal medical data is lawful, when consent is required and in what form, in addition to a number of other conditions. The current information and communication technology must not be allowed to dictate the possibilities and limitations of legally applicable standards, but the technical implementation of the electronic patient file must be made to suit and assure the legal standards of medical professional secrecy and patient privacy. Otherwise the advantages of an electronic patient file will be out-weighed by patients withholding important information from their physician and by their being less inclined to consent to their data being used for e.g. scientific investigations. PMID- 10707749 TI - [Somatic and psychological symptoms in soldiers after military clashes and peace keeping missions]. PMID- 10707750 TI - [Somatic and psychological symptoms in soldiers after military clashes and peace keeping missions]. PMID- 10707751 TI - [Clinical results and cost due to improved antibiotics policies]. PMID- 10707752 TI - Long-term effects of clinical outcome with low and high dose in the Captopril in Heart Insufficient Patients Study (CHIPS). AB - BACKGROUND: Although angiotensin-converting enzyme inhibitors are recommended as first line therapy in patients with chronic heart failure, the target doses proven to be effective in major morbidity and mortality trials (e.g. captopril 50 mg b.i.d), are generally not used in daily practice in Belgium. AIM: The objective of this study (CHIPS, Captopril in Heart Insufficient Patients Study) was to compare the long-term effects of a low dose (25 mg b.i.d.) and a high dose (50 mg b.i.d.) of captopril in mild to moderate heart failure. After a titration period of at least 10 days, patients who tolerated 50 mg b.i.d., were randomly assigned to receive either the low dose or the high dose of captopril and followed up to 2 years. RESULTS: 298 patients were included and were followed up for a mean of 12 months. Progression in heart failure seems to be favourably influenced by therapy with high dose in comparison to low dose; a relative difference of 29% in the rates of heart failure worsening was observed between the two doses, 31.5% and 22.4% for low and high dose (p = 0.088), respectively. Treatment with high dose showed also a trend to benefit as compared to low dose in reducing the number of hospitalizations for all causes from 22.4 to 14.5% (p = 0.1) and for congestive heart failure from 14.7 to 7.2% (p = 0.06); moreover, the incidence of fatal and nonfatal cardiac events showed a trend in favour of the high dose of 22% (p = 0.142). The total number of adverse events was comparable for both doses, but dizziness and hypotension were a little more frequently reported in the high-dose group. Serum creatinine values showed no significant changes either in the low-dose or in the high-dose group. CONCLUSION: In the CHIPS-study, in comparison to a low dose, therapy with a high dose of captopril tends to improve the long-term clinical outcome of patients with mild to moderate heart failure without significantly more toxicity. PMID- 10707753 TI - Effects of infusion of glucose-insulin-potassium on myocardial function after a recent myocardial infarction. AB - We studied the effects of glucose, insulin, and KCl infusion (GIK), on regional myocardial perfusion and function by 99m-Tc-tetrofosmin-gated SPECT. METHODS: We studied 21 male patients with their first uncomplicated acute myocardial infarction (AMI). All patients underwent a rest and submaximal stress before and after 24-hour infusion of GIK-solution (group A) or saline solution (group B). RESULTS: Group A showed better stress tolerance and ischaemic threshold improvement after GIK infusion whilst no statistical differences were found between basal and post-infusion test in group B. At first the stress test in group A, of the 192 segments analysed, 52 (27%) showed reversible perfusion defect. In group B, of 144 segments analysed, 31 (21%) showed reversible perfusion defect. A post-infusion analysis in group A showed a post-GIK end diastolic significant count improvement in 21 segments, and a post-GIK end systolic count improvement in 22 segments. In group B, perfusion increase was observed only in 4 segments, whilst systolic thickening increase was observed only in 1 segment. CONCLUSION: These data demonstrate the efficacy of GIK infusion to improve regional myocardial perfusion and function mainly in segments adjacent to the recently infarcted area. PMID- 10707754 TI - Clinical epidemiology of acute myocardial infarction in Kuwait. AB - OBJECTIVE: We studied the clinical epidemiological features of patients with AMI treated at a major hospital in Kuwait. The objectives of the study were to determine (i) personal and clinical characteristics of patients; (ii) prevalence of major risk factors among the patients; and (iii) factors associated with in hospital morbidity and mortality. METHODS AND RESULTS: All patients (n = 126) who fulfilled the standard diagnostic criteria for AMI and treated at the CCU of the study hospital during the calendar year 1996 were included in the study. Patients were identified from the CCU register and information was extracted from the medical records. Multiple logistic regression was performed to study the factors independently associated with in-hospital morbidity and mortality. Of the 126 patients, 84.9% were men and 15.1% were women; and 40.5% were Kuwaiti nationals and 59.5% were expatriates. On average, male patients were younger than females (mean age = 52.4 +/- 10.4 years vs. 60.2 +/- 10.2 years), and male expatriates were the youngest sub-group in the study (mean age = 49.7 +/- 8.7 years). Overall, history of diabetes, hypertension, and CHD was recorded in 44.4%, 29.6%, and 16.8% of the patients, respectively. About 58% of the male patients were current smokers and the prevalence of smoking was significantly higher in expatriates compared with Kuwaiti patients (62% vs 36%, respectively). Kuwaiti nationals had a significantly high prevalence of diabetes compared with the expatriates (57% vs 36%, respectively). There was a significant trend in increasing prevalence of diabetes, hypertension and CHD with age whereas smoking was most prevalent (87%) in the youngest age group. As for the clinical features, 70.5% of the patients presented within 6 hours of the onset of symptoms, 73% presented with ST-segment elevation (40.5% with inferior, 32.5% with anterior ST segment elevation), and fibrinolytic therapy was given to 63.6% of the patients. About half of the patients had an admission blood glucose level of > or = 10 mmol/l, and 70.6% had a fasting blood glucose level of > or = 6.1 mmol/l one day after admission. In the multiple logistic regression analysis, old age (> 60 years), anterior MI, and admission blood glucose level of > or = 10 mmol/l were found to be significantly associated with in-hospital cardiac morbidity. The in hospital case fatality was about 6%. Old age and history of CHD were found to be the significant predictors of in-hospital mortality. CONCLUSION: Control of smoking and early diagnosis and appropriate treatment of diabetes may reduce the burden of AMI-related morbidity and mortality. PMID- 10707755 TI - Exercise-induced ST-segment depression: imbalance between myocardial oxygen demand and myocardial blood flow. AB - OBJECTIVE: ST-segment depression is believed as a common electrocardiographic sign of myocardial ischemia during exercise testing. Ischemia is generally defined as oxygen deprivation due to reduced perfusion. However, the exact relationship of the ischemic definition to ST-segment depression remains unclear. This study was conducted to evaluate the correlation between myocardial oxygen demand and myocardial blood flow (MBF) when ischemic (horizontal or downsloping) ST-segment depression of > or = 0.1 mV 80 ms after the J point developed during low-level exercise. METHODS AND RESULTS: Seventy-two patients with angiographically proven coronary artery disease (CAD) and 9 healthy volunteers underwent exercise positron emission tomography (PET). Myocardial oxygen demand was defined as a rate-pressure product (RPP, heart rate x systolic blood pressure) during exercise and MBF was quantified by nitrogen-13 ammonia perfusion PET. The myocardial demand-supply balance (MDSB) index was calculated from the MBF ratio (values during exercise/values at rest) against the RPP ratio (values during exercise/values at rest). The MDSB index was significantly lower in patients with ischemic ST-segment depression than in patients with non-ischemic ST depression and healthy volunteers (0.82 +/- 0.16 vs. 1.02 +/- 0.17, p < 0.0001 and vs. 0.99 +/- 0.14, p = 0.0109). Further, the presence of inadequate increase in MBF of < or = 10% (2 SD below the mean % value of healthy volunteers) during exercise in regional myocardium perfused by stenotic CAD significantly correlated with exercise-induced ischemic ST-segment depression (p = 0.0105). CONCLUSIONS: Our study could demonstrate that exercise-induced ischemic ST-segment depression is associated with myocardial ischemia due to exercise-induced imbalance between myocardial oxygen demand and global and regional MBF supply in patients with proven CAD. PMID- 10707756 TI - Stroke in a hospital-derived cohort of patients with chronic Chagas' disease. AB - The aim of this study was to determine the prevalence of stroke in a hospital derived cohort of patients with chronic Chagas' disease. Seventy-nine patients with chronic Chagas' disease were prospectively followed at the Cardiomyopathy Clinic of the Santa Casa Hospital from January 1990 to June 1993 (mean follow up = 17 +/- 12 months). Mean New York Heart Association functional class was 2.42 +/ 1.24. Fifty-six (70%) patients were on angiotensin-converting enzyme inhibitors at maximum tolerated doses, but no patient was on anticoagulation therapy. Atrial fibrillation was detected on the resting ECG in twelve (15%) patients. On echocardiography, mean left ventricular ejection fraction was 49.07 +/- 17.96% and mean left ventricular diastolic dimension 60.12 +/- 10.97 mm; mitral regurgitation was detected in 20 (29%) patients. Left ventricular thrombus was seen in three (4%) patients; all of them were in sinus rhythm and had left ventricular dysfunction on echocardiography. No thromboembolic event, however, was detected during the follow-up. One patient (1%) had a fatal stroke during the study period; she was in sinus rhythm on the resting ECG, and had mild mitral regurgitation, normal left ventricular function and no intracavitary thrombus on Doppler echocardiography. The prevalence of stroke is low in a hospital-derived cohort of patients with mild to moderate heart failure due to chronic Chagas' disease. Routine prophylactic anticoagulation, therefore, seems not to be warranted. PMID- 10707757 TI - Double-chamber right ventricle in a 63-year-old woman. AB - Double-chamber right ventricle (DCRV) is a rare congenital heart disease consisting in right ventricular obstruction due to one or several anomalous muscle bundles that divide the right ventricle into two chambers. Because of the rarity of this anomaly in adults, we present the case of a 63-year-old woman suffering from this heart disease, being on the other hand, one of the few cases described in the literature in such an old patient. PMID- 10707758 TI - Congestive heart failure treated by the upgrade from VVI to DDD pacing. AB - The case is presented of an elderly woman with normal left ventricular (LV) systolic function and VVI pacing complicated by severe congestive heart failure. The symptoms and findings of congestive heart failure became refractory to medical treatment and resolved with the upgrade of the VVI to a DDD system. Right heart catheterization during VVI pacing showed increased mean pulmonary capillary wedge and right atrial pressures both being normalized under DDD pacing. This case report illustrates the need to consider permanent physiological pacing in elderly patients, even in presence of normal LV systolic function, to ensure AV synchrony when the atrium can be paced, since diastolic LV dysfunction is quite common in these subjects. PMID- 10707759 TI - Flecainide-associated pneumonitis with acute respiratory failure in a patient with the LEOPARD syndrome. AB - Antiarrhythmic agents can cause pneumonitis. Flecainide is a rare cause of hypersensitivity pneumonitis, and few cases have been reported. We describe a case of interstitial hypoxaemiant pneumonitis probably related to flecainide in a patient with the LEOPARD syndrome, a rare congenital disorder. PMID- 10707760 TI - Stenosis of a mechanical mitral prosthetic valve in a patient with systemic lupus erythematosus. AB - A young woman with systemic lupus erythematosus (SLE) developed progressive heart failure several years after mitral valve replacement with a Bjork-Shiley prosthesis for treating mitral stenosis due to Libman-Sacks endocarditis. She was admitted to the hospital with pulmonary oedema. Transoesophageal echocardiography revealed stenosis of the mitral prosthesis, which was covered by fibrous tissue. Replacement of the prosthesis was done but the patient died from cerebral haemorrhage three days later. Although three cases of prosthetic valve dysfunction in SLE have been documented so far, this is to our knowledge the first report of a SLE recurrence on a tilting disc mechanical valve. PMID- 10707761 TI - Twiddler's syndrome. PMID- 10707762 TI - Enhanced taurine release in cultured cerebellar granule cells in cell-damaging conditions. AB - The release of taurine from cultured cerebellar granule neurons was studied in different cell-damaging conditions, including hypoxia, hypoglycemia, ischemia, oxidative stress and in the presence of free radicals. The effects of both ionotropic and metabotropic glutamate receptor agonists on the release were likewise investigated. The release of [3H]taurine from the glutamatergic granule cells was increased by K+ (50 mM) and veratridine (0.1 mM), the effect of veratridine being the greater. Hypoxia and ischemia produced an initial increase in release compared to normoxia but resulted in a diminished response to K+. Hypoglycemia, oxidative stress and free radicals enhanced taurine release, and subsequent K+ treatment exhibited a correspondingly greater stimulation. A common feature of taurine release in all the above conditions was a slow response to the stimulus evoked by K+ and particularly to that evoked by veratridine. All ionotropic glutamate receptor agonists potentiated taurine release, but only the action of kainate seemed to be receptor-mediated. Metabotropic receptor agonists of group I slightly stimulated the release. The prolonged taurine release seen in both normoxia and cell-damaging conditions may be of importance in maintaining homeostasis in the cerebellum and reducing excitability for a longer period than other neuroprotective mechanisms. PMID- 10707763 TI - Production of tyrosine and other aromatic compounds from phenylalanine by rumen microorganisms. AB - Rumen contents from three fistulated Japanese native goats fed Lucerne hay cubes (Medicago sativa) and concentrate mixture were collected to prepare the suspensions of mixed rumen bacteria (B), mixed protozoa (P) and a combination of the two (BP). Microbial suspensions were anaerobically incubated at 39 degrees C for 12 h with or without 1 mM of L-phenylalanine (Phe). Phe, tyrosine (Tyr) and other related compounds in both supernatant and microbial hydrolysates of the incubations were analyzed by HPLC. Tyr can be produced from Phe not only by rumen bacteria but also by rumen protozoa. The production of Tyr during 12 h incubation in B (183.6 mumol/g MN) was 4.3 times higher than that in P. One of the intermediate products between Phe and Tyr seems to be p-hydroxyphenylacetic acid. The rate of the net degradation of Phe incubation in B (76.0 mumol/g MN/h) was 2.4 times higher than in P. In the case of all rumen microorganisms, degraded Phe was mainly (> 53%) converted into phenylacetic acid. The production of benzoic acid was higher in P than in B suspensions. Small amount of phenylpyruvic acid was produced from Phe by both rumen bacteria and protozoa, but phenylpropionic acid and phenyllactic acid were produced only by rumen bacteria. PMID- 10707764 TI - Evidence that taurine modulates osmoregulation by modification of osmolarity sensor protein ENVZ--expression. AB - Although the involvement of taurine in osmoregulation is well-documented and widely accepted, no detailed mechanism for this function has been reported so far. We used subtractive hybridization to study mRNA steady state levels of genes up- or downregulated by taurine. Rats were fed taurine 100 mg/kg body weight per day for a period of three days and hearts (total ventricular tissue) of experimental animals and controls were pooled and used for mRNA extraction. mRNAs from two groups were used for subtractive hybridization. Clones of the subtractive library were sequenced and the obtained sequences were identified by gen bank assignment. Two clones were found to contain sequences which could be assigned to the osmolarity sensor protein envZ, showing homologies of 61 and 65%. EnvZ is an inner membrane protein in bacteria, important for osmosensing and required for porine gene regulation. It undergoes autophosphorylation and subsequently phosphorylates OmpR, which in turn binds to the porine (outer membrane protein) promoters to regulate the expression of OmpF and OmpC, major outer membrane porines. This is the first report of an osmosensing mechanism in the mammalian system, which was described in bacteria only. Furthermore, we are assigning a tentative role for taurine in the osmoregulatory process by modifying the expression of the osmoregulatory sensor protein ENVZ. PMID- 10707765 TI - Antimicrobial activity of o-carboranylalanine. AB - Functionalized polyhedral carboranes, including amino acid analogs, have unique physicochemical properties and are used as experimental anticancer agents. However, our current knowledge on their effect in nonmammalian biological systems is limited. We investigated the activity spectrum in vitro of o-carboranylalanine (o-Cba), considered to be a highly lipophilic analog of phenylalanine, against representative plant pathogenic bacteria and fungi of various taxonomic position. The antibacterial effect of o-Cba against some species was comparable to that of the widely used agricultural antibiotic, streptomycin. The sensitivity of individual bacterial species to o-Cba within the same genus varied to a greater extent than the average sensitivity of various genera. In general, this carborane containing amino acid was more toxic to Gram positive bacteria (Bacillus, Corynebacterium, Curtobacterium, Micrococcus, Rhodococcus, and Staphylococcus) than to Gram negative ones (Agrobacterium, Erwinia, Escherichia, Pseudomonas, Rhizobium, and Xanthomonas). Compared to the commercial fungicide, prochloraz, o Cba was weakly toxic against various fungi (Zygo- and Ascomycota). It was also inferior to the commercial fungicide metalaxyl in inhibiting the vegetative growth of oomyceteous plant pathogens (Pythium irregulare, Phytophthora cryptogea and Plasmopara halstedii). Against the asexual spores of P. halstedii, o-Cba, however, was over a thousandfold more active than tridemorph, a selective zoospore inhibitor fungicide. For all taxonomic groups, the observed antimicrobial effect of o-Cba could be diminished with histidine, but not with phenylalanine. In studies on healthy and mildew-infected sunflower and tobacco plants o-Cba showed neither fungicidal nor phytotoxic effects at 500 ppm. This is the first report on the biological activity spectrum of a carborane-containing amino acid. PMID- 10707766 TI - Synthesis of non proteinogenic dipeptides by asymmetric hydrogenation. AB - N-Boc protected non-proteinogenic dipeptides with D,L- and L,L-configuration were prepared by catalytic asymmetric hydrogenation of the corresponding dehydrophenylalanyl-(L)-phenylalanine derivatives. The configuration of the new stereogenic centre depends first of all on the catalyst configuration and is less influenced by the substrate configuration. Diastereomeric excesses in the range of 80-96% de could be increased up to 99% by recrystallization. Analytical data of selected new compounds are given. PMID- 10707767 TI - Production of polyclonal antibodies towards the immunodetection of insecticide phosalone. AB - Hapten synthesis for the production of specific insecticide phosalone polyclonal antibodies was carried out starting from an intermediate of the phosalone synthesis, 6-chloro-2-benzoxazolone 1. Two haptens containing different spacers have been prepared: N-5-carboxypentyl-6-chloro-2-benzoxazolone 7 and N-(2-oxo-3 aza-5-carboxypentyl)-6-chloro-2-benzoxazolone 12. Each of these two haptens conjugated to bovine serum albumine (BSA) was used to immunize four rabbits. Immunoassays of phosalone were performed with ELISA using solid-phase bound hapten thyroglobulin conjugate and horseradish peroxidase labelled goat antirabbit IgG. The more sensitive response was observed when the antiserum obtained from the rabbit immunized with the hapten-BSA conjugate containing the N 2-oxo-3-aza-5-carboxypentyl spacer was in competition with the same hapten coupled to thyroglobulin. An identical response was obtained under the same conditions when using benzoxazolone instead of phosalone as competitor, showing a good recognition of the specific aromatic part of the organophosphate insecticide phosalone. Reduction of coating conjugate concentration (from 2 to 0.05 micrograms/well) and also the use of heterologous coating protein instead of homologous did improve the sensitivity, resulting in a concentration of phosalone required to inhibit binding by 50% of 2 mg/l and a detection limit of 0.02 mg/l. PMID- 10707768 TI - Inhibitory effect of arginine-derivatives from ginseng extract and basic amino acids on protein-arginine N-methyltransferase. AB - Protein-arginine N-methyltransferase (protein methylase I) catalyzes methylation of arginyl residues on substrate protein posttranslationally utilizing S-adenosyl L-methionine as the methyl donor and yields NG-methylarginine residues. Arginyl fructose and arginyl-fructosyl-glucose from Korean red ginseng were found to inhibit protein methylase I activity in vitro. This inhibitory activity was shown to be due to arginyl moiety in the molecules, rather than that of carbohydrates. Several basic amino acids as well as polyamines were also found to inhibit protein methylase I activity. Interestingly, the intensity of the inhibitory activity was correlated with the number of amino-group in polyamines, thus, in the order of spermine > spermidine > putrescine > agmatine-sulfate, with IC50 at approximately 15 mM, 25 mM, 35 mM, and 50 mM, respectively. On the other hand, neutral amino acids or NaCl did not inhibit the enzyme activity. Lineweaver-Burk plot analysis of the protein methylase I activity in the presence of arginine and spermidine indicated that the inhibition was competitive in nature in respect to protein substrate, with the Ki values of 24.8 mM and 11.5 mM, respectively. PMID- 10707770 TI - Proceedings of the 7th International Conference on Malignant Lymphoma. Lugano, Switzerland, 2-5 June 1999. PMID- 10707769 TI - Where does indolylacrylic acid come from? AB - In addition to the main catabolic routes of tryptophan (Trp), there exist minor and less thoroughly investigated pathways; one of these leads to indolylacrylic acid (IAcrA). IAcrA is a plant growth hormone, whereas its biological role in animals is still obscure, as is the way and site where it is formed in the organism. A two-stage production is likely: Intestinal microorganisms catabolize Trp to indole derivatives which are then absorbed and converted to IAcrA and its glycine conjugate, indolylacryloylglycine (IAcrGly). Our finding of IAcrGly in the urine of proven germ-free piglets points to the possibility that Trp can be converted to IAcrA without the intervention of intestinal microorganisms. Seasonal and age variations, influence of light and connection with photodermatoses have been reported. Besides other pathological conditions the differences in IAcrGly excretion relative to normal controls were especially pronounced in some myopathies, namely in boys with Duchenne muscular dystrophy. PMID- 10707771 TI - Lymphoma classification--from controversy to consensus: the R.E.A.L. and WHO Classification of lymphoid neoplasms. AB - BACKGROUND: Controversy in lymphoma classification dates back to the first attempts to formulate such classifications. Over the years, much of this controversy arose from the assumption that there had to be a single guiding principle--a 'gold standard'--for classification, and from the existence of multiple different classifications. DESIGN: The International Lymphoma Study Group (I.L.S.G.) developed a consensus list of lymphoid neoplasms, which was published in 1994 as the 'Revised European-American Classification of Lymphoid Neoplasms' (R.E.A.L.). The classification is based on the principle that a classification is a list of 'real' disease entities, which are defined by a combination of morphology, immunophenotype, genetic features, and clinical features. The relative importance of each of these features varies among diseases, and there is no one 'gold standard'. In some tumors morphology is paramount, in others it is immunophenotype, a specific genetic abnormality, or clinical features. An international study of 1300 patients, supported by the San Salvatore Foundation, was conducted to determine whether the R.E.A.L. Classification could be used by expert pathologists and had clinical relevance. Since 1995, the European Association of Pathologists (EAHP) and the Society for Hematopathology (SH) have been developing a new World Health Organization (WHO) Classification of hematologic malignancies, using an updated R.E.A.L. Classification for lymphomas and applying the principles of the R.E.A.L. Classification to myeloid and histiocytic neoplasms. A Clinical Advisory Committee (CAC) was formed to ensure that the WHO Classification will be useful to clinicians. RESULTS: The International Lymphoma Study showed that the R.E.A.L. Classification could be used by pathologists, with inter-observer reproducibility better than for other classifications (> 85%). Immunophenotyping was helpful in some diagnoses, but not required for many others. New entities not specifically recognized in the Working Formulation accounted for 27% of the cases. Diseases that would have been lumped together as 'low grade' or 'intermediate/high grade' in the Working Formulation showed marked differences in survival, confirming that they need to be treated as distinct entities. Clinical features such as the International Prognostic Index were also important in determining patient outcome. The WHO Clinical Advisory Committee concluded that clinical groupings of lymphoid neoplasms was neither necessary nor desirable. Patient treatment is determined by the specific type of lymphoma, with the addition of grade within the tumor type, if applicable, and clinical prognostic factors such as the International Prognostic Index (IPI). CONCLUSIONS: The experience of developing the WHO Classification has produced a new and existing degree of cooperation and communication between oncologists and pathologists from around the world, which should facilitate progress in the understanding and treatment of hematologic malignancies. PMID- 10707772 TI - EORTC classification for primary cutaneous lymphomas: a comparison with the R.E.A.L. Classification and the proposed WHO Classification. AB - Primary cutaneous lymphomas differ significantly from their nodal equivalents in clinical behaviour and prognosis, and often require a different therapeutic approach. Since currently used classification systems for non-Hodgkin lymphomas do not or insufficiently recognize the special character of these lymphomas, primary cutaneous lymphomas are not uncommonly diagnosed incorrectly, and/or treated inappropriately with unnecessarily aggressive therapies. For that reason the Cutaneous Lymphoma Group of the European Organization for Research and Treatment of Cancer (EORTC) has recently proposed a separate classification for the group of primary cutaneous lymphomas. The EORTC Classification is consistently based on a combination of clinical, histological, immunophenotypical and genetic criteria, and includes well-defined and recognizable disease entities. It contains a limited number of cutaneous T-cell lymphomas and cutaneous B-cell lymphomas, which comprise more than 95% of all primary cutaneous lymphomas. The clinical significance of this classification has been validated by long-term follow-up data of more than 800 patients with a primary cutaneous lymphoma. The basic principles of the EORTC Classification will be presented, and current controversies between the EORTC Classification on the one hand, and the R.E.A.L. Classification and the proposed WHO Classification on the other will be discussed. PMID- 10707773 TI - Cutaneous lymphomas: a proposal for a unified approach to classification using the R.E.A.L./WHO Classification. AB - BACKGROUND: The classification of cutaneous lymphomas has been controversial. The EORTC has proposed that conventional classification schemes are not suitable for cutaneous lymphomas, and that a unique classification system is required. DESIGN: The authors review the suitability of the R.E.A.L. Classification for cutaneous lymphomas, and compare it with the newly proposed EORTC system. The principles of the R.E.A.L. Classification have been adopted by the WHO committees for the classification of hematopoietic and lymphoid neoplasms. Each disease is defined as a distinct entity based on an integration of morphology, immunophenotypic and genetic features, clinical presentation and course, and normal cellular counterpart. If either primary or secondary involvement of the skin is a constant factor, this aspect is considered integral to disease definition. RESULTS: Organ specific classification schemes may impede the recognition of common features of diseases involving multiple anatomic sites. For example, cutaneous marginal zone B-cell lymphomas (formerly designated cutaneous immunocytomas) mirror the features of MALT lymphomas in other anatomic sites. While the EORTC Classification for cutaneous lymphomas attempts to emphasize certain aspects of these neoplasms of importance to dermatologists, the use of multiple classification systems is a step backward, and may lead to confusion among hematologists/oncologists, and dermatologists. Nevertheless, cutaneous lymphomas often have a more indolent natural history than nodal lymphomas, and may require different therapeutic approaches. Clinical features are an important prognostic factor and should be utilized in guiding therapy. For cutaneous lymphomas the presence or absence of systemic spread is particularly important. Additionally, the site of origin is often important in the definition of disease entities. CONCLUSIONS: Organ-specific classification schemes, such as the EORTC Classification for cutaneous lymphomas, are not required, and indeed may impede the recognition of common features of diseases involving multiple anatomic sites. A common classification system, such as the R.E.A.L./WHO Classification, should be utilized for all lymphomas, regardless of the site of origin. PMID- 10707774 TI - Follicular lymphoma: have we made any progress? AB - Follicular lymphomas are characterized by relatively long median survivals and a continuous pattern of relapse. The heterogeneity in these diseases is increasingly appreciated, leading to concerted efforts to define prognostic factors and risk-adapted strategies. The status of multiple options for treatment including interferon, fludarabine, dose intensification with autologous transplantation, therapy targeting the CD20 antigen and novel approaches is reviewed. The long natural history of follicular lymphoma requires mature data for accurate analysis. However, the achievement of molecular remission as a surrogate endpoint is under active investigation. This is an exciting era for the clinical investigation of follicular lymphoma given the large number of candidate therapies and their potential combinations and permutations. Although the goal of primary treatment remains durable remission and cure, the sequential application of effective, non-cross-resistant treatments may also result in a prolongation of median survival time. It is essential that physicians treating patients with follicular lymphoma demonstrate restraint in the application of new treatments and cooperate in the study of new therapies in carefully designed phase II and phase III trials. PMID- 10707775 TI - Diffuse large-cell lymphoma. AB - BACKGROUND: New treatments are desperately needed to improve the results obtained using CHOP chemotherapy for patients with diffuse large cell lymphoma. In order to develop successful new strategies we need to understand why prior promising therapies have failed and to develop new testable hypotheses based on our current knowledge. PATIENTS AND METHODS: The International Non-Hodgkin's Lymphoma Prognostic Factors Index has provided us with a methodology to compare the expected prognosis of patients on different clinical trials. Many prior, apparent improvements in treatment outcome can now be attributed to the inclusion of patients with better prognoses. RESULTS: Current areas of investigation include: 1) the identification of new active drugs for the treatment of lymphoma, 2) the use of colony stimulating factors to allow dose escalation of the active myelotoxic drugs, 3) the use of strategies which may overcome the problem of resistance to chemotherapy, 4) the combination of monoclonal antibodies with combination chemotherapy and 5) ablative chemotherapy with autologous stem-cell support. CONCLUSIONS: Based on all of the available data, the North American Lymphoma Intergroup has developed the hypothesis that high-intermediate and high risk patients with aggressive lymphoma who receive full course standard induction therapy will benefit form the addition of high dose therapy and has antedated a clinical trial testing that hypothesis. PMID- 10707776 TI - Burkitt lymphoma is immunophenotypically different from Burkitt-like lymphoma in young persons. AB - INTRODUCTION: Burkitt-like lymphoma (BLL) is a provisional category of B-cell lymphoma which is morphologically intermediate between Burkitt lymphoma (BL) and large B-cell lymphoma (LBCL). The clinical significance of this morphology is controversial. PATIENTS AND METHODS: We examined 41 cases of pediatric B-cell lymphoma by immunohistochemistry for proteins associated with proto-oncogenes c myc, BCL-2 and BCL-6 and a subset of cases (with adequate slides) for a proliferation-associated marker (Ki-67) and for apoptosis (Apop-Tag). Sixteen cases of BLL, thirteen cases of BL and twelve cases of LBCL were examined. RESULTS: Our results showed BCL-6 expression in 16 of 16 BLL, 4 of 13 BL, and 9 of 12 LBCL; c-myc expression in 14 of 15 BLL, 9 of 13 BL, and 12 of 12 LBCL; and BCL-2 expression in 2 of 16 BLL, 9 of 13 BL, and 6 of 12 LBCL. Mean apoptotic index for BLL was 10.3% (n = 6); for BL was 17.1% (n = 5); and for LBCL was 10.9% (n = 6). Ki-67 was diffusely reactive in all cases tested. There was a significantly higher proportion of BLL than BL which expressed BCL-6 (P = 0.0001). CONCLUSIONS: Labeling for BCL-6 distinguishes BLL from BL. It is likely that in children in North America, BLL is biologically distinct from BL and more closely resembles a subset of LBCL. PMID- 10707777 TI - Primary cerebral lymphomas: unsolved issues regarding first-line treatment, follow-up, late neurological toxicity and treatment of relapses. The FNCLCC. French Federation Nationale des Centres de Lutte contre le Cancer. AB - BACKGROUND: Primary cerebral non-Hodgkin's lymphomas (NHL) in immunocompetent patients (PCL) are located exclusively in the central nervous system, the eye, or meninges. Clinical management of these patients remains controversial. PATIENTS AND METHODS: Clinical characteristics of the patients and parameters influencing their outcome as of December 1998 were investigated and registered in a database of 226 patients treated in the French Federation of Cancer Centers between 1980 and 1995. RESULTS: Most PCL are diffuse large-cell NHL with a B phenotype. The incidence of PCL has been steadily increasing over the past 20 years in some but not all countries. The overall survival of primary cerebral lymphoma (PCL) patients in the published series, a median of 12-16 months and a five-year survival of 5%-20%, is poor. Several series have now reported long-term survivals of more than 10 years and PCL may therefore be a curable tumor in some patients. The optimal treatment of PCL is not known. Complete resection of the tumor does not improve outcome and multidisciplinary approaches combining chemotherapy and radiotherapy are now commonly used, although the superiority of combination over radiotherapy- or chemotherapy-alone has never been demonstrated in a phase III trial. The optimal chemotherapy regimen, the dose and even the usefulness of brain radiotherapy after chemotherapy are therefore still matters of debate. Recently, several authors have reported a relatively high incidence of late neurological sequelae after PCL treatment. CONCLUSIONS: The optimal treatment of PCL patients remains to be defined. Large cooperative international phase III trials are now required to define and improve the optimal treatment of PCL and reduce its sequelae. PMID- 10707778 TI - Diagnosis and treatment of transplant-related lymphoma. AB - Immunodeficiency-related B-cell disorders are seen after organ transplantation and in congenital and acquired immunodeficiency states. Post-transplant lymphoproliferative disorders (PTLD) comprise a histologic spectrum ranging from hyper-plastic appearing lesions to frank non-Hodgkin's lymphoma or multiple myeloma histology. Multiple clones may co-exist, representing a uniquely different mechanism for lymphomagenesis. The incidence varies from 1% in renal recipients to 8% in lung recipients, but can be markedly increased by the use of anti-T-cell therapies, or by T-cell depletion in bone marrow transplantation. Pre transplant EBV seronegativity increases risk to as high as 30%-50%. More than 90% of tumors are EBV-associated. Mechanisms for viral lymphomagenesis remain incompletely defined; LMP-1 may function as an oncogene and coprecipitates with TRAF, BCL-2 overexpression has also been identified. A possible direct tumorigenic effect has recently been suggested for cyclosporine. PTLD has a highly variable clinical picture, certain patterns are however seen. Reversibility of PTLD with reduction in immunosuppressives has long been recognized. Predicting reversibility has been difficult. The presence or absence of BCL-6 mutations has recently been identified as being of predictive value. Surgical resection can be curative. Cytotoxics, although problematic, can also be curative. Long term remission has been achieved with anti CD21 and CD24 antibodies; efficacy has been reported anecdotally for interferon alpha and for rituximab. In vitro expanded EBV-specific T cells have been effective as treatment and as prophylaxis in the setting of bone marrow transplantation. EBV viral load measured in blood appears to correlate with the emergence of PTLD and may facilitate prophylactic studies. PTLD is a model of immunodeficiency related EBV lymphomagenesis. Pathogenetic, therapeutic, and prophylactic insights gained from the study of PTLD are likely to be applicable to other immunodeficiency states and to EBV-related lymphoid neoplasia in general. PMID- 10707779 TI - Stem-cell transplantation for chronic lymphocytic leukemia: the 1999 perspective. AB - BACKGROUND: The use of myeloablative intensive therapy followed by autologous or allogeneic stem-cell transplantation (SCT) for treatment of chronic lymphocytic leukemia (CLL) has largely increased over the last years. DESIGN: The present overview updates the available clinical results and discusses important aspects of SCT in patients with CLL including the type of SCT (autologous vs. allogeneic), myeloablative regimens, purging, the predictive value of molecular monitoring of residual disease, and prognostic factors for the outcome of transplant approaches. RESULTS: With appropriate supportive care, autologous SCT is safe and can induce long-lasting clinical and molecular remissions, which may improve the prognosis of patients with CLL. Feasibiliy and efficacy of autologous SCT appears to be best early during the course of the disease, but it is still unclear if autotransplantation can cure the disease even in this favorable subgroup. The role of purging is still unclear. The better disease control observed after allografting appears to be due to graft-versus-leukemia activity and may allow cure in at least a subset of poor-risk patients. Due to the extraordinarily high treatment-related mortality, however, the outcome after allogeneic SCT is still inferior to that after autologous SCT. CONCLUSIONS: Autologous transplantation is a valuable treatment option for younger patients with early or sensitive poor-risk CLL. Selected patients with advanced poor-risk disease and low probability of successful auto-SCT should be considered for allografting. However, it must be kept in mind that both autologous and allogeneic stem-cell transplantation are still experimental procedures and clinical trials further elucidating their value in the treatment of patients with CLL are warranted. PMID- 10707780 TI - The role of high-dose chemotherapy in the treatment of multiple myeloma: a controversy. AB - BACKGROUND: Minimal criteria for the diagnosis of multiple myeloma are provided. Monoclonal gammopathy of undetermined significance, smoldering multiple myeloma, primary systemic amyloidosis and metastatic carcinoma must be included in the differential diagnosis. Patients with multiple myeloma should not be treated unless they have an increasing M-protein in the serum or urine, development of anemia, hypercalcemia, renal insufficiency, lytic lesions, fractures or extra medullary plasmacytomas. PATIENTS AND METHODS: This is a review of patients treated with chemotherapy, autologous stem-cell transplantation and allogeneic transplantation. RESULTS: Comparisons of melphalan and prednisone with a variety of combinations of therapeutic agents produces a higher response rate than with melphalan and prednisone but no significant difference in overall survival. Autologous stem-cell transplantation produces a higher response rate and some prolongation of survival but is not curative. Allogeneic transplantation is associated with a mortality of 40% and is not curative. CONCLUSIONS: If the patient is younger than 70 years, the physician should consider the possibility of an autologous peripheral blood stem-cell transplant. Ideally, this should be done as part of a prospective study. Hematopoietic stem cells are damaged by alkylating agents so they must be collected before these agents are given. Autologous stem-cell transplantation does not produce a cure and most patients will relapse. The appropriate timing of an autologous stem-cell transplant has not been ascertained. Hopefully, better preparative regimens and the removal of contaminated tumor cells from the peripheral blood will make an autologous transplant more effective. Another major question is whether double (tandem) transplants are superior to a single autologous stem-cell transplant. A current French Myeloma Group Study randomized study should answer this question. Allogeneic transplantation for multiple myeloma must be made safer because the transplant-related mortality is 40%. The relapse of multiple myeloma following allogeneic transplant is a major problem and consequently the preparative regimens must be improved. The infusion of donor lymphocytes following relapse after an allogeneic transplant is useful. New approaches with immunologic aspects including the use of dendritic cells and vaccines are of potential importance for the future. PMID- 10707781 TI - Allografting for indolent lymphoid neoplasms. AB - BACKGROUND: Allogeneic bone marrow transplantation (BMT) has been used in patients with low-grade lymphoma (LGL) and chronic lymphocytic leukemia (CLL) with the goal of achieving long-term disease-free survival. PATIENTS AND METHODS: Twenty-nine patients with these diagnoses (LGL = 19, CLL = 10) received allogeneic BMT between September 1995 and January 1999. Median age was 42 (range 20-52) years. Twenty-three of twenty-nine patients (79%) were Ann Arbor or Rai stage IV at the time of transplant; twenty-four (83%) had never achieved complete remission (CR). Donor source was HLA-matched sibling (20), unrelated (8) and syngeneic (1). RESULTS: Seventeen patients are currently alive, a median of 29 months (range 1-85) post-BMT with a median KPS of 90%. Twenty-three of twenty seven evaluable patients (85%) achieved CR post-BMT. Six patients had refractory/recurrent disease. Death occurred related to transplant complications in eight patients and underlying disease in four. Overall and event-free survival for the whole group is 51% and 44%, respectively. CONCLUSIONS: Allogeneic BMT for young patients with advanced stage LGL or CLL is a feasible strategy that can result in achievement of long-term disease-free survival. PMID- 10707782 TI - Distribution of various subtypes of non-Hodgkin's lymphoma in India: a study of 2773 lymphomas using R.E.A.L. and WHO Classifications. AB - INTRODUCTION: The distribution of the major subtypes of non-Hodgkin's lymphoma (NHL) differs across geographic regions. This is the first study from India that has incorporated immunophenotypic findings while investigating the distribution of NHL subtypes. PATIENTS AND METHODS: All cases diagnosed as NHL between January 1995 and June 1998 in the Department of Pathology and in the Lymphoma Registry, Tata Memorial Hospital, Bombay, were selected for the study. The cases were reviewed by three pathologists and diagnostic problems were discussed by a panel of pathologists with a special interest in lymph node pathology. Of a total of 2831 cases, the diagnosis of NHL was accepted in 2773 cases. RESULTS: B-cell lymphomas formed 79.1% of the NHLs, whereas T-cell lymphomas formed 16.2% of the total. Diffuse large B-cell lymphoma was the most common subtype (34% of all NHLs). Follicular centre-cell lymphomas, B-cell small lymphocytic lymphoma, mantle-cell lymphoma, and marginal zone B-cell lymphomas (including MALT lymphomas) amounted to 12.6%, 5.7%, 3.4%, and 8.2%, respectively. Among the T cell lymphomas, T-cell lymphoblastic lymphoma, anaplastic large-cell lymphomas of T/null-cell type, and other nodal peripheral T-cell lymphomas accounted for 6%, 4.3%, and 2.9% of all cases, respectively. CONCLUSIONS: The distribution of NHL subtypes in India shows important differences with those from the rest of the world. Follicular lymphoma and mantle-cell lymphoma are less common in India compared to Europe and the USA. Peripheral T-cell lymphomas and T/NK-cell lymphomas of nasal and nasal types, which are common in many other Asian countries, are also less prevalent. T-cell lymphoblastic lymphoma and anaplastic large T/null cell lymphoma are more prevalent in India. PMID- 10707783 TI - Non-Hodgkin's lymphoma and occupational exposure to chemicals in Canada. Canadian Cancer Registries Epidemiology Research Group. AB - BACKGROUND: The incidence of non-Hodgkin's lymphoma (NHL) has been increasing in Canada. This study assessed the effect of occupational exposure to specific chemicals on the risk of NHL. PATIENTS AND METHODS: Mailed questionnaires were used to obtain data on 1469 newly diagnosed, histologically confirmed NHL cases and 5073 population controls between 1994 and 1997 in eight Canadian provinces. Data was collected on socioeconomic status, life-style, diet, occupation, and years of exposure to any of 17 chemicals. Odds ratios (OR) and 95% confidence intervals (95% CI) were derived by logistic regression. RESULTS: The study found an increased risk of NHL among males exposed to benzidine, mineral, cutting, or lubricating oil, pesticides, and herbicides. Compared with non-exposure to each specific chemical, the adjusted ORs were 1.9 (95% CI: 1.1-3.4) for benzidine, 1.3 (95% CI: 1.0-1.5) for mineral, cutting, or lubricating oil, 1.3 (95% CI: 1.0-1.6) for herbicides, and 1.3 (95% CI: 1.0-1.6) for pesticides. Excess risk of NHL among females was associated with exposure to pesticides and wood dust. ORs increased with increasing exposure in years to benzidine and herbicides for males and with increasing exposure years to wood dust for females. These trends were statistically significant (P < 0.05). CONCLUSIONS: The findings in this study suggest that occupational exposure to specific chemicals plays an important role in the development of NHL in Canada. PMID- 10707784 TI - Translocation t(14;18) in healthy individuals: preliminary study of its association with family history and agricultural exposure. AB - BACKGROUND: The t(14;18) translocation, present in 90% of follicular non Hodgkin's lymphomas (NHL), has been found to exist in low levels in healthy persons. Its clinical/prognostic significance in healthy populations is unknown, and risk factors for its development have not been determined. Our objectives were to assess the prevalence of t(14;18) in individuals without NHL, comparing residents of agricultural settlements (kibbutzim) with city dwellers, as well as first degree relatives of NHL cases. PATIENTS AND METHODS: Residents of kibbutzim and members of two control groups: 1) Jerusalem residents--randomly selected hospital administrative workers and 2) first degree family members of lymphoma patients were interviewed extensively regarding exposures and had blood drawn for t(14;18) determination. The translocation was detected after B-cell purification of blood samples with CD-19 microbeads (Mini-Macs) using nested PCR. The method detects the translocation in a BCL2 positive cell line after dilutions of up to 1:10(5) with normal peripheral blood lymphocytes. RESULTS: Nineteen of two hundred thirty healthy individuals (8.3%) tested were found to be positive for t(14;18). No statistically significant differences in the prevalence of t(14;18) were detected among the rural and urban populations. Five of thirty-four (11.9%) family members tested positive for t(14;18). No age or sex differences between t(14;18) positive and negative individuals were found. No significant association with exposure to specific agricultural or other chemicals was found. CONCLUSIONS: The presence of the t(14;18) translocation in healthy individuals was not associated with agricultural residence in this preliminary study. Whether relatives of patients with NHL are at increased risk will require further study in larger populations. Specific exposures affecting the onset of this translocation have not been ruled out. The significance of this translocation in healthy individuals remains unknown. PMID- 10707785 TI - Treatment of Hodgkin's disease: results and current concepts of the German Hodgkin's Lymphoma Study Group. AB - BACKGROUND: Treatment strategies in Hodgkin's disease (HD) are changing fundamentally over the last decades. Both radiotherapy and combination chemotherapy are effective treatment modalities. However, the optimal choice of treatment or combinations of treatment is still debated for different prognostic groups. PATIENTS AND METHODS: The German Hodgkin's Lymphoma Study Group (GHSG) initiated randomized clinical trials since 1978. Over the past 20 years, more than 6000 patients with HD in all stages were randomized, treated and followed by the GHSG. Patients are now being recruited from more than 300 clinical centers. RESULTS: As a consequence of different clinical trials, it is now the policy of the GHSG to tailor treatment to the individual risk of patients, giving favorable patients less intensive and less toxic therapy than unfavorable patients. The treatment for early and intermediate stage HD becomes quite similar with few cycles of polychemotherapy followed by involved field irradiation. In advanced stage HD, the introduction of dose intensified chemotherapy (BEACOPP), has improved treatment results and thus will substitute the MOPP or ABVD regimens. CONCLUSIONS: Although most of the patients with HD will be cured by modern treatment strategies, several questions are still subjects of ongoing clinical trials: 1) which chemotherapy regimen in which quantity will be the best with respect to efficacy and toxicity and 2) which dose and field size of radiotherapy is adequate within the combined modality. PMID- 10707786 TI - A randomized British National Lymphoma Investigation trial of CHOP vs. a weekly multi-agent regimen (PACEBOM) in patients with histologically aggressive non Hodgkin's lymphoma. AB - BACKGROUND: Between 1987 and 1991, the British National Lymphoma Investigation randomized 459 patients with non-Hodgkin's lymphoma with a large-cell component to either CHOP or the PACEBOM regimen. PATIENTS AND METHODS: Four hundred fifty nine eligible patients were included in this trial, four hundred one with diffuse large-cell lymphoma and fifty-eight with diffuse mixed-cell lymphoma according to the Working Formulation. Two hundred twenty-six patients were randomized to receive CHOP and two hundred thirty-three to receive PACEBOM. The two arms of the trial were well balanced for all potential prognostic factors. RESULTS: The complete remission rate with PACEBOM was 64% and with CHOP 57% (NS). At eight years, the actuarial cause specific survival (CSS) in the PACEBOM arm is 59% compared to 49% in the CHOP arm (P = 0.09). Patients < 50 years of age fared significantly better in the PACEBOM arm both for CSS and overall survival (P = 0.002) and the CSS was also significantly improved in stage IV disease (P = 0.02). CONCLUSIONS: The mature data from this trial suggest that an etoposide containing multi-agent weekly regimen can be superior to CHOP. PMID- 10707787 TI - The CUP trial: a randomized study analyzing the efficacy of high dose therapy and purging in low-grade non-Hodgkin's lymphoma (NHL). AB - BACKGROUND: The CUP trial was initiated to analyze the value of high-dose therapy and stem-cell transplantation and purging in patients with relapsed chemosensitive follicular NHL. PATIENTS AND METHODS: After three cycles of chemotherapy responsive patients were randomized to either three more cycles of the same chemotherapy (C), high-dose therapy followed by autologous unpurged (U) or purged (P) stem-cell transplantation. Purging was performed using a cocktail of monoclonals. Pretransplant conditioning consisted of cyclophosphamide (60 mg/kg x 2) and total body irradiation. RESULTS: Of the 140 patients registered from 26 centers in Europe, 89 fulfilled the criteria for randomization (C: 24, U: 33 and P: 32). Reasons for failure to randomize were: no response (28), persistent marrow infiltration (4), patient refusal (7), other (7), no data (5). With the current follow up (median 26 months from randomization) 16 (66%) in C are known to have progressed or relapsed, in contrast to 13 (39%) of U and 12 (37%) of the P patients (P-value 0.002). Overall survival is premature with the current available data. CONCLUSIONS: Patients in U and P arms had higher progression/relapse-free survival rate. There are some suggestions of some improvement in overall survival rate. PMID- 10707788 TI - A T-cell epitope determined with random peptide libraries and combinatorial peptide chemistry stimulates T cells specific for cutaneous T-cell lymphoma. AB - BACKGROUND: Mycosis fungoides is the most frequent T-cell lymphoma of the skin. Despite numerous attempts, no tumour antigens have yet been identified. Only in one case has an idiotype-derived peptide been found to trigger CTL of the respective patient. The identification of natural antigens requires the cultivation of large amounts of tumour cells in vitro, which has been possible in two exceptional cases. The identification of synthetic epitopes for tumour specific CTL with random peptide libraries can overcome this limitation and is a powerful tool for application in the development of immune therapies for a wide range of patients. MATERIALS AND METHODS: The critical amino acids for the construction of epitopes for the CTCL-specific CTL clone My-La CTL were determined with synthetic peptide libraries in positional scanning OX8 format in a standard 61chromium release assay. Sixteen different peptides could be synthesized from the combinatoric of these amino acids with the canonical anchor amino acids for MHC binding. These peptides were tested for their capacity to stimulate My-La CTL and PBMC of an HLA-matched CTCL patient. RESULTS: A synthetic epitope could be identified for My-La CTL, which was recognized in a HLA restricted manner. The response towards this epitope was comparable to the response towards their natural target My-La. Using these synthetic epitopes, T cells of a HLA-matched patient could be induced in vitro and led to the establishment of different cell lines and clones. Some of these lines recognized the peptides as well as the allogenic but HLA-matched tumour cell line My-La, indicating that they are specific for a naturally expressed tumour antigen. CONCLUSIONS: The identification of synthetic epitopes for tumour-specific CTL clones can be used for the development of vaccines for immune therapies of cancer; such peptides can be applied inter-individually. Synthetic epitopes must not correspond to the natural ones, but they can be even more potent as stimulation of specific T cells and can be fine-tuned to increase the success of the therapy. PMID- 10707789 TI - T-cell receptors for cytokines: targets for immunotherapy of leukemia/lymphoma. AB - BACKGROUND: Cytokine receptors are exceptionally valuable targets for immunotherapy. For example, the high affinity IL-2 receptor is expressed by abnormal T cells in patients with certain lymphoid malignancies or autoimmune disorders and in individuals rejecting allografts whereas it is not expressed by normal resting cells. DESIGN: To exploit this difference in receptor expression in normal resting cells and leukemic cells we have introduced different forms of IL-2 receptor directed therapy including an unmodified murine antibody to the alpha subunit of the IL-2 receptor (anti-Tac), humanized anti-Tac as well as this antibody armed with truncated Pseudomonas exotoxin or alpha- and beta-emitting radionuclides (e.g., 211At and 90Y). In particular, unmodified murine anti-Tac was used in the therapy of HTLV-I-associated adult T-cell leukemia (ATL). RESULTS: Six of nineteen patients treated with this antibody underwent a partial (four) or complete (two) remission. In a subsequent clinical trial involving anti Tac armed with 90Y over 50% of the patients with ATL treated underwent a partial or complete remission. CONCLUSIONS: New agents under development include humanized antibodies directed toward shared cytokine receptors such as IL-2/15R beta used by both IL-2 and IL-15 as well as to a shared signal transduction element Jak3 utilized by the T-cell stimulatory cytokines IL-2, IL-4, IL-7, IL-9 and IL-15. Thus our emerging understanding of cytokine receptors and their signaling pathways taken in conjunction with the ability to produce humanized antibodies armed with radionuclides or toxins are providing novel perspectives for the treatment of leukemia and lymphoma. PMID- 10707790 TI - Treatment of multiple myeloma by antibody mediated immunotherapy and induction of myeloma selective antigens. AB - BACKGROUND: In view of the successful use of serotherapy in many B-cell malignancies, we and others have sought to identify tumor selective antigens for the serotherapy of plasma cell dyscrasias (PCD) including multiple myeloma (MM), and Waldenstrom's macroglobulinemia (WM). We recently identified Muc-1 core protein as a MM selective antigen. Though Muc-1 core protein is abundantly expressed on most MM plasma cells, expression of this antigen can be absent, or weak on some plasma cells which could potentially result in the selection of Muc 1 core protein negative clones following serotherapy of PCD. In addition to Muc-1 core protein, we have also been examining the use of CD20 directed serotherapy for PCD. DESIGN: As part of these efforts, we recently initiated a phase II clinical trial examining the use of Rituximab (Rituxan, MabThera) as a single agent in MM patients; as well several WM patients have been treated with Rituximab at our Institutions. RESULTS: In previous studies, we have shown that CD20 is abundantly expressed on the plasma cells of most WM patients; in contrast, CD20 is expressed on plasma cells from a minority of MM patients, and in these patients expression of CD20 can be weak or heterogeneous with both CD20+ and CD20- plasma cells present. As such, we have sought out clinically useful inducers of Muc-1 core protein, and of CD20 on malignant plasma cells. CONCLUSIONS: These efforts resulted in the identification of dexamethasone (Dex) as a potent inducer of Muc-1 core protein on MM plasma cells, and interferon gamma (IFN-gamma) as a potent inducer of CD20 on MM plasma cells and B-cells. Importantly, these agents induced their respective antigens at pharmacologically achievable doses. PMID- 10707791 TI - Humanized anti-CD20 monoclonal antibody (Rituximab) in post transplant B lymphoproliferative disorder: a retrospective analysis on 32 patients. AB - BACKGROUND: B-lymphoproliferative post-transplant disorder (BLPD) is a severe complication of organ and bone marrow transplantation. The reduction of immuno suppressive therapy or surgery for localized disease may cure some BLPDs. Other therapeutic approaches such as chemotherapy and antiviral drugs are toxic and of limited efficacy. Adoptive immunotherapy with donor T-cell infusions has yielded promising results but is, at the present time, easily applicable only in bone marrow-transplanted patients. Anti-B-cell Murine monoclonal antibodies (MoAbs) have proven effective but are no longer available for human use. We report the activity of a humanized anti CD 20 Mo Ab (Rituximab-MABTHERA Roche) in 32 episodes of BLPD treated in 14 French centers. PATIENTS AND METHODS: Between November 1997 and September 1998, 32 patients were diagnosed with BLPD. Twenty six patients had undergone solid organ transplants (liver 8, kidney 8, heart 4, lung 3, heart lung 1, kidney-pancreas 1, liver-kidney 1) and six patients had received bone marrow transplantations. The median age of the patients was 34 years (3-67 years) and the median delay between graft and tumor 5 months (1-156 months). In organ recipients, tumors were classified as polymorphic and monomorphic in 10 and 15 cases, respectively; 4 of 6 bone marrow transplant recipients were treated without pathology documentation because of a rise in EBV load, fever and lymph node enlargement. Tumors were associated with EBV in 22 of 26 tested cases. Rituximab was used as first-line therapy in 30 patients (after reduction of immunosuppressive treatment in 27 patients) and as salvage therapy in 2 patients (after failure of chemotherapy). The median time from diagnosis of BLPD to treatment with Rituximab was 14 days (1-110 days). Two patients received eight infusions, twenty-six patients four infusions, one patient three infusions and three patients two infusions of 375 mg/m2. RESULTS: The tolerance of rituximab was good. The overall response rate was 69%, with 20 complete responses and 2 partial responses. In solid organ transplant the response rate was 65% (15 CR and 2 PR) while it was 83% in bone marrow-transplanted patients (5 CR). With a median follow-up of 8 months (1-16 months) 24 patients are still alive. The one year projected survival is 73%. Of the 22 patients who achieved response, 15 patients (11 solid organ transplant and 4 bone marrow transplant) are alive with no evidence of disease, 4 patients relapsed a median of 7 months (3-10 months) after treatment and 3 died while in CR of concurrent diseases. Of the 10 patients who did not respond to Rituximab 5 are alive with no evidence of disease after salvage therapy. CONCLUSIONS: The use of rituximab appears to be a safe and relatively efficient therapy in BLPDs. The results need to be confirmed in a prospective multicentric trial. PMID- 10707792 TI - Treatment of mantle-cell lymphoma with Rituximab (chimeric monoclonal anti-CD20 antibody): analysis of factors associated with response. AB - BACKGROUND: A retrospective analysis was performed to delineate the factors associated with response, and to determine the duration of response, in 87 patients with CD20-positive mantle-cell lymphoma (MCL) treated with Rituximab (chimeric monoclonal anti-CD20 antibody) in two prior studies. PATIENTS AND METHODS: Patients with newly-diagnosed MCL (MCL1, n = 37), and previously-treated MCL (MCL2, n = 50), received single-agent Rituximab, in the context of two multicentre clinical studies using different schedules and doses, conducted in 1996 and 1997. A follow-up analysis was performed at the end of 1998, including all 81 patients who completed therapy. Statistical modeling of factors associated with response was performed using ordered logistic regression. The duration of complete (CR) and partial response (PR), and the time to disease progression (TTP), were also derived. RESULTS: The overall response rate (RR) was 34% (30 of 87) (81 evaluable patients, RR 37%; CR 14%), and was equivalent for MCL1 and MCL2. On univariate analysis, elevated LDH (P = 0.004); prior therapy with alkylating agents (P = 0.01) or fludarabine phosphate (P = 0.04); WHO performance status = 2 (P = 0.02); MCL2 refractory to last prior therapy (P = 0.04); and splenomegaly (P = 0.04), each at the time of treatment with Rituximab, were significantly associated with a lower RR. On multivariate analysis, only LDH (P = 0.007) and prior alkylating agents (P = 0.03) retained statistical significance. At a median follow-up of 1.4 years, the median TTP was 7 months. The median duration of response was one year, and was significantly longer for patients achieving CR vs. PR (P = 0.04). CONCLUSIONS: Rituximab is active in MCL, and can induce complete responses in a minority of patients. Elevated LDH at the time of therapy, and prior therapy with alkylating agents, are associated with a significantly lower RR. The duration of response of one year is similar to that previously reported in follicular lymphoma. PMID- 10707794 TI - The ATM gene in the pathogenesis of mantle-cell lymphoma. AB - BACKGROUND: Mantle-cell lymphoma (MCL) is genetically characterized by the translocation t(11;14)(q13;q32) leading to an overexpression of cyclin-D1, but additional chromosomal abnormalities appear to be required for MCL pathogenesis. PATIENTS AND METHODS: Deletions involving chromosome 11q, which were recently found as recurrent aberrations in MCL, were analyzed at the molecular level in a series of 81 MCL by fluorescence in situ hybridization (FISH) with probes from a contiguous set of yeast artificial chromosomes (YACs) spanning bands 11q14-q24. RESULTS AND CONCLUSIONS: Loss of chromosome 11 material was observed in 37 of the 81 MCL cases (46%). The consensus deletion comprised YAC 801e11 containing the ATM gene. The minimal region of loss was further narrowed with P1-derived artificial chromosome (PAC) probes. This allowed the identification of a deletion confined to the genomic region of ATM, which, together with intragenic mutations found in the coding sequence, suggests a role of ATM as a tumor suppressor gene in MCL. PMID- 10707793 TI - The effect of Rituximab on patients with follicular and mantle-cell lymphoma. Swiss Group for Clinical Cancer Research (SAKK). AB - BACKGROUND: Clinical activity of the anti CD-20 monoclonal antibody Rituximab has been reported in patients with follicular lymphoma (FL) and mantle-cell lymphoma (MCL). PATIENTS AND METHODS: 120 patients with bi-dimensionally measurable FL or MCL (R.E.A.L. Classification) were treated with Rituximab 375 mg/m2/week for 4 weeks. A central pathology review confirmed the diagnosis of FL in 76 of 78 and of MCL in 39 of 42 cases. The response was evaluated after 8 weeks and confirmed after 12 weeks from the start of treatment. RESULTS: The toxicity of the treatment was, as expected, grade 1-2 fever and rigors during the first infusion and mild asthenia during the treatment period. Serious adverse events, probably or possibly related to the study treatment, included four deaths (3 of cardiac origin, 1 caused by P. carinii pneumonia) and 10 further nonfatal cases, including a permanent agranulocytosis and one case of heart failure. Response rate at week 12 was 52% for FL and 22% for MCL. After treatment, the BCL-2 rearrangement disappeared in 15 of 29 blood but only in 5 of 23 bone marrow samples; BCL-1 disappeared in 5 of 12 blood and 0 of 7 bone marrow specimens, as determined by PCR. CONCLUSIONS: Rituximab is an active agent for the treatment of FL, while its efficacy is modest in MCL. The effect in reducing minimal residual disease is more pronounced on the blood than it is on the bone marrow. PMID- 10707795 TI - Recurrent immunoglobulin gene translocations identify distinct molecular subtypes of myeloma. AB - BACKGROUND: Chromosome translocations involving the immunoglobulin heavy chain gene (IgH) on 14q32 are a seminal event in the pathogenesis of many B-cell malignancies. Since myeloma is a post-germinal center tumor of mature, isotype switched plasma cells, we hypothesized that 14q32 translocations would usually involve IgH switch regions. MATERIALS AND METHODS: We analyzed a panel of 21 human myeloma cell lines using a Southern blot assay to detect illegitimate rearrangements involving the switch regions. We then cloned the breakpoints, developed probes for FISH analysis, and characterized the oncogenes dysregulated by the translocations. RESULTS: Only half of the cell lines demonstrated a 14q32 abnormality by conventional karyotypic analysis, but we were able to identify translocations involving IgH switch regions in 15 of 21 lines, including all of the lines in which a 14q32 translocations was not identified by conventional karyotypic analysis. Six cell lines have an Ig translocation involving 11q13 with overexpression of cyclin D1. Six cell lines have an Ig translocation involving 16q23 with overexpression of c-maf. Five lines have an Ig translocations involving 4p16 with overexpression of FGFR3 and a novel gene, MMSET. The 4p16 breakpoints occur within the 5' introns of MMSET, and are associated with IgH MMSET hybrid mRNA transcripts. The remaining five cell lines have translocations involving other loci, including: 6p25 (MUM1), 8q24 (c-myc), and 21q22 (?AML1). CONCLUSIONS: Recurrent Ig translocations identify at least three distinct molecular subtypes of myeloma. Our long-term goal is to determine if there are phenotypic, prognostic and therapeutic differences associated with these molecular subtypes. PMID- 10707796 TI - Molecular response assessed by PCR is the most important factor predicting failure-free survival in indolent follicular lymphoma: update of the MDACC series. AB - BACKGROUND: We have observed that molecular response, as defined by a PCR negative status during the first year of therapy, along with beta 2-microglobulin (beta 2M), was the most important variable associated with failure-free survival (FFS) in follicular lymphoma (FL). Herein, we present an update of the previously published MDACC series. PATIENTS AND METHODS: A total of 116 patients (male:female ratio 64:52; median age: 52 years) with indolent FL and BCL-2 rearrangement (at MBR or mcr breakpoints) assessable in peripheral blood (pb) by PCR prior to treatment, and with two or more PCR determinations during the first year, were selected for the present study. RESULTS: Of the 116 patients, 4 who presented with progression and 1 who died of unrelated causes during the first year were excluded from the landmark analysis. One hundred patients (86%) achieved clinical CR and 80 (69%) achieved a negative PCR status within first year. Median FFS was 6.4 years. Five-year FFS was 73% and 28% for molecular responders and nonresponders, respectively (P = 0.001). In spite of this strikingly higher FFS favoring molecular responders, no clearcut plateau was evident in this group. Molecular response assessed in pb (P = 0.001) and serum beta 2M (P < 0.001) were the most important factors to predict FFS in the multivariate analysis. In the subset of patients with normal beta 2M and molecular CR, there was a trend for a plateau in the FFS curve. No significant difference between the groups has been observed so far in terms of survival. CONCLUSIONS: Molecular response assessed in pb using a PCR technique is, along with beta 2M, the most important factor to predict FFS in FL. PMID- 10707797 TI - Non-Hodgkin's lymphoma in pediatric patients with chromosomal breakage syndromes (AT and NBS): experience from the BFM trials. AB - BACKGROUND: Lymphoma and leukemia are the commonest malignant diseases in patients with chromosomal breakage syndromes and immunodeficiency (Ataxia teleangiectasia (AT) and Nijmegen breakage syndrome (NBS)). With improved management of infections, malignant disease is more frequently diagnosed and has become one of the commonest causes of death in pediatric AT and NBS. PATIENTS AND METHODS: In three consecutive multicenter therapy trials for pediatric non Hodgkin's lymphoma (NHL) (NHL-BFM), 1569 patients with newly diagnosed NHL have been registered between 1986 and 1997. Nine patients with AT (n = 5) and NBS (n = 4) were identified and analysed. RESULTS: Median age of patients with AT and NBS at diagnosis of NHL was nine years. NHL-entities differed from non-AT/NBS patients: diffuse large B-cell lymphomas, n = 7 (78%); ALCL, n = 1; lymphoblastic T-cell lymphoma, n = 1. Cervical nodes, paranasal sinuses and epipharynx were the sites most frequently involved. Stages were: I and II in three patients, III in five and IV in one patient. All patients received polychemotherapy according to tumor-entity and stage, none received radiation. Dose reductions according to individual tolerance concerned mainly ethotrexate, alkylating agents and epipodophyllotoxines. One patient died of toxic complications, two patients relapsed and died, one patient suffered from second malignancy. Five of nine patients are in 1. CCR after a median follow-up of five years. CONCLUSIONS: Patients with AT and NBS suffer from rare entities of pediatric NHL. Curative treatment is possible and should be attempted. Intensity of therapy should be adjusted to individual risk factors and tolerance. Alkylating agents, epipodophyllotoxines should be omitted, dose of MTX should be limited to 1 g/m2. Further cooperative trials using standardized approaches are required. PMID- 10707798 TI - 18-FDG-PET for the assessment of residual masses on CT following treatment of lymphomas. AB - BACKGROUND: The problem of residual masses on post-treatment CT scans is a continuing dilemma for the oncologist treating malignant lymphomas. These masses may contain active disease or represent only necrotic tumour which continues to shrink without further treatment or post-treatment fibrosis which remains stable on continued follow-up. 18-FDG-PET offers a novel metabolic imaging modality, which can differentiate malignant from benign tissue on the basis of increased glycolytic activity. PATIENTS AND METHODS: Thirty-two patients (15 with Hodgkin's disease (HD) and 17 with aggressive histology non-Hodgkin's lymphoma (NHL)) who had residual masses on their post-treatment CT scans underwent 18-FDG-PET. The post-treatment CT and PET scans were compared and the accuracy of the 18-FDG-PET in assessing residual masses was evaluated using clinical and pathological follow up data. RESULTS: Nine patients had positive post-treatment 18-FDG-PET, eight (89%) of whom have relapsed. Twenty-three patients had negative post-treatment PET with only two relapses in this group. The 2 patients who relapsed had aggressive NHL while none of the 11 HD patients with negative PET relapsed. The median follow-up of patients in continued complete remission is 38 months. CONCLUSIONS: 18-FDG-PET can differentiate between residual masses containing viable lymphoma where further treatment will be required to achieve cure and those representing ablated disease, where unnecessary treatment and additional morbidity may be avoided. PMID- 10707799 TI - [Ketolides--erythromycin derivatives with activity against macrolide-resistant bacteria]. PMID- 10707800 TI - [The effect of heat shock on the formation and composition of actinomycins]. AB - The effect of various conditions of heat shock on production of actinomycins by Streptomyces chrysomallus 2 and their composition was studied. The actinomycin biosynthesis was shown to be the function of the growing mycelium and changed in accordance with changes in the volume of the mycelium and its morphological features after heat shock at various suboptimal temperatures. The temperature shock had a specific action on the antibiotic synthesis: the index of the actinomycin maximum quantity increased after the heat shock at 35 and 38 degrees C and lowered more sharply than that of the biomass volume after the heat shock at the temperatures of 40, 42, 45 and 50 degrees C for 1 hour. After the shock at 38 degrees C the component composition of the actinomycin complex did not significantly change while with addition of exogenic amino acids such as L valine, L-leucine and L-isoleucine the shock effect on the component composition of the actinomycin complex was marked. PMID- 10707801 TI - [Calcium ions and the differentiation of Streptomyces hygroscopicus 155, a producer of an antibiotic complex]. AB - The effect of Ca2+ on differentiation of Streptomyces hygroscopicus 155 and its inactive variant 155-0 was studied. Addition of Ca2+ to the medium induced formation of the aerial mycelium in the inactive variant and accelerated formation of the aerial mycelium in the parent strain. The inhibitory effect of EGTA, verapamil, nifedipin, chlorpromazine and dilthiazeme on the aerial mycelium formation demonstrated the physiological role of Ca2+ in the process. Addition of pandavir (nigericin) and azalomycin B, the antibiotics produced by the streptomycete, induced formation of the aerial mycelium in the inactive variant. The effect was higher in the presence of Ca2+. Streptomyces hygroscopicus 155 and its inactive variant synthesized a proteolytic complex containing metalloproteases and trypsin-like proteases. The total proteolytic activity of the inactive variant was lower than that of the parent strain. Addition of Ca2+ to the medium stimulated their proteolytic activity. The inducing action of the antibiotics produced by the parent strain on differentiation of S.hygroscopicus 155-0 and the increase of their action in the presence of Ca2+ suggested that they controlled the differentiation and that such a function of the antibiotics expressed itself through the Ca2+ signal system. PMID- 10707802 TI - [The immunological aspects of predicting antibiotic therapy efficacy in peritonitis patients]. AB - The analysis of the impact of various group antibiotics on the mechanisms of development and correction of pathogenetically heterogeneous immune deficiency in 235 patients at age of 17 to 85 years with local (80 patients) and general (155 patients) peritonitis is presented. Cephalosporins and fluoroquinolones promoted restoration of the immune and interleukin (IL-1 and IL-2 of donors, IL-1r and IL 2r of patients) status and lowered the immunodepressive effect of glucocorticoids (GC) at the organism (cortisol, ACTH and cortisol-binding globulin) and cellular (GC receptors III) levels. Aminoglycosides and penicillins had no significant action on the immune and interleukin status but lowered the EG effect at the organism (aminoglycosides) and cellular (penicillins) levels. It is recommended that the antibiotics be used with an account of their involvement in the systemic reactions of the host. PMID- 10707803 TI - [The clinical laboratory assessment of vancomycin (Edicin) efficacy in treating suppurative wounds of the skin and soft tissues, burn wounds and the infectious complications of burns]. AB - Clinical and bacteriological efficacies of vancomycin (Edicin, LEK) in the treatment of 17 patients with wound infection and 13 patients with thermal affections were studied. The clinical efficacy in the group of the patients with purulent wounds of the soft tissues amounted to 94.1 per cent and that in the patients with thermal affections was 92.3 per cent. The bacteriological effect was recorded in 86.6 per cent of the patients with purulent wounds of the soft tissues and in 69.3 per cent of the patients with burn infections. The drug intolerability was observed in two cases. PMID- 10707804 TI - [The metabolites of anaerobic bacteria (volatile fatty acids) and the reactivity of the macroorganism]. PMID- 10707805 TI - [The immunotropic properties of I-->3; I-->6-beta-D-glucans]. PMID- 10707806 TI - Enzyme self-inactivation is a main limitation of the preparation of eicosanoids. Enzymatic synthesis of thromboxane and 12(S)-hydroxytetraenoic acid. AB - Synthesis of prostanoids is accompanied by various processes reducing the product yield. These processes are also known to affect syntheses of thromboxane (TX) and 12(S)-hydroxy-5(Z),8(Z),10(E),14(Z)-eicosatetraenoic acid (12-HETE). Partially purified preparations of TX synthase and prostaglandin (PG) synthase were used to optimize TX synthesis with respect to concentrations of the enzymes and eicosapolyenoic acid (EPA). Conditions for the maximum product yield and the minimum consumption of enzymes were determined. Consumption of the TX synthase was large owing to its inactivation during the reaction and the nonenzymatic destruction of the intermediate product PG-endoperoxide. Separate addition of PG and TX synthases increased the product yield by preventing EPA sorption on ballast proteins. Microsomal 12-lipoxygenase (12-LO) was also shown to be inactivated during the reaction, and this process was the major limitation of 12 HETE synthesis. Lipoxygenase reaction in the presence of some reducing agents led to a considerable increase of the 12-HETE yield, supposedly by preventing further oxidation of the 12-LO reaction product 12-hydroperoxy derivative of eicosatetraenoic acid. The possibility of using human blood platelet microsomes for preparation of some derivatives of EPAs is discussed. PMID- 10707807 TI - Modification of the electrokinetic properties of reversible electrophoresis gels for the separation and preparation of DNA. Addition of linear polymers. AB - The effect of adding linear polymers to a novel reversible electrophoretic was measured. Reversible gels are formed using the polyanionic carbohydrate polymer, gellan gum. Gellan gum forms strong stable gels in the presence of divalent cations or diamines. The gels are reversible (return to solution) by changing the ionic environment or pH. Gellan gum is an anionic polymer, and the electrophoresis gels have considerable electroosmotic flow (EOF) toward the negative electrode. We measured the EOF in gellan gum electrophoresis gels as a function of gel concentration, buffer composition, and linear polymer additive. The linear polymers used in this study were polyethylene oxide and hydroxyethyl cellulose. Both polymers reduced EOF in the gels, in a manner dependent on molecular weight. Polymers with high molecular weight were more effective at reducing EOF. The addition of polymers increased the resolution of low molecular weight DNA. Native gellan gum resolved DNA from approx 50,000 to 1000 bp. Addition of the polymers resolved DNA down to approx 50 bp, in some instances. The influence of the polymers on circular plasmid DNA was also investigated. Addition of high molecular weight polyethylene oxide reduced the electrophoretic mobility of the nicked circular form compared to the supercoiled form. PMID- 10707808 TI - Imaging membrane potential with voltage-sensitive dyes. AB - Membrane potential can be measured optically using a variety of molecular probes. These measurements can be useful in studying function at the level of an individual cell, for determining how groups of neurons generate a behavior, and for studying the correlated behavior of populations of neurons. Examples of the three kinds of measurements are presented. The signals obtained from these measurements are generally small. Methodological considerations necessary to optimize the resulting signal-to-noise ratio are discussed. PMID- 10707809 TI - Germ-cell warfare in ascidians: sperm from one species can interfere with the fertilization of a second species. AB - Ascidians (invertebrate chordates) are very abundant in many marine subtidal areas. They often live in dense multispecies clumps; thus, interspecific competition for space may be intense. Although most noncolonial species are broadcast spawners, their eggs can be fertilized only by sperm of the same species (1). Multiple fertilization is lethal and all animals have evolved blocks to polyspermy. Ascidian eggs block polyspermy by enzymatic (2) and electrical mechanisms (3). Sperm bind to N-acetylglucosamine groups on the vitelline coat (4, 5, 6, 7). Follice cells surrounding the vitelline coat release N acetylglucosaminidase during egg activation (8), preventing the binding of all sperm but a few (2). I show here that this interaction is not species-specific; sperm from one species can cause glycosidase release from follicle cells of a second species. Furthermore, once glycosidase release has been induced, the subsequent addition of sperm from the egg-producing species fails to fertilize a substantial proportion of these eggs. This leads to the hypothesis that sperm from one species of ascidian can interfere with fertilization of a second species. While intraspecific sperm competition has been well documented in several taxa (9, 10), this is the first record of sperm competition between species, or interspecific sperm competition. PMID- 10707810 TI - Light-sensitive voltage responses in the neurons of the cerebral ganglion of Ciona savignyi (Chordata: Ascidiacea). AB - Light-responsive behaviors such as siphon contraction (1), phototropism (2), and gamete release (3, 4) have been described in several ascidian species. The pigmented spots around the siphon openers (5), the epithelial cells of the sperm duct (6, 7), and the cerebral ganglion (8) have been suggested to be the photoreceptor candidates underlying these behaviors. However, these arguments have not yet been settled because no direct electrophysiological recordings of light-induced receptor potentials have been reported. In this study, we focused on the cerebral ganglion and performed intracellular recordings from the neurons in the ventral side of the cerebral ganglion in an isolated in vitro preparation of the neural complex in Ciona savignyi. We found that 24% (n = 115) of the recorded neurons showed various types of voltage responses to light stimuli. Almost all (27/28) of the recorded voltage responses were "on" responses that included hyperpolarizing and depolarizing responses and could be categorized into five types, except for a complex response recorded in one cell; the remaining one (1/28) was a depolarizing "off" response. This is the first report of electrophysiological recordings of light-sensitive voltage responses from ascidian cerebral ganglion neurons. PMID- 10707811 TI - Parasitic diatoms inside antarctic sponges. AB - Antarctic sponges may host large populations of planktonic and benthic diatoms. After settling on the sponge, these diatoms enter its body through pinacocytes (1) and form, there, large mono- or pauci-specific assemblages. Yet the total amount of carbohydrates in the invaded sponge tissue is inversely correlated with that of chlorophyll-a. We suggest, therefore, that endobiont diatoms utilize the products of the metabolism of their host as an energy source. This is the first evidence indicating that an endobiotic autotrophic organism may parasitize its animal host. Moreover, this unusual symbiotic behavior could be a successful strategy that allows the diatom to survive in darkness. PMID- 10707812 TI - Maintaining the line of defense: regeneration of Cuvierian tubules in the sea cucumber Holothuria forskali (Echinodermata, Holothuroidea). AB - When irritated, individuals of the sea cucumber Holothuria forskali expel a few Cuvierian tubules which lengthen, instantly become sticky, and rapidly immobilize most organisms with which they come into contact. After expulsion, the lost tubules are readily regenerated. When only a few tubules have been expelled, there is often a latent period before the regeneration starts. In contrast, when many tubules have been expelled, the regenerative process starts immediately but proceeds in successive waves of 10 to 30 tubules that begin to regenerate at 10 day intervals. However, in all cases, the complete regeneration of a given tubule takes about 5 weeks and may be divided into three successive phases: an initial repair phase including the overall 48-h post-autotomy period, a true regenerative phase taking about 4 weeks to complete, and a growth phase of about one more week. Initial regeneration events occur by epimorphosis, cell proliferation being essential to the regenerative process, whereas late events occur mainly by morphallaxis, with migration of the newly differentiated cells. The mesothelium is the tissue layer in which cell proliferation is the most precocious and the most important, involving both peritoneocytes and undifferentiated cells (which seem to be dedifferentiated peritoneocytes). As regeneration proceeds, the percentage of undifferentiated cells regularly decreases in parallel with the differentiation of granular (adhesive-secreting) cells and myocytes. The myocytes then separate off from the mesothelium and migrate within the connective tissue layer. Three types of pseudopodial cells follow one another in the tubule connective tissue during regeneration. Type 1 cells have all the characteristics of echinoderm phagocytes and may have a fibroblastic function, cleaning the connective tissue compartment before new collagen synthesis starts. Type 2 cells are rather undifferentiated and divide actively. The presence of type 3 cells is closely associated with the appearance of collagen fibers, and it is suggested that they have a fibroblastic function. In the inner epithelium, cells also divide actively, but only those in which spherules have not yet differentiated in the basal intraconnective processes. It appears, therefore, that in the three tissue layers of the tubules, regeneration proceeds by cell dedifferentiation, then proliferation, and finally by differentiation. Cuvierian tubules thus constitute a very efficient defensive mechanism: their large number, sparing use, and particular regeneration dynamics make them an almost inexhaustible line of defense maintained at limited energy cost. PMID- 10707813 TI - Nucleation and growth of calcite on native versus pyrolyzed oyster shell folia. AB - The thin sheets of calcite, termed folia, that make up much of the shell of an oyster are covered by a layer of discrete globules that has been proposed to consist of agglomerations of protein and mineral. Foliar fragments, treated at 475 degrees C for 36 h to remove organic matter, were imaged by atomic force microscopy (AFM) as crystals grew on the foliar surfaces in artificial seawater at calcite supersaturations up to 52-fold. Crystals were also viewed later by scanning electron microscopy. After pyrolysis, the foliar globules persisted only as fragile remnants that were quickly washed away during AFM imaging, revealing an underlying morphology on the foliar laths of a tightly packed continuum of nanometer-scale protrusions. At intermediate supersaturations, crystal formation was seen immediately almost everywhere on these surfaces, each crystal having the same distinctive shape and orientation, even at the outset with crystals as small as a few nanometers. In contrast, nucleation did not occur readily on non pyrolyzed foliar surfaces, and the crystals that did grow, although slowly at intermediate supersaturations, had irregular shapes. Possible crystallographic features of foliar laths are considered on the basis of the morphology of ectopic crystals and the atomic patterns of various surfaces. A model for foliar lath formation is presented that includes cycles of pulsed secretion of shell protein, removal of the protein from the mineralizing solution upon binding to mineral, and mineral growth at relatively high supersaturation over a time frame of about 1 h for each turn of the cycle. PMID- 10707814 TI - The apical sensory organ of a gastropod veliger is a receptor for settlement cues. AB - On the basis of anatomy and larval behavior, the apical sensory organ (ASO) of gastropod veliger larvae has been implicated as the site of perception of cues for settlement and metamorphosis. Until now, there have been no experimental data to support this hypothesis. In this study, cells in the ASO of veliger larvae of the tropical nudibranch Phestilla sibogae were stained with the styryl vital dye DASPEI and then irradiated with a narrow excitatory light beam on a fluorescence microscope. When its ASO cells were bleached by irradiation for 20 min or longer, an otherwise healthy larva was no longer able to respond to the usual metamorphic cue, a soluble metabolite from a coral prey of the adult nudibranch. The irradiated cells absorbed the dye acridine orange, suggesting that they were dying. When larvae not stained with DASPEI were similarly irradiated, or when stained larvae were irradiated with the light beam focused on other parts of the body, there was no loss of ability to metamorphose. Together these data provide strong support for the hypothesis. Potassium and cesium ions, known to induce metamorphosis in larvae of many marine-invertebrate phyla, continue to induce metamorphosis in larvae that have lost the ability to respond to the coral inducer due to staining and irradiation. These results demonstrate that (1) the ASO-ablated larvae have not lost the ability to metamorphose and (2) the ions do not act only on the metamorphic-signal receptor cells, but at other sites downstream in the metamorphic signal transduction pathway. PMID- 10707815 TI - Reproduction cycles and strategies of the cold-water sponges Halisarca dujardini (Demospongiae, Halisarcida), Myxilla incrustans and Iophon piceus (Demospongiae, Poecilosclerida) from the White Sea. AB - The reproductive development of the Demospongiae species Halisarca dujardini (Halisarcida), Myxilla incrustans and Iophon piceus (Poecilosclerida) from Chupa Inlet (Kandalaksha Bay, the White Sea) was studied histologically during 1982 1994 and 1997. These species are all viviparous. Halisarca dujardini inhabits shallow waters (1.5-5 m); M. incrustans and I. piceus are common in a more stable environment at depths between 15 and 25 m. Initiation of sexual reproduction stages is dependent upon water temperature. Reproductive effort is low in Myxilla incrustans and I. piceus (reproductive elements contribute 7.3% and 12% of maternal tissue volume respectively), but much higher in H. dujardini (up to 69% of the parental tissue volume). Reproduction leads to localized destruction of maternal tissue for M. incrustans and I. piceus and complete disorder of central and basal parts of the choanosoma of H. dujardini after each period of reproduction. Myxilla incrustans and I. piceus reproduce throughout the hydrological summer, but reproduction in H. dujardini is restricted to 3 weeks. The average life span of M. incrustans and I. piceus is more than 4 years, and that of H. dujardini is about 7-12 months. The data suggest that M. incrustans and I. piceus are K-strategists, whereas H. dujardini is an r-strategist. PMID- 10707816 TI - Walking versus breathing: mechanical differentiation of sea urchin podia corresponds to functional specialization. AB - The podia of sea urchins function in locomotion, adhesion, feeding, and respiration; but different podia on a single urchin are often specialized to one or more of these tasks. We examined the morphology and material properties of podia of the green sea urchin, Strongylocentrotus droebachiensis, to determine whether, despite apparent similarities, they achieve functional specialization along the oral-aboral axis through the differentiation of distinct mechanical properties. We found that oral podia, which are used primarily for locomotion and adhesion, are stronger and thicker than aboral podia, which are used primarily for capturing drift material and as a respiratory surface. The functional role of ambital podia is more ambiguous; however, they are longer and are extended at a lower strain rate than other podial types. They are also stronger and stiffer than aboral podia. In addition, all podia become stronger and stiffer when extended at faster strain rates, in some cases by nearly an order of magnitude for an order of magnitude change in strain rate. This strain-rate dependence implies that resistance to rapid loading such as that imposed by waves is high compared to resistance to slower, self-imposed loads. Thus, the serially arranged podia of S. droebachiensis are functionally specialized along an oral-aboral axis by differences in their morphology and mechanical properties. PMID- 10707817 TI - Egg brooding by deep-sea octopuses in the North Pacific Ocean. AB - Videotapes made from the submersible Alvin on Baby Bare, a 2600-m-deep North Pacific basalt outcrop, and at two other deep-sea localities document that octopuses of the genera Graneledone and Benthoctopus attach their eggs to hard substrate and apparently brood them through development. The behavior of brooding females was generally similar to that of shallow-water octopuses, but the genera showed apparent differences. In addition to the high density of brooding females observed at Baby Bare, which may relate to the increased availability of exposed hard substrates for egg attachment and of prey, females are suggested to increasingly associate with hard substrates as they mature. The biology of Baby Bare may seem unduly unique because the outcrop is isolated on a sedimented plain and is among the few exposures of hard substrate other than hydrothermal vents that have been explored by submersible. On the sediment-covered ocean floor, the availability of hard substrate may strongly affect the distribution of brooding octopuses. The size and shape of boreholes in 19 of over 400 thyasirid clam shells collected from Baby Bare support the hypothesis that octopuses had preyed upon the clams. PMID- 10707818 TI - Filamentous fungi associated with holothurians from the sea of Japan, off the primorye coast of Russia. AB - Holothurians (Holothurioidea, Echinodermata) are known to contain triterpene glycosides, which show antifungal activity. Nevertheless, fungi can be isolated from all organs of holothurians. During 1995-1996, mycelial fungi from several Far-Eastern holothurians--Apostichopus japonicus, Eupentacta fraudatrix, Cucumaria japonica--were collected from the Sea of Japan near the coast of Primorye (Russia) and studied. Twenty-seven species of marine fungi, mostly facultative ones belonging to the mitosporic fungi, were isolated from the holothurians and identified. Fungi isolated from the holothurian surface were more diverse and abundant than those from internal organs and coelomic fluids. Of the holothurians studied, Cucumaria japonica was poorest in abundance and diversity of fungi. The fungi Cladosporium brevicompactum and C. sphaerospermum were common in the holothurian coelom. Because of their high proteolytic activity, these fungi may be pathogenic to holothurians. The detritovorus holothurian A. japonicus was shown to modify the fungal assemblages within the marine bottom sediments. PMID- 10707819 TI - Cellular growth of host and symbiont in a cnidarian-zooxanthellar symbiosis. AB - The hydroid Myrionema ambionense, a fast-growing cnidarian (doubling time = 8 days) found in shallow water on tropical back-reefs, lives in symbiosis with symbiotic dinoflagellates of the genus Symbiodinium (hereafter also referred to as zooxanthellae). The symbionts live in vacuoles near the base of host digestive cells, whereas unhealthy looking zooxanthellae are generally located closer to the apical end of the host cell. Cytokinesis of zooxanthellae occurred at night, with a peak in number of symbionts with division furrows (mitotic index, MI = 12% 20%) observed at dawn. The MI of zooxanthellae decreased to near zero by the middle of the afternoon and remained there until the middle of the next night. Densities of live zooxanthellae living inside of host digestive cells peaked following cytokinesis, whereas densities of unhealthy looking symbionts were highest just before the division peak. Mitosis of host digestive cells was highest in the evening, also preceding the peak in zooxanthellar MI. This is the first study relating phased host cell division to diel zooxanthellar division in marine cnidarians. Food vacuoles were prevalent inside of digestive cells of field-collected hydroids within a few hours after sunset and throughout the night, coinciding with digestion of captured demersal plankton. Laboratory experiments showed that food vacuoles appeared in digestive cell cytoplasm within 2 h of feeding with nauplii of Artemia. The number and size of food vacuoles per digestive cell and the percentage of digestive cells with food vacuoles all decreased 5-7 h following feeding in laboratory experiments, and by mid-day in field-collected hydroids. Light and external food supply were important in maintaining phased division of the symbionts, with a lag in response time to both parameters of 11-36 h. Altering light and feeding during the night did not influence the level of the peak MI the next morning, though in one experiment the absence of light slowed final separation of daughter cells at the end of cytokinesis. In another experiment, hydroids starved for 3-7 d and "pulse-fed" Artemia nauplii for 1 h at the beginning of the dark period showed continued low symbiont division (< 5%) after 11 h, whether maintained in constant light or darkness, implying that most algal division is set more than 24 h prior to actual cytokinesis. Transferred to a 14:10 h light:dark cycle for another 24 h (36 h after feeding), the same hydroids exhibited a "normal" peak MI (ca. 15%) at dawn, but zooxanthellae from hydroids kept in constant darkness still showed a low MI. These results show that mitosis of symbiotic dinoflagellates requires three factors: external food; a minimum period of time following feeding (11-36 h), presumably for digestion; and a period of light following feeding, presumably to provide carbon skeletons necessary for completing cytokinesis. PMID- 10707820 TI - Discrimination and phylogeny of solenogaster species through the morphology of hard parts (Mollusca, Aplacophora, Neomeniomorpha). AB - Ten species in five genera and three families from continental shelf and deep-sea collections of neomenioid Aplacophora (Mollusca) are described, emphasizing external anatomy and hard parts--body shape, radula, epidermal spicules, and copulatory spicules--as well as the reproductive system. One genus and seven species are new: Plawenia n.g., Plawenia sphaera, P. argentinensis, Dorymenia tortilis, Eleutheromenia bassensis, E. mimus, Kruppomenia levis, and K. delta. Also included are redescriptions of three published species, emphasizing hard parts for comparisons with the new species and genus: Dorymenia sarsii (Koren & Danielssen), Simrothiella margaritacea (Koren & Danielssen), and Plawenia schizoradulata (Salvini-Plawen). A cladistic analysis of species described here demonstrates the usefulness of hard parts for phylogeny. Specimens came from collections made in the southwest Pacific and the southwest and northeast Atlantic. PMID- 10707821 TI - A dynamical model of communities and a new species-abundance distribution. AB - It is known to many field biologists that biosurveys of natural communities tend to produce a J-shaped curve when the numbers of species are plotted against abundance. In other words, when the number of species of abundance k is plotted against k (running from 1 to some large number), the resulting distribution peaks at the lowest abundance, then forms a concave ramp as it approaches zero at the far end of the abundance axis. Does this distribution represent a single formula operating behind the scenes, or does it represent several formulas, appropriate for different types of community? Or does it represent no particular formula at all? The research reported here has three components: (1) The analysis of a new dynamical system that simulates multispecies communities (producing J-curves in the process) and the derivation of the "logistic-J" distribution, as the underlying community equilibrium curve; (2) the summary of a general theory of sampling as a bridge between natural communities and samples of them; (3) the evaluation of extant proposals for species-abundance distributions by application of a general theory of sampling or by cross-comparison via 100 biosurveys randomly selected from the literature. PMID- 10707822 TI - Regulation of vascular tone by plant polyphenols: role of nitric oxide. AB - Vascular endothelium plays a crucial role in regulating blood flow and vascular tone. It can synthesize and release different relaxant factors including nitric oxide (NO). This article summarizes pharmacological properties of red wine polyphenol extracts (RWPC) with respect to endothelial NO. It is shown that RWPC produces endothelium-dependent relaxation as a result of enhanced NO synthesis rather than enhanced biological activity of NO or protection against breakdown by O2-. The mechanisms involve influx of Ca2+ and production of O2- within the endothelial cells. These results suggest that RWPC, by releasing endothelial NO, may have therapeutically relevant effects against cardiovascular diseases. PMID- 10707823 TI - Structural alterations in the heart after long-term L-NAME and D-NAME treatment. AB - N(G)-nitro-D-arginine methyl ester (D-NAME), considered as an inactive enantiomer of NAME, is generally used as a negative control for NO synthase inhibition with L-NAME. The aim of this work was to compare the effect of L-NAME (20 and 40 mg/kg/day), and D-NAME (40 mg/kg/day) on hemodynamic and structural parameters in the rat cardiovascular system. After 4 weeks of treatment, blood pressure and left ventricle weight/body weight ratio increased significantly in all studied groups versus control. Myocardial fibrosis (in %) represented 0.94 +/- 0.04 in control, 4.70 +/- 0.39 in L-NAME (20 mg/kg/day), 10.54 +/- 0.91 in L-NAME (40 mg/kg/day) and 5.25 +/- 0.46 in D-NAME (40 mg/kg/day) group. We conclude that in a long-term experiment D-NAME provokes similar changes in cardiovascular system like L-NAME. PMID- 10707824 TI - Functional alterations of cardiac (Na,K)-ATPase in L-NAME induced hypertension. AB - (Na,K)-ATPase, an enzyme involved in the active translocation of Na+ and K+ ions across cell membranes was shown to be affected by nitric oxide (NO) in various tissues. In the present study the functional alterations of (Na,K)-ATPase after chronic inhibition of nitric oxide synthesis were studied in rat hearts. Four weeks lasting administration of an L-arginine analogue, the N(G)-nitro-L-arginine methyl ester (L-NAME) induced an increase in the systolic blood pressure of about 36%. In this hypertension the kinetic parameters Km and Vmax for ATP-activation of the (Na,K)-ATPase did not show any significant changes. Activation of the enzyme by its cofactor Na+ revealed no change in the Vmax, but the K(Na) increased by 50%. Two weeks after terminating the administration of L-NAME the blood pressure returned to control values. In these conditions the activity of (Na,K)-ATPase increased, due to enlarged affinity of the ATP-binding site as revealed from the diminished Km value for ATP. The K(Na) value for activation with Na+ returned to control value. Our findings indicate that there is no change in energy utilization by the (Na,K)-ATPase during L-NAME induced hypertension in the heart. The transport properties of the enzyme are deteriorated, due to its decreased sensitivity to Na+. This inhibition of the (Na,K)-ATPase might be responsible for the increase of [Na+]i during lowered NO synthesis. In hearts from rats that recovered from the hypertension, the (Na,K)-ATPase increases its activity due to improved ATP binding properties. PMID- 10707825 TI - Cardiac membrane proteins and phospholipids in L-NAME induced hypertension. AB - This study deals with qualitative alterations of membranes in cardiac cells after chronic inhibition of NO synthesis in rats induced by administration of N(G) nitro-L-arginine methyl ester (L-NAME) in a dose 40 mg/kg/day for 4 weeks. Concentration of proteins did not change either in the cytosolic fraction or in the particulate fraction of the cardiac homogenate from L-NAME treated rats. The concentration of phospholipids and consequently the ratio of phospholipids to membrane proteins increased by 100% and 88%, respectively. The concentration of conjugated dienes (CD), often used as an indirect marker for the production of free oxygen radicals, increased by 141% after calculation per gram of tissue. However, evaluation of CD concentration directly in phospholipids revealed no change, suggesting that phospholipids in cardiac tissue after L-NAME treatment were not damaged additionally, by increased level of free oxygen radicals. The increase of the CD concentration in the cardiac tissue is therefore, a consequence of the elevated phospholipid concentration. PMID- 10707826 TI - Hypokalemia-induced ultrastructural, histochemical and connexin-43 alterations resulting in atrial and ventricular fibrillations. AB - Perfusion of the isolated guinea pig heart with hypokalemic solution provide simple model for examination of the molecular mechanisms involved in the incidence of atrial and/or ventricular fibrillations. The results point out that dispersion of the metabolic and subcellular alterations and heterogenously impaired intercellular coupling might account for electrical disturbances and desynchronization of the myocardium thus facilitate occurrence of fibrillation. PMID- 10707828 TI - Cerebral angiogenesis shows nestin expression in endothelial cells. AB - The class VI intermediate filament protein nestin has been generally considered as a specific marker for neural precursor cells or developing muscles. In the prenatal developing rat central nervous system (CNS), we localized immunoreactivity for the nestin in blood vessels. Although the widespread nestin expression in cerebral blood vessels persisted in early postnatal periods, it was down-regulated in the adulthood. However, when the adult rat brains were subjected to procedures that trigger neovascularization, e.g. grafting fetal nervous tissue or C6 glioma, the abundant immunoreactivity was detected in all newly formed vessels and adjacent host vasculature. Our results demonstrate that nestin expression in endothelial cells lining cerebral vessels accompanies the process of angiogenesis. PMID- 10707827 TI - The role of sarcoplasmic reticulum in the protective effect of class III drugs against Ca2+ overload. AB - Various studies on humans and experimental mammals showed that d-sotalol and tedisamil (class III antiarrhythmic drugs with positive inotropic effect) facilitate spontaneous ventricular defibrillation. Following our previous results, we summarized that spontaneous ventricular defibrillation requires high level of intercellular coupling and synchronization, both of which depends on intracellular free Ca2+ concentration. We hypothesized that any antiarrhythmic compound that facilitates spontaneous defibrillation, including d-sotalol and tedisamil, should prevent intracellular free Ca2+ overload most likely by elevating cAMP level and enhancing cAMP-related Ca2+ uptake of the sarcoplasmic reticulum (SR). The aim of the present study was to examine the role of the SR uptake function in their effect against Ca2+ overload. METHODS: The effect of d sotalol, tedisamil and dbcAMP on increased intracellular Ca2+ level were examined in cultured rat cardiomyocytes during blockade of SR Ca2+ uptake by administration of thapsigargin (TG), a selective inhibitor of Ca2+-ATPase. RESULTS: Administration of 3 x 10(-6) mol/l TG, prior to d-sotalol, tedisamil and dbcAMP, significantly increased intracellular free Ca2+ concentration and prevented the effect of d-sotalol, tedisamil or dbcAMP to decrease intracellular Ca2+ level to its beseline, while 10(-6) mol/l TG prevented it only partially. Administration of either d-sotalol or tedisamil (at concentration of 10(-5) mol/l) before the administration of 10(-6) mol/l TG prevent the TG induced elevation of [Ca2+]i. CONCLUSION: These results support our hypothesis that d sotalol and tedisamil prevent Ca2+ overload by the cAMP dependent SR Ca2+ uptake. PMID- 10707829 TI - The determination of the collagen and elastin amount in the human varicose vein by the computer morphometric method. AB - The results of the works dealing with alterations of the connective tissue in varicose vein wall are not ambiguous, so the exact cause of the vein dilatation has still not been established. We were determining the collagen and elastin amounts in human varicose vein wall in comparison with non-dilated long saphenous vein by the light microscopy and computer morphometric method. We have found the lesser amount of collagen in varicose veins than in non-dilated veins, the amounts of the elastin in both the varicose and non-varicose veins were without the statistical significance. PMID- 10707830 TI - CD26 and DPP IV expression in T acute lymphoblastic leukemia cells: immunocytochemistry and enzyme cytochemistry. AB - Recent studies have revealed the dipeptidyl peptidase IV (DPP IV) enzymatic activity of CD26 antigen. In this paper, the possible identity of DPP IV and CD26 expression in phenotypically defined T-ALL has been examined. The combination of enzyme cytochemistry and immunocytochemistry was used. The correlation between the CD26 antigen expression and DPP IV positivity in the vast majority of T lymphoblasts in T-ALL patients was observed. No CD26 was expressed on DPP IV negative T cells. The variable CD4 and/or CD8 antigen expression, frequent CD7 positivity and absence of membrane CD3 antigen expression were the characteristic immunophenotypic features of CD26/DPP IV positive T cells. CD26/DPP IV activity strongly paralleled the CD71 antigen (transferrin receptor, T cell activation/proliferation antigen) expression. The phenotypic features of CD26/DPP IV positive T cells are characteristic for the relative immature cell population. Noteworthy was the slight disassociation between the very high CD26 antigen J expression and moderate DPP IV activity in cells of some T-ALL patients. The possible existence of enzymatically inactive structures of CD26 antigen or inactive precursors of DPP IV detectable only by immunocytochemistry was discussed. Our study indicates that CD26 antigen expression is tended to identify cells with DPP IV enzymatic activity in T-ALL patients. The results provide information of CD26 antigen possible involvement in the pathology of leukemic cells via its DPP IV enzyme activity. PMID- 10707831 TI - Mola invasiva--special form of GTD. AB - Invasive hydatidiform mole is a relative rare form of gestational trophoblastic disease (GTD). Most of hydatidiform moles remit after evacuation but some of them have the tendency to invade the myometrium. In some rare cases the trophoblastic tissue can be found in other tissues like lungs, vulva, vagina or broad ligament. The aim of the study was to demonstrate some of clinical, immunohistochemical and DNA analysis findings of a patient with a previous diagnosis of a complete hydatidiform mole. PMID- 10707832 TI - The normal female and the male breast epithelium does not express prostate specific antigen. Preliminary immunohistochemical observations of autopsy breast tissues. AB - In the normal female and male breast epithelial structures any prostate-specific antigen (PSA) immunohistochemical positivity was observed. Variable PSA expression, which often borders the positivity, was observed in membranes of adipocytes of fat tissue and in the endothelium of small vessels in a female and a male breast. Based on these initial observations, tissue of the normal breast, male or female, can not be considered to be the principal source of PSA. PMID- 10707833 TI - The role of adenylate cyclase in ischemic preconditioning in the rat heart: a cytochemical study. AB - Using catalytic cytochemistry the AC activity was studied during ischemic preconditioning (IP) (5 min occlusion of LAD and 10 min reperfusion) followed by 30 min regional ischemia in isolated Langendorff-perfused rat heart. In controls the specific precipitate of AC reaction was found on the sarcolemma (SL) and the junctional sarcoplasmic reticulum (JSR) of cardiomyocytes. After prolonged ischemia the reaction product was absent, whereas IP followed by prolonged ischemia protected the AC activity on SL and JSR. IP-induced enhancement of AC activity in this model was accompanied by significant reduction of ischemia/reperfusion fibrillation. The results suggest involvement of AC system in mechanisms of IP. PMID- 10707834 TI - Detection of cytoskeletal proteins in small cell lung carcinoma. AB - Small cell lung carcinoma (SCLC) is the most aggressive of lung tumors, metastasize widely and are virtually incurable by surgical means. Therefore, the classification of lung cancer into SCLC and non-small cell lung carcinoma is essential for disease prognosis and treatment. For this purpose we have compared the immunohistochemical distribution of different cytoskeletal proteins as tumor markers. Analysis was performed by using of monoclonal antibodies directed against cytokeratins, neurofilaments, betaIII-tubulin, epithelial membrane antigen and neuron-specific enolase. Our results indicate that keratin and epithelial membrane antigen are reliable epithelial markers for SCLC. In addition, the positive staining with monoclonal antibodies TU-20 against betaIII tubulin and neuron-specific enolase was found in some cases of SCLC. We suggest, that these antibodies could be a useful tool for complex immunohistochemical diagnosis of SCLC. PMID- 10707835 TI - Restructuring and extrusion of nuclear ribonucleoproteins (RNPs) during apoptosis. AB - To investigate the fate of nuclear ribonucleoprotein (RNP) during apoptosis, we performed a cytochemical and immunocytochemical study of apoptotic mammalian cells by fluorescence and electron microscopy using specific antibodies which recognize different RNP-associated proteins. Light and electron microscopy showed that during apoptosis nuclear RNPs are rearranged, with the formation of fibrogranular heterogeneous clusters which are extruded from the nucleus into the cytoplasm, and finally released at the cell surface, as apoptotic blebs. Restructuring and extrusion of nuclear RNPs apparently determine the arrest of RNP maturation, thus effectively blocking protein synthesis in apoptotic cells. PMID- 10707836 TI - Growth and differentiation of the vascular smooth muscle and endothelial cells cultured on fluorine ion-implanted polystyrene. AB - The rat vascular (SMCs) and bovine endothelial cells (BECs) were cultured on conventional or fluorine ion-implanted polystyrene (5 x 10(12) and 5 x 10(14) fluorine ions/cm2). The cells grown on the implanted growth supports showed better adherence, higher volume and higher total protein content. The immunocytochemical analysis revealed that SMCs contained more of the cytoskeletal vimentin and the vascular SMC-specific alpha-actin as well as several cell adhesion-mediating molecules (vinculin, talin, alpha(v)-integrin and ICAM-1). In BECs, only the content of vimentin and talin increased, while expression of ICAM 1 was unchanged. The data suggest that cells on the ion implanted polymers could be more viable and that increased expression of some adhesion molecules mediating interactions with the host immune system is cell type-dependent. PMID- 10707837 TI - Different responsiveness of male and female rat aortic smooth muscle cells (SMCs) to repeated passaging in culture. AB - The smooth muscle cell (SMC) cultures were prepared from the aorta of male and female 8-week-old rats and used at passage 5-7 or 40-45. On day 1, low-passaged cells of both sex groups adhered to growth supports at similar numbers while after repeated passaging the adherence of female-derived cells was higher. These cells had also higher total protein content and contained more of the SMC specific alpha-actin, vimentin and alpha(v) integrins. Compared to the male type of cultures, the high passaged cells of female origin cycled at a slower rate and were undergoing massive polyploidization. Male-derived cells remained of the same morphology, ploidy and the differentiation status at all passages. Their passage response consisted mainly in faster cycling and growth to higher population densities. The data could be of importance for explanation of different incidence of hyperplastic vascular diseases in males and females. PMID- 10707838 TI - Ultrastructural response of the nuclear envelope (NE) of C6-glioma cells to cisplatin-induced apoptosis. AB - The early response of the nuclear envelope of C6-glioma cells (t < or = 24 h), treated with a cytostatic dose of cisplatin in culture (5 microg/ml) included formation of slim and deep invaginations formed by either the inner or both membranes. The invaginations made of the complete NE often extended up to the enlarged nucleoli. In some of them, nucleus-like material occurred at their cytoplasmic site suggesting its enhanced nucleus-to-cytoplasm transport. Some nuclear pores in the invagination-forming cells were covered by dome-shaped "caps" protruding into the karyoplasm. The "capped" pores were absent in the cells that were initially more damaged. At 48 to 72 h, we found a small number of large and hyperlobulated cells with some small lobules containing a rarefied chromatin and focally disintegrated NE. The lamina-free remnants of the NE with a swollen perinuclear cistern were still present at 72-96 h when the population entered the execution phase of apoptotic death. PMID- 10707839 TI - Immunohistochemical localization of laminin-1 in the acellular nerve grafts is associated with migrating Schwann cells which display corresponding integrin receptors. AB - The presence of laminin-1, collagen-IV, alpha6 and beta1 integrin chains was detected by indirect immunohistochemistry using biotin/streptavidin/HRP or gold conjugated secondary antibody at the light and electron microscope level, respectively. Cryo-treated segment of the peripheral stump without living Schwann cells (S-100-) did not display immunoreactivity for laminin-1 and integrin's chains, while the migrating Schwann cells in the marginal regions were immunostained for the antigens. Isolated acellular nerve segments protected from migration of Schwann cells (S-100-) exhibited laminin-1-, beta1-, and alpha6- integrin chains immunoreactivities. Position of the basal lamina was verified by collagen-IV+ immunoreactivity. Results indicate that presence of the laminin in the peripheral nerve is related with living Schwann cells. PMID- 10707841 TI - Difference of pericentral NADPH-d positive neurons in the rabbit spinal cord segments. AB - The purpose of the present investigation was to characterize and determine the number of NADPH-diaphorase positive neurons around the central canal in the rabbit spinal cord. These neurons are known to function as interneurons and are present in all spinal cord segments. They differ in shape of their bodies and in length and branching of their processes. The main differentiation was observed in their number, depending on the place of their localization. The highest number of these NADPH-diaphorase positive neurons was in sacral part (6 in average), the lowest one was noticeable in thoracic spinal cord (1-2 in average). It can be concluded that pericentral neurons of the rabbit spinal cord are capable of synthesizing nitric oxide and that they differ in number, depending on the place of their localization in each spinal cord segment. PMID- 10707840 TI - Immunohistochemical localization of some extracellular molecules and their integrin receptors in the rat Pacinian corpuscles. AB - The present results suggest that laminin-1 and 3 are localized in the specialized Schwann cells of Pacinian corpuscles, in spite of incomplete deposition of the basal lamina on the surface of their cytoplasmic processes. In addition, laminin 3 is concentrated and probably function as a stop protein not only in the neuromuscular junction, but also in the specialized Schwann cells enveloping the dendritic zone of the afferent axon. No significant changes of immunostaining for both laminins and their integrin receptors following denervation of Pacinian corpuscles indicate that their synthesis is independent to afferent axon as a prerequisite for successful reinnervation. PMID- 10707842 TI - Coexistence of nitrergic and acetylcholinesterase (ACHE)-positive nerve structures of the phaesant spleen. AB - The coexistence of neuronal NADPH-diaphorase and ACHE activities were investigated in the phaesant spleen by successive double histochemical staining of the same sections. Two types of nerve structures were found in pheasant the spleen: nerve cells and nerve fibres. NADPH-d and ACHE-positive nerve fibres in colocalization enter the spleen in its hilum in the vicinity of splenic artery branches and are gradually distributed in periarterial topography in the white pulp. Only NADPH-d positive nerve cells were seen around the splenic vessels. In the red pulp and splenic capsule, only ACHE-positive nerve fibres were present. PMID- 10707843 TI - The influence of transient spinal ischemia on substance P positive nerve structures. AB - The aim of this study was to investigate, if transient spinal ischemia and a period of 4-day reperfusion will change the distribution pattern of substance P in the spinal cord of rabbits. Strongly enhanced staining of substance P positive nerve structures appeared in the superficial dorsal horn (laminae I, II), the Lissauer's tract, the pericentral region (lamina X), and in the areas of autonomic nuclei (sympathetic-intermediolateral--IML nucleus and parasympathetic sacral parasympathetic nucleus--SPN) in the control group. Transient spinal ischemia was produced by occlusion of the abdominal aorta just below the left renal artery. Neuropathology of the lesion 4 days after transient ischemia was characterized by selective necrosis of gray matter in the central part of dorsal horn and medial portions of anterior gray matter. Areas with the most dense accumulation of substance P positive structures stayed almost intact. Therefore, no significant change in the distribution pattern of substance P was found in the spinal cord of animals with ischemia-reperfusion-induced injury. PMID- 10707844 TI - Selective sparing of NADPH-d positive spinal cord neurons affected by ischemia. AB - Histochemical characterization of NADPH diaphorase positive neuronal pools in the rabbit lumbosacral segments was performed during and after transient spinal cord ischemia. Strongly enhanced staining of NADPH diaphorase positive structures appeared in the superficial dorsal horn, the pericentral region and in the neurons of the sacral parasympathetic nucleus at the end of 40 min of abdominal aorta ligation or after 1 day reperfusion. Four days after ischemia, NADPH-d positive neurons and vessels were detected in the central gray matter despite well developed necrosis in this location. Regional nitric oxide synthesis and its vasodilatatory effect during the period of aortic occlusion may account for the observed selective resistance of these spinal cord neurons to transient ischemia. PMID- 10707845 TI - Expression of myosin heavy chain (MyHC) isoforms in rat intrafusal muscle fibres after neonatal deefferentation and subsequent denervation. AB - The analysis of developing intrafusal fibres is not feasible in the absence of primary sensory axons, as neonatal denervation leads to the disintegration of muscle spindles. On the other hand, neonatal deefferentation does not arrest their differentiation and, moreover, it leads to the neomyogenesis of supernumerary intrafusal profiles. If the sciatic nerve was sectioned in 4-week old rats deefferented at the birth, muscle spindles survived, the neomyogenesis proceeded and the denervated intrafusal fibres expressed the spindle specific slow tonic (STO) MyHC. The expression of MyHC pattern in individual fibres and the differentiation of the fibre type characteristics were, however, less obvious compared to the control or deefferented spindles. The newly formed intrafusal profiles (which differentiated from satellite cells in the absence of innervation) expressed the STO MyHC particularly when they developed in a spatial relation to nuclear bag fibres. PMID- 10707846 TI - Stereological evaluation of the soleus muscle isografted into fast extensor digitorum longus (EDL) muscle in rats with different thyroid status. AB - The 2-D stereology can be used advantageously in the case of muscle cross sections stained by routine histochemical and immunocytochemical methods, such as mATPase reaction, when the quality of the image is often not sufficient for using image analysis techniques without considerable individual intervention. Other advantages of stereological methods in muscle morphometry are that measurements are made directly on specimens under the microscope and in their simplest arrangement they do not require sophisticated and expensive technical equipment. Furthermore, unbiased results are obtained, no segmentation and edge effect problems arise and the quantity of work invested in stereological estimation is reasonable. Therefore, we have used the stereological methods as our standard technique for assessment of fibre type composition in regenerated soleus muscles grafted from 21- to 28-day-old rats into fast EDL muscles of adult inbred recipients with different plasma levels of thyroid hormones. PMID- 10707847 TI - The up-regulation of utrophin is not limited to muscular dystrophies. AB - Immunohistochemical reactivity for utrophin has been recorded in 45 biopsies from patients with various neuromuscular diseases. The upregulation of utrophin on the extrajunctional sarcolemma has been found in dystrophinopathies, other muscular dystrophies, congenital myopathies, inflammatory myopathies, neurogenic muscle disorders (diabetic neuropathy, amyotrophic lateral sclerosis and spinal muscular atrophies), minimal change myopathies as well as in some normal biopsies. PMID- 10707848 TI - Apoptosis and expression of proliferative proteins in the developing nervous system and orofacial region of human embryos. AB - We have studied expression of PCNA and Ki-67 in the developing nervous system, sensory organs and orofacial regions in human embryos and fetuses, using monoclonal antibodies PC-10 and MIB-1 in three-step immunohistochemical method and apoptosis performed by TUNEL technique. Expression of PCNA and Ki-67 increased with the age. Apoptosis was rare in above mentioned regions. PMID- 10707849 TI - Detection of ischemic changes in the cytoplasm of neurocytes from rat brain and spinal cord by densitometric measurement of methylene blue binding. AB - Ischemic changes in neurocytes from brain and spinal cord of rats were studied by densitometric measurement of bound basic stain--methylene blue. Statistically significant differences in integrated optical density (I.O.D.) of cytoplasm near to cell nucleus in brain and spinal cord neurocytes were detected after ischemia. After 10 minutes of ischemia, the average values of I.O.D. decreased to 65% and to 69.9% of I.O.D. values of controls. After 2 hours of ischemia, the average values of I.O.D. in brain cell cytoplasm reached only 43.6% and in the spinal cord cells they fell to 54.5% of control values. PMID- 10707850 TI - The role of the apoptosis and the genes regulating apoptosis in the early differentiation of human embryo. AB - Apoptosis (programmed cell death) is an important process participating in the formation of organs and tissues during embryogenesis. Our aim of the work is studying the role of the apoptosis during the human embryonic differentiation. We tend to give acquired findings into the correlation with expression of proteins Bcl-2 and Bax (products of genes regulating apoptosis). Detection of the apoptosis was carried out on 25 routinely processed human embryos by means of TUNEL technique. The level of expression of Bcl-2 and Bax was determined using standard three-step immunohistochemical procedure. Results were achieved by the comparison of apoptoic index and the level expression of Bcl-2 and Bax was semiquantitatively evaluated. The low value of apoptotic index was mostly accompanied by the high expression of Bcl-2 and the Bax expression was not proportionally related to the value of apoptic index. PMID- 10707851 TI - Immunohistochemical demonstration of vimentin and S-100 protein in the kidneys. AB - Vimentin and S-100 protein expression was studied in the kidneys of adult sheep and goat using immunohistochemistry. Vimentin was detected in the podocytes, mesangial cells of the glomerulus, in the endothelium of renal capillaries and renal stromal cells. In collecting tubules, ducts and nerves of the renal papilla, S-100 protein was expressed. PMID- 10707852 TI - Expression of cytokeratins in the urinary passages. AB - The distribution of Pan cytokeratin and cytokeratin 18 in the dog and sheep urinary bladder and ureter as seen by immunohistochemistry using monoclonal antibodies is described. Both cytokeratins were observed in the urinary bladder and ureter of the studied species. Differences in their localization are described. PMID- 10707853 TI - Functional specialization of the epithelium in the mesonephric tubules. AB - An electron microscopy study was aimed to correlate structural differentiation of the epithelium in mesonephric proximal tubules (PT) with the expression of membrane activities of alkaline phosphatase (AP) and 5'-nucleotidase (AMP). Tissue samples of mesonephros were taken from 5 to 16 days old chick embryos. Both enzymes were detected with cerium technique, Mayahara modification of lead capture method was used also for localization of AP. Control incubation was performed with levamisole. The formation of absorptive apparatus was characterized by the differentiation of PT epithelium. Activities of AP and AMP appeared to increase rapidly with the differentiation of epithelium. Reaction products of AP and AMP were detected on brush border as well as on membranes of tubular invaginations, transport tubules and endocytotic vacuoles. The basolateral cell surfaces of epithelium were projected in short interdigitating microvilli and the expression of AP and AMP activities on their membranes suggested the transport role of this structural specialization. PMID- 10707854 TI - Total and local changes in the arthritis adjuvans. AB - Arthritis adjuvans was studied in the murine model. An effect of different treatment (methotrexate, tauredon, collagen hydrolysate) was estimated in the course of developing disease (day 3, 5, 11 and 21). Repeated evaluation of body weight and peripheral blood leukograms as a total response of organism was performed. Oedema of paw periarticular and tail regions, light- and electron microscopical screening and immunohistochemical investigation of prevalence of interleukin-1-beta (IL-1beta) and tumour necrosis factor (TNF-alpha) were estimated. The most pronounced benefit effect of methotrexate at stabilization of the monocytes blood level, synovial membrane cell invasion and TNF-alpha immunopositivity was ascertained. PMID- 10707855 TI - Dynamics of beta-glucosidase Zm-p60.1 ectopic expression during transgenic pollen development: a histochemical approach. AB - Zm-p60.1 is maize cDNA coding cytokinin-glucoside specific beta-glucosidase. Indigogenic method was used for histochemical localization of Zm-p60.1 beta glucosidase activity in various developmental stages of transgenic tobacco anthers. Expression of Zm-p60.1 cDNA in T7 tobacco plants is controlled by the CaMV 35S promoter. Another type of tobacco transformant expresses Zm-p60.1 under the control of LAT 52 promoter. Histochemical detection has proved different patterns of beta-glucosidase activity during tobacco pollen development in these two types of transformants. Zm-p60.1 beta-glucosidase activity had not direct influence on pollen germinability. PMID- 10707856 TI - Serum autoantibodies in patients with primary sclerosing cholangitis. AB - BACKGROUND/AIM: Primary sclerosing cholangitis is a chronic cholestatic syndrome with a presumed autoimmune basis frequently associated with inflammatory bowel disease. The aim of this study was to determine the profile and significance of serum autoantibodies in patients with primary sclerosing cholangitis. METHODS: Serum samples taken from 73 untreated patients (32 female and 41 male, median age 45 years) with well-defined primary sclerosing cholangitis, and from 75 healthy age- and sex-matched controls were assayed for 20 different autoantibodies. RESULTS: Of 73 patients, 71 (97%) were positive for at least 1 autoantibody; whereas 59/73 patients (81%) were positive for > or =3 antibodies. Patients with primary sclerosing cholangitis had a significantly greater rate of positivity than controls for antinuclear, anticardiolipin, antineutrophil cytoplasmic, and antithyroperoxidase antibodies as well as rheumatoid factor. The rate of positivity and serum levels of any of these 20 autoantibodies were not significantly different between patients with primary sclerosing cholangitis and inflammatory bowel disease and those without inflammatory bowel disease. Anticardiolipins were the single group of antibodies that had a significant correlation with the Mayo risk score (r=0.49, p<0.001) and histologic stage of disease (r=0.30, p<0.01). CONCLUSIONS: Primary sclerosing cholangitis is associated with a high proportion of non-organ specific autoantibodies. Anticardiolipin antibodies appear to be related to the severity of primary sclerosing cholangitis and may be a useful prognostic marker. PMID- 10707857 TI - Superoxide dismutase activity in children with chronic liver diseases. AB - BACKGROUND/AIMS: Liver disease in infancy has multiple etiologies. As reactive oxygen intermediates are involved in several types of tissue damage, we have investigated whether different forms of liver disease in infancy are associated with increased free radical generation, using an indirect approach in which superoxide dismutase (a free radical scavenger) activity is determined in the liver tissue. METHODS: A total of 48 liver biopsies performed at diagnosis were evaluated retrospectively. Nine infants had biliary atresia, eight Alagille syndrome, seven alantitrypsin deficiency and 12 cryptogenic hepatitis. As controls we studied 12 biopsies with normal histology obtained from seven children with portal vein thrombosis and five children who underwent biopsy for management reason but had no liver disease. Superoxide dismutase activity in liver biopsy specimens was measured using the cytochrome C method by spectrophotometry and expressed as U SOD/mg protein. RESULTS: Superoxide dismutase activity was significantly increased in biliary atresia (1.25 +/- 0.56 U SOD/mg protein, p<0.0001) and Alagille syndrome (1.31 +/- 0.56 U SOD/mg protein, p<0.0001) as compared with al-antitrypsin deficiency (0.75 +/- 0.3 U SOD/mg protein), neonatal hepatitis (0.72 +/- 0.37 U. SOD/mg protein) and normal controls (0.4 +/- 0.7 U. SOD/mg protein). The highest level of SOD activity was found, however, in control children with portal vein thrombosis (2.09 +/- 0.96 U SOD/mg protein; p<0.0001 as compared to the other groups). CONCLUSION: Superoxide dismutase, a key enzyme in free radical protection, is increased significantly in the liver tissue of infants with cholestatic liver disease due to bile duct damage and in children with portal vein thrombosis, suggesting that products of free radical reactions are involved in the pathogenesis of these disorders. PMID- 10707858 TI - Dissolution of copper-rich granules in hepatic lysosomes by D-penicillamine prevents the development of fulminant hepatitis in Long-Evans cinnamon rats. AB - BACKGROUND/AIM: The Long-Evans cinnamon rat has a mutation homologous to the human Wilson disease gene, leading to gross copper accumulation and the development of hepatitis. D-penicillamine, a copper-chelating drug widely and efficiently used in treating Wilson disease, has also been shown to prevent hepatitis in Long-Evans cinnamon rats. The objectives of this study were: i) to investigate the effectiveness of D-penicillamine when administered to the already affected animals, and ii) to elucidate the mechanism of action of the drug. METHODS: Long-Evans cinnamon rats were divided into groups according to age and treatment with D-penicillamine. The drug was administered orally before and after the onset of hepatitis. Livers were examined by light and electron microscopy. The effect of D-penicillamine on the subcellular distribution and binding of copper was investigated in more detail. Finally, the interaction between D penicillamine and specific hepatic copper-binding proteins was studied in vitro. RESULTS: D-penicillamine when given to either healthy or diseased animals prevented or reversed hepatitis, respectively. The drug particularly inhibited the disease-specific accumulation of copper in lysosomes of hepatocytes, tissue macrophages and Kupffer cells. When administered to diseased animals, the drug sequestered copper particularly from insoluble lysosomal particles. According to results obtained in vitro, the mobilization of this copper is likely to proceed through the solubilization of these particles. In contrast and as supported by the in vitro data, D-penicillamine had only a minor effect on copper bound to metallothionein in the cytosol. CONCLUSION: Our findings on the Long-Evans cinnamon rat provide some conclusions on the mechanism of action of D penicillamine in Wilson disease therapy. The drug prevents the formation or promotes the solubilization of copper-rich particles which occur in lysosomes of hepatocytes and Kupffer cells in the livers of patients with Wilson disease. Once chelated with D-penicillamine copper might then be excreted into urine. However, the mobilization of copper by D-penicillamine seems to be limited due to the binding of the metal to metallothionein in liver cytosol. This copper, even at relatively high concentrations, apparently may be well tolerated. PMID- 10707859 TI - Recurrence of esophageal varices following endoscopic treatment and its impact on rebleeding: comparison of sclerotherapy and ligation. AB - BACKGROUND/AIMS: Endoscopic variceal ligation is superior to sclerotherapy because of its lower rebleeding and complication rates. However, ligation is not without drawbacks due to a higher tendency to variceal recurrence. We conducted a randomized cohort study to delineate the long-term history of variceal recurrence following ligation and sclerotherapy, and to clarify the impact of recurrence on rebleeding and on the consumption of endoscopic treatment resources. METHODS: Two hundred cirrhotic patients with esophageal variceal bleeding were randomized to undergo maintenance endoscopic variceal sclerotherapy or ligation. RESULTS: One hundred and forty-one patients achieved variceal eradication and were regularly followed up for 2.2 to 6.7 (mean: 5.1 +/- 1.2) years. The demographic data, hepatic reserve, bleeding severity, and endoscopic features of both sclerotherapy (n=70) and ligation (n=71) showed no difference. Forty (57.1%) patients who underwent sclerotherapy experienced 58 recurrences of esophageal varices, in contrast to the 46 (64.8%) patients who underwent ligation and experienced 81 episodes of recurrence. Kaplan-Meier analysis showed that within 2 years variceal recurrence was more frequent for ligation than sclerotherapy, and the difference decreased thereafter. Multiple recurrence appeared more common with ligation (1/2/3/4/5 episodes of recurrence: 46/23/8/3/1 vs. 40/14/3/1/0, p=0.08). On multifactorial analysis, the endoscopic treatment method and red wale markings were the two factors determining variceal recurrence. Rebleeding from recurrent esophageal varices was unusual and showed no difference between the two groups (7/58 vs. 6/81, p>0.05). Rebleeding from gastric varices was more common after eradication by sclerotherapy (7/19 vs. 1/16, p=0.085) than by ligation. The number of sessions required for eradication of recurrent varices was no different between the two groups. CONCLUSIONS: Early recurrence and multiple recurrence of esophageal varices are more likely in patients undergoing endoscopic ligation, compared to sclerotherapy; however, the recurrence did not lead to a higher risk of rebleeding or require more endoscopic treatment. PMID- 10707860 TI - Prevalence of diabetes mellitus in patients with end-stage liver cirrhosis due to hepatitis C, alcohol, or cholestatic disease. AB - BACKGROUND/AIMS: The aims were to study: 1) the prevalence of diabetes mellitus in patients with end-stage liver cirrhosis due to hepatitis C, alcohol, or cholestatic liver disease, 2) viral and host immunogenetic factors that may predispose to diabetes, and 3) liver transplantation outcome in patients with or without diabetes. METHODS: Fasting blood glucose values of patients who underwent liver transplantation because of hepatitis C-related cirrhosis (73 patients) were compared with those of patients with cirrhosis due to cholestatic (78 patients) or alcoholic liver disease (53 patients) and to a general population. Data on diabetes prevalence in a population without liver cirrhosis was based on the prevalence of diabetes in Olmsted County, Minnesota, residents. HLA was determined using serologic assays. Hepatitis C virus genotypes were determined with polymerase chain reaction amplification and direct sequencing. Hepatitis G RNA was detected with polymerase chain reaction. Liver transplantation outcome in patients with or without diabetes was determined with rejection, retransplantation, or death at 1 year after transplantation as end points. RESULTS: Of 64 patients with hepatitis C alone, 16 (25%) had diabetes before transplantation compared with 1 of 78 (1.3%) with cholestatic liver disease (p= 0.0001) and 10 of 53 (19%) with alcoholic liver disease (p=0.36). Nine patients had hepatitis C plus cholestatic liver disease; one of these (11%) had diabetes. The prevalence of diabetes in patients with cholestatic liver cirrhosis was not different from that of the general population. The frequency of hepatitis G virus coinfection, HLA-DR3, or HLA-DR4 in hepatitis C and diabetes was not different from that of hepatitis C alone. The distribution of hepatitis C virus genotype was similar in those with and those without diabetes. Diabetes was not associated with increased risk of rejection, retransplantation, or death at 1 year after transplantation, and had no impact on overall survival after transplantation. CONCLUSIONS: 1) The risk of diabetes is not increased in patients with liver cirrhosis due to cholestatic liver disease but is in patients with liver cirrhosis due to hepatitis C or alcoholic liver disease; 2) cofactors (age, sex, body mass index, hepatitis G virus coinfection, hepatitis C virus genotype, or HLA-DR3/DR4) did not explain the increased risk of diabetes in patients with hepatitis C; 3) diabetes before liver transplantation did not change the outcome at 1 year after transplantation or survival. PMID- 10707861 TI - Oncostatin M: a cytokine upregulated in human cirrhosis, increases collagen production by human hepatic stellate cells. AB - BACKGROUND/AIMS: Hepatic stellate cells are predominantly responsible for the increased extracellular matrix seen in cirrhosis. The cytokine oncostatin M has been implicated in fibrogenesis in vitro in other cell types and in vivo in other tissues, although its effect on hepatic stellate cells or in cirrhosis is unknown. METHODS: To examine the effect of oncostatin M on collagen production by human hepatic stellate cells in culture, collagen protein was measured and collagen alpha2(1) mRNA was quantified by Northern analysis. Tissue inhibitor of metalloproteinase-1 (an inhibitor of collagen degradation) mRNA was measured in response to oncostation M stimulation. To explore the potential biological significance of this work to human liver disease, oncostatin M messenger RNA in normal and cirrhotic human liver was measured. RESULTS: Oncostatin M induced in a 2-fold increase in collagen secretion. The potency of induction of collagen protein secretion was equal to that observed after transforming growth factor beta stimulation. An increase in endogenous collagen alpha2(1) mRNA could not be detected. This suggested a post-transcriptional mechanism for the increase in collagen protein. In response to oncostatin M stimulation, there was a 2-fold increase in the tissue inhibitor or metalloproteinase-1 mRNA. Oncostatin M mRNA was detected in 6/6 cirrhotic livers and 1/7 normal livers after 28 PCR cycles. CONCLUSION: These results suggest that oncostatin M expression is upregulated in cirrhosis where it may have a role as a profibrogenic cytokine in hepatic stellate cells. PMID- 10707862 TI - Phosphatidylinositol-3 kinase and extracellular signal-regulated kinase mediate the chemotactic and mitogenic effects of insulin-like growth factor-I in human hepatic stellate cells. AB - BACKGROUND/AIM: Several studies have shown that proliferation of hepatic stellate cells is stimulated by insulin-like growth factor-I. The aim of this study was to investigate the effect of insulin-like growth factor-I on human hepatic stellate cells chemotaxis and the intracellular pathways involved in both mitogenic and chemotactic effects. METHODS/RESULTS: Insulin-like growth factor-I, at the concentration of 100 ng/ml, was able to induce a 2- to 3-fold increase in human hepatic stellate cells migration in a modified Boyden chamber system. This effect was associated with a marked activation of phosphatidylinositol 3-kinase by insulin-like growth factor-I, as evaluated by measurement of phosphatidylinositol 3-kinase activity in phosphotyrosine immunoprecipitates In order to establish a functional link between these observations, we then performed experiments employing two selective phosphatidylinositol 3-kinase inhibitors, namely wortmannin and LY294002. These compounds blocked activation of phosphatidylinositol 3-kinase and inhibited insulin-like growth factor-I-induced hepatic stellate cells migration. Since phosphatidylinositol 3-kinase activation has been shown to be necessary for platelet-derived growth factor-induced mitogenesis in hepatic stellate cells, we verified the effects of phosphatidylinositol 3-kinase inhibition on insulin-like growth factor-I-induced DNA synthesis. Incubation with either wortmannin or LY294002, dose-dependently reduced the mitogenic potential of insulin-like growth factor-I. Since phosphatidylinositol 3-kinase is involved, at least in part, in the activation of the Ras/extracellular signal-regulated kinase pathway in hepatic stellate cells, the role of extracellular signal-regulated kinase activation in mediating the biological effects of insulin-like growth factor-I was explored. Insulin-like growth factor-I induced mitogenesis and chemotaxis were markedly reduced by pre incubation of hepatic stellate cells with PD-98059, a selective inhibitor of MEK. CONCLUSIONS: Activation of phosphatidylinositol 3-kinase and extracellular signal regulated kinase is required for both insulin-like growth factor-I-dependent hepatic stellate cells proliferation and chemotaxis. Insulin-like growth factor I, together with other soluble mediators, may contribute to the hepatic wound healing response by modulating hepatic stellate cells migration and proliferation. PMID- 10707863 TI - Telomerase activity of needle-biopsied liver samples: its usefulness for diagnosis and judgement of efficacy of treatment of small hepatocellular carcinoma. AB - BACKGROUND/AIMS: High values for telomerase activity in malignant tumors have been reported. The clinical usefulness of measurements of telomerase activity as a diagnostic tool and to evaluate treatment efficacy in small hepatocellular carcinoma was investigated. METHODS: We investigated 22 patients (26 nodules) with intrahepatic abnormal nodules < or =20 mm in size determined by abdominal ultrasound. All underwent needle biopsies of tumorous nodules and extranodular regions of the liver by ultrasound guidance for histopathological diagnosis and measurement of telomerase activity by the fluorescence-based telomeric repeat amplification protocol. Re-biopsy of the same nodule was performed 1 week after percutaneous ethanol injection therapy to measure telomerase activity in 10 patients (10 nodules) found to have hepatocellular carcinoma. Liver-biopsied samples from 30 patients with chronic hepatitis C were used as a control. RESULTS: Telomerase activity increased with statistical significance stepwise: chronic hepatitis (n=30, mean: 0.00 U) extranodular regions (pre-cirrhosis or cirrhosis, n=22, mean: 1.80 U), atypical hyperplasia (borderline or premalignant lesions, n= 15, mean: 7.02 U) and low-grade malignant hepatocellular carcinoma (n=11, mean: 31.96 U) (p<0.0001 by the Kruskal-Wallis test). Percutaneous ethanol injection therapy resulted in loss (0.00 U) of telomerase activity in 9 nodules and persistence in 1 nodule. CONCLUSIONS: Measurement of telomerase activity appeared useful for diagnosis of intrahepatic abnormal nodules and assessment of the efficacy of percutaneous ethanol injection therapy and may be used as an alternative diagnostic method, especially when pathohistological discrimination between atypical hyperplasia and well-differentiated hepatocellular carcinoma is difficult. PMID- 10707864 TI - Fibronectin regulates morphology, cell organization and gene expression of rat fetal hepatocytes in primary culture. AB - BACKGROUND/AIMS: The extracellular matrix regulates hepatic development and regeneration, modulating the maintenance of liver architecture in the differentiated state. The aim of this work was to analyze how different extracellular matrix molecules modulate fetal hepatocyte morphology, growth and differentiation. METHODS: We cultured fetal hepatocytes either on plastic or on different extracellular matrix proteins, i.e., collagen I, fibronectin or E-C-L (entactin-collagen IV-laminin) and we analyzed cell attachment, morphological organization, proliferative response and gene expression. RESULTS: Cell attachment was increased by all the extracellular matrix proteins to a similar extent. However, only fibronectin facilitated the formation of elongated cord like structures, reminiscent of liver plate organization. Immunocytochemical analysis of the cells in these structures revealed high levels of albumin and cytokeratin 18, phenotypical markers of parenchymal hepatocytes. Fibronectin did not block the mitogenic stimuli induced by epidermal growth factor in these cells and the elongated structures appeared either in the absence or in the presence of the mitogen. Cells cultured on fibronectin, regardless of whether epidermal growth factor was present or not, also presented the maximal levels of expression for liver specific genes, such as albumin or alpha-fetoprotein. This expression was coincident with an increased expression of hepatocyte nuclear factor (HNF)-4 and a higher HNF-1alpha/HNF-1beta ratio, when compared with those cells that were cultured on collagen or E-C-L extracellular matrix. CONCLUSIONS: These results suggest that fibronectin might play a differential role, as compared to other extracellular matrix proteins, in fetal hepatocyte organization and gene expression. PMID- 10707865 TI - Differential regulation of activin A for hepatocyte growth and fibronectin synthesis in rat liver injury. AB - BACKGROUND/AIMS: Both hepatocyte growth and production of extracellular matrix such as fibronectin are essential for liver regeneration. Although activin A is reported to inhibit DNA replication in rat hepatocytes, the role of activin A for liver regeneration after acute injury has not been fully assessed. This study investigated the mechanism by which hepatocyte growth is regulated by activin A during liver regeneration and the effects of activin A on extracellular matrix production. METHODS: The mRNA for betaA subunit of activin A and activin receptors in hepatocytes and hepatic stellate cells after CCl4 administration were studied by Northern blotting. Binding of 125I-activin A was tested in these cells. Effects of activin A were examined by DNA, collagen and fibronectin synthesis. RESULTS: betaA mRNA was expressed in quiescent hepatocytes, and this expression peaked 12 h after CCl4 administration. Activin receptor mRNAs and cross-linked ligand/receptor complexes were expressed in hepatocytes and hepatic stellate cells However, these levels decreased specifically in hepatocytes at 24 h and had normalized by 72 h. The down-regulation of activin receptor was also observed after partial hepatectomy. Antiproliferative response to activin A decreased in hepatocytes at 24 h. Activin A stimulated production of fibronectin by hepatic stellate cells, but the synthesis of collagen was only slightly elevated in hepatic stellate cells following activin stimulation. CONCLUSIONS: The down-regulation of activin receptors in hepatocytes may be partly responsible for these cells becoming responsive to mitogenic stimuli. The increase of activin A at the early stage of liver injury has the potential to contribute to the regulation of fibronectin production in hepatic stellate cells. PMID- 10707866 TI - Hepatic artery embolisation with a novel radiopaque polymer causes extended liver necrosis in pigs due to occlusion of the concomitant portal vein. AB - BACKGROUND/AIM: In an attempt to overcome some of the problems encountered with the materials available for liver embolisation, we investigated a novel radiopaque polymer of the polyurethane family (Degra-Bloc). METHODS: Hepatic artery embolisation of one liver lobe using polyurethane was performed in 19 healthy pigs. Microcirculatory changes were assessed by laser Doppler flowmetry. Radiological and pathological examinations of the livers, hearts and lungs removed provided information about the extent and effect of the embolisation. RESULTS: None of the pigs died due to hepatic failure or toxicity of polyurethane. Microcirculation of embolised liver lobes significantly decreased from 106 (+/-15) perfusion units (PU) to 45 (+/-6) PU immediately after embolisation and further to 28 (+/-7) PU before euthanasia. At this time conventional and angiographic X-ray controls demonstrated the radiopaque casts extending up to the peripheral arteries with signs of degradation over time but without formation of collateral vessels. The main pathological findings consisted of destruction of the portal tract structures and also of large areas of liver necrosis. Polyurethane was encountered in arterioles as small as 10-20 microm, but not in liver sinusoids, hearts or lungs. CONCLUSIONS: The novel polymer called DegraBloc is a biocompatible, slowly degradable, radiopaque embolic agent. The occlusion of the arterial tree up to the smallest arteriolar diameter combined with concomitant portal vein occlusion leads to sharp segmental necrosis in pig livers without formation of significant collaterals and without systemic embolism. In the treatment of liver tumours polyurethane might provide a promising alternative to conventional embolic materials, provided that it is used with care in patients with advanced liver cirrhosis. PMID- 10707867 TI - Prospective analysis of risk factors for early intrahepatic recurrence of hepatocellular carcinoma following ethanol injection. AB - BACKGROUND/AIM: Time-dependent intrahepatic recurrence of hepatocellular carcinoma is frequent after different treatment modalities, including percutaneous ethanol injection. We attempted to prospectively analyze the possible risk factors for early intrahepatic recurrence of hepatocellular carcinoma after percutaneous ethanol injection. METHODS: Sixty-five patients with 65 solitary hepatocellular carcinoma nodules < or =6 cm in diameter underwent initial treatment with percutaneous ethanol injection and were examined to ascertain the factors related to recurrence, local and distant, within the liver. A number of clinical and tumor parameters were analyzed. RESULTS: Cumulative overall recurrence rates 12 and 24 months after percutaneous ethanol injection were 15.6% and 45.1%, respectively, irrespective of clinical and tumor parameters. Overall recurrence rates 12 and 24 months after percutaneous ethanol injection were 40% and 67.5%, for tumor > or =3 cm and 7.5% and 37.5%, for tumor <3 cm. Cumulative local recurrence rates at 12 and 24 months were 26.3% and 43.5%, respectively, for tumor > or =3 cm and 11.7% and 18.2%, respectively, for tumor <3 cm. The log-rank test indicated that a tumor size of > or =3 cm and the presence of capsule for a tumor of <3 cm in diameter were significant risk factors for intrahepatic recurrence. A pretreatment serum PIVKA-II level of > or =0.02 AU/ml was the only clinical parameter associated with overall recurrence (p=0.0041) and distant intrahepatic recurrence (p=0.0307). Distant intrahepatic recurrence rates 12 and 24 months after percutaneous ethanol injection were 22.5% and 31.4%, respectively, for PIVKA-II levels of > or =0.02 AU/ml and 8% and 17.8%, for PIVKA-II of <0.02 AU/ml. Cox's proportional hazard model identified that tumor size, tumor capsule and baseline serum PIVKA-II levels were independently related to intrahepatic recurrence. CONCLUSIONS: These data demonstrate that tumor size and peritumoral capsule were associated with overall and local recurrence of hepatocellular carcinoma. Moreover, pretreatment serum levels of PIVKA-II can indicate the risk of early intrahepatic recurrence and may assist in patient selection and appropriate therapy. PMID- 10707868 TI - Combined gene therapy with suicide gene and interleukin-12 is more efficient than therapy with one gene alone in a murine model of hepatocellular carcinoma. AB - BACKGROUND/AIMS: Gene therapy has emerged as a new form of treatment for unresectable hepatocellular carcinoma (HCC). We evaluate here the effect of IL-12 and the suicide gene thymidine kinase as single agents and in combination to treat experimental liver cancer. METHODS: Recombinant adenoviruses expressing mouse interleukin-12 (AdCMVIL-12) or thymidine kinase of herpes simplex virus (AdCMVtk) or lacZ reporter gene (AdCMVlacZ) were constructed. The efficacy of the treatment was evaluated in a murine HCC model based on subcutaneous implantation of liver tumor cells (BNL). RESULTS: Transduction of BNL cells after in vitro infection with AdCMVlacZ was very low at multiplicity of infection (moi) of 100, whereas 10-15% of cells were transduced when using moi 1,000. Similarly, production of IL-12 was detectable only in BNL cells infected with AdCMVIL-12 at moi 1,000. In vitro infection of BNL cells with AdCMVIL-12 at moi 100 did not abrogate tumorigenicity, whereas moi 1,000 resulted in inhibition of tumor growth in all mice as well as in abrogation of tumor formation in 3 out of 8 animals. In vivo studies showed that intratumor injection of AdCMVIL-12 induced a dose dependent effect on tumor regression. However, none of the animals exhibited complete tumor elimination with this treatment. We observed that suppression of tumor growth was more intense in animals treated with the combination of AdCMVIL 12 plus AdCMVtk than in animals which received AdCMVtk or AdCMVIL-12 alone. The combined treatment resulted in a significant increase in animal survival, and 25% of treated animals were free of tumor for over 100 days without recurrence of the disease. CONCLUSIONS: Combination of AdCMVIL-12 and AdCMVtk is more efficient than either of the two vectors alone for the treatment of the murine model of HCC used in this study. PMID- 10707869 TI - Effect of preoperative interventions on outcome following liver resection in a rat model of cirrhosis. AB - BACKGROUND/AIMS: High morbidity and mortality rates in cirrhotic patients undergoing resections for hepatocellular malignancies underscore the need for identifying a therapy that will decrease fibrosis or enhance hepatic regenerative activity in the perioperative period. Thus, in the present study, 104 carbon tetrachloride-induced cirrhotic rats received either saline (untreated cirrhotic controls) or one of the following agents that have been reported to decrease hepatic fibrosis or increase hepatic regeneration; pentoxifylline, ciprofloxacin or a traditional Chinese herbal remedy (TCHR). Twelve additional rats served as healthy, non-cirrhotic controls. METHODS: Treatments were administered daily by gavage for 4 weeks followed by a 70% partial hepatectomy. Hepatic fibrosis was documented at the time of surgery by computer-assisted quantitation of collagen content. Liver function and hepatic regenerative activity were documented 24 h post partial hepatectomy by serum bilirubin determinations and a combination of 3[H]-Thymidine incorporation into hepatic DNA and proliferating cell nuclear antigen (PCNA) quantitation, respectively. RESULTS: Compared to untreated cirrhotic controls (8.1 +/- 0.7%), fibrosis was significantly reduced in the pentoxifylline- and ciprofloxacin-treated groups (4.6 +/- 0.2%, p<0.005 and 5.5 +/- 0.6%, p<0.05) but unchanged in the TCHR-treated group (6.6 +/- 11.0%). Post operatively, total serum bilirubin levels were lower in the pentoxifylline (1.40 +/- 0.15 mg/dl,p<0.01) and ciprofloxacin (1.87 +/- 0.25 mg/dl, p<0.05)-treated groups, but unchanged in the TCHR group (2.20 +/- 0.45 mg/dl), when compared to untreated cirrhotic controls (3.00 +/- 0.37 mg/dl). Hepatic regenerative activity was also significantly improved in the pentoxifylline-treated group (17.8 +/- 2.2 versus 9.9 +/- 1.9 DPM/microg DNA in untreated cirrhotic controls, p<0.05), but unchanged in the ciprofloxacin (16.1 +/- 1.8 DPM/microg DNA) and TCHR (10.9 +/- 1.2 DPM/microg DNA)-treated groups. PCNA protein determinations were in keeping with the 3[H]-Thymidine results CONCLUSIONS: Pre-operative pentoxifylline holds promise as a useful therapeutic intervention for patients with cirrhosis requiring hepatic resection. PMID- 10707870 TI - Decrease in serum ALT and increase in serum HCV RNA during pregnancy in women with chronic hepatitis C. AB - BACKGROUND/AIMS: The natural history of chronic hepatitis C infection during pregnancy has not been clearly established, and thus our aim was to assess serum alanine aminotransferase levels and serum HCV RNA levels during pregnancy. METHODS: Twenty-six pregnant women with chronic hepatitis C were studied. Serum alanine aminotransferase was assessed within the 3 months before, monthly during and within the 3 months after pregnancy. In 12 women, serum HCV RNA levels were quantified by the branched DNA assay. Twenty-six age-matched non-pregnant women with chronic hepatitis C were followed up for 1 year, and used as a comparison group. RESULTS: During pregnancy, serum alanine aminotransferase levels decreased in the second and third trimesters. The third trimester levels were significantly lower than serum alanine aminotransferase levels before pregnancy (p=0.0001). Seventy-seven percent of the pregnant women with increased pre-pregnancy levels had normalization of serum alanine aminotransferase levels. In the second or third trimesters, serum HCV RNA levels increased. The third trimester serum HCV RNA levels were significantly higher than levels before pregnancy (p=0.01). No significant change in serum alanine aminotransferase or HCV RNA levels was observed in the control group. CONCLUSION: In pregnant women with chronic hepatitis C, serum alanine aminotransferase levels decrease, and serum HCV RNA levels increase during the second and third trimesters. PMID- 10707871 TI - Long-term follow-up of patients with anti-HBe/HBV DNA-positive chronic hepatitis B treated for 12 months with lamivudine. AB - BACKGROUND/AIMS: Interferon alpha provides benefit in only a limited number of patients with chronic anti-HBe-positive hepatitis B. The aim of this study was to verify the long-term efficacy of lamivudine treatment of these patients and the incidence of lamivudine-resistant hepatitis B virus mutants. METHODS: Fifteen consecutive patients with chronic anti-HBe-positive hepatitis B were treated with lamivudine 100 mg once daily for 52 weeks. Levels of alanine aminotransferase, HBV DNA, hepatitis B surface antigen, and IgM antibodies to hepatitis B core antigen were monitored during therapy and 12-month follow up. The polymerase gene was amplified by polymerase chain reaction and the region coding for YMDD amino acid motif was directly sequenced. RESULTS: Only 2/15 patients (13%) had a sustained virological and biochemical response and improved histologically. Eleven out of 15 (74%) showed inhibition of viral replication and normalization of alanine aminotransferase levels during lamivudine treatment but relapsed 1-12 months after terminating therapy. In the two remaining patients (13%), HBV DNA initially became negative but reappeared in the serum after 24 weeks, and in both patients the emergence of YMDD mutants was demonstrated. CONCLUSIONS: Our data confirm the antiviral efficacy of lamivudine in anti-HBe-positive patients, but response to a 1-year course was only transient as the majority of patients relapsed after therapy withdrawal. The lack of a sustained effect and the emergence of lamivudine-resistant mutants suggest that therapy for chronic hepatitis B should be based on a combination of several therapeutic agents. PMID- 10707872 TI - Persistence of viral replication after anti-HBe seroconversion during antiviral therapy for chronic hepatitis B. AB - BACKGROUND/AIMS: Hepatitis B virus genome mutants may be selected during the immune-mediated clearance of infection or during long-term nucleoside analog administration and may escape both antiviral pressures. The pattern of anti-HBe seroconversion was analyzed in patients receiving new nucleoside analogs, lamivudine or famciclovir, in comparison with patients treated with interferon alpha. METHODS: Eighteen consecutive patients who seroconverted to anti-HBe were included in the study. Serial serum samples were studied with the quantitative determination of HBV DNA by the branched DNA assay (Chiron) and by a quantitative PCR assay (Roche diagnostics), determination of pre-S1 Ag, the genetic analysis of the viral genome with the determination of pre-core promoter or pre-core region mutations with a line probe assay (Innogenetics) and, in selected samples of polymerase gene mutations. RESULTS: The quantitative PCR assay was found to be more sensitive than the bDNA assay, allowing a 25-log decrease in viral DNA levels to be demonstrated after anti-HBe seroconversion. Viral persistence after anti-HBe seroconversion induced by interferon, lamivudine or famciclovir, was often associated with circulating HBV genomes harboring mutations in the precore promoter. The clinical significance of these findings was demonstrated by the observation of reversion to HBeAg in two patients treated with interferon and one with lamivudine. CONCLUSION: Persistence of significant levels of viremia that are not detected by the branched DNA assay may be observed after anti-HBe seroconversion. A precise monitoring of viremia levels with more sensitive assays and HBV mutant strains is warranted in patients undergoing antiviral therapy. PMID- 10707873 TI - A phase I/II study of recombinant human interleukin-12 in patients with chronic hepatitis B. AB - BACKGROUND/AIMS: Interleukin-12 (IL-12) may be active against hepatitis B virus (HBV). The objective of the study was to assess the tolerability, activity, pharmacokinetics, and pharmacodynamics of three dose levels (0.03 microg/kg b.w., n=15; 0.25 microg/kg b.w., n=15; 0.50 microg/kg b.w., n=16) of recombinant human (rHu) IL-12 given s.c. once a week for 12 consecutive weeks. METHODS: Forty-six patients with chronic hepatitis B, HBV DNA positivity and aminotransferase elevation were included in a multicenter prospective randomized phase I/II study. RESULTS: Compared with the baseline, HBV DNA levels had decreased significantly at the end of rHuIL-12 treatment and after the 12-week follow-up period (p<0.001). The response to rHuIL-12 treatment was dose-dependent: at the end of the study HBV DNA clearance was greater in patients treated with 0.50 microg/kg b.w. (25%) or with 0.25 microg/kg b.w. (13%) compared with those given 0.03 microg/kg b.w. (7%). Moreover, HBeAg became undetectable at the end of follow-up in five of the patients given the 0.25microg/kg (2/15) or the 0.50 microg/kg (3/16) dose. The drug pharmacology showed that IL-12 had an estimated half-life of 30 h with levels remaining detectable for more than 48 h after rHuIL-12 administration. The serum levels of IL-12, interferon-gamma, IL-10, neopterin and beta2-microglobulin as well as the area under the curve (AUC) were rHuIL-12 dose related. Side effects were observed more frequently with higher doses, including moderate decreases in lymphocyte and neutrophil counts; three patients withdrew prematurely from treatment. The local reaction observed at the injection site was unrelated to the drug dose. Only one patient showed detectable antibody levels to rHuIL-12 without clinical impact. CONCLUSIONS: Treatment with rHuIL-12 at the doses investigated is safe and tolerable, and appears to be active against HBV in patients with chronic hepatitis B. PMID- 10707874 TI - Basement membrane peptides as markers of liver disease in chronic hepatitis C. AB - BACKGROUND/AIM: Chronic hepatitis C is characterised by slow progression to liver fibrosis. In liver fibrosis, basement membrane components are increasingly deposited around the vessels and in the portal tracts. Serum assays can measure the two major components of the basement membrane, type IV collagen and laminin. The aim of this study was to determine whether serum levels of type IV collagen and laminin are related to severity of liver injury in chronic hepatitis C. METHODS: Thirty-seven patients with chronic hepatitis C (CHC) and five healthy controls were studied. Serum type IV collagen was measured by a one-step sandwich EIA kit (Fuji, Japan) and serum laminin was measured by RIA (CIS, UK). Liver biopsies in patients with CHC were scored using a previously described grading and staging system. Liver biopsy scores were compared to serum levels of laminin, type IV collagen and alanine aminotransferase (ALT). Receiver operating characteristic (ROC) analysis was used to compare the ability of the assays to detect advanced liver injury. RESULTS: The median serum concentration of type IV collagen was 127.1 ng/ml (range 17.7 to 317.4) in CHC patients compared to a median of 61.3 ng/ml (range 11.5 to 102.3) in controls, p=0.006. The median serum concentration of laminin was 1.12 U/ml (range 0.74 to 2.46) in CHC compared to a median of 0.87 U/ml (range 0.83 to 1.06) in controls, p=0.07. Both serum type IV collagen and laminin were significantly correlated with the fibrotic stage and also with the necroinflammatory injury scores- histological activity index, portal inflammation and periportal hepatitis. Serum ALT was significantly correlated with portal inflammation. Using ROC analysis, the area under the curve for type IV collagen and laminin was 0.83 (p=0.001) and 0.82 (p=0.0017), respectively, while the area under the curve for ALT was 0.54 (p=0.1). CONCLUSIONS: Serum assays of basement membrane peptides are accurate non-invasive markers of liver fibrosis and liver inflammation in chronic hepatitis C. These markers are superior to serum ALT in reflecting liver injury and they have high specificity and sensitivity in detecting advanced liver disease in chronic hepatitis C. PMID- 10707875 TI - Intrahepatic accumulation of nitrotyrosine in chronic viral hepatitis is associated with histological severity of liver disease. AB - BACKGROUND/AIMS: The toxicity of nitric oxide is thought to be engendered, at least in part, by its reaction with superoxide yielding peroxynitrite, a potent oxidant that promotes the formation of nitrotyrosine within cells and tissue lesions. In this study we assessed the intrahepatic localization and distribution of the inducible nitric oxide synthase (iNOS) and nitrotyrosine (NTY) in patients with viral and non-viral liver disease. METHODS: We carried out single and double immunostaining experiments on cryostat liver biopsy sections using monoclonal antibodies against iNOS and NTY. We also performed a comparative analysis between the intrahepatic immunostaining score of NTY and the histological activity index of chronic viral hepatitis. RESULTS: We found a marked hepatocellular expression of iNOS with a diffuse lobular pattern in all liver samples from patients with viral liver disease, whereas NTY localization was mainly restricted to cellular foci consisting of hepatocytes and Kupffer cells. Interestingly, we demonstrated by means of double immunostaining experiments the existence of hepatocellular co localization of iNOS and NTY in the majority of NTY-expressing liver cells. The amount of NTY was significantly higher in liver biopsies from viral liver disease than in non-viral liver disease. In addition, a statistically significant association between the intrahepatic amount of NTY and the severity of viral liver disease was found. CONCLUSIONS: Nitric oxide-mediated nitration of hepatocellular proteins is markedly induced in the inflamed liver tissue from patients with chronic viral hepatitis, and appears to be associated with the histological severity of viral chronic liver disease. PMID- 10707876 TI - TT virus infection in patients with chronic hepatitis C virus infection--effect of primers, prevalence, and clinical significance. Hepatitis Interventional Therapy Group. AB - BACKGROUND/AIM: A novel DNA virus, TT virus (TTV), was recently identified in patients with post-transfusion non-A-G hepatitis. The aim of this study was to determine the prevalence and clinical significance of TTV infection in patients with chronic hepatitis C virus (HCV) infection. METHODS: We analyzed pretreatment serum samples from 171 United States and European patients who relapsed after interferon-alpha treatment and were recruited into an interferon-alpha 2b/ribavirin combination treatment trial. TTV DNA was detected by PCR using two different set of primers (TTV-A and TTV-B) derived from open reading frames 1 and 2, respectively. RESULTS: TTV was detected in 29.2% of the patients with the TTV A primer set, 70.8% with the TTV-B primer-set, and 72.5% if positive by either/both sets of the primers. The amplicons generated by primer set A were sequenced and a phylogenetic tree was constructed. The 50 isolates belonged to group la (n=8), 1b (n=17), 2a (n=21), 2b (n=3), and 4 (n=1). There was no difference in demographic (age, sex distribution, estimated duration of HCV infection), biochemical (serum ALT levels), virologic (serum HCV RNA levels, HCV genotype distribution), or histologic scores, and their subsequent response to either interferon-alpha-2b or interferon-alpha-2b/ribavirin combination treatment. CONCLUSIONS: The prevalence of TTV infection reported previously may have been significantly underestimated, based on the primers originally described and used by most studies. Although TTV infection is very common in patients with chronic HCV infection, it has no identifiable clinical significance. PMID- 10707877 TI - The potential of gene therapy in the treatment of hepatocellular carcinoma. PMID- 10707878 TI - Neoadjuvant photodynamic therapy before curative resection of proximal bile duct carcinoma. AB - BACKGROUND: Hilar bile duct carcinoma has an 80% probability of local recurrence after curative resection, which might be reduced if neoadjuvant photodynamic therapy is feasible. CASE AND TREATMENT: After intravenous injection of sodium porfimer we treated an adenocarcinoma of the proximal common bile duct (T2 N0 M0, Bismuth type II) in a 72-year-old man with red laser light (applied from the lumen at a dose 250 Joules/cm2), and the adjacent right and left hepatic and common bile duct at a dose of 125 Joules/cm2. After 23 days the tumor was completely resected (adenocarcinoma pT2 pNO; G2). RESULTS: In the lumenal, 4-mm thick layer the bile duct specimen exhibited complete tumor necrosis with pigmentation of photodegraded porfimer and no viable tumor cells, while in the outer layer of the wall (at 5-8-mm depth) viable cancer cell nests without degraded porfimer were seen. The bile duct tissue showed little damage. Eighteen months after surgery, neither tumor recurrence nor stricture formation was found at the pretreated bilioenteric anastomoses. CONCLUSIONS: a) Photodynamic therapy with sodium porfimer seems to be confined to the superficial 4-mm layer of bile duct cancer. b) Neoadjuvant photodynamic therapy is feasible for hilar bile duct carcinoma. PMID- 10707879 TI - Polymyalgia rheumatica as a manifestation of a large hepatic cavernous hemangioma. AB - A 59-year-old woman presented with polymyalgia rheumatica which was refractory to conventional anti-inflammatory and steroid therapy. A full investigation for an underlying occult malignancy showed only the presence of a giant cavernous hepatic hemangioma. To our knowledge, polymyalgia rheumatica has never been described in association with giant cavernous hepatic hemangioma; resection of the latter lesion resulted in complete and, to this date, definite resolution of rheumatologic complaints in our patient. PMID- 10707880 TI - Images in hepatology. Intrahepatic bleeding due to undifferentiated (embryonal) hepatic sarcoma. PMID- 10707881 TI - Rapid screening of the Cys282Tyr and His63Asp mutations in the HFE gene. PMID- 10707882 TI - HLA-A2 transgenic mouse model: potential utility for development of an HCV vaccine. PMID- 10707883 TI - Fatal lactic acidosis and liver steatosis associated with didanosine and stavudine treatment: a respiratory chain dysfunction? PMID- 10707884 TI - Discovery of a cholecystokinin-releasing peptide: biochemical characterization and physiological implications. AB - Recent studies indicate that the secretion of CCK is mediated by a trypsin sensitive peptide secreted by the proximal small intestine that has been designated "CCK-releasing factor" (CCK-RF). This CCK-RF was found to be identical to the porcine diazepam binding inhibitor by peptide sequencing and mass spectrometry analysis. This peptide is present in abundance in the epithelial cells in the duodenal mucosa. Its release into the lumen is mediated by intestinal submucosal cholinergic neurons. Functionally, this peptide appears to mediate feedback regulation of pancreatic secretion and CCK release in response to peptone and lipid stimulation. It fulfills all the criteria as a physiological regulator of CCK secretion. This represents the first chemical characterization of a luminally secreted enteric peptide functioning as an intraluminal regulator of intestinal hormone release. PMID- 10707885 TI - Hypothesis--origin of parietal cells: transfer of the H+K+-ATPase gene from parasitic microorganisms to Cnidaria? AB - Parietal cells present in the stomach and terminal ileum secrete a highly concentrated hydrochloric acid into the lumen. The cells are characterized by the enzyme P-type H+K+-ATPase, which has an alpha-subunit with a high homology (>85%) for the amino acid sequences of frog, mouse and pig stomachs. Gastric H+K+-ATPase also exhibits a high homology to H+-ATPase in yeast and Na+K+-ATPase in many tissues, suggesting origination from a common ancestral ATPase. It is known that parietal cells first appeared in fish and were later expressed in evolutionarily higher organisms. Primitive organisms, such as Cnidaria and Ctenophora, that possessed digestive organs, but not parietal cells, were abundant in the ocean more than 600 million years ago (Pre-Cambrian period). The author thus hypothesized that the genes of either H+-ATPase or H+K+-ATPase that were present in parasitic microorganisms, such as yeast, were transferred to the interstitial cells of host organisms, such as Cnidaria, eventually leading to the evolution of parietal cells. It appears that although parietal cells in the stomach developed by chance, such cells have greatly contributed to the evolution of advanced organisms, including humans, by affording safe ingestion of a large volume of various foods. PMID- 10707886 TI - Effects of endothelin-1 on duodenal bicarbonate secretion and mucosal integrity in rats. AB - Effects of endothelin-1 on gastric acid secretion, duodenal HCO3- secretion, and duodenal mucosal integrity were investigated in anesthetized rats, in comparison with those of TY-10957, a stable analogue of prostacyclin. A rat stomach mounted on an ex-vivo chamber or a proximal duodenal loop was perfused with saline, and gastric acid or duodenal HCO3- secretion was measured using a pH-stat method and by adding 100 mM NaOH or 10 mM HCl, respectively. Duodenal lesions were induced by mepirizole (200 mg/kg) given subcutaneously. Intravenous administration of endothelin-1 (0.6 and 1 nmol/kg) caused an increase of duodenal HCO3- secretion with concomitant elevation of blood pressure; this effect was antagonized by co administrahon of BQ-123 (ET(A) antagonist; 3 mg/kg, i.v.) and significantly mitigated by vagotomy. Likewise, endothelin-1 caused a significant decrease in histamine-stimulated acid secretion, and this effect was also significantly antagonized by BQ-123. Although TY-10957 (10 and 30 mg/kg, i.v.) produced a temporal decrease of blood pressure, this agent caused not only an increase of duodenal HCO3- secretion, independent of vagal nerves, but also a decrease of acid secretion as well. In addition, both endothelin-1 and TY-10957 significantly prevented mepirizole-induced duodenal lesions at the doses that caused an increase of duodenal HCO3- secretion and a decrease of gastric acid secretion. These results suggest that endothelin-1 affects the duodenal mucosal integrity by modifying both gastric acid and duodenal HCO3- secretions, the effects being mediated by ET(A) receptors. PMID- 10707887 TI - Interactions of EGF and ornithine decarboxylase activity in the regulation of gastric mucus synthesis in cigarette smoke exposed rats. AB - Cigarette smoking has been shown to aggravate ulceration and delay ulcer healing. Smokers had a lower level of mucus in their stomachs. In the present study, we examined whether cigarette smoke or its extract reduced mucus production through the suppression of epidermal growth factor (EGF) associated with the reduction of polyamine biosynthesis both in vivo and in vitro. Ornithine decarboxylase (ODC) activities and mucus synthesis were determined in rat gastric mucosa and in human MKN-28 cells. Incubation of MKN-28 cells with EGF (0.01-1.00 ng/mL) significantly increased mucus synthesis in vitro, which was accompanied by an increase of ODC activity. Removal of salivary glands decreased the circulated EGF level and induced a significant reduction of mucus-secreting layer thickness in the gastric mucosa. Cigarette smoke or its extract markedly decreased mucus synthesis in vivo and in vitro, both of which could be completely reversed by intravenous administration of EGF (20 microg/kg) in rats or co-incubation with EGF (1 and 2 ng/mL) in MKN-28 cells. However, ODC activities, which were suppressed by cigarette smoke or its extract, were unaffected by intravenous administration of EGF in rats, or only partially reversed by co-incubation with EGF in MKN-28 cells. These findings indicate that both EGF and ODC activity represent two different entities in the modulation of cigarette smoking on gastric mucus synthesis. The action of EGF on mucus synthesis may only be partially if not dependent on ODC activity in the stomach. PMID- 10707888 TI - Role of gastric microcirculation in the gastroprotection by glucocorticoids released during water-restraint stress in rats. AB - Our previous investigations demonstrated that glucocorticoids released in response to stress protect gastric mucosa against stress-induced ulceration. This study was designed to determine whether gastric microcirculation is involved in the mechanism of gastroprotective glucocorticoid action. For this we evaluated the effects of deficiency of glucocorticoid production during 3 hr water restraint stress and corticosterone replacement on the stress-induced gastric erosions, gastric microcirculation and arterial pressure in rats. The stress was produced in awake rats and gastric microcirculation and arterial pressure were evaluated in animals anesthetized in 3 hr after the onset of water-restraint stress. An in vivo microscopy technique for the direct visualization of gastric microcirculation was employed. The gastric submucosal and the superficial mucosal microvessels were monitored on television screen through a microscope and the pictures were stored by microfilming for the analysis of red blood cell velocity and vessel diameter. Gastric microcirculation was estimated on the base of both the volume blood flow velocity in submucosal microvessels and the diameter of superficial mucosal venous microvessels. Gastric erosions were quantitated by measuring the area of damage. Plasma corticosterone levels were also measured after 3 hr stress by fluorometry. Water-restraint stress induced an increase in corticosterone level, an appearance of gastric erosions, a decrease in volume blood flow velocity of submucosal microvessels, a dilatation of superficial mucosal microvessels, a decrease in arterial pressure. The deficiency of glucocorticoid production during water-restraint stress promoted the stress induced gastric ulceration, a dilatation of mucosal microvessels, a decrease of blood flow velocity in submucosal microvessels and of arterial pressure. Corticosterone replacement eliminated the effects of deficiency of glucocorticoid production on all of the parameters under study. Thus, the stress-induced corticosterone rise decreased gastric ulceration, restricted both the reduction of blood flow velocity in submucosal microvessels and a dilatation of superficial mucosal venous microvessels during water-restraint stress. These data suggest that the gastroprotective action of glucocorticoids during stress may be provided by the maintenance of gastric blood flow. PMID- 10707889 TI - N(omega)-nitro-L-arginine decreases resting cytosolic [Ca2+] and enhances heat stress-induced increase in cytosolic [Ca2+] in human colon carcinoma T84 cells. AB - N(omega)-nitro-L-arginine (LNNA) inhibits the synthesis of heat shock proteins in animals and cultured cells exposed to heat stress. Heat shock protein synthesis is known to be Ca2+-dependent. In this study, we have characterized the effect of LNNA on [Ca2+]i before and after heat stress in human colon carcinoma T84 cells. In untreated cells incubated in the presence of external Ca2+, the resting [Ca2+]i was 201+/-3 nM. If these cells were exposed to 45 degrees C for 10 min, [Ca2+]i increased by 50+/-2%. Preincubation with LNNA (100 microM) without subsequent heating led to a decrease in [Ca2+]i in a LNNA concentration-dependent manner. Preincubation with LNNA followed by heating increased [Ca2+]i to levels 88+/-5% greater than cells heated without LNNA pretreatment. Incubating cells in medium without external Ca2+ (no heating, no LNNA treatment) lowered resting [Ca2+]i to 115+/-2 nM and greatly reduced the increase in [Ca2+]i observed if cells were heated in the presence of Ca2+, indicating that external Ca2+ plays an important role in the maintenance of [Ca2+]i in T84 cells. With external Ca2+ absent, LNNA pretreatment further reduced [Ca2+]i in unheated cells, and heating failed to enhance [Ca2+]i. We determined (with external Ca2+ present) that the heat-stress induced increase in [Ca2+]i in T84 cells was blocked by dichlorobenzamil, a Na+/Ca2+ exchanger inhibitor, suggesting that the exchanger mediates Ca2+ entry. The median inhibitory concentration (IC50) in cells not treated with LNNA was 0.970+/-0.028 microM. With LNNA pretreatment, the IC50 was 5.099+/-0.107 microM. Heat stress of T84 cells did not affect the binding affinity of the Na+/Ca2+ exchanger for external Ca2+, but it increased the maximal velocity of the exchanger. In unheated cells, preincubation with LNNA decreased the binding affinity of the exchanger for Ca2+, but after heat treatment, both the binding affinity and maximal velocity of the exchanger increased. Our data are consistent with the idea that LNNA affects the activity of the Na+/Ca2+ exchanger. We also determined there are intracellular Ca2+ pools in T84 cells sensitive to thapsigargin, monensin, and ionomycin. Treatment with TMB-8, a blocker of Ca2+ sequestration and mobilization, or ionomycin inhibited the LNNA-induced decrease in [Ca2+]i observed in the absence of external Ca2+, suggesting that LNNA promotes Ca2+ sequestration. PMID- 10707890 TI - Protective effects of several amino acid-nutrients on gastric hemorrhagic erosions in acid-irrigated stomachs of septic rats. AB - Our previous report demonstrates that severe gastric mucosal damage is produced in lipopolysaccharide (LPS)-intoxicated rats. In the present study, we examined protective effects of several amino acids including taurine, phenylalanine and L Arginine on gastric hemorrhagic erosions in acid-irrigated stomachs of LPS rats. The animals were deprived of food for 24 hr. Intravenous LPS (3 mg/kg) was challenged 12 hr after withdrawal of food. Gastric vagotomy was performed, followed by irrigation the stomachs for 3 hr with a physiological acid solution containing 100 mM HCl and 54 mM NaCl. The ulcerogenic parameters including increased gastric acid back-diffusion, mucosal histamine concentrations, lipid peroxide productions, luminal hemoglobin contents, stomach erosions and the lowered glutathione levels were markedly enhanced in LPS rat stomachs irrigated with acid solution. Both phenylalanine and taurine caused dose-dependent attenuations of these ulcerogenic parameters in LPS rats. L-arginine also was effective in inhibition. The inhibitory effect was restored by pretreatment of nitric oxide synthase inhibitors, such as N(G)-nitro-L-arginine-methyl ester or L N(G)-(1-iminoethyl)-lysine. Furthermore, marked amelioration of hemorrhagic erosions in LPS rats was observed when a combination of these amino acid nutrients was used. The results provide evidence that these amino acid nutrients may ameliorate gastric hemorrhagic erosion via GSH synthesis stimulation, histamine cell membrane stabilization and antioxidant actions in LPS rat stomachs. PMID- 10707891 TI - The effect of a novel pentadecapeptide BPC 157 on development of tolerance and physical dependence following repeated administration of diazepam. AB - A novel gastric pentadecapeptide BPC 157 with different beneficial activities and anticonvulsant effect interacting with GABAergic system could improve diazepam efficacy coadministered (10 microg/kg, 10 ng/kg i.p.) with diazepam (5.0 mg/kg i.p.) twice daily for 10 days, since diazepam chronic medication would otherwise predispose for diazepam- tolerance/withdrawal development (shorter latency to convulsion after convulsant). In diazepam chronically treated mice, it attenuated diazepam tolerance (provoked by later acute administration of diazepam together with convulsant) and postponed physical dependence/withdrawal effects (provoked by later administration of isoniazid). In tolerance assay, at 42 h after the end of conditioning regimen, shorter preconvulsive latencies than in healthy (non diazepam conditioned) mice following isoniazid (800 mg/kg i.p.) (as hallmark of tolerance) were observed if diazepam (5.0 mg/kg i.p.) was again given acutely to mice previously conditioned with diazepam alone (use of picrotoxin 3.0 mg/kg i.p., as convulsant, with acute application of diazepam in previously diazepam conditioned mice did not lead to tolerance hallmark). This was completely avoided in diazepam+BPC 157 10 microg or diazepam+BPC 157 10 ng chronically treated animals. In physical dependence assay (isoniazid challenge assessed at 6, 14, 42 and 72 h after conditioning medication), when compared to diazepam non conditioned healthy mice, in diazepam conditioned mice residual anticonvulsive activity was not present already at the earliest post-conditioning interval (i.e., not different latency to isoniazid-convulsions), whereas shorter preconvulsive latencies (as physical dependence/withdrawal hallmark) were noted in diazepam conditioned mice following isoniazid challenge at 42 h and at 72 h after end of conditioning treatment. In diazepam+BPC 157 10 microg- conditioned mice, a residual anticonvulsive activity (i.e., longer latency to isoniazid convulsion) was noted at 6 h post-conditioning, whereas shorter preconvulsive latencies appeared only at 72 h-post-conditioning period. In conclusion, taken together these data (lack of tolerance development (tolerance studies), prolonged residual anticonvulsive activity, and postponed physical dependence/withdrawal hallmark in diazepam+BPC 157 chronically treated mice) with common benzodiazepines tolerance/withdrawal knowledge, it could be speculated that BPC 157 acts favoring the natural homeostasis of the GABA receptor complex as well as enhancing the GABAergic transmission, and having a mechanism at least partly different from those involved in diazepam tolerance/withdrawal, it may be likely used in further therapy of diazepam tolerance and withdrawal. PMID- 10707893 TI - Calcium rather than calcitonin is dominant to mediate gastric emptying in thyroidectomized rats. AB - Both calcium and calcitonin are important in mediating gastrointestinal motility. Present study tried to study what was the dominant role of calcitonin or calcium replacement on the gastric emptying in thyroidectomized animals. Adult Sprague Dawley male rats received thyroidectomy or sham operation and then housed for two weeks until motility study, which was conducted using radiochromium to measure gastric emptying. Before motility study these rats were i.p. injected with saline or human calcitonin in the doses of 0.1, 1 and 10 microgM/kg, respectively. Another group of thyroidectomized rats received i.v. infusion of saline or CaCl2 for 30 min before motility study. Among thyroidectomized rats, neither saline nor various doses of calcitonin treatment disturbed gastric emptying compared to this of sham operated rats. Thyroidectomy diminished plasma calcium level, however, additional calcitonin treatment did not restore the suppressed calcium level (P<0.01). Of rats following saline or CaCl2 infusion, thyroidectomy did not change gastric emptying, whereas CaCl2 infusion enhanced gastric emptying (P<0.05). In conclusion, exogenous calcium treatment further enhances gastric emptying in thyroidectomized rats, whereas calcitonin replacement has no effect on gastric emptying. We suggest that calcium rather than calcitonin is dominant to mediate gastric emptying. PMID- 10707892 TI - Effects of L-glutamic acid on acid secretion and mucosal blood flow in the rat stomach. AB - The effect of intravenous administration of L-glutamic acid (L-Glu) on gastric acid secretion and gastric mucosal blood flow (GMBF) in anesthetized rats were investigated. Infusion with synthetic L-Glu alone had no effect on spontaneous acid secretion. However, L-Glu reduced histamine- (2 mg/kg/hr) or oxotremorine- (1 microg/kg/hr) stimulated acid secretion, whereas L-Glu had no effect on acid secretion induced by pentagastrin (8 microg/kg/hr). Furthermore, this inhibitory effect of L-Glu on histamine- or oxotremorine-stimulated acid secretion was blocked by 6,7-dinitroquinoxaline-2,3-dione (DNQX), a non-NMDA receptor antagonist. The effect of L-Glu on gastric mucosal microcirculation in the anesthetized rats was evaluated by using Laser Doppler Flowmetry (LDF). The results showed that L-Glu did not significantly reduce both mucosal and serosal blood flow in stomach. No significant modulatory effect on histamine- or oxotremorine-stimulated increase in GMBF was noted after infusion with L-Glu. It is concluded that L-glutamic acid is capable of the modulating of gastric acid secretion via ionotropic non-NMDA receptors, but do not affect on GMBF. However, L-glutamic acid showed no effect on acid secretion by itself. PMID- 10707894 TI - Differential distribution of 5-hydroxytryptamine3 receptor in the colon between human and guinea pig. AB - Localization of 5-hydroxytryptamine3 (5-HT3) receptor in the human colon was examined by in vitro receptor autoradiography using [125I](S)iodozacopride, and compared with that in the guinea pig colon. [125I](S)iodozacopride binding sites were found with high densities around the myenteric plexus, but with low ones in the muscle layer and mucosa of the human colon, and the binding was abolished by granisetron, a specific 5-HT3 receptor antagonist. While in the guinea pig colon, specific [125I](S) iodozacopride binding was not detected in either the myenteric plexus or the muscle layers. Thus, the 5-HT3 receptors are present in the human colon, especially densely located in the myenteric plexus, but not in the guinea pig colon, and those may participate in the colonic motility. The results of functional studies of 5-HT3 receptor obtained from experiments using guinea pig are not always applying to the human. PMID- 10707895 TI - Neurotransmission at the interface of sympathetic and enteric divisions of the autonomic nervous system. AB - The sympathetic and enteric divisions of the autonomic nervous system are interactive in the determination of the functional state of the digestive tract. Activation of the sympathetic input suppresses digestive function primarily through release of norepinephrine at its synaptic interface with the enteric nervous system. The enteric nervous system functions like an independent minibrain in the initiation of the various programmed patterns of digestive tract behavior and moment-to-moment control as the neural microcircuits carry-out the behavioral patterns. Most of the postganglionic projections from sympathetic prevertebral ganglia terminate as synapses in myenteric and submucous ganglia of the enteric nervous system. Two primary actions of the sympathetic input are responsible for suppression of motility and secretion. First is presynaptic inhibitory action of norepinephrine to suppress release of neurotransmitters at fast and slow excitatory synapses in the enteric neural microcircuits and this effectively shuts-down the circuit. Second is inhibitory synaptic input to submucosal secretomotor neurons to the intestinal crypts. The alpha, adrenergic receptor subtype mediates both actions. Axons of secretomotor neurons to the crypts bifurcate to innervate and dilate the submucosal vasculature. Dilitation of the vasculature increases blood flow in support of increased secretion. Sympathetic inhibitory input to the secretomotor neurons therefore suppresses both secretion and blood flow. Activation of the sympathetic nervous system cannot explain the symptoms of secretory diarrhea and abdominal discomfort associated with psychologic and other forms of stress. Current evidence suggests that brain to mast cell connections account for stress-induced gastrointestinal symptoms. Degranulation of enteric mast cells by neural inputs releases inflammatory mediators that enhance excitability of intestinal secretomotor neurons while suppressing the release of norepinephrine from postganglionic sympathetic axons. This is postulated to underlie the secretory diarrhea and abdominal discomfort associated with stress. PMID- 10707896 TI - Comparison between effects of dantrolene and nifedipine on lipopolysaccharide induced endotoxemia in the anesthetized rats. AB - Intracellular calcium is an important mediator for regulating the cellular response in endotoxemia. In this study, we investigated the effects of dantrolene and nifedipine, two agents of reducing intracellular calcium levels, on bacterial endotoxin (lipopolysaccharide, LPS; 10 mg/kg i.v.)-induced production of tumor necrosis factor-alpha (TNF-alpha) and nitric oxide (NO) as well as hemodynamic changes in the anesthetized rat. Injection of LPS (i) induced biphasic changes of blood glucose and rectal temperature: an initial increased phase (<180 min after injection of LPS) followed by a decreased phase (at 240 or 360 min), (ii) caused a significant fall in mean arterial blood pressure from 119+/-3 mmHg (at time 0) to 73+/-67 mmHg (at 360 min) with a concomitant increase of heart rate, (iii) resulted in a substantial hyporeactivity to norepinephrine (NE) (1 microg/kg i.v.), (iv) increased plasma nitrate (an indicator of NO formation) in a time dependent manner, and (v) induced bell-shape changes in plasma TNF-alpha levels which reached a peak at 60 min. Pretreatment of animals with dantrolene (1 mg/kg i.v. at 20 min prior to LPS) or nifedipine (20 microg/kg i.v. infusion for 20 min at 20 min prior to LPS) not only attenuated the delayed circulatory failure (e.g. delayed hypotension and vascular hyporeactivity to NE), but also prevented the overproduction of NO caused by LPS in the rat. However, the prevention of NO overproduction by dantrolene, but not by nifedipine, was associated with an inhibition of TNF-alpha production elicited by LPS. Thus, both dantrolene and nifedipine have beneficial hemodynamic effects, although through different mechanisms, in animals with endotoxic shock. PMID- 10707897 TI - Two subgroups of lung vagal C-fibers with different vulnerabilities to blockades by perivagal capsaicin and vagal cooling in dogs. AB - Perivagal capsaicin treatment and vagal cooling are two techniques that have been widely used to study the respiratory reflexes mediated by lung vagal C-fibers because they can block the neural conduction of unmyelinated fibers. We hypothesized that there are two subgroups of lung vagal C-fibers which have different vulnerabilities to blockades by these two techniques. To test this hypothesis, afferent activity arising from lung vagal C-fibers was recorded in 29 anesthetized, paralyzed, and artificially ventilated dogs. Afferent C-fiber activity was recorded before and after various concentrations of perivagal capsaicin treatment or before and during various temperatures of vagal cooling. Of the 89 lung vagal C-fibers studied, 73 fibers were classified as the group of "low resistance" to capsaicin, while the other 16 were classified as the group of "high resistance". The former group differed from the latter due to their afferent activity being blocked at relatively low concentrations of perivagal capsaicin and at relatively low temperatures of vagal cooling. Our results suggest that lung vagal C-fibers can be categorized into two subgroups, based upon their different blocking thresholds for perivagal capsaicin and vagal cooling. Our data may provide information for researchers to further differentiate the respiratory reflexes originating from these two subgroups of lung vagal C-fibers. PMID- 10707898 TI - Changes in the level of glial fibrillary acidic protein (GFAP) after mild and severe focal cerebral ischemia. AB - In the present study, we examined the temporal and spatial expression profiles of GFAP mRNA and protein in a focal cerebral ischemia model with ischemic injury confined to the cerebral cortex in the right middle cerebral artery (MCA) territory. Northern blot analysis showed a respective 5.5-fold and 7.2-fold increase in the GFAP mRNA in the ischemic right MCA cortex in rats subjected to 30-min (mild) or 60-min (severe) ischemia followed by 72-hr reperfusion. The GFAP mRNA signal remained elevated up to 2-week reperfusion. Interestingly, increased GFAP mRNA signal was clearly demonstrated for the first time in the left MCA cortex. A significant 1.5-fold and 5-fold increase was observed after 72-hr reperfusion following mild and severe ischemia, respectively. However, unlike the ischemic right MCA cortex, this induction was transient in the non-ischemic left MCA counterpart. In situ hybridization studies further revealed characteristic spatial induction profile following mild vs. severe ischemia. In mild ischemia, following 24-hr reperfusion, increase in GFAP mRNA was observed mainly within the ischemic right MCA cortex. Following 72-hr reperfusion, GFAP mRNA signal was observed in virtually the entire ischemic cortex, particularly the amygdala region, then gradually reduced and restricted to right MCA territory and subcortical thalamic nucleus following 2-week reperfusion. On the other hand, in severe ischemia, following 24-hr reperfusion increased GFAP mRNA signal was observed in area surrounding right MCA territory (infarct region) and outer cortical layers within the right MCA territory. Following 72-hr reperfusion, no signal was detected within right MCA cortex; however, increased GFAP signal was detected throughout the remaining ipsilateral cortex and subcortical region, as well as the contralateral cerebral cortex. GFAP mRNA signals then gradually reduced its intensity and was restricted to area surrounding necrosis and ipsilateral thalamic nucleus following 2-week reperfusion. GFAP-like immunoreactivity was also detected in area expressing GFAP mRNA. It is very likely that de novo synthesis was responsible for this increase. In summary, increased GFAP signal was noted in both ipsilateral and contralateral cerebral following mild and severe ischemia. Although the temporal induction profile for mild vs. severe ischemia was similar, the spatial induction profile was different. The mechanism leading to this differential induction and their physiological and functional significance are not clear at present. It is very likely that some local factors may involve, nevertheless, the detailed mechanisms remain to be fully explored. PMID- 10707899 TI - Cl--dependent and Cl--independent Na+/ HCO3- acid extrusion in cultured rat cerebellar astrocytes. AB - It is still uncertain whether the Na+-dependent Cl--HCO3- exchanger (NCBE) is expressed in mammalian astrocytes. Using fluorescent indicators to monitor the intracellular pH (pHi) and intracellular Na+ or Cl- levels, the NCBE in cultured rat cerebellar astrocytes was examined in detail. In nominally bicarbonate-free (Hepes-buffered) medium, a marked pHi recovery from internal acid load was seen which could be blocked completely by 30 microM HOE 694, a specific Na+-H+ exchanger isoform 1(NHE-1) inhibitor, at a pHi above 6.9. These conditions were therefore used to block NHE activity in CO2/HCO3-buffered media when the NCBE was being studied at pHi above 6.9. After internal acid loading in completely Cl- free bicarbonate-buffered medium (containing HOE 694), the rates of pHi recovery and transient Na+ influx were considerably slowed, and the Cl--dependent acid extrusion was both Na+- and 4,4-diisothiocyano-stilbene-disulphonic acid (DIDS) sensitive. Moreover, a HCO3-dependent Cl- efflux during internal acid injection was seen. These results suggest that the NCBE is present in astrocytes. Following repetitive internal acid loading by addition of 5% CO2 to internal Cl- depleted cells, a similar rate of pHi recovery was consistently seen, suggesting Cl- independent pHi regulation also occurred in astrocytes. Moreover, this pHi recovery was completely blocked in the absence of sodium or on addition of DIDS, confirming that the Na+-HCO3 cotransporter (NBC) is present. Thus, the present study provides evidence that both the NCBE and NBC play important roles in acid extrusion in cultured mammalian astrocytes. PMID- 10707900 TI - Promotion of forskolin-induced long-term potentiation of synaptic transmission by caffeine in area CA1 of the rat hippocampus. AB - Caffeine which is present in soft drinks has been shown to increase alertness and allays drowsiness and fatigue. The aim of this study is to investigate whether caffeine could produce a long-term effect on the synaptic transmission using extracellular recording technique in the hippocampal slices. Bath application of caffeine (100 microM) reversibly increased the slope of field excitatory postsynaptic potential (fEPSP). Forskolin (25 microM) by its own did not affect the fEPSP significantly. However, in the presence of caffeine, forskolin induced a long-term potentiation (LTP) of fEPSP. Enprofylline which has been shown to exhibit some actions like caffeine but with a low adenosine antagonistic potency did not affect the normal synaptic transmission or the effect of forskolin at a lower concentration (10 microM). However, when the concentrations were increased to 20 and 50 microM, enprofylline significantly enhanced the fEPSP slope and promoted forskolin-induced LTP. The parallel increase of fEPSP and promotion of LTP observed with enprofylline suggests that adenosine A1 antagonism is the primary mechanism behind caffeine's effect. This hypothesis was further strengthened by the finding that promotion of forskolin-induced LTP was mimicked by the non-xanthine adenosine antagonist 9-chloro-2-(furyl)[1,2,4]triazolo [1,5 c]quinazolin-5-amine (CGS 15943). The promotion of forskolin-induced LTP provides a cellular basis behind caffeine's increase in capacity for sustained intellectual performance. PMID- 10707901 TI - A cystatin-related gene, testatin/cresp, shows male-specific expression in germ and somatic cells from the initial stage of murine gonadal sex-differentiation. AB - Sex-differentiation in mammals initiates at mid-gestation when the differentiation of Sertoli cells is triggered by the expression of the testis determining gene, Sry. However, little is known about the succeeding germ-soma interaction that directs the sex-differentiation of germ cells. We carried out subtraction and differential screening between male and female gonads at 13.5 dpc (days post coitum). A novel cystatin-related gene was identified and named cresp (cystatin-related expressed in Sertoli and spermatogonia), and has recently been reported independently under the name testatin (Tohonen et al., 1998). The presumed amino acid sequence of testatin/cresp showed considerable homology to the cystatin family, but it lacked a few critical amino acid residues for the cysteine-protease inhibitory activity. A 0.7 kb RNA was detected by northern blotting specifically in the fetal and adult testes from 11.5 dpc and expression increased between 11.5 dpc and 12.5 dpc. Using RT-PCR analysis, the testatin/cresp mRNA was first detectable at 9.5 dpc in both male and female embryos but it was maintained only in the male. In females, the expression became weaker at 11.5 dpc and was undetectable after 12.0 dpc. In situ hybridization and immunohistochemical analyses, as well as single cell RT-PCR analysis, showed that the testatin/cresp mRNA was localized specifically in both the (pro)spermatogonia and Sertoli cells in the testis from 12.5 dpc to adult. Thus, expression of the testatin/cresp gene is upregulated in male gonads but downregulated in females immediately after the initiation of sex-differentiation, suggesting roles in the early developmental cascade of testis such as the germ-soma interaction. PMID- 10707902 TI - Pleiotropic effects of zebrafish protein-tyrosine phosphatase-1B on early embryonic development. AB - Protein tyrosine phosphorylation is an important mechanism of eukaryotic cell signalling which is regulated by protein-tyrosine kinases and protein-tyrosine phosphatases. Here we report the molecular cloning of the first zebrafish protein tyrosine phosphatase, zf-PTP-1B, the homologue of human PTP-1B. Zf-PTP-1B was catalytically active and localised to the endoplasmic reticulum, like human PTP 1B. Zf-PTP-1B was maternally expressed in zebrafish embryos, and low ubiquitous expression was detected up to day 7 of development. Microinjection of zf-PTP-1B RNA induced pleiotropic, but reproducible developmental defects. Evaluation of the live embryos at 24 h post fertilisation indicated that zf-PTP-1B induced defects in somite formation. The phenotype was dependent on protein-tyrosine phosphatase activity of zf-PTP-1B, since embryos injected with catalytically inactive zf-PTP-1B-C213S developed normally. Co-injection of wild type and inactive zf-PTP-1B led to a rescue of the zf-PTP-1B-induced phenotype, suggesting that zf-PTP-1B-C213S had dominant negative activity. The zf-PTP-1B-induced phenotype suggests that proper tyrosine phosphorylation of key proteins is essential for early development, most notably somitogenesis. PMID- 10707904 TI - Fate maps of the primitive streak in chick and quail embryo: ingression timing of progenitor cells of each rostro-caudal axial level of somites. AB - Developmental fates of cells emigrating from the primitive streak were traced by a fluorescent dye Dil both in chick and in quail embryos from the fully grown streak stage to 12-somite stage, focusing on the development of mesoderm and especially on the timing of ingression of each level of somitic mesoderm. The fate maps of the chick and quail streak were alike, although the chick streak was longer at all stages examined. The anterior part of the primitive streak predominantly produced somites. The thoracic and the lumbar somites were shown to begin to ingress at the 5 somite-stage and 10 somite-stage in a chick embryo, and 6 somite-stage and 9 somite-stage in a quail embryo, respectively. The posterior part of the streak served mainly as the origin of more lateral or extra embryonic mesoderm. As development proceeded, the fate of the posterior part of the streak changed from the lateral plate mesoderm to the tail bud mesoderm and then to extra embryonic, allantois mesoderm. The fate map of the primitive streak in chick and quail embryo presented here will serve as basic data for studies on mesoderm development with embryo manipulation, especially for transplantation experiments between chick and quail embryos. PMID- 10707903 TI - Convergence of the BMP and EGF signaling pathways on Smad1 in the regulation of chondrogenesis. AB - Bone morphogenetic protein 4 (BMP4) induces, whereas epidermal growth factor (EGF) inhibits chondrogenesis. We hypothesize that BMP4 and EGF mediated intracellular signals are both coupled in the regulation of Meckel's cartilage development. Two chondrogenic experimental model systems were employed to test the hypothesis: (1) an ex vivo, serum-free, organ culture system for mouse embryonic mandibular processes, and (2) a micromass culture system for chicken embryonic mandibular processes. Chondrogenesis was assayed by alcian blue staining and expression of Sox9 and type II collagen. Exogenous EGF inhibited and BMP4 induced ectopic cartilage in a dose-dependent manner. When BMP4- and EGF soaked beads were implanted in juxtaposition within embryonic day 10 mouse mandibular processes, the incidence and amount of ectopic cartilage, and Sox9 and type II collagen expression induced by BMP4, were significantly reduced as the concentration of EGF was increased. Similarly, in chicken serum-free micromass cultures, expression of a constitutively active BMP receptor type IB by replication competent avian retrovirus system promoted the rate and extent of chondrogenesis; however, exogenous EGF attenuated this effect. In micromass cultures, BMP signaling resulted in nuclear translocation and accumulation of the signaling molecule Smad1, whereas the addition of EGF inhibited this event. Our results suggest that BMP4 and EGF function antagonistically, yet are coupled in the regulation of initial chondrogenesis. Smad1 serves as a point of convergence for the integration of two different growth factor signaling pathways during chondrogenesis. PMID- 10707905 TI - In Hydra magnipapillata the activator of protein kinase C diC8 causes multiple head formation along the body axis only when accompanied by feeding, but heavy feeding alone is sufficient to cause multiple head formation. AB - In Hydra magnipapillata additional head structures can be induced to form by daily feeding accompanied by a daily treatment with diC8, an activator of protein kinase C. Based on these results, it was proposed that the PKC- pathway plays a central role in head formation in hydra. The results described here show that ectopic structures, as well as the ectopic localization of nerve cells, can be induced by heavy feeding alone. Furthermore, diC8 treatment does not induce ectopic head structures in starved animals. DiC8 reduces the rate of budding, leading to an unusual lengthening of the body column in reasonably fed animals. PMID- 10707906 TI - A somatic gene transfer approach using recombinant fusion proteins to map muscle motoneuron projections in Xenopus spinal cord. AB - A combination of somatic gene transfer with fusion protein technology has been developed, thus providing an innovative means of mapping muscle-motoneuronal connections in Xenopus tadpole spinal cord. We analyzed whether a neuronal tracer created by the fusion of the LacZ gene to the tetanus toxin C fragment (LacZ-TTC) could be produced from plasmid DNA injected into muscle, and whether it could be released and undergo retrograde transport into motoneurons. Plasmids encoding various fusion protein constructions, with or without a signal peptide, were injected into dorsal or caudal muscles of premetamorphic tadpoles. The marker was produced in the muscle at constantly high levels. At one month post-injection, the fusion protein passed the neuromuscular junction and underwent retrograde transport into motoneurons. Transfer into motoneurons was seen for every animal injected, emphasizing the high reproducibility and efficiency of the process. No uptake of beta-gal protein into motoneurons was observed in the absence of the TTC fragment. Furthermore, no enhancement was obtained by adding a signal peptide. These results provide the first demonstration of the synthesis and transport of a TTC fusion protein produced directly from exogenous DNA in a vertebrate system. PMID- 10707907 TI - The generation and in vivo differentiation of murine embryonal stem cells genetically null for either N-cadherin or N- and P-cadherin. AB - Many mutations of the murine genome are recessive embryonic lethals precluding phenotype analysis at subsequent stages of development. This is true for embryos genetically lacking either N-cadherin or N- and P-cadherin. To circumvent this, we have generated pluripotent embryonal stem (ES) cells of the same genotype in vitro and differentiated them in vivo in the form of teratomas. All of the ES cells isolated in this study had a normal ES cell morphology in vitro and were able to generate teratomas. Histological analysis revealed that some differentiation and histogenesis had occurred within the teratomas. Epithelial formation was, for example, unaffected in all cadherin null cells. Surprisingly, however, the differentiation of cells lacking both N- and P-cadherin was, in general, even more pronounced both quantitatively and qualitatively. Tumours lacking either N- cadherin or N- and P-cadherin contained more striated muscle (apparently cardiac muscle) than heterozygote controls, and this was most strikingly conspicuous in teratomas from N- and P-cadherin null cells. This more pronounced differentiation was not seen for all tissues, however, as structures with a simple neural tube-like morphology were never found in teratomas lacking both N- and P-cadherin and organoid-like structures were rare in Ncad-/-tissue. PMID- 10707908 TI - Restricted expression of the zebrafish hsp90alpha gene in slow and fast muscle fiber lineages. AB - Members of the heat shock protein 90 (Hsp90) family of molecular chaperones play important roles in allowing a select group of intracellular signaling molecules reach and maintain functionally active conformations. We have previously shown that hsp90alpha gene expression in early zebrafish embryos is restricted to a subgroup of paraxial-mesoderm derived somitic cells prior to muscle formation and that the gene is downregulated in mature trunk and tail muscle fibers. Here we have compared the expression of the hsp90alpha gene to muscle regulatory genes during development of slow and fast muscle fibers in normal embryos and in embryos carrying mutations which affect somitic muscle formation. We show that hsp90alpha is first expressed early during the development of slow somitic muscle progenitors shortly following myoD activation and at a point prior to or co incident with the expression of other known muscle regulatory genes. Expression of hsp90alpha is also activated in the midline of flh mutants when these cells switch from a notochord to a muscle fate. Conversely, expression is not detectable in cells of the paraxial mesoderm lineage which fail to converge in spt mutants and which do not activate expression of other muscle specific marker genes. Finally, expression of hsp90alpha is downregulated in slow muscle fibers by 24 h of age but becomes detectable in the later developing fast fibers at this time. Thus, hsp90alpha is expressed in developing muscle progenitors during short temporal and spatial windows of both slow and fast fiber lineages in the zebrafish somite. PMID- 10707909 TI - Age and gonadotropins control Ca2+-spike acquisition in mouse oocytes isolated from early preantral follicles. AB - The action of gonadotropins upon the oocyte is known to be crucial at later stages of follicular development in mammals. However, its influence on oocytes at early preantral stages is still a matter of debate. In the present study we evaluated the onset of mouse oocyte's capacity to exhibit calcium spikes during preantral stages of follicular development, prior to meiotic competence acquisition. In particular, through the use of confocal microscopy, we probed for the specific effects of age and gonadotropin stimulation upon the calcium dynamics of preantral follicle oocytes. We found that important developmental changes on the Ca2+ signalling mechanisms take place early during follicular development. Specifically we demonstrate that both age and gonadotropin stimulation increase the capacity of oocytes recovered from preantral follicles to exhibit calcium spikes. We propose that a strictly morphological staging of follicular development is insufficient to predict oocyte behaviour and must take in consideration animal age and gonadotropin environment. PMID- 10707910 TI - Changes in the placenta and in the rat embryo caused by the demethylating agent 5 azacytidine. AB - DNA methylation is an important mechanism for regulation of gene expression during vertebrate development. 5-azacytidine is used as an experimental tool for demethylation. In this work, a single dose of 5-azacytidine (5 mg/kg body weight) was administered to rats at different stages of development. After 5-azacytidine administration on the first or third day of pregnancy, no changes were detected. After administration on the fourth day of pregnancy or later, a reduction in growth was observed. After treatment on day five and on any other day till day eleven of pregnancy, no living fetuses were found. Of those treated on day twelve, 24% of fetuses survived, but forelimb and hindlimb malformations were present. Administered on day thirteen, 5-azacytidine did not interfere with survival, but malformations were still present. From day fourteen on, 5 azacytidine caused no gross external malformations. Placentas were also influenced by 5-azacytidine. They were significantly smaller and histological evaluation showed the labyrinthine part to be severely reduced. In contrast, trophoblast giant cells were more abundant than in controls. PMID- 10707911 TI - Harvesting and long term exposure effects in the relation between air pollution and mortality. AB - While time series analyses have demonstrated that airborne particles are associated with early death, they have not clarified how much the deaths are advanced. If all of the pollution-related deaths were advanced by only a few days, one would expect little association between weekly averages of air pollution and daily deaths. The author used the STL algorithm to classify data on air pollution, daily deaths, and weather from Boston, Massachusetts (1979-1986) into three time series: one reflecting seasonal and longer fluctuations, one reflecting short term fluctuations, and one reflecting intermediate patterns. By varying the cutoff point between short term and intermediate term, it was possible to examine harvesting on different time scales. For chronic obstructive pulmonary disease, there was evidence that most of the mortality was displaced by only a few months. For pneumonia, heart attacks, and all-cause mortality, the effect size increased with longer time scales. The percentage increase in all deaths associated with a 10-microg/m3 increase in PM2.5 rose from 2.1% (95% confidence interval: 1.5, 4.3) to 3.75% (95% confidence interval: 3.2, 4.3) as the focus moved from daily patterns to monthly patterns. This is consistent with the larger effect seen in prospective cohort studies, rather than harvesting's playing a major role. PMID- 10707912 TI - Beyond the body count: air pollution and death. PMID- 10707913 TI - Contribution of genetic and environmental influences to ankle-brachial blood pressure index in the NHLBI Twin Study. National Heart, Lung, and Blood Institute. AB - The ankle-brachial index (ABI) is widely used in the clinical diagnosis of peripheral arterial disease. The contributions of genetic and environmental influences to normal and abnormal ABI values are unknown. In this study, the authors used available data on 94 monozygotic pairs and 90 dizygotic pairs of elderly, White, male twins examined in 1995-1997 to investigate the contributions of genetic and environmental influences to normative ABI values. Within-twin-pair correlations for normative ABI values were statistically significant, and the correlation in monozygotic twin pairs was significantly greater than that in dizygotic pairs. Structural equation modeling of the variance-covariance matrices of monozygotic and dizygotic twins indicated that 48% of the observed variability in ABI values could be attributed to additive genetic effects. In contrast, concordance rates for low ABI values (ABI< or =0.9) for both monozygotic and dizygotic twins were significantly greater than would be expected by chance alone, but within-pair monozygotic similarity was not significantly greater than dizygotic similarity. A matched-cotwin analysis in 21 pairs that were discordant for low ABI values found that twins with low ABI values were physically less active and more likely to be persistent smokers than their normal-control brothers. These findings reinforce the role of individual health practices (e.g., physical activity, smoking) in the manifestation of peripheral arterial disease among subjects matched for age, genetics, and early shared environment. PMID- 10707914 TI - Does heavy physical exertion trigger myocardial infarction? A case-crossover analysis nested in a population-based case-referent study. AB - To study possible triggering of first events of acute myocardial infarction by heavy physical exertion, the authors conducted a case-crossover analysis (1993 1994) within a population-based case-referent study in Stockholm County, Sweden (the Stockholm Heart Epidemiology Program). Interviews were carried out with 699 myocardial infarction patients after onset of the disease. These cases represented 47 percent of all cases in the study base, and 70 percent of all nonfatal cases. The relative risk from vigorous exertion was 6.1 (95% confidence interval: 4.2, 9.0). The rate difference was 1.5 per million person-hours, and the attributable proportion was 5.7 percent. The risk was modified by physical fitness, with an increased risk being seen among sedentary subjects as in earlier studies, but the data also suggested a U-shaped association. In addition, the trigger effect was modified by socioeconomic status. Premonitory symptoms were common, and this implies risks of reverse causation bias and misclassification of case exposure information that require methodological consideration. Different techniques (the use of the usual-frequency type of control information, a pair matched analysis, and a standard case-referent analysis) were applied to overcome the threat of misclassification of control exposure information. A case-crossover analysis in a random sample of healthy subjects resulted in a relative risk close to unity, as expected. PMID- 10707915 TI - Serum albumin level as a predictor of incident coronary heart disease: the Atherosclerosis Risk in Communities (ARIC) study. AB - Various studies have reported an inverse association between serum albumin level and incident coronary heart disease (CHD), though biologic mechanisms have not been established. The authors examined the association between serum albumin level and CHD in the Atherosclerosis Risk in Communities cohort, comprising 14,506 White and African-American middle-aged men and women. The mean albumin level in this population was 3.9 g/dl (standard deviation 0.3). During 5.2 years of follow-up, 470 incident CHD events occurred. The hazard ratio for incident CHD associated with a 1-standard deviation decrease in serum albumin level was 1.26 (95% confidence interval (CI): 1.15, 1.38) after adjustment for age, gender, and ethnicity and 1.18 (95% CI: 1.07, 1.30) after additional adjustment for covariates related to CHD. Hazard ratios were similar across gender and ethnic groups. However, there was statistically significant effect modification by smoking status, with hazard ratios of 1.01 (95% CI: 0.84, 1.22) among never smokers, 1.09 (95% CI: 0.92, 1.30) among former smokers, and 1.35 (95% CI: 1.17, 1.54) among current smokers. Further adjustment for factors related to renal disease, nutrition, platelet aggregation, inflammation, use of angiotensin converting enzyme inhibitors, and hemostasis factors attenuated the albumin-CHD relation only slightly. In this study, serum albumin was inversely associated with incident CHD at the baseline examination in current smokers but not in never or former smokers. Albumin level may be a marker of susceptibility to the inflammatory response that results from smoking. PMID- 10707916 TI - Carotid wall thickness is predictive of incident clinical stroke: the Atherosclerosis Risk in Communities (ARIC) study. AB - Few studies have determined whether carotid artery intima-media thickness (IMT) is associated prospectively with risk of first ischemic stroke. In the Atherosclerosis Risk in Communities Study, carotid IMT, an index of generalized atherosclerosis, was defined as the mean of IMT measured by B-mode ultrasonography at six sites of the carotid arteries. The authors assessed the relation of mean IMT to stroke incidence over 6-9 years' follow-up (1987-1995) among 7,865 women and 6,349 men aged 45-64 years without prior stroke at baseline in four US communities. There were 90 incident ischemic stroke events for women and 109 for men. In sex-specific Cox proportional hazards models adjusting only for age, race, and community, the hazard rate ratios comparing extreme mean IMT values (> or =1 mm) to values less than 0.6 mm were 8.5 for women (95% confidence interval: 3.5, 20.7) and 3.6 for men (95% confidence interval: 1.5, 9.2). The relation was graded, and models with cubic splines indicated significant nonlinearity, with hazards increasing more rapidly at lower IMTs than at higher IMTs. Thus, models using linear IMT values substantially underestimate the strength of the association at lower IMTs. The strength of the association was reduced by the inclusion of putative stroke risk factors, but it remained elevated at higher IMTs. Hence, mean carotid IMT is a noninvasive predictor of future ischemic stroke incidence. PMID- 10707917 TI - Nonsteroidal antiinflammatory drugs and acute renal failure in elderly persons. AB - Renal prostaglandin inhibition by nonsteroidal antiinflammatory drugs (NSAIDs) may decrease renal function, especially under conditions of low effective circulating volume. To evaluate the risk of important deterioration of renal function due to this effect, the authors performed a nested case-control study using Tennessee Medicaid enrollees aged > or =65 years in 1987-1991. Cases were patients who had been hospitalized with community-acquired acute renal failure; they were selected on the basis of medical record review of Medicaid enrollees with selected discharge diagnoses. Information on the timing, duration, and dose of prescription NSAIDs used, demographic factors, and comorbidity was gathered from computerized Medicaid-Medicare data files. Of the 1,799 patients with acute renal failure (4.51 hospitalizations per 1,000 person-years), 18.1% were current users of prescription NSAIDs as compared with 11.3% of 9,899 randomly selected population controls. After control for demographic factors and comorbidity, use of NSAIDs increased the risk of acute renal failure 58% (adjusted odds ratio = 1.58; 95% confidence interval (CI): 1.34, 1.86). For ibuprofen, which accounted for 35% of NSAID use, odds ratios associated with dosages of < or =1,200 mg/day, >1,200-<2,400 mg/day, and > or =2,400 mg/day were 0.94 (95% CI: 0.58, 1.51), 1.89 (95% CI: 1.34, 2.67), and 2.32 (95% CI: 1.45, 3.71), respectively (test for linear trend: p = 0.009). Prescription NSAID use resulted in an estimated 25 excess hospitalizations associated with renal failure per 10,000 years of use. Thus, NSAIDs represent a relatively uncommon but avoidable cause of acute renal failure in frail elderly persons. PMID- 10707918 TI - Risk factors for cortical, nuclear, and posterior subcapsular cataracts: the POLA study. Pathologies Oculaires Liees a l'Age. AB - The POLA (Pathologies Oculaires Liees a L'Age) Study is a population-based study of cataract and age-related macular degeneration and their risk factors being carried out among 2,584 residents of Sete, southern France, aged 60-95 years. Recruitment took place between June 1995 and July 1997. Cataract classification was based on a standardized lens examination by slit lamp, according to Lens Opacities Classification System III. This paper presents results obtained from cross-sectional analysis of the first phase of the study. In polytomous logistic regression analyses, an increased risk of cataract was found for female sex (cataract surgery: odds ratio (OR) = 3.03; cortical cataract: OR = 1.67), brown irises (cortical, nuclear, and mixed cataracts: OR = 1.61), smoking (cataract surgery: OR = 2.34 for current smokers and OR = 3.75 for former smokers), known diabetes of 10 or more years' duration (posterior subcapsular, cortical, and mixed cataracts and cataract surgery: OR = 2.72), use of oral corticosteroids for at least 5 years (posterior subcapsular cataract: OR = 3.25), asthma or chronic bronchitis (cataract surgery: OR = 2.04), cancer (posterior subcapsular cataract: OR = 1.92), and cardiovascular disease (cortical cataract: OR = 1.96). Decreased risk of cataract was found with higher education (all types of cataract and cataract surgery: OR = 0.59), hypertension (cataract surgery: OR = 0.57), and high plasma retinol levels (nuclear and mixed cataracts and cataract surgery: OR = 0.75 for a 1-standard-deviation increase). Most of the risk factors identified in this study confirm the findings of other studies. The association of cataract with plasma retinol level requires further investigation. PMID- 10707919 TI - Prior spontaneous abortion, prior elective termination, interpregnancy interval, and risk of neural tube defects. AB - A woman with a history of spontaneous abortion in her immediately prior pregnancy may be at increased risk for a pregnancy affected by a neural tube defect (NTD). A short interpregnancy interval may further increase this risk. Using data from a population-based case-control study (1989-1991), the authors investigated NTD risk resulting from a prior spontaneous abortion or elective termination and a short interpregnancy interval. Of 538 interviewed case mothers and 539 interviewed control mothers, 408 case mothers and 433 control mothers reported having a prior pregnancy. Analysis showed a slightly decreased NTD risk among mothers whose immediately prior pregnancy had ended in a spontaneous abortion or elective termination in comparison with a live birth (odds ratio (OR) = 0.82; 95% confidence interval (CI): 0.61, 1.1). This decreased risk was consistent across strata of short or long interpregnancy intervals. Additional analysis revealed an increased NTD risk for mothers with an interpregnancy interval of < or =6 months compared with >12-< or =24 months (OR = 1.5; 95% CI: 0.93, 2.4). This latter risk was greatest among mothers whose immediately prior pregnancy had resulted in a live birth (OR = 2.0; 95% CI: 1.0, 3.8) rather than a spontaneous abortion or elective termination (OR = 0.96; 95% CI: 0.44, 2.1). Adjustment for potential covariates did not substantially alter observed risk estimates. PMID- 10707920 TI - Are children living near high-voltage power lines at increased risk of acute lymphoblastic leukemia? AB - In the National Cancer Institute/Children's Cancer Group case-control study of childhood acute lymphoblastic leukemia (1989-1993), living in a home with a high voltage wire code was not associated with disease risk. To further investigate risk near power lines, the authors analyzed distance to transmission and three phase primary distribution lines within 40 m of homes and created an exposure index of distance and strength of multiple power lines (408 case-control pairs). Neither distance nor exposure index was related to risk of childhood acute lymphoblastic leukemia, although both were associated with in-home magnetic field measurements. Residence near high-voltage lines did not increase risk. PMID- 10707921 TI - Relation between psychiatric syndromes and behaviorally defined sexual orientation in a sample of the US population. AB - Most surveys of the prevalence of psychiatric disorders among lesbians and gay men find no increased risk in comparison with heterosexuals. However, the majority of this work has relied on convenience samples drawn from the visible lesbian and gay community. The authors examined differences in 1-year prevalence of six psychiatric syndromes among sexually active individuals in the 1996 National Household Survey of Drug Abuse who reported either exclusive heterosexuality (n = 9,714) or having any same-gender sex partners (n = 194) in the prior year. Although nearly three quarters of homosexually active individuals did not meet criteria for any of the six syndromes assessed, in multivariate logistic regression analyses, homosexually active men were more likely than other men to evidence major depression and panic attack syndromes. In contrast, homosexually active women were more likely than other women to be classified with alcohol or drug dependency syndromes. Both men and women reporting any same gender sex partners were more likely than others to have used mental health services in the year prior to interview. These findings suggest a small increased risk among homosexually active populations in 1-year psychiatric morbidity and use of mental health care services. PMID- 10707922 TI - Outbreak of aseptic meningitis associated with mass vaccination with a urabe containing measles-mumps-rubella vaccine: implications for immunization programs. AB - A mass immunization campaign with a Urabe-containing measles-mumps-rubella vaccine was carried out in 1997 in the city of Salvador, northeastern Brazil, with a target population of children aged 1-11 years. There was an outbreak of aseptic meningitis following the mass campaign. Cases of aseptic meningitis were ascertained through data collected from the records of children admitted to the local referral hospital for infectious diseases between March and October of 1997, using previously defined eligibility criteria. Vaccination histories were obtained through home visits or telephone calls. Eighty-seven cases fulfilled the study criteria. Of those, 58 cases were diagnosed after the vaccination campaign. An elevated risk of aseptic meningitis was observed 3 weeks after Brazil's national vaccination day compared with the risk in the prevaccination period (relative risk = 14.3; 95% confidence interval: 7.9, 25.7). This result was confirmed by a case series analysis (relative risk = 30.4; 95% confidence interval: 11.5, 80.8). The estimated risk of aseptic meningitis was 1 in 14,000 doses. This study confirms a link between measles-mumps-rubella vaccination and aseptic meningitis. The authors discuss the implications of this for the organization and planning of mass immunization campaigns. PMID- 10707923 TI - Problems due to small samples and sparse data in conditional logistic regression analysis. AB - Conditional logistic regression was developed to avoid "sparse-data" biases that can arise in ordinary logistic regression analysis. Nonetheless, it is a large sample method that can exhibit considerable bias when certain types of matched sets are infrequent or when the model contains too many parameters. Sparse-data bias can cause misleading inferences about confounding, effect modification, dose response, and induction periods, and can interact with other biases. In this paper, the authors describe these problems in the context of matched case-control analysis and provide examples from a study of electrical wiring and childhood leukemia and a study of diet and glioma. The same problems can arise in any likelihood-based analysis, including ordinary logistic regression. The problems can be detected by careful inspection of data and by examining the sensitivity of estimates to category boundaries, variables in the model, and transformations of those variables. One can also apply various bias corrections or turn to methods less sensitive to sparse data than conditional likelihood, such as Bayesian and empirical-Bayes (hierarchical regression) methods. PMID- 10707924 TI - Values and limitations of 18F-fluorodeoxyglucose-positron-emission tomography with preoperative evaluation of patients with pancreatic masses. AB - The aim of this study was to determine the value and limitations of 18F fluorodeoxyglucose (FDG)-position-emission tomography (PET) for differentiating benign and malignant pancreatic disease and for staging malignant disease. One hundred fifty-nine patients with 89 malignant and 70 benign pancreatic lesions all received PET, computed tomography (CT), and endoscopic retrograde cholangiopancreatography (ERCP) before pancreatic surgery. The original reports were compared for all patients (group I; N = 159), for a subgroup that neither had fasting plasma glucose levels > or =130 mg/dL or known elevated levels of C reactive protein ([CRP], group II; n = 123), and for the remaining patients (group III; n = 36). For group I, accuracy values (areas under receiver operating characteristic [ROC] curves) for differentiation of benign/malignant masses were 0.86 (PET), 0.93 (ERCP), 0.82 (CT), and 0.95 for ERCP + PET (N = 159). For group II, ROC areas increased to 0.92 (PET), 0.94 (p < 0.05; n = 123) (ERCP), 0.82 (CT), 0.97 (p < 0.05; n = 123) (ERCP + PET). The results for group III were 0.71 (PET), 0.81 (CT), and 0.93 (ERCP); (n = 36). With 54 patients of group II that either had contradictory or indeterminate/technically unsuccessful CT/ERCP, PET was correct in 43 patients (84%). Sensitivity/specificity for lymph node staging was 49%/63%, respectively. For patients with hepatic metastasis, PET was 70% sensitive and 95% specific, missing some metastasis that were <1 cm. PET detected peritoneal metastasis in 25% of patients, missing poorly localized microscopic spread. For selected patients who have indeterminate pancreatic masses but no hyperglycemia or serologic evidence of active inflammation, FDG-PET is an independent functional assay that significantly adds to the diagnostic accuracy of ERCP and CT in the differentiation of benign and malignant pancreatic disease. PET can reliably detect hepatic, peritoneal, and other distant metastases that are > or =1 cm. PMID- 10707925 TI - Spontaneous apoptosis and proliferation in human pancreatic cancer. AB - Several studies have documented the role of programmed cell death in the development and/or progression of cancer. The aims of this study were to analyze (a) the spontaneous apoptosis in human pancreatic duct carcinoma by terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-digoxigenin nick end labeling (TUNEL); (b) its correlation with the proliferation rate of the tumor (determined by immunohistochemistry by using monoclonal antibody MIB-1); and (c) the association of apoptotic and mitotic index with the histologic features of the tumor and the outcome of patients. In pancreatic cancer, the apoptotic index (AI) was 4.9 +/- 4.8, and the mitotic index (MI) was 1.3 +/- 1.0 (mean +/- SD). AI was higher in small (<4 cm) than in large (>4 cm) size primary tumors (p = 0.02) and in undifferentiated as compared with differentiated cancers (p = 0.05). Significantly higher values of MI were detected in advanced as compared with early-stage carcinomas (p = 0.03) and when perineural invasion was present (p = 0.03). No correlation was found between AI and MI. Patients with AI > 2.3 survived significantly less than those with lower AI values (p = 0.03). Mitotic index >0.5 was associated with a worse survival (p = 0.006). These results suggest that in pancreatic cancer, spontaneous apoptosis is present and is more evident in small and undifferentiated tumors. Proliferation is increased in the advanced stage of cancer and seems to be independent of apoptosis. Higher levels of apoptosis and proliferation are negative prognostic indexes. PMID- 10707926 TI - Coexpression of FAS and FAS-ligand in chronic pancreatitis: correlation with apoptosis. AB - Activation of the Fas receptor by Fas-ligand (FasL) results in apoptosis, and dysregulation of this pathway may contribute to abnormal cell proliferation and cell death. The aim of this study was to compare the expression of Fas and FasL in the normal pancreas and chronic pancreatitis (CP). By Northern blotting, Fas messenger RNA (mRNA) levels were increased in CP in comparison to the normal pancreas. Immunostaining revealed that faint Fas and FasL immunoreactivity was present in ductal and islet cells of the normal pancreas. In CP, there was faint Fas and strong FasL immunoreactivity in the proliferating ductal cells. Additionally, many of these ductal cells in the CP samples exhibited an apoptotic signal, as determined by DNA 3'-OH end labeling. These findings suggest that activation of apoptosis through the Fas receptor may contribute to the pathobiology of CP. PMID- 10707927 TI - Beta1 integrins play an essential role in adhesion and invasion of pancreatic carcinoma cells. AB - To investigate the role of beta1 integrins in pancreatic carcinoma invasion, we analyzed the relationship between the activity of beta1 integrins and the invasive ability of human pancreatic carcinoma cell lines. AsPC1, BxPC3, PANC1, SU8686, KP1NL, KP2, and H48N cells had high expression of beta1 and alpha6 subunits, and various levels of alpha2, alpha3, and alpha5 expression as determined by flow cytometry. Cell adhesion assay revealed that alpha2beta1, alpha5beta1, and alpha6beta1 integrins were the predominant adhesion receptors for collagen, fibronectin, and laminin, respectively. Beta1 integrins on different cell types showed a wide range of constitutive activity. Anti-beta1 monoclonal antibody (MAB) TS2/16 rapidly activated beta1 integrins, and thus TS2/16 requirement in cell adhesion represented the levels of constitutive activity of beta1 integrins. Notably, as the result of in vitro chemoinvasion assay, the levels of constitutive activity of beta1 integrins correlated with the invasive ability of pancreatic carcinoma cells. The inhibitory anti-beta1 MAB 13 completely blocked the invasion of these cell lines. Alternatively, the stimulatory anti-beta1 MAB TS2/16 strongly inhibited the invasion. These results show an essential role of beta1 integrins in invasion of pancreatic carcinoma cells and also suggest subtle regulatory mechanisms of cell invasion. PMID- 10707928 TI - Transforming growth factor-beta1 acts as a potent inhibitor of complement C3 biosynthesis in human pancreatic cancer cell lines. AB - In this study, we attempted to determine how transforming growth factor (TGF) beta1 affects complement C3 secretion in the pancreatic cancer cell lines PANC-1 and BxPC-3. We also compared the responses in C3 secretion with those in interleukin (IL)-8 secretion. The C3 and IL-8 expression was evaluated at the protein and messenger RNA (mRNA) levels. The activation of nuclear factor-kappaB (NF-kappaB) was assessed by an electrophoretic gel mobility shift assay (EMSA). IL-1beta and tumor necrosis factor (TNF)-alpha both induced a marked increase in C3 and IL-8 secretion. However, TGF-beta1 potently decreased the IL-1beta- and TNF-alpha-induced C3 secretion, whereas the IL-8 secretion was weakly but significantly enhanced. These responses were also observed at the mRNA level. In PANC-1 cells, IL-1beta and TNF-alpha induced a rapid activation of nuclear factor (NF)-kappaB, and TGF-beta1 enhanced this activation slightly. The induction of Fos protein has been reported to be required for the inhibitory action of TGF beta1, and the translocation of Fos protein into the nucleus was associated with TGF-beta1 stimulation in PANC-1 cells. Our results suggest that TGF-beta1 may act as a potent inhibitor of C3 secretion in pancreatic cancer cell lines under inflammatory conditions. This action of TGF-beta1 did not correlate with NF kappaB activation, but associated with the translocation of Fos protein into the nucleus. PMID- 10707929 TI - Experimental pancreatitis induced by synthetic prooxidant tert-butyl hydroperoxide. AB - The purpose of this study was to verify whether injection of tert-butyl hydroperoxide (Bu(t)OOH, a well-known prooxidant agent) into the bile-pancreatic duct can induce acute pancreatitis. A rapid blockade of the secretion was observed in the majority of the animals after 3 hours of observation. After 6 hours, the secretion reached a very low level, significantly different compared with controls. In groups of rats injected with Bu(t)OOH, pancreatic weight gain was observed compared with the rats injected with physiologic saline. Histology of pancreata removed 3 hours after injection of Bu(t)OOH showed acinar cell vacuolization, interstitial edema, focal necrosis of pancreatic acini, fat-tissue necrosis, and leukocyte infiltration of the organ. These changes were considerably greater after the 6-hour observation period. Electron-microscopic inspection revealed profound morphologic changes 3 hours after Bu(t)OOH injection. The control rats receiving physiologic saline alone had well-preserved pancreatic tissue structure. In conclusion, injection of the prooxidant agent, tert-butyl hydroperoxide, into common bile-pancreatic duct induces acute necrotizing pancreatitis, which indicates the crucial role of free radical reactions in pathogenesis of this disease. PMID- 10707930 TI - Homologous orthotopic implantation models of pancreatic ductal cancer in Syrian golden hamsters: which is better for metastasis research--cell implantation or tissue implantation? AB - With a nitrosamine induced hamster pancreatic cancer cell line (HaP-T1), survival time and metastatic rates were compared between orthotopic cell implantation (OCI; n = 5) and orthotopic tissue implantation (OTI; n = 5) models. All the tumors were palpable (100% tumor take) after 1 week in both groups. Hamsters in the OCI group survived 71 +/- 2.17 days (range, 69-75 days), and in the OTI group, 73.8 +/- 4.03 days (range, 58-80 days). After necropsy, spontaneous metastases were noted in 100% of the animals. Direct invasion to adjacent organs was observed in four animals, and liver metastases, in three in the OTI group, which were significantly higher compared with the OCI group. On the other hand, peritoneal dissemination was observed only in the OCI group. Other metastatic sites showed no significant difference between the groups. All the histologically noted metastases had K-ras point mutation confirmed by polymerase chain reaction restriction fragment length polymorphism (PCR/RFLP) analysis. We conclude that the homologous OTI model may be more useful than the OCI model. The OTI model may contribute to the development of therapeutic strategies in the field of pancreatic cancer research because of the capacity for invasion to adjacent organs, higher liver metastatic rate, and similarity to the clinical picture of the disease. PMID- 10707931 TI - Familial pancreatic hyperenzymemia. AB - I recently described a new form of pancreatic hyperenzymemia in healthy subjects in the absence of any pancreatic disease. The aim of this study was to determine whether this pancreatic hyperenzymemia has a familial distribution. From January 1996 to January 1999, 25 subjects with chronic nonpathological pancreatic hyperenzymemia were seen, in 23 of whom it was possible to investigate family members. A total of 102 subjects was studied. In addition to clinical history, physical examination and routine blood analysis, serum amylase, pancreatic isoamylase, lipase, and amylasuria were determined. Abdominal ultrasonography was also performed. Seven of the 23 families studied were found to have more than one member with pancreatic hyperenzymemia. In all, 19 persons (eight female and 11 male) were found to have this trait (mean age, 32.7 years; range, 3-84). The increase in enzyme concentration over the upper normal limit was in the range of 1.3- to 5.2-fold for amylase, 1.4- to 8.6-fold for pancreatic isoamylase, and 1.6 to 18.0-fold for lipase. Amylasuria was also increased. Clinically, no subject had symptoms or signs of pancreatic or other disease. Abdominal ultrasonography was normal in all of them. The results of this study show that pancreatic hyperenzymemia in healthy subjects may present with a familial distribution. The underlying defect in this disorder is not known. Although it is a benign condition, awareness of it is important to avoid unnecessary concern, examinations, and expenditure. PMID- 10707932 TI - N-acetylcysteine decreases severity of acute pancreatitis in mice. AB - Oxidative stress plays a major role in the early stage of acute pancreatitis. This study assessed the effects of N-acetylcysteine (NAC), a reduced glutathione (GSH) provider and a direct scavenger of reactive oxygen intermediates, in the course of acute pancreatitis in mice. Acute pancreatitis (AP) was induced by intraperitoneal (i.p.) injections of cerulein. Mice received NAC (1,000 mg/kg, i.p.) every 3 h, starting either 1 h before the first cerulein injection (prophylactic group) or 1 h after the first cerulein injection (therapeutic group), or i.p. saline injections for controls. Severity of AP was evaluated by histology, serum hydrolase levels, and serum and intrapancreatic levels of MCP-1 and interleukin 6 (IL-6). Pancreatic conjugated dienes and intrapancreatic and intrahepatic GSH levels were measured to assess the local and systemic oxidative processes. Acute pancreatitis was also induced with a CDE diet in controls and mice receiving either both NAC ad libidum in drinking water and 1,000 mg/kg i.p. injection once daily. The severity of pulmonary lesions was assessed by arterial blood gases (pO2) and intrapulmonary myeloperoxidase (MPO content) measurements as well as the survival of mice. The severity of cerulein-induced AP was significantly decreased in the prophylactic group compared with the therapeutic and control groups. Prophylactic administration of NAC also decreased the intrapancreatic levels of conjugated dienes compared with controls. The intrapancreatic and systemic release of MCP- 1 and IL-6 was also decreased in the prophylactic group 3 and 6 hours after AP induction. In addition, NAC pretreatment also reduced hepatic IL-6 production at 3 and 6 hours after starting cerulein challenge. In CDE-induced AP, the severity of lung injury (hypoxemia, MPO content) was decreased, and survival was improved by NAC. NAC administered in a prophylactic protocol limits the severity of experimental acute pancreatitis in mice, as well as its systemic complications and related mortality. PMID- 10707933 TI - Effects of intraluminal trypsin and bile on the exocrine and endocrine pancreas after pancreaticobiliary diversion and biliodigestive shunt. AB - Pancreaticobiliary diversion (PBD) and biliodigestive shunt (BDS) cause long standing hypercholecystokininemia followed by pancreatic hyperplasia. These changes have been suggested to be due to the lack of intraluminal trypsin and bile, respectively, in the upper small intestine. The aim of these experiments was to study the effect of restoration of intraluminal trypsin and bile on plasma levels of cholecystokinin (CCK) and the changes found in exocrine and endocrine pancreas after PBD and BDS. Male Sprague-Dawley rats were used. PBD was done in 16 rats, eight of which had trypsin dissolved in 50 mM sodium bicarbonate (SB), and eight had SB only by gastric intubation twice daily. BDS was done in another 16 rats, eight of which had bile dissolved in SB, and eight had SB in a similar manner. Sham-operated rats had SB and served as controls. After 4 weeks, the rats were killed, and the concentrations of circulating CCK, gastrin, glucose, glucagon, and insulin were determined. The pancreas was removed, weighed, and analyzed for contents of water, protein, and DNA. In another study, PBD-operated rats got trypsin in varying dosages or trypsin and taurocholate in combination for 2 weeks before death. The concentrations of plasma CCK and glucagon were elevated after both PBD and BDS. PBD decreased the concentration of gastrin in plasma. PBD caused an increase of pancreatic weight and the contents of protein and DNA. Trypsin substitution to PBD-operated rats did not affect plasma CCK or glucagon levels, but the PBD-induced increases in weight and DNA content were counteracted by trypsin. Higher dosages of trypsin did not further influence the effects seen after PBD. Pancreatic weight and DNA content were increased after BDS. Bile administration completely abolished the increase in plasma CCK and glucagon, as well as the gain in pancreatic weight, and reduced the increase in pancreatic DNA. Substitution with bile to BDS-operated rats abolished the increase in the plasma levels of CCK and glucagon, as well as the trophic effects on the pancreas. Trypsin substitution to PBD-operated rats partly reversed the trophic effects on the pancreas but not the hormonal changes in plasma. Thus the trophic effects on the pancreas exerted by BDS seem to be dependent on the lack of bile in the upper small intestine, whereas the effects of PBD only partly are a consequence of the absence of intraluminal trypsin. PMID- 10707934 TI - Role of adrenergic receptors in veratridine-stimulated amylase secretion from rabbit pancreatic lobules. AB - Sympathetic inhibition of pancreatic enzyme secretion has been attributed to vasoconstriction and direct inhibition of acinar cells. We observed both adrenergic inhibition and facilitation of cholinergic transmission in rabbit pancreatic ganglia, which innervate acini. Here we used pancreatic lobules to determine whether adrenergic receptors also regulate synaptic transmission between pancreatic nerves and acini. Stimulation of pancreatic nerve terminals with veratridine (Ver), an activator of voltage-dependent Na+ channels, resulted in a 102% increase in amylase secretion, which was unaffected by alpha and beta receptor antagonists but inhibited 65% by atropine. At a concentration of 10 microM, norepinephrine inhibited (38%) and epinephrine potentiated (40%) Ver stimulated secretion. At the same concentration, the alpha2 agonist clonidine (Clon) inhibited (39%), whereas the nonselective beta agonist isoproterenol (Iso) and the selective beta3 agonist BRL 37344 potentiated (71 and 67%, respectively) nerve-stimulated secretion. The effects of Clon and Iso and BRL 37344 were antagonized by yohimbine and propranolol, respectively. Phenylephrine, dobutamine, and terbutaline had no effect. Neither basal, bethanechol-stimulated, nor noncholinergic nerve-stimulated secretion was significantly altered by Clon or Iso. Thus, cholinergic nerve terminals innervating pancreatic acini exhibit both inhibitory alpha2 and atypical facilitatory beta adrenergic receptors. The apparent lack of adrenergic innervation suggests that adrenergic receptors on the nerve terminals of cholinergic pancreatic neurons are under hormonal control by circulating catecholamines. These results provide further evidence that intrinsic pancreatic neurons, which supply most, if not all, of the cholinergic innervation of acini, are important sites of sympathetic regulation of pancreatic exocrine secretion. PMID- 10707935 TI - Improved quality and yield of islets isolated from human pancreata using a two step digestion method. AB - A new approach, involving a two-step digestion process and Los Angeles preservation solution #1 (LAP-1), a cold storage solution, was developed for isolation of high-quality islets from human pancreata for transplantation. This approach markedly improves the islet yield, purity and viability, and the isolation success rate. In this method, the pancreas was digested first in warm collagenase solution for up to 20 minutes. After decanting the enzyme solution, partially digested tissue was dissociated by gentle agitation in cold LAP-1 solution without additional collagenase. The digested tissues were stored in cold LAP-1 solution until islet purification on Euro-Ficoll. Forty-six islet isolations were performed consecutively by the new method (group 1). These results were compared to those obtained earlier with 46 consecutive isolations, using our previous method that had been used before development of the new method (group 2). Our old method was a modification of Ricordi's method involving only warm collagenase digestion and the storage of digested tissues in cold Hanks balanced salt solution. All pancreata were partial, containing the body and tail. There were no significant differences in both groups with regard to the donor age, cold ischemic time, harvesting conditions, and pancreatic weight. Pancreas digestion was completed in approximately 1 hour in both groups. The isolation success rate as determined by viable islets after 2 days in culture was 93.5% (43 of 46 cases) in group 1, and 56.5% (26 of the 46) in group 2. Immediately after isolation, the new method yielded a total of 335,739 +/- 36,244 islets equivalent to 150 microm (IEQ) and 6,233 +/- 681 IEQ/g of pancreas with 83 +/- 2.5% purity, whereas the old method yielded a total of 195,587 +/- 25,242 IEQ and 3,763 +/- 5,509 IEQ/g with 69.2 +/- 4.7% purity. Isolated islets in group 1 maintained a good three-dimensional structure, displayed normal insulin release to high glucose stimulation in vitro, and restored euglycemia after transplantation into streptozotocin-diabetic athymic mice. The two-step digestion method provides a sufficient number of islets for transplantation from a single pancreas. PMID- 10707936 TI - Cytokine balance and lipid antigen presentation in the NOD mouse pancreas during development of insulitis. AB - The role of cytokine balance and lipid antigen presentation in the development of diabetes was studied using immunohistochemistry of cytokines in the pancreas of non-obese diabetic mice (NOD) and BALB/c mice at various ages. In both the NOD and BALB/c mice, interleukin 10 (IL-10) was expressed in the islets. IL-10 was also present in the epithelial cells of the exocrine tissue in both strains. In the NOD mice, IL-10 disappeared from both the islets and the exocrine tissue at 16 weeks of age. At this age, IL-10 was still present in the islets and exocrine tissue of the BALB/c pancreata. IL-10 was not present in the pancreata of diabetic NOD mice. IL-6 first appeared in the pancreas at 10 weeks of age and disappeared at the age of 16 weeks in both NOD and BALB/c mice. It was present in the endothelial cells. Neither the pancreata of normal BALB/c mice nor NOD mice at 2-16 weeks of age contained tumor necrosis factor alpha (TNF-alpha), interferon gamma (IFN-gamma), IL-4, or IL-12. At 8 weeks of age, a few IL-2+ cells were found in the pancreas of one of three NOD mice. CD1d was already present in both strains at 2 weeks of age but disappeared from the NOD mice at 16 weeks of age. CD1d localized to walls of tubular structures probably representing collecting tubules. These results suggest that in the NOD mice the disappearance of the T(H0), T(H1), and T(H2) responses inhibiting IL-10 from the islets at the age of 16 weeks may trigger the final stage of the immune response leading to overt diabetes. The simultaneous disappearance of CD1d suggests that activation of immune responses against lipid antigens does not play a role in this stage of the disease. PMID- 10707937 TI - The role of lipid antigen presentation, cytokine balance, and major histocompatibility complex in a novel murine model of adoptive transfer of insulitis. AB - After adoptive transfer of insulitis from nonobese diabetic (NOD) mice, leukocytes accumulate in the pancreas of SCID/SCID and NOD/SCID mice. These cells express classical antigen-presenting molecules and costimulators of T-cell activation and adhesion molecules involved in homing. The aim of the present study was to study the expression of cytokines involved in regulation of the T(H1)/T(H2) balance by these cells, the role of lipid antigen presentation in the local immune system activation in the pancreas during onset of insulitis, and the role of major histocompatibility complex in this process. Splenocytes from NOD and BALB/c mice were injected intraperitoneally to SCID/SCID and NOD/SCID mice. Sections from the pancreata of these injected mice were stained for cytokines (tumor necrosis factor alpha [TNF-alpha], interferon gamma [IFN-gamma], CD1d, interleukin 2 [IL-2], IL-4, IL-6, IL-10, and IL-12). Some SCID/SCID and NOD/SCID mice injected with NOD splenocytes developed a severe disease. IL-10 was expressed in almost all the animals: in exocrine pancreas, large groups of infiltrating lymphocytes, endothelia of blood vessels, pancreatic islets, and interstitial tissue. CD1d was found in most of the mice: in the endothelia of collecting ducts and blood vessels of the pancreas, lymphocytic infiltrates, interstitial tissue, septae, islets, and a pancreatic lymph node. TNF-alpha was expressed notably more often in the pancreata of NOD/SCID than SCID/SCID mice. It was found between pancreatic lobules, in the epithelia of collecting ducts, endothelia of blood vessels, islets, capillaries, infiltrates, and septae. IL-6 was expressed more in the SCID/SCID than in the NOD/SCID mice. It was seen in infiltrates, walls of blood vessels, around islets, and in connective tissue. IFN gamma was found only in the pancreata of SCID/SCID and NOD/SCID mice injected with NOD splenocytes. The expression of IL-2 and IL-12 was very scarce. IL-4 was not expressed at all. The present study clearly shows that antigen presentation has a role in the development of autoimmune diseases after adoptive transfer of splenocytes from diseased mice to intact ones and that IL-10 may have a central role in the control of the disease process. PMID- 10707939 TI - Cryptosporidiosis: the treatment dilemma. PMID- 10707938 TI - Multiple polypeptide hormone expression in pancreatic islet cell carcinomas derived from phosphoenolpyruvatecarboxykinase-SV40 T antigen transgenic rats. AB - Transgenic rats carrying a PEPCK-SV40 large T-antigen (TAg) transgene rapidly develop numerous pancreatic islet cell neoplasms, the cells of which express TAg. Although many of the larger neoplasms contain relatively undifferentiated cells, many tumors contain areas of well-differentiated cells with abundant endoplasmic reticulum (ER) and secretory granules for endocrine hormones like those observed in normal pancreatic islets. In the well-differentiated lesions, glucagon producing alpha-cells, insulin-producing beta-cells, and somatostatin-producing delta-cells are readily identifiable morphologically under the electron microscope. Beta-cells were observed in all normal and hyperplastic islets, and nests of these cells were scattered throughout the larger neoplasms. These nests varied from small clusters of epithelium-like cells that stain intensely for insulin, to sheets of small, basophilic cells that stain more diffusely for the hormone. Alpha-cells were also present in all of the normal and hyperplastic islets, but in larger hyperplastic islets, the peripheral localization was absent. Larger neoplasms contained many nests of glucagon-expressing cells, as well as scattered glucagon-producing single cells. Delta-cells were rarely observed in the hyperplastic islets and in the neoplasms. Blood-glucose levels were unaltered in the transgenic animals relative to their nontransgenic litter mates. Thus although these islet cell neoplasms express several polypeptide hormones, there is no obvious clinical effect of such expression in vivo. PMID- 10707940 TI - Influence of bacteria from the duodenal microbiota of patients with symptomatic giardiasis on the pathogenicity of Giardia duodenalis in gnotoxenic mice. AB - Recent studies have shown that the intestinal microbiota is essential for the pathogenicity but not for the multiplication of Giardia duodenalis in the intestinal lumen. The microbial components responsible for this phenomenon are not known. Twenty-eight facultative and three strictly anaerobic micro-organisms were isolated from the dominant duodenal microbiota of five patients with symptomatic giardiasis. The bacterial combinations from each patient were associated with groups (GN) of germ-free mice. Five days after the association, when their faecal populations ranged from 10(7) to 10(9) cfu/g, all groups were inoculated intragastrically with 10(5) viable trophozoites of G. duodenalis strain BT6. Two groups of germ-free (GF) and conventional (CV1) mice were also infected. Gnotobiotic animals were killed 10 days after infection and GF and CV1 animals were killed 10, 20 and 30 days after infection. More marked pathological alterations were detected in CV1 mice when compared with GF animals. Gnotobiotic animals showed intermediate pathological alterations between CV1 and GF mice. The CV1 and GF groups became infected by day 3 and faecal cyst levels were similar in both groups throughout the experiment. Total and G. duodenalis-specific IgA levels in the intestinal fluid and G. duodenalis-specific IgM and IgG levels in the serum increased during the infection and were higher in CV1 animals at all times tested when compared with GF mice. The present results confirm the stimulatory activity of the intestinal microbiota on the pathogenicity of G. duodenalis, and some combinations of microbial components of the dominant duodenal ecosystem from patients with symptomatic giardiasis can partially develop this function. However, none of these combinations was able to stimulate the protozoan pathogenicity in the same manner as the entire intestinal microbiota. PMID- 10707941 TI - Binding of von Willebrand factor by coagulase-negative staphylococci. AB - Coagulase-negative staphylococci (CNS) are the most common infectious micro organisms isolated from prosthetic devices. To determine whether von Willebrand factor (vWF) acts as an adhesin in bacterial recognition, bacterial binding of recombinant vWF (rvWF) was studied. Eleven CNS strains, belonging to S. epidermidis, S. haemolyticus and S. hominis species, bound soluble rvWF, but to a lesser extent than S. aureus. S. epidermidis strain H2-W bound 125I-labelled rvWF in a dose-dependent manner. The binding could be inhibited by unlabelled rvWF and thrombospondin, but not by fibrinogen, vitronectin or the carbohydrates N acetylgalactoseamine, D-galactose, D-glucose, and D-fucose. Pre-incubation of rvWF with type I collagen and Arg-Gly-Asp-Ser (RGDS) peptides did not inhibit binding, whereas pre-incubation of rvWF with heparin decreased binding significantly. The interaction between CNS and rvWF was sensitive to proteinase treatment of bacterial cells. CNS strains bound to immobilised rvWF an extent greater or equal to the positive control strain S. aureus Cowan I. rvWF binding structures from bacterial cell wall were detected by immunoblot. Cowan I strain had 140-, 90- and 38-kDa binding molecules. S. haemolyticus strain SM131 and S. epidermidis strain H2-W had two (120 and 60 kDa) and five (120, 90, 60, 52 and 38 kDa) binding molecules, respectively. Similar binding structures were formed when cell wall extracts from these strains were incubated with thrombospondin. These results indicate that specific ligand-receptor interaction between CNS and rvWF may contribute to bacterial adhesion and colonisation on biomaterial surfaces. Heparin-binding domains of rvWF might be the crucial regions for bacterial attachment. rvWF and thrombospondin may recognise similar molecules in staphylococcal cell wall extracts. PMID- 10707942 TI - Bactericidal activity of a monocytic cell line (THP-1) against common respiratory tract bacterial pathogens is depressed after infection with respiratory syncytial virus. AB - Non-typable Haemophilus influenzae, Streptococcus pneumoniae, Moraxella catarrhalis and respiratory syncytial virus (RSV) are commonly isolated from patients during the course of chronic obstructive pulmonary disease (COPD). Earlier studies found that virus infection enhanced binding of bacterial respiratory pathogens to epithelial cells in vitro. The objective of the present study was to assess the effect of RSV infection of a human monocytic cell line on bactericidal activity and cytokine production in response to these bacterial respiratory pathogens. The effect of RSV infection on binding, uptake and intracellular killing of bacteria by a human monocytic leukaemia cell line, THP 1, was assessed. Cell culture supernates were examined with a mouse fibroblast cell assay for tumour necrosis factor-alpha (TNF-alpha) bioactivity. Expression of CD14, CD11a, CD18, CD15 and CD29 on uninfected and RSV-infected THP-1 cells was assessed by flow cytometry in relation to differences in bacterial binding. RSV infection of THP-1 cells significantly decreased their ability to bind and kill bacteria. Compared with uninfected cells, fewer bacteria bound to RSV infected THP-1 cells and the surface antigens that have been reported to bind bacteria were expressed at lower levels on RSV-infected cells. RSV-infected cells incubated with bacteria exhibited less TNF-alpha bioactivity than uninfected cell incubated with bacteria. The results elucidate some of the mechanisms involved in the increased susceptibility of virus-infected patients to secondary bacterial infection. Reduced bacterial killing by virus-infected monocytes might contribute to reduced clearance of bacteria from the respiratory tract and damage elicited by the bacteria or cytokine response in COPD patients. PMID- 10707943 TI - Isolation and characterisation of sialidase from a strain of Streptococcus oralis. AB - Streptococcus oralis, the most virulent of the viridans streptococci, produces a sialidase and this exo-glycosidase has been implicated in the disease process of a number of pathogens. The sialidase of S. oralis strain AR3 was purified in order to understand the characteristics of this putative virulence determinant. The enzyme isolated as a high mol. wt aggregate (c. 325 kDa) was purified 4520 fold from late exponential phase cultures by a combination of ultrafiltration, ammonium sulphate precipitation, ion-exchange and gel filtration chromatography. The sialidase component had a mol.wt of 144 kDa as determined by SDS-PAGE analysis. The purified sialidase released N-acetylneuraminic acid from a range of sialoglycoconjugates including human alpha1-acid glycoprotein, bovine submaxillary mucin, colominic acid and sialyl-alpha2,3- and sialyl-alpha2,6 lactose. Also, N-glycolylneuraminic acid was cleaved from bovine submaxillary mucin. The sialidase had a Km of 11.8 microM for alpha1-acid glycoprotein, was active over a broad pH range with a pH optimum of 6.0 and cleaved alpha2,3-, alpha2,6- and alpha2-8-sialyl glycosidic linkages with a marked preference for alpha2,3-linkages. The enzyme was competitively inhibited by the sialic acid derivative, 2,3-dehydro-N-acetylneuraminic acid, with a K(IC) of 1.2 microM. The characteristics of the purified sialidase would support a nutritional role for this enzyme that may be significant in the proliferation of this organism in the oral cavity and at extra-oral sites in association with life-threatening infections. PMID- 10707944 TI - Epithelial cell response to challenge of bacterial lipoteichoic acids and lipopolysaccharides in vitro. AB - Accumulating dental plaque at the gingival margin contains lipoteichoic acids (LTAs) from the cell walls of gram-positive bacteria. In subgingival plaque associated with periodontal disease the amount of lipopolysaccharides (LPSs) from gram-negative bacteria increases. As the gingival junctional epithelium (JE) is an important structural and functional tissue, participating in the first line defence against apical advancement of dental plaque, this study examined the direct effects of LTAs (from Streptococcus mutans and S. sanguis) and LPSs (from Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Treponema denticola and Escherichia coli) on two epithelial cell lines (HaCaT and ERM) and a culture model for human JE. The cells were exposed to the LTAs or LPSs (10-50 microg/ml) for variable periods of time. None of the bacterial surface components had any effect on primary adhesion or on the epithelial attachment of the JE cultures. However, cell growth and mitotic activity were consistently reduced in all cultures treated with LTAs. In contrast, LPSs showed only slight or no effects on cell growth and mitotic activity depending on the epithelial cells used. This suggests that LPSs, despite their established role as modulators of inflammation, do not have direct harmful effects - at the concentrations found in dental plaque and gingival crevicular fluid - which would explain the mechanism of epithelial degeneration and detachment from the tooth surface. However, the LTAs appear to inhibit the renewal of epithelium and may thus contribute to degeneration of coronal JE and subgingival colonisation by periodontal pathogens. PMID- 10707945 TI - Protective features of monoclonal antibodies to Escherichia coli during experimental infection of mice with homologous and heterologous serotypes of E. coli. AB - Murine monoclonal antibodies (MAbs) MT1F and ARM1-4, recognising proteins on the surface of untreated Escherichia coli O6:K-, protected 100% of mice challenged intraperitoneally with 2 x LD50 of the same strain. MAb MT1F protected 70% of animals challenged with 2 x LD50 of E. coli O111:B4, whereas ARM1-4 gave complete protection. Lower survival was observed in mice given either MAb and challenged with E. coli O128:K-, with values ranging from 30 to 42%. However, the protection afforded against E. coli O111:B4 and E. coli O128:K- was significantly improved when the mice were pre-treated with a mixture of the two MAbs. Control mice, pre treated with unrelated ascitic fluid and challenged with any of the E. coli serotypes, showed 100% mortality and organ histological lesions resembling those of the early stages of septic shock. The mice had high levels of circulating endotoxin and tumour necrosis factor-alpha (TNF-alpha) at 90 min after challenge. In contrast, mice treated with MAbs and surviving the infection displayed moderate histological lesions, enhanced bacterial clearance and lower serum levels of TNF-alpha, despite circulating endotoxin levels that were higher than in the control group. Protection by the MAbs was probably due to the prevention of the bacterial spread to organs and of the cascade of events leading to septic shock. This occurred in spite of the presence of high levels of circulating endotoxin. PMID- 10707946 TI - Development, characterisation and diagnostic application of monoclonal antibodies against Yersinia pestis fibrinolysin and coagulase. AB - A library of monoclonal antibodies (MAbs) which recognised different epitopes of Yersinia pestis fibrinolysin (Fib) was developed. These MAbs were species specific and demonstrated no cross-reaction in indirect immunofluorescence tests (IIFT) with other gram-negative bacteria possessing plasminogen activator activity. All the MAbs provided equally high levels of immunofluorescence with pPst+ Y. pestis strains cultivated at 37 degrees C and at 28 degrees C. In all cases, the MAbs inhibited both fibrinolytic and coagulase (Coag) activities of Y. pestis in Fib-activity inhibition and coagulase-activity inhibition reactions, and reacted with 35- and 37-kDa proteins of Y. pestis in immunoblotting, demonstrating bifunctional activity possibly similar to the properties of MAbs produced by hybrid hybridomas. On the basis of these and earlier studies, the immunochemical identity of Fib and Coag, two distinct subunits of a bifunctional fusion protein whose specific functional activity depends upon the temperature factor, was established. A new rapid, cheap, strictly specific and safe dot-ELISA based on the use of MAb against Y. pestis Fib (MAb-Fib) for reliable identification of Y. pestis strains was developed. This technique has great advantages over monoclonal diagnostic kits based on the use of MAb against Y. pestis fraction I (FI) because it allows detection of plague bacilli grown at 37 degrees C as well as at 28 degrees C. This dot-ELISA will be valuable as a clinical diagnostic tool and might be applicable to field studies and plague surveillance. PMID- 10707947 TI - A clinical, microbiological and economic analysis of a national service for the rapid molecular diagnosis of tuberculosis and rifampicin resistance in Mycobacterium tuberculosis. AB - A clinical, microbiological and economic study of a national rapid molecular service for the identification of Mycobacterium tuberculosis and the determination of rifampicin resistance in smear-positive sputum samples (and other primary specimens) was performed. Ninety-one primary specimens, of which 55 were smear-positive sputum, were examined by molecular and conventional assays. Concordance of molecular results from smear-positive sputum specimens with tuberculosis diagnosis and rifampicin resistance by conventional analysis was 52 (94.5%) of 55 and 44 (91.7%) of 48, respectively. Concordance of molecular analysis on all primary specimens was 81 (89.0%) of 91 (diagnosis) and 55 (90.2%) of 61 (rifampicin resistance). Approximately 28 days were saved in the time to diagnosis by using the molecular assay. Hospitals can reduce the cost of inappropriate isolation of patients with risk factors for multiple drug-resistant tuberculosis (MDRTB) who subsequently are shown to have drug-sensitive tuberculosis. At one hospital potential annual savings were between pound sterling 50000 and pound sterling 150000. Of the nine MDRTB cases identified, all had a previous diagnosis of tuberculosis, 78% were born overseas, 44% were known to be non-compliant with therapy, but only one case (12.5%) was HIV positive. HIV status was not significantly different between MDRTB and drug-sensitive tuberculosis cases. Over 75% of specimens were taken while the patient was on therapy. Isolates from >50% of the MDRTB cases were resistant to three or more drugs and one was resistant to seven drugs. All patients were placed on additional therapy once the molecular result was known; this was subsequently modified based on the results of in-vitro drug susceptibility testing. All survived at least 6 months of follow-up. There was no difference in the proportion of successful cultures from smear-positive samples from patients with drug-sensitive tuberculosis or MDRTB who were on therapy. Molecular rifampicin resistance assays are reliable for diagnosis in cases with smear-positive disease. PMID- 10707948 TI - Use of rep-PCR to define genetic relatedness among Bacteroides fragilis strains. AB - Bacteroides fragilis, a component of the normal flora and an important anaerobic pathogen in non-intestinal endogenous infections, has recently been associated with enteric diseases. In this study, 41 B. fragilis strains were analysed in relation to their genetic diversity. This collection included two reference strains (ATCC 23745 and 25285), 20 isolates from non-intestinal infections, six from intestinal infections, five from intestinal microflora and eight from an aquatic environment. The fingerprints were generated by using two repetitive sequences (REP and ERIC) as primers to PCR (rep-PCR). A dendrogram was obtained with the Taxotron Program. Three clusters (threshold genotypes I, II and III) were observed when the genetic distance was 0.30. These results confirm previous data found regarding the genotypical diversity of B. fragilis. PMID- 10707949 TI - In-situ detection of Aspergillus fumigatus. AB - An in-situ hybridisation (ISH) technique to detect Aspergillus fumigatus in infected tissues was developed in which 568-bp, 333-bp and 154-bp PCR products of the alkaline proteinase gene were employed. Dot-blot hybridisation with the 568 bp probe on a membrane containing genomic DNA from several different fungi including A. flavus, A. niger, Penicillium spp., Mucor racemosus or Pseudallescheria boydii gave negative results. ISH was done on formalin-fixed, paraffin-embedded pulmonary tissues from rats infected with A. fumigatus and renal tissues from mice infected with A. fumigatus, A. flavus or A. niger. The 568-bp probe reacted strongly in ISH with both A. fumigatus and A. flavus, and weakly with A. niger. The 333-bp probe also reacted in ISH with A. fumigatus and A. flavus, although the intensity was weaker. However, in ISH with the 154-bp probe, there was no positive signal with any Aspergillus spp. These results demonstrate that A. fumigatus and A. flavus can be specifically detected in infected tissues by ISH with the 568-bp probe. This technique could be applicable to clinical specimens for molecular diagnosis of aspergillus infections. PMID- 10707950 TI - Factors affecting the adhesion of Candida albicans to epithelial cells of insulin using diabetes mellitus patients. AB - This study investigated the influence of the carbon source of the growth medium, strains of Candida albicans and source of epithelial cells, and the influence of smoking and gender, on the adhesion of C. albicans to epithelial cells from insulin-using diabetic patients. Adhesion was determined by an autologous adhesion assay with exfoliated buccal or palatal epithelial cells and one strain of C. albicans isolated from each patient. The type strain CBS 562 was also used. Glucose or sucrose were used as the predominant carbon sources of the growth medium. The autologous strain of C. albicans adhered selectively to the oral mucosa of diabetic patients. Palatal epithelial cells retained significantly more C. albicans in vivo and adhesion was influenced by the availability of sugars in the growth medium and the strain of C. albicans. PMID- 10707951 TI - Rapid discrimination between methicillin-sensitive and methicillin-resistant Staphylococcus aureus by intact cell mass spectrometry. AB - Rapid, accurate discrimination between methicillin-sensitive Staphylococcus aureus (MSSA) and methicillin-resistant S. aureus (MRSA) strains is essential for appropriate therapeutic management and timely intervention for infection control. A rapid method involving intact cell mass spectrometry (ICMS) is presented that shows promise for identification, discrimination of MSSA from MRSA and typing. In ICMS, cells from a bacterial colony are emulsified in a chemical matrix, added to a sample slide, dried and analysed by matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOF-MS). This technique examines the chemistry of the intact bacterial cell surface, yielding spectra consisting of a series of peaks from 500 to 10000, which represent the mass:charge (m:z) ratios. Each peak corresponds to a molecular fragment released from the cell surface during laser desorption. Specimens can be prepared in a few seconds from plate cultures and a spectrum can be obtained within 2 min. ICMS spectra for 20 staphylococcal isolates showed characteristic peaks, some of which were conserved at species level, some at strain level and some were characteristic of the methicillin susceptibility status of the strain. ICMS may have potential for MRSA identification and typing, and may improve infection control measures. PMID- 10707952 TI - Nuclear hormone receptor coregulators in action: diversity for shared tasks. PMID- 10707953 TI - Progesterone induces focal adhesion in breast cancer cells MDA-MB-231 transfected with progesterone receptor complementary DNA. AB - Since the effects of progesterone are mediated mainly via estrogen-dependent progesterone receptor (PR), the expression of the effects of progesterone may be masked or overridden by the influence of estrogen under conditions in which priming with estrogens is required. We have established a PR-positive but estrogen receptor-alpha (ER-alpha) negative breast cancer cell model by transfecting PR cDNA into ER-alpha- and PR-negative MDA-MB-231 cells in order that the functions of progesterone can be studied independently of estrogens. We have demonstrated using this model that progesterone markedly inhibited cell growth. We have also discovered that progesterone induced remarkable changes in cell morphology and specific adhesion structures. Progesterone-treated cells became considerably more flattened and well spread than vehicle-treated control cells. This was associated with a striking increase of stress fibers, both in number and diameter, and increased focal contacts as shown by the staining of focal adhesion proteins paxillin and talin. There were also distinct increases in tyrosine phosphorylation of focal adhesion protein paxillin and focal adhesion kinase in association with increased focal adhesion. The staining of tyrosine phosphorylated proteins was concentrated at focal adhesions in progesterone treated cells. More interestingly, monoclonal antibody (Ab) to beta1 integrin was able to inhibit progesterone-induced cell spreading and formation of actin cytoskeleton. To our knowledge, this is the first report describing a direct effect of progesterone in inducing spreading and adhesion of breast cancer cells, and beta1-integrin appeared to play an essential role in the effect. It is known that the initial step of tumor metastasis is the breakaway of tumor cells from primary tumor mass when they lose the ability to attach. Hence, progesterone induced cell spreading and adhesion may have significant implications in tumor metastasis. PMID- 10707954 TI - C/EBPbeta (CCAAT/enhancer binding protein) controls cell fate determination during mammary gland development. AB - Deletion of the transcription factor CCAAT/enhancer binding protein (C/EBP)beta results in a severe inhibition of lobuloalveolar development in the mouse mammary gland. Because progesterone receptor (PR) is requisite for alveolar development, the expression of PR was investigated in C/EBPbeta-/- mice. Unexpectedly, the number of PR-positive cells, as well as the levels of PR mRNA, were elevated 3 fold in the mammary glands of C/EBPbeta-/- mice. Furthermore, in contrast to wild type nulliparous mice, in which PR distribution shifted from a uniform to nonuniform pattern between 8-12 weeks of age, C/EBPbeta-/- mice exhibited uniform PR distribution throughout all stages of mammary development analyzed. No change in C/EBPbeta mRNA levels was observed in the mammary glands of PR-/- mice, suggesting that PR acts in a pathway either in parallel to or downstream of C/EBPbeta. The overexpression and disrupted cellular distribution of PR in C/EBPbeta-/- mice were coincident with a striking 10-fold decrease in cell proliferation after acute steroid hormone treatment, assayed by incorporation of bromodeoxyuridine. In wild-type mice, PR and bromodeoxyuridine-positive cells were adjacent to each other and rarely colocalized. No differences in the level or pattern of PR expression were observed in the uterus, suggesting that C/EBPbeta influences PR in a mammary-specific fashion. Together, these data suggest that C/EBPbeta may control cell fate decisions in the mammary gland through the appropriate temporal and spatial expression of molecular markers, such as PR, that induce the proliferation of alveolar progenitor cells via juxtacrine mechanisms. PMID- 10707955 TI - Tamoxifen-bound estrogen receptor (ER) strongly interacts with the nuclear matrix protein HET/SAF-B, a novel inhibitor of ER-mediated transactivation. AB - The estrogen receptor (ER) is a ligand-dependent transcription factor that acts in a cell- and promoter-specific manner. Evidence suggests that the activity of the ER can be regulated by a number of other stimuli (e.g. growth factors) and that the effects of the ER are modulated by nuclear factors termed coregulators. While the interplay among these factors may in part explain the pleiotropic effects elicited by the ER, there are several other less well described mechanisms of control, such as interactions with the nuclear matrix. Here we report that the nuclear matrix protein/scaffold attachment factor HET/SAF-B is an ER-interacting protein. ER and HET/SAF-B interact in in vitro binding assays, with HET binding to both the ER DNA-binding domain and the hinge region. Coimmunoprecipitation experiments reveal that HET/SAF-B and ER associate in cell lines in the presence or absence of estradiol, but binding is increased by the antiestrogen tamoxifen. HET/SAF-B enhances tamoxifen antagonism of estrogen induced ER-mediated transactivation, but at high concentrations can inhibit both estrogen and tamoxifen-induced ER activity. HET/SAF-B-mediated repression of ER activity is dependent upon interaction with the ER-DBD. While the existence of high-affinity binding sites for the ER in the nuclear matrix has been known for some time, we now provide evidence of a specific nuclear matrix protein binding to the ER. Furthermore, our data showing that HET/SAF-B binds to ER particularly strongly in the presence of tamoxifen suggests that it may be important for the antagonist effect of tamoxifen. PMID- 10707956 TI - Human ERRgamma, a third member of the estrogen receptor-related receptor (ERR) subfamily of orphan nuclear receptors: tissue-specific isoforms are expressed during development and in the adult. AB - The nuclear receptor protein superfamily is a large group of transcription factors involved in many aspects of animal development, tissue differentiation, and homeostasis in the higher eukaryotes. A subfamily of receptors, ERRalpha and beta (estrogen receptor-related receptor alpha and beta), closely related to the ER, were among the first orphan nuclear receptors identified. These receptors can bind DNA as monomers and are thought to activate transcription constitutively, unaffected by beta-estradiol. Studies of the expression patterns of ERRalpha and gene disruption experiments of ERRbeta indicate that they play an important role in the development and differentiation of specific tissues in the mouse. In this work we demonstrate the existence in humans of a third member of this subfamily of receptors, termed ERRgamma, which is highly expressed in a number of diverse fetal and adult tissues including brain, kidney, pancreas, and placenta. The ERRgamma mRNA is highly alternatively spliced at the 5'-end, giving rise to a number of tissue-specific RNA species, some of which code for protein isoforms differing in the N-terminal region. Like ERRalpha and beta, ERRgamma binds as a monomer to an ERRE. A GAL4-ERRgamma fusion protein activates transcription in a ligand-independent manner in transfected HEK293 cells to a greater degree than either the GAL4-ERRalpha or -beta fusion proteins. PMID- 10707957 TI - Regulation of tissue factor gene expression in human endometrium by transcription factors Sp1 and Sp3. AB - Prior studies indicate that tissue factor (TF), the primary cellular initiator of hemostasis, is persistently up-regulated in human endometrial stromal cells (HESCs) undergoing progestin-induced decidualization in vivo and in vitro. The mechanism underlying progestin enhancement of TF mRNA and protein levels in these cells involves transcriptional activation of the TF gene. Transient transfections of HESCs with the truncated TF promoters driving the luciferase reporter gene have demonstrated that the region spanning -111 to +14 bp retained differential progestin-enhancing effects. We now demonstrate that RU486 displays inhibitory effects on the progestin-induced TF promoter activity, confirming the involvement of the progesterone receptor. Since the TF minimal promoter (pTF 111 spanning 111 to +14 bp) contains three overlapping Sp1 and three Egr-1 sites, the present study determined whether Sp1 and/or Egr-1 were required for progestin-regulated TF expression. The results indicate that the three Sp1 sites are primarily responsible for both the constitutive and progestational activity of the pTF 111 promoter, whereas the Egr-1 sites have only a minor involvement in both activities. Overexpression of the Sp1 protein resulted in greater than a 6-fold induction in TF promoter activity. In contrast, no enhancement was observed when the Sp3 protein was overexpressed. The concomitant overexpression of Sp1 and Sp3 demonstrated that Sp3 completely blocked the induction of TF promoter activity by Sp1. Moreover, the addition of 10 nM mithramycin, a concentration that inhibits Sp1 binding to target DNA, blocked the progestational induction of TF mRNA expression. Immunohistochemical studies demonstrated increased Sp1 levels in perivascular stromal cells in secretory phase compared with proliferative phase endometria. In contrast, Sp3 expression was greatly decreased in stromal cells of secretory, compared with proliferative phase tissues. The levels of Egr-1 were low in both proliferative and secretory endometria. Immunocytochemistry of E2 vs. E2 + medroxyprogesterone acetate-treated HESCs demonstrated a dramatic reduction in Sp3 expression after progestin treatment, and Northern blots demonstrated progestational increases in Sp1 and reduction in Sp3 mRNA expression compared with controls. Taken together, our results demonstrate that progestin enhancement of TF gene expression in HESCs is mediated principally by Sp1. We propose that progestins regulate HESC TF gene expression in vivo by altering the ratio of Sp1 to Sp3 nuclear factors. PMID- 10707958 TI - The polymorphic N terminus in human vitamin D receptor isoforms influences transcriptional activity by modulating interaction with transcription factor IIB. AB - The human vitamin D receptor (hVDR) is a ligand-regulated transcription factor that mediates the actions of the 1,25-dihydroxyvitamin D3 hormone to effect bone mineral homeostasis. Employing mutational analysis, we characterized Arg-18/Arg 22, hVDR residues immediately N-terminal of the first DNA binding zinc finger, as vital for contact with human basal transcription factor IIB (TFIIB). Alteration of either of these basic amino acids to alanine also compromised hVDR transcriptional activity. In contrast, an artificial hVDR truncation devoid of the first 12 residues displayed both enhanced interaction with TFIIB and transactivation. Similarly, a natural polymorphic variant of hVDR, termed F/M4 (missing a FokI restriction site), which lacks only the first three amino acids (including Glu-2), interacted more efficiently with TFIIB and also possessed elevated transcriptional activity compared with the full-length (f/M1) receptor. It is concluded that the functioning of positively charged Arg-18/Arg-22 as part of an hVDR docking site for TFIIB is influenced by the composition of the adjacent polymorphic N terminus. Increased transactivation by the F/M4 neomorphic hVDR is hypothesized to result from its demonstrated enhanced association with TFIIB. This proposal is supported by the observed conversion of f/M1 hVDR activity to that of F/M4 hVDR, either by overexpression of TFIIB or neutralization of the acidic Glu-2 by replacement with alanine in f/M1 hVDR. Because the f VDR genotype has been associated with lower bone mineral density in diverse populations, one factor contributing to a genetic predisposition to osteoporosis may be the F/f polymorphism that dictates VDR isoforms with differential TFIIB interaction. PMID- 10707959 TI - Endocrine disrupting chemicals, phthalic acid and nonylphenol, activate Pregnane X receptor-mediated transcription. AB - Recently, Pregnane X receptor (PXR), a new member of the nuclear receptor superfamily, was shown to mediate the effects of several steroid hormones, such as progesterone, glucocorticoid, pregnenolone, and xenobiotics on cytochrome P450 3A genes (CYP3A) through the specific DNA sequence for CYP3A, suggesting that PXR may play a role in steroid hormone metabolism. In this paper, we demonstrated that phthalic acid and nonylphenol, endocrine-disrupting chemicals (EDCs), stimulated PXR-mediated transcription at concentrations comparable to those at which they activate estrogen receptor-mediated transcription using a transient reporter gene expression assay in COS-7 cells. However, bisphenol A, another EDC, had no effect on PXR-mediated transcription, although this chemical significantly enhanced ER-mediated transcription. In the yeast two-hybrid protein interaction assay, PXR interacted with two nuclear receptor coactivator proteins, steroid hormone receptor coactivator-1 and receptor interacting protein 140, in the presence of phthalic acid or nonylphenol. Thus, EDC-occupied PXR may regulate its specific gene expression through the receptor-coactivator interaction. In contrast, these EDCs had no effect on the interaction between PXR and suppressor for gal 1, a component of proteasome. Finally, the expression of CYP3A1 mRNA in the liver of rats exposed to phthalic acid or nonylphenol markedly increased compared with that in rats treated with estradiol, bisphenol A, or ethanol as assessed by competitive RT-PCR. These data suggest that EDCs may affect endocrine functions by altering steroid hormone metabolism through PXR. PMID- 10707960 TI - Synergy of activin and ciliary neurotrophic factor signaling pathways in the induction of vasoactive intestinal peptide gene expression. AB - Activin, a member of the transforming growth factor-beta superfamily, can regulate neuropeptide gene expression in the nervous system and in neuroblastoma cells. Among the neuropeptide genes whose expression can be regulated by activin is the gene encoding the neuropeptide vasoactive intestinal peptide (VIP). To investigate the molecular mechanisms by which activin regulates neuronal gene expression, we have examined activin's regulation of VIP gene expression in NBFL neuroblastoma cells. We report here that NBFL cells respond to activin by increasing expression of VIP mRNA. Activin regulates VIP gene transcription in NBFL cells through a 180-bp element in the VIP promoter that was previously characterized to be necessary and sufficient to mediate the induction of VIP by the neuropoietic cytokines and termed the cytokine response element (CyRE). We find that the VIP CyRE is necessary and sufficient to mediate the transcriptional response to activin. In addition, ciliary neurotrophic factor (CNTF), a neuropoietic cytokine, synergizes with activin to increase VIP mRNA expression and transcription through the VIP CyRE. Mutations in either the Stat (signal transducer and activator of transcription) or AP-1 sites within the CyRE that reduce the response to CNTF, also reduce the response to activin. However, mutating both the Stat and AP-1 sites within the wild-type CyRE, while reducing the separate responses to either activin or CNTF, eliminates the synergy between them. These data suggest that activin and CNTF, two factors that appear to signal though distinct pathways, activate VIP gene transcription through a common transcriptional element, the VIP CyRE. PMID- 10707961 TI - Thyrotropin induces SOCS-1 (suppressor of cytokine signaling-1) and SOCS-3 in FRTL-5 thyroid cells. AB - TSH has multiple physiological roles: it is required for growth, differentiation, and function of the thyroid gland, and it regulates transcription of interferon gamma (IFN-gamma)-responsive genes in thyrocytes, including genes for the major histocompatibility complex and intercellular adhesion molecule-1. This report demonstrates that TSH induces the expression of suppressor of cytokine signaling (SOCS)-1 and -3 proteins and alters the phosphorylation state of signal transducer and activator of transcription (STAT) proteins STAT1 and STAT3. The expression of SOCS-1 and SOCS-3 and the phosphorylation state of STAT1 and STAT3 were examined after treatment with TSH or IFN-gamma in either TSH-sensitive FRTL 5 thyroid cells or TSH-insensitive FRT and buffalo rat liver (BRL) cells, which lack functional TSH receptors. SOCS-1 and SOCS-3 are constitutively expressed in FRTL-5 cells, but not in FRT and BRL cells. IFN-gamma up-regulated SOCS-1 and SOCS-3 RNA and protein in FRTL-5 cells, as reported previously for nonthyroid cells. Interestingly, TSH also significantly induced SOCS-1 and SOCS-3 in FRTL-5 cells, but not in FRT and BRL cells. When SOCS-1 or SOCS-3 was overexpressed in FRTL-5 cells, STAT1 phosphorylation at Y701 and STAT1/DNA complex formation in response to IFN-gamma were reduced. Furthermore, overexpression of either SOCS-1 or SOCS-3 significantly inhibited the IFN-gamma-mediated transactivation of the rat ICAM-1 (intercellular adhesion molecule-1) promoter. TSH and IFN-gamma had different effects on STAT1 and STAT3 phosphorylation. The phosphorylation of Y701 in STAT1, which is responsible for homodimer formation, nuclear translocation, and DNA binding, was specifically stimulated by IFN-gamma, but not by TSH or forskolin. However, the phosphorylation of S727 in STAT1 was induced by IFN gamma, TSH, and forskolin. TSH induced phosphorylation of both Y705 and S727 in STAT3, while IFN-gamma phosphorylated only the Y705. In addition, we found that SOCS-3 was associated with JAK1 and JAK2 and that these associations were stimulated by TSH. These findings demonstrate that TSH induces SOCS in thyroid cells and provides the evidence of signal cross-talk between TSH and cytokines in thyroid cells. PMID- 10707962 TI - TFE3, a transcription factor homologous to microphthalmia, is a potential transcriptional activator of tyrosinase and TyrpI genes. AB - Microphthalmia gene encodes a basic helix-loop-helix-leucine zipper (bHLH-Zip) transcription factor involved in the development of the melanocyte lineage and plays a key role in the transcriptional regulation of the melanogenic enzymes, tyrosinase and TyrpI. Recently, we have shown that Microphthalmia mediates the melanogenic effects elicited by alphaMSH that up-regulates the expression of tyrosinase through the activation of the cAMP pathway. Therefore, Microphthalmia appears as a principal gene in melanocyte development and functioning. Among the transcription factors of the bHLH-Zip family, TFE3 and TFEB show a remarkably elevated homology with Microphthalmia. These observations prompted us to investigate the role of TFE3 and TFEB in the regulation of tyrosinase and TyrpI gene transcription. We show in this report that overexpression of TFE3 stimulates the tyrosinase and TyrpI promoter activities, while TFEB acts only on the TyrpI promoter. TFE3 and TFEB elicit their effects mainly through the binding to Mbox (AGTCATGTGCT) and Ebox motifs (CATGTG) of tyrosinase and TyrpI promoters. In B16 melanoma cells, the high basal expression of TFE3 is down-regulated by forskolin and by alphaMSH. Interestingly, endogenous TFE3 cannot bind as homodimers to the Mbox, and we did not detect TFE3/Mi heterodimers. According to these data, TFE3 is clearly endowed with the capacity to regulate tyrosinase and TyrpI gene expression. However, TFE3 binding to the melanogenic gene promoters is hindered, thereby preventing its potential melanogenic action. In specific physiological or pathological conditions, the recovery of its binding function would make TFE3 an important element in melanogenesis regulation. PMID- 10707963 TI - Specificity and promiscuity among proneural proteins. PMID- 10707964 TI - Bipolar cells in the spotlight: cause for excitement. PMID- 10707965 TI - Learning to like your voice: developing selectivity to birdsong. PMID- 10707966 TI - Ion channel surprises: prokaryotes do it again! PMID- 10707967 TI - CREB couples neurotrophin signals to survival messages. PMID- 10707968 TI - Moving colors in the lime light. PMID- 10707969 TI - Molecular insights into mRNA transport and local translation in the mammalian nervous system. PMID- 10707970 TI - Neuropilin-2 is required in vivo for selective axon guidance responses to secreted semaphorins. AB - Neuropilins are receptors for class 3 secreted semaphorins, most of which can function as potent repulsive axon guidance cues. We have generated mice with a targeted deletion in the neuropilin-2 (Npn-2) locus. Many Npn-2 mutant mice are viable into adulthood, allowing us to assess the role of Npn-2 in axon guidance events throughout neural development. Npn-2 is required for the organization and fasciculation of several cranial nerves and spinal nerves. In addition, several major fiber tracts in the brains of adult mutant mice are either severely disorganized or missing. Our results show that Npn-2 is a selective receptor for class 3 semaphorins in vivo and that Npn-1 and Npn-2 are required for development of an overlapping but distinct set of CNS and PNS projections. PMID- 10707971 TI - Neuropilin-2 regulates the development of selective cranial and sensory nerves and hippocampal mossy fiber projections. AB - Neuropilin-1 and neuropilin-2 bind differentially to different class 3 semaphorins and are thought to provide the ligand-binding moieties in receptor complexes mediating repulsive responses to these semaphorins. Here, we have studied the function of neuropilin-2 through analysis of a neuropilin-2 mutant mouse, which is viable and fertile. Repulsive responses of sympathetic and hippocampal neurons to Sema3F but not to Sema3A are abolished in the mutant. Marked defects are observed in the development of several cranial nerves, in the initial central projections of spinal sensory axons, and in the anterior commissure, habenulo-interpeduncular tract, and the projections of hippocampal mossyfiber axons in the infrapyramidal bundle. Our results show that neuropilin-2 is an essential component of the Sema3F receptor and identify key roles for neuropilin-2 in axon guidance in the PNS and CNS. PMID- 10707972 TI - The proneural gene amos promotes multiple dendritic neuron formation in the Drosophila peripheral nervous system. AB - In the Drosophila peripheral nervous system, proneural genes direct the formation of different types of sensory organs. Here, we show that amos is a novel proneural gene that promotes multiple dendritic (MD) neuron formation. amos encodes a basic-helix-loop-helix (bHLH) protein of the Atonal family. During embryonic development, amos is expressed in patches of ectodermal cells, and the expression is quickly restricted to sensory organ precursors. Loss of amos function eliminates MD neurons that remain in ASC;atonal mutants. Misexpression of amos generates ectopic MD and other types of neurons. Amos interacts with the ubiquitously expressed Daughter-less protein in vivo and in vitro. Our final misexpression experiments suggest that a domain located outside the DNA-binding domain of Amos determines the MD neuronal specificity. PMID- 10707973 TI - amos, a proneural gene for Drosophila olfactory sense organs that is regulated by lozenge. AB - In a variety of organisms, early neurogenesis requires the function of basic helix-loop-helix (bHLH) transcription factors. For the Drosophila PNS, such transcription factors are encoded by the proneural genes (atonal and the achaete scute complex, AS-C). We have identified a proneural gene, amos, that has strong similarity with atonal in its bHLH domain. We present evidence that amos is required for olfactory sensilla and is regulated by the prepattern gene lozenge. Between them, amos, atonal, and the AS-C can potentially account for the origin of the entire PNS. PMID- 10707974 TI - Cysteine-rich domain isoforms of the neuregulin-1 gene are required for maintenance of peripheral synapses. AB - Neuregulin-1 (NRG-1) signaling has been implicated in inductive interactions between pre- and postsynaptic partners during synaptogenesis. We used gene targeting to selectively disrupt cysteine-rich domain-(CRD-) containing NRG-1 isoforms. In CRD-NRG-1-/-mice, peripheral projections defasciculated and displayed aberrant branching patterns within their targets. Motor nerve terminals were transiently associated with broad bands of postsynaptic ACh receptor (AChR) clusters. Initially, Schwann cell precursors accompanied peripheral projections, but later, Schwann cells were absent from axons in the periphery. Following initial stages of synapse formation, sensory and motor nerves withdrew and degenerated. Our data demonstrate the essential role of CRD-NRG-1-mediated signaling for coordinating nerve, target, and Schwann cell interactions in the normal maintenance of peripheral synapses, and ultimately in the survival of CRD NRG-1-expressing neurons. PMID- 10707975 TI - Molecular modification of N-cadherin in response to synaptic activity. AB - The relationship between adhesive interactions across the synaptic cleft and synaptic function has remained elusive. At certain CNS synapses, pre- to postsynaptic adhesion is mediated at least in part by neural (N-) cadherin. Here, we demonstrate that upon depolarization of hippocampal neurons in culture by K+ treatment, or application of NMDA or alpha-latrotoxin, synaptic N-cadherin dimerizes and becomes markedly protease resistant. These properties are indices of strong, stable, enhanced cadherin-mediated intercellular adhesion. N-cadherin retained protease resistance for at least 2 hr after recovery, while other surface molecules, including other cadherins, were completely degraded. The acquisition of protease resistance and dimerization of N-cadherin is not dependent on new protein synthesis, nor is it accompanied by internalization of N cadherin. By immunocytochemistry, we found that high K+ selectively induces surface dispersion of N-cadherin, which, after recovery, returns to synaptic puncta. N-cadherin dispersion under K+ treatment parallels the rapid expansion of the presynaptic membrane consequent to the massive vesicle fusion that occurs with this type of depolarization. In contrast, with NMDA application, N-cadherin does not disperse but does acquire enhanced protease resistance and dimerizes. Our data strongly suggest that synaptic adhesion is dynamically and locally controlled, and modulated by synaptic activity. PMID- 10707976 TI - Compromised neural selectivity for song in birds with impaired sensorimotor learning. AB - Anterior forebrain (AF) neurons become selective for song as songbirds learn to produce a copy of a memorized tutor song. We report that development of selectivity is compromised when birds are prevented from matching their output to the tutor song. Finches with denervated vocal organs developed stable song, but it usually did not resemble the tutor song. In those birds, numerous neurons in Area X responded selectively to both tutor and bird's own song (BOS), indicating the importance of both in shaping AF responses. The degree of selectivity for BOS was less, however, than that of normal adults. In contrast, neurons in denervated birds that successfully mimicked tutor song exhibited normal adult selectivity for BOS. Thus, during sensorimotor learning, selectivity for complex stimuli may be influenced by how well motor output matches internal sensory models. PMID- 10707977 TI - GABA synchronizes clock cells within the suprachiasmatic circadian clock. AB - The master clock in the suprachiasmatic nuclei (SCN) is composed of multiple, single-cell circadian clocks. We test the postulate that these individual "clock cells" can be synchronized to each other by the inhibitory transmitter gamma aminobutyric acid (GABA). For these experiments, we monitored the firing rate rhythm of individual clock cells on fixed multielectrode plates in culture and tested the effects of GABA. The results show that the daily variation in responsiveness of the SCN to phase-shifting agents is manifested at the level of individual neurons. Moreover, GABA, acting through A-type receptors, can both phase shift and synchronize clock cells. We propose that GABA is an important synchronizer of SCN neurons in vivo. PMID- 10707978 TI - Rest in Drosophila is a sleep-like state. AB - To facilitate the genetic study of sleep, we documented that rest behavior in Drosophila melanogaster is a sleep-like state. The animals choose a preferred location, become immobile for periods of up to 157 min at a particular time in the circadian day, and are relatively unresponsive to sensory stimuli. Rest is affected by both homeostatic and circadian influences: when rest is prevented, the flies increasingly tend to rest despite stimulation and then exhibit a rest rebound. Drugs acting on a mammalian adenosine receptor alter rest as they do sleep, suggesting conserved neural mechanisms. Finally, normal homeostatic regulation depends on the timeless but not the period central clock gene. Understanding the molecular features of Drosophila rest should shed new light on the mechanisms and function of sleep. PMID- 10707979 TI - The mle(napts) RNA helicase mutation in drosophila results in a splicing catastrophe of the para Na+ channel transcript in a region of RNA editing. AB - The mle(napts) mutation causes temperature-dependent blockade of action potentials resulting from decreased abundance of para-encoded Na+ channels. Although maleless (mle) encodes a double-stranded RNA (dsRNA) helicase, exactly how mle(napts) affects para expression remained uncertain. Here, we show that para transcripts undergo adenosine-to-inosine (A-to-I) RNA editing via a mechanism that apparently requires dsRNA secondary structure formation encompassing the edited exon and the downstream intron. In an mle(napts) background, >80% of para transcripts are aberrant, owing to internal deletions that include the edited exon. We propose that the Mle helicase is required to resolve the dsRNA structure and that failure to do so in an mle(napts) background causes exon skipping because the normal splice donor is occluded. These results explain how mlen(napts) affects Na+ channel expression and provide new insights into the mechanism of RNA editing. PMID- 10707980 TI - Postsynaptic target specificity of neurotrophin-induced presynaptic potentiation. AB - The role of the target cell in neurotrophin-induced modifications of glutamatergic synaptic transmission was examined in cultured hippocampal neurons. Brain-derived neurotrophic factor (BDNF) induced rapid and persistent potentiation of evoked glutamate release when the postsynaptic neuron was glutamatergic, or excitatory (E-->E), but not when it was GABAergic, or inhibitory (E-->1). This target-specific action of BDNF was also found at divergent outputs of a single presynaptic neuron innervating both glutamatergic and GABAergic neurons, suggesting that individual terminals can be independently modified. Surprisingly, BDNF increased the frequency of miniature postsynaptic currents at both E-->E and E-->I, although it had no effect on evoked currents at E-->I. Finally, potentiation by neurotrophin-3 (NT-3) was also target specific. The selective effect at E-->E suggests that retrograde signaling by the postsynaptic target cell endows a localized presynaptic action of neurotrophins. PMID- 10707982 TI - The I-II loop of the Ca2+ channel alpha1 subunit contains an endoplasmic reticulum retention signal antagonized by the beta subunit. AB - The auxiliary beta subunit is essential for functional expression of high voltage activated Ca2+ channels. This effect is partly mediated by a facilitation of the intracellular trafficking of alpha1 subunit toward the plasma membrane. Here, we demonstrate that the I-II loop of the alpha1 subunit contains an endoplasmic reticulum (ER) retention signal that severely restricts the plasma membrane incorporation of alpha1 subunit. Coimmunolabeling reveals that the I-II loop restricts expression of a chimera CD8-I-II protein to the ER. The beta subunit reverses the inhibition imposed by the retention signal. Extensive deletion of this retention signal in full-length alpha1 subunit facilitates the cell surface expression of the channel in the absence of beta subunit. Our data suggest that the beta subunit favors Ca2+ channel plasma membrane expression by inhibiting an expression brake contained in beta-binding alpha1 sequences. PMID- 10707981 TI - Blocking the NGF-TrkA interaction rescues the developmental loss of LTP in the rat visual cortex: role of the cholinergic system. AB - Although nerve growth factor (NGF) is a crucial factor in the activity-dependent development and plasticity of visual cortex, its role in synaptic efficacy changes is largely undefined. We demonstrate that the maintenance phase of long term potentiation (LTP) is blocked by local application of exogenous NGF in rat visual cortex at an early stage of postnatal development. Long-term depression (LTD) and bidirectional plasticity are unaffected. At later postnatal ages, blockade of either endogenous NGF by immunoadhesin (TrkA-IgG) or TrkA receptors by monoclonal antibody rescues LTP. Muscarinic receptor activation/inhibition suggests that LTP dependence on NGF is mediated by the cholinergic system. These results indicate that NGF regulates synaptic strength in well-characterized cortical circuitries. PMID- 10707983 TI - Syntaphilin: a syntaxin-1 clamp that controls SNARE assembly. AB - Syntaxin-1 is a key component of the synaptic vesicle docking/fusion machinery that forms the SNARE complex with VAMP/synaptobrevin and SNAP-25. Identifying proteins that modulate SNARE complex formation is critical for understanding the molecular mechanisms underlying neurotransmitter release and its modulation. We have cloned and characterized a protein called syntaphilin that is selectively expressed in brain. Syntaphilin competes with SNAP-25 for binding to syntaxin-1 and inhibits SNARE complex formation by absorbing free syntaxin-1. Transient overexpression of syntaphilin in cultured hippocampal neurons significantly reduces neurotransmitter release. Furthermore, introduction of syntaphilin into presynaptic superior cervical ganglion neurons in culture inhibits synaptic transmission. These findings suggest that syntaphilin may function as a molecular clamp that controls free syntaxin-1 availability for the assembly of the SNARE complex, and thereby regulates synaptic vesicle exocytosis. PMID- 10707984 TI - Piccolo, a presynaptic zinc finger protein structurally related to bassoon. AB - Piccolo is a novel component of the presynaptic cytoskeletal matrix (PCM) assembled at the active zone of neurotransmitter release. Analysis of its primary structure reveals that Piccolo is a multidomain zinc finger protein structurally related to Bassoon, another PCM protein. Both proteins were found to be shared components of glutamatergic and GABAergic CNS synapses but not of the cholinergic neuromuscular junction. The Piccolo zinc fingers were found to interact with the dual prenylated rab3A and VAMP2/Synaptobrevin II receptor PRA1. We show that PRA1 is a synaptic vesicle-associated protein that is colocalized with Piccolo in nerve terminals of hippocampal primary neurons. These data suggest that Piccolo plays a role in the trafficking of synaptic vesicles (SVs) at the active zone. PMID- 10707985 TI - Light evokes Ca2+ spikes in the axon terminal of a retinal bipolar cell. AB - Bipolar cells in the vertebrate retina have been characterized as nonspiking interneurons. Using patch-clamp recordings from goldfish retinal slices, we find, however, that the morphologically well-defined Mb1 bipolar cell is capable of generating spikes. Surprisingly, in dark-adapted retina, spikes were reliably evoked by light flashes and had a long (1-2 s) refractory period. In light adapted retina, most Mb1 cells did not spike. However, an L-type Ca2+ channel agonist could induce periodic spiking in these cells. Spikes were determined to be Ca2+ action potentials triggered at the axon terminal and were abolished by 2 amino-4-phosphonobutyric acid (APB), an agonist that mimics glutamate. Signaling via spikes in a specific class of bipolar cells may serve to accelerate and amplify small photo-receptor signals, thereby securing the synaptic transmission of dim and rapidly changing visual input. PMID- 10707986 TI - The role of mitochondria in presynaptic calcium handling at a ribbon synapse. AB - Mitochondria are thought to be important in clearing calcium from synaptic terminals. It is unclear, however, whether the principal role of mitochondria in pre-synaptic calcium handling is to take up Ca2+ directly or to fuel Ca2+ removal by other mechanisms. We used patch clamp techniques and fluorescence imaging to examine calcium clearance mechanisms, including mitochondrial uptake, in single synaptic terminals of retinal bipolar neurons. We found that extrusion through the ATP-dependent Ca2+ pump of the plasma membrane is the dominant form of Ca2+ removal in the synaptic terminal. Calcium uptake into mitochondria was sometimes evident with large Ca2+ loads but was consistently observed only when plasma membrane extrusion was inhibited. We conclude that mitochondria act primarily as an energy source in clearance of Ca2+ from bipolar cell synaptic terminals. PMID- 10707988 TI - Autonomic nervous system dysfunction may explain the multisystem features of fibromyalgia. PMID- 10707987 TI - Mice lacking alpha-synuclein display functional deficits in the nigrostriatal dopamine system. AB - alpha-Synuclein (alpha-Syn) is a 14 kDa protein of unknown function that has been implicated in the pathophysiology of Parkinson's disease (PD). Here, we show that alpha-Syn-/- mice are viable and fertile, exhibit intact brain architecture, and possess a normal complement of dopaminergic cell bodies, fibers, and synapses. Nigrostriatal terminals of alpha-Syn-/- mice display a standard pattern of dopamine (DA) discharge and reuptake in response to simple electrical stimulation. However, they exhibit an increased release with paired stimuli that can be mimicked by elevated Ca2+. Concurrent with the altered DA release, alpha Syn-/- mice display a reduction in striatal DA and an attenuation of DA-dependent locomotor response to amphetamine. These findings support the hypothesis that alpha-Syn is an essential presynaptic, activity-dependent negative regulator of DA neurotransmission. PMID- 10707989 TI - Trauma and fibromyalgia: is there an association and what does it mean? AB - OBJECTIVES: The primary objective is to review current research with respect to the role of trauma in fibromyalgia (FM). A secondary objective is to hypothesize which steps need to be taken, first to determine whether such an association truly exists, and second to clarify what such an association might mean. METHODS: An extensive literature review was undertaken, including Medline from 1979 to the present. RESULTS: The strongest evidence supporting an association between trauma and FM is a recently published Israeli study in which adults with neck injuries had greater than a 10-fold increased risk of developing FM within 1 year of their injury, compared with adults with lower extremity fractures (P= .001). Several other studies provide a hypothetical construct for such an association. These include studies on (1) postinjury sleep abnormalities; (2) local injury sites as a source of chronic distant regional pain; and (3) the concept of neuroplasticity. There are, however, several primary arguments against such an association: (1) FM may not be a distinct clinical entity; (2) FM may be a psychological, rather than physical, disease; (3) the evidence supporting any association is limited and not definitive; (4) the Israeli study, itself, has some methodological limitations; and (5) other factors may be more important than the injurious event in determining chronic symptoms after an acute injury. CONCLUSIONS: Although there is some evidence supporting an association between trauma and FM, the evidence is not definitive. Further prospective studies are needed to confirm this association and to identify whether trauma has a causal role. PMID- 10707990 TI - Autonomic dysfunction in patients with fibromyalgia: application of power spectral analysis of heart rate variability. AB - OBJECTIVES: To assess the interaction between the sympathetic and parasympathetic systems in patients with fibromyalgia syndrome (FM), using power spectrum analysis (PSA) of heart rate variability (HRV). In addition, we explored the association between HRV, measures of tenderness, FM symptoms, physical function, psychological well being and quality of life. METHODS: We studied 22 women with FM and 22 age-matched healthy women. Twenty-minute electrocardiogram recordings were obtained in a supine position during complete rest. Spectral analysis of R-R intervals was done by the fast-Fourier transform algorithm. RESULTS: Heart rate was significantly higher in FM patients compared with controls (P < .006). FM patients had significantly lower HRV compared with controls (P= .001), and higher low-frequency (LF) and lower high-frequency (HF) components of PSA than controls (P < .001). Quality of life, physical function, anxiety, depression, and perceived stress were moderately to highly correlated with LF, HF (in normalized units), and LF/HF. No association was observed between HRV parameters and measures of tenderness and FM symptoms. CONCLUSIONS: The basal autonomic state of patients with FM is characterized by increased sympathetic and decreased parasympathetic tones. Autonomic dysregulation may have implications regarding the symptomatology, physical and psychological aspects of health status. PMID- 10707991 TI - Management of corticosteroid-induced osteoporosis. AB - OBJECTIVES: To educate scientists and health care providers about the effects of corticosteroids on bone, and advise clinicians of the appropriate treatments for patients receiving corticosteroids. METHODS: This review summarizes the pathophysiology of corticosteroid-induced osteoporosis, describes the assessment methods used to evaluate this condition, examines the results of clinical trials of drugs, and explores a practical approach to the management of corticosteroid induced osteoporosis based on data collected from published articles. RESULTS: Despite our lack of understanding about the biological mechanisms leading to corticosteroid-induced bone loss, effective therapy has been developed. Bisphosphonate therapy is beneficial in both the prevention and treatment of corticosteroid-induced osteoporosis. The data for the bisphosphonates are more compelling than for any other agent. For patients who have been treated but continue to lose bone, hormone replacement therapy, calcitonin, fluoride, or anabolic hormones should be considered. Calcium should be used only as an adjunctive therapy in the treatment or prevention of corticosteroid-induced bone loss and should be administered in combination with other agents. CONCLUSIONS: Bisphosphonates have shown significant treatment benefit and are the agents of choice for both the treatment and prevention of corticosteroid-induced osteoporosis. PMID- 10707992 TI - Fibronectin fragments and their role in inflammatory arthritis. AB - OBJECTIVES: To identify potential immunopathogenic links between fibronectin (Fn) fragmentation and the inflammatory response in chronic joint disease. METHODS: Scientific papers involving studies of Fn fragments and inflammatory processes important in the pathogenesis of arthritis, including chondrolysis, synoviocyte growth and adhesion, polymorphonuclear leukocyte (PMN) and monocyte function, proteolysis, and immune complex activation were reviewed. In addition, reports identifying Fn fragments in synovial fluid (SF) were assessed. RESULTS: A series of Fn fragments have been identified in arthritic SF by several investigators. Fn and fragments ranging from 30 to 200 kd are present in elevated concentrations in inflammatory SF. SF Fn fragments display reduced affinity for fibrin and collagen. The 29- and 50-kd amino terminal fragments mediate release of proteoglycan from articular cartilage by RGD-independent mechanisms. Fn fragments can induce fibroblast gene expression of metalloproteinases or can act as proteinases themselves. A 90-kd plasmin generated fragment possesses homology with streptokinase. Fragments mediate PMN chemotaxis and enhance proliferation of CD4+ lymphocytes as well as binding to the C1q component of complement and influencing the behavior of immune complexes. CONCLUSIONS: Fn fragments can be functionally and biochemically characterized in diseased SF. Modification of fragment formation and inhibition of fragment function may have potential therapeutic value in the interruption of chronic synovial inflammation. PMID- 10707993 TI - High prevalence of hepatitis C antibody and RNA in patients with Sjogren's syndrome. PMID- 10707994 TI - A review of autologous hematopoietic cell transplantation. PMID- 10707995 TI - Complexity of effector mechanisms in cyclosporine-induced syngeneic graft-versus host disease. AB - Administration of the immunosuppressive drug cyclosporine after syngeneic or autologous bone marrow transplantation elicits a T-lymphocyte-dependent autoimmune syndrome similar to graft-versus-host disease (GVHD). The onset of this autoaggression syndrome, termed syngeneic GVHD, is associated with the development of a highly restricted repertoire of CD8+ autoreactive T cells that recognize a peptide from the invariant chain, termed CLIP, presented by major histocompatibility complex (MHC) class II molecules. Clonal analysis reveals 2 distinct subsets of autoreactive T cells defined by their activation requirement for either the N-terminal or the C-terminal flanking regions of CLIP and by their cytokine profile. The studies here reveal that the autoreactive T-cell clones requiring the N-terminal flanking region of CLIP produce type 1 cytokines (interferon [IFN]-gamma, interleukin [IL]-2, and tumor necrosis factor-alpha). In contrast, the autoreactive T-cell clones that require the C-terminal flanking region of CLIP produce type 2 cytokines (IL-4, IL-10, transforming growth factor beta). As assessed in a local graft-versus-host reaction assay, the N-terminal flanking-restricted clones mediate changes consistent with acute GVHD, whereas the clones responsive to the C-terminal flanking region do not. Moreover, the autoreactive T-cell clones restricted by the C-terminal flanking region of CLIP ameliorate the pathogenic potential of the cells responsive to the N-terminal flanking region of CLIP. The mechanism accounting for this regulatory affect appears to be the downregulation of mRNA message for type 1 cytokines (IFN-gamma and IL-2). The C-terminal-restricted autoreactive T-cell clones, however, could manifest disease with dermal changes similar to those seen in chronic syngeneic GVHD, provided that IFN-gamma was present. Consistent with these observations was the demonstration that type 1 cytokines are preferentially detected during the acute phase of syngeneic GVHD, whereas type 2 cytokines dominate during the chronic phase. The results suggest that acute and chronic syngeneic GVHD is mediated by distinct autoreactive T cells, which are separated by their fine specificity for the CLIP-MHC class II complex and by their cytokine profiles. PMID- 10707996 TI - A phase I trial of recombinant human thrombopoietin in patients with delayed platelet recovery after hematopoietic stem cell transplantation. AB - Delayed platelet recovery is a significant complication after both autologous and allogeneic hematopoietic stem cell transplantation (HSCT). A multicenter, phase I dose-escalation study of recombinant human thrombopoietin (rhTPO) was conducted to assess its safety and to obtain preliminary data on its efficacy in patients with persistent severe thrombocytopenia (<20,000/microL) >35 days after HSCT. Thirty-eight patients, 37 of whom were evaluable, were enrolled in the study from April 1996 through January 1997. rhTPO was administered at doses of 0.6, 1.2, and 2.4 microg/kg as a single dose (group A) or in multiple doses every 3 days for a total of 5 doses (group B). No significant adverse effects were observed. Ten patients had recovery of platelet counts during the 28-day study period; 3 of these 10 had an increase in marrow megakaryocyte content 7 days after completing treatment with rhTPO. When all baseline marrows were compared with samples after rhTPO treatment, there was no difference in marrow megakaryocyte content (P = 0.49). This study design could not answer the question of whether the recoveries of platelet counts observed in some patients were spontaneous or influenced by rhTPO treatment; nonetheless, the authors found no correlation between the dose of rhTPO and the recovery of platelet counts. Increases in serum TPO levels were dose-dependent and remained significantly elevated for up to 72 hours after treatment. To evaluate response, further studies of treatment strategies with rhTPO in patients with delayed platelet recovery are required. PMID- 10707997 TI - Aerosolized pentamidine as pneumocystis prophylaxis after bone marrow transplantation is inferior to other regimens and is associated with decreased survival and an increased risk of other infections. AB - Pneumocystis carinii pneumonia (PCP) is a life-threatening but preventable infection that may occur after bone marrow transplantation (BMT). Although various prophylactic regimens have been used in this setting to prevent active infection, their efficacy, toxicity profile, and impact on outcomes are poorly described in this patient group. We undertook a retrospective cohort study in which we reviewed the records of 451 adult patients who underwent BMT for hematologic malignancies, aplastic anemia, or myelodysplasia over a 7-year period at the Brigham and Women's Hospital. Post-BMT PCP prophylaxis consisted of aerosolized pentamidine (AP) 150 mg every 2 weeks or 300 mg per month, trimethoprim/sulfamethoxazole (TMP/SMX) 160/800 mg orally b.i.d. 3 times per week, or dapsone 100 mg orally each day. Prophylaxis was continued for 1 year post-BMT in all patients when clinically feasible. One hundred twenty-one patients were unevaluable because of death or relapse <60 days after BMT (n = 89), loss to follow-up upon hospital discharge (n = 20), or other reasons (n = 12). Three eligible patients did not receive any prophylaxis and were not further evaluated. Of the 327 patients analyzed, 133 underwent autologous BMT, 4 syngeneic BMT, 159 related allogeneic BMT, and 31 unrelated allogeneic BMT. Graft versus-host disease prophylaxis in the 190 patients receiving allogeneic BMT consisted of T-cell depletion with anti-CD5 and complement in 58 patients and cyclosporine/methotrexate or FK506 with or without steroids in 132 patients. Eight of 327 (2.4%) documented PCP cases were identified, 0 of 105 in patients receiving only TMP/SMX. Four cases occurred in patients receiving only AP (4/44, 9.1%; odds ratio [OR] relative to TMP/SMX 23.4, 95% confidence interval [CI] 1.2, 445.2); 1 in patients receiving only dapsone (1/31, 3.2%; OR not significant); 2 in patients receiving more than 1 prophylactic regimen (2/147 1.4%; OR not significant); and 1 >1 year post-BMT in a patient who was off PCP prophylaxis. Although the patients receiving only AP had a significantly lower probability of treatment-related toxicity than those receiving TMP/SMX (OR 0.19 [95% CI 0.04, 0.851), the probability of their acquiring other serious non-PCP infections was increased (OR 2.2 [95% CI 1.0, 4.6]), and the probability of their dying by 1 year post-BMT was significantly higher (OR 5.2 [95% CI 2.4, 26.6]), even when adjusted for variables such as type of BMT (autologous versus allogeneic; high versus low risk) and sex. Although AP is associated with fewer toxicities, the data show that it is inferior to TMP/SMX in preventing PCP in the post-BMT setting and is associated with an increased risk of other infections and a higher mortality at 1 year after BMT. PMID- 10707998 TI - Varicella zoster virus infections following allogeneic bone marrow transplantation: frequency, risk factors, and clinical outcome. AB - Reactivation of varicella zoster virus (VZV) is a common event in patients undergoing allogeneic bone marrow transplantation (BMT) and may lead to life threatening complications. We retrospectively analyzed the incidence, clinical outcome, and risk factors for VZV infections occurring within the first 5 years of transplantation in 100 consecutive adults undergoing allogeneic BMT between 1992 and 1997. Forty-one patients (41%) developed VZV reactivation a median of 227 days (range 45-346 days) post-transplantation. Twelve percent of VZV reactivation occurred in the first 100 days and 88% within the first 24 months. Among those who survived for 2 or more years after transplantation (n = 47), 59% developed VZV infection. Forty percent of patients with VZV reactivation required admission with a mean hospital stay of 7.2 days. Two patients developed encephalitis, and 1 died despite antiviral therapy. The most frequent complications were post-herpetic neuralgia and peripheral neuropathy (68%). Thoracic dermatomal zoster represented 41% of the infections; disseminated cutaneous involvement was observed in 17% of patients. No clinical or epidemiologic risk factors were associated with recurrence. Administration of ganciclovir for prevention of cytomegalovirus infection delayed the onset of VZV infection beyond 4 months (P = .06). In a further subset analysis, patients with a limited chronic graft-versus-host disease (GVHD) had a lower estimated incidence of VZV reactivation compared with those with extensive chronic GVHD (P = .11). We conclude that complications from reactivation of VZV infection are common and associated with considerable morbidity and mortality in patients undergoing allogeneic BMT. PMID- 10707999 TI - Autologous stem cell transplantation for acute myeloid leukemia in first remission. AB - We studied the feasibility, toxicity, and efficacy of a 2-step approach to autologous stem cell transplantation for patients with acute myeloid leukemia in first remission. Step 1 consisted of consolidation chemotherapy including cytarabine 2000 mg/m2 twice daily for 4 days concurrent with etoposide 40 mg/kg by continuous infusion over 4 days. During the recovery from this chemotherapy, peripheral blood stem cells were collected under granulocyte colony-stimulating factor stimulation. Step 2, autologous stem cell transplantation, involved the preparative regimen of busulfan 16 mg/kg followed by etoposide 60 mg/kg and reinfusion of unpurged peripheral blood stem cells. A total of 128 patients were treated. During step 1, there was 1 treatment-related death. A median CD34+ cell dose of 14 (x10(6)/kg) was collected in 3 aphereses. Ten patients suffered relapse before transplantation, and 117 patients (91%) proceeded to transplantation. During step 2, there were 2 treatment-related deaths, and 35 patients subsequently suffered relapse. With median follow-up of 30 months, 5 year disease-free survival for all patients entered in the study is projected to be 55%. By cytogenetic risk group, 5-year disease-free survival is 73% for favorable-risk patients, 51% for intermediate-risk patients, and 0% for poor-risk patients. We conclude that this 2-step approach to autologous transplantation produces excellent stem cell yields and allows a high percentage of patients to receive the intended therapy. Preliminary efficacy analysis is very encouraging, with outcomes that appear superior to those of conventional chemotherapy. PMID- 10708000 TI - Four-cycle high-dose therapy with hematopoietic support for metastatic breast cancer: no improvement in outcomes compared with single-course high-dose therapy. AB - Multiple-cycle high-dose therapy with autologous hematopoietic progenitor cell (AHPC) support has been used to deliver dose-intensive therapy. We have used this approach as well as single-cycle high-dose therapy in treating patients with metastatic breast cancer. We present the outcomes of multiple-cycle high-dose therapies and compare them with those resulting from single-course high-dose therapies performed at a single institution. Fifty-five patients received 4 cycles of intensive chemotherapy with AHPC support. Three multicycle regimens were sequentially applied. Twenty patients were enrolled to receive 4 cycles of high-dose mitoxantrone, thiotepa, and cyclophosphamide. Nineteen subsequent patients received this regimen modified by the incorporation of paclitaxel. Sixteen patients received 2 cycles of high-dose melphalan, thiotepa, and paclitaxel and 2 cycles of mitoxantrone, thiotepa, and paclitaxel. The results of all 3 multiple-cycle therapies are compared with those of 55 contemporaneous patients with metastatic breast cancer who received a single course of high-dose cyclophosphamide and thiotepa or cyclophosphamide, cisplatin, and BCNU (carmustine) with hematopoietic cell rescue. Multiple-cycle therapy was associated with more infectious complications, increased transfusion requirements, and increased hospital admissions. However, there were no significant differences in outcomes between the groups. For 55 patients who received multiple-cycle therapy, the actuarial 3-year overall survival rate was 36% (95% confidence interval [CI] 23%-49%); freedom from progression and event free survival were both 15% (CI 5%-25%). The median time to disease progression and median survival were 1.0 and 1.6 years, respectively. For the 55 patients who underwent a single course of high-dose therapy, the 3-year overall survival was also 36% (CI 18%-54%), whereas freedom from progression and event-free survival were both 19% (CI 7%-31%). The median time to progression and median survival were 0.8 and 2.2 years, respectively. Within the constraints of this patient population, the outcomes of 4 cycles of high-dose therapy with AHPC support were not superior to those resulting from single courses of high-dose therapy in the treatment of patients with metastatic breast cancer. PMID- 10708001 TI - Intralesional autotherapy of cutaneous leishmaniasis with buffy coat cells: cytological findings. AB - The skin lesions of five patient volunteers with dry-type cutaneous leishmaniasis were treated by intralesional injection of auto-leukocytes prepared from buffy coat of the patient's own blood. Giemsa stained, air-dried cytological smear preparations were prepared from scrapings taken from the margins of the lesions. The cellular interaction between the organism and the inflammatory response of the host was studied. All lesions showed clinical evidence of regression. The cytological findings suggested progressive degradation of the Leishman donovan (LD) bodies within the parasitophorous vacuoles of the activated macrophages. The parasiticidal effect appeared to be induced by synergistic action of the injected neutrophils and lymphocytes. Due to lack of placebo controls in this study the possibility that, healing might not be related to therapy can not be excluded. This study illustrates the potential for intralesional autotherapy with buffy coat in dry-type cutaneous leishmaniasis. PMID- 10708002 TI - Immunodiagnosis of human eosinophilic meningitis using an antigen of Angiostrongylus cantonensis L5 with molecular weight 204 kD. AB - An antigen from Angiostrongylus cantonensis fifth-stage larvae was purified by immuno-affinity chromatography with a specific monoclonal antibody. The purified antigen showed only a single band with a molecular weight of 204 kD in SDS-PAGE, and no cross-reactivity to antibodies induced by several other species of helminths were observed in ELISA. When the purified antigen was used to examine serum and cerebrospinal fluid (CSF) specimens by ELISA, the antibody levels in patients with eosinophilic meningitis or meningoencephalitis (EME) were significantly higher than those of control subjects. The antibody levels in serum were slightly higher than those in CSF, and the levels in serum were positively correlated with the levels in CSF. The reliability in detection of antibodies in serum was slightly higher than that in the detection of antibodies in CSF specimens. The purification of a specific A. cantonensis antigen and its subsequent use in the development of an ELISA for detection of A. cantonensis specific antibodies in serum specimens constitute an important step towards improvement in the accuracy of diagnosis for A. cantonensis infections. PMID- 10708003 TI - Acute adenolymphangitis due to bancroftian filariasis in Rufiji district, south east Tanzania. AB - A longitudinal prospective surveillance for acute adenolymphagitis (ADL) was carried out in three villages in Rufiji district. A sample population of 3000 individuals aged 10 years and above was monitored fortnightly for a period of 12 months. The annual incidence of ADL was found to be 33 per 1000 population and was significantly higher in males than females (52.7/1000 and 18.7/1000 respectively). ADL episodes were more frequent in the age group of 40 years and above. Individuals with chronic manifestations seemed to be more vulnerable to ADL attacks with 62.2% of the total episodes occurring in this group. Furthermore, individuals with lymphoedema experienced more frequent acute episodes compared to those with hydrocele and 'normal exposed'. ADL episodes ranged from one to five per annum and the majority of the affected (60.4%) experienced a single episode. The average duration of an ADL episode was 8.6 days and in 72.5% of the episodes the affected individuals were incapacitated and unable to do their normal activities for an average duration of 3.7 days. The physical incapacitation associated with ADL episodes emphasizes the significance of lymphatic filariasis as a major public health problem of substantial socio economic consequences. PMID- 10708004 TI - Perinatal outcome in pregnancies booked for antenatal care but delivered outside health facilities in Calabar, Nigeria. AB - Pregnancies that were booked for antenatal care but delivered outside the health facilities were studied. The aim was to determine the perinatal outcome of these pregnancies, and also to compare the outcome with that of pregnancies that were booked and delivered in the University of Calabar Teaching Hospital (UCTH). Birth asphyxia was the commonest perinatal morbidity in both the study (14.3%) and control (4.8%) groups and was significantly higher in the study group than in the control (P < 0.01-P < 0.05). Incidence of neonatal infection tetanus and birth trauma was also significantly higher in the study than in the control groups (P < 0.01-P < 0.05). The incidence of prematurity, neonatal jaundice and congenital abnormality did not show any significant difference in the two groups (P > 0.05). The risk of perinatal death was three times higher in the study group than in the control. Proper public enlightenment campaigns and the establishment of a national health insurance scheme which may strengthen the use of orthodox health facilities for delivery, may improve the poor perinatal outcome in our community. PMID- 10708005 TI - High levels of C-reactive protein in the peripheral blood during visceral leishmaniasis predict subsequent development of post kala-azar dermal leishmaniasis. AB - Post kala-azar dermal leishmaniasis (PKDL) is a known sequel to visceral leishmaniasis in India and East Africa, and in Sudan about 50% of the kala-azar patients develop PKDL. In this study we followed kala-azar patients from diagnosis and up to 2 years after initiation of treatment. During the first 6 months some developed PKDL (group 1), while some did not develop PKDL (group 2). We measured the plasma levels of C-reactive protein (CRP) at diagnosis of kala azar (day 0), during treatment (day 15), after treatment (day 30) and later during the follow up period. At day 0, plasma CRP levels were higher in patients who later developed PKDL (group 1) than in patients who did not develop PKDL subsequently (group 2) (P = 0.008). At days 15 and 30, the CRP levels were comparable in the two groups, and lower than at day 0. We have previously shown that high plasma levels of IL 10 and in keratinocytes during visceral leishmaniasis predict subsequent development of PKDL. The method however requires expensive equipment and reagents. The results of the present study indicate that kala-azar patients, who have a high risk of developing PKDL after treatment can be identified by measuring plasma CRP. PMID- 10708006 TI - Immunogenicity and effectiveness of post-exposure rabies prophylaxis with a new chromatographically purified Vero-cell rabies vaccine (CPRV): a two-stage randomised clinical trial in the Philippines. AB - Recent improvements in chromatographic purification procedures have made it possible to develop a new chromatographically purified rabies vaccine (CPRV) by further purifying the current rabies vaccine prepared from Vero-cell culture (PVRV) (Verorab; Pasteur Merieux Connaught). The immunogenicity and effectiveness of post-exposure rabies prophylaxis with this new vaccine were evaluated in a two stage clinical trial conducted in the Philippines. In both study stages. post exposure treatment consisted of five injections of vaccine [(D)ays 0, 3, 7, 14, 28], together with a dose of rabies immunoglobulin (RIG) of equine or human origin on D0. In stage 1, 231 subjects with low-risk rabies exposure (WHO category I or II), and who had a negative ERIG skin test, were treated with either CPRV (n = 114) or PVRV (n = 117). By D14, all subjects in each group had achieved rabies antibody titres over ten times that recommended by the WHO as indicating seroconversion (> or = 0.5 IU/ml). The kinetics of the immune response to vaccination were very similar in the two groups, and at D28, the immunogenicity of CPRV was equivalent to that of PVRV (one-sided equivalence test). Following these positive results, 132 subjects with severe rabies exposure were included in the second stage of this trial. All were scheduled to receive four vaccine doses with CPRV. After D14, only those 57 patients with confirmed rabies exposure (animal with positive FA test) and seven patients for whom rabies exposure could not be excluded (animal lost or not tested) completed the treatment and were followed for one year to assess survival. After 1 year, 62 patients treated for confirmed or possible severe rabies exposure had been examined and were still alive. Two patients contacted by letter and telephone confirmed good health 7 and 16 months after exposure. No severe local or systemic reactions were reported in either stage of the study, and no treatment-related serious adverse event occurred. This two-stage clinical trial attests to the safety and satisfactory immunogenicity of CPRV in post-exposure rabies treatment, and confirms the effectiveness of a new rabies vaccine in patients with severe confirmed exposure. PMID- 10708007 TI - Clinical management and prevention of sexually transmitted diseases: a review focusing on women. AB - This review highlights some of the difficulties inherent in controlling sexually transmitted diseases (STDs) in developing countries--especially amongst women. Considerable efforts have been made to improve the syndromic approach to STD management but the poor performance of the algorithm for managing vaginal discharge limits the effectiveness of this strategy. The facilitating role of the human immunodeficiency virus (HIV) has been the main impetus to STD control rather than reduction of morbidity in women, especially pregnant women and their children. There are no easy solutions--but action on several fronts, with more attention to core groups, men and adolescents is indicated. PMID- 10708008 TI - RAPD in the analysis of isolates of Entamoeba histolytica. AB - Genetic variability in Entamoeba histolytica was analyzed by random amplified polymorphic DNA (RAPD) using ten arbitrary primers. Due to intrinsic characteristics of the RAPD technique only axenic samples were analyzed. since the presence of any microorganism in the cultures interfered in the DNA profile by generating RAPDs not pertaining to E. histolytica. The RAPD profiles of E. histolytica samples isolated from patients with different clinical manifestations from different regions of the Americas shared about 70% of the bands produced. These profiles were compared to those obtained for E. moshkorskii, and E. invadens. The combined data for the ten primers were used in the phenetic analysis of all the isolates studied by using the Dice similarity coefficient as the genetic distance measure between the samples. Three distinct groups could be separated by phenon line: one including E. moshkovskii samples, which shared > 90% of the RAPDs produced by the different primers; one consisting solely of E. invadens; and a third comprising samples of E. histolytica, which showed considerable intraspecific variability. PMID- 10708009 TI - Paleoparasitology of Chagas disease revaled by infected tissues from Chilean mummies. AB - Mummified tissues were sampled from bodies stored at the Museo Arqueologico de San Pedro de Atacama, northern Chile, dated from 2000 years BP-1400 AD, and Trypanosoma cruzi DNA was recovered using polymerase chain reaction (PCR) methodology. Amplification of the conserved region of the minicircle molecule of T. cruzi was achieved in four of the six samples tested. Amplified products corresponding to genetic fragments of the parasite were tested by hybridization experiments with positive results for T. cruzi specific molecular probe. The origin and dispersion of T. cruzi human infection is discussed as well as the molecular paleoparasitological approach, and what it may represent in an evolutionary perspective. PMID- 10708010 TI - A comparison of Bulinus africanus group species (Planorbidae; Gastropoda) by use of the internal transcribed spacer 1 region combined by morphological and anatomical characters. AB - The morphological and anatomical characters applied to determine species identity in the Bulinus africanus group species are insufficient to unambiguously discriminate between arbitrary species of given populations. In order to solve this problem, four snail populations from Kenya have been investigated morphologically and anatomically, and the species status compared with the result of molecular methods. We have amplified the entire ITS region and found that the investigated populations showed intra-specific genetic polymorphism, thereby giving the taxa an identity which were indistinct when the region was cut with restriction enzymes. Instead, an amplification of the sub-region ITS 1 revealed an unambiguous identification. because the amplification revealed only one single fragment. We also found that the observed heterogeneity of the entire ITS region could be confined to the sub-region ITS 2. Furthermore, the micro-sculpture of the shell and penis to preputium proportion, which are normally applied as morphological characters, might be considered as inadequate, because of the lack of a significant difference in those characters between the two well established species, namely B. africanus and B. nasutus in the Kisumu area. Instead, these two taxa were determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) to be one single species with a highly variable morphology. This result was further confirmed by random amplified polymorphic DNA (RAPD) data and the mitochondrial cytochome oxidase subunit I (COI). PMID- 10708011 TI - Study of the dog population and the rabies control activities in the Mirigama area of Sri Lanka. AB - The national health authorities of Sri Lanka have adopted a combined strategy of rabies vaccination and stray dog removal to control endemic dog rabies. Despite the control efforts, an increase of animal and human rabies cases has occurred since 1994. As a consequence, a project to evaluate the national rabies control program has been started and a study focussing on the dog population and rabies control activities in a limited area of Mirigama was conducted. Information on canine abundance and the accessibility of dogs for rabies vaccination was obtained by a household survey, vaccination of dogs against rabies at several vaccination points, collar-marking, and transect line recapture. The number of unvaccinated dogs was estimated by using Bayesian methodology. The estimated number of dogs per square kilometre was 87 (95% credibility interval: 80, 93) for owned dogs and 108 (100, 116) for owned and ownerless dogs. Coverage after the immunisation campaign was 57.6% (53.3, 61.9%) if vaccination at the vaccination points was considered and 66% (60.4, 72.0%) if recently provided vaccination by private veterinarians was also taken into account. The proportion of households with at least one dog vaccinated varied between 59.1 and 94.2% within the catchment area of the different vaccination points. Unvaccinated dogs were puppies (12%), ownerless dogs (57%), and owned dogs, which were not presented for vaccination (31%). In order to improve the rabies immunisation coverage among dogs and to achieve complete elimination of rabies it was recommended that the 95% catchment area of each vaccination point be assessed, the distribution of vaccination points in the vaccination area be redefined if necessary, a system for the vaccination of dogs missing the vaccination campaign for dog owner specific reasons be established, and an inexpensive marking system be used for vaccinated dogs. PMID- 10708012 TI - Feeding patterns of three sympatric tsetse species (Glossina spp.) (Diptera: Glossinidae) in the preforest zone of Cote d'Ivoire. AB - The feeding patterns of Glossina longipalpis Wiedemann 1830, G. medicorum Austen 1911, G. palpalis gambiensis Vanderplank 1949 and G. p. palpalis Robineau Desvoidy 1830 are described from natural habitats in central Cote d'Ivoire where these tsetse species occurred sympatrically. Blood-meal identification of tsetse flies revealed that in natural habitats wild ruminants were by far the most frequent source of food for each Glossina species, but there were significant differences between the nutritional spectra of single fly species. G. p. gambiensis fed significantly less often on bushbuck and significantly more often on monitor lizard (Varamus niloticus) than both, G. longipalpis and G. medicorum. In G. p. gambiensis the blood of wild ruminant species was significantly more often found than in G. p. palpalis, whereas the latter fed significantly more often on domestic animals. Peridomestic populations of G. longipalpis and G. p. palpalis fed mostly on domestic pig and therefore had significantly reduced host spectra in comparison to natural populations. The significant differences in feeding patterns among the investigated species, subspecies and populations seem not to depend on species specific feeding preferences. Rather, they can be attributed to microhabitat specialization of the various tsetse groups and hence mainly to the different availability of hosts. This implies that environmental factors should be taken more into account when analysing and comparing the feeding patterns of tsetse. PMID- 10708013 TI - Why do we need standard genetic nomenclature for parasite genes and gene products? PMID- 10708014 TI - Patterns of reinfection following praziquantel treatment of urinary schistosomiasis at a period of low transmission. PMID- 10708016 TI - 5th Oxford Conference on Biomedicine in Developing Countries organised by the Oxford International Biomedical Centre (OIBC): 12-15 April 1999. PMID- 10708017 TI - The achromatic mechanism and mechanisms tuned to chromaticity and luminance in visual search. AB - The purpose of the study was to determine whether visual search can be mediated by an achromatic, or luminance, mechanism in which signals are independent of the chromaticity of the stimuli. Experiments were designed to determine whether variability in the chromaticity of distractor stimuli made it more difficult to search for a target that differed from the distractor stimuli in luminance. Variability in the chromaticity of the distractors had little or no effect on search times when the target stimulus was white. Variability in the chromaticity of the distractors increased search times when the target was a reddish or bluish chromaticity. Results obtained with white targets suggest that these searches are mediated by an achromatic mechanism in which the signals are independent of the chromaticity of the stimuli. Results obtained with reddish and bluish targets suggest that searches for those targets may be mediated by mechanisms tuned to both chromaticity and luminance. Further experiments in which observers searched for targets that differed from distractors in both chromaticity and luminance provided additional support for the second conclusion. PMID- 10708015 TI - Treatment with ivermectin reduces the high prevalence of scabies in a village in Papua New Guinea. PMID- 10708018 TI - Imaging properties of scanning holographic microscopy. AB - Scanning heterodyne holography is an alternative way of capturing three dimensional information on a scattering or fluorescent object. We analyze the properties of the images obtained by this novel imaging process. We describe the possibility of varying the coherence of the system from a process linear in amplitude to a process linear in intensity by changing the detection mode. We illustrate numerically the properties of the three-dimensional point-spread function of the system and compare it with that of a conventional imaging system with equal numerical aperture. We describe how it is possible, by an appropriate choice of the reconstruction algorithm, to obtain an ideal transfer function equal to unity up to the cutoff frequency, even in the presence of aberrations. Some practical implementation issues are also discussed. PMID- 10708019 TI - Computationally efficient and statistically robust image reconstruction in three dimensional diffraction tomography. AB - Diffraction tomography (DT) is an inversion scheme used to reconstruct the spatially variant refractive-index distribution of a scattering object. We developed computationally efficient algorithms for image reconstruction in three dimensional (3D) DT. A unique and important aspect of these algorithms is that they involve only a series of two-dimensional reconstructions and thus greatly reduce the prohibitively large computational load required by conventional 3D reconstruction algorithms. We also investigated the noise characteristics of these algorithms and developed strategies that exploit the statistically complementary information inherent in the measured data to achieve a bias-free reduction of the reconstructed image variance. We performed numerical studies that corroborate our theoretical assertions. PMID- 10708020 TI - Network approaches to two-dimensional phase unwrapping: intractability and two new algorithms. AB - Two-dimensional (2-D) phase unwrapping, that is, deducing unambiguous phase values from a 2-D array of values known only modulo 2pi, is a key step in interpreting data acquired with synthetic aperture radar interferometry. Noting the recent network formulation of the phase unwrapping problem, we apply here some well-established ideas of network theory to formalize the problem, analyze its complexity, and derive algorithms for its solution. It has been suggested that the objective of phase unwrapping should be to minimize the total number of places where unwrapped and wrapped phase gradients differ. Here we use network constructions to show that this so-called minimum L0-norm problem is NP-hard, or one that complexity theory suggests is impossible for efficient algorithms to solve exactly. Therefore we must instead find approximate solutions; we present two new algorithms for doing so. The first uses the network ideas of shortest paths and spanning trees to improve on the Goldstein et al. residue-cut algorithm [Radio Sci. 23, 713 (1988)]. Our improved algorithm is very fast, provides complete coverage, and allows user-defined weights. With our second algorithm, we extend the ideas of linear network flow problems to the nonlinear L0 case. This algorithm yields excellent approximations to the minimum L0 norm. Using interferometric data, we demonstrate that our algorithms are highly competitive with other existing algorithms in speed and accuracy, outperforming them in the cases presented here. PMID- 10708021 TI - Rotational-diversity phase estimation from differential-interference-contrast microscopy images. AB - An iterative phase-estimation method for the calculation of a specimen's phase function or optical-path-length (OPL) distribution from differential-interference contrast (DIC) microscopy images is presented. The method minimizes the least squares discrepancy measure by use of the conjugate-gradient technique to estimate the phase function from multiple DIC images acquired at different specimen rotations. The estimate is regularized with a quadratic smoothness penalty. Results from testing the method with simulations and measured DIC images show improvement in the estimated phase when at least two rotationally diverse DIC images instead of a single DIC image are used for the estimation. The OPL of a cell that is estimated from two DIC images was found to be much more reliable than the OPL computed from single DIC images (which had a coefficient of variation equal to 15.8%). PMID- 10708022 TI - Background estimation in nonlinear image restoration. AB - One of the essential ways in which nonlinear image restoration algorithms differ from linear, convolution-type image restoration filters is their capability to restrict the restoration result to nonnegative intensities. The iterative constrained Tikhonov-Miller (ICTM) algorithm, for example, incorporates the nonnegativity constraint by clipping all negative values to zero after each iteration. This constraint will be effective only when the restored intensities have near-zero values. Therefore the background estimation will have an influence on the effectiveness of the nonnegativity constraint of these algorithms. We investigated quantitatively the dependency of the performance of the ICTM, Carrington, and Richardson-Lucy algorithms on the estimation of the background and compared it with the performance of the linear Tikhonov-Miller restoration filter. We found that the performance depends critically on the background estimation: An underestimation of the background will make the nonnegativity constraint ineffective, which results in a performance that does not differ much from the Tikhonov-Miller filter performance. A (small) overestimation, however, degrades the performance dramatically, since it results in a clipping of object intensities. We propose a novel general method to estimate the background based on the dependency of nonlinear restoration algorithms on the background, and we demonstrate its applicability on real confocal images. PMID- 10708023 TI - Higher-order extrema in two-dimensional wave fields. AB - Higher-order extrema with topological indices greater than unity are discussed. Explicit constructions are given for their wave functions, and simple geometric rules are presented for analysis of their topological indices. Experimental means for verifying the theory with use of Gaussian laser beams are considered, unusual properties of optical vortices constructed from this new type of critical point are described, and applications to topologically based optical arithmetic are suggested. PMID- 10708024 TI - Axially symmetric on-axis flat-top beam. AB - A synthesis method for arbitrary on-axis intensity distributions from axially symmetric fields is developed in the paraxial approximation. As an important consequence, a new pseudo-nondiffracting beam, the axially symmetric on-axis flat top beam (AFTB), is given by an integral transform form. This AFTB is completely determined by three simple parameters: the central spatial frequency S(c), the on axis flat-top length L, and the on-axis central position z(c). When LS(c) >> 1, this AFTB can give a nearly flat-top intensity distribution on the propagation axis. In particular, this AFTB approaches the nondiffracting zero-order Bessel J0 beam when L--> infinity. It is revealed that the superposition of multiple AFTB fields can give multiple on-axis flat-top intensity regions when some appropriate conditions are satisfied. PMID- 10708025 TI - Singularities and Rayleigh's hypothesis for diffraction gratings. AB - The foci or branch points are found for a sinusoidal boundary. It is shown how they determine the range of validity of Rayleigh's hypothesis for diffraction gratings. PMID- 10708026 TI - Electromagnetic fields for a spheroidal particle with an arbitrary embedded source. AB - A spheroidal coordinate separation-of-variables solution has been developed for the determination of the internal, near-surface, and scattered fields of a spheroid (either prolate or oblate) with an embedded source of arbitrary type, location, and orientation. Presented results for (1,0) and (1,1) electric multipoles embedded in 2:1 axis ratio prolate and oblate spheroids (equal volume sphere size parameter equal to 20) illustrate that the presence of the spheroid interface can have a profound effect on the corresponding far-field scattering pattern. The calculation procedure could be used, for example, to model the emission of inelastic scattered light (Raman, fluorescence, etc.) from biological particles of appreciably elongated (prolatelike) or appreciably flattened (oblatelike) geometries. PMID- 10708027 TI - Analytical model of Doppler spectra of coherent light backscattered from rotating cones and cylinders. AB - The interaction between a rotating object and laser beams has been studied in the field of laser Doppler velocimetry, where two incident laser beams are focused on one small spot of the rotating surface and interferometry is used. In the case of a single incident laser beam illuminating a relatively large area of the rotating surface, both the Doppler broadening and the reflected-power level are dictated by points distributed over a wide curved area at varying angles of incidence. An analytical model of spectra in backscatter from cones and cylinders rotating around their axes is presented. This analytical solution may contribute to laser Doppler velocimetry as well as ladar applications. PMID- 10708028 TI - Confocal laser scanning ophthalmoscope and spherical harmonics used as a possible aid to detect glaucoma. AB - We present a procedure whereby the confocal laser scanning ophthalmoscope can be used to extract information about the three-dimensional structure of the central excavated area or the cup of the optic nerve head of the eye. The data are analyzed in terms of spherical harmonics. It is hypothesized that the shape of the cup of the optic nerve head for a normal eye can be parameterized by a specific set of spherical harmonic coefficients and is different from the set of coefficients describing a glaucomatous eye. The sets of coefficients are analyzed by using multivariate statistics and can in turn be used to classify new observations. Preliminary results indicate that there are significant differences in the coefficients and that the procedure might have potential as a diagnostic aid for the detection or the screening of glaucoma. PMID- 10708029 TI - Analysis of optical coherence tomography systems based on the extended Huygens Fresnel principle. AB - We have developed a new theoretical description of the optical coherence tomography (OCT) technique for imaging in highly scattering tissue. The description is based on the extended Huygens-Fresnel principle, valid in both the single- and multiple-scattering regimes. The so-called shower curtain effect, which manifests itself in a standard OCT system, is an inherent property of the present theory. We demonstrate that the shower curtain effect leads to a strong increase in the heterodyne signal in a standard OCT system. This is in contrast to previous OCT models, where the shower curtain effect was not taken into account. The theoretical analysis is verified by measurements on samples consisting of aqueous suspensions of microspheres. Finally, we discuss the use of our new theoretical model for optimization of the OCT system. PMID- 10708030 TI - Convergence of electromagnetic field components across discontinuous premittivity profiles. AB - We provide further insight into why the inverse rule [J. Opt. Soc. Am. A 13, 1870 (1996)] for multiplying two finite Fourier series of two pairwise discontinuous functions yields correct results at the point of discontinuity. PMID- 10708031 TI - Flicker-photometric electroretinogram estimates of L:M cone photoreceptor ratio in men with photopigment spectra derived from genetics. AB - Relative proportions of long-wavelength-sensitive (L) to middle-wavelength sensitive (M) cones were estimated by use of the flicker-photometric electroretinogram (ERG). It has been demonstrated that a major source of error in estimates of cone proportions from spectral luminosity functions is the known variation in the lambda(max) of the photopigments [Vision Res. 38, 1961 (1998)]. To correct for these errors, estimates of cone proportions were derived by use of individualized L-cone spectral sensitivity curves deduced from photopigment gene sequences from each subject. For some individuals this correction made a large difference in the estimated cone proportions compared with the value obtained when a fixed standard L cone was assumed. The largest discrepancy occurred in a man estimated to have 62% L cones (L:M ratio 1.6:1) when a standard L pigment was assumed but a value of 80% L cones (L:M ratio 4:1) when his individualized L-cone spectrum was used. From repeated measurements made with the ERG, it was determined that individual estimates of the relative L-to-M cone contributions, expressed as %L cones, are usually reliable within approximately 2%. The average L:M ratio for 15 male subjects was estimated at 2:1 (67% L cones). Previously, a large range of individual variability was reported for L:M ratios obtained from photometry. An unresolved issue concerns how much of the range might be attributed to error. Here efforts have been taken to markedly reduce measurement error. Nonetheless, a large range of individual differences persists. Estimated L:M ratios for individuals ranged from 0.6:1 to 12:1 (40% L to 92% L). PMID- 10708032 TI - Relative number of long- and middle-wavelength-sensitive cones in the human fovea. AB - Flicker photometric measurements yield spectral sensitivity curves that are well fitted by sums of the spectral sensitivity curves of long-wavelength-sensitive (L) cones and middle-wavelength-sensitive (M) cones if the L cones are given twice the weight of the M cones. This result has been interpreted as implying that L cones are more numerous than M cones but is also consistent with a different numerical ratio, say, 1:1, and with the assignment of greater weight to the L cone input than to the M cone input by the mechanism subserving flicker photometry. Measurements of temporal sensitivity are presented for lights that modulate the inputs of either only the L cones or only the M cones. Sensitivity to modulation of the L cones is approximately twice that of modulation of the M cones at approximately 30 Hz, but that advantage disappears at approximately 2 Hz. Thus flicker sensitivity is equivocal with regard to cone numerosity. Electrophysiological, anatomical, and psychophysical evidence is reviewed, with particular weight placed on the statistics of color appearance of small, brief, monochromatic lights and on increment thresholds measured on the same observers. It is concluded that, in the central fovea, the ratio of L:M cone numbers is close to unity and may not be so variable as is usually supposed. PMID- 10708033 TI - L/M cone ratios in human trichromats assessed by psychophysics, electroretinography, and retinal densitometry. AB - Estimates of the relative numbers of long-wavelength-sensitive (L) and middle wavelength-sensitive (M) cones vary considerably among normal trichromats and depend significantly on the nature of the experimental method employed. Here we estimate L/M cone ratios in a population of normal observers, using three psychophysical tasks-detection thresholds for cone-isolating stimuli at different temporal frequencies, heterochromatic flicker photometry, and cone contrast ratios at minimal flicker perception--as well as flicker electroretinography and retinal densitometry. The psychophysical tasks involving high temporal frequencies, specifically designed to tap into the luminance channel, provide average L/M cone ratios that significantly differ from unity with large interindividual variation. In contrast, the psychophysical tasks involving low temporal frequencies, chosen to tap into the red-green chromatic channel, provide L/M cone ratios that are always close to unity. L/M cone ratios determined from electroretinographic recordings or from retinal densitometry correlate with those determined from the high-temporal-frequency tasks. These findings suggest that the sensitivity of the luminance channel is directly related to the relative densities of the L and the M cones and that the red-green chromatic channel introduces a gain adjustment to compensate for differences in L and M cone signal strength. PMID- 10708034 TI - Cone pigment gene expression in individual photoreceptors and the chromatic topography of the retina. AB - Human trichromatic vision is based on three classes of cones: L, M, and S (long-, middle-, and short-wavelength sensitive, respectively). Individuals can have more than one M and/or more than one L pigment gene on the X chromosome along with an S pigment gene on chromosome 7. In some people the X-linked pigment gene array can include polymorphic variants that encode multiple, spectrally distinct cone photopigment subtypes. A single-cell, polymerase chain reaction approach was used to examine visual pigment gene expression in individual human cone cells and identify them as L or M. The ratio of L:M pigment gene expression was assayed in homogenized retinal tissues taken from the same eyes. Results indicate that there is a close correspondence between the cone ratio determined from counting single cells and the L:M pigment mRNA ratio estimated from homogenized pieces of retina. The results also show that the different pigment genes in one array are often expressed at very different levels, giving rise to unequal numbers of L and M cones. Expression of only one photopigment gene was detected in each cone cell. However, individual males can have more than the classically described three spectrally distinct cone types in their retinas. PMID- 10708036 TI - Comparison of red-green equiluminance points in humans and macaques: evidence for different L:M cone ratios between species. AB - The human spectral luminosity function (V(lambda)) can be modeled as the linear sum of signals from long-wavelength-selective (L) and middle-wavelength-selective (M) cones, with L cones being weighted by a factor of approximately 2. This factor of approximately 2 is thought to reflect an approximate 2:1 ratio of L:M cones in the human retina, which has been supported by studies that allow for more direct counting of different cone types in the retina. In contrast to humans, several lines of retinally based evidence in macaques suggest an L:M ratio closer to 1:1. To investigate the consequences of differences in L:M cone ratios between humans and macaques, red-green equiluminance matches obtained psychophysically in humans (n = 11) were compared with those obtained electrophysiologically from single neurons in the extrastriate middle temporal visual area of macaques (M. mulatta, n = 5). Neurons in the middle temporal visual area were tested with sinusoidal red-green moving gratings across a range of luminance contrasts, with equiluminance being defined as the red-green contrast yielding a response minimum. Human subjects were tested under analogous conditions, by a minimally distinct motion technique, to establish psychophysical equiluminance. Although red-green equiluminance points in both humans and macaques were found to vary across individuals, the means across species differed significantly; compared with humans, macaque equiluminance points reflected relatively greater sensitivity to green. By means of a simple model based on equating the weighted sum of L and M cone signals, the observed red-green equiluminance points were found to be consistent with L:M cone ratios of approximately 2:1 in humans and 1:1 in macaques. These data thus support retinally based estimates of L:M cone ratios and further demonstrate that the information carried in the cone mosaic has functional consequences for red-green spectral sensitivity revealed perceptually and in the dorsal stream of visual cortex. PMID- 10708035 TI - Interindividual and topographical variation of L:M cone ratios in monkey retinas. AB - We analyzed the ratio of L:M cone photopigment mRNA in the retinas of Old World monkeys, using the method of rapid polymerase chain reaction-single-strand conformation polymorphism. The L:M cone pigment mRNA ratio in whole retina ranged from 0.6 to 7.0, with a mean of approximately 1.6 (standard deviation, +/- 0.56; n = 26). There was no change in this ratio with eccentricity up to 9 mm (approximately 45 degrees), though the ratio was approximately 30% greater in temporal than in nasal retina. The mRNA ratios are in good agreement with the L:M cone ratio in these same retinas, inferred from electrophysiological recordings of cone signal gain in horizontal cell interneurons. The correlation between mRNA ratios and physiological cone gain ratio supports the conclusion that both measures reflect the relative number of L and M cones. PMID- 10708037 TI - Spatial order in short-wavelength-sensitive cone photoreceptors: a comparative study of the primate retina. AB - We compared the spatial distribution of short-wavelength-sensitive (SWS or blue) cone photoreceptors in the retinas of eight primate species. The regularity of the SWS cone array was quantified with a statistic (packing factor) that varies between a random distribution (0) and a triangular array (1). We find wide variability among species, with packing factors varying between 0.06 and 0.3. The SWS cone array in at least two New World monkey species is indistinguishable from a random array. The SWS cone density gradient across the retina was measured in the capuchin monkey Cebus apella and the squirrel monkey Saimiri sciureus. Both species show a peak density of 5,000-8,000 cells/mm2 at the fovea and a 50-fold central-peripheral density gradient. In contrast to the wide variation in local regularity, the spatial density and the topography of SWS cones are well preserved across primates. PMID- 10708038 TI - Photoreceptor distribution in the retinas of subprimate mammals. AB - Relevant data on the distribution of color cones are summarized, with special emphasis on the marked dorsoventral asymmetries observed in a number of mammalian species. In addition, an overview is given of studies that demonstrate the coexistence of two visual pigments within the same cone cell. The biological significance of these phenomena is discussed in conjunction with comparative immunocytochemical analyses of subprimate retinas. Based on various cone distribution patterns and temporal and spatial visual pigment coexpression, two models of cone photoreceptor differentiation are suggested. PMID- 10708039 TI - Irregular S-cone mosaics in felid retinas. Spatial interaction with axonless horizontal cells, revealed by cross correlation. AB - In most mammals short-wavelength-sensitive (S) cones are arranged in irregular patterns with widely variable intercell distances. Consequently, mosaics of connected interneurons either may show some type of correlation to photoreceptor placement or may establish an independent lattice with compensatory dendritic organization. Since axonless horizontal cells (A-HC's) are supposed to direct all dendrites to overlying cones, we studied their spatial interaction with chromatic cone subclasses. In the cheetah, the bobcat, and the leopard, anti-S-opsin antibodies have consistently colabeled the A-HC's in addition to the S cones. We investigated the interaction between the two cell mosaics, using autocorrelation and cross-correlation procedures, including a Voronoi-based density probe. Comparisons with simulations of random mosaics show significantly lower densities of S cones above the cell bodies and primary dendrites of A-HC's. The pattern results in different long-wavelength-sensitive-L- and S-cone ratios in the central versus the peripheral zones of A-HC dendritic fields. The existence of a related pattern at the synaptic level and its potential significance for color processing may be investigated in further studies. PMID- 10708040 TI - Physiology of L- and M-cone inputs to H1 horizontal cells in the primate retina. AB - In the primate retina, H1 horizontal cells form an electrically coupled network and receive convergent input from long- (L-) and middle- (M-) wavelength sensitive cones. Using an in vitro preparation of the intact retina to record the light-evoked voltage responses of H1 cells, we systematically varied the L- and M cone stimulus contrast and measured the relative L- and M-cone input strength for 137 cells across 33 retinas from three Old World species (Macaca nemestrina, M. fascicularis, and Papio anubis). We found that the L- and the M-cone inputs were summed by the H1 cell in proportion to the stimulus cone contrast, which yielded a measure of what we term L- and M-cone contrast gain. The proportion of L-cone contrast gain was highly variable, ranging from 25% to 90% [mean +/- standard deviation, (60 +/- 14)%]. This variability was accounted for by retinal location within an individual, with the temporal retina showing a consistently higher percentage of L-cone gain, and by large overall variation across individuals, with the mean percentage of L-cone gain ranging from 32% to 80%. We hypothesize that the relative L- and M-cone contrast gain is determined simply by the relative number of L and M cones in the H1 cell's receptive field and that the variability in L- and M-cone contrast gain reflects a corresponding variability in the mosaic of L and M cones. PMID- 10708041 TI - Representation of cone signals in the primate retina. AB - Vision begins with specialized retinal circuits that encode diverse types of information. For Old World primates, these circuits sample three submosaics formed by cone photoreceptors sensitive to short, middle, and long wavelengths. For spatial acuity, the photon catch between any two cones is compared for discrimination of patterns as fine as the cone mosaic. For color vision, the photon catch between different cone types is compared for discrimination of fine spectral differences on the basis of hue. The retinal circuits for these two tasks differ at the synaptic level to form distinct representations of signals from the cone mosaic. PMID- 10708042 TI - Functional consequences of the relative numbers of L and M cones. AB - Direct imaging of the retina by adaptive optics allows assessment of the relative number of long-wavelength-sensitive (L) and middle-wavelength-sensitive (M) cones in living human eyes. We examine the functional consequences of variation in the relative numbers of L and M cones (L/M cone ratio) for two observers whose ratios were measured by direct imaging. The L/M cone ratio for the two observers varied considerably, taking on values of 1.15 and 3.79. Two sets of functional data were collected: spectral sensitivity measured with the flicker electroretinogram (ERG) and the wavelength of unique yellow. A genetic analysis was used to determine L and M cone spectra appropriate for each observer. Rayleigh matches confirmed the use of these spectra. We determined the relative strength of L and M cone contributions to ERG spectral sensitivity by fitting the data with a weighted sum of L and M cone spectra. The relative strengths so determined (1.06 and 3.38) were close to the cone ratios established by direct imaging. Thus variation in L/M cone ratio is preserved at the sites tapped by the flicker ERG. The wavelength of unique yellow varied only slightly between the two observers (576.8 and 574.7 nm). This small variation indicates that neural factors play an important role in stabilizing unique yellow against variation in the L/M cone ratio. PMID- 10708043 TI - L and M cone relative numerosity and red-green opponency from fovea to midperiphery in the human retina. AB - The relative numerosity of the long-wavelength-sensitive (L) and middle wavelength-sensitive (M) cones and the red-green color appearance, as assessed by means of unique yellow, are stable from fovea to midperiphery (+/- 28 deg nasotemporal). As foveal tests decrease in size, unique yellow progressively shifts toward longer wavelengths, favoring a model of red-green opponency carried by cells whose centers receive input from either L or M cones and whose surrounds receive mixed contributions from both. Individual differences in unique yellow over a 20-nm range and the relative numerosity of L and M cones can be linked by means of this model, suggesting that the relative number of L and M cones is a factor that regulates individual variations in red-green color appearance. PMID- 10708044 TI - Effect of the short-wavelength-sensitive-cone mosaic and rods on the locus of unique green. AB - A primary goal of this study was to establish whether the magnitude of the short wavelength-sensitive- (S-) cone signal into the yellow/blue (Y/B) mechanism was influenced by the absolute or the relative numbers of S cones. This was assessed by measuring the locus of unique green for various test sizes at four eccentric locations chosen to exploit differences in the underlying mosaic of S cones. In general, the locus of unique green was unaffected by test size, retinal quadrant, or rod input but was influenced by retinal eccentricity. The locus of unique green shifted to shorter wavelengths as retinal eccentricity increased from 1 degrees to 8 degrees. The data do not support a model whereby the S-cone signal is determined by the absolute number of S cones, but a model based on the relative number of S cones cannot be eliminated. PMID- 10708045 TI - Cost of cone coupling to trichromacy in primate fovea. AB - Cone synaptic terminals couple electrically to their neighbors. This reduces the amplitude of temporally uncorrelated voltage differences between neighbors. For an achromatic stimulus coarser than the cone mosaic, the uncorrelated voltage difference between neighbors represents mostly noise; so noise is reduced more than the signal. Here coupling improves signal-to-noise ratio and enhances contrast sensitivity. But for a chromatic stimulus the uncorrelated voltage difference between neighbors of different spectral type represents mostly signal; so signal would be reduced more than the noise. This cost of cone coupling to encoding chromatic signals was evaluated using a compartmental model of the foveal cone array. When cones sensitive to middle (M) and long (L) wavelengths alternated regularly, and the conductance between a cone and all of its immediate neighbors was 1,000 pS (approximately 2 connexons/cone pair), coupling reduced the difference between the L and M action spectra by nearly fivefold, from about 38% to 8%. However, L and M cones distribute randomly in the mosaic, forming small patches of like type, and within a patch the responses to a chromatic stimulus are correlated. In such a mosaic, coupling still reduced the difference between the L and M action spectra, but only by 2.4-fold, to about 18%. This result is independent of the L/M ratio. Thus "patchiness" of the L/M mosaic allows cone coupling to improve achromatic contrast sensitivity while minimizing the cost to chromatic sensitivity. PMID- 10708047 TI - Principles of community psychiatry. AB - Our primary goal in community psychiatry is to satisfy the service needs of a bounded population for whose mental health we have accepted responsibility and accountability. We base our programs on public health practice models: These direct us to focus on segments of our population which are currently exposed to harmful biopsycho-social factors that increase their risk of becoming mentally ill. We focus on preventing psychosocial problems or their consequences by reducing their population rates: either the incidence of new cases (primary prevention), the prevalence of all existing cases (secondary prevention), and the rates of residual disability (tertiary prevention). We increase our efficiency and effectiveness by organizing our program on the basis of crisis theory which demands that we reach out to people in crisis and provide them with immediate guidance and help to master their current difficulties during the short period when they are open to influence and amenable to change in ways that have long term mental health consequences. We spread our own influence by organizing support groups and we multiply many-fold our impact on the huge problems involved in covering the needs of our population by recruiting the collaboration of other professional caregivers and non-professional helpers. We enhance the mental health component in the daily work of all caregiving agencies and institutions and individual professionals in the community through education and mental health consultation and collaboration. We also reach out to assist non-professional caregiving individuals and organizations, especially those who provide mutual help to fellow sufferers. In our latest work we are currently identifying harmful practices in our caregiving systems that actually harm those people whom we are trying to help. We are in the process of developing methods for reducing this system-generated damage. PMID- 10708046 TI - Spatial summation in human cone mechanisms from 0 degrees to 20 degrees in the superior retina. AB - The maximum area of complete spatial summation (i.e., Ricco's area) for human short-wavelength-sensitive-(S-) and long-wavelength-sensitive- (L-) cone mechanisms was measured psychophysically at the fovea and at 1.5 degrees , 4 degrees , 8 degrees , and 20 degrees along the vertical meridian in the superior retina. Increment thresholds were measured for three observers by a temporal two alternative forced-choice procedure. Test stimuli ranging from -0.36 to 4.61 log area (min2) were presented on concentric 12.3 degrees adapting and auxiliary fields, which isolated either an S- or an L-cone mechanism on the plateau of its respective threshold versus intensity function. Test flash durations were 50 and 10 ms for the S- and L-cone mechanisms, respectively. The data indicate that, from 0 degrees to 20 degrees, Ricco's area increases monotonically for the L-cone mechanism, is variable for the S-cone mechanism, and is larger for the S-cone mechanism than for the L-cone mechanism for essentially all retinal locations. This pattern of results most likely reflects differences in ganglion cell density and changes in neural convergence with retinal eccentricity. PMID- 10708048 TI - Four studies of crisis in parents of prematures. 1965. AB - The responses of 86 families to the birth of a premature baby have been investigated in four linked studies in order to refine the concept and understanding of crisis. Patterns of the grappling behavior during the crisis were identified which enabled accurate predictions of the short-term mental health outcome. Psychological tasks presented by the stress of premature delivery were also identified. The adequacy with which these tasks were accomplished was predictive of the patterns of early maternal care and mother-child relationships. Results indicate that this type of study is relevant to studies of the causation of mental health and mental illness and to preventive intervention. Certain methodologic and research implications are derived from these studies and point to further research effort which is now practical and desirable. PMID- 10708049 TI - The delivery of mental health services in the 21st century: bringing the community back in. AB - The community mental health movement of the 1960s enjoyed widespread public support but poorly served its intended target population of seriously mentally ill individuals because: (1) its professional values and technology were, at least initially, not well-oriented toward serving people with severe mental illness; (2) organizational structures linking Community Mental Health Centers with State Mental Health Agencies, State Hospitals, and other relevant service agencies were lacking; (3) ideologically driven aspirations diverted energies and resources into diffuse goals related to the achievement of social justice; and (4) performance objectives were not operationally defined or monitored. Since that time professional technologies and organizational linkages have substantially improved, but there has been a loss of public support for safety net services for the least well off, in part due to a general ascendence of individualist market values, declining civic engagement and reduced support for specialized services for the disadvantaged. A new community mental health movement would be less oriented towards stimulating broad community change, and more narrowly focused on building support among decision makers and the public at large to expand the availability of costly but effective and improved services for people with severe and persistent mental illness. PMID- 10708050 TI - Natural analogue peptides of an HIV-1 GP120 T-helper epitope antagonize response of GP120-specific human CD4 T-cell clones. AB - Neutralizing antibodies and specific cytotoxic T lymphocytes (CTL) may contribute to controlling viral spread, and ideally, to virus clearance in HIV infection. Both effector mechanisms depend on specific CD4 T-helper (Th) cells. Nevertheless, HIV hypervariability facilitates appearance of escape mutants for antibodies and for CTL responses. Here we also show that natural mutations (i.e., from sequences of different HIV strains) in an immunodominant Th epitope recognized by human CD4 clones specific for the envelope glycoprotein gp120 escape CD4 T-cell recognition. Furthermore, several natural analogue peptides exert an antagonistic function by inhibiting proliferative response of T cells specific to gp120 with a wild-type sequence. If similar events occur in vivo, they may represent an additional escape mechanism for HIV. In fact, antagonism for CD4 Th response may occur during superinfection with a different strain, or with the appearance of a variant carrying a mutated antagonistic sequence. In both cases, impaired Th cell function could lead to reduced immune control of HIV infection by interfering with CTL and antibody response. PMID- 10708051 TI - Neurophysiologic and immunologic abnormalities associated with feline immunodeficiency virus molecular clone FIV-PPR DNA inoculation. AB - Although direct feline immunodeficiency virus (FIV) proviral DNA inoculation has been shown to be infectious in cats, long-term studies to assess the pathogenic nature of DNA inoculation are lacking. We have recently reported that direct feline leukemia virus (FeLV) DNA inoculation resulted in infection and the development of FeLV-related disease end points with similar temporal expression and virulence to that of cats infected with whole virus. We show in this study that pFIV-PPR DNA inoculation resulted in infection of cats and the development of FIV-related immunologic and neurologic abnormalities. Infected cats demonstrated progressive loss of CD4+ lymphocytes resulting in decreased CD4:CD8 ratios. Neurologic dysfunction was demonstrated by increased bilateral frontal lobe slow-wave activity. Prolongation of the visual evoked potential peak latency onset response pattern also supported a similar progression of abnormal cortical response. Furthermore, histopathologic examination revealed lesions attributed to FIV infection in lymph node, thymus, brain, and lung. Finally, nested polymerase chain reaction detected FIV provirus in brain, bone marrow, mesenteric lymph node, thymus, spleen, tonsil, and liver. These results confirm that FIV DNA inoculation is an efficient model for study of the pathogenic nature of molecular clones in vivo and offers the opportunity to measure temporal genomic stability of a homogeneous challenge material. PMID- 10708052 TI - Effects of early antiretroviral treatment on HIV-1 RNA in blood and lymphoid tissue: a randomized trial of double versus triple therapy. Swiss HIV Cohort Study. AB - To assess the effects of early initiation of antiretroviral therapy on cell-free and cell-associated viral load in blood and lymphoid tissue, we performed a randomized, open-label, multicenter trial comparing a double (zidovudine + lamivudine) and triple (zidovudine + lamivudine + ritonavir) drug combination in treatment-naive, asymptomatic patients with CD4 counts >400 cells/microl. HIV-1 RNA was measured in plasma, peripheral blood mononuclear cells, and sequential tonsil or lymph node biopsies (27 patients); the study follow-up was 2 years. Among 42 randomized patients, the proportion with plasma HIV-1 RNA <50 copies/ml was 16% and 74% at week 24 (p<.001) in those randomized to double and triple therapy, respectively, necessitating frequent treatment intensification in the double arm. After a rapid decline within 4 weeks in both arms, cell-associated HIV-1 RNA decreased further only in those patients with sustained suppression of plasma viral load, but remained almost always detectable at low levels, indicating persisting transcription of viral RNA. CD4 counts increased by 200 to 250 cells/microl at week 96 in both arms without significant differences (intent to-treat analyses). Thus, even if treatment is initiated early in asymptomatic patients with preserved CD4 counts, three drugs are necessary to achieve sustained decreases of HIV load in blood and lymphoid tissue. PMID- 10708053 TI - Emergence of dual resistance to zidovudine and lamivudine in HIV-1-infected patients treated with zidovudine plus lamivudine as initial therapy. AB - Presence of mutations associated with resistance to zidovudine or lamivudine was determined in isolates of HIV-1 obtained after long-term follow-up of 64 infected individuals who received zidovudine, lamivudine, or both drugs as initial antiretroviral therapy. Zidovudine resistance mutations were less frequent in isolates from patients treated with combination lamivudine plus zidovudine compared with zidovudine alone, but these mutations accumulated over time. Phenotypic resistance to both drugs was found in isolates from 3 of 23 patients. In 3 other patients, lamivudine-resistant virus detected at week 12 was replaced by wild-type virus after longer follow-up, which correlated with a return to baseline levels of plasma HIV-1 RNA. These results show that dual resistance to zidovudine and lamivudine develops over time despite the delayed emergence of zidovudine-resistant mutations. These results also suggest a selective advantage in vivo for HIV-1 species that are wild-type at RT codon 184. PMID- 10708054 TI - Hyperlipidemia and insulin resistance are induced by protease inhibitors independent of changes in body composition in patients with HIV infection. AB - Although protease inhibitor (PI) therapy has improved the clinical status of patients with HIV infection, concerns have arisen that such treatment may have deleterious effects on glucose control, lipid metabolism, and body fat distribution. To determine whether initiation of PI therapy uniquely affects glucose and lipid metabolism, we analyzed paired data in HIV-infected patients before and after beginning antiretroviral therapy that included a PI (PI; N = 20) or lamivudine (3TC) but no PI (3TC; N = 9); and a control group on stable regimens that included neither of these agents (CONT: N = 12). Measurements included fasting glucose; insulin; triglycerides; total, high-density lipoprotein (HDL) and low-density lipoprotein (LDL) cholesterol; HIV RNA; CD4+ lymphocytes; weight; and total and regional body composition. Neither weight nor total or regional fat content changed significantly in any group during the period of observation. Nonetheless, in patients beginning PI therapy, there were significant increases in glucose (+9+/-3 mg/dl; p = .0136), insulin (+12.2+/-4.9 U/ml; p = .023), triglycerides (+53+/-17 mg/dl; p = .0069), and total and LDL cholesterol (+32+/-11 and +18+/-5 mg/dl; p = .0082 and .0026, respectively). None of these parameters changed significantly in the 3TC or CONT groups. The PI and 3TC groups experienced comparable increases in CD4+ lymphocytes, suggesting that the observed metabolic effects may be associated with PIs uniquely, rather than improvement in clinical status. However, it is also possible that the metabolic effects of PIs are associated with more effective viral suppression, because a greater proportion of patients in the PI group achieved undetectable levels of virus. We conclude that changes in glucose and lipid metabolism are induced by PI therapy in the absence of significant changes in weight or fat distribution. Longer periods of follow-up will be required to determine the clinical significance of these changes. However, at the moment, the risks associated with these metabolic effects do not appear to outweigh improvements in survival seen with PI therapy. PMID- 10708055 TI - Prediction of imminent complications in HIV-1-infected patients by markers of lymphocyte apoptosis. AB - OBJECTIVE: The aim of the study was to compare accepted surrogate markers of HIV disease progression with markers of lymphocyte apoptosis in their ability to predict short-term disease progression. METHODS: In all, 40 HIV-positive patients were studied prospectively and observed during follow-up for HIV-related adverse clinical events. Ex vivo apoptosis was measured with the markers CD95 expression, annexin V binding, and Apostain dye uptake by flow cytometry at baseline. Established markers of disease progression (CD4 count, HIV-RNA level, and CD8/38 count), CD8, B-cell, and natural killer (NK) cell counts were determined by standard procedures at baseline and after 6 months. RESULTS: In HIV-infected patients, CD95 expression and annexin V binding showed significantly elevated apoptosis in peripheral blood lymphocytes and all lymphocyte subsets at baseline compared with HIV-negative, healthy controls. Apostain failed to differentiate between HIV-infected patients and healthy controls. HIV-related complications could be predicted by CD4 and CD8/38 counts, but not HIV viral load as assessed by relative operating characteristic (ROC) analysis (CD4, p = .003; CD8/38, p = .031). A similar or even better diagnostic accuracy was found for CD95 expression in total lymphocytes (p<.001), the CD4+ (p = .003) and CD8+ (p = .005) T-cell subsets and for annexin V binding in CD4+ T cells (p = .005). When patients with CD4 counts <200 cells/microl were analyzed separately, only annexin V binding in CD4+ T cells, but none of the other prognostic markers could predict complications (p = .001). CONCLUSION: Determination of annexin V binding on CD4+ T cells may be a useful tool to monitor HIV-infected patients with low (<200 cells/microl) CD4 counts, as it can reliably assess the risk for imminent complications in such patients. PMID- 10708056 TI - Protective effect of CCR2-64I and not of CCR5-delta32 and SDF1-3'A in pediatric HIV-1 infection. AB - The effects of chemokine and chemokine receptor genetic polymorphisms such as stromal derived factor 1 (SDF1-3'A), CCR2-64I, and CCR5-delta32 associated with HIV-1 transmission and/or rate of disease progression in infected study subjects remain highly controversial and have been analyzed primarily only in adults. We have investigated whether these polymorphisms may provide similar beneficial effects in children exposed to HIV-1 perinatally. The prevalence of CCR2-64I allele was significantly increased (p = .03) and the CCR2-64I genotype distribution was not in Hardy-Weinberg equilibrium, among HIV-1-exposed uninfected infants. Moreover, in the HIV-1-infected group, a delay to AIDS progression was observed among carriers of CCR2-64I allele. This is the first report that suggests a protective role of CCR2-64I allele in mother-to-infant HIV 1 transmission and documents a delay in disease progression, after the child has been infected with HIV-1. However, SDFI-3'A and CCR5-delta32 alleles did not modify the rate of HIV-1 transmission or disease progression in HIV-1-infected children. PMID- 10708057 TI - Molecular epidemiology of HIV-1 in Switzerland: evidence for a silent mutation in the C2V3 region distinguishing intravenous drug users from homosexual men. Swiss HIV Cohort Study. AB - OBJECTIVES: To study the molecular epidemiology of HIV-1 strains found in Switzerland and to determine possible genetic linkages among strains sorted by risk group or geographic region. DESIGN: A cross-sectional, clinic-based survey of HIV-1 molecular sequences and linked patient history from Swiss people. METHODS: Specimens were collected from 215 HIV-1-infected people in HIV outpatient clinics of four tertiary referral centers (Lausanne, St. Gallen, Zurich, and Basel) between May and August 1996, mainly from homosexual men, injecting drug users (IDU), and heterosexually infected people. In addition, specimens collected between 1991 and 1995 in the HIV outpatient clinic at University of Geneva were included into this survey. These specimens were collected primarily for an ongoing, prospective cohort (Swiss HIV Cohort Study). Direct C2V3C3 sequences of the env gene were determined from 158 samples of peripheral blood mononuclear cells. Genetic data were analyzed with the available patient history on each specimen. RESULTS: As found in other previous studies in Europe, primarily subtype B viruses were identified, whereas seven (4%) of 158 were non-subtype B: one subtype D, four subtype A, and two subtype E. Five of seven non-B subtypes occurred in immigrants from African or Asian countries and all seven were found exclusively in individuals who had been infected by heterosexual contact. No significant clustering of strains within different study sites or risk groups was found. A silent mutation (LAI env 834) occurred significantly more often in IDU than in homosexual men (p<.001). CONCLUSIONS: Although the lack of significant clustering of strains by risk group or geographic region may result from early introduction of subtype B viruses in Switzerland, the strong association of a silent mutation with IDU suggests that, early in the epidemic, there was a unique founder virus among IDUs. The HIV epidemic in Switzerland is still predominantly caused by subtype B viruses. PMID- 10708058 TI - Genetic analysis of HIV-1 samples from Spain. AB - To characterize the viruses responsible for the HIV-1 epidemic in Spain, we genetically characterized 79 samples obtained from Spanish patients with different risk practices (injecting drug users and male homosexuals) in two regions (Madrid and Navarra). Genetic characterization was carried out by nucleotide sequencing in the C2-V3-C3 region and by phylogenetic analysis. All samples were of subtype B except one that clustered with clade F. Because no segregation of samples was determined according to the risk group of patients nor to their geographic origin, the Spanish samples analyzed constitute a single group of viruses. These data, along with the starlike topology of the phylogenetic tree, support the existence of a single introduction of HIV-1 subtype B in Spain. The mean genetic distance among subtype B sequences was of 13.9%+/-0.06% (range, 5%-25%), suggesting an epidemic of long evolution. Analysis of sequences in relation to isolation dates revealed an increase in the heterogeneity of the nucleotide sequences with time. According to these data, a divergence rate of 0.49%+/-0.11% per year was calculated for the Spanish samples during the period analyzed. PMID- 10708059 TI - Risk factors for developing tuberculosis in HIV-1-infected adults from communities with a low or very high incidence of tuberculosis. AB - OBJECTIVE: To estimate the incidence rate of tuberculosis in HIV-1-infected adults resident in a region with a high tuberculosis prevalence and to identify clinical and laboratory parameters associated with increased risk of developing tuberculosis. METHODS: Adult patients going to the University of Cape Town HIV clinics between January 1986 and May 1996. The following variables were assessed for the risk of developing tuberculosis: ethnicity, employment and education status, World Health Organization (WHO) clinical stage, erythrocyte sedimentation rate (ESR), CD4+ count, and total lymphocyte count. Tuberculin skin test data were not available. RESULTS: There were 198 prevalent and 144 incident cases of tuberculosis in the cohort of 1206 patients. The incidence rate of tuberculosis risk was 10.4/100 person years. WHO clinical stages 3 and 4 (risk ratio [RR], 3.4; 95% confidence interval [CI], 1.8-6.4), ESR >75 mm/hour (RR, 3.5; CI, 1.8 6.5) and being a member of a high-prevalence tuberculosis community (RR, 2.5; CI, 1.2-5.1) were independently associated with the risk of developing tuberculosis. CONCLUSIONS: HIV-infected adults in Cape Town are at high risk of developing tuberculosis irrespective of tuberculin skin testing. The risk increases markedly with HIV disease progression. Patients at extremely high risk can be identified on the basis of demographic and clinical features. Such individuals would be suitable for targeted tuberculosis prophylaxis. PMID- 10708060 TI - When obesity is desirable: a longitudinal study of the Miami HIV-1-infected drug abusers (MIDAS) cohort. AB - Despite widespread nutrient deficiencies, a substantial proportion of the MIDAS cohort exhibits obesity, which has been linked to immune dysregulation in other clinical settings. Herein, the effects of obesity on immune function, disease progression, and mortality were evaluated longitudinally in 125 HIV-1 seropositive drug users, with comparison measures in 148 HIV-1-seronegative controls. Data were collected at a community clinic from 1992 to 1996, before administration of highly active antiretroviral therapy. Results indicated that overweight/obesity, defined as body mass index (BMI; kg/m2) > or =27, was evident in 18% of the HIV-1-seropositive patients and 29% of the seronegative patients. At baseline, no significant immunologic differences were observed among lean, nonobese, and obese groups. Over an 18-month period, 60.5% of the nonobese HIV-1 seropositive patients exhibited a 25% decline in CD4 cell count, compared with 18% of the obese patients (p<.004). During the follow-up period, 38% of the lean and 13% of the nonobese study subjects died of HIV-1-related causes. Measurements of BMI were inversely associated with progression to death, independent of CD4 count <200 cells/mm3 (p<.02). These data suggest that mild-to-moderate obesity in HIV-1-infected chronic drug users does not impair immune function and is associated with better HIV-1-related survival. PMID- 10708061 TI - Prevalence and route of transmission of infection with a novel DNA virus (TTV), hepatitis C virus, and hepatitis G virus in patients infected with HIV. AB - OBJECTIVES: To evaluate the prevalence, route of transmission and clinical significance that current co-infection with TT virus (TTV), hepatitis C virus (HCV), and hepatitis G virus (HGV) have in HIV-1-infected patients. DESIGN: Presence of TTV, HCV, and HGV was analyzed in plasma samples from 160 HIV-1 infected patients with parenteral (38 intravenous drug users [IVDUs] and 41 patients with hemophilia) or sexual (39 homosexuals and 42 heterosexuals) risk of exposure, and in 168 volunteer blood donors. Alanine aminotransferase (ALT) levels and CD4+ counts were also analyzed. METHODS: HCV and HGV RNA were detected by specific reverse transcriptase (RT) nested polymerase chain reaction (PCR) and TTV DNA by specific heminested PCR. RESULTS: TTV DNA was detected in 39% of the patients and in 14% of the volunteer blood donors. HCV and HGV infections were detected in 42% and in 14% of the patients, and in 0% and 3% of the blood donors, respectively. Prevalences of TTV and HCV infection were higher among patients with parenteral (62% and 68%) than in those with sexual (17% and 16%) risk of exposure. A higher prevalence of TTV infection (but not of HCV or HGV infection) was observed among patients with hemophilia (76%) than IVDUs (47%), and among homosexuals (26%) than among heterosexuals (10%). Abnormal ALT levels were related with the presence of HCV infection, independently of the detection of TTV DNA. TTV infection did not seem to alter the levels of CD4+ T cells. CONCLUSIONS: Prevalence of current TTV infection is high among HIV-infected patients with parenteral risk of exposure, but TTV is also transmitted through sexual routes; detection of TTV does not seem to influence the clinical or immune status of HIV infected patients. PMID- 10708062 TI - Switch to an antiretroviral treatment of expected lower potency after effective highly active antiretroviral therapy (HAART) PMID- 10708063 TI - HIV viral load and viral cultures in sexually active heterosexual men. PMID- 10708064 TI - Perimenopausal symptomatology among HIV-infected women at least 40 years of age. PMID- 10708065 TI - Factors associated with the decline in sexually transmitted infections among HIV positive women. PMID- 10708066 TI - Polymerase chain reaction monitoring of HIV-1 infection: should the CCR5 genotype be considered? PMID- 10708067 TI - Peripheral blood lymphocyte proliferation specific for HTLV-Tax peptides in enzyme-linked immunosorbent assay-negative HTLV DNA-positive blood donors. PMID- 10708068 TI - Commentary on monaural and binaural loudness measures in cochlear implant users with contralateral residual hearing by P. Blamey, G. Dooley, C. James, and E. Parisi. PMID- 10708069 TI - Monaural and binaural loudness measures in cochlear implant users with contralateral residual hearing. AB - OBJECTIVE: The aim was to measure the loudness of monaural and binaural stimuli in a group of cochlear implant users who had residual hearing in the nonimplanted ear, and to consider the implications of these measures for a binaural fitting consisting of a hearing aid and an implant in opposite ears. Three independent hypotheses were addressed: that the shapes of the electric and acoustic loudness growth functions would be similar, although the dynamic ranges would differ; that standard implant and hearing aid fittings would result in substantial loudness mismatches between the acoustic and electric signals; and that loudness summation would occur for binaural combinations of electric and acoustic signals. DESIGN: A modified version of the "Loudness Growth in 1/2-Octave Bands" method (Allen, Hall, & Jeng, 1990) was used to measure loudness growth for each ear of nine subjects. At the time of the experiment, the subject group included all implant users in Melbourne and Denver who were available for research and who also had sufficient residual hearing to use a hearing aid in the nonimplanted ear. Five acoustic frequencies and five electrodes were measured for each subject. The same subjects also estimated the loudness of a set of stimuli including monaural and binaural signals chosen to cover the loudness range from very soft to loud. RESULTS: The shapes of the averaged loudness growth functions were similar in impaired and electrically stimulated ears, although the shapes of iso-loudness curves were quite different in the two ears, and dynamic ranges varied considerably. Calculations based on the psychophysical data demonstrated that standard fitting procedures for cochlear implants and hearing aids lead to a complex pattern of loudness differences between the ears. A substantial amount of loudness summation was observed for the binaural stimuli, with most summation occurring when the acoustic and electric components were of equal loudness. This is consistent with observations for subjects with normal hearing and subjects with bilaterally impaired hearing. CONCLUSIONS: These experiments provide data on which criteria and methods for the binaural fitting of cochlear implants and hearing aids may be based. It is unlikely that standard monaural fitting methods for cochlear implants and hearing aids will result in balanced loudness between the two ears across a reasonably broad range of frequencies and levels. It is also likely that output levels of both devices will need to be reduced relative to a monaural fitting to compensate for the binaural summation of loudness in some listeners. PMID- 10708070 TI - Preimplant measures of preverbal communicative behavior as predictors of cochlear implant outcomes in children. AB - OBJECTIVE: Comparison of preverbal communication behavior in young children before receiving cochlear implants with outcomes 3 yr after implantation on speech identification and speech production tasks, to assess whether outcomes can be predicted from characteristics inherent to the child before implantation. DESIGN: Video recordings of preverbal communicative behavior were examined before use of the implant to quantify turn-taking and demonstration of autonomy by the child. Speech identification ability was measured 3 yr after implantation based on the Iowa Closed-Set Speech Perception Sentence Test, continuous discourse tracking, and an observational measure of telephone use. Speech production ability was measured 3 yr after implantation by the Edinburgh Articulation Test. Associations between the preimplant measures and the 3-yr outcomes were assessed by correlation analysis of data from 33 children. RESULTS: The 3-yr performance measures of speech identification were correlated with the preimplant measure of autonomy. Telephone use and speech production ability were not significantly associated with the preimplant measures. CONCLUSIONS: Up to a fourth of the variance in speech identification performance 3 yr after cochlear implantation of young children may be predicted from characteristics that are inherent to the child before implantation. Those characteristics are represented by the demonstration of autonomy in preverbal communicative interactions, whether by means of vocalization or by gesture. If those characteristics are acquired during infancy, outcomes in children with auditory prostheses including cochlear implants may be enhanced by activities that encourage autonomy in early years. PMID- 10708071 TI - The effect of reduced dynamic range on speech understanding: implications for patients with cochlear implants. AB - OBJECTIVE: To determine the effect of reduced dynamic range on speech understanding when the speech signals are processed in a manner similar to a 6 channel cochlear implant speech processor. DESIGN: Signals were processed in a manner similar to a 6-channel cochlear implant processor and output as a sum of sine waves with frequencies equal to the center frequencies of the analysis filters. The amplitudes of the sine waves were compressed in a systematic fashion to simulate the effect of reduced dynamic range. The compressed signals were presented to 10 normal-hearing listeners for identification. RESULTS: There was a significant effect of compression for all test materials. The effect of the compression on speech understanding was different for the three test materials (vowels, consonants, and sentences). Vowel recognition was affected the most by the compression, and consonant recognition was affected the least by the compression. Feature analysis indicated that the reception of place information was affected the most. Sentence recognition was moderately affected by the compression. CONCLUSIONS: Dynamic range should affect the speech perception abilities of cochlear implant users. Our results suggest that a relatively wide dynamic range is needed for a high level of vowel recognition and a relatively small dynamic range is sufficient to maintain consonant recognition. We infer from this outcome that, if other factors were held equal, an implant patient with a small dynamic range could achieve moderately high scores on tests of consonant recognition but poor performance on vowel recognition, and that it is more likely for an implant patient with a large dynamic range to obtain high scores on vowel recognition than for an implant patient with a small dynamic range. PMID- 10708072 TI - Comparison of linear gain and wide dynamic range compression hearing aid circuits II: aided loudness measures. AB - OBJECTIVES: The goal of this study was to test the theoretical advantages of a single-channel wide dynamic range compression (WDRC) circuit fitted using the DSL method for increased dynamic range and normalized loudness growth. DESIGN: Ten adolescents and young adults with moderate to severe sensorineural hearing loss were fitted monaurally with the Siemens Viva 2 Pro behind-the-ear instrument set to DSL 4.0 targets for both linear gain and WDRC processing. Threshold, upper limit of comfort and loudness growth were measured in the unaided, linear gain and WDRC conditions for warble tones, environmental sounds and speech. Twelve adult listeners with normal hearing also were tested monaurally in the unaided condition to provide normative data for comparison purposes. RESULTS: The WDRC hearing aid provided a greater input dynamic range than the linear circuit for all stimuli. The dynamic range was normalized for more subjects with the WDRC than the linear hearing aid. In addition, exponential loudness growth functions fitted to the loudness growth data showed that, on average, loudness growth was more normalized with the WDRC hearing aid fitted to DSL[i/o] targets than the linear hearing aid fitted to DSL[i/o] targets. CONCLUSIONS: WDRC processing, fitted using the DSL[i/o] method, has potential applications in hearing aid fittings for listeners with moderate to severe hearing loss because it provides an audible, comfortable and tolerable amplified signal across a wider range of inputs than linear gain processing, without the need for volume control adjustments. PMID- 10708073 TI - Directivity quantification in hearing aids: fitting and measurement effects. AB - OBJECTIVE: To evaluate the impact of venting, microphone port orientation, and compression on the electroacoustically measured directivity of directional and omnidirectional behind-the-ear hearing aids. In addition, the average directivity provided across three brands of directional and omnidirectional behind-the-ear hearing aids was compared with that provided by the open ear. DESIGN: Three groups of hearing aids (four instruments in each group) representing three commercial models (a total of 12) were selected for electroacoustic evaluation of directivity. Polar directivity patterns were measured and directivity index was calculated across four different venting configurations, and for five different microphone port angles. All measurements were made for instruments in directional and omnidirectional modes. Single source traditional, and two-source modified front-to-back ratios were also measured with the hearing aids in linear and compression modes. RESULTS: The directivity provided by the open (Knowles Electronics Manikin for Acoustic Research) ear was superior to that of the omnidirectional hearing aids in this study. Although the directivity measured for directional hearing aids was significantly better than that of omnidirectional models, significant variability was measured both within and across the tested models both on average and at specific test frequencies. Both venting and microphone port orientation affected the measured directivity. Although compression reduced the magnitude of traditionally measured front-to-back ratios, no difference from linear amplification was noted using a modified methodology. CONCLUSIONS: The variation in the measured directivity both within and across the directional microphone hearing aid brands suggests that manufacturer's specification of directivity may not provide an accurate index of the actual performance of all individual instruments. The significant impact of venting and microphone port orientation on directivity indicate that these variables must be addressed when fitting directional hearing aids on hearing-impaired listeners. Modified front-to-back ratio results suggest that compression does not affect the directivity of hearing aids, if it is assumed that the signal of interest from one azimuth, and the competing signal from a different azimuth, occur at the same time. PMID- 10708074 TI - Customized acoustic transform functions and their accuracy at predicting real-ear hearing aid performance. AB - OBJECTIVE: The purpose of the study was to evaluate the validity of predicting the real-ear aided response by adding customized acoustic transform functions to the performance of a hearing aid in a 2-cc coupler. DESIGN: The real-ear hearing aid response, the real-ear-to-coupler difference (RECD/HA2), and field to behind the-ear microphone transfer functions were measured in both ears of 24 normally hearing subjects using probe-tube microphone equipment. The RECD/HA2 transform function was obtained using both insert earphones and with the hearing aid/ pressure comparison method. An RECD/HA2 transfer function was also obtained with a customized earmold, ER-3A foam tip, and an oto-admittance tip. RESULTS: Validity estimates were calculated as the difference between the derived and measured real-ear response. The derived response was generally within 5 dB of the measured real-ear response when it incorporated an RECD/HA2 transform function obtained with a customized earmold for the specific ear in question. Discrepancies increased when the RECD/HA2 transfer function was obtained from the same subject but the opposite ear. There were significant differences between the RECD/HA2 transform function obtained with customized and temporary earmolds. As a result, the derived response incorporating these transforms differed significantly from the measured real-ear response obtained with the customized earmold. The insert earphone and the hearing aid RECD/HA2 transfer function were equally valid. CONCLUSIONS: The derived response may be used as a substitute for in situ hearing aid response procedures when it incorporates acoustic transform functions obtained with a customized earmold from the specific ear in question. PMID- 10708075 TI - Cognitive factors and cochlear implants: some thoughts on perception, learning, and memory in speech perception. AB - Over the past few years, there has been increased interest in studying some of the cognitive factors that affect speech perception performance of cochlear implant patients. In this paper, I provide a brief theoretical overview of the fundamental assumptions of the information-processing approach to cognition and discuss the role of perception, learning, and memory in speech perception and spoken language processing. The information-processing framework provides researchers and clinicians with a new way to understand the time-course of perceptual and cognitive development and the relations between perception and production of spoken language. Directions for future research using this approach are discussed including the study of individual differences, predicting success with a cochlear implant from a set of cognitive measures of performance and developing new intervention strategies. PMID- 10708076 TI - Clinical and radiographic evaluation of the Harris-Galante cup: incidence of wear and osteolysis at 7 to 9 years follow-up. AB - A retrospective evaluation of the clinical and radiographic results of the Harris Galante acetabular cups was performed in 112 patients with 127 total hip arthroplasties. Patients with 14 hips had died, and patients with 20 hips were lost to follow-up. A total of 82 patients with 93 hips was available for follow up. There were 67 men and 45 women. The mean follow-up was 87 months (range, 48 113 months). There were a total of 9 revisions: 2 for recurrent dislocations, 1 for a loose cup, and 6 for wear and osteolysis. Radiographic evaluation demonstrated that 22 (24%) hips had periacetabular osteolysis, and 16 of these 22 (73%) were associated with the screws. Twenty-two hips (23%) demonstrated osteolytic lesions around the femoral stem. Mean femoral head displacement was of 1.00 mm (range, 0.40-4.5 mm) with a rate of 0.16 mm/yr (range, 0.05-0.44 mm/yr). The Harris-Galante socket has maintained a low implant failure rate at intermediate term, even in these young patients. As follow-up increases, wear of the polyethylene and periprosthetic osteolysis may become growing concerns. PMID- 10708077 TI - Primary total hip replacement with a noncemented acetabular component: minimum 5 year clinical follow-up. AB - The results of 123 total hip replacements with a noncemented Harris-Galante I acetabular component were reviewed (minimum 5-year clinical follow-up). The average clinical follow-up was 7 years (range, 5-10.8 years). No acetabular components were revised for loosening. One cup was revised for recurrent dislocation. In 3 cases, the acetabular liner was replaced at the time of femoral component revision (aseptic loosening), and in 1 case, a liner was revised because of recurrent dislocations. Of the hips, 92 had a complete set of radiographs for analysis. None of the acetabular components had migrated. Of the 92 acetabular components, 90 were considered radiographically stable (98%). Of these hips, 24 had linear radiolucencies of < or =2 mm in < or =2 of 4 zones and were considered stable. Two cups (2%) were considered possibly unstable. One of these had a linear radiolucent line in 3 zones, and the other had an osteolytic lesion measuring 6 x 11 mm in greatest dimensions. No acetabular components were definitely unstable. The average Harris Hip Score improved from 50 points (range, 17-89 points) preoperatively to 95 points (range, 74-99 points) at the latest follow-up examination. The average Hospital for Special Surgery hip score improved from 21 points (range, 10-31 points) preoperatively to 38 points (range, 27-40) at the last follow-up examination. Noncemented acetabular fixation with the Harris-Galante I component showed excellent clinical results at a minimum of 5 years' follow-up. PMID- 10708078 TI - Evaluation of the safety and efficacy of enoxaparin and warfarin for prevention of deep vein thrombosis after total knee arthroplasty. AB - Of 263 patients who underwent total knee arthroplasty, 122 received adjusted low dose warfarin and 141 received enoxaparin as deep vein thrombosis (DVT) prophylaxis. Three patients in the warfarin group and 3 in the enoxaparin group developed ultrasound-detectable DVT (P > .05). Although the amount of perioperative blood transfused was equivalent in both groups, the overall hemoglobin drop was greater (P < .005) in the enoxaparin group (2.9 g/dL) as compared with the warfarin group (2.3 g/dL). Five patients (4.6%) in the warfarin group and 16 (11.3%) in the enoxaparin group had bleeding complications (P < .05). Our data support earlier published reports suggesting that reductions, if any, in the incidence of DVT associated with enoxaparin are offset by a significant increase in bleeding complications as compared with adjusted-dose warfarin. We continue to use adjusted-dose warfarin as primary thromboembolic prophylaxis after total knee arthroplasty. PMID- 10708079 TI - Collarless polished tapered impaction grafting of the femur during revision total hip arthroplasty: pitfalls of the surgical technique and follow-up in 31 cases. AB - Impacting morcellized allograft bone into the femur during revision total hip arthroplasty is a simple concept with the goal of rebuilding femoral bone stock and providing secure fixation to the femoral stem. Using the collarless polished tapered (CPT, Zimmer, Warsaw, IN) stem impaction grafting system, we became concerned about the discrepancy between the straightforward concept and precise execution of the technique. In this study, we examined 31 consecutive procedures to determine intraoperative difficulties and report on the clinical outcome of 30 cases at an average follow-up of 31 months. Modified Harris Hip Scores averaged 41 points preoperatively and improved to 86 points at follow-up. Nineteen cases were performed on intact femora, whereas 12 cases had disrupted femoral integrity, either extended trochanteric osteotomy or periprosthetic fracture. Successful outcome was seen in all cases with an intact femur, and restoration of femoral integrity was key to successful outcome in cases with compromised femoral integrity. Among cases with disrupted femoral integrity, 3 distal fractures occurred as a result of the rigid CPT cement plug, and 2 complete femoral fractures occurred as a result of bone impaction, for a technique-related fracture rate of 16%. Difficulty packing bone distally occurred in 94% of cases and was associated with varus and valgus stem alignment and medial and lateral stem displacement. Complete cement mantles were seen in 77% of cases. No stem subsidence was seen in 15 of 30 cases (50%). Stem subsidence of <5 mm was seen in 10 of 30, stem subsidence of 6 to 8 mm was seen in 4 of 30, and stem subsidence of >10 mm was seen in 1 patient (4%). Of the patients, 87% thought the procedure improved their function, and 97% would recommend it to a friend with a failed femoral component. Although we hope that the instruments for this procedure can improve, we endorse the concept of impaction grafting with the CPT stem as a successful way of dealing with revision femoral surgery. PMID- 10708080 TI - Survivorship of AGC knee replacement in juvenile chronic arthritis: 13-year follow-up of 77 knees. AB - This study analyzed the survivorship and results of 77 knee replacements in 52 patients with juvenile chronic arthritis using the nonconstrained Anatomically Graduated Components (AGC; Biomet, Warsaw, IN) prosthesis design. Patients were operated on between the years 1985 and 1995. The mean duration of the general disease was 24 years (range, 10-56 years), and the mean age of the patients at the time of surgery was 33 years (range, 16-64 years). Bone-grafts were installed into 15 knees, custom-made components were used in 5 knees, and cemented fixation in 4 knees. The patella was resurfaced in 23 knees. Clinical follow-up examinations were conducted 3 months, 1 year, 4 years, and 8 years postoperatively. An interview was arranged at the end of 1998, 3 to 13 years after surgery; 2 patients were not reached, and 2 died during the follow-up. Fifty-five of 73 (75%) knees were subjectively excellent, 18 (25%) were fair, and none was poor. Radiolucent lines of 1.0 to 1.5 mm were found under 14 tibial trays but not adjacent to femoral components. No deep infections were detected. One knee was revised 4 years after the implantation. The overall survival was 99% (95% confidence interval, 92-100) at 5 years. We consider these results excellent in this demanding patient material. The nonconstrained AGC prosthesis with cementless fixation proved to be feasible in knee replacement in patients with juvenile chronic arthritis. PMID- 10708081 TI - Core decompression in atraumatic osteonecrosis of the hip. AB - Core decompression for osteonecrosis of the femoral head continues to be a controversial procedure. We report the results of core decompression in the treatment of hip osteonecrosis. Forty-two patients (67 hips) were evaluated. Minimum follow-up was 2 years. Preoperative outcome instruments were assessed. Volume of involvement (%) from magnetic resonance imaging was assessed. Failure was described as a total hip arthroplasty (THA). Mean patient age was 40.26 years. The average clinical and radiologic follow-up was 40.7 months and 33.1 months. The average Harris Hip Scores preoperatively and postoperatively were 49 and 58. None of the hips classified as Ficat I progressed to THA, whereas 17% of Ficat II hips and 66% of Ficat III hips progressed to THA. Our results demonstrate no relationship between the volume of involvement of the femoral head or the location of the lesion in progression to collapse. Staging with the Ficat classification demonstrated the most statistically significant correlation with progression to THA. The SF-36 scores at last follow-up on our patients were significantly worse than patients undergoing THA. PMID- 10708082 TI - Heterotopic ossification after total shoulder arthroplasty. AB - Fifty-eight primary ingrowth total shoulder arthroplasties, performed between 1989 and 1992, with a minimum of 2 years' radiographic and clinical follow-up (mean, 4.7 years), were reviewed to determine the frequency and clinical significance of heterotopic ossification after total shoulder arthroplasty. Fourteen of the 58 shoulders had radiographic evidence of heterotopic ossification: grade I (12 shoulders) and grade II (2 shoulders). Heterotopic ossification was present on the early postoperative radiographs (1-2 months) in 12 of the 14 shoulders. Among these 12 shoulders, there was no increase in the grade of ossification comparing the early postoperative radiographs with those obtained at a minimum of 2 years. There were no identifiable preoperative patient characteristics associated with the development of heterotopic ossification (P > .05). Range of motion, pain, and result rating were not statistically different comparing patients with and without heterotopic ossification (P > .05). The data from this study suggest that when heterotopic ossification develops after elective total shoulder arthroplasty, it is usually low grade, is present in the early postoperative period, is nonprogressive, and does not adversely affect the clinical results. PMID- 10708083 TI - Routine follow-up office visits after total joint replacement: do asymptomatic patients wish to comply? AB - A total of 100 patients presenting for routine office follow-up after total hip or knee arthroplasty completed questionnaires evaluating whether they preferred to come to the office for routine follow-up evaluation or whether they would have preferred an evaluation without an office visit. Of 100 patients, 45 would have preferred not to come into the office for a routine evaluation. They were content to mail completed questionnaires and radiographs to their physicians. The other 55 patients preferred office visits. These 2 groups were comparable for age, sex, height, weight, and number of surgeries (P > .11) Preoperative and postoperative scores were similar between the 2 groups (P > .39). None of the patients that would have preferred not to come in to the office believed that quality of care would be compromised. A significant number (45%) of patients would prefer not to come to the office because of the wages saved and time spared. Routine office visits may be eliminated for these patients through the use of health outcome devices, such as the SF-36, along with routine radiographs. The potential to decrease healthcare costs and increase patient satisfaction warrants the identification of these patients. Assessment of the effect on quality of care with elimination of routine follow-up visits requires further study. PMID- 10708084 TI - Hip arthroplasty with a collared straight cobalt-chrome femoral stem using second generation cementing technique: a 10-year-average follow-up study. AB - Clinical and radiographic results of 116 patients who had undergone 132 hip arthroplasties at our institution from 1983 to 1988 with a collared cemented straight cobalt-chrome femoral stem using second-generation cementing technique were reviewed. Twenty hips in 20 patients who were part of the original cohort were lost to follow-up. Mean age at the time of surgery was 68.2 years. Mean radiographic follow-up was 9.6 years with a minimum follow-up of 5 years. Ten year survivorship of the component was 96.5% with revision considered as an endpoint and 94.2% with either revision or radiographic loosening considered the endpoint. Three implants (2.3%) were revised for aseptic loosening at a mean of 8.1 years after implantation. One implant (0.8%) was revised for septic loosening at 10.5 years after surgery. Of the implants not revised, 1 showed evidence of circumferential bone-cement radiolucencies, and 1 had radiolucencies at the implant-cement interface. Five of the surviving femoral components (5.0%) showed focal areas of cystic osteolysis, and proximal femoral bone resorption under the collar was seen in 32 patients (31.7%). There were no cases of cement fracture or stem subsidence. The biomechanical and material properties of this stem combined with second-generation cementing technique look promising for long-term survivorship. PMID- 10708085 TI - The effect of posterior capsulorrhaphy in primary total hip arthroplasty: a prospective randomized study. AB - Between 1994 and 1997, 180 cases of primary total hip arthroplasty (THA) were performed with the posterior (Moore) approach for a variety of indications and studies prospectively. The cases were separated randomly into 2 groups to evaluate the effect of posterior capsulorrhaphy in the prevention of postoperative dislocation. In group 1 (96 cases), closure of the arthroplasty was performed with a posterior capsulorrhaphy; in group 2 (84 cases), closure was performed without capsulorrhaphy. The follow-up period was 38 months (range, 12 60 months). No dislocations occurred in group 1, whereas 2 dislocations (2.3%) occurred in group 2. Although the factors affecting dislocation in primary THA are many, a posterior capsulorrhaphy may be helpful in the prevention of posterior dislocation of primary THA performed with a posterior approach. PMID- 10708086 TI - The accuracy of assessing total hip arthroplasty outcomes: a prospective correlation study of walking ability and 2 validated measurement devices. AB - The Western Ontario and McMaster University Osteoarthritis Index (WOMAC) and the SF-36 are used to assess subjective outcome after total hip arthroplasty (THA). Although these indices have been validated, neither the WOMAC nor the SF-36 has been tested for accuracy against objective data in this clinical setting. Thirty osteoarthritic patients undergoing elective primary THA were subjectively evaluated preoperatively and 1 year postoperatively with the WOMAC and the SF-36 and objectively evaluated at the same interval with basic stride analysis and the 6-minute walk test. Correlation analysis of the subjective and objective data (both perioperative improvement and postoperative absolute scores) yielded Pearson coefficients of r = 0.50-0.81. This work demonstrates a sound statistical relationship between walking ability and the functional aspects of the WOMAC and the SF-36, supporting the use of these instruments in assessing the functional outcome after THA. PMID- 10708087 TI - Low-friction arthroplasty with Boneloc bone-cement: outcome at 2 to 4 years. AB - We report the clinical and radiologic outcome of 109 Chamley low-friction arthroplasties implanted with Boneloc bone-cement (Biomet, Bridgend, South Wales, UK) into 104 patients. The mean follow-up was 30 months (range, 2-48 months). There were 72 women (mean age, 71 years) and 32 men (mean age, 72). Cartridge packed cement was used in 37 cases and vacuum-packed cement in 72 cases. Survivorship analysis based on revision for aseptic loosening showed 79% survival at 4 years. Seventeen (15.5%) hips have been revised for aseptic loosening to date, in which all stems and 4 cups were loose. Extensive femoral osteolysis was always present and resulted in 4 cases of femoral cortical perforation at revision. Survivorship analysis based on revision and radiologic failure showed only 55% survival over the same period. When radiologic loosenings were included as failures, the vacuum-packed cement performed significantly worse than the cartridge-packed cement it replaced. These poor results were consistent with the withdrawal of Boneloc from clinical use in 1995, and we recommend indefinite follow-up for surviving prostheses. PMID- 10708088 TI - Charnley low-friction arthroplasty for Paget's disease of the hip. AB - This study investigated the perceived risks and complications associated with total hip arthroplasty for Paget's disease. A total of 98 Charnley low-friction arthroplasties were performed on 76 patients, 27 men (37 hips) and 49 women (61 hips), whose average age was 67.4 years (range, 51-79 years). Intraoperative blood loss averaged 388 mL (range, 110-1,730 mL), and minor heterotopic ossification occurred in 24 hips (25%), with significant changes in 4 hips (4%). After average follow-up of 10.4 years (range, 5.5-20 years), 10 acetabular (10%) and 8 femoral (8%) implants had loosened aseptically, but there was no evidence of progressive protrusio acetabuli or femoral deformity after operation. Survivorship to revision was 98% at 10 years (95% confidence interval [CI], 95% 100%) and 91% at 15 years (95% CI, 80%-100%) for the acetabular component and 93% (95% CI, 87%-99%) and 89% (95% CI, 80%-99%) for the femur. The only increased risk identified was nonunion of the trochanteric osteotomy (13%). PMID- 10708089 TI - The association of excessive warfarin anticoagulation and postoperative ileus after total joint replacement surgery. AB - Patients undergoing joint replacement who show signs of ileus in the postoperative period that require insertion of a nasogastric tube (NGT) must be monitored closely to avoid bleeding complications. The diagnosis of postoperative ileus was documented in 40 of 2,526 (1.6%) consecutive joint replacement operations between January 1, 1990, and March 1, 1998, at 1 hospital. Of the 40 patients with postoperative ileus, 34 received warfarin postoperatively. Of these 34 patients, 19 required a NGT for >48 hours, and 15 patients required a NGT for <48 hours or did not require a NGT. Of the 19 patients who required a NGT for >48 hours and who received warfarin anticoagulation, 17 had a prothrombin time of >20 seconds or an international normalized ratio (INR) of >2.0. None of the 15 patients who required a NGT for <48 hours and who received warfarin anticoagulation had a prothrombin time of >20 seconds or an INR of >2.0. This difference was highly statistically significant (P < .001). PMID- 10708091 TI - Potential errors in axial alignment using intramedullary instrumentation for total knee arthroplasty. AB - Accurate restoration of normal limb alignment is crucial for the long-term survivorship of total knee arthroplasty (TKA). A mathematical model was used to evaluate the maximum error in varus and valgus alignment that could occur when cutting the tibia or femur during TKA using intramedullary (IM) guides of varying length and diameter. Minor deviations in the insertion point of IM instrumentation during TKA can result in malalignment of several degrees. This error can be minimized by careful attention to the entry point of the IM instrumentation or by increasing the IM rod diameter and length used during primary TKA. PMID- 10708090 TI - Posterior slope of tibial plateau in Chinese. AB - The posterior slope of tibial plateau in 25 pairs of Chinese cadaveric tibia was studied. A digital photograph of the lateral profile of each cadaveric tibia was taken, and the posterior slope angles of medial and lateral plateau relative to the anterior tibial cortex were measured. The posterior slope of the medial plateau was 14.8 degrees, and the posterior slope of the lateral plateau was 11.8 degrees. Lateral radiographs of each cadaveric tibia were taken, and the posterior tibial slope angles relative to simulated extramedullary and intramedullary alignment were determined. With radiographic measurement, the posterior slope was 11.5 degrees using intramedullary method, and it was 14.7 degrees using extramedullary method. The posterior slopes increased with the presence of degenerative changes. PMID- 10708092 TI - A device for removal of femoral distal cement plug during hip revision arthroplasty: a high-powered drill equipped with a centralizer. AB - A removal procedure of a femoral cement mantle in hip revision arthroplasty has a risk of causing perforation or fracture, especially when removing a well-fixed distal cement plug. A high-powered drill equipped with a centralizer was developed to remove the distal cement plug safely. Using the drill equipped with a centralizer, the cement plug was removed well enough to insert a new component without causing perforation during the operation. PMID- 10708093 TI - Failure of a cemented all-polyethylene patellar component of a Press-Fit Condylar Total Knee arthroplasty. AB - A 65-year-old woman had a posterior cruciate-retaining cemented Press-Fit Condylar (PFC; Johnson & Johnson Professional, Inc., Raynham, MA) knee replacement with resurfacing of the patella using a cemented all-polyethylene, 3 peg component. Five years postoperatively, the patient developed anterior knee pain. The patella tracked normally, but radiographs showed a shift of the patellar component. At revision, there was shear failure of all 3 fixation pegs and loosening of the component. Revision with a cemented Press-Fit Condylar Sigma patellar 3-peg component (Johnson & Johnson Professional, Inc., Raynham, MA) was performed. PMID- 10708094 TI - Necrotizing fasciitis: a rare complication of total hip replacement. AB - Necrotizing fasciitis is a rare, but life-threatening emergency. We report a case history of necrotizing fasciitis in a previously healthy woman. A review of the literature suggests that this is the first report of necrotizing fasciitis complicating total hip replacement surgery. PMID- 10708095 TI - Silent compartment syndrome complicating total knee arthroplasty: continuous epidural anesthesia masked the pain. AB - Posterior dislocation is an uncommon complication of total knee arthroplasty (TKA) using a posterior stabilized total knee prosthesis, and it usually results from flexion instability. Acute posterior dislocation of a posterior stabilized prosthesis complicated by compartment syndrome of the leg has not previously been reported in the literature. We report a 62-year-old woman with posterior dislocation of her posterior stabilized TKA when her knee was in extension. It was further complicated by compartment syndrome with severe muscle necrosis. The diagnosis of compartment syndrome was delayed, partly because of continuous epidural anesthesia that completely abolished the pain and partly because of the low index of suspicion, as compartment syndrome is not well recognized as a possible complication of TKA. This case report strongly emphasizes that continuous epidural anesthesia is contraindicated in the case of complicated TKA because important clinical cues to neurovascular complications could be masked. PMID- 10708096 TI - Immunosuppressive-associated leukoencephalopathy in organ transplant recipients. AB - Immunosuppressive-associated leukoencephalopathy is a significant complication of cyclosporine (CsA) or tacrolimus therapy. However, the precise time of onset, role of putative risk factors, differences, if any, in presentation in various types of organ transplantation and outcome of this entity, remain poorly defined. Fifty cases of immunosuppressive-associated leukoencephalopathy reported in the literature in organ transplant recipients, were reviewed. Of 50 cases, 31 occurred in liver, 8 in renal, 6 in lung, and 5 in heart transplant recipients. Median time to onset was 28 days (range 3-1512 days); 82% occurred within 90 days of transplantation. Lesions tended to occur earlier in the liver transplant recipients, compared with other organ transplant recipients (median 9 vs. 29 days, P=.19). Seizures 74%, altered mental status 50%, and visual abnormalities 28% were the most frequently presenting features. Ten percent of the patients had fever with no documented source of infection. Systemic hypertension (P=.001), and lesions in the presence of therapeutic drug levels (P=.11) were more likely to occur with CsA than tacrolimus. Neuroimaging and clinical abnormalities were reversible on cessation or reduction of CsA or tacrolimus in all but two cases. Resolution of neurologic signs/symptoms occurred a median of 4 days and neuroimaging abnormalities in a median of 20 days on reduction/cessation of the drug. Immunosuppressive-associated leukoencephalopathy is a unique entity that can usually be diagnosed on the basis of its distinctive time of onset, and clinical and neuroimaging characteristics, and it is potentially reversible if promptly diagnosed. Despite identical clinical presentation of this syndrome in the recipients of CsA and tacrolimus, above noted variations in risk factors suggest that a difference in pathophysiologic mechanism may exist. PMID- 10708097 TI - Carotid atheroma: intransigent plaque or suppressable inflammatory pathology? New thoughts from a study of Australian SPK recipients. PMID- 10708098 TI - Expression of tissue factor mRNA in cardiac xenografts: clues to the pathogenesis of acute vascular rejection. AB - BACKGROUND: Acute vascular rejection destroys vascularized xenografts over a period of hours to days and is now considered the major hurdle to the clinical application of xenotransplantation. The hallmark of acute vascular rejection is diffuse intravascular coagulation; however, the pathogenesis of coagulation is a matter of controversy. One line of evidence points to activated endothelial cells and another to activated inflammatory cells as a source of tissue factor and thus as a primary cause of this lesion. The distinction between the two mechanisms inducing coagulation in the xenograft provides an opportunity for specific intervention. METHODS: To explore these mechanisms, we studied the expression of tissue factor mRNA by in situ reverse transcriptase-polymerase chain reaction in relation to the histopathologic manifestations of acute vascular rejection in guinea pig hearts transplanted into rats treated by cobra venom factor to avoid the hyperacute rejection. RESULTS: Three hours after transplantation and before the deposition of fibrin, tissue factor mRNA was expressed in the endothelial cells lining small and medium blood vessels and in smooth muscle cells of guinea pig cardiac xenografts. Sixteen hours after transplantation, while rat tissue factor mRNA was expressed only in occasional infiltrating cells, cardiac xenografts showed prominent deposits of fibrin in small vessels. Maximum expression of tissue factor on rat infiltrating cells was observed 48 hr after transplantation. CONCLUSIONS: These results suggest that in acute vascular rejection, coagulation is initiated on the donor vascular system, while the procoagulant characteristics of infiltrating cells may reflect a response to tissue injury rather than a cause. PMID- 10708099 TI - CD4+ T cells are critical for corneal, but not skin, allograft rejection. AB - BACKGROUND: The relative contribution of CD4+ or CD8+ T cells in allograft rejection remains to be fully characterized. Some reports indicate that there is an absolute requirement for CD4+ T cells in allogeneic rejection, whereas others report that CD4-depleted mice are capable of rejecting certain types of allografts. METHODS: We compared the ability of CD4- knockout (KO), CD8- KO, and normal CD4+/CD8+ mice to reject allogeneic corneal or skin grafts. We also examined delayed-type hypersensitivity and CTL responses to donor alloantigens. RESULTS: Engraftment of C57BL/6 corneas to C.B6-(n5-7) CD4-KO mice resulted in significantly higher rates of acceptance (>85%) than either C.B6-(n5-7) CD8- KO (30%) or normal BALB/c mice (40%). Likewise, mean survival times for B6 skin grafts placed on C.B6-(n5-7) CD4- KO mice (29.2 +/- 3.5 days) were significantly increased over those of normal BALB/c mice (13.2 +/- 1 days), although most CD4- KO mice (70%) eventually reject their grafts. C.B6-(n5-7) CD4- KO mice that reject allogeneic grafts fail to develop a delayed-type hypersensitivity response, but they did demonstrate significantly greater cytotoxic T lymphocyte precursor (CTLp) frequencies than did CD4- KO mice that accepted such grafts or that were not grafted. CONCLUSIONS: This study indicates that mice lacking CD4+ T cells have a significantly impaired ability to reject corneal allografts, but are able, in most cases, to reject allogeneic skin grafts. Thus, in the absence of CD4+ T cells, the likely mechanism for rejection appears to involve the generation of CD8+ CTLs. PMID- 10708100 TI - Prevention of acute allograft rejection in nonhuman primate lung transplant recipients: induction with chimeric anti-interleukin-2 receptor monoclonal antibody improves the tolerability and potentiates the immunosuppressive activity of a regimen using low doses of both microemulsion cyclosporine and 40-O-(2 hydroxyethyl)-rapamycin. AB - BACKGROUND: In previous studies of cynomolgus monkey lung allograft recipients, we demonstrated significant immunosuppressive efficacy but reduced tolerability after combined treatment with high doses of microemulsion cyclosporine (CsA) and SDZ RAD (40-O-(2-hydroxyethyl)-rapamycin). The current study was designed to compare efficacy and tolerability of a combination of low-dose CsA and high-dose SDZ RAD (CTL group) to triple therapy using the chimeric anti-interleukin-2 (IL 2) receptor (CD25) monoclonal antibody (mAb) basiliximab (anti-IL-2 receptor mAb) for induction therapy (basiliximab: 5 mg intravenously on days 0 and 4) plus low dose CsA and low-dose SDZ RAD for maintenance immunosuppression (CD25 group). CsA and anti-IL-2 receptor mAb are drugs that reduce cytokine synthesis and block IL 2-mediated lymphocyte stimulation, respectively. SDZ RAD blocks lymphocyte stimulation by other cytokines (e.g., IL-15) that are not inhibited by anti-IL-2 receptor mAb. METHODS: Twelve unilateral lung transplants were performed. Recipients were observed for 49 days by daily weight assessment, hemograms, blood chemistries, radiographs, and lung biopsies. Monkeys were euthanized before day 49 in the event of excessive weight loss (>25%) or organ failure. Target CsA trough levels were 100-200 ng/ml. Target SDZ RAD trough levels in the CTL group (no mAb) were 20-40 ng/ml, and 10-20 ng/ml in the CD25 group. RESULTS: None of the monkeys in the CD25 group needed to be euthanized early due to signs of drug toxicity. In contrast, four monkeys in the CTL group were sacrificed on days 28 35 as a result of excessive weight loss (n=3) and renal functional impairment (n=1). Three recipients in the CD25 group were euthanized on days 36, 38, and 46 as a result of persistent high fever associated with severe rejection. The median animal survival in the CTL group was 32 vs. 46 days in the CD25 group (P<0.04). The only two long-term survivors in the CTL group showed moderate rejection at day 49. The median rejection scores at day 14 (A0) and day 28 (A2) were identical in the two groups, despite the fact that the mean SDZ RAD trough level was significantly lower in the CD25 group (CTL: 38+/-3 ng/ml, CD25: 18+/-2 ng/ml, P<0.0001). After basiliximab levels fell below the minimum therapeutic level (1 mg/ml) on day 28, the median rejection score at day 49 increased to A4 in the CD25 group. CONCLUSION: This is the first study to combine an anti-IL-2 receptor mAb with a drug from the rapamycin class plus CsA. Our study shows that induction therapy with basiliximab enabled SDZ RAD blood levels to be significantly reduced, which led to improved tolerability without the penalty of increased rejection. PMID- 10708101 TI - SDZ-RAD prevents manifestation of chronic rejection in rat renal allografts. AB - BACKGROUND: Chronic rejection remains the most frequent cause of renal graft loss over the long term. However, effective treatment of this process is not yet available. SDZ-RAD (40-O-[2-hydroxyethyl]-rapamycin) is a new, orally active rapamycin derivative with potent immunosuppressive activity. We have examined the effects of SDZ-RAD in a well-established model of chronic renal allograft rejection in rats. METHODS: Kidneys of Fisher (F334) rats were orthotopically transplanted into bilaterally nephrectomized Lewis recipients. To suppress an initial episode of acute rejection, rats were briefly treated with low doses of cyclosporine for the first 10 days. Thereafter they received either SDZ-RAD (0.5 mg/kg(day) or vehicle. At 24 weeks, functional evaluations were performed, kidneys were harvested, and histological, immunohistological, and reverse transcription-polymerase chain reaction evaluations were performed. RESULTS: Animals treated with SDZ-RAD developed lower proteinuria and less glomerulosclerosis as compared with controls. Additionally SDZ-RAD reduced the infiltration of macrophages and lymphocytes and the expression of intercellular adhesion molecule-1, laminin, and fibronectin. Furthermore, we observed a reduced expression of growth factor mRNA (transforming growth factor-beta and platelet derived growth factor-AA) in these animals. CONCLUSION: Our results demonstrated that SDZ-RAD effectively ameliorates chronic renal allograft rejection in rats, probably mediated by suppression of growth factors. PMID- 10708102 TI - Ganciclovir-sensitive acute graft-versus-host disease in mice receiving herpes simplex virus-thymidine kinase-expressing donor T cells in a bone marrow transplantation setting. AB - BACKGROUND: The use of donor T cells expressing the herpes simplex thymidine kinase (HSV-TK) gene followed by ganciclovir (GCV) treatment could allow for specific modulation of the alloreactivity occurring after bone marrow transplantation. We are presently exploring such an approach in a phase I clinical trial. METHODS: To examine the beneficial effect of administrating HSV TK-expressing donor T lymphocytes +/- GCV treatment on acute graft-versus-host disease (aGVHD) control, irradiated Balb/c or C57BL/6 mice underwent transplantation with allogeneic bone marrow cells in conjunction with CD3+ allogeneic splenocytes from transgenic mice expressing an HSV-TK transgene. GCV treatment was initiated upon the occurrence of severe aGVHD. RESULTS: GCV treatment resulted in a 40-60% long-term survival rate of GVHD-free recipients having received HSV-TK-expressing T cells, whereas only 0-6% of mice survived without GCV treatment. Lethal aGVHD occurred in all the control animals having received non-HSV-TK-expressing T cells, irrespective of GCV treatment. CONCLUSION: Our results demonstrate that the administration of donor HSV-TK expressing T cells to hematopoietic stem cell graft recipients followed by GCV treatment at the onset of severe aGVHD significantly reduces aGVHD-induced mortality and results in GVHD-free surviving recipients. PMID- 10708103 TI - Peritoneal dialysis favorably influences early graft function after renal transplantation compared to hemodialysis. AB - BACKGROUND: Delayed graft function (DGF) and acute renal failure (ARF) after renal transplantation negatively influence short- and long-term graft outcome. Peritoneal dialysis as pretransplantation dialysis modality was reported to influence favorably the recovery of renal function immediately after kidney transplantation. It has been hypothesized that fluid status was the factor explaining this better outcome. This hypothesis was tested in this study by multivariate analysis, also including other factors related to DGF and ARF. METHODS: The records of peritoneal dialysis (PD; n=40) and hemodialysis (HD; n=79) patients receiving a first cadaveric kidney transplantation at the University Hospital Gent were analyzed. RESULTS: DGF and ARF were observed in 33 (27 HD and 6 PD, P=0.03) and 14 (14 HD and 0 PD, P=0.01) patients, respectively. The number of days needed to reach a serum creatinine 50% below that before transplantation (T1/2(SCr)), was correlated with cold ischemia time (CIT) (P<0.001) and body weight gain (BWG) (P<0.01) and was inversely correlated with urinary output in the first 24 hr (P<0.001), fluid load (P<0.001), and central venous pressure (P<0.001). A multivariate model with CIT (P<0.001), PD as pretransplantation dialysis mode (P=0.01), urinary output in the first 24 hr (P=0.001), BWG (P=0.05), and fluid load (P=0.01) resulted in an R2 of 0.32 (P<0.001). Using Cox regression analysis, the relative risk for a prolonged T1/2(SCr) increased with 4%/hr CIT (P=0.01) and with 1%/kg BWG (P=0.02). Fluid load decreased the relative risk with 5%/liter (P<0.001) and PD as pretransplantation modality favorably modified the relative risk by a factor of 1.6 (P=0.01). CONCLUSION: PD as pretransplantation dialysis modality can reduce the incidence and the severity of delayed recovery of renal function after renal transplantation. This protective effect was independent from CIT, and fluid status, two other major influencing factors. PMID- 10708104 TI - Analysis of the French liver transplant waiting list, 1992-1996. AB - BACKGROUND: In organ transplantation, each country has specific rules of allocation. We have retrospectively evaluated the French liver transplant waiting list for the period 1992-1996, during which time the rules for allocation remained stable. Mortality while on the waiting list and waiting times were the two principal endpoints. METHODS: Using the Etablissement Francais des Greffes (EFG) registry, the study was conducted in three steps: (1) description of the waiting list population; (2) analysis of the outcome of patients, with the use of a logistic model to explain death while on the waiting list; (3) estimation of waiting times and use of a Cox model to explain waiting times. RESULTS: The distribution of patients with regard to degree of urgency, age, blood type, and liver disease was variable according to the the EFG region. The outcome of patients was variable according to blood type and to EFG region. Patients classed as "extremely urgent" did not have a different outcome compared to elective patients. The logistic model indicated that two factors influenced the death: blood group and EFG region. Waiting times were variable according to EFG region; age and blood group had an influence on waiting times for a graft. The Cox model indicated the independent influence of EFG region on waiting times. CONCLUSION: We found geographical disparities between patients with respect to time on the waiting list. However, the database must be improved by including the risk profile of each patient, leading to changes in rules for a better allocation of transplants. PMID- 10708105 TI - Risk factors in the development of cyclosporine-induced gingival overgrowth. AB - BACKGROUND: Severe gingival hyperplasia (GH) is one of the most frequent side effects associated with the prescription of cyclosporine-A (CsA). Using the largest group of renal allograft recipients assembled for this purpose, in this study, we statistically modeled the genetic (HLA), medical, and dental risk factors for the development of GH subsequent to administration of CsA. METHODS: Two hundred thirty-six renal transplant patients underwent full dental examination to quantify the extent and distribution of hyperplasia and dental disease (gingivitis, plaque, and calculus). Computerized data from all patients included pre-transplant medical history and dosage of nifedipine and azathioprine, as well as dose and serum levels of CsA and CsA microemulsion. Donor and host HLA haplotype were studied to investigate potential association of haplotype and donor-host mismatching with the development of GH. We evaluated the data by multivariate regression analysis, using Statistical Package for Social Sciences (SPSS). RESULTS: There was no association with age, sex, duration of renal replacement therapy, or interval since transplantation or pre-transplant disease (P>0.05). There also was no association of disease with host HLA haplotype, but degree of HLA-A mismatching was protective for GH development (P<0.002). GH was associated with the dose and serum levels of CsA (P<0.001) and the last dose of CsA microemulsion (P=0.009) but not nifedipine (P=0.10). Gingival inflammation and plaque were also strongly associated with GH (P<0.0003). In multivariate analysis, however, the last recorded dose of CsA (P<0.0001), presence of local gingival inflammation (P<0.0001), and gingivitis (P<0.003) were the independent predictors of the extent and severity of GH. CONCLUSIONS: Inter-patient variation in the extent and severity of GH is related to CsA dose and serum levels. Differences in host HLA phenotype do not explain individual susceptibility to GH, but donor-host HLA-A mismatching may be important. Inter-site variation in the extent and severity of the disease is related to local gingival inflammation. PMID- 10708106 TI - Recombinant human granulocyte colony-stimulating factor after kidney transplantation: a retrospective analysis to evaluate the benefit or risk of immunostimulation. AB - BACKGROUND: Leukopenia due to immunosuppressive drugs represents a well-known complication in graft recipients, which might put patients at an increased risk for infections. In this study, recombinant human granulocyte colony-stimulating factor (rhG-CSF), a hematopoietic growth factor that selectively stimulates neutrophil colony formation and neutrophil cell differentiation, was tested for safety and efficacy. METHODS: We evaluated 30 episodes of leukopenia (<2000/mm3) in 19 kidney graft recipients treated with rhG-CSF. This cohort was compared with an age- and sex-matched historical control group without therapy. Peripheral and differential blood cell counts were analyzed, and the duration of leukopenia was estimated. Furthermore, the occurrence of infections associated with leukopenia was investigated. RESULTS: All patients responded to rhG-CSF therapy. Peripheral leukocyte counts increased from 1756+/-582 to a peak of 8723+/-3038/mm3 (P<0.0001). On the average, the peak was reached after 2.7 days (range 1 to 8). Furthermore, the effect was fairly persistent, because in 22 of 30 episodes leukocyte counts were within the normal range after 7 days. The elevation of total leukocytes was mainly due to a specific increase in neutrophil granulocytes from 1143+/-514 to 6895+/-1950/mm3 on the peak day (P<0.0001). Patients in the G CSF group were leukopenic for a mean of 1.29+/-0.59 days, whereas in the control group leukopenia persisted for at least 7 days. Consequently, the rate of infections was significantly higher (P<0.045) in nontreated patients. CONCLUSION: rhG-CSF was safe and effective in leukopenic kidney graft recipients. Leukopenic episodes in treated patients were significantly shorter, and infections occurred at a significantly lower rate. No evidence was found that rhG-CSF therapy might trigger rejection episodes, and no side effects were observed. PMID- 10708107 TI - Chylomicron metabolism in patients submitted to cardiac transplantation. AB - BACKGROUND: Development of coronary graft disease is the most important cause of late heart graft failure. Alterations in plasma lipid profile are frequent in heart transplant (HT) patients, but they seem not to be prominent. Currently, the metabolism of chylomicrons, the lipoproteins that carry dietary lipids absorbed by the intestine, was evaluated because chylomicron remnants are considered atherogenic. METHODS: An emulsion labeled with 3H-triolein and 14C-cholesteryl oleate and known to mimic the metabolic behavior of chylomicrons was injected intravenously after a 12-hr fast into 34 HT patients, 24 patients with end-stage heart failure (ESHF), and 30 healthy normolipidemic subjects. The plasma disappearance curves of the radioisotopes were determined from blood samples collected over 1 hr. In some of the patients and in controls, in vitro postheparin lipolytic activity was measured and an oral fat load test with postprandial measurement of triglyceridemia was performed. RESULTS: Fractional clearance rate (in m(-1), median [25%; 75%]) of both emulsion 3H-triolein and 14C cholesteryl oleate was extremely diminished in HT patients (HT: 0.0114 [0.0114; 0.0179] and 0.2x10(-8) [0.2x10(-8); 0.0041, respectively]; ESHF: 0.0226 [0.0223; 0.0568] and 0.0160 [0.0055; 0.0189]; control subjects: 0.0270 [0.0226; 0.0392] and 0.0090 [0.0042; 0.0180], respectively, P<0.05). HT patients also had reduced postheparin lipolysis and marked elevation of postprandial triglyceridemia compared with the controls. CONCLUSIONS: HT patients develop accumulation in the plasma of chylomicrons and their remnants. The observed alterations were so intense that they may suggest an important involvement of atherogenic chylomicron remnants in coronary graft disease. PMID- 10708108 TI - Chimerism in peripheral blood of sensitized patients waiting for renal transplantation: clinical implications. AB - BACKGROUND: Potential renal transplant recipients with preformed antibodies to HLA resulting from previous transplants, pregnancy, and/or transfusions are unlikely to receive an allograft. The factors contributing to the long-term maintenance of antibody titers in these individuals are still unknown. In the present study, we sought to determine whether chimerism in the blood correlates with maintenance of HLA sensitization in highly sensitized patients. METHODS: Qualitative analysis of chimerism in blood of sensitized patients was assessed by polymerase chain reaction-sequence specific oligonucleotide probes (PCR-SSOP) to HLA-DR. PCR single-strand conformation polymorphism (PCR-SSCP) was used to confirm the extra HLA-DR antigens detected by PCR-SSOP. RESULTS: Fourteen of 36 patients (38.9%) were positive for more than two HLA-DR indicative of chimerism. The presence of extra HLA-DR was confirmed by PCR-SSCP. When patients were analyzed on the basis of their panel-reactive antibody (PRA) status, 10 of 15 (66.7%) were positive for chimerism in the sensitized group, compared with only two of eight (25%) in the unsensitized group. Of the five males in the sensitized group who had received a blood transfusion but not a transplant, three were positive for chimerism. An association was observed between chimerism and maintenance of sensitization. None of the eight normal subjects studied demonstrated chimerism. CONCLUSIONS: The results obtained with sensitized patients suggest an association between blood chimerism and maintenance of HLA sensitization. We speculate that chimerism may lead to long-term maintenance of anti-HLA antibody titers. This finding implies that abolition of chimerism could result in the eventual elimination of antigenic stimuli for antibody production against HLA antigens. PMID- 10708109 TI - Accuracy and significance of computed tomographic scan assessment of hepatic volume in patients undergoing liver transplantation. AB - BACKGROUND: A small liver volume is considered to be a poor prognostic factor in cirrhosis, often indicative of advanced liver disease. Radiologic assessment of liver volume before liver transplant is routinely performed in many transplant centers. We sought to assess the accuracy and significance of computed tomographic (CT) scanning in hepatic volumetric analysis by correlating CT derived estimation of liver volume with that of corresponding liver explants. METHODS: A chart review of all patients aged 17 years or older undergoing liver transplant at Mount Sinai Medical Center between 1989 and 1995 was performed. Each patient underwent conventional CT scanning with measurement of liver volume (CTLV). Recipient liver volume (RLV) was defined as weight of liver explant after all attached ligaments, portal structures, and gallbladder were dissected free. Expected liver volume was calculated pretransplant based on age, gender, height, and weight. Patients were categorized into three groups based on etiology of liver disease: (1) hepatocellular (e.g., viral hepatitis, alcohol-related), (2) cholestatic (e.g., primary biliary cirrhosis), and (3) cryptogenic. The ratio of CTLV to RLV was used as a measure of the accuracy of CT volumetric analysis. RESULTS: A total of 579 patients was studied (group 1=376, group 2=139, group 3=64). All three groups were statistically similar with regard to age, prothrombin time and total bilirubin. Median CT liver volume was 1308 ml (range: 338-3847), 1651 ml (range: 641-3861), and 1210 ml (range: 348-2575) in groups 1 3, respectively; median ratio of CTLV to RLV was 1.02 (range: 0.50-2.31), 1.05 (range: 0.52-2.22), and 1.05 (range: 0.50-1.56) for groups 1-3, respectively. When RLV was small, it tended to be overestimated by CTLV. In contrast, when RLV was large, it was often underestimated. Clinical features such as history of esophageal variceal bleed, encephalopathy or ascites, and laboratory data did not influence accuracy of CT volumetric analysis. CONCLUSIONS: CT-derived estimation of liver volume appears to correlate closely with actual weight of liver explant regardless of the etiology of chronic liver disease. With extremes in CT volumetric analysis, actual liver volume tends to be under- or overestimated. For patients with end-stage liver disease, both CT-derived and actual liver volume are greater in cholestatic than in hepatocellular disorders. PMID- 10708110 TI - Long-term outcome of simultaneous kidney-pancreas transplantation: analysis of 61 patients with more than 5 years follow-up. AB - BACKGROUND: The long-term outcome of simultaneous kidney pancreas transplant recipients is not well established. METHODS: We retrospectively reviewed all patients who underwent simultaneous kidney-pancreas transplantation with bladder drainage at our center between January 1989 and December 1991. A total of 57 patients (93%) were alive with functioning grafts 1 year after transplantation and were followed for a minimum of 5 years. These patients formed the study group. RESULTS: Five-year actual patient, kidney and pancreas survival rates were 95%, 85%, and 88%, respectively. Fasting serum glucose fell from 198 mg/dL preoperatively to 94 mg/dL and remained stable thereafter. Glycohemoglobin levels decreased from 9.8% preoperatively to 4.8% 1 year after transplantation and remained normal thereafter. Kidney function remained good, with mean serum creatinine of 2.0 and creatinine clearance of 56 ml/min throughout the follow-up period. Hospital admissions decreased significantly with increasing time after transplantation from a mean of 1.2 admissions per patient in the 1st year to a mean of 0.2 admissions per patient 6 years after transplantation. Of the readmissions, 42% were for <48 hr and the most common reasons for readmission were infection, surgery, and dehydration. Mean systolic blood pressure decreased from 166 mm Hg before the transplant to 142 mm Hg 1 year after the transplant. CONCLUSIONS: Simultaneous kidney pancreas transplantation is a safe and effective method to treat advanced diabetic nephropathy and is associated with stable metabolic function, decreased cholesterol, improved hypertension control, improved rehabilitation over time, and little morbidity or mortality after the 1st year. PMID- 10708111 TI - En block combined reduced-liver and small bowel transplants: from large donors to small children. AB - BACKGROUND: The critical shortage of size-matched donor organs for infants and small children in need of combined liver and intestinal transplantation has lead to long waiting times and a high risk of dying before transplantation. Utilizing grafts from larger donors could alleviate this problem, but using larger composite grafts in small children has been challenging and unsuccessful in the past. METHODS: We conducted a pilot study for evaluating the results of transplanting into small recipients a composite graft (reduced-size liver and whole small bowel, including duodenum and pancreas head) procured from large donors. Liver size reduction was performed ex situ using the extrahilar approach, which leaves the liver hilum untouched. Straightforward implantation of the graft was performed by simple, two-step vascular anastomoses. The preservation of the donor duodenum in continuity with the combined graft avoided the need for biliary reconstruction, thus preserving maximal bowel length for gut continuity restoration in the recipient. RESULTS: Two children, weighing 7.6 and 9.8 kg, respectively, underwent transplantation of a composite graft procured from donors weighing 35 kg. Their waiting time (68 and 97 days, respectively) was shorter compared with our previous experience with conventional techniques. Both are currently alive and well, at home and on full enteral feeds, 15 and 11 months after transplantation, respectively. CONCLUSION: This new technique has extended the range of possible donors for small candidates waiting for combined grafts and was successful in two patients. It should be considered for small recipients in the future. PMID- 10708112 TI - Immunopathogenesis of hepatitis B virus recurrence after liver transplantation. AB - BACKGROUND AND AIMS: Hepatitis B virus (HBV) recurrence after orthotopic liver transplantation is associated with inflammatory graft changes, despite immunosuppression and donor/recipient HLA mismatch. We investigated whether immune mechanisms are involved in the pathogenesis of hepatitis B after liver transplantation. METHODS: The virus-specific T helper (Th) cell response, activation of Th1/Th2 subpopulations, donor/recipient HLA, and expression of tumor necrosis factor (TNF)-alpha/TNF receptors were determined in 28 patients who underwent transplantation for HBV-related cirrhosis (17 with HBV recurrence and 11 without recurrence) in comparison to 30 nontransplant patients with chronic hepatitis B. RESULTS: Orthotopic liver transplantation recipients with HBV recurrence showed significant hepatitis B core antigen-specific T-cell proliferation, comparable to nontransplant patients, which was not present in transplant recipients without recurrence. In addition, hepatic and serum interleukin (IL)-2, interferon-gamma, and TNF-alpha were enhanced, without changes in IL-4 and IL-10. Phenotypically, hepatic infiltrates in allografts with HBV recurrence were comprised of CD4+ lymphocytes and macrophages with a correlation between interferon-gamma- and TNF-alpha-producing cells and the degree of necroinflammatory activity. There was a marked up-regulation of both TNF-alpha receptors, significantly greater than in nontransplant patients. CONCLUSIONS: These findings suggest that despite immunosuppression, HLA class I independent immune mechanisms have a significant pathogenic role in liver damage associated with HBV recurrence after liver transplantation. PMID- 10708113 TI - Different time course of circulating adhesion molecules and hyaluran during hepatic allograft rejection. AB - BACKGROUND: Inducible adhesion molecules are involved in cell-mediated allograft rejection. In addition, the endothelium is the main target of this process. This study investigated, whether soluble (s) forms of vascular cell adhesion molecule 1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) are elevated during cellular rejection and whether hyaluran is a useful marker of endothelial function in liver transplantation. METHODS: Serum levels of sICAM-1, sVCAM-1, and hyaluran were determined in 24 controls and 27 hepatic transplant recipients. These patients were divided in two groups: group I, 14 patients without rejection; and group II, 13 patients with rejection. Samples were collected on day 1 and 7 after transplantation, on the day of liver biopsy, and after treatment of the rejection. RESULTS: We found a significant increase in sICAM-1 levels in the postoperative period in the rejection group compared with the non rejection group. It persisted significantly elevated until the diagnosis of rejection was made. In contrast, sVCAM-1 was only significantly elevated in the rejection group when diagnosis of rejection was evident. Hyaluran levels were also significantly elevated in the rejection group at diagnosis of rejection. We noticed a significant decline in sICAM-1, sVCAM-1, and hyaluran levels after successful treatment of rejection. In addition, we observed in the non-rejection group a stable lower levels of hyaluran during the entire postoperative period. CONCLUSIONS: The release of circulating adhesion molecules is a prominent feature coinciding with the first episode of hepatic rejection. Differential patterns of sICAM-1 and sVCAM-1 exist during rejection. In addition, hyaluran levels may be a sensitive marker of liver endothelial cell function in the postoperative period of liver transplantation. PMID- 10708114 TI - Progression of macrovascular disease after transplantation. AB - INTRODUCTION: Cardiovascular and cerebrovascular disease are major causes of morbidity and mortality after kidney transplantation. The aim of this longitudinal study was to examine the natural history of carotid plaque and to determine risk factors for the progression of vascular disease in uremic, type 1 diabetic patients who received a combined kidney and pancreas transplant. METHODS: Carotid artery (n=765) and lower limb vascular duplex scanning (n=656) were prospectively undertaken in 82 recipients before transplantation, at 6 months, and then at annual intervals for up to 10 years. Plaque in the internal carotid artery (ICA), external carotid artery, and common carotid artery was classified by type, location, extent, and degree of functional obstruction, and evaluated using multivariate analysis. RESULTS: Carotid plaque was present in 22.5% of patients at initial scanning, but increased to 56.6% by 7-10 years after transplantation, especially in the ICA and common carotid artery. Both the severity and extent of plaque increased, and plaque became more complex and heterogeneous with time after transplantation (P<0.001). Carotid plaque was associated with older age, current cigarette smoking, hyperphosphatemia, hypoalbuminemia, duration of pretransplantation dialysis, and presence of lower limb plaque (P<0.05-0.001). The severity of carotid plaque increased in older, hypertensive recipients and was associated with metabolic acidosis and hyperphosphatemia (all P<0.05). Severity of ICA disease correlated with disease in the contralateral ICA (r=0.57, P<0.001) and femoral arteries (r=0.42, P<0.001). Paradoxically, each carotid artery progressed independently of the other. ICA disease severity progressed when heterogenous, calcified, or new plaque was present on scanning, and with reduced renal transplant function (P<0.01-0.001). The mean ICA blood flow remained stable with time but was progressively impaired by hypertension, fasting hyperglycemia, and a lower prednisolone dose (P<0.05). Cerebrovascular events occurred in only four patients and were unrelated to carotid disease, implying relative plaque stability. CONCLUSION: Extensive carotid vascular wall abnormalities increased significantly despite kidney and pancreas transplantation. Initiation of plaque was associated with systemic factors, whereas progression of established plaque was largely influenced by local factors within the arterial wall. PMID- 10708115 TI - Effects of cold and warm ischemia on the mitochondrial oxidative phosphorylation of swine lung. AB - BACKGROUND: The aim of the study was to investigate the consequence of warm and cold ischemia on lung mitochondria in order to define bioenergetic limits within lung could be suitable for pulmonary transplantation. METHODS: Twenty-two pigs underwent lung harvesting after lung flush with Euro-Collins solution. Mitochondria were isolated from fresh lungs, from lungs submitted to 24 or 48 hr of cold ischemia, to 30 or 45 min of warm ischemia, and to 30 min of warm ischemia followed by 24 or 48 hr of cold ischemia. Mitochondrial oxidative phosphorylation parameters were determined in isolated mitochondria by in vitro measurement of oxygen consumption. RESULTS: Relative to controls, mitochondria submitted to cold ischemia showed an alteration in the oxidoreductase activities of the respiratory chain but no membrane permeability alteration. After 48 hr of cold ischemia, there was a decrease in the yield of the oxidative phosphorylation. Thirty minutes of warm ischemia did not alter the mitochondrial respiratory parameters. However, lung submitted to 45 min of warm ischemia showed mitochondrial damage as a decrease in the oxidative phosphorylation efficiency and ADP availability but no change in the oxidoreductase activities. Relative to cold ischemia alone, 30 min of warm ischemia preceding cold ischemia promoted no significant change in the respiratory parameters. CONCLUSIONS: On bioenergetic basis, lung submitted to warm ischemia could be suitable for transplantation if the warm ischemia duration does not exceed 30 min. This could be a major concern in lung procurement from non-heart beating donors. PMID- 10708117 TI - Activation of immediate early gene, c-fos, and c-jun in the rat small intestine after ischemia/reperfusion. AB - BACKGROUND: Activated immediate early genes (IEGs) play key roles in mediating cellular response after ischemia/reperfusion (I/R) injuries in some organs such as liver, heart and kidney. However, there is no report investigating an association between the activation of IEGs and cellular regeneration or programmed cell death after I/R in small intestine. METHODS: We examined a sequential expression of c-fos and c-jun after I/R in rat small intestine using reverse transcription-polymerase chain reaction and Northern blot analysis, and compared the patterns with coexistent two parameters: (1) regeneration determined by immunohistochemical detection of proliferating cell nuclear antigen, (2) programmed cell death determined with the terminal deoxynucleotidyltransferase mediated dUTP-biotin nick end-labeling (TUNEL) method and DNA fragmentation. RESULTS: The expression of c-fos and c-jun mRNA increased markedly 15 min after reperfusion and was, respectively, 6.3 and 4.4 times higher than in controls. Proliferating cell nuclear antigen expression was significantly elevated between 5 min and 4 hr, peaking at 30 min after reperfusion. Apoptosis showed a peak 60 min after reperfusion. Apoptosis after I/R was detected in the nuclei of absorptive epithelial cells by the TUNEL method, and these apoptotic signals were consistent with the expression of c-Fos and c-Jun proteins using an immunohistochemical method. CONCLUSIONS: These results suggest that overexpression of c-fos and c-jun after I/R in the small intestine correlates with programmed cell death and subsequent cellular regeneration. PMID- 10708116 TI - In vitro analysis of verapamil-induced immunosuppression: potent inhibition of T cell motility and lymphocytic transmigration through allogeneic endothelial cells. AB - BACKGROUND: Cyclosporine A (CsA) and tacrolimus prevent proliferation but not transendothelial migration of alloreactive lymphocytes into donor organs. As a result, serious adverse effects, such as nephrotoxicity and neurotoxicity, have been observed under CsA/tacrolimus therapy. The incorporation of new drugs with infiltration blocking properties might enhance the efficacy of the current immunosuppressive protocol, allowing lower CsA/tacrolimus dosage. Because Ca2+ plays a critical role in cell-cell interaction, the Ca2+-channel blocker verapamil might be a good cany. didate for supporting CsA/tacrolimus-based therapy. METHODS: A T-cell endothelial cell coculture model or immobilized immunoglobulin G globulin chimeras were employed to investigate how S- and R- verapamil interfere with the lymphocytic infiltration process. The expression and arrangement of membranous adhesion receptors and cytoskeletal F-actin filaments were analyzed by fluorometric method in the presence of. verapamil. RESULTS: Both verapamil enantiomers strongly inhibited lymphocyte infiltration. CD4+ and CD8+ T cells were influenced to a similar extent with regard to horizontal locomotion (CD4+=CD8+), but to a different extent with regard to adhesion and penetration (CD4+ > CD8+). Moreover, penetration was blocked to a higher extent than was adhesion. ID50-values were 31 microM (CD4+-adhesion) and 11 microM (CD4+ penetration). Verapamil reduced P-selectin expression on endothelial cells and effectively down-regulated binding of T-cells to immobilized P-selectin immunoglobulin G globulins (ID50=4.4 microM; CD4+). A verapamil-induced reduction of intracellular F-actin in T-lymphocytes was proven to be mainly responsible for diminished cell locomotion. CONCLUSIONS: The prevention of CD4+ T-cell penetration by verapamil might argue for its use as an adjunct to CsA/tacrolimus based immunosuppressive therapy. PMID- 10708118 TI - Identification and characterization of the antigen-specific subpopulation of alloreactive CD4+ T cells in vitro and in vivo. AB - We report the identification and characterization of the small subpopulation of alloantigen-specific T cells in vitro and in vivo. This subpopulation of T cells was distinguished by up-regulation of cell surface CD4 expression. These CD4high T cells were alloantigen specific in proliferation assays in vitro, and they expressed memory/activation markers, including CD44high and CD69high. Further studies demonstrated that these allospecific CD4high cells were also present (< or = 1% of CD4+ T cells) in vivo in BALB/c (H-2d) recipients of C57BL/6 (H-2b) skin allografts. CD4high T cells isolated from regional draining lymph nodes in these skin graft recipients reacted in a donor-specific fashion to C57BL/6 splenocyte stimulator cells in mixed lymphocyte culture. Adoptive transfer of CD4high, but not CD4normal T cells, just before skin engraftment in CD4 knockout mice, reconstituted rejection. The discovery that a small subpopulation of CD4high lymph node cells contained all of the alloantigen-specific T cells may allow study of tissue-specificity and subsequent alloantigen identification in transplantation. PMID- 10708119 TI - Immunobiology of xenogeneic cornea grafts in mouse eyes. I. Fate of xenogeneic cornea tissue grafts implanted in anterior chamber of mouse eyes. AB - BACKGROUND: The cornea is an immune privileged tissue that, when grafted orthotopically, forms the anterior surface of the immune privileged anterior chamber. We have recently reported that allogeneic cornea fragments implanted in the anterior chamber of mouse eyes resist immune rejection, although such graft fragments are rejected outside the eye. We wished to determine the extent to which xenogeneic cornea fragments placed in the eyes of normal mice are vulnerable to immune rejection. METHODS: Guinea pig corneas, deprived surgically of epithelium, were cut into fragments and inserted into the anterior chamber of eyes of BALB/c and severe combined immune deficient (SCID) mice, adjacent to the central cornea of the recipient. The fate of the grafts was assessed clinically by biomicroscopy and histologically for 8 weeks postimplantation. RESULTS: The majority of guinea pig cornea fragments devoid of epithelium came to rest with the raw stroma adjacent to recipient endothelium. These fragments remained clear for the 8-week observation interval in both BALB/c and severe combined immune deficient mice. Clear grafts displayed viable guinea pig keratocytes and endothelial cell layers for 4 weeks. The endothelium was then replaced by murine cells by 8 weeks. A minority of guinea pig cornea fragments were oriented with donor endothelium adjacent to recipient endothelium. Although these grafts in severe combined immune deficient eyes eventually acquired an endothelial layer that faces the anterior chamber and remained clear, similar fragments in BALB/c eyes became opaque, failed to acquire a proper lining of endothelium that faces the anterior chamber, and incited an inflammatory reaction in adjacent recipient cornea. CONCLUSIONS: Immune privilege is afforded to xenografts of guinea pig cornea placed as stromal: endothelial cell fragments in the anterior chamber of mouse eyes, but only if the surface of the fragments that faces the anterior chamber is promptly covered with corneal endothelium. The possible roles of corneal endothelium in promoting immune privilege of corneal xenografts are discussed. PMID- 10708120 TI - Immunobiology of xenogeneic cornea grafts in mouse eyes. II. Immunogenicity of xenogeneic cornea tissue grafts implanted in anterior chamber of mouse eyes. AB - BACKGROUND: Xenogeneic corneal fragments (guinea pig) are highly resistant to immune rejection in the anterior chamber of mouse eyes. Because guinea pigs and mice are discordant (i.e., mouse serum normally contains guinea pig reactive xenoantibodies), we wished to determine the extent to which xenogeneic corneal fragments placed intraocularly in normal and specifically sensitized mice activated xenoreactive T and B cells. METHODS: Guinea pig corneas, deprived surgically of epithelium, were cut into fragments and inserted into the anterior chamber of eyes of BALB/c mice, adjacent to the central cornea of the recipient. Antibody (immunoglobulin [Ig]M, IgG) and delayed type hypersensitivity (DTH) immune responses of recipient mice to guinea pig xenoantigens were assessed. The fate of xenogeneic cornea implants was assessed in mice immunized systemically to guinea pig antigens. RESULTS: Guinea pig spleen cells and corneal fragments implanted s.c. induced within 2 weeks of immunization both DTH and IgG antibodies to guinea pig xenoantigens. By contrast, xenogeneic corneal fragments implanted in the anterior chamber of mouse eyes evoked no change in recipient humoral immune status and induced mild guinea pig-specific DTH only after 5 weeks. Presensitization of mice to guinea pig xenoantigens failed to increase the proportion of grafts that were regarded as rejected, but the onset of rejection in failed grafts occurred earlier than in unsensitized recipients. Active systemic immunization of mice bearing intracameral guinea pig corneal fragments failed to curtail the grafts' survival. Guinea pig corneal fragments implanted in the anterior chamber of normal mice failed to induce anterior chamber-associated immune deviation. CONCLUSIONS: Xenogeneic corneal fragments implanted in the anterior chamber of eyes of normal mice display strikingly reduced immunogenicity, and an inability to induce anterior chamber-associated immune deviation. These properties are discussed in terms of the vulnerability of guinea pig corneal tissue to immune rejection within the eye. PMID- 10708121 TI - Alleviation of graft-versus-host disease after conditioning with cobalt protoporphyrin, an inducer of heme oxygenase-1. AB - BACKGROUND: Recently, we demonstrated that elevated expression of heme oxygenase 1 (HO-1 or Hsp-32) resulted in the modulation of several immune effector functions. Here we evaluated whether induction of HO-1 after administration of cobalt protoporphyrin (CoPP) can prevent the development of acute graft-versus host-disease (GVHD). METHODS: Acute GVHD was initiated by injection of unfractionated spleen cells from C57BL/6 into B6D2/F1 mice. RESULTS: Administration of CoPP resulted in increased survival: 85% of CoPP-treated animals survived for >100 days compared with only 29% of saline-treated control animals (P<0.05). In contrast, administration of ZnPP, a well-known inhibitor of HO, accelerated GVHD development. The protective effect of CoPP therapy seemed to be caused by immunomodulation of donor cells, because treatment of cell donors prevented development of acute GVHD in 80% of recipients compared with 0% in control animals. Spontaneous lymphocyte proliferation could be measured with splenocytes harvested from animals developing GVHD but not with splenocytes from recipients of CoPP-treated donor cells. CoPP-treatment had no effect on interleukin-2 or interleukin-4 synthesis but inhibited interferon-gamma production. Mice with active GVHD demonstrated a defective lympho-proliferative response to alloantigens or concanavalin A. However, spleen cells isolated from survivors (on day 100) responded normally. Flow cytometric analysis of splenic T cell populations revealed a severe reduction in recipient type (H-2b,d) cells in mice with active GVHD, whereas in protected mice the number of cells remained normal. CONCLUSION: The results from this study confirmed our previous observation that up-regulation of HO-1 activity is associated with down regulation of several immune effector functions. This resulted in protection from acute GVHD in a parent into F1 mouse model. PMID- 10708122 TI - Natural killer cell and alphabeta and gammadelta lymphocyte traffic into the liver graft immediately after liver transplantation. AB - BACKGROUND: The persistence and migration of donor leukocytes has been well established, but cellular kinetics immediately after revascularization and the potential relevance of these different lymphocyte populations to spontaneous tolerance remain unclear. During the early hours of revascularization, there is a transitory "congestion" of the liver graft, which is evidence of an early phase that we have termed "first cellular contact." METHODS: We have carried out by flow cytometry a prospective comparative study of the peak kinetics of lymphocyte subpopulations contained in: (a) peripheral blood and liver grafts at the time of multi-organ extraction from 14 brain-dead donors, (b) recipient peripheral blood before transplantation, and (c) recipient peripheral blood and liver grafts after (t=2 h) declamping and vascularization of the liver graft. RESULTS: Before transplantation, the liver grafts contained large numbers of natural killer (NK) and NK-like cells with early lymphocyte activation. Immediately after revascularization, there was an influx of recipient NK and NK-like cells into the liver. CONCLUSIONS: NK and CD3+CD56+ (NK-like) cells flooding into the liver graft immediately after revascularization could rapidly destroy allogeneic cells. However, spontaneous tolerance and the persistence of donor lymphocytes after orthotopic liver transplant could be a result of donor TCRalphabeta NK1.1 liver graft lymphocytes, which may be involved in the destruction of CD8+ T lymphocytes that would have received the apoptosis signal, and to NK and NK-like cell inhibition via inhibitory NK receptors. The decrease in gammadelta T lymphocytes in the two compartments suggests a mechanism of recirculation and capture in other lymphoid organs. PMID- 10708124 TI - Cyclosporine facilitates chimeric and inhibits nonchimeric tolerance after posttransplant total lymphoid irradiation. AB - BACKGROUND: Previous studies showed that Lewis rats given posttransplant total lymphoid irradiation, antithymocyte globulin, and a single infusion of ACI peripheral blood or bone marrow cells develop tolerance to ACI heart allografts. METHODS: To determine the effects of cyclosporine on these tolerance induction protocols, groups of Lewis hosts, given either ACI blood or marrow infusions, were given a 60-day course of daily cyclosporine immediately after the cell infusion. RESULTS: Cyclosporine treatment was associated with uniform graft rejection in the groups given an ACI blood transfusion, and was associated with uniform graft acceptance in the groups given an ACI bone marrow infusion. Studies of donor-type T and B cell chimerism in the host blood showed that cyclosporine facilitated chimerism in the hosts given ACI bone marrow cells, and stable chimerism over a 300-day observation period was predicted by detectable chimerism by day 30. None of the hosts given ACI blood cells developed chimerism. CONCLUSION: Cyclosporine facilitated long-term graft acceptance in a tolerization protocol that induced mixed chimerism, but prevented long-term graft acceptance in a tolerization protocol that did not induce chimerism. PMID- 10708123 TI - Visualization of the in vivo generation of donor antigen-specific effector CD8+ T cells during mouse cardiac allograft rejection: in vivo effector CD8+ T cell generation during allograft rejection. AB - BACKGROUND: An adoptive transfer system was used to study the fate of alloreactive CD8+ H-2Kb-specific TCR transgenic (DES+) T cells in vivo after transplantation. METHODS: A trace population of 2.0x10(6) CD8+DES+ T cells were adoptively transferred into syngeneic CBA.Ca (H-2k) mice 24 hr before transplantation of an H-2Kb+ or H-2Kb- cardiac allograft. RESULTS: H-2Kb specific T cells proliferated and produced interleukin-2 and interferon-gamma in response to H-2Kb+, but not H-2Kb- cardiac allografts. CD8+DES+ T cells that infiltrated the H-2Kb+ cardiac allografts developed a distinct cell surface and cytokine phenotype compared with the CD8+DES+ T cells that remained in the periphery. H 2Kb-specific graft infiltrating T cells (a) underwent a larger number of cell divisions (> =3), (b) increased in size, (c) up-regulated CD69, and (d) down regulated CD62L. CONCLUSIONS: These results demonstrate that alloantigen-specific T cells can be monitored in vivo during the immune response to an allograft and that the fate of CD8+ T cells specific for the allogeneic class I molecules expressed by the graft is different between cells in the periphery and those that infiltrate the graft. PMID- 10708125 TI - Determination of acceptable HLA mismatches in highly sensitized patients by soluble HLA class I ELISA inhibition. AB - BACKGROUND: Acceptable HLA mismatches for highly sensitized patients are determined so as to increase their chances of receiving transplants. The disadvantage of the current procedures is that the antibody reactivity of the patients' sera is tested against HLA antigens expressed on cells or HLA antigens isolated from cell lysates. Therefore, two (homozygous for HLA-A and -B) to four (heterozygous for HLA-A and -B) different HLA class I antigens are present in the test. This might cause reactivity toward nonacceptable mismatches to mask the determination of acceptable mismatches. METHODS: Recently we observed that the detection of soluble HLA class I antigens is inhibited by HLA-specific antibodies. In the present study, inhibition of soluble HLA-specific ELISAs (anti soluble HLA-A2, -B7, -B12) was evaluated as a tool used to determine acceptable mismatches. The results were compared with current determination of acceptable mismatches (which is by complement-dependent cytotoxicity and/or fluorescence activated cell sorter analysis). RESULTS: In the case of acceptable mismatches determined by conventional methods, sera from the patients were not interfering in these ELISAs, whereas in the case of nonacceptable mismatches (thus specific antibodies), significant inhibition was observed in most instances. Among the nonacceptable mismatches, the test showed significant inhibition in 20 of 24 cases, whereas among acceptable mismatches, no inhibition was observed (in eight of eight), indicating the lack of specific antibodies. CONCLUSIONS: In highly sensitized patients, the introduction of soluble HLA-specific ELISAs is of additional and confirmatory value for the determination of acceptable mismatches. The major advantage of this approach is that antibody reactivity is tested against single antigens only. PMID- 10708126 TI - The role of respiratory epithelium in a rat model of obliterative airway disease. AB - BACKGROUND: The etiology and pathogenesis of obliterative bronchiolitis after lung transplantation remain to be fully elucidated. Using a rat model of heterotopically transplanted trachea grafts, we have examined the role airway epithelium plays in obliterative airway disease (OAD). METHODS: Rat trachea isografts were denuded of epithelium by protease digestion. Grafts were inoculated either with or without native airway epithelial cells and transplanted into the omentum of recipient animals. RESULTS: Airway epithelium removal resulted in OAD in denuded isogeneic trachea grafts. Reseeding of the denuded grafts with epithelial cells significantly reduced airway obliteration from 78% to 22% luminal occlusion. CONCLUSIONS: Non-immune-mediated injury will cause OAD, and epithelial cell replacement in denuded isografts can significantly reduce the fibrotic progression of the disease. PMID- 10708127 TI - Blocking of CD44-hyaluronic acid interaction prolongs rat allograft survival. AB - BACKGROUND: Lymphocyte activation and infiltration into a transplanted organ is an integral component of the rejection process. Graft infiltration of lymphocytes requires adhesion of leukocytes to the endothelium, diapedesis, and transmigration. One of several proteins involved in this process is CD44, which is known to interact with endothelial hyaluronan (HA). Blockade of cell-matrix and cell-cell interactions have been used extensively for modulation of immune responses and graft rejection. Based on these observations, we evaluated the effects of blocking CD44-HA interactions in a transplantation model. METHODS: We used a low molecular weight hyaluronic acid formulation (LMWHA) for the treatment of rat renal and cardiac allograft recipients. LMWHA was administered intraperitoneally at 0.5-5 mg/kg for 5-10 days after transplantation with or without a subtherapeutic dose of cyclosporine. RESULTS: LMWHA monotherapy prolonged allograft survival significantly, but only for a few days. In combination with low-dose cyclosporine, long-term survival of allografts was observed in some of recipients. CONCLUSION: Further definition of the underlying mechanism of LMWHA therapy may provide a rationale for the development of novel, nontoxic, nonimmunogenic immunotherapies. PMID- 10708128 TI - Primary posttransplant lymphoproliferative disorder of the gallbladder in a lung transplant patient presenting with acute cholecystitis. AB - BACKGROUND: Acute cholecystitis in an immunocompromised host is potentially devastating. Posttransplant lymphoproliferative disorder (PTLD) is a well described complication of immunosuppressive therapy used after solid organ transplantation; however, isolated involvement of the gallbladder has not been described. METHODS: Case report format is used. RESULTS: We report a case of PTLD isolated to the gallbladder, as well as histological evidence of acute cholecystitis, in a patient who presented with signs and symptoms of acute cholecystitis 1 year after single lung transplant. CONCLUSIONS: PTLD can occur in the setting of acute cholecystitis and may be missed if careful pathological examination is not undertaken. PMID- 10708129 TI - Cytomegalovirus infection may cause ureteral necrosis. AB - Cytomegalovirus (CMV) infection has protean presentation among immunocompromised patients, but the urinary tract is rarely involved. We report a case of extensive ureteral necrosis in a renal transplant, 12-year-old patient with typical histological feature of CMV inclusions. The role of CMV was confirmed by immunohistochemical analysis and concomitant CMV DNA detection in peripheral blood leukocytes by polymerase chain reaction analysis. CMV infection can, therefore, be regarded as a possible cause of ureteral necrosis in renal transplant recipients. PMID- 10708130 TI - Reversal of intrapulmonary arteriovenous shunting detected by two-dimensional contrast-enhanced echocardiography after liver transplantation. AB - BACKGROUND: Intrapulmonary arteriovenous shunting (IPS), occasionally associated with advanced liver disease, may reverse after liver transplantation (LTx). Two dimensional contrast-enhanced echocardiography, a convenient noninvasive study, has never been used to demonstrate disappearance of IPS after LTx. METHODS: For an 8-month-old girl undergoing living-related LTx, two-dimensional contrast enhanced echocardiography was performed with the microbubble injection. The opacification of the microbubble in the left heart emerging within 3-6 beats after detection in the right heart was compared with that in the right heart. RESULTS: Microbubble opacification in the left heart was almost the same as that in the right heart (grade 3) shortly after LTx. However, the contrast in the left heart diminished (grade 1) as the respiratory condition improved and subsequently disappeared (grade 0). CONCLUSIONS: Two-dimensional contrast-enhanced echocardiography may be a feasible noninvasive method to evaluate the degree of IPS in the peritransplant period and observe disappearance of IPS after LTx. PMID- 10708131 TI - Liver transplantation in a patient with acute liver failure due to sickle cell intrahepatic cholestasis. AB - BACKGROUND: Sickle cell intrahepatic cholestasis is a potentially catastrophic complication of sickle cell anemia Once acute liver failure develops, transplantation is the only option. We describe a patient with sickle cell intrahepatic cholestasis who underwent liver transplantation. METHODS: Data were obtained from the chart. Serial hemoglobin S levels were monitored, and measures were taken to maintain hemoglobin S <20% to prevent sickle cell crisis. RESULTS: Although the allograft functioned well initially, the patient developed veno occlusive disease and required repeat transplantation at 5 months after transplant. Histologic examination of the explant revealed occlusion of the terminal hepatic venules due to fibrosis and packed red cells. Repeat transplant was complicated by thrombosis of the intrahepatic portion of the hepatic artery, and sepsis. The patient died of sepsis after a third transplant. CONCLUSION: Liver transplantation for sickle cell disease involving the liver may carry a high risk of graft loss due to vascular problems. Repeat transplantation may not be feasible if disease recurs. PMID- 10708132 TI - Large de novo renal cell carcinoma in a 10-year-old transplanted kidney: successful organ-preserving therapy. AB - BACKGROUND: A recent review of the Cincinnati Transplant Tumor Registry recorded 24 de novo renal cell carcinomas developing in renal allografts. However, late development of these tumors after transplantation is very rare. Only four reports exist regarding conservative surgery on kidney transplant tumors. METHODS: This is a report on a case of a large 6-cm de novo renal cell carcinoma in a 10-year old transplanted kidney. Optimal therapy by transplant nephrectomy or tumor enucleation was discussed. RESULTS: Partial resections or enucleations of renal cell carcinoma are still less than ideal in carcinomas larger than 3 cm considering the higher risk of local recurrence. But the recipient in this case had done so well and had had such a high quality of life after transplantation that partial nephrectomy as therapy of choice was selected. Now the patient is 2 years tumor free. CONCLUSION: The case report demonstrates that in certain select cases of large tumors, organ-preserving surgery could be an alternative approach in combining complete tumor removal with preservation of graft function. PMID- 10708133 TI - Identification of alpha3, alpha4, and alpha5 chains of type IV collagen as alloantigens for Alport posttransplant anti-glomerular basement membrane antibodies. AB - BACKGROUND: Alport syndrome is a hereditary disorder of basement membranes especially affecting the kidneys, ears, and eyes. Some patients who undergo renal transplantation lose their kidneys as a result of posttransplant anti-glomerular basement membrane (anti-GBM) disease. METHODS: In the present study, we analyzed serum from 21 unselected Alport patients who underwent renal transplantation. Eleven samples were from patients without posttransplant anti-GBM nephritis, and 10 were from patients with this disease. RESULTS: Thirteen serum samples [10 alport posttransplant nephritis serum (APTN) and three Alport posttransplant serum (APT)] revealed linear binding to the GBM by indirect immunofluorescence. By using direct ELISA and immunoblotting with GBM constituents and type IV collagen NC1 domains from bovine, human, and recombinant sources, we detected anti-GBM antibodies in all Alport patients in varying titers. Five samples showed specific reactivity to the alpha3 chain, four to the alpha5 chain, six to both alpha3 and alpha5 chains, one to the alpha3 and alpha4 chains, and two to the alpha3, alpha4, and alpha5 chains of type IV collagen. The varied spectrum of reactivities was present equally in nephritic and non-nephritic sera. Ten control samples from non-Alport transplant patients did not exhibit specific binding to the GBM. CONCLUSIONS: These results suggest that the absence of alpha3, alpha4, and alpha5 chains of type IV collagen in the Alport kidney leads to alloantibodies in all Alport patients who receive transplants, irrespective of whether they develop nephritis or not. Although all Alport transplant patients develop this humoral response, only a select few develop anti-GBM disease. We suggest that this difference could be attributable to a genotypic effect on the ability of some individuals to launch a cell-mediated immune response. PMID- 10708134 TI - Renal allograft rejection: beta-chemokine involvement in the development of tubulitis. AB - BACKGROUND: Tubulitis is a defining feature of renal allograft rejection. Graft dysfunction may result from damage inflicted on tubular epithelial cells by intratubular cytotoxic T lymphocytes. Graft cells are known to produce chemokines during acute rejection, but it is not known whether changes in expression of specific chemokines can influence the composition of the intratubular lymphocyte population. We examined expression of individual chemokines in biopsy sections showing different pathological rejection grades. METHODS: Sections from Banff graded transplant biopsies were examined for the presence of beta-chemokines (MCP 1, MIP-1alpha, MIP-1beta, and RANTES) by immunofluorescence and semiquantitative confocal laser scanning microscopy. RESULTS: Beta-chemokines were expressed predominantly at the basolateral surface of tubular epithelial cells. Expression of MCP-1 and MIP-1beta was significantly higher in sections showing grade 2 rather than grade 1 acute rejection. RANTES and MIP-1alpha showed no significant variation in level of expression between rejection grades. CONCLUSIONS: Beta chemokines are expressed by tubular epithelial cells during acute rejection. Consistent expression of RANTES and MIP-1alpha suggests a general role in recruiting T lymphocytes. However, MCP-1 and MIP-1beta may play a more subtle role in recruitment of specific T-cell subsets, such as Th1 cells, during acute cellular rejection. PMID- 10708135 TI - Human cytomegalovirus blocks interferon-gamma stimulated up-regulation of major histocompatibility complex class I expression and the class I antigen processing machinery. AB - Interferon-gamma stimulates major histocompatibility complex (MHC) class I antigen processing and presentation by inducing the expression of major histocompatibility complex class I heavy chains, beta2-microglobulin, the transporter associated with antigen processing, and components of the proteasome complex. We demonstrate that this effect of interferon-gamma on the major histocompatibility complex class I pathway is inhibited in human cytomegalovirus infected fibroblasts and endothelial cells. This is the result of a direct human cytomegalovirus/cell interaction leading to a block in interferon-gamma signal transduction beginning at early times after infection and peaking at 72 hr after infection. These observations suggest a novel level of herpesvirus interference with antigen processing: protection of infected cells from the immunoregulatory effects of interferon-gamma. Thus protected, human cytomegalovirus persists and may exacerbate graft rejection or lead to fulminant infection in the immunocompromised transplant recipient. PMID- 10708136 TI - G-CSF modulates cytokine profile of dendritic cells and decreases acute graft versus-host disease through effects on the donor rather than the recipient. AB - Allogeneic peripheral blood stem cell transplantation (PBSCT) is increasingly used instead of bone marrow transplantation, particularly in HLA identical sibling pairs. Despite the presence of significantly increased numbers of T cells in the PBSC graft, acute graft-versus-host disease (GVHD) is not increased. We have investigated whether granulocyte-colony stimulating factor (G-CSF) administration to PBSCT recipients, both with and without donor G-CSF pretreatment, further modulates acute GVHD in a murine model of PBSCT. Recipients of G-CSF mobilized splenocytes showed a significantly improved survival (P<0.001) and a reduction in GVHD score and serum LPS levels compared with control recipients. G-CSF treatment of donors, rather than recipients, had the most significant effect on reducing levels of tumor necrosis factor (TNFalpha) 7 days after transplantation. As a potential mechanism of the reduction in TNFalpha, we demonstrate G-CSF decreased dendritic cells TNFalpha, and interleukin-12 production to lipopolysaccharide. In conclusion, G-CSF modulates GVHD predominantly by its effects on donor cells, reducing the production of TNFalpha. G-CSF treatment of bone marrow transplantation recipients, without pretreatment of the donor, does not have an impact on acute GVHD. PMID- 10708137 TI - Observations regarding the effect of targeted gene deletions (knockouts) on graft rejection. PMID- 10708138 TI - U-stitch ureteroneocystostomy: a new renal transplantation ureteral reimplantation technique associated with reduced urologic complications. AB - Urologic complications represent the most frequent complications following renal transplantation and are associated with significant morbidity. We present the results of the first 105 patients who underwent ureteroneocystostomy at our institution using a new surgical ureteral reimplantation technique designed to reduce the incidence of urologic complications after renal transplantation. PMID- 10708139 TI - Chordee repair utilizing a novel technique ensuring neurovascular bundle preservation. AB - Penile chordee, with and without hypospadias, is amenable to surgical correction. The Nesbit technique of dorsal plication of the ventral tunica albuginea is effective in correcting most cases of corporal disproportion. A hazard with this approach is the potential inclusion of the dorsal neurovascular bundle, with resultant erectile and sensory dysfunction. We developed a simple technique using the Freer elevator to isolate the neurovascular bundle prior to plication. This ensures that no injury occurs to the neurovascular bundle during plication. Since 1994, 37 boys with chordee have been repaired using this approach. Their ages at the time of operation ranged from 5 months to 28 years (mean 9 months). Following standard degloving of the penis, an incision through Buck's fascia is made lateral and parallel to the neurovascular bundle at the maximum level of the chordee. A similar incision is carried out on the contralateral side. A 4-mm-wide Freer elevator is positioned under Buck's fascia while hugging the tunica albuginea. The Freer elevator slides across the midline to the contralateral side, separating Buck's fascia and underlying layers from the tunica albuginea. Following isolation of the bundle, each corporal body is plicated by creating a longitudinal incision through the tunica albuginea, which then is closed transversely with a 5-0 polydioxanone suture. Buck's fascia subsequently is closed with an absorbable suture following confirmation of chordee correction. No complications have been encountered during a mean follow-up of 21 months (range 5 51 months). No patients have required reoperation for persistent chordee. We developed a technique that elevates the neurovascular bundle prior to plication, thereby ensuring no injury to this structure. We have successfully used this modified Nesbit technique since 1994 and have had no complications. Utilization of the Freer elevator adds an estimated 5 minutes to chordee correction compared to a standard plication lateral to the neurovascular bundles. Although long-term follow-up needs to be performed to confirm any erectile or sensory advantage, this approach should be considered whenever plication is to be performed. PMID- 10708140 TI - Laparoscopic radical nephrectomy: a single-center experience of 51 cases. AB - A modified technique of laparoscopic radical nephrectomy for treatment of renal cell carcinoma makes surgery easier, faster, and safer in terms of tumor cell spillage. We report our experience with this procedure in 51 consecutive cases. A transperitoneal approach was used in all cases. The average patient age was 62 years. The solid renal mass diameter was between 2 and 9 cm. Extrafascial mobilization of the kidney included limited lymph node dissection. In six patients the adrenal gland was removed simultaneously. The specimen was removed intact through a small muscle-splitting incision in the lower abdominal wall. The procedure was successful without conversion to open surgery in all 51 patients. The average operating time was 125 minutes, and the average postoperative hospital stay was 7.2 days. Major complications were seen in 4% of patients. Neither local recurrences nor metastases were observed in the following 7.9 (1 19) months. In our experience, laparoscopic radical nephrectomy is safe and efficient. Removing the specimen intact through a small muscle-splitting incision reduces operating time, avoids tumor cell spillage, and allows exact pathological staging. PMID- 10708141 TI - Retrograde ureteroscopic incision for the treatment of nonureteroenteric ureteral strictures. AB - A variety of methods are available for the management of patients with ureteral strictures. Ureteroscopic, retrograde incision using the holmium laser was performed on an outpatient basis or with hospitalization for <24 hours in three patients with strictures of varying etiologies. With follow-up of 4 to 12 months, all patients have remained asymptomatic without radiographic evidence of recurrent strictures. Retrograde ureteroscopic incision is an effective, minimally invasive option for patients with benign ureteral strictures. PMID- 10708142 TI - Ultrasound-guided drainage of perirenal and periureteral urine collections. AB - Thirty patients with perirenal and periureteral collections were evaluated. The reasons for these collections were after open surgery on the kidney in 11, open surgery on the ureter in 10, percutaneous nephrostolithotomy in 2, after renal transplantation in 5, and after ureteroscopy in 2. The presenting symptoms and signs included flank pain in 12, urine leakage in 5, fever in 5, masses in the iliac fossa in 5, and flank mass in 3. Ultrasound-guided single-step drainage of these collections was done using 10-12 F catheters. Ultrasound-guided single-step percutaneous nephrostomy (PCN) was done prior to drainage of the collection in seven patients in whom ultrasound revealed back pressure changes in their renal units. Pigtail catheters (7-10 F) were used for PCN. The aspirated fluid was clear urine in 12 cases and turbid in 18. The amount of fluid drained ranged from 150-500 mL immediately after the procedure. We used color Doppler sonography to map the site of puncture. No complications were encountered after drainage. The period of drainage varied from 1 week to 37 days. Further intervention was needed in 12 cases due to development of ureteric stricture in 7, prolonged leakage in 3 (one after transplantation, one after ureterocolic anastomosis, and one after ureterolithotomy), and residual stones in 2. Double-J stenting was done in four cases. We conclude that ultrasound-guided drainage of perirenal and periureteral collections is a safe, rapid, and easy method of treatment and should eliminate the need for exploration to drain these collections. Whenever backpressure exists in the renal unit, ultrasound-guided PCN should be done prior to drainage of the collection. PMID- 10708143 TI - Local transrectal enzymatic treatment for chronic nonbacterial prostatitis and prostatodynia: initial clinical experience. AB - We report our initial clinical experience with local transrectal application of enzymatic treatment for chronic nonbacterial prostatitis and prostatodynia in 20 patients. Using a specially designed symptom score for evaluation of subjective treatment parameters, a statistically significant improvement of symptoms was found in the areas of pain, micturition, and recreational activities. No statistically significant differences were noted in laboratory values before and after treatment. Minimal local side effects were seen in only one patient. A favorable clinical response was noted in 75% of patients, whereas the remaining 25% showed only moderate improvement of symptoms. No patient experienced complete treatment failure. PMID- 10708144 TI - Use of residual fraction instead of residual volume in the evaluation of lower urinary tract symptoms. AB - Assessment of postvoid residual volume (PVR) has become a valuable routine investigation in the evaluation of bladder outlet obstruction. PVR has been shown to have interindividual and intraindividual variation and dependence on prevoid urinary volume, thus raising a question about its significance. The aim of this study was to investigate an alternate parameter more reliable than PVR, described as residual fraction (RF) and calculated as (PVR x 100)/prevoid volume. Ninety three adult patients with lower urinary tract symptoms (LUTS) presenting to the urology outpatient clinic were evaluated for bladder outlet obstruction. Patients with urinary retention and neurological disorders were excluded. Evaluation was by clinical assessment, uroflowmetry, and ultrasound bladder for prevoid and postvoid urine volume estimation. The latter was compared with RF. Results were statistically analyzed using bivariate analysis and Spearman's test. In 93 evaluable patients, there were 87 (94%) males and 6 (6%) females. Residual volume ranged from 4 to 450 mL (mean 91.4+/-92.7 mL). Peak flow rate (Qmax) varied from 3 to 49 mL/s (mean 9+/-15 mLis). Two-tailed correlation between PVR and Qmax was significant at the .05 level, whereas a more significant R value at .01 was observed between RF and Qmax. There was a strong positive correlation between RF and PVR with Qmax. RF statistically correlated better with Qmax than PVR. It is recommended that RF be used instead of PVR in the routine noninvasive evaluation of LUTS. PMID- 10708145 TI - Transverse retubularized ileovesicostomy continent urinary diversion to the umbilicus. AB - The transverse retubularized ileovesicostomy is useful as a continent urinary diversion when the appendix is unusable or unavailable for an appendicovesicostomy continent urinary diversion. Eight patients (mean age 29 years) with difficulty catheterizing their native urethras underwent creation of a transverse retubularized ileovesicostomy continent urinary diversion to the umbilicus. Diagnoses included myelomeningocele (3), multiple sclerosis (1), and spinal cord injury (4). Concomitant procedures included ileocystoplasty, antegrade continence enema procedure, and pubovaginal sling. All patients were able to catheterize their ileovesicostomy conduit and stoma easily with a 14F catheter. Six patients were completely dry, and two patients needed to catheterize every 3 to 4 hours to prevent urinary leakage. Mean follow-up was 3 years. Experience with the transverse retubularized ileovesicostomy continent urinary diversion to the umbilicus has been favorable. When the Mitrofanoff appendicovesicostomy continent urinary diversion is not an option, the transverse retubularized ileovesicostomy has several advantages as a second choice. PMID- 10708146 TI - Prostate cryoablation without an insertion kit using direct transperineal placement of ultrathin freezing probes. AB - A modified technique for prostate cryoablation is described. Prostate cryoablation was performed with a gas-based cryomachine using multiple 17-gauge probes. The 17-gauge probes were inserted via a transperineal route directly into the prostate without using an insertion kit. Probe insertion and positioning are simplified, and operative time is reduced. The perineum is less crowded with devices and tubes. Miniaturizing the cryoprobes and their tubing is feasible. The smaller diameter enables direct insertion without an insertion kit. The surgical result does not seem to be inferior to that of conventional ablation, and penetrating trauma to the prostate is minimal. To the best of our knowledge, this is the first report of transperineal prostate cryoablation performed without an insertion kit. PMID- 10708147 TI - Inline tug on a through-and-through glidewire to reposition open-ended catheter during percutaneous nephrolithotomy. AB - We present a simple technique to reposition an open-ended ureteric catheter in the pelvicalyceal system during percutaneous nephrolithotomy. A through-and through glidewire is straightened using a stone-grasping forceps. The open-ended catheter is advanced in the pelvicalyceal system over this taut glidewire. PMID- 10708148 TI - Use of a laparoscopic instrument to improve urethrovesical anastomosis quality during retropubic radical prostatectomy. AB - Urethrovesical or urethroileal anastomosis is a critical step during radical surgery to obtain good postoperative continence without restenosis of the lumen. A number of surgical maneuvers and technical devices have been proposed for safer reconstruction of the urethrovesical junction. We experimentally used a 25 charrier cystoscope sheath as a guide for exact placement of sutures on the urethral stump with an instrument designed to perform laparoscopic sutures. Among the advantages of this technique are easy and quick placement of the stitches, which can be placed to an exact depth and in the desired direction. We believe that future modifications of laparoscopic instruments will make oncologic pelvic surgery easier. PMID- 10708150 TI - Retrograde ureteroscopic endopyelotomy for the treatment of primary and secondary ureteropelvic junction obstruction in children. AB - The use of endopyelotomy in children with ureteropelvic junction (UPJ) obstruction remains controversial. Although most investigators reported good results with percutaneous or retrograde balloon cautery incision, there are distinct advantages associated with a ureteroscopic approach. Three male children, ages 11, 12 and 17 years, underwent ureteroscopic endopyelotomy for treatment of UPJ obstruction (one primary and two secondary). The procedures were performed using 6F to 8.5F semirigid instruments and the holmium laser. All three patients underwent endopyelotomy without complication. The mean operative time was 80 minutes. Two patients were discharged home the day of the procedure, and the third patient was hospitalized for less than 24 hours postoperatively. With follow-up of 6 to 11 months, two patients are asymptomatic, with no radiographic evidence of obstruction. The 12-year-old boy had continued obstruction following endopyelotomy. At the time of open pyeloplasty, a large crossing vessel was noted, which appeared to be the source of obstruction. Ureteroscopic endopyelotomy can be performed with minimal morbidity and hospitalization in children. Further clinical experience is needed to assess the relative efficacy of this procedure in comparison with other forms of endopyelotomy in children. PMID- 10708149 TI - Total pelvic exenteration and reconstruction for locally invasive recurrent sarcoma of the perineum. AB - Extensive primary tumors and locally recurrent tumors of the pelvis or perineum are difficult to manage. We describe the techniques necessary to perform total pelvic exenteration with en bloc resection of the perineum and genitalia for treatment of recurrent sarcoma of the perineum. Wide excision of the sarcoma with negative margins can be achieved by resecting the inferior portion of the pubic symphysis. An absorbable mesh sling may be used to suspend the small bowel above the pelvis, facilitating postoperative radiation. A catheterizable continent urinary reservoir avoids the necessity of two stomas and improves quality of life. Adequate tissue coverage can be attained by myocutaneous gracilis flaps that promote wound healing. PMID- 10708152 TI - Case no. 1. Newborn scrotal swelling and testicular mass. PMID- 10708151 TI - Primary localized amyloidosis of the ureter and bladder managed by ileal interposition. AB - Amyloidosis of the ureter is a rare condition. It is even rarer when it involves both the ureter and bladder. The case presented is the second known case of combined amyloidosis of the bladder and ureter and the first combined case to be treated successfully by ileal ureter replacement. Historically, amyloidosis of the ureter has been treated by nephroureterectomy. Based on the benign nature of the disease, amyloidosis of the ureter is optimally treated with a kidney-sparing procedure such as ileal ureter replacement. PMID- 10708153 TI - Case no. 2. Renal mass. PMID- 10708154 TI - Case no. 3. Ambiguous genitalia. PMID- 10708155 TI - Case no. 4. 55-year-old man presenting with sudden right loin pain, loin mass, and sepsis. PMID- 10708156 TI - Prone positioning attenuates and redistributes ventilator-induced lung injury in dogs. AB - BACKGROUND: We previously demonstrated a markedly dependent distribution of ventilator-induced lung injury in oleic acid-injured supine animals ventilated with large tidal volumes and positive end-expiratory pressure > or =10 cm H2O. Because pleural pressure distributes more uniformly in the prone position, we hypothesized that the extent of injury induced by purely mechanical forces applied to the lungs of normal animals might improve and that the distribution of injury might be altered with prone positioning. OBJECTIVE: To compare the extent and distribution of histologic changes and edema resulting from identical patterns of high end-inspiratory/low end-expiratory airway pressures in both supine and prone normal dogs. DESIGN/SETTING: We ventilated 10 normal dogs (5 prone, 5 supine) for 6 hrs with identical ventilatory patterns (a tidal volume that generated a peak transpulmonary pressure of 35 cm H2O when implemented in the supine position before randomization, positive end-expiratory pressure = 3 cm H2O). Ventilator-induced lung injury was assessed by gravimetric analysis and histologic grading. MEASUREMENTS AND MAIN RESULTS: Wet weight/dry weight ratios (WW/DW) and histologic scores were greater in the supine than the prone group (8.8+/-2.8 vs. 6.1+/-0.7; p = .01 and 1.4+/-0.3 vs. 1+/-0.3; p = .037, respectively). In the supine group, WW/DW and histologic scores were significantly greater in dependent than nondependent regions (9.4+/-1.9 vs. 6.7+/ 0.9; p = .01 and 2.0+/-0.4 vs. 0.9+/-0.4; p = .043, respectively). In the prone group, WW/DW also was greater in dependent regions (6.7+/-1.1 vs. 5.8+/-0.5; p = .054), but no significant differences were found in histologic scores between dependent and nondependent regions (p = .42). CONCLUSION: In this model of lung injury induced solely by mechanical forces, the prone position resulted in a less severe and more homogeneous distribution of ventilator-induced lung injury. These results parallel those previously obtained in oleic acid-preinjured animals ventilated with higher positive end-expiratory pressure. PMID- 10708157 TI - Short-term effect of inhaled nitric oxide and prone positioning on gas exchange in patients with severe acute respiratory distress syndrome. AB - OBJECTIVE: To compare the short-term effects of inhaled nitric oxide (NO) and prone positioning in improving oxygenation in acute respiratory distress syndrome (ARDS). METHODS: Charts of consecutive ARDS patients (lung injury score >2) during a 2-yr period, tested for both inhaled NO and prone positioning efficacy were retrospectively reviewed. Variations in the Pao2/Fio2 ratio induced by inhaled NO and prone positioning were evaluated. MEASUREMENTS AND MAIN RESULTS: Twenty-seven patients (age, 42+/-17 yrs) were included. Simplified Acute Physiology Score II was 45+/-14. Mortality rate in the intensive care unit was 63%. The causes of ARDS were pneumonia (n = 14), extra-lung infection (n = 5), and noninfectious systemic inflammatory response syndrome (n = 8). Lung injury score was 2.7+/-0.3. At baseline, before the initiation of inhaled NO, the Pao2/Fio2 ratio was 97+/-46 torr and before prone positioning, 92+/-26 torr. Variations in the Pao2/Fio2 ratio were lower at start of NO therapy (11+/-4 ppm) than that observed at prone positioning initiation (23+/-31 vs. 62+/-78 torr, p<.05). An increase in variations in the Pao2/Fio2 ratio of >15 torr was associated with prone positioning in 16 patients (59%) and with NO inhalation in 13 patients (48%) (not significant). An increase in variations in the Pao2/Fio2 ratio of >15 torr was associated with both techniques in only six patients (22%). There was no correlation between the response to prone positioning and the response to inhaled NO (r2 = .005; p = .73). CONCLUSIONS: Prone positioning improves hypoxemia significantly better than does inhaled NO. The response to one technique is not predictive of the response to the other technique. PMID- 10708158 TI - Incidence of atrial fibrillation after mild or moderate hypothermic cardiopulmonary bypass. AB - OBJECTIVES: Atrial fibrillation remains a significant source of morbidity after coronary artery bypass grafting (CABG). Whether cardiopulmonary bypass (CPB) temperature influences the occurrence of postoperative atrial fibrillation in CABG patients has not been specifically examined. In the present study, we reviewed postoperative data from patients who were prospectively randomized to mild or moderate hypothermic CPB for elective CABG to determine the incidence of postoperative atrial fibrillation. DESIGN: Randomized, single center, observational study. SETTING: Tertiary university medical center. PATIENTS: Adults undergoing elective CABG surgery. INTERVENTIONS: Enrolled patients were prospectively randomized to mild (34 degrees C [93.2 degrees F]) or moderate (28 degrees C [82.4 degrees F]) hypothermic CPB. MEASUREMENTS AND MAIN RESULTS: The incidence of postoperative atrial fibrillation was determined by review of ICU and hospital records. There was a significantly higher incidence of atrial fibrillation in the moderate compared with the mild hypothermic CPB group. Patients who had postoperative atrial fibrillation were significantly older than those without atrial fibrillation. Furthermore, a significant increase in the relative risk of developing postoperative atrial fibrillation was found for both age and CPB temperature. CONCLUSIONS: Our results indicate that the temperature of systemic cooling during CPB is an important factor in the development of atrial fibrillation after CABG surgery. In addition, this study confirms that increasing age is a significant determinant of postoperative atrial fibrillation. PMID- 10708159 TI - Effects of a heat and moisture exchanger and a heated humidifier on respiratory mucus in patients undergoing mechanical ventilation. AB - OBJECTIVE: To evaluate the effects of a heat and moisture exchanger and a heated humidifier on respiratory mucus and transportability by cilia and cough in patients undergoing invasive mechanical ventilation (up to 72 hrs). DESIGN: Prospective, randomized, clinical study. SETTING: General intensive care unit and university research laboratory. PATIENTS: A total of 32 consecutive patients with acute respiratory failure, who were intubated and mechanically ventilated in the intensive care unit setting, were enrolled in the study. INTERVENTIONS: Patients were randomly assigned to receive as a humidifying system a heat and moisture exchanger (HME) or heated humidified water (HHW) at the onset of mechanical ventilation (time 0). Respiratory mucus samples were collected by suction using a sterile technique at time 0, 24, 48, and 72 hrs of mechanical ventilation. MEASUREMENTS AND MAIN RESULTS: Eleven patients were excluded from this study because of either extubation or death before 72 hrs of mechanical ventilation, leaving 12 patients in the HME group and nine patients in the HHW group. Ventilatory variables including minute volume, mean airway pressure, positive end expiratory pressure, Fio2, as well as Pao2/Fio2 ratio, fluid balance (last 6 hrs), furosemide, and inotrope administration (last 4 hrs) were recorded. In vitro mucus transportability by cilia was evaluated on the mucus-depleted frog palate model, and the results were expressed as the mucus transport rate. Cough clearance (an estimation of the interaction between the flow of air and the mucus lining the bronchial walls) was measured using a simulated cough machine, the results being expressed in millimeters. Mucus wettability was measured by the contact angle between a mucus sample drop and a flat glass surface. Mucus rheologic properties (mechanical impedance [log G*] and the ratio between viscosity and elasticity [tan delta]) were measured using a magnetic microrheometer at 1 and 100 cGy/sec deformation frequency. The two humidification groups were comparable in terms of the Acute Physiology and Chronic Health Evaluation II score, age, gender, ventilatory variables, fluid balance, use of inotropes, and furosemide. CONCLUSION: Ours results indicate that air humidification with either HME or HHW at 32 degrees C (89.6 degrees F) has similar effects on mucus rheologic properties, contact angle, and transportability by cilia in patients undergoing mechanical ventilation, except for transportability by cough, which diminished after 72 hrs of mechanical ventilation in the HME group (p = .0441). PMID- 10708160 TI - Cardiovascular response and stress reaction to flumazenil injection in patients under infusion with midazolam. AB - OBJECTIVES: To evaluate the cardiovascular response and acute stress reaction after arousal induced by a benzodiazepine antagonist, flumazenil, in patients sedated with midazolam. DESIGN: Prospective study. SETTING: Emergency center in a university hospital. PATIENTS: A total of 12 patients were ventilated mechanically under sedation with midazolam. INTERVENTIONS: We monitored the consciousness level, heart rate, systemic blood pressure, pulmonary artery pressure, and pulmonary artery occlusion pressure before and after a bolus injection of 0.5 mg of flumazenil. The score for the consciousness level represents the sum of the scores for eye opening and best motor response, as determined by the Glasgow Coma Scale. We measured the cardiac output, concentrations of norepinephrine, epinephrine, and 3-methoxy-4 hydroxyphenylethyleneglycol in plasma, and concentration of cortisol in serum. We calculated the left ventricular ejection fraction, cardiac index, systemic vascular resistance index, pressure-rate product, systemic oxygen delivery, and systemic oxygen consumption at 0, 10, 30, and 60 mins after injection of flumazenil. MEASUREMENTS AND MAIN RESULTS: The serum benzodiazepine's receptor binding activity in serum was in the range from 50 to 1000 ng/mL before injection of flumazenil. Flumazenil improved the consciousness level from 6.7+/-2.0 to 8.9+/-1.6 and induced transient elevations in heart rate, blood pressure, systolic pulmonary artery pressure, and pulmonary artery occlusion pressure. Left ventricular ejection fraction, oxygen delivery index, and pressure-rate product increased significantly, from 61%+/-8%, 640+/-170 mL/min/m2, and 13,300+/-2600 mm Hg/min at 0 mins to 67% +/-5%, 710+/-220 mL/min/m2, and 16,500+/-4400 mm Hg/min at 10 mins, respectively. Concentrations of norepinephrine and epinephrine in plasma increased significantly, from 890+/-840 pg/mL and 220+/-360 pg/mL, respectively, at 0 mins to 990+/-850 pg/mL and 270+/-300 pg/mL, respectively, at 10 mins. There were no significant changes in the plasma concentration of 3 methoxy-4-hydroxyphenylethyleneglycol, the serum concentration of cortisol after the administration of flumazenil. CONCLUSIONS: Flumazenil did not result in a significant acute stress reaction in midazolam-sedated patients, but it increased myocardial oxygen consumption by enhancing sympathetic nervous activity or antagonizing cardiovascular depression induced by midazolam. PMID- 10708161 TI - Altered plasma cytokines and total glutathione levels in parenterally fed critically ill trauma patients with adjuvant recombinant human growth hormone (rhGH) therapy. AB - OBJECTIVES: Glutathione (GSH) is a potent endogenous antioxidant that serves as one of the body's most important defenses against oxygen metabolites. Plasma levels of GSH are maintained primarily by a balance between secretion from the liver and degradation in the kidney. The ability to maintain and enhance tissue GSH may be of particular importance in controlling cytokine production in response to a stimulus like injury. The interaction after severe trauma between GSH and cytokines, tumor necrosis factor (TNF) -alpha, and interleukin (IL)-6, are not known. The purpose of the study was to investigate the levels of plasma GSH and cytokines TNF-alpha and IL-6 in adult patients admitted to the intensive care unit of our level I trauma center who were treated with recombinant human growth hormone (rhGH) for > or =7 days. DESIGN: Prospective, randomized, controlled trial. SETTING: Trauma intensive care unit. PATIENTS: Twenty-eight patients with multiple injuries and 14 normal postabsorptive controls. INTERVENTIONS: From 48-60 hrs after injury, when resuscitation was complete, a stable hemodynamic status was achieved and the patients were receiving maintenance fluid without nitrogen or calories, a blood sample was drawn for basal, plasma GSH, TNF-alpha, and IL-6 measurement. Intravenous feeding was then started and continued for 7 days. The patients were randomized to receive or not to receive daily intramuscular doses of recombinant human growth hormone (0.15 mg rhGH/kg/day). Daily morning plasma was obtained for analysis of GSH, TNF-alpha, and IL-6 levels. RESULTS: In the early catabolic "flow phase" of severe injury, the plasma levels of GSH were not altered but plasma TNF-alpha and IL-6 levels were increased significantly, compared with uninjured controls. Seven days of total parenteral nutrition alone enhanced plasma GSH levels (76%), but no change in TNF-alpha was observed. Supplementation with rhGH enhanced GSH (180%), and TNF (65%) with no changes in IL-6 levels. There is a significant linear relationship between plasma GSH and TNF-alpha levels in our rhGH-supplemented trauma patients. CONCLUSION: Modification of plasma GSH and TNF-alpha levels by adequate nutritional support with adjuvant rhGH during the postinjury period demonstrates the beneficial role of GSH in enhancing antioxidant defenses. PMID- 10708162 TI - Prospective evaluation of pulmonary edema. AB - OBJECTIVES: To describe the clinical profile and hospital outcome of successive unselected patients with pulmonary edema hospitalized in an internal medicine department. DESIGN: Prospective, consecutive, unsolicited patients diagnosed with pulmonary edema. SETTING: An internal medicine department in a 900 tertiary care center. PATIENTS: A total of 150 consecutive unselected patients (90 males, 60 females; median age, 75 yrs). RESULTS: Ischemic heart disease, hypertension, various valvular lesions and diabetes mellitus were present in 85%, 70%, 53%, and 52% of patients, respectively. Acute myocardial infarction at admission was observed in 15% of patients. The most common precipitating factors associated with the development of pulmonary edema included: high blood pressure (29%), rapid atrial fibrillation (29%,) unstable angina pectoris (25%), infection (18%), and acute myocardial infarction (15%). Twenty-two patients (15%) were mechanically ventilated. Eighteen patients (12%) died while in the hospital, and the cause of death was cardiac pump failure in 82%. The median hospital stay was 10 days. Predictors for increase rate of in-hospital mortality included: diabetes (p<.05), orthopnea (p<.05), echocardiographic finding of moderate-to-severely depressed global left ventricular systolic function (p<.001), acute myocardial infarction during hospital stay (p<.001), hypotension/shock (p<.05), and the need for mechanical ventilation (p<.001). CONCLUSIONS: Most patients with pulmonary edema in the internal medicine department are elderly, having ischemic heart disease, hypertension, diabetes, and a previous history of pulmonary edema. The overall mortality is high (in-hospital, 12%) and the predictors associated with high in-hospital mortality are related to left ventricular myocardial function. The long median hospital stay (10 days) and the need for many cardiovascular drugs, impose a considerable cost in the management and health care of these patients. PMID- 10708163 TI - Changes in circulating blood volume after cardiac surgery measured by a novel method using hydroxyethyl starch. AB - OBJECTIVE: To determine the incidence and extent of postoperative blood volume (BV) changes in patients after elective cardiac surgery using a new method based on dilution of hydroxyethyl-starch. DESIGN: Prospective, clinical, and laboratory investigation. SETTING: University hospital intensive care unit. PATIENTS: A total of thirty-five patients undergoing cardiac surgery requiring cardiopulmonary bypass (CPB). INTERVENTIONS: Perioperative measurements of circulating BV, systemic hemodynamics, lactate, and collection of clinical data. MEASUREMENTS AND MAIN RESULTS: Measurements were made before and 1 to 72 hrs after CPB. The majority of patients undergoing cardiac surgery showed postoperative BV deficits compared with preoperative BV despite marked positive fluid balances after CPB. At 1 hr and 5 hrs after CPB, 18% and 33% of the patients, respectively, had BV deficits in the range of 0.5 L and 1.5 L, and in 3% to 10% of the cases, postoperative BV deficits exceeded 1.5 L. Concomitantly, at 5 hrs after CPB, mean arterial pressure was maximally reduced, and heart rate and lactate levels were maximally elevated. Thereafter, BV began to normalize, and at 24 hrs after CPB, pre- and postoperative mean BV were no longer significantly different. At 48 hrs and 72 hrs, even a BV surplus of more than 1 L could be observed in 6% and 14% of the patients, respectively. CONCLUSIONS: During the first hours after CPB, a high percentage of patients had significantly reduced BV and, concomitantly, showed cardiovascular dysfunction and hyperlactemia. Because hypovolemia is associated with increases of perioperative morbidity and mortality, rapid determination of BV is warranted to guide fluid therapy and optimize treatment in patients undergoing cardiac surgery. PMID- 10708164 TI - The impact of long-term acute-care facilities on the outcome and cost of care for patients undergoing prolonged mechanical ventilation. AB - OBJECTIVES: To compare the 6-month mortality rate of chronically ventilated patients treated either exclusively in a traditional acute-care hospital or transferred during hospitalization to a long-term acute-care facility. To analyze the hospital cost of care and estimate the amount of uncompensated care incurred by acute-care hospitals under the Medicare prospective payment diagnostic related groups system. DESIGN: Retrospective chart review and questionnaire. SETTING: Fifty-four acute-care referral hospitals and 26 longterm acute-care institutions. PATIENTS: A total of 432 ventilated patients selected from 3,266 patients referred but not transferred to a study long-term acute-care facility and 1,702 ventilated patients from 4,174 patients referred and then subsequently transferred to the long-term acute-care facility. Six-month outcomes were determined for the subgroup of patients > or =65 yrs old (279 and 1,340 patients, respectively). Hospital charges were available for 192 of the 279 nontransferred patients who were > or =65 yrs old and 1,332 of the 1,340 transferred patients. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: The 6-month mortality rate was 67.4% for the 279 nontransferred patients and 67.2% for the 1,340 transferred patients. On multiple regression analysis, variables associated with the 6-month mortality rate included initial admitting diagnosis, age, the acute physiology score, and presence of decubitus ulcer. After controlling for these variables, there was no significant difference in 6-month mortality rate, but admission to the long-term acute-care facility was associated with a longer mean survival time. Average total hospital costs for the 192 nontransferred patients was $78,474, and estimated Medicare reimbursement was $62,472, resulting in an average of $16,002 of uncompensated care per patient. Estimated costs for the long-term acute-care facility admissions were $56,825. CONCLUSIONS: Patients undergoing prolonged ventilation have high hospital and 6-month mortality rates, and 6-month outcomes are not significantly different for those transferred to long-term acute-care facilities. These patients generate high costs, and acute care hospitals are significantly underreimbursed by Medicare for these costs. Acute-care hospitals can reduce the amount of uncompensated care by earlier transfer of appropriate patients to a long-term acute-care facility. PMID- 10708165 TI - Factors predicting perioperative cytokine response in patients undergoing liver transplantation. AB - OBJECTIVES: An exaggerated production of proinflammatory cytokines during liver transplantation stimulates the inflammatory process within the graft, and eventually promotes liver failure. This study was conducted to evaluate factors predicting perioperative response of proinflammatory cytokines during liver transplantation. DESIGN: Prospective, consecutive entry study of liver transplant candidates. SETTING: University hospital. PATIENTS: Thirty liver transplant recipients. INTERVENTIONS: Arterial blood samples were obtained perioperatively. MEASUREMENTS: Interleukin (IL)-1beta, IL-6, tumor necrosis factor-alpha were measured by ELISA. Endotoxin was determined by a chromogenic endotoxin-specific method. MAIN RESULTS: The peak concentrations of IL-1beta and IL-6 in the patients with complications were significantly higher than those in the patients without complications. The peak concentration of IL-1beta was significantly correlated with the level of bilirubin at admission and the intraoperative blood product requirement. The peak concentration of IL-6 was significantly correlated with the admission bilirubin and the intraoperative blood product requirement. A multivariate regression model revealed that the serum bilirubin and the intraoperative blood product requirement were the independent factors that influenced the peak concentration of IL-1beta or IL-6. The severely jaundiced patients had a significantly higher plasma concentration of endotoxin at the end of the anhepatic phase. In addition, there was a tendency for these patients to have a higher postoperative peak concentration of endotoxin. CONCLUSIONS: Serum level of bilirubin may be a potent preoperative factor influencing perioperative cytokine response in patients undergoing liver transplantation. An enhanced perioperative response of endotoxin seen in severely jaundiced patients suggests the clinical implication of endotoxin removal during the anhepatic phase in liver transplant surgery. PMID- 10708166 TI - Insertion time of the pulmonary artery catheter in critically ill patients. AB - OBJECTIVES: Measurement of the time elapsed from the decision to use a pulmonary artery catheter to the onset of the adapted treatment. DESIGN: Prospective study. SETTING: Critical care unit of a university hospital. PATIENTS: A total of 104 critically ill patients. INTERVENTIONS: The time elapsed from the decision to use a pulmonary artery catheter to the onset of the adapted treatment. Five time intervals (availability, preparation, catheterization, data collection, and therapeutic intervals) were individualized according to the times of decision of pulmonary artery catheter insertion, operator's hand washing, venipuncture, postoperative dressing, data collection, and the effective onset of subsequent therapy. MEASUREMENTS AND MAIN RESULTS: Among 120 used pulmonary artery catheters, seven could not be inserted. The time to use the pulmonary artery catheter was never shorter than 45 mins (median value = 120 mins). For availability, preparation, catheterization, data collection, and therapeutic intervals, the median values were 30, 20, 20, 20, and 10 mins, respectively. The availability and data collection intervals were shortened during the night period and the fourth quarter of the study, respectively. CONCLUSIONS: The pulmonary artery catheter use is time consuming. However, the availability and data collection intervals could be shortened. PMID- 10708167 TI - Effect of acute moderate changes in PaCO2 on global hemodynamics and gastric perfusion. AB - OBJECTIVE: To describe global hemodynamics and splanchnic perfusion changes in response to acute modifications in Paco2 in hemodynamically stable patients. DESIGN: Prospective, randomized crossover study. SETTING: Medical-surgical intensive care unit at a community hospital (400,000 inhabitants). PATIENTS: Ten critically ill patients who were sedated, paralyzed, and mechanically ventilated. INTERVENTIONS: Hypercapnia and hypocapnia were obtained by increasing and reducing instrumental deadspace in random order. After each intervention, patients returned to the basal condition. Each period lasted 80 min: 20 min to achieve stable Paco2 and 60 min for tonometer equilibration. In each period, global hemodynamic variables and tonometric data were collected. The periods were compared using analysis of variance. MEASUREMENTS AND MAIN RESULTS: Acute hypercapnia (Paco2 from 40+/-3 to 52+/-3 torr, p<.05) increased cardiac index (3.43+/-0.37 vs. 3.97+/-0.43 mL/min/m2, p<.05), heart rate (95+/-6 vs. 105+/-3 beats/min, p<.05), and mean pulmonary artery pressure (21+/-1 vs. 24+/-1 mm Hg, p<.05) and reduced systemic vascular resistance (992+/-98 vs. 813+/-93 dyne x sec/ cm5, p<.05) and oxygen extraction ratio (27+/-3% vs. 22+/-2%, p<.05). Standardized intramucosal Pco2 increased from 49+/-2 to 61+/-3 torr (p<.05) with an associated decrease in calculated intramucosal pH ([pHi] 7.35+/-0.03 vs. 7.25+/-0.02, p<.05), but the gastro-arterial Pco2 gradient (deltaPco2) did not change. Acute hypocapnia (Paco2 from 41+/-3 to 34+/-3 torr, p<.05; pH 7.41+/-0.01 to 7.47+/-0.02, p<.05) induced slight increments in systemic vascular resistance (995+/-117 vs. 1088 +/- 160 dyne x sec/cm5, p<.05) and oxygen extraction ratio (28+/-2% vs. 30+/-2%, p<.05). Standardized intramucosal Pco2 decreased (50+/-4 vs. 44+/-3 torr, p<.05), pHi increased (7.33+/-0.03 vs. 7.36+/-0.02; p<.05), but deltaPco2 did not change. CONCLUSIONS: In this small group of stable patients, moderate acute variations in Paco2 had a significant effect on global hemodynamics, but splanchnic perfusion, assessed by deltaPco2, did not change. In these conditions, the use of pHi to evaluate gastric perfusion appears unreliable. PMID- 10708168 TI - Evaluation of an antiseptic triple-lumen catheter in an intensive care unit. AB - OBJECTIVE: To evaluate a decrease in catheter-related bloodstream infection rate in patients with antiseptic triple-lumen catheters in an intensive care unit. DATA SOURCES: Retrospective review of surveillance records, patient medical records, laboratory and microbiological reports, and antibiotic administration records. STUDY SELECTION: Patients admitted to the intensive care unit with triple-lumen catheters. DATA EXTRACTION: A subset of one entry per patient was extracted from 2 yrs of primary bloodstream infection surveillance data. Data collection included risk factors, laboratory and microbiological data, and insertion sites and dates of all intravascular catheters present during triple lumen catheterization. DATA SYNTHESIS: The catheter-related bloodstream infection rate was 5.4 and 11.3 per 1000 catheter days in antiseptic and nonantiseptic triple-lumen catheter groups, respectively (p = .06). By multivariate analysis using a Cox Proportional Hazards Model, the antiseptic triple-lumen catheters were associated with a significant reduction in catheter-related bloodstream infection (p = .03). Model expansion to include intrajugular site was significant by a likelihood ratio test [2(log likelihood diff) = 4.26 P<.05 chi2(1)] CONCLUSIONS: The use of antiseptic triple-lumen catheters may substantially reduce catheter-related bloodstream infections in an intensive care population and may be subsequently associated with a decrease in length of stay. PMID- 10708169 TI - Accuracy of oscillometric blood pressure measurement according to the relation between cuff size and upper-arm circumference in critically ill patients. AB - OBJECTIVE: To evaluate the accuracy of oscillometric blood pressure measurement according to the relation between cuff size and upper-arm circumference in critically ill patients. DESIGN: Prospective data collection. SETTING: Emergency department in a 2,000-bed inner city hospital. PATIENTS: Thirty-eight patients categorized into three groups according to their upper-arm circumference (group I: 18-25 cm; group II: 25.1-33 cm; and group III: 33.1-47.5 cm) were enrolled in the study protocol. INTERVENTIONS: In each patient, all three cuff sizes (Hewlett Packard Cuff 40401 B, C, and D) were used to perform an oscillometric blood pressure measurement at least within 3 mins until ten to 20 measurements for each cuff size were achieved. Invasive mean arterial blood pressure measurement was done by cannulation of the contralateral radial artery with direct transduction of the systemic arterial pressure waveform. The corresponding invasive blood pressure value was obtained at the end of each oscillometric measurement. MEASUREMENT AND MAIN RESULTS: Overall, 1,494 pairs of simultaneous oscillometric and invasive blood pressure measurements were collected in 38 patients (group I, n = 5; group II, n = 23; and group III, n = 10) over a total time of 72.3 hrs. Mean arterial blood pressure ranged from 35 to 165 mm Hg. The overall discrepancy between oscillometric and invasive blood pressure measurement was -6.7+/-9.7 mm Hg (p<.0001), if the recommended cuff size according to the upper-arm circumference was used (539 measurements). Of all the blood pressure measurements, 26.4% (n = 395) had a discrepancy of > or =10 mm Hg and 34.2% (n = 512) exhibited a discrepancy of > or =20 mm Hg. No differences between invasive and noninvasive blood pressure measurements were noted in patients either with or without inotropic support (-6.6 + 7.2 vs. -8.6 + 6.8 mm Hg; not significant). CONCLUSION: The oscillometric blood pressure measurement significantly underestimates arterial blood pressure and exhibits a high number of measurements out of the clinically acceptable range. The relation between cuff size and upper arm circumference contributes substantially to the inaccuracy of the oscillometric blood pressure measurement. Therefore, oscillometric blood pressure measurement does not achieve adequate accuracy in critically ill patients. PMID- 10708170 TI - Nosocomial endocarditis in the intensive care unit: an analysis of 22 cases. AB - OBJECTIVES: To review the intensive care unit experience of patients with admitted or acquired nosocomial endocarditis (NE) defined according to the Duke criteria. DESIGN: Prospective, cohort study. SETTING: University teaching hospital. PATIENTS: We reviewed the records of 22 patients documented with NE during a 6-yr period from 1992 to 1997. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Twenty-two patients (9 women/13 men) aged 38-83 yrs (mean 65+/-9 yrs) had a NE (prevalence of 5 per 1,000 admissions). For six patients, NE was the reason for the admission to the intensive care unit. For 17 patients, the time elapsed between admission and diagnosis of NE was 39+/-25 days. Sixteen patients were predisposed to infection and seven had underlying heart conditions that put them at risk for acute endocarditis: three prosthetic valves, two valvular diseases, and two cardiac pacemakers. In 21 cases (one unknown portal of entry), NE was the consequence of bacteremia related to a medical or surgical procedure: 11 intravascular devices, eight surgical wounds, one tracheal procedure, and one leg ulceration. The bacteriologic agents detected in blood cultures were: staphylococci (n = 17), Streptococcus (n = 2), Pseudomonas aeruginosa (n = 2), and Candida (n = 2). Fourteen patients underwent echocardiography according to cardiac signs (cardiac failure, new cardiac murmur, or embolic event). For the eight remainders, echocardiography was performed systematically because of fever and positive blood cultures. The lesions detected by 21 transthoracic and 17 transesophageal echocardiographs were the following: vegetations (n = 19), myocardial abscesses (n = 5), and valvular perforation (n = 1). On 16 surgical indications, only five patients underwent surgery because the others were in too poor of a condition. The overall mortality was 68% (n = 15) and was directly associated with NE in 36% of cases (n = 8). Seven patients (28%) were discharged 34 days after the diagnosis of endocarditis. CONCLUSIONS: NE is a frequent nosocomial infection that occurs late during hospitalization. Persistent fever with positive blood cultures is sufficient symptomology to promptly perform an echocardiogram. The poor prognosis is related to the poor condition of those patients who cannot be referred for surgical treatment. PMID- 10708171 TI - Bedside chest radiography as part of a postcardiac surgery critical care pathway: a means of decreasing utilization without adverse clinical impact. AB - OBJECTIVE: To evaluate the use of bedside chest radiography and patient outcome before and after implementation of a cardiac surgery critical care pathway that included guidelines for bedside radiography. DESIGN: A cohort observational study. SETTING: A university hospital in the midwest. PATIENTS: Three groups, of 100 patients each, undergoing cardiac surgery in 1990, 1991, and 1995. INTERVENTION: Introduction of a critical care pathway. MEASUREMENTS: Medical records were retrospectively reviewed in three groups of 100 patients each: before the introduction of the critical care pathway; 2 months after introduction of the pathway in 1991; and 4 yrs after introduction in 1995. Data were analyzed to determine operative risk for each group. Subsequent analyses determined bedside radiography use, total length of hospital stay, and patient outcome (mortality rate, complications requiring intervention, and reoperation) during hospitalization and at outpatient follow-up 15-30 days postdischarge. RESULTS: Total length of hospital stay was shorter for the 1995 group (7.6+/-6.6 days) compared with other groups (prepathway, 11.1+/-10.3 days; 1991 postpathway, 10.2+/-9.6 days; p<.05). The mean numbers of radiographs per patient were as follows: prepathway, 5.1; 1991 postpathway, 5.2; and 1995 postpathway, 3.3. The mean number of radiographs in the 1995 group was significantly lower (p = .02). More patients had the proposed number of two bedside radiographs described in the pathway in the 1995 group compared with the other groups (prepathway, p<.0001; the two-month postpathway group, p = .01). Twenty-three malpositioned catheters/tubes were found in the prepathway and 1991 groups compared with 11 in the 1995 group (p = .02). No statistically significant difference was found in inpatient complications (mediastinal bleeding, pneumothoraces, and pleural effusions), postdischarge complications, reoperations, or mortality rate. CONCLUSION: Introduction of a critical care pathway can decrease the use of bedside radiography without adversely affecting near-term patient outcomes. PMID- 10708172 TI - Should the pre-sedation Glasgow Coma Scale value be used when calculating Acute Physiology and Chronic Health Evaluation scores for sedated patients? Scottish Intensive Care Society Audit Group. AB - OBJECTIVE: To assess the effect on the performance of Acute Physiology and Chronic Health Evaluation (APACHE) II and APACHE III of two different approaches to scoring the Glasgow Coma Scale (GCS) in sedated patients. The first approach was to assume that the GCS score was normal, and the second was to use the GCS value recorded before the patient was sedated. DESIGN: Prospective cohort study over 2 yrs. SETTING: Twenty-two general adult intensive care units in Scotland. PATIENTS: 13,291 consecutive admissions to the participating intensive care units. MEASUREMENTS AND MAIN RESULTS: After exclusion of patients according to standard, predefined criteria, the Acute Physiology and Chronic Health Evaluation II and III systems were used to calculate the probability of hospital mortality for patients included in the study. In patients whose GCS scores could not be assessed accurately during the first 24 hrs, the APACHE II and III predictions were calculated twice: first, assuming that the GCS score was normal; and second, substituting the GCS score recorded before sedation. This generated two different databases for each system, and the predictions for both were compared with the observed hospital mortality rate. The effect of the two different approaches to the GCS on the performance of both APACHE II and APACHE III was assessed using measures of discrimination (area under the receiver operating characteristic curve) and goodness of fit (calibration curves and the Hosmer-Lemeshow statistic). Analysis was undertaken for both the entire cohort and for the group of patients whose APACHE scores were altered. There was a wide variation in the number of patients who had their scores altered between participating units. There were also differences between diagnostic groups. Overall, however, 50% of the patients were sedated and 22% had their scores altered. Using the presedation GCS score increased the discrimination of both APACHE II and APACHE III. The calibration of APACHE III was also improved but that of APACHE II deteriorated. The calibration improved, however, in those patients with altered scores, suggesting that the overall deterioration is attributable to other limitations in the fit of the model to these data. Although changes had the greatest effect in patients with a neurologic or trauma diagnosis, the changes were important in most diagnostic groups. CONCLUSIONS: The GCS is an important component of both APACHE II and APACHE III. It should be assessed directly whenever possible. When patients are sedated, using the GCS score recorded before sedation is preferable to the assumption of normality. The variations between different units and different diagnostic groups highlight the possible effects of case mix on the performance of prognostic scoring systems. PMID- 10708173 TI - Oxygen consumption in the early postinjury period: use of continuous, on-line indirect calorimetry. AB - OBJECTIVE: To determine the patterns of oxygen consumption (Vo2) using indirect calorimetry (IC) for the first 24 hrs after serious blunt traumatic injury. DESIGN: Prospective, observational study. SETTING: Surgical intensive care unit of a Level 1 trauma center. PATIENTS: Sixty-six mechanically ventilated patients with blunt traumatic injury and Injury Severity Score >15. INTERVENTIONS: IC for 24 hrs postinjury. Patients were resuscitated to standard parameters of perfusion. MEASUREMENTS AND MAIN RESULTS: Mean patient age was 50.1+/-18.7 yrs with a mean Injury Severity Score 30.7+/-11.3). Mean Vo2 for all patients for the 24-hr study period was 168.5+/-29.5 mL/min/m2. The level of Vo2 was not related to Injury Severity Score, the number or combination of organ systems injured, or to the use of vasoactive agents. Patients >65 yrs of age had significantly lower Vo2 (P = .0038) compared with patients < or =50 yrs. Vo2 did not change over time after resuscitation to normal parameters of perfusion. Mean Vo2 was 156.5+/-63.2 mL/min/m2 in patients who developed multiple organ dysfunction, and 172.4+/-33.3 mL/min/m2 in those who did not develop multiple organ dysfunction (p = .16). CONCLUSIONS: Seriously injured patients are hypermetabolic in the early postinjury period. The level of Vo2 is unrelated to injury severity or number of organ systems involved. Elderly patients can be expected to have lower levels of Vo2. Vo2 does not change significantly in response to resuscitation to normal parameters of perfusion. Vo2 measured by IC did not predict the development of multiple organ dysfunction. PMID- 10708174 TI - Autotriggering caused by cardiogenic oscillation during flow-triggered mechanical ventilation. AB - OBJECTIVES: We noticed that in some patients after cardiac surgery, when flow triggering was used, cardiogenic oscillation might be autotriggering the ventilatory support. In a prospective study, we evaluated the degree of cardiogenic oscillation and the frequency rate of autotriggering. We suspected that autotriggering caused by cardiogenic oscillation was more common than clinically appreciated. DESIGN: Prospective, nonrandomized, clinical study. SETTING: Surgical intensive care unit in a national heart institute. PATIENTS: A total of 104 adult patients were enrolled after cardiac surgery. INTERVENTIONS: During the study period, patients were paralyzed and ventilated with intermittent mandatory ventilation at a rate of 10 breaths/min, pressure support of 10 cm H2O, and flow triggering with a sensitivity of 1 L/min. MEASUREMENTS AND MAIN RESULTS: Because the patients would not be able to breathe spontaneously, we counted pressure-support (PS) breaths as instances of autotriggering. Then, we classified the patients into two groups according to the number of PS breaths: an "AT group" (PS breaths of >5/min) and a "non-AT group" (PS breaths of < or =5/min). If autotriggering occurred, we decreased the sensitivity so autotriggering disappeared (threshold triggering sensitivity). The intensity of cardiogenic oscillation was assessed as the flow and airway pressure at the airway opening. A total of 23 patients (22%) demonstrated more than five autotriggered breaths/min. During mechanical ventilation, the inspiratory flow fluctuation caused by cardiogenic oscillation was significantly greater in the AT group than in the non AT group (4.67+/-1.26 L/min vs. 2.03+/-0.86 L/min; p<.01). The AT group also showed larger cardiac output, higher ventricular filling pressures, larger heart size, and lower respiratory system resistance than the non-AT group. As the inspiratory flow fluctuation caused by cardiogenic oscillation increased, the level of triggering sensitivity also was increased to avoid autotriggering. In the AT group with 1 L/min of sensitivity, the respiratory rate increased (19.9+/ 2.7 vs. 10+/-0 breaths/min, p<.01), Paco2 decreased (30.8+/-4.0 torr [4.11+/-0.36 kPa] vs. 37.6+/-4.3 torr [5.01+/-0.57 kPa]; p < .01), and mean esophageal pressure increased (7.7+/-3.0 vs. 6.9+/-3.0 cm H2O; p<.01) compared with the threshold triggering sensitivity. CONCLUSIONS: Autotriggering caused by cardiogenic oscillation is common in postcardiac surgery patients when flow triggering is used. Autotriggering occurred more often in patients with more dynamic circulation. Autotriggering caused respiratory alkalosis and hyperinflation of the lungs. PMID- 10708175 TI - The distribution of costs of care in mechanically ventilated patients with chronic obstructive pulmonary disease. AB - OBJECTIVES: To delineate the costs of care of patients with Chronic Obstructive Pulmonary Disease (COPD) and respiratory failure and to compare them with those of other mechanically ventilated patients. DESIGN: A post hoc analysis of a prospective investigation. SETTING: Medical and coronary intensive care units (ICUs) of an 804-bed, university-based hospital. PATIENTS: A total of 300 mechanically ventilated patients, 44 with COPD and 256 others, were included. MEASUREMENTS AND MAIN RESULTS: Despite similar lengths of ICU stay (9 days) and mechanical ventilation (5.5 days COPD vs. 5 days other, p = .11), ICU respiratory care costs for patients with COPD were $2,422 ($1,157-$6,110) [median U.S. dollars (interquartile range)] vs. $1,580 ($738-$3,322) for the others (p = .01). Ventilator costs for COPD patients were $1,795 ($943-$5,782) vs. $1,574 ($613 $3,112) (p = .12). Total hospitalization respiratory care costs for COPD patients were higher, $4,064 ($2,422-$9,572) vs. $2,342 ($1,186-$4,591), (p = .0001), and 74.4% of the median difference in cost between COPD patients and others was accounted for by spontaneous nebulizers (p = .001), metered dose inhalers (p = .01), and pulse oximetry (p = .02). By using multiple linear regression analyses, we found that COPD remained associated with higher respiratory costs (p<.05). Respiratory Care was the third largest category of hospital costs after beds (27%) and pharmacy expenses (25%), and it comprised approximately 14% of total cost. Total hospital costs were large for both groups, but did not differ between COPD and the others [$24,217 ($16,211-$58,834) vs. $27,672 ($15,692-$53,766), respectively (p = .96)]. Linear regression analyses showed that only Acute Lung Injury score was significantly related to total ICU and hospital costs of care (p<.05). CONCLUSIONS: Costs of ICU and non-ICU respiratory care for patients with COPD are higher than costs of care for other mechanically ventilated patients. Although the increased cost of bronchodilators and oximetry in these patients may serve as target areas for reductions in respiratory care costs, it may also be true that these modalities of therapy and management are necessary and need to be used with even greater intensity to achieve better outcomes. The predominant contributions of bed and pharmacy costs in all of our patients with respiratory failure support research efforts addressing these aspects of care. PMID- 10708176 TI - Predictive value of a rapid semiquantitative D-dimer assay in critically ill patients with suspected venous thromboembolic disease. AB - OBJECTIVE: To evaluate the performance of a new, rapid semi-quantitative assay for the detection of circulating D-dimer in whole blood from critically ill patients with suspected venous thromboembolic disease. DESIGN: Prospective, blinded, single-center study. SETTING: Medical intensive care unit (ICU) of Barnes-Jewish Hospital, St. Louis, MO, a university-affiliated urban teaching hospital. PATIENTS: Two hundred thirty-nine adult patients with clinical suspicion of venous thromboembolic disease admitted to a medical ICU. INTERVENTIONS: Collection of blood samples within 24 hrs of clinical suspicion of venous thromboembolic disease. MEASUREMENTS AND MAIN RESULTS: The main outcome measures evaluated included the occurrence of venous thromboembolic disease (i.e., lower extremity venous thrombosis, pulmonary embolism, catheter-associated venous thrombosis) and hospital mortality. Fifty-seven patients (23.8%) were classified as having venous thromboembolic disease during their ICU stays (pulmonary embolism, 21 patients; lower extremity thrombosis, 44 patients; line associated venous thrombosis, 3 patients). The semiquantitative whole-blood assay for circulating D-dimer had a 96.4% sensitivity and a negative predictive value of 92.1% for identifying patients with venous thromboembolic disease. The specificity of this assay was 19.7%, and its positive predictive value was 26.9%. There was a strong correlation between the semiquantitative assay for circulating D-dimer and the quantitative measurement of circulating D-dimer using an enzyme immunoassay (Spearman's correlation coefficient, 0.8643; p<.001). We also identified a strong correlation between both the quantitative and semiquantitative measurements of circulating D-dimer with the sepsis classification proposed by the American College of Chest Physicians/Society of Critical Care Medicine (i.e., systemic inflammatory response syndrome, sepsis, severe sepsis, septic shock) for patients without venous thromboembolic disease (n = 182; quantitative measure: Spearman's correlation coefficient, 0.207; p = .002; semiquantitative measure: Spearman's correlation coefficient, 0.3519; p<.001). CONCLUSIONS: These preliminary findings suggest that a rapid whole-blood assay for the semiquantitative detection of circulating D-dimer may be a useful diagnostic tool for the exclusion of venous thromboembolic disease among critically ill patients. PMID- 10708177 TI - Nadroparin versus dalteparin anticoagulation in high-volume, continuous venovenous hemofiltration: a double-blind, randomized, crossover study. AB - OBJECTIVES: To compare filter survival times during high-volume, continuous venovenous hemofiltration in patients with normal coagulation variables, using anti-factor Xa bioequivalent doses of nadroparin and dalteparin. To evaluate which other factors influence filter survival time. DESIGN: Randomized, prospective, double-blind, crossover study. SETTING: An 18-bed intensive care unit in a 530-bed teaching hospital. PATIENTS: Thirty-two critically ill patients with renal failure, treated with high-volume, continuous venovenous hemofiltration. INTERVENTIONS: High-volume, postdilutional continuous venovenous hemofiltration, with a standard blood flow rate of 200 mL/min and an ultrafiltrate volume of 100 L in 24 hrs, was performed with a highly permeable, large-surface cellulose triacetate membrane. Anticoagulation with anti-Xa bioequivalent doses of nadroparin and dalteparin was administered in the extracorporeal line before the filter. Blood was sampled for determination of coagulation variables before hemofiltration, 0.5, 2, 4, 6, and 12 hrs after starting the treatment, and at the end of the hemofiltration run. MEASUREMENTS AND MAIN RESULTS: Anti-Xa peak activity, time of anti-Xa peak activity, area under the curve for 0-3 hrs and filter survival time were not significantly different using nadroparin or dalteparin. When analyzing the patients according to the length of filter survival time, no relationship among anti-Xa peak activity, area under the curve for 0-3 hrs, and filter survival time was found. However, there was a strong trend toward a negative correlation between baseline platelet count and filter survival time (r2 = .11; p = .07). Mean blood urea nitrogen decreased from 81.0+/-31.9 to 41.1+/-21.2 mg/dL (p<.01) and mean creatinine decreased from 3.4+/-1.8 to 1.9+/-1.2 mg/dL (p<.01). There were no clinically important bleeding complications. CONCLUSIONS: Nadroparin and dalteparin are bioequivalent with respect to their anti-Xa activities. Using either drug, we did not find a difference in filter survival time during high volume, continuous venovenous hemofiltration. No relationship between anti-Xa activity and filter survival time could be found. However, there is a strong trend toward a negative correlation between baseline platelet count and filter survival time. This suggests that during high-volume, continuous venovenous hemofiltration, patients with a higher baseline platelet count might need a different anticoagulation regimen to obtain longer filter survival times. PMID- 10708178 TI - Comparison of Acute Physiology and Chronic Health Evaluation II (APACHE II) and Simplified Acute Physiology Score II (SAPS II) scoring systems in a single Greek intensive care unit. AB - OBJECTIVE: To evaluate Acute Physiology and Chronic Health Evaluation (APACHE) II and Simplified Acute Physiology Score (SAPS) II scoring systems in a single intensive care unit (ICU), independent from the ICUs of the developmental sample; and to compare the performance of APACHE II and SAPS II by means of statistical analyses in such a clinical setting. DESIGN: Prospective, cohort study. SETTING: A single ICU in a Greek university hospital. PATIENTS: In a time interval of 5 yrs, data for 681 patients admitted to our ICU were collected. The original exclusion criteria of both systems were employed. Patients <17 yrs of age were dropped from the study to keep compatibility with both systems. Eventually, a total of 661 patients were included in the analysis. INTERVENTIONS: Demographics, clinical parameters essential for the calculation of APACHE II and SAPS II scores, and risk of hospital death were recorded. Patient vital status was followed up to hospital discharge. MEASUREMENTS AND MAIN RESULTS: Both systems showed poor calibration and underestimated mortality but had good discriminative power, with SAPS II performing better than APACHE II. The evaluation of uniformity of fit in various subgroups for both systems confirmed the pattern of underprediction of mortality from both models and the better performance of APACHE II over our data sample. CONCLUSIONS: APACHE II and SAPS II failed to predict mortality in a population sample other than the one used for their development. APACHE II performed better than SAPS II. Validation in such a population is essential. Because there is a great variation in clinical and other patient characteristics among ICUs, it is doubtful that one system can be validated in all types of populations to be used for comparisons among different ICUs. PMID- 10708179 TI - Inflammatory cytokine response in patients with septic shock secondary to generalized peritonitis. AB - OBJECTIVES: The aims of this study were the following: a) to assess the proinflammatory cytokine (tumor necrosis factor [TNF]-alpha, interleukin [IL]-1, and IL-6) response in patients with septic shock secondary to generalized peritonitis; and b) to evaluate the influence of bacteremic status, type of peritonitis (acute perforation or postoperative), and peritoneal microbial status (mono- or polymicrobial) on cytokine expression and mortality. DESIGN: Prospective study. SETTING: Surgical intensive care unit of a university hospital. PATIENTS: Fifty-two consecutive patients with septic shock caused by generalized peritonitis. INTERVENTIONS: Routine blood tests, blood cultures, and cytokine assays were performed during the first 3 days after onset of shock. MEASUREMENTS AND MAIN RESULTS: Serum TNF-alpha and IL-6 concentrations were measured by using a radioimmunoassay, and IL-1 concentrations were measured by using ELISA. Median serum concentrations on day 1 were: TNF-alpha, 90 pg/mL; IL 1, 7 pg/mL; and IL-6, 5000 pg/mL. TNF-alpha and IL-6 concentrations decreased significantly between the first and third days of septic shock (p = .0001), whereas IL-1 concentrations remained low. The decrease in IL-6 tended to be more pronounced in the survivors group (p = .057). Median TNF-alpha serum concentrations were higher in bacteremic compared with nonbacteremic patients (151 vs. 73 pg/mL, p = .003). TNF-alpha, IL-1, and IL-6 serum concentrations and mortality were not different between acute perforation vs. postoperative peritonitis and mono- versus polymicrobial peritonitis. CONCLUSIONS: The systemic release of TNF-alpha and IL-6 during septic shock caused by generalized peritonitis was maximal on day 1 and decreased rapidly during the next days. No systemic release of IL-1 was observed. IL-6 serum concentrations remained higher in patients who subsequently died. Among the different features of peritonitis studied, only bacteremia influenced the systemic cytokine response (higher TNF alpha). PMID- 10708180 TI - Sequential single doses of cisapride, erythromycin, and metoclopramide in critically ill patients intolerant to enteral nutrition: a randomized, placebo controlled, crossover study. AB - OBJECTIVE: To evaluate the comparative efficacy of enteral cisapride, metoclopramide, erythromycin, and placebo for promoting gastric emptying in critically ill patients with intolerance to gastric enteral nutrition (EN). DESIGN: A randomized, crossover study. SETTING: Adult medical intensive care unit at a university-affiliated private hospital and trauma intensive care unit at a university teaching hospital. PATIENTS: Ten adult, critically ill, mechanically ventilated patients not tolerating a fiber-containing EN product defined as a single aspirated gastric residual volume >150 mL or two aspirated gastric residual volumes >120 mL during a 12-hr period. INTERVENTIONS: Patients received 10 mg of cisapride, 200 mg of erythromycin ethylsuccinate, 10 mg of metoclopramide, and placebo as 20 mL of sterile water every 12 hrs over 48 hrs. Acetaminophen solution (1000 mg) was administered concurrently. Gastric residual volumes were assessed, and plasma acetaminophen concentrations were serially determined by TDx between 0 and 12 hrs to evaluate gastric emptying. MEASUREMENTS AND MAIN RESULTS: Gastric residual volumes during the study were not significantly different between agents. No differences in area under the concentration vs. time curve or elimination rate constant were identified between agents. Metoclopramide and cisapride had a significantly shorter mean residence time of absorption than erythromycin (6.3+/-4.5 [SEM] mins and 10.9+/-5.8 vs. 30.1+/-4.5 mins, respectively [p<.05]). Metoclopramide (9.7+/-15.3 mins) had a significantly shorter time to peak concentration compared with erythromycin and placebo (60.7+/-8.1 and 50.9+/-13.5 mins, respectively [p<.05]). The time to onset of absorption was significantly shorter for metoclopramide vs. cisapride (5.7+/-4.5 vs. 22.9+/-5.7 mins [p<.05]). CONCLUSION: In critically ill patients intolerant to EN, single enteral doses of metoclopramide or cisapride are effective for promoting gastric emptying in critically ill patients with gastric motility dysfunction. Additionally, metoclopramide may provide a quicker onset than cisapride. PMID- 10708181 TI - Changes of the hemostatic network in critically ill patients--is there a difference between sepsis, trauma, and neurosurgery patients? AB - OBJECTIVE: To study the time course of coagulation data in intensive care patients. DESIGN: Prospective, descriptive study. SETTING: Clinical investigation on a surgical and neurosurgical intensive care unit of a university hospital. PATIENTS: Fifteen patients with severe trauma (injury severity score, 15 to 25), 15 sepsis patients secondary to major surgery, and 15 neurosurgery patients (cancer surgery) were studied. INTERVENTIONS: Standardized intensive care therapy. MEASUREMENTS AND MAIN RESULTS: Standard coagulation data and molecular markers of coagulation activation and fibrinolytic activity (soluble thrombomodulin, protein C, free protein S, thrombin/antithrombin III complex, plasmin-alpha 2-antiplasmin complex, tissue plasminogen activator, platelet factor 4, beta-thromboglobulin were measured from arterial blood samples on the day of admission to the intensive care unit (trauma/neurosurgery patients) or on the day of diagnosis of sepsis (baseline value) and serially during the next 5 days. Antithrombin III, fibrinogen, and platelet counts were highest in neurosurgery patients but without significant differences between sepsis and trauma patients. Thrombin/antithrombin III complex increased in the sepsis patients (from 22.6+/-4.2 microg/L to 39.9+/-6.8 microg/L), but decreased in trauma (from 40.2+/-5.1 microg/L to 17.6+/-4.0 microg/L) and neurosurgery patients (from 28.2+/-4.2 microg/L to 16.2+/-3.8 microg/L). Tissue plasminogen activator increased in the sepsis patients (from 14.4+/-3.9 microg/L to 20.7+/ 3.8 microg/mL) and remained almost unchanged in the other two groups. Soluble thrombomodulin plasma concentration increased significantly in the sepsis group (max, 131.8+/-22.5 ng/mL), while it remained elevated in the trauma (max, 75.5+/ 5.9 ng/mL) and was almost normal in the neurosurgery patients. Protein C and free protein S remained decreased only in the sepsis group. CONCLUSIONS: Alterations of the hemostatic network were seen in all three groups of critically ill patients. Hemostasis normalized in the neurosurgery patients and posttraumatic hypercoagulability recovered within the study period. By contrast, monitoring of molecular markers of the coagulation process demonstrated abnormal hemostasis in the sepsis patients during the entire study period indicating ongoing coagulation disorders and abnormalities in fibrinolysis in these patients. PMID- 10708182 TI - Coagulation system and platelets are fully activated in uncomplicated sepsis. AB - OBJECTIVE: To test the hypothesis that the coagulation system and platelets are activated in sepsis, the uncomplicated and usually earliest stage of the septic process, and to compare the findings detected in sepsis with those found in severe sepsis and septic shock. DESIGN: Prospective study comparing patients with sepsis, severe sepsis, and septic shock, and healthy volunteers. SETTING: General intensive care unit in a tertiary university hospital. PATIENTS: Seventy-four consecutive septic patients (45 with sepsis, 15 with severe sepsis, and 14 with septic shock). Fourteen healthy volunteers served as control subjects. INTERVENTION: None. MEASUREMENTS AND MAIN RESULTS: After blood sampling, molecular activation markers of coagulation (prothrombin fragments 1 and 2, fibrinopeptide A, thrombin-antithrombin complexes, and monomers of fibrin) and of platelets (beta-thromboglobulin and platelet factor 4), several coagulation factors, global tests of coagulation (prothrombin time and activated partial thromboplastin time), and platelet count (PTL) were measured. In sepsis, prothrombin fragments 1 and 2, fibrinopeptide A, thrombin-antithrombin complexes, and monomers of fibrin were increased to 2.52+/-0.21 nmol/L, 20.88+/-2.52 ng/mL, 33.8+/-2.9 microg/L, and 69% positive, respectively, compared with control subjects (0.86+/-063 nmol/L, 1.14+/-0.15 ng/mL, 16.07+/-1.01 microg/L, and 0%, respectively). Beta-Thromboglobulin and the beta-thromboglobulin-to-platelet factor 4 ratio were also increased to 107.87+/-11.87 IU/mL and 8.86+/-1.06, compared with controls (18.36 +/-2.99 IU/mL and 2.67+/-0.52, respectively). With the exception of a decrease in factor XII and an increase in fibrinogen, coagulation factors, global coagulation tests, and PTL were not changed in sepsis. In severe sepsis and mainly in septic shock, coagulation factors were markedly decreased, global coagulation tests were prolonged, and PTL was reduced. All changes were independent of the causative infectious pathogen. CONCLUSION: Coagulation system and platelets are strongly activated in sepsis. In this stage, only factor XII is decreased. In contrast, in severe sepsis and mainly in septic shock, most of the coagulation factors are depleted, PTL is decreased, and global coagulation tests are prolonged, indicating exhaustion of hemostasis. Finally, Gram-positive, Gram-negative, and other microorganisms produce identical impairment of coagulation. PMID- 10708183 TI - Procalcitonin behaves as a fast responding acute phase protein in vivo and in vitro. AB - OBJECTIVES: Procalcitonin (PCT) is a 13 kD protein of which plasma concentrations are strongly increased in inflammatory states. PCT concentrations are claimed to have a more powerful discriminatory value for bacterial infection than the acute phase proteins serum amyloid A (SAA) or C-reactive protein (CRP). The source of production and its mechanism of induction are unknown. We investigated the inducibility of PCT both in vivo and in vitro and compared the behavior of PCT with those of SAA and CRP. DESIGN: A prospective descriptive patient sample study and a controlled liver tissue culture study. SETTING: A university hospital. PATIENTS: Cancer patients who were treated with human tumor necrosis factor-alpha (rhTNF-alpha; 5 patients) or interleukin-6 (rhIL-6; 7 patients). MEASUREMENTS AND MAIN RESULTS: Serial serum samples were collected for analysis of concentrations of PCT, SAA, and CRP. In the TNF-alpha group, frequent sampling was performed on the first day to allow analysis of initial responses. In a human liver slice model, the release of PCT, SAA, and CRP was measured on induction with rhTNF alpha and rhIL-6 for 24 hrs. We found that PCT displayed acute phase reactant behavior in vivo after administration of both rhTNF-alpha and rhIL-6. After rhTNF alpha-administration, PCT reached half-maximal concentrations within 8 hrs, 12 hrs earlier than either SAA or CRP did. PCT, SAA, and CRP were produced in detectable quantities by liver tissue in vitro. PCT production by liver slices was enhanced after stimulation with rhTNF-alpha or rhIL-6; SAA and CRP concentrations were elevated after stimulation with rhTNF-alpha. CONCLUSIONS: We found that PCT and acute phase proteins such as CRP are induced by similar pathways. The liver appears to be a major source of PCT production. Thus, PCT may be considered an acute phase protein. The different kinetics of PCT, rather than a fundamentally different afferent pathway, may explain its putative diagnostic potential to discriminate bacterial infection from other causes of inflammation. PMID- 10708184 TI - Accuracy of mucosal pH and mucosal-arterial carbon dioxide tension for detecting mesenteric hypoperfusion in acute canine endotoxemia. AB - OBJECTIVE: To determine the level of mucosal-arterial Pco2 (Pco2 gap) that is both sensitive and specific for the detection of mesenteric hypoperfusion as defined by either a >50% reduction in portal blood flow or release of lactate by the gut. DESIGN: Animal experiment. SUBJECTS: Seven anesthetized, intubated, mechanically ventilated, and surgically instrumented mongrel dogs. INTERVENTION: Escherichia coli endotoxin (1 mg/kg) given intravenously for 5 mins. MEASUREMENTS AND MAIN RESULTS: Tonometric Pco2, arterial blood gases, arterial and portal venous lactates, and portal and systemic hemodynamic variables were measured. Mucosal pH (pHi) was calculated according to the manufacturers' instructions. From these data, receiver operating characteristics were calculated. Although animals were resuscitated to maintain a constant cardiac output, portal flow decreased from 350+/-101 to 152+/-75 mL/min (p<.01) and the gut released lactate into the portal circulation in all animals. Pco2 gap increased from 13.1+/-3.9 to 40.2+/-39.2 torr (p<.01) and was inversely correlated with portal blood flow (r2 = .20; p<.05). For detection of a >50% reduction in portal blood flow, a Pco2 gap of 20 torr yielded a maximum accuracy of 67% (sensitivity, 55%; specificity, 73%) and was less accurate than a pHi of 7.20, which yielded a maximum accuracy of 76% (sensitivity, 90%; specificity, 70%), although this difference was not significant (p = .24). There was also a correlation between pHi and portal blood flow (r2 = .31; p<.01). For detection of lactate release by the gut, a Pco2 gap of 20 torr was also 67% accurate (sensitivity, 53%; specificity, 78%), whereas a pHi of 7.10 achieved an accuracy of 64% (sensitivity, 40%; specificity, 83%), which was not significantly different. CONCLUSION: Pco2 gap measurements are neither sensitive nor specific for mesenteric hypoperfusion with regard to total gut blood flow reductions of >50% or the release of lactate into the portal circulation. PMID- 10708185 TI - Dobutamine restores intestinal mucosal blood flow in a porcine model of intra abdominal hyperpressure. AB - OBJECTIVE: To assess the effects of dopamine and dobutamine administration on the systemic and mesenteric (macro- and microvascular) circulatory disturbances induced by intra-abdominal hyperpressure. DESIGN: Prospective, randomized study. SETTING: Animal research laboratory in a university hospital. SUBJECTS: Twenty five pigs of either gender, weighing 30-35 kg. INTERVENTIONS: Animals were anesthetized, and their lungs were mechanically ventilated. Pulmonary artery flotation and carotid artery catheters were inserted for hemodynamic monitoring and blood sampling. A perivascular flow probe was placed around the superior mesenteric artery, and a laser Doppler probe was positioned in the lumen of the ileum to measure arterial and intestinal mucosal blood flows, respectively. CO2 was insufflated into the peritoneal cavity to reach an intra-abdominal pressure of 15 mm Hg, and 60 mins later, animals received dopamine (5 microg/kg/min; n = 10), dobutamine (5 microg/kg/min; n = 10), or saline (n = 5) for 30 mins. MEASUREMENTS AND MAIN RESULTS: Peritoneal CO2 insufflation induced significant increases in heart rate, arterial pressure, and systemic vascular resistance with concomitant decreases in cardiac output and superior mesenteric arterial and intestinal mucosal blood flows. Although dobutamine infusion reversed the decrease in cardiac output, it failed to restore superior mesenteric artery blood flow; however, intestinal mucosal blood flow returned to baseline levels. Dopamine also attenuated the decrease in cardiac output, but it had no beneficial effect on splanchnic hemodynamic variables. CONCLUSIONS: Low-dose infusion of dobutamine, but not dopamine, corrects the intestinal mucosal perfusion impairment induced by moderate increases in intra-abdominal pressure. PMID- 10708186 TI - Optimal level of pressure support ventilation for recovery from diaphragmatic fatigue in rabbits. AB - OBJECTIVE: To determine whether there is an optimal level of pressure support ventilation (PSV) for recovery from acute diaphragmatic fatigue. DESIGN: Prospective laboratory trial. SETTING: Experimental laboratory. SUBJECTS: Twenty healthy adult New Zealand White rabbits. INTERVENTIONS: Diaphragmatic fatigue was induced with 50 Hz of phrenic nerve stimulation for 30 mins. Recovery was compared between inspiratory load + PSV of 0 cm H2O (L0), inspiratory load + PSV of 60 cm H2O (L60), inspiratory load + PSV of 80 cm H2O (L80), and PSV of 0 cm H2O without inspiratory load (SB) for 90 mins immediately after the end of the fatigue-inducing procedure. To add inspiratory load during the recovery phase, three pressure threshold valves, each having an opening pressure of -20 cm H2O, were used. MEASUREMENTS AND MAIN RESULTS: After the fatigue-inducing procedure, diaphragmatic electromyogram and transdiaphragmatic pressure remained at baseline in both SB and L60, decreased in L80, and increased in L0. Recovery was assessed by abdominal cavity pressure (Pabd) generated by high-frequency (100 Hz) and low frequency (20 Hz) stimulation. Pabd at 100 Hz recovered to baseline in L60 and SB but not in L0 and L80 (69.1%, 81.3%, 100.3%, and 100.7% of the baseline at 90 mins for L0, L80, L60, and SB, respectively). Pabd at 20 Hz did not differ among ventilatory settings. CONCLUSION: There is an optimal range of PSV assist level to improve recovery from diaphragmatic fatigue. Recovery was hampered not only by inadequate PSV but also by excessive PSV. PMID- 10708187 TI - Partial liquid ventilation decreases serum tumor necrosis factor-alpha concentrations in a rat acid aspiration lung injury model. AB - OBJECTIVE: To examine the hypothesis that partial liquid ventilation (PLV) with perfluorocarbon would decrease serum tumor necrosis factor-alpha concentrations in a rat acid aspiration lung injury model. DESIGN: Prospective, controlled animal study. SETTINGS: Research laboratory in a university setting. SUBJECTS: Male Sprague-Dawley rats. INTERVENTIONS: Treatment with intratracheal perflubron or control mechanical ventilation beginning 30 mins after acid aspiration. MEASUREMENTS AND MAIN RESULTS: PLV with perfluorocarbon compared with control ventilation resulted in significantly greater mean arterial blood pressures at 3 and 4 hrs and greater arterial Po2 at all times. Serum tumor necrosis factor alpha at 2, 3, and 4 hrs was significantly less than that observed in the control group (4-hr values: 80+/-64 pg/mL vs. 658+/-688 pg/mL; p<.05), although no significant difference in tracheal fluid tumor necrosis factor-alpha concentrations (1425+/-1347 pg/mL vs. 2219+/-1933 pg/mL) was found. CONCLUSION: We conclude that the effects of PLV with perfluorocarbon can extend beyond improvements in pulmonary physiology and that PLV may be beneficial in reducing systemic sequelae of acute lung injury and inflammation. PMID- 10708188 TI - Lidocaine attenuates acute lung injury induced by a combination of phospholipase A2 and trypsin. AB - OBJECTIVE: Acute severe pancreatitis is often associated with acute lung injury, including acute respiratory distress syndrome. Acute lung injury induced by phospholipase A2 (PLA2) or trypsin, a pancreatic enzyme, is an experimental model resembling acute respiratory distress syndrome. Neutrophils and platelets are thought to play a pivotal role in the pathogenesis of acute respiratory failure. Lidocaine inhibits some aspects of neutrophil and platelet functions. We conducted the current study to assess the effects of pretreatment with lidocaine on acute lung injury induced by a combination of PLA2 and trypsin. DESIGN: Prospective, randomized animal study. SETTING: University research laboratory. SUBJECTS: Twenty-one adult male Japanese White rabbits (weight range, 2.0-2.4 kg). INTERVENTIONS: The animals were mechanically ventilated with a tidal volume of 10 mL/kg and an Fio2 of 0.4, and thereafter, they were randomly assigned to three groups. Acute lung injury was induced by a combination of PLA2 (1000 units/kg/hr) and trypsin (5000 units/kg/hr) infused intravenously for 4 hrs. Immediately before induction of the acute lung injury, the lidocaine treatment group received intravenous lidocaine (2 mg/kg bolus followed by 2 mg/kg/hr) until they were killed. In the nontreatment group, saline was given instead of lidocaine. Rabbits in the nonlung-injury group received saline infusion instead of the pancreatic enzymes. MEASUREMENTS AND MAIN RESULTS: During the experimental period (4 hrs), arterial blood gases, lung mechanics, and peripheral neutrophil and platelet counts were measured. Immediately after killing, the wet weight/dry weight ratio of the lung was recorded. Light microscopic findings (lung injury score and number of neutrophils) were compared between the three groups. The combination of PLA2 and trypsin decreased Pao2, lung compliance, and peripheral counts of neutrophils and platelets and increased alveolar/arterial oxygen tension difference, lung resistance, wet weight/dry weight ratio, and the number of neutrophils in the lung. Lidocaine treatment attenuated these changes. The two pancreatic enzymes caused extensive morphologic lung damage, which was lessened by lidocaine. CONCLUSIONS: We conclude that pretreatment with intravenous lidocaine attenuated the lung injury induced by the pancreatic enzymes. However, further studies are required to determine whether this drug has a therapeutic effect once the lung injury has developed. PMID- 10708189 TI - Site-specific effect of guanosine 3',5'-cyclic monophosphate phosphodiesterase inhibition in isolated lamb lungs. AB - OBJECTIVE: To determine the effect of combining inhaled nitric oxide (NO) with an inhibitor of guanosine 3',5'-cyclic monophosphate-specific phosphodiesterase on total and segmental lung resistances. STUDY DESIGN: A controlled laboratory study in isolated blood-perfused lungs prepared from lambs. SETTING: Animal research facility affiliated with a university teaching hospital. SUBJECTS: Five newborn lambs at <48 hrs of life. INTERVENTIONS: Isolated blood-perfused lungs were prepared and treated with indomethacin (40 microg/mL) to inhibit prostaglandin synthesis. After a baseline period of normoxia (28% oxygen), pulmonary hypertension was induced with the thromboxane mimetic U46619 (0.1-0.4 microg/kg/min). During pulmonary hypertension, lungs were studied with inhaled NO only, with infusion of zaprinast only (0.25 mg/kg bolus and 0.05 mg/kg/min infusion), and with a combination of the two. For each study condition, the total pressure decrease across the lung was measured, and the inflow-outflow occlusion technique was used to partition the total pressure gradient measured at constant flow (100 mL/kg/min) into gradients across relatively noncompliant large arteries and veins and more compliant small arteries and veins. MEASUREMENTS AND MAIN RESULTS: U46619 infusion produced significant pulmonary vasoconstriction. The combination of inhaled NO and zaprinast decreased the total pressure decrease across the lung significantly more than NO alone. This effect was primarily attributable to a significantly greater decrease in gradient across the small artery segment after inhaled NO and zaprinast compared with NO alone. CONCLUSIONS: Guanosine 3',5'-cyclic monophosphate phosphodiesterase inhibition with zaprinast enhances the effect of inhaled NO, particularly in conditions in which small arteries represent the site of resistance. Phosphodiesterase inhibition may be a promising adjunct to inhaled NO for the treatment of persistent pulmonary hypertension. PMID- 10708190 TI - Spontaneous high systemic oxygen delivery increases survival rate in awake sheep during sustained endotoxemia. AB - OBJECTIVE: To study the natural evolution of systemic oxygen delivery (Do2) and oxygen consumption (Vo2) in sheep infused with low or high doses of endotoxin. DESIGN: Prospective, controlled experimental study. SETTING: Animal research laboratory at a medical university. SUBJECTS: Twenty-nine chronically instrumented awake sheep (25-35 kg). INTERVENTIONS: Awake animals were continuously infused with saline (n = 8) or two doses of Escherichia coli endotoxin (20 or 40 ng/kg/min; n = 21) for 72 hrs. No attempt was made to increase Do2, but respiratory failure was treated by mechanical ventilation and metabolic acidosis was corrected. MEASUREMENTS AND MAIN RESULTS: The mortality rate was 25% in the group infused with the low dose and 89% in the group infused with the high dose of endotoxin. During the first 12 hrs of endotoxemia, both surviving (S group; n = 10) and nonsurviving (NS group; n = 11) sheep developed similar pulmonary hypertension, left ventricular failure, and hypotension with low systemic vascular resistance. However, S sheep had less interstitial lung edema (pulmonary lymph protein clearance at 8 hrs was 13+/-3 mL/hr vs. 27+/-6 mL/hr in the NS group and 4+/-1 mL/hr in the control group). During this early phase of endotoxemia, Do2, Vo2, and oxygen extraction ratio did not change significantly in any group. After this phase, animals that ultimately survived had a persistent hyperdynamic syndrome with high cardiac output and hypotension. In this group, the Do2 increase was greater than the Do2 measured in controls and remained steady up to 48 hrs after the start of the endotoxin infusion. Because systemic Vo2 did not change significantly, oxygen extraction ratio decreased progressively to values less than those measured in controls. In contrast, animals that ultimately died had a hypotensive and normokinetic syndrome associated with pulmonary hypertension, persistent depressed left ventricular function, hypothermia, and a progressive deterioration of gas exchange. Systemic Do2 was not significantly different from that in the control group. In contrast, Vo2 decreased progressively to values significantly lower than those measured in controls and remained low until death. CONCLUSIONS: Our results indicate that in the absence of treatment such as fluid challenge or inotropic drugs in sheep infused with endotoxin, the occurrence of spontaneous hyperdynamic syndrome and high Do2 improves the survival rate. PMID- 10708191 TI - Hepatocyte growth factor modulates the hepatic acute-phase response in thermally injured rats. AB - OBJECTIVE: Hepatocyte growth factor (HGF) has been shown to modulate the acute phase response in vitro. The specific in vivo role of HGF in this multifactorial response, however, remains unknown. This study examines the effects of exogenous HGF on the acute-phase response in thermally injured rats. DESIGN: Prospective, randomized, laboratory study. SETTINGS: Shriners Hospital for Children and University of Texas Medical Branch laboratories. SUBJECTS: Fifty-six male Sprague Dawley rats (weight range, 300-325 g). INTERVENTION: Animals received a 60% total body surface area third-degree scald burn and were randomly divided to receive either 400 microg/kg/day i.v. HGF or saline (control). MEASUREMENTS AND MAIN RESULTS: Serum acute-phase proteins, cytokines, and insulin-like growth factor (IGF)-I concentrations, as well as liver weight, protein and triglyceride content, IGF-I concentrations, and cytokine gene expression were measured 1, 2, 5, or 7 days after burn. Serum albumin was increased on days 2, 5, and 7 after burn, and transferrin was increased on day 7 after burn in HGF-treated rats compared with controls (p<.05). HGF increased alpha2-macroglobulin concentrations on postburn days 2, 5, and 7 compared with controls (p<.05). Serum interleukin-6 and tumor necrosis factor-alpha were significantly higher within 2 days of burn in rats treated with HGF (p<.05). HGF increased the hepatic gene expression of tumor necrosis factor-alpha compared with controls (p<.05). Serum IGF-I decreased in rats receiving HGF 1, 2, and 5 days after burn, whereas liver IGF-I concentrations were higher on days 1 and 7 after burn compared with controls (p<.05). Hepatic protein concentrations were higher in the HGF group compared with controls on postburn days 1, 2, and 7, with a concomitant increase in total liver weight (p<.05). HGF exerted a strong mitogenic effect on hepatocytes 1 and 2 days after thermal injury compared with controls (p<.05). CONCLUSIONS: These findings suggest that HGF modulates the acute-phase response in vivo after burn and causes changes in liver morphology. PMID- 10708192 TI - Bedside monitoring of cerebral blood flow in patients with acute hemispheric stroke. AB - OBJECTIVE: To test the practicability of a new double indicator dilution method for bedside monitoring of cerebral blood flow (CBF) and to assess the clinical value of CBF monitoring as a prognostic tool for outcome and in therapy of elevated intracranial pressure (ICP) in patients with acute hemispheric stroke. DESIGN: Prospective study. Clinical evaluation of a new method. SETTING: Neurological intensive care unit of a university hospital. PATIENTS: Ten patients with acute complete middle cerebral artery territory- or hemispheric infarctions. INTERVENTIONS: Two combined fiberoptic thermistor catheters were placed in the right jugular bulb and in the thoracic aorta. Central venous injections of ice cold indocyanine green dye were performed. CBF was estimated by calculating the mean transit times of the cold bolus and dye. MEASUREMENTS AND MAIN RESULTS: A total of 104 reproducible CBF measurements were obtained. No complications associated with the method were observed. Twelve pairs of measurements were performed within 30 mins with unchanged clinical conditions. The standard deviation of repeated measurements was 2.7 mL/100 g/min; the interrater reliability was between 0.95 and 0.99. The median CBF in patients who died (n = 4) was lower (27 mL/100g/min) than in those who survived (n = 6) (45 mL/100g/ min). Patients who died more frequently had low CBF values of <30 mL/100g/min (22 of 38; 58%) than patients who survived (10 of 54; 19%). A total of 37 CBF measurements were done during ICP elevation of >20 mm Hg. In patients who survived, ICP elevations were only associated with low CBF values in 5 of 26 events; whereas in patients who died, ICP elevations were associated with low CBF values in 8 of 11 events. CONCLUSIONS: The new double indicator dilution technique may be suitable for serial bedside CBF measurement. It is easy to perform and can be rapidly repeated in the ICU environment. Validation of the method by comparison with standard methods is needed. The preliminary data indicate that bedside monitoring of CBF may give prognostic information for outcome and may guide therapy of elevated ICP in patients with malignant hemispheric infarction. PMID- 10708193 TI - Long-term outcomes of burned children after in-hospital cardiac arrest. AB - OBJECTIVE: Cardiopulmonary resuscitation (CPR) in severely burned patients experiencing cardiac arrest (CA) has been considered by some as futile. The objective of this article is to report predisposing factors and the outcomes of burned children experiencing in-hospital CA at our institution. DESIGN: The records of 595 children admitted from 1985 to 1998 with burns covering >35% of their total body surface area were reviewed. Thirty-four children receiving CPR after in-hospital CA were studied for predisposing factors and long-term outcomes. SETTING AND PATIENTS: Shriners Burns Hospital. Burned children of both genders, 0.5-19 yrs of age, who experienced in-hospital CA and received CPR. INTERVENTION: Standard burn care and CPR. MEASUREMENTS AND MAIN RESULTS: Predisposing factors of CA, mortality, and long-term outcomes were measured. The incidence of CA in burned children with burns on >35% total body surface area was 5.7%. No significant difference in age or burn size could be shown between long term CA survivors (n = 17) and nonsurvivors (n = 17). CPR was successful (defined as survival for at least 1 day after CA) in 22 of 34 children (65%), with 17 of the 22 survivors (77%) experiencing long-term survival, currently from 2-14 yrs. Significant predisposing factors of CA were sepsis, identified in 53% of the nonsurvivors vs. 12% of the survivors (p<.05), and delayed fluid resuscitation (>2 hrs after burn injury), identified in 82% of the nonsurvivors vs. 6% of the survivors (p<.001). There was only one morbid long-term survivor. This survivor was diagnosed as having anoxic brain injury with persistent neurologic deficiencies. CONCLUSION: In this study, 50% of the burned children experiencing CA are long-term survivors. We suggest that all burned children with CA should be afforded cardiopulmonary resuscitation. PMID- 10708194 TI - Venovenous versus venoarterial extracorporeal life support for pediatric respiratory failure: are there differences in survival and acute complications? AB - OBJECTIVES: To examine the Extracorporeal Life Support Organization (ELSO) registry database of infants and children with acute respiratory failure to compare outcome and complications of venovenous (VV) vs. venoarterial (VA) Extracorporeal Life Support (ECLS). DESIGN: Retrospective cohort study. SETTING: ELSO registry for pediatric pulmonary support. PATIENTS: All nonneonatal pediatric pulmonary support ECLS cases treated at U.S. centers and reported to the ELSO registry as of July 1997. Patients were excluded if they had one or more of the following diagnoses: hematologic-oncologic, cardiac, abdominal surgical, burn, metabolic, airway, or immunodeficiency disorder. INTERVENTIONS: Venoarterial or venovenous extracorporeal life support for severe pulmonary failure. MEASUREMENTS AND MAIN RESULTS: From 1986 to June of 1997, 763 pediatric patients met the inclusion criteria. Overall, 595 were initially managed with VA bypass, and 168 with VV bypass. The VA group was younger (mean +/- SD, 26.1+/ 42.2 months for VA vs. 63.5+/-68.7 months for VV) and smaller (11.8+/-15.1 kg vs. 22.9+/-23.8 kg) (p<.001). There were no differences between groups in number of days on mechanical ventilation before ECLS, number of hours on ECLS, or number of hours on mechanical ventilation post-ECLS in survivors. Mean pH and Paco2 values, positive end-expiratory pressure, and mean airway pressure just before placing the patient on ECLS were also similar. VA-treated patients had higher Fio2 requirements (p = .034), lower Pao2 (p = .047), and lower Pao2/Fio2 ratio (p = .014) just before cannulation. There was a trend of higher peak inspiratory pressure in VA-treated patients (p = .053). Overall, survival rate was not different for the two groups (55.8% for VA vs. 60.1% for VV; p = .33). Central nervous system complications were not different between the two groups. Examination of the same variables was then conducted after dividing the patients into four subgroups. There were no significant differences in survival or complications during bypass between VV and VA modes of ECLS in any subgroup. Stepwise logistic regression modeling was performed to control for variables associated with the outcome survival for VV and VA-treated groups, and variables measured before bypass were identified as being associated with improved survival. There was a trend of improved survival in the VV-treated patients (p = .12). CONCLUSIONS: Overall survival of pediatric patients with acute respiratory failure supported by VA or VV ECLS was comparable. A randomized clinical trial may be useful in clarifying these observations. PMID- 10708195 TI - Impact of multiple organ system dysfunction and nosocomial infections on survival of children treated with extracorporeal membrane oxygenation after heart surgery. AB - OBJECTIVES: To evaluate whether cardiac and noncardiac variables may be used to predict survival in children treated with extracorporeal membrane oxygenation (ECMO) after cardiopulmonary bypass and to determine when to discontinue ECMO support. DESIGN: Retrospective review. SETTING: Neonatal and pediatric intensive care units of Kosair Children's Hospital. PATIENTS: Fifty-nine children treated with ECMO after cardiopulmonary bypass from 1987 through 1996. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Medical, nursing, operative, and perfusion records for each patient were reviewed. The primary outcome measure was survival to hospital discharge. Cardiac and noncardiac variables were recorded at serial times. Nineteen of 59 patients (32%) survived. No cardiac variable was a clinically useful predictor of survival or marker for when to discontinue ECMO. Among the noncardiac variables, progressive multiple organ system dysfunction and development of a nosocomial infection were significantly associated with nonsurvival. No patient with a positive blood culture (n = 3) within the first 24 hrs of ECMO survived, and 21 of 24 children with a positive culture from any site during ECMO died (p = .007). Despite their higher mortality, children with positive cultures were supported with ECMO significantly longer than those with negative cultures (275+/-168 vs. 135+/-108 hrs, respectively; p = .0004). For all patients, the longest duration of ECMO that resulted in survival was 256 hrs. For children with a positive culture, the longest duration of support that resulted in survival was 200 hrs. CONCLUSIONS: Support with ECMO beyond 256 hrs was not associated with survival. Progressive multiple organ system dysfunction and nosocomial infections have a negative impact on survival. Serious consideration should be given to discontinuing ECMO support whenever there is a progressive increase in the number of abnormally functioning organ systems, a nosocomial infection occurs, or native cardiac function has not improved significantly by 250 hrs of ECMO support. PMID- 10708196 TI - Comparison of infrared thermometer with thermocouple for monitoring skin temperature. AB - OBJECTIVE: To test the hypothesis that the infrared thermometer (Genius) is comparably useful with thermocouples that are routinely used for skin temperature monitoring. DESIGN: Prospective, controlled, not blinded study. SETTING: Operating room of a university hospital. SUBJECTS: Ten healthy male volunteers. INTERVENTIONS: Volunteers were minimally clothed and were initially warmed by a forced air warmer until they became vasodilated at the finger and the foot for approximately 30 mins. Subsequently, they were kept in the room with no blanket. MEASUREMENTS AND MAIN RESULTS: Skin temperatures were measured continuously with the Mon-a-Therm thermocouple and were also measured with the Genius thermometer just before and after the warming and subsequently every 10 mins for 70 mins. Forearm and finger-tip skin temperatures and skin-surface temperature gradients (from arm to finger and from calf to toe) measured by the Genius thermometer were compared with those measured by the Mon-a-Therm thermocouple using linear regression and Bland and Altman statistics. Forearm temperature and finger-tip temperature ranged from approximately 31 degrees to approximately 36.5 degrees C (87.8-97.7 degrees F) and approximately 22.5 degrees to approximately 36 degrees C (72.5-96.8 degrees F), respectively. Gradients (from arm to finger and from calf to toe) ranged from approximately -3 degrees to approximately 10 degrees C (26.6-50.0 degrees F) and approximately -3 degrees to approximately 11 degrees C (26.6-51.8 degrees F), respectively. Correlations between the temperatures measured by the Genius thermometer and those by the Mon-a-Therm thermocouple were similar and reliable. The correlation coefficients were as follows: 0.78 at forearm, 0.97 at finger-tip, and 0.97 at skin-surface temperature gradients. CONCLUSIONS: The infrared thermometer with a special probe is useful to measure the change of skin-surface temperatures and to evaluate the severity of shock in patients. PMID- 10708197 TI - Does programmed cell death (apoptosis) play a role in the development of multiple organ dysfunction in critically ill patients? a review and a theoretical framework. AB - OBJECTIVES: To critically review the current understanding of the pathophysiologic events leading to the development of secondary multiple organ dysfunction (MODS) in critical illness and to examine the role of apoptosis (programmed cell death) as a mechanism involved in the progression of MODS. DATA SOURCES: Research and review articles published since 1982 on the pathophysiology of MODS, particularly the role of cytokines, reactive oxygen species, heat shock proteins, and apoptosis. Research and review articles on the physiology of apoptosis. Articles include human/animal and in vitro/in vivo studies. DATA EXTRACTION: The most prevalent mediating factors of MODS were examined for their potential to induce apoptosis, as reported in the literature. The combination of several of the above factors was also examined in terms of apoptosis-triggering potential. DATA SYNTHESIS: Specific pathophysiologic conditions related to the onset of MODS have been shown to affect apoptotic rates in organ tissue cells and their respective endothelial cells in animal and in vitro models. These conditions include the following: a) increased release of inflammation-related cytokines; b) increased production of oxygen free radicals associated with ischemia/reperfusion injury and states of low tissue perfusion; c) expression and release of heat shock proteins from tissue cells and the liver; d) elevated glucocorticoid concentrations after adrenal cortex activation; and e) release of bacterial products into the systemic circulation. CONCLUSION: The most important MODS-related pathophysiologic conditions known to date have been shown to affect programmed cell death rates in almost all cell types. Organ-specific cell death involving both parenchymal and microvasculature endothelial cells is conceivably underlying organ dysfunction. The hypothesis that increased apoptotic rates are involved in organ dysfunction may provide a unifying theory for the pathophysiology of MODS. PMID- 10708198 TI - Medical students can learn the basic application, analytic, evaluative, and psychomotor skills of critical care medicine. AB - OBJECTIVE: To determine whether fourth-year medical students can learn the basic analytic, evaluative, and psychomotor skills needed to initially manage a critically ill patient. DESIGN: Student learning was evaluated using a performance examination, the objective structured clinical examination (OSCE). Students were randomly assigned to one of two clinical scenarios before the elective. After the elective, students completed the other scenario, using a crossover design. SETTING: Five surgical intensive care units in a tertiary care university teaching hospital. PARTICIPANTS: Forty fourth-year medical students enrolled in the critical care medicine (CCM) elective. INTERVENTIONS: All students evaluated a live "simulated critically ill" patient, requested physiologic data from a nurse, ordered laboratory tests, received data in real time, and intervened as they deemed appropriate. MEASUREMENTS AND MAIN RESULTS: Student performance of specific behavioral objectives was evaluated at five stations. They were expected to a) assess airway, breathing, and circulation in appropriate sequence; b) prepare a manikin for intubation, obtain an acceptable airway on the manikin, demonstrate bag-mouth ventilation, and perform acceptable laryngoscopy and intubation; c) provide appropriate mechanical ventilator settings; d) manage hypotension; and e) request and interpret pulmonary artery data and initiate appropriate therapy. OSCEs were videotaped and reviewed by two faculty members masked to time of examination. A checklist of key behaviors was used to evaluate performance. The primary outcome measure was the difference in examination score before and after the rotation. Secondary outcomes included the difference in scores at each rotation. The mean preelective score was 57.0%+/ 8.3% compared with 85.9%+/-7.4% (p<.0001) after the elective. Significant improvement was demonstrated at each station except station I. CONCLUSION: Fourth year medical students without a CCM elective do not possess the basic cognitive and psychomotor skills necessary to initially manage critically ill patients. After an appropriate 1-month CCM elective, students' thinking and application skills required to initially manage critically ill patients improved markedly, as demonstrated by an OSCE using a live simulated "patient" and manikin. PMID- 10708199 TI - Procalcitonin in fever of unknown origin after liver transplantation: a variable to differentiate acute rejection from infection. AB - OBJECTIVE: Does procalcitonin (PCT) differentiate between infection and rejection after liver transplantation in patients with fever of unknown origin? DESIGN: Open prospective trial. SETTING: Transplant intensive care unit at a university hospital. PATIENTS: Forty patients after liver transplantation. INTERVENTIONS: Liver biopsy for the diagnosis of rejection and transcutaneous aspiration cytology for monitoring of lymphocyte activation. MEASUREMENTS: Procalcitonin from EDTA plasma, Acute Physiology and Chronic Health Evaluation II, and sepsis score. RESULTS: Eleven patients experienced an infectious complication resulting in an increase in PCT concentrations (2.2-41.7 ng/mL). Eleven patients had a rejection episode; none of these patients showed a rise in PCT concentrations. The statistical difference between PCT concentrations in rejection and infection was significant (p<.05) on the day of diagnosis. CONCLUSION: PCT allows for differentiation between rejection and infection in patients with fever of unknown origin. Elevation of PCT plasma concentrations develops early postoperatively from operation trauma, and in the case of fever of unknown origin, with no rise in PCT, a rejection may be suspected. PMID- 10708200 TI - Resolution of mucus plugging and atelectasis after intratracheal rhDNase therapy in a mechanically ventilated child with refractory status asthmaticus. AB - OBJECTIVE: To report the dramatic resolution of unilateral mucus plugging and atelectasis in a mechanically ventilated child with refractory status asthmaticus after intratracheal recombinant human DNase (rhDNase) therapy. DESIGN: Case report. SETTING: Critical care unit. PATIENT: A 7-yr-old boy with status asthmaticus, severe respiratory failure and barotrauma unresponsive to conventional therapy. Fiberoptic bronchoscopy confirmed widespread mucus impaction of the subsegmental bronchi of the left lung without response to bronchoscopic lavage. INTERVENTIONS: Two 10-mg doses of intratracheal rhDNase were administered 8 hrs apart. MAIN RESULTS: The left-sided atelectasis resolved 3 hrs after the first dose of rhDNase. Improvements in gas exchange and tidal volumes were sustained and particularly noticeable after the second dose. The patient was successfully extubated 26 hrs after receiving the rhDNase treatment without any adverse effects. CONCLUSIONS: rhDNase should be considered as a potential therapy for refractory mucus plugging and atelectasis in intubated patients with status asthmaticus. PMID- 10708201 TI - Disseminated fatal human cytomegalovirus disease after severe trauma. AB - OBJECTIVE: Disseminated human cytomegalovirus (HCMV) disease is considered to be uncommon in critically ill but otherwise not immunosuppressed patients. We describe the case of a trauma victim who developed fatal HCMV disease that initially presented as pseudomembranous colitis and resulted in sudden cardiac death. DESIGN: Case report of fatal HCMV disease in a previously healthy patient after multiple trauma. SETTING: Surgical intensive care unit (ICU). PATIENT: A 63 yr-old male patient with multiple injuries. INTERVENTIONS AND MEASUREMENTS: Under ICU treatment, symptoms of HCMV reactivation presenting as pseudomembranous colitis appeared 32 days after trauma. Detailed laboratory examinations for HCMV infection were performed, including complement fixation titer, immunoglobulin G and M, polymerase chain reaction, and virus isolation. RESULTS: The intravital detection of HCMV DNA in serum, leukocytes, and a colonic biopsy specimen indicated HCMV reactivation. Postmortem examination findings, including positive viral cultures, showed severe disseminated HCMV disease with involvement of the colon and myocardium. CONCLUSIONS: The lack of specific clinical symptoms of HCMV disease and the delay until viral culture results are available make an exact and timely diagnosis of HCMV disease difficult. Its prevalence in critically ill but otherwise not immunosuppressed patients is currently unknown and possibly underestimated. Because severe illness or trauma can cause immunodysfunction and, thus, may contribute to an increased rate of HCMV disease, detailed studies are warranted to evaluate the real risk in the ICU setting. PMID- 10708202 TI - Acute isoniazid neurotoxicity during preventive therapy. AB - OBJECTIVE: To describe rare side effects of treatment with isoniazid. DESIGN: Descriptive case report. SETTING: Medical intensive care unit in a university medical center. PATIENT: A 14-yr old previously healthy girl receiving preventive isoniazid therapy who suddenly developed generalized tonic-clonic seizures and coma. INTERVENTIONS: Patient was sedated and mechanically ventilated. She also received pyridoxine intravenously. MEASUREMENTS AND MAIN RESULTS: An isoniazid overdose was not confirmed. Computed tomography of the brain and electroencephalogram revealed nothing abnormal. Seizures gradually disappeared within 2 hrs after sedation and treatment with pyridoxine. The patient was discharged on day 14 without consequences and has been well for 10 mos. No seizures reappeared after isoniazid was discontinued. CONCLUSIONS: We caution against possible isoniazid neurotoxicity in healthy individuals using recommended preventive doses. PMID- 10708203 TI - Acute adrenal insufficiency after cardiac surgery. AB - Adrenal insufficiency after cardiac surgery can easily be confused during the course of an immediate unstable postoperative period. If unrecognized, this condition may cause serious morbidity and can be fatal. We report on a 43-yr-old female patient with chronic known adrenal insufficiency, who, despite her adequate preoperative replacement therapy, presented with one episode of acute hypoadrenal crisis after elective open heart surgery, which could serve as a model to illustrate the salient clinical features and possible problems in this setting for diagnosing this problem to patients in whom chronic adrenal insufficiency remains unknown. PMID- 10708204 TI - Turnabout may be more than fair play. PMID- 10708205 TI - Inhaled nitric oxide versus prone positioning in acute respiratory distress syndrome. PMID- 10708206 TI - Prevention of postoperative atrial fibrillation: lukewarm progress. PMID- 10708207 TI - The emperor has no clothes: a misguided case for long-term acute-care facilities? PMID- 10708208 TI - Postinjury oxygen consumption: new method, no new answers. PMID- 10708209 TI - Flow triggering, pressure triggering, and autotriggering during mechanical ventilation. PMID- 10708210 TI - Cost reduction and quality improvement: it takes two to tango. PMID- 10708211 TI - The perils of D-dimer in the medical intensive care unit. PMID- 10708212 TI - Coagulation activation in sepsis. PMID- 10708213 TI - Procalcitonin: new insights on regulation and origin. PMID- 10708214 TI - Of Goldilocks and ventilatory muscle loading. PMID- 10708215 TI - Lidocaine for acute lung injury: questions still to answer. PMID- 10708216 TI - Animal models: the sheep. PMID- 10708217 TI - Should seriously burned children who suffer cardiac arrest be subjected to cardiopulmonary resuscitation? PMID- 10708218 TI - Pediatric cardiac extracorporeal membrane oxygenation: supporting life or prolonging death? PMID- 10708219 TI - To suction or not to suction, above the cuff. PMID- 10708220 TI - Is recombinant tissue plasminogen activator an option in the treatment of meningococcus-induced purpura? PMID- 10708221 TI - Excessive predicted mortality in acute hypoxemic respiratory failure? PMID- 10708222 TI - Drugs and brain death testing. PMID- 10708223 TI - Improvement in pediatric critical care outcomes. PMID- 10708224 TI - Structural-functional relationships of the optic nerve in glaucoma. PMID- 10708225 TI - Digital imaging and microtexture analysis of the nerve fiber layer. AB - PURPOSE: To describe the design of a digital retinal nerve fiber layer (RNFL) imaging techniques and present a new approach to measure the differences in RNFL patterns. METHODS: A digital camera body is connected to a wide-angle camera to obtain images of the RNFL, which are displayed in workstations throughout the clinic. In the on-line archive, images in Joint Photographics Experts Group (JPEG) format (100 KB per frame) are used. The hypothesis that changes in RNFL structure can be seen as changes in the microtexture of digital images was tested using an information theoretical approach (Kullback Information Distance, KID). A large KID value indicates a large difference, and a small KID value indicates a small difference in microtexture between the two regions. The material of this pilot study consists of 9 patients with glaucoma, 8 patients with ocular hypertension, and 7 normal subjects. RESULTS: The median KID value in the glaucoma group was 3.5, compared with the median KID values of 0.6 in the control groups. Although a trend could be seen in the measured values, because of a small sample size, the differences were not statistically significant. Five of 24 (21%) KID values overlapped between the glaucomatous group and the other two groups. CONCLUSION: Although digital imaging produces good quality RNFL images, further research is needed to establish minimum accepted specifications for digital imaging. In this pilot study, only the microtexture of the RNFL was measured in digital images. In the future, the approach can be expanded to include also properties of macrotexture and full color palette. PMID- 10708226 TI - Objective VEP perimetry in glaucoma: asymmetry analysis to identify early deficits. AB - PURPOSE: The multifocal visual evoked potential (VEP) shows markedly symmetrical responses between the two eyes of control subjects. Patients with glaucoma and patients considered at high risk for glaucoma were examined to determine if VEP asymmetry could be identified and used for diagnosis and detection of early damage. METHODS: Multifocal pattern VEP recordings were performed using a single channel bipolar occipital electrode position and the Visual Evoked Response Imaging System (VERIS). There were 125 subjects: 24 control subjects, 70 patients with glaucoma, and 31 patients considered at high risk for glaucoma. A between eye relative asymmetry coefficient (RAC) was determined for each of the 60 test points in the VEP field. The RAC for patients with glaucoma and patients considered at risk for glaucoma were compared with values from control subjects. Correlation between Humphrey thresholds and RAC scores was performed. RESULTS: Patients with glaucoma and patients considered at risk for glaucoma both showed significantly larger mean quadrant RAC values. When point by point analysis was performed, 69 out of 70 scotomas were identified with a cluster of at least 3 points of P < 0.05. For those considered at high risk for glaucoma, 10 out of 31 patients had abnormal areas in the VEP field. There was a strong correlation (r = 0.82) between quadrantic RAC mean values and Humphrey quadrant threshold scores in an asymmetric glaucoma subgroup. Abnormal VEP responses were identified in parts of the visual field that were still normal on perimetry. CONCLUSIONS: Asymmetry analysis correctly identifies patients with glaucomatous field loss and shows abnormalities in many patients considered at high risk for glaucoma who still have normal fields. Asymmetry analysis is able to identify objectively the extent of glaucomatous damage and may be able to detect changes before subjective field loss occurs. PMID- 10708227 TI - Comparison of the Humphrey swedish interactive thresholding algorithm (SITA) and full threshold strategies. AB - PURPOSE: To compare the Swedish interactive thresholding algorithm (SITA) strategy with the full threshold strategy in routine clinical practice. METHODS: Using the Humphrey visual field analyzer model 750 (Allergan Humphrey, San Leandro, CA), 108 subjects were tested with 24-2 SITA (version A9) and 24-2 full threshold strategies. Test results were compared for time taken and reliability and on the basis of seven criteria of abnormality. RESULTS: The SITA required on average 48.8% less time than the full threshold strategy. Patient reliability parameters were somewhat better with SITA. There was a strong correlation between mean deviation and pattern standard deviation. Average threshold sensitivity at each point was increased by 1.31 dB with SITA, but greater differences were seen at points with lower sensitivity. Using the full threshold strategy as our standard for comparison, the sensitivity of SITA varied from 83.0% to 93.2% in detecting the variously defined abnormalities. Fields shown as normal with full threshold strategy corresponded with those found to be normal with SITA in 79.0 to 96.3% cases depending on criteria for abnormality. There were a few cases in which SITA suggested an early abnormality but results of full threshold testing remained normal. On average, the size and depth of scotomas decreased slightly with SITA, but this difference was not statistically significant. Of the 70 patients surveyed about their preference, 65 (92.9%) preferred SITA. CONCLUSION: Full threshold and SITA strategies are comparable in detecting glaucomatous defects. The SITA strategy requires significantly less time to perform and is a satisfactory alternative to full threshold algorithms in clinical practice for diagnosis and management of glaucoma. PMID- 10708228 TI - Concordance between results of optic disc tomography and high-pass resolution perimetry in glaucoma. AB - PURPOSE: To evaluate concordance between results obtained with the Heidelberg Retina Tomograph (HRT) (Heidelberg Engineering GmbH, Heidelberg, Germany) and those obtained with the high-pass resolution perimeter (HRP) in glaucoma diagnosis. METHODS: A total of 217 patients from the glaucoma services at St. Erik Eye Hospital, Stockholm, Sweden (n = 107) and Sahlgrenska University Hospital, Goteborg, Sweden (n = 110) were included in the study. All patients were examined because of known or suspected glaucoma in at least one eye. The conventional indices presented by the instruments were used, i.e., glaucoma index from the HRT (negative values = abnormal, positive = normal) and combined deviation (upper normal limit 2.1 dB) from the HRP. RESULTS: The concordance regarding the classification of normal or glaucomatous by the two instruments was 71% (153/217 eyes). The HRT indicated glaucoma but HRP findings were normal in 47 patients, and the reverse occurred in 17 patients. There was a significant difference in optic disc area between patients with abnormal HRT findings and normal HRP findings and in patients with normal HRT findings and abnormal HRP findings. The concordance could not be improved by adjusting for disc size. The correlation between combined deviation on HRP and the HRT glaucoma index was r = 0.53. CONCLUSION: A significant correlation was found between the combined index given by HRP and the HRT glaucoma index in 217 patients examined for known or suspected glaucoma. Discordant findings were observed in 64 patients; in 22 this discordance was explained by the influence of disc size. No other reasons for differences in examination results could be detected. PMID- 10708229 TI - Study of the contralateral eye in patients with glaucoma and a unilateral perimetric defect. AB - PURPOSE: To study the unaffected fellow eye in patients with glaucoma and unilateral visual field defect, using conventional automated achromatic perimetry, blue-yellow short-wavelength automated perimetry (SWAP), and a nerve fiber layer analyzer (GDx; Laser Diagnostic Technologies, San Diego, CA). METHODS: Eighteen patients in whom a unilateral visual field defect was detected on conventional computerized threshold perimetry were selected. The contralateral eyes of these patients were studied with normal conventional threshold perimetry using blue-yellow perimetry and also were studied with the nerve fiber layer analyzer. Also, 18 eyes of 18 sex- and age- (+/-3 years) matched persons without glaucoma were selected as a control group. RESULTS: Of the 18 contralateral eyes, seven (38.8%) showed a visual field defect on blue-yellow conventional perimetry, and 10 (55.5%) showed a defect of the nerve fiber layer when evaluated with the nerve fiber analyzer. Of the 10 eyes with abnormal visual fields on the nerve fiber analyzer, six (60.0%) also showed a defect on blue-yellow perimetry. In the control group, no eyes showed visual field defect on SWAP, but three eyes (16.6% false positive rate) showed a visual field defect on the nerve fiber layer analyzer. CONCLUSION: These findings suggest that what appeared to be unilateral visual field defects may in fact have been bilateral in at least 33.3% of our patients (n = 6) for whom there was agreement between results of SWAP and the nerve fiber layer analyzer. PMID- 10708230 TI - Which patients are treated for glaucoma? An observational analysis. AB - PURPOSE: More than one million patients in the United States are treated for glaucoma, although little is known about the typical clinical characteristics of this group of patients and the type of therapy they receive. This study was conducted to describe the demographic and diagnostic characteristics of patients beginning long-term drug therapy for glaucoma. METHODS: This cross-sectional study included 544 patients beginning topical glaucoma medication regimens who received care at a group model health-maintenance organization (HMO) located in central Massachusetts. The primary medical records of 544 patients beginning topical glaucoma medication between 1987 and 1990 were reviewed to ascertain the presence of three clinical findings: intraocular pressure (IOP) > or = 22 mmHg; optic disc changes including cup-to-disc ratio > or = 0.8, cup-to-disc asymmetry > or = 0.2, or morphologic disc changes consistent with glaucomatous optic neuropathy; and visual field defect consistent with glaucoma. RESULTS: A majority of the 544 patients (86%) were diagnosed as having primary open-angle glaucoma (POAG) by their physicians. Almost half (44.7%) of these patients had only an elevation in IOP without other clinical findings, and 9% met none of the above criteria for glaucoma according to information in the medical record. CONCLUSION: In this setting, most patients who were prescribed drug therapy for POAG were treated for an elevation in IOP alone in the absence of other ophthalmologic characteristics of glaucoma. PMID- 10708231 TI - Comparison of the effects of 0.5% timolol maleate, 2% carteolol hydrochloride, and 0.3% metipranolol on intraocular pressure and perimetry findings and evaluation of their ocular and systemic effects. AB - PURPOSE: To compare the effects of 0.5% timolol maleate, 2% carteolol, and 0.3% metipranolol on intraocular pressure (IOP) in 45 patients with primary open-angle glaucoma (POAG) and ocular hypertension. A secondary goal of this study was to evaluate the ocular and systemic side effects of these medications. METHODS: Measurements of IOP were taken at baseline (pretreatment) and 2, 6, and 12 hours after instillation on treatment days 15, 30, 60, and 90. Mean sensitivity (MS) and mean defect (MD) values of perimetry before and after treatment and the effects of the three beta blockers on serum lipid profiles were determined. Ocular and systemic side effects were recorded. RESULTS: The most prominent IOP lowering effect was noted with metipranolol at 2 and 6 hours on day 15, and with timolol maleate at 12 hours on day 15 and at all hours of the subsequent days on which measurements were taken. Timolol maleate produced a significant decrease in IOP at 12 hours on day 15 compared with carteolol. There was not a statistically significant difference between the MS and MD values on perimetry before and after treatment for any treatment. There was a statistically significant decrease in levels of total cholesterol and high-density lipoprotein (HDL) cholesterol and a significant increase in triglyceride levels; these changes were observed for all treatments. CONCLUSION: The effects of the three medications were not statistically different from each other in terms of IOP reduction and visual field changes. Careful monitoring of blood lipid levels is necessary with long term treatment with beta blockers, because these agents reduced serum levels of HDL and total cholesterol while increasing triglycerides. Such changes in lipid levels could lead to increased incidence of complications, particularly in patients with atherosclerosis or coronary heart disease. PMID- 10708232 TI - Changes in anterior chamber depth and angle width after filtration surgery: a quantitative study using ultrasound biomicroscopy. AB - PURPOSE: To evaluate the anterior chamber configuration by means of ultrasound biomicroscopy in patients with glaucoma and control subjects, and to determine quantitative changes in this configuration after glaucoma filtration surgery (trabeculectomy) and combined cataract and filtration surgery. METHODS: The study included 33 eyes of 33 patients with glaucoma (diagnosed with primary open-angle or exfoliative glaucoma) in which filtration surgery (n = 17) or combined cataract and filtration surgery (n = 16) was performed, and 25 eyes of 25 age matched control subjects. Ultrasound biomicroscopy was used to measure anterior chamber depth and the angle width at 500 microm from the scleral spur in all eyes. The patients with glaucoma were examined 2 days before surgery and 3 and 6 months after surgery. RESULTS: There were no significant differences in anterior chamber depth and angle width between patients with glaucoma before surgery and control subjects. Postoperative values for anterior chamber depth were significantly greater in patients with glaucoma who underwent combined surgery, but no significant changes were observed in those who underwent filtration surgery alone. In contrast, angle width was significantly greater after surgery both in patients who underwent combined surgery and in those who underwent filtration surgery alone. CONCLUSION: On ultrasound biomicroscopic evaluation, there were no differences in anterior chamber depth and angle width between patients with glaucoma and control subjects. Trabeculectomy alone widens the angle but does not affect the anterior chamber depth; however, combined surgery both deepens the anterior chamber depth and widens the angle. PMID- 10708233 TI - An optical model of the human retinal nerve fiber layer: implications of directional reflectance for variability of clinical measurements. AB - PURPOSE: The reflectance of the retinal nerve fiber layer is highly directional- that is, it depends strongly on the angles of illumination and viewing. This study explored and illustrated the implications of this directional reflectance for nerve fiber layer measurements in the human eye. METHODS: The retina was modeled as a sphere centered on the optic axis of a schematic eye. Nerve fiber ribbons were projected onto the retina and cylindrical light scattering was calculated along each ribbon. The reflectance along the ribbon was then determined for the illuminating and viewing apertures of two hypothetical optical instruments, a fundus camera and a confocal scanning laser ophthalmoscope. Results were displayed as reflectance maps. RESULTS: Uniformly illuminated nerve fiber ribbons exhibited a nonuniform reflectance pattern that was very sensitive to the location in the pupil of the instrument apertures. Ribbon reflectance at the superior and inferior disc margins varied with ribbon orientation, being higher with temporal tilt and lower with nasal tilt. Ribbons nasal to the disk could be quite dim. CONCLUSIONS: In quantitative nerve fiber layer assessment technologies, the observed reflectance depends on the configuration of the illuminating and viewing apertures of the measuring instrument and on the retinal position and orientation of each nerve fiber bundle. In clinical practice, this dependence may cause significant measurement variability that can be reduced by specific measurement maneuvers. PMID- 10708234 TI - Isolation of short-wavelength sensitive mechanisms in normal and glaucomatous visual field regions. AB - PURPOSE: To determine whether "isolation" of short wavelength sensitive mechanisms (i.e., exclusive detection of a threshold stimulus by a short wavelength sensitive mechanism) is maintained in areas of glaucomatous visual field damage as measured with short-wavelength automated perimetry (SWAP). METHODS: Data from conventional automated perimetry and SWAP were analyzed for both eyes of 60 normal control subjects, 38 patients with ocular hypertension, and 22 patients with early to moderate glaucomatous field damage (mean defect better than -12 dB). Comparisons of results of SWAP and conventional perimetry were performed by determining the deviation from the mean normal sensitivity for the two procedures. Locations with sensitivity <3 dB for either procedure were rejected, as 3 dB is near the maximum stimulus luminance and may have introduced a bias by underestimating defects. The interval between deviation from normal (the isolation interval) on conventional perimetry and SWAP was examined to determine the likelihood of short wavelength mechanism isolation loss for different levels of glaucomatous visual field damage. RESULTS: Using normal isolation estimates of 13 dB and 10 dB as bases for determining the likelihood that isolation of short wavelength sensitive mechanisms may have been lost, it was found that this was an infrequent possibility, as low as 0.39 to 1.63% for normal control subjects and 2.53 to 10.44% for patients with glaucoma. CONCLUSION: Analyses indicate that isolation of short wavelength sensitive mechanisms is mostly maintained for SWAP, even in areas of moderate glaucomatous field damage. One limitation of SWAP for evaluating extensive glaucomatous damage is its dynamic range. This could be overcome by using a more intense stimulus light source. PMID- 10708235 TI - Performance of a new, low-volume, high-surface area aqueous shunt in normal rabbit eyes. AB - PURPOSE: This experimental study was conducted to report perfusion characteristics of small diameter, cylindrical aqueous shunts in normal rabbit eyes and to test the hypothesis that decreasing bleb diameter would decrease capsular fibrosis, as evidenced by a thinner capsule forming around the implant. These two properties increase hydraulic conductivity of the fibrous membrane forming around the device, resulting in a more effective filtering shunt. METHODS: Cylindrical latex tubes with the distal portion of the sidewall removed were implanted under the conjunctiva. The proximal, intact end of tubing was inserted into the anterior chamber and ligated to prevent hypotony. The ligature was released after 1 week to inflate the bleb. Animals were again anesthetized at 6 or 12 weeks after ligature release and in vivo perfusion experiments conducted using a miniperfusion system and a water manometer. Perfusion of the implant with latex microspheres was performed before the animals were killed. Capsule diameters were measured in situ using calipers under a dissecting microscope after excision of orbital tissues and before fixation for histologic study. Membrane hydraulic conductivity (outflow per unit membrane area at unitary pressure gradient, microL/min/mm2/mmHg) was calculated using data obtained during perfusion experiments and compared with results of other studies. Capsule thickness was measured histologically. RESULTS: Cylindrical filtration membranes with thin (15-20 microm) capsules formed around latex implants. Bleb diameters consistently measured 1 mm at all points along their length before formaldehyde fixation. Hydraulic conductivity was measured and found to be eight times higher than that reported for capsules around conventional implants. Decreased diameter (16:1 for Baerveldt and 13:1 for Molteno implants) produced a proportional decrease in surface tension on the wall. This resulted in decreased capsule thickness, from 222 microm for Baerveldt implants in rabbit eyes and from 150 microm for Molteno implants in monkey eyes to less than 20 microm for the shunting device studied here. CONCLUSION: As a consequence of Laplace's law, reduction in bleb diameter reduces surface tension on the bleb, reducing capsular fibrosis and consequently capsule thickness, thus increasing hydraulic conductivity. Increased hydraulic conductivity increases the effectiveness of the filtering surface. Cylindrical geometry allows reduced bleb volume yet maintains total surface area that is proportional to the length of the implant, which is unlimited and customizable for each eye by simply cutting the length of implant needed. PMID- 10708236 TI - Initial medical management of open-angle glaucoma. PMID- 10708237 TI - Health-related quality of life in patients with cataract and glaucoma. AB - The patient's perspective of his or her own health status as it relates to functioning and well-being is referred to as health-related quality of life. Various generic and ophthalmology-specific survey instruments have been used to gain an understanding of patient-oriented health status in patients with cataract or with glaucoma. Improvement in vision-targeted quality of life has been shown following cataract surgery; however, an improvement in self-perceived overall health status following cataract surgery has not been established. Increasing severity of glaucoma has been shown to be negatively related to vision-targeted quality of life; the relationship between increasing severity of glaucoma and overall self-perceived health status is inconclusive. Integration of the concepts of health-related quality of life into clinical practice will require the development of better measurement instruments that can demonstrate notable outcome advantages for patients. PMID- 10708238 TI - Latanoprost-induced iris color darkening: a case report with long-term follow-up. AB - PURPOSE: To determine the effect on iris color of discontinuing latanoprost (LP) treatment in a patient with pronounced iris color darkening, and to assess the possible role of sympathetic innervation. METHODS: In a patient demonstrating pronounced iris color darkening in both eyes after treatment with LP for 6 months, magnified iris color photographs were taken at 3- to 6-month intervals for 5 years after discontinuation of LP treatment. Pupillary testing for sympathetic insufficiency was performed with cocaine 10% or hydroxyamphetamine 1%. RESULTS: The iris color did not appreciably change after discontinuing LP. The cocaine-induced increase in pupillary diameter was considerably greater for the control subject than for the patient who demonstrated the LP-induced color change. CONCLUSIONS: Latanoprost-induced iris color darkening does not appreciably change for several years after discontinuing treatment. Some eyes that show LP-induced darkening may have relative ocular sympathetic insufficiency. PMID- 10708239 TI - Regulation of Ped gene expression by TAP protein. AB - The Ped (Preimplantation embryo development) gene regulates fast or slow cleavage of preimplantation mouse embryos and their subsequent survival. The protein product of the Ped gene is the major histocompatibility complex (MHC) class Ib protein Qa-2. MHC class I expression on the cell surface requires the assembly within the endoplasmic reticulum (ER) of an alpha heavy chain, a beta2 microglobulin light chain, and a small peptide. Small peptides are primarily produced in the cytosol by the ubiquitin-proteasome pathway and then are transported into the ER by the transporter associated with antigen processing (TAP) protein. However, some peptides can bind to MHC class I heavy chains in a TAP-independent manner. In this study, we assessed whether TAP protein regulates Qa-2 expression on the cell surface of preimplantation mouse embryos thereby influencing the PED phenotype of the embryos. We chose Tap 1 knockout mice and their control mice (B6.129) as our experimental system. We analyzed Qa-2 mRNA expression by RT-PCR, total Qa-2 protein expression by Western blotting, and cell surface Qa-2 protein expression by Immuno-PCR in preimplantation embryos of both Tap 1 knockout mice and control mice. Then we determined the PED phenotype of both Tap 1 knockout mouse embryos and control mouse embryos. The results showed that Qa-2 expression on the cell surface of preimplantation embryos is dependent on TAP protein, and that Qa-2 expression on the cell surface is required for expression of the fast PED phenotype. PMID- 10708240 TI - The metrial gland is more than a mesometrial lymphoid aggregate of pregnancy. PMID- 10708241 TI - Meconium-stained amniotic fluid exhibits chemotactic activity for polymorphonuclear leukocytes in vitro. AB - We have studied whether meconium-stained, turbid amniotic fluid (turbid AF) obtained during term pregnancy possesses chemotactic activity for polymorphonuclear leukocytes (PMNs) in the absence of clinically apparent infection. Eight samples of turbid AF were obtained from eight women who underwent a cesarean section (four emergency and four elective cesarean sections) in the absence of signs of clinical infection or fetal distress. Samples of clear AF obtained from nine women during an elective cesarean section served as a control. We used also a negative control (medium only) and a positive control containing 10 nM N-formyl-methionyl-leucyl-phenylalanine. The control or turbid AF specimen was placed in the lower compartment of a blind well chamber, and the PMN suspension was placed in the upper compartment. Following incubation, the number of PMNs that had migrated and passed through the filter to the AF was counted. The number of control PMNs that migrated to the turbid AF (200+/-59) was comparable to that of the positive (162+/-24) but significantly exceeded that of the clear AF (17+/-11; P < 0.0001) and of the negative control (25+/-9; P < 0.0001). Checkerboard assay indicated that the turbid AF exhibited a dose dependent chemotactic activity for PMNs. The turbid AF contained higher levels of TNFalpha, IL-1beta and IL-8 than the clear AF. The concentration of IL-8 in the AF was correlated positively with the chemotactic activity of the AF (r = 0.733, P = 0.0005). Anti-human IL-8 antibody added in the turbid AF dose-dependently abolished the chemotactic activity of the turbid AF. It is concluded that meconium-stained AF is a chemoattractant for PMNs and that cytokines such as an IL-8 may be involved in this phenomenon. PMID- 10708242 TI - HLA-DR and HLA-DQ gene typing of infertile women possessing sperm-immobilizing antibody. AB - Thirty-eight infertile women, possessing sperm-immobilizing antibody (SIA), were examined for their HLA-DR and -DQ types using DNA obtained from peripheral blood cells. The typing of HLA-DR and DQ was performed by polymerase chain reaction sequence specific oligonucleotide probes (PCR-SSOP) and PCR-restriction fragment length polymorphism (RFLP), respectively. In comparison to the normal Japanese population, the SIA positive patient population had higher genes frequencies in HLA-DRB1*0901 (26.3 vs. 13.6%, P<0.005), DQB1*0602 (13.2 vs. 6.2%, P<0.05) and DQB1*0303 (26.3 vs. 14.8%, P<0.01), but not in any HLA-DQA1 gene types by chi2 test. After Bonferroni correction, the high frequency of HLA-DRB1*0901 remained significant (P<0.05) and HLA-DQB1*303 was slightly significant (P<0.07) but no other genes had a gene frequency significantly higher than that of the normal Japanese population. HLA-DRB1*0901 and HLA-DQB1*0303 are very rare among Caucasians but characteristically high among Japanese. The high frequency of HLA DRB1*0901 and DQB1*0303 genes in the Japanese population may account for higher frequency of sperm-immobilizing antibody in Japanese compared to other ethnic groups. PMID- 10708243 TI - Thyroid autoimmunity and its association with non-organ-specific antibodies and subclinical alterations of thyroid function in women with a history of pregnancy loss or preeclampsia. AB - Following the observation that non-organ-specific antibodies are related with pregnancy loss and preeclampsia, the role of organ-specific antibodies is currently being extensively investigated. The aim of this study was on the one hand to evaluate the incidence of antithyroid antibodies in a study group of 69 women with a history of early pregnancy loss (subgroup 1), foetal death (subgroup 2) or preeclampsia (subgroup 3) and in a control group, on the other hand to assess the possible association of these autoantibodies with non-organ-specific antibodies and subclinical alterations of thyroid function in the study group. Antithyroid antibodies were present in 26/69 (37.7%) women of the study group (37.9% in subgroup 1; 40.9% in subgroup 2; 33.3% in subgroup 3) and in 10/69 (14.5%) of controls, the difference being statistically significant. A significant difference in the distribution of antibodies to thyroglobulin and thyroid peroxidase was found in subgroup 2. In the study group, the incidence of antiphospholipid antibodies was not significantly different in women positive (26.9%) and negative (34.9%) for antithyroid antibodies. Also, the overall incidence of subclinical alterations of thyroid function in the study group was significantly different in women positive (53.8%) and negative (16.2%) for thyroid autoimmunity (P<0.02). The results of this study seem to confirm the association between thyroid autoimmunity and obstetric complications and suggest the usefulness of undertaking prospective studies in order to evaluate the reproductive outcome of women with a history of recurrent abortion, foetal death or preeclampsia and positivity for antithyroid antibodies. PMID- 10708244 TI - Neonatal thrombocytopenia induced by maternal anti-HLA antibodies: a potential side effect of allogenic leukocyte immunization for unexplained recurrent aborters. AB - Allogenic leukocyte immunization is one of several treatments tried for unexplained recurrent aborters, and is reported to have few maternal and neonatal side effects after the immunotherapy having been reported to date. In the present study, we report a rare case of neonatal thrombocytopenia (41000 cells/microl) observed in a female infant delivered by an unexplained habitual aborter. The mother was immunized with her husband's leukocytes once before pregnancy and twice at the 5th and 6th week of her successful pregnancy. Serological studies using mixed passive hemagglutination assays (MPHA) showed that maternal serum did not contain any antibodies which were reactive to 11 platelet-specific antigens, or to granulocyte antigens extracted from 9 persons. Lymphocyte cytotoxicity tests, however, showed that maternal serum but not infant serum had anti-HLA antibodies against both paternal and infant lymphocytes. Moreover, the maternal serum was found to have anti-HLA IgGs against platelet antigens extracted from the father and the infant. It is highly likely that this case of neonatal thrombocytopenia was caused by transplacental perfusion of maternal anti-HLA antibodies whose production was induced or enhanced by the allogenic leukocytes immunizations. PMID- 10708245 TI - Report from the IFPA meeting in conjunction with the 8th EPG meeting, Schladming, Austria, September 26-29, 1999. AB - Some 350 participants from 35 countries gathered in the small town of Schladming in the Austrian Alps, where the meeting of the International Federation of Placental Associations took place in conjunction with the 8th meeting of the European Placenta Group. The congress was organized by Gernot Desoye and Gottfried Dohr (both from Graz, Austria). The topics covered all aspects of placenta research. One symposium, two workshops and a number of posters were specifically devoted to placental immunology. PMID- 10708246 TI - Effect of the oxidative stress induced by adriamycin on rat hepatocyte bioenergetics during ageing. AB - We have investigated the effect of ageing and of adriamycin treatment on the bioenergetics of isolated rat hepatocytes. Ageing per se, whilst being associated with a striking increase of hydrogen peroxide in the cells, induces only minor changes on mitochondrial functions. The adriamycin treatment induces a decrease of the mitochondrial membrane potential in situ and a consistent increase of the superoxide anion cellular content independently of the donor's age, whilst the hydrogen peroxide is significantly higher in aged than in adult rat hepatocytes. Kinetic studies in isolated mitochondria show that the mitochondrial respiratory chain activity (NADH --> O2) of 50 microM adriamycin-treated hepatocytes is lowered both in adult and aged rats. The same adriamycin concentration induces a slight decrease of the maximal rate of ATP hydrolysis in both young and aged rats, without affecting the Km for the substrate. However, at drug concentrations lower than 50 microM, both ATPase and NADH oxidation activities decrease significantly in aged rats only. The results suggest that free radicals increase during ageing in rat hepatocytes but are unable to induce major modifications of mitochondrial bioenergetics. This contrasts with the damaging effect of adriamycin, suggesting that some effects of the drug may be due to other reasons besides oxidative stress. PMID- 10708247 TI - Effects of long-term, light exercise under restricted feeding on age-related changes in physiological and metabolic variables in male Wistar rats. AB - The effects of long-term, light exercise under restricted feeding on age-related changes in physiological and metabolic functions were examined in male Wistar rats. Adult (100 days old) rats were divided into sedentary (R10S) and exercise (R11E) groups, and given 10 and 11 g/day, respectively, of a 20% casein diet until they reached 900 days of age. Group R11E simultaneously underwent 3000 m/day of running exercise throughout the test period. As compared with the sedentary group, long-term, light exercise significantly increased body nitrogen retention and serum protein levels, decreased body fat and plasma insulin levels, prevented age-related decline in the basal metabolic rate, and reduced age associated histopathological changes in the kidney and liver. Long-term, light exercise further enhanced the benefits of restricted feeding on age-related deterioration in physiological and metabolic variables and improved body composition, but did not prolong survival at 900 days of age. PMID- 10708248 TI - Metabolic mass, metabolic rate, caloric restriction, and aging in male Fischer 344 rats. AB - Previous investigators have found the metabolic rate to be the same in calorically-restricted and ad-libitum fed rodents, and hence concluded that the Rate of Living Theory does not help explain the longer lifespan of the calorically-restricted (CR) animal. However, these previous instigators may not have used reliable estimates of metabolic mass in their calculations of metabolic rate. Hence the present study investigated the reliability of ten different estimates of metabolic mass (MM) in 21-month-old male Fischer 344 rats fed three different diets to yield a wide range of body compositions. Two criteria were used to rank each estimate of metabolic mass: strong correlation with daily caloric intake (DCI); and zero Y-intercept on the regression curve of DCI versus the MM. The combined weight of the heart, liver, kidneys and brain (OW) was found to be the best estimate of MM. Statistical analysis of the differences in metabolic rate in the three groups of rats showed that the significance of these differences depended on the estimate of MM used. OW yielded different results than did fat-free mass (FFM), body weight (BW), BW(0.75), and BW(0.67). Therefore, because previous investigators used FFM, BW, BW(0.75), or BW(0.67), rather than a more reliable estimate such as OW, their finding that metabolic rate was not different in the CR and ad-lib groups, and their conclusion that the Rate of Living Theory does not help explain the longer lifespan of the CR animal, are called into question. PMID- 10708249 TI - Glucose-6-phosphatase and age: biochemical and histochemical studies. AB - Glucose-6-phosphatase catalyzes the final reactions in both gluconeogenesis and glycolysis. It occurs mainly in glycogenic tissues, such as the liver, where it plays an important role in the synthesis of glucose, a carbohydrate essential for tissue functioning. The effect of age on liver glucose-6-phosphatase activity was evaluated in male Wistar rats treated with mixed function oxidase system (MFO) inducers. The rats were divided into the following age groups: 0.5, 1, 2, 4, 8, 12, 20 and 28 months of age. Glucose-6-phosphatase activity was evaluated biochemically and histochemically. Biochemical glucose-6-phosphatase activity increased up to the 20th month of rat life and then decreased rapidly. A similar tendency was observed in inducer-treated groups, though only dexamethasone stimulated this enzyme activity in all age groups studied. Histochemical glucose 6-phosphatase activity was strongest in the periportal zones. Glucose-6 phosphatase activity decreased significantly at month 8 and then it increased significantly until month 20. In the oldest age group, glucose-6-phosphatase activity decreased again. On histochemical analysis, the inducers used variably affected glucose-6-phosphatase activity. PMID- 10708250 TI - Possible involvement of proteasome inhibition in aging: implications for oxidative stress. AB - Oxidative stress may contribute to the cellular alterations, which occur as the result of aging, and the nervous system is particularly vulnerable to aging associated oxidative injury. The multicatalytic proteasome (MCP) is responsible for the majority of protein degradation and is sensitive to oxidative stress. To determine if MCP activity is altered during aging, studies were conducted in multiple tissues from aged Fisher 344 rats. Analysis of heart, lung, kidney, and liver revealed decreased MCP activity in 12, 24, and 28 month old rats, compared with 3 week or 3 month old animals. The spinal cord, hippocampus, and cerebral cortex demonstrated age dependent decreases in MCP activity, but at no timepoint was MCP activity decreased in either the brain stem or cerebellum. Oxidative injury and the lipid oxidation product 4-hydroxynonenal caused decreased MCP activity in neural PC6 cells, while application of MCP inhibitors was sufficient to induce cell death in neural PC6 cells. Together, these data indicate a role for MCP inhibition in cellular dysfunction associated with aging and oxidative injury. PMID- 10708251 TI - Neural network modeling for surgical decisions on traumatic brain injury patients. AB - Computerized medical decision support systems have been a major research topic in recent years. Intelligent computer programs were implemented to aid physicians and other medical professionals in making difficult medical decisions. This report compares three different mathematical models for building a traumatic brain injury (TBI) medical decision support system (MDSS). These models were developed based on a large TBI patient database. This MDSS accepts a set of patient data such as the types of skull fracture, Glasgow Coma Scale (GCS), episode of convulsion and return the chance that a neurosurgeon would recommend an open-skull surgery for this patient. The three mathematical models described in this report including a logistic regression model, a multi-layer perceptron (MLP) neural network and a radial-basis-function (RBF) neural network. From the 12,640 patients selected from the database. A randomly drawn 9480 cases were used as the training group to develop/train our models. The other 3160 cases were in the validation group which we used to evaluate the performance of these models. We used sensitivity, specificity, areas under receiver-operating characteristics (ROC) curve and calibration curves as the indicator of how accurate these models are in predicting a neurosurgeon's decision on open-skull surgery. The results showed that, assuming equal importance of sensitivity and specificity, the logistic regression model had a (sensitivity, specificity) of (73%, 68%), compared to (80%, 80%) from the RBF model and (88%, 80%) from the MLP model. The resultant areas under ROC curve for logistic regression, RBF and MLP neural networks are 0.761, 0.880 and 0.897, respectively (P < 0.05). Among these models, the logistic regression has noticeably poorer calibration. This study demonstrated the feasibility of applying neural networks as the mechanism for TBI decision support systems based on clinical databases. The results also suggest that neural networks may be a better solution for complex, non-linear medical decision support systems than conventional statistical techniques such as logistic regression. PMID- 10708252 TI - Computerized reminders to monitor liver function to improve the use of etretinate. AB - OBJECTIVE: to determine whether computerized reminders during the process of prescribing can improve the use of drugs requiring prior laboratory testing according to the indications listed in the Drug Package Insert. MEASURES: Change in proportion of appropriate prescribing and frequency of severe hepatotoxicity between pre- and post-intervention. METHODS: etretinate, a medication indicated for psoriasis, was selected as a monitored drug because it was the most prescribed of all the identified drugs that require specific prior laboratory tests. Computerized reminders are designed to alert a physician who is about to prescribe etretinate either without the alanine aminotransferase (ALT) test or the aspartate aminotransferase (AST) test within 3 months or despite abnormality in ALT or AST. Data on alerts were gathered by using electronic mail whenever alerts occurred. RESULTS: prescriptions of etretinate with normal ALT or AST results within the previous three months increased suddenly from 25.9% (127/491) in the pre-intervention period to 66.2% (353/533) in the post-intervention period (P < 0.0001). Moreover, three patients who used etretinate had markedly abnormal tests in the pre-intervention period, but none of the patients were classified in this way in the post-intervention period. CONCLUSIONS: the computerized reminders appear to improve physicians' use of a drug requiring specific prior laboratory tests. PMID- 10708253 TI - Mobile information and communication tools in the hospital. AB - Mobile information and communication systems in clinical routine have the potential to greatly improve communication, facilitate information access, eliminate double documentation, and increase quality of patient care in the long run. Projects to date have focused, for the most part, on highly specialized applications of the mobile computer. In our research project, 'Cooperative Problem Solving in Health Care', we have, among other things, designed a multifunctional mobile information and communication assistant. A prototype version of this system was implemented. This article outlines the close-to reality evaluation of our prototype in a 1-week simulation study in a Heidelberg University hospital. We describe methods, aims, design and results of the simulation study, as well as discuss our methodology and the results we have obtained. We argue that the diverse requirements of different professional groups cannot be fulfilled by a single multifunctional device and propose, therefore, a 'multi-device mobile computer architecture'. Finally, we present consequences for the future computing infrastructure. PMID- 10708254 TI - Improving diagnostic accuracy using a hierarchical neural network to model decision subtasks. AB - A number of quantitative models including linear discriminant analysis, logistic regression, k nearest neighbor, kernel density, recursive partitioning, and neural networks are being used in medical diagnostic support systems to assist human decision-makers in disease diagnosis. This research investigates the decision accuracy of neural network models for the differential diagnosis of six erythematous-squamous diseases. Conditions where a hierarchical neural network model can increase diagnostic accuracy by partitioning the decision domain into subtasks that are easier to learn are specifically addressed. Self-organizing maps (SOM) are used to portray the 34 feature variables in a two dimensional plot that maintains topological ordering. The SOM identifies five inconsistent cases that are likely sources of error for the quantitative decision models; the lower bound for the diagnostic decision error based on five errors is 0.0140. The traditional application of the quantitative models cited above results in diagnostic error levels substantially greater than this target level. A two-stage hierarchical neural network is designed by combining a multilayer perceptron first stage and a mixture-of-experts second stage. The second stage mixture-of experts neural network learns a subtask of the diagnostic decision, the discrimination between seborrheic dermatitis and pityriasis rosea. The diagnostic accuracy of the two stage neural network approaches the target performance established from the SOM with an error rate of 0.0159. PMID- 10708255 TI - Framework for a clinical information system. AB - The current status of our work towards the design and implementation of a reference architecture for a Clinical Information System is presented. This architecture has been developed and implemented based on components following a strong underlying conceptual and technological model. Common Object Request Broker and n-tier technology featuring centralised and departmental clinical information systems as the back-end store for all clinical data are used. Servers located in the 'middle' tier apply the clinical (business) model and application rules to communicate with so-called 'thin client' workstations. The main characteristics are the focus on modelling and reuse of both data and business logic as there is a shift away from data and functional modelling towards object modelling. Scalability as well as adaptability to constantly changing requirements via component driven computing are the main reasons for that approach. PMID- 10708256 TI - IGF-1 bioavailability is increased by resistance training in older women with low bone mineral density. AB - We investigated if long-term resistance training would increase insulin-like growth factor-1 (IGF-1) bioavailabilty at rest in older women (68+/-1 years) with low bone mineral density. IGF-1 levels were significantly lower (P<0.05), and insulin-like growth factor binding proteins -1 and -3 (IGFBP-1 and IGFBP-3) significantly higher than an age-matched healthy normal group. Resistance training resulted in significant (P<0.05) increases in repetition maximums across all exercises (range 41-78%). Resting IGF-1 levels were significantly (P<0.05) elevated (70%) by the resistance training whereas no significant changes occurred in IGFBP-1 and IGFBP-3 levels. IGFBP-1/IGF-1 and IGFBP-3/IGF-1 ratios were significantly decreased (approximately - 50%) as a result of resistance training (P<0.05). Thus, IGF-1 bioavailability was increased as a result of resistance training induced increases in IGF-1 levels in older women with low bone mineral density. These alterations in the IGF-1 system may be contributing to the significant strength gain observed with the resistance training in this population. PMID- 10708257 TI - Impaired signal transduction in mitogen activated rat splenic lymphocytes during aging. AB - Mitogen activated protein kinases (MAPK) are activated by a wide variety of signals leading to cell proliferation and differentiation in different cell types. With aging, there is a marked decrease in proliferation of T-lymphocytes in response to a variety of mitogens. Several age-related changes in the activation of MAPK pathways in T-lymphocytes activated via the T-cell receptor (TCR) have been described in different species. This way, some TCR proximal defects in tyrosine kinase activity have been delineated. In this study, we have used rat splenic lymphocytes to measure the effect of aging on the activation of two MAP kinase families: ERK and JNK. In order to bypass the receptor-proximal age-dependent defects previously described, we used phorbol ester (PMA) and Ca2+ ionophore (A23187) as co-mitogens. Our results demonstrate that splenic lymphocytes from old rats have a disturbance in the activation of the ERK and JNK MAPK signal transduction pathways, that are located downstream of the receptor proximal events. At least part of the age-related defect leading to decreased ERK activity appears to be located upstream of ERK itself, since activation of MEK is also impaired. On the other hand, the observed defects in MAPK activation do result in decreased activation of downstream events, such as c-Jun phosphorylation. Thus, we conclude that aging of splenic lymphocytes results in a functional decline in signal transduction, and at least some of these defects are located downstream of the receptor-proximal events previously described by others. The impaired activity of these two MAP kinase pathways is likely to play a role in the diminished lymphoproliferation observed in old individuals. PMID- 10708258 TI - Isolating aging mutants: a novel method yields three strains of the nematode Caenorhabditis elegans with extended life spans. AB - We designed a novel procedure for the isolation of mutant strains with significantly increased life spans in the nematode Caenorhabditis elegans. This procedure involves using heat-shock to screen a large number of animals and isolate a few which are more resistant to heat-shock stress. From the heat-shock resistant animals, three mutant strains, HG25, HG96, and HG246, all exhibiting increased life span, were isolated. One mutant strain (HG246) develops more slowly than the wild-type strain, N2. Two mutant strains, HG96 and HG246, exhibit lower fertility than the wild-type. Each of the three mutant strains has a normal appearance. Their locomotive behavior also appears normal; only HG246 shows slightly slower movement. Their feeding behavior appears normal, and the males of HG25 and HG96 show normal mating behavior. However, the males of HG246, either are defective in their mating ability or their sperm are defective. The results indicate that heat-shock can be used as a means to facilitate the isolation of mutants which have longer life expectancy. PMID- 10708260 TI - Age-related changes of gene expression in mouse kidney: fluorescence differential display--PCR analyses. AB - We used a fluorescence differential display--PCR (FDD-PCR) technique to analyze the genes expressed in mouse kidneys collected at nine different developmental stages ranging from 3 days to 15 months after birth. We found ten genes that were age-dependent and differentially-expressed in the kidneys during our experimental period. We confirmed by comparative RT-PCR that of the ten cDNAs, seven showed reproducible age-dependent expression. Four of the nucleotide sequences of these cDNA clones, had high homology with known genes (fibronectin, soluble guanylyl cyclase alpha-1 subunit, cytosolic aldehyde dehydrogenase and mitochondrial DNA), and three with expressed sequence tags of unknown genes. The FDD-PCR method was very useful for detecting new age-related genes expressed differentially in the mouse kidney. PMID- 10708259 TI - The MHC influences NK and NKT cell functions associated with immune abnormalities and lifespan. AB - The lifespans of H-2 congenic mice differ significantly. The B10.AKM (H-2m) strain has a median survival time (MST) of 15 months, whereas the B10.BR (H-2k) strain has an MST of 24 months. It was previously shown that B10.AKM mice at 13 15 months of age have immunological function comparable to those of B10.BR mice at 22-26 months of age. These functions include: a low proliferative response, reduced levels of intracellular calcium release [Ca2+]i, and an increase in the frequency of memory helper T-cells (CD4+ CD44hiCD45RBlo). In this report similar deficiencies were demonstrated in B10.AKM mice at 2-4 months of age and show that activated spleen NK1.1+CD4+ T (NKT) cells from young B10.AKM mice produce a significantly higher level of IL-4 but a lower level of IFN-gamma as compared to NKT cells from B10.BR mice of the same age. Also, the cytotoxic activity of natural killer (NK) cells from spleens of young (2-4 months) as well as adult (12 16 months) B10.AKM mice is significantly lower (P < 0.01) than that of NK cells from B10.BR mice. These findings suggest that the NKT activity in young B10.AKM mice is a factor for the early onset of immune dysfunction leading to a shorter lifespan. PMID- 10708261 TI - How the United Nations mortality tables hang together. AB - The mortality statistics of 80 member states of the United Nations were used to calculate Gompertz constants. When compared with a model based on the cumulant of failures of 46 human biological functions, they were found to form essentially one set, their slopes spreading out from a constant value like spokes from the axle of a wheel. Data for the longest lived regions agree with a prediction based on the above model, and those for less long lived regions are consistent with such a model having a constant normal cumulant average but increased S.D.s. Several different approaches support the view that the data are based on a probabilistic life-span. PMID- 10708262 TI - Attentional deficits in patients with schizophrenia and in their non-psychotic first-degree relatives. AB - The aim of this study was to investigate whether non-psychotic relatives of schizophrenic probands have deficits in sustained attention as measured by the Continuous Performance Test, Identical Pairs version (CPT-IP) and whether such deficits are associated with negative schizotypal personality disorders. The study subjects were 23 schizophrenic probands, 45 of their first-degree relatives and 36 normal controls. For each subject, attention was assessed during five conditions (2 standard, 2 slow, 1 easy) of visual stimuli (numbers and shapes). Schizotypy status was determined with the physical anhedonia and social anhedonia scales of Chapman et al. (Chapman, L.J., Chapman, J.P., Raulin, M.L., 1976. Scales for physical and social anhedonia. Journal of Abnormal Psychology 42, 374 382). The CPT-IP sensitive index d' in the standard shape condition was significantly lower in schizophrenics and in their relatives than in controls. For all d' values, the percentage of impaired first-degree relatives was at an intermediate level between patients and control individuals. Furthermore, the schizophrenic probands made more random errors in the standard and in the slow number conditions than the other two groups. None of the schizotypy measures correlated with the CPT-IP deficits. These results suggest that spatial sustained attention deficit may be a vulnerability marker for schizophrenia; however, this deficit and the negative dimension of schizotypal personality disorders may be distinct traits. PMID- 10708263 TI - Working memory and Wisconsin Card Sorting Test performance in schizotypic individuals: a replication and extension. AB - The present study examined spatial working memory and Wisconsin Card Sorting Test (WCST) performance in psychosis-prone individuals, either those with extremely high scores on the Social Anhedonia Scale (SocAnh; n = 49) or deviant scores on the Perceptual Aberration-Magical Ideation Scales (Per-Mag; n = 66). Sixty-three individuals with normal scores on the Chapman Psychosis-Proneness Scales served as control subjects. In order to evaluate working memory performance, participants were administered three tasks, namely, sensorimotor, degraded stimulus, and delayed-response tasks. Although the SocAnh and Per-Mag groups displayed poorer performance than control subjects on the working memory task, they did not differ significantly from each other. The SocAnh group exhibited slower reaction times on the working memory task compared to the control group. The groups did not differ in their performance on sensorimotor or degraded stimulus control tasks. Both psychosis-prone groups differed significantly from control subjects in terms of their WCST performance. Working memory performance was inversely associated with the number of perseverative errors (r = -0.17) and the number of trials to complete the first category on the WCST (r= -0.15). These findings extend the literature by indicating that some psychosis-prone individuals with social-interpersonal schizotypal deficits also display subtle spatial working memory impairments. PMID- 10708264 TI - Executive functioning and verbal memory in young patients with unipolar depression and schizophrenia. AB - Although neuropsychological studies have consistently reported executive deficits in schizophrenia, studies of executive functions in depression have produced equivocal results. The aim of this study was to examine the profile and the specificity of the executive impairment and its association with memory performance in young patients with unipolar depression. We compared patients with depression to normal control subjects and schizophrenics. Twenty young inpatients with unipolar depression, 14 schizophrenics and 20 age-, education- and IQ matched control subjects were assessed with a neuropsychological battery including: (1) verbal memory task; (2) frontal tasks (WCST, Cognitive Estimate, Verbal fluency, verbal and visuo-spatial span) and a new complex sorting test (Delis test). Depressed patients and schizophrenics exhibited executive deficits. Unlike schizophrenics, depressed patients did not show memory impairment. Deficits in several 'higher-level' functions combined to produce executive impairments in patients with depression including complex integration for concept formation, spontaneous cognitive flexibility and initiation ability. Impaired functions in schizophrenia and in depressed patients were similar but were differently related to clinical variables. The pattern of memory failure in our schizophrenics is believed to reflect retrieval and encoding deficits. Our findings highlight the heterogeneity of skills grouped under the term 'executive functions' that are vulnerable in depression or schizophrenia. PMID- 10708265 TI - Agonist-stimulated Ca2+ response in neutrophils from patients with primary alcoholism and alcoholized healthy subjects. AB - The sensitivity of the inositol phosphate (IP)/Ca2+-second messenger generating system was assessed in neutrophils from healthy volunteers before and after ingestion of approximately 1%o ethanol for 2 h. In addition, isolated neutrophils from healthy subjects were incubated with ethanol in vitro. Furthermore, the sensitivity of the IP/Ca2+ system was evaluated in neutrophils from alcoholic patients in the state of active drinking, and after 2-3 weeks and 6 months of abstinence. EC50 values of the concentration-response curves obtained by agonist stimulation with formyl-methionyl-leucylphenylalanine (fMLP) of the intracellular Ca2+ accumulation were determined as an indicator of the sensitivity of the system. Ingestion of ethanol by healthy volunteers (both in the ex vivo and in vitro experiments) induced a rightward shift of the concentration-response curve (higher EC50 values) in neutrophils, indicating a reduced sensitivity to agonist stimulation evoked by ethanol. The sensitivity of the Ca2+ response in neutrophils from alcoholic patients decreased intraindividually after a period of 2-3 weeks of abstinence (higher EC50 values) and was at this time also significantly lower compared to a group of matched healthy controls In contrast, the maximal Ca2+ release induced by a saturating concentration of fMLP was increased after 2-3 weeks of abstinence, both intraindividually and in comparison to healthy controls. These alterations of the EC50 values and the maximal Ca2+ response were normalized after 6 months of abstinence. It is concluded that ethanol attenuates the sensitivity of the IP/Ca2+ system in neutrophils in healthy subjects. In neutrophils from alcoholic subjects complex alterations appear to persist up to several weeks, which are only normalized after a prolonged period of abstinence. PMID- 10708266 TI - EEG theta activity and pain insensitivity in self-injurious borderline patients. AB - The principal aim of this study was to investigate possible neurophysiological underpinnings of self-injurious behavior in women with borderline personality disorder (BPD). Pain report and EEG power spectrum density during a laboratory pain procedure, a 4-min 10 degrees C cold pressor test (CPT), were compared among four groups; female inpatients with BPD who do (BPD-P group, n = 22) and do not (BPD-NP group, n = 19) report pain during self-injury, female inpatients with major depression (n = 15), and normal women (n = 20). The BPD-NP group reported less pain intensity during the CPT compared to the other groups. Total absolute theta power was significantly higher in the BPD-NP group compared to the Depressed (P = 0.0074) and Normal (P = 0.0001) groups, with a trend toward being significantly higher compared to the BPD-P group (P = 0.0936). Dissociative Experience Scale scores were significantly higher in the BPD-NP group compared to the Depressed and Normal groups (maximum P = 0.0004), and significantly higher in the BPD-P group compared to the Normal group (P = 0.0016). Beck Depression Inventory and Sheehan Patient Rated Anxiety Scale scores were significantly lower in the Normal group compared to all patient groups. Theta activity was significantly correlated with pain rating (Pearson partial r = -0.43, P = 0.0001) and Dissociative Experiences Scale score (Pearson partial r = 0.32, P = 0.01). PMID- 10708267 TI - Age of onset anticipation in anxiety disorders. AB - Anticipation, an increase in severity or a decrease in the age of onset inherent in the transmission of a disease gene from an affected parent to a child, is being increasingly described in human diseases. In this study we searched for possible anticipation in anxiety disorders. Seventeen unilineal families who had anxiety disorders were compared across two successive generations as to age at the onset of anxiety disorders. Life table analyses revealed a significant decrease in the onset of anxiety disorders from older to younger generations. No evidence of a difference in the type of anxiety disorder was found. Anticipation was thus found in families with anxiety disorders and, if it is confirmed by other studies, trinucleotide repeat sequences may be considered to account for the familial aggregation of anxiety disorders. PMID- 10708268 TI - Platelet [3H]paroxetine binding in patients with OCD-related disorders. AB - The recently introduced notion of clinical conditions being related one to another, the spectrum concept, permits the testing of the involvement of serotonergic systems in a broad range of disorders tentatively linked to obsessive-compulsive disorder (OCD) for which no pathophysiological hypotheses yet exist. We therefore compared the binding of [3H]paroxetine ([3H]Par), a ligand that specifically labels the serotonin (5-HT) transporter, in platelets of drug-free outpatients suffering from various OCD-related disorders with binding in platelets of OCD patients and healthy subjects. Diagnoses were made according to DSM-IV criteria. The most frequent diagnosis was that of body dysmorphic disorder, followed by impulse control disorder, kleptomania, Tourette's syndrome and trichotillomania. Platelet membranes and [3H]Par binding were studied according to standardized protocols. The results, showing a similarly decreased density of [3H]Par binding sites in both patient groups as compared with healthy subjects, suggest the presence of a shared abnormality at the level of the presynaptic 5-HT transporter, probably linked to a common dimension yet to be identified. PMID- 10708269 TI - Predictors of course in obsessive-compulsive disorder. AB - Systematic studies of course of illness in obsessive-compulsive disorder (OCD) using standardized diagnostic criteria are relatively rare. In the present study, 100 patients diagnosed with OCD were prospectively followed for up to 5 years. Other comorbid conditions included anxiety disorders (76%), major depressive disorder (33%), and at least one personality disorder (33%), mainly in the anxious cluster. Approximately 20% of patients had full remission and 50% had partial remission during follow-up. Significant predictors of partial remission included being married and having lower global severity scores at intake; the presence of major depression was marginally predictive of poorer course. Adequate serotonergic medication was associated with worse course, but findings are likely spurious. Only marital status and global severity were retained as predictors in a final regression model. Findings are discussed with regard to sample characteristics and similarity to other reports on predictors of course and of treatment outcome. PMID- 10708270 TI - Treatment of severe mania with intravenous magnesium sulphate as a supplementary therapy. AB - Ten patients with severe, therapy-resistant manic agitation received magnesium sulphate infusions with a continuous magnesium (Mg) flow of approximately 200 mg/h (4353+/-836 mg/day; daily monitored Mg plasma level: 2.44+/-0.34 mmol/l) for periods ranging from 7 to 23 days. Concomitant psychotropic treatment consisted of lithium (n = 10), haloperidol (n = 5) and clonazepam (n = 10). During i.v. Mg treatment the mean values of the maximum dosages of neuroleptics (in chlorpromazine equivalents) and benzodiazepines (in diazepam equivalents) were significantly lower than during the last day of pretreatment (= baseline). Seven patients showed a marked improvement in the Clinical Global Impression scale. In case of bradycardia detected by the ECG monitor (n = 5), Mg flow was reduced and bradycardia disappeared promptly. Mg i.v. may be a useful supplementary therapy for the clinical management of severe manic agitation. This open study needs double-blind confirmation. PMID- 10708271 TI - Non-adherence with long-term prophylaxis: a 6-year naturalistic follow-up study of affectively ill patients. AB - In a retrospective 6-year follow-up, we assessed the reasons for and the frequency and consequences of non-adherence in 76 affectively ill patients receiving lithium prophylaxis in two lithium clinics. Thirty-eight bipolar (50%), 21 unipolar (27.6%) and 17 schizoaffective patients (22.4%) diagnosed according to DSM-III-R, were investigated with a specialized follow-up documentation. Of the patients 53.9% discontinued prophylaxis at some time; 43.2% of the discontinuations occurred during the first 6 months. In contrast to other studies the main reason reported for non-adherence was resistance against long-term treatment. According to the Lithium Attitudes Questionnaire non-adherent patients showed significantly less acceptance of the prophylaxis in general, of the effectiveness of lithium and of the severity of their illness than adherent patients. In a multivariate analysis of various parameters, only the negative attitude to prophylaxis correlated significantly with non-adherence. Significant correlation was found between treatment outcome and duration of initial prophylaxis. During the 6-year follow-up only the adherent patients showed a significant reduction of the number and duration of admissions. Our findings confirmed non-adherence as a major problem in the effectiveness of lithium prophylaxis. The authors recommend prospective investigations of attitudes and the impact of psychoeducation on long-term adherence. PMID- 10708272 TI - Use of BPRS-A percent change scores to identify significant clinical improvement: accuracy of treatment response classification in acute psychiatric inpatients. AB - Use of Brief Psychiatric Rating Scale [Overall J.E., Gorham D.R., 1988. The Brief Psychiatric Rating Scale (BPRS): recent developments in ascertainment and scaling. Psychopharmacology Bulletin 24, 97-99] percent change scores (PCSs) to measure treatment effects may be problematic because two different item-weighting systems (0-6 and 1-7) have been employed to represent the seven rating options and PCSs have demonstrated sensitivity to the item-weighting system used. This study compared the ability of a range of BPRS total scale PCS categories generated by both item-weighting procedures to predict estimates of clinical improvement in a large (N = 1415) heterogeneous acute sample of adult psychiatric inpatients. Results revealed significant differences between the two scaling systems in the proportion of patients classified into categories of PCS symptom improvement. Additional analysis suggested different optimal predictive PCS classifications for each item-weighting system: > 19% for 1-7 and > 39% for 0-6. Guidelines for BPRS publications are presented to facilitate study interpretation and replication. In light of their demonstrated limitations, it is suggested that the use of BPRS PCSs to measure treatment effects be reconsidered. PMID- 10708273 TI - Interactive risk factors for treatment adherence in a chronic psychotic disorders population. AB - This study identified the unique and primary contributions of several concurrent risk factors for poor adherence to treatment recommendations in a clinic population of individuals with chronic psychotic disorders, i.e. 48% had DSM-IV diagnoses of schizoaffective disorder, 38% had schizophrenia, paranoid type, 12% had schizophrenia, undifferentiated type, and 2% had affective disorder with psychotic features. The target cohort consisted of 87 consecutive admissions to a continuing day treatment program. As part of a services-oriented quality assurance program, clinical staff completed rating scales for all patients. These included the BASIS-32 rating scale, which consisted of the following five subscales: psychosis; depression/anxiety; impulsive/addictive behavior; relation to self and others; and daily living and role functioning, and the Working Alliance Inventory-short form (therapist version), which consisted of the following three subscales: goal; task; and bond. These data were used to identify risk factors that weaken a patient's adherence to medication and non-medication treatment during the first 2 weeks of treatment in the clinic. Medication treatment consisted of both typical and atypical neuroleptic medications, with most patients being on multiple medications. Correlational analyses suggested that many of the risk factor variables were significantly associated with poor treatment adherence. Regression analyses suggested that the degree of psychoticism was most strongly associated with poor adherence to medication treatment and that difficulties relating to self and others were the strongest predictor of poor adherence to non-medication treatment. A large-sample services research design such as this can begin to determine patterns of associations between previous identified risk factors and poor treatment adherence in individuals with chronic psychotic disorders. PMID- 10708274 TI - Antipsychotic effects and tolerability of the sigma ligand EMD 57445 (panamesine) and its metabolites in acute schizophrenia: an open clinical trial. AB - Antipsychotic efficacy and side effects of the selective sigma ligand EMD 57445 (panamesine) were investigated in 12 patients (6 males, 6 females) who met DSM III-R criteria for schizophrenia. A 4-week open clinical study revealed only modest effects of EMD 57445 and its metabolites on positive and negative symptoms of schizophrenia. Extrapyramidal and other side effects were moderate, although a significant increase in mild dyskinetic movements was found. Five patients, four of whom were females, completed the trial. Dropouts were mainly due to treatment failure. Antipsychotic effects were significantly greater in female than male patients. PMID- 10708275 TI - Traumatic brain injury in individuals convicted of sexual offenses with and without bipolar disorder. AB - The authors examined the occurrence of traumatic brain injury (TBI) in individuals convicted of sexual offenses with and without bipolar disorder and a comparison group of patients with bipolar disorder without a history of sexual offending behaviors. Individuals convicted of sexual offenses and diagnosed with bipolar disorder had greater rates of brain injury resulting from head trauma than individuals convicted of sexual offenses without bipolar disorder and comparison patients with bipolar disorder. TBI predated the first sexual offense and/or the onset of bipolar disorder in most subjects. PMID- 10708276 TI - Non-nucleoside reverse transcriptase inhibitor resistance among patients failing a nevirapine plus protease inhibitor-containing regimen. AB - OBJECTIVE: To determine the rate of nevirapine resistance in patients failing a nevirapine plus protease inhibitor (PI)-based regimen, and whether these isolates remain susceptible to other non-nucleoside reverse transcriptase inhibitors (NNRTI). DESIGN AND SETTING: A retrospective cohort study in two tertiary university hospitals. PATIENTS: Eighty-eight HIV-infected, NNRTI-naive patients receiving nevirapine plus PI as a rescue regimen after PI treatment failure. MAIN OUTCOME MEASURES: Genotypic and phenotypic resistance data at inclusion (73 and 60 plasma samples, respectively) and after 24 weeks (53 and 42 samples). RESULTS: Baseline phenotypic susceptibility to nevirapine was found in 70% of patients, and similar data were observed for efavirenz (91%) and delavirdine (71%). NNRTI resistance-associated mutations were found in 11 patients (12.5%). At 24 weeks, resistant isolates to nevirapine were found in 92% of patients, and correlated with similar resistance to efavirenz (68%) and delavirdine (73%). In the genotypic analysis, the Y181 C mutation was observed in 76% of mutants, and the most common changes were a combination of mutations at positions Y181C/K103N (23%) and the single mutation Y181C (15%). The development of nevirapine resistance was associated with baseline resistance to PI included in the regimen (P= 0.01). For isolates containing the single amino acid substitution Y181C, 29% remained fully susceptible to efavirenz, whereas 14% showed intermediate resistance to efavirenz and delavirdine. CONCLUSION: The failure of a nevirapine plus PI-containing regimen is associated with nevirapine resistance in most patients, with the most common mutation occurring at amino acid residue 181. Although there is a high degree of cross-resistance among NNRTI, nearly one third of resistant isolates carrying the single Y181C mutation remain susceptible to efavirenz. PMID- 10708277 TI - Phenotypic and genotypic resistance patterns of HIV-1 isolates derived from individuals treated with didanosine and stavudine. AB - OBJECTIVE: To evaluate the phenotypic susceptibilities and genotypic resistance patterns to both didanosine and stavudine of baseline and follow-up HIV-1 isolate pairs, derived from antiretroviral naive subjects treated with this dual nucleoside combination. DESIGN AND METHODS: Phenotypic drug susceptibility testing was performed in peripheral blood mononuclear cells on 34 viral isolate pairs derived from patients participating in the BMS AI-460 trial. Sequencing of the complete reverse transcriptase of 36 study isolate pairs, baseline and follow up, was performed using standard dideoxy techniques. RESULTS: The mean fold change in susceptibilities to didanosine was 1.6 (P= 0.278) and to stavudine 1.9 (P= 0.002, Wilcoxon's signed rank test). Mutations classically associated with zidovudine resistance were observed to emerge in 7 out of 36 isolates, T215Y/F (four), M41L +T215Y/F (two) and D67N (one). Other mutations observed included the A62V, V751, F77L, F116Y, Q151 M multinucleoside resistance complex (one), the Q151M mutation (two) and the rare V75T mutation (two). No mutations classically associated with didanosine exposure and resistance were observed. No relationship was evident between the emergence of zidovudine associated mutations and the level of phenotypic resistance to either stavudine or didanosine or between the emergence of zidovudine associated mutations and changes in plasma HIV RNA levels. CONCLUSION: These comprehensive data demonstrate modest (< twofold) mean reductions in didanosine and stavudine susceptibilities at follow-up. The emergence of zidovudine associated mutations in this retroviral-naive population treated with combination didanosine and stavudine therapy is notable. Furthermore, the emergence of these mutations and of the Q151 M multinucleoside resistance complex raise concerns for potential nucleoside analog cross resistance. The potential mechanisms driving the selection of the zidovudine associated mutations in the setting of didanosine and stavudine therapy and the relevance of these findings to current three and four drug regimens merit further evaluation. PMID- 10708278 TI - Prevalence of HIV-1 resistant to antiretroviral drugs in 81 individuals newly infected by sexual contact or injecting drug use. Investigators of the Quebec Primary Infection Study. AB - OBJECTIVE: Prolonged treatment with antiretroviral drugs results in the selection of HIV-1 variants with mutations conferring resistance to nucleoside and non nucleoside reverse transcriptase inhibitors (NRTI and NNRTI) or to protease inhibitors (PI). There is serious concern about transmission of resistant viruses to newly infected persons. This study monitored the prevalence of resistant viruses in individuals undergoing primary HIV infection. DESIGN: Resistance testing was performed on 81 individuals infected between 1997 and 1999 by injecting drug use (n =21), sexual (n = 56), or unknown (n = 4) transmission. METHODS: Automated sequencing was used to genotype the reverse transcriptase (RT) and protease regions of virus isolated from patients' plasma. The phenotypic susceptibility of stimulated peripheral blood mononuclear cells to antiretroviral drugs was assayed. Line probe assays detected quasispecies variations in wild type and mutated RT codons. RESULTS: A high prevalence of PI and RT genotypic variants, associated with high-level resistance to antiretroviral drugs, was observed in individuals newly infected by injecting drug use (PI = 24%, RT = 24%) or sexual transmission (PI = 12%, RT = 22%). The PI mutations, L101, V82A, and L90M, were found in 10.5, 3 and 4% of cases, respectively; whereas for RT, primary mutations at positions T215Y (zidovudine), M184V (lamivudine), T69D/A (zalcitabine), and K103N (multi-NNRTI) were present in 8, 5, 4, and 4% of subjects, respectively. Resistance to NRTI was demonstrated by phenotypic, genotypic, and line probe analyses. Transmission of multidrug (NRTI/NNRTI/PI) resistance in eight subjects (9.9%) was confirmed by showing that source partners possessed viruses of similar genotype. CONCLUSIONS: The transmission of drug resistant HIV is a serious problem that merits further attention by public health officials as well as virologists and clinicians. PMID- 10708279 TI - CCR5 promoter polymorphisms, CCR5 59029A and CCR5 59353C, are under represented in HIV-1-infected long-term non-progressors. The Australian Long-Term Non Progressor Study Group. AB - OBJECTIVE: To determine the influence of CCR5 promoter polymorphisms on HIV-1 progression to AIDS and to evaluate the interaction between CCR5 structural polymorphisms and those occurring in the regulatory region of the same gene. PARTICIPANTS: Seventy-one HIV-1-infected long-term non-progressors with a CD4+ T cell count of > 500 x 10(6)/I more than 8 years after infection were compared with 75 HIV-1-infected individuals who had progressed to AIDS and/or death within 8 years and with a further 119 HIV-1-positive patients who had CD4+ T cell counts of 200-500 x 10(6)/l. An additional 92 HIV-negative individuals were also studied. METHODS: CCR5 delta32 genotype was determined by PCR with primers spanning the 32 base pair deletion. CCR2-64I, CCR5 59029A/G and CCR5 59353C/T genotypes were determined by PCR followed by restriction fragment length polymorphism analysis. RESULTS: Strong linkage disequilibrium between the CCR5 59029A and CCR5 59353C polymorphic variants was identified. CCR5 59029A and CCR5 59353C homozygotes were found to be significantly under-represented in the long term non-progressor group as compared with the other HIV-1-positive groups, with the effect being more marked in the absence of the CCR5 delta32 and CCR2 64I mutations. CONCLUSIONS: This study provides the first evidence for an association between CCR5 promoter polymorphisms and long-term asymptomatic HIV-1 infection, with individuals lacking the CCR5 59029A/CCR5 59353C homozygous genotype likely to progress more slowly towards AIDS and/or death. PMID- 10708280 TI - Different immunologic profiles characterize HIV infection in highly active antiretroviral therapy-treated and antiretroviral-naive patients with undetectable viraemia. The Master Group. AB - BACKGROUND: Suppression of human immunodeficiency virus (HIV) replication can be obtained in chronically infected individuals by highly active antiretroviral therapy (HAART) and can also be observed in antiretroviral-naive patients. The immunological correlates of these two situations were examined. DESIGN AND METHODS: Cross-sectional study involving 32 HIV-infected patients with undetectable HIV plasma viraemia (< 500 copies/ml) and either antiretroviral naive (n = 14) or undergoing HAART therapy with two nucleoside reverse transcriptase inhibitors (NRTI) plus one (n = 13) or two (n = 5) protease inhibitors (PI). CD4 counts, disease duration, and CDC clinical stage were comparable between the two groups of individuals. Immune parameters (antigen- and mitogen-stimulated proliferation and cytokine production; cytokine mRNA; beta chemokine production; HIV coreceptors mRNA) were analysed in all patients. RESULTS: Results showed immune profiles to be profoundly different in antiretroviral-naive in comparison with HAART-treated patients. Thus: (1) T-cell proliferation to HIV-specific and HIV-unrelated antigens is potent in antiretroviral-naive but suppressed in HAART-treated individuals; (2) interleukin (IL)2, IL-12 and interferon gamma (IFNgamma) production is robust in naive patients; and (3) a high CCR5/low CXCR4 pattern of HIV coreceptors-specific mRNA is observed in naive but not in HAART-treated patients. In contrast with these observations, no clear differences were detected when beta chemokine production by either peripheral blood mononuclear cells or purified CD8+ T-cells was analysed. Results from HAART-treated patients undergoing therapy with one PI and two NRTI or two PI and two NRTI were in very close agreement. CONCLUSIONS: These data suggest that control over HIV replication can be independently achieved by pharmacological or immunologic means. HAART is associated with weaker HIV specific and -non-specific immune responses. PMID- 10708281 TI - Potent antiretroviral treatment of HIV-infection results in suppression of the seminal shedding of HIV. The Swiss HIV Cohort Study. AB - OBJECTIVE: The amount of HIV in semen likely influences infectiousness. Antiretroviral therapy decreases HIV-RNA in semen, but data on HIV concentrations in semen in a large cohort of men with suppressed HIV-RNA in blood is unavailable. METHODS: Male patients with a treatment-induced reduction of HIV-RNA load in plasma below 400 copies/ml were asked to donate a semen and blood sample. Blood and seminal plasma were tested for the presence of HIV-RNA by the NucliSens method (detection limit 400 copies/ml). Seminal cell samples from 67 patients were further analysed for the presence of HIV-DNA using a nested DNA-polymerase chain reaction. Results of RNA and DNA testing in semen were compared with 55 HIV positive antiretroviral therapy-naive men. RESULTS: A total of 114 patients participated in the study. Seminal plasma HIV-RNA was detectable in only two patients [1.8%, 95% confidence ratio (CI), 0-4.2%] compared with a detection frequency of 67% in untreated controls [Odds ratio (OR), 0.01; 95% CI, 0-0.03]. Detection of cell-associated HIV-DNA in semen was significantly less frequent (16 versus 38%) in patients receiving suppressive therapy compared with untreated controls (OR, 0.32; 95% CI, 0.12-0.80). CONCLUSION: In patients with treatment induced suppression of blood viral load the likelihood of having detectable HIV in semen is very low (< 4%). In addition, seminal shedding of cell-free and cell associated HIV is significantly lower than in an untreated population of HIV infected asymptomatic men. On a population basis, this effect of therapy may help to reduce sexual transmission of HIV. However, individual patients may still be infected as evidenced by continued shedding of cells harbouring the HIV provirus. PMID- 10708282 TI - Natural history of serum HIV-1 RNA levels in 330 patients with a known date of infection. The SEROCO Study Group. AB - OBJECTIVE: To describe the spontaneous course, before the introduction of highly active antiretroviral therapy (HAART), of HIV-1 RNA during the AIDS-free period of the disease. To assess the predictive value of changes in HIV-1 RNA levels. DESIGN: A total of 330 patients with a known date of infection followed in the SEROCO cohort. METHODS: HIV-1 RNA levels (threshold, 200 copies/ml) were evaluated from 2243 frozen sera obtained from enrolment until the onset of AIDS or until February 1996. Lowess curves were used to describe the variations of viraemia during follow-up. A Cox regression model was used to assess the predictive value of early and updated CD4 cell count and viral load. RESULTS: In addition to a lower early viral load, patients who remained AIDS-free had, on average, a longer period of viral load decrease after infection (36 versus 18 months), followed by a slower viral load increase compared with those who progressed to AIDS. A true plateau-phase after the seroconversion period, lasting approximately 4 years, was identified only in patients who remained AIDS-free for at least 90 months. In multivariate analysis, both early viral load and later changes were significant predictors of progression to AIDS. A decrease in the CD4 cell count to less than 200 cells/microl and the onset of a group B condition remained significant predictors of progression. CONCLUSION: Our study extends to the early post-seroconversion phase the prognostic value of extracellular HIV-1 RNA levels. Moreover, our data suggest that, in most HIV-infected individuals, a progressive loss of control of viral replication arises during the early years of HIV-1 infection. PMID- 10708283 TI - B-cell stimulation and prolonged immune deficiency are risk factors for non Hodgkin's lymphoma in people with AIDS. AB - OBJECTIVES: To identify risk factors for non-Hodgkin's lymphoma (NHL) in people with HIV infection. DESIGN AND SETTING: Case-control study in Sydney, Australia. PARTICIPANTS AND METHODS: Two hundred and nineteen patients with AIDS-related NHL were compared with 219 HIV-infected controls without NHL, matched for CD4 positive cell count and date of specimen collection. Data on demographic, infectious, treatment-related and immunological factors were abstracted by medical record review. The association between demographic factors, sexually transmissible diseases, HIV-related opportunistic infections, anti-viral therapy, duration of immune deficiency and indices of immune stimulation and risk of NHL were derived for these groups. RESULTS: In a multivariate model, there were two independent groups of predictors of NHL risk. The first was duration of immunodeficiency, as measured by longer time since seroconversion (P for trend 0.008), and lower CD4 positive cell count 1 year prior to the time of NHL diagnosis (P for trend 0.009). The second predictor was B-cell stimulation, as indicated by higher serum globulin (a surrogate marker for serum immunoglobulin, P for trend 0.044) and HIV p24 antigenaemia [odds ratio (OR) for p24 positivity, 1.82; 95% confidence interval (CI), 1.15-2.88]. Indices of B-cell stimulation preceded the diagnosis of NHL by several years. Factors not related to NHL risk included clinical indices of Epstein-Barr virus infection and receipt of individual nucleoside analogue antiretroviral agents. Combination therapy with these agents was associated with a non-significant reduction in NHL risk (OR, 0.68; 95% CI, 0.39-1.18). CONCLUSIONS: Markers of long-standing immune deficiency and B-cell stimulation were associated with an increased risk of developing NHL. Unless the strongest risk factor for NHL, immune deficiency, can be reversed, NHL is likely to become proportionately more important as a cause of morbidity and mortality in people with HIV infection. PMID- 10708285 TI - Adherence to HAART in French HIV-infected injecting drug users: the contribution of buprenorphine drug maintenance treatment. The Manif 2000 study group. AB - OBJECTIVES: To assess adherence to highly active antiretroviral therapies (HAART) in a cohort of French patients infected by HIV through injection drug use (IDU), and the impact on adherence of buprenorphine ambulatory drug maintenance treatment (DMT) which has been widely introduced since 1996. DESIGN: Adherence assessment at first visit after initiation of HAART in the MANIF2000 cohort study. METHODS: Patient's face-to-face and self-administered questionnaires. Univariate and logistic regression adjusted odds ratios (OR) to compare characteristics of non-adherent versus adherent patients. RESULTS: Of the 164 patients, 34.8% took less than 80% of the prescribed HAART doses during the previous week. Decrease in viral load titres after initiation of HAART was significantly lower among non-adherent patients. After adjustment by logistic regression, non-adherence was associated with younger age, alcohol consumption, frequency of negative life-events during the prior 6 months and active drug use. However, IDU in buprenorphine DMT reached higher levels of adherence (78.1%) than ex-IDU (65.5%), although this difference did not reach statistical significance. CONCLUSION: Prescription of buprenorphine DMT may increase adherence to HAART among HIV-infected opiate-dependent patients. Reducing the negative impact of stressful life-events through psychosocial interventions should be considered, even for those who have stopped using drugs. PMID- 10708284 TI - Factors associated with clinical and virological failure in patients receiving a triple therapy including a protease inhibitor. AB - OBJECTIVE: To determine the predictors of virological and clinical failure in patients receiving a protease inhibitor as part of triple therapy. METHODS: From the French Hospital Database on HIV, 1402 protease inhibitor-naive patients starting a highly active antiretroviral therapy regimen with ritonavir, saquinavir-hard gel capsule (hgc) or indinavir between July 1996 and March 1997, and with measured HIV RNA at baseline and at 12 months, were studied for progression to a new AIDS-defining event (ADE) or death. Virological failure was defined as plasma HIV RNA > 1000 copies/ml at 12 months. Multivariate analyses were performed using Cox models for clinical outcomes and logistic regression for virological outcomes. RESULTS: During a median follow-up of 14.1 months, 94 (6.7%) patients experienced an ADE or died. At 12 months, 700 patients (49.9%) had virological failure. In the multivariate analysis, baseline CD4 cell count and viral load were found to be independent predictors of both virological and clinical failure. Neither the type of the first protease inhibitor taken nor previous antiretroviral therapy experience was associated with risk of clinical progression. For virological failure, the use of saquinavir-hgc was associated with a significant 1.96-fold increase in risk compared with indinavir; pre treated patients were at higher risk than antiretroviral therapy-naive patients. CONCLUSION: In this study with large samples of patients, the use of saquinavir hgc was associated with higher risk of virological failure at 12 months than were ritonavir and indinavir; no differences between protease inhibitors were found for clinical progression. As biases cannot be excluded, a longer duration of follow-up will be necessary to answer the question of whether the results are really discrepant or simply reflect the delay between virological failure and clinical manifestations. PMID- 10708286 TI - A randomized, comparative study of lamivudine plus stavudine, with indinavir or nelfinavir, in treatment-experienced HIV-infected patients. AB - OBJECTIVE: To compare adherence and clinical outcome with two modalities of highly active antiretroviral therapy (HAART), in HIV-infected patients. DESIGN: Randomized, open-label, prospective study. SETTING: Tertiary care centre in Spain. PATIENTS: A total of 112 non-naive HIV-infected patients, recruited from March 1998 through August 1998, were studied. INTERVENTIONS: Triple drug therapy with stavudine and lamivudine, plus indinavir or nelfinavir. MAIN OUTCOME MEASURES: Adherence, side-effects, and immunological, virological, and clinical efficacy of treatment were assessed at 3-month intervals. RESULTS: After a median follow-up of 9 months, 32% of patients in the indinavir group versus 50% of those in the nelfinavir group showed adequate adherence in all clinical appointments (P= 0.0559). Adherence was superior in the nelfinavir group in every visit. After 6 months of treatment 48% of subjects in the indinavir group and 70% of those in the nelfinavir group exhibited adequate adherence (P= 0.0311). After 9 months 35% of patients in the indinavir group and 59% of those in the nelfinavir group showed adequate adherence (P= 0.0291). Side-effects provoked discontinuation of treatment in 34% of patients in the indinavir group and 12% of patients in the nelfinavir group (P= 0.0073). Immunological and virological efficacy were similar in both groups. CONCLUSIONS: Adherence to a HAART regimen with stavudine plus lamivudine plus nelfinavir was superior to a regimen with stavudine plus lamivudine plus indinavir. Side-effects provoked more discontinuation of treatment in the indinavir group than in the nelfinavir group. PMID- 10708287 TI - HIV-1 reverse transcriptase (RT) genotype and susceptibility to RT inhibitors during abacavir monotherapy and combination therapy. AB - OBJECTIVE: To examine changes in HIV-1 susceptibility (genotype and phenotype) during an initial abacavir monotherapy phase followed by the addition of zidovudine and lamivudine. DESIGN: Sixty HIV-1 infected, antiretroviral therapy naive subjects were randomized to receive 100, 300 or 600 mg abacavir twice daily. Subjects completing 24 weeks of randomized therapy or meeting a protocol defined switch criterion could switch to open label abacavir/zidovudine/lamivudine. METHODS: Plasma HIV-1 reverse transcriptase was genotyped at baseline, week 12, and at the last time point on ABC monotherapy. Drug susceptibility was analysed at baseline and on subsequent samples with sufficient HIV-1 RNA levels using the recombinant virus assay. Virological responses (week 24) were correlated to week 24 genotypes. RESULTS: Mutant viruses were not detected before week 12 with the exception of one subject. At the latest time point on abacavir monotherapy (range, weeks 6-48), 21 out of 43 subjects harboured virus with resistance conferring mutations including single, double and triple combinations of K65R, L74V, Y115F and M184V. The most common mutational pattern was L74V + M184V (11/21 cases). Twenty of the 21 subjects with isolates containing abacavir-associated mutations reached week 48, and upon addition of lamivudine/zidovudiine, 16 out of 20 (80%) had week 48 plasma HIV-1 -RNA below 400 copies/ml. At week 48, 16 out of 46 genotypes were obtained; one of these was wild-type; 15 contained M184V either alone, in combination with K65R and/or L74V and/or Y115F or with thymidine analogue-associated mutations. Week 48 viral load levels for these 15 subjects was low (median 3.43 log10 copies/ml or -1.99 log10 copies reduction from baseline). Genotype correlated well with phenotypic resistance to ABC; four samples with three abacavir-associated mutations had high level abacavir resistance (> 8-fold) and six samples with two or three mutations showed intermediate (4-8-fold) resistance. All samples with single mutations retained full ABC susceptibility. CONCLUSIONS: Resistance conferring mutations to abacavir were relatively slow to develop during the monotherapy phase, and did not preclude durable efficacy of abacavir/lamivudine/zidovudine up to 48 weeks. PMID- 10708288 TI - CMV retinitis recurs after stopping treatment in virological and immunological failures of potent antiretroviral therapy. AB - OBJECTIVES: To determine predictors of clinical relapse of cytomegalovirus (CMV) end-organ disease in a cohort of 17 HIV-infected patients with healed and treated CMV retinitis (CMVR) who responded to HAART with an increase in CD4 cell counts to above 70 cells/mm3 and discontinued CMV maintenance therapy (MT). DESIGN: Seventeen patients were monitored for reactivation of retinitis. The CD4 cell counts, HIV RNA and peripheral blood mononuclear cell (PBMC) lymphoproliferative assays to CMV at 3 month intervals were compared between patients with and without reactivation of CMVR. Positive lymphoproliferative responses were defined as a stimulation index of 3 or greater. RESULTS: Five out of 17 (29%) patients experienced a recurrence of CMVR a mean of 14.5 months after stopping CMV MT and between 8 days and 10 months after CD4 cell counts fell below 50 cells/mm3. Median CD4 cell counts and plasma HIV RNA at reactivation were 37 cells/mm3 and 5.3 log10 copies/ml. Three patients recurred at a previously active site of the retina, one had contralateral CMVR, and one a recurrence of retinitis and pancreatitis simultaneously. Mean lymphoproliferative responses to CMV were 2.4 in patients with reactivation versus 21.0 stimulation index (SI) in patients without reactivation (P= 0.01). A model incorporating four variables (CD4 cell counts and HIV RNA at maintenance discontinuation, highest CD4 cell count, nadir HIV RNA and median lymphoproliferative responses) identified correctly 88% of patients with and without reactivation. CONCLUSION: CMV disease recurs after virological and immunological failure of HAART if CD4 cell counts drop below 50. In this situation, anti-CMV agents should be resumed before clinical reactivation ensues, because of the risk of contralateral retinal involvement and systemic disease. PMID- 10708289 TI - Quality of life in asymptomatic- and symptomatic HIV infected patients in a trial of ritonavir/saquinavir therapy. The Prometheus Study Group. AB - OBJECTIVE: To compare the impact on quality of life (QoL) of treatment with ritonavir (RTV)/saquinavir (SQV) versus RTV/SQV/stavudine (d4T) in asymptomatic [Centers for Disease Control (CDC) class A] and symptomatic HIV-infected patients (CDC B and C) who did or did not receive antiretroviral therapy (ARVT) before entry into the study. DESIGN: A multicenter randomized clinical trial. PATIENTS: Protease inhibitor- and d4T-naive patients were allocated to RTV/SQV (n = 84) versus RTV/SQV/d4T (n = 83). MAIN OUTCOME MEASURE: Changes from baseline in QoL assessed by the Medical Outcomes Study Health Survey for HIV (MOS-HIV) and a symptom checklist administered at baseline and after 12, 24, 36 and 48 weeks. RESULTS: Changes in QoL were comparable in both treatments, although more neuropathy was reported in the RTV/SQV/d4T group. QoL improved significantly in both groups regarding health distress, energy/fatigue, mental health, health perceptions, physical function and overall QoL, despite an increase in reported symptoms. More favourable changes in cognitive and social function were observed in symptomatic compared with asymptomatic patients, with symptomatic patients showing an improvement and asymptomatic patients showing a decline in function after baseline. ARVT-naive patients showed more favourable changes in mental health, health distress and social function compared with patients with previous ARVT. CONCLUSION: RTV/SQV and RTV/SQV/d4T were equally effective in improving the QoL of patients over 48 weeks, despite an increase in reported symptoms. Symptomatic patients reported more QoL benefit than asymptomatic patients, and ARVT-naive patients benefitted more than those with previous ARVT. The impact on patients' QoL should be considered in the search for the optimal management of HIV infection. PMID- 10708290 TI - High HIV seroprevalence associated with gonorrhea: New York City Department of Health, sexually transmitted disease clinics, 1990-1997. AB - OBJECTIVE: To measure trends in HIV seroprevalence associated with gonorrhea in patients presenting to New York City Department of Health sexually transmitted disease (STD) clinics, 1990-1997 (n = 94 577). METHOD: Unlinked HIV-1 serosurvey using remnant serum originally drawn for routine serologic tests for syphilis (STS). Demographic, risk factor, clinical and laboratory data were abstracted from clinic charts. No other data sources were used. Patients were not interviewed. RESULTS: During 1990-1997 HIV seroprevalence declined from 9 to 6% (P for trend < 0.01) in the STD clinic sample. Gonorrhea incidence city-wide declined from 481 per 100 000 to 194 per 100 000. HIV seroprevalence in patients with a diagnosis of gonorrhea (n = 11 914) remained stable at 10-11% during the period, whereas HIV seroprevalence associated with all other STDs combined declined from 8 to 5% (P for trend < 0.01). Seroprevalence in women with gonorrhea (n = 2243) declined from 8 to 4% (P for trend < 0.001), whereas seroprevalence in men with gonorrhea was stable at 11-12%. Seroprevalence in men aged less than 25 years and diagnosed with gonorrhea declined from 5 to 3% (P for trend = 0.02). In contrast, in men aged 25 years and older and diagnosed with gonorrhea, seroprevalence remained at 14-16% throughout the period 1990-1997. Among men with gonorrhea, seroprevalence was associated with same gender or bisexual contact [odds ratio (OR), 9.2; 95% confidence interval (CI), 8.1-10.4], age > 25 years (OR, 5.1; 95% CI, 4.6-5.7), and white race/ethnicity (OR, 1.3; 95% CI, 1.2-1.4). CONCLUSIONS: In this 9-year serosurvey the association between HIV and gonorrhea remained strong during a period when HIV seroprevalence and gonorrhea incidence declined. The data suggest that a gonorrhea diagnosis is an important risk marker in this era of 'safe sex' and that behavior patterns of patients with gonorrhea warrant further study. PMID- 10708291 TI - Estimating the population impact in Australia of improved antiretroviral treatment for HIV infection. AB - OBJECTIVE: To estimate the reduction in AIDS incidence, if any, which has occurred in Australia following the availability of new combination antiretroviral treatments from 1995. DESIGN: Analyses were based on national surveillance data. METHODS: Back-projection analyses based on quarterly AIDS counts to the end of 1994 were used to estimate the numbers of AIDS diagnoses which would have occurred if new treatments had not reduced the rate of progression to AIDS. Estimates of the reduction in AIDS diagnoses between 1995 and 1998 were made by subtracting the observed delay-adjusted AIDS counts from the predicted AIDS incidence. RESULTS: AIDS incidence between 1995 and 1998 was estimated to have been reduced by 1093 cases (33%) following the availability of new antiretroviral treatments (95% confidence interval 831 (25%) to 1425 (43%) cases). The majority of this reduction in AIDS incidence was estimated to have occurred during 1997 (434 cases) and 1998 (427 cases). CONCLUSIONS: AIDS incidence in Australia has declined since 1995 coincidental with introduction of new antiretroviral treatments. In particular, the more rapid decline in AIDS incidence since mid-1996 coincided with the availability and widespread uptake of combinations including protease inhibitors. PMID- 10708292 TI - Long-term effects of therapeutic vaccination with rgp 120. PMID- 10708293 TI - Fas and Fas ligand are not involved in the suppression of HIV replication by CD8 cells. PMID- 10708294 TI - HIV-1 infection transmitted by serum droplets into the eye: a case report. PMID- 10708295 TI - Hydroxyurea plus didanosine as maintenance therapy after 1 year on highly active antiretroviral therapy. PMID- 10708296 TI - HIV drug-resistance testing on archived samples to help current clinical decisions. PMID- 10708297 TI - Prevalence of HIV and syphilis among high-risk groups in Bangladesh. PMID- 10708298 TI - Hepatitis C viraemia in HIV-HCV co-infected patients having immune restoration with highly active antiretroviral therapy. PMID- 10708299 TI - Synthesis and autoradiographic evaluation of a novel high-affinity Tc-99m ligand for the 5-HT2A receptor. AB - The synthesis and in vitro autoradiography of a novel Tc-99m ligand with subnanomolar affinity to the 5-HT2A receptor is reported. The complex combines the 4-(4-fluoro)-benzoyl piperidine portion derived from the 5-HT2A receptor antagonist ketanserin with a neutral oxotechnetium(V) chelate in form of a mixed ligand "3 + 1" unit containing the SNS/S donor set. The analogous rhenium compound has been synthesized as a surrogate for the Tc-99m complex for use in receptor binding assays and for complete structural characterization. In competition experiments the Tc-99 complex as well as its Re analogue display subnanomolar affinity toward the 5-HT2A receptor (Ki 0.44 nM for Tc, 0.25 nM for Re). The subnanomolar 5-HT2A receptor binding of the Re complex was confirmed by functional in vitro antagonism of contractile effects evoked by 5-HT in rat arterial tissue. Re 1 inhibited 5-HT-induced, 5-HT2A receptor-mediated contractions of isolated rat tail artery in a competitive fashion and possessed nanomolar affinity (pA2 = 9.08, pA2 representing the negative decadic logarithm of the Re 1/5-HT2A-receptor dissociation constant [mol/L]). Like ketanserin, Re 1 displayed moderate affinity for adrenergic alpha1D (pA2 = 8.23) and histamine H1 receptors (pA2 = 8.00), and was >600-fold up to 10,700-fold less active at several neurotransmitter receptor subtypes. In vitro autoradiographic studies clearly indicate the accumulation of the Tc-99m compound in 5-HT2A-receptor-rich areas of the brain. This enrichment can be blocked by 5-HT2A receptor antagonists such as mianserin and ketanserin and is therefore specific. PMID- 10708300 TI - Somatostatin receptor scintigraphy during treatment with lanreotide in patients with neuroendocrine tumors. AB - To investigate possible changes in somatostatin receptor expression during treatment with high dose lanreotide, eight patients with neuroendocrine tumors were investigated by [(111)In-DTPA-D-Phe1]-octreotide scintigraphy before and during treatment. The spleen-to-background ratio decreased in all patients, whereas tumor-to-background ratio revealed a heterogeneous pattern with an average increase of 50% (-79% to +1,087%). This finding indicates that lanreotide treatment may influence the binding of radioactively labeled somatostatin to the spleen, while changes in the binding to functioning somatostatin receptors in tumor cells are more complex and not clearly related to treatment. PMID- 10708301 TI - The integrity of the disulfide bond in a cyclic somatostatin analog during 99mTc complexation reactions. AB - Recent development of a variety of thiol-free chelating agents has facilitated the design of 99mTc-labeled somatostatin analogs suitable for receptor imaging of somatostatin-positive tumors. However, it remains ambiguous whether the disulfide bonds in cyclic peptides are stable during 99mTc complexation reactions, and contradictory results have been reported regarding the integrity of disulfide bonds in cyclic somatostatin analogs. To estimate the stability of the disulfide bond in a synthetic somatostatin analog at low peptide concentrations, [125I]I-RC 160, in which radioiodine was incorporated into the 3-Tyr residue, was synthesized and the integrity of the disulfide bond of the peptide was investigated in the presence of reducing agents such as ascorbic acid, dithionite, and stannous ions. The disulfide bond in [125I]I-RC-160 remained stable in the presence of ascorbic acid in boiling water. The disulfide bond was also stable when treated with stannous ions at concentrations sufficient to reduce 99mTc for complexation with a thiol-free chelating agent, bis(hydroxamamide) analog when the 99mTc complexation reaction was performed at room temperature. However, the disulfide bond of [125I]I-RC-160 was slightly cleaved in the presence of a small amount of stannous ions when the reaction was performed in boiling water. Treatment of [125I]I-RC-160 with dithionite in boiling water markedly reduced the disulfide bond of the parental peptide. These findings indicated that synthetic somatostatin analogs may be labeled with 99mTc with stannous ions as the reducing agent without impairing their structure after conjugation of thiol-free chelating agents that provide 99mTc chelates under mild reaction conditions. PMID- 10708302 TI - Synthesis and biodistribution of R- and S-isomers of [18F]-fluoropropranolol, a lipophilic ligand for the beta-adrenergic receptor. AB - The S and R isomers of [18F]-fluoropropranolol (1-[1-fluoro-2-isopropylamino]-3 naphthalen-1-yloxy-propan-2 -ol) have been prepared by reductive alkylation of the appropriate aminoalcohols. The radiosynthesis provides a reasonable yield (approximately 25%) to give products of 99% enantiomeric excess and specific activities of 1-3 Ci/micromol. The dissociation constants for the beta2 adrenergic receptor are 0.5 and 2.5 nM for the S and the R isomers, respectively. The biodistribution data in rats show that uptake and egress of the tracer is rapid but that the result of blocking studies and the difference between the R and the S isomers suggest receptor-mediated uptake in receptor-rich tissue. PMID- 10708303 TI - 2Beta-carbomethoxy-3beta-(4- and 2-[18F]fluoromethylphenyl)tropanes: specific probes for in vivo quantification of central dopamine transporter sites. AB - Dopamine reuptake transporter binding kinetics of 2beta-carbomethoxy-3beta-(4 [18F]fluoromethylphenyl)tropane (p-FWIN) and 2beta-carbomethoxy-3beta-(2 [18F]fluoromethylphenyl)tropane (o-FWIN) were determined in vervet monkeys using positron emission tomography (PET). Ligand localization was rapid and specific to the striatum with kinetic estimates comparable with those of 11C-labeled WIN 35,428 (CWIN). Binding was more specific with p-FWIN than with CWIN or o-FWIN. The relatively longer half-life of the 18F radiolabel enabled longer acquisition times with p-FWIN, resulting in less variability in the kinetic estimates. PMID- 10708304 TI - Interactions of 16alpha-[18F]-fluoroestradiol (FES) with sex steroid binding protein (SBP). AB - Fluorine-18 16alpha-Fluoroestradiol ([18F]-FES) is a positron-emitting tracer for the estrogen receptor that is used for positron emission tomography (PET) studies of tumor tissues rich in the estrogen receptor. The role of the sex steroid binding protein (SBP or SHBG) in the transport of the [18F]-FES to the estrogen receptor-rich tissue in breast cancer patients in vivo was investigated. To determine the extent to which [18F]-FES is bound to SBP in the blood, we performed a series of studies using blood samples obtained from patients undergoing [18F]-FES PET scans. The binding of [18F]-FES to the SBP was measured using a simple protein precipitation assay. The binding of [18F]-FES metabolites to SBP was also measured. These measurements showed that the tracer was distributed between albumin and SBP, and the binding capacity of SBP was sufficient to ensure that the protein was not saturated when the tracer was fully mixed with the plasma; however, local saturation of SBP may occur when [18F]-FES is administered intravenously. Typically about 45% of [18F]-FES in circulating plasma was bound to SBP, but this fraction was dependent on the concentration of SBP in plasma. The transfer of the tracer between the two proteins was rapid, complete in less than 20 s at 0 degrees C, suggesting that the equilibrium was maintained under most circumstances and that local saturation resolved quickly when blood from the injection site entered the central circulation. These data suggest that SBP binding of [18F]-FES is significant and will affect the input function of the tracer for any model that is used for the quantitative evaluation of [18F]-FES uptake in PET studies. Estimates of equilibrium binding in blood samples are sufficient to characterize [18F]-FES binding to SBP in the circulation. PMID- 10708305 TI - Synthesis and in vivo evaluation of [11C]SA6298 as a PET sigma1 receptor ligand. AB - The potential of a 11C-labeled selective sigma1 receptor ligand, 1-(3,4 dimethoxyphenethyl)-4-[3-(3,4-dichlorophenyl)propyl]piperazine ([11C]SA6298), was evaluated as a positron emission tomography (PET) ligand for mapping sigma, receptors in the central nervous system and peripheral organs. [11C]SA6298 was synthesized by methylation of the desmethyl SA6298 with [11C]CH3I, with the decay corrected radiochemical yield of 39 +/- 5% based on [11C]CH3I and with the specific activity of 53 +/- 17 TBq/mmol within 20 min from end of bombardment (EOB). In mice, the uptake of [11C]SA6298 was significantly decreased by carrier loading in the brain, liver, spleen, heart, lung, small intestine, and kidney in which sigma receptors are present as well as in the skeletal muscle. Pretreatment with SA6298 also blocked the uptake of [11C]SA6298 by these organs except for the small intestine, but significant displacement of [11C]SA6298 by posttreatment with SA6298 was observed only in the heart, lung, and muscle. In the blocking study with one of the eight sigma receptor ligands, including haloperidol, SA6298, NE-100, (+)-pentazocine, SA4503, (-)-pentazocine, (+)-3-PPP, and (+)-SKF 10,047 (in the order of the affinity for sigma1 receptor subtype), only SA6298 and an analog SA4503 significantly reduced the brain uptake of [11C]SA6298 to approximately 80% of the control, but the other six ligands did not. Peripherally, the uptake of [11C]SA6298 by the organs described above was decreased predominantly by SA6298 or SA4503, but the blocking effects of the other five ligands except for NE-100 depended on their affinity for sigma1 receptors. The saturable brain uptake of [11C]SA6298, approximately 20%, was also observed by tissue dissection method in rats and by PET in a cat. Ex vivo autoradiography of the rat brain showed a high uptake in the cortex and thalamus. In the cat brain a relatively high uptake was found in the cortex, thalamus, striatum, and cerebellum. These results have indicated a receptor-mediated uptake of the tracer to some extent in the brain and peripheral organs. However, the tracer has a limited potential for the PET study of the brain receptors because of a relatively high nonspecific binding. PMID- 10708306 TI - High yield direct 76Br-bromination of monoclonal antibodies using chloramine-T. AB - Monoclonal antibody (MAb) A33 was labeled with the positron emitter 76Br (T(1/2) = 16.2 h). Direct labeling was done using the conventional chloramine-T method. After optimization of the labeling conditions, a maximum yield (mean +/- max error) of 77 +/- 2% was obtained at pH 6.8. In vitro binding of 76Br-A33 to SW1222 colonic cancer cells showed that the immunoreactivity was retained. Also, the MAbs 38S1 and 3S193 and the peptide hEGF were 76Br-labeled, resulting in labeling yields (mean +/- max error) of 75 +/- 3%, 63 +/- 4%, and 73 +/- 0.1%, respectively. We conclude that antibodies and peptides can be labeled conveniently with 76Br for the purpose of whole-body tumour imaging by positron emission tomography. PMID- 10708307 TI - Biodistribution of liposomal 131I-VIP in rat using gamma camera. AB - Vasoactive intestinal peptide (VIP), a 28 amino-acid peptide was labeled with 131I and encapsulated into liposomes. 131I-VIP or liposomal 131I-VIP was administered intravenously into the rats. The distribution was studied by a gamma camera and established by counting the radioactivity in the removed organs. The elimination half-life for the liposomal 131I-VIP in both blood and lungs was significantly longer (5.29 and 9.28 min, respectively) than that obtained after the administration of 131I-VIP (0.62 and 3.18 min, respectively). Dynamic scans using a gamma camera after the administration of liposomal 131I-VIP showed a higher uptake of the liposomal form into the lungs compared with 131I-VIP. The lack of VIP in asthmatics has been shown in previous studies. However, the clinical investigations using VIP were disappointing most probably due to the rapid degradation of the peptide in the bronchial tract. This in fact is supported by our previous study, in which we demonstrated that VIP had a half life of 0.45 min in blood. We conclude that the encapsulation of VIP in liposomes prolongs its elimination half-life in plasma and enhances its uptake in lungs. This observation may increase the clinical use of VIP in both diagnostic and therapy. PMID- 10708308 TI - Interaction of a monoclonal antibody against hEGF with a receptor site for EGF. AB - Epidermal growth factor (EGF) has been detected by radioimmunoassay (RIA) in different body fluids such as serum, amniotic fluid, and urine. Human tumor tissues with EGF receptors (EGF-Rc) may be saturated with EGF, which may be of prognostic value. An RIA was envisaged to measure human epidermal growth factor (hEGF) levels using EGF-Rc as capture agent and a monoclonal antibody anti-hEGF (MAb anti-hEGF) labeled with 125Iodine as a marker for this binding. The purpose of this work was to study the feasibility of MAb anti-hEGF to detect the receptor binding sites and to investigate the interaction between MAb anti-hEGF and the EGF-Rc. Various binding experiments were performed to study possible interference and interactions in the complex MAb anti-hEGF and the receptor. Affinity constants were determined by means of Scatchard plot analysis to interpret the complex stability challenged with other compounds for a better understanding of the interaction process. Binding constants were of the same order for all the ligands tested separately involving the EGF-Rc, but were significantly higher (t = 15.7, p < 0.05) for hEGF in its binding to MAb anti-hEGF. It was possible with equilibrium studies and competition experiments to evaluate the interaction of EGF and MAb anti-hEGF with the EGF receptor. This observation makes the MAb anti hEGF a potential tracer for the quantitation of receptors in vitro, and possibly for the detection of membrane receptors on tumor cells in vivo. PMID- 10708309 TI - A triglycine linker improves tumor uptake and biodistributions of 67-Cu-labeled anti-neuroblastoma MAb chCE7 F(ab')2 fragments. AB - The peptide-linked copper chelators CPTA-triglycyl-L-p-isothiocyanato phenylalanine (CPTA-R1-NCS) as well as DOTA-triglycyl-L-p-isocyanato phenylalanine (DOTA-R1-NCS) were synthesized and coupled to F(ab')2 fragments of the anti-neuroblastoma monoclonal antibody (MAb) chCE7. 67Cu-labeled conjugates were compared with the original CPTA- and DO3A-F(ab')2 in vitro and in vivo in mice bearing neuroblastoma xenografts. With the CPTA-R1-F(ab')2, biodistributions were improved, because radioactivity present in the kidney was reduced. With the DOTA-R1-F(ab')2, clearance from the blood was slower and tumor uptake was higher compared with the other conjugates. DOTA-R1-F(ab')2 achieved the best tumor/tissue ratios. PMID- 10708310 TI - Rhodium-105 tetrathioether complexes: radiochemistry and initial biological evaluation. AB - 105Rhodium(III) complexes with three different acyclic tetrathioether ligands containing pendant carboxylic acid groups have been synthesized and characterized. The complexes were evaluated for stability under physiological conditions and the most promising complexes were evaluated in vivo in normal mice. The primary route of clearance for these complexes was the renal/urinary system, consistent with the presence of pendant carboxylate groups. The results indicate that the cis-[Rh(III)Cl2(2,5,8,11-tetrathiadodecane-1,12-dicarboxylic acid)]+ complex shows the most promising in vivo characteristics on which to base a potential therapeutic radiopharmaceutical. PMID- 10708311 TI - Effects of methoxyflurane anesthesia on the pharmacokinetics of 125I-IAZA in Sprague-Dawley rats. AB - Effects of methoxyflurane anesthesia on the pharmacokinetics of intravenous 125I IAZA in rats are reported. No significant differences in t(1/2alpha), t(1/2beta), V(SS), and ClTB for total radioactivity (125I-IAZA and metabolites) were observed between the anesthetized (Group 1, n = 4) and nonanesthetized (Group 2, n = 3) animals. For 125I-IAZA, ClTB increased from 646 +/- 52 mL/h/kg to 2250 +/- 351 mL/h/kg and t(1/2beta) decreased from 97.7 +/- 17.5 min to 35.6 +/- 5.4 min, for Groups 1 and 2, respectively. There were no differences in V(SS) or t(1/2alpha) between the two groups. These findings support literature reports of anesthetic effects on xenobiotic pharmacokinetics, and indicate a need for caution in the evaluation of preclinical imaging studies in which animals are immobilized with anesthetics. PMID- 10708312 TI - Rhenium-188-Labeled DTPA: a new radiopharmaceutical for intravascular radiation therapy. AB - Balloon angioplasty is a standard treatment for artherosclerotic coronary artery disease. However, its clinical value is reduced by a high restenosis rate. A new concept in preventing restenosis is the use of a liquid-filled balloon containing a beta-emitting radioisotope. In this study, we performed biodistribution studies of Re-188 perrhenate and Re-188 diethylenetriaminopentaacetate (DTPA) to assess the resulting organ dose values in the event of balloon rupture if these agents are used for the clinical inhibition of restenosis after percutaneous transluminal coronary angioplasty (PTCA). After injecting Re-188 preparations intravenously, rats were killed at 10 min, 30 min, 60 min, 2 h, and 6 h (n = 5 per group). Tissue concentrations were calculated and expressed as percent injected dose per gram or per milliliter (%ID/g or %ID/mL). In addition, urine excretion and thyroid gland uptake were evaluated in rats (n = 5 per group) with a gamma camera after administration of 37 MBq (1 mCi) of each agent. Our data showed that both agents were excreted primarily via urine. However, the excretion of Re-188 DTPA was much faster than that of Re-188 perrhenate via the urinary system. The biodistribution data revealed that radioactivity levels in the stomach and the thyroid gland were high in the perrhenate group but low in the Re 188 DTPA group. The concentration levels in other tissues including lung, liver, testis, muscle, and blood were low throughout this study for both agents. The thyroid radiation value in the Re-188 perrhenate group was 0.163 mGy/MBq, which was much higher than that of the Re-188 DTPA group (0.0167 mGy/MBq). The stomach radiation value was as high as 0.127 mGy/MBq for Re-188 perrhenate, compared with 0.013 mGy/MBq for Re-188 DTPA. In conclusion, in the event of balloon rupture, the release of Re-188 DTPA results in lower radiation doses than Re-188 perrhenate, especially to the thyroid gland and the stomach. Our data suggest that Re-188 DTPA is a useful radiopharmaceutical for endovascular irradiation. PMID- 10708313 TI - Comparison of various rhenium-188-labeled diphosphonates for the treatment of bone metastases. AB - In the past, many diphosphonates were introduced as bone scan radiopharmaceuticals. In addition, diphosphonates have been labeled with beta emitted isotopes and developed into useful therapeutic drugs for bone metastases. However, it is not clear which diphosphonate is the best choice when labeling with Re-188. In this study, we labeled methylene diphosphonate (MDP), hydroxyethylidene diphosphonate (HEDP), and hydroxymethane diphosphonate (HDP) with Re-188. Each radiopharmaceutical was further evaluated in two conditions (with and without carrier). Twenty-four rabbits were used (four in each group) for the analysis of the biodistributions and bone uptakes of these radiopharmaceuticals to assess their potential for clinical applicability. Four hours after intravenous injection of approximately 37 MBq (1 mCi) Re-188-labeled diphosphonate preparations, whole body scans were performed using a large-field gamma camera equipped with a high resolution collimator. Bone-to-soft tissue ratios (B/S ratio) were calculated using a computer program. Our data showed that Re-188 HEDP with carrier (10(-4) M carrier) could accumulate in the skeletal system whereas very little absorption by bone was observed in the rabbits that were injected with carrier-free Re-188 HEDP. In addition, no significant bone uptake was demonstrated for Re-188 MDP or Re-188 HDP, with or without carrier. The B/S ratio was 25.06 in the Re-188 HEDP with carrier group but less than 3 in the other groups. In conclusion, HEDP is the best choice among these three bone seeking drugs when labeled with Re-188. But, it is necessary to add carrier when preparing Re-188 HEDP for the treatment of bone metastases. PMID- 10708314 TI - Spectrophotometric method for determination of bifunctional macrocyclic ligands in macrocyclic ligand-protein conjugates. AB - A simple spectrophotometric assay for determination of bifunctional polyazacarboxylate-macrocyclic ligands of different sizes that are conjugated to proteins has been developed for: 12-membered macrocycle DOTA (2-[4-nitrobenzyl] 1, 4, 7, 10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid) and analogs, the 15-membered PEPA macrocycle (2-[4-nitrobenzyl]-1,4,7,10,13 pentaazacyclopentadecane-N,N',N'',N ''',N''''-pentaacetic acid), and the large 18 membered macrocycle HEHA (1,4,7,10,13,16-hexaazacyclooctadecane N,N',N'',N''',N'''',N'''''- hexaacetic acid). The method is based on titration of the blue-colored 1:1 Pb(II)-Arsenazo II (AAIII) complex with the polyazacarboxylate macrocyclic ligand in the concentration range of 0-2.5 microM, wherein color change occurring upon transchelation of the Pb(II) from the AAIII to the polyazamacrocyclic ligand is monitored at 656 nm. The assay is performed at ambient temperature within 20 min without any interfering interaction between the protein and Pb(II)-AA(III) complex. Thus, this method also provides a ligand to-protein ratio (L/P ratio) that reflects the effective number of ligands per protein molecule available to radiolabeling. The method is not suitable for 14 membered TETA macrocycle (2-[4-nitrobenzyl]-1, 4, 8, 11-tetraazacyclotetradecane N,N',N'',N'''-tetraacetic acid) because of low stability constant of Pb(II)-TETA complex. The method is rapid, simple and may be customized for other polyazacarboxylate macrocyclic ligands. PMID- 10708315 TI - Using interactive voice response technology and timeline follow-back methodology in studying binge eating and drinking behavior: different answers to different forms of the same question? AB - As part of a study of the relationship of binge eating, alcohol use, mood, and stressors, we compared the results of two forms of reporting on binge eating and drinking behavior. Forty-three first-year college women participated in an interactive voice response (IVR) study for 12 weeks. Participants answered computer-administered questions daily via IVR technology on number of eating binges and number of alcoholic drinks consumed. After 12 weeks, participants completed a Timeline Follow-back (TLFB) interview retrospectively for number of binges and drinks in the past 12 weeks. Results of this distally retrospective methodology (commonly used in drinking research and applied here also to binge eating) were compared to the results of daily IVR reporting. There was convergence across measures for drinking behavior, but divergence between IVR and TLFB for binge eating reports. TLFB reports underrepresented actual binge eating frequency, which calls into question the validity of applying this methodology to the assessment of binge eating. PMID- 10708316 TI - The relationship between alcohol problems and use of tranquilizing drugs: longitudinal patterns among American women. AB - A previous community study of older adults (Graham et al., 1996) indicated a relationship between alcohol problems and use of tranquilizing drugs despite no relationship between alcohol consumption and tranquilizer use. The present paper explores this issue further using longitudinal data from a representative sample of American women. The results replicated previous findings of a significant relationship between alcohol problems and tranquilizer use that was unrelated to alcohol consumption. Analyses of longitudinal patterns indicated that alcohol problems in 1981 predicted subsequent use of tranquilizing drugs and that this relationship may be moderated by anxiety, with the relationship being strongest for respondents who reported few or no problems with anxiety. The results indicated no support for the relationship being due to: a pharmacological interaction of alcohol with tranquilizing drugs; use of tranquilizing drugs precipitating alcohol problems; or depression, anxiety, poor health or childhood sexual abuse being common causes of both alcohol problems and tranquilizer use. The link between alcohol problems and use of tranquilizing drugs needs to be investigated further to increase understanding of addictive behaviors. PMID- 10708317 TI - Direct and indirect effects of nicotine/smoking on cognition in humans. AB - Acute nicotine administration and acute nicotine withdrawal have been shown to have effects on mood, arousal, and cognition in humans. However, given what is known about the interrelationships between mood, arousal, and cognition in the psychology literature, it is unclear to what extent these effects on a given response are direct, or mediated through other responses (indirect). This article discusses this issue with particular emphasis on the question of whether nicotine has direct effects on cognition. Five ways are suggested as to how this question can be resolved, the relevant literature is reviewed, and some research ideas are proposed. PMID- 10708318 TI - Addictive behaviors and depression among African Americans residing in a public housing community. AB - Numerous studies have indicated that there is an association between cigarette smoking, alcohol use, and depression. However, little attention has been devoted to understanding how demographic factors, such as socioeconomic status and ethnicity, influence these relationships. To address this gap in the literature, cigarette and alcohol use were examined in a sample of African Americans from an urban area. A single public-housing community in Washington, DC was selected for complete ascertainment of the adult population. A total of 126 African American subjects were recruited. Semi-structured interviews were conducted to assess depressive symptoms and to characterize cigarette and alcohol use patterns. Cigarette smoking was not related to the severity of depressive symptoms. By contrast, increased symptoms of depression were related to alcohol use patterns. Light drinkers had a mean score of 5.77 on the Centers for Epidemiologic Studies Depression Scale, compared to a mean of 8.30 for abstainers and 10.07 for heavy drinkers (F = 4.968, p < .003). An analysis of patterns of substance use revealed that subjects were more likely to either abstain from both substances (30.2%) or to use both substances (32.5%) (chi2 = 8.516, df = 1. p < .004). It is unclear which specific processes work to link alcohol use and depressive symptoms in this group of urban African Americans from a low-income community. What is clear is that alcohol use is clearly related to depressive symptoms in the sample. It is hypothesized that both self-medicating processes and substance-induced depressive symptoms may be responsible for these findings. Important factors to consider in developing effective intervention programs that target this specific population are discussed. PMID- 10708319 TI - Development of the drug abuse screening test for adolescents (DAST-A). AB - The development and initial validation of the Drug Abuse Screening Test for Adolescents (DAST-A) is summarized. The DAST-A, derived from a modification of the original adult version called the Drug Abuse Screening Test (DAST: Skinner, 1982), was psychometrically tested in a study group of adolescent inpatients. The DAST-A demonstrated good internal consistency, high test-retest reliability, unidimensional factor structure, and good concurrent validity. Using the classification system of the fourth edition of the Diagnostic and Statistical Manual of Mental Disorders (American Psychiatric Association 1994), DAST-A scores of greater than 6 yielded sensitivity, specificity, and positive predictive powers of 78.6%, 84.5%, and 82.3%, respectively, in differentiating adolescent psychiatric inpatients with and without drug-related disorders. These findings suggest that the DAST-A holds promise as a drug abuse screening measure in psychiatrically impaired adolescent populations. PMID- 10708320 TI - Perceived invulnerability and cigarette smoking among adolescents. AB - Adolescent perceptions of invulnerability toward smoking and nonsmoking-related health risks were examined among 442 continuation high school students. Smokers were less likely than nonsmokers to report feeling invulnerable to both smoking and nonsmoking-related health risks. Among the smokers, those who reported feeling invulnerable to smoking-related health risks, compared to those who reported feeling vulnerable, smoked fewer cigarettes, were less addicted, were less likely to intend to smoke more in the future, attempted to quit fewer times in the past, valued their health more, and reported higher public body awareness. In a multiple logistic regression model, only high public body awareness, fewer previous attempts to quit, and being in the action stage of change (compared to being in the precontemplation stage of change) remained significant independent concurrent predictors of being in the invulnerable group. These results suggest, contrary to some previous work, that perceived invulnerability may be predictive of quitting tobacco use and may reflect relative invulnerability; that is, lighter use of tobacco. PMID- 10708321 TI - Twelve-month follow-up of a smoking relapse prevention intervention for postpartum women. AB - This study examined the long-term effectiveness of a postpartum smoking relapse prevention intervention by evaluating the smoking status and smoking cessation self-efficacy of original study participants at 12 months following delivery. Two hundred and thirty-eight women who had participated in a randomized clinical trial, a nurse-delivered relapse prevention intervention, were visited in their homes. Data were collected on smoking status, self-efficacy, mental health, alcohol use, breast feeding, social support, smoking in the social environment, and sociodemographics. Smoking status was verified with measures of carbon monoxide in expired air. The 12-month continuous smoking abstinence rate was 21.0% in the treatment group and 18.5% in the control group; odds ratio (OR) = 1.17, 95% confidence interval (CI) = 0.62-2.22. One half (50.4%) of the control group and 41.2% of the treatment group reported smoking daily at 12 months; OR = 1.45, 95% CI = 0.87-2.43. The treatment group attained higher self-efficacy. Four variables were associated with relapse to daily smoking; breast feeding and mental health had protective effects, while partners who smoked and greater amount smoked prior to pregnancy had adverse effects. PMID- 10708322 TI - Expectancies for alcohol to affect tension and anxiety as a function of time. AB - Alcohol outcome expectancies have been linked to drinking behavior on both empirical and theoretical grounds. Although typically measured as a static construct, we hypothesized that expectancies may be time-specific. Subjects rated their expectancies for a moderate amount of alcohol to increase, decrease, or not change their level of tension and anxiety. Ratings were repeated for when the intoxicating effects of the drinking would be: (1) "at their peak;" (2) "nearly worn off;" and (3) "completely worn off" (Time Epochs 1-3, respectively). As predicted, most subjects (72%) expected alcohol to reduce tension and anxiety at Time Epoch 1; however, significantly fewer subjects expected this effect at Time Epochs 2 and 3 (25% and 2%, respectively). Conversely, few subjects expected alcohol to worsen tension and anxiety at Time Epoch 1 (3.5%); however, significantly more subjects expected this effect at Time Epochs 2 and 3 (31% and 34%, respectively). Expectancies for Time Epoch 1 related most strongly to several measures of alcohol use, including drinking for the purpose of reducing tension (whole sample) and drinking frequency (men but not women). These findings show that tension-reduction expectancies are not stable over the course of a drinking episode and suggest the possibility of a treatment approach aimed at amplifying attention to expectancies for alcohol's more negative longer-term effects. PMID- 10708323 TI - Influence of individual differences in craving and obsessive cocaine thoughts on attentional processes in cocaine abuse patients. AB - In the present pilot-study, the relation between craving, obsessive thoughts about cocaine, experienced control, and attentional bias for cocaine related words is investigated. Sixteen cocaine abuse patients participated in a reaction time (RT) experiment which was employed to measure the ability of subjects to shift their attention away from cocaine related words. Postexperiment craving was found to be positively correlated with reaction times on drug related cues, in contrast to RT on neutral cues. Furthermore, obsessive thoughts about cocaine use and the experienced cocaine use control, in the week before the experiment, were correlated in a higher degree with RTs on drug cues than postexperiment craving. Attentional bias for drug cues was evidenced in patients with higher scores on obsessive cocaine thoughts and higher craving scores. This study shows that individual differences on information processing, within a cocaine abuse patient population, are present. PMID- 10708324 TI - Smokers can learn to influence their urge to smoke. AB - Forty heavy smokers participated in a within-subject experiment in which the association between smoking-related cues and nicotine intake was made conditional on two neutral stimuli. Two colored cards indicated whether smoking-related cues, placed on the cards, would or would not be followed by nicotine intake. In the presence of each card, subjects were asked to rank their urge to smoke before and during the exposure to the smoking cues. The results of the present study revealed that the predictive value of a cue, in regard to the occurrence of nicotine intake, strongly determines its ability to generate craving. It was concluded that participants learned a modified predictive value of smoking cues, through a process of conditioning, and in this way influenced their urge to smoke. Furthermore, the findings suggest the reconditioning of CS-US associations as an aid in the treatment of smoking addiction. PMID- 10708325 TI - Sensation seeking and drinking game participation in heavy-drinking college students. AB - Previous research has identified differences between heavy-drinking students who play drinking games and those who do not. Johnson, Wendel, and Hamilton (1998) suggested that heavy-drinking players may correspond to Cloninger's (1987) Type II alcoholic and that heavy-drinking nonplayers resemble Type I. The current study predicted that (a) sensation seeking would be associated with greater frequency of play and greater frequency of negative consequences from play and that (b) heavy-drinking students who play drinking games would be higher in sensation seeking than heavy-drinking students who do not play. A sample of 172 female and 84 male college students completed the Sensation Seeking Scale Form V, questions about quantity and frequency of alcohol consumption, and questions regarding drinking game participation. Higher levels of sensation seeking predicted greater frequency of play even after controlling for overall quantity and frequency of consumption. Sensation seeking was also related to specific motives for play. Men who were higher in sensation seeking experienced more negative alcohol-related consequences as a result of play. In women, but not in men. heavy-drinking players were higher in sensation seeking than heavy nondrinking nonplayers. The results of the current study do not clearly support Cloninger's model, but they are consistent with other research concerning the role of sensation seeking and risk taking in contributing to negative alcohol related consequences. Personality style likely interacts with social norms and contextual factors in influencing drinking game participation and consequences of play. PMID- 10708326 TI - Modulating effect of alcohol use on cocaine use. AB - Clinical observations have indicated that alcohol may be employed by cocaine/crack users to attenuate negative effects of cocaine, especially when "coming down" from a cocaine binge. This issue was examined by interviewing 66 dual cocaine/alcohol users, with opiate dependence histories, enrolled in methadone treatment. A path analysis model was specified to test several hypotheses concerning the possible modulating effects of alcohol use on cocaine use. About 60% of the subjects reported often employing alcohol to ameliorate discomfort associated with tapering or ceasing cocaine/crack use. The main findings were: (a) more intense cocaine/crack craving and feeling that cocaine/crack use was out of control both led to increased use of alcohol to come down; (b) the more frequently alcohol was used to come down, the less use of cocaine/crack; and (c) more cocaine/crack use and more use of alcohol to come down both led to increased heavy alcohol use. Thus, treating alcohol abuse in this population must take into account the important function it serves in modulating cocaine/crack use. PMID- 10708327 TI - Evaluation of substance use outcomes in demonstration projects for pregnant and postpartum women and their infants: findings from a quasi-experiment. AB - We evaluated the impact of nine community-based drug prevention, education, and treatment projects for pregnant and postpartum women using a quasi-experimental design. Projects provided case management and referral to services or provided day treatment. Self-reported measures of six substances were collected (a) from intake to delivery and (b) delivery to 6 months postpartum. We hypothesized that women who received project services (n = 370) between intake and follow-ups would be more likely to deter or reduce substance use than women who received an alternative or no intervention (n = 288). Data showed that project clients had significantly lower 30-day use rates on four of the measures-alcohol, any illicit drug(s), marijuana, and crack-from intake to delivery, with preintervention alcohol and other drugs use controlled. However, none of these results was maintained from intake through 6 months postpartum. Findings are discussed in terms of the difficulty of detecting and documenting promising intervention program effects over time in this population. PMID- 10708328 TI - Correlates of life stress in an alcohol treatment sample. AB - Men in alcohol dependence treatment commonly report elevated levels of stressful life experiences prior to entering treatment. Clinical researchers have argued that training patients to cope with stress is an important aspect of therapy for alcohol dependence. Current age and age of onset of alcohol dependence are two patient characteristics that may relate to stress and allow mental health care providers to anticipate patient needs more readily. This study examines whether current age and age of onset of alcohol dependence are related to the type of stress experienced preceding treatment. Participants were 350 male veterans receiving inpatient treatment for alcohol dependence who completed a semistructured interview-based life stress assessment. The likelihood of life events related to achievement, relationship, and legal problems decreased with age; the likelihood of health difficulties increased with age; and age and age of onset of alcohol dependence interacted to predict health difficulties. These results indicate that the nature of some stress experiences prior to treatment for alcohol dependence may be related to current age and the course of alcohol dependence. PMID- 10708329 TI - New perspectives on the manipulation of opiate urges and the assessment of cognitive effort associated with opiate urges. AB - Behavioral models of drug urges assume that conditioned urges are strongly associated with drug consumption. An alternative, cognitive model assumes that urges represent the operation of cognitively demanding processes devoted to either supporting or blocking the automatized drug-use behavior. In Study 1, the effect of verbal drug cues and mood induction on self-reported opiate urges were examined. Twenty-four opiate addicts were either instructed to listen to verbal drug cures or neutral cues. Negative mood induction was applied on 12 addicts. Study 2 examined the cognitive processes underlying these urges. In a dual task paradigm, participants responded to a probe stimulus and listened simultaneously to an imagery script. It was predicted that response times to the probe should increase to the extent that urge-related nonautomatic processing is invoked. Although negative affect was not associated with nonautomatic processing, the findings suggest that drug urges can be activated by drug cues and that cue related urges are supported by nonautomatic cognitive processes. PMID- 10708330 TI - Cultural, social, and intrapersonal factors associated with substance use among alternative high school students. AB - The purpose of this study was to identify cultural, social, and intrapersonal factors associated with tobacco, alcohol, and illicit drug use among students attending dropout prevention/recovery high schools. Four mutually exclusive categories of substance use were used as outcome measures, and religiosity, educational achievement, educational aspiration, family caring, others caring, self-esteem, optimism, coping, depression, loneliness, and self-efficacy were used as predictor variables. In the final multivariate model more family caring and loneliness were inversely associated with marijuana use; young age, more family caring, less coping ability, church attendance, and low educational aspirations were significantly associated with cocaine use. This study demonstrates the importance of health education and health promotion programs for students attending alternative high schools which include prevention of initiation, as well as treatment. PMID- 10708331 TI - A comparison of motives for marijuana and alcohol use among experienced users. AB - Motivational models suggest that individuals use substances to achieve desired effects. Given different pharmacological effects across drug classes, and variations in social context, one would expect that the motives instigating use differ by drug class. However, commonalties in motives across drugs have also been observed. The purpose of this study was to examine similarities and differences across a common set of motives for alcohol and marijuana among experienced users of both drugs. Participants were 46 college students (21 women) who completed a motives assessment twice, once for marijuana and once for alcohol. All had used each drug 60 or more times in their lifetime. Social motives were more highly endorsed for alcohol than marijuana. Expansion motives were more highly endorsed for marijuana. Enhancement motives were more highly endorsed for marijuana than alcohol among women but not men. Endorsement of coping and conformity motives did not differ across drugs. Experienced users of marijuana and alcohol discriminate between their reasons for using the two drugs. These findings are discussed with regard to the differentiation between and commonalties among substances of abuse. PMID- 10708332 TI - Variables modulating stress and coping that discriminate drug consumers from low or nondrug consumers. AB - The present study assesses how certain stress and coping variables relate to drug use. A total of 124 subjects (85 men, 39 women) took part. They were divided into two groups: consumers and low or nonconsumers. Results indicate that consumers show significantly lower scores in self-control than low or nonconsumers. This research is of interest because it analyzes the relationship among assertiveness, self-control, self-efficacy, and drug use. We consider it worthwhile to take these variables into account when planning prevention programs. PMID- 10708333 TI - Mechanisms of parenchymal cell death in-vivo after microvascular hemorrhage. AB - OBJECTIVE: In vitro studies suggest that microhemorrhages with escape of red cells into the tissue may be cytotoxic to parenchymal cells due to oxygen free radical formation. We examined in the rat mesentery the impact of microhemorrhages on parenchymal cell death, as detected by propidium iodide staining, using an intravital approach. METHODS AND RESULTS: Postcapillary venules were punctured with a closed-end micropipette, permitting escape of blood cells and plasma into the mesentery interstitium. Over a period of 2 h, no significant increase in parenchymal cell death was encountered in tissues with hemorrhagic sites compared with nonhemorrhagic control sites. Interstitial microinjections of plasma derived from whole blood incubated for several hours with and without a combination of sodium azide (2 mM) and hydrogen peroxide (1 mM) led to significantly increased levels of cell death compared to control experiments. Interventions against the hydroxyl radical with dimethylthiourea (DMTU, 2 mM) or 2,2'-dipyridyl (DPD, 2 mM), a lipid soluble iron chelator, provided no protective effect against the parenchymal cell death. DMTU slightly delayed tile cytotoxic reaction. CONCLUSIONS: These observations suggest that a newly formed microhemorrhage is not necessarily cytotoxic to parenchymal tissue cells. Interstitial microinjections of plasma, derived from whole blood after prolonged exposure to oxygen free radicals or just aging under in vitro conditions, may be cytotoxic to mesenteric parenchymal cells without effective blockade by interventions against the hydroxyl radical. PMID- 10708334 TI - Endothelial cells exposed to anoxia/reoxygenation are hyperadhesive to T lymphocytes: kinetics and molecular mechanisms. AB - OBJECTIVE: The objectives of this study were to 1) determine the time-course of T lymphocyte adhesion to monolayers of human umbilical vein endothelial cell (HUVEC) that were exposed to 60 min of anoxia followed by 24 h of reoxygenation, and 2) define the mechanisms responsible for the hyperadhesivity of postanoxic HUVEC to human T-lymphocytes. METHODS: Human peripheral blood mononuclear leukocytes were isolated from heparinized peripheral blood. T-lymphocytes were obtained by negative selection using a MACS column. HUVEC monolayers were exposed to anoxia/reoxygenation (A/R), and then reacted with 51Cr -labeled T-lymphocytes in adhesion assays. RESULTS: A/R leads to an increased adhesion of T-lymphocytes to HUVEC monolayers, with peak responses occurring at 8 h after reoxygenation. This adhesion response was largely attributed to the CD4+ T-cell subset. The hyperadhesivity of A/R-exposed HUVEC was inhibited by monoclonal antibodies directed against either LFA-1, VLA-4, ICAM-1, or VCAM-1, indicating a contribution of these adhesion molecules and their ligands. Moreover, T-cell hyperadhesivity was attenuated by anti- IL-8. consistent with a role for this chemokine in the adhesion response. Protein synthesis inhibitors (actinomycin D and cycloheximide) as well as chemical inhibitors of (and binding ds oligonucleotides to) NFkappaB and AP-1 significantly attenuated the A/R-induced T lymphocyte adhesion responses. The kinetics of VCAM-1 on post-anoxic HUVEC correlated with the T-lymphocyte adhesion response. CONCLUSIONS: A/R elicits a T lymphocyte-endothelial cell adhesion response that involves transcription dependent surface expression of VCAM-1. PMID- 10708335 TI - Adaptation of ion channels in the microcirculation to exercise training. PMID- 10708336 TI - The inflammatory reaction during venous hypertension in the rat. AB - OBJECTIVE: Numerous studies have examined arterial occlusion followed by reperfusion but few studies have reported about venular occlusion which, in contrast to arterial occlusion, is associated with elevation of the capillary blood pressure. Here we examine leukocytes infiltration and tissue injury in rat mesentery during local venular occlusion and venous hypertension followed by reperfusion, and determine the level of protection offered by pretreatment with micronized, purified flavonoid fraction (MPFF). METHODS: Leukocyte rolling, adhesion, migration, and parenchymal cell death as detected by propidium iodide labeling were determined during venular occlusion using a micropipette followed by reperfusion in the rat mesenteric microcirculation pretreated with 0, 50, or 100 mg/kg MPFF for 7 days. Spontaneous leukocyte activation by nitroblue tetrazolium reduction and expression of CD18 and CD62L on naive donor neutrophils incubated with plasma from each treatment group were determined. RESULTS: Venous occlusion led to elevated levels of leukocyte rolling, adhesion, and migration as well as parenchymal cell death. These injurious processes were significantly inhibited by MPFF in a those-dependent fashion. MPFF reduced spontaneous leukocyte NBT reduction and the neutropil expression of CD62L, even though CD18 was not affected. CONCLUSION: These results suggested that microvascular occlusion in venules with elevation of the micropressure followed by reperfusion is a highly cytotoxic process in the rat mesentery which can be attenuated by MPFF pretreatment. PMID- 10708337 TI - Endothelium-dependent, shear-induced vasodilation is rate-sensitive. AB - OBJECTIVES: To quantify the relative contributions of the rate of change and the magnitude of shear stress to endothelium-mediated arteriolar dilation. METHODS: A feedback control system was designed in which shear stress (tau) and the temporal shear gradient (TSG) were prescribed and dynamically controlled in isolated rat cremaster 1A arterioles. The TSG was the quotient of the maximum shear stress and the ramp duration. This system was used to assess the roles of tau and TSG in the initial, transient vasodilations and the secondary, sustained vasodilations in response to steps and ramps in shear stress. RESULTS: Both step- and ramp-shear experiments revealed time-dependent hiphasic vasodilations that we report for the first time. Application of a step-shear stress of 20 dynes/cm2 elicited an initial transient vasodilation that peaked at about 4 min. When the shear stress was applied as a ramp that reached the maximum value of 20 dynes/cm2 over 5 min, a vasodilation was observed over the ramp period, which reached a peak at the end of the ramp period that was much lower than that observed after step shear. After 20 dynes/cm2 was attained, the vessel diameter decreased despite constant maintenance of the maximum shear stress. In both step- and ramp-shear experiments, after the decrease of the initial vasodilation, a second phase of vasodilation began approximately 15 min after the beginning of the shear application. The second phase of vasodilation reached a steady state that was essentially the same for both the step and the ramp shear. By refining the ramping apparatus further, we varied the TSG up to 40 dynes/cm2 per second and showed that the early vasodilation was highly rate sensitive to TSGs greater than 5 dynes/cm2 per second for a given intermediate value of final shear stress (20 dynes/cm2) and was magnitude sensitive when shear was increased gradually (TSG < 5 dynes/cm2 per second). CONCLUSIONS: Our results suggest that two fundamentally different responses to shear stress are mediated by microvascular endothelium: one vasodilation is elicited by shear stress changes on a time scale of a few seconds or less and another is elicited by shear stress changes on a longer time scale. The former response is potent, transient, and rate sensitive; the latter is more modest, sustained, and magnitude sensitive. PMID- 10708338 TI - Determinants of endothelial cell phenotype in venules. AB - Inflammatory stimuli cause plasma leakage and leukocyte adhesion in venules but not in capillaries or arterioles. The specific response of venules is governed by phenotypic specialization of the venular endothelial cells. What regulates this specialized phenotype? Several recent developments have shed new light on this question and may challenge our thinking about regulation of the venular endothelial cell phenotype. In this review, we consider some of the molecular markers of venular endothelial cells, the hemodynamic and molecular factors that may regulate the phenotype of venular endothelial cells, and abnormalities in endothelial cell phenotype in disease-related angiogenesis and microvascular remodeling. The expanding list of molecular markers may help clarify the physiologic and molecular factors that regulate the phenotype of venular endothelial cells in normal development and disease. PMID- 10708339 TI - Superior temporal gyrus in schizophrenia: a volumetric magnetic resonance imaging study. AB - The left superior temporal gyrus (STG) has been reported to be smaller in patients with schizophrenia. The volume of the STG has been found to correlate negatively with severity of hallucinations and thought disorder. In this study, we measured the STG volume of 20 normal controls and 20 patients with schizophrenia using 3 mm contiguous coronal T1 magnetic resonance images. We found that patients had a significantly smaller left anterior STG, and that the volume of this region negatively correlated with the severity of hallucinations. The left posterior STG was not significantly smaller in patients than in controls, but its volume negatively correlated with severity of thought disorder. We also found that the left anterior STG was smaller than the right STG in patients but not in controls. The STG has at least three histologically distinct areas, each with different connections to the rest of the brain. These data are consistent with the proposition that dysfunction of the primary auditory cortex in the anterior and middle STG and auditory association cortex in the posterior STG may play a role in the production of auditory perceptual abnormalities and poor organization of thought respectively. PMID- 10708340 TI - Increased dendritic MAP2 expression in the hippocampus in schizophrenia. AB - Microtubule associated proteins (MAPs) are central to the development of normal neuronal cytoarchitecture and have been reported to be altered in schizophrenia. In 12 schizophrenic (DSM-III-R criteria) and 12 control hippocampi, we estimated the MAP2 immunoreactive dendritic length using antibodies that recognize total MAP2 (MAP2-T), and a non-phosphorylated form of MAP2 (MAP2-NP). Within the corona ammonis (CA) subregions, and the subiculum, we estimated, for each antibody, the length of the immunoreactive dendritic arborisation using a stereological length estimation technique based on Bouffon's Needle principle and image analysis computer software. Controlling for the confounding effects of age and post-mortem delay, we have found an elevation in overall MAP2-NP immunoreactive dendritic length among schizophrenic subjects in the CA3 (F=5.9, p=0.03), CA2 (F=6.5, p=0.02), CA1 (F=8.3, p=0.01) and subicular (F=9.5, p=0.008) hippocampal subregions. Similar analyses of MAP2-T immunoreactive dendritic length demonstrated significant elevations in the CA1 (F=8.3, p=0.02), CA4 (F=4.9, p=0.04) and subicular (F=7.4, p=0.01) regions. The findings of this quantitative study of increased MAP2 immunoreactive dendritic arborisation in schizophrenia are most likely to reflect either an altered dendritic arborisation or a generalised increase in levels of MAP2 with the hippocampal pyramidal neurons. These findings add to the growing literature indicating the presence of synaptodendritic abnormalities in schizophrenia. PMID- 10708341 TI - Alteration of event related potentials in siblings discordant for schizophrenia. AB - This study was aimed at confirming that auditory event related potential (ERP) abnormalities are indicators of vulnerability to schizophrenia. Auditory ERP performances were assessed at Fz, Cz, and Pz, with an oddball paradigm, in 21 clinically stable patients with schizophrenia, 21 of their healthy biological full siblings and 21 control subjects. The evoked response did not differ between the three groups on N200 waves. Compared to controls, patients with schizophrenia exhibited reduced amplitudes of N100 and P300, and prolonged latency of P300, while their siblings showed prolonged latency of P200 and P300. Among the patients with schizophrenia, ERP abnormalities did not correlate with age, clinical state, duration of illness or antipsychotic treatments. Although other conditions also accounted for alterations of the same type, ERP abnormalities may represent a neurobiological marker of the genetic vulnerability to schizophrenia, independent of phenotypic expression. PMID- 10708342 TI - Cerebrospinal fluid angiotensin-converting enzyme (ACE) correlates with length of illness in schizophrenia. AB - The aim of the study was to evaluate a possible progression with time of cerebrospinal fluid (CSF) angiotensin-converting enzyme (ACE) levels in treated schizophrenia patients. CSF ACE was determined in duplicate by a sensitive inhibitor-binding assay (IBA) from morning CSF samples of 56 acute and chronic in patients with schizophrenic psychoses diagnosed according to DSM-IV. CSF ACE correlated significantly with length of schizophrenic psychosis (r=0.39, p=0.003). There was also a positive significant correlation between CSF ACE and duration of current psychotic episode (r=0.39, p=0.003) as well as duration of current hospitalization (r=0.66, p<0.001). These significances were maintained even when patients who were not treated with antipsychotics at the time of sampling were excluded. The correlations also remained significant when controlling for current neuroleptic dose in chlorpromazine equivalents. Serum ACE did not correlate with any clinical variable. No significant correlations between serum or CSF ACE and age, diagnostic subgroup, gender, serum ACE, CSF to serum albumin ratios, or neuroleptic dose in chlorpromazine equivalents were detected. The elevation of CSF ACE seemed to be confined to a subgroup of chronic patients with few positive symptoms. Elevated CSF ACE may reflect an increased solubilization of ACE from cell membranes in the central nervous system or constitute an increased expression of the ACE gene in response to some stimuli. This may be a function of treatment or a result of the deteriorating schizophrenic process. PMID- 10708343 TI - Efficacy of beta-blocker supplementation for schizophrenia: a systematic review of randomized trials. AB - Adrenergic beta-receptor antagonists, commonly used in the field of cardiovascular diseases, have also been recommended for treatment-resistant schizophrenia. We systematically review quality assessed trials on beta-blocker supplementation of antipsychotic treatment for schizophrenia. All randomized controlled trials comparing any beta-blocking agent added to any antipsychotic with a placebo added to any antipsychotic, and lasting for at least 1 week, were located through electronic searches in all languages of several databases. The trials were assessed by at least two independent reviewers for inclusion, quality score, and data extraction. The reviewers located five studies with 117 participants. The data were poorly presented in these short-term studies and did not evidence any effect of beta-blockers as an adjunct to conventional antipsychotic medication. At present beta-blockers cannot be recommended in the treatment of schizophrenia, and schizophrenia treatment guidelines advocating use of beta-blockers should be revised. PMID- 10708344 TI - Subjective response to antipsychotics in schizophrenic patients: clinical implications and related factors. AB - Awad defined subjective response to medication as the subjective interpretation of the physiological changes that follow its intake. This response is involved in drug compliance and may relate to clinical outcome of the disease. This study examined the variables hypothesized to be related to subjective response to antipsychotics. Sixty schizophrenic in-patients were evaluated with a protocol that examined compliance, hospitalizations, psychopathology, familial and social relationships, degree of autonomy and motivation for life during the year prior to the study. Overall functioning in the previous year was assessed with the Global Assessment Scale (GAS), psychopathology with the Brief Psychopathology Rating Scale (BPRS), insight with the Birchwood scale and side-effects with the Uscandinavian Kociety of Usypharmacology (UKU) side-effects rating scale. Subjective response was evaluated with the Drug Attitude Inventory (DAI-30). The multiple regression analysis revealed that insight and the BPRS paranoid subscale predicted subjective response to antipsychotics (R2=0.31). No relationship was found between subjective response and sociodemographic variables or side-effects. A positive subjective response was related to drug compliance and variables that indicate a more benign clinical course over the previous year. Subjective response to antipsychotics in schizophrenia is related to insight and paranoid ideation. PMID- 10708345 TI - Confusion between silent and overt reading in schizophrenia. AB - The present study was aimed at investigating whether schizophrenic patients are impaired in monitoring their own speech. In particular, we attempted to assess their ability to discriminate between overt and covert speech in a reading task, in order to verify whether they can correctly recollect the modality in which an internally generated action is produced. Subjects were asked to read either silently or aloud, items from a list of words. After a delay of 5 min, they were required to indicate in a new list which words had been read previously (either silently or overtly), or had never been presented during the reading task. With respect to normal controls, schizophrenic patients showed a significant bias to report that they had read aloud words which they had actually read silently, or which were absent during the reading task. The results are discussed in relation to recent neuroimaging studies on inner and overt speech in hallucinating schizophrenic patients. Our data favour the hypothesis that the inability to correctly discriminate between inner and overt speech may play a role in the onset of schizophrenic hallucinations. PMID- 10708346 TI - Lack of effect of an early stressful life event on sensorimotor gating in adult rats. AB - Hypotheses of the etiology of schizophrenia emphasize the important role of perinatal insults in predisposing individuals to the development of the disease, so that an animal model in which a discrete postnatal manipulation of the infant social environment yields schizophrenia-like behavior in adulthood would be valuable in terms of the study of the neural substrate and treatment of schizophrenia. Schizophrenics demonstrate a deficit in sensorimotor gating (prepulse inhibition), and a similar phenomenon has been described in adult rats following the administration of direct and indirect dopamine agonists. Recently it has been reported that a 24 h separation of rat pups from the mother results in a disruption of prepulse inhibition at adulthood. Here we report a study which investigated the same phenomenon but which, in contrast to the previous study, utilized unrelated subjects all derived from different dams. Maternal separation was conducted for 24 h with pups aged 4, 9 or 18 days and these subjects, together with non-separated controls, were tested at age 3 months in terms of their prepulse inhibition in the acoustic startle response paradigm. Maternal separation did not disrupt prepulse inhibition. Comparison of males and females (with a maximum of one opposite-sex sibling) demonstrated that acoustic startle response and prepulse inhibition of this response was enhanced in males relative to females. This study indicates that 24 h maternal separation does not provide a robust model for studying the effects of early environmental insults on the long term abnormal development of sensorimotor gating. PMID- 10708347 TI - Determinants of smoking behaviour in outpatients with schizophrenia. AB - The present study is an assessment of the rate and severity of tobacco consumption in outpatients with schizophrenia, and the determinants of smoking behaviour. Sixty-four patients, assessed by the Item Group Checklist section of the SCAN interview and fitting DSM-IV criteria, were evaluated with CGI and the PANSS scales. In addition, they completed STAI (Spielberger), EPQ (Eysenck), and TPQ (Cloninger) questionnaires. Tobacco dependence was assessed by the Fagerstrom test. One hundred and thirty-seven consecutive outpatients were psychiatric controls. Forty-one out of 64 patients with schizophrenia (64.1%) were current smokers, this rate being significantly higher than in other psychiatric patients and general population. The severity of cigarette consumption in smokers was greater (mean of 22.4 cigarettes/day) than in the general population, but it was not different from that of other psychiatric patients. For patients with schizophrenia, no one variable (except male sex) was different between smokers and non-smokers, but the number of cigarettes/day correlated with state anxiety, trait anxiety, and neuroticism. In the multivariate analysis, the only variable that remained significant was neuroticism. The relationship between clinical features and severity of smoking behaviour may be linked to non-specific variables such as neuroticism and anxiety, but not to psychotic symptoms. PMID- 10708348 TI - Plasma antioxidants and schizophrenia. PMID- 10708349 TI - Schizophrenia associated with psoriasis vulgaris: three case reports. PMID- 10708350 TI - Psychopathological and auditory evoked potential correlates of ketamine psychosis -a single case report. PMID- 10708351 TI - Central nervous system abnormalities in pediatric human immunodeficiency virus infection. AB - With the recent improvements in treatment of HIV, the disease has become a chronic one. The mean survival time of HIV-infected children is now 9-10 years, which is more than 4 times the mean age of such children who died in 1990. Yet, the prevalence of HIV encephalopathy has not decreased despite use of HAART. Rather, it is expected that as patients live longer, the prevalence of CNS manifestations will actually increase. Thus, more children can be expected to manifest encephalopathy, cerebral aneurysms and CNS lymphoma. As AIDS patients live longer, newer and more effective drugs to combat the neurological effects of HIV infection will be required. There is little evidence that HIV damages neurons directly; rather, the damage appears to occur indirectly via viral proteins or neurotoxic factors causing excessive stimulation by excitatory amino acids such as glutamate and quinolinate. This common pathway is similar to that seen in acute neuronal injury and CNS degenerative diseases and may make HIV encephalopathy amenable to pharmacotherapy. The very complexity of HIV CNS entry mechanisms and neuropathogenesis provides a host of sites for potential therapeutic interventions and hope for the future. PMID- 10708352 TI - Syndrome of overdrainage associated with disconnection of a ventriculoperitoneal shunt. AB - We present the case of a child who developed the syndrome of cerebrospinal fluid (CSF) overdrainage with slit-like ventricles on CT in the setting of a disconnected distal shunt valve. Upgrading the shunt alleviated his symptoms. It is suggested that the presence of a patent fibrous tract allowed the overdrainage of CSF. PMID- 10708353 TI - Malignant evolution of choroid plexus papilloma. AB - Choroid plexus tumors are rare CNS neoplasms. The distinction between choroid plexus papilloma (CPP) and choroid plexus carcinoma (CPC) is made on the basis of clinical and histological criteria. Malignant evolution of CPP may occur, and the presence of mitotic figures in CPP may predict the likelihood of recurrence or malignant evolution. Close surveillance is mandated for these patients. We report on two such cases of CPP that transformed to CPC at the time of recurrence. PMID- 10708354 TI - Posterior fossa syndrome: identifiable risk factors and irreversible complications. AB - Cerebellar mutism was first described by Rekate et al. in 1985 as a transient condition which occurs after posterior fossa operations in children. Posterior fossa syndrome (PFS) and cerebellar mutism are often used interchangeably in the literature. In our experience, we found cerebellar mutism to be a reversible component of a persistent neurologic syndrome. The cause and identifiable risk factors have not been clearly elucidated in the literature. To further characterize PFS, we reviewed 253 children with posterior fossa tumors who underwent surgical resection. We documented 20 cases of PFS (8%), 12 males and 8 females. Age ranged from 1.5 to 13 years (mean = 6.5). Of the 20, 16 were medulloblastoma, 3 ependymoma and 1 astrocytoma. There was a 21 % incidence (16/76) of PFS in medulloblastoma of the posterior fossa. The incidence for ependymoma was 13% (3/24) and 1% (1/102) for astrocytoma. All 20 cases (100%) had brainstem involvement by the tumor. The most frequent postoperative findings included mutism, ataxia, 6th and 7th nerve palsies and hemiparesis. Mutism had a latency range of 1-7 days (mean = 1.7) and a duration of 6-365 days (mean = 69.2, median = 35). Although mutism resolved in all cases, the remaining neurologic complications which characterized our findings of PFS were rarely reversible. We describe potential risk factors for developing PFS after surgery with hopes of making neurosurgeons more aware of potential problems following the removal of lesions in this area. Early recognition of PFS would further promote patient and family understanding and coping with this syndrome. PMID- 10708355 TI - Intrauterine myelomeningocele repair reverses preexisting hindbrain herniation. AB - BACKGROUND: It has been reported that intrauterine myelomeningocele repair reduces the amount of hindbrain herniation normally seen in association with the Chiari type II malformation. It is not yet known, however, whether hindbrain herniation is prevented, or whether preexisting herniation is reversed. The following study was designed to elucidate this issue. METHODS: A series of 9 patients underwent intraoperative ultrasound examinations immediately prior to intrauterine myelomeningocele repair. These same patients were then evaluated postnatally using ultrasound and/or MRI. The degree of hindbrain herniation before and after repair was compared using a grading system devised by the authors. RESULTS: Eight patients had clear evidence of moderate to severe hindbrain herniation on intraoperative scans while one was mild. In contrast, on postnatal studies 5 of 9 patients had no evidence of hindbrain herniation, while the other 4 had only mild herniation. CONCLUSION: Intra-uterine myelomeningocele repair appears to reverse preexisting hindbrain herniation. It is postulated that continuous flow of cerebrospinal fluid through the neural placode is the force responsible for inducing migration of the cerebellum and brain stem downward through the foramen magnum. By interrupting that flow during gestation, intrauterine myelomeningocele repair enables the cerebellum and brain stem to resume a normal, or nearly normal, configuration. PMID- 10708356 TI - Management of Chiari I malformations with holocord syringohydromyelia. AB - The management of patients with Type I Chiari malformations (CM 1) with or without syringohydromyelia (SHM) has remained quite controversial, and many different surgical procedures have been advocated. Over the past several years, the authors have treated 7 children presenting with CM 1 and holocord syringohydromyelia with suboccipital decompression and duraplasty alone without intradural procedures. All children received MRI imaging at 2-4 months and 1 year postoperatively. On the early postoperative MRI examination, marked reduction in the syringohydromyelia was seen in 6 children, with minimal change in syrinx size in 1 child who was clinically improving after the operation. At 1 year, all children with early collapse remained collapsed, and the child with minimal early collapse demonstrated an approximately 50% reduction in syrinx size. Clinical follow-up (mean 30 months, range 21-50 months) showed good results in all patients: none of the children have required further neurosurgical intervention, and all have shown improvement in their preoperative function. One child with a 46 degrees scoliosis had a complete collapse of her SHM, but ultimately required spinal fusion. The presenting clinical findings, operative technique, MRI imaging and clinical outcomes will be discussed. The results from these 7 patients with CM 1 and holocord syringomyelia suggest that posterior fossa decompression alone (without intradural procedures) can provide excellent radiographic and clinical outcome. PMID- 10708357 TI - Symptomatic calcified subdural hematomas. AB - Two unique cases of chronic calcified subdural hematomas are reported in children as a long-term complication of a ventriculoperitoneal shunt. Both the patients had undergone shunt procedures in infancy for congenital hydrocephalus. In one patient, the cause of the hydrocephalus was aqueduct stenosis, while in the second patient, a lumbar meningomyelocele was associated with hydrocephalus. In both these patients, a ventriculoperitoneal shunt was done in infancy. In one of them, following the shunt surgery, a bilateral subdural collection was noticed which required burr hole evacuation. Both the patients remained asymptomatic for 9 years, when they presented to our center with acute raised intracranial pressure and contralateral hemiparesis. Both the patients had a relatively short history and had altered sensorium at admission. Surprisingly, in both the patients, the CT scan showed significant mass effect producing calcified subdural hematomas. The shunt systems were found to be working well at surgery. Craniotomy and excision of the calcified subdural hematomas was undertaken. Postoperatively, the patients showed satisfactory recovery, and at discharge the patients were doing well. At the follow-up at the outpatient clinic, the patients were asymptomatic. PMID- 10708358 TI - Multiple recurrent gram-negative cerebrospinal fluid shunt infections associated with a patient with a retained ventricular foreign body. AB - A 3-year-old boy experienced 10 recurrent gram-negative ventriculoperitoneal shunt (VPS) infections with identical strains over a 17-month period. Multiple cranial MRI and CT scans to identify a retained foreign body were negative. CT myelography and pressure infusion radionuclide cisternography were also unhelpful. Ventriculoscopy revealed a small retained foreign body which was successfully removed, and cultures subsequently yielded gram-negative organisms identical to the previously identified bacteria by pulsed field gel electrophoresis. No further infections were noted after removal of the retained fragment. Exploratory ventriculostomy should be considered in patients with recurrent VPS infections where other techniques fail to reveal a cause. PMID- 10708359 TI - Medical management of eosinophilic granuloma of the cervical spine. AB - We report a case of eosinophilic granuloma involving the vertebral bodies of the cervical spine in a 33-month-old girl. This lesion was diagnosed by needle biopsy and treated with prednisone and vinblastine therapy along with immobilization in a Minerva brace. The child has done well over a 9-month follow-up and has shown MRI evidence of resolution of the lesion, reestablishment of structural integrity within the cervical spine and potential reconstitution of the involved vertebral bodies. PMID- 10708360 TI - Ruptured arteriovenous malformation in a boy with Beckwith-Wiedemann syndrome. AB - We report here the first case of ruptured arteriovenous malformation (AVM) in a patient with Beckwith-Wiedemann syndrome (BWS). The subject was a 9-year-old boy exhibiting various abnormal features characteristic of BWS, who had undergone frequent surgical management for conditions such as umbilical hernia, polydactyly, inguinal hernia, cleft palate and lip, and undescended testis. The patient was suffering from ruptured AVM in the right frontal lobe, which was successfully managed with surgical interventions. The possible genetic mechanisms that formed the cerebral vascular malformations in this patient are discussed. PMID- 10708361 TI - Bilateral optic nerve glioma. PMID- 10708362 TI - Presence of the Cpx system in bacteria. PMID- 10708363 TI - Autolysins of Bacillus subtilis: multiple enzymes with multiple functions. PMID- 10708364 TI - Regulation of the transport system for C4-dicarboxylic acids in Bacillus subtilis. AB - Transport systems for C4-dicarboxylates, such as malate, fumarate and succinate, are poorly understood in Gram-positive bacteria. The whole genome sequence of Bacillus subtilis revealed two genes, ydbE and ydbH, whose deduced products are highly homologous to binding proteins and transporters for C4-dicarboxylates in Gram-negative bacteria. Between ydbE and ydbH, genes ydbF and ydbG encoding a sensor-regulator pair, were located. Inactivation of each one of the ydbEFGH genes caused a deficiency in utilization of fumarate or succinate but not of malate. Expression of ydbH, encoding a putative transporter, was stimulated in a minimal salt medium containing 0-05% yeast extract but repressed by the addition of malate to the medium. Inactivation of the putative sensor-regulator pair or solute-binding protein, ydbFG or ydbE, caused complete loss of ydbH expression. The utilization of fumarate and stimulation of ydbH expression resumed in a ydbE null mutant in which ydbFGH were overproduced. Based on these observations, together with analysis of the sequence similarities of the deduced product, we conclude that YdbH is a C4-dicarboxylate-transport protein and its expression is regulated by a C4-dicarboxylate sensor kinase-regulator pair, YdbF and YdbG. Furthermore, it is suggested that YdbE does not directly participate in transport of C4-dicarboxylates, but plays a sensory role in the ydbF-ydbG two-component system, giving rise to specificity or increased efficiency to the system. Deletion analysis of the promoter region of ydbH revealed that a direct repeat sequence was required for the activation of ydbH expression. A catabolite responsive element (CRE) was also found in the -10 region of the promoter, suggesting negative regulation by a CRE-binding protein. PMID- 10708365 TI - S-layer gene sbsC of Bacillus stearothermophilus ATCC 12980: molecular characterization and heterologous expression in Escherichia coli. AB - The cell surface of Bacillus stearothermophilus ATCC 12980 is completely covered with an oblique S-layer lattice. To investigate sequence identities and a common structure-function relationship in S-layer proteins of different B. stearothermophilus wild-type strains, the nucleotide sequence encoding the S layer protein SbsC of B. stearothermophilus ATCC 12980 was determined by PCR techniques. The entire sbsC sequence showed an ORF of 3297 bp predicted to encode a protein of 1099 aa with a theoretical molecular mass of 115409 Da and an isoelectric point of 5.73. Primer extension analysis suggested the existence of two promoter regions. Amino acid sequence comparison between SbsC and SbsA, a previously characterized S-layer protein of B. stearothermophilus PV72/p6 which assembles into a hexagonally ordered lattice, revealed an identical secretion signal peptide, 85% identity for the N-terminal regions (aa 31-270) which do not carry any S-layer homologous motifs, but only 21% identity for the rest of the sequences. Affinity studies demonstrated that the N-terminal part of SbsC is necessary for recognition of a secondary cell wall polymer. This was in accordance with results obtained in a previous study for SbsA, thus confirming a common functional principle for the N-terminal parts of both S-layer proteins. The sbsC coding region cloned into the pET3a vector without its own upstream region, the signal sequence and the 3' transcriptional terminator led to stable expression in Escherichia coli. PMID- 10708366 TI - Vibrio harveyi bioluminescence plays a role in stimulation of DNA repair. AB - Although the genetics and biochemistry of bacterial luminescence have been investigated extensively, the biological role of this phenomenon remains unclear. Here it is shown that luxA, luxB and luxD mutants (unable to emit light) of the marine bacterium Vibrio harveyi are significantly more sensitive to UV irradiation when cultivated in the dark after irradiation than when cultivated under a white fluorescent lamp. This difference was much less pronounced in the wild-type (luminescent) V. harveyi strain. Survival of UV-irradiated Escherichia coli wild-type cells depended on subsequent cultivation conditions (in the dark or in the presence of external light). However, after UV irradiation, the percentage of surviving E. coli cells that bear V. harveyi genes responsible for luminescence was significantly higher than that of non-luminescent E. coli, irrespective of the subsequent cultivation conditions. Moreover, it is demonstrated that luminescence of V. harveyi can be stimulated by UV irradiation even in diluted cultures, under conditions when light emission by these bacteria is normally impaired due to quorum sensing regulation. It is proposed that luminescent bacteria have an internal source of light which could be used in DNA repair by a photoreactivation process. Therefore, production of internal light ensuring effective DNA repair seems to be at least one of the biological functions of bacterial luminescence. PMID- 10708367 TI - InhA, a target of the antituberculous drug isoniazid, is involved in a mycobacterial fatty acid elongation system, FAS-II. AB - Most drug-resistant clinical isolates of the tubercle bacillus are resistant to isoniazid, a first-line antituberculous drug. This antibiotic was shown to act on Mycobacterium tuberculosis by inhibiting a 2-trans-enoyl-acyl carrier protein reductase, called InhA. However, the exact role played by InhA in mycobacteria remained unclear. A mycobacterial enzyme fraction containing InhA was isolated. It displays a long-chain fatty acid elongation activity with the characteristic properties described for the FAS-II (fatty acid synthetase II) system. Inhibition of this activity by InhA inhibitors, namely isoniazid, hexadecynoyl-CoA or octadecynoyl-CoA, showed that InhA belongs to the FAS-II system. Moreover, the InhA inhibitors also blocked the biosynthesis of mycolic acids, which are major lipids of the mycobacterial envelope. The data strongly suggest that isoniazid acts on the mycobacterial cell wall by preventing the FAS-II system from producing long-chain fatty acid precursors for mycolic acid biosynthesis. PMID- 10708368 TI - A new single-copy mycobacterial plasmid, pMF1, from Mycobacterium fortuitum which is compatible with the pAL5000 replicon. AB - A 9.2 kb cryptic Mycobacterium fortuitum plasmid, pMF1, was isolated from strain 110 and its restriction map constructed. A 4.2 kb HindIII fragment of pMF1 was found to support replication in mycobacteria and this fragment was cloned and sequenced to characterize the replication elements of the plasmid. Computer analysis identified a putative Rep protein (362 amino acids) with high homology to the putative Rep protein of the Mycobacterium celatum plasmid pCLP and limited homology, mostly in the N-terminal region, to the Rep proteins of Mycobacterium avium pLR7, M. fortuitum pJAZ38 and Mycobacterium scrofulaceum pMSC262. A region containing a putative ori site was located upstream of the rep gene; this region displayed high homology at the nucleotide level with the predicted ori of pCLP and pJAZ38. A plasmid carrying the 4.2 kb HindIII fragment and a kanamycin resistance marker, designated pBP4, was maintained as a single-copy plasmid in Mycobacterium smegmatis and was stably inherited in the absence of antibiotic selection. Plasmid pBP4 was incompatible with the pJAZ38 replicon but was compatible with the widely used pAL5000 replicon, indicating that among the mycobacterial vectors now available there are two incompatibility groups. Significantly, the plasmid was able to replicate in the pathogen Mycobacterium tuberculosis, making it a useful tool for gene expression studies. To provide a choice of restriction sites and easy manipulation, a 2.1 kb fragment containing the minimal replication region was cloned to make the mycobacterial shuttle vector pBP10, which showed similar stability to pBP4. PMID- 10708369 TI - Analysis of the internal replication region of a mycobacterial linear plasmid. AB - Linear plasmids have previously been identified by the authors in mycobacteria, the telomeres of which have terminal inverted repeats and covalently attached proteins. In this study, the replication of these unusual molecules was investigated by studying a 25 kb linear plasmid from the slow-growing species Mycobacterium celatum called pCLP. An internal region of pCLP responsible for replication in Mycobacterium smegmatis was identified. The nucleotide sequence of the minimum replication region of pCLP, which was 2.8 kb long, contained a putative replication gene, rep, and a putative origin of replication consisting of an 18 bp direct repeat and an AT-rich region. A short section of the pCLP replication region was also found to have sequence identity with the replication regions of mycobacterial circular plasmids, suggesting that these linear and circular plasmids are related. It was found that pCLP replicated in Mycobacterium bovis BCG and was compatible in M. smegmatis with pAL5000- and pJAZ38-derived plasmids from Mycobacterium fortuitum, which belong to two different compatibility groups. Thus, this new Escherichia coli-mycobacteria shuttle vector may be used in both slow- and fast-growing mycobacteria and in co-transformation experiments with other mycobacterial vectors. PMID- 10708370 TI - Cryptosporidium parvum appears to lack a plastid genome. AB - Surprisingly, unlike most Apicomplexa, Cryptosporidium parvum appears to lack a plastid genome. Primers based upon the highly conserved plastid small- or large subunit rRNA (SSU/LSU rRNA) and the tufA-tRNAPhe genes of other members of the phylum Apicomplexa failed to amplify products from intracellular stages of C. parvum, whereas products were obtained from the plastid-containing apicomplexans Eimeria bovis and Toxoplasma gondii, as well as the plants Allium stellatum and Spinacia oleracea. Dot-blot hybridization of sporozoite genomic DNA (gDNA) supported these PCR results. A T. gondii plastid-specific set of probes containing SSU/LSU rRNA and tufA-tRNA(Phe) genes strongly hybridized to gDNA from a diverse group of plastid-containing organisms including three Apicomplexa, two plants, and Euglena gracilis, but not to those without this organelle including C. parvum, three kinetoplastids, the yeast Saccharomyces cerevisiae, mammals and the eubacterium Escherichia coli. Since the origin of the plastid in other apicomplexans is postulated to be the result of a secondary symbiogenesis of either a red or a green alga, the most parsimonious explanation for its absence in C. parvum is that it has been secondarily lost. If confirmed, this would indicate an alternative evolutionary fate for this organelle in one member of the Apicomplexa. It also suggests that unlike the situation with other diseases caused by members of the Apicomplexa, drug development against cryptosporidiosis targeting a plastid genome or metabolic pathways associated with it may not be useful. PMID- 10708371 TI - Distribution of IS1358 and linkage to rfb-related genes in Vibrio anguillarum. AB - The insertion sequence IS1358 is linked to the rfb regions of both Vibrio cholerae O1 and O139, and its location was suggestive of a role in generating new combinations of rfb genes. This provoked an examination of the distribution and localization of IS1358 in Vibrio anguillarum. S11358 was widely distributed in a number of V. anguillarum serogroups. In particular, when cosmid clones of V. anguillarum O1 were screened with IS1358 and subsequently subcloned and sequenced, it was found that rfb-like genes were linked to this region. Furthermore, when the previously identified genes virA and virB from V. anguillarum O1, now known to be involved in LPS biosynthesis, were used as probes, it was discovered that they too are present on the same large EcoRI fragment as IS1358. This clearly indicated that IS1358 was linked to the rfb region of V. anguillarum O1. Further analysis of the location of IS1358 in other serotypes indicated that V. anguillarum O2 also has IS1358 associated with rfb like genes. In V. anguillarum O2 there is more than one copy of IS1358, suggesting that this element is a site for recombination, gene duplication or that it may be capable of transposition. Following this latter premise, IS1358 elements from a variety of V. anguillarum strains have been cloned and sequenced. Only those strains with multiple copies of IS1358 produce a full-length putative transposase, as shown by protein overexpression, further strengthening the argument that the element is transposing within these strains. PMID- 10708372 TI - Multiple paralogous genes related to the Streptomyces coelicolor developmental regulatory gene whiB are present in Streptomyces and other actinomycetes. AB - The whiB sporulation gene of Streptomyces coelicolor was shown [Davis, N. K. & Chater, K. F. (1992). Mol Gen Genet 232, 351-358] to encode a small, cysteine rich putative transcription factor unlike any that had been described previously. The large database of DNA sequences of mycobacteria (like Streptomyces, members of the Actinomycetales) has revealed a family of genes encoding proteins related to WhiB. Mycobacterium tuberculosis contains at least six such genes (whiB homologues in mycobacteria: whmA-F) and a likely seventh, whmG. Using conserved features of Whm proteins, a PCR-based approach led to the discovery that S. coelicolor A3(2) contains several similar genes. Cloning and sequencing of these whiB-like (wbI) genes revealed likely orthologues of four of the whm genes of M. tuberculosis. In all, S. coelicolor contains at least five wbI genes in addition to whiB itself. All five were shown by RT-PCR to be transcribed. A Southern blotting survey using each wbI gene as a probe showed that nearly all of a series of representatives of ten actinomycete genera (including morphologically simple organisms) contain close homologues of several wbI genes, suggesting that the ancient progenitor of all these organisms already contained a family of such genes, which have not been found in any other organisms. PMID- 10708373 TI - A novel valanimycin-resistance determinant (vlmF) from Streptomyces viridifaciens MG456-hF10. AB - A novel valanimycin-resistance determinant (vImF) was isolated from a cosmid containing Streptomyces viridifaciens DNA that leads to valanimycin production in Streptomyces lividans. Expression of the vImF gene in both Escherichia coli and S. lividans provided valanimycin resistance. The nucleotide sequence of vImF consists of 1206 bp and the deduced amino acid sequence encodes a polypeptide with 12 putative transmembrane-spanning segments and a calculated pI of 10.1. VImF shows significant similarities to other known or putative transmembrane efflux proteins that confer antibiotic resistance, but it appears to be specific for valanimycin. The sequence similarities suggest that VImF is a member of the DHA12 family within the major facilitator superfamily of transport proteins and that it is probably involved in active valanimycin efflux energized by a proton dependent electrochemical gradient. PMID- 10708374 TI - Single allele knock-out of Candida albicans CGT1 leads to unexpected resistance to hygromycin B and elevated temperature. AB - Almost all eukaryotic mRNAs are capped at their 5'-terminus. Capping is crucial for stability, processing, nuclear export and efficient translation of mRNA. We studied the phenotypic effects elicited by depleting a Candida albicans strain of mRNA 5'-guanylyltransferase (mRNA capping enzyme; CGT1). Construction of a Cgt1 deficient mutant was achieved by URA-blaster-mediated genetic disruption of one allele of the CGT1 gene, which was localized on chromosome III. The resulting heterozygous mutant exhibited an aberrant colony morphology resembling the 'irregular wrinkle' phenotype typically obtained from a normal C. albicans strain upon mild UV treatment. Its level of CGT1 mRNA was reduced two- to fivefold compared to the parental strain. Proteome analysis revealed a large number of differentially expressed proteins confirming the expected pleiotropic effect of CGT1 disruption. The disrupted strain was significantly more resistant to hygromycin B, an antibiotic which decreases translational fidelity, and showed increased resistance to heat stress. Proteome analysis revealed a 50-fold overexpression of Ef-1alphap and a more than sevenfold overexpression of the cell wall heat-shock protein Ssa2p. Compared to a reference strain, the cgt1/CGT1 heterozygote was equally virulent for mice and guinea pigs when tested in an intravenous infection model of disseminated candidiasis. PMID- 10708375 TI - Very low amounts of glucose cause repression of the stress-responsive gene HSP12 in Saccharomyces cerevisiae. AB - Changing the growth mode of Saccharomyces cerevisiae by adding fermentable amounts of glucose to cells growing on a non-fermentable carbon source leads to rapid repression of general stress-responsive genes like HSP12. Remarkably, glucose repression of HSP12 appeared to occur even at very low glucose concentrations, down to 0.005%. Although these low levels of glucose do not induce fermentative growth, they do act as a growth signal, since upon addition of glucose to a concentration of 0.02%, growth rate increased and ribosomal protein gene transcription was up-regulated. In an attempt to elucidate how this type of glucose signalling may operate, several signalling mutants were examined. Consistent with the low amounts of glucose that elicit HSP12 repression, neither the main glucose-repression pathway nor cAMP-dependent activation of protein kinase A appeared to play a role in this regulation. Using mutants involved in glucose metabolism, evidence was obtained suggesting that glucose 6-phosphate serves as a signalling molecule. To identify the target for glucose repression on the promoter of the HSP12 gene, a promoter deletion series was used. The major transcription factors governing (stress-induced) transcriptional activation of HSP12 are Msn2p and Msn4p, binding to the general stress-responsive promoter elements (STREs). Surprisingly, glucose repression of HSP12 appeared to be independent of Msn2/4p: HSP12 transcription in glycerol-grown cells was unaffected in a deltamsn2deltamsn4 strain. Nevertheless, evidence was obtained that STRE-mediated transcription is the target of repression by low amounts of glucose. These data suggest that an as yet unidentified factor is involved in STRE-mediated transcriptional regulation of HSP12. PMID- 10708376 TI - Long-chain alkyl ester of AMP acts as an antagonist of glucose-induced signal transduction that mediates activation of plasma membrane proton pump in Saccharomyces cerevisiae. AB - One of the long-chain alkyl esters of AMP, adenosine 5'-hexadecylphosphate (AMPC16), exhibited a cytotoxic growth inhibitory effect on cells of various yeast strains. The growth inhibitory effect of AMPC16 on Saccharomyces cerevisiae cells was observed only in medium containing Mg2+, which accelerated cellular uptake of the nucleotide analogue. In the presence of Mg2+, AMPC16 completely inhibited glucose-induced extracellular acidification by the intact cells and also interfered with activation of the plasma membrane ATPase, but did not directly inhibit the ATPase activity itself. AMPC16 treatment prevented cells from increasing their intracellular sn-1,2-diacylglycerol (DAG) level in response to glucose, whereas the inhibition of proton extrusion by the cells could be largely reversed by the coaddition of a membrane-permeable DAG analogue. The DAG analogue, a physiological activator of protein kinase C (PKC), was not protective against the inhibition of glucose-induced proton extrusion by staurosporine, which is capable of directly interfering with the action of PKC. These results implied that AMPC16 caused a Mg(2+)-dependent cytotoxic effect on Sac. cerevisiae cells by interfering with a phosphatidylinositol type of signal that mediates activation of the plasma membrane proton pump. PMID- 10708377 TI - The yeast Chs4 protein stimulates the trypsin-sensitive activity of chitin synthase 3 through an apparent protein-protein interaction. AB - Inducible overexpression of the CHS4 gene under the control of the GAL1 promoter increased Chs3p (chitin synthase 3) activity in Saccharomyces cerevisiae several fold. Approximately half of the Chs3p activity in the membranes of cells overexpressing Chs4p was extracted using CHAPS and cholesteryl hemisuccinate. The detergent-extractable Chs3p activity appeared to be non-zymogenic because incubation with trypsin decreased enzyme activity in both the presence and absence of the substrate, UDP-N-acetylglucosamine. Western blotting confirmed that Chs3p was extracted from membranes by CHAPS and cholesteryl hemisuccinate and revealed that Chs4p was also solubilized using these detergents. Yeast two hybrid analysis with truncated Chs4p demonstrated that the region of Chs4p between amino acids 269 and 563 is indispensable not only for eliciting the non zymogenic activity of Chs3p but also for binding of Chs4p to Chs3p. Neither the EF-hand motif nor a possible prenylation site in Chs4p was required for these activities. Thus, it was demonstrated that stimulation of non-zymogenic Chs3p activity by Chs4p requires the amino acid region from 269 to 563 of Chs4p, and it seems that Chs4p activates Chs3p through protein-protein interaction. PMID- 10708378 TI - Identification and characterization of anaerobic gut fungi using molecular methodologies based on ribosomal ITS1 and 185 rRNA. AB - The gut fungi are an unusual group of zoosporic fungi occupying a unique ecological niche, the anaerobic environment of the rumen. They exhibit two basic forms, with nuclear migration throughout the hyphal mass for polycentric species and with concentration of nuclear material in a zoosporangium for monocentric species. Differentiation between isolates of these fungi is difficult using conventional techniques. In this study, DNA-based methodologies were used to examine the relationships within and between two genera of monocentric gut fungi gathered from various geographical locations and host animals. The ribosomal ITS1 sequence from 16 mono- and 4 polycentric isolates was PCR-amplified and sequenced; the sequences obtained were aligned with published sequences and phylogenetic analyses were performed. These analyses clearly differentiate between the two genera and reflect the previously published physiological conclusions that Neocallimastix spp. constitute a more closely related genus than the relatively divergent genus Piromyces. The analyses place two type species N. frontalis and N. hurleyensis together but, contrary to a recent suggestion in the literature, place them apart from the other agreed species N. patriciarum. In situ hybridization and slot-blotting were investigated as potential methods for detection of and differentiation between monocentric gut fungi. DNA slot-blot analysis using ribosomal sequences is able to differentiate between gut fungal genera and thus has considerable potential for use in ecological studies of these organisms. PMID- 10708379 TI - Degradation of pentachlorophenol by Phanerochaete chrysosporium: intermediates and reactions involved. AB - Under nitrogen-limiting, secondary metabolic conditions, the lignin-degrading basidiomycete Phanerochaete chrysosporium rapidly degrades pentachlorophenol. The pathway for the degradation of pentachlorophenol has been elucidated by the characterization of fungal metabolites and oxidation products generated by purified lignin peroxidase (LiP) and manganese peroxidase (MnP). The multi-step pathway is initiated by a LiP- or MnP-catalysed oxidative dechlorination reaction to produce tetrachloro-1,4-benzoquinone. Under primary or secondary metabolic conditions, the quinone is further degraded by two parallel pathways with cross links. The quinone is reduced to tetrachlorodihydroxybenzene, which can undergo four successive reductive dechlorinations to produce 1,4-hydroquinone, and the latter is o-hydroxylated to form the final aromatic metabolite, 1,2,4 trihydroxybenzene. Alternatively, the tetrachloro-1,4-benzoquinone is converted, either enzymically or nonenzymically, to 2,3,5-trichlorotrihydroxybenzene, which undergoes successive reductive dechlorinations to produce 1,2,4 trihydroxybenzene. Finally, at several points, hydroxylation reactions convert chlorinated dihydroxybenzenes to chlorinated trihydroxybenzenes, linking the two pathways at each of these steps. Presumably, the 1,2,4-trihydroxybenzene produced in each pathway is ring-cleaved with subsequent degradation to CO2. In contrast to the oxidative dechlorination step, the reductive dechlorinations and hydroxylations occur during both primary and secondary metabolic growth. Apparently, all five chlorine atoms are removed from the substrate prior to ring cleavage. PMID- 10708380 TI - Glucoamylase::green fluorescent protein fusions to monitor protein secretion in Aspergillus niger. AB - A glucoamylase::green fluorescent protein fusion (GLA::sGFP) was constructed which allows the green fluorescent protein to be used as an in vivo reporter of protein secretion in Aspergillus niger. Two secretory fusions were designed for secretion of GLA::sGFP which employed slightly different lengths of the glucoamylase protein (GLA499 and GLA514). Expression of GLA::sGFP revealed that fluorescence was localized in the hyphal cell walls and septa, and that fluorescence was most intense at hyphal apices. Extracellular GLA::sGFP was detectable by Western blotting only in the supernatant of young cultures grown in soya milk medium. In older cultures, acidification of the medium and induction of proteases were probably responsible for the loss of extracellular and cell wall fluorescence and the inability to detect GLA::sGFP by Western analysis. A strain containing the GLA::sGFP construct was subjected to UV mutagenesis and survivors screened for mutations in the general secretory pathway. Three mutants were isolated that were unable to form a halo on either starch or gelatin medium. All three mutants grew poorly compared to the parental strain. Fluorescence microscopy revealed that for two of the mutants, GLA::sGFP accumulated intracellularly with no evidence of wall fluorescence, whereas for the third mutant, wall fluorescence was observed with no evidence of intracellular accumulation. These results indicate that the GLA::sGFP fusion constructs can be used as convenient fluorescent markers to study the dynamics of protein secretion in vivo and as a tool in the isolation of mutants in the general secretory pathway. PMID- 10708381 TI - Role of K+ and amino acids in osmoregulation by the free-living microaerophilic protozoon Hexamita inflata. AB - The primitive free-living protozoon Hexamita inflata was found to maintain a cell volume of approximately 260 fI under standard culture conditions. On increasing the extracellular osmolality the volume decreased and the cells remained shrunken for >30 min. By contrast, a decrease in the external osmolality resulted in a transient increase in cell volume which was followed by an efficient 'regulatory volume decrease' (RVD). H. inflata contains high concentrations of amino acids, with alanine constituting over 70% of the total amino acid pool. Exposure to hypo osmotic medium resulted in the loss from the cell of both amino acids and K+, via one or more swelling-activated pathways. The efflux of amino acids and K+, together with a charge-balancing counter-anion, accounted almost fully for the observed RVD. The pharmacological properties of the swelling-activated pathways differ from those of volume-sensitive transporters and channels described previously in other cell types. PMID- 10708382 TI - Molecular characterization of the lactococcal plasmid pCIS3: natural stacking of specificity subunits of a type I restriction/modification system in a single lactococcal strain. AB - A 6.1 kb plasmid from the Lactococcus lactis subsp. cremoris strain UC509.9, named pCIS3, was found to mediate a restriction/modification (R/M) phenotype. Nucleotide sequence analysis of pCIS3 revealed the presence of an hsdS gene, typical of type I R/M systems. The presence of this plasmid resulted in a 10(4) fold reduction in the efficiency of plating (e.o.p.) of unmodified phage. In addition to the hsdS gene of pCIS3, two more hsdS genes were identified in strain UC509.9, one located on the chromosome downstream of a gene highly homologous to hsdM genes and a third on the smallest (4 kb) plasmid, named pCIS1. The replication region of pCIS3 was highly similar to that of a large family of lactococcal theta replicons. In addition, pCIS3 was found to encode a member of the CorA family of magnesium transporters. PMID- 10708383 TI - Microbiological and molecular impacts of AbiK on the lytic cycle of Lactococcus lactis phages of the 936 and P335 species. AB - The lactococcal abortive infection mechanism AbiK was previously shown to be highly effective against the small isometric-headed bacteriophage ul36 of the P335 species, as evidenced by an efficiency of plaquing (e.o.p.) of 10(-6), a 14 fold reduction in the burst size and an efficiency at which centres of infection form (e.c.o.i.) of 0.5%. No phage DNA was detected in the infected AbiK+ cells [Emond, E., Holler, B. J., Boucher, I., Vandenbergh, P. A., Vedamuthu, E. R., Kondo, J. K. & Moineau, S. (1997). Appl Environ Microbiol 63, 1274-1283]. Here, the effects of AbiK are compared on the small isometric-headed phages p2 and P008 (936 species) and on the phage P335 (P335 species). The microbiological impacts of AbiK on p2 were relatively similar to those reported for ul36, with an e.o.p. of 10(6), an 11-fold reduction in the burst size and an e.c.o.i. of 5%. Contrary to phage ul36, replication of phage p2 DNA was observed in the AbiK+ cells. Only immature forms (concatemeric and circular DNA) of phage p2 DNA were found, indicating that the presence of AbiK prevented phage DNA maturation. These distinct molecular consequences of AbiK were also observed for phages P335 and P008, two phages that propagate on the same host. To the knowledge of the authors, this is the first time that different phage responses towards an Abi system have been reported. PMID- 10708384 TI - Genes for the synthesis of the osmoprotectant glycine betaine from choline in the moderately halophilic bacterium Halomonas elongata DSM 3043, USA. AB - The genes involved in the oxidative pathway of choline to glycine betaine in the moderate halophile Halomonas elongata DSM 3043 were isolated by functional complementation of an Escherichia coli strain defective in glycine betaine synthesis. The cloned region was able to mediate the oxidation of choline to glycine betaine in E. coli, but not the transport of choline, indicating that the gene(s) involved in choline transport are not clustered with the glycine betaine synthesis genes. Nucleotide sequence analysis of a 4.6 kb segment from the cloned DNA revealed the occurrence of three ORFs (betIBA) apparently arranged in an operon. The deduced betI gene product exhibited features typical for DNA-binding regulatory proteins. The deduced BetB and BetA proteins showed significant similarity to soluble glycine betaine aldehyde dehydrogenases and membrane-bound choline dehydrogenases, respectively, from a variety of organisms. Evidence is presented that BetA is able to oxidize both choline and glycine betaine aldehyde and therefore can mediate both steps in the synthesis of glycine betaine. PMID- 10708385 TI - Catalase deficiency in Staphylococcus aureus subsp. anaerobius is associated with natural loss-of-function mutations within the structural gene. AB - Degenerate oligonucleotide primers based on internal peptide sequences obtained by HPLC from purified Staphylococcus aureus catalase were used to locate the S. aureus and S. aureus subsp. anaerobius kat regions by PCR. Southern hybridization analysis with a probe derived from a 1.1 kb PCR-amplified fragment showed that a single copy of the putative catalase gene was present in the S. aureus and S. aureus subsp. anaerobius chromosome. The nucleotide sequence of S. aureus katA revealed a 1518 bp open reading frame for a protein with 505 amino acids and a predicted molecular mass of 58347 Da, whereas S. aureus subsp. anaerobius katB is 1368 nt long and encodes a polypeptide of 455 amino acids with a predicted molecular mass of 52 584 Da. These catalases are highly homologous to typical monofunctional catalases from prokaryotes. The active-site residues, proximal and distal haem-binding ligands and NADPH-binding residues of the bovine liver catalase-type enzyme were highly conserved in S. aureus KatA. Escherichia coli cells carrying cloned katA had a catalase activity approximately 1000 times that of untransformed E. coli, but no detectable increase in catalase activity was observed with E. coli carrying cloned katB. Northern blotting showed the presence of a kat-specific transcript in S. aureus subsp. anaerobius, suggesting that the lack of catalase activity in this bacterium is due to a post-transcriptional alteration. Compared to the nucleotide sequence of katA, katB showed a single base-pair deletion and six mis-sense mutations, and these alterations were present in three other S. aureus subsp. anaerobius strains analysed. The deletion, located at 1338 bp from the initiation codon, originates a shift of the nucleotide reading frame and is responsible for the premature translation termination at 1368 bp, generating a KatB polypeptide 50 amino acid residues shorter than KatA. Moreover, four of the mis-sense mutations present in katB lead to non-conservative amino acid replacements, the most significant being that located at residue 317 (Pro in KatA-->Ser in KatB) because the affected amino acid is involved in determining the proximal haem-binding site. Both the main alterations found in KatB (the deletion and the substitution in residue 317) seem to contribute to the lack of catalase activity in S. aureus subsp. anaerobius, as deduced from results obtained with chimeric catalase constructs. PMID- 10708386 TI - Genomic and antigenic differences between the European and African/Australian clusters of Mycoplasma mycoides subsp. mycoides SC. AB - Mycoplasma mycoides subsp. mycoides small-colony type (SC), the aetiological agent of contagious bovine pleuropneumonia (CBPP), can be grouped into two major, epidemiologically distinct, clusters. One cluster contains strains isolated from different European countries since 1980 and a second cluster contains African and Australian strains collected over the last 50 years. Genetic analysis of representative strains from the two clusters revealed a genomic segment of 8.84 kb, located close to a copy of IS1296, which is present in all strains of the African cluster but lacking in all strains of the European cluster. This segment contains a copy of IS1634, a gene for a potential lipoprotein, IppB, open reading frames encoding a putative surface-located membrane protein and a hypothetical proline-rich membrane protein, and two open reading frames showing similarity to putative ABC transporters. The product of the IppB gene, lipoprotein B (LppB), has an apparent molecular mass of 70 kDa and was shown to be surface located. It is detected with monospecific antibodies in all strains of the African cluster tested, but not in European-cluster strains. DNA sequence analysis of the splicing site at which European strains differ from African-cluster strains by the lack of the 8.84 kb segment showed that the European cluster has arisen by deletion from a strain of the African cluster. Hence, M. mycoides subsp. mycoides SC strains isolated in different European countries from the newly reemerging outbreaks of CBPP, which occurred after the eradication of the epizootic in Europe in the middle of the 20th century, represent a phylogenetically newer cluster that has been derived from a strain of the older cluster of M. mycoides subsp. mycoides SC which is still endemic on the African continent. PMID- 10708387 TI - The genes for erythritol catabolism are organized as an inducible operon in Brucella abortus. AB - Erythritol utilization is a characteristic of pathogenic Brucella abortus strains. The attenuated vaccine strain B19 is the only Brucella strain that is inhibited by erythritol, so a role for erythritol metabolism in virulence is suspected. A chromosomal fragment from the pathogenic strain B. abortus 2308 containing genes for the utilization of erythritol was cloned taking advantage of an erythritol-sensitive Tn5 insertion mutant. The nucleotide sequence of the complete 7714 bp fragment was determined. Four ORFs were identified in the sequence. The four genes were closely spaced, suggesting that they were organized as a single operon (the ery operon). The first gene (eryA) encoded a 519 aa putative erythritol kinase. The second gene (eryB) encoded an erythritol phosphate dehydrogenase. The function of the third gene (eryC) product was tentatively assigned as D-erythrulose-1-phosphate dehydrogenase and the fourth gene (eryD) encoded a regulator of ery operon expression. The operon promoter was located 5' to eryA, and contained an IHF (integration host factor) binding site. Transcription from this promoter was repressed by EryD, and stimulated by erythritol. Functional IHF was required for expression of the operon in Escherichia coli, suggesting a role for IHF in its regulation in B. abortus. The results obtained will be helpful in clarifying the role of erythritol metabolism in the virulence of Brucella spp. PMID- 10708388 TI - A flagellar gene cluster from the oral spirochaete Treponema maltophilum. AB - A flagellar gene cluster from the oral spirochaete Treponema maltophilum ATCC 51939T was cloned. Sequence analysis revealed six putative ORFs, two of which encode the flagellar subunit proteins FlaB2 (286 aa) and FlaB3 (285 aa). Northern blot analysis revealed two flagellin transcripts with the expected size of monocistronic mRNAs. Sequence analysis and primer extension experiments indicated that the transcription of the flaB2 gene is directed by a sigma28-like FliA factor. Using fliA and fliA+ Escherichia coli K-12 strains, it was shown that flaB2 expression in E. coli required the sigma28 factor using an initiation site identical to that in Treponema maltophilum. Primer extension analysis revealed two transcriptional start sites 5' of the flaB3 gene, a strong promoter with a sigma28-like -10 promoter element and a weak promoter with a putative sigma54 promoter consensus sequence. Downstream of flaB3, a putative fliD homologue was found, probably encoding the flagellar cap protein of Treponema maltophilum. Flagellin-gene-specific DNA probes hybridized to all 13 Treponema strains investigated, whereas a fliD-specific DNA probe only hybridized to Treponema maltophilum, other treponemal group IV isolates and Treponema brennaborense. PMID- 10708389 TI - Transcriptional analysis of the nirS gene, encoding cytochrome cd1 nitrite reductase, of Paracoccus pantotrophus LMD 92.63. AB - The gene for cytochrome cd1 nitrite reductase of Paracoccus pantotrophus, a protein of known crystal structure, is nirS. This gene is shown to be flanked by genes previously recognized in other organisms to encode proteins involved in the control of its transcription (nirI) and the biosynthesis of the d1 cofactor (nirE). Northern blot analysis has established under anaerobic conditions that a monocistronic transcript is produced from nirS, in contrast to observations with other denitrifying bacteria in which arrangement of flanking genes is different and the messages produced are polycistronic. The lack of a transcript under aerobic conditions argues against a role for cytochrome cd1 in the previously proposed aerobic denitrification pathway in Pa. pantotrophus. A putative rho independent transcription termination sequence immediately following nirS, and preceding nirE, can be identified. The independent transcription of nirS and nirE indicates that it should be possible to produce site-directed mutants of nirS borne on a plasmid in a nirS deletion mutant. The transcript start point for nirS has been determined by two complementary techniques, 5'-RACE (Rapid amplification of cDNA 5' ends) and primer extension. It is 29 bp upstream of the AUG of nirS. An anaerobox, which presumably binds Nnr, is centred a further 41.5 bp upstream of the transcript start. No standard sigma70 DNA sequence motifs can be identified, but a conserved sequence (T-T-GIC-C-G/C-G/C) can be found in approximately the same position (-16) upstream of the transcript starts of nirS and nirI, whose products are both involved in the conversion of nitrite to nitric oxide. PMID- 10708390 TI - Glutamate residues in the putative transmembrane region are required for the function of the VirS sensor histidine kinase from Clostridium perfringens. AB - The causative agent of gas gangrene, Clostridium perfringens, is a Gram-positive anaerobe which produces a number of extracellular toxins and enzymes. The production of several of these toxins is regulated by the VirS/VirR two-component signal transduction system. The sensor histidine kinase, VirS, contains motifs that are conserved amongst sensor histidine kinases, although not in the same relative positions. In this study, the conserved histidine residue (H255), the GXGL and DXGXG motifs, and two glutamate residues located in putative transmembrane domains were altered by site-directed mutagenesis to examine their significance for VirS function. Introduction of the mutated virS genes into the virS::Tn916 mutant, JIR4000, showed that the altered virS genes were not able to complement the host mutation. These results demonstrate that the conserved motifs, including the cytoplasmic DXGXG motif which is located between the putative transmembrane domains 4 and 5, are functional. Furthermore, it is concluded that charged residues located within two of these transmembrane domains are also required for the structural or functional integrity of the VirS sensor kinase. PMID- 10708391 TI - Oxidase and periplasmic cytochrome assembly in Escherichia coli K-12: CydDC and CcmAB are not required for haem-membrane association. AB - The mechanism(s) that bacteria use to transport haem into and across the cytoplasmic membrane to complete the assembly of periplasmic cytochromes is unknown. The authors have tested directly the role(s) of two ATP-binding cassette (ABC) transporters - the cydDC and ccmAB gene products - in Escherichia coli by measuring haem uptake in everted (inside-out) membrane vesicles. If haem is exported to the periplasm in vivo, the same process should result in active accumulation in such everted vesicles. [14C]Haemin (chloride) with bovine serum albumin (BSA) as a carrier protein was accumulated in intact everted membrane vesicles by an energy-independent mechanism. The kinetics of this process were biphasic: rapid uptake/binding was followed by a slower uptake of haem, which was inhibited by a large excess of unlabelled haemin-BSA, but not by BSA. However, accumulated haemin was not chased out of the vesicles by unlabelled haemin-BSA, suggesting specific binding of haemin with the membrane or transport into the lumen of the vesicle. Neither ATP nor a protonmotive force (delta(p)) generated by lactate oxidation was required for haemin binding or subsequent transport, and carbonyl cyanide m-chlorophenylhydrazone (CCCP), sodium vanadate and monensin had no effect on haemin transport. The rate of haemin uptake following the initial rapid binding was proportional to the external haemin concentration, suggesting that the uptake process was driven by the haemin concentration gradient across the cell membrane. The kinetics of [14C]haemin uptake were similar in wild-type and cydD1 or delta(ccmA) mutants, suggesting that the activity of neither the CydDC nor CcmAB transporters is essential for haem export to the periplasm. Cytochrome d levels were unaffected by mutations in trxB (encoding thioredoxin reductase), trxA (thioredoxin), or grx (glutaredoxin), suggesting that the CydDC transporter does not export these components of reducing pathways for cytochrome assembly. PMID- 10708392 TI - Regulation of production of the antifungal metabolite 2,4-diacetylphloroglucinol in Pseudomonas fluorescens F113: genetic analysis of phlF as a transcriptional repressor. AB - The antifungal metabolite 2,4-diacetylphloroglucinol plays a major role in the biocontrol capabilities of Pseudomonas fluorescens. The phloroglucinol biosynthetic locus of P. fluorescens F113 has been isolated previously. From nucleotide sequence data, a putative regulator gene (phlF) was identified upstream and divergently transcribed from the phlACBD phloroglucinol biosynthetic genes. PhlF shows similarity to various transcriptional repressors in the EMBL database and exhibits a helix-turn-helix motif in its amino acid sequence. phlF was cloned into an expression vector and the PhlF protein product was purified. Gel retardation experiments demonstrated PhlF to be a DNA-binding protein and showed that it binds to the phlA-phlF intergenic region. Introduction of phlF into P. fluorescens F113 in multiple copies resulted in repression of phloroglucinol production in this strain. This effect was mediated at the transcription level since the expression of a phloroglucinol biosynthetic gene fusion in this background was equally repressed. Furthermore, the inactivation of phlF results in derepression of phloroglucinol production in this strain. PMID- 10708393 TI - Quantitation of low level unconjugated polysaccharide in tetanus toxoid-conjugate vaccine by HPAEC/PAD following rapid separation by deoxycholate/HCl. AB - A simple and rapid acid precipitation method has been applied successfully for separating free capsular polysaccharide of Haemophilus influenzae type b (polyribosyl ribitol phosphate, PRP) from PRP tetanus toxoid conjugate (PRP-T) in a final dosage amount of low-level materials. The unconjugated PRP was found to stay in the supernatant without precipitation, while conjugated PRP-T was fully precipitated. High performance anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD) has been applied for analysis of the PRP content in the supernatant after the separation. This method requires minimum sample handling and is specific, sensitive and reproducible making it suitable for release and stability testing of PRP-T in final containers. PMID- 10708394 TI - Mechanisms of metal-catalyzed oxidation of histidine to 2-oxo-histidine in peptides and proteins. AB - The metal-catalyzed oxidation of histidine (His) to 2-oxo-histidine (2-oxo-His) represents an important pathway of protein oxidation in vivo and in vitro. In the pharmaceutical literature this pathway has received less attention. However, this fact may not necessarily represent reality as, in some cases, the analysis of His oxidation in proteins may be compromised by aggregation and precipitation of the target protein. For predicting the susceptibility of His residues in proteins it is important to understand in detail how protein sequence and conformation control the mechanisms of His oxidation to 2-oxo-His, reviewed in this paper. PMID- 10708395 TI - Validation of a peptide mapping method for a therapeutic monoclonal antibody: what could we possibly learn about a method we have run 100 times? AB - Peptide mapping is a key analytical method for studying the primary structure of proteins. The sensitivity of the peptide map to even the smallest change in the covalent structure of the protein makes it a valuable 'finger-print' for identity testing and process monitoring. We recently conducted a full method validation study of an optimised reverse-phase high-performance liquid chromatography (RP HPLC) tryptic map of a therapeutic anti-CD4 IgG1 monoclonal antibody. We have used this method routinely for over 1 year to support bioprocess development and test production lots for clinical trials. Herein we summarize the precision and ruggedness of the testing procedure and the main findings with respect to 'coverage of amino acid sequence' and limits-of-detection for various hypothetical structural variants. We also describe, in more detail, two unanticipated insights into the method gained from the validation study. The first of these is a potentially troublesome side-product arising during the reduction/alkylation step. Once the cause of this side-product was identified, it was easily prevented. We also report on subtle changes to the peptide map upon extended storage of the digest in the autosampler. These findings helped us to develop a 'robust' method for implementation in a quality control laboratory. PMID- 10708397 TI - Simultaneous determination of atenolol and amlodipine in tablets by high performance thin-layer chromatography. AB - A new simple, precise, rapid and selective high-performance thin-layer chromatographic (HPTLC) method has been developed for the simultaneous determination of atenolol (ATL) and amlodipine (AMLO) in tablets, using methylene chloride:methanol:ammonia solution (25% NH3) (8.8:1.3:0.1; v/v) as the mobile phase and Merck HPTLC plates (0.2 mm thickness) precoated with 60F254 silica gel on aluminium sheet as the stationary phase. Detection was carried out densitometrically using a UV detector at 230 nm. The retention factors of ATL and AMLO were 0.33 and 0.75, respectively. Calibration curves were linear in the range 10-500 microg ml(-1) for both. Assays of ATL and AMLO were 49.87 mg per tablet (relative standard deviation (R.S.D.), 1.3%) and 4.90 mg per tablet (R.S.D., 1.38%) for brand I, and 49.27 mg per tablet (R.S.D., 1.12%) and 4.98 mg per tablet (R.S.D., 1.42%) for brand II, respectively. The percentage recoveries for ATL and AMLO for brands I and II were 99.06 and 99.30%, and 99.27 and 99.15%, respectively. PMID- 10708396 TI - Isoaspartate in peptides and proteins: formation, significance, and analysis. AB - Formation of isoaspartyl peptide bonds (isoAsp) is one of the most common forms of non-enzymatic degradation of peptides and proteins under mild conditions. IsoAsp arises when certain Asn-Xaa and Asp-Xaa sites undergo a spontaneous intramolecular rearrangement to form a succinimide which subsequently hydrolyzes to generate a mixture of isoAsp-Xaa and Asp-Xaa linkages in a ratio of approximately 2:1. This pathway is responsible for the much greater susceptibility of asparagine, compared with glutamine, to deamidation at neutral and alkaline pH. Rearrangement occurs most readily at Asn-Gly, Asn-Ser, and Asp Gly sequences where the local polypeptide chain flexibility is high. Formation of isoAsp can decrease the biological activity of a protein pharmaceutical, alter its susceptibility to proteolytic degradation, and elicit autoimmunity. The enzyme protein L-isoaspartyl methyltransferase can be used to measure isoAsp sites in the low pmol range with or without the use of radioisotopes. PMID- 10708398 TI - Direct thin layer chromatography enantioresolution of some basic DL-amino acids using a pharmaceutical industry waste as chiral impregnating reagent. AB - Direct enantiomeric resolution of DL-arginine, DL-histidine, DL-lysine, DL-valine and DL-leucine into their enantiomers was achieved by thin layer chromatography (TLC) on silica gel plates impregnated with optically pure (1R, 3R, 5R)-2 azabicyclo[3,3,0]octan-3-carbo-xylic acid (0.011 M) as a chiral selector which is a waste of a pharmaceutical industry. Different combinations of acetonitrile methanol-water were found to be successful in resolving the DL-amino acids. The spots were detected by ninhydrin (0.2% in acetone) and the detection limit was 0.66 microg. PMID- 10708399 TI - A validated quantitative colorimetric assay for gentamicin. AB - A colorimetric procedure was developed for the quantification of gentamicin. The method was based on the ninhydrin reaction with primary and secondary amines present in the gentamicin. This reaction produces a purple colour. The effects of several factors including pH, ninhydrin concentration and reaction time were investigated to optimize the assay method. Using the assay protocol, the absorption of the gentamicin-ninhydrin mixtures at 400 nm had a linear relationship with the gentamicin concentration ranging from 30 to 120 microg/ml. The colorimetric gentamicin assay reported herein is of great practical value because it is reproducible, sensitive, simple and extremely inexpensive. PMID- 10708400 TI - Determination of possible impurities in piracetam using FTIR spectroscopy. AB - Piracetam (2-oxo-1-pyrrolidine acetamide (PAm)) may contain some structurally related impurities deriving from synthesis or degradation, mainly 2-oxo-1 pyrrolidine acetic acid (PAc). A rapid and sensitive Fourier transform infrared (FTIR) spectrophotometric method was developed for determination of possible impurities in piracetam. The results showed that the method was effective for the simultaneous determination of PAc in piracetam by FTIR spectrophotometry and may be a real alternative to HPLC. The apparent spectral resolution was first enhanced by using the Fourier self-deconvolution (FSD) method and the profiles were then fully deconvoluted by using a curve fitting procedure. FSD method followed by curve fitting allowed to evaluate quantitatively the areas under the pyrrolidine acetic acid peaks. Mixtures of known composition were used as standards to minimise errors due to the presence of both compounds in the same mixture. The detection limit of pyrrolidine acetic acid was estimated to be 0.1% with respect to piracetam. PMID- 10708401 TI - Multiwavelength spectrophotometric determination of acid dissociation constants part V: microconstants and tautomeric ratios of diprotic amphoteric drugs. AB - The acid-base equilibria of several diprotic amphoteric drugs, namely, niflumic acid, norfloxacin, piroxicam, pyridoxine and 2-methyl-4-oxo-3H-quinazoline-3 acetic acid have been characterized in terms of microconstants and tautomeric ratios. A multiwavelength spectrophotometric (WApH) titration method for determination of acid dissociation constants (pKa values) of ionizable compounds developed previously was applied for this purpose. Microspeciation was investigated by three approaches: (1) selective monitoring of ionizable group by spectrophotometry, (2) deductive method and (3) k(z) method for determination of tautomeric ratio from co-solvent mixtures. The formulation for (3) has been derived and found to invoke fewer assumptions than a reported procedure (K. Takacs-Novak, A. Avdeef, K.J Box, B. Podanyi, G. Szasz, J. Pharm. Biomed. Anal., 12 (1994) 1369-1377). It has been shown that the WApH technique, for such types of ampholytes, is able to deduce the microconstants and tautomeric ratios which are in good agreement with literature data. PMID- 10708402 TI - Determination of ipratropium bromide in vials using kinetic and first-derivative spectrophotometric methods. AB - Two sensitive and accurate spectrophotometric methods are presented for the determination of ipratropium bromide (IPB). The first method, kinetic method, is based on the alkaline oxidation of IPB with KMnO4. At a fixed time of 20 min, the formed manganate ion is measured at 608 nm. The concentration of IPB is calculated using the regression equation for the fixed-time method, at 20 min. The determination of IPB by fixed-concentration and rate-constant methods is feasible with regression equations obtained, but the fixed-time method was found to be more applicable. The second method uses first-derivative (D1-) spectrophotometry for the determination of IPB at 254-268 nm. The applicability of the proposed methods was examined by analyzing Atrovent unit dose vials and the percentage recoveries were 100.01+/-1.16, 100.02+/-0.97, for kinetic and D1- methods, respectively. PMID- 10708403 TI - Extractive analysis of aluminum traces in dialysis solutions. AB - A very sensitive procedure for the fluorimetric determination of aluminum traces in dialysis solutions by means of Mordant Red 19 dyestuff is described with the extraction of the Al complex in isobutylmethylketone. The experimental conditions were studied, in order to obtain the best extraction yield. The emission intensity of the metal chelate, extracted in the organic layer, was measured at 549 nm, exciting at 485 nm. Linearity between emission intensity and Al concentration was found in the 1-30 ng/ml range. The limit of detection was 0.25 ng/ml. The method resulted to be suitable for the determination of Al traces in commercial dialysis solutions for toxicological purposes. PMID- 10708404 TI - Use of mass spectrometry for assessing similarity/diversity of natural products with unknown chemical structures. AB - An evaluation whether mass spectral data contain useful information for assessing similarity/diversity of drug compounds is presented. A comparative study was carried out between Ward's hierarchical agglomerative clustering, based on the 2D Daylight fingerprints or on the mass spectra, of a small database of 66 synthetic substances. The influence of normalization of the mass spectral data on the clustering result has also been studied. The results were subsequently compared with an expert's classification of the same small dataset, based on own evaluation according to known structure and pharmacological activity. PMID- 10708405 TI - Quantitative analysis of urinary acylglycines for the diagnosis of beta-oxidation defects using GC-NCI-MS. AB - The analysis of acylglycines is an important biochemical tool for the diagnosis of inherited disorders of mitochondrial fatty acid beta-oxidation. A stable isotope dilution gas chromatography negative chemical ionisation mass spectrometry method for the quantitative analysis of short- and medium-chain acylglycines as their bis(trifluoromethyl)benzyl (BTFMB) ester derivatives is described. The diagnostic usefulness of the method was demonstrated in nine patients with medium-chain acyl-coenzyme A (CoA) dehydrogenase (MCAD) deficiency, and seven patients with multiple acyl-CoA dehydrogenation defect (MAD). The urinary acylglycine profiles in these patients were compared to those in controls (n = 19), children on a medium-chain triglyceride (MCT) supplemented diet (n = 4), and patients with various other diseases (n = 5). PMID- 10708406 TI - Automated sample preparation of Roxifiban tablets: transfer of a manual method to an automated workstation. AB - Automation offers obvious advantages for the preparation of tablets prior to analysis by HPLC including unattended operation, minimization of human intervention and an electronic audit trail. However, significant effort has to be put in up front to develop and validate an automated method, particularly if it is required to closely follow an existing manual method. Here, method transfer for Roxifiban, a fibrinogen receptor antagonist, will be discussed. A Zymark tablet processing workstation II (TPWII) was used for all automated sample preparations. Manual methods for composite assay, content uniformity, weight variation and degradation products testing of a tablet formulation were transferred to the TPWII. The method involved weighing of the sample, disintegration of the dosage form by homogenization, extraction of the analyte in the homogenate solution, filtration of the homogenate, dilution of the filtrate and transfer to autosampler vials. Obstacles to a quick transfer included limitations in the volume capabilities of the TPWII, poor analyte solubility and achieving proper conditioning of the transfer lines and filter. After resolving these issues, a validated method was achieved. Spiked recoveries were from 99.4 to 101.1% (RSD's <0.5%). A cross-validation between automated and manual assay methods was compared by Westlake analysis giving a 0.7% calculated interval at the 95% confidence level. Carryover was 0.07% after 20 sample preparations at the highest tablet strength. PMID- 10708407 TI - Rapid, real-time sampling of R-84760 in blood by in vivo microdialysis with tandem mass spectrometry. AB - A new technique involving rapid sampling of R-84760 in real-time was achieved using a combination of microdialysis (MD) and tandem mass spectrometry (MS/MS). After collecting the analyte in real-time by MD and separating it by MS/MS, the ion intensities are adapted to the data without any subsequent chromatographic separation or flow injection analysis. The R-84760 concentration was obtained from the plateau part of the ion intensities or the corrected values using an internal standard, after immersing the MD probe into the dialysis solution containing the drug for a definite time. Since contamination of the ion source was prevented by using an organic solvent for the perfused solution, it was possible to establish a stable analysis method. For an MD membrane of length 4 mm, the R-84760 concentration in saline was linear over the range 5-541.1 ng/ml (r2 = 0.9997). Moreover, the R-84760 concentration in rat whole blood was linear over the range 24.9-1868.9 ng/ml (r2 = 0.9993). As this method allowed the measurement of free drug concentration in rat blood, the analysis was also able to provide data needed for determining the protein-binding ratio. The protein binding ratio obtained from the calibration curve in saline and rat whole blood was 87-90%, which was close to the result obtained by another analysis method. The concentration profile of R-84760 in blood as obtained by the MD-MS/MS method correlated well with the concentration profile in plasma, which was simultaneously monitored by LC-MS/MS. PMID- 10708408 TI - Second-derivative UV spectrometric determination of simvastatin in its tablet dosage form. AB - Simvastatin, a highly effective cholesterol-lowering agent, has been widely used for the treatment of hypercholesterolemia. During the development of simvastatin solid dosage form, formulation compositions were constantly varied to define a suitable matrix. A fast and reliable method for the dissolution and release testing of simvastatin was highly desirable to support formulation screening. A second derivative UV spectroscopic method was developed for determination of simvastatin in the tablet dosage form. After carefully choosing a zero-crossing technique of second derivative UV measurement at 243 nm, the selectivity and sensitivity of simvastatin was comparable to the previously developed HPLC method. In comparison with the direct UV method, second derivative UV spectroscopy eliminates the interference from UV absorbing excipients such as ascorbic acid, which often results in a bias of 2-10%. This method is also fast and economical in comparison to the more time-consuming HPLC method regularly used for formulation screening. Finally, this method has been validated to be precise and accurate, and is demonstrated to be an excellent alternative to HPLC method for the dissolution and release testing of simvastatin in the solid dosage form. PMID- 10708409 TI - Validation of immunoassays for bioanalysis: a pharmaceutical industry perspective. AB - Immunoassays are bioanalytical methods in which quantitation of the analyte depends on the reaction of an antigen (analyte) and an antibody. Although applicable to the analysis of both low molecular weight xenobiotic and macromolecular drugs, these procedures currently find most consistent application in the pharmaceutical industry to the quantitation of protein molecules. Immunoassays are also frequently applied in such important areas as the quantitation of biomarker molecules which indicate disease progression or regression, and antibodies elicited in response to treatment with macromolecular therapeutic drug candidates. Currently available guidance documents dealing with the validation of bioanalytical methods address immunoassays in only a limited way. This review highlights some of the differences between immunoassays and chromatographic assays, and presents some recommendations for specific aspects of immunoassay validation. Immunoassay calibration curves are inherently nonlinear, and require nonlinear curve fitting algorithms for best description of experimental data. Demonstration of specificity of the immunoassay for the analyte of interest is critical because most immunoassays are not preceded by extraction of the analyte from the matrix of interest. Since the core of the assay is an antigen-antibody reaction, immunoassays may be less precise than chromatographic assays; thus, criteria for accuracy (mean bias) and precision, both in pre-study validation experiments and in the analysis of in-study quality control samples, should be more lenient than for chromatographic assays. Application of the SFSTP (Societe Francaise Sciences et Techniques Pharmaceutiques) confidence interval approach for evaluating the total error (including both accuracy and precision) of results from validation samples is recommended in considering the acceptance/rejection of an immunoassay procedure resulting from validation experiments. These recommendations for immunoassay validation are presented in the hope that their consideration may result in the production of consistently higher quality data from the application of these methods. PMID- 10708410 TI - Clink, a nanovirus-encoded protein, binds both pRB and SKP1. AB - Clink, a 20-kDa protein of faba bean necrotic yellows virus, a single-stranded DNA plant virus, interacts with pRB family members and a SKP1 homologue from Medicago sativa. An LxCxE motif and an F-box of Clink mediate the interactions with the respective proteins. The capacity of Clink to bind pRB correlates with its ability to stimulate viral replication. Interaction of a single protein with the cell cycle regulator pRB and SKP1, a constituent of the ubiquitin-protein turnover pathway, appears to be a novel feature. Hence, Clink may represent a new class of viral cell cycle modulators. PMID- 10708411 TI - Classical swine fever virus E(rns) deletion mutants: trans-complementation and potential use as nontransmissible, modified, live-attenuated marker vaccines. AB - An SK6 cell line (SK6c26) which constitutively expressed the glycoprotein E(rns) of classical swine fever virus (CSFV) was used to rescue CSFV E(rns) deletion mutants based on the infectious copy of CSFV strain C. The biochemical properties of E(rns) from this cell line were indistinguishable from those of CSFV E(rns). Two E(rns) deletion mutants were constructed, virus Flc23 and virus Flc22. Virus Flc23 encoded only the utmost N- and C-terminal amino acids of E(rns) (deletion of 215 amino acids) to retain the original protease cleavage sites. Virus Flc22 is not recognized by a panel of E(rns) antibodies, due to a deletion of 66 amino acids in E(rns). The E(rns) deletion mutants Flc22 and Flc23 could be rescued in vitro only on the complementing SK6c26 cells. These rescued viruses could infect and replicate in SK6 cells but did not yield infectious virus. Virus neutralization by E(rns)-specific antibodies was similar for the wild-type virus and the recombinant viruses, indicating that E(rns) from SK6c26 cells was incorporated in the viral particles. Pigs vaccinated with Flc22 or Flc23 were protected against a challenge with a lethal dose of CSFV strain Brescia. This is the first demonstration of trans-complementation of defective pestivirus RNA with a pestiviral structural protein and opens new ways to develop nontransmissible modified live pestivirus vaccines. In addition, the absence of (the antigenic part of) E(rns) in the recombinant viral particles can be used to differentiate between infected and vaccinated animals. PMID- 10708412 TI - Reovirus-induced apoptosis requires activation of transcription factor NF-kappaB. AB - Reovirus infection induces apoptosis in cultured cells and in vivo. To identify host cell factors that mediate this response, we investigated whether reovirus infection alters the activation state of the transcription factor nuclear factor kappa B (NF-kappaB). As determined in electrophoretic mobility shift assays, reovirus infection of HeLa cells leads to nuclear translocation of NF-kappaB complexes containing Rel family members p50 and p65. Reovirus-induced activation of NF-kappaB DNA-binding activity correlated with the onset of NF-kappaB-directed transcription in reporter gene assays. Three independent lines of evidence indicate that this functional form of NF-kappaB is required for reovirus-induced apoptosis. First, treatment of reovirus-infected HeLa cells with a proteasome inhibitor prevents NF-kappaB activation following infection and substantially diminishes reovirus-induced apoptosis. Second, transient expression of a dominant negative form of IkappaB that constitutively represses NF-kappaB activation significantly reduces levels of apoptosis triggered by reovirus infection. Third, mutant cell lines deficient for either the p50 or p65 subunits of NF-kappaB are resistant to reovirus-induced apoptosis compared with cells expressing an intact NF-kappaB signaling pathway. These findings indicate that NF-kappaB plays a significant role in the mechanism by which reovirus induces apoptosis in susceptible host cells. PMID- 10708413 TI - Sequence heterogeneity of TT virus and closely related viruses. AB - TT virus (TTV) is a recently discovered infectious agent originally obtained from transfusion-related hepatitis. However, the causative link between the TTV infection and liver disease remains uncertain. Recent studies demonstrated that genome sequences of different TTV strains are significantly divergent. To assess genetic heterogeneity of the TTV genome in more detail, a sequence analysis of PCR fragments (271 bp) amplified from open reading frame 1 (ORF1) was performed. PCR fragments were amplified from 5 to 40% of serum specimens obtained from patients with different forms of hepatitis who reside in different countries (e.g., China, Egypt, Vietnam, and the United States) and from normal human specimens obtained from U.S. residents. A total of 170 PCR fragments were sequenced and compared to sequences derived from the corresponding TTV genome region deposited in GenBank. Genotypes 2 and 3 were found to be significantly more genetically related than any other TTV genotype. Moreover, three sequences were shown to be almost equally related to both genotypes 2 and 3. These observations suggest a merger of genotypes 2 and 3 into one genotype, 2/3. Additionally, five new groups of TTV sequences were identified. One group represents a new genotype, whereas the other four groups were shown to be more evolutionary distant from all known TTV sequences. The evolutionary distances between these four groups were also shown to be greater than between TTV genotypes. The phylogenetic analysis suggested that these four new genetic groups represent closely related yet different viral species. Thus, TTV exists as a "swarm" of at least five closely related but different viruses. These observations suggest a high degree of genetic complexity within the TTV population. The finding of the additional TTV-related species should be taken into consideration when the association between TTV infections and human diseases of unknown etiology is studied. PMID- 10708414 TI - Evolution of the Sabin strain of type 3 poliovirus in an immunodeficient patient during the entire 637-day period of virus excretion. AB - A 20-year-old female hypogammaglobulinemic patient received monotypic Sabin 3 vaccine in 1962. The patient excreted type 3 poliovirus for a period of 637 days without developing any symptoms of poliomyelitis, after which excretion appeared to have ceased spontaneously. The evolution of Sabin 3 throughout the entire period of virus excretion was studied by characterization of seven sequential isolates from the patient. The isolates were analyzed in terms of their antigenic properties, virulence, sensitivity for growth at high temperatures, and differences in nucleotide sequence from the Sabin type 3 vaccine. The isolates followed a main lineage of evolution with a rate of nucleotide substitution that was very similar to that estimated for wild-type poliovirus during person-to person transmission. There was a delay in the appearance of antigenic variants compared to sequential type 3 isolates from healthy vaccines, which could be one of the possible explanations for the long-term excretion of virus from the patient. The distribution of mutations in the isolates identified regions of the virus possibly involved in adaptation for growth in the human gut and virus persistence. None of the isolates showed a full reversion of the attenuated and temperature-sensitive phenotypes of Sabin 3. Information of this sort will help in the assessment of the risk of spread of virulent polioviruses from long-term excretors and in the design of therapies to stop long-term excretion. This will make an important contribution to the decision-making process on when to stop vaccination once wild poliovirus has been eradicated. PMID- 10708415 TI - Attenuation markers of a candidate dengue type 2 vaccine virus, strain 16681 (PDK 53), are defined by mutations in the 5' noncoding region and nonstructural proteins 1 and 3. AB - The genome of a candidate dengue type 2 (DEN-2) vaccine virus, strain PDK-53, differs from its DEN-2 16681 parent by nine nucleotides. Using infectious cDNA clones, we constructed 18 recombinant 16681/PDK-53 viruses to analyze four 16681 to-PDK-53 mutations, including 5' noncoding region (5'NC)-57 C-to-T, premembrane (prM)-29 Asp-to-Val (the only mutation that occurs in the structural proteins), nonstructural protein 1 (NS1)-53 Gly-to-Asp, and NS3-250 Glu-to-Val. The viruses were studied for plaque size, growth rate, and temperature sensitivity in LLC MK(2) cells, growth rate in C6/36 cells, and neurovirulence in newborn mice. All of the viruses replicated to peak titers of 10(7.3) PFU/ml or greater in LLC MK(2) cells. The crippled replication of PDK-53 virus in C6/36 cells and its attenuation for mice were determined primarily by the 5'NC-57-T and NS1-53-Asp mutations. The temperature sensitivity of PDK-53 virus was attributed to the NS1 53-Asp and NS3-250-Val mutations. The 5'NC-57, NS1-53, and NS3-250 loci all contributed to the small-plaque phenotype of PDK-53 virus. Reversions at two or three of these loci in PDK-53 virus were required to reconstitute the phenotypic characteristics of the parental 16681 virus. The prM-29 locus had little or no effect on viral phenotype. Sequence analyses showed that PDK-53 virus is genetically identical to PDK-45 virus. Restriction of the three major genetic determinants of attenuation markers to nonstructural genomic regions makes the PDK-53 virus genotype attractive for the development of chimeric DEN virus vaccine candidates. PMID- 10708416 TI - Chimeric dengue type 2 (vaccine strain PDK-53)/dengue type 1 virus as a potential candidate dengue type 1 virus vaccine. AB - We constructed chimeric dengue type 2/type 1 (DEN-2/DEN-1) viruses containing the nonstructural genes of DEN-2 16681 virus or its vaccine derivative, strain PDK 53, and the structural genes (encoding capsid protein, premembrane protein, and envelope glycoprotein) of DEN-1 16007 virus or its vaccine derivative, strain PDK 13. We previously reported that attenuation markers of DEN-2 PDK-53 virus were encoded by genetic loci located outside the structural gene region of the PDK-53 virus genome. Chimeric viruses containing the nonstructural genes of DEN-2 PDK-53 virus and the structural genes of the parental DEN-1 16007 virus retained the attenuation markers of small plaque size and temperature sensitivity in LLC-MK(2) cells, less efficient replication in C6/36 cells, and attenuation for mice. These chimeric viruses elicited higher mouse neutralizing antibody titers against DEN-1 virus than did the candidate DEN-1 PDK-13 vaccine virus or chimeric DEN-2/DEN-1 viruses containing the structural genes of the PDK-13 virus. Mutations in the envelope protein of DEN-1 PDK-13 virus affected in vitro phenotype and immunogenicity in mice. The current PDK-13 vaccine is the least efficient of the four Mahidol candidate DEN virus vaccines in human trials. The chimeric DEN-2/DEN 1 virus might be a potential DEN-1 virus vaccine candidate. This study indicated that the infectious clones derived from the candidate DEN-2 PDK-53 vaccine are promising attenuated vectors for development of chimeric flavivirus vaccines. PMID- 10708417 TI - A single-amino-acid substitution of a tyrosine residue in the rubella virus E1 cytoplasmic domain blocks virus release. AB - Rubella virus particles, consisting of a nucleocapsid surrounded by a lipid envelope in which two virus-encoded glycoproteins E1 and E2 are embedded, assemble on intracellular membranes and are secreted from cells, possibly via the cellular secretory pathway. We have recently demonstrated that the cytoplasmic domain of E1 (residues 469 to 481, KCLYYLRGAIAPR) is required for virus release. Alteration of cysteine 470 to alanine did not affect virus release, whereas mutation of leucine 471 to alanine reduced virus production by 90%. In the present study, substitutions of remaining amino acids in the E1 cytoplasmic domain were made in order to investigate the role of each amino acid in regulating rubella virus release. Generated mutants were analyzed in the context of infectious full-length cDNA clone and virus-like particles using combined genetic, biochemical, and electron microscopic approaches. Substitution of a single residue of tyrosine 472 to alanine or tyrosine 473 to serine resulted in a block in virus release without affecting protein transport and virus budding into the lumen of the Golgi complexes. Infectious RNA transcripts bearing these mutations were incapable of forming plaques. Mutants with substitutions at the amino-terminal region (leucine 474, arginine 475, and glycine 476) in the E1 cytoplasmic domain had reduced virus release and small-plaque phenotype, while mutants with substitutions at the carboxy-terminal region (alanine 477, isoleucine 478, alanine 479, proline 480, and arginine 481) had only marginal defects in virus release. Plaque-forming revertants could be isolated from mutants Y472A and Y473S. Sequencing analysis revealed that the substituted serine residue in mutant Y473S reverted to the original tyrosine residue, whereas the substituted alanine residue in mutant Y472A was retained. These results indicate that the E1 cytoplasmic domain modulates virus release in a sequence-dependent manner and that the tyrosine residues are critical for this function. We postulate that residues YYLRG constitute a domain in the E1 tail that may interact with other proteins and this interaction is involved in regulating virus release. PMID- 10708418 TI - Erythroid cells rendered erythropoietin independent by infection with Friend spleen focus-forming virus show constitutive activation of phosphatidylinositol 3 kinase and Akt kinase: involvement of insulin receptor substrate-related adapter proteins. AB - The erythroleukemia-inducing Friend spleen focus-forming virus (SFFV) encodes a unique envelope glycoprotein which allows erythroid cells to proliferate and differentiate in the absence of erythropoietin (Epo). In an effort to understand how SFFV causes Epo independence, we have been examining erythroid cells rendered factor independent by SFFV infection for constitutive activation of signal transducing molecules. Previous studies from our laboratory showed that various signal-transducing molecules known to be activated by Epo, including Stat proteins and components of the Raf-1/MAP kinase pathway, are constitutively activated in SFFV-infected erythroid cells in the absence of Epo. Since another signal transduction pathway involving activation of phosphatidylinositol 3-kinase (PI 3-kinase) after Epo stimulation plays an important role in erythroid cell proliferation and differentiation, we carried out studies to determine if this pathway was also activated in SFFV-infected cells in the absence of Epo. Our studies show that PI 3-kinase is constitutively activated in erythroid cells rendered factor independent by infection with SFFV and that PI 3-kinase activity, but not Epo receptor tyrosine phosphorylation, is required for the proliferation of these cells in the absence of Epo. We further show that in SFFV-infected erythroid cells grown in the absence of Epo, PI 3-kinase associates with the insulin receptor substrate (IRS)-related adapter molecules IRS-2, Gab1, and Gab2, which are constitutively tyrosine phosphorylated in SFFV-infected cells. Finally, Akt, a protein kinase that is one of the downstream effectors of PI 3-kinase, and SHIP, a lipid phosphatase that is important for Akt activation through PI 3 kinase, are both tyrosine phosphorylated in SFFV-infected cells grown in the absence of Epo. Our results indicate that induction of Epo independence by SFFV requires the activation of PI 3-kinase and suggest that constitutive activation of this kinase in SFFV-infected cells may occur primarily through interaction of PI 3-kinase with constitutively phosphorylated IRS-related adapter molecules. PMID- 10708419 TI - Basic residues in human immunodeficiency virus type 1 nucleocapsid promote virion assembly via interaction with RNA. AB - Retroviral Gag polyproteins drive virion assembly by polymerizing to form a spherical shell that lines the inner membrane of nascent virions. Deletion of the nucleocapsid (NC) domain of the Gag polyprotein disrupts assembly, presumably because NC is required for polymerization. Human immunodeficiency virus type 1 NC possesses two zinc finger motifs that are required for specific recognition and packaging of viral genomic RNA. Though essential, zinc fingers and genomic RNA are not required for virion assembly. NC promiscuously associates with cellular RNAs, many of which are incorporated into virions. It has been hypothesized that Gag polymerization and virion assembly are promoted by nonspecific interaction of NC with RNA. Consistent with this model, we found an inverse relationship between the number of NC basic residues replaced with alanine and NC's nonspecific RNA binding activity, Gag's ability to polymerize in vitro and in vivo, and Gag's capacity to assemble virions. In contrast, mutation of NC's zinc fingers had only minor effects on these properties. PMID- 10708420 TI - Hypervariable region 1 sequence stability during hepatitis C virus replication in chimpanzees. AB - The putative envelope 2 (E2) gene of hepatitis C virus (HCV) contains a highly variable region referred to as hypervariable region 1 (HVR1). We hypothesized that this genetic variability is driven by immune selection pressure, rather than representing the accumulation of random mutations in a region with relatively little functional constraint. To test this hypothesis, we examined the E2 sequence of a human inoculum that was passaged through eight chimpanzees, which appear to have a replicative rate (opportunity for chance mutation) similar to that of humans. Acute-phase plasma samples from a human (the inoculum) and six of eight serially infected chimpanzees were studied. For each, 33 cloned cDNAs were examined by a combined heteroduplex-single-stranded conformational polymorphism assay to assess quasispecies complexity and optimize selection of clones with unique gel shift patterns (clonotypes) for sequencing. The sequence diversity of HCV was significantly lower in the chimpanzees than in the humans, and during eight serial passages there was no change in the sequence of the majority clonotype from each animal examined. Similarly, the rates of protein sequence altering (nonsynonymous) substitution were lower in the chimpanzees than in the humans. These findings demonstrate that nonsynonymous mutations indicate selection pressure rather than being an incidental result of HCV replication. PMID- 10708421 TI - Alternative proteolytic processing of mouse mammary tumor virus superantigens. AB - Mouse mammary tumor viruses express a superantigen essential for their life cycle. It has been proposed that viral superantigens (vSags) require processing by prohormone convertases (PCs) for activity. We now observe, using a panel of mutant forms of potential PC cleavage sites and in vitro cleavage assays, that only the CS1 (position 68 to 71) and CS2 (position 169 to 172) sites are utilized by furin and PC5. Other members of the convertase family that are expressed in lymphocytes are not endowed with this activity. Furthermore, mutant forms of two different viral superantigens, vSag7 and vSag9, which completely abrogated in vitro processing by convertases, were efficient in functional presentation to responsive T-cell hybridomas. This effect was observed in both endogenous presentation and paracrine transfer of the vSag. Processing by convertases thus appears not to be essential for vSag function. Finally, we have identified the purified endosomal protease cathepsin L as another protease that is able to cleave convertase mutant vSag in vitro, yielding fragments similar to those detected in vivo, thus suggesting that proteases other than convertases are involved in the activation of vSags. PMID- 10708422 TI - Mengovirus and encephalomyocarditis virus poly(C) tract lengths can affect virus growth in murine cell culture. AB - Many virulent aphthoviruses and cardioviruses have long homopolymeric poly(C) tracts in the 5' untranslated regions of their RNA genomes. A panel of genetically engineered mengo-type cardioviruses has been described which contain a variety of different poly(C) tract lengths. Studies of these viruses have shown the poly(C) tract to be dispensable for growth in HeLa cells, although the relative murine virulence of the viruses correlates directly and positively with tract length. Compared with wild-type mengovirus strain M, mutants with shortened poly(C) tracts grow poorly in mice and protectively immunize rather than kill recipient animals. In the present study, several murine cell populations were tested to determine whether, unlike HeLa cells, they allowed a differential amplification of viruses with long or short poly(C) tracts. Replication and cytopathic studies with four hematopoietically derived cell lines (CH2B, RAW 264.7, A20.J, and P815) and two murine fibroblast cell lines [L929 and L(Y)] demonstrated that several of these cell types indeed allowed differential virus replication as a function of viral poly(C) tract length. Among the most discerning of these cells, RAW 264.7 macrophages supported vigorous lytic growth of a long-tract virus, vMwt (C(44)UC(10)), but supported only substantially diminished and virtually nonlytic growth of vMC(24) (C(13)UC(10)) and vMC(0) short-tract viruses. The viral growth differences evident in all cell lines were apparent early and continuously during every cycle of virus amplification. The data suggest that poly(C) tract-dependent attenuation of mengovirus may be due in part to a viral replication defect manifest in similar hematopoietic-type cells shortly after murine infection. The characterized cultures should provide excellent tools for molecular study of poly(C) tract-mediated virulence. PMID- 10708423 TI - Structure and coding content of CST (BART) family RNAs of Epstein-Barr virus. AB - CST (BART BARF0) family viral RNAs are expressed in several types of Epstein-Barr virus (EBV) infection, including EBV-associated cancers. Many different spliced forms of these RNAs have been described; here we have clarified the structures of some of the more abundant splicing patterns. We report the first cDNAs representing a full-length CST mRNA from a clone library and further characterize the transcription start. The relative abundance of splicing patterns and genomic analysis of the open reading frames (ORFs) suggest that, in addition to the much studied BARF0 ORF, there may be important products made from some of the upstream ORFs in the CST RNAs. Potential biological functions are identified for two of these. The product of the RPMS1 ORF is shown to be a nuclear protein that can bind to the CBF1 component of Notch signal transduction. RPMS1 can inhibit the transcription activation induced through CBF1 by NotchIC or EBNA-2. The protein product of another CST ORF, A73, is shown to be a cytoplasmic protein which can interact with the cell RACK1 protein. Since RACK1 modulates signaling from protein kinase C and Src tyrosine kinases, the results suggest a possible role for CST products in growth control, perhaps consistent with the abundant transcription of CST RNAs in cancers such as nasopharyngeal carcinoma. PMID- 10708424 TI - A protein kinase activity associated with Epstein-Barr virus BGLF4 phosphorylates the viral early antigen EA-D in vitro. AB - The Epstein-Barr virus (EBV) open reading frame BGLF4 was identified as a potential Ser/Thr protein kinase gene through the recognition of amino acid sequence motifs characteristic of conserved regions within the catalytic domains of protein kinases. In order to investigate this potential kinase activity, BGLF4 was expressed in Escherichia coli and the purified protein was used to generate a specific antiserum. Recombinant vaccinia virus vTF7-3, which expresses the T7 RNA polymerase, was used to infect 293 and 293T cells after transient transfection with a plasmid containing BGLF4 under the control of the T7 promoter. Autophosphorylation of the BGLF4 protein was demonstrated using the specific antiserum in an immune complex kinase assay. In addition, EBNA-1-tagged BGLF4 and EBNA-1 monoclonal antibody 5C11 were used to demonstrate the specificity of the kinase activity and to locate BGLF4 in the cytoplasm of transfected cells. Manganese ions were found to be essential for autophosphorylation of BGLF4, and magnesium can stimulate the activity. BGLF4 can utilize GTP, in addition to ATP, as a phosphate donor in this assay. BGLF4 can phosphorylate histone and casein in vitro. Among the potential viral protein substrates we examined, the EBV early antigen (EA-D, BMRF1), a DNA polymerase accessory factor and an important transactivator during lytic infection, was found to be phosphorylated by BGLF4 in vitro. Amino acids 1 to 26 of BGLF4, but not the predicted conserved catalytic domain, were found to be essential for autophosphorylation of BGLF4. PMID- 10708425 TI - Human immunodeficiency virus type 1 vpr induces apoptosis through caspase activation. AB - Human immunodeficiency virus type 1 (HIV-1) Vpr is a 96-amino-acid protein that is found associated with the HIV-1 virion. Vpr induces cell cycle arrest at the G(2)/M phase of the cell cycle, and this arrest is followed by apoptosis. We examined the mechanism of Vpr-induced apoptosis and found that HIV-1 Vpr-induced apoptosis requires the activation of a number of cellular cysteinyl aspartate specific proteases (caspases). We demonstrate that ectopic expression of anti apoptotic viral proteins, which inhibit caspase activity, and addition of synthetic peptides, which represent caspase cleavage sites, can inhibit Vpr induced apoptosis. Finally, inhibition of caspase activity and subsequent inhibition of apoptosis results in increased viral expression, suggesting that therapeutic strategies aimed at reducing Vpr-induced apoptosis in vivo require careful consideration. PMID- 10708426 TI - Tissue sites of persistent infection and active replication of equine infectious anemia virus during acute disease and asymptomatic infection in experimentally infected equids. AB - Equine infectious anemia virus (EIAV) infection of horses is characterized by recurring cycles of disease and viremia that typically progress to an inapparent infection in which clinical symptoms are absent as host immune responses maintain control of virus replication indefinitely. The dynamics of EIAV viremia and its association with disease cycles have been well characterized, but there has been to date no comprehensive quantitative analyses of the specific tissue sites of EIAV infection and replication in experimentally infected equids during acute disease episodes and during asymptomatic infections in long-term inapparent carriers. To characterize the in vivo site(s) of viral infection and replication, we developed a quantitative competitive PCR assay capable of detecting 10 copies of viral DNA and a quantitative competitive reverse transcription-PCR assay with a sensitivity of about 30 copies of viral singly spliced mRNA. Animals were experimentally infected with one of two reference viruses: the animal-passaged field isolate designated EIAV(Wyo) and the virulent cell-adapted strain designated EIAV(PV). Tissues and blood cells were isolated during the initial acute disease or from asymptomatic animals and analyzed for viral DNA and RNA levels by the respective quantitative assays. The results of these experiments demonstrated that the appearance of clinical symptoms in experimentally infected equids coincided with rapid widespread seeding of viral infection and replication in a variety of tissues. During acute disease, the predominant cellular site of viral infection and replication was the spleen, which typically accounted for over 90% of the cellular viral burden. In asymptomatic animals, viral DNA and RNA persisted in virtually all tissues tested, but at extremely low levels, a finding indicative of tight but incomplete immune control of EIAV replication. During all disease states, peripheral blood mononuclear cells (PBMC) were found to harbor less than 1% of the cellular viral burden. These quantitative studies demonstrate that tissues, rather than PBMC, constitute the predominant sites of virus replication during acute disease in infected equids and serve as resilient reservoirs of virus infection, even in the presence of highly effective immune responses that maintain a stringent control of virus replication in long-term inapparent carriers. Thus, these observations with EIAV, a predominantly macrophage-tropic lentivirus, highlight the role of tissues in sequestering lentiviral infections from host immune surveillance. PMID- 10708427 TI - An adeno-associated virus (AAV) initiator protein, Rep78, catalyzes the cleavage and ligation of single-stranded AAV ori DNA. AB - The Rep78 protein of adeno-associated virus (AAV) contains amino acid sequence motifs common to rolling-circle replication (RCR) initiator proteins. In this report, we describe RCR initiator-like activities of Rep78. We demonstrate that a maltose-binding protein (MBP)-Rep78 fusion protein can catalyze the cleavage and ligation of single-stranded DNA substrates derived from the AAV origin of replication. Rep-mediated single-stranded DNA cleavage was strictly dependent on the presence of certain divalent cations (e.g., Mn(2+) or Mg(2+)) but did not require the presence of a nucleoside triphosphate cofactor. Electrophoretic mobility shift assays demonstrated that binding of single-stranded DNA by MBP Rep78 was influenced by the length of the substrate as well as the presence of potential single-stranded cis-acting sequence elements. Site-directed mutagenesis was used to examine the role of specific tyrosine residues within a conserved RCR motif (motif 3) of Rep78. Replacement of Tyr-156 with phenylalanine abolished the ability of MBP-Rep78 to mediate the cleavage and ligation of single-stranded DNA substrates but not the ability to stably bind single-stranded DNA. The cleaving joining activity of Rep78 is consistent with the mechanism of replicative intermediate dimer resolution proposed for the autonomous parvoviruses and may have implications for targeted integration of recombinant AAV vectors. PMID- 10708428 TI - Evolutionarily related Sindbis-like plant viruses maintain different levels of population diversity in a common host. AB - The levels of population diversity of three related Sindbis-like plant viruses, Tobacco mosaic virus (TMV), Cucumber mosaic virus (CMV), and Cowpea chlorotic mottle virus (CCMV), in infections of a common host, Nicotiana benthamiana, established from genetically identical viral RNA were examined. Despite probably having a common evolutionary ancestor, the three viruses maintained different levels of population diversity. CMV had the highest levels of diversity, TMV had an intermediate level of diversity, and CCMV had no measurable level of diversity in N. benthamiana. Interestingly, the levels of diversity were correlated to the relative host range sizes of the three viruses. The levels of diversity also remained relatively constant over the course of serial passage. Closer examination of the CMV and TMV populations revealed biases for particular types of substitutions and regions of the genome that may tolerate fewer mutations. PMID- 10708429 TI - Amphotericin B inhibits the generation of the scrapie isoform of the prion protein in infected cultures. AB - Transmissible spongiform encephalopathies form a group of fatal neurodegenerative disorders that have the unique property of being infectious, sporadic, or genetic in origin. Although some doubts about the nature of the responsible agent of these diseases remain, it is clear that a protein called PrP(Sc) plays a central role. PrP(Sc) is a conformational variant of PrP(C), the normal host protein. Polyene antibiotics such as amphotericin B have been shown to delay the accumulation of PrP(Sc) and to increase the incubation time of the disease after experimental transmission in laboratory animals. Unlike for Congo red and sulfated polyanions, no effect of amphotericin B has been observed in infected cultures. We show here for the first time that amphotericin B can inhibit PrP(Sc) generation in scrapie-infected GT1-7 and N2a cells. Its activity seems to be related to a modification of the properties of detergent-resistant microdomains. These results provide new insights into the mechanism of action of amphotericin B and confirm the usefulness of infected cultures in the therapeutic research of transmissible spongiform encephalopathies. PMID- 10708430 TI - Complex effects of deletions in the 5' untranslated region of primate foamy virus on viral gene expression and RNA packaging. AB - Due to various advantageous features there is current interest in retroviral vectors derived from primate foamy viruses (PFVs). Two PFV cis-acting sequences have been mapped in the 5' region of the RNA (pre-)genome and in the 3' pol genomic region. In order to genetically separate PFV packaging constructs from vector constructs, we investigated the effect of deletions in the 5' untranslated region (UTR) of PFV packaging constructs and vectors on gene expression and RNA incorporation into viral particles. Our results indicate that the 5' UTR serves different previously unknown functions. First, the R region of the long terminal repeat was found to be required for PFV gag gene expression. This regulation of gene expression appeared to be mainly posttranscriptional. Second, constructs with sequence deletions between the R region and the gag gene start codon packaged as much viral mRNA into particles as the undeleted construct, and RNA from such a 5'-UTR-deleted packaging construct was copackaged into vector-virus particles, together with vector RNA which was preferentially packaged. Finally, in the U5 region a sequence was identified that was required to allow cleavage of the Gag precursor protein by the pol gene-encoded protease, suggesting a role of RNA in PFV particle formation. Taken together, the results indicate that complex interactions of the viral RNA, capsid, and polymerase proteins take place during PFV particle formation and that a clear separation of PFV vector and packaging construct sequences may be difficult to achieve. PMID- 10708431 TI - Complete nucleotide sequence and genome organization of hibiscus chlorotic ringspot virus, a new member of the genus Carmovirus: evidence for the presence and expression of two novel open reading frames. AB - The complete nucleotide sequence of hibiscus chlorotic ringspot virus (HCRSV) was determined. The genomic RNA (gRNA) is 3,911 nucleotides long and has the potential to encode seven viral proteins in the order of 28 (p28), 23 (p23), 81 (p81), 8 (p8), 9 (p9), 38 (p38), and 25 (p25) kDa. Excluding two unique open reading frames (ORFs) encoding p23 and p25, the ORFs encode proteins with high amino acid similarity to those of carmoviruses. In addition to gRNA, two 3' coterminated subgenomic RNA (sgRNA) species were identified. Full-length cDNA clones derived from gRNA and sgRNA were constructed under the control of a T7 promoter. Both capped and uncapped transcripts derived from the full-length genomic cDNA clone were infectious. In vitro translation and mutagenesis assays confirmed that all the predicted ORFs except the ORF encoding p8 are translatable, and the two novel ORFs (those encoding p23 and p25) may be functionally indispensable for the viral infection cycle. Based on virion morphology and genome organization, we propose that HCRSV be classified as a new member of the genus Carmovirus in family Tombusviridae. PMID- 10708432 TI - Isolation and characterization of an arterivirus defective interfering RNA genome. AB - Equine arteritis virus (EAV), the type member of the family Arteriviridae, is a single-stranded RNA virus with a positive-stranded genome of approximately 13 kb. EAV uses a discontinuous transcription mechanism to produce a nested set of six subgenomic mRNAs from which its structural genes are expressed. We have generated the first documented arterivirus defective interfering (DI) RNAs by serial undiluted passaging of a wild-type EAV stock in BHK-21 cells. A cDNA copy of the smallest DI RNA (5.6 kb) was cloned. Upon transfection into EAV-infected BHK-21 cells, transcripts derived from this clone (pEDI) were replicated and packaged. Sequencing of pEDI revealed that the DI RNA was composed of three segments of the EAV genome (nucleotides 1 to 1057, 1388 to 1684, and 8530 to 12704) which were fused in frame with respect to the replicase reading frame. Remarkably, this DI RNA has retained all of the sequences encoding the structural proteins. By insertion of the chloramphenicol acetyltransferase reporter gene in the DI RNA genome, we were able to delimitate the sequences required for replication/DI based transcription and packaging of EAV DI RNAs and to reduce the maximal size of a replication-competent EAV DI RNA to approximately 3 kb. PMID- 10708433 TI - E1A blocks hyperphosphorylation of p130 and p107 without affecting the phosphorylation status of the retinoblastoma protein. AB - The phosphorylation status of the pRB family of growth suppressor proteins is regulated in a cell cycle entry-, progression-, and exit-dependent manner in normal cells. We have shown previously that p130, a member of this family, exhibits patterns of phosphorylated forms associated with various cell growth and differentiation stages. However, human 293 cells, which are transformed cells that express the adenoviral oncoproteins E1A and E1B, exhibit an abnormal pattern of p130 phosphorylated forms. Here we report that, unlike pRB, the phosphorylation status of both p130 and p107 is not modulated during the cell cycle in 293 cells as it is in other cells. Conditional overexpression of individual G(1)/S cyclins in 293 cells does not alter the phosphorylation status of p130, suggesting that the expression of E1A and/or E1B blocks hyperphosphorylation of p130. In agreement with these observations, transient cotransfection of vectors expressing E1A 12S, but not E1B, in combination with pocket proteins into U-2 OS cells blocks hyperphosphorylation of both p130 and p107. However, the phosphorylation status of pRB is not altered by cotransfection of E1A 12S vectors. Moreover, MC3T3-E1 preosteoblasts stably expressing E1A 12S also exhibit a block in hyperphosphorylation of endogenous p130 and p107. Direct binding of E1A to p130 and p107 is not required for the phosphorylation block since E1A 12S mutants defective in binding to the pRB family also block hyperphosphorylation of p130 and p107. Our data reported here identify a novel function of E1A, which affects p130 and p107 but does not affect pRB. Since E1A does not bind the hyperphosphorylated forms of p130, this function of E1A might prevent the existence of "free" hyperphosphorylated p130, which could act as a CDK inhibitor. PMID- 10708434 TI - DNA methylation of helper virus increases genetic instability of retroviral vector producer cells. AB - Retroviral vector producer cells (VPC) have been considered genetically stable. A clonal cell population exhibiting a uniform vector integration pattern is used for sustained vector production. Here, we observed that the vector copy number is increased and varied in a population of established LTKOSN.2 VPC. Among five subclones of LTKOSN.2 VPC, the vector copy number ranged from 1 to approximately 29 copies per cell. A vector superinfection experiment and Northern blot analysis demonstrated that suppression of helper virus gene expression decreased Env receptor interference and allowed increased superinfection. The titer production was tightly associated with helper virus gene expression and varied between 0 and 2.2 x 10(5) CFU/ml in these subclones. In one analyzed subclone, the number of integrated vectors increased from one copy per cell to nine copies per cell during a 31-day period. Vector titer was reduced from 1.5 x 10(5) CFU to an undetectable level. To understand the mechanism involved, helper virus and vectors were examined for DNA methylation status by methylation-sensitive restriction enzyme digestion. We demonstrated that DNA methylation of helper virus 5' long terminal repeat occurred in approximately 2% of the VPC population per day and correlated closely with inactivation of helper virus gene expression. In contrast, retroviral vectors did not exhibit significant methylation and maintained consistent transcription activity. Treatment with 5-azacytidine, a methylation inhibitor, partially reversed the helper virus DNA methylation and restored a portion of vector production. The preference for methylation of helper virus sequences over vector sequences may have important implications for host virus interaction. Designing a helper virus to overcome cellular DNA methylation may therefore improve vector production. The maintenance of increased viral envelope-receptor interference might also prevent replication-competent retrovirus formation. PMID- 10708435 TI - A recombinant human parainfluenza virus type 3 (PIV3) in which the nucleocapsid N protein has been replaced by that of bovine PIV3 is attenuated in primates. AB - The shipping fever (SF) and Kansas (Ka) strains of bovine parainfluenza virus type 3 (BPIV3) are restricted in their replication in rhesus monkeys 100- to 1,000-fold compared to human parainfluenza virus type 3 (HPIV3), and the Ka strain also was shown to be attenuated in humans. To initiate an investigation of the genetic basis of the attenuation of BPIV3 in primates, we produced viable chimeric HPIV3 recombinants containing the nucleoprotein (N) open reading frame (ORF) from either BPIV3 Ka or SF in place of the HPIV3 N ORF. These chimeric recombinants were designated cKa-N and cSF-N, respectively. Remarkably, cKa-N and cSF-N grew to titers comparable to those of their HPIV3 and BPIV3 parents in LLC MK2 monkey kidney and Madin-Darby bovine kidney cells. Thus, the heterologous nature of the N protein did not impede replication in vitro. However, cKa-N and cSF-N were each restricted in replication in rhesus monkeys to a similar extent as Ka and SF, respectively. This identified the BPIV3 N protein as a determinant of the host range restriction of BPIV3 in primates. These chimeras thus combine the antigenic determinants of HPIV3 with the host range restriction and attenuation phenotype of BPIV3. Despite their restricted replication in rhesus monkeys, the chimeric viruses induced a level of resistance to HPIV3 challenge in these animals which was indistinguishable from that conferred by immunization with HPIV3. The infectivity, attenuation, and immunogenicity of these BPIV3/HPIV3 chimeras suggest that the modified Jennerian approach described in the present report represents a novel method to design vaccines to protect against HPIV3 induced disease in humans. PMID- 10708436 TI - Human immunodeficiency virus type 1 pathogenesis in SCID-hu mice correlates with syncytium-inducing phenotype and viral replication. AB - Human immunodeficiency virus type 1 (HIV-1) patient isolates and molecular clones were used to analyze the determinants responsible for human CD4(+) thymocyte depletion in SCID-hu mice. Non-syncytium-inducing, R5 or R3R5 HIV-1 isolates from asymptomatic infected people showed little or no human CD4(+) thymocyte depletion in SCID-hu mice, while syncytium-inducing (SI), R5X4 or R3R5X4 HIV-1 isolates from the same individuals, isolated just prior to the onset of AIDS, rapidly and efficiently eliminated CD4-bearing human thymocytes. We have mapped the ability of one SI HIV-1 isolate to eliminate CD4(+) human cells in SCID-hu mice to a region of the env gene including the three most amino-terminal variable regions (V1 to V3). We find that for all of the HIV-1 isolates that we studied, a nonlinear relationship exists between viral replication and the depletion of CD4(+) cells. This relationship can best be described mathematically with a Hill type plot indicating that a threshold level of viral replication, at which cytopathic effects begin to be seen, exists for HIV-1 infection of thymus/liver grafts in SCID-hu mice. This threshold level is 1 copy of viral DNA for every 11 cells (95% confidence interval = 1 copy of HIV-1 per 67 cells to 1 copy per 4 cells). Furthermore, while SI viruses more frequently achieve this level of replication, replication above this threshold level correlates best with cytopathic effects in this model system. We used GHOST cells to map the coreceptor specificity and relative entry efficiency of these early- and late stage patient isolates of HIV-1. Our studies show that coreceptor specificity and entry efficiency are critical determinants of HIV-1 pathogenesis in vivo. PMID- 10708438 TI - Development of an effective polyvalent vaccine against both Marek's and Newcastle diseases based on recombinant Marek's disease virus type 1 in commercial chickens with maternal antibodies. AB - An earlier report (M. Sakaguchi et al., Vaccine 16:472-479, 1998) showed that recombinant Marek's disease virus type 1 (rMDV1) expressing the fusion (F) protein of Newcastle disease virus (NDV-F) under the control of the simian virus 40 late promoter [rMDV1-US10L(F)] protected specific pathogen-free chickens from NDV challenge, but not commercial chickens with maternal antibodies against NDV and MDV1. In the present study, we constructed an improved polyvalent vaccine based on MDV1 against MDV and NDV in commercial chickens with maternal antibodies. The study can be summarized as follows. (i) We constructed rMDV1 expressing NDV-F under the control of the MDV1 glycoprotein B (gB) promoter [rMDV1-US10P(F)]. (ii) Much less NDV-F protein was expressed in cells infected with rMDV1-US10P(F) than in those infected with rMDV1-US10L(F). (iii) The antibody response against NDV-F and MDV1 antigens of commercial chickens vaccinated with rMDV1-US10P(F) was much stronger and faster than with rMDV1 US10L(F), and a high level of antibody against NDV-F persisted for over 80 weeks postvaccination. (iv) rMDV1-US10P(F) was readily reisolated from the vaccinated chickens, and the recovered viruses were found to express NDV-F. (v) Vaccination of commercial chickens having maternal antibodies to rMDV1-US10P(F) completely protected them from NDV challenge. (vi) rMDV1-US10P(F) offered the same degree of protection against very virulent MDV1 as the parental MDV1 and commercial vaccines. These results indicate that rMDV1-US10P(F) is an effective and stable polyvalent vaccine against both Marek's and Newcastle diseases even in the presence of maternal antibodies. PMID- 10708437 TI - Pathogenesis of primary R5 human immunodeficiency virus type 1 clones in SCID-hu mice. AB - We studied the replication and cytopathicity in SCID-hu mice of R5 human immunodeficiency virus type 1 (HIV-1) biological clones from early and late stages of infection of three patients who never developed MT-2 cell syncytium inducing (SI; R5X4 or X4) viruses. Several of the late-stage non-MT-2 cell syncytium-inducing (NSI; R5) viruses from these patients depleted human CD4(+) thymocytes from SCID-hu mice. Earlier clones from the same patients did not deplete CD4(+) thymocytes from SCID-hu mice as well as later clones. We studied three R5 HIV-1 clones from patient ACH142 in greater detail. Two of these clones were obtained prior to the onset of AIDS; the third was obtained following the AIDS diagnosis. In GHOST cell infection assays, all three ACH142 R5 HIV-1 clones could infect GHOST cells expressing CCR5 but not GHOST cells expressing any of nine other HIV coreceptors tested. Furthermore, these patient clones efficiently infected stimulated peripheral blood mononuclear cells from a normal donor but not those from a homozygous CCR5Delta32 individual. Statistical analyses of data obtained from infection of SCID-hu mice with patient ACH142 R5 clones revealed that only the AIDS-associated clone significantly depleted CD4(+) thymocytes from SCID-hu mice. This clone also replicated to higher levels in SCID-hu mice than the two earlier clones, and a significant correlation between viral replication and CD4(+) thymocyte depletion was observed. Our results indicate that an intrinsic property of AIDS-associated R5 patient clones causes their increased replication and cytopathic effects in SCID-hu mice and likely contributes to the development of AIDS in patients who harbor only R5 quasispecies of HIV-1. PMID- 10708439 TI - Evolutionary relationships of endemic/epidemic and sylvatic dengue viruses. AB - Endemic/epidemic dengue viruses (DEN) that are transmitted among humans by the mosquito vectors Aedes aegypti and Aedes albopictus are hypothesized to have evolved from sylvatic DEN strains that are transmitted among nonhuman primates in West Africa and Malaysia by other Aedes mosquitoes. We tested this hypothesis with phylogenetic studies using envelope protein gene sequences of both endemic/epidemic and sylvatic strains. The basal position of sylvatic lineages of DEN-1, -2, and -4 suggested that the endemic/epidemic lineages of these three DEN serotypes evolved independently from sylvatic progenitors. Time estimates for evolution of the endemic/epidemic forms ranged from 100 to 1,500 years ago, and the evolution of endemic/epidemic forms represents relatively recent events in the history of DEN evolution. Analysis of envelope protein amino acid changes predicted to have accompanied endemic/epidemic emergence suggested a role for domain III in adaptation to new mosquito and/or human hosts. PMID- 10708440 TI - The BFRF1 gene of Epstein-Barr virus encodes a novel protein. AB - Computer analysis of the Epstein-Barr virus (EBV) genome indicates there are approximately 100 open reading frames (ORFs). Thus far about 30 EBV genes divided into the categories latent and lytic have been identified. The BamHI F region of EBV is abundantly transcribed during lytic replication. This region is highly conserved among herpesviruses, thus suggesting that some common function could be retained in the ORFs encompassed within this viral fragment. To identify putative novel proteins and possible new markers for viral replication, we focused our attention on the first rightward ORF in the BamHI F region (BFRF1). Histidine and glutathione S-transferase-tagged BFRF1 fusion proteins were synthesized to produce a mouse monoclonal antibody (MAb). Analysis of human sera revealed a high seroprevalence of antibodies to BFRF1 in patients affected by nasopharyngeal carcinoma or Burkitt's lymphoma, whereas no humoral response to BFRF1 could be detected among healthy donors. An anti-BFRF1 MAb recognizes a doublet migrating at 37 to 38 kDa in cells extracts from EBV-infected cell lines following lytic cycle activation and in an EBV-negative cell line (DG75) transfected with a plasmid expressing the BFRF1 gene. Northern blot analysis allowed the detection of a major transcript of 3.7 kb highly expressed in EBV-positive lytic cycle induced cell lines. Treatment with inhibitors of viral DNA polymerase, such as phosphonoacetic acid and acyclovir, reduced but did not abolish the transcription of BFRF1, thus indicating that BFRF1 can be classified as an early gene. Cell fractionation experiments, as well as immunolocalization by immunofluorescence microscopy, immunohistochemistry, and immunoelectron microscopy, showed that BFRF1 is localized on the plasma membrane and nuclear compartments of the cells and is a structural component of the viral particle. Identification of BFRF1 provides a new marker with which to monitor EBV infection and might help us better understand the biology of the virus. PMID- 10708441 TI - Asymmetric subunit organization of heterodimeric Rous sarcoma virus reverse transcriptase alphabeta: localization of the polymerase and RNase H active sites in the alpha subunit. AB - The genes encoding the alpha (63-kDa) and beta (95-kDa) subunits of Rous sarcoma virus (RSV) reverse transcriptase (RT) or the entire Pol polypeptide (99 kDa) were mutated in the conserved aspartic acid residue Asp 181 of the polymerase active site (YMDD) or in the conserved Asp 505 residue of the RNase H active site. We have analyzed heterodimeric recombinant RSV alphabeta and alphaPol RTs within which one subunit was selectively mutated. When alphabeta heterodimers contained the Asp 181-->Asn mutation in their beta subunits, about 42% of the wild-type polymerase activity was detected, whereas when the heterodimers contained the same mutation in their alpha subunits, only 7.5% of the wild-type polymerase activity was detected. Similar results were obtained when the conserved Asp 505 residue of the RNase H active site was mutated to Asn. RNase H activity was clearly detectable in alphabeta heterodimers mutated in the beta subunit but was lost when the mutation was present in the alpha subunit. In summary, our data imply that the polymerase and RNase H active sites are located in the alpha subunit of the heterodimeric RSV RT alphabeta. PMID- 10708442 TI - cis- and trans-acting elements in flavivirus RNA replication. AB - Most of the seven flavivirus nonstructural proteins (NS1 to NS5) encoded in the distal two-thirds of the RNA positive-sense genome are believed to be essential components of RNA replication complexes. To explore the functional relationships of these components in RNA replication, we used trans-complementation analysis of full-length infectious RNAs of Kunjin (KUN) virus with a range of lethal in-frame deletions in the nonstructural coding region, using as helper a repBHK cell line stably producing functional replication complexes from KUN replicon RNA. Recently we showed that replication of KUN RNAs with large carboxy-terminal deletions including the entire RNA polymerase region in the NS5 gene, representing 34 to 75% of the NS5 coding content, could be complemented after transfection into repBHK cells. In this study we have demonstrated that KUN RNAs with deletions of 84 to 97% of the NS1 gene, or of 13 to 63% of the NS3 gene including the entire helicase region, were also complemented in repBHK cells with variable efficiencies. In contrast, KUN RNAs with deletions in any of the other four nonstructural genes NS2A, NS2B, NS4A, and NS4B were not complemented. We have also demonstrated successful trans complementation of KUN RNAs containing either combined double deletions in the NS1 and NS5 genes or triple deletions in the NS1, NS3, and NS5 genes comprising as much as 38% of the entire nonstructural coding content. Based on these and our previous complementation results, we have generated a map of cis- and trans-acting elements in RNA replication for the nonstructural coding region of the flavivirus genome. These results are discussed in the context of our model on formation and composition of the flavivirus replication complex, and we suggest molecular mechanisms by which functions of some defective components of the replication complex can be complemented by their wild-type counterparts expressed from another (helper) RNA molecule. PMID- 10708443 TI - Evidence for budding of human immunodeficiency virus type 1 selectively from glycolipid-enriched membrane lipid rafts. AB - A number of recent studies have demonstrated the significance of detergent insoluble, glycolipid-enriched membrane domains or lipid rafts, especially in regard to activation and signaling in T lymphocytes. These domains can be viewed as floating rafts composed of sphingolipids and cholesterol which sequester glycosylphosphatidylinositol (GPI)-linked proteins, such as Thy-1 and CD59. CD45, a 200-kDa transmembrane phosphatase protein, is excluded from these domains. We have found that human immunodeficiency virus type 1 (HIV-1) particles produced by infected T-cell lines acquire the GPI-linked proteins Thy-1 and CD59, as well as the ganglioside GM1, which is known to partition preferentially into lipid rafts. In contrast, despite its high expression on the cell surface, CD45 was poorly incorporated into virus particles. Confocal fluorescence microscopy revealed that HIV-1 proteins colocalized with Thy-1, CD59, GM1, and a lipid raft-specific fluorescent lipid, DiIC(16)(3), in uropods of infected Jurkat cells. CD45 did not colocalize with HIV-1 proteins and was excluded from uropods. Dot immunoassay of Triton X-100-extracted membrane fractions revealed that HIV-1 p17 matrix protein and gp41 were present in the detergent-resistant fractions and that [(3)H]myristic acid-labeled HIV Gag showed a nine-to-one enrichment in lipid rafts. We propose a model for the budding of HIV virions through lipid rafts whereby host cell cholesterol, sphingolipids, and GPI-linked proteins within these domains are incorporated into the viral envelope, perhaps as a result of preferential sorting of HIV Gag to lipid rafts. PMID- 10708444 TI - The T-cell receptor zeta chain contains two homologous domains with which simian immunodeficiency virus Nef interacts and mediates down-modulation. AB - We have recently demonstrated that simian immunodeficiency virus (SIV) Nef binds to the zeta chain of the T-cell receptor (TCR), leading to its down-modulation from T-cell surfaces (I. Bell, C. Ashman, J. Maughan, E. Hooker, F. Cook, and T. A. Reinhart, J. Gen. Virol. 79:2717-2727, 1998). Using a panel of human as well as rhesus macaque TCR zeta cytoplasmic domain mutants, we have identified in this report two linear peptides in the cytoplasmic domain of TCR zeta which independently interact with SIV Nef. Each SIV Nef interaction domain was sufficient in the absence of the other for interaction with SIV Nef in a yeast two-hybrid assay. In parallel, we demonstrated that Nef down-modulation of CD8 TCR zeta fusion proteins containing full-length or truncated portions of the TCR zeta cytoplasmic domain occurs in transiently transfected 293T cells. Furthermore, using proteins expressed in Escherichia coli, a glutathione S transferase-Nef fusion protein coprecipitated histidine-tagged portions of the TCR zeta cytoplasmic domain which contained SNID-1 or SNID-2. The peptides targeted by SIV Nef, YNELNL and YSEIGMKGERRR, are portions of the first and second of three immunoreceptor tyrosine-based activation motifs which are important in signal transduction, thymocyte development, and TCR biogenesis. These results demonstrate that SIV Nef binds to two distinct domains on TCR zeta in the absence of other T-cell-specific factors, and that interaction with either domain is sufficient to cause down-modulation of TCR zeta. PMID- 10708445 TI - Virus-induced diabetes in a transgenic model: role of cross-reacting viruses and quantitation of effector T cells needed to cause disease. AB - Virus-specific cytotoxic T lymphocytes (CTL) at frequencies of >1/1, 000 are sufficient to cause insulin-dependent diabetes mellitus (IDDM) in transgenic mice whose pancreatic beta cells express as "self" antigen a protein from a virus later used to initiate infection. The inability to generate sufficient effector CTL for other cross-reacting viruses that fail to cause IDDM could be mapped to point mutations in the CTL epitope or its COO(-) flanking region. These data indicate that IDDM and likely other autoimmune diseases are caused by a quantifiable number of T cells, that neither standard epidemiologic markers nor molecular analysis with nucleic acid probes reliably distinguishes between viruses that do or do not cause diabetes, and that a single-amino-acid change flanking a CTL epitope can interfere with antigen presentation and development of autoimmune disease in vivo. PMID- 10708446 TI - The lymphocytic choriomeningitis virus RING protein Z associates with eukaryotic initiation factor 4E and selectively represses translation in a RING-dependent manner. AB - Only a few host cell proteins that associate with arenaviruses have been identified. To date, the arenavirus Z protein associates with the promyelocytic leukemia protein PML and the ribosomal P proteins. The majority of PML is present in nuclear bodies which are translocated to the cytoplasm by infection with the arenavirus, lymphocytic choriomeningitis virus (LCMV). The Z protein is a small zinc-binding RING protein with an unknown function which is required for the viral life cycle. Here, we demonstrate an association between Z and the host cell translation factor, eukaryotic initiation factor 4E (eIF-4E) in infected and transfected cells. Z's association with both ribosomal proteins and this translation factor led us to investigate whether Z could modulate host cell translation. In cell culture, Z selectively represses protein production in an eIF-4E-dependent manner. Specifically, we see reduction in cyclin D1 protein production with no effect on glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in cells transfected with Z. Previous reports indicate that cyclin D1 is sensitive to eIF-4E levels, whereas GAPDH is not. Consistent with this, we observe preferential downregulation of cyclin D1 during infection and no effect on GAPDH. Further, no changes in RNA levels were observed for cyclin D1 or GAPDH transcripts. The interaction between eIF-4E and Z may provide a mechanism for slower growth observed in infected cells and a viral strategy for establishing chronic infection. PMID- 10708447 TI - Distinctions between bovine herpesvirus 1 and herpes simplex virus type 1 VP22 tegument protein subcellular associations. AB - The alphaherpesvirus tegument protein VP22 has been characterized with multiple traits including microtubule reorganization, nuclear localization, and nonclassical intercellular trafficking. However, all these data were derived from studies using herpes simplex virus type 1 (HSV-1) and may not apply to VP22 homologs of other alphaherpesviruses. We compared subcellular attributes of HSV-1 VP22 (HVP22) with bovine herpesvirus 1 (BHV-1) VP22 (BVP22) using green fluorescent protein (GFP)-fused VP22 expression vectors. Fluorescence microscopy of cell lines transfected with these constructs revealed differences as well as similarities between the two VP22 homologs. Compared to that of HVP22, the BVP22 microtubule interaction was much less pronounced. The VP22 nuclear interaction varied, with a marbled or halo appearance for BVP22 and a speckled or nucleolus bound appearance for HVP22. Both VP22 homologs associated with chromatin at various stages of mitosis and could traffic from expressing cells to the nuclei of nonexpressing cells. However, distinct qualitative differences in microtubule, nuclear, and chromatin association as well as trafficking were observed. The differences in VP22 homolog characteristics revealed in this study will help define VP22 function within HSV-1 and BHV-1 infection. PMID- 10708448 TI - Rotavirus spike protein VP4 is present at the plasma membrane and is associated with microtubules in infected cells. AB - VP4 is an unglycosylated protein of the outer layer of the capsid of rotavirus. It forms spikes that project from the outer layer of mature virions, which is mainly constituted by glycoprotein VP7. VP4 has been implicated in several important functions, such as cell attachment, penetration, hemagglutination, neutralization, virulence, and host range. Previous studies indicated that VP4 is located in the space between the periphery of the viroplasm and the outside of the endoplasmic reticulum in rotavirus-infected cells. Confocal microscopy of infected MA104 monolayers, immunostained with specific monoclonal antibodies, revealed that a significant fraction of VP4 was present at the plasma membrane early after infection. Another fraction of VP4 is cytoplasmic and colocalizes with beta-tubulin. Flow cytometry analysis confirmed that at the early stage of viral infection, VP4 was present on the plasma membrane and that its N-terminal region, the VP8* subunit, was accessible to antibodies. Biotin labeling of the infected cell surface monolayer with a cell-impermeable reagent allowed the identification of the noncleaved form of VP4 that was associated with the glycoprotein VP7. The localization of VP4 was not modified in cells transfected with a plasmid allowing the expression of a fusion protein consisting of VP4 and the green fluorescent protein. The present data suggest that VP4 reaches the plasma membrane through the microtubule network and that other viral proteins are dispensable for its targeting and transport. PMID- 10708449 TI - An envelope glycoprotein of the human endogenous retrovirus HERV-W is expressed in the human placenta and fuses cells expressing the type D mammalian retrovirus receptor. AB - A new human endogenous retrovirus (HERV) family, termed HERV-W, was recently described (J.-L. Blond, F. Beseme, L. Duret, O. Bouton, F. Bedin, H. Perron, B. Mandrand, and F. Mallet, J. Virol. 73:1175-1185, 1999). HERV-W mRNAs were found to be specifically expressed in placenta cells, and an env cDNA containing a complete open reading frame was recovered. In cell-cell fusion assays, we demonstrate here that the product of the HERV-W env gene is a highly fusogenic membrane glycoprotein. Transfection of an HERV-W Env expression vector in a panel of cell lines derived from different species resulted in formation of syncytia in primate and pig cells upon interaction with the type D mammalian retrovirus receptor. Moreover, envelope glycoproteins encoded by HERV-W were specifically detected in placenta cells, suggesting that they may play a physiological role during pregnancy and placenta formation. PMID- 10708450 TI - Degradation of tobacco mosaic virus movement protein by the 26S proteasome. AB - Cell-to-cell spread of tobacco mosaic virus is facilitated by the virus-encoded 30-kDa movement protein (MP). This process involves interaction of viral proteins with host components, including the cytoskeleton and the endoplasmic reticulum (ER). During virus infection, high-molecular-weight forms of MP were detected in tobacco BY-2 protoplasts. Inhibition of the 26S proteasome by MG115 and clasto lactacystin-beta-lactone enhanced the accumulation of high-molecular-weight forms of MP and led to increased stability of the MP. Such treatment also increased the apparent accumulation of polyubiquitinated host proteins. By fusion of MP with the jellyfish green fluorescent protein (GFP), we demonstrated that inhibition of the 26S proteasome led to accumulation of the MP-GFP fusion preferentially on the ER, particularly the perinuclear ER. We suggest that polyubiquitination of MP and subsequent degradation by the 26S proteasome may play a substantial role in regulation of virus spread by reducing the damage caused by the MP on the structure of cortical ER. PMID- 10708451 TI - Tumor necrosis factor alpha-deficient, but not interleukin-6-deficient, mice resist peripheral infection with scrapie. AB - In most peripheral infections of rodents and sheep with scrapie, infectivity is found first in lymphoid tissues and later in the central nervous system (CNS). Cells within the germinal centers (GCs) of the spleen and lymph nodes are important sites of extraneural replication, from which infection is likely to spread to the CNS along peripheral nerves. Here, using immunodeficient mice, we investigate the identity of the cells in the spleen that are important for disease propagation. Despite possessing functional T and B lymphocytes, tumor necrosis factor alpha-deficient (TNF-alpha(-/-)) mice lack GCs and follicular dendritic cell (FDC) networks in lymphoid tissues. In contrast, lymphoid tissues of interleukin-6-deficient (IL-6(-/-)) mice possess FDC networks but have impaired GCs. When the CNSs of TNF-alpha(-/-), IL-6(-/-), and wild-type mice were directly challenged with the ME7 scrapie strain, 100% of the mice were susceptible, developing disease after closely similar incubation periods. However, when challenged peripherally (intraperitoneally), most TNF-alpha(-/-) mice failed to develop scrapie up to 503 days postinjection. All wild-type and IL 6(-/-) mice succumbed to disease approximately 300 days after the peripheral challenge. High levels of scrapie infection and the disease-specific isomer of the prion protein, PrP(Sc), were detectable in spleens from challenged wild-type and IL-6(-/-) mice but not from TNF-alpha(-/-) mice. Histopathological analysis of spleen tissue demonstrated heavy PrP accumulations in direct association with FDCs in challenged wild-type and IL-6(-/-) mice. No PrP(Sc) accumulation was detected in spleens from TNF-alpha(-/-) mice. We conclude that, for the ME7 scrapie strain, mature FDCs are critical for replication in lymphoid tissues and that in their absence, neuroinvasion following peripheral challenge is impaired. PMID- 10708452 TI - Readministration of adenovirus vector in nonhuman primate lungs by blockade of CD40-CD40 ligand interactions. AB - The interaction between CD40 on B cells and CD40 ligand (CD40L) on activated T cells is important for B-cell differentiation in T-cell-dependent humoral responses. We have extended our previous murine studies of CD40-CD40L in adenoviral vector-mediated immune responses to rhesus monkeys. Primary immune responses to adenoviral vectors and the ability to readminister vector were studied in rhesus monkeys in the presence or absence of a transient treatment with a humanized anti-CD40 ligand antibody (hu5C8). Adult animals were treated with hu5C8 at the time vector was instilled into the lung. Immunological analyses demonstrated suppression of adenovirus-induced lymphoproliferation and cytokine responses (interleukin-2 [IL-2], gamma interferon, IL-4, and IL-10) in hu5C8 treated animals. Animals treated with hu5C8 secreted adenovirus-specific immunoglobulin M (IgM) levels comparable to control animals, but did not secrete IgA or develop neutralizing antibodies; consequently, the animals could be readministered with adenovirus vector expressing alkaline phosphatase. A second study was designed to examine the long-term effects on immune functions of a short course of hu5C8. Acute hu5C8 treatment resulted in significant and prolonged inhibition of the adenovirus-specific humoral response well beyond the time hu5C8 effects were no longer significant. These studies demonstrate the potential of hu5C8 as an immunomodulatory regimen to enable administration of adenoviral vectors, and they advocate testing this model in humans. PMID- 10708453 TI - Vaccinia virus envelope H3L protein binds to cell surface heparan sulfate and is important for intracellular mature virion morphogenesis and virus infection in vitro and in vivo. AB - An immunodominant antigen, p35, is expressed on the envelope of intracellular mature virions (IMV) of vaccinia virus. p35 is encoded by the viral late gene H3L, but its role in the virus life cycle is not known. This report demonstrates that soluble H3L protein binds to heparan sulfate on the cell surface and competes with the binding of vaccinia virus, indicating a role for H3L protein in IMV adsorption to mammalian cells. A mutant virus defective in expression of H3L (H3L(-)) was constructed; the mutant virus has a small plaque phenotype and 10 fold lower IMV and extracellular enveloped virion titers than the wild-type virus. Virion morphogenesis is severely blocked and intermediate viral structures such as viral factories and crescents accumulate in cells infected with the H3L( ) mutant virus. IMV from the H3L(-) mutant virus are somewhat altered and less infectious than wild-type virions. However, cells infected by the mutant virus form multinucleated syncytia after low pH treatment, suggesting that H3L protein is not required for cell fusion. Mice inoculated intranasally with wild-type virus show high mortality and severe weight loss, whereas mice infected with H3L( ) mutant virus survive and recover faster, indicating that inactivation of the H3L gene attenuates virus virulence in vivo. In summary, these data indicate that H3L protein mediates vaccinia virus adsorption to cell surface heparan sulfate and is important for vaccinia virus infection in vitro and in vivo. In addition, H3L protein plays a role in virion assembly. PMID- 10708454 TI - Alpha/beta interferon protects adult mice from fatal Sindbis virus infection and is an important determinant of cell and tissue tropism. AB - Infection of adult 129 Sv/Ev mice with consensus Sindbis virus strain TR339 is subclinical due to an inherent restriction in early virus replication and viremic dissemination. By comparing the pathogenesis of TR339 in 129 Sv/Ev mice and alpha/beta interferon receptor null (IFN-alpha/betaR(-/-)) mice, we have assessed the contribution of IFN-alpha/beta in restricting virus replication and spread and in determining cell and tissue tropism. In adult 129 Sv/Ev mice, subcutaneous inoculation with 100 PFU of TR339 led to extremely low-level virus replication and viremia, with clearance under way by 96 h postinoculation (p.i.). In striking contrast, adult IFN-alpha/betaR(-/-) mice inoculated subcutaneously with 100 PFU of TR339 succumbed to the infection within 84 h. By 24 h p.i. a high-titer serum viremia had seeded infectious virus systemically, coincident with the systemic induction of the proinflammatory cytokines interleukin-12 (IL-12) p40, IFN-gamma, tumor necrosis factor alpha, and IL-6. Replicating virus was located in macrophage-dendritic cell (DC)-like cells at 24 h p.i. in the draining lymph node and in the splenic marginal zone. By 72 h p.i. virus replication was widespread in macrophage-DC-like cells in the spleen, liver, lung, thymus, and kidney and in fibroblast-connective tissue and periosteum, with sporadic neuroinvasion. IFN alpha/beta-mediated restriction of TR339 infection was mimicked in vitro in peritoneal exudate cells from 129 Sv/Ev versus IFN-alpha/betaR(-/-) mice. Thus, IFN-alpha/beta protects the normal adult host from viral infection by rapidly conferring an antiviral state on otherwise permissive cell types, both locally and systemically. Ablation of the IFN-alpha/beta system alters the apparent cell and tissue tropism of the virus and renders macrophage-DC-lineage cells permissive to infection. PMID- 10708455 TI - Four proteins processed from the replicase gene polyprotein of mouse hepatitis virus colocalize in the cell periphery and adjacent to sites of virion assembly. AB - The replicase gene (gene 1) of the coronavirus mouse hepatitis virus (MHV) encodes two co-amino-terminal polyproteins presumed to incorporate all the virus encoded proteins necessary for viral RNA synthesis. The polyproteins are cotranslationally processed by viral proteinases into at least 15 mature proteins, including four predicted cleavage products of less than 25 kDa that together would comprise the final 59 kDa of protein translated from open reading frame 1a. Monospecific antibodies directed against the four distinct domains detected proteins of 10, 12, and 15 kDa (p1a-10, p1a-12, and p1a-15) in MHV-A59 infected DBT cells, in addition to a previously identified 22-kDa protein (p1a 22). When infected cells were probed by immunofluorescence laser confocal microscopy, p1a-10, -22, -12, and -15 were detected in discrete foci that were prominent in the perinuclear region but were widely distributed throughout the cytoplasm as well. Dual-labeling experiments demonstrated colocalization of the majority of p1a-22 in replication complexes with the helicase, nucleocapsid, and 3C-like proteinase, as well as with p1a-10, -12, and -15. p1a-22 was also detected in separate foci adjacent to the replication complexes. The majority of complexes containing the gene 1 proteins were distinct from sites of accumulation of the M assembly protein. However, in perinuclear regions the gene 1 proteins and nucleocapsid were intercalated with sites of M protein localization. These results demonstrate that the complexes known to be involved in RNA synthesis contain multiple gene 1 proteins and are closely associated with structural proteins at presumed sites of virion assembly. PMID- 10708456 TI - The primary sequence of rhesus monkey rhadinovirus isolate 26-95: sequence similarities to Kaposi's sarcoma-associated herpesvirus and rhesus monkey rhadinovirus isolate 17577. AB - The primary sequence of the long unique region L-DNA (L for low GC) of rhesus monkey rhadinovirus (RRV) isolate 26-95 was determined. The L-DNA consists of 130,733 bp that contain 84 open reading frames (ORFs). The overall organization of the RRV26-95 genome was found to be very similar to that of human Kaposi sarcoma-associated herpesvirus (KSHV). BLAST search analysis revealed that in almost all cases RRV26-95 coding sequences have a greater degree of similarity to corresponding KSHV sequences than to other herpesviruses. All of the ORFs present in KSHV have at least one homologue in RRV26-95 except K3 and K5 (bovine herpesvirus-4 immediate-early protein homologues), K7 (nut-1), and K12 (Kaposin). RRV26-95 contains one MIP-1 and eight interferon regulatory factor (vIRF) homologues compared to three MIP-1 and four vIRF homologues in KSHV. All homologues are correspondingly located in KSHV and RRV with the exception of dihydrofolate reductase (DHFR). DHFR is correspondingly located near the left end of the genome in RRV26-95 and herpesvirus saimiri (HVS), but in KSHV the DHFR gene is displaced 16,069 nucleotides in a rightward direction in the genome. DHFR is also unusual in that the RRV26-95 DHFR more closely resembles HVS DHFR (74% similarity) than KSHV DHFR (55% similarity). Of the 84 ORFs in RRV26-95, 83 contain sequences similar to the recently determined sequences of the independent RRV isolate 17577. RRV26-95 and RRV17577 sequences differ in that ORF 67.5 sequences contained in RRV26-95 were not found in RRV17577. In addition, ORF 4 is significantly shorter in RRV26-95 than was reported for RRV17577 (395 versus 645 amino acids). Only four of the corresponding ORFs between RRV26-95 and RRV17577 exhibited less than 95% sequence identity: glycoproteins H and L, uracil DNA glucosidase, and a tegument protein (ORF 67). Both RRV26-95 and RRV17577 have unique ORFs between positions 21444 to 21752 and 110910 to 114899 in a rightward direction and from positions 116524 to 111082 in a leftward direction that are not found in KSHV. Our analysis indicates that RRV26-95 and RRV17577 are clearly independent isolates of the same virus species and that both are closely related in structural organization and overall sequence to KSHV. The availability of detailed sequence information, the ability to grow RRV lytically in cell culture, and the ability to infect monkeys experimentally with RRV will facilitate the construction of mutant strains of virus for evaluating the contribution of individual genes to biological properties. PMID- 10708457 TI - Virus clearance through apoptosis-dependent phagocytosis of influenza A virus infected cells by macrophages. AB - Some cultured cell lines undergo typical apoptosis upon infection with influenza virus. However, the release of replicated virus into the culture medium does not change when apoptosis is inhibited. Since apoptotic cells are heterophagically eliminated at early stages of the apoptosis pathway, we anticipated that the coexistence of phagocytic cells with virus-infected cells affects the extent of virus growth. When influenza A virus-infected HeLa cells were mixed with activated mouse peritoneal macrophages, efficient phagocytosis, which was abrogated in the presence of a caspase inhibitor, occurred. At the same time, the release of virus into the culture medium was completely inhibited, and this required direct contact between virus-infected cells and macrophages. Furthermore, an immunoelectron microscopic analysis detected influenza virus particles associated with phagosome-like structures within macrophages. These results indicate that apoptosis-dependent phagocytosis of virus-infected cells may lead to direct elimination of the pathogen. PMID- 10708458 TI - Impaired antiviral response and alpha/beta interferon induction in mice lacking beta interferon. AB - We have generated mice lacking the gene for beta interferon and report that they are highly susceptible to vaccinia virus infection. Furthermore, in cultured embryo fibroblasts, viral induction of alpha interferon and of 2-5A synthetase genes is impaired. We also show that beta interferon does not prime its own expression. PMID- 10708459 TI - Fatty acid-depleted albumin induces the formation of echovirus A particles. AB - Picornavirus infection requires virus uncoating, associated with the production of 135S "A" particles and 80S empty particles from 160S mature virions, to release the RNA genome into the cell cytoplasm. Normal albumin inhibits this process. We now show that when depleted of fatty acids, albumin induces the formation of echovirus A particles. PMID- 10708460 TI - Characterization of the P protein-binding domain on the 10-kilodalton protein of Borna disease virus. AB - The Borna disease virus (BDV) is the prototype member of the Bornaviridae, and it replicates in the cell nucleus. The BDV p24P and p40N proteins carry nuclear localization signals (NLS) and are found in the nuclei of infected cells. The BDV p10 protein does not have an NLS, but it binds with P and/or N and is translocated to the nucleus. Hence, p10 may play a role in the replication of BDV in the cell nucleus. Here, we show that the P-binding domain is located in the N terminus of p10 and that S(3) and L(16) are important for the interaction. PMID- 10708461 TI - Assembly and processing of human immunodeficiency virus Gag mutants containing a partial replacement of the matrix domain by the viral protease domain. AB - We constructed human immunodeficiency virus (HIV) mutants by replacing the matrix domain with sequences encoding the viral protease or p6* and protease. The chimeras retaining matrix myristylation and processing signals underwent efficient autoprocessing with severely defective particle budding. The budding defects of the chimeras were rescued by suppressing the chimera protease activity either through addition of an HIV protease inhibitor or through inactivating the chimera protease via a substitution mutation of the catalytic aspartic acid residue. This resulted in the release of chimeric virus-like particles with the density of a wild-type retrovirus particle. In addition, the assembly-competent but processing-defective chimeras produced proteolytically processed particles with significant reverse transcriptase activity when a downstream native pol gene was present. These results suggest that HIV has the potential to adapt heterologous sequences in place of the matrix sequence without major effects on virus-like particle budding. In addition, the positions of the protease and substrate accessibility may contribute significantly toward avoiding a premature Gag or Gag-Pol process, which leads to severe defects in both particle budding and incorporation. PMID- 10708462 TI - Uncoating kinetics of hepatitis A virus virions and provirions. AB - When the growth kinetics of immature hepatitis A virus provirions and mature virions were monitored, distinct eclipse phases were noted for both types of particles. Strikingly, uncoating of virions occurred around 4 h postinfection, while uncoating of provirions occurred predominantly between 8 and 10 h postinfection. It is proposed that the heterogeneous mixture of infectious hepatitis A virus particles (virions and provirions) typically present in inocula is responsible for the normally asynchronous nature of hepatitis A virus uncoating kinetics. PMID- 10708464 TI - Abnormal uterine bleeding in the reproductive years. Part II-medical management. PMID- 10708463 TI - Modulation of antigen-specific humoral responses in rhesus macaques by using cytokine cDNAs as DNA vaccine adjuvants. AB - An important limitation of DNA immunization in nonhuman primates is the difficulty in generating high levels of antigen-specific antibody responses; strategies to enhance the level of immune responses to DNA immunization may be important in the further development of this vaccine strategy for humans. We approached this issue by testing the ability of molecular adjuvants to enhance the levels of immune responses generated by multicomponent DNA vaccines in rhesus macaques. Rhesus macaques were coimmunized intramuscularly with expression plasmids bearing genes encoding Th1 (interleukin 2 [IL-2] and gamma interferon)- or Th2 (IL-4)-type cytokines and DNA vaccine constructs encoding human immunodeficiency virus Env and Rev and simian immunodeficiency virus Gag and Pol proteins. We observed that the cytokine gene adjuvants (especially IL-2 and IL-4) significantly enhanced antigen-specific humoral immune responses in the rhesus macaque model. These results support the assumption that antigen-specific responses can be engineered to a higher and presumably more desirable level in rhesus macaques by genetic adjuvants. PMID- 10708465 TI - Real-time 3-dimensional echocardiography evaluation of congenital heart disease. AB - We evaluated the ability of real-time 3-dimensional (RT3D) echocardiography to diagnose congenital heart defects and its potential for presenting structural abnormalities in novel views. Seventy-five patients with suspected congenital heart defects were examined with the use of RT3D echocardiography. Images were reviewed off-line as multiple slices of the raw data or as volume-rendered images by a blinded observer. Diagnoses made from blinded review of the RT3D images were compared with the clinical report of the 2D echocardiogram obtained at the same visit. Real-time 3D echocardiography identified all structural abnormalities except for small atrial septal defects in 2 patients and coronary artery anatomy in D-transposition of the great arteries. Less than 5 minutes were needed to acquire RT3D images in all cases, and sedation was never required. Unique region oriented views obtained from the 3D data set can be acquired quickly and have the potential to enhance understanding of complex cardiac anatomy. PMID- 10708466 TI - Reconstruction of 3-dimensional right ventricular shape and volume from 3 orthogonal planes. AB - This study validates a reconstructive technique that describes 3-dimensional right ventricular (RV) shape and volume with the use of 3 standard echocardiographic planes. The volume of 24 cast models of lamb right ventricles (12 normal, 12 hypertensive) was determined by water displacement. Reconstruction of the cast shapes was calculated from 2 sets of digitized data: cast cross sectional digitized tracings and echocardiographic cross-sectional tracings. Regional volume ratios from both data sets were assessed to quantitatively specify RV regional volume differences between normotensive and hypertensive right ventricles. This method described the 3-dimensional RV shape with no differences between reconstructed volumes and true volumes for either normotensive or hypertensive casts. Between hypertensive and normal groups, regional volume ratios yielded a difference in free wall ratios that was observed to be greater in the hypertensive cast group (P =.007). PMID- 10708467 TI - A new quantitative method for the diagnosis of right ventricular hypertensive disorders in 3 dimensions. AB - A new 3-dimensional (3D) method is described for the diagnosis of normal and hypertensive right ventricular (RV) conditions on the basis of similarity of RV structure to models of normal average shape or hypertensive average shape. Right ventricular quantification in multiple views (coronal, sagittal, and transverse) was obtained by measuring tangent angle differences (TADs) between RV tracings and average shapes at 128 points around the ventricular contour in each view. The TAD measurements of all views were then combined to quantify the closest 3D fit of the ventricle to a normal or hypertensive model. RESULTS: In 24 lamb casts measured in vitro, an accurate diagnosis was obtained in 11 of 12 normotensive casts (specificity 92%) and 11 of 12 hypertensive casts (sensitivity 92%). CONCLUSION: Accurate 3D diagnosis of in vitro normotensive and hypertensive RV conditions can be realized by measuring the TADs between the ventricle and average-shaped models. PMID- 10708468 TI - Normal regional right ventricular function and its change with age: a Doppler myocardial imaging study. AB - BACKGROUND: Doppler Myocardial Imaging (DMI) is a new technique currently being studied for the assessment of regional systolic and diastolic left ventricular (LV) function. No normal values or data on age-related changes in regional myocardial right ventricular (RV) velocities are available. METHODS AND RESULTS: Color DMI was used in 32 healthy volunteers (aged 16-76 years) to derive regional velocities from basal, medial, and apical segments of the RV free wall in the apical 4-chamber view, and from distal segments as well as from the tricuspid annulus in the parasternal long-axis view. Both mitral annular and regional LV velocities (4-chamber, long-axis parasternal view) were also recorded and compared with corresponding RV regional velocities. The M-mode displacement of the cardiac base was measured. Corresponding RV and LV DMI data sets were compared. For longitudinal function, RV free wall systolic velocities were consistently higher than velocities recorded in corresponding LV segments (analysis of variance, P <.05). Older subjects (40-76 years; 13 men, 2 women) had lower RV long-axis regional velocities than younger subjects (16-39 years; 15 men, 2 women), but had higher short-axis RV systolic velocities. For diastolic velocities, a negative correlation between age and the ratio of regional early diastolic to late diastolic velocity was shown for all RV free wall segments (eg, basal segment: r = -0.63, P <.0001). CONCLUSIONS: The right ventricle has higher long-axis regional velocities, a greater excursion of its lateral atrioventricular valve ring, and reduced circumferential shortening velocities compared with the left ventricle. Right ventricular longitudinal shortening is dominant over short-axis function in healthy young subjects. Normal age-related changes of diastolic velocities for each segment of the normal RV free wall have been defined. PMID- 10708469 TI - Transesophageal 3-dimensional echocardiography: in vivo determination of left ventricular mass in comparison with magnetic resonance imaging. AB - The objective of this study was to assess the accuracy and reproducibility of transesophageal 3-dimensional echocardiography (3DE) in comparison with magnetic resonance imaging (MRI) for the in vivo calculation of left ventricular mass (LVM). In addition, mass values obtained by M-mode echocardiography were compared with those calculated by MRI. Three-dimensional reconstruction of the left ventricle was performed from a transesophageal and transgastric transducer position with a multiplane transducer in 20 patients. Left ventricular mass was calculated from both transducer positions by using slices of various thicknesses, ranging from 5 to 20 mm. Reproducibility was determined by 5 repeated measurements of mass in each of 5 randomly selected left ventricles. M-mode echocardiography was performed according to the method described by Devereux. For MRI, multiple short-axis views with 10-mm slice thickness were acquired in inspiration hold. Correlation was high for mass determined by 3DE and MRI (for 10 mm slice thickness: r = 0.99; y = 0.99 x - 0.7 g; standard error of estimate = 8.5 g; P <.001). There was no statistical bias, and the limits of agreement ranged from +/-16.4 g to +/-27.2 g, depending on the slice thickness. Variability was lowest for a slice thickness of 10 mm (SD +/- 8.2 g). The reproducibility of mass determination was excellent (mean width of the 95% CI 12.8 g). Left ventricular mass values calculated from the transgastric and transesophageal transducer position were not different from each other (mean bias 0.6 +/- 9.1 g; P = ns). M-mode-based LVM calculations showed systematic overestimation and large measurement variability (bias 23.7 g; 95% CI +/- 92.8 g). Compared with MRI, transesophageal 3DE is an accurate and reproducible method for the determination of LVM and clearly superior to M-mode echocardiography. PMID- 10708470 TI - Feasibility and accuracy of left ventricular volumes and ejection fraction determination by fundamental, tissue harmonic, and intravenous contrast imaging in difficult-to-image patients. AB - The need to enhance the echocardiographic determination of left ventricular ejection fraction is greatest in patients with suboptimal images. Intravenous contrast (CON) and tissue harmonic imaging (THI) are 2 important methods for enhancing endocardial border definition. However, the comparative feasibility and accuracy of THI and contrast-enhanced power harmonic imaging in difficult-to image patients have not been examined. We assessed the comparative accuracy of THI and CON in determining EF and ventricular volumes in patients with suboptimal fundamental images. We demonstrated that CON is feasible and exhibits a greater correlation with ejection fraction and ventricular volumes determined by radionuclide angiography (standard of comparison) than THI in this difficult-to image population, with no reported side effects. For both ejection fraction and ventricular volumes, the observer variability was least for CON, intermediate with THI, and greatest for fundamental imaging. PMID- 10708471 TI - Diagnosis of a left coronary artery to right ventricular fistula with progression to spontaneous closure. AB - Coronary artery fistulas in structurally normal hearts are rare. The natural history of these lesions depends on their size and can cause congestive heart failure, infective endocarditis, ischemia, or accelerated atherosclerosis. These fistulas are usually closed either in the catheterization laboratory or surgically. This case demonstrates the prenatal diagnosis of a left coronary to right ventricular fistula and documents its natural history to spontaneous closure by 1 year of age. This may help confirm the rationale of observation rather than closure of small fistulas in selected cases of patients without symptoms. PMID- 10708472 TI - Intra-aortic balloon pump associated with dynamic left ventricular outflow tract obstruction after valve replacement for aortic stenosis. AB - An unstable patient with critical aortic stenosis had an intra-aortic balloon pump placed preoperatively for hemodynamic support and alleviation of symptoms. After separation from cardiopulmonary bypass following aortic valve replacement, the patient was hypotensive with increased pulmonary artery pressures. Transesophageal echocardiography revealed left ventricular outflow tract obstruction associated with systolic anterior motion of the mitral valve and severe mitral regurgitation. This pathophysiology was present when ventricular systole was preceded by balloon counterpulsation, but was absent during unassisted systole. This case report demonstrates a potentially significant untoward effect of intra-aortic balloon pump augmentation after aortic valve replacement for aortic stenosis. The timely diagnosis of this iatrogenic condition in the operating room permitted the prompt implementation of appropriate management strategies and avoided unnecessary surgical intervention. PMID- 10708473 TI - The echocardiographic diagnosis, characterization, and extraction guidance of cardiac foreign bodies. AB - Echocardiography is ideal for localizing cardiac foreign bodies and for characterizing associated cardiac and vascular injury before and during extraction. We report 5 cases of traumatic and iatrogenic cardiac foreign bodies that illustrate the central role of transthoracic and transesophageal ultrasonography in the management of these patients. PMID- 10708474 TI - Incorporating ultrasound contrast in the laboratory: a series on contrast echocardiography, article 1. PMID- 10708475 TI - Echocardiographic features of genetic diseases: part 3. Shunts. PMID- 10708477 TI - An approach to proteomic analysis of human tumors. AB - A strategy for proteomic analysis of microdissected cells derived from human tumor specimens is described and demonstrated by using esophageal cancer as an example. Normal squamous epithelium and corresponding tumor cells from two patients were procured by laser-capture microdissection and studied by two dimensional polyacrylamide gel electrophoresis (2D-PAGE). Fifty thousand cells resolved approximately 675 distinct proteins (or isoforms) with molecular weights ranging between 10 and 200 kDa and isoelectric points of pH 3-10. Comparison of the microdissected protein profiles showed a high degree of similarity between the matched normal-tumor samples (98% identical). However, 17 proteins showed tumor-specific alterations, including 10 that were uniquely present in the tumors and seven that were observed only in the normal epithelium. Two of the altered proteins were characterized by mass spectrometry and immunoblot analysis and were identified as cytokeratin 1 and annexin I. Acquisition of 2D-PAGE protein profiles, visualization of disregulated proteins, and subsequent determination of the identity of selected proteins through high-sensitivity MS-MS microsequencing are possible from microdissected cell populations. These separation and analytical techniques are uniquely capable of detecting tumor-specific alterations. Continued refinement of techniques and methodologies to determine the abundance and status of proteins in vivo holds great promise for future study of normal cells and associated neoplasms. Mol. PMID- 10708476 TI - The paradox of E2F1: oncogene and tumor suppressor gene. AB - Cancer cells often contain mutations that lead to the loss of retinoblastoma tumor suppressor (Rb) function and the activation of E2F-dependent transcription. As a result, proliferation is deregulated, and sensitivity to apoptotic stimuli is increased. In cell culture studies, the transcription factor E2F1 has been shown to be equally adept at inducing proliferation and apoptosis. Several groups using mouse models have been examining how these E2F1-regulated processes impact the development of cancer. The conclusion from these studies is that E2F1 can function as both oncogene and tumor suppressor gene and that both positive and negative effects on tumorigenesis can be observed whether E2F1 is absent or overexpressed. These findings are discussed in the context of a model in which pathways controlling cell-cycle progression and apoptosis are intimately linked. PMID- 10708478 TI - The protein kinase C beta-specific inhibitor LY379196 blocks TPA-induced monocytic differentiation of HL60 cells the protein kinase C beta-specific inhibitor LY379196 blocks TPA-induced monocytic differentiation of HL60 cells. AB - The ability of the promyelocytic leukemia HL60 cell line to differentiate in response to various stimuli has provided a widely used model of differentiation. The phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA), acting via its cellular receptor protein kinase C(PKC), induces these cells to acquire a monocytic phenotype. We set out to identify the specific isoform of the multigene PKC family that is involved in this differentiation event. To do so, we utilized a highly specific PKCbeta inhibitor, LY379196. We found that LY379196 could prevent the growth arrest, cellular adherence, and changes in several marker proteins that occur after the addition of TPA to HL60 cells and that these effects were not simply due to nonspecific cytotoxicity. Thus, the present studies provide strong evidence that the beta isoform of PKC plays a critical role in TPA-induced HL60 monocytic differentiation. PMID- 10708479 TI - Molecular cloning of human Hic-5, a potential regulator involved in signal transduction and cellular senescence. AB - By using differential display, we cloned the human counterpart of the murine gene Hic-5 from senescent human keratinocytes. The full-length cDNA contained a short GC-stretch proceeding a consensus Kozak sequence followed by a single open reading frame of 1338 bp encoding a 461-amino acid protein with a predicted molecular weight of 50 kDa. The expression of this gene was prominent in cells of epithelial origin but low or absent in lymphoid tissues and hematopoietic cells. The deduced protein contained four LIM domains at the carboxyl-terminal end and four LD motifs at the amino-terminal half, sharing high similarities with the focal adhesion protein paxillin. Hic-5 may therefore function, like paxillin, as a potential adapter for the recruitment of structural and signaling molecules to certain subcellular sites or in focal adhesions. Isolation of the genomic sequence revealed that the gene covered a segment of 6 kb and spanned 11 exons from the translation initiation site ATG to the termination signal TGA. Fluorescent in situ hybridization by using a human Hic-5 specific probe localized the gene to human chromosome 16p11. PMID- 10708480 TI - Activation of the MN/CA9 gene is associated with hypomethylation in human renal cell carcinoma cell lines. AB - The MN/CA9 (G250) gene expressed in the normal alimentary tract in a tissue specific manner is often activated in renal cell carcinomas. To cast light on the activation mechanism, we examined the methylation status of this gene in seven human renal cell carcinoma cell lines (SKRC-01, -06, -10, -12, -14, -44, and -59) and three normal kidney tissue samples by using the bisulfite genomic sequencing protocol. CpG methylation was measured at seven locations in the MN/CA9 5' region. MN/CA9 transcripts were detected by reverse transcription-polymerase chain reaction in five of the renal cell carcinoma cell lines (SKRC-01, -06, -10, -44, and -59). These MN/CA9 positive cell lines showed hypomethylation, whereas the remaining two cell lines (SKRC-12, and -14), and three normal kidney tissue samples without transcripts demonstrated hypermethylation. Treatment with the demethylating agent 5-aza-2'-deoxycytidine resulted in activation of the MN/CA9 gene in the negative cell lines (SKRC-12 and -14). These data suggest that hypomethylation in the 5' region may have a major role in expression of the MN/CA9 gene in renal cell carcinoma cells. PMID- 10708481 TI - The carcinogen 2-acetylaminofluorene inhibits activation and nuclear accumulation of cyclin-dependent kinase 2 in growth-induced rat liver. AB - Growth arrest in G(1) is a common cellular response to DNA damage. In the present study, liver regeneration was combined with continuous exposure for 2 acetylaminofluorene (AAF) to study mechanisms of carcinogen-induced growth arrest in vivo. Growth arrest of uninitiated hepatocytes is central for AAF-induced promotion of premalignant lesions in rat liver. To characterize this growth arrest, we examined the activity of cyclin-dependent kinase (Cdk) 2 in unexposed liver and in AAF-exposed liver after growth induction by partial hepatectomy (PH). Rats were fed either a control diet or an AAF-supplemented diet. After 7 d, a two-third PH was performed and the animals were killed after 0, 12, 18, 24, and 36 h. Kinase assays showed that cyclin E- and Cdk2-associated activities were lower in AAF-exposed liver than in unexposed liver after PH. Although the total cellular levels of cyclin E and Cdk2 were similar, cyclin E-Cdk2 assembly was markedly reduced. In unexposed hepatocytes, Cdk2 translocated to the nuclei after PH. Much of the nuclear Cdk2 was in a rapidly migrating form, presumably representing the Thr160-phosphorylated form of Cdk2. In contrast, in AAF-exposed liver both nuclear Cdk2 accumulation and Thr160-phosphorylation of Cdk2 were reduced. Although p53 and p21(waf1/cip1) were induced by AAF, the binding of p21 to cyclin E and Cdk2 was not increased in growth arrested liver. In conclusion, hepatocyte growth arrest caused by AAF exposure was characterized by a lowered Cdk2 activity that was accompanied by a reduced assembly of cyclin E-Cdk2 complexes but not by binding of p21. PMID- 10708482 TI - Transgenic coexpression of v-Ha-ras and transforming growth factor alpha increases epidermal hyperproliferation and tumorigenesis and predisposes to malignant conversion via endogenous c-Ha-ras activation. AB - Previously, transgenic mice were generated that overexpressed v-Ha-ras or human transforming growth factor alpha (TGFalpha) exclusively in the epidermis, by means of a targeting vector based on the human keratin 1 gene (HK1). Both transgenics exhibited a similar neonatal phenotype of epidermal hyperplasia/hyperkeratosis and, in adults, spontaneous and 12-O tetradecanoylphorbol-13-acetate (TPA)-induced papilloma formation. To assess the synergism in vivo between Ha-ras and TGFalpha, mating experiments were performed. All ras/TGFalpha double genotype progeny (HK1 less than, with dotras/alpha) exhibited an increased epidermal hyperplasia/hyperkeratosis in neonates and accelerated spontaneous papillomatogenesis, compared with single transgenic siblings. HK1 less than, with dotras/alpha mice from the mild lines of HK1 less than, with dotrasxHK1 less than, with dotTGFalpha developed spontaneous papillomas that were not shown in either their parental mice or single transgenic littermates. Unlika in parental or single-genotype siblings, in which TPA promotion-elicited papillomas remained benign, TPA promotion elicited autonomous papillomas in HK1 less than, with dotras/alpha mice and exhibited a novel susceptibility to malignant conversion. Sequence analysis of the endogenous c-Ha ras from spontaneous and TPA-induced HK1 less than, with dotras/alpha papillomas revealed wild-type sequence. However, carcinomas exhibited c-Ha-ras mutations at codon 61. All tumors analyzed to date expressed wild-type p53. These data provide in vivo evidence that Ha-ras and TGFalpha cooperate in the induction of epidermal hyperplasia and spontaneous tumor formation and predispose to malignant conversion via endogenous c-Ha-ras activation. PMID- 10708483 TI - Azoxymethane induces KI-ras activation in the tumor resistant AKR/J mouse colon. AB - A differential susceptibility phenotype to the organotropic colon carcinogen azoxymethane (AOM) has been described in mice. The following studies were undertaken to test the hypothesis that intraspecific susceptibility can be accounted for by the specific complement of genetic alterations acquired by precancerous colon lesions referred to as aberrant crypt foci (ACF). As an initial approach to this question, mutations in codons 12 and 13 of the Ki-ras proto-oncogene were assessed in ACF, normal-appearing AOM-treated colonic epithelium, and tumors from A/J and SWR/J (susceptible) as well as AKR/J (resistant) mice. Four-week-old male mice were injected intraperitonealy, with AOM once a week for a total of 6 wk and killed 4 and 24 wk after the last injection. DNA was isolated from microdissected tissue, and polymerase chain reaction (PCR)-amplified products of Ki-ras exon 1 (codons 12 and 13) were directly sequenced from microdissected tissues. At 4 wk after AOM exposure, there was no significant difference in the frequency of Ki-ras activation (20-33%) between the three strains. Ki-ras mRNA expression was also evaluated by reverse transcription (RT)-PCR analysis and was comparably reduced (40-50%) in all three strains at the 4 wk time point. However, Ki-ras expression returned to normal by 24 wk after treatment. Finally, to gain further insight into the molecular pathogenesis underlying this experimental tumor model, analysis of the adenomatous polyposis coli (APC) protein within the colonic epithelium was undertaken by using an immunohistochemical approach. Although the APC protein was lost to a varying extent in tumors from A/J and SWR/J mice, the full-length form of the protein was still present in precancerous ACF isolated from each of the three strains, regardless of the degree of dysplasia of the lesion. A further molecular genetic analyses of ACF will be required to gain a more complete understanding of the molecular basis of tumor susceptibility phenotype in this murine model. PMID- 10708484 TI - Somatic cell hybrids for high-density mapping of chromosome 2 breakpoints in radiation-induced myeloid leukemia cell lines from inbred mice. AB - Chromosome 2 (chr 2) deletions are recurrent abnormalities in acute myeloid leukemia (AML) induced by ionizing radiation in the mouse. The localization of deletion sites has proven extremely useful in providing information on the molecular mechanisms of leukemogenesis. The models available for the study of AML are mostly represented by inbred mouse strains, in which the molecular resolution of breakpoints is problematic. In this study, we have examined five leukemic cell lines exhibiting hemizygous chr 2 loss, derived from CBA, C3H, or (C57BLxCBA/H) F1 mice in which AML had been induced by a whole-body dose of radiation. By application of a somatic cell hybridization technique, we have generated interspecific cell hybrids retaining the deleted murine chr 2 homologue. This strategy permitted a very detailed genetic analysis allowing the utilization of any genetic marker on chr 2 without a requirement for polymorphism. Somatic cell hybrid clones were subjected to a high-density polymerase chain reaction-based microsatellite screening using 62-106 informative markers for each cell line. Detailed maps accurately defining chr 2 breakpoints were obtained. The identification of critical breakpoint markers allowed the construction of partial yeast artificial chromosome contigs across chr 2 breakpoints. These maps represent an essential resource for cloning of the breakpoint regions. PMID- 10708485 TI - The human PCPH proto-oncogene: cDNA identification, primary structure, chromosomal mapping, and expression in normal and tumor cells. AB - We identified a human cDNA encoding a 47-kDa protein that shares 78% and 87% identity with the products of the Syrian hamster and mouse PCPH proto-oncogenes respectively. The human homolog was localized by radiation-hybrid mapping to chromosome band 14q24.3, a region syntenic to the Pcph location on mouse chromosome 12. Northern analyses revealed that PCPH mRNA was widely distributed in normal human adult tissues, but its expression varied significantly among human tumor cells and cell lines of several tissue types, regardless of the level of expression in the corresponding normal tissues. The highest levels of PCPH mRNA and protein were detected in kidney and liver. However, PCPH was not expressed in the majority of human neoplasms tested, including kidney tumors. These data provide suggestive evidence for a possible association of the lack of PCPH expression to the neoplastic phenotype of human tumor cells. Our results should prove instrumental in designing studies to define the cellular function of the human PCPH proto-oncogene. PMID- 10708487 TI - Karol Joseph Mysels (1914-1998). PMID- 10708486 TI - The expansion of the CAG repeat in exon 1 of the human androgen receptor gene is associated with uterine endometrial carcinoma. AB - Uterine endometrial carcinoma is common among women. Several reports indicated that this cancer is influenced by androgens mediated through their receptors. The human androgen receptor gene contains a polymorphic CAG repeat in the coding region of exon 1. Other studies suggested a possible association between the CAG repeat of this gene and the development of several cancers. DNA samples isolated from 29 patients with sporadic endometrial cancer were analyzed for allelic changes in 12 highly polymorphic microsatellite loci on nine chromosomes, containing CAG repeat in the exon 1 of the androgen receptor gene. A significantly high rate of allelic change in the CAG repeat was observed (51.7%) in these patients, although the frequencies of additional loci were similar to those reported by others (0-22.2%). The changed alleles of the tumor tissue were always longer than that of normal tissue except in only one case. Since only one allele on the X chromosome is commonly active in female cells, a differential methylation assay was carried out by using genomic DNA cut with HpaII. In 14 of 15 cases, we found that the activated allele was longer in these samples from tumor tissues than those from normal tissue; in the remaining case, the length of this repeat was unchanged. The expression assays were done by using poly(A+) RNA from tumor and normal uterine tissues, revealing that an activated allele in these tumor tissues was longer than that in the normal tissues in all the cases examined. These findings suggest that expansions of the CAG repeat in the androgen receptor gene may play an important role in the carcinogenesis of uterine endometrial cells. PMID- 10708488 TI - A Titration Microcalorimetric Study of the Effects of Halide Counterions on Vesicle-Forming Aggregation in Aqueous Solution of Branched-Chain Alkylpyridinium Surfactants. AB - Titration microcalorimetry is used to study the influences of iodide, bromide, and chloride counterions on the aggregation of vesicle-forming 1-methyl-4-(2 pentylheptyl)pyridinium halide surfactants. Formation of vesicles by these surfactants was characterised using transmission electron microscopy. When the counterion is changed at 303 K through the series iodide, bromide, to chloride, the critical vesicular concentration (cvc) increases and the enthalpy of vesicle formation changes from exo- to endothermic. With increase in temperature to 333 K, vesicle formation becomes strongly exothermic. Increasing the temperature leads to a decrease in enthalpy and entropy of vesicle formation for all three surfactants. However the standard Gibbs energy for vesicle formation is, perhaps surprisingly, largely unaffected by an increase in temperature, as a consequence of a compensating change in both standard entropy and standard enthalpy of vesicle formation. Interestingly, standard isobaric heat capacities of vesicle formation are negative, large in magnitude but not strikingly dependent on the counterion. We conclude that the driving force for vesicle formation can be understood in terms of overlap of the thermally labile hydrophobic hydration shells of the alkyl chains. Copyright 2000 Academic Press. PMID- 10708489 TI - Heterogeneous Nucleation of n-Butanol Vapor on Submicrometer Charged and Neutral Particles of Lactose and Monosodium Glutamate. AB - Condensation of a supersaturated vapor of n-butanol on monodisperse submicrometer particles of lactose and monosodium glutamate is investigated in a flow cloud chamber (FCC). The dependence of critical supersaturation S(cr) on the particle size in the range 30 to 90 nm is experimentally examined. The results show that the size dependence of S(cr) qualitatively agrees with that predicted by the Fletcher version of the Volmer theory of heterogeneous nucleation, but to a lesser degree. The experimental S(cr) is smaller than the theoretical prediction even with the line tension and surface diffusion taken into account, and they induce heterogeneous nucleation better than perfectly wetted particles. The discrepancy can not be fully accounted for by the effects of line tension and surface diffusion and the existing theory concerning the curvature-dependent surface tension. The condensation on single positive-charged particles of diameter 30, 60, and 90 nm is also examined. A lowering of S(cr) at an efficiency much larger than the prediction by Volmer's theory for ion-induced nucleation is observed, and the charge effect fades away as particle size increases. Copyright 2000 Academic Press. PMID- 10708490 TI - The Surface Structure of Hydroxyapatite Single Crystal and the Accumulation of Arachidic Acid. AB - Single crystals of hydroxyapatite (HAp) were grown by a flux method using beta tricalcium phosphate and Ca(OH)(2) under hot isostatic pressure. After chemical etching by 0.05 N HCl aqueous solution, the HAp crystal surfaces were observed by an atomic force microscope (AFM) and shown dominantly to consist of steps with heights corresponding to a lattice distance d(100) or its 2/3. Arachidic acid films were accumulated on the etched HAp crystal by a Langmuir-Blodgett method. From section analyses by AFM, a distance between the carboxyl groups of arachidic acid and the HAp surface was estimated to be approximately 0.04 nm, sufficiently adjacent for a chemical interaction to take place. Copyright 2000 Academic Press. PMID- 10708491 TI - Modes of Nonaxisymmetry in the Stability of Fixed Contact Line Liquid Bridges and Drops. AB - A method is presented for predicting the onset and stability character of nonaxisymmetric modes in liquid bridges and drops. The analysis applies to any fixed contact line axisymmetric interface in a steady force field. The onset and stability character of nonaxisymmetric equilibria in liquid bridges and drops is determined. Perturbation analysis is used to locate branches to nonaxisymmetry, and the configuration of the branches then gives the stability character. The number of unstable modes to both constant pressure and constant volume disturbances can be determined, so that changes in stability beyond the primary loss of stability may be examined. Although the first nonaxisymmetric mode tends to dominate higher order modes are significant for liquid bridges where length is less than radius and for drops at higher Bond numbers. At Bond numbers significantly greater than unity, the onset of the least unstable nonaxisymmetric modes tend to collapse between the fixed pressure and fixed volume axisymmetric modes of instability. For liquid bridges, two non-singular classes of nonaxisymmetric mode are distinguished: the predominant, classical shift mode; and a previously unreported tilt mode. The range over which the stability character of fixed contact line liquid bridges and drops is understood is significantly extended. Copyright 2000 Academic Press. PMID- 10708492 TI - Resonance Behavior of Oscillating Bubbles. AB - A convolution-type equation has been derived to describe the behavior of a bubble under periodical pressure oscillations. This equation holds for a diffusion controlled adsorption mechanism and small disturbances of the equilibrium state, and it describes both the established and transition regimes of bubble oscillation. Systems free of any surfactant and in the presence of a surfactant are considered. The results obtained allow all aspects of surfactant influence on the bubble oscillation resonance to be analyzed. The sharp increase in the bubble oscillation amplitude may result in bubble detachments, even at rather low harmonic pressure oscillations. The presence of surfactant can result in a depression of the resonance amplitudes. Copyright 2000 Academic Press. PMID- 10708493 TI - Calcium Carbonate Deposition on Cellulose. AB - Cellulose powder was found to be a substrate favoring the deposition of calcite crystals from stable supersaturated solutions at pH 8.50 and at 25 degrees C. Kinetic analysis of the initial rates showed that they were proportional with the relative supersaturation with respect to calcite. Analysis of the dependence of the induction periods on the initial solution conditions showed that the number of ions forming the critical nucleus was 5. The second-order dependence of the rate of precipitation of calcite on cellulose on the solution supersaturation suggested a surface controlled mechanism. The surface energy of the calcite nuclei growing on cellulose was calculated to be 46 mJ m(-2) from the dependence of the induction time on the solution supersaturation. The overgrowth of calcite on cellulose was done selectively on the macromolecules possibly through active sites formation at ionizable functional groups (-OH). The nucleating capability of cellulose was found to be comparable with that of sulfonated polystyrene and significantly lower in comparison with sulfonated polystyrene divinyl benzene copolymer on which vaterite was formed. This fact together with the selective growth of the most stable calcite suggested that stereochemical factors are very important in determining both the kinetics and the nature of the polymorph formed. Copyright 2000 Academic Press. PMID- 10708494 TI - Thermophoretic Motion of a Sphere Parallel to an Insulated Plane. AB - An analytical study is presented for the thermophoresis of a sphere in a constant applied temperature gradient parallel to an adiabatic plane. The Knudsen number is assumed to be small so that the fluid flow can be described by a continuum model with a thermal creep and a hydrodynamic slip at the particle surface. A method of reflections is used to obtain the asymptotic formulas for the temperature and velocity fields in the quasisteady situation. The thermal insulated plane may be a solid wall (no-slip) and/or a free surface (perfect slip). The boundary effect on the thermophoretic motion is found to be weaker than that on the axisymmetric thermophoresis of a sphere normal to a plane with constant temperature. In comparison with the motion driven by gravitational force, the interaction between the particle and the boundary is less significant under thermophoresis. Even so, the interaction between the plane and the particle can be very strong when the gap thickness approaches zero. For the thermophoretic motion of a particle parallel to a solid plane, the effect of the plane surface is to reduce the translational velocity of the particle. In the case of particle migration parallel to a free surface due to thermophoresis, the translating velocity of a particle can be either greater or smaller than that which would exist in the absence of the plane surface, depending on the relative thermal conductivity and the surface properties of the particle and its relative distance from the plane. Not only the translational velocity but also the rotational velocity of the thermophoretic sphere near the plane boundary is formulated analytically. The rotating direction of the particle is strongly dominated by its surface properties and the internal-to-external thermal conductivity. Besides the particle motion, the thickness of the thermophoretic boundary layer is evaluated by considering the thermophoretic mobility. Generally speaking, a free surface exerts less influence on the particle movement than a solid wall. Copyright 2000 Academic Press. PMID- 10708495 TI - Adsorption of Water-Soluble Proteins onto Bubbles in Continuous Foam Separation. AB - The mechanism of water-soluble protein enrichment in continuous foam separation was studied. The liquid flow rate and the protein concentration in the foam phase were measured at various heights from the interface between the bulk liquid and foam layer, and the intrinsic values at the interface were estimated by the extrapolation method to determine the accurate adsorption density on the bubble surface. Ovalbumin (OA) and hemoglobin (HB) were used as the soluble proteins. The solution pH values were varied from 3.5 to 6.0 for OA and from 6.0 to 8.0 for HB. The experimental isotherms for OA and HB were compared to the Langmuir isotherm, and the two adsorption parameters of the equilibrium constant, K, and the saturated density, gamma, at each pH were determined. Both gamma values obtained for OA and HB showed maxima at their isoelectric point (pH 4.6 for OA and pH 6.8 for HB). Assuming that OA and HB molecules are spherical in shape and are adsorbed on the bubble surface in a close-packed structure at saturation, the calculated diameters for OA and HB molecules were quite similar to the literature values. The variation in gamma for both OA and HB is discussed qualitatively in relation to the net charge of the protein molecule. Copyright 2000 Academic Press. PMID- 10708496 TI - Self-Association of Poly AB - Poly[2-(beta-D-glucosyloxy)ethyl acrylate] (PGEA) was prepared by radical polymerization of 2-(2',3',4',6'-tetra-O-acetyl-beta-D-glucosyloxy)ethyl acrylate (AcGEA) followed by O-deacetylation. Fluorescence measurements revealed that PGEA tended to associate into aggregates in an aqueous medium. The critical aggregation concentration (cac) of PGEA in water was determined by using N-phenyl 1-naphthalamine (PNA) as a fluorescent probe. The dependence of cac on the molecular weight of PGEA and temperature was observed. A copolymer of GEA with a hydrophobic monomer, i.e., stearyl acrylate (SA), was also synthesized in order to increase the hydrophobicity of PGEA and to understand the hydrophobic effect on cac. It was found that the cac of P(GEA-co-SA) was much lower than that of PGEA, and it decreased with increasing ratio of SA in the copolymer. Size exclusion chromatography (SEC), dynamic light scattering (DLS), and transmission electron microscopy (TEM) measurements indicated that the aggregates have hydrophobic interiors; they were spherical in shape and their sizes varied in a broad range. This phenomenon was explained in terms of a combination of hydrophobic interaction and cooperative hydrogen bonding. Copyright 2000 Academic Press. PMID- 10708497 TI - Film Formation and Redispersion of Waterborne Latex Coatings. AB - Poly(vinyl acetate-co-ethylene) latex dispersions are prepared and their films investigated with a focus on the effect of composition upon redispersion. Films of dispersions containing sufficient amounts of poly(vinyl alcohol) (PVA) can be redispersed in water. This property is lost in the presence of surfactant, a fact which suggests a procedure to control film formation. It is demonstrated that redispersion is due to a PVA-membrane which separates the particles. Loss of redispersibility in the presence of surfactant proceeds with the breakup of the membranes and a corresponding change of film properties. Experimental data is provided by light microscopy, mechanical testing, and TEM in conjunction with a staining method new to the field. The hypothesis is developed that interaction with surfactant leads to imperfect PVA-membranes that are no longer able to prevent latex polymer interdiffusion. Fluorescence correlation spectroscopy demonstrates the formation of surfactant micelles, as well as the simultaneous adsorption and aggregation of PVA onto the micelles. It is concluded that the competing surface of the surfactant micelles traps enough PVA to cause thinning and fragmentation of the membranes surrounding the particles, which enables interdiffusion of latex polymer. This effect can be used to convert the system from one forming a redispersible coating to one forming a nonredispersible (permanent) film. Copyright 2000 Academic Press. PMID- 10708498 TI - Dynamic Mobility of Colloidal Particles with Thick Double Layers. AB - The dynamic mobility spectra of several colloidal systems having a ratio of particle radius to double-layer thickness between 1 and 20 have been measured using the technique of electroacoustics. Good agreement is found between the experimental mobility spectra and the theoretical spectra generated by the computer program of Manglesdorf and White for spherical monodisperse suspensions with sizes in the neighborhood of 0.1 um. Smaller and larger particles show some minor discrepancies which are more likely to be due to limitations of the model systems being used for the test than to any basic limitation of the theoretical analysis. Copyright 2000 Academic Press. PMID- 10708499 TI - Determination of the Cation-Exchange Capacity of Muscovite Mica. AB - High cation-exchange capacity (CEC) muscovite mica with a homoionic surface was prepared by replacing the Li(+) surface ions of partially delaminated Li-mica with K(+). The CEC of this K-mica was determined by exchanging its surface cations with Cs(+), NH(+)(4), methylene blue (MB(+)), and copper triethylenetetramine [Cu(trien)(2+)]. The kinetics of these exchange reactions were studied and showed large differences depending on their relative affinities to mica. The NH(4)(+)/K(+) exchange was slow, while the Cs(+) and Cu(trien)(2+)/K(+) exchange was fast. The MB(+)/K(+) exchange was quite slow and was not completed even after 99 h. Insufficient reaction time is one of the main reasons for the contradictory results reported in the literature for the CEC of aluminosilicates obtained by different methods. The CEC of mica can be photometrically measured by exchanging its surface cations with Cu(trien)(2+). Copyright 2000 Academic Press. PMID- 10708500 TI - Deposition of Oil Drops on a Glass Substrate in Relation to the Process of Washing. AB - The attachment of emulsion drops to glass substrates is investigated in relation to the redeposition of oil drops in the process of washing. It turns out that the drops of a surfactant-stabilized oil-in-water emulsion cannot be attached to an immersed glass plate simply by the buoyancy force. However, the same drops can be deposited on the plate when the latter is pulled out of the emulsion, i.e., when the drops are pressed against the substrate by a receding meniscus. We measured the amount of the oily deposit as a function of the pH, ionic strength, and composition of an amphoteric-anionic surfactant mixture. The enhanced oil deposition at low pH correlates with the domain in which the emulsion drops and the solid substrate bear opposite electric charges. This was established by zeta potential measurements with oil drops and glass particles. The anionic surfactant brings negative surface charge to the oil droplets and suppresses the oil deposition on the negatively charged glass. With the increase of the fraction of the amphoteric surfactant in the mixture, the zeta-potential is converted from negative to positive, and the oil deposition grows almost linearly with the potential. In general, the deposition of oil drops by a receding meniscus is governed by an interplay of electrostatic and hydrodynamic factors. Copyright 2000 Academic Press. PMID- 10708501 TI - The Effect of Adsorbed Sodium Dodecylsulfate at the Alumina-Water Interface on the Luminescence Quenching of Tris(2,2'-bipyridine) ruthenium(II) by Nitrophenols. AB - The luminescence quenching of tris(2,2'-bipyridine)ruthenium(II) (Ru(bpy)(3)(2+)) by 2-, 3-, and 4-nitrophenol (NO(2)PhOH) was studied in the absence and presence of aggregates of sodium dodecyl sulfate (SDS) in the bulk solution and in the adsorbed state on alpha-alumina at pH 2. At this pH, the alumina has a superficial excess of positive charge. The uncorrected luminescence spectrum of Ru(bpy)(3)(2+) did not show any spectral shift in alumina suspension in the absence of SDS in relation to the spectrum in aqueous solution at pH 2 (lambda(max)=609 nm). In the presence of 1 g/L alumina and concentrations of SDS between 0.6 and 0.8 mM and the critical micellar concentration (CMC), the surfactant forms surface aggregates-hemimicelles or admicelles. In these surface aggregates, the emission spectrum showed a red shift (lambda(max)=637 nm). At SDS concentrations higher than the CMC, the surfactant forms micelles in the bulk solution and surface aggregates onto alpha-alumina. However, from the spectral shift observed with the increase in SDS concentration, it seems that, at high surfactant concentrations in the bulk, the metallic complex prefers to remain in the micellar instead of in the hemimicellar phases. The emission maximum in micelles was in 627 nm. The Stern-Volmer constants (K(SV)) of luminescence quenching were determined from steady-state emission intensity measurements at the maximum of emission in each case. The luminescence quenching of Ru(bpy)(3)(2+) showed that the NO(2)PhOHs are incorporated into both types of aggregates. On the basis of the K(SV) values as a function of the SDS concentration, the process of luminescence quenching of Ru(bpy)(3)(2+) by these compounds is more efficient in premicelles/micelles than in the surface aggregates onto alpha-alumina. This was observed up to a concentration of 5 mM SDS. These results can be interpreted as an effect of the increase in polarity of the micellar microenvironment on the electron-transfer mechanism for the quenching process. At SDS concentrations higher than 5 mM, the plot of K(SV) vs SDS concentration in the presence of 1 g/L alumina is shifted to higher SDS concentrations in relation to the observed plot in the absence of 1 g/L alumina. This fact can be explained in terms of the surfactant that forms surface aggregates onto alumina which do not contain the metallic complex. Copyright 2000 Academic Press. PMID- 10708502 TI - Interaction of 2-Amino-, 3-Amino-, and 4-Aminopyridines with Chromium and Manganese Ferrocyanides. AB - The present investigation deals with the interaction of 2-aminopyridine, 3 aminopyridine, and 4-aminopyridine with chromium and manganese ferrocyanides. Chromium ferrocyanide possesses better adsorbing properties than manganese ferrocyanide. Maximum uptake was observed at neutral pH (pH 7.0). The adsorption data obtained at neutral pH are fitted in a Langmuir adsorption isotherm. The adsorption behavior of the aminopyridines studied follows the order 3 aminopyridine >4-aminopyridine >2-aminopyridine. The infrared spectral studies of adsorption adducts indicate that adsorption takes place through interactions between the adsorbate molecule and the outer divalent metal ion of metal ferrocyanides. From these studies, it is clear that metal ferrocyanides and metal ions play a major role in the stabilization of organic molecules through their surface activity in the prebiotic environment. Copyright 2000 Academic Press. PMID- 10708503 TI - The Effect of Protein Concentration on Electrophoretic Mobility. AB - The electrophoretic mobility of haemoglobin was measured in a novel membrane electrophoresis cell and by electrophoretic light scattering. The effect of protein concentration was investigated at different ionic strengths in two different buffer systems. The results indicated that although the effect of the concentration is weak, the mobility did decrease linearly with an increase in volume fraction throughout the range of volume fractions investigated (φ<0.06). This dependence is more pronounced at lower kappaa values where double-layer interactions between the particles are more significant. Protein contribution to the solution ionic strength alone cannot explain the observed reduction in electrophoretic mobility. The theory of Shugai et al. (Shugai, A. A., Carnie, S. L., Chan, D. Y. C., and Anderson, J. L., J. Colloid Interface Sci. 191, 357 (1997)) was found to be adequate in describing particle interactions. The agreement with Shugai's theory is somewhat surprising considering the polydispersed nature of the samples, uncertainties in protein size, changes in ionic strength at high protein concentrations, and possible membrane-protein interactions not accounted for in the theory. Copyright 2000 Academic Press. PMID- 10708504 TI - Creaming and Rheology of Oil-in-Water Emulsions Containing Sodium Dodecyl Sulfate and Sodium Caseinate. AB - The creaming and rheology of fine n-tetradecane oil-in-water emulsions at pH 6.8 containing the commercial protein sodium caseinate and the ionic surfactant sodium dodecyl sulfate (SDS) have been studied, and an overview diagram relating surfactant composition and creaming stability has been constructed. The presence of both SDS and sodium caseinate in an emulsion system increases the overall stability with respect to creaming. Excess SDS promotes destabilization through fast creaming; this can be attributed to depletion flocculation brought about by unadsorbed surfactant micelles. Addition of sodium caseinate was found to reduce this effect, even at relatively high SDS concentrations. The behavior of the caseinate + SDS emulsions is thus different from the behavior of the previously reported caseinate + Tween 20 systems, where the combination of the two surface active agents was found to reduce the emulsion stability, as indicated by fast creaming and shear-thinning rheology. Addition of sodium chloride was found to increase the extent of non-Newtonian behavior and to enhance the degree of creaming for SDS-containing emulsions. Increased caseinate levels in these systems seem to offer some stabilization through reduction of the shear-thinning character and improvement in creaming stability. These phenomena can be explained in terms of a considerable amount of SDS binding to the protein, which reduces the amount of SDS available to promote protein displacement and depletion flocculation. In contrast to the SDS systems, the properties of equivalent emulsions containing caseinate + nonionic surfactant Tween 20 are relatively insensitive to salt content. Copyright 2000 Academic Press. PMID- 10708505 TI - Sorption of the Pesticide Endosulphan on Two Indian Soils. AB - Adsorption of endosulphan on uncontaminated sandy loam and silt clay loam soils in acetone-water and methanol-water mixtures at different f(s) values has been studied by the batch technique. Higher adsorption of endosulphan was observed on sandy loam soil than on silt clay loam soil, as was predicted from Freundlich constant K values and partition coefficient K(D) values. The K and K(D) values also confirm that adsorption of endosulphan was higher in acetone-water mixtures than in methanol-water mixtures and decreases with increases in the volume fraction of solvents (acetone, methanol), the f(s) values. The data were used to evaluate the cosolvent theory for describing adsorption of endosulphan in acetone water and methanol-water mixtures. The aqueous phase partition coefficient K(DW) (mol/g) normalized on f(OC) (fraction of soil organic carbon) for endosulphan was evaluated by extrapolating to f(s)=0. Copyright 2000 Academic Press. PMID- 10708506 TI - Electroactive Films of Alternately Layered Polycations and Iron-Sulfur Protein Putidaredoxin on Gold. AB - Layered, electrochemically active films of bacterial iron-sulfur protein putidaredoxin (Pdx) and poly(dimethyldiallyammonium) (PDDA) polycations were constructed on gold electrodes coated with mercaptopropane sulfonate (MPS) and on quartz slides. Second-derivative UV-vis spectra suggested similar structures of Pdx in films and solutions at pH 7. Direct electrochemistry was achieved between Pdx and gold electrodes in these films, with significantly better electrochemical reversibility than in cast Nafion-lipid-Pdx films. A formal potential dispersion model gave a good fit to square wave voltammograms by regression analysis and was used to estimate an average apparent rate constant of 4.5 s(-1). Reduced Pdx in the polyion films did not react with its natural redox partner cytochrome P450(cam) because of unfavorable thermodynamics in the film environment. Copyright 2000 Academic Press. PMID- 10708507 TI - Surface Properties of Vitreous Fibers. AB - The surface properties of various vitreous fibers, suspected to be toxic to humans and animals, were investigated by means of paramagnetic labels covalently linked to the surface. Computer-aided analysis of the electron paramagnetic resonance (EPR) spectra provided structural and dynamic information on the label and its environment. Calorimetric measurements provided information on the hydration mechanism. The results were analyzed in terms of (a) different polarity and interaction abilities of surface regions, (b) presence of ions at the surface, (c) silica contents, (d) vicinity of the interacting sites, (e) fiber dimension and morphology of the surfaces, and (f) water hydration. The mobility of the labels decreased due to interaction of the fibers with ions or ionic and polar groups at the surface. Close interacting sites were identified on the basis of spin-spin effects and were distinguished and quantified in strongly and weakly interacting sites. The spin-labeling technique indicated decreased ability of the surface to interact with decreased silicon concentration and in the presence of contaminants at the surface. The interaction with water revealed in all cases a substantial heterogeneity in hydrophilicity of surface sites. The labels were not easily hydrated. Vitreous fibers of various compositions adsorbed much more water than crystalline or amorphous silica; water coordinated to surface cations played a major role in the overall adsorption. The surface reaction mechanisms were the same on fibers of different compositions, but the surface composition affected the extent of adsorption. Glass wool exhibited a much higher adsorption capacity than rock wool under the same experimental conditions. In conclusion, the combination of EPR and calorimetric measurements provided insight into the surface properties of silica-based fibers. Copyright 2000 Academic Press. PMID- 10708508 TI - Flow-Induced Anisotropy in Mixtures of Associative Polymers and Latex Particles. AB - The effect of associative polymers on the structure and rheological behavior of colloidal suspensions is discussed. Adding associative polymer is known to increase the viscosity of the suspensions. At high shear rates the increase is close to what could be expected on the basis of the hydrodynamic effects of the added polymer. At low shear rates the viscosity increases much more. Small-angle light scattering (SALS) during flow is used here to investigate the underlying structural mechanisms. The SALS patterns indicate that the associative polymer changes the particulate structure: characteristic butterfly patterns appear even at relatively low particle volume fractions. They are not present in the suspensions without associative polymer. The patterns indicate that fluctuations in particle concentration are more pronounced in the flow direction than in the vorticity direction and that anisotropic particulate structures with an orientation along the vorticity direction develop. The evolution of their characteristic length scale during flow has been followed over time. Changing the hydrophilic part of the polymer from polyacrylamide to polyacrylic acid induces stronger associative interactions. In the suspensions this results in a reduction of the relative viscosity rather than an increase. The difference in degree of associativity between the polymers also has an effect on the SALS patterns in the suspensions both at rest and during flow. The rheology as well as the SALS suggest the presence of a strong polymer network in the second system. The competition between adsorption of the associative polymer on the particles with the intermolecular associations between the polymer chains seems to be responsible for the observed differences. Copyright 2000 Academic Press. PMID- 10708509 TI - Mixing Properties of Two-Dimensional Lattice Solutions of Amphiphiles. AB - Lattice Monte Carlo simulations of two-dimensional amphiphile solutions are used to examine the accuracy of the mixing properties predicted by lattice theories such as the Flory-Huggins theory, random-solution approximation, and quasichemical approximation. The internal energy, Helmholtz free energy, and entropy of mixing have been calculated from the configurational energy data obtained from the simulations, and the effect of nonrandom mixing on these properties has been determined. The quasichemical approximation predicts the entropy and Helmholtz free energy of mixing accurately for the amphiphile solution, but fails to predict the energy of mixing, due to the presence of microphase (self-aggregation) separation, which is beyond the reach of the quasichemical approximation, a mean-field theory. Helmholtz free energy of mixing is predicted accurately, and the shielding of the solvophobic segments in the microphase leads to small energies of mixing compared to the entropy of mixing. Copyright 2000 Academic Press. PMID- 10708510 TI - Reexamination of 2-Naphthol Adsolubilization on Alumina with Sodium Dodecyl Sulfate Adsorption. AB - Adsolubilization behavior of 2-naphthol on alumina with adsorption of sodium dodecyl sulfate (SDS) at pH 3.5 in the presence of 10 mmol dm(-3) NaCl was reexamined. The adsolubilized amount of 2-naphthol increased sharply and reached a maximum, then decreased with SDS concentration. The decrement of the adsolubilized amount began below the critical micelle concentration of SDS. From the dispersion state of the alumina suspension and the SDS adsorption isotherm, it is demonstrated that the decrement of adsolubilization of 2-naphthol is not due to the partition of 2-naphthol between the SDS adsorbed layer and SDS micelles, but is due to the difference of SDS adsorption states such as monolayers and admicelles. Copyright 2000 Academic Press. PMID- 10708511 TI - Cobalt Ion-Doped TiO(2) Photocatalyst Response to Visible Light. AB - Photocatalytic activity under visible light irradiation was generated by doping a small amount of Co(2+) ions into TiO(2) particles. Nanometer-sized particles with the composition xCoO-(100-x) TiO(2) (xCo/TiO(2); 0300 nm) light irradiation but also induced the visible light (lambda>400 nm) response. The highest photocatalytic activities were obtained at x=0.03 for both irradiations. Copyright 2000 Academic Press. PMID- 10708512 TI - On a Simple Nonisothermal Adsorption Experiment with Organic Vapors and an Inertial Microbalance To Study the Surface Properties of Hybrid (Organic/Inorganic) Porous Materials. AB - A nonisothermal adsorption experiment using a controlled flow of cyclopentane in the 333-313 K range is used to simultaneously estimate the specific surface area and micropore volume of a hybrid (organic/inorganic) alcogel. For reference, the method is also applied to an all-inorganic material with a more rigid structure, namely, a high surface area SiO(2)-Al(2)O(3). The proposed data analysis provides guidelines to determine whether adsorption data on a certain adsorbate/adsorbent system can be modeled effectively as a convolution of BET (meso- and macropore) and Dubinin-Radushkevitch (DR, micropore) contributions. Copyright 2000 Academic Press. PMID- 10708513 TI - A 6-Mb high-resolution physical and transcription map encompassing the hereditary prostate cancer 1 (HPC1) region. AB - Several hereditary disease loci have been genetically mapped to the chromosome 1q24-q31 interval, including the hereditary prostate cancer 1 (HPC1) locus. Here, we report the construction of a 20-Mb yeast artificial chromosome contig and a high-resolution 6-Mb sequence-ready bacterial artificial chromosome (BAC)/P1 derived artificial chromosome (PAC) contig of 1q25 by sequence and computational analysis, STS content mapping, and chromosome walking. One hundred thirty-six new STSs, including 10 novel simple sequence repeat polymorphisms that are being used for genetic refinement of multiple disease loci, have been generated from this contig and are shown to map to the 1q25 interval. The integrity of the 6-Mb BAC/PAC contig has been confirmed by restriction fingerprinting, and this contig is being used as a template for human chromosome 1 genome sequencing. A transcription mapping effort has resulted in the precise localization of 18 known genes and 31 ESTs by database searching, exon trapping, direct cDNA hybridization, and sample sequencing of BACs from the 1q25 contig. An additional 11 known genes and ESTs have been placed within the larger 1q24-q31 interval. These transcription units represent candidate genes for multiple hereditary diseases, including HPC1. PMID- 10708514 TI - Cloning and expression of nope, a new mouse gene of the immunoglobulin superfamily related to guidance receptors. AB - The novel mouse gene Nope was identified due to its proximity to the Punc gene on chromosome 9. With a domain structure of four immunoglobulin domains, five fibronectin type III repeats, a single transmembrane domain, and a cytoplasmic domain, Nope encodes a new member of the immunoglobulin superfamily of cell surface proteins. It displays a high level of similarity to Punc, as well as to guidance receptors such as the Deleted in Colorectal Cancer protein and Neogenin. Nope is expressed during embryonic development in the notochord, in developing skeletal muscles, and later in the ventricular zone of the nervous system. In the adult brain, Nope can be detected in the hippocampus. Radiation hybrid mapping of Nope, Punc, and Neogenin placed all three genes in close vicinity on mouse chromosome 9. PMID- 10708515 TI - Identification of 18 mouse ABC genes and characterization of the ABC superfamily in Mus musculus. AB - ATP-binding cassette (ABC) genes encode a family of transport proteins known to be involved in a number of human genetic diseases. In this study, we characterized the ABC superfamily in Mus musculus through in silico gene identification and mapping and phylogenetic analysis of mouse and human ABC genes. By querying dbEST with amino acid sequences from the conserved ATP-binding domains, we identified and partially sequenced 18 new mouse ABC genes, bringing the total number of mouse ABC genes to 34. Twelve of the new ABC genes were mapped in the mouse genome to the X chromosome and to 10 of the 19 autosomes. Phylogenetic relationships of mouse and human ABC genes were examined with maximum parsimony and neighbor-joining analyses that demonstrated that mouse and human ABC orthologs are more closely related than are mouse paralogs. The mouse ABC genes could be grouped into the seven previously described human ABC subfamilies. Three mouse ABC genes mapped to regions implicated in cholesterol gallstone susceptibility. PMID- 10708516 TI - Detailed comparative gene map of rat chromosome 1 with mouse and human genomes and physical mapping of an evolutionary chromosomal breakpoint. AB - We report the localization of 92 new gene-based markers assigned to rat chromosome 1 by linkage or radiation hybrid mapping. The markers were chosen to enrich gene mapping data in a region of the rat chromosome known to contain several of the principal quantitative trait loci in rodent models of human multifactorial disease. The composite map reported here provides map information on a total of 139 known genes, including 80 that have been localized in mouse and 109 that have been localized in human, and integrates the gene-based markers with anonymous microsatellites. The evolutionary breakpoints identifying 16 segments that are homologous regions in the human genome are defined. These data will facilitate genetic and comparative mapping studies and identification of novel candidate genes for the quantitative trait loci that have been localized to the region. PMID- 10708517 TI - A transcript map of the chromosome 19q-arm glioma tumor suppressor region. AB - Allelic loss of the chromosome 19q arm is a frequent event in human diffuse gliomas, suggesting that it contains a tumor suppressor gene. Recent deletion mapping studies have broadly implicated a 1.6-Mb interval between D19S241E and D19S596, with a limited subset of tumors, suggesting that the region may be as narrow as 150 kb. Focusing on this smaller interval, we have used cDNA selection, exon amplification, and genomic sequencing to identify three novel transcripts (EHD2, GLTSCR1, and GLTSCR2) and to map two known genes (SEPW1 and CRX). A partial transcript map of 19 transcripts and two EST markers has been constructed for the 1.6-Mb interval D19S241E-D19S596. Ten of these transcripts, including the 5 mapped to the 150-kb deletion interval, have been examined for alterations in a panel of gliomas with allelic loss of 19q. Tumor-specific alterations have not been identified in the transcripts examined thus far. Collectively, these data should facilitate subsequent efforts to identify and characterize the remaining transcripts in the 1.6-Mb interval. PMID- 10708518 TI - The prepro vasoactive intestinal contractor (VIC)/endothelin-2 gene (EDN2): structure, evolution, production, and embryonic expression. AB - Murine vasoactive intestinal contractor (VIC) and its human analog endothelin-2 (ET2) are potent vasoactive hormones composed of 21 amino acids. To study the structural characteristics of the VIC/ET2 gene (HGMW-approved symbol EDN2), we isolated the full length of the mouse VIC gene. Sequence analysis indicates that a biologically active mature VIC peptide is produced from a 175-residue precursor protein; preproVIC (PPVIC). Several remarkable similarities of the PPVIC gene to the human preproendothelin-1 gene strongly suggest that the two genes have arisen from a common progenitor by gene duplication. Transfection of ACHN adenocarcinoma cells with the cDNA resulted in the production of VIC peptide. VIC production was increased by the deletion of the 3'-untranslated region, which contains an AU rich mRNA destabilizing sequence. Increased PPVIC gene expression during the late embryonic stage suggests an important function in development. This study provides the basis for disruption and regulation analysis of the gene, which may lead to a better understanding of VIC/ET2's physiological significance. PMID- 10708519 TI - Evolutionary divergence of the mouse and human Lgn1/SMA repeat structures. AB - The orthologous genomic segments on mouse chromosome 13D1-D3 and human chromosome 5q11.2-q13.3 have been extensively studied because of their involvement in two distinct disease phenotypes, spinal muscular atrophy (SMA) in human and susceptibility to Legionella pneumophila (determined by Lgn1) in mice. The overlapping intervals in both species contain genomic amplifications of distinct structure, indicating an independent origin. We have endeavored to construct a comprehensive comparative gene map of the mouse and human Lgn1/SMA intervals in the hopes that the origins and maintenance of the genomic amplifications may become clear. Our comparative gene map demonstrates that the only regional gene in common between the amplified segments in mouse and human is the Lgn1 candidate Naip/NAIP. We have also determined that mice of the 129 haplotype harbor seven intact and three partial Naip transcription units arranged in a closely linked direct repeat on chromosome 13. Several, but not all, of these Naip loci are contained within the Lgn1 critical interval. We present a model for the origins of the mouse and human repetitive arrays from a common ancestral haplotype. PMID- 10708520 TI - Solh, the mouse homologue of the Drosophila melanogaster small optic lobes gene: organization, chromosomal mapping, and localization of gene product to the olfactory bulb. AB - The Drosophila melanogaster small optic lobes gene (sol) is required for normal development of the neuropiles of the medulla and lobula complexes of the adult optic lobes. The predicted protein products of sol and its human homologue SOLH contain zinc-finger-like repeats, a calpain-like protease domain, and a C terminal domain of unknown function. Long-distance PCR was used to amplify genomic DNA for Solh, the mouse homologue of sol, following the identification of mouse Solh expressed sequence tags. The nucleotide sequence of the Solh coding region (6.0 kb) was determined. The predicted Solh protein of 1095 amino acid residues shows 89% identity (93% similarity) to the human homologue. Solh was localized by in situ hybridization to band A3.3 on mouse Chromosome 17, in a region of maintained homology with human 16p13.3. Antipeptide antibodies were prepared and verified by demonstration of specific reactivity with recombinant human SOLH protein prepared by in vitro transcription/translation and expression in insect cells using the baculovirus system. The antibodies were used to show that the Solh protein localizes to the olfactory bulb in mouse and rat brain, suggesting that it could have an analogous role in development of sensory system neurons in Drosophila and in mammals. PMID- 10708521 TI - Multiple potential intragenic regulatory elements in the CFTR gene. AB - The CFTR gene exhibits a complex pattern of expression that shows temporal and spatial regulation though the control mechanisms have not been fully elucidated. We have mapped DNase I hypersensitive sites (DHS) flanking the CFTR gene to identify potential regulatory elements. We previously characterized DHS at -79.5 and -20.9 kb with respect to the CFTR translational start site, DHS 3' to the gene at 4574 + 5.4-7.4 and 4574 + 15.6 kb, and a regulatory element in the first intron of the gene at 185 + 10 kb. We generated a cosmid contig to provide probes to evaluate the whole of the CFTR gene for DHS and have now mapped novel sites in introns 2, 3, 10, 16, 17a, 18, 20, and 21. These DHS show different patterns of cell-specific expression. PMID- 10708522 TI - Mapping and structure of DMXL1, a human homologue of the DmX gene from Drosophila melanogaster coding for a WD repeat protein. AB - The DmX gene was recently isolated from the X chromosome of Drosophila melanogaster. TBLASTN searches of the dbEST databases revealed sequences with a high level of similarity to DmX in a variety of different species, including insects, nematodes, and mammals showing that DmX is an evolutionarily highly conserved gene. Here we describe the cloning of the cDNA and the chromosomal localization of one of the human homologues of DmX, Dmx-like 1 (DMXL1). The human DMXL1 gene codes for a large mRNA of 11 kb with an open reading frame of 3027 amino acids. The putative protein belongs to the superfamily of WD repeat proteins, which have mostly regulatory functions. The DMXL1 protein contains an exceptionally large number of WD repeat units. The DMXL1 gene is located on chromosome 5q22 as determined by radiation hybrid mapping and fluorescence in situ hybridization. Although the function of the DMXL1 gene and its homologues in other species remains to be discovered, the high level of evolutionary conservation together with the unusual structure suggests that it probably has an important function. PMID- 10708523 TI - HSA4 and GGA4: remarkable conservation despite 300-Myr divergence. AB - The chicken (GGA) and human (HSA) genomes diverged around 300-350 Myr ago. Due to this large phylogenetic distance, significant synteny conservation has not been anticipated between the genomes of the two species. However, Zoo-FISH with HSA4 chromosome-specific paint on chicken metaphase chromosomes shows that the human chromosome corresponds largely to the GGA4cen-->q26 region. Comparative gene mapping data in the two species, though limited, provide strong support for these observations. The findings, together with the very recently published data on HSA9-GGAZ and HSA12-GGA1, show that some large chromosomal segments share conserved synteny in the two species. These syntenies are considerably disrupted in the mouse. This makes us believe that despite very early divergence, parts of the human and chicken genomes are more conserved than those of human and mouse, which radiated only 100-120 Myr ago. Moreover, the HSA4-GGA4q correspondence points to a "candidate" chromosome from the karyotype of a mammal-bird ancestor. The findings are thus a small but important step toward understanding the evolution of the two genomes. PMID- 10708524 TI - Structure and localization of mouse Pmscl1 and Pmscl2 genes. AB - Sera from some patients with polymyositis-scleoderma overlap syndrome (PM-SCL) recognize two antigenically unrelated proteins, PMSCL1 and PMSCL2. Complete mouse Pmscl1 and Pmscl2 cDNA sequences, chromosomal localizations, exon/intron structure, and promoter region sequences of the mouse Pmscl2 gene are presented. The PMSCL1 gene was found to overlap significantly with cyclin A2 in both human and mouse. As such, it may be deduced that PMSCL1 sequences map to human chromosome 4q27 and the proximal portion of mouse chromosome (Chr) 3 where human and mouse cyclin A2 genes reside. Analysis of human and mouse PMSCL1 cDNA sequences provides evidence that the PMSCL1 protein is 68 amino acids longer than previously thought. A BAC containing mouse Pmscl2 was localized to distal mouse Chr 4 by FISH. This BAC contains the microsatellite D4Mit310. D4Mit310 colocalizes with a number of genes that map to human 1p36. In fact, a STS (G25404) located 54.6 cR from the top of human chromosome 1 was found to contain PMSCL2 sequence upon BLAST search. PMID- 10708525 TI - BAC trimming: minimizing clone overlaps. AB - Bacterial vectors containing large inserts of genomic DNA are now the standard substrates for large-scale genomic sequencing. Long overlaps between some clones lead to considerable redundant effort. A method for deleting defined regions from bacterial artificial chromosome (BAC) inserts, using homologous recombination, was applied to minimize the overlap between successive BAC clones. This procedure, called trimming, was carried out in the recA(-) BAC host. We have precisely deleted up to 70 kb of DNA from BACs that were to be sequenced. This method requires minimal prior characterization of the clones: collections of BAC end sequences or STS-based maps will accelerate the process. BAC trimming will be useful in both small and large genome sequencing projects and will be of particular utility for gap closure in finishing phases. PMID- 10708526 TI - Exon sharing of a novel human zinc-finger gene, ZIM2, and paternally expressed gene 3 (PEG3). AB - We have identified a novel human gene, ZIM2 (zinc-finger gene 2 from imprinted domain), located 25 kb downstream of PEG3 (paternally expressed gene 3). ZIM2 produces two different-size transcripts, 2.5 and 9.0 kb in length, with highest levels of expression in adult testis and modest levels in fetal kidney and brain. The 2.5-kb transcript of ZIM2 consists of 11 exons and encodes a Kruppel-type (C2H2) zinc-finger protein with a conserved Kruppel-associated box (KRAB) domain. Rapid amplification of cDNA ends and cDNA sequencing studies showed that ZIM2 and PEG3 transcripts share identical 5'-ends, composed of 7 small exons. Alternative splicing events connect these 7 exons either with the remaining 2 exons of PEG3 or with the remaining 4 exons of ZIM2. Interestingly, the third among the 7 shared exons exhibits sequence similarity to leucine-rich domains that are found at the N-terminal region of a subset of KRAB-containing zinc-finger genes. Sequencing of the 5'-termini of both transcripts indicates that ZIM2 and PEG3 share identical transcription start sites and may also share upstream regulatory elements, although the two genes show distinct patterns of tissue-specific expression. PMID- 10708528 TI - The Microwave Spectrum of m-Tolunitrile: Methyl Internal Rotation and (14)N Nuclear Quadrupole Coupling. AB - The microwave spectrum of m-tolunitrile (3-methylbenzonitrile, m-C(6)H(4)CH(3)CN) has been investigated in the frequency range from 1 to 4 and 8 to 26.5 GHz. The spectra in the two lowest states of internal methyl rotation (m = 0, +/-1) were recorded by means of pulsed molecular beam Fourier transform microwave (MB-FTMW) spectrometers. The interpretation of the spectra was based on an asymmetric frame symmetric top Hamiltonian with inclusion of centrifugal distortion terms and first-order contributions from (14)N nuclear quadrupole coupling. A least-squares analysis yielded the rotational constants A = 3295.9103(10) MHz, B = 1199.1188(2) MHz, C = 883.9223(1) MHz, all elements of the nuclear quadrupole coupling tensor chi(aa) = -3.626(1) MHz, chi(bb) = 1.684(1) MHz, chi(cc) = 1.943(1) MHz, and chi(ab) = -1.870(3) MHz, as well as the threefold barrier to internal rotation, V(3) = 14.2 cm(-1), and the angle between the internal rotor axis and the principal moment of inertia a axis, θ = 42.66 degrees, using fixed values for the sixfold barrier term V(6) (-11 cm(-1)) and the moment of inertia of the methyl top I(alpha) (3.16 u A(2)). Copyright 2000 Academic Press. PMID- 10708527 TI - Characterization of a zebrafish/mouse somatic cell hybrid panel. AB - We have characterized a collection of zebrafish/mouse somatic cell hybrids with 211 genes and markers chosen from the 25 zebrafish linkage groups. Most of the zebrafish genome is represented in this collection with 88% of genes/markers present in at least one hybrid cell line. Although most hybrids contain chromosomal fragments, there are a few instances where a complete or nearly complete zebrafish chromosome has been maintained in a mouse background, based on multiple markers covering the entire chromosome. In addition to their use in mapping studies, this collection of somatic cell hybrids should constitute an important tool as a source of specific chromosome fragments and for assessing the function of genome regions. PMID- 10708529 TI - 13C(16)O(2): Global Treatment of Vibrational-Rotational Spectra and First Observation of the 2nu(1) + 5nu(3) and nu(1) + 2nu(2) + 5nu(3) Absorption Bands. AB - The effective operator approach is applied to the calculation of both line positions and line intensities of the (13)C(16)O(2) molecule. About 11 000 observed line positions of (13)C(16)O(2) selected from the literature have been used to derive 84 parameters of a reduced effective Hamiltonian globally describing all known vibrational-rotational energy levels in the ground electronic state. The standard deviation of the fit is 0.0015 cm(-1). The eigenfunctions of this effective Hamiltonian have then been used in fittings of parameters of an effective dipole-moment operator to more than 600 observed line intensities of the cold and hot bands covering the nu(2) and 3nu(2) regions. The standard deviations of the fits are 3.2 and 12.0% for these regions, respectively. The quality of the fittings and the extrapolation properties of the fitted parameters are discussed. A comparison of calculated line parameters with those provided by the HITRAN database is given. Finally, the first observations of the 2nu(1) + 5nu(3) and nu(1) + 2nu(2) + 5nu(3) absorption bands by means of photoacoustic spectroscopy (PAS) is presented. The deviations of predicted line positions from observed ones is found to be less than 0.1 cm(-1), and most of them lie within the experimental accuracy (0.007 cm(-1)) once the observed line positions are included in the global fit. Copyright 2000 Academic Press. PMID- 10708530 TI - The Sextic Centrifugal Distortion Terms for an Open-Shell Complex Consisting of a Diatomic Molecule in a (2S+1)Sigma Electronic State and a Closed-Shell Partner. AB - An effective Hamiltonian for calculating rotational energy levels of an open shell diatomic molecule, in a (2S+1)Sigma electronic state, weakly bonded to a closed-shell partner was presented (W. M. Fawzy, J. Mol. Spectrosc. 191, 68-80 (1998)). The Hamiltonian was given as H = H(ev) + H(rot) + H(sr) + H(ss) + H(cd) + H(srcd) + H(sscd), where all the quartic centrifugal distortion correction terms were included in the Hamiltonian term H(cd) but the sextic centrifugal distortion terms were ignored. This Hamiltonian is useful in cases where the complex has a well-defined equilibrium geometry and if the barrier to large amplitude motion is large compared to the rotational constant of both the closed shell molecule and its paramagnetic partner; if the barrier to large-amplitude motion is small compared to the rotational constant of one or both of the fragments, then a different treatment is required. In this paper, we introduce the new Hamiltonian terms H(sex(A))(cd) and H(sex(S))(cd), which represent the sextic centrifugal distortion correction terms for an asymmetric rotor. We also introduce all the nonvanishing matrix elements of each of the H(sex(A))(cd) and H(sex(S))(cd) operators. These operators and their matrix elements are required for calculating the rotational energy levels of relatively high J values in the described type of weakly bonded open-shell complexes. The terms H(sex(A))(cd) and H(sex(S))(cd) and their matrix elements are also valid for any stable asymmetric rotor in a nondegenerate electronic state. A brief discussion of the new Hamiltonian terms and their matrix elements is given. Copyright 2000 Academic Press. PMID- 10708531 TI - The Mechanism of Magnetically Tuned Singlet-Triplet Avoided Crossings in the A(1)A(2)-&Xtilde;(1)A(1) 4(1)(0) Band of Thioformaldehyde H(2)C&dbond;S. AB - Magnetically tuned singlet-triplet perturbations in the 4(1)A(1)A(2) 2(1)3(1)a(3)A(2) system of thioformaldehyde, found in ortho-rotational states (I = 1, the two hydrogen spins parallel) have been identified as being caused by vibronic spin-orbit coupling. This perturbation mechanism has been confirmed in several avoided crossings observed in this work for para states (I = 0, hydrogen spins antiparallel) which are much stronger. Parametrization of the theory has led to a quantitative understanding of the experimental frequency-field relations, and to an accurate prediction of the rovibrational energies of the triplet state. This in turn permitted the detection of about 100 Doppler-limited 2(1)3(1)a(3)A(2)-0(0) &Xtilde;(1)A(1) rovibronic transitions which led into fine structure states. The combined data was then used to determine a set of rotational, fine, and hyperfine triplet-state parameters, the term value T(0)(2(1)3(1)a(3)A(2)) = (16 685.385 +/- 0.002) cm(-1), and the spin-orbit vibronic singlet-triplet coupling constant, W(ST) = (0.0691 +/- 0.0016) cm(-1). A large number of frequency perturbations observed in the crossings, ranging from 2 to 300 MHz, can be explained with this single parameter. Copyright 2000 Academic Press. PMID- 10708532 TI - Submillimeter-Wave Spectroscopy of CO in the a(3)Pi State. AB - Submillimeter-wave absorption spectrum of CO in electronically excited a(3)Pi state was observed in the 540-830 GHz region by using a phase-locked BWO spectrometer. New rotational transitions up to J = 9-8 in the vibrational excited states up to v = 5 were analyzed accompanied with previous observations in the RF and millimeter-wave regions. A multivibrational states fit among a' (3)Sigma(+) (v = 0-3) and a(3)Pi (v = 0-7) states was performed in order to analyze overall perturbation between the a(3)Pi and a' (3)Sigma(+) states. As a result, the deperturbed rotational parameters were derived precisely to improve the RKR potential. Copyright 2000 Academic Press. PMID- 10708533 TI - High-Resolution Infrared and Millimeter-Wave Study of the v(3) = 1 State of HSiF(3) and DSiF(3). AB - Millimeter-wave spectra of HSiF(3) and DSiF(3) in the v(3) = 1 excited state have been measured from 100 to 490 GHz. Infrared spectra have been recorded in the nu(3) regions, nu(0) 424.0301 and 420.9320 cm(-1) in HSiF(3) and DSiF(3), respectively, with a resolution of 2.4 x 10(-3) cm(-1). Since in both species the parameters alpha(B)(3) and alpha(C)(3) have very similar values, no K structure could be resolved in the (Q)P and (Q)R clusters for low-to-medium K values. For high J the effect of the ground state D(JK) term more and more dominates and spreads the J clusters into opposite directions such that medium-to-high K components, particularly those with K = 3p, are resolved. Rotational and infrared data have been fitted together using a model up to sextic centrifugal distortion constants. No perturbations were indicated. Hot bands (nu(3) + nnu(6))-nnu(6) with n = 1, 2, and 3 have been detected and analyzed. Copyright 2000 Academic Press. PMID- 10708534 TI - Rotational Spectrum of the AsH(2) Radical in Its Ground State, Studied by Far Infrared Laser Magnetic Resonance. AB - The rotational spectrum of AsH(2) in its ground &Xtilde;(2)B(1) electronic state has been recorded using a far-infrared laser magnetic resonance spectrometer. The AsH(2) radical was produced inside the spectrometer cavity by the reaction of arsine (AsH(3)) with fluorine atoms. Hyperfine splittings from both (75)As and (1)H nuclei were observed, and analysis of the spectra has yielded accurate values for rotational, hyperfine, and Zeeman parameters. Copyright 2000 Academic Press. PMID- 10708535 TI - High-Resolution Laser Spectroscopy of YbCl: The A(2)Pi-X(2)Sigma(+) Transition. AB - The A(2)Pi-X(2)Sigma(+) transition of (174)Yb(35)Cl and (172)Yb(35)Cl has been rotationally analyzed for the first time. Doppler-limited laser excitation spectroscopy with selective detection of fluorescence was used to obtain spectra of the 0-0 and 1-0 bands with a measurement accuracy of approximately 0.0035 cm( 1). Resolved fluorescence was used to record the 0-1, 0-2, and 0-3 bands and to unequivocally assign the rotational numbering, N, to the laser excitation spectra. In total, over 1300 line positions have been measured and assigned for each of the two isotopomers and employed in least-squares fits of molecular parameters. The principal results for the A(2)Pi state are A(e) = 1491.494(2) cm( 1) and R(e) = 2.4433(1) A, and for the X(2)Sigma(+) state, R(e) = 2.4883(2) A and gamma(e) = 4.59(2) x 10(-3) cm(-1). The interaction between the X(2)Sigma(+) and A(2)Pi states has been investigated and is shown to be the main contributor to the spin-rotation splitting in the ground state. Copyright 2000 Academic Press. PMID- 10708536 TI - Measurements of (18)O-Enriched Ozone Isotopomer Abundances Using High-Resolution Fourier Transform Far-IR Spectroscopy. AB - The distribution of ozone isotopomers in ozone mixtures produced by electric discharge in mixtures of (16)O(2) and (18)O(2) at 77 K was measured by high resolution FTIR spectroscopy. It was of key importance to assess not only the total amount of isotopomers of a certain mass but also the relative amounts of corresponding asymmetric and symmetric ozone species of the same mass given as the ratios [(16)O(16)O(18)O]/[(16)O(18)O(16)O] and [(16)O(18)O(18)O]/[(18)O(16)O(18)O]. For many purposes both ratios have been assumed to have the statistical value 2.00. Pure rotational spectra in the far-IR region (30-100 cm(-1)) were recorded for three different (18)O-enriched ozone mixtures, all at 0.00185 cm(-1) resolution. All the spectra were corrected for thermal emission. Linestrengths for individual lines in a particular spectrum were measured by means of a fitting technique taking into account contributions from all other lines in the spectrum. For this purpose theoretical linestrengths for all six ozone species containing (16)O and (18)O obtained from a quantum number-dependent dipole operator were used. The ratios between observed and theoretical linestrengths were used to determine the abundances of individual isotopomers in a particular ozone mixture. For one of the ozone samples the abundances of all six ozone species were determined within 1% relative uncertainty. For the three ozone mixtures studied, the ratio between asymmetric and symmetric species of mono-(18)O ozone were determined to 1.99(2), 2.01(2), and 2.10(6). The ratio between asymmetric and symmetric species of di-(18)O ozone were determined to 2.51(4), 2.42(10), and 2.46(3). Copyright 2000 Academic Press. PMID- 10708537 TI - The nu(1) and nu(3) Bands of the (17)O(16)O(17)O Isotopomer of Ozone. AB - Using 0.002 cm(-1) resolution Fourier transform absorption spectra of an (17)O enriched ozone sample, an extensive analysis of the nu(3) band together with a partial identification of the nu(1) band of the (17)O(16)O(17)O isotopomer of ozone has been performed for the first time. As for other C(2v)-type ozone isotopomers [J.-M. Flaud and R. Bacis, Spectrochim. Acta, Part A 54, 3-16 (1998)], the (001) rotational levels are involved in a Coriolis-type resonance with the levels of the (100) vibrational state. The experimental rotational levels of the (001) and (100) vibrational states have been satisfactorily reproduced using a Hamiltonian matrix which takes into account the observed rovibrational resonances. In this way precise vibrational energies and rotational and coupling constants were deduced and the following band centers nu(0)(nu(3)) = 1030.0946 cm(-1) and nu(0)(nu(1)) = 1086.7490 cm(-1) were obtained for the nu(3) and nu(1) bands, respectively. Copyright 2000 Academic Press. PMID- 10708538 TI - High-Resolution Study of the BaI A(2)Pi Electronic State. AB - Near-infrared and visible spectra of the A(2)Pi-X(2)Sigma(+), C(2)Pi(1/2) A(2)Pi(1/2), C(2)Pi(1/2)-B(2)Sigma(+), and C(2)Pi(1/2)-X(2)Sigma(+) band systems of the BaI molecule were recorded by using Fourier transform spectroscopy (FTS). The spectra were produced from the chemiluminescent reaction Ba + I(2) and also by using laser-induced fluorescence (LIF) technique in which the laser sources were a Ti:sapphire single-mode laser, a dye single-mode laser, and a Kr(+) multimode ion laser. Resolved rotational data, originating from 19 vibrational levels (0 Cys at 88alpha (rHb A88alphaC), Cys-->Ala at 93beta (rHb C93betaA) and Cys-->Thr at 93beta (rHb C93betaT). These Hbs showed increased oxygen affinities and impaired allosteric effects. The spectral data indicated that the R to T transition upon deoxygenation was partially restricted in these Hbs. The number of titratable -SH groups of liganded form was 3.2-3.5 for rHb A88alphaC compared with 2.2 for Hb A, whereas those for rHb C93betaA and rHb C93betaT were negligibly small. The reduction of rate of reaction with 4,4'-dipyridine disulfide upon deoxygenation in rHb A88alphaC was smaller than that in Hb A. Our experimental data have shown that the residues at 88alpha and 93beta have definite roles but they have no functional homology. Structure-function relationships in our mutant Hbs are discussed. PMID- 10708651 TI - Protein engineering of cytochrome p450(cam) (CYP101) for the oxidation of polycyclic aromatic hydrocarbons. AB - Mutations of the active site residues F87 and Y96 greatly enhanced the activity of cytochrome P450(cam) (CYP101) from Pseudomonas putida for the oxidation of the polycyclic aromatic hydrocarbons phenanthrene, fluoranthene, pyrene and benzo[a]pyrene. Wild-type P450(cam) had low (<0.01 min(-1)) activity with these substrates. Phenanthrene was oxidized to 1-, 2-, 3- and 4-phenanthrol, while fluoranthene gave mainly 3-fluoranthol. Pyrene was oxidized to 1-pyrenol and then to 1,6- and 1,8-pyrenequinone, with small amounts of 2-pyrenol also formed with the Y96A mutant. Benzo[a]pyrene gave 3-hydroxybenzo[a]pyrene as the major product. The NADH oxidation rate of the mutants with phenanthrene was as high as 374 min(-1), which was 31% of the camphor oxidation rate by wild-type P450(cam), and with fluoranthene the fastest rate was 144 min(-1). The oxidation of phenanthrene and fluoranthene were highly uncoupled, with highest couplings of 1.3 and 3.1%, respectively. The highest coupling efficiency for pyrene oxidation was a reasonable 23%, but the NADH turnover rate was slow. The product distributions varied significantly between mutants, suggesting that substrate binding orientations can be manipulated by protein engineering, and that genetic variants of P450(cam) may be useful for studying the oxidation of polycyclic aromatic hydrocarbons by P450 enzymes. PMID- 10708652 TI - Structural adaptation to selective pressure for altered ligand specificity in the Pseudomonas aeruginosa amide receptor, amiC. AB - The AmiC protein in Pseudomonas aeruginosa is the negative regulator and ligand receptor for an amide-inducible aliphatic amidase operon. In the wild-type PAC1 strain, amidase expression is induced by acetamide or lactamide, but not by butyramide. A mutant strain of P. aeruginosa, PAC181, was selected for its sensitivity to induction by butyramide. The molecular basis for the butyramide inducible phenotype of P.aeruginosa PAC181 has now been determined, and results from a Thr-->Asn mutation at position 106 in PAC181-AmiC. In the wild-type PAC1 AmiC protein this residue forms part of the side wall of the amide-binding pocket but does not interact with the acetamide ligand directly. In the crystal structure of PAC181-AmiC complexed with butyramide, the Thr-->Asn mutation increases the size of the ligand binding site such that the mutant protein is able to close into its 'on' configuration even in the presence of butyramide. Although the mutation allows butyramide to be recognized as an inducer of amidase expression, the mutation is structurally sub-optimal, and produces a significant decrease in the stability of the mutant protein. PMID- 10708653 TI - Structural characterization of protein-denaturant interactions: crystal structures of hen egg-white lysozyme in complex with DMSO and guanidinium chloride. AB - A variety of physico-chemical methods employ chemical denaturants to unfold proteins, and study different biophysical processes involved therein. Chemical denaturants are believed to induce unfolding by stabilizing the unfolded state of proteins over the folded state, either macroscopically or through specific interactions. In order to characterize the nature of specific interactions between proteins and denaturants, we have solved crystal structures of hen egg white lysozyme complexed with denaturants, and report here dimethyl sulfoxide and guanidinium chloride complexes. The dimethyl sulfoxide molecules and guanidinium ions were seen to bind the protein at specific sites and were involved in characteristic interactions. They share a major binding site between them, the C site in the sugar binding cleft of the enzyme. Although the overall conformations of the complexes were very similar to the native structure, spectacular conformational changes were seen to occur locally. Temperature factors were also seen to drop dramatically in the local regions close to the denaturant binding sites. An interesting observation of the present study was the generation of a sodium ion binding site in hen egg-white lysozyme in the presence of denaturants, which was hitherto unknown in any of the other lysozyme structures solved so far. Loss of some of the crucial side chain-main chain interactions may form the initial events in lysozyme unfolding. PMID- 10708654 TI - Prospects for pneumococcal vaccination in African children. AB - Streptococcus pneumoniae (pneumococcus) remains a major cause of morbidity and mortality in both developed and undeveloped countries. Accurate disease burden estimates for developing countries and Africa in particular, where diagnostic facilities are less adequate and a disease surveillance system virtually non existent, is difficult. However, from conservative estimates, the pneumococcus is probably responsible for at least 1 million of the 4 million deaths that occur from acute lower respiratory infections in children aged less than 5 years. The global burden of disease has been accentuated by the rising menace of multi-drug resistant strains, which defy geographic and racial borders. Thus, now more than ever before, there is an urgent need to identify and implement preventive measures to avert this problem. The currently licensed pneumococcal polysaccharide vaccine, comprises 23 capsular polysaccharides of the pneumococcus, many of which are poorly immunogenic in the very vulnerable age group of under-fives. A possible solution to the problem of poor immunogenicity is to use a protein/polysaccharide conjugate vaccine similar to that recently introduced successfully for Haemophilus influenzae type b (Hib) and using this approach, several workers have reported promising results from safety and immunogenicity studies. However, unlike Hib, the development of conjugate vaccine against pneumococcal disease is complicated by the existence of more serotypes than can be feasibly incorporated in a single conjugate vaccine formulation. Whilst this challenge has been taken on by some vaccine manufacturers, novel approaches such as the identification or construction of protective protein antigen, common to all clinically important strains are being explored. Novel application of the pneumococcal polysaccharide vaccines in pregnancy for protection of disease in early infancy is an approach that has not been evaluated. For maximum impact, the ultimate vaccine formulation should be affordable and available to resource poor countries where the burden of disease is highest. Establishing disease surveillance systems in such countries now will greatly facilitate the introduction of the vaccines. PMID- 10708655 TI - Aetiology of visceral leishmaniasis in Mexico. AB - Two children with visceral leishmaniasis (VL), were studied by DNA analysis. DNA from liver biopsy samples from both patients, was amplified by PCR with broad primers specific for the Leishmania subgenus. DNA from the patient from Chiapas was also amplified with primers specific for the Leismania donovani complex and hybridised with a probe specific for L. donovani complex. The second patient, who is the first reported case of visceral leishmaniasis in the Mexican state of Tabasco, where localised cutaneous leishmaniasis and DCL predominate, had a co infection with Toxoplasma gondii. The DNA from this patient was not amplified with primers specific for the L. donovani complex, did not hybridise with a probe specific for the L. donovani complex, but did hybridise with kDNA from a Mexican Leishmania mexicana strain used as a probe. We therefore, suggest that members of the L. donovani or L. mexicana complexes cause VL in Mexico. PMID- 10708656 TI - A study of the urban malaria transmission problem in Khartoum. AB - A study of malaria prevalence and transmission was carried out in Khartoum, the capital of Sudan. The sentinel sites were El manshia, an urban area on the Blue Nile and Ed dekheinat, a lower-income peri-urban area bordering the White Nile. Anopheles arabiensis, the only malaria vector encountered, was present throughout the year although vector density varied seasonally. Plasmodium falciparum was the only species found in El manshia. In Ed dekheinat P. falciparum, Plasmodium ovale and Plasmodium vivax constituted 84.9, 8.2 and 6.9% of the cases, respectively. Plasmodium ovale appears to have recently spread into Khartoum since it has not previously been reported there. We conclude that focal transmission of malaria in the districts bordering both Niles has become established and that the reservoir of human infections has increased in recent years leading to increased risk of malaria epidemics, particularly in the aftermath of seasonal flooding. PMID- 10708657 TI - Malaria control in central Malaita, Solomon Islands. 1. The use of insecticide impregnated bed nets. AB - The present study investigated the use of insecticide-impregnated bed nets by communities in central Malaita, Solomon Islands. Qualitative and quantitative data were collected by: (1) questionnaire administration to 124 care-givers of children aged 0-10 years of age; (2) 20 focus group discussions; (3) two structured observations of bed net re-impregnation, and (4) interviews with key informants. Ninety-four percent of all care-givers had bed nets, but only 62% had sufficient bed nets for all household members. Fifty-two percent used bed nets throughout the year and 70% of care-givers reported that all their children slept under bed nets. Although coastal householders considered malaria and mosquitoes more of a problem than inland householders, overall bed net compliance did not differ. Factors affecting bed net ownership were cost and community expectation of free bed nets. Bed net use was affected by four factors: (1) seasonality (99% used bed nets during the rainy season, 52% used them all year); (2) mosquito nuisance (59% of respondents reported that protection against mosquitoes was the main reason for using a bed net); (3) weather (68% of care-givers would not use a bed net if the weather was hot), and (4) low density of mosquitoes (respondents who used bed nets as protection against mosquito nuisance were more likely not to use bed nets when mosquitoes were few than those who used bed nets for malaria protection (odds ratio (OR), 3.9; 95% confidence interval (CI), 1.4-12.0). Protection against malaria was the main reason children slept under bed nets. Children from households where bed nets were used for malaria protection were more likely to sleep under bed nets than children from households where nets were used as protection from mosquitoes only (OR, 2.7; 95% CI, 1.3-5.9). Other factors that affected children's bed net use were, age (users were significantly younger than non-users; chi(2)=7.9, degrees of freedom=1, P=0.005) and sufficiency of bed nets (OR, 2.0; 95% CI, 1. 3-7.0). PMID- 10708658 TI - Malaria control in central Malaita, Solomon Islands 2. Local perceptions of the disease and practices for its treatment and prevention. AB - Government health policy for malaria control in Solomon Islands has three main objectives: (1) early diagnosis and treatment of malaria at a health service; (2) reduction of human-vector contact through widespread use of insecticide impregnated bed nets; and (3) provision of malaria chemoprophylaxis for pregnant women. Social research was carried out in thirteen villages in central Malaita to determine local attitudes toward malaria and to estimate the level of participation in malaria control activities. Interviews with 124 care-givers who had children 0-10 years of age, 20 focus group discussions and four evening structured observations were research methods used. Antimalarial drugs were the most favoured treatment, and use of traditional medicines and healers were reportedly minimal. Twenty-five percent of respondents reported keeping chloroquine at home and 42% said they would use chloroquine before seeking diagnosis and treatment from a health service. Structured observations suggest that protection against mosquitoes is poor during the evening. Fifty-two percent of respondents reported using fire and 32% said they used bed nets to protect themselves from mosquitoes. Participants had contradictory beliefs on the threat of malaria during pregnancy and the safety of taking chloroquine prophylaxis. Implications of malaria treatment and prevention practices are discussed, and recommendations for improving malaria control are presented. PMID- 10708659 TI - PAIR as percutaneous treatment of hydatid liver cysts. AB - Hydatid disease of the liver remains an important and challenging problem in rural areas; although, surgery is considered the treatment of choice, percutaneous treatment of hydatid cysts is relatively new, and the data related to it are limited. The purpose of the study was to present the results of percutaneous treatment of liver hydatid cysts. Thirty-four patients (13 male and 21 female), ranging in age between 14 and 80 years, with 55 liver hydatid cysts underwent percutaneous treatment with albendazole prophylaxis. Cysts were treated with a one-stage procedure that consisted of puncture of the cysts under guidance with computed tomography, aspiration of fluid, injection of hypertonic saline solution as scolicidal agent and reaspiration. Follow-up examinations showed progressive reduction and solidification of the cysts. The mean reduction in volume was 72%. No mortality, abdominal dissemination, or tract seeding occurred. Minor complications were urticaria with pruritus in two patients. One patient had a subcapusular hematoma without problem. Hospitalization courses varied from ambulatory procedures to 15 days of in-patient, mean hospital stay was 1.82 days. The results of percutaneous liver hydatid cyst treatment, indicating that the procedure is efficient and safe and offers complete cure in selected patients with a short hospitalization and that this technique should be considered an alternative to surgery. PMID- 10708660 TI - Plasticity of the histone H2A genes in a Brazilian and six Colombian strains of Trypanosoma cruzi. AB - The analysis of three recombinant clones containing the histone H2A locus isolated from a genomic library of Trypanosoma cruzi DNA shows that the H2A gene loci are formed by 1.2 and 0.76 kb long intercalated units organized in a head-to tail tandem array. The difference in length between the two gene units is due to the presence of a short interspersed nucleotide element (SINE)-like DNA sequence inserted at the 3' end of some of these units. Southern, northern and chromosomal blot analysis of a Brazilian Y strain and six Colombian strains demonstrated the existence of polymorphisms regarding the relative copy number of the H2A gene units, the relative abundance of the H2A transcripts and their chromosomal location. These results show the existence of a dynamic organization in the H2A loci among T. cruzi strains in which a SINE-like sequence may be involved and support the fact that T. cruzi has a high degree of plasticity in its genome. PMID- 10708661 TI - Changes of anti-Trypanosoma cruzi antibodies after gamma-irradiation of mice in the chronic phase of the infection. AB - The effect of sublethal whole body irradiation (800 rads) on the level and biological activities of antibodies in mice chronically infected with the CL strain of Trypanosoma cruzi was studied. Irradiated mice died, although a high parasitemia did not always preceded death. Before and after irradiation, a constant level of antibodies was detected by enzyme-linked immunosorbent assay and complement mediated lysis, but after irradiation the level of clearance antibodies was decreased. These results suggest that clearance antibodies are important in the control of the chronic phase of the infection. PMID- 10708662 TI - Effect of the alkyl-lysophospholipids on the proliferation and differentiation of Trypanosoma cruzi. AB - Alkyl-lysophospholipids (ALPs), designed as potential immunomodulators, have been shown to be cytotoxic for a variety of tumour cells and are under clinical studies for cancer chemotherapy. ET-18-OCH(3), hexadecylphosphocholine and ilmofosine were assayed against the three forms of Trypanosoma cruzi. Incubation with bloodstream trypomastigotes resulted, under different experimental conditions, in higher activity of the compounds in comparison with crystal violet. The ED(50)/24 h values were 13.4+/-2.8 microM and 11. 7+/-0.6 microM for amastigotes and epimastigotes, respectively. ET-18-OCH(3) (0.3 and 0.6 microM) inhibited the differentiation of epimastigotes to trypomastigotes (Dm28C clone) in the range 40-57%. This drug (3.75-15 microM) also caused a time- and dose dependent inhibition of the intracellular proliferation of amastigotes in heart muscle cells with ED(50) values of 14.3+/-4.2, 8.9+/-1.9 and 6. 8+/-0.4 microM, after 1, 2 and 3 days of treatment. Pre-treatment of the parasite with this drug inhibited its interiorization into the host cell. Interestingly, the intracellular differentiation of amastigotes to trypomastigotes was not hampered by the drug. The present results demonstrate the lytic effect of ALPs on the three forms of T. cruzi, as well as the inhibition of both the differentiation to the infective form and the proliferation of parasites interiorized in heart cells. Ultrastructural analysis of epimastigotes treated with the three ALPs showed extensive blebing of the flagellar membrane. As described in tumour cells, the membrane seems to be a primary target of the drugs. PMID- 10708663 TI - Some biochemical changes following Trypanosoma congolense infection in Djallonke ewe lambs and breeding ewes fed on two levels of nutrition. AB - The effects of artificial Trypanosoma congolense infection and dietary level on biochemical changes were observed in 24 ewe lambs (Experiment 1) and 42 breeding ewes (Experiment 2). All animals belonged to the Djallonke breed which is known to be trypanotolerant. For both experiments, there were four treatment combinations, of which two were kept on a restricted diet (L), the other two on an at libitum diet (H). Half of each dietary group was infected with T. congolense (LI, HI), while the remainder served as uninfected controls (LC, HC). Artificial T. congolense infection took place at the age of 200+/-7 days in Experiment 1 and at the peak of oestrus in Experiment 2. Irrespective of dietary levels offered, total proteins in lambs and ewes and albumin in lambs declined significantly (P<0.001) post infection. Plasma glucose concentration was reduced by the low dietary level and not by infection. Although plasma urea concentrations were slightly increased in the infected ewe lambs, adult ewes in the HI group demonstrated increased plasma urea concentrations (P<0.05) due to an interaction between infection and diet. Neither infection nor the imposed diet induced significant changes on plasma creatinine concentrations. Transitory peaks in non-esterified fatty acids (NEFA) and beta-hydroxy butyric acid (BHBA) levels in infected ewes on low dietary level indicated temporary changes in the energy metabolism of the host. It was concluded from this study that, inspite of their trypanotolerance, Djallonke lambs and ewes demonstrated an infection effect on host metabolism pattern due to T. congolense infection. These changes reflected to some extent trypanosome-induced alteration of the nutrient metabolism, which could not always be negated by diet supplements. Nutrition, as an independent factor, did confer added benefits against the debilitating effects of trypanosomosis under the conditions of the present study. PMID- 10708664 TI - Efficacy of aqueous suspension and granular formulations of Bacillus thuringiensis (Vectobac) against mosquito vectors. AB - The efficacy of aqueous suspension (AS) and granular (G) formulation of Bacillus thuringiensis var. israelensis (Vectobac) was tested against the immatures of mosquito vectors in the laboratory and under field conditions. Laboratory tests showed that the aqueous suspension was relatively more effective against Culex quinquefasciatus than Aedes aegypti and Anopheles stephensi, the respective LC(50) values being 0.046, 0.060 and 0.190 mg/l. In stream pools, with the application of Vectobac AS at 1.2 l/ha, more than 80% reduction in immature density of Anopheles larvae (Anopheles fluviatilis and Anopheles culicifacies) was observed for 2-8 days, and at 2.4 l/ha for 3.5-9.0 days. At the dosage of 7.0 kg/ha of the granular formulation, a reduction in immature density by more than 80% was observed for 2-9 days. In polluted habitats such as cesspits, U-drains and cement tanks, the effectiveness of Vectobac AS lasted for 1-4 days when applied at 1.2 and 2.4 l/ha, and Vectobac G was effective for 1-3 days at application rates of 7. 0 and 14.0 kg/ha against C. quinquefasciatus. There was no significant difference in the effectiveness between the two formulations and the two application rates. PMID- 10708665 TI - An analysis of trypanocidal drug use in the Eastern Province of Zambia. AB - As part of the development of a strategy for the control of bovine trypanosomosis in Zambia, a survey was conducted to quantify and qualify the current use of trypanocidal drugs (diminazene aceturate and isometamidium chloride) in a tsetse controlled and a tsetse-infested area of the Eastern Province. A total of 207 trypanocide users were interviewed. Questions were posed on herd structure, trypanocidal drug preference, treatment strategy, reason for treatment, method of treatment and treatment frequency. The majority of the cattle owners preferred to use diminazene aceturate rather than isometamidium chloride. Both trypanocides were mainly used to treat clinically sick animals (not necessarily infected with trypanosomes) and preference was given to the treatment of oxen and cows. The proportion of animals treated and the frequency of drug application did not differ between the two areas. Hence, in the tsetse-controlled area, a high proportion of the trypanocide treatments was inappropriate. In the tsetse infested area, on the other hand, the treatment of clinically sick animals significantly reduced the trypanosomosis-related mortality but was insufficient to boost reproduction in cows. Despite the fact that the cattle owners administered most trypanocides themselves, evidence from the survey suggests that most of the farmers did not under-dose with either diminazene aceturate or isometamidium chloride. Moreover, other factors enhancing the development of resistance to trypanocides in trypanosomes were not present in the areas surveyed. Conclusions are drawn on the usefulness of this type of survey in determining appropriate methods to control bovine trypanosomosis. PMID- 10708666 TI - The effect of minocycline on the metabolism of androgens by human oral periosteal fibroblasts and its inhibition by finasteride. AB - The antimicrobial minocycline has matrix-stimulatory effects on connective tissue and bone. The aim here was to study the effect of minocycline on 5alpha reduction of androgen substrates to 5alpha-dihydrotestosterone (DHT) in periosteal fibroblasts and the influence of the antiandrogen finasteride on this conversion. Confluent cultures of periosteal fibroblasts established from oral periosteum isolated from the bone surface were incubated in duplicate in multiwell dishes with two androgen substrates, [(14)C]-testosterone/[(14)C]-4-androstenedione, in the presence or absence of serial concentrations of minocycline or the antiandrogen finasteride or the two in combination for 24 h. The metabolites formed were solvent-extracted with ethyl acetate, separated by thin-layer chromatography and quantified using a radioisotope scanner. Both androgen substrates were metabolized to DHT and 4-androstenedione or testosterone. Minocycline stimulated the synthesis of DHT from these substrates by 75-83% at 20 30 microg/ml (n=4; p<0.01). Finasteride inhibited the 5alpha-reductase activity of these substrates by 3-5-fold at 1 microg/ml and 40-80% at 0.01 and 0.1 microg/ml (n=4; p<0.01), with little change in 17beta-hydroxysteroid dehydrogenase activity. Minocycline and finasteride in combination showed an intermediate response with one substrate. As finasteride inhibits the type 2, 5alpha-reductase isoenzyme associated with anabolic functions, these findings demonstrate target-tissue androgen metabolic activity in periosteal fibroblasts at baseline and in response to minocycline. This has implications for the reparatory potential of the diseased periodontium during adjunctive treatment with minocycline. PMID- 10708667 TI - Associations between incisor and mandibular condylar movements during maximum mouth opening in humans. AB - This study evaluated the common clinical assumption that condylar translation and mouth opening at the incisor are closely related. The sample studied comprised 27 adult females (23-35 years), selected for normal temporomandibular function, occlusion, and skeletal patterns. Condylar and mandibular central incisor movements (straight-line distances and curvilinear pathways) were recorded in three dimensions (3D) for 20 s with an optoelectric (Optotrak(R)) jaw-tracking system while each participant performed multiple maximum opening cycles. Masticatory analysis and multilevel statistical programs computed the 3D movements of the incisors and condylar hinge axis during opening and closing. The incisor point moved an average straight-line distance of 46.6 mm during opening and 45.9 mm during closing; the lengths of the opening and closing curvilinear pathways were 48.6 and 47.7 mm, respectively. The condyles moved average straight line distances of 11.9 and 12.2 mm during opening and closing, respectively. The condyles' curvilinear pathways during opening and closing were 14 and 14.6 mm, respectively. Ranges of condylar movement varied widely between individuals. The straight-line distances and curvilinear pathways were highly correlated for the incisors (R=0.98) and the condyles (R=0.98). Neither the straight-line distances nor curvilinear pathways of the incisors were correlated with those of the condyles. Incisor straight-line distances and curvilinear pathways were moderately correlated with mandibular rotation (R(between subjects)=0.82 and R(between repeats)=0.88). During repeated openings, both condylar and incisor excursions increased, but during repeated closings only incisor excursions increased. It is concluded that (1) maximum incisor opening does not provide reliable information about condylar translation and its use as a diagnostic indicator of condylar movement should be limited, (2) healthy individuals may perform normal opening with highly variable amounts of condylar translation, (3) the straight-line distances of the incisor and condyles provide adequate information about the length of the curvilinear pathway, and (4) variation in maximum incisor opening is largely explained by variation in the amount of mandibular rotation. PMID- 10708668 TI - In situ studies of pellicle formation on hydroxyapatite discs. AB - The formation of acquired enamel pellicle on hydroxyapatite (HA) discs of known surface area carried in the mouth was studied; discs were carried in the mouth for 30 s, 1, 5, 10 and 20 min. Similar amounts of protein were found on the discs at each time-point, as determined by ninhydrin analyses. The amounts of amylase and lysozyme detected remained stable after 5 min of exposure of the discs to the mouth. Assay of the discs for fructosyl- and glucosyltransferase activities revealed that fructosyltransferase activity increased up to 1 min of exposure to the mouth and decreased when kept in the mouth for longer periods; glucosyltransferase activity, in contrast, increased the longer the discs were kept in the mouth. This in situ model provides insight into the activities of various enzymes during the first 20 min of pellicle formation. The effects of rinsing with sucrose and sugar alcohols on pellicle formation on the discs were also explored. The discs were placed in the mouth for 30 s, 1, 5, 10 and 20 min, preceded by rinsing with either distilled deionized water, sucrose, sorbitol, xylitol or phosphate-buffered saline. Western blot analyses of disc eluates with antiserum/antibody preparations to various salivary components revealed distinct patterns of deposition of bacterial and salivary components depending on the composition of the rinse. These studies confirm that salivary molecules and bacteria are deposited on apatitic surfaces in a selective manner and reveal that pellicle formation may be influenced by composition of diet. It is apparent that this in situ model could be used in screening potential antiplaque agents. PMID- 10708669 TI - Response of immortalized murine cementoblasts/periodontal ligament cells to parathyroid hormone and parathyroid hormone-related protein in vitro. AB - Cementum is an essential component of the periodontium, but the mechanisms involved in regulating the activity of this tissue are poorly understood. As one approach to better defining the cellular and molecular properties of cementum and the associated ligament, immortalized murine cell populations expressing gene markers associated with both cementoblasts (CM) and periodontal ligament cells (PDL), termed CM/PDL cells, were established. To further characterize these cells, their responsiveness to parathyroid hormone (PTH) and parathyroid hormone related protein (PTHrP) was examined. CM/PDL cells were tested for the presence of steady state PTH-1 receptor mRNA using Northern blot analysis. In addition, the ability of PTH and PTHrP to stimulate cAMP production and c-fos mRNA expression in CM/PDL cells was determined, using a cAMP-binding assay and northern blot hybridization, respectively. Rat osteosarcoma cells (ROS 17/2.8) were used as a positive control and human periodontal ligament cells as a negative control. Northern blot analysis demonstrated that cells within the CM/PDL cell population expressed PTH-1 receptor mRNA. Both PTH (1-34) and PTHrP (1-34) increased cAMP and c-fos mRNA in CM/PDL cells. Furthermore, PTHrP treatment for either 24 or 48 h downregulated expression of transcripts for bone sialoprotein, osteocalcin and PTH-1 receptor by CM/PDL cells and abolished CM/PDL cell-mediated mineralization in vitro. These results indicate that cells within the CM/PDL population are targets for PTH and PTHrP action and that PTHrP may play an important part in regulating the biomineralization of cementum. PMID- 10708671 TI - A histological study of root-resected and root-transected rat incisors when eruption ceases, shortly before they are exfoliated from the socket. AB - Resection of the odontogenic region or root transection of normal (impeded) rat lower incisors showed that eruption ceased from 1 to 13 weeks when the base of the resected teeth (87.5%) or of the distal segment of the transected ones (86%) reached the alveolar-crest region. When the operated teeth reached the crestal region, the enamel-related periodontal tissues were absent and the periodontal ligament (PDL) was the only periodontal tissue that remained. The PDL of the crestal region may be considered as mature PDL, showing a length of approx. 5-6 6 mm at the mesial face of the tooth, 4-5 mm at lingual face and 1 mm at distal face; from these limits towards the apical end of the socket the PDL becomes gradually immature. The mature PDL seems not to have a role in the process of tooth eruption. Several factors can be suggested to explain the more frequent retention, at the crestal region of the socket, of impeded rather than unimpeded incisors submitted to the same procedures. The connective tissue that develops between the base of the tooth and the bone that fills the alveolus may have more time to organize itself in impeded than in unimpeded teeth, which erupt at a faster rate; this tissue could support and retain the impeded operated teeth longer than the unimpeded ones. The decrease in the mechanical properties of the PDL in the unimpeded condition may ease the traumatic effects and lead to exfoliation. Eruption might be stopped by the increase in occlusal forces, per unit area of root surface, as the root becomes shorter; this effect is likely to be greater in impeded than unimpeded teeth. PMID- 10708670 TI - Active detachment of Streptococcus mutans cells adhered to epon-hydroxylapatite surfaces coated with salivary proteins in vitro. AB - Although the formation of biofilms has been much studied, detachment of adherent cells from biofilms has been relatively neglected. Recent results have shown that adherent Streptococcus mutans cells can actively detach from epon-hydroxylapatite (EHA) rods conditioned with hog gastric mucin. The mechanisms for adherence and detachment of Strep. mutans cells in this system was uncertain. In the present study, resting Strep. mutans cells were used to form a simple monolayer on EHA rods coated with saliva and salivary agglutinin (SAG). Preliminary experiments defined the variables for conditioning EHA with saliva and SAG and establishing the adherence of Strep. mutans to the conditioned surfaces. The results showed that salivary proteins including SAG adsorbed rapidly to EHA and that a relatively stable Strep. mutans NG8 monolayer was formed within 60 min of incubation. The monolayers were subsequently used for detachment studies. The results showed that adherent Strep. mutans cells detached in a temperature dependent manner and responded to the addition of a preparation of surface protein-releasing enzyme (SPRE) obtained from Strep. mutans in a dose-dependent fashion. The effect of the exogenous SPRE on detachment could be abrogated by pronase treatment. Two putative SPRE-defective mutants (A and E) were generated by Tn917 mutagenesis. Both mutants possessed a single transposon insertion as demonstrated by Southern hybridization and appeared to be different from one another based on the hybridization patterns. Mutant A displayed an increased quantity of cell-surface antigen P1, an adhesin that interacts with SAG. At the same time mutant A was unable to release P1 and other high molecular-weight proteins from the cell surface. Mutant A detached at a significantly lower rate (21%) than the parent strain (37%) (p=0.05). SPRE prepared from mutant A was unable to release Strep. mutans NG8 adherent cells as compared to SPRE obtained from the wild-type cells. Collectively, these results suggest that the detachment of Strep. mutans adherent cells formed on salivary protein-coated EHA was an active process mediated by the action of SPRE. PMID- 10708672 TI - The effects of local trauma to the enamel-related periodontal tissues in the eruption of the rat incisor. AB - The periodontal tissues related to enamel (PTE) of the rat incisor comprise a connective tissue derived from the dental follicle and the enamel organ with its successive stages of development. Localized damage to these tissues in rat lower incisors was done surgically in three ways: with an endodontic file introduced into the labial periodontal space through either (i) its basal or (ii) its incisal extremities, or (iii) by the partial removal of the mandibular lower border, at the level of the molar teeth, together with the introduction of an endodontic file into the incisal part of that space. The lesions in the molar region of the PTE produced first a variable period of retarded eruption, and, depending upon their extent or degree were followed by a cessation of the eruptive movement and, in the majority of the operated teeth, a recovery of the normal eruption rate before the end of the experiment (17 weeks after surgery). Access to the PTE through the basal portion of the socket was erratic, but when the tissues were damaged produced similar effects. Effects on eruption of lesions produced through the alveolar crest were minimal or even absent. Localized injury to the periodontal ligament of either lower or upper incisors did not produce similar effects on tooth eruption. The dental follicle and the enamel organ of teeth of limited growth when their crown is completed are similar to the PTE in the molar region of continuously growing rodent incisors. In teeth of limited growth these tissues play an essential part in the intraosseous stage of eruption. The results here suggest that the PTE may also have a role in the supraosseous stage of eruption, which is continuous in teeth such as rat incisors due to the presence of a continuously functioning odontogenic organ. PMID- 10708673 TI - Calcitonin gene-related peptide and substance P immunoreactivity in rat trigeminal ganglia and brainstem following adjuvant-induced inflammation of the temporomandibular joint. AB - The immunoreactivity of two inflammatory mediators, calcitonin gene-related peptide (CGRP) and substance P, was measured in the trigeminal ganglia and brainstem to characterize an adjuvant-induced inflammation within the rat temporomandibular joint at various acute (6, 24 and 48 h) and intermediate (10 day) time intervals. Concentrations of adjuvant-related neuropeptides were compared to those in both contralateral vehicle-related tissues and non-injected controls. By 6 h, CGRP immunoreactivity in the trigeminal ganglia was significantly above that in contralateral vehicle-injected tissue. The CGRP had decreased at each of the following time-points, but remained significantly elevated at 10 days. Substance P in the ganglion on the injected side was significantly increased for all four time periods. In brainstem subnucleus caudalis, CGRP was significantly increased for all four time periods. Substance P immunoreactivity in the subnucleus caudalis was significantly increased for the initial three time periods, but by day 10 had been reduced to that of the control. These data show that the pattern of changes in neuropeptides following the induction of inflammation is different between substance P and CGRP. Moreover, the pattern of change varies between the brainstem and the trigeminal ganglion. This suggests that the two neuropeptides may have different roles in the inflammatory process, and that this process may be modulated by different mechanisms at the brainstem and ganglion. PMID- 10708674 TI - Increased histamine release and granulocytes within the thalamus of a rat model of Wernicke's encephalopathy. AB - The current study examined the possible role of increased histamine release and granulocyte activity in the vascular changes that precede the onset of necrotic lesions with the thalamus of the pyrithiamine-induced thiamine deficiency (PTD) rat model of Wernicke's encephalopathy (WE). An increase in histamine release and the number of granulocytes was observed in lateral thalamus on day 9 and in medial thalamus on day 10 of PTD treatment, a duration of thiamine deficiency associated with perivascular edema in this brain region. Within the hippocampus, histamine release was significantly increased on day 9, declined to control levels on days 10-12, and was significantly elevated on days 12-14. No granulocytes were observed in hippocampus of either PTD or control rats. These observations suggest that the release of histamine from nerve terminals and histamine and other vasoactive substances from granulocytes may be responsible for thiamine deficiency-induced vascular breakdown and perivascular edema within thalamus. PMID- 10708675 TI - Distribution of beta-amyloid and amyloid precursor protein in the brain of spawning (senescent) salmon: a natural, brain-aging model. AB - Brain amyloid precursor protein (APP), a normal constituent of neurons, glial cells and cerebrospinal fluid, has several proposed functions (e.g., in neuronal growth and survival). It appears, however, that altered processing of APP is an initial or downstream step in the neuropathology of brain aging, Alzheimer's disease (AD), and Down's syndrome (DS). Some studies suggest that proteolytic cleavage of APP, producing beta-amyloid (Abeta(1-42)), could have neurotoxic or neuroprotective effects. In this study, we utilized antibodies to human APP(695) and Abeta(1-42,) and Congo red staining, to search for amyloid deposition in the brain of semelparous spawning kokanee salmon (Oncorhynchus nerka kennerlyi). Intracellular APP(695) immunoreactivity (APP-ir) was observed in brain regions involved in gustation (glomerulosus complex), olfaction (putative hippocampus, olfactory bulb), vision (optic tectum), the stress response (nucleus preopticus and nucleus lateralis tuberis), reproductive behavior (nucleus preopticus magnocellularis, nucleus preopticus periventricularis, ventral telencephalon), and coordination (cerebellum). Intra- and extra-neuronal Abeta(1-42) immunoreactivity (Abeta-ir) were present in all APP-ir regions except the nucleus lateralis tuberis and Purkinje cells of the cerebellum (coordination). Thus, the relationship between APP and Abeta deposition during brain aging could shed light on the processing of APP into Abeta, neurodegeneration, and possible protection of neurons that are functioning in spawning but senescent salmon. Pacific salmon, with their predictable and synchronized life history, could provide research options not available with the existing models for studies of brain aging and amyloidosis. PMID- 10708676 TI - The effect of serotonin(1A) receptor agonism on antipsychotic drug-induced dopamine release in rat striatum and nucleus accumbens. AB - Serotonin (5-HT)(1A) receptor agonism may be of interest in regard to both the antipsychotic action and extrapyramidal symptoms (EPS) of antipsychotic drugs (APD) based, in part, on the effect of 5-HT(1A) receptor stimulation on the release of dopamine (DA) in the nucleus accumbens (NAC) and striatum (STR), respectively. We investigated the effect of R(+)-8-hydroxy-2-(di-n-propylamino) tetralin (R(+)-8-OH-DPAT) and n-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-n-(2 pyridinyl)cyclohe xanecarboxamide trihydrochloride (WAY100635), a selective 5 HT(1A) receptor agonist and antagonist, respectively, on basal and APD-induced DA release. In both STR and NAC, R(+)-8-OH-DPAT (0.2 mg/kg) decreased basal DA release; R(+)-8-OH-DPAT (0.05 mg/kg) inhibited DA release produced by the 5 HT(2A)/D(2) receptor antagonists clozapine (20 mg/kg), low dose risperidone (0.01 and 0. 03 mg/kg) and amperozide (10 mg/kg), but not that produced by high dose risperidone (0.1 and 1.0 mg/kg) or haloperidol (0.01-1.0 mg/kg), potent D(2) receptor antagonists. This R(+)-8-OH-DPAT-induced inhibition of the effects of clozapine, risperidone and amperozide was antagonized by WAY100635 (0.05 mg/kg). WAY100635 (0.1-0.5 mg/kg) alone increased DA release in the STR but not NAC. The selective 5-HT(2A) receptor antagonist M100907 (1 mg/kg) did not alter the effect of R(+)-8-OH-DPAT or WAY100635 alone on basal DA release in either region. These results suggest that 5-HT(1A) receptor stimulation inhibits basal and some APD induced DA release in the STR and NAC, and that this effect is unlikely to be mediated by an interaction with 5-HT(2A) receptors. The significance of these results for EPS and antipsychotic action is discussed. PMID- 10708677 TI - X-linked transmission of the shaker mutation in rats with hereditary Purkinje cell degeneration and ataxia. AB - This study reports on the mode of inheritance of the shaker mutation and the development of an inbred strain of the shaker rat mutation from Sprague Dawley outbred stock onto a Wistar Furth background. Neuroanatomical and behavioral expression of the affected phenotype, through seven generations of backcross and intercross breeding, has confirmed the mode of inheritance to be X-linked. Behaviorally, affected mutants present with a wide-based ataxic gait and whole body tremor. In affected mutants calbindin immunostaining for surviving cerebellar Purkinje cells revealed widespread degeneration in the anterior lobe and in limited areas of the posterior lobe. Fast Fourier transform analysis of the tremor revealed a frequency of 3-5 Hz. As predicted by X-linked inheritance, female descendants of an affected male are carriers for the genotype and the phenotype is expressed in one-half of her male offspring. There was spatially random and limited degeneration of Purkinje cells in carrier females, but they did not display overt clinical signs of ataxia and tremor. These data provide further support for using the shaker mutant rat as an animal model for studies of mechanisms underlying human heredodegenerative diseases. PMID- 10708678 TI - Modulation of triceps surae H-reflexes as a function of the reflex activation history during standing and stepping. AB - The facilitatory effectiveness of spindle afferent feedback is controlled by modulation of segmental reflex excitability such that the level of muscle activation is appropriate for the task. Phase-dependent modes of reflex modulation have been well-characterized. We hypothesized that segmental reflex excitability of the triceps surae was also modulated in a manner associated with the activation history of the spindle afferents and the segmental reflex pathway during isometric contractions, standing and stepping. In the first experiment. pairs of soleus (S) H-reflexes were evoked 80 ms apart with equal strength stimuli at rest and while subjects isometrically contracted their S against loads of 10%. 20%. and 50% of their maximum voluntary efforts. The percent depression of the second H-reflex relative to the first was used as a measure of the effect of reflex activation history. At rest, the second H-reflexes were depressed an average of 73% relative to the first. The degree of depression was progressively reduced as the plantarflexion torque increased. In the second experiment, paired H-reflexes were obtained from the S and medial (MG) and lateral gastrocnemii (LG) muscles while subjects were standing and during the stance phase of step initiation. The degree of depression of the second H-reflex during standing ( > 78%) was similar in magnitude to that produced at rest in Experiment I. At the end of the stance phase of stepping. depression of the second H-reflex of all three muscles was reduced to less than 25%. We conclude that the segmental reflex excitability is modulated as a function of the reflex activation history during these tasks. PMID- 10708679 TI - Prenatal exposure to the dopamine agonist SKF-38393 disrupts the timing of the initial response of the suprachiasmatic nucleus to light. AB - The abuse of social drugs such as cocaine during pregnancy represents enormous risks to the offspring. Recent studies showed that drugs administered to the pregnant rat can activate cell populations in the fetal brain, possibly altering the timing of key neuronal developmental events. The current study examined the ontogeny of light-responsiveness of the neonatal rat suprachiasmatic nucleus using c-FOS protein in SCN nuclei as a marker. The effect of acute administration of the dopamine D1 agonist, SKF-38393, on the development of light responsiveness was also examined. Pregnant dams received either SKF-38393 (10 mg/kg) or vehicle 7 h after dawn on gestational day 20. Litters were then assigned to one of seven experimental time points from 4 h after subjective dark onset on the day of birth (P0-CT16) at 4-h intervals until CT16 on the day after birth (P1-CT16). Half of the pups in each litter were exposed to a 200 lux/2 h light pulse and the other half remained in darkness. Three time points (P1-CT0, P1-CT8 and P1-CT16) were used to examine the prenatal drug effects on light-responsiveness. Light exposure at the time of subjective lights on, the day after birth (P1-CT0), resulted in a significant increase in c-FOS-positive cells. The number of positive cells recorded in the SCN after a light pulse at P1-CT0 and P1-CT8 was significantly less in SKF-38393 pretreated pups compared to vehicle treated animals. The exposure to dopaminergic stimulation during gestation may have altered the timing of development of afferent connections to the fetal SCN, resulting in alteration of the initial response of the circadian timing system to light. PMID- 10708680 TI - Spatio-temporal profile of DNA fragmentation and its relationship to patterns of epileptiform activity following focally evoked limbic seizures. AB - The specific electrographic activity responsible for seizure-induced DNA damage remains little explored. We therefore examined the regional and temporal appearance of DNA fragmentation and cell death and its relationship to specific electrographic seizure patterns in a rat model of focally evoked limbic epilepsy. Animals received intra-amygdaloid injection of kainic acid (KA) to induce seizures for 45 min during continuous electroencephalographic (EEG) monitoring, after which diazepam (30 mg/kg) was administered. DNA polymerase I-mediated biotin-dATP nick translation (PANT) and terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL) were used to detect single- and double stranded DNA breaks, respectively. Injection of 0.01 microg KA induced seizures characterized by ictal fast activity but without consequent brain injury. By contrast, 0.1 microg KA induced an additional pattern of seizure activity characterized by bursts of high frequency polyspike paroxysmal discharges. In these animals, there was a significant reduction in numbers of pyramidal neurons within the ipsilateral and contralateral CA3 subfield of the hippocampus, detectable as little as 4 h following seizures. PANT- and TUNEL-positive cells appeared in similar numbers 16 h following seizure cessation within the CA3, declining after 72-96 h. Varying the duration of polyspike paroxysmal discharges determined that as little as 30 s elicited maximal injury. These data suggest single- and double-stranded DNA breaks are generated during the cell death process and are consequent on a specific component of seizure activity electrographically determined. PMID- 10708681 TI - GM1 ganglioside treatment protects against long-term neurotoxic effects of neonatal X-irradiation on cerebellar cortex cytoarchitecture and motor function. AB - Exposure of neonatal rats to a 5 Gy dose of X-irradiation induces permanent abnormalities in cerebellar cortex cytoarchitecture (disarrangement of Purkinje cells, reduction of thickness of granular cortex) and neurochemistry (late increase in noradrenaline levels), and motor function (ataxic gait). The neuroprotective effects of gangliosides have been demonstrated using a variety of CNS injuries, including mechanical, electrolytic, neurotoxic, ischemic, and surgical lesions. Here, we evaluated whether systemically administered GM1 ganglioside protects against the long-term CNS abnormalities induced by a single exposure to ionizing radiation in the early post-natal period. Thus, neonatal rats were exposed to 5 Gy X-irradiation, and subcutaneously injected with one dose (30 mg/kg weight) of GM1 on h after exposure followed by three daily doses. Both at post-natal days 30 and 90, gait and cerebellar cytoarchitecture in X irradiated rats were significantly impaired when compared to age-matched controls. By contrast, both at post-natal days 30 and 90, gait in X-irradiated rats that were treated with GM1 was not significantly different from that in non irradiated animals. Furthermore, at post-natal day 90, cerebellar cytoarchitecture was still well preserved in GM1-treated, X-irradiated animals. GM1 failed to modify the radiation-induced increase in cerebellar noradrenaline levels. Present data indicate that exogenous GM1, repeatedly administered after neonatal X-irradiation, produces a long-term radioprotection, demonstrated at both cytoarchitectural and motor levels. PMID- 10708682 TI - Subarachnoid hemolysate produces DNA fragmentation in a pattern similar to apoptosis in mouse brain. AB - Stroke and traumatic brain/spinal cord injuries are often associated with hemorrhage. Despite the relative frequency of hemorrhage in the central nervous system (CNS), little is known about what role blood and hemoglobin (Hb) play in mediating cellular injury. Since Hb and hemolysate have been associated with generation of oxidative stress and cell injury, we examined whether apoptosis was present after cortical exposure to subarachnoid hemolysate. Subarachnoid hemorrhage (SAH) was induced in CD-1 mice (n=25) by injection of 50 microl of autologous hemolysate over the right parietal cortex. Saline-injected mice (n=13) were used as controls. Subjects were sacrificed at 24 h. Transcardiac perfusion fixation was performed on a subgroup of hemolysate- (n=15) and saline-injected (n=9) animals. Sections were stained for DNA fragmentation using the terminal deoxyuridine nick end-labeling (TUNEL) method and also immunostained for the hemeoxygenase-1 (HO-1) protein to assess blood distribution. In the remaining animals (n=6 SAH, n=4 saline), DNA was extracted and precipitated from 40 mg of tissue and subjected to electrophoresis on a 1.5% agarose gel. DNA fragmentation was evident on TUNEL staining in 10/15 subjects injected with hemolysate as compared to 0/9 subjects injected with saline (p<0.01, Fisher exact test). TUNEL positive cells were most abundant closest to the site of cortical SAH, as evidenced by HO-1 immunoreactivity. TUNEL-positive cells were also seen remotely in the hippocampus and basal forebrain. The presence of apoptosis was suggested by DNA laddering on electrophoresis in the hemolysate-injected subjects (4/6 animals). No laddering was evident in saline-injected subjects (n=4). These results provide evidence that the presence of subarachnoid blood products is associated with DNA fragmentation and apoptotic cell death. PMID- 10708683 TI - Differential neuroendocrine responsiveness to morphine in Lewis, Fischer 344, and ACI inbred rats. AB - Preclinical evidence suggests there is a link between the responsiveness to stress and the propensity to self-administer drugs of abuse. Our previous findings, for example, have shown a significant positive correlation between the locomotor response to novelty and the acquisition of morphine self-administration in Lewis (LEW), Fischer 344 (F344) and ACI inbred rat strains. As an extension of this work, we now report on the neuroendocrine responses (i.e., corticosterone and prolactin secretion) evoked by morphine administration in these same inbred strains. Male LEW, F344, and ACI rats were surgically prepared with indwelling jugular catheters 7 days prior to the study. Following a habituation period, rats were treated with i.p. saline or morphine (1, 5 or 10 mg/kg). Repeated blood samples were withdrawn via the catheters immediately before and at 20, 40, 60 and 120 min after injection. Plasma samples were assayed for hormone levels by radioimmunoassay. No differences in baseline corticosterone levels were found across strains. There was a significant effect of genotype on the corticosterone response to saline injection (i.e., mild stress), with F344 rats exhibiting sustained elevations in corticosterone compared to LEW and ACI rats. Morphine induced stimulation of corticosterone release differed significantly across strains, and in this case LEW rats displayed a reduced sensitivity to morphine. Similar to the corticosterone results, LEW rats also had blunted prolactin responses to morphine when compared to F344 rats. Our data demonstrate that genotype is an important factor modulating the neuroendocrine sensitivity to morphine. It is noteworthy that LEW rats acquire self-administration more rapidly than F344 or ACI rats, yet LEW rats display reduced corticosterone responses to stress and morphine. Taking into account the particular conditions of this study (high i.p. doses used here vs. low i.v. doses in self-administration studies), our results do not suggest that corticosterone response to stress and morphine is related to vulnerability to intravenous opiate self-administration. The data, however, are consistent with the idea of that genetic factors might influence the sensitivity to the morphine-induced effects of glucocorticoids across these inbred strains. PMID- 10708684 TI - Bilateral lesions of the gustatory thalamus disrupt morphine- but not LiCl induced intake suppression in rats: evidence against the conditioned taste aversion hypothesis. AB - Rats decrease intake of a saccharin conditioned stimulus (CS) when followed by: (1) the administration of an aversive agent such as lithium chloride (referred to as a conditioned taste aversion, CTA); (2) access to a very palatable concentration of sucrose (referred to as an anticipatory contrast effect, ACE); or (3) the administration of a drug of abuse. It is not clear, however, whether the suppressive effects of drugs of abuse are mediated by their aversive or rewarding properties. The present set of experiments addressed this issue by examining the suppressive effects of morphine in rats with a lesion thought to dissociate the two phenomena (i.e., CTA and ACE). The results show that bilateral ibotenic acid lesions of the gustatory thalamus eliminate the suppressive effects of morphine, but fail to disrupt the suppressive effects of the aversive agent, lithium chloride. This pattern of results argues against the CTA account in favor of the reward comparison hypothesis. Specifically, the data suggest that rats suppress intake of a saccharin CS in anticipation of the availability of a preferred drug of abuse and that the gustatory thalamus is essential for this type of reward comparison process. PMID- 10708685 TI - Increased cerebral glucose utilization and decreased glucose transporter Glut1 during chronic hyperglycemia in rat brain. AB - Whereas acute hyperglycemia has been shown to result in an unchanged local cerebral glucose utilization (LCGU) the changes of LCGU during chronic hyperglycemia are a matter of dispute. The present study had three aims: (1) To compare the effects of acute and chronic hyperglycemia on LCGU and to investigate in vivo the lactate level as a potential indicator of glycolytic flux. (2) To investigate local changes in brain Glut1 and/or Glut3 glucose transporter densities during chronic hyperglycemia. (3) To analyze the relationship between LCGU and local Glut densities during chronic hyperglycemia. To induce chronic hyperglycemia in rats steptozotocin was given i.p. and experiments were performed 3 weeks later. LCGU was measured by the 2-[14C]deoxyglucose method and intraparenchymal lactate concentration by MR-spectroscopy. Local densities of the glucose transport proteins were determined by immunoautoradiographic methods. During chronic hyperglycemia weighted average of LCGU increased by 13.9% whereas it remained unchanged during acute hyperglycemia. The cerebral lactate/choline ratio was increased by 143% during chronic hyperglycemia. The average density of glucose transporters Glut1 decreased by 7.5%. Local densities of Glut1 were decreased in 12 of 28 brain structures. Glut3 remained unchanged. Positive correlations were found between LCGU and local Glut densities during control conditions and during chronic hyperglycemia. It was concluded that (1) Chronic, but not acute hyperglycemia is followed by an increased LCGU. (2) The capacity to transport glucose is decreased during chronic hyperglycemia. (3) Increased LCGU and decreased densities of Glut1 are matched on a local level. PMID- 10708686 TI - Biotin and biocytin uptake into cultured primary calf brain microvessel endothelial cells of the blood-brain barrier. AB - The uptake of biotin and the closely related biocytin was characterized in primary cultures of calf brain microvessel endothelial (CBME) cells. Biotin uptake was found to be Na(+)-gradient dependent and independent of changes in the membrane potential. Concentration dependence revealed a single saturation mechanism with a K(m) of 47 microM and a V(max) of 101 pmol/min/mg. Inhibition studies demonstrated dependence on metabolic energy and the necessity for a free carboxyl group for transport activity. The anticonvulsants primidone and carbamazepine had no inhibitory effect. Biotin uptake into CBME cells is a secondary active, electroneutral, saturable and specific process. Biocytin which accumulates in biotinidase deficiency, a human congenital disorder, did not inhibit biotin uptake and was not transported into these cells. The presence of human serum with normal biotinidase activity significantly reduced biotin uptake by about 50%. Further, added biocytin was hydrolyzed to biotin, which accumulated intracellularly but to a lesser extent than added free biotin. Biotin uptake after addition of plasma of biotinidase-deficient patients was not different from that in the presence of normal serum. These results indicate that the absence of biotinidase activity in serum does not reduce blood-brain barrier transport of biotin. PMID- 10708687 TI - 3-nitropropionic acid induced in vivo protein oxidation in striatal and cortical synaptosomes: insights into Huntington's disease. AB - 3-nitropropionic acid (3-NP) administered systemically daily for 4 days to rats inhibits mitochondrial oxidative phosphorylation and induces selective lesions in the striatum in a manner reminiscent of Huntington's disease (HD). To investigate the potential oxidative nature of these lesions, rats were injected with 3-NP (20 mg/kg, i.p. daily for 4 days) and subsequently isolated brain synaptosomal membranes were examined for evidence of oxidative stress. Brain synaptosomal membrane proteins from rats injected with 3-NP exhibited a decreased in W/S ratio, the relevant electron paramagnetic resonance (EPR) parameter used to determine levels of protein oxidation (76% of control), and Western blot analysis for protein carbonyls revealed direct evidence of increased synaptosomal membrane protein oxidation (248% of control). Similar results were obtained in synaptosomes isolated from striatum and from cerebral cortex, demonstrating that the oxidative changes are not restricted to the lesion site. Moreover, increased oxidative stress was evident prior to the appearance of morphological lesions. These data are consistent with the hypothesis that 3-NP-induced striatal lesions, and perhaps those in HD, are associated with oxidative processes. PMID- 10708688 TI - Permanent cerebral hypoperfusion: 'preconditioning-like' effects on rat energy metabolism towards acute systemic hypotension. AB - Chronic cerebrovascular disorders are often complicated by additional temporary ischaemic insults, resulting in substantial deterioration of brain energy metabolism. In the present study, chronic limitations of oxygen supply were induced in Wistar rats by 2 weeks of permanent bilateral common carotid artery occlusion (2-vo) to initiate a 'preconditioning-like' effect that protects rat brain energy metabolism against further acute systemic hypotension (15 min). Haemodynamic parameters, arterial blood gases and body temperature were monitored. Energy metabolites were determined in rat parietotemporal cerebral cortex: adenosine 5'-triphosphate (ATP), adenosine 5'-diphosphate (ADP), adenosine 5'-monophosphate (AMP), phosphocreatine (PCr), and adenosine by the high-pressure liquid chromatography (HPLC) technique and lactate spectrophotometrically. After 2 weeks, permanent 2-vo led to a significant decrease in the concentrations of cortical tissue ATP and PCr, from 3.06+/-0.48 to 2. 09+/-0.28 and from 4.27+/-0.63 to 3.35+/-0.41 micromol/g, respectively. These changes were associated with a two-fold increase in AMP and adenosine. Acute systemic hypotension alone (non-preconditioning) reduced ATP and PCr drastically, to 0.97+/-0. 51 and 1.76+/-1.23 micromol/g. Tissue concentrations of lactate, AMP, and adenosine were markedly increased, three- to five-fold, in 'non preconditioned' brain tissue. In contrast, after 2 weeks of 2-vo acute hypotension did not significantly alter the cortical energy state any further. The effects of preconditioning on tissue ATP and PCr were most pronounced at 5 min and 48 h after reperfusion. In conclusion, permanent 2-vo seems to activate compensatory mechanisms, which effectively protect the rat's cortical energy metabolism against an additional ischaemic attack ('preconditioning-like' effect). PMID- 10708689 TI - Prolonged intracerebroventricular infusion of neurosteroids affects cognitive performances in the mouse. AB - The effects of prolonged intracerebroventricular (i.c.v.) steroid infusions on memory performances (Y-maze arm discrimination test) and on neurosteroids brain levels were studied in young adult male mice. The Y-maze test consisted of two trials separated by a time interval. In the first trial, one arm of the maze (subsequently called novel arm) was closed, and mice were allowed to visit the two accessible arms. After a short 2-h intertrial interval (ITI), control mice explored preferentially the novel arm, whereas with a longer 6-h ITI, they did not remember the location of the novel arm and performed at random level (33% of time spent in each arm). Using a 2-h ITI, allopregnanolone (THPROG, 0.5 and 1 ng/h) decreased memory performances to random level after 3 and 6 days of infusion. Conversely, with a 6-h ITI, pregnenolone sulfate (PREG S, 10, 50, and 100 ng/h) significantly increased memory performances after 3 days, but only the smallest dose was still effective after 6 days. THPROG infusion (1 ng/h) increased the forebrain concentration of 5alpha-dihydroprogesterone (DHPROG) and tended to increase its own level. PREG S administration (10 ng/h) increased its own concentration and tended to increase those of pregnenolone (PREG) and of further metabolites. In conclusion, the memory-enhancing effects of PREG S and the inhibitory ones of THPROG have been confirmed. A persistent, however moderate, increase of PREG S brain concentration might be of interest for the treatment of amnesic deficits. PMID- 10708690 TI - GM1 ganglioside restores abnormal responses to acute thermal and mechanical stimuli in aged rats. AB - We investigated the effect of aging on the responses to thermal and mechanical stimuli in rats. Young (3-5 months old) and aged (22-24 months old) male Sprague Dawley rats were tested in the hot plate, high- and low-intensity radiant heat tail flick, and von Frey hair assays. Compared to young rats, aged rats displayed longer latencies in the hot plate and the high-intensity tail flick assays (hypoalgesia), but there was no difference in the low-intensity tail flick assay. In addition, aged rats had decreased thresholds to mechanical stimuli produced by von Frey hairs compared with young rats (mechanical allodynia). Administration of GM1 ganglioside, 30 mg/kg, i.p., once daily for 30 days, to aged rats partially restored the responses in the hot plate and von Frey hair assays. GM1 had no effect on the altered responses in the tail flick test in aged rats, and in general, had no effect on any sensory modality tested in young rats. PMID- 10708691 TI - Suppression of proteasome C2 contralateral to ischemic lesions in rat brain. AB - Functional as well as structural reorganization takes place in the surrounding and remote brain areas after focal ischemic lesions. In particular, reactive or regenerative processes have been described to occur in the contralateral hemisphere. We used mRNA differential display to gain more insight into the molecular mechanisms underlying this type of neuronal plasticity. Circumscribed unilateral infarcts consistently affecting the forelimb area of the primary motor cortex were induced photochemically in adult male Wistar rats. The lesion produced significant behavioral asymmetry with subsequent partial recovery within 1 week. Cloning the genes with altered expression profiles identified the 20S proteasome subunit C2 as a gene whose expression level is decreased in contralateral homotopic cortex. Semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) revealed approximately twofold lower proteasome C2 mRNA levels in the lesion group as compared with the sham-operated group. The proteasome serves as the central enzyme of non-lysosomal protein degradation. It is responsible for intracellular protein turnover and is critically involved in a variety of regulation processes, such as cell cycle, metabolism and differentiation. Our results suggest that proteasome activity may play also a role in contralateral cortical plasticity occurring after focal cerebral ischemia. PMID- 10708692 TI - Effect of a 5-HT(1A) receptor agonist, flesinoxan, on the extracellular noradrenaline level in the hippocampus and on the locomotor activity of rats. AB - We have studied effects of 5-hydroxytryptamine 1A (5-HT(1A)) receptor-selective compounds on the extracellular noradrenaline (NA) level in the hippocampus of rats using microdialysis and on their locomotor activity. A selective 5-HT(1A) receptor agonist, flesinoxan (5 mg/kg, i.p.) increased the extracellular NA level in the hippocampus, and increased the locomotor activity. Both responses were blocked by pretreatment with a 5-HT(1A) receptor antagonist, WAY100635 (1 mg/kg, i.p.) and an alpha(2) adrenoceptor agonist, clonidine (50 microg/kg, i.p.). Bilateral intrahippocampal injection of flesinoxan (200 nmol in 2 microl, respectively) increased the locomotor activity of rats and the intrahippocampal perfusion of flesinoxan (1 mM, 2 microl/min) increased the extracellular NA level in the hippocampus. Bilateral intrahippocampal injections of a small amount of WAY100635 (0.1 nmol in 2 microl, respectively) blocked the flesinoxan (5 mg/kg, i.p.)-induced hyperactivity. Flesinoxan (5 mg/kg, i.p.) did not significantly influence the level of serotonin or its major metabolite in the hippocampus, or dopamine or its metabolites in the striatum. In conclusion, these behavioral as well as pharmacological results indicate that postsynaptic 5-HT(1A) receptor activation by flesinoxan increase the extracellular NA level in the hippocampus, which may be the cause of the increase of the locomotor activity. PMID- 10708693 TI - Blockade of alcohol-induced locomotor sensitization and conditioned place preference in DBA mice by 7-nitroindazole. AB - Our previous studies indicated that inhibition or ablation of the neuronal nitric oxide synthase (nNOS) prevents the development of sensitization to the locomotor stimulating effect of cocaine and cocaine-induced conditioned place preference (CPP). The present study was undertaken to investigate the effect of the nNOS inhibitor, 7-nitroindazole (7-NI), on ethanol-induced locomotor sensitization and CPP in DBA/2J mice. Administration of ethanol (1.5 g/kg; i.p.) for 7 days resulted in a progressive increase in the locomotor-stimulating effect of ethanol. Pretreatment with 7-NI (25 mg/kg) blocked the expression of the sensitized response to ethanol. A challenge injection of ethanol given 1 week and then 4 weeks following withdrawal from ethanol indicated that (a) ethanol sensitization was long lasting, and (b) the co-administration of 7-NI and ethanol attenuated the sensitized response to ethanol challenge. The CPP experiments showed that pairing four ethanol (2.5 g/kg) injections with a specific environment resulted in a marked preference for the drug-paired environment. The pretreatment with 7-NI (25 mg/kg) completely blocked ethanol-induced CPP. 7-NI alone produced neither rewarding nor aversive effects. Taken together, results of the present study indicate that blockade of nNOS by 7-NI-attenuated ethanol induced behavioral sensitization and completely blocked the rewarding effect of ethanol. These findings support the role of NO in ethanol actions and further suggest that the nNOS system is relevant to the rewarding effects of various drugs of abuse. PMID- 10708694 TI - GABA-mediated corticofugal inhibition of taste-responsive neurons in the nucleus of the solitary tract. AB - The nucleus of the solitary tract (NST) receives descending connections from several forebrain targets of the gustatory system, including the insular cortex. Many taste-responsive cells in the NST are inhibited by gamma-aminobutyric acid (GABA). In the present study, we investigated the effects of cortical stimulation on the activity of gustatory neurons in the NST. Multibarrel glass micropipettes were used to record the activity of NST neurons extracellularly and to apply the GABA(A) antagonist bicuculline methiodide (BICM) into the vicinity of the cell. Taste stimuli were 0.032 M sucrose (S), 0.032 M NaCl (N), 0.00032 M citric acid (H), and 0.032 M quinine hydrochloride (Q), presented to the anterior tongue. Each of 50 NST cells was classified as S-, N-, H-, or Q-best on the basis of its response to chemical stimulation of the tongue. The ipsilateral insular cortex was stimulated both electrically (0.5 mA, 100 Hz, 0.2 ms) and chemically (10 mM DL-homocysteic acid, DLH), while the spontaneous activity of each NST cell was recorded. The baseline activity of 34% of the cells (n=17) was modulated by cortical stimulation: eight cells were inhibited and nine were excited. BICM microinjected into the NST blocked the cortical-induced inhibition but had no effect on the excitatory response. Although the excitatory effects were distributed across S-, N-, and H-best neurons, the inhibitory effects of cortical stimulation were significantly more common in N-best cells. These data suggest that corticofugal input to the NST may differentially inhibit gustatory afferent activity through GABAergic mechanisms. PMID- 10708695 TI - Pharmacological characterization of a novel AVP(4-9) binding site in rat hippocampus. AB - pGlu-Asn-Cys (Cys)-Pro-Arg-Gly-NH(2) (AVP(4-9)), a major metabolite C-terminal fragment of Arginine(8)-vasopressin (AVP), improves the disruption of the learning and memory, and is a far more potent in the mnemonic function than AVP. In this study, we pharmacologically characterized its putative binding site and mechanism of intracellular signaling. Radioligand binding assay showed that [35S]AVP(4-9) could detect specific binding sites in the rat hippocampus membrane preparations, and the binding site was specifically displaced by AVP(4-9) but not by either V(1) or V(2) antagonists. Furthermore, [35S]AVP(4-9) could not detect the cloned rat V(1a), V(1b) and V(2) vasopressin receptors. Even at a low doses (10-100 pM), AVP(4-9) caused an increase in both inositol(1,4, 5)-trisphosphate (Ins(1,4,5)P(3)) and intracellular calcium concentrations ([Ca(2+)](i)) in rat hippocampal cells. The AVP(4-9)-induced [Ca(2+)](i) increase was partially inhibited by the absence of Ca(2+) or by Ca(2+)-channel blocker, suggesting that AVP(4-9) caused the [Ca(2+)](i) increase via release from intracellular calcium store as well as influx from extracellular calcium. For the first time, this study provides evidence to show that AVP(4-9) activates Ins(1,4,5)P(3)/[Ca(2+)](i) pathway through a novel type of receptor in rat hippocampus, which might be potentially important in improving the mnemonic function. PMID- 10708696 TI - A 5-HT(1B) receptor agonist inhibits light-induced suppression of pineal melatonin production. AB - Serotonin (5-HT) modulates the phase adjusting effects of light on the mammalian circadian clock through the activation of presynaptic 5-HT(1B) receptors located on retinal terminals in the suprachiasmatic nucleus (SCN). The current study was conducted to determine whether activation of 5-HT(1B) receptors also alters photic regulation of nocturnal pineal melatonin production. Systemic administration of the 5-HT(1B) receptor agonist TFMPP attenuated the inhibitory effect of light on pineal melatonin synthesis in a dose-related manner with an apparent ED(50) value of 0.9 mg/kg. The effect of TFMPP on light-induced melatonin suppression was blocked by the 5-HT(1) receptor antagonist, methiothepin, but not by the 5-HT(1A) antagonist, WAY 100,635, consistent with the involvement of 5-HT(1B) receptors. The results are consistent with the interpretation that activation of presynaptic 5-HT(1B) receptors on retinal terminals in the SCN attenuates the effect of light on pineal melatonin production, as well as on circadian phase. PMID- 10708697 TI - Monosynaptic and disynaptic projections from the substantia nigra pars reticulata to the parafascicular thalamic nucleus in the rat. AB - We examined a direct pathway and an indirect pathway via the reticular thalamic nucleus (RT) from the substantia nigra pars reticulata (SNr) to the parafascicular thalamic nucleus (PF) by using anterograde and retrograde tract tracing methods. After biotinylated dextranamine (BDA) injection into the dorsolateral part of the SNr, many labeled fibers and axon terminals were distributed in the ventral part of the RT, as well as in the ventrolateral part of the PF, bilaterally with an ipsilateral dominance. After BDA injection into the ventral part of the RT, a plexus of labeled axons was found bilaterally with an ipsilateral dominance in the ventrolateral part of the PF. After combined injections of BDA into the dorsolateral part of the SNr and cholera toxin B subunit (CTb) into the ventrolateral part of the PF on the same side, overlapping distribution of BDA-labeled fibers and CTb-labeled neurons was observed in the ventral part of the RT ipsilateral to the injection sites, where the BDA-labeled axon terminals made symmetrical synaptic contacts with soma and dendrites of the CTb-labeled neurons. PMID- 10708698 TI - Effect of quinolinic acid on endogenous antioxidants in rat corpus striatum. AB - The response of endogenous antioxidants to the N-methyl-D-aspartate (NMDA) receptor agonist and excitotoxin, quinolinic acid (QUIN), was investigated in rat corpus striatum. Animals treated with QUIN (240 nmol/microl), were sacrificed at 120 min after a single intrastriatal injection to examine the alterations in the levels of both reduced (GSH) and oxidized (GSSG) glutathione, and the activities of the antioxidant enzymes, superoxide dismutase (SOD) and glutathione peroxidase (Gpx). Changes in the rate of lipid peroxidation (LP) were also measured after exposure to different doses of QUIN (60, 120, 240 and 480 nmol/microl) as an index of oxidative stress. When compared to control, lipid peroxidation was increased at QUIN doses of 240 and 480 nmol/microl. Striatal levels of GSH and GSSG were decreased and increased, respectively, after QUIN injection; whereas GPx activity was unchanged. Cytosolic copper/zinc SOD (CuZn-SOD) activity decreased after treatment, while mitochondrial manganese SOD (Mn-SOD) was unchanged. The alterations observed on these antioxidant systems suggest that QUIN toxicity is mediated by specific mechanisms leading to oxidative stress. PMID- 10708699 TI - Conditioned fear to environmental context: cardiovascular and behavioral components in the rat. AB - This study compares the time course of the cardiovascular changes (mean arterial blood pressure, heart rate) and behavioral changes (freezing, rearing, grooming and activity) evoked by 30 min long exposures to a footshock chamber before and after conditioning with footshocks. The main finding is that the conditioned fear evoked by re-exposure to the footshock chamber after conditioning is associated with a prolonged freezing response, a marked rise in mean arterial pressure (+35 mm Hg above a resting baseline of 105 mm Hg) and a delayed rise in heart rate. The pattern of behavioral and cardiovascular changes is the same as with conditioned fear to a discrete stimulus but the effect is a lot longer. PMID- 10708700 TI - Acute delta-opioid receptor activation induces CREB phosphorylation in NG108-15 cells. AB - A growing body of evidence supports an important role of the transcription factor cAMP responsive element binding protein (CREB) in mediating opioid-induced changes in the cAMP pathway. Regulation of CREB and subsequent changes in gene expression may underlie some long-term cellular adaptations associated with the administration of opioid drugs. The effect of morphine on the level of the transcription factor CREB, as well as CREB phosphorylation, was investigated in NG108-15 cells. Morphine and the delta-opioid receptor agonist [D Pen(2,5)]enkephalin (DPDPE) produced a dose-dependent increase in CREB phosphorylation. The effect was reversed by naloxone and naltrindole, respectively. The calmodulin antagonist N-(6-aminohexyl)-5-chloro-1 naphthalenesulfonamide hydrochloride (W-7), the protein kinase inhibitor staurosporine, as well as 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride (H-7), an inhibitor of protein kinase C and cAMP-dependent protein kinase, but not N-[2-(methylamino)ethyl]-5-isoquinolinesulfonamide dihydrochloride (H-8), an inhibitor of cAMP- and cGMP-dependent protein kinase, blocked the opioid-induced CREB phosphorylation. The obtained results suggest that in the cells studied opioids affect, via the delta-opioid receptor, stimulatory intracellular mediator systems involving Ca(2+)/calmodulin and the protein kinase C pathway. PMID- 10708701 TI - Effects of CI-1002 and CI-1017 on spontaneous synaptic activity and on the nicotinic acetylcholine receptor of Torpedo electric organ. AB - The effect of azepino[2,1-b]quinazoline 1,3-dichloro-6,7,8,9,10, 12-hexahydro-, mono-hydrochloride (CI-1002), a tacrine derivative, and 1-azabicyclo[2.2.1]heptan 3-one, O-[3-(methoxyphenyl)-2-propynyl]oxime [R-(Z)]-2-butenedioate (CI-1017), a muscarinic M(1) receptor agonist, on spontaneous synaptic activity was investigated by measuring amplitude, rise time, velocity of rising, half-width, and electrical charge of miniature endplate potentials (m.e.p.p.) recorded extracellularly in Torpedo electric organ fragments. The effect of CI-1002 and CI 1017 on the nicotinic acetylcholine receptor was investigated by measuring the current induced by acetylcholine in Xenopus laevis oocytes transplanted with membranes from Torpedo electric organ. CI-1002, at a concentration of 1 microM, altered the m.e.p.p. by increasing the amplitude (from 1.08+/-0.01 to 2.76+/-0.03 mV), rise time (from 0. 700+/-0.006 to 1.02+/-0.01 ms), rising rate (from 1.79+/ 0.02 to 3. 45+/-0.05 mV/ms), half-width (from 0.990+/-0.008 to 2.40+/-0.02 ms), and electrical charge (from 304+/-4 to 784+/-11 mV s). CI-1017, at a concentration of 1 microM, altered the m.e.p.p. by decreasing the amplitude (from 1.08+/-0.01 to 0.650+/-0.007 mV), rise time (from 0. 700+/-0.006 to 0.530+/-0.007 ms), rising rate (from 1.79+/-0.02 to 1. 53+/-0.02 mV/ms), half-width (from 0.990+/-0.008 to 0.670+/-0.007 ms), and electrical charge (from 304+/-4 to 75+/-1 mV s). CI-1002 inhibited the acetylcholine-induced current of nicotinic acetylcholine receptors with an IC(50) of 3.4+/-0.3 microM. CI-1017 inhibited the acetylcholine-induced current of nicotinic acetylcholine receptors with an IC(50) of 0.8+/-0.1 microM. These results indicate that, although both drugs interacted negatively with nicotinic acetylcholine receptors, CI-1002 overcame this inhibition by recruiting more acetylcholine to build a quantum. PMID- 10708702 TI - Characterisation of human melatonin mt(1) and MT(2) receptors by CRE-luciferase reporter assay. AB - A cyclic AMP response element (CRE)-luciferase reporter gene assay was used to characterise the functional responses of human melatonin mt(1) and human melatonin MT(2) receptors, stably expressed in the human embryonic kidney cell line HEK293, to a series of six naphthalenic analogues of melatonin. By comparison to the observed melatonin-mediated inhibition of stimulated luciferase levels the naphthalenic series was identified as comprising agonists, partial agonists and one antagonist of melatonin mt(1) and melatonin MT(2) receptor function. Three of the agonist/partial agonist members of this series were also identified as displaying a functional selectivity for the melatonin MT(2) receptor. Competitive displacement of 2-[125I]iodomelatonin binding to the ovine pars tuberalis melatonin ML(1) receptor demonstrated a close correlation to the observed functional luciferase responses of the human melatonin mt(1) receptor. We conclude that the CRE-luciferase reporter gene assay provides an effective functional screening method for the pharmacological characterisation of human melatonin receptor subtypes. PMID- 10708703 TI - Characterization of receptor-mediated [35S]GTPgammaS binding to cortical membranes from postmortem human brain. AB - The [35S]GTPgammaS binding assay represents a functional approach to assess the coupling between receptors and G-proteins. The optimal conditions for [35S]GTPgammaS binding to human brain homogenates were established in postmortem samples of prefrontal cortex. The influence of protein content, incubation time, GDP, Mg(2+), and NaCl concentrations on the [35S]GTPgammaS binding were assessed in the absence and presence of the alpha(2)-adrenoceptor agonist UK14304 5-bromo N-(4,5-dihydro-1H-imidazol-2-yl)-6-quinoxalinamine). In conditions of 50 microM GDP and 100 mM NaCl, UK14304 increased the apparent affinity of the specific [35S]GTPgammaS binding without changing the apparent density. Concentration response curves to agonists of alpha(2)-adrenoceptors, mu-opioid, 5-HT(1A), cholinergic muscarinic, and GABA(B) receptors displayed, in the presence of NaCl, maximal stimulations between 24% and 61% with EC(50) values in the micromolar range. Selective antagonists shifted to the right the agonist-induced stimulation curves. The G(i)/G(o)-protein alkylating agent N-ethylmaleimide decreased basal [35S]GTPgammaS binding in a concentration-dependent manner and inhibited the stimulation induced by the different agonists. In cortical sections, [35S]GTPgammaS binding to gray matter was stimulated by the agonist UK14304. The present study demonstrates that functional studies of the receptor coupling to G(i)/G(o)-proteins can be performed in postmortem human brain samples. PMID- 10708704 TI - Inhibitory effect of cepharanthine on fibronectin production in growth factor stimulated rat mesangial cells. AB - We examined the effect of cepharanthine, a biscoclaurine alkaloid, on extracellular matrix production in rat mesangial cells in response to platelet derived growth factor (PDGF) or transforming growth factor-beta (TGF-beta). Stimulation of the cells with PDGF increased the amounts of fibronectin, one of extracellular matrix components. Pretreatment with cepharanthine (0.1-2 microM) suppressed the PDGF-stimulated increase in fibronectin in a dose-dependent manner. At a concentration of 2 microM, the alkaloid almost completely suppressed the production. Under the conditions, the alkaloid inhibited tyrosine phosphorylation of several proteins including PDGF beta receptor in PDGF stimulated cells, and also tyrosine kinase activity of the receptor prestimulated with PDGF in a cell-free assay system. Furthermore, cepharanthine suppressed TGF beta-stimulated fibronectin production at the same concentration ranges. Our results suggest that cepharanthine inhibits fibronectin production induced by growth factors, probably through suppression of receptor autophosphorylation. PMID- 10708705 TI - The role of adenosine A(1) receptors in the ATP-evoked Ca(2+) response in rat thyroid FRTL-5 cells. AB - The effect of adenosine A(1) receptor activation on the ATP-induced increase in intracellular free Ca(2+) was studied in control and protein kinase C down regulated Fisher rat thyroid (FRTL-5) cells. Long-term phorbol ester treatment, which leads to protein kinase C down-regulation, enhanced the ATP-evoked extracellular Ca(2+) influx. The increased Ca(2+) influx was antagonized by the adenosine A(1) receptor antagonist 8-cyclopentyl-1, 3-dipropylxanthine (DPCPX). [3H]DPCPX binding studies revealed that phorbol ester-treatment increased the number of adenosine A(1) receptors. The adenosine A(1) receptor-mediated inhibition of the cyclic AMP formation was not affected by the increased receptor number. We conclude that the enhanced ATP-evoked Ca(2+) influx in protein kinase C down-regulated cells is mediated by adenosine formed by hydrolysis of ATP, and that this adenosine interacts with the increased number of A(1) receptors. The mechanism by which adenosine enhances Ca(2+) entry is not known. Thus, the larger number of adenosine A(1) receptors broadens the spectrum of adenosine A(1) receptor affected signaling systems in FRTL-5 cells. PMID- 10708706 TI - Functional characterization of corticotropin-releasing factor type 1 receptor endogenously expressed in human embryonic kidney 293 cells. AB - The endogenous expression in human embryonic kidney 293 (HEK293) cells of corticotropin-releasing factor (CRF) receptors was detected. High-affinity binding sites for human CRF (K(i)=3.6 nM), ovine CRF (K(i)=4.6 nM), rat urocortin (K(i)=2.2 nM), sauvagine (K(i)=2.4 nM) and astressin (K(i)=4.3 nM) with the pharmacological characteristics for CRF type 1 (CRF(1)) receptors and B(max) values of approximately 30 fmol/mg protein were determined. The four CRF receptor agonists nonselectively stimulated cAMP production in HEK293 cells at low agonist concentrations, whereas the antagonist astressin shifted the dose-response curve for ovine CRF significantly rightward. Transfection of the pcDNA3 vector into HEK293 cells strongly reduced the expression of the endogenous CRF receptor. Northern blot analysis revealed the expression of a CRF(1) transcript in human neuronal tissues, HEK293, human NTera-2 (NT2) carcinoma, Y-79 retinoblastoma and African green monkey kidney (COS-7) cells. Neither by Northern blot analysis nor by reverse transcriptase PCR (RT-PCR), the expression of CRF(2) could be detected. In cAMP stimulation experiments, functional CRF receptors were detected in these cell lines. These data show that HEK293 and other cell lines endogenously express CRF(1) receptors. PMID- 10708707 TI - Activation of p38 mitogen-activated protein kinase by formyl-methionyl-leucyl phenylalanine in rat neutrophils. AB - The signaling pathways leading to p38 mitogen-activated protein kinase (MAPK) activation in formyl-methionyl-leucyl-phenylalanine (fMLP)-stimulated rat neutrophils were examined. Immunoblot analysis with antibodies against a phosphorylated form of p38 MAPK showed that fMLP-stimulated p38 MAPK activation was dependent on a pertussis toxin-sensitive G protein. Two phosphatidylinositol 3-kinase inhibitors, wortmannin and 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4 one (LY294002), did not affect the p38 MAPK activation. Phosphorylation of p38 MAPK was concentration dependently attenuated by a tyrosine kinase inhibitor, genistein, and by a Ca(2+)-dependent protein kinase C inhibitor, 13-cyanoethyl-12 methyl-6,7,12,13-tetrahydroindolo[2,3-a]pyrrolo[3 , 4-c]carbazole-7-one (Go6976). However, the protein kinase C inhibitors with a broader spectrum, 2-[1-(3 dimethylaminopropyl)-5-methoxy-1H-indol-3-yl]-3-(1H-indol-3-y l)-maleimide (Go6983) and 2-[1-(3-dimethylaminopropyl)-1H-indol-3-yl]-3-(1H-indol-3-yl) maleimi de (GF109203X), had no inhibitory effect. fMLP-stimulated p38 MAPK phosphorylation was also reduced in cells pretreated with a phospholipase C inhibitor, 1-[6-((17beta-3-methoxyestra-1,3, 5(10)-trien-17-yl)amino)hexyl]-1H pyrrole-2,5-dione (U73122), or preloaded with an intracellular Ca(2+) chelator, 1, 2-bis-(o-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid (BAPTA). We conclude that phosphorylation of p38 MAPK by fMLP stimulation in rat neutrophils is dependent on G(i/o) protein, nonreceptor tyrosine kinase, phospholipase C/Ca(2+), and probably Ca(2+)-dependent protein kinase C pathways. PMID- 10708708 TI - Inactivation of 5-HT(1A) receptors in hippocampal and cortical homogenates. AB - 5-HT(1A) receptor function can be assessed in rat hippocampal and cortical membrane preparations as agonist-stimulated [35S]GTPgammaS binding. Membranes were preincubated in vitro with N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ). R(+)-8-hydroxy-2-(di-n-propylamino)tetralin [R(+)-8-OH-DPAT]-stimulated [35S]GTPgammaS binding and [3H]8-OH-DPAT binding assays were used to assess 5 HT(1A) receptor function and density, respectively. EEDQ decreased both R(+)-8-OH DPAT-stimulated [35S]GTPgammaS and [3H]8-OH-DPAT binding in hippocampal and cortical membranes. The E(max) but not the EC(50) of R(+)-8-OH-DPAT to stimulate [35S]GTPgammaS binding was decreased by EEDQ in both preparations. Additionally, the IC(50) for EEDQ to reduce R(+)-8-OH-DPAT-stimulated [35S]GTPgammaS and [3H]8 OH-DPAT binding was the same for both brain regions in both assays. In contrast to EEDQ alone, agonist-stimulated [35S]GTPgammaS binding was not reduced in hippocampal membranes preincubated with EEDQ and the 5-HT(1A) receptor antagonist N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-2-pyridinyl- cyclohexanecarboxamide maleate (WAY 100,635), suggesting that EEDQ acts directly on the receptor. Due to parallel reductions in receptor density and maximal functional response, it is concluded that there is little or no reserve for 5 HT(1A) receptor coupling to G(alpha) in these preparations. In addition, the sensitivity of hippocampal and cortical 5-HT(1A) receptors to inactivation by EEDQ in vitro is the same. PMID- 10708709 TI - Effects of abciximab and tirofiban on vitronectin receptors in human endothelial and smooth muscle cells. AB - human umbilical venous endothelial cells. 7E3 binding correlated with alphavbeta3 expression in all cell types. Integrin-mediated cell functions were analysed with adhesion and spreading assays on vitronectin. In human umbilical venous endothelial cells, these functions were mediated by alphavbeta3 and in human iliac arterial smooth muscle cells by alphavbeta5. In human umbilical venous smooth muscle cells, both vitronectin receptors were involved. Abciximab potently inhibited alphavbeta3-mediated cell adhesion and spreading. With tirofiban, no significant inhibition of vascular cell functions was observed. The present data demonstrate that vitronectin-cell interactions in vascular cells are mediated via two distinct integrin-receptors, alphavbeta3 and alphavbeta5. Abciximab, which solely inhibits alphavbeta3-mediated cell functions, may be particularly effective in human endothelium and in beta3-integrin expressing vascular smooth muscle cells. PMID- 10708710 TI - The p55 tumor necrosis factor receptor (CD120a) induces endothelin-1 synthesis in endothelial and epithelial cells. AB - Synthesis of the vasoconstrictor peptide endothelin-1 by endothelial and epithelial cells is strongly induced by tumor necrosis factor alpha (TNF-alpha). The actions of TNF-alpha are mediated by two transmembrane receptors of approximately 55 (p55, CD120a) and 75 kDa (p75, CD120b). Reagents activating selectively these receptor subtypes have been used to identify which TNF receptor mediates the induction of endothelin-1 synthesis. Stimulation of bovine aortic endothelial cells or human HEp-2 epithelial cells with a p55-selective mutant of human TNF-alpha (R32W-S86T) induced significant and concentration-dependent increases in endothelin-1 release. A p75 receptor-selective TNF-alpha mutant (D143N-A145R) was ineffective alone or in combination with the p55-selective mutant. Competitive binding experiments with [125I]TNF-alpha showed the p55 selective mutant, but not the p75-selective mutant, to inhibit the binding of [125I]TNF-alpha to endothelial and HEp-2 cells. Similar results were obtained with the p55 agonist antibody htr1 in both cell lines. These results establish the p55 TNF receptor as the main receptor involved in the induction of endothelin 1 synthesis by TNF-alpha. PMID- 10708711 TI - Opposite effects of halothane on guinea-pig ventricular action potential duration. AB - Halothane protects the heart against the reperfusion injury observed after an ischemia. In ischemic or anoxic conditions, a large ATP-sensitive K(+) (K(ATP)) conductance is supposed to provide an endogenous protection to the myocardium. In this study, we tested the possibility that halothane acted by modulating this conductance. Isolated guinea-pig cardiomyocytes were successively studied in current clamp and in voltage-clamp conditions. Action potentials regulation by halothane was tested in control conditions and in situations where the K(ATP) channels were activated. In control conditions, halothane decreased action potential duration of myocytes but did not significantly alter the inward rectifying K(+) current. Conversely, halothane lengthened action potential of cells in which the K(ATP) conductance was activated, by inhibiting the K(ATP) current. In ischemic conditions, simultaneous shortening of long action potentials and lengthening of shortened ones would be expected to homogenize the absolute refractory period at the border between normoxic and anoxic zones. This effect, together with a decrease in calcium load, could protect the myocardium against re-entrant arrhythmias. PMID- 10708712 TI - T(84) epithelial cells respond to 5-hydroxytryptamine when grown in serum-free media. AB - The aim of this study was to establish the cause of insensitivity of T(84) human colonic epithelial cells to 5-hydroxytryptamine (5-HT). Monolayers of T(84) cells were placed in modified Ussing chambers for measurement of short-circuit current, an index of secretion. When grown in serum-supplemented media, T(84) cells gave secretory responses to acetylcholine and forskolin but not to 5-HT. When grown in AIM V serum-free media, T(84) cells responded to 5-HT. Chromatographic analysis with fluorimetric detection showed high levels of 5-HT (1.8 microM) in the serum. This contamination is probably responsible for subsequent desensitization of T(84) cells to 5-HT. PMID- 10708713 TI - Highly potent neurotensin analog that causes hypothermia and antinociception. AB - The tridecapeptide neurotensin has long been proposed as an endogenous neuroleptic. However, for neurotensin [or neurotensin(8-13) [NT(8-13)], the active fragment] to cause its effects, it must be administered centrally. Here, we report on an analog of NT(8-13), (N-methyl-Arg),Lys,Pro,L-neo-Trp,tert-Leu,Leu (NT69L), which contains a novel amino acid, L-neo5 degrees C (rectal), with a significant effect persisting for over 7 h. NT69L also caused a rapid (within 15 min) and persistent (for over 5 h) antinociceptive effect, as determined by the hot plate test. NT69L was overall the most potent and longest lasting neurotensin analog that has been reported. These studies provide the background for further testing of a stable, potent and long lasting neurotensin analog as a potential neuroleptic. PMID- 10708714 TI - Effect of diabetes on bradykinin-induced thermal hyperalgesia in mice. AB - To investigate the role of protein kinase C in the attenuation of bradykinin induced thermal hyperalgesia in diabetic mice, we examined the effects of a protein kinase C activator or inhibitor on the i.t. bradykinin-induced hyperalgesia in diabetic and non-diabetic mice. Intrathecal injection of bradykinin caused a transient antinociceptive effect, which diminished within 30 min, and then produced a thermal hyperalgesia, which lasted about 120 min, in non diabetic mice. Although the duration of the antinociceptive phase was longer in diabetic mice than in non-diabetic mice, the hyperalgesic response was not observed in diabetic mice. The bradykinin-induced hyperalgesia was dose dependently and significantly enhanced by pretreatment with calphostin C (0.3 to 3 pmol, i.t.), a specific protein kinase C inhibitor, in diabetic mice. However, calphostin C (3 pmol, i.t.) had no significant effect on bradykinin-induced hyperalgesia in non-diabetic mice. On the other hand, pretreatment with phorbol 12, 13-dibutyrate (12.5 to 50 pmol, i.t.), a protein kinase C activator, significantly and dose-dependently reduced bradykinin-induced hyperalgesia in non diabetic mice. However, phorbol-12, 13-dibutyrate (50 pmol, i.t. ) had no significant effect on bradykinin-induced hyperalgesia in diabetic mice. These results suggest that the change in bradykinin-induced thermal hyperalgesia in diabetic mice may be due, at least in part, to the modification of nociceptive transmission in the spinal cord by the activation of protein kinase C. PMID- 10708715 TI - Responses of the extrapyramidal and limbic substance P systems to ibogaine and cocaine treatments. AB - Ibogaine is an indolamine found in the West Africa shrub, Tabernanthe iboga, and has been proposed for the treatment of addiction to central nervous system (CNS) stimulants such as cocaine and amphetamine. The mechanism of ibogaine action and its suitability as a treatment for drug addiction still remains unclear. Since previous studies demonstrated differential effects of stimulants of abuse (amphetamines) on neuropeptide systems such as substance P, we examined the impact of ibogaine and cocaine on extrapyramidal (striatum and substantia nigra) and limbic (nucleus accumbens and frontal cortex) substance P-like immunoreactivity. Ibogaine and cocaine treatments altered substance P systems by increasing striatal and nigral substance P-like immunoreactivity concentration 12 h after the last drug treatment. However, substance P-like immunoreactivity content was not significantly increased in nucleus accumbens after treatment with either drug. The ibogaine- and cocaine-induced increases in substance P-like immunoreactivity in striatum and substantia nigra were blocked by coadministration of selective dopamine D(1) receptor antagonist (SCH 23390; R(+) 7-Chloro-8-hydroxy-3-methyl-1-phenyl-2,3,4, 5-tetrahydro-1H-3-benzazepine hydrochloride) or dopamine D(2) receptor antagonist (eticlopride; S(-)-3-Chloro-5 ethyl-N-[(1-ethyl-2-pyrrolidinyl)methyl]-6-hydroxy-2- methoxy-benzamide hydrochloride). Most of the responses by substance P systems to ibogaine administration resembled those caused by cocaine, except in cortical tissue where multiple administration of cocaine, but not ibogaine increased substance P-like immunoreactivity. These data suggest that substance P systems may contribute to the effects of ibogaine and cocaine treatment. PMID- 10708716 TI - Effect of amphetamine on extracellular acetylcholine and monoamine levels in subterritories of the rat medial prefrontal cortex. AB - The present study sought to investigate the contributions of the dorsal prelimbic/anterior cingulate and ventral prelimbic/infralimbic cortices to the reverse microdialysis of amphetamine (1, 10, 100, 500, and 1000 microM) on dialysate acetylcholine, choline, norepinephrine, and serotonin levels. The results demonstrate that basal levels of acetylcholine, choline, and serotonin were homogeneous within subregions of the medial prefrontal cortex. In contrast, dialysate norepinephrine levels were significantly higher in the anterior cingulate cortex compared with the infralimbic cortex. Reverse microdialysis of amphetamine in both subareas of the medial prefrontal cortex produced a dose dependent increase in norepinephrine and serotonin levels; the magnitude of this effect was similar in both subterritories of the medial prefrontal cortex. Microinfusion of amphetamine increased dialysate acetylcholine levels in a dose dependent manner only in the infralimbic cortex. Finally, amphetamine decreased choline levels in both subregions of the medial prefrontal cortex. The magnitude of this effect was larger in the anterior cingulate cortex compared with its infralimbic counterpart. Since depletions of frontal cortical acetylcholine result in severe cognitive deficits, the present data raise the possibility that the type of neural integrative processes that acetylcholine mediates depends, at least in part, on the subterritories that characterize the medial prefrontal cortex. PMID- 10708718 TI - Differential effect of dehydroepiandrosterone and its steroid precursor pregnenolone against the behavioural deficits in CO-exposed mice. AB - The neuroactive steroids pregnenolone (3beta-hydroxy-5-pregnen-20-one) and dehydroepiandrosterone (DHEA, 3alpha-hydroxy-5-androstene-17-one) are negative allosteric modulators of the GABA(A) receptors and positive modulators of acetylcholine, NMDA and sigma(1) receptors. Pregnenolone was recently shown to potentiate the neuronal damage induced by excessive glutamate in cell culture models, whereas dehydroepiandrosterone was reported to present some neuroprotective activity. The in vivo relevance of these effects was investigated in mice submitted to an hypoxic insult, the repeated exposure to carbon monoxide (CO) gas, a model that leads to neurodegeneration in the CA(1) hippocampal area and learning deficits. Recording spontaneous alternation behaviour in the Y-maze assessed short-term memory and long-term memory was examined using a passive avoidance task. After exposure to CO, mice showed a progressive deterioration of their learning ability, reaching significance after 3 days and being maximal after 7 days. Pregnenolone administered before CO significantly facilitated the hypoxia-related deficits, which could be measured 1 day after CO and appeared maximal after 3 days. Dizocilpine blocked the deficits in vehicle- and pregnenolone-treated CO-exposed animals, showing that pregnenolone selectively facilitated the NMDA receptor-dependent excitotoxicity. Dehydroepiandrosterone blocked the appearance of the CO-induced deficits, even after 7 days. Interestingly, the sigma(1) receptor antagonist N, N-dipropyl-2-(4-methoxy-3-(2 phenylethoxy)phenyl)ethylamine (NE-100) failed to affect the dehydroepiandrosterone-induced protection, showing the lack of involvement of sigma(1) receptors. Cresyl violet-stained sections of the mouse hippocampal formation showed that the neurodegeneration observed in the CA(1) area after exposure to CO was augmented by pregnenolone and blocked by dehydroepiandrosterone. These results show that pregnenolone and dehydroepiandrosterone, although being similarly involved in modulating the excitatory/inhibitory balance in the brain, do not equally affect the extent of excitotoxic insults. PMID- 10708717 TI - Potentiation of opioid-induced conditioned place preference by the selective serotonin reuptake inhibitor fluoxetine. AB - The ability of the selective serotonin reuptake inhibitor, fluoxetine, to modify the effects of morphine, N-((S)-2-benzyl-3[(S) 2-amino-4-methylthio)butyldithio-] 1-oxopropyl)-L-alanine benzylester (RB 120; mixed inhibitor of enkephalin metabolism), and 4-?[2-[[3-(1H-indol-3-yl))-2-methyl-1-oxo-2-[[(tricyclo[3,3,1,1] dec-2-yloxy) carbonyl] amino? propyl] amino]-1-phenylethyl] amino?-4-oxo-[R (R*,R*)]-butanoate N-methyl-D-glucamine (CI 988; cholecystokinin receptor subtype [CCK(2)] antagonist), was assessed using conditioned place preference. RB 120 and morphine both induced significant, dose-dependent conditioned place preference, whilst CI 988 failed to elicit conditioned place preference. A subthreshold dose of fluoxetine (2.5 mg/kg) potentiated the morphine submaximal response. Notably, the combination of a subthreshold dose of fluoxetine (2.5 mg/kg) with RB 120 (5 mg/kg) or CI 988 (3 mg/kg) was devoid of any significant conditioned place preference properties. Fluoxetine may act via enhanced serotonergic activity to modulate enkephalinergic tone. Agents that increase enkephalinergic tone more directly such as RB 120 and CI 988, at submaximal doses, did not induce conditioned place preference when co-administered with fluoxetine. These data suggest that fluoxetine, in combination with CI 988 or RB 120, might prove to be a beneficial treatment strategy for opioid drug addiction, though further studies are necessary. PMID- 10708719 TI - Force-length relationship in dogs as a measure of protective effect of imidapril on regional myocardial ischemia and reperfusion injury. AB - Our laboratory previously reported that the end-systolic force-length relationship of the left ventricle provides a better method of evaluating myocardial contractile properties than the left ventricular end-systolic pressure volume relationship, because it avoids deficiencies of the latter parameter such as dependence of its slope (E(max)) on the volume intercept (V(0)). The slope (E(c)) of the left ventricular end-systolic force-length relationship represents the contractility of functioning myocardium, while its length intercept (L(0)) reflects the length of non-functioning myocardium. However, the effect of regional myocardial ischemia on these parameters, as evaluated by the force length relationship, remains unknown. To clarify the effects of regional ischemia and angiotensin-converting enzyme inhibition on the myocardium during ischemia reperfusion, the changes in E(c) and L(0) were determined in anesthetized open chest dogs. (1) Control group (n=26): Before and after 15 min of complete coronary artery occlusion, as well as after 15 min of reperfusion, left ventricular pressure and volume were simultaneously recorded during inferior vena cava occlusion. The left ventricular force-length relationship was obtained from the pressure and volume of three cylindrical segments of the ventricle, and E(c) and L(0) were calculated. (2) Imidapril group (n=14): Imidaprilat (1 microg/kg/min) was continuously infused from 30 min before ischemia to the end of the experiment, and the same procedures were followed as in the control group. Fourteen out of the 26 dogs (54%) in the control group died of reperfusion induced ventricular arrhythmias, while only two of the 14 dogs (14%) in the imidapril group did so (P<0.05). In the control group, E(c) was increased during ischemia and remained at the same level after reperfusion. However, E(c) was not altered in the imidapril group. Although L(0) was increased during ischemia and decreased after reperfusion in both groups, the percent increase of L(0) in the imidapril group was significantly smaller than in the control group (8% vs. 32%, P<0.05). With the improvement of these indices, the bradykinin concentration of coronary venous blood increased in the imidapril group (P<0.01). These findings suggest that regional myocardial ischemia increased the average contractility of overall functioning myocardium despite the increased non-functioning myocardium. Moreover, imidapril has a cardioprotective effect against ischemia-reperfusion injury by decreasing infarct size, and through the antiarrhythmic effect and the reversal of increased overall contractility. PMID- 10708720 TI - Positive inotropic effects of adrenomedullin on rat papillary muscle. AB - Adrenomedullin is a peptide recently isolated from pheochromocytoma that has vasorelaxant and long-lasting hypotensive activities. Plasma levels of adrenomedullin are elevated in patients with congestive heart failure, but the effects of adrenomedullin on the cardiac function are unclear. We, thus, investigated the effects of adrenomedullin on the contraction of rat papillary muscles. We measured the isometric tension and cAMP contents of isolated rat papillary muscles. Adrenomedullin exhibited concentration-dependent inotropic effects. Adrenomedullin also significantly increased intracellular contents of cAMP. Addition of the calcitonin gene-related peptide (CGRP) receptor antagonist inhibited both contractile force and cAMP generation of papillary muscles stimulated by adrenomedullin. The adrenomedullin-induced inotropic effect was further increased in the presence of the phosphodiesterase inhibitor, 3-isobutyl 1-methyl-xanthine (IBMX), while the effect was significantly suppressed by KT5720 and Rp-8-bromoadenosine-3',5'-cyclic monophosphorothioate (Rp-8-Br-cAMPS), protein kinase A inhibitors. These results indicate that adrenomedullin has positive inotropic effects on the heart, at least partially through a cAMP dependent pathway. PMID- 10708721 TI - P2X receptors counteract the vasodilatory effects of endothelium derived hyperpolarising factor. AB - Dilatory responses of extracellular nucleotides were examined in the precontracted isolated rat mesenteric artery. Dilatation mediated by endothelium derived hyperpolarising factor (EDHF) was studied in the presence of Nomega-nitro L-arginine (L-NOARG) and indomethacin, and was most potently induced by the selective P2Y(1) receptor agonist adenosine 5'-O-thiodiphosphate (ADPbetaS), while 2-methylthioadenosine triphosphate (2-MeSATP) and adenosine triphosphate (ATP) were almost inactive. However, after P2X receptor desensitisation (with alphabeta-methylene-adenosine triphosphate, alphabeta-MeATP), 2-MeSATP and ATP potently stimulated EDHF-mediated dilatation. This can be explained by simultaneous activation of endothelial P2Y receptors that release EDHF, and depolarising P2X receptors on smooth muscle cells. Uridine triphosphate (UTP) also induced potent dilatation, suggesting EDHF release via P2Y(2)/P2Y(4) receptors. Uridine diphosphate (UDP) had only minor dilatory effects, and when pretreated with hexokinase it was almost inactive, suggesting a minor role for P2Y(6) receptors. The nitric oxide (NO) mediated dilatation was studied in the presence of charybdotoxin, apamin and indomethacin. ADPbetaS, 2-MeSATP, ATP and UTP were all potent relaxant agonists suggesting NO release via P2Y(1) and P2Y(2)/P2Y(4) receptors, while UDP was much less potent and efficacious. P2X receptor desensitisation had only minor effect on the NO-mediated dilatations. In conclusion, both EDHF and NO-mediated dilatation can be induced by activation of P2Y(1) and P2Y(2)/P2Y(4) receptors. P2X receptor stimulation of smooth muscle cells selectively counteracts the dilatory effect of EDHF. PMID- 10708722 TI - Effects of tamoxifen on oxyhemoglobin-induced cerebral vasoconstriction. AB - The effect of tamoxifen on oxyhemoglobin-mediated cerebral vasoconstriction was examined. Tamoxifen caused a concentration-dependent relaxation of cerebral artery preparations contracted with oxyhemoglobin and phorbol myristate acetate with the IC(50) values 0.66+/-0.1 and 1.1+/-0.1 microM, respectively. In cerebrovascular smooth muscle cells, oxyhemoglobin and phorbol myristate acetate induced protein kinase C activation, which was 220+/-7% and 203+/-8% of control, respectively. The increase in protein kinase C activity was prevented by tamoxifen. The results suggest that the ability of tamoxifen to reverse vasoconstriction is mediated, at least in part, via inhibition of protein kinase C. PMID- 10708723 TI - The mechanisms of alpha(2)-adrenoceptor agonist-induced contraction in longitudinal muscle of the porcine uterus. AB - The aim of the present study was to clarify the cellular mechanisms underlying the alpha(2)-adrenoceptor-mediated contraction of porcine myometrium (nonvascular smooth muscle). Acetylcholine (3 nM-1 microM), clonidine (1 nM-10 microM) and 5 bromo-N-[2-imidazolin-2-yl]-6-quinoxalinamine (UK14304) (1 nM-10 microM) in Krebs solution caused a concentration-dependent contraction in the longitudinal muscles of the porcine uterus with similar EC(50) values and maximum responses. A lowered external Ca(2+) concentration and verapamil (10 nM-10 microM) decreased the contractile response to clonidine and UK14304 more markedly than the response to acetylcholine. However, in Kumagai solution, neither clonidine nor UK14304 caused contractile responses, but acetylcholine remained effective. The effects of alpha(2)-adrenoceptor agonists on intracellular Ca(2+) concentration ([Ca(2+)](i)) and smooth muscle force were measured simultaneously using fura-PE3 loaded muscle preparations. Clonidine and UK14304 caused increases in [Ca(2+)](i) and force of the longitudinal muscle. The increases in [Ca(2+)](i) and muscle force were markedly inhibited by verapamil and in Ca(2+)-free solution (EGTA, 1 mM). In the absence of external Ca(2+), clonidine caused only a small increase in [Ca(2+)](i) in Ca(2+)-loaded preparations compared with those increases caused by carbachol, histamine, and oxytocin. Ca(2+) (2.5 mM) caused increases in [Ca(2+)](i) and force of the longitudinal muscles in a Ca(2+)-free high K(+) solution. Clonidine concentration dependently potentiated the Ca(2+)-induced contraction without significantly changing the increase in [Ca(2+)](i), and this potentiation was inhibited by yohimbine. These results suggested that clonidine increases the Ca(2+) sensitivity of the contractile elements through activation of alpha(2)-adrenoceptors. During the development of the contractile response to clonidine (1 microM, 0-5 min), tissue cyclic AMP levels did not change significantly. In vitro treatment with pertussis toxin (1 microg/ml for 2 h) significantly decreased the contraction induced by clonidine without affecting the responses to carbachol and high K(+). The present results indicate that in porcine myometrium, alpha(2)-adrenoceptor stimulation caused contraction of the longitudinal muscles by mechanisms largely dependent on the influx of extracellular Ca(2+), probably through voltage-dependent Ca(2+) channels (VDCCs), and that the potentiation of the Ca(2+) sensitivity of the contractile elements is another mechanism of the contractile responses. These actions involve a pertussis-toxin-sensitive G protein (probably G(i) type) in the signal transduction pathway. PMID- 10708724 TI - Neutrophil elastase inhibitor, ONO-5046 suppresses ozone-induced airway mucus hypersecretion in guinea pigs. AB - To investigate the role of neutrophil elastase in ozone-induced airway hypersecretion, we measured goblet cell secretion by using a semiquantitative morphometric technique in guinea pigs. The magnitude of mucus discharge was estimated from the mucus score, which is inversely related to the degree of mucus discharge in histological sections of trachea stained for mucus glycoprotein with periodic acid Schiff/Alcian blue. Mucus hypersecretion of goblet cells was induced by ozone exposure and persisted for up to 5 h after exposure. Pretreatment with N-[2-?4-(2,2-dimethyl-propionyloxy) phenyl-sulfonylamino? benzoyl] aminoacetic acid (ONO-5046), a specific neutrophil elastase inhibitor (200 mg/kg, intraperitoneally), significantly inhibited goblet cell hypersecretion both just after and 5 h after ozone-exposure, but the latter inhibition was not complete. In bronchoalveolar lavage fluid, ozone exposure significantly increased the number of neutrophils just after and 5 h after exposure, while ONO-5046 significantly inhibited the increase in neutrophils only 5 h after ozone-exposure. These results indicate that neutrophil elastase may play an important role in the ozone-induced tracheal goblet cell hypersecretion and influx of neutrophils. PMID- 10708725 TI - Effect of human plasma-type platelet-activating factor acetylhydrolase in two anaphylactic shock models. AB - The effect of human recombinant plasma-type platelet-activating factor (PAF) acetylhydrolase was examined in two murine models, PAF-induced death and active anaphylactic models. In the PAF-induced death model where mice were injected intravenously with 40 microg/kg of PAF, the administration of PAF acetylhydrolase reduced mortality in a dose-dependent manner, showing complete prevention of mortality at 1.0 mg/kg. Myeloperoxidase activity, the marker for neutrophils, was increased in the lung by PAF injection, and the PAF acetylhydrolase treatment significantly reversed the increase in myeloperoxidase activity. As in the PAF induced model, PAF acetylhydrolase also decreased mortality in the active anaphylactic shock model where bovine serum albumin was injected intravenously to mice previously immunized with bovine serum albumin. The protective effect of PAF acetylhydrolase on mortality in this model was significant at 1.0 mg/kg. These results suggest that PAF is an important mediator in the lethality of systemic anaphylaxis, and that PAF acetylhydrolase may be beneficial for treatment of anaphylactic shock. PMID- 10708726 TI - Effects of tempol, a membrane-permeable radical scavenger, in a rodent model of carrageenan-induced pleurisy. AB - Carrageenan causes enhanced formation of reactive oxygen species, which contribute to the pathophysiology of inflammation. We have investigated the effects of tempol, a membrane-permeable radical scavenger, in rats subjected to carrageenan-induced pleurisy. Treatment of rats with tempol (10, 30, or 100 mg/kg 15 min prior to carrageenan) attenuated the pleural exudation and the migration of polymorphonuclear cells caused by carrageenan dose dependently. Tempol also attenuated the lung injury (histology) as well as the increase in the tissue levels of myeloperoxidase and malondialdehyde caused by carrageenan in the lung. However, tempol did not inhibit the activity of inducible nitric oxide synthase in the lungs. Immunohistochemical analysis for nitrotyrosine revealed positive staining in lungs from carrageenan-treated rats. Lung tissue sections from carrageenan-treated rats also showed positive staining for poly-(ADP-ribose) synthetase (PARS). The degree of staining for nitrotyrosine and PARS was markedly reduced in tissue sections obtained from carrageenan-treated rats, which had received tempol (100 mg/kg). Furthermore, treatment of rats with tempol significantly reduced (i) the formation of peroxynitrite, (ii) the DNA damage, (iii) the impairment in mitochondrial respiration, and (iv) the fall in the cellular level of NAD(+) observed in macrophages harvested from the pleural cavity of rats treated with carrageenan. Tempol also attenuated the cell injury caused by hydrogen peroxide (1 mM) in cultured human endothelial cells. This study provides the first evidence that tempol, a small molecule which permeates biological membranes and scavenges ROS, attenuates the degree of inflammation and tissue damage associated with carageenan-induced pleurisy in the rat. The mechanisms of the anti-inflammatory effect of tempol are discussed. PMID- 10708727 TI - Anti-inflammatory action of a novel peptide, SEK-1005, isolated from a Streptomyces. AB - The pharmacological activity of a novel cyclic peptide (SEK-1005: C(45)H(70)N(8)0(13)) isolated from Streptomyces nobilis was studied in rats during the development of inflammation. SEK-1005 (0.1-0.5 mg/kg, i.p.) suppressed the passive Arthus reaction and the carrageenin-induced oedema. A steroidal anti inflammatory drug, prednisolone (10 mg/kg, i.p.), also was effective on both inflammations. However, indomethacin (5 mg/kg, i.p.), a cyclooxygenase inhibitor, was less effective on the passive Arthus reaction. Also interesting was that the SEK-1005 effect showed its maximum level after a 24-h lag period and that its effect, as well as the prednisolone effect, was reduced by the treatment with a protein synthesis inhibitor, cycloheximide. SEK-1005 and prednisolone also showed marked protection against the adjuvant-induced arthritis, but failed to prevent the tuberculin response. These findings indicate that SEK-1005 is a new type of non-steroidal anti-inflammatory agent with an action similar to that of prednisolone. PMID- 10708728 TI - Leu(10) of alpha-conotoxin PnIB confers potency for neuronal nicotinic responses in bovine chromaffin cells. AB - Two alpha-conotoxins PnIA and PnIB (previously reported as being "mollusc specific") which differ in only two amino acid residues (AN versus LS at residues 10 and 11, respectively), show markedly different inhibition of the neuronal nicotinic acetylcholine receptor response in bovine chromaffin cells, a mammalian preparation. Whereas alpha-conotoxin PnIB completely inhibits the nicotine-evoked catecholamine release at 10 microM, with IC(50) = 0.7 microM, alpha-conotoxin PnIA is some 30-40 times less potent. Two peptide analogues, [A10L]PnIA and [N11S]PnIA were synthesized to investigate the extent to which each residue contributes to activity. [A10L]PnIA (IC(50) = 2.0 microM) completely inhibits catecholamine release at 10 microM whereas [N11S]PnIA shows little inhibition. In contrast, none of the peptides inhibit muscle-type nicotinic responses in the rat hemi-diaphragm preparation. We conclude that the enhanced potency of alpha conotoxin PnIB over alpha-conotoxin PnIA in the neuronal-type nicotinic response is principally determined by the larger, more hydrophobic leucine residue at position 10 in alpha-conotoxin PnIB. PMID- 10708729 TI - Troglitazone does not initiate hypertrophy but can sensitise cardiomyocytes to growth effects of serum. AB - Chronic administration of troglitazone might predispose to cardiac hypertrophy. The aims of the study were to determine if troglitazone could (i) initiate a trophic response directly in ventricular cardiomyocytes and (ii) modify responses to other trophic stimuli. After 24 h, troglitazone (10 nM-10 microM) (i) did not increase cellular protein mass and decreased incorporation of [14C]phenylalanine, a marker of protein synthesis, (ii) interacted with serum (10% v/v) and insulin like growth factor-1 (10 nM) to produce small trophic responses, (iii) increased cellular protein mass but not protein synthesis with insulin (1 unit/ml). Troglitazone (1 microM) attenuated responses to phorbol-12-myristate-13-acetate (PMA) (100 nM), and noradrenaline (5 microM) and endothelin-1 (100 nM), which also activate protein kinase C. In summary, troglitazone does not initiate cardiomyocyte growth directly in vitro, and can inhibit protein kinase C-mediated growth mechanisms. However, the interaction of troglitazone with serum growth factors may contribute modestly to the development of hypertrophy. As troglitazone produced a moderate hypertrophic effect per se in re-differentiated cardiomyocytes, it may directly increase the severity of established hypertrophy. PMID- 10708730 TI - Decreased binding affinity of olanzapine and clozapine for human muscarinic receptors in intact clonal cells in physiological medium. AB - The binding affinity of olanzapine, clozapine and atropine for muscarinic receptor subtypes in clonal Chinese hamster ovary (CHO) cell lines was compared in intact cells in physiological media to disrupted cells in hypotonic buffer. The affinity of olanzapine and clozapine, but not atropine, for muscarinic receptor subtypes (M(1)-M(5)) was significantly reduced in intact cells in physiological medium compared to disrupted cells in hypotonic buffer. The affinity of olanzapine for muscarinic M(1) receptors was most affected with a reduction of K(i) value from 2.5 to 73 nM in intact cells. These data suggest that the affinity of olanzapine and clozapine for muscarinic receptors have been significantly overestimated. PMID- 10708731 TI - N-tosyl-L-phenylalanyl-chloromethylketone reduces hypoxic-ischemic brain injury in rat pups. AB - N-tosyl-L-phenylalanyl-chloromethylketone (TPCK) in vitro blocks apoptotic pathways leading to cell death. We wished to see if TPCK would reduce brain injury in vivo. Seven-day-old rat pups had the right carotid artery ligated and then received either vehicle or TPCK (5 to 100 mg/kg i.p.). They were then given 8% oxygen for 2.25 h. Twenty-two days later, the cerebral hemispheres were weighed to determine the reduction in size in the right hemisphere. TPCK decreased the reduction in right hemisphere weight from 15+/-3% (vehicle, n=20), to 4+/-2% (10 mg/kg, n = 19, P<0.01). TPCK reduced the number of cells staining for DNA breaks 3 days after injury from 1729+/-275 mm(-2) (vehicle, n = 8) to 550+/-236 mm(-2) (10 mg/kg TPCK, n = 9, P<0.01), decreased the amount of DNA fragmentation 3 days after injury by gel electrophoreses (20 mg/kg, n = 16, P<0.01) and eliminated the increase in nitric oxide metabolites 6 h after injury (vehicle 1.5+/-0.4, n = 10; and 20 mg/kg TPCK 0.0+/-0.1 nM/mg protein, n = 10, P<0.001). TPCK pretreatment in the newborn rat model of hypoxic-ischemic brain injury reduces DNA fragmentation, nitric oxide production and brain injury. PMID- 10708732 TI - Possible involvement of the locus coeruleus in inhibition by prostanoid EP(3) receptor-selective agonists of morphine withdrawal syndrome in rats. AB - We examined the mechanism of the inhibitory effect of prostanoid EP(3) receptor agonists on naloxone-precipitated withdrawal syndrome in morphine-dependent rats. Rats were rendered morphine dependent by subcutaneous (s.c.) implantation of two pellets containing 75 mg morphine for 5 days. Morphine withdrawal syndrome was precipitated by i.p. injection of naloxone (3 mg/kg). Intracerebroventricular (i.c.v.) administration of (+/-)-15alpha-hydroxy-9-oxo-16-phenoxy-17,18, 19,20 tetranorprost-13-trans-enoic acid (M&B28,767: prostanoid EP(3) receptor agonist) or sulprostone (prostanoid EP(1)/EP(3) receptor agonist) significantly suppressed many withdrawal signs. Northern blotting and in situ hybridization studies revealed that i.c.v. administration of M&B28,767 (1 pg/rat) attenuated the elevation of c-fos mRNA during naloxone-precipitated withdrawal in many brain regions, including the cerebral cortex, thalamus, hypothalamus and locus coeruleus. Double in situ hybridization analysis revealed that in the locus coeruleus most of the tyrosine hydroxylase mRNA-positive neurons expressed mu opioid receptor mRNA and more than half of these neurons were positive for prostanoid EP(3) receptor mRNA. These results indicate that the suppression by prostanoid EP(3) receptor agonists of naloxone-precipitated morphine withdrawal syndrome can be attributed to the inhibition of neuronal activity in several brain regions, including the locus coeruleus, the largest source of central noradrenergic neurons. PMID- 10708733 TI - Systemic administration of 4-chlorokynurenine prevents quinolinate neurotoxicity in the rat hippocampus. AB - The synthetic compound 4-chlorokynurenine has been shown to be enzymatically transaminated to the selective glycine(B) receptor antagonist 7-chlorokynurenate. Since 4-chlorokynurenine, in contrast to 7-chlorokynurenate, readily penetrates the blood-brain barrier, the present study evaluated its neuroprotective properties after systemic administration in rats. Intrahippocampal injection of the NMDA receptor agonist quinolinate (15 nmol/l microl) was used as the neurotoxic paradigm. Serum and hippocampal tissue measurements confirmed that 4 chlorokynurenine serves as an effective pro-drug of 7-chlorokynurenate both in the periphery and in the brain. These studies and complementary hippocampal microdialysis experiments compared the effects of single and repeated injections of 4-chlorokynurenine (50 or 200 mg/kg, intraperitoneal (i.p.), 10 min prior to an intrahippocampal quinolinate injection; or 50 mg/kg, i.p., 10 min before and 30, 120 and 360 min after quinolinate). With the multiple-dosing regimen, extracellular 7-chlorokynurenate levels in the hippocampus reached a maximum of approximately 750 nM 7 h after quinolinate and gradually decreased with a half life of about 3 h. In contrast, a single injection of 200 mg/kg 4 chlorokynurenine resulted in a considerably shorter rise in extracellular 7 chlorokynurenate without yielding higher peak levels. In separate animals, repeated treatment with 50 mg/kg 4-chlorokynurenine, but not a single injection of 200 mg/kg of the pro-drug, provided total protection against quinolinate induced excitotoxicity. These data suggest that a prolonged and functionally relevant blockade of hippocampal glycine(B) receptors can be achieved after the systemic administration of 4-chlorokynurenine. PMID- 10708734 TI - Characterization of D-fenfluramine-induced hypothermia: evidence for multiple sites of action. AB - The effects of D-fenfluramine on core body temperature has been largely investigated under conditions of either high or low ambient temperature, whereas little research has focused on this response under normal environmental conditions. Moreover, there has been neglect in research on the mechanisms underlying changes in body temperature. In this study, we demonstrate that D fenfluramine (5 and 10 mg/kg) induces a sustained decrease in body temperature in the rat under normal ambient temperatures. Pre-treatment with the selective serotonin reuptake inhibitor sertraline (5 mg/kg), the full 5-HT(1A) receptor antagonist 4-fluoro-N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-2-pyridinyl benzamide], WAY 100635 (0.15 mg/kg) and the 5-HT(2C) receptor antagonist benzofuran-2-carboxamidine, RO 43-0440 (2.5 mg/kg) blocked D-fenfluramine-induced hypothermia. Depletion of 5-hydroxytryptamine (5-HT) stores following treatment with the serotonergic neurotoxin parachlorophenylalanine reversed the initial hypothermic effects of D-fenfluramine but not the later effects, as D120 min post challenge) in animals pre-treated with parachlorophenylalanine. Such findings are consistent with a requirement for D-fenfluramine uptake into 5-HT neurons followed by release of 5-HT from intracellular stores and stimulation of post synaptic 5-HT receptors to reduce body temperature. The hypothermic response to D fenfluramine was potentiated by ketanserin pre-treatment 30 min post-challenge but then antagonized at later time intervals. Pre-treatment with the dopamine, D(2) antagonist, haloperidol (1 mg/kg) and sulpiride (30 mg/kg) had a similar effect in blocking the hypothermia as WAY 100635, suggesting a role for dopamine D(2) receptors in the response. Pre-treatment with the alpha(2)-adrenoceptor antagonist yohimbine failed to block the hypothermic response. These results suggest multiple sites of action mediating D-fenfluramine-induced hypothermia and may be the result of a combined effect of D-fenfluramine and its active metabolite norfenfluramine affecting not only the release of 5-HT but also stimulation of post-synaptic receptors. PMID- 10708735 TI - Effects of ketamine on synaptic transmission and long-term potentiation in layer II/III of rat visual cortex in vitro. AB - The effects of ketamine, which has NMDA receptor antagonist properties, on synaptic transmission and long-term potentiation in layer II/III of adult rat visual cortex were examined in vitro. Field potentials were recorded in layer II/III following layer IV stimulation. Primed-burst stimulation was used for induction of long-term potentiation. Stimulation of layer IV resulted in a two component response in layer II/III, a population excitatory postsynaptic potential1 (EPSP1) and a population excitatory postsynaptic potential2 (EPSP2). DL-2-Amino-5-phosphono-valeric acid (AP5), a competitive NMDA receptor antagonist, reduced the amplitude of the population EPSP1 while ketamine increased the amplitude of the population EPSP2. The results showed that primed burst stimulation induced long-term potentiation in layer II/III of the visual cortex in vitro. Preincubation for 30 min with AP5 (25-100 microM) reduced the extent of long-term potentiation of the population EPSP2 and blocked the induction of long-term potentiation of the population EPSP1. When ketamine (100 200 microM) was present for 30 min prior to tetanic stimulation, it blocked the induction of long-term potentiation of the population EPSP1 and reduced the extent of long-term potentiation of the population EPSP2. We conclude that ketamine can interfere with synaptic transmission in the visual cortex. Primed burst stimulation is an effective protocol for neocortical potentiation. NMDA receptors are involved in the induction of long-term potentiation by primed-burst stimulation of the population EPSP1 and population EPSP2 in adult rat visual cortex in vitro. PMID- 10708736 TI - Nimesulide limits kainate-induced oxidative damage in the rat hippocampus. AB - Kainate induces a marked expression of cyclooxygenase-2 after its systemic administration. Because cyclooxygenase-2 activity is associated to the production of reactive oxygen species, we investigated the effects of nimesulide, a selective cyclooxygenase-2 inhibitor, on kainate-induced in vivo oxidative damage in the rat hippocampus. A clinically relevant dose of nimesulide (6 mg/kg, i.p. ) was administered three times following kainate application (9 mg/kg, i.p.). After 24 h of kainate administration, the drastic decrease in hippocampal glutathione content and the significant increase in lipid peroxidation were attenuated in nimesulide-treated rats, suggesting that the induction of cyclooxygenase-2 is involved in kainate-mediated free radicals formation. PMID- 10708737 TI - Melatonin treatment does not prevent decreases in brain glutathione levels induced by sleep deprivation. AB - Recent findings from this laboratory revealed that sleep deprivation reduces total glutathione (GSH) levels in hypothalamus, suggesting an increased vulnerability to oxidative damage. Since melatonin has been shown to prevent oxidative damage in other experimental situations, the present study tested the effects of exogenous melatonin on sleep deprivation-induced GSH decreases. Rats were deprived of sleep for 96 h on small platforms, and melatonin (10 mg/kg body weight; i.p.) or vehicle was given twice a day. Hypothalamic GSH levels were significantly reduced in sleep-deprived groups, irrespective of melatonin treatment. Indeed, unexpectedly, melatonin treatment resulted in lower hypothalamic GSH levels in all groups, including cage controls. These results confirm that sleep deprivation reduces hypothalamic GSH and further indicate that melatonin treatment not only is ineffective in reversing this effect but may actually potentiate it. PMID- 10708738 TI - Effects of NMDA receptor antagonists on cocaine-conditioned motor activity in rats. AB - NMDA receptor antagonists have been reported to affect learned behaviors conditioned with abused drugs, with the outcome dependent, in part, on the class of NMDA receptor antagonist used. The present study tested the ability of various site-selective NMDA receptor antagonists to modify cocaine-conditioned motor activity. Two procedures were used for independently assessing drug effects on spontaneous activity and expression of cocaine-conditioned behavior. In the conditioning experiments, rats were administered i.p. injections of cocaine (30 mg/kg) or saline paired with distinctive environments. Spontaneous horizontal activity was dose-dependently enhanced by dizocilpine (0.03-0.3 mg/kg) and memantine (1-30 mg/kg), but not by D-CPPene (3-(2-carboxypiperazin-4-yl)-1 propenyl-1-phosphonic acid; SDZ EAA 494; 1-10 mg/kg), ACEA-1021 (5-nitro-6,7 dichloro-1,4-dihydro-2, 3-quinoxalinedione; 3-56 mg/kg), or eliprodil (3-30 mg/kg). Higher doses of memantine, D-CPPene (1-10 mg/kg), eliprodil (3-30 mg/kg), or ACEA-1021 reduced vertical activity. Following five cocaine-environment pairings, rats displayed significant increases in motor activity when exposed to the cocaine-paired environment. The following antagonists were administered prior to the conditioning test: dizocilpine (MK-801; 0.03-0.1 mg/kg), memantine (1-10 mg/kg), D-CPPene (0.3-3 mg/kg), ACEA-1021 (3-10 mg/kg), and eliprodil (1-10 mg/kg). Of these, memantine, ACEA-1021 and, to the lesser degree, eliprodil attenuated expression of cocaine-conditioned motor activity at doses that did not significantly affect spontaneous motor activity. These results show that cocaine conditioned behaviors can be selectively modulated by some, but not all, NMDA receptor antagonists. PMID- 10708739 TI - Pre-training blocks the improving effect of tetrahydroaminoacridine and D cycloserine on spatial navigation performance in aged rats. AB - We investigated the effect of pre-training on the improvement of spatial navigation performance provided by a cholinesterase inhibitor, tetrahydroaminoacridine (3 mg/kg, i.p.), and a positive modulator of NMDA receptor, D-cycloserine (10 mg/kg, i.p.), or their combination in aged rats. Pre training consisted of spatial or non-spatial conditions and took place in either the same or a separate room. We found that any kind of pre-training was able to eliminate the enhancing effect of tetrahydroaminoacridine and D-cycloserine on spatial navigation. However, none of these pre-training conditions was able to block the age-related deficit in spatial navigation. These results indicate that tetrahydroaminoacridine and D-cycloserine, separately or in combination, do not themselves alleviate the age-related spatial memory deficit, but may enhance procedural aspects of water maze learning in aged rats. PMID- 10708740 TI - Vasorelaxant effect of harman. AB - The in vivo cardiovascular effect and in vitro vasorelaxant effect of harman, one of harmala alkaloids isolated from Peganum harmala, were examined in this study. Harman (1-10 mg/kg, i.v.) dose-dependently produced transient hypotension and long-lasting bradycardia in pentobarbital-anesthetized rats, which were attenuated by N(G)-nitro-L-arginine pretreatment. In isolated rat endothelium intact thoracic aortic rings, harman concentration dependently relaxed phenylepherine-induced contractions with an IC(50) value around 9 microM. This vasorelaxant effect was attenuated by endothelium removal or N(omega)-nitro-L arginine methyl ester pretreatment, but not by tetraethylammonium or indomethacin pretreatment. In cultured rat aortic endothelial cells, harman (1-100 microM) concentration dependently increased nitric oxide (NO) release, which was dependent on the presence of external Ca(2+). Harman pretreatment (3-30 microM) also concentration dependently inhibited the contractions induced by phenylephrine, 5-hydroxytryptamine (5-HT), and KCl in endothelium-denuded aortic rings in a non-competitive manner. In addition, harman pretreatment reduced both phasic and tonic phases of phenylephrine-induced contractions. Receptor binding assays further indicated that harman (K(i) values around 5-141 microM) interacted with the cardiac alpha(1)-adrenoceptors, brain 5-HT(2) receptors, and cardiac 1, 4-dihydropyridine binding site of L-type Ca(2+) channels. Therefore, the present results suggested that the vasorelaxant effect of harman was attributed to its actions on the endothelial cells to release NO and on the vascular smooth muscles to inhibit the contractions induced by the activation of receptor-linked and voltage-dependent Ca(2+) channels. The vasorelaxant effect may be involved in the hypotensive effect of harman. PMID- 10708741 TI - Protective effect of oral L-arginine administration on gentamicin-induced renal failure in rats. AB - We investigated the effects of orally supplemented L-arginine, the substrate of nitric oxide (NO) and N(omega)-nitro-L-arginine methyl ester (L-NAME), a nitric oxide-synthase inhibitor in gentamicin-induced renal failure. Rats were given gentamicin (100 mg/kg/day s.c.), gentamicin and L-arginine (2 g/l, drinking water), gentamicin and L-NAME (100 mg/l, drinking water) or gentamicin plus L arginine and L-NAME. After 8 days, the gentamicin group developed marked renal failure, characterized by a significantly decreased creatinine clearance and increased blood creatinine, fractional excretion of sodium, fractional excretion of lithium, urine gamma glutamyl transferase, systolic blood pressure and daily urine volume when compared to controls. Renal histological analysis confirmed tubular necrosis. L-arginine administration caused normalization of these parameters, whereas L-NAME led to aggravation of the failure. Concomitant administration of L-NAME and L-arginine to gentamicin-treated rats caused no significant changes when compared to the rats receiving gentamicin alone. We conclude that L-arginine supplementation has beneficial effects in gentamicin induced renal failure in rats and that these effects are reversed by the NO synthase inhibitor, L-NAME. PMID- 10708742 TI - Dual effect of [D-Pen(2), D-Pen(5)]enkephalin on ion transport in guinea pig colon. AB - Effects of the delta-opioid receptor ligand, [D-Pen(2), D-Pen(5)]enkephalin (DPDPE) on basal and endothelin-1-induced ion secretion in guinea pig colon were investigated. Muscle-stripped segments of guinea pig colon were mounted in Ussing flux chambers and changes in the short-circuit current (I(sc)) were monitored continuously. DPDPE significantly reduced baseline I(sc) at a low dose, 1 nM; however DPDPE increased I(sc) at 10 and 100 microM. Endothelin-1 stimulated ion secretion that was unaltered in tissues pretreated with DPDPE. In guinea pig colon, delta-opioid receptor activation evoked both a proabsorptive and prosecretory response. PMID- 10708743 TI - 4-Hydroxyisoleucine: effects of synthetic and natural analogues on insulin secretion. AB - 4-Hydroxyisoleucine, a peculiar amino acid extracted from fenugreek seeds and never found in mammalian tissues, exhibits interesting insulinotropic activity. To investigate the structural requirements for this stimulating effect, the insulinotropic activity of the major isomer (2S,3R,4S) of 4-hydroxyisoleucine, in the presence of 8. 3 mM glucose, was compared to that of (1) its minor isomer (2R,3R, 4S) (2) its lactone form, (3) classical structurally related amino acids, and (4) synthetic monomethylated analogues. In the isolated, ex vivo, perfused rat pancreas, only the major isomer of 4-hydroxyisoleucine (200 microM) potentiated insulin release. On incubated isolated rat islets, the threshold concentration for a significant increase (P<0.05) in insulin release was 200 microM for (2S,3R,4S) 4-hydroxyisoleucine, 500 microM for (2S,4R) and (2S,4S) gamma-hydroxynorvalines as well as (2S,3S) and (2S,3R) gamma-hydroxyvalines, and 1 mM or more for other congeners. In conclusion, the insulinotropic properties of 4-hydroxyisoleucine, in the micromolar range, are seen only in the presence of the linear major isoform; they also require carbon alpha in S-configuration, full methylation and carbon gamma-hydroxylation. PMID- 10708744 TI - Expression of type XVI collagen in cultured skin fibroblasts is related to cell growth arrest. AB - The expression of type XVI collagen in various phases of cell growth in cultured skin fibroblasts was studied. A marked increase in type XVI collagen mRNA level was found in stationary phases of cell growth (non-adherent and confluent phases), whereas the expression of type I and III collagens was undetectable in the non-adherent phase but became greater in the confluent phase. When suspended cells were further cultured over 72 h (suspension arrest), mRNA level and gene transcription of type XVI collagen were time-dependently increased whereas those of type I collagen remained undetectable. When the confluent cells were further cultured for 72 h under the condition of serum deprivation (serum deprivation arrest), mRNA levels of both type XVI collagen and type I collagen were elevated. The level of type XVI collagen polypeptide in the culture media of suspension arrested and serum deprivation-arrested cells paralleled the mRNA level of type XVI collagen. The results indicate that expression of type XVI collagen (a member of the fibril-associated collagens with interrupted triple helices), unlike interstitial collagens (type I collagen), is related to cell growth arrest brought about by two different growth inhibiting systems, suspension arrest and serum deprivation arrest. PMID- 10708745 TI - Lactoferrin inhibits the binding of lipopolysaccharides to L-selectin and subsequent production of reactive oxygen species by neutrophils. AB - The activation of leukocytes by lipopolysaccharides (LPS), resulting in the oxidative burst, contributes to the pathogenesis of septic shock. The binding of LPS to L-selectin, which was reported as a serum-independent LPS receptor on neutrophils, induces the production of oxygen free radicals. Human lactoferrin (hLf), an anti-inflammatory glycoprotein released from neutrophil granules during infection, binds to LPS. In this study, we investigated the capacity of hLf to inhibit the L-selectin-mediated activation of neutrophils. Our experiments revealed that hLf prevents the binding of LPS to L-selectin in a concentration dependent manner. Inhibition was maximum (87.7+/-0.5%) at a concentration of 50 microg/ml of hLf. Furthermore, hLf inhibited up to 55.4+/-0.5% of the intracellular hydrogen peroxide production induced by LPS in neutrophils. These findings suggest that the anti-inflammatory properties of hLf are due, at least in part, to their ability to prevent the binding of LPS to neutrophil L-selectin. PMID- 10708746 TI - Novel activity of a phycobiliprotein lyase: both the attachment of phycocyanobilin and the isomerization to phycoviolobilin are catalyzed by the proteins PecE and PecF encoded by the phycoerythrocyanin operon. AB - The structure of phycoviolobilin, the photoactive chromophore of alpha phycoerythrocyanin, is incompatible with a chromophore ligation to the apoprotein via SH-addition (cysteine) to a Delta3, 3(1)-double bond of the phycobilin. The two putative phycoerythrocyanin lyase genes of Mastigocladus laminosus, pecE and pecF, were overexpressed in Escherichia coli. Their action has been studied on the addition reaction of phycocyanobilin to apo-alpha-phycoerythrocyanin (PecA). In the absence of the components of alpha-PEC-phycoviolobilin lyase PecE and PecF, or in the presence of only one of them, phycocyanobilin binds covalently to PecA forming a fluorescent chromoprotein with a red-shifted absorption (lambda(max)=641 nm) and low photoactivity (<10%). In the presence of both PecE and PecF, a chromoprotein forms which by its absorption (lambda(max)=565 nm) and high photoreversible photochromism (100% type I) has been identified as integral alpha-phycoerythrocyanin. We conclude that PecE and PecF jointly catalyze not only the addition of phycocyanobilin to PecA, but also its isomerization to the native phycoviolobilin chromophore. PMID- 10708747 TI - Identification of a vascular endothelial growth factor (VEGF) antagonist, sFlt-1, from a human hematopoietic cell line NALM-16. AB - An antagonistic activity against vascular endothelial growth factor (VEGF) was identified in the culture supernatants of certain human hematopoietic cell lines and the antagonistic protein was purified from NALM-16 (B cell) culture supernatant. Amino acid sequencing of the N-terminus and Western blot analysis confirmed that the antagonist was identical to a soluble truncated form of Flt-1 (sFlt-1). Seventeen of 52 leukemia and lymphoma cell lines investigated expressed sFlt-1 mRNA, and 16 of the sFlt-1 expressing cells also expressed VEGF and membrane-bound Flt-1 (mFlt-1). This report is the first showing that sFlt-1 can be produced by malignant hematopoietic cells, suggesting that the production of VEGF antagonist by hematopoietic cells may play some role in the regulation of VEGF activity in normal and malignant hematopoietic cell proliferation. PMID- 10708748 TI - Molecular cloning of MINK, a novel member of mammalian GCK family kinases, which is up-regulated during postnatal mouse cerebral development. AB - A new germinal center kinase (GCK) family kinase, Misshapen/NIKs-related kinase (MINK), has been cloned and its expression has been characterized in several tissues and various developmental stages of the mouse brain. MINK encodes a 1300 amino acid polypeptide, consisting of an N-terminal kinase domain, a proline-rich intermediate region, and a C-terminal GCK homology region. The expression of MINK is up-regulated during the postnatal development of the mouse brain. MINK activates the cJun N-terminal kinase and the p38 pathways. PMID- 10708749 TI - Analysis of the fine specificity of Tn-binding proteins using synthetic glycopeptide epitopes and a biosensor based on surface plasmon resonance spectroscopy. AB - Using synthetic Tn (GalNAc-O-Ser/Thr) glycopeptide models and a biosensor based on surface plasmon resonance spectroscopy we have determined that isolectin B4 from Vicia villosa (VVLB4) binds to one Tn determinant whereas the anti-Tn monoclonal antibodies 83D4 and MLS128 require at least two Tn residues for recognition. When an unglycosylated amino acid is introduced between the Tn residues, both antibodies do not bind. MLS128 affinity was higher on a glycopeptide with three consecutive Tn residues. These results indicate that Tn residues organized in clusters are essential for the binding of these antibodies and indicate a different Tn recognition pattern for VVLB4. PMID- 10708750 TI - The high stability of cruzipain against pH-induced inactivation is not dependent on its C-terminal domain. AB - Unlike mammalian lysosomal cysteine proteases, the trypanosomal cysteine protease cruzipain contains a 130-amino acid residue C-terminal domain, in addition to the catalytic domain, and it is stable at neutral pH. The endogenous (with C-terminal domain) and recombinant (without C-terminal domain) cruzipains exhibit similar stabilities at both acid (k(inac)=3.1x10(-3) s(-1) and 4.4x10(-3) s(-1) at pH 2.75 for endogenous and recombinant cruzipain, respectively) and alkaline pH (k(inac)=3.0x10(-3) s(-1) and 3. 7x10(-3) s(-1) at pH 9.15 for endogenous and recombinant cruzipain, respectively). The pH-induced inactivation, which is a highly pH dependent first order process, is irreversible and accompanied by significant changes of secondary and tertiary structure as revealed by circular dichroism measurements. The different stability of cruzipain as compared to related proteases, is therefore due mainly to the different number, nature and distribution of charged residues within the catalytic domain and not due to addition of the C-terminal domain. PMID- 10708752 TI - Molecular cloning of fresh water and deep-sea rod opsin genes from Japanese eel Anguilla japonica and expressional analyses during sexual maturation. AB - We have determined the complete cDNA sequences of fresh water rod opsin gene (fwo) and deep-sea rod opsin gene (dso) from Japanese eel Anguilla japonica. The cDNA clones of fwo and dso consisted of 1437 and 1497 nucleotides, respectively. The predicted opsins of both genes consisted of 352 amino acid residues. Southern blot and PCR analyses of genomic DNA indicated that the Japanese eel genome contains only one fwo and one dso and they are intronless. Quantitative RT-PCR analyses revealed that the expression of fwo decreases with sexual maturation while that of dso increases. PMID- 10708751 TI - cDNA cloning of turtle prion protein. AB - Cloning of the cDNA coding for the 270-residue turtle prion protein is reported. It represents the most remote example thus far described. The entire coding region is comprised in a single exon, while a large intron interrupts the 5' UTR. The common structural features of the known prion proteins are all conserved in turtle PrP, whose identity degree to mammalian and avian proteins is about 40 and 58%, respectively. The most intriguing feature, unique to the turtle prion, is the presence of an EF-hand Ca(2+) binding motif in the C-terminal half of the protein. PMID- 10708754 TI - The effect of glutathione on the ATPase activity of MRP1 in its natural membranes. AB - The transport mechanism by which the multidrug resistance protein 1 (MRP1) effluxes cytotoxic agents out of cells is still not completely understood. However, the cellular antioxidant glutathione (GSH) has been shown to have an important role in MRP1-mediated drug transport. In this study we show that GSH stimulates the ATPase activity of MRP1 in a natural plasma membrane environment. This stimulation was dose-dependent up to 5 mM. The MRP1 substrates vincristine and daunorubicin do not induce MRP1 ATPase activity. In addition, the effect of GSH on the MRP1 ATPase activity is not increased by daunorubicin or by vincristine. In contrast, a GSH conjugate of daunorubicin (WP811) does induce the ATPase activity of MRP1. In the presence of GSH the effect of WP811 was not significantly increased. Finally, (iso)flavonoid-induced MRP1 ATPase activity is not synergistically increased by the presence of GSH. In conclusion, we show that GSH has no apparent influence on the ATPase reaction induced by several MRP1 substrates and/or modulators. The subclasses of molecules had different effects on the MRP1 ATPase activity, which supports the existence of different drug binding sites. PMID- 10708755 TI - CD63 associates with CD11/CD18 in large detergent-resistant complexes after translocation to the cell surface in human neutrophils. AB - CD63 antibody binding to the neutrophil surface triggers a transient activation signal that regulates the adhesive activity and surface expression of CD11/CD18. Gel permeation chromatography demonstrated that all of the cell surface CD11/CD18 associated with CD63 eluted in the void volume, indicating that they were present in large detergent-resistant complexes. In contrast, the majority of the total cellular CD63, CD11 and CD18, which are largely intracellular, was not present in complexes. The data suggest that intracellular CD11, CD18 and CD63 are not in detergent-resistant complexes, but enter such complexes following translocation to the cell surface. PMID- 10708753 TI - A lyso-platelet activating factor phospholipase C, originally suggested to be a neutral-sphingomyelinase, is located in the endoplasmic reticulum. AB - Recently a putative mammalian neutral-sphingomyelinase was cloned [Tomiuk et al. (1998) Proc. Natl. Acad. Sci. USA 95, 3638-3643; GenBank accession number AJ222801]. We have overexpressed this enzyme in cultured cells and demonstrate, using four different tagged constructs, that it is localized at the endoplasmic reticulum and not at the plasma membrane. This localization precludes a role for enzyme AJ222801 in the sphingomyelin cycle. Furthermore, a recent publication demonstrated that this enzyme has lyso-platelet activating factor (PAF) phospholipase C activity [Sawai et al. (1999) J. Biol. Chem. 274, 38131-38139]. Together, these data suggest a role for enzyme AJ222801 in the regulation of PAF metabolism. PMID- 10708756 TI - Properties of an insulin receptor with an IGF-1 receptor loop exchange in the cysteine-rich region. AB - The insulin receptor (IR) and the insulin-like growth factor-I receptor (IGF-1R) show differential binding of insulin and IGFs. The specificity determinants for IGF-1 binding are known to be located in the cysteine-rich (Cys-rich) region between residues 223 and 274 of human IGF-1R, which includes a loop that protrudes into the putative ligand binding site. In this report we have replaced residues 260-277 of human IR with residues 253-266 of the human IGF-1R to produce an IR-based, cysteine loop exchange chimaera, termed hIR-Cys loop exchange (CLX), in which all 14 amino acid residues in the exchanged loop differ from wild-type insulin receptor. This loop exchange had a detrimental effect on the efficiency of pro-receptor processing and on the binding of the mouse monoclonal antibody 83 7. However, this antibody, which binds hIR but not hIGF-1R, was still capable of immunoprecipitating the mature chimaeric receptor, indicating that the conformational epitope recognised by this antibody is not primarily determined by the loop region exchanged. The loop exchange did not significantly affect the ability of insulin to displace bound radiolabelled insulin, but increased the capacity of IGF-1 to competitively displace labelled insulin by at least 10 fold. PMID- 10708757 TI - A chloroplast envelope-transfer sequence functions as an export signal in Escherichia coli. AB - The small subunit precursor of pea ribulose-1,5-bisphosphate carboxylase/oxygenase engineered with prokaryotic elements was expressed in Escherichia coli. This resulted in a dependable level of synthesis of the precursor protein in E. coli. The bacterially synthesised plant precursor protein was translocated from the cytoplasm and targeted to the outer membrane of the envelope zone. During the translocation step, a significant proportion of the precursor was processed to a soluble, mature SSU and found localised in the periplasm. The determined amino acid sequence of the isolated precursor showed that it had a deletion of an arginine residue at position -15 in the transit peptide. Expression of this transit peptide-appended mammalian cytochrome b(5) in E. coli displayed a targeting profile of the chromogenic chimera that was similar to that observed with the plant precursor protein. PMID- 10708758 TI - Phenotype-dependent expression of cadherin 6B in vascular and visceral smooth muscle cells. AB - We used mRNA subtraction of differentiated and dedifferentiated smooth muscle cells (SMCs) to reveal the molecular mechanisms underlying the phenotypic modulation of SMCs. With this approach, we found that a 10 kb mRNA encoding a homotypic cell adhesion molecule, cadherin 6B, was strongly expressed in differentiated vascular and visceral SMCs, but not in the dedifferentiated SMCs derived from them. In vivo, cadherin 6B was expressed in vascular and visceral SMCs, in addition to brain, spinal cord, retina and kidney, at a late stage of chicken embryonic development. These results suggest that cadherin 6B is a novel molecular marker for vascular and visceral SMC phenotypes and is involved in the late differentiation of SMCs. PMID- 10708759 TI - Direct interaction of nerve growth factor receptor, TrkA, with non-receptor tyrosine kinase, c-Abl, through the activation loop. AB - The nerve growth factor receptor, TrkA, is essential for the survival and differentiation of neurons in the central and peripheral nervous systems. To understand the molecular principles underlying this differentiation step, we employed a yeast two-hybrid screening protocol using human TrkA as bait. We isolated c-Abl as a TrkA-interacting protein, in addition to known proteins such as phospholipase Cgamma and SH2-B. This interaction was confirmed by an in vitro binding assay using glutathione S-tranferase-Abl fusion protein. Furthermore, we show here that c-Abl binds to phosphotyrosine residue(s) in the kinase activation loop of TrkA. PMID- 10708760 TI - Characterisation of receptor-specific TNFalpha functions in adipocyte cell lines lacking type 1 and 2 TNF receptors. AB - Tumour necrosis factor-alpha (TNFalpha) is a multifunctional cytokine that exerts a myriad of biological actions in numerous different tissues including adipocytes through its two distinct cell surface receptors. To address the role of each TNF receptor in the biological actions of TNFalpha in adipocytes, we have developed four new preadipocyte cell lines. These were established from wild type controls (TNFR1(+/+)R2(+/+)) and from mice lacking TNFR1 (TNFR1(-/-)), TNFR2 (TNFR2(-/-)) or both (TNFR1(-/-)R2(-/-)). All four new cell lines can fully differentiate to form mature adipocytes, under appropriate culture conditions, as judged by cell morphology, expression of multiple adipogenic markers and the ability to mediate agonist-stimulated lipolysis and insulin-stimulated glucose transport. In wild type (TNFR1(+/+)R2(+/+)) and TNFR2(-/-) adipocytes, TNFalpha stimulated lipolysis and inhibited insulin-stimulated glucose transport as well as insulin receptor autophosphorylation. In contrast, these activities were completely lost in the TNFR1(-/-)R2(-/-) and TNFR1(-/-) cells. Taken together, these studies demonstrate that TNFalpha-induced lipolysis, as well as inhibition of insulin-stimulated glucose transport are predominantly mediated by TNFR1 and that the presence of TNFR2 is not necessary for these functions. This new experimental system promises to be useful in dissecting the molecular pathways activated by each TNF receptor in mediating the biological functions of TNFalpha in differentiated adipocytes. PMID- 10708761 TI - Requirement of autocrine signaling by bone morphogenetic protein-4 for chondrogenic differentiation of ATDC5 cells. AB - Mouse EC cell line ATDC5 undergoes differentiation to form cartilage nodules via the cellular condensation stage in the presence of insulin. ATDC5 cells expressed transcripts for bone morphogenetic protein-4 (BMP-4), and type IA and type II BMP receptors. Moreover, cells retained responsiveness to BMP-4, which induced the formation of chondrocytes in the culture. When transfected with a kinase domain truncated type IA BMP receptor construct, cells failed to undergo differentiation beyond the condensation stage even in the presence of insulin. The soluble form of type IA BMP receptor also blocked the formation of chondrocytes in a dose dependent manner. These lines of evidence suggested that autocrine BMP-4 signaling is required for the conversion of chondrogenic precursor cells into chondrocytes. PMID- 10708762 TI - Identification of a novel protein complex containing annexin VI, Fyn, Pyk2, and the p120(GAP) C2 domain. AB - p120(GAP) (RasGAP) has been proposed to function as both an inhibitor and effector of Ras. Previously we have shown that RasGAP contains a C2 domain which mediates both Ca(2+)-dependent membrane association and protein-protein interactions. Specifically, three proteins have been isolated in a complex with the C2 domain of RasGAP; these are the Ca(2+)-dependent lipid binding protein annexin VI (p70) and two previously unidentified proteins, p55 and p120. Here we provide evidence that p55 is the Src family kinase Fyn and p120 is the focal adhesion kinase family member Pyk2. In addition, in vitro binding assays indicate that Fyn, but not Pyk2 binds directly to annexin VI. Finally, co immunoprecipitation studies in Rat-1 fibroblasts confirm that Fyn, Pyk2, annexin VI and RasGAP can form a protein complex in mammalian cells. PMID- 10708763 TI - MOMP (major outer membrane protein) of Campylobacter jejuni; a versatile pore forming protein. AB - The great majority of trimeric porins of Gram-negative bacteria cannot be dissociated into monomers without disrupting their folded conformation. The porin of Campylobacter jejuni, however, displays two folded structures, a classical oligomer and a monomer resistant to detergent denaturation. We probed the transition of trimer to monomer using light scattering experiments and examined the secondary structures of these two molecular states by infra-red spectroscopy. The channel-forming properties of both trimer and monomer were studied after incorporation into artificial lipid bilayers. In these conditions, the trimer induced ion channels with a conductance value of 1200 pS in 1 M NaCl. The pores showed marked cationic selectivity and sensitivity to low voltage. Analysis of the isolated monomer showed nearly the same single-channel conductance and the same selectivity and sensitivity to voltage. These results indicate that the folded monomer form of C. jejuni MOMP displays essentially the same pore-forming properties as the native trimer. PMID- 10708764 TI - Voltage and substrate dependence of the inverse transport mode of the rabbit Na(+)/glucose cotransporter (SGLT1). AB - Properties of the cytoplasmic binding sites of the rabbit Na(+)/glucose cotransporter, SGLT1, expressed in Xenopus oocytes were investigated using the giant excised patch clamp technique. Voltage and substrate dependence of the outward cotransport were studied using alpha-methyl D-glucopyranoside (alphaMDG) as a substrate. The apparent affinity for alphaMDG depends on the cytoplasmic Na(+) concentration and voltage. At 0 mV the K(M) for alphaMDG is 7 mM at 110 mM Na(+) and 31 mM at 10 mM Na(+). The apparent affinity for alphaMDG and Na(+) is voltage dependent and increases at positive potentials. At 0 mV holding potential the outward current is half-maximal at about 70 mM. The results show that SGLT1 can mediate sugar transport out of the cell under appropriate concentration and voltage conditions, but under physiological conditions this transport is highly improbable due to the low affinity for sugar. PMID- 10708765 TI - One-codon alternative splicing of the CpG MTase Dnmt1 transcript in mouse somatic cells. AB - The genomic methylation patterns in the mammalian somatic cells are presumably maintained by a single enzyme, dnmt1. In mouse, this DNA (cytosine-5) methyltransferase, or CpG MTase, is encoded by the Dnmt1 gene. We now present evidence that in different tissues and cell types, the primary transcript of mouse dnmt1 is alternatively spliced to generate two poly-(A) RNAs of approximately similar abundance. This alternative splicing most likely originates from the existence of two tandemly arranged acceptor sites separated by only 3 nt. The two Dnmt1 mRNAs thus encode two CpG MTases differing by two amino acids. We discuss the implications of the discovery of two dnmt1 isozymes, instead of one enzyme as previously thought, in the somatic cells of both mouse and human. PMID- 10708766 TI - Trimeric ring-like structure of ArsA ATPase. AB - ArsA protein is the soluble subunit of the Ars anion pump in the Escherichia coli membrane which extrudes arsenite or antimonite from the cytoplasm. The molecular weight of the subunit is 63 kDa. In the cell it hydrolyzes ATP, and the energy released is used by the membrane-bound subunit ArsB to transport the substrates across the membrane. We have obtained two-dimensional crystals of ArsA in the presence of arsenite on negatively-charged lipid monolayer composed of DMPS and DOPC. These crystals have been studied using electron microscopy of negatively stained specimens followed by image processing. The projection map obtained at 2.4 nm resolution reveals a ring-like structure with threefold symmetry. Many molecular assemblies with the same ring-shape and dimensions were also seen dispersed on electron microscopy grids, prepared directly from purified ArsA protein solution. Size-exclusion chromatography of the protein sample with arsenite present revealed that the majority of the protein particles in solution have a molecular weight of about 180 kDa. Based on these experiments, we conclude that in solution the ArsA ATPase with substrate bound is mainly in a trimeric form. PMID- 10708768 TI - Retinoic acid stimulates HIV-1 transcription in human neuroblastoma SH-SY5Y cells. AB - Although the brain is an important target for the human immunodeficiency virus type 1 (HIV) and viral infection causes neuronal degeneration and dementia, the mechanisms responsible for HIV transcription in neuronal cells are largely unknown. We show here that retinoic acid (RA) stimulates HIV transcription in human neuronal SH-SY5Y cells. The steroid receptor coactivator 1 (SRC-1) enhances the transcriptional response to RA, and the viral protein Tat cooperates with RA and SRC-1 to induce a strong transactivation. These results suggest that retinoid receptors could play an important role as activators of viral gene expression in the human brain. PMID- 10708767 TI - Inactivation of glycogen synthase kinase-3 by protein kinase C delta: implications for regulation of tau phosphorylation. AB - The role of the phosphatidylinositol 3-kinase (PI3K) pathway in the hyperphosphorylation of tau was investigated in SY5Y human neuroblastoma cells. Wortmannin, an inhibitor of PI3K, induced transient (after 1 h) activation of glycogen synthase kinase-3 (GSK-3), hyperphosphorylation of tau and dose dependent cytotoxicity. However, continuous inactivation of protein kinase (PK) B was observed from 1 to 24 h, suggesting the involvement of protein kinase(s) other than PKB in the phosphorylation and inactivation of GSK-3 after 3 h. In cells treated with wortmannin, PKC delta fragments were observed, and the PKC activity increased after 3 h, whereas treatment of cells with z-DEVD-fmk, an inhibitor of caspase 3, also inhibited fragmentation of PKC delta and induced continuous activation of GSK-3. It is suggested that fragmentation of PKC delta during the process of apoptosis results in the phosphorylation and inactivation of GSK-3 and consequently inhibition of the phosphorylation of tau. PMID- 10708769 TI - The novel MMS-inducible gene Mif1/KIAA0025 is a target of the unfolded protein response pathway. AB - In a search for genes induced by DNA-damaging agents, we identified two genes that are activated by methyl methanesulfonate (MMS). Expression of both genes is regulated after endoplasmic reticulum (ER) stress via the unfolded protein response (UPR) pathway. The first gene of those identified is the molecular chaperone BiP/GRP78. The second gene, Mif1, is identical to the anonymous cDNA KIAA0025. Treatment with the glycosylation inhibitor tunicamycin both enhances the synthesis of Mif1 mRNA and protein. The Mif1 5' flanking region contains a functional ER stress-responsive element which is sufficient for induction by tunicamycin. MMS, on the other hand, activates Mif1 via an UPR-independent pathway. The gene encodes a 52 kDa protein with homology to the human DNA repair protein HHR23A and contains an ubiquitin-like domain. Overexpressed Mif1 protein is localized in the ER. PMID- 10708770 TI - Triple helix assembly and processing of human collagen produced in transgenic tobacco plants. AB - The use of tobacco plants as a novel expression system for the production of human homotrimeric collagen I is presented in this report. Constructs were engineered from cDNA encoding the human proalpha1(I) chain to generate transgenic tobacco plants expressing collagen I. The recombinant proalpha1(I) chains were expressed as disulfide-bonded trimers and were shown to fold into a stable homotrimeric triple helix. Moreover, the recombinant procollagen was subsequently processed to collagen as it occurs in animals. Large amounts of recombinant collagen were purified from field grown plant material. The data suggest that plants are a valuable alternative for the recombinant production of collagen for various medical and scientific purposes. PMID- 10708771 TI - The influence of foot positioning on ankle sprains. AB - The goal of this study was to examine the influence of changes in foot positioning at touch-down on ankle sprain occurrence. Muscle model driven computer simulations of 10 subjects performing the landing phase of a side shuffle movement were performed. The relative subtalar joint and talocural joint angles at touchdown were varied, and each subject-specific simulation was exposed to a set of perturbed floor conditions. The touchdown subtalar joint angle was not found to have a considerable influence on sprain occurrence, while increased touchdown plantar flexion caused increased ankle sprain occurrences. Increased touchdown plantar flexion may be the mechanism which causes ankles with a history of ankle sprains to have an increased susceptibility to subsequent sprains. This finding may also reveal a mechanism by which taping of a sprained ankle or the application of an ankle brace leads to decreased ankle sprain susceptibility. PMID- 10708772 TI - A three-dimensional mechanical analysis of a stentless fibre-reinforced aortic valve prosthesis. AB - Failure of bioprosthetic and synthetic three-leaflet valves has been shown to occur as a consequence of high tensile and bending stresses, acting on the leaflets during opening and closing. Moreover, in the stented prostheses, whether synthetic or biological, the absence of contraction of the aortic base, due to the rigid stent, causes the leaflets to be subjected to an unphysiological degree of flexure, which is related to calcification. It is shown that the absence of the stent, which gives a flexible aortic base and leaflet attachment, and leaflet fibre-reinforcement result in reduced stresses in the weaker parts of the leaflets in their closed configuration. It is postulated that this leads to a decrease of tears and perforations, which may result in a improved long-term behaviour. The effect of a flexible leaflet attachment and aortic base of a synthetic valve is investigated with a finite element model. Different fibre reinforced structures are analysed with respect to the stresses that are likely to contribute to the failure of fibre-reinforced prostheses and compared with the results obtained for a stented prosthesis. Results show that for the stentless models a reduction of stresses up to 75% is obtained with respect to stented models with the same type of reinforcement. PMID- 10708773 TI - The history dependence of force production in mammalian skeletal muscle following stretch-shortening and shortening-stretch cycles. AB - The purpose of this study was to determine the history dependence of force production during and following stretch-shortening and shortening-stretch cycles in mammalian skeletal muscle. Thirty-three different isometric, stretch, shortening, stretch-shortening and shortening-stretch experiments were preformed in cat soleus (n=8) using previously established methods. Stretch-shortening and shortening-stretch cycles are not commutative with respect to the isometric forces following the length changes. Whereas force depression following shortening is virtually unaffected by previous stretching of the muscle, force enhancement following stretch depends in a dose-dependent manner on the amount of muscle shortening preceding the stretch. The history dependence of isometric force following shortening-stretch cycles can conveniently be modelled using an elastic (compressive and tensile) element that engages at the length of muscle activation. Such an "elastic" mechanism has been proposed by Edman and Tsuchiya (1996) (Edman, K.A. P., Tsuchiya, T., 1996. Strain of passive elements during force enhancement by stretch in frog mucle fibres. Journal of Physiology 490. 1, 191-205) based on experimental observations, and has been implemented theoretically in a rheological model of muscle (Forcinito et al., 1997) (Forcinito, M., Epstein, M., Herzog, W., 1997. Theoretical considerations on myofibril stiffness. Biophysics Journal 72, 1278-1286). The history dependence of isometric force following stretch-shortening cycles appears independent of the stretch preceding the shortening, except perhaps, if stretching occurs at very high speeds (i.e. 6-10 times fibre length per second). The results of this study are hard to reconcile with the two major mechanisms associated with history dependence of force production: sarcomere length non-uniformity (Edman et al., 1993) and stress-induced cross-bridge inhibition (Marechal and Plaghki, 1979) (Marechal, G., Plaghki, L., 1979. The deficit of the isometric tetanic tension redeveloped after a relase of frog muscle at a constant velocity. Journal of General Physiology 73, 453-467). It appears that studying the history dependence of force production under more functionally relevant conditions than has been done to date may provide new information that contributes to our understanding of possible mechanisms associated with force depression and force enhancement following muscular length changes. PMID- 10708774 TI - A critical parameter for transcapillary exchange of small solutes in countercurrent systems. AB - Small solute transport by a countercurrent capillary loop was studied using a theoretical model. In the model, the afferent and the efferent limbs of the loop share a common interstitial space, with which exchange of solute occurs. Sources of solute, epithelial cells, exist near capillaries and secret solute into the interstitial fluid. Parameters based on experimental measurements on young Sprague-Dawley rats were used in the model, and asymptotic solutions were derived. Comparison of the solute distribution in the interstitium between a capillary loop and a single capillary reveals that the ratio of the product of permeability (P(1)) and surface area (A(1)) to flow (F(1)) of the afferent limb, gamma(1)=P(1)A(1)/F(1) is a critical parameter for the countercurrent exchange system. It alone determines whether the countercurrent arrangement of capillaries facilitates clearance of solute from the interstitial fluid, a greater axial gradient of solute in the interstitium from the base to the tip of the capillary loop and a greater effect of flow, F, upon this gradient. The properties of the efferent limb affect the results, but it is gamma(1) that determines the characteristic difference between a capillary loop and a single capillary. PMID- 10708775 TI - Computational model of the fluid dynamics in systemic-to-pulmonary shunts. AB - A systemic-to-pulmonary shunt is a connection created between the systemic and pulmonary arterial circulations in order to improve pulmonary perfusion in children with congenital heart diseases. Knowledge of the relationship between pressure and flow in this new, surgically created, cardiovascular district may be helpful in the clinical management of these patients, whose survival is critically dependent on the blood flow distribution between the pulmonary and systemic circulations. In this study a group of three-dimensional computational models of the shunt have been investigated under steady-state and pulsatile conditions by means of a finite element analysis. The model is used to quantify the effects of shunt diameter (D), curvature, angle, and pulsatility on the pressure-flow (DeltaP-Q) relationship of the shunt. Size of the shunt is the main regulator of pressure-flow relationship. Innominate arterial diameter and angles of insertion have less influence. Curvature of the shunt results in lower pressure drops. Inertial effects can be neglected. The following simplified formulae are derived: DeltaP=(0. 097Q+0.521Q(2))/D(4) and DeltaP=(0.096Q+0.393Q(2))/D(4) for the different shunt geometries investigated (straight and curved shunts, respectively). PMID- 10708776 TI - The effects of stress enhancement on the extracellular matrix and fibroblasts in the patellar tendon. AB - The purpose of the present study is to determine the effect of the stress enhancement and intrinsic fibroblasts on the extracellular matrix of the patellar tendon. Thirty-two female Japanese White rabbits were divided into four groups. In Group 1, the patellar tendon underwent the in situ freeze-thaw treatment to kill intrinsic fibroblasts of the patellar tendon and the patellar tendon underwent the wrapping treatment with nylon membrane filters to inhibit extrinsic cell infiltration. In Group 2, the medial and the lateral portions of the frozen thawed patellar tendon were resected to enhance the stress, and then the central two-thirds of the patellar tendon underwent the wrapping treatment. In Group 3, the patellar tendon without the freeze/thaw treatment underwent the wrapping treatment. In Group 4, the patellar tendon was narrowed and wrapped in the same manner. All rabbits were killed 6 weeks after surgery. While the elastic modulus and the tensile strength of the patellar tendon in Group 2 were significantly less than those in Group 1, we could not find any significant differences in these parameters between Groups 3 and 4. Histologically, while no fibroblasts were observed in Groups 1 and 2, fibroblasts were found in Groups 3 and 4. This study revealed that stress enhancement decreases the elastic modulus and the tensile strength of the extracellular matrix of the patellar tendon and that intrinsic fibroblasts prevent the detrimental effect of stress enhancement on mechanical properties of the patellar tendon. PMID- 10708777 TI - Time-frequency analysis and filtering of kinematic signals with impacts using the Wigner function: accurate estimation of the second derivative. AB - Biomechanical signals are represented in the time-frequency domain using the Wigner distribution function. Filtering of this representation for the case of a non-stationary displacement signal with impact is studied. Smoothed displacement data are then double differentiated and compared with references accelerometer data. It is shown that this technique is able to remove noise from these signals in a better way than conventional filtering techniques currently used in biomechanics. PMID- 10708778 TI - Effect of removing the nucleus pulposus on the deformation of the annulus fibrosus during compression of the intervertebral disc. AB - Eighteen frozen ovine discs were bisected, in the mid-sagittal plane, to produce 36 specimens. The cut surfaces were marked at the inner and outer annulus boundaries of the annulus fibrosus, both anteriorly and posteriorly, with Alcian blue stain. The sections were sealed by a transparent plate, and thawed. A compression of 1mm at a rate of 0.2mms(-1) was applied. The displacements of the Alcian blue marks were measured from the video images, recorded during the tests, using interactive image analysis software. Before removal of the nucleus, the inner boundaries of the annulus moved outwards during compression (P<0.001, anterior; P=0.01, posterior). However, after removal of the nucleus, both inner boundaries moved inwards (P<0.001, anterior and posterior). The outer boundaries moved outwards both before and after removal of the nucleus (P<0.001). The results showed that total removal of the nucleus changes the response of the annulus to compression. PMID- 10708779 TI - Middle-ear dynamics before and after ossicular replacement. AB - The mechanism of hearing involves conduction of mechanical vibrations along the ossicular chain to the inner ear. An acoustic wave is collected and transformed as it passes down the ear canal and impacts on the tympanic membrane (ear drum). The drum is connected to the inner-ear by three ossicle bones (malleus, incus, and stapes) in a complex arrangement, which serves to further transform the mechanical vibration before it reaches the cochlea of the inner ear. What is the mechanical function of the ossicular chain, and what are the biomechanical consequences of surgical reconstruction with prostheses? To answer these questions, a three-dimensional finite element model of the outer ear canal and middle ear was generated. The dynamical behaviour was predicted for the normal ear, and an ear reconstructed with partial and total ossicular replacement prostheses. For the normal ear, stapes amplitudes of 1x10(-8) m at low frequencies decrease to 4x10(-10)m at approximately 3kHz with several resonance peeks in between, most significantly at approximately 1kHz. Thereafter a further resonance is predicted at 4kHz associated with the ear canal. The behaviour is changed fundamentally by adding a prosthesis; the partial replacement increases the vibratory coupling of the drum and the stapes compared to the normal ear whereas the total replacement does the opposite, and is predicted to have the disadvantage of bringing several new resonances of the ossicular chain into the hearing range. It is hypothesised that the function of the malleus-incus-stapes arrangement is to link the drum to the oval window with the flexibility required for impedance matching but the rigidity to prevent unconstrainable resonances from occurring in the hearing range. If this is true, then the structural stiffness of ossicular chain is the critical design variable for middle-ear replacement prostheses. PMID- 10708780 TI - Elbow and wrist joint contact forces during occupational pick and place activities. AB - A three-dimensional, mathematical model of the elbow and wrist joints, including 15 muscle units, 3 ligaments and 4 joint forces, has been developed. A new strain gauge transducer has been developed to measure functional grip forces. The device measures radial forces divided into six components and forces of up to 250N per segment can be measured with an accuracy of +/-1%. Ten normal volunteers were asked to complete four tasks representing occupational activities, during which time their grip force was monitored. Together with kinematic information from the six-camera Vicon data, the moment effect of these loads at the joints was calculated. These external moments are assumed to be balanced by the internal moments, generated by the muscles, passive soft tissue and bone contact. The effectiveness of the body's internal structures in generating joint moments was assessed by studying the geometry of a simplified model of the structures, where information about the lines of action and moment arms of muscles, tendons and ligaments is contained. The assumption of equilibrium between these external and internal joint moments allows formulation of a set of equations from which muscle and joint forces can be calculated. A two stage, linear optimisation routine minimising the overall muscle stress and the sum of the joint forces has been used to overcome the force-sharing problem. Humero-ulnar forces of up to 1600N, humero-radial forces of up to 800N and wrist joint forces of up to 2800N were found for moderate level activity. The model was validated by comparison with other studies. PMID- 10708781 TI - Effect of resistance load on biomechanical characteristics of racing wheelchair propulsion over a roller system. AB - The purpose of this study was to examine how resistance load influenced the kinematic characteristics and the activity of selected muscles (flexor and extensor carpi radialis, biceps brachii, triceps brachii, antero-middle and postero-middle deltoids, pectoralis major, and upper trapezius) during maximum effort racing wheelchair stroking using 3D videographic and surface EMG techniques. Fifteen male experienced wheelchair racers served as subjects and three consecutive stroke cycles were analyzed for two load conditions. In contrast to previous studies where variations in speed were a result of variations in pushing effort or disability classification, a reduction in stroking speed caused by increasing load did not result in a decrease in stroking frequency. Increases in load significantly influenced the push and recovery times but not the stroke time or frequency. The vertical ranges of motion and vertical velocities at initial hand contact of the upper extremity joints decreased significantly from light to heavy resistance conditions. These results suggest that the vertical motion is influenced greatly by the load. Various degrees of muscle co-contractions were observed in most phases of the stroke cycle. The activation pattern of the deltoid muscle was different from what has been previously reported, probably because of the exaggerated forward lean trunk position adopted by our subjects. Although the overall EMG activity remained the same or decreased when the resistance was increased, stroking under a heavy resistance load is likely to be more demanding physiologically because of the greater push time-recovery time (work-rest) ratio with increasing resistance. PMID- 10708782 TI - Glenohumeral translation during active and passive elevation of the shoulder - a 3D open-MRI study. AB - Despite its importance for the understanding of joint mechanics in healthy subjects and patients, there has been no three-dimensional (3D) in vivo data on the translation of the humeral head relative to the glenoid during abduction under controlled mechanical loading. The objective was therefore to analyze humeral head translation during passive and active elevation by applying an open MR technique and 3D digital postprocessing methods. Fifteen healthy volunteers were examined with an open MR system at different abduction positions under muscular relaxation (30-150 degrees of abduction) and during activity of shoulder muscles (60-120 degrees ). After segmentation and 3D reconstruction, the center of mass of the glenoid and the midpoint of the humeral head were determined and their relative position calculated. During passive elevation, the humeral head translated inferiorly from +1.58mm at 30 degrees to +0. 36mm at 150 degrees of abduction, and posteriorly from +1.55mm at 30 degrees to -0.07mm at 150 degrees of abduction. Muscular activity brought about significant changes in glenohumeral translation, the humeral head being in a more inferior position and more centered, particularly at 90 and 120 degrees of abduction (p<0.01). In anterior/posterior direction the humeral head was more centered at 60 and 90 degrees of abduction during muscle activity. The data demonstrate the importance of neuromuscular control in providing joint stability. The technique developed can also be used for investigating the effect of muscle dysfunction and their relevance on the mechanics of the shoulder joint. PMID- 10708783 TI - Trabeculated embryonic myocardium shows rapid stress relaxation and non-quasi linear viscoelastic behavior. AB - Passive viscoelastic behavior is important in embryonic cardiovascular function, influencing the rate and magnitude of contraction and relaxation. We hypothesized that if viscoelastic behavior is influenced by interstitial fluid flow, then the stage-21 (312d) and stage-24 (4d) chick myocardium with large intertrabecular spaces will exhibit much different viscoelastic behavior than stage-16 (212d) and stage-18 (3d) compact myocardium and a non-quasi-linear response. Excised left ventricular sections were tested with ramp-and-hold stress relaxation tests at axial stretch ratios of 1.05:1.1:1.2:1.3. The measured stress relaxation was much more rapid than previously observed in the compact, non-trabeculated myocardium. The reduced relaxation curves depended significantly on the stretch level. A continuous-spectrum quasi-linear relaxation function described their shape well but the model-fit parameters also depended on the stretch level. Sinusoidal stretching of ventricular sections at rates from 0.2 to 25Hz showed that the steepening of stress-strain curves with increasing strain rate was half as much as predicted by a quasi-linear model. Hysteresis ranged from 25-35%, varied little with loading rate from 0.2 to 8Hz, and was twice that predicted from a quasi-linear model. Doubling the viscosity of the perfusate in stress-relaxation tests produced increased stiffness and decreased relaxation rate. These results demonstrate that the passive viscoelastic behavior of the trabeculated embryonic myocardium is markedly different from that of younger, compact myocardium and is not quasi-linear. PMID- 10708784 TI - Simulated pathline visualization of computed periodic blood flow patterns. AB - Improvements in computer hardware and software have made it possible to model pulsatile blood flow in realistic arterial geometries. Such studies produce enormous amounts of velocity data, which are often difficult to interpret and communicate using traditional contour and/or vector field plots. Inspired by in vitro flow visualization techniques such as particle image velocimetry (PIV), we describe a simple and effective method for visualizing periodic three-dimensional velocity data, based on the subdivision and sequential display of computed particle trajectories. Analogous to a PIV experiment, the length and spacing of such simulated particle pathlines are controlled by user-specified shutter-speed and frame rate variables. Strategies for color-coding pathlines to highlight important hemodynamic features such as recirculation zones and branch flow division are presented. PMID- 10708785 TI - A new method for the representation of articular surfaces using the influence surface theory of plates. AB - The traditional approach to the representation of an articular surface is by using piecewise polynomial functions with a limited continuity to fit the surface from ordered data points. In this study, we introduce a new method, which is based on the influence surface theory of plates, for the representation of articular surfaces. The most significant advantage of this method is that it can effectively represent an articular surface from non-ordered data points. The effectiveness of the present method was shown by reconstruction of a human femoral surface and a mathematical cone. PMID- 10708787 TI - How do changes to plate thickness, length, and face-connectivity affect femoral cancellous bone's density and surface area? An investigation using regular cellular models PMID- 10708786 TI - Simulation of elbow and forearm motion in vitro using a load controlled testing apparatus. AB - The purpose of this study was to compare passive to active testing on the kinematics of the elbow and forearm using a load-controlled testing apparatus that simulates muscle loading. Ten fresh-frozen upper extremities were tested. Active control was achieved by employing computer-controlled pneumatic actuators attached to the tendons of the brachialis, biceps, triceps, brachioradialis and pronator teres. Motion of the radius and ulna relative to the humerus was measured with an electromagnetic tracking system. Active elbow flexion produced more repeatable motion of the radius and ulna than when tested passively (p<0.05). The decrease in variability, as determined from the standard deviation of five successive trials in each specimen, was 76.5 and 58.0% for the varus valgus and internal-external motions respectively (of the ulna relative to the humerus). The variability in flexion during simulated active forearm supination was 30.6% less than during passive testing. Thus under passive control, in the absence of stability provided by muscular loading across the joint, these uncontrolled motions produce increased variability amongst trials. The smooth and repeatable motions resulting from active control, that probably model more closely the physiologic state, appear to be beneficial in the evaluation of unconstrained kinematics of the intact elbow and forearm. PMID- 10708788 TI - Ultrastructure and affinity of Lower Carboniferous megaspores from the Moscow Basin, Russia. AB - Ten megaspore species isolated from Moscow Basin lignites of Lower Carboniferous (Visean) age have been studied by scanning and transmission electron microscopy (SEM and TEM). These species belong to seven megaspore genera: Lagenicula, Sublagenicula, Crassilagenicula, Setosisporites, Zonalesporites, Caudatosporites, and Cystosporites. Megaspores of the genus Caudatosporites have only been described previously from the Duckmantian (Westphalian B); a new species is duly erected. The ultrastructure of megaspore walls from the genera Crassilagenicula and Zonalesporites has not been previously described. This study also places them in context with other contemporaneous megaspores. The study shows that during the Visean, in the Moscow Basin, megaspores expressed a similar wall ultrastructure despite large differences in external appearance. The genus Crassilagenicula may represent a group of megaspores from plants that had evolved from those bearing gulate megaspores here typified by Lagenicula acuminata, Setosisporites brevispinosus, and Sublagenicula hirsutoida. Zonalesporites brasserti also appears to show affinities to this group, and may be representative of a plant species in a transitional state between the Lagenicula bearing lycopsids and those more isoetalean in nature. PMID- 10708789 TI - Diversity of exine structure in Upper Carboniferous (Westphalian) selaginellalean megaspores. AB - Studies of wall structure in Mesozoic and Recent selaginellalean megaspores have been well documented. However, Palaeozoic examples have received minimal attention. The principal Palaeozoic megaspore genus of likely selaginellalean affinity is Triangulatisporites, extending from the Upper Devonian to the Upper Carboniferous. The particulate wall ultrastructure of a previously published Carboniferous (Duckmantian) megaspore assigned to this genus suggested that this form of wall construction may have been the ancestral wall structure of the group, an observation which posed difficulties in relating selaginellalean ultrastructure to that of other contemporaneous lycopsid megaspores. Subsequent investigation showed that the genus also contains more laminate exines similar to those of other extinct lycopsids and extant Selaginella species. Our new examples of Triangulatisporites ultrastructure from the Langsettian, Duckmantian and Westphalian D yield more information regarding early variation of wall structure within Carboniferous selaginellalean megaspores and suggest that a more laminate wall composition is at least as old as the particulate form. However, without further investigation of Lower Carboniferous forms, we are unable to state which is indeed ancestral. The laminate structure reported here and elsewhere is, none the less, more easily related to comparable ultrastructure in other groups of Carboniferous lycopsid megaspores and could suggest a link with such genera as Zonalesporites and early Lagenicula. This would be in keeping with current concepts regarding the most primitive ultrastructural type within lycopsid megaspore walls. PMID- 10708790 TI - Paleoenvironmental history of the Popayan area since 27 000 yr BP at Timbio, Southern Colombia. AB - A pollen record from Timbio, located at an elevation of 1750m on the high plain of Popayan (2 degrees 24'N, 76 degrees 36'W) is presented. This forms the basis for reconstructing the vegetation and climate history for the periods from 27000 to 9200 radiocarbon years before the present (14Cyr BP) and 2100 14Cyr BP to sub recent. The 5m sediment core has time control based on seven AMS radiocarbon dates. Four pollen assemblage zones (TIM-1 to TIM-4) are recognized. During the period of 27200 to 26000 14Cyr BP, an Andean forest was near the site. The vegetation consisted of forest and open herb-rich vegetation, climatic conditions were moist and temperatures some 6 degrees C lower than compared to those of today. During the period of 26000 to 16000 14Cyr BP forest was less open. The observed succession from a Podocarpus-Weinmannia dominated forest to a Hedyosmum dominated forest, and finally to a forest with Ilex, Myrica and ferns indicates a progressive decrease of temperature during this period, with a maximum temperature depression of ca. 5-7.5 degrees C compared to present-day conditions. During the period of 16000 to 9200 14Cyr BP, temperature decrease is estimated at ca. 7.5 degrees C and the climate was the driest. During the period of 2100 to 600 14C2600m altitude (ca. 8 degrees C) and those at sea-level (2.5-6 degrees C) and supports the observation that glacial lapse rates were higher than in modern times. PMID- 10708791 TI - Variations in dinoflagellate cyst morphology under conditions of changing salinity during the last 2000 years in the Limfjord, Denmark. AB - Morphological variations are examined in the dinoflagellate cysts Spiniferites spp., Lingulodinium polyedrum and Protoceratium reticulatum (=Operculodinium centrocarpum) from a core taken in the Bjornsholm Bay, the Limfjord, Denmark. The fjord has a history of changing salinity, and unusual cyst morphotypes are found in the greatest numbers during periods of inferred low salinity. Variation occurs primarily in cyst process morphology, and the aberrant morphotypes have processes that are shorter, thicker and/or more membranous. The different morphotypes are described and compared with other varieties and forms of the three taxa and to other closely related taxa. PMID- 10708792 TI - Erratum to: "Palaeofloristic and palaeovegetational changes across the Paleocene/Eocene boundary in northern South America" PMID- 10708793 TI - Cloning and characterization of a cDNA sequence encoding the precursor of a chlorotoxin-like peptide from the Chinese scorpion Buthus martensii Karsch. AB - A full-length cDNA sequence encoding the precursor of a venom peptide with homology to chlorotoxin (named BmKCT) was isolated from a cDNA library made from the venom glands of the Chinese Scorpion Buthus martensii Karsch. The encoded precursor of BmKCT was 59 amino acid residues long including a signal peptide of 24 residues and a mature toxin of 35 residues with four disulfide bridges. The sequence of BmKCT is similar (68% identities) to that of chlorotoxin isolated from Leiurus quinguestriatus quinquestriatus. BmKCT is the first report of the cDNA sequences encoding four-disulfide-bridged short-chain toxins from Buthus martensuii Karsch so far. PMID- 10708794 TI - Portuguese Man-of-war (Physalia physalis) venom induces calcium influx into cells by permeabilizing plasma membranes. AB - Portuguese Man-of-war (Physalia physalis) nematocyst venom dose-dependently stimulates calcium (45Ca(2+)) influx into L-929, GH(4)C(1), FRL, and embryonic chick heart cells. Venom-induced calcium influx is not blocked by ouabain, vanadate, nor organic calcium channel blockers, but is blocked by transition metal cations, such as lanthanum and zinc. Venom-induced calcium influx is accompanied in a dose-dependent manner by the release of intracellular lactate dehydrogenase, indicating a loss in plasma membrane integrity and cytolysis. Concentrations of zinc that block 45Ca(2+) influx also block lactate dehydrogenase release. Lanthanum, which also blocks 45Ca(2+) uptake, does not neutralize the cytolytic activity of the venom, but rather inhibits the venom's cytolytic action at the level of the target cell plasma membrane. Our findings indicate that Man-of-war venom causes an influx of calcium into several different cells types, not just those of the cardiovascular system, and this influx likely occurs by permeabilizing the plasma membranes of cells. PMID- 10708795 TI - Habutobin recognizes Thr(7) in the sequence of fibrinopeptide A of rabbit fibrinogen. AB - Habutobin, a thrombin-like enzyme from Trimeresurus flavoviridis venom, cleaves only the Arg(16)-Gly(17) bond in the rabbit Aalpha chain and releases fibrinopeptide A (FPA). To investigate the role of amino acid residues in the rabbit FPA sequence upon habutobin action, we examined the inhibitory effects of FPA and peptides containing partial sequences of FPA on the habutobin action. Fibrinopeptides from rabbit, human, bovine and dog were isolated and rabbit FPA was fragmented using dilute HCl. Rabbit FPA inhibited the action of habutobin although FPA from human, bovine and dog did not. Among the fragments of rabbit FPA, a heptapeptide Aalpha 3-9, the N-terminal region of rabbit FPA, competitively inhibited the release of FPA by habutobin, whereas the C-terminal hexapeptide of FPA (Aalpha 11-16) exerted no effect on the habutobin action. Synthetic tripeptides Ser-Thr-Phe corresponding to Aalpha 6-8 and Ala-Thr-Phe also inhibited the habutobin action, but Ser-Asp-Phe and Ala-Thr-Gly did not. It is concluded that habutobin would recognize the region around Thr(7)-Phe(8) in the sequence of rabbit FPA (Aalpha 1-16) prior to the cleavage of the Arg(16) Gly(17) bond. PMID- 10708796 TI - Okadaic acid production from the marine benthic dinoflagellate Prorocentrum arenarium Faust (Dinophyceae) isolated from Europa Island coral reef ecosystem (SW Indian Ocean). AB - Okadaic acid was isolated from a strain of Prorocentrum arenarium Faust (Prorocentrales, Dinophyceae) collected from Europa Island (40 degrees 22'E, 22 degrees 20'S, SW Indian Ocean). The presence of okadaic acid in the algal extract was suspected after cytotoxicity and phosphatase 2A inhibition testing. It was confirmed by ADAM derivatization, immunoaffinity extraction and liquid chromatography with fluorimetric detection analysis as well as by liquid microchromatography with mass spectrometric detection. Results indicate that the P. arenarium strain was toxinogenic and could be potentially involved in the toxin production associated with the human diseases, diarrhetic shellfish poisoning and possibly ciguatera fish poisoning in the SW Indian Ocean area. PMID- 10708797 TI - Sensitivity of alpha-amanitin to oxidation by a lactoperoxidase-hydrogen peroxide system. AB - Oxidation of alpha-amanitin - a potent hepatotoxin found in the mushroom Amanita phalloides - by a lactoperoxidase-hydrogen peroxide system was investigated by different techniques. (i). UV spectroscopy of the mixture after 24 h incubation reveals a significant decrease in the absorbance range characteristic of the putative reactive moiety of the toxin, the tryptathionine group. (ii). Formation of a new product was detected by Thin Layer Chromatography. (iii). In vivo experiments with non-inbred male albino mice showed a lowered toxicity of the modified toxin in comparison with that of the native one. Taking into account the latter results concerning the sensitivity of the toxin towards an oxidising system, the formation and reactivity of an alpha-amanitin derivative is discussed in the course of A. phalloides poisoning (inhibition of RNA polymerase type II and development of damaging radical species). PMID- 10708798 TI - The multiplicity of cardiotoxins from Naja naja atra (Taiwan cobra) venom. AB - Four novel cardiotoxins were isolated from Naja naja atra (Taiwan cobra) venom by successive separation on a SP-Sephadex C-25 column and a reverse phase column. Amino acid sequences of the cardiotoxins were determined by Edman degradation and carboxypeptidase digestion. It shows that these cardiotoxins comprise 60 amino acid residues. Comparative analyses on the amino acid sequences of cardiotoxins from the venoms of N. naja atra and other Naja species indicated that amino acid substitutions of cardiotoxin isoforms frequently occurred at positions 7-11, 27 32 and 45-47. The hypervariable segments encoded by the second and third exon of cardiotoxin genes are located at or near the tips of loop structure of cardiotoxin molecules. These results, together with the suggestions that the residues at the tips of cardiotoxins' loop structure were involved in the manifestation of the biological activities of cardiotoxins, reflect that the preferential mutations may contribute to alterations in the function of cardiotoxin molecules. Analysis on the secondary structure of pre-mRNAs of N. naja atra cardiotoxin 4 gene and N. naja sputatrix cardiotoxin 3 gene has shown that the hypervariable regions of the exon 2 pertain to form intra-exon pairings and are not involved in the formation of intron-exon pairings. Since the pairings of splice sites and gene architecture were supposed to be associated with intron exon recognition, it is likely that the preferred loci of mutations occurring with the evolution of cardiotoxin genes would not affect the processing of cardiotoxin precursors. PMID- 10708799 TI - Melittin-mediated release of [3H]-oleic acid from E. coli cells is dependent upon heat- and trypsin-sensitive factor(s) in human serum. AB - Synthetic melittin mediated the release of [3H]-oleic acid ([3H]-OA) or its acylated lipids from [3H]-OA-labeled E. coli cells exposed to human serum. This phenomenon was not observed in the absence of serum and was calcium independent. The addition of serum was not required for melittin-mediated lysis of erythrocytes, although lysis was greater in the presence of serum than in its absence (P<0.001). Trypsin treatment of human serum reduced the melittin-mediated release of [3H]-OA/acylated lipids, and this effect was more pronounced upon boiling the serum (P<0.01). A kinetic study showed that maximum release of [3H] OA/acylated lipids occurred within 3-6 min. Thin layer chromatography (TLC) analysis showed the lipids to be phosphatidyl ethanolamine (PE), phosphatidylethanol (PEt) and phosphatidic acid (PA). There was no detectable level of oleic acid (OA), diacylglycerol (DAG), phosphatidyl choline (PC) or phosphatidyl serine (PS). These findings suggested that a trypsin and heat sensitive enzyme/factor present in the serum had a role in melittin-mediated action. These findings further showed that melittin activated phospholipase D (PLD), without affecting phospholipase A(2) (PLA(2)) or phospholipase C (PLC) activity. PMID- 10708800 TI - Purification and characterization of a thrombin-like enzyme, elegaxobin, from the venom of Trimeresurus elegans (Sakishima-habu). AB - A thrombin-like enzyme, named elegaxobin, was purified from the venom of Trimeresurus elegans (Sakishima-habu) by gel filtration on Sephadex G-100, and ion-exchange chromatographies on Q-Sepharose Fast Flow and S-Sepharose Fast Flow. By this procedure, about 8.5 mg of purified enzyme was obtained from 1.1 g of the venom. The purified enzyme showed a single protein band in SDS-polyacrylamide electrophoresis under reducing condition and its molecular weight is 30,000. The specific activity of this enzyme toward tosyl-L-arginine methyl ester (TAME) was 490 TAME units/mg of protein. Elegaxobin clotted only rabbit fibrinogen whereas human and bovine fibrinogens were unaffected. In the fibrinogen-fibrin convertion, the enzyme released only fibrinopeptide A from rabbit fibrinogen, whereas fibrinopeptide B was not released. The N-terminal sequences (Val-Ile-Gly Gly) of this enzyme was identical to typical sequence of serine proteinases. PMID- 10708801 TI - Frequent occurrence of paralytic shellfish poisoning toxins as dominant toxins in marine puffer from tropical water. AB - Considerably high toxicity was detected in marine puffers collected from Masinloc Bay, Philippines. The toxicity was detected in the liver, intestine, muscle and skin. Noteworthy, the specimens, the muscle of which showed high toxicity, appeared in high frequency, indicating that puffers from this area is not safe for human consumption. These puffer specimens contained paralytic shellfish poisoning (PSP) toxins, often as major toxin components, the profile of which was similar to that of freshwater puffers reported from tropical areas. These results indicate that PSP toxins are common in tropical puffers both from marine and freshwater. PMID- 10708802 TI - Sex differences in the pharmacological activity of venom from the white-tailed spider (Lampona cylindrata). AB - This study compared the pharmacological activity of venom from male and female white-tailed spiders (L. cylindrata). In guinea-pig ileum, male L. cylindrata venom (1-10 microg/ml) caused dose-dependent contractions. The response to venom (5 microg/ml) was significantly inhibited by mepyramine (0.5 microM). Venom (5-50 microg/ml) from female L. cylindrata had no contractile activity in this tissue. However, female L. cylindrata venom (50 microg/ml) inhibited electrically-evoked twitches of guinea-pig ileum. This inhibitory effect was attenuated by 8 phenyltheophylline (10 microM) or by prior exposure of venom to adenosine deaminase. In the rat vas deferens, male (5 microg/ml) and female (50 microg/ml) L. cylindrata venom inhibited electrically-evoked twitches. 8-Phenyltheophylline (20 microM) significantly attenuated the response to female L. cylindrata venom, while the histamine H(2)- and H(3)-receptor antagonists ranitidine (10 microM) and thioperamide (0.2 microM) significantly attenuated the response to male L. cylindrata venom. Male L. cylindrata venom (5-20 microg/ml) caused dose-dependent contractions in the epididymal segment of the rat vas deferens. The response to male L. cylindrata venom (10 microg/ml) was significantly inhibited by prazosin (0.3 microM) but was unaffected by depleting monoamine stores with reserpine. Male L. cylindrata venom (5-15 microg/ml) caused dose-dependent increases in rate and force of rat atria which were significantly inhibited by propranolol (5 microM) but not by reserpine. Female L. cylindrata venom (50 microg/ml) had no effect in atria. In the anaesthetised (pentobarbitone, 100 mg/kg, i.p.) rat, male L. cylindrata venom (10-300 microg/kg, i.v.) caused dose-dependent depressor responses while venom (up to 1 mg/kg, i.v.) from female L. cylindrata had no effect on arterial pressure. A histamine content of 5 and 0.01% (dry weight) was detected in venom from male and female L. cylindrata, respectively. Venom from male L. cylindrata was found to contain 56 pg noradrenaline/microg whereas venom from the female contained negligible noradrenaline. The results of this study show the presence of histamine and noradrenaline in venom from male L. cylindrata. Although devoid of significant quantities of these amines, female L. cylindrata venom has activity at adenosine receptors. PMID- 10708803 TI - Molecular mechanism of lung hemorrhage induction by VRV-PL-VIIIa from Russell's viper (Vipera russelli) venom. AB - The basic phospholipase A(2) (VRV-PL-VIIIa) from Vipera russelli venom induces multiple toxic effects including neurotoxicity, myotoxicity, edema and hemorrhage. This phospholipase A(2) has been extensively characterized for its pharmacological properties except for hemorrhagic activity. In the present investigation, the lung hemorrhagic activity was assayed using lung dye diffusion method. The investigations to understand the mechanism of lung hemorrhage induction by VRV-PL-VIIIa was followed by chemical modification studies and also by interaction with an antihemorrhagic factor p-anisic acid (4-methoxy benzoic acid). In presence of 1:2 mol:mol PLA(2): anisic acid, the lung hemorrhagic and edema inducing activities were completely neutralized in experimental animals; however, catalytic and anticoagulant activities were not neutralized. Carbamylation of VRV-PL-VIIIa resulted in the loss of lung hemorrhage and edema inducing activities. In contrast, carbamylation of VRV-PL-VIIIa in the presence of anisic acid could not neutralize the lung hemorrhage and edema inducing activities. The anticoagulant and enzyme activities were only partially neutralized when carbamylated both in the presence and absence of anisic acid. PMID- 10708804 TI - Neutralization of Thalassophryne nattereri (niquim) fish venom by an experimental antivenom. AB - T. nattereri (niquim) is a venomous fish involved in many human accidents in Brazil. The clinical picture includes mild local erythema, severe edema, intense pain and rapid progression to necrosis. The present therapy with anti inflammatory and analgesic drugs is ineffective and, therefore, we decided to assess serum therapy as an alternative treatment using an experimental antivenom. The antivenom used was raised in rabbits showing an ELISA antibody titer of 1:8,192,000 and its ability to neutralize lethality, necrosis, nociception and edema was evaluated both by pre-incubating the venom with antivenom before injection into mice or by independent injections of venom and antivenom. Lethality was completely neutralized by pre-incubation (ED(50)=141.5 microl/mg) while necrosis and nociception were neutralized by pre-incubation or the independent injection of antivenom. Edema was only partially prevented even when large amounts of antivenom were used. These data suggest that antivenom may be a promising treatment for patients stung by T. nattereri and suggest the viability of producing a horse antivenom for use in clinical trials. PMID- 10708805 TI - Study of acute chagasic mice under immunosuppressive therapy by cyclosporin A : modulation and confocal analysis of inflammatory reaction. AB - The in vivo effects of cyclosporin A (CsA) on Trypanosoma cruzi infection were examined using different schedules of the drug in mice infected with the Y strain. Parasitaemia at day 8 after infection among CsA-treated animals was usually higher than control infected non-treated mice. On the other hand, mortality analysis showed that animals CsA-treated either with 200 mg/kg 2 days before infection or with therapeutic doses (10 mg/kg every other day) showed almost the same mean time of death (35.8 and 38.2 days, respectively). In these groups mice died 50% less than control infected non-treated ones. The mean time of death in the animals treated with 200 mg/kg 5 days after infection and in infected non-treated control mice were respectively 29.0 and 22.6 days. The kinetics analysis of the leukocyte population of animals treated with a single dose of 200 mg/kg of CsA before or after infection did not show the alternate pattern of leukopenia/leukocytosis observed in control groups of infected mice but differential cell counts indicated a modulatory action upon circulating leukocytes of therapeutic doses of CsA. The animals treated with any of the CsA schedules showed a moderate to intense diffuse inflammatory reaction exhibiting mainly mononuclear cells in the heart. Immunofluorescence analysis by confocal microscopy revealed that macrophages are a major component of the inflammatory infiltrate in all groups of CsA-treated mice and also in the control group. PMID- 10708806 TI - Extracts of smokeless tobacco induce pro-inflammatory changes in cultured human vascular endothelial cells. AB - Habitual use of smokeless tobacco leads to accumulation of inflammatory leukocytes at the site of placement, which may contribute to tissue damage. Recruitment of leukocytes is facilitated by the endothelial lining of blood vessels, which can be activated to express adhesion molecules and to produce chemoattractants. The ability of aqueous extracts of chewing tobacco, dry snuff, and moist snuff to stimulate such changes was investigated using cultured human umbilical vein endothelial cells (HUVEC). All three extracts caused HUVEC to express the adhesion molecule E-selectin and to produce the chemokines interleukin-8 and monocyte chemoattractant protein-1. Neutrophils migrated avidly across HUVEC monolayers that had been previously exposed to the extracts, whereas migration across unstimulated monolayers was negligible. The smokeless tobacco extracts contained relatively high concentrations of bacterial lipopolysaccharide (LPS). Although LPS appeared to be the major stimulatory component in extracts of chewing tobacco, it accounted for only part of the activity found in extracts of moist and dry snuffs. These observations suggest that smokeless tobacco may induce inflammatory changes in vivo by activating endothelium in a manner that promotes recruitment of leukocytes. PMID- 10708807 TI - Apoptosis inhibitory effect of the isothiourea compound, tri-(2-thioureido-S ethyl)-amine. AB - Sulfhydril compounds, some of them with immunomodulatory activity have also been shown to modulate the induction of apoptosis. This study was performed to assess the possible apoptosis inhibitory effect of the immunomodulatory compound tri-(2 thioureido-S-ethyl)-amine (K-1) in U-937 and HL-60 leukaemia cell lines as model systems. Treatment of U-937 and HL-60 cells with K-1 inhibited etoposide (ETO) induced apoptosis in both cell lines in a dose-dependent manner, IC(50)=200 microg/ml. The results indicate that inhibition of ETO-induced apoptosis occurs downstream of ETO-mediated cleavable complex formation but early in, or at the level of the mitochondria events of the apoptotic pathway. Further, like other isothioureas, K-1 proved to be a potent inhibitor of nitric oxide synthase. This raises the possibility that where the activation of nitric oxide synthase is involved in apoptosis induction, K-1 might also be effective. These findings suggest that K-1 may serve as a potent inhibitor of apoptosis initiated by ETO or nitric oxide. PMID- 10708808 TI - Methotrexate suppresses the interleukin-6 induced generation of reactive oxygen species in the synoviocytes of rheumatoid arthritis. AB - Various cytokines and reactive oxygen species (ROS) play a fundamental role in the inflammatory and immunologic processes of rheumatoid arthritis (RA). Methotrexate (MTX) is one of the disease-modifying anti-rheumatic drugs and its effect may be partly due to the modulation of immunologic or inflammatory reactions by some cytokines. In the present study, we investigated the effects of MTX on the gene expression and synthesis of interleukin-6 (IL-6), and the proliferative activity and the production of ROS in the fibroblast-like synoviocytes (FLSs) obtained from the patient of RA. The expression or production of IL-6 was induced spontaneously, and augmented by the addition of recombinant human IL-6 or recombinant human IL-1 beta and TNF-alpha in FLSs. These spontaneous and augmented IL-6 expressions or productions were suppressed by treatment with low-concentration of MTX (1 microg/ml). Also, IL-6 stimulated the proliferation of FLSs, and this IL-6 driven proliferation was inhibited with the treatment of MTX or N-acetylcysteine (NAC, 1 mM). Furthermore, ROS production in FLSs was increased significantly by IL-6, and its effect was also abrogated in the presence of MTX or NAC. These results suggest that inflammatory reaction in the synovium of RA patients could be augmented by the autocrine or other cytokine induced production of IL-6 with subsequent generation of ROS in the synoviocytes, and the modulations of IL-6 synthesis and ROS production may contribute to the therapeutic effects of MTX for RA. PMID- 10708809 TI - Immunological effects of the orally administered neuraminidase inhibitor oseltamivir in influenza virus-infected and uninfected mice. AB - Oseltamivir (GS4104), the ethyl ester prodrug of the carbocyclic transition state sialic acid analog GS4071, has been reported to be a striking inhibitor of influenza A and B virus infections in mice and ferrets. Multiple studies indicate this material to also be active against the disease in humans, and it has recently been approved for human use. The effect of oral gavage (p.o.) therapy of oseltamivir on various immune factors considered to be of importance in primary influenza virus infection was studied in mice. Both uninfected animals and those infected with influenza A/NWS/33 (H1N1) virus were used. Doses of 100 mg kg(-1) day(-1) were administered twice daily for 5 days beginning 16 h pre-virus exposure. Two hours after end of treatment, the mice were killed and their spleens assayed for cytotoxic T lymphocyte (CTL) and natural killer (NK) cell activity. Subpopulations of splenic T, T-helper, T-cytotoxic and B lymphocytes as well as macrophages were determined using flow cytometry. Similar significant (P<0.01) increases in CTL activity were seen at effector:target cell ratios of 60:1 and 30:1 in the infected mice treated with oseltamivir or with placebo. NK cell activity was greater in the infected mice than in uninfected mice; the levels in all animals were not significantly affected by treatment with oseltamivir. Macrophage, T, T-helper, T-cytotoxic and B lymphocyte populations were similar in both treated and untreated animals. These data indicate treatment with oseltamivir does not adversely affect the primary in vivo cellular immune responses to influenza virus infection assayed in this study. The experiment was repeated to show that treatment with this compound significantly prevented the development of the infection and inhibited virus titers in the lung. Surviving treated mice on day 21 had mean neutralizing antibody titers of 1:208, and withstood rechallenge with the virus at this time, indicating the initial virus inhibitory effect also did not prevent the animals from developing an adequate humoral immunity to the virus. PMID- 10708810 TI - Opioids suppress chemokine-mediated migration of monkey neutrophils and monocytes - an instant response. AB - Opioid users having acquired human immunodeficiency syndrome (AIDS) are at a greater risk than non-users of contracting opportunistic infections. Opioid administered and simian immunodeficiency virus (SIV)-infected rhesus monkeys have been an excellent model for studying AIDS and drug abuse in humans. In this study, chemotaxis of monkey leukocytes was evaluated using the chemokines interleukin-8 (IL-8) and regulated upon activation, normal T cell expressed (RANTES) as the chemoattractants, and the effects of various opioid agonists and antagonists on the efficiency of chemotaxis were examined. Opioids were either incubated with monkey leukocytes or added directly to chemokines, and the number of cells migrating toward IL-8 (for neutrophils) or RANTES (for monocytes) was scored. Inhibition of chemotaxis was seen with both assay conditions, and the inhibition was mediated by opioids binding to mu or kappa receptors. Binding to delta opiod receptors was rarely, if ever, observed. Although opioids themselves may act as weak chemoattractants for monkey leukocytes, addition of opioid agonists to chemokines would reduce the chemoattractant ability of the chemokines. Opioids did not cause the same inhibitory effect on the chemotactic migration of neutrophils when the complement component C5a or the chemotactic peptide N-formyl-MET-LEU-PHE (fMLP) was used as chemoattractant. These studies suggest that the presence of opioids during SIV infection immediately alters chemokine-mediated immune functions. PMID- 10708811 TI - Relationship between depression and cerebrovascular disease: conceptual issues. AB - OBJECTIVES: The association between cerebrovascular disease (CVD) and depression has been well described, but our understanding of various aspects of the relationship between these two disorders remains limited. METHOD: Based on a selective literature review, this paper examines empirical evidence and discusses conceptual issues concerning hierarchical, interactive, and co-morbid relationships between CVD and depression. RESULTS: The concept of vascular depression minimizes the importance of the contribution of psychosocial factors. The interactive and co-morbid relationships have been largely neglected in psychiatric research. There is evidence that depression may increase the risk of CVD morbidity in patients with vascular disease and delay recovery in stroke patients, implying an interactive relationship. The concurrent existence of these two disorders based on common etiological factors such as genetic vulnerability, alcoholism and personality traits seems plausible. CONCLUSIONS: A modified comorbidity model may guide investigation into the hierarchical, interactive and common etiological relationships between CVD and depression. PMID- 10708812 TI - The current status of the platelet 5-HT(2A) receptor in depression. AB - The author reviews the current status of the platelet serotonin (5-HT)(2A) receptor in depression. Considered are studies of receptor binding, and 5-HT induced platelet activation and aggregation. 5-HT(2A) receptor density tends to increase in depression, although this more clearly relates to suicidality than depression per se. Indeed, data are consistent with the hypothesis that increased density of platelet 5-HT(2A) receptors may be a marker for increased risk of suicide. 5-HT-induced calcium mobilization is enhanced in unipolar depression; however, unlike in bipolar depression, baseline calcium levels are not. Despite inconsistencies, 5-HT-induced aggregation appears inhibited in depression. This may manifest as a relative inhibition, i.e. no change in aggregation response despite a higher density of 5-HT(2A) receptors. The inhibited aggregation response is state dependent, and acute phase proteins or components of the stress response may be factors. It is unclear if differences between depressed and normal subjects in disposition of 5-HT(2A) receptors are generally indicative of traits or states. Nonetheless, there is little evidence that the degree of departure from normal density or activity of platelet of 5-HT(2A) receptors reflects severity of depression. PMID- 10708813 TI - Evidence for genetic influences common and specific to symptoms of generalized anxiety and panic. AB - BACKGROUND: Generalized anxiety disorder (GAD) and panic disorder (PD) often co occur and have been shown to be heritable. Researchers have debated the validity of the distinction between GAD and PD. To test for distinction between disorders, we estimated the genetic and environmental contributions which were specific and common to GAD and PD in a cohort of male-male twin pairs. METHODS: Data were obtained from a telephone interview performed in 1992 utilizing the Diagnostic Interview Schedule Version 3-Revised. Interviews were administered to 6724 male male monozygotic and dizygotic twin pair members of the Vietnam Era Twin Registry. We defined lifetime GAD by the report of six or more DSM-III-R symptoms and lifetime PD by the report of four or more DSM-III-R symptoms. RESULTS: The lifetime co-occurrence of GAD and PD was best explained by a model which did not include family environmental influences. The variance in risk for GAD was due to a 37.9% influence from additive genetic factors with the remainder due to unique environmental influences. The variance in risk for PD was due to a 22.6% additive genetic contribution which was common with GAD and a 21.2% non-additive genetic contribution specific to PD with the remainder of variance in risk for PD due to unique environmental influences. LIMITATIONS: Results may be limited to middle aged males. Model fitting with full diagnostic criteria was not possible due to low prevalences. CONCLUSIONS: Our data suggest a distinction in liability for GAD versus PD. The common genetic influence to GAD and PD may partially account for the risk of the co-occurrence of these disorders in a lifetime. PMID- 10708814 TI - Diurnal variations in endocrine and psychological responses to 0.2 mg/kg naloxone administration in patients with major depressive disorder and matched controls. AB - BACKGROUND: There is evidence that the endogenous opioid system (EOS) is involved in the modulation of mood and neuroendocrine function. Furthermore, the possible involvement of the EOS in major depression has been postulated, although a clear role has not been established. METHODS: The affective and endocrine responses to naloxone administration in seven female depressives and in seven matched controls and their diurnal variations were investigated. Subjects had an i.v. bolus of either 0.2 mg/kg naloxone or saline at two time points (09:00 or 18:00 h) and for 2 days in a single-blind, cross-over design. RESULTS: The basal cortisol plasma levels, both in the morning and in the afternoon, showed higher values (P<0.05) in the depressives. There was a naloxone-induced increase in the adrenocorticotrophic hormone (ACTH), cortisol, and luteinizing hormone (LH) plasma levels, plus a subjective dysphoric effect in both groups. The depressives showed a greater dysphoric effect both in the morning and afternoon (P<0.05), and a blunted cortisol response in the afternoon (P<0.05). There were no differences between groups or time of day in the ACTH or LH responses. LIMITATIONS: The sample size was small, but by studying each patient as their own control, plus a matched control for every patient, softens this effect. Finding patients with a major depressive episode free of medication is difficult, and this aspect contributes to the size of the sample. CONCLUSIONS: These results suggest that opioid mechanisms may be involved in the HPA axis changes and possibly in mood changes found in depression. The discrepancy between increased sensitivity in depression to mood changes and decreased change in cortisol may indicate a ceiling effect for the latter. PMID- 10708815 TI - The course of depression in elderly subjects with and without dementia. AB - BACKGROUND: A persistent course of depression has been described in subjects with and without dementia. Up to the present it is unclear to what extent dementia affects the prognosis of depression. METHOD: At baseline and at 6 and 12 months follow-up AGECAT depression diagnoses were made in 49 subjects with and 72 subjects without DSM-III-R dementia living in homes for the elderly. RESULTS: Adjusting for demographic characteristics and physical health, dementia was not associated with the severity of depression at follow up. The baseline depression severity and to a lesser extent somatic complaints predicted a bad prognosis of depression in the total sample. LIMITATIONS: Because of the high vulnerability of the residents the results cannot be generalised to other populations of elderly subjects. CONCLUSION: Depression is persistent in residents of homes for the elderly. Dementia does not affect its course. PMID- 10708816 TI - Bright light improves vitality and alleviates distress in healthy people. AB - BACKGROUND: The relative shortage of light during the decreasing photoperiod may compromise well-being. Earlier studies suggest that bright-light exposure may be of help to alleviate winter-bound symptoms. METHODS: We carried out a field study with exposure to bright light on office employees during winter. RESULTS: Repeated bright-light exposure improved vitality and reduced depressive symptoms. The benefit was observed not only in healthy subjects with season-dependent symptoms but also in those not having the seasonal variation. CONCLUSIONS: Bright light exposure during winter appears to be effective at improving the health related quality of life and alleviating distress in healthy subjects. CLINICAL IMPLICATIONS: Administration of bright light is a useful option to improve vitality and mood among subjects working indoors in wintertime. LIMITATIONS OF STUDY: Our field setting used self-reports, not interviews, for the assessment of outcome. PMID- 10708817 TI - Is cognitive behavior therapy just a 'nonspecific' intervention for depression? A retrospective comparison of consecutive cohorts treated with cognitive behavior therapy or supportive counseling and pill placebo. AB - BACKGROUND: There is a dearth of placebo-controlled studies of cognitive behavior therapy (CBT) of depression and the largest such study, by Elkin et al. (Arch. Gen. Psychiatry 46 (1989) 971-982), failed to find a significant difference between CBT and a clinical management plus placebo condition. METHODS: The outcomes of two consecutive cohorts of out-patients with major depressive disorder, treated with either CBT (n=90) or a nonspecific control condition (support-counseling-placebo; SCP: n=100), were compared. Although the principal comparisons between the CBT and SCP conditions were delimited to the first 4 weeks of treatment, a secondary set of analyses addressed the subset of 16 patients who received 12 additional weeks of supportive therapy. RESULTS: A consistent pattern of statistically and clinically significant differences favoring CBT over SCP was found in both weeks 4 and 16. LIMITATIONS: Interpretation of these findings are subject to several potential confounds, including the non-randomized nature of the groups and the greater amount of therapeutic contact during the first 4 weeks of CBT. CONCLUSIONS: While these results do not lessen the need for additional prospective studies, our findings do suggest that CBT has therapeutic effects beyond those attributable to placebo expectancy and other nonspecific factors. PMID- 10708818 TI - The benefit from whole body acupuncture in major depression. AB - BACKGROUND: In a single-blind placebo-controlled study design we investigated the efficacy of acupuncture additionally applied to drug treatment in major depression. METHODS: We randomly included 70 inpatients with a major depressive episode in three different treatment groups: verum acupuncture, placebo acupuncture and a control group. All three groups were pharmacologically treated with the antidepressant mianserin. The verum group received acupuncture at specific points considered effective in the treatment of depression. The placebo group was treated with acupuncture at non-specific locations and the control group received pharmacological treatment plus clinical management. Acupuncture was applied three times a week over a period of 4 weeks. Psychopathology was rated by judges blind to verum/placebo conditions twice a week over 8 weeks. RESULTS: Patients who experienced acupuncture improved slightly more than patients treated with mianserin alone. CONCLUSIONS: Additionally applied acupuncture improved the course of depression more than pharmacological treatment with mianserin alone. However, we could not detect any differences between placebo and verum acupuncture. PMID- 10708819 TI - Motor and cognitive aspects of motor retardation in depression. AB - BACKGROUND: Motor retardation is a common feature of major depressive disorder having potential prognostic and etiopathological significance. According to DSM IV, depressed patients who meet criteria for psychomotor retardation, must exhibit motor slowing of sufficient severity to be observed by others. However, overt presentations of motor slowing cannot distinguish slowness due to cognitive factors from slowness due to neuromotor disturbances. METHODS: We examined cognitive and neuromotor aspects of motor slowing in 36 depressed patients to test the hypothesis that a significant proportion of patients exhibit motor programming disturbances in addition to psychomotor impairment. A novel instrumental technique was used to assess motor programming in terms of the subject's ability to program movement velocity as a function of movement distance. A traditional psychomotor battery was combined with an instrumental measure of reaction time to assess the cognitive aspects of motor retardation. RESULTS: The depressed patients exhibited significant impairment on the velocity scaling measure and longer reaction times compared with nondepressed controls. Approximately 40% of the patients demonstrated abnormal psychomotor function as measured by the traditional battery; whereas over 60% exhibited some form of motor slowing as measured by the instruments. Approximately 40% of the patients exhibited parkinsonian-like motor programming deficits. A five-factor model consisting of motor measures predicted diagnosis among bipolar and unipolar depressed patients with 100% accuracy. LIMITATIONS: The ability of motor measures to discriminate bipolar from unipolar patients must be viewed with caution considering the relatively small sample size of bipolar patients. CONCLUSIONS: These findings suggest that a subgroup of depressed patients exhibit motor retardation that is behaviorally similar to parkinsonian bradykinesia and may stem from a similar disruption within the basal ganglia. PMID- 10708820 TI - Reliability and validity of the DSM-IV diagnostic category of schizoaffective disorder: preliminary data. AB - BACKGROUND: Concerns have been expressed about the reliability and validity of the DSM-IV criteria for schizoaffective disorder, but no systematic study has been published up to now. METHODS: The Cohen's kappa for the individual items of the DSM-IV definition of schizoaffective disorder, manic episode and major depressive episode was evaluated in 150 patients independently interviewed by two psychiatrists using the Composite International Diagnostic Interview. The two year outcome of patients with a consensus DSM-IV diagnosis of schizoaffective disorder was compared to that of patients with DSM-IV schizophrenia and schizophreniform disorder, using the Strauss-Carpenter Outcome Scale. RESULTS: The Cohen's kappa was 0.22 for the diagnosis of schizoaffective disorder, 0.71 for that of manic episode, and 0.82 for that of major depressive episode. Schizoaffective patients had a significantly better outcome than those with schizophrenia but a worse outcome than those with schizophreniform disorder. CONCLUSIONS: The inter-rater reliability of the DSM-IV criteria for schizoaffective disorder is not satisfactory. The better outcome of DSM-IV schizoaffective disorder compared with schizophrenia seems to depend more on the inclusion, in the definition of schizophrenia but not in that of schizoaffective disorder, of the six-month duration and functional impairment criteria than on the different symptomatological patterns of the two conditions. LIMITATION: The size of the sample of patients fulfilling DSM-IV criteria for schizoaffective disorder was small. CLINICAL RELEVANCE: The study suggests that the clinical implications of the currently problematic diagnosis of schizoaffective disorder may be modest. PMID- 10708821 TI - Urban/rural and gender differentials in suicide rates: east and west. AB - BACKGROUND: Many epidemiological studies indicate suicide rates are higher for males than females and for urban than rural. Here we re-examine gender, urban and rural differentials in suicide in Australia and Beijing (China). More specifically, to test the two hypotheses (i) that the male to female ratio is larger than one; (ii) that the urban suicide rate is higher than the rural in both places. METHODS: Suicide data with information of gender, rural and urban regions for Australia and Beijing (China) for the period of 1991-1996 were used. Ratios between the gender-specific urban and rural suicides rates with the associated confidence intervals were constructed to examine gender, urban and rural differentials in Australia and Beijing. RESULTS: The rural suicide rate in Beijing for both genders was higher than for their urban counterparts. Further, the elderly had the highest suicide rate followed by women aged 20-29. Also, the male to female ratio in China was less than one. In Australia, the rural male suicide rate was higher than the urban whereas the urban female suicide rate was higher than the rural. The male to female ratio was 4 to 1. The differences in rural to urban and male to female ratios between Australia and Beijing are statistically significant. CONCLUSIONS: In contrast to the west, male suicide rates are not higher than female rates in China. Urban rates are not necessarily higher than rural rates --not even in a western setting. Cultural factors and regional differences in socio-economic situation are significant in explaining the low gender ratio and the relatively higher suicide rates in rural China. LIMITATIONS: The suicide rate in the Beijing region might not exactly reflect the same for the whole of China. PMID- 10708822 TI - A control study of the cutaneous side effects of chronic lithium therapy. AB - BACKGROUND: To assess the nature and prevalence of skin disorders among psychiatric patients on chronic lithium therapy and to compare them with patients on other psychotropic medications. METHOD: 51 patients on lithium and 57 patients on other psychotropics were recruited. Dermatological assessment included a semi structured questionnaire and clinical examination of the subjects by two dermatologists who were blind to the psychiatric diagnosis and treatment. Secondary cutaneous reaction was defined as skin eruption that developed or deteriorated after commencement of psychiatric medication. RESULTS: Lithium treated patients developed significantly more secondary cutaneous reactions than the control group. This applied particularly to acne and psoriasis. Male patients on lithium were more likely to be affected than female patients. CONCLUSION: Lithium aggravates or triggers cutaneous conditions that are characterized by the pathological findings of neutrophilic infiltration. Since these cutaneous problem can be distressing to patients and may affect medication compliance, there should be heightened attention to skin problems in patients receiving lithium treatment. PMID- 10708823 TI - Differential clinical features of late-onset panic disorder. AB - OBJECTIVES: The aim was to analyse the sociodemographic and clinical characteristics of panic disorder (PD) in patients with a PD onset after 60 years of age, at two outpatient psychiatric clinics in Barcelona (northeastern Spain). MATERIAL AND METHODS: All patients presenting with PD at two outpatient clinics over a 4-year period were assessed by the same team. Patients with PD onset at 60 or after were grouped (late-onset), and compared with the group with an earlier onset. The instruments administered to the sample were: Global Assessment of Functioning scale, Panic-Associated Symptom Scale, Hamilton's Depression and Anxiety Scales and Marks-Matthews' Fear and Phobia scale. RESULTS: Of 5301 patients attended over a 4-year period, 64 (1.2%) were PD patients aged 60 or above. Age at PD onset was over 60 in 27 cases (0.4% of the total population, and 6.1% of all PD patients). The mean age in the late-onset group was 67.0+/-4.9 years. Late-onset PD patients were less likely to report family history of PD. They scored lower on most scales assessing clinical severity (excepting GAF and agoraphobia scores), and they exhibited fewer and milder panic symptoms during the attacks. However, dysthymic disorder, but not major depressive disorder, was more common among late-onset PD patients (P<0.05). COMMENTS: The most notable findings in our late-onset PD subgroup of patients were: lesser severity of the disorder, greater comorbidity with dysthymia, and less family history of PD. Prevalence rates of late-onset PD in our sample appeared to be rather high. Physical illness and less severe panic symptoms may contribute to underdiagnosing PD in this particular subpopulation. PMID- 10708824 TI - Effects of pregnancy and delivery on serum prolyl endopeptidase (PEP) activity: alterations in serum PEP are related to increased anxiety in the early puerperium and to postpartum depression. AB - BACKGROUND: There is now some evidence that anxiety or anxiety disorders are related to increased activity of serum prolyl endopeptidase (PEP) and that major depression is related to lower serum PEP. The aims of the present study were to examine (i) the effects of pregnancy and delivery on serum PEP and (ii) the relationships between serum PEP and postpartum depression, anxiety in the early puerperium and a past history of depression. METHODS: Serum PEP activity was measured in 11 healthy nonpregnant and in 98 pregnant women 3 days before delivery and 1 and 3 days after delivery. On the same occasions, pregnant females completed the Spielberger State Anxiety Inventory (STAI) and were divided into high and low anxiety responders, as defined by changes in the STAI. The presence of a previous depression and postpartum depression within 3 months of delivery was assessed by means of DSM-IV criteria. RESULTS: Serum PEP activity was significantly higher 1 and 3 days after delivery than before. Women with a past history of depression as well as anxiety responders had significantly increased serum PEP activity over nonpregnant women and puerperae with a negative history and anxiety nonresponders, respectively. Parturients who developed a postpartum major, but not minor, depression had significantly lower serum PEP than parturients without postpartum depression. The results were controlled for maternal and labor variables, such as type of analgesia and delivery, induction of labor, breast feeding, parity, and duration of pregnancy and labor. CONCLUSIONS: Our results show that, in puerperae, increased serum PEP is related to increased state anxiety in the early puerperium and that lowered serum PEP is related to a subsequent postpartum major depression. INTERPRETATION: The results suggest that increased serum PEP may be related to postpartum anxious blues and that lowered serum PEP may predispose toward postpartum major depression. PMID- 10708825 TI - Predictors of recurrence in affective disorder--analyses accounting for individual heterogeneity. AB - BACKGROUND: Previous studies suggest that gender, age at onset, and marital status act as risk factors for further recurrence initially during the course of affective disorder but not at a later stage. These studies did, however, not take the individual liability to recurrence into account. METHOD: The effect of predictors of recurrence was estimated with the use of generalised linear mixed models in a case register study including a random sample of all patients admitted with primary affective disorder in Denmark during 1971-1993. RESULTS: In total, 7047 first admission patients with a diagnosis of affective disorder, depressive or manic/circular type were included in the analyses. The study confirmed that the effect of the type of disorder, age at first admission, and never being married decreased during the course of illness even when the individual liability to recurrence was taken into account. No differences in the effect of gender and in the effect of a recent divorce were found between early and later episodes and the effect of a recent death of a spouse seemed to increase during the course of illness. The risk of recurrence increased with every new episode for all sub-groups of patients. CONCLUSION: The effect of some, but not all, predictors of recurrence decline during the course of affective illness. The number of previous episodes predicts recurrence in most subgroups of patients. LIMITATION: The data relate to re-admissions rather than recurrence. CLINICAL RELEVANCE: The study underscores the importance of the illness process itself. PMID- 10708826 TI - The directed forgetting task: application to emotionally valent material. AB - Three experiments are reported which investigate the application of the directed forgetting task to emotionally valent material and to different mood states. In this task, some subjects are told when halfway through an intentional or incidental learning task that the trials presented so far are to be forgotten because they were merely practice. However, at the end of the subsequent list, the subjects are then asked to recall all of the previous items including those that they were instructed to forget. The results typically show that significantly fewer directed forgetting items are recalled whether the task is an intentional or incidental learning one. In the first experiment, normal and 'depressed' students rated positive and negative material for pleasantness; although directed forgetting effects were obtained, there were no differential effects of mood state nor of valence of the material. In order to investigate this effect further, a variant of this task was used in Experiment 2 in which the positive and negative material had to be processed in relation to the self. The results showed that differential forgetting now occurred; healthy students recalled more positive than negative information, but this positive bias was not obtained for 'depressed' students who showed an even-handed level of recall. In Experiment 3, groups of clinically depressed, clinically anxious, and normal controls were presented with the directed forgetting task. The key finding showed that the depressed subjects showed a retrieval facilitation for to-be-forgotten negative adjectives, an effect that was not present for the other two groups. It is concluded therefore, that the directed forgetting task could be usefully extended to investigate cognition-emotion interactions in clinical populations. PMID- 10708827 TI - Remission onset and relapse in depression. An 18-month prospective study of course for 100 first admission patients. AB - BACKGROUND: Few prospective studies of course for first admission depressives are reported. METHODS: One hundred consecutive depressed inpatients were followed prospectively over 18 months. Course was defined operationally using the Hamilton Depression scale and ICD-10 criteria. Results were analysed using life-tables. RESULTS: The cumulative probabilities of remission onset by 3 and 18 months were 0.67 (95% C.I.=0.57-0.77) and 0.82 (95% C.I.=0.74-0.90). The cumulative probability of relapse was 0.25 (95% C.I.=0.15-0.35); 53% of those relapsing did so in the first 2 months. Younger age at onset, longer illness length, higher depression and anxiety ratings, predicted delayed remission onset. ICD-10 episode severity predicted relapse. CONCLUSIONS: The chances of remission onset at 3 months and relapse were increased relative to other studies; risk of chronicity was similar. Predictors of outcome to emerge were similar to other studies. CLINICAL IMPLICATIONS: Adoption of these remission onset criteria may identify earlier (at 3 months), subjects at high risk of chronicity. After remission onset, subjects with severe illnesses warrant careful follow-up to detect relapse, particularly during the first 2 months. LIMITATIONS: The operational criteria used were different to other prospective studies. Relatively few psychosocial variables were included in the analysis. PMID- 10708828 TI - 'Acting out' and 'acting in' behavioural stress responses: the relevance of anxiety and personality style. AB - OBJECTIVE: To look for links between 'acting out' and 'acting in' behavioural stress responses with anxiety and disordered personality function. METHOD: Depressed patients completed a self-report measure of behavioural responses to stress 1 year after baseline assessment of anxiety levels, personality functioning and other study variables. RESULTS: Patients were assigned to four factorial groups on the basis of variable 'acting out' and 'acting in' scale scores, and effects of individual scale scores also examined. 'Acting in' styles were indicative of high trait anxiety, disordered personality functioning and, in particular, a Cluster C personality disorder style. 'Acting out' was linked most clearly with a Cluster B personality disorder style. CONCLUSIONS: We demonstrate links between behavioural stress responses and anxiety levels and disordered personality functioning. Assessing behavioural stresses responses may shape the clinical expression of some depressive disorders and inform the clinician about the likely salience of anxiety and personality styles. PMID- 10708829 TI - Paternal remarriage as a modifier of proneness to depression in young adulthood. AB - BACKGROUND: The remarriage of a parent is a major event experienced by many children. Its role in children's depressive symptoms was examined in a follow-up study of a cohort from the age of 16 to 22 years. METHODS: The study population consisted of young people who had experienced parental divorce in childhood (N=356). Associations between a parent's remarriage and potential modifying factors in adolescence (atmosphere at home, school performance, dating behavior, life-events, the importance of siblings, and socioeconomic status) and depressive symptoms at 22 years of age were studied. RESULTS: A poor atmosphere at home at 16 years and father's remarriage (but not mother's) in childhood were associated with subsequent depression. Girls whose father had not remarried, but boys whose father had remarried, were more depressive than others. Dating behavior in adolescence modified proneness to depression in these groups. CONCLUSION: The findings indicate the importance of the father in adolescent development after divorce and that the processes involved may differ by gender. PMID- 10708830 TI - Lifetime psychiatric comorbidity rate in Israeli non-help-seeking patients with combat-related post-traumatic stress disorder. AB - BACKGROUND: Most of the data on lifetime psychiatric comorbidity in combat related posttraumatic stress disorder (CR-PTSD) were collected in help-seeking patients. METHODS: In the present study we used the Schedule for Affective Disorder and Schizophrenia-Lifetime Version to examine a relatively large sample (n=80) of Israeli non-help-seeking CR-PTSD patients. The diagnosis of PTSD was based on the DSM-III-R criteria. RESULTS: We found a low rate of lifetime psychiatric comorbidity, especially drug dependence (2.25%), alcoholism (2.25%) and major depressive disorders (5%). CONCLUSION: It seems that in contrast to help-seeking CR-PTSD, non-help-seeking CR-PTSD is associated with a low frequency of comorbid psychiatric disorders. LIMITATION: Only non-help seeking CR-PTSD patients who agreed to participate in the study were included in this investigation. CLINICAL RELEVANCE: The detection and diagnosis of CR-PTSD comorbidity is important for establishing appropriate psychotherapeutic and pharmacological treatment. PMID- 10708831 TI - Lack of coordination of nonverbal behaviour between patients and interviewers as a potential risk factor to depression recurrence: vulnerability accumulation in depression. AB - BACKGROUND: Coordination of nonverbal behaviour during interactions is a prerequisite for satisfactory relationships. Lack of coordination may form a risk factor for depression. The 'vulnerability-accumulation hypothesis' assumes that vulnerability to recurrence of depression will increase with increasing experience of depressive episodes. Therefore it is expected that interviewers and patients remitted from a recurrent episode of depression would reach less coordination during a clinical interview compared to interviewers and patients remitted from a first lifetime episode. Moreover, we assumed that prior severity of depression modifies this reciprocal coordination process. METHODS: During discharge interviews, interviewers were videotaped in interaction with remitted patients with unipolar major depression recurrent depression (REC); n=28; first episode (SEP); n=22. Durations and frequencies of nonverbal involvement behaviour was registered during the first 15 min. Involvement of the patients consisted of gesticulating, looking at the interviewer, and general head movements; yes- nodding and hm, hm, yes, yes reflected involvement of the interviewer. Coordination between patients and interviewers was analyzed per 3-min epochs and defined as 'attunement': the absolute difference between patients' and interviewers' involvement. Averaged attunement, its time course and variability (presumably reflecting control of the attunement process) were assessed. RESULTS: The time course of nonverbal attunement differed between the REC and SEP condition. A larger variability of attunement was found in patients remitted from a relatively severely depressed episode, compared to patients remitted from a severe first life time episode. No other significant differences were found. CONCLUSIONS: Partial support was found for the notion that nonverbal vulnerability accumulates in depression and that the severity of prior depression modifies this process. PMID- 10708832 TI - Oral d-fenfluramine test in treatment-refractory depression. Plasma prolactin response compared in patients with and without suicide attempts and in a healthy reference group. AB - BACKGROUND: Fenfluramine (d-FEN) has been used as a serotonin challenge agent to assess central serotonin availability. Blunted serum prolactin (PRL) response to d-FEN has been reported in depressed patients, in suicide-prone patients, and in patients with aggression and personality disorders. We have analyzed suicidality in relation to central serotonergic events by comparing the PRL response to d-FEN in chronically depressed patients with and without suicide attempts and in healthy volunteers. METHODS: In 56 inpatients (10 patients with and 46 without suicide attempts) with at least 2 years of treatment-refractory depression (TRD) (DSM-IV) and a reference group of 30 healthy adults, the PRL response after an oral dose of 30 mg d-FEN was followed for 5 h. RESULTS: Controlling for group differences in age, sex, and weight, the PRL response to d-FEN did not differ significantly between the three groups. Far from confirming the hypothesis of a blunted PRL response in depressed patients, our results suggest: (1) that duration and treatment resistance of depression may affect the PRL secretion, and (2) that TRD and major depression may differ in biological relationship to suicidal behavior. LIMITATIONS: The findings require corroboration in larger and more closely matched study populations. The fenfluramine concentration was not analyzed in blood. CONCLUSIONS: PRL responses to d-FEN challenge did not differ between TRD patients with and without suicidality and the healthy reference group. Chronicity/treatment refractoriness per se may be related to a serotonergic mechanism. PMID- 10708833 TI - Do early adverse experiences establish a cognitive vulnerability to depression on exposure to mirroring life events in adulthood? AB - BACKGROUND: We pursue a 'lock and key' hypothesis which posits that early adverse events ('locks') create an increased vulnerability to depression in the face of mirroring life events ('keys') in adulthood. Here we examine whether any such vulnerability links are cognitively mediated. METHODS: We study a sample of 96 clinically depressed patients who reported an identifiable 'cognitive schema' being activated when depressed. We examine for significant associations between early adverse events and later precipitants to the patients' depression, and then assess the extent to which any identified links are cognitively mediated. RESULTS: Qualitative analyses suggested quite strong associations between early childhood experiences and identified schemas, while the quantitative analyses identified few links. LIMITATIONS: These contrasting results may present a challenge to the hypothesis or reflect methodological limitations, and we therefore detail some of the complexities involved in identifying cognitive schemas. PMID- 10708834 TI - Quality of well being in panic disorder: the assessment of psychiatric and general disability. AB - BACKGROUND: Panic disorder is a common and debilitating anxiety disorder which significantly disrupts the lives of patients and their family members. Recent epidemiological studies and analyses of data from clinical trials suggest that patients with panic disorder suffer significant work and social dysfunction. The authors hypothesized that this dysfunction could be characterized using both a well-validated, generalized scale of functioning and a specifically designed scale for assessing function in psychiatric patients and that these findings would correlate with symptomatology. METHOD: Fifty-six patients with panic disorder were characterized using the Sheehan Disability Scale, the Anxiety Sensitivity Index, and the Spielberger State Trait Anxiety Scale. Measures of health related quality of life from the Quality of Well Being Scale were compared with ratings for matched, historical, and population controls. RESULTS: Patients with panic disorder lost 39 quality-adjusted days for each year that they lived with the disorder. This decrease in quality of life is similar to what is observed in patients with non-insulin dependent diabetes. Diminished quality of life is correlated with the number of panic attacks, state anxiety, and depressive symptoms. These patients also demonstrated significant dysfunction in Sheehan total disability and subscale scores, including work-related functioning. CONCLUSIONS: This study demonstrates that the specific disabilities inherent in panic disorder can be linked to declines in quality of life as measured by the Quality of Well Being Scale and by measures of work-related dysfunction. Such an association between disease specific measures and a generalized measure of health related quality of life may offer clinicians a new tool to understand panic disorder and to conceptualize it within the broader context of disease and disability. PMID- 10708835 TI - Is self-criticism unique for depression? A comparison with social phobia. AB - BACKGROUND: This study further examined the diagnostic specificity of the self critical personality dimension, as measured by the Depressive Experiences Questionnaire (DEQ; Blatt et al., 1976. The Depressive Experiences Questionnaire. Yale University Press, New Haven). METHODS: Patients with major depression (n=26) were compared to social phobia patients (n=32). RESULTS: Depressed patients scored significantly higher on the DEQ Self-Criticism dimension. However, when current level of depressed mood was controlled for, self-criticism was not a significant predictor of diagnostic status. Further, the level of DEQ self criticism reported by patients with social phobia was almost three times greater than the level reported in an earlier diagnostic specificity study with panic disorder patients [Bagby et al., 1992. Diagnostic specificity of the dependent and self-critical personality dimensions in major depression. J. Affect. Disord. 26, 59-64]. LIMITATIONS: Only one measure of self-criticism was used in this study, and the research design was cross-sectional rather than prospective. CONCLUSIONS: Self-criticism is not unique to major depression, and this personality dimension may be implicated in other forms of psychopathology [Blatt, 1991. A cognitive morphology of psychopathology. J. Nerv. Ment. Dis. 179, 449 458]. Some cognitive features believed to play an important role in depression may also be salient in persons with social phobia. PMID- 10708836 TI - Parental child-rearing behavior as measured by the Parental Bonding Instrument in a Japanese population: factor structure and relationship to a lifetime history of depression. AB - BACKGROUND: Recent factor analyses showed that the variance of the Parental Bonding Instrument (PBI) was more satisfactorily explained by the use of three factors (one factor corresponding to the original care factor and two factors derived from the original protection factor), casting doubt as to the accuracy of previous estimations regarding the associations between parental rearing behavior and depressive disorders. METHODS: 418 employed Japanese adults completed the PBI and the Inventory to Diagnose Depression, lifetime version. Associations of PBI scores with lifetime history of depression were explored by performing logistic regression analyses. The analyses were carried out using the original two PBI dimensions and the three new PBI dimensions validated in this sample. RESULTS: The results of the analyses using the three new dimensions did not differ markedly from those using the original two dimensions. Parental low care was always associated with having a lifetime history of depression. Analyses using the three new PBI dimensions provided some evidence that overprotective aspects of parenting may also be associated with a lifetime history of depression (a higher score on paternal denial of the psychological autonomy dimension predicted a lifetime history of depression in female subjects). LIMITATIONS: Subjects were Japanese adults, which may limit the validity of the conclusion to Western cultures. Data regarding both child-rearing behavior and lifetime major depression were obtained by self-rating instruments, which also may have influenced the results. CONCLUSION: Low parental care may be the most important factor associated with depression, even though the factor-analytically appropriate three PBI dimensions are used. Evidence for an association between overprotective aspects of child-rearing behavior and a lifetime history of depression can be newly recognized using the three new PBI dimensions. PMID- 10708837 TI - Health-related quality of life using the SF-36 in patients with bipolar disorder compared with patients with chronic back pain and the general population. AB - BACKGROUND: The purpose of this study was to assess and compare the health related quality of life of patients with bipolar disorder and chronic back pain and, in turn, to compare these results with those previously generated for the general population. METHODS: Subjects were patients with bipolar disorder (n=44), a comparison group of chronic back pain patients (n=30), and a population-based control sample (n=2,474). Health-related quality of life was assessed using the Medical Outcomes Study 36-Item Short-Form Health Survey (SF-36), a self administered questionnaire in which lower scores are indicative of greater impairment. RESULTS: Patients with bipolar disorder had lower mean scores than the general population on all scales except Physical Functioning. Bipolar patients had significantly higher scores than chronic back pain patients in the categories of Physical Functioning, Role Limitations--Physical, Bodily Pain, and Social Function. There were no significant differences between bipolar disorder and chronic back pain groups in the Mental Health and Role Limitations - Emotional categories. LIMITATIONS: The results of the study are limited by the relatively small sample sizes of the bipolar and back pain patient groups. CONCLUSIONS: Patients with bipolar disorder had substantial impairment in health related quality of life in comparison with the general population. Bipolar patients were less compromised in areas of physical and social functioning than chronic back pain patients but had similar impairment in mental health. PMID- 10708838 TI - Tridimensional personality questionnaire and treatment response in major depressive disorder: a negative study. AB - BACKGROUND: Recent studies have found that the Tridimensional Personality Questionnaire can be used to help predict antidepressant treatment response in depressed outpatients. As this finding could be of great clinical importance, we attempted to replicate these findings. METHODS: Our study included 199 outpatients with major depressive disorder in an 8-week open trial with fluoxetine 20 mg/day. The Tridimensional Personality Questionnaire (TPQ) was administered to all patients before treatment. RESULTS: There was a significant correlation between pre-treatment scores on the TPQ subscale of harm avoidance and severity of depression at baseline as determined by Hamilton Depression Rating Scale-17 (HAM-D-17) scores. There was no correlation of harm avoidance scores and percent improvement of HAM-D-17 after treatment with fluoxetine. There was also no correlation of baseline HAM-D-17 scores or percent improvement with the subscales of reward dependence and novelty seeking. LIMITATIONS: Our study's limitations include a possible selection bias, lack of controls and fixed dosing of fluoxetine. CONCLUSIONS: In contrast to previous studies, we failed to find a relationship between temperament type as defined by the TPQ and antidepressant response. Our failure to replicate the findings of other studies may in large part be related to the use of different classes of antidepressants. Further studies using similar antidepressants may be helpful to clarify this discrepancy. PMID- 10708839 TI - Mexiletine in treatment-resistant bipolar disorder. AB - BACKGROUND: The purpose of this study was to evaluate the efficacy and safety of mexiletine, a medication with antiarrhythmic, anticonvulsant and analgesic properties, in treatment-resistant bipolar disorder patients. METHODS: Twenty subjects with rapid-cycling bipolar disorder who had failed to respond or were intolerant to lithium, valproic acid and carbamazepine were entered into the 6 week, open label study. Subjects were followed on a weekly basis for dosing of mexiletine, blood levels, and completion of the Hamilton Depression Rating Scale (HAM-D) and the Manic State Rating Scale (MSRS). "Burden of Mood Symptoms" (BMS) was calculated by combining scores for the HAM-D and MSRS. RESULTS: Thirteen subjects (10 female, 3 male), mean age 41 years (S.D.=7.6), and mean duration of illness 20 years (S.D.=7.7) completed the study. The dose range of mexiletine was 200-1200 mg/day. Full response (>/=50% reduction in BMS) was seen in 46% of the subjects, and a partial response (25-49% reduction in BMS) in 15%. Of note, 5/5 subjects with a mixed or manic state demonstrated a full or partial response. LIMITATIONS: This study has an open label design, and a small number of subjects. CONCLUSIONS: Mexiletine may be effective and safe in patients with highly treatment-resistant, chronic bipolar disorder. Randomized, controlled trials are required to confirm the current results. PMID- 10708840 TI - Antidepressant efficacy of Sudarshan Kriya Yoga (SKY) in melancholia: a randomized comparison with electroconvulsive therapy (ECT) and imipramine. AB - BACKGROUND: Sudarshan Kriya Yoga (SKY) is a procedure that involves essentially rhythmic hyperventilation at different rates of breathing. The antidepressant efficacy of SKY was demonstrated in dysthymia in a prospective, open clinical trial. This study compared the relative antidepressant efficacy of SKY in melancholia with two of the current standard treatments, electroconvulsive therapy (ECT) and imipramine (IMN). METHODS: Consenting, untreated melancholic depressives (n=45) were hospitalized and randomized equally into three treatment groups. They were assessed at recruitment and weekly thereafter for four weeks. RESULTS: Significant reductions in the total scores on Beck Depression Inventory (BDI) and Hamilton Rating Scale for Depression (HRSD) occurred on successive occasions in all three groups. The groups, however, did not differ. Significant interaction between the groups and occasion of assessment occurred. At week three, the SKY group had higher scores than the ECT group but was not different from the IMN group. Remission (total HRSD score of seven or less) rates at the end of the trial were 93, 73 and 67% in the ECT, IMN and SKY groups, respectively. No clinically significant side effects were observed. DISCUSSION: Within the limitations of the design (lack of double blind conditions), it can be concluded that, although inferior to ECT, SKY can be a potential alternative to drugs in melancholia as a first line treatment. PMID- 10708841 TI - The use of the Beck Depression Inventory to screen for depression in the general population: a preliminary analysis. AB - OBJECTIVE: The aim of the present paper is to study the performance of Beck's Depression Inventory (BDI) as a screening instrument for depressive disorders in a general population sample. METHODS: 1250 subjects, from 18 to 64 years old, were randomly selected from the Santander (Spain) municipal census. A two-stage method was used: in the first stage, all individuals selected completed the BDI; in the second, 'probable cases' (BDI cut-off>/=13) and a random 5% sample of the total sample with a BDI score less than 13 were interviewed by psychiatrists using the Schedules for Clinical Assessment in Neuropsychiatry (SCAN), which generates diagnoses of depressive disorders. RESULTS: Our data confirm the predictive value of the selected cut-off point (12/13): 100% sensitivity, 99% specificity, 0. 72 PPV, 1 NPV, and 98% overall diagnostic value. The area under ROC (AUC) was found to be 0.99. There were no statistical differences in terms of sex or age. We conclude that the BDI is a good instrument for screening depressive disorders in community surveys. PMID- 10708843 TI - Special issue in honor of hans Neurath's 90th birthday PMID- 10708842 TI - Psychiatric features of individuals with problematic internet use. AB - BACKGROUND: Problematic internet use has been described in the psychological literature as 'internet addiction' and 'pathological internet use'. However, there are no studies using face-to-face standardized psychiatric evaluations to identify behavioral characteristics, psychiatric comorbidity or family psychiatric history of individuals with this behavior. METHODS: Twenty individuals with problematic internet use were evaluated. Problematic internet use was defined as (1) uncontrollable, (2) markedly distressing, time-consuming or resulting in social, occupational or financial difficulties and (3) not solely present during hypomanic or manic symptoms. Evaluations included a semistructured interview about subjects' internet use, the Structured Clinical Interview for Diagnostic and Statistical Manual of Mental Disorders-IV (SCID-IV), family psychiatric history and the Yale-Brown Obsessive-Compulsive Scale (Y-BOCS) modified for internet use. RESULTS: All (100%) subjects' problematic internet use met DSM-IV criteria for an impulse control disorder (ICD) not otherwise specified (NOS). All 20 subjects had at least one lifetime DSM-IV Axis I diagnosis in addition to their problematic internet use (mean+/-SD=5.1+/-3.5 diagnoses); 14 (70.0%) had a lifetime diagnosis of bipolar disorder (with 12 having bipolar I disorder). LIMITATIONS: Methodological limitations of this study included its small sample size, evaluation of psychiatric diagnoses by unblinded investigators, and lack of a control group. CONCLUSIONS: Problematic internet use may be associated with subjective distress, functional impairment and Axis I psychiatric disorders. PMID- 10708844 TI - Biography. Hans Neurath. PMID- 10708845 TI - From colloids to proteases. AB - The present autobiographical review describes my professional experiences as a graduate student in Vienna, Austria, the postdoctoral experiences at the University of London, University of Minnesota, and at Cornell University, Ithaca, NY. This was followed by a faculty appointment at Duke University where I rose through the ranks from assistant professor to professor of physical biochemistry from 1938 to 1950. This account includes both scientific and cultural episodes and anecdotes. In 1950 I moved to Seattle to become founding chairman and professor in the Department of Biochemistry as will be described elsewhere. PMID- 10708846 TI - Structural and biochemical studies of retroviral proteases. AB - Retroviral proteases form a unique subclass of the family of aspartic proteases. These homodimeric enzymes from a number of viral sources have by now been extensively characterized, both structurally and biochemically. The importance of such knowledge to the development of new drugs against AIDS has been, to a large extent, the driving force behind this progress. High-resolution structures are now available for enzymes from human immunodeficiency virus types 1 and 2, simian immunodeficiency virus, feline immunodeficiency virus, Rous sarcoma virus, and equine infectious anemia virus. In this review, structural and biochemical data for retroviral proteases are compared in order to analyze the similarities and differences between the enzymes from different sources and to enhance our understanding of their properties. PMID- 10708847 TI - Bacterial proteinases as targets for the development of second-generation antibiotics. AB - The emergence of bacterial pathogen resistance to common antibiotics strongly supports the necessity to develop alternative mechanisms for combating drug resistant forms of these infective organisms. Currently, few pharmaceutical companies have attempted to investigate the possibility of interrupting metabolic pathways other than those that are known to be involved in cell wall biosynthesis. In this review, we describe multiple, novel roles for bacterial proteinases during infection using, as a specific example, the enzymes from the organism Porphyromonas gingivalis, a periodontopathogen, which is known to be involved in the development and progression of periodontal disease. In this manner, we are able to justify the concept of developing synthetic inhibitors against members of this class of enzymes as potential second-generation antibiotics. Such compounds could not only prove valuable in retarding the growth and proliferation of bacterial pathogens but also lead to the use of this class of inhibitors against invasion by other infective organisms. PMID- 10708849 TI - Leucaena leucocephala serine proteinase inhibitor: primary structure and action on blood coagulation, kinin release and rat paw edema. AB - A serine proteinase inhibitor isolated from Leucaena leucocephala seeds (LlTI) was purified to homogeneity by acetone fractionation, ion exchange chromatography, gel filtration and reverse phase chromatography (HPLC). SDS-PAGE indicated a protein with M(r) 20000 and two polypeptide chains (alpha-chain, M(r) 15000, and beta-chain, M(r) 5000), the sequence being determined by automatic Edman degradation and by mass spectroscopy. LlTI is a 174 amino acid residue protein which shows high homology to plant Kunitz inhibitors, especially those double chain proteins purified from the Mimosoideae subfamily. LlTI inhibits plasmin (K(i) 3.2 x 10(-10) M), human plasma kallikrein (K(i) 6.3 x 10(-9) M), trypsin (K(i) 2.5 x 10(-8) M) and chymotrypsin (K(i) 1.4 x 10(-8) M). Factor XIIa activity is inhibited but K(i) was not determined, and factor Xa, tissue kallikrein and thrombin are not inhibited by LlTI. The action of LlTI on enzymes that participate in the blood clotting extrinsic pathway is confirmed by the prolongation of activated partial thromboplastin time, used as clotting time assay. The inhibition of the fibrinolytic activity of plasmin was confirmed on the hydrolysis of fibrin plates. LlTI inhibits kinin release from high molecular weight kininogen by human plasma kallikrein in vitro and, administered intravenously, causes a decrease in paw edema induced by carrageenin or heat in male Wistar rats. In addition, lower concentrations of bradykinin were found in limb perfusion fluids of LlTI-treated rats. PMID- 10708848 TI - Calcium-binding EGF-like modules in coagulation proteinases: function of the calcium ion in module interactions. AB - Epidermal growth factor (EGF)-like modules are involved in protein-protein interactions and are found in numerous extracellular proteins and membrane proteins. Among these proteins are enzymes involved in blood coagulation, fibrinolysis and the complement system as well as matrix proteins and cell surface receptors such as the EGF precursor, the low density lipoprotein receptor and the developmentally important receptor, Notch. The coagulation enzymes, factors VII, IX and X and protein C, all have two EGF-like modules, whereas the cofactor of activated protein C, protein S, has four EGF-like modules in tandem. Certain of the cell surface receptors have numerous EGF modules in tandem. A subset of EGF modules bind one Ca(2+). The Ca(2+)-binding sequence motif is coupled to a sequence motif that brings about beta-hydroxylation of a particular Asp/Asn residue. Ca(2+)-binding to an EGF module is important to orient neighboring modules relative to each other in a manner that is required for biological activity. The Ca(2+) affinity of an EGF module is often influenced by its N-terminal neighbor, be it another EGF module or a module of another type. This can result in an increase in Ca(2+) affinity of several orders of magnitude. Point mutations in EGF modules that involve amino acids which are Ca(2+) ligands result in the biosynthesis of biologically inactive proteins. Such mutations have been identified, for instance, in factor IX, causing hemophilia B, in fibrillin, causing Marfan syndrome, and in the low density lipoprotein receptor, causing hypercholesterolemia. In this review the emphasis will be on the coagulation factors. PMID- 10708850 TI - The human mast cell tryptase tetramer: a fascinating riddle solved by structure. AB - Tryptases, the predominant proteins of human mast cells, have been implicated as pathogenetic mediators of allergic and inflammatory conditions, most notably asthma. Until recently, the fascinating properties that distinguish tryptases among the serine proteinases, particularly their activity as a heparin-stabilized tetramer, resistance to most proteinaceous inhibitors, and preference for peptidergic over macromolecular substrates presented a riddle. This review solves this riddle with the help of the crystal structure of the human beta(2)-tryptase tetramer, but also indicates controversies between the unique quaternary architecture and some experimental data. PMID- 10708851 TI - S-nitrosylated human alpha(1)-protease inhibitor. AB - Alpha(1)-protease inhibitor (alpha(1)PI) is an acute phase plasma protein, and possesses a single cysteine residue at position 232. A single cysteinyl sulfhydryl of human alpha(1)PI is found to be readily S-nitrosylated by nitric oxide (NO) in vitro without affecting the inhibitory capacity against bovine trypsin or elastase, a major target protease of alpha(1)PI in vivo. S-nitroso alpha(1)PI (S-NO-alpha(1)PI) was also formed by the reaction of alpha(1)PI with NO produced excessively by a murine macrophage cell line (RAW264 cells) upon infection with Salmonella typhimurium and in an ex vivo perfusion system of the liver obtained from lipopolysaccharide-treated rats. S-NO-alpha(1)PI (10(-9)-10( 6) M) induces a dose-dependent relaxation of the ring preparation of rabbit aorta. Also, S-NO-alpha(1)PI but not alpha(1)PI shows a potent inhibitory effect on platelet aggregation. Unprecedented observation is that S-NO-alpha(1)PI showed a potent bacteriostatic effect against a wide range of bacteria at the concentration of 1-10 microM, which was 10-1000-fold stronger than that of NO and other S-nitrosylated compounds including S-nitrosylated albumin and S nitrosylated glutathione. These results suggest that S-NO-alpha(1)PI is produced as an NO sink under inflammatory conditions, where production of both alpha(1)PI and NO is highly up-regulated, and it may function as a soluble factor which consists of an innate defense system through not only the protease inhibitory activity but also its antibacterial activity and facilitating the peripheral blood flow. Therefore, S-nitrosylation of alpha(1)PI occurring under physiological conditions in vivo should diversify the biological functions contributing to cytoprotective effects of alpha(1)PI. PMID- 10708852 TI - Lysosomal cysteine proteases: more than scavengers. AB - Lysosomal cysteine proteases were believed to be mainly involved in intracellular protein degradation. Under special conditions they have been found outside lysosomes resulting in pathological conditions. With the discovery of a series of new cathepsins with restricted tissue distributions, it has become evident that these enzymes must be involved in a range of specific cellular tasks much broader than as simple housekeeping enzymes. It is therefore timely to review and discuss the various physiological roles of mammalian lysosomal papain-like cysteine proteases as well as their mechanisms of action and the regulation of their activity. PMID- 10708853 TI - The systemin signaling pathway: differential activation of plant defensive genes. AB - Systemin, an 18-amino-acid polypeptide released from wound sites on tomato leaves caused by insects or other mechanical damage, systemically regulates the activation of over 20 defensive genes in tomato plants in response to herbivore and pathogen attacks. Systemin is processed from a larger prohormone protein, called prosystemin, by proteolytic cleavages. However, prosystemin lacks a signal sequence and glycosylation sites and is apparently not synthesized through the secretory pathway, but in the cytoplasm. The polypeptide activates a lipid-based signal transduction pathway in which the 18:3 fatty acid, linolenic acid, is released from plant membranes and converted to the oxylipin signaling molecule jasmonic acid. A wound-inducible systemin cell surface receptor with an M(r) of 160,000 has recently been identified. The receptor regulates an intracellular cascade including, depolarization of the plasma membrane, the opening of ion channels, an increase in intracellular Ca(2+), activation of a MAP kinase activity and a phospholipase A(2) activity. These rapid changes appear to play important roles leading to the intracellular release of linolenic acid from membranes and its subsequent conversion to jasmonic acid, a potent activator of defense gene transcription. Although the mechanisms for systemin processing, release, and transport are still unclear, studies of the timing of the synthesis and of the intracellular localization of wound- and systemin-inducible mRNAs and proteins indicates that differential syntheses of signal pathway genes and defensive genes are occurring in different cell types. This signaling cascade in plants exhibits extraordinary analogies with the signaling cascade for the inflammatory response in animals. PMID- 10708854 TI - Structure-function relationships of glutamine synthetases. AB - As a highly regulated enzyme at the core of nitrogen metabolism, glutamine synthetase has been studied intensively. We review structural and functional studies of both bacterial and eukaryotic glutamine synthetases, with emphasis on enzymatic inhibitors. PMID- 10708855 TI - Snake venom proteases affecting hemostasis and thrombosis. AB - The structure and function of snake venom proteases are briefly reviewed by putting the focus on their effects on hemostasis and thrombosis and comparing with their mammalian counterparts. Up to date, more than 150 different proteases have been isolated and about one third of them structurally characterized. Those proteases are classified into serine proteases and metalloproteinases. A number of the serine proteases show fibrin(ogen)olytic (thrombin-like) activities, which are not susceptible to hirudin or heparin and perhaps to most endogenous serine protease inhibitors, and form abnormal fibrin clots. Some of them have kininogenase (kallikrein-like) activity releasing hypotensive bradykinin. A few venom serine proteases specifically activate coagulation factor V, protein C, plasminogen or platelets. The venom metalloproteinases, belonging to the metzincin family, generally show fibrin(ogen)olytic and extracellular matrix degrading (hemorrhagic) activities. A few venom metalloproteinases show a unique substrate specificity toward coagulation factor X, platelet membrane receptors or von Willebrand factor. A number of the metalloproteinases have chimeric structures composed of several domains such as proteinase, disintegrin-like, Cys rich and lectin-like domains. The disintegrin-like domain seems to facilitate the action of those metalloproteinases by interacting with platelet receptors. A more detailed analysis of snake venom proteases should find their usefulness for the medical and pharmacological applications in the field of thrombosis and hemostasis. PMID- 10708856 TI - Structure and function of the methionine aminopeptidases. AB - The removal of the N-terminal methionine from proteins and peptides is dependent upon a novel class of proteases typified by the dinuclear metalloenzyme methionine aminopeptidase from Escherichia coli (eMetAP). Substantial progress has recently been made in determining the structures of several members of this family. The identification of human MetAP as the target of putative anti-cancer drugs reiterates the importance of this family of enzymes. Determination of the modes of binding to E. coli MetAP of a substrate-like bestatin-based inhibitor, as well as phosphorus-containing transition-state analogs and reaction products has led to a rationalization of the substrate specificity and suggested the presumed catalytic mechanism. The conservation of key active site residues and ligand interactions between the MetAPs and other enzyme of the same fold suggest that avoidance of cross-reactivity may be an important consideration in the design of inhibitors directed toward a single member of the family. PMID- 10708857 TI - HIV protease as a target for retrovirus vector-mediated gene therapy. AB - The dimeric aspartyl protease of HIV has been the subject of intense research for almost a decade. Knowledge of the substrate specificity and catalytic mechanism of this enzyme initially guided the development of several potent peptidomimetic small molecule inhibitors. More recently, the solution of the HIV protease structure led to the structure-based design of improved peptidomimetic and non peptidomimetic antiviral compounds. Despite the qualified success of these inhibitors, the high mutation rate associated with RNA viruses continues to hamper the long-term clinical efficacy of HIV protease inhibitors. The dimeric nature of the viral protease has been conducive to the investigation of dominant negative inhibitors of the enzyme. Some of these inhibitors are defective protease monomers that interact with functional monomers to form inactive protease heterodimers. An advantage of macromolecular inhibitors as compared to small-molecule inhibitors is the increased surface area of interaction between the inhibitor and the target gene product. Point mutations that preserve enzyme activity but confer resistance to small-molecule inhibitors are less likely to have an adverse effect on macromolecular interactions. The use of efficient retrovirus vectors has facilitated the delivery of these macromolecular inhibitors to primary human lymphocytes. The vector-transduced cells were less susceptible to HIV infection in vitro, and showed similar levels of protection compared to other macromolecular inhibitors of HIV replication, such as RevM10. These preliminary results encourage the further development of dominant-negative HIV protease inhibitors as a gene therapy-based antiviral strategy. PMID- 10708858 TI - Targeting the HIV-protease in AIDS therapy: a current clinical perspective. AB - This review deals with clinical applications of compounds that inhibit the action of the protease encoded within the genome of human immunodeficiency virus (HIV). The HIV-protease is essential for viral maturation and represents an important therapeutic target in the fight against AIDS. Following a brief overview of the enzyme structure and function, the article focuses on a number of peptide and non peptide based HIV-protease inhibitors that are in current clinical use. These drugs are discussed both with respect to their efficacy in treatment of AIDS, and to problems related to insurgence of viral resistance and side effects seen to date in patient populations. PMID- 10708859 TI - Cell surface complex of cathepsin B/annexin II tetramer in malignant progression. AB - The cysteine protease cathepsin B is upregulated in a variety of tumors, particularly at the invasive edges. Cathepsin B can degrade extracellular matrix proteins, such as collagen IV and laminin, and can activate the precursor form of urokinase plasminogen activator (uPA), perhaps thereby initiating an extracellular proteolytic cascade. Recently, we demonstrated that procathepsin B interacts with the annexin II heterotetramer (AIIt) on the surface of tumor cells. AIIt had previously been shown to interact with the serine proteases: plasminogen/plasmin and tissue-type plasminogen activator (tPA). The AIIt binding site for cathepsin B differs from that for either plasminogen/plasmin or tPA. AIIt also interacts with extracellular matrix proteins, e.g., collagen I and tenascin-C, forming a structural link between the tumor cell surface and the extracellular matrix. Interestingly, cathepsin B, plasminogen/plasmin, t-PA and tenascin-C have all been linked to tumor development. We speculate that colocalization through AIIt of proteases and their substrates on the tumor cell surface may facilitate: (1) activation of precursor forms of proteases and initiation of proteolytic cascades; and (2) selective degradation of extracellular matrix proteins. The recruitment of proteases to specific regions on the cell surface, regions where potential substrates are also bound, could well function as a 'proteolytic center' to enhance tumor cell detachment, invasion and motility. PMID- 10708860 TI - The two sides of enzyme-substrate specificity: lessons from the aspartic proteinases. AB - Like most proteolytic enzymes, the aspartic proteinases bind substrates and most inhibitors within an extended active site cleft. Bound ligands typically adopt a beta-strand conformation. Interactions with groups on both sides of the cleft determine the primary as well as secondary specificity of the enzymes. We have pursued the discovery of the sometimes subtle distinctions between members of the aspartic proteinase family by two routes. In the first case, we have constructed sets of oligopeptide substrates with systematic variation in each position to assess interactions at one position at a time. In the second type of experiment, we have altered residues of the enzymes in order to test theories of selectivity. The combination of the two approaches has provided a better understanding of the forces involved in determining specificity of enzyme action. PMID- 10708861 TI - Structural basis of the endoproteinase-protein inhibitor interaction. AB - Proteolytic enzymes are potentially hazardous to their protein environment, so that their activity must be carefully controlled. Living organisms use protein inhibitors as a major tool to regulate the proteolytic activity of proteinases. Most of the inhibitors for which 3D structures are available are directed towards serine proteinases, interacting with the active sites in a 'canonical' i.e. substrate-like manner via an exposed reactive site loop of conserved conformation. More recently, some non-canonically binding serine proteinase inhibitors directed against coagulation factors, in particular thrombin, a few cysteine proteinase inhibitors inhibitory towards papain-like proteinases, and three zinc endopeptidase inhibitors directed against metzincins and thermolysin have been characterised in the free and complexed state, displaying novel mechanisms of inhibition with their target proteinases. These different interaction modes are presented and briefly discussed with respect to the different strategies applied by nature. PMID- 10708862 TI - Nerve growth factor alpha subunit: effect of site-directed mutations on catalytic activity and 7S NGF complex formation. AB - Mouse alpha- and gamma-nerve growth factor (NGF) are glandular kallikreins that form a non-covalent complex (7S NGF) with beta-NGF. gamma-NGF is an active arginine-specific esteropeptidase; the alpha-subunit is catalytically inactive and has a zymogen-like conformation. Site-directed mutagenesis of alpha-NGF to alter the N-terminus and three residues in loop 7, a region that contributes to the catalytic center, restored substantial catalytic activity against N-benzoyl arginine-p-nitroanilide as substrate in two derivatives although they were not as active as recombinant gamma-NGF. Seven of the 15 derivatives that remained more alpha-like were able to substitute for native alpha-NGF in reforming 7S complexes; the other eight derivatives that were more gamma-like showed greatly reduced ability to do so. However, the most gamma-like alpha-NGF derivative could not substitute for native gamma-NGF in 7S complex formation. These findings suggest that the alpha-NGF backbone can be corrected to a functional enzyme by the addition of a normal N-terminal structure and two catalytic site substitutions and that the 7S complex requires one kallikrein subunit in the zymogen form and one in an active conformation. PMID- 10708863 TI - Tissue inhibitors of metalloproteinases: evolution, structure and function. AB - The matrix metalloproteinases (MMPs) play a key role in the normal physiology of connective tissue during development, morphogenesis and wound healing, but their unregulated activity has been implicated in numerous disease processes including arthritis, tumor cell metastasis and atherosclerosis. An important mechanism for the regulation of the activity of MMPs is via binding to a family of homologous proteins referred to as the tissue inhibitors of metalloproteinases (TIMP-1 to TIMP-4). The two-domain TIMPs are of relatively small size, yet have been found to exhibit several biochemical and physiological/biological functions, including inhibition of active MMPs, proMMP activation, cell growth promotion, matrix binding, inhibition of angiogenesis and the induction of apoptosis. Mutations in TIMP-3 are the cause of Sorsby's fundus dystrophy in humans, a disease that results in early onset macular degeneration. This review highlights the evolution of TIMPs, the recently elucidated high-resolution structures of TIMPs and their complexes with metalloproteinases, and the results of mutational and other studies of structure-function relationships that have enhanced our understanding of the mechanism and specificity of the inhibition of MMPs by TIMPs. Several intriguing questions, such as the basis of the multiple biological functions of TIMPs, the kinetics of TIMP-MMP interactions and the differences in binding in some TIMP-metalloproteinase pairs are discussed which, though not fully resolved, serve to illustrate the kind of issues that are important for a full understanding of the interactions between families of molecules. PMID- 10708864 TI - Metallocarboxypeptidases and their protein inhibitors. Structure, function and biomedical properties. AB - Among the different aspects of recent progress in the field of metallocarboxypeptidases has been the elucidation of the three dimensional structures of the pro-segments (in monomeric or oligomeric species) and their role in the expression, folding and inhibition/activation of the pancreatic and pancreatic-like forms. Also of great significance has been the cloning and characterization of several new regulatory carboxypeptidases, enzymes that are related with important functions in protein and peptide processing and that show significant structural differences among them and also with the digestive ones. Many regulatory carboxypeptidases lack a pro-region, unlike the digestive forms or others in between from the evolutionary point of view. Finally, important advances have been made on the finding and characterization of new protein inhibitors of metallocarboxypeptidases, some of them with interesting potential applications in the biotechnological/biomedical fields. These advances are analyzed here and compared with the earlier observations in this field, which was first explored by Hans Neurath and collaborators. PMID- 10708865 TI - Caspases - controlling intracellular signals by protease zymogen activation. AB - Animal development and homeostasis is a balance between cell proliferation and cell death. Physiologic, and sometimes pathologic, cell death - apoptosis - is driven by activation of a family of proteases known as the caspases, present in almost all nucleated animal cells. The enzymatic properties of these proteases are governed by a dominant specificity for substrates containing Asp, and by the use of a Cys side chain for catalyzing peptide bond cleavage. The primary specificity for Asp turns out to be very rare among proteases, and currently the only other known mammalian proteases with the same primary specificity is the physiological caspase activator granzyme B. Like most other proteases, the caspases are synthesized as inactive zymogens whose activation requires limited proteolysis or binding to co-factors. To transmit the apoptotic execution signal, caspase zymogens are sequentially activated through either an intrinsic or an extrinsic pathway. The activation of caspases at the apex of each pathway, the initiators, occurs by recruitment to specific adapter molecules through homophilic interaction domains, and the activated initiators directly process the executioner caspases to their catalytically active forms. In the present communication we review the different mechanisms underlying the selective activation of the caspases. PMID- 10708866 TI - Granzymes (lymphocyte serine proteases): characterization with natural and synthetic substrates and inhibitors. AB - Natural killer (NK) and cytotoxic T-lymphocytes (CTLs) kill cells within an organism to defend it against viral infections and the growth of tumors. One mechanism of killing involves exocytosis of lymphocyte granules which causes pores to form in the membranes of the attacked cells, fragments nuclear DNA and leads to cell death. The cytotoxic granules contain perforin, a pore-forming protein, and a family of at least 11 serine proteases termed granzymes. Both perforin and granzymes are involved in the lytic activity. Although the biological functions of most granzymes remain to be resolved, granzyme B clearly promotes DNA fragmentation and is directly involved in cell death. Potential natural substrates for Gr B include procaspases and other proteins involved in cell death. Activated caspases are involved in apoptosis. The search continues for natural substrates for the other granzymes. The first granzyme crystal structure remains to be resolved, but in the interim, molecular models of granzymes have provided valuable structural information about their substrate binding sites. The information has been useful to predict the amino acid sequences that immediately flank each side of the scissile peptide bond of peptide and protein substrates. Synthetic substrates, such as peptide thioesters, nitroanilides and aminomethylcoumarins, have also been used to study the substrate specificity of granzymes. The different granzymes have one of four primary substrate specificities: tryptase (cleaving after Arg or Lys), Asp-ase (cleaving after Asp), Met-ase (cleaving after Met or Leu), and chymase (cleaving after Phe, Tyr, or Trp). Natural serpins and synthetic inhibitors (including isocoumarins, peptide chloromethyl ketones, and peptide phosphonates) inhibit granzymes. Studies of substrate and inhibitor kinetics are providing valuable information to identify the most likely natural granzyme substrates and provide tools for the study of key reactions in the cytolytic mechanism. PMID- 10708867 TI - What can the structures of enzyme-inhibitor complexes tell us about the structures of enzyme substrate complexes? AB - Proteinases perform many beneficial functions that are essential to life, but they are also dangerous and must be controlled. Here we focus on one of the control mechanisms: the ubiquitous presence of protein proteinase inhibitors. We deal only with a subset of these: the standard mechanism, canonical protein inhibitors of serine proteinases. Each of the inhibitory domains of such inhibitors has one reactive site peptide bond, which serves all the cognate enzymes as a substrate. The reactive site peptide bond is in a combining loop which has an identical conformation in all inhibitors and in all enzyme-inhibitor complexes. There are at least 18 families of such inhibitors. They all share the conformation of the combining loops but each has its own global three-dimensional structure. Many three-dimensional structures of enzyme-inhibitor complexes were determined. They are frequently used to predict the conformation of substrates in very short-lived enzyme-substrate transition state complexes. Turkey ovomucoid third domain and eglin c have a Leu residue at P(1). In complexes with chymotrypsin, these P(1) Leu residues assume the same conformation. The relative free energies of binding of P(1) Leu (relative to either P(1) Gly or P(1) Ala) are within experimental error, the same for complexes of turkey ovomucoid third domain, eglin c, P(1) Leu variant of bovine pancreatic trypsin inhibitor and of a substrate with chymotrypsin. Therefore, the P(1) Leu conformation in transition state complexes is predictable. In contrast, the conformation of P(1) Lys(+) is strikingly different in the complexes of Lys(18) turkey ovomucoid third domain and of bovine pancreatic trypsin inhibitor with chymotrypsin. The relative free energies of binding are also quite different. Yet, the relative free energies of binding are nearly identical for Lys(+) in turkey ovomucoid third domain and in a substrate, thus allowing us to know the structure of the latter. Similar reasoning is applied to a few other systems. PMID- 10708869 TI - Regulation of blood coagulation. AB - The protein C anticoagulant pathway converts the coagulation signal generated by thrombin into an anticoagulant response through the activation of protein C by the thrombin-thrombomodulin (TM) complex. The activated protein C (APC) thus formed interacts with protein S to inactivate two critical coagulation cofactors, factors Va and VIIIa, thereby dampening further thrombin generation. The proposed mechanisms by which TM switches the specificity of thrombin include conformational changes in thrombin, blocking access of normal substrates to thrombin and providing a binding site for protein C. The function of protein S appears to be to alter the cleavage site preferences of APC in factor Va, probably by changing the distance of the active site of APC relative to the membrane surface. The clinical relevance of this pathway is now established through the identification of deficient individuals with severe thrombotic complications and through the analysis of families with partial deficiencies in these components and an increased thrombotic tendency. One possible reason that even partial deficiencies are a thrombotic risk is that the function of the pathway can be down-regulated by inflammatory mediators. For instance, clinical studies have shown that the extent to which protein C levels decrease in patients with septic shock is predictive of a negative outcome. Initial clinical studies suggest that supplementation with protein C may be useful in the treatment of acute inflammatory diseases such as sepsis. PMID- 10708868 TI - Evolution of the prohormone convertases: identification of a homologue of PC6 in the protochordate amphioxus. AB - Many of the protein precursors traversing the secretory pathway undergo cleavage at multibasic sites to generate their bioactive forms. The proprotein convertases (PCs), a family of subtilisin-like proteases, are the major endoproteases that serve this function. Genes encoding seven distinct members of this family have so far been characterized in vertebrates: furin, PC2, PC1/PC3, PC4, PACE4, PC5/PC6 and PC7/PC8/LPC. Multiple PC genes have also been cloned from a number of invertebrates, including Drosophila melanogaster and Caenorhabditis elegans. These findings suggest that gene duplication and diversification of the PCs have occurred throughout metazoan evolution. To investigate the structural and functional changes which have occurred during vertebrate development, we have analyzed the expression of PC genes in the protochordate amphioxus. We have previously shown that amphioxus express homologous PC2 and PC1/PC3 genes [Proc. Natl. Acad. Sci. USA 92 (1995) 3591]. Here we report the characterization of amphioxus cDNAs encoding proteases with a high degree of similarity to mammalian PC6. Three cDNAs encoding three PC6 isoforms differing only in their carboxy terminal sequences were found, derived by alternative splicing. Two isoforms appear to be soluble enzymes, whereas the third contains a transmembrane hydrophobic segment and thus is likely to be membrane-bound. All three variants contain many repeats of a cysteine-rich motif that is found in several other PC family members. Thus, amphioxus, like the vertebrates, expresses two types of PCs, e.g., PC2 and PC1/PC3 which function in the regulated secretory pathway in neuroendocrine cells, and the more widely expressed PC6 which functions mainly in the constitutive pathway. PMID- 10708873 TI - Oral dosage forms fabricated by three dimensional printing. AB - Three Dimensional Printing is a novel technique used in the fabrication of complex oral dosage delivery pharmaceuticals. It is possible to engineer devices with complicated internal geometries, varying densities and diffusivities, and multiple actives and excipients. Samples were fabricated using this technique using standard pharmaceutical materials. Erosion mechanism delayed-release tablets were constructed with varying polymer content from 8.9 to 17. 9%. Lag times varied between 25 and 50 min with a corresponding decrease in release rate as polymer content increased. Diffusion mechanism tablets were constructed with varying polymer content from 9.0 to 16.7%. The peak release rate decreased and the time to exhaustion increased with polymer content, whereas lag time was not affected. Active delivery studies with fluorescein indicated that Three Dimensional Printing is capable of accurately constructing dosage forms with active content as low as 10(-12) moles per tablet. Hardness and friability testing indicated that samples fabricated with this technique are comparable to other standard pharmaceutical products. PMID- 10708874 TI - Multimechanism oral dosage forms fabricated by three dimensional printing. AB - Four types of complex oral drug delivery devices have been fabricated using the three dimensional printing process. Immediate-extended release tablets were fabricated which were composed of two drug-containing sections of different pH based release mechanisms. Pulsed release of chlorpheniramine maleate occurred after a lag time of 10 min followed by extended release of the compound over a period of 7 h. Breakaway tablets were fabricated composed of three sections. An interior fast-eroding section separating two drug-releasing sub-units eroded in 30-45 min in simulated gastric fluid. Enteric dual pulsatory tablets were constructed of one continuous enteric excipient phase into which diclofenac sodium was printed into two separated areas. These samples showed two pulses of release during in vitro USP dissolution at 1 and 8 h with a lag time between pulses of about 4 h. Dual pulsatory tablets were also fabricated. These samples were composed of two erosion based excipient sections of opposite pH based solubility. One section eroded immediately during the acid dissolution stage releasing diclofenac during the first 30 min, and the second section began eroding 5 h later during the high pH stage. PMID- 10708875 TI - Drug release from poly(acrylic acid) grafted poly(vinylidene fluoride) membrane bags in the gastrointestinal tract in the rat and dog. AB - Stomach-specific drug delivery systems would be of value in treating diseases of the upper gastrointestinal tract. The present study measured in vitro and in vivo drug release from pH-sensitive membrane bags, constructed of poly(acrylic acid) grafted onto a poly(vinylidene fluoride) (PAA-PVDF) membrane, which might be suitable for stomach-specific drug delivery. The used model drugs were propranolol-HCl (1.0 mg) and FITC-dextran MW 4400 (1.0 mg). Drug release in vivo was studied by inserting membrane bags into the stomach and proximal duodenum of anesthetized rats and dogs. At 30 and 180 min, the bags were removed from the lumens and residual drug content was determined. The release of either propranolol or FITC-dextran were comparable in both stomach and duodenum, showing that in vivo drug release did not depend on environmental pH. In vitro results suggested that these results could be explained by interactions between PAA and the mucous layers of the stomach and duodenum. PMID- 10708876 TI - First order release rate from porous PLA microspheres with limited exit holes on the exterior surface. AB - We applied the finite element method (FEM) to calculate release profiles from computer simulated slabs, one with a limited number of exit holes on the exterior surface, and the other with uniform structure. The former slab showed a first order release rate, and a nearly uniform drug concentration distribution within the device during the release process. It was concluded that circulation of the drug molecules within the slab resulted in the uniform concentration and consequently first order release rate. This theoretical work was used to explain the first order release rate of an active ingredient (flourescin-4-isothiocyanate dextran, M(W)=71000 Da) from porous PLA (poly(D,L)-lactic acid) microspheres, which by canning electron microscopy (SEM) examination showed only a few exit holes on their exterior surface. Calculations indicated that the internal surface adsorption of the active ingredient, or the pore size distribution of the microspheres, could not influence the mechanism for the first order release rate, and the small number of exit holes on the exterior surface was likely to be the rate-determining factor. The exit holes could be observed by SEM and their size and number is consistent with our interpretations. PMID- 10708877 TI - Development of controlled drug release systems based on thiolated polymers. AB - The purpose of the present study was to generate mucoadhesive matrix-tablets based on thiolated polymers. Mediated by a carbodiimide, L-cysteine was thereby covalently linked to polycarbophil (PCP) and sodium carboxymethylcellulose (CMC). The resulting thiolated polymers displayed 100+/-8 and 1280+/-84 micromol thiol groups per gram, respectively (means+/-S.D.; n=6-8). In aqueous solutions these modified polymers were capable of forming inter- and/or intramolecular disulfide bonds. The velocity of this process augmented with increase of the polymer- and decrease of the proton-concentration. The oxidation proceeded more rapidly within thiolated PCP than within thiolated CMC. Due to the formation of disulfide bonds within thiol-containing polymers, the stability of matrix-tablets based on such polymers could be strongly improved. Whereas tablets based on the corresponding unmodified polymer disintegrated within 2 h, the swollen carrier matrix of thiolated CMC and PCP remained stable for 6.2 h (mean, n=4) and more than 48 h, respectively. Release studies of the model drug rifampicin demonstrated that a controlled release can be provided by thiolated polymer tablets. The combination of high stability, controlled drug release and mucoadhesive properties renders matrix-tablets based on thiolated polymers useful as novel drug delivery systems. PMID- 10708878 TI - Development of new drug delivery system for protein drugs using silicone (I). AB - A novel technique, by which protein drugs effective in small doses can be released over a long period, was developed using silicone and a water-soluble substance. In this study, interferon (IFN) was used as a model of the protein drugs. The IFN-silicone formulation released IFN over long periods of time in vitro and suppressed tumor growth in nude mice for about 100 days after a single administration. This indicates that physiologically active IFN is released over a prolonged period of time from the IFN-silicone formulation in vivo. Silicone formulations are expected to be a practically feasible sustained-release formulation. PMID- 10708879 TI - Ion pair skin transport of a zwitterionic drug, cephalexin. AB - The ion pair skin transport of cephalexin was investigated using various counter ions and solvents. The permeability of cephalexin was enhanced by 1 alkylsulfonates (ASs) at pH 3.0 and by tetraalkylammoniums (AAs) at pH 7.0; the enhancing ratio increased with the number of carbon atoms in their alkyl chains. The corresponding effects of these additives were observed on the partitioning of cephalexin. Most of the additives did not affect the skin transport of D-mannitol and cortisone. These results suggest that the enhanced transport of cephalexin results from the ion pair formation with additives. Although ASs increased the partitioning of cephalexin above that of AAs, the transport enhancement effect of ASs was lower than AAs having the same number of carbon atoms in their alkyl chains, indicating higher diffusivity of the ion pairs with AAs in skin. Moreover, the transport enhancement by AAs increased even more when ethanol buffer solutions were used as solvents. The conductivity measurement of dissolving solutes in donor solvents showed that the further enhancement might be caused by the increasing ion pair formation in solvents with low dielectric constants. To obtain the maximum enhancement of skin transport of zwitterionic drugs via ion pair concept, one should select a counter ion having high lipophilicity and small volume, and a solvent with suitable pH and low dielectric constant. PMID- 10708880 TI - Prolonged delivery of nicotine in rats via nasal administration of proliposomes. AB - In order to achieve a prolonged delivery of nicotine to the systemic circulation, proliposomes containing nicotine base (NB-proliposomes) or nicotine hydrogen tartarate salt (NS-proliposomes) and a mixture of powdered nicotine hydrogen tartarate salt and sorbitol (1:9 mixture, MP) were administered intranasally to rats at a nicotine dose of 1 mg/kg. Proliposomes, lipid-sorbitol mixtures that form liposomes upon contact with water, were prepared according to previously established methods, and the mixture (MP) was prepared by mixing NS powder with sorbitol particles (105-350 micrometer in size). Nasal absorption of nicotine from these formulations was very rapid (i.e. less than 10 min was required to reach plasma peaks) and showed substantially sustained plasma nicotine levels compared to saline solutions of NB and NS, and previously reported nasal nicotine sprays. The AUC values from the proliposomes and MP were comparable to those from the saline solutions of NB and NS. However, the mean residence time (MRT) and plasma half-life (T(1/2beta)) of nicotine in the present study were much larger than those from the saline solutions. Thus, a prolonged delivery of nicotine to systemic circulation via the application of proliposomes or MP intranasally appears feasible. NB-proliposomes exhibited the best characteristics in terms of the area under the plasma concentration (AUC), MRT and T(1/2beta) of nicotine, which was followed by NS-proliposomes and MP. Retarded conversion of proliposomes and MP to liposomal emulsions and solution in the nasal cavity seems responsible, in part, for the sustained plasma nicotine concentrations, since the emulsions and solution yielded very short MRT and T(1/2beta) of nicotine. In addition, reduced metabolism to cotinine from the proliposomes and MP was apparently responsible for the sustained plasma nicotine levels. These dosage forms of nicotine appear to circumvent some of the shortcomings of transdermal patches (i.e. long T(max)) and nasal sprays (i.e. short T(1/2beta) and physicochemical instability). PMID- 10708881 TI - NMR microscopy of the uptake, distribution and mobility of dissolution media in small, sub-millimetre drug delivery systems. AB - NMR techniques were used to study the drug release process from small (sub-mm) lipophilic matrix theophylline beads. NMR microscopy was used to follow the ingress of the dissolution medium into the beads. Pulsed gradient spin echo (PGSE) NMR and 3D NMR imaging were used to measure the mobility and distribution of liquid within fully liquid penetrated beads. These techniques were used to rationalise the increase in the drug release rate with increasing proportion of GMS (glycomonosaccharide):paraffin in the matrix composition. A well-defined penetrant front was seen to move towards the centre of the bead during the dissolution process and the rate of liquid uptake showed the same trend with increasing GMS content as the rate of drug release. The total concentration of absorbed liquid increased and its T(2) relaxation time decreased with increasing GMS content of the bead matrix. This combined with the interpretation of PGSE results suggested that liquid resides in the matrix material as well as in pores left by the dissolved drug, and that this tendency increases with increasing GMS content. Heterogeneities in liquid distribution within the beads were quantified using percolation analysis and were related to the lipid matrix composition and may be a contributory factor in determining the rate of drug release. PMID- 10708882 TI - Generation of antibodies directed against the low-immunogenic peptide-toxins microcystin-LR/RR and nodularin. AB - The preparation of antibodies against the liver toxin microcystin, as described here, is of major importance for its detection and purification in food and water, and for a therapeutic approach to neutralize the toxin by passive immunization. Microcystin-LR (MLR) and microcystin-RR (MRR) were purified from cyanobacterial cell materials by extraction, Sephadex LH-20-, ODS silica gel-, ionic exchange and RP-HPLC-chromatography. In order to reduce the toxicity for parenteral administration, microcystins were coupled by the carbodiimide method to poly-L-lysine (PLL(50.000)). Mice and rabbits were immunized with the conjugates in the presence of two lipopeptide immunoadjuvants (P(3)CSK(4) and P(3)CS-T(h)). High MLR-specific antibody levels were observed after parenteral coadministration of antigen and lipopeptides, whereas no anti-MLR antibodies were obtained with free microcystin or the microcystin-PLL(50.000)-conjugate in the absence of lipopeptide. In oral immunization, coadministration of antigen and adjuvants resulted in an accelerated development of anti MLR-specific antibodies and high antibody levels. Using the antisera, we could detect different microcystins and nodularin down to a concentration range of 10-50 ng/ml by a competitive inhibition ELISA; detection of microcystins in crude cell preparations was also possible. Furthermore, microcystins from different sources could be detected and discriminated from cyclic cyanopeptolines. PMID- 10708883 TI - Effect of Salmonella typhi wild type and O-antigen mutants on human natural killer cell activity. AB - We investigated the effect of glutaraldehyde-fixed Salmonella typhi Ty2 (Vi(-)) wild-type (World Health Organization's vaccine strain) and mutant strains MEI028 (rough, O-antigen(-)) and MEI012 [smooth (O-antigen(+)95%), immunomagnetically isolated NK cell preparations. Incubation of PBMC with each and every one of the S. typhi strains studied consistently and significantly, increased this cellular immune function, as well as the supernatant level of the various cytokines tested e.g. IFN-gamma, TNF-alpha, IL-10 and IL-12 (ELISA). In similar experiments, a significant increase in the cytolytic activity of HPNK cells was elicited by S. typhi Ty2 but not by mutant strain MEI028; neither of the cytokines assayed (IFN gamma and TNF-alpha) was detected in the supernatant. Our results suggest that S. typhi O-antigen plays an essential role in a mechanism resulting in the direct activation of NK cell activity in HPNK cell preparations. However, the relative quantitative significance of this antigen in the direct stimulation of NK cell cytotoxicity expression in PBMC samples is less clear, as it appears that in this case bacterial-induced monocyte-released cytokines plays a most important role. Incubation with S. typhi Ty2 or MEI028 elicited significant expression of CD69, an early marker of NK cell activation, in PBMC but not in HPNK cell samples (flow cytometry); in similar experiments, the expression of CD16/56 and activation marker CD25 remained essentially unchanged. PMID- 10708884 TI - In vivo macrophage activation in chickens with Acemannan, a complex carbohydrate extracted from Aloe vera. AB - Acemannan (ACM 1), a beta-(1,4) -acetylated mannan isolated from Aloe vera, can be used as an effective adjuvant in vaccination against some avian viral diseases. Our results demonstrate a quick and lasting in vivo priming effect of ACM 1 on macrophage response after intramuscular inoculation in chickens (500 microg per 2-month-old bird). In response to IFN-gamma in vitro, monocytes from ACM 1-treated chickens exhibited a strong enhancement of NO production from 3 to 9 days p.i., but a weaker effect on MHC II cell surface antigen expression on day 3 p.i. A stimulating effect of ACM 1 treatment was also observed on spontaneous and inducible NO production for splenocytes only on day 3 p.i. By that time, splenocytes exhibited a strong higher capacity to proliferate in response to the T cell-mitogen PHA. At the same time, the in vivo capacity to produce NO, measured by the (NO(-)(2)+NO(-)(3)) serum level after intravenous LPS injection, increased greatly from 3 to 9 days p.i. In conclusion, ACM 1 was able efficiently and durably to increase the activation capacity of macrophages from the systemic immune compartment (in particular from the blood and spleen after an intramuscular injection) in chickens, especially for NO production. These findings provide a better understanding of the adjuvant activity of ACM 1 for viral and tumoral diseases. PMID- 10708885 TI - Delta(9)-tetrahydrocannabinol selectively increases aspartyl cathepsin D proteolytic activity and impairs lysozyme processing by macrophages. AB - Delta(9)-tetrahydrocannabinol (THC) causes an antigen-dependent defect in the ability of macrophages to activate helper T cells, and this drug-induced impairment is mediated through the peripheral CB2 receptor. Various requirements for the processing of the antigen, lysozyme, were examined to determine where along the pathway THC exerts its influence. A THC-exposed macrophage hybridoma inefficiently stimulated interleukin-2 secretion by a helper T cell hybridoma in response to native lysozyme and its reduced form, suggesting that disulfide bond reduction was unaffected. Cell surface expression of major histocompatibility complex class II molecules was normal on THC-exposed macrophages. The drug exposed macrophages also competently presented a lysozyme peptide to the T cells, indicating that the class II molecules were functional. The proteolytic activity of two thiol cathepsins was unaltered, but aspartyl cathepsin D activity was significantly increased in THC-exposed macrophages. Thus, selective up-regulation of aspartyl cathepsin activity accompanied the deficiency in lysozyme processing and may contribute, at least in part, to the antigen-dependent processing defect in THC-exposed macrophages. PMID- 10708886 TI - Immunopharmacological and immunotoxicological activities of a water-soluble (1- >3)-beta-D-glucan, CSBG from Candida spp. AB - We have established a convenient, two-step procedure to solubilize the yeast cell wall (1-->3)-beta-D-glucan using the combination of NaClO oxidation and DMSO extraction. Candida soluble beta-D-glucan (CSBG) was mainly composed of a linear beta-1,3 glucan with a linear beta-1,6-glucan moiety. In this study, we screened for several immunopharmacological activities of CSBG and found the following activities: (1) interleukin-6 synthesis of macrophages in vitro; (2) antagonistic effect for zymosan mediated-tumor necrosis factor synthesis of macrophages; (3) augmentation for lipopolysaccharide mediated tumor necrosis factor and nitrogen oxide syntheses of macrophages; (4) activation of alternative pathway of complement; (5) hematopoietic response on cyclophosphamide induced leukopenia; (6) the antitumor effect on ascites form tumor; (7) Enhanced vascular permeability; (8) priming effect on lipopolysaccharide triggered TNF-alpha synthesis; and (9) adjuvant effect on antibody production. These results strongly suggested that CSBG possessed various immunopharmacological activity. PMID- 10708887 TI - Nitrogen contamination in the Yangtze river system, China. AB - The data at 570 monitoring stations during 1990 were studied. The results indicate as follows: (i) the contents of nitrogen in the Yangtze mainstream has a raising trend from the upper reaches to the lower reaches; (ii) total nitrogen content at a lot of stations during the middle 1980s is 5-10 times more than that during the 1960s; (iii) seasonal variances of nitrogen content vary with watersheds; and (iv) the difference of nitrogen contamination level is related to the regional population and economic development. PMID- 10708889 TI - Oxygen uptake rate inhibition with PACT sludge. AB - Oxygen uptake rate (OUR) experiments were performed with sludge from six laboratory-scale, continuously fed, activated sludge and PACT reactors (sludge ages of 4-, 8-, and 12-days) to evaluate the sludge's resistance to inhibitory compounds. Three inhibitory compounds with varied ability to sorb on activated carbon were tested: 3,5-dichlorophenol (3,5-DCP, strongly adsorbed), phenol (moderately adsorbed), and zinc (poorly adsorbed). The inhibitory compound concentration that reduced the unacclimated sludge's specific oxygen uptake 50% from its maximum rate was determined (IC(50)). For the organic compounds, PACT sludge resisted acute inhibition better for all sludge ages; sorption studies indicate that phenol sorbed onto the PACT sludge could account for the IC(50) difference at the higher sludge ages. With 3,5-DCP, the 4- and 8-day-old PACT and activated sludge solids sorbed similar amounts of 3,5-DCP at concentrations near the IC(50) values, yet the PACT sludge exhibited higher IC(50) values; biomass differences may have accounted for the improved resistance to inhibition. With the poorly adsorbed zinc, no difference in IC(50) or sorption was noted with the 4-day-old sludge. For the 12-day-old sludge, the PACT sludge was much more resistant to zinc exposure, with changes in the biomass rather than sorption on carbon the apparent reason. PMID- 10708888 TI - Assessing risk communication effectiveness: perspectives of agency practitioners. AB - A study conducted by the Agency for Toxic Substances and Disease Registry (ATSDR), a US public health agency, evaluated ATSDR's risk communication process, specifically the roles and responsibilities, planning, implementation, and coordination of activities in response to illegal indoor spraying of methyl parathion, a hazardous pesticide, in Pascagoula, MS. Interviews of staff members involved in the intervention were conducted and an analysis revealed strengths and areas in need of improvement in the design and implementation of risk communication strategies. Key recommendations included developing a clear strategy for planning and conducting communication activities; determining staff roles and responsibilities for coordination; and developing clear and consistent health messages, a dissemination strategy, and training in the delivery and evaluation of messages, effects, and outcomes. PMID- 10708890 TI - Experimental investigation into the incineration of wool scouring sludges in a novel rotating fluidised bed. AB - The main purpose of this research was to investigate the possibility of incineration of wool scouring sludges in a novel vertical axis rotating fluidized bed (RFB). A small-scale RFB was designed and constructed with an internal diameter (ID) of 200 mm and height of 50 mm to carry out the experiments. In phase one of the experiments, a cold test was conducted to investigate the fluidization performance of the RFB, which eventually led to the optimisation of the operating parameters, i.e., sand particle size, rotation speed and bed loading (bed thickness) which ensures complete fluidization and minimum particle elutriation. Sand particle size of 0.5 to 0.6 mm, rotation speed of 200 to 400 rpm and bed loading of 1 kg (equivalent to bed thickness of 27 mm) were found optimal. These information generated were useful for the second phase of the experiments, which was the hot test, in investigating the possibility of incinerating wool scouring sludges in the RFB. Nine wool sludges from different process routes generated from the wool scouring industries were analysed for their compositions. Most of these sludges were highly moist, had high volatile matter and high ash content with low level of fixed carbon. These characteristics made incineration difficult. Hence, the effect of varying the moisture content, rotation speed and sludge feed rate on the incineration of the three selected sludges were studied in the hot test. With 5% support methane, all sludges with a maximum moisture up to 70% as-received could be successfully burned in the RFB at rotating speeds of 200 and 300 rpm. The combustion was found to be intense with a high efficiency due to the good turbulence and mixing in the RFB. The combustion gases produced, i.e., CO, CO(2) and NO(x) were reasonably low due to the high combustion intensity and efficiency. To study the dynamics of the bed and freeboard region in the RFB, the velocity flow field was simulated using a computational fluid dynamics (CFD) model to generate information of the flow pattern. The special advantages of swirling flow would benefit the gas combustion in the RFB. The experimental results obtained have suggested that the incineration was successful and the ash particles elutriated were fine due to the good mixing and turbulence in the RFB. This also reflects the RFB as an effective incinerator. PMID- 10708891 TI - Kinetics of p-hydroxybenzoic acid photodecomposition and ozonation in a batch reactor. AB - The decomposition of p-hydroxybenzoic acid, an important pollutant present in the wastewaters of the olive oil industry, has been carried out by a direct photolysis provided by a polychromatic UV radiation source, and by ozone. In both processes, the conversions obtained as a function of the operating variables (temperature, pH and ozone partial pressure in the ozonation process) are reported. In order to evaluate the radiation flow rate absorbed by the solutions in the photochemical process, the Line Source Spherical Emission Model is used. The application of this model to the experimental results provides the determination of the reaction quantum yields which values ranged between 8.62 and 81.43 l/einstein. In the ozonation process, the film theory allows to establish that the absorption process takes place in the fast and pseudo-first-order regime and the reaction is overall second-order, first-order with respect to both reactants, ozone and p-hydroxybenzoic acid. The rate constants are evaluated and vary between 0.18x10(5) and 29.9x10(5) l/mol s depending on the temperature and pH. PMID- 10708892 TI - Attenuation mechanisms of N-nitrosodimethylamine at an operating intercept and treat groundwater remediation system. AB - The North Boundary Containment System (NBCS), an intercept-and-treat system, was established at Rocky Mountain Arsenal (RMA), Commerce City, CO, to remove low level organic contaminants from a groundwater plume exiting RMA to the north and northwest. N-nitrosodimethylamine (NDMA) was detected in groundwater collected from the dewatering and recharge zones of the NBCS system. Concern over the fate of NDMA, in terms of potentially exiting the boundaries of the arsenal, prompted an investigation to evaluate potential attenuation mechanisms for NDMA within the alluvial aquifer system and within the NBCS itself. Groundwater, soil, and granular activated carbon (GAC) samples were taken from key locations in the NBCS system. Soil and GAC samples were assayed for sorption kinetics and for adsorption and desorption properties using 14C-labeled NDMA. NDMA biodegradation experiments were conducted by following 14CO(2) evolution from 14C-labeled NDMA in soils and GAC samples under aerobic and anaerobic conditions. The sorptive capacity of the site soils for NDMA was insignificant. Furthermore, the adsorption of the NDMA by the soil was almost completely reversible. Evaluation of the degradation potential of the native microbial consortia indicated a high level of NDMA mineralization when measured using bench-scale microcosms. The native consortia had capability to mineralize the NDMA under both aerobic and anaerobic incubations, indicating facultative characteristics. Testing of the local groundwater chemistry revealed that the area of the aquifer of interest was microaerobic and neutral in pH. These conditions were optimal for NDMA removal. While sorption was insignificant, degradation was a significant attenuation mechanism, which may be the reason that no NDMA has migrated off-site. This gives rise to the potential of a long-term sink for attenuating NDMA within the recharge zone of the treatment system. PMID- 10708893 TI - Photocatalytic TiO(2) deposition by chemical vapor deposition. AB - Dip-coating, spray-coating or spin-coating methods for crystalline thin film deposition require post-annealing process at high temperature. Since chemical vapor deposition (CVD) process is capable of depositing high-quality thin films without post-annealing process for crystallization, CVD method was employed for the deposition of TiO(2) films on window glass substrates. Post-annealing at high temperature required for other deposition methods causes sodium ion diffusion into TiO(2) film from window glass, resulting in the degradation of photocatalytic efficiency. Anatase-structured TiO(2) thin films were deposited on window glass by CVD, and the photocatalytic dissociation rates of benzene with CVD-grown TiO(2) under UV exposure were characterized. As the TiO(2) film deposition temperature was increased, the (112)-preferred orientations were observed in the film. The (112)-preferred orientation of TiO(2) thin film resulted in a columnar structure with a larger surface area for benzene dissociation. Obviously, benzene dissociation rate was maximum when the degree of the (112) preferential orientation was maximum. It is clear that the thin film TiO(2) should be controlled to exhibit the preferred orientation for the optimum photocatalytic reaction rate. CVD method is an alternative for the deposition of photocatalytic TiO(2). PMID- 10708894 TI - A review on reproduction in South American camelids. AB - In this paper, aspects of reproductive physiology and endocrinology, as well as sexual behaviour in South American camelids are reviewed. Because of the many unique features of reproduction in these animals, the application of advanced breeding techniques that are routinely used in other domestic species has been slow and, in some cases, are not applicable. Relatively high embryonic loss and the capacity to carry only one offspring at a time limit production in females. Furthermore, some 20% of females do not conceive following mating. Research is needed to elucidate the causes of embryonic loss, particularly in relation to the preferential location of embryos in the left uterine horn and the apparent differential luteolytic activity of the two uterine horns. A fuller understanding of the endocrine changes and mechanisms accompanying folliculogenesis, estrus, induction of ovulation and luteal regression may led to treatments that provide better control of ovulation and enhance the quality and viability of eggs shed. In the male, the hormonal interactions involved in facilitating libido need to be established and the underlying causes of declining libido resulting from continued exposure to estrous females, identified. More importantly, there is a need to develop a routine method to collect semen from animals on farm, or in studs. Deficiencies in this area have long hampered the evaluation of sires through proper assessment of their semen quality and in the application of artificial insemination. In addition, establishment of techniques to freeze semen from these species has obvious advantages for breeding, including crossbreeding between species to improve products such as fibre and to assist in preservation of some of the more endangered New World camelids. PMID- 10708895 TI - Regulation of seasonal reproductive activity in the stallion, ram and hamster. AB - This review considers seasonal reproduction in male animals with emphasis on the stallion, ram and hamster. The pineal hormone melatonin is the common link between photoperiod and reproduction. An increase in the daily diurnal period of melatonin secretion is associated with a decrease in GnRH release in long-day breeders, but an increase in GnRH release in short-day breeders. Melatonin influences GnRH release within or close to the mediobasal hypothalamus in rams; whereas melatonin receptors have not been found in the hypothalamus of horses. Prolactin release is positively correlated with daylength. Prolactin concentrations are consequently low during the breeding season of sheep and high during the breeding season of horses and hamsters. Prolactin stimulates testicular function in rams. Seasonal changes in GnRH release in the horse are regulated by changes in a GnRH-inhibitory opioidergic tone. Opioids are at least, in part, responsible for the decrease in testicular function during winter. An opioidergic inhibition of LH release is present during the breeding season in rams; but dopaminergic pathways inhibit LH release during long daylight hours. A dopaminergic inhibition of LH release does not exist in stallions. PMID- 10708896 TI - Immunolocalization of proacrosin/acrosin in bovine sperm and sperm penetration through the zona pellucida. AB - The aim of the present work was to immunolocalize acrosin in bull spermatozoa incubated for up to 6 h in capacitating culture medium (TALP-heparin), in order to study the kinetics of its release during the acrosome reaction and in vitro sperm penetration. Six replicates from semen of one bull were used. Acrosin was localized by the silver-enhanced immunogold technique using anti-bovine acrosin monoclonal antibody ACRO-C2E5. Spermatozoa thus showed the presence of acrosin only at the acrosomal region. Four different patterns were seen: (1) no labeling: (2) intense labeling on the rim of the portion of the acrosome; (3) diffuse label over the entire acrosomal region; and (4) intense label over the entire acrosomal region. Spermatozoa incubated in capacitating medium for 4 h showed that unlabeled (pattern 1) spermatozoa decreased from 72% to 28% difference that was found to be significant (p<0.05). Patterns 3 and 4 increased from about 10% to 20 29%, (p<0.05). With further incubation (4-6 h), pattern 1 increased while patterns 3 and 4 decreased differences were not significant (p0.05). The incidence of pattern 2 did not change through the whole incubation period. Sperm penetration through the zona pellucida of in vitro matured bovine oocytes (57%) or empty zonae pellucida (70.5%) increased (p<0.05) as a function of sperm incubation time in capacitating medium. The presence of acrosin, as determined by the silver-enhanced immunogold technique, was highly correlated with sperm penetration of in vitro mature bovine oocyte (r=0.98) and cryopreserved zonae pellucidae (r=0.93) (p<0.01). PMID- 10708897 TI - The effect of macromolecular supplementation on the surface tension of TCM-199 and the utilization of growth factors by bovine oocytes and embryos in culture. AB - The objectives of this study were to determine the surface tension of bovine follicular fluid (BFF) and TCM-199, and the effects of the synthetic surfactant, Twin-80, or FCS, in TCM-199 on surface tension measurements and subsequent effects on bovine oocyte and embryo development. The surface tension of BFF was determined to be approximately 45.5 mN m(-1) at 25 degrees C and approximately 42.7 mN m(-1) at 39 degrees C, which was comparable to the surface tension of TCM 199 containing Twin-80 (45.7 and 43.2 mN m(-1), respectively). There was no difference in surface tension measurements of BFF from follicles 2-7 mm or 8-15 mm in diameter. Both Millipore water and phosphate buffered saline (PBS) had a surface tension measurement of 69.5 mN m(-1) at 39 degrees C. Although the presence of Twin-80 in TCM-199 resulted in a reduction in surface tension measurement as compared with unsupplemented TCM-199, there was no effect on the number of oocytes reaching metaphase II. However, the addition of epidermal growth factor (EGF) to TCM-199 containing Twin-80 did result in increased maturation rates of oocytes in vitro. There was no effect of insulin, transferrin, and selenium (ITS), or EGF on surface tension measurement of TCM 199, but significantly more zygotes cleaved and developed to morulae/blastocysts in TCM-199 containing both Twin-80 and growth factors. The addition of 20 microg EGF ml(-1) to TCM-199 containing Twin-80 was as efficacious in supporting bovine embryos in culture as was the addition of 5% or 10% fetal calf serum. This study demonstrates that surface-active components in culture media positively affect bovine oocyte maturation and embryo development in culture. Data also suggest that non-ionic surfactants, such as Twin-80 in TCM-199, may successfully replace the surface-active properties but not the embryotrophic properties of serum in embryo maturation/culture media. Although commercial TCM-199 (containing Twin-80) did not require the addition of other surface-active compounds to lower surface tension, it did benefit from the addition of growth promoting factors, which were also provided by serum. PMID- 10708898 TI - Endocrine profiles of dairy cows following experimentally induced clinical mastitis during early lactation. AB - Concentrations of LH, cortisol, estradiol-17beta (E(2)), prolactin and 13,14 dihydro-15-keto-prostaglandin F(2alpha) (PGFM) were determined in cows with experimentally induced clinical mastitis during early lactation. Cows free of intramammary infection (IMI) and in the luteal phase of the estrous cycle were balanced by lactation number and days in milk and assigned to either control (n=5) or treatment (n=5) groups. Treated cows were infected experimentally (day 0), in two mammary quarters, with Streptococcus uberis and developed clinical mastitis within 60 h after inoculation as evidenced by increased mastitis scores, elevated rectal temperatures, mammary swelling and isolation of S. uberis pathogen. Four days following bacterial challenge, blood samples were collected every 20 min for 8 h for determination of PGFM and LH following administration of oxytocin and GnRH, respectively. Blood samples were also collected on days 0, 4 and 7 of the experiment to determine concentrations of E(2), prolactin and cortisol. Four days after bacterial challenge, concentrations of cortisol were higher (P=0.04) in experimentally infected cows than controls. Experimentally challenged cows had increased (P=0.02) concentrations of cortisol on days 4 and 7 compared with day 0. Control cows had no significant increase in blood cortisol during the experimental period. Baseline concentrations of PGFM did not differ between groups; however, peak concentrations of PGFM following oxytocin challenge were elevated (P=0.006) in cows with clinical mastitis compared with control animals. Prolactin, E(2) and LH did not differ between cows with clinical mastitis or controls. Experimentally induced mastitis during early lactation elevated concentrations of cortisol during the luteal phase of the estrous cycle. Furthermore, mastitic cows demonstrated an increased PGFM response following oxytocin administration. Altered reproductive efficiency in cows with clinical mastitis caused by Gram-positive pathogens may be the result of increased uterine sensitivity to prostaglandin F(2alpha) (PGF(2alpha)). PMID- 10708899 TI - Norgestomet- and oestradiol valerate-induced luteolysis is dependent upon the uterus. AB - Beef heifers were assigned to three groups: (1) untreated controls (n=4), (2) Syncro-Mate B(R) (SMB)-treated (n=5), and (3) hysterectomized and SMB-treated (n=4). SMB was administered 8 or 9 days after oestrus, approximately 30 days after hysterectomy. This study was conducted to determine if the uterus was necessary for SMB to induce luteolysis. SMB induced premature luteolysis as only 20% of the intact SMB-treated heifers had >/=0.75 ng/ml of progesterone 7 days after the time of SMB treatment, compared to all (100%) of the untreated heifers (p<0.05). By 9 days after the time of SMB treatment, 25% of the untreated heifers and none (0%) of the intact SMB-treated heifers had >/=0.75 ng/ml of progesterone; however, all (100%) of the hysterectomized SMB-treated heifers had >/=0.75 ng/ml of progesterone (p<0.05). Therefore, SMB-induced luteolysis required the involvement of the uterus. The luteolysin, prostaglandin F(2alpha), is probably the secretion from the uterus that mediates the SMB-induced luteolysis. SMB treatment, however, required 7-8 days to induce luteolysis. PMID- 10708900 TI - Secretion of IGF-1 by ovine granulosa cells: effects of growth hormone and follicle stimulating hormone. AB - Insulin-like growth factor-1 (IGF-1) is implicated in follicle development and is considered to mediate the actions of growth hormone (GH) and gonadotrophins at the ovarian level. However, the expression and secretion of IGF-1 by the ovary are controversial, partly because of species and cell-type specificity. The present study investigated whether IGF-1 is produced by ovine granulosa cells and whether its production is regulated by GH and follicle stimulating hormone (FSH). Follicles (>/=4.0 mm) were obtained from ewes during seasonal anoestrus. Granulosa cells were cultured for a total period of 96 h in Dulbecco's modified Eagle's medium (DMEM)/Ham's F-12 medium supplemented with BSA (0.1%, w:v), transferrin (0.5 microg/ml) and testosterone (100 ng/ml). In the first set of experiments, cells were incubated in the presence of bovine calf serum (BCS) (2.5%) for the initial 48 h of culture. The cells were then cultured for the next 48 h in medium without BCS, but containing either GH (0, 2, 20, and 200 ng/ml) or FSH (0, 20, 200, and 2000 ng/ml). The medium was assayed for oestradiol (E), progesterone (P) and IGF-1. There were six wells per treatment and the experiment was carried out four times. Control granulosa cells maintained both IGF-1 and E secretion, with only low levels of progesterone output. In all experiments, both GH and FSH produced significant (P<0.001) dose-related increases in E, IGF-1 and P secretion into the medium. The maximum responses to GH (20 or 200 ng/ml) were 402% for E and 528% for IGF-1 compared with controls. The maximum responses to FSH (200 or 2000 ng/ml) were 460% for E and 514% for IGF-1. The objective of the second set of experiments was to determine the effect of the progestogenic status of cells on IGF-1 production. Granulosa cells were cultured both in the presence and absence of BCS (2.5% in the medium) during the initial 48 h of culture. For the next 48 h, cells were cultured in serum-free medium. Addition of BCS to the medium during the initial 48 h of culture stimulated progesterone production. However, it did not affect either IGF-1 or oestradiol secretion between 49 and 96 h of culture, or the cell numbers at the end of culture. In conclusion, (1) IGF-1 is secreted by granulosa cells irrespective of their progestogenic status and (2) concomitant increases in E and IGF-1 production by granulosa cells as a result of GH and/or FSH treatment suggest a role for GH and FSH in the regulation of ovarian function. PMID- 10708901 TI - Ovarian antral follicular dynamics and their associations with peripheral concentrations of gonadotropins and ovarian steroids in anoestrous Finnish Landrace ewes. AB - Daily transrectal ultrasonography of ovaries was done in seven Finn ewes during three 17-day periods from May to July. Blood samples were collected each day for estimation of the serum follicle-stimulating hormone (FSH), oestradiol and progesterone concentrations, and also every 15 min for 6 h, halfway through each period of ultrasonographic examination, to determine the patterns of gonadotropic hormone secretion. Four ewes ceased cycling from March to mid-April (ewes entering anoestrus early) and three in May (ewes entering anoestrus late). In all ewes cyclicity resumed during the period from mid-August to mid-September. The growth of ovarian antral follicles to periovulatory sizes of >/=5 mm in diameter was seen at all stages of anoestrus. An average of four waves of follicular development (follicles growing from 3 to >/=5 mm in diameter before regression) with a periodicity of 4 days were recorded during each of the three scanning periods. There was a close temporal relationship between days of follicular wave emergence and peaks of successive FSH fluctuations. Ewes entering anoestrus late exceeded ewes that became anoestrus early in numbers of large (>/=5 mm in diameter) ovarian antral follicles and maximum follicle diameter. Peak concentrations of transient FSH increases were higher (P<0.05) in ewes entering anoestrus late than in ewes entering anoestrus early. The secretion of luteinising hormone, (LH; mean and basal level, and LH pulse frequency, but not amplitude) was lowest during the month of June in all ewes. Oestradiol production was markedly suppressed throughout anoestrus. Peaks of progesterone secretion appeared to occur at regular intervals and were associated with the end of the growth phase of the largest follicles of sequential waves. In conclusion, the growth of ovarian follicles to ostensibly ovulatory diameters is maintained throughout anoestrus in Finn ewes and periodic emergence of follicular waves is correlated with an endogenous rhythm of FSH secretion. The present study also provides evidence for the inverse relationship between the time of the onset of seasonal anoestrus and the number and size of antral follicles developing throughout anoestrus in Finn ewes, and indicates that differences exist in both the secretion of and ovarian responsiveness to gonadotropic hormones among early and late anoestrous ewes. PMID- 10708902 TI - Fish neurotrophins and Trk receptors. PMID- 10708903 TI - Comparative postnatal development of dopamine D(1), D(2) and D(4) receptors in rat forebrain. AB - Postnatal development of dopamine D(1), D(2) and D(4) receptors in the caudate putamen, nucleus accumbens, frontal cortex and hippocampus was assessed in rat brain between postnatal days 7 and 60. In the caudate-putamen and nucleus accumbens, density of all three receptor subtypes increased to a peak at postnatal day 28, then declined significantly in both regions (postnatal days 35 60) to adult levels. In the frontal cortex and hippocampus, these receptors rose steadily and continuously to stable, maximal adult levels by postnatal day 60. Evidently, D(1), D(2) and D(4) receptors follow a similar course of development in several cortical, limbic and extrapyramidal regions of rat forebrain, with selective elimination of excess dopamine receptors at the time of puberty in the caudate-putamen and accumbens but not other brain regions. PMID- 10708904 TI - Antiproliferative actions of inhalational anesthetics: comparisons to the valproate teratogen. AB - The antiproliferative potential of the volatile anesthetics isoflurane, enflurane and sevoflurane was determined and compared to the valproate teratogen. The in vitro system employed, a G1 phase proliferative arrest endpoint in C6 glioma, has served previously to discriminate agents with known teratogenic potential in vivo. Based on estimated IC(50) values that were within twice the estimated minimum aveolar concentration value, the rank antiproliferative potency of the inhalational anesthetics employed was isoflurane=enflurane>>sevoflurane. Flow cytometric analysis of growth-arrested cell populations failed to reveal specific accumulation in any cell cycle phase and the lack of a G1 phase-specific effect was confirmed by the absence of a transient, time-dependent sialylation event in synchronized cells. The antiproliferative mechanism of volatile anesthetics, and valproate, was mediated at hydrophobic binding sites, as increasing the hydration sphere of the drug-micelle complex, using the hygroscopic qualities of the dimethylsulfoxide vehicle, completely reversed this effect. Our findings suggest inhalational anesthetics lack the specific in vitro characteristics of the valproate teratogen. PMID- 10708905 TI - Thyrotropin-releasing hormone immunoreactivity in the brain and the pituitary during Bufo arenarum development. AB - The ontogeny of the thyrotropin releasing hormone (TRH) neuronal system was evaluated by immunocytochemistry in Bufo arenarum. The first appearance of TRH immunoreactive fibers was at early premetamorphosis. These fibers were found in small numbers and weakly stained in the median eminence and pars nervosa. With the advance of larval development, TRH-like material stained intensely and tended to aggregate in the median eminence, pars nervosa and pars intermedia. At climax stages immunoreactive fibers and perikarya (weakly stained) were also identified in the preoptic area. In adult specimens TRH perikarya and neuronal fibers were found in the preoptic and infundibular nuclei of the hypothalamus and in the amygdala, septum and diagonal band of Broca of the telencephalon. In addition, TRH neuronal fibers and endings were found in the preoptic-hypophyseal tract, the external zone of the median eminence, the pars nervosa and pars intermedia. Fibers were absent in the pars distalis. This study represents the first immunocytochemical demonstration of TRH in Bufo species, and serves as a basis for clarification of the neuroendocrine regulation of metamorphosis. PMID- 10708906 TI - Ontogeny of epinephrine metabolic pathways in the rat: role of glucocorticoids. AB - Recent studies suggest that the initial expression of adrenal phenylethanolamine N-methyltransferase (PNMT) and epinephrine (E) are dependent upon stimulation of adrenal glucocorticoid receptors. However, evidence suggests that the expression of heart and brain PNMT is independent of glucocorticoids. We measured PNMT activity and E levels in adrenal, heart and head over the latter half of gestation in rat fetuses treated chronically with glucocorticoids, and in normal controls. Chronic glucocorticoid treatment ending on embryonic day (e)12 did not affect heart, head or trunk PNMT activity or E levels. In contrast, chronic glucocorticoid exposure ending e19 or e20 resulted in marked increases in both PNMT and E in adrenal, heart and head tissues. The elevation of E in all three tissues was unaffected by maternal adrenalectomy, indicating enhanced fetal E synthesis. In the absence of exogenous glucocorticoid treatment heart PNMT activity peaked on e12, prior to the earliest reported appearance of glucocorticoid receptors. We conclude that expression of PNMT in all three tissues is glucocorticoid independent until the latter part of gestation when it is readily enhanced by glucocorticoids. PMID- 10708907 TI - AMPA receptor subunit mRNAs and intracellular [Ca(2+)] in cultured mouse and rat cerebellar granule cells. AB - Cultured mouse cerebellar granule cells differ from their rat counterparts in that they survive well when grown in non-depolarising medium (5 mM K(+)). However, when chronically stimulated by added glutamate agonists, including (RS)alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA), rat cerebellar granule cells also survive well in non-depolarising medium. We hypothesised that the relatively good survival of mouse cerebellar granule cells in the absence of added glutamate agonists might reflect AMPA receptors resistant to desensitisation. These receptors might be stimulated by endogenous glutamate. We tested this hypothesis by comparing cultured mouse and rat cerebellar granule cells grown in depolarising (25 mM K(+)) and non-depolarising (5 mM K(+)) medium. We studied the AMPA-induced increase in intracellular Ca(2+) concentration ([Ca(2+)](i)), using the fluorescent Ca(2+) chelator, Fluo-3, and the relative concentrations of mRNAs for the four AMPA receptor subunits, GluR1-4. GluR1-4 mRNAs were measured by restriction enzyme analysis of a PCR product containing cDNA with a composition proportional to the four subunit mRNAs. We found that the [Ca(2+)](i)-response to AMPA receptor activation in cultured cerebellar granule cells is determined mainly by the desensitisation properties of the AMPA receptors rather than by their ion permeability. We also found that mouse cerebellar granule cells express AMPA receptors which are more resistant to desensitisation than the corresponding rat AMPA receptors. Thus, relatively slow AMPA receptor desensitisation kinetics may contribute to the survival of mouse cerebellar granule cells in non-depolarising medium. PMID- 10708908 TI - Expression of insulin-like growth factor-I (IGF-I) and IGF-II in the avian brain: relationship of in situ hybridization patterns with IGF type 1 receptor expression. AB - Insulin-like growth factors (IGFs) are expressed in defined spatiotemporal patterns during the development of the mammalian central nervous system (CNS). Since IGF expression in avian species is less well documented, we studied here the expression of IGF-I and IGF-II during chicken CNS development, using in situ hybridization and reverse transcriptase-PCR, and compared the results with the expression of the IGF type 1 receptor (IGF-1R). IGF-II expression started early in embryonic life, shortly after the onset of IGF-1R expression. During organogenesis, IGF-II was strongly expressed in kidney, liver and gut primordia, in contrast with IGF-1R mRNA, which is highly enriched in proliferating neuroepithelia. During the second half of embryonic development, IGF-I and IGF-II had distinct expression patterns, suggesting specific roles for each ligand during brain maturation. IGF-II mRNA was found in numerous brainstem nuclei and in the optic tectum, whereas IGF-I mRNA was found predominantly in telencephalic regions. Both ligands were expressed in the cerebellum, but each by different cell layers. Some brain regions (olfactory bulb and olivo-cerebellar system) did not exhibit the postnatal downregulation typical of extrahepatic IGF-I expression, but continued to express IGF-I into adulthood. Purkinje cells expressed IGF-II in the embryo, but switched to IGF-I expression in the adult. The conservation of embryonic and postnatal IGF expression patterns in the CNS between avians and mammals suggests that the involvement of the IGF system in neurogenesis and differentiation, and possibly in neural plasticity and learning, may have arisen early during tetrapode/vertebrate evolution. PMID- 10708909 TI - Levels of amino acid neurotransmitters during mouse olfactory bulb neurogenesis and in histotypic olfactory bulb cultures. AB - The developmental changes in the levels of amino acid neurotransmitters were analyzed by high pressure liquid chromatography during mouse olfactory bulb neurogenesis, from embryonic day (E)13 until the young adult age, between postnatal days (P)30 and P40. During the embryonic period, high levels of glutamate, aspartate and GABA were observed, with the values of GABA about 2-fold higher than those of glutamate and aspartate. At P0, the production of these neurotransmitters experienced birth stress as shown by a significant 2-fold reduction in their levels. During the first two postnatal weeks, a progressive increase in the glutamate content was detected diminishing slightly in the adult stage. The aspartate concentrations showed a maximal value at P3 and then decreased gradually until the second postnatal week; in the young adult age, its concentration was comparable with that of glutamate. The postnatal GABA contents increased progressively from birth to maturity, showing maximal levels at P3, P11 and in the adult. Throughout the studied developmental period, the concentration of glycine remained relatively low. With regard to taurine, very low concentrations were detected during the prenatal period but after birth, the taurine content gradually increased with age, and in the adult animal, its concentration was comparable with those of GABA and glutamate. Our data demonstrate the predominance of GABA and glutamate during olfactory bulb synaptogenesis, however, in the adult animal, both glutamate and aspartate exert the same influence in the excitatory synaptic transmission; in the adult inhibitory synaptic transmission, taurine appears to play an important neuromodulatory or neurotransmitter role as that of GABA. To determine the intrinsic neurotransmitter production, primary histotypic olfactory bulb cultures were prepared from mice at P10. The comparative analysis of in vitro neurotransmitter contents with those in in situ adult animal showed higher levels of endogenously produced glutamate, glycine and GABA in the olfactory bulb than the extrinsic ones coming from olfactory nerve axons and higher olfactory brain centers. On the other hand, most of aspartate and taurine neurotransmitters apparently come from extrinsically located neurons. PMID- 10708910 TI - Activation of neurospecific gene expression by antennapedia homeobox peptide. AB - Antennapedia homeobox peptide has been reported to enhance neurite outgrowth and branching. Thus it is of interest to investigate whether antennapedia peptide is capable of modulating the expression of genes related to different events of neuronal development. In this paper we report the enhancement of a 68 KDa neurofilament subunit, choline acetyltransferase and acetylcholinesterase expression in spinal cord neurons, elicited by antennapedia peptide. Modulation of gene expression is different with respect to each gene product analyzed, suggesting a specific action of the peptide on diverse genes controlling different events of neuronal differentiation. PMID- 10708911 TI - Histochemical and electrophysiological evidence for estrogen receptors on cultured astrocytes: colocalization with cholinergic receptors. AB - By means of autoradiographic and immunohistochemical methods it was demonstrated that astrocytes in explant and primary cultures of rat neocortex, hippocampus, preoptic area and spinal cord express estrogen alpha- and beta-receptors. Immunoreactivity was mainly distributed over the soma, the nuclei being more intensely stained. Combined autoradiographic and immunohistochemical studies as well as double-immunostaining revealed a colocalization of estrogen alpha- and beta-receptors on many astrocytes. There was also a coexistence of estrogen receptors and cholinergic muscarinic and nicotinic sites. Electrophysiological investigations have shown that 17beta-estradiol induced hyperpolarizations on the majority of astrocytes in explant cultures of hippocampus and spinal cord, providing evidence for the existence of functional estrogen receptors on these cells. Furthermore, on the same astrocytes, 17beta-estradiol, muscarine and nicotine caused hyperpolarizations, suggesting a coexistence of receptors for estrogen and the cholinergic agonists on glial cells. The presence of glial estrogen receptors and their colocalization with cholinergic receptors is discussed with respect to the effects of these neurotransmitters/neuromodulators in development and maturation of the central nervous system, as well as to neurodegenerative events such as Alzheimer's disease. PMID- 10708912 TI - Phosphorylated MAP-1B isoforms in the developing mouse barrel cortex. AB - Developmental expression of two phosphorylation modes of microtubule-associated protein 1B (MAP-1B) has been studied in the barrel cortex of mice at postnatal days (P)5, P12, P21 and P90 using immunocytochemistry with antibodies 125 and 150 that recognize phosphorylation modes II and I, respectively. The antibody 125 immunoreactive processes, identified as dendrites, are not yet detectable at P5; they are already present at P12 and become more evident at P21. In the barrel cortex of P90 animals the antibody 125 immunopositive dendrites are still present, although they are much less pronounced. The antibody 150 punctate immunostaining seen at P5 is not detectable at P12. At P21, however, thin immunopositive fibres appear, implicating a re-expression of the microtubule associated protein 1B phosphorylation mode I in a portion of axons. The antibody 150 immunopositive axons are no longer present in the P90 barrel cortex. The re expression of the MAP-1B phosphorylation mode I, which is a juvenile isoform characteristic for growing axons, may imply induction of mechanisms providing mouse barrel cortex neurons with the potency for plastic changes at a terminal stage of synaptogenesis. PMID- 10708913 TI - Developmental maturation of the N-methyl-D-aspartic acid receptor channel complex in postnatal rat brain. AB - The N-methyl-D-aspartate (NMDA) receptor plays an important role in developmental plasticity. Previous studies have reported differences between the NMDA receptor channel complex in the rat pup brain and the adult brain. In the present study, modulation of the NMDA channel complex as a function of age was measured to determine when the temporal switching of the NMDA receptor from the immature form to the adult mature form takes place. [(3)H]MK-801 binding was measured in the rat forebrain from postnatal day 1 to day 21. Our data suggest the presence of two types of NMDA receptors - an immature type and a mature type. The immature NMDA receptor, seen during the early postnatal period (day 1-day 14) is highly sensitive to spermidine, L-glutamate alone potentiates [(3)H]MK-801 binding, and glycine failed to potentiate an L-glutamate-induced increase in [(3)H]MK-801 binding. During the late postnatal period (after day 14) spermidine alone did not increase [(3)H]MK-801 binding as potently as it did during the early postnatal period, high-affinity [(3)H]MK-801 binding was not seen in the presence of L glutamate alone, and L-glutamate and glycine or L-glutamate and spermidine or L glutamate, glycine and spermidine together, significantly increased [(3)H]MK-801 binding in a manner similar to that reported in the adult brain. Together, the pharmacology of the NMDA receptor during the early postnatal period differs from the adult-like receptor seen during the late postnatal period, and that in rats the apparent switching of the NMDA receptor from the immature type to the mature type takes place after the second postnatal week. PMID- 10708914 TI - Distribution of dopamine-immunoreactive fibers in the rat brainstem. AB - We describe the distribution of axons immunoreactive for dopamine in pons and medulla oblongata of rat under normal conditions or after inhibition of monoamine oxidase or dopamine beta-hydroxylase. In the pons of non-treated animal, fairly dense plexuses of dopamine-immunoreactive varicose fibers were found in the locus coeruleus, dorsal parabrachial and dorsal raphe nuclei, central gray and reticular formation dorsal to the superior olive. In the medulla oblongata, the immunoreactive fibers were abundant in the dorsal vagal complex, lateral paragigantocellular nucleus, midline raphe nuclei and spinal trigeminal nucleus. Monoamine oxidase inhibition made it possible to increase the intensity of immunoreactivity and consequently the number of labeled fibers in these areas, indicating that dopamine is perpetually oxidized by monoamine oxidase, and consequently in low concentration under normal conditions. Sparse dopamine immunoreactive fibers were observed in the pontine gray, motor trigeminal nucleus, inferior olive and major axon bundles such as the dorsal and ventral tegmental bundles, where numerous noradrenergic fibers have been reported. In axons of these areas, intense dopamine-immunoreactivity was seen only after inhibition of dopamine-beta-hydroxylase. It appears that dopamine is released and oxidized in response to autonomic changes such as hypoxia, hemorrhage, and cardiovascular variation in the caudal brainstem, as we have described elsewhere. PMID- 10708915 TI - Ontogeny of (D-Ala(2))-deltorphin I-like immunoreactive neurones in foetal rat brain. AB - Foetal rat brain from embryonic day (ED) 12-22 was immunohistochemically studied to describe the time of first appearance and further distribution patterns of (D Ala(2))-deltorphin-I-immunoreactive (DADTI-IR) nerve elements. The primary antiserum used in this study was a polyclonal antibody against DADTI previously used in adult and postnatal rat brain mapping. DADTI-IR nerve elements first appeared in the neuroepithelium of ventral mesencephalon on ED 13. From there, positive cell bodies migrated towards the mantle layer until they invaded the whole ventral mesencephalic tegmentum. They then reached their definitive position, corresponding to a subpopulation of the A8, A9 and A10 dopaminergic neurones that had been constantly observed also in the adult age. From ED 15-17, DADTI-positive nerve fibres appeared in the medial forebrain bundle, the neostriatum anlage, the accumbens nucleus, the olfactory tubercle, the fasciculus retroflexus, and the prefrontal cortex. All these locations have also been found in adult rats. From ED 14 onwards, transient DADTI-IR somata and nerve fibres were observed in retinal neuroepithelium, optic pathways as far as the superior colliculus, CA3 hippocampal field, reticular formation in the medulla oblongata. All these locations gradually disappeared either before birth (medulla oblongata) or within the first 3 weeks after birth. These results suggest that the DADT-like molecule recognised by our antibody has during the embryonic development a regulatory function in neuronal growth and differentiation. PMID- 10708916 TI - The ascending reticular activating system--from aminergic neurons to nitric oxide. AB - The cholinergic neurons of the laterodorsal and pedunculopontine tegmental neurons are thought to comprise an important portion of the ascending reticular activating system. More recent work has demonstrated that the neurons of this cell group also released a number of neruoactive peptides and can produce nitric oxide in response to increases in intracellular calcium. The release of NO from the nerve terminals of these cells within the thalamus varies with behavioural state, being much lower during slow wave sleep than during wake and paradoxical sleep states. The NO release in the thalamus appears to act via the type II cGMP dependent protein kinase present at high levels in the thalamic neurons. Thus the NO-cGMP signal transduction system can play an important role in regulating thalamic activity across behavioural states. PMID- 10708917 TI - Immunohistochemical demonstration of choline acetyltransferase of a peripheral type (pChAT) in the enteric nervous system of rats. AB - Using a recently developed antiserum against a splice variant (pChAT) of choline acetyltransferase, the enzyme which synthesizes acetylcholine, we carried out an immunohistochemical examination in the digestive canal of rats. Positive staining was exclusively localized to neuronal cells and fibers. Positive somata were distributed widely in the intramural ganglia throughout the digestive tract from the esophagus to the rectum. Double staining indicated that, in the rat, virtually all pChAT immunoreactive somata exhibited histochemical activity for acetylcholinesterase but not for NADPH-diaphorase. In the guinea pig, however, there were a few neurons possessing both pChAT and NADPH-diaphorase. We also found a few neuronal somata which were positive for acetylcholinesterase but not for pChAT. The results suggest that pChAT immunohistochemistry is useful for studying the enteric cholinergic system. PMID- 10708918 TI - Serotoninergic neurons and serotonin receptors: gains from cytochemical approaches. AB - Serotonergic systems, their phylogeny and ontogeny have been thoroughly described up to the ultrastructural level, thanks to the multiplicity of methodological approaches. They have often been referred to as a 'Rosetta stone', as several features first described for serotonin neurons or paraneurons have been then extended to other neurotransmitter systems: coexistence with neuropeptides or even a canonical neurotransmitter (GABA), volume transmission, regrowth after lesioning, and characterization of multiple receptor subtypes. This review deals with the contributions of neuroanatomical approaches for studying serotoninergic systems, and focuses on recent advances concerning the topological relationships between serotonergic innervation, receptors and target cells. This aspect is particularly important with regard to the possibility for serotonin to act through classical synaptic transmission and/or non-junctional transmission. Serotonin then can selectively regulate different neuronal systems through the activation of distinct receptor subtypes, which in turn can be linked to different transduction pathways. Neurocytochemical approaches constitute unique tools to analyse both anatomical and functional characteristics of complex neuronal systems. PMID- 10708919 TI - The locus coeruleus: history. AB - The author outlines the history of the locus coeruleus from its discovery to the latest research findings. Although Russel's report in Texas has been believed to be the first break-through scientific article of this neuronal structure, there were two preceding, anatomical and physiological, studies made by Japanese groups. One of the aims of this review is to shed brief light on their works, which being written in Japanese so that might have been unfamiliar to most researchers in the world. PMID- 10708920 TI - The histaminergic system in the brain: structural characteristics and changes in hibernation. AB - Histaminergic neurons in adult vertebrate brain are confined to the posterior hypothalamic area, where they are comprised of scattered groups of neurons referred to as the tuberomammillary nucleus. Histamine regulates hormonal functions, sleep, food intake, thermoregulation and locomotor activity, for example. In the zebrafish, Danio rerio, histamine was detected only in the brain, where also the histamine synthesizing enzyme L-histidine decarboxylase (HDC) was expressed. It is possible that histamine has first evolved as a neurotransmitter in the central nervous system. We established sensitive quantitative in situ hybridization methods for histamine H(1) and H(2) receptors and HDC, to study the modulation of brain histaminergic system under pathophysiological conditions. A transient increase in H(1) receptor expression was seen in the dentate gyrus and striatum after a single injection of kainic acid, a glutamate analog. H(1) antagonists are known to increase duration of convulsions, and increased brain histamine is associated with reduced convulsions in animal models of epilepsy. No HDC mRNA was detected in brain vessels by in situ hybridization, which suggests lack of histamine synthesis by brain endothelial cells. This was verified by lack of HDC mRNA in a rat brain endothelial cell line, RBE4 cells. Both H(1) and H(2) receptor mRNA was found in this cell line, and the expression of both receptors was downregulated by dexamethasone. The findings are in agreement with the concept that histamine regulates blood-brain barrier permeability through H(1) and H(2) receptor mediated mechanisms. Hibernation is characterized by a drastic reduction of central functions. The activity of most transmitter systems is maintained at a very low level. Surprisingly, histamine levels and turnover were clearly elevated in hibernating ground squirrels, and the density of histamine containing fibers was higher than in euthermic animals. It is possible that histamine actively maintains the low activity of other transmitters during the hibernation state. PMID- 10708921 TI - Multiple messengers in descending serotonin neurons: localization and functional implications. AB - In the present review article we summarize mainly histochemical work dealing with descending bulbospinal serotonin neurons which also express a number of neuropeptides, in particular substance P and thyrotropin releasing hormone. Such neurons have been observed both in rat, cat and monkey, and may preferentially innervate the ventral horns of the spinal cord, whereas the serotonin projections to the dorsal horn seem to lack these coexisting peptides. More recent studies indicate that a small population of medullary raphe serotonin neurons, especially at rostral levels, also synthesize the inhibitory neurotransmitter gamma-amino butyric acid (GABA). Many serotonin neurons contain the glutamate synthesizing enzyme glutaminase and can be labelled with antibodies raised against glutamate, suggesting that one and the same neuron may release several signalling substances, causing a wide spectrum of post- (and pre-) synaptic actions. PMID- 10708922 TI - Sexual dimorphism in the human brain: evaluation of tissue volume, tissue composition and surface anatomy using magnetic resonance imaging. AB - Magnetic resonance imaging (MRI) was used to evaluate sex differences in brain morphology by comparing measures of brain tissue volume, brain tissue composition (proportions of gray and white matter), and measures of cortical surface anatomy. A large and well-matched sample of healthy women (n=42) and healthy men (n=42) were evaluated. There was a significant gender effect on intracranial volume, males being larger. However, this increase in size was limited to the cerebrum as there was no sex difference in the volume of the cerebellum. The gender difference in size of the cerebral volume was evenly distributed, with all four lobes equally larger in males compared to females. Gray and white matter tissue proportions were similar between the sexes globally. Regional tissue composition analysis showed sex differences within the parietal lobes with females having proportionately more gray matter on the right side. There were no differences between the sexes in cortical surface anatomy measures. Overall, against the background of similarity in morphology, there are differences between the sexes with regard to general and regional brain measures. The functional significance of these sex differences is unclear, but may represent the differential effects of gonadal hormones during brain growth and development. PMID- 10708923 TI - Effect of ecstasy [3,4-methylenedioxymethamphetamine (MDMA)] on cerebral blood flow: a co-registered SPECT and MRI study. AB - 3,4-methylenedioxymethamphetamine (MDMA), an illicit recreational drug, damages serotonergic nerve endings. Since the cerebrovasculature is regulated partly by the serotonergic system, MDMA may affect cerebral blood flow (CBF) in humans. We evaluated 21 abstinent recreational MDMA users and 21 age- and gender-matched healthy subjects with brain SPECT and MRI. Ten of the MDMA subjects also had repeat SPECT and MRI after receiving two doses of MDMA. Abstinent MDMA users showed no significantly different global or regional CBF (rCBF) compared to the control subjects. However, within 3 weeks after MDMA administration, rCBF remained decreased in the visual cortex, the caudate, the superior parietal and dorsolateral frontal regions compared to baseline rCBF. The decreased rCBF tended to be more pronounced in subjects who received the higher dosage of MDMA. Two subjects who were scanned at 2-3 months after MDMA administration showed increased rather than decreased rCBF. Low-dose recreational MDMA use does not cause detectable persistent rCBF changes in humans. The lack of long-term rCBF changes may be due to a non-significant effect of serotonergic deficits on rCBF, or regeneration of serotonergic nerve terminals. The subacute decrease in rCBF after MDMA administration may be due to the direct effect of MDMA on the serotonergic system or the indirect effects of its metabolites on the dopaminergic system; the preliminary data suggest these effects may be transient. PMID- 10708924 TI - Reduced regional cerebral blood flow in non-psychotic violent offenders. AB - The present study was designed to replicate previously reported findings of abnormal frontal and/or temporal cerebral blood flow in violent offenders and to control for the influence of major mental disorder (MMD), substance abuse, and current medication. HMPAO-SPECT-CBF and MRI scans from pretrial forensic psychiatric investigations of 21 subjects convicted of impulsive violent crimes were retrospectively re-evaluated. In 16/21 subjects, visual assessment of SPECT scans showed some hypoperfusion in the temporal and/or frontal lobes. MRI showed no corresponding structural damage. Quantified regional cerebral blood flow (rCBF) in defined regions of interest was compared between index cases and 11 healthy control subjects. Index subjects had significant reductions in the right angular gyrus and the right medial temporal gyrus, bilaterally in the hippocampus, and in the left white frontal matter, but they had significantly increased rCBF in the parietal association cortex bilaterally. The aberrations were as frequent and severe among the subjects without MMD, substance abuse, and current medication (n=7) as in the entire group of index subjects. PMID- 10708925 TI - Preliminary findings of simultaneous 18F-FDG and 99mTc-HMPAO SPECT in patients with depressive disorders at rest: differential correlates with ratings of anxiety. AB - The assumption of a dynamic coupling between regional cerebral blood flow (rCBF) and cerebral glucose metabolic rates (rCMRGlu) has been challenged by simultaneous measurements of both. Through the use of a dual-headed gamma camera with a 511-keV collimator applying the double isotope 18F-FDG and 99mTc-HMPAO SPECT technique, the uptake rates of these isotopes can be semi-quantitatively evaluated. Sixteen depressed patients, diagnosed by ICD-10 criteria and assessed with the 17-item Hamilton Rating Scale for Depression (HRSD), were studied. Based on the severity of HRSD-rated anxiety (item 10: low=1-21; high=3-4), two eight patient subgroups were formed and compared with 12 age- and handedness-matched healthy control subjects. As regions of interest, we selected areas implicated in the neuroanatomy of anxiety and depression: hippocampus (hippo), basal ganglia (BG) and gyri temporales superiores (G.t.s.). In the control subjects, a significant statistical coupling between rCBF and rCMRGlu was revealed by the Spearman correlation coefficient only in left hippo and left BG. Patients in the low-anxiety subgroup demonstrated a marked dynamic coupling bilaterally for the G.t.s., while patients in the high-anxiety subgroup showed a significant statistical correlation of rCBF and rCMRGlu only in the left G.t.s. These findings indicate that a dynamic coupling between blood flow and glucose metabolism exists only in distinct brain regions, and that the depressive illness has an uncoupling effect on this correlation in the left BG. Furthermore, our results suggest that the HRSD anxiety score might interact with the underlying depressive illness to influence the relationship of rCBF and rCMRGlu. PMID- 10708926 TI - Modeling brain compartmental lactate response to metabolic challenge: a feasibility study. AB - Magnetic resonance spectroscopy has been used to characterize abnormal brain lactate response in panic disorder (PD) subjects following lactate infusion. The present study integrated water quantification and tissue segmentation to evaluate compartmental lactate response within brain and cerebrospinal fluid (CSF). As there is evidence of brain parenchymal pH changes during lactate infusion, water scans were collected at baseline and post-infusion to address brain water stability. Water levels remained essentially stable across the protocol suggesting internal water provides an improved reference signal for measuring dynamic changes in response to metabolic challenge paradigms such as lactate infusion. To model brain lactate changes by compartments, we took the null hypothesis that lactate rises occur only in tissue. The approach referenced lactate amplitude (potentially from both compartments) to 'voxel' water (water scan corrected for differential T(2) between CSF brain at long-echo times - synonymous to a short-echo water scan). If the magnitude of lactate rise in CSF was equal to or greater than brain, voxels with substantial CSF fractions should demonstrate an equivalent or elevated response to voxels comprised only of tissue. The magnitude of lactate increases paralleled voxel tissue fraction suggesting the abnormal lactate rise observed in PD is tissue-based. The feasibility of lactate quantification and compartmental modeling are discussed. PMID- 10708927 TI - Energy balance, cancer and the sympathetic nervous system. PMID- 10708928 TI - Do we need better prognostic factors in node-negative breast cancer? Pro:. PMID- 10708929 TI - Do we need better prognostic factors in nodal-negative breast cancer? Contra. PMID- 10708930 TI - Do we need prognostic factors in nodal-negative breast cancer? Arbiter. PMID- 10708931 TI - Evaluation of the carcinogenic risks to humans associated with surgical implants and other foreign bodies - a report of an IARC Monographs Programme Meeting. International Agency for Research on Cancer. AB - A meeting was held within the International Agency for Research on Cancer (IARC) Programme on the Evaluation of Carcinogenic Risks to Humans of surgical implants and other foreign bodies. This meeting report summarises the types of materials considered, their wear and degradation, their cancer epidemiology in both humans and other animals, the published experimental carcinogenicity data and selected data on their toxic, including genotoxic, effects. Evaluations resulting in a classification of Group 2B (possibly carcinogenic to humans) were reached for: (1) polymeric implants prepared as thin smooth films [with the exception of poly(glycolic acid)]; (2) metallic implants prepared as thin smooth films; and (3) implanted foreign bodies consisting of metallic cobalt, metallic nickel and a particular alloy powder consisting of 66-67% nickel, 13-16% chromium and 7% iron. Group 3 classifications (not classifiable as to their carcinogenicity to humans) were made for: (1) organic polymeric materials as a group; (2) orthopaedic implants of complex composition and cardiac pacemakers; (3) silicone breast implants; (4) dental materials; and (5) ceramic implants. PMID- 10708932 TI - Cancer chemoprevention through interruption of multistage carcinogenesis. The lessons learnt by comparing mouse skin carcinogenesis and human large bowel cancer. AB - Whilst in the early stages, neoplastic development is predominantly triggered by environmental genotoxic and non-genotoxic carcinogens, tumour progression becomes more and more autonomous at later stages. In this context a dysregulation of arachidonic acid metabolism seems to play a disastrous role. Conversely, non steroidal anti-inflammatory drugs (NSAIDs) rank among the most potent and most promising agents for cancer chemoprevention probably because of their ability to inhibit prostaglandin biosynthesis, in particular, at the level of the 'pro inflammatory' enzyme cyclooxygenase-2 (COX-2). A pathological overexpression of COX-2 resulting in excessive prostaglandin production has been found already in early stages of carcinogenesis and seems to be a consistent feature of neoplastic development in a wide variety of tissues. COX-2 overexpression is thought to occur along signalling pathways of inflammation and tissue repair which become activated in the course of tumour promotion and, due to autocrine and auto stimulatory mechanisms, finally lead to some autonomy of tumour development (self promotion). Prostaglandins formed along a dysregulated COX pathway have been shown to mediate tumour promotion in animal experiments and may play a role, in addition, in other processes involved in tumour growth such as angiogenesis, metastasis and immunosuppression. Moreover, genotoxic byproducts such as organic free radicals, reactive oxygen species and malondialdehyde produced in the course of prostanoid biosynthesis may contribute to genetic instability (mutator phenotype) of neoplastic cells thereby promoting malignant progression. Such mixtures of physiologically highly active mediators and genotoxic byproducts are, in addition, formed along the various lipoxygenase-catalysed pathways of arachidonic acid metabolism some of which also become dysregulated during tumour development and, therefore, provide novel targets of future chemopreventive approaches. PMID- 10708933 TI - Beta-adrenoceptor activity and resting energy metabolism in weight losing cancer patients. AB - This study was aimed at comparing the blocking of beta-adrenoceptor activity to changes in the resting energy metabolism of 10 cancer patients with progressive weight loss due to solid malignant tumours. Resting energy expenditure (REE) as well as whole body carbohydrate and fat oxidation were investigated and related to plasma substrate levels (glucose, glycerol, free fatty acids (FFA)) before and after 5 days of oral administration of specific beta1 receptor blocker (atenolol, 50 mg/day) and non-specific beta1,beta2-adrenoceptor (propranolol, 80 mg/day) blockade. The administration order of the drugs was random, and a 3-day washout period was used in all individuals between the provision of the first and the second drug in order to minimise the risk of carry-over effects. Resting measurements in the morning after an overnight fast were performed by indirect calorimetry. Atenolol treatment reduced REE by 77+/-14 kcal/day and propranolol by 48+/-13 kcal/day, respectively (P<0.05 versus pretreatment values). Whole body oxygen uptake and carbon dioxide production were decreased similarly by both atenolol and propranolol treatment (P<0.05). Carbohydrate oxidation was increased by atenolol and decreased by propranolol, whilst fat oxidation was decreased by atenolol and unchanged by propranolol. The decrease in REE, accounting for the decline in heart rate, was significantly more pronounced following treatment with propranolol compared with atenolol (P<0.05). Atenolol and propranolol had no effect on blood glucose, plasma glycerol and FFA. We conclude that wastage in cancer patients is in part explained by increased beta(1) and beta(2) adrenoceptor activity, in part secondary to elevated cardiovascular activity as a result of anaemia, loss of cardiac contractile capacity and altered host metabolism. PMID- 10708934 TI - Low alpha-linolenic acid content of adipose breast tissue is associated with an increased risk of breast cancer. AB - Data derived from experimental studies suggest that alpha-linolenic acid may have a protective effect in breast cancer. Observations obtained from epidemiological studies have not allowed conclusions to be drawn about a potential protective effect of dietary alpha-linolenic acid on breast cancer, possibly because of methodological issues. This case-control study conducted in an homogeneous population from a central area in France was designed to explore the hypothesis that alpha-linolenic acid inhibits breast cancer, using fatty acid levels in adipose breast tissue as a biomarker of past qualitative dietary intake of fatty acids. Biopsies of adipose breast tissue at the time of diagnosis were obtained from 123 women with invasive non-metastatic breast carcinoma. 59 women with benign breast disease served as controls. Individual fatty acids were analysed by capillary gas chromatography. An unconditional logistic regression model was used to obtain odds ratio estimates whilst adjusting for age, menopausal status and body mass index (BMI). No association was found between fatty acids (saturates, monounsaturates, long-chain polyunsaturates n-6 or n-3) and the disease, except for alpha-linolenic acid which showed an inverse association with the risk of breast cancer. The relative risk of breast cancer for women in the highest quartile of adipose breast tissue alpha-linolenic acid level was 0.36 (95% confidence interval=0.12-1.02) compared with those in the lowest quartile (P trend=0.026), suggesting a protective effect of alpha-linolenic acid in the risk of breast cancer. The effects of dietary alpha-linolenic on the risk of breast cancer warrant further study. PMID- 10708935 TI - Chronotherapy with 5-fluorouracil, folinic acid and carboplatin for metastatic colorectal cancer; an interesting therapeutic index in a phase II trial. AB - The aim of this study was to develop a phase II study gauging the contribution of a daily low-dose of carboplatin combined with 5-fluorouracil (5-FU) and folinic acid (FOL) in a chronotherapy schedule for advanced colorectal cancer patients. 60 patients with metastatic colorectal cancer were included in a phase II trial in which 50% of patients had received prior chemotherapy of up to three regimens. Treatment consisted of a combination of 5-FU (700 mg/m(2)/day) and FOL (300 mg/m(2)/day) infused from 10.00 pm to 10. 00 am peaking at 4.00 am with carboplatin infused from 10.00 am to 10.00 pm at 40 mg/m(2)/day with a peak at 4.00 pm. 4-day courses were repeated every 2 weeks in an ambulatory setting with a programmable pump. Patients experienced excellent tolerance (grades III-IV%): diarrhoea, 8.1; nausea/vomiting, 4.8; mucositis, 3.2; skin or neurological 1.7; granulocytes 29.0; platelets and haemoglobin (Hb) 9.7. Major tumour responses were observed in 47% of cases, 4 complete response (CR), 24 partial response (PR); 3 CR and 6 PR (69%) were recorded in 13 previously untreated patients; 11 (18%) underwent subsequent surgical resection of residual metastases. Median survival was 14.6 months with 22% patients surviving over 2 years (35% survival for responders versus 0 for non-responders). In conclusion, this chronotherapy determined administration of 5-FU/FOL and carboplatin yielded an excellent therapeutic index for the combination and warrants further evaluation in the first-line treatment of metastatic colorectal cancer. PMID- 10708936 TI - p53 and Bcl-2 as significant predictors of recurrence and survival in rectal cancer. AB - The aim of this study was to evaluate the prognostic value of p53 nuclear accumulation and Bcl-2 expression after curative surgery for rectal cancer. Immunohistochemistry was performed using monoclonal antibodies (MAb) (DO-1 for p53; anti-human Bcl-2 MAb, clone 124, for Bcl-2) on formalin-fixed, paraffin embedded tissues of 160 rectal carcinomas (UICC stages I-III), and results were compared with data from the prospective registry of rectal cancer by univariate and multivariate logistic regression model focusing specifically on recurrence. Survival was calculated by the Kaplan-Meier method and proportional hazards model. p53 nuclear accumulation was documented in 39% (n=63) of tumours and was associated with a higher incidence of tumour progression (local or distant recurrence) and poorer disease-free survival (P<0.0001). Bcl-2 expression was detected in 29% (n=47), and was associated with longer disease-free survival and lower incidence of recurrence (P<0.0086). Multivariate logistic regression analysis demonstrated that gender (P=0.0136), UICC stage (P=0.0002), p53 expression (P=0.0002) and Bcl-2 expression (P=0. 0243) were independent factors predictive of recurrence. The proportional hazards model identified p53 (P=0.0009), UICC stage (P=0.0480), gender (P=0.0049), but not Bcl-2 (P=0.1503), as independently related to disease-free survival. Looking at the p53/Bcl-2 subgroups, the poorest prognosis was observed in the p53+/Bcl-2- subgroup, whereas patients whose tumours were p53-/Bcl-2+ had the best prognosis (P<0.0001). Immunohistochemical assessment of both p53 and Bcl-2 status may be valuable in predicting recurrence and survival after curative surgery for rectal cancer. Therefore, they play a role as prognostic factors in rectal cancer. p53 is a stronger predictor of prognosis than Bcl-2. PMID- 10708938 TI - Weekly alternating combination chemotherapy for good prognosis AIDS-related lymphoma. AB - Early studies reported that the major adverse prognostic factors in AIDS-related non-Hodgkin's lymphoma (ARL) are low CD4 cell count, prior AIDS defining diagnosis and poor performance status. Since 1989 we have adopted a prognosis stratified approach for ARL. In this study, we identified a group of good prognosis patients. These patients with one or no adverse prognostic factors were treated with alternating weekly chemotherapy using the bleomycin, etoposide, vincristine, methotrexate, prednisolone/cyclophosphamide, doxorubicin (BEMOP/CA) schedule (Bower M, Block C, Gulliford T, et al. Cancer Chemother Pharmacol 1995, 38, 106-109). Modifications to the regimen with the aim of reducing toxicity were a briefer duration (12 weeks) and the addition of prophylaxis against pneumocystis and mycobacteria. Intrathecal methotrexate was administrated fortnightly to patients with Burkitt's histology, meningeal involvement or base of skull disease. 30 patients were treated, including 5 with prior AIDS, 3 with ECOG status 3 and 1 with CD4 <100/microl. The mean age was 40 (range 22-60 years), the median CD4 cell count at ARL diagnosis was 262/microl (range 44 588/microl). The International non-Hodgkin's lymphoma (NHL) prognostic factors project classifications were low risk 8 (maximum 5.4 years) (27%), low intermediate risk 6 (20%), high-intermediate risk 11 (37%) and high risk 5 (17%). The median follow-up was 2.1 years. 3 patients withdrew from treatment within 2 weeks due to toxicity, 2 patients died within 2 weeks of starting chemotherapy. The major toxicity was myelosuppression with 14 patients developing grade 4 neutropenia. The 2-year overall survival rate is 46% (95% confidence interval (CI)=27-65%), and lymphoma-specific survival at 2 years is 59% (95% CI: 40-78%). For selected patients with good prognosis ARL 12 weeks BEMOP/CA therapy produces good overall survival at 2 years. PMID- 10708937 TI - Prognostic value of the expression of E-cadherin and beta-catenin in bladder cancer. AB - The purpose of this study was to assess the prognostic effect of the expression of E-cadherin, beta-catenin and CD44 adhesion molecules in bladder carcinoma. 22 superficial and 18 invasive bladder tumour samples were studied by immunohistochemistry. The median follow-up was 24 months (range: 1-50 months). Loss of E-cadherin and beta-catenin immunoreactivity was found in 14 (35%) and 17 (43%) tumours, respectively, and was significantly associated with invasiveness, high grade and p53 overexpression. There was no correlation between CD44 variant expression and clinicopathological findings. Loss of E-cadherin expression was an independent predictor of poor survival in a multivariate analysis, when assessed with age, grade, stage and p53 status (hazards ratio adjusted (HRa)=4.45 [95% confidence interval (CI), 1.06-18.63]). This effect was particularly augmented in patients with invasive bladder cancer. When expression of E-cadherin and beta catenin were evaluated simultaneously, loss of immunoreactivity of both proteins was a strong predictor of poor survival (HRa=13.06 [95% CI, 0.95-178.55]). The same pattern was found when progression-free survival in relation to these variables was assessed. In conclusion, assessment of E-cadherin and beta-catenin immunoreactivity may be a useful prognostic marker in bladder cancer complementary to established prognostic factors. PMID- 10708939 TI - Bone marrow transplantation for children with acute myelogenous leukaemia in the first complete remission. AB - Of 52 children aged 9 months to 16 years old with acute myelogenous leukaemia (AML) in first complete remission undergoing bone marrow transplantation at our institution, 31 received allogeneic transplants (allo-BMT) and 21 received autologous transplants (ABMT). Initial induction and consolidation chemotherapy were not uniform. BMT was performed at a median of 7 months (range: 2.5 to 22.5 months) from the diagnosis. Conditioning included chemotherapy (n=43: 4 x 4 mg/kg of busulfan and 3 x 60 to 70 mg/m(2) of melphalan) or total body irradiation (12 Gy) plus chemotherapy (n=9). Graft-versus-host disease (GVHD) prophylaxis in allo BMT cases consisted of methotrexate +/- cyclosporin A. Unpurged marrow was used in ABMT cases. All patients showed sustained engraftment. Amongst allograft cases, acute or chronic GVHD developed in 7 patients each (23%). 8 patients (15%) died (5 with allo-BMT, 3 with ABMT), including transplant-related mortality in 3 of the allo-BMT patients. 7 patients had relapses (3 with allo-BMT, 4 with ABMT). As of June 1999, 43 patients are alive and well 13 to 160 months after BMT (median, 71), with 5-year disease-free survival rates after BMT of 84% for allo BMT, 81% for ABMT and 83% altogether. Although the presented data are based on a retrospective evaluation, we consider BMT for childhood AML during first complete remission an effective treatment for eradicating leukaemia. PMID- 10708940 TI - Non-seminomatous ovarian germ cell tumours in children. AB - In this study, we report the results of two consecutive protocols. TGM 55 and TGM 90, of the Societe Fran?aise d'Oncologie Pediatrique ( SFOP) for patients with non-seminomatous germ cell tumours of the ovary and analyse the rationale for surgical indications. neoadjuvant or adjuvant chemotherapy. TGM 55 and 90 both utilised six drugs, bleomycin, cyclophosphamide, vinblastine, dactinomycin, etoposide and either cisplatin (TGM 55) or carboplatin TGM 90). Chemotherapy was given in ease of unresectable or incompletely resected tumour. Patients who had a complete resection of a localised tumour underwent expectant management and were only treated if progression occurred. 63 patients aged less than 18 sears old were enrolled between January 1955 and December 1994. 49 patients had alpha fetoprotein (alphaFP) +/- beta-human chorionic gonadotropic hormone (betaHCG) secreting tumours and 14 had immature teratomas. Median follow-up for surviving patients is 60 months (range: 19-154). The 5-year overall survival is 85% +/- 5%. 13 out of 14 patients (93%) with immature teratoma are alive, including 3 of 4 patients (75%) who received chemotherapy for advanced disease. 41 patients (54%) with secreting tumours are alive, including 2 patients who required salvage treatment. Most failures occurred amongst patients with high initial alphaFP secretion ( > 15,000 ng/ml). 39 of 41 survivors (95%) in thc non-teratoma group had conservative surgery, allowing the possibility of future pregnancy. High cure rate can he achieved with a conservative approach in non-seminomatous germ cell tumour of the ovary. Whenever possible, fertility should he preserved during the primary operation in children suffering from these tumours. PMID- 10708941 TI - Long-term survival in Hodgkin's disease patients. A comparison of relative survival in patients in trials and those recorded in population-based cancer registries. AB - The prognosis of Hodgkins disease (HD) has improved during the last 30 years. This study was planned to analyse long-term survival of LID patients and to compare survival rates estimated from clinical trials and population-based data. Individual data were analysed on 2,755 adult HD patients entering randomised clinical trials of the British National Lymphoma Investigation BN LI) between 1970 and 1987, and 5,064 patients with HD incident 1978-1984 recorded in the UK population-based cancer registries participating in the EUROCARE study. Relative survival of Hodgkins disease patients allowing for mortality expected from general population rates was analysed by a proportional hazards regression model including covariates. Although relative mortality decreased with longer follow up, it was still significantly positive at 9-10 years after diagnosis in both the clinical trials and the population-based data sets. Relative mortality was worse for late stage than for early stage patients even at 10-15 years after first treatment (BNLI data). Whereas 10-year relative survival was identical in trials and population-based patients at ages under 45 years (> 69%), it was much higher in BNLI older patients than in the population-based patients. In the older age group (65-74 years) the BNLI patients had 39% relative survival whilst for the population-based patients it was only 27%, Generalisation of clinical trials results to the general population must be done with caution, especially for older patients. PMID- 10708942 TI - Prognostic factors after curative resection for gastric cancer. A population based study. AB - The aim of this study was to document patterns of survival after resection for cure for gastric cancer in a well-defined population. A population-based series of 649 gastric cancers resected for cure between 1976 and 1995 in a 494000 population, was used. Resection for cure was performed in 44.4% of the diagnosed cases. This proportion increased from 36.8% (1976-1979) to 45.0% (1992-1995) (P=0.03) whilst operative mortality decreased from 18.3 to 12.7% (P=0.003). The overall crude 5-year survival rate (excluding operative mortality) was 32.6% (95% confidence interval (CI) 28.7-36. 5) and the corresponding relative survival rate was 40.9%. Prognosis did not improve during the study period. Stage at diagnosis was the most important prognostic factor, the 5-year relative survival rate being 81.2% (+/-5.9) in TNM stage IA, 76.9% (+/-8.0) in stage IB, 50. 4% (+/-4.6) in stage II, 24.4% (+/-3.7) in stage IIIA, 5.6% (+/-3.2) in stage IIIB and 5.2% (+/- 2.2) in stage IV. Stage at diagnosis, age, subsite and macroscopic type of growth were independent prognostic factors, in a multivariate relative survival model. Earlier detection or development of an effective adjuvant therapy could contribute to improvement in prognosis. PMID- 10708943 TI - An antisense oligonucleotide targeting the alphaV integrin gene inhibits adhesion and induces apoptosis in breast cancer cells. AB - The aim of this study was to show the anti-adhesive potential of an antisense oligodeoxynucleotide (ODN) approach when designed to suppress the cellular function of the alphaV integrin subunit in breast cancer cells. The alphaV integrins play major roles in favouring breast cancer spreading. In this study, we inhibited alphaV subunit synthesis in the human breast carcinoma cell line, MDA-MB231, by a partially phosphorothioated antisense oligodeoxynucleotide (5543 ODN). The alphaV antisense 5543-ODN reduced alphaV, but not actin, mRNA transcription and protein expression by 55% and 65% respectively (1 microM, 72 h). Control sense and mismatch reagents were inactive. The antisense, but not the sense and mismatch, 5543-ODN induced dose- and time-dependent inhibition of MDA MB231 adhesion to serum, vitronectin, fibrinogen and fibronectin substrates but was inactive on adhesion to laminin. Thus, the alphaV integrin was located in adhesion structures, which were disrupted by treatment with the alphaV antisense 5543-ODN. Antisense treated cells also showed evidence of programmed cell death with the appearance of apoptotic bodies. MDA-MB231 cells express a mutant form of the pro-apoptotic factor p53; however, no changes in the expression of p53 were observed by Western blotting. Immunofluorescence did reveal an increased nuclear translocation of p53 suggesting activation of the protein, but such a translocation did not lead to significant changes in either the expression of the cyclin dependent kinase inhibitor, p21(WAF1/CIP1) the cell survival factor Bcl-2 or the pro-apoptotic factor Bax. PMID- 10708944 TI - In vitro and in vivo effects of a cyclic peptide with affinity for the alpha(nu)beta3 integrin in human melanoma cells. AB - Expression of the integrin alpha(nu)beta3 has been shown to be associated with increasing metastatic potential in malignant melanoma. It also has a functional role on vascular endothelial cells during angiogenesis. The cyclic oligopeptide cRGDfV is known to bind with high affinity to alpha(nu)beta3. We have investigated the cellular effects of cRGDfV on a panel of human melanoma cell lines in vitro and also on the A375 melanoma cell line growing as xenografts in nude mice. cRGDfV is a potent inhibitor of alpha(nu)beta3-mediated cell adhesion, however, we have found no convincing evidence that integrin ligation by cRGDfV induces apoptosis in melanoma cell lines. However, cRGDfV when administered subcutaneously into nude mice did inhibit the growth of A375 melanoma xenografts. Histological examination of the tumours indicated that this effect was primarily one of angiogenesis inhibition. The results suggest that agents which target the alpha(nu)beta3 integrin may have a useful role as anti-angiogenesis agents in clinical oncology, but that they may not exert a direct effect on alphavbeta3 expressing tumour cells. PMID- 10708945 TI - Effect of 1-(gamma)linolenyl-3-eicosapentaenoyl propane diol on the growth of human pancreatic carcinoma in vitro and in vivo. AB - Essential fatty acids such as (gamma)linolenic (GLA) and eicosapentaenoic (EPA) acids have been proposed as anticancer drugs. The aim of this study was to test the effect of a lipid emulsion containing both GLA and EPA in a novel chemical formulation of 1-(gamma)linolenyl-3-eicosapentaenoyl propane diol on the growth of human pancreatic carcinoma in vitro and in nude mice. This compound had a dose dependent growth-inhibitory effect on human pancreatic cancer cell lines MIA PaCa 2 and Panc-1 in vitro. The concentration necessary for 50% growth inhibition was 25 micromol/l for MIA PaCa-2 and 68 micromol/l for Panc-1 (95% CI 20-29 and 59-77 micromol/l respectively). Nude mice bearing subcutaneous pancreatic tumours produced with the MIA PaCa-2 cell line were treated with the maximum tolerated dose (6.75 mg GLA and 7.3 mg EPA per g of body weight) administered over 10 days by daily intravenous (i.v.) bolus injections. No antitumour effect or major alteration in tumour lipid fatty acid composition was seen in comparison with control animals. Concurrent treatment with parenteral iron (iron saccharate, 5 microg/gram body weight daily) did not make a significant difference. Further improvements in fatty acid delivery mechanisms are necessary before they can become useful anticancer agents. PMID- 10708946 TI - Regulation of cellular glutathione modulates nuclear accumulation of daunorubicin in human MCF7 cells overexpressing multidrug resistance associated protein. AB - Multidrug resistance (MDR) is frequently associated with the overexpression of P glycoprotein (Pgp) and/or multidrug resistance associated protein (MRP1), both members of the ABC superfamily of transporters. Pgp and MRP1 function as ATP dependent efflux pumps that extrude cytotoxic drugs from tumour cells. Glutathione (GSH) has been considered to play an important role in the MRP1 mediated MDR. In our study, we examined the effects of buthionine sulphoximine (BSO), an inhibitor of GSH biosynthesis, on the nuclear accumulation of daunorubicin (DNR), in etoposide (VP16) and doxorubicin (ADR) resistant MCF7 cell lines, overexpressing respectively MRP1 (MCF7/VP) and Pgp (MCF7/ADR). The study of DNR transport was carried out using scanning confocal microspectrofluorometry. This technique allows the determination of the nuclear accumulation of anthracyclines in single living tumour cells. Treatment of MCF7/VP cells with BSO increased the sensitivity of these cells to DNR whilst the cytotoxicity of the drug in MCF7/ADR cells remained unchanged. In MCF7 resistant cells treated with BSO, their GSH level decreased as observed by confocal microscopy. DNR nuclear accumulation in MCF7/VP cells was increased by BSO whereas in MCF7/ADR cells BSO was unable to significantly increase the DNR nuclear accumulation. These data suggest a requirement for GSH in MRP1-mediated resistance whilst the nuclear efflux of GSH conjugates is probably not the primary mechanism of Pgp-mediated MDR. Finally, BSO might be a useful agent in clinical assays for facilitating detection of MRP1 expression. PMID- 10708947 TI - Introduction PMID- 10708948 TI - The Smad pathway. AB - Transforming growth factor-beta superfamily member signals are conveyed through cell-surface serine/threonine kinase receptors to the intracellular mediators known as Smads. Activation of Smads causes their translocation from the cytoplasm to the nucleus where they function to control gene expression. In this review we will focus on proteins that modulate Smad activity, including SARA, for Smad Anchor for Receptor Activation, which functions during the initiation of signalling and on components of the ubiquitin-proteasome pathway, such as Smurf1, which can negatively regulate Smad signalling. In addition, we will summarize recent findings on the role of Smads as transcriptional co-modulators. PMID- 10708949 TI - TGF-beta signaling by Smad proteins. AB - Smads are signal transducers for the members of the transforming growth factor beta (TGF-beta) superfamily. Bone morphogenetic proteins (BMPs) and their receptors induce differentiation of C2C12 cells into osteoblast-like cells. Using an adenoviral expression vector system, we showed that receptor-regulated Smads (R-Smads) activated by BMPs can induce the differentiation of C2C12 cells. Inhibitory Smads (I-Smads) interfere with the osteoblast differentiation of C2C12 cells by preventing the nuclear translocation of R-Smads. After translocation into the nucleus, Smad oligomers regulate the transcription of target genes through binding to DNA directly, interaction with other DNA binding proteins, and recruitment of transcriptional co-activators or co-repressors. Through interaction with different transcription factors and transcriptional co activators or co-repressors, Smads may exhibit specific effects in various cell types. PMID- 10708950 TI - Role of Ras and Mapks in TGFbeta signaling. AB - Normal signaling by TGFbeta, in the absence of serum or exogenous factors, involves a rapid activation of Ras, Erks, and Sapks in proliferating cultures of TGFbeta-sensitive untransformed epithelial cells and human carcinoma cells. Expression of either RasN17 or dominant-negative (DN) MKK4, or addition of the MEK1 inhibitor PD98059, can block the ability of TGFbeta to induce AP-1 complex formation at the TGFbeta(1) promoter and to autoinduce its own production. The primary components present in this TGFbeta-stimulated AP-1 complex are JunD and Fra-2, although c-Jun, and possibly Fos B, may also be present. While there are two potential Smad binding elements (SBE's) in the TGFbeta(1) promoter, supershift assays suggest that at least one of these does not bind Smad4, and the other is unable to bind factors activated by TGFbeta. In contrast, TGFbeta autoinduction is Smad3-dependent, as DN Smad3 inhibits the ability of TGFbeta to stimulate TGFbeta(1) promoter activity. Our results indicate that TGFbeta can activate both the MKK4/Sapk and MEK/Erk pathways, through Ras and TGFbeta R(I) and R(II), to induce TGFbeta(1) production; Smad4 does not appear to be involved, and Smad3 appears to function independently of this Smad4. We also demonstrate that activation of the Ras/Mapk pathway by TGFbeta positively modulates Smad1 signaling-pathway activation by TGFbeta. In addition, Smad1 could enhance TGFbeta activation of the SBE reporter SBE-luc and this effect could be blocked by co expression of a DN TGFbeta R(I) receptor or by the MEK1 inhibitor PD98059. This cross-talk between the MEK/Erk and Smad1 pathways was mediated through the four Erk consensus phosphorylation sites in the linker region of Smad1. Mutation of these sites resulted in a loss of the ligand-dependence of both Smad1-Smad4 interactions and nuclear accumulation of Smad1, as well as a loss of the ability of Smad1 to enhance TGFbeta-mediated SBE activation. Our results provide evidence that Erk-mediated phosphorylation of Smad1 in response to TGFbeta is critical for regulating Smad1 subcellular localization; this may be a key determinant in maintaining TGFbeta-dependent transcriptional activation. PMID- 10708951 TI - The Smads: transcriptional regulation and mouse models. AB - The field of transforming growth factor-beta (TGF-beta) signaling sees periodic discoveries that revolutionize our thinking, redirect our experiments, and peak our excitement. One of the first such discoveries was less than a decade ago: the molecular cloning of the type I and type II TGF-beta receptors. This breakthrough defined a novel family of serine/threonine kinase receptors, which led to the description of an ever-expanding superfamily. The discovery of how these receptors are grouped on the cell surface, bind TGF-beta and are activated by specific phosphorylation events further defined the uniqueness of this system in comparison to other families of growth factor receptors. Now, once again, the TGF beta field has been revolutionized. This time, the discovery is the Smad family of proteins. Although one can hardly imagine TGF-beta without the Smads, the cloning of the Smads and their implication in TGF-beta signaling was only four years ago. Since that time, great advances have been made in our understanding of the Smads as transcription factors, which are activated by receptor mediated phosphorylation. In addition, animal models for a loss of Smad function have provided insight into the role of specific Smads in a variety of physiologic systems. The Smad field has been growing exponentially. A comprehensive review of all aspects of the Smads, therefore, would be beyond the scope of a single review. Instead, this review highlights some of the general aspects of Smad function, and then focuses on the role of specific Smad family members in transcriptional regulation, animal physiology, and disease processes. PMID- 10708952 TI - Functions of mammalian Smad genes as revealed by targeted gene disruption in mice. AB - The Smad genes are the intracellular mediators of TGF-beta signals. Targeted mutagenesis in mice has yielded valuable new insights into the functions of this important gene family. These experiments have shown that Smad2 and Smad4 are needed for gastrulation, Smad5 for angiogenesis, and Smad3 for establishment of the mucosal immune response and proper development of the skeleton. In addition, these experiments have shown us the importance of gene dosage in this family, as several of its members yielded haploinsufficiency phenotypes. These include gastrulation and craniofacial defects for Smad2, accelerated wound healing for Smad3, and the incidence of gastric cancer for Smad4. Combinatorial genetics has also revealed functions of Smads in left/right isomerism and liver development. PMID- 10708953 TI - Activation of latent TGF-beta by thrombospondin-1: mechanisms and physiology. AB - Regulation of the activation of latent TGF-beta is essential for health as too much or too little TGF-beta activity can have serious, deleterious consequences. The processes that control conversion of the precursor to the biologically active form of TGF-beta in vivo are not well characterized. We have identified a mechanism for the activation of latent TGF-beta that involves binding of the secreted and extracellular matrix protein, thrombospondin-1 (TSP-1), to the latent precursor. Specific sequences in TSP-1 and in the precursor portion (the latency associate peptide-LAP) have been determined to be essential for activation of latent TGF-beta by TSP-1. It is thought that binding of TSP-1 to the latent complex induces a conformational rearrangement of the LAP in such a manner as to prevent the LAP from conferring latency on the mature domain of TGF beta. A TSP-dependent mechanism of activation may be locally important during wound healing and in post-natal development of epithelial structures. The possible involvement of TSP-1 in TGF-beta activation during several disease processes is also discussed. PMID- 10708954 TI - TGF-beta influences the life and death decisions of T lymphocytes. AB - TGF-beta is a powerful mediator of immune cell phenotype and function. In TGF beta1 homozygous null mice, aberrant regulation of the immune response culminates in lethal cardiopulmonary inflammation. In dissecting the underlying mechanisms leading to the attack of self, a role for TGF-beta1 in controlling apoptosis and T cell selection patterns was uncovered. Increased levels of apoptosis and TCR mediated cell death disrupted normal negative and positive T cell selection in the thymus. Moreover, in peripheral T cell populations, increased T lymphocyte death was associated with increased expression of apoptosis-inducing receptors. Persistent activation of T cells engendered unchecked apoptosis which, rather than reducing, further exacerbated, tissue inflammation due to the absence of TGF beta1. TGF-beta, normally generated by macrophages during clearance of apoptotic cells contributes to dampening of inflammatory sequelae associated with phagocytosis. Collectively, these data demonstrate a pivotal role for TGF-beta in multiple stages of T cell apoptosis, selection, activation and clearance. PMID- 10708955 TI - Murine models define the role of TGF-beta as a master regulator of immune cell function. AB - Many members of transforming growth factor-beta (TGF-beta) superfamily, including not only TGF-beta, but also the activins, and bone morphogenetic proteins (BMPs), have been demonstrated to affect the development and function of immune cells. From the proliferation and differentiation of pluripotent stem cells, to the activation and migration of mature lymphoid and myeloid lineages, the TGF-betas have been recognized for their ability to modulate the manner in which such cells respond to stimuli in their environment. Recent studies involving disruption of this pathway in genetically engineered mice now emphasize the importance of this activity and validate functional models predicted by in vitro studies. Phenotypic differences between mice harboring mutations in the TGF-beta1 ligand and the TGF beta receptor-activated signaling intermediate Smad3 are presented and serve to highlight the valuable role of these in vivo genetic tests of function. PMID- 10708956 TI - When engineered to produce latent TGF-beta1, antigen specific T cells down regulate Th1 cell-mediated autoimmune and Th2 cell-mediated allergic inflammatory processes. AB - To determine whether T cells which produce large amounts of latent TGF-beta1 are capable of down-regulating autoimmune and allergic disease, myelin basic protein (MBP)-specific and ovalbumin (OVA)-specific BALB/c cloned Th1 cells were transduced with cDNA for murine TGF-beta1 by coculture with fibroblasts producing a genetically engineered retrovirus. The transduced MBP-specific Th1 cells were found to lose the capacity to provoke EAE in BALB/c mice, and to gain instead the ability to protect against EAE in (SJLxBALB/c) F1 mice immunized with proteolipid protein (PLP). This protective effect was not obtained with OVA-specific TGF beta1 transduced Th1 cells. The transduced OVA-specific Th1 cells did protect against airway hyperreactivity induced by Th2-cell mediated responses to inhaled OVA. This effect was again antigen specific and it also could not be obtained with untransduced OVA-specific Th1 cells. In both cases these effects of antigen specific TGF-beta1 transduced T cells were nullified by administration of neutralizing anti-TGF-beta mAb. Thus, the antigen specificity of the cloned T cells allows the site-specific local delivery of therapeutic active TGF-beta1 to both Th1 and Th2 cell-mediated inflammatory infiltrates. PMID- 10708957 TI - Co-activation of TGF-ss and cytokine signaling pathways are required for neurotrophic functions. AB - This article summarizes and interprets recent data from our laboratories suggesting that transforming growth factor-ss (TGF-ss1, -ss2, -ss3) is essentially required, in vitro and in vivo, for the neurotrophic signaling of glial cell line-derived neurotrophic factor (GDNF). TGF-ss, which is synthesized by and released from neurons, also synergizes with neurotrophins and members of the neurokine and fibroblast growth factor families by increasing their efficacies. However, when applied to purified neuron populations without other factors being added, TGF-ss does not promote survival or differentiation. Together, these data suggest that neither TGF-ss nor GDNF fulfil essential criteria of a typical neurotrophic factor, as e.g. nerve growth factor (NGF). Moreover, the neurotrophic activity of NGF and other classic neurotrophic factors is apparently based, to a significant extent, on their co-operativity with TGF ss. Mechanisms, by which TGF-ss generates neurotrophic effects and synergizes with other cytokines are beginning to emerge. Recruitment and/or stabilization of receptors and cross-talks at different levels of signal transduction are likely to be implied in generating the neurotrophic potential of the TGF-ss/cytokine synergisms. Together, these data outline a novel role of TGF-ss in a key event of nervous system development, ontogenetic neuron death. Conceptually more important, however, may be the broadening of the neurotrophic factor concept, which now has to imply the possibility that two cytokines, each being ineffective by itself, become neurotrophically active when acting in concert. PMID- 10708958 TI - TGF-betas and TGF-beta receptors in atherosclerosis. AB - Based on diverse evidence in animals and humans, it has been hypothesized that atherosclerosis, and other injury-induced hyperplasias such as restenosis, may result from a failure in endogenous inhibitory systems that normally limit wound repair and induce regression of wound repair cells. A key defect in one of these inhibitory pathways, the TGF-beta system, has been identified and characterized in both animal models and in human lesions and lesion-derived cells. Cells derived from human lesions are resistant to the antiproliferative and apoptotic effects of TGF-beta, while their normal counterparts from the vascular media are potently inhibited and killed. Both cell types increase PAI-1 production, switch actin phenotypes in response to TGF-beta1, and produce similar levels of TGF-beta activity. Membrane cross-linking of (125)I-TGF-beta1 indicates that normal human SMC express Type I, II and III receptors. The Type II receptor is strikingly decreased in lesion cells, with little change in the Type I or III receptors. RT PCR confirmed that the Type II TGF-beta1 receptor mRNA is reduced in lesion cells. Subsequent analysis of human lesion vs normal tissues confirmed that the Type I receptor was consistently present in the lesion, while the Type II receptor was much more variable, and commonly absent in both coronary artery and carotid artery lesions. Transfection of the Type II receptor into lesion cells partially restores the growth inhibitory response to TGF-beta1, implying that signaling remains intact. A subset of patients, and cells derived from their lesions, exhibit acquired mutations in the Type II receptor that would explain their resistance, though the majority of cells are resistant without obvious mutational defects. Thus, it is currently being tested whether transcriptional defects or abnormalities in receptor processing may explain the low levels of the Type II receptor. Because TGF-beta1 is overexpressed in fibroproliferative vascular lesions, receptor-negative cells would be allowed to grow in a slow, but uncontrolled fashion, while overproducing extracellular matrix components. PMID- 10708959 TI - TGF-beta in diabetic kidney disease: role of novel signaling pathways. AB - Diabetic nephropathy is the leading cause of end-stage renal disease in the United States and is a major contributing cause of morbidity and mortality in patients with diabetes. Despite conventional therapy to improve glycemic and blood pressure control the incidence of diabetic nephropathy is reaching epidemic proportions worldwide. As the major pathologic feature of diabetic nephropathy is diffuse mesangial matrix expansion, the pro-sclerotic cytokine transforming growth factor-beta, TGF-beta, is a leading candidate to mediate the progression of the disease. Numerous studies have now demonstrated that TGF-beta is a key factor in experimental models of diabetic kidney disease as well as in patients with diabetic nephropathy. Recent studies have begun to explore the mechanisms by which TGF-beta is stimulated by high glucose and how TGF-beta exerts its matrix stimulating effects on renal cells. TGF-beta may also be involved in mediating the vascular dysfunction of diabetic kidney disease via its effects on the key intracellular calcium channel, the inositol trisphosphate receptor (IP(3)R). As there is substantial evidence for a cause and effect relationship between upregulation of TGF-beta and the progression of diabetic kidney disease, future studies will seek to establish specific targets along these pathways at which to intervene. PMID- 10708960 TI - Loss of Smad3 modulates wound healing. AB - TGF-beta plays a central and critical role in tissue repair. The recent identification of TGF-beta signal transduction pathways involving Smad proteins has now made it possible to explore their contribution to the activities of TGF beta in vivo. Both Smad3 and its closely related homolog Smad2 act as latent nuclear transcriptional activators and mediate intracellular signaling by TGF betas and activin, each of which regulates cellular functions pivotal to cutaneous wound healing. Mice null for Smad3 (Smad3(ex8/ex8)) survive into adulthood and show accelerated cutaneous wound healing characterized by an increased rate of re-epithelialization and a reduced local inflammatory infiltrate. These data implicate Smad3 in specific pathways of tissue repair and suggest that it could be a target for the development of therapeutic strategies to modulate wound healing. PMID- 10708961 TI - TGF-beta in blood: a complex problem. AB - The cytokine transforming growth factor-beta (TGF-beta) was initially purified from human platelets, a rich source of this protein. In addition to platelets, TGF-beta1 is also found in other blood fractions, including plasma and the circulating leukocytes. However, more than 15 years after the initial isolation of TGF-beta1, there remains no consensus on how much TGF-beta1 is present in normal human plasma. Here we review the difficulties associated with measuring TGF-beta concentrations in complex biological fluids, and discuss the current state of knowledge on the distribution of TGF-beta isoforms in various blood fractions as well as the nature of the TGF-beta-containing protein complexes. PMID- 10708962 TI - Gastro-intestinal tumorigenesis in Smad4 mutant mice. AB - The SMAD4 gene plays a key role in the TGF-beta signaling pathway. We inactivated its mouse homolog Smad4. The homozygous mutants were embryonically lethal, whereas the heterozygotes were viable and fertile. Although young heterozygotes appeared normal, old mice developed gastric and duodenal polyps similar to human juvenile polyps characterized by abundant stroma and eosinophilic infiltrations. These data are consistent with the reports that a subset of human juvenile polyposis kindreds carry germline mutations in the SMAD4 gene. We then introduced the Smad4 mutation into the Apc(Delta716) knockout mice, a model for human familial adenomatous polyposis. Because both Apc and Smad4 are located on mouse chromosome 18, we constructed by meiotic recombination compound heterozygotes carrying both mutations on the same chromosome. In such mice, intestinal polyps developed into more malignant tumors than those in the simple Apc(Delta716) heterozygotes, showing an extensive stromal cell proliferation and strong submucosal invasion. These results indicate that mutations in SMAD4 play a significant role in the malignant progression of colorectal tumors. PMID- 10708963 TI - Molecular mechanisms of inactivation of TGF-beta receptors during carcinogenesis. AB - Signals from the TGF-betas are mediated by the TGF-beta receptors and their substrates, the Smad proteins. Inactivation of either of the two transmembrane serine/threonine kinases called the TGF-beta type I and type II receptors is now known to underlie a wide variety of human pathologies including, especially carcinogenesis. Numerous studies have now demonstrated that the TGF-beta receptor complex and its downstream signaling intermediates constitute a tumor suppressor pathway. We review here a specific pathway of mutational inactivation of the TGF beta type II receptor resulting from microsatellite instability and demonstrate that, by contrast, the most common mechanism of loss of expression of the TGF beta type II receptor involves transcriptional repression. This provides a new target for therapeutic intervention. PMID- 10708965 TI - The comet assay in eight mouse organs: results with 24 azo compounds. AB - The genotoxicity of 24 azo compounds selected from IARC (International Agency for Research on Cancer) groups 2A, 2B, and 3 were determined by the comet (alkaline single cell gel electrophoresis, SCG) assay in eight mouse organs. We treated groups of four mice once orally at the maximum tolerated dose (MTD) and sampled stomach, colon, liver, kidney, bladder, lung, brain, and bone marrow 3, 8, and 24 h after treatment. For the 17 azo compounds, the assay was positive in at least one organ; (1) 14 and 12 azo compounds induced DNA damage in the colon and liver, respectively, (2) the genotoxic effect of most of them was greatest in the colon, and (3) there were high positive responses in the gastrointestinal organs, but those organs are not targets for carcinogenesis. One possible explanation for this discrepancy is that the assay detects DNA damage induced shortly after administration of a relatively high dose, while carcinogenicity is detected after long treatment with relatively low doses. The metabolic enzymes may become saturated following high doses and the rates and pathways of metabolic activation and detoxification may differ following high single doses vs. low long-term doses. Furthermore, considering that spontaneous colon tumors are very rare in rats and mice, the ability to detect tumorigenic effects in the colon of those animals might be lower than the ability to detect genotoxic events in the comet assay. The in vivo comet assay, which has advantage of reflecting test chemical absorption, distribution, and excretion as well as metabolism, should be effective for estimating the risk posed by azo dyes to humans in spite of the difference in dosage regimen. PMID- 10708964 TI - Aluminium triggers genotoxic adaptation to methyl mercuric chloride and ethyl methane sulfonate, but not to maleic hydrazide in plant cells in vivo. AB - Non-toxic, conditioning doses of aluminium chloride were tested for induction of adaptive response to the genotoxic challenge doses of methyl mercuric chloride (MMCl), maleic hydrazide (MH) and ethyl methane sulfonate (EMS). Embryonic shoot cells of Hordeum vulgare and root meristem cells of Allium cepa were employed as the assay systems. Plant tissues fixed at different recovery hours following the challenge treatments with or without prior Al-conditioning were analyzed for cells with genotoxicity markers that include spindle and/or chromosome aberrations and micronuclei (MNC). The results provided evidence that Al(3+) triggered adaptive response that protected the plant cells from the genotoxicity of MMCl and EMS. Al(3+), however, failed to induce adaptive response against the genotoxicity of MH. A comparison of Al-induced adaptive response with that induced by heavy metals: Cd(2+), Cu(2+), Hg(2+), Ni(2+), Pb(2+), Zn(2+) and oxidative agents: hydrogen peroxide (H(2)O(2)) and paraquat (PQ) pointed to the similarity of Al-adaptive response to that of PQ rather than to other heavy metals or H(2)O(2). Al-induced adaptive response demonstrated in the present study to MMCl and EMS possibly involved antioxidant defense and DNA repair systems, respectively. PMID- 10708966 TI - Rec effect of certain textile dyes in Bacillus subtilis. AB - A large number of compounds are toxic, genotoxic, mutagenic, teratogenic and/or carcinogenic. The genotoxicity of four textile dyes commonly used in India namely Sulphur Red Brown 360 (SRB), Jade Green 2G (JG), Reactofix Turquoise Blue 5GFL (RTB) and Direct Scarlet 4BS (DS) was determined by Bacillus subtilis spore Rec assay, both in the presence and absence of metabolizing activation mixture (S9 mix). Each dye was toxic at higher dose levels. A dose-dependent increase in the depth of growth inhibition zones was observed for all dyes. Zones of inhibition were usually clearer at higher doses of the dyes and with Rec- bacteria, but were translucent with Rec+ bacteria. SRB and DS were toxic to Rec+ and Rec- bacteria. JG was less genotoxic in the absence of S9 mix, however, its genotoxic potential increased in the presence of S9 mix. Reactofix T blue was more genotoxic in the absence of S9 mixture. PMID- 10708967 TI - Mutagenicity of benzo[a]pyrene-deoxyadenosine adducts in a sequence context derived from the p53 gene. AB - Mutations in the human p53 tumor suppressor gene are prominently linked to sporadic cancers in breast, lung and other tissues. Recent research has shown that tobacco-associated cancer in the human lung is related to mutation of the p53 gene mediated by the carcinogen benzo[a]pyrene (BaP), and the mutations are targeted to DNA "hot spots" at specific codons. In order to gain insight into the relation between the structures of the adducts formed by BaP at these sites and their mutagenic activities, we have synthesized site-specifically modified oligo nucleotide adducts of the active BaP diol epoxide metabolite (anti-BaPDE). This manuscript reports on the mutagenic consequences of replication past anti-BaPDE deoxyadenosine adducts located within a sequence context related to codon 157 in exon 5 of the p53 gene. In this sequence context, the adduct derived from the carcinogenic 7R,8S-dihydrodiol 9S,10R-epoxide was much more active as a mutagen than the adduct derived from the noncarcinogenic 7S,8R-dihydrodiol 9R,10S-epoxide and the mutation found most frequently was an A-->G transition. Since previous studies in other sequence contexts have yielded somewhat different findings, these studies further emphasize the key role played by sequence context in determining the mutational properties of carcinogen-DNA adducts. PMID- 10708968 TI - Sequential chromosome aberration analysis following radiotherapy - no evidence for enhanced genomic instability. AB - Chromosome analysis of peripheral blood lymphocytes using block staining was performed on 18 cancer patients who had received fractionated radiotherapy doses totalling 35-80 Gy. Samples were obtained from 13 individuals within 1 year of treatment and thereafter approximately annually up to a maximum of eight times (range: three to eight samples per individual). Sampling of the remaining five patients started later. Frequencies of cells with unstable chromosome aberrations showed a steady decline whereas frequencies of cells with just chromatid aberrations and gaps were initially low and remained so. There was no subsequent rise in any aberrant cell type in later years and thus no suggestion that the radiation exposure had induced a persistent or late manifesting state of genomic instability. PMID- 10708969 TI - Cytogenetic studies of three triazine herbicides. I. In vitro studies. AB - Atrazine, simazine, and cyanazine are widely used pre-emergence and post emergence triazine herbicides that have made their way into the potable water supply of many agricultural communities. Because of this and the prevalence of contradictory cytogenetic studies in the literature on atrazine, simazine, and cyanazine, a series of in vitro experiments was performed to investigate the ability of these three triazines to induce sister chromatid exchanges (SCEs) and chromosome aberrations (CAs) in human lymphocyte cultures. Our results showed that all three triazines failed to produce any significant increases in SCEs or CAs up to the limits of solubility [using 0.5% dimethyl sulfoxide (DMSO)]. Our results are discussed in light of contradictory results in the literature. PMID- 10708970 TI - The somatostatin analogue peptide TT-232 induces apoptosis and chromosome breakage in cultured human lymphocytes. AB - Somatostatin receptors are supposed to be important in the regulation of apoptosis. In this study, we measured apoptosis occurring spontaneously, or induced by the synthetic somatostatin analogue, the peptide TT-232. We examined isolated human peripheral blood lymphocytes (PBL) from 32 nurses exposed bedside to cytostatic drugs, 12 chronic lymphoid leukaemia (CLL) patients prior to treatment, and 19 unexposed, healthy donors without anamnestic occupational exposure to genotoxic agents. Cells were stimulated by phytohaemagglutinin-P (PHA) and cultured for 69 h with or without 15 microg/ml TT-232, respectively. Cell kinetic parameters and apoptosis were determined by flow cytometry after staining with FITC-labeled anti-BrdU and propidium iodide (PI) and the results on spontaneous and peptide-induced apoptosis were compared with the obtained chromosome aberration frequencies (CA). The peptide TT-232 unexpectedly induced chromosome breakage in addition to apoptosis. The mean spontaneous apoptotic fractions were 6.65+/-0.89%, 6.46+/-0. 53%, and 3.07+/-0.57%, and the mean CA yields in the samples without TT-232 were 1.74+/-0.46%, 2.44+/-0.40%, and 4.50+/ 1.05%, for healthy subjects, nurses, and CLL patients, respectively. A total of 15 microg/ml TT-232 treatment in healthy subjects increased the mean CA frequency (10.38+/-1.57%), as well as the apoptotic cell fraction (2.63+/-0.45 times higher than the corresponding untreated sample). In TT-232-treated PBLs of nurses, CA remained unchanged and the mean apoptotic cell fraction showed only a slight increase (1.24+/-0.11 times higher than the untreated). Among CLL patients, TT 232 treatment significantly increased both CA (up to 17.83+/-4.04%) and the ratio of apoptotic cells (21.78+/-11.00 times higher than the untreated). These results demonstrated significant differences in apoptosis sensitivity in controls, nurses and CLL donors, after 15 microg/ml TT-232 treatment. Data also indicate that the induced CA yields in CLL donors with high CA are in correlation with TT-232 induced apoptosis. PMID- 10708971 TI - The influence of GSTM1 and GSTT1 genotypes on the induction of sister chromatid exchanges and chromosome aberrations by 1,2:3,4-diepoxybutane. AB - Cytogenetic tests - chromosome aberrations (CA), sister chromatid exchanges (SCE) and micronuclei (MN) - are most often applied in biomonitoring of the genotoxicity of potentially carcinogenic chemicals in human cells. One of the extensively studied genotoxins is diepoxybutane (DEB) - reactive biometabolite of butadiene (BD). Several studies showed a high SCE induction in human lymphocytes exposed in vitro to various concentrations of DEB. DEB also proved to be a potent inducer of chromosome aberrations and micronuclei. A bimodal distribution of SCE frequency after in vitro DEB treatment was observed. The aim of the present study was to examine the ability of DEB to induce different individual cytogenetic response measured by SCE and CA frequency. The possible influence of genetic polymorphism has also been taken into account, by including donors representing positive or null GSTM1 and GSTT1 genotypes. Our study supported the earlier results showing that DEB is an effective inducer of SCEs and CAs, causing also the decrease in replication index (RI). DEB bioactivity measured by SCE induction - but not by CA test - was significantly higher in GSTT1 negative than in GSTT1 positive donors. GSTM1 polymorphism had no influence on these endpoints. The donors GSTT1-/GSTM1+ were shown to be slightly more sensitive to DEB than GSTT1 /GSTM1- individuals. There was also observed a unimodal distribution of DEB induced SCEs and CAs in the group, despite the fact that the experiment was performed on the lymphocytes obtained from both GSTT1 positive and negative donors. PMID- 10708972 TI - Genotoxicity testing of five herbicides in the Drosophila wing spot test. AB - Four triazine herbicides: amitrole, metribuzin, prometryn and terbutryn, and the bipyridal compound diquat dibromide have been evaluated for genotoxicity in the wing somatic mutation and recombination test of Drosophila melanogaster, following standard procedures. Third-instar larvae trans-heterozygous for the third chromosome recessive markers multiple wing hairs (mwh) and flare-3 (flr(3)) were chronically fed with different concentrations of the test compounds. Feeding ended with pupation of the surviving larvae. Genetic changes induced in somatic cells of the wing's imaginal discs lead to the formation of mutant clones on the wing blade. Point mutation, chromosome breakage and mitotic recombination produce single spots; while twin spots are produced only by mitotic recombination. Exposure to 0.5 mM and 1 mM of amitrole clearly increased the frequency of small single, large single and total spots. Terbutryn, at the concentration of 5 mM, induced a slight increase in the frequency of small single and total spots, but this result could be false positive. The other three herbicides tested did not show any genotoxic effect. When heterozygous larvae for mwh and the multiple inverted TM3 balancer chromosomes were treated, significant increases in the frequency of mutant spots were only detected for amitrole. The observed spot frequencies were lower than those found in mwh/flr(3)50%) of the total spot induction was due to mitotic recombination. PMID- 10708973 TI - Bioactivation of the carcinogen 11-methoxy-16, 17-dihydro-15H cyclopenta[a]phenanthrene. AB - The title compound is a more potent carcinogen than would be anticipated from its simple phenanthrene structure lacking further D-ring conjugation. In vitro it undergoes microsomal metabolism to yield as major metabolites its 15- and 17 alcohols and its 16, 17-diol; other minor metabolites are also derived from attack at the 5-membered ring, but no evidence of aromatic oxidation is apparent. The title compound is a weak mutagen in the Ames' test with Salmonella typhimurium TA100, but only with microsomal bio-activation. The 17-ol and 16,17 diol are inactive, with or without biological activation. By contrast the 15 alcohol, a rather reactive compound, is a strong mutagen both in the presence and absence of the bio-activation system. This, therefore, may be the proximate carcinogen, and its structural analogy to the naturally occurring hepato carcinogen safrole is noted. PMID- 10708974 TI - Enumeration of micronucleated reticulocytes in rat peripheral blood: a flow cytometric study. AB - Micronuclei (MN) are routinely enumerated in mouse peripheral blood to index genotoxicity. Recent data from the Collaborative Study Group for the Micronucleus Test (CSGMT) [CSGMT (The Collaborative Study Group for the Micronucleus Test), Evaluation of the rat micronucleus test with bone marrow and peripheral blood: summary of the 9th collaborative study by CSGMT/JEMS MMS, Environ. Mol. Mutagen. 32 (1998) 84-100] suggest that rat peripheral blood may also be appropriate for the enumeration of MN, if scoring is limited to the youngest fraction of reticulocytes. The experiments described herein were designed to test whether modifications to a flow cytometric scoring procedure for measuring micronucleated reticulocytes (MN-RET) in mouse peripheral blood could be extended to accurately enumerate MN in rat peripheral blood. Rats were treated with saline or one of three genotoxic agents (6-mercaptopurine, ethyl methanesulfonate or propane sultone) in an acute dosing protocol. Peripheral blood samples were subsequently collected for both microscopic and flow cytometric analysis. Micronucleus frequencies were scored in the youngest fraction of reticulocytes: scoring by microscopy was restricted to the types I and II reticulocytes based on RNA content utilizing acridine orange supravital staining; flow cytometric measurements were restricted to the youngest fraction of reticulocytes based on transferrin receptor (CD71) staining. A statistically significant dose-related increase in the incidence of MN was observed, irrespective of scoring method. A higher level of statistical discrimination between control and genotoxin-treated groups was observed for the flow cytometric data and can most likely be explained by the increased number of cells scored (10x more than microscopy) and the lower scoring variability. Together, these data suggest that (i) rat peripheral blood represents an appropriate compartment for evaluating genotoxin-induced MN when the analysis is restricted to young reticulocytes, and (ii) the measurement of MN in rat peripheral blood reticulocytes benefits from the high throughput methodology of flow cytometry. PMID- 10708975 TI - Role of maternal exposures and newborn genotypes on newborn chromosome aberration frequencies. AB - Maternal exposures may induce chromosome damage and birth defects in the fetus. Polymorphic variation in genes coding for enzymes involved in metabolic activation and detoxification of environmental procarcinogens may account for some of the differences in chromosome aberration frequencies in newborns. In this study, 40 mothers completed questionnaires regarding exposures they received during their pregnancy. Umbilical cord blood samples were analyzed for chromosome aberrations. An average of 1020 metaphase cell equivalents (equal to 1020 G banded cells) were examined from each newborn. In 26 of the newborns, genotyping analysis was performed for genes functioning in metabolic activation and detoxification (cytochrome P450 genes: CYP2D6 and CYP1A1, and phase II genes: NAT1, NAT2, GSTT1, GSTM1, GSTP1, and epoxide hydrolase). A significant association between the CYP1A1 MspI polymorphism and chromosome aberration frequencies was observed in the newborns (p=0.02), with heterozygotes showing higher aberration frequencies than the wild type homozygotes. Some large differences in chromosome aberration frequencies for other genotypes were also noted, but these were not statistically significant. Exposure to tobacco smoke in utero also appeared to increase translocation frequencies. The mean frequency of translocations per 100 cell equivalents from newborns of mothers who smoked during pregnancy was significantly higher than that of newborns whose mothers did not smoke (0.21 vs. 0.11, respectively, p=0.045). PMID- 10708976 TI - Flow cytometric detection of micronuclei and cell cycle alterations in fish derived cells after exposure to three model genotoxic agents: mitomycin C, vincristine sulfate and benzo(a)pyrene. AB - The measurement of cytogenetic alterations in vitro is considered an initial step in the risk assessment procedures for genotoxic agents. The concern about genotoxic pollutants in natural fish population makes the use of fish-derived cells an useful tool for these purposes. The technological improvements in well established cytogenetic endpoints, such as micronuclei (MN) estimations by means of flow cytometry, have been proposed in the later years using mammalian cells. In this work, we test the capability of flow cytometry to evaluate MN induction and cell cycle alterations in an established fish cell line (RTG-2) using three agent-inductor models at different concentrations and exposure periods. For mitomycin C, an inverse relationship between length of exposure period and concentrations was observed. A dose-response relationship was observed after exposing RTG-2 cells to vincristine sulfate and benzo(a)pyrene. As this study shows, RTG-2 cells respond to clastogenic and aneugenic effects of the tested chemicals through the induction of MN at similar doses to mammalian cells and without the addition of exogenous metabolic activity. The possibility to check cell cycle alterations, in the same sample, gives the opportunity to evaluate early signals of cytotoxicity. The use of flow cytometry improves the assay by means of its speed and objectivity, which makes the assay very useful for genotoxicity assessment of aquatic chemicals. PMID- 10708977 TI - Induction of chromosome aberrations, micronucleus formation and sperm abnormalities in mouse following carbofuran exposure. AB - Carbofuran was tested to study in vivo cytogenetic effects in mouse bone marrow cells and morphological alterations in sperms. The acute oral and intraperitoneal (i.p.) LD(50) of carbofuran was determined to be 9.5 or 2.0 mg/kg b.w. in mice, respectively. The animals were orally administered 1.9, 3.8 or 5.7 mg/kg b.w. (20, 40 and 60% of LD(50)) of carbofuran for 24 h or 1.9 mg/kg b.w. for 4 consecutive days (cumulative 7.6 mg/kg or 80% of LD(50)) to analyse chromosome aberrations (CAs). For micronucleus test (MT) animals were orally exposed to 5.7 mg/kg b.w. for 24 and 48 h or 1.9 mg/kg b.w. for 4 consecutive days. For reference mice were exposed to peanut oil (negative control) and cyclophosphamide (20 mg/kg) or ethyl methanesulfonate (EMS: 100 mg/kg) positive control for CAs and MT respectively. To analyse the effect on sperm morphology mice were exposed to single i.p. dose of 1 and 2 mg/kg b.w. of carbofuran and repeatedly to 0.5 mg/kg for 5 consecutive days. Cytogenetic analysis revealed that all the test doses induced mitotic inhibition, CAs, micronucleus (MN) formation and sperm abnormalities in a dose dependent manner. Present observations concurrent with earlier reports substantiate the genotoxic potential of carbofuran and possible risk to human beings. PMID- 10708978 TI - Effects of the antioxidants curcumin and vitamin C on cisplatin-induced clastogenesis in Wistar rat bone marrow cells. AB - The use of dietary antioxidants to prevent antitumor agent-induced chromosomal damage in nontumor cells is currently eliciting considerable interest. Curcumin (CMN) is a dietary antioxidant that has been reported to protect against clastogenesis in in vivo and in vitro assays. This study was undertaken to investigate the modulatory effects of CMN on cisplatin-induced chromosomal aberrations in Wistar rat bone marrow cells and whether there is any potentiation of these effects with the combination between CMN and vitamin C (VC), which has been reported to reduce the clastogenic effect of many antitumor agents in in vivo assays. Animals treated with CMN plus a single dose of cisplatin, at 18, 24 or 72 h following treatment, presented a statistically significant reduction in the total amount of chromosomal damage and in the number of abnormal metaphases. The results also indicate that the combination between antioxidants would not be effective in protecting against cisplatin-induced chromosomal damage in animals sacrificed 24 h after cisplatin treatment. Under the present experimental conditions, CMN could prevent cisplatin-induced clastogenesis by acting as a free radical scavenger. PMID- 10708979 TI - Assessment of radiation-induced DNA damage caused by the incorporation of 99mTc radiopharmaceuticals in murine lymphocytes using single cell gel electrophoresis. AB - The DNA damage induced by the 99mTc-radiopharmaceuticals incorporation to the cell was determined by the single-cell gel electrophoresis in murine lymphocytes in vitro. The 99mTc-hexamethyl-propylene amine oxime (99mTc-HMPAO) and 99mTc-2, 5 dihydroxybenzoic acid (99mTc-gentisic acid) induced nearly 100% of cells with breaks and/or alkali labile sites, which is explained by the action of the Auger electrons produced by the decay of the 99mTc. These results agree with the doses of 1.6 and 1.0 Gy estimated by subcellular dosimetry for 99mTc-HMPAO that is incorporated in the cytoplasm, and the 99mTc-gentisic acid, which remains bonded to the cell membrane, respectively. The results imply that Auger electrons are able to cause important DNA damage, when the radionuclide is incorporated in the range of a few microns from the nuclei. PMID- 10708980 TI - Detection of micronuclei in haemocytes of zebra mussel and great ramshorn snail exposed to pentachlorophenol. AB - The frequency of micronuclei (MN) induced by pentachlorophenol (PCP) in haemocytes of zebra mussel, Dreissena polymorpha Pall. and great ramshorn snail, Planorbarius corneus L. was determined over a 14 days of exposure (sampling after 4, 7 and 14 days) under laboratory conditions. PCP doses for zebra mussel ranged from 10 to 150 microg/l, and for ramshorn snail from 10 to 450 microg/l. Micronuclei were detected after bisbenzimide fluorescent staining. Positive responses were observed in both species. The mean MN frequencies in treated mussels ranged between 0.69 and 7.50 per thousand, and between 2.07 and 13.80 per thousand in treated snails. The spontaneous MN levels in mussels averaged from 0.5 to 2.75 per thousand, and in snails from 1.56 to 2.00 per thousand. Our results suggest that haemolymph of both species represent an appropriate test tissue in environmental genotoxicity assessment. PMID- 10708981 TI - Inhibitory activity of vitamin E and alpha-naphthoflavone on beta-carotene enhanced transformation of BALB/c 3T3 cells by benzo(a)pyrene and cigarette-smoke condensate. AB - We previously found that beta-carotene (betaCT) can act as a co-carcinogenic agent enhancing the cell transforming activity of powerful carcinogens such as benzo(a)pyrene (B(a)P) and cigarette-smoke condensate (TAR) in an in vitro medium term ( approximately 8 weeks) experimental model utilizing BALB/c 3T3 cells (Mutat. Res. 440 (1999) 83-90). Here, we investigated whether vitamin E (VitE) and alpha-naphthoflavone (alphaNF) are able to affect the co-carcinogenic activity of betaCT in terms of inhibiting B(a)P and TAR cell transforming potential. The following experimental schedules were performed: (i) cultures treated for 72 h with chemicals in various experimental combinations (acute treatment); (ii) cultures grown in presence of tester agents for the whole period of the assay (chronic treatment) to more closely mimic human exposure. While the co-carcinogenic potential of betaCT was confirmed on both B(a)P and TAR, the latter being ineffective by itself, we found in repeated experiments that the presence of VitE or alphaNF significantly reduced the betaCT's enhancing effect in the formation of transformation foci by B(a)P and TAR. The mechanism of the inhibition could be explained by the known ability of alphaNF to inhibit cytochrome P450-linked B(a)P-bioactivating monooxygenases, while VitE may contrast the prooxidant activity of betaCT (e.g., oxygen radicals overgeneration). While highlighting the importance of increasing knowledge of the role of single provitamins, vitamins and micronutrients, our findings also underline the potential advantages of combining several dietary supplements in in vitro preventive investigations. PMID- 10708982 TI - Sister chromatid exchange rate from pleural fluid cells in patients with malignant mesothelioma. AB - This study assessed the impact of malignant mesothelioma on the frequencies of sister chromatid exchange (SCE) in the pleural effusion cells. Ten patients with mesothelioma and 20 control subjects were included in the study. The control subjects were the patients with tuberculosis pleurisy, and the remaining 10 subjects of control group were healthy volunteers and only heparinized blood samples were collected from these subjects. The pleural effusion cells were cultured with conventional culture methods. The samples were obtained from the patients after histopathologic confirmation of the malignancy but before the initiation of chemotherapy or radiotherapy. At the end of the culture period and 48 h prior the harvesting, BrdU was added into flasks. Totally, 100 metaphases were scored for each sample. In this study, we found that the SCE frequencies of malignant pleural mesotheliomas were significantly higher than the control subjects (P<0.001). Six of 10 patients came from central Anatolia, which is of great importance due to high rate of exposure to asbestosis in this region. PMID- 10708983 TI - Comparison of the mutagenic specificity induced by four nitro-group-containing aromatic amines in Salmonella typhimurium his genes. AB - Four nitrated aromatic amines (2-nitro-p-phenylenediamine [2NPD], 3-nitro-o phenylenediamine [3NPD], 4-nitro-o-phenylenediamine [4NPD] and 4,4'-dinitro-2 biphenylamine [DNBA]) are direct-acting mutagens in Salmonella typhimurium strain TA100. These compounds were tested further using the Xenometrix strains of S. typhimurium: TA7001, TA7002, TA7003, TA7004, TA7005, and TA7006, with and without S9 mix in the plate incorporation assay. The direct-acting mutagenicity of 2NPD, 4NPD, and DNBA was detected with TA7002, TA7004 and TA7005. 2NPD and DNBA showed some activity in TA7006; DNBA also showed some activity in TA7003. Mutagenicity was generally decreased in these strains when S9 was added. 3NPD was mutagenic in TA7004 without S9 and in TA7005 with and without S9. These data suggest that 2NPD, 4NPD and DNBA induced TA-->AT and CG-->AT transversions as well as GC-->AT transitions in the his gene. 3NPD induced CG-->AT transversions and GC-->AT transitions. 2NPD and DNBA also induced a small portion of CG-->GC transversions. PMID- 10708984 TI - Anti-mutagenic structural modification by fluorine-substitution in highly mutagenic 4-methylquinoline derivatives. AB - We have previously shown that fluorine-substitution at position 3 of quinoline deprived this molecule of mutagenicity, possibly due to interference with the yield of its metabolically activated form, the 1,4-hydrated 2,3-epoxide (enamine epoxide), which is directly responsible for the mutagenic modification of DNA. To further explore the possibility of a method for anti-mutagenic modification of mutagens by fluorine-substitution, 4-methylquinoline (4-MeQ), the most mutagenic form of all the quinoline derivatives examined so far, was used as a target in the present study. Five mono- and di-fluorinated derivatives of 4-MeQ, 2-fluoro-4 methylquinoline (2-F-4-MeQ), 6-F-4-MeQ, 7-F-4-MeQ, 2,6-difluoro-4-methylquinoline (2, 6-diF-4-MeQ), and 2,7-diF-4-MeQ, were subjected to analysis of their structure-mutagenicity relationships. The 2-fluorinated derivatives (2-F-4-MeQ, 2,6-diF-4-MeQ, and 2,7-diF-4-MeQ) were all non-mutagenic in the Ames test. 7-F-4 MeQ was as highly mutagenic as, and 6-F-4-MeQ was less mutagenic than non fluorinated 4-MeQ. Metabolic studies were also conducted with 4-MeQ, 2-F-4-MeQ, 6 F-4-MeQ, and 7-F-4-MeQ, using a liver microsomal enzyme fraction prepared from the 3-methylcholanthrene-treated rat. The HPLC analytical data showed that, although the metabolic patterns (hydroxylation at 4-methyl group as a main metabolic pathway and 3-hydroxylation as a minor pathway) of these four F-MeQs were similar to one another, only the 3-hydroxy metabolite of 2-F-4-MeQ was not produced under the present experimental conditions employed. These results suggest that fluorine-substitution at position 2 of 4-MeQ inhibited the formation of the enamine epoxide in the pyridine moiety and deprived this molecule of mutagenicity as in the case of quinoline. PMID- 10708985 TI - Effect of food reductones on the generation of the pyrazine cation radical and on the formation of the mutagens in the reaction of glucose, glycine and creatinine. AB - One of the possible pathways of the formation of mutagens in heated foods is through the pyrazine cation radical generated in the early stage of the Maillard reaction. The aim of the present study was to elucidate how food reductones contribute to the pyrazine cation radical generation in the reaction of glucose (Glc) and glycine (Gly), and to the formation of the mutagens in the reaction of Glc, Gly and creatinine. Electron spin resonance (ESR) studies showed that fragrant reductones, 2,5-dimethyl-4-hydroxy-3(2H)-furanone (DMHF) and 4-hydroxy 2(or 5)-ethyl-5(or 2)-methyl-3(2H)-furanone (HEMF), generated in the Maillard reactions, enhanced the generation of the pyrazine cation radical in the reaction of Glc and Gly, and the reaction of DMHF or HEMF with Gly generated a larger amount of the pyrazine cation radical than the reaction of Glc and Gly, indicating that the furanones were intermediates of the pyrazine cation radical. By contrast, food antioxidants, ascorbic acid and erythorbic acid, effectively scavenged the pyrazine cation radical generated in the reaction of Glc and Gly. DMHF and HEMF were not effective to modulate the mutagen formation in the reaction of Glc, Gly and creatinine, and the mutagenicity produced in the reaction of DMHF or HEMF, Gly and creatinine was lower than that produced in the reaction of Glc, Gly and creatinine. On the other hand, ascorbic acid and erythorbic acid were effective to decrease the mutagen formation in the reaction of Glc, Gly and creatinine. PMID- 10708986 TI - Comparison of environmental tobacco smoke concentrations and mutagenicity for several indoor environments. AB - Environmental tobacco smoke (ETS) is a major source for indoor air pollution. Although ETS-caused indoor air pollution has been well studied in the developed countries, few studies have examined ETS indoor air pollution in China, which currently has the largest population of tobacco smokers. In this study, respirable-particulate (RP) from ETS-contaminated (RP-ETS) indoor air was collected and measured in 5 different indoor environments during the winter in the northwestern Liaoning Province, China. The extractable portion of RP-ETS (ERP ETS) was obtained by dichloromethane extraction and used in the Salmonella mutagenicity assay in the presence of S9 using strains TA98, TA100, and TA1538. The percentage of RP-ETS attributable to ETS (ETS-RP) and the percentage of ERP ETS attributable to ETS (ETS-ERP) were estimated by measuring the concentration of solanesol, an ETS marker. Comparative results in 5 different indoor environments were: (1) the concentration of RP-ETS ranged from 197.3 to 1227.6 microg/m(3) and approximately 64.7 to 92. 0% of the RP-ETS originated from ETS; (2) the concentration of ERP-ETS ranged 88.8 to 601.5 microg/m(3) and approximately 83.1 to 95.4% of the ERP-ETS originated from ETS; (3) the mutagenic potency (revertants/m(3)) of ERP-ETS ranged from 60.4 to 595.5 for TA98, from 33.7 to 312.8 for TA100, and from 49.7 to 475.2 for TA1538. The data indicate that the extent of ETS pollution and the potential health hazards of ETS to humans in the five indoor environments are in the following increasing order: rural bedrooms, urban living rooms, office rooms, restaurants, and passenger cars in that area. PMID- 10708987 TI - Modeling the mouse lymphoma forward mutational assay: the Gene-Tox program database. AB - An SAR model of the induction of mutations at the tk(+/-) locus of L5178Y mouse lymphoma cells (MLA, for mouse lymphoma assay) was derived based upon a re evaluation of experimental results reported by a Gene-Tox (GT) working group [A.D. Mitchell, A.E. Auletta, D. Clive, P.E. Kirby, M.M. Moore, B.C. Myhr, The L5178Y/tk(+/-) mouse lymphoma specific gene and chromosomal mutation assay. A phase III report of the U.S. Environmental Protection Agency Gene-Tox Program, Mutation Res. 394 (1997) 177-303.]. The predictive performance of the GT MLA SAR model was similar to that of a Salmonella mutagenicity model containing the same number of chemicals. However, the structural determinants (biophores) derived from the GT MLA SAR model include both electrophilic as well as non-electrophilic moieties, suggesting that the induction of mutations in the MLA may occur by both direct interaction with DNA and by non-DNA-related mechanisms. This was confirmed by the observation that the set of biophores associated with MLA overlapped significantly with those associated with phenomena related to loss of heterozygosity, chromosomal rearrangements and aneuploidy. The MLA SAR model derived from the GT data evaluation was significantly more predictive than an SAR model previously derived from MLA data reported by the US National Toxicology Program [B. Henry, S.G. Grant, G. Klopman, H.S. Rosenkranz, Induction of forward mutations at the thymidine kinase locus of mouse lymphoma cells: evidence for electrophilic and non-electrophilic mechanisms, Mutation Res. 397 (1998) 331 335.]. Moreover, the latter model appeared to be more complex than the former, suggesting that the GT induction data was both simpler mechanistically and more homogeneous than that of the NTP. PMID- 10708989 TI - Degenerative Spondylolisthesis: Diagnosis and Treatment. AB - Degenerative spondylolisthesis is most often seen at the L4-5 level. The most common complaint is back pain, but the advent of leg symptoms, such as claudication and restless legs syndrome, is often the reason for seeking specialized medical attention. Conservative treatment usually suffices; extended bed rest is of little value. The 15% of patients who are surgical candidates are those with clinical signs and symptoms of cauda equina abnormality, progressive muscular weakness, or progressive incapacitating radicular pain or claudication. The author advocates pedicle-to-pedicle decompression with preservation of the articular facets as the essential operation. The indications for fusion have been debated, but recent prospective studies show improved outcomes after fusion. The risk of significant morbidity associated with laminectomy and fusion increases as a function of age and magnitude of operation; therefore, careful patient selection for surgical intervention is vital. PMID- 10708988 TI - Lateral and Medial Epicondylitis of the Elbow. AB - Epicondylitis of the elbow involves pathologic alteration in the musculotendinous origins at the lateral or medial epicondyle. Although commonly referred to as "tennis elbow" when it occurs laterally and "golfer's elbow" when it occurs medially, the condition may in fact be caused by a variety of sports and occupational activities. The accurate diagnosis of these entities requires a thorough understanding of the anatomic, epidemiologic, and pathophysiologic factors. Nonoperative treatment should be tried first in all patients, beginning with an initial phase of rest, ice, nonsteroidal anti-inflammatory agents, and possibly corticosteroid injection. A second phase includes coordinated rehabilitation, consisting of range-of-motion and strengthening exercises and counterforce bracing, as well as technique enhancement and equipment modification if a sport or occupation is causative. Nonoperative treatment has been deemed highly successful, yet the few prospective reports available suggest that symptoms frequently persist or recur. Operative treatment is indicated for debilitating pain that is diagnosed after the exclusion of other pathologic causes for pain and that persists in spite of a well-managed nonoperative regimen spanning a minimum of 6 months. The surgical technique involves excision of the pathologic portion of the tendon, repair of the resulting defect, and reattachment of the origin to the lateral or medial epicondyle. Surgical treatment results in a high degree of subjective relief, although objective strength deficits may persist. PMID- 10708990 TI - Modularity of Prosthetic Implants. AB - The vast majority of total-joint-replacement components currently utilized are modular to some degree. Modularity reduces inventory and increases the surgeon's options in both primary and revision total-joint arthroplasty. Use of a modular interface, however, increases the risk of fretting, wear debris, and dissociation and mismatching of components. The use of modular heads in total hip replacement is firmly established. The occurrence of corrosion and fretting has been recognized, and most manufacturers have improved the quality of the interface to minimize these problems. Modular polyethylene liners also offer advantages, particularly in revision procedures, where the option of additional screw fixation remains important. Many uncemented acetabular components are inserted without screws, which may generate renewed interest in one-piece factory preassembled components. The conformity, locking mechanism, and nonarticular interface of modular acetabular components have all been studied and improved. Modular tibial components offer additional flexibility in the performance of total knee replacement but introduce the risk of dissociation and increased polyethylene wear; in revision procedures, modularity provides a valuable option for dealing with bone loss and an additional method of fixation by means of press fit stems. Modular humeral components offer a significant advantage with limited apparent risk; however, longer clinical experience is required to assess potential problems. PMID- 10708991 TI - Anterior Cruciate Ligament Insufficiency: Principles of Treatment. AB - Anterior cruciate ligament (ACL) injuries often result in functional disability, particularly in jumping, cutting, and deceleration activities. Some patients can accommodate to this functional loss, while others require surgical reconstruction of the ligament to provide stability and to protect the meniscus from further injury. Nonoperative management involves an intensive rehabilitation program, patient counseling about high-risk activities, and measures to prevent recurrent injuries. Surgical reconstruction of the ACL involves the technical factors of graft selection, positioning, fixation, and tensioning and the avoidance of stress risers. A supervised and intensive rehabilitation program is necessary to achieve optimal results. PMID- 10708992 TI - Acute Calcaneal Fractures: Treatment Options and Results. AB - The treatment of choice for acute displaced intra-articular calcaneal fractures remains controversial. The authors present a brief historical review of treatment options and results, coupled with the biomechanical rationale for open reduction and internal fixation. Their current management protocol and surgical technique are outlined, along with preliminary functional results at an average follow-up of 2.5 years. PMID- 10708993 TI - Infected Total Knee Replacements. AB - Deep infection is a devastating complication following total knee arthroplasty. Prompt diagnosis and definitive treatment of this complication are essential for a successful outcome. The treatment options for an infected total knee replacement include (1) antibiotic suppression alone; (2) aggressive wound debridement, drainage, and antibiotic suppression therapy; (3) resection arthroplasty; (4) arthrodesis; (5) two-stage reimplantation; and (6) amputation. Successful salvage of this complication can be accomplished only by extensive investment of surgical and infectious disease efforts in eradicating the infection. Two-stage reimplantation has been the most successful functional option and should be used whenever possible to definitively eradicate the infection and ensure good function of the knee joint. PMID- 10708994 TI - Displaced Proximal Humeral Fractures: Evaluation and Treatment. AB - Successful treatment of proximal humeral fractures relies on the surgeon's ability to make an accurate diagnosis. Treatment must be predicated on a thorough understanding of the complex shoulder anatomy, a precise radiographic evaluation, and use of a well-designed classification system. Appropriate and realistic goals must be established for each patient. The patient's general medical health, physiologic age, and ability to cooperate with intense and prolonged rehabilitation are all considerations when selecting the optimal treatment. PMID- 10708995 TI - Reflex Sympathetic Dystrophy of the Knee. AB - Reflex sympathetic dystrophy (RSD) of the knee frequently does not present with the classic combination of signs and symptoms seen in the upper extremity. Pain out of proportion to the initial injury is the hallmark symptom. Symptom relief by sympathetic block is the current standard for confirmation of the diagnosis. Because invasive diagnostic procedures, such as arthroscopy, are likely to increase symptoms, evaluation with a noninvasive diagnostic modality, such as magnetic resonance imaging, is preferred. Generally, RSD should be treated before surgical intervention to correct any underlying intra-articular pathologic condition. However, surgery may sometimes be necessary before RSD symptoms resolve; in these cases, use of intra- and postoperative continuous epidural block can be successful. The initial treatment of RSD of short duration should be conservative; physical therapy modalities, including exercise and contrast baths, and non-steroidal anti-inflammatory drugs are indicated. In the authors' experience, an indwelling epidural block using bupivacaine for several days followed by use of a narcotic agent, combined with functional rehabilitation, is the most effective management when noninvasive treatment has failed. Surgical sympathectomy can be successful, but should be reserved until repeated lumbar sympathetic block or more than one trial of inpatient epidural block has failed. Early diagnosis and early institution of treatment (prior to 6 months) are the most favorable prognostic indicators in the management of RSD. PMID- 10708996 TI - Full-Thickness Rotator Cuff Tears: Factors Affecting Surgical Outcome. AB - Eighty-five percent to 95% of patients who undergo primary surgical repair of full-thickness rotator cuff tears have a significant decrease in shoulder pain and improvement in shoulder function. The results of surgery are dependent on the surgical technique, the extent of pathologic changes in the rotator cuff, and the postoperative rehabilitation protocol. Preoperative factors associated with a less favorable result are the size of the tear, the quality of the tissues, the presence of a chronic rupture of the long head of the biceps tendon, and the degree of preoperative shoulder weakness. Surgical factors associated with a less favorable result include inadequate acromioplasty, residual symptomatic acromioclavicular arthritis, inadequate rotator cuff tissue mobilization, deltoid detachment or denervation, and failure of rotator cuff healing. Clinical evaluation and preoperative imaging of the shoulder will improve patient selection and counseling. Meticulous surgical technique and postoperative rehabilitation will optimize the final result. PMID- 10708997 TI - Acute Slipped Capital Femoral Epiphysis: Treatment Alternatives. AB - Acute slipped capital femoral epiphysis represents a unique type of proximal femoral epiphyseal instability. The potential for complications and unsatisfactory outcomes is high, especially due to avascular necrosis. A newly proposed classification based on epiphyseal stability provides a rational assessment of this acute physeal fracture. Improvements in imaging and fixation techniques have reduced the morbidity of this condition. Choice of treatment must be based on the surgeon's experience and expertise. Vigilance is particularly required in young patients with underlying endocrine or metabolic conditions that predispose them to bilateral hip involvement. PMID- 10708998 TI - Nonreamed Intramedullary Nailing of Open Tibial Fractures. AB - The development of small-diameter interlocking intramedullary nails that can be inserted without reaming provides a fixation option for open tibial-shaft fractures. Nonreamed intramedullary nailing of these injuries facilitates soft tissue management without an increase in infection or nonunion rates relative to external fixation. Reaming is not required, which means less injury to the tibial endosteal blood supply. Proximal and distal interlocking maintains better bone alignment than is possible with semirigid or noninterlocking intramedullary nails. The technique of using these devices with static interlocking is described, as are some suggested techniques for avoiding complications. PMID- 10708999 TI - Nerve Entrapment Syndromes in the Wrist. AB - The patient with compression neuropathies of the median and ulnar nerves at the wrist commonly presents with pain, paresthesias, and weakness in the hand and digits. Diagnosis of these conditions is becoming more widespread with the increased attention given to "cumulative trauma disorders" during the past decade. Successful management requires a thorough understanding of the pathophysiology of compression neuropathy and how it relates to the various diagnostic tests available today. The authors review the epidemiology, etiology, and evaluation of compression neuropathy and discuss common clinical presentations, treatment recommendations, and controversies surrounding carpal and ulnar tunnel syndromes. PMID- 10709000 TI - Patellofemoral Pain Disorders: Evaluation and Management. AB - Patellofemoral pain disorders can be difficult to diagnose. Careful attention to the history and physical examination is central to accurate diagnosis. Standardized office radiographs are sufficient in most cases. Computed tomography of the patellofemoral joint (precise midpatellar transverse images through the posterior femoral condyles with the knee at 15, 30, and 45 degrees of knee flexion) will provide valuable objective information regarding subtle abnormalities of patellar alignment. Magnetic resonance imaging and radionuclide scanning may be helpful in selected cases. By differentiating between rotational (tilt) and translational (subluxation) components of patellar malalignment, the clinician will be better able to prescribe appropriate treatment. It is also extremely important to localize and quantitate articular and retinacular abnormalities. While nonoperative treatment is usually successful, surgery is sometimes required. Lateral release will relieve tilt and associated pain in the lateral retinaculum. Realignment of the extensor mechanism, usually at the level of the tibial tubercle, is necessary to control lateral tracking (subluxation) of the patella. If there is lateral or distal medial articular damage related to chronic lateral tilt and/or subluxation, shift of the tibial tubercle will help to unload damaged cartilage while realigning the extensor mechanism. PMID- 10709001 TI - Corticosteroid Injections: Their Use and Abuse. AB - Local injections of corticosteroids are commonly used in orthopaedic practice on the assumption that they will diminish the pain of inflammation and accelerate healing. Less often considered is the possibility that their use may delay the normal repair response. Among the multitude of conditions treated with corticosteroids are acute athletic injuries, overuse syndromes, nerve compression, bone cysts, and osteoarthritis. Unfortunately, there is a paucity of well-controlled studies that provide definitive recommendations for nonrheumatologic use of corticosteroids. Also troubling are the significant potential complications that can occur with their use. The authors believe that use of corticosteroids should be limited to the few conditions that have been proved to be positively influenced by them. Their use must be accompanied by a well-orchestrated treatment plan including close follow-up, physical therapy, and limitation of activities. PMID- 10709002 TI - Hip Fractures: I. Overview and Evaluation and Treatment of Femoral-Neck Fractures. AB - Hip fractures remain a major source of morbidity and mortality in the elderly, and their incidence is increasing as the population ages. Surgical management followed by early mobilization is the treatment of choice for most patients with hip fractures. However, all comorbid medical conditions, particularly cardiopulmonary and fluid- electrolyte imbalances, must be evaluated and stabilized prior to operative intervention. Nondisplaced femoral-neck fractures should be stabilized with multiple parallel lag screws or pins. The treatment of displaced femoral-neck fractures is based on the patient's age and activity level: young active patients should undergo open reduction and internal fixation; older, less active patients are usually treated with hemiarthroplasty, either uncemented or cemented. Regardless of treatment method, the goal is to return the patient to his or her prefracture level of function. PMID- 10709003 TI - Hip Fractures: II. Evaluation and Treatment of Intertrochanteric Fractures. AB - Surgical stabilization followed by early mobilization is the treatment of choice for both nondisplaced and displaced intertrochanteric fractures. Fracture stability is dependent on the status of the posteromedial cortex. The sliding hip screw is the device mostly commonly used for fracture stabilization. The most important aspect of its insertion is secure placement within the femoral head. Although the sliding hip screw allows postoperative fracture impaction, it is essential to obtain an impacted reduction at the time of surgery. If there is a large posteromedial fragment, an attempt should be made to internally fix the fragment with a lag screw or cerclage wire. Although intramedullary hip screws have not been shown to be superior to the sliding hip screw, they may have selected indications. PMID- 10709004 TI - The Multidisciplinary Approach to Occupational Low Back Pain and Disability. AB - Chronic disability generates most of the growing costs of occupational low back injuries. When back problems persist for more than a few months, traditional diagnostic and therapeutic approaches are rarely curative. Beyond the challenges of physical impairment, disabling back pain is commonly complicated by psychosocial issues, including depression, fear of reinjury, family discord, and vocational dissatisfaction. The biopsychosocial complexity of chronic disability often demands integrated care from physicians, physical and occupational therapists, psychologists, and vocational counselors. In the past decade, the care of back-injured workers has shifted emphasis from symptom palliation toward functional restoration. This evolution has been possible, in part, through improved quantification of physical capacities. Repeated objective measurements of function guide rehabilitation and recommendations for return to work and other activities. Published results of function-oriented multidisciplinary care depend on the outcome variables reported and the particular socioeconomic setting. PMID- 10709005 TI - Periprosthetic Femoral Fractures. AB - Fracture of the femoral shaft around a hip prosthesis presents the simultaneous problems of prosthetic stability and femoral- fracture management. Treatment options include nonoperative stabilization (traction) and operative stabilization by means of intramedullary fixation, extramedullary fixation, or proximal femoral prosthetic replacement. PMID- 10709006 TI - Spontaneous Osteonecrosis of the Knee. AB - Spontaneous osteonecrosis of the knee is a common cause of knee pain, principally seen in women over 60 years of age. This condition is distinguished from secondary conditions with known causes, such as corticosteroid-induced osteonecrosis. Although originally described and most common in the medial femoral condyle, it can also occur in the tibial plateaus and on the lateral side of the femur. The radionuclide bone scan will show focally increased uptake before the radiographs are abnormal. Magnetic resonance imaging can also be diagnostic, but the findings may be normal early in the course of the disease. The etiology remains unknown, but it is speculated that primary vascular ischemia or microfractures in osteoporotic bone are causative. Many patients have a benign course followed by resolution of symptoms. Therefore, conservative management is indicated initially. If progressive collapse accompanied by severe symptoms occurs, high tibial osteotomy, unicompartmental replacement, and total knee replacement are therapeutic alternatives. Recognition of this entity is important to avoid needless surgical intervention. PMID- 10709007 TI - Endoscopic Carpal Tunnel Release. AB - On the basis of clinical outcome measures, endoscopic carpal tunnel release is an effective operation for treating idiopathic carpal tunnel syndrome. Patients who have undergone bilateral carpal tunnel operations have routinely preferred endoscopic release over the open release. An endoscopic release allows many patients to return to work sooner. However, the benefits of more rapid functional recovery and return to work are tempered by the increased cost and higher complication rate of the procedure. Endoscopic carpal tunnel release is a technically demanding procedure with low tolerances for error. Despite its widespread use, its role is not yet clearly defined. PMID- 10709008 TI - Scaphoid Nonunion. AB - The natural history and treatment of scaphoid fractures and subsequent nonunions have occupied a substantial portion of the orthopaedic literature. The authors examine the role of modern diagnostic tools in making an earlier diagnosis of scaphoid nonunion, in more accurately determining the displacement and angulation of the fragments, and in identifying the presence of avascular necrosis. They also consider the various available treatment modalities, including immobilization, electrical stimulation, both conventional and vascularized bone grafting, and internal fixation. Finally, a brief review of salvage procedures and the authors' preferred treatment are presented. PMID- 10709009 TI - Restoration of Injured or Degenerated Articular Cartilage. AB - Intra-articular fractures, ligamentous and meniscal injuries, and articular cartilage breakdown are major causes of degenerative joint disease. Lesions on the articular surface seem to have a limited capacity for repair and often progress inexorably toward osteoarthritis. Recent studies on joint immobilization and cartilage atrophy, however, have shown that repair and remodeling of articular cartilage may be possible. Currently used clinical methods of stimulating cartilage repair and remodeling include alteration of the loading on degenerated joints (primarily by using osteotomies), introduction of new cartilage-forming cells by perforation of subchondral bone, and soft-tissue arthroplasty. These procedures provide temporary relief in selected patients, but they often do not predictably restore long-term joint function. Experimentally, cartilage repair has been stimulated successfully with the use of allografts of periosteum and perichondrium, which serve as sources of cells with chondrogenic potential; introduction of cells grown in culture (stem cells or chondrocytes); stimulation by fibrin clot formation; artificial collagen matrices combined with cell transplants; and chondrogenic growth factors. The long-term success of all these methods has not been explored thoroughly, even in animal studies. Nevertheless, some research results are sufficiently encouraging to suggest that repair of the degenerating articular cartilage may be possible in the future. PMID- 10709010 TI - Soft-Tissue Tumors: Diagnosis, Evaluation, and Management. AB - Benign soft-tissue neoplasms and tumorlike conditions of the musculoskeletal system are common. Sarcomas are less frequent, with only 5,000 new cases diagnosed each year in the United States. After plain radiographs of the affected area have been obtained, magnetic resonance (MR) imaging (both T1- and T2 weighted sequences) is the best imaging modality for detecting and characterizing the lesion. Although MR imaging is not specific in determining whether lesions are benign or malignant, it can be useful in evaluating other characteristics, such as size, pattern of growth, integrity of natural boundaries, and homogeneity. Biopsy must be done carefully, so as not to adversely affect the outcome. Technical considerations include proper location and orientation of the biopsy incision, meticulous hemostasis, and frozen-section analysis to ensure that diagnostic material has been obtained. Effective treatment requires close coordination between the surgeon, the radiation oncologist, the pathologist, the plastic surgeon, and the diagnostic radiologist. Limb-salvage surgery has resulted in a local control rate greater than 90%. High-grade tumors that are larger than 5 cm in diameter have the worst prognosis. The role of chemotherapy remains controversial and unresolved. PMID- 10709011 TI - Wear Debris in Total Joint Replacements. AB - In vivo degradation of prosthetic implant materials is increasingly recognized as a major factor limiting the durability of total joint arthroplasty. In vivo degradation occurs primarily by means of wear processes that can generate large quantities of particulate debris. This debris can stimulate an adverse local host response leading to periprosthetic bone loss, which can compromise implant fixation and bone stock. The authors review the basic mechanisms of implant degradation and the host response to particulate degradation products, particularly in the context of the pathogenesis of osteolysis. Submicron polyethylene particles (mean size, 0.5 um) are the dominant type of wear particle present in periprosthetic tissues associated with uncemented hip replacements. Polyethylene wear can be minimized by improving the quality of the polyethylene, avoiding use of large-diameter (greater than 28 mm) femoral heads in total hip arthroplasty, and improving the design and fabrication of modular connections, which can be important sources of three-body wear particles. Advances in the understanding of the basic mechanisms of osteolysis are critical to the development of preventive measures that will minimize the clinical impact of this phenomenon. PMID- 10709012 TI - Isokinetic Muscle Testing: Is It Clinically Useful? AB - The use of computer-driven muscle-testing devices has become increasingly popular during the past two decades. This expensive equipment allows evaluation of muscles and muscle groups in an isokinetic manner. Isokinetic muscle testing is performed with a constant speed of angular motion but variable resistance. Isokinetic dynamometers have been shown to produce relatively reliable data when testing simple, uniaxial joints, such as the knee, as well as when testing the spine in flexion and extension. Isokinetic strength data are generally not helpful in the diagnosis of orthopaedic abnormalities. Isokinetic testing can be helpful during the rehabilitation of orthopaedic patients, since it allows easy monitoring of progress. It also enables the patient to work on muscle rehabilitation in a controlled manner at higher speeds than are possible with more conventional exercise equipment. An isokinetic rehabilitation program can be easily tailored with concentric and eccentric components that closely resemble muscle actions during occupational and sports activities. PMID- 10709013 TI - Management of Traumatic Foot Wounds. AB - The foot is frequently exposed to direct trauma due to its role in weight bearing. The soft-tissue wounds that commonly result interfere with ambulation due to complications such as tissue necrosis, scar formation, infection, and deformity. The five most common categories of foot injury are (1) low-velocity blunt trauma, (2) high-velocity blunt trauma, (3) low-velocity penetrating trauma, (4) high-velocity penetrating trauma, and (5) thermal injuries. For major wounds, the treatment is early aggressive debridement, copious irrigation, and skeletal stabi-lization with early coverage of skin defects. Local and systemic antibiotics are adjunctive to debridement to prevent infection. Prompt recognition and release of compartment syndrome of the foot are extremely important. Close observation is appropriate for wounds that appear minor on initial evaluation. PMID- 10709014 TI - Musculoskeletal Injuries in the Workplace: Defining Quality Care. AB - Quality health care for a specific medical condition may be defined as adherence to an algorithm in which decision points are based on established medical practice as supported in the literature. The decision points can be considered either a "stan-dard of care" if there is definitive scientific evidence for their validity or a "guide-line for care" if there is only a consensus of medical opinion available. Algorithms for musculoskeletal injuries can be and have been successfully applied to patients in the workers' compensation setting. They can function as a concurrent surveil-lance system and are well accepted by physicians, patients, and industry if imple-mented by unbiased medical experts. A high level of quality care is attained by following such algorithms. Other goals achieved are early functional restoration as measured by return to work, a more efficient use of diagnostic studies, and avoidance of unnecessary therapeutic interventions, with the result that treatment is more cost-effective. Such a program that strives for high-quality care and emphasizes appropriate utilization will realize cost savings that may be far greater and longer lasting than the financial saving seen with arbitrary spending caps and fee controls. PMID- 10709015 TI - Patellofemoral Pain After Total Knee Arthroplasty. AB - The incidence of patellofemoral complications after total knee arthroplasty has been reported to range from 2% to 7%. Such complications include pain, sub luxation, dislocation, loosening, and wear. Usually these complications are attrib-utable to prosthetic design or surgical technique. Today, it is understood that patellofemoral prostheses must have a degree of congruence; must allow smooth, not abrupt, motion; and must restore a relatively normal size relationship between the patella and the femur. Surgical technique requires strict attention to (1) restoration of the patellofemoral spacing while avoiding "overstuffing" of the patellofemoral compartment; (2) accurate superior and medial positioning of the patellar component; (3) restoration of the rotational alignment of the femoral and tibial components; and (4) appropriate balancing of the patellofemoral soft tissues. PMID- 10709016 TI - Indirect Fracture Reduction: A Technique for Minimizing Surgical Trauma. PMID- 10709017 TI - Nonsteroidal Anti-inflammatory Drugs: Making the Right Choices. AB - Nonsteroidal anti-inflammatory drugs (NSAIDs) are among the most prescribed pharmacologic agents in medicine. The ability of these drugs to decrease inflammation is linked to their inhibitory effect on the synthesis of prostaglandins. This mechanism also results in toxicity that can cause gastrointestinal ulceration and bleeding, renal failure, and worsening of preexisting congestive heart failure. The superiority of one NSAID over another has not been clinically demonstrated in musculoskeletal conditions, nor has the efficacy of NSAIDs in noninflammatory rheumatic conditions been shown to be better than that of simple analgesics, such as acetaminophen. The use of these drugs, particularly in the elderly patient with osteoarthritis, should be carefully considered, and alternative, less toxic therapies should be sought whenever possible. PMID- 10709018 TI - Valgus Extension Injuries of the Elbow in the Throwing Athlete. AB - Valgus extension injuries of the elbow are common among throwing athletes. There is a wide spectrum of these injuries, ranging from early soft-tissue inflammatory changes to attenuation and incompetence of the ulnar collateral ligament, as well as bone changes, including chondromalacia, osteochondritis, and osteophyte formation. Early treatment should be directed toward decreasing pain and inflammation, followed by global strengthening and conditioning of the elbow with special emphasis on plyometric strengthening of the flexor-pronator musculature. In patients who remain symptomatic after an initial course of nonoperative treatment, arthroscopy of the elbow can address many of the later changes, including chondromalacia, osteochondritis, and osteophyte formation. Reconstruction of the ulnar collateral ligament should be reserved for those patients who wish to remain active at a highly competitive level and for whom rehabilitation and less invasive procedures have failed. Appropriate rehabilitation remains the cornerstone of successful treatment of these patients, facilitating their return to competitive play. PMID- 10709020 TI - Knee Bracing. AB - The authors present an overview of the design and functional features of knee braces and their relationship to knee biomechanics. Four types of knee braces prophylactic, rehabilitative, functional, and patellofemoral-have been developed to cover the wide variety of indications in patients who have suffered knee injuries or hope to prevent them. Important considerations when choosing specific brace types are discussed, and summaries of relevant research are presented. Clinical criteria for brace selection are offered to help physicians and sports medicine professionals in choosing the right brace for each patient. PMID- 10709019 TI - Persistently Painful Sprained Ankle. AB - Chronic discomfort sufficient to limit activity may affect 20% to 40% of patients after an ankle sprain. These patients complain of vague and diffuse pain, most often localized to the lateral and/or anterolateral aspect of the ankle. They may also complain of a giving-way sensation, swelling, stiffness, and locking and crepitation. Examination may show tenderness, swelling, and reduced range of motion, especially in dorsiflexion. Ankle instability is sometimes demonstrable. Severe cases exhibit discoloration, glossy skin, and temperature changes suggestive of reflex sympathetic dystrophy. Incomplete rehabilitation is the most common cause of chronic pain. Other common problems are intra-articular lesions (e.g., osteochondral and meniscoid lesions), chronic instability, undetected syndesmotic or deltoid sprains, chronic tendon degeneration, stress fractures, and, in rare cases, congenital lesions and tumors. Reflex sympathetic dystrophy occurs occasionally, even after minor trauma. With correct diagnosis and appropriate treatment, it is often possible to restore acceptable ankle function PMID- 10709021 TI - Heterotopic Ossification: Two Rare Forms and What They Can Teach Us. AB - Heterotopic ossification is characterized by the formation of normal bone at ectopic soft-tissue locations. Regardless of the etiology of heterotopic ossification, requisite pathogenetic conditions include an inductive signal capable of stimulating morphogenesis, a population of inducible osteoprogenitor cells, and a heterotopic environment conducive to osteogenesis. Two rare heritable and developmental forms of heterotopic ossification, fibrodysplasia ossificans progressiva and progressive osseous heteroplasia, provide valuable clinical and pathogenetic insights into heterotopic ossification in humans. A fundamental understanding of the developmental and molecular pathology of these disorders may lead to more effective strategies for preventing and treating heterotopic ossification in humans. PMID- 10709022 TI - Tibial Plafond Fractures: Changing Principles of Treatment. AB - Tibial plafond fractures from axial loading are high-energy injuries with significant associated soft-tissue damage. New classification methods include detailed anatomic subgroupings and highlight the soft-tissue injury. The traditional treatment of this intra-articular fracture with open reduction and internal fixation resulted in high rates of wound breakdown and infection. Treatment of these complications is lengthy and costly and not infrequently results in a poor outcome. Newer techniques using external fixation minimize disturbance of the soft-tissue envelope and have decreased these complications. Because the long-term outcome with all techniques is variable and often depends on factors beyond the surgeon's control, it is particularly important to avoid complications of initial treatment. Longer follow-up will determine whether patients treated with these techniques have a different rate of arthrosis. PMID- 10709023 TI - Radiolucent Lesions of the Extremities. AB - There are numerous conditions that produce a radiolucent lesion in a bone. Many of these are benign and of little consequence and need only occasional observation, as they usually heal spontaneously. A few are benign but do not heal spontaneously and require a limited operation. Others are malignant and must be removed surgically or irradiated. The physician evaluating the radiolucent lesion must be able to distinguish lesions that should be observed from those that should be further evaluated or treated. It is unnecessary to evaluate every radiolucent lesion as if it were a malignant tumor. With an understanding of the potential lesions and how they present, it is possible to construct an algorithm that can be used to organize an efficient and appropriate evaluation. PMID- 10709024 TI - Thoracic Outlet Syndrome. AB - The manifestations of thoracic outlet syndrome vary according to which of the neurovascular structures are affected. To provide optimal treatment, the pathogenesis must be understood in terms of both the anatomic variants and the dynamic factors. The diagnosis is primarily clinical, although ancillary diagnostic studies are useful in selected patients. Following a careful examination, the orthopaedic surgeon should be able to initiate a program of appropriate therapy depending on the nature and severity of the clinical manifestations. Initial treatment is oriented toward postural reeducation and periscapular muscle strengthening. Glenohumeral instabilities and painful upper limb conditions that cause disuse atrophy must be addressed. Operative treatment is reserved for patients in whom a conservative program has failed and for those with significant neural or vascular deficits. The surgeon must be cognizant of the potential complications of the various procedures used to correct thoracic outlet syndrome. Proper selection of surgical candidates should produce significant improvement in most patients. PMID- 10709025 TI - Rheumatoid Arthritis of the Foot and Ankle. AB - Rheumatoid arthritis of the foot and ankle can be a debilitating problem, particularly for patients who have undergone successful hip or knee arthroplasty. Optimal medical management, use of orthotic devices, and surgical intervention are essential components of patient care. Forefoot involvement with hallux valgus and lesser metatarsophalangeal joint subluxation and dislocation are the most common findings. Reconstruction usually requires lesser metatarsophalangeal joint excisional arthroplasty and first metatarsophalangeal joint arthrodesis. Midfoot tarsometatarsal and intertarsal involvement is treated with orthotic devices and intertarsal fusion for advanced arthropathy. Hindfoot involvement frequently leads to pes planovalgus deformity, which may require isolated talonavicular arthrodesis if treated early or triple arthrodesis for advanced destruction. Ankle involvement is less frequent; when it is unresponsive to conservative measures, ankle symptoms may be improved by arthrodesis. Although great advances have been made in medical and surgical management of rheumatoid arthritis, the disease remains a serious problem. Through prudent use of medical management, orthotic devices, and other conservative measures as well as surgical intervention, long-term function can be enhanced greatly. PMID- 10709026 TI - Pediatric Hematogenous Osteomyelitis: New Trends in Presentation, Diagnosis, and Treatment. AB - The character of acute hematogenous osteomyelitis (AHO) in North American children has changed significantly during the past several decades. Although the typical clinical picture of established acute osteomyelitis in children (illness, dehydration, and an acutely painful limb) is still seen, more subtle presentations appear more frequently. Children often present with subacute osteomyelitis. Less common variants include Brodie's abscess, subacute epiphyseal osteomyelitis, and chronic recurrent multifocal osteomyelitis. Some patients present with a bone lesion that may be confused with other disease entities, including neoplasms. Biopsy is often needed to clarify the diagnosis. With the trend toward more invasive procedures in the neonatal intensive care unit, neonatal osteomyelitis is also seen more frequently. Advances in imaging technology, particularly improvements in technetium bone scanning and the advent of magnetic resonance imaging, have contributed to more precise diagnosis and better management of AHO. With the increased concern about medical economics, the recent trend toward decreasing the duration of intravenous antibiotic treatment of these infections appears to be appropriate as long as certain criteria are met. Neither surgery nor antibiotics alone will be associated with successful treatment in all cases, and this fact may explain the rare but continued morbidity that is still seen in children with AHO. PMID- 10709028 TI - Southeastern academy of prosthodontics-founded in 1952 PMID- 10709027 TI - Lower-Extremity Local Flaps. AB - Some soft-tissue defects of the lower extremities can be covered reliably with local flaps. Five such flaps--the tensor fascia lata, gastrocnemius, soleus, posterior tibial artery fasciocutaneous, and dorsalis pedis flaps--are described. If the indications for each flap are understood and the vascular pedicle is carefully preserved, these flaps can be used to provide relatively simple and reliable coverage of selected soft-tissue defects on the lower extremities. However, the indications must not be overextended in an attempt to avoid a free tissue transfer. The gastrocnemius flap is most often used. It reliably covers common defects about the knee and the proximal tibia. A skin graft is required for the gastrocnemius flap, as well as the soleus flap, which covers the midportion of the tibia. The soleus requires deeper dissection of the calf for elevation. The tensor fascia lata flap and the more recently described posterior tibial artery fasciocutaneous flap are relatively easy to raise, but there are fewer orthopaedic indications for their use. The dorsalis pedis cutaneous flap is technically more demanding, but it can be used to cover difficult defects around the ankle. PMID- 10709029 TI - American academy of maxillofacial prosthetics-founded in 1953 PMID- 10709030 TI - American equilibration society- founded in 1955 PMID- 10709031 TI - Measurement in dentistry. PMID- 10709033 TI - New impression system for ITI solid abutment cementable restoration. PMID- 10709032 TI - Fixed partial dentures combining both resin-bonded and conventional retainers: a clinical report. PMID- 10709034 TI - Impression procedure for a progressive sclerosis patient: a clinical report. PMID- 10709035 TI - Prosthetic rehabilitation after maxillectomy and temporalis flap reconstruction: a clinical report. PMID- 10709036 TI - Tooth wear treated with direct composite restorations at an increased vertical dimension: results at 30 months. AB - STATEMENT OF PROBLEM: Severe tooth wear localized to the anterior maxillary or mandibular teeth with loss of interocclusal space is difficult to manage. PURPOSE: This study evaluated the outcome of composite restorations placed at an increased vertical dimension of occlusion in such patients. METHODS AND MATERIAL: Sixteen patients were restored with 104 restorations in 2 groups. In group A, Durafill composite and Scotchbond Multipurpose dentine adhesive system were used to place direct anterior restorations (N = 52). In group B, Herculite XRV composite and Optibond dentine bonding agent was used (N = 52). The restorations were placed at an increased vertical dimension of occlusion creating a posterior disclusion of 1 to 4 mm. RESULTS: Clinical follow-up showed that the posterior occlusion remained satisfactorily restored after a mean duration of 4.6 months (range 1 to 11 months). Mean follow-up of 30 months has shown a combined success rate of 89.4% for both groups with 93 of the restorations remaining in service. Maintenance in group A was high with 33 failures, but low in group B with 6 failures. Patient satisfaction was reported as good. CONCLUSION: Direct composite restorations may be a treatment option for localized anterior tooth wear. PMID- 10709037 TI - Clinical evaluation of resin-bonded gold alloy veneers. AB - STATEMENT OF PROBLEM: Clinical performance of resin-bonded alumina-abraded type III cast gold alloy veneers has not been reported. PURPOSE: This retrospective study evaluated the survival of such restorations for the management of tooth wear and other restorative problems. MATERIAL AND METHOD: This clinical study evaluated 25 patients between 14 and 60 years of age treated with a total of 158 cast gold veneers cemented with Panavia Ex cement. Restorations placed between January 1990 and February 1996 (mean age 48 months) were evaluated. Factors, including age, gender, operator, site, design, the extent of dentin exposure, the presence of previous restorations, dynamic and static occlusion, rubber dam, cause, and methods of interocclusal space creation, were evaluated with Cox regression. Survival probability was evaluated with Kaplan-Meier analyses. Significance was taken as P <.05. RESULTS: Failures occurred for 7.6% of restorations in 28% of patients. Alumina-abraded gold alloys cemented with Panavia Ex cement had an estimated 89% survival probability at 60 months using Kaplan-Meier analyses. The only variable to statistically influence survival was operator (P <.0001). Operator differences were due to a single operator who placed only 2 restorations in the same patient, both of which failed unusually quickly. CONCLUSION: Resin-bonded alumina-abraded type III cast gold alloys can be successfully used to restore both anterior and posterior teeth and were associated with an estimated 89% survival probability at 60 months. PMID- 10709038 TI - Dimensional accuracy of an epoxy resin die material using two setting methods. AB - STATEMENT OF PROBLEM: Resinous die materials have several important advantages including strength, abrasion resistance, and detail reproduction. Despite these advantages, the shrinkage of resinous die materials during polymerization has limited their widespread acceptance. PURPOSE: This study determined whether a retarded setting reaction could improve the accuracy of an epoxy resin die system, and compared the accuracy of this epoxy resin system with gypsum-based die materials. MATERIAL AND METHODS: Four groups were compared: an epoxy resin manipulated according to manufacturer's instructions (Ivoclar, Schaan, Liechtenstein); the same epoxy resin manipulated to undergo a retarded set; a high-strength high-expansion gypsum (Die Keen); and a resin-filled gypsum (Resin Rock). Ten dies were fabricated for each material from a master metal model using conventional prosthodontic laboratory techniques. The mean of 3 independent measurements recorded using a toolmaker's microscope and digital positioners was used to describe each die. RESULTS: One-way ANOVA revealed that significant differences existed among the materials (P <.0001). Tukey's multiple comparisons testing ranked the dies as follows, from largest to smallest: high-strength high expansion gypsum, resin-filled gypsum, master metal model, retarded epoxy, and manufacturers epoxy (P <.05). CONCLUSION: Retarding the setting reaction of an epoxy resin die material improved its accuracy. Of the materials tested, retarded set epoxy dies had the least mean dimensional change from the metal master. Epoxy resin die materials had a net shrinkage, but the gypsum-based materials had a net expansion. The epoxy resin materials exhibited more variability than the gypsum based materials. PMID- 10709039 TI - Margin adaptation of indirect composite inlays fabricated on flexible dies. AB - STATEMENT OF PROBLEM: Indirect composite restorations can be made in 1 appointment using a flexible die. Interactions between different impression materials and flexible die materials may affect the accuracy of fit and margin adaptation of the restoration. PURPOSE: This study compared the margin adaptation of composite inlays made using the following 5 impression/flexible die material combinations; condensation silicone/polyvinyl siloxane (CS/PVS), wash viscosity polyvinyl siloxane/medium or heavy viscosity polyvinyl siloxane (PVS/PVS), irreversible hydrocolloid impression/medium viscosity polyvinyl siloxane (IH/PVS), wash viscosity polyvinyl siloxane impression/polyether (PVS/PE), with composite inlays made using a control system of a wash viscosity polyvinyl siloxane impression and a type IV stone die. MATERIAL AND METHODS: For each test and control system, 10 impressions were made of a class II composite inlay preparation in a metal master die. One die was made from each impression and one composite inlay was made and finished on each die (a total of 60 inlays). Inlays were placed on the master die and the margin opening at the buccal, distal, and gingival sites was recorded with a measuring microscope (x40 magnification). RESULTS: The overall mean +/- SD margin openings of inlays made from the systems were as follows: PVS wash/PVS heavy viscosity 149.5 +/- 107. 4 microm; PVS wash/PVS medium viscosity 87.4 +/- 63.0 microm; IH/PVS medium viscosity 76.7 +/- 48.9 microm; CS/PVS 73.3 +/- 48.7 microm, PVS wash viscosity/PE 64.0 +/- 44.3 microm, PVS wash viscosity/stone 53.9 +/- 48.3 microm. Composite inlays made using the PVS wash viscosity/PVS heavy viscosity system had significantly larger distal, gingival, and overall mean margin openings than all other inlays (ANOVA and Fisher PLSD test; P =.05). The separating medium required between some impression and die materials did not work consistently. Composite inlays fabricated on dies made of material different than the impression material had mean buccal, distal, gingival, and overall margin openings < or =100 microm. CONCLUSION: Composite inlays made on the CS/PVS, IH/PVS medium viscosity, PVS wash viscosity/PE flexible dies, and control PVS wash viscosity/stone dies had statistically similar (P =.05) mean buccal, distal, gingival, and overall mean margin openings that were < or =100 microm. Composite inlays made on dies that were made of the same type of material as the impression material (PVS/PVS) had mean gingival margin openings >100 microm that were significantly larger than all other systems tested (P =.05). PMID- 10709040 TI - Fit of titanium and a base metal alloy metal-ceramic crown. AB - STATEMENT OF PROBLEM: Titanium has been widely used in dental implantology. However, problems remain for use of titanium in prosthodontics. PURPOSE: This study evaluated the marginal and inner fit of metal-ceramic restorations and frameworks made with a nickel-chromium alloy (Remanium CS) and a commercially pure titanium (Rematitan). Fit was compared with SEM before and after firing of ceramic materials. MATERIAL AND METHODS: Sixteen metal frameworks were cast with each casting material, 8 frameworks were used directly to evaluate measurements for fit (RCS: Remanium CS frameworks and TI: Rematitan frameworks). Porcelain was applied on the other 8 frameworks (RCSP: Remanium CS-porcelain and TIP: Rematitan porcelain), fired, and then the fit was measured. Measurements were recorded at 3 points for marginal fit evaluation and at 3 points for inner fit. RESULTS: Significant differences were recorded among the occlusal (JK) values of groups tested (P <.05). Marginal fit and inner fit of Remanium CS frameworks were better than Rematitan frameworks. However, there were no substantial differences detected among groups after firing the ceramic superstructures. CONCLUSION: The fit of the base metal alloy metal-ceramic crowns was better than the commercially pure titanium metal-ceramic crowns. However, both artificial crowns were clinically acceptable. PMID- 10709041 TI - Proximal plate in conventional circumferential cast clasp retention. AB - Improper guiding planes are commonly used with conventional removable partial denture cast clasps. Short guiding planes with improper proximal plates decrease retention and cause repetitive traumatic movement of abutments. This article discusses the importance of guiding planes and proximal plates for conventional tooth-supported removable partial dentures with circumferential clasp. The retention force of the clasps was analyzed geometrically. If the guiding plane is short or the prepared surface is oblique, the contact between the proximal plate and the guiding plane or oblique plane diminishes at an early stage, which results in reduction of retention. The guiding plane should be parallel to the path of insertion and must be of a certain length for adequate retention. An elongated proximal plate that is parallel to the path of insertion may compensate for retention deficiency when an adequate guiding plane cannot be prepared. PMID- 10709042 TI - Comparison of impression materials for direct multi-implant impressions. AB - STATEMENT OF PROBLEM: Given that meticulous implant prosthodontic procedures are recommended to obtain the best possible intraoral fit, impression materials that are suitable for use with a direct impression technique warrant further investigation. PURPOSE: This in vitro study compared the amount of torque required to rotate a square impression coping in an impression and evaluated the accuracy of solid implant casts fabricated from different impression materials. MATERIAL AND METHODS: Two direct transfer implant impressions were made using 8 impression materials; the torque required to rotate an impression coping in the impressions was calculated. Ten direct transfer implant impressions were made from the master model and poured in a die stone (Resin Rock) for 3 of the 8 initial impression material groups. Linear distances between steel balls placed on each abutment replica were measured with a traveling microscope to determine distortion in the impression procedure for each group. Data were analyzed (P =.05) with ANOVA and Ryan-Einot-Gabriel-Welsch multiple range test for post hoc. RESULTS: With a 1-way ANOVA, average torque values among the material groups differed significantly (P =.001). Polyether (medium consistency) was found to produce the highest overall torque values, followed by addition silicone (high consistency), and then polysulfide (medium consistency). Statistically significant difference was also found among the 3 material groups' mean absolute cast error using a 1-way ANOVA (P =.0086). Implant casts made from polyether (medium) or addition silicone (high) impressions were significantly more accurate than casts made from polysulfide medium impressions. CONCLUSION: On the basis of the results of this study, the use of either polyether (medium) or addition silicone (high) impression is recommended for direct implant impressions. PMID- 10709043 TI - Theoretical cantilever lengths versus clinical variables in fifty-five clinical cases. AB - STATEMENT OF PROBLEM: Cantilever loading increases loads distributed to implants, potentially causing biomechanical complications. The implemented length is often less than what is considered to be optimal. PURPOSE: This study investigated the effects of clinical variables on predicted cantilever lengths. Theoretically, calculated maximum cantilever was defined as the length that would not cause gold screw loosening or fatigue failure. The variables investigated included number and distribution of implants, arches placed, and the clinician's "optimal" cantilevers. MATERIAL AND METHODS: Implant and prosthesis location coordinates of 55 clinical cases were determined from casts. The distribution of an applied 143 N vertical load to implants was calculated through the Skalak model for more than 500 loading sites. Gold screw joint overload was assumed to occur at 200 and 250 N in compression and tension. Calculated lengths were compared with clinical variables. RESULTS: For a set number of implants, the relationship between calculated cantilever length and anterior-posterior spread was linear. The sum of length on both sides versus prosthesis length between the most distal implants was linear, regardless of the number of implants. Predicted satisfaction was defined as calculated length greater than the clinicians' optimal length. Satisfaction rates were 100%, 56%, 33%, 8%, and 0% for cases supported by 8 and 7, 6, 5, 4, and 3 implants (44% overall), respectively. Ninety-eight percent of cases with anterior-posterior spreads greater than 11.1 mm were satisfied. CONCLUSION: Within the limitations of the model, predicted complications of the gold screw joint may be reduced if: (1) cantilever length is less than calculated from linear equations, and (2) anterior-posterior spread is greater than 11.1 mm. PMID- 10709044 TI - Effects of abutment size and luting cement type on the uniaxial retention force of implant-supported crowns. AB - STATEMENT OF PROBLEM: The assumption that increasing the diameter of the abutment/crown components will provide greater resistance to crown loosening forces than standard-sized components has not been reported either with clinical trials or in the laboratory. PURPOSE: This study attempted to determine what effect abutment dimension and type of luting agent have on the retention of the prosthetic crown. METHODS AND MATERIAL: Test specimens consisted of standard, wide, and "experimental" CeraOne titanium abutments and matching CeraOne gold cylinders cemented with a zinc phosphate permanent or a zinc oxide eugenol provisional cement. The mean uniaxial force (Newtons) and the load (MPa) required to dislodge the cylinder from the abutment was determined. Statistical analysis of the sample data was performed using a 2-way analysis of variance test (alpha=.05). RESULTS: Mean uniaxial resistance force (Newtons) was significantly greater for zinc phosphate cement than for zinc oxide cement (P <. 001). Abutment size was a significant factor when permanent luting cement is used (P <.001). Retention strength per unit area (MPa) of the wide abutments was lower than the standard size and "experimental" abutments. CONCLUSION: Permanent luting cement produced uniaxial retention forces approximately 3 times greater than provisional cement. The increase in surface area provided by a wide abutment did not result in an improvement in retention strength over the standard abutment. PMID- 10709045 TI - Pilot study of conditioner/primer effects on resin-dentin bonding after provisional cement contamination using SEM, energy dispersive x-ray spectroscopy, and bond strength evaluation measures. AB - STATEMENT OF PROBLEM: Previous studies have shown that provisional cement cannot be completely removed from the dentin surface and the remnants inhibit adhesion of resin cement to dentin. PURPOSE: This study investigated the effects of dentin conditioners on resin-dentin bonding after provisional cement contamination. MATERIAL AND METHODS: A total of 216 bovine dentin specimens were divided into 2 groups with and without provisional cement coating on the dentin surface. Each group was further divided into 3 subgroups with 2 conditioning methods and without dentin conditioning (36 specimens per subgroup). The 2 methods applied were (1) phosphoric acid etching and (2) phosphoric acid etching, followed by sodium hypochlorite gel. Provisional cement was completely removed with a curette before dentin conditioning. Stainless steel rods were luted to dentin surfaces with Panavia 21 resin cement. Tensile bond strengths were measured before and after thermocycling. The dentin surface was analyzed by SEM and energy dispersive x-ray spectroscopy. Three-way ANOVA and Scheffe's comparison test were performed (P =. 05). RESULTS: Bond strength was significantly influenced by provisional cement application, type of dentin conditioning, and thermocycling. Bond strength decreased when treated with provisional cement, but the decreased strengths were restored to their original values or higher by both dentin conditioners used. Specimens treated with sodium hypochlorite gel applied after phosphoric acid etching demonstrated the highest bond strengths with good stability after thermocycling. CONCLUSION: Application of sodium hypochlorite gel after phosphoric acid etching was more effective than etching with phosphoric acid alone to eliminate the inhibitory effect of provisional cement remnants on adhesion between resin cement and dentin. PMID- 10709046 TI - Comparative study of water sorption, solubility, and tensile bond strength of two soft lining materials. AB - STATEMENT OF PROBLEM: Two soft denture lining materials with distinct chemical compositions were evaluated to determine whether these compositional variations manifest themselves in property differences. PURPOSE: This study evaluated and compared the water sorption, solubility, and tensile bond strength of a recently introduced silicone-based soft liner (Luci-sof) and a plasticized acrylic resin soft liner (Permasoft) using 2 processing techniques, laboratory-processed and autopolymerized at chairside, for the latter. MATERIAL AND METHODS: For water sorption and solubility testing, 24 disks (45 mm in diameter and 1 mm in thickness) were prepared for each group, stored in distilled water at 37 degrees C, and tested after 1, 4, and 6 weeks. The disks used for the first week were again tested after 4 and 6 weeks of continuous storage. Their weight was recorded and sorption and solubility were calculated using 2 methods. Two PMMA cylinders (25 mm in diameter, 25 mm in length), joined together by the soft liner, were used to determine the bond strength in tension at a loading rate of 2 mm/min after 48 hours and 12 weeks aging in distilled water. RESULTS: Permasoft had higher solubility (1.50% +/- 0.20% laboratory-processed and 1.42% +/- 0.16% autopolymerized at chairside) and sorption (2.45% +/- 0.36% laboratory-processed and 1. 76% +/- 0.08% autopolymerized at chairside) than Luci-sof (0.17% +/- 0.09% solubility and 0.41% +/- 0.17% sorption) after 6 weeks of aging. For tensile bond strength, there was no significant difference between each respective control and the 12-week bond strength for each material. However, Luci-sof (1.16 +/- 0.27 MPa) had a significantly higher tensile bond strength than Permasoft (0. 50 +/- 0.18 MPa laboratory-processed and 0.44 +/- 0.09 MPa autopolymerized at chairside). CONCLUSION: On the basis of lower water sorption and solubility and higher tensile bond strength, Luci-sof may provide better clinical success. PMID- 10709047 TI - Method for marginal measurements of restorations: accessory device for toolmakers microscope. AB - Standardization of measurements for marginal fit of castings is critical. This study describes the fabrication of a device that allowed fixation of specimens on a Toolmakers microscope with identical conditions according to tri-dimensional positioning of specimens, measuring location, and seating force. The device also allows mapping of the marginal discrepancies on the entire marginal perimeter of the tooth preparation. PMID- 10709048 TI - New approach for making accurate cross sections of casts. AB - This article describes a new procedure for producing cross sections of a cast, a denture, or an impression and a method of presenting the results in graphic form to show variance. To test the method, identically colored, duplicate dentures were embedded in plaster blocks formed from a precisely engineered and calibrated brass box. A modified jig and saw allowed the plaster blocks to be positioned, held, and sectioned at any required location with a high degree of accuracy. The sectioned surfaces of the duplicate dentures were scanned. The resultant profiles were registered in location frames on the computer screen, traced by computer to produce outlines, and printed onto transparent film for the purpose of comparison. In vitro results showed the relationship between congruent outer forms and provided clear visual evidence of the accuracy of the procedure used and a way to further in vivo use. PMID- 10709049 TI - Immediate maxillary denture base extension for posterior palatal seal. AB - A procedure for extension of the maxillary denture base for development of a posterior palatal seal is described. The technique involves provisional extension with paraffin wax and adding direct relining resin supported by a silicone putty core. This simple, quick procedure achieves immediate recovery of retention for underextended maxillary dentures without additional laboratory procedures. PMID- 10709050 TI - Chairside fabrication of provisional implant-supported prosthesis using impression copings. PMID- 10709051 TI - Fabricating a fit checker. PMID- 10709052 TI - Prosthetic above-knee femoropopliteal bypass grafting: five-year results of a randomized trial. AB - PURPOSE: This trial was designed to identify factors affecting patency rates of primary prosthetic above-knee femoropopliteal bypass grafts at 5 years. METHODS: A multi-institutional, prospective trial randomized 240 patients to compare patency rates of Gore-tex and Hemashield above-knee femoropopliteal bypass grafts at 5 years. Univariate comparisons of patency between levels of each prognostic variable were made with the Kaplan-Meier method. Variables that had a univariate P value less than.25 or those known to be important were submitted to a Cox regression analysis. RESULTS: The patient survival rate at 5 years was 59.4%. There were no differences in primary or secondary patency rates at 5 years between the two graft materials (primary, 45% vs 43% and secondary, 68% vs 68%). The risk for graft occlusion was significantly increased for patients younger than 65 years (2.1; P =.001) and for grafts with a diameter less than 7 mm (1.65; P =.0219). Variables with no apparent independent effect on patency rates were smoking status, runoff, diabetes mellitus, sex, presenting symptoms, and postoperative treatment with aspirin or Coumadin. Noninvasive test results were not predictive of subsequent graft function. CONCLUSION: Although the type of prosthetic used for above-knee femoropopliteal bypass grafts does not affect 5 year patency rates, age and graft size do influence results. These factors should be considered before a prosthetic bypass grafting procedure. Furthermore, these data should serve as a contemporary standard, with which evolving and conventional procedures can be compared. PMID- 10709053 TI - Long-term results of arterial allograft below-knee bypass grafts for limb salvage: a retrospective multicenter study. AB - PURPOSE: Arterial allografts (AAs) have been recently reconsidered in the treatment of critical limb ischemia when vein material is absent, because of the disappointing results with artificial grafts. The aim of this study was to report the results observed in three centers where AAs were used for infrainguinal reconstruction in limb-threatening ischemia. METHODS: Between 1991 and 1997, 165 AA bypass procedures were performed in 148 patients (male, 90) with a mean age of 70 years (range, 20-93 years). Indications for operation were rest pain in 54 cases and tissue loss in 111 cases. Mean resting ankle pressure was 53 mm Hg in 96 patients who did not have diabetes and mean transcutaneous pressure of oxygen was 10 mm Hg in 52 patients who did have diabetes. In 123 cases (75%), there was at least one previous revascularization on the same limb. AAs were obtained from cadaveric donors. The distal anastomosis was to the below-knee popliteal artery in 34 cases, to a tibial artery in 114 cases, and to a pedal artery in 17 cases. RESULTS: At 30 days, the mortality rate was 3.4%; the primary patency rate was 83.3%; the secondary patency rate was 90%; and the limb salvage rate was 98%. During follow-up (mean, 31 months), 65 grafts failed primarily. Causes of primary failure were thought to be progression of the distal disease in 15 cases, myointimal hyperplasia in 16 cases, graft degradation in 10 cases (four dilations, three stenoses, two ruptures, and one dissection), miscellaneous in eight cases, and not known in 16 cases. Primary patency rates at 1, 3, and 5 years were, respectively, 48.7% +/- 4%, 34.9% +/- 6%, and 16.1% +/- 7%. Secondary patency rates at 1, 3, and 5 years were, respectively, 59. 8% +/- 4%, 42.1% +/- 5%, and 25.9% +/- 8%. Limb salvage rates at 1, 3, and 5 years were, respectively, 83.8% +/- 3%, 76.4% +/- 5%, and 74.2 % +/- 8%. CONCLUSION: AA leads to an acceptable limb salvage rate but poor patency rates. A randomized trial that will compare AAs and polytetrafluoroethylene should be undertaken. PMID- 10709054 TI - The tourniquet revisited as an adjunct to lower limb revascularization. AB - PURPOSE: The purpose of this study was to evaluate the role and efficacy of the tourniquet in lower limb revascularization. METHODS: During a 3-year period, 195 patients underwent 205 infrainguinal reconstruction operations in the lower extremity. These patients underwent bypass with a tourniquet and inflow occlusion (group 1) or bypass without a tourniquet (group 2). The type of infrainguinal reconstruction, tourniquet ischemia time, blood loss, and complications related to tourniquet use were recorded. A subset of patients underwent serial muscle biopsies. Specimens from calf muscle were taken just (1) before application of the tourniquet, (2) before tourniquet release, and (3) once wound closure was initiated. These biopsy specimens were studied by histochemical staining and also analyzed for phosphorylase enzyme, a marker for subcellular ischemia. RESULTS: One hundred eleven patients underwent 117 infrainguinal reconstruction procedures in which the tourniquet and inflow occlusion were used. These patients were matched against 84 patients who underwent 88 infrainguinal reconstructions without the use of the tourniquet. Complete hemostatic control in group 1 was obtained in 108 of the procedures (92%). Eight percent of the procedures required minor additional techniques to obtain complete hemostasis; in two instances, the tourniquet was removed because it did not provide hemostasis. Mean tourniquet time was less than 1 hour for all reconstruction groups. There were no instances of neurologic deficit, thrombosis of distal vessels, or vascular injury that was related to the use of a tourniquet. A comparison of the two groups revealed no differences with regard to overall blood loss (P =.63) or duration of operation (P = 0.60), observations that reflect the complexity of the cases rather than the use or nonuse of a tourniquet. When tourniquet control was used, we noted a definite decrease in the time for the distal dissection, because total vascular control with extensive dissection was unnecessary. Histochemical analysis with phosphorylase revealed a conversion of tissue with active enzyme activity to a low level with tourniquet use (P <.05). CONCLUSION: The use of a tourniquet for lower limb revascularization is safe and effective and improves visualization of the operative field. Less dissection of the target vessels is required. With a combination of the nonuse of clamps and other occluding devices, we project a decrease in host hyperplastic response that will, in turn, impact favorably on patency rates. The possibility exists that early failure may be prevented by avoiding the application of traumatic forces to diseased and brittle or calcified arteries. In this study, tourniquet time had no impact on overall operative procedural time, although certain phases of the operation were clearly shortened and facilitated, particularly in complex and difficult reconstructions. Histochemical changes found in muscle biopsy specimens did not adversely impact patients clinically, but further investigation is required to elucidate subcellular events. PMID- 10709055 TI - Expression of myosin heavy chain isoforms in skeletal muscle of patients with peripheral arterial occlusive disease. AB - PURPOSE: Peripheral arterial occlusive diseases (PAODs) not only compromise blood flow but lead to a series of subsequent metabolic and structural changes in the relevant muscles. Changes in myofibrillar proteins (eg, of myosin heavy chain [MHC] isoforms), one of the determinants of muscle structure as well as of muscular function, have not been reported in patients with PAOD and were therefore the aim of this study. METHODS: Thirteen consecutive patients with PAOD were examined (clinical stage according to Fontaine II, three patients; III, three patients, and IV, seven patients) and compared with five age-matched control patients who had been in traffic accidents. A calf muscle sample (gastrocnemius muscle) in the ischemic region was taken for MHC isoform analysis by sodium dodecyl sulfate polyacrylamide gel electrophoresis and silver stain, and the relative content of MHC isoforms was measured. RESULTS: Compared with the control patients, there was no significant change of MHC isoforms in patients with PAOD II. In patients with PAOD III, MHC IIb decreased significantly (P <.05) although MHC IIa remained unchanged; in patients with PAOD IV, both MHC IIa and IIb decreased significantly (P <.05). Accordingly, there was a progressive increase of the relative amount of MHC I with more critical ischemia in PAOD. CONCLUSION: In patients with PAOD, the content of MHC II decreased with a higher grade of ischemia. That seems to be consistent with an increased resistance to ischemia for myosin isoforms in the order of I more than in IIa more than IIb. Whether the decrease of MHC II in patients with PAOD is related to atrophy of muscle fibers or to muscle-fiber transition must be investigated further. PMID- 10709056 TI - Superficial femoral popliteal vein: An anatomic study. AB - OBJECTIVE: The superficial femoral popliteal vein (SFPV) has been used as an alternative conduit for both arterial and venous reconstructive surgery. Its popularity continues to grow, despite concern about the potential for venous morbidity after harvest. The purpose of this study was to determine an anatomic "safe" length of SFPV for harvest, assuming that the preservation of at least one valve and one significant collateral vein in the remaining popliteal vein (PV) segment can minimize venous morbidity. METHODS: Forty-four SFPVs were harvested from 39 cadaveric specimens. The length of both the superficial femoral vein (SFV) and PV was measured, and the number and location of valves and significant side branches (more than 2 mm in diameter) of the PV were measured. The Student two-tailed t test was used as a means of comparing vein lengths between the sexes. Correlation coefficients were determined for the effect of patient height on vein length, stratified by means of sex. RESULTS: Vein length (SFV mean, 24.4 +/- 4 cm; PV mean, 18.8 +/- 4 cm) varied with sex (male SFV mean, 28.1 +/- 5 cm; male PV mean, 21. 5 +/- 3 cm; female SFV mean, 22.6 +/- 4 cm; female PV mean, 18.4 +/- 3 cm; P =.01). Valve number (mean, 1.8 +/- 0.5) and location and collateral vein number (mean, 5 +/- 1.8) and location were variable and independent of height or sex. CONCLUSION: An anatomic "safe" length of SFPV for harvest to minimize venous morbidity would include all the SFV and 12 cm of PV in 95% of women and 15 cm of PV in 95% of men. We found that the male sex was a significant determinant for a longer safe length of vein that can be harvested. PMID- 10709057 TI - Effect of oral micronized purified flavonoid fraction treatment on leukocyte adhesion molecule expression in patients with chronic venous disease: a pilot study. AB - PURPOSE: The purpose of this study was to determine the effects of a micronized purified flavonoid fraction treatment on surface expression of leukocyte adhesion molecules in chronic venous disease (CVD). METHODS: Twenty patients with chronic venous disease were assessed with the use of clinical and Duplex scanning criteria. Consenting patients were treated for 60 days with a micronized purified flavonoid fraction treatment (500 mg twice daily). Blood was collected from a foot vein immediately before the start of treatment and within 1 week after the treatment was stopped. Neutrophil and monocyte surface adhesion molecule expression was determined by flow cytometry using the monoclonal antibodies to CD11b and CD62L. RESULTS: Neutrophil CD11b (248:212), monocyte CD11B (204:190), neutrophil CD62L (130:97 [P =.002]), and monocyte CD62L (170:121 [P =.03]) were determined, respectively, before and after treatment. All values are arbitrary units and represent median values. CONCLUSION: Micronized purified flavonoid fraction treatment for 60 days seems to decrease the surface expression of CD62L by neutrophils and by monocytes. The clinical significance of this finding needs to be explored further. It is feasible to use changes in the levels of these molecules as a marker for response to therapy in chronic venous disease. PMID- 10709058 TI - Prevalence of deep venous anomalies in congenital vascular malformations of venous predominance. AB - PURPOSE: The overall incidence of congenital vascular malformations in the general population is 1.5%. Approximately two thirds of them are malformations of venous predominance. Abnormalities of the deep venous trunks have been observed in association with large superficial compensatory varices in these type of malformations. Knowledge of the integrity of the deep venous system is important in their management because excision of the enlarged superficial veins may be deleterious if there is aplasia or hypoplasia of the deep venous trunks. The objective was to investigate the prevalence and nature of deep venous anomalies that occur in patients with congenital vascular malformations of venous predominance both in our series and in the series from the medical literature. METHODS: From the last 35 years of medical literature, we reviewed seven series of congenital vascular malformations that provided pertinent information on the subject of our study. We also reviewed our own series of 392 patients with congenital vascular malformations studied at Children's Hospital of Mexico City (1963-1983; n = 223 children) and at Walter Reed Army and National Naval Medical Centers (1984-1998; n = 169 children). Of 392 patients, 257 (65.5%) had malformations of venous predominance; these were the subject of our analysis. Prevalence of the following deep venous anomalies was recorded: phlebectasia, aplasia or hypoplasia of venous trunks, aneurysms, and avalvulia. Diagnosis was made by one or more of the following methods: Doppler scanning, duplex scanning, plethysmography, computerized tomography, magnetic resonance imaging, and angiography. RESULTS: At least one anomaly of the deep venous system was present in 47% of the congenital vascular malformations of venous predominance reviewed. Phlebectasia was recorded in 36% of the cases, and aplasia or hypoplasia of deep venous trunks was observed in 8% of the cases. Venous aneurysms also were present in 8% of the cases; avalvulia was recorded in 7% of the cases. CONCLUSION: Anomalies of the deep venous system occur in almost one half of congenital vascular malformations of venous predominance. The most common is the relatively innocuous phlebectasias that occur in over one third of cases. Aplasia/hypoplasia, venous aneurysms, and avalvulia were less frequent, each less than 10%; but failure to detect the latter three anomalies may lead to serious therapeutic errors. PMID- 10709059 TI - Venous outflow and inflow resistance in health and venous disease. AB - PURPOSE: The purpose of this study was to develop a physiologic method to measure outflow and inflow from the lower extremities and thus to quantify the degree of venous valvular insufficiency and venous obstructive disease. METHODS: Calibrated photoplethysmography was used in combination with passive changes in hydrostatic pressure, by leg elevation followed by repositioning of the leg to the original sitting position. With the principle of venous occlusion plethysmography, timed volume changes were then used to calculate the outflow and inflow. The inflow and outflow units were the percentage of optical reflectance (%OR) per minute. The respective resistances were calculated by identifying the hydrostatic pressure distance from the third intercostal space to the probe site that is inducing these site changes. The resistance units were millimeters of Mercury x minutes per %OR. RESULTS: Four groups of subjects were examined: normal individuals, patients with venous valvular insufficiency, deep venous thrombosis, and a combination of both. The most significant differences in outflow values were found between the control group (81.77% OR/min) and the deep venous thrombosis group (28.47% OR/min). In contrast, the most significant differences in inflow values were found between the control group (9. 67% OR/min) and the venous valvular insufficiency group (108.61% OR/min). The resistances changed correspondingly. CONCLUSION: The application of calibrated photoplethysmography in conjunction with induced changes in leg hydrostatic pressure proved to be an effective physiologic method to noninvasively quantify venous hemodynamics in normal control subjects, patients with venous valvular insufficiency, venous obstructive disease, or both. PMID- 10709060 TI - High diastolic flow velocities in severe internal carotid artery stenosis: a sign of increased surgical risk? AB - PURPOSE: We reviewed the history and preoperative investigations of patients with early postoperative neurologic events after carotid thromboendarterectomy in an attempt to identify risk factors for neurologic complications. METHODS: Patients with neurologic events/complications (S group, n = 14 patients) were compared with an age- and disease-matched control group (C group, n = 42 patients) selected from the whole carotid thromboendarterectomy material between 1987 and 1996. In this retrospective study, we re-evaluated the maximum systolic and end diastolic flow velocities within the internal carotid artery (ICA) using video recordings of preoperative Duplex ultrasound scan investigations. The flow velocity variables were compared with preoperative carotid angiography and intraoperative ICA stump pressure measurement. RESULTS: S-group did not differ from C-group concerning either cardiovascular risk factors or diseases, ipsilateral and contralateral angiographic grade of ICA stenosis, or history of cerebral infarctions. Nevertheless, in contrast to control subjects, patients with early postoperative major stroke had higher end diastolic flow velocities and lower ICA stump pressures. Patients with postoperative minor stroke, transient ischemic attack, or amaurosis fugax did not differ significantly from the control subjects. Among patients with ICA stenosis of 75% or more, end diastolic flow velocities were correlated to the diastolic stump pressures. CONCLUSION: Diastolic flow velocities within severe internal carotid artery stenosis are dependent on the level of the collateral perfusion pressure distally to the stenosis (ie, high values indicate a low internal carotid artery stump pressure), which seems to be a risk factor for early postoperative strokes. PMID- 10709061 TI - Validation of flow convergence region method in the assessment of carotid artery stenoses during color-flow duplex studies. AB - PURPOSE: The flow convergence region (FCR) method (also known as the proximal isovelocity surface area method) is currently used in echocardiography to evaluate the flow through cardiac valves and septal defects. The FCR method is based on the characteristic alterations in flow dynamics that occur proximal to a stenotic orifice. Blood converges uniformly and radially towards an orifice that is small relative to the section of the vessel and forms concentric isovelocity hemispheric shells where velocity progressively increases and flow remains laminar. The purpose of the article is to validate the use of this principle in the detection and assessment of carotid stenoses in the course of color-flow duplex studies. METHODS: In this prospective study, 80 patients affected by unilateral or bilateral carotid artery stenoses were evaluated for the presence of the FCR from February 1997 to March 1999. The results were compared with digital subtraction angiography. RESULTS: Color-flow duplex diagnosis of carotid artery stenoses of 70% or more was confirmed in 100% of the carotid artery stenoses (40/40 patients) with angiography. The FCR was detected in 72.2% (13/18) of carotid arteries affected by stenoses greater than 80%, in 54.4% (12/22) of carotid arteries affected by stenoses 70% to 80%, and in 13.6% (6/44) of carotid arteries affected by stenoses 50% to 69% (P <.001). In 5% of cases (2/40 of stenoses) the FCR was the only detectable sign of carotid stenosis. CONCLUSION: Our data suggest that a routine search for FCR in the course of color-flow duplex study of carotid arteries may further improve the reliability of this examination in the detection of carotid artery stenoses, particularly in the presence of heavily calcified lesions. PMID- 10709062 TI - The abdominal aortic aneurysm sac after endoluminal exclusion: a medium-term morphologic follow-up based on volumetric technology. AB - PURPOSE: The purpose of this study was to evaluate the role of three-dimensional spiral computed tomographic angiography (SCTA) for the assessment of the feasibility and results of endoluminal repair of infrarenal abdominal aortic aneurysm. METHODS: Laboratory studies: Phantom glass aneurysms, filled with contrast, underwent SCTA. The correlation between SCTA and laboratory measurements of linear dimensions and volumes was highly accurate (r(2) = 1.0). CLINICAL STUDIES: From the first 7 patients that were suitable for endoluminal repair, the correlation between SCTA and angiocatheter measurements was 0.85 to 0.99 (P <.04), but there was poor agreement between individual values. As determined from the measurements by 2 experienced investigators, intraobserver and interobserver errors for volume calculation in 12 randomly chosen scans from a total of 120 scans were 5.7 and 4.4 mL, respectively (range of volumes, 100-403 mL). The conditions of 53 patients were judged suitable for endoluminal repair of which 30 patients reached 1 year or more follow-up. The median aneurysm neck length and diameter were 24.5 mm (range, 11.5-60.8 mm) and 23.4 mm (18.3-31.5 mm), respectively. The fate of the sac after endografting by two techniques (pre expanded polytetrafluoroethylene [PTFE] fixed with Palmaz stents and endografts) was defined with three-dimensional SCTA. RESULTS: The sac volume after endografting by pre-expanded PTFE (n = 12 patients) showed a significant median increase (P =.02) from 129 mL before surgery to 141 mL at 5 days after the operation with no change at 6 (139 mL), 12 (137 mL), and 18 (159 mL) months later. With the endografts (n = 18), there was an initial increase in median volume at 5 days (179-194 mL; P =.02) and then a significant shrinkage at 6 (148 mL; P =.012) and 12 (94.9 mL; P =.02) months. CONCLUSION: Three-dimensional SCTA has been validated and is both precise and reliable. Interobserver and intraobserver errors are within acceptable ranges. Angiocatheter measurements are less accurate and may give misleading information when used for patient selection and endograft construction. The sac volume increased after endografting and later shrank in patients who were treated with endografts, but not in those patients treated with pre-expanded PTFE. We propose that three-dimensional SCTA should be regarded as the gold standard for linear and volumetric measurement for infrarenal abdominal aortic aneurysm. PMID- 10709063 TI - Thrombus within an aortic aneurysm does not reduce pressure on the aneurysmal wall. AB - PURPOSE: The role of thrombus within an aneurysm in relation to the risk of rupture is controversial. In literature, reports describing reduction and increase of rupture risk can be found. In the era of endovascular treatment of abdominal aortic aneurysms, a possible reduction of risk of rupture by the presence of thrombus within the aneurysmal sac can be important in relation to the location of an endoleak to the aneurysmal wall and in relation to the effect of the thrombosis of the endoleak, either spontaneously or by intervention. METHODS: In nine patients who underwent operation for an infrarenal aortic aneurysm by open procedure at the level of the thickest thrombus lining, the pressure within the aneurysmal thrombus (just inside the aneurysmal wall) was measured and compared with the systemic pressure. RESULTS: Pressure within systemic circulation and aneurysmal thrombus correlated well for the mean pressure (r = 0.90; P <.001) and for pulse pressure (r = 0.74; P <.01) Also, there was agreement between the levels of the mean pressure. Conduction of mean and pulse pressure to the aneurysmal wall was not related to the thickness of the thrombus at the level of the pressure measurement (r = 0.18 and r = 0.08, respectively). CONCLUSION: We conclude that thrombus within the aneurysm does not reduce both the mean and the pulse pressure near the aneurysmal wall and thus will not reduce the risk of rupture of the aneurysm. PMID- 10709064 TI - Ultrasonic measurement of abdominal aortic aneurysm wall compliance: a reproducibility study. AB - PURPOSE: The purpose of this study was to examine the intraobserver and interobserver error associated with ultrasonic echo-tracking compliance measurement in patients with abdominal aortic aneurysm. METHODS: Two observers independently measured brachial blood pressure by sphygmomanometer and maximum aortic diameter, pressure strain elastic modulus (Ep) and stiffness using an ultrasonic echo-tracker. The observer was blind to several variables: pulse pressure, diameter change, Ep, and stiffness. In study 1, observer A measured compliance in 13 patients at 30 to 60 minutes apart. In study 2, observers A and B each measured compliance on 23 patients at two visits, 2 weeks apart. RESULTS: There were no significant differences within observer A's compliance measurements. The coefficients of variation of method error (CV(ME)) for directly measured variables were systolic blood pressure, 7.3%; diastolic blood pressure, 5.4%; and maximum aortic diameter, 2.6%. CV(ME) values for derived variables were Ep, 21.2%, and stiffness, 17.6%. No differences were found between observers A and B and visits 1 and 2. CV(ME) values were 7.9% or less for directly measured variables and 32.7% or less for derived variables. These CV(ME) values were greatly reduced when the calculation was made with the use of log transformed data. CONCLUSION: The high CV(ME) value for derived variables is largely due to their wide variation within this population. This technique can measure abdominal aortic aneurysm diameter and compliance with an acceptable level of intraobserver and interobserver error. PMID- 10709065 TI - Elevated levels of soluble tumor necrosis factor receptors are associated with increased mortality rates in patients who undergo operation for ruptured abdominal aortic aneurysm. AB - PURPOSE: Elevated levels of soluble tumor necrosis factor receptors (sTNF-Rs) are associated with multiple organ failure and increased mortality rates in critically ill patients. Paradoxically, experimental data suggest exogenous sTNF Rs may improve outcome in patients who undergo elective abdominal aortic aneurysm (AAA) repair. This study examines, for the first time, changes in sTNF-R levels during repair of ruptured and nonruptured AAA. METHODS: Sixteen patients who underwent surgical procedures for ruptured AAA and 10 patients who underwent surgical procedures for nonruptured AAA were studied. Levels of sTNF-Rs p55 and p75 were measured before the operation and immediately before and 5 minutes, 6 hours, and 24 hours after aortic clamp release. RESULTS: When compared with nonruptured AAA, levels of sTNF-R p55 were significantly higher in ruptured AAA 5 minutes (P <.02) and 24 hours after aortic clamp release (P <.05). Levels of sTNF R p75 were significantly higher in ruptured AAA before (P <.05), during (P <.001), and after the surgical procedure (P <.01). Six hours after aortic clamp release, sTNF-R p75 levels were significantly higher in nonsurvivors of ruptured AAA when compared with survivors (P <.05) and patients who underwent surgical procedures for nonruptured AAA (P <.01). CONCLUSION: Ruptured AAA repair is associated with increased sTNF-R expression. Furthermore, elevated levels of sTNF R p75 are associated with increased postoperative mortality rates. PMID- 10709066 TI - Endotoxemia during supraceliac aortic crossclamping is associated with suppression of the monocyte CD14 mechanism: possible role of transforming growth factor-beta1. AB - PURPOSE: Monocyte CD14 and its soluble form (sCD14) mediate the proinflammatory response to endotoxemia. The aim of this study was to measure the changes to these factors after major aortic surgery and the possible inhibitory role of transforming growth factor-beta(1) (TGF-beta(1)) during these procedures. METHODS: Twenty-four patients with supraceliac aortic crossclamping during thoracoabdominal aortic aneurysm (TAAA) repair and 12 patients with infrarenal aortic crossclamping as part of infrarenal aneurysm repair (AAA) were studied. Blood was collected at incision, aortic clamping, and reperfusion and at 1, 8, and 24 hours after reperfusion. Samples were assayed for endotoxin, peripheral blood monocyte CD14 expression, sCD14, tumor necrosis factor-alpha, and TGF beta(1). RESULTS: Although there was significant endotoxemia on reperfusion in both groups of patients, peak plasma endotoxin levels were significantly higher in patients with TAAA (P =.001). Monocyte CD14 and plasma sCD14 were significantly decreased in patients with TAAA at reperfusion and 1 hour after reperfusion (P <.01, both points). In patients with AAA, a significant upregulation of CD14 was observed at 24 hours after reperfusion (P <.01), but no significant changes in sCD14 were observed. TNF-alpha showed no significant changes during the study period in both groups. In patients with TAAA, TGF beta(1) showed significant elevation at all time points (P <.01); whereas in patients with AAA, TGF-beta(1) showed no significant changes. CONCLUSION: Splanchnic ischemia reperfusion in patients who undergo supraceliac aortic clamping is associated with peripheral blood monocyte CD14 suppression and significant elevation of TGF-beta(1). TGF-beta(1) may play an important role in modulating the immune response to endotoxemia during major aortic aneurysm surgery. PMID- 10709067 TI - Vascular complications of osteochondromas. AB - PURPOSE: Osteochondromas are the most common benign tumor of the bone. They are sometimes responsible for vascular complications involving either veins or arteries, principally around the knee. METHODS: We report six cases of such complications. An extensive review of literature through a computerized research was performed. RESULTS: We found 97 cases that were previously reported in the English literature giving sufficient details and providing data on 103 cases for analysis. CONCLUSION: Surgical treatment of vascular complications of osteochondromas is recommended as an urgent procedure to avoid irreversible damages, such as arterial occlusion, embolism, or phlebitis. Prophylactic resection of osteochondromas in the vicinity of a vessel must be performed. PMID- 10709068 TI - An in vitro comparison of the hemodynamics of two inferior vena cava filters. AB - PURPOSE: The effectiveness of an inferior vena cava (IVC) filter in preventing pulmonary embolism while preserving caval flow is significantly affected by its hemodynamic characteristics. Flow fields surrounding two types of IVC filters were compared to assess how the design of a filter may influence performance. METHODS: The 12F Titanium Greenfield and VenaTech LGM inferior vena cava filters were studied in vitro with a noninvasive flow visualization technique, the photochromic flow visualization and measurement technique. Axial velocity profiles and wall shear stress distributions were measured. These results were compared with analytical data corresponding to the flow field in the absence of a filter to determine the relative extent of the flow disturbances. RESULTS: The reductions in near-wall axial velocity and wall shear stress caused by the VenaTech filter were more extensive and severe than those caused by the Greenfield filter. These changes were the consequence of differences in the geometry and dimensions of the struts of the two filters. The measurements showed the flow fields to be laminar, with no evidence of turbulence in both cases. CONCLUSION: Two factors that have been linked to thrombogenesis, near-wall velocity and wall-shear stress, were significantly affected by the larger frontal profile area of the VenaTech filter. Although a larger area may increase clot trapping efficiency, as shown by previous studies, the reduced near-wall velocities and wall shear stresses may increase the potential for thrombogenesis and, thus, caval occlusion. In contrast to other in vitro flow visualization studies, no turbulence was observed with either filter. PMID- 10709069 TI - Embolic risk of the different stages of carotid bifurcation balloon angioplasty: an experimental study. AB - PURPOSE: Embolic events during carotid angioplasty are a challenging problem. This experimental study was undertaken to determine the embolic risk after each stage of carotid angioplasty procedure. METHODS: Five ex vivo carotid artery balloon angioplasties were performed on fresh carotid specimens. The carotid specimens were obtained from five patients who underwent an internal carotid artery bypass for stenosis >75%. Before the endovascular maneuvers and after each stage of the procedures, the specimens were flushed with 20 mL of saline solution. Small particulate emboli (diameter, <60 microm) were searched in all the effluents according to the Coulter technique. After this procedure, each effluent was also submitted to scanning electron microscopy. RESULTS: When the stenosis was crossed with the guidewire or the balloon catheter, the number and the mean diameter of embolic particles did not change with three plaques (CP1, CP2, and CP3) and were increased with two plaques (CP4 and CP5). The maximal size of particles was 220 microm (CP5). After balloon angioplasty, the number and the mean diameter of particles increased with CP1, CP2, and CP3. With CP4 and CP5, the number of particles decreased, but their size increased. The maximal size of particles was 1100 microm (CP4). CONCLUSION: Carotid balloon angioplasty generates embolic particles after each stage of the procedure. Techniques of prevention should then be effective from the initial step of the angioplasty procedure, and the selection of patients for carotid angioplasty remains crucial. PMID- 10709070 TI - Adenoviral-mediated gene transfer of ICP47 inhibits major histocompatibility complex class I expression on vascular cells in vitro. AB - PURPOSE: Many viruses have evolved mechanisms to evade detection by the host immune system. The herpes simplex gene ICP47 encodes a protein that binds to the host antigen-processing transporter, inhibiting the formation of major histocompatibility complex class I (MHC-I) antigens in infected cells. MHC-I antigen expression is also important in acute allograft rejection. This study was designed to quantitate the effect of adenoviral-mediated gene transfer of ICP47 on MHC-I cell surface expression of human vascular cells. We hypothesized that the transduction of vascular cells with a replication-incompetent adenoviral vector that was expressing ICP47 (AdICP47) would inhibit constitutive and inducible MHC-I expression and thereby reduce the rate of cytolysis of ICP47 transduced vascular cells by sensitized cytotoxic T lymphocytes (CTL). METHODS: A replication-incompetent adenoviral vector, AdICP47, was created to express ICP47 driven by the cytomegalovirus immediate early promoter. Cultured human vascular endothelial and smooth muscle cells and human dermal fibroblasts were transduced with either AdICP47 or the control empty vector AddlE1. Cell surface constitutive and gamma-interferon-induced MHC-I expression were quantitated by flow cytometry. A standard 4-hour chromium release cytotoxicity assay was used to determine the percent cytolysis of transduced and nontransduced endothelial cells by sensitized CTL. Finally, to quantitate the specificity of the effect of ICP47 on MHC-I expression, adhesion molecule expression was quantitated in both transduced and nontransduced cells. RESULTS: Constitutive MHC-I expression in AdICP47-transduced endothelial cells was inhibited by a mean of 84% +/- 5% (SEM) in five experiments. After 48 hours of exposure to gamma-interferon, AdICP47-transduced cells exhibited a mean of 66% +/- 8% lower MHC-I expression than nontransduced cells. Similar inhibition in MHC-I expression was achieved in AdICP47-transduced vascular smooth muscle cells and dermal fibroblasts. Percent cytolysis of AdICP47 transduced endothelial cells by CTL was reduced by 72%. Finally, the specificity of the effect of transduction of ICP47 on vascular cell MHC-I expression was confirmed by a lack of significant change in either constitutive or tumor necrosis factor-induced vascular cell adhesion molecule/intercellular adhesion molecule expression. CONCLUSION: Transduction of vascular cells with AdICP47 strongly inhibits both constitutive and inducible MHC-I expression in human vascular cells. AdICP47-transduced cells exhibited a substantial reduction in cytolysis by CTL. Thus AdICP47 transduction holds promise as a technique to characterize the role of MHC-I expression in acute vascular allograft rejection in vivo and as a potential therapeutic intervention. PMID- 10709071 TI - Vascular smooth muscle cell apoptosis in aneurysmal, occlusive, and normal human aortas. AB - PURPOSE: Apoptosis is a physiologic mechanism of cell death that regulates mass and architecture in many tissues. Apoptosis has been described as a feature in human vascular atherosclerosis and large vessel structural integrity. We examined the extent of vascular smooth muscle cell (VSMC) apoptosis in aneurysmal, occlusive, and normal human aortic tissue. METHODS: Tissue samples of aneurysmal, occlusive, and normal human infrarenal aorta were evaluated. DNA fragmentation detection methods, immunohistochemistry, and DNA electrophoresis determined VSMC density, VSMC apoptosis, and apoptosis markers. Apoptotic cells and VSMC nuclei were counted with the use of computer-generated image analysis. Aortic subtypes were compared statistically by analysis of variance. RESULTS: Seventeen aneurysmal, ten occlusive, and five normal human aortas were evaluated. By alpha(1)-actin immunostaining, VSMC density was least in aneurysmal aortas (271.8 +/- 13.5 cells/high-power field [HPF]) compared with occlusive aorta (278.2 +/- 39.4 cells/HPF) and normal aortas (291.0 +/- 25.4 cells/HPF; P = not significant). Presence of apoptotic VSMCs was demonstrated by terminal deoxynucleotidyl transferase fragment end labeling and propidium iodide nuclear staining. VSMC apoptosis was greatest within aneurysmal aortas with 11.7 +/- 1.5 cells/HPF compared with occlusive aortas with 3.3 +/- 0. 8 cells/HPF (P <.05) and normal aortas with 3.75 +/- 4.6 cells/HPF (P <.05). Significant differences in apoptosis markers, p53 or bcl-2, could not be demonstrated by immunohistochemistry or DNA electrophoresis in aortic subtypes. CONCLUSION: Apoptosis of VSMCs is increased and VSMC density is decreased within the medial layer of aneurysmal aortic tissue. Structural degeneration of aortic tissue at the cellular level contributes to aneurysmal formation. PMID- 10709072 TI - The effect of growth factors, cytokines, and extracellular matrix proteins on fibronectin production in human vascular smooth muscle cells. AB - PURPOSE: Although 60% to 80% of the mature intimal hyperplastic plaque is composed of extracellular matrix (ECM) proteins, little is known about the factors that stimulate smooth muscle cells (SMCs) to produce these proteins. A major component of the ECM protein is fibronectin. Thus we studied fibronectin production and its response to various growth factors, cytokines, and other ECM proteins that are released at the time of vascular injury. METHODS: Quiescent cultured human SMCs were stimulated for varying intervals with increasing concentrations of agonist. Fibronectin in the cell medium was assayed by immunoblotting with a fibronectin-specific antibody. RESULTS: After 72 hours of stimulation, transforming growth factor-beta (10 ng/mL) had the most profound effect on fibronectin production (9.6- +/- 2.1-fold; P <.05), followed by epidermal growth factor (100 ng/mL; 5.0- +/- 0.1-fold; P <.05, for both). Surprisingly, the platelet-derived growth factors (AA, AB, and BB) and fibroblast growth factor did not stimulate fibronectin production. Among the matrix proteins studied, only collagen type I (20 microg/mL) stimulated fibronectin production (1.9- +/- 0.1-fold; P <.05), whereas collagen type IV and laminin had no effect. The contractile protein angiotensin II (100 ng/mL) was a weak stimulant of fibronectin (1.6- +/- 0.2-fold; P <.05). Time course studies of fibronectin production up to 72 hours revealed kinetics that varied with each agonist. Transforming growth factor-beta stimulated significant early production of fibronectin, whereas fibronectin production in response to epidermal growth factor and collagen type I was initially modest but increased with time. The effect of angiotensin II did not become evident until 72 hours. CONCLUSION: Cytokines, growth factors, and matrix proteins have varying quantitative effects on ECM protein production by human vascular SMCs. Knowledge of the factors that influence ECM protein production may allow for the design of specific inhibitors that can prevent intimal hyperplasia. PMID- 10709073 TI - Abdominal aortic aneurysm after pulmonary transplantation: a case report. AB - We present the case of a 39-year-old man who underwent repair of a symptomatic 5 cm abdominal aortic aneurysm. This patient had received a bilateral lung transplant for cystic fibrosis 10 years before this event. He was receiving cyclosporine, prednisone, and azathioprine as immunosuppression therapy. To our knowledge, this is the first reported abdominal aortic aneurysm after lung transplantation, and we note that our patient had a rapidly enlarging aneurysm, as seen in recipients of heart transplants. We postulate that immunosuppression may be related to the development and/or rapid growth of abdominal aortic aneurysms. PMID- 10709074 TI - Dorsalis pedis artery aneurysm: case report and literature review. AB - The presentation, diagnostic evaluation, and treatment of a patient with a dorsalis pedis artery aneurysm is discussed, and the literature is reviewed on this rare entity. PMID- 10709075 TI - Reversal of twice-delayed neurologic deficits with cerebrospinal fluid drainage after thoracoabdominal aneurysm repair: a case report and plea for a national database collection. AB - Delayed neurologic deficits are an uncommon yet devastating complication of thoracoabdominal aortic aneurysm repair. The mechanisms involved in the development of delayed spinal cord ischemia remain ill defined. We report a case of complete reversal of delayed neurologic deficits with postoperative cerebrospinal fluid (CSF) drainage. After a thoracoabdominal aneurysm extent I repair, the patient experienced delayed paraplegia at 6 hours and again at 34 hours after the operation, with elevated CSF pressure (>10 mm Hg) on both occasions. Prompt CSF decompression completely reversed the neurologic deficits within hours after onset. The findings in this case further support the role of CSF drainage in spinal cord protection for patients who undergo thoracoabdominal aneurysm repair and make a plea for a national database collection. PMID- 10709076 TI - Late abdominal aortic aneurysm rupture after AneuRx repair: a report of three cases. AB - Rupture due to device failure and/or endoleak is the most feared complication of endoluminal grafting for exclusion of abdominal aortic aneurysm. We present three previously unreported cases of abdominal aortic aneurysm rupture 23 months after AneuRx "repair" and describe the mechanisms of failure and discuss instructive technical aspects of their management. PMID- 10709077 TI - Guidewire entrapment during deployment of the over-the-guidewire stainless steel Greenfield filter: a device design-related complication. AB - The Greenfield filter (Medi-tech/Boston Scientific, Watertown, Mass) is widely used for the prevention of pulmonary embolism. The latest version is a stainless steel over-the-wire filter. The purpose of the guidewire is to facilitate placement of the device down through the atrium or through tortuous vessels and to prevent tilting of the released filter. A case of entrapment of the guidewire in the filter after deployment through the right internal jugular approach and its recovery by reentering it into the sheath is presented. To prevent this complication, the guidewire should be removed completely before releasing this type of filter. The potential risk of tilting the filter does not outweigh the risk of guidewire entrapment. PMID- 10709078 TI - Regarding: "Expression of myosin heavy chain isoforms in skeletal muscle of patients with peripheral arterial occlusive disease". PMID- 10709079 TI - Treatment options for venous thrombosis. PMID- 10709080 TI - Regarding "Adventitial cystic disease: a unifying hypothesis". PMID- 10709081 TI - Regarding "Randomized study of carotid angioplasty and stenting versus carotid endarterectomy: a stopped trial". PMID- 10709083 TI - Reply PMID- 10709082 TI - Reply PMID- 10709084 TI - Regarding "In situ replacement of the aorta in a contaminated field with the infrarenal inferior vena cava". PMID- 10709085 TI - Reply PMID- 10709086 TI - Regarding "Early history of aortic surgery". PMID- 10709087 TI - Reply PMID- 10709088 TI - Regarding: "Anastomotic tissue response associated with expanded polytetrafluoroethylene access grafts constructed by using nonpenetrating clips". PMID- 10709089 TI - L-myc polymorphism in cancer patients, healthy blood donors and elderly, tumor free individuals in Russia. AB - L-myc polymorphism was investigated in 95 breast cancer (BC), 63 colorectal cancer (CC) and 58 lung cancer (LC) patients, as well as in 122 healthy, middle aged blood donors (HBDs) and 184 elderly, tumor-free individuals. The occurrence of the S allele in the BC cohort (57%) was significantly higher than that in middle-aged, healthy females (41%) and elderly, non-affected women (47%), implying involvement of the L-myc genotype in BC susceptibility (age-adjusted OR = 1.74, 95% CI 1.11-2.73, p = 0.016). L-myc allele distribution in CC and LC was similar to that in controls. Contrary to earlier reports, L:S allele frequencies ratio in elderly blood donors (EBDs) did not significantly differ from that in HBDs (0.49:0. 51 and 0.54:0.46, respectively). However, the S allele had a tendency to be over-represented among elderly compared with middle-aged smokers (55% vs. 44%; OR = 1.57, 95% CI 0.98-2.50, p = 0. 059), which implies that it may be linked with tolerance to smoking effects. PMID- 10709090 TI - Tumor targeting with radiolabeled antibodies in a human carcinoembryonic antigen transgenic mouse model. AB - Mice transgenic for the carcinoembryonic (CEA) gene were used to study the biodistribution and tumor targeting of a radioiodinated monoclonal antibody (MAb), T84.66. The specificity of antibody uptake in tumors was assessed in mice bearing a CEA-transfected syngeneic tumor as well as the antigen-negative parental tumor. With high CEA-expressing tumors, the percent injected dose per gram (%ID/g) approached 30% at 48 hr. Tumor uptake in antigen-positive tumors was 5-8-fold higher than that observed in the antigen-negative parental tumors. Only antigen-positive tumors were visualized by immunoscintigraphy. The tumor targeting obtained in athymic nude mice bearing human tumor xenografts was similar to that observed with CEA-expressing murine tumors implanted in either athymic nude or transgenic mice. The degree of localization of CEA-transfected murine tumors was related with the level of antigen expression. Circulating antigen-radio-antibody complexes were not detected while blood clearance of radio antibody was similar between transgenic and non-transgenic mice. With the exception of the large bowel, the distribution of radioiodinated MAb in normal tissues was similar in both CEA transgenic and non-transgenic mice. Increased localization of intact antibody was observed in the large bowel from transgenic mice, suggesting specific targeting to antigen-positive normal tissues. These results suggest that the CEA transgenic mouse model will be useful in the development of antibodies for radio-immunodetection and treatment of carcinomas expressing CEA. PMID- 10709091 TI - Alteration of beta-catenin expression in esophageal squamous-cell carcinoma. AB - beta-catenin regulates cadherin-mediated cell-cell adhesion and also functions as a signaling molecule. In this study, we examined the expression pattern of E cadherin, alpha-catenin and beta-catenin in 22 cases of esophageal squamous-cell carcinoma by Western-blot analysis. Expression of E-cadherin, alpha-catenin and beta-catenin was lower in carcinomas than in normal esophageal mucosa in 4 cases (18.2%) for E-cadherin, 6 cases (27.3%) for alpha-catenin and 9 cases (40.9%) for beta-catenin. Expression of beta-catenin was not always correlated with that of E cadherin. Over-expression of beta-catenin was observed in 3 cases (13.6%). Of 3 cases that presented with over-expression of beta-catenin, 2 showed cytoplasmic staining by immunohistochemistry. Nuclear localization of beta-catenin was observed in one case that had higher beta-catenin level in tumor tissue (1.4-fold higher than normal mucosa). The genomic DNA sequences of the beta-catenin and the APC gene were analyzed. No mutation of the beta-catenin gene was observed in any cases. Silent mutation of the APC gene was found in all the cases that showed over-expression or nuclear localization of the beta-catenin protein. These results indicate that alterations of the cadherin-catenin complex may play an important role in a sub-set of esophageal carcinogenesis. Furthermore, it is suggested that beta-catenin over-expression is not caused by genetic alteration of either the beta-catenin or the APC gene. PMID- 10709092 TI - Human retrovirus 5 sequences in peripheral blood cells of patients with B-cell non-Hodgkin's lymphoma. AB - A recently described sequence from a probable 5th human exogenous retrovirus, HRV 5, is related to type A, B and D retroviruses. It was initially detected in a salivary gland biopsy from a patient with Sjogren's syndrome, but it is not consistently associated with this disease. We searched for the HRV-5 sequence in DNA extracted from whole blood of 300 blood donors, 81 patients with hematological malignancy and 21 patients with neurological disease using PCR. While samples from none of the blood donors and the neurological patients became positive, 3 of the 81 patients with hematological malignancy were HRV-5 DNA positive. All 3 had B-cell non-Hodgkin's lymphoma of low grade. The difference in frequency between NHL and controls is statistically significant. HRV-5 DNA was found in DNA from whole blood and in plastic-adherent cells but not in tumor cell DNA. Thus, monocytes/macrophages may be preferred targets for HRV-5. Our result, together with a previous finding of HRV-5 DNA in 2 NHL cases, is compatible with an association between HRV-5 and NHL, whether causal or not. PMID- 10709093 TI - Characterization and modulation of a prolactin receptor mRNA isoform in normal and tumoral human breast tissues. AB - The role of prolactin (PRL) and its specific receptor (R-PRL) in human breast tumorigenesis remains unclear. We have investigated here the presence of extracellular-deleted hPRL-R isoforms in normal human breast, fibrocystic disease, primary breast carcinoma (ductal carcinoma, ductulo-lobular and lobular) and breast cancer cell lines (T47-D and MCF-7). RT-PCR and Southern blot analysis demonstrated the expression of full-length hPRL-R transcript in all samples tested. We also detected a hPRL-R transcript generated by alternative exon 6 splicing. This isoform has a 170 bp deletion in its extracellular sub-domain that induces a frameshift. Thus, the predicted amino-acid sequence should encode a putative soluble protein with the N-terminal sub-domain of the hPRL-R and 10 additional carboxy-terminal residues. This isoform should not bind PRL as previously demonstrated by other experiments. Moreover, the ratio of full-length to deleted form of hPRL-R transcripts differs from normal to tumoral breast tissue. This ratio is higher in tumoral mammary gland than in normal tissue. Our data suggest that the alternative splicing of the hPRL-R gene towards the deleted transcript may be a mechanism to down- or up-regulate the expression of the native transcript of hPRL-R in accordance to the physiological or pathological state of the mammary gland. PMID- 10709094 TI - Role of metallothionein in cisplatin sensitivity of germ-cell tumours. AB - Cisplatin (CDDP) is an extremely active drug in the treatment of germ-cell tumours. Earlier, we found an unexpected inverse correlation between the total amount of sulfhydryl groups and CDDP sensitivity in a panel of 3 human germ-cell tumour and 3 colon-carcinoma cell lines. Major components of the sulfhydryl groups are glutathione and metallothionein (MT). We further investigated a possible role of MT in the CDDP sensitivity of germ-cell tumours. MT levels and functionality of the germ-cell-tumour and colon-carcinoma cell lines were found to be inversely correlated with CDDP sensitivity. No difference in sub-cellular localization of MT could be observed among the types of cell lines. In agreement with the in vitro data, immunohistochemical detection of MT was high in 11/14 primary human germ-cell tumours and low in 7/7 human colon carcinomas. MT-protein expression in primary germ-cell tumours did not discriminate between responding and non-responding patients. As compared with the primary tumours, MT-protein expression decreased in 5/7 post-chemotherapy residual vital tumours or remained undetectable (2/7). MT-protein expression in the germ-cell tumours was not related to total p53-protein expression. In summary, over-expression of MT was found in germ-cell tumours, both in cell lines and in human tumours. Although MT protein over-expression seems to be associated with the CDDP sensitivity of germ cell tumours, MT-protein expression in primary germ-cell tumours did not differ between responding and non-responding patients and therefore cannot be used to predict response to chemotherapy. PMID- 10709095 TI - Changes of the p16 gene but not the p53 gene in human chondrosarcoma tissues. AB - The role of two important tumour suppressor genes, p16 and p53, was evaluated in cartilaginous tumour tissues. Genomic DNA from 22 chondrosarcomas, 5 benign chondroid tumours, 1 sample of reactive proliferative cartilage and 2 samples of normal cartilage were analysed using polymerase chain reaction, single strand conformational polymorphism, DNA sequencing and methylation-specific polymerase chain reaction. The p16 gene was found to be partly methylated in 5 high-grade chondrosarcomas and homozygously deleted in 1 chondrosarcoma. Moreover, a polymorphism was detected in 3 malignant tumours, but not in benign tumours or normal cartilage. Analysis of the p53 gene revealed an unchanged structure in all samples. These findings show a role for p16, but not p53, in chondrosarcoma. PMID- 10709096 TI - Cessation of alcohol drinking and risk of cancer of the oral cavity and pharynx. AB - A strong, dose-dependent association exists between alcohol consumption and risk of cancer of the oral cavity and pharynx. The impact on risk of temporal aspects of drinking habits has been inadequately evaluated. Our case-control study included 754 individuals with incident cancer of the oral cavity and pharynx (median age 57) and 1,775 controls (median age 57) in the hospital for acute, non neoplastic diseases who were interviewed in 2 Italian areas and in the Swiss Canton of Vaud between 1992 and 1997. The questionnaire included lifetime drinking and smoking habits. No influence of age at starting or duration of alcohol drinking was found. Risk increased substantially with the increase of weekly alcoholic drinks [Odds Ratios (OR) for >/= 91 drinks/week vs. never drinkers = 11.6]. Risk in former compared with current drinkers was 1.9-fold elevated. However, among individuals who had also stopped smoking, former drinkers showed lower ORs than current drinkers. The persistence of risk elevation several years after drinking cessation suggests that the role of alcohol is complex and it probably affects more than one stage of oral carcinogenesis. It remains to be clarified which impact prevention-driven drinking cessation may have on the excess of cancer of the oral cavity and pharynx due to elevated alcohol intake. PMID- 10709097 TI - Circulating p53 antibodies, p53 gene mutational profile and product accumulation in esophageal squamous-cell carcinoma in India. AB - Esophageal cancer (EC) in the Indian population presents in advanced stages with poor prognosis and warrants the identification of a non-invasive marker for early detection and better prognostic assessment. We have previously reported high prevalence of p53 protein accumulation in esophageal squamous-cell carcinomas (ESCCs). The present study was designed to determine (i) if esophageal cancer patients elicit a humoral immune response to intra-tumoral p53 protein accumulation and (ii) their relationship with p53 gene mutations. The goal was to compare the cellular events, p53 protein accumulation and gene mutations with the presence of serum anti-p53 antibodies (p53-Abs) and to assess the utility of serological p53-Ab analysis as a surrogate marker for p53 alterations in esophageal cancer. A high prevalence of circulating p53-Abs was observed in 36 of 60 (60%) ESCC patients. In a subset of 44 ESCCs, exons 5-9 of the p53 gene were examined for mutations by PCR and direct sequencing of PCR products. Mutational data have been correlated with p53-Abs and p53 protein accumulation in ESCCs. Circulating p53-Abs in ESCC patients were significantly associated with intra tumoral p53 protein accumulation (p=0.0005). A strong correlation observed between humoral immune response against p53 protein, missense gene mutations and protein accumulation warrants the application of serological p53-Abs as a non invasive surrogate marker in screening high-risk populations for early detection of malignancy. PMID- 10709098 TI - Germline BRCA1 and HMLH1 mutations in a family with male and female breast carcinoma. AB - Hereditary non-polyposis colorectal cancer (HNPCC) is an autosomal dominant disease, predisposing to the development of colorectal cancer and other tumor types such as endometrial, small bowel, stomach, ovary and urinary tract carcinoma, while most investigators find no association between HNPCC and cancer of the breast. We have identified hMLH1 mutations in 2 Amsterdam-criteria HNPCC families where both male and female gene carriers were affected with breast cancer. To investigate whether these breast cancers were caused by other genetic factors, we analyzed the 2 breast cancer susceptibility genes BRCA1 and BRCA2. In one family we did not find any mutation in the breast cancer genes, while in the other, a BRCA1 mutation segregated in the breast cancer cases. Hereditary breast cancer, and in particular BRCA1 tumors, display discrete histo-pathological and immunohistological characteristics. The tumor from a woman with both hMLH1 mutations and a BRCA1 mutation exhibited typical BRCA1 histology, e.g., grade 3 invasive ductal carcinoma with dense lymphocytic infiltration, and immunohistology, estrogen receptor (ER) negative, progesterone receptor (PgR) negative, strongly p53 positive, c-erbB-2 negative and highly Ki67 positive (>50% stained cells). The histology of the breast tumor from the man with both one hMLH1 mutation and a BRCA1 mutation was a grade 2 invasive ductal carcinoma without any special BRCA1 features. Immunohistology was also different. This might merely reflect a true difference in male breast tumor progression vs. female. We cannot exclude that the combined effect of BRCA1 and hMLH1 dysfunction has a bearing on tumor progression. PMID- 10709099 TI - Matrix-metalloproteinases and their inhibitors in plasma and tumor tissue of patients with renal cell carcinoma. AB - Matrix-metalloproteinases (MMPs) are associated with invasive and metastatic behavior of several human malignant tumors. We have determined MMP-2 and MMP-9 and tissue inhibitors of MMPs (TIMP-1 and TIMP-2) in blood plasma and in renal tissue samples of patients with renal cell carcinoma (RCC) from cancerous and non cancerous parts of the same kidney. In tumor tissue, MMP-9 and TIMP-1 were significantly higher than in normal counterparts. MMP-2 was not different between tumor tissue and normal counterparts. TIMP-2 values could not be measured. In plasma, MMP-9 concentrations were significantly higher in RCC patients than in healthy controls, MMP-2 and TIMP-2 concentrations were higher in healthy controls and TIMP-1 concentrations were not different. PMID- 10709100 TI - Epigenetic silencing of maspin gene expression in human breast cancers. AB - Maspin is a tumor suppressor whose expression is lost in many advanced breast cancers. Maspin has been shown to inhibit cell motility, invasion and metastasis; however, its precise role in normal mammary epithelium remains to be elucidated. Although expression of maspin mRNA is low or absent in most human breast cancer cells, the maspin gene is rarely re-arranged or deleted. We hypothesized that aberrant cytosine methylation and chromatin condensation of the maspin promoter participates in the silencing of maspin expression during neoplastic progression. To test this hypothesis, we compared cultured normal human mammary epithelial cells (HMECs) to 9 cultured human breast cancer cell lines. HMECs expressed maspin mRNA and displayed a completely non-methylated maspin gene promoter with an open chromatin structure. In contrast, 7 of 9 breast cancer cell lines had no detectable maspin expression and 6 of these 7 maspin-negative breast cancer cell lines also displayed an aberrant pattern of cytosine methylation of the maspin promoter. Interestingly, the maspin promoter was completely methylated in maspin negative normal peripheral blood lymphocytes. This indicates that the maspin promoter is not a functional CpG island and that cytosine methylation of this region may contribute to normal tissue-restricted gene expression. Chromatin accessibility studies with MCF-7 cells, which lack maspin expression and have a methylated maspin promoter, showed a closed chromatin structure compared with HMECs. Moreover, maspin gene expression could be re-activated in MCF-7 cells by treatment with 5-aza-2;-deoxycytidine, a DNA demethylating agent. Thus, aberrant cytosine methylation and heterochromatinization of the maspin promoter may silence maspin gene expression, thereby contributing to the progression of human mammary cancer. PMID- 10709101 TI - Association of inverted sinonasal papilloma with non-sinonasal head-and-neck carcinoma. AB - The nature and pathogenesis of inverted papilloma of the nose and paranasal sinuses are debated. Evidence suggesting a viral association is controversial, and epidemiological evidence has pointed to tobacco smoking as a potential etiologic factor. A retrospective regional cohort of 197 patients with sinonasal papilloma was compared with a cohort of 1583 patients with nasal polyps showing a similar distribution by age and sex. All instances of head-and-neck carcinoma diagnosed in both cohorts during a 38-year calendar period were culled from the regional cancer registry, the incidence rate ratio was computed (papilloma:polyp, on tumors detected at the time of or prior to the index diagnosis), and the clinical details were obtained. Nine instances of oral or laryngeal squamous-cell carcinoma, all in men, were identified in the papilloma cohort, and 7 labial, oral or laryngeal carcinomas (2 in women) in patients with polyps. In addition, 5% of the papillomas progressed to sinonasal carcinoma, including 2 cases among those with other primary head-and-neck carcinomas. The incidence-rate ratio for non-sinonasal head-and-neck carcinoma was 12.8 (95% CI, 3. 7 to 50; p < 0.0001). Among the papilloma patients with oral/laryngeal carcinoma, 8 smoked tobacco. Inverted sinonasal papilloma is associated with an increase in non-sinonasal head and-neck carcinoma, and tobacco may be a causative link. PMID- 10709102 TI - Antibodies against oncoproteins E6 and E7 of human papillomavirus types 16 and 18 in patients with head-and-neck squamous-cell carcinoma. AB - Human papillomaviruses (HPVs) have been recognized as an essential pathogenic factor in anogenital cancer. HPV DNA has also been found in a subgroup of head and-neck squamous-cell carcinomas (HNSCCs), and a causative role of the virus in the development of these tumors has been suggested by the concomitant inactivation of the tumor-suppressor protein pRb. Using 4 second-generation ELISAs, we found antibodies against at least 1 of the oncoproteins E6 and E7 of the high-risk HPV types 16 and 18 in 11 of 92 sera (12%) taken from HNSCC patients at or near diagnosis, in 1 of 52 sera (2%) taken from HNSCC patients >6 months after diagnosis and in 10 of 288 sera (3. 5%) taken from sex- and age matched healthy control individuals of the normal population. In 11 of the 12 seropositive HNSCC cases, antibodies were directed against HPV16 proteins. In patients, the HPV16 antibodies were mostly of high titer, and in 6 cases, antibodies against both HPV16 oncoproteins were present. Seven of the 8 HPV16 antibody-positive sera from the control group were of low titer, and none of the 10 antibody-positive sera reacted with both oncoproteins of the same HPV type. The HPV type of the antigens detected by the antibodies in HNSCC patients correlated well with that of the HPV DNA found in the tumor. Of 19 patients known to have HPV16 DNA-positive tumors, 7 (37%) also had HPV16 E6 and/or E7 antibodies. Our finding suggests that the antibodies were formed in an immune response against HPV E6 and E7 proteins expressed in the HNSCC and thus strongly supports the concept of a biologically active role of HPV in the development of a subgroup of HNSCC. PMID- 10709103 TI - Modulation of phenotype and induction of irregular vessels accompany high tumorigenic potential of clonal human glioma cells xenografted to nude-rat brain. AB - Three phenotypically different clonal human glioma cell lines were injected stereotactically into nude-rat brains, to determine their individual growth potential and to establish an in vivo system in which different therapeutic modalities could be tested. As assessed by serial sectioning, microscopic evaluation, and computer analysis, the mean approximate tumour volume after 3-7 weeks in vivo was 0.42 mm(3) for U-343 MG, 2.6 mm(3) for U-343 MGa Cl2:6, and 50.3 mm(3) for U-343 MGa 31L. When compared with the initial injected cell volume, only U-343 MGa 31L had increased in size, U-343 MGa Cl2:6 remained approximately the same but showed a certain proliferative potential, and U-343 MG regressed. Thus, only U-343 MGa 31L cells had high tumorigenic potential, invaded and replaced brain tissue in every direction, while U-343 MGa Cl2:6 cells grew in sheet-like tumour extensions along white-matter nerve-fibre tracts, in this respect mimicking foetal astrocytes. The tumorigenic potential of the U-343 MGa 31L cell clone was associated with a variable phenotype, as observed when the in vivo and in vitro characteristics were compared. The in vivo phenotype was characterized by the loss of GFAP immunoreactivity, the gain of heterogeneously distributed cellular tenascin, fibronectin, and laminin, but absence of extracellularly deposited material, and by the formation of irregular vessels. It appears that the intrinsic capacity of glioma cells to adapt to in vivo conditions is decisive for their tumorigenicity in the brain, rather than any single phenotypic property in itself. Moreover, the 2 glioma cell clones best suited for in vitro growth were no longer tumorigenic. PMID- 10709104 TI - Agonist peptide from a cytotoxic t-lymphocyte epitope of human carcinoembryonic antigen stimulates production of tc1-type cytokines and increases tyrosine phosphorylation more efficiently than cognate peptide. AB - The identification of an agonist peptide (YLSGADLNL, designated CAP1-6D) to an immunodominant cytotoxic T-lymphocyte (CTL) epitope (designated CAP1) of human carcinoembryonic antigen (CEA) has previously been reported. The agonist peptide harbors a single amino acid substitution at a non-MHC anchor residue and is proposed to exert its effects at the level of the T-cell receptor (TCR). The type and magnitude of cytokines produced by CAP1-reactive CTL upon stimulation with the agonist peptide, CAP1-6D, were compared to those obtained upon stimulation with the cognate CAP1 peptide. In addition, early events in the TCR signaling pathway were examined for differences in tyrosine phosphorylation. Upon stimulation with the agonist peptide CAP1-6D, several different CEA-specific CTL lines exhibited a marked shift in the peptide dose response, which resulted in as much as a 1,000-fold increase in the levels of GM-CSF and gamma-IFN produced as compared with the use of the CAP1 peptide. However, levels of IL-4 and IL-10, which are associated with anti-inflammatory effects, were very low or non existent. The cytokine profile of CAP1- and CAP1-6D-specific CTL is consistent with a Tc1-type CTL. Consistent with these findings, CEA-specific CTL showed increased tyrosine phosphorylation of TCR signaling proteins ZAP-70 and TCR zeta chains in response to both peptides. However, when CAP1-6D was compared with the wild-type peptide, the increase in ZAP-70 phosphorylation was greater than the increase in zeta phosphorylation. CTL generated with the CAP1-6D agonist were shown capable of lysis of human carcinoma cells expressing native CEA. The ability to upregulate the production of GM-CSF, gamma-IFN, TNFalpha and IL-2 with the agonist peptide, as compared with CAP1, may help in initiating or sustaining anti-tumor immune responses and thus potentially prove to be useful in the treatment of CEA-positive tumors. PMID- 10709105 TI - Immunomodulation with FK506 around the time of intravenous re-administration of an adenoviral vector facilitates gene transfer into primed rat liver. AB - Although adenoviruses are an attractive vehicle for gene transfer into tissues including various tumors, in vivo adenoviral administration elicits a neutralizing antibody response which eliminates or substantially reduces the efficacy of subsequent treatments. Transiently immunosuppressive strategies at the time of initial adenoviral exposure have shown to prevent the formation of neutralizing antibodies and permit the successful adenoviral readministration in animals. Initial treatment in humans may, however, correspond to adenoviral readministration into animals, because the exposure to wild-type adenoviruses is common in humans. In the present study, we infused Adex1CAlacZ adenoviruses carrying the lacZ gene into the tail vein of rats, and examined whether a transient treatment with the immunosuppressant FK506 around the time of i.v. readministration of adenoviruses could induce the re-expression of the lacZ gene in animals primed with adenoviruses. Although i.v. infusion of adenoviruses carrying the lacZ gene resulted in transiently high levels of transgene expression in rat liver, i.v. reinfusion of adenoviruses failed to induce detectable levels of transgene expression. Conversely, when animals were treated transiently with FK506 around the time of adenoviral reinfusion, development of neutralizing antibodies and antigen-specific T cell proliferation in response to adenoviral reinfusion were significantly suppressed, and re-expression of the transgene was achievable. PMID- 10709106 TI - Comparison of DNA repair protein expression and activities between human fibroblast cell lines with different radiosensitivities. AB - In order to investigate the molecular basis of variation in response to ionising radiation (IR) in radiotherapy patients, we have studied the expression of several genes involved in DNA double-strand break repair pathways in fibroblast cell lines. Ten lines were established from skin biopsies of cancer patients with different normal-tissue reactions to IR, and 3 from a control individual. For all 10 test cell lines, the cellular radiosensitivity was also known. Using Western blots we measured, in non-irradiated cells, the basal expression levels of ATM, Rad1 and Hus1, involved in the control of cellular DNA damage checkpoints, together with DNA-PKcs, Ku70, Ku80; XRCC4, ligaseIV and Rad51, involved in radiation- induced DSB repair. We also analysed the in vitro enzymatic activities, under non-irradiated conditions, of the DNA-PK and XRCC4/ligaseIV complexes. The levels of expression of the different proteins were similar in all the cell lines, but the activities of the DNA-PK and XRCC4/ligaseIV complexes showed some differences. These differences did not correlate with either the normal tissue response of the patient in vivo or with cellular radiation sensitivity in vitro. The activity differences of these enzyme complexes, therefore, do not account for the variation of responses seen between patients. PMID- 10709107 TI - Anti-beta4 integrin antibodies enhance migratory and invasive abilities of human colon adenocarcinoma cells and their MMP-2 expression. AB - Integrin-mediated adhesion of cells to extracellular matrix proteins has been shown to activate various intracellular signaling events. In the present study, we demonstrate that the addition of a monoclonal antibody raised against the beta4 integrin subunit in the culture medium of a clone derived from the colon adenocarcinoma cell line LoVo specifically results in stimulation of cell migration and invasion through reconstituted basement membrane matrices. Moreover, an increase in MMP-2 activity is observed. Conversely, monoclonal anti alpha6 and anti-beta1 have no effect on MMP-2 expression. The s. c. co-injection of adenocarcinoma cells with antibodies raised against the beta4 integrin subunit to immunosuppressed newborn rats gives rise to tumors displaying altered and disorganized peri-tumoral basement membranes compared with tumors obtained when cells are injected with adenocarcinoma cells alone. Higher metastatic capacity of cells results when they are co-injected with antibodies to the beta4 integrin subunit. Our results suggest that the beta4 subunit of alpha6beta4 integrin, a laminin receptor in colon adenocarcinoma, may be responsible for the specific signals which stimulate cell motility, expression of MMP-2 and tumor invasion. PMID- 10709108 TI - Human glucose-6-phosphate dehydrogenase (G6PD) gene transforms NIH 3T3 cells and induces tumors in nude mice. AB - The main physiological function of glucose-6-phosphate dehydrogenase (G6PD) is to produce NADPH and ribose 5-phosphate, which are essential for reductive biosynthesis and nucleic acid synthesis. In normal cells, G6PD expression is tightly controlled; however, in many tumors, regulation of its expression is altered, resulting in a significant increase in G6PD activity. To investigate the potential role of G6PD in tumorigenesis, we transfected NIH 3T3 cells with human G6PD cDNA. Cells overexpressing G6PD showed altered cell morphology and exhibited tumorigenic properties. In contrast to the control cells or cells transfected with mutated G6PD cDNA, G6PD-overexpressing cells were not contact inhibited and exhibited anchorage-independent growth. They divided more quickly and induced rapidly growing, large fibrosarcomas in nude mice. Moreover, the induced tumorigenic properties were positively correlated with the level of G6PD activity. Interestingly, treatment with buthionine SR-sulfoximine (BSO), a glutathione depletion agent, decreased the colony-forming efficiency of G6PD overexpressing cells in soft agar, which implicates that alteration of the redox balance may be involved in G6PD-induced tumorigenesis. A comparative analysis of the expression level of G6PD in a variety of human cancer cell lines was also performed. Northern- and Western-blot analyses revealed that G6PD was particularly overexpressed in human esophageal cancer cell lines. Our observations indicate that G6PD may act as a potential oncogene, whose overexpression plays a critical role in neoplastic transformation. PMID- 10709110 TI - The ubiquitin-activating enzyme E1-like protein in lung cancer cell lines. AB - The UBE1L gene isolated from the chromosome 3p21 region has an extremely reduced level of mRNA in lung cancer. Sequence analysis showed a 45% homology to the human ubiquitin-activating enzyme E1 at the amino acid level. To further characterize the protein product, we generated UBE1L protein-specific antibodies. Immunoblot analysis revealed a full-length gene product of approximately 112 kDa. Assessment of the level and distribution pattern of the UBE1L protein in normal and tumor tissue using the generated antibodies showed that the UBE1L protein was present in normal lung cells and non-lung cancer cell lines, but was undetectable in all 14 human lung cancer cell lines analyzed. This difference in expression of the UBE1L protein between normal lung tissue and lung tumor-derived cell lines suggests a possible involvement of an E1-like protein in the origin and/or progression of lung tumors. PMID- 10709109 TI - Molecular cloning and immunogenicity of renal cell carcinoma-associated antigen G250. AB - The molecular cloning of the cDNA and gene encoding the renal cell carcinoma (RCC)-associated protein G250 is described. This protein is one of the best markers for clear cell RCC: all clear-cell RCC express this protein, whereas no expression can be detected in normal kidney and most other normal tissue. Antibody studies have indicated that this molecule might serve as a therapeutic target. In view of the induction/up-regulation of G250 antigen in RCC, its restricted tissue expression and its possible role in therapy, we set out to molecularly define the G250 antigen, which we identified as a transmembrane protein identical to the tumor-associated antigen MN/CAIX. We determined, by FISH analysis, that the G250/MN/CAIX gene is located on chromosome 9p12-13. In view of the relative immunogenicity of RCC, we investigated whether the G250 antigen can be recognized by TIL derived from RCC patients. The initial characterization of 18 different TIL cultures suggests that anti-G250 reactivity is rare. PMID- 10709111 TI - Expression analysis of endogenous menin, the product of the multiple endocrine neoplasia type 1 gene, in cell lines and human tissues. AB - We have investigated the endogenous expression of menin, a protein encoded by the gene mutated in multiple endocrine neoplasia type 1 (MEN1). Western blot analysis showed strong expression of menin as a 68 kDa protein in all of 7 human and primate cell lines tested. In a panel of 12 fetal human tissue extracts, 68 kDa menin was readily detected in brain cortex, kidney, pituitary, testis and thymus and weakly detected in thyroid. Reproducible bands other than 68 kDa were observed in adrenal and heart, whereas menin was undetectable in liver, lung, pancreas and skin. Analysis of synchronized HeLa cells revealed no variation in the amount or size of menin throughout the cell cycle. Protein expression was compared between lymphoblastoid cell lines from healthy controls and MEN1 patients carrying nonsense mutations on 1 allele. No truncated protein was detected in either cytoplasmic or nuclear fractions in mutation-carrying cells. The expression level and cellular location of full-length menin did not differ between cell lines derived from MEN1 patients and healthy donors. This suggests that the wild-type allele has been up-regulated in mutation-carrying cells to compensate for the loss of 1 functional allele. PMID- 10709112 TI - Changes in phospholipase D isoform activity and expression in multidrug-resistant human cancer cells. AB - Multidrug resistance (MDR) is a major cause of failure of cancer chemotherapy and is often associated with elevated expression of drug transporters such as P glycoprotein (P-gp) in the cancer cells. MDR is, however, accompanied by additional biochemical changes including modifications of membrane composition and properties. We have shown that MDR is associated with a massive up-regulation of caveolin expression and an elevated surface density of caveolae. We report that phospholipase D (PLD), a constituent enzyme of caveolae and detergent insoluble glycolipid-rich membranes (DIGs), is up-regulated in human MDR cancer cells. Lysates of HT-29-MDR human colon adenocarcinoma cells, MCF-7 AdrR human breast adenocarcinoma cells and the corresponding parental drug-sensitive cells, were fractionated on discontinuous sucrose density gradients. PLD activity was found to be enriched in low density fractions that contain DIGs and caveolar membranes, and the activity in these fractions was 4- to 6-fold higher in the MDR cells compared with the parental drug- sensitive cells. Utilizing specific antibodies to PLD1 and PLD2, the distribution of PLD isoforms along the gradient was determined and the PLD localized in DIGs and caveolar membranes has been identified as PLD2. Northern blot analysis of PLD1 and PLD2 mRNA levels has indicated that PLD2 mRNA is elevated in both HT-29-MDR and MCF-7 AdrR cells. PLD1 mRNA levels were either unchanged or reduced in the MDR cells. Finally, in vivo experiments have confirmed previous results showing that activation of PLD by phorbol esters is markedly potentiated in the MDR cells. We conclude that MDR is accompanied by an increase in PLD2 activity in DIGs and caveolar membranes. PMID- 10709113 TI - Hormone-dependent nuclear localization of the tyrosine kinase iyk in the normal human breast epithelium and loss of expression during carcinogenesis. AB - iyk, a member of the frk family of non-receptor tyrosine kinases, was originally isolated from normal mouse mammary glands and is characterized by a nuclear localizing signal within the SH2 domain. We have investigated the expression and subcellular localization of iyk in the normal human breast and in malignant breast diseases. Immuno-histochemical analyses revealed that in normal tissue iyk localizes to both cytoplasmic and nuclear compartments of breast epithelial cells. The subcellular distribution was dependent on the hormonal state, being mostly cytoplasmic during the follicular, proliferative phase of the menstrual cycle, whereas frequent nuclear staining was observed in the resting stages during the luteal phase and, most prominently, after menopause. Strikingly, invasive carcinomas, irrespective of tumor type or hormonal status of the patient, exhibited almost complete loss of iyk expression in both the cytoplasm and the nucleus. In contrast, in situ breast carcinomas from post-menopausal patients showed a clear reduction of the nuclear iyk localization while retaining cytoplasmic staining. Our results indicate that iyk expression is gradually lost during carcinogenesis; thus, iyk may be classified as a tumor-suppressor gene. PMID- 10709114 TI - Major changes in social characteristics in oesophageal cancer patients in France. PMID- 10709115 TI - A functional and quantitative mutational analysis of p53 mutations in yeast indicates strand biases and different roles of mutations in DMBA- and BBN-induced tumors in rats PMID- 10709116 TI - Directory of on-going research in cancer prevention PMID- 10709117 TI - From the Editor's Pen PMID- 10709118 TI - Obesity in children in developing societies: indicator of economic progress or a prelude to a health disaster? PMID- 10709119 TI - Recombinant human erythropoietin in anemia of prematurity. AB - OBJECTIVE: To evaluate safety and efficacy of recombinant human erythropoietin (r HuEPO)in reducing the need for red cell transfusions in anemia of prematurity. METHODS: forty -two preterm infants (gestational age <32 weeks) were randomly assigned to a "treatment" group (r-HuEPO 400 units/kg every alternate day * 10 doses) or "no treatment" (control) group. All infants on enteral feeds received oral iron 3 mg/kg/day, graded up to 6 mg/kg/day. RESULTS: Higher reticulocyte counts in week 2 and 3 and higher hemoglobin levels in week 4 were noted after treatment with r-HuEPO. Despite stumulated erythropoiesis, the frequency of transfusions could not be reduced with r-HuEPO therapy.Overall, Phlebotomy losses, frequency and volume of redcell transfusions were significantly more in neonates with birthweight <1000 grams compared with those with birthweight >1000 grams (p<0.05). Associated side effects of r-HuEPO such as neutropenia,sepsis, hypertension or increased risk of late death did not occur. CONCLUSION: r-HuEPO therapy was safe without any side effects. Inability of r-HuEPO therapy to minimize red cell transfusions for anemia of prematurity may be explained by a relatively strict red-cell transfusion policy and the desired degree of treatment effect. PMID- 10709120 TI - Langerhans cell histiocytosis in children less than 2 years of age. AB - OBJECTIVE: To study the clinical profile and outcome of langerhans cell histiocytosis in children upto 2 years of life. DESIGN: Retrospective analysis. METHODS: Medical records of Children upto 2 years of age with a diagnosis of langerhans cell histiocytosis (LCH) were analyzed. Their clinical pattern, treatment modalities and outcome were studied. The patients Were categorized into 2 groups according to their clinical presentation: (i) Subject without organ dysfunction; and (ii) cases with organ dysfunction. Treatment considered of surgical intervention, radiotherapy, chemotherapy or combination of all these modalities depending upto the extent of disease. RESULTS: There were 20 children upto 2 years of age with histiocytosis during the 12 year period (January 1983 - December 1994). The median age at diagnosis was 18 months (range 52 days - 24 months). Of the twenty patients,13 patients didn't have organ dysfunction and 7 had organ dysfunction. Out of the 13 children without organ dysfunction eleven patients received treatment and all of them are alive free of disease with a median follow up of 62 months. But all children with organ dysfunction succumbed to disease within a few weeks. CONCLUSION: Children under 2 years of age with localised and or multifocal LCH without organ dysfunction have a good prognosis and they should not be exposed to aggressive form of treatment. All children with organ dysfunction require multi-agent chemotherapy. PMID- 10709121 TI - Nutritional status and dietary intake in tribal children of Bihar. AB - OBJECTIVE: To assess the dietary intake and nutritional status in children of the tribal areas of Bihar. DESIGN: Cross sectional survey with two stage probability proportional to size sampling. SETTING: Study covered 396 villages from 17 tribal districts of Bihar. SUBJECTS: 1847 preschool children (0-6 Years) were studied. METHODS: 24 hours recall method was used to assess the nutrition intake and anthropometric measurements included height and weight. Nutritional intake was compared with Indian Council of Medical Research recommended dietary allowances (RDA) and nutritional status assessed by SD classification. RESULTS: The intake of protein was broadly in line with the recommended dietary allowances (RDA) in all age groups among children. However, the average intake of energy and other nutrients was lower in allage groups as compared to RDA. Calorie deficiency was 38% whereas protein deficiency was about 19%. More than half of the children were caloric deficient in Katihar, Bokaro, Godda and Singhbhum (east and west). The overall prevalence of stunting was about 60% and underweight about 55% and was comparable in boys and girls. However, wasting was more frequent in girls (urban 34.5% vs. 16.3% and rural - 34.9% vs 18%). The level of malnutrition was not very different in rural and urban areas. CONCLUSION: The nutritional status and dietary intakes of tribal children in Bihar is very poor. Urgent remedial measures are required in this context, particularly on a war footing in especially vulnerable districts identified by this survey. PMID- 10709122 TI - Undergraduate pediatric education at BPKIHS integrated with an innovative curriculum. PMID- 10709123 TI - The role of intravenous immunoglobulins in pediatric diseases. PMID- 10709124 TI - Latex agglutination test (LAT) for the diagnosis of typhoid fever. PMID- 10709125 TI - Malignant recessive osteopetrosis. PMID- 10709126 TI - Health status of school girls from affluent population of Mumbai. PMID- 10709127 TI - Angiotensin converting enzyme inhibitor fetopathy. PMID- 10709128 TI - Class I histiocytosis: response to combination of etoposide and prednisolone. PMID- 10709129 TI - Paroxysmal torticollis of infancy. PMID- 10709130 TI - Lateral cranial meningocele. PMID- 10709131 TI - Sturge Weber syndrome. PMID- 10709132 TI - Congenital lumbar hernia. PMID- 10709133 TI - Routine hepatitis B and typhoid vaccination. PMID- 10709134 TI - Reply PMID- 10709135 TI - Immunogenecity of hepatitis A vaccine in children below 2 years of age. PMID- 10709136 TI - Carbamazepine hypersensitivity syndrome. PMID- 10709137 TI - Sunlight exposure in young infants. PMID- 10709139 TI - Lateral thinking in clinical practice. PMID- 10709138 TI - Rational drug therapy (RDT) - from where to embark on? PMID- 10709140 TI - Long-term performance of heart valve prostheses. PMID- 10709141 TI - Muscle strength and tissue composition in women as assessed by isokinetic dynamometry and dual energy X-ray absorptiometry. Experimental and clinical investigations within the field of rheumatology. PMID- 10709142 TI - 5-Aminosalicylic acid containing drugs. Delivery, fate, and possible clinical implications in man. AB - Since the beginning of the 1940s salazosulfapyridine (SASP) has been used in the treatment of chronic inflammatory bowel disease (CIBD). Almost 40 years later 5 aminosalicylic acid (5-ASA), which is split off by azo-reducing enzymes in the colon, was identified as the therapeutically active moiety of SASP. Thus different 5-ASA containing drugs were produced from which 5-ASA is released in the small and large intestine in a pH-dependent manner. Since there is a firm clinical indication that the 5-ASA concentration in the gut lumen is decisive for the therapeutic effect, a method was developed to evaluate the 5-ASA concentration at different levels in the intestine. The method was subsequently used to clarify factors of importance for the release of 5-ASA from the preparations. Ileostomy patients and healthy volunteers were investigated during continuous treatment with the three 5-ASA containing drugs with pH-dependent 5 ASA release: Asacol, Mesasal (Salofalk, Claversal), and Pentasa. The study confirmed release of 5-ASA in the small intestine from all preparations, but at different levels and speeds. Despite similar peroral dosage, very different 5-ASA concentration profiles were found in the ileostomy effluents, reflecting not only the difference in the release pattern of the preparations, but also the influence of the gastric residence time for larger sized tablets. The 5-ASA concentrations increased in the faeces of healthy volunteers. Furthermore the systemically absorbed fraction of 5-ASA was larger than previously found after SASP. The 5-ASA release from the preparation with the most proximal release, Pentasa, was less influenced by acceleration of the intestinal transit time than previously demonstrated after SASP in a similar study design. A comparative study of children given SASP and Pentasa showed similar results as in adults: a tendency for smaller 5-ASA concentration at rectal level after Pentasa than after SASP, and also larger systemic absorption. Despite higher 5-ASA dose per kg body weight, lower 5-ASA concentrations were seen in the faeces after both preparations, compared with adults. A peroral dose increase of Pentasa in healthy adults resulted in higher intraluminal 5-ASA concentration in the gut lumen, but also in saturation of the local and probably also systemic acetylation capacity, demonstrated by higher plasma concentrations and larger urinary excretion of 5 ASA. Similar faecal water concentrations were found after Pentasa 4 g and the azo bond preparation with colonic 5-ASA release, Dipentum (olsalazine) 2 g, confirming the substantial 5-ASA release from Pentasa in the small intestine. Investigation of pregnant patients treated with different 5-ASA containing drugs showed a similar pattern to SASP treatment: small amounts of 5-ASA cross the placenta, whereas the concentration of the metabolite Ac-5-ASA is similar in the maternal and fetal circulations. Only minute amounts of 5-ASA were found in milk from nursing mothers, while the concentrations of Ac-5-ASA were considerably higher. The decrease in the semen quality during SASP treatment was improved by changing to a controlled-release 5-ASA drug. The concentrations of 5-ASA in seminal plasma were similar during the two treatment periods, but higher of its metabolite Ac-5-ASA during treatment with the controlled-release preparation. That indicates that the toxic effect after SASP is not caused by the 5-ASA or Ac 5-ASA moiety. All the preparations have proved effective in the treatment of ulcerative colitis, but data concerning the 5-ASA treatment of Crohn's disease are conflicting. Knowledge of the demonstrated differences in the release profiles of the 5-ASA containing drugs is therefore important when designing future clinical trials. PMID- 10709143 TI - [In vitro activities of carbapenem antibiotics against the various clinical isolates]. AB - The annual changes of antibacterial activities of beta-lactam antibiotics, mainly carbapenem antibiotics, were investigated against 5 bacterial species, S. aureus (MSSA), methicillin-resistant S. aureus (MRSA), Klebsiella pneumoniae, Serratia marcescens, Pseudomonas aeruginosa, which had been isolated from the clinical materials at Toho University Omori Hospital during the period of 1995 to 1997. In addition, antibacterial activities against other main bacterial strains isolated from the clinical materials during 1997 were also determined. The five bacterial species on which annual changes of the sensitivity were investigated did not show any remarkable trend to increase in resistance to the carbapenem antibiotics tested. The antibacterial activities of the carbapenem antibiotics against MRSA were weak, and MIC90 values were between 25 and 50 micrograms/ml. In S. marcescens and P. aeruginosa on which high resistance by the production of metallo-beta-lactamase has become a problem in recent years, there were no remarkable changes in annual changes of sensitivities. Especially, MIC90 valuses of the carbapenem antibiotics against P. aeruginosa were between 12.5 and 25 micrograms/ml, 4 to 8 times better than that of PIPC, like the case of CAZ. Furthermore, the carbapenem antibiotics showed strong antibacterial activities against clinically important 16 bacterial species, from Gram-positive to Gram negative bacteria. PMID- 10709144 TI - [Clinical efficacy of cefpirome sulfate against Bacteroides species, Prevotella species and Porphyromonas species. Society of Anaerobic Bacterial Infections in the fields of obstetrics and gynecology in Gifu]. AB - The injectable cephalosporin cefpirome (CPR) was launched in Japan in 1993. It has widely been used in the various infectious diseases. We therefore studied the clinical and bacteriological efficacy of CPR against infections caused by Bacteroides species, Prevotella species and Porphyromonas species frequently isolated from the obstetric and gynecologic infections. Thirteen institutions were involved in this study which ran from March 1994 to January 1999. The administration dosage of CPR was 2 to 4 gram per day administered by drip infusion or intravenous infusion. The duration of treatment was from 3 to 15 days. The evaluations were performed before and after the treatment. CPR was administered to 194 patients with obstetric and gynecologic infections, and 146 of 194 cases were acceptable for the evaluation of drug efficacy. Bacteroides species were identified in 102 patients. Clinical efficacy in 146 cases was excellent in 12 patients, good in 110, fair in 9 and poor in 15 patients. The eradication rate for Bacteroides species could be in 37 cases out of 54 evaluable cases; Prevotella species in 38 out of 49; and Porphyromonas species in 5 out of 5. The overall assessment of bacteriological efficacy was "eradicated" in 91 cases out of 133 (68.4%). Adverse reactions including abnormal findings in laboratory tests were seen in 8 patients (4.76%). Based on these results, CPR promises efficacy and safety in the treatment of obstetric and gynecologic infections due to Bacteroides species. PMID- 10709145 TI - [The resistance of recent clinical isolates against isepamycin, other aminoglycosides and injectable beta-lactams]. AB - Clinical isolates collected from clinical facilities across Japan in 1998 were tested against five aminoglycosides and three beta-lactams. The resistance of 50 strains each of methicillin sensitive Staphylococcus aureus, methicillin resistant Staphylococcus aureus (MRSA), Staphylococcus epidermidis, Escherichia coli, Citrobacter freundii, Klebsiella pneumoniae, Enterobacter sp., Serratia sp., Pseudomonas aeruginosa and Proteus sp. (P. mirabilis 25 strains and P. vulgaris 25 strains) to the aminoglycosides isepamicin (ISP), amikacin (AMK), gentamicin, tobramycin and dibekacin, and to the beta-lactams imipenem, ceftazidime and piperacillin (all three known to be effective against P. aeruginosa) were investigated using a micro liquid dilution method with the following results: 1. ISP was effective against all strains except for 14% of MRSA, 2% of Proteus sp., and 4% of P. aeruginosa. 2. Six strains of MRSA were resistant to all eight drugs; however, in these cases ISP exhibited a relatively low minimum inhibitory concentration (MIC) compared to the other compounds. 3. Four strains of MRSA were resistant to all drugs except ISP. MRSA was the only isolate to demonstrate a resistance to seven or more drugs. 4. Twenty-one strains of MRSA and 1 strain of P. aeruginosa were resistant to six drugs; however, all of these were susceptible to both ISP and AMK. 5. Against all strains tested, ISP generally exhibited a lower MIC compared to AMK. These results suggest that, even ten years after its entering the market, ISP is still an aminoglycoside having a high anti-bacterial activity against a wide range of clinical isolates. PMID- 10709146 TI - The art and science of chart review. AB - BACKGROUND: Explicit chart review was an integral part of an ongoing national cooperative project, "Using Achievable Benchmarks of Care to Improve Quality of Care for Outpatients with Depression," conducted by a large managed care organization (MCO) and an academic medical center. Many investigators overlook the complexities involved in obtaining high-quality data. Given a scarcity of advice in the quality improvement (QI) literature on how to conduct chart review, the process of chart review was examined and specific techniques for improving data quality were proposed. METHODS: The abstraction tool was developed and tested in a prepilot phase; perhaps the greatest problem detected was abstractor assumption and interpretation. The need for a clear distinction between symptoms of depression or anxiety and physician diagnosis of major depression or anxiety disorder also became apparent. In designing the variables for the chart review module, four key aspects were considered: classification, format, definition, and presentation. For example, issues in format include use of free-text versus numeric variables, categoric variables, and medication variables (which can be especially challenging for abstraction projects). Quantitative measures of reliability and validity were used to improve and maintain the quality of chart review data. Measuring reliability and validity offers assistance with development of the chart review tool, continuous maintenance of data quality throughout the production phase of chart review, and final documentation of data quality. For projects that require ongoing abstraction of large numbers of clinical records, data quality may be monitored with control charts and the principles of statistical process control. RESULTS: The chart review module, which contained 140 variables, was built using MedQuest software, a suite of tools designed for customized data collection. The overall interrater reliability increased from 80% in the prepilot phase to greater than 96% in the final phase (which included three abstractors and 465 unique charts). The mean time per chart was calculated for each abstractor, and the maximum value was 13.7 +/- 13 minutes. CONCLUSIONS: In general, chart review is more difficult than it appears on the surface. It is also project specific, making a "cookbook" approach difficult. Many factors, such as imprecisely worded research questions, vague specification of variables, poorly designed abstraction tools, inappropriate interpretation by abstractors, and poor or missing recording of data in the chart, may compromise data quality. PMID- 10709147 TI - Reconciling quality measurement with financial risk adjustment in health plans. AB - BACKGROUND: Initiatives to improve quality measurement (QM) and to create systems for financial risk adjustment (RA) have developed in response to concerns about price competition's threat to quality and stimulation of risk selection. QM is designed to help purchasers identify best plans, to aid plans in their selection of providers, to facilitate quality improvement by plans and providers, and to assist patients faced with choices among plans and providers. The goal of RA is to eliminate incentives for plans and providers to avoid sick, high-cost patients in favor of healthy, low-cost patients. CONFLICTS BETWEEN QM AND RA: For QM it is often necessary to identify all patients with a particular condition, and many quality measures involve intervening on patients early in the course of their disease. Identifying patients through utilization decisions (for example, identifying patients with depression through receipt of an antidepressant prescription) may bias QM. For RA, the focus is on the highest-cost patients, and patient capture through resource utilization is more likely to be appropriate. DISCUSSION: Achieving QM and RA depends on improving information systems and patient identification processes and developing standard definitions for important variables. QM and RA could both be improved, and the conflicts between them reduced, if they were based more on detailed clinical data, if consensus definitions of quality of care for specific diagnoses could be achieved, if the number of QM measures that target acute and chronic care (versus preventive care) were increased, and if information systems were enhanced. PMID- 10709148 TI - Infant mortality review as a vehicle for quality improvement in a local health department. AB - BACKGROUND: Many communities across the United States have established fetal and infant mortality review (FIMR) programs as a way of gaining insight into the causes of such deaths and of devising and implementing ways to improve the health of pregnant women and their infants. IMR PROCESS: The IMR process in the Jefferson County Department of Health in Birmingham, Alabama, evolved in a somewhat different fashion than that in other communities. A technical review team reviews all the infant deaths in the county, with particular attention to each woman's pregnancy history. A community review team reviews composite cases that illustrate some particular problem that might lead to infant mortality, such as teenage pregnancy or short intervals between pregnancies. This team provides insights into cultural patterns and a community perspective on the problems. Recommendations from the two teams are acted on by the health department, with the assistance of other agencies as needed. IMPACT OF THE IMR PROCESS: The IMR process has been used to increase community agency participation in health department activities, improve health department procedures, increase health department staff acceptance of a new and controversial program (Healthy Start), and offer services to women who need them. CONCLUSIONS: IMR has become a mechanism for CQI in the health department, embodying many of the principles of CQI, including the use of teams, focus on a team mission, and examination of processes, not individuals. The program offers a model of how to reduce rates of fetal and infant mortality. PMID- 10709149 TI - The accreditation status of student health services at academic medical centers. AB - BACKGROUND: Student health services (SHSs) on campuses with academic medical centers (AMCs; schools affiliated with teaching hospitals) often provide clinical services (including occupational health services) for students and staff. There are few aggregate data about the accreditation status of these health services. METHOD: A cross-sectional survey of SHSs at the 124 medical schools in the United States was conducted in spring 1999. SHSs were questioned about their accreditation status. RESULTS: One-hundred three SHSs were included in the survey. Forty-seven (46%) of the SHSs were accredited, 30 (64%) through the Joint Commission on Accreditation of Healthcare Organizations, and the remainder through the Accreditation Association of Ambulatory Health Care (AAAHC). The Joint Commission predominated among private SHSs (13/15 [87%]), but public institutions were more evenly divided. Most of the SHSs accredited as freestanding ambulatory care centers were AAAHC accredited (16/27 [59%]). Nearly all the SHSs accredited as satellites of the AMCs were Joint Commission accredited. CONCLUSION: Although fewer than one-half of SHSs at AMCs are accredited either through the Joint Commission or AAAHC, this is a far higher percentage than of accredited SHSs in general (134/1,500 [9%]). The Joint Commission dominates as the accrediting body for SHSs at AMCs largely because of the high-proportion of services accredited as satellites of the AMCs. Although the accreditation process is costly and time-consuming and may appear daunting, SHSs at AMCs should pursue accreditation to ensure quality in health care processes. PMID- 10709150 TI - Gender-specific effects on verapamil pharmacokinetics and pharmacodynamics in humans. AB - Pharmacokinetic studies of i.v. and oral racemic verapamil and 14C-erythromycin breath tests (ERBT) were performed in 84 healthy men (n = 42) and women (n = 42). Verapamil was measured by HPLC, concentration versus time data were analyzed by noncompartmental models, protein binding was measured by equilibrium dialysis, and statistical analyses were performed by ANOVA. Clearance of i.v. and p.o. verapamil was 13.7 +/- 4.3 and 58.4 +/- 35 ml/min/kg (mean +/- SD) in women compared to 12.6 +/- 3.4 and 82.6 +/- 70 ml/min/kg in men (p = 0.076). Bioavailability was higher in women (0.25 +/- 0.09) compared to men (0.20 +/- 0.09, p = 0.019) with a significant Gender x Formulation interaction (p = 0.04). ERBT were higher in women (p < 0.0001). Verapamil (i.v. and p.o.) decreased blood pressure in all subjects with greater heart rate increases after p.o. verapamil in women compared to men (p = 0.041). The findings suggest that sex-specific differences in drug metabolism may occur in both the gut and the liver and involve multiple metabolic pathways and that pharmacokinetic differences will alter pharmacodynamic responses. PMID- 10709151 TI - A pharmacokinetic and pharmacodynamic study of the potential drug interaction between tasosartan and atenolol in patients with stage 1 and 2 essential hypertension. AB - The primary objective of this study was to evaluate the pharmacokinetics (PK) and pharmacodynamics (PD) of tasosartan and atenolol administered alone and concomitantly under steady-state conditions in 17 patients ages 18 to 65 years diagnosed with stage 1 to 2 essential hypertension. After a 3- to 14-day qualification period, all patients received placebo tasosartan on days--1 through 5 and 25 through 34, atenolol alone (50 mg) on days 1 through 5, atenolol (50 mg) + tasosartan (50 mg) on days 6 through 19, and tasosartan (50 mg) alone on days 20 through 24. A PK and PD evaluation of atenolol alone was performed on study day 5. On study day 19, PK and PD of both tasosartan and atenolol were assessed. PK and PD evaluation for tasosartan alone was assessed on study day 24. The coadministration of atenolol + tasosartan did not affect the pharmacokinetics of tasosartan, its major metabolite (enoltasosartan), or atenolol when compared with tasosartan or atenolol administered separately. For area under the change in diastolic blood pressure curve, the reduction was significantly greater after tasosartan + atenolol compared with that after atenolol alone (336 +/- 85 and 190 +/- 71 mmHg.24 h; p < 0.05 for combination and atenolol alone, respectively; mean +/- SEM). Combination therapy also caused a maximal reduction in diastolic blood pressure that is significantly more than with monotherapy with atenolol (-27 +/- 2 mmHg and -20 +/- 2 mmHg, respectively, p < 0.05). The additive effects of tasosartan and atenolol in decreasing diastolic blood pressure may provide a rationale for combination antihypertensive therapy. PMID- 10709152 TI - A population pharmacokinetic analysis of zanamivir in subjects with experimental and naturally occurring influenza: effects of formulation and route of administration. AB - The pharmacokinetics of zanamivir were evaluated in subjects from three phase I single-center and two phase II multicenter, randomized, double-blind, multidose, placebo-controlled trials. A total of 96 phase I subjects received zanamivir (3.6 to 16 mg) intranasally two or six times daily for 4 to 5 days beginning 4 hours before or 1 to 2 days after inoculation with influenza virus. A total of 75 phase II subjects with influenza or a history of exposure to naturally occurring influenza virus were administered zanamivir as an intranasal spray (3.4 mg/nostril), inhaled powder (10 mg), or combination of intranasal and inhaled formulations twice daily for 5 days. Population parameters (including demographic factors, zanamivir formulation, infection-related variables, and concurrent medication use) were estimated by a nonlinear mixed-effect modeling software program (NONMEM) using a one-compartment model with first-order absorption and conditional estimation algorithm. Formulation and route of administration were the most significant factors affecting the pharmacokinetics of zanamivir. Relative bioavailability of the inhaled powder to the intranasal drops and spray was 2.3 and 1.6, respectively. No significant differences in pharmacokinetic parameters were observed when demographic variables, indices of infection, or concurrent medication use were considered in either phase I or phase II population analyses. PMID- 10709154 TI - Dose-proportional pharmacokinetics of risedronate on single-dose oral administration to healthy volunteers. AB - Risedronate is a pyridinyl bisphosphonate approved for the treatment of Paget's disease (US-FDA) and in development for the treatment and prevention of osteoporosis. This study examined risedronate pharmacokinetics and tolerability after oral administration using a randomized, double-blind, parallel-group design. Healthy male and female volunteers (n = 22-23 subjects per dose) received a single oral dose of 2.5, 5, or 30 mg risedronate. Serum and urine samples were collected for 72 and 672 hours, respectively, and risedronate concentrations were determined by ELISA. Safety was evaluated by monitoring adverse events, clinical laboratory measurements, vital signs, and electrocardiograms. Mean Cmax (0.41, 0.94, and 5.1 ng/mL for 2.5, 5, and 30 mg, respectively), AUC (1.8, 3.9, and 21 ng.h/mL for 2.5, 5, and 30 mg, respectively), and urinary excretion (22, 63, and 260 micrograms for 2.5, 5, and 30 mg, respectively) were dose proportional, and there were no significant differences in tmax or CLR among the three doses. All doses were well tolerated; no serious adverse events occurred, and all but one of the adverse events were mild or moderate in severity. There was no evidence of an acute phase reaction occurring after oral administration of risedronate. PMID- 10709153 TI - Pharmacokinetics of diclofenac sodium in chronic active hepatitis and alcoholic cirrhosis. AB - The objective of this study was to assess the pharmacokinetics of diclofenac sodium and its five metabolites following administration of a 150 mg oral dose to healthy subjects and patients with either chronic active hepatitis of varying morphology or alcoholic cirrhosis. Six healthy subjects, 6 chronic active hepatitis patients, and 6 alcoholic cirrhosis patients were enrolled in this prospective, open-label, parallel study. Blood samples were drawn at 0, 0.25, 0.5, 0.75, 1, 2, 4, 6, 8, 12, 24, 48, 72, 144, 312, and 480 hours, and urine samples were collected for 144 hours after administration of a single oral dose of diclofenac sodium. The mean area under the serum concentration-time curve extrapolated to infinity, oral clearance, half-life, maximal concentration, and time to peak concentration for diclofenac and its metabolites were determined and compared using analysis of variance. Cirrhotics had a mean +/- SD diclofenac AUC value (19,114 +/- 6806 ng.h/ml) significantly different (p < 0.02) from hepatitis patients (6071 +/- 1867 ng.h/ml) and healthy subjects (7008 +/- 2006 ng.h/ml), whereas healthy subjects and hepatitis patients had similar values. Comparable results were found for 4'-hydroxydiclofenac. The AUC values for 3' hydroxydiclofenac and 3'-hydroxy-4'methoxydiclofeanc were significantly different when healthy subjects were compared to cirrhotics. However, hepatitis subjects were not significantly different from either group. The results indicate that hepatitis does not alter the pharmacokinetics of diclofenac. Alcoholic cirrhosis increased the mean diclofenac AUC approximately three times compared to normal subjects, indicating that one-third of the usual dose in cirrhotics would produce equivalent AUC values in normal subjects and subjects with alcoholic cirrhosis. However, since pharmacodynamic measurements were not made and no increase in untoward or side effects was noted in the alcoholic cirrhosis patients after a single dose, maintenance doses should be titrated to patients response. PMID- 10709155 TI - Pharmacokinetic-pharmacodynamic modeling of testosterone and luteinizing hormone suppression by cetrorelix in healthy volunteers. AB - Cetrorelix (CET), a potent luteinizing hormone-releasing hormone (LH-RH) antagonist, was recently approved for the prevention of premature ovulation in patients undergoing a controlled ovarian stimulation (COS), followed by oocyte pickup and assisted reproductive techniques (ART), and is currently under clinical trials for benign prostate hyperplasia, endometriosis, and tumors sensitive to sex hormones. CET suppresses luteinizing hormone (LH), follicle stimulating hormone (FSH), and testosterone (T) in men. The purpose of this study was to evaluate the pharmacokinetics and absolute bioavailability of 3 mg intravenously and subcutaneously administered CET in healthy male and female volunteers and to develop a pharmacokinetic-pharmacodynamic (PK-PD) model to link the plasma concentrations of CET to the T and LH suppression in males. Following intravenous (IV) (n = 5) and subcutaneous (SC) (n = 6) administration of CET acetate, CET and hormone plasma levels were measured by radioimmunoassay (RIA) and enzyme immunoassay (EIA) methods, respectively. Pharmacokinetics of CET was explained by a three-compartment model for the IV route and by a two-compartment model with first-order absorption for the SC route. Average absolute bioavailability after SC administration was 85%. There were no differences in the pharmacokinetics between male and female subjects (ANOVA, p > 0.05). Single IV and SC doses of CET caused immediate and distinct suppression of LH, FSH, and T levels by 80%, 45% and 95% of their baseline levels, respectively. The duration of hormone suppression was longer for the SC route. An indirect-response PK-PD Emax model was developed to link the measured CET plasma concentrations with the respective T or LH levels. In addition, the circadian rhythm of T levels was accounted by including a cosine function in a second separate PD model. The PD model with cosine function was applied to T baseline levels as well as to the suppressed concentrations after CET dosing. The two models adequately described the PK-PD relationship between plasma levels of CET and T suppression following IV and SC dosing. The EC50 values (mean +/- SD) for the suppression of T were similar (p > 0.05) between the two routes of administration and the two models. PMID- 10709156 TI - Population pharmacokinetics/toxicodynamics (PK/TD) relationship of SAM486A in phase I studies in patients with advanced cancers. AB - SAM486A (previously termed CGP 48664), a potent inhibitor of S-adenosylmethionine decarboxylase, is under clinical development for the treatment of advanced refractory malignancies. Hematological toxicity manifested by dose-dependent neutropenia has been observed in phase I studies. Population methods were used to investigate pharmacokinetics (PK) as a prognostic factor for safety end point (hematological toxicity) in patients with advanced cancers. SAM486A plasma concentrations and neutrophil counts were collected from three ascending-dose tolerability and PK studies (study 1: single 5-day continuous intravenous (IV) infusion with doses ranging from 24-700 mg/m2/cycle; study 2: 10-minute to 3-hour IV infusion once weekly with doses ranging from 16-325 mg/m2/week; study 3: 1 hour IV infusion once daily for 5 days with doses ranging from 3.6-202.8 mg/m2/day). The PK of SAM486A were best estimated by a population linear three compartment model with NONMEM (version 5) using data from 9 patients in studies 1 through 3. The population pharmacokinetic parameters (SD) were CL = 6.2 (0.4) l/h/m2, Q2 = 15.4 (1.5) l/h/m2, Q3 = 33.6 (5.3) l/h/m2, V1 = 9.5 (1.6) l/m2, V2 = 672 (52) l/m2, and V3 = 39.9 (8.3) l/m2, and the corresponding intersubject variability was 45.4%, 74.0%, 85.3%, 80.1%, 37.0%, and 103%, respectively, where CL is total body clearance, Q2 and Q3 are intercompartmental clearances, and V1, V2, and V3 are the volumes of distribution in central and peripheral compartments, respectively. The intrasubject variability was 24.0%. The cumulative AUC before the onset of neutrophil nadir count (AUC) and the duration of exposure over threshold SAM486A concentrations in the range of 0.05 to 0.1 microM based on Bayesian PK parameter estimates significantly correlated with absolute neutrophil count nadir (< 5 x 10(9)/l). AUC showed the best correlation (R2 = 0.72) with absolute neutrophil count nadir by an inhibitory sigmoid Emax model and also correlated with percent decrease in neutrophil count from baseline to nadir by a simple Emax model (R2 = 0.53). Logistic regression analysis indicated that AUC and the duration of exposure over 0.05 to 0.1 microM, but not Cmax, were strong predictors of grade 4 neutropenia (< 0.5 x 10(9)/l). Drug exposure parameters such as AUC derived from population analysis may be used clinically as a useful predictor of drug-induced neutropenia. PMID- 10709157 TI - Higher incidence of elevated body temperature or increased C-reactive protein level in asthmatic children showing transient reduction of theophylline metabolism. AB - The authors investigated whether theophylline metabolism is decreased in asthmatic patients and what condition may be related to its reduction. Fifty-two children with asthma were given 15 mg/kg/day aminophylline intravenously at a constant rate. Blood and spot urine samples were collected at 24 hours, 48 hours, and 72 hours after beginning infusion. The ratio of plasma theophylline concentration at 72 hours to that at 24 hours (C72h/C24h) varied from 0.42 to 1.51 (average 0.894). Plasma theophylline concentration of patients with lower C72h/C24h than average reduced significantly, while the concentration of those with higher C72h/C24h remained unchanged. The urinary ratio of the sum of the metabolites to theophylline was significantly increased in the patients with the lower ratio. Among the demographic characteristics examined, significant difference was found only in the incidence of patients with C-reactive protein (CRP) of 0.5 mg/dl or greater or patients with a fever of 37.5 degrees C or greater when admitted. Acute febrile illness accompanied by increased CRP level may affect theophylline metabolism. PMID- 10709158 TI - Cerebrospinal fluid pharmacokinetics of cefpodoxime proxetil in piglets. AB - Cefpodoxime is an oral third-generation cephalosporin used for the treatment of acute upper-respiratory tract infections caused by susceptible bacteria in children. Although not indicated for the treatment of bacterial meningitis, it is used to treat other infections produced by organisms associated with meningitis and may obscure the result of cerebrospinal fluid (CSF) cultures in children who develop meningitis while receiving oral antibiotics if sufficient concentrations are achieved in the CSF. This study evaluated the disposition of cefpodoxime and penetration into CSF in piglets. Fifteen Landacre-Camborough cross piglets (10-20 days old) received cefpodoxime proxetil oral suspension (10 mg/kg). Repeated plasma and CSF samples were collected over 24 hours for quantitation of cefpodoxime by HPLC. Pharmacokinetic analysis was performed on both plasma and CSF data. The plasma concentration versus time data for cefpodoxime were best characterized using a one-compartment model with first-order absorption. The mean (+/- SD) pharmacokinetic parameters for Cmax, tmax, and AUC0-infinity were 23.3 +/- 12.9 mg/L, 3.9 +/- 1.4 h, and 237 +/- 129 mg/L.h, respectively. CSF/plasma ratios for AUC0-infinity demonstrated a mean cefpodoxime penetration of approximately 5%. CSF penetration of cefpodoxime was evident following a single oral dose of cefpodoxime proxetil suspension. Despite the small percentage of total cefpodoxime dose distributing into the CSF, the resultant concentrations approached or exceeded the MIC90 for many bacterial pathogens considered susceptible to cefpodoxime. Accordingly, clinicians should use caution in the interpretation of CSF cultures in patients who develop clinical signs and symptoms consistent with meningitis and who have been previously treated with cefpodoxime. PMID- 10709159 TI - Acute encephalopathy due to aluminum toxicity successfully treated by combined intravenous deferoxamine and hemodialysis. AB - Acute aluminum intoxication is uncommon in clinical practice but can be fatal. This limited experience is reflected in the paucity of data assessing a viable approach to the treatment of these patients. In this report, the authors describe the clinical course and successful, pharmacokinetic-based deferoxamine hemodialysis treatment regimen of a patient with severe aluminum encephalopathy following alum bladder irrigation. The combined use of deferoxamine and appropriately timed hemodialysis appears to be a very reasonable means of treating patients with severe acute aluminum intoxication. PMID- 10709160 TI - Lack of interaction between lansoprazole and propranolol, a pharmacokinetic and safety assessment. AB - Due to the prevalence of both gastrointestinal and cardiovascular diseases, it is likely that patients may be coprescribed gastric parietal cell proton pump inhibitors and beta-adrenergic antagonists. Therefore, the objectives of this phase I study were to assess the potential effects of the coadministration of lansoprazole on the pharmacokinetics of propranolol and to evaluate the safety of propranolol with concomitant lansoprazole dosing. In a double-blind fashion, 18 healthy male nonsmokers were initially randomized to receive either 60 mg oral lansoprazole, each morning for 7 days, or an identical placebo (period 1). On day 7, all subjects were concomitantly administered oral propranolol, 80 mg. After a minimum of 1 week following the last dose of either lansoprazole or placebo, subjects were crossed over to the opposite treatment for another 7 days (period 2). Subjects were again administered oral propranolol on day 7. During both treatment periods, blood samples for the determination of plasma propranolol and 4-hydroxy-propranolol were obtained just before the dose and at 0.5, 1, 2, 3, 4, 6, 8 12, 16, 20, and 24 hours postdose. Plasma propranolol and 4-hydroxy propranolol concentrations were determined by using HPLC with fluorescence detection. The Cmax, tmax, AUC0-infinity, and t1/2 values for propranolol, as well as the AUC0-infinity for 4-hydroxy-propranolol, were calculated and compared between the lansoprazole and placebo regimens. The mean age of the 15 subjects who successfully completed the study was 31 years (range: 24-38 years), and their average weight was 174.8 pounds (range: 145-203 pounds). There were no statistically significant differences between the lansoprazole and placebo regimens for the propranolol Cmax, tmax, AUC0-infinity, and t1/2 values. Also, there were no statistically significant differences between regimens for the 4-OH propranolol AUC0-infinity. Safety evaluations, which included adverse events, vital signs, clinical laboratory determinations, ECG, and physical examinations, revealed no unexpected clinically significant findings and did not suggest a drug drug interaction. In conclusion, lansoprazole does not significantly alter the pharmacokinetics of propranolol, suggesting that it does not interact with the CYP2D6- or CYP2C19-mediated metabolism of propranolol. Modification of a propranolol dosage regimen in the presence of lansoprazole is not indicated, based on the pharmacokinetic analysis and the lack of a clinically significant alteration in the pharmacodynamic response. PMID- 10709161 TI - A double-blind, placebo-controlled evaluation of the effect of oral doses of rizatriptan 10 mg on oral contraceptive pharmacokinetics in healthy female volunteers. AB - Rizatriptan (MAXALT), a potent, oral 5-HT1B/1D agonist with a rapid onset of action, is available now for the acute treatment of migraine. This study examined the pharmacokinetic and clinical interaction between rizatriptan 10 mg and the components (ethinyl estradiol [EE] 35 micrograms and norethindrone [NET] 1.0 mg) of a well-established oral contraceptive combination product, ORTHO-NOVUM 1/35. Levels of sex hormone binding globulin (SHBG), a protein increased by EE to which NET binds, were also examined. In this two-period crossover study, 20 healthy young female subjects received a coadministration of 8 days of rizatriptan treatment (6 days of single-dose 10 mg rizatriptan and 2 days of multiple-dose rizatriptan, 10 mg q 4 hours for three doses, giving a total daily dose of 30 mg on Days 7 and 8) or matching placebo along with their daily dose (one tablet) of ORTHO-NOVUM 1/35 within their oral contraceptive cycle. Plasma was sampled for EE, NET, and SHBG concentrations. Safety evaluations included routine laboratory safety studies, physical examinations, and monitoring for ECG, vital signs, and adverse events. There were no statistically significant differences in any of the pharmacokinetic parameters of EE or NET between the rizatriptan and placebo treatment periods, thus indicating that rizatriptan had no meaningful effect on the disposition of either the EE or the NET component of ORTHO-NOVUM 1/35. The SHBG concentration did not change throughout the entire study. Clinically, coadministration of rizatriptan with ORTHO-NOVUM 1/35 was well tolerated. Blood pressure, heart rate, and temperature showed no consistent trend or clinically important changes. Adverse events following coadministration of rizatriptan with ORTHO-NOVUM 1/35 were similar to those reported when placebo was given with ORTHO NOVUM 1/35. The findings of this study indicate that there is little potential for dosages as high as 30 mg/day, the maximum recommended dosing schedule, of rizatriptan to alter the plasma concentrations of oral contraceptives. PMID- 10709162 TI - Lack of a clinically significant pharmacokinetic interaction between fenofibrate and pravastatin in healthy volunteers. AB - This study was conducted to evaluate the potential pharmacokinetic interaction between fenofibrate and pravastatin. A total of 23 healthy adult volunteers received single-dose 201 mg fenofibrate alone, 201 mg fenofibrate + 40 mg pravastatin, and 40 mg pravastatin alone in a three-period crossover experiment. Plasma samples were collected at predetermined times and were analyzed with validated methods for the quantitation of fenofibric acid, pravastatin, and 3 alpha-hydroxy-isopravastatin (3 alpha-iso-PV). Pharmacokinetic parameters of these three compounds were calculated using noncompartmental methods and compared by analyses of variance and bioavailability assessments. Concomitant administration of fenofibrate and pravastatin did not affect the pharmacokinetics of either fenofibric acid or pravastatin. However, the AUC0-infinity and Cmax of 3 alpha-iso-PV were increased by 26% and 29%, respectively. The moderate increase in the formation of this pravastatin metabolite should not raise any clinical concerns due to its much lower pharmacological potency compared to pravastatin and lack of toxicity. PMID- 10709163 TI - Family and medicine. PMID- 10709164 TI - Recurrence of symptoms following treatment of posterior semicircular canal benign positional paroxysmal vertigo with a particle repositioning manoeuvre. AB - A consecutive series of 51 patients (34 females and 17 males) with posterior semicircular canal benign positional paroxysmal vertigo (BPPV) were treated with the modified Epley particle positioning manoeuvre (PRM). Follow-up data were available on all 51 patients. After one manoeuvre, 42 patients had a negative Dix Hallpike test, and after a second manoeuvre, 8 of the remaining 9 patients had a negative Dix-Hallpike test (testing was conducted 1 to 2 weeks after the PRM was performed). Therefore, the overall short-term success rate after two manoeuvres was 50 of 51 patients (98%), which is similar to other series. A follow-up questionnaire to determine the incidence of recurrence of symptoms was administered after a minimum period of 30 weeks. Twenty-three patients reported a recurrence (or, in the case of the one treatment failure, persistence) of their symptoms (45%). Therefore, although virtually all patients can be treated successfully with the PRM, almost half of these patients can be expected to experience a further recurrence of symptoms. This long-term recurrence rate is higher than has previously been reported and is a significant factor clinicians must be aware of in their treatment of this condition. In particular, this finding emphasizes the need for patient counselling with regard to the likelihood of recurrence and access to follow-up treatment if recurrence occurs. PMID- 10709165 TI - Outcome analysis of endoscopic sinus surgery for chronic sinusitis in patients having Samter's triad. AB - A study was conducted to assess outcome analysis in acetylsalicylic acid (ASA) triad patients after endoscopic sinus surgery (ESS). The control group consisted of patients with chronic sinusitis, with or without asthma, who had also undergone ESS. The study group contained 18 patients with the classic triad who were compared with 22 controls. The study was conducted in retrospective fashion highlighting clinical presentation, radiologic evaluation, surgical findings, and recurrence rate of nasal polyps. Although both groups had a relatively similar age of onset of symptoms, the symptomatic picture was different in the two groups. Radiologic evaluation of the nose and paranasal sinuses revealed more extensive involvement of the sinuses in ASA triad patients. Furthermore, ASA triad patients underwent a greater number of repeat operations. This review suggests that ASA triad patients respond less well to surgical intervention and that other treatment modalities should perhaps be explored. PMID- 10709166 TI - Submucosal arytenoidectomy: new surgical technique and review of the literature. AB - OBJECTIVE: Arytenoidectomy is indicated in cases of bilateral median vocal cord paralysis (most commonly due to recurrent laryngeal nerve paralysis), ankylosis of the cricoarytenoid joint due to arthritis, and tumours of the arytenoid cartilage. We propose the use of the submucosal approach, to excise the arytenoid cartilage in cases of vocal cord paralysis. We present the surgical technique and review the history and relevant literature, as well as the pros and cons of various surgical techniques for arytenoidectomy. SETTING: Department of Otolaryngology-Head and Neck Surgery, Rambam Medical Center, Haifa, Israel. METHOD: We present six cases: five cases of bilateral vocal cord paralysis and one case of a chondroma of the arytenoid with mechanical fixation of the cord. All patients suffered from dyspnea on mild exertion. An arytenoidectomy using the submucosal approach was performed on all six patients. RESULTS: Airway results were evaluated via fibre-optic videotape laryngoscopy and direct microlaryngoscopy. Voice was evaluated subjectively by the patients and by a speech therapist before and after surgery. Following the surgery, all six patients showed clinical improvement, they no longer suffered from dyspnea at rest or upon mild exertion, and they retained reasonable voice quality. CONCLUSION: The submucosal approach is not difficult to perform and preserves an intact laryngeal mucosa, which prevents the formation of granulation tissue and scarring, which may further obstruct the lumen. The resulting airway is good, with minimal compromise of phonation. We feel that the submucosal approach to arytenoidectomy is an important addition to the arsenal of many surgical techniques for the treatment of bilateral vocal cord paralysis. PMID- 10709167 TI - Assessment and management of idiopathic facial (Bell's) palsy: comparison of Nova Scotia family physicians and otolaryngologists. AB - Acute idiopathic facial paralysis, or Bell's palsy, is frequently encountered in clinical practice. The present study compares knowledge of Bell's palsy assessment and management between a group of family physicians and otolaryngologists practising in Nova Scotia. Respondents completed a questionnaire and statistical analyses were performed on selected data. There were similarities regarding Bell's palsy assessment and management, but there were notable differences in the ability to distinguish Bell's palsy on the basis of symptomatic complaints, specific counselling strategies, length of patient follow-up, and use of appropriate diagnostic tests. This needs assessment suggests several areas where a family physician continuing medical education program on management of acute facial paralysis may be beneficial. PMID- 10709168 TI - Effects of radiotherapy for nasopharyngeal carcinoma on the paranasal sinuses: study based on computed tomography scanning. AB - OBJECTIVE: The purpose of this study was to determine if radiotherapy for nasopharyngeal carcinoma causes mucosal disease of the paranasal sinuses. DESIGN: This study was a retrospective study. SETTING: This study was conducted at a tertiary care centre. METHOD: A series of 69 newly diagnosed patients, without pre-existing sinus disease, who were treated with high-dose radiotherapy participated. MAIN OUTCOME MEASURES: The prevalence, severity, and time course of mucosal abnormalities were analyzed, as judged by consecutive computed tomographies (CTs). RESULTS: The CT study revealed that 58.8% of the postirradiation scans had mucosal disease of the sinuses. The maxillary sinus had the highest prevalence (42.3%) without statistical significance (p = .10). The difference by McNemar test for two follow-up scans was not significant (p = .48) and by Kappa test was significant (p = .04). The relationship between the prevalence and the time course post radiotherapy revealed that it remained a high prevalence until after the 4-year follow-up scans. CONCLUSIONS: The results of this study confirm that chronic sinus disease is a common late complication of radiotherapy and it persists for years. Thus, aggressive treatment is indicated. PMID- 10709169 TI - Clinical evaluation of a commercially available oral moisturizer in relieving signs and symptoms of xerostomia in postirradiation head and neck cancer patients and patients with Sjogren's syndrome. AB - A major complication of irradiation therapy for head and neck cancer is salivary gland dysfunction and xerostomia. The purpose of this clinical investigation was to evaluate the effects of a commercially available oral moisturizer (Optimoist) on salivary flow rate, symptoms of xerostomia, oral pH, oral microflora, and swallowing in postirradiation head and neck cancer patients (XRT) and patients with Sjogren's syndrome (SS). Subjects who were post-XRT and subjects with SS (n = 24; mean age = 54.1) discontinued their use of any salivary substitute or moisturizer for 2 weeks prior to entering the study. Baseline whole unstimulated saliva was collected for 5 minutes using a standard sialometric technique. Candida albicans and Lactobacillus cultures were performed using kits from Orion Diagnostica, Inc., and a pH analysis was performed on the salivary sample using a Markson (model 00663) pH meter. Swallowing was assessed by clinical measures by videofluoroscopic techniques. Several subjective assessments were performed to evaluate symptoms of xerostomia. Subjects were instructed in the use of a daily diary and to use only the provided article ad libitum for a period of 2 weeks. After the 2-week period, the results indicated significant subjective and objective improvements in signs and symptoms of xerostomia. Whole unstimulated salivary flow rate improved from (mean +/- SEM) 0.1150 +/- 0.02 to 0.2373 +/- 0.09 mL/min. Salivary pH did not change. Global subjective improvement in xerostomia improved in 58% of the subjects. Candida colonization decreased in 43% of the subjects. There was no change in Lactobacilli colonization. Swallowing objectively improved in 75% of subjects. These results indicate significant improvement in both signs of hyposalivation and symptoms of xerostomia with the use of Optimoist in postirradiation head and neck cancer patients and patients with SS. PMID- 10709170 TI - Low-dose intratympanic gentamicin treatment for dizziness in Meniere's disease. AB - OBJECTIVE: Intratympanic gentamicin is used to control dizziness of Meniere's disease, with a low rate of morbidity and a high success rate. We aimed to develop a new technique and schedule of therapy using a lower dose. DESIGN: A retrospective chart review in Meniere's disease patients treated for intractable dizziness. SETTING: A tertiary/quaternary care outpatient setting. METHODS: Patients were administered intratympanic gentamicin using a low-dose protocol on 2 successive days and evaluated with pre- and post-treatment audiovestibular assessment. MAIN OUTCOME MEASURES: Standard evaluation methods of audiovestibular function measured pre- and postfunction of hearing and balance to determine the effects of treatment and morbidity in the form of hearing loss. A telephone follow-up survey was also undertaken. RESULTS: Patients reported satisfactory control of dizziness, with little morbidity in the form of hearing loss. We also found that the use of a myringotomy tube could be precluded. Post-treatment symptoms of imbalance reported by patients settled as patients compensated. In a telephone survey conducted some years after treatment, patient satisfaction was found to be high. CONCLUSIONS: This two-dose regime was shown to be effective in controlling dizzy spells. In patients refractory to the initial two-dose treatment, a follow-up course of treatment usually proved effective. Long-term follow-up of patients seems to show that failure of treatment usually occurs within the first few months, and that symptoms, once controlled, rarely recur. PMID- 10709171 TI - Induction of facial muscle neurotization by temporalis muscle transposition: literature review and animal model evaluation using horseradish peroxidase uptake. AB - OBJECTIVE: To study the concept of facial muscle reinnervation from the trigeminal pathway following facial nerve paralysis. DESIGN AND METHODS: We studied this phenomenon in an animal model using the neuronal marker, horseradish peroxidase (HRP). The temporalis transposition procedure was performed at varying intervals post facial nerve transection. To evaluate the trigeminal-facial reinnervation process and its timing, the zygomaticus major muscle was injected with HRP at varied periods after temporalis transposition, and histologic sections of the brainstem nuclei were examined for the final location of the HRP. RESULTS: The presence of HRP in the trigeminal nucleus provided evidence of trigeminal-facial neurotization in those animals that underwent temporalis transposition up to 2 months following facial denervation and in which the HRP injection was performed 4 months after temporalis transposition. CONCLUSIONS: The findings of our pilot study are strongly supportive of the trigeminal-facial neurotization hypothesis in those animals that underwent temporalis transposition up to 2 months post facial denervation and in which 4 months were allowed thereafter for adequate neurite ingrowth and neurotization to occur. This suggests that the neurotrophic signals are greatest up to 2 months post denervation and denotes the optimal time for performance of reconstructive procedures. Future studies with a larger number of animals in each group will be necessary to ensure more potent statistical significance and to augment our experimental evidence that trigeminal-facial crossover does occur and can be used as an adjunctive concept to maximize early rehabilitation of the paralyzed face. PMID- 10709172 TI - Focal myositis in the mylohyoid muscle of an adult. PMID- 10709173 TI - Triple primary malignancies of the head and neck. PMID- 10709174 TI - Leiomyoma of the inferior turbinates. PMID- 10709175 TI - Suprastomal puncture for voice restoration after laryngectomy. PMID- 10709176 TI - Shy-Drager syndrome: an otolaryngology perspective. PMID- 10709177 TI - Golden earth against cough: a 14th-century instruction for inhalation. PMID- 10709178 TI - Sentence comprehension is mediated by content-addressable memory structures. AB - Studies of working memory demonstrate that some forms of information are retrieved by a content-addressable mechanism (McElree & Dosher, 1989; McElree, 1996, 1998), whereas others require a slower search-based mechanism (McElree & Dosher, 1993). Measures of the speed and accuracy of processing sentences with filler-gap dependencies demonstrate that the probability of maintaining a representation of a filler item decreases as additional material is processed, but that the speed with which a preserved representation is accessed is unaffected by the amount of interpolated material. These results suggest that basic binding operations in sentence comprehension are mediated by a content addressable memory system. PMID- 10709179 TI - On bound variable interpretations: the LF-only hypothesis. AB - Under what conditions do perceivers prefer to assign a bound variable interpretation to a pronominal that is ambiguous between a bound variable and a coreferential interpretation? Several experiments were designed to test the hypothesis that language perceivers prefer a bound variable over a coreferential interpretation of a pronoun because the former only requires consultation of a logical form (LF) representation, while the latter requires access to a discourse representation. The hypothesis was disconfirmed in two respects. First, although bound variable interpretations show a processing advantage over coreferential interpretations in VP ellipsis constructions, the preference for bound variable interpretations is not general--it does not extend to other quantificational contexts. Second, the preference for bound variable interpretations in VP ellipsis constructions is not limited to examples in which the antecedent and the ellipsis site occur in the same sentence. If the bound variable advantage were due to the ready availability of the LF for the current sentence, the advantage should disappear across sentence boundaries. An alternative hypothesis is then considered which could explain the source of the bound variable advantage in VP ellipsis contexts. PMID- 10709180 TI - Brain potentials in the processing of complex sentences: an ERP study of control and raising constructions. AB - In the present study we made use of Event-Related Potentials (ERPs) to examine raising and subject control constructions in German. Our most salient result is that the ERPs elicited at the empty subject position of a raising construction are clearly different from those elicited at the corresponding position of an otherwise identical subject control construction, the former producing a stronger P600. We argue that this result provides an electrophysiological correlate of the theoretical distinction between NP trace and PRO. PMID- 10709181 TI - The temporal unfolding of local acoustic information and sentence context. AB - The purpose of this study was to investigate the temporal unfolding of local acoustic information and sentence context using both cross-modal interference (CMI) and word-monitoring tasks. The timing of sentence context effects have important theoretical implications for models of language processing (e.g., initial context independence vs. initial interaction). Yet, different tasks tend to yield different results. For both experiments, stimuli from an acoustically manipulated "goat-to-coat" continuum were embedded in sentences whose interpretation was biased toward either "goat" or "coat." In experiment 1 (CMI), the primary task was listening to sentences for comprehension; the interference task was a word/nonword decision to an unrelated visual probe that appeared at one of three positions within the sentence. Results showed immediate effects of the acoustic manipulation, but only delayed effects of sentence context. These results were interpreted to indicate that phonological processing is initially context-independent but is followed by rapid context integration. Experiment 2 used a word-monitoring task: Response times were significantly longer when sentence context was incongruent with the monitoring target, showing an immediate effect of context. The apparently contradictory results of the two experiments together support an account of language processing in which phoneme categorization is initially independent of sentence context unless an explicit judgment about the identity of the target is required. PMID- 10709182 TI - Intonational disambiguation in sentence production and comprehension. AB - Speakers' prosodic marking of syntactic constituency is often measured in sentence reading tasks that lack realistic situational constraints on speaking. Results from such studies can be criticized because the pragmatic goals of readers differ dramatically from those of speakers in typical conversation. On the other hand, recordings of unscripted speech do not readily yield the carefully controlled contrasts required for many research purposes. Our research employs a cooperative game task, in which two speakers use utterances from a predetermined set to negotiate moves around gameboards. Results from a set of early versus late closure ambiguities suggest that speakers signal this syntactic difference with prosody even when the utterance context fully disambiguates the structure. Phonetic and phonological analyses show reliable prosodic disambiguation in speakers' productions; results of a comprehension task indicate that listeners can successfully use prosodic cues to categorize syntactically ambiguous fragments as portions of early or late closure utterances. PMID- 10709183 TI - Search strategies in syntactic reanalysis. AB - We first consider the nature of syntactic reanalysis, paying particular attention to questions about whether and how it differs from the construction of an initial analysis, such as whether it is encapsulated or whether alternatives are considered in parallel or not. We then outline different strategies that the processor may use in reanalysis, and review the relevant evidence. We show that our experimental evidence is compatible with an encapsulated serial search strategy. PMID- 10709184 TI - The concomitant effects of phrase length and informational content in sentence comprehension. AB - Recent evidence suggests that phrase length plays a crucial role in modification ambiguities. Using a self-paced reading task, we extended these results by examining the additional pragmatic effects that length manipulations may exert. The results demonstrate that length not only modulates modification preferences directly, but that it also necessarily changes the informational content of a sentence, which itself affects modification preferences. Our findings suggest that the same length manipulation affects multiple sources of constraints, both structural and pragmatic, which can each exert differing effects on processing. PMID- 10709185 TI - Activation of syntactic information during language production. AB - In order to produce utterances, people must draw upon syntactic information. This paper considers how evidence from syntactic priming experiments casts light upon the nature of syntactic information activation during language production. We examine three issues: the way in which syntactic information is initially activated, the circumstances under which activation may persist or dissipate, and the effects of residual activation of syntactic information on subsequent language production. Evidence from dialog experiments suggests that the information that is initially activated is the same in both production and comprehension. Evidence about the persistence of activation following initial activation is more complex. We suggest that persistence may be related to the potential relevance of the information for subsequent syntactic processing. We show that current evidence is inconclusive about how long syntactic information remains activated. PMID- 10709186 TI - Structural priming as implicit learning: a comparison of models of sentence production. AB - Structural priming reflects a tendency to generalize recently spoken or heard syntactic structures to different utterances. We propose that it is a form of implicit learning. To explore this hypothesis, we developed and tested a connectionist model of language production that incorporated mechanisms previously used to simulate implicit learning. In the model, the mechanism that learned to produce structured sequences of phrases from messages also exhibited structural priming. The ability of the model to account for structural priming depended on representational assumptions about the nature of messages and the relationship between comprehension and production. Modeling experiments showed that comprehension-based representations were important for the model's generalizations in production and that nonatomic message representations allowed a better fit to existing data on structural priming than traditional thematic role representations. PMID- 10709188 TI - Falsifying serial and parallel parsing models: empirical conundrums and an overlooked paradigm. AB - When the human parser encounters a local structural ambiguity, are multiple structures pursued (parallel or breadth-first parsing), or just a single preferred structure (serial or depth-first parsing)? This note discusses four important classes of serial and parallel models: simple limited parallel, ranked limited parallel, deterministic serial with reanalysis, and probabilistic serial with reanalysis. It is argued that existing evidence is compatible only with probabilistic serial-reanalysis models, or ranked parallel models augmented with a reanalysis component. A new class of linguistic structures is introduced on which the behavior of serial and parallel parsers diverge the most radically: multiple local ambiguities are stacked to increase the number of viable alternatives in the ambiguous region from two to eight structures. This paradigm may provide the strongest test yet for parallel models. PMID- 10709187 TI - Distinguishing serial and parallel parsing. AB - This paper discusses ways of determining whether the human parser is serial maintaining at most, one structural interpretation at each parse state, or whether it is parallel, maintaining more than one structural interpretation in at least some circumstances. We make four points. The first two counterclaims made by Lewis (2000): (1) that the availability of alternative structures should not vary as a function of the disambiguating material in some ranked parallel models; and (2) that parallel models predict a slow down during the ambiguous region for more syntactically ambiguous structures. Our other points concern potential methods for seeking experimental evidence relevant to the serial/parallel question. We discuss effects of the plausibility of a secondary structure in the ambiguous region (Pearlmutter & Mendelsohn, 1999) and suggest examining the distribution of reaction times in the disambiguating region. PMID- 10709189 TI - [The historical unifying congress of anatomists, histologists and embryologists of the Soviet Union (on the 50th anniversary of the formation of the All-Union Scientific Society of Anatomists, Histologists and Embryologists)]. PMID- 10709190 TI - [Changes in the enzyme activity in the brain structures of August rats under the influence of the delta sleep-inducing peptide against a background of prolonged L dihydroxyphenylalanine administration]. AB - Quantitative cytochemical methods revealed the decrease of MAO activity (substrate--tryptamine) in the hippocampus of L-dioxytryptamine treated August rats genetically predisposed to emotional stress under the effect of delta sleep inducing peptide (DSIP). Activities of aminopeptidase and glutamate dehydrogenase were decreased in n.accumbens while changes of the activities of these enzymes were not significant in the layers III and V of sensomotor cortex and n.caudatus. In all brain structures Ache and MAO (substrate 5-hydroxytryptamine) activities were not influenced by DSIP. PMID- 10709191 TI - [The reaction of the population of mitochondria in the sensorimotor cortical neurons of rats to the prolonged, continuous action of low-frequency vibration]. AB - The investigations revealed that long term action of vibration causes the following changes: growth of mitochondria size (the appearance of hypertrophied ones) clearing of their matrix, vacuoles formation, destruction of cysts. Similar changes were described in extreme conditions connected with deficiency of energy supply. Reaction of mitochondrial population to the action of vibration is phased and lies in change of relative content of mitochondria with different functional activity (orthodoxal, activated and destroying). General direction of these reorganization is a creation of stable and reactive functional subcellular system. PMID- 10709192 TI - [The modular organization of the granular layer of the human cerebellar cortex in postnatal ontogeny]. AB - Age changes of microstructure of granular layer of the cortex of human cerebellum superior lobe (the area of clivus, posterior lateral regions of right and left tetrangular lobules) were studied in annual intervals from birth up to 20 yrs using computer analysis of optic images. Enlargement of the area of profile fields of cell groups in apical and basal regions of cerebellar folia from birth up to 14-15 yrs and differences between hemispheres in clasterization of granular layer were demonstrated. The data on age changes of cellular aggregations obtained allowed to formulate a hypothesis on autonomic local sensory networks of human cerebellar cortex associated with granular layer of cerebellar cortex that are considered as structural elements of module organization of the latter. PMID- 10709194 TI - [The structure of the initial inputs into the metasympathetic nervous system of the rat uterus]. AB - Different populations of sympathetic neurons exerting modulating influence on neurons of nervous plexuses of proper metasympathetic nervous system of the uterus in albino laboratory rats were detected using the method on retrograde transport of fluorescent marker primulin. Following the injection of the marker into uterovaginal plexus, labelled neurons were found as aggregations in caudal mesenterial sympathetic ganglia, ganglia of coeliac plexus, renal ganglia and ganglia of coeliac trunk. The structure of nervous paths of external control of uterus functioning was analysed. PMID- 10709193 TI - [Neuronal changes in the dorsal motor nucleus of the vagus nerve during intervals of hypoxic exposures in animals with diabetes mellitus]. AB - Changes of neurons of dorsal motor nucleus of nervus vagus were studied in adaptation to hypoxia, experimental diabetes mellitus and its correction by means of interrupted hypoxic effects. It was established previously that interrupted hypoxic training exerted stimulating effect on insulin synthesizing function of pancreas. As a result of the present study the increase of morphofunctional activity of neurons was found in all experimental series although it was greater manifested in animals with experimental diabetes mellitus who were subjected to actions of hypoxia. The changes of morphofunctional activity of dorsal motor nucleus of nervus vagus established allow to conclude on the significant role these structure plays in realization of stimulating effect of interrupted actions of hypoxia on the state of insulin synthesizing function of pancreas and clinical characteristics of the experimental diabetes mellitus. PMID- 10709195 TI - [The structural organization of the sensory system of the statocysts in nudibranch mollusks (Coryphella rufibranchialis)]. AB - Using silver nitrate impregnation after Golgi and Colognier, staining with methylen blue and HRP labelling organization of peripheral and central regions of statocyst sensory system in nudibranch mollusc Coryphella rufibranchialis was investigated. Ramification of processes of the majority of statocyst receptor cells in its wall, adjacent optic and both cerebropleural ganglia was demonstrated. Special attention was drawn to structural bases of possible intersensory interactions between statocyst. Common zone of distribution of afferent fibres from statocysts, eyes, rhinophores and neuron processes of optic ganglia were distinguished and neurons innervating these zones and pedal ganglia were discovered. PMID- 10709196 TI - [The age-related characteristics of the uterine arteries]. AB - The investigation was performed in 120 uterus preparations of the 16-80-yrs women without pathology of the internal sexual organs. A complex of morphological and morphometrical methods was used. The changes of the uterine wall vascularization, connected with ageing and with the functional reconstruction of arterial bed of parous women were described. PMID- 10709197 TI - [The artificial lymph node]. AB - Adsorption of lymphocytes on the sorbents in correction of inflammatory lesions has significant clinical perspectives as the cells and active biological substances they secrete function actively in inflamed tissues acting in the site of its application: every cell adsorbs and inactivates foreign substances, every cytokin affects certain target cells. The products of these cells functioning and dissociation as well as the products of immune system reaction to them remain on the sorbent and are eliminated together with granules during bandaging or by the end of the treatment thus do not entering the organism. Lymphocytic monolayer on the sorbent does not prevent its specific action in pathologic nidus. In this case sorbent does not only drain the tissue duplicating certain primitive non specific functions of regional lymph node (barrier, filtrating, transport, draining and protective functions) but also performs certain highly specialized functions of immunocompetent organs--selective adsorption and inactivation of antigenic substances. It is reasonable to use sorbents for adsorbing lymphocytes in kapron containers which exclude the preparation leak into wounds and cavities and act as a filter preventing fast inactivation of the preparation by large fragments of tissue detritus. PMID- 10709198 TI - [The development of the lymphatic vessel wall in the early period of swine postnatal ontogeny]. AB - Using histological methods and transmission electron microscopy differences in architecture of lymphatic vessel wall between newborn and 2 months old pigs were established. The number of free ribosomes and polysomes is smaller and that of pinocytotic vesicles is higher in 2 months pigs. Contacts between endotheliocytes are presented in three types. Smooth myocytes are specified for contracting: the majority of cytoplasm contains microfilament bundles, dense bodies, great number of pinocytotic vesicles are present. In lymphatics external coat collagen fibril bundles are thicker than in that of newborns and the number of nerve fibres is higher. PMID- 10709199 TI - [Remodeling of the cortical layer of the tibia after osteotomy of the femur in the same extremity]. AB - Chronobiological characteristics of temporal [correction of temper]-dimensional organization of compact bone remodelling in right tibia after the femoral diaphysis osteotomy on the same side were studied using roentgenometry and roentgenodensitometry. These characteristics were studied in experiments on 179 outbred male albino rats. Circaseptan periodicity of osseous tissue reorganization was established. PMID- 10709200 TI - [The morphological dynamics and correlation-regression functions of the morphometric indices of the lesion and regeneration in an experimental embolic bone infarct]. AB - Histological changes and dynamics of morphometric parameters of ischemic lesion, organization and regeneration of bone marrow and osseous tissue were studied experimentally in rabbits with embolic aseptic infarction of long bones (on d 3 60). Correlation dependences were determined and regression equations were derived between morphometric parameters of lesion and regeneration of bone tissues. Close or medium connections were demonstrated between volume of ischemic necrosis of bone marrow and compact substance (exponential regression), those of compact substance and periosteal regenerate (exponential regression), volume of necrosis of compact substance and manifestation of reorganization (power regression), volume of periosteal regenerate and manifestation of reorganization of compact substance (rectilinear regression). PMID- 10709201 TI - [The morphofunctional changes in the Langerhans cells of the human epidermis in inflammatory and cancerous diseases]. AB - Using ATPase reaction Langerhans cells were studied in human epidermis obtained from healthy volunteers, corpses of people who died from trauma and were autopsied during first 3 postmortem hrs and patients operated for inflammatory and oncological diseases of different localization. The extent of Langerhans cells alteration was shown to correlate directly with the severeness of the disease the patients has been operated for. Least changes of Langerhans cells developed in people operated for hernia of the anterior abdominal wall while most pronounced changes were found in those operated for stomach cancer of IV stage. The studying of Langerhans cells may be used in clinical practice for evaluation of efficiency of the treatment and in prognostic purposes. PMID- 10709202 TI - [Thymus changes in chronic heliotrine-induced hepatitis]. AB - The structural and morphometrical characteristics of thymus were studied in the dynamics of experimental heliotrine hepatitis. The decrease of areas and cell hypoplasia of cortical zone of the thymus were noted. The destructive processes of the thymocytes were intensified and, in response to it, thymic macrophages activity was enhanced. The facts obtained indicated that the use of the thymic peptides is necessary for correction of immune disorders in chronic hepatitis. PMID- 10709203 TI - [The structure of the white pulp of the spleen and the peripheral blood indices in rats under increased muscle activity]. AB - The white pulp structure was studied in rat spleen white pulp in conditions of increased muscle activity as well as certain parameters of peripheral blood. 36 male outbred albino rats were involved. Experimental rats were subjected to the increased physical load (everyday swimming seances lasting from 10 to 40 hrs during 4 weeks). The number of erythrocytes, leukocytes and level of hemoglobin were determined in peripheral blood. Perimeter squares and density of lymphoid nodules (LN) and marginal zone (MZ) as well as LN minimal and maximal diameter and MZ minimal and maximal width were determined in histological sections prepared at the level of the organ portae using TV-computer set. Besides, marginal sinuses width were measured in nine sites in every LN as well as the capsule thickness in three sites and at the level of portae. The data obtained indicate that increased muscle activity causes reorganization of red and white pulp in the direction of growth of the red pulp square which is associated with the increase of erythropoietic function. The appearance of lymphoid nodules with germination centres indicates lymphocytopoietic function intensification while the significant increase of marginal zone morphometric parameters witnesses its participant in primary immune response. High correlation between certain blood indexes and the parameters of lymphoid nodules and marginal zones supports these data. PMID- 10709204 TI - [Lung tissue changes in experimentally induced inflammation and its treatment]. AB - As a result of the investigation of experimental models of pneumonia, three stages of its development and peculiarities of epitheliostromal interrelations in walls of bronchi and in lung respiratory region characteristic for every stage were established. Administration of benzonal as an inductor of the surfactant system along with polyunsaturated fatty acids exerted stimulating effect on the adaptive processes at acute and subacute stages in pneumonia. The number of macrophages increased, their function enhanced, which, along with type II alveolocytes proliferation indicated the activation of regenerative processes. PMID- 10709205 TI - [Changes to the hepatocytes and muscle fibers of the diaphragm during adaptation to cold under normal conditions and in combination with high-altitude hypoxia]. AB - Comparative morphometric analysis of muscular fibres of the diaphragm and hepatocytes of albino rats was carried out in adaptation to cold (0-2 degrees C for 42 d) and to combined effect of cold and high-mountain hypoxia (2-5 degrees C temperature, 4000 m height, 40 d duration of the experiment). It was demonstrated that in adaptation to cold in normobaric conditions muscular fibres of the diaphragm develop moderate hypertrophy of mitochondria, accumulation of lipid inclusions with glycogen content remaining close control level. In combined effect of cold and high-mountain hypoxia direction of these changes was remained while their manifestation was increased. Relative volume of mitochondria was increased in hepatocytes in both experiments while volumetric indexes of rough endoplasmic reticulum and glycogen grew smaller. On one hand, the peculiarities of the dynamics of quantitative parameters of diaphragm muscular fibres and hepatocytes revealed in conditions of the long-term effect of the cold and in its combination with high mountain hypoxia indicate different efficiency of adaptive processes in the organ depending on the character of the effect and their organ specificity on the other. PMID- 10709206 TI - [Modern tends in the alteration of human spermatogenic activity]. PMID- 10709207 TI - [Experience with the use of testing control over student knowledge of human anatomy]. PMID- 10709208 TI - [The individualization of student instruction in a histology department at the current stage of college development]. PMID- 10709209 TI - Long-lasting effects of GABA infusion into the cerebral cortex of the rat. AB - In electrophysiological terms, experimental models of durable information storage in the brain include long-term potentiation (LTP), long-term depression, and kindling. Protein synthesis correlates with these enduring processes. We propose a fourth example of long-lasting information storage in the brain, which we call the GABA-withdrawal syndrome (GWS). In rats, withdrawal of a chronic intracortical infusion of GABA, a ubiquitous inhibitory neurotransmitter, induced epileptogenesis at the infusion site. This overt GWS lasted for days. Anisomycin, a protein synthesis inhibitor, prevented the appearance of GWS in vivo. Hippocampal and neocortical slices showed a similar post-GABA hyperexcitability in vitro and an enhanced susceptibility to LTP induction. One to four months after the epileptic behavior disappeared, systemic administration of a subconvulsant dose of pentylenetetrazol produced the reappearance of paroxysmal activity. The long-lasting effects of tonic GABAA receptor stimulation may be involved in long-term information storage processes at the cortical level, whereas the cessation of GABAA receptor stimulation may be involved in chronic pathological conditions, such as epilepsy. Furthermore, we propose that GWS may represent a common key factor in the addiction to GABAergic agents (for example, barbiturates, benzodiazepines, and ethanol). GWS represents a novel form of neurono-glial plasticity. The mechanisms of this phenomenon remain to be understood. PMID- 10709210 TI - The GABA-withdrawal syndrome: a model of local status epilepticus. AB - The GABA-withdrawal syndrome (GWS) is a model of local status epilepticus following the interruption of a chronic GABA infusion into the rat somatomotor cortex. GWS is characterized by focal epileptic electroencephalographic discharges and associated contralateral myoclonus. In neocortical slices obtained from GWS rats, most neurons recorded in the GABA-infused area are pyramidal neurons presenting bursting properties. The bursts are induced by white-matter stimulation and/or intracellular depolarizing current injection and correlate with a decrease of cellular sensitivity to GABA, caused by its prolonged infusion. This effect is related to a calcium influx that may reduce the GABAA receptor-mediated inward current and is responsible for the bursting properties. Here we present evidence for the involvement of calcium- and NMDA-induced currents in burst genesis. We also report modulatory effects of noradrenaline appearing as changes on firing patterns of bursting and nonbursting cells. Complementary histochemical data reveal the existence of a local noradrenergic hyperinnervation and an ectopic expression of tyrosine hydroxylase mRNAs in the epileptic zone. PMID- 10709211 TI - GABA and muscimol as reversible inactivation tools in learning and memory. AB - Reversible inactivation of brain areas is a useful method for inferring brain behavior relationships. Infusion of GABA or of the GABA receptor agonist muscimol is considered one interesting reversible inactivation method because it may not affect fibers of passage and may therefore be compared to axon-sparing types of lesions. This article reviews the data obtained with this method in learning and memory experiments. A critical analysis of data, collected in collaboration with Simon Brailowsky, with chronic GABA infusion is presented, together with an illustration of data obtained with muscimol-induced inactivation. PMID- 10709212 TI - GABA withdrawal modifies network activity in cultured hippocampal neurons. AB - Dissociated hippocampal neurons, grown in culture for 2 to 3 weeks, tended to fire bursts of synaptic currents at fairly regular intervals, representing network activity. A brief exposure of cultured neurons to GABA caused a total suppression of the spontaneous network activity. Following a washout of GABA, the activity was no longer clustered in bursts and instead, the cells fired at a high rate tonic manner. The effect of removing GABA could be seen as long as 1 to 2 days after GABA withdrawal and is expressed as an increase in the number of active cells in a network, as well as in their firing rates. Such striking effects of GABA removal may underlie part of the GABA withdrawal syndrome seen elsewhere. PMID- 10709213 TI - Myoclonia in Papio papio: are they all "epileptic"? PMID- 10709214 TI - Functional and anatomic correlates of two frequently observed temporal lobe seizure-onset patterns. AB - Intracranial depth electrode EEG records of 478 seizures, recorded in 68 patients undergoing diagnostic monitoring with depth electrodes, were evaluated to investigate the correlates of electrographic onset patterns in patients with temporal lobe seizures. The seizure onsets in 78% of these patients were identified as either hypersynchronous onsets, beginning with low-frequency, high amplitude spikes, or low-voltage fast (LVF) onsets, increasing in amplitude as the seizure progressed. The number of patients (35) having hypersynchronous seizure onsets was nearly twice that of patients (18) having LVF onsets. Three major differences were seen among patients with the two seizure-onset patterns. When compared with patients having LVF onsets, patients with hypersynchronous seizure onsets had a significantly greater probability of having (1) focal rather than regional seizure onsets (p < 0.01), (2) seizures spreading more slowly to the contralateral mesial temporal lobe (p < 0.003), and (3) cell counts in resected hippocampal tissue showing greater neuronal loss (p < 0.001). The results provide evidence that the most frequent electrographic abnormality associated with mesial temporal seizures is local hypersynchrony, a condition associated with major neuronal loss in the hippocampus. The results also indicate that LVF seizure onsets more frequently represent widely distributed discharges, which interact with and spread more rapidly to surrounding neocortical areas. PMID- 10709215 TI - Parallel information processing in motor systems: intracerebral recordings of readiness potential and CNV in human subjects. AB - We performed intracerebral recordings of Readiness Potential (RP) and Contingent Negative Variation (CNV) with simple repetitive distal limb movement in candidates for epilepsy surgery. In 26 patients (in Paris), depth electrodes were located in various cortical structures; in eight patients (in Brno), in the basal ganglia and the cortex. RPs were displayed in the contralateral primary motor cortex, contralateral somato-sensory cortex, and bilaterally in the SMA and the caudal part of the anterior cingulate cortices. CNVs were recorded in the same cortical regions as the RP, as well as in the ipsilateral primary motor cortex, and bilaterally in the premotor fronto-lateral, parietal superior, and middle temporal regions. In the basal ganglia, the RP was recorded in the putamen in six of seven patients, and in the head of the caudate nucleus and the pallidum in the only patient with electrodes in these recording sites. We suggest that our results are consistent with a long-lasting, simultaneous activation of cortical and subcortical structures, before and during self-paced and stimulus-triggered movements. The particular regions that are simultaneously active may be determined by the task context. PMID- 10709216 TI - Effects of the novel NMDA receptor antagonist gacyclidine on recovery from medial frontal cortex contusion injury in rats. AB - Gacyclidine, a novel, noncompetitive NMDA receptor antagonist, was injected (i.v.) into rats at three different doses to determine if the drug could promote behavioral recovery and reduce the behavioral and anatomical impairments that occur after bilateral contusions of the medial frontal cortex (MFC). In the Morris water maze, contused rats treated with gacyclidine at a dosage of 0.1 mg/kg performed better than their vehicle-treated conspecifics. Rats given gacyclidine at either 0.3 or 0.03 mg/kg performed better than brain-injured controls, but not as well as those treated with 0.1 mg/kg. Counts of surviving neurons in the nucleus basalis magnocellularis (NBM) and the medial dorsal nucleus (MDN) of the thalamus were used to determine whether gacyclidine treatment attenuated secondary cell death. In both the NBM and the MDN, the counts revealed fewer surviving neurons in untreated contused rats than in gacyclidine-treated rats. Increases in the size and number of microglia and astrocytes were observed in the striatum of gacyclidine-treated contused brains. Although most consequences of MFC contusions were attenuated, we still observed increases in ventricle dilation and thinning of the cortex. In fact, the ventricles of rats treated with 0.1 mg/kg of gacyclidine were larger than those of their vehicle treated counterparts, although we observed no behavioral impairment. PMID- 10709217 TI - Intracerebral transplants and memory dysfunction: circuitry repair or functional level setting? AB - Intracerebral grafting techniques of fetal neural cells have been used essentially with two main types of lesion paradigms, namely damage to long projection systems, in which the source and the target are clearly separate, and damage to neurons that are involved in local circuits within a small (sub)region of the brain. With the first lesion paradigm, grafts placed homotopically (in the source) are not appropriate because their fibers grow poorly through the host parenchyma and fail to reach their normal target. To be successful, the grafts must be placed ectopically in the target region of the damaged projection systems, where generally they work as level-setting systems. Conversely, with the second paradigm, the grafts are supposed to compensate for a local loss of neurons and must be placed homotopically to induce functional effects that are based on the reconstruction of a point-to-point circuitry. By inserting a biological or artificial bridging-substrate between the source and the target of long projection systems, it might be possible to combine the positive effects of both homotopic and ectopic grafting by achieving both target reinnervation and normal control of the grafted neurons within the source area. These issues are illustrated and discussed in this review. PMID- 10709218 TI - Alleviation of brain injury-induced cerebral metabolic depression by amphetamine: a cytochrome oxidase histochemistry study. AB - Measurements of oxidative metabolic capacity following the ablation of rat sensorimotor cortex and the administration of amphetamine were examined to determine their effects on the metabolic dysfunction that follows brain injury. Twenty-four hours after surgery, rats sustaining either sham operations or unilateral cortical ablation were administered a single injection of D amphetamine (2 mg/kg; i.p.) or saline and then sacrificed 24 h later. Brain tissue was processed for cytochrome oxidase histochemistry, and 12 bilateral cerebral areas were measured, using optical density as an index of the relative amounts of the enzyme. Compared with that of the control groups, cytochrome oxidase in the injured animals was significantly reduced throughout the cerebral cortex and in 5 of 11 subcortical structures. This injury-induced depression of oxidative capacity was most pronounced in regions of the hemisphere ipsilateral to the ablation. Animals given D-amphetamine had less depression of oxidative capacity, which was most pronounced bilaterally in the cerebral cortex, red nucleus, and superior colliculus; and in the nucleus accumbens, caudateputamen, and globus pallidus ipsilateral to the ablation. The ability of D-amphetamine to alleviate depressed cerebral oxidative metabolism following cortical injury may be one mechanism by which drugs increasing noradrenaline release accelerate functional recovery in both animals and humans. PMID- 10709219 TI - Neural mechanisms of attention: the northern California years. PMID- 10709221 TI - Silent atrial fibrillation--another Pandora's box. PMID- 10709220 TI - Through the looking glass: differential noradenergic modulation of prefrontal cortical function. PMID- 10709222 TI - Proarrhythmic effects of ibutilide in a canine model of pacing induced cardiomyopathy. AB - The authors developed a canine model of pacing induced cardiomyopathy to study the possible mechanisms of ibutilide induced torsades de pointes (TP) in heart failure. Thirteen dogs received intravenous ibutilide after acute AV block for 60 minutes, and after implantation of a VVI pacemaker, with a rate of 270 beats/min for 2-3 weeks. Twelve-lead ECG and right and left ventricle monophasic action potentials were recorded at different right ventricle pacing cycle lengths from 600 ms to 1200 ms during the study. The results showed ibutilide could significantly prolong ventricular repolarization and increase the dispersion in a dose dependent and reverse use dependent manner. Furthermore, after ibutilide administration, cardiomyopathic dogs had a greater dispersion of ventricular repolarization, and also had higher incidences of early afterdepolarizations and spontaneous or pacing induced TP than acute AV block dogs. PMID- 10709223 TI - Consistency of multicenter measurements of heart rate variability in survivors of acute myocardial infarction. AB - Heart rate variability (HRV) analysis from 24-hour ambulatory ECG has been widely used in risk stratification of patients after myocardial infarction (MI). The accuracy of HRV assessment is known to potentially vary when different commercial systems are used. However, the consistency of HRV measurements has never been fully investigated. Twenty-six post-MI patients (mean age 59 +/- 8 years, 22 men) were studied, of whom 13 succumbed to sudden cardiac death (SCD) within 1 year and 13 remained alive for at least 3 years (MI survivors). Each patient had a 24 hour Holter ECG recorded before hospital discharge. HRV analysis was performed four times from the same recordings using three different Holter tape analysis systems (Marquette, Reynolds, and CardioData) by four independent operators (CardioData system was used twice, once in the United Kingdom and once in the United States). Mean normal-to-normal RR intervals (mNN) and 3 HRV parameters (SDNN, RMSSD, and HRV triangular index [HRVi]) were derived from each recording. The consistency of mNN and HRV measurements was evaluated by coefficient of variance (CV) and by the Bland-Altman method. The results demonstrated that (1) all indices measured by different systems were statistically similar (P = NS) except the measurement of RMSSD (P = 0.01), (2) the measurements of mNN were highly reproducible with a maximum mean difference of 1.8 +/- 13.8 ms and maximum limits of agreement from -14.6 to +15.6 ms. The maximum mean differences were- 1.8 +/- 1.4 unit and 4.4 +/- 9.6 ms for HRVi and SDNN, respectively, and RMSSD was less reproducible with a maximum mean difference of--11.1 +/- 11.5 ms, and limits of agreement from -16.2 to +9.6 ms; and (3) the consistency of mNN (CV 0.9% +/- 0.9%) was significantly higher than that of HRVi, SDNN, and RMSSD (P < 0.0001). The consistency of HRVi was similar to that of SDNN (4.8% +/- 2.1% vs 5.7% +/- 4.8%, P = 0.4), and the consistency of RMSSD (26.6% +/- 13.3%) was significantly lower than that of the other measurements (P < 0.00001). In conclusion, the measurements of mNN by different analytical systems are the most consistent among the parameters studied. The global 24-hour measurements of HRV (SDNN and HRVi) are highly reproducible, whereas the measurement of short-term HRV components (RMSSD) is significantly less reproducible. PMID- 10709224 TI - How to program rate responsive pacemakers. AB - Because oxygen uptake (VO2) increases linearly with heart rate during exercise, the oxygen pulse reserve (OPR) method (VO2 reserve divided by heart rate reserve) may provide a valid guide for rate responsive parameter tailoring. Using custom made software (Pacing Rate Profile Software [PRPS]) it is possible to predict the exercise pacing rate profile with significant accuracy, according to the patient's functional class when ergospirometry apparatus is not available for a cardiopulmonary stress test (CPX). PRPS for Windows is based on the OPR method and some known workload/metabolic cost of exercise relationships during effort. The present study had two aims; first, to evaluate the reliability of PRPS in accurately predicting pacing rate profiles; and second, the suitability of activity and metabolic rate responsive sensors in supplying pacing rates sufficiently near to those predicted using CPX or PRPS. To test the reliability of PRPS we studied 244 patients, NYHA Class I-II, under two different stress test protocols. In one, the bicycle protocol (25 W, 2-minute steps), we tested 137 normal patients (94 men and 43 women, mean age 67 +/- 15 years). Sixty-eight of these were simultaneously CPX tested. PRPS predicted pacing rates were matched against the patients' sinus rhythms or their theoretical CPX measured VO2 heart rates (OPR method). Linear regression analysis was highly significant (r = 0.93 and r = 0.97, respectively). The other, the treadmill protocol, consisted of three different protocols. (1) Speed Incremental Treadmill Stress Test (SITST): 57 patients underwent CPX (33 men and 24 women, mean age 67 +/- 15 years, NYHA Class I-II). All had been pacemaker implanted for SSS and/or advanced atrioventricular block (AVB). PRPS pacing rates were matched against CPX VO2 OPR calculated heart rates (r = 0.93), (linear regression analysis). (2) CAEP: 30 patients underwent CPX (26 men and 4 women, mean age 61 +/- 11 years, NYHA Class I-II). Thirteen of them had been pacemaker implanted for SSS and/or advanced AVB. In all 30 patients the PRPS rates were matched against CPX VO2 calculated rates (r = 0.90). In the 17 normal nonimplanted patients, the PRPS rates were also matched against sinus rhythms, (r = 0.80). (3) Weber: 20 patients underwent CPX (16 men and 4 women, mean age 68 +/- 8 years, NYHA Class I-II). As above, in six normal nonimplanted patients, statistical analysis between PRPS rates and sinus rhythms was performed (r = 0.89). The comparison between PRPS theoretical pacing rates and VO2 predicted rates in all 20 patients was also statistically significant (r = 0.93). Finally, to test the reliability of PRPS also in NYHA Class III-IV patients, we tested 22 implanted patients (15 men and 7 women, mean age 70 +/- 9 years) and compared PRPS predicted rates against VO2 CPX measured rates (r = 0.92). To determine if the wide variety of RR pacers were able to supply pacing rates near to those predicted, whether by means of CPX or PRPS, we studied a total of 89 patients: 49 of these (26 men and 23 women, mean age 66 +/- 12 years) had been implanted with activity sensors; 12 patients (11 men and one woman, mean age 70 +/- 7 years) had been implanted with metabolic sensors, and finally 28 patients (19 men and 9 women, mean age 70 +/- 12 years) had been implanted with dual sensors (activity + QT or minute ventilation). Linear regression analysis showed r = 0.93 for activity sensors, r = 0.94 for metabolic sensors, and r = 0.92 for dual sensor. In conclusion, when rate responsive pacing causes symptoms or functional impairment, physicians must provide a personalized rate response tailoring derived from precise, simple physiological testing. OPR is an easy physiological method for tailoring rate response settings, suitable for activity and metabolic sensors. When ergospirometry apparatus is not available, PRPS can successfully replace CPX testing for tailoring. PMID- 10709225 TI - Importance of AV synchronous pacing during low intensity exercise evaluated by oxygen kinetics. AB - It has been shown that dual chamber pacing with preservation of AV synchrony (DDD) is superior to fixed rate ventricular (VVI) or rate responsive ventricular (VVIR) pacing modes, as evaluated by ventilatory response to exercise. Previous studies have focused on the benefits of maintained AV synchrony at maximal exercise. However, there are limited data comparing O2 kinetics in different pacing modes during low intensity exercise, representing the majority of daily activities. This study aimed to provide an evaluation of different pacing modes using O2 kinetics during low intensity exercise. Nineteen patients (age 61 +/- 18 years) with complete AV block underwent low intensity treadmill exercise (35 W) with simultaneous evaluation of symptoms and O2 kinetics in three pacing modes. The first test was performed in DDD mode followed by a second test in VVIR mode with a programmed heart rate corresponding to the sinus rate during the first test. After 6 minutes of each test, the mode was switched from DDD to VVIR and vice versa. The third test was performed in VVI mode at 70 beats/min. O2 kinetics were defined as O2 deficit (time [rest to steady state] x delta VO2-sigma VO2 [rest to steady state]) and mean response time (MRT) of oxygen consumption (O2 deficit/delta VO2). The O2 deficit was 551 +/- 134 mL in DDD pacing, 634 +/- 139 mL in VVIR pacing, and 648 +/- 179 mL in VVI pacing (P = 0.001). MRT was 49 +/- 7.8 seconds in DDD pacing, 54.7 +/- 9.5 seconds in VVIR pacing, and 57.4 +/- 11.0 seconds in VVI pacing (P = 0.002). Ten (53%) patients developed symptoms during switch from DDD to VVIR mode whereas the switch from VVIR to DDD mode was not perceived by any patient (P < 0.001). In conclusion, our study shows an impact of AV synchronous pacing and heart rate adaptation on O2 kinetics during low intensity exercise that correspond to casual daily life activities. Our observations may have clinical implications for the management of patients with complete AV block. PMID- 10709226 TI - Scaling structure of electrocardiographic waveform during prolonged ventricular fibrillation in swine. AB - Ventricular fibrillation (VF) is the most common arrhythmia causing sudden cardiac death. However, the likelihood of successful defibrillation declines with increasing duration of VF. Because the morphology of the electrocardiogram (ECG) waveform during VF also changes with time, this study examined a new measure that describes the VF waveform and distinguishes between early and late VF. Surface ECG recordings were digitized at 200 samples/s from nine swine with induced VF. A new measure called the scaling exponent was calculated by examining the power-law relationship between the summation of amplitudes of a 1,024-point (5.12 second) waveform segment and the time scale of measurement. The scaling exponent is a local estimate of the fractal dimension of the ECG waveform. A consistent power law relationship was observed for measurement time scales of 0.005-0.040 seconds. Calculation of the scaling exponent produced similar results between subjects, and distinguished early VF (< 4-minute duration) from late VF (> or = 4-minute duration). The scaling exponent was dependent on the order of the data, supporting the hypothesis that the surface ECG during VF is a deterministic rather than a random signal. The waveform of VF results from the interaction of multiple fronts of depolarization within the heart, and may be described using the tools of nonlinear dynamics. As a quantitative descriptor of waveform structure, the scaling exponent characterizes the time dependent organization of VF. PMID- 10709227 TI - Automated classification of human atrial fibrillation from intraatrial electrograms. AB - The assessment of the degree of organization and the classification of atrial fibrillation (AF) according to the types defined by Wells usually resorts to the visual inspection of bipolar intraatrial electrograms. The focus of this study was to test seven parameters aimed to quantify the degree of organization of the electrograms, and then to design a final classification scheme based on a multidimensional, minimum-distance analysis. The following parameters were tested: mean atrial period (AP) and its coefficient of variation (CV); number of points lying at the baseline (NO) and the Shannon entropy (EN) of the amplitude probability density function (APDF); depolarization width (F-WIDTH); and correlation waveform analysis (CWA) and electrogram bandwidth (BW). The signal database consisted in a set of 160 AF strips of Type I, II, and III AF, scored by an expert cardiologist (60 Type I, 40 Type II, 60 Type III) and further divided in a training set (60) and a test set (100). Strips were 6 seconds long and were recorded with 5-mm interspace bipolar catheters from electrically induced (n = 13) and chronic (n = 10) patients. A classification algorithm based on a minimum distance (Mahalanobis distance) discriminant analysis was tested. Using a single parameter, the best discriminations were provided by NO, F-WIDTH, and CV. F-WIDTH was found strongly inversely correlated to NO (r = -0.90). Of all the two parameter combinations, CV-NO provided the best classification: 92 of 100 segments were correctly classified with sensitivity > 90% and specificity > 92%. A further improvement was obtained by including BW as a third parameter (93/100 correctly classified). The use of more than three parameters not only failed to improve, but even degraded the classification. PMID- 10709228 TI - Prospective comparison of lesions created using a multipolar microcatheter ablation system with those created using a pullback approach with standard radiofrequency ablation in the canine atrium. AB - The aim of this study was to compare the lesions created using a multipolar microcatheter (MICRO) ablation system in the right canine atrium to a pullback approach with a standard radiofrequency (STND RF) ablation and to determine the value of electrogram amplitude and pacing threshold in predicting transmurality of lesions. Ten dogs underwent right atrial ablation using a MICRO (6 dogs) or STND RF (4 dogs) ablation system in each animal. Attempts were made to create linear RF lesions at four predetermined atrial sites. RF energy was delivered for 60 seconds using closed-loop, temperature control to achieve a target temperature of 60 degrees C for STND RF and 50 degrees C for MICRO. Unipolar atrial electrogram amplitude and atrial pacing threshold were obtained before and after ablation. Pathological analysis was determined at 4 weeks after ablation. Lesions created with MICRO were narrower, more likely to be continuous, and more likely to be anchored to an anatomic structure than those lesions which were created using a STND RF. No difference was observed in overall lesion length or in the proportion of lesions that were transmural over at least 50% of their length. Of lesions created using MICRO, a significant relation was observed between transmurality of lesion and unipolar electrogram amplitude as well as pacing threshold. Further studies are needed to determine if this type of ablation technique and parameters during ablation may facilitate a successful catheter based MAZE procedure. PMID- 10709229 TI - Reproducibility of response to programmed atrial stimulation. AB - The induction of atrial tachyarrhythmias (ATAs) is used to guide the medical or ablative treatment of these tachycardias. To date no information is available regarding the reproducibility of programmed atrial stimulation (PAS) induced ATA. The purpose of the study was to look for the reproducibility of PAS. Two baseline electrophysiological tests were performed in the drug-free state and within 6 months to 3 years of one another (mean 18 months) in 62 patients. Twenty-six patients had spontaneous documented ATAs (group I); 36 patients did not have spontaneous ATAs (group II). PAS used one and two extrastimuli delivered during three cycle lengths (sinus rhythm, 600 ms, 400 ms). The results were as follows. In group I, sustained (> 1 minute) ATA was induced in 23 patients on the first PAS and remained inducible in 22 patients in the second study. In three patients with noninducible ATA, PAS remained negative in only one; the reproducibility of PAS was 88%. In 17 (47%) group II patients, a sustained ATA was induced in the first study, and the ATA remained inducible in 10 patients in the second study. Nineteen other patients did not have inducible ATA on the first study, but 10 of them had an inducible ATA on the second PAS; the reproducibility of PAS was 53%. In conclusion, long-term reproducibility of PAS induced ATA in patients with spontaneous and documented ATA was good. In patients without spontaneous ATA, the reproducibility of PAS induced ATA was low and the induction of ATA in these patients should be interpreted cautiously in light of this observed variability in induced atrial arrhythmias. PMID- 10709230 TI - The effect of ibutilide on retrograde accessory pathway conduction. AB - Ibutilide is a compound with Class III effects marketed for rapid conversion of atrial fibrillation and atrial flutter. The Class III effect is primarily mediated by blockade of the rapid component of the cardiac delayed rectifier of potassium current, Ikr. Ibutilide was used in three patients with concealed accessory pathways during electrophysiological evaluation for ablation of symptomatic atrioventricular reentry tachycardia. Each pathway (mid-septal, left posterior, and left lateral) exhibited a mean retrograde effective refractory period of 240 +/- 20 ms. Each patient had atrioventricular reentry tachycardia that consistently degenerated to recurrent sustained atrial fibrillation. One to two milligrams of intravenous ibutilide converted atrial fibrillation to sinus rhythm and maintained sinus rhythm throughout the procedure. Retrograde accessory pathway conduction was unchanged. Maintenance of sinus rhythm allowed for successful mapping and catheter ablation of the concealed accessory pathways. No direct current cardioversion was needed. In these patients, ibutilide was effective in converting and controlling atrial fibrillation induced by atrioventricular reentry tachycardia without masking retrograde pathway conduction. Antegrade accessory pathway conduction could not be assessed in this study. PMID- 10709231 TI - Catheter ablation approach on the right side only for paroxysmal atrial fibrillation therapy: long-term results. AB - We report the long-term follow-up of a right side only catheter ablation approach for paroxysmal AF. Eighteen patients with AF refractory to drugs entered the study. Ablation was attempted in the right atrium only by creating linear lesions based on a specific design including from two to four linear lesions. Induction of AF was attempted before ablation and after placement of the lesions. A septal lesion was performed in nine patients. In ten patients atrial defibrillation thresholds (ADFTs) before ablation and following creation of the linear lesions were compared. After a mean follow-up of 22 +/- 11 months, seven patients had recurrence of AF, and another nine patients experienced atrial flutter or atrial tachycardia. Five patients remained in sinus rhythm without medications and four required the use of drugs. Three patients had sporadic AF and six were in chronic AF. The recurrence rate was similar in patients with and without the septal lesion. However, a cure with right side ablation appeared to be predicted by the presence of disorganized and earlier activity in the high right atrium and crista terminalis. Linear lesions in the right atrium were associated with a lower ADFT (pre 2.6 +/- 04 J vs post 1.7 +/- 0.6 J). In conclusion, in a small number of patients, control of AF can be obtained with a right side only approach. Certain activation patterns may identify patients suitable to this approach. No specific lesion pattern appeared more effective. Right atrial linear lesions resulted in lower ADFT. PMID- 10709232 TI - QRS amplitude and shape variability in magnetocardiograms. AB - In magnetocardiography, averaging of QRS complexes is often used to improve the signal-to-noise ratio. However, averaging of QRS complexes ignores the variation in amplitude and shape of the signals caused, for example, by respiration. This may lead to suppression of signal portions within the QRS complexes. Furthermore, for inverse source, reconstructions of dipoles and of current density distributions errors in the spacial arrangement may occur. To overcome these problems we developed a method for separating and selective averaging QRS complexes with different shapes and amplitudes. The method is based on a spline interpolation of the QRS complex averaged by a standard procedure. This spline function then is fitted to each QRS complex in the raw data by means of nonlinear regression (Levenberg-Marquardt method). Five regression parameters are applied: a linear amplitude scaling, two parameters describing the baseline drift, a time scaling parameter, and a time shift parameter. We found that both amplitude and shape of the QRS complex are influenced by respiration. The baseline shows a weaker influence of the respiration. The regression parameters of two neighboring measurement channels correlate linearly. Thus, selective averaging of a larger number of sensors can be performed simultaneously. PMID- 10709233 TI - Effect of underlying heart disease on the frequency content of ventricular fibrillation in the dog heart. AB - Although prior studies have examined the frequency content of local electrogram characteristics during fibrillation, little is know about the effects of underlying heart disease on these parameters. This study was designed to compare the frequency content of local electrograms during VF in canine models of acute ischemia, subacute infarction, and chronic myocardial infarction (MI) to those in control animals to test the hypothesis that underlying heart disease can alter the basic characteristics of VF. VF was induced using burst pacing in three groups of mongrel dogs. Five dogs were evaluated 8 weeks after LAD occlusion MI, five were evaluated 5 days after experimental MI, and 5 had VF induced before (control) and immediately after LAD occlusion (ischemia). During VF, unipolar electrograms were recorded from 112 sites on the anterior LV and electrograms were evaluated 15 and 30 seconds after VF initiation in each group. Electrograms were analyzed by fast Fourier transform. No significant time dependent changes in VF characteristics were noted. The peak frequency was highest in control animals and 8-week MI, intermediate in 5-day MI, and lowest in acute ischemia (P < 0.01 for pairwise comparisons). In contrast, the fractional of energy within a bandwidth of 25% peak amplitude was highest in acute ischemia, (P < 0.001) and similar in the other three groups. Infarction decreased total energy by approximately 50%. In conclusion, the pressure of ischemia or infarction alters the frequency content of VF in a complex fashion. In addition to decreasing the peak frequency, the shape of the power spectral curve is altered in models of structural heart disease. These results suggest that the electrophysiological changes produced by infarction or ischemia alter the structural organization of ventricular fibrillation. PMID- 10709234 TI - Popping phenomena in temperature-controlled radiofrequency ablation: when and why do they occur? AB - During temperature-controlled radiofrequency (RF) ablation a popping sound sometimes occurs. This popping phenomenon is known to be associated with unwanted effects like blood boiling, endocardial rupture, catheter dislocation, and impedance rise. The present in vitro study determined the influence of cooling, electrode contact, and tip temperature on the occurrence of popping phenomena. Pieces of porcine ventricle were immersed in a bath of saline solution at 37 degrees C. Forty-two RF ablations were performed with different electrode-tissue contact forces (i.e., 0.0-0.44 N) in a temperature-controlled mode (70 degrees C setpoint, 30 s, 50 W maximum power output, 4-mm tip, thermocouple). Half of the 42 ablations were performed with fluid flow (0.1 m/s, group I), the other half without flow (group II). In group I, mean tip temperature and power were 55.6 +/- 8.5 degrees C and 36.2 +/- 13.8 W, resulting in a lesion volume of 121 +/- 57 mm3. In group II, the respective values were 67.3 +/- 1.5 degrees C and 9.9 +/- 5.2 W resulting in a volume of 42 +/- 18 mm3. The differences between groups were statistically significant. Overall, ten popping phenomena occurred in group I and none in group II. Pops occurred significantly more often when the contact force was < 0.1 N (8/10) and the tip temperature was < 60 degrees C (8/10). Two endocardial ruptures occurred, both were associated with a popping phenomenon. Using temperature control, the probability of pops is significantly higher when the ablation electrode and the endocardial tissue surface are exposed to fluid flow and the electrode-tissue contact is poor. Under these conditions the tissue temperature can be much higher than the temperature measured at the tip electrode and can potentially reach 100 degrees C causing intramyocardial steam formation and a popping phenomenon. PMID- 10709235 TI - Signal-averaged P wave duration predicts early recurrence of atrial fibrillation after cardioversion. AB - Thirty-two patients had signal-averaged P wave duration measured after electrical cardioversion of AF, and were followed for 1 year or until there was a recurrence. The use of antiarrhythmic medications was left to the discretion of the attending physician. Among 20 patients not taking antiarrhythmic medication, the 11 patients who had a recurrence of AF within 3 months of cardioversion had a significantly longer signal-averaged P wave duration compared to the 9 patients who did not (148 +/- 17 vs 135 +/- 20 ms, P = 0.005). There was no difference in clinical parameters or left atrial diameter. A signal-averaged P wave duration cutoff anywhere between 130 and 135 ms correctly classified 85% of patients with a sensitivity of 81% and a specificity of 89%. In patients taking antiarrhythmic medications, signal-averaged P wave duration did not correlate with the risk of recurrence. In patients not taking antiarrhythmic medications, signal-averaged P wave duration can be used to predict the risk of an early recurrence of AF after cardioversion. The poor predictive value in patients taking antiarrhythmics may be due to changes in the atrial refractory period, which are not reflected in P wave duration. PMID- 10709236 TI - Spontaneous recurrent ventricular fibrillation in a patient with a structurally normal heart. PMID- 10709237 TI - Basket catheter localization of the origin of atrial tachycardia with atypical morphology after atrial flutter ablation. AB - Atrial activation from a site in the low lateral right atrium will typically proceed in a superior direction. We present a case of a low lateral right atrial tachycardia with a surface electrocardiographic P wave morphology that appeared to have an inferiorly directed axis. The tachycardia occurred 2 years after successful atrial flutter ablation. The use of a multipolar basket catheter allowed confirmation of the focal origin of the tachycardia, permitted its rapid localization, facilitated catheter ablation, and provided clues to atrial activation that helped describe the appearance of the P wave. PMID- 10709238 TI - Catheter ablation of atrioventricular junction via retrograde route in a patient with single ventricle. AB - Radiofrequency catheter ablation of the atrioventricular junction (AVJ) was performed by the retrograde route in a 19-year-old woman with atrial fibrillation and single ventricle following the bidirectional Glenn procedure. Two energy applications resulted in complete atrioventricular block and dependence on an epicardial ventricular pacemaker. PMID- 10709239 TI - Orthodromic tachycardia with atrioventricular dissociation: evidence for a nodoventricular (Mahaim) fiber. AB - We describe a patient in whom two tachycardias with AV dissociation were inducible by ventricular extrastimulation. The first tachycardia was characterized by a narrow QRS preceded by a His deflection with an HV interval identical to that recorded in sinus rhythm (40 ms). Premature ventricular depolarization delivered when the His bundle was refractory advanced the next His deflection. These findings suggest the presence of a nodoventricular bypass tract involved in an orthodromic tachycardia. The second tachycardia was induced after propafenone infusion and exhibited a wide QRS complex with left bundle branch block morphology; each ventricular complex was consistently associated with a His deflection with a HV interval of -15 ms. The second tachycardia may be considered to represent an antidromic tachycardia through the nodoventricular tract. However, a ventricular tachycardia cannot be excluded. PMID- 10709240 TI - Late onset of accessory pathway conduction in a patient with complete AV block. AB - This case report discusses a patient with complete AV block in early childhood. The patient required a permanent pacemaker. At 6 years of age, intermittent preexcited beats were noted on telephonic transmissions. At 7 years of age, 1:1 preexcitation was noted in sinus rhythm. Therefore, late onset of antegrade accessory pathway function is demonstrated. This case provides evidence of developmental changes in accessory pathways. This may explain age related differences in the onset of narrow complex tachycardia in the school age years. PMID- 10709241 TI - Atrial tachycardia as the presenting sign of a left atrial appendage aneurysm. AB - A patient presented with atrial tachycardia. The work-up, guided by the tachycardia morphology, led to the diagnosis of left atrial appendage aneurysm. Surgical removal of the atrial appendage resulted in cure of the tachycardia and associated symptoms. PMID- 10709242 TI - The diagnostic dilemma of "pseudopacemaker spikes". AB - In a patient who sustained sudden collapse, later attributed to pulmonary embolization, an ECG during her evaluation demonstrated sinus tachycardia and stimulus artefacts at a rate of 250 per minute which did not capture the heart. Implanted pacemaker malfunction was considered the cause until a chest X ray showed a transcutaneous electrical nerve stimulation (TENS) device, which was the source of the artefacts. In instances of rapid stimulus artefacts on the ECG that do not capture the heart, the presence of a TENS device should be considered. PMID- 10709243 TI - A superfamily of small potassium channel subunits: form and function of the MinK related peptides (MiRPs). PMID- 10709245 TI - [In Process Citation] PMID- 10709244 TI - Conserved geometrical base-pairing patterns in RNA. PMID- 10709246 TI - [Calponin: biological, chemical and structural properties]. AB - Calponin distribution in smooth muscle cells and its properties in experiments in vitro are described. A comparison of these properties with those of other regulatory proteins and of proteins presumed to play this role suggest that calponin has another, yet unknown function. On the basis of existing experimental and theoretical data, an attempt has been made to quantitatively estimate the content of structural elements of the polypeptide chain and their localization. With consideration of the structural calponin domains, a general model of the secondary structure of all known calponin sequences is suggested. Based on the known roentgenographic data of the CH-domain of other proteins a possible organization of the calponin molecule hydrophobic core has been proposed. PMID- 10709247 TI - [Role of myosin light chains in regulating muscle contraction]. AB - Current review is focused on regulatory functions of myosin light chains from different muscle types. Special attention is paid to myosin light chains from striated muscles. The present review considers mainly the relevant data provided after 1986. PMID- 10709248 TI - [Structural and functional characteristics of muscle allotransplants, formed under different conditions of laser irradiation]. AB - It is known that a low-energy laser radiation can cause reflex suppression of immunity. The present experiments were designed to determine the plastic activity of allogenic muscle tissue in different conditions of a previous action of laser rays. The cross homotransplantation of gastrocnemius muscles was carried out between intact rats, or between rats in which 14 days before transplantation each hind leg was subjected to low-energy He-Ne laser radiation in dose of 7.5-9 J/cm2 (10 procedures, the duration of each exposure was equal to 5 min), or between intact and radiated rats. It was shown that the donor muscle tissue survived longer when a nonradiated muscle was transplanted into the radiated muscle bed. The axons grew into the donor muscle tissue. More allogenic muscle tissue was involved in contractile reaction when stimulation was carried out via the nerve. Laser radiation of a homotransplant alone, or that of a homotransplant and a muscle bed in the recipient was less effective. So, He-Ne laser radiation of the area of a planned allotransplantation decreased the transplant immunity response and favoured a longer development of allogenic muscle tissue. The viability of donor muscle tissue therewith increased, if the muscle allograft had not been subjected to a previous laser radiation. PMID- 10709249 TI - [Assessment of pathological changes in fish muscle tissue, forming as a result of exposure to toxic factors in the environment]. AB - Our investigation of muscle tissue of fishes, inhabiting the regions with unfavorable ecological conditions (the river Volga), permitted to select four types of degenerative changes in muscle tissue. These alterations are associated with both the phylogenetic status of fish species and ecological dispositions of species. Using different methods of investigation several types of muscle destruction were shown. I. Destruction of myofibrillar apparatus (lysis of protofibrils), with sarcolemma remaining intact. II. Destruction of the myofibrillar apparatus, with sarcolemma, T-system, and sarcoplasmic reticulum being disrupted. III. Invasion of muscle fibers by lymphoid cells and macrophages; with sarcolemma being intact. IV. Lysis of sarcolemma by proteolytic enzymes of lymphoid elements; with muscle fibers being disintegrated. The objects of this study were muscle tissues of 8 fish species (Acipenser gueldenstadti, A. stellatus, A. ruthenus, Lucioprerca lucioperca, Esox lucius, Perca fluviatilis, Tinca tinca, Caprinus carpio). The white muscle degeneration followed the patterns of types I and II, while that of red muscles corresponded to types III and IV. White and red muscles of the Chondrostei fishes (sturgeon, stellate, sterlet) undergo destruction more frequently, than muscles of the Holostei fishes (pike, perch, zander, sazan, tench). Degenerative processes of white and red muscles of fish-eating fishes were more obvious than those of herbivorous fishes. PMID- 10709250 TI - Secophalloidin and phalloidin-(S)-sulfoxide as contraction modifiers for comparative study of skeletal and cardiac muscles. AB - Phalloidin, a toxic product of the mushroom Amanita phalloides, binds specifically to F-actin resulting in strong stabilization of F-actin structure (for review, see; Wieland, 1986). Binding to a specific site on the muscle thin filament F-actin, phalloidin modifies contraction in a tissue specific manner. Phalloidin induced changes depend on functionally important parameters (thin filament activation, cross-bridge kinetics), indicating changes in essential steps of the contractile mechanism. Moreover, there is a different action with different phalloidin derivatives. Such properties make phallotoxins (phalloidin and its derivatives) powerful modifiers for muscle research (for review, see: Bukatina, 1996). Phalloidin-induced changes vary qualitatively with muscle types. In all types of skinned skeletal muscle preparations that have been studied (fast and slow muscles from evolutionarily distant animals), the most general effect of phalloidin is to cause a decrease in tension (Bukatina, Morozov, 1979; Alievskaya et al., 1987; Bukatina et al., 1993). In mammalian skeletal muscles, this decrease in tension may be followed by a slowly developing increase in tension. The resulting tension may considerably exceed the tension before phalloidin administration. In contrast, skinned cardiac muscle responds to phalloidin only by increasing isometric tension from the onset of the response. Moreover, the phalloidin response is completed in approximately one-tenth the time in cardiac muscle that it takes in skeletal muscle. These phalloidin effects in cardiac muscle result in an enhanced Ca2+ responsiveness (Boels, Pfitzer, 1992) with an increase in both the force at maximum Ca2+ activation and the Ca2+ sensitivity (Bukatina et al., 1995). PMID- 10709251 TI - [Spreading of porcine kidney epithelial cells under normal condition and under the effect of inhibitors of energy metabolism. I. Dynamics of spreading]. AB - In the present work dynamics of cell spreading on a solid substratum has been investigated in normal conditions and under ATP-synthesis inhibitors. Sodium aside (20 mM), which blocks ATP-synthesis in mitochondria, and N,N dicyclohexylcarbodiimide (DCCD) (100 mM), which blocks both ATP-synthesis and glycolysis, were ATP-synthesis inhibitors of choice. In the range from the moment of cell plating to 24 h three stages of cell spreading could be distinguished according to the dynamical change of the average projected cell area. At the first stage, within 1.5 h in the control culture the cell area increases rapidly. The slowing of spreading occurs at the second stage, within 1.5-4.0 h. The third stage is characterized by a slow but pronounced increase in cell spreading, which ceases in 24 h. Under inhibitory treatment, the pattern of cell spreading during the first 4 h is essentially the same as in control conditions. The subsequent DCCD action results in cell spreading inhibition; sodium aside, on the contrary, accelerates the spreading. The cell shape analysis has demonstrated that even as early as in 0.5 h the first small polarized cells appear simultaneously with the nonpolarized cells. In control cells, the share of polarized cells increases in almost 2 h, conversely, under drug actions the process of polarization begins earlier to be more pronounced in the presence of sodium aside. Thus, it has been shown that the spreading of PK cells does not require any additional ATP synthesis. At early stages in normal conditions and under inhibitory treatment the picture of cell spreading is the same. A complete inhibition of ATP-synthesis slow down the process of cell spreading. However, an activation of these processes was observed in cells with low content of ATP, resulting from glycolysis retaining. PMID- 10709252 TI - [Spreading of porcine kidney epithelial cells under normal conditions and under the effect of inhibitors of energy metabolism. II. Behavior of cellular organelles]. AB - In the present work the behavior of mitochondria and lysosomes during cell spreading has been investigated in normal conditions and under ATP-synthesis inhibitors: sodium aside and N,N-dicyclohexylcarbodiimide (DCCD). In the control culture, microtubules run along the stable edge and perpendicular to the leading edge in most of spreading cells. As a whole, microtubules form a dense network in these cells. However, the radial cells contain bundles of microtubules, radiating from the perinuclear area or form circular arrays around the nucleus. The microtubule network is more dense under inhibitory treatment, than in control conditions. In the control culture the spherical cells display numerous small mitochondria (staining with Rhodamine 123). In the process of cell spreading some elongated mitochondria appear, most of them being localized in the perinuclear area. The mitochondria of cells with radial microtubule organization are directed towards the cell periphery, while in cells with circular bundles of microtubules the mitochondria are localized chaotically. Under DCCD treatment the mitochondria retain the staining for 2-3 h. In the spreading cells, round mitochondria may be distributed all over the cytoplasm. In the presence of sodium aside the mitochondria are not stained. However, by means of phase contrast microscopy some disoriented thread-shaped structures are observed, obviously corresponding to mitochondria. In the control conditions, lysosomes (stained with Acridine orange) in spreading cells are dispersed chaotically, all over the cytoplasm, or are localized in the perinuclear area. In the presence of sodium aside lysosomes are observed only in the perinuclear area. Under DCCD treatment lysosomes do not accumulate the dye. Thus, the cytoskeleton modification and changes in the properties of membrane organelles, induced by ATP-synthesis inhibitors, do not prevent attachment, spreading or cell polarization. PMID- 10709253 TI - [Migration of fibroblasts into heart valve leaflet tissue in vitro]. AB - Heart valve allografts are widely used for surgical treatment of the heart. In recent years a new field of research has emerged dealing with allograft modification by cells of recipient by means of tissue engineering. This method involves culturing fibroblasts and endothelial cells, using recipient tissue, followed by introduction of the fibroblasts into tissues of allograft and coating its surface by the endothelial cells. This modification is expected to ensure the structural maintenance of implanted tissues and to reduce its thrombogenecity. This procedure may promote the allograft adhering to the recipient tissues, thus prolonging the terms of the valve normal functioning after implantations. For this purpose, methods of luminescent microscopy are suggested using double staining of tissue with fluorescent dyes Hoechst 33,342 and ethidium bromide, or with fluorescein diacetate and ethidium bromide. Experimental results are presented indicative of fibroblast migration from the surface to the human heart valve leaflets. PMID- 10709254 TI - [Analysis of elemental composition (Na/K/Ca) of muscle cells in ischemia in a model of the perfused heart]. AB - It has been hypothesized that cardiac ischaemia induces some changes in the cardiac myocyte element. In the present study we analysed whether the imbalance of potassium, sodium and calcium in cardiomyocyte may be coupled with ischaemic conditions, using a perfused heard as a model. Electron Probe Microanalysis (EPMA) of papillary muscle cryosection was employed to examine the intracellular content of elements. Following a 30 min acute ischaemia the intracellular potassium was not changed and sodium was reduced. During a prolonged ischaemia (45 min) [K] loss was shown and [Na] concentration was seen reestablished. These results demonstrate that the active transport of potassium and sodium is possible at the beginning of ischaemia. This suggests that under abaerobic conditions Na-K ATPase may be functionally coupled with an ATP-sensitive K channel through intracellular messenger, possibly ATP. PMID- 10709255 TI - [Nonenzymatic glycosylation of and oxidative damage to actin in vitro and in vivo]. AB - A study was made the influence exerted by non-enzymatic glycosylation (glycation) and oxidative destruction on structural and functional parameters of actin (free NH2-groups, advanced glycation end product and bityrosine cross-linking content, DNase inhibition by G-actin and myosin Mg(2+)-ATPase activation by F-actin). The functional properties of actin were shown to change under high molecular weight product formation and oxidative destruction: the extent of DNAase I inhibition decreases (from 70 to 40%) and the extent of myosin Mg(2+)-ATPase decreases (by 40%). Carnosine prevents actin oligomer formation and oxidative destruction which favours preservation of the protein functional properties. PMID- 10709256 TI - [Nitric oxide as a contrast modulator of basic elements of the cytoskeleton]. AB - Early it was shown that nitric oxide induced in the cerebellum neuronal net both degenerative and compensatory-adaptive changes: 1) bouton encapsulation of spines, and 2) spiral wraps formed by glial cell processes around synapses and boutons. All these morphological changes were produced with cytoskeleton involvement. In the present work we have found that a NO-generative compound enhanced the contrast of cytoskeleton elements which depended on the concentration of this compound. The best contrast was observed at 1 mM concentration. The reason of the contrast enhance may be due presumably to protein transition from a soluble to a membrane-bound state. Using the contrast enhance effect we carried out a comparative analysis of cytoskeleton elements (CE) composition. Results of the analysis showed the specificity of CE in different cell structures: bouton, spine, glial cell. The obtained data support our proposal about the leading role of cytoskeleton in compensatory-adaptive morphological changes in extremal conditions. PMID- 10709257 TI - [Concentration of macroergic phosphates and oxidative potential of skeletal muscles]. AB - It is known that a long-duration decline of high-energy phosphate (HP) level in skeletal muscles, induced by administration of beta-guanidinpropionic acid (beta GPA), is followed by an increase in mitochondrial enzyme activities (MEA). The same increase in MEA was observed in the course of physical exercise training. Under gravitational inloading decrease in MEA and increase in the level of high energy phosphates occurred. If changes in (HP) level are believed to trigger the alterations in MEA, the increase in high-energy phosphate levels in muscles is to lead to a decline in MEA as well. The present work was purposed to reveal if changes in HP level under different contractile activity levels may be associated with changes in oxidative potential in the skeletal muscles. PMID- 10709258 TI - [Cytomechanical control of morphogenesis]. AB - The role of mechanically strained state of cells and multicellular structures in morphogenesis regulating in vertebrate embryos is discussed. Regular changes in patterns of mechanical strain during embryonic development are described. Artificial relaxation of mechanical strain performed on definite developmental stages and retension of embryonic tissues in arbitrary directions considerably affects morphogenesis and cell differentiation patterns. Cytomechanical models of morphogenesis are reviewed and a concept of hyperrestoration of mechanical strain as a possible driving force of morphogeneiss is suggested. PMID- 10709259 TI - [Activity of the genome of cardiomyocytes as an indicator of the development of adaptive changes in the myocardium following exposure of the central nervous system to electromagnetic fields]. AB - Methods of cardiomyocyte nuclei isolation from the myocard homogeneous mixture, and of cardiomyocyte genome activity estimation were elaborated. In the experiments with hyperlipoproteidemic rats, cardiomyocyte genome activity was shown to reflect the primary adaptive changes in the myocard, and to serve a reliable index of their influence on the CNS regulatory centres exposed to electromagnetic field, which is used for hyperlipoproteidemia treatment. The cardiomyocyte genome activity was used to distinguish between three types of development of adaptive reactions in the myocard. PMID- 10709260 TI - [Ultrastructure of skeletal muscle fibers in monkeys after space flight]. AB - It is known that exposure of humans and animals to microgravity causes reduction in the cross-sected area of muscle fibers and muscle atrophy. These changes also involve ultrastructural alterations in muscle fibers. Therefore primates, that are physiologically close to humans, are to be examined to help a better understanding of the nature of these ultrastructural changes is muscles and muscle fibers. Although failed to find any relevant published data on the quantitative aspects of ultrastructural changes in muscle fibers of space-flown primates we believe that it is important to examine these aspects. The postflight study of monkey's m. soleus, and m. vastus lateralis did not reveal any significant changes in volume density of the myofibrillar apparatus. Mitochondria of m. soleus showed a distinct reduction in volume density, being more obvious in the subsarcolemmal zone than in the central one. Mitochondria of m. vastus lateralis showed a decrease (P > 0.05) in volume density. Following the flight, m. soleus and m. vastus lateralis of the monkeys showed a significant increase in the mean area of myofibrils, and a trend towards a decrease in the number of myofibrils per 100 micron 2. Besides, m. soleus showed a significant increase in the mean area of mitochondria, and a trend towards a decrease in the number of mitochondria per 100 micron 2. In m. vastus lateralis of the monkeys after space flight the number opf mitochondria tended to decrease and the mean area showed differential changes. It can be postulated that these phenomena may be associated with a reduction in the diffusion surface of mitochondria resulting from the diminished myofibrillar volume. PMID- 10709261 TI - [The immediate results of laparoscopic treatment in perforated gastroduodenal ulcers]. AB - The article presents an analysis of immediate results of laparoscopic interventions (suturing, vagotomy) in 40 patients with perforated ulcers in the pylorobulbar zone. Postoperative complications were noted in 7.5% of the patients. There were no lethal outcomes. The authors recommend wider introduction of the laparoscopic technology as a method of optimization of surgical treatment for perforated ulcers. PMID- 10709262 TI - [The prognosis and prevention of the occurrence of acute ulcers and erosions of the stomach and duodenum in surgical patients]. AB - Acute ulceration of the gastric and duodenal mucosa was found in 20.3% among 404 patients who died at the surgical department of the hospital. In 89.7% of the patients acute erosive-ulcerous lesions were localized in the gastric mucosa. In 39 patients (9.7%) acute ulcers of the stomach and duodenum complicated by bleedings or perforations were the direct cause of death. Based on results of an analysis of severity of the main and concomitant diseases as well as their complications the authors have determined the factors responsible for the appearance of the ulcers. A complex of prophylactic measures aimed at the main links of the pathogenesis of acute ulcers is proposed. PMID- 10709263 TI - [The acid-forming function of the stomach and its hormonal regulation after truncal and combined vagotomies]. AB - The acid-producing function of the stomach and the state of its hormonal regulation were assessed on the basis of the immediate and long-term follow-up results obtained after truncal (334 patients) and combined (96 patients) vagotomy for duodenal ulcer. According to the data of aimed endoscopic pH-metry and gastroimpedancemetry the acid level changes in different parts of the stomach and duodenum were the same at the nearest and remote periods as well as the dynamics of the main hormones responsible for the acid production. PMID- 10709264 TI - [The pathogenesis of achlorhydria of the operated stomach and the functionally optimal volumes for distal resection in areflux gastrointestinal anastomosis]. AB - Examinations of 68 patients after operations of selective proximal vagotomy and 59 patients after the Billroth-II and Roux resections were performed within the periods of about 16 years. It was found that reflux was of main significance in the genesis of achlorhydria of the operated stomach. Gastric changes caused by Helicobacter pylori did not result in so rapid suppression of functional activity of the operated stomach. Hemiresection is thought to be optimal for the areflux Roux gastrojejunal anastomosis with hypersecretion. In patients with normal secretion a resection of 2/3 of the stomach is expedient. In cases with hypersecretion and unknown boundaries of the antrum a resection of 50-60% of the stomach must be supplemented with vagotomy. PMID- 10709265 TI - [Lipid peroxidation and antioxidant system functions in patients with gastroduodenal ulcerous hemorrhages undergoing conservative and surgical treatment]. AB - The results of examination of 53 patients have shown that greater intensity of lipid peroxidation and less activity of the antioxidant system of blood are more pronounced in the operated patients. The authors recommend to include antioxidants in the complex of medical measures. PMID- 10709266 TI - [The lipid profile in patients with obesity who have undergone vertical gastroplasty]. AB - The effect of vertical banded gastroplasty (VBG) on the lipid profile in obese patients is considered. The antiatherogenic effect of VBG and of the following weight loss consists in significantly decreased plasma triglycerides, increased content of high-density lipoproteins and decreased atherogenic coefficient. Patients with hypercholesterolemia have significantly lower total plasma cholesterol by the period of the body mass stabilization. The effect of VBG is compared with that of other methods of correction of dyslipidemia. The possibility to improve this effect is also discussed. PMID- 10709267 TI - [The dynamics of blood serum middle-molecule peptides in the prognosis of the course of acute pancreatitis]. AB - The work is devoted to studying the dynamics of endogenous toxemia by the level of middle mass peptides of blood serum in patients with acute pancreatitis. The level of middle mass peptides was investigated in 116 patients spectrophotometrically after Gabrielian with the wave length 254 nm (1st fraction, MMP-1) and with the wave length 282 nm (2nd fraction, MMP-2). It was found that the ratio of these fractions could be used as a prognostic index of the complicated course of acute pancreatitis. When the initial ration MMp-2/MMP-1 is less that 1 the risk of the development of complications of the second phase of acute pancreatitis is real. The decrease of the level of MMP-1 and MMP-2 during treatment by the 3rd-4th days is considered to be an indicator of the favorable outcome of the disease. The elevation of the level of MMP-1 and MMP-2 by the 3rd-4th days despite the treatment predicts an unfavorable outcome of the disease. PMID- 10709268 TI - [The clinical picture and surgical treatment of adhesive intestinal obstruction]. AB - Results of examination and treatment of 128 operated patients were analyzed. In addition to clinical and roentgenological data the ultrasonic investigation and laparoscopy were also made for diagnosis of commissural ileus. Prevailing was terminal ileus (109 patients). Dissection of the commissures was performed in 115 patients, dissection of the commissures with resection of the intestine was made in 29 patients. Combined operations were fulfilled in 17% of the patients. Sonic stimulation of the intestine was used parallel with medication at the postoperative period which led to earlier restoration of the intestine function. Serious complications were noted in 19 patients, 14 patients died. Main causes of the death were intoxication and peritonitis, hepato-renal and cardio-vascular insufficiency. PMID- 10709269 TI - [The prognosis of the immediate results of surgical treatment in colonic obstructive ileus of tumor origin]. AB - For making prognosis of high risk of the lethal outcome the authors used a correlative analysis of 23 factors influencing the treatment results. Ten of them have high correlation coefficients and reliably influence the level of postoperative lethality. The rule of the prognosing of the nearest results of operative treatment is formulated which allows the adequate assessment of the patient's state in dynamics and the timely correction of the process of treatment. The elaborated scheme of the score assessment is simple in use, informative and statistically reliable. It can be recommended for wide clinical practice. PMID- 10709270 TI - [The effect of tunneling on the pressure in the tibial cavity in arterial insufficiency in the extremity]. AB - The intraosseous pressure in the diaphyseal cavity of the tibia was investigated in 33 adult dogs after the introduction of the needle for the investigations, tunneling of the bone with a wire and modelling of the arterial insufficiency in the extremity. It was found that perforated defects made in compact substance led to a certain lowering of the pressure. The arterial insufficiency is a cause of stable decrease of the intraosseous pressure. PMID- 10709271 TI - [The diagnosis and surgical treatment of neck wounds]. AB - The authors share their experiences with treatment of wounds of the neck in 210 patients. Major vessels were injured in 85 of them (40.5%), arterial vessels being injured in 19 patients, venous vessels in 23 patients. 43 patients had wounds of the hollow, parenchymatous organs and nerves. All the patients with wounds of the hollow and parenchymatous organs recovered. Lethality among the patients with wounds of the major vessels was 11.9% and was due to a combination of injuries of the vessels and blood loss incompatible with life. Active surgical treatment of penetrating wounds of the neck with careful revision of the wound, use of emergency esophagogastroduodenoscopy, laryngotracheobronchoscopy, X-ray examination allow the amount of diagnostic errors and lethality to be reduced. PMID- 10709272 TI - [The diagnosis of disorders of the colorectal innervation in children]. AB - The problem of congenital and acquired defects of innervation of the distal colon has many unsolved questions for making clinical and histological diagnosis, classification and using methods of treatment. Clinical, radiological and morphological aspects of treatment of 153 patients aged from 2 days to 14 years with disturbances of the colorectal innervation were analyzed. Radiological examination was not sufficient for making the diagnosis. Histological signs of aganglionosis, hypoganglionosis, dysganglionosis, type-A and type-B neuronal intestinal dysplasia in full-thickness biopsy specimens were used to confirm the diagnosis of congenital defects of innervation of the distal colon. The scheme of patho- and morphogenesis and clinico-morphological classification of disturbances of the colorectal innervation were proposed. Hirschsprung's disease was classified as a variant of the disturbance of the colorectal innervation. PMID- 10709273 TI - [The surgical treatment of degenerative-dystrophic diseases of the hip joint]. AB - An analysis of long-term results of surgical treatment of 609 patients with degenerative-dystrophic diseases of the hip-joint was made. Choice of the optimum methods of surgical treatment of such patients must be based on the prognosis of effectiveness of the operative intervention taking into account general regularities associated with the degree of the arthrosis alterations, dyscongruence of the articulation surfaces and duration of the disease, the degree of disturbances of the femur head blood supply, the disturbed supporting ability and severity of the trauma. When choosing the method of surgical treatment of young patients with coxarthrosis the operative interventions aimed at saving the hip joint should be considered. PMID- 10709274 TI - [Variants in the methods of plastic compensation for different defects of the middle face with pedicled soft-tissue flaps]. AB - The author describes some original methods of reconstructive and restorative operations for correction of complex defects and deformations of the middle area of the face by using pedicled flaps from the cheeks, chin, lips and lateral parts of the nose. Methods of restoration of the hairy area over the upper lip in men are described as well as plasty of a defect of the red margin of the upper lip, restorative operations for through defects of the nose and elimination of absolute stenosis of the nose passage with a pedicled flap from the cheek with the simultaneous performing of radical cheilorhinoseptoplasty. The method gives optimal aesthetic results and reestablishes the respiratory function of the nose. PMID- 10709275 TI - [Erythropoietin in the prevention and treatment of anemia in heart surgery patients]. AB - The authors present results of using recombinant human erythropoietin in patients operated on the heart under conditions of extracorporeal blood circulation. It was found that the intravenous infusions of erythropoietin at the postoperative period accelerated the restitution of circulating erythron indices. The volume of transfusion of the donor erythrocyte-containing media to the patient is given erythropoietin was reliably less than that in the control group. The results obtained allow using erythropoietin to be recommended as an effective method of prophylactics and treatment of anemia in cardiosurgical patients. PMID- 10709276 TI - [The rheological properties of the peripheral blood in acute diffuse suppurative peritonitis and their changes during infusion therapy using Modegel]. AB - The rheological properties of blood and deformability of erythrocytes were investigated in 85 patients with acute diffuse peritonitis. It was found that higher viscosity of blood mainly with slow shift rates were observed in the reactive and toxic phases of the disease. In the reactive phase these changes can be considered as adaptational, in the toxic phase--as pathological ones. Lessened viscosity in the terminal phase of the disease is an unfavorable prognostic sign. PMID- 10709277 TI - [A new method for detoxifying the lymph in patients with cholestatic endotoxicosis]. AB - A method is proposed for lymph detoxication in patients with cholestatic endotoxicosis by indirect electrochemical oxidation. The method is simple and cheap. After the treatment of the toxic lymph with the solution of sodium chloride during 4 hours the concentration of main toxic metabolites in it was substantially decreased while the total protein content was practically not changed. Under study were the results of using electrochemical lymph detoxication in 11 patients with cholestatic endotoxicosis which was followed by better condition of the patients and by faster dynamics of lowering the level of the main toxic metabolites. PMID- 10709278 TI - [The combined use of fentanyl and clofelin in the anesthesiological support of adrenalectomy]. AB - The analgetic effect of a combination of phentanyl and clonidine was investigated in 68 cases of adrenalectomy as an analgetic component of anesthesia. The analgesic potency of the used combination and stability of the autonomous functions were proved to be sufficient in surgical interventions for various types of adrenal tumors. It was found that the mechanism of the clonidine effect was not associated with vasodilatation and was based on its antinociceptive properties. The role of hemodynamic monitoring in prevention of possible complications and adverse effects of the described anesthetic technique was stressed. PMID- 10709279 TI - [A method for plastic repair in oblique inguinal hernia]. AB - Experimental and clinical investigations enabled the authors to develop a method of hernioplasty using a pi-shaped flap from aponeurosis of the external oblique abdominal muscle on the feeding pedicle. In this case the anatomo-functional interrelations of the tissues in the ilio-inguinal area get minimum injuries. It does not make a substantial obstacle for the following regeneration and restitution and so the method is rather efficient and can be used in the everyday practice. PMID- 10709280 TI - [The cure of multiple abscesses of the right lobe of the liver]. PMID- 10709281 TI - [Mesenteric arteriovenous fistula with the formation of an aneurysm and portal hypertension]. PMID- 10709282 TI - [What the journal reported during the first year of its existence]. PMID- 10709283 TI - [Clinico-physiological analysis of cranial dystonia]. AB - To study afferent and efferent mechanisms of both pathogenesis and forming of the symptoms of dystonia a clinical-electrophysiologic examination was performed in 22 aged 35-69 years patients with cranial dystonia. A wide range of different factors in anamnesis of the patients was found, that had had either short or long influence on the facial zone and that had preceded the development of cranial dystonia (2 month-5 years before its rise). This was called by the authors the peripheral factor of dystonia pathogenesis. This peripheral factor may have a starting and maintaining role in pathogenesis of dystonia. The results of electrophysiologic studies (electromyography, method of dermal sympathetic potentials, registration of abdominal reflexes) suggest the presence of diffuse hyperactivity of interneuronal apparatus on different levels of brain stem being, probably, an efferent link in both pathogenesis and symptoms of dystonia as a whole and cranial dystonia, in particular. The conclusion is made that a removal of the peripheral factor as a stable link of pathogenesis of cranial dystonia together with the influence on the efferent link of pathogenesis may be a new aspect in therapy of this disease. PMID- 10709284 TI - [Differentiated types of chronic agoraphobia]. AB - The study included 60 patients aged 18-65 years with anxious-phobic disorder (APD)--agoraphobia with and without panic disorders. Diagnosis of the disease was performed according to ICD-10. It was established that dynamics and outcome of APD with stable agoraphobia depend on some comorbid psychic disorders (neurotic, nonpsychotic affective, personality disorders, slow-progredient schizophrenia). Influence of such disorders on APD depends on level of realization of the comorbid correlations: symptomatic (nondelirious hypochondria) and intersyndromal ones (affective and personal disorders as well as slow-progredient schizophrenia). Typology of APD with stable agoraphobia is proposed, that is determined by comorbid correlations with the phenomena of nondelirious (overvaluable or neurotic) hypochondria. Agoraphobia, comorbid with the phenomena of overvaluable hypochondria, is characterized by isolated phobic avoidance (1-2 situations), lack of the signs of progredience of psychopathologic disorders and relatively favourable outcome (lack of the cases of invalidism, decrease of a professional status in 8% of the patients). Agoraphobia, comorbid with the phenomena of neurotic hypochondria, is characterized by total phobic avoidance, syndromal comorbidity with polymorphic and generalized somatophormic disorders, depressive disorders syndromally completed, slow-progredient schizophrenia and less favourable outcome (avoidance behaviour of severe degrees was found in 57% of the patients, including 18% of patients disabled due to mental disease). PMID- 10709285 TI - [Personal behavior and mental state in anorexia nervosa]. AB - Personality of the patients with anorexia nervosa is characterised by high neurotization and anxiety that resulted in diffuse anxiety in stress situation. Personal peculiarities included also high total hostility and intrapunitivity, inclination to obsessive-phobic and dysthymic reactions and tendency to somatization of anxiety. Such peculiarities promoted the choice of intrapunitivity type of reaction in situation of frustration. Its manifestation increases with an increase in the disease duration. The range of psychotic disorders in anorexia nervosa is restricted to anxious-depressive, obsessive and asthenic sphere. PMID- 10709286 TI - [Follow-up study of patients operated for traumatic subdural hematoma]. AB - In 56 patients operated for traumatic subdural hematomas clinical manifestations were analysed (8 main syndromes of the remote period) as well as the degree of neurologic rehabilitation and the level of social-occupational adaptation. Such adaptation appeared to be rather high: 66.1% of the patients were able to resume work. The highest lethality was in the acute period; the worst rehabilitation and follow-up adaptation were observed in elderly and old patients who were in comatose state before the operation and had severe accompanying contusion of the brain. Policy of drug treatment was determined in patients with traumatic subdural hematomas regarding peculiarities and manifestations of the syndromes (urgent operation, if necessary--cranioplasty, psychologic support in the remote period, resorption therapy and symptomatic drugs). PMID- 10709287 TI - [The role of group psychotherapy in combined modality therapy of juvenile endogenous depression]. AB - The paper presents the study in of the combined psychopharmaco-psychocorrective method designed specially for juvenile patients with endogenic depressive disorders. Psychocorrective measures included the method of "rehabilitation training of social habits" used on the stage of hospital therapy and psychocorrective training according to "social support" method used in group psychotherapy outpatiently. A 5 year follow-up study performed in 110 showed that psychotherapy has increased efficiency of the therapy in terms of both reduction of clinic disorders in the acute period and prophylaxis of the relapses as well as social readaptation. PMID- 10709288 TI - [Acoustic brain stem and cognitive evoked potentials (P300) in patients with hepatolenticular degeneration]. AB - 18 patients with hepatolenticular degeneration (Wilson's disease, WD) aged 15-38 years were subjected to an overall clinical and neurophysiologic examinations. As a result, the data obtained enable to evaluate functional reserves of CNS of the WD patients in correlation with the illness duration and severity of neurologic symptoms. Correlation between an increase of interpeak I-V and the degree of neurological deficit and, also, level of ceruloplasmin was established (r = 0.45; p < 0.05). Correlation between an increase of latency P300 and the degree of manifestation of neurologic symptoms was identified as well (r = 0.63; p < 0.05). Positive dynamics of evoked potentials was followed in 4 WD patients during copper-eliminative drugs treatment. PMID- 10709289 TI - [Serotonin receptor gene allele polymorphism (5HTR2A) and clinical pathogenetic characteristics in patients with schizophrenia and schizophrenia spectrum disorders]. AB - Serotonin receptor 5HTR2A gene polymorphism was reported to be associated with psychiatric disorders, in particular schizophrenia, in numerous studies. This study aimed to analyze a possible association between 5HTR2A gene polymorphism and clinical and pathogenetic characteristics in schizophrenia and schizophrenia spectrum disorders. We studied 209 individuals with schizophrenia and related disorders (107 male and 102 female, mean age 34.7 +/- 17.2 years) and 116 healthy controls (44 males, 72 females, mean age = 33.6 +/- 14.4 years). Diagnoses were made according diagnostic criteria of ICD-10. Positive and Negative Symptoms Scale (PANSS) assessed clinical symptoms. Significant difference (p < 0.01) was found for 5HTR2A genotype distribution between affected and control groups. The frequencies of genotypes A1A1, A1A2 and A2A2 for schizophrenia were 13.3%, 44.0% and 42.7%, respectively, versus 33%, 47% and 27% in controls. These results support the evidence for association between 5HTR2A A2A2 genotype and schizophrenia. Schizophrenics with A2A2 genotype were characterized by significantly higher mean values of the PANSS negative symptoms subscale than those with A1A1 genotype (22.6 vs 17.8; p < 0.05) and, consequently, by majority of deficit patients and patients with more severe forms of schizophrenia. Patients with A1A1 genotype were younger compare to those with A2A2 genotypes and had the least familial factors (35.7% vs 46.1%). In agreement with the results obtained in the study the 5HTR2A gene polymorphism appears to be considered as additional diagnostic or prognostic trait in the medical genetic studies of schizophrenia. PMID- 10709290 TI - [Multiple sclerosis in Northern-West region of Russia: results of HLA-typing]. AB - Distribution of antigens of A, B, DR loci of HLA system in standard lymphocytotoxic test was studied in 59 patients with a significant diagnosis of multiple sclerosis (MS) and in 138 healthy donors. In the patients elevated frequency of the next antigens was found as compared with the controls: A10 (37%; chi 2 = 6.31; p < 0.05; relative risk--RR = 2.34), B7 (37%; chi 2 = 4.62; p < 0.05; RR = 2.05), B13 (29%; chi 2 = 10.86; p < 0.01; RR = 3.59), B35 (17%; chi 2 = 4.27; p < 0.05; RR = 2.61), DR2 (68%; chi 2 = 11.61; p < 0.001; RR = 2.99), as well as DR6 (5%; chi 2 = 3.95; p < 0.05; RR = 7.34) and also DRw52 (24%; chi 2 = 27.49; p < 0.001; RR = 21.16). The highest value of etiologic fraction was found for DR2 antigen. Analysis of intralocus and extralocus combinations of antigens in MS revealed that significantly elevated frequency had only one combination- B7DR2 (25.4%; chi 2 = 9.77; p < 0.01; RR = 3.58), relative risk was higher for this combination than for each individual antigen separately: B7 (RR = 2.05), DR2 (RR = 2.99). Significant negative associations with a possible protective effect of separate alleles were established in MS for antigens HLA A2 (34%; chi 2 = 5.55; p < 0.05; RR = 0.47), A11 (7%; chi 2 = 4.66; p < 0.05; RR = 0.31), A30 (0%; chi 2 = 4.50, p < 0.05; RR = 0.01), B18 (8%; chi 2 = 4.55; p < 0.05; RR = 0.35), DR5 (59%; chi 2 = 10.17; p < 0.01; RR = 0.36). The most significant was a decrease of the frequency of DR5 antigen (p < 0.01). Patients with the recurrent course had prevailed antigens A11, B21, B35 and decreased frequencies of antigens A9, B13, DR7. However, only the difference in the frequency of DR7 (16% in remitting and 57% in progredient course, chi 2 = 10.02; p < 0.001; RR = 0.14) was significant. PMID- 10709291 TI - [Theoretical-methodological foundation of prophylaxis of nervous and mental diseases]. PMID- 10709292 TI - [Forerunners in neurology: Jackson, Sechenov, Bernstein, Habbard]. PMID- 10709294 TI - [The role of neurodynamic factors in reparative morphogenesis]. PMID- 10709293 TI - [Changes of immunological blood indices in patients with glial tumors of the brain]. PMID- 10709295 TI - [Central pontine and extrapontine myelinolysis]. PMID- 10709296 TI - [Trichobezoar of gastrointestinal tract in child neurosis]. PMID- 10709297 TI - [Syndrome of inadequate antidiuretic hormone secretion in diseases of central nervous system]. PMID- 10709298 TI - [Comments on the letter by V.L. Feigin and A.V. Sukhanov on the article "Risk factors of the development of dementias of Alzheimer's type"]. PMID- 10709299 TI - [Meeting of the committee of the World Federation of Neurologists on education: graduate education in neurology]. PMID- 10709300 TI - [National Association against stroke]. PMID- 10709301 TI - Reflections on complementary medicine research in the UK. PMID- 10709302 TI - A systematic review of craniosacral therapy: biological plausibility, assessment reliability and clinical effectiveness. AB - OBJECTIVES: The objective of this research was to review critically the scientific basis of craniosacral therapy as a therapeutic intervention. DESIGN: A systematic search for and critical appraisal of research on craniosacral therapy was conducted. Medline, Embase, Healthstar, Mantis, Allied and Alternative Medicine, Scisearch and Biosis were searched from their start date to February 1999. MAIN OUTCOME MEASURES: A three-dimensional evaluative framework with related appraisal criteria: (A) craniosacral interventions and health outcomes; (B) validity of craniosacral assessment; and (C) pathophysiology of the craniosacral system. RESULTS: The available research on craniosacral treatment effectiveness constitutes low-grade evidence conducted using inadequate research protocols. One study reported negative side effects in outpatients with traumatic brain injury. Low inter-rater reliability ratings were found. CONCLUSIONS: This systematic review and critical appraisal found insufficient evidence to support craniosacral therapy. Research methods that could conclusively evaluate effectiveness have not been applied to date. PMID- 10709303 TI - Monitoring of liver enzymes in patients treated with traditional Chinese drugs. AB - BACKGROUND: Use of traditional Chinese drug treatment is widespread. While cases of hepatotoxic effects have been reported, little is known about the frequency of such effects. OBJECTIVES: To investigate the frequency, magnitude and course of liver enzyme elevations in patients treated with traditional Chinese drugs. DESIGN: Retrospective study. SETTING: Hospital for traditional Chinese medicine in Germany. PATIENTS: All 1507 patients admitted for inpatient treatment between February 1994 and July 1995. MAIN OUTCOME MEASURE: Number of patients who presented at discharge with a more than 2-fold elevation of alanine amino transferase (ALT) levels (over maximum normal values or elevated admission values). RESULTS: A more than 2-fold elevation of ALT values was observed in 14 patients (0.9%). In 13 cases a causal relationship with Chinese drug therapy seemed possible and, for one patient, likely. However, all patients were also on non-Chinese drug treatment. Follow-up values of ALT within 8 weeks after discharge were normal in 11 patients (six of them had continued to take traditional Chinese drugs) and close to normal in the remaining three. In five patients there were indications for previous liver damage. CONCLUSIONS: In the population and setting studied, clinically relevant liver enzyme elevations occurred in about 1/100 patients treated with traditional Chinese drugs. PMID- 10709304 TI - The costs of treating rheumatoid arthritis patients with complementary medicine: exploring the issue. AB - OBJECTIVES: To measure the marginal costs of providing complementary medicine services (mostly homoeopathy) in outpatient clinics for patients with rheumatoid arthritis (RA) and to illustrate how parameters to which the cost of complementary medicine may be sensitive can be identified. DESIGN: Retrospective, observational costing study. SETTING: The outpatient clinic of the Royal London Homoeopathic Hospital. SUBJECTS: Random sample of 89 patients from the 427 (RA) patients attending outpatient clinics from April 1995 to March 1996. MAIN OUTCOME MEASURES: The marginal costs incurred by the hospital of treating 89 patients attending outpatient clinics and the relative contribution of the different resources to the total costs. RESULTS: The total costs of treating 89 patients were 7,124 Pounds of which 543 Pounds was assumed to be fixed and the remainder variable. The marginal costs of treating additional patients, starting from zero patients treated, are presented. Consultation time (doctors and dietician) contributed to 29% of the total costs, non-conventional drugs contributed to 22% of the total costs. CONCLUSIONS: Understanding the marginal costs of providing complementary care to RA patients will inform the debate over whether these therapies are likely to be cost-effective. In addition, those who would like to explore the practicalities of establishing a service involving complementary medicine will gain an understanding of the likely provider costs. The cost of complementary medicine appears to be most sensitive to the time spent with the patient by the doctor. PMID- 10709305 TI - New approaches to treating pollenosis--a pilot study. AB - OBJECTIVES: To evaluate the effectiveness and safety of a homoeopathic drug combination in the treatment of pollenosis under condition of daily practice. DESIGN: Care review. SETTING: A general practice in Germany. PATIENTS: 35 patients with pollenosis. MAIN OUTCOME MEASURES: Objective and subjective symptoms of acute pollenosis. RESULTS: During an average treatment period of 61 days it was found that 28 of 35 patients experienced an improvement in pollenosis symptoms. The medication was well tolerated thought patients preserved oral to subcutaneous administration. CONCLUSION: Patients receiving the homoeopathic drug combination experienced improvements without side effects. Whether this improvement is due to the medicine or some other factor would need to be determined in a controlled trial. PMID- 10709306 TI - Effect of Traumeel S, a homeopathic formulation, on blood-induced inflammation in rats. AB - OBJECTIVE: To evaluate the activity of Traumeel S (TRS), a homeopathic formulation containing Arnica montana and other plant extracts and minerals on an animal model of traumatic inflammation. DESIGN: TRS and individual components thereof were administered locally to rats 1 h before hind-paw injection with 0.1 ml of homologous blood and the development of oedema was measured over five hours. In each experiment, a control group was treated with saline. MAIN OUTCOME MEASURES: Paw volume of each rat was measured before oedema and 1, 3, and 5 h after oedema induction. Serum levels of IL-6 were determined at hour 5. RESULTS: The decrease of paw oedema, associated with the process of healing, was more rapid in rats treated with TRS (P < 0.05 after 3 h and P < 0.01 after 5 h). Similar effects were also induced by separate injection of most, but not all, TRS ingredients. The efficacy of complete mixture of TRS was higher than the combination of a selection of active components. TRS also reduced oedema development when administered after the oedema induction. The therapeutic effect of TRS was associated with a significant decrease of systemic interleukin-6 production. CONCLUSION: TRS seems to act by speeding up the healing process instead of blocking the development of oedema from the beginning. Moreover, its effect cannot be considered as the 'sum' of its active components and probably a synergistic interaction occurs to determine the final effect. PMID- 10709307 TI - Effects of non-invasive stimulation of acupoints on the cardiovascular system. AB - OBJECTIVE: To study the effect of two non-invasive methods to stimulate acupoints on the cardiovascular system. DESIGN: Blind randomized-controlled trial. SETTING: An experimental setting in a university-affiliated hospital. INTERVENTIONS: The subjects (24 healthy male volunteers) were randomized to receive either an active stimulation consisting of pressure on acupoints (P), an active stimulation consisting of stroking along the meridians (S) or a control stimulation (C). MAIN OUTCOME MEASURES: Data on skin blood flow, arterial pressure, heart rate and EKG were recorded continuously from 20 min before stimulation to 30 min after. RESULTS: In P group there was a decrease in systolic arterial pressure, diastolic arterial pressure, mean arterial pressure, heart rate and skin blood flow. These changes were significantly different from those in C group and, as regards diastolic pressure and mean pressure, also from those in S group. There were no significant differences between S and C groups. CONCLUSIONS: Pressure on acupoints can significantly influence the cardiovascular system. PMID- 10709308 TI - A pilot study to evaluate the effects of floatation spa treatment on patients with osteoarthritis. AB - OBJECTIVE: To conduct a preliminary investigation of the effects on floatation spa therapy on quality of life in patients with osteoarthritis to see if controlled trials are warranted. DESIGN: Uncontrolled clinical trial. SETTING: Private floatation spa therapy centre. PATIENTS: Fourteen patients with chronic osteoarthritis of the weight-bearing joints, of whom four dropped out. INTERVENTION: Six weekly sessions of floatation spa therapy. OUTCOME MEASURES: SF36, AIMS2 and MYMOP quality-of-life questionnaires. MAIN RESULTS: All patients improved. Differences between baseline and discharge scores showed statistically significant improvement for MYMOP, but not AIMS2 or SF-36. CONCLUSIONS: Controlled trials of floatation spa therapy for patients with osteoarthritis are warranted. PMID- 10709309 TI - Evaluation and attributional analysis of an aromatherapy service for older adults with physical health problems and carers using the service. AB - OBJECTIVE: To evaluate an aromatherapy service for older adults with physical health problems and their carers. The aromatherapy service was based in a carer support unit at a hospital in Birmingham. The research question was, 'What types of benefits do clients/carers report from aromatherapy?' DESIGN: Qualitative analysis of interview scripts and written descriptions. SETTING: The interviews were carried out either at the carer support unit, at a connected day centre or at the client/carer's home. PARTICIPANTS: The participants were six clients and four carers who were in contact with the carer support unit and had received aromatherapy from the aromatherapist in the past year. MAIN OUTCOME MEASURES: The participants were interviewed using a semi-structured questionnaire to explore which part of a session they liked best, perceived benefits of aromatherapy and a smell attribution to certain essential oils. The aromatherapist was also asked to write descriptions of her perceived benefits for the clients and carers. The interviews were analysed qualitatively and compared to the aromatherapist's written descriptions. RESULTS: All clients and carers said they benefited from the aromatherapy and felt more relaxed after a session. The qualitative analysis revealed a 70% area of overlap and a 30% 'hidden' area of congruence. The smell analysis revealed individual differences in attribution depending on past experience and expectation of oil presented. CONCLUSION: The findings of this evaluation suggest the aromatherapy service offered was valuable to clients and carers and their perception of its benefits for them were largely congruent with those of the aromatherapist. PMID- 10709310 TI - Guidelines for authors of books and papers on complementary medicine. AB - Writing about complementary medicine, whether intended for book or journal publication, has often suffered from three flaws: inadequate attention to the origin and nature of knowledge claims; careless use of problematic concepts such as 'holistic' or 'natural' and poor application of basic principles of good scholarship. In this paper I present guidelines to promote better writing. With respect to knowledge claims, authors need to be explicit about the origin of the claim; that is, the reasons why they believe it to be true. Many books and papers on complementary medicine are flawed because their authors avoided explaining why they have made particular claims. Claims can be justified by using personal experience or by quoting scientific research. When using personal experience, authors need to: describe the practitioners concerned; use generalities about practitioners with care and be wary when referring to 'classical' or 'traditional' practitioners. When quoting scientific research, authors should: cite references and use these sparingly and specifically; avoid 'secondary sourcing', reliance on abstracts and the referencing of authorities; use 'weasel words' ('may' or 'can') with extreme care; think carefully about causal inferences and take care with laboratory-based research. Many of the concepts found in books and papers on complementary medicine are used rather carelessly. Concepts such as holism or 'natural' medicine, or even the concept of 'complementary' and 'conventional' medicine, are often taken to have simple and obvious meanings. Yet these concepts are extremely slippery and open to differing interpretations, especially if they are used with insufficient care. Authors should apply basic principles of good scholarship by displaying thoroughness and attention to detail; reflecting on the validity of each point made; reflecting on the limitations of their arguments and maintaining a level of disinterestedness. However, authors should avoid trying to sound academic for the sake of it: the use of long words, obscure jargon and dense and lengthy prose does not make a work scholarly. PMID- 10709311 TI - Funding research into complementary medicine: the situation in Britain. AB - This article outlines the theoretical options for research funding for complementary medicine in Britain. Sponsorship from the National Health Service, the Medical Research Council, Medical Research Charities, the industry and private donations constitute the main potential sources. Surveys of the actual funding spent by these sources are evaluated. The data thus generated suggest that funds for research into complementary medicine are not readily available in the UK. Several suggestions are made to ameliorate this situation and the case for 'ring-fenced' funding is put forward, not least on the basis of international experience. PMID- 10709312 TI - Integrating traditional medicine in Japan: the case of Kampo medicines. PMID- 10709313 TI - Schizophrenia is (not simply) a neurodevelopmental disorder. PMID- 10709314 TI - The concept of schizophrenia: pro et contra. PMID- 10709315 TI - Dysregulation of dopamine and pathology of prefrontal neurons: neuroimaging studies in schizophrenia and related animal models. PMID- 10709316 TI - Subthreshold affective disorders: a useful concept in psychiatric epidemiology? AB - OBJECTIVE: In recent years an extensive literature has grown up around the concepts of subthreshold, subsyndromal, minor and brief recurrent affective disorder and their applications in population-based research. The aim of this short review is to examine the definitions and current status of these proposed categories with special reference to depression, and to assess their potential contribution to psychiatric epidemiology. METHOD: A Medline search was carried out for the period 1965-1999, based on the above four terms. Relevant references found in all identified publications were also followed up. RESULTS: In great measure these constructs have been developed as a response to deficiencies in the DSM classification system and to a lesser extent in the ICD. The groups are all defined by having fewer criterial symptoms, or a shorter duration of symptoms, than the 'official' diagnostic categories. Use of these definitions has resulted in widely varying prevalence estimates. CONCLUSION: Improved methods are badly needed for classifying all those persons in the wider community who are in need of medical treatment and help for psychological disorder, but do not satisfy operational criteria laid down in the official guidelines. This cannot, however, be achieved simply by lowering operational thresholds in these systems. Further research on clinical and psycho-social characteristics of the common mental disorders is called for, and in many societies a favourable setting is that of primary health care, where a move towards pragmatic, comprehensive classification of community health problems is already under way. PMID- 10709317 TI - [The use of psychotropic drugs in an Italian psychiatric hospital: a two-year long follow-up study]. AB - OBJECTIVE: Following the introduction of guidelines of rational drug use, the pharmacoepidemiology of psychotropic drugs was investigated in a sample of long stay patients living in a Italian psychiatric hospital. DESIGN: A prospective, longitudinal two-year follow-up study was carried out. Information about sociodemographic and clinical characteristics of the inpatient population, and about medications prescribed, was collected at baseline and after one and two years of follow-up. SETTING: Three wards of the psychiatric hospital of Milan. MAIN OUTCOME MEASURES: Number of patients taking psychotropic drugs, number of patients taking more than one neuroleptic or benzodiazepine, mean neuroleptic dose, psychopathological status according to the Brief Psychiatric Rating Scale (BPRS). RESULTS: 70 patients were recruited and followed for two years. At follow up a reduction in the number of patients taking neuroleptic drugs was recorded, together with a 50% decrease in the number of patients taking more than one neuroleptic. A reduction in the use of depot formulations was in addition shown. Patients taking benzodiazepines decreased of 50%. According to the BPRS, no psychopatological changes were observed during the study. CONCLUSIONS: These data suggest that education in psychopharmacology may guide towards a more rational use of drugs; longitudinal clinical audits should be implemented to monitor everyday practice. PMID- 10709318 TI - [Cognitive-behavioral group treatment of panic attacks disorder: a description of the results obtained in a public mental health service]. AB - OBJECTIVE: This article describes the short-term and at 6 months follow-up results of an intensive cognitive-behavioural group treatment on subjects affected by Panic Disorder with or without Agoraphobia (DSM IV criteria). DESIGN AND SETTING: We studied a group of 22 subjects treated in the public Centro Psico Sociale of Zogno (Bergamo) and valuated them with self-rated instruments inherent the life satisfaction (SF/36) and symptoms andament (PAAAS; MSPS, STAI-X1, STAI X2). The results indicate significant improvements at the end of treatment and at a 6 months follow up. CONCLUSIONS: We are studying the long-term results with others follow up valuations. The most important results, anyway, is the demonstration that also in an Italian public mental health centre, as in many foreign countries, is possible to treat patients affected by Anxiety Disorders with effectual and relatively low cost techniques and that is possible to introduce objective results indicators in the routinary clinical activity. PMID- 10709319 TI - [The use of the Italian version of the Parental Bonding Instrument (PBI) in a clinical sample and in a student group: an exploratory and confirmatory factor analysis study]. AB - OBJECTIVE: The aim of this study was to verify the construct validity of the Italian version of Parental Bonding Instrument (PBI) a questionnaire which estimates the parental style as reported by the son or daughter. METHOD: The questionnaire was administered to a group of 102 students (62 males and 40 females) attending University of L'Aquila and to a sample of 128 patients (76 males and 52 females) consecutively admitted to a psychiatric unit for an index episode. We compared the means of the two factors (care, protection) separately for each parent in the two groups using a t-test for independent samples. After having estimated the internal consistency of items of each scale by calculating Cronbach's coefficient alpha, a factor analysis was performed for students and patients to find the structural factors of the questionnaire; then, we conducted a confirmatory factor analysis of the PBI items, for the students only, to evaluate the fit of the real items to models proposed in the literature. RESULTS: The Italian version of the Parental Bonding Instrument, demonstrated the ability to discriminate between patients and controls; it showed an high internal consistency. The factor analysis identified a two factors solution which accounted for 44.6% and 44.3% of the variance of the mother's and father's PBI scores respectively for the group of students and it identified two factors which accounted for 49.3% and 46.6% of the variance in the group of patients. CONCLUSIONS: The psychiatric patients showed a low "care"-high "protection" confirming an association between the "affectionless control" pattern and psychiatric disorders. PMID- 10709320 TI - Gangliosides: potential diagnostic as well as therapeutic target for cancer. PMID- 10709321 TI - Phototoxicity evaluation--Tetrahymena thermophila as an alternative model. AB - Interest in utilizing an alternative to animal method for toxicological evaluation has received considerable attention due to cost effectiveness and the ethical issues involving animal experimentation. Alternative methods for phototoxicity evaluation are significant because of growing concern over increasing health effects due to stratospheric ozone depletion resulting in an increasing penetration of ultraviolet light-B radiation (UVB, 290-320 nm) which contributes to activation of chemical and biological molecules to potential phototoxic agents. The classic rabbit eye-irritancy test referred to as Draize test has been the subject of severe criticism by animal welfare groups. Dermal toxicity test using guinea pigs and mouse tail phototoxicity test is time consuming and requires a large number of laboratory animals. In photohaemolysis assay some of the phototoxic agents (such as riboflavin) react with the membrane proteins of the erythrocyte. However, in vitro test system using protozoa offers a promising alternative means of phototoxicity evaluation. Our previous studies have demonstrated that synergistic action of photochemically reactive agents and sunlight produces lethal effects to Paramecium but the protozoan has not received serious consideration for use as an alternative model for phototoxicity evaluation. In the present communication we have described the potential application of Tetrahymena as an alternative model to study the radiation-induced changes both in the presence or absence of photoreactive chemical agents. This model is likely to provide scope for studying the biological effects of environmental UVB radiation, DNA damage and defence against oxidative stress. PMID- 10709322 TI - Apoptosis of rat decidual cells: site specific initiation and related biochemical changes. AB - The artificially induced rat deciduoma serves as a model to study cellular changes associated with implantation in the endometrium. The stromal cells differentiate to form two types of decidual cells and are restricted to specific anatomical sites of the uterus. Programmed cell death starts in the antimesometrial area and expression of glutathione-S-transferase, an antioxidant enzyme, enhances in these cells as the deciduoma enters the regressive phase. The enzyme activity is significantly high compared with that of mesometrial decidual cells. Similarly, lipid peroxide content of antimesometrial decidual cells is high during this phase. DNA fragmentation, a feature of cells undergoing programmed cell death, is initiated in the antimesometrial area during regression of deciduoma. PMID- 10709323 TI - Hepatoprotective effects of Liv-52 on ethanol induced liver damage in rats. AB - The mechanism of protective effects of Liv-52, a multiherbal hepatoprotective drug, on ethanol induced hepatic damage has been investigated. The results indicate that Liv-52 treatment prevents ethanol induced increase in the activity of the enzyme gamma-glutamyl transpeptidase. Concomitantly there was also a decrease in ethanol accentuated lipid peroxidation in liver following Liv-52 treatment. The activity of antioxidant enzymes; superoxide dismutase, glutathione peroxidase and the levels of glutathione were decreased following ethanol ingestion. Liv-52 treatment was found to have protective effects on the activity of superoxide dismutase and the levels of glutathione. The results obtained from the study indicate hepatoprotective nature of Liv-52 which might be attributed to its ability to inhibit lipid peroxidation. PMID- 10709324 TI - Male accessory gland secretory proteins in nasuta subgroup of Drosophila: nature and SDS-PAGE patterns. AB - Male accessory gland secretory proteins in seven members of Drosophila nasuta subgroup were analyzed by SDS-PAGE in combination with different staining techniques such as CBB-R250, Silver, PAS, PAS-silver and zinc-imidazole reverse staining. Based on coomassie blue patterns the protein fractions could be classified in to 3 major groups namely group I, group II as well as group III; with high molecular weight fractions falling into group I and low molecular weight fractions into group III. All the three groups of fractions are post translationally modified by way of glycosylation and group III fractions are found to be highly glycosylated. Fractions of groups I and II when localized with silver stain and group III fractions when localized with PAS-silver stain appear yellow; suggesting that they are sialoglycoproteins. A 40 kD fraction of group II shows differential staining property with zinc-imidazole stain in closely related species namely D. n. nasuta and D. n. albomicans. Analysis of this protein fraction in F1 males of an interspecific cross revealed that it is synthesized by X-chromosomal gene. PMID- 10709325 TI - Epichlorohydrin induced biochemical changes in the rose-ringed parakeet, Psittacula krameri Scopoli. AB - Intraperitoneal administration of epichlorohydrin (ECH) at the dose level of 20 and 50 mg/kg body weight inhibited spermatogenesis in the testis of parakeet during breeding season. A total load of 60 mg/kg body weight of ECH given on 3 consecutive days proved to be lethal. Testicular proteins, nucleic acids (DNA and RNA), phospholipids and acid phosphatase activity were decreased, while the lipids, total cholesterol and alkaline phosphatase activity increased after ECH administration. The results suggest that the testicular atrophy caused by ECH was associated with an alteration in the activities of macromolecules and enzymes related to specific events of spermatogenesis. PMID- 10709326 TI - Spectrophotometric analysis of resazurin reduction test and semen quality in men. AB - The resazurin reduction test (RRT) was performed on semen samples obtained from 225 untreated subfertile and 10 pregnancy confirmed fertile males. The results of RRT were determined visually using resazurin colour chart and again the extent of resazurin reduction in each sample was additionally read by spectrophotometer to assess the quality of samples. Absorption spectra was scanned for resazurin and resorufin and two most sensitive wavelengths (572 and 600 nm) were selected. The ratio of the two optical densities was used as a probe to discriminate the various grades of semen samples. In azoospermic samples, RRT ratio ranges from 0.7 to 1.16, in oligoasthenozoospermic samples from 1.10 to 1.35, in oligozoospermic samples from 1.5 to 2.0 (characterised visually as grades from 1 4) in normozoospermic and proven fertile samples from 2.25 to 5.9 (characterized visually as grades from 5 to 11). The highest correlation of RRT ratio was observed with sperm motility (r = 0.889, P < 0.001), followed by concentration (r = 0.848, P < 0.001) morphology (r = 0.660, P < 0.001) and viability (r = 0.544, P < 0.01). The test ratio had a positive predictive value of 95% for a sperm concentration of > 20 x 10(6)/ml and motility > 40% and a negative predictive value of 90% for a sperm concentration of < 20 x 10(6)/ml and motility < 40%. Therefore the evaluation of RRT results using spectrophotometric ratio method may provide a tool for obtaining a wider range of seminological diagnosis more accurately than the routine semen analysis. It is suggested that the method is simple and reliable, it can be performed in any andrology laboratories. PMID- 10709327 TI - Cloning and expression of a nitroaryl reductase gene from Streptomyces aminophilus strain MCMB411 in E. coli JM109 and Streptomyces lividans TK64. AB - A partial genomic library was prepared in E. coli JM109 using pBR322 as vector and 2.4 kb Sau 3A I chromosomal fragment, encoding a nitroaryl reductase (nbr A) gene, from Streptomyces aminophilus strain MCMB 411. From the library, 2.4 kb fragment was recloned in E. coli JM109 and S. lividans TK64 using pUC18 and pIJ702 as vectors respectively. The recombinant plasmids pSD103 and pSD105 expressed the reductase gene and exported the enzyme in periplasmic space of E. coli and in cytoplasm of S. lividans TK64. The proteins expressed by E. coli and S. lividans had the same molecular mass (70 kD) as that expressed by parent strain, which suggested that the enzyme was processed similarly by all strains. Activities of the enzymes cloned in E. coli JM109 and S. lividans TK64 containing recombinant plasmids pSD103 and pSD105 respectively were optimum at 30 degrees C and pH 9 and requirement of cofactors was same as that of the parent strain. PMID- 10709328 TI - Identification of volatile compounds from cheek glands of lesser bandicoot rats and assessment of behavioural response for identified compounds. AB - Check gland secretions were collected from sexually mature male and female rats to identify the volatile compounds using gas chromatography linked mass spectroscopy with a modified head space technique. Twenty one volatile fractions including alkanes, aliphatic acids, esters and alcohols were found. Three compounds viz. 3-octen-1-ol (E) (I), cyclopentane undecanoic acid (II) and 2,4,6,8 tetramethyl-l-undecane (III) were present in very high concentrations in both male and female rats. In addition, the estrus female contained another compound, L-alanine, 1,1-dimethyl ethyl ester. On the basis of odour preference tests the compounds I, II and III were found to attract both male and female of the homotypic species. In contrast, the fourth compound present in the female attracts the opposite sex. The results suggest that the volatile fractions have unique function and the level of attraction varies from compound to compound. PMID- 10709329 TI - Experimental amebiasis: molecular weight analysis of Entamoeba histolytica antigens recognized by IgG antibodies. AB - The reactivity of sera from experimentally infected animal was studied from 5-60 days postinoculation to determine which of the E. histolytica antigens are recognized frequently to infection. Crude extract of E. histolytica trophozoites was used and sera were examined by immunoelectrotransference assay. It was observed that sera recognized polypeptide with 70 kDa molecular mass after 15 days postinoculation onward and later 14 to 24 polypeptide with molecular weight of 110-22 kDa were recognized. All the amebic antigens (polypeptides) could be recognized by sera till 60 days postinoculated animals. Significance of expression of different amebic polypeptides in terms of pathogenesis needs further investigations. PMID- 10709330 TI - Protective effect of capsaicin on the osmotic fragility of human erythrocytes. AB - Capsaicin exerts a stabilizing effect on erythrocytes making them more resistant to lysis under hypotonic stress. The protective action of capsaicin on osmotic fragility (OF) was not receptor mediated since no dose responsive effect was observed. The results suggest that this protective effect of capsaicin on OF is due to a direct interaction of capsaicin with the erythrocyte membrane rather than due to any alteration in the intracellular metabolism of erythrocytes. PMID- 10709331 TI - Evaluation of serological status of rubella & mumps in children below five years. AB - A cross-sectional study was carried out on 321 serum samples to detect rubella and mumps antibodies in children below five years and to assess the optimum age for immunization against rubella and mumps. Seropositivity to rubella was 33.3 per cent in children below nine months, 16.9 per cent at 9-12 months and 25.5 per cent by two years. Mean antibody levels for rubella were low at nine months to one year and remained so till five years of age. Similarly, seropositivity for mumps was 53.3 per cent below nine months, 20.3 per cent at 9-12 months and 40 per cent by two years. Mean antibody levels for mumps were low between nine months to two years with a slight rise by five years. The findings suggest that a large majority of children are at risk by the age of nine months in our population and the measles, mumps and rubella (MMR) vaccination at this age may be most beneficial. PMID- 10709332 TI - The clearance of tubercle bacilli & mycobacterial antigen vis a vis the granuloma in different organs of guinea pigs. AB - In the present study, an attempt was made to define the relationship of intact tubercle bacilli and/or their antigenic fragments to a granuloma in the guinea pig in order to distinguish an active from a resolving granuloma. In one set of animals, granuloma was induced in the skin by injecting heat-killed Mycobacterium tuberculosis intradermally and in another set, granuloma was produced in the lung and spleen by injecting live M. tuberculosis intramuscularly. The animals were sacrificed at various time points and skin, lung and spleen from the two groups were subjected to histological examination for the presence of granuloma, bacilli and antigenic fragments. In the dermal lesion, intact acid fast bacilli were cleared first by day 42 followed by the removal of their antigenic fragments by day 63 and finally by day 84, the granuloma had resolved completely. In the guinea pigs infected with live M. tuberculosis, removal of the bacilli followed by the clearance of antigen was observed. Though the granuloma itself did not subside completely in these animals, it was found that there was a reduction in congestion and oedema of the granulomatous area. It is concluded from the results that the demonstration of antigen at the site of lesion may be potentially useful to discriminate between a persisting and a resolving tuberculous granuloma. PMID- 10709333 TI - Role of yeasts as nosocomial pathogens & their susceptibility to fluconazole & amphotericin B. AB - A total of 213 and 208 yeasts were isolated as nosocomial pathogens from various infected specimens during 1996 and 1997 respectively. Yeasts ranked fifth among uropathogens in both the years and from eighth to eleventh in other specimens. Increasing trend in nosocomial urinary tract yeast infection (11.9 in 1996 to 12.6 in 1997) and decreasing trend in wound and other infections (5.1 in 1996 to 2.9 in 1997) per 1000 patients' discharges were observed; blood stream infection remained unchanged (2/1000 discharges) in both the years. Eighty two (41 from each year) randomly selected yeasts were identified to species level following standard protocol and tested for antifungal susceptibility against fluconazole and amphotericin B by reference broth macrodilution technique and agar dilution (AD) method. The frequency of various yeast species identified was Candida albicans 39 (47.6%), C.tropicalis 29 (35.4%), C. krusei 4 (4.9%), C. glabrata 3 (3.7%), C. zeylanoides 2 (2.4%), C. guilliermondii 2 (2.4%), one strain (1.2%) each of C. kefyr, C. parapsilosis, and Trichosporon beigelii. Resistance to fluconazole (MIC > or = 64 micrograms/ml) as per NCCLS criteria was observed in 2 Candida sp. (2.4%). Significantly higher number of non-albicans Candida sp. (8/43; 18.6%) had MIC > 8 micrograms/ml as compared to C. albicans (2/39; 5.1%) (P < 0.05). Only one strain of C. tropicalis had MIC 8 micrograms/ml to amphotericin B and none had MIC > 8 micrograms/ml. Agreement between the reference and the AD methods for fluconazole was 88 per cent and for amphotericin B was 94 per cent. The present study indicates that Candida sp. are emerging as important nosocomial pathogens and the tendency of yeasts to develop resistance to antifungal agents appears to be a challenge for patient management. PMID- 10709334 TI - Diagnostic accuracy of rapid enzyme linked immunosorbent assay for the diagnosis of human hydatidosis. AB - Rapid enzyme linked immunosorbent assay (ELISA) was compared with the standard ELISA and indirect haemagglutination (IHA) techniques for the diagnosis of human hydatidosis. Eighty nine serum samples including 17 from hydatidosis patients (10 surgically confirmed and 7 clinically suspected), 50 from patients with other parasitic diseases and 22 samples from normal healthy individuals were analysed for anti-hydatid IgG antibodies using sheep hydatid cyst fluid antigen. The sensitivity and specificity respectively was found to be 82.3 and 100 per cent by rapid ELISA; 88.23 and 90.27 per cent by standard ELISA and 70.58 and 100 per cent by IHA technique. No cross reactions were observed with rapid ELISA technique using samples from cysticercosis and amoebiasis patients and normal healthy controls. The present study indicates that rapid ELISA can easily be performed in place of the standard ELISA for the serodiagnosis of human hydatidosis with the advantage of minimising reporting time and manpower hours. PMID- 10709335 TI - A modified radiographic method for estimating segmental colonic transit time in subjects with rapid gut transit. AB - In this study on Indian subjects, the single X-ray method was assessed for its reliability in measuring the transit of particulate matter through the colon, and if inaccurate a suitable and simple alternative was to be devised. Radio-opaque markers were serially followed in 20 normal male volunteers. This was done by three 12 hourly radiographs and by stool collection to determine the transit time through the colon and its segments. It was compared with similar parameters calculated from the same data using one radiograph and three combinations of two radiographs each. The mean +/- SD colonic transit time determined by using three X-rays was 18.0 +/- 6.6 h which agreed well with the mean mouth-to-anus transit time of 24.2 +/- 6.8 h (mean +/- SDdiff = -6.2 +/- 2.9). When two of the three X rays were used in various combinations, the best results were obtained with the method including radiographs at 12 and 36 h. Parameters calculated from a single radiograph done 36 h after the ingestion of makers showed lesser agreement with the results of the three radiograph method. Therefore in subjects with rapid gut transit, the simplified method for estimating the colonic and segmental transit times using a single X-ray is inaccurate. Using two radiographs enhances the accuracy. PMID- 10709336 TI - Effect of banana on cold stress test & peak expiratory flow rate in healthy volunteers. AB - The effect of banana on cold stress induced hypertension, peak expiratory flow rate and plasma ACE activity in healthy human volunteers was tested. Systolic blood pressure (P < 0.005), diastolic blood pressure (P < 0.025) and mean arterial blood pressure (P < 0.005) were significantly decreased during cold stress after banana treatment compared to controls subjected to cold stress. There was no significant changes in heart rate and peak expiratory flow rate but only significant decrease in plasma ACE activity after banana treatment. Banana decreased the rise of systolic blood pressure and diastolic blood pressure in healthy volunteers subjected to cold stress test without much effect on heart rate and peak expiratory flow rate. PMID- 10709337 TI - Current and potential pulp therapies for primary and young permanent teeth. AB - OBJECTIVES: This paper aims to alert the dental practitioner to the rapidly evolving therapies for treating the pulps of primary and young permanent teeth. DATA SOURCES: Experimental research on animals, clinical studies and case reports. STUDY SELECTION: Indirect pulp capping, direct pulp capping, pulpotomies, and pulpectomies are standard procedures for treating primary teeth. However, direct pulp capping, heretofore not very successful, is being revisited. Based on studies in animals and clinical findings in humans, there has been a movement in pediatric dentistry to find alternatives to formocresol and calcium hydroxide for pulpotomy therapy. Venues range from eradication by cautery to the possibility of healing with growth factors. New studies with iodoform paste for pulpectomies are confirming the success rates of previous publications. The new dental adhesives are being tested as agents for direct pulp capping, as well as partial and complete pulpotomy protocols. CONCLUSIONS: More thought is being given by clinicians to preserving pulp, either through more ambitious indirect pulp therapy or partial pulpotomy. Formocresol and calcium hydroxide pulpotomies, while still popular, may soon be challenged by other chemical treatments, electrocautery or stimulation of reparative dentine by growth factors. Iodoform pastes are promising easier and more successful pulpectomy therapy. Total etch direct bonding materials could soon transform direct pulp capping, as well as partial and complete pulpotomy protocols. PMID- 10709338 TI - Porcelain veneers: a review of the literature. AB - OBJECTIVES: Porcelain veneers are steadily increasing in popularity among today's dental practitioners for conservative restoration of unaesthetic anterior teeth. As with any new procedure, in vitro and in vivo investigations are required to assess the ultimate clinical efficacy of these restorations. The current literature was therefore reviewed in search for the most important parameters determining the long-term success of porcelain veneers. DATA SOURCES: Laboratory studies focusing on parameters in prediction of the clinical efficacy of porcelain veneers such as the tooth preparation for porcelain veneers, the selection and type of the adhesive system, the quality of marginal adaptation, the resistance against microleakage, the periodontal response, and the aesthetic characteristics of the restorations have been reviewed. The clinical relevance of these parameters was then determined by reviewing the results of short and medium to long-term in vivo studies involving porcelain veneers performed during the last 10 years. CONCLUSIONS: The adhesive porcelain veneer complex has been proven to be a very strong complex in vitro and in vivo. An optimal bonded restoration was achieved especially if the preparation was located completely in enamel, if correct adhesive treatment procedures were carried out and if a suitable luting composite was selected. The maintenance of aesthetics of porcelain veneers in the medium to long term was excellent, patient satisfaction was high and porcelain veneers had no adverse effects on gingival health inpatients with an optimal oral hygiene. Major shortcomings of the porcelain veneer system were described as a relatively large marginal discrepancy, and an insufficient wear resistance of the luting composite. Although these shortcomings had no direct impact on the clinical success of porcelain veneers in the medium term, their influence on the overall clinical performance in the long term is still unknown and therefore needs further study. PMID- 10709340 TI - Ultrasonic measurement of enamel thickness: a tool for monitoring dental erosion? AB - OBJECTIVES: Wear of dental hard tissues, e.g. dental erosion, is reported to be a growing problem. A non-destructive measurement of enamel layer thickness would provide the opportunity for both early diagnosis, and longitudinal measurement of progressive enamel loss. It was the aim of this study to investigate the potential of ultrasonic pulse-echo measurements for the enamel thickness measurement. METHODS: Nine extracted human incisor teeth were selected and stored in physiological saline. Mesial and distal tooth parts were removed, resulting in a central tooth slice of about 2 mm thickness. Where possible three buccal, and one palatal measuring sites were selected and indicated by pencil marks on one of the section planes. Ultrasonic pulse-echo measurements were made at each site using a Panametrics 25DL thickness gauge (Panametrics, Waltham, MA, USA), using a perspex delay line transducer (15 MHz) and glycerine coupling medium. Ultrasonic measurements were validated by measuring the thickness of the enamel layer at the marked side of the tooth slices with a light stereomicroscope at 120 x magnification. Two observers performed independent measurements. RESULTS: Limits of agreement for measurements by two observers (n = 42) were -0.09 and 0.09 mm. Measurements performed at 21 degrees C and 34 degrees C were not significantly different, as analysed by paired Student's t-test (p = 0.19). Pearson's correlation coefficient between ultrasonic and microscopic measurements was 0.90. Analysis of all measurements from both observers at both temperatures yielded a sound velocity in enamel of 6.5 x 10(3) m/s (standard error 0.1 x 10(3) m/s). CONCLUSIONS: It was concluded that the ultrasonic measurement of the enamel thickness is feasible without enamel preparation. PMID- 10709339 TI - Scanning electron microscopic observations of human dentine after mechanical caries excavation. AB - OBJECTIVES: The structural integrity and surface characteristics of dentine remaining after caries excavation may be relevant to the subsequent bonding of adhesive restorative materials to the prepared cavity. This in vitro investigation aimed to analyse the different surface characteristics of the dentine cavity floor created after preparation using five different mechanical and chemo-mechanical methods of excavation: hand excavation, slow-speed bur, sono abrasion, air-abrasion and Carisolv gel. METHODS: Ten cavities were prepared using each excavation method in extracted teeth with occlusal carious lesions. Epoxy resin replicas of the 50 cavities were manufactured from silicone impressions and then analysed using secondary electron scanning electron microscopy (SEM) to ascertain the surface characteristics of the dentine at the cavity floor. RESULTS AND CONCLUSIONS: Results from the 50 cavities examined suggested that each alternative excavation technique produced a different and characteristic dentine surface. Carisolv gel was the only method examined that consistently removed the smear layer during excavation to leave exposed dentine tubules at the end of cavity preparation. PMID- 10709341 TI - Monkey pulpal response and microtensile bond strength beneath a one-application resin bonding system in vivo. AB - OBJECTIVES: The aim of this in vivo study was to investigate the biocompatibility and microtensile bond strength of a one-application resin bonding system. METHODS: Class V cavities were prepared on the facial surfaces of 36 intact monkey teeth, and the cavities were restored with an experimental one-application resin bonding system (TOF-1; Tokuyama Corp., Tokuyama, Japan) and a hybrid resin composite (PALFIQUE ESTELITE; Tokuyama Corp., Tokuyama, Japan). Histopathological changes of the restored teeth were evaluated at 3, 30 and 90 days after operation (N = 10). Microtensile bond tests were performed at 3 and 90 days after operation (N = 10). RESULTS: Only two of 30 pulps showed a slight inflammatory cell infiltration. There were no statistically significant differences in the incidence of slight inflammatory cell infiltration among time periods. Bacterial penetration along the cavity walls could not be detected in any specimen. The mean microtensile bond strength at 3 days after operation was 20.6 MPa, and that at 90 days was 14.9 MPa. Differences in bond strengths between the 3 day specimen and the 90 day specimen were statistically significant (p < 0.05). CONCLUSIONS: The one-application resin bonding system exhibited acceptable biologic compatibility to the monkey pulp. Although there were statistically significant differences in bond strengths between the 3 day specimen and the 90 day specimen, this material provided a hermetic seal, eliminating bacterial microleakage. PMID- 10709342 TI - Bacterial morphotypes and early cellular responses in clinically infected and non infected sites after combination therapy of guided tissue regeneration and allograft. AB - OBJECTIVES: To compare the bacterial morphotypes and early cellular responses in periodontally treated sites with and without pus formation after a combination of guided tissue regeneration (GTR) and allograft therapy. METHODS: 45 subjects with 80 sites having periodontal lesions with moderate to deep pockets and angular bone defects participated. 28 treated sites in 25 patients were included in the studies. 14 sites suffered from symptoms and signs of infection with pus formation during the healing period were assigned to the pus (P) group. Another 14 sites had asymptomatic healing and were assigned to the non-pus (NP) group. The GTR membranes were retrieved 4-6 weeks after surgery and processed for SEM examination. The bacterial morphotypes on the membranes were observed and photographed. Bacterial adhesion score (BAS, 0-5) and the presence of leukocytes and fibroblasts were estimated from photographs. RESULTS: The results showed that large numbers of bacteria (high BAS) were present on both sides of the coronal 2/3 of the membrane in both groups, irrespective of clinical conditions. At the apical 1/3 of the membrane, moderate numbers of bacteria were still found on the outer side in the P group. The BAS of rod-shaped bacteria were significantly higher in the P group than that of the NP group on the outer coronal 2/3 of the membrane. The frequency of the presence of fibroblasts (18.5%) at the apical 1/3 of the inner (tooth facing) side of the P group was much lower than that of the same location (28.6-29.6%) in the NP group. The presence of leukocytes and fewer numbers of fibroblasts on the GTR membrane were associated with greater BAS for rod- and filament-shaped bacteria. CONCLUSIONS: GTR membranes are commonly colonized by oral bacteria during retention, even on uncomplicated and tissue covered portions. The overt infection clinically (pus group) of the membrane allograft treated sites is associated with a significantly elevated BAS of rod shaped bacteria, and may be closely related to the occurrence of its adverse early healing responses (inflammation, pus formation, fewer fibroblasts and greater accumulation of leukocytes). PMID- 10709343 TI - Ultraviolet radiation and health: from hazard identification to effective prevention. AB - The increased exposure to ultraviolet (UV) radiation due to ozone depletion is one of the most serious global health problems. The UV exposure is known to cause skin carcinoma, cataract and deteriorated immune function, but for countries like Japan, the magnitude of health effects of UV radiation is yet to be elucidated. The International Workshop on the Health Effects of Ultraviolet Radiation was held in Tokyo, Japan, on February 17-19, 1999, in attempts to visualize the size of this problem and to identify better solutions. Through this workshop, several lines of scientific evidence were provided, which clearly show that the risk of cataract and skin cancer among people living in Japan increases with the increasing level of sun exposure. We must seek, therefore, the extent to which the UV exposure of given intensity causes adverse health effects in Japanese population. Through the workshop, the importance of preventive measure was confirmed. The scientific basis of prevention is, of course, the knowledge of dose-response relationship and the current exposure status in Japanese population. It is hoped that the communications between researchers in Japan and other countries are strengthened through this workshop. PMID- 10709344 TI - International workshop on the health effects of ultraviolet radiation opening remarks. PMID- 10709345 TI - Sun exposure, skin cancers and related skin conditions. AB - Skin cancer is the most commonly occurring cancer in humans. Solar keratoses are related benign tumours that are at least ten times commoner than skin cancers and photoageing of the skin is still more common. Descriptive studies show that incidence rates of the main types of skin cancer, basal cell carcinoma, squamous cell carcinoma and melanoma are maximal in populations in which ambient sun exposure is high and skin (epidermal) transmission of solar radiation is high, suggesting strong associations with sun exposure. Analytic epidemiological studies confirm that exposure to the UV component of sunlight is the major environmental determinant of skin cancers and associated skin conditions and evidence of a causal association between cumulative sun exposure and SCC, solar keratoses and photodamage is relatively straightforward. Results for BCC and melanoma are complicated by several factors including the existence of subgroups of these diseases which do not appear to be caused by sun exposure yet have been included in most aetiological studies to date. Complementary to epidemiological data is the molecular evidence of ultraviolet (UV) mechanisms of carcinogenesis such as UV-specific mutations in the DNA of tumour suppressor genes in skin tumours. With increased UV irradiation resulting from thinning of the ozone layer, skin cancer incidence rates have been predicted to increase in the future- unless, as is hoped, human behaviour to reduce sun exposure can offset these predicted rises. PMID- 10709346 TI - Incidence of skin cancers and precancerous lesions in Japanese--risk factors and prevention. AB - An examination of the occurrence of skin cancers and precancerous lesions among residents of Kasai City (34 degrees 56' N) since 1992, and of le-island (25 degrees 10' N) since 1993, has been conducted to characterize the prevalence and incidence of skin cancers in Japanese people and to evaluate risk and preventive factors. The mean prevalence of actinic keratosis (AK) in residents of Kasai City and le-island was 203.33 and 756.26, respectively, indicating that twice the dose of UVB radiation causes a 3-4 fold higher incidence of AK, although life styles, including types of occupations, differ in these two locations. Working outdoors, having skin type I and/or a history of severe sunburns during childhood were found to be important risk factors, while the use of cosmetics after 20 years of age was a protective factor, for AK and possibly for skin cancers. Further, sunscreen use among males over 60 years of age in Kasai City from 1994 through 1998 suggested that sunscreen use may reduce AK development in older people. Four and 12 cases of skin cancers were found in residents of Kasai City (from 1992 to 1997) and on le-island (from 1993 to 1998), respectively. These numbers are too small to establish the prevalence of skin cancer in Japanese, but indicate that people living in areas of higher ambient solar radiation have a higher incidence of skin cancer. This epidemiological study strongly indicates that sun protection is the major modality to reduce sun-induced cutaneous tumors in Japanese. PMID- 10709347 TI - Geometrical assessment of ocular exposure to environmental UV radiation- implications for ophthalmic epidemiology. AB - Epidemiological studies of the influence of environmental ultraviolet radiation (UVR) in the development of cataract, pterygium, droplet keratopathies and age related macular degeneration have produced inconsistent findings. The lack of consistent results may be due largely to either incomplete or erroneous estimates of outdoor UV exposure dose. Geometrical factors dominate the determination of UVR exposure of the eye. The degree of lid opening limits ocular exposure to only those rays entering at angles near the horizon. Clouds redistribute overhead UVR to the horizon sky. Mountains, trees and building shield the eye from direct sky exposure. Most ground surfaces reflect little UVR. The result is that the highest UVR exposure occurs during light overcast where the horizon is visible and ground surface reflection is high. By contrast, exposure in a high mountain valley with green foliage results in a much lower ocular dose. Other findings of these studies show that retinal exposure to light and UVR in daylight occurs largely in the superior retina. PMID- 10709348 TI - Epidemiological studies on UV-related cataract in climatically different countries. AB - Cataract epidemiological surveys applying objective judgement through lens images in the climatically different places of Noto and Amami, Japan, Singapore and Reykjavik, Iceland yielded several significant results about the influence of solar UV. 1) The percentage of transparent and of lens opacification was significantly higher in the Reykjavik subjects than in the Singaporeans. 2) The percentages including early changes were higher in Amami and Singapore than in Noto and Reykjavik. 3) Progressed lens opacification was highest in Singapore. While the main type of lens opacification was cortical in Noto and Reykjavik, that of Singapore was nuclear. 4) A significant correlation between cortical opacification and the history of time spent outdoors was noticed. The UV risk for formation and/or progression of cortical opacification should be acceptable from the epidemiological standpoint. PMID- 10709349 TI - UV damage to the eye lens: further results from animal model studies: a review. AB - UV irradiation has the potential to induce the development of lens opacities. This has been demonstrated since long with animal experiments. Unfortunately these animal cataracts did not explain or elucidate the epidemiological observation that the frequency of human cataracts--such as the so called senile cataract--is remarkably higher in regions with increased cosmic UV irradiation or in the population being in close professional contact with UV-irradiation. The main problem was that the type of UV induced animal cataracts differs remarkably with respect to onset, localization of the opacity, size and its timely progression from the cataract classes observed in human. The research of the last 10 years comes to the conclusion that beside the direct (acute) damage--as seen in animal studies due to high UV dosages--we have to realize a syn- or co cataractogenic potential of UV irradiation even below the threshold dose which is able to accumulate in the lens and to initiate together with other risk factors (chronic damage) the opacification of the lens. The mechanism for the animal cataract and the human cataract (with an UV risk participation) are different. The epidemiological research about cataract frequency in different regions of the world have to take into account that UV irradiation--even below a threshold dose- is a possible risk among the multifactorial pathogenesis of human cataract. PMID- 10709350 TI - Possible effects of sunlight on human lymphocytes. AB - The human population is exposed to both the ultraviolet A (UVA) and B (UVB) regions of the solar spectrum. UVB induces mainly dipyrimidine photoproducts in DNA by a direct photochemical mechanism, whereas UVA is absorbed by other cellular constituents and induces mainly oxidative damage indirectly. The proportions of the different dipyrimidine photoproducts, and the ratio of dipyrimidine to oxidative damage depend on the exact spectral output of a UV source. Irradiation of human epidermal keratinocytes induces release of cytokines, with cyclobutane pyrimidine dimers playing a significant role in the process. These cytokines may then modulate the activity of cells of the immune system. Freshly isolated human lymphocytes are exquisitely sensitive to UVB irradiation, because of their low deoxyribonucleotide pools. They also have a separate defect in removal of cyclobutane pyrimidine dimers from their DNA. We have observed that frequencies of mutations at the hprt locus in human T lymphocytes and translocations involving the bcl2 locus in B-lymphocytes appear to be associated with sunlight levels over the period before the blood sample was taken. This may be an indirect cytokine-mediated effect, and may be relevant to the possible link between non-Hodgkin's lymphoma and sunlight. On the other hand, sunlight can have beneficial effects, and may protect against autoimmune diseases including type I diabetes and multiple sclerosis. PMID- 10709351 TI - Mutations in cancer genes of UV-induced skin tumors of hairless mice. AB - Ultraviolet (UV) radiation is a very common carcinogen in our environment. Epidemiological data on the relationship between skin cancers and ambient solar UV radiation are very limited. Hairless mice provide the possibility to study the process of UV carcinogenesis in more detail. Experiments with this animal model have yielded quantitative data on how tumor development depends on dose, time and wavelength of the UV radiation. In addition, at the molecular level the interactions between UV, specific cancer genes-like the Ras oncogene family and the p53 tumor suppressor gene, together with the role of DNA repair in this process have been addressed recently. In wildtype hairless mice mutations in the p53 gene are clearly linked to UVB but not to UVA radiation. Furthermore, the p53 alterations seem to be essential early in tumor development. However, in Xpa deficient mice this dependency on p53 alterations appeared to be different as is the tumor type induced by UVB. Research using genetically modified hairless mice should enable us to further unravel the mechanisms of UV-induced skin cancer. PMID- 10709352 TI - Somatic cell mutation induced by sunlight in Drosophila. AB - There is ample epidemiological evidence showing that sunlight can cause skin cancer in the human. In experimental studies, simulated sunlight or UV lamps are used for demonstrating carcinogenesis and other biological effects. Little studies, however, have been performed using natural sunlight itself. In this work, we have examined the mutagenicity of natural sunlight in Drosophila. The Drosophila wing spot test is useful to detect somatic cell mutations. Third instar larvae in petri dishes were exposed to sunlight (ultraviolet region with < 290 nm wavelength cut off by a plastic cover) in the yard of Okayama University campus (north latitude: 34 degrees 39', east longitude: 133 degrees 55'). The sunlight was mutagenic in Drosophila larvae and produced pyrimidine dimers in their DNA. In the observed mutagenicity, there was dependence on the exposure period and UV fluence. During the two-year monitoring, the highest induction of mutant spot observed was 1.98 total spots/wing on June 25, 1998, and the lowest was 0.64 on December 29, 1998, while non-exposure spontaneous spots were 0.29 and 0.32 on these days, respectively. Thus, solar radiation was mutagenic both in summer and in winter. PMID- 10709353 TI - Cytotoxicity and mutagenicity of UVB assessed using cultured rat fibroblast. AB - A retroviral vector carrying both positive (neo) and negative (herpes simplex virus thymidine kinase or HSV-tk) selection markers was constructed as a substrate for mutational assay in mammalian cells. Using a population of rat fibroblast cells carrying a single copy per cell of retroviral DNA randomly integrated in their chromosomes, we examined the cytotoxic and mutagenic activities of ultraviolet light (UV) at four wavelengths (254, 290, 300, and 320 nm). The action spectra for these activities are similar to some of the previously reported spectra for photochemical DNA modifications, erythema, cell killing, and mouse skin carcinogenesis, except at 290 and 320 nm. At 290 nm, no significant mutagenicity was observed. At 320 nm, both cytotoxic and mutagenic activities were 10 times higher than the values expected from the absorption spectrum for DNA and the action spectrum for bacterial inactivation and mutagenesis. Structural comparison of some of the HSV-tk mutants obtained after irradiation with 300 and 320 nm UV revealed partially different patterns of mutation specificity, suggesting the involvement of multiple molecular mechanisms in the genotoxicity associated with this range of UV. PMID- 10709354 TI - Intrinsic and extrinsic biomarkers for the assessment of risks from environmental UV radiation. AB - In the last decades the knowledge of the effects of UV radiation on human health, especially in skin cancerogenesis, but also in immunsuppression, photoaging, eye damages, has enlarged strongly. The increasing solar UV radiation and changes in life style strengthen the necessity to identify and quantitate intrinsic biomarkers which are indicative for the individual UV susceptibility and the accumulated individual UV burden. For the risk assessment of potentially deleterious UV effects extrinsic biomarkers have to be developed and tested as personal biological UV dosimeters. One example for such a well characterized biological UV dosimeter is the DLR-biofilm which consists of spores of the bacterium Bacillus subtilis as UV sensitive target. PMID- 10709355 TI - UV-induced changes in the immune response to microbial infections in human subjects and animal models. AB - Exposure to UV is a recognised risk factor for skin cancer and it also induces immunosuppression to a variety of antigens encountered following the irradiation. The latter property has been demonstrated in rodent models of infections with the microbial agents including viruses, bacteria, protozoa and helminths. In the majority of cases the severity of the symptoms and the microbial load in the host are increased as a result of the immunomodulation. UV can also affect the pathogenesis of some natural microbial infections of human subjects, such as causing recrudescence of herpes simplex virus and contributing to the oncogenic potential of papillomaviruses. Sufficient data have been generated from the animal models to construct a risk assessment in humans for suppression of microbial immune responses induced by sunlight exposure. This estimation requires verification from epidemiological studies and from further work to assay modulation in human immunity to particular pathogens experienced before and after the UV radiation. PMID- 10709356 TI - Increased susceptibility of mice to malarial infection following UV-B irradiation. AB - Using a mouse model, we examined whether UV-B was a risk factor for malarial infection. Two mouse strains, susceptible (BALB/c) and resistant (C57BL/10) to murine malaria (Plasmodium chabaudi), were UV-preirradiated and infected with a sub-lethal dose of malaria parasite (104 and 105, respectively). Parasite growth was assayed with tail-blood smears counting parasitized red blood cells. Mice resistant to malaria were bled by heart puncture and the plasma cytokines were determined. Our results showed that UV-B irradiation worsened the malarial infection and 100% of the malaria-resistant mice strains died due to a usual infection at sub-lethal dose following UV-B irradiation. In the resistant mice strain infected with the parasite, the plasma IFN-gamma production was inhibited by UV-B irradiation and the maximum titer was about one-fifth of the non irradiated mice. Furthermore, activation of macrophages from UV-irradiated mice also decreased compared with that of non-irradiated mice. IFN-gamma administration prevented the death of UV-B irradiated resistant mice and the cure ratio was 60%. In conclusion, UV-B increased the susceptibility of both strains of mice and impaired IFN-gamma production in the malaria-resistant mice strain. PMID- 10709357 TI - Ocular ultraviolet B exposure and lens opacities: a review. AB - A review of the epidemiological evidence linking cataract and chronic ultraviolet B radiation (UV-B) exposure was carried out. The majority of ecological studies suggest an increased risk of cataract with residence in areas of greater ambient UV-B. Studies which have measured personal ocular exposure to UV-B have found that even low exposures, as encountered in the general populations of developed countries, confer a measurable risk of cortical cataract. There is sufficient evidence of increased risk of cortical lens opacity with ocular exposure to UV-B to warrant public health messages about simple measures to decrease exposure. PMID- 10709358 TI - Environmental factors in nonmelanoma and melanoma skin cancer. AB - We discuss the role of sunlight, mostly ultraviolet light (UV), in the induction of nonmelanoma and melanoma skin cancer. Whilst the former seems to be correlated with accumulated exposure, the causation of melanoma is more complex, and may also involve the pattern of, and age at, exposure. The efficacy of sunscreens is debatable; while they protect against UVB wavelengths (290-320 nm), and so extend the time that may be spent in the sun before becoming sunburnt, their use may subject wearers to excessive exposure to UVA (320-400 nm) and visible light. Both epidemiological surveys and experiments with animal models suggest that UVA, and perhaps the visible, may induce melanomas. Although Japanese have a much lower incidence of skin cancer than Caucasians, the dramatic rise in skin cancer in Japanese-Americans in Hawaii exposed to high-intensity irradiation raises concerns. If the Japanese people adopt sun-seeking behavior, or should the levels of UV irradiation rise significantly through depletion of the ozone layer, then this could become an important health problem in future. PMID- 10709359 TI - Ambient solar UVR, personal exposure and protection. AB - Ambient solar ultraviolet radiation (UVR) has been monitored around Australia by the Australian Radiation Laboratory (ARL) and its successor ARPANSA since the mid 1980's using a network of radiometric detectors and a spectroradiometer (SRM) for spectral measurements, based in Melbourne. In a continent the size of Australia, the levels vary markedly, basically following a latitude gradient increasing towards the equator but with local geographical and weather effects also evident. ARL also conducts personal exposure studies of various population groups in collaboration with other research centres to gather information on what fraction of the ambient UVR people receive. ARL also undertakes studies on the UVR protection provided by sunscreens, clothing, hats, sunglasses and other materials in an attempt to improve UVR protection used by the public. PMID- 10709361 TI - Time trends of incidence for cutaneous melanoma among the Japanese population: an analysis of Osaka Cancer Registry data, 1964-95. AB - Cutaneous malignant melanoma occurs much less frequently among non-white populations than among white populations. Little is known of the descriptive epidemiology of melanoma among Japanese. We investigated time trends of incidence of invasive cutaneous malignant melanoma using data from the Osaka Cancer Registry (Japan) among 321 men and 313 women diagnosed between 1964-95. Average, annual, age-standardized incidence rates per 1,000,000 population were 2.45 (95% confidence interval (CI): 2.17-2.72) for men and 2.04 (95% CI: 1.81-2.28) for women. The age-standardized rate ratio among men from 1964-71 as a reference was almost constant during the study period, whereas that among women increased up to 1.8 fold (95% CI: 1.25-2.56) in 1980-87 and seems to have reached a plateau recently. Among men, the ratio for head and neck lesions decreased to 0.5 fold (95% CI: 0.26-0.99) in 1988-95. Among women, the ratio for lesions of the extremities steeply increased up to 4.7 fold (95% CI: 2.68-8.35) in 1980-87 from the reference period of 1964-71, whereas a slight increase for trunk lesions and no increase for head and neck lesions were noted during the same period. Possible explanations for the subsite-specific time trends are discussed. PMID- 10709360 TI - Regional correlation between estimated UVB levels and skin cancer mortality in Japan. AB - Most ecological studies investigating the relationship between incidence and/or mortality of skin cancer and surrogate measures of ultraviolet radiation B (UVB) have been conducted among the Caucasian population. The objective of the present study was therefore to assess the geographical correlation between ambient UVB estimates and regional mortality rates for skin cancer in Japan. The standardized mortality ratio (SMR) for malignant melanoma and other malignant neoplasms of the skin was calculated by sex, region and time-period for all deaths occurring in the period 1973-1994. The Spearman's correlation coefficient was calculated between estimated ambient UVB and regional SMRs for the two types of skin cancer. There was no geographical correlation between UVB and skin cancer mortality, except for a significantly negative correlation in malignant melanoma among males and a significantly negative correlation in other malignant neoplasms of the skin confined to unexposed anatomic sites of the body among females. The characteristic ecological relationship adds to the importance of conducting further epidemiological studies at the individual level in Japan. PMID- 10709362 TI - Influence of chronic UV exposure and lifestyle on facial skin photo-aging- results from a pilot study. AB - In order to better understand the effect of chronic sun exposure on facial skin photo-aging and to identify the factors affecting it, we planned a study in two areas in Japan, Akita and Kagoshima, which correspond to the low and high sun exposure environments, respectively. As a first step, we conducted a pilot study in the two areas, examining 195 subjects. Hyper-pigmentation and wrinkling were measured with a high-resolution digital video imaging system. As expected, people in Kagoshima had darker skin, higher visual grades of facial hyper-pigmentation, and more facial wrinkles than people in Akita, reflecting the difference of UV exposure levels in the two areas. Both the grades of hyper-pigmentation and number of wrinkles increased in a roughly linear fashion with the advancement of age. On the other hand, the effect of gender was different in those two skin photo-aging parameters. Women had higher hyper-pigmentation grades (P = 0.012) and less wrinkles (P = 0.004) than men. Interestingly, post-menopausal women had higher grades of hyper-pigmentation than pre-menopausal women. Neither sun exposure index for darkness nor wrinkling showed any significant differences by menopausal status. In this pilot study, we collected information on various factors, including life-styles. The results of detail analysis will be presented elsewhere. In the present analysis, we found that the grade of hyper-pigmentation was not related to total hours spent outside in life but was affected by various factors, including toe-nail zinc levels. On the other hand, the number of wrinkles was significantly related to total hours spent outside in life. The most important risk factors of non-melanoma skin cancer are chronic sun exposure, age and male sex. All of them are strongly related to higher levels of UV exposure. The present study confirmed that chronic sun exposure, age and male sex were strong risk factors of the wrinkle number. The number of wrinkles was significantly related to total hours of sun exposure in life, increased in a roughly linear fashion with the advancement of age, and was larger in men than in women. In epidemiological studies of UV-related skin cancer, the number of wrinkles, which can be easily measured with a high-resolution digital video imaging system as shown in this study, may be a good marker of total sun exposure in life. PMID- 10709363 TI - Keio University International Symposia for Life Sciences and Medicine: Strategy for Cardio-aortic and Aortic Surgery. Tokyo, Japan, December 1-3, 1999. Abstracts. PMID- 10709364 TI - Call for diazepam replacement. PMID- 10709365 TI - Concern over battlefield use of intravenous fluids. PMID- 10709366 TI - International military human immunodeficiency virus/acquired immunodeficiency syndrome policies and programs: strengths and limitations in current practice. AB - A survey was conducted to evaluate military human immunodeficiency virus/acquired immunodeficiency syndrome (HIV/AIDS) policies and programs in 119 countries. Ninety-eight percent of the 62 respondents provide prevention education, 95% in group settings but only 53% individually. Predeployment briefings are more common than postdeployment briefings. Condoms are promoted more often than provided. Seventy-eight respondents report some form of mandatory HIV testing, and 58% perform mandatory recruit testing, with recruitment denied to HIV-positive individuals in 17%. Counseling accompanies mandatory testing less than voluntary testing. In-service care for AIDS patients is universal. Many military prevention programs can be improved through postdeployment briefings and proactive interventions involving education, condom distribution, and counseling combined with testing. Mandatory testing is often inconsistent with stated goals, and AIDS care policies may strain military budgets. Testing based on cost-benefit assessments may increase efficiency in military HIV control. Military budgets may benefit from greater civil-military cost sharing in AIDS care. PMID- 10709368 TI - A suicide prevention advisory group at an academic medical center. AB - During a 15-month period, there were seven suicides among patients who were in active treatment or who had been seen recently by providers in the Department of Psychiatry of Tripler Army Medical Center, Honolulu, Hawaii. As a result, a Suicide Prevention Advisory Group was formed to identify possible causes and make recommendations aimed at improving the identification and treatment of suicidal patients. The group made 11 specific recommendations. No known suicides occurred during the 22 months after the implementation of the Suicide Prevention Advisory Group's recommendations. PMID- 10709367 TI - Surgical treatment of war lesions to the posterior segment of the eye. AB - OBJECTIVE: The aim of this study was to assess the outcome of surgical treatment of war lesions to the posterior eye segment. METHODS: During a 1-year period, 96 patients with severe eye lesions, 54 of them with intraocular foreign bodies (56.2%), were operated on at the Institute for Posterior Eye Segment Diseases and Trauma, Zagreb Clinical Hospital. Surgical procedures performed included high quality primary microsurgery of the wound, extraction of intraocular foreign bodies, treatment of double perforating wounds by silicone oil tamponade and gas, and emergency vitrectomy when necessary. RESULTS: A varying degree of vision function improvement was achieved in 67 patients (69.7%). Most of the foreign bodies removed were metal and nonmagnetic. Only six foreign bodies were made of glass, and two 2 were of plastic material. In 29 patients (30.3%), the vision function remained unchanged or deteriorated after vitrectomy. CONCLUSION: Although satisfactory results were achieved, definitive evaluation of the procedures will be possible only after long-term follow-up. PMID- 10709369 TI - Circadian disturbances after night-shift work onboard a naval ship. AB - The aim of the present study was to investigate how night duties can affect the circadian rhythms of military personnel working onboard a naval ship. Twenty individuals on a regular day-work schedule from 9:00 a.m. to 6:00 p.m. (serving as controls) and 40 individuals on night-shift duties participated in the study. Salivary melatonin and cortisol profiles were established within two 24-hour periods from 2-hour saliva samplings. Under the condition of abrupt shift in work/rest schedule, the majority of the navy officers (52%) retained their normal melatonin profiles. Twelve percent displayed a right phase shift in melatonin rhythm after night work. Nineteen percent exhibited distortions in the form of abnormal peaks or troughs, and 17% showed signs of disrupted rhythm in the form of low daytime levels of melatonin throughout the sampling period. No consistent relationship was found between the melatonin changes and various work stations of the ship. Prominent changes in the cortisol profile included unexpected peaks or troughs that may be related to the conditions that individuals were exposed to, i.e., high noise level in the engine room, as well as to performing intense tracking operations. The findings of this study (1) show the possible detrimental effects of shift duties on circadian rhythms, (2) highlight a wide interindividual variation in the manner in which the circadian systems respond to an abrupt phase shift in work/rest schedules, and (3) form the basis for further investigations into effective strategies to help military personnel cope with shift work, thereby maintaining health and high working standards while on duty. PMID- 10709370 TI - Patient compliance and blood pressure control on a nuclear-powered aircraft carrier: impact of a pharmacy officer. AB - The impact of a pharmacy officer on patient compliance and blood pressure control on a deployed nuclear-powered aircraft carrier for a 2-week at-sea period was evaluated. Before any counseling by a pharmacy officer, 43 crewmembers on chronic medications anonymously completed a compliance questionnaire. The pharmacy officer then counseled these crewmembers. A follow-up compliance questionnaire was completed 2 weeks later. After counseling, compliance had increased 58% (p < 0.0001) from compliance measured before counseling. The pharmacy officer also initiated therapeutic interventions. Among 26 crewmembers diagnosed as hypertensive, preintervention blood pressure (BP) measurements were obtained. Ten to 14 days after the initial BP measurement, BP was remeasured. After intervention, 31% (p < 0.02) more crewmembers were at BP goal compared with before intervention. A pharmacy officer, working closely with a medical officer, improved patient compliance and blood pressure control. One problem identified was that these warships require computer software that can prospectively identify drug-drug interactions. PMID- 10709371 TI - Problematic intubation in soldiers: are there predisposing factors? AB - Attempts by a combat medical officer to secure the airway of a multiple-injury patient in the field may frequently end in failure. The recurrence of such failures, despite Advanced Trauma Life Support training, is perplexing; therefore, we studied the prevalence of clinical criteria that could predispose active soldiers to difficult intubation. Such known anatomical features and the Mallampati classification were assessed by experts in 250 soldiers at a military outpatient clinic of the Israel Defense Forces. It was found that most soldiers had normal airways. Limitations of head and neck movement, or in opening the mouth, were not observed. Other risk factors were noted in only a small percentage of the study population. Mallampati classes I and II were noted in 40% and 31%, respectively. No statistically significant differences between young (18 21 years) and older (40-44 years) soldiers were found. It was concluded that difficult intubations among soldiers are unlikely to be associated with anatomical causes. Complicated scenarios and deficient skills of the physicians are the most important factors that contribute to in-field failures to secure airway control. Several recommendations to remedy this situation are offered. PMID- 10709372 TI - Modeling access, cost, and perceived quality: computer simulation benefits orthodontic clinic staffing decisions. AB - Given limited financial resources, simulation permits a financial analysis of the optimum staffing levels for orthodontists and dental assistants in an orthodontic clinic. A computer simulation provides the information for managerial review. This study, by building a computer simulation of an orthodontic service, set out to determine the most efficient mix between providers and support staff to maximize access, maximize perceived quality, and minimize expenditures. Six combinations of providers and support staff were compared during an animated, computer-generated what-if analysis. Based on the clinic workload and size, on the cost per patient, and on the cost per quality point, the research team recommended a staffing mix of one orthodontist and three assistants. This study shows that computer simulation is an enormous asset as a decision support tool for management. PMID- 10709373 TI - Physiological changes in pigs exposed to a blast wave from a detonating high explosive charge. AB - The aim of this project was to study respiration, circulation, and brain activity in pigs during and after a blast wave exposure. Ten anesthetized pigs were used. Seven were exposed to blast and three were controls. Physiological parameters of respiration and circulation as well as cortical activity were followed from 30 minutes before until 120 minutes after the real or simulated blast. There were no significant changes in heart rhythm, cardiac output, arterial oxygen or carbon dioxide tension, blood pH, or mixed venous saturation during the experiment. The blast exposure caused intestinal injuries but no lung damage. A transient flattening of the electroencephalogram was seen immediately after the blast in four experimental animals, in contrast to the unchanged baseline electroencephalogram of the control animals. This momentary depression of cortical activity accompanied by short-lasting apnea indicates a blast wave induced effect on the brainstem or higher controlling center. PMID- 10709374 TI - Endodontics and dental readiness. AB - The primary mission of the Dental Corps--maintaining the dental health of our soldiers at an optimum level to ensure their readiness to deploy and fight--is more important than ever in today's downsized, high-operational-tempo Army. A review of the literature indicates that 20 to 25% of all soldiers will report on dental sick call during a 1-year deployment, and approximately half will require endodontic intervention. This could cost a division more than 18,000 man-days of combat effectiveness in a theater of battle, an unacceptable loss. Approximately three-fourths of these sick call visits could be prevented if selected conditions were identified and treated before deployment. Endodontic conditions, which result in the majority of dental sick call complaints, should be given priority for treatment in garrison, and the identified soldier should be placed in dental fitness class 3 until the tooth is definitively treated. PMID- 10709375 TI - An oral history of the "joint" nursing experience at Landstuhl Regional Medical Center. AB - In 1993, the U.S. Army and U.S. Air Force undertook long-term joint staffing of Landstuhl Regional Medical Center, a 180-bed facility in Germany. With little historical precedence, the Nurse Corps from each service worked diligently to integrate. Initial collaboration suffered because of confusion, ignorance of the other service's capabilities, and differences in mission emphasis and personnel management. Eventually, the nursing staffs built a cohesive team. Lessons learned for future multi-service nursing integration include extensive planning before implementation, flexibility and a willingness to learn, and full integration of service members throughout the organization. Staffing by two or more military medical services can be done successfully. PMID- 10709376 TI - Tattooed Army soldiers: examining the incidence, behavior, and risk. AB - Primary prevention is a priority for medical personnel. Despite societal popularity and a long association of the military with tattooing, little is known about the tattooed Army soldier, which hampers primary health planning. Basic recruits and advanced individual training students (N = 1,835) at one mid-western military installation completed a questionnaire about any tattooing experiences. Almost half (48%) of the soldiers were serious/very serious about getting a tattoo, with 31% stating that there were "no reasons" to keep them from getting a tattoo. More than one-third (36%) were tattooed, with 22% possessing three or more tattoos. Many soldiers (64%) entered the military with the tattoos. Limited use (15%) of alcohol and/or drugs before tattooing was reported. Findings included a high incidence of tattooing, a strong determination to obtain tattoos, the possession of tattoos for self-identity reasons, and the supportive role of friends. Reported procedural bleeding (76%) documents the potential for blood borne disease transmission. These results confirm the need for targeted health education regarding the safety and potential risks of tattooing. PMID- 10709377 TI - Stress fractures of the pelvis in female navy recruits: an analysis of possible mechanisms of injury. AB - The purpose of our study was to investigate possible risk factors and mechanisms for the development of pelvic stress fractures in female Navy recruits. We used a case-control retrospective study of female Navy recruits undergoing basic military training. We compared anthropometric and activity data between recruits with pelvic stress fractures (N = 25) and female recruits who completed training without injury (N = 61). Recruits developing pelvic stress fractures were significantly (p < 0.05) shorter and lighter and were more frequently Asian or Hispanic than recruits without stress fractures. In addition, recruits with pelvic stress fractures reported marching in the back of their training division, were road guards, and felt that their stride was too long during training activities more often than recruits without injury. Self-reported fitness, activities before recruit training, or a history of amenorrhea was not found to be associated with the development of a pelvic stress fracture in our population. PMID- 10709378 TI - Cadet basic training: an ethnographic study of stress and coping. AB - Cadet basic training (CBT) at the U.S. Military Academy at West Point is an initial cadet experience designed to transition freshmen (new cadets) into the military. Challenge is an inherent component of CBT, and some challenging activities may be stressful. However, the nature and the impact of stress on health status have not been systematically investigated. An ethnographic technique, participant observation, was used to identify stressors and coping strategies among cadets aged 18 to 21 years participating in CBT. A company of 183 cadets, consisting of 123 new cadets and 60 supervising upperclass cadets from the U.S. Military Academy, was followed throughout the 6-week CBT in the summer of 1993. The investigator observed daily activities and participated in select field training experiences. Daily field observations were taped, and field notes were generated chronicling the experience. After CBT, 10 of the 60 upperclass cadets participated in a 20-minute structured interview. Field and interview notes were systematically reviewed to identify and categorize stressors and coping techniques. Stressors included anticipatory stress, time management pressures, sleep deprivation, performance evaluations, conflicts between teamwork and competitive grading, and inexperience in the leadership role. Coping techniques identified included perceiving social support, humor, and rationalization. Three new hypotheses were generated from the observations. PMID- 10709379 TI - Intra-abdominal sound pressure levels during impulse noise exposure in sheep. AB - Ambient sound pressure levels (SPLs) created with intense blasts were compared with SPLs recorded in the abdomen of euthanized sheep. Hydrophones were placed in the abdominal cavity at locations referred to as proximal, medial, and distal with respect to a shock tube that created 169-dB peak SPL (pSPL). No differences in pSPL, duration, or rise time were found between recordings in air and at the intra-abdominal proximal position. Significant differences were noted in these variables when recordings in air were compared with recordings made at the medial and distal locations. Intra-abdominal pSPL varied by 20 dB depending on recording location. PMID- 10709380 TI - Left coronary artery anomaly: an often unsuspected cause of sudden death in the military athlete. AB - More than 300,000 cases of sudden cardiac death (SCD) occur in the United States each year. Left coronary artery anomaly (LCAA), although rare, is second only to hypertrophic cardiomyopathy as the most common cause of SCD associated with structural cardiovascular abnormalities. This case illustrates SCD secondary to LCAA in a military athlete. A 19-year-old soldier collapsed after an 8-km run. On arrival at the emergency room, he was unresponsive and in asystole. Despite successful resuscitation and aggressive management, the patient died the next morning. Autopsy revealed an anomalous left coronary artery. LCAA-associated SCD is rare and usually seen in young individuals who collapse (and/or die) while exercising. A substantial proportion of these individuals experience prodromal symptoms of exertional chest pain, syncope, and/or sudden collapse. Early recognition and intervention are key to survival. Rapid, early imaging and invasive therapeutic measures leading to surgical correction may be the difference between life and death. PMID- 10709381 TI - Self-limited recurrent multifocal neurological symptoms, headache, and cerebrospinal fluid lymphocytic pleocytosis: a benign syndrome with a predilection for young adult men. AB - Two young men, aged 34 and 30 years, developed transient recurrent multifocal neurological symptoms with associated severe headache over a 2-week period. Both had a lymphocytic pleocytosis in their cerebrospinal fluid. Cranial imaging studies were normal. All symptoms resolved without recurrence. Although the cause and pathogenesis are undefined, this self-limited benign neurological syndrome may be more common than previously recognized and has a predilection for young adult men. PMID- 10709382 TI - Myomectomy during early pregnancy. AB - Abdominal pain during early pregnancy may be caused by leiomyoma of the uterus. Inconsistency of uterine size and gestational dates in a pregnant patient with acute abdominal pain may be the first sign of leiomyoma. This 31-year-old primigravida presented with progressively worsening lower abdominal pain at 12 weeks gestational age. Ultrasonography and magnetic resonance imaging demonstrated a large fundal heterogeneous mass and an intrauterine gestation compatible with her menstrual dates. Exploratory surgery and myomectomy confirmed a large leiomyoma showing benign degenerative changes. The operative procedure was successful, and the pregnancy progressed normally. An elective cesarean section was performed at 37 weeks gestation after confirming fetal maturity by amniocentesis and serial ultrasonography. Abdominal pain in a pregnant patient with leiomyoma uteri may be attributable to degenerative changes in the myoma. Surgical intervention during pregnancy is occasionally necessary in uncommon cases of intractable pain. PMID- 10709383 TI - Do we need new supraglottic devices? Clinical appraisal of the cuffed oropharyngeal airway (COPA) AB - BACKGROUND: The laryngeal mask airway (LMA) has been widely studied for both conventional and nonconventional uses, while the literature on the cuffed oropharyngeal airway (COPA) is still limited. The purpose of this manuscript was to review the initial appraisal of efficacy, safety, effects on hemodynamics and respiratory function, induction agents and drug requirements of this new supraglottic device. METHODS: We reviewed main results of studies recently published on peer reviewed journals concerning the clinical uses of COPA. RESULTS: When used in healthy adults undergoing general anesthesia for routine minor procedures, the COPA and LMA are substantially equivalent. The LMA is associated with a higher first-time placement rate and fewer manipulations during usage, but the incidence of airway untoward events during COPA anesthesia is equivalent to that reported when using an LMA. The quality of breathing and capnography during COPA ventilation is similar to that provided by the LMA ventilation, with clinically relevant decrease in the physiological deadspace/tidal volume ratio and arterial to end-tidal CO2 tension difference compared with facemask ventilation. In selected patients without risk factors for regurgitation of gastric content, positive-pressure ventilation is similarly successful and safe with the COPA as with the LMA. The COPA seems to be less stimulating than LMA because it has been demonstrated to cause a lower incidence of pharyngeal trauma and sore throat in the immediate postoperative period, requires shorter exposure to an inhalational anesthetic and lower concentrations of propofol to be successfully placed, and is associated with lower effects on the patient's hemodynamic homeostasis than LMA. CONCLUSIONS: More extensive clinical evaluations should be advocated to better understand the risk/benefit ratio of this new supraglottic device; however, it may be concluded that in healthy adults receiving general anesthesia for short procedures the COPA allows for an effective and safe control of the patient's airway and ventilation. PMID- 10709384 TI - Anesthesia in geriatric patients. PMID- 10709385 TI - [Laryngeal mask as alternative to facial mask in the newborn]. AB - OBJECTIVE: The present study demonstrates that the use of laryngeal mask airway (LMA) is an alternative to face-mask (FM) during induction of general anesthesia with halothane. In all patient the induction of general anesthesia is carried out by halotane and N2O/O2 50%, using only the LMA, preceding topical anesthesia of pharynx. METHODS: EXPERIMENTAL DESIGN: prospective study. SETTING: this study was carried out at the surgical-division of the Pediatric Clinic, of University "La Sapienza", Rome. PATIENTS: a total of 80 newborns, average age 14.8 +/- 2.4 days and average body weight 2280 +/- 110 g were examined. INTERVENTIONS: newborns were submitted to surgery for congenital malformations, diagnostic research and positioning of a central venous catheter (CVC). MEASUREMENTS: Heart rate, non-invasive arterial pressure through cardiomonitor Hewlett Packard 78352A, oxygen saturation through Nellcor N3000, time of induction of general anesthesia and respiratory rate were assessed. RESULTS: Blood pressure and heart rate were increased during the positioning of LMA; oxygen saturation remained > or = 94% and respiratory rate was constant during the whole observation. Muscular relaxing, as an index of anesthesia, was observed after 33 +/- 1.5 sec after positioning of LMA. CONCLUSIONS: In the light of the results obtained, the use of the LMA for airway ventilation during the induction period of pediatric anesthesia is suggested. PMID- 10709386 TI - [The use of bronchial fibroscopy for difficult intubations in maxillofacial surgery]. AB - BACKGROUND: Evaluation of difficulties and modalities of tracheal intubation in maxillofacial surgery. METHODS: DESIGN: retrospective study. SETTING: maxillofacial operation room. 2152 patients who underwent elective maxillofacial surgery during a five-year time, from 1994 to 1998. Indications and alternative modalities of tracheal intubation through fiberoptic bronchoscope in eight patients (0.37%) with preoperatively evaluated difficult intubation due to temporomandibular ankylosis (3 patients), burns sequelae (1 patient), craniofacial congenital malformations (2 patients), unstables fractures of the cervical spine (2 patients), are discussed. Fiberoptic bronchoscope was used through nasotracheal route under topical nasal and laryngeal anaesthesia, combined with appropriate benzoanalgesia, in order to maintain spontaneous breathing. Proper positioning of tracheal tube was directly checked by fiberoptic bronchoscope, through visualization of the tracheal carina. RESULTS: In seven patients tracheal intubation was easily performed without complications in less than 20 minutes. In the eight patients the time spent was 35 minutes, due to important nasal bleeding, which caused a change of the nostril. CONCLUSION: Among the alternative methods of elective tracheal intubation, previously evaluated as difficult or impossible using conventional laryngoscopy, the use of fiberoptic bronchoscope seems to be safest and easiest to use, although not the cheapest. PMID- 10709387 TI - [Target controlled infusion (TCI): applications of the "TCI visual" program in anesthesia and sedation with propofol]. AB - BACKGROUND: Systems for Target Controlled Infusion accepting not only patient' data, like Diprifusor, but also a pharmacokinetic model have not been available in Italy in the last years. Therefore a program which controls a Pilot Anesthesia Vial pump and accepts any pharmacokinetic model was developed and applied to propofol infusion for anaesthesia and sedation. METHODS: Two versions of the Visual TCI program have been developed. The first, at intervals, supplies the anaesthetist with the values for the pump; the second directly interacts with the pump. The program also supplies the anaesthetist with the current amount of drug in each compartment and with the estimated awakening time. DESIGN: preliminary prospective study. SETTING: operatory theatre and Intensive Care Unit in a University Hospital. PATIENTS: 6 patients undergoing total intravenous anaesthesia with propofol and fentanyl for abdominal surgery; 6 patients undergoing sedation with propofol in an Intensive Care Unit (the first 4-hour period was taken into account). INTERVENTIONS: propofol infusion was regulated by the Visual TCI program. The first version was employed in three patients of each group and the second one in the others. Hypo- and hypertensive episodes (systolic pressure less than 80 mmHg or higher than basal value plus 25%) were recorded during anaesthesia and sedation. Propofol concentration was measured in plasma three times at defined intervals and per cent differences between measured and computer-calculated values (Predictive error, PE) were calculated. RESULTS: No hypo- or hypertensive episodes were recorded. PE was 27.4 +/- 17.9%. CONCLUSIONS: The program was easily employed, caused no inconvenience, and its use was associated with a remarkable cardiovascular stability. PE distribution was acceptable on the ground of the criteria reported in the literature. The program can be applied to drugs other than propofol, with both two and three compartment pharmacokinetic models and the anaesthetist can choose the most suitable model for the patient. PMID- 10709388 TI - Postoperative analgesia for early extubation after cardiac surgery. A prospective, randomized trial. AB - BACKGROUND: Early extubation after cardiac surgery is a procedure recently gaining interest due to its ability to shorten intensive care unit and hospital stay and to limit the operation-related costs. Its use, however, raised new problems in terms of pain control in the early postoperative course, due to the need for limiting opioid analgesia. This study deals with non-opioid pain control after cardiac surgery and early extubation. METHODS: Prospective, randomized trial aimed to investigate the effectiveness of three intravenous analgesic drugs (ketorolac, 60 mg i.v.; propacetamol, 2 g i.v.; tramadol, 200 mg i.v.) for the management of postoperative pain in early extubated cardiac surgical patients. Each treatment group comprised 20 patients. RESULTS: The pain assessment (5-item verbal scale) demonstrated a significant (p < 0.05) lower value in patients treated with ketorolac vs propacetamol, while patients treated with tramadol did not significantly differ from the other two groups. There was a significantly (p < 0.05) higher rate of patients with severe pain in propacetamol group. Patients treated with tramadol had a significantly (p < 0.01) higher PaCO2 (48 +/- 6 mmHg) versus patients treated with ketorolac (43.4 +/- 3.7 mmHg) or propacetamol (42.9 +/- 3.4 mmHg). CONCLUSIONS: Tramadol and ketorolac seem to be the best options for treating postoperative pain in the specific setting of early extubation after cardiac surgery; high doses of tramadol may result in a significant even if clinically not relevant respiratory depression. PMID- 10709389 TI - Prevention of acute renal failure in major vascular surgery. AB - The aim of this study was to analyse the main problems which involve renal function in major vascular surgery and can lead to postoperative acute renal failure. The factors responsible for renal damage in this surgical branch are at first analysed, then followed by the physiological changes which characterize the renal injury, the techniques employed to detect and monitor them and finally the therapeutic tools available to prevent acute renal failure. The most significant data of personal experience on the use of nifedipine and low-dose dopamine during abdominal aortic surgery are then presented. It is concluded that: a) an ischemic attack is the main cause of acute renal failure in mayor vascular surgery; b) prevention of ischemic renal damage is far superior to treatment; c) the optimal management of the cardiovascular function by means of the invasive hemodynamic monitoring, is the main tool to protect the kidneys and prevent acute renal failure; d) the pharmacological protection by diuretics and low-dose dopamine is of minor importance and anyway subordinate to the maintenance of adequate hemodynamics, as well as for calcium antagonist whose employment however seems to protect the kidneys better against an ischemic attack. PMID- 10709390 TI - [Rhabdomyolysis caused by improper intraoperative positioning. Clinical and medico-legal aspects]. AB - The etiology, diagnosis, pathology and treatment of rhabdomyolysis due to intraoperative malpositioning and the medico-legal implications of physicians involved in the surgical treatment and anesthesia of the patient are described. According to the Italian law, the anesthesiologist is the only physician of the surgery-anesthesia team responsible for the patient's positioning. The anesthesiologist must assume primary responsibility for protecting the patient from iatrogenic injuries due to improper positioning, and/or inadequate preventive measures. PMID- 10709391 TI - [The Revista Espanola de Salud Publica in the SciELO Virtual Library (Scientific Electronic Library On-Line)]. PMID- 10709392 TI - [The performance and perception of informed consent in a health-care section of Catalonia]. AB - BACKGROUND: Informed consent entails a process which involves more than signing a form to give one's consent. This process involves the mutual exchange of information, understanding, trust and consent between physician and patient. The purpose of this study is that of ascertaining the degree to which those consumers who have gone through the informed consent process in a health care district in Catalonia have filled out and fully understood this form. METHODS: Telephone survey conducted among 314 former surgery patients at hospitals in a given district, ages 18-75, who had undergone surgery within the three months immediately prior to the date on which the survey was conducted for the purpose of ascertaining the opinion of these consumers regarding the consent process. A review of the clinical records of 30% of these individuals was conducted for the purpose of ascertaining the degree of compliance with the consent form process. RESULTS: Sixty-one percent (61%) of the patients surveyed remembered having signed the consent form, 59.2% recalling explanations regarding risks or complications of the surgery they were to undergo. Nine percent (9%) did not understand well enough what was going to be done to them during the surgery, and 36% were of the understanding that the consent form released the health care professionals from liability. A statistically significant relationship was found to exist among the age group, educational level and admissions channel. Seventy eight percent (78%) of the clinical records reviewed included informed consent forms. The diagnosis was stated on 14.9% of these forms, and 48.9% had been signed by the attending physician. A statistically significant relationship was found between the channels through which patients were admitted to the hospital. CONCLUSIONS: The informed consent process is not being implemented correctly at the two hospitals analyzed. Physicians are not totally involved in the process. PMID- 10709393 TI - [A health study of older persons in Extremadura: drug taking and the most frequent chronic diseases]. AB - BACKGROUND: To ascertain the use of drugs among the non-institutionalised elderly population, the factors related to polypharmacy and pinpointing the relationship thereof with chronic diseases. METHODS: Cross-sectional epidemiological study by means of door-to-door survey. The population systematically selected totalled 960 individuals age 65-93. The questionnaire included demographic data, self-assessed health condition, quantitative and qualitative aspects of the use of medications, chronic diseases and assessment of functional ability. RESULTS: 91.62% of those surveyed were taking medication, a greater number of females (p < 0.002) than males. The medications taken to the greatest extent were blood pressure drugs (42.4%), analgesics (38.7%) and heart drugs (24.8%). Drugs were used to a greater extent among the oldest age group (p < 0.0001), those having the lowest educational level (p < 0.001), those with impaired eyesight and hearing (p < 0001), those having the lowest social status (p < 001), more contacts with the health care services (p < 0001), worse self-assessed health condition (p < 0001), a greater number of chronic diseases (p < 0.0001) and depressive disorders (p = 0.004). The linear regression analysis revealed a positive relationship between the number of medications taken and the number of chronic diseases (r = 0.518; p < 0.0001). Females report worse health conditions (p < 0.05). The variables analyzed with regard to polypharmacy by means of logistic regression are age (over age 75; OR = 1.1478), three or more chronic diseases (OR = 1.83) and poor self-assessed health condition (OR = 1.22). CONCLUSIONS: Physical checkups on the elderly must include a review of the medications being taken, especially among those over age 75 who have a worse self-assessed health condition and a larger number of chronic diseases. PMID- 10709395 TI - [What is your diagnosis? Pott disease--tuberculous spondylodiscitis]. PMID- 10709394 TI - [AIDS prevention in secondary school: program summary and evaluation]. AB - BACKGROUND: In the past decade a number of school AIDS prevention programs addressed to adolescents have been published in Spain. This paper aims to compile, describe and assess these programs. METHODS: AIDS prevention programs addressed to secondary school students which were published in Spain from 1990 to 1997 were included in the study. The collection of the programs was made by contacting aids prevention heads of national and regional governments. Each program was described in a standardized form and assessed according to the following quality criteria: preventive skills training, teachers or peer delivered, active pedagogical methodology, and at least administered along four sessions. RESULTS: 19 programs were included. Seventy-four percent (n = 14) are specific AIDS prevention programs and 26% (n = 5) sexual health programs. Among the AIDS programs, 57% include skills training, 93% are delivered by teachers, 100% propose active learning methodology and 64% are recommended to be developed in four or even more sessions. Only one of the sexual health programs includes skills training, all of them are delivered by teachers and use active learning methodology and 60% (n = 3) are recommended to be applied in four or even more sessions. CONCLUSIONS: Nearly all regional governments have developed or adapted secondary school preventive materials. Most of them are AIDS-specific programs. They use active learning methodology and suitable providers, but an important percentage of the programs lack a higher number of sessions and skills training. Programs should include peer-delivered activities. PMID- 10709396 TI - [Potential drug interactions and number of prescription drugs with special instructions at hospital discharge]. AB - OBJECTIVE: Up to 6% of all hospitalizations are due to adverse drug reactions and 20% of these are caused by drug-drug interactions. There is only little information on the prescription frequency of drug-combinations with the potential to induce dangerous drug-drug interactions and drugs with the need for special patient instruction (e.g. inhalers). The aim of our study was to investigate the frequency of such drug prescriptions at hospital discharge. PATIENTS AND METHODS: In a retrospective, descriptive study drug prescriptions of 100 patients discharged consecutively from the department of internal medicine of a 300 bed hospital were analysed. Possible drug-drug interactions were detected using a special computer program. Furthermore, the number of prescriptions warranting patient instruction such as anticoagulants, antidiabetics, hormones, immunosuppressive drugs, chemotherapeutics, antituberculotic and antiepileptic drugs as well as inhalatives and injections was recorded. RESULTS: The mean age of the 100 patients (61 men, 39 women) was 61.7 years, the mean duration of the hospital stay was 9.2 days. At discharge, patients took an average of 3.5 different drugs. Half of the patients were given drug-combinations with the potential for drug-drug interactions, whereby 5% were at risk for the development of interactions of severe and 42% of intermediate degree. All drug-combinations with potentially severe interactions were prescribed deliberately. 31% of all patients took medications with the need for special education, with inhalatives being the most frequent. The prescription of drugs with potential interactions and the necessity for special patient instruction was more frequent in the elderly. CONCLUSIONS: Drug-combinations with the potential of harmful interactions and drugs with the requirement for special patient instruction are frequently prescribed at hospital discharge. The frequency of prescribing these drugs increases with age. Detection of potentially dangerous drug-drug interactions is simplified by special computer programs. Careful patient instruction about the use of certain drugs is a key issue to improve patient compliance and to guarantee an optimal treatment effect. PMID- 10709397 TI - [Urticaria]. AB - Urticaria may affect as many as 25% of all people at some time during their lives. The hallmark of urticaria are transitory and completely reversible wheals (intracutaneous oedema). There are multiple causes of urticaria. It has various clinical expressions. Corresponding to the time course it is possible to differentiate between acute, acute relapsing and chronic urticaria. Acute and acute relapsing urticaria is often caused by an allergy or "pseudoallergy". Causal factors of chronic urticaria are usually elusive and therefore only a symptomatic treatment is possible. H1 antihistamines with a low potential for sedation are the most important first-line treatment. PMID- 10709398 TI - [Idiopathic entrapment neuropathy of the ilio-inguinalis nerve--a differential diagnosis in inguinal pain]. AB - We report the case of a 41-year old man, who developed leftsided and later rightsided inguinal pain radiating to the scrotum within three months. There was also sensory disturbance in the supply area of the left ilioinguinal nerve. No surgical procedures in the inguinal region had been done, so that we supposed an idiopathic entrapment neuropathy of the ilioinguinal nerve. Infiltration with local anesthetics resolved the complaints on the left side completely. On the right side neurolysis was necessary, to ablate symptoms. The entrapment neuropathy of the ilioinguinal nerve--often called ilioinguinal-syndrome--is a relevant differential diagnosis of inguinal pain. Anatomy, pathology, clinical presentation and therapy of this syndrome are discussed in the article. Knowledge of this syndrome prevents unnecessary investigation. PMID- 10709399 TI - [75-year-old man with rapid deterioration of his general condition]. AB - A 75 year old man was hospitalized because of rapid deterioration of his general condition, weight loss, night sweat and subfebrile temperature. The patient presented with severely reduced general condition, tachyarrythmia with previously known atrial fibrillation. Laboratory investigations revealed among others a moderately increased CRP-value, normal sedimentation rate and, initially, a normal white cell count. The differential diagnosis included neoplastic and infectious diseases (i.e. endocarditis, tuberculosis), endocrinopathies (i.e. thyreoditis) and general inflammatory or rheumatic diseases. An initially increased CMV-IgM-titer, a rising anti CMV IgG-titer and gradually developing atypical lymphocytosis in particular suggested diagnosis of severe CMV-infection. The diagnosis was confirmed by PCR positive for CMV-DNA in blood. The course of a CMV-primary infection may develop from asymptomatic infection over a mononucleosis-like syndrome to disseminated CMV-infection with multi organ involvement. PMID- 10709401 TI - Pituitary tumours--a new look at an old problem. PMID- 10709400 TI - [Irregular pulse as an incidental finding in a 30-year-old athlete]. PMID- 10709402 TI - Some recent developments and concerns in occupational health. PMID- 10709403 TI - Epidemiology, presentation and management of congenital muscular torticollis. AB - AIM OF STUDY: Congenital muscular torticollis is a condition of debatable aetiology and management. Untreated, cervical function and facial cosmesis may be severely compromised. The aim of this study was to establish the epidemiology, presentation and management of congenital muscular torticollis in Singapore. PATIENTS: Ninety-one patients with torticollis were seen at the National University Hospital (NUH) from January 1994 to December 1997. Torticollis was first noted at a median age of 2 months with the median age of presentation being 6 months. At presentation, a sternomastoid tumour was noted in 33 patients and 62 patients had facial asymmetry. Thirteen of 22 patients with neonatal records available had mandibular hypoplasia at birth on the side where the sternomastoid was affected. Half of the patients (45) had a right sided lesion, with 46 being left sided. The rates of assisted breech delivery, instrumental deliveries (forceps and vacuum) and Caesarean section were higher in the study group. Nine (59.1%) of 13 patients with vertex presentation, had a lesion on the side of the presenting shoulder. Forty-eight of 72 patients responded well to therapy with improvement; 20 underwent surgery and the median age of presentation of 19.5 months in this group was significantly later than that of 4 months in the group which responded to physiotherapy alone. CONCLUSION: Birth trauma appears to be the main aetiological factor in congenital muscular torticollis. Patients generally respond well to physiotherapy. This study revealed 2 findings hitherto unreported: (1) mandibular hypoplasia may be an useful early sign of this condition, and (2) the side affected may depend on the side of shoulder delivered first. More studies, however, are required to confirm these findings. PMID- 10709404 TI - Comparison of the diagnostic utility of CK, CK-MB (activity and mass), troponin T and troponin I in patients with suspected acute myocardial infarction. AB - OBJECTIVE: The aim of the study was to investigate the clinical performance of serum creatine kinase (CK), CKMB (mass and activity), Troponin T (TnT) and Troponin I (TnI) in the diagnosis of acute myocardial infarction (AMI) in patients admitted to the Coronary Care Unit at Tan Tock Seng Hospital between June and July 1998. METHODS: Routine blood samples sent to the laboratory for cardiac enzyme determination (CK, CKMB activity) were stored at -20 degrees C for later determination of CKMB mass (Abbott Axsym, Ortho Clinical Diagnostics (OCD) ECi and Roche Elecsys), Troponin I (Abbott Axsym) and Troponin T (Roche Elecsys). For CKMB mass measurements, the relative index (RI = CKMB mass/CK) was calculated. The diagnosis of acute myocardial infarction was obtained from inspection of clinical notes and/or discharge diagnosis for each patient. RESULTS: Forty-four of fifty-nine specimens were from AMI patients. Area under Receiver Operating Curve values were: CK 0.56, CKMB activity 0.72, percentage of CKMB activity 0.73, CKMB mass (Abbott) 0.76, CKMB mass (Roche) 0.77, CKMB mass (OCD) 0.78, RI (Roche) 0.83, RI (Abbott) 0.86, RI (OCD) 0.87, TnT 0.94, TnI 0.95. Sensitivity: TnI 88%, TnT 93%; specificity TnI 100%, TnT 92%. There was no significant difference in performance between Troponin T and Troponin I assays or between any of the CKMB mass measurements. CONCLUSION: Troponin T and I are superior to CKMB (mass or activity) and CK in the identification of patients with AMI. Combining multiple sampling of the percentage of CKMB with single confirmatory troponin testing may provide a cost-effective testing protocol for suspected AMI patients. PMID- 10709405 TI - Chronic haemodialysis with PTFE arterio-venous grafts. AB - BACKGROUND/AIM OF STUDY: End-stage renal failure (ESRF) patients requiring long term haemodialysis need a durable vascular access. The arterio-venous fistula (AVF) with its long patency rate and low complication profile is usually the first choice procedure for vascular access creation. However when superficial veins are not suitable for AVF creation or have all been exhausted as a result of repeated AVF procedures, arterio-venous grafts (AVG) using polytetraflouroethylene (PTFE) to bridge arteries and veins is an alternative for provision of continued vascular access for haemodialysis. This study is a review of our experience in using PTFE AVGs for vascular access in patients requiring chronic haemodialysis. METHODS AND MATERIALS: A retrospective review was done on 92 grafts in 77 patients placed by 3 vascular access surgeons at the Singapore General Hospital from January 1989 to December 1994. RESULTS: There were 58 female and 19 male patients with a median age of 43 years (range 15-76 years). Twelve patients (16%) were diabetic and 6 patients (8%) had systemic lupus erythematosis requiring long-term steroids. Seventy-three percent of patients had up to 2 previous AVF creations before placement of AVG over the forearm (64%), upper arm (23%) or thigh (6.5%). Complications include graft infection (19), pseudoaneurysm formation (10), graft thrombosis (24), steal syndrome (1), venous congestion (1) and venous end stenosis (1). Diabetic status and long-term steroid therapy did not significantly increase the incidence of graft infection. The patency rates at 24 months and 36 months were 77% and 58% respectively. However serviceability rates were 61% at 2 years and 38% at 3 years mainly due to infective complications. CONCLUSION: PTFE AVGs offer reasonable patency and serviceability rates as a vascular access modality but in view of their complication profile, the native vein arteriovenous fistula should continue to be the first choice procedure for vascular access in patients requiring chronic haemodialysis. PMID- 10709406 TI - Primary non-compliance in a Singapore polyclinic. AB - AIM OF STUDY: To determine the rate of patients not collecting their medication despite being given a prescription (primary non-compliance) in a Singapore polyclinic; to compare the results with similar studies done abroad and to identify the characteristics of patients most likely to be non-compliant. METHODS: Out of 500 prescriptions issued consecutively by the author, those that were not presented to the pharmacy for collection of medication were identified. The case sheets of these patients were then retrieved for further analysis. FINDINGS: The primary non-compliance rate in this study was 4.0%. Patients who were less than 30 years old or who consulted for an acute complaint were significantly more likely to be non-compliant. While the overall non-compliance rate was similar to that of studies done in the UK, the number of non-compliant patients with chronic illnesses was markedly lower in the local population. PMID- 10709407 TI - Comparison of the grade of CIN in colposcopically directed biopsies with that in outpatient loop electrosurgical excision procedure (LEEP) specimens--a retrospective review. AB - BACKGROUND: It has been suggested that LEEP should be the standard for the diagnosis of cervical dysplasia, rather than colposcopically directed biopsy as traditionally so, since it is both diagnostic and therapeutic. PATIENTS: Seventy eight patients who underwent LEEP at the Gynaecological Cancer Centre, KK Hospital from 1 January 1995 to 31 December 1997 for cervical dysplasia diagnosed by colposcopically directed biopsy were retrospectively reviewed. The mean age of the patients was 40.3 years (SD 8.4), with 78.2% of them with CIN I or CIN II. The mean operating time was 11.8 minutes (SD 4.9) and 53.8% (42/78) were given prophylactic antibiotics, with the only complication being moderate postoperative haemorrhage in 3.8% (3/78). The mean follow-up period was 19.1 months (SD 9.3) with the cure rate being 97.4% (76/78). RESULTS: Only half of the patients had corresponding histologies on biopsy and LEEP, with 28.2% (22/78) undergraded and 21.8% (17/78) overgraded. CONCLUSION: Significant discrepancies may be found between the results of colposcopically directed biopsy and loop excision, and LEEP, which is safe and effective may be the choice procedure for the diagnosis of cervical dysplasia. PMID- 10709408 TI - Benign cephalic histiocytosis in Singapore--a review of 8 cases. AB - AIM OF STUDY: Benign cephalic histiocytosis (BCH) is a rare, benign, self-healing papular eruption that affects mainly children. The aim of this study was to characterise our Singapore children affected by this condition and to review some of the recent literature on non-Langerhans cell histiocytoses. METHODOLOGY: The case records of all cases of BCH seen between January 1990 and December 1996 in our centre were retrieved and analysed. Further details not available in the case records were obtained via a telephone interview with the patients or their parents. RESULTS: A total of 8 cases were seen. There was no sex preponderance or ethnic predilection. The average age of onset was 29 months. All patients began with small brownish papules on the face and sometimes the upper trunk, and most of the lesions regressed in less than 84 months. CONCLUSION: BCH is a benign non Langerhans cell histiocytosis in which spontaneous resolution is the rule. A small skin punch biopsy is simple, diagnostic and helpful in its management. The management of BCE is expectant and reassurance is sufficient. However the regression and resolution of the lesions may take up to several years. PMID- 10709409 TI - Right and left bundle branch block in acute inferior myocardial infarction. AB - An 88-year-old man presented with acute Q wave inferior, posterior and right ventricular myocardial infarction which was associated with intermittent complete right and left bundle branch block. PMID- 10709410 TI - Posterior cerebral artery territory infarct presenting as acute psychosis. AB - An 82-year-old man with a past medical history of hypertension was admitted to a psychiatric hospital for sudden onset of acute psychosis. He was then transferred to an acute geriatric unit for further evaluation. During the admission the patient was noted to be very restless, agitated and noisy and was shouting and screaming incessantly. This was interspersed with occasional short periods of calm and quiet. Clinically, no obvious focal neurological deficits were detected. A CT scan of the brain was performed and it revealed an acute infarct involving the area supplied by the left posterior cerebral artery. This was a rather atypical presentation for an infarct involving this area. PMID- 10709411 TI - Simultaneous EEG and ECG recording during a Stokes-Adams attack. AB - Simultaneous electroencephalographic and electrocardiographic recordings were obtained from a 77-year-old patient during a Stokes-Adams attack. The recordings showed a clear temporal relationship between symptoms, electroencephalographic and electrocardiographic changes during the Stokes-Adams attack. This case shows the usefulness of simultaneous EEG and ECG recordings in the investigation of patients with unexplained episodes of disturbed consciousness. PMID- 10709412 TI - Treatment and prevention of sudden cardiac death--what have we learnt from randomised clinical trials? AB - Sudden cardiac death is most commonly caused by ventricular tachycardia or fibrillation. Three groups of patients at highest risk for sudden cardiac death are survivors of previous sudden cardiac death, those with recurrent documented episodes of sustained ventricular tachycardia and patients with recurrent syncope of unknown origin. The experience with antiarrhythmic drugs has been discouraging. Only beta-blockers have been shown to unequivocally reduce both arrhythmic and total mortality in randomised trials. Class I antiarrhythmic drugs increase mortality, especially in an ischemic substrate. Class III drugs such as sotalol and amiodarone have had variable success. Racemic sotalol has both beta blocker as well as Class III actions and some of the benefits may be due to the former effect. D-sotalol which has only pure Class III action, increases mortality in the post myocardial infarction patient. Amiodarone is superior to Class I antiarrhythmic drugs for patients with previous cardiac arrest. In the high-risk myocardial infarction patient, it seems to reduce sudden death but not total mortality. In the cardiac failure patient, the effect of amiodarone on total mortality is controversial. Several randomised trials of implantable cardioverter-defibrillator (ICD) therapy versus drugs have however concluded that the ICD is superior to drugs in reducing total mortality. In comparison with many other high volume therapies used in medicine today, ICD is still a cost-effective therapy. PMID- 10709413 TI - Clinics in diagnostic imaging (43). Right adrenal myelolipoma. AB - A 53-year-old woman who presented with drug-induced jaundice and urinary tract infection was incidentally found to have a large abdominal mass. Radiographs and ultrasonography showed a large fatty mass located between the right lobe of the liver and the right kidney. Diagnosis of right adrenal myelolipoma was made on computed tomography. The patient was treated conservatively. The causes of large fatty masses of the abdomen in adults are discussed, with emphasis on the imaging appearances of myelolipoma, renal angiomyolipoma, cystic teratoma and liposarcoma. PMID- 10709414 TI - Electrocardiographic case: a middle aged, seriously ill woman with an unusual ECG and wide complex tachycardia. PMID- 10709415 TI - Health without borders. PMID- 10709416 TI - No, no to the commercialization of medicine. PMID- 10709417 TI - Restorative neurology--making patients whole again. PMID- 10709418 TI - Orthopaedic surgery. PMID- 10709419 TI - Alzheimer's disease--the rising tide. PMID- 10709420 TI - Challenges in family medicine. AB - Family medicine has matured over the last 30 years. It has become a discipline but acceptance as a specialty is still not universal. It needs to pay attention to developing intellectual vigour and capacity building to meet peoples' needs. Opportunities to do so abound, namely in preventive care, geriatric care and care of chronic diseases. Future challenges lie in capacity building in practice, teaching and research. The setting up of a vocational register for trained family physicians and charging an adequate fee are important mechanisms to improve its image. PMID- 10709421 TI - Confidence and confidentiality--aspects of the law. PMID- 10709422 TI - Antenatal HIV screening--knowledge, attitudes and practices of obstetricians in KKH. AB - AIM OF STUDY: To determine the knowledge, attitudes and practices of obstetricians with regards to HIV screening in pregnant women. METHOD: This is a cross-sectional study based on a questionnaire survey of all obstetricians in KKH from January to August 1997. MAIN OUTCOME MEASURES: Obstetricians' knowledge and beliefs of HIV epidemiology and HIV perinatal transmission, and attitudes and practices with regards to antenatal HIV screening. RESULTS: Forty-one (77.4%) obstetricians responded to the survey. At the time of the survey, only 43.9% of the respondents had offered the HIV serology test to their patients with only 17.1% and 19.5% providing pre-test and post-test counselling respectively. Seventy-eight percent of them were aware of reports that zidovudine could reduce the vertical HIV transmission rate. All the respondents felt that HIV testing should be performed in pregnant women. The majority (70.7%) felt that antenatal HIV testing should be voluntary rather than mandatory and 56.1% felt that the patients' civil rights and confidentiality could be more assured if HIV testing is voluntary than if it was mandatory. Most respondents (56.1%) felt that antenatal HIV screening should be universally offered rather than targeted to those with risk factors. Most obstetricians did not feel comfortable (58.5%) or competent (80.5%) to manage HIV infection in pregnancy. Those who felt competent were more likely to feel comfortable, more likely to have provided HIV test in the clinic, and less likely to opt out of caring for an HIV-infected patient. CONCLUSION: The majority of the surveyed obstetricians would support a program of voluntary antenatal HIV screening that is universally offered to all pregnant women. The feelings of discomfort and incompetence of the obstetricians towards caring for an HIV-infected pregnant woman need to be addressed further. There is a need for continuing medical education to help obstetricians keep abreast with the advances in HIV screening and its management. PMID- 10709423 TI - One week triple therapy for Helicobacter pylori associated duodenal ulcer disease. AB - INTRODUCTION: Eradication of Helicobacter pylori (H. pylori) cures and prevents the relapse of duodenal ulceration. Different treatment regimes for the eradication of H. pylori have been used and the most successful eradication regimens have been one week treatments with a proton pump inhibitor and two antibiotics. AIM OF STUDY: To examine the eradication rate of H. pylori with a one week regimen consisting of OCT (Omeprazole 20 mg BD, Clarithromycin 250 mg BD, Tinidazole 500 mg BD). This treatment regimen has been used for H. pylori eradication in our department since the end of 1996. METHODS: Patients diagnosed to have duodenal ulcer in 1997 were retrospectively reviewed. Infection with H. pylori must be documented either by gastric biopsy or by a positive CLO test. Eradication of H. pylori was confirmed by negative 14C urea breath test or by histology at least four weeks after cessation of therapy. RESULTS: The review was performed on 251 patients. There were 177 males, 74 females. The median age was 51 (18-77) years. H. pylori infection was confirmed by CLO test in 170 patients and by histology in 72 patients. Thirty patients did not undergo further investigation after therapy to confirm the eradication. Of the remaining 221 patients, H. pylori was successfully eradicated in 198 patients (89.6%) as confirmed by 14C urea breath test (190 patients) or repeat gastroscopy and gastric biopsy (31 patients). There were no serious adverse events documented. CONCLUSIONS: Our retrospective study showed that the one week regimen used in our department is effective for the eradication of H. pylon in nearly 90% of infected cases. PMID- 10709424 TI - The presentation of elderly people at an emergency department in Singapore. AB - OBJECTIVE: This study aims to provide demographic and clinical data of elderly people attending one of the hospital emergency departments in Singapore. DESIGN: Patients aged 60 years and above who attended the Emergency Department (ED) at Alexandra Hospital, Singapore, during 4 randomly selected one-week periods in 1996 were retrospectively studied. MAIN OUTCOME MEASURES: Patient profile; presenting symptoms; diagnoses; types of investigations done, and outcome following attendance. RESULTS: A total of 455 ED attendance cards were analysed. The age of patients ranged from 60 to 102 years with a mean age of 72.8 years. Two hundred and sixty-one (57.4%) were males and 194 (42.6%) were females. Four hundred and twenty-seven (93.9%) were emergencies, 25 (5.5%) were non-emergencies and in 3 (0.6%) the priority rating was unknown. Two hundred and ninety (63.7%) were admitted, of whom 166 (57.2%) were males. One (0.2%) was admitted for social reason. The 3 most common symptoms were abnormalities of breathing (10.6%), falls (9.4%) and musculoskeletal pain (8.2%). Majority had 1 (40.4%) or 2 (41.6%) symptoms. The symptoms were mainly acute (1 day, 45.2%) or less than a week (25.7%). The 3 most common diagnoses were chest infection or pneumonia (8.2%), non-fracture head injury (7.2%) and heart failure (6.6%). Most patients (90.5%) had only 1 diagnosis. The 3 most common tests done were chest X-ray (172 patients, 37.8%), electrocardiography (119 patients, 26.2%) and blood glucose (86 patients, 19.0%). One hundred and twenty (26.4%) patients did not require any investigation. CONCLUSION: The elderly constituted 12.4% of attendance at the ED but formed 34.5% of admissions. They were more likely to have emergency problems. Understanding the common presenting symptoms and diagnoses of the elderly will help doctors at the ED provide better care. Elderly patients with complex problems who are not hospitalised would probably benefit from further geriatric assessment. PMID- 10709425 TI - Pacemaker implantation in Singapore in 1997. AB - INTRODUCTION: Previous reports on pacemaker implantation in Singapore have been from a single institution and hence may not accurately reflect the practice in Singapore. As part of the World survey on pacemaker implantation, a survey of all pacemaker implantations in Singapore in 1997 was performed. METHOD: Information was obtained from the pacemaker manufacturers and a survey form was sent to all doctors involved in pacemaker implantation. RESULT: In 1997, 206 pacemakers were implanted or replaced in Singapore. This gives a pacemaker rate of 69 per million. For new implants only, there were 61 implants per million. More detailed information regarding the patient and implantation was obtained in 160 (78%) patients. The mean age of the patients was 68.5 +/- 14.4 years (range 2-97 years). There were 142 (89%) new implants and 18 (11%) replacements. 62.5% of the patients were females. Seventy-nine percent of the patients were older than 60 years old and 17.5% were older than 80 years. Seventy-five percent of the pacemakers were single chamber pacemakers. Twenty-five percent were dual chamber pacemakers. Only 1.4% of the pacemakers used epi-myocardial leads and all these were in children. Heart block was the most common indication for pacing and consisted of 52.8% of the patients while 43.0% of patients were implanted for the sick sinus syndrome. CONCLUSION: Pacemaker implantation in Singapore in 1997 was 69 per million. Heart block remains the most common indication for implant and single chamber pacing is still the most commonly used mode of pacing. The majority of the implants were in persons older than 60 years. With an increasing ageing population in Singapore, the implant rate for pacemakers will be expected to increase significantly. PMID- 10709426 TI - Does the piriformis compress the sciatic nerve during limb length equalization? AB - BACKGROUND: Correction of limb length inequality during reconstruction procedures at the hip has the inherent danger of sciatic nerve injury. Among others, we think that constriction by a tightened Piriformis tendon may be a significant cause during longitudinal traction at the time of surgery. PATIENTS: A specific protocol involving the use of staged surgical release at the time of intraoperative traction/limb length equalization gave only partial nerve compromise in one out of 11 cases; three other patients treated without this protocol developed sciatic nerve problems. CONCLUSION: The authors feel that prior to the application of longitudinal traction, routine sectioning of the contracted piriformis tendon is important to prevent this complication; it is suggested that further investigations like intra operative nerve conduction studies during traction may perhaps substantiate these findings. PMID- 10709427 TI - Nafarelin acetate for pituitary suppressions in in-vitro fertilisation cycles--a Singaporean experience. AB - AIM OF STUDY: The aim of this prospective clinical trial was to determine if intranasal nafarelin acetate (NA) is as effective as leuprolide (LA), our standard GnRHa, in IVF cycles. In addition, we believe that this may be the first report of such a trial in an Asian IVF population. METHOD: Midluteal GnRHa administration (LA = 0.5 mg/d; NA = 800 micrograms/d) was used with a standardised hMG ovarian stimulation protocol for all 88 consecutive cycles, randomised at recruitment. RESULTS: There were no significant differences between LA and NA in terms of the mean duration of agonist to reach pituitary suppression, total hMG dosage, number of embryos produced or frozen and the clinical pregnancy rate (LA = 21.4% and NA = 16.3% per cycle). CONCLUSION: Intranasal nafarelin acetate was as effective as leuprolide acetate in our series of IVF patients of Asian origin, and may be offered as an alternative choice for pituitary suppression. PMID- 10709428 TI - Clinics in diagnostic imaging (44). Testicular tumour with retroperitoneal lymphadenopathy and inferior vena cava thrombosis. AB - A 20-year-old Indian man presented with a two week history of non-specific abdominal pain. Abdominal ultrasonograpy incidentally detected a thrombus in the inferior vena cava (IVC). Computed tomography revealed the presence of extensive para-aortic lymph node disease as well as a filling defect in the IVC. Scrotal ultrasonography located a heterogeneous intra-testicular tumour in an otherwise palpably-normal testis. The extent of the IVC thrombus was evaluated by the use of magnetic resonance imaging. Inguinal orchidectomy was performed and histology revealed a non-seminomatous germ cell tumour. Combination chemotherapy led to complete resolution of lymph node disease and IVC thrombus. The patient remained well 9 months after diagnosis. The causes of IVC obstruction, role of imaging in investigating IVC obstruction and the management of tumour involvement of the IVC are discussed. PMID- 10709429 TI - Ambulatory care education. PMID- 10709430 TI - Promote self-esteem in young girls and women. PMID- 10709431 TI - [Colorectal liver metastases: does the number of metastases determine of resection is oncologically indicated?]. AB - Complete surgical removal of colorectal liver metastases offers the patient the only curative option as long as there is no extrahepatic spread. Apparently the number of parameters of prognostic significance is unlimited. Consequently their importance for the individual prognosis remains questionable. One of the most frequently discussed parameters is the number of lesions in the liver. The aim of this study was therefore to review the literature and to analyze our own patients with colorectal liver metastases. For 302 patients a complete follow-up was available. The patients were grouped according to the number of lesions and the status of resection (R0, R1, R2, Exploration). The groups were compared by Kaplan Meier Analyses with log rank tests. The literature was reviewed until December 1998. The number of metastases was confirmed to be of prognostic value. Three or more metastases made a resection without residual tumor more unlikely than if there were only two or less (17.8% versus 67.2%). 5-year survival after curative resection was reduced in the group with more metastases from 36% to 9% and 10 year survival was reduced from 18% to 0% without reaching statistical significance (p < 0.07). The number of liver metastases may have some prognostic impact for patients after colorectal cancer. However, even if more than four metastases were resected with clear margins some patients did survive for a long time. The most important parameter to predict an oncological benefit is resection without residual tumor. If this is technically and functionally possible the patient should not be denied surgery, his only chance for cure. PMID- 10709432 TI - Relevance of locoregional chemotherapy in patients with liver metastases from colorectal primaries. AB - Progression of colorectal cancer can occur primarily isolated in the liver. But, only the minority of the affected patients is eligible for surgery. Initially, systemic chemotherapy was ineffective in the treatment of unresectable hepatic metastases. For this reason, intraarterial chemotherapy was introduced as treatment alternative to the systemic chemotherapy. Long-term intraarterial chemotherapy regimens with FUDR in patients with colorectal liver metastases, using implantable pumps and ports, resulted in improved response rates, which was confirmed by several randomized trials. However, an improvement in median survival has not yet been demonstrated after regional chemotherapy of hepatic metastases. Since the intraarterial therapy with floxuridine (FUDR) had been reported to result in a high rate of local toxicity, 5-fluorouracil (5-FU) was introduced into regional chemotherapy of the liver. A randomized trial demonstrated superiority of intraarterial 5-FU versus intraarterial FUDR therapy. Despite these reports about high response rates, the benefit of intraarterial chemotherapy remains questionable, because it has not yet resulted in a prolongation of median survival. For this reason, long-term regional chemotherapy cannot be considered as standard treatment and should therefore not be conducted outside controlled clinical trials. Further evaluations on this technique should only be performed in experienced centers. PMID- 10709433 TI - The effects of dimethylsulfoxide on experimental hepatic ischemia. AB - AIMS: The aim of the study is to investigate the effects of dimethylsulfoxide (DMSO), a non-enzymatic free oxygen radical detoxifier, on the alterations observed during hepatic ischemia. METHODS: Twenty four albino rabbits were entered into the study. DMSO (500 mg/kg) was administered through inferior vena cava following dissection of the portal triad and immediately prior to clamping. The alterations on liver glycogen, blood glucose, ALT, AST, LDH, intracellular ATP, GSH-px, SOD, MDA within the erythrocyte and hepatic tissue MDA were investigated. RESULTS: In the control group, following ischemia, a reduction in blood glucose, hepatic glycogen and ATP levels within the cell and an increase in AST and LDH levels, MDA, GSH-px and SOD levels within the erythrocyte and hepatic tissue MDA level were observed (p < 0.01). In the study group, the blood glucose values increased at 30th minute following ischemia. The increase in LDH level was not statistically significant (p > 0.05). The increase in AST level was significantly lower when compared to the control group (p < 0.05). No effect was observed on ALT and ALP levels. DMSO lowered the reduction in hepatic glycogen level (p < 0.05), GSH-px (p < 0.01), MDA (p < 0.05) and ATP (p < 0.05) levels within the erythrocyte and hepatic tissue MDA level (p < 0.01) that were increased following ischemia. The increase in blood SOD level was not statistically significant (p > 0.05). CONCLUSION: DMSO can be administered in attempt to prolong the duration of ischemia or to reduce the adverse effects of ischemia on the hepatic tissue during the existing ischemic period. PMID- 10709434 TI - [Incarcerated and strangulated hernias--surgical approach and management]. AB - Acute symptomatic groin hernias with potential or definite ischemia represent a special group of all the groin hernias. The method of choice to treat these hernias has to fulfill the following criteria: 1. Easy reduction of the hernia sac and its contents without causing damage. 2. Good exposure and easy access for possible resection. 3. Safe hernia repair through the same access. According to our experience with 44 incarcerated and strangulated groin hernias operated between 1993 and 1997 and after a literature review, we took the following procedure as our routine: Posterior approach and mesh repair. We do not use a meshgraft only in the presence of colonic necrosis or peritonitis. PMID- 10709435 TI - [Mesenteric cyst as the etiology of abdominal pain--a case report]. AB - INTRODUCTION: Mesenteric cysts are part of the differential diagnosis of abdominal tumors. We want to remember this diagnosis with the following case report. CASE REPORT: A 35-year-old woman was admitted for abdominal pain that had begun two weeks previously. Sonographic examination and CT scan of the abdomen showed a 14 x 12 x 3 cm abdominal tumor without any relation to the uterus, adnexa or organs of the epigastrium. Laparotomy was performed and the cystic tumor removed. DISCUSSION: Mesenteric cysts are rare. The pathogenesis is not uniform and the clinical and radiologic diagnosis is difficult. The symptoms of this condition vary from acute abdominal signs to non-specific abdominal features or incidental findings. Mesenteric cysts can be located anywhere in the mesentery from the duodenum to the rectum. The treatment of choice is resection. CONCLUSIONS: Mesenteric cysts are rare abdominal conditions. The resection of the cyst and the verification of the diagnosis is the treatment of choice. PMID- 10709436 TI - ["Recurrent appendicitis" 19 years after perityphlitic abscess]. AB - We report about a patient who was admitted with acute lower right quadrant pain. She underwent an undetermined operation for appendiceal abscess formation 19 years ago. Our investigations including ultrasound, CT-scan, conventional radiography and finally coloscopy revealed a pericoecal mass formation. Due to worsening of the symptoms, emergency laparotomy was performed. An inflammatory process and a partial necrosis of the coecum wall at the site of the appendix basis were identified und managed by ileocoecal resection and drainage. We took this case to review the literature concerning the treatment of appendiceal abscess and appendiceal mass, and consecutively redesigned our own treatment concept. PMID- 10709437 TI - [Quality control in patient education. Results of a patient survey about the patient education protocol of the Swiss Society of Surgery in 6 Swiss hospitals]. AB - In the past years the number of malpractice suits due to lack of patient information has increased. Because there have been no generally accepted guidelines for preoperative patient information, the Swiss Society of Surgery has decided to work out an informative brochure tailored to the needs of patients. It takes into account that the need to know beforehand is increasing rapidly. In collaboration with the judiciary service of the Swiss Medical Federation all the items and points of legal relevance have been compiled to establish an informative brochure. Based on this protocol, patients in surgical departments of 6 Swiss community hospitals were asked before discharge to qualify the preoperative information offered to them. 2660 questionnaires were evaluated. The majority of patients considered the information regarding their diagnosis, the complications, risks, treatment and postoperative care, the sketches describing the operation and the overall degree of information as good or very good. Almost 60% of all patients stated that no alternative treatment had been discussed with them other than the planned procedure. In most of these patients operative procedures were chosen and carried out for which there were few or no other acceptable options. 2/3 of the patients asked for immediate preoperative written information, especially if they had malignant disease. Barely 4% of the patients were not reassured by the information provided to them. The fact that 2/3 of all patients re-read the informative protocol before the operation underlines how important it is to hand out a copy of the protocol to satisfy the informative needs of the patients. To our surprise the vast majority of patients uttered little concern about giving their signature to forms that were presented to them. Only 2% of the patients felt that giving a signature would cause them grave reservations. The informative protocol devised by the Swiss Society of Surgery is well adapted to the informative needs of the patients and allows for a structured conversation. It facilitates documentation and offers valid legal proof for the physician that he/she has provided adequate information. PMID- 10709438 TI - [Undesirable effects of spa treatments: apropos of a systematic, prospective spa surveillance study at Bagneres-de-Bigorre]. AB - The aim of the study was a systematic prospective survey of adverse events at health resorts using the French method of assessment of imputability of adverse drug reactions. The work was performed over a period of one month in Grands Thermes at Bagneres-de-Bigorre (France, Pyrenees). Seventy-one adverse events were recorded in 1794 patients, i.e. 0.3 pour cent per day of treatment. Only one adverse event was considered as 'serious'. Six adverse events were evaluated as 'possible' and six others 'likely'. Most of the adverse events were general or neurological (such as asthenia, malaise) and were observed during the six first days at health resorts. These data show that health resorts in Bagneres-de Bigorre are associated with a low number of adverse events. They show that an epidemiological survey of health resorts is possible. This work underlines the necessity of genuine surveillance for a modern evaluation of the benefit/risk ratio of health resorts. PMID- 10709439 TI - [Role of suindac in the treatment of familial adenomatous polyposis coli]. AB - Familial adenomatous polyposis is a rare genetic disease characterized by the development of colorectal adenomatous polyps. Extracolonic digestive and extra digestive manifestations can also appear. Inevitably, colorectal cancer occurs if a colectomy is not performed. Sulindac is an indolic non-steroidal anti inflammatory indole drug which decreases colonic tumoral proliferation. The different trials published since 1983 have shown that sulindac caused regression of colorectal adenomatous polyps, but it does not affect the other manifestations of familial adenomatous polyposis. However, colorectal polyps recurred after cessation of this therapy; the effect of long-term sulindac therapy is unknown; and sulindac may cause, as a non-steroidal anti-inflammatory drug, digestive side effects. Moreover, treatment with sulindac does not completely eliminate the risk of cancer. For patients with familial adenomatous polyposis, total colectomy and ileal pouch-anal anastomosis is the recommended procedure for most patients. However, sulindac is useful for patients who have had subtotal colectomy and ileorectal anastomosis, if these patients have only a few rectal stump polyps and accept regular and strict follow-up of the rectal stump. Sulindac is also indicated for patients who have not undergone colectomy because surgery is contraindicated or has been refused. PMID- 10709440 TI - [Awareness of 125 patients of the rules for using their non-steroidal anti inflammatory agent]. AB - A total of 125 inpatients from a rheumatology unit were interviewed to assess their awareness of the rules for using a prescribed NSAID. Two per cent were unaware that they were receiving an NSAID, and 13 per cent and 71 per cent respectively did not know the name or the dosage. Twenty-two per cent admitted excessive intake, and 2 per cent simultaneous intake of two NSAIDs. Some declared that in the absence of adequate improvement they would take another NSAID (17 per cent) or aspirin (26 per cent) simultaneously. Fifty-nine per cent were unaware that aspirin is an anti-inflammatory drug, 3 per cent thought that taking a double dosage on alternate days was safe, and 46 per cent that taking a stronger dose at the beginning of treatment was an efficient and safe procedure. The replies of patients who had read the directions for use (69 per cent) did not differ from those of patients who had not (31 per cent). Fourteen per cent of patients declared that they would not advise their physicians after buying NSAIDs over the counter. PMID- 10709441 TI - [Renal tolerance of cidofovir]. AB - Renal failure, proteinuria and proximal tubular acidosis are the features of cidofovir renal toxicity, its main side-effect. Proteinuria occurs in more than 40 per cent of patients and correlates with early renal dysfunction. A fall in serum potassium, bicarbonate, uric acid, calcium and phosphorus levels associated with glucosuria is the hallmark of proximal tubular acidosis. Most of the patients exhibit only glucosuria. Renal failure, diagnosed in 12 per cent of treated patients, is a late feature, usually discovered after the onset of proteinuria and glucosuria. Prevention of cidofovir-induced renal toxicity involves a search for other risk factors, probenecid treatment, and requires an optimal hydration status. PMID- 10709442 TI - [Evaluation of the effect of tobacco on trandolapril tolerance]. AB - The objective of this study was to evaluate the impact of smoking habits on safety of trandolapril assessed by interrogation and by visual analogue scales (VAS). A total of 3402 hypertensive smokers (> or = 1 cigarette/d for at least 6 months) and non-smokers (no smoking or ceased at least 6 months previously) received trandolapril 2 mg/d for 4 weeks. The safety profile of trandolapril was assessed by both interrogation and by VAS. The VAS completed by the patients at D0 and D28 explored the following symptoms: asthenia, nausea, cough, headaches and dizziness. A significant change in cough VAS was previously defined by an at least 19 mm change. VAS analysis was performed on 2840 patients (1296 smokers and 1544 non-smokers), mean age 59 +/- 12 years. Smokers and non-smokers were significantly different for age 56 +/- 12 years vs. 62 +/- 12 years, sex ratio 74 per cent males vs. 45 per cent, history of hypertension 4.5 +/- 6.1 years vs. 5.3 +/- 6.5 years and cough VAS score at D0 35 +/- 26 mm vs. 20 +/- 21 mm. In the total population, 214 adverse events were reported by 177 patients (5.2 per cent). The most frequent adverse events were a cough (2.1 per cent), bronchitis (0.6 per cent), headaches (0.5 per cent), rhinitis (0.4 per cent), nausea (0.4 per cent) and asthenia (0.3 per cent). Cough was reported by 23 smokers (1.5 per cent) and by 49 (2.6 per cent) non-smokers (p = 0.02). In the VAS population, 151 adverse events were reported by 130 patients, 47 smokers (3.6 per cent) and 83 non-smokers (5.4 per cent, p = 0.03). The difference between the two groups was mainly due to a cough: 15 smokers (1.2 per cent) reported a cough vs. 38 non smokers (2.5 per cent, p = 0.01) and 77 smokers (5.9 per cent) presented a significant change of cough VAS score vs. 124 non-smokers (8.0 per cent, p = 0.03). In this large scale study, 1.9 per cent of patients treated with trandolapril exhibited a cough. Smokers were less likely to present a cough. Use of VAS confirmed this trend. PMID- 10709443 TI - [Acute toxicity of 10 Moroccan plants reported to be hypoglycemic agents]. AB - An acute toxicity study of twelve medicinal plants on Swiss albino mice has been carried out. The extracts were administered orally at a dose of 1 g/kg body weight. Animals were observed for 7 days and changes in weight and behaviour were noted. On the eighth day, the animals were sacrificed and an anatomo histopathologic survey was undertaken. The results showed a change in behaviour and some histologic modifications following the administration of certain plant extracts. PMID- 10709444 TI - Toxicology of Astragalus lusitanicus Lam. AB - Astragalus lusitanicus is a toxic legume grown in Morocco and in some other Mediterranean countries. In small ruminants, poisoning by this plant is dominated by nervous signs characterized by many cycles of excitement-depression. Macroscopic examination of poisoned animals showed congestive lesions and oedema in the brain and lungs. Microscopic lesions consisted mainly of vacuolar degeneration in neurons, hepatocytes and in spleen and kidney cells. Serum activity of AST and CK as well as blood glucose and urea were increased as a result of poisoning. However, serum activity of alpha-mannosidase was not modified as is the case in locoism. Chemical investigations showed that A. lusitanicus does not contain swainsonine or miserotoxin and its selenium concentration is very low. However, this legume contains indolizidin alkaloids and a first compound was purified and identified. PMID- 10709445 TI - Pharmacological studies of two antidiabetic plants: Globularia alypum and Zygophyllum gaetulum. AB - Investigations were carried out to evaluate the hypoglycaemic activity of the infusions of Globularia alypum and Zygophyllum gaetulum. Oral and intraperitoneal administration of the plants (0.7 g/kg) produced a significant hypoglycaemic effect in normal as well as in hyperglycaemic rats. The infusions increased significantly plasma insulin levels in normal rats. It is suggested that the hypoglycaemic activity of these plants may be mediated through enhancement of peripheral metabolism of glucose and an increase in insulin release. PMID- 10709446 TI - Effects of Olea europea var. oleaster leaves in hypercholesterolemic insulin resistant sand rats. AB - Sand rats fed a hypercaloric diet manifest obesity and diabetes. We have used this model to develop hypercholesterolaemia and describe the beneficial action of Olea europea var. oleaster leaves. Twenty-eight sand rats submitted to a high cholesterol diet for four months were assigned to control and treated groups. Plant decoction at 10 per cent was given orally for two months. Results showed that the control group exhibited hyperglycaemia, glucose intolerance, hypercholesterolaemia and moderate hyperinsulinaemia. Light microscopic study showed thickening of capillary walls in skin, pancreas and kidney. The treatment produced hypoglycaemic (43 per cent, p < 0.001), antihyperglycaemic (48 per cent, p < 0.001) and hypoinsulinaemic (39 per cent, p < 0.01) activities. In addition, the plant presented a hypocholesterolaemic effect (47 per cent, p < 0.001) accompanied by lowering of oxidized LDL (30 per cent, p < 0.01). Accordingly, capillary wall thickening was reduced in skin and pancreas and completely prevented in kidney. The data demonstrate that oleaster leaves possess at least two active compounds to treat hypercholesterolaemia and diabetes. PMID- 10709448 TI - Antibacterial and antifungal activities of Cistus incanus and C. monspeliensis leaf extracts. AB - In this study, the antimicrobial activity of leaf extracts obtained from two species of genus Cistus L. was examined in vitro against five strains of bacteria and five strains of fungi. The species studied are Cistus villosus L. = incanus and Cistus monspeliensis L. All extracts showed inhibitory activity against microorganisms. These results encourage us towards further biological investigation. PMID- 10709447 TI - Effect of Suaeda fruticosa aqueous extract in the hypercholesterolaemic and insulin-resistant sand rat. AB - To study the effect of Suaeda fruticosa in lipid and carbohydrate metabolism in the hypercholesterolaemic and insulin resistant sand rat, 25 rats were subjected to a high cholesterol diet for 90 days. On the 45th day the animals were divided into two groups: control and treated. Aqueous extract prepared in infusion at 10 per cent was administered orally at 1.5 ml per 100 g of body weight per day. On the 90th day, the control group developed a severe hyperlipidaemia, impaired glucose tolerance test and insulin resistance. Treatment by Suaeda fruticosa extract showed hypoglycaemic (41 per cent) and antihyperglycaemic (53 per cent) effects. Furthermore, the plant led to a decrease in plasma levels of insulin (31 per cent), total cholesterol (50 per cent), LDL cholesterol (55 per cent), VLDL cholesterol (49 per cent), oxidized LDL (40 per cent), lecithin cholesterol acyl transferase (44 per cent), triglycerides (57 per cent) and free fatty acids (36 per cent). We concluded that Suaeda fruticosa aqueous extract contains at least two compounds responsible for hypoglycaemic and hypolipidaemic activities. PMID- 10709449 TI - Effects of saponins from Herniaria glabra on blood pressure and renal function in spontaneously hypertensive rats. AB - Experiments were performed on male and female spontaneously hypertensive rats weighing 310-340 g (10 animals per group). The oral administration of 200 mg/kg/day of saponins from Herniaria glabra for 30 days, resulted in a significant decrease in blood pressure in hypertensive rats. The systolic and diastolic blood pressure decreased significantly and respectively from 187.60 +/- 20.63/119.00 +/- 7.09 mmHg at day 0 (D0) to 141.60 +/- 7.51/90.40 +/- 7.68 mmHg at day 30 (D30), p < 0.001 (vs. 186.30 +/- 11.27/114.10 +/- 12.00 mm Hg at D0 to 154.50 +/- 6.38/132.3 +/- 7.68 mmHg at D30 in furosemide-treated group, p < 0.001). Control animals receiving placebo did not show any significant variation in the mean arterial pressure. The effect of saponins of Herniaria glabra on renal function was evaluated in spontaneously hypertensive rats using clearance techniques. Glomerular filtration rate was constant in the control rats and increased significantly in the hypertensive rats after saponins treatment (5.55 +/- 0.32 vs. 6.03 +/- 0.43 ml.min-1.kg-1 in the control (C) and saponins (S) groups, respectively, p < 0.05). Saponins administration provoked an increase in urinary flow (59.38 +/- 5.85 ml.kg-1.24 h-1 vs. 36.92 +/- 5.17 ml.kg-1.24 h-1, p < 0.001). Saponins also increased potassium excretion (6.89 +/- 0.81 mmol.kg-1.24 h-1 vs. 5.40 +/- 0.51 mmol.kg-1.24 h-1, p < 0.001) and sodium excretion (10.74 +/ 1.21 mmol.kg-1.24 h-1 vs. 7.25 +/- 0.54 mmol.kg-1.24 h-1, p < 0.001) as well as chloride excretion (13.59 +/- 1.04 mmol. kg-1.24 h-1 vs. 9.67 +/- 0.77 mmol.kg 1.24 h-1, p < 0.001). It is concluded that chronic oral administration of saponins from Herniaria glabra decreased the arterial blood pressure and affected salt and water transport in renal tubules. PMID- 10709450 TI - [Update of a database on plants involved in the composition of 825 drugs: Pharmaplantes-Kenitra 98]. AB - The main objective of this work lies in the setting up of a database on plants used in medicines which is aimed at pharmacological development of plant resources in Morocco. We have, as a first step, made an inventory of different plant species involved in medicine making in Morocco. This survey dealt with 825 miscellaneous pharmaceutical products and reveals the use of 445 different plant species in medicine making. For each plant species, we have also noted the parts of the plant which are used in the pharmaceutical industry. Furthermore, we have taken an interest in plant extracts involved in this medicine making. The results show that in all these medicines contain 46 extracts of a vegetal nature. At the top of the list, menthol is used in the manufacture of 110 pharmaceutical products. Finally, an analysis per pharmaco-therapeutic family reveals the impact of the involvement of these plants on each of these families. In fact, it turns out that 204 plant species play a part in medicines classified in the family of gastro-entero-hepatology whereas only one plant is involved in anti-inflammatory medicines. PMID- 10709451 TI - [Anticoagulant activity of coumarins from Ferula communis L]. AB - Ferula communis is an ombelliferous plant of the Mediterranean regions. It is represented in Morocco by two varieties: brevifolia and genuina. The later is very rich in a soap or resinous gum. This product, collected from the roots, is largely used in traditional medicine. It is know as fessoukh in Morocco and other Arab countries. This plant is also well known for its toxicity and its anticoagulant activity. In the present review, are discussed: (1) the ethnobotany of the plant, especially medicinal uses of fessoukh in traditional medicine as well as alimentary use of young stems as legumes; (2) clinical and biochemical data of intoxication by this plant, which are dominated by haemorrhage as a consequence of blood coagulation disturbance; (3) 4-hydroxycoumarins isolated from Ferula communis L. and their anticoagulant activity; (4) the role of vitamin K1 in the treatment of poisoning by this plant. PMID- 10709452 TI - Antitumour principles from Peganum harmala seeds. AB - From ancient times, Peganum harmala was claimed to be an important medicinal plant. Its seeds were known to possess hypothermic, and essentially hallucinogenic properties. Various authors have undertaken studies on the antibacterial, antifungal and antiviral effects of Peganum harmala seeds, but studies on the antitumour activity are not to be found in the literature. In Moroccan traditional medicine, seed powder is sometimes used on skin and subcutaneous tumours. This work was designed to investigate some aspects of the antineoplastic properties of the plant Peganum. Varying concentrations (10 to 120 micrograms/ml) of total alkaloid extracts of Peganum harmala seeds (collected in Morocco) were tested in vitro on four tumoural cell-lines: Med-mek and UCP-Med carcinoma, UCP-Med sarcoma and Sp2/O-Ag14. In vivo experiments were performed with the Sp2/O cell-line grafted subcutaneously in syngenic BALB/c mice. In vitro, proliferation of tumoural cell lines was significantly reduced by all tested concentrations of the Peganum alkaloid extracts during the first 24 h of contact. A cell lysis effect occurred after 24 h and progressed to complete cell death within 48 to 72 h depending on the alkaloid concentration. Results obtained indicate that alkaloids of Peganum have a high cell toxicity in vitro. The active principle at a dose of 50 mg/kg given orally to mice for 40 days was found to have significant antitumoural activity. Peganum harmala alkaloids thus possess significant antitumour potential, which could prove useful as a novel anticancer therapy. PMID- 10709453 TI - Preliminary screening of antiprotozoal activity of extracts from Cotula cinerea L. AB - Ethyl ether, ethyl acetate and n.butanol extracts of Cotula cinerea L. were tested for their antiprotozoal activity against two species of Trichomonas: Trichomonas intestinalis and Trichomonas vaginalis. It has been found that the growth of both Trichomonas was significantly inhibited. PMID- 10709454 TI - Screening of antibacterial and antiparasitic activities of six Moroccan medicinal plants. AB - The extracts of six plants selected on the basis of folk-medicine reports were examined for their antibacterial effects against eight pathogenic bacteria. The results showed that n-butanol extract of Calotropis procera proved to be the most effective against the bacteria tested using the paper disc diffusion method. The antiprotozoal activity was also examined and showed that ethyl ether extract of Sium nodiflorum exhibits a parasiticidal effect against Trichomonas intestinalis and vaginalis. PMID- 10709455 TI - [Serotoninergic syndrome after combining tramadol and iproniazid]. PMID- 10709456 TI - Citalopram pharmacokinetic interaction with clomipramine. UDP glucuronosyltransferase inhibition? A case report. PMID- 10709457 TI - [Accidental ingestions of paracetamol in the form of EFFERALGAN pediatric syrup: experience of the Marseille Anti-poison Center during 1998]. AB - In 1998, 77 cases of accidental ingestion of paracetamol paediatric syrup (Efferalgan) in children were notified to the Marseille Poison Centre. In a quarter of them, the alleged dose taken was greater than the toxic dose. Ingestion was mainly due to the child opening the bottle. The proximate marketing of a product with a child-proof top, which should allow the number of accidents to be reduced. Doctors and pharmacists should be informed rapidly, so that they can warn the families who still have the old type of bottle. PMID- 10709458 TI - [Monitoring of life-threatening infection pathogens in relation to the problem of prediction of critical situations]. AB - The epidemic situation in the context of many infectious diseases caused by bacteria is presently assessed as being poor in Russia and other countries. The spectrum of the pathogens that can deteriorate epidemic well-being is highly wide. The epidemic situation in terms of many infectious diseases, including those caused by such causative agents as Bacillus anthracis, Vibrio cholerae, Yersinia pestis, Francisella tularensis and others may deteriorate due to the emergence of their modified forms owing to their specific variability. The above generates the necessity of improving controlling measures or developing the techniques for monitoring the pathogens of infectious diseases, including those in the framework of international cooperation. PMID- 10709459 TI - [Elements of rickettsiosis pathogenesis in the light of currently available data]. PMID- 10709460 TI - [Problem in the diagnosis of anaerobic infections and dysbacteriosis in clinical dentistry]. AB - The fluorescence technique was proposed for rapid diagnosis of anaerobic infection. It determines the microbiocenosis of a wound, the gastrointestinal tract, maxillofacial region, and other pathologically changed organs and tissues. The technique allows one to follow the time course of changes in the microflora and in the pathological process during the treatment and follow-up of a patient. An attempt was made to establish a rapid diagnosis of intestinal dysbacteriosis and to evaluate its severity. A real-time feed-back unit was developed for the diagnosis of anaerobic infection. An anaerobic microflora model in dental caries was used to work out criteria for the informative value of the proposed technique and it was clinically tested. PMID- 10709461 TI - [Design of human liposomal recombinant alpha2-interferon-containing preparation]. AB - The composition of liposomal lipid phase preparation was selected. The optimum ratio of liposomal lipid phase components was found to produce a a human liposomal recombinant alpha 2-interferon-containing preparation by extrusion across the polycarbonate membranes. The study gave rise to a lyophilized sterile liposomal recombinant alpha 2-interferon preparation. The agent was shown to be stable and retains its antiviral activity within a year. PMID- 10709462 TI - [Staphylococcus aureus-lysing enzymes: isolation and some properties of lysostaphine]. AB - The authors investigated a procedure for isolating lysostaphine, a complex of enzymes that lyse the cells of virtually any S. aureus strains in the fermenter and large bottles. It was shown that the activity of the enzyme in the fermenter might be much higher than in the flask. The enzyme can be concentrated on the hollow fibers, purified on the ion exchanger and kept without decreasing its activity within a year. The process can be scaled and, lysostaphine may be used in the treatment of man and animals after its proper biological tests. PMID- 10709463 TI - [Examining interaction of phages with microorganisms by fluorometry and electro orientation spectroscopy]. AB - Bacterial sensitivity to different various phages was examined by electro orientation spectroscopy, fluorometry, and electron microscopy. The strains of Pseudomonas aeruginosa, Staphylococcus aureus, Yersinia pestis, Mycobacterium smegmatis, and Xanthomonas campestris were used. The fluorescence intensity of a membranotropic agent in the ANS-cell-phage system was shown to depend on the interaction of a bacterial virus and a microorganism. Fluorometric data correlated with electro-orientation spectroscopic findings. An analysis of the low-frequency site makes it possible to determine phage adsorption on the bacterial surface. The changes in electro-orientation effects at high frequencies suggest that there are barrier dysfunctions in the external membranes and that there is cellular phage reproductions. Whether fluorometry and electro orientation spectroscopy can be further used for rapid identification of microorganisms by using phages is discussed. PMID- 10709464 TI - [pCSE4 plasmid for cloning of promoter-containing DNA fragments in francisella tularensis]. AB - Recombinant pCSE4 plasmid has been constructed, which contains the cat gene of the Tn9 without its own promoter and with the restriction BamH1 site in front of the Shine-Delgarno region of this gene. pCSE4 is consistently inherited in the cells of F. tularensis and E. coli and makes the cells of both microorganisms to be resistant to tetracycline. By cloning the Sau3A fragments of F. tularensis chromosomal DNA at the BamH1 site of pCSE4. Promoter-containing DNA fragments were selected. The plasmids with such chromosomal DNA fragments retained their structural integrity and functional activity in F. tularensis. PMID- 10709465 TI - [Temperature-dependent changes in immunochemical properties of lipopolysaccharides of yersinia pestis]. AB - The preparations of pilopolysaccharides (LPS) of virulent and avirulent Y. pestis strains, which have a different composition, were cultivated at different temperatures. Despite the cultivation temperature of parental cells, all LPS preparation inhibited the passive hemagglutination reaction (PHR) of the red blood cells sensitized by LPS isolated from the cultures grown at 26 degrees C by using homologous antisera. In contrast, the homologous system consisting of the red blood cells sensitized by LPS from the EV NIIEG, cultured at 37 degrees C, and the antiserum to these cells proved to be more specific and it was inhibited only by the homologous LPS isolated from the strain EV NIIEG. The similar reaction of the interaction of the red blood cells sensitized by high temperature LPS agents from plasmid-free strains with the same serum was inhibited by all plague LPS preparations. The LPS preparations from the strains Y. pestis 1146 and Y. pseudotuberculosis 9532 obtained when cultivated at 37 degrees C. RHR of the red blood cells sensitized by these preparations was inhibited by homologous LPS irrespective of the temperature of cell cultivation. In all cases, the reaction was specific for Yersinia strains and it was inhibited by LPS from Ra-Rd2 cultures, variants of Salmonella and E. coli. PMID- 10709466 TI - [Physicochemical and antimicrobial properties of chlorine-containing compositions ciaref and DP-1]. AB - The paper presents the results of the physicochemical properties and corrosive activity of the new chlorine-containing compositions Ciaref and DP-1. Various microbiological, biochemical, and biophysical studies were conducted to examine different aspects of the mechanism of action of Ciaref and DP-1 on vegetative (Francisella tularensis) and sporal (Bacillus turingiensis) microorganisms. Suspension, test-surface irrigation, and aerosol techniques were employed under close industrial conditions to study the disinfecting properties of the agents in bacterial and sporal contamination. PMID- 10709467 TI - [Response of immune system and lymphoid tissue of respiratory and gastrointestinal organs to space flight factors]. AB - The studies demonstrated that gamma-radiation drastically enhanced destructive processes and suppressed the mitotic activity of lymphocytes in the thymus and spleen. This resulted in the altered morphological picture of immune organs: the inversion of layers occurred in the thymus, the splenic white pulp increased by three times, lymphoid nodules with germinating centers disappeared, the marginal area became thinner. Following gamma-radiation, restorative processes in the thymus and spleen were noticeable just on day 3 and 7, respectively. However, the cell composition of murine immune organs failed to achieve control values by day 60 after exposure. Examining the responses of respiratory and digestive lymphoid tissue to acetaldehyde and drinking water organisms indicated that as the concentration of an acting agent and the time of exposure increased, there was lymphocytopoietic inhibition in the lymphoid formations whereas its small doses activated a local immune response. PMID- 10709468 TI - [Maternal blood fetal cells: new noninvasive approach in prenatal diagnosis of hereditary diseases]. AB - A new noninvasive approach to prenatal diagnosis of hereditary diseases is being actively developed, which is based on the use of different fetal cells contained in pregnant females. Due to the fact that the native concentration of fetal cells is extremely low, their isolation requires the application of different know-how enrichment and sorting techniques. Either the FISH method with chromosome specific probes or PCR is used to examine the cell fraction isolated, which detects fetal sex, Mendelian disorders such as beta-globin mutations. Promising results in the prenatal diagnosis of chromosomal aneuploidies were achieved in isolating and examining fetal erythroblasts. The results of numerous studies on the optimization of a protocol for isolating and reliably examining fetal cells from the blood of pregnant females allow the new noninvasive approach to the prenatal diagnosis of hereditary diseases to be considered to be highly promising. PMID- 10709469 TI - Use of a tapered, porous-surfaced dental implant in combination with osteotomes to restore edentulism in the difficult maxilla. AB - The maxilla is the more difficult arch to restore with endosseous dental implants because of hurdles such as low bone density, narrow buccopalatal width, minimal bone height, and proximity to the maxillary sinus. In this article, a technique to resolve all of these problems using a short, tapered, porous-surfaced implant and a placement protocol using hand osteotomes rather than surgical burs is presented. PMID- 10709470 TI - Fixed prosthodontics in skeletal Class III patients with partially edentulous jaws and age-related prognathism: the basal osseointegration procedure. AB - Today, prognathism in the partially or completely edentulous jaw can be treated with endosteal implants and fixed prostheses. The preferred procedure uses basal osseointegration. If the distribution of available bone is favorable, the prosthodontic suprastructures can be loaded early, taking the various phases of bone regeneration into account. Invasive surgical interventions, specifically iliac crest transplants, are rarely indicated and can be avoided in most cases. Patients are able to return to their everyday lives within a few days. PMID- 10709471 TI - Treatment of peri-implantitis: longitudinal clinical and microbiological findings -a case report. AB - Failing implants can be successfully treated by surgical procedures that use either bone fillers or membranes combined with an antimicrobial treatment. In this report, we present a case of failing implants with the corresponding treatment and results of 8 years of follow-up. PMID- 10709472 TI - Three-dimensional guidance system for implant insertion: Part II. Dual axes table -problem solving. AB - The three-dimensional guidance system for implant insertion is a technique for placing a radiopaque vertical orientation pin over the crest of the alveolar ridge on the stone cast during fabrication of the radiographic guide. The cross sectional and panoramic reformatted images were reproduced on a Polaroid or 35-mm print. The true vertical orientation pin facilitates (1) identification and the exact planned location of each implant in the reformatted images of the CT scan, (2) establishment of the internal starting point for the osteotomy on a photographic print, (3) optimum implant orientation, and (4) measurement of the angulation between the true vertical orientation line and optimum implant orientation. With the aid of a newly developed dual-axes base and transfer of the internal starting point of each implant to the stone cast, the buccolingual and mesiodistal implant inclinations for each implant were transferred to a surgical guide in the form of surgical steel drill guide tubes. The resulting pilot osteotomy transfers to the alveolar bone the exact starting point and the buccolingual and mesiodistal inclination for each implant. The technique provides a three-dimensional guidance system for implant insertion that is extremely accurate and yet practical. PMID- 10709473 TI - Why do dental implants fail? Part II. AB - Dental implant failure has led to continuous innovations of various implant systems and to different interceptive treatment modalities. These concerns have also led to selection of implant designs that best suit the various types of bone. A checklist has been created to facilitate collection of data on the different factors associated with dental implant failure. The data gathered from this list are the basis of a multinational statistical analysis. This analysis will provide accurate information about the percentage of each element causing implant failure. Different causes of failure, such as host factors, surgical placement, and improper implant selection, were reviewed in Part I of this two part series. This article discusses failure categories in terms of etiology, failure mode, failure type, failure origin, failure timing, responsible personnel, and different tissue types. PMID- 10709474 TI - Block autografts for localized ridge augmentation: Part I. The posterior maxilla. AB - This two-part series focuses on localized ridge augmentation using block autografts from the symphysis and the ramus buccal shelf for posterior maxillary (part I) and mandibular (part II) reconstruction. The various aspects of the surgical technique necessary for predictable results and minimal morbidity are discussed in detail. Emphasis is placed on staging, incision design, and recipient site preparation. PMID- 10709475 TI - Improvement of osseointegration of titanium dental implants by a modified sandblasting surface treatment: an in vivo interfacial biomechanics study. AB - To study the effects of a modified sandblasting surface treatment on the osseointegration of dental implants at the level of interfacial biomechanics, an in vivo pullout test was conducted using bone-interfacial shear strength as a criterion. Titanium implants were inserted into the medialis condyli of dogs and harvested 2, 4, and 12 weeks after insertion. Shear strength was determined with an Instron pullout tester. Observation and analysis of the surface of modified sandblasted implants after pullout at 12 weeks were performed with scanning electron microscopy and x-ray spectroscopy. Results showed that the shear strength of implants with a modified sandblasted surface was about five times as high as that of implants with a smooth surface. We concluded that the rough surface of titanium dental implants created by the modified sandblasting treatment can greatly enhance the shear strength at the dental implant-bone interface and that, with this enhancement, the secondary micropores play a much more important role in implant-bone bonding. PMID- 10709476 TI - Histomorphometric analyses of hydroxyapatite-coated and uncoated titanium dental implants in rabbit cortical bone. AB - In this article, we report the results of analyses of bone healing around four types of dental implants. Five implants of each type were inserted into the proximal tibia metaphysis of adult New Zealand rabbits and were analyzed using computerized histomorphometry 12 weeks after implantation. Hydroxyapatite-coated implants showed more direct bone contact and more lamellar bone in the threads than the titanium implants. There was a significant correlation between an increase in the percentage of mineralized tissue in the threads of metallic implants and cellular density around the implant, indicating less lamellar bone in contact with metallic implants. PMID- 10709477 TI - Evaluation of bone tissue on metallic implants by energy-dispersive x-ray analysis: an experimental study. AB - Osseointegration capacity of the different metallic implants depends on several variables. Osseointegration can be evaluated by using different methodologies, such as light microscopy and scanning or transmission electron microscopy. The aim of this study was to develop a qualitative and quantitative method to evaluate the presence of bone tissue on large metallic surfaces. A laminar implant was placed in each tibia of 10 Wistar rats. The animals were killed 30 days after surgery. Tibiae were resected, one for embedding in methyl methacrylate and the other for evaluation by energy-dispersive x-ray analysis. Light microscopy revealed osseointegration. Observation of the implant surface by scanning electron microscopy revealed the presence of a coating on the metallic surface that was rough in some areas and smooth in others. Analysis of the coating by energy-dispersive x-ray analysis showed the presence of Ca and P. Eighty percent (+/- 10%) of the metallic implant surface exhibited bone tissue. After confirmation of the occurrence of osseointegration capacity using light microscopy, the method described here allows qualitative and quantitative evaluation of the bone tissue found on large metallic surfaces. PMID- 10709478 TI - Evaluation of the effects of restorative materials used for occlusal surfaces of implant-supported prostheses on force distribution. AB - In this study, the functional stresses in alveolar bone created by restorative materials used in implant-supported prostheses were determined by the photoelastic stress analysis method. A photoelastic resin mandibular model, hollow-cylinder implants (3.5 mm diameter, 10 mm length), and fixtures and superstructures made of five different prosthetic materials were used. Vertical and 45-degree inclined loads were applied to the model. Subsequently, color photographs of the force line distribution observed on a polariscope were taken. Differences between vertical and inclined loads were statistically significant for all prosthetic materials (P < 0.01). The greatest stress distribution around the apex of the implant was observed with Ceramco II porcelain, followed by Biodent acrylic. The ArtGlass and Elcebond CCV composite materials had equal values, whereas Verebond Ni-Cr alloy showed minimum stress distribution. PMID- 10709479 TI - Antibiotics and anti-inflammatory agents in dental implantology. PMID- 10709480 TI - Dental implants in the diabetic patient: a retrospective study. AB - It has become increasingly common for controlled diabetic patients to be considered as candidates for dental implants. This study reports on the results of placing implants in 34 patients with diabetes who were treated with 227 Branemark implants. At the time of second-stage surgery, 214 of the implants had osseointegrated, a survival rate of 94.3%. Only one failure was identified among the 177 implants followed through final restoration, a clinical survival rate of 99.9%. Screening for diabetes and trying to ensure that implant candidates are in metabolic control are recommended to increase the chances of successful osseointegration. Antibiotic protection and avoidance of smoking should also be considered. PMID- 10709481 TI - Potential of recombinant human bone morphogenetic protein-2 in bone regeneration. AB - Recombinant human bone morphogenetic protein (rhBMP-2) has been used as a bone substitute. This article describes the rhBMP-2 structure, mechanisms of action, carriers, advantages, safety, and recent clinical studies relevant to dentistry. PMID- 10709482 TI - Titanium foil-guided tissue regeneration in the treatment of periimplant bone defects. AB - Resorbable membranes are often not stable enough for complete corrections of deep periimplant bony defects. Nonresorbable polytetrafluoroethylene membranes require removal after osseous healing. In this study the use of titanium foils for reconstruction of deficient alveolar ridge structures around dental implants is described. The advantages and disadvantages of the titanium foil-guided bone regeneration technique is discussed. Forty-two patients with deep intra alveolar periimplant defects were treated by means of a titanium foil-guided bone regeneration technique. Autologous bone in combination with a demineralized freezedried bone allotransplant was used for augmentation. Clinical and radiological control was performed 3, 6, and 12 months after surgery. In 37 cases, the average 12-month postoperative increase in bone was 4.2 mm, and the decrease in augmented bone was only 4% compared with the postoperative situation. Acceptable augmentative results were achieved in 88% of patients after the first operative treatment using a titanium foil-guided bone regeneration technique for reconstruction of periimplant defects and in all patients after the second augmentation. The main problem with foil loss was denudation and infection 6 weeks after surgery. PMID- 10709483 TI - Implant design considerations for the posterior regions of the mouth. AB - The posterior regions of the mouth sustain greater forces, yet often present poorer bone density. A biomechanical approach, often presented to decrease risk factors in such regions, is to increase implant surface area. Most manufacturers provide implants in various lengths. The longest implants are typically inserted into the anterior regions of the mouth, where forces of less magnitude and superior bone quality are present. A finite element analysis supports the hypothesis that implant length is a secondary parameter for stress distribution. A common approach is to enhance implant surface area in the posterior regions primarily by focusing on diameter. However, this increases surface area by only 30% for conventional thread designs despite the fact that forces increase by > 300% in the posterior regions. A change in implant diameter and thread design may increase surface area by > 300%. Such increases in surface area may decrease stresses to the crestal bone regions and reduce both crestal bone loss and early loading implant failure. PMID- 10709484 TI - Restoration of congenitally missing lateral incisors: a case report. AB - Parents and dentists are forced to make a decision early in a young patient's life when it is learned that the lateral incisors are missing. For many years the treatment has been to either move the cuspids into the lateral incisor sites or retain the teeth in their natural environment and restore the defect with a bonded or fixed bridge. With the advent of new designs in dental implants and their abutments, it is possible to consider replacing missing single teeth with implant-borne prostheses. Often-times, because of the limited residual bone and proximity of adjacent roots, placing conventional cylinder or screw-type implants is difficult. This article demonstrates the advantages of using a tapered-step implant, immediate one-stage surgery, and temporization in replacing congenitally missing laterals. PMID- 10709485 TI - Mandibular smooth staple implant: a case report. AB - This article reviews the development of staple bone plates for mandibular reconstruction over the past 25 years. A case study utilizing the Smooth Staple Implant System in a severely resorbed mandible is presented. Benefits of this system include reduction of the surgical operating and reconstruction time, elimination of the need to harvest a bone graft, the ability to create a functional prosthesis within 24 to 48 hours of the implant surgery, and increased cost-effectiveness. PMID- 10709486 TI - Nine-year follow-up of successful placement of endosseous implants in a mandibular bone graft. AB - Facial trauma injuries secondary to gunshot wounds present a unique challenge. These wounds are avulsive and typically involve the destruction of soft tissue with bone loss. A technique in bone transplantation is that of particulate cancellous bone and marrow. Initial form and stability can be provided by a titanium mesh tray or reconstruction plates while the graft undergoes maturation and consolidation. Dental implants can then be placed in this grafted site to provide stabilization for a functional and comfortable prosthesis and for the support of the peri-oral soft tissues. PMID- 10709487 TI - A simple prosthetic approach using cement-retained implant prosthesis after surgical treatment of ameloblastoma. AB - Ameloblastoma is an odontogenic tumor of epithelial origin that manifests itself in the maxillofacial area with marked deformity. After removal of the tumor, prosthetic reconstruction may be challenging because of extensive tissue loss, which necessitates careful treatment planning. We describe a case in which a patient lost considerable supporting tissue after excision of the tumor. The mandible was grafted with autogenous corticocancellous bone harvested from the left anterior iliac crest in a delayed manner. After healing, six implants were placed. After second-stage surgery, a new method was used to reduce thickness of the soft tissue around the implants. A cement-retained nine-unit fixed partial denture was fabricated in three clinical visits. The patient has been monitored for 5 years with no complications despite the use of cement to retain the prosthesis and the unfavorable crown-to-root ratio. PMID- 10709488 TI - Clinically based implant selection. AB - A hierarchy of implant selection is presented, based on overcoming specific clinical challenges in a variety of situations, including maximization of the esthetic, comfort, and functional potentials of therapy. PMID- 10709489 TI - In vitro susceptibility of Candida albicans to four disinfectants and their combinations. AB - AIM: The aim of this study was to evaluate the susceptibility of seven strains of Candida albicans to four disinfectants: iodine potassium iodide, chlorhexidine acetate, sodium hypochlorite and calcium hydroxide. In addition, all possible pairs of the disinfectants were tested in order to compare the effect of the combination and its components. METHODOLOGY: Filter paper discs were immersed in standardized yeast suspensions and then transferred to disinfectant solutions of different concentrations and incubated at 37 degrees C for 30 s, 5 min, 1 h and 24 h. After incubation the filter paper discs were transferred to vials with PBS and glass beads that were then vigorously shaken for dispersal of the yeast cells. PBS with resuspended yeasts was serially diluted 10-fold. Droplets of 25 microL from each dilution were inoculated on TSB agar plates and incubated in air at 37 degrees C for 24 h. The number of colony-forming units was then calculated from appropriate dilutions. RESULTS: C. albicans cells were highly resistant to calcium hydroxide. Sodium hypochlorite (5% and 0.5%) and iodine (2%) potassium iodide (4%) killed all yeast cells within 30 s, whilst chlorhexidine acetate (0.5%) showed complete killing after 5 min. Combinations of disinfectants were equally or less effective than the more effective component. All C. albicans strains tested showed similar susceptibility to the medicaments tested. CONCLUSIONS: This study indicates that sodium hypochlorite, iodine potassium iodide and chlorhexidine acetate are more effective than calcium hydroxide against C. albicans in vitro. However, combining calcium hydroxide with sodium hypochlorite or chlorhexidine may provide a wide-spectrum antimicrobial preparation with a long-lasting effect. PMID- 10709490 TI - The shear bond strength of glass ionomer cement sealers to bovine dentine conditioned with common endodontic irrigants. AB - AIM: The aim of this in vitro study was to evaluate the shear bond strength of commercially available and experimental glass ionomer cement (GIC) sealers to dentine exposed to common endodontic irrigants. METHODOLOGY: The enamel of 90 bovine incisor crowns, randomly divided into nine equal groups, was ground to expose the superficial dentine layer. The exposed surface was conditioned with either: (i) distilled H2O; (ii) 2.6% NaOCl; or (iii) 17% EDTA followed by 2.6% NaOCl. Five cc of each irrigant was applied over a 30-second period. The following GIC sealers were tested: (i) Ketac-Endo; (ii) KT-308, an experimental sealer; and (iii) ZUT, a combination of KT-308 and an antibacterial agent (0.2% by weight). The test sealers were applied to form cylinders with a standardized contact surface area (17.8 mm2) on the conditioned dentine surfaces. Specimens were bench set for 90 min and stored in distilled water at 37 degrees C for 48 h, then tested to failure for shear bond strength (MPa) in an Instron machine. RESULTS: KT-308 and ZUT had significantly higher MPa values than Ketac-Endo (two factor ANOVA, P < 0.0001), regardless of the dentine conditioning. All specimens conditioned with 17% EDTA and 2.6% NaOCl combined had significantly lower MPa values than those conditioned with water or 2.6% NaOCl alone (P < 0.02). CONCLUSIONS: It was concluded that the experimental sealers KT-308 and ZUT bonded better to bovine dentine than Ketac-Endo, and that the bond of all three GIC sealers was better with the smear layer present. PMID- 10709491 TI - Rotary Ni-Ti profile systems for preparing curved canals in resin blocks: influence of operator on instrument breakage. AB - AIM: The aim of this study was to determine the incidence of fracture of ProFile 0.4 and 0.6 taper Series 29 nickel-titanium instruments with respect to operator experience. METHODOLOGY: A total of 125 simulated root canals in resin blocks with the same geometrical shape in terms of angle and radius of curvature and coronal and apical orifice diameter were used. Five operators prepared all the specimens using an identical step-down instrument sequence, each one preparing 25 canals. The operators included two endodontists and three general practitioners. Statistical data concerning the incidence of instrument failure was compiled using Statlab and Fisher's partial least square difference analysis of variance. RESULTS: A total of 21 (16.8%) instruments fractured, all had 0.04 tapers. Nine size 25 instruments failed, 9 size 20 instruments failed and 3 size 15. During the study, the Binary Tree analysis of instrument failure revealed two operator populations belonging to two different study periods. The first period, which represented the first 13 root canal preparations, was called the 'learning period', and the second period, which represented the next 12 sample preparations, was called the 'application period'. A greater number of instruments failed during the first period than during the second. In the 'learning period', both groups of operators learned the same way. In the 'application period', two groups could be distinguished; the first group consisted of a general practitioner who produced worse results, and the second group consisted of the other four operators. CONCLUSIONS: The results indicate the necessity of mastering this rotary canal preparation technique, and the importance of improving competence through learning and experience. PMID- 10709492 TI - A comparison in vitro of fibroblast attachment to resected root-ends. AB - AIM: The aim of this study was to investigate the effect of surface morphology of root ends resected with various bur configurations on fibroblast attachment. METHODOLOGY: Seventy-two human single-rooted teeth were collected and decoronated. The root canals were instrumented, and then obturated with thermoplasticized gutta-percha using AH-26 as the sealer. The roots were randomly divided into eight different groups, and apical root-end resections were performed using eight different instruments, which included high and low speed burs and a scalpel blade. After each root was resected, the surface area of the root end was measured. Cultured human periodontal ligament fibroblasts were radiolabelled and then allowed to attach, in vitro, to the root-ends. Cell attachment to the resected root-ends was determined by counting in a liquid scintillation system and expressed as the number of decays per min/mm2 (DPM/mm2) of root surface. RESULTS: There was no significant difference in fibroblast attachment to the root-ends prepared with various instruments. CONCLUSIONS: These findings indicate that the choice of instrument for root-end resection has little influence on the initial attachment of fibroblasts, and thus may have little effect on healing following root-end resection. PMID- 10709494 TI - Effects of alternating and direct electrical current application on the odontoblastic layer in human teeth: an in vitro study. AB - AIM: The aim of this study was to investigate the influence of a low intensity alternating current on the odontoblasts and odontoblast layer and compare this with the effects of a direct current. METHODOLOGY: Teeth extracted for orthodontic reasons were immersed in physiological saline stabilized with thymol crystals. Within 1 h of extraction, an alternating or direct current was applied on the crown in the direction of the apex of the tooth for 120-360 s. The current doses were 12, 30, 60, 600, 1800, 3600, 7200, 24,000 and 144,000 microC. The teeth were fixed in Bouin or Baker fluids, the pulps removed, dehydrated and immersed in paraffin, then sectioned, stained with haematoxylin and eosin, and studied under a light microscope. RESULTS: Neither direct nor alternating current, similar to that applied in electrical caries diagnosis caused histological changes in the odontoblasts. CONCLUSIONS: There was no difference between direct and low intensity alternating current in the response of the odontoblast. PMID- 10709493 TI - A scanning electron microscopic evaluation of root surfaces and the gutta-percha interface following root-end resection in vitro. AB - AIM: The aim of this study was to evaluate the effects that various commonly used instruments have on root surface morphology, the obturating material, and the interface between the obturation and the root canal walls following root-end resection in vitro. METHODOLOGY: Sixty human single-rooted teeth with fully formed apices were collected and decoronated. The root canals were instrumented, and then obturated with thermoplasticized gutta-percha using AH-26 as the sealer. The roots were randomly divided into 12 different groups, and apical root-end resections were performed using eight different instrument configurations, and two different directions in which the bur moved across the root face in relation to its direction of rotation. Epoxy resin replicas of the resected root ends were examined using scanning electron microscopy. RESULTS: Each instrument produced a characteristic surface finish on the resected root end that mirrored its cutting profile. Irrespective of the design of the bur used, smearing and shredding of the gutta-percha across the root face occurred only when the handpiece was moved across the root face in reverse direction in relation to the direction of rotation of the bur. CONCLUSIONS: To ensure minimal disruption and distortion of the root filling and to prevent shredding of the gutta-percha interface, care should be taken to ensure that the final pass of the bur across the root canal is in the correct direction in relation to its direction of rotation. PMID- 10709495 TI - Adaptation and sealability of two contemporary obturation techniques in the absence of the dentinal smear layer. AB - AIM: The aim of this study was to evaluate the adaptation and short- and long term sealability of two different thermoplastic techniques--a core carrier technique, Thermafil; and a warm vertical continuous wave of compaction technique, System B. METHODOLOGY: Fifty-one mesial roots of mandibular molars with separate canals, patent canal orifices and curvature greater than 15 degrees were cleaned and shaped with Orifice Shapers and ProFile.04/.06 taper Ni-Ti rotary files using 5.25% NaOCl and 17% REDTA to a size 30.04 taper Profile at the apex to create a continuous tapered preparation. Canals were randomly obturated with Sealapex root canal sealer and either alpha-phase gutta-percha on a plastic Thermafil carrier, or nonstandardized beta-phase gutta-percha using the System B heat source. Proximal radiographs of roots were evaluated by three examiners based on established criteria for overall material adaptation, apical adaptation and filling material extrusion. Teeth were randomly separated into three groups of 17 each and placed in black India ink for 10 days, 24 h, or after 67 days storage in Hank's Balanced Salt Solution. All roots were demineralized and rendered transparent. Three examiners evaluated the apical leakage by the linear measurement of dye penetration under the stereo-microscope. The movement of filling material into canal irregularities was also evaluated. RESULTS: Both obturation techniques were not significantly different in the overall canal obturation and in the apical third adaptation (P > 0.05). Significantly more filling material extrusion beyond the apex was noted with the Thermafil technique (P < 0.001). No significant difference was found amongst the 67-day, 10-day and 24 h System B groups (P > 0.05). The 67-day Thermafil group showed significantly more leakage than the 10-day and 24 h Thermafil groups. There was a significant difference in the degree of leakage between the 67-day Thermafil group and the 67 day System B group (P < 0.05), but not between the 10-day and 24 h groups (P > 0.05). Both obturation techniques produced substantial filling material movement into canal irregularities. CONCLUSIONS: It was concluded that Thermafil demonstrated more filling material extrusion beyond the apex and significantly more long-term apical leakage. PMID- 10709496 TI - Newly developed resinous direct pulp capping agent containing calcium hydroxide (MTYA1-Ca). AB - AIM: The aim of this study was to evaluate a newly developed resin (MTYA1-Ca) for direct pulp capping. METHODOLOGY: The powder of MTY1-Ca is composed of 89.0% microfiller, 10.0% calcium hydroxide and 1.0% benzoyl peroxide and was mixed with liquid (67.5% triethyleneglycol dimethacrylate, 30.0% glyceryl methacrylate, 1.0% o-methacryloyl tyrosine amide, 1.0% dimethylaminoethylmethacrylate, and 0.5% camphorquinone). The shear bond, diametral tensile, bending and compressive strengths were measured. The alkaline activity of the elute dissolved from MTYA1 Ca was calculated. Cell viability by MTT assay and alkaline phosphatase (ALPase) activity were evaluated from dental pulp fibroblast reaction to the eluate dissolved from MTYA1-Ca. Histopathological studies of the response to exposed dental pulp of beagle dogs were completed with Dycal as a control. RESULTS: The physical properties of MTYA1-Ca were significantly superior to those of Dycal. It was impossible to measure these properties with Dycal because of poor physical properties. Both MTYA1-Ca and Dycal maintained high levels of alkaline activity (pH 10.96-12.20) over the 168-h duration of the study. Cell viability by MTT assay in the intact eluate of MTYA1-Ca was significantly higher than that of Dycal, whilst ALPase showed no difference between MTYA1-Ca and Dycal. A dentine bridge formed more slowly under MTYA1-Ca than under Dycal, but similar amounts had formed at 90 days. CONCLUSIONS: MTYA1-Ca has the potential to be used as a direct pulp capping material. PMID- 10709497 TI - Scanning electron microscope study on the efficacy of root canal wall debridement of hand versus Lightspeed instrumentation. AB - AIM: The aim of this in vitro study was to compare the efficacy of root canal wall debridement following hand versus LightSpeed instrumentation. METHODOLOGY: Twenty recently extracted single-rooted teeth were paired and randomly placed into two treatment groups of 10 teeth each. In group 1, a step-back instrumentation without initial coronal flaring with stainless steel Hedstroem files was used; group 2 was instrumented with Ni-Ti LightSpeed instruments. Both groups had the same irrigation regimen: 2.5% NaOCl and a 15% EDTA solution. The teeth were then decoronated and each root split longitudinally into two halves to be examined using the scanning electron microscope (SEM). The presence of superficial debris and smear layer was evaluated by a standardized grading system, and the resulting scores submitted to nonparametric statistics. RESULTS: Under the conditions of this study, the removal of superficial debris was generally excellent with both canal preparation techniques. Both techniques resulted in variable presence of residual smear layer, with a canal wall covered by smear layer as the predominant characteristic. Generally, the amount of smear layer was greater in the apical than in the middle third of the root, however, this difference was statistically significant (P < 0.005) only in hand instrumented teeth. The use of LightSpeed instruments was associated with significantly more (P < 0.05) smear layer presence in the middle region of the root when compared with hand instrumentation. In addition, less smear layer was present in the apical region following LightSpeed instrumentation than stainless steel hand files, but this difference was not statistically significant. Differences in debridement between the two halves of the same root were more evident with LightSpeed than manual instrumentation, however, there was no statistical significance. CONCLUSIONS: It may be inferred that the choice between hand and LightSpeed instrumentation should be based on factors other than the amount of root canal debridement, which does not vary significantly according to the instruments used. PMID- 10709498 TI - Unusual case of bilateral talon cusp associated with dens invaginatus. AB - CASE REPORT: This paper presents a rare case of bilateral talon cusp in permanent maxillary central incisors, one on the labial and another on the lingual surface of each tooth. The condition was associated with dens invaginatus in a maxillary permanent lateral incisor, but no developmental syndrome was identified. The left central incisor required no treatment because the aesthetic appearance was satisfactory and neither occlusal interferences nor caries were present. After careful investigation of the right lateral incisor, a sealant was applied. Root canal treatment was indicated for the right central incisor that had a wide open apex with thin, weak, divergent walls, and an apical radiolucency. PMID- 10709499 TI - Distraction osteogenesis to achieve mandibular vertical bone regeneration: a case report. AB - In this case report a surgical technique for vertical ridge augmentation is presented. The procedure, performed in a 30-year-old woman with an atrophied alveolar ridge in the anterior portion of the mandible, is based on the biologic concept of osteogenesis distraction previously introduced in orthopedic and maxillofacial surgery. After elevation of a full-thickness flap a horizontal osteotomy was performed 7 to 8 mm from the top of the ridge. Two vertical osteotomies were prepared with drills of increasing diameter (2, 2.8, and 3.25 mm), tapping was performed for the first 5 to 6 mm, and two distractor base plugs were placed at the base of the osteotomies with a repositioning tool. An intraosseous distraction implant was then inserted and 2 inward vertical cuts were made in the bone to allow proper distraction to take place. Correct functioning of the device was checked by distracting the bone fragment 1 mm using the axial distraction screw. A latency distraction healing screw was inserted in each of the distraction implants and the area was left to heal for 5 days. Once primary healing had occurred, the distraction of the newly formed bone callus was activated each day for 10 days (1 mm per day). At the end of the distraction period a final distraction screw was left in place and a final healing screw was inserted. During this time there were no complications and the patient on no occasion complained of discomfort. The distractor device was removed 30 days later, leaving the base plugs in place. One month later a vertical augmentation of 7 mm had been achieved; the base plugs were removed, 3 intraosseous implants were inserted, and a biopsy of the newly formed tissue was obtained. Histologic evaluation of the biopsy specimen showed woven bone formation approximately 75 days after the initial procedure. PMID- 10709500 TI - Surgical crown-lengthening procedure to enhance esthetics. AB - This article presents the rationale for the use of a preprosthetic surgical crown lengthening procedure, particularly in the anterior region where esthetics is of great concern. Clinical cases illustrate the procedure and demonstrate how it can be used to provide enough sound tooth structure to restore teeth without impingement on the biologic width, at the same time reducing a "gingival smile" and creating new papillae. PMID- 10709501 TI - Immediate loading of modular transitional implants: a histologic and histomorphometric study in dogs. AB - Modular Transitional Implants (MTI) are made from pure titanium and are used to support fixed provisional restorations during the osseointegration of definitive implants. This study histologically examined the jaw response to loaded MTIs in the dog mandible. Three implants were inserted transmucosally into each side of the mandible in 3 dogs. Stability was examined using a Periotest. Anterior and posterior implants were splinted using a cemented acrylic resin fixed partial denture to allow immediate loading. The middle implant remained unloaded and was used as a control. Dogs were sacrificed 11 to 12 weeks after implantation, and tissue blocks containing the implants were removed. Histologic examination showed that 10 of the 18 implants had good bone-to-implant contact, with the percentage of bone contacting the threaded portion of the implant varying from 30% to 65%. There was no statistical difference (p > 0.1) in percentage of bone-to-metal contact between loaded and unloaded implants. Six implants were entirely surrounded by connective tissue with or without inflammation; two implants were lost during the study. The success rate did not differ between loaded and unloaded implants. In the successful implants trabecular bone made good contact with the implant, forming supporting struts. There was bone remodeling in some bone-to-metal contact areas. It is believed that success was mainly influenced by the initial bone density at the implant site and by the uncontrolled load that the animals applied to the implants during the early healing stage. PMID- 10709502 TI - Improved efficacy of calculus removal in furcations using ultrasonic diamond coated inserts. AB - Dentsply Cavitron Diamond Inserts provide improved efficacy in removing calculus from furcations. A total of 60 extracted human mandibular molar teeth had artificial calculus applied to the furcations, then were randomly treated with either sharp universal Gracey curettes (HAND), a plain ultrasonic TFI-10 tip in a cavitron instrument (CAV), or one of 2 diamond-coated cavitron instruments (TFI 10 fine-grit (FIN) and TFI-10 medium-grit (MED)) When the time needed to completely clean the furcations was evaluated, MED was the fastest, followed by FIN, CAV, and HAND, respectively. All of the powered instruments were faster than hand curettes with regard to effective in vitro calculus removal in furcations. The use of these types of instruments would reduce the time required to perform periodontal surgery and might improve regenerative therapy. PMID- 10709503 TI - Periodontal regeneration of intrabony defects using resorbable membranes: determinants of the healing response. An observational clinical study. AB - The safety and bone-regenerative capacity of a resorbable membrane (Resolut) was evaluated by a nonrandomized prospective clinical study of patients with periodontal defects. Prior to surgical management all patients underwent scaling and root planing and were instructed on oral hygiene. The study included 18 patients (31 periodontal defects) who received surgical treatment by guided tissue regeneration (GTR) using resorbable membranes. The results were evaluated 12 months after surgery in terms of Plaque Index, bleeding index, probing depth, gingival recession, clinical attachment level, and dental mobility. The results obtained show that the use of resorbable membranes in GTR causes few complications. The mean gain in clinical attachment level (4.06 +/- 1.91 mm), with an attachment level gain of more than 3 mm in 81.2% of the defects, suggests the presence of "new attachment." This difference was clinically and statistically significant (t = 11.03, P = 0). The multivariate regression study showed that 60% of the observed variability (F = 11.48, P < 0.001) in clinical attachment level gain was accounted for by the variable's initial probing depth, the Plaque Index of the tooth subjected to GTR, and the infrabony component of the defect. PMID- 10709504 TI - Skin-prick test for severe marginal periodontitis. AB - The present study tested the hypothesis that treatment-resistant periodontitis patients present with a more intense inflammatory response to marginal bacterial plaque as a sign of an inflammatory overreaction. Patients with severe marginal periodontitis (Gingival Index > 20%) who had not responded to treatment showed almost no positive response to lipid A in a skin-prick test, which was significantly different from the results from patients with severe marginal periodontitis who had responded to treatment and from healthy control individuals without marginal periodontitis. This finding can be interpreted as an impaired inflammatory reactivity to periodontitis pathogens in treatment-resistant patients, rejecting the hypothesis. PMID- 10709505 TI - Preliminary study of the forces developed by practitioners during amalgam condensation. AB - The aim of this preliminary study was to analyze, in vitro, the forces developed by practitioners during amalgam condensation. Standardized Class II cavities were drilled into 40 freshly extracted teeth. The forces exerted during condensation were then measured by means of a new device, the Endographe, and plotted online or offline as a function of time on Endogrammes. The work techniques of the different practitioners revealed similarities. The mean values of the vertical forces (+/- SEM) were: 15 +/- 2 N for manual compaction of capsule amalgam; 8 +/- 3 N for manual compaction of amalgam with a higher mercury-to-alloy ratio; 6 +/- 4 N for mechanical compaction of encapsulated amalgam; and 4 +/- 2 N for mechanical compaction of the modified amalgam. For the horizontal components, the forces were, respectively, 5 +/- 2 N, 0.1 +/- 0.05 N, 3 +/- 1 N, and 0.7 +/- 0.2 N. The mean values of condensation time were 20 +/- 15 s for each incremental application, with a great variation in durations between the beginning and end of condensation. By visually depicting forces as a function of time, the Endographe can be used to analyze the process of amalgam insertion and condensation. Future research will study the relationship between the forces developed during condensation and the adaptation of the amalgam to the walls of the cavity. PMID- 10709506 TI - Split palatal flap. II. A surgical approach for maxillary implant uncovering in cases with reduced keratinized tissue: technique and clinical results. AB - It is generally accepted that a more ideal and functional soft tissue-implant interface can be established if there is an adequate zone of keratinized mucosa around endosseous dental implants. The purpose of this article was to describe a surgical procedure, based on the use of a split palatal flap, which predictably creates or increases the zone of keratinized tissue around implants at the time of implant uncovering. It is especially useful for maxillary implants with a nonexistent or minimal width of keratinized buccal tissue. The study comprised 34 implants in the maxillae of 8 patients, who were chosen because they had minimal or nonexistent buccal keratinized gingiva prior to implant uncovering. Following healing, between 2 and 5 mm of keratinized gingiva (mean 3.7 mm) could be measured buccally at all abutments. Postsurgical inconveniences were minimal. The use of a split palatal flap at implant uncovering minimizes the number of surgical stages and sites necessary, while predictably providing an adequate zone of buccal keratinized gingiva. PMID- 10709507 TI - Reconstruction of the lost interproximal papilla--presentation of surgical and nonsurgical approaches. AB - Modern esthetic dentistry involves not only the restoration of lost teeth and their associated hard tissues, but increasingly the management and reconstruction of the encasing gingiva with adequate surgical techniques. The loss of interproximal dental papillae may cause functional, phonetic, and devastating esthetic problems. Complete and predictable restoration of lost interdental papillae remains one of the biggest challenges in periodontal reconstructive surgery. On reviewing the literature, publications involving surgical and nonsurgical techniques for papilla reconstruction are basically case presentations. Very little scientific data concerning long-term success and predictability of specific techniques has been published so far. Starting with facts about the anatomy and morphology of the interdental tissues, this article gives an overview of surgical and nonsurgical techniques to restore lost interproximal dental papillae. PMID- 10709508 TI - Histologic evaluation of guided vertical ridge augmentation around implants in humans. AB - Recent experimental and clinical case reports demonstrated vertical ridge regeneration in atrophic posterior mandibles and maxillae. Although the results from these clinical cases are quite encouraging there is a lack of human histologic data on the newly regenerated tissue around commercially available titanium implants. The aim of the present study was to perform a qualitative and quantitative histologic analysis of the bone response to previously exposed implant threads after treatment with guided bone regeneration in a series of patients. A total of 30 Nobel Biocare implants were consecutively placed in 6 patients with partially edentulous mandibles. Of these implants, 6 were planned for removal after 1 year, whereas the remaining 24 implants were inserted to function as support for a fixed partial denture. The 6 experimental implants were intentionally allowed to protrude occlusally 5 to 7 mm from the bone crest without counter-sinking. The exposed implant threads were completely covered by autogenous bone chips. After a 12-month healing period the 6 experimental implants were removed with trephine burs. Bone-to-metal contact and bone density in the implant threads were measured. Clinically, all implants were stable and there was complete tissue fill of the space underneath the membranes. Histologically, a substantial amount of new bone had formed underneath the membrane in all cases. Histomorphometrically, there was a lower bone-to-metal contact percentage in the exposed compared to the nonexposed region in every case. With respect to bone density, there was a mean of 43.2% in previously exposed regions compared to 60.3% in previously nonexposed regions. PMID- 10709509 TI - Successful root coverage: a human histologic evaluation of a case. AB - Connective tissue grafts combined with pedicle grafts (subepithelial grafts) have been shown to be effective in obtaining root coverage. Unfortunately, little is known about the histology of the results in humans. This is a case report of a tooth with a recession defect that was treated with a subepithelial graft. Complete root coverage was obtained. However, at 5 months postoperative the tooth had to be extracted because of a vertical root fracture. With the patient's permission, a small collar of tissue was removed with the tooth. The sample was processed and evaluated histologically. The results revealed areas of regeneration, with new bone, cementum, and connective tissue attachment coronal to the original gingival margin. No bone grafts or guided tissue regeneration membranes were used. This case report confirms that regeneration is possible with subepithelial grafts. PMID- 10709510 TI - Root coverage and papilla reconstruction in Class IV recession: a case report. AB - While root coverage is predictable for Miller Class I and II recessions, the surgical regeneration or reconstruction of a lost interdental papilla is more difficult to obtain. To date only a few successful case reports have been reported, and there are no studies that report a predictable technique to obtain papilla reconstruction or root coverage on Class IV recessions. This case report, which is part of a preliminary study, outlines a surgical technique to obtain simultaneous root coverage and papilla reconstruction. PMID- 10709511 TI - Immediate implant placement and GBR in humans: a case report and histologic evaluation. AB - An IMZ titanium plasma-sprayed implant was placed at the time of removal of a fractured mandibular left first premolar. Porous hydroxyapatite (Interpore 200) was placed on one side of the residual defect around the implant, and the entire defect was covered with a Gore-Tex membrane. The implant, with surrounding newly reformed hard tissues, was removed in a block section 13 months postoperative. Histologic examination demonstrated regeneration of living bone tissues, the attainment of osseointegration, and incorporation of the Interpore 200 into surrounding bone. PMID- 10709512 TI - Efficacy of Carisolv-assisted caries excavation. AB - As a possible alternative to conventional techniques for excavating caries chemomechanical methods have been developed. Caridex has so far been the dominating product. However, a new system, Carisolv, was recently introduced. The purpose of the present study was to evaluate the caries-dissolving efficacy of Carisolv in vitro. After excavation with Carisolv all dentin surfaces were caries free. However, 6 of the 10 cavities showed residual caries in the dentinoenamel junction. The dentin and enamel surfaces were covered by smear and debris. Since there may be a risk of leaving caries in the dentinoenamel junction proper case selection appears to be of importance to ensure a successful result. PMID- 10709513 TI - The bilateral pedicle flap-tunnel technique: a new approach to cover connective tissue grafts. AB - A new surgical technique for the treatment of adjacent soft tissue marginal recession is presented. This technique combines the use of a tunnel procedure with double lateral pedicle flaps to cover a connective tissue graft. This approach combines the advantages of the tunnel technique with the increased blood supply and protection provided by pedicle flaps. Indications include adjacent Class I and II deep, wide recessions; however, the procedure may also be applied to mild Class III recessions. Two case reports are presented to illustrate this new technique. PMID- 10709514 TI - Combined treatment of a large periodontal defect using GTR and DFDBA. AB - The regeneration of periodontal structures lost to inflammatory disease is an elusive yet attainable goal of periodontal therapy. This article reports the successful treatment of a large periodontal defect using a combination of demineralized freeze-dried bone allograft (DFDBA) and guided tissue regeneration (GTR). The case presents endodontic and mucogingival complications in the combined GTR osseous graft technique. The combined techniques used in this 27 year-old patient achieved a reduction in probing depth, radiographic evidence of bone fill, and a reduction in clinical mobility. PMID- 10709515 TI - Histologic observations of periodontal wound healing after treatment with PerioGlas in nonhuman primates. AB - The effect of PerioGlas (synthetic bone particulate) on wound healing of experimental palatal periodontal defects in monkeys was evaluated. Chronic periodontal defects were created on the palatal aspects of maxillary molars and premolars. Open-flap debridement was performed. Experimental sites received PerioGlas, while control sites received no further treatment. Histologic measurements were performed on new bone, new cementum, epithelial downgrowth, and recession. Results showed significantly more new cementum and less epithelial downgrowth in the sites that received PerioGlas (P < 0.05). The present results indicate that PerioGlas may enhance periodontal wound healing outcomes. PMID- 10709516 TI - GTR with bioresorbable membranes in the treatment of intrabony defects: a clinical and histologic study. AB - The aim of the present study was to evaluate clinically and histologically the treatment of intrabony periodontal defects with a bioresorbable membrane barrier. Fifty-two intrabony periodontal defects were treated according to the principles of guided tissue regeneration (GTR) with a bioresorbable membrane. Results were evaluated by assessing probing pocket depth, recession of the gingival margin, and clinical attachment level at baseline and at 1 and 2 years after therapy. Bone level changes were evaluated radiographically. The postoperative phase was uneventful in all cases. There was a mean probing pocket depth reduction from 8.4 to 3.6 mm, a mean increase of gingival margin recession from 1.5 to 3.0 mm, and a mean clinical attachment level change from 9.9 to 6.5 mm. Mean attachment gain was 3.4 mm. Two teeth scheduled for extraction were also treated with the same bioresorbable membrane. The histologic analysis 6 months after treatment revealed the formation of new connective tissue attachment and new alveolar bone in both cases. Based on the histologic findings it can be concluded that the clinical improvements following GTR with this type of bioresorbable membrane may represent, at least in part, true periodontal regeneration. PMID- 10709517 TI - Implant-supported removable overdentures in the edentulous maxilla: clinical and technical aspects. AB - PURPOSE: The aim of this article is to describe the indication criteria and the treatment planning for a maxillary implant-supported removable overdenture. Prostheses are designed according to the requirements of the bar system and the factors influencing the extension of the prosthesis base. MATERIALS AND METHODS: The decisive factors in determining whether a bar-retained overdenture prosthesis is indicated should be evaluated during the initial clinical examination and with the help of a reformatted computed tomographic (CT) scan that is performed with a radiologic template in place. Titanium markers represent the ideal location of the denture teeth in the diagnostic setup so that the implant position can be selected and the available space for the bar system can be assessed vertically and horizontally. RESULTS: For the overdenture prosthesis that is solely implant supported 6 to 8 implants are placed ideally at a distance of about 10 to 14 mm from center to center. A prefabricated bar system that allows the clips to be inserted between the implants can then be used. When the available bone restricts implant placement to adjacent tooth positions an individually milled bar that includes additional frictional pins and/or retentive elements needs to be planned. The prosthesis design, in particular its buccal and palatal flange extension, is determined during setup try-in, taking into account the patient's smile line, their need for facial support, and their phonetic requirements. CONCLUSION: The removable implant-supported overdenture offers flexibility in placing implants in either adjacent tooth positions or with greater distances between them depending on the available bone, as either conventional bar and clip systems or individually milled bars can be used. Adjustment of the buccal prosthesis flange and the palatal prosthesis base is made to fulfill the patient's requirements concerning esthetics, phonetics, comfort, and function. PMID- 10709518 TI - Prosthetic restoration following maxillary resection without an oroantral defect: a case report. AB - PURPOSE: The aim of this article was to present the oral rehabilitation of a patient with a large, created intraoral defect after the surgical extirpation of a pathologic process. MATERIALS AND METHODS: On osseointegrated oral implants, a bar was fabricated to support a partial overdenture by means of attachments. RESULTS: The treatment was successful and the construction has been in service for more than 5 years. CONCLUSION: In cases with large intraoral defects removable dentures may have clinical advantages over fixed partial dentures. In this patient the restoration of missing teeth, alveolar crest, and soft tissue was accomplished at the same time oral function was restored, and good oral hygiene access was obtained. PMID- 10709519 TI - Measurement of the margins of partial-coverage tooth preparations for CAD/CAM. AB - PURPOSE: This study tested the hypothesis that a scanning laser 3-dimensional digitizer is a precise and accurate instrument to measure chamfered and beveled margins of partial coverage tooth preparations for computer-aided design/computer aided manufacturing (CAD/CAM). MATERIALS AND METHODS: The margins were measured by the digitizer on stone dies and calculated by triangulation into a 3-D representation. Instrument precision was defined as the ability to reproduce the same margin in repeated measurements and expressed as the coefficient of variation as a percentage. Instrument accuracy for chamfered and beveled margins was estimated by correlating their measurements to the measurement of the margin of a spherical calibration "phantom" with known dimensions. Accuracy was expressed as the standard deviation. RESULTS: The precision errors for the box- and cusp-chamfered margins and cusp-beveled margins were 3.9%, 3.4%, and 2.4%, respectively. With regard to accuracy the standard deviations of the measurements of the box- and cusp-chamfered margins and cusp-beveled margins were 19 microns, 21 microns, and 24 microns, respectively, compared to 15 microns for the phantom. CONCLUSION: Measurements of chamfered and beveled margins by a scanning laser 3-D digitizer for CAD/CAM are (1) precise (error < 4%) and (2) accurate, with a standard deviation of less than 9 microns compared to optimal measurements of the spherical margin of the phantom. PMID- 10709520 TI - A difference in perspective--the North American and European interpretations of tooth wear. AB - PURPOSE: There is considerable interest in the European dental research literature about the problem of tooth wear and specifically about dental erosion, but this interest does not appear to be matched in North America based on the volume of the literature there. The purpose of this article is to consider the possible explanations for this difference. MATERIALS AND METHODS: This article examines the reasons for this disparity and attempts to explain the difference by reviewing the North American and European literature on the etiology, pathogenesis, and prevalence of tooth wear. RESULTS: It would appear from the literature that the reason for the difference in interest between the 2 continents is a reflection of how the appearance, etiology, and terminology are interpreted and used to define tooth wear, attrition, and erosion. CONCLUSION: Attrition is the wear of teeth against teeth; therefore, by definition any worn surface that does not contact the opposing tooth must have another etiology. An appropriate descriptive term is "tooth wear" when the etiology is multifactorial or cannot be determined. A search of the literature shows more studies in the European literature of the etiology and prevalence of tooth wear than in the North American literature. The thrust of the European studies supports the view that erosion is more important than attrition in the etiology of tooth wear. PMID- 10709521 TI - Wear behavior of precision attachments. AB - PURPOSE: The purpose of this article was to compare the wear behavior of precision attachments with plastic inserts to conventional metal-alloy precision attachments. MATERIALS AND METHODS: In a comparative study attachments of various designs were subjected to alternating load cycles in a wear simulator. In addition to conventional adjustable attachments with metal-alloy matrix and patrix elements, attachments with female elements that are lined with plastic inserts were investigated for the first time. In each wear test 10,000 separating and joining movements were performed in an axial direction under a continuous spray of artificial saliva at 37 degrees C. RESULTS: The attachments with metal surfaces showed a rapid loss of approximately 60% of the required separating/joining forces during the first 1,000 cycles; after a further 9,000 cycles these forces fell to 25% and 35%, respectively, of the initial value. The attachments with plastic inserts, by contrast, showed only a 4% and 8% loss, respectively, of the required separating/joining forces even after 10,000 wear cycles. With one attachment type a reproducible 20% increase of retention occurred during the testings. CONCLUSION: The precision attachments with plastic female inserts showed only negligible amounts of wear and the most consistent retentive force in comparison with conventional precision attachments consisting of metal-alloy matrix and patrix components. PMID- 10709522 TI - Surface energy of etched ceramic. AB - PURPOSE: In a previous article the authors examined the evolution of the bond strengths of 2 dental feldspathic ceramics. The objective of the present study was to evaluate the effect of surface modifications with hydrofluoric acid gel (concentration 10%) on the surface energy of 2 dental feldspathic ceramics (GC and PVS). MATERIALS AND METHODS: For an energy characterization, 30 samples of GC and 30 samples of PVS were built. This study comprised the measurement of contact angles to determine the work of adhesion (WA) of the 2 ceramics. The evolution of the work of adhesion depended on the action of the hydrofluoric acid gel on the roughness of the surface of the 2 ceramics. RESULTS: In a polished state PVS presented a higher work of adhesion than GC. Etching the ceramics with hydrofluoric acid gel increased the work of adhesion, especially for GC, but this treatment was not sufficient to obtain a high work of adhesion. CONCLUSION: Etching with hydrofluoric acid gel was not sufficient to raise the work of adhesion of the 2 ceramics. Silanization is preferable to etching. PMID- 10709523 TI - Integrated electromyography of the masseter on incremental opening and closing with audio biofeedback: a study on mandibular posture. AB - PURPOSE: The purpose of this study was to test the hypothesis of a minimum electromyographic (EMG) rest position based on masseter surface EMG recordings of incremental opening and closing of the mandible with simultaneous audio EMG biofeedback. MATERIALS AND METHODS: Nineteen alert subjects in an upright seated position opened and closed the mandible in 1-mm increments 20 mm interincisally. An electronic recording device allowed each subject to maintain the vertical dimension of each increment while simultaneously reducing right masseteric muscle activity to the minimum possible level using audio EMG biofeedback. Integrated EMG masseteric activity was recorded at each static opening and closing increment. RESULTS: A mean plateau of integrated EMG output for all subjects with no minimum EMG point or circumscribed minimum EMG range for any of the nineteen subjects was shown. Analysis of variance for repeated measures showed no difference in opening and closing EMG levels (P = 0.27) and no interaction between opening, closing, and change in vertical dimension (P < 0.0001). CONCLUSION: These results, with those of other studies, raise questions regarding the validity of the concept of a unique physiologic rest position of the mandible with the masseter or associated muscles at minimum muscle activity. The idea of overlapping postural ranges appears to be more appropriate. PMID- 10709524 TI - Prosthodontic decision making among general dentists in Sweden. I: The choice between crown therapy and filling. AB - PURPOSE: The purpose of this study was to analyze dentists' evaluations of factors related to the choice between crown therapy and filling and to possibly explain this by social and demographic attributes, job situation, and dentists' attitudes. MATERIALS AND METHODS: Questionnaires were sent to a random sample of 2,059 Swedish dentists. The response rate was 76%. In the questionnaire the choice between crown therapy and filling in a clinical situation was presented. The dentists were asked to mark their assessments of the relative importance of the different items on 14-item visual analogue scales (VAS). Multiple regressions were run for all 14 items. RESULTS: Large individual variations were seen among the dentists regarding the stated importance of the various items. The items rated as the most important were "patient's wish" and "treatment prognosis," and the items rated as least important were "treatment time required" and "number of visits required for treatment." The differences between groups were small, and for no item exceeded 0.7 step on the 8-grade VAS scale. No data reduction was possible using principal components analysis. CONCLUSION: The study showed great variations among individual dentists. The dentist-related factors explained little of the variance. The results indicated that the questionnaire instrument did not fully capture the real influences on the treatment choice between a filling and an artificial crown. PMID- 10709525 TI - Two-body wear resistance and degree of conversion of laboratory-processed composite materials. AB - PURPOSE: The purpose of this investigation was to evaluate the relative 2-body abrasive wear and degree of conversion of 4 laboratory-processed composites (Targis, Concept, belleGlass, and Artglass) and 2 direct placement composites (Herculite and Heliomolar) after 7 days of storage. MATERIALS AND METHODS: Human enamel was used as a positive control for 2-body abrasive wear, and 10 cylindric specimens (3.5-mm diameter, 8-mm height) of each material were prepared and stored in distilled water at 37 +/- 2 degrees C for wear testing. Relative 2-body abrasive wear rates were determined using a 30-micron diamond disk and a 2-body pin-on-disk apparatus. Subsequently, 3 polymerized specimens that had been stored in sealed polyethylene vials for 7 days were prepared for degree of conversion testing. The degree of conversion was determined on an infrared spectrometer using standard baseline techniques and various internal standards. RESULTS: Statistical analysis using analysis of variance and the Tukey-Kramer multiple range test indicated significant differences between several of the materials tested for both 2-body abrasive wear and degree of conversion. CONCLUSION: Concept exhibited significantly less 2-body abrasive wear compared to the direct and indirect composites (P < 0.01). Concept and belleGlass exhibited a mean degree of conversion that was significantly higher than any of the other composites tested (P < 0.01). PMID- 10709526 TI - The effects of tooth preparation on pressure measured in the pulp chamber: a laboratory study. AB - PURPOSE: The purpose of this study was to determine pressure changes in the pulp space during tooth preparation with either diamond or tungsten carbide burs in a high-speed dental handpiece in the laboratory. MATERIALS AND METHODS: Twenty premolar teeth were collected and randomly assigned to two groups: group 1 received preparation with diamond burs and group 2 with tungsten carbide burs. The teeth were mounted on a pressure transducer and the pulp chambers were filled with saline. A 0.1-mm thickness of tooth tissue was removed from the proximal surface of each tooth, alternating dry and wet cutting, until the pulp chamber was exposed. Pressure and temperature changes were recorded during tooth preparation. RESULTS: At 0 to 1 mm of remaining dentin depth dry cutting with diamond and tungsten carbide burs generated a mean positive pulpal pressure of 12 kPa and 6 kPa, respectively. Wet cutting under the same conditions produced 0.6 kPa and 0.15 kPa, respectively. The difference between wet and dry cutting was highly significant (P < 0.001). Diamond burs produced significantly higher pressure increases than carbide burs at all levels for both wet and dry techniques (P < 0.05). When cutting farther than 2 mm from the pulp, tooth preparation created a mean 0.09-kPa pressure increase, which was not influenced by either coolant use or bur type. The temperature change was minimal during wet cutting and only minor temperature increases were recorded during dry cutting. CONCLUSION: From this laboratory study it is concluded that significant pressure changes occur in the pulp chamber during tooth preparation of extracted teeth when the remaining dentin thickness is less than 2 mm. PMID- 10709527 TI - An intraindividual clinical comparison of 2 metal-ceramic systems. AB - PURPOSE: It has been questioned whether the surface and color of the ceramic and the metal-ceramic bond strength of a titanium-ceramic system are comparable to those of a conventional noble alloy-ceramic system. It was therefore the aim of this study to carry out an intraindividual clinical comparison between crowns fabricated according to the Procera system (titanium copings veneered with a low fusing ceramic) and noble-alloy copings veneered with a medium-fusing ceramic. MATERIALS AND METHODS: Twenty-one crown pairs were fabricated for eighteen patients; three of the patients were each provided with two crown pairs. After 2 years nineteen crown pairs in sixteen patients could be compared. Clinical examinations were performed by two calibrated dentists who are long experienced in prosthetic dentistry. The crowns were rated according to the California Dental Association system. In addition, Bleeding Index and Margin Index were evaluated. RESULTS: After 2 years the quality of surface and color of the ceramic material seemed to have deteriorated more in titanium-ceramic crowns than in conventional metal-ceramic crowns, although the difference was not statistically significant. Regarding anatomic form, margin integrity, Bleeding Index, and Margin Index the differences between the two crown systems were small. CONCLUSION: The low-fusing ceramics have been subject to improvements during the last few years. Their bond strength to titanium seems to be comparable to that of conventional metal-ceramic systems. However, in the long run one problem may be the surface and color stability of low-fusing ceramics. To make extended long-term comparisons between the two metal-ceramic systems possible the present patient material will be followed for a longer period than the current 2 years. PMID- 10709535 TI - Assessing an active distracting technique for local anesthetic injection in pediatric dental patients: repeated deep breathing and blowing out air. AB - The aim of the study was to assess the effect of an active distraction technique that included the repeated breathing and blowing out of air on the pain behavior and facial display of children receiving local anesthesia injections prior to dental treatment. Fifty children between the ages of 3 and 7 years and who were undergoing dental treatment in a pediatric dental clinic were selected for this study. The children were randomly assigned to an intervention group or to a control group. The intervention group of 25 children was told to repeatedly breathe deeply before and during the administration of the injection and to blow the air out. The 25 control group children were given the injection in the slow manner without the repeated breathing and air blowing. Children in the intervention group demonstrated significantly less eyelid squeezing (P = 0.04). Also, more children in the intervention group than in the control group significantly expressed their wish to have the same technique used during the second visit (p = 0.033). Children in the intervention group generally demonstrated less hand and torso movements, less eyebrow bulging, and expressed less pain than in the control group. Boys significantly reported less pain after the injection. The results of this study indicate some advantages of distraction techniques (deep breaths and blowing air) prior to and during the administration of a local anesthetic injection in children. PMID- 10709536 TI - Efficacy of the nocturnal bite plate in the control of bruxism for 3 to 5 year old children. AB - Bruxism occurs in nearly 60% of children between 3 and 5 years, with important repercussions to the different components of the stomatognathic system. Nevertheless, there is little information in the literature about this topic. The aim of this study was to compare two groups of children with bruxism. One group was not submitted to treatment, serving as a control. To the other group, nocturnal bite plate was made. Cast models were made for both groups, to evaluate the progression of wear facets, during 8 months. The results are as follows: The 4 children of the control group displayed increased wear facets during the study period. On the other hand, of the 5 children that used nocturnal bite plate, showed no increase of wear facets, even after the removal of the device. From this study, we can conclude that the use of nocturnal bite plate is efficient against bruxism in 3- to 5-year-old children. PMID- 10709537 TI - Correction of a Class III malocclusion with a four-way sagittal appliance: case report. AB - This case report demonstrates the treatment of a Class III malocclusion, in a twelve-year-old girl, using a unique combination of a removable functional appliance, the "four-way sagittal" appliance and fixed appliance therapy. Efficient and effective correction of malocclusion was attained, following use of the "four-way sagittal" appliance. This appliance was designed on the Denar Witzig articulator, a powerful instrument in providing the clinician with a three dimensional perspective plus the ability of changing the location of the TMJ on the articulator. A significant increase in the maxillary length and maxillary width was achieved. PMID- 10709538 TI - Ectopic eruption of maxillary canine and premolars: case report. AB - This article presents a case of ectopic eruption of permanent teeth in the maxilla in the late mixed dentition. The authors advocate early diagnosis of ectopic eruption with a closer follow-up during the transition from primary to permanent dentition and space maintenance, as a way to avoid active orthodontic treatment for alignment ectopic erupted teeth. PMID- 10709539 TI - Transient extraocular muscle palsy resulting from inferior alveolar nerve block in children. AB - Transient extraocular muscle palsy after inferior alveolar nerve block was described in adults. Two children who had dental treatment under inferior alveolar nerve block developed this complication. One child suffered from lateral rectus muscle palsy, and the other child from levator palpebral muscle palsy. Both cases recovered spontaneously after a short time. PMID- 10709540 TI - Nonsurgical treatment of a sublingual ranula in a ten-month-old baby. AB - A clinical case of nonsurgical treatment of a sublingual ranula in a ten-month old baby is presented. PMID- 10709541 TI - Preservation of the dentition following removal of a central giant cell granuloma: a case presentation. AB - Clinical and radiographic presentation of a child with central giant cell granuloma in the early mixed dentition and a suggested conservative management technique for preserving the developing dentition is presented. PMID- 10709542 TI - Dental management of a pediatric patient with myelodysplastic syndrome. AB - Myelodysplastic syndrome in the pediatric population is an extremely rare hematological disorder. An eleven-year-old girl with a remarkable, past medical history of myelodysplastic syndrome is presented. She was treated for a falling hematocrit and platelet count, with splenectomy as a lifesaving measure. The features of the syndrome and treatment options are described. The clinical protocol for the dental management of the pediatric patient with myelodysplastic syndrome is discussed. PMID- 10709543 TI - Comparisons between cervical vertebrae and hand-wrist maturation for the assessment of skeletal maturity. AB - It was claimed that, skeletal maturity could be determined by using anatomical changes of the cervical vertebrae observed on the lateral cephalometric radiographs. In this method of assessment cervical vertebrae C2, C3, and C4 are observed and each patient is placed in a cervical vertebrae maturation index (CVMI). Fishman developed a system of hand-wrist skeletal maturation indicators (SMI), using four stages of bone maturation at six anatomic sites. The purpose of this study was to analyze associations between cervical vertebrae maturation index (CVMI) and skeletal maturation index (SMI). The second objective was to determine the reproducibility of the identifications on the lateral cephalograms and hand-wrist films. Lateral cephalometric and left hand-wrist radiographs of 180 untreated subjects (99 girls and 81 boys) aged from 8 to 18 years were obtained from the files of the Marmara University School of Dentistry, Department of Orthodontics. The results of this study indicated that cervical vertebrae maturation and hand-wrist skeletal maturation were significantly related. PMID- 10709544 TI - The prevalence of crowding, attrition, midline discrepancies and premature tooth loss in the primary dentition of children in Jeddah, Saudi Arabia. AB - The aim of the present study is to evaluate the prevalence of crowding, attrition, midline discrepancies and premature loss of primary molars in primary dentition of children resident in Jeddah, Saudi Arabia. Five hundred and two (502) children aged 4-6 years old residing in the city of Jeddah, Saudi Arabia provide the data for the analysis. Crowding was found in 14.7% of the subjects crowding in the maxilla occurred in 27 (5.4%) of the children, and 67 (13.4%) in the mandible. Simultaneous crowding in maxilla and mandible was present in 20 (3.98%) of the subject. The prevalence of crowding was significantly higher in the mandible than the maxilla (P > 0.01) and higher in male (P < 0.05). Midline shift was present in 10% of the children with females showing a significantly higher prevalence than the males (P < 0.001). Attrition was present in 167 (33.3%) of the children. One hundred and sixty children (31.9%) had attrition in enamel, and only 7(1.4%) had attrition, which exposed the dentin. Thirty-one (6.2%) out of 502 children had 51 (0.5%) missing teeth out of total number of 10,040 teeth. Tooth #84 was most commonly lost tooth (P < .025). Overall premature loss of 1st primary molars was found to be significantly higher than 2nd primary molars (P < .001). PMID- 10709545 TI - Temporomandibular joint dysfunction and occlusion in the mixed and permanent dentition. AB - The aim of this study was to investigate the relation between occlusal factors: Angle classification, overbite, overjet, openbite, anterior and posterior crossbite, scissors bite or buccal crossbite and lateral openbite--and the presence of mandibular dysfunction in a sample of 359 Turkish children with mixed and permanent dentition. Z Test was used to compare the results. It was found that, Class III malocclusion in the permanent dentition and openbite, overbite = 0, overjet = 0, anterior-posterior crossbite in the mixed dentition were related with TMD. PMID- 10709546 TI - Microleakage evaluation of pit and fissure sealants done with different procedures, materials, and laser after invasive technique. AB - This study evaluated the microleakage of pit and fissure sealants after different surface preparation (invasive technique and laser irradiation) and the use of different materials (fluoride resin-filled sealant, resin-modified glass ionomer cement and adhesive system). Eighty-four pre molars were used in this study, which were divided into seven groups. After the accomplishment of the different treatments, these were submitted to thermocycling process and assess for microleakage by examination under an epifluorescent microscope and scored zero to seven. Two specimens of each group were observed under scanning electron microscope (SEM). The results showed that laser irradiation did not lessen microleakage in pit and fissures when using a filled-resin sealant with fluoride or a resin-modified glass ionomer cement. The use of laser irradiation and adhesive system, followed by a resin-filled sealant with fluoride, showed the lowest microleakage scores in pit and fissures. Comparing this group to the resin modified glass ionomer cement group, there was statistical significance. The use of a adhesive system decreased microleakage when using a fluoride resin-filled sealant with or without previous laser irradiation; although it was not statistically significant. PMID- 10709547 TI - Effect of saliva contamination on micromorphological adaptation of single-bottle adhesives to etched enamel. AB - This study explored the topography of the interface of composite resin bonded with four single-bottle adhesives and enamel under salivary contaminated conditions. Prime & Bond 2.1, One Step, Tenure Quik and Syntac Single Component were evaluated. Thirty six sound molars were divided into three subgroups for each of the four adhesives. Small flat areas on the buccal and lingual surface were wet ground in enamel. The adhesives were tested under: uncontaminated condition (Group 1), contamination of the bonding site with fresh whole saliva for 20 seconds (Group 2) and rinsing the saliva for 20 seconds before adhesive application and after enamel etching (Group 3). After adhesive application button of composite resin was applied and light cured. The roots of the teeth were removed and the crowns with the bonded composite immersed in HCl acid till the composite resin buttons become free of enamel. The buttons were immersed in NaC10(4) and prepared for SEM examination. The bonding surface of the composite resin buttons revealed the formation of resin tags reflecting the different etching pattern of enamel. Saliva contamination either washed or unwashed did not affected the resin tags formation except for Syntac Single Component with contaminated unwashed enamel. The clinical significance of this in vitro research was that saliva contamination did not affect the resin tags formation of Prime & Bond 2.1, One Step, and Tenure Quick. For Syntac Single Component, only contaminated unwashed enamel decreased resin tags formation. PMID- 10709548 TI - Myofibroma of gingiva: report of a case with immunohistochemical and ultrastructural study. AB - Oral myofibroma is an uncommon, benign, solitary proliferation of myofibroblastic tissue. Few cases affecting maxillofacial region have been reported. We present a case of gingival myofibroma, diagnosed on clinical, histopathological, immunohistochemical, and ultrastructural basis. PMID- 10709549 TI - "He who fizzles at the finish was faulty from the first!" Above the gumline; the struggle to be the ruler of disease rages. PMID- 10709550 TI - The effects of managed care on the quality of dental hygiene care. AB - PURPOSE: This study examined whether patients insured by a managed dental care plan receive lower quality dental hygiene care than those not enrolled in a managed dental care plan. METHODS: Questionnaire data were gathered from 193 dental hygienists in the Chicago, Illinois area. Managed care was measured by a questionnaire item that assessed the proportion of managed care patients treated by the subject; quality of dental hygiene care was measured by items that assessed the extent to which the subject performed each of 23 dental hygiene tasks. RESULTS: A factor analysis of the 23 items assessing the quality of dental hygiene care indicated four meaningful factors named: periodontal procedures, appointment time, visual examinations, and oral examinations. Measures based on these factors were the dependent variables in regression analysis that included managed care as the primary independent variable and demographic characteristics of the subjects and their practices as control variables. Managed care had a significant (p < .05) negative relationship with appointment time, but did not have a significant relationship with periodontal procedures, visual examinations, or oral examinations. CONCLUSION: This study suggests that managed dental care program patients may have inadequate appointment time with a dental hygienist, which may affect whether they receive important services, such as oral health education. Dental hygienists in managed-care environments should be certain they are making effective use of the scheduled appointment time and procedures to ensure managed-care patients receive adequate time for dental hygiene care. PMID- 10709551 TI - Factors that affect health and well-being in dental hygienists; a comparison of Swedish dental practices. AB - PURPOSE: This study was designed to focus on the health and well-being of dental hygienists as a function of work environment and background. METHODS: A questionnaire was mailed to 471 randomly selected dental hygienists from members of the Swedish Dental Hygienists' Association (SDHA) to collect data on health, demographics, lifestyles, and working conditions in various delivery systems. Seventy-seven percent responded. RESULTS: Small dental practices were associated with better job control, work relations, management support, and development of job skills. In contrast, large practices were associated with higher finance related and personal job demand, anxiety, collegial pressure, and demands on social job skills. In discriminant analysis, finance-related job demands, physically demanding patients, and colleague cooperation showed significant differences in working condition variables between the delivery systems and dental practices. Dental hygienists' control over their job functions and the clarity of information explained by management had a variance in the applied competencies dimension by 41 percent. Thirty-seven percent of the variance of musculoskeletal problems in the upper-body region were attributed to physical exposure from clinical job tasks, work breaks, and anxiety over role competition with dentists. CONCLUSION: More job control and clearer information from management enables dental hygienists to optimally apply their competencies in the workplace. Physical job exposure, such as demands on manual/motor job skills, should be decreased and work breaks, as well as working relationships, with dentists should be improved to promote health and well-being. PMID- 10709552 TI - An assessment of issues related to clinical skill remediation in dental hygiene education. AB - PURPOSE: Within the confines of a lock-step dental hygiene education curriculum, the remediation of clinical skills poses a challenge for both the faculty responsible for assuring clinical competence and for the students expected to meet the stringent clinical performance criteria. To gain an understanding of how dental hygiene programs meet remediation challenges, a survey of U.S. dental hygiene programs was conducted. METHODS: A questionnaire designed to elicit information on specific remediation protocols, the type of instructional methods used in clinical remediation, and the management of faculty work load and compensation when the need for remediation was identified was developed. The questionnaire was pretested by dental hygiene program administrators and then distributed to 227 U.S. dental hygiene programs via mail and Internet services with a follow-up mailing to non-respondents. Data were analyzed and reported as descriptive statistics using the Statistical Package for Social Science (SPSS). RESULTS: An 80 percent (n = 181) response rate was obtained. A chi-square analysis of goodness of fit demonstrated no statistically significant difference between the respondents and the survey population in relationship to type of degree granted, educational setting, or geographic location. Results revealed an average student to clinical faculty ratio of five to one regardless of year of curriculum, educational setting, or geographic location. Just over half (53.6%, n = 97), reported having a written policy on clinical remediation with a clinical course syllabus being the most frequently cited mode of distribution. Ninety eight percent (n = 177) indicated that clinical faculty met regularly to discuss student clinical progress. Issuing an incomplete grade, requiring the student to attend additional clinical sessions, and allowing the student to continue with his or her peers was the most frequent action taken when clinical remediation was needed at the end of the academic term (32.6%, n = 59). The most frequent remediation intervention strategy employed was one-on-one faculty instruction (87.3%, n = 158). Typodont practice and extra clinical time under one-on-one supervision were also identified by over half of the programs as clinical remediation methods. The majority reported that faculty responsible for managing remediation activities received no financial compensation for the remedial instruction (83%, n = 150). CONCLUSIONS: The study demonstrates that dental hygiene programs do engage in student clinical skill remediation. Educators are encouraged to apply principles of psychomotor skill acquisition to increase remediation effectiveness. PMID- 10709553 TI - Thoracic outlet syndrome: a review of the literature. AB - Dental hygienists perform tasks that are repetitive, strenuous, and can contribute to cumulative trauma disorders (CTDs) of the upper body. Besides prevalent disorders of the hand and wrist, such as carpal tunnel syndrome (CTS), there are many other upper extremity disorders, including thoracic outlet syndrome (TOS). If a dental hygienist can identify symptoms and seek treatment early, prognosis for recovery is greatly improved. PMID- 10709554 TI - Incorporating veterinary preventive dental instruction into a traditional dental hygiene program. AB - A traditional dental hygiene program at the University of Texas Health Science Center, San Antonio, (UTHSCSA) incorporates veterinary preventive dental instruction into its community dentistry course. Students in the Certificate in Dental Hygiene Program attend two hours of didactic instruction on veterinary dentistry during the fall semester of their final year. The following spring semester students may choose a rotation in a veterinary clinic as one of their optional alternative practice sites. In addition, students are given the opportunity to participate in community service activities that are structured around veterinary dental care. PMID- 10709556 TI - Dental print media and their value in forensic odontology. AB - Teeth are one of the most important factors in the identification of unknown cadavers. One of the most efficient methods the police have of publicizing a case is to publish the victim's dental x-rays of treatment records in the dental print media which dentists will often recognize and thus be able to contribute to the identification. To evaluate the efficiency of this procedure, 177 cases published between 1975 and 1995 were reviewed. The results show that only 3% of the cases were identified from recognition in the dental print media. A survey of 500 dentists was carried out to gauge the efficacy of the presentation of identification cases in their dental print media. According to these results a revised method of presentation needs to be developed to improve the efficiency of this odontoforensic publicity medium. New technologies could also open new avenues for forensic identification such as digital x-raying or online data transfer. The Internet itself could be useful regarding international cooperation of forensic odontologists in the identification of unknown corpses. PMID- 10709555 TI - The effects of extreme temperatures on composite, compomer and ionomer restorations. AB - The composite resin, compomer and glass ionomer restorative materials used in modern dental practice are fire resistant and remain radiopaque. They shrink significantly and are likely to fall out of the cavities after being burnt, but they remain extremely important for identification purposes and it may be possible to identify the white filling materials used to restore teeth by their radiopacity and morphology. The radiographic density of these filling materials do not change significantly when exposed to heat and although they may be difficult to see with the naked eye, they will remain visible when radiographed. Furthermore, special care has to be taken when handling heated materials as their compressive strengths decrease significantly depending on temperature and time of exposure and damage is possible. PMID- 10709557 TI - A comparison of bitemark injuries between fatal wolf and domestic dog attacks. AB - Bitemark patterns in adult human victims following a fatal wolf pack attack and a domestic dog pack attack are compared. Both victims exhibited a concentration of wounds to the extremities, left and right torso, but not to the groin or internal organs. The neck and face of the domestic dog attack victim were primary sites of attack while the feral wolf pack victim was spared damage to the neck, but had facial tissue destroyed postmortem. Most punctures were found on the ventral aspect of the domestic victim and dorsal aspect of the feral victim. It is speculated that most wounds were attributable to dominant animals of both packs and in both victims and this suggests a co-ordination of activity between. Differences in bitemark patterns may well have been caused in part by differences in genetics, training, breeding, socialization and impetus of attack between wolves and dogs. Distinct differences in bitemark patterns were found in these two human victims of a wolf and domestic dog attacks. PMID- 10709558 TI - An approach to person identification by means of dental prostheses in a burnt corpse. AB - The identification of a burnt corpse is described. In this case it is suggested that the composition and properties of alloys used for dental prostheses are useful in determining the country of origin of the deceased in addition to the role they could play in an ordinary dental identification process. PMID- 10709559 TI - On denture marking. AB - During the last decades in Sweden dentures have been permanently marked with a stainless steel metal band incorporated into the acrylic and containing the patient's birth date, a special number, and "S" for Sweden. The last recommendation issued by the National Board of Health and Welfare states that "the patients shall always be offered denture marking and be informed about the benefit thereof. Denture marking is not permitted if the patient refuses it". Requirements for denture markers have been that they should be biologically inert (when incorporated into the denture), not be expensive, be easy to inscribe, be possible to retrieve after an accident, and survive elevated temperatures for a reasonable time under normal circumstances. Although the frequency of edentulousness has decreased in recent years due to the improvement in oral health there remains a need to address the issue of marking of complete dentures, because there is a large variation in the oral status of populations in different countries. Given that only one marked denture can reveal the identity of a deceased person when all other methods fail to do so, makes it worthwhile. Furthermore, denture marking is important in long-term care facilities. We have investigated the issue of denture marking in Europe and in the United States. The results from the European survey show that denture marking is, to our knowledge regulated by law only in Sweden and Iceland. In the US denture marking is so far mandatory in 21 states while New York State requires dentures to be marked if the patient requests it and several other states impose the obligation to mark dentures on long-term care facilities. Since there is no international consensus regarding the issue of denture marking it is important to address it. A survey from the Nordic countries has shown that if denture marking was in general use, the contribution to the establishment of identity by forensic odontology in cases of fire would increase by about 10%. This means that about 25 more individuals could have been identified if their dentures were marked. Increased international collaboration is needed to solve the issue of denture marking for clinical and forensic purposes. PMID- 10709560 TI - Gender determination by odontometrics in a Swedish population. AB - Gender determination of skeletal remains is fraught with uncertainty, especially in subadults. Many anatomical structures have been studied, but the teeth and their measurements seem to be the most reliable method in individuals whose secondary sexual characteristics have not yet developed. The purpose of this study was to investigate the accuracy with which gender can be differentiated by odontometric analyses in the Swedish population. The material consisted of 58 dental casts, 29 male and 29 female, ranging in age from 14 to 38 (mean 19) years. Measurements were made on the mesio-distal, bucco-lingual, mesiobuccal distolingual and distobuccal-mesiolingual diameters. The mean diameters for males were larger than those for females in all variables and 27 out of the 56 differences were statistically significant (p < 0.05). The upper canine had significant mean differences in all measurements. Lower canines, second upper and lower premolars, upper second molars and the lower first molars all had significant mean values in three of four variables. These findings support the usefulness of especially the canines in gender determination by odontometric analyses. It also shows high significant dimorphic values for some of the other variables investigated. PMID- 10709561 TI - The effect of temperature on sex determination using DNA-PCR analysis of dental pulp. AB - Forensic applications often necessitate the identification of human remains. This is made more difficult when the tissues have been exposed to high temperatures. Previously, metrical and non-metrical assessments of skeletal remains have been used to assess gender. Recent advances in molecular biology allow amplification of DNA from human blood, dental pulp and other tissues using the polymerase chain reaction (PCR), thus facilitating gender identification. The aim of this study was to investigate the efficacy of utilising DNA retrieved from the pulp of human teeth that had been exposed to different temperatures for different lengths of time, in order to assess gender. DNA was obtained from 94 teeth, 88 of which were isolated (44 male and 44 female), and six male teeth embedded in bone and soft tissue. A 106 base pair fragment from the X chromosome and a 112 base pair fragment from the Y chromosome was amplified from the amelogenin gene. PCR was shown to be 100% reliable when used to assess the gender of teeth which had been heated at 100 degrees C for 15 minutes but less reliable when the teeth were heated at higher temperatures for longer periods of time. Teeth encased in bone and soft tissue yielded better results when subjected to higher temperatures than did the isolated teeth. PMID- 10709562 TI - Evaluation of a bitemark using clear acrylic replicas of the suspect's dentition- a case report. AB - An assault occurred during which a bite was inflicted on the left ear of the victim, producing a laceration and severing a portion of tissue from the ear. During the course of their investigation police recovered a lacerated fragment of tissue thought to be of a person's left ear. Impressions of a suspect's dentition were made and cast in dental stone. Positive replicas of the occlusal surfaces of the suspect's dentition were subsequently made using acrylic resin. The ear fragment displayed a lacerated border and a surface which exhibited indentations. When compared with the details of the suspect's lower anterior teeth, correspondence was visible between the shape of the indentations and characteristics of the suspect's dentition. The use of transparent acrylic replicas of the suspect's dentition facilitated the interpretation and comparison between the marks retained in the ear fragment and the features of the suspect's dentition. PMID- 10709563 TI - Odontological identification in two high-impact, high-temperature accidents. AB - Here we report on two high-impact accidents both of which were complicated by fire. The first accident involved a light aircraft which crashed in a gorge while performing a low, slow flight. Both victims were found to have experienced extensive dental fracturing. The second accident was a high speed motor vehicle crash followed by incineration which left the two victims without dental fracturing. In the absence of comparative data on the effect of burning on teeth the cause of the foregoing remains unanswered. PMID- 10709564 TI - Radiography in forensic dental identification--a review. AB - It is recognised that reliable objective evidence is provided by medical and dental radiographs in forensic identification. This depends on the observation and comparison of anatomical and artificial structures recorded in both ante mortem and post-mortem radiographs, and which can be regarded as unequivocal evidence. The introduction of new technology into the area of radiographic imaging provides both clear advantages and also some cause for concern where image enhancement can be carried out which may give rise to dispute if this technology is to be applied to forensic dental identification. To date there have been no published forensic case reports involving the use of directly acquired digital radiographs but it is anticipated that this will change. The technical advantages and known limitations need to be considered if this method of radiography is to be applied in forensic dental identification. PMID- 10709565 TI - Operator agreement in the use of a descriptive index of complete denture quality. AB - BACKGROUND AND DESIGN: No generally accepted method for classification of clinical quality of complete dentures exists. While various methods have been proposed, little effort has been made to test observer agreement. The purpose of this study was to test observer agreement in the clinical assessment of complete denture quality using a systematic classification. RESULTS: Fifty seven (57) complete denture-wearing patients were examined on two occasions separated by two weeks. Observer agreement was measured using Cohen's? (kappa). High levels of observer agreement were found for upper and lower retention and stability and for quality of retruded jaw relationship. CONCLUSION: High levels of operator agreement were found for a method for classification of clinical quality of complete dentures suggesting that the classification may be useful as a research tool. PMID- 10709566 TI - Characteristics of children referred to a general anaesthetic service in Northern Ireland. AB - General anaesthetics are still frequently given to children for dental extractions and this method of treatment is most prevalent in regions such as Northern Ireland where high levels of dental disease persist in children. The aim of this study was to establish the social and dental characteristics of the children receiving general anaesthetics for dental extractions. Parents of children referred to the community dental extraction service in the Craigavon and Banbridge area of Northern Ireland completed a closed-ended questionnaire. In the sample there was a significantly lower level of maternal education than seen in the general population. There was a significantly higher level (p < 0.01) of dental anxiety seen in the sample group of children compared to the general population. For the primary dentition the corrected dmft values were higher than in the general population as was the untreated decay component while lower numbers of filled and of extracted teeth were seen. In the permanent dentition the caries experience and levels of extractions were similar to those seen in the general population while the level of untreated decay was higher and the mean number of fillings was lower. There was a similar pattern of attendance as that seen in the general child population. PMID- 10709568 TI - Environmental Mutagen Society 31st annual meeting. New Orleans, Louisiana, USA. April 8-13, 2000. Abstracts. PMID- 10709567 TI - Dental treatment in Romania. PMID- 10709569 TI - Prevention of coronary heart disease. Part II. Secondary prevention, detection of subclinical disease, and emerging risk factors. AB - The prevention of CHD should be a major priority among primary care physicians and subspecialists who have any dealing with the cardiovascular system. There is ample evidence from epidemiologic studies for the impact of specific risk factors on CHD events. There is also ample evidence from observational studies and clinical trials that interventions of lifestyle and pharmacologic therapy can decrease morbidity and mortality from CHD before or after the first event. It behooves the physician who wishes to practice good medicine to understand the pathophysiologic roles of the risk factors and the evidence from epidemiologic studies and clinical trials for their association with cardiovascular disease. It is important to determine the efficacy of interventions, both lifestyle and pharmacologic, in modifying CHD risk. To be effective in doing so, the practicing physician has to have the motivation to determine target goals for risk factor modification in each patient, to understand the patient's own motivations in modifying risk factors, and to define clearly with the patient the expectations of such interventions. Although there are guidelines for risk factor modification in modification of cholesterol and in hypertension, the periodic renewal of these guidelines reflects the changing concepts of risk and its modification. A cardiovascular risk factor intervention categorization is presented in Table 12. The physician must be convinced that such intervention is beneficial to the patient, cost-effective, and thus fulfills the expectations of medical practice. The practice of medicine in the evaluation and treatment of coronary heart disease has always been challenging and stimulating. The prevention of CAD disease should ultimately provide the greatest accomplishment. PMID- 10709570 TI - New agents in glaucoma therapy. PMID- 10709571 TI - Glaucoma associated with penetrating keratoplasty. PMID- 10709572 TI - Modeling glaucomatous optic nerve damage. PMID- 10709573 TI - Recent developments in glaucoma drainage implant surgery. AB - Refractory glaucoma remains a challenging problem for the ophthalmologist. Drainage implant surgery is a valuable and effective option for its treatment. Recent developments in implant design and surgical technique have decreased the incidence and severity of postoperative complications while increasing the efficacy of glaucoma drainage implant devices. PMID- 10709574 TI - Glaucomatous neurodegeneration and the concept of neuroprotection. AB - Glaucomatous neurodegeneration is a progressive pathological affair morphologically characterized by RGC death resulting in functional visual deterioration. Although a variety of primary insults ranging from mechanical trauma to ischemia to genetic susceptibility can initiate disease onset, apparently loss of trophic support and excitotoxin release remain the common themes responsible for neuronal damage and eventual cell death by apoptosis. The spread of this damage is perpetuated by secondary degeneration that allows directly injured neurons to release noxious and degenerative substances into the surrounding cells. Neuroprotection is a collective therapeutic approach the aim of which is to provide resilience to such neurons in an effort to prevent or delay progressive neuronal degeneration. At present, definitive neuroprotective therapy remains unachieved despite a variety of approaches having been defined and intensely pursued. However, the evolution in our understanding of glaucoma continues to offer reason for optimism. To this end, we remain unreservedly hopeful in the labor and committed work of dedicated investigators. We are all indebted to them. PMID- 10709575 TI - Phacoemulsification in the glaucoma patient. PMID- 10709576 TI - Prevention and management of hypotony after glaucoma surgery. AB - Postoperative hypotony is a common complication of glaucoma filtering surgery, particularly with adjunctive use of antifibrotic agents. Associated structural sequelae and reduced visual function may occur in some eyes, resulting in the low pressure syndrome. Precautions may be taken intraoperatively and postoperatively to decrease the likelihood of hypotony. Sometimes, despite these measures, the low-pressure syndrome still can occur, the management of which can be difficult. When simple observation does not result in spontaneous resolution, several noninvasive and invasive techniques are available, targeted at the cause of low IOP. PMID- 10709577 TI - Nonpenetrating deep sclerectomy: collagen implant and viscocanalostomy procedures. PMID- 10709578 TI - Optic nerve and retinal nerve fiber layer imaging in glaucomatous optic neuropathy. PMID- 10709579 TI - Macular edema. PMID- 10709581 TI - White-dot chorioretinal inflammatory syndromes. PMID- 10709580 TI - Evaluating macular function. PMID- 10709582 TI - Infectious maculopathy. PMID- 10709584 TI - Hereditary macular diseases. PMID- 10709583 TI - Drug-induced maculopathy. PMID- 10709585 TI - Molecular genetics of macular degeneration. PMID- 10709586 TI - Vision rehabilitation for age-related macular degeneration. AB - Though the numbers of patients with ARMD are high, associated referrals for vision rehabilitation are not. Practitioners need to refer patients with age related maculopathy when medical and surgical treatment are no longer possible, and patients need to be educated to that fact. The impact of improving activities of daily living may be monumental and benefits society as a whole. People who are visually impaired are often ill-prepared to deal with the substantial adjustment involved, further stressing their entire support system. It may not be safe for visual and systemic reasons for older adults to cook, clean, and maintain their home. Poor vision contributes to the already increased risk of falls and subsequent fractures in these patients. Individuals who may have already been told they can no longer drive now face the possibility of being unable to live in their houses. Their independence may be threatened dramatically and abruptly. All these circumstances contribute to anxiety and depression. Patients with ARMD need to be educated about their disease process (teaching that can never be assumed to have been initiated). They need to be educated that they will not go completely blind and that, with assistance, they can accomplish a great deal. With today's technology, it is not difficult to help visually impaired individuals with ARMD, unless they are not referred or lack motivation. The primary complaint of an individual with ARMD is recognition of central detail. This affects all activities of daily living, and patient performance is subject to the duration and severity of the disease (including the size, density, and location of the central scotoma) and to their understanding of the disease. Rubin and coworkers, found that slow reading performance of patients with a dense central scotoma might reflect inherent limitations of peripheral retina for complex visual tasks. ARMD in most cases lends itself to magnification that enlarges the object beyond the blind spot for visual recognition. Visual devices for distance, intermediate, and near tasks are usually helpful after patient education regarding their predicament and education for adaptation. Eccentric fixation techniques should be one of the first exercises mastered prior to further visual rehabilitation. Activities of daily living should be addressed with every individual, and appropriate assessment of existing problems and modifications to those problems should be implemented. Orientation and mobility should be offered to any individual who is legally blind or has difficulty with safe travel. A great deal of empathy is required on the part of the vision rehabilitation team. However, when patients lack of motivation, feel despair, or exhibit psychosocial overtones of reliance on others, they needs to be confronted, and appropriate action must be taken. Social work consultation and access to a support group can go a long way in mental strengthening and socialization. The author conducted a support group that, over a 10-year period, had a negligible dropout rate owing to the positive socialization obtained from attending the meetings. Older adults who are still working should be referred to an agency for vocational and financial resources if so desired. There is the issue of driving. In the United States, maintaining a driver's license is an important part of the quality of life. Older adults are the most rapidly growing segment of the driving population in the United States. The percentage of drivers older than 65 is expected to increase 17% by the year 2020. The rate of traffic fatalities among older adults has increased substantially, although the overall rate of fatalities is declining. The elderly drive fewer miles but have the highest rate of crashes per miles driven. Many important issues regard the older adult driver. (ABSTRACT TRUNCATED) PMID- 10709587 TI - History of photodynamic therapy. PMID- 10709588 TI - Diagnosis and management of macular holes. PMID- 10709589 TI - Surgical treatment of clubfoot: the significance of talocalcaneonavicular malposition correction. AB - In a retrospective study on the surgical management of clubfoot based on clinical and radiographic assessments, 171 feet in 137 patients were reviewed. The surgical procedure was selected according to the degree of the deformity. The more severe cases (group A, 75 feet) were surgically treated according to Turco's one-stage posteromedial release, whereas the milder degrees of deformity (group B, 96 feet) were corrected by elongation of the Achilles tendon with posterior capsulotomy. The mean age of the patients at surgery was 12.5 months in group A and 5.2 months in group B. The mean follow-up time for both groups was 12.2 years. At follow-up, 24 feet (41%) in group A and 52 feet (68%) in group B required repeat surgery. In group A the results were good in 51 feet (68%), fair in 15 (20%), and poor in 9 (12%). In group B, good results were obtained in 44 feet (45%), barely satisfactory results in 25 (27%), and poor results in 27 (28%). It is suggested that the accurate correction of talocalcaneonavicular and calcaneocuboid malposition is a prerequisite for successful surgical treatment of clubfoot. There was a tendency for a better result in group A when the patients were surgically treated between 6 and 12 months of age. PMID- 10709590 TI - Clubfoot analysis with three-dimensional foot models. AB - The purpose of this study was to develop a method of defining, in mathematical terms, the interpositional relationships of the bones of the hindfoot complex in the idiopathic clubfoot and the neurogenic clubfoot. The neurogenic clubfoot and contralateral normal-appearing foot of a stillborn infant with myelomeningocele, and the normal foot of a 10-year-old were sectioned with a cryomicrotome. Magnetic resonance images (MRIs) of the clubfoot and the normal foot of a 3-month old boy were obtained. Using a computer program, three-dimensional foot models were generated from the digitized cryomicrotome sections and from the MRIs. The central principal axes were determined for the talus and calcaneus. The long central principal axes of the talus and calcaneus were neutrally rotated with reference to the bimalleolar axis in the idiopathic clubfoot while in the neurogenic clubfoot the long central principal axis of the talus was medially rotated 52 degrees and that of the calcaneus 10 degrees. The talocalcaneal angles defined by the long central principal axes in the superior and medial views were 0 degree and 10 degrees, respectively, in the idiopathic clubfoot, and 42 degrees and 56 degrees, respectively, in the neurogenic clubfoot. PMID- 10709591 TI - Retinoic acid-induced clubfoot-like deformity: pathoanatomy in rat fetuses. AB - The purpose of this assay was to study the hindfoot patho-dynamic in clubfoot like deformity during fetal development. Experimental induction of clubfoot-like deformity in rat fetuses was produced by maternal administration of retinoic acid (120 mg/kg body weight) as a single intragastric dose on day 10 of pregnancy. Hindlimbs from fetuses at 17, 19, and 21 days were removed, and serial sections in three planes were made. Experimental and control hindlimbs were studied. There was clubfoot-like deformity in 86.5% of the experimental fetuses and none in the controls. Other associated malformations found were craniofacial (96.3%), neural tube (75.7%), and club-hand (40.3%) defects. Persistence of the embryonic position of the talus and tibia in fetuses with severe clubfoot-like deformity was observed. No overlapping between talus and calcaneus was seen. An equinus position, medialization of anterior segment, and lateralization and inward torsion of the posterior body of the calcaneous were observed. Results of this study showed that there are rotational anomalies in the hindfoot and full hindlimb from the beginning of the fetal period, and these anomalies increase during development. This simple model may allow us to gain better knowledge in congenital clubfoot deformity. PMID- 10709592 TI - Blount's disease: classification and treatment. AB - Forty-three tibia vara in 27 patients were analyzed retrospectively in two centers. The criteria for diagnosis of the child form are discussed. A simple classification is suggested to facilitate the choice of treatment. In stage 0 (possible Blount's disease), the patient is younger than 2 1/2 years, and an observation period is indicated for gathering data. In stage 1 (confirmed Blount's disease and absence of medial metaphyseal bony bridge), known as physis+, a valgization osteotomy is proposed. In stage 2 (evidence of a medial metaphysoepiphyseal bony bridge) known as physis-, valgization osteotomy with lateral epiphysiodesis and treatment of the lower limb discrepancy is proposed. For stages 1 and 2, there are two possibilities: normal medial tibial plateau or sloping of the medial tibial plateau, indicating a transphyseal elevation osteotomy. When one-step correction is proposed for stage 2 disorder, external fixators such as Orthofix or Ilizarov devices are useful. PMID- 10709593 TI - Natural course of osteochondritis dissecans in children. AB - Results are reported from an absence of physiotherapic, orthopaedic, or surgical treatment in 31 cases of osteochondritis dissecans in 24 children. The mean age at diagnosis was 11 years and 4 months, and all the children were suffering from pain for an average of 3 months. None of these children were treated, except for instructions to discontinue involvement in sports activities until their pain had disappeared. In all cases pain disappeared, and these children have all returned to their former activities. According to x-ray findings, 30 lesions disappeared totally, although there was one case of a loose body. As a result, absence of treatment is recommended for osteochondritis dissecans in children. PMID- 10709594 TI - Patellar height ratios in children: an interobserver study of three methods. AB - Several indices for patellar height measurement have been described to relate patellofemoral instability and maltracking. No known study has proved interobserver reliability of these indices when applied to growing knees. This study included comparisons of three of these indices: Caton-Deschamps, Blackburne Peel, and Koshino. Three observers measured patellar height on 36 lateral radiographs of children's knees. The best interobserver agreement was achieved by the Caton-Deschamps method, a simple, reliable, and reproducible index that is not affected by skeletal maturation. PMID- 10709595 TI - Gait pattern in patients with spastic diplegic cerebral palsy who underwent staged operations. AB - Fifteen patients with spastic diplegic cerebral palsy (CP) were monitored for a mean length of 9.5 years after they underwent staged operations and were evaluated by gait analysis, including joint motion in the sagittal plane and the ground reaction force (GRF) in three dimensions. Results showed an increased hip flexion (132%) at midstance, a reduction of peak knee flexion (PKF) during swing (45%) accompanied by an augmented time of PKF during swing (50%), and an increased dorsiflexion of the ankle during swing (293%) as well as its time during the gait cycle, in comparison with normal values. Moreover, significant decreases of the vertical GRF at the terminal stance and the forward and backward GRF were present. Additionally, it was found that a bilateral popliteal angle < 20 degrees is acceptable in spastic CP. Staged operations gave unpredictable results in the correction of contracture of the hamstrings, the Achilles tendon, and the iliopsoas. The authors are convinced that gait analysis is useful in evaluating these patients and enhances the results of operative treatment, and they have since changed their approach toward multilevel simultaneous corrections. PMID- 10709596 TI - Management of double-layered patellae by compression screw fixation. AB - Tripartite patella is a form of double-layered patella in which the anterior layer is bipartite. It was first described by Buttner in 1925 (1). More recently, an association with multiple epiphyseal dysplasia has been described (2,3). In this article, we describe a case of symptomatic tripartite patella in a 13-year old child who was subsequently found to have features of multiple epiphyseal dysplasia. Her patellae had an abnormal excursion with a click and a visible jump in the longitudinal line of movement, but no lateral instability. We present a surgical solution not previously described by fusion of the two main components and report an excellent result. PMID- 10709597 TI - Spontaneous ankylosis of the contralateral hip after unilateral adductor tenotomy in cerebral palsy. AB - This is the case report of a 15-year-old black male with spastic quadriplegia cerebral palsy who developed heterotopic ossification and spontaneous ankylosis of his contralateral nonoperative hip after unilateral adductor tenotomy. To the authors' knowledge, this is the only reported case of such an occurrence. The mechanism and possible risk factors are discussed as well as management of this complication. PMID- 10709598 TI - Left-sided congenital pseudarthrosis of the clavicula. AB - This a well-documented case of a 13-year-old girl with unilateral pseudarthrosis of the left clavicle without the presence of cervical ribs or dextrocardia. Magnetic resonance imaging could not detect any abnormality that could be related to the left-sided pseudarthrosis. PMID- 10709599 TI - Hypertrophic osteitis of the medial end of the clavicle. AB - A 9-year-old boy had a spontaneous onset of enlargement of the medial end of the clavicle due to extensive sclerosis and periosteal reaction. There was no clinical or laboratory evidence of infection. Biopsy revealed an inflammatory exudate, and histochemical staining for Langerhans'-cell histiocytosis was negative. Hypertrophic sclerosis causing painful enlargement of the medial end of the clavicle in isolation should be distinguished from condensing osteitis and chronic recurrent multifocal osteomyelitis. An early biopsy to exclude neoplasia is recommended. PMID- 10709600 TI - Osteofibrous dysplasia of the tibia: case report and review of the literature. AB - A case of osteofibrous dysplasia (OFD) of the tibia with 10 years of follow-up is presented. Spontaneous healing of this lesion occurred without any surgical intervention at the age of 10 years. The diagnosis was made retrospectively on the basis of clinical and radiographic appearance and evolution. The capricious nature and indolent course of this neoplasm has led to uncertainty regarding its etiology, evolution, and treatment. A review of the literature and the ongoing discussion about this matter is presented. PMID- 10709601 TI - Amputation stump overgrowth in a congenital above-elbow agenesis: a short report. AB - This is a case of bony overgrowth in a congenital agenesis stump, treated successfully with an autologenous stump plasty. PMID- 10709602 TI - Os vesalianum as a cause of lateral foot pain: a familial case and its treatment. AB - This is a rare case of bilateral symptomatic os vesalianum in a 13-year-old girl whose mother had the same condition unilaterally. We performed osteosynthesis and bone grafting instead of a simple resection to preserve the peroneus brevis tendon, with excellent results. PMID- 10709603 TI - Bilateral trigger thumbs in identical twins. AB - Bilateral trigger thumbs in 4-year-old identical male twins are reported. To the authors' best knowledge, this is the first true description of this condition in identical twins. All four thumbs were treated by surgical release of the A1 pulley, with good results. The causes proposed for congenital and acquired trigger thumb are discussed, and it is concluded that the cases described here support a genetic predisposition to the condition. PMID- 10709604 TI - Focal fibrocartilaginous dysplasia of the femur. AB - Two additional cases of femoral fibrocartilaginous dysplasia are reported. The condition produces such significant angulation of the articular surfaces of the knee that progressive deformity is of concern to parents and surgeon alike. In both the reported cases, corrective distal femoral osteotomy confirmed the histologic diagnosis and ensured satisfactory gait in later childhood. PMID- 10709605 TI - Catastrophic familial rhabdomyolysis: compartment syndrome with muscle fiber regeneration. AB - Two reports of patients with rhabdomyolysis are described. Patient 1 was a 4-year old-girl who had a 48-hour history of pyrexia and a 24-hour history of vomiting, drowsiness, polydipsia oliguria, and back pain. She could not walk easily because of tenderness in the calves. She was treated with furosemide and dopamine. On day 9, she was mobilized with the aid of physiotherapy. After 2 years, she tired easily but could walk normally. Patient 2 was a 3-year-old girl who had a 24-hour history of general malaise, peripheral aches and pain, and increasing drowsiness. She had severe swelling in the calves. Full-leg four compartment fasciotomies were performed on both calves. After patient 2 healed, she was observed to have muscle regeneration, which is very rare. PMID- 10709606 TI - Double-stress fracture of the tibia in a ten-year-old child. AB - A double-stress fracture of the tibia in a 10-year-old girl is described. Double stress fracture of the tibia has previously been described in association with osteoarthritic varus deformity of the knee but not, to our knowledge, in a child. It is important to establish the diagnosis of stress fracture in childhood because the differential diagnosis, both clinically and on imaging, includes malignancy that must be excluded while avoiding unnecessary invasive investigations. The site of the lesions, their appearance on magnetic resonance imaging, the absence of any soft-tissue involvement, and the clinical history made the diagnosis possible. The characteristics of stress fracture shown on magnetic resonance imaging are described. PMID- 10709608 TI - Results after preoperative traction and pinning in slipped capital femoral epiphysis: K wires versus cannulated screws. PMID- 10709607 TI - Osteochondritis of the medial cuneiform. AB - Osteochondritis of tarsal cuneiforms is a rare entity, barely reported in the medical literature. To the nine cases previously reported, we add our series with four cases of bilateral lesion of the medial cuneiform. Medial cuneiform osteochondritis may be the cause of foot pain and limping in children ages 4 to 6 years. In three of our four cases, it was a casual finding in feet being studied for some other problems. Only boys were affected, with bilateral lesions and coexistent navicular injury in half of our cases. Evolution was always satisfactory, with clinical and radiologic resolution in a period ranging from 4 to 72 months, and needing only symptomatic relief. PMID- 10709609 TI - IX Congress of the International Society of Hematology, Asian-Pacific Division. Bangkok, Thailand, October 24-28, 1999. Abstracts. PMID- 10709610 TI - 8th Workshop on Cell Biology of Bone and Cartilage in Health and Disease. Davos, Switzerland, April 1-4, 2000. 5th Workshop on Bisphosphonates--From Laboratory to the Patient. Davos, Switzerland, April 5-7, 2000. Abstracts. PMID- 10709611 TI - 4th Symposium on Probing Disorders of the White Matter. Abstracts. PMID- 10709612 TI - 4th World Congress on Inflammation. Paris, France, 27-30 June 1999. Abstracts. PMID- 10709613 TI - 31st Annual General Meeting of the European Brain and Behavior Society (EBBS). Rome, Italy, 29 September-2 October 1999. Abstracts. PMID- 10709614 TI - 7th International Congress of the European Association of Endoscopic Surgery (EAES). Linz, Austria, 23-26 June 1999. Abstracts. PMID- 10709615 TI - 3rd Joint meeting of the German, the Austrian and the Swiss Societies for Thoracic and Cardiovascular Surgery. Lucerne, Switzerland, February 9-12, 2000. Abstracts. PMID- 10709616 TI - Improved HPLC determination of acidic opines by phenylisothiocyanate derivatization and its application to marine animals. AB - We present here a reliable and sensitive method for the determination of acidic opines such as meso-alanopine, beta-alanopine, tauropine and strombine in biological samples. Interfering primary amino acids were eliminated by reaction with o-phthalaldehyde, and the derivatized compounds were passed through Sep-Pak Plus PS-1 cartridges. The acidic opines were recovered by flushing the cartridges with water, then determined by high performance liquid chromatography after a second derivatization with phenylisothiocyanate. All 4 acidic opines were detected within 30 min. This method ensured good separation and guaranteed almost full recovery of all acidic opines. This method was applied to analyze opines in marine animals and to test whether opines are metabolized in the livers of the rat and fish. PMID- 10709617 TI - Effects of thyroxine on L-cysteine desulfuration in mouse liver. AB - The effect of exogenous thyroxine (T4) administration on the activity of rhodanese, cystathionase, and 3-mercaptopyruvate sulfurtransferase (MPST) in the mitochondrial and cytosolic fractions of mouse liver was investigated. Three groups of mice were treated for 6 consecutive days with subcutaneous injections of T4 (50 micrograms, 100 micrograms, and 250 micrograms per 100 g of body wt, respectively). The other 3 groups were given 100 micrograms of T4 per 100 g of body wt for 1, 2, or 3 days. The dose of 100 micrograms T4 per 100 g of body wt given for 6 days exerted the strongest effect on the activity of all of the investigated enzymes. In comparison to the control, rhodanese activity diminished in the mitochondrial fraction by 40% (P < 0.05), cystathionase activity diminished in the cytosolic fraction by 15% (P < 0.05), and MPST activity in the mitochondrial fraction was reduced by 34% (P < 0.05), whereas cytosolic MPST activity was unaltered. Simultaneously, in the liver homogenate, elevated levels of ATP and sulfate were observed after 6 days of T4 administration. Thus, the present results seem to suggest that in the mouse liver, after 6 days of administration of 100 micrograms T4 per 100 g of body wt, the desulfuration metabolism of L-cysteine is diminished, which is probably accompanied by an increase in oxidative L-cysteine metabolism. The dose of 100 micrograms per 100 g of body wt administered for a shorter period, and the use of a lower dosage (50 micrograms T4 per 100 g of body wt) for 6 days had a stimulatory effect upon MPST activity level, and an increased level of sulfane sulfur was observed. PMID- 10709618 TI - Increase in cerebral blood flow as a predictor of hyperbaric oxygen-induced convulsion in artificially ventilated rats. AB - In spontaneously breathing rats, a transient increase in cerebral blood flow (CBF) has been shown to be a predictor of hyperbaric oxygen (HBO)-induced convulsion. In the present study, we evaluated whether artificially ventilated animals also show an increase in CBF prior to the onset of HBO-induced convulsion. Rats were ventilated with 100% oxygen in 5 atmospheres. CBF, blood pressure, and an electroencephalogram were monitored continuously. Convulsion was observed at 41 +/- 12 min after the initiation of HBO treatment. A single abrupt increase in CBF, reaching 223 +/- 39% of the control level, was observed at 29 +/ 13 min after the initiation of HBO exposure and lasted until the onset of convulsion 12 +/- 2 min later. The time of the increase in CBF correlated strongly with the onset of convulsion (r = 0.99, P < 0.001). Further, the logistic regression curve demonstrated a close relationship between the duration of increased CBF and percentage of epileptiform electrical-discharge incidence (r = 0.92, P < 0.006). The durations of increased CBF causing convulsion in 10%, 50%, and 90% of the rats were 8.4 min, 11.7 min, and 15.1 min, respectively. These results indicate that an increase in CBF is a predictor of HBO-induced convulsion in artificially ventilated rats. The increase in CBF may be involved in the pathogenesis of HBO-induced convulsion. PMID- 10709619 TI - Forensic study of sex determination using PCR on teeth samples. AB - In this study, sex determination using polymerase chain reaction (PCR) on tooth material was evaluated from the viewpoint of forensic medicine. The sensitivity of PCR for detection of the Y chromosome-specific alphoid repeat sequence and the X chromosome-specific alphoid repeat sequence was 0.5 pg of genomic DNA. Sex could be determined by PCR of DNA extracted from the pulp of 16 freshly extracted permanent teeth and dentine including the surface of the pulp cavity of 6 freshly extracted milk teeth. Sex could be determined using the pulp in all 20 teeth (10 male and 10 female) preserved at room temperature for 22 years. For the pulp of teeth stored in sea water, the sex could be determined in all 8 teeth immersed for 1 week and in 5 of 6 teeth immersed for 4 weeks. In the remaining 1 tooth, in which sex determination based on the pulp failed, the sex could be determined correctly when DNA extracted from the tooth hard tissue was examined. For teeth stored in soil, the sex could be determined accurately in all 8 teeth buried for 1 week, 7 of 8 teeth buried for 4 weeks, and in all 6 teeth buried for 8 weeks. When teeth were heated for 30 min, sex determination from the pulp was possible in all teeth heated to 100, 150, and 200 degrees C, and even in some teeth heated to 250 degrees C. When this method was applied to actual forensic cases, the sex of a mummified body estimated to have been discovered half a year to 1 year after death could be determined readily by examination of the dental pulp. In the skeletons of 2 bodies placed under water for approximately 1 year and approximately 11 years and 7 months, pulp tissues had been dissolved and lost, but sex determination was possible using DNA extracted from hard dental tissues. These results indicate that this method is useful in forensic practices for sex determination based on teeth samples. PMID- 10709620 TI - Work conditions as risk factors for varicose veins of the lower extremities in certain professions of the working population of Rijeka. AB - This research aims to establish the effect of working conditions on the appearance of varicose veins. The epidemiological study was carried out on 1,324 examinees, 530 males and 794 females, employed in 5 highly represented groups of professional activities in Rijeka (catering, trade, light industry, heavy industry and finances). The data were collected by survey and clinical examination. Varicose veins were more prevalent in the trade than in the office workers (odds ratio (OR) = 2.08; 95% confidence interval (CI) = 1.31-3.31), and more prevalent in catering industries than in the office workers (OR = 1.56; 95% CI = 1.001-2.43). chi 2-testing suggested that standing in the workplace (OR = 1.35; 95% CI = 0.95-1.92), weight handling while working (OR = 1.29; 95% CI = 1.01-1.64) and working indoors (OR = 1.61; 95% CI = 1.02-2.53) were risk factors for varicose veins. By multiple logistic regression, the following risk factors were isolated in the total population: female sex (OR = 1.92; 95% CI = 1.37 2.67), workplace (OR = 0.89; 95% CI = 0.78-0.99), age (OR = 1.05; 95% CI = 1.03 1.07), body mass index (OR = 1.04; 95% CI = 1.01-1.07) and family history of the disease (OR = 1.99; 95% CI = 1.55-2.57). PMID- 10709621 TI - Helicobacter pylori infection: relationship between seroprevalence and dietary preference in a rural area. AB - In order to evaluate the relationship between Helicobacter pylori (H. pylori) infection and dietary preference, a cross-sectional study was performed among 626 residents in a rural area of Japan. Seropositive rates were 88.7% in males and 71.4% in females, and these increased with age for both sexes [male P < 0.05 and female P < 0.01]. The relationship between H. pylori-seropositivities and salted food intake, after adjustment for age, demonstrated a significant result in the "almost every day" group in males with an odds ratio (OR) of 8.39 and with 95% confidence intervals (CI) of 1.02-69.30. As regards an association between seropositivities of H. pylori and levels of serum pepsinogens for the screening of chronic atrophic gastritis (low pepsinogen values used were a pepsinogen I level below 70 ng/ml and a pepsinogen I/pepsinogen II ratio below 3.0), the ORs of H. pylori-seropositivities for low pepsinogen cases were 6.32 [95% CI: 1.42 28.03] in males and 12.72 [95% CI: 4.57-35.46] in females. With regard to the relationship between low pepsinogen cases and light-colored vegetables intake, a significant low OR for the low pepsinogen cases was obtained in the "almost every meal" group in females after adjustment for age and seropositivities of H. pylori with an OR of 0.37 and with 95% CI of 0.15-0.92. PMID- 10709622 TI - Influence of exposure to new circumstances on pharmacokinetics of plasma drugs concentrations in rats. AB - The influences of emotional changes induced by being exposed to a new environment on the pharmacokinetics of plasma drug concentration were studied in male Wistar rats. Transfer from a familiar home cage to a new home cage was considered to induce psychological (non-physical) emotional changes. First, nicorandil and zonisamide, drugs that act on the peripheral system and central nervous systems, were used, respectively. Immediately after oral administration of nicorandil (10 mg/kg) or zonisamide (50 mg/kg), the animals were transferred to new home cages. Plasma nicorandil and zonisamide concentrations were determined by high performance liquid chromatography at 1 and 4 h after administration. Plasma nicorandil concentration in the group transferred to new home cages was significantly decreased relative to levels in the non-transferred control group. However, zonisamide concentrations were unchanged. These findings suggest that the pharmacokinetics of nicorandil, but not those of zonisamide, tend to be influenced by non-physically induced emotional changes. PMID- 10709623 TI - Biogenesis of the rat liver mitochondrial carnitine palmitoyltransferase I. PMID- 10709624 TI - Subcellular distribution of mitochondrial carnitine palmitoyltransferase I in rat liver. Evidence for a distinctive N-terminal structure of the microsomal but not the peroxisomal enzyme. AB - Mitochondria, microsomes and peroxisomes all express overt (cytosol-facing) carnitine palmitoyltransferase activities that are inhibitable by malonyl-CoA and are collectively termed CPTo. In order to quantify the relative contribution of the different membrane systems towards overall hepatocyte activity, all three membrane fractions and a high-speed supernatant (soluble) fraction were prepared quantitatively from rat liver homogenates. The overt (malonyl-CoA-sensitive) carnitine palmitoyltransferase activity (CPTo) associated with the different fractions were measured. In parallel experiments, rat livers were perfused in situ with oxygenated medium containing dinitrophenyl (DNP)-etomoxir in order to label covalently (with DNP-etomoxiryl-CoA) and quantitatively the molecular species responsible for CPTo activity in each of the membrane systems under near physiological conditions. Mitochondria accounted for only 65% of total cellular overt CPT activity, with the microsomal and peroxisomal contributions accounting for the remaining 25% and 10%, respectively. A single major protein with an identical molecular size (Mr 88,000) was labelled by DNP-etomoxir perfusion in all three membrane fractions. The abundance of this 88 kDa protein in each fraction was quantitatively positively related to the respective specific activities of overt CPT. The same protein was immunoreactive with three anti peptide antibodies raised against linear epitopes within the N- and C-terminal and loop (L) domains of the mitochondrial outer membrane CPT I of the liver mitochondrial outer membrane (L-CPT I). However, whereas reaction with anti-L and anti-C antipeptide antibodies were proportional to the respective overt CPT activities and DNP-etomoxir labelling in all three membrane fractions, reaction with anti-N peptide antibody was much stronger for microsomal CPT. We conclude that in all three membrane systems overt CPT activity is associated with the same or highly similar molecular species to mitochondrial outer membrane CPT I, but that the protein expressed in microsomes has a modified N-terminal domain, which gives the microsomal enzyme its higher malonyl-CoA sensitivity and may target the protein to its microsomal location. PMID- 10709625 TI - Topology of hepatic mitochondrial carnitine palmitoyltransferase I. AB - Our earlier work using intact mitochondria and isolated mitochondrial outer membranes confirms the observations of Murthy and Pande that CPT-I is located on the mitochondrial outer membranes and supports the notion that this enzyme has a malonyl-CoA binding domain facing the cytosol and an acyl-CoA binding domain facing the inter membrane space. Our data also suggests that coenzyme A binds at the active site of CPT-I, as does acyl-CoA, 2-bromopalmitoyl-CoA, and (+) hemipalmitoylcarnitinium, but malonyl-CoA does not bind at that site. Inhibition of CPT-I at the malonyl-CoA binding site by HPG and Ro 25-0187, which have no CoA moiety, contributes to a resolution of this question in that the CoA itself is not essential for the binding of malonyl-CoA to its regulatory site, but the dicarbonyl function which is present in malonyl-CoA, HPG, and Ro 25-0187 is absolutely essential. Our re-evaluation of the topology of hepatic mitochondrial CPT-I confirms the original observations that this enzyme has at least two different binding domains, one domain binding malonyl-CoA, HPG, and Ro-25-187 and the other domain binding acyl-CoA and other inhibitors of CPT-I. Furthermore, the malonyl-CoA binding domain is exposed to the cytosolic face of the membrane. Our data showing that treatment of the intact mitochondria with trypsin causes release of adenylate kinase which indicates that trypsin has damaged the mitochondrial outer membrane, possibly allowing trypsin to enter the intermembrane space and act on CPT from within the outer membrane. Since trypsin's action is limited to arginine and lysine residues, an alternative explanation could be that the portion of the protein domain responsible for malonyl-CoA inhibition may not contain these residues. The latter explanation is plausible, since malonyl-CoA was able to protect against loss of activity and sensitivity to inhibition, but did not protect against loss of adenylate kinase, suggesting that rupture of the outer membrane is not necessarily related to loss of CPT activity. These results suggest that some protein domain that is necessary for CPT activity is exposed on the outer surface of the outer membranes. Therefore, it seems likely that trypsin would have to be able to hydrolyse protein domains of CPT that are inaccessible to Nagarse and papain. PMID- 10709626 TI - Possible involvement of cytoskeletal components in the control of hepatic carnitine palmitoyltransferase I activity. PMID- 10709627 TI - Effects of 3-thia fatty acids on beta-oxidation and carnitine palmitoylatransferase I activity in cultured rat hepatocytes. PMID- 10709628 TI - Carnitine acyltransferases and associated transport processes in the endoplasmic reticulum. Missing links in the VLDL story? PMID- 10709629 TI - Reciprocal enzymatic interference of carnitine palmitoyltransferase I and glycerol-3-phosphate acyltransferase in purified liver mitochondria. AB - (i) Highly purified mitochondrial fractions were practically devoid of microsomal contamination and of acyl-CoA ligase activity. (ii) In mitochondria, glycerol-3 phosphate acyltransferase (GPAT) activity was supported by two enzymes, the first being very active at low palmitoyl-CoA/albumin ratios and sensitive to external agents (external form), the second being detected only at higher palmitoyl CoA/albumin ratios and insensitive to external agents (internal form). (iii) Carnitine palmitoyltransferase I (CPT I) activity was shown to inhibit external GPAT activity only. (iv) Glycerol-3-phosphate exerted an inhibitory effect on CPT I, even when GPAT was inactive. Reciprocal interaction of CPT I and GPAT was discussed with regard to the balance existing between fatty acid oxidation and esterification metabolic pathways. PMID- 10709630 TI - Characterization of a response element for peroxisomal proliferator activated receptor (PPRE) in human muscle-type carnitine palmitoyltransferase I. PMID- 10709631 TI - Kinetic investigation of carnitine palmitoyltransferases in homogenates of human skeletal muscle using L-amino-carnitine and malonyl-CoA. PMID- 10709632 TI - Processing of carnitine octanoyltransferase pre-mRNAs by cis and trans-splicing. AB - Trans-splicing is a mechanism by which two pre-mRNAs are processed to produce a mature transcript that contains exons from both precursors. This process has been described mostly in trypanosoma, nematodes, plant/algal chloroplasts and plant mitochondria [Bonen et al. (1993) FASEB J. 7, 40-46]. Our studies clearly demonstrate that a trans-splicing reaction occurs in the processing of the carnitine octanoyltransferase (COT) gene in rat liver. Three different mature transcripts of COT have been found in vivo, the canonical cis-spliced mRNA and two trans-spliced transcripts, in which either exon 2 or exons 2 and 3 are repeated. Splicing experiments in vitro also indicate the capacity of exon 2 to act either as a donor or as an acceptor of splicing, allowing the trans-splicing reactions to occur. PMID- 10709633 TI - Selective modulation of carnitine long-chain acyltransferase activities. Kinetics, inhibitors, and active sites of COT and CPT-II. AB - Carnitine acyltransferases in mitochondria, peroxisomes and the endoplasmic reticulum are different gene products and serve different metabolic functions in the cell. Here we summarize briefly evidence that carnitine octanoyltransferase (COT) from the peroxisomes and carnitine palmitoyltransferase II (CPT-II) from the mitochondria (both matrix facing enzymes) differ kinetically and demonstrate that they differ in their sensitivity to conformationally constrained inhibitors that mimic the reaction intermediate. Medium chain inhibitors are 15 times more effective on COT than on CPT-II and long chain inhibitors, such as hemipalmitoylcarnitinium, 80 times more effective on the mitochondrial enzyme. Thus, it may be possible to develop inhibitors to inhibit mitochondrial beta oxidation with minimal effects on peroxisomal beta-oxidation and other acyl-CoA dependent reactions. PMID- 10709634 TI - Confocal laser scanning microscopy of human skin fibroblasts showing transient expression of a green fluorescent carnitine palmitoyltransferase 1 fusion protein. PMID- 10709635 TI - Carnitine biosynthesis. Purification of gamma-butyrobetaine hydroxylase from rat liver. AB - gamma-Butyrobetaine hydroxylase catalyse the last step in carnitine biosynthesis, the formation of L-carnitine from gamma-butyrobetaine, a reaction dependent on Fe2+, alpha-ketoglutarate, ascorbate and oxygen. Initial attempts to purify the protein from rat liver showed that gamma-butyrobetaine hydroxylase is unstable. We, therefore, determined the influence of various compounds on the stability of gamma-butyrobetaine hydroxylase at different storage temperatures. The enzyme activity was best conserved by storing the protein at 4 degrees C in the presence of 200 g/l glycerol and 10 mM DTT. We subsequently purified the enzyme from rat liver to apparent homogeneity by liquid chromatography. PMID- 10709636 TI - Hypolipidemic 3-thia fatty acids. Fatty acid oxidation and ketogenesis in rat liver under proliferation of mitochondria and peroxisomes. PMID- 10709637 TI - Molecular mechanisms of fatty acid beta-oxidation enzyme catalysis. PMID- 10709638 TI - Control of mitochondrial beta-oxidation at the levels of [NAD+]/[NADH] and CoA acylation. PMID- 10709639 TI - Production and export of acylcarnitine esters by neonatal rat hepatocytes. PMID- 10709640 TI - Tissue specific differences in intramitochondrial control of beta-oxidation. PMID- 10709641 TI - Endotoxin-induced changes in very-low-density lipoprotein and myocardial utilisation of triacylglycerol from abnormal VLDL in the rat. PMID- 10709642 TI - Effect of valproic acid on the expression of acyl-CoA dehydrogenases in various tissues. PMID- 10709643 TI - Formation of a human "electron transferring flavoprotein". Medium chain acyl coenzyme A dehydrogenase complex, preliminary evidence from crosslinking studies. PMID- 10709644 TI - Cloning and regulation of peroxisome proliferator-induced acyl-CoA thioesterases from mouse liver. AB - 1.1. Acyl-CoA thioesterases hydrolyze acyl-CoAs to the corresponding free fatty acid plus CoASH. The activity is strongly induced in rat and mouse liver after feeding the animals peroxisome proliferators. To elucidate the role of these enzymes in lipid metabolism, we have cloned the cDNAs corresponding to the inducible cytosolic and mitochondrial type I enzymes (CTE-I and MTE-I) and studied tissue expression and nutritional regulation of expression of the mRNAs in mice. The constitutive expression of both mRNAs was low in liver, with CTE-I being expressed mainly in kidney and brown adipose tissue and MTE-I expressed in brown adipose tissue and heart. As expected, the expression in liver of both the CTE-I and MTE-I mRNAs was strongly induced (> 50-fold) by treatment with clofibrate. A similar level of induction was observed by fasting and a time course study showed that both mRNAs were increased already at 6 hours after removal of the diet. Refeeding normal chow diet to mice fasted for 24 hours normalized the mRNA levels with a T1/2 of about 3-4 hours. Feeding mice a fat free diet further decreased the expression, possibly indicating repression of expression. The strong expression of MTE-I and CTE-I in the heart was increased about 10-fold by fasting. To further characterize these highly regulated enzymes, we have cloned the corresponding genes and promoter regions. The structures of the two genes were found to be very similar, consisting of three exons and two introns. Exon-intron borders conform to general consensus sequences and especially the first exon appears to be highly conserved. The promoter regions of both the CTE-I and MTE-I genes contain putative peroxisome proliferator response elements (PPREs), suggesting an involvement of peroxisome proliferator-activated receptors in the regulation of these genes. PMID- 10709645 TI - Metabolic effects of 3-thia fatty acid in cancer cells. PMID- 10709646 TI - Poorly oxidizable fatty acid analogues inhibit the proliferation of cancer cells in culture. PMID- 10709647 TI - The role of PPAR alpha as a "lipostat" transcription factor. PMID- 10709648 TI - The hypolipidaemic effect of EPA is potentiated by 2- and 3-methylation. PMID- 10709649 TI - Is it time to reconsider the role of CPT I in control of hepatic ketogenesis? PMID- 10709650 TI - Developmental comparison of human and rat hepatic mitochondrial 3-hydroxy-3 methylglutaryl-CoA synthase. PMID- 10709651 TI - Regulation of the ketogenic enzyme mitochondrial 3-hydroxy-3-methylglutaryl-CoA synthase in astrocytes and meningeal fibroblasts. Implications in normal brain development and seizure neuropathologies. PMID- 10709652 TI - Studies on regulation of the peroxisomal beta-oxidation at the 3-ketothiolase step. Dissection of the rat liver thiolase B gene promoter. AB - The peroxisomal 3-oxoacyl-CoA thiolase (thiolase) is the last enzyme involved in the beta-oxidation of fatty acids. The enzyme cleaves long chain fatty acyl-CoA to generate acetyl-CoA and shortened acyl-CoA. The enzyme is nuclear encoded, synthesized in the cytoplasm and transported into peroxisomes. The thiolase B gene is inducible by the peroxisome proliferator compounds, like other genes involved in beta-oxidation of fatty acids in peroxisomes. The importance of studying thiolase is that it generates acetyl-CoA which is the precursor for the synthesis of molecules like cholesterol and fatty acids. The structural and functional analysis of thiolase at molecular level may add to the knowledge of fatty acid metabolism and further the obesity phenomenon. It is known that several genes mediate lipid homeostasis in target organs like liver, adipose tissue and are regulated by peroxisome proliferator activated receptors (PPAR alpha and PPAR gamma). To elucidate the mechanism of induction of rat liver thiolase B gene, an upstream 2.8 kb fragment containing promoter element has been subcloned and partially sequenced. The sequence analysis revealed a putative PPRE (Peroxisome Proliferator Response Element) of AGACCT T TGAACC sequence at -681 to -668 [Kliever et al. (1992) Nature 358:771-774]. By transient expression of a luciferase reporter gene in HeLa cells, we conclude that the identified PPRE could be functional in induction of thiolase B gene, but other sequences of genes might be involved. PMID- 10709653 TI - Role and organization of peroxisomal beta-oxidation. AB - In mammals, peroxisomes are involved in breakdown of very long chain fatty acids, prostanoids, pristanic acid, dicarboxylic fatty acids, certain xenobiotics and bile acid intermediates. Substrate spectrum and specificity studies of the four different beta-oxidation steps in rat and/or in man demonstrate that these substrates are degraded by separate beta-oxidation systems composed of different enzymes. In both species, the enzymes acting on straight chain fatty acids are palmitoyl-CoA oxidase, an L-specific multifunctional protein (MFP-1) and a dimeric thiolase. In liver, bile acid intermediates undergo one cycle of beta oxidation catalyzed by trihydroxycoprostanoyl-CoA oxidase (in rat), or branched chain acyl-CoA oxidase (in man), a D-specific multifunctional protein (MFP-2) and SCPX-thiolase. Finally, pristanic acid is degraded in rat tissues by pristanoyl CoA oxidase, the D-specific multifunctional protein-2 and SCPX-thiolase. Although in man a pristanoyl-CoA oxidase gene is present, so far its product has not been found. Hence, pristanoyl-CoA is believed to be desaturated in human tissues by the branched chain acyl-CoA oxidase. Due to the stereospecificity of the oxidases acting on 2-methyl-branched substrates, an additional enzyme, 2-methylacyl-CoA racemase, is required for the degradation of pristanic acid and the formation of bile acids. PMID- 10709654 TI - Hepatic alpha-oxidation of phytanic acid. A revised pathway. AB - Synthetic 3-methyl-branched chain fatty acids were used to decipher the breakdown of phytanic acid. Based on results obtained in intact or permeabilized rat hepatocytes, rat liver homogenates or subcellular fractions, a revised alpha oxidation pathway is proposed which appears to be functioning in man as well. In a first step, the 3-methyl-branched chain fatty acid is activated by an acyl-CoA synthetase. This reaction requires CoA, ATP and Mg2+. Subsequently, the acyl-CoA ester is hydroxylated at position 2 by a peroxisomal dioxygenase. This step is dependent on alpha-oxoglutarate, ascorbate (or glutathione), Fe2+ and O2. The 2 hydroxy-3-methylacyl-CoA intermediate is cleaved by a peroxisomal lyase to formyl CoA and a 2-methyl-branched fatty aldehyde. Formyl-CoA is (partly enzymically) hydrolyzed to formate, which is then converted, most likely in the cytosol, to CO2. In the presence of NAD+, the aldehyde is dehydrogenated to a 2-methyl branched fatty acid, presumably by a peroxisomal aldehyde dehydrogenase. This acid can--after activation--be degraded via a D-specific peroxisomal beta oxidation system. PMID- 10709655 TI - Functions and dysfunctions of peroxisomes in fatty acid alpha- and beta oxidation. New insights. PMID- 10709656 TI - Enzymology of beta-oxidation of (poly)unsaturated fatty acids. PMID- 10709657 TI - The effect of beta-oxidable and non-beta-oxidable thia fatty acids on fatty acid metabolism. PMID- 10709658 TI - EPA and DHA possess different metabolic properties. PMID- 10709659 TI - The use of [9,10-3H]myristate, [9,10-3H]palmitate and [9,10-3H]oleate for the detection and diagnosis of medium and long-chain fatty acid oxidation disorders in intact cultured fibroblasts. PMID- 10709660 TI - Rapid diagnosis of organic acidemias and fatty-acid oxidation defects by quantitative electrospray tandem-MS acyl-carnitine analysis in plasma. AB - The analysis of circulating free carnitine and acyl-carnitines provides a powerful selective screening tool for genetic defects in mitochondrial fatty acid oxidation and defects in the catabolism of branched chain amino acids. Using electrospray tandem mass spectrometry (ESI/MS/MS) we developed a sensitive quantitative analysis of free carnitine and acyl-carnitines in plasma and/or serum. This method was evaluated by analyzing 250 control samples and 103 samples of patients suffering from twelve different defects in either mitochondrial fatty acid oxidation or the catabolism of branched chain amino acids. The reproducibility of the method was acceptable with a day-to-day coefficient of variation ranging from 6-15% for free carnitine and the different acylcarnitines. Except for one patient with a mild form of short chain acyl CoA dehydrogenase (SCAD) deficiency and a single sample from a patient with a mild form of multiple acyl CoA dehydrogenase (MAD) deficiency all patient samples were clearly abnormal under a wide variety of clinical conditions, illustrating the high sensitivity and specificity of the method. PMID- 10709661 TI - Genetics of carnitine palmitoyltransferase II deficiencies. PMID- 10709662 TI - Identification of a missense mutation in a patient with lethal carnitine acyl carnitine carrier deficiency. PMID- 10709663 TI - MCAD deficiency. Acylcarnitines (AC) by tandem mass spectrometry (MS-MS) are useful to monitor dietary treatment. PMID- 10709664 TI - D-hydroxyacyl-CoA dehydrogenase deficiency. Identification of a new peroxisomal disorder with implications for other disorders of beta-oxidation. AB - The second and third steps of peroxisomal beta-oxidation are catalysed by two multifunctional enzymes: D-bifunctional protein and L-bifunctional protein. Here we show that fibroblasts of a patient described as being deficient in the 3 hydroxyacyl-CoA dehydrogenase component of D-bifunctional protein and fibroblasts of a patient described as being deficient in L-bifunctional protein do not complement one another. Using a newly developed method to measure the activity of D-bifunctional protein in fibroblast homogenates, we found that the activity of the D-bifunctional protein was completely deficient in the patient with presumed L-bifunctional protein deficiency. PMID- 10709665 TI - Phytanoyl-CoA hydroxylase deficiency. Enzymological and molecular basis of classical Refsum disease. PMID- 10709666 TI - Rationale for a conditional knockout mouse model to study carnitine palmitoyltransferase I deficiencies. AB - Several severe congenital cardiomyopathies are known to be associated with deficiencies in long-chain fatty acid transport and oxidation. Our studies are focused on a key enzyme in the regulation of intracellular long-chain fatty acid transport: carnitine palmitoyltransferase 1. Of this enzyme, two isoforms are expressed in the neonatal heart: L-CPT1 (the "liver-type" isoform) and M-CPT1 (the "muscle-type" isoform). It is known from studies in rats that chemical inhibition of both CPT1 isoforms results in hypertrophy of the cardiomyocytes, leading to an increase in heart-weight of up to 25%. With the aid of expressed sequence tag database analyses, cDNA- and genomic sequence information, we analysed the human gene for M-CPT1 in detail, and obtained partial clones of the murine genes for both CPT1 isoforms. We now started the development of a conditional knockout model to analyse and dissect deficiencies in these genes. While of the other mitochondrial components of the carnitine system deficiencies are known, some with severe cardiac consequences, M-CPT1 deficiencies have never been described. This suggests that M-CPT1 deficiency either (1) has not been recognised within the pool of congenital disorders, (2) is detrimental in an early stage of reproduction or embryogenesis, or (3) does not lead to physiological problems, probably due to the existence of a rescue system. If (1) is the case, the phenotypic effects of M-CPT1 deficiency have to be studied in order to generate criteria for clinical decision making and diagnosis. Option (2) demonstrates the necessity to use novel vector systems to create conditional gene disruptions. Hypothesis (3) implies a possible role for L-CPT1, and a knockout model allows a study of the interaction between the genes for L-CPT1 and M-CPT1. Applicable strategies to develop such a model system will be discussed. PMID- 10709667 TI - Biochemical characterisation of mutations of human medium-chain acyl-CoA dehydrogenase. PMID- 10709668 TI - Lessons learned from the mouse model of short-chain acyl-CoA dehydrogenase deficiency. AB - The SCAD deficient mouse model has been useful to investigate mechanisms of deficient fatty acid oxidation disease in human patients. This mouse model has been thoroughly characterized and is readily available from the Jackson Laboratory. Using the new technologies of gene-knockout mouse modeling, we envisage developing additional members of the acyl-CoA dehydrogenase family of enzyme deficiencies in mice and furthering our understanding of fatty acid metabolism in health and disease. PMID- 10709669 TI - Transcriptional regulation in intestinal development. Implications for colorectal cancer. AB - Deciphering the complex mechanisms of intestinal epithelial development will require multiple cell and molecular approaches in both in vitro and whole animal systems. Additionally, the use of model organisms such as D. melanogaster, C. elegans, and zebrafish will help describe paradigms that may be investigated in mammals as well as serve as test systems for findings from mammals. This manuscript reviewed only one approach to understanding intestinal development. However, the Cdx story and the information to be mined from an understanding of SI gene transcription is not at an end. As the other pieces of the transcriptional puzzle of the SI gene are assembled there will be new information to generate hypotheses on the relationship of transcriptional mechanisms to cancer pathogenesis. PMID- 10709670 TI - Colonic cell proliferation, differentiation, and apoptosis. PMID- 10709671 TI - Defects in the regulation of beta-catenin in colorectal cancer. AB - The molecular events that contribute to the progression of colon cancer are beginning to unravel. An initiating and probably obligatory event is the oncogenic activation of beta-catenin. This can come about by the loss of its negative regulator the adenomatous polyposis coli (APC) protein, or by mutations in the beta-catenin gene that result in a more stable protein product. The interaction between APC and beta-catenin, and additional proteins that affect assembly and signaling along this pathway, are discussed. PMID- 10709672 TI - Dietary lipids, inflammation, and colon cancer. PMID- 10709673 TI - Sulindac sulfone induced regression of rectal polyps in patients with familial adenomatous polyposis. AB - Sulindac sulfone (Exisulind), a metabolite of the non-steroidal anti-inflammatory drug, sulindac, was evalauted for its effects on the development of rectal polyps in patients with familial adenomatous polyposis. Three cohorts of 6 patients each were given doses of 200, 300, or 400 mg Exisulind twice daily. Hepatotoxicity, shown by elevation in blood transaminase levels, was the dose-limiting toxicity and occurred at the 400 mg bid dose. Due to this toxicity, all patients treated with the 400 mg dose were subsequently reduced to the 200 mg dose level. Subsequently, 2 of the 6 patients were dose-escalated to 400 mg bid dose. The patients were treated with Exisulind for a period of six months. Sixteen of 18 patients had regression of small polyps (> or = 6 mm in diameter) characterized by a flattening of the polyps and a macular "halo" appearance. Histopathologic examination of the polyp biopsy specimens showed a marked increase in the proportion of mucin producing cells in the glands after treatment with Exisulind at all dose levels. Ki-67 staining, a measure of cell proliferation, was higher in the polyps than in normal mucosa. There was no significant change in the proliferation index between baseline and six month values in any of the groups treated with Exisulind or in normal tissues. The median apoptotic labeling index, as determined by the TUNEL technique, was higher in the polyps than in normal appearing mucosa. Overall, there was no significant change in the apoptotic labeling index between base-line and 6 months in normal-appearing mucosa however, the index in polyps was increased. These results suggest that treatment of FAP patients with Exisulind for a period of six months may lead to regression of small polyps, and that the mechanisms of Exisulind--induced regression appear to be through stimulation of mucus differentiation and apoptosis in glandular epithelium. PMID- 10709674 TI - Prevention of colon cancer and modulation of aberrant crypt foci, cell proliferation, and apoptosis by retinoids and NSAIDs. AB - The effect of the NSAIDs, retinoids and DFMO on AOM-induced colon tumors, and ACF, cell proliferation, and apoptosis is summarized in Table 1. The ability to prevent AOM-induced ACF has been used as an assay to screen agents for chemoprevention. As discussed above, all six potential chemopreventive agents, aspirin, 2-CPR, DFMO, 4-HPR, piroxicam, and 9-cis-retinoic acid, decreased the level of AOM-induced ACF. However, two of the agents, aspirin (at doses that greatly reduced the yield of ACF) and 2-CPR did not prevent AOM-induced colon tumors. Hence, aspirin and 2-CPR would appear to be false positive in the ACF assay. Besides being a false positive in the ACF assay, 2-CPR actually had the opposite effect of doubling the yield of colon tumor. The false positive result for aspirin could be due to the lower sensitivity of the AOM-induced colon cancer assay compared to the ACF assay. However, aspirin [table: see text] significantly reduced the yield of ACF at a dose (600 mg/kg diet) one-third the dose (1800 mg/kg diet) that did not reproducibly reduce the yield of colon tumors. Thus, although there were no false negative results, two of the six agents gave false positive results in the AOM-induced ACF assay with respect to their ability to prevent colon cancer. Two other potential biomarkers for chemopreventive activity are the ability to reduce the level of cell proliferation and to enhance the level of apoptosis. All six of the agents including aspirin and 2-CPR reduced the level of cell proliferation in adenomas. Thus, similar to their ability to prevent ACF, the ability of aspirin and 2-CPR to decrease cell proliferation were also false positive responses with respect to prevention of colon cancer, but not with respect to the prevention of ACF. Piroxicam, the most potent of the six agents in preventing AOM-induced colon cancer, did not significantly affect the level of cell proliferation in adenomas which is a false negative response. Hence, only three of the six agents (50%) were correctly identified as potential chemopreventive agents by their ability to reduce the level of cell proliferation. In contrast, retinoids, including the three discussed here, demonstrated good correlation between the ability to prevent AOM-induced ACF and the ability to decrease cell proliferation in colonic mucosa or ACF. Thus, within some classes of agents such as the retinoids, the ability to prevent ACF and to reduce cell proliferation appear to correlate, while in other classes including the NSAIDs, the correlation appeared not to exist. The four agents that prevented colon cancer all enhanced the level of apoptosis, while the two agents that did not prevent colon cancer did not effect apoptosis. Three other chemopreventive agents, including phenylethyl-3-methylcaffeate and the NSAIDs, curcumin and sulindac, have been shown by Samaha et al. to enhance apoptosis in AOM-induced colon tumors. Thus, although a very limited number of chemopreventive agents have been evaluated for the ability to enhance apoptosis in the colon, there appears to be an association between the ability to enhance apoptosis and the ability to prevent colon cancer. The use of the AOM-induced ACF assay to screen agents for the ability to prevent colon tumors would appear to result in false positive responses including agents (2-CPR and quercetin) that actually promote colon cancer. However, our results suggest that false positive responders could be distinguished by their inability to enhance apoptosis while potential chemopreventive agents would enhance it. It is therefore proposed that a Two Step Procedure be used to screen agents for the ability to prevent colon cancer. Step 1 would be the determination of the ability to prevent ACF. Because the ACF assay appears to suffer more from false positive than from false negative responders, apparently few potent chemopreventive agents would be missed. Also the ACF assay could be the source of foci for evaluation of the effect PMID- 10709676 TI - Mechanisms by which energy restriction inhibits carcinogenesis. AB - Cancer that occurs at numerous organ sites, including the colon and breast, is inhibited by energy restriction, and the inhibition is proportional to the degree of restriction imposed. In an effort to identify the mechanism(s) by which energy restriction exerts this effect, a short term model system of experimentally induced mammary carcinogenesis was used. Given that carcinogenesis is known to involve a dysregulation to tissue size homeostasis in which cell proliferation and cell death are in dysequilibrium, we hypothesized that energy restriction exerts its effect by altering one or more aspects of cell cycle regulation. It was observed that energy restriction inhibited cell proliferation and increased cell death due to apoptosis. Thus attention was next focused on aspects of cell cycle regulation that might be affected by energy restriction. It was observed that the amount of p27 protein, one member of the Cip/Kip family of genes that are involved in cell cycle arrest, was increased dose dependently by energy restriction. Based on this and related observations, the hypothesis is advanced that energy restriction inhibits carcinogenesis, at least in part, by delaying cell cycle progression via shifting cell populations into a G(0)/G(1)state. Ongoing work indicates that corticosteroids, which are produced in increased amounts in response to energy restriction, may be involved in mediating this effect. PMID- 10709675 TI - Digested fiber from wheat bran induces cdk inhibitors which block colon epithelial cells in G1. PMID- 10709677 TI - Dietary intervention studies of colorectal cancer. PMID- 10709678 TI - The genetics of hereditary non-polyposis colorectal cancer and non-polypotic colon cancer. PMID- 10709679 TI - Familial association. AB - Familial risk of colon cancer is a commonly encountered issue, involving both rare inherited syndromes of colon cancer and common familial clustering of cases. The genes that give rise to the rare syndromes of FAP and HNPCC are now known and current research is addressing the cellular mechanisms of these genes and the proper application of genetic testing in families with one of the syndromes. Common familial clustering of cases appears to arise from an interaction of mildly to moderately severe inherited susceptibility factors with certain environmental factors to give rise to adenomatous polyps and then finally colorectal cancer. Research in this area involves identification of these purportedly more common susceptibility genes, and determination of how each interacts with environmental factors to give rise to polyps and cancer. Optimal application of varying degrees of familial risk to screening strategies is also being determined. PMID- 10709680 TI - Colonic cell proliferation and apoptosis in rodent species. Modulation by diet. PMID- 10709681 TI - Nonsteroidal anti-inflammatory drugs (NSAIDs), cyclooxygenases, and the cell cycle. Their interactions in colon cancer. PMID- 10709682 TI - Clinical trials of pacing mode selection. AB - Current recommendations in favor of dual-chamber over single-chamber ventricular pacing for patients with sinus node dysfunction or AV conduction disorders were made largely based on observational data and expert opinions. The first randomized pacing mode selection study was relatively small and suggested survival advantage with physiologic pacing only after an extended follow-up duration of 5.5 years. Preliminary results of the first large-scale multicenter randomized pacing mode selection trial revealed only modest reduction in atrial fibrillation without survival advantage after 3 years of physiologic pacing. Two other large-scale multicenter randomized trials comparing physiologic versus ventricular pacing are currently ongoing. They may provide further scientific evidence based on which more objective recommendations can be made with respect to pacing mode selection. PMID- 10709683 TI - Pacing to prevent atrial fibrillation. AB - Present therapies for the treatment of atrial fibrillation (AF) are often ineffective or not well-tolerated. Atrial-based pacing reduces the development of AF in the general pacemaker population. Presently, a number of studies are exploring the role of atrial pacing for prevention of AF. This article reviews the rationale for atrial pacing as a treatment for AF and the results of recent studies evaluating atrial pacing for prevention of AF. PMID- 10709684 TI - Management of atrial tachyarrhythmias in patients with implantable devices. AB - The incidence of atrial fibrillation in patients with conduction system disease is high and the management of patients with pacemakers and atrial fibrillation is discussed. The use of mode switch algorithms to avoid tracking of atrial arrhythmias is explained in detail and programming and evaluation of different mode switch algorithms is presented. PMID- 10709685 TI - Pacing for patients with congestive heart failure and dilated cardiomyopathy. AB - Considerable evidence has now accumulated that permanent pacing may provide symptomatic benefit for at least some patients with CHF. Recently, the most promising results with left ventricular or biventricular pacing have been obtained. The data for improvement in survival with pacing is less compelling. The mortality of CHF associated with systolic dysfunction of the left ventricle remains high and arrhythmic deaths are frequent. Clinical trials such as the Sudden Cardiac Death Heart Failure Trial (SCD-HeFT) are currently underway to investigate the role of the implantable defibrillator in patients with heart failure. The development and general availability of ICDs with biventricular pacing capability may play an increasingly important role in the overall therapeutic plan for this group of patients to allow for optimization of functional status with pacing and protection from sudden cardiac death with defibrillation. PMID- 10709686 TI - Pacing in hypertrophic cardiomyopathy. AB - Dual chamber pacing has been proposed as an alternative to surgery in the management of hypertrophic cardiomyopathy. Reports have documented hemodynamic and symptomatic benefit from dual chamber pacing, raising the question of whether or not all patients with drug-refractory symptoms should undergo a trial of pacing before consideration of surgery. The enthusiasm for pacing in hypertrophic cardiomyopathy has generated a number of investigations addressing this issue, including several recently concluded clinical trials. This article reviews the recent experience with dual chamber pacing in hypertrophic cardiomyopathy. PMID- 10709687 TI - Pacing to prevent vasovagal syncope. AB - Patients with frequent vasovagal syncope have markedly poor quality of life and are often resistant to treatment with standard pharmacologic approaches. Vasovagal syncope is due to combinations of bradycardia and hypotension. There is accumulating evidence that many of these patients may respond to permanent cardiac pacing. Several controlled open-label studies suggest that about half of paced patients no longer faint, and most of the rest are improved. At this point, we do not know the role of placebo, and specific pacing modes in this improvement are not known. Ongoing trials will clarify how to select patients and how best to pace them. PMID- 10709688 TI - Cardiac pacing leads. AB - Many of the advances that have been seen in the last decade concerning the functionality, size, and longevity of cardiac pacemakers have been dependent upon concomitant advances in cardiac pacing leads. The most difficult component of a pacing lead to develop has been the insulator. There are many choices for physicians implanting pacing leads: active versus passive fixation, standard impedance versus high impedance and polyurethane versus silicone. The current state of affairs of cardiac pacing leads is quite good in that we have leads that have excellent electrical properties and appear to be more resistant to the hostile environment into which the lead is placed. In spite of this, the goal of a perfect lead remains elusive and there continues to be many challenges in lead design. PMID- 10709689 TI - Developments in sensor-driven pacing. AB - This article reviews the recent major developments in the field of rate adaptive pacing. Including, the improved instrumentation of existing sensors, the use of multiple sensors to enhance sensor specificity or sensitivity, and the automation of sensor calibration. The physiologic benefits and programming of rate adaptive pacing are reviewed. PMID- 10709690 TI - Programming of sensor driven pacemakers. AB - The chronotropic response is the most important means by which cardiac output is increased and oxygen delivery is maintained in response to increased oxygen consumption during exercise or stress. When the chronotropic response is suboptimal or absent, exercise intolerance results. This condition, called chronotropic incompetence can be treated effectively with a sensor-driven rate responsive pacemaker. The effectiveness of this therapy assumes that the pacemaker is programmed appropriately. This article focuses on the programming of sensor-driven pacemakers and provides additional suggestions for follow-up testing to ensure maximal benefit from these devices. PMID- 10709691 TI - Automated pacemaker function. AB - Pacemakers have used automated functions since the introduction of the inhibited mode more than 30 years ago. Currently, virtually all aspects of pacemaker function are subject to automated control, including automated threshold tracking and sensitivity adjustment. These features are designed to enhance patient safety and quality of life, extend battery life, and simplify pacemaker programming and follow-up for health care providers. Many of these automated algorithms are still in evolution and the clinical benefits are not clearly demonstrated for all functions. Although pacemaker function will become increasingly automated, these features should not be accepted uncritically and without demonstrating benefit to the pacemaker patient. PMID- 10709692 TI - Current concepts in extraction of transvenous pacing and ICD leads. AB - Extraction of chronically implanted pacing leads involves a thorough understanding of the pathophysiology of lead maturation and the problems that may occur. It also requires specific knowledge of lead construction and the idiosyncrasies of individual lead models. Though we have evolved to use a standardized approach to lead extraction, each patient and lead removal is unique. The operator must be ready to adapt the technique and tools used to the situation at hand. The more experience and the more tools available to the operator, the more likely that there will be a safe and successful outcome to the procedure. Preparation for disaster is mandatory, such that when a complication does occur, one may respond quickly and therefore salvage the patient. PMID- 10709693 TI - Interference with cardiac pacing. AB - Most exposures to electromagnetic interference are transient and pose no threat to patients with pacemakers and implantable cardioverter defibrillators. Prolonged exposure may be catastrophic in pacemaker dependent patients. New technologies (wireless phones, electronic antitheft surveillance) are safe if proper precautions are takes. Radiofrequency ablation requires concomitant temporary pacing. MR imaging remains contraindicated in patients with these devices until further study is undertaken. PMID- 10709694 TI - Three-dimensional computer vision for tooth restoration. AB - If a person with carious lesions needs or requests crowns or inlays, these dental fillings have to be manufactured for each tooth and each person individually. We survey computer vision techniques which can be used to automate this process. We introduce three particular applications which are concerned with the reconstruction of surface information. The first one aims at building up a database of normalized depth images of posterior teeth and at extracting characteristic features from these images. In the second application, a given occlusal surface of a posterior tooth with a prepared cavity is digitally reconstructed using an intact model tooth from a given database. The calculated surface data can then be used for automatic milling of a dental prosthesis, e.g. from a preshaped ceramic block. In the third application a hand-made provisoric wax inlay or crown can be digitally scanned by a laser sensor and copied three dimensionally into a different material such as ceramic. The results are converted to a format required by the computer-integrated manufacturing (CIM) system for automatic milling. PMID- 10709695 TI - Robust retrieval of three-dimensional structures from image stacks. AB - Robust high-breakdown-point location estimators are employed to analyze image stacks under the piecewise constant image structure model. To reduce the effect of bias along the Z-axis, the class parameters are extracted using three consecutive slices, The segmentation algorithm first determines the most reliable seed regions, which are then used in a region-growing procedure supported by local evidence. The robustness and stability of the proposed technique is shown with both synthetic and real data, the latter consisting of one MRI and one confocal microscopy set. The performance of the algorithm is consistent with the ground truth obtained with manual segmentation by physicians. PMID- 10709696 TI - Model-based detection of spiculated lesions in mammograms. AB - Computer-aided mammographic prompting systems require the reliable detection of a variety of signs of cancer. In this paper we concentrate on the detection of spiculated lesions in mammograms. A spiculated lesion is typically characterized by an abnormal pattern of linear structures and a central mass. Statistical models have been developed to describe and detect both these aspects of spiculated lesions. We describe a generic method of representing patterns of linear structures, which relies on the use of factor analysis to separate the systematic and random aspects of a class of patterns. We model the appearance of central masses using local scale-orientation signatures based on recursive median filtering, approximated using principal-component analysis. For lesions of 16 mm and larger the pattern detection technique results in a sensitivity of 80% at 0.014 false positives per image, whilst the mass detection approach results in a sensitivity 80% at 0.23 false positives per image. Simple combination techniques result in an improved sensitivity and specificity close to that required to improve the performance of a radiologist in a prompting environment. PMID- 10709697 TI - Evaluating a robust contour tracker on echocardiographic sequences. AB - In this paper we present an evaluation of a robust visual image tracker on echocardiographic image sequences. We show how the tracking framework can be customized to define an appropriate shape space that describes heart shape deformations that can be learnt from a training data set. We also investigate energy-based temporal boundary enhancement methods to improve image feature measurement. Results are presented demonstrating real-time tracking on real normal heart motion data sequences and abnormal synthesized and real heart motion data sequences. We conclude by discussing some of our current research efforts. PMID- 10709698 TI - Knowledge-based tensor anisotropic diffusion of cardiac magnetic resonance images. AB - We present a general formulation for a new knowledge-based approach to anisotropic diffusion of multi-valued and multi-dimensional images, with an illustrative application for the enhancement and segmentation of cardiac magnetic resonance (MR) images. In the proposed method all available information is incorporated through a new definition of the conductance function which differs from previous approaches in two aspects. First, we model the conductance as an explicit function of time and position, and not only of the differential structure of the image data. Inherent properties of the system (such as geometrical features or non-homogeneous data sampling) can therefore be taken into account by allowing the conductance function to vary depending on the location in the spatial and temporal coordinate space. Secondly, by defining the conductance as a second-rank tensor, the non-homogeneous diffusion equation gains a truly anisotropic character which is essential to emulate and handle certain aspects of complex data systems. The method presented is suitable for image enhancement and segmentation of single- or multi-valued images. We demonstrate the efficiency of the proposed framework by applying it to anatomical and velocity-encoded cine volumetric (4-D) MR images of the left ventricle. Spatial and temporal a priori knowledge about the shape and dynamics of the heart is incorporated into the diffusion process. We compare our results to those obtained with other diffusion schemes and exhibit the improvement in regions of the image with low contrast and low signal-to-noise ratio. PMID- 10709699 TI - The development and evaluation of CADMIUM: a prototype system to assist in the interpretation of mammograms. AB - We have developed CADMIUM, a novel approach for the design of systems to assist in the interpretation of medical images. CADMIUM uses symbolic reasoning to relate information obtained from image processing to the decisions radiologists take. The approach is based on a symbolic decision procedure which has already been used successfully in a variety of nonimaging clinical decision systems. In CADMIUM this decision procedure is extended with models of three generic image interpretation tasks: detection, measurement and classification of image features. The extended procedure is used to construct the lines of reasoning needed in each task and to control the acquisition of information by image processing. CADMIUM has been evaluated as an aid to the differential diagnosis of microcalcifications on mammographic images. Radiographers who had been trained to interpret images performed better when using the advice provided by the system. PMID- 10709700 TI - A comparison of freehand three-dimensional ultrasound reconstruction techniques. AB - Three-dimensional freehand ultrasound imaging produces a set of irregularly spaced B-scans, which are typically reconstructed on a regular grid for visualization and data analysis. Most standard reconstruction algorithms are designed to minimize computational requirements and do not exploit the underlying shape of the data. We investigate whether an approximation with splines holds any promise as a better reconstruction method. A radial basis function approximation method is implemented and compared with three standard methods. While the radial basis approach is computationally expensive, it produces accurate reconstructions without the kind of visible artefacts common with the standard methods. The other potential advantages of radial basis functions, such as the direct computation of derivatives, make further investigation worthwhile. PMID- 10709701 TI - Model tags: direct three-dimensional tracking of heart wall motion from tagged magnetic resonance images. AB - Although magnetic resonance tissue tagging is a useful tool for the non-invasive measurement of three-dimensional (3-D) heart wall motion, the clinical utility of current analysis techniques is limited by the prohibitively long time required for image analysis. A method was therefore developed for the reconstruction of 3 D heart wall motion directly from tagged magnetic resonance images, without prior identification of ventricular boundaries or tag stripe locations. The method utilized a finite-element model to describe the shape and motion of the heart. Initially, the model geometry was determined at the time of tag creation by fitting a small number of guide points which were placed interactively on the images. Model tags were then created within the model as material surfaces which defined the location of the magnetic tags. An objective function was derived to measure the degree of match between the model tags and the image stripes. The objective was minimized by allowing the model to deform directly under the influence of the images, utilizing an efficient method for calculating image derived motion constraints. The model deformation could also be manipulated interactively by guide points. Experiments were performed using clinical images of a normal volunteer, as well as simulated images in which the true motion was specified. The root-mean-squared errors between the known and calculated displacement and strain for the simulated images were similar to those obtained using previous stripe-tracking and model-fitting methods. A significant improvement in analysis time was obtained for the normal volunteer and further improvements may allow the method to be applied in a 'real-time' clinical environment. PMID- 10709702 TI - Comparative evaluation of multiresolution optimization strategies for multimodality image registration by maximization of mutual information. AB - Maximization of mutual information of voxel intensities has been demonstrated to be a very powerful criterion for three-dimensional medical image registration, allowing robust and accurate fully automated affine registration of multimodal images in a variety of applications, without the need for segmentation or other preprocessing of the images. In this paper, we investigate the performance of various optimization methods and multiresolution strategies for maximization of mutual information, aiming at increasing registration speed when matching large high-resolution images. We show that mutual information is a continuous function of the affine registration parameters when appropriate interpolation is used and we derive analytic expressions of its derivatives that allow numerically exact evaluation of its gradient. Various multiresolution gradient- and non-gradient based optimization strategies, such as Powell, simplex, steepest-descent, conjugate-gradient, quasi-Newton and Levenberg-Marquardt methods, are evaluated for registration of computed tomography (CT) and magnetic resonance images of the brain. Speed-ups of a factor of 3 on average compared to Powell's method at full resolution are achieved with similar precision and without a loss of robustness with the simplex, conjugate-gradient and Levenberg-Marquardt method using a two level multiresolution scheme. Large data sets such as 256(2) x 128 MR and 512(2) x 48 CT images can be registered with subvoxel precision in <5 min CPU time on current workstations. PMID- 10709703 TI - Model extraction from magnetic resonance volume data using the deformable pyramid. AB - A general framework for automatic model extraction from magnetic resonance (MR) images is described. The framework is based on a two-stage algorithm. In the first stage, a geometrical and topological multiresolution prior model is constructed. It is based on a pyramid of graphs. In the second stage, a matching algorithm is described. This algorithm is used to deform the prior pyramid in a constrained manner. The topological and the main geometrical properties of the model are preserved, and at the same time, the model adapts itself to the input data. We show that it performs a fast and robust model extraction from image data containing unstructured information and noise. The efficiency of the deformable pyramid is illustrated on a synthetic image. Several examples of the method applied to MR volumes are also represented. PMID- 10709704 TI - A new variational shape-from-orientation approach to correcting intensity inhomogeneities in magnetic resonance images. AB - A new intensity inhomogeneity correction algorithm based on a variational shape from-orientation formulation is presented. Unlike most previous methods, the proposed algorithm is fully automatic, widely applicable and very efficient. Since no prior classification knowledge about the image is assumed in the proposed algorithm, it can be applied to correct intensity inhomogeneities for a wide variety of medical images. In this paper, a finite-element method is used to model the smooth bias-field function. Orientation constraints for the bias-field function are computed at the nodal locations of the regular discretization grid away from the boundary between different class regions. The selection of reliable orientation constraints is facilitated by the goodness of fit of a first-order polynomial model to the neighborhood of each nodal location. The automatically selected orientation constraints are integrated in a regularization framework, which leads to minimization of a convex and quadratic energy function. This energy minimization is accomplished by solving a linear system with a large, sparse, symmetric and positive semi-definite stiffness matrix. We employ an adaptive preconditioned conjugate-gradient algorithm to solve the linear system very efficiently. Experimental results on a variety of magnetic resonance images are given to demonstrate the effectiveness and efficiency of the proposed algorithm. PMID- 10709705 TI - Rigid point feature registration using mutual information. AB - We have developed a new mutual information-based registration method for matching unlabeled point features. In contrast to earlier mutual information-based registration methods, which estimate the mutual information using image intensity information, our approach uses the point feature location information. A novel aspect of our approach is the emergence of correspondence (between the two sets of features) as a natural by-product of joint density estimation. We have applied this algorithm to the problem of geometric alignment of primate autoradiographs. We also present preliminary results on three-dimensional robust matching of sulci derived from anatomical magnetic resonance images. Finally, we present an experimental comparison between the mutual information approach and other recent approaches which explicitly parameterize feature correspondence. PMID- 10709706 TI - Are routine arrhythmia inductions necessary in patients with pectoral implantable cardioverter defibrillators? AB - INTRODUCTION: The value of ventricular arrhythmia inductions as part of routine implantable cardioverter defibrillator (ICD) follow-up in new-generation pectoral ICDs is unknown. METHODS AND RESULTS: We performed a retrospective analysis of a prospectively collected database analyzing data from 153 patients with pectoral ICDs who had routine arrhythmia inductions at predismissal, and 3 months and 1 year after implantation. Routine predismissal ventricular fibrillation (VF) induction yielded important findings in 8.8% of patients, all in patients with implantation defibrillation threshold (DFT) > or = 15 J or with concomitant pacemaker systems. At 3 months and 1 year, routine VF induction yielded important findings in 5.9% and 3.8% of tested patients, respectively, all in patients who had high DFT on prior testing. Ventricular tachycardia (VT) induction at predismissal, and 3 months and 1 year after implantation resulted in programming change in 37.4%, 28.1%, and 13.8% of tested patients, almost all in patients with inducible VT on baseline electrophysiologic study and clinical episodes since implantation. CONCLUSION: Although helpful in identifying potentially important ICD malfunctions, routine arrhythmia inductions during the first year after ICD implantation may not be necessary in all cases. VF inductions have a low yield in patients with previously low DFTs who lack concomitant pacemakers. VT inductions have a low yield in patients without baseline inducible VT and in the absence of clinical events. Definite recommendations regarding patient selection must await larger prospective studies as well as consensus in the medical community about what comprises an acceptable risk justifying avoidance of the costs and inconveniences of routine arrhythmia inductions. PMID- 10709707 TI - Ablation of atypical atrial flutter guided by the use of concealed entrainment in patients without prior cardiac surgery. AB - INTRODUCTION: Mapping techniques have not been systematically evaluated with respect to atypical atrial flutter (AF) not involving the inferior vena cava isthmus. The purpose of this study was to assess prospectively the use of concealed entrainment (CE) in mapping of AF and to assess the clinical benefit of ablation of clinically relevant atypical AF. METHODS AND RESULTS: In seven consecutive patients without prior cardiac surgery presenting with atypical AF, mapping was performed in the right and, if necessary, left atrium. At sites with CE, radiofrequency energy was delivered. In a posthoc analysis, the endocardial activation time, stimulus-flutter wave (F) interval, presence of split potentials and diastolic potentials, and postpacing interval were assessed, and effective sites were compared to ineffective sites. A total of 22 forms of atypical AF either could be induced or were present at the time of the study. Eleven of the 13 targeted atypical AFs (85%) were successfully ablated. The positive predictive value of CE increased from 45% to 75% in the presence of matching electrogram-F and stimulus-F intervals or if flutter terminated during entrainment pacing, and to 88% in the presence of split atrial electrograms or diastolic potentials. During short-term clinical follow-up, none of the patients had recurrence of the ablated AF. However, the majority of patients required either medication for atrial fibrillation or repeated interventions for new forms of AF. CONCLUSION: Mapping and ablation of atypical AF is feasible if sites with CE can be identified. However, the clinical benefit of successful ablations in patients with atypical flutter appears to be limited. PMID- 10709708 TI - Unipolar electrogram characteristics predictive of successful radiofrequency catheter ablation of accessory pathways. AB - INTRODUCTION: The purpose of this study was to determine the characteristics of the unipolar electrogram that are most helpful in predicting successful radiofrequency ablation of accessory pathways. METHODS AND RESULTS: The unipolar electrogram was analyzed at 185 ablation sites in 53 patients; 94 attempts were directed at the site of earliest atrial activation ("atrial group") and 91 at the site of earliest ventricular activation ("ventricular group"). The electrogram was analyzed for several features, including pattern ("QS" or "initial R"). Unipolar pattern: Overall, a "QS" pattern was seen at 55% of unsuccessful, 75% of temporarily successful, and 90% of permanently successful sites. For the atrial group, the respective frequencies were 53%, 77%, and 92%, and for the ventricular group, 57%, 73%, and 86%. The difference in pattern distribution between unsuccessful and permanently successful sites was significant for all groups: overall, P < 0.0001; atrial group, P = 0.0005; ventricular group, P = 0.02. Absence of a "QS" pattern (i.e., "initial R") predicted a 92% chance of unsuccessful ablation. Additional features: Activation times were significantly shorter at permanently successful than at unsuccessful (P < 0.0001) or temporarily successful sites (P = 0.0002). No significant differences were found in atrial or ventricular amplitudes or in A/V ratios. Intrinsic deflection slew was lower at temporarily successful sites (P = 0.03 vs all other sites). CONCLUSION: Ablation at sites revealing an "initial R" pattern (i.e., absent "QS") is very unlikely to be successful. Activation time is shorter at successful sites. These features are equally applicable when mapping the atrial potential as when mapping the ventricular potential. PMID- 10709709 TI - Comparison of endocardial activation times at effective and ineffective ablation sites within the pulmonary veins. AB - INTRODUCTION: Recent studies demonstrated that atrial arrhythmias may be generated within pulmonary veins. The purpose of this study was to compare the endocardial activation times at effective and ineffective ablation sites during radiofrequency catheter ablation of arrhythmias initiated or generated within pulmonary veins. METHODS AND RESULTS: Twenty-one of 28 patients without structural heart disease underwent successful ablation of 23 arrhythmogenic foci within a pulmonary vein. Electrograms were recorded at 75 pulmonary venous sites and categorized into three groups: 23 successful ablation sites; 28 unsuccessful target sites within an arrhythmogenic pulmonary vein; and 24 sites within nonarrhythmogenic pulmonary veins. The endocardial activation time of premature depolarizations arising at successful target sites was significantly earlier than at other sites. During premature depolarizations, an endocardial activation time of -75 msec or earlier had a sensitivity of 83% and a specificity of 79% for identification of a successful ablation site. Endocardial activation times earlier than -100 msec were recorded only at successful ablation sites, and endocardial activation times later than -30 msec were recorded only at sites within nonarrhythmogenic pulmonary veins. The presence of a split potential during sinus rhythm or premature depolarizations was not a specific indicator of a successful ablation site. CONCLUSION: The endocardial activation times of premature depolarizations that arise within pulmonary veins and initiate atrial tachycardia/fibrillation are useful in identifying successful ablation sites. PMID- 10709710 TI - Signal-averaged isoharmonic body surface maps of patients with ischemic cardiomyopathy. AB - INTRODUCTION: Prevention of sudden arrhythmic cardiac death depends on accurate identification of individuals at high risk. Previous studies of signals recorded directly from arrhythmogenic tissue suggested that the predictive value of the signal-averaged ECG could be enhanced by expanded temporal, spectral, and spatial analysis. Accordingly, we performed a prospective study of 192-lead signal averaged body surface maps from 43 patients with ischemic cardiomyopathy referred for electrophysiologic study. Three groups were included: 15 patients with clinical ventricular tachycardia (VT), 12 patients with inducible VT, and 16 patients with non-VT. METHODS AND RESULTS: The patients were well matched with regard to age, gender, infarct location, ejection fraction (28% +/- 9%), QRS duration, and incidence of nonsustained VT (96%). Isoharmonic maps of the entire cardiac cycle were constructed for each patient. The peaks of the 1-7 Hz isoharmonic maps distinguished patients with clinical VT from non-VT and inducible VT patients (1,152 +/- 273, 852 +/- 283, and 808 +/- 272, respectively; P = 0.003). After prospective observation for 22 +/- 16 months, the combined endpoint of spontaneous sustained VT, ventricular fibrillation, appropriate defibrillator therapy, and death was predicted by inducibility of VT (relative risk 3.8, P = 0.008) and by the signal-averaged isoharmonic body surface map (relative risk 3.1, P = 0.02). CONCLUSION: These results confirm the diagnostic utility of signal-averaged isoharmonic body surface maps in a rigorously defined patient population. PMID- 10709711 TI - Direct discrimination and full-day disclosure of ventricular parasystole on new heart rate tachograms. AB - INTRODUCTION: To discriminate ventricular parasystole from fixed coupling interval ventricular premature complexes (VPCs), we developed a new diagnostic method using a dot distribution pattern corresponding to VPCs recorded on a heart rate tachogram using ambulatory ECG monitoring data. We tested our hypothesis that widely scattered VPC dots on instantaneous heart rate tachograms indicate a constant VPC-VPC interval compatible with parasystole. METHODS AND RESULTS: Patients with frequent VPCs > 5,000/day) were divided into two groups depending on the tachogram dot distribution patterns: group S (n = 10, aged 61 +/- 16 years) showed widely scattered VPC dot distribution, whereas group F (n = 10, 60 +/- 17 years) showed fixed VPC dot distribution limited to a narrow zone. Using digitized R-R interval data, full-day heart rate tachograms and VPC-VPC intervals were depicted simultaneously. Group S demonstrated constant basic VPC-VPC intervals (1,285 to 2,052 msec, mean 1,738 +/- 219), with a coefficient of variation (CV) of 0.061 +/- 0.018. Their VPC coupling intervals were markedly variable (651 +/- 113 msec; CV = 0.193 +/- 0.034). Each patient's basic VPC-VPC intervals showed small diurnal alterations (minimum -13% +/- 3% to maximum +15% +/- 6%). VPC-VPC intervals in group F were not constant and showed marked variation. Group F VPC coupling intervals were shorter and constant (480 +/- 30 msec, P = 0.0002; with CV = 0.076 +/- 0.013, P < 0.0001). CONCLUSION: Ventricular parasystole with constant VPC-VPC intervals consistently became evident based on VPC dot patterns recorded on heart rate tachograms. PMID- 10709712 TI - Parasystolic alienation: an impression or a reality? PMID- 10709713 TI - Altered transient outward current in human atrial myocytes of patients with reduced left ventricular function. AB - INTRODUCTION: Electrophysiologic remodeling is involved in the self-perpetuation of atrial fibrillation. To define whether differences in atrial electrophysiology already are present in patients with increased susceptibility for atrial fibrillation, we compared patients in sinus rhythm with and without heart failure. METHODS AND RESULTS: Atrial specimens were obtained from patients with reduced left ventricular ejection fraction (LVEF; n = 10) and normal LVEF (n = 16) who were undergoing aortocoronary bypass surgery and from donor hearts (n = 4). Enzymatically isolated atrial myocytes were investigated by whole cell, patch clamp techniques. Total outward current was significantly larger in myocytes of hearts with low LVEF than normal LVEF (19.4 +/- 1.3 vs 15.1 +/- 1.2 pA/pF at pulses to +60 mV, respectively). Analysis of inactivation time courses of different outward current components revealed that the observed current difference is due to the transient calcium-independent outward current I(to1) which is twice as large in the low LVEF group than in the normal LVEF group (9.4 +/- 0.9 vs 4.7 +/- 0.4 pA/pF at pulses to +60 mV, respectively). I(to1) recovery from inactivation was significantly more rapid in myocytes of hearts with low LVEF, and action potential plateau in these cells was significantly shorter. The results of I(to1) and action potential measurements in atrial myocytes of donor hearts were very similar to the results of patients with preserved heart function. CONCLUSION: I(to1) in human atrial myocytes of patients with reduced LVEF has an increased density and altered kinetics in sinus rhythm. These differences in outward current may explain the reduced plateau phase of action potentials. PMID- 10709714 TI - Microwave ablation using a spiral antenna design in a porcine thigh muscle preparation: in vivo assessment of temperature profile and lesion geometry. AB - INTRODUCTION: Theoretical studies have suggested that microwave energy can increase the depth of heating compared with radiofrequency energy. A spiral microwave antenna design may have advantages over previous designs using smaller designs because the resulting power deposition pattern is considerably larger than the catheter diameter. We tested the efficacy of a spiral antenna using microwave energy in a porcine thigh muscle preparation. METHODS AND RESULTS: In five anesthetized pigs, the thigh muscle was exposed and bathed in heparinized bovine blood (36 degrees to 37 degrees C). A helical microwave catheter with a fiberoptic thermometer attached to the distal end was positioned perpendicular to the thigh muscle. The antenna-tissue interface and tissue temperatures at depths of 3.0 and 6.0 mm were measured. A 915-MHz microwave generator delivered energy at one of three power outputs (50, 100, or 150 W) for 60 seconds. Seventy lesions were created: 50 W (n = 23), 100 W (n = 24), and 150 W (n = 23). The mean depths at 50, 100, and 150 W were 4.3 +/- 1.8 mm, 7.2 +/- 1.7 mm, and 9.4 +/- 0.9 mm, respectively. Lesion depth (R = 0.96, P = 0.05), maximum surface dimension (R = 0.99, P = 0.06), and volume (R = 0.99, P = 0.04) were closely correlated to the power applied. CONCLUSION: Power is an important determinant of lesion size using a spiral microwave antenna. A novel, spiral microwave antenna design can create lesions of significant depth that may be applicable for the ablative therapy of ventricular tachycardia. PMID- 10709715 TI - Pacing in right ventricular dysplasia after disconnection surgery. AB - This report describes a 33-year-old patient with arrhythmogenic right ventricular (RV) dysplasia who had a dual chamber pacemaker implanted at age 23 years for drug-induced bradycardia. Pacing was continued after right ventricular free-wall disconnection (RVFWD) at age 24 years. Her pacemaker was not replaced after battery depletion 7 years later. She presented the following year in severe right sided heart failure. Her old pacemaker generator was replaced. This was followed by rapid resolution of her clinical failure and return to a full, active, physical lifestyle. This observation suggests the potential benefit of dual chamber pacing in patients with RV dysplasia after RVFWD. PMID- 10709716 TI - Radiofrequency catheter ablation of upper septal idiopathic left ventricular tachycardia exhibiting left bundle branch block morphology. AB - Idiopathic left ventricular (LV) tachycardia usually exhibits right bundle branch block morphology. There are only a few sporadic cases that exhibit left bundle branch block (LBBB) morphology. We report a patient whose QRS complex during ventricular tachycardia (VT) was relatively narrow (100 msec) and exhibited LBBB (precordial R wave transition between V3 and V4) and a normal frontal plane axis. This VT was ablated successfully by radiofrequency current applied to the LV upper septum, where the earliest endocardial activation was recorded. PMID- 10709717 TI - Nonsurgical transthoracic mapping and ablation in a child with incessant ventricular tachycardia. AB - We report the case of an 11-month-old child with incessant ventricular tachycardia who underwent two unsuccessful endocardial ablations with standard catheters and in whom the ventricular tachycardia was interrupted only during transthoracic epicardial catheter ablation. This report outlines the usefulness and safety of this novel approach in pediatric patients before surgery when endocardial ablation fails. PMID- 10709718 TI - Electrophysiologic characteristics and radiofrequency ablation of concealed nodofascicular and left anterograde atriofascicular pathways. AB - INTRODUCTION: True nodoventricular or nodofascicular pathways and left-sided anterograde decremental accessory pathways (APs) are considered rare findings. METHODS AND RESULTS: Two unusual patients with paroxysmal supraventricular tachycardia were referred for radiofrequency (RF) ablation. Both patients had evidence of dual AV nodal conduction. In case 1, programmed atrial and ventricular stimulation induced regular tachycardia with a narrow QRS complex or episodes of right and left bundle branch block not altering the tachycardia cycle length and long concentric ventriculoatrial (VA) conduction. Ventricular extrastimuli elicited during His-bundle refractoriness resulted in tachycardia termination. During the tachycardia, both the ventricles and the distal right bundle were not part of the reentrant circuit. These findings were consistent with a concealed nodofascicular pathway. RF ablation in the right atrial mid septal region with the earliest atrial activation preceded by a possible AP potential resulted in tachycardia termination and elimination of VA conduction. In case 2, antidromic reciprocating tachycardia of a right bundle branch block pattern was considered to involve an anterograde left posteroseptal atriofascicular pathway. For this pathway, decremental conduction properties as typically observed for right atriofascicular pathways could be demonstrated. During atrial stimulation and tachycardia, a discrete AP potential was recorded at the atrial and ventricular insertion sites and along the AP. Mechanical conduction block of the AP was reproducibly induced at the annular level and at the distal insertion site. Successful RF ablation was performed at the mitral annulus. CONCLUSION: This report describes two unusual cases consistent with concealed nodofascicular and left anterograde atriofascicular pathways, which were ablated successfully without impairing normal AV conduction system. PMID- 10709719 TI - Radiofrequency catheter ablation of atrial fibrillation initiated by pulmonary vein ectopic beats. AB - Ectopic beats from the pulmonary veins (PVs) have been demonstrated to initiate atrial fibrillation (AF). This article describes the conceptual approach to mapping, interpretation of different electrograms, and ablation of AF initiated by PV ectopic beats. PMID- 10709720 TI - Diversity of gap junctional proteins: does it play a role in cardiac excitation? PMID- 10709721 TI - Palpitations and near-syncope in a 34-year-old man: what is the tachycardia? PMID- 10709722 TI - Atrial tachycardia and "kissing catheters". PMID- 10709723 TI - The chaos about heart rate chaos. PMID- 10709724 TI - Cholesterol synthesis and skeletal formation. PMID- 10709725 TI - MRI predicts neurobehavioral outcome in the very premature infant. PMID- 10709726 TI - Antenatal glucocorticoids and programming of the developing CNS. AB - Glucocorticoids (GCs) are essential for many aspects of normal brain development. However, there is growing evidence from a number of species that exposure of the fetal brain to excess GC, at critical stages of development, can have life-long effects on behavior and neuroendocrine function. The hypothalamo-pituitary adrenal axis, which is central to the integration of the individual's endocrine and behavioral response to stress, appears highly sensitive to excess GC exposure during development. A number of animal studies have shown that exposure to synthetic GCs in utero results in adult offspring that exhibit hyperactivity of the hypothalamo-pituitary-adrenal axis. This will have a long-term impact on health, inasmuch as increased life-long exposure to endogenous GC has been linked to the premature onset of diseases associated with aging. The mechanisms involved in the permanent programming of hypothalamo-pituitary-adrenal function and behavior are not well understood. Synthetic GCs are used extensively to promote pulmonary maturation in fetuses at risk of being delivered before term. Therefore, it is important that we understand the potential long-term consequences of prenatal GC exposure on brain development as well as the underlying mechanisms involved. This review will explore the current state of knowledge in this rapidly expanding field. PMID- 10709727 TI - Labor-associated changes in Fas ligand expression and function in human placenta. AB - Fas ligand (FasL)-dependent apoptosis has been implicated in the control of tissue-damaging inflammatory responses in several immune privileged sites. Here, we present data demonstrating that FasL is expressed on human trophoblast cells throughout pregnancy and transduces growth inhibitory/death signals in cells bearing its receptor, Fas (CD95). Immunohistochemical analysis detected FasL positive staining in the trophoblast layer of villi of first- and second trimester and term (no labor) placental tissues, as well as in freshly isolated cytotrophoblasts representing these gestational ages. In contrast, term placental tissues and cytotrophoblasts from labor-associated deliveries exhibited significantly reduced FasL expression, suggesting that parturition altered the characteristics of trophoblast cells. FasL-specific immunoblotting of cytotrophoblast cell lysates further confirmed these results. To assess the functionality of FasL expressed on cytotrophoblasts, we co-cultured these cells with Fas-bearing Jurkat T cells. Cytotrophoblasts from early pregnancy, or term with no labor, significantly inhibited growth in Jurkat cells, evident even at a 1:1 effector:target cell ratio, as determined by the incorporation of [3H]thymidine. Similar results were obtained when a FasL-positive colon carcinoma cell line, SW620, was used in place of cytotrophoblasts. In contrast, term cytotrophoblasts from labor deliveries exhibited poor FasL expression and were quantitatively much less proficient in inhibiting [3H]thymidine incorporation in Jurkat cells. These data indicate that FasL could participate in modulating the inflammatory responses associated with labor and suggest intrinsic molecular differences in the placental microenvironment before and after labor . PMID- 10709728 TI - Glucose metabolism is elevated and vascular resistance and maternofetal transfer is normal in perfused placental cotyledons from severely growth-restricted fetuses. AB - We hypothesized that placental resistance was elevated and transfer reduced in cotyledons from intrauterine growth-restricted (IUGR) fetuses. We perfused 10 cotyledons from term, normally grown fetuses, six from preterm, normally grown fetuses with normal umbilical arterial end-diastolic velocities (EDV), and six from preterm IUGR fetuses (<3rd centile) with absent or reversed umbilical arterial EDV. Perfused cotyledons were pressure-fixed, and villi were observed by scanning electron microscopy. The groups did not differ in fetoplacental resistance at baseline; neither did they differ in the change in resistance that followed the administration of nitroglycerin or angiotensin II. The increase in resistance during hypoxia was similar in the two preterm groups but greater in the term than in the preterm normally grown group (p < 0.05). Groups did not differ in net maternofetal transfer of oxygen or glucose, or in clearance of aminoisobutyric acid or antipyrine. However, glucose consumption was doubled in cotyledons of preterm IUGR versus preterm normally grown fetuses (p < 0.05). Terminal villi of perfused cotyledons from preterm IUGR fetuses displayed less terminal villous branching and budding than preterm controls, as anticipated from previous work. IUGR fetuses with absent or reversed umbilical arterial EDV in vivo may have high placental resistance due to a vasoconstrictive rather than anatomic abnormality and an elevated placental glucose consumption that may impair glucose transfer. PMID- 10709729 TI - Continuing positive secular growth change in The Netherlands 1955-1997. AB - Since 1858, an increase of mean stature has been observed in the Netherlands, reflecting the improving nutritional, hygienic, and health status of the population. In this study, stature, weight, and pubertal development of Dutch youth, derived from four consecutive nationwide cross-sectional growth studies during the past 42 y, are compared to assess the size and rate of the secular growth change. Data on length, height, weight, head circumference, sexual maturation, and demographics of 14,500 boys and girls of Dutch origin in the age range 0-20 y were collected in 1996 and 1997. Growth references for height and weight were constructed with a method that summarizes the distribution by three smooth curves representing skewness (L curve), the median (M curve), and coefficient of variation (S curve). The relationship between height and demographic variables was assessed by multivariate analysis. Reference curves for menarche and secondary sex characteristics were estimated by a generalized additive model using a logit transformation. A positive secular growth change has been present in the past 42 y for children, adolescents, and young adults of Dutch origin, although at a slower rate in the last 17 y. Height differences according to region, educational level of child and parents, and family size have remained. In girls, median age at menarche has decreased by 6 mo during the past four decades to 13.15 y. Environmental conditions have been favorable for many decades in the Netherlands, and the positive secular change in height has not yet come to a halt, in contrast to Scandinavian countries. Main contributors to the increase in height may be improved nutrition, child health, and hygiene, and a reduction of family size. PMID- 10709730 TI - Correlation between electrocardiographic and ultrasonographic time-interval measurements in fetal lamb heart. AB - The objective of this study was to establish the echocardiographic modality that best correlates with electrical events in the fetal heart. No documentation on the relationship between electrical events recorded with a surface ECG and fetal M-mode or Doppler echocardiographic measurements is available. The following ultrasound tracings were recorded simultaneously with a surface ECG on six exteriorized near-term fetal lambs: 1) M-mode echocardiography of atrial and ventricular contractions; and 2) Doppler flow velocity waveforms in the right superior vena cava (SVC) either alone or 3) in association with those of the ascending aorta. In the SVC, the onset of the retrograde A wave and the beginning of the forward wave during ventricular systole were used as markers for the start of the P wave and QRS complex, respectively. For the simultaneous SVC and ascending aorta tracings, the beginnings of the A and of the aortic ejection waves were used as markers. On average, the atrioventricular interval was 84 ms longer than the PR interval with the M-mode, corresponding to an increase of 107%. A similar observation was made for the simultaneous Doppler signals from SVC and ascending aorta, but the difference between the atrioventricular and PR intervals was smaller, averaging 35 ms. When the SVC Doppler was taken alone, no significant difference was found between atrioventricular and ventriculoatrial compared with PR and RP intervals, respectively, and a strong correlation was found between the two methods of measurement, both for the atrioventricular (r = 0.91) and ventriculoatrial (r = 0.89) intervals. Doppler interrogation of the SVC alone and, to a lesser degree, of the SVC and ascending aorta are reliable indirect markers for the timing of electrical events of the fetal lamb heart in sinus rhythm. PMID- 10709731 TI - Heart rate recovery after exercise and cardiac autonomic nervous activity in children. AB - To investigate the difference in heart rate (HR) recovery after exercise between children and young adults, we administered a constant load of light exercise intensity and progressive treadmill exercise tests to nine children (aged 9 to 12 y, group A) and eight young adults (six male and two female, aged 17 to 21 y, group B) who had a history of Kawasaki disease without significant coronary arterial lesions. HR after both exercise protocols was analyzed. The low frequency (LF) and high-frequency (HF) components of HR variability were measured, and LF/HF was calculated (log LF, log HF, log L/H). Arterial baroreflex sensitivity was assessed by the phenylephrine method. There were no differences between groups A and B in resting HR, peak HR, peak oxygen uptake, and decreases in systolic blood pressure during the recovery period. HR 1 and 2 min after peak exercise and 1 min after constant-load exercise was significantly lower in group A than in group B (p < 0.05), and the changes in HR from peak values after both exercise tests were also greater in group A than in group B (p < 0.05-0.01). Although no difference in arterial baroreflex sensitivity was observed, log HF was significantly higher in group A than in group B (p < 0.01), and log L/H was significantly lower in group A than in group B (p < 0.05). The value of log HF correlated inversely with the decrease in HR immediately after both exercise protocols (p < 0.05-0.01). Although log L/H correlated with the decrease in HR after peak exercise (p < 0.05-0.0005), the early decline in HR after constant load exercise did not correlate with log L/H. Arterial baroreflex sensitivity did not correlate with the decrease in HR at any recovery time. These data suggest that the early phase of HR recovery after light to severe exercise is influenced by the cardiac parasympathetic nervous activity at rest and that the greater central cholinergic modulation of HR in children than in young adults may be responsible in part for children's faster HR recovery after exercise. PMID- 10709732 TI - Autoimmune lymphoproliferative syndrome (ALPS) in a child from consanguineous parents: a dominant or recessive disease? AB - Autoimmune lymphoproliferative syndrome (ALPS) is characterized by autoimmune features and lymphoproliferations and is generally caused by defective Fas mediated apoptosis. This report describes a child with clinical features of ALPS without detectable Fas expression on freshly isolated blood leukocytes. Detection of FAS transcripts via real-time quantitative PCR made a severe transcriptional defect unlikely. Sequencing of the FAS gene revealed a 20-nucleotide duplication in the last exon affecting the cytoplasmic signaling domain. The patient was homozygous for this mutation, whereas the consanguineous parents and the siblings were heterozygous. The patient reported here is a human homologue of the Fas-null mouse, inasmuch as she carries an autosomal homozygous mutation in the FAS gene and she shows the severe and accelerated ALPS phenotype. The heterozygous family members did not have the ALPS phenotype, indicating that the disease-causing FAS mutation in this family is autosomal recessive. PMID- 10709733 TI - Evidence of chronic inflammation in morphologically normal small intestine of cystic fibrosis patients. AB - Cystic fibrosis (CF) is caused by mutations in the CF transmembrane conductance regulator gene and characteristically leads to prominent lung and pancreatic malfunctions. Although an inflammatory reaction is normally observed in the CF airways, no studies have been performed to establish whether a chronic inflammatory response is also present in the CF intestine. We have investigated whether immunologic alterations and signs of inflammation are observed in CF small intestine. Fourteen CF, 20 negative, and four disease controls underwent duodenal endoscopy for diagnostic purposes. Two CF patients were rebiopsied, one after 3 mo of an elemental diet and the other after 2 wk of pancreatic enzyme withdrawal. In three CF and 10 controls, in vitro small intestine organ cultures were also performed. Expression of ICAM-1, IL-2 receptor, IL-2, IFN-gamma, CD80, and transferrin receptor was studied by immunohistochemistry before and after in vitro organ culture. In CF small intestine, an increased number of lamina propria mononuclear cells express ICAM-1 [mean 114 (SD 82.8), p < 0.001 versus controls], CD25 [20.2 (18.7), p < 0.01], IL-2 [23.6 (13.7), p < 0.05], and IFN-gamma [19 (15.9), p < 0.05], whereas villus enterocytes highly express transferrin receptor. Reduced expression of immunologic markers was observed after 24 h of in vitro culture in all three CF patients as well as in the patient kept on elemental diet for 3 mo. These results indicate that chronic inflammation is observed in CF duodenum and suggest that the perturbation of local mucosal immune response may contribute to the overall clinical picture in CF patients. PMID- 10709734 TI - A humanized monoclonal antibody against respiratory syncytial virus (palivizumab) inhibits RSV-induced neurogenic-mediated inflammation in rat airways. AB - Respiratory syncytial virus (RSV) is the most important respiratory pathogen in infancy and early childhood and may predispose to subsequent lower respiratory tract illness. Recent data indicate that RSV up-regulates the substance P receptor, making the airways abnormally susceptible to the proinflammatory effects of this peptide released from sensory nerves. The present study was designed to determine whether the administration of RSV antibodies prevents the potentiation of neurogenic inflammation in rat airways. Five days after inoculation, sensory nerve-mediated extravasation of Evans blue-labeled albumin was significantly greater in the airways of RSV-infected rats than in pathogen free controls. Polyclonal immune globulin enriched for RSV-neutralizing antibodies (RSVIG) reduced neurogenic extravasation when injected 24 h before intranasal inoculation of the virus but not when injected before endotracheal inoculation. A humanized MAb against RSV fusion protein (palivizumab) was twice as potent as RSVIG when given before intranasal inoculation and also caused significant inhibition after endotracheal inoculation. Furthermore, palivizumab inhibited neurogenic inflammation in RSV-infected rats when given 72 h after virus inoculation. These data suggest that palivizumab protects the respiratory tract from RSV-induced inflammation when given before or in the early phase of the viral infection. The administration of palivizumab to high-risk infants may limit the severity of the acute airway inflammation and may protect against subsequent lower respiratory tract illness. PMID- 10709735 TI - Activation of human granulocytes by intravenous immunoglobulin preparations is mediated by FcgammaRII and FcgammaRIII receptors. AB - Previous studies from our laboratory have shown that i.v. Ig (IVIG) exposure triggers superoxide anion (O2) generation by and increases bactericidal capacity of human blood granulocytes. However, the molecular interactions between IVIG and granulocytes have not been evaluated before. The objective of this study was to investigate the role of FcgammaRII and FcgammaRIII receptors in the immunomodulatory effects of IVIG concentrates on granulocytes. We found that four different IVIG preparations (concentration range, 1-10 mg/mL) shared the ability to stimulate O2- release in vitro by granulocytes in a dose-dependent manner. Dimers fractionated from IVIG were significantly more potent in inducing activity of the respiratory burst than were monomers. MAb (concentration range, 0.1-10 microg/mL) specific for FcgammaRII and FcgammaRIII receptors inhibited the IVIG induced O2- release, with a more profound inhibitory effect observed with anti FcgammaRIII. These findings suggest the involvement of Fcgamma receptors in triggering O2- release by granulocytes exposed to IVIG. We also report that IVIG added to granulocyte suspensions elicited a rapid and vigorous increase in the concentration of cytosolic free calcium, a finding suggesting direct activation and not priming of granulocytes by IVIG through FcgammaRII and FcgammaRIII receptors. The in vitro effects described here might occur in patients treated with IVIG and may, in part, be responsible for inflammatory reactions evoked by infused Ig concentrates as well as the immunomodulatory effect of Ig in patients with autoimmune and inflammatory diseases. PMID- 10709736 TI - Prenatal exposure to toluene results in abnormal neurogenesis and migration in rat somatosensory cortex. AB - Toluene inhalant abuse during pregnancy may result in growth-retarded microcephalic newborns who subsequently demonstrate developmental impairment. By using a rat model of toluene-abuse embryopathy, we studied the effects of prenatal toluene exposure on the generation and migration of cortical neurons. Dams were exposed by gavage to either corn oil or toluene diluted in corn oil on d 6-21 of gestation. The time of origin of cortical neurons was determined in the mature pups of dams injected with the thymidine analogue 5'-bromodeoxyuridine on 1 d during the period from d 13-21 of gestation. 5'-Bromodeoxyuridine-labeled neurons were identified by immunohistochemistry in a 400-microm-wide column of somatosensory cortex. The brains of the toluene-exposed pups had a significant reduction in the number of neurons within each cortical layer (p < 0.001). Depending on the cortical layer, the generation of neurons in the toluene-exposed pups was delayed by 1 or 2 d. In addition, the brains of the toluene-exposed pups also showed evidence of abnormal neuronal migration. However, there were no differences in either brain weight or body weight between the control and toluene exposed pups. These observations suggest that although prenatal toluene exposure results in abnormal neuronal proliferation and migration, brain weight in the toluene-exposed pups may be preserved by enhanced development of glia or the neuropil. PMID- 10709737 TI - Maturational differences in soluble guanylate cyclase activity in ovine carotid and cerebral arteries. AB - Basal cGMP concentrations are greater in immature than in mature cranial arteries, which may help explain why cerebrovascular resistance is lower in neonates than in adults. The present studies explore the hypothesis that this difference derives from age-related differences in soluble guanylate cyclase (sGC) activity. Maturation depressed (p < 0.01) maximal sGC activity (pmol cGMP/mg/min) in both carotid (from 11.10 +/- 0.50 to 3.60 +/- 0.20) and cerebral (from 3.10 +/- 0.31 to 1.45 +/- 0.08) arteries. Western blot analysis of relative sGC abundance (relative to sGC expression in adult kidney) found that sGC abundance was significantly greater (p < 0.05) in newborn carotid (0.38 +/- 0.04) and cerebral arteries (0.37 +/- 0.06) than in adult arteries (0.25 +/- 0.05 and 0.17 +/- 0.03, respectively). Basal Km values in carotid and cerebral arteries did not differ significantly between newborns (3- to 7-d old) and adults. Activation of sGC with nitrosylated heme significantly reduced Km values 3- to 5 fold in both types of artery and in both age groups. Within artery type, maturation had no significant effect on activated Km. Between artery types, activated Km values were greater (p < 0.05) in cerebral (200 +/- 40 microM) than in carotid (80 +/- 10 microM) arteries. Together, these data suggest that variations in sGC substrate affinity contribute to observed differences in sGC activity between artery types but not those between age groups. In contrast, variations in enzyme abundance, and possibly also enzyme-specific activity, appear responsible for differences in sGC activity associated with both age and artery type. PMID- 10709738 TI - Preservation of cerebrovascular tone and reactivity by sodium channel inhibition in experimental prolonged asphyxia in piglets. AB - Sodium channels using cAMP as a second messenger play a role in the regulation of cerebral circulation and metabolism. Cerebrospinal fluid (CSF) cAMP levels have been shown to correlate with the degree and duration of hypoxic injury and outcome and to be an indicator of cerebral vascular reactivity. We hypothesize that sodium channel inhibition either before or at termination of experimental asphyxia will attenuate cerebrovascular alterations and maintain CSF cAMP levels. Three groups of piglets with closed cranial windows were studied: asphyxia or group 1 (n = 5) and two treatment groups. Pigs were treated with 50 mg/kg of sodium channel blocker before asphyxia (group 2, n = 6) and after the termination of asphyxia and start of reventilation (group 3, n = 6). Asphyxia was sustained over 60 min by ventilating piglets with 10% O2 gas mixture and decreasing minute ventilation followed by 60 min of reventilation with room air. Every 10 min, pial arterial diameters were measured, and CSF samples were collected for cAMP determination. Vascular reactivity to topically applied isoproterenol (10(-4) M) was evaluated 60 min after recovery. During asphyxia, cAMP levels in group 2 peaked and declined at a later time with mean values remaining significantly higher than those of groups 1 and 3. During reventilation, CSF cAMP concentrations were highest in group 3 and lowest in group 1. Pial arteriolar dilation occurred during asphyxia in all three groups but to a lesser degree in the pretreated group compared with groups 1 and 3. Pial arteriolar reactivity to isoproterenol postasphyxia was preserved in both groups 2 and 3. In summary, in newborn pigs, pretreatment with sodium channel blocker resulted in higher CSF cAMP levels and a lesser degree of pial arteriolar dilation during prolonged asphyxia. Pretreatment or treatment at reventilation restored vascular tone and reactivity. PMID- 10709739 TI - Effect of pneumolysin on rat brain ciliary function: comparison of brain slices with cultured ependymal cells. AB - This study compares two models for examining ependymal ciliary function: rat brain slices cut from the fourth ventricle and primary ependymal cells in culture. The cilia from both preparations were very reproducible; each preparation had cilia beating at a constant frequency of between 38 and 44 Hz. With the brain slices, ciliary stasis occurred after 5 d in culture. However, ependymal cells had fully functional cilia for up to 48 d in culture. The pneumococcal toxin, pneumolysin, caused a dose-dependent inhibition of cilia beat frequency within 15 min in both models. There were no significant differences in the mean log 50% inhibitory concentration (pIC50) slice = 0.65 +/- 0.05, equivalent to 4.4 hemolytic units (HU)/mL; cells = 0.57 +/- 0.14, equivalent to 3.7 HU/mL. There were also no significant differences in the mean Hill slope factors for the curves (slice = 1.4 +/- 0.05; cells = 1.6 +/- 0.4). These data demonstrate that both models can be used to examine the acute (15-min) effects of pneumolysin on cilia beat frequency. The main advantage of the primary ependymal culture model is that considerably more cultured ependymal cells (approximately 70%) are available, compared with the number of ependymal cells on the brain slices (approximately 2%), thus reducing the number of animals used. A pure ependymal culture was not achieved (approximately 30% of the cells were not ciliated). The increased survival time of the ependymal cells compared with the brain slices make cultured ependymal cells more useful for examining long-term ciliary function, whereas brain slices may be more useful for examining the interactions between ependymal and other nearby cells. PMID- 10709740 TI - Effects of hypothermia on hypoxia-induced apoptosis in cultured neurons from developing rat forebrain: comparison with preconditioning. AB - In neuronal cultures from the forebrain of 14-d-old rat embryos, transient hypoxia (95% N2/5% CO2, 37 degrees C) for 6 h has been shown to trigger delayed apoptotic death through sequential changes in protein synthesis, whereas preconditioning by a brief episode of hypoxia can rescue neurons. Because hypothermia has been reported to be neuroprotective, the present study was designed to test the influence of reduced temperature on the consequences of lethal hypoxia in our culture model, and cellular mechanisms involved were compared with those underlying preconditioning effects. After 6 d in vitro, cultures were subjected to hypoxia for 6 h. They were either placed at 32 degrees C concomitantly with hypoxia for 6 h or preconditioned the day before by a 1-h episode of hypoxia. The hypoxic insult decreased cell viability by 38% at 96 h after reoxygenation, and 23% of the neurons showed morphologic features of apoptosis. Both hypothermia and preconditioning prevented neuronal death and reduced apoptosis. Preconditioning led to time-dependent changes in leucine incorporation, with persistent overexpression of the survival proteins Bcl-2 and heat-shock protein 70. It also increased thymidine incorporation, in line with induction of the cofactor for DNA polymerase, proliferating cell nuclear antigen. Hypothermia reduced basal apoptosis and necrosis, but did not affect thymidine incorporation, and abolished hypoxia-associated protein synthesis. Therefore, both treatments were protective against neuronal injury consecutive to hypoxia in developing brain neurons in vitro. Whereas preconditioning activated a program that stimulated the expression of anti-apoptotic gene products and regulatory components of the cell cycle, hypothermia did not trigger active processes, but depressed cell activity, which in turn may impair the apoptotic phenomenon. PMID- 10709741 TI - Dynamics of breathing during partial liquid ventilation in spontaneously breathing rabbits supported by elastic and resistive unloading. AB - Partial liquid ventilation (PLV) has been shown to improve gas exchange in paralyzed animals and in humans with lung disease. This study tests the hypothesis that PLV combined with respiratory mechanical unloading results in stable ventilation and gas exchange in spontaneously breathing animals. Ten adult anesthetized, intubated, and spontaneously breathing rabbits received ventilatory support by respiratory mechanical unloading (Fi(O2) 1.0). Minute ventilation, respiratory rate, esophageal pressure, heart rate, and arterial blood pressure were recorded continuously during gas ventilation for 1 h. Next, 30 mL/kg of perfluorocarbon was instilled into the endotracheal tube. Thereafter, data were recorded again for 1 h (PLV). Arterial blood gases were obtained at the end of each period. Variability of recorded data was assessed by calculating coefficients of variation using data obtained each minute. Compared with gas ventilation, minute ventilation was larger during PLV (275 +/- 93 versus 368 +/- 89 mL/kg/min.; p < 0.01). This was because of a higher respiratory rate during PLV (58 +/- 23 versus 74 +/- 18 breaths/min; p < 0.05), while tidal volume was similar. Compared with gas ventilation, Pa(O2) was lower during PLV (61.31 +/- 5.32 versus 47.35 +/- 8.38 kPa; p < 0.05). Pa(CO2), peak esophageal pressure deflections, heart rate, mean arterial blood pressure, and coefficients of variation for minute ventilation, tidal volume, respiratory rate, and peak esophageal pressure were not significantly different between modes. Compliance was decreased and resistance and work of breathing were increased during PLV. We conclude that stable ventilation and gas exchange may be achieved during PLV combined with mechanical unloading in spontaneously breathing animals without lung disease. PMID- 10709742 TI - Lung injury and surfactant metabolism after hyperventilation of premature lambs. AB - We asked whether lung injury and surfactant metabolism differed in preterm lambs after a 1-h period of hyperventilation to P(CO2) values of 25-30 mm Hg. The lambs then were surfactant treated and conventionally ventilated (CV) or high-frequency oscillatory ventilated (HFOV) for an additional 1 or 8 h. The results were compared with lambs that were not hyperventilated or surfactant treated but were ventilated with CV or HFOV. The 1-h hyperventilation resulted in increased alveolar protein, increased recovery of intravascular [131I]albumin in the lungs, and an increase in tumor necrosis factor-alpha mRNA. There were no differences between CV or HFOV in alveolar or total lung recoveries of saturated phosphatidylcholine (Sat PC), tracer doses of [14C]Sat PC and [125I]surfactant protein-B, or in percent Sat PC in large aggregate surfactant in surfactant treated lambs. The lambs not hyperventilated or treated with surfactant had lower large aggregate pools and lower recoveries of [14C]Sat PC and [125I]surfactant protein-B in total lungs than for the surfactant-treated lungs, but there were no differences between the CV and HFOV groups. Hyperventilation followed by surfactant treatment resulted in a mild injury, but the subsequent use of CV or HFOV did not result in differences in surfactant metabolism. PMID- 10709743 TI - Surfactant modulates calcium response of neutrophils to physiologic stimulation via cell membrane depolarization. AB - Pulmonary surfactant (PS) reduces inflammation in the lung by poorly understood mechanisms. We have observed that surfactant-associated proteins (SAP) insert monovalent cation channels in artificial membranes. Neutrophils are primary mediators of acute pulmonary inflammation, and their functions are activated by increases in cytosolic ionized calcium concentration ([Ca2+]) and by changes in membrane potential. We hypothesize that PS inserts SAP-dependent cation channels in neutrophils, causing membrane depolarization, altered [Ca2+] response, and depressed activation. Human neutrophils were isolated, exposed to PS+SAP (1% Survanta), PS-SAP (1% Exosurf), or buffer, and washed before activating with selected stimulants. PS+SAP reduced phorbol ester- and formyl peptide-stimulated adherence and aggregation by 38% (p < 0.05) and 54% (p < 0.02), respectively. PS+SAP also inhibited the formyl peptide-induced [Ca2+] response of neutrophils (p < 0.01), but only in the presence of external Ca2+. Further characterization of this inhibition demonstrated that PS+SAP blocked formyl peptide-induced influx of both Ca2+ and Mn2+, and that this inhibition was present during activation by other neutrophil stimulants (IL-8, immune complexes). Prior depolarization of neutrophils with gramicidin-D similarly inhibited the [Ca2+] response of neutrophils to formyl peptide, and analysis of neutrophil membrane potential by 3,3'-dipentyloxaearbocyanine iodide (diOC5(3)) fluorescence revealed that PS+SAP induced rapid neutrophil depolarization. In contrast, PS-SAP exhibited little effect on neutrophil function, [Ca2+], or membrane potential. We conclude that PS+SAP decreases neutrophil adherence and aggregation responses, blocks Ca2+ influx after physiologic stimulation, and decreases membrane potential. We speculate that these effects are caused by membrane depolarization via SAP dependent cation channel insertion, and that all of these effects contribute to the antiinflammatory properties of PS+SAP. PMID- 10709744 TI - Novel pathway of metabolism of alpha-linolenic acid in the guinea pig. AB - Docosahexaenoic acid (DHA) plays an important role in the nervous system. The capacity of the infant to use the essential fatty acid alpha-linolenic acid (ALA) as a substrate for neural DHA has been the subject of much debate recently. In this study, we explored the metabolic fate of an oral dose of 14C-labeled ALA in guinea pigs fed a defined diet for 3 wk from weaning. Of the 14C-labeled ALA administered, more than 46% was associated with the skin and fur lipids, mostly in the FFA fraction, and less than 0.1% was in brain lipids. About 39% of the label was not recovered in the body lipids and was assumed to be expired as CO2 or unabsorbed. The fur and skin were almost equally labeled; however, because of the very low mass of ALA in the fur, the specific activity of the fur was very high. These data identify a new route of metabolism of ALA in this species, presumably through the sebaceous glands onto fur. If this pathway exists in other species, including humans, it may account for the poor efficiency of conversion of ALA to DHA, because dietary ALA would not be available for anabolic pathways such as DHA synthesis. The relevance of these data to infants is that ALA may play an important hitherto unidentified role in the skin related to barrier function or epidermal integrity. This calls for more research into the importance of ALA as an essential fatty acid in its own right in human infants. PMID- 10709745 TI - Treatment-resistant expansion of CD8+CD28-cells in pediatric HIV infection. AB - There is a disease stage-dependent loss of CD28 expression on T cells in HIV infected children. In this study, T cell recovery, in particular CD28 expression on T cells, was analyzed after initiation of highly active antiretroviral therapy in a group of eight mostly treatment-naive HIV-infected children. Plasma HIV-RNA levels were recorded, and numbers of CD4, CD8, CD4+CD28+, and CD8+CD28+ cells were determined by two-color flow cytometry. Values after 12 mo of therapy were compared with age-matched, seronegative control subjects. CD4 recovery to subnormal values was observed in all children. CD8+CD28+ cells recovered and were within the normal range after 12 mo of therapy (patients, 703 +/- 250 cells/microL; controls, 789 +/- 269 cells/microL), whereas CD8+CD28- cells (546 +/- 269 cells/microL) remained significantly expanded compared with age-matched controls (140 +/- 35 cells/microL). Expansions of CD8+CD28- cells persisted even in cases with long-term suppression of viral replication. Highly active antiretroviral therapy in HIV-infected children induces substantial but incomplete T cell recovery. PMID- 10709746 TI - Production economics analysis of investment initiated to improve working environment. AB - This article describes the results of an evaluation of a new work place for ladle preparation at Swedish Steel in Lulea, Sweden. The company initiated a development project related to ladle service work, in order to come to grips with the difficult working environment and problems associated with absenteeism due to illness and occupational injuries. The evaluation was performed for the first three years after implementation of the project and it shows that the new work place considerably improved working conditions and increased both the quality and efficiency of production. The purpose of this article is also to discuss some methodological problems. The follow-up of the various changes in working environment and personnel statistics was fairly simple to carry out. But in terms of production effects, the company's in-house production follow-up system proved to be too unspecified and oversimplified. It was also difficult to decide which changes should count as effects of the new work place and to value these in monetary terms. The profitability calculation shows that an investment initiated to improve the working environment can yield good profitability. PMID- 10709747 TI - Biomechanical analysis of the dimensions of pilot seats in civil aircraft. AB - The dimensions of pilot seats from five different types of civil aircraft were measured and the results compared with existing standards and biomechanical criteria. It was apparent that these seats failed to meet requirements, particularly in the effective depth and inclination of the seat and in the height of the lumbar support and the armrests. Hence, none of these seats made it possible for the pilot to establish a comfortable sitting posture. In comparison with aviation standards, the anthropometric dimensions were not satisfactory, meeting only 4-7 out of 10 requirements. The dimensions based on biomechanics were even less satisfactory, meeting only between 1 and 3 requirements out of 7. PMID- 10709748 TI - Determining fatigue allowances for grocery order selectors. AB - This paper compares and contrasts four fatigue allowance worksheets commonly used to establish fatigue (relaxation) allowances for production standards; the factors, weights, degree of documentation and other operational characteristics of these worksheets are also examined. We briefly review the fatigue literature including objective physical measures of fatigue. Results of an experiment in which 11 industrial engineers independently applied the four worksheets to a standardized job analysis and video tape of a grocery order selector are reported. We conclude that inter-rater reliability and cross-validation are very low for the four worksheets and suggest validation studies using objective physiological measures of fatigue would be appropriate. PMID- 10709749 TI - Maximum forces sustained during various methods of exiting commercial tractors, trailers and trucks. AB - Many commercial vehicles have steps and grab-rails to assist the driver in safely entering/exiting the vehicle. However, many drivers do not use these aids. The purpose of this study was to compare impact forces experienced during various exit methods from commercial equipment. The study investigated impact forces of ten male subjects while exiting two tractors, a step-van, a box-trailer, and a cube-van. The results showed that exiting from cab-level or trailer-level resulted in impact forces as high as 12 times the subject's body weight; whereas, fully utilizing the steps and grab-rails resulted in impact forces less than two times body weight. An approach that emphasizes optimal design of entry/exit aids coupled with driver training and education is expected to minimize exit-related injuries. PMID- 10709750 TI - Psychophysical assessment of assistive devices for transferring patients/residents. AB - This article reports the psychophysical assessment of nine battery-powered lifts, a sliding board, a walking belt, and a baseline manual method for transferring nursing home patients/residents from a bed to a chair. A separate article reports the biomechanical evaluation of the same task and devices. The objectives of the psychophysical assessment were to investigate the effects of resident transferring methods on the psychophysical stress to nursing assistants performing the transferring task, and to identify transfer methods that could reduce the psychophysical stress reported by nursing assistants. Nine nursing assistants served as test subjects. Two elderly persons participated as residents. The results indicated that the psychophysical stresses on nursing assistants were significantly lower when performing resident transfers with some of the assistive devices than when performing transfers with the baseline manual transfer method. The nursing assistants generally preferred the basket-sling lift and stand-up lift to other methods. The residents' comfort and security ratings indicated the comfort and security with most of the assistive devices were greater than or equal to the baseline manual method. Most of the comments of the nursing assistants and residents on the assistive devices were favourable. PMID- 10709751 TI - Postural analysis of paramedics simulating frequently performed strenuous work tasks. AB - Paramedics who perform emergency rescue functions are highly susceptible to musculoskeletal injuries. Through an interview and survey process firefighters, many of whom are cross-trained paramedics in a consortium of 14 suburban fire departments, identified and rated tasks that were perceived to be both strenuous and frequently performed. The objective of the current study was to describe the working postures and the forces applied as firefighter/paramedics (FF/Ps) simulated specific roles within the following tasks identified by the survey: (1) transferring a patient from a bed to a stretcher using bedsheets, (2) transferring a patient from the ambulance stretcher to a hospital gurney, (3) carrying a victim down a set of stairs and around a landing using a stairchair, (4) carrying a victim down a set of stairs and around a landing using a backboard, and (5) carrying a victim down a set of stairs using a stretcher. Ten two-person teams of FF/Ps participated and were videotaped to obtain postural data for the upper and lower extremities as they performed each role in the simulated two-person tasks. Trunk postures were obtained using lumbar motion monitors. Static hand forces were estimated using a hand-held dynamometer at the most physically demanding points for each role within each task. The postural and force data were averaged across subjects performing identical roles to quantify the postures assumed by the FF/Ps at the most strenuous moments during task performance. Based on these analyses we concluded that: (1) when transferring victims from a bed to a stretcher the FF/P on the bed was able to maintain an upright and more stable posture by standing as opposed to kneeling, (2) an interface board should be used to reduce the frictional forces when transferring victims from a bed to a stretcher or from a stretcher to a gurney, thereby reducing the need to lift the victim with flexed torsos and/or shoulders, and (3) equipment and training that encourages the FF/P in the leader role to walk facing forward during victim transport, especially when descending stairs, potentially results in safer transit. PMID- 10709752 TI - The evaluation of workplaces subjected to heat stress: can ISO 7933 (1989) adequately describe heat strain in industrial workplaces? AB - The International Standard ISO 7933 (1989) Hot environments--Analytical determination and interpretation of thermal stress using calculation of required sweat rate has been proposed for the evaluation of climatic stress within the European system of CEN standards. Comparison of results of studies performed in climatic chambers and those in the field with the predictions of ISO 7933 show that there are considerable problems in using this index in practice. In its present state of development, ISO 7933 seems to be rather a step towards a useable index for evaluating climatic conditions rather than an established climatic index which is applicable in practice. Within the CEN standards, the deficiencies of ISO 7933 are reflected mainly by a restriction of the limits of application within EN 12 515 (1997) which is based on ISO 7933. PMID- 10709753 TI - A participant-observer study of ergonomics in engineering design: how constraints drive design process. AB - Too often, ergonomics is relegated to being a "post-design" evaluation, leaving ergonomists little opportunity to make significant and important design changes. One way to start attacking this problem is to study the process of design and, in particular, ergonomics in design. This article describes the findings from a four month long participant-observer study of the relationship between ergonomics and engineering design. The study was conducted in the context of a large, interdisciplinary project consisting of design of a control room for a nuclear power plant. It was observed that designers and ergonomists must negotiate through a changing web of constraints from many sources. The impact that these constraints had on the course of the design was documented. A model is developed based on the abstraction hierarchy (Rasmussen, 1985, IEEE Trans. Systems Man Cybernet. SMC-15, 234-243; 1990, Int. J. Ind. Ergon. 5, 5-16) which shows the interaction of conflicting goals as ergonomists and other designers attempt to solve a complex design problem. This model leads to several insights: (1) locally optimal ergonomic designs may not be globally optimal, (2) ergonomists can improve their solutions by understanding the goals of other designers, and (3) future tools to aid ergonomists must be compatible with the constraint-rich environments in which they work. PMID- 10709754 TI - Evaluation of an 8 hour versus a 12 hour shift roster on employees at a power station. AB - Several studies exist that have conducted research into the effects of different shiftwork patterns on the individual, especially regarding 8 and 12 h rosters. The findings of these studies have been largely supportive of longer shifts, however, the effects on work performance are not as clear cut. This study aimed to examine the changeover from an 8 h roster to a 12 h roster in a power station via monitoring on-shift performance, general health and well-being, sleep and mood behaviour, as well as absence and accident data. Results suggest that the domestic and social life of workers was markedly improved under the 12 h roster. Improvements in physical health, sleeping behaviour and mood state of employees were also documented. On-shift performance measures showed an increase in error rates at the end of a 12 h shift. Ways of reducing the risk of error towards the end of a 12 h shift should be explored. The results of this study suggest that 12 h shifts are a valid alternative to 8 h shifts in this particular workplace, although tasks that require error-free activities should not be performed towards the end of a 12 h shift. PMID- 10709755 TI - Length of the spine while sitting on a new concept for an office chair. AB - Changes in spinal length were used to evaluate a new concept for an office chair. This so-called dynamic chair imparts passive forced motion to the seated subject. The passive forced motion is a rotary movement about an axis, perpendicular to the seat with amplitude of 0.6 degrees and a frequency of 0.08 Hz. Change of stature is assumed to provide a measure for spinal load. Eight subjects were measured in two situations: static (without motion) and dynamic. In both situations the same office tasks were performed and the duration of the sitting period was 1 h. To allow for the normal shrinkage curve the starting time was the same on each of the measurement days. The results indicated a significant difference: when sitting on the dynamic chair the average spinal length increased in comparison to the spinal length in the static chair, where average spinal length decreased. It was concluded that there is spinal distress relief due to the passive motion of the chair. PMID- 10709756 TI - Warning! Nearby construction can profoundly affect your experiments. AB - This is meant to alert people to potentially major effects of construction projects on research results. Because we study the effects of stress on regulation of ACTH and corticosterone secretion and of serotonin receptors and stress on energy balance, we serve as an early warning system when things go awry. Most of our experiments include taking daily, or twice daily, measurements of rat or mouse weights and food intake as well as stress hormone levels. We are highly sensitized to environmental disruption and we've shown previously the effects of construction on stress hormones (1). However, we did not anticipate the change and disruption in energy balance that may occur in response to environmental perturbation. We provide two examples of these, below. PMID- 10709757 TI - Mammary gland sympathetic innervation is a major component in type 1 deiodinase regulation. AB - Recent observations have shown that in lactating rats previously deprived of suckling, either suckling stimulus or ip injection of norepinephrine was capable of increasing mammary deiodinase type 1 (M-D1) mRNA content and enzyme activity. In the present work, we show that intact efferent sympathetic mammary innervation is required to restore both mammary D1 mRNA content and enzyme activity, whereas suckling-induced secretion of catecholamines from the adrenal glands does not seem to participate in M-D1 enzyme regulation. The data also indicate that the sympathetic reflex activation in response to suckling involves two complementary autonomic components: (1) activation, presumably through mammary segmental arrangement affecting neighboring mammary glands; and (2) an individual reflex regulatory mechanism capable of maintaining M-D1 activity within each mammary gland. In addition to these findings, we show that the suckling-induced sympathetic activation of M-D1 activity could be blocked by prior activation of ductal mechanoreceptors. This set of regulatory and counterregulatory mechanisms seems to ensure the optimal control of mammary energetic expenditure according to litter size. PMID- 10709758 TI - Estrogen downregulates neuronal nitric oxide synthase in rat anterior pituitary cells and GH3 tumors. AB - The anterior pituitary gland produces neuronal nitric oxide synthase (nNOS) and nitric oxide regulates secretion of various anterior pituitary hormones. Estrogen has many functions in anterior pituitary cells including stimulation of prolactin (PRL) cell proliferation and secretion of various anterior pituitary hormones. However, the role of estradiol-17beta (E2) in regulating pituitary nNOS expression has not been previously examined. We studied the regulation of nNOS in normal pituitaries, and neoplastic GH3 pituitary tumors in order to analyze the effects of E2 on nNOS in pituitary cells. GH3 tumors expressed higher levels of nNOS proteins compared to normal pituitaries. Estrogen downregulated nNOS mRNA and protein in both estrogen-treated pituitaries with PRL cell hyperplasia and in GH3 tumors implanted into the flank of rats treated with E2 in silastic tubing. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis demonstrated three alternatively spliced nNOS transcript isoforms--nNOSa, nNOSb, and nNOSc mRNAs--with distinct 5' untranslated first exons that arose from alternative splicing to a common second exon. All three spliced isoforms were found in the normal rat pituitary, whereas nNOSa and nNOSb, but not nNOSc, were expressed in GH3 tumors implanted into Wistar-Furth rats. E2 also downregulated the nNOSa alternative mRNA transcript isoform in vivo. These results indicate that the biological activity of nNOS in the normal rat anterior pituitary and in pituitary tumors is regulated by a complex pattern of alternative splicing and that some of these mRNA isoforms as well as nNOS protein are regulated by estrogen. Our results also indicate that the levels of nNOS and the alternatively spliced nNOS transcript between normal and GH3 pituitary tumors are different. PMID- 10709759 TI - Differential influences of gender and physiological status on calcium dynamics and prolactin gene expression in rat mammotropes. AB - The rate of prolactin (PRL) secretion is influenced by the gender and physiological state of an animal, but little is known about the mechanisms involved. In the present study, we assessed possible contributions of Ca2+ dynamics and PRL gene expression to these differences. This was accomplished by monitoring spontaneous [Ca2+]i changes and PRL promotor-driven reporter activity in pituitary cultures derived from rats comprising a broad spectrum of PRL secretory capacities: male, cycling female, and lactating rats. We found that Ca2+ oscillatory activity exhibited a rank order of lactating > cycling females > males, consistent with the reported secretory capacities of mammotropes from these sources. Interestingly, we observed that the basal level of PRL promotor driven reporter activity was the same for all three models, but that mammotropes from males were the most responsive to stimulation of PRL gene expression by elevation of [Ca2+]i. Collectively, our findings reveal gender- and state specific differences in Ca2+ dynamics and induction of PRL gene expression. These likely contribute to reported differences in secretory capacity. PMID- 10709760 TI - Steroid feedback on gonadotropin release and pituitary gonadotropin subunit mRNA in mice lacking a functional estrogen receptor alpha. AB - Steroid hormones regulate levels of gonadotropin mRNA in the pituitary, and gonadotropic hormones in plasma. To determine whether estrogen receptor alpha (ERalpha) mediates steroid negative feedback, wild type (WT) and estrogen receptor alpha knockout (ERalphaKO) mice of both sexes were gonadectomized and implanted with a Silastic capsule containing either estradiol (E2), dihydrotestosterone (DHT), testosterone, or a blank capsule. Ten days later, plasma luteinizing hormone (LH) and follicle-stimulating hormone (FSH) levels were measured. Pituitary mRNA levels of gonadotropin subunit (alpha, LHbeta, FSHbeta) and prolactin (PRL) were quantified. LH levels in gonad-intact ERalphaKO females were elevated, similar to values seen following gonadectomy. By contrast, serum LH concentrations in gonad-intact ERalphaKO males were low and rose following gonadectomy, suggesting androgen feedback. Estradiol treatment significantly decreased plasma LH in WT animals, but not in ERalphaKOs. In fact, in female ERalphaKOs, our dose of E2 increased plasma levels of LH as compared with untreated, ovariectomized ERalphaKOs. All the steroid treatments suppressed LH in WT animals whereas only DHT consistently suppressed LH concentrations in ERalphaKO mice. The postgonadectomy rise in plasma FSH was prevented by steroid treatments in WT females, but not in any of the other groups. Gonadotropin subunit and PRL mRNA responses to E2 treatment (both inhibitory and stimulatory) were absent in ERalphaKO mice, suggesting a critical role for ERalpha. Although E2 can exert negative feedback effects on LH release in both males and females by actions at the ERalpha, the androgen receptor plays the primary physiological role in the male mouse. PMID- 10709761 TI - Responses of serum corticosterone and corticosteroid-binding globulin to acute and prolonged stress in the rat. AB - Responses of serum corticosterone (B) and corticosteroid-binding globulin (CBG) to ether anesthesia (a "classic" acute stress) and to a number of stressors influencing metabolic homeostasis--fasting, physical exercise, cold exposure, and water deprivation--were studied in male and female rats. Metabolic stressors included placing in an ice bath, physical exercise (swimming), fasting for 2 d, swimming after fasting for 2 d, cold-room (4 degrees C) exposure for 2 d, fasting in combination with cold-room exposure for 1 d, and water deprivation for 2 d. The study demonstrated clear differences between males and females in basal B levels and B responses to some stressors. Only ether anesthesia and fasting resulted in similar B levels in males and females whereas in control and all other groups serum B levels were higher in females. Serum CBG was considerably higher in females. In females, ether, swimming, swimming after fasting, fasting, and fasting during cold exposure resulted in a decrease in circulating CBG. Ice bathing and cold exposure did not influence CBG, and water deprivation elevated serum CBG. In males, animals subjected to fasting and fasting during cold exposure had CBG levels lower than control animals. Other groups did not differ from the control. Higher CBG levels in females counterbalanced higher total B in setting circulating free B: significant sex differences in free B were observed only after swimming or fasting during cold exposure. Stress-responsive changes in CBG levels seem to contribute little to changes in free B; the main contributing factor is the rise in total B. However, CBG may play a special role, independent of the functions of corticosteroids. It is proposed that the need for substantial mobilization of spare fuel (as it takes place during physical exercise or fasting) is critical in involving CBG in the stress response. PMID- 10709762 TI - Induction of somatotroph differentiation in vivo by corticosterone administration during chicken embryonic development. AB - Somatotroph differentiation in the embryonic pituitary of avian and mammalian species can be stimulated by glucocorticoids in vitro, and this effect can be augmented by concomitant treatment with growth hormone-releasing hormone (GHRH). Owing to its isolation from maternal influences, the chick embryo is a useful model for studying humoral regulation of pituitary cell differentiation. Somatotroph differentiation in chickens occurs between embryonic day (e-) 14 and e-16, and treatment of e-12 pituitary cells with e-16 serum or corticosterone induces growth hormone (GH) cell differentiation within 2 d in culture. The objective of the present study was to determine whether direct administration of embryonic serum and corticosterone to developing chick embryos was effective in vivo in inducing somatotroph differentiation prematurely. The albumen of fertile eggs was injected on e-11 with 300 approximately microL of 0.9% saline or 150 microL of serum from e-12 or e-16 chick embryos diluted 1:1 with saline. The embryos were allowed to develop until e-14, when pituitaries were dispersed and the resulting pituitary cells were subjected to reverse hemolytic plaque assays (RHPA) and immunocytochemistry to detect GH-secreting and GH-containing cells, respectively. Injection of e-16 serum increased (p < 0.01) GH-secreting and GH containing cells to 11.5 +/- 1.0% and 1 7.4 +/- 3.3% of all pituitary cells, compared to 5.0 +/- 0.3% and 5.5 +/- 0.9% for saline-injected controls, respectively. Day 12 serum increased GH-containing cells to 9.8 +/- 0.9%, without changing percentages of GH-secreting cells. In experiment 2, saline, e-16 serum, and corticosterone were injected on e-11, and pituitary cells were subjected to GH RHPA on e-14. GH secretors were increased by e-16 serum and corticosterone. In experiment 3, we tested whether GHRH would magnify the effect of corticosterone, as we had seen in extended 6-d cultures previously. Saline, corticosterone, and corticosterone plus GHRH were injected on e-11, and pituitary cells were subjected to GH RHPA on e-18. Treatment with corticosterone alone and combined with GHRH increased the percentage of GH-secreting cells. However, combined treatment with corticosterone and GHRH was not more effective than corticosterone alone. The present findings demonstrate that glucocorticoid administration can stimulate somatotroph differentiation in living vertebrate embryos isolated from maternal interactions. PMID- 10709763 TI - The effects of histone acetylation on estrogen responsiveness in MCF-7 cells. AB - Because histone acetylation is implicated in the facilitation of specific gene transcription, the effect of increasing histone acetylation on the expression of an endogenous gene was investigated. The ability of trichostatin A (TSA), a histone deacetylase inhibitor, to potentiate the estradiol (E2) induction of progesterone receptor (PR) levels in MCF-7 cells was studied. Although TSA alone had no effect on PR synthesis, measured by a whole-cell binding assay with [3H]R5020, TSA potentiated the effect of 10(-11) ME2 such that 10 ng of TSA/mL approximately doubled the hormone response. When TSA was removed from the cells after various incubation times (24 and 48 h) by successive washings with TSA-free medium, it was determined that TSA was required throughout the 96-h incubation period in order to achieve maximal potentiation for the E2 response. In addition, TSA potentiated E2 induction of pS2 mRNA. These results suggested that the estrogen receptor (ER) complex might alter histone acetylation as part of the gene activation process. To test this directly, MCF-7 cells were incubated for 48 h with E2 followed by incubation with sodium [3H]acetate for 1 h. From two dimensional polyacrylamide gel electrophoresis, an increase in total acetate incorporation into histones in estrogen- treated cells compared to control was observed as well as a preferential increase in the mono- and diacetylated histone H4. Experiments with lysine-specific antiacetylated H4 antibodies suggest a preferential increase in acetylation at lysine 16, but not 5, 8, or 12. The results of this study support an important role for histone acetylation in the mechanism of action of the ER. PMID- 10709764 TI - Inhibition of iodine organification and regulation of follicular size in rat thyroid tissue in vitro. AB - The factors mediating the accumulation of thyroglobulin are of great importance to the understanding of the pathogenesis of human and experimentally induced colloid goiters. To elucidate further the underlying cellular mechanism, thyroid fragments from newborn rats were incorporated into semisolid alginate beads and were cultured as three-dimensional organoids for up to 21 d. In five parallel cultures, the medium contained either no supplements (group A), Nal (group B), thyroid-stimulating hormone (TSH) (group C), Nal plus TSH in the same concentrations as B and C (group D), or Nal and TSH (as in group D) plus methimazole (MMI, group E). The thyroid organoids maintained morphological integrity, functional activity, and ability to proliferate in vitro. Addition of iodine to the cultures significantly increased mean (+/-SEM) follicular diameters from 19.5 +/- 0.7 microm in controls to 33.9 +/- 2.2 microm (p < 0.0001) when NaI was added alone (group B), and 30.4 +/- 1.7 microm (p < 0.0001) when combined with TSH (group D). The effect of NaI on follicular size was abolished by MMI (group E, follicular diameter 23.5 +/- 1.3 microm). The results presented support the recent finding, using a rat colloid goiter model, that not only TSH but also iodine organification or its inhibition are important factors in modulating follicular morphology. PMID- 10709765 TI - Effects of overexpression of growth hormone-releasing hormone on the hypothalamo pituitary-gonadal function in the mouse. AB - In this investigation, the neuroendocrine alterations induced by high, chronic circulating levels of endogenous growth hormone (GH) were studied in transgenic mice with ectopic overexpression of the human growth hormone-releasing hormone (h GH-RH) gene. In comparison with their normal littermates, transgenic h-GH-RH mice had elevated plasma levels of GH, prolactin (PRL), and corticosterone. In addition, they had elevated body, liver, kidney, spleen, and pituitary weights compared with normal mice. Testis and seminal vesicle weights were also increased in transgenic mice. Although basal plasma luteinizing hormone (LH) levels, plasma estradiol levels in females, and plasma testosterone levels in males did not differ significantly between normal and transgenic animals, the LH response to castration was severely impaired in transgenic mice of both sexes. Among the biogenic amines studied in the hypothalamus, only dopamine concentrations were significantly lower in transgenic animals compared with their normal littermates. This decrease in hypothalamic dopamine may be related to the hyperprolactinemia in transgenic animals. In vitro, pituitaries from transgenic mice released significantly higher amounts of GH, and although the basal release of LH was not different in both normal and transgenic mice, the response to gonadotropin releasing hormone was significantly smaller in transgenic mice. Cultured anterior pituitary cells from transgenic mice secreted high quantities of GH and PRL in vitro, but these quantities significantly decreased from 1 to 8 wk in culture. These results show that high, persistent levels of circulating endogenous GH induce alterations in neuroendocrine functions related to the hypothalamo pituitary-gonadal and the hypothalamo-pituitary-adrenal axes. PMID- 10709766 TI - Perinatal hypoxia-ischemia decreased neuronal but increased cerebral vascular endothelial IGFBP3 expression. AB - In adults, insulin-like growth factor binding protein 3 (IGFBP3) is the main carrier protein for circulating insulin-like growth factors (IGFs) (IGF-I and II). While most IGFBP3 is synthesized in the liver, it is also expressed locally by many cell types including vascular endothelial cells. The regulation of this endothelial IGFBP3 expression, especially in response to hypoxic-ischemic injury, has not been investigated in vivo. Using in situ hybridization histochemistry, we studied the cellular distribution of IGFBP3 mRNA in rat brains following hypoxic ischemic injury at 1, 5, 24, and 72 h of recovery. In normal P7 rat brain, IGFBP3 mRNA was found in neurons within the thalamus, hippocampus, and amygdaloid. Low levels of IGFBP3 mRNA were also detected in cerebral vascular endothelial cells. After the hypoxic-ischemic injury, the levels of neuronal IGFBP3 mRNA substantially decreased within 24 h in areas that were normally supplied by the middle cerebral artery. In the meantime, there was an immediate increase in IGFBP3 expression in vascular endothelial cells throughout the affected hemisphere. This vascular IGFBP3 expression was further enhanced with the highest level at 24 h of recovery whereas neuronal IGFBP3 expression was further decreased. By 72 h of recovery, IGFBP3 was no longer expressed in vascular endothelial cells. Taken together, the activation of IGFBP3 is a likely mechanism by which vascular endothelial cells respond to hypoxic-ischemic insult. In addition, increased endothelial IGFBP3 may modulate the interaction of IGFs with IGF-I receptors at the site of injury and/or act independently on endothelial cell growth. PMID- 10709767 TI - Short- and long-term effects of beta-cellulin and transforming growth factor alpha on beta-cell function in cultured fetal rat pancreatic islets. AB - The polypeptide beta-cellulin, identified in conditioned media from insulinoma cell cultures and produced by pancreatic islet cells, was recently identified as a possible autocrine growth factor for the pancreatic islet beta-cell. In this study, we investigated the short- and long-term actions of beta-cellulin, and the structurally related transforming growth factor-alpha (TGF-alpha), on beta-cell function in fetal rat pancreatic islets in vitro. We found that neither beta cellulin nor TGF-alpha (10 nM each), in contrast to glucose (20 mM), acutely influenced beta-cell levels of cytosolic-free Ca2+. Additionally, whereas glucose markedly increased short-term (60-min) insulin release, neither beta-cellulin nor TGF-alpha (10 nM each) influenced the rate of hormone secretion at basal (3 mM) or stimulatory (20 mM) concentrations of glucose. Likewise, long-term (24-h) exposure of islets to a high glucose concentration significantly augmented the secretion of insulin. This effect was slightly potentiated by TGF-alpha (10 nM), but not beta-cellulin (10 nM), at high (but not low) glucose concentrations. Conversely, the islet insulin content was not significantly affected by beta cellulin or TGF-alpha at any glucose concentration tested. We conclude that, although beta-cellulin is produced by islet cells, the peptide does not seem to be of importance for the regulation of insulin production by isolated pancreatic beta-cells. PMID- 10709769 TI - Freedom, responsibility, and care: Hong Kong's health care reform. PMID- 10709768 TI - Differential induction of androgen receptor transactivation by different androgen receptor coactivators in human prostate cancer DU145 cells. AB - Recently identified androgen receptor (AR) coactivators were used in this study to determine whether the specificity of sex hormones and antiandrogens could be modulated at the coactivator level. We found that ARA70 is the best coactivator to confer the androgenic activity on 17beta-estradiol. Only ARA70 and ARA55 could increase significantly the androgenic activity of hydroxyflutamide, a widely used antiand rogen for the treatment of prostate cancer. None of the AR coactivators we tested could significantly confer androgenic activity on progesterone and glucocorticoid at their physiological concentrations (1-10nM). We also found that ARA70, ARA55, and ARA54, but not steroid receptor coactivator-1 (SRC-1) and Rb, could significantly enhance the delta5-androstenediol-mediated AR transactivation. Furthermore, in comparing the relative specificity of these coactivators to AR in DU145 cells, our results suggested that ARA70 has a relatively higher specificity and that SRC-1 can enhance almost equally well many other steroid receptors. Finally, our data demonstrated that AR itself and some select AR coactivators such as ARA70 or ARA54 could, respectively, interact with CBP and p300/CBP-associated factors that have histone acetyl-transferase activity for assisting chromatin remodeling. Together, our data suggest that the specificity of sex hormones and antiandrogens can be modulated by some selective AR coactivators. These findings may not only help us to better understand the specificity of the sex hormones and antiandrogens, but also facilitate the development of better antiandrogens to fight the androgen-related diseases, such as prostate cancer. PMID- 10709770 TI - Does it really care? The Harvard report on health care reform for Hong Kong. AB - This paper aims to provide a rendition of the care ethic in Confucian philosophy and to argue that social policy developments in Hong Kong society, including health care policy, have been significantly shaped and justified in terms of the ideal of care in the Confucian moral tradition. On the basis of this analysis, the paper raises a number of questions about a recent proposal for health care reform for Hong Kong put forth by the Harvard School of Public Health which argues for adopting the principle of equity as the overriding value for the moral foundation of Hong Kong's health care system. The paper examines how the over emphasis on equity in the Harvard Report proposals can lead to the erosion of care and ultimately the eclipse of the vision of care in Hong Kong's health care system. It argues that the pursuit of equity, which is itself a valuable principle, should not displace the importance of the value of care or undermine the ideal of care and that health care decisions must be firmly embedded in local cultures and moral traditions. PMID- 10709771 TI - Power of politics and reasonableness in policy study: on some methodological problems with the Harvard Team Report. AB - The so-called "Harvard Team Report," commissioned by the Hong Kong government (Hong Kong SAR Government, 1999), suggests significant institutional changes to the local health care system, including a partial shift of the financial burden directly to the citizens. I argue that 1) the Report's adoption of the contextuality principle as its research framework encounters practical problems in collecting data for a reliable analysis; 2) the existing health care system already satisfies the Report's first guiding principle; 3) the Report's employment of the "working assumption" of the government (i.e., not increasing its financial support of health care) as its second guiding principle is questionable, for the share of the percentage of GDP as represented by the existing system (4.6% in 1996) is small enough; and 4) because of 3), the Report is unnecessarily constrained in its choices of considered options and seems to overlook some feasible ones. In conclusion, the methodological reasonableness of the Report is questioned. PMID- 10709772 TI - Constructing options for health care reform in Hong Kong. AB - The Harvard Report, published in April 1999 for public consultation in Hong Kong, proposed a fundamental restructuring in its health care delivery and financing systems. The Report claims to be evidence-based in its approach (Hsiao et al., 1999a). While 'evidence' has been widely collected by the consultancy team through surveys, consultations and focus groups, the recommendations put forth are not value-free. They carry clear ideological preferences. The value assumptions and ethical presuppositions underlying the report are discussed in this paper. The Harvard consultancy study is in favor of a positive government role in regulation and control, a single central body to administer compulsory health insurance for all citizens, and a purchaser-provider split to induce competition. Such preference is based on pre-existing ideology and generic health care management concepts, which are still in the experimental phase internationally. While value and ideology are inevitable factors in any policy choice, the challenge is to lay these values open for reflection and public debate. For Hong Kong, the challenge is also to take on local substantive issues in health care and deal with them head-on, rather than putting hope in a universal, generic solution. PMID- 10709773 TI - Free choice, equity, and care: the moral foundations of health care. AB - The aims of this paper are threefold. The first aim is to provide a critique of the reform proposal of the Harvard School of Public Health for Hong Kong's health care system through privatization of the public sector services. The second aim is to argue for the duty of society to guarantee every member equal access to a basic level of health care based on the values of equity, care and free choice. The third aim is to explore some suggestions about delivery structures and financial arrangements of a dual sector health care system which will better enable society to provide a basic level of health care that is sustainable and affordable, while being at the same time consistent with the values of care, equity and free choice. PMID- 10709774 TI - Health care reform and societal values. AB - Hong Kong is undergoing a public debate on the need to reform and future directions of reforming its health care system. This paper highlights the debates and considerations brought up by the Hospital Authority, the largest provider of public health care in Hong Kong, on the ethical principles and societal values underlying the upcoming reform. It is recognized that the exact meanings behind each ethical principle and value must be debated and clarified during the reform process. In a modern day society like Hong Kong, societal values are likely to be diversified. A health care system also has to fulfil different and often conflicting objectives of equity, efficiency, quality and choice. It would be difficult for a health care system to satisfy these different values and objectives based on a single value parameter. The Hong Kong experience shows that a society may prefer a combination of strategies in addressing different societal values. The re-structuring of the health care system in Hong Kong should therefore be based on a balanced and optimum combination of various financing and delivery strategies. PMID- 10709775 TI - Drugs in nails: physiology, pharmacokinetics and forensic toxicology. AB - In recent years, drug analysis in keratinised matrices, such as hair and nails, has received considerable attention because of several advantages over drug testing methodologies employing body fluids, such as urine or serum. For example, keratinic matrices, such as finger- and toenails, can accumulate drugs during long term exposure. Drugs are incorporated into nails by a double mechanism: (i) deposition into the root of the growing nail via the blood flow in the nail matrix; and (ii) incorporation via the nail bed during growth from the lunula to the beginning of the free margin. Together, these account for a wide retrospective window of drug detection. Nails can provide a good forensic matrix for the detection of drugs of abuse. Indeed, the international literature has reported the use of nail analysis in postmortem detection of drugs of abuse, drug testing in the workplace and drug screening to detect prenatal exposure, even though further studies are needed for correct interpretation of the data obtained. Another application of drug analysis in nails consists of the possibility of detecting the presence of an antimycotic at the site of action during antifungal therapy for patients with onychomycosis. When available, this evidence has permitted drug treatment of a shorter duration and reduced toxicity. However, so far the potential of drug monitoring in nails still lacks harmonisation and validation of analytical methodologies and a better comprehension of the possible correlation between drug concentrations in the matrix and period of exposure. PMID- 10709777 TI - Target-controlled infusion systems: role in anaesthesia and analgesia. AB - Drug delivery by target-controlled infusion (TCI) allows automatic adjustments of the infusion rate of a drug to maintain a desired target concentration. Since drug effect is more closely related to blood concentration than to infusion rate, drug delivery via TCI is capable of creating stable blood concentrations of intravenous anaesthetics and analgesics. In this article the concept and history of TCI are described. The rational administration of TCI requires an appropriate pharmacokinetic data set and knowledge of the concentration-effect relationship; therefore, general pharmacokinetic and pharmacodynamic aspects of intravenous anaesthetics and analgesics are also addressed. Intraoperative investigations have demonstrated that TCI drug delivery allows rapid titration to a desired effect. The use of TCI for postoperative analgesia is still experimental, but TCI can, in part, overcome the disadvantages associated with continuous infusions and patient-controlled analgesia regimens in the postoperative period. Although TCI is capable of creating stable blood concentrations, when the target concentration is changed the resulting effect correlates better with a theoretical effect site concentration. The efficacy of TCI systems that can perform effect-site steering are still to be explored. PMID- 10709778 TI - Human toll-like receptors 2 and 4 are targets for deactivation of mononuclear phagocytes by interleukin-4. PMID- 10709779 TI - Dendritic cell migration in different micropore filter assays. AB - Dendritic cells (DC) are highly motile and have been shown to migrate in vitro or in vivo towards various chemoattractants. Micropore filter methods with polycarbonate filters are generally used in these in vitro experiments. Among others, the main drawback of these filters is their thickness, which does not allow any assessment of effects of absolute concentration compared to gradients. The aim of this study was to establish a chemotaxis assay for dendritic cells using nitrocellulose filters, which can be adapted for checkerboard studies to distinguish between chemokinesis and chemotaxis. Immature DC were generated by culture of peripheral blood mononuclear cells using granulocyte-macrophage colony stimulating factor and interleukin-4. We tested cell migration into nitrocellulose in a Boyden microchemotaxis chamber (leading front assay) and compared this method to the commonly used polycarbonate filter technique. Dendritic cells migrated well into nitrocellulose towards gradients of formyl peptide, complement fragment 5a, and monocyte chemotactic protein-3. The nitrocellulose method appeared to be more sensitive as compared to experiments testing migration across polycarbonate filters. Subsequent checkerboard analyses confirmed chemotactic activities of formyl peptide and complement fragment 5a. However, depending on the assay system, chemotaxis in polycarbonate filters but chemokinesis in nitrocellulose filters were observed for monocyte chemotactic protein-3. Measurement of DC migration in a cellulose nitrate micropore filter assay is more sensitive than the commonly used polycarbonate method and can be adapted for checkerboard analyses. PMID- 10709780 TI - Cysticercosis: towards the design of a diagnostic kit based on synthetic peptides. AB - Cysticercosis caused by Taenia solium is a very common disease in developing countries that seriously affects human health. Diagnosis can only be confirmed with the aid of computerized tomography or nuclear magnetic resonance (NMR) creating obvious difficulties for epidemiological studies. Reliable immunoassays employing cerebrospinal fluid (CSF) have been developed, based on the use of cysticercal antigens. However, the reliance on parasite material is restrictive. Herein, we report the advances in the design of a diagnostic kit based on immunodominant synthetic peptides, targeting four candidate epitopes KETc1, KETc12, 410 and 413 which were identified from three different clones (KETc1, 12 and 4) selected from a cDNA library of Taenia crassiceps. CSF antibodies against T. solium cysticercal antigens (TCA) as well as the four peptides were determined by enzyme-linked immunoabsorbent assays (ELISA) using two panels of CSF from patients with confirmed neurocysticercosis and other neurological diseases. In the first CSF panel which included patients with high level of antibodies against TCA, KETc12 exhibited almost the same sensitivity (87.5%) as TCA (93.7%) and 100% specificity. In the second panel of 110 CSF collected at random, two peptides (KETc1 and KETc12) exhibited sensitivities of 40 and 36% respectively, and were 100% specific. PMID- 10709782 TI - The fungal metabolite gliotoxin: immunosuppressive activity on CTL-mediated cytotoxicity. AB - Gliotoxin, a potential etiologic agent which is synthesized by Aspergillus fumigatus and other pathogenic fungi, exhibits a variety of immunosuppressive activities. We have found that gliotoxin markedly inhibits both perforin dependent and Fas ligand-dependent cytotoxic T-lymphocyte (CTL)-mediated cytotoxicity. Gliotoxin blocked granule exocytosis and the production of inositol phosphates in response to anti-CD3 stimulation. Apparently, activation signals were not efficiently received by the gliotoxin-treated CTL clone, perhaps because gliotoxin profoundly disturbed CTL cell attachment to immobilized anti-CD3. Although the expression of surface molecules of the CTL clone such as CD3 was unaffected by gliotoxin, the effector/target conjugate formation was inhibited dose-dependently by gliotoxin treatment of the effector CTL clone. These results suggest that gliotoxin prevents CTL from interacting with target cells. PMID- 10709781 TI - Kinetics of cellular and cytokine responses in a chimeric mouse model for the study of staphylococcal enterotoxin B pathogenesis. AB - The mechanisms by which superantigens, such as staphylococcal enterotoxin B (SEB), contribute to microbial pathogenicity have been poorly defined. The study of such pathogenic processes has been hampered by the lack of an adequate animal model. We utilized a previously described murine chimeric model to determine the cytokines and cell populations that might be involved in SEB toxicity. In the absence of bone marrow transplantation (BMT), all total body irradiated (TBI) mice died, while all transplanted mice survived up to 6 months. Compared with non TBI and non-BMT mice, chimeric mice had an increased percentage of CD11b (Mac-1) positive splenocytes (17 vs. 59%, P < 0.05) and decreased CD45R-positive (B) cells (33 vs. 6%, P < 0.05) at 6 weeks after BMT. The relative numbers of splenocyte CD4 and CD8 cells were similar in chimeric and normal mice. Susceptibility of chimeric animals to 10 or 100 microg SEB was time-dependent: no mice challenged at 2 weeks post-BMT died, but 15% of mice challenged at 4 weeks and 50% of those challenged at 6-8 weeks died. Compared with TBI and non-BMT C3H/HeJ mice, SEB-challenged chimeric mice at 6-8 weeks had (1) increased splenocyte mRNA expression for: IFN-gamma (3.5 x optimally at 1 h), TNF-alpha (6.5 x at 2 h), IL-6 (4.8 x at 4 h), IL-1beta (8.4 x at 4 h), IL-2 (4.7 x at 4 h), and IL-10 (3 x at 16 h), and (2) increased and earlier peak serum levels of IFN-gamma, IL-6, IL-1beta and IL-2, but no increase in serum TNF-alpha or IL-4. These data support the hypothesis that the decreased percentage of B cells and increased macrophages in chimeric mice lead to enhanced T cell-macrophage interactions after SEB administration and a lethal burst of T cell and macrophage cytokine release. This model will provide insight into cell populations and mechanisms that mediate superantigen-induced toxicity. PMID- 10709783 TI - CD99 monoclonal antibody induce homotypic adhesion of Jurkat cells through protein tyrosine kinase and protein kinase C-dependent pathway. AB - CD99 is a 32 kDa cell surface glycoprotein which is involved in cell adhesion. Engagement of the CD99 molecule by CD99 monoclonal antibodies has been shown to induce homotypic aggregation of various cell types. By using a newly established CD99 monoclonal antibody, MT99/3, we show here that LFA-1/ICAM-1 independent cell adhesion pathways are activated via CD99. Engagement of the CD99 molecule by MT99/3 induced homotypic cell aggregation of Jurkat T-cells within 30 min reaching its maximal level within 4 h. The Jurkat cell aggregation was not blocked by addition of CD11a (LFA-1) and CD54 (ICAM-1) mAbs. Furthermore, MT99/3 treatment did not alter the expression of LFA-1 and ICAM-1 molecules. Induction of Jurkat homotypic aggregation by MT99/3 was, however blocked by the protein kinase C inhibitor, sphingosine, the protein tyrosine kinase inhibitor, genistein, and by actin filament polymerization blocking agent, cytochalasin B. Thus, these observations suggest that CD99 can mediate beta2-integrin independent cell adhesion that depends on activation of protein kinases and reorganization of the cytoskeleton. PMID- 10709784 TI - Natural antibodies against alliinase in human serum and polyclonal antibodies elicited in rabbit share the same immunogenic determinants. AB - Human serum contains natural antibodies against alliinase, a protein abundantly found in garlic (Allium sativum) cloves. In order to study the epitope(s) of this protein recognized by anti-alliinase antibodies, we used a random hexapeptide library displayed on filamentous M13 phage. Analysis of the phagotopes selected on rabbit anti-alliinase antibodies revealed that the motif-GKXVXX- was common for all peptides. The most frequent phage displaying -GKHVAV- sequence has a 50% identity with the original alliinase sequence (amino acid residues 156-161). The position of this epitope is only nine amino acids apart from the oligosaccharide chain attached to the N146. The rabbit anti-alliinase immunoglobulin G (IgG), which bound the phages displaying this phagotope, also bound the corresponding peptide derived from the alliinase sequence. Affinity-purified natural antibodies against alliinase, present in normal human serum (which can specifically recognize the native and denaturated protein) also bound the selected phagotope. Thus, our results indicate that specific natural anti-dietary protein antibodies presented in human serum can have the same. or overlapping. epitopes with the IgG evoked during the active (experimental) immunization in animals. PMID- 10709785 TI - Ultraviolet-irradiated apoptotic lymphocytes produce interleukin-10 by themselves. AB - Since inflammatory responses are rarely associated with apoptotic cell death, it is plausible that cells undergoing apoptosis may signal the immune system to suppress inflammatory responses. By employing intracytoplasmic cytokine staining in conjunction with annexin V-binding, we examined the representative pro inflammatory cytokine tumor necrosis factor-alpha (TNFalpha) and anti inflammatory cytokine interleukin-10 (IL-10) expression in ultraviolet (UV) irradiated lymphocytes and analyzed them with apoptosis induction at a single cell level. We show here that lymphocytes exposed to UV resulted in IL-10 expression with marginal TNFalpha expression, and these IL-10-expressing cells underwent apoptosis. Addition of inhibitors for caspases blocked UV-induced apoptosis but not IL-10. These results indicate that UV elicited at least two types of signals: one which was caspase dependent, leading to apoptosis; and another which was caspase independent, leading to IL-10 production. Lymphocyte apoptosis was thus found to link anti-inflammatory cytokine secretion, and thereby may contribute to preventing unwanted immune responses. PMID- 10709786 TI - HPV type 16 protein E7 HLA-A2 binding peptides are immunogenic but not processed and presented. AB - Human papillomaviruses (HPV) have been implicated in the etiology of cervical malignancies and a high percentage of cervical carcinoma cells express HPV-16 E6 and E7 oncoproteins. These proteins are attractive targets for cytolytic T lymphocyte (CTL) mediated immunotherapy. We screened peptides derived from the HPV-16 E7 protein for binding to HLA-A2 and tested their potential to induce specific CTL responses in chimeric HLA-A2/H2-Kb transgenic mice. From eight potential binding peptides four displayed binding and were tested for immunogenicity. CTL activity was tested using target cells pulsed with peptide or expressing E7 protein. While there was no CTL induction observed with the peptides 7-15, 66-74 and 82-90, CTL from mice immunized with 86-93 lysed targets presenting the peptide in the context of the HLA-A2/H2-Kb molecule or wild-type HLA-A2. In contrast, 86-93 induced CTL showed no cytolytic activity against cells expressing the protein E7 and vaccination with the E7 protein did not lead to cytotoxicity against targets pulsed with the 86-93 peptide. Therefore the peptide 86-93, which binds to HLA-A2, is able to induce CTL responses in context of HLA A2, but the peptide appears not to be processed or presented by HPV type 16 infected cells. PMID- 10709787 TI - Establishment of efficient reaggregation culture system for gene transfection into immature T cells by retroviral vectors. AB - To overcome low efficiency of retroviral infection into immature T cells, we modified reaggregation fetal thymus organ culture by closely packed co-culture with virus-producing cells (VPC). The viral vector was constructed in chimeric vector, pMX, with IRES and tailless-rat CD2 as a surface marker of infected cells. A rearranged TCR beta gene (Vbeta8.2) was further inserted into the construct for investigating effect of the introduced gene in T cell development. Using this system, we succeeded to transfer the viral vector into immature thymocytes at a remarkably higher efficiency compared to conventional methods using medium containing retrovirus. Moreover, the introduced TCR beta gene was expressed on thymocytes of RAG2-deficient mice to induce in the transition of CD4 CD8- double-negative (DN) into CD4+CD8+ double-positive (DP) cells by transducing beta-selection signaling. Thus, our modified reaggregation culture system is useful for studying the molecular mechanism of T cell development due to a highly efficient gene transfer into immature T cells. PMID- 10709788 TI - Selective metabolism of kynurenine in the spleen in the absence of indoleamine 2,3-dioxygenase induction. AB - The kynurenine pathway of L-tryptophan degradation is differentially regulated dependent on the level of immune system activation. During inflammation and disease, activity of the hepatocellular enzyme tryptophan 2,3-dioxygenase (TDO) decreases and a second enzyme, indoleamine 2,3-dioxygenase (IDO), is induced in extrahepatic sites. Substantial formation of a metabolise downstream of this step, quinolinic acid (Quin), subsequently occurs only in select regions of the lymphoid tissues, such as spleen, in a temporally restricted manner. The goal of this study was to determine the localization of Quin in unstimulated mice under conditions where rate-limiting control of the pathway by both TDO and IDO was by passed. Supplementation of drinking water with L-kynurenine, a pathway intermediate that lies between tryptophan and Quin, resulted in a dose-dependent increase in Quin immunoreactivity in the follicles and discontinuous regions of the marginal zones of the spleen. Strongly immunoreactive cells in the periarteriole lymphoid sheaths adopted a highly reactive morphology despite the lack of immunostimulation and IDO induction. In contrast, a patchy to diffuse pallor of staining was observed in the liver parenchyma with 1 and 10 mM L kynurenine ingestion, respectively. These data show that selective tryptophan metabolism can occur in discrete subcompartments of the lymphoid tissues beyond the level of IDO. In vivo manipulation of Quin synthesis in the absence of IDO induction may serve as a model for studying regulation and function of the kynurenine pathway activation in the immune system. PMID- 10709789 TI - Dilemmas in conducting qualitative sex research in applied field settings. AB - Although resources are available to inform researchers of the many technical skills necessary to conduct qualitative research, individuals working in applied field settings often encounter ethical, moral, and sociopolitical dilemmas that cannot be resolved through the application of technical skills. The purpose of this article is to present examples of dilemmas faced by qualitative research methodologists studying sexual behavior in applied field settings. Possible solutions to these dilemmas are discussed within a theoretical and conceptual framework. The examples and discussion are organized around four broad topic areas: informed consent, privacy, confidentiality, and personal relationships. PMID- 10709790 TI - Ethical principles for analyzing dilemmas in sex research. PMID- 10709776 TI - Effects of the antifungal agents on oxidative drug metabolism: clinical relevance. AB - This article reviews the metabolic pharmacokinetic drug-drug interactions with the systemic antifungal agents: the azoles ketoconazole, miconazole, itraconazole and fluconazole, the allylamine terbinafine and the sulfonamide sulfamethoxazole. The majority of these interactions are metabolic and are caused by inhibition of cytochrome P450 (CYP)-mediated hepatic and/or small intestinal metabolism of coadministered drugs. Human liver microsomal studies in vitro, clinical case reports and controlled pharmacokinetic interaction studies in patients or healthy volunteers are reviewed. A brief overview of the CYP system and the contrasting effects of the antifungal agents on the different human drug-metabolising CYP isoforms is followed by discussion of the role of P-glycoprotein in presystemic extraction and the modulation of its function by the antifungal agents. Methods used for in vitro drug interaction studies and in vitro-in vivo scaling are then discussed, with specific emphasis on the azole antifungals. Ketoconazole and itraconazole are potent inhibitors of the major drug-metabolising CYP isoform in humans, CYP3A4. Coadministration of these drugs with CYP3A substrates such as cyclosporin, tacrolimus, alprazolam, triazolam, midazolam, nifedipine, felodipine, simvastatin, lovastatin, vincristine, terfenadine or astemizole can result in clinically significant drug interactions, some of which can be life threatening. The interactions of ketoconazole with cyclosporin and tacrolimus have been applied for therapeutic purposes to allow a lower dosage and cost of the immunosuppressant and a reduced risk of fungal infections. The potency of fluconazole as a CYP3A4 inhibitor is much lower. Thus, clinical interactions of CYP3A substrates with this azole derivative are of lesser magnitude, and are generally observed only with fluconazole dosages of > or =200 mg/day. Fluconazole, miconazole and sulfamethoxazole are potent inhibitors of CYP2C9. Coadministration of phenytoin, warfarin, sulfamethoxazole and losartan with fluconazole results in clinically significant drug interactions. Fluconazole is a potent inhibitor of CYP2C19 in vitro, although the clinical significance of this has not been investigated. No clinically significant drug interactions have been predicted or documented between the azoles and drugs that are primarily metabolised by CYP1A2, 2D6 or 2E1. Terbinafine is a potent inhibitor of CYP2D6 and may cause clinically significant interactions with coadministered substrates of this isoform, such as nortriptyline, desipramine, perphenazine, metoprolol, encainide and propafenone. On the basis of the existing in vitro and in vivo data, drug interactions of terbinafine with substrates of other CYP isoforms are unlikely. PMID- 10709791 TI - Project Northland high school interventions: community action to reduce adolescent alcohol use. AB - Project Northland is a randomized community trial initially implemented in 24 school districts and communities in northeastern Minnesota, with goals of delaying onset and reducing adolescent alcohol use using community-wide, multiyear, multiple interventions. The study targets the Class of 1998 from the 6th to 12th grades (1991-1998). The early adolescent phase of Project Northland has been completed, and reductions in the prevalence of alcohol use at the end of 8th grade were achieved. Phase II of Project Northland, targeting 11th- and 12th grade students, uses five major strategies: (1) direct action community organizing methods to encourage citizens to reduce underage access to alcohol, (2) youth development involving high school students in youth action teams, (3) print media to support community organizing and youth action initiatives and communicate healthy norms about underage drinking (e.g., providing alcohol to minors is unacceptable), (4) parent education and involvement, and (5) a classroom-based curriculum for 11th-grade students. This article describes the background, design, implementation, and process measures of the intervention strategies for Phase II of Project Northland. PMID- 10709792 TI - Results of language for health: cardiovascular disease nutrition education for Latino English-as-a-second-language students. AB - This report presents the final evaluation of Language for Health, part of a federally funded initiative to develop heart disease prevention interventions for low-literate populations. Language for Health specifically intervened with recent immigrants enrolled in English-as-a-second-language classes, incorporating nutritional behavior change materials into English-language curricula. Latino participants (n = 732) were exposed to either nutrition education or stress management classes (attention-placebo group) designed specifically for low English-literate adults. Participants completed physiological measures assessing blood pressure, total and high-density lipoprotein (HDL) cholesterol, waist and hip circumference, and weight. Self-report surveys were administered to collect students' nutrition-related knowledge, attitudes, self-reported fat avoidance behaviors, and demographic information. Data were collected at baseline, 3-month posttest, and 6-month follow-up. Results indicated long-term effects of the intervention on nutrition knowledge and fat avoidance, yet only short-term effects on total cholesterol:HDL ratio and systolic blood pressure. PMID- 10709793 TI - Teacher training as a behavior change process: principles and results from a longitudinal study. AB - For students to realize the benefits of behavior change curricula for disease prevention, programs must be implemented effectively. However, implementation failure is a common problem documented in the literature. In this article, teacher training is conceptualized as a behavior change process with explicit teacher motivation components included to help effect the intended behavior (i.e., implementation). Using this method, the Hutchinson Smoking Prevention Project, a randomized controlled trial in school-based smoking prevention, conducted 65 in-service programs, training nearly 500 teachers (Grades 3-10) from 72 schools. Implementation was monitored by teacher self-report and classroom observations by project staff. The results were favorable. All eligible teachers received training, virtually all trained teachers implemented the research curriculum, and 89% of observed lessons worked as intended. It is concluded that teacher training conceptualized as a behavior change process and including explicit teacher motivation components can promote effective implementation of behavior change curricula in public school classrooms. PMID- 10709794 TI - Positive versus negative framing of a hypothetical infant immunization: the influence of involvement. AB - Framing studies dealing with health messages show mixed results, although a tendency in favor of negative framing. Involvement has been hypothesized to account for these conflicting results. The authors selected a realistic issue (immunization of infants) deemed high or low involving depending on the respondent's circumstances: women with an infant or who were pregnant or intending to get pregnant in the next 12 months were deemed to be high involved; women in none of these categories were deemed to be low involved. A convenience sample of adult women was presented with a hypothetical "new" immunization that protected infants against respiratory complaints such as bronchitis and pneumonia Side effects (the common flu) were framed positively (90% chance of no side effects) or negatively (10% chance of side effects). The authors found positive framing to be superior for low-involved respondents, but there was no framing effect for high-involved respondents. PMID- 10709795 TI - Gimme 5 fruit, juice, and vegetables for fun and health: outcome evaluation. AB - A theory-based multicomponent intervention (Gimme 5) was designed and implemented to impact fourth- and fifth-grade children's fruit, juice, and vegetable (FJV) consumption and related psychosocial variables. Gimme 5 was a randomized controlled intervention trial with school (n = 16 elementary) as unit of random assignment and analysis. Participants included the cohort of students who were in the third grade in the winter of 1994 and students who joined them in the fourth and fifth grades. The intervention included a curriculum, newsletters, videotapes, and point-of-purchase education. Evaluation included 7-day food records and psychosocial measures from students, telephone interviews with parents, and observational assessments. Favorable results were observed for consumption of FJV combined, FJV consumed at weekday lunch, eating FJV self efficacy, social norms, asking behaviors, and knowledge. A theory-based school nutrition education program can help change children's FJV consumption and impact factors at home that predispose to FJV consumption, but changes were small, and their persistence is unknown. PMID- 10709796 TI - Mediating factors in dietary change: understanding the impact of a worksite nutrition intervention. AB - This report, based on 1,795 participants in the Next Step Trial, examines how a dietary intervention program affected mediating factors for dietary change. The model tested whether intervention increased predisposing (skills, knowledge, and beliefs) and enabling (social support and norms) factors for change and advanced participants into action and maintenance stages of change. The intervention significantly increased both predisposing factors for dietary change and the likelihood of moving into or remaining in action and maintenance stages of change. Changes in predisposing and enabling factors and stage of change at follow-up (regardless of stage at baseline) were associated with significant dietary change. Changes in mediating variables explained between 34% and 55% of the effects of the dietary intervention. These results support the value of measuring mediating factors as part of dietary intervention evaluations and suggest that interventions that target norms and eating environments in addition to skills and knowledge may further increase intervention effectiveness. PMID- 10709797 TI - Multiple-objective designs in a dose-response experiment. AB - This article is an extension of the work of Huang and Wong (1), who found dual objective designs for models with a continuous outcome. We consider quantal dose response experiments with a binary outcome and develop multiple-objective designs for two or more Bayesian optimality criteria. Using the logit model as an illustrative example, we construct numerically optimal designs for estimating model parameters and percentiles, with possibly unequal interest in each of the objectives. We also show that the popular equal dosage assignment rule can be a rather inefficient design for estimating model parameters under a Bayesian setup. PMID- 10709798 TI - Identifying the maximum safe dose: a multiple testing approach. AB - Identifying the maximum safe dose (MAXSD) is an objective of both randomized clinical dose-finding studies for the safety endpoint and toxicological studies. MAXSD is defined as the highest experimental dose with no significant increased safety effect relative to the placebo or control group. In safety assessment, the primary control of the false-negative error rate is more important than that of the false-positive rate. Therefore, we propose a multiple testing procedure for equivalence in the many-to-one design with a priori ordered contrasts (shifted control vs. dose), where the acceptable risk delta is defined in advance. Tests for shifted and ratio hypotheses are presented and discussed. PMID- 10709799 TI - Optimal sampling times in bioequivalence tests. AB - In bioequivalence studies, drug formulations are compared in terms of bioavailability parameters such as the area under the concentration-time curve (AUC), the maximum concentration (Cmax), and the time to maximum concentration (t(max)). Accuracy in measuring these parameters directly affects the accuracy of bioequivalence tests. Because the number of blood draws per patient is limited, the blood collection times must be spaced so that concentration-time curve measurements can produce accurate bioavailability parameter estimates. This paper describes an optimization approach for calculating optimal time designs for one compartment models, but is sufficiently general for other compartmental models. Simulation indicates that the optimal design improves the accuracy of AUC estimation. PMID- 10709800 TI - Designs for testing lack of fit for a nonlinear dose-response curve model. AB - We would like to estimate the parameters of a dose-response function with the greatest precision as possible. For a two-parameter model, this is equivalent to minimizing the area of the confidence ellipsoid, i.e., a D-optimal design. Previous work on this particular model has included minimal designs. These designs are unable to determine lack of fit. We introduce a distinct dose level to the design to be able to estimate the lack of fit. The minimal and new designs will be compared, and the sample size needed to achieve adequate power for the lack-of-fit test will be derived. PMID- 10709801 TI - Why are pharmacokinetic data summarized by arithmetic means? AB - The main aim of many studies in clinical pharmacology is to describe the pharmacokinetic activity of a given compound. This pharmacokinetic activity for an individual is then evaluated through a series of summary parameters, such as area under the concentration-time curve (AUC), maximum concentration (Cmax) and the rate constant lambda, and it is evaluated across individuals by descriptive statistics of these parameters, such as the mean and range and a measure of spread such as the standard deviation. How the pharmacokinetic parameters are derived is described here. It is demonstrated that the assumption of an exponential half-life is often fundamental to the derivation of pharmacokinetic parameters. Given this fact, one would think it logical that data are analyzed with the appropriate statistics on the log-scale and not by summary statistics, such as arithmetic means, on the original scale. Why arithmetic means are used to describe the data is explored and the special nature of the log-transformation highlighted. PMID- 10709802 TI - Some considerations about stability study design. AB - The aim of a stability study is to check whether or not a drug maintains its initial properties (over a given range) in a period of time. There are many parameters considered in a stability study, but in this paper only the active content will be considered. Evaluation of this parameter is performed by measuring the content of the active ingredient in the drug at different times and on expiration to see if it is within specification (1). The problem also involves estimating whether the drug will perform satisfactorily and maintain its initial properties in the future. In this paper, we provide a criterion for the evaluation of experimental design used in a stability study for predicting the content of an active ingredient in a drug, starting from the confidence limit calculated following International Conference on Harmonisation of Technical Requirements for the Registration of Pharmaceuticals for Human Use (ICH) recommendations. The point of view adopted is akin to hypothesis testing. The main question is this: "If the drug fulfills the requirements at a given time in the future, what is the probability that the data collected for that drug will show its suitability?" For this purpose, two concepts are used-producer gain and consumer loss-which are defined as follows: producer gain is the advantage (for the producer) of recognizing a drug as "good" (provided it is good and within specification); consumer loss is the loss (for the consumer) when the drug is no longer good, but the data collected indicate that it is good. The aim of the experimental design in this study is to increase the probability of producer gain and maximize it on the basis of a given consumer loss. PMID- 10709803 TI - Setting specifications for non-normally distributed data. AB - In this paper, we focus on estimating an upper specification for non-normally distributed data. Cornish-Fisher expansions were exploited for derivations of the method. The method was applied for upper specification calculations of two particle-size data sets, and simulation studies were also performed to demonstrate the accuracy of the approximations. The method was also used to calculate the critical values for delta(min) for the Anderson-Hauck test for equivalence using bootstrap samplings. The main advantage of the method is in calculating specifications with limited data. PMID- 10709804 TI - A comparison of urn designs for randomized clinical trials of K > 2 treatments. AB - Response-adaptive designs in clinical trials involve incorporating accruing information from patient responses to treatment into the randomization scheme in order to assign more patients to the treatment that has performed better in the trial up to that point. One probability model useful in generating an adaptive randomization scheme is an urn model. We will give a short overview of such adaptive models and compare four of them. We will be interested in how these four models minimize the number of treatment failures in a clinical trial with dichotomous response treatments. Comparison will be done via simulations for four treatments and exactly for three treatments for moderate sample sizes. We compare designs under the assumption that the results of treatments are known immediately, and we also allow some delay in response. Power is analyzed under various alternatives. Our results indicate that a birth and death urn with immigration is the best unless success probabilities are very small, in which case a randomized version of Polya's urn is preferred. PMID- 10709805 TI - Conditional and exact tests in crossover trials. AB - Generalized linear models are developed for crossover trials with no carryover effects and fixed subject effects. A general multinominal model for the distribution of data is considered. This subsumes both binary and categorical data. Conditional inferences eliminate subject effects by conditioning on their sufficient statistics. For normal data, least-squares analysis is exact with identical treatment inferences from unconditional and conditional analyses. For Poisson data, unconditional and conditional analyses are also identical, but for multinomial data this is not the case and the unconditional analysis is invalid. For multinomial data, asymptotic tests of both treatment effects and goodness of fit are unreliable with small samples. Procedures for exact tests are developed to overcome such problems, using enumeration, random sampling, and a hybrid of importance sampling and enumeration. A four-period binary crossover trial is used to illustrate an exact test of treatment effects by a two-stage sampling procedure based on a factorization of the conditional distribution of the sufficient statistics. An exact test of goodness of fit on the same data illustrates a two-stage scheme mixing importance sampling and enumeration. PMID- 10709806 TI - EEG and functional neuroimaging. PMID- 10709807 TI - Neurophysiologic basis of functional neuroimaging: animal studies. AB - Functional neuroimaging adds metabolic or biochemical information to that obtained with anatomic imaging, allowing localization of a neural function. Positron emission tomography and single photon emission tomography make use of radioactive tracers tagged to a molecule which can indicate glucose metabolism, oxygen consumption, or blood flow. Functional magnetic resonance imaging uses the different magnetic properties of oxyhemoglobin and deoxyhemoglobin to identify areas of increased blood flow, which, in turn, reflects neuronal activation. Magnetic resonance spectroscopic imaging, with magnetically labeled molecules, can be used to follow biochemical pathways. Functional neuroimaging is based on the experimental data that neuronal activation leads to increased metabolism. Uptake of glucose and oxygen increases to meet increased energy needs. The fractionally increased glucose appears to be taken up mostly by glia, which metabolize it through glycolysis. The end product, lactate, is released for neuronal uptake and subsequent oxidative phosphorylation. To meet these metabolic needs, blood flow increases to such an extent that overall capillary oxyhemoglobin concentration increases. This changes the magnetic signal in the region and permits functional magnetic resonance imaging studies. Recent data suggest that there is an initial decrease in the concentration of oxyhemoglobin which may be more spatially specific to the area of neuronal activation. Further refinements in functional neuroimaging will lead to improved understanding of the normal functional anatomy of the brain and will shed further light on the pathophysiology of many neurologic disorders. PMID- 10709808 TI - The EEG evaluation of single photon emission computed tomography abnormalities in epilepsy. AB - Single photon emission computed tomography (SPECT) has increasingly been used as a diagnostic procedure for localizing epileptic seizure foci and as a research tool for investigating the physiologic mechanisms underlying seizure activity. With increasing use of SPECT in localizing the seizure focus for epilepsy surgery, there arises a need to critically assess its current role in the evaluation of patients for epilepsy surgery, especially as it relates to other clinical and laboratory data used in presurgical evaluation. Ictal EEG discharge has traditionally been used as the "gold standard" against which SPECT studies are compared in assessing the latter's localizing value. However, this practice presents a major challenge because SPECT studies are often reserved for patients with nonlocalizing EEG or magnetic resonance imaging findings. Nonetheless, SPECT studies in evaluation for epilepsy surgery should always be performed with the knowledge of the patient's EEG activity preceding, during, and after the injection of the radiotracer. The advent of techniques such as subtraction SPECT with co-registration on magnetic resonance imaging (SISCOM) and computer image guided surgery has great potential in enhancing the clinical electrophysiologic evaluation of SPECT-detected abnormalities in epilepsy. These techniques permit accurate spatial correlation between intracranial EEG activity and SPECT perfusion patterns. The techniques can also be used to evaluate the effect of the extent of EEG focus resection compared with that of SISCOM focus resection to determine which has more prognostic importance in postsurgical control of seizures. Both animal and human studies are warranted to advance our knowledge of the electrophysiology associated with the various SPECT perfusion patterns. PMID- 10709810 TI - EEG-Linked functional magnetic resonance imaging in epilepsy and cognitive neurophysiology. AB - The ability to trigger functional magnetic resonance imaging (fMRI) acquisitions related to the occurrence of EEG-based physiologic transients has changed the field of fMRI into a more dynamically based technique. By knowing the temporal relationship between focal increases in neuronal firing rates and the provoked focal increase in blood flow, investigators are able to maximize the fMR-linked images that show where the activity originates. Our mastery of recording EEG inside the bore of a MR scanner has also allowed us to develop cognitive paradigms that record not only the fMR BOLD images, but also the evoked potentials (EPs). The EPs can subsequently be subjected to localization paradigms that can be compared to the localization seen on the BOLD images. These two techniques will most probably be complimentary. BOLD responses are dependent on a focal increase in metabolic demand while the EPs may or may not be related to energy demand increases. Additionally, recording EPs require that the source or sources of that potential come from an area that is able to generate far-field potentials. These potentials are related to the laminar organization of the neuronal population generating that potential. As best we know the BOLD response does not depend on any inherent laminar neuronal organization. Therefore, by merging these two recording methods, it is likely that we will gain a more detailed understanding of not only the areas involved in certain physiologic events, e.g. focal epilepsy or cognitive processing, but also on the sequencing of the activation of the various participating regions. PMID- 10709809 TI - Relationship between EEG and positron emission tomography abnormalities in clinical epilepsy. AB - Positron emission tomography (PET) is a relatively noninvasive neuroimaging method by means of which a large variety of human brain functions can be assessed. Localized neurochemical abnormalities detected by PET were found in patients with partial epilepsy and suggested the use of this modality for localizing epileptogenic regions of the brain. The clinical usefulness of PET is determined by its sensitivity and specificity for identifying epileptogenic areas as defined by ictal surface and intracranial EEG recordings. The findings obtained from comparative EEG and glucose PET data are reviewed with special emphasis on patients undergoing presurgical evaluation because of medically intractable temporal and extratemporal lobe epilepsy. The utility of glucose PET studies for identifying regions of seizure onset is presented, and the limited specificity of glucose metabolic abnormalities for the detection of various EEG patterns in clinical epilepsy is discussed. The authors review the available intracranial EEG and PET comparisons using [11C]flumazenil (FMZ) PET, a tracer for the assessment of tau-amino-butyric acid/benzodiazepine receptor function. They also summarize their experience with [11C]flumazenil PET in identifying cortical regions that show various ictal and interictal cortical EEG abnormalities in patients with extratemporal seizure origin. Finally, the authors demonstrate that further development of new PET tracers, such as alpha [11C]methyl-L-tryptophan, is feasible and clinically useful and may increase the number of patients in whom PET studies can replace invasive EEG monitoring. PMID- 10709812 TI - Pattern-evoked potential latencies from central and peripheral visual fields. AB - Median P100 and N70 latencies for peripheral field (>8 degrees) TV-generated pattern visual-evoked potentials were 6 and 8 milliseconds less than for central field patterns in subjects with normal pattern visual-evoked potentials. These differences held for patient subgroups with P100 latency maxima from 140 to 100 milliseconds, and for left and right eye data separately compiled. Latencies for central field stimulation exceeded those for peripheral field stimulation in 233 (85%) of 274 eyes for the N70 potential and in 210 (77%) of 274 eyes for the P100 potential. Such data suggest that the faster conducting peripheral visual system, conveying location and motion information, prepares the occipital cortex for the later arriving central data conveying more intricate details. PMID- 10709811 TI - An international survey of long-term video/EEG services. AB - To determine current practices in the provision of video-EEG services, the authors conducted an international survey by post. The aim of the survey was to evaluate, by reference to other centres, how and why certain things are done, be assured that their own center is providing a quality service, identify weaknesses in their service, and from this, set improvement goals and objectives. A purposive sampling method was used by sending questionnaires to 78 hospitals where it was believed a long-term video-EEG monitoring service existed. Completed survey questionnaires were returned from 42 centers. Although the survey mechanism may have resulted in self-selection bias, evaluation of the responses provides information on patient management, staffing levels, equipment, and equipment management. Ultimately, these data may aid in identifying a minimum set of requirements for the provision of a video-EEG telemetry service. PMID- 10709813 TI - The macrostructure and microstructure of sleep in patients with autosomal dominant nocturnal frontal lobe epilepsy. AB - The electroclinical features of autosomal dominant nocturnal frontal lobe epilepsy have been recently described. Although some patients reported a poor quality of sleep, daytime tiredness, and sleepiness, their sleep macrostructure appeared to be indistinguishable from those of the control group. The aim of this study was to evaluate the macro- and microstructure of sleep in a sample of autosomal dominant nocturnal frontal lobe epilepsy patients, diagnosed by videopolysomnography. The authors selected 16 patients, 8 with daytime complaints (morning tiredness and/or excessive sleepiness) (group 1) and 8 without those complaints (group 2). The classical macrostructure of sleep and the microstructure, according to the cyclic alternating pattern (CAP) scoring rules, were compared with another group of 8 healthy controls. In group 1 the motor attacks during sleep took place more frequently during CAP and were significantly related to phase A of the CAP cycle in comparison to group 2 (P = 0.04). Group 2 had a sleep microstructure similar to the controls, whereas group 1 showed higher CAP/nonrapid eye movement sleep (CAP rate) and a higher number of CAP cycles with respect to controls (P = 0.012 and P = 0.001) and to group 2 (P = 0.05 and P = 0.04). The analysis of sleep microstructure showed an increase in sleep instability in patients with autosomal dominant nocturnal frontal lobe epilepsy and daytime sleep complaints and indicated the relationship between sleep fragmentation, nocturnal motor seizures, and daytime symptoms. PMID- 10709814 TI - Using time domain characteristics to discriminate physiologic and parkinsonian tremors. AB - Tremor amplitude and frequency do not always clearly differentiate subjects with particular pathologies from control subjects or from subjects with other pathologies, especially in early stages of a disease. For patients with early stages of Parkinson's disease (PD) the discriminative power of amplitude was compared with that of other time domain characteristics of tremor recordings that are probably not evident clinically. Postural tremor with and without visual feedback and rest tremor were recorded in both hands of a group of patients with Parkinson's disease (n = 21) and a group of healthy control subjects (n = 30) using displacement lasers. Velocity and acceleration data were derived from displacement data. Twelve time domain characteristics were calculated on each recording and the discriminating power of each was evaluated using the worse hand in each case. Postural tremor with no visual feedback separates the two groups of subjects most efficiently, especially in velocity and acceleration. Tremor in Parkinson's disease (in comparison to normal physiologic tremor) has a specific morphology, has a distinctive histogram, is more periodic, and contains indications of nonlinearity in the underlying dynamics. There may also be greater difference in amplitude between the two hands and time asymmetry in tremor of patients with PD. A series of finger flexions seems to enhance normal tremor but not tremor in PD and may thus aid in discrimination. Discrimination of tremor attributable to PD from normal physiologic tremor can be enhanced by measuring time domain characteristics subtler than amplitude, particularly when amplitude itself is not large. Tremor measurement should not be limited to acceleration data because some information is more visible in other variables. PMID- 10709815 TI - Computer-assisted EEG monitoring during carotid endarterectomy. AB - The purpose of this study was to develop a reliable method of EEG analysis during carotid endarterectomy. EEGs of 104 patients under general anesthesia were processed by three different methods: a) "on-line" visual analysis during surgery, b) "off-line" visual analysis in laboratory, and c) computer analysis. To identify pathological EEGs, variability and asymmetry indexes of the 0.5-3.5 Hz and 8-15 Hz bands, absolute power and variability indexes of spectral edge frequency (SEF), and main dominant frequency were evaluated. On-line visual analysis showed clamp-related modifications in 29 EEGs (27.9%). Off-line visual analysis detected 24 pathological EEGs (23.1%): 18 with major changes and 6 with moderate changes. Computer analysis showed 21 EEGs (20.19%) with at least one altered index and 7 (6.7%) with altered variability for both SEF and 8-15 Hz power. The statistical analysis was significant for SEF variability and for 8-15 Hz power variability and asymmetry (P < 0.0001, analysis of variance test). While SEF and 8-15 Hz power variability did not appear influenced by anesthesia and single electrode artifacts, 8-15 Hz power asymmetry index was confounded by the presence of contralateral internal carotid occlusion. The data show that the use of these spectral indexes adds objective information to visual analysis, supporting and making easier intraoperative strategies. Their routine clinical use does not involve additional costs remaining technical requirements unchanged compared to traditional recording. PMID- 10709816 TI - The last word. PMID- 10709817 TI - Results of right coronary artery endarterectomy with or without patchplasty. AB - We report preoperative and early postoperative findings of 286 coronary bypass patients operated between 1988 and 1998 who had endarterectomy and/or patchplasty to the right coronary artery. In this retrospective study there were 61 cases with only saphenous vein patchplasty to the right coronary artery (patch group), 57 patients who underwent endarterectomy and patchplasty (open-patch group), and 229 patients having closed endarterectomy to the right coronary artery (closed group). A group of 150 patients having a saphenous vein graft to the right coronary artery without endarterectomy were chosen as a control group. Gender, age, family history, smoking history, diabetes, hyperlipidemia, hypertension, nature of the angina, severity of the coronary artery disease, left ventricular functions, preoperative rhythm, and electrocardiographic patterns were evaluated for their effect on early mortality among groups. No significant difference was detected. Positive inotropic and mechanical support need was higher in the closed group at the end of the operation and in the intensive care unit. Duration of cardiopulmonary bypass and clamp time was higher in the open-patch group. Atrial fibrillation in the early postoperative period was more frequent in the patch and closed groups. Complete atrioventricular block development and the need for a pacemaker were higher in the open-patch and closed groups. Non-Q wave myocardial infarction was more frequent in the closed group. Mortality rates were higher in the open-patch and closed groups. We conclude that endarterectomy to right coronary artery should be avoided if possible, and patchplasty with saphenous vein should be preferred. PMID- 10709818 TI - Complete repair of Tetralogy of Fallot with absent pulmonary valve including the role of airway stenting. AB - Tetralogy of Fallot (TOF) with absent pulmonary valve (APV) represents an extreme form of tetralogy where pulmonary insufficiency and mild annular stenosis often results in massive pulmonary arterial (PA) dilatation. The aneurysmal left and right PAs often compress the adjacent trachea and bronchi, leading to airway obstruction and respiratory failure in infancy. Between 1991 and 1997, 11 patients underwent a single stage repair of TOF and APV using a valved (10 patients) or nonvalved (1 patient) homograft conduit and PA reduction arterioplasty. There was one (1/11 [9.1%) perioperative and one (9.1%) late death. Both deaths were related to airway complications. Morbidity associated with postoperative respiratory complications and ventilator-dependency due to underlying tracheobronchomalacia is an important problem. Intermediate follow-up shows a high incidence of reintervention for conduit stenosis and/or insufficiency and tracheobronchial compression. These infants also required multiple hospitalizations for recurrent respiratory infections secondary to their tracheobronchomalacia. Stenting of the right and left main bronchi with balloon expandable metallic stents is a new experimental therapy that has been useful in two recent patients with respiratory failure despite satisfactory intracardiac repair. It may provide an attractive alternative therapy to prolonged mechanical ventilation with positive end expiratory pressure in patients with severe tracheobronchomalacia. Complete repair with a valved homograft conduit and reduction pulmonary arterioplasty in infancy at the time of diagnosis is the procedure of choice for infants with TOF with APV. With this approach the patient outcome is essentially determined by their airway status and airway management. PMID- 10709819 TI - Reduction of blood loss and transfusion requirements after coronary artery bypass grafting: similar efficacy of tranexamic acid and aprotinin in aspirin-treated patients. AB - BACKGROUND: In patients with coronary artery disease, continuation of aspirin may reduce the incidence of unstable angina and preoperative myocardial infarction before surgery, but the risk of perioperative bleeding may be increased. METHODS: The efficacy of aprotinin and tranexamic acid (TXA) was examined in a prospective, randomized, double-blind trial involving 56 patients scheduled for coronary artery bypass grafting and who received aspirin 100 mg/day until the day of the operation. Group I received high-dose aprotinin whereas group II received 10 g of tranexamic acid (TXA) over 20 minutes before sternotomy. Heparinization during cardiopulmonary bypass was controlled with HDTT (high-dose thrombin time) to eliminate interference of aprotinin on ACT (celite activated clotting time). Postoperative blood loss and transfusion requirements were registered during the first 24 hours. RESULTS: The demographics, coagulation, and intraoperative parameters were similar in both groups. Postoperative blood loss (aprotinin 840 mL /24 hours, TXA 880 mL/24 hours, p = 0.481), and transfusion requirements (2.18 units/patient in the aprotinin group, 2.11 units/patient in the TXA group) were not remarkably different between the two regimen protocols. No perioperative myocardial infarction, pulmonary embolism, cerebrovascular event, or other thrombotic events were observed. CONCLUSIONS: In this trial, we were not able to demonstrate any difference in postoperative bleeding in patients pretreated with aspirin after high-dose aprotinin or TXA. From a practical point of view, TXA is safe, less expensive than aprotinin, and easy to handle, and can be recommended in patients pretreated with aspirin to improve postoperative hemostasis. PMID- 10709820 TI - Surgical treatment of vascular ring including right cervical aortic arch. AB - Cervical aortic arch is a developmental entity consisting of persistence of the right or left third branchial arch and regression of the fourth branchial arches. In most cases, the aorta is redundant and crosses behind the esophagus to the opposite side. In the presence of an aberrant subclavian artery contralateral to the side of the aortic arch and a ligamentum arteriosum, a vascular ring is formed around the trachea and esophagus. Two young patients with right-sided cervical aortic arch, aberrant left subclavian artery, and ligamentum arteriosum presented with dysphagia and exertional dyspnea. In one patient, through a left thoracotomy, the ligamentum arteriosum was divided, and the trachea and esophagus were dissected thoroughly above and below the level of the ring. In addition, the aberrant left subclavian artery was divided at its origin from a large diverticulum and implanted into the left common carotid artery; the aortic diverticulum was resected. In the other patient, who had associated 22q11 chromosomal deletion, in addition to left-sided compression of the trachea and esophagus, there was additional marked compression of the right anterolateral trachea by the redundant ascending aorta. Through a median sternotomy, the ligamentum arteriosum was divided, and the trachea and esophagus were widely mobilized; an additional aortopexy of the ascending aorta to the right of the sternum resulted in the absence of tracheal compression. The cases of the two reported patients illustrate the clinical variability of vascular ring, including a right cervical aortic arch and the consequently versatile surgical approach that is needed to successfully address this combination of vascular anomalies. PMID- 10709821 TI - Partial maze procedure is effective treatment for chronic atrial fibrillation associated with valve disease. AB - The maze procedure may be performed in combination with valve operations to treat chronic atrial fibrillation associated with valve dysfunction. Although we initially used the modified Cox maze III procedure, a more limited partial maze procedure is now preferred because the left atrium might be considered as the electrical impetues for atrial fibrillation. In this study we compared the results of 30 patients (group I) who underwent the full biatrial modified Cox maze III and 20 (group II) patients the partial maze procedure. While the rates of restored sinus rhythm were the same in both groups at 6-month follow-up (I: 83.3%, vs II: 80%), the following advantages were noted in the patients undergoing the partial maze procedure: shorter operative times, lesser elevations of creatine phosphokinase, lower rate of blood transfusion, lower rate of junctional rhythm soon after the operation, and a higher P wave in those patients with restored sinus rhythm. The effectiveness of the partial maze procedure seems equal to that of the biatrial modified Cox maze III procedure for atrial fibrillation associated with valve disease. The partial maze procedure is simple and less invasive, and thus might be applied more frequently as an additional procedure to valve operations without additional risk. PMID- 10709822 TI - Mitral valve replacement in patients after aortic valve replacement. AB - BACKGROUND: Mitral valve replacement in patients who previously had undergone aortic valve replacement is a technical challenge. The rigid aortic prosthesis limits visualization of the anterior mitral annulus and placement of sutures. METHODS: Reoperative mitral valve replacement was performed in five patients after aortic valve replacement. Two patients underwent resternotomy to allow verification of normal aortic prosthetic valve function. Anterolateral right thoracotomy was used for reentry in the remaining three patients. Exposure of the anterior mitral annulus was accomplished by initial traction on the intact anterior leaflet, with resection of this leaflet only after placement of sutures. RESULTS: All patients survived the surgical procedure and are well 2 to 30 months after operation. In one patient it was impossible to open one cusp of the mitral prosthesis, nor was it possible to rotate the valve. The valve was reimplanted, but sutures were tied only after testing for full free cusp motion. CONCLUSIONS: When appropriate, right thoracotomy incision offers excellent exposure of the mitral valve with minimal dissection. Placement of sutures along the anterior portion of the annulus is facilitated by traction downwards on the anterior leaflet. Full range of motion of the prosthetic cusps should be verified before tying the sutures. PMID- 10709823 TI - Approach to the mitral valve through a right thoracotomy in potentially hazardous reoperation. AB - BACKGROUND: Repeat sternotomy for mitral valve surgery may be hazardous in some patients. A right thoracotomy avoids the densely scarred area beneath the sternum and provides adequate in-line exposure of the mitral valve. METHODS: Between 1994 and 1997, five patients were reoperated for a mitral valve or prosthesis dysfunction through a right thoracotomy. Indications were three second redo mitral valve surgeries and two first redo, once in a patient with an aortic prosthesis and once in a patient with patent aortocoronary grafts. The operation was performed without clamping the ascending aorta in moderate hypothermic (four patients) or normothermia (one patient). RESULTS: Exposure of the mitral valve for replacement (four patients) or for repair of a paraprosthetic leak (one patient) was optimal in all patients. Resumption of cardiac function occurred rapidly after repair without specific support. Postoperative course was uncomplicated. Blood loss ranged from 300 to 700 mL. Patients were discharged from 7 to 12 days postoperatively. They are in New York Heart Association (NYHA) functional Class I (four patients) and II (one patient), from 3 to 42 months postoperatively. CONCLUSION: Right thoracotomy provides a direct "in the line of vision" access to the mitral valve. Because complete de-airing of the heart is difficult and respiratory function depressed after a right thoracotomy, this approach seems suitable when technical difficulties are expected in sternal reopening. PMID- 10709825 TI - New horizons on the surgical treatment of dilated cardiomyopathy. PMID- 10709826 TI - The Batista operation in patients with dilated cardiomyopathy. AB - Between December 1996 and October 1998, 34 patients with nonischemic dilated cardiomyopathy (DCM) received cardiac volume reduction surgery. The patients' ages ranged from 14 to 67 years (mean = 48 years) and included 28 males and 6 females. Associated mitral regurgitation was present in 31 patients, tricuspid regurgitation in 19 patients, and aortic regurgitation in 4 patients. We performed a partial left ventriculectomy (PLV) using antegrade intermittent warm blood cardioplegia in 15 patients (group A), and in 19 patients (group B) PLV was performed using the on-pump beating heart technique. In group A, the mean aortic clamping time was 79+/-33 minutes and the total cardiopulmonary bypass time was 155+/-58 minutes. In group B the mean cardiopulmonary bypass time was 121+/-43 minutes. There were eight hospital deaths (five in group A and three in group B). Five of 10 survivors of group A required inotropic support for 13.8+/-25.3 days after the operation, while 5 of 12 survivors in group B required inotropes for 4.2+/-3.1 days. Hospital mortality was 86% in 7 emergent cases and 7% in 27 elective cases. Echocardiographic study showed that the left ventricular ejection fraction improved from a mean of 18.7% to 30.3% and the left ventricular diameter decreased from a mean of 80.2 mm to 62.3 mm after the operation. All 26 hospital survivors were followed for 1 to 20 months. Three patients died at early follow up because of congestive heart failure, thrombosed valve, and hepatic failure, respectively. Nineteen patients were in New York Heart Association (NYHA) Class I or II and four were in NYHA Class III. In conclusion, cardiac volume reduction surgery is effective when the operative technique and proper judgment of patient selection are established, and emergent operation is avoided. PMID- 10709824 TI - Surgery for aortic regurgitation caused by Behcet's disease: a clinical study of 11 patients. AB - Surgical treatment for aortic regurgitation (AR) caused by Behcet's disease is difficult due to the need to manipulate fragile, inflamed tissue. Valve detachment following aortic valve replacement (AVR) and suture detachment are serious postoperative complications. We investigated the surgical results in 11 patients. Between 1981 and July 1999, 11 patients, 9 males and 2 females, with AR caused by Behcet's disease underwent surgery. The age of these patients ranged from 33 to 60 years (mean, 45+/-8 years). The surgical procedures for AR were AVR in six patients and valved conduit operation in five patients. No patient died during the hospital stay. In a follow-up period ranging from 3 to 204 months (mean, 93+/-64 months) two patients died. Prosthetic valve detachment or suture detachment necessitating redo-operation occurred in four patients (36%) who then underwent a valved conduit procedure as a reoperation. Prosthetic valve detachment was higher in patients with AVR than in patients with a valved conduit operation. Valved conduit reconstruction is indicated in patients with AR caused by Behcet's disease in whom prevention of valve detachment is difficult even by current valve fixation methods. PMID- 10709827 TI - Results after partial left ventriculectomy in a European heart failure population. AB - BACKGROUND: Forty-nine consecutive patients undergoing partial left ventriculectomy (Batista) surgery between January 1995 and June 1998 were studied. METHODS: Patient ages ranged from 12 to 85 years, and all patients were in New York Heart Association functional Class III or IV. Thirty-three patients had ischemic cardiomyopathy, and 16 had idiopathic myopathy. Inclusion criteria were left ventricular end diastolic volume index of > 150 mL/m2, left ventricular ejection fraction of < 20%, or left ventricular end-diastolic diameter of > 70 mm. Sixteen patients were transplant candidates. Partial left ventriculectomy and mitral valve repair by means of a Cosgrove annuloplasty ring plus the Alfieri repair constituted only part of the complex cardiac reconstruction in 38 patients. RESULTS: Five patients died early and five patients died late between 3 and 30 months postoperatively. The actuarial 1-year survival rate was 81%. Twenty seven patients with coronary artery disease underwent one to five bypass grafts when appropriate. In addition, three patients received aortic valve replacement, four received tricuspid valve repair, two received mitral valve replacement, and two underwent dynamic cardiomyoplasty. Left ventricular (LV) diameter could be reduced from a preoperative mean of 71 to 56 mm postoperatively. LV ejection fraction increased to 36% postoperatively. Ninety percent of patients are in New York Heart Association functional Class I or II. CONCLUSIONS: Patients with end stage idiopathic or ischemic cardiomyopathies can be improved considerably with partial left ventriculectomy. Any cardiac comorbidity should be repaired simultaneously. PMID- 10709828 TI - Perioperative care in left ventricular volume reduction. AB - BACKGROUND: While the operative technique of left ventricular volume reduction (LVVR) is rapidly becoming standardized, the optimal perioperative management strategy is yet to be determined. We present our experience with the care of patients undergoing LVVR. METHODS: LVVR was performed in 21 patients (mean age = 65.5 years) with congestive heart failure. Our management strategy was initially based on afterload reduction with sodium nitroprusside, but was later modified to include routine preoperative intra-aortic balloon support, normothermic cardiopulmonary bypass, antegrade intermittent warm blood cardioplegia, and postoperative support with phosphodiesterase-III inhibitors. Hemodynamic manipulations are aimed to attain systemic vascular resistance between 600 and 800 dyne/sec per cm(-5) and the lowest mean blood pressure that is able to maintain satisfactory systemic perfusion. Postoperatively, aggressive antifailure medical therapy with a high dose of angiotensin converting enzyme inhibitors, nitrates, and diuretics was initiated early and maintained indefinitely. RESULTS: Using this approach, postoperative progress was characterized by hemodynamic stability. IABP support was used for 59.6+/-9 hours following surgery, and inotropic support for 103+/-12 hours. In our series there were four (19%) in hospital deaths, two of which were related to heart failure. CONCLUSION: The described approach is associated with an acceptable early outcome. However, further advances in myocardial protection methods and pharmacological and mechanical support techniques are necessary for a wider adoption of this procedure. PMID- 10709829 TI - Myocardial inflammatory cell infiltrates in cases of dilated cardiomyopathy: light microscopic, immunohistochemical, and virological analyses of myocardium specimens obtained by partial left ventriculectomy. AB - OBJECTIVE: Partial left ventriculectomy was introduced for the treatment of refractory dilated cardiomyopathy (DCM). To determine the presence and degree of inflammatory cell infiltrates in DCM and the correlation between the underlying myocardial injury and early clinical outcomes after the operation, we performed histopathological, immunohistochemical, and virological studies of the resected myocardium. METHODS: Posterolateral walls of the left ventricle from 13 idiopathic DCM patients (9 males and 4 females; mean age = 53+/-14 years) were examined. Qualitative and quantitative analyses of the interstitial fibrosis and of the infiltrating inflammatory cells were conducted. For the immunohistochemistry, leukocyte surface markers and antibodies to adhesion molecules and cytokines were used. The histopathological findings were compared with the clinical results, including outcome within 1 year, and pre- and postoperative hemodynamic data. Genomic analysis of the myocardium with polymerase chain reaction was performed for enterovirus, mumps, influenza A, cytomegalovirus, and hepatitis C virus. RESULTS: (1) The three patients who died of cardiac insufficiency after surgery had a higher count of infiltrating inflammatory cells than the eight survivors (32.1+/-10.4 vs 16.3+/-11.9 cells/mm2, p = 0.07). The severity of interstitial fibrosis (percent fibrosis) did not differ significantly between these two groups (28.3+/-15.0 vs 24.0+/ 11.7%). (2) In patients who died of myocardial dysfunction, focal accumulations of lymphocytes were common, in which cytotoxic/suppressor T cells and helper/inducer T cells were observed. (3) Enterovirus genome was detected in the myocardium of two patients, both of them died after surgery. CONCLUSIONS: Inflammatory cell infiltrates or active myocarditis appear in some cases to play an important role in the etiology and pathophysiology of clinically diagnosed DCM. There is a possibility that those patients with a more severe or ongoing inflammatory process might have poor outcomes after partial left ventriculectomy. PMID- 10709830 TI - Heart transplantation in perspective. AB - Heart disease remains one of the leading causes of death in the western world. In the 35 years since the first human heart transplants, cardiac transplantation has become established as the therapeutic option of choice in the management of terminal cardiac failure. Since 1981, the introduction of cyclosporin for immunosuppression has dramatically increased cardiac transplantation. However, several obstacles limit further utilization, including limited availability of donor hearts, limited ischemic time tolerated by donor hearts, and chronic rejection. Research is underway into donor heart preservation and new immunosuppressant drugs in an effort to increase donor organ availability. Due to these constraints, alternative therapies are under development. More than 2,000 circulatory assist devices have been implanted with >25% used as a bridge to heart transplantation. The University of Ottawa Heart Institute began the first Canadian implantation of circulatory assist devices in 1986 and has implanted 23 total artificial hearts and 23 ventricular assist devices. The Heart Institute is also developing a totally implantable electrohydraulic ventricular assist device (EVAD) for long-term mechanical support outside the hospital. Another alternative being evaluated for clinical use is xenotransplantation. The major obstacle for widespread use of clinical xenotransplantation remains graft rejection, and fundamental research is ongoing to address hyperacute and delayed xenograft rejection. While cardiac transplantation is the most effective treatment of terminal heart failure, limited donor hearts compel us to rely on alternatives. In the future, the research underway on xenotransplantation and mechanical circulatory assist devices will provide new options for the clinical treatment of terminal cardiac failure. PMID- 10709831 TI - Intraoperative assessment of acute hemodynamic changes after partial left ventriculectomy. AB - Partial left ventriculectomy (PLV) has been introduced as an alternative surgical therapy for patients with end-stage dilated cardiomyopathy. The physiological benefits of PLV are relatively unknown. Therefore, the objective of this study was to determine the acute effects of PLV by measuring cardiac function before and after PLV. Aortic and left ventricular pressures and aortic flow were measured in eight patients. Continuous, beat-to-beat data were recorded and compared pre-PLV and post-PLV with and without inferior vena cava (IVC) occlusions. PLV increased cardiac output (0.93+/-0.5, p = 0.01) as a result of increased stroke volume (5.12+/-4.24, p = 0.06) and heart rate (14.5+/-8.44, p = 0.02). Contractility (+/- dP/dt, 240.33+/-74.28, p = 0.001) and external work (650.8+/-320.4, p = 0.01) were also improved. Left ventricular end-diastolic elastance (0.15+/-0.14, p = 0.10) nearly doubled after PLV. Our results indicated an improved cardiac function as measured by increased cardiac output, stroke volume, ejection fraction (EF), and contractility. PMID- 10709832 TI - Doppler frequency shift and time-domain velocity enhancement induced by an ultrasonographic contrast agent. AB - The aim of this study was to evaluate in rabbit aorta the effect of three bolus doses of Levovist on velocity values measured with spectral Doppler sonography and with time-domain correlation method (color velocity imaging). At each step, a mean peak systolic velocity was calculated from five measurements. These measurements were taken before injection, at 20 s after, at every 30 s till the third minute, and at every minute until return to peak systolic velocity at baseline value. Total duration of enhancement was noted after each injection. After each injection, once the systolic velocity values return to baseline values, a 3 min delay was observed before the following intravenous contrast agent injection was done. With Doppler spectral analysis, after the first injection, peak systolic velocity enhancement was 15 +/- 8.4% (5 to 28%), with a 6.4 +/- 4.3 min duration. After the second injection, peak systolic velocity enhancement was 15.8 +/- 8.4% (5 to 28%) with an 8.8 +/- 4 min duration. After the third injection, it was 14 +/- 9.8% (5 to 34%) with a 13.6 +/- 7.6 min duration (P = 0.04). Peak systolic velocity measured with color velocity imaging remained unchanged after every injection. Doppler velocities were increased by a bolus injection of a contrast agent. Amplitude was not cumulative with the number of injections but was cumulative on its duration. Velocity measurement with time domain correlation was not influenced by repeated injections. PMID- 10709833 TI - Dilatation of the biliary tree in children: sonographic diagnosis and its clinical significance. AB - We evaluated sonographically 162 children who met the criteria for biliary tract dilatation in the past 18 years. Of these, 131 patients were diagnosed as having anomalous dilatations of the biliary tree (including 112 with choledochal cysts and 19 with biliary duct dilatation and biliary atresia). Biliary tract dilatations in the other 31 patients were due to secondary causes or normal variants. All cases of intrahepatic biliary tree dilatation and those with both intra- and extrabiliary duct dilatations were anomalous. In 117 cases of extrahepatic biliary tract dilatation only, the mean diameter was widest in cases of choledochal cyst (21.4 +/- 12.1 mm, compared with cases of biliary tract dilatation with biliary atresia (10 +/- 2.4 mm), secondary biliary duct dilatation (8.5 +/- 1.5 mm), and normal variants (4.4 +/- 1.2 mm) (P < 0.001). Of the 43 infants with biliary tree dilatation, 24 (56%) had choledochal cysts and 19 (44%) had biliary tract dilatation associated with biliary atresia. Excluding cases associated with biliary atresia, the accuracy of diagnosing choledochal cysts in extrahepatic biliary tract dilatation was 71% and 97% using cutoffs of 7 mm and 10 mm as the minimum diameter, respectively. PMID- 10709834 TI - Comparison of standard and second harmonic B-mode sonography in the detection of segmental renal infarction with sonographic contrast in a rabbit model. AB - This study compares contrast-enhanced fundamental and second harmonic B-mode sonography using a rabbit renal infarct model. Segmental renal infarctions were produced in 13 rabbits by embolizing a 0.7 mm bead into the renal artery 1 day prior to imaging. An ultrasonographic unit equipped with an L10-5 transducer and second harmonic imaging capability was used. Real-time recordings were made during the injection of 0.5 ml of an experimental formulation of a perfluorohexane vapor-stabilized microbubble (AF0145) given into the ear vein, and the imaging technique alternated between standard and harmonic imaging every 20 s. Each rabbit received two injections 1 h apart. To control for the effect of peak bolus enhancement, the initial imaging technique used for the first injection was randomized, and the other technique was used initially for the second injection. The videointensity difference between the infarcted and the normal cortex was then calculated and evaluated as a function of time. The infarcted segment could not be seen before administration of contrast agent with either technique. Although the infarction could be seen after injection of contrast agent with either technique, image contrast and contrast duration were nearly 75% greater for the harmonic technique than for the standard technique. AF0145 allows the visualization of segmental renal infarction on standard B-mode imaging. The second harmonic B-mode technique significantly increases image contrast and contrast duration. PMID- 10709835 TI - Enlarged lymph nodes of the neck: evaluation with parallel extended field-of-view sonographic sequences. AB - A standardized extended field-of-view sonographic examination technique of the neck is evaluated. In a prospective study we screened 50 patients suspected of having carcinoma or lymphoma for enlarged cervical lymph nodes. After conventional CT of the neck, extended field-of-view sonography was performed using defined axial parallel scanning sequences. The results were interpreted separately by two radiologists. Of 245 lymph nodes (diameter 1 cm or greater) diagnosed with conventional CT, 218 were correctly identified by extended field of-view ultrasonography. With respect to the entire neck, the sensitivity of extended field-of-view sonography was 92%, and the correlation coefficient between the methods was r = 0.98 (P < 0.001). Fifteen of 17 false-negative lymph nodes were located in the mandibular angle region. False-positive results (N = 10) were caused by misinterpretation of primary tumors, blood vessels, lobulated salivary glands, and double imaged lymph nodes. Our results indicate that extended field-of-view sonography in parallel scanning sequences represents a reliable method for the detection of cervical lymphadenopathy. PMID- 10709836 TI - Foot length in fetuses with abnormal growth. AB - Sonographic fetal foot length is highly predictive of gestational age. In order to assess the reliability of this parameter in predicting gestational age in cases of abnormal fetal growth, we examined fetal foot length in small- and large for-gestational-age fetuses. A nomogram of foot length versus gestational age between 15 and 37 weeks was constructed using cross-sectional data obtained from 5372 singleton fetuses. Fetal foot lengths for small-for-gestational-age fetuses (estimated fetal weight below the 10th percentile) and large-for-gestational-age fetuses (above the >90th percentile) fetuses were plotted against the foot length nomogram in order to determine the number of small-for-gestational-age fetuses and large-for-gestational-age fetuses with foot lengths below the 10th and above the 90th percentiles, respectively. Of the 586 small-for-gestational-age fetuses, 355 (60.6%) had foot lengths below the 10th percentile on the nomogram. When foot lengths from large-for-gestational-age fetuses were plotted on the foot length nomogram, 29.4% (219 of 744) had measurements above the 90th percentile. Fetal foot length can be influenced by growth restriction as well as states of accelerated fetal growth. Our findings imply that there are limitations to the use of fetal foot length for gestational age assessment, particularly in fetuses with growth abnormalities. PMID- 10709837 TI - Sonographic appearance of the ventriculus terminalis cyst in the neonatal spinal cord. AB - The ventriculus terminalis or "fifth ventricle" is an ependyma-lined residual lumen of the caudal portion of the spinal cord (the conus medullaris). We present the cases of three neonates with asymptomatic cystic dilatation of the ventriculus terminalis as seen on spinal sonography. Over a 4 year period (1996 1999), we prospectively found three cases in which spinal sonograms demonstrated cystic dilatation of the ventriculus terminalis of the conus medullaris in normal term neonates. Sonograms of the lumbosacral spine of two of the infants demonstrated cystic dilatation of the ventriculus terminalis of the conus medullaris. The third infant had cystic dilatation at the distal tip of the conus medullaris at the origin of the filum terminale. No other abnormalities were noted. The three infants have remained asymptomatic during clinical follow-up periods of up to 3 years. Cystic dilatation of the ventriculus terminalis is an unusual but normal anatomic variant of the conus medullaris that can be visualized on spinal sonograms in neonates. PMID- 10709838 TI - Transperineal ultrasonography in imperforate anus: identification of the internal fistula. AB - The purpose of this study was to assess the usefulness of transperineal ultrasonography in identifying the internal fistula in cases of imperforate anus. Transperineal ultrasonography was performed in 19 infants (13 neonates and 6 older infants; 13 were male and 6 were female) with imperforate anus to identify the internal fistula. Sagittal plane images were obtained through the anal dimple, and the internal connection of the rectal fistula was traced. The ultrasonographically traced internal fistula was compared with that observed on distal loopography after colostomy or with surgical findings. The internal fistula was identified as a hypoechoic linear tract, containing linear echogenicity in some cases. Of 19 patients, internal fistulas were correctly identified in 16 patients; these were rectourethral (n = 12), rectovaginal (n = 1), rectovestibular (n = 1), rectovesical (n = 1), and rectocloacal (n = 1). In three patients, internal fistulas were incorrectly defined; these cases consisted of rectovestibular (n = 2) and rectovaginal (n = 1) fistulas. Internal fistulas were correctly identified in all of the 13 male patients and in 3 of 6 female patients. Transperineal ultrasonography is an excellent diagnostic modality to define the type of the internal fistula in imperforate anus. PMID- 10709839 TI - Narrowing of the upper abdominal inferior vena cava in patients with elevated intraabdominal pressure: sonographic observations. AB - Focal narrowing of the upper intrahepatic inferior vena cava on computed tomography performed in patients with elevated intraabdominal pressure has been reported recently. The purpose of this study was to assess this phenomenon further with gray scale and duplex Doppler sonography. Seven patients with elevated intraabdominal pressure due to massive ascites in whom computed tomography showed narrowing of the intrahepatic inferior vena cava were studied. Sonography confirmed focal narrowing of the intrahepatic inferior vena cava that persisted throughout the respiratory and cardiac cycles. The narrowed inferior vena cava segment had a characteristic slitlike appearance on transverse images, slightly oblique to the sagittal plane. A focal flow jet typical of stenosis was observed in the three patients in whom Doppler sonography technically was feasible. In three of four patients who were able to sit or to turn into the left lateral decubitus position, the narrowing immediately resolved after the positional change but promptly recurred upon return to the supine position. In patients with elevated intraabdominal pressure, focal slitlike narrowing of the intrahepatic inferior vena cava is an anticipated finding that should not be misinterpreted as evidence of stenosis of the inferior vena cava or of a juxtacaval liver lesion. PMID- 10709840 TI - Umbilical cord hematoma associated with an umbilical cord cyst and fetal death at 28 weeks of gestation. PMID- 10709841 TI - Ultrasonographic findings of an intratesticular adenomatoid tumor. PMID- 10709843 TI - The role of inflammatory mediators in the mechanism of the host immune response induced by ischemia-reperfusion injury. AB - Our previous study suggested that inflammatory mediators released due to IRI lead to host's immune response by upregulating MHC II in the host's peripheral T lymphocytes. This study hypothesized the role of platelet-activating factor (PAF) in the mechanism of induced MHC II upregulation due to IRI on peripheral T lymphocytes. The objectives of this study were to investigate the role of PAF in the induction of host immune reactivity and the protective effect of PAF antagonist TCV-309 in combination with prostaglandin E1 (PGE1) against the host's immune response caused by IRI. Thirty female domestic swine were divided into three groups. Group A (6 donors, 6 recipients) had no pharmacological intervention. Group B (6 donors, 6 recipients) was the experimental group treated with TCV-309 + PGE1. Group C underwent sham operation. The ex vivo preservation time for groups A and B was 4 hr at 4 degrees C. To detect the changes in MHC II expression on T cells due to IRI, blood samples were collected before reperfusion (baseline level), 1, 2, and 3 days post-reperfusion. Two-colour flow cytometry analysis (FACS) was used to study MHC II-DR-beta expression in peripheral T lymphocytes. Swine anti-MHC II and anti-CD3 antibodies were used for this purpose. The FACS analyses demonstrated that in group A, there was a significant increase (p < 0.05) in MHC II intensity on peripheral T lymphocytes on day 2 post reperfusion. By the third day post-reperfusion, MHC intensity had a tendency to decrease but did not reached the baseline level. In group B and C, however, there was no significant change in the level of MHC II in T lymphocytes at any of the post-reperfusion times. In group A, the number of CD3+MHC+ T lymphocytes significantly decreased (p < 0.05) by one day post-reperfusion and remained at this level until the third day post-reperfusion. In groups B and C, no significant change in the number of CD3+MHC+ T cells was observed. The results of this study suggested that the release of inflammatory mediators (e.g. PAF) due to IRI played a role in the mechanism of IRI-induced host's immune response. The results also suggested that the combination of TCV-309 + PGE1 could reduce this immune response. PMID- 10709842 TI - Protective immunity induced by a Trypanosoma cruzi soluble extract antigen in experimental Chagas' disease. Role of interferon gamma. AB - CBA/J mice can be protected against lethal infection with Trypanosoma cruzi by treatment using T. cruzi soluble extract antigen (TCSE). In vivo administration of TCSE (400 microg/mouse) into naive mice increased the cellular proliferative response to Con A and elevated the levels of IFN-gamma. The production of IFN gamma was extremely important in controlling the replication of the parasite since the protective activity of TCSE was completely abrogated by in vivo treatment with an anti IFN-gamma neutralizing antibody. These results suggest that depending on the level, cytokine production results in the control of replication of the parasite in experimental Chagas' disease. PMID- 10709844 TI - Soluble T cell receptors modulate cytokine production and oxygen metabolism by peritoneal macrophages. AB - Preincubation of peritoneal macrophages and their subsequent culture with recombinant soluble T cell receptor (sTCR) results in significant increase of: TNF-alpha, IL-1beta, IL-6, IL-10, IL-12 production and nitric oxide (NO) synthesis and this phenomenon was dose dependent. Moreover, treatment of macrophages with sTCR showed two to three fold increase of luminol dependent chemiluminescence (LCL) when compared to untreated macrophages (Mf). In contrast, in our study we did not find any influence of sTCR on co-stimulatory (B7.1 and B7.2), adhesion molecule (ICAM-1) or FcRII/III expression by macrophages. However, macrophages treated with control supernatants received after phosphatidylinositol-specific phospholipase C (PI-PLC) treatment of BW1100 cells or thymocytes termed s-BW or s-Th did not influence their biological activity. PMID- 10709845 TI - Epitope-vaccine induces high levels of ELDKWA-epitope-specific neutralizing antibody. AB - Based on the fact that mAb 2F5 recognizing ELDKWA-epitope on the C-domain of HIV 1 gp41 has significant neutralization potency against 90% of the investigated viruses of African, Asian, American and European strains, we attempted to characterise immunogenicity of the ELDKWA-epitope on an epitope-vaccine, and to produce ELDKWA-epitope-specific monoclonal antibodies (mAb) induced by the epitope-vaccine. The C-domain peptide (P2) and the ELDKWA-tetramer peptide [C (ELDKWAG)4] were conjugated with BSA or P24-EC (GPKEPFRDYVDRFYK, a peptide of HIV 1 gag-protein P24, proved to be a good carrier peptide to induce an immune response to the hapten on the conjugates[18])by different methods. After the vaccination course, two P2-BSA peptide-vaccines both induced a strong antibody response against the P2-peptide by about 1:12800-25600 dilution, and a weak antibody response against the ELDKWA-epitope (1:1600-3200). The P2-P24EC and P2 (conjugated with itself) peptide-vaccines could also induce a weak antibody response against the ELDKWA-epitope (1:1600-3200), while an rgp160 subunit vaccine induced a very weak antibody response (1:400). Interestingly, the ELDKWA tetramer epitope-vaccine [C-(ELDKWAG)4-BSA] could induce a strong antibody response against the ELDKWA-epitope (1:12800-25600), i.e. It increased the level of ELDKWA-antibody eight-fold, clearly better than the P2 peptide-vaccine, and much better than the rgp160 subunit vaccine, which indicates that the immunogenicities of the ELDKWA-epitope on the ELDKWA-tetramer peptide, the C domain peptide and rgp160 are very different. These results suggest that the ELDKWA-epitope-vaccine may be a new strategy for inducing high levels of epitope specific neutralizing antibodies against HIV-1. Using hybridoma-technique, a mouse monoclonal antibody recognizing the ELDKWA-epitope on ELDKWA-peptide and C domain peptide was produced by immunization with the C-(ELDKWAG)4-BSA epitope vaccine, which indicates a new way to produce an epitope-specific mAb, namely immunization with epitope-vaccine instead of a natural or recombinant protein immunogen. PMID- 10709846 TI - Murine bone marrow-derived mast cells exhibit evidence of both apoptosis and oncosis after IL-3 deprivation. AB - IL-3 deprivation has been reported to induce apoptosis of bone marrow-derived mast cells. In order to evaluate this type of cell death further, we employed trypan blue and propidium iodide stainings, photometric enzyme immunoassay, fluorescence measurement of caspase-3, DNA electrophoresis, flow cytometry and transmission electron microscopy. In this experiment, although several evidences supporting apoptosis were demonstrated some findings were not consistent with typical apoptosis. On the other hand, electron microscopical observation demonstrated that most cells from all the time phases after IL-3 deprivation showed the morphology of typical oncosis, i.e. cell swelling, disintegration of ultrastructure and subsequent karyolysis. Only a small number of cells from the later time phases showed apoptotic morphology. We here suggest that BMMCs undergo both apoptosis and oncosis after IL-3 deprivation and that the dominant type of prelethal change is oncosis in all time phases, although apoptosis also plays a partial role in the late time phases. PMID- 10709847 TI - Kinetics of peptide uptake and tissue distribution following a single intranasal dose of peptide. AB - We have previously used intranasal (i.n.) peptide application to induce mucosal tolerance in experimental autoimmune encephalomyelitis (EAE). This strategy, however, appeared to give rise to similar phenomena of tolerance observed as a result of systemic administration of soluble antigenic peptide. We were interested, therefore, in the uptake and tissue distribution of peptide following i.n. treatment. In the H-2u mouse model of EAE, the highly tolerogenic peptide analogue Ac1-9[4Y] of myelin basic protein (MBP) displays high affinity binding to Au MHC class II. For the purpose of the present study this peptide was synthesised to contain a tritiated acetyl group and a protocol was developed to recover radioactivity in solubilised tissues taken at various times after [3H]Ac1 9[4Y] i.n.. Radiolabel loads of the lung and gastro-intestinal tract were initially high but declined rapidly. Radiolabel uptake by blood and lymphoid tissues followed similar kinetics with peak levels around 2.5-4 hours after i.n. administration. Concentrations were high in the draining cervical lymph nodes (CLN) but also reached significant levels in the spleen and 'nondraining' inguinal lymph nodes. The presence of intact antigenic peptide was demonstrated in spleens and CLN from Ac1-9[4Y] i.n. treated mice. Cell suspensions prepared from these tissues at selected time points after peptide i.n. were able to stimulate peptide-specific T cell lines up to at least one day after peptide i.n., suggesting long lasting formation of stable Au-Ac1-9[4Y] complexes in vivo. PMID- 10709848 TI - Induction of PYK-2 phosphorylation during LFA-1/ICAM-1-dependent homotypic adhesion of fresh human B-cells. AB - Stimulation with the combination of PDB plus ionomycin induced LFA-1/ICAM-1 dependent homotypic adhesion of tonsil B cells. Adhesion of tonsil B cells in our system induced tyrosine phosphorylation of Pyk-2. Disruption of homotypic adhesion and concomitant inhibition of induction of protein tyrosine phosphorylation was achieved by physical separation of the cells and by treatment with methyl-2.5-dihydroxycinnamate (MDHC), an inhibitor of protein tyrosine phosphorylation. These results suggest that protein tyrosine phosphorylation that is associated with homotypic adhesion is mediated by LFA-1/ICAM-1 interactions. PMID- 10709849 TI - Cellular signaling with nitric oxide and cyclic guanosine monophosphate. AB - The understanding of the formation and biological actions of nitric oxide (NO) has grown extensively during the past two decades. With the discoveries of the biological effects of NO and nitrovasodilators on cyclic guanosine monophosphate, with the elucidation of the biochemical mechanisms of NO synthesis, and with the growing knowledge of regulation of NO synthases, the complexities of this signal transduction cascade and its participation in numerous cell signaling processes continues. NO can be recognized as an intracellular second messenger, a local substance for regulation of neighboring cells, a neurotransmitter, and probably a hormone acting at distant sites. PMID- 10709850 TI - Angiotensin converting enzyme: history and relevance. AB - The renin angiotensin system (RAS) is now recognized as the body's most powerful hormone system for controlling renal hemodynamics and sodium excretion and, therefore, body fluid volumes and arterial pressure. The discovery of angiotensin converting enzyme inhibitors (ACEi) was a keystone for the understanding of the significance of the RAS since ACEi proved to be effective in controlling hypertension and heart failure and in preventing the development of the vascular injury of chronic diseases like scleroderma and diabetes mellitus. The success of ACEi stimulated the research into inhibitors of other actors of the RAS like renin or angiotensin receptor antagonists. It is not often realized that the discovery of ACEi owes a great deal to basic research in which the venom of a Brazilian viper, Bothrops Jararaca, was instrumental for the discovery of bradykinin by Rocha e Silva and the bradykinin potentiating factor. This article reviews the contribution of the converting enzyme inhibitors for the demonstration of the relevance of the RAS to several human pathologies. PMID- 10709851 TI - Hemodynamic changes in pregnancy. AB - The basic mechanisms that underlie alterations in the physiology of pregnancy are virtually unknown. Basal oxygen consumption increases by some 50 mL/min in pregnant women at term. Blood volume increases gradually over gestation as does red cell mass. Cardiac output increases by some 50% by mid-third trimester. Stroke volume and heart rate increase over the course of pregnancy with heart rate increasing gradually until term. The heart of the pregnant woman remodels dramatically in the first few weeks of pregnancy; end diastolic volume increases. Stroke volume is augmented by the increase in end diastolic volume and maintenance of ejection fraction through a possible increase in contractile force. Systolic and diastolic blood pressures drop during normal pregnancy. There is evidence of blood vessel remodeling in all vessels. Venous compliance and venous blood volume are increased. Renal plasma flow increases by some 70% in pregnancy with glomerular filtration rate increasing by 50% by unknown mechanisms. The complex hormonal environment is changing throughout pregnancy. In summary, under the influence of circulating chemical mediators blood flow is redistributed to the uterus, breast, and kidney. PMID- 10709852 TI - EDHF: a cytochrome P450 metabolite in coronary arteries. AB - Endothelium-dependent relaxation cannot be fully attributed to the release of nitric oxide or prostacyclin (PGI2). In resistance-sized vessels and coronary arteries a high proportion of endothelium-dependent relaxation, in response to agonist-induced or mechanical stimulation of endothelial cells, can be attributed to the release of 1 or more endothelium-derived hyperpolarizing factor (EDHF). In coronary arteries EDHF has been pharmacologically characterized as a cytochrome P450 (CYP)-derived metabolite of arachidonic acid. We show here that a CYP 2C arachidonic acid epoxygenase, homologous to CYP 2C8/9, is expressed in cultured human endothelial cells and native porcine coronary artery endothelial cells. Down-regulation of CYP 2C protein by transfection of porcine coronary arteries with anti-sense oligonucleotides decreased EDHF-mediated vascular responses while EDHF-mediated hyperpolarisation and relaxation were potentiated by the CYP inducing compound beta-naphthoflavone. Thus, CYP 2C appears to play a crucial role in the generation of EDHF-mediated responses in porcine coronary arteries. PMID- 10709853 TI - Effects of oxidative and nitrosative stress in cytotoxicity. AB - Nitric oxide is a key bioregulatory agent in a wide variety of biological processes, yet it also can have cytotoxic properties. This dichotomy raises the question of how this potentially toxic species can be involved in so many fundamental physiological processes. This articles discusses how the chemistry of nitric oxide might pertain to its observed biology as it relates to oxidative and nitrosative stress in different mechanisms of cytotoxicity. PMID- 10709854 TI - Preeclampsia: what we know and what we do not know. AB - Preeclampsia remains a major health problem for mothers and infants. Studying the entire pathophysiology of preeclampsia rather than "pregnancy-induced hypertension" has greatly expanded our knowledge of the disorder. Current thinking approaches preeclampsia as a 2 stage disorder: reduced placental perfusion usually secondary to abnormal implantation and a consequent maternal disorder characterized by endothelial dysfunction and subsequent pathophysiological changes. We know much about the 2 stages and less about their linkage. It is evident that reduced placental perfusion is not sufficient to account for the pathophysiology. Reduced perfusion and abnormal implantation occur in other conditions (intrauterine growth restriction and preterm labor) without the maternal syndrome. This leads to the hypothesis that reduced placental perfusion must interact with maternal constitutional factors to generate the systemic pathophysiology of preeclampsia. The similarities of these risk factors and metabolic alterations between preeclampsia and atherosclerosis suggest a common pathophysiology. Oxidative stress is postulated as the genesis of endothelial dysfunction in atherosclerosis. The author proposes that oxidative stress secondary to reduced placental perfusion leads to endothelial dysfunction, linking the 2 stages of the syndrome. PMID- 10709855 TI - The role of lipid oxidation and oxidative lipid derivatives in the development of preeclampsia. AB - Preeclampsia develops as a consequence of an exceptionally complex interaction between a multiplicity of factors that originate in 2 genetically different individuals (the mother and the fetus). Oxidative stress/oxidative lipid derivatives may represent one group of such factors. The evidence for a role of oxidative stress/oxidative lipid derivatives in the pathogenesis of preeclampsia may be summarized as follows: In women with established preeclampsia there is good evidence of increased oxidative stress/oxidative lipid derivatives in the decidual-placental tissues. Likewise, in the systemic circulation of women with established preeclampsia most of the studies although not all, indicate elevated levels of oxidative lipid derivatives and reduced anti-oxidative capacity. Because almost all studies on the role of oxidative stress in the pathogenesis of preeclampsia have been among women with established preeclampsia, it remains uncertain whether enhanced oxidative stress is present before clinical signs of preeclampsia develops. One controlled randomized study in which antioxidants were used to treat severe preeclampsia showed nonsignificant differences in clinical outcomes in favor of antioxidants. Controlled studies on the effectiveness of antioxidants in preventing preeclampsia are lacking. Such studies are of major interest in order to evaluate more definitely the role of oxidative stress/oxidative lipid derivatives in the pathogenesis of preeclampsia. PMID- 10709856 TI - Calcium, nitric oxide, and preeclampsia. AB - A relationship between calcium dietary intake and incidence of preeclampsia was proposed. In the Andean Ecuadorian population, the average calcium intake, evaluated by a 24 hours dietary recall range between 52.3% of the US RDA to 77%. The calcium intake in women with preeclampsia was significantly lower in relation with normal pregnant women. Three prospective, randomized, double-blind, placebo controlled clinical trials to investigate the effect of calcium supplementation (2 g/day of elemental calcium) in the incidence of pregnancy-induced hypertension and preeclampsia were conduced between 1984 and 1995. All the subjects included were nulliparous, younger that 25 years old, first prenatal visit before 24 weeks' gestation, residency in Quito, and normotensives. These clinical trials showed a risk reduction in pregnancy induced hypertension and preeclampsia in the calcium group. Calcium supplementation was associated with an increase in the serum ionized calcium concentrations. Moreover, women with preeclampsia showed a significant decrease in the levels of the serum ionized calcium. Ionic calcium is crucial for the synthesis of vasoactive substances in the endothelium as prostacyclin and nitric oxide. Recent results suggest that an alteration in the action of NO may be related to a high inactivation by free radical superoxide, secondary to an inflammatory process. PMID- 10709857 TI - Nitric oxide and peroxynitrite in the perinatal period. AB - Many of the actions of nitric oxide are not due to nitric oxide itself, but rather by the secondary formation of oxidants like peroxynitrite. Peroxynitrite leaves a footprint in the nitration of tyrosine, which helps track the formation of reactive nitric oxide-derived species in diseases and even normal development. PMID- 10709858 TI - The preferred source of arginine for high-output nitric oxide synthesis in blood vessels. AB - L-arginine is the substrate for nitric oxide (NO) production by each of the 3 NO synthase (NOS) isoforms encoded by the mammalian genome. Despite the pivotal roles of NO in mammalian physiology and pathophysiology, the source of arginine for NO synthesis is not clearly defined. In this context, it is notable that cell types that do not have a complete urea cycle often possess the urea cycle enzymes argininosuccinate synthase and argininosuccinate lyase; together, these enzymes confer the ability to regenerate arginine from the NOS product, L-citrulline. Herein, the authors summarize evidence to support the view that argininosuccinate synthase and argininosuccinate lyase function in an arginine-citrulline cycle, providing a ready source of arginine for high-output NO synthesis. The arginine citrulline cycle is induced in vascular cells by the same cytokines that trigger iNOS expression and provides the preferred source of substrate for NO production. Evidence suggests that argininosuccinate synthase activity is rate-limiting to high-output NO synthesis and, hence, represents a novel target for the treatment of pathophysiological conditions arising from NO overproduction. PMID- 10709860 TI - Nitric oxide synthase gene therapy in vascular pathology. AB - Nitric oxide (NO) is intimately involved in vascular homeostasis through its antiplatelet, antiproliferative, and vasodilating actions. Because of these beneficial properties, methods of harnessing NO for the prevention of vascular injury responses, such as intimal hyperplasia, are being explored. One such method involves gene transfer of an NO synthase (NOS) to sites of vascular injury to provide for local NO synthesis. Gene delivery of the inducible NOS (iNOS) cDNA to sites of vascular injury in animal models dramatically reduced smooth muscle proliferation and intimal hyperplasia. The cellular mechanisms by which NO inhibits smooth muscle cell proliferation appear to be independent of cyclic guanosine monophosphate production but are linked to the upregulation of the cell cycle inhibitor p21. p21 upregulation occurs independent of p53 expression. Instead, p42/44 mitogen activated protein kinase activation by NO results in reduced cellular proliferation and increased p21 expression, suggesting NO inhibits intimal hyperplasia through cell cycle arrest as mediated by p21 and the signaling pathway involved in p21 upregulation may be regulated by p42/44 mitogen activated protein kinase. PMID- 10709859 TI - The role of nitric oxide in apoptosis. AB - Nitric oxide (NO) has been shown to play important roles in modulating cell viability. Both proapoptotic and antiapoptotic functions of NO have been reported. This article discusses our current understanding on the mechanisms responsible for the dual actions of NO in regulating apoptosis and propose that one common mechanism for NO-mediated inhibition of apoptosis is the inhibition of caspases via nitrosylation. PMID- 10709861 TI - Inhaled nitric oxide. AB - Nitric oxide (NO) is a free-radical gas that is an important signaling molecule in pulmonary vessels. Endogenous NO produced in endothelial cells from oxygen and L-arginine diffuses into smooth muscle cells in the vascular wall and causes vasodilatation. NO that diffuses into the blood vessel lumen is avidly bound by hemoglobin and does not cause important systemic vasodilatation. Inhaling low levels of NO rapidly and safely decreases pulmonary artery hypertension in many patients without causing systemic hypotension. In hypoxemic newborns with pulmonary hypertension, clinical studies indicate that inhaled NO increases systemic oxygen levels and decreases the requirement for extracorporeal membrane oxygenation. NO also has been observed to regulate cell proliferation. Recent studies suggest that inhaled NO selectively modulates the pulmonary artery proliferative response that is associated with lung injury. These later studies may indicate that inhaled NO can be applied to attenuate or prevent pulmonary artery disease in patients with injured lungs. PMID- 10709862 TI - Nitric oxide therapy for the newborn infant. AB - Inhaled nitric oxide (INO) is a novel selective pulmonary vasodilator without significant effects on the systemic circulation. Initial case studies of near term newborn infants with hypoxic respiratory failure and persistent pulmonary hypertension of the newborn showed that INO was associated with improvements in oxygenation. There have now been at least 11 prospective randomized controlled trials evaluating the use of INO in the near-term neonate with hypoxic respiratory failure, 10 of which have been published. A meta-analysis of these trials provides evidence that INO improved the PaO2 in the INO treated infants by 46.4 torr (weighted mean difference) compared with controls (95% CI, 34.2, 58.5) and significantly decreased the oxygenation index by 10.7 compared with controls (95% CI, -14.1, -7.4). The incidence of death or need for extracorporeal membrane oxygenation (ECMO) was significantly reduced by treatment with INO, relative risk (RR) 0.72 compared to control (95% CI, 0.6, 0.87) with the majority of the improvement seen in the reduction in the need for ECMO. Infants with congenital diaphragmatic hernia do not appear to benefit from early INO therapy. The only prospective trials evaluating INO in premature infants to date have not found that this therapy is associated with significant clinical benefit. The long-term evaluations of near-term and full-term infants who have received INO suggest that this therapy does not increase the incidence of adverse neurodevelopmental sequelae in these high-risk infants. INO is an effective therapy for the hypoxic term neonate and will reduce the occurrence of death or the need for ECMO in this population. Further research is required to evaluate the benefit of this therapy in the hypoxic preterm infant. PMID- 10709863 TI - An update on estrogen receptors. AB - The discovery of a second estrogen receptor (ER), ERbeta, has led to a complete change in our views on estrogen action. The previous dogmatic view that ERalpha represented the only estrogen receptor led to a static and simplistic concept of mechanisms of estrogen action with conceptual limitations in the development of novel estrogenic and antiestrogenic drugs. It is now realized that estrogen signaling represents a complex and multi facetted signal transduction pathway with, at least in many cases, quite different roles of ERalpha and ERbeta. For instance, the two receptors appear to behave quite differently on AP1, antioxidant and Sp1-response elements where ERbeta mediates positive regulation by antiestrogens whereas ERalpha is silent under these conditions. ERalpha and ERbeta also appear to be differentially distributed in the body and within tissues. They are regulated differently and seem to have distinct biological roles, at least in certain contexts. Data are currently rapidly generated with respect to these issues from knockout animals with either of the two receptors deleted. Also double knockouts have been generated and apparently survive. ERbeta may well have significant roles in the etiology of the following diseases and symptoms: prostate cancer, osteoporosis, depression, as well as urinary incontinence in postmenopausal women. Attempts are ongoing in several labs to develop specific ligands to the two receptors. Such ligands may well turn out to be extremely important in treating the mentioned diseases and symptoms as well as possibly others. PMID- 10709864 TI - Novel role of estrogen receptors in vascular endothelium. AB - Estrogen has a variety of effects on the vascular wall including rapid vasodilation due to the stimulation of endothelial nitric oxide synthase (eNOS). Studies in cultured endothelium indicate that the hormone cause acute, direct activation of eNOS that is unaffected by actinomycin D but fully inhibited by estrogen receptor (ER) antagonism. Overexpression of ERalpha leads to marked enhancement of the acute response to estrogen, and this process is blocked by ER antagonism and requires the ERalpha hormone binding domain. The acute response of eNOS to estrogen can also be reconstituted in COS-7 cells cotransfected with ERalpha and eNOS, but not by transfection with eNOS alone. The inhibition of calcium influx, or tyrosine kinases or mitogen-activated protein (MAP) kinase prevents eNOS stimulation by estrogen, and estrogen causes rapid ER-dependent activation of MAP kinase. Thus, the acute effect of estrogen on eNOS is mediated by ERalpha functioning in a novel, nongenomic manner to activate the enzyme via calcium-dependent, MAP kinase-dependent mechanisms. PMID- 10709865 TI - VPF/VEGF and the angiogenic response. AB - Vasculogenesis, the generation of new blood vessels de novo, and angiogenesis, the formation of new blood vessels from preexisting vessels, are mediated by a number of cytokines and growth factors among which vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) is one of the most important. VPF/VEGF is secreted by many tumor cells, at sites of wound healing and chronic inflammation, and in physiological angiogenesis as in corpus luteum formation. VPF/VEGF is a multifunctional cytokine that interacts with two high affinity tyrosine kinase receptors that are selectively expressed on vascular endothelium. This interaction triggers an angiogenic cascade whose steps, among others, include increased microvascular permeability, leading to deposition of a pro-angiogenic extracellular fibrin matrix, and the formation of mother/daughter vessels. PMID- 10709866 TI - Importance of angiogenesis in reproductive physiology. AB - The cyclic angiogenesis that occurs uniquely within the female reproductive tract is critical for normal reproduction. Two families of endothelial cell-specific growth factors and their receptors have been identified in the ovary, uterus, and placenta: vascular endothelial growth factor/vascular permeability factor and the angiopoietins. These appear to have complementary actions on the vasculature and to be involved during intrauterine development as well as in the adult. Within the ovary, a complex cascade of events required for angiogenesis may play a role in follicular maturation and selection as well as in normal corpus luteum function. Aberrant expression of angiogenic factors plays a role in a wide variety of abnormalities in the ovary. In the uterus, angiogenesis is required for endometrial growth and remodeling. Peptide growth factors such as vascular endothelial growth factor may serve as local mediators of the effects of reproductive hormones on the endometrial vasculature. Disease states such as dysfunctional uterine bleeding, endometriosis, and endometrial hyperplasia or cancer may be associated with aberrant uterine angiogenesis. PMID- 10709867 TI - Angiogenic growth factor expression in placenta. AB - New blood vessel growth is generally a rare event in the healthy adult. However, a notable exception to this is the female reproductive tract where cyclic angiogenesis occurs. Striking new vessel growth and remodeling also occurs during placentation; thus angiogenesis is essential for reproductive success. Vascular endothelial growth factor is a potent stimulator of this process and its production and action is tightly regulated. Indeed the placenta is a rich source of a soluble variant of the flt-1 receptor which seems to protect the placenta from the effects of excess vascular endothelial growth factor. The balance between new vessel growth (in the placental villi for example) and endothelial cell loss in the spiral arteries within the decidua is a delicate one. This is influenced by the local production of promotors and inhibitors of endothelial cell activation. Perturbation of this may lead to maternal pathology during pregnancy. PMID- 10709868 TI - Lessons learned from nitric oxide synthase knockout animals. AB - Nitric oxide (NO) is generated by 3 major isoforms of nitric oxide synthase (NOS) with complex and overlapping patterns of expression. This article presents several examples of how gene targeted mice lacking endothelial and neuronal isoforms have showed various roles of NO. Neuronal NOS knockout mice are resistant to global and focal cerebral ischemia, confirming a role for neuronal NO in cellular toxicity after stroke. Endothelial NOS knockout mice have increased susceptibility to stroke consistent with a vascular protective effect of NO. They are hypertensive and lack endothelium dependent relaxing factor activity. Analysis of cardiac function shows roles for NO in suppression of inotropic responses to beta-adrenergic agonists and in mediating basal diastolic relaxation. Endothelial NOS knockout mice respond to vascular injury with increased neointimal proliferation, consistent with a physiological role for NO to suppress smooth muscle cell proliferation. PMID- 10709869 TI - Temporal profile of release of neurobiochemical markers of brain damage after traumatic brain injury is associated with intracranial pathology as demonstrated in cranial computerized tomography. AB - This study aimed at the investigation of release patterns of neuron specific enolase (NSE) and protein S-100B after traumatic brain injury (TBI) and their association with intracranial pathologic changes as demonstrated in computerized tomography (CT). We analyzed NSE and S-100B concentrations in serial venous blood samples taken one to three days after TBI in 66 patients by the use of immunoluminometric assays. These markers are considered to be specific neurobiochemical indicators of damage to glial (S-100B) or neuronal (NSE) brain tissue. Standardized neurological examination and plani- and volumetric evaluation of computerized tomography scans were performed in all patients. Patients with medium severe to severe TBI [Glasgow Coma Scale (GCS) score at the site of accident < or =12] exhibited significantly higher NSE and S-100B concentrations and a significantly longer release compared to patients with minor head injury (GCS: 13-15). Both, patients with and without visible intracerebral pathology in CT scans exhibited elevated concentrations of NSE and S-100B after TBI and a significant decrease in the follow-up blood samples. Release patterns of S-100B and NSE differed in patients with primary cortical contusions, diffuse axonal injury (DAI), and signs of cerebral edema (ICP) without focal mass lesions. All serum concentrations of NSE and S-100B were significantly correlated with the volume of contusions. The data of the present study indicate that the early release patterns of NSE and S-100 may mirror different pathophysiological consequences of traumatic brain injury. PMID- 10709870 TI - Altered expression of amyloid precursors proteins after traumatic brain injury in rats: in situ hybridization and immunohistochemical study. AB - The expression of alternatively spliced mRNAs for amyloid precursor protein (APP) isoforms and their translation products were examined in the rat cerebral cortex 1, 3, 6, and 12 h and 1, 3, and 7 days (n = 4-5 in each group) after fluid percussion brain injury. In situ hybridization studies demonstrated that the expression of APP695 mRNA decreased in and around the damaged area of the cerebral cortex exposed to fluid-percussion injury 1 h after the insult. On the other hand, APP751/770 mRNAs were increased in the regions surrounding the damaged cortical areas 1 day after the injury. An increase of immunoreactive APP was detected in the regions around the damaged cortical areas 3 h after traumatic injury and maintained for the following 3 days. The APP immunoreactivity in the damaged cortices declined to the level of sham-operated animals by post experimental day 7. Using an anti-amyloid beta (Abeta) protein (17-24) antibody, no deposits of immunoreactive Abeta (17-24) were observed in any of the samples examined in these experiments. These results suggest that the induction of Kunitz type protease inhibitor (KPI) domain-containing APP mRNAs and the increased accumulation of APP are involved in the physiological and neuropathological responses of brains under various neurodegenerative conditions, including head trauma. PMID- 10709871 TI - Lactate/glucose dynamics after rat fluid percussion brain injury. AB - Traumatic brain injury (TBI) places enormous early energy demand on brain tissue to reinstate normal ionic balance. Clinical studies have demonstrated a decline in extracellular fluid (ECF) glucose and an increase in lactate after TBI. In vitro studies suggest that this increase in lactate is mediated by increased glutamate and may provide a metabolic substrate for neurons, to aid in ionic restoration. This led us to hypothesize that high ECF lactate may be beneficial in recovery following TBI, where major ionic flux has been shown to occur. In this study, we measured cerebral dialysate lactate and glucose, and arterial lactate and glucose, before and after rat lateral fluid percussion brain injury (FPI; 2.06 +/- 0.13 atm) with and without IV lactate infusion (100 mM X 0.65 mL/h X 5 h) to test the hypothesis that arterial lactate can influence ECF lactate. Dialysate lactate increased within 10 min following FPI, with higher values in the lactate infusion group. Following FPI, the dialysate lactate increase was 238% with lactate infusion versus 171% increase with saline infusion. Dialysate glucose fell immediately following FPI, with a more severe decline in the saline group. The glucose decrease was 231% greater in the IV saline group. Furthermore, in the lactate infusion group, the dialysate glucose levels recovered to baseline levels by 4 h after injury, whereas they remained depressed through out the experiment, in the saline infusion group. We conclude that arterial lactate augmentation can increase brain dialysate lactate, and result in more rapid recovery of dialysate glucose after FPI. This may indicate a beneficial role for lactate, that may be potentially useful in the clinical situation, after TBI. PMID- 10709872 TI - Real-time monitoring of glutamate following fluid percussion brain injury with hypoxia in the rat. AB - In the present study, extracellular glutamate (Glu) was monitored in real time using an enzyme electrode biosensor following traumatic brain injury (TBI) either with or without inducing hypoxia in the rat. We also measured the cortical contusion volume at 3 days after insult by staining with 2,3,5 triphenyltetrazolium chloride (TTC). Male Sprague-Dawley rats (300-400 g) were anesthetized and then subjected to lateral fluid percussion (FP) brain injury of moderate severity (3.5-4.0 atm), using the Dragonfly device model (no. HPD-1700). The experimental animals were divided into four groups. Group 1 (n = 10) was subjected to TBI only, group 2 (n = 10) to TBI followed by 20 min of moderate hypoxia (FiO2: 10%), group 3 (n = 4) to 20 min of moderate hypoxia without TBI, and group 4 (n = 4) to sham. Seventy-two hours after the insults, the animals were sacrificed, their brains were stained with TTC, and the lesion volumes were calculated. A surge in the extracellular Glu concentration occurred immediately after TBI in groups 1 and 2. There was no significant difference between the two groups. Group 2 showed a prolonged efflux of Glu during hypoxia ( < 0.05). In group 3, Glu continued to show a mild increase. The cortical contusion volume in group 2 was significantly larger than that in group 1. To evaluate the possible involvement of apoptosis in groups 1 and 2, separate rats were sacrificed under the same procedures after 1, 6, 24, and 72 h after insult (n = 2/group). Immunohistochemical analysis demonstrated an increased number of both the cysteine protease caspase-3-positive cells at 24 h and TUNEL-positive cells at 72 h in group 2. These results suggest that TBI with moderate hypoxia induced the prolonged efflux of Glu, which thus resulted in more cortical damage due to necrosis and apoptosis. PMID- 10709873 TI - Brain injury after gunshot wounding: morphometric analysis of cell destruction caused by temporary cavitation. AB - In addition to the primary destruction of brain tissue readily visible at autopsy (permanent track), gunshot wounding to the brain creates a pulsating temporary cavity due to radial expansion along the bullet's track. To determine the maximum extent of this temporary cavitation in brains of victims of gunshots from weapons with low muzzle velocity, we carried out morphometric studies on 20 cases of death from gunshot wounding to the head from bullets with a muzzle energy <500 J and a survival time of <90 min. The brains were fixed in formalin, examine macroscopically and microscopically, and subjected to morphometric analyses. Surrounding the permanent track, a narrow mantle-like zone of astrocyte destruction was found within an area of hemorrhagic extravasation. Axons near the permanent track had been broken into tiny fragments. The axonal damage lessened with increasing distance from the permanent track, although axons continued to be fragmented and to exhibit varicose changes and clumping until assuming their normal structure beyond 18 mm. Nerve cells were extremely shrunken close to the permanent track but gradually took on their normal shape with increasing distance. We also assessed the loss of glial fibrillary acid protein expression by astrocytes in the white matter, the extent of traumatic bleeding, and damage to axons and neurons as measured radially from the permanent track. Axonal and neuronal damage were found to extend about 18 mm radially from the permanent track, tapering gradually along the track from entry point to exit point. The destruction was probably produced by the temporary cavitation and accords with theoretical considerations and experimental observations. PMID- 10709874 TI - Thrombin inhibitor ameliorates secondary damage in rat brain injury: suppression of inflammatory cells and vimentin-positive astrocytes. AB - The effects of the thrombin inhibitor argatroban on the number of inflammatory cells and reactive astrocytes were investigated in a rat brain injury model. Gelatin sponge soaked with thrombin inhibitor (treatment group) or saline (control group) was placed in the brain defect to assess the infiltration of inflammatory cells by hematoxylin-eosin and immunohistochemical staining. Expression of polymorphonuclear leukocytes (PMNs) and monocyte/macrophage (Mo/Mo) cells, and vimentin (VIM)-positive astrocytes and glial fibrillary acidic protein (GFAP)-positive astrocytes were compared between groups. In the treatment group, infiltration of both PMNs and Mo/Mo cells, and the number of VIM-positive astrocytes were significantly reduced, but the number of GFAP-positive astrocytes was not different from the control group. Thrombin inhibitor suppresses the infiltration of inflammatory cells and excessive gliosis caused by VIM-positive astrocytes, but not expression of GFAP-positive astrocytes, suggesting minimization of secondary brain damage and promotion of the conditions required for neural regeneration. PMID- 10709875 TI - Implications of p53 protein expression in experimental spinal cord injury. AB - In order to clarify the role of p53, known as a tumor suppressor protein and also as a key molecule of apoptotic cell death, we have studied p53 expression in relation to localization, time course, cell type, and TUNEL reaction in a rat model of transectional spinal cord injury. Other apoptosis related molecules, p21, Bcl-2 and Bax, that are in the cascade of p53 pathway, were also examined. p53 was expressed in cells residing in the vicinity of transection as early as 30 min. For the next 2 days, the positive cells spread in distribution, increased in number, and thereafter decreased. p53 immunoreactivity was localized primarily to the nucleus but not to cytoplasm. Double-staining with glial cell markers revealed that p53 immunoreactivity was often co-localized in microglia, oligodendrocytes and astrocytes, but not in neurons. In view of the results of the double-staining of p53 and Bcl-2, Bax or TUNEL, a variety of apoptosis related molecules are expressed with p53, all within the first three days of injury. Further, the process of apoptosis via the p53, pathway appears complex even in this simple model of CNS injury. Our study suggests that the manipulation of these apoptosis-related molecules may prove useful in modifying the cell and tissue damage in traumatic CNS injury. PMID- 10709876 TI - Myocardial growth before and after birth: clinical implications. AB - Perinatal changes in myocardial growth have recently evoked considerable interest with regard to cardiac chamber development with congenital cardiac lesions and to myocardial development in preterm infants. It is suggested that cardiac chamber development is influenced by blood flow. Experimental pulmonary stenosis in fetal lambs may induce either greatly reduced or markedly increased right ventricular volume. Ventricular enlargement appears to be associated with a large ventricular volume load resulting from tricuspid valve regurgitation. A small competent tricuspid valve is associated with reduced flow through the ventricle due to outflow obstruction and a small right ventricle. Postnatal growth of the ventricles in congenital heart disease is discussed. Increase in myocardial mass prenatally is achieved by hyperplasia, both during normal development and when myocardial mass is increased by right ventricular outflow obstruction. Postnatally, increases in myocardial mass with normal growth, as well as with ventricular outflow obstruction, are largely due to hypertrophy of myocytes. Myocardial capillary numbers do not increase in proportion with myocyte numbers in ventricular myocardium in association with outflow obstruction. The postnatal effects of these changes in congenital heart lesions are considered. Studies in fetal lambs suggest that the late gestational increase in blood cortisol concentrations is responsible for the change in the pattern of myocardial growth after birth. The concern is raised that prenatal exposure of the premature infant to glucocorticoids, administered to the mother to attempt to prevent hyaline membrane disease in the infant, may inhibit myocyte proliferation and result in a heart with fewer than normal myocytes. This would necessitate that each myocyte would have to hypertrophy abnormally to achieve a normal cardiac mass postnatally. PMID- 10709877 TI - Psychosocial adjustment to cancer treatment and other chronic illnesses. PMID- 10709878 TI - Growth patterns of breastfed infants. PMID- 10709879 TI - Intensive nursing care of kwashiorkor in Malawi. PMID- 10709880 TI - The controversial epidemiology of coeliac disease. PMID- 10709881 TI - Long-chain polyunsaturated fatty acids in human milk and brain growth during early infancy. AB - Long-chain polyunsaturated fatty acids are essential for growth and development, and their crucial role in the development of the central nervous system and in retinal function has been the subject of many studies. As the balance between n-6 and n-3 fatty acids has to be optimal, their concentrations in the milk given to infants who are exclusively breastfed is of major importance. In this study, the composition of fatty acids in mothers' milk and the growth rate of the infant brain were analysed. Nineteen mother-term infant pairs from Stockholm, Sweden, were studied from birth to 1 mo and 3 mo of age, during which time the infants were breastfed exclusively. The dietary intake of the mothers was calculated and found to concur with the recommended daily dietary allowances of Swedish lactating women as regards energy, protein, fat and carbohydrates. The amounts of linoleic acid and alpha-linolenic acid in the diet were similar to those reported for European and North American women. The ratio between arachidonic acid (AA) and docosahexaenoic acid (DHA) in the milk from Swedish mothers is approximately the same as in the brain of infants, and was found to be positively correlated with the rate of gain of the occipito-frontal head circumference and of the calculated brain weight at 1 mo (p < 0.01) and 3 mo (p < 0.01) of age, respectively. However, further studies are needed to establish the exact requirements of AA and DHA for optimal growth and development during early infancy in exclusively breastfed infants. PMID- 10709882 TI - Effects of growth during infancy and childhood on bone mineralization and turnover in preterm children aged 8-12 years. AB - To investigate the effect of growth on later bone mass and turnover, bone mineral content (BMC) and density (BMD: dual X-ray absorptiometry (QDR 1000W) and single photon absorptiometry (Lunar SP2)) and bone turnover (plasma osteocalcin, urine deoxypyridinoline) were measured at 8-12 y in 244 preterm children who had weight and height measured at 18 mo and 7.5-8 y corrected age. Weight and length at birth, 18 mo, 7.5-8 y and current follow-up showed increasingly strong, positive correlations with bone area, BMC and BMD. After adjusting for current size, there were significant negative associations between earlier size measurements and later whole body and lumbar spine bone mass which were stronger for length than for weight, and a negative relationship between birthweight for gestation and later radial bone mass; but no relationship with bone turnover. Current calcium intake and activity level had no independent effect on bone mass. Bone mass at 8 12 y is related to current bone and body size, which tracks throughout childhood. However, amongst children of the same current size, those who have shown the greatest increase in size, particularly in height, have the highest bone mass. These findings raise the hypothesis that improving linear growth in vulnerable children may be important in maximizing bone mass. PMID- 10709883 TI - Effect of helicobacter pylori eradication on sideropenic refractory anaemia in adolescent girls with Helicobacter pylori infection. AB - We investigated the effect of Helicobacter pylori eradication on sideropenic refractory anaemia in adolescent girls with H. pylori-associated antral gastritis without evidence of haemorrhage or clinical symptoms other than sideropenic anaemia. We conducted an open therapeutic trial in 21 adolescent girls aged 15-17 y with sideropenic refractory anaemia, which was defined as iron-deficiency anaemia refractory to oral iron therapy for 3 mo. All subjects underwent gastroduodenal endoscopy. Thirteen patients with confirmed H. pylori infection were given a 2-wk course of triple therapy and 6 wk of oral ferrous sulfate. We compared the mean levels of haemoglobin and serum ferritin among the "initial sample" (the time when the sampling was done before treatment with oral iron), "before eradication" (the time prior to triple therapy for eradication after subjects had been given oral iron for 3 mo) and "after eradication" (the time when the follow-up endoscopy was performed) data in 11 subjects in whom H. pylori infection was eradicated. Haemoglobin levels (g/dl) were 8.6+/-1.9, 8.6+/-1.4 and 11.3+/-2.3, respectively. Serum ferritin levels (microg/l) were 4.6+/-2.4, 4.2+/ 2.3 and 17.5+/-6.7, respectively. After eradication of H. pylori, mean levels of haemoglobin (p = 0.0002) and serum ferritin (p = 0.0002) showed a prominent increase between "before eradication" and "after eradication". In conclusion, adolescent girls with sideropenic refractory anaemia should be evaluated for H. pylori infection. If H. pylori infection is coexistent, its eradication along with iron supplementation could correct the anaemia. PMID- 10709884 TI - Protein intake and metabolism in formula-fed infants given Swedish or Italian weaning foods. AB - The aim of the study was to compare protein intake and metabolism between infants from two countries given similar infant formulae but different weaning foods. Healthy Swedish and Italian infants were studied between 3 and 12 mo. Infants in both populations were assigned to 1 of 3 infant formulae, containing 13, 15 or 18/20 g l(-1) of protein, given in addition to Swedish or Italian weaning foods. Protein intake from weaning foods was higher in Italian than in Swedish infants at 6 and 12 mo, whereas protein intake from formula at 6 mo and from formula/milk at 12 mo was similar in both populations. Plasma isoleucine, leucine, lysine, histidine and valine at 6 mo were lower in Italian than in Swedish infants fed formula with 13 g l(-1) of protein. All essential plasma amino acids were similar in Italian and Swedish groups at 12 mo. Serum urea was similar at 6 mo in corresponding formula groups, but was higher at 12 mo in the Italian than in the Swedish formula group. Serum albumin and growth were normal in both populations throughout infancy. In conclusion, formula with protein content of 13 g l(-1) seems to provide sufficient protein intake when combined with Swedish or Italian weaning foods during the second half of infancy, as indicated by normal serum albumin and normal growth. However, the bioavailability of protein and amino acids from weaning foods, in addition to their protein content, should be considered, as indicated by some indices of protein metabolism in the Italian infants. PMID- 10709885 TI - Epidemic of coeliac disease in Swedish children. AB - Coeliac disease has emerged as a public health problem. The aim of the present study was to analyse trends in the occurrence of symptomatic coeliac disease in Swedish children from 1973 to 1997, and to explore any temporal relationship to changes in infant dietary patterns. We established a population-based prospective incidence register of coeliac disease in 1991, and, in addition, retrospective data from 1973 were collected. A total of 2151 cases fulfilled the diagnostic criteria. Furthermore. We collected national data on a yearly basis on duration of breastfeeding, intake of gluten-containing cereals and recommendations on when and how to introduce gluten into the diet of infants. From 1985 to 1987 the annual incidence rate in children below 2 y of age increased fourfold to 200-240 cases per 100000 person years, followed from 1995 by a sharp decline to the previous level of 50-60 cases per 100000 person years. This epidemic pattern is quite unique for a chronic disease of immunological pathogenesis, suggesting that prevention could be possible. The ecological observations made in this study are compatible with the epidemic being the result, at least in part, of a change in and an interplay among three factors within the area of infant feeding, i.e. amount of gluten given, age at introduction of gluten, and whether breastfeeding was ongoing or not when gluten was introduced. Other factor(s) may also have contributed, and the search for these should be intensified. PMID- 10709886 TI - Fatty acid composition of plasma lipids in HIV-infected children. Comparison with seroreverters. AB - Children infected with the type-1 human immunodeficiency virus (HIV) are at risk of nutritional deficiencies leading to an impaired polyunsaturated fatty acid (PUFA) status. The aim of the present study was to compare the PUFA composition of plasma lipid classes (total lipids, phospholipids (PL), cholesteryl esters (CE) and triglycerides) in well-growing HIV-infected children with an age-matched group of HIV-seroreverter children born to infected mothers. Eighteen HIV children, of both sexes, mean age 4.6 y, most of whom under combined antiretroviral regimen, were compared with 18 seroreverters, mean age 5.4 y, comparable for demographic, anthropometric and dietary characteristics. All children had adequate growth parameters (weight and height > 3rd percentile). The plasma fatty acid content was similar in the two groups. HIV seropositive subjects showed lower linoleic acid (LA) levels in all the plasma lipid fractions, with higher 20:3n-9 and 20:5n-3 levels in PL and CE. The plasma PL triene/tetraene ratio (marker of relative LA deficiency) related positively to the viral load and negatively to the blood CD4+ lymphocyte count. Compared to age matched seroreverter subjects, HIV-seropositive children show a lipid fatty acid status suggestive of relative LA deficiency and increased turnover of the PUFA series. PMID- 10709887 TI - Clinical, laboratory and molecular characteristics of children with Familial Mediterranean Fever-associated vasculitis. AB - Familial Mediterranean Fever (FMF) is an autosomal recessive disease characterized by recurrent self-limited attacks of fever accompanied by peritonitis, pleuritis and arthritis. Approximately 5% of individuals with FMF have been reported to have Henoch-Schonlein purpura (HSP) and about 1% have polyarteritis nodosa (PAN). Protracted febrile myalgia is another vasculitis associated clinical entity among patients with FMF. Recently, the gene responsible for FMF, MEFV, has been cloned and four missense mutations (M680I, M694V, V726A and M694I) have been described. In this report, we present clinical and laboratory findings and mutation results of 23 children with FMF-associated vasculitis. HSP, PAN and protracted febrile attacks have been diagnosed in 11, 2 and 10 children, respectively. Mutation analysis shows that 3 children are homozygotes for the M694V mutation and 11 are compound heterozygotes for 2 of the studied mutations. M694V/V726A mutations were identified in 8, M694V/M694I in 2 and M680I/M694V in 1 of these children. In six children only one mutation was found and in three none of the studied mutations were identified. This study confirms that most children with FMF-associated vasculitis have identifiable mutations in the MEFV gene. Environmental and/or other genetic factors are possibly involved in the pathogenesis of vasculitis in FMF; elucidation of these mechanisms will help to understand pathogenesis of childhood vasculitides. PMID- 10709888 TI - Long-term outcome of classical 21-hydroxylase deficiency: diagnosis, complications and quality of life. AB - A nationwide search of patients with classical 21-hydroxylase deficiency (21-OHD) was performed in Finland to determine the long-term outcome of the disease. In total, 108 patients were found. Fifty-four patients (50%, 31F, 23M) had deficiency of a salt-wasting form and another 54 (50%, 29F, 25M) had a simple virilizing form of 21-OHD. A significant number of severe complications suggestive of glucocorticoid deficiency was found. There were five deaths (4.6% of all) possibly connected with cortisol deficiency. Ten additional patients (9.3% of all) had been acutely admitted 14 times in all due to symptoms of glucocorticoid deficiency. These symptoms included sudden loss of consciousness, convulsions and severe fatigue. Afterwards, permanent neurological defects were detected in two of these patients. Finally, a cross-sectional study was carried out to establish an estimate of the long-term outcome of the disease. Thirty-two (55%) of the 58 patients aged 16 y or more participated in this study. The patient group did not differ from the general Finnish population in terms of education. Three of the patients (5%) had retired prematurely. Surprisingly, the patients felt that their health-related quality of life, as reported in the RAND 36 questionnaire, was better than that of the general Finnish population (p = 0.023). However, as a significant number of all patients did not participate in this study, the quality of life evaluation results must be interpreted with caution. In conclusion, a significant number of complications was found among patients treated for classical 21-OHD. Nevertheless, the disease has a favourable outcome in terms of quality of life. PMID- 10709889 TI - Pamidronate treatment of bone fibrous dysplasia in nine children with McCune Albright syndrome. AB - McCune-Albright syndrome is a rare genetic disorder consisting of skin and bone dysplasia and peripheral endocrinopathies. Little data have been collected regarding bisphosphonate treatment of bone fibrous dysplasia in paediatric patients with this syndrome. The aim of our study was to investigate the therapeutic efficacy of pamidronate in these patients. Nine patients with moderate to severe forms of bone fibrous dysplasia were treated with pamidronate intravenously (0.5-1 mg/kg/daily for 2-3 d) at 0.5-1-y intervals. Patients were treated over a time period of 0.5-3.5 y. During treatment no spontaneous fracture occurred. Bone pain and gait abnormality due to pain disappeared after 2-3 therapeutic cycles. Cranial asymmetry and limb length discrepancy remained unchanged. Elevated serum alkaline phosphatase and urine hydroxyproline values were reduced by the treatment, demonstrating drug activity at the lesional level. The effectiveness of pamidronate was also seen at the non-lesional level through an increase in bone density. Radiographic and scintigraphic evidence of lesion healing was not attained. Pamidronate treatment can ameliorate the course of bone fibrous dysplasia in children and adolescents with McCune-Albright syndrome. PMID- 10709890 TI - Utility of brainstem auditory evoked potentials in children with spastic cerebral palsy. AB - In a retrospective study of 75 children with spastic cerebral palsy (CP), brainstem auditory evoked potentials (BAEP) were recorded and subsequently correlated with birthweight, gestational age, aetiology and type of CP, neuroradiological findings, additional impairments and disabilities (including the inability to walk independently). Seventeen patients (22.7%) had abnormal BAEP recordings. Thirteen of these 17 patients (76.5%) had spastic tetraplegia, 16 patients (94.1%) were full-term infants, 12 patients (70.6%) had myoskeletal problems, 9 (52.9%) had epilepsy, 16 (94.1%) had visual impairment, 13 patients (76.5%) were unable to walk independently, while all 17 patients (100%) had speech impairment and mental retardation. The aetiology of CP was prenatal in 2 of these 17 patients (11.8%) and perinatal in 15 patients (88.2%). Thirteen patients (76.5%) had cortical atrophy determined by either computed tomography or magnetic resonance imaging, two patients (11.8%) had an infarct picture and two patients (11.8%) had maldevelopment of the central nervous system. There was a definite statistically significant association between abnormal BAEP recordings and full-term delivery, perinatal aetiology of CP, spastic tetraplegia, speech, visual and myoskeletal impairments, epilepsy, mental retardation, inability to walk independently and cortical atrophy on neuroimaging (p < 0.001). We conclude that abnormal BAEP recordings in children with spastic CP are indicative of poor prognosis and associated with a "multihandicap state". BAEP testing should be incorporated into the diagnostic plan of all children with spastic CP newly referred to neurodevelopmental centres. PMID- 10709891 TI - Head growth in Rett syndrome. AB - The longitudinal development of head growth was investigated in girls with Rett syndrome. Measurements were taken retrospectively from different kinds of records. Growth retardation was expressed in standard deviation (SD) scores. In classic types, the mean head circumference fell successively to 2 SD scores below the norm at the age of 4 y. After the age of 8 y it stabilized close to -3 SD scores. The degree of deceleration correlated strongly to the age at which a deceleration of 1 SD score had occurred. In forme fruste variants, the mean head circumference was within normal limits; however, it was significantly below the norm (-0.8 SD scores). Body height deviated to -2 SD scores at the age of 6 y and was highly correlated to decline in head growth. When head growth was related to the severity of motor disability, there was a continuum from almost normal head growth with well-preserved gross motor function and some preserved fine motor function to a marked deceleration in head growth with maximum gross and fine motor disability. PMID- 10709892 TI - Intensive nursing care of kwashiorkor in Malawi. AB - The case fatality rate for children with kwashiorkor in central hospitals in Malawi was 30.5% (275/901) in 1995. The purpose of this study was to determine whether improved case management with intensive nursing care could lower this case fatality rate. A total of 75 children admitted with kwashiorkor in Blantyre, Malawi, received intensive nursing care. This included nursing in individual clean beds with blankets, a nurse:child ratio of 1:3, supervised feedings every 2 h, a paediatrician with expertise in treating kwashiorkor always available for consultation, laboratory evaluation for systemic infection and empiric use of ceftriaxone. Nineteen of these children died (25%). The causes of death were life threatening electrolyte abnormalities (hypokalaemia, hyponatraemia, hypophosphataemia) in nine cases, overwhelming infection in eight cases and congestive heart failure in two children. Children infected with the human immunodeficiency virus were more likely to die (9/20), as were children with life threatening electrolyte abnormalities (9/15) and children with more severe wasting. When compared with 225 children treated in the same year at the same institution, who were carefully matched for severity of kwashiorkor, intensive nursing did not improve overall survival. PMID- 10709893 TI - Living conditions in early infancy in Denmark, Norway and Sweden 1992-95: results from the Nordic Epidemiological SIDS study. AB - The objective was to study living conditions of infants and their families in Scandinavia in the 1990s and to assess similarities and differences among the three Scandinavian countries. The emphasis is on health and normality rather than on diseases and other deviations from well-being. The subjects are the 869 controls in the Nordic Epidemiological SIDS study carried out between 1 September 1992 and 31 August 1995 in Norway, Denmark and Sweden. The controls were matched with the 244 SIDS cases for sex, age and maternity hospital. Parents of the SIDS cases and the controls filled in the same questionnaire on family, pregnancy, delivery, the neonatal and the post-perinatal period. The most striking findings were that 99% of the mothers went to regular maternity controls, 97% to well-baby clinics and 84% breastfed exclusively. On the other hand, 11% drank alcohol more than once a month during pregnancy and 29% smoked during pregnancy. Compared to official statistics, to the extent they exist, the differences were small. The material contains valuable information on normal infant care in Scandinavia in the 1990s. Living conditions of infants in Scandinavia are similar in the three countries. Differences exist, but only to a small extent. PMID- 10709894 TI - Growth patterns of breastfed infants in seven countries. AB - An international effort is underway to develop a new international growth reference for assessing the growth of young children, especially breastfed infants who appear to falter relative to the currently recommended National Center for Health Statistics/World Health Organization reference. While limited data from high socioeconomic status children from different parts of the world suggest that their growth patterns are similar, there is no comprehensive study of breastfed infants. The WHO Multinational Study of Breastfeeding and Lactational Amenorrhea provides bi-weekly weights and 2-4 weekly length measurements on breastfed babies from selected sites in Australia, Chile, China, Guatemala, India, Nigeria and Sweden. Multi-level modelling was used to analyse between-site differences in the growth of approximately 120 infants per site, after adjustment for maternal stature and infant feeding pattern. All mothers were literate and mean educational levels were well above national averages, but the study was not restricted to infants of high socioeconomic status. Maternal education was significantly associated with infant weight only in India. The growth curves of infants from most sites were strikingly similar, but relative to the Australians (the reference category), the Chinese babies were about 3% shorter at 12 mo of age and the Indians up to 15% lighter. The present results suggest that breastfed babies from reasonably well-off families in different continents show very similar growth patterns. However, it is important that the growth of children from South and East Asian populations be rigorously assessed in the process of developing the new international growth reference. This paper discusses the relative importance of environmental versus genetic influences in the growth of young children and illustrates the complexities involved in the analysis of growth data. PMID- 10709895 TI - Sleep problems of school-aged children: a complementary view. AB - The aim of this population-based multicentre study was to evaluate the prevalence rates of sleep problems among 8-9-y-old children. The sample consisted of 5813 Finnish children, making up 10% of the age cohort. Both parents and children provided information. Disturbed sleep was reported by 21.7% of parents. Most of the problems were mild; only 0.3% were serious. Dyssomnias were frequent: 11.1% had difficulties with sleep onset, 7.1% with night waking and 2.3% with waking too early. Multiple sleep problems were present in 9.1% of the children. 17.8% of children reported disturbed sleep, 12.7% had problems many nights and 5.1% every night. In 32.0% of cases, either the parent or the child reported disturbed sleep; 7.4% of these reports came from both the parent and the child, 14.1% from the parent only and 10.3% from the child only. The correspondence between informants was poor (kappa = 0.224). Sleeping problems were associated with somatic and psychiatric problems. It is concluded that by restricting questioning to parents only, one-third of all potential cases of sleep problems may go unnoticed. In order to increase the sensitivity of screening children's sleep problems, both parents and children should provide information in epidemiological settings as well as in clinical work. PMID- 10709896 TI - Self-esteem, depression and anxiety among Swedish children and adolescents on and off cancer treatment. AB - Self-esteem, depression and anxiety were investigated in 51 Swedish children and adolescents, 8-18 y, on (n = 16) and off (n = 35) cancer treatment. The self report measures "I Think I Am" (ITIA), the Children's Depression Inventory (CDI) and the Revised Children's Manifest Anxiety Scale (RCMAS) were used. Data were compared with data previously obtained by others for healthy Swedish children. Children and adolescents on treatment showed levels of self-esteem, depression and anxiety comparable to those of healthy children. However, children and adolescents off treatment reported higher depression and anxiety levels and lower psychological well-being and physical self-esteem than have been reported for healthy Swedish children. Seven children (14%) reported a high level of depression, six of whom were off treatment. The findings suggest that the period after treatment termination is characterized by a higher risk of psychosocial problems than is the actual treatment period. PMID- 10709898 TI - Surgery is safe in very low birthweight infants with necrotizing enterocolitis. AB - The aim of this study was to determine how the operative event itself affects very low birthweight (VLBW) infants (< 1500 g) with necrotizing enterocolitis (NEC) undergoing surgery, and to try to identify preventable factors leading to perioperative morbidity and mortality. Eighty-five VLBW infants developed NEC during a 6-year period; 34 of those required emergent celiotomies. Data were collected retrospectively from hospital charts available on 33 infants. Birthweight ranged from 566 g to 1415 g (mean +/- SD: 961+/-262 g) and gestational age from 24 to 34 wk (28+/-3.2 wk). Thirty infants had been fed premature formula (first feed at 5+/-3.6 d) prior to the onset of symptoms and three had not been fed at all. Age at NEC symptoms was 19+/-15 d. Infants < 1000 g developed NEC much longer after the first feed compared to infants > 1000 g (p < 0.002; t-test). In 42% of the children, intraoperative blood pressure fell at least 20% from the preoperative value. Body temperature dropped from a preoperative 36.5+/-0.340 degrees C to 35.5+/-1.20 degrees C (p < 0.005), although in all children two or more heating devices were employed in the operating room. All infants survived the procedure. Six infants with pannecrosis died within 72 h of the operative event. In an appropriate setting, operative intervention under general anesthesia is well tolerated by VLBW infants with NEC. Since hypothermia was a major problem, the authors have modified their approach and now no longer transport these infants to the operating room. Instead, these infants are operated upon in the neonatal intensive care unit, directly on an infant radiant warmer system. PMID- 10709897 TI - Randomized trial of two levels of fluid input in the perinatal period--effect on fluid balance, electrolyte and metabolic disturbances in ventilated VLBW infants. AB - The aim of this study was to determine whether fluid restriction does indeed significantly increase acute adverse effects. One-hundred-and-sixty-eight ventilated infants, median gestational age 27 wk (range 23-33) and birthweight 953 g (range 486-1500), entered into a randomized controlled trial of two fluid regimes. Infants on regime A were to be prescribed 60 ml/kg of fluids on day 1 which was gradually increased over the first week to 150 ml/kg, infants on fluid regime B were to be prescribed approximately 20% less fluid over the first week. Daily fluid input and output were recorded. Serum electrolytes, bilirubin, creatinine and urine osmolalities were measured daily. Arginine vasopressin levels were assessed on days 1, 3 and 5. Episodes of jaundice, hypoglycaemia and hypotension requiring treatment were noted. Infants on regime B actually received overall 11% and, in the first 4 days, 19% less fluid than those on regime A (p < 0.001). There were no statistically significant differences in the occurrence of episodes of jaundice, hypotension, hypoglycaemia, hypernatraemia or hyponatraemia between infants on the two regimes. Although the infants on regime B had significantly higher urine osmolalities and lower urine output for most of the perinatal period, their median creatinine and arginine vasopressin levels did not differ significantly from those on regime A. We conclude that fluid restriction to less than 90% of usual maintenance fluids is not associated with an excess of acute adverse effects. PMID- 10709899 TI - A local cluster of achalasia in a province of Crete. AB - Achalasia has a largely obscure aetiology and is uncommon in childhood. We report three cases of otherwise well children, residents of a small province of Crete, two of them female cousins. This cluster probably indicates an autosomal recessive trait of inheritance. All three children were surgically treated (Heller cardiomyotomy combined with Nissen fundoplication), with excellent results. PMID- 10709900 TI - Transient recovery of endogenous immune function following haploidentical peripheral stem cell transplantation in a patient with severe combined immunodeficiency without evidence of engraftment. PMID- 10709901 TI - Multivariate analysis-based prediction rule for pulmonary embolism. AB - The diagnosis of pulmonary embolism (PE) is still an unresolved problem. The aim of this prospective observational study was to derive and validate a prediction rule (PEscore) by which PE can be diagnosed by easily obtainable and rapidly available investigations. Included were consecutive patients with a clinical suspicion of PE admitted to a community hospital. Risk factors and clinical and instrumental investigations were registered. PE was diagnosed by angiography, scintigraphy, or autopsy. In 168 patients, PE was either diagnosed (angiography, n = 28; autopsy, n = 18) or excluded (angiography, n = 12; scintigraphy, n = 99; autopsy, n = 11). Based on the results of clinical and instrumental findings, a PEscore was derived by a multiple regression analysis, calculated as: [0.29 x proven leg vein thrombosis (0 = no, 1 = yes)] + [0.25 x ECG right heart strain (0 = no, 1 = yes)] + [0.22 x neck vein distension (0 = no, 1 = yes)] + [0.20 x dyspnoea (0 = no, 1 = yes)] + [0.13 x suspicious chest X-ray (0 = no, 1 = yes)] - [0.17 (constant)]. The PEscore was tested further in 139 subsequent cases. In these patients, the PEscore was 0.65+/-0.17 (diagnosed PE, n = 47) and 0.18+/ 0.17 (excluded PE, n = 92), respectively (p = 0.0001). Depending on a given PE score, the level of probability of PE can be assessed. Calculation of the PEscore can be helpful in clinical decisions when PE is suspected. PMID- 10709902 TI - The -323Ins10 polymorphism for factor VII is not associated with coronary atherosclerosis in symptomatic men. The REGRESS study group. AB - Elevated factor VII coagulant activity (FVII:C) has been associated with an increased risk of ischaemic heart disease, particularly for fatal events. Results of studies on the association between FVII:C and atherosclerosis are not consistent. FVII:C levels are influenced by several environmental factors and by genetic factors. One of the genetic factors is the -323Ins10 polymorphism in the promoter region of the factor VII gene, which is strongly related to FVII:C, and thus may be associated with ischaemic heart disease. We studied the association of this polymorphism with the severity and progression of atherosclerosis. In 511 male patients of the Regression Growth Evaluation Statin Study, the genotype for the -323Ins10 polymorphism was determined. The minimum obstruction diameter and the mean segment diameter were determined at baseline and after a 2-year follow up period, and new lesion formation was assessed as well. Cardiovascular events were recorded. No relationship was observed between the -323Ins10 polymorphism and angiographic measures of disease progression, nor on the risk of new cardiovascular events. The results suggest that there is no association between the -323Ins10 polymorphism for factor VII and the severity or progression of coronary atherosclerosis in male patients with symptomatic coronary artery disease. PMID- 10709903 TI - Effect of iron therapy on the whole blood platelet aggregation in infants with iron deficiency anemia. AB - This study was performed to investigate the platelet aggregation alterations in whole blood samples of infants with iron deficiency anemia. Platelet aggregation induced by various concentrations of adenosine diphosphate (ADP) and collagen was studied with impedance aggregometry in 25 patients before and after oral iron therapy and in 12 children of the control group. The posttreatment mean maximum aggregation values were significantly higher (p<0.01) and the posttreatment mean aggregation times were significantly lower (p<0.01) in the study group at all concentrations of ADP and collagen. The aggregation time and maximum aggregation values revealed no significant difference except for the maximum aggregation value at 5 microM ADP (p<0.05) between the study group after therapy and the control group. The differences between the pretreatment and posttreatment mean platelet counts and mean platelet volume values in the study group were statistically significant (p<0.01), whereas those values in the study group after therapy and in the control group were not significantly different. We conclude that iron deficiency anemia in infants, even without clinically meaningful platelet abnormality, may cause dysfunction of the ex vivo whole blood platelet aggregation, and can be reversed by iron therapy. Further studies should be carried out at the enzymatic level to determine whether this platelet aggregation dysfunction in iron deficiency anemia is due to a deficiency in the activation of iron-containing enzymes. PMID- 10709904 TI - Platelet aggregation induced by ADP or epinephrine is enhanced in habitual smokers. AB - A novel system has been developed to evaluate biochemically induced platelet aggregation by means of a particle-counting technique that uses laser light scattering. Using this system, we compared the differences in platelet aggregability between 90 smoking healthy males after 10 hours of smoking abstinence and 141 age-matched nonsmoking healthy males. Smokers had more small spontaneous platelet aggregates and more medium and large aggregates induced by 1 or 5 microM of epinephrine than nonsmokers. No large aggregates with 5-microM epinephrine-induced platelet aggregation were seen in 10% of smokers and 24% of nonsmokers; these subjects showed no small aggregates in spontaneous aggregation at all. Smokers had significantly more small, medium, and large aggregates induced by 1 microM of adenosine diphosphate (ADP) than nonsmokers. Smokers showed a positive correlation between age and 1-microM epinephrine-induced large platelet aggregates, percent reduction of optical density, and 1-microM ADP induced medium and large aggregates. Smokers also showed a positive correlation between fibrinogen concentration in plasma and small spontaneous aggregates. On the other hand, nonsmokers showed a significant positive correlation between age and small spontaneous aggregates, and a positive correlation between fibrinogen and 1- or 5-microM epinephrine-induced large aggregates, and between 1 microM ADP induced large aggregates and percent reduction of optical density. These results confirmed that platelet aggregability is enhanced in smokers, and we speculate that long-term smoking might enhance the sensitivity of platelets to epinephrine or ADP. PMID- 10709905 TI - Production of macromolecular activators of phagocytosis by lysed platelets. AB - Macromolecular activators of phagocytosis from platelets (MAPP: 1-MAPP and s MAPP) are released from activated fresh platelets and enhance leukocyte phagocytosis via the Fcgamma receptors. In this study, production of MAPP was investigated in lysate of freeze-thawed stored platelets (PL). Incubation of PL and thrombin with precursors of MAPP (pre-MAPP: pre-1-MAPP and pre-s-MAPP) produced 1-MAPP and s-MAPP, whereas products released from stored platelets by stimulation with thrombin or collagen did not produce MAPP after incubation with pre-MAPP. The action of thrombin in MAPP formation with PL and pre-MAPP was inhibited by antithrombin III and heparin, and sequential incubation studies indicated that the key site of action of thrombin was on a component of PL. Other serine proteases such as trypsin could be substituted for thrombin in this reaction, whereas the action of thrombin was specific when whole platelets were used instead of PL. Gel filtration of PL before and after treatment with thrombin suggested that a macromolecule in PL (PMA-I) is digested by thrombin and liberates a 700 to 800 Da substance (PMA-II) which converts pre-MAPP to MAPP. PMID- 10709906 TI - Antiplatelet and antithrombotic effects of orbofiban, a new orally active GPIIb/IIIa antagonist, in guinea pigs. AB - Antiplatelet and antithrombotic effects of orbofiban, a new orally active glycoprotein IIb/IIIa antagonist, were evaluated in guinea pigs. SC-57101A (0.03 1 microM), the hydrochloride salt of the active form of orbofiban, inhibited in vitro ADP- and collagen-induced platelet aggregation in a concentration-dependent manner. Oral administration of orbofiban (3-30 mg/kg) resulted in dose-dependent inhibition of ADP- and collagen-induced platelet aggregation. The inhibition peaked at 1-2 hours postdose and then declined slowly. The agent also showed similar inhibition of platelet aggregation in guinea pigs with dietary-induced hypercholesterolemia. In contrast, the antiaggregatory effects of acetylsalicylic acid differed more widely between normal and hyperlipidemic animals compared to those of orbofiban. Plasma concentration of the active form, measured by a column switching HPLC method, correlated well with the inhibition of platelet aggregation. Orbofiban (3-100 mg/kg, p.o.) caused dose-dependent inhibition of thrombus formation in an arteriovenous-shunt-thrombosis model. Orbofiban at high doses (> or =30 mg/kg) and acetylsalicylic acid (100 mg/kg) both prolonged cutaneous bleeding time measured by the template method. These results demonstrate that orbofiban is an orally active and potent inhibitor of platelet aggregation with an efficacy that correlates well with the plasma concentration of its active form. PMID- 10709907 TI - PS-liposome and ox-LDL bind to different sites of the immunodominant domain (#155 183) of CD36: a study with GS95, a new anti-CD36 monoclonal antibody. AB - CD36, a multifunctional adhesive receptor on a variety of cells such as monocytes and platelets, has been implicated in clearance of modified LDL and in the removal of apoptotic or senescent cells. We recently developed a new anti-CD36 monoclonal antibody, GS95. We determined the binding site of phosphatidylserine (PS)-liposome on CD36 by flow cytometric analysis of competitive bindings between phospholipid-liposomes or synthetic CD36 peptides and FITC-labeled anti-CD36 antibodies (GS95, OKM5, and FA6-152). The epitope of GS95 was mapped to the amino acid sequence #162-183 of CD36 that was partially overlapped with, but distinct from, #155-183, which has been reported as the epitopes of two commercially available antibodies, OKM5 and FA6-152. Oxidized-LDL dose-dependently inhibited bindings of both GS95 and OKM5 antibodies to platelet CD36, while PS-liposome inhibited the binding of GS95 but not OKM5 or FA6-152. These results indicate that the binding site of PS-liposome on platelet CD36 is not identical to that of oxidized-LDL and may be located in the amino acid sequence #162-183. PMID- 10709908 TI - Adjuvant effect of antibodies against von Willebrand Factor, fibrinogen, and fibronectin on staphylokinase-induced thrombolysis as measured using mural thrombi formed in rat mesenteric venules. AB - The change in thrombus mass during thrombolytic therapy is thought to be the difference between its growth and its degradation induced by thrombolytic agents. Platelets play a pivotal role in arterial thrombosis and bind to each other and to exposed subendothelial matrices via adhesive proteins such as von Willebrand Factor, fibrinogen, and fibronectin. The aim of the present study was to assess whether administration of antibodies against these adhesive proteins, in conjunction with plasminogen activator, could enhance the degradation of platelet rich thrombus. Mural platelet-rich thrombi were formed in rat mesenteric venules using He-Ne laser irradiation. Recombinant staphylokinase was infused continuously and polyclonal antibodies against adhesive proteins were given by bolus injection. The thrombolytic process was analysed using computer-enhanced image analysis software. Administration of each of the antibodies enhanced staphylokinase-induced thrombolysis. PMID- 10709909 TI - Expression of prothrombin fragment 1+2 in cancer tissue as an indicator of local activation of blood coagulation. AB - Immunohistochemistry was applied to AMeX-fixed tissue sections of 12 adenocarcinomas of the stomach (seven intestinal adenocarcinomas and five diffuse carcinomas), 12 adenocarcinomas of the pancreas (nine ductal adenocarcinomas and three signet ring carcinomas), and 12 squamous cell carcinomas of the larynx obtained at surgical resection to examine the possibility of extravascular activation of blood coagulation in cancer tissues by exploring the in loco patterns of distribution of fibrinogen, a final product of blood coagulation, fibrin, and a by-product of coagulation reactions (prothrombin fragment 1+2). Gastric, pancreatic, and laryngeal cancers exhibited fibrinogen antigen in abundance throughout the tumor stroma. Fibrin was detected along the edges of nests of carcinoma cells and at the host-tumor interface. Prothrombin fragment 1+2 was present in the blood vessels in areas of neoangiogenesis at the host tumor interface (gastric and pancreatic cancer tissues) and on the tumor cell bodies (pancreatic and laryngeal cancer tissues). The presence of prothrombin fragment 1+2 in cancer tissues appears to be a good indicator of coagulation activation and thrombin generation at the tumor burden. PMID- 10709910 TI - Induction of urokinase-type plasminogen activator by lipopolysaccharide in PC-3 human prostatic cancer cells. AB - Urokinase-type plasminogen activator (uPA) plays a central role in many aspects of cellular regulation, such as fibrinolysis, cell migration, and metastasis. The uPA induction by inflammatory stimuli such as IL-1, TNFalpha, and lipopolysaccharide (LPS) has been reported in a number of human cells. We examined the effects of LPS on uPA expression in human PC-3 prostatic cancer cells that express uPA in a conditioned medium. LPS increased the uPA accumulation in PC-3 cells, whereas IL-1, IL-6, and TNFalpha did not. Northern blot analysis revealed that the peak induction of uPA mRNA occurred 5 hours after LPS stimulation. A protein synthesis inhibitor, cycloheximide, amplified the LPS induced uPA mRNA, suggesting that LPS induces uPA by activating the gene expression in which de novo protein synthesis is not necessary. PMID- 10709911 TI - The potential mechanism for the effect of heparin on tissue plasminogen activator mediated plasminogen activation. AB - The effects and possible role of heparin on tissue plasminogen activator-mediated plasminogen activation was thoroughly investigated. Direct analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis demonstrated that heparin increased the conversion of plasminogen to plasmin. Experiments by fluorescence quenching suggested that the stimulation of tissue plasminogen activator activity probably was due to a direct binding of heparin to tissue plasminogen activator, causing a conformational change of tissue plasminogen activator and rendering it more accessible to plasminogen interaction. The absence of additive stimulation effects on tissue plasminogen activator-mediated plasminogen activation when both heparin and fibrinogen were present also implied that both compounds interacted with tissue plasminogen activator via the same domain; it appeared to be most likely via the kringle-2 domain in tissue plasminogen activator based on studies using epsilon-aminocaproic acid as an inhibitor. Unlike heparin-induced stimulation of antithrombin-thrombin interaction, the heparin-induced stimulation of tissue plasminogen activator did not seem to follow a template model. Only in the presence of a high plasminogen or a low tissue plasminogen activator concentration, massive stimulation of tissue plasminogen activator activity was observed via a pseudotemplate model. The results suggest that precautions concerning high heparin dose should be given during its conjunctive clinical use with tissue plasminogen activator in thrombolytic therapy to reduce the risk of hemorrhage. PMID- 10709912 TI - The prothrombin nt20210 A allele as a risk factor for venous thromboembolism: detection of heterozygous and homozygous carriers by alternative methods. AB - The A allele of a G/A dimorphism at position nt20210 in the 3'-untranslated region of the prothrombin gene has been recently identified as a common risk factor for venous thrombosis. It is detected mostly by PCR amplification using a mutagenic primer and subsequent restriction analysis. We have applied two alternative methods--DGGE and SSCP-to analyse the nt20210 dimorphism in a series of 383 consecutive patients with deep venous thrombosis and in 144 controls. Both screening methods resulted in the unambiguous identification of carriers of the nt20210 A allele not only in heterozygotes but in two cases as well in homozygous state. The frequency of the A allele was 0.7% in the control group but 3.7% in the patient group, demonstrating a substantial increase in risk for venous thrombosis in carriers of the 20210 A allele. Both methods have the potential to detect other DNA sequence variations in the vicinity of the 20210 G/A dimorphism. This was demonstrated by the identification of a new mutation T20122G in the 3' untranslated region of the prothrombin gene in a single patient. PMID- 10709913 TI - 40 mg of aspirin are not sufficient to inhibit platelet function under conditions of limited compliance. PMID- 10709914 TI - In situ von Willebrand factor staining in human arteries and veins. PMID- 10709915 TI - A latent inhibitor of fibrin polymerization with ancillary anticoagulant activity. PMID- 10709916 TI - Effects of preshipping vs. arrival medication with tilmicosin phosphate and feeding chlortetracycline on health and performance of newly received beef cattle. AB - Our objective was to determine the effects of preshipping (PRE) vs. arrival (ARR) medication with tilmicosin phosphate (MIC; Exp. 1 and 2) and feeding chlortetracycline (CTC; 22 mg/kg of BW from d 5 to 9; Exp. 2) on health and performance of beef calves received in the feedlot. Ninety-six steers (Exp. 1; pay weight 236 kg) and 240 (Exp. 2; average pay weight 188 kg) steer and bull calves were used. For Exp. 1, treatments included no MIC (CON), PRE, and ARR. For Exp. 2, treatments were arranged in a 3x2 factorial. Treatments included CON, PRE, and ARR, either with CTC or without CTC. For Exp. 2, serum concentrations of immunoglobulin (Ig)G and alpha-1-acid glycoprotein (AGP) were determined on samples collected on d 0, 5, 10, and 28 and d 0, 5, and 10, respectively. No MIC x CTC interactions were observed. No differences were noted among MIC or CTC treatments in any of the experiments for ADG, daily DMI, or gain:feed ratio for the overall receiving periods. For Exp. 1, percentage of steers treated for bovine respiratory disease (BRD) was decreased (P<.05) for MIC-treated animals vs CON (71.9, 45.2, and 46.9 for CON, PRE, and ARR, respectively), and the week that calves were treated for BRD differed (P<.10) among treatments. For Exp. 2, the number of calves treated for BRD was decreased (P<.01) for MIC-treated steers vs CON and decreased (P<.05) for ARR vs. PRE (40.0, 18.7, and 7.5% for CON, PRE, and ARR, respectively). Averaged across days, serum IgG was decreased (P<.05) for MIC treated steers vs. CON, with no differences noted among treatments for AGP. Results suggest that preshipping medication programs are no more effective than arrival medication programs using tilmicosin phosphate. PMID- 10709917 TI - Comparison of models for genetic evaluation of scrotal circumference in crossbred bulls. AB - The aim of this study was to evaluate the effect of including concomitant body weight and(or) a random dam effect in genetic evaluation models on variance component estimates and standard error of prediction for scrotal circumference (SC) at 6, 8, 10, and 12 mo. Variance components and average standard errors of prediction were compared under models differing in either the number of related traits (M11 [SC], M12 [SC and BW]) or an uncorrelated random dam effect (M21 [SC], M22 [SC and BW]) using records on 1,547 bull calves. In a single-trait model (M11), estimates of direct heritabilities (h2a) for SC were .45, .49, .57, and .66 at 6, 8, 10, and 12 mo, respectively. In a two-trait model (M12), h2a were similar to those in M11 model. In M21, h2a for SC were .37, .42, .54, and .65, whereas the proportions of phenotypic variance due to dams (d2) were .12, .11, .04, and .02 at 6, 8, 10, and 12 mo, respectively. Similarly, in M22, h2a for SC were .36, .44, .56, and .65 and d2 were .13, .10, .02, and .02. Standard errors of prediction for SC EBV from M22 were reduced by 2.86, 1.21, 3.02, and 1.99% relative to M21 and by 6.45, 2.70, 2.72, and 1.21% relative to M11 at 6, 8, 10, and 12 mo, respectively. Standard errors of prediction for SC EBV from M12 were reduced by .06, .73, 1.56, and .87% relative to M11 at 6, 8, 10, and 12 mo, respectively. The importance of the dam effect decreased with age for both SC and BW. These results demonstrate that a two-trait (SC and BW) animal model would result in more accurate evaluations of yearling SC EBV in beef cattle than a single-trait model. PMID- 10709918 TI - Evaluation of Egyptian sheep production systems: I. Breed crosses and management systems. AB - Our objective was to evaluate life-cycle performance of flocks of two Egyptian breeds, Rahmani (R) and Ossimi (O), and their crosses with Finnish Landrace (F) in two management systems. Management systems were one mating season per year (1M) and three mating seasons per 2 yr (3M). Breeds and crosses studied included purebred R and O, F1 crosses 1/2F-1/2R (FR) and 1/2F-1/2O (FO), and inter se matings of 1/4 F-3/4 R (RFR) and 1/4 F-3/4 O (OFO). A dynamic computer model was used to simulate animal performance and enterprise efficiency and profit. Two measures of lifecycle feed conversion (biological efficiency) were computed: kilograms of TDN input per kilograms of empty body weight output (TDN/EBW) and kilograms of TDN input per kilogram of carcass lean output (TDN/CLN). Profit was measured as gross margin (income minus variable costs per ewe per year, GM/EWE). Input parameters for the model were obtained from published results and analyses of data collected from experimental flocks of the same genetic stocks in Egypt. Profit for FR and RFR was 42 and 6% higher in 1M than in 3M. However, profit for all other genetic types was 4 to 8% greater in 3M than in 1M. Breed rankings changed depending on the measure of evaluation (i.e., biological efficiency or profit). Maximization of system output did not necessarily improve efficiency. Under accelerated lambing systems, greater overhead costs associated with labor and feed offset gains in ewe productivity. Genetic stocks should be matched to resources and management systems. PMID- 10709919 TI - Evaluation of Egyptian sheep production systems: II. Breeding objectives for purebred and composite breeds. AB - Objectives for this study were to estimate relative economic weights for performance traits for two native and two composite sheep breeds under two management systems in Egypt. Breeds studied were Rahmani (R), Ossimi (O), 3/4R 1/4Finnish Landrace (RFR), and 3/4O-1/4Finn (OFO); OFO and RFR were composite breeds. Management systems were one mating season per year (1M) and three mating seasons per 2 yr (3M). A dynamic computer model was used to simulate animal performance and enterprise efficiency and profit. Input parameters for the model were obtained from published results and analyses of data collected from experimental flocks of the same genetic stocks in Egypt. Responses for two measures of life-cycle feed conversion and one measure of enterprise profit were evaluated. Life-cycle feed conversion was calculated as kilograms of TDN input per kilogram of empty body weight output (TDN/EBW) and kilograms of TDN input per kilogram of carcass lean output (TDN/CLN). Profit was measured as annual gross margin/ewe (GM/EWE). Traits evaluated were conception rate (CR), lambing rate (LR), mortality rate (MR), mature weight (MW), and milk production (MK). Based on responses to percentage changes in trait means, CR was most important for TDN/EBW, followed by LR and MR. For TDN/CLN, LR, MR, and CR were most important. For GM/EWE, CR was most important, followed by LR, MW, and MR. In the systems studied, there was little response to changes in MK. Based on changes in GM/EWE per genetic standard deviation change, LR was most important, followed by CR, MR, MW, and MK in all systems. Relative economic weights for O and OFO were similar, as were weights for R and RFR. Differences in economic weights between management systems for the same breed were not large enough to justify separate selection lines within breeds. PMID- 10709920 TI - Social effects and boar taint: significance for production of slaughter boars (Sus scrofa). AB - A study was conducted to elucidate the effects of social factors on the concentrations of boar taint substances, androstenone and skatole, in boars. The factors included dominance (social rank) and the effects of strongly tainted animals on other members of the group. Four successive replicates of 100 pigs (50 boars + 50 gilts) with an average live weight of 24 kg were randomly allocated to 10 pens of 10. Data for this study were collected during the period of 67 to 114 kg of live weight and included the repetitive recording of agonistic behavior during competitive feeding; blood sampling for determination of plasma androstenone, skatole and testosterone in boars; feces sampling for determination of skatole content; and collection of bulbourethral glands in boars, and uteri plus ovaries in gilts at slaughter, for the assessment of sexual maturity. Results show an influence of social rank on plasma concentrations of androstenone (P = .0001) and testosterone (P = .0001), the weight of the bulbourethral glands (P = .0001), and plasma skatole (P = .02). Pens were classified according to the pig with the highest concentration of androstenone in the pen into high, medium, and low maximum pens. In pens with high maximum concentrations of androstenone, the second-highest androstenone concentration (P = .0001), and the average concentration (P = .0003) in the pen were higher than those in pens with medium or low maximum concentrations of androstenone. Mean aggression level was also higher (P = .02), but pens with high maximum aggression level did not have higher mean androstenone concentration. Rank effect on androstenone was more important than aggression effect. Neither maximum androstenone concentration nor maximum aggression level in a pen was related to the pen mean stage of sexual maturity in either sex. No influences of rank, aggression, or aggression received were found on the feces skatole level, and no pheromonal communicative function was demonstrated for skatole. High androstenone concentrations did not have a suppressive effect on androstenone concentrations in other males of the group; on the contrary, the levels were increased. This may be due to a stimulating effect of androstenone and, possibly, mating activity. Consequently, in the production of boars for slaughter, strongly tainted animals should be avoided or removed and mating activity minimized. This could be facilitated by, for instance, slaughtering before sexual maturity or separate rearing of the sexes. PMID- 10709921 TI - Expression of two variants of growth hormone receptor messenger ribonucleic acid in porcine liver. AB - Transcription of GH receptor (GHR) mRNA is initiated from multiple promoters. Most GHR mRNA arise from GHR Promoter 1 (GHR P1) and GHR P2, which transcribe GHR 1A and GHR 1B mRNA, respectively. Our objective was to characterize the expression of GHR 1A and GHR 1B mRNA in liver of neonatal intact (1 d of age) and castrated (14, 28, and 42 d of age) male pigs (Exp. 1; n = 6 per age group), intact male pigs treated with recombinant porcine ST (rpST) or control (Exp. 2; 21, 42, and 77 d of age; n = 4 pigs per treatment per age), and pregnant gilts treated with rpST (n = 6) or control (n = 7) (Exp. 3). Tissue samples were collected at slaughter for mRNA analyses. The porcine GHR 1A and GHR 1B cDNA were cloned and were homologous to GHR cDNA isolated from other species. Ribonuclease protection assays were used to measure GHR 1A and GHR 1B mRNA. Liver expressed GHR 1A and GHR 1B mRNA, whereas muscle, uterus, and ovary expressed GHR 1B mRNA. The GHR 1A mRNA in the liver of neonatal intact and castrated male pigs (Exp. 1) was expressed at very low levels on d 1, 14, and 28, and two of six pigs expressed a high level of GHR 1A on d 42. The GHR 1B mRNA, however, was detected at all ages (d 1 through 42), and the amount of GHR 1B increased (P<.05) on d 42. The liver of intact male pigs (Exp. 2) expressed GHR 1B mRNA by 21 d, whereas a high level of GHR 1A mRNA was not detected until d 42 (P<.10). Administration of rpST had no effect on expression of GHR 1A or GHR 1B mRNA in pigs younger than 77 d (Exp. 2), but it tended to decrease (P<.10) GHR 1A mRNA but not GHR 1B mRNA in pregnant gilts (Exp. 3). In conclusion, GHR mRNA in porcine liver was composed of at least two variants (GHR 1A and GHR 1B). The GHR 1B mRNA was the major GHR mRNA in pig liver before 77 d of age. The GHR 1A mRNA increased after 42 d of age and tended to undergo specific down-regulation in response to rpST in pregnant gilts. PMID- 10709922 TI - Feeding value of an enzymatically digested protein for early-weaned pigs. AB - Weanling pigs were used in a series of studies to determine the feeding value of an enzymatically digested protein product developed from a blend of swine and poultry abattoir by-products. The initial study used 156 pigs weaned at approximately 22 d of age to compare the product with menhaden fish meal in Phase II diets. The product supported equal growth rate, and there was no preference for diet exhibited based on inclusion level of the enzymatically digested protein product. The second study used 100 pigs weaned at approximately 21 d of age to compare the product with spray-dried animal blood cells in Phase II diets. The product supported a growth rate equal to that with the blood cells, and the combination of products enhanced growth rate (P<.05). The third study used 265 pigs to compare the product with spray-dried porcine plasma in a slope ratio growth assay. Results demonstrated a relative feeding value of 91% for the product over a 4-wk feeding period. The fourth study used 290 pigs to compare the product with spray-dried porcine plasma in Phase II diets; results demonstrated comparable growth performance. The final study used 180 pigs to compare the product with spray-dried porcine plasma in Phase I diets; results demonstrated comparable growth performance. These data indicate that the enzymatically digested abattoir by-product is a high-quality protein source for weanling pigs. PMID- 10709923 TI - A dynamic model of protein digestion in the small intestine of pigs. AB - A dynamic mathematical model of the digestion of proteins in the small intestine of pigs was developed. The model integrates current knowledge on the transit of digesta along the small intestine, endogenous secretions, digestion of proteins, and absorption of amino acids into a mechanistic representation of digestion. The main characteristics of the model are the following: the small intestine is divided into several segments of variable length but with equal digesta retention time; the rate of transfer of digesta between segments is based on the progression of myoelectric migration complexes; pancreatic and biliary secretions are poured into the first segment, whereas intestinal secretions enter all intestinal segments; protein hydrolysis is described by first-order equations; and an intestinal absorption capacity is used to estimate absorption of hydrolyzed protein. Simulation results are consistent with observed data, although more information is needed to represent reality more closely. The sensitivity analysis shows that parameters for protein hydrolysis largely determine protein digestibility. The absorption capacity of the small intestine limits the absorption of amino acids at the beginning of a meal and modulates the appearance of amino nitrogen in the portal vein. It also shows that amino acid absorption can be limiting to protein digestibility when large amounts of protein are eaten in a single daily meal. The model is useful in evaluating the dynamics of protein digestion and absorption of feedstuffs. The model can be used in evaluating protein digestion of different feedstuffs and feeding strategies. PMID- 10709924 TI - Effects of pelleting and storage of a complex nursery pig diet on lysine bioavailability. AB - The effects of pelleting and storage of a complex nursery pig diet (28% lactose and 1.4% lysine) on lysine bioavailability were assessed in a chick bioassay. The nursery diet was steam-conditioned at 60 degrees C for 45 s and then pelleted through a 5-mm die with a depth of 38 mm. Samples of meal and pelleted diet were placed in metallic feeders in an occupied nursery facility for 1 wk (warm) or were stored at 4 degrees C (cool). For the standard-curve bioassay, a total of 144 8-d-old chicks were offered the following dietary treatments: 1 to 3) a basal diet (lysine deficient) and two levels (.08 and .16%) of added lysine (from L lysine-HCl); 4 and 5) two positive controls (.7% added lysine with or without 10% of the nursery diet); and 6 to 9) basal diet plus 10% of one of the four nursery diet samples (meal or pellet stored cool or warm for 1 wk). Pelleting had no effect (P>.10) on lysine bioavailability, probably because pelleting conditions (temperature, humidity, and pellet size) were not aggressive enough to result in detectable effects on lysine utilization. However, storage in the nursery facility for 1 wk reduced (P<.03) lysine bioavailability by an average of 10%. No significant (P>.10) interactions were observed. Furthermore, true digestibility of lysine in the four pig diet samples was estimated in a cecectomized cockerel digestibility assay using 15 adult Single-Comb White Leghorn cockerels. Lysine digestibility in all samples was high (average of 94%) and was not affected (P>.10) by treatment. We conclude that the pelleting conditions used in our experiments did not decrease lysine utilization. More research is needed to define thermal processing conditions that might cause protein quality deterioration. However, typical warm and humid environmental conditions encountered in modern nursery facilities have a negative effect on protein quality of diets rich in reducing sugars and lysine. PMID- 10709925 TI - Lactational and subsequent reproductive responses of lactating sows to dietary lysine (protein) concentration. AB - Gilts (n = 208) were used to evaluate the effect of lysine (protein) intake over three parities on lactation and subsequent reproductive performance. Sows were assigned randomly to one of five experimental diets at each farrowing. The five corn-soybean mealbased lactation diets contained increasing concentrations of total lysine (.60, .85, 1.10, 1.35, and 1.60%) and CP (14.67, 18.15, 21.60, 25.26, and 28.82%). Other amino acids were provided at a minimum of 105% of the NRC (1988) ratio to the lysine requirement. Sows had ad libitum access to their assigned diets from parturition until weaning (19.5+/-.2 d postpartum). All sows were fed a common gestation diet (14% CP and .68% lysine) from weaning to next farrowing. Litter size was standardized by d 3 postpartum to 10 pigs in parity 1 and 11 pigs in parity 2 and 3. Increasing dietary lysine (protein) linearly decreased (P<.05) voluntary feed intake of parity 1 (from 5.4 to 4.6 kg/d), 2 (from 6.5 to 5.8 kg/d), and 3 sows (from 6.8 to 6.2 kg/d). With the increase of dietary lysine (protein) concentration during lactation, litter weight gain responded quadratically (P<.05) in all three parities. Maximal litter ADG was 2.06, 2.36, and 2.49 kg/d in parities 1, 2, and 3, respectively, which occurred at about 44, 55, and 56 g/d of lysine intake for parity 1, 2, and 3 sows, respectively. Increasing dietary lysine (protein) had no effect (P>.1) on sow weight change, weaning-to-estrus interval, and farrowing rate in all three parities and no effect on backfat change in parity 2 and 3, but tended to increase backfat loss linearly (P<.1) in parity 1. A linear decrease of second litter size (total born, from 11.7 to 10.1, P<.1; born alive, from 11.0 to 8.9, P<.01) was observed when dietary lysine (protein) increased during the first lactation. Lysine (protein) intake during the second lactation had a quadratic effect on third litter size (P<.05; total born: 13.3, 11.2, 11.6, 11.9, and 13.6; born alive: 11.8, 10.1, 10.3, 11.2, and 12.4). However, fourth litter size was not influenced by lysine (protein) intake during the third lactation. These results suggest that the lysine (protein) requirement for subsequent reproduction is not higher than that for milk production. Parity influences the lysine (protein) requirement for lactating sows and the response of subsequent litter size to previous lactation lysine (protein) intake. PMID- 10709926 TI - Influence of consumption of endophyte-infested tall fescue hay on performance of heifers and lambs. AB - Two experiments were conducted to evaluate performance and physiological responses of heifers and lambs to Neotyphodium coenophialum-infested tall fescue hay fed under European rearing conditions. Endophyte-free (E-) or 100% endophyte infested (E+) hay was derived from the same cultivar (cv. Clarine) so that the effect of the endophytic fungus could be clearly separated from a possible cultivar effect. In Exp. 1, starting in June 1996, 20 age- and body weight-paired Holstein dairy heifers were assigned for 97 d to one of two treatments consisting of ad libitum access to either E- or E+ hay, corresponding to 0 and .41 mg/kg ergovaline, respectively. During the experimental period, no significant difference (P>.20) in forage consumption, rectal temperature, or behavioral status of the animals was observed between the two treatments. The E+ diet induced a 10% apparent decrease in ADG and a clear reduction in prolactin (PRL) plasma concentration compared to the E- diet. When animals were all reassigned to a common endophyte-free diet, the E+ group recovered body weight and PRL to levels similar to those in animals fed E- after 7 wk. In Exp. 2, 30 Texel ram lambs were assigned to two treatments consisting of dietary E- or E+ tall fescue hay. The E- and E+ hays were harvested from the same plots as used in Exp. 1 and contained 0 and .96 mg/kg ergovaline, respectively. No effect of the endophyte was found on intake or carcass or testicle weight (P>.20) after the 95-d feeding period. The E+ treatment resulted in a slight reduction in BW at slaughter, mainly explained by a lower ruminal fill (P<.01). In E+ treated animals, prolactin concentrations dropped significantly (P<.001) from d 27. Hay assessment in both experiments showed no difference in chemical composition and IVDMD. The endophytic fungus strongly lowered the palatability of the E+ hay, although there was no effect on intake with heifers (Exp. 1) or with lambs (Exp. 2). The potential of severe heat stress, as expressed by the temperature humidity index, was not high in our experimental conditions, although they were considered rather unusually stressful for the western part of northern Europe. Yet, no economic effect on cattle was observed, in disagreement with results obtained in many previous U.S. studies. PMID- 10709927 TI - Serum concentrations of luteinizing hormone, growth hormone, testosterone, estradiol, and leptin in boars treated with n-methyl-D,L-aspartate. AB - Three experiments were conducted to determine the effects of n-methyl-D,L aspartate (NMA), an agonist of the excitatory amino acid glutamate, on secretion of hormones in boars. In Exp. 1, boars (185.0+/-.3 d of age; mean +/- SE) received i.v. injections of either 0, 1.25, 2.5, 5, or 10 mg of NMA/kg BW. There were no effects of NMA (P>.1) on secretion of LH and testosterone. Treatment with NMA, however, increased (P<.01) circulating GH concentrations in a dose-dependent manner. In Exp. 2, boars (401 d of age) received an i.v. challenge of NMA at a dose of 10 mg/kg BW or .9% saline. Treatment with NMA, but not saline (P>.1), increased serum concentrations of LH (P<.01), GH (P <.01), and testosterone (P<.06). In Exp. 3, boars that were 152, 221, or 336 d of age were treated i.v. with NMA (10 mg/kg BW). Across ages, treatment with NMA increased circulating concentrations of LH (P<.07) and testosterone (P<.01). However, NMA increased (P<.01) mean GH concentrations in only the oldest boars. Treatment with NMA had no effect (P>.1) on circulating concentrations of estradiol or leptin; however, estradiol concentrations increased (P<.03) with age. In summary, NMA increased secretion of LH, GH, and testosterone in boars. However, endocrine responses to treatment with NMA may be influenced by age of the animal. Finally, NMA did not influence circulating concentrations of estradiol or leptin. PMID- 10709928 TI - Investigation of the primary cause of hypoadrenocorticism in South African angora goats (Capra aegagrus): a comparison with Boer goats (Capra hircus) and Merino sheep (Ovis aries). AB - Our objective was to identify the primary site of the reduced adrenal function in South African Angora goats (Capra aegagrus) that causes a decrease in cortisol production and leads to severe losses of Angora goats during cold spells. Angora goats, Boer goats (Capra hircus), and Merino sheep (Ovis aries) were assigned to three intravenous treatments: 1) insulin, 2) corticotropin-releasing factor (CRF), and 3) ACTH. Blood cortisol concentrations were determined over a 90-min period to determine any differences in the response of the experimental animals to these treatments. For both the insulin and ACTH treatments, cortisol concentrations were less in Angora goats than in the other experimental animals. The adrenal gland was subsequently investigated as a possible cause for the observed hypoadrenocorticism. Primary adrenal cell cultures were prepared from these species, subjected to different treatments, and the cortisol production determined. Upon pregnenolone (PREG) addition, all the experimental animals' cortisol production increased significantly, with the production in Boer goats higher (P<.01) when compared with that in the other species. The stimulation of cortisol biosynthesis by ACTH was only obtained for Boer goats and Merino sheep. The stimulation of cortisol production by forskolin and cholera toxin were compared with ACTH, and, for Angora goats, only cholera toxin caused a significant increase in cortisol production. For Boer goats, no difference (P>.05) between the PREG, ACTH, forskolin, or cholera toxin treatments were observed. The Merino adrenal cells were increasingly stimulated in the following order: PREG, ACTH, forskolin, and cholera toxin (forskolin and cholera toxin stimulated cortisol production to the same extent). This investigation of the hypothalamic-pituitary-adrenocortical axis, therefore, identified the adrenal gland as the primary site of the Angora's hypoadrenocorticism. PMID- 10709929 TI - Subclinical infection with the nematode Trichostrongylus colubriformis increases gastrointestinal tract leucine metabolism and reduces availability of leucine for other tissues. AB - Gastrointestinal (GI) tract leucine metabolism was measured in 6- to 9-mo-old lambs subjected to trickle infection with Trichostrongylus colubriformis larvae and in separate animals that were not infected. Animals prepared with a jejunal catheter and with indwelling catheters into the aorta and the portal- (PDV) and mesenteric- (MDV) drained viscera were infused simultaneously with [1-13C] and [5,5,5-2H3] leucine to determine GI tract sequestration of leucine from arterial and luminal amino acid pools by tracer and tracee arteriovenous concentration differences. Leucine oxidative losses and net fluxes were also determined across the GI tract. Infection had no detectable effect on whole-body leucine flux, but it increased total GI tract leucine sequestration by 24% (P<.05) and GI tract oxidative losses of leucine by 22 to 41% (P<.01). Net PDV fluxes of leucine were decreased by 20 to 32% during the infection. The infection did not alter either the proportion of precursor leucine used by GI tract metabolism that was derived from the arterial leucine pool (.84 to .88) or the proportional sequestration of digesta-derived leucine during "first pass" absorptive metabolism (.12 to .18). These findings help to elucidate the metabolic basis for the reduced growth rates and nitrogen retention observed when animals are subjected to subclinical nematode infection. PMID- 10709930 TI - Lysine uptake by mammary gland tissue from lactating sows. AB - Kinetic properties and substrate specificity of the lysine transport system in porcine mammary gland were studied using mammary tissue explants from nine lactating sows. Sodium dependence of lysine uptake was determined by replacing sodium in the medium with choline. Kinetic parameters of lysine uptake were determined using lysine concentrations from 5 microM to 5.12 mM. Competition of lysine uptake by other amino acids was determined using the cationic amino acids, arginine and ornithine, and using other essential amino acids. Transport of lysine was time-dependent and was unaffected by replacing sodium with choline. Lysine uptake occurred by a transport mechanism with a Km of approximately 1.4 mM and a Vmax of 7.9 mmol x kg cell water(-1) x 30 min(-1). Lysine uptake was inhibited by arginine and ornithine and by high concentrations of L-alanine, L methionine, L-leucine, cycloleucine, and D-lysine, but not by 2-(methylamino) isobutyric acid. This transport mechanism is the primary system responsible for uptake of cationic amino acids in lactating sow mammary tissue. The relatively high Km, compared with physiological blood concentrations of lysine, indicates that the kinetic properties of the lysine transport system should not be limiting to milk protein synthesis. Transmembrane transport of lysine by lactating sow mammary tissue should be a direct function of plasma concentrations. However, interactions of other amino acids with the uptake system may affect lysine uptake. PMID- 10709931 TI - Dietary energy source at two feeding levels during lactation of primiparous sows: I. Effects on glucose, insulin, and luteinizing hormone and on follicle development, weaning-to-estrus interval, and ovulation rate. AB - Our objective was to study the effects of dietary-induced insulin enhancement during and after lactation on the reproductive performance of primiparous sows. During a 21-d lactation period, 48 sows were allotted to a 2x2 factorial experiment. Treatments were feeding level (high or low; 44 MJ or 33 MJ NE/d) and dietary energy source (fat or starch). After weaning, all sows received the same amount of feed (31 MJ NE/d from weaning to estrus and 17.5 MJ NE/d from breeding until slaughter) of the same energy source as fed during lactation. On d 7, 14, and 21 of lactation and d 22 (weaning), blood samples were taken every 12 min for 12 h and analyzed for plasma glucose, insulin, and LH. Sows were slaughtered on d 35 of the subsequent pregnancy, and ovulation rate was assessed. During lactation, postprandial plasma glucose and insulin concentrations were higher for sows fed the starch diet than for those fed the fat diet (P<.001), whereas feeding level had no effect. Basal and mean LH concentrations were not affected by treatments. The LH pulse frequency on d 7 of lactation was greater for sows fed the starch diet than for those fed the fat diet (.52 vs .17 pulses/12 h; P = .03). The high compared with the low feeding level resulted in a greater LH pulse frequency on d 21 of lactation (.89 vs .47 pulses/12 h; P = .05) and on d 22 (8.63 vs 5.77 pulses/12 h; P = .02), in a higher percentage of sows that exhibited estrus within 10 d after weaning (96 vs. 63%; P = .01), and a tendency for a higher ovulation rate (18.0 vs. 16.2; P = .09). Plasma glucose and insulin concentrations were not related to any of the LH traits. The LH pulse frequency after weaning was related to the weaning-to-estrus interval (WEI) and was best explained by a linear-plateau model. In sows fed the low feeding level, follicle size after weaning was correlated with LH pulse frequency after weaning and with the WEI, whereas in sows fed the high feeding level these correlations were not significant. Our results indicate that an improved dietary-induced insulin status during and after lactation does not overcome the inhibitory effects of lactation on subsequent reproduction at any of the feeding levels. PMID- 10709932 TI - Dietary energy source at two feeding levels during lactation of primiparous sows: II. Effects on periestrus hormone profiles and embryonal survival. AB - Our objective was to study the effects of dietary energy source (fat or starch) on periestrus hormone profiles and embryonal survival in primiparous sows. During lactation, 48 primiparous sows were fed either a starch-rich or a fat-rich diet, at either a high (44 MJ NE/d) or a low (33 MJ NE/d) feeding level. After weaning, all sows received the same amount of feed (31 MJ NE/d from weaning to estrus and 17.5 MJ NE/d from breeding to slaughter) of the same dietary energy source fed during lactation. Around estrus, blood samples were taken to analyze the preovulatory LH surge, estradiol (E2), and progesterone (P4). Sows were inseminated on each day of standing estrus. On d 35 after last insemination, all 35 pregnant sows were slaughtered and their reproductive tracts were removed. The number, weight, and length of the embryos and placentas were determined as well as the weight and length of the uterus. The LH, E2, and P4 profiles were similar for the treatment groups, except for the E2 levels at 16, 12, and 8 h before the LH surge, which were lower in the sows fed the fat-rich diet at a low level. Ovulation rate tended to be higher in sows fed the high compared to the low feeding level during lactation (18.0 vs. 16.2; P = .09), but the number of total and viable embryos as well as embryonal survival rate were not influenced by the treatments. Neither uterine length and weight nor length and weight of the embryos and placentas were affected by treatments. However, after removal of the embryo-placental units, uterine weight was greater in sows fed the high than in those fed the low feeding level during lactation (1.8 vs. 1.6 kg; P = .03). Plasma insulin concentration during lactation was not related to any of the uterine, placental, or embryo traits. Mean progesterone concentration between 24 and 250 h after the LH surge was positively correlated with embryonal survival. Differences in progesterone concentration between sows with high and low embryonal survival were evident from 172 h after the LH surge. From the present study, we conclude that altering feeding level during lactation or dietary energy source from farrowing until d 35 of subsequent pregnancy did not affect embryonic development and embryonal survival. PMID- 10709933 TI - Influence of bovine growth hormone and growth hormone-releasing factor on messenger RNA abundance of lipoprotein lipase and stearoyl-CoA desaturase in the bovine mammary gland and adipose tissue. AB - Our objective was to determine the influence of bovine growth hormone (bGH) and bovine growth hormone-releasing factor (bGRF) administration on the mRNA abundance of lipoprotein lipase (LpL) and stearoyl-CoA desaturase (SCD). Primiparous Holstein cows received bGH, bGRF, or no treatment from 118 to 181+/-1 d postpartum. We hypothesized that bGH and bGRF treatment would increase the mRNA abundance of both SCD and LpL in the mammary gland with a corresponding reduction in adipose tissue. Milk yield significantly increased but milk fat percentage did not change as a result of bGH or bGRF treatment. Short-, medium-, and long-chain fatty acid concentrations in milk were not affected by either bGH or bGRF treatments, with the exception of a modest, but significant, increase in C16:1 and C18:1 following bGH treatment. Analysis was conducted on the genes encoding LpL (E.C. 3.3.1.34), a key enzyme involved in the uptake of fatty acids into tissues, and SCD (E.C. 1.14.99.5), which is the enzyme responsible for introducing delta9 double bonds in fatty acids of 16 and 18 carbons in length. In adipose tissue, treatment with bGH and bGRF reduced the mRNA abundance of LpL to 14.6 and 25.7% respectively, of that observed for control animals. Similarly, these treatments reduced the SCD mRNA abundance to undetectable levels in adipose tissue. In mammary gland, bGH and bGRF had no significant impact on LpL mRNA abundance. Bovine GH did not significantly affect SCD mRNA abundance in the mammary gland, and bGRF reduced SCD mRNA abundance. From this study to examine the role of bGH and bGRF on the expression of the genes encoding these key lipogenic enzymes in cattle, we conclude that the increased substrate required for enhanced milk fatty acid yield may have been provided through redirection of nutrients to the mammary gland away from adipose tissue and through overall increased metabolism in the mammary gland. PMID- 10709934 TI - Assessment of hypothalamic-pituitary-adrenal axis and sympathetic nervous system activity in pregnant sows through the measurement of glucocorticoids and catecholamines in urine. AB - We validated the use of urine to monitor changes in the activity of both the hypothalamic-pituitary-adrenal (HPA) axis and the sympathetic nervous system (SNS) in swine. Ten pregnant sows were fitted with venous catheters 3 wk after mating. In the early (wk 6), middle (wk 9), and late (wk 14) stages of gestation, blood and urine were collected over 24 h to monitor diurnal changes in plasma cortisol, urinary cortisol, and urinary catecholamines (norepinephrine [NE] and epinephrine [EPI]). Dexamethasone suppression tests (DST) and ovine corticotropin releasing hormone (CRH) challenge tests were also performed at each stage of gestation. All plasma and urinary values changed markedly around the clock. Diurnal variations of urinary cortisol were comparable to those in plasma, with a late nocturnal peak and a trough occurring in the evening. During the dark period, urinary catecholamines were lower than during the light period. Norepinephrine increased sharply after lights came on and peaked after meal time. Epinephrine began to rise at the end of the dark period and peaked just before meal time. Average plasma cortisol increased with the stage of gestation, due to higher levels during daylight hours. Dexamethasone at 2000 (20 microg/kg i.v.) decreased plasma cortisol at 0830 and nocturnal cortisol excretion. The magnitude of the decrease in plasma ACTH and urinary cortisol after DST was lower in late than in early and midgestation, indicating increased feedback resistance at that stage. The CRH (1 microg/kg i.v.) increased plasma and urinary cortisol. Peak levels occurred 30 min and 2 to 3 h after the injection, respectively. Catecholamines and cortisol in urine produced during the night (2000 to 0800) and the early morning (0400 to 0800 and 0800 to 0900) were highly correlated with their 24-h excretion rate. These results indicate that it is possible to monitor changes in the HPA axis and SNS activity through urinary measurements in pigs. PMID- 10709935 TI - The effect of acute nutritional change on follicle wave turnover, gonadotropin, and steroid concentration in beef heifers. AB - The effects of acute nutritional change on endocrine and ovarian characteristics were studied in cyclic (intact; n = 20) and long-term ovariectomized (ovx; n = 18) heifers being fed 1.2 x maintenance (1.2M). On d 7 of an 8-d progesterone and estradiol treatment, intact and ovx heifers were randomly allocated to diets providing .4, 1.2, or 2.0M until emergence of the second follicular wave after ovulation in intact heifers. In intact heifers, two of eight fed .4M failed to ovulate. In the other six, growth rate and maximum diameter (1.1+/-.09 mm/d and 10.1+/-.7 mm, respectively) of the first dominant follicle (DF) postovulation were less (P<.05) than in heifers fed either 1.2 (1.6+/-.18 mm/d; 12.9+/-.44 mm) or 2.0M (1.6+/-.08 mm/d; 12.7+/-.7 mm). In intact heifers, LH pulse frequency and amplitude were not affected by diet (P>.10). In ovx heifers, the frequency of LH pulses was unaffected by diet (P>.10), but heifers fed .4M had a greater pulse amplitude (P<.05) and mean concentration of LH (P<.001) than those fed 1.2 or 2.0M. Plasma concentrations of FSH were greater (P<.05) in ovx heifers fed .4M than in those fed 1.2 or 2.0M and increased linearly with time (P<.01). The FSH concentrations in heifers fed 1.2 and 2.0M were similar (P>.10) and decreased linearly with time (P<.001). In intact heifers, concentrations of FSH preceding follicle wave emergence were greater in heifers fed .4M (P<.001), but basal concentrations were not affected (P>.10). Concentrations of progesterone and estradiol were unaffected by diet (P>.10). Significant diet x ovarian status interactions in plasma IGF-I concentrations existed. Plasma concentrations of insulin increased as the level of nutrition increased, whereas concentrations of NEFA decreased. In conclusion, growth rate and maximum diameter of the DF were decreased by acute nutritional restriction, without affecting the concentration of LH. The magnitude of the FSH increase preceding new follicle wave emergence increased following dietary restriction, but concentrations of FSH were unaffected during the other stages of DF growth. The results of this study may have important implications for the feeding strategies adapted for high-yielding dairy cows in the early postpartum period when feed intake is often physiologically restricted. PMID- 10709936 TI - The importance of seminal plasma on the fertility of subsequent artificial inseminations in swine. AB - Yorkshire x Landrace sows and gilts were used in a 3x2 factorial arrangement of treatments to determine the effect of uterine inflammation induced by either killed spermatozoa (KS) or bacterial lipopolysaccharide (LPS) on the fertility of a subsequent, optimally timed AI. Estrus was detected with a mature boar twice daily. Twelve hours after the first detection of estrus, females received intrauterine infusions of an inflammatory stimulus consisting of a 100-mL dose of extender containing 3x10(9) KS (n = 40), 20 microg of LPS (n = 40; positive control) or extender alone (n = 40; negative control). An insemination was performed 12 to 18 h later with 3x10(9) motile spermatozoa (i.e., fertile AI) suspended in either 100 mL of seminal plasma (SP; n = 60) or extender replenished with of estrogens (5 microg of estradiol-17beta, 4.5 microg of estrone sulfate, and 2 microg of estrone; n= 60). Transcutaneous ultrasound was performed at the time of fertile AI and again 24 h later to detect the presence or absence of preovulatory follicles. A fertile AI performed within 24 h before ovulation was considered optimal. Conception (CR) and farrowing rates (FR) were greater in females that received a fertile AI diluted with SP compared with extender (P<.01), and there was a significant (P<.05) treatment x fertile AI dilution medium interaction for both CR and FR. Females that received a fertile AI 12 h after infusion of extender had similar CR and FR regardless of fertile AI dilution medium. After inducing an inflammatory response with either KS or LPS, CR and FR were higher in females that received a fertile AI diluted with SP compared with fertile AI dilution with extender (P<.05). The effects of treatment and AI dilution media and their interactions were not significant for litter size in females that farrowed. These results show that the fertility of a subsequent AI can be impaired when semen is deposited into an inflamed environment created by an earlier AI, and this impairment was offset by inclusion of SP in the subsequent insemination. PMID- 10709937 TI - Undegraded intake protein supplementation: I. Effects on forage utilization and performance of periparturient beef cows fed low-quality hay. AB - Hereford x Angus cows (n = 36; initial wt = 568+/-59 kg) were used to evaluate effects of undegradable intake protein (UIP) supplementation on forage utilization and performance of beef cows fed low-quality hay. Treatments were control (unsupplemented) or one of three protein supplements. Supplements were fed at 1.3 kg DM/d and included UIP at low, medium, or high levels (53, 223, or 412 g UIP/kg supplement DM, respectively). Supplements were formulated to be isocaloric (1.77 Mcal NEm/kg) and to contain equal amounts of degradable intake protein (DIP; 211 g DIP/kg supplement DM). Intake of forage was measured daily during six 7-d collection periods, which approximated mo 7, 8, and 9 of gestation and mo 1, 2, and 3 of lactation. Prairie hay (5.8% CP) was offered daily for ad libitum consumption. Cows were weighed and condition-scored on d 7 of each period. Supplemented cows had greater (P = .01) total organic matter intake (g/kg BW) compared with control animals during gestation. Forage organic matter intake (g/kg BW) was greater (P< or =.02) for control cows than for supplemented cows during lactation. Digestion of OM and NDF was lower (P<.10) for control than for supplemented cows. Body weight of supplemented cows was greater (P = .01) than that of control cows on four of six weigh dates. Supplemental UIP did not affect (P> .10) cow body weight or condition score. Body condition scores of supplemented cows were higher (P = .02) during mo 9 of gestation and during mo 3 of lactation compared with controls. Reproductive performance was similar (P>.10) among treatment groups, and there were few differences in calf performance. These data were interpreted to suggest that supplemental protein can increase total tract OM and NDF digestion by beef cows and increase body weight. Increasing the level of UIP in the supplement had little effect on forage utilization or animal performance. PMID- 10709939 TI - Effect and absorption of histamine in sheep rumen: significance of acidotic epithelial damage. AB - The significance of ruminal histamine for the induction of epithelial damage and systemic histaminosis during the ruminal lactic acidosis syndrome was investigated using the Ussing chamber technique. Histamine did not affect the electrophysiological characteristics of ovine ruminal epithelia under shortcircuit conditions. In contrast, mucosal acidification to pH 5.1 induced pronounced effects on tissue conductance (Gt) and short-circuit current (Isc). Using [3H]histamine for flux determination (hist-rad fluxes), significant net absorption of hist-rad (.40+/-.07 nmol x cm(-2) x h(-1); n = 6) was evident under short-circuit conditions in the presence of a mucosal-to-serosal (ms) histamine gradient (80 microM:12 microM). In comparison to hist-rad, absorption of native histamine (ms histamine gradient 80 microM:0 microM) measured with HPLC under open circuit conditions was smaller (.010+/-.003 nmol x cm(-2) x h(-1); n = 10). Mucosal acidification to pH 5.1 led to an increase (P<.05) in net absorption of hist-rad (to .67+/-.06 nmol x cm(-2) x h(-1); n = 6) and a dramatic increase (P<.01) in the absorption of native histamine (to .27+/-.04 nmol x cm(-2) x h( 1); n = 10). Absorption of ruminal histamine should be considered an important cause of systemic histaminosis in acidotic ruminants. Histamine absorption is linked to ruminal epithelial damage, which is primarily induced by luminal acidity and not by histamine. PMID- 10709938 TI - Undegraded intake protein supplementation: II. Effects on plasma hormone and metabolite concentrations in periparturient beef cows fed low-quality hay during gestation and lactation. AB - Hereford x Angus cows (n = 36; initial wt 568+/-59 kg) were used to evaluate the effects of undegradable intake protein (UIP) supplementation on plasma hormone and metabolite concentrations. Treatments were control (unsupplemented) or one of three protein supplements. Supplements were fed at 1.3 kg DM/d and included UIP at low, medium, or high levels (53, 223, or 412 g UIP/kg supplement DM, respectively). Supplements were formulated to be isocaloric (1.77 Mcal NEm/kg) and to contain equal amounts of degradable intake protein (DIP; 211 g DIP/kg supplement DM). Prairie hay (5.8% CP) was offered for ad libitum consumption. Jugular blood samples were collected daily from each cow during six 7-d collection periods (corresponding to mo 7, 8, and 9 of gestation and to mo 1, 2, and 3 of lactation). Plasma glucose concentrations were similar between control and supplemented cows during mo 2 and 3 of lactation; however, the low UIP treatment group had consistently higher plasma glucose (P< or =.02) than cows fed medium or high UIP supplements during gestation and the last month of lactation. During gestation, cows fed the high UIP supplement had higher (P< or =.08) plasma glucose than cows fed the medium UIP supplement. During gestation, plasma insulin concentration was increased (P = .01) by supplementation; insulin also increased (P<.01; mo 8 and 9) as supplemental UIP increased. During lactation, plasma insulin was greater (P = .01) in supplemented than in control cows. During mo 2 and 3 of lactation, insulin was lower (P< or =.04) in cows fed low UIP supplement compared with cows fed medium or high UIP supplements. Growth hormone concentration was higher (P< or =.03) in control cows than in supplemented cows in all periods measured except mo 7 of gestation. Plasma nonesterified fatty acid concentrations were higher (P< or =.03) in control cows than in supplemented cows in all periods measured except the 1st mo of lactation. These data are interpreted to suggest that protein supplementation and level of UIP can alter plasma concentrations of hormones and metabolites in gestating and lactating beef cows consuming low-quality hay. PMID- 10709940 TI - Determination of parotid urea secretion in sheep by means of ultrasonic flow probes and a multifactorial regression analysis. AB - For determination of the dynamics of parotid urea secretion in conscious sheep, a previously standardized transit time ultrasonic flow metering system was used to measure bilateral parotid flow. Six ewes fed for ad libitum consumption were prepared under halothane anesthesia with ultrasonic probes around both parotid ducts; these ducts were also cannulated orally. After probe encapsulation (8 d), parotid flows were recorded during 24 h, and samples of saliva and blood for urea determination were obtained hourly. Jaw movements were recorded by means of a submandibular balloon to monitor feeding behavior. Urea concentration in parotid saliva was 60 to 74% of that in plasma (a positive linear correlation existed) and was poorly influenced by the parotid flow. The amount of urea secreted with parotid saliva was directly related to the salivation rate. To calculate the urea secretion in parotid saliva, a multiple linear regression model was developed from computer-calculated parotid flows over 1-min periods and plasma urea concentration. The model was accurate because the plot of calculated vs measured values was not significantly different from the line of identity. The daily parotid urea N varied from .35 to 1.02 g among ewes. The higher urea secretion rate found during rumination and eating (1.32+/-.42 and .98 +/-.33 mg/min, respectively) vs. during rest (.60+/-.39 mg/ min, P<.05) was due to higher salivation rates (5.17 +/-1.46, 3.56+/-.90, and 2.04+/-.52 mL/min, respectively, P<.05) rather than to changes in saliva urea concentrations (saliva:plasma urea ratio = .65+/-.04, .67+/-.04, and .68+/-.03, respectively). Of the daily parotid urea output, 40.8% was secreted during rest. The contribution of parotid urea N to the ruminal N pool was relatively small (1.2 to 3.7% of the N intake, which was 23.0 to 33.6 g/d). These techniques allowed direct and precise measurements of parotid urea secretion without disturbing the animal or altering the physiological regulation of salivary secretion. PMID- 10709941 TI - Intraruminal supplementation with increasing levels of exogenous polysaccharide degrading enzymes: effects on nutrient digestion in cattle fed a barley grain diet. AB - The effects of supplying increasing ruminal doses of exogenous polysaccharide degrading enzymes (EPDE) on rumen fermentation and nutrient digestion were studied using eight ruminally cannulated heifers, four of which were also duodenally cannulated, in a replicated Latin square. The heifers were fed a diet of 85.5% rolled barley grain and 14% barley silage (DM basis), and once daily they were given intraruminal doses of 0 (Control), 100, 200, or 400 g of a preparation containing polysaccharide-degrading enzymes. Enzyme treatment decreased ruminal pH (linear, P<.001) and increased ammonia N (quadratic, P<.001) concentration. The ruminally soluble fraction and effective degradability of feed DM in situ were increased (quadratic response, P<.001) by enzyme treatment. Ruminal administration of EPDE increased ruminal fluid carboxymethylcellulase and xylanase activities linearly (P<.001) and beta-glucanase activity quadratically (P<.01), decreased (quadratic response, P<.05) ruminal fluid viscosity, and did not affect (P>.05) ruminal fluid amylase activity. Elevated levels of fibrolytic activities in the rumen resulted in increased (quadratic, P<.001) carboxymethylcellulase, xylanase, and beta-glucanase (P<.01) activities in duodenal digesta. Duodenal amylase activity and reducing sugar concentration were also increased (quadratic responses, P<.001 and P<.05, respectively) by EPDE. Xylanase activity of fecal DM was increased linearly (P<.05) with increasing ruminal EPDE levels. Apparent digestibilities of DM, crude protein, and NDF were not affected by EPDE supplementation. Enzyme treatment did not affect (P>.05) urinary excretion of allantoin and uric acid, or concentrations of glucose and urea in blood. PMID- 10709942 TI - Rapid communication: microsatellite DNA markers (BFRO010 and BFRO011) for grayling. PMID- 10709943 TI - Rapid communication: two highly polymorphic dinucleotide microsatellites in rainbow trout (Oncorhynchus mykiss): OmyRGT18TUF and OmyRGT23TUF. PMID- 10709944 TI - Long-term maintenance of weight loss: current status. AB - Intervention strategies for promoting long-term weight loss are examined empirically and conceptually. Weight control research over the last 20 years has dramatically improved short-term treatment efficacy but has been less successful in improving long-term success. Interventions in preadolescent children show greater long-term efficacy than in adults. Extending treatment length and putting more emphasis on energy expenditure have modestly improved long-term weight loss in adults. Fresh ideas are needed to push the field forward. Suggested research priorities are patient retention, natural history, assessment of intake and expenditure, obesity phenotypes, adolescence at a critical period, behavioral preference-reinforcement value, physical activity and social support, better linkage of new conceptual models to behavioral treatments, and the interface between pharmacological and behavioral methods. PMID- 10709945 TI - Relapse and maintenance issues for smoking cessation. AB - This article reviews short-term (6 months) and longer term (12-24 months) maintenance of cessation and relapse in adult smokers and the factors and treatments that affect these outcomes. MedLine and PsycLIT searches were done for research published in English between 1988 and 1998 meeting a defined set of criteria. Intensive intervention, telephone counseling, and use of pharmacotherapy were found to improve outcomes; however, compared with public health approaches, they reach relatively few smokers. Brief interventions during medical visits are cost-effective and could potentially reach most smokers but are not consistently delivered. Predictors of relapse include slips, younger age, nicotine dependence, low self-efficacy, weight concerns, and previous quit attempts. Potential areas for research, recommendations for longer follow-up assessments, and standard definitions for slip, relapse, and long-term maintenance are discussed. PMID- 10709946 TI - Physical activity behavior change: issues in adoption and maintenance. AB - The many benefits of participation in regular moderate- or vigorous-intensity physical activity are well established, yet more than 60% of the population is sedentary or insufficiently active. Published studies have revealed that behavior modification and cognitive-behavior modification can be successfully used to assist patients, healthy adults, and youth in the adoption of physically active lifestyles. However, few studies with adults and youth have examined the maintenance of physical activity behavior beyond 6 months of adoption of this behavior. Maintenance of physical activity is critically important because ongoing participation in the behavior is necessary to sustain health benefits. Knowledge of effective intervention strategies for long-term maintenance of physical activity is at an early stage. The authors provide a summary of what is known about the maintenance of physical activity behavior in adults and youth and how physical activity behavior relates to other health behaviors such as smoking, as well as recommendations for research on physical activity behavior change and maintenance. PMID- 10709947 TI - Maintenance of dietary behavior change. AB - Reducing dietary fat, saturated fat, and sodium and increasing intakes of dietary fiber and fruits and vegetables are important for cardiopulmonary risk reduction. Behaviorally, these dietary changes are very challenging, and in different ways. Fewer than half of U.S. adults have diets meeting recommended intakes of these constituents, and many do not see a need to align their diets with recommendations. Various nutrition education and behavioral counseling approaches have been shown to facilitate changes in fat, fiber, sodium, and fruits and vegetables, but primarily in research settings and among the highly motivated. Practice-based and interdisciplinary studies are needed to refine strategies to effect long-term dietary changes, to differentiate behavioral issues for changes involving additions versus deletions from the diet, and to elucidate the roles of sensory, psychosocial, and contextual factors in adoption and maintenance. PMID- 10709948 TI - A learning theory perspective on lapse, relapse, and the maintenance of behavior change. AB - Behavior change processes studied in the learning laboratory, such as extinction and counterconditioning, do not involve destruction of the original learning. Instead, they often result in new behavior that is strongly dependent on the current context, whether provided by external cues, internal state, recent events, or time. Lapse and relapse effects may therefore occur after various manipulations of the context. Theory and preliminary evidence suggests that long term maintenance of changed behavior may be promoted by a number of factors, including situating the new learning in the most relevant contexts, providing retrieval cues after the new learning is complete, and varying the contexts in which the new learning takes place. Furthermore, because original learning is often more context free than the learning that replaces it, the most efficient way to reduce risk behavior in the general population may be to find ways to ensure that healthy behaviors and attitudes are learned first. PMID- 10709949 TI - Toward a theory-based analysis of behavioral maintenance. AB - Intervention strategies that can produce successful rates of long-term behavior change have proven elusive and indicate the need for new approaches to this vexing problem. However, the development of new intervention strategies is constrained by our current conceptualization of behavioral maintenance. This article reviews how the dominant models of health behavior change have operationalized the psychological processes that guide the initiation and maintenance of a new pattern of behavior. In light of this review, an alternative framework is proposed based on the premise that the decision criteria that lead people to initiate a change in their behavior are different from those that lead them to maintain that behavior. Decisions regarding behavioral initiation are predicted to depend on favorable expectations regarding future outcomes, whereas decisions regarding behavioral maintenance are predicted to depend on perceived satisfaction with received outcomes. The implications of this framework for behavioral interventions are addressed. PMID- 10709950 TI - Maintenance of behavior changes in cardiorespiratory risk reduction: a clinical perspective from the Ornish Program for reversing coronary heart disease. AB - This article is an edited version of extemporaneous remarks invited by the organizers of the National Heart, Lung, and Blood Institute-sponsored conference "Maintenance of Behavior Change in Cardiorespiratory Risk Reduction" (July 1998). The request for this author was an "outside-the-box" reaction from a clinical perspective to the working groups on the reports presented at the conference. The specific clinical perspective is one gained from 15 years of experience in the Ornish Heart Disease Reversal Program. The Ornish Program was of interest to conference organizers because of its success in reducing cardiac risk factors, altering the progression of coronary artery disease, and maintaining long-term adherence to lifestyle change (K. L. Gould et al., 1992, 1995; D. M. Ornish et al., 1979, 1983, 1990, 1998; D. Ornish & Multicenter Lifestyle Demonstration Project Study Group, 1998). PMID- 10709951 TI - Promoting the maintenance of health behavior change: recommendations for the next generation of research and practice. AB - This article highlights several broad themes that emerged from the series of papers presented at the National Heart, Lung, and Blood Institute conference, "Maintenance of Behavior Change in Cardiorespiratory Risk Reduction," with a view to generating recommendations for the next generation of research and practice. Major recommendations center around the need for (a) new models of population health behavior change and maintenance that integrate individual-level with broader environmental and macro-level policy influences; (b) a fuller model of the maintenance process, which views maintenance more as a journey than as a destination; and (c) more theory-based and interdisciplinary research on the maintenance process and on strategies for assisting special populations and addressing more than one behavioral risk at a time. PMID- 10709952 TI - Cross-cutting themes in maintenance of behavior change. AB - This article reviews the cross-cutting themes and research recommendations that emerged from the National Heart, Lung, and Blood Institute "Maintenance of Behavior Change in Cardiorespiratory Risk Reduction" conference. Throughout the conference participants emphasized that maintenance should be conceptualized as a process rather than merely as the last step in the behavior change process. Researchers should focus on understanding how those who are successful at long term maintenance complete this process and on developing new approaches that can be used to help those who are not successful. Further attention to the following topics was encouraged: observational studies of the natural history of successful long-term behavior change, development of new technologies for measuring health behaviors, better theoretical understanding of the differences between initial behavior change and maintenance, development of strategies to increase participation rates and long-term effectiveness of formal treatment programs, new approaches to increase the effectiveness of self-help interventions, and better understanding of the development of behavioral patterns in children and adolescents. PMID- 10709953 TI - Brain research bulletin special section on the proceedings of the neurochemical and peptidergic pathways of the baroreflex arc in the medulla oblongata symposium. PMID- 10709954 TI - Neurochemical and peptidergic pathways of the baroreflex arc in the medulla oblongata: an introduction. PMID- 10709955 TI - Anatomical substrates for baroreflex sympathoinhibition in the rat. AB - The fundamental neuronal substrates of the arterial baroreceptor reflex have been elucidated by combining anatomical, neurophysiological, and pharmacological approaches. A serial pathway between neurons located in the nuclei of the solitary tract (NTS), the caudal ventrolateral medulla (CVL), and the rostral ventrolateral medulla (RVL) plays a critical role in inhibition of sympathetic outflow following stimulation of baroreceptor afferents. In this paper, we summarize our studies using tract-tracing and electron microscopic immunocytochemistry to define the potential functional sites for synaptic transmission within this circuitry. The results are discussed as they relate to the literature showing: (1) baroreceptor afferents excite second-order neurons in NTS through the release of glutamate; (2) these NTS neurons in turn send excitatory projections to neurons in the CVL; (3) GABAergic CVL neurons directly inhibit RVL sympathoexcitatory neurons; and (4) activation of this NTS-->CVL- >RVL pathway leads to disfacilitation of sympathetic preganglionic neurons by promoting withdrawal of their tonic excitatory drive, which largely arises from neurons in the RVL. Baroreceptor control may also be regulated over direct reticulospinal pathways exemplified by a newly recognized sympathoinhibitory region of the medulla, the gigantocellular depressor area. This important autonomic reflex may also be influenced by parallel, multiple, and redundant networks. PMID- 10709956 TI - Neurochemical transmission of baroreceptor input in the nucleus tractus solitarius. AB - Baroreceptor activation has been found to produce different types of discharge patterns in neurons in the nucleus tractus solitarius (NTS). The contribution of different glutamate receptor subtypes, neuropeptide modulators and input from different baroreceptor subtypes to the generation of firing patterns in NTS barosensitive neurons was examined in a series of studies. Results from these studies indicate that both subtypes of ionotropic glutamate receptors contribute to discharge in barosensitive neurons, and the role of each subtype can vary for different neurons. The neuropeptide neurotensin was found to modulate baroreceptor control of BP and discharge of central barosensitive neurons, both through modulation of baroreceptor afferent input and possibly through release of neurotensin by baroreceptor afferent fibers in the NTS. Finally, selective modulation of input from baroreceptor subtypes indicates that there is some degree of divergent baroreceptor innervation of NTS neurons that could contribute to initiation of their different discharge patterns in response to baroreceptor input. PMID- 10709957 TI - Angiotensin peptides and baroreflex control of sympathetic outflow: pathways and mechanisms of the medulla oblongata. AB - The baroreceptor reflex is a relatively high gain control system that maintains arterial pressure within normal limits. To a large extent, this is accomplished through central neural pathways responsible for autonomic outflow residing in the medulla oblongata. The circulating renin-angiotensin system also contributes to the regulation of blood pressure, predominantly through its effects on the control of hydromineral balance and fluid volume. All the components of the renin angiotensin system are also found in the brain. One of the principal products of the renin-angiotensin system cascade (brain or blood), angiotensin II, modulates the baroreceptor reflex by diminishing the sensitivity of the reflex and shifting the operating point for regulation of sympathetic outflow to higher blood pressures. This paper reviews our current knowledge about the neuronal pathways in the medulla oblongata through which angiotensin peptides alter the baroreceptor reflex control of sympathetic nerve activity. Emphasis is placed on the probable components and neural mechanisms of the medullary baroreflex arc that account for the ability of angiotensin peptides to change the sensitivity of the baroreceptor reflex and to shift the baroreceptor reflex control of sympathetic outflow to higher blood pressures in a pressure-independent manner. PMID- 10709958 TI - Baroreflex dependent and independent roles of the caudal ventrolateral medulla in cardiovascular regulation. AB - The caudal ventrolateral medulla (CVLM) plays a critical role in cardiovascular regulation. Convincing data now support the hypothesis that inhibition of sympathoexcitatory neurons in the rostral ventrolateral medulla (RVLM) by CVLM neurons constitutes the necessary inhibitory link in baroreceptor reflex mediated control of sympathetic vasomotor outflow. Inhibition or destruction of the CVLM produces severe acute hypertension, consistent with blockade of baroreceptor reflexes and withdrawal of inhibition of RVLM sympathoexcitatory neurons. However, other data indicate that the CVLM also tonically inhibits RVLM sympathoexcitatory neurons in a manner not driven by baroreceptor input. In some studies, inhibition of the CVLM results in an increase in arterial pressure (AP) without inhibiting baroreceptor reflexes, possibly reflecting baroreceptor independent and baroreceptor-dependent sub-regions of the CVLM. Furthermore, in baroreceptor-denervated rats, inhibition of the CVLM still leads to large increases in AP. In addition, in spontaneously hypertensive rats (SHR) central processing of baroreceptor reflexes appears normal but CVLM-mediated inhibition of the RVLM seems to be attenuated, suggesting that it is specifically a baroreceptor-independent mechanism of cardiovascular regulation in SHR that is altered. Taken together, these findings support an important, tonic, baroreceptor independent inhibition of RVLM sympathoexcitatory neurons exerted by the CVLM. PMID- 10709960 TI - Computational modeling of the baroreflex arc: nucleus tractus solitarius. AB - In this study, we examine the utility of computational modeling in understanding nervous system function. We start by examining the reasons for, and major approaches to, computational modeling. We then chose a modeling approach and applied different variations to understanding nucleus tractus solitarius (NTS) neuronal responses to various baroreceptive stimuli. We examine the results in light of our objectives and with regard to the known parameters of the system under investigation. Our results demonstrate that modeling can be a useful tool in analysis of (and examination of underlying mechanisms for) NTS behavior on many levels. PMID- 10709959 TI - Superior laryngeal neurons directly excite cardiac vagal neurons within the nucleus ambiguus. AB - The aim of this study was to test whether superior laryngeal neurons have axon collaterals that synapse upon cardiac vagal neurons. Superior laryngeal neurons were tested as likely mediators of cardio-respiratory interaction because these neurons are active in post-inspiration, co-localized with cardiac vagal neurons, and have many axon collaterals within the nucleus ambiguus. Nontoxic fluorescent tracers were utilized to identify, in vitro, both superior laryngeal neurons that innervated the crico-thyroid muscle, and cardiac vagal neurons that projected to cardiac ganglia. Co-localization of these two populations of neurons demonstrated that cardiac vagal and superior laryngeal neurons are both co-localized in the nucleus ambiguus. Simultaneous dual patch clamp recordings were used to either inject depolarizing current and evoke an action potential (current clamp configuration) or control the voltage and depolarize an identified single superior laryngeal neuron (voltage clamp configuration) while simultaneously recording from a cardiac vagal neuron. Depolarization of some, but not all, individual superior laryngeal neurons elicited post-synaptic excitatory currents in cardiac vagal neurons, indicating that at least some superior laryngeal neurons monosynaptically synapse upon cardiac vagal neurons within the nucleus ambiguus. PMID- 10709961 TI - Intra-medial prefrontal cortex injections of scopolamine increase instrumental responses for cocaine: an intravenous self-administration study in rats. AB - The present experiments examined the effects of muscarinic cholinergic receptor blockade in the nucleus accumbens (NAC) and medial prefrontal cortex (MPC) on intravenous cocaine self-administration. Adult male Sprague-Dawley rats were implanted with chronic indwelling jugular catheters and guide cannulae stereotaxically aimed at the NAC or MPC. The rats were then given the opportunity to intravenously self-administer cocaine (0.8 mg/kg/infusion) during daily 2-h sessions. Intra-NAC microinjections of methyl-scopolamine (2, 4, 8, 16, and 32 microg/side) or vehicle did not affect either the number of lever presses made or infusions delivered. On the other hand, intra-MPC injections of scopolamine significantly increased responding, although there was only a trend for an increase in the number of cocaine infusions. The effects of intra-MPC injections of scopolamine (8 and 16 microg/side) on locomotor activity were also evaluated. Intra-MPC injections of scopolamine (16 microg/side) produced significant increases in locomotor activity. However, these same microinjections decreased locomotor activity when the animals also received cocaine (15 mg/kg, i.p.). These results suggest that cholinergic neurotransmission at muscarinic receptors in the MPC is involved in regulating cocaine-maintained responding. PMID- 10709962 TI - Electrical interaction between neurons in the pigeon isthmo-optic nucleus. AB - The present study used brain slices to investigate interneuronal communication in the isthmo-optic nucleus in pigeons. Electrical stimulation of the isthmo-optic tract generated a transmembrane potential in isthmo-optic cells that was obtained by subtracting the extracellular potential from the intracellular potential. This transmembrane potential resulted in enhancement of excitability and/or in production of spikes in 42 (63%) cells. In most cases, proximal axons marked in brain slices by Lucifer yellow were too short to reach the stimulation site, indicating that spikes were evoked by electrical field effect or ephaptic interaction produced by nearby cells whose axons were activated by stimulation. Eleven (16%) cells discharged a spikelet, or spike that was abolished by hyperpolarizing current injection leaving a spikelet. Markings of five of these cells all indicated the presence of dye-couplings, each of which consisted of a pair of cells. Fourteen (21%) cells only produced antidromic spikes with a short and constant latency. Four of these cells were marked and their axons passed through the stimulation site, implying that their nearby cells' axons might be cut too short to be electrically stimulated or they were in a sparse-cell area. The present results provide electrophysiological and neuroanatomical evidence that both electrical field effect and electrical coupling may play important roles in interneuronal communication within the pigeon isthmo-optic nucleus. These findings are supported by anatomical arrangement of densely packed cells and their oriented dendrites in this centrifugal nucleus. PMID- 10709963 TI - Feature detection of visual neurons in the nucleus of the basal optic root in pigeons. AB - Previous studies have shown that the nucleus of the basal optic root in birds is involved in optokinetic nystagmus, and its neurons respond not only to large field stimuli but also to a single object moving through their excitatory receptive fields. The present study provides electrophysiological evidence that basal optic neurons in pigeons respond vigorously to motion of a black leading edge. The orientation of the edge is also an essential factor affecting visual responses of these cells, showing that any deviation of the edge from the direction perpendicular to the preferred direction decreases visual responses in most cases. Furthermore, visual responses increase as the edge is lengthened within the excitatory receptive field. However, a square, semicircle and isosceles with an area ratio of 1.00: 0.39: 0.50 but with an identical leading edge elicit almost the same responses, which are not dependent on the shape and area of visual stimuli. It suggests that these feature extraction properties, similar to those of neurons in the nucleus lentiformis mesencephali, may be specialized for detecting optokinetic stimuli rich in luminance contrasts, but not for realizing pattern recognition. PMID- 10709964 TI - Distribution of prolactin-releasing peptide mRNA in the rat brain. AB - In order to identify the distribution of prolactin-releasing peptide (PrRP) mRNA in the rat brain, we independently cloned cDNA of PrRP. Brains were removed from three adult males, and brains from three females each at 0200 and 1400 h on day 7 of pregnancy were obtained. By the nonradioactive in situ hybridization method, the location of PrRP mRNA was detected in very restricted brain areas. The distribution of PrRP mRNA signals was very similar in both sexes. In the hypothalamus, only the ventral part of the caudal dorsomedial nucleus had PrRP mRNA signals. Other forebrain areas did not show any positive signals. In the medulla oblongata, two discrete areas contained PrRP mRNA signals. No positive signal was found in the rostral part of the medulla oblongata extending to the anterior part of the area postrema. The caudal part of the nucleus of the solitary tract (NTS) had neurons with very strong signals of PrRP mRNA. The reticular nucleus showed a few PrRP mRNA positive neurons. The number of PrRP mRNA positive cells in the NTS was not different between experimental groups, although plasma prolactin levels in these animals were different. This anatomical information on the location of PrRP mRNA in the brain provides the framework to understand the physiological functions of PrRP in vivo. PMID- 10709965 TI - Lateral parabrachial nucleus lesions in the rat: long-and short-duration gustatory preference tests. AB - The present study reports two experiments that evaluated the influence of bilateral ibotenic acid lesions of the viscerosensory neurons in the lateral parabrachial nucleus (LPBN) on intake of four prototypical taste stimuli (sucrose, sodium chloride, citric acid, and quinine hydrochloride). In the 24-h, two-bottle tests of Experiment 1, rats with lesions of the LPBN were severely impaired in their concentration-dependent consumption of sucrose, displayed a mild disturbance of sodium chloride intake, and drank normal amounts of citric acid and quinine hydrochloride. These lesion-induced deficits were less pronounced when assessed with the 15-min, 1-bottle tests of Experiment 2. The results suggest that destruction of the viscerosensory neurons within the LPBN disrupt the processing of gastrointestinal feedback. PMID- 10709966 TI - Neither acute nor chronic exposure to a naturalistic (predator) stressor influences the interleukin-1beta system, tumor necrosis factor-alpha, transforming growth factor-beta1, and neuropeptide mRNAs in specific brain regions. AB - Physical (neurogenic) stressors may influence immune functioning and interleukin 1beta (IL-1beta) mRNA levels within several brain regions. The present study assessed the effects of an acute or repeated naturalistic, psychogenic stressor (predator exposure) on brain cytokine and neuropeptide mRNAs. Acute predator (ferret) exposure induced stress-like behavioral effects, including elicitation of a startle response and reduced exploratory behaviors; these responses diminished after 30 sessions. Moreover, acute and repeated predator exposure, like acute restraint stress, increased plasma corticosterone levels measured 5 min later, but not 2 h after stressor exposure. In contrast, none of the stressors used influenced IL-1beta, IL-1 receptor antagonist, IL-1 receptor type I, IL-1 receptor accessory proteins I and II, or tumor necrosis factor-alpha mRNA levels in the prefrontal cortex, amygdala, hippocampus, or hypothalamus. Likewise, there were no stressor effects on transforming growth factor-beta1, neuropeptide Y, glycoprotein 130, or leptin receptor mRNAs in brain regions. Thus, the naturalistic/psychogenic stressor used does not affect any of the brain cytokine component mRNAs studied. It is suggested that this type of stressor activates homeostatic mechanisms (e.g., glucocorticoid release), which act to preclude brain cytokine alterations that would otherwise favor neuroinflammatory/neuroimmunological responses and the consequent increase of brain sensitivity to neurotoxic and neurodegenerative processes. PMID- 10709967 TI - Influence of melatonin on free radical-induced changes in rat pancreatic beta cells in vitro. AB - Free radicals may produce cytotoxicity to pancreatic islets under pathophysiological conditions. The aim of our in vitro investigations was to compare functional and morphological changes in pancreatic beta-cells induced by reactive oxygen species (ROS) generated by alloxan or xanthine oxidase/hypoxanthine (XO/HX), respectively. We demonstrate that short-term exposure to alloxan or to XO/HX leads to a temporarily elevated insulin release from isolated pancreatic islets. On application of alloxan, this effect is caused by beta-cell necrosis and can be prevented by administration of melatonin, while in contrast, XO/HX did not lead to long-term morphological changes in the majority of the cells. Among the cells destroyed by alloxan, only necrosis could be detected, while in contrast, some apoptotic cells were identified by the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) reaction and electron microscopic examinations of cells treated with XO/HX. Melatonin was able to prevent the changes caused by alloxan, but failed to influence the alterations caused by XO/HX. Using electron spin resonance and lipid peroxidation assay, respectively, it was confirmed that melatonin effectively detoxifies hydroxyl radicals. Therefore, we believe that hydroxyl radicals are the toxic principle of alloxan, but not of XO/HX toxicity. PMID- 10709968 TI - Ototoxicity caused by cisplatin is ameliorated by melatonin and other antioxidants. AB - The mechanism of the ototoxicity caused by cisplatin is based in the generation of reactive oxygen species, which interferes with the antioxidant protection of the organ of Corti. Conversely, the protection of the cochlea with antioxidants ameliorates the ototoxicity by cisplatin. The ototoxicity produced by cisplatin can be reversible or persistent, depending on the age of the patient, cumulative doses, number of chemotherapy cycles, history of noise exposure, and deteriorating renal function. We have obtained in rats an ototoxic chart utilizing cisplatin (10 mg/kg body weight injected intraperitoneally, once only). Together with this treatment, the animals were treated with melatonin in the drinking water (10 mg/L) or injected subcutaneously (250 microg), and with an antioxidant mixture, injected subcutaneously, composed of 0.25 mg alpha tocopherol acid succinate, 3 mg ascorbic acid, 1 mg glutathione, and 60 mg N acetylcysteine. The distortion product otoacoustic emissions were determined for a prolonged period of time for each animal. The ototoxicity produced by cisplatin was maximal from days 7 to 10 post-treatment, returning to normal values in a month. When melatonin and the antioxidant mixture were present, the recovery was between days 10 and 15 post-treatment, independent of the means of administration of the pineal product. We conclude that the ototoxicity caused by cisplatin is ameliorated by melatonin and other antioxidants. PMID- 10709969 TI - Neuroprotection by melatonin from glutamate-induced excitotoxicity during development of the cerebellum in the chick embryo. AB - This work investigated the ability of melatonin to prevent cell damage in the cerebellar cortex of chick embryo caused by glutamate administration. Cell injury was evaluated estimating, at ultrastructural level, the phenomenon of cell death and the synaptogenesis of the Purkinje cells and the cerebellar glomerular synaptic complex. Administration of glutamate during cerebellar development of the chick provokes excitotoxic neuronal degeneration characterized by a phenomenon of neuronal cell death that exhibits essentially the features of a death pattern described as necrosis and the deletion of synaptogenic processes. Our results show that melatonin has a neuroprotective effect against glutamate induced excitotoxicity. This effect is morphologically revealed by the lack of neural cell death in the embryos treated with melatonin prior to glutamate injection and also by the degree of a synaptogenesis similar to that exhibited by the control group. Likewise, we corroborate the absence of teratological effects of melatonin on chick cerebellar development. Although the possible mechanisms involved in the neuroprotective effect of melatonin are discussed, i.e., direct antioxidant effects, up-regulating endogenous antioxidant defenses, and inhibiting nitric oxide formation activated by glutamate, further studies are required to establish the actual mechanism involved in the neuroprotective effect of melatonin. PMID- 10709970 TI - Melatonin increases activities of glutathione peroxidase and superoxide dismutase in fetal rat brain. AB - Melatonin is a powerful scavenger of oxygen free radicals. In humans, melatonin is rapidly transferred from the maternal to the fetal circulation. To investigate whether or not maternal melatonin administration can protect the fetal rat brain from radical-induced damage by increasing the activities of antioxidant enzymes, we administered melatonin to pregnant rats on day 20 of gestation. Melatonin (10 mg/kg) was injected intraperitoneally at daytime (14:00 hr) and, to remove the fetuses, a laparotomy was performed at 1, 2, or 3 hr after its administration. We measured the melatonin concentration in the maternal serum and in fetal brain homogenates and determined the activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) in fetal brain homogenates. Melatonin administration markedly increased melatonin concentrations in the maternal serum and fetal brain homogenates, with peak levels achieved 1 hr after melatonin administration (serum: 538.2+/-160.7 pM/mL; brain homogenates: 13.8+/-2.8 pM/mg protein). Between 1 and 3 hr after melatonin administration, GSH-Px activity in fetal brain homogenates increased significantly (P<0.01). Similarly, SOD activity increased significantly between 1 and 2 hr after melatonin administration (P<0.01). These results indicate that melatonin administration to the mother increases antioxidant enzyme activities in the fetal brain and may thereby provide indirect protection against free radical injury. Thus, melatonin may potentially be useful in the treatment of neurodegenerative conditions that may involve excessive free radical production, such as fetal hypoxia and preeclampsia. PMID- 10709971 TI - Nocturnal 6-hydroxymelatonin sulfate excretion in female workers exposed to magnetic fields. AB - The objective of this study was to determine whether daytime occupational exposure to extremely low frequency magnetic fields (MFs) suppresses nocturnal melatonin production. Sixty female volunteers were recruited. Thirty-nine worked in a garment factory, and 21 office workers served as a reference group. Exposure assessment was based on the type of sewing machine used and MF measurements around each type of machine. Eye-level MF flux density was used to classify the operators to higher (>1 microT) and lower (0.3-1 microT) exposure categories. A third group of factory workers had diverse MF exposures from other sources. The reference group had average exposure of about 0.15 microT. Urine samples were collected on Friday and Monday for three consecutive weeks. Melatonin production was assessed as urinary 6-hydroxymelatonin sulfate (6-OHMS) excretion. The ratio of Friday morning/Monday morning 6-OHMS was used to test the hypothesis that melatonin production is suppressed after 4 days of occupational MF exposure with significant recovery during the weekend. Possible chronic suppression of melatonin production was evaluated by studying exposure-related differences in the Friday values by multivariate regression analysis. The Monday/Friday ratios were close to 1.0, suggesting that there is no increase in melatonin production over the weekend. The average 6-OHMS excretion on Friday was lower among the factory workers than in the reference group, but no monotonous dose-response was observed. Multivariate regression analysis identified MF exposure, smoking, and age as significant explanatory variables associated with decreased 6-OHMS excretion. PMID- 10709972 TI - Identification and sequencing of pineal-specific enhancing region in the mouse tryptophan hydroxylase promoter. AB - Pineal-specific expression of the tryptophan hydroxylase (TPH) gene has been demonstrated by a number of studies. However, little is known about the regulatory mechanism for pineal-specific expression of the TPH gene. To identify the cis-acting region responsible for pineal-specific expression of the TPH gene, we investigated a 6.1-kb 5'-flanking region of the mouse TPH gene using an immortalized pineal cell line (PGT-beta) derived from transgenic mice. By deletion analysis, it was demonstrated that the pineal-specific enhancing region resides approximately between -6.1 and -4.7 kb upstream from the transcription initiation site of the mouse TPH gene. Additionally, nucleotide sequence analysis of this region showed that the (AC/TG)22 repetitive sequence is located approximately -5.78 kb upstream of the mouse TPH gene, and several known tissue specific cis-acting elements, such as Pit-1 and the pituitary specific element (PSE), have also been identified in the region. We believe that the analysis of the sequence and several cis-acting elements in the pineal-specific enhancing region of the mouse TPH promoter would enhance our understanding of the precise mechanism of pineal-specific expression. PMID- 10709973 TI - Changes in nocturnal melatonin secretion in perimenopausal women: correlation with endogenous estrogen concentrations. AB - Although age-related decrease in melatonin secretion in humans and animals is well documented, there is a paucity of data on the precise changes in melatonin secretion that occur during the perimenopausal period. The present study was designed to measure changes in nocturnal melatonin and to characterize the role played by estrogen in controlling nocturnal melatonin secretion in perimenopausal women. Nocturnal serum melatonin concentrations were determined every 2 hr in 46 premenopausal women, 44 postmenopausal women, and 11 premenopausal women with uterine leiomyoma scheduled for hysterectomy and bilateral salpingo-oophorectomy. Nocturnal serum melatonin secretion in premenopausal women declined moderately from 17 to 45 years of age, and increased during the period from 46 to 50 years of age. Among postmenopausal women, a steep, age-related decline in nocturnal melatonin secretion was found for up to 15 years postmenopause, followed by an extremely gradual decline thereafter. A significant negative correlation was observed between the peak serum melatonin concentration and the serum 17 beta estradiol concentration in premenopausal women aged 40-50 years (r = -0.661, P<0.0005). Daily oral administration of conjugated estrogen (0.625 mg) to postmenopausal women suppressed nocturnal melatonin secretion (P<0.005). A low estrogen state, induced by oophorectomy of premenopausal women with uterine leiomyoma led to an increase in nocturnal melatonin secretion (P<0.0001). Our findings suggest that transient elevated nocturnal melatonin secretion during menopause may be related to the existence of a low estrogen environment. The age related decrease in melatonin secretion observed in other conditions is most likely attributable to other age-related factors. PMID- 10709975 TI - Recent developments in the characterization and biotechnological production of sweet-tasting proteins. AB - The state of the art regarding the six known sweet-tasting proteins (thaumatin, monellin, mabinlin, pentadin, brazzein and curculin) and the taste-modifying protein miraculin is reviewed. Their biochemical properties, molecular genetics and biotechnological production are assessed. All of these proteins have been isolated from plants that grow in tropical rainforests. They share no sequence homology or structural similarities. Nonetheless, one of them, thaumatin, shares extensive homology with certain non-sweet proteins found in other plants. The potential industrial applications of the sweet-tasting proteins are also discussed, placing special emphasis on the barriers that a recombinant product of these characteristics will have to overcome before it reaches the market. PMID- 10709974 TI - Therapeutic effect of melatonin on carbon tetrachloride-induced acute liver injury in rats. AB - The therapeutic effect of melatonin on acute liver injury was examined in rats intoxicated with carbon tetrachloride (CCl4). Melatonin (10, 50, or 100 mg/kg body weight [BW]) was intraperitoneally administered to male Wistar rats 6 hr after intraperitoneal injection of CCl4 (1.6 g/kg BW) at which time an apparent liver injury occurred. This post-melatonin administration dose dependently prevented the progression of liver injury at 24 hr after CCl4 injection, judging from the levels of serum transaminases, indices of liver cell damage. Rats injected with CCl4 alone showed an increase in liver lipid peroxide (LPO) content and a decrease in liver reduced glutathione content at 6 and 24 hr after the injection. The post-melatonin administration dose dependently ameliorated both changes found at 24 hr after CCl4 injection. Rats injected with CCl4 alone showed an increase in liver triglyceride (TG) content and decreases in serum TG concentration and liver tryptophan 2,3-dioxygenase (TDO) activity, a marker of the inhibition of liver protein synthesis by CCl4, at 6 and 24 hr after the injection, and also a decrease in serum albumin concentration at 24 hr. The changes in serum TG, albumin concentration, liver TG content, and TDO activity found at 24 hr after CCl4 injection were not ameliorated by the post administration of melatonin. The same administration of melatonin dose dependently reduced liver LPO content in CCl4-untreated rats. These results indicate that melatonin exerts a therapeutic effect on CCl4-induced acute liver injury in rats, possibly through its antioxidant action. PMID- 10709977 TI - Influence of complex nutrients, temperature and pH on bacteriocin production by Lactobacillus sakei CCUG 42687. AB - The effects of process conditions and growth kinetics on the production of the bacteriocin sakacin P by Lactobacillus sakei CCUG 42687 have been studied in pH controlled fermentations. The fermentations could be divided into phases based on the growth kinetics, phase one being a short period of exponential growth, and three subsequent ones being phases of with decreasing specific growth rate. Sakacin P production was maximal at 20 degrees C. At higher temperatures (25-30 degrees C) the production ceased at lower cell masses, when less glucose was consumed, resulting in much lower sakacin P concentrations. With similar media and pH, the maximum sakacin P concentration at 20 degrees C was seven times higher than that at 30 degrees C. The growth rate increased with increasing concentrations of yeast extract, and the maximum concentration and specific production rate of sakacin P increased concomitantly. Increasing tryptone concentrations also had a positive influence upon sakacin P production, though the effect was significantly lower than that of yeast extract. The maximum sakacin P concentration obtained in this study was 20.5 mg l(-1). On the basis of the growth and production kinetics, possible metabolic regulation of bacteriocin synthesis is discussed, e.g. the effects of availability of essential amino acids, other nutrients, and energy. PMID- 10709976 TI - Synthesis of alpha-ketoglutaric acid by Yarrowia lipolytica yeast grown on ethanol. AB - The ability of yeast to synthesize alpha-ketoglutaric acid (KGA) from ethanol has been studied. Thiamine-auxotrophic yeasts of different genera and species may be able to produce KGA; the main condition of synthesis is growth limitation by thiamine. Using a model culture, mutant Yarrowia lipolytica N 1, the principal conditions affecting KGA oversynthesis were identified. These were: thiamine concentration in medium and in cells, nitrogen and oxygen concentration in medium, and pH level. A KGA concentration of 49 g/l and a yield from ethanol consumed of 42% were achieved. Based on the results of the analysis of the activities of the key enzymes participating in ethanol metabolism and KGA synthesis, a concept of the mechanism of KGA biosynthesis by Y. lipolytica yeast is suggested and discussed. PMID- 10709978 TI - Production of polyhydroxyalkanoic acids by Ralstonia eutropha and Pseudomonas oleovorans from an oil remaining from biotechnological rhamnose production. AB - Screening experiments identified several bacteria which were able to use residual oil from biotechnological rhamnose production as a carbon source for growth. Ralstonia eutropha H16 and Pseudomonas oleovorans were able to use this waste material as the sole carbon source for growth and for the accumulation of polyhydroxyalkanoic acids (PHA). R. eutropha and P. oleovorans accumulated PHA amounting to 41.3% and 38.9%, respectively, of the cell dry mass, when these strains were cultivated in mineral salt medium with the oil from the rhamnose production as the sole carbon source. The accumulated PHA isolated from R. eutropha consisted of only 3-hydroxybutyric acid, whereas the PHA isolated from P. oleovorans consisted of 3-hydroxyhexanoic acid, 3-hydroxyoctanoic acid, 3 hydroxydecanoic acid, and 3-hydroxydodecanoic acid. The composition was confirmed by gas chromatography of the isolated polyesters. Batch and fed-batch cultivations in stirred-tank reactors were done. PMID- 10709979 TI - Particle size effects in bioleaching of pyrite by acidophilic thermophile Sulfolobus metallicus (BC). AB - The effect of mineral particle size on the bioleaching of pyrite by the acidophilic thermophile Sulfolobus metallicus was investigated in a batch bioreactor. Decreasing the particle size from a mean diameter of 202 micron (size fraction: 150-180 micron) to a mean diameter of 42.5 micron (size fraction: 25-45 micron) enhanced the bioleaching rate from 0.05 kg m(-3) h(-1) to 0.098 kg m(-3) h(-1). The particle size distribution of the mineral in this range did not influence the morphology and growth kinetics of the cells. The values of specific growth rate (mu) and yield factor (Y) were 0.018-0.025 h(-1) and 0.67x10(11) 1.45x10(11) cells (g iron)(-1), respectively. Decreasing the particle size of the mineral to a mean diameter of 6.40 micron (size fraction <25 micron) adversely influenced the activity of the cells. The presence of fine particles apparently damaged the structure of the cells, resulting in their inability to oxidise pyrite. PMID- 10709980 TI - Studies on nutritional and oxygen requirements for production of L-asparaginase by Enterobacter aerogenes. AB - The carbon and nitrogen sources most suitable for L-asparaginase production by Enterobacter aerogenes were selected and their concentrations optimized in shake flask cultures. Sodium citrate (1.0%) and diammonium hydrogen phosphate (0.16%) proved to be the best sources of carbon and nitrogen, respectively. Nitrogen catabolite repression of enzyme formation was absent in this bacterium. Cultivation in a reactor showed that the dissolved oxygen level is the limiting factor for L-asparaginase production by E. aerogenes. Glucose was found to be a repressor of enzyme synthesis. Asparagine was absent intracellularly when the L asparaginase level was high. An increase in the extracellular alanine level when the dissolved oxygen remained low indicated a shift from aerobic to fermentative metabolism. PMID- 10709981 TI - Novel regioselective hydroxylations of pyridine carboxylic acids at position C2 and pyrazine carboxylic acids at position C3. AB - We have previously described the isolation of the new bacterial species, Ralstonia/Burkholderia sp. strain DSM 6920, which grows with 6-methylnicotinate and regioselectively hydroxylates this substrate in the C2 position by the action of 6-methylnicotinate-2-oxidoreductase to yield 2-hydroxy-6-methylnicotinate (Tinschert et al. 1997). In the present study we show that this enzymatic activity can be used for the preparation of a series of hydroxylated heterocyclic carboxylic acid derivatives. The following products were obtained from the unhydroxylated educts by biotransformation using resting cells: 2 hydroxynicotinic acid, 2-hydroxy-6-methylnicotinic acid, 2-hydroxy-6 chloronicotinic acid, 2-hydroxy-5,6-dichloronicotinic acid, 3-hydroxypyrazine-2 carboxylic acid, 3-hydroxy-5-methylpyrazine-2-carboxylic acid and 3-hydroxy-5 chloropyrazine-2-carboxylic acid. Thus the respective educts were all regioselectively mono-hydroxylated at the carbon atom between the ring-nitrogen and the ring-carbon atom carrying the carboxyl group. In contrast to its relatively broad biotransformation abilities, the strain shows a limited heterocyclic nutritional spectrum. It could grow only with three of the seven transformed educts: 6-methylnicotinate, 2-hydroxy-6-methylnicotinate and 5 methylpyrazine-2-carboxylate. 2-Hydroxynicotinate, 2-hydroxy-6-chloronicotinate, 2-hydroxy-5,6-dichloronicotinate, 3-hydroxypyrazine-2-carboxylate and 3-hydroxy-5 chloropyrazine-2-carboxylate were not degraded by the strain. Therefore, unlike 6 methylnicotinate-2-oxidoreductase, which has a broad substrate spectrum, the second enzyme of the 6-methylnicotinate pathway seems to have a much more limited substrate range. Among 28 aromatic heterocyclic compounds tested as the sole source of carbon and energy, only pyridine-2,5-dicarboxylate was found as a further growth substrate, and this was degraded by a pathway which did not involve 6-methylnicotinate-2-oxidoreductase. To the best of our knowledge the microbial production of 2-hydroxy-6-chloronicotinic acid, 2-hydroxy-5,6 dichloronicotinic acid and 3-hydroxy-5-methylpyrazine-2-carboxylic acid have not been reported before. Strain DSM 6920 is so far the only known strain which allows the microbial production of both these compounds and 3-hydroxypyrazine-2 carboxylic acid and 3-hydroxy-5-chloroypyrazine-2-carboxylic acid. PMID- 10709982 TI - Keratinase of Doratomyces microsporus. AB - The fungus Doratomyces microsporus produced an extracellular keratinase during submerged aerobic cultivation in a medium containing a protein inducer for enzyme synthesis. The keratinase was purified to homogeneity using hydrophobic interaction chromatography followed by gel chromatography. The molecular weight was estimated to be 33 kDa (from SDS-PAGE analysis) or 30 kDa (by gel chromatography), suggesting a monomeric structure. The isoelectric point of the enzyme was determined to be around 9. The optimal pH and temperature for keratinolytic activity were pH 8-9 and 50 degrees C, respectively. The serine protease inhibitor PMSF totally inhibited the keratinase. The enzyme was not glycosylated. It was capable of hydrolysing different keratinous materials as well as some non-keratinous proteins. Hydrolysis of some synthetic substrates, specific for known proteinases, suggested that the keratinase of D. microsporus is close to proteinase K. PMID- 10709983 TI - Production characteristics of interferon-alpha using an L-arabinose promoter system in a high-cell-density culture. AB - Using high-cell-density culture of Escherichia coli under the control of an L arabinose promoter (ParaB), several factors affecting the production of recombinant protein and the formation of inclusion bodies were studied. The inducer, L-arabinose, showed a maximal induction level above 10.7 mM in the final concentration. The concentration of inducer also affected the partition of interferon-alpha (IFN-alpha) into the soluble form and inclusion bodies. Induction kinetics of the rate of accumulation of IFN-alpha on the ParaB promoter showed a slower rate than those of other promoter systems, for example T7, lac or tac. These innate characteristics of ParaB enabled cells to grow continuously in spite of the metabolic burden induced by the expression of foreign protein. The duration time of induction could control the expression of both soluble and insoluble protein. The ratio of yeast extract to glycerol (N/C ratio) in feeding media significantly affected both the production level of recombinant protein and inclusion body formation. The reason for decreasing specific bioactivity during induction can be explained by the increased proportion of inclusion bodies in the total expressed IFN-alpha. PMID- 10709984 TI - Characterization and cloning of an (R)-specific trans-2,3-enoylacyl-CoA hydratase from Rhodospirillum rubrum and use of this enzyme for PHA production in Escherichia coli. AB - An (R)-trans-2,3-enoylacyl-CoA hydratase was purified to near-homogeneity from Rhodospirillum rubrum. Protein sequencing of enriched protein fractions allowed the construction of a degenerate oligonucleotide. The gene encoding the (R) specific hydratase activity was cloned following three rounds of colony hybridization using the oligonucleotide, and overexpression of the gene in E. coli led to the purification of the enzyme to homogeneity. The purified enzyme used crotonyl-CoA, trans-2,3-pentenoyl-CoA, and trans-2,3-hexenoyl-CoA with approximately equal specificity as substrates in the hydration reaction. However, no activity was observed using trans-2,3-octenoyl-CoA as a substrate, but this compound did partially inhibit crotonyl-CoA hydration. Based on the nucleotide sequence, the protein has a monomeric molecular weight of 15.4 kDa and is a homotetramer in its native form as determined by gel filtration chromatography and native PAGE. The hydratase was expressed together with the PHA synthase from Thiocapsa pfennigii in E. coli strain DH5alpha. Growth of these strains on oleic acid resulted in the production of the terpolyester poly(3-hydroxybutyrate-co-3 hydroxyvalerate-co-3-hydroxyhexanoate) . PMID- 10709985 TI - Influence of a synbiotic mixture consisting of Lactobacillus acidophilus 74-2 and a fructooligosaccharide preparation on the microbial ecology sustained in a simulation of the human intestinal microbial ecosystem (SHIME reactor). AB - Lactobacillus acidophilus 74-2, which is used in probiotic products, was administered, with fructo-oligosaccharide in a milk-based product, to the second vessel (duodenum/jejunum) of the SHIME reactor, an in vitro simulation of the human intestinal microbial ecology. The main focus of this study was to monitor the changes of the population density of selected bacterial species in the intestine and the changes of metabolic activities during the supplementation of L. acidophilus and fructooligosaccharide in the SHIME reactor. Interestingly, the addition of L. acidophilus 74-2 with fructooligosaccharide gave rise to an increase of bifidobacteria. Moreover, major positive changes occurred in the production of volatile fatty acids: a strong upward trend was observed especially in the case of butyric acid and propionic acid. Furthermore a noticeable increase of beta-galactosidase activity was monitored, while the activity of beta glucuronidase, generally considered undesirable, declined. PMID- 10709986 TI - Biocompatible alginate from freshly collected Laminaria pallida for implantation. AB - A simple procedure is described for the extraction and purification of alginate from the inner stipes of the kelp Laminaria pallida. Alginate yield was about 10 15% of the dry mass, with a 70:30 mannuronic/guluronic acid ratio. Analysis of the purified alginate revealed a low polyphenol content while proteins were below detection level. The purified alginate was highly viscous, with 10-15 mPa s and 281 mPa s for a 0.1% and 0.5% solution, respectively, indicating a very high molecular mass (larger than 250 kDa). Bead formation occurred in the presence of divalent cations, but also in the presence of artificial serum (FCSIII) without added divalent cations. The biocompatibility of the alginate was tested with the in vitro mice lymphocyte test as well as by implantation of Ba2+ cross-linked beads beneath the kidney capsule of BB/OK rats. There was no evidence for significant mitogenic activity or fibrotic reaction. Biocompatibility of the alginate was also demonstrated by the encapsulation of human chondrocytes into Ca2+ cross-linked alginate beads. Immobilized chondrocytes grew and remained functional (i.e. they produced collagen). PMID- 10709987 TI - Reactions of pentachlorophenol with laccase from Coriolus versicolor. AB - Laccase, purified from Coriolus versicolor, removed pentachlorophenol (PCP) from solution at pH 5, depending on initial PCP concentration and amount of laccase. With 100 units of laccase, 100% of 25 microg ml(-1) PCP and 60% of 200 microg ml( 1) PCP were removed respectively over 72 h. No free chloride was released in the reaction. In reaction with 100 microg PCP, products were primarily polymers (about 80,000 MW) with only 2-3 pg of o- and p-chloranils formed. Polymers were stable to acid hydrolysis and no release of PCP, or other low-molecular-weight products, was detected over several weeks. Laccase has a potential use in the biotreatment of aqueous effluents containing PCP, with polymerised products being removed from solution due to their high molecular weight. PMID- 10709988 TI - Role of proteases in autolysis of Penicillium chrysogenum chemostat cultures in response to nutrient depletion. AB - An industrial strain of Penicillium chrysogenum was subjected to carbon or nitrogen limitation in a chemostat and the response monitored in terms of the "classical" indicators of autolysis (biomass decline and ammonia release), culture degradation (as measured by image analysis) and by obtaining profiles for three classes of proteases implicated in autolysis. Under both sets of conditions (carbon or nitrogen limitation), once started, autolysis involved a succession of different protease activities. The first stages of the process of autolysis in starved chemostat cultures was associated with peaks in the activities of both serine and aspartyl proteases, coinciding with the mobilisation of endogenous energy reserves. Conversely, a peak in the activity of metalloproteases was associated with the later stages of autolysis, perhaps occurring in response to depletion of endogenous energy reserves; the activity of these enzymes led to gross culture degradation, disintegration of ordered mycelial structures and signalled the end of metabolic activity (respiration) within the culture. These findings indicate that strategies intended to control/regulate autolysis in large scale industrial fungal cultures might profitably be focused on regulation of the activity of key classes of proteases involved in the series of events leading to culture degradation. PMID- 10709989 TI - Role of the reactor configuration in the biological detoxification of a dump site polychlorobiphenyl-contaminated soil in lab-scale slurry phase conditions. AB - The biotreatability of a xenobiotic contaminated soil is frequently determined through a bioslurry treatment usually performed in lab-scale shaken baffled flasks. In this study, a 3-1 unconventional stirred tank reactor was developed and tested in the slurry-phase treatment of a soil heavily contaminated by polychlorobiphenyls (PCBs) derived from an Italian dump site, in the absence and in the presence of biphenyl and of the exogenous PCB aerobically dechlorinating co-culture ECO3. The data obtained were compared with those obtained on the same soil in experiments performed in parallel in 3-1 baffled shaken flask reactors. Considerably higher PCB removal and soil detoxification yields (determined through the Lepidium sativum germination test and the Collembola mortality test) were attained in the stirred tank reactors, which generally displayed a higher slurry-phase homogeneity and a higher availability of biphenyl- and chlorobenzoic acid-degrading bacteria compared to the corresponding shaken flask reactors. Moreover, enhanced soil PCB biodegradation and detoxification yields were observed when the developed reactor was supplemented with biphenyl and the exogenous ECO3 bacteria. In conclusion, the results of the soil biotreatability experiments commonly performed in bioslurry lab-scale reactors are significantly influenced by the reactor configuration; the use of the unconventional stirred tank reactor system developed in this work is recommended. PMID- 10709990 TI - Azo-dye degradation in an anaerobic-aerobic treatment system operating on simulated textile effluent. AB - Decolorisation of azo dyes during biological effluent treatment can involve both adsorption to cell biomass and degradation by azo-bond reduction during anaerobic digestion. Degradation is expected to form aromatic amines, which may be toxic and recalcitrant to anaerobic treatment but degradable aerobically. Methods for the quantitative detection of substituted aromatic amines arising from azo-dye cleavage are complex. A simple qualitative method is suggested as a way in which to investigate whether decolorisation is actually due to degradation, and whether the amines generated are successfully removed by aerobic treatment. Samples from a combined anaerobic-aerobic system used for treating a simulated textile wastewater containing the reactive azo dye Procion Red H-E7B were analysed by high-performance liquid chromatography/ultraviolet (HPLC-UV) methods. Anaerobic treatment gave significant decolorisation, and respiration-inhibition tests showed that the anaerobic effluent had an increased toxicity, suggesting azo-dye degradation. The HPLC method showed that more polar, UV-absorbing compounds had been generated. Aerobically, these compounds were removed or converted to highly polar compounds, as shown by HPLC analysis. Since the total organic nitrogen (TON) decreased aerobically as organic N-containing compounds were mineralised, aromatic amine degradation is suggested. Although only a simple qualitative HPLC method was used, colour removal, toxicity and TON removal all support its usefulness in analysing biotreatment of azo dyes. PMID- 10709991 TI - The gene encoding the mouse homologue of the human osteoclast-specific 116-kDa V ATPase subunit bears a deletion in osteosclerotic (oc/oc) mutants. AB - Osteosclerosis (oc) is an autosomal recessive lethal mutation that impairs bone resorption by osteoclasts, and induces a general increase of bone density in affected mice. Genetic mapping of the oc mutation was used as a backbone in a positional cloning approach in the pericentromeric region of mouse chromosome 19. Perfect cosegregation of the osteopetrotic phenotype with polymorphic markers enabled the construction of a sequence-ready bacterial artificial chromosome (BAC) contig of this region. Genomic sequencing of a 200-kb area revealed the presence of the mouse homologue to the human gene encoding the osteoclast specific 116-kDa subunit of the vacuolar proton pump. This gene was located recently on human 11q13, a genomic region conserved with proximal mouse chromosome 19. Sequencing of the 5' end of the gene in oc/oc mice showed a 1.6-kb deletion, including the translation start site, which impairs genuine transcription of this subunit. The inactivation of this osteoclast-specific vacuolar proton ATPase subunit could be responsible for the lack of this enzyme in the apical membranes of osteoclast cells in oc/oc mice, thereby preventing the resorption function of these cells, which leads to the osteopetrotic phenotype. PMID- 10709992 TI - Reversal of malignant phenotype in human osteosarcoma cells transduced with the alkaline phosphatase gene. AB - Alkaline phosphatases are a family of glycoproteins that are able to hydrolize various monophosphate esters at a high pH optimum. Liver/bone/kidney (L/B/K) alkaline phosphatase (ALP) is one of the four major isoenzymes that belong to this family. Apart from its role in normal bone mineralization, other functions of L/B/K ALP remain obscure, both in physiological and in neoplastic conditions, including the bone-forming tumor osteosarcoma. In this study, we transfected the U-2 OS osteosarcoma cell line, which does not show any basal expression of this enzyme, with the full-length gene of L/B/K ALP, and analyzed the in vitro and in vivo features of four transfectants showing different expression of L/B/K ALP. A reduced in vitro ability to invade Matrigel and to grow in a semi-solid medium, together with a lower tumorigenic and metastatic ability in athymic mice, was found to be associated with a high level of cell surface L/B/K ALP activity. Moreover, L/B/K ALP transfectants showed a reduced secretion of matrix metalloproteinase-9 enzyme. These findings indicate a loss of aggressiveness of osteosarcoma cells after the expression of L/B/K ALP on their surface and suggest a new role for this enzyme. PMID- 10709993 TI - Nuclear localization of the type 1 parathyroid hormone/parathyroid hormone related peptide receptor in MC3T3-E1 cells: association with serum-induced cell proliferation. AB - We have recently demonstrated that the receptor for parathyroid hormone (PTH) and PTH-related peptide (PTHrP), PTHR, can be localized to the nucleus of cells within the liver, kidney, uterus, gut, and ovary of the rat. We set out to determine the localization of the PTHR in cultured osteoblast-like cells. MC3T3 E1, ROS 17/2.8, UMR106, and SaOS-2 cells were cultured in alpha-modified eagle medium containing 15% fetal calf serum under standard conditions. Untreated cells were grown on glass coverslips to 75-95% confluence and fixed in 1% paraformaldehyde. For experiments designed to examine cells synchronized by serum starvation, cells were grown on glass coverslips, starved of serum for 46 h, and then fixed at 2-h intervals for a total of 26 h after the addition of serum to the medium. Parallel sets of cells were pulsed with [3H]thymidine to track the DNA duplication interval. The PTHR was localized by immunocytochemistry using a primary antibody raised against a portion of the N-terminal extracellular domain of the PTHR. The results presented herein indicate that the PTHR attains a nuclear localization in each cell line examined. In UMR106 cells, PTHR immunoreactivity was restricted to the nucleolus. After cell synchronization, MC3T3-E1 cells double approximately 24 h after the addition of serum. Immunocytochemistry for the PTHR in these cells showed that the receptor staining is initially diffuse for the first 6 h, then becomes more perinuclear in distribution by 12-16 h. Nuclear localization of the receptor is achieved approximately 16-20 h after the addition of serum and remains there throughout the mitotic phase. Intense staining of mitotic and postmitotic cells was observed. No change in cell proliferation kinetics was observed in MC3T3-E1 cells cultured in the presence of 25 nM PTH(1-34). These data suggest an important role for the PTHR in the nucleus of MC3T3-E1 cells at the time of DNA synthesis and mitosis. PMID- 10709994 TI - The expression of the nuclear matrix proteins NuMA, topoisomerase II-alpha, and beta in bone and osseous cell culture: regulation by parathyroid hormone. AB - Bone cells undergo changes in cell structure during phenotypic development. Parathyroid hormone (PTH) induces a change in osteoblast shape, a determinant of collagen expression. We hypothesize that alterations in bone cell shape reflect and direct gene expression as governed, in part, by nuclear organization. In this study, we determined whether the expression of nuclear matrix proteins that mediate nuclear architecture, NuMA, topoisomerase II (topo II)-alpha, and -beta, were altered during osteoblast development and response to PTH in vivo. NuMA forms an interphase nuclear scaffold in some cells, the absence of which may accommodate alterations in nuclear organization necessary for specific functions. Topo II enzymes are expressed in bone cells; the alpha-isoform is specific to proliferating cells. We used immunohistochemistry and flow cytometry to determine whether NuMA is expressed in the primary spongiosa of the rat metaphyseal femur and whether expression of NuMA, topo II-alpha, and II-beta changes during osteoblast development or with PTH treatment. NuMA and topo II-beta were expressed in marrow cells, osteoblasts, osteocytes, and chondrocytes. These proteins were not detected in osteoclasts in vivo, but were observed in cultured cells. Bone marrow cells expressed topo II-alpha. All three proteins were expressed in cultures of rat osteoblast-like UMR-106 cells. PTH treatment downregulated the number of topo II-alpha-immunopositive cells, correlated with a decrease in S-phase cells, in both bone tissue and cell culture. We conclude that, in vivo, nuclear matrix composition is altered during bone cell development and that anabolic doses of PTH attenuate the proliferative capacity of osteogenic cells, in part, by targeting topo II-alpha expression. PMID- 10709995 TI - Activin release from bone coupled to bone resorption in organ culture of neonatal mouse calvaria. AB - Activin, a member of the transforming growth factor-beta (TGF-beta) superfamily, is present in the bone matrix and assumed to be involved in the regulation of bone formation. In the present study, we investigated whether the release of activin from bone is coupled with bone resorption. Neonatal mouse calvaria were cultured in the presence of various stimulators of bone resorption (parathyroid hormone [PTH], interleukin-1beta, prostaglandin E2) for up to 72 h, and the activin activity in the medium was measured using a specific bioassay for activin. Activin activity was accumulated in proportion to the time- and dose dependent increase in calcium release from bone into the medium (bone resorption). An inhibition of PTH-dependent bone resorption by a bisphosphonate, disodium dichlormethane-1,1-bisphosphonic acid (Cl2MBP), completely blocked release of activin activity from bone into the medium. In primary culture of calvarial cells, however, neither PTH nor Cl2MBP affected activin production. These findings indicate that release of activin activity from bone tissue is strongly coupled to bone resorption. Because activin possesses osteogenic activities, activin released locally from bone might be involved in the regulation of bone formation in the physiological process of bone remodeling, as has been suggested for TGF-beta. PMID- 10709996 TI - Proteolysis of human bone collagen by cathepsin K: characterization of the cleavage sites generating by cross-linked N-telopeptide neoepitope. AB - An immunoassay for cross-linked N-telopeptides of type I collagen (NTx) in urine or serum has proven to give a sensitive index of osteoclast-mediated bone resorption. We show that recombinant human cathepsin K is highly active in releasing the NTx neoepitope in 100% yield from bone type I collagen. Cathepsins S, L, and B were also active but at 57%, 36%, and 27% of the yield of K, respectively. The matrix metalloproteinases that were tested, stromelysin, collagenase 3, or matrilysin, did not produce any immunoreactivity. Cathepsin K also acted on demineralized bone matrix, releasing NTx epitope and completely dissolving the bone particles in 24-48 h. Proteolytic cleavage of a G-L peptide bond in the alpha2(I)N-telopeptide was shown to be required for recognition by monoclonal antibody 1H11. Peptide analysis identified bonds in the N-telopeptide and helical cross-linking domains adjacent to the cross-linking residues at which cathepsin K cleaved in bone collagen. The sites were consistent with the known substrate specificity of cathepsin K, which prefers a hydrophobic residue or proline in the critical P2 position. The NTx peptides generated by cathepsin K were of low molecular weight, in the range previously found in human urine. Because cathepsin K appears to be essential for the normal resorption of mineralized bone matrix by osteoclasts, these findings help explain the specificity and responsiveness of NTx as a marker of osteoclastic bone resorption in vivo. PMID- 10709997 TI - Transforming growth factor-beta increases interleukin-6 transcripts in osteoblasts. AB - Bone remodeling is regulated by local factors and cytokines. Among them, interleukin-6 (IL-6) plays a critical role in bone resorption, and its synthesis is stimulated by osteoresorptive factors. Transforming growth factor-beta (TGF beta) is present in high amounts in the bone matrix and is a local regulator of bone formation. However, its role in bone resorption remains unclear. In this paper, we report that TGF-beta stimulates IL-6 transcripts in a time- and dose dependent manner in primary rat osteoblasts isolated from 22-day-old calvariae (Ob cells). The TGF-beta effect on IL-6 mRNA levels does not require de novo protein synthesis because cycloheximide, a protein synthesis inhibitor, does not block the induction. The mechanisms of IL-6 stimulation by TGF-beta is at least partially transcriptional because TGF-beta induces IL-6 heterogenous nuclear RNA, and, to a lesser extent, IL-6 transcription rate as determined by a nuclear run on assay. Transforming growth factor-beta upregulation of IL-6 may be critical in conditions of increased bone resorption, such as myeloma. PMID- 10709999 TI - Improvement of pagetic bone lesions with risedronate treatment: a radiologic study. AB - Risedronate is a potent pyridinyl bisphosphonate in clinical development for treatment and prevention of osteoporosis, and has been recently approved for treatment of Paget's disease in the United States. An open-label study was conducted to determine the effect of risedronate treatment on pagetic bone lesions in patients with moderate to severe Paget's disease (mean serum alkaline phosphatase levels [ALP] approximately seven times the upper limit of normal). Patients were treated with 30 mg/day oral risedronate for 84 days followed by a 112-day nontreatment period. This 196-day cycle was repeated once in patients whose ALP did not normalize or who experienced relapse, defined as a > or =25% increase in ALP from the lowest value measured. Radiographs of affected anatomical sites in 26 patients were collected at baseline, 6 months, and/or 12 months. Eleven patients received one course and 15 patients received two courses of treatment. Radiographs were examined by a skeletal radiologist who was blinded to their time sequence. Changes in pagetic lesions were categorized as "improved," "deteriorated," or "no change." Between baseline and 6 months, 16 patients improved and 3 deteriorated; at 12 months, 11 patients improved and 2 deteriorated. Most lesions remained unchanged between 6 and 12 months. Improvements were noted in all skeletal sites (tibia, femur, humerus, forearm, pelvis, spine, and skull), but were most pronounced in weight-bearing long bones. In weight-bearing bones, nine lesions had osteolytic fronts. Of these, seven improved and two remained unchanged at 6 months; at 12 months, all but one lesion (which improved) remained unchanged. This radiographic assessment demonstrates that oral risedronate, 30 mg/day in one or two 3-month courses, is highly effective for improving bone lesions in patients with Paget's disease. Risedronate treatment had no deleterious effect on osteolytic lesions in weight bearing bones; indeed, the majority of lesions with osteolytic fronts were improved after 6 months of risedronate treatment. PMID- 10709998 TI - Skeletal changes in rats bearing mammosomatotrophic pituitary tumors: a model of acromegaly with gonadal dysfunction. AB - Growth hormone (GH) exerts potent effects on bone metabolism, resulting in an increased bone formation in animals and humans. Acromegaly has been associated with increased bone turnover, whereas the net effect of the increased bone metabolism has been obscured because patients with acromegaly are often associated with hypogonadism. We investigated changes in cortical and cancellous bone in adult rats implanted mammosomatotrophic pituitary tumor cells (GH3) as a model of acromegaly with gonadal dysfunction. Acromegaly model rats were prepared by implanting GH3 cells into female Wistar-Furth rats at 17 weeks of age. At 28 weeks of age, GH3-bearing rats (GH rats) showed very high serum GH levels and a moderate increase in serum prolactin levels, resulting in low circulating estradiol levels. The GH rats showed significant increases in body weight and in length and volume of both the femur and vertebral body. Bone mineral content values of either the midfemur or the whole lumbar body were significantly greater in the GH rats compared with littermate controls, while the areal bone mineral density values of the respective bones were not different between the two groups. The parameters of mechanical strength of the femur were significantly larger in the GH rats than in controls, whereas those of the lumbar vertebral body cylinder specimen were not different between the two groups. Respective normalized mechanical parameters of the femur and the vertebral body were the same in the GH rats as in controls. In the midfemur, the GH rats showed a significant increase in the total cross-sectional area without influencing the bone marrow area, resulting in an increase in the cortical bone area and the moment of inertia compared with controls. The indices of periosteal bone formation in the midfemur were greater in the GH rats compared with controls, but the endocortical bone formation and resorption were not different between the two groups. In the vertebral body cancellous bone, the GH rats had an increase in bone turnover rate, whereas the structural parameters were not different between the two groups. These results from GH3-bearing rats demonstrate that an excess of GH increases cortical bone mass in rats accompanied with estrogen deficiency, while no large effect on vertebral body cancellous bone mass is seen. PMID- 10710000 TI - Intermittent versus continuous clodronate administration in postmenopausal women with low bone mass. AB - The aim of the study was to compare the effects on bone mass and turnover of continuous vs. intermittent clodronate administration on 120 postmenopausal women (average age 61 years) with low bone mass (femoral neck bone mineral density [BMD] of at least -1 SD or more, T-score), with another 30 women as a control group. Participants were given 1800 mg of clodronate every 6 months over 2 years using different treatment patterns: a) two continuous regimens, consisting of a daily oral dose of 400 mg or 100 mg every 10 days by intramuscular injection, the latter being considered continuous because the interval between injections is shorter than the time employed by each bone remodelling unit to complete the resorption phase of a remodelling cycle; and b) two intermittent regimens, consisting of 1800 mg every 6 months administered either as a single 18-h intravenous infusion or by separate infusions of 300 mg over 6 consecutive days. All women, including those in the control group, received calcium and vitamin D supplementation. After 2 years, continuous clodronate regimens caused an increase in BMD both at lumbar spine and proximal femur (L(1-4) BMD = 3.07% and 2.69%; femoral neck = 2.12% and 2.09%, respectively, with intramuscular and oral regimens). Intermittent clodronate administration was associated with a small increase or a stabilization in bone mass (L(1-4) BMD = 0.53% and 1.22%; femoral neck = 0.30% and 0.77%, respectively, with 1- and 6-day intravenous infusion regimens). From the 12th month, changes in spine and femoral neck BMD after continuous regimens were statistically different compared with that obtained with intermittent ones. Twenty-five of the 150 women (16.7%) discontinued the study before the end of the 2-year follow-up, but of these, only 7 dropped out because of adverse events related to the treatment itself. To summarize, intermittent clodronate administration could be a suitable option for the prevention of osteoporosis. PMID- 10710001 TI - The short-term changes of bone mineral metabolism following bone marrow transplantation. AB - Organ transplantation is now the treatment of choice for many patients with life threatening chronic diseases. A new set of side effects unique to these groups of patients has become recognized, and bone disease is one of these complications. However, little is known about the effects of myeloablative treatment followed by bone marrow transplantation (BMT) on bone mineral metabolism. We have prospectively investigated 31 patients undergoing BMT for hematologic diseases. Serum concentrations of calcium, phosphorus, creatinine, gonadotropins, sex hormones, and the biochemical markers of bone turnover were measured. The samples were collected before BMT and 1, 2, 3, 4, and 12 weeks, 6 months, and 1 year after BMT. Bone mineral density (BMD) was measured with dual-energy X-ray absorptiometry before BMT and 1 year after BMT. The serum carboxy-terminal cross linked telopeptide of type I collagen increased progressively until 4 weeks after BMT. Thereafter, it began to decrease and reached basal values after 1 year. Serum osteocalcin decreased progressively until 3 weeks after BMT. After that, it increased and reached basal values after 3 months. No distinct differences were observed in the serum biochemical turnover markers between males and females, or between patients who received total body irradiation and those who did not. One year after BMT, lumbar spine BMD had decreased by 2.2%, and total proximal femoral BMD had decreased by 6.2%. Eighty-six percent of the women (12/14) went into a menopausal state immediately after BMT. This was caused by high gonadotropin levels and low estradiol levels. In contrast, gonadotropin levels and testosterone levels did not change significantly in the male patients after BMT. In conclusion, the rapid impairment of bone formation and the increase in bone resorption, as shown by the biochemical markers in this study, might play a role in post-BMT bone loss. PMID- 10710002 TI - Altered biochemical markers of bone turnover in humans during 120 days of bed rest. AB - Microgravity induces significant and progressive bone loss in both humans and animals. This is the consequence of disturbed bone remodeling. We performed a bed rest experiment to simulate microgravity and tried to clarify bone metabolism by measuring biochemical markers of bone turnover. Six healthy volunteers participated in 120 days of bed rest. The parameters of calcium homeostasis, calcitropic hormones, and biochemical markers of bone turnover were examined. After ambulatory control evaluation, all subjects underwent 120 days of bed rest. Metabolic evaluation was performed in a baseline period, and on days 7, 16, 50, 72, 92, and 108 during bed rest, and on days 10 and 25 during a recovery period. Bed rest induced an increase in urinary calcium (Ca) excretion and serum Ca and bone resorption markers. Urine pyridinoline, deoxypyridinoline, and type I collagen cross-linked N-telopeptide increased more rapidly than urinary Ca excretion and serum Ca. Tartrate-resistant acid phosphatase (TRAP) increased even in the recovery period. Carboxy-terminal propeptide of type I collagen, a bone formation marker, significantly decreased on days 50, 92, and 108 of bed rest. These changes of biochemical markers of bone metabolism, except for TRAP, rapidly returned toward control levels in the recovery period. Immunoreactive parathyroid hormone showed a modest decrease during bed rest and a significant increase in the recovery period. Insulin-like growth factor I (IGF-I) and its binding protein, insulin-like growth factor binding protein-3, increased during bed rest, indicating the possibility of resistance to IGF-I in bones under reduced mechanical stress and strain. Bone loss from unloading results from the combination of acceleration of bone resorption and subsequent retardation of bone formation. PMID- 10710003 TI - COLIA1 polymorphism contributes to bone mineral density to assess prevalent wrist fractures. AB - Wrist fractures associated with postmenopausal women are only partially explained by osteoporosis. Recent studies have shown that polymorphism of an Spl binding site in the first intron of the collagen I alpha 1 gene (COLIA1) may determine risk for vertebral and nonvertebral fractures in post-menopausal women independent of bone mass. We investigated the relationship between the COLIA1 polymorphism, lumbar spine and femoral neck bone mineral density (BMD), ultrasound stiffness of the heel, anthropometric variables, and risk for wrist fractures in 126 Czech postmenopausal women with low bone mass who suffered one or more wrist fracture in the last 5 years and in 126 postmenopausal women with low bone mass without any fracture. Genotypes for the Spl COLIA1 polymorphism were determined by polymerase chain reaction, digestion with Ball restriction enzyme, and agarose gel electrophoresis. The test discriminates two alleles, S and s, which correspond to the presence of guanine and thymidine, respectively, at the first bases in the Spl-binding site in the first intron of the gene for CO LIA1. No significant differences were found between the fracture and control group with regard to age, weight, and years since menopause. However, BMD of the lumbar spine and femoral neck and ultrasound stiffness of the heel were significantly lower in patients with prevalent wrist fracture. Femoral neck BMD was the strongest determinant of prevalent fracture of the wrist. COLIA1 genotyping significantly strengthened prediction of prevalent fracture of the wrist. After multivariate adjustment, women in the Ss group had 2.0 times the risk of the women in the SS group (95% confidence interval [CI] = 1.1-3.8), and the women in the ss group had 2.8 times the risk of the women in the SS group (95% CI = 0.5-14.6). The overall gene-dose effect was an odds ratio of 2.1 per copy of the "s" allele (95% CI = 1.2-3.8). In the stepwise logistic regression, COLIA1 acted synergistically with femoral neck BMD and weight in increasing prediction of wrist fracture. The results demonstrate that COLIA1 Sp1 polymorphism is associated with an increased risk of wrist fracture in postmenopausal women independent of BMD and may be helpful in clinical practice by identifying patients with an increased fracture risk. PMID- 10710004 TI - Quantification of age-related changes in the structure model type and trabecular thickness of human tibial cancellous bone. AB - Structure model type and trabecular thickness are important characteristics in describing cancellous bone architecture. It has been qualitatively observed that a radical change of trabeculae from plate-like to rod-like occurs in aging, bone remodeling, and osteoporosis. Thickness of trabeculae has traditionally been measured using model-based histomorphometric methods on two-dimensional (2-D) sections. However, no quantitative study has been published based on three dimensional (3-D) methods on the age-related changes in structure model type and trabecular thickness for human peripheral (tibial) cancellous bone. In this study, 160 human proximal tibial cancellous bone specimens from 40 normal donors, aged 16 to 85 years, were collected. These specimens were micro-computed tomography (micro-CT) scanned, then the micro-CT images were segmented using optimal thresholds. From accurate 3-D data sets, structure model type and trabecular thickness were quantified by means of novel 3-D methods. Structure model type was assessed by calculating the structure model index (SMI). The SMI was quantified based on a differential analysis of the triangulated bone surface of a structure. This technique allows quantification of structure model type, such as plate, rod objects, or mixture of plates or rods. Trabecular thickness is calculated directly from 3-D images, which is especially important for an a priori unknown or changing structure. Furthermore, 2-D trabecular thickness was also calculated based on the plate model. Our results showed that structure model type changed towards more rod-like in the elderly, and that trabecular thickness declined significantly with age. These changes become significant after 80 years of age for human tibial cancellous bone, whereas both properties seem to remain relatively unchanged between 20 and 80 years. Although a fairly close relationship was seen between 3-D trabecular thickness and 2-D trabecular thickness, real 3-D trabecular thickness was significantly underestimated using 2 D method. PMID- 10710005 TI - Discriminant capacity of quantitative ultrasound versus dual X-ray absorptiometry to determine cancellous bone loss in ovariectomized rats. AB - The capacity of dual x-ray absorptiometry and quantitative ultrasound to discriminate bone loss and to predict the mechanical and microarchitectural properties of cancellous bone in an animal model of osteopenia was evaluated. Thirty-five female Sprague-Dawley rats (10 months old) were randomized into three groups: baseline group, 10 rats killed at the beginning of the study; ovx group, 15 rats ovariectomized; and sham group, 10 rats sham operated. At the beginning and end of the study, all the animals underwent osteosonography to record the proximal tail (C3 vertebra) bone speed of sound. Sixteen weeks after surgery, the animals were euthanized and the L5-6 lumbar vertebrae of each rat were excised for densitometric, biomechanical (compression test), and histomorphometric studies. Significant differences were found among the groups for final speed of sound (p = 0.01). The L5 bone mineral density of the ovx group decreased by 12.1% (p = 0.049) and 12.6% (p = 0.035) compared, respectively, with baseline and sham groups. The biomechanical parameters of the ovx group decreased by 15-47% compared with the other groups, showing significant differences between the ovx and sham groups both for maximal stress (p = 0.026) and elastic modulus (p = 0.013). Histomorphometric parameters of the ovx group showed significant decreases in comparison with other groups. Logistic regression analysis showed that dual X-ray absorptiometry and quantitative ultrasound discriminate ovariectomized and healthy rats with a similar capacity, classifying correctly all rats used in the model in a range of 61-70%. This similar capacity seems to derive from two different capacities to detect bone changes. Dual X-ray absorptiometry, depending on bone mineralization and density, is able to detect modifications in bone stiffness and strength, confirmed also by the correlation with biomechanical data. On the contrary, quantitative ultrasound seems to depend more on cancellous bone microarchitecural changes because it is correlated to histomorphometric parameters. PMID- 10710006 TI - Spatial clustering of remodeling osteons in the femoral neck cortex: a cause of weakness in hip fracture? AB - Intracapsular femoral neck fractures are associated with decreased cortical width and increased proportions of Haversian canals with diameters greater than the normal mean plus 3 SD (i.e., >385 microm). Such canals might be formed if closely associated resorbing osteons merge; a cortical event analogous with the loss of cancellous connectivity. To test this, we investigated the pattern of osteon distribution in the aging femoral neck to determine if remodeling osteons were distributed in anatomical clusters. Femoral neck biopsies from female patients with intracapsular hip fractures (n = 13) were compared with age/gender-matched cadaveric controls (n = 13). Solochrome-stained sections were analyzed for Haversian canal location, canal diameter, and the presence of an osteoid surface. Clustering was investigated using statistical software with a cluster defined as two or more osteoid-bearing osteon centers within 0.75 mm of each other. Clusters occurred more frequently than would be expected by chance (p < 0.001). Fracture cases had more clusters per unit area (3.14 +/- 0.31 clusters/25 mm2 of cortical bone) than controls (1.89 +/- 0.22) (p = 0.002). In fracture cases, the antero inferior, antero-superior, and infero-anterior regions had more clusters per 25 mm2 than comparable control regions (ant/inf: 4.12 +/- 0.79, 1.70 +/- 0.60,p = 0.025; ant/sup: 5.31 +/- 1.1, 1.80 +/- 0.59,p = 0.013; inf/ant: 3.15 +/- 0.49, 1.27 +/-0.29, p = 0.004). The mean number of clusters per 25 mm2 per region correlated with the mean porosity per region (adjusted r2 = 0.60;p = 0.014), and the total number of giant canals per region correlated with the total number of clusters per region (adjusted r2 = 0.58; p = 0.011). In conclusion, remodeling osteons are clustered or grouped anatomically, and fracture cases have more clusters than controls. Our data suggest that merging of adjacent, clustered osteons during resorption could lead to the rapid development of canals with excessive diameters and focal weakness. Clustering is greatest in those regions that we have previously shown to have the largest relative reductions in bone strength compared with controls and known to be maximally loaded during a sideways fall. This implicates the remodeling process underlying clustering of remodeling osteons in the aetiology of hip fracture. PMID- 10710007 TI - Use of robotic technology for diathrodial joint research. AB - Knowledge of diarthrodial joint mechanics and specific function of the ligaments are needed in order to understand injury mechanisms, improve surgical procedures and design better post-surgical rehabilitation protocols. To facilitate these needs, a robotic/universal force-moment sensor (UFS) testing system was developed to measure joint kinematics in multiple degree-of-freedom and the in situ forces in the ligaments. When operated in the position control mode, the testing system applies a known load to the intact joint while the motion and force data are recorded. After transection of a ligament, the recorded motion for the intact joint is repeated and new force and moment data is recorded by the UFS. Since the robot reproduces the identical initial position as well as path of joint motion before and after a ligament is transected, the in situ force in the ligament is the difference between the two sets of force and moment data. In force control mode, a known force is applied to the intact knee while the kinematics are recorded. After ligament transection, the same force is applied while the changes in kinematics are again recorded. Testing in this mode is similar to a clinical examination that diagnoses ligament injury. To date, this testing system has been used for experimental studies that examine the anterior cruciate ligament & posterior cruciate ligament of the knee and ligaments of the shoulder. A three dimensional finite element model has also been constructed based on CT/MRI scans of a knee specimen and validated using data obtained with the testing system. Once in vivo kinematics (such as during gait analysis or throwing activities) are available, the robotic/UFS testing system can be programmed to reproduce these joint kinematics on young human cadaveric specimens in order to generate a database for in situ forces in the ligaments, or Ligament replacement grafts. With appropriate computational models, the stresses and strains in these tissues in vivo can also be determined. Potential applications of this combined approach include pre-operative surgical planning, improvement of surgical procedures as well as development of appropriate post-operative rehabilitation protocols. PMID- 10710009 TI - The effect of intramuscular iron injections on serum ferritin levels and physical performance in elite netballers. AB - The purpose of this study was to determine the effect of iron supplementation by intramuscular injection on both serum ferritin (SF) levels and exercise performance in iron depleted, non-anaemic elite female netballers. Fifteen iron depleted (Serum Ferritin <40 ug x L(-1). Haemoglobin >125 g x L(-1)) subjects (19+/-3 y) first performed their routine test battery: a vertical jump test, a 10s power and 5x6s repeat sprint test on a cycle ergometer and a 20m multi-stage shuttle run. Subjects were matched on the basis of height, mass, and playing position and then assigned to either a Ferritin Group (FG) or Placebo Group (PG) (single blind design). Subjects then underwent a course of 5x2ml intramuscular injections of either Ferrum H (FG) or normal saline (PG) over a period of 8-10 days before repeating the blood and physical performance tests. Five and 10 days following supplementation, SF levels in the FG increased significantly from baseline levels (P<0.05) and were also significantly greater than levels measured in the PG (P<0.01). Haemoglobin levels remained unchanged in both groups. All test scores remained unchanged from baseline values and were not different between the two groups. These results demonstrate that a course of 5x2ml intramuscular iron injections significantly increased SF concentration within 2 weeks without increasing Hb levels, but this rapid elevation did not enhance the physical performance in selected tests of iron depleted, non-anaemic athletes. PMID- 10710008 TI - Biomechanics: an integral part of sport science and sport medicine. AB - Biomechanics is one of the disciplines in the field of Human Movement and Exercise Science and it can be divided into three broad categories from a research perspective. Clinical biomechanics involves research in the areas of gait, neuromuscular control, tissue mechanics, and movement evaluation during rehabilitation from either injury or disease. Occupational biomechanics typically involves research in the areas of ergonomics and human growth or morphology as they influence movement. While these two categories will briefly be discussed, the primary aim of this paper is to show the role of biomechanics in sports science and sports medicine. Research in sports biomechanics may take the form of describing movement from a performance enhancement (such as matching of impulse curves in rowing) or injury reduction perspective (such as diving in swimming or the assessment of knee joint loading during downhill walking). However, the strength of sports biomechanics research is the ability to establish an understanding of causal mechanisms for selected movements (such as the role of internal rotation of the upper arm in hitting or striking, and the influence of elastic energy and muscle pre-stretch in stretch-shorten-cycle actions). The growth of modelling and computer simulation has further enhanced the potential use of sports biomechanics research (such as quantification of knee joint ligament forces from a dynamic model and optimising gymnastics performance through simulation of in-flight movements). Biomechanics research may also play an integral role in reducing the incidence and severity of sporting injuries (such as identification of the causes of back injuries in cricket, and the causes of knee joint injuries in sport). In the following discussion no attempt will be made to reference all papers published in each of these areas because of the enormity of the task. Published and current work from the biomechanics laboratory at the Department of Human Movement and Exercise Science at The University of Western Australia will generally be used to illustrate the scope of biomechanics research within that institution. PMID- 10710010 TI - Do changing patterns of heat and humidity influence thermoregulation and endurance performance? AB - The purpose of this project was to determine whether changing patterns of temperature and humidity, as expected in the morning versus afternoon, had a differential effect on thermoregulation and endurance performance. Eight male distance runners each participated in two heat pattern tests consisting of two hours treadmill running at 70%-maximum oxygen consumption. The mean heat load for each test was identical (22.2 degrees C wet bulb temperature) but either dry bulb temperature increased (24 to 27.5 degrees C) or decreased (27.5 to 24 degrees C) over the course of the two hour heat stress test. Whole body sweat rate was 10.7% higher (p<0.05) and there was greater plasma volume loss (2.7 versus 1.6%, p<0.05) in the cooling versus warming pattern test. Mean skin and body temperature changed in a significantly different (p<0.05) manner between the two patterns and closely followed ambient dry bulb temperature change. The thermoregulatory variables of heart rate and rectal temperature were not affected and performance did not differ between pattern tests. Ratings of perceived exertion (RPE) and oxygen consumption were also not significantly different between cooling and warming test. In summary, although some minor differences were noted, thermal homeostasis was maintained equally well during either warming or cooling for wet bulb temperatures between 24 and 27 degrees C. The mean heat load is therefore more important than changing patterns of temperature and humidity in determining an individual's physiological response to exercise in a warm environment. PMID- 10710011 TI - The reliability of muscle biopsies taken from vastus lateralis. AB - The purpose of this study was to examine whether a single biopsy sample of vastus lateralis could provide an accurate estimate of capillary density (CD) which is indicative of the entire muscle, or whether capillary density is distributed unevenly and varies with muscle depth. Whole muscle cross sections of vastus lateralis were excised post mortem (n=11) for analysis of capillary density at three muscle depths, (superficial, mid and deep regions). Muscle thickness varied widely (17-79 mm) across subjects. The distribution of CD throughout the depth of the muscle was homogeneous in 8 subjects, but in 3 subjects it was heterogeneous (p<0.05). In 3 of these subjects there was a significant (p<0.05) effect of sample depth on CD. These data indicate that tissue from a single biopsy will not adequately represent the CD of the entire vastus lateralis in some individuals. Single biopsies from unspecified muscle depth, have routinely been used to estimate CD and fibre type in vastus lateralis. The present study indicates that a more reliable method of analysis would be to use the tissue from two needle biopsies taken at the superficial and deep portions of the muscle from a group of at least 10 subjects. Sampling theory analysis supported this conclusion. PMID- 10710012 TI - Muscular strength, body composition and health responses to the use of testosterone enanthate: a double blind study. AB - To determine the effect the steroid, testosterone enanthate (TE) had on upper body strength, body composition and health. Twenty one male weight training subjects were randomly assigned in a double blind method to either a 3.5 mg(-1) x kg(-1) TE (n=11) or placebo (n=10) weight training group. The subjects were monitored during a 12 week administration phase and a subsequent 12 week follow up phase. Subjects were tested on a number of strength and size measurements, whilst having their health monitored. The results from the study revealed that the testosterone/weight training group improved significantly (p<0.05) more than the placebo/weight training group during and immediately after the administration phase on a 1 repetition maximum bench press. With regards to body composition, body weight, arm girth and rectus femoris circumference all increased significantly greater in the TE group compared to the placebo. Furthermore, the abdomen skinfold showed significant decreases in the TE group compared to the placebo group at post testing, follow up mid testing and the follow up post testing occasions. With the exception of the abdomen skinfold no within or between group differences were evident following a cycling off period of 12 weeks. Changes to baseline health indicators were reported in some subjects following testosterone usage. This included an average elevation in systolic blood pressure in all TE subjects by 10 mm Hg, a mild increase in hereditary frontal alopecia, increased muscle tightness (hamstrings and pectorals), a mild increase in libido over the first two weeks with a subsequent fall to normal, mild acne, subjective changes to personality including an increase in aggression, irritability and positive mood responses. Consequently, moderate doses of TE combined with weight training can result in short term significant changes in upper body strength and body composition, with corresponding changes to baseline health in some individuals. PMID- 10710013 TI - Prolonged incremental tests do not necessarily compromise VO2max in well-trained athletes. AB - Existing literature suggests that tests for maximal oxygen uptake (VO2max) should last 8-12 minutes and that prolonged tests do not produce valid measurements. The research underlying this suggestion has been performed with non-athletic populations and trained athletes may be more tolerant of longer protocols. Eleven rowers (8 males, 3 females) each underwent four different incremental tests on a standard rowing ergometer in randomised counterbalanced order. One of the tests was continuous with workload increments each minute (IT1MIN). This test lasted an average of 12 min+/-4 s (SEM). The other three tests were discontinuous and consisted of 7 stages separated by 1-minute recovery intervals. Stage durations of 3, 4 and 5 min were used in the different tests (IT3MIN, IT4MIN and IT5MIN). Mean values for VO2max were almost identical for IT1MIN (4.56+/-0.22 L x min( 1)), IT3MIN (4.60+/-0.23 L x min(-1)) and IT4MIN (4.60+/-0.21 L x min(-1)), while IT5MIN produced a significantly lower value (4.47+/-0.21 L x min(-1), p<0.05). There was no significant difference between protocols in peak post-exercise blood lactate concentration (approx 13 mmol x L(-1) in each case), but IT1MIN produced lower peak heart rates and higher respiratory exchange ratios. We conclude that with well trained rowing athletes discontinuous test protocols involving 7 stages of 3-4 minutes duration can provide valid measurements of VO2max. PMID- 10710014 TI - The relationship between dynamic, isokinetic and isometric strength and bone mineral density in a population of 45 to 65 year old women. AB - This study investigated the relationship between age, lumbar spine bone mineral density (LS BMD) and muscular strength of peri and postmenopausal women between 45 and 65 years either taking or not taking hormone replacement therapy (HRT). Ninety six women were tested for LS BMD (L2-L4), one repetition maximum (1RM) bench press and squat, maximal voluntary isometric contraction (MVC) of the knee extensors and peak torque of back extensor muscles at a speed of 30 degrees s( 1). Bone and strength data were analysed to evaluate the relationships in incrementing five year age groups and based on groups either taking or not taking HRT. ANOVA revealed significant differences in LS BMD between the 45-49 and 55-59 (F[3,92]=2.6411, p<0.05; -8%) age groups amounting to an annual bone loss of 0.8% for this Australian based population. Non significant LS BMD results were evident after controlling for the influence of age and menopausal status on the groups either taking or not taking HRT. Significant differences between the 45-49 and 60 64 (F[3,92]=2.7463, p<0.05) age groups for 1RM bench press and the 45-49 and 60 64, 50-54 and 60-64, and, 55-59 and 60-64 (F[3,92]=4.2816, p<0.05) age groups for 1RM squat amounting to an 18.8% and 37.5% loss of dynamic upper and lower body strength, respectively. Group correlation coefficients ranged between (r=-0.20 and -0.34, p<0.05) for LS BMD, strength and age. The conclusions demonstrate a concomitant decline in maximal muscle strength and bone density between women 45 and 65 years irrespective of HRT. These results also demonstrate a 50% greater decline in lower body strength compared to upper body strength between women 45 and 65 years. PMID- 10710015 TI - The physical demands of Olympic yacht racing. AB - The primary purpose of this study was to quantify the up wards forces of the feet on the hiking strap and the forces in the mainsheet of four Olympic classes of racing dinghies (Europe, Laser. Finn and 470) during realistic on-water sailing in varying wind conditions. The secondary aim of the study was to measure the joint angles adopted by the sailors and boat heel angles. The tertiary aim was to identify events and sailing conditions associated with large or patterned force production. Forces in the hiking strap and mainsheet of four classes of Olympic sailing dinghies were measured on eleven New Zealand sailors during simulated on water racing in a range of wind conditions. Up-wind hiking strap forces reached an average of 73-87% of predicted maximal voluntary contraction (pred MVC), with peak forces exceeding 100% pred MVC. Mainsheet forces reached 25-35% pred MVC, with peak forces reaching 40-50% pred MVC. Off-wind hiking strap and mainsheet forces were considerably lower than up-wind forces. Ankle and hip joint angles increased and knee joint angles decreased with increasing wind speed during up wind sailing. Large forces occurred in the hiking strap and mainsheet when boats reached the tops of wave during up-wind sailing in high wind speeds and when a gust of wind hit the boat. During off-wind sailing large forces were observed in the mainsheet when surfing down waves. It is recommended that the intensities and joint angles found in this study be used as a basis for the development of class specific off-water physical conditioning programmes. PMID- 10710016 TI - The evolution of Australian football. AB - Australian football has undergone considerable change over the past century. This evolution seems to have accelerated more recently since the introduction and major influence of the media, increased professionalism and the start of a national competition. In this study we have attempted to quantify the evolution in game 'style' by measuring events during elite football games (from video analysis) and gathering physical information on players involved at the highest level. These data are important to gain insight into the game demands so that player preparation may be enhanced and when predicting the nature of the game in the future. Understanding the patterns of play within the game may also be useful when assessing the possible impact of rule changes, for example, increasing the number of interchange players on the potential for injury. Four games were selected, one from each of the past 4 decades to determine the rate at which specific, measurable events occurred in the games. Height and mass data on players were also obtained from official records of registered players in the VFL/AFL competitions. The results indicate the 'speed' of the game has approximately doubled in the period 1961-1997. The proportion of the total game which involves 'play' time has been reduced significantly while breaks in play are more frequent and longer. Despite this pattern, however, the average game tempo has increased along with player height and mass and we present a case which suggests these are likely determinants of the increased incidence of player injuries and lost match time. PMID- 10710017 TI - Decreasing sedentary behaviors in treating pediatric obesity. AB - BACKGROUND: Epidemiological studies have shown television watching to be a risk factor for the development of obesity in children. The effect of reducing television watching and other sedentary behaviors as a component of a comprehensive obesity treatment program has not been thoroughly tested. OBJECTIVE: To compare the influence of targeting decreases in sedentary behavior vs. increases in physical activity in the comprehensive treatment of obesity in 8 to 12-year-old children. DESIGN: Randomized, controlled outcome study. SETTING: Childhood obesity research clinic. DESIGN: Ninety families with obese 8- to 12 year-old children were randomly assigned to groups that were provided a comprehensive family-based behavioral weight control program that included dietary, and behavior change information but differed in whether sedentary or physically active behaviors were targeted and the degree of behavior change required. RESULTS: Results during 2 years showed that targeting either decreased sedentary behaviors or increased physical activity was associated with significant decreases in percent overweight and body fat and improved aerobic fitness. Self-reported activity minutes increased and targeted sedentary time decreased during treatment. Children substituted nontargeted sedentary behaviors for some of their targeted sedentary behaviors. CONCLUSION: These results support reducing sedentary behaviors as an adjunct in the treatment of pediatric obesity. PMID- 10710018 TI - An evaluation of a safety education program for kindergarten and elementary school children. AB - OBJECTIVE: To determine the effectiveness of a safety education program, Safety City, that is designed to teach kindergarten and first grade children how to cross the street, call 911 in an emergency, and avoid strangers. PARTICIPANTS/SETTING: Kindergarten students at 10 urban elementary schools. DESIGN: Each school was randomized to either the intervention or control group. An evaluation tool was administered to all participants as a pretest. The Safety City program was then presented to the intervention schools. Afterward, the same evaluation tool was used as a post-test. The posttest was administered to the intervention group 6 months after the Safety City program was presented. The control group took the posttest 6 months after the pretest. MAIN OUTCOME MEASURE: Change in individual test scores. RESULTS: One hundred eighty-one children completed the pretest and posttest evaluations. There was no statistical difference in the change between pretest and posttest scores of children who participated in the Safety City program and those in the control group (crossing the street, P = .29; calling 911, P = .41; stranger avoidance, P = .57). CONCLUSIONS: Exposure to the Safety City program did not achieve the desired changes in safety knowledge among participants. This is most likely owing to the fact that Safety City attempts to convey a large amount of relatively complex information to young children in a brief period. We conclude that programs such as Safety City are not sufficient to teach children these behaviors. This report also emphasizes the importance of building an evaluation component into educational programs. PMID- 10710019 TI - Workplace toxic exposures involving adolescents aged 14 to 19 years: one poison center's experience. AB - BACKGROUND: While many previous reports describe injuries to adolescents in the workplace, few focus on toxic substance exposures among such injuries. Yet low skill, entry-level jobs pose a particular hazard of toxic exposure owing to the frequent use of cleaning agents, solvents, and/or other chemicals in carrying out assigned tasks. OBJECTIVE: To analyze the types and severity of adolescent occupational toxic exposures. DESIGN: Secondary analysis of calls to a single regional poison control center (PCC). SETTING: Massachusetts PCC poisoning consultations between 1991 and 1996. SUBJECTS: Children aged 19 years or younger reporting toxic exposures occurring in the workplace. RESULTS: Of 7024 occupational toxic exposures recorded by the PCC in the 6 years of study, 269 incidents (3.8%) involved adolescents aged 14 to 19 years (median age, 18 years; 124 aged 14-17 years and 145 aged 18-19 years; 65% were male). The most frequently involved agents were cleaning compounds (27.8%); paints, solvents, and glues (9.0%); caustics (8.7%); hydrocarbons (8.7%); and bleaches (7.3%). Of 88 cases (32.7%) in which a worksite was identified, food services (30.7%), automotive services (14.8%), and general retail stores (12.5%) were the most common locations. One hundred fifty-six patients (58.0%) were triaged to an emergency department; 7 were hospitalized. Forty-three subjects (16.0%), 18 who were between the ages of 14 and 17 years and 25 who were aged 18 or 19 years, were judged to have moderate to severe injuries. There were no deaths. CONCLUSIONS: This study confirmed the usefulness of PCC surveillance as a source of information about adolescent toxic exposures occurring in the workplace. The occupational toxic exposures reported here most commonly involved cleaning agents, solvents, paints, caustics, and bleach used in those entry-level jobs most frequently filled by adolescents. We conclude that occupational toxic exposures are an underrecognized adolescent injury, and that PCC experience can be used to fill a gap in the surveillance of such workplace-associated events. PMID- 10710021 TI - Current expectations for survival in pediatric burns. AB - BACKGROUND: Conventional wisdom and published reports suggest that children, particularly those younger than 48 months, have higher mortality rates after burns than young adults. However, coincident with refinements in resuscitation, operative techniques, and critical care, survival rates for children with burns seem to have improved. To document this change and to define current expectations, a review of deaths during two 7-year intervals separated by a decade was done. DESIGN: We examined the clinical course of children who died after admission for care of acute thermal burns during two 7-year intervals: calendar years 1974 to 1980 inclusive (group 1) and 1991 to 1997 inclusive (group 2). Dying children were stratified by total body surface area (TBSA) burned: small (0%-39%), midsize (40%-59%), and large (60%-100%) TBSA burns. Children who arrived with anoxic brain injury or in a moribund state with refractory shock were excluded from analysis (4 children in group 1 and 5 in group 2); 2 of these children in group 2 died and became solid organ donors. SETTING: Regional pediatric burn center. PATIENTS: Six hundred seventy-eight children in group 1 and 1150 children in group 2. MAIN OUTCOME MEASURE: Survival. RESULTS: In children with 0% to 39% TBSA burns, mortality was 0.6% in group 1 and 0% in group 2 (Fisher exact test, P = .04; chi2 test, P = .02). In children with 40% to 59% TBSA burns, mortality was 7.7% in group 1 and 0% in group 2 (Fisher exact test, P = .07; chi2 test, P = .047). In children with 60% to 100% TBSA bums, mortality was 33.3% ingroup 1 and 14.3% in group 2 (Fisher exact test, P = .04; chi2 test, P = .02). Although 59% of the children in group 2 were younger than 48 months, including 55% of those with 40% to 59% TBSA burns and 41% of those with 60% to 100% TBSA burns, there were no deaths in this age group. CONCLUSION: Survival rates after burns have improved significantly for children. At present, most children, even young children and children with large burns, should survive. PMID- 10710020 TI - Psychiatric diagnoses in adolescents seropositive for the human immunodeficiency virus. AB - OBJECTIVE: To provide a descriptive analysis of the prevalence of past and current psychiatric disorders in adolescents positive for the human immunodeficiency virus (HIV). DESIGN: Structured interview in a convenience sample in a primary care urban adolescent clinic in Washington, DC. PARTICIPANTS: Thirty-four HIV-seropositive adolescents ranging in age from 16 to 21 years. MAIN OUTCOME MEASURES: The Structured Clinical Interview for DSM-IV Axis I Disorders Patient Edition (SCID-P) was administered by a child psychiatrist or a clinical child psychologist. Extensive review of medical records was also conducted. RESULTS: A majority of the HIV-infected adolescents in our sample had received psychiatric diagnoses prior to their treatment at the clinic (53%), had a documented history of sexual abuse (50%), and had a history of substance use (82%). Psychiatric diagnoses determined by the SCID-P indicated that 85% of the sample had a current Axis I disorder, with 44% reporting ongoing depressive disorders. CONCLUSIONS: The majority of subjects in this sample had had a previous psychiatric diagnosis, and almost half had a current affective disorder. Psychiatric disorders, especially affective disorders, may be a risk factor for high-risk sexual behaviors and substance use that increases the risk for HIV infection in adolescent populations. PMID- 10710022 TI - Predictors of intussusception in young children. AB - OBJECTIVE: To identify predictors of intussusception in young children. DESIGN: A retrospective cross-sectional study. SETTING AND PATIENTS: A consecutive sample of children younger than 5 years on whom contrast enemas were performed because of suspected intussusception seen at an urban children's hospital from 1990 to 1995. METHODS: We evaluated historical, clinical, and radiographic variables. Variables documented in 75% or more of the medical records and associated with intussusception (P< or =.20) in the univariate analysis were evaluated in a multiple logistic regression analysis. Variables retaining significance (P< or =.05) in the multivariate analysis were considered independent predictors of intussusception. We used bootstrap resampling techniques to validate the multivariate model. RESULTS: Sixty-eight (59%) of the 115 patients had intussusception. Univariate predictors of intussusception included male sex, age younger than 2 years, history of emesis, rectal bleeding, lethargy, abdominal mass, and a highly suggestive abdominal radiograph. In the multivariate analysis, we identified only 4 independent predictors (adjusted odds ratio; 95% confidence interval): a highly suggestive abdominal radiograph (18.3; 4.0-83.1), rectal bleeding (17.3; 2.9-104.0), male sex (6.2; 1.2-32.3), and a history of emesis (13.4; 1.4-126.0). We identified 3 of these 4 variables (all but emesis) as independent predictors in more than 50% of 1000 bootstrap data samples. CONCLUSIONS: Rectal bleeding, a highly suggestive abdominal radiograph, and male sex are variables independently associated with intussusception in a cohort of children suspected of having this diagnosis. Knowledge of these variables may assist in clinical decision making regarding diagnostic and therapeutic interventions. PMID- 10710023 TI - Information collected during the residency match process does not predict clinical performance. AB - OBJECTIVE: To determine whether information collected during the National Resident Matching Program (NRMP) predicts clinical performance during residency. METHODS: Ten faculty members rated the overall quality of 69 pediatric house officers as clinicians. After rating by the faculty, folders were reviewed for absolute rank on the NRMP match list; relative ranking (where they ranked in their postgraduate year 1 [PGY-1] group); scores on part I of the National Board of Medical Examiners (NBME) examination; grades during medical school pediatrics and internal medicine rotations; membership in the Alpha Omega Alpha Medical Honor Society; scores of faculty interviews during intern application; scores on the pediatric in-service examination during PGY-1; and scores on the American Board of Pediatrics certification examination. RESULTS: There was substantial agreement among faculty raters as to the overall quality of the residents (agreement rate, 0.60; kappa = 0.50; P = .001). There was little correlation between faculty ratings and absolute (r = 0.19; P = .11) or relative (r = 0.20; P = .09) ranking on the NRMP match list. Individuals ranked in the top 10 of the match list had higher faculty ratings than did their peers (mean +/- SD, 3.66+/ 1.22 vs. 3.0+/-1.27; P = .03), as did individuals ranked highest in their PGY-1 group (mean +/- SD, 3.88+/-1.45 vs. 3.04+/-1.24; P = .03). There was no correlation between faculty ratings and scores on part I of the NBME examination (r = 0.10; P = .49) or scores on the American Board of Pediatrics certification examination (r = 0.22; P = . 11). There were weak correlations between faculty ratings and scores of faculty interviews during the intern application process (r = 0.27; P = .02) and scores on the pediatric in-service examination during PGY-1 (r = 0.28; P = .02). There was no difference in faculty ratings of residents who were elected to Alpha Omega Alpha during medical school (mean +/- SD, 3.32+/ 1.21) as compared with those who were not (mean +/- SD, 3.08+/-1.34) (P = .25). CONCLUSIONS: There is significant agreement among faculty raters about the clinical competence of pediatric residents. Medical school grades, performance on standardized examinations, interviews during the intern application process, and match-list ranking are not predictors of clinical performance during residency. PMID- 10710024 TI - Psychosocial morbidity: the economic burden in a pediatric health maintenance organization sample. AB - OBJECTIVES: To evaluate psychosocial morbidity in pediatric primary care and to determine displaced health care utilization. DESIGN AND SETTING: A cross sectional sample of parent-child dyads was screened using the Pediatric Symptom Checklist (PSC) at 6 pediatric sites of a health maintenance organization (HMO). Cost and utilization data were retrieved from regional databases for this sample. PARTICIPANTS: Parent-child dyads from an HMO in northern California (N = 1840). The children ranged in age from 2 to 18 years. RESULTS: In all, 13.0% of children exhibited psychosocial dysfunction. The rate of children's chronic illness was 18.4%. Multiple regression analyses measured utilization and cost of health and psychiatric care for the selected population for the previous year; the average log cost of health care per child was $393. The average health care cost for children with anxious, depressed symptoms was $805. Chronically ill children were the highest utilizers of health care, with an average log cost of $1138. When psychosocial dysfunction was present, regression models showed that health care spending was highest for young children. CONCLUSIONS: Health care utilization was higher for children with psychosocial morbidity, was higher among younger children, and decreased with age as psychiatric costs progressively increased. PMID- 10710025 TI - Palatability of oral antibiotics among children in an urban primary care center. AB - OBJECTIVE: To evaluate the palatability of antimicrobial agents effective against beta-lactamase-producing bacteria in American children. DESIGN: In a taste test of 4 antimicrobial agents, azithromycin (cherry flavored), cefprozil (bubble gum flavored), cefixime (strawberry flavored), and amoxicillin-clavulanic acid (banana flavored) were compared. SETTING: An urban inner-city primary care clinic. SUBJECTS: A volunteer sample of 30 healthy children (aged 5-8 years). INTERVENTION: Palatability was determined using a single-blind taste test of 4 flavored antimicrobial agents. The 4 antimicrobial agents used were azithromycin, cefprozil, cefixime, and amoxicillin-clavulanic acid. MAIN OUTCOME MEASURES: After each antimicrobial test dose, subjects rated the taste on a 10-cm visual analog scale incorporating a facial hedonic scale. Preference assessments for the best-tasting and worst-tasting agent were also conducted. RESULTS: Of the 20 children who expressed a preference, significantly more children (9 [45%], P<.05) selected the cefixime preparation as the best-tasting formulation compared with the other preparations. The cefixime preparation was also significantly the least likely to be selected as the worst-tasting preparation (2 [10%], P<.05). There were no significant differences between the other 3 preparations with respect to being selected as either the best or worst tasting. The mean (+/- SD) visual analog scale score for cefixime was highest (8.53 [2.49]) compared with the scores for azithromycin (6.78 [3.45]), cefprozil (6.26 [4.04]), and amoxicillin clavulanic acid (6.24 [4.01]). CONCLUSION: The cefixime preparation was most commonly rated as best tasting by children. PMID- 10710026 TI - Pulmonary hemorrhage in an infant following 2 weeks of fungal exposure. AB - BACKGROUND: Exposure to indoor fungus growth and tobacco smoke has been epidemiologically linked to unexplained pulmonary hemorrhage in infants. OBJECTIVE: To describe the 40-day-old male infant who had been exposed to fungi for a discrete 2-week period followed by acute exposure to environmental tobacco smoke prior to development of a life-threatening pulmonary hemorrhage. PATIENT AND METHODS: History and clinical evaluation of the infant immediately followed the pulmonary hemorrhage. Air and surface sampling for isolation and identification of fungal growth in the dwelling where the infant resided before the acute hemorrhage was accomplished when the homeowner returned from vacation 4 months after the clinical event. RESULTS: Two fungi associated with mycotoxin production were cultured from surface samples collected in the residence: Penicillium (possibly Penicillium purpurogenum) and a Trichoderma species. Stachybotrys atra was not isolated from air or surface samples. Environmental tobacco smoke exposure occurred over a discrete several-hour period prior to onset of the acute pulmonary hemorrhage. CONCLUSIONS: Avoidance of unnecessary exposure of infants to fungus growth in water-damaged environments or exposure to tobacco smoke is prudent. Further investigation into the toxic effects of indoor fungi as causes of infantile pulmonary hemorrhage is warranted. PMID- 10710027 TI - Computer crash simulations in the development of child occupant safety policies. AB - OBJECTIVE: To address the predictability of injury from air bag activation by use of crash simulation software. METHODS: Using current, validated crash simulation software, the effect of air bag activation on injury risk was assessed for the 6 year-old child, both restrained and unrestrained. Results were compared with those for adult occupants in similar crash scenarios. RESULTS: For the unrestrained child passenger, crash simulations predicted serious head, neck, and chest injuries with air bag activation, regardless of crash severity. For the restrained child passenger, crash simulations predicted similar severe injuries for high-severity crashes only. No serious injuries were predicted for unrestrained male adults exposed to air bags or for child passengers restrained in the rear seat for the crash scenarios simulated. CONCLUSIONS: Using current crash simulation software, this study demonstrated that the risk of air bags to school-aged children could be predicted. Our results confirmed the previously identified risks to unrestrained children and provided the first evidence that air bags, in their current design, are not beneficial to restrained children. This study illustrates that computer crash simulations should be used proactively to identify injury risks to child occupants, particularly when limited real-world data are available. PMID- 10710028 TI - Latex hypersensitivity in a child with diabetes. AB - BACKGROUND: A 6-year-old girl who was diagnosed with diabetes mellitus 20 months previously developed erythematous, raised lesions at the site of her insulin injections. The reactions occurred when isophane and lispro insulin were administered individually or combined but not when insulin was obtained from the bottle after the septum had been removed. OBJECTIVES: To describe latex hypersensitivity in a child with diabetes and to review the literature. DESIGN: Case report. RESULTS: Findings from intradermal testing confirmed latex hypersensitivity. A change to insulin administration by insulin pen decreased the frequency of the reactions. CONCLUSION: Latex hypersensitivity should be considered in children with type 1 diabetes who develop local reactions to insulin injections. PMID- 10710029 TI - Injury prevention practices as depicted in G-rated and PG-rated movies. AB - BACKGROUND: Previous studies on alcohol, tobacco, and violence suggest that children's behavior can be influenced by mass media; however, little is known about the effect of media on unintentional injuries, the leading cause of death among young persons in the United States. OBJECTIVE: To determine how injury prevention practices are depicted in G-rated (general audience) and PG-rated (parental guidance recommended) movies. DESIGN: Observational study. SETTING: The 25 movies with the highest domestic box-office grosses and a rating of G or PG for each year from 1995 through 1997. Movies that were predominantly animated or not set in the present day were excluded from analysis. SUBJECTS: Movie characters with speaking roles. MAIN OUTCOME MEASURES: Safety belt use by motor vehicle occupants, use of a crosswalk and looking both ways by pedestrians crossing a street, helmet use by bicyclists, personal flotation device use by boaters, and selected other injury prevention practices. RESULTS: Fifty nonanimated movies set in the present day were included in the study. A total of 753 person-scenes involving riding in a motor vehicle, crossing the street, bicycling, and boating were shown (median, 13.5 person-scenes per movie). Forty two person-scenes (6%) involved falls or crashes, which resulted in 4 injuries and 2 deaths. Overall, 119 (27%) of 447 motor vehicle occupants wore safety belts, 20 (18%) of 109 pedestrians looked both ways before crossing the street and 25 (16%) of 160 used a crosswalk, 4 (6%) of 64 bicyclists wore helmets, and 14 (17%) of 82 boaters wore personal flotation devices. CONCLUSIONS: In scenes depicting everyday life in popular movies likely to be seen by children, characters were infrequently portrayed practicing recommended safe behaviors. The consequences of unsafe behaviors were rarely shown. The entertainment industry should improve its depiction of injury prevention practices in G-rated and PG rated movies. PMID- 10710030 TI - Prevalence, impact, and trends in childhood disability due to asthma. AB - BACKGROUND: Although not widely recognized as such, asthma is the single most prevalent cause of childhood disability and has contributed to a substantial rise in the overall prevalence of disability among children during the past 25 years. OBJECTIVE: To provide a national profile of the prevalence, impact, and trends in childhood disability due to asthma. (Disability is a long-term reduction in the ability to participate in children's usual activities, such as attending school or engaging in play, due to a chronic condition.) METHODS: We derived our primary findings from a cross-sectional, descriptive analysis of 62171 children younger than 18 years who were included in the 1994-1995 National Health Interview Survey. MAIN OUTCOME MEASURES: Outcome measures include the presence of disability, degree of disability, restricted activity days, school absence days, and use of hospital and physician services. We also used data from the 1969-1970, 1979-1981, and 1994-1995 National Health Interview Surveys to assess trends in the prevalence of disability due to asthma. RESULTS: A small, but significant, proportion of children, estimated at 1.4% of all US children, experienced some degree of disability due to asthma in 1994-1995. Prevalence of disability due to asthma was higher for adolescents (odds ratio [OR], 1.64), black children (OR, 1.66), males (OR, 1.23), and children from low income (OR, 1.46) and single parent families (OR, 1.37). Disabling asthma resulted in an annual average of 20 restricted activity days, including 10 days lost from school-almost twice the level of illness burden as experienced by children with disabilities due to other types of chronic conditions. Finally, prevalence of disabling asthma, as reported in the National Health Interview Survey, has increased 232% since 1969, the first year that electronic data are available from the survey. In contrast, prevalence of disability due to all other childhood chronic conditions increased by 113% over the same period. CONCLUSIONS: Disabling asthma has profound effects on children. The social costs of asthma are likely to rise in the future if current trends in the prevalence of disabling asthma continue. PMID- 10710031 TI - Antecedents and neonatal consequences of low Apgar scores in preterm newborns: a population study. AB - BACKGROUND: To examine the antenatal and early neonatal correlates of low Apgar scores (<3 and <6 at 1 and 5 minutes) in preterm newborns (23-34 weeks' gestation). OBJECTIVE: The use of Apgar scoring for premature newborns remains widespread, despite controversy regarding its reliability as a measure of morbidity and mortality in the neonatal period. DESIGN: A cohort of 852 preterm newborns born during a 34-month period between 1984 and 1987 was studied. Newborns were stratified into 2 groups by gestational age (23-28 weeks and 29-34 weeks), and data were analyzed, controlling for gestational age in single weeks. SETTING: Two academic and 1 community hospital, which together accounted for 83% of all preterm births in a tri-county area of central New Jersey during the study period. PATIENTS: All premature newborns (birth weight <2000 g and gestational age <35 weeks) born in the participating hospitals during the study period were evaluated. MAIN OUTCOME MEASURES: Antecedents included maternal illness during pregnancy, maternal complications of labor and delivery, and fetal heart rate abnormalities during labor and delivery. Consequences included delivery room resuscitation, abnormal physical findings, diagnoses, and therapeutic interventions in the first 6 to 8 hours of life. RESULTS: Premature newborns with low Apgar scores received more cardiopulmonary resuscitation in the delivery room and in the first 6 to 8 hours of neonatal intensive care. Mortality was significantly increased among newborns with low Apgar scores (54% vs. 26% in the 23- to 28-week stratum, 30% vs 6% in the 29- to 34-week stratum). Newborns with low Apgar scores in the 29- to 34-week stratum more often required intubation, positive pressure ventilation, and umbilical vessel catheterization. Newborns with low Apgar scores had higher rates of bradycardia, pneumothoraces, acidosis, and increased oxygen requirement during the first 6 to 8 hours of life. Maternal illness, complications of labor and delivery;, and fetal heart rate decelerations did not correlate with subsequent Apgar scores of newborns. The presence of severe bradycardia (<90/min) and fetal heart rate accelerations correlated with low Apgar scores in the 29- to 34-week group. CONCLUSION: Low Apgar scores are associated with increased neonatal morbidity and mortality in preterm newborns. Antenatal maternal history, and pregnancy complications are not clearly associated with low Apgar scores. Therefore, the Apgar score is a useful tool in assessing neonatal short-term prognosis and the need for intensive care among preterm newborns. PMID- 10710032 TI - Parental attitudes toward varicella vaccination. The Puget Sound Pediatric Research Network. AB - OBJECTIVES: To evaluate parental health beliefs regarding the varicella vaccine and to identify potential areas for interventions designed to increase immunization against varicella. SETTING: Data were collected in the offices of pediatricians who are members of the Puget Sound Pediatric Research Network, a regional practice-based research group in the Seattle, Wash, area. METHODS: At the time of an office visit, parents were asked to complete a survey on the varicella vaccine. Respondents indicated level of agreement with 10 health belief statements regarding the immunization using a 6-point Likert scale from "completely agree" to "completely disagree"; responses were subsequently transformed to an ordinal scale from 1 to 6, with 6 corresponding to highly positive beliefs. A composite health belief score for each respondent was computed by averaging responses to all statements. Parents also were asked to indicate the level of influence of their child's pediatrician on their decision to use the varicella vaccine. RESULTS: A total of 598 surveys were completed. Generally, parents agreed that the vaccine was worthwhile even if the only benefit was preventing a rare complication. Conversely, the majority of parents disagreed that varicella vaccine was worthwhile if the only benefit was preventing lost time from work, and that the immunization was worthwhile even if immunity was not lifelong. Parents who indicated that their child's pediatrician's opinion significantly influenced their decision to use the vaccine had higher composite health belief scores than those who indicated less influence (median scores, 4.3 and 4.0, respectively; P<.001). CONCLUSIONS: In this sample, parents had more positive health beliefs about the ability of varicella vaccine to prevent rare complications than to save time lost from work. These data also suggest that pediatricians can have an important role in increasing positive health beliefs about the vaccine. These findings may help future interventions to increase the immunization rate against varicella. PMID- 10710033 TI - Radiological case of the month. Hemangioendothelioma of the liver. PMID- 10710034 TI - Picture of the month. Sacrococcygeal teratoma. PMID- 10710035 TI - Pathological case of the month. Disseminated brucellosis. PMID- 10710036 TI - Foreign body aspiration: an unusual complication of antibiotic therapy. PMID- 10710037 TI - Presentation style as important as the message. PMID- 10710038 TI - Screening for serious bacterial infections in young febrile infants. PMID- 10710039 TI - Expandable biliary stents: more questions than answers. PMID- 10710040 TI - Analyzing hospital costs for patients with inflammatory bowel disease. PMID- 10710041 TI - How common is celiac disease in South America? PMID- 10710042 TI - What's good for the goose should be good for the gander--6-MP use in fathers with inflammatory bowel disease. PMID- 10710043 TI - Doing battle with HCV. PMID- 10710044 TI - The myth of informed consent. AB - Informed consent is an integral element of ethical medical practice. However, unlike gastrointestinal endoscopy, the consent process is not standardized. Moreover, informed consent is often viewed as a legal necessity rather than as an expression of patients' autonomy. Elevating informed consent to its rightful place can enhance doctor-patient relationship. PMID- 10710045 TI - Gastrointestinal bleeding in the elderly patient. AB - The elderly patient with acute gastrointestinal bleeding requires a management approach that attends to age-dependent demands, such as functional status, the independent impact of functional status on outcomes such as short term mortality, and the consent process. This review examines issues such as patient triage, resuscitation, diet and medications, anticoagulation, endoscopy and conscious sedation, informed consent in the impaired patient, and discharge planning in light of these age-dependent demands. PMID- 10710046 TI - Peutz-Jeghers syndrome. AB - Peutz-Jeghers syndrome (PJS) is an unusual polyposis syndrome that has enjoyed a rich and somewhat confusing history. Mucocutaneous pigmentation and diffuse gastrointestinal hamartomas are the hallmark features of this autosomal dominant inherited condition. Peutz-Jeghers syndrome is now also recognized as a cancer predisposition syndrome. In this review, we highlight the historical aspects of PJS polyposis with special emphasis on its extraintestinal manifestations, particularly genital tract tumors. A PJS management scheme for clinicians is included. PMID- 10710047 TI - Gastrointestinal miniprobe sonography: the current status. AB - Endoscopic ultrasonography (EUS) represents a major advance in endoscopic imaging. The usefulness and effectiveness of EUS have been established during the past few years. However, endosonography using dedicated echoendoscopes (7.5/12 MHz) has some serious drawbacks, as follows: 1) Combining endoscopy and ultrasonography in one instrument increases the diameter of such echoendoscopes (12-13 mm); 2) Because of the large diameter, complete passage of severe strictures is often not possible and, for examination of the pancreatobiliary duct system, is not feasible at all; 3) Image quality and resolution for small lesions is not always satisfactory; and 4) Conventional endosonography requires a second examination separate from the previous routine endoscopy. Recently developed ultrasonographic miniprobes (diameters about 2 mm; frequencies 12-20 MHz) can be passed through the working channel of standard endoscopes to provide high frequency ultrasound images. These miniprobes might overcome some of the above-mentioned drawbacks and contribute to patients' security and convenience. Moreover, in various diseases of the GI tract and the pancreatobiliary duct system, diagnostic accuracy of miniprobe ultrasonography has been shown to be even superior to that of EUS. In summary, miniprobe ultrasonography seems to be a promising tool in the armamentarium of gastroenterological diagnostics. PMID- 10710048 TI - Intestinal and pancreatic metaplasia at the esophagogastric junction in patients without Barrett's esophagus. AB - OBJECTIVES: A distinctive type of columnar epithelium with intestinal metaplasia is considered diagnostic for Barrett's esophagus. The neoplastic potential of pancreatic metaplasia at the esophagogastric junction is unknown. The aims of the present study were: 1) to characterize both forms of metaplasia at the esophagogastric junction, and to estimate their prevalence; 2) to investigate c erbB-2 expression and K-ras mutations in pancreatic metaplasia; and 3) to study the relationship between metaplasia, inflammatory changes in the cardiac mucosa, and presence of H. pylori. METHODS: A total of 76 esophagogastrectomy specimens of patients with a normally located squamocolumnar junction, were investigated immunohistochemically. K-ras mutations were evaluated using PCR. RESULTS: Intestinal metaplasia in the cardia was found in 12% of patients: six complete type, and three incomplete-type. Pancreatic metaplasia was demonstrated in 14% of patients, and neither c-erbB-2 expression nor K-ras mutations were found. Intestinal and pancreatic metaplasia were associated with mucosal inflammation. In contrast to generalized gastritis, isolated "carditis" was not associated with H. pylori infection. CONCLUSIONS: When intestinal metaplasia occurs in a biopsy from the esophagogastric junction, it is not necessarily a marker for Barrett's esophagus. No indication was found that pancreatic metaplasia has neoplastic potential. Both forms of metaplasia reflect mucosal inflammation. Carditis may be a distinct inflammatory condition of the gastric mucosa that is not related to H. pylori infection. PMID- 10710049 TI - Oral and intravenous dosage forms of pantoprazole are equivalent in their ability to suppress gastric acid secretion in patients with gastroesophageal reflux disease. AB - OBJECTIVE: The aim of this study was to assess the ability of pantoprazole to maintain gastric acid suppression in patients with gastroesophageal reflux disease who are switched from an oral (p.o.) to an intravenous (i.v.) dosage form. METHODS: A total of 65 patients with gastroesophageal reflux disease were administered either 40 or 20 mg of p.o. pantoprazole daily for 10 days, then were switched to either a matching dose of i.v. pantoprazole or to placebo for 7 days. Acid output (basal and maximal) was measured at the end of the p.o. treatment period and on the first and last days of i.v. therapy. In the primary efficacy analysis, the acid output values at the end of the p.o. pantoprazole treatment were compared with those at the end of the i.v. treatment. Safety was monitored by periodic vital sign measurements, clinical laboratory evaluations, ophthalmic examinations, electrocardiograms, and reports of adverse events. The data were tested by an analysis of covariance and by Wilcoxon signed rank and t tests. RESULTS: Maximal acid output (mean +/- SD) in the 40 mg and 20 mg pantoprazole group after p.o. treatment was 6.5 +/- 5.6 mEq/h and 14.5 +/- 15.5 mEq/h, respectively; whereas, at the end of the i.v. treatment period, the values were 6.6 +/- 6.3 mEq/h and 11.1 +/- 10.2 mEq/h, respectively. In patients given i.v. placebo, acid output was significantly (p < 0.05) increased to 29.2 +/- 13.0 mEq/h by day 7. Both p.o. and i.v. pantoprazole dosage forms had similar favorable safety and tolerability profiles. CONCLUSIONS: The p.o. and i.v. formulations of pantoprazole (40 and 20 mg) are equivalent in their ability to suppress gastric acid output. The i.v. form of pantoprazole offers an alternative for gastroesophageal reflux disease patients who are unable to take the p.o. formulation. PMID- 10710050 TI - Upper gastrointestinal toxicity of alendronate. AB - OBJECTIVE: Alendronate is rapidly gaining widespread use in the treatment of osteoporosis. However, recent postmarketing surveys and endoscopic studies suggest that its use may be associated with significant predictable esophageal and gastric mucosal toxicity, similar to that of aspirin and nonsteroidal anti inflammatory drugs. Because treatment of osteoporosis may be needed in as many as 30% of all postmenopausal women, and considering that alendronate could be used in all postmenopausal women as prevention, definition of potential mucosal toxicity is crucial. Our aim was to study the upper gastrointestinal toxicity of alendronate in an age-appropriate female population using a clinically applicable dose (10 mg/day) to determine whether it causes predictable damage in the proximal gastrointestinal mucosa in a fashion similar to that seen with aspirin and nonsteroidal anti-inflammatory drugs. METHODS: We conducted a double-blind, randomized, placebo-controlled trial in 32 healthy female volunteers between the ages of 40 and 65 yr recruited by newspaper advertisement. Endoscopic mucosal abnormalities in the esophagus, stomach, and duodenum both before and after 1 month of treatment were scored and compared using validated endoscopic grading systems. Symptom scores before and after treatment were determined. Noninvasive measurements of gastrointestinal permeability were obtained before, during, and after treatment using sucrose and mannitol/lactulose urinary excretions. RESULTS: Endoscopic scores before and after treatment with alendronate were not significantly different. Similarly, mean symptom scores in the alendronate group did not change significantly after treatment. There were no significant mucosal permeability changes in the stomach or small intestine after treatment. CONCLUSION: Alendronate does not cause predictable esophageal, gastric, or duodenal mucosal damage when used as directed. PMID- 10710051 TI - A single drug for Helicobacter pylori infection: first results with a new bismuth triple monocapsule. AB - OBJECTIVE: In this pilot study we investigated the efficacy and tolerability of a new monocapsule that contains a bismuth compound, tetracycline, and metronidazole. If proven to be effective, this monotherapy would turn the well accepted multidrug regimen of standard bismuth-based triple therapy into an easy and more patient-friendly regimen. It can be used in patients allergic to penicillin. METHODS: A total of 53 consecutive H. pylori-infected patients (30 with proven ulcer disease, 23 with gastritis only) from a single center were prescribed two monocapsules q.i.d. after the three meals and after an evening snack during 10 days. Each capsule contained 60 mg colloidal bismuth subcitrate (as Bi2O3 equivalent), 125 mg tetracycline, and 125 mg metronidazole. Repeat endoscopy with biopsies for urease test, Giemsa stain, and culture was carried out > or =5 wk later. Side effect data were collected. RESULTS: One patient was lost to follow-up, two failed to respond, and 50 were cured. The intention-to treat cure rate was 50 of 53 (94.4%, 95% CI 88.1-100%). Antibiotic sensitivity was available from 51 isolates. The cure rate in the metronidazole sensitive strains was 44 of 45 (97.8%, 95% CI: 93.5-100%), whereas it was four of five in the resistant strains. The regimen was well tolerated, with only two drop-outs (4%) because of side effects. CONCLUSIONS: The new monocapsule is an inexpensive, well tolerated, and patient-friendly formulation of a bismuth based triple therapy. A 10-day course with this multidrug capsule reached a very high cure rate in metronidazole-sensitive strains. The number of cases with resistant strains was insufficient to allow firm conclusions about its efficacy in case of resistance. The results are in agreement with previous data with bismuth triple therapy using separate drugs. From the high cure rate, we can conclude that the new capsule dissolves adequately, with proper delivery of its ingredients at the site of action. PMID- 10710052 TI - Validation of the 13C-urea breath test for the diagnosis of Helicobacter pylori infection in children: a multicenter study. AB - OBJECTIVE: The 13C-urea breath test (13C-UBT) is a safe, noninvasive, and accurate test for the detection of Helicobacter pylori (H. pylori) infection in adults. The aim of this study was to evaluate sensitivity and specificity of 13C UBT in children using different types of test meal, doses of 13C-urea and breath sampling intervals. As yet, a validated, standardized 13C-UBT protocol for children has not been formulated. METHODS: 13C-UBT was performed in 115 children and repeated within 3 days, modifying the test meal or the dose of 13C-urea. H. pylori status was assessed by histology and rapid urease test. 13C-UBT was performed using 100 mg or 50 mg of 13C-urea and a fatty test meal (100 FA; 50 FA), 50 mg of 13C-urea, and a carbohydrate test meal (50 CA). Breath samples were collected every 10 min for 60 min. RESULTS: The 13C-UBT in children was highly sensitive and specific with all three protocols used. The best combination of sensitivity (97.92%) and specificity (97.96%) was obtained with Protocol 50 FA at 30 min with a cut-off of 3.5 per mil. CONCLUSIONS: The 13C-UBT is an accurate test for the detection of H. pylori infection also in children. Administration of 50 mg of 13C-urea, a fatty test meal, and breath sampling at 30 min appears to be the most convenient protocol. PMID- 10710053 TI - Is upper gastrointestinal radiography a cost-effective alternative to a Helicobacter pylori "test and treat" strategy for patients with suspected peptic ulcer disease? AB - OBJECTIVE: Current clinical consensus supports an initial Helicobacter pylori (HP) "test and treat" approach when compared to immediate endoscopy for patients with suspected peptic ulcer disease. Alternative diagnostic approaches that incorporate upper GI radiography (UGI) have not been previously evaluated. We sought to determine the cost effectiveness of UGI compared to a HP test and treat strategy, incorporating recent data addressing the reduced prevalence of HP, lower cost of diagnostic interventions, and reduced attribution of PUD to HP. METHODS: Using decision analysis, three diagnostic and treatment strategies were evaluated: 1) Test and Treat--initial HP serology, treat patients who test positive with HP eradication and antiulcer therapy; 2) Initial UGI series--treat all patients with documented ulcer disease with HP eradication and antiulcer therapy; and 3) Initial UGI series, HP serology if ulcer present--treat ulcer and HP based on diagnostic test results. RESULTS: The estimated cost per ulcer cured for each strategy were as follows: test and treat, $3,025; initial UGI, $3,690; and UGI with serology, $3,790. The estimated cost per patient treatment were: test and treat, $498; initial UGI, $610; and UGI with serology, $620. When UGI reimbursement was decreased to less than $50, the UGI strategies yielded a lower cost per patient treated than the test and treat strategy. CONCLUSION: At the current level of reimbursement, UGI should not be considered a cost-effective alternative to the HP test and treat strategy for the initial evaluation of patients with suspected peptic ulcer disease. PMID- 10710054 TI - Association of Helicobacter pylori genotype with gastroesophageal reflux disease and other upper gastrointestinal diseases. AB - OBJECTIVE: Helicobacter pylori (H. pylori) is a recognized pathogen, but it may also have a protective effect for gastroesophageal reflux disease (GERD). We compared the prevalence of potential virulence factors (cagA, cagE, vacA genotypes) in GERD to other upper gastrointestinal diseases and controls. METHODS: A total of 405 patients underwent gastroscopy with H. pylori isolation and serum testing. Patient diagnostic subgroups were prospectively defined. Genotypes were determined by amplification using polymerase chain reaction. CagA antibodies were determined by western blot, enzyme-linked immunosorbent, and flow microsphere immunofluorescent assays. RESULTS: Patients were grouped as follows: nonulcer dyspepsia (26%), GERD (20%), gastric ulcer (17%), duodenal ulcer (12%), gastric cancer (6%), or controls (19%). The cagA gene was present in 94-97% of subjects in all categories, but the cagA antibody was less prevalent in nonulcer dyspepsia (69%, 95% CI: 48-86%, p = 0.02) and GERD (69%, CI: 39-91%, p < 0.05) than in those with gastroduodenal pathology including gastric ulcer, duodenal ulcer, and gastric cancer (92%, CI: 81-98%). The cagE gene and vacA S1 genotype were more frequent in patients with gastroduodenal pathology (p < 0.01). GERD was associated with a significantly lower rate of vacA S1 genotype than controls (29% (CI: 10-56%) versus 80% (CI: 59-93%), p < 0.01). The vacA S1 genotype was associated with the presence of cagA antibodies. CONCLUSIONS: The cagE and vacA S1 genotypes are more prevalent in patients with peptic ulcer or gastric cancer, suggesting a potential function in virulence for these genes. However, the vacA S1 genotype was also more prevalent in controls than GERD, suggesting a potential protective effect against GERD. PMID- 10710055 TI - A comparison of Ultraflex Diamond stents and Wallstents for palliation of distal malignant biliary strictures. AB - OBJECTIVE: The objective of this study was to compare the effectiveness of the Ultraflex Diamond stent and the Wallstent for the drainage of distal malignant biliary strictures. METHODS: The results obtained in 23 consecutive patients in whom the insertion of a Ultraflex Diamond stent had been attempted were compared with those obtained in 23 patients matched for age, gender, serum bilirubin, and diagnosis who had been treated with Wallstents. RESULTS: Biliary drainage was obtained in 100% of cases. More than one stent was required in 4% and 12% of patients treated with Ultraflex Diamond stents and Wallstents, respectively (p > 0.05). The first stent inserted did not provide adequate biliary drainage in four patients, because of the impaction of the proximal end of Wallstents into the bile duct wall (n = 2) and obstruction of the stent lumen by tumor tissue (one in each group). Procedure-related morbidity and mortality were 4%. Patients were followed-up for a mean of 228 days (range, 1 to 1262 days). During follow-up, bile duct obstruction relapsed in 5/22 and 6/21 patients treated with Ultraflex Diamond stents and Wallstents, respectively. Life table analysis of bile duct patency was similar with both stent models. CONCLUSIONS: Ultraflex Diamond stents are easy to insert and provide a high success rate of biliary drainage with minimal complication. Although long-term patency rates obtained with this stent were similar to those observed with Wallstents, no firm conclusion can be drawn in this regard due to the relatively small number of patients studied. PMID- 10710056 TI - Direct hospital costs for patients with inflammatory bowel disease in a Canadian tertiary care university hospital. AB - OBJECTIVE: We set out to determine the direct costs of hospitalizations of patients with Crohn's disease and ulcerative colitis admitted to a university affiliated tertiary care hospital and to contrast the costs of medical versus surgical inpatient care, Crohn's disease versus ulcerative colitis, and to identify dominant components of inpatient costs. METHODS: We used a patient specific case costing system at Saint Boniface General Hospital, Winnipeg, Manitoba, for fiscal years 1994 and 1995. We extracted all inpatients whose hospital discharge abstracts included ICD-9-CM codes 555 (Crohn's disease) and 556 (ulcerative colitis) among the top eight discharge diagnoses, and performed a chart review on all cases to ensure that the hospitalization was for inflammatory bowel disease and the diagnoses were accurate. We analyzed cases based on their disease diagnosis, primary mode of therapy associated with the hospitalization (medical vs surgical), and their major diagnosis-related group (DRG). This study evaluated direct patient care costs only and costs are expressed in Canadian dollars. RESULTS: Of 362 hospital admissions, 325 were eligible and of these admissions 275 belonged to the digestive system DRGs. Seventy-one (37%) were admitted more than once during the 2 yr of the study, accounting for 202 (62%) of the total number of admissions. The mean cost per admission of all cases of Crohn's disease was $3,149 (95% confidence interval [CI], $2,665-$3,634) and for ulcerative colitis was $3,726 (95% CI $3,008-$4,445). Surgical therapy cases accounted for 49.8% of all admissions, 57.8% of all hospital days, and 60.5% of all costs. Patients treated surgically had more costly hospitalizations than those treated medically, particularly when analyzing only nontotal parenteral nutrition (TPN) cases. Surgical treatment admissions were significantly more costly for ulcerative colitis digestive DRG admissions than Crohn's disease. The nondigestive DRG admissions were more costly than the digestive DRGs in all categories although this was only statistically different among medically treated Crohn's disease. Patients treated medically were similarly costly whether they had Crohn's disease or ulcerative colitis. There was no significant difference in cost per admission among cases admitted multiple times, compared with those admitted only once. TPN cases accounted for 9.5% of cases but 27.1% of costs. TPN associated hospitalizations were more costly than non-TPN-use hospitalizations but these costs were primarily driven by duration of stay rather than TPN use itself. For all cases, the top five cost categories in descending order were nursing unit bed-days, drugs and pharmacy, diagnostic lab tests, operating room, and diagnostic imaging and endoscopy. CONCLUSIONS: Using our system we could determine direct costs for inpatients with inflammatory bowel disease and the factors that determined increased costs. Medical therapy admissions were similarly costly between Crohn's disease and ulcerative colitis; however, surgical therapy admissions were costlier among ulcerative colitis patients. Admissions for nondigestive DRGs were more costly than those for digestive DRGs. TPN use identified a sicker group of patients who remained in the hospital longer than nonusers and, not surprisingly, these were the costliest patients. PMID- 10710057 TI - Outcome of pregnancies when fathers are treated with 6-mercaptopurine for inflammatory bowel disease. AB - OBJECTIVE: The outcomes of pregnancies after maternal use of 6-mercaptopurine (6 MP) for inflammatory bowel disease (IBD) during pregnancy have been reported, but data are lacking for outcomes when the fathers use this drug. METHODS: Subjects were male patients with IBD seen at one center between 1970 and 1997. Patients and their wives were interviewed. Group 1 comprised pregnancies fathered by men who were taking 6-MP. This group was further subdivided into those conceived within 3 months of 6-MP use and those conceived at least 3 months after 6-MP was stopped. Group 2 comprised pregnancies fathered by men with IBD, similar in characteristics to group 1, who had not taken 6-MP before fertilization. Information was collected regarding the fathers, the mothers, and the pregnancies, as well as the health of the children, in a historical cohort study. RESULTS: There were 50 pregnancies in group 1 (13 in 1A and 37 in 1B) and 90 pregnancies in group 2. Four of the 13 pregnancies in group 1A were associated with complications. There were two spontaneous abortions, and two congenital anomalies including a missing thumb in one and acrania with multiple digital and limb abnormalities in the other. Risk of complications was significantly increased when compared with group 1B (p < 0.013) and group 2 (p < 0.002). CONCLUSION: The incidence of pregnancy-related complications was significantly increased when the fathers used 6-MP within 3 months of conception. PMID- 10710058 TI - Prevalence of celiac disease among blood donors in Brazil. AB - OBJECTIVE: There are no studies on the prevalence of celiac disease (CD) in either Brazil or, as far as we know, South America. The aim of this study was to determine the prevalence of CD in healthy blood donors in the city of Brasilia, Brazil. METHODS: Sera were obtained, independently of age and gender, from an unselected group of 2045 blood donors attending the Hematological Center of Brasilia. An IgG antigliadin antibody (AGA) test was used as a first-level screening step, followed by IgA-AGA test, serum IgA antiendomysium (EMA), and total serum IgA determination performed in all sera showing abnormally high IgG AGA results. Jejunal biopsy was suggested for all subjects showing at least one of the following: IgA-EMA positivity; IgG-AGA and IgA-AGA positivity; IgG-AGA positivity and selective IgA deficiency. AGA was determined by an enzyme-linked immunosorbent assay (ELISA) technique and IgA-EMA was ascertained by indirect immunofluorescence on cryostat sections of monkey esophagus. Jejunal mucosa samples were obtained with a Watson capsule. RESULTS: Sixty-two (3.03%) blood donors had IgG-AGA above the cut-off values. Fifty-eight individuals showed isolated high values of IgG-AGA, whereas four had simultaneously increased IgG and IgA-AGA. Three patients had positive IgA-EMA test (one with both IgG- and IgA AGA and two with only IgG-AGA) and underwent jejunal biopsies that disclosed complete villous atrophy associated with an increased number of intraepithelial lymphocytes and hypertrophic criptae. In this study group, the prevalence of biopsy-proven celiac disease was 1.47 +/- 1.66 in 1000 subjects. CONCLUSIONS: We found a prevalence of undiagnosed CD of 1:681 among apparently healthy blood donors. These preliminary results support the view that CD is not a rare disease in Brazil. PMID- 10710059 TI - Epidemiology of ulcerative colitis in Israel: a survey of Israeli kibbutz settlements. AB - OBJECTIVE: The incidence of ulcerative colitis ranges from 3 to 15 cases per 100,000 persons per year with a prevalence of 50-80 cases per 100,000, and the disease is 3-5 times more common among Jews. In Israel, Ashkenazi Jews have a higher incidence than Sephardi Jews, but a lower incidence than Ashkenazi Jews in the United States or Northern Europe. The aim of this study was to examine the prevalence, mean annual incidence, and clinical patterns of ulcerative colitis in a stable population of communal settlements (kibbutz). METHODS: We repeated a community-based survey in 124,400 kibbutz residents, 10 yr after our first study. This population represents 2.5% of the Jewish population of Israel. All ulcerative colitis patients were located by contacting the kibbutz clinics of the 269 kibbutz settlements (100% compliance). Data were updated to December 31st, 1997, which was designated the point prevalence date, and included information on gender, age, origin, education, profession, extent of the inflammatory process, clinical spectrum of the disease, therapy, complications of the disease, and treatment. The mean annual incidence for the 10 yr was calculated from the prevalence data. Only cases with a definite diagnosis of ulcerative colitis made in a recognized gastroenterology unit were accepted into the study. RESULTS: There were 208 confirmed cases of ulcerative colitis disease, 113 women and 95 men (female:male ratio = 1.19). The prevalence rate rose from 121.0/100,000 in 1987 to 167.2/100,000 in 1997 (p < 0.001). The prevalence rates were higher in women than men. Prevalence was highest in Israeli-born members in 1987 but in European/American-born members in 1997. The average annual incidence rate for the 10-yr period was 5.04/100,000/yr. In 1987, 146 cases of ulcerative colitis were collected. During 10 yr of surveillance 17 patients left the kibbutz, 12 died, and 62 new cases were added. The mean age at presentation of the disease was lower in 1987 than in 1997, 46.14 +/- 11.10 and 51.43 +/- 16.67 yr, respectively. Prevalence was highest in men with >16 yr and in women with 9-10 yr of education, 259.3 and 242.9/100,000, respectively. CONCLUSIONS: The prevalence of ulcerative colitis in this Israeli population increased and has reached the upper range for European and American populations. The mean annual incidence rate of ulcerative colitis is in the lower range of that reported for the Western countries. PMID- 10710060 TI - Increased urinary N-telopeptide cross-linked type 1 collagen predicts bone loss in patients with inflammatory bowel disease. AB - OBJECTIVE: Reduced bone mineral density (BMD) is common in patients with inflammatory bowel disease (IBD), but the factors associated with its longitudinal rate of change have not been established. We prospectively assessed the rate of change in BMD, and its association with biochemical markers of bone turnover. METHODS: Twenty-two patients with Crohn's disease and 14 ulcerative colitis patients age 37.1 +/- 11.6 yr were followed for 2 yr. Lumbar spine (L2 L4) and femoral neck BMD were measured by dual x-ray absorptiometry at baseline and 24 months. Bone-specific alkaline phosphatase, osteocalcin, urinary N telopeptide crosslinked type 1 collagen (NTx), parathyroid hormone, and 25 hydroxyvitamin-D were determined at baseline. RESULTS: At baseline, 59% of Crohn's patients and 43% of ulcerative colitis patients were osteoporotic, with spine or femoral neck BMD T-score < -2.5. Spine BMD, and spine and femoral neck T scores were lower and disease duration was longer in nine patients with ileal resection compared with nonoperated patients (0.84 +/- 0.15 g/cm2 vs 0.96 +/- 0.11 g/cm2, -3.0 +/- 1.5 vs -1.7 +/- 1.3, -3.2 +/- 1.5 vs -2.2 +/- 1.0, respectively; all p < 0.05). At 24 months, 13/36 (36%) and 14/36 (39%) patients experienced spinal and femoral neck bone loss, respectively, with mean annual percent BMD changes of -2.0% and -1.5%, respectively. NTx, a bone resorption marker, inversely correlated with spinal BMD rate of change (r = -0.4, p < 0.05). Using quartiles analysis, patients with the highest NTx (Q4) experienced the greatest decrease in spine BMD compared with patients with the lowest NTx (Q1). CONCLUSIONS: Spine and femoral neck bone loss continues over time in more than one-third of IBD patients. Increased NTx level predicts spinal bone loss in IBD patients. PMID- 10710061 TI - Focal gastric inflammatory infiltrates in inflammatory bowel diseases: prevalence, immunohistochemical characteristics, and diagnostic role. AB - OBJECTIVES: To date, few studies have evaluated gastric histology in patients with inflammatory bowel disease (IBD). The aim of this prospective controlled study was to establish the frequency of focal gastritis in Crohn's disease (CD) and ulcerative colitis (UC) patients, as well as to evaluate its immunohistochemical characteristics and clinicoanatomical determinants. METHODS: We evaluated 141 consecutive patients with known CD of the large and/or small bowel, 79 patients with UC, and 141 CD- and UC-free controls; all underwent upper gastrointestinal (GI) endoscopy and 13C urea-breath test. Biopsy specimens taken from the antrum, angulus, and gastric body were evaluated by histology and immunohistochemistry. A series of variables, including CD activity index, duration, extent and location of disease, intestinal resection, number of recurrences, and previous and current medical therapy, as well as the presence of dyspeptic symptoms and mucosal lesions at endoscopy, were determined in all CD patients and correlated with the presence or absence of focal gastritis. RESULTS: Helicobacter pylori-associated gastritis was found in 47 patients with CD (33%), in 37 patients with UC (47%), and in 60% of CD-/UC-free controls (p < 0.01). In H. pylori-negative CD patients focal gastritis was found in 43% of cases (40/94), compared with 12% (5/42) of UC patients and 19% (11/57) of controls (p < 0.05). Specificity and positive predictive value of focal gastritis in CD were 84% and 71%, respectively. It was characterized by a focal perifoveolar or periglandular lymphomonocytic infiltrate, with CD8+/CD4+ cells predominant both in CD and UC patients. There were no significant correlations between the occurrence of focal gastritis and any clinicoanatomical CD features. CONCLUSIONS: Focal gastritis is relatively common in CD patients although it is not exclusive to this condition. Its recognition could be useful in the diagnostic workup of any patient with suspected or indeterminate inflammatory bowel disease, as it makes a diagnosis of CD more likely. PMID- 10710062 TI - Disappearance of endomysial antibodies in treated celiac disease does not indicate histological recovery. AB - OBJECTIVE: Although serum IgA-class endomysial antibody (EmA) has high sensitivity for villous atrophy (VA) in patients with untreated celiac disease, few studies have attempted to correlate EmA seroconversion with histological recovery after starting a gluten-free diet. We prospectively studied changes in EmA status and in duodenal histology of seropositive patients after dietary treatment. METHODS: Patients with VA and EmA had repeat EmA testing at 3, 6, and 12 months after starting gluten-free diet, plus assessment of dietary compliance by dietitians and follow-up duodenal biopsy at 12 months. VA before and after treatment was classified as partial (P), subtotal (ST), and total (T). RESULTS: Of 77 patients with newly diagnosed VA and without IgA deficiency, 62 (81%) had EmA: 46 of 57 (81%) with T or STVA and 16 of 20 (80%) with PVA. Of 53 initially EmA-positive patients who completed study criteria, EmA was undetectable in 31 patients (58%) after 3 months' diet, in 40 (75%) after 6 months, and in 46 (87%) after 12 months. However, only 21 patients (40%), all seronegative by 12 months, had complete villous recovery. Only three (33%) of 10 patients with persisting ST or TVA and two (9%) of 22 with PVA remained EmA positive. Four of the five patients with persisting EmA had poor dietary compliance. CONCLUSIONS: EmA is a poor predictor of persisting VA after patients have started gluten-free diet, although it may be of value in monitoring dietary compliance. Although there are no clear guidelines regarding the need for follow-up biopsy, EmA seroconversion cannot substitute. The apparent association between dietary compliance and seroconversion suggests that gluten intake may determine whether untreated celiac patients are EmA positive or negative for a given degree of small bowel damage. PMID- 10710063 TI - Coloanal motor coordination in association with high-amplitude colonic contractions after pharmacological stimulation. AB - OBJECTIVE: The rectoanal inhibitory reflex facilitates defecation by relaxation of the internal anal sphincter during rectal distention by gas or stool. Defecation is sometimes preceded by high-amplitude propagated contractions (HAPCs). Our objective was to seek evidence for motor coordination between human colonic and anal sphincter functions. METHODS: As part of a study of alpha2 modulation of colonic and anal motor functions in 32 healthy volunteers, we studied the relationship between high HAPCs and anal sphincter pressure with colonic manometry, barostat, and a Dent sleeve in the anal canal. RESULTS: Twenty two HAPCs were observed; in 19/22 HAPCs there was optimal positioning of the Dent sleeve to assess the anal sphincter. Eighteen of 19 HAPCs occurred postprandially; 14 HAPCs occurred after administration of yohimbine, three after clonidine, and one before any drug administration. Seven followed experimental balloon distention. Anal sphincter relaxation occurred (14 +/- 4 s) before the recorded onset of HAPC in the descending colon and 88 +/- 7 s before the arrival of the HAPC in the rectum. After or during the HAPCs, anal sphincter pressure decreased by 40 +/- 4% and increased by 56 +/- 8% in the postrelaxation phase. CONCLUSIONS: The close temporal association between anal sphincter relaxation and onset of HAPC in the descending colon suggests a coloanal reflex that may facilitate defecation during mass movements independently of the rectoanal inhibitory reflex. PMID- 10710064 TI - Magnetic resonance imaging with ferumoxil, a negative superparamagnetic oral contrast agent, in the evaluation of ulcerative colitis. AB - OBJECTIVE: The introduction of new oral contrast agents that enhance image quality has increased the importance of magnetic resonance imaging (MRI) in the management of ulcerative colitis. The aim of our study was to investigate the usefulness of a new negative superparamagnetic oral contrast (ferumoxil) alone or in association with gadolinium i.v. in the assessment of the disease. METHODS: Twenty-eight patients with clinically active ulcerative colitis and 10 control subjects entered the study. In each patient a clinical, endoscopic, histological, and MRI evaluation was performed. In particular, in 14 patients affected by ulcerative colitis (group A) and in five controls, magnetic resonance images were acquired 1 h after the oral administration of 900 ml of ferumoxil, while the remaining 14 patients (group B) and five controls were submitted to double contrast MRI (ferumoxil and gadolinium). In both groups, wall thickness, length of affected bowel segments, and, in group B, also percent contrast enhancement were calculated. RESULTS: The comparison of endoscopic and MRI extent of disease was statistically significant. Wall thickness and, in group B, also percent contrast enhancement were significantly correlated with clinical and endoscopic activities. In each group wall thickness was significantly different in the activity phases of the disease. CONCLUSIONS: MRI with negative superparamagnetic oral contrast is comparable to endoscopy in the assessment of ulcerative colitis. The double-contrast imaging does not provide more information than single oral contrast, so we concluded that the latter is preferable in the follow-up of the disease and in patients unable or with a poor compliance to undergo endoscopy. PMID- 10710065 TI - A meta-analysis of antimycobacterial therapy for Crohn's disease. AB - OBJECTIVE: Various therapies have been studied for the treatment of Crohn's disease, including antimycobacterial therapy. Meta-analysis was used to evaluate the effect of antimycobacterial therapy in patients with Crohn's disease. METHODS: Randomized, controlled trials comparing antimycobacterial therapy with placebo were identified. Key outcome data were abstracted and the results were pooled to yield odds ratios for maintenance of remission in treated versus control groups. RESULTS: A total of eight randomized trials were identified. Six trials were fully published and were included in the primary analysis. Two trials used antimycobacterial therapy in combination with corticosteroids to induce remission in patients with active Crohn's disease, followed by maintenance therapy with antimycobacterial agents. In these trials, control patients received corticosteroids to induce remission but no antimycobacterial therapy. Pooling of these trials yielded an odds ratio of maintenance of remission in treatment versus control of 3.37 (95% confidence interval [CI], 1.38-8.24) in favor of antimycobacterial therapy. The remaining four trials used antimycobacterial therapy combined with standard therapy in patients with Crohn's disease. In these trials, control patients received standard therapy alone. Pooling of these trials yielded an odds ratio of maintenance of remission in treatment versus control of 0.69 (95% CI, 0.39-1.21) in favor of standard therapy. CONCLUSIONS: These results suggest that antimycobacterial therapy is effective in maintaining remission in patients with Crohn's disease after a course of corticosteroids combined with antimycobacterial therapy to induce remission. Treatment of Crohn's disease with antimycobacterial therapy does not seem to be effective without a course of corticosteroids to induce remission. Because of the small number of studies included in this meta-analysis, the results should be interpreted with caution. PMID- 10710066 TI - The use of induced sputum in the assessment of pulmonary involvement in Crohn's disease. AB - OBJECTIVE: Our aim was to evaluate lung involvement in Crohn's disease (CRD) patients by induced sputum (IS). Extraintestinal manifestations are frequent in CRD, but lung involvement is rare. Induced sputum is a reliable noninvasive method of investigating the pathogenesis, pathophysiology, and treatment of lung disease. METHODS: Twenty-four CRD patients and nine control subjects (all nonsmokers) without respiratory symptoms were tested. Sputum was induced by 20' inhalation of 3.5% saline using ultrasonic nebulizer. Samples were studied by differential counts of 200 cells on cytopreps stained by Giemsa. T-lymphocyte subset analyses were done by FACS using three monoclonal antibodies: CD3 = total T cells, CD4 = T helper cells, and CD8 = T suppressor-cytotoxic cells. CD4/CD8 >2.5 was considered abnormal. RESULTS: Four patients did not produce sputum. Of the remaining 20 patients, seven had active CRD and 13 were in remission. They were divided into two groups: Group A patients had abnormal CD4/CD8 ratio of 6.7 +/- 2.5; Group B (seven patients) had normal CD4/CD8 ratio of 1.7 +/- 0.52 (p = 0.0001). The differential counts of IS samples were similar in both groups, but lymphocyte count was significantly higher in CRD patients than in the control group (13.2 +/- 11.2 vs 4.8 +/- 3.6, p = 0.036). There were no differences in spirometry, treatment, extent, or activity of CRD. CONCLUSION: Using a simple noninvasive method, we found that among CRD patients without respiratory symptoms there was a high (65%) incidence of lung involvement. PMID- 10710067 TI - Polymeric versus elemental diet as primary treatment in active Crohn's disease: a randomized, double-blind trial. AB - OBJECTIVE: Enteral feeding is now an established primary therapy for active Crohn's disease. This first-double blind randomized trial was designed to compare the therapeutic efficacy of a polymeric diet (PD) with an elemental diet (ED). METHODS: Patients with active Crohn's disease (Crohn's disease activity index [CDAI] > 150, increased bowel uptake of Tc-HMPAO-labeled leukocytes, and abnormal C-reactive protein [CRP]), were randomized to receive either an ED or a PD. The two preparations were identical except for the nitrogen source, which was amino acid based in ED and intact protein in PD. Enteral feeding was considered successful if clinical remission was achieved as defined by a final CDAI of < or = 150, a reduction in the CDAI by at least 100 points from baseline level, and a normal CRP. RESULTS: Twenty-one patients were enrolled of whom 11 were randomized to PD and 10 to ED. The two groups were comparable at entry. Clinical remission was obtained in eight (80%) patients receiving ED and six (55%) patients receiving PD, p = 0.1. The treatment failed in three and two patients in the PD and ED groups, respectively. Another two patients were intolerant to the feed (PD). Reduction in the CDAI after treatment with ED (359 +/- 67 to 112 +/- 19) was similar to that seen with PD (303 +/- 27 to 97 +/- 11). Similar changes in the CRP were also observed (16 +/- 5 to 4 +/- 1.6) and (62 +/- 20 to 9 +/- 6), respectively. Overall, enteral feeding was successful in 14 patients (63%). CONCLUSIONS: Enteral nutrition is effective in treatment of active Crohn's disease. Differences in nitrogen sources of enteral feeds are not relevant to their therapeutic efficacy, as polymeric and elemental diets are equally effective. PMID- 10710068 TI - Epidemiology of hepatitis C virus infection in American veterans. AB - OBJECTIVE: This study reports the findings of hepatitis C virus (HCV) infection in a large Department of Veterans Affairs Health Care System in suburban Northern California. METHODS: All veterans who had anti-HCV (EIA II) tested during a 6-yr period (7/92 to 6/98) were included in this study. To estimate the seroprevalence of anti-HCV among our population, 126 consecutive bloodborne pathogen exposure accidents were studied. RESULTS: Of 8558 veterans tested for anti-HCV (EIA II), 2985 (35%) veterans were positive with a mean age of 48.4 yr (range, 28-89 yr). Sixty percent were between the age of 41 and 50 yr. Risk factors for HCV infection identified in 409 consecutive veterans were intravenous drug abuse (81%), unknown (11%), blood transfusion (3%), sexual/household contact (2%), transfusion and intravenous drug use (2%), and tattoo (1%). Of 215 consecutive anti-HCV-positive veterans whose sera were tested by polymerase chain reaction, 96% were viremic. The most common HCV genotypes were 1a (50.5%), 1b (22.8%), 3a (12.1%), 2b (9.7%), 2a (1.9%), undetermined (1.9%), and mixed infection (1%). Veterans infected with genotype 1b were significantly older. Among 126 consecutive bloodborne pathogen exposure accidents, hepatitis C serology was available for 72 index veterans involved in the accidents and 18% were positive. CONCLUSIONS: We found the epidemiology of hepatitis C infection was different in the veteran population when compared to other published data on nonveterans. Hepatitis C infection was much more common among veteran, within a very narrow age distribution and intravenous drug use was the major risk factor. PMID- 10710069 TI - The role of telomerase activity in hepatocellular carcinoma. AB - OBJECTIVE: The aim of this study was to clarify the role of telomerase activity in hepatocellular carcinoma (HCC). METHODS: Specimens from both HCC and noncancerous liver were obtained from 39 patients with HCC using a 14-gauge biopsy needle immediately after laparotomy. Telomerase activity was determined using a telomeric repeat amplification protocol assay. The 3+ of telomerase activity in HCC was defined as a high telomerase group, and 2+ or less of HCC telomerase activity was defined as a low telomerase group. In noncancerous liver, 2+ or more of telomerase activity was defined as an increased telomerase group, and 1+ or less of telomerase activity was defined as a nonincreased telomerase group. The correlation between telomerase activity in HCC or noncancerous liver and clinicopathological factors, including prognosis, was investigated. RESULTS: Telomerase activities in HCCs were 0 in one patient, 1+ in two, 2+ in seven, and 3+ in 29 patients. The disease-free survival rate in the high telomerase group was significantly worse than that in the low telomerase group. The des-gamma carboxy prothrombin level in a high telomerase group (median, 330 mAU/ml) was significantly higher than that in the low telomerase group (median, 150 mAU/ml). A multivariate analysis revealed that higher TNM stage, high telomerase activity in HCC, female gender, and high alpha-fetoprotein value were independent significant factors related to be early recurrence. The incidence of multicentric HCC occurrence in the increased telomerase group (53.3%) tended to be higher than that in the nonincreased telomerase group (27.3%). CONCLUSION: A high telomerase activity in HCC correlated with the potential of HCC to be more malignant, which was expressed as both a high level of des-gamma-carboxy prothrombin and an earlier recurrence after hepatectomy than that of HCC with a low telomerase activity. PMID- 10710070 TI - Hyperbilirubinemia and cholestatic liver injury in hepatitis C-infected liver transplant recipients. AB - OBJECTIVE: A cholestatic pattern of liver injury has been observed in liver transplant recipients with rapidly progressive hepatitis C. We assessed the frequency and causes of cholestasis in hepatitis C-infected liver transplant patients, and evaluated the clinical and pathological course of those with cholestatic hepatitis C. METHODS: Sixty-nine sequential liver transplant recipients who had detectable hepatitis C viremia were studied retrospectively. Records and diagnostic tests were examined from patients who developed hyperbilirubinemia. RESULTS: Hyperbilirubinemia occurred in 33 of 69 (48%) hepatitis C-infected liver transplant patients. A thorough evaluation including review of clinical and laboratory data, ultrasound with Doppler, cholangiogram, and liver biopsy identified causes of hyperbilirubinemia other than hepatitis C in 26 of 33 patients. Seven patients developed cholestatic hepatitis C characterized by histological features of recurrent hepatitis C and cholestatic liver injury with ballooning of centrilobular hepatocytes, bile ductular proliferation, and canalicular cholestasis, in the absence of other causes of cholestasis. Five progressed rapidly to bridging fibrosis and two died of complications related to liver failure. Four patients with cholestatic hepatitis C showed extended survival after the onset of hyperbilirubinemia. CONCLUSIONS: 1) Hepatitis C is a relatively infrequent cause of cholestasis in liver transplant recipients. 2) The diagnosis of cholestatic hepatitis C requires a multimodality approach to exclude other causes of cholestasis. 3) Cholestatic hepatitis C ranges in severity and is not always associated with rapid development of graft failure, although significant histological abnormalities are frequent. PMID- 10710071 TI - Impact of fatigue on the quality of life of patients with primary biliary cirrhosis. AB - OBJECTIVE: Fatigue is a frustrating symptom frequently reported by patients with primary biliary cirrhosis (PBC), but it is still poorly understood and not well evaluated. Our aim was to determine its importance and its impact on the quality of life and mental health status of patients with PBC. METHODS: Patients with PBC (103 women and 13 men with a mean age of 52.6 yr) completed self-report questionnaires to evaluate the impact of fatigue on their quality of life (Fatigue Impact Scale, FIS), the perception of their own mental health (Symptom Check list-90-R, SCL), and depression (Beck Depression Inventory, BDI). A cohort of age-matched healthy blood donors served as controls. RESULTS: Fatigue was present in 99 patients (85.3%) and was the worst or one of the worst symptoms in about half of them. In PBC patients, the mean FIS and SCL indexes were significantly increased, compared to healthy controls (1.49 +/- 1.11 vs 0.6 +/- 0.6 and 0.72 +/- 0.55 vs 0.36 +/- 0.35, respectively). Unexpectedly, 52 patients (44.8%) could be classified as having depression (BDI score > 10). Significant correlations were found between the FIS and SCL indexes, between the FIS index and the BDI score, as well as between the BDI score and the SCL index. Finally, fatigue was not related to the disease severity parameters, that is, clinical, biochemical, metabolic, and pathological. CONCLUSIONS: Fatigue is a frequent and disabling complaint that impairs the quality of life of PBC patients and their perception of their own mental health, which may be associated with an unexpected depressive condition. In addition, the FIS questionnaire can be considered as a useful tool to assess fatigue in PBC patients and may be used in the evaluation of specific treatments aimed at reducing this complaint in such patients. PMID- 10710072 TI - Comparison of endoscopic variceal sclerotherapy alone and in combination with octreotide in controlling acute variceal hemorrhage and early rebleeding in patients with low-risk cirrhosis. AB - OBJECTIVE: Efficacy of endoscopic variceal sclerotherapy (EVS) alone and in combination with octreotide in controlling acute variceal bleeding and preventing early rebleeding was compared in a double-blind study. METHODS: Consecutive patients presenting with variceal bleeding with low-risk liver cirrhosis were randomized into two groups. Group A received EVS with 3-5 ml of ethanolamine oleate per varix and placebo injection at 50 microg/h; group B received the combined therapy of EVS and octreotide 50 microg/h continuously for 5 days. A total of 70 patients (mean age, 38.4 +/- 8.6 yr) were selected for the study, which included 56 men (mean age, 37.9 +/- 8.5 yr) and 14 women (mean age, 40.6 +/ 9.0 yr). Thirty-five patients were allocated in each group. RESULTS: In group A bleeding was controlled in 30 patients (85.7%) and in group B in 33 (94.3%) (p = 0.24). The number of patients who rebled during the first 5 days after sclerotherapy was eight (22.9%) and two (5.7%) in groups A and B, respectively (p = 0.04). The mean packs of blood transfused to the patients of groups A and B were 2.1 +/- 1.2 packs and 1.5 +/- 0.7 packs, respectively (p = 0.03). The mean hospital stay of group A was 6.6 +/- 1.3 days, whereas that in group B was 5.9 +/ 1.2 days (p = 0.04). One patient from each group died during the course of the study. CONCLUSIONS: No significant difference was observed in arrest of bleeding in the two groups, but episodes of early rebleeding, blood transfusions, and hospital stay was significantly less in group B. PMID- 10710073 TI - Prevalence of TT virus infection in blood donors with elevated ALT in the absence of known hepatitis markers. AB - OBJECTIVE: Recently a novel DNA virus (TT virus) has been identified in Japan and shown to be associated with elevated aminotransferase levels after blood transfusion. The exact role of TTV in the pathogenesis of liver disease is yet to be established. Our aim was to determine the prevalence and role of TTV in the pathogenesis of elevated transaminases in healthy blood donors in the absence of markers for viral hepatitis A-C. METHODS: Stored sera were collected from 99 healthy blood donors with elevated alanine amino transferase (ALT) values that were discovered at the time of blood donation. A total of 146 samples were obtained from healthy donors with normal ALT values who were used as controls. None of the patients or controls had a history of blood transfusion or had clinical signs of acute or chronic hepatitis. Serological markers for hepatitis A, hepatitis B, and hepatitis C viruses were negative. TTV DNA was amplified and detected using polymerase chain reaction followed by gel electrophoresis. RESULTS: Five of 99 (5%) samples obtained from donors with elevated ALT had TTV DNA detected by PCR, as compared to one of 146 (0.7%) of those with normal ALT (p = 0.006). Among those with elevated ALT, mean ALT values in patients with TTV (296 +/- 305 U/L) were higher than in patients without TTV (95 +/- 37 U/L), but the difference was not statistically significant (p = 0.08). The two samples with highest ALT values (both >450 U/L) were among the five samples with detectable TTV DNA in serum. CONCLUSIONS: Although TTV is not likely to explain the majority of elevated ALT cases in otherwise healthy blood donors, TTV infection may potentially be associated with some cases. Based on these findings, we propose that the role of TTV in the pathogenesis of acute and chronic liver diseases merits further investigation. PMID- 10710074 TI - Why are internal medicine residents at university medical centers not pursuing fellowship training in gastroenterology? A survey analysis. AB - OBJECTIVE: The decrease in available GI fellowship positions appears to be associated with a disproportionate decrease in the quality of applicants. Thus, the aims of this study were: 1) to determine the current interest in advanced training of nonprimary care internal medicine residents at university medical centers, and 2) to identify the reasons why fellowship-bound residents are not pursuing GI. METHODS: Postage prepaid survey cards were distributed directly to the campus mailboxes of 1862 internal medicine residents at 61 university medical centers at the beginning of their second postgraduate year of training. E-mail questionnaires were then sent to 144 residents planning fellowship training and careers in academia other than in GI/hepatology. RESULTS: A total of 592 residents (32% response) returned completed survey cards. Overall, 392 (66%) indicated that they will pursue fellowship training and approximately 60% wanted to remain in academia. However, <10% indicated an interest in GI/hepatology. E mail replies were then obtained from 122 residents (87% response) planning academic careers but not in GI. The major reasons for disinterest in GI/hepatology include the perception that jobs are not widely available, that the field is intellectually unchallenging, as well as that is too procedure-oriented. CONCLUSIONS: The majority of residents at university training programs plan advanced training and want to pursue careers in academia, but not in GI/hepatology. Efforts to attract highly qualified residents to GI must emphasize the improved job market, especially as it exists in academia; must advertise research opportunities; and must de-emphasize the procedural nature of this subspecialty. PMID- 10710075 TI - Typhoid carriers among patients with gallstones are at increased risk for carcinoma of the gallbladder. AB - OBJECTIVE: The aim of this study was to identify risk factors for carcinoma of the gallbladder (CaGB) among patients with gallstones (GS) with special reference to role of chronic Salmonella typhi carrier state. METHODS: A prospective case control study was conducted in a tertiary care center in India. Cases were defined as consecutive patients with CaGB and GS, whereas controls were patients with GS alone. All were assessed clinically and their demographic data, diet, and smoking history recorded. Patients were detected to be typhoid carriers on the basis of Vi serology by enzyme linked immunosorbent assay. Cases (n = 37) and controls (n = 80) were compared by univariate and logistic regression analysis. RESULTS: The mean age of the cases and the controls were 53.4 +/- 11 yr and 43.5 +/- 14 yr, respectively. Among the cases, six (16%) patients were detected to be typhoid carriers, in contrast to two (2.5%) among controls (p = 0.01). Compared to controls, cases were more often older (p = 0.0002), of a lower socioeconomic status (p = 0.0005), and smokers (p = 0.0002). Stepwise logistic regression analysis identified typhoid carrier state (OR = 14; CI 2-92), age > or =47 yr (OR = 5; CI 2-14) and smoking (OR = 11; CI 2-71) as the three independent risk factors for development of CaGB among patients with GS. CONCLUSION: Chronic typhoid carrier state was the most important risk factor among patients with CaGB and gallstones. PMID- 10710076 TI - Work loss costs due to peptic ulcer disease and gastroesophageal reflux disease in a health maintenance organization. AB - OBJECTIVE: The aim of this study was to estimate the value of work time and productivity loss because of peptic ulcer disease (PUD) and gastroesophageal reflux disease (GERD). METHODS: A total of 300 adult members of Northern California Kaiser Permanente Medical Care Program with outpatient diagnoses of PUD or GERD were randomly selected for a record review to confirm diagnosis. A telephone survey was conducted soliciting information about work loss because of their disease. Reported work losses were valued at self-reported hourly wage to derive work loss costs. A total of 117 PUD and 102 GERD patients participated. RESULTS: About 75% of each sample was employed full-time or part-time. In all, 42% of potentially working PUD patients and 41% of GERD patients reported some lost work productivity because of their disease. The average loss (per person working) was $606 for PUD and $237 for GERD over a 3-month period. Reduced productivity while at work and part-time work because of the disease were the most costly productivity losses for PUD, whereas time off for physician visits and reduced productivity while at work were the most costly losses for GERD. CONCLUSIONS: Work loss costs for patients with PUD and GERD may be nearly as large as direct medical care costs, and are consistent with the more acute nature of PUD and the chronic pattern of GERD. The work losses resulting from these diseases are large enough to warrant consideration in guideline development and policy decisions for patients with PUD and GERD. PMID- 10710077 TI - Mean corpuscular volume may be a useful index of risk for colorectal adenoma in middle-aged Japanese men. AB - OBJECTIVE: It has been reported that alcohol intake and folate deficiency are associated with an increased risk of colorectal adenomas and carcinomas. Mean corpuscular volume (MCV) of red blood cells has been reported to be increased in these conditions. The purpose of this study was to assess the association between MCV and risk of colorectal adenoma. METHODS: The subjects were 497 middle-aged (45-65 yr old) men who underwent both barium enema examination and total colonoscopy. The subjects answered a questionnaire regarding their alcohol consumption history, and their blood samples were analyzed. The subjects were divided into four groups three times: with or without alcoholism, and with or without adenoma according to alcohol intake, and according to the MCV value. Various variables were compared among the groups, and the odds ratios of adenoma were calculated. RESULTS: The MCV was higher in the alcoholic group than in the nonalcoholic group (p < 0.01) and in patients with adenoma than in those without adenoma (p < 0.0001). When the subjects were stratified by alcohol intake, the MCV value had a higher significant difference than alcohol intake, between patients with adenoma and those without adenoma. As for the MCV value, the odds ratio (95% confidence interval) of adenoma was 1.00 (referent); (<92), 1.20 (0.71 1.69); (> or =92 but <95), 2.61 (2.07-3.15); (> or =95 but <98); and 3.62 (2.99 4.25); (> or =98). CONCLUSION: A high MCV value may be used as a simple index of the risk of colorectal adenomas, regardless of alcohol consumption. PMID- 10710078 TI - Doctor's doll. PMID- 10710079 TI - Syncytial giant-cell hepatitis due to autoimmune hepatitis type II (LKM1+) presenting as subfulminant hepatitis. AB - Giant cell hepatitis (GCH) in adults is a rare event. The diagnosis of GCH is based on findings of syncytial giant hepatocytes. It is commonly associated with either viral infection or autoimmune hepatitis type I. A patient with GCH due to autoimmune hepatitis type II (LKM1+) is described, a combination that has not been previously reported. Corticosteroid therapy was effective in decreasing serum liver enzymes; however, the patient deteriorated rapidly and developed subfulminant hepatic failure. Although an emergency orthotopic liver transplantation was performed, the patient died because of reperfusion injury. Interestingly, only a few giant hepatocytes were noted in the explanted liver. This case stresses the association of GCH with autoimmune disorders, the possible immune mechanism involved in the formation of giant cell hepatocytes, and illustrates the rapidly progressive course and unfavorable prognosis that these patients can develop. PMID- 10710080 TI - Usefulness of exchange transfusion in acute liver failure due to severe falciparum malaria. AB - Acute hepatic failure is a rare and serious complication of severe falciparum malaria. The management of uncomplicated falciparum malaria comprises of specific antimalarial drugs and supportive therapy. In a few patients who are critically ill because of severe falciparum malaria and heavy parasitaemia, exchange transfusion has been used. We describe a young male Saudi patient who presented with a 2-day history of fever, jaundice, and confusion. On examination he was deeply jaundiced, confused, and irritable. There were no signs of chronic liver disease. His laboratory workup revealed a markedly raised direct hyperbilirubinaemia and transaminases with prolonged prothrombin time. His serology was negative for HbsAg, HBc IgM, anti-HCV, HAV IgM, HEV IgM, and IgG. He was initially treated with parenteral quinine and other supportive treatment, without any improvement of his clinical and laboratory parameters. At this stage he was treated with whole blood exchange transfusion. He slowly improved, with complete normalization of his liver function tests and prothrombin time. PMID- 10710081 TI - Cholecystitis caused by hemocholecyst from underlying malignancy. AB - Massive hemobilia is a well recognized clinical entity, particularly when it presents with jaundice, GI bleeding, and biliary pain. However, occult hemobilia is more difficult to diagnose and has seldom been reported because of its clinically silent nature. In fact, this is usually overlooked until complications arise. Hemocholecyst or clot within the gallbladder may rarely occur in this setting, leading to cystic duct obstruction and cholecystitis. Most previous reports describe cholecystitis resulting from hemocholecyst after iatrogenic trauma. We describe two cases in which hemocholecyst occurred from underlying malignancies, both resulting in cholecystitis (acute or chronic). PMID- 10710082 TI - Completion of upper endoscopic procedures despite paradoxical reaction to midazolam: a role for flumazenil? AB - Paradoxical excitation after benzodiazepine administration is well described. Although it is relatively uncommon, its occurrence can severely impede or even prevent the performance of upper endoscopy. We describe three cases in which paradoxical reactions to midazolam responded so well to flumazenil administration that the procedure was successfully completed in each instance. We review the limited literature on this topic and suggest that flumazenil may have greater utility in the management of this particular problem than is considered at present. PMID- 10710083 TI - Should we colonoscope women with gynecologic cancer? AB - Retrospective analysis of the Surveillance Epidemiology and End Results (SEER) program database for the period 1974 through 1995 identified 101,734 white and African-American women, age > or = 25 yr, with prior cervical, endometrial, or ovarian cancer. Subsequent follow-up demonstrated no increased risk of colorectal cancer in women with cervical cancer. For endometrial cancer patients, increased risk of colorectal cancer was confined to women whose diagnosis of endometrial cancer was before age 50, but the increased risk was substantial in this group (RR 3.39; 95% CI 2.73-4.17). For ovarian cancer patients, increased risk for colorectal cancer was substantial for those diagnosed with ovarian cancer before age 50 (RR 3.67; 95% CI 2.74-4.80), and there was some increased risk for women diagnosed at ages 50-64 yr (RR 1.52; 95% CI 1.25-1.83). PMID- 10710084 TI - Just a little heartburn? AB - Lagergren et al. performed a population-based, case-control study to investigate the possible association between gastroesophageal reflux (GER) and adenocarcinoma of the esophagus and gastric cardia. They demonstrated an odds ratio of 7.7 for esophageal adenocarcinoma in patients with GER symptoms. The frequency, severity, and duration of symptoms correlated with an increased risk of esophageal adenocarcinoma. A weaker association was noted for GER and adenocarcinoma of the gastric cardia. No association surfaced between GER and squamous cell carcinoma of the esophagus. PMID- 10710085 TI - Hyperammonemia and intracranial hypertension: lying in wait for patients with hepatic disorders? AB - In the past decade, a dozen patients with acute liver injury, with idiopathic hyperammonemia and intracranial hypertension in the absence of acute liver failure have been reported, as discussed below. If one combines these patients to those with acute liver failure and Reye's syndrome, in both of which cerebral herniation is a common complication, liver disease may be one of the most common causes of cerebral herniation. Indeed, these reports have similarities to the patients of Clemmesen et al. who found that increased blood ammonia levels are frequently elevated, which led to the observation that increased blood ammonia concentrations may be early warning signs of impending cerebral herniation. PMID- 10710086 TI - US academic digestive disease sites on the internet. PMID- 10710087 TI - GI sources on the internet. PMID- 10710088 TI - Re: Peppers at treatment for Helicobacter pylori infection. PMID- 10710089 TI - Obstructive jaundice as a recurrent symptom of small cell lung cancer. PMID- 10710090 TI - Re: Amylase normal, lipase elevated: is it pancreatitis? PMID- 10710091 TI - Profound hypocalcemia in fulminant hepatic failure. PMID- 10710092 TI - Initial presentation of ulcerative colitis with acute pancreatitis. PMID- 10710093 TI - Erythromycin eye ointment: effect on gastrointestinal motility. PMID- 10710094 TI - Ventricular fibrillation after insertion of a self-expanding metallic stent for malignant dysphagia. PMID- 10710095 TI - Complete regression of hepatocellular carcinoma under tamoxifen. PMID- 10710096 TI - Topical pharyngeal anesthesia for upper gastrointestinal endoscopy. PMID- 10710098 TI - Liver hemangioma with systemic inflammatory manifestations. PMID- 10710097 TI - Cyclosporin for steroid-refractory ulcerative colitis. PMID- 10710099 TI - Once-daily Helicobacter pylori treatment to family members of gastric cancer patients. PMID- 10710100 TI - In vitro activity of rifabutin against strains of Helicobacter pylori resistant to metronidazole and clarithromycin. PMID- 10710101 TI - Age, uncomplicated dyspepsia, endoscopy, and gastric cancer. PMID- 10710102 TI - Coinfection with hepatitis C virus and TT virus in a case of late onset hepatic failure. PMID- 10710103 TI - Gallstone migration into the pleural space. PMID- 10710104 TI - pp60c-src activation in gastric carcinoma: a preliminary study. PMID- 10710105 TI - Aortoenteric fistula revisited: an endoscopic image. PMID- 10710106 TI - Re: Shad and Schindler's report on spontaneous peritonitis due to Streptococcus bovis. PMID- 10710107 TI - Does administration of infliximab increase susceptibility to listeriosis? PMID- 10710108 TI - Gastric duplication cyst with markedly elevated concentration of carbohydrate antigen 19-9. PMID- 10710109 TI - Lower gastrointestinal bleed in a patient with typhoid fever. PMID- 10710110 TI - Cumulative hepatotoxicity induced by continuous low-dose cyclophosphamide therapy. PMID- 10710111 TI - Are myocardial functional abnormalities biventricular in nonalcoholic cirrhotics with or without ascites? PMID- 10710112 TI - Immunoproliferative small intestinal disease. PMID- 10710114 TI - Crohn's disease of the bladder--a new type of metastatic granulomatous inflammatory disease? PMID- 10710113 TI - Metastatic gastrointestinal cancer compressing the spinal cord or cauda equina. PMID- 10710115 TI - Asymptomatic ileal carcinoid detected during colonoscopy. PMID- 10710116 TI - Bone marrow transplantation: hope for the treatment of chronic hepatitis B? PMID- 10710117 TI - Roles of tyrosine kinase and protein kinase C in infarct size limitation by repetitive ischemic preconditioning in the rat. AB - In this study, we examined the possibility that infarct-size limitation by repetitive preconditioning (PC) is achieved by activation of both protein kinase C (PKC) and tyrosine kinase. In addition, we assessed whether such kinase activation is triggered by angiotensin II type 1 (AT1) and alpha1-adrenergic receptors and whether sarcolemmal and mitochondrial adenosine triphosphate (ATP) sensitive potassium (K(ATP)) channels play roles as effectors of cardioprotection in the rat. Under pentobarbital anesthesia, myocardial infarction was induced by 20-min coronary occlusion and 3-h reperfusion in the rat. Infarct size was determined by tetrazolium and expressed as a percentage of area at risk (%IS/AR). PC with one cycle of 5-min ischemia/5-min reperfusion before 20-min ischemia significantly reduced %IS/AR from the control value of 49.4 +/- 2.0 to 35.4 +/- 2.8, and repetitive PC with two cycles of 5-min ischemia/5-min reperfusion further limited %IS/AR to 3.2 +/-0.9. Infarct-size limitation by single-cycle PC was completely abolished by a PKC inhibitor, staurosporine (100 microg/kg; %IS/ AR, 45.7 +/- 5.0). In contrast, the cardioprotection by repetitive PC was only partially blocked by staurosporine (%IS/AR, 19.8 +/- 2.4), another PKC inhibitor, polymyxin B (5 mg/kg; %IS/AR, 16.2 +/- 3.1), or a tyrosine kinase inhibitor, genistein (5 mg/kg; %IS/AR, 21.8 +/- 1.4). However, a combined injection of genistein and staurosporine additively inhibited protection of repetitive PC (%IS/AR, 36.4 +/- 1.7). Staurosporine, polymyxin B, or genistein alone did not modify %IS/AR in nonpreconditioned rat hearts. Infarct-size limitation by repetitive PC was not attenuated by pretreatment with a selective AT1-receptor blocker (CV11974, 10 mg/kg), prazosin (0.6 mg/kg; %IS/AR, 6.4 +/- 3.2 and 1.6 +/- 0.5, respectively). A selective blocker of mitochondrial K(ATP) channels, 5 hydroxydecanoate (3 mg/kg), completely abolished the cardioprotective effect (%IS/AR, 50.8 +/-3.5), but HMR1883 (3 mg/kg), a selective blocker of sarcolemmal K(ATP) channels, failed to inhibit the preconditioning effect (%IS/AR, 4.4 +/- 0.7). These findings suggest that repetition of PC provokes activation of both PKC and tyrosine kinase, leading to enhanced antiinfarct tolerance by opening of mitochondrial but not sarcolemmal K(ATP) channels. It is unlikely that activation of either AT1 or alpha1-adrenergic receptor alone is crucial to trigger preconditioning. Key Words: Tyrosine kinase-Genistein-Angiotensin II-alpha1 Adrenergic receptor-Sarcolemmal K(ATP) channel-Mitochondrial K(ATP) channel. PMID- 10710118 TI - Regional and temporal profiles of phorbol 12,13-dibutyrate binding after myocardial infarction in rats: effects of captopril treatment. AB - Phosphoinositide turnover and protein kinase C (PKC) mediate the signaling of angiotensin II, which plays a pivotal role in ventricular remodeling after myocardial infarction (MI). To determine whether PKC is activated after MI, rat hearts after MI were subjected to in vitro quantitative autoradiography with [3H]phorbol 12,13-dibutyrate (PDBu), which is highly selective for PKC. [3H]PDBu binding in the infarcted area increased significantly compared with the non infarcted region 7 and 21 days after MI, but not 1 and 3 days and 10 months after MI. [3H]PDBu binding in the noninfarcted area was similar to that in the sham operated rats. Immunohistochemical analysis revealed that abundant macrophages (7 days after MI), fibroblasts, and myofibroblasts (7 and 21 days after MI) occupied the infarcted region. To investigate whether myocardial [3H]PDBu binding is affected by captopril, hearts were subjected to in vitro autoradiography with [3H]PDBu after 1- or 3-week captopril treatment or no treatment. Captopril treatment significantly suppressed [3H]PDBu binding in the infarcted area 3 weeks after MI, but not 1 week after MI nor in the noninfarcted areas. These results suggest that PKC is upregulated during the healing and fibrogenic process after MI and that captopril treatment suppresses the upregulation in the infarcted area. PMID- 10710119 TI - Efficacy and tolerability of fluvastatin and bezafibrate in patients with hyperlipidemia and persistently high triglyceride levels. AB - Hyperlipidemia is an important cardiovascular risk factor. Lipid-lowering therapy has been shown to decrease morbidity and mortality in these patients. Combination therapy with a 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor and a fibric-acid derivative has been reported to be more efficacious to reduce low-density lipoprotein (LDL) cholesterol and triglycerides but may be associated with an increased risk of myositis. The aim of this study was to investigate the efficacy and tolerability of fluvastatin, an HMG-CoA reductase inhibitor, alone and in combination with bezafibrate, a fibric-acid derivative. In a randomized controlled trial with 454 hypercholesterolemic patients (mean cholesterol, 8.6 +/- 1.6 mM), fluvastatin (20 mg/day) significantly lowered total plasma cholesterol levels (-12.5%; p < 0.0001 vs. placebo), LDL cholesterol ( 14%; p < 0.0001), and triglycerides (-4%; p = 0.05). A small increase in high density lipoprotein (HDL) cholesterol levels (3%, NS) also was observed. Combination therapy with fluvastatin and bezafibrate (400 mg/day) in 71 patients with persistent hypertriglyceridemia during treatment with the statin resulted in a more pronounced reduction in triglyceride (-47%; p < 0.0001) and total cholesterol levels (-15%; p < 0.0001) than did fluvastatin alone. Furthermore, the additional bezafibrate significantly increased HDL cholesterol (+5%; p < 0.001). No significant increases in creatine phosphokinase levels or in frequency of myalgia were observed. In summary, fluvastatin decreases both cholesterol and triglyceride levels. In patients with persistent hypertriglyceridemia, combination therapy with fluvastatin and bezafibrate may be safely used to lower triglyceride and cholesterol levels more efficiently. PMID- 10710120 TI - Monophosphoryl lipid A: a novel nitric oxide-mediated therapy to attenuate platelet thrombosis? AB - Nitric oxide (NO) is a potent inhibitor of platelet aggregation. However, the benefits of NO-based therapies can be confounded by concomitant hypotension. Monophosphoryl lipid A (MLA) is a nontoxic derivative of endotoxin that purportedly increases nitric oxide synthase (NOS) activity and, presumably, NO production, yet has a hemodynamically benign profile. Thus our aims were to determine whether (a) MLA attenuates in vivo platelet aggregation in damaged and stenotic canine coronary arteries by a NO-mediated mechanism but without reductions in arterial pressure; and (b) the platelet inhibitory effects are manifest in vitro. To address the first aim, anesthetized dogs underwent coronary injury + stenosis, resulting in cyclic variations in coronary blood flow (CFVs) caused by the formation/dislodgement of platelet-rich thrombi. In protocol I, dogs received MLA (100 microg/kg + 40 microg/kg/h) or vehicle beginning 15 min before stenosis. Protocol II was identical, except the NOS inhibitor aminoguanidine was coadministered with MLA/vehicle. Coronary patency was assessed throughout the initial 3 h after injury + stenosis. Infusion of MLA did not result in hypotension. However, in protocol I, the median nadir of the CFVs was higher (2.1 vs. 0.8 ml/min; p < 0.05), median duration of total thrombotic occlusion tended to be reduced (0 vs. 10.4 min; p = 0.1), and mean flow-time area, expressed as a percentage of baseline flow, was increased (53 +/- 9% vs. 33 +/- 3%; p < 0.05) in MLA-treated versus vehicle-treated dogs. In contrast, in protocol II, vessel patency was comparable in both groups. Finally, whole blood impedance aggregometry (protocol HI) revealed a significant reduction in the in vitro platelet aggregation in blood samples receiving exogenous MLA, which was blocked by coadministration of exogenous aminoguanidine. Thus MLA attenuates platelet-mediated thrombosis in both damaged and stenotic canine coronary arteries and in vitro, possibly by an NO-mediated mechanism, but without concomitant hypotension. PMID- 10710121 TI - Effects of fluvastatin treatment on red blood cell Na+ transport systems in hypercholesterolemic subjects. AB - This study was performed to ascertain the effects of short-term cholesterol lowering therapy with fluvastatin on red blood cells Na+ transport systems. Forty familial hypercholesterolemic subjects (FH; 19 men and 21 women) without hypertension or cardiovascular disease were given a placebo for 4 weeks, and then randomized in two groups. Twenty (fluvastatin group) were given fluvastatin (40 mg/day), and the other 20 (placebo group) continued placebo administration. After the placebo period and after 4 and 12 weeks of placebo or fluvastatin treatment, we measured Na+/K+ pump activity, Na+/K+ cotransport (Na+/K+ Ct), Na+/Li+ countertransport (Na+/Li+ Cnt), passive Na+ permeability (Na+PP), and internal Na+ content (Na+i). The same parameters were measured in 23 control subjects (C) with normal cholesterolemic values, who were matched for sex and age. FH had higher Na+/Li+ Cnt values than C (193.2 +/- 59.4 vs. 139.8 +/- 48.7 microM cells/h; p < 0.01), an increase in Na(+)PP (0.034 +/- 0.012/h vs. 0.018 +/- 0.004/h; p < 0.001), and higher Na(+)i (7.5 +/- 1.5 vs. 6.2 +/- 0.9 mM cells; p < 0.001). In hypercholesterolemic subjects, Na(+)i values were correlated with cholesterol (total and LDL) and apo B levels, whereas an inverse correlation was found for HDL-c and apo AI levels. Reduced total and LDL cholesterol and apo B levels after fluvastatin treatment caused a decrease in both Na(+)/Li(+) Cnt (from 186.1 +/- 60.5 to 125.1 +/- 34.0 microM cells/h; p < 0.001) and Na(+) PP (from 0.035 +/- 0.013/h to 0.02 +/- 0.016/h; p < 0.01), and an increase in Na+/K+ pump activity (from 1,549.0 +/- 507.7 to 1,894.2 +/- 536.2 microM cells/h; p < 0.04), with a significant reduction in the internal Na+ content (from 7.5 +/- 1.6 to 5.8 +/- 2.4 mM cells; p < 0.001). Our findings show that hypercholesterolemia affects red blood cell Na+ transport systems, with an increase in Na+/Li+Cnt, Na+PP, and the internal Na+ content. Cholesterol-lowering treatment with fluvastatin influences Na+ transport systems and reduces the internal Na+ content. This might also be responsible for the greater vascular reactivity observed in hypercholesterolemic patients, and its amelioration after a reduction in cholesterol levels. PMID- 10710122 TI - Pharmacologic profile of UR-7247, an orally active angiotensin II AT1 receptor antagonist, in healthy volunteers. p6. AB - This study was conducted to assess the pharmacologic properties of the new orally active angiotensin II subtype I (AT1) antagonist UR-7247, a product with a half life >100 h in humans. The experiment was designed as an open-label, single-dose administration study with four parallel groups of four healthy men receiving increasing single oral doses (2.5, 5, and 10 mg) of UR-7247 or losartan, 100 mg. Angiotensin II receptor blockade was investigated < or =96 h after drug intake, with three independent methods [i.e., the inhibition of blood pressure (BP) response to exogenous Ang II, an in vitro Ang II-receptor assay (RRA), and the reactive increase in plasma angiotensin II. Plasma drug levels also were measured. The degree of blockade observed in vivo was statistically significant < or = 96 h with all UR-7247 doses for diastolic BP (p < 0.05) and < or =48 h for systolic BP. The maximal inhibition achieved with 10 mg UR-7247 was measured 6-24 h after drug intake and reached 54 +/- 17% and 48 +/- 20% for diastolic and systolic responses, respectively. Losartan, 100 mg, induced a greater short-term AT1-receptor blockade than 2.5- and 5.0-mg doses of UR-7247 (p < 0.001 for diastolic BP), but the UR-7247 effect was longer lasting. In vivo, no significant difference was observed between 10 mg UR-7247 and 100 mg losartan 4 h after drug intake, but in vitro, the blockade achieved with 100 mg losartan was higher than that seen with UR-7247. Finally, the results confirm that UR-7247 has a very long plasma elimination half-life, which may be due to a high but also tight binding to protein binding sites. In conclusion, UR-7247 is a long-lasting, well tolerated AT1 receptor in healthy subjects. PMID- 10710123 TI - The vasodilator-stimulated phosphoprotein (VASP): target of YC-1 and nitric oxide effects in human and rat platelets. AB - The effects of the different types of soluble guanylate cyclase (sGC) stimulators on the phosphorylation status of vasodilator-stimulated phosphoprotein (VASP) in both human and rat platelets were studied under in vitro and in vivo conditions. sGC-dependent VASP phosphorylation (at Ser(239) and Ser(157)) both by the new direct sGC stimulator YC-1 and by NO donors was examined by sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS/PAGE) with different antibodies. One antibody, which recognizes VASP independent of its phosphorylation state, was used to detect the mobility shift of VASP caused by Ser(157) phosphorylation. The other antibody was specifically directed against VASP phosphorylated at Ser(239), the cGMP-dependent protein kinase (PKG) preferred phosphorylation site of VASP. In vitro YC-1 increased both VASP phosphorylation and cyclic guanosine monophosphate (cGMP) levels as did the NO donors 2-(N,N-diethylamino)-diazenolate 2-oxide (DEA/NO) and sodium nitroprusside (SNP). The combination of both types induced a synergistic effect in both VASP phosphorylation and cGMP increase. In rat platelets, similar effects could be shown in vitro. In vivo we observed a significant increase in cGMP and a distinct effect on VASP phosphorylation in rat platelets 1 h after oral administration of YC-1. These biochemical alterations are supported by a significant prolongation in rat-tail bleeding time. Direct stimulators of sGC like YC-1 are on the one hand direct potent stimulators of the cGMP/PKG/VASP pathway in platelets and on the other hand synergize with NO, the physiologic stimulator of sGC. Therefore YC-1-like substances are interesting tools for the development of new cardiovascular drugs with vasodilatory and antithrombotic properties. PMID- 10710124 TI - 5-hydroxytryptamine stimulates phosphorylation of p44/p42 mitogen-activated protein kinase activation in bovine aortic endothelial cell cultures. AB - 5-Hydroxytryptamine (5-HT) is sequestered and released by endothelial cells, acts as an endothelial cell mitogen, promotes the release of nitric oxide (NO), and has been associated with the p44/p42 mitogen-activated protein kinase (MAPK) cascade. NO also acts as a cell mitogen and promotes signals that culminate in the phosphorylation of MAPK. The aim of this study was to test whether endothelial 5-HT receptors stimulate dual (tyrosyl- and threonyl-) phosphorylation of MAPK through a mitogen-activated protein kinase kinase-1 (MEK 1) and eNOS-dependent pathway in bovine aortic endothelial cells (BAECs). As shown by Western blot analysis, 5-HT and the 5-HT1B-selective agonist 5 nonyloxytryptamine (5-NOT) stimulate time- and concentration-dependent (0.001-10 microM) phosphorylation of MAPK in these cells. The agonist-stimulated phosphorylation of MAPK was blocked by the 5-HT1b-receptor antagonist isamoltane (0.01-10 p3M) and the MEK-1 inhibitor PD 098059 ([2-(2'-amino-3'-methoxy-phenyl) oxanaphthalen-4-one]; 0.01-10 microM?. The eNOS inhibitor L-N(omega)-iminoethyl-L ornithine (L-NIO; 0.01-10 microM) failed to block the 1 microM 5-NOT-stimulated responses. Our findings suggest that the 5-HT receptors (specifically 5-HT1B) mediate signals to MEK-1 and subsequently to MAPK through an eNOS-independent pathway in BAECs. PMID- 10710125 TI - Effects of an endothelin receptor antagonist TAK-044 on myocardial energy metabolism in ischemia/reperfused rat hearts. AB - The purpose of this study was to investigate the effects of an endothelin receptor antagonist TAK-044 on functional defects and metabolic derangement in myocardial ischemia/reperfusion injury. We sequentially measured high-energy phosphate metabolites and intracellular pH by phosphorus magnetic resonance spectroscopy during 35-min global ischemia followed by 60-min reperfusion in Langendorff-perfused rat hearts. TAK-044 (initial loading by 3 mg/kg followed by perfusion with 100 nM solution) was administered in two different ways: before ischemia or immediately after reperfusion. In addition, we investigated the effects of TAK-044 on functional defects and metabolic alterations induced by hydrogen peroxide (200 microM, 30 min). The recoveries of left ventricular developed pressure after reperfusion in TAK-044 groups (51 +/-12% in TAK-I, 61 +/ 12% in TAK-R) were better than in control (10 +/- 5% in control; p < 0.01). Increases in left ventricular end-diastolic pressure (LVEDP) in TAK-044 groups (22 +/- 5 mm Hg in TAK-I, 24 +/- 5 mm Hg in TAK-R) were less than in control (38 +/- 3 mm Hg; p < 0.01). Adenosine triphosphate (ATP) (33 +/- 5% in TAK-I, 28 +/- 4% in TAK-R) in TAK-044 groups were higher than in control (13 +/- 3%; p < 0.01). The creatine phosphokinase (CPK) release during reperfusion in TAK-044 groups (3.3 +/- 1.5 IU/g wet wt/60 min in TAK-I, 3.5 +/- 2.5 IU/g wet wt/60 min in TAK R) were lower than in control (13.8 +/- 3.9 IU/g wet wt/60 min; p < 0.05). In contrast, TAK-044 did not attenuate the myocardial injury induced by hydrogen peroxide. TAK-044, even if administered simultaneous with coronary reperfusion, attenuated myocardial ischemia/ reperfusion injury. The energy-preservative effect of TAK-044 could be associated with the good functional recovery in ischemia/reperfused rat hearts. PMID- 10710126 TI - FTY720 prevents development of experimental autoimmune myocarditis through reduction of circulating lymphocytes. AB - FTY720 is a new immunosuppressant agent and selectively decreases the number of circulating lymphocytes. In this study, we compared the effects of FTY720 with those of tacrolimus on experimental autoimmune myocarditis (EAM) in rats. A significant decrease in circulating lymphocyte counts was noted after a single administration of FTY720 in normal rats. At day 0, 7-week-old male Lewis rats were immunized with purified porcine cardiac myosin emulsified in complete Freund's adjuvant. FTY720 or tacrolimus was administered intraperitoneally daily. The number of myocarditis-affected areas in the FTY720 treatment groups with doses of 0.1 mg/kg/ day was significantly lower than those in control groups at days 14 and 28. In addition, at day 28, the myocarditis-affected areas in the FTY720 treatment group were significantly smaller than those in the tacrolimus treatment group receiving the same dose. Effects of early administration (days 0 10) and delayed administration (days 11-20) of FTY720 also were examined. At day 28, the myocarditis-affected areas in the early therapy group were significantly lower than those in the control group. In conclusion, we demonstrated that the development of EAM could be prevented by FTY720. These data also indicated that lymphocyte-mediated immunity is critically involved in the development of EAM. PMID- 10710127 TI - Effects of moxonidine on corticocerebral blood flow under normal and ischemic conditions in conscious rabbits. AB - Hypertension associated with excessive liberation of circulating and tissue catecholamines is an independent risk factor for further cardiovascular complications and an important predictor of stroke. Moxonidine is a centrally acting anti-hypertensive drug with potent action on I1-imidazoline receptors. It inhibits catecholamine release and is therefore expected to exert an antiadrenergic effect at various levels in the regulation of the cardiovascular system. The aim of this study was to investigate the effect of moxonidine (0.025 0.1 mg/kg, i.v.) on the normal and unilateral carotid occlusion-induced impaired corticocerebral blood flow (cCBF) determined by hydrogen polarography, on mean arterial blood pressure (MABP) and heart rate (HR) in conscious rabbits. Moxonidine produced a reduction of MABP and HR. On the other hand, after administration of the drug, a significant increase in the normal and impaired cCBF was observed. Because the improvement in cCBF was conspicuous in both normal and ischemic conditions, moxonidine might be beneficial not only in the treatment of hypertension but also in the management of cerebral ischemia. PMID- 10710129 TI - The positive inotropic effects of milrinone but not of digoxin are attenuated at short cycle lengths. AB - The effects of inotropically active agents on the left ventricular force-interval relation are a potential determinant of their clinical utility and safety. However, little information is available concerning the effects of noncatecholamine positive inotropic agents on this relation. Therefore this study compared the short-term effects of digoxin and milrinone on resting hemodynamics, frequency potentiation (FP), and mechanical restitution (MR) in patients undergoing nonemergency cardiac catheterization. Both digoxin and milrinone produced similar increases in LV + dP/dt at rest (12.2 +/- 1.3%, p < 0.000001 and 11.4 +/- 3.2%, p < 0.01, respectively). The positive inotropic effects of digoxin were marginally attenuated during FP (by 8.5 +/- 4.2% and 4.6 +/- 2.9% at 10 and 60 s, respectively, both p = NS compared with baseline). Similarly, on MRC analysis, the parameter c (a measure of sensitivity of contractile performance to reductions in cycle length) increased by 3.6 +/- 3.7% (p = NS). Whereas the positive inotropic effects of milrinone were not significantly attenuated during FP, they were abolished and possibly reversed at short cycle lengths on MR curve construction (6.8 +/- 5.9% negative inotropic effect at 60% of resting cycle length; p = NS; p < 0.05 vs. resting cycle length). In conclusion, in patients with well-preserved left ventricular systolic function, the positive inotropic effects of milrinone but not of digoxin are markedly dependent on heart rate. These properties may influence both relative safety and efficacy of both agents. PMID- 10710128 TI - Effect of olprinone, phosphodiesterase III inhibitor, on cerebral blood flow assessed with technetium-99m-ECD SPECT. AB - Few clinical reports exist regarding the effects of phosphodiesterase III inhibitors on cerebral arteries. Therefore we used a [99mTc]-ECD brain SPECT and an ultrasound method to quantitatively evaluate cerebral and systemic flow dynamics after the administration of olprinone, a phosphodiesterase III inhibitor. In 15 patients (65 +/- 8 years, M/F = 13/2) with no abnormalities on a brain computed tomography (CT), cerebral blood flow and cardiac output were measured using [99mTc]-ECD brain SPECT and Doppler echocardiography, respectively. Measurements were repeated at baseline and after the administration of olprinone. Significant increases in cerebral blood flow (p = 0.0007) and cardiac output (p = 0.001) were observed, and systolic blood pressure (p = 0.02) and systemic vascular resistance (p = 0.002) significantly decreased. No significant correlation between the increase in cerebral blood flow and the increase in cardiac output was observed. The data indicate that olprinone has a direct vasodilator effect on the cerebral artery, which was little related to the increase in cardiac output in patients who had not experienced stroke. PMID- 10710130 TI - Blockade of cardiac ATP-sensitive K+ channel by cibenzoline targets its pore forming subunit. AB - Several antiarrhythmic agents with Na-channel blocking action have been shown to inhibit cardiac K(ATP) channels. We used cibenzoline to examine its precise target site using patch-clamp techniques and receptor binding assays in guinea pig ventricular myocytes. Exposure of myocytes to a glucose-free perfusate containing 1 mM cyanide produced a time-dependent shortening of the action potential duration (APD) in the current-clamp mode. Cibenzoline (30 microM) slowed the development of APD shortening (APD90 to approximately 91% vs. approximately 55% control 16 min after metabolic inhibition) at pHo 7.4, but not at pHo 6.4 (to approximately 60%). The pinacidil (30 microM)-induced K(ATP) currents were inhibited by cibenzoline in a pHo-dependent manner: the higher the pHo, the stronger the blocking effect of cibenzoline. The binding of [3H]-labeled cibenzoline was prevented by cibenzoline, but not by glibenclamide. Alkalinization produces a higher concentration of the uncharged form of cibenzoline, which can more easily permeate the cell membrane than the charged form. In NIH3T3 cells stably expressing Kir6.1, a putative pore-forming subunit of K(ATP) channel, cibenzoline but not glibenclamide inhibited the K conductance. Thus cibenzoline interacts with the channel pore-forming subunit of the K(ATP) channel (Kir6.2), but not the sulfonylurea receptor, from the cytosolic side after it permeates into the cell interior via the membrane lipid bilayer. PMID- 10710131 TI - Isoproterenol specifically modulates reverse rate-dependent effects of d,l sotalol, d-sotalol, and dofetilide. AB - The modulation of antiarrhythmic and proarrhythmic properties of antiarrhythmic compounds by increased sympathetic activity is of experimental and clinical interest. However, the interaction of adrenergic stimulation with the rate response pattern of class III antiarrhythmic agents is not well established. Using standard microelectrode techniques, we evaluated the effects of isoproterenol (iso) on the action of d,l-sotalol (d,l-sot), d-sotalol (d-sot), and dofetilide (dof) on action-potential parameters recorded from isolated canine cardiomyocytes. The cell-isolation procedure was performed from the endocardial layers of left ventricular myocardium from healthy beagle dogs. The following electrophysiologic parameters were recorded: resting membrane potential (RMP), action-potential amplitude (APA), action-potential duration at 90% repolarization (APD 90), and effective refractory period (ERP). After exposure to iso, the class III activity of d,l-sot was well maintained over the entire range of frequencies studied. In contrast, iso differentially antagonized the action of d-sot and dof. In comparison to dof, the class III action of d-sot was particularly sensitive to iso, predominantly at faster stimulation rates. Our observations demonstrate specific rate regulation of the class III action of d,l-sot, d-sot, and dof in response to adrenergic stimulation. The unfavorable rate-response pattern of d sot compared with d,l-sot and dof might prove disadvantageous in high catecholamine states. PMID- 10710132 TI - Short-term effects of smoking and nicotine chewing gum on endothelium-dependent vasodilation in young healthy habitual smokers. AB - Smoking is a major risk factor for coronary and peripheral vascular disease. This study was designed to investigate the short-term effects of smoking and nicotine gum on endothelium-dependent (EDV) and -independent (EIDV) vasodilation in the forearm of young habitual smokers. In 10 subjects, forearm blood flow (FBF) during local infusion of metacholine (4 microg/min, evaluating EDV) and sodium nitroprusside (10 microg/min, evaluating EIDV) was assessed before and at 10 min (early phase) and 30-50 min (plateau phase) after the initiation of smoking, using forearm venous occlusion plethysmography. Six subjects underwent similar measurements of FBF before and 30 min after chewing a nicotine gum (4 mg). As a change in blood pressure was expected, forearm vascular resistance (FVR) was used to calculate EDV and EIDV. FVR during metacholine infusion increased from 4.6 +/- 1.4 SD to 5.9 +/- 2.1 mm Hg/ml/min/100 ml tissue during the early and to 5.0 +/- 1.6 mm Hg/ml/min/100 ml tissue at the plateau phase of smoking (p < 0.01 for both vs. baseline) and from 4.5 +/- 1.6 to 5.2 +/- 1.6 mm Hg/ml/min/100 ml tissue after chewing the nicotine gum (p < 0.01). No significant changes in EIDV were seen after smoking or the nicotine gum. When all data were analyzed together, plasma nicotine levels and blood pressure were both independent predictors of endothelial function (p < 0.001 for both). In conclusion, cigarette smoking induced a dose-dependent attenuation in EDV, being maximal shortly after initiation of smoking and persisting up to 30-50 min. Nicotine chewing gum induced a similar impairment in EDV. PMID- 10710133 TI - The inhibitory effect of KT3-671, a nonpeptide angiotensin-receptor antagonist, on rabbit and rat isolate vascular smooth muscles: a possible involvement of K(ATP) channels. AB - The vasoinhibitory effect of KT3-671, a recently synthesized nonpeptide angiotensin II (Ang II), AT1-receptor antagonist, and the factors affecting insurmountable antagonism of Ang II were examined in rabbit and rat isolated vascular smooth muscle preparations. In rabbit and rat aortic rings, KT3-671 caused insurmountable antagonism of Ang II. In addition, KT3-671 inhibited contractile responses to angiotensin III (Ang III). In rabbit isolated smooth muscles, KT3-671 was most effective in reducing the maximal contraction induced by Ang II in the renal artery followed by the basilar artery and the aorta. In rat renal arterial rings, KT3-671 (10(-5) M) inhibited the concentration-response curves of prostaglandin F2alpha and STA2. In rabbit and rat aortic rings without endothelium, the insurmountable antagonisms of Ang II by KT3-671 and EXP 3174 were changed to surmountable antagonism by pretreatment with DuP 753 and KT3-671, respectively. In addition, KT3-671 abolished the inhibitory effect of CV- 11974 in the rat aorta but not in the rabbit aorta. Indomethacin (10(-5) M) or the removal of endothelium did not affect the inhibitory effect of Ang II by CV-11974 or EXP 3174 but enhanced the insurmountable antagonism by KT3-671. ODQ (3 x 10( 6) M), N(G)-nitro-L-arginine (3 x 10(-4) M), 4-aminopyridine (3 x 10(-3) M), tetraethylammonium (TEA; 10(-3) M), or iberiotoxin (10(-7) M) did not affect the inhibitory action of KT3-671 or CV-11974. Methylene blue (3 x 10(-6) M), KCl (10(2) M), TEA (10(-2) M), or BaC12 (10(-4) M) changed the insurmountable antagonism by KT3-671 to surmountable antagonism and abolished the inhibitory effect of CV-11974. However, glibenclamide (3 x 10(-6) M) did not affect the inhibitory action of KT3-671 but reduced the insurmountable antagonism by CV- 11974. These results indicate that KT3-671 is an insurmountable antagonist of Ang II in the rabbit and rat aorta. The results in the rat aorta also suggest that K(ATP) channels may be involved in insurmountable antagonism of Ang II by KT3-671 and CV-11974. Key Words: KT3-671-Rabbit-Rat-Vascular smooth muscle-Angiotensin II Insurmountable antagonist-K(TP)channels. PMID- 10710134 TI - Increased endothelium--monocyte interactions in salt-sensitive hypertension: effect of L-arginine. AB - Although adhesion of monocytes to endothelial cells is considered as one of the initial factors leading in the long term to the development of atherosclerosis, the effects of hypertension on monocyte-endothelial cell interactions are still largely unknown. Thus we evaluated whether hypertension affects adhesion of monocytes on rat carotid endothelium, and whether this adhesion may be modified by long-term treatment with L-arginine, the physiologic precursor of nitric oxide (NO). Hypertension was induced in Dahl rats using a sodium-rich diet (8%), in the absence or the presence of L-arginine (1.25 g/L in drinking water). After 1 month, the carotid arteries were isolated, opened longitudinally, and incubated in the presence of 2 x 10(6) monocytes previously rendered fluorescent by incubation with tetramethyl rhodamine isothiocyanate (TRITC), and adherent cells were counted under fluorescence microscopy. In parallel, the production of NO was evaluated in vitro in isolated aorta and isolated hearts. Hypertension markedly increased adhesion of monocytes on carotid endothelium, and this was reduced by L arginine. Hypertension also reduced an index of NO release at the level of the aorta and the coronary circulation. This impaired release of NO was partially prevented by L-arginine. Thus hypertension was associated with an increased adhesion of monocytes, which is probably due at least in part to a decreased production of NO. The increased adhesion was partly reduced by L-arginine, possibly secondary to an increased production of NO. Such an increased adhesion of monocytes may contribute the increased cardiovascular risk in hypertension. PMID- 10710135 TI - Inhibition of the cardiac p38-MAPK pathway by SB203580 delays ischemic cell death. AB - We report that SB203580 (SB), a specific inhibitor of p38-MAPK, protects pig myocardium against ischemic injury in an in vivo model. SB was applied by local infusion into the subsequently ischemic myocardium for 60 min before a 60-min period of coronary occlusion followed by 60-min reperfusion (index ischemia). Infarct size was reduced from a control value of 69.3 +/- 2.7% to 36.8 +/- 3.7%. When SB was infused systemically for 10 min before index ischemia, infarct size was reduced to 36.1 +/- 5.6%. We measured the content of phosphorylated p38-MAPK after systemic infusion of SB and Krebs-Henseleit buffer (KHB; negative control) and during the subsequent ischemic period using an antibody that reacts specifically with dual-phosphorylated p38-MAPK (Thr180/ Tyr182). Ischemia with and without SB significantly increased phospho-p38-MAPK, with a maximum reached at 20 min but was less at 30 and 45 min under the influence of the inhibitor. The systemic infusion of SB for 10 min before index ischemia did not significantly change the p38-MAPK activities (compared with vehicle, studied by in-gel phosphorylation) < or =20 min of ischemia, but activities were reduced at 30 and 45 min. Measurements of p38-MAPK activities in situations in which SB was present during in-gel phosphorylation showed significant inhibition of p38-MAPK activities. The systemic infusion of SB significantly inhibited the ischemia induced phosphorylation of nuclear activating transcription factor 2 (ATF-2). Using a specific ATF-2 antibody, we did not observe significant changes in ATF-2 abundance when nuclear fractions from untreated, KHB-, and SB-treated tissues were compared. We investigated also the effect of local and systemic infusion of SB on the cardioprotection induced by ischemic preconditioning (IP). The infusions (local or systemic) of SB before and during the IP protocol did not influence the infarct size reduction mediated by IP. The observed protection of the myocardium against ischemic damage by SB points to the negative role of the p38-MAPK pathway during ischemia. PMID- 10710136 TI - The angiotensin-converting enzyme gene polymorphism and responses to angiotensins and bradykinin in the human forearm. AB - The deletion (D) allele of the angiotensin-converting enzyme (ACE) is associated with high ACE levels. Subjects homozygous for the D allele should therefore exhibit enhanced angiotensin I-induced vasoconstrictor responses and diminished bradykinin-induced vasodilator responses as compared with subjects homozygous for the insertion (I) allele. In eight II and eight DD normotensive male subjects, angiotensin I, bradykinin, and angiotensin II were infused in the forearm. Changes in forearm blood flow were registered with venous occlusion plethysmography. Blood was sampled to quantify angiotensin I to II conversion. Plasma ACE levels were 60% higher, and DD subjects showed an enhanced response to angiotensin I infusion (p < 0.05). No differences in angiotensin I to II conversion, angiotensin H vasoconstriction, and bradykinin vasorelaxation were found. The ACE-inhibitor enalaprilate inhibited angiotensin I-induced vasoconstriction, but did not significantly affect bradykinin-induced vasodilation. The AT1-receptor antagonist losartan (3,000 ng/kg/min) inhibited angiotensin II-induced vasoconstriction. In conclusion, subjects with the DD genotype display an enhanced vasoconstrictor response to angiotensin I, which cannot be explained on the basis of a similarly enhanced angiotensin I to II conversion rate or a difference in vascular reactivity. Possibly therefore, differences in angiotensin I to II conversion occur within the vascular wall only, at a site that does not readily equilibrate with blood plasma. PMID- 10710137 TI - Preconditioning of coronary artery against vasoconstriction by endothelin-1 and prostaglandin F2alpha during repeated downregulation of epsilon-protein kinase C. AB - The cellular mechanisms of coronary vasospasm are unclear, and a role for protein kinase C (PKC) activation by the endogenous vasoconstrictors endothelin-1 (ET-1) and prostaglandin F2alpha (PGF2alpha) has been suggested. In this study, we developed a phorbol ester-induced PKC downregulation protocol to investigate the relation between the amount and activity of specific PKC isoforms in coronary arterial smooth muscle and coronary vasoconstriction by ET-1 and PGF2alpha. Isometric tension was measured in deendothelialized porcine coronary artery strips, [Ca2+]i was monitored in single coronary smooth muscle cells loaded with fura-2, and the whole tissue, cytosolic, and particulate fractions were examined for PKC activity and reactivity with isoform-specific anti-PKC antibodies using Western blot analysis. In Ca(2+)-free (2 mM EGTA) Krebs solution, ET-1 (10(-7) M), PGF2alpha (10(-5) M) and PKC activator phorbol 12,13-dibutyrate (PDBu) (10( 6) M) caused significant contractions that were completely inhibited by the PKC inhibitors staurosporine and calphostin C, no significant change in [Ca2+]i, and significant activation and translocation of the Ca(2+)-independent epsilon-PKC but not the Ca(2+)-dependent alpha-PKC. In Ca(2+)-free Krebs, a single application of PDBu produced maximal contraction and PKC activity after 30 min, which declined to basal levels in 3 h and remained steady for 24 h, but did not prevent subsequent increases in contraction and PKC activity with a new addition of PDBu and did not significantly decrease the amount of alpha- or epsilon-PKC. Repeated (five to eight) applications of PDBu in Ca(2+)-free Krebs at 3-h intervals completely inhibited subsequent increases in contraction and PKC activity to PDBu, ET-1, or PGF2alpha, and significantly decreased the amount of epsilon-PKC but not that of alpha-PKC. These results provide evidence that a Ca(2+)-independent coronary vasoconstriction induced by ET-1 and PGF2alpha is associated with activation of the epsilon-PKC isoform. The results suggest that, in coronary artery smooth muscle, downregulation of PKC is isoform specific and is more dependent on the frequency rather than the duration of PKC activation. The results also suggest that repeated downregulation of epsilon-PKC might play a role in preconditioning of the coronary artery against vasoconstriction by ET-1 and PGF2alpha. PMID- 10710138 TI - Effect of stable fish oil on arterial thrombogenesis, platelet aggregation, and superoxide dismutase activity. AB - We examined the influence of dietary stable fish oil on aortic thrombosis, platelet aggregation, and superoxide dismutase (SOD) activity in a rat model. Twenty-nine Sprague-Dawley rats were fed regular chow supplemented with stable fish oil preparation (for 1 or 3 weeks), and 37 rats fed regular chow served as controls. The abdominal cavity was opened, and the abdominal aorta isolated. Whatman paper impregnated with 35% FeCl3 was wrapped around the surface of the aorta, and aortic flow was continuously recorded. In control rats, an occlusive platelet-fibrin-rich thrombus was formed in 21 +/- 3 min. Dietary fish oil in a time-dependent fashion delayed time to thrombus formation (24 +/- 2 min in rats fed fish oil for 1 week and 31 +/- 2 min in rats fed fish oil for 3 weeks), inhibited platelet aggregation (21 +/- 5% vs. 45 +/- 6%; p < 0.01) and increased SOD activity (p < 0.01). We conclude that dietary supplementation with stable fish oil delays formation of arterial thrombus, probably by reducing platelet aggregation and oxidative stress-associated arterial injury. PMID- 10710139 TI - Estrogen modulates a large conductance chloride channel in cultured porcine aortic endothelial cells. AB - Estrogen is known to exert a protective effect on cardiovascular disease, but the mechanism for this effect is unclear. It has, however, been reported that estrogen and antiestrogen modify ionic currents and membrane potential in various cells. The aim of this study was to clarify whether the chloride channel of aortic endothelial cells was, in fact, modified by estrogen and antiestrogen with inside-out patch and cell-attached patch recording methods. Tamoxifen activated a large-conductance (368 +/- 23 pS, n = 6, in symmetric 150 mM Cl- solution) chloride channel of endothelial cells grown in the presence of 1 microg/ml colchicine. The channels were activated mainly between +/-40 mV, but were inactivated at more extreme potentials. The open probability of channels in cell attached patches increased from <0.01 to 0.37 +/- 0.08 (n = 8) when cells were treated with 15 microM tamoxifen. This effect can be blocked by 17beta-estradiol, but not by progesterone. The results showed that tamoxifen increased chloride channel activity in the presence of colchicine in cultured endothelial cells, and this action was suppressed by 17beta-estradiol but not by progesterone. This rapid effect by estrogens suggests that these hormones exert nongenomic, short term activity and do not appear to affect the nuclear estrogen receptor. With these effects, estrogen and antiestrogen bind to the endothelial cells plasma membrane site and subsequently may activate an intracellular second messenger pathway. PMID- 10710140 TI - Increases in intracellular calcium of arterial smooth muscle cells in transgenic mice overexpressing Na+/H+ exchanger. AB - We produced transgenic mice overexpressing Na+/ H+ exchanger as a model of salt sensitive hypertension and reported that dietary salt loading elevates blood pressure in these transgenic mice. We speculate that this blood pressure elevation may be attributed to the elevation of intraarterial smooth muscle Ca2+ concentration through Na+/Ca2+ exchange. To test this hypothesis, we measured the isometric tension of aortic rings and intracellular free calcium ([Ca2+]i) of cultured smooth muscle cells. In the transgenic mice, the aortic ring contraction induced by 5 mM caffeine (percentage of 60 mM K-induced contraction) was significantly greater than control mice (60.1 +/- 5.5% vs. 44.8 +/- 3.1%). The mean [Ca2+]i in vascular smooth muscle cells (VSMCs) of transgenic mice (123.1 +/ 19.7 nM) was higher than those in VSMCs of control mice (66.6 +/- 7.2 nM). These observations suggest that dietary salt loading increases the concentration of calcium in arterial smooth muscle cells in this transgenic mice. These findings are helpful in tracing the causes of salt-sensitive hypertension. PMID- 10710141 TI - Anterior cervical diskectomy and fusion without plate instrumentation in 178 patients. AB - Between 1989 and 1996, fusion, pseudarthrosis, repeated operation rates, and outcomes were studied in 178 patients undergoing one- to four-level (average, 2.2 levels) anterior cervical diskectomy and fusion (ADF) without plating. Dynamic radiographs taken 3 and 6 months after operation showed fusion or pseudarthrosis without motion in 99% of patients after one-level ADF (78 patients), in 90% after two-level ADF (84 patients), and in 100% after three-level ADF (12 patients) and four-level ADF (4 patients). Pseudarthrosis with motion was noted in 1% after one level ADF and in 10% after two-level ADF (statistically significant with a lower pseudarthrosis rate in the 1-level; by Fisher's exact test, p = 0.0351). Three patients required secondary posterior wiring and fusion. Good or excellent outcomes (by Odom's criteria) were achieved in 96% of patients within an average of 82 months. Although fusion rates for one-level ADF without plates appear adequate, high pseudarthrosis rates after two-level ADF warrant that plating be considered. PMID- 10710142 TI - The value of anterior cervical plating in preventing vertebral fracture and graft extrusion after multilevel anterior cervical corpectomy with posterior wiring and fusion: indications, results, and complications. AB - Anterior cervical plates were added to anterior corpectomy and fusion (ACF) with posterior wiring and fusion (PWF) to prevent vertebral fracture and graft extrusion in patients with ossification of the posterior longitudinal ligament and spondylostenosis. From January 1989 to March 1997, 22 patients had an average 2.5-level ACF without plates and an average 5-level PWF with halo placement (average follow-up, 4 years). From April 1997 to October 1998, 22 patients had an average 2.8-level ACF with Orion plating and an average 5.4-level PWF with halo devices (patients were followed for an average of 11 months). Vertebral fracture and graft extrusion requiring revision developed in three (14%) patients without plates within 24 hours of surgery, whereas neither of these occurred in patients with plates (by Fisher's test: nonsignificant p value = 0.2326). Ultimately, all 44 patients had fusion. Thus far, vertebral fractures and graft extrusions have not been observed for 22 patients undergoing plated circumferential cervical surgery. PMID- 10710143 TI - Anterior transvertebral herniotomy for cervical disk herniation. AB - This study analyzed the results of anterior transvertebral herniotomy for cervical disk herniation to assess the utility of this procedure. Anterior transvertebral herniotomy was performed in 24 patients who had cervical disk herniation without spinal canal stenosis. In most patients, a good result was obtained, but simultaneous or subsequent anterior intervertebral fusion was necessary in four patients. In one (4%) patient, the two adjacent vertebrae had fused spontaneously. The best indication for this treatment judging from the postoperative results is a large disk hernia associated with either myelopathy or radiculopathy, but without spinal canal stenosis. PMID- 10710144 TI - Ossification of the ligamentum flavum as a cause of myelopathy in North America: report of three cases. AB - Myelopathy caused by ossification of the ligamentum flavum is a rare condition in North America. The authors describe three patients whose myelopathy was attributed to posterior cord compression warranting laminectomy to decompress the cervical spine (in one patient) and the thoracic spine (in two patients). The spinal computed tomographic scan (especially after myelography) can be instrumental in guiding the management of this condition. PMID- 10710145 TI - Cervical curvature after laminoplasty for spondylotic myelopathy--involvement of yellow ligament, semispinalis cervicis muscle, and nuchal ligament. AB - To assess the consequences of cervical laminoplasty on postoperative lordosis, a retrospective radiographic analysis of 31 patients undergoing laminoplasty for spondylotic myelopathy was completed. Special attention was paid to lordotic changes occurring at each level over more than 2 years. Preoperative lordosis remained unchanged with the patients wearing a cervical orthosis 1 week postoperatively. However the lordosis subsequently demonstrated a significant decrease in 87% of patients over an average of 3.1 years. Lordotic alignment at C2-C3 and C6-C7 before surgery significantly decreased in 81% and 58% of patients 1 week postoperatively, and 84% and 81% at last follow up, respectively, while lordotic alignment at other levels pre- and postoperatively did not significantly change. Loss of lordotic alignment was largely attributed to detachment of semispinalis cervicis muscle on C2 and nuchal ligament on C6/C7 with a posterior approach and/or section of yellow ligament at C2-C3. PMID- 10710146 TI - Spontaneous interbody fusion after controlled injuries to the spine: an experimental study in rabbits. AB - To evaluate the rationale of spontaneous spine fusion after a spinal injury, the authors conducted an experimental study that consisted of three types of controlled injuries to a rabbit spine model. The first was injury to the intervertebral disk (type I injury). The second was injury of the intervertebral disk along with injury to one of the adjacent vertebral end plates (type II). In type III injury, both the opposing end plates were injured along with the intervertebral disk. In 38 rabbits, a total of 82 injuries of these three types were inflicted. Twenty-six injuries were of type I (n = 22 rabbits), 26 were type II (n = 24 rabbits), and 30 were type III (n = 26 rabbits). Spontaneous fusion occurred only in type III injuries. From the 30 type III injuries, fusion occurred in 20 (66.6%). For an autofusion to occur, both epiphyseal plates may be injured. In the clinical situation, this observation suggests that a radiographically obscure lesion of both neighboring vertebrae may proceed to autofusion of that spinal segment observed later. PMID- 10710147 TI - Acute cervical spinal cord injury secondary to air bag deployment without proper use of lap or shoulder harnesses. AB - The authors present a case report of a patient with cervical central spinal cord syndrome caused by a hyperextension injury after a motor vehicle collision in which the air bag deployed in the absence of shoulder or lap belt harnesses. The potential for cervical spine and spinal cord hyperextension injuries in passengers positioned in front of air bags without proper use of shoulder or lap belt harnesses is discussed. Cervical central spinal cord quadriplegia occurred with cervical spondylosis and kyphosis that was managed by early three-level cervical corpectomy in a 58-year-old patient. Early improvement in the patient's neurological status occurred but was incomplete at the time of this report. Cervical hyperextension injuries are possible in passengers positioned in the front seat of cars with air bags when shoulder or lap belt harnesses are not used properly. Previous biomechanical studies have documented the potential for these types of injuries. PMID- 10710148 TI - Subarachnoid drainage of an established or chronic pseudomeningocele. AB - Incidental dural tears are a common complication of lumbar spine surgery, but if unrecognized cerebrospinal fluid (CSF) leakage can lead to the formation of a fistula or pseudomeningocele. Traditionally, fistulae and pseudomeningoceles have been treated with open revision surgery, but acute fistulae with direct CSF leakage through the incision site have reportedly been treated by extracorporal drainage. This report presents a case in which an established or chronic pseudomeningocele was successfully managed without open surgical repair. PMID- 10710149 TI - Postoperative deep wound infection in adults after posterior lumbosacral spine fusion with instrumentation: incidence and management. AB - The authors reviewed 817 instrumented lumbosacral fusions in adults and found an incidence of 3.2% deep wound infections. The primary focus of this study was the management of these infections, with particular attention to whether the implants needed to be removed. A consulting infectious disease specialist indicated that an acute infection of a low back fusion wound could not be healed without removal of the metallic implants. This opinion was in contrast to the authors' daily experience and prompted this study. The authors identified and reviewed 817 cases of instrumented posterior lumbosacral arthrodeses in adults. A detailed analysis of any case with a deep wound infection was performed and yielded and infection rate of 3.2% (26 patients). Of these, 24 achieved a clean, closed wound without removal of instrumentation through a protocol of aggressive debridement and secondary closure. Instrumentation removal is not necessary to obtain a clean, closed wound using an aggressive approach with early diagnosis, vigorous debridement in the operative room under general anesthesia, delayed primary or secondary closure, and appropriate antibiotic coverage. PMID- 10710150 TI - Delayed postoperative epidural hematoma formation after heparinization in lumbar spinal surgery. AB - Treatment of thromboembolic disease in the postoperative lumbar spine patient is controversial. This case report describes an epidural hematoma with neurologic sequelae in an elderly patient who received intravenous heparin therapy over 2 weeks after lumbar decompression. Implications for treatment of thromboembolic disease in the postoperative lumbar spine is reviewed. PMID- 10710151 TI - Factor analysis of the effectiveness of transfixation and rod characteristics on the TSRH screw-rod instrumentation. AB - The effects of Texas Scottish Rite Hospital (TSRH) hardware parameters (rod length and diameter and cross-link) and their interaction on the stiffness of the TSRH pedicle screw-rod construct were evaluated. Four TSRH screws were assembled in pairs to two polymethyl-methacrylate blocks to resemble a one-level or more corpectomy model and the construct underwent nondestructive torsional, extension, and flexion loading. In every loading test, each construct was tested using TSRH rods of different lengths (10, 15, and 20 cm) and diameters (4.9 and 6.5 mm) and different cross-links (TSRH and two new types made for this experiment). We compared the stiffness of the construct without cross-linking with that with single or double TSRH cross-linking, or either the closed new-type cross-link (closed NTC) or the open new-type cross-link (open NTC) using factor analysis. There was no axial slipping of one rod versus the other up to a force of 100 kg. The stiffness of the construct in all three loading modes increased as the rod length decreased, the rod diameter increased, and the construct was augmented with a cross-link. The closed NTC provided the greatest stiffness and the single TSRH provided the least stiffness. Unaugmented 10-cm-long rods showed two or three times more torsional stiffness than did that of the longer unaugmented rods independent of rod diameter. In addition, the closed NTC offered the maximal increase in flexion stiffness of the construct with thick rods and 10-, 15-, and 20-cm-long rods at a maximum of 40%, 27%, and 30%, respectively. This rigid closed NTC increased the extension stiffness of the same construct with 10- and 15-cm-long rods at 40% and 6%, respectively, whereas it had no influence on the extension stiffness of 20-cm-long rods. PMID- 10710152 TI - The biomechanical significance of anterior column support in a simulated single level spinal fusion. AB - This study examines the biomechanical effects of interbody cages and variations in posterior rod diameter in a simulated single-level spinal fusion. A single level spinal fusion model composed of polyethylene cylinders, posterior pedicular instrumentation, and variously positioned single or dual interbody cages was used for biomechanical testing. Constructs were tested under compressive flexural load, with measurement of stiffness, rod strain, cage strain, and intracage pressure. A strong linear correlation emerged between the mean construct stiffness and cage positioning within the sagittal plane that was inversely related to posterior rod strain. Two small titanium mesh cages were equivalent to one large cage. In a single-level spine model, the presence of and sagittal position of interbody cages significantly influences overall construct stiffness. Cage strain increased with more anterior positions and was inversely related to rod strain. PMID- 10710153 TI - Surgical anatomy of the cervical pedicles: landmarks for posterior cervical pedicle entrance localization. AB - The posterior entrance to the cervical pedicle is described using quantitative and descriptive parameters. Fifty-three spines (C2-C7) were evaluated using a digital caliper and by visual inspection using four bony landmarks: the lateral vertebral notch and inferior articular process (C2-C7), the medial pedicle cortex at C2, and the transverse process at C7. Three distances were defined. (1) At C2, the average medial pedicle cortex-pedicle distance was 7.2 mm. (2) The lateral vertebral notch-pedicle distances showed that the entrances were located close to the notch at C2, almost at the notch at C3 and C4, and gradually moved medially away from the notch from C5 to C7. The pedicles were rarely located lateral to the lateral vertebral notch. (3) The inferior articular process-pedicle distance was large at C2, the shortest at C3, and gradually increased toward C7. Three relations were defined. (1) The pedicles were located mostly in the intermediate third of the inferior facet at C2; in the lateral third at C3, C4, and C7; or in the lateral or intermediate thirds at C5 and C6. Only C2 and C6 pedicles were located in its medial third. (2) The pedicles were located mostly below the lateral vertebral notch at C2, at C3-C6, or almost equally above and at the notch at C7. (3) Most of the C7 pedicles were located below the midline of the transverse process. The location of the pedicle entrance was unique at each cervical level. Their distribution followed the cervical spinal cord enlargement. These landmarks should assist with safe placement of pedicle screws. PMID- 10710154 TI - Psoas abscess secondary to discitis: a case report of conservative management. AB - We report a case of secondary psoas abscess in a 37-year-old man with a 3-week history of severe low backache managed conservatively without surgical drainage. Apart from bilaterally restricted straight leg raising (<70 degrees), his neurologic examination was within normal limits. Magnetic resonance imaging showed discitis of the L3-L4 space and a left-sided secondary psoas abscess. Aspiration biopsy of the abscess material under radiologic control isolated Staphylococcus aureus, which responded to appropriate antibiotic therapy with complete resolution. A high index of suspicion is necessary for diagnosis of psoas abscess, which should be considered in patients with pyrexia and backache with a neurologic examination that is otherwise normal. We discuss the recommendations for surgical and nonsurgical approaches. PMID- 10710155 TI - Esophageal perforations after anterior cervical surgery. AB - An esophageal perforation after anterior cervical surgery is an uncommon but well recognized complication. During the past 25 years, 44 patients have presented to Craig Hospital (Rocky Mountain Regional Spinal Injury Center) with esophageal perforations; this is the largest series reported to date. There were 34 patients whose esophageal injury was related to the operations performed for cervical fractures, of which 28 patients had plate and screw fixation. The most frequently occurring clinical symptoms were that of neck and throat pain, odynophagia, dysphagia, hoarseness, and aspiration. The most common clinical findings were an elevated temperature, localized induration and neck tenderness, crepitus or subcutaneous air in the neck and anterior chest wall, an unexplained tachycardia, and blood in the nasogastric tube. Imaging studies indicated an esophageal injury in only 32 (72.7%) patients. Twenty-two patients experienced cervical osteomyelitis or an abscess of the neck. Nonoperative treatment is fraught with a high mortality, and 42 patients required surgical repair of their esophageal injury. The length of hospital stay averaged 253 days. Successful management of esophageal perforations depends on the physicians' awareness of the causes, prompt recognition of the symptoms and clinical findings, and immediate institution of treatment. PMID- 10710156 TI - Paraplegia after epidural anesthesia in a patient with peripheral vascular disease: case report and review of the literature with a description of an original technique for hematoma evacuation. AB - Epidural hematoma after epidural anesthesia is a rare and uncommon complication in patients with peripheral vascular disease who require perioperative anticoagulation therapy. A low index of suspicion makes its diagnosis difficult and often delayed. Treatment usually involves extensive laminectomy, increasing the chances for patient complications. In this article, the authors report a case of epidural hematoma with secondary paraplegia after epidural anesthesia. Also described is an original technique for evacuating the epidural space. PMID- 10710157 TI - Lumbar facet joint morphology. PMID- 10710158 TI - Variability of human brain structure size: ages 4-20 years. AB - Understanding variability of human brain structure sizes during development is important for the design and interpretation of pediatric neuroimaging studies. In this study we analyze the effects of hemisphere, sex and age on size variability of the total cerebrum, cerebellum, lateral ventricles, temporal lobe, amygdala, hippocampus, superior temporal gyrus, corpus callosum, caudate, putamen, and globus pallidus in 115 healthy children and adolescents, ages 4-20 years. Variability differed significantly across structures, with the lateral ventricles demonstrating the highest coefficient of variation and the putamen the lowest. Males varied significantly more than females in the left cerebrum and left superior temporal gyrus, whereas females varied more than males in the right caudate and right putamen. Age effects were seen in increased variability after puberty for the lateral ventricles, hippocampus and superior temporal gyrus. These variances are important determinants of minimum sample sizes required to detect group differences in both cross-sectional and longitudinal studies. PMID- 10710159 TI - Functional magnetic resonance imaging in schizophrenia: initial methodology and evaluation of the motor cortex. AB - The purpose of the present study was to evaluate the differential activation of the motor cortex during finger tapping in patients with schizophrenia using the newly available imaging method of functional magnetic resonance imaging (fMRI). Nine patients with DSMIII-R schizophrenia and 9 well-matched healthy volunteer subjects underwent fMRI examination on a conventional MR unit; activation of the primary motor cortex was evaluated during performance of a finger motion task. Localized activation of the motor cortex was observed in 17 of 18 subjects during fMRI. Patients and controls were, however, indistinguishable with respect to signal intensity or area thereof within the motor cortex. fMRI did not reveal motor cortical dysfunction in schizophrenia. Despite its infancy, fMRI holds considerable promise to advance understanding of the neurodynamics of psychiatric disorders, particularly schizophrenia. PMID- 10710160 TI - Caudate regional cerebral blood flow in obsessive-compulsive disorder, panic disorder and healthy controls on single photon emission computerised tomography. AB - We compared regional cerebral blood flow (rCBF) in 15 patients with DSM IIIR obsessive-compulsive disorder (OCD), 15 patients with DSM IIIR panic disorder and 15 healthy controls matched for age, sex and hand preference, using uptake of technetium-99m-D,L-hexamethyl-propylene amine oxime (99mTc HMPAO), on single photon emission computerised tomography (SPECT). Caudate rCBF was significantly reduced in OCD patients compared to healthy subjects and panic disorder patients. When four patients were excluded from each group, right caudate rCBF remained significantly lower in OCD patients than in panic disorder patients or healthy subjects. The data suggest functional involvement of the right caudate nucleus is present in OCD. PMID- 10710161 TI - The continuous performance test, identical pairs version (CPT-IP): III. Brain functioning during performance of numbers and shapes subtasks. AB - The numbers and shapes subtasks of the CPT-IP are difficulty-matched measures of independent aspects of attentional skill that have been used to differentiate the impairments of schizophrenics and major depressives. Previous studies suggest that they tap into lateralized aspects of attentional performance. To investigate this hypothesis, seven subjects free of psychiatric illness were presented with these CPT-IP subtasks during a SPECT procedure. Subtasks--4-digit number strings and nonsense shapes--were administered on successive weeks, in counterbalanced order, simultaneous with administration of 10 mCi 99mTc HMPAO. Scanning took place after 10 min of test performance. Quantitative data were extracted from each scan via a semi-automated region of interest (ROI) analysis which defined eight cortical and four subcortical ROI on each of five transverse slices. Data for each ROI were normalized and compared between task conditions. Results indicate that the two tasks produce different patterns of functioning within two general areas of the brain. First, during Numbers task performance, left-sided activity was increased on multiple transverse slices in an anterior subcortical region that incorporated the anterior cingulate, frontal white matter, and much of the basal ganglia. Left-sided activity was also increased in a posterior subcortical region including the left side of the thalamus. Lateralization of function, defined as relative activity on the left and right sides, changed within these regions across tasks, but primarily as a result of the contribution of increased or decreased activity on the left side alone. Second, relative perfusion to occipital regions, bilaterally, was more extensive during the Shapes task. These results suggest that subtle alterations in stimulus parameters can affect activation patterns in regions that are critically associated with task performance. They also suggest that the Numbers task may provide more robust activation of anterior attention systems, that may better discriminate the functioning of these systems in normal and psychopathological groups. PMID- 10710162 TI - Clinical symptoms of schizophrenia affect reference-independent measures of task induced EEG alpha asymmetry. AB - Differential patterns of EEG alpha asymmetry during verbal and spatial cognitive activity are commonly described and are thought to reflect predominance of left- vs. right-sided cortical activation. Although these patterns have been difficult to elicit reliably in schizophrenics, the authors have previously suggested that clinical status may have confounded results. Therefore, EEG data from 17 additional schizophrenic patients, 16 mood disorder patient controls, and 17 normal controls were used to examine the relationship between severity of schizophrenic symptoms and task induced alpha asymmetry. Subjects performed verbal and spatial tasks during recording of 16-channel EEG. After transformation to the average reference, log alpha power from central and temporal leads was analyzed by MANOVA and MANCOVA. The expected task-side interaction (P < 0.02) was present for the total sample and for each control group when analyzed alone. However, it was only present in schizophrenics when the BPRS score was entered as a covariable. Patterns of correlations between BPRS scores, left temporal alpha power, right/left temporal alpha ratios, and task differences in ratios suggested that higher symptom levels were associated with excessive left-sided activation during spatial activity. This is consistent with other evidence of left hemisphere overactivity in schizophrenics. PMID- 10710163 TI - Proton magnetic resonance spectroscopy of the lenticular nuclei in bipolar I affective disorder. AB - Proton magnetic resonance spectroscopy (1H-NMS) was used to examine the ratio of choline-containing compound (Cho) to creatine (Cr) in the basal ganglia. Subjects comprised 10 bipolar I affective disorder patients and 10 healthy control subjects. No significant difference was found in the Cho/Cr, N-acetyl-aspartate (NAA)/Cr, or NAA/Cho ratios between bipolar patients and control subjects. Within the bipolar group, negative correlations emerged between the NAA/Cr ratio in the right lenticular nuclei and both age at onset and age at the time of study. The results suggest that a late onset of illness and older age are associated with neuronal cell loss in the right lenticular nuclei in bipolar patients. PMID- 10710164 TI - A quantitative magnetic resonance imaging study of caudate and lenticular nucleus gray matter volume in primary unipolar major depression: relationship to treatment response and clinical severity. AB - The authors investigated whether there were differences in caudate and lenticular nucleus volumes in depressed patients relative to comparison subjects, and whether differences in basal ganglia volume were associated with treatment response to fluoxetine. Brain magnetic resonance images were obtained from 38 unipolar depressed patients and 20 matched comparison subjects. Patients were divided into groups of 'responders' and 'non-responders' based on change in the 17-item Hamilton Depression Rating Scale (HDRS) score after a 10-week open trial of fluoxetine, 20 mg/day. There were no group mean differences in caudate and lenticular nucleus volumes between patients and comparison subjects. Female treatment responders tended to have larger caudate nucleus volumes than male 'responders', and also larger right caudate nucleus volumes than their female 'non-responder' counterparts. Baseline HDRS scores correlated negatively with left caudate nucleus volume in depressed patients. Thus, in mild to moderately depressed patients, we were unable to find differences in caudate and lenticular nucleus-gray matter volumes relative to comparison subjects. One possible reason is that caudate nucleus-gray matter volume and severity of depression are inversely correlated, suggesting that severity of depression may be an important covariate when comparing caudate volumes in depressed patients and control subjects. PMID- 10710165 TI - Association of brain structural change with the heterogeneous course of schizophrenia from early childhood through five years subsequent to a first hospitalization. AB - Fifty first-episode patients with schizophrenia were followed for 5 years subsequent to their first hospitalization. The course of illness was charted prospectively and premorbid childhood histories were obtained retrospectively at the initial evaluation, and MRI scans were obtained initially and at each follow up. Fifteen different life-time patterns of illness course emerged, although none were specifically associated with structural brain change. A deterioration in premorbid scores was positively correlated with larger ventricular volume at the first hospitalization, and the larger the ventricles, the less the subsequent change in ventricular size thereafter. An analysis to see whether initial hemispheric and ventricular size could predict different course types only revealed that patients with an acute onset and complete recovery had significantly smaller ventricles than all others. No differences emerged for initial hemispheric size. Thirty-four percent of patients individually showed some association of brain ventricular size and 28% hemisphere volume reductions with fluctuation in psychotic symptoms. Paradoxically, most showed larger ventricles and smaller hemispheres to be associated with clinical improvement, rather than the predicted reverse. These latter data question the notion that the structural brain changes seen over time in some patients are related to poor outcome, although small ventricular size in those patients with acute onset may be predictive of recovery. Thus, brain structural change is occurring early in the course of illness and may be a consequence of the process leading to resolution. PMID- 10710166 TI - Is there a relationship between delta sleep at night and afternoon cerebral blood flow, assessed by HMPAO-SPECT in depressed patients and normal control subjects? Preliminary data. AB - We wished to explore the relationships between waking HMPAO uptake and visually scored polysomnography. We hypothesized that HMPAO activity would correlate positively with slow wave sleep measures the same night. Eight unmedicated unipolar patients with current DSM-IV major depression (17-item Hamilton Depression Rating Scale score 21.5+/-2.9) and seven control subjects received polysomnography on 2 consecutive nights. On the afternoon following the adaptation night, subjects received cerebral SPECT, with 15 mCi Tc-99m-HMPAO injected while subjects performed the Continuous Performance Task. Patients and control subjects did not significantly differ on demographic, polysomnographic, and SPECT variables. Slow wave sleep measures correlated positively (Spearman's) with global and regional tracer activity for depressed (n = 8), control (n = 7) and combined groups (n = 15); in other words, the greater the global or regional afternoon HMPAO uptake, the greater the slow wave sleep measures were the same night. In addition, the greater the waking afternoon global or regional HMPAO activity, the faster subjects fell asleep and the less Stage 2% they had. In patients, global and regional HMPAO activity correlated positively with REM density. Positive correlations between waking tracer activity and subsequent slow wave measures are consistent with previous hypotheses linking slow wave sleep with brain energy conservation and restoration. Further study is needed to determine whether these functional relationships differ in depression. PMID- 10710167 TI - Brain function in spider phobia. AB - Measurements of regional cerebral blood flow (rCBF) were performed in 16 women suffering from spider phobia. The non-invasive 133Xe inhalation method, giving information about the blood flow of superficial areas, was used. The subjects were studied under three conditions: during resting, when exposed to a videotape showing nature scenery, and finally when watching a video with living spiders. During the rCBF measurements the subjects' behaviour was registered systematically and respiration, blood pressure, Pco2, and heart rate were monitored. Eight subjects who showed and reported severe panic during the spider exposure had marked rCBF decreases in frontal areas, especially in the right hemisphere. The remaining eight subjects displayed a more efficient control of their emotions and became frightened, but not panic-stricken, during the spider exposure. These showed a consistent rCBF increase in the right frontal area compared to neutral stimulation. Thus, results revealed significant functional changes in the frontal cortex in subjects with spider phobia during phobogenic exposure. It seems likely that these frontal changes are related to the experience and control of phobic anxiety. PMID- 10710168 TI - A method of basal forebrain anatomical standardization for functional image analysis. AB - Functional as well as structural assessment of the basal forebrain has mostly focused on the dorsal caudate and putamen in axial slices where they are easily outlined or their centers located with stereotaxic methods. The more ventral extent of the basal forebrain, where the irregular form and indistinct boundaries of the nucleus accumbens and substantia innominata are difficult to trace and where the brain's ventral surface may contribute partial volume artifacts to measurement, has been less studied. We present a method based on coronal sections, landmarks placed on clearly visible anchor points, and the computational technique of thin-plate spline warping which allows the alignment of groups of individuals to common coordinates for pixel-by-pixel statistical mapping. The reliability of the landmarks across independent raters yields a median absolute difference of 1.3-1.6 mm. The validity of the method is confirmed by variance maps which reveal significant decreases in variance over spindle and bounding box alignment. PMID- 10710169 TI - Platelet paroxetine binding in post-traumatic stress disorder. AB - Sixty-two subjects, 45 with post-traumatic stress disorder (PTSD) and 17 healthy control subjects, were examined in a study of serotonin function measured by [3H]paroxetine binding to platelet membranes. Subjects were selected from male combat exposed veterans. The mean (+/- S.D.) Kd was 0.078 +/- 0.045 nM for the PTSD patient group and 0.064 +/- 0.037 nM for the control group. The mean Bmax was 934 + 238 fmol/mg protein for the PTSD patient group and 1011 +/- 363 fmol/mg protein for the control group. There was no significant difference between the groups for either Kd or Bmax before or after controlling for season of sampling. There were no significant differences between subjects with current PTSD and those with PTSD in the past, or between PTSD subjects with or without concurrent major depressive disorder. This study finds no relationship between PTSD, major depressive disorder and peripheral serotonin function measured by [3H]paroxetine binding to blood platelets. PMID- 10710170 TI - Saccadic eye movements in schizophrenic patients. AB - The nature of saccadic abnormalities in schizophrenia was investigated in three different paradigms: (1) the visually guided saccade; (2) the antisaccade; and (3) the remembered saccade paradigm. Subjects comprised 14 schizophrenic patients and 14 normal volunteers. Deficits in the schizophrenic group were observed in the antisaccade and remembered saccade tasks, both of which were characterized by increased latency and reduced gain. Moreover, in the antisaccade task, schizophrenic patients showed an increased number of errors compared with control subjects. Saccadic abnormalities in the patients were correlated with impaired performance on the Wisconsin Card Sorting Test. These data suggest that schizophrenic patients have difficulty in inhibiting reflexive saccades and in producing voluntary saccades. The implications of these findings for a prefrontal cortex dysfunction involved in oculomotor control in schizophrenia are discussed. PMID- 10710171 TI - Schizophrenic syndromes and clozapine response in treatment-resistant schizophrenia. AB - Relationships between symptom profile and clozapine response were studied. Symptom scores on the Scale for the Assessment of Positive Symptoms and the Scale for the Assessment of Negative Symptoms were subjected to principal component analysis (PCA) in a group of 66 treatment-resistant schizophrenic patients, 49 of whom were treated with clozapine. Factor scores were compared among responders, non-responders and partial responders. The PCA yielded a four-factor solution, with positive symptoms, negative symptoms, cognitive disorganization and behavioral disorganization components. Cognitive and behavioral disorganization syndrome scores showed significant differences across groups. Cognitive disorganization was higher in non-responders, while behavioral disorganization was higher in partial responders. The results support the possibility of using clinical profiles to predict therapeutic response to clozapine. PMID- 10710172 TI - Superoxide anion production by neutrophils derived from peripheral blood of schizophrenic patients. AB - Evidence indicates that excess free radicals formation may occur in patients with schizophrenia. A study comparing the production of superoxide anion by peripheral blood neutrophils of 29 schizophrenic patients with that of 17 healthy volunteers detected a significant statistical increase in superoxide anion production in schizophrenic patients compared to the healthy control group. Despite the fact that oxidative mechanisms may play a role in schizophrenia, further studies are needed to define their involvement. Such studies would shed light on the etiology and pathogenesis of schizophrenia and may lead to new therapeutic approaches using antioxidants, which might partially alleviate or prevent the symptoms of schizophrenia. PMID- 10710173 TI - Effects of acute intravenous cocaine on cardiovascular function, human learning, and performance in cocaine addicts. AB - Continuous non-invasive cardiovascular monitoring in eight healthy cocaine addicts receiving intravenous cocaine (0.325 mg/kg or 0.650 mg/kg) or placebo in double-blind, randomized, cross-over fashion demonstrated significant dose dependent increases in pulse and mean arterial pressure following cocaine. Pulse and mean arterial pressure peaked 5 min post-cocaine injection and maximal response was sustained for a further 15 min and 35 min afterwards, respectively. Cocaine administration had no significant effect on peripheral oxygen saturation, and no clinically significant abnormalities of rhythm or conduction were seen on the electrocardiogram. These doses and method of single-dose intravenous cocaine administration, and our procedures for cardiovascular monitoring, appear relatively safe for laboratory studies of healthy cocaine addicts with no pre existing cardiovascular disease. In addition, cocaine-taking (0.325 mg/kg i.v. and 0.650 mg/kg i.v.) was associated with enhanced attention (i.e. increased numbers of correct responses on the Rapid Visual Information Processing Task), but the trend towards reduced reaction time did not achieve statistical significance. Cocaine-taking resulted in a small but statistically insignificant improvement in learning on the Digit Symbol Substitution Task. These results suggest that cocaine-taking in rested subjects is associated with some cognitive enhancement. PMID- 10710174 TI - The use of the Panic and Agoraphobia Scale in a clinical trial. AB - A new scale for assessing severity in PDA (Panic Disorder with/without Agoraphobia) has recently been developed: the Panic and Agoraphobia Scale [P & A (Bandelow, 1995)]. The objective of this study was to test whether the scale is sensitive to changes during a treatment trial. Thirty-seven patients (mean age, 32.7; S.D., 6.3) with PDA were treated with imipramine (75-150 mg/day) for 8 weeks in an open prospective trial. Patients with concurrent agoraphobia were instructed in practising self-exposure to agoraphobic situations. The total scores on the P & A, the Hamilton Anxiety Scale (HAMA) and the Clinical Global Impression Scale (CGI) were used as the main efficacy criteria. Treatment results were excellent, as could be shown by a decrease in the average severity scores of the P & A observer-rated version from 28.9 (S.D., 8.1) to 13.3 (S.D., 11.8; rank statistic T(N) = 6.7; P < 0.0001). The largest effect size r(w) of all clinician rated scales was seen with the observer-rated version of the P & A, although closely followed by the CGI and the HAMA. Among the self-rated scales, the P & A (self-rated version) also showed the largest effect size. All five subscores of the P & A showed significant improvements. The highest treatment effect sizes could be seen in the 'panic attacks' subscore, followed by the 'anticipatory anxiety' subscore. The new Panic and Agoraphobia Scale (P & A) is a useful tool for measuring treatment efficacy in panic disorder trials. PMID- 10710175 TI - Adaptation of a simple patient satisfaction instrument to mental health: psychometric properties. AB - The authors describe the psychometric properties of a simple patient satisfaction self-report instrument originally developed for use in primary care patients, adapted for use in mental health clinic patients of varied educational and socioeconomic backgrounds. The instrument demonstrated a single major principal component, high internal consistency reliability, high test-retest reliability, and sensitivity to change with experimental manipulations in clinical programming. The results also indicate that patient satisfaction is unidimensional in mental health patients as it is for primary care patients. PMID- 10710176 TI - Melatonin treatment of winter depression: a pilot study. AB - Five patients with winter depression received low doses of melatonin in the afternoon, and five patients received placebo capsules. Melatonin treatment significantly decreased depression ratings compared to placebo. If these findings are replicated in a larger sample with documentation of expected phase shifts, the phase shift hypothesis will be substantially supported. PMID- 10710177 TI - Interleukin-2 and schizophrenia. PMID- 10710178 TI - Live interdisciplinary teaching via the internet. AB - The independence of teachers and students is one of the main advantages of teleteaching. Specialties considered unsuitable for combined lessons are manageable using the internet. This study outlines simultaneous communication with students and lecturers over long distances between the anatomical dissection laboratory, the operating theatre, and the lecture hall. In several three directional on-line lectures, different equipment was used. Students could also participate using personal computers from other locations. During the presentations, the participants have the opportunity to discuss problems with any lecturer. It was possible to demonstrate sufficient transmission capability for real-time application with the use of the new internet technology. No important qualitative differences can be reported between: hardware and software based solutions; or commercial and free offers. Although it is often difficult to reconcile the timetable of surgeries and lectures, multimedia on-line teaching via the internet provides new potential for interdisciplinary medical education. PMID- 10710179 TI - Source of error in calculation of optical diffuse reflectance from turbid media using diffusion theory. AB - Diffusion theory and similarity relations were used to calculate the optical diffuse reflectance of an infinitely narrow laser beam incident upon a semi infinite turbid medium. The results were analyzed by comparison with the accurate results from Monte Carlo simulations. Because a large number of photon packets were traced, the variance of the results from Monte Carlo simulations was small enough to reveal the detailed defects of the diffusion theory and the similarity relations, which are broadly used in photomedicine. We demonstrated that both diffusion theory and similarity relations provide very accurate results when the photon sources are isotropic and buried more deeply than one transport mean free path in turbid media. We found that the key factor affecting the accuracy of the diffusion theory application was the conversion from the infinitely narrow laser beam to an isotropic point source in turbid media. PMID- 10710180 TI - Detection of high-frequency respiratory movements during sleep. AB - Sleep-related breathing disorders are common in adults and they have a significant impact on vigilance and quality of life. Previous studies have shown the validity of the static-charge-sensitive bed (SCSB) in monitoring breathing abnormalities during sleep. A whole nights sleep study produces a signal with considerable length, and therefore an automated analysis system would be of great need. In this work we focus on detection of high-frequency respiratory movement (HFRM) patterns which are related to increased respiratory efforts. The paper documents four methods to automatically detect these patterns. The first two are based on classical statistical tests applied to the SCSB signal, and the other two use spectral characteristics in order to adaptively segment the SCSB signal. Finally we adjust each method to detect patterns that coincide with the HFRMs determined by an expert, and evaluate the performance of the methods using independent test data. PMID- 10710181 TI - A new program to compute and evaluate continuously monitored stopping boundaries for clinical trials. AB - We present code for the calculation and evaluation of continuously monitored stopping boundaries for use in one-arm and two-arm clinical trials. These designs were first developed for one-arm trials by Thall, Simon and Estey (TSE) (P.F. Thall, R. Simon, E.H. Estey, Bayesian sequential monitoring designs for single arm clinical trials with multiple outcomes, Stat. Med. 14 (1995) 357-379). Our code corrects some problems in the original TSE algorithms and extends these algorithms for use in a two-arm trial setting. It is written in S-Plus to improve interactivity for the statistically adept user, and employs external routines, dynamically loaded into S-Plus, to improve calculation efficiency. Efficient versions of our code require both a C compiler and the S-Plus program. Our code has been tested in UNIX and Microsoft Windows environments, and compiled code is available from our website. A numerical integration routine for the convolution of beta distributions is included. PMID- 10710182 TI - Integration of a hematopoietic progenitor cell program using the ACT/DB database system. AB - Infusion of hematopoietic progenitor cells following high-dose chemotherapy is frequently used to treat patients with hematological malignancies and solid tumors. We have developed a comprehensive software system to monitor these patients once they are entered into an experimental protocol. The captured data encompasses all phases of progenitor cell therapy including progenitor cell mobilization and collection, stem cell processing, as well as cell infusion and engraftment kinetics. Particular attention was paid to the quality assurance and quality control functionality of the software during development of data entry forms and reports. The system was developed using the ACT/DB client-server database, which utilizes Microsoft Access as a front-end and accesses either an Oracle or SQL Server database. ACT/DB has been modified for deployment on the Internet in order to take advantage of Web-based technology. Information technology can help to integrate the diverse data requirements of complex therapeutic trials. PMID- 10710184 TI - Windows software for cardiac electrophysiology studies and ablation monitoring. AB - A system for cardiac electrophysiology (EP) studies consisting of a Windows software package, a standard 120 MHz Pentium PC with a high-performance video card and a data acquisition card has been developed during this study. The system is capable of real time data acquisition and storage of 24 channels with simultaneous display of 1-16 arbitrarily chosen channels at a sampling rate of 500 Hz. It can be used clinically in electrophysiology studies and during catheter radio-frequency ablation treatment for monitoring the ablation and its effects. The built-in ablation monitoring capability enables combined EP study and ablation treatment, thus helping to reduce exposure times and the total time needed per patient. For clinical use the software includes versatile tools for data analysis and reduction. Our system has been developed in association with Department of Cardiology of Tampere University Hospital and has been in regular clinical use there. PMID- 10710183 TI - Development and maintenance of guideline-based decision support for pharmacological treatment of hypertension. AB - The objective was to build a computer-based decision support system (DSS), which could apply the formal rules embedded in guidelines regarding pharmacological treatment of hypertension. The aim was also to test VISUAL BASIC as a development tool for DSS's in health care. From the Swedish guidelines for treatment of hypertension, the most widely accepted and scientifically best proved treatment strategies were chosen and implemented as rules. A DSS that is capable of applying the evidence-based rules extracted from guidelines regarding drug treatment of hypertension, to any patient's medical profile, was constructed. The output consists of a recommendation regarding preferred generic drug class and also a written report, reflecting decision steps provided by the rule-base and inference engine. We also provide methods for formalising an implementable language of guidelines. A mainstream programming language like VISUAL BASIC can be an alternative when building complicated decision support systems. A logic formal notation can facilitate communication between the expert and the programmer. The program is a stand-alone product independent of computerized medical records and thereby easy to install and maintain. PMID- 10710185 TI - Effects of aging and dietary restriction on the structural integrity of rat articular cartilage. AB - The objectives of this study were to investigate the effects of aging and diet restriction on the biomechanical properties of articular cartilage, using a well controlled rat model (Fischer 344). This animal model is recommended by the National Institute of Aging specifically to study aging and diet issues. The intrinsic biomechanical properties of articular cartilage were obtained using a creep indentation approach. The ages chosen (6, 12, 18, 24 months of age) correspond to approximate human ages of 20 to 80 years old. The diet regimen employed in this study used either an ad libitum fed group or a group fed 60% of the mean food intake of the ad libitum group. The results demonstrate that, unlike bone, rat articular cartilage biomechanical properties are not affected in a discernible manner by diet restriction, despite the fact that diet-restricted animals were significantly lighter in terms of body weight. Age effects on biomechanical properties are found only at 6 and 12 months probably due to developmental reasons, but not at later ages. It appears that aging and diet restriction have profoundly different effects on articular cartilage and bone. Another significant result of this study was to establish the rat as a suitable animal model to study cartilage biomechanical properties. Thus, the rat can be added to the list of animals that can be used to study structure-function and pathophysiological relationships in articular cartilage. PMID- 10710186 TI - Interstitial fluid pressurization during confined compression cyclical loading of articular cartilage. AB - The objective of this study was to experimentally verify the well-accepted but untested hypothesis that cartilage interstitial fluid pressurizes variously under the action of an applied cyclical stress in confined compression over a range of loading frequencies, contributing significantly to the cartilage dynamic stiffness. Eighteen bovine cartilage cylindrical samples were tested under load control using a porous indenter in a confined compression chamber fitted with a microchip pressure transducer at its bottom. Over a static stress of 130 kPa, a cyclical stress of amplitude 33 kPa was applied with the indenter at frequencies ranging from 0.0001 to 0.1 Hz. The cartilage interstitial fluid pressure and deformation were measured simultaneously as a function of time. The displacement response at the lowest tested frequency was curvefitted in the time domain to determine the linear biphasic material properties, H(A) = 0.70+/-0.10 MPa and k0=2.4x10(-16)+/-0.64x10(-16) m4/N s. These properties were employed in the biphasic theory to predict the interstitial fluid pressure response and compare it to experiment, resulting in nonlinear coefficients of determination ranging from r2 = 0.89+/-0.15 to 0.96+/-0.03 depending on frequency. It was found for the samples of this study that above a characteristic frequency of 0.00044 Hz, the magnitude and phase of fluid pressurization matched the applied stress, reducing the tissue strain at the impermeable bottom surface to nearly zero. The findings of this study verify the hypothesis that cartilage dynamic stiffness derives primarily from flow-dependent viscoelasticity as predicted by the linear biphasic theory; they demonstrate experimentally the significance of interstitial fluid pressurization as the fundamental mechanism of cartilage load support over a wide range of frequencies. PMID- 10710187 TI - Experimental evaluation of a model for oxygen exchange in a pulsating intravascular artificial lung. AB - Intravascular oxygenation and carbon dioxide removal remains a potentially attractive means for respiratory support in patients with acute or chronic respiratory failure. Our group has been developing an intravascular hollow fiber artificial lung that uses a pulsating balloon located within the fiber bundle to augment gas transfer. We previously reported on a simple compartmental model for simulating O2 exchange in pulsating intravascular artificial lungs. In this study we evaluate the O2 exchange model with gas exchange and PO2 measurements performed on an idealized intravascular artificial lung (IIVAL) tested in a water perfusion loop. The IIVAL has well-defined bundle geometry and can be operated in balloon pulsation mode, or a steady perfusion mode for determining the mass transfer correlation required by the model. The O2 exchange rates and compartmental O2 tensions measured with balloon pulsation in the IIVAL are within 10% of model predictions for flow and pulsation conditions relevant to intravascular oxygenation. The experiments confirmed that a significant buildup of PO2 occurs within the fiber bundle, which reduces the O2 exchange rate. The agreement between experiments and predictions suggests that the model captures the cardinal processes dictating gas transfer in pulsating intravascular artificial lungs. PMID- 10710188 TI - Reproducibility of parameters of postocclusive reactive hyperemia measured by near infrared spectroscopy and transcutaneous oximetry. AB - The purpose of this study was to investigate postocclusive hyperemic response using near infrared spectroscopy (NIRS) and transcutaneous oximetry (TcpO2). Five minute arterial occlusion on the calf muscle was performed in six healthy volunteers (mean age 29, range 23-34 years, mean TcpO2 at rest 53 mm Hg, range 47 58 mmHg, and ankle brachial index between 1 and 1.2). Oxygen partial pressure at rest, oxygen consumption (VO2) during ischemia, recovery times and resaturation rates after arterial occlusion were determined and new parameters for evaluation of the level of vascular disorders of lower limbs are suggested. The reproducibility of the signals was studied by repeating the same protocol on each subject four to six times. Repeated measurements showed no significant difference among trials, indicating that the measurements were reproducible. The mean values of the coefficient of variability for suggested parameters varied between 6% and 30% (mean value 17%). Interindividual variations of parameters are higher and can be explained by differences in fat/muscle ratio and in the measured tissue volume of the NIRS signal. Simultaneous measurements of NIRS and TcpO2 showed different responses to ischemic conditions, due to the different physiological levels of oxygen assessment. The combined use of both methods yields deeper insight into conditions of blood flow and tissue oxygenation. PMID- 10710189 TI - Influence of agonist, shear rate, and perfusion time on nitric oxide inhibition of platelet deposition. AB - Nitric oxide (NO) is a physiological species involved in inhibition of platelet adhesion and aggregation. A novel NO delivery device was utilized to quantitatively assess the effects of gaseous NO on platelet deposition to agonist coated biomaterials in the presence of a platelet suspension. Platelet deposition was evaluated as a function of agonist (collagen, fibrinogen, or IgG), shear rate (250, 500, and 750 s(-1)), and perfusion time (5, 7.5, and 15 min). The minimal aqueous surface NO concentrations and fluxes necessary for significant inhibition of platelet deposition were quantified. Platelet deposition was completely inhibited at a gaseous NO exposure of 0.1 ppm, irrespective of the platelet agonist, shear rate, and perfusion time. The corresponding aqueous surface NO concentration was 0.09 nM at 250 s(-1) as predicted by a validated model. Surface fluxes ranged between 0.3 and 0.6 femtomoles cm(-2) s(-1). The results of this study are useful for establishing generalized guidelines (i.e., NO flux requirements in the presence of agonists, shear rate, and perfusion time) for the design and development of suitable biomaterials incorporating NO to reduce platelet deposition. Further studies incorporating blood, rather than platelet suspensions, are required to provide a more complete assessment of the required NO flux necessary to inhibit platelet deposition. PMID- 10710190 TI - Enhancement of stent-induced thromboembolism by residual stenoses: contribution of hemodynamics. AB - In vitro stent-induced thromboembolism was altered by the presence of residual stenoses placed upstream or placed upstream and downstream of the stent. Heparinized (3 micro/ml) bovine blood was gravity fed through a conduit with a deployed coronary stent. Embolism was continuously monitored using a light scattering microemboli detector, and the thrombus accumulated on the stent at the conclusion of the experiment was assessed gravimetrically. Gaussian stenoses (75% reduction in the cross-sectional area) were placed upstream or upstream and downstream of the stent to alter flow characteristics in the stent region. The presence of stenoses enhanced embolization from the stent in all cases, while end point thrombus accumulation on the stent decreased with only an upstream stenosis present, and increased when upstream and downstream stenoses were present. Computational fluid dynamics with and without hypothetical model thrombi were used to ascertain the alterations in the flow environment caused by the stenoses and thrombi. Combining the computed hemodynamic parameters with experimental results indicated that increased radial transport of blood components and low wall shear stress provided by the stenoses and thrombi may explain the enhancement of end-point thrombus accumulation. Analysis further showed that thrombi growing at the stenosis-induced reattachment and separation points will be subjected to high shear forces which may explain the increased embolism when stenoses are present. PMID- 10710191 TI - Laser Doppler anemometry study of bidimensional flows downstream of three 19 mm bileaflet valves in the mitral position, under kinematic similarity. AB - Three small-size (nominal size: 19 mm) bileaflet valves, CarboMedics R (CM), St Jude Standard (SJ) and Sorin Bicarbon (SB), have been tested by means of a two component laser Doppler anemometry (LDA) system, in the mitral position, in order to assess the potential damage to blood elements entailed by the turbulent flow through them. A high regime (6 l/min cardiac output) was chosen to perform measurements for the worst case in generated turbulence. Two half-diameter profiles, at 13 and 26 mm downstream of the valve plane, have been investigated for each model. Besides velocity profiles, turbulence shear stresses (TSS) are reported, after the application of the stress analysis technique, in order to assess the maximum values of TSS (TSSmax) exerted on blood particles. Results show the typical bileaflet-type velocity profile for SB and SJ, with three jets exiting the valve, whereas CM lacks the central jet, due to instabilities of its flow field. As for TSSmax, CM reaches the highest values, presumably due to leaflets fluttering. SJ's TSSmax profiles maintain similar shapes at the two downstream locations, whereas SB presents an unexpected increase in the peak value of TSSmax, from 13 to 26 mm downstream of the valve plane, probably due to the curved leaflet design. The three prosthetic heart valves (PHVs) tested show many differences as for their turbulence properties, although they are similarly constructed. PMID- 10710192 TI - Prediction of solute kinetics, acid-base status, and blood volume changes during profiled hemodialysis. AB - A mathematical model of solute kinetics oriented to the simulation of hemodialysis is presented. It includes a three-compartment model of body fluids (plasma, interstitial and intracellular), a two-compartment description of the main solutes (K+, Na+, Cl-, urea, HCO3-, H+), and acid-base equilibrium through two buffer systems (bicarbonate and noncarbonic buffers). Tentative values for the main model parameters can be given a priori, on the basis of body weight and plasma concentration values measured before beginning the session. The model allows computation of the amount of sodium removed during hemodialysis, and may enable the prediction of plasma volume and osmolarity changes induced by a given sodium concentration profile in the dialysate and by a given ultrafiltration profile. Model predictions are compared with clinical data obtained during 11 different profiled hemodialysis sessions, both with all parameters assigned a priori, and after individual estimation of dialysances and mass-transfer coefficients. In most cases, the agreement between the time pattern of model solute concentrations in plasma and clinical data was satisfactory. In two sessions, blood volume changes were directly measured in the patient, and in both cases the agreement with model predictions was acceptable. The present model can be used to improve the dialysis session taking some characteristics of individual patients into account, in order to minimize intradialytic unbalances (such as hypotension or disequilibrium syndrome). PMID- 10710193 TI - Vegetable and fruit consumption and lung cancer risk in the Netherlands Cohort Study on diet and cancer. AB - OBJECTIVE: The purpose was to study the association between vegetable and fruit consumption and lung cancer incidence using 1074 cases after 6.3 years of follow up in the Netherlands Cohort Study. METHODS: Dietary intake was assessed using a 150-item food-frequency questionnaire. Multivariate models were used including age, sex, family history of lung cancer, highest educational level attained, and smoking history. RESULTS: Statistically significant inverse associations were found with total vegetables and most vegetable groups. Rate ratios (RRs) based on consumption frequency showed the strongest effect of vegetables from the Brassica group (RR 0.5, 95% confidence interval (95% CI) 0.3-0.9, for consumption > or = 3 times per week versus < or = once a month). RR of highest versus lowest quintile of total vegetable consumption was 0.7 (95% CI 0.5-1.0, p-trend 0.001). Statistically significant inverse associations were found for all fruits listed in the questionnaire. RRs for quintiles of total fruit intake were 1.0, 0.7, 0.6, 0.6 and 0.8 respectively (p-trend < 0.0001). Protective effects of fruits and vegetables were stronger in current than in former smokers, and weaker for adenocarcinomas than for other types of tumors. CONCLUSIONS: Inverse associations with lung cancer are found for both vegetable and fruit intake, but no specific type of vegetable or fruit seems to be particularly responsible. PMID- 10710194 TI - Hydatidiform moles and the long-term risk of breast cancer (Sweden). AB - OBJECTIVES: The etiology of breast cancer is only partially understood. Based on the findings that pregnancies reduce breast cancer risk, a possible inverse association between exposure to the placental hormone human chorionic gonadotropin (hCG) and the risk of breast cancer has been suggested. Hydatidiform mole, a gestational trophoblastic disease, is associated with a high expression of hCG. We performed a population-based cohort study in which women with a history of hydatidiform mole were followed up for future cancer outcomes. METHODS: All 3371 women with a notification of hydatidiform mole in the Swedish Cancer Registry between 1958 and 1993 were followed up for future cancer outcomes by record linkages within the registry. RESULTS: In a total of 57,075 person years of follow-up, 59 women had a diagnosis of breast cancer during follow-up, yielding an overall standardized incidence ratio of 1.3 (95% CI 1.0-1.7). CONCLUSION: This finding is not consistent with the hypothesis of a protective effect of hCG exposure on breast cancer risk, but rather suggests an adverse association. PMID- 10710195 TI - Hypothesis: iodine, selenium and the development of breast cancer. AB - BACKGROUND: In this paper we examine some of the evidence linking iodine and selenium to breast cancer development. Seaweed is a popular dietary component in Japan and a rich source of both of these essential elements. We hypothesize that this dietary preference may be associated with the low incidence of benign and malignant breast disease in Japanese women. In animal and human studies, iodine administration has been shown to cause regression of both iodine-deficient goiter and benign pathological breast tissue. Iodine, in addition to its incorporation into thyroid hormones, is organified into anti-proliferative iodolipids in the thyroid; such compounds may also play a role in the proliferative control of extrathyroidal tissues. Selenium acts synergistically with iodine. All three mono deiodinase enzymes are selenium-dependent and are involved in thyroid hormone regulation. In this way selenium status may affect both thyroid hormone homeostasis and iodine availability. CONCLUSION: Although there is suggestive evidence for a preventive role for iodine and selenium in breast cancer, rigorous retrospective and prospective studies are needed to confirm this hypothesis. PMID- 10710196 TI - Serum tocopherols, selenium and lung cancer risk among tin miners in China. AB - OBJECTIVE: To evaluate the association of prediagnostic serum antioxidants and lung cancer risk we conducted a case-control study nested in an occupational cohort of tin miners. METHODS: Male workers free of cancer enrolled in the cohort. During up to 6 years of follow-up, 339 lung cancer cases were diagnosed and, among these cases, those who donated blood prospectively (n = 108) were eligible for this study. For each case, two controls alive and free of cancer at the time of case diagnosis were matched on age and date of blood collection. RESULTS: Overall, we observed no association between serum alpha-tocopherol, gamma-tocopherol or selenium levels and lung cancer risk. However, a significant gradient of decreasing lung cancer risk with increasing serum alpha-tocopherol was apparent for men less than 60 years old (odds ratio by tertile: 1.0, 0.9, 0.2; trend p = 0.002). Alpha-tocopherol was also protective in men who reported no alcohol drinking (OR by tertile: 1.0, 0.6, 0.3; trend p = 0.008). CONCLUSION: Although there were no significant overall associations between prospectively collected serum alpha-tocopherol, gamma-tocopherol or selenium and incidence of lung cancer, results from this study suggest that higher alpha-tocopherol levels may be protective in men less than 60 years old and in those who do not drink alcohol. PMID- 10710198 TI - Crohn's disease and cancer risk (Denmark). AB - OBJECTIVES: The large number of studies of intestinal cancer among patients with Crohn's disease have provided inconsistent risk estimates in regard to risk of both colorectal and small intestinal cancer. We investigated incidence of cancer among Crohn's disease patients in comparison with the incidence in the general population of Denmark. METHODS: From the Danish National Registry of Patients we identified 2645 patients who had been hospitalized with Crohn's disease during 1977-1989. Cancer incidence for up to 17 years was determined in the cohort and compared to an expected number derived from national cancer incidence rates. RESULTS: The 15 observed cases of colorectal cancer were close to the expected number of 13.1 (SIR = 1.1; 95% CI 0.6-1.9), whereas the five cases of small intestinal cancer (three adenocarcinomas and two carcinoids) observed corresponded to an 18-fold increased risk (SIR = 17.9; 95% CI 5.8-42). CONCLUSIONS: A potential excess of colorectal cancer among subgroups of patients with Crohn's disease was not detectable in the overall risk estimate for colorectal cancer. Only for small intestinal cancer was a significantly elevated risk found among these patients hospitalized with Crohn's disease. PMID- 10710197 TI - A pooled analysis of thyroid cancer studies. V. Anthropometric factors. AB - OBJECTIVE: To assess the relation between anthropometric factors and thyroid cancer risk in a pooled analysis of individual data from 12 case-control studies conducted in the US, Japan, China and Europe. METHODS: 2056 female and 417 male cases, 3358 female and 965 male controls were considered. Odds ratios (OR) were derived from logistic regression, conditioning on age, A-bomb exposure (Japan) and study, and adjusting for radiotherapy. RESULTS: Compared to the lowest tertile of height, the pooled OR was 1.2 for females for the highest one, and 1.5 for males, and trends in risk were significant. With reference to weight at diagnosis, the OR for females was 1.2 for the highest tertile, and the trend in risk was significant, whereas no association was observed in males. Body mass index (BMI) at diagnosis was directly related to thyroid cancer risk in females (OR = 1.2 for the highest tertile), but not in males. No consistent pattern of risk emerged with BMI during the late teens. Most of the associations were observed both for papillary and follicular cancers, and in all age groups. However, significant heterogeneity was observed across studies. CONCLUSIONS: Height and weight at diagnosis are moderately related to thyroid cancer risk. PMID- 10710199 TI - Parental occupational exposure to magnetic fields and childhood cancer (Sweden). AB - OBJECTIVES: To test the hypothesis that parental occupational exposure to magnetic fields before conception and during pregnancy increases the risk of cancer in the offspring. METHODS: The study is designed as a cohort study based on a population of 235,635 children born shortly after two different censuses in Sweden. The children were followed from birth to 14 years and cases of cancer were identified in the Swedish cancer registry. The parents' occupational titles in the censuses were linked to a job-exposure matrix with information about magnetic field levels in different occupations. The cancer incidence among the exposed was compared to that among the unexposed using Cox proportional hazards modeling. RESULTS: There was no association between childhood cancer and maternal occupational magnetic field exposure. Paternal exposure was associated with an increased risk of childhood leukemia, with a relative risk of 2.0 (95% CI 1.1 3.5) for exposures > or = 0.30 microT. A decreased risk was found for brain tumors (RR = 0.5; 95% CI 0.3-1.0). CONCLUSIONS: The results do not support previous findings of an increased risk of childhood brain tumors associated with paternal occupational exposure to magnetic fields. The finding for childhood leukemia has to be interpreted with caution. PMID- 10710200 TI - Deprived areas and attendance to screening of cervix uteri cancer in a French region. AB - OBJECTIVES: To examine the relationship between deprivation and attendance to cervical cancer screening. METHODS: Three deprivation indices (Carstairs, UnderPrivileged Area, Department of Environment) were calculated for women aged 25-65 attending a 1993-95 cervical cancer screening program (Doubs "departement", France), with 594 municipalities as statistical units. Weighted multivariate linear regressions were performed, with attendance rate as the dependent variable, and the three deprivation indices in turn as independent variables along with women's mean age, average net income, density of (para)medical amenities, density of population and proportion of women. RESULTS: Per municipality women were numbered 1-29,822 (mean 210). In multivariate models, the three deprivation indices were negatively linked to attendance rate, and so were mean age of women and density of population. Average net income, proportion of women, and density of (para)medical amenities (nurses, laboratories, ambulances, physicians, dentists) were positively associated with attendance rate. CONCLUSIONS: In early stages, cervical cancer screening programs should account for populations living in deprived areas, through focused health promotion efforts and easier access to screening facilities. PMID- 10710201 TI - Descriptive epidemiology of thyroid cancer in Los Angeles County, 1972-1995. AB - OBJECTIVE: To examine the descriptive epidemiologic features of incident thyroid cancers diagnosed among Los Angeles County residents between 1972 and 1995. METHODS: The Los Angeles County/University of Southern California Cancer Surveillance Program (CSP) collected data on 8820 newly diagnosed thyroid cancer of cases occurring in Los Angeles County. Average annual age-adjusted incidence rates were calculated to study the epidemiology of each of the major histologic types of thyroid carcinoma. RESULTS: For all races combined, the age-adjusted incidence rates were 2.5 per 100,000 males and 6.0 per 100,000 females. Differences in incidence by ethnicity were particularly striking, with Filipinos demonstrating the highest incidence rates (4.44 per 100,000 males, 11.3 per 100,000 females). Over the 23-year period, thyroid cancer incidence rates increased 1.5% per year for males and 1% per year for females. By histology, papillary carcinoma rates increased over time and follicular carcinoma rates decreased. Non-Spanish surnamed white men employed in certain white-collar occupations and radiologic technicians were at greater risk of thyroid cancer. CONCLUSIONS: Additional research on the epidemiologic risk factors for thyroid cancer, particularly for gender and ethnicity, is needed to explain the marked elevated incidence rates among females and the Filipino population in Los Angeles County. PMID- 10710202 TI - Hepatitis B and C viruses in the etiology of hepatocellular carcinoma; a study in Greece using third-generation assays. AB - OBJECTIVES: The purpose of this study was to describe the role of hepatitis B virus (HBV) and hepatitis C virus (HCV) in the etiology of hepatocellular carcinoma (HCC). METHODS: During a 4-year period from January 1995 to December 1998, blood samples and questionnaire data were obtained from 333 incident cases of HCC from Athens, Greece, as well as from patients in two control groups, also from Athens. Controls were 272 metastatic liver cancer (MLC) patients and 360 patients hospitalized for injuries or eye, ear, nose or throat conditions. Coded sera were tested for hepatitis B surface antigen (HBsAg) and antibodies to hepatitis C virus (anti-HCV) by third-generation enzyme immunoassays. RESULTS: The odds ratios (with 95% confidence intervals) in logistic regression modeling comparing the HCC cases to the combined control series were 48.8 (30.5-78.3) for the presence of HBsAg and 23.2 (11.4-47.3) for the presence of anti-HCV. The odds ratio for concurrent infection with HBV and HCV was 46.2 (9.9-216.6) compared to infection with neither virus. CONCLUSIONS: Although HBV and HCV are both important causes of HCC in this study population the data do not suggest, neither do they conclusively refute, a super-additive interaction between the two infections in the development of this malignancy. In this population, 58% of HCC cases can be attributed to HBV, 12% to HCV, and 3% to dual infection with these viruses. PMID- 10710203 TI - Repeated measurements of organochlorine exposure and breast cancer risk (Denmark). AB - OBJECTIVE: To prospectively evaluate if repeated measurements of organochlorine exposure provide a more precise measure of breast cancer risk. METHODS: In the Copenhagen City Heart Study (CCHS) participants donated blood twice, in 1976-1978 and 1981 1983. Information on breast cancer risk factors was obtained through standardized questionnaires. A cohort nested case-control study of 155 cases and 274 matched breast cancer-free controls who had participated in both CCHS examinations was conducted. The average serum organochlorine concentration over the course of the two examinations was used, testing a possible association between organochlorine exposure and breast cancer risk. RESULTS: A high serum concentration of p,p'-DDT over the course of the two examinations was associated with a more than three-fold significantly increased risk of breast cancer, and a dose-response relationship was apparent. Furthermore, the risk of breast cancer increased with increasing serum concentrations of PCB congener 118 and 138 and the total amount of DDT isomers (sigmaDDT), but the trends were not significant. CONCLUSION: This study provides new evidence of the adverse effect of some organochlorines on breast cancer risk. Furthermore, repeated assessment of exposure during a relevant time period may provide a more precise risk estimate than a single measurement. PMID- 10710204 TI - Body size in different periods of life, diabetes mellitus, hypertension, and risk of postmenopausal endometrial cancer (Sweden). AB - OBJECTIVE: To measure the association between endometrial cancer risk and obesity at age 18 and recently, adult weight gain, diabetes mellitus and hypertension. METHODS: We performed a population-based, nationwide case-control study among postmenopausal women aged 50-74 years in Sweden, including 709 incident cases with histopathologically verified endometrial cancer and 3368 controls. RESULTS: Compared to lean women (recent body mass index (BMI), i.e. kg/m2 below 22.5), overweight women (recent BMI 28-29.99) had a 50% increase in risk for endometrial cancer (OR 1.5, 95% CI 1.0-2.1). Obese women (recent BMI 30-33.99) had a 3-fold increased risk (OR 2.9, 95% CI 2.0-4.0), and markedly obese women (recent BMI > or = 34) a 6-fold increased risk (OR 6.3, 95% CI 4.2-9.5). The OR for Type 2 diabetes mellitus was 1.5 (95% CI 1.0-2.1) and for Type 1 diabetes mellitus it was 13.3 (3.1-56.4). The effect of recent BMI was similar for tumors having different degrees of differentiation and myometrial invasion, and did not vary with age, time since menopause, smoking status, diabetes mellitus, and use of contraceptives. Hypertension increased risk only among obese women. BMI at age 18, height, and adult weight change were not independent risk factors. CONCLUSIONS: Recent overweight/obesity and diabetes mellitus (Types 1 and 2) are associated with endometrial cancer risk. Hypertension increases risk among obese women. PMID- 10710205 TI - A research agenda for tobacco control. PMID- 10710206 TI - A randomized trial of interferon alpha therapy for HIV type 1 infection. AB - The immunologic and virologic efficacy and safety of interferon a (IFN-alpha) administered in combination with zidovudine (ZDV) and zalcitabine (ddC) was evaluated in HIV-infected subjects with CD4+ cell counts between 300 and 500 cells/ml and no more than 14 weeks of prior antiretroviral therapy. A total of 256 subjects enrolled in an open-label, randomized controlled trial. Subjects were randomized equally into treatment groups. All subjects received ZDV and ddC, while half also receive IFN-alpha (3 MU subcutaneously every 24 hr). At 48 weeks the median average area under the curve minus baseline (AAUCMB) for plasma HIV-1 RNA for the two-drug group was -0.68 versus -0.75 log10 copies/ml for the IFN alpha group (p = 0.046). Mean HIV-1 RNA changes from baseline to 48 weeks for these groups were -0.65 and -1.12 log10 copies/ml, respectively (p = 0.010). The median AAUCMB for CD4+ cell count for the two-drug group was 28 versus -1 cells/mm3 for the IFN-alpha group (p = 0.011). Neutropenia, anemia, and drug intolerance were more common in the IFN-alpha group. This study demonstrates that IFN-alpha inhibits HIV-1 replication but attenuates the CD4+ cell response to dual therapy with ZDV and ddC. PMID- 10710207 TI - Normalization of CD4+ cell numbers and reduced levels of memory CD8+ cells in blood and tonsillar tissue after highly active antiretroviral therapy in early HIV type-1 infection. AB - Antiretroviral therapy increases the number of both CD4+ and CD8+ T cells in the blood of HIV-1-positive patients with advanced disease. In the present study, we have examined the kinetics of CD4+ and CD8+ T cell restoration in blood and lymphoid tissue in asymptomatic HIV-1-positive individuals with high CD4+ cell counts during highly active antiretroviral treatment. Tonsillar biopsies and blood samples were collected at baseline and at regular intervals during the following 48 weeks and from HIV-1-negative controls. Mononuclear cells from blood and tonsils were phenotyped and quantified by three-color flow cytometry. After 48 weeks of therapy, blood CD4+ cell counts in the HIV-1-infected group were comparable to those found in uninfected controls. Naive CD4+ T cells in blood increased during the initial 2 weeks in parallel with reduced plasma viremia. Both naive and memory CD4+ T cells in blood reached normal numbers by week 48, whereas the CD4+ naive/memory cell ratio in tonsils was within normal range throughout the study. The level of memory CD8+ T cells in blood declined during the first 8 weeks in parallel with a reduction in the tonsillar memory CD8+ T cells. Naive CD8+ T cells in the blood increased after 4 weeks, while the level of naive CD8+ T cells in tonsils remained unaltered. Our data indicate that in the early stages of HIV-1 infection antiretroviral therapy normalizes CD4+ cell counts and causes a decrease in the level of memory CD8+ cells in blood and lymphoid tissue, suggesting reduced CD8+ cell turnover in response to reduced viral replication. PMID- 10710208 TI - Massive loss of sulfur in HIV infection. AB - Skeletal muscle tissue from SIV-infected macaques was previously found to contain abnormally high sulfate and low glutathione levels indicative of an excessive cysteine catabolism. We now confirm the peripheral tissue as a site of massive cysteine catabolism in HIV infection and have determined the urinary loss of sulfur per time unit. The comparison of the sulfate concentrations of the arterial and venous blood from the lower extremities of 16 symptomatic HIV+ patients and 18 HIV- control subjects (study 1) revealed (1) that the peripheral tissue of HIV+ patients with or without highly active antiretroviral therapy (HAART) releases large amounts of sulfate and (2) that plasma sulfate, thioredoxin, and interleukin-6 levels are elevated in these patients. A complementary investigation of 64 asymptomatic HIV+ patients and 65 HIV- subjects (study 2) revealed increased plasma sulfate levels in the asymptomatic patients. The analysis of the daily urinary excretion of sulfate and urea of another group of 19 HIV+ patients and 22 healthy HIV- subjects (study 3) confirmed (1) that HIV+ patients experience a massive loss of sulfur and (2) that this loss is not ameliorated by HAART. The sulfur loss of asymptomatic patients was equivalent to a mean loss of about 10 g of cysteine per day. If extrapolated, this would correspond to an alarming negative balance of approximately 2 kg of cysteine per year under the assumption that the normal sulfate excretion equivalent to approximately 3 g of cysteine per day is balanced by a standard Western diet. The abnormally high sulfate/urea ratio suggests that this process drains largely the glutathione pool. PMID- 10710209 TI - HIV type 1-induced inhibition of CD45 tyrosine phosphatase activity correlates with disease progression and apoptosis, but not with anti-CD3-induced T cell proliferation. AB - The tyrosine phosphatase CD45 is a key positive element in multiple lymphocyte signaling pathways. To understand the contribution of CD45 to HIV-1-induced T cell hyporesponsiveness and apoptosis we evaluated the CD45-associated tyrosine phosphatase activity of lymphocytes from patients with different stages of HIV-1 disease and compared it with CD45 expression, spontaneous and Fas-induced apoptosis, anti-CD3-induced T cell proliferation, distribution of CCR5 delta32/wt, and cytokine production. The proliferative response to anti-CD3 as well as the CD45-associated phosphatase activity were significantly reduced in progressors. In long-term nonprogressors (LTNPs) the proliferative response to anti-CD3 was also diminished, although to a lesser extent, while the tyrosine phosphatase activity was not significantly impaired. One-third of LTNPs were found positive for the 32-bp deletion of the CCR5 gene. This mutation had no effects on anti-CD3 proliferative response or CD45 phosphatase activity. A significant reduction in IL-2 and IFN-gamma was observed in both LTNPs and in normal progressors, whereas IL-4 production was significantly decreased only in progressors. Last, we observed a significant correlation between CD45 phosphatase activity and apoptosis. We therefore conclude that the impairment of CD45 tyrosine phosphatase activity correlates with disease progression and the level of T cell apoptosis, but not with anti-CD3-induced T cell proliferation. Moreover, we suggest that evaluation of CD45 tyrosine phosphatase activity may represent an additional tool with which to assess disease progression. PMID- 10710210 TI - Increased levels of soluble Fas receptor and Fas ligand in the cerebrospinal fluid of HIV-infected patients. AB - Analyses of serum samples and blood cells have revealed a dysregulation of the Fas/Fas ligand (FasL) system during HIV infection, which may be related to disease progression. As Fas and FasL have been suggested to participate in brain injury in a variety of CNS disorders, the aim of this study was to determine (1) whether soluble Fas and FasL can be detected in cerebrospinal fluid (CSF) samples from HIV-infected patients, (2) whether levels of these molecules are related to disease progression, and (3) whether levels of sFasL are related to other laboratory findings. Soluble Fas was detected in 38 of 56 (68%) and soluble Fas ligand in 17 of 56 (30%) CSF samples from HIV-infected patients. CSF levels of both molecules correlated neither with the CSF-to-serum albumin ratio nor with corresponding serum concentrations. This finding suggests that they are at least in part produced intrathecally. Levels of both CSF sFas and sFasL correlated significantly and inversely with the blood CD4+ cell counts, suggesting that the intrathecal release of both molecules is increased during progression to advanced immunodeficiency. PMID- 10710211 TI - Production and characterization of simian--human immunodeficiency virus-like particles. AB - We have produced and characterized, in a baculovirus expression system, simian human immunodeficiency virus-like particles (SHIV VLPs) containing SIV Gag and HIV envelope (Env) proteins. Recombinant SIV gag (SIVmac239) and full-length or cytoplasmic domain-truncated HIV env from either HIV BH10 or HIV 89.6 virus were coexpressed in insect cells and Env incorporation into released SHIV VLPs was characterized. The expression level of the Env protein was found to be about 20 50% higher in both strains producing the truncated Env. Cell surface expression of the truncated Env proteins was found to be about eightfold higher than that of the full-length Env proteins. Furthermore, the truncated Env proteins exhibited higher levels of cleavage into gp120 and gp41 compared with the full-length Env. The SHIV VLPs produced by the coexpression of SIV gag and truncated HIV env contained both precursor (gp160) and gp120, while predominantly gp160 was found in the VLPs containing full-length Env. Coinfection of a recombinant virus expressing the protease furin also resulted in more efficient cleavage of gp160 to gp120. Both full-length and truncated Env were found to induce CD4+ cell fusion. Analysis of VLPs by immunoelectron microscopy demonstrated the incorporation of both full-length and truncated Env on the surface of VLPs. Truncated Env also was incorporated at higher levels on the surfaces of VLPs than full-length Env. The assembly of VLPs containing biologically active Env proteins may be useful in vaccine development and in functional studies of the HIV envelope protein. PMID- 10710212 TI - The HB-19 pseudopeptide 5[Kpsi(CH2N)PR]-TASP inhibits attachment of T lymophocyte and macrophage-tropic HIV to permissive cells. AB - The HB-19 pseudopeptide 5[Kpsi(CH2N)PR]-TASP[psi(CH2N) indicating a reduced peptide bond], which binds the cell surface-expressed nucleolin, is a potent inhibitor of HIV infection. Here, by using primary T lymphocyte cultures and an experimental cell model to monitor HIV entry, we show that HB-19 inhibits in a dose-dependent manner both T lymphocyte- and macrophage-tropic HIV isolates. Similar positively charged control pseudopeptides have no effect on HIV infection even at high concentrations. These observations, and the fact that HB-19 has no effect on SIV-mac and HIV-1 pseudotyped with VSV envelope glycoproteins, confirm the specific nature of this inhibitor against the entry process mediated by the HIV envelope glycoproteins. Finally, association of low doses of HB-19 with beta chemokines or AZT results in an increased inhibitory effect on HIV infection. HB 19 has no inhibitory effect when added to cells a few hours after HIV entry. On the other hand, in HB-19-pretreated cells, the inhibitory effect persists for several hours, even after washing cells to remove away the unbound pseudopeptide. Under such conditions, the attachment of HIV particles to cells is inhibited as efficiently as by neutralizing monoclonal antibodies directed against the V3 loop. In view of its specific mode of action on various HIV isolates, HB-19 represents a potential anti-HIV drug. PMID- 10710213 TI - Role of nitric oxide in the promoting effect of HIV type 1 infection and of gp120 envelope glycoprotein on interleukin 4-induced IgE production by normal human mononuclear cells. AB - Increased levels of serum IgE have been described in HIV-1 infection; however, mechanisms implicated in this immunoglobulin production remain unknown. In this study, we demonstrate that in vitro infection of human peripheral blood mononuclear cells (PBMCs) by HIV-1 monocytotropic (Ba-L) or lymphocytotropic (LAI) strains promotes IL-4-induced IgE production, indicating that the HIV-1 infectious process may participate in the IgE production observed in vivo. The effect of membrane glycoproteins (gp160, gp120, and gp41) was also evaluated. It was found that gp120 specifically potentiates in a dose-dependent manner IL-4 induced IgE production and does not affect IL-4-induced IgG, IgA, or IgM production. In these experiments, gp160 was also found to upregulate IL-4-induced IgE production, whereas gp41 was ineffective. This effect of gp120, gp160, and HIV-1 infection on IgE synthesis was not observed in the absence of IL-4. In the presence of IL-4, the inducing effect of gp120 appeared to be indirect because gp120 did not modify purified B lymphocyte IgE production after IL-4 and anti CD40 monoclonal antibody stimulation. As HIV-1 infection is associated with alterations of PBMC redox metabolism, the role of nitric oxide (NO) in this IgE production by human PBMCs was evaluated. In the presence of a specific inhibitor of NO synthase pathways (L-NAME), IgE production induced by IL-4 and gp120 was abolished. Taken together, these data indicate that HIV-1 envelope glycoprotein gp120 (and gp160) specifically enhances IL-4-induced IgE production by normal human PBMCs, probably through the regulation of the nitric oxide pathway. PMID- 10710214 TI - Parameters influencing measurement of the Gag antigen-specific T-proliferative response to HIV type 1 infection. AB - We have analyzed factors that might influence the in vitro quantitation of the T proliferative response to HIV-1 Gag antigens, a common and increasingly used clinical measurement of helper T cell function in the context of HIV-1 infection. We have compared the rate and extent of T cell proliferation in freshly prepared and previously frozen PBMC samples, and have concluded that frozen cells can be used successfully; we have assessed whether the suppression of any HIV-1 replication in the PBMC cultures affects the extent of T cell proliferation; we have studied which forms of the Gag antigens are the most efficient at inducing T cell proliferation. From the latter experiments, we conclude that Gag proteins that include p17, and perhaps also p7, sequences flanking the central p24 capsid protein, are better stimulants than proteins that comprise only p24 sequences. PMID- 10710215 TI - Priming of strong, broad, and long-lived HIV type 1 p55gag-specific CD8+ cytotoxic T cells after administration of a virus-like particle vaccine in rhesus macaques. AB - Despite advances in the clinical management of HIV infection, using combinations of antiretroviral pharmaceuticals, a safe and efficacious vaccine is needed to limit the AIDS pandemic. It is now thought that an effective HIV-1 vaccine should prime both cross-neutralizing antibodies and long-lasting cytotoxic CD8+ T lymphocytes (CTLs) recognizing multiple codominant HIV-1 epitopes. To that end, many novel vaccine strategies have been tested. However, only a few of these strategies, beside those relying on live-attenuated viruses, are able to prime strong CTL responses in nonhuman primates and humans. In this study, three rhesus macaques were immunized with HIV-1 p55gag virus-like particles (VLPs) in the absence of adjuvant to assess the potential of such a vaccine to prime CTL responses. After intramuscular injection of p55gag VLP, all three animals mounted CTL responses against HIV-1 p55gag. Notably, these CTLs primed by vaccination recognized naturally processed peptides and were long lived (>8.5 months) both in the peripheral blood and draining lymph node. Furthermore, these CTLs were directed against multiple HIV-1 p55gag epitopes. This indicated that immunization with p55gag VLP primes strong MHC class I-restricted, CD8+ cell-mediated immune responses and suggested that HIV-1 p55gag VLPs should be a reasonable vaccine candidate, when combined with strategies priming cross-neutralizing antibodies. PMID- 10710216 TI - Highly variable sequences in the env V3 region of HIV type 1 distributing among Thai carriers from 1995 to 1997. AB - The amino acid sequences of the Env V3 region of HIV-1 subtype E in Thailand were highly variable in the samples obtained from 1995 to 1997, compared with the previously reported sequences in samples obtained from 1990 to 1993. The sequences of the V3 region in the samples from five provinces in Thailand revealed that the variability was much higher in the samples from Bangkok and Ubonrachathani than in those from Chiangmai, Prathumthani, and Trang. There was no apparently different level of diversity at the V3 region in the samples from symptomatic and asymptomatic individuals. The V3 loop motif in most (66.7%) of the samples was GPGQ, although this motif was more heterogeneous in the samples from Bangkok and Ubonrachathani than in those from the other three provinces. The N-linked glycosylation sites in the V3 region among these samples were relatively conserved. There was no apparent difference in the presence of positively charged amino acids at positions 306 and 320 between the samples from symptomatic and asymptomatic individuals. PMID- 10710217 TI - Silent mutation in the V3 region characteristic of HIV type 1 env subtype B strains from injecting drug users in the former Soviet Union. AB - New independent states of the former Soviet Union are facing a rapidly growing epidemic of HIV-1 among injecting drug users (IDUs). This epidemic is caused by three HIV-1 populations, one belonging to HIV-1 subtype A (IDU-A), another to subtype B (IDU-B), and the third being a recombinant of the IDU-A and IDU-B viruses (IDU-A/B, gagA/envB). Each of these populations is characterized by a high level of genetic homogeneity. We identified a unique synonymous nucleotide substitution in the first isoleucine codon at the IHIGPGR motif (ATT), which was observed in the env subtype B V3 sequences derived from IDUs in Russia and the Ukraine. This substitution was observed in none of 179 sequences obtained from IDUs in western Europe, northern America, and Asia. Molecular epidemiological analysis of HIV-1 strains based on this sequence pattern could be useful for tracing the origin and spread of the IDU-B viruses to other countries and risk groups. PMID- 10710218 TI - Genetic analysis of HIV type 2 in monotypic and dual HIV infections. AB - A significant level of genetic variation among HIV-1 and HIV-2 has been described. The interaction of specific HIV-2 subtypes with HIV-1 may serve to identify potential biological properties associated with dual infection. To genetically characterize the HIV-2 strains circulating in Senegal and their relationship to coinfection with HIV-1, we sequenced the HIV-2 envelope C2-C3 region of 12 subjects coinfected with HIV-1 and HIV-2 and 9 subjects singly infected with HIV-2. The phylogenetic analysis showed that all subjects were infected with HIV-2 subtype A, confirming its predominance in West Africa. We did not observe specific sequences or genetic clustering based on coinfection status. PMID- 10710219 TI - Different distribution of HIV type 1 genetic variants in European patients with distinct risk practices. AB - The use of two genetic markers has permitted the analysis of the distribution of two different human immunodeficiency virus type 1 (HIV-1) variants in patients of the homosexual (HO) and intravenous drug user (IDU) groups in distinct European countries. In Germany, Holland, and Italy the variants circulating in each risk group of HO and IDU patients were genetically distinguishable according to the genetic markers used. In contrast, in France and Spain, the same variant has been recovered from patients with different risk practices. These data highlight the diversity of the HIV-1 epidemic in Europe and the different patterns of HIV-1 variant distribution in European countries. PMID- 10710220 TI - Occurrence of additional NF-kappaB-binding motifs in the long terminal repeat region of South African HIV type 1 subtype C isolates. PMID- 10710221 TI - Macrocephaly-Cutis marmorata telangiectatica congenita without cutis marmorata? AB - We report on two patients with clinical manifestations consistent with a diagnosis of macrocephaly-cutis marmorata telangiectatica congenita (M-CMTC). Both showed macrocephaly with high forehead, overgrowth, capillary hemangiomata involving philtrum, nose, and lips, and redundant skin. In addition, the first had cutis marmorata and joint laxity. The second had postaxial polydactyly of hands and feet, cutaneous syndactyly of third and fourth right fingers and of second and third right toes without evident cutis marmorata. A magnetic resonance imaging scan showed cerebral alterations in both patients. The first had bilateral cortical dysplasia with frontal bilateral myelinization defect of corona radiata. The second had mild intertonsillar widening, cavum septi pellucidi, small porencephalic areas in the anterolateral region of cellae, and subsequently developed a nonobstructive hydrocephalus. Reviewing all reported cases we propose a new criterion for M-CMTC diagnosis. PMID- 10710222 TI - De novo 46,XX,t(6;7)(q27;q11;23) associated with severe cardiovascular manifestations characteristic of supravalvular aortic stenosis and Williams syndrome. AB - Supravalvular aortic stenosis may present as an isolated finding or as part of Williams syndrome. Williams syndrome is a contiguous gene syndrome associated with neurodevelopmental and multisystemic manifestations caused by hemizygous deletion at 7q11.23. We report on the prenatal and histopathological findings in a patient with a chromosome translocation involving the Williams syndrome critical region. The initial abnormality on fetal ultrasound was hydrops fetalis detected at 30 weeks and echocardiography showed narrowing of the aorta and the pulmonary arteries. The baby died shortly after delivery and an autopsy revealed diffuse tubular thickening with luminal narrowing of the aorta, aortic branches, and the pulmonary arteries. Histopathology showed dysplasia of the media with reduced elastic content and "cartwheel" arrangement of collagen, elastic, and muscle fascicles. The karyotype was 46,XX,t(6;7)(q27;q11.23). Three signals were detected using the Oncor fluorescent in situ hybridization probe for elastin Williams syndrome (WSCR) suggesting that the break in chromosome 7 is within the elastin-Williams gene. This patient is of special interest because of the prenatal presentation and the chromosomal translocation involving the elastin Williams syndrome locus. PMID- 10710224 TI - Heterogeneity in Wiedemann-Beckwith syndrome: anthropometric evidence. AB - Wiedemann-Beckwith syndrome (WBS) has attracted a great deal of attention because of its genetic complexity. Individuals with WBS can be identified objectively by anthropometric analysis. Craniofacial anthropometry in conjunction with multivariate statistical analysis can be used to define patterns of variability that appear to relate to specific modes of inheritance that have been proposed for WBS. Our data on 19 affected individuals and their first-degree relatives indicate that the pattern of inheritance rather than the age of subjects may be responsible for the highly variable craniofacial phenotype found in individuals diagnosed with WBS. PMID- 10710223 TI - Preferential involvement of the short arm in chromosome 8-derived supernumerary markers and ring as identified by chromosome arm painting. AB - We report on the use of fluorescence in situ hybridization (FISH) with specific chromosome 8 arm painting to characterize further small supernumerary chromosome 8-derived markers/rings (SMC/SRC) identified in three patients. Two patients (patients 1 and 2) who carried the marker (SMC) were evaluated because of mental retardation and minor facial anomalies. The patient (patient 3) who carried the ring (SRC) had ventriculomegaly. Parental blood chromosomes of patients 2 and 3 were normal and unavailable on patient 1. The identification of the SMC/SRC was first characterized by FISH specific alpha-repeat centromeric probes, second by FISH whole chromosome painting (WCP), and finally by FISH chromosome arm painting (CAP). The latter showed involvement of only the short arm of chromosome 8 in all three SMC/SRC cases, suggesting a U-type exchange mechanism. PMID- 10710225 TI - Paternal isodisomy 13 in a normal newborn infant after trisomy rescue evidenced by prenatal diagnosis. AB - Maternal and paternal uniparental disomy of chromosome 13 have been associated with normal phenotypes. We report on a new case of paternal isodisomy 13 in a phenotypically normal girl. Prenatal diagnosis had shown a 46,XX,-13,der(13;13) karyotype in chorionic villi and a 45,XX,der(13;13) karyotype in amniocytes and fetal blood. Molecular studies demonstrated that the de novo der(13;13) was an isochromosome 13 of paternal origin. This observation supports the lack of imprinting effects on chromosome 13 and trisomy rescue as a mechanism leading to uniparental disomy in cases involving isochromosomes. PMID- 10710226 TI - Mental retardation and seizure disorder in Schimke immunoosseous dysplasia. AB - Schimke immunoosseous dysplasia (SID) is a rare, pleiotropic disorder compromising spondyloepiphyseal dysplasia, nephrotic syndrome, defective T-cell mediated immunity, and vascular changes which can lead to cerebral infarcts. The cause is unknown but an autosomal recessive inheritance pattern has been suggested. Understanding of the clinical phenotype is evolving; however, the neurologic spectrum is not well known. We report on a 17-year-old woman who presented with behavior changes, developmental regression, and partial complex seizures in early childhood. Computed tomographic scan of the brain was normal at that time. Short stature and cognitive deficits became evident several months later. At 4 1/2 years, she developed nephrotic syndrome and later malignant hypertension. Recent magnetic resonance imaging of the brain showed focal encephalomalacia in the parietal regions and a magnetic resonance angiography documented narrowing of the middle cerebral arteries. A skeletal survey showed evidence of spondyloepiphyseal dysplasia. We have not been able to identify an immune defect. To our knowledge this is the first reported patient with SID, profound mental retardation, and a seizure disorder. This case supports the theory that an intrinsic vascular defect may be more important in the pathogenesis of SID than a T-cell-mediated immune deficit. PMID- 10710227 TI - Family-based study of the association of the dopamine D2 receptor gene (DRD2) with habitual smoking. AB - A recent study showed an association between the dopamine D2 receptor gene (DRD2) and smoking. The purpose of this study was to determine if the familial transmission of smoking is linked to variation at the DRD2 locus in a genetically informative sample. Subjects were identified in alcohol treatment centers and their relatives were recruited for study. All subjects were interviewed to assess alcohol dependence, smoking habits, and psychiatric disorders. Two polymorphisms within the DRD2 gene were analyzed, including the TaqIA polymorphism. The sample consisted of 138 nuclear families with at least one offspring with habitual smoking, and analysis was by the transmission disequilibrium test (TDT), which avoids problems due to population stratification. There was no significant difference in the frequency between DRD2 alleles transmitted and not transmitted to habitual smokers. There also was no evidence for unequal transmission of DRD2 alleles for the phenotypes "ever smoker" or comorbid alcohol dependence and habitual smoking. This study does not support linkage of the DRD2 with smoking. PMID- 10710228 TI - Oculoauriculovertebral abnormalities in children of diabetic mothers. AB - Maternal diabetes is known to have teratogenic effects. Malformations including neural tube defects, caudal dysgenesis, vertebral defects, congenital heart defects, femoral hypoplasia, and renal anomalies are described in infants of diabetic mothers. However, craniofacial anomalies have rarely been reported in such infants. Here we document craniofacial anomalies of patients born to diabetic mothers. We describe two patient populations: individuals evaluated through our genetics services for multiple malformations and individuals identified through a database search in our craniofacial clinic. The first group consists of 14 individuals evaluated in our genetics clinics who were born to diabetic mothers and had craniofacial anomalies. The second group consists of seven individuals who were identified from a craniofacial database search of patients with hemifacial microsomia and who were born to diabetic mothers. Thus, both groups were born to diabetic mothers and had hemifacial microsomia (67%), microtia (52%), hearing loss (43%), epibulbar dermoids (24%), and fused cervical vertebrae (24%). Therefore, the teratogenic effects of maternal diabetes probably include such craniofacial malformations as the oculoauriculovertebral/Goldenhar complex. Infants of diabetic mothers should be evaluated for craniofacial anomalies. Conversely, mothers of infants with craniofacial anomalies should be evaluated for diabetes to aid in counseling concerning cause and recurrence risks. PMID- 10710229 TI - Jeune asphyxiating thoracic dystrophy and short-rib polydactyly type III (Verma Naumoff) are variants of the same disorder. AB - Jeune syndrome (JS) and short-rib polydactyly syndrome type III (SRP type III) are autosomal recessive disorders characterized by short ribs and polydactyly. They are distinguished from each other by the more severe radiological and histological bone findings as well as the occurrence of facial anomalies, ambiguous genitalia, and occasionally, cloacal abnormalities in SRP type III. We present a family in which two children have mild JS and one has SRP type III as evidence that JS and SRP type III are variants of the same disorder. The intrafamilial variability may reflect the effects of modifying loci on gene expression. PMID- 10710231 TI - Risk factors for cytogenetically normal holoprosencephaly in California: a population-based case-control study. AB - Holoprosencephaly is a developmental field defect manifested by a spectrum of abnormalities of the forebrain and midface. Approximately 50% of holoprosencephaly cases are associated with a cytogenetic abnormality or a monogenic syndrome. Suggested risk factors for the remaining 50% of cases have been described in case reports, but have not been confirmed in systematically conducted studies. We report the results of a population-based case-control study of holoprosencephaly. Live births, fetal deaths, and terminations with a diagnosis of cytogenetically normal holoprosencephaly were identified by the California Birth Defects Monitoring Program. Telephone interviews were conducted with the mothers of 58 cases and 107 live born, nonmalformed controls. Women were questioned about their health and reproductive histories, family demographics, and exposures occurring during their pregnancies. Among nonsyndromic cases, increased risks were observed for females (OR=1.8, 95% C.I. 0.9-3.9), foreign born vs. U.S. or Mexico-born women (OR=3.1, 95% C.I. 1.1-8.6), and women with early menarche (OR=2.3, 95% C.I. 0.9-5.7). Maternal periconceptional exposures associated with increased risks for nonsyndromic holoprosencephaly included alcohol consumption (OR=2.0, 95% C.I. 0.9-4.5), cigarette smoking (OR=4.1, 95% C.I. 1.4-12.0), and combined alcohol and smoking (OR=5.4, 95% C.I. 1.4-20.0), insulin-dependent diabetes (OR=10.2, 95% C.I. 1.9-39.4), medications for respiratory illnesses (OR=2.3, 95% C.I. 0.9-6.0), and salicylate-containing medications (OR=2.5, 95% C.I. 0.8-7.9). These findings are consistent with risk factors identified in some previous reports, and identify several new potential risk factors that require confirmation in future studies. PMID- 10710230 TI - Holoprosencephaly: molecular study of a California population. AB - Holoprosencephaly (HPE) is a common developmental anomaly of the forebrain and midface in which the cerebral hemispheres fail to separate into distinct left and right halves. HPE is extremely heterogeneous. In addition to teratogenic agents, several genes are implicated in the cause of HPE. Using samples from a population based birth defects registry in California, we performed a mutational analysis of the known HPE genes Sonic Hedgehog (SHH), ZIC2, and SIX3, in addition to two HPE candidate genes, TG-interacting factor (TGIF), and Patched (PTC), on a group of sporadic HPE patients. This is the first molecular study of HPE in a population based sample of patients. Among these patients, a deletion in the homeodomain of SIX3 and several polymorphisms in SIX3 and TGIF were identified. No sequence changes were detected in SHH, ZIC2, and PTC. Our results suggest that mutations in the currently recognized HPE genes may explain <5% of all sporadic HPE cases. PMID- 10710232 TI - Anodontia of permanent teeth (OMIM # 206780) and pegged/missing maxillary lateral incisors (OMIM # 150400) in the same family. PMID- 10710233 TI - CHILD syndrome caused by deficiency of 3beta-hydroxysteroid-delta8, delta7 isomerase. AB - CHILD (congenital hemidysplasia, ichthyosis, and limb defects) syndrome is a rare, usually sporadic disorder associated with unilateral distribution of ichthyosiform skin lesions, limb defects, punctate calcifications of cartilaginous structures, and visceral anomalies. CHILD syndrome shares some manifestations with X-linked dominant Conradi-Hunermann syndrome (CDPX2), although the skeletal defects and skin lesions in CDPX2 are bilateral and asymmetric. Because CDPX2 patients have abnormal 8-dehydrosterol metabolism caused by mutations in 3beta-hydroxysteroid-delta8,delta7-isomerase, we measured plasma sterols in a patient with CHILD syndrome and found levels of 8 dehydrocholesterol and 8(9)-cholestenol increased to the same degree as in CDPX2 patients. Subsequently, we identified a nonsense mutation in exon 3 of the patient's 3beta-hydroxysteroid-delta8,delta7-isomerase gene. We speculate that at least some cases of CHILD syndrome are allelic with CDPX2 caused by 3beta hydroxysteroid-delta8,delta7-isomerase deficiency. PMID- 10710234 TI - Behold the CHILD. PMID- 10710235 TI - Mutations in the NSDHL gene, encoding a 3beta-hydroxysteroid dehydrogenase, cause CHILD syndrome. AB - We report for the first time that CHILD syndrome (MIM 308050), an X-linked dominant, male-lethal trait characterized by an inflammatory nevus with striking lateralization and strict midline demarcation, as well as ipsilateral hypoplasia of the body is caused by mutations in the gene NSDHL located at Xq28 (NAD(P)H steroid dehydrogenase-like protein) encoding a 3beta-hydroxysteroid dehydrogenase functioning in the cholesterol biosynthetic pathway. SSCA and genomic sequence analysis of NSDHL identified in 6 patients with CHILD syndrome, including one boy as well as a mother and her daughter, mutations potentially impairing protein function. This phenotype is distinct from, but shares various clinical and biochemical findings with chondrodysplasia punctata (CDPX2, MIM 302960). CDPX2 is due to mutations affecting a delta8-delta7 sterol isomerase (EBP, emopamil binding protein, at Xp11.22-p11.23) that functions downstream of NSDHL in a later step of cholesterol biosynthesis. EBP was unaffected in the patients analyzed by us demonstrating that CHILD syndrome and CDPX2 are not caused by allelic mutations. Two mouse X-linked dominant male-lethal traits, bare patches (Bpa) and striated (Str) had previously been associated with mutations in Nsdhl. They provide animal models for the study of CHILD syndrome, a further human condition due to mutations in a gene of the cholesterol synthesis pathway. PMID- 10710236 TI - Detection of a common mutation in the RSH or Smith-Lemli-Opitz syndrome by a PCR RFLP assay: IVS8-G-->C is found in over sixty percent of US propositi. AB - The RSH or Smith-Lemli-Opitz syndrome (SLOS) is a relatively common autosomal recessive disorder of cholesterol biosynthesis resulting from a deficiency of the enzyme 7-dehydrocholesterol delta7-reductase (7-DHCR). Mutations in 7-DHCR gene cause SLOS. Among these, a G-->C transversion in the splice acceptor site of exon 9 (IVS8-1G-->C) was suspected to be a frequent mutation, having been detected in about 18% of SLOS patients so far. This mutation results in the elimination of a AlwN1 restriction endonuclease site. We report a simple PCR-RFLP assay to detect the IVS8-1G-->C mutation. Using this method, we identified the IVS8-1G-->C mutation in 21 of 33 SLOS propositi. This mutation was detected in one of 90 normal adult Caucasian Americans; but not among 121 Africans from Sierra Leone, 120 Caucasians from Finland, 95 Chinese or 103 Japanese adults. The results of this study provide further evidence that IVS8-1G-->C transversion is a very common mutation in SLOS patients from the US and that the carrier rate in US caucasians may be high. The simple PCR-RFLP assay developed makes identification of this mutation convenient for diagnosis and for carrier detection. PMID- 10710237 TI - Binocular interactions in visual evoked cortical potentials with two light emitting-diodes. AB - Binocular interaction for a central field was studied with transient scalp visual evoked cortical potentials (VECPs) using two light-emitting-diodes. VECPs were obtained for binocular and monocular visions with dominant and non-dominant eyes, and arithmetical sums of monocular VECPs with dominant and non-dominant eyes were calculated. Amplitude and latency of remarkable initial three peaks were tested with the multivariate analysis of variance. Significant differences were noted among the four VECPs. Pairwise comparisons showed that (1) the amplitude of the first peak for the binocular VECPs was larger than that for the monocular VECPs but smaller than that for the sum-VECPs; the latency of the first peaks for the binocular VECPs were earlier than that for the monocular VECPs with the non dominant eye; (2) the amplitude of the first negative peak for the sum-VECPs was larger than that for the binocular VECPs, and the peak latency for the sum-VECPs showed later than that for the binocular VECPs; (3) the amplitude of the second positive peak for the binocular VECPs and monocular VECPs with the dominant eye was larger than that with the non-dominant eye, but smaller for the binocular VECPS than that for the sum-VECPs; the latency for the binocular VECPs showed earlier than that for the monocular VECPs with the dominant eye and for the sum VECPs. Binocular suppression was noted in amplitude for the three peaks and binocular facilitation was noted in latency for the latter two peaks. PMID- 10710238 TI - Cone positive off-response in normal and dystrophic cats. AB - Light-adapted cone ERG in response to white stimuli with long duration (200 ms) was studied in seven normal cats and six cats with early to moderate, inherited retinal dystrophy. The stimuli typically elicited an ERG consisting of an a- and b-wave in response to light onset, whereas light-offset was followed by a cornea positive d-wave and subsequent negative dip and occasionally a second positive peak in both normal and dystrophic cats, although b- and d- waves had less distinct peaks in cats with moderately advanced retinal dystrophy. Linear regression models indicated a positive correlation between d-wave amplitude and stimulus luminance, whereas a negative correlation was found between amplitude and background light luminance in normal cats. The d-wave implicit time was independent of both stimulus and background light luminance in normal cats. The d wave amplitude was not significantly different in dystrophic cats, whereas the implicit time was increased when affected cats were compared to normal cats. The significant increase in implicit time in dystrophic cats could not be explained with a reduced sensitivity of the off-pathway to background or stimulus light. This is supported by the finding that d-wave amplitude was not significantly altered in early to moderately advanced dystrophic cats. PMID- 10710239 TI - Cone properties of the light-adapted murine ERG. AB - PURPOSE: Because the mouse lacks a typical Purkinje shift, we have examined its light-adapted ERG to determine whether there was other evidence in addition to tolerance to background light, that could be used to identify cone function in the ERG. METHODS: Full field corneal ERGs to white flashes, double flashes and flash trains were examined in the presence of a strong full field light adaptation and compared with the human cone ERG. RESULTS: The following cone-like properties could be identified. (1) The light-adapted murine ERG increases in amplitude gradually during the first 10 minutes of light-adaptation; (2) It is capable of responding to a 50 Hz stimulus, although its overall frequency response is slower than that of the human cone ERG; (3) A corneal positive d-wave occurs to the termination of a flash train; (4) The response increases linearly with light intensity. CONCLUSION: The light-adapted murine ERG has several properties of cones but it has a slower response than the human cone ERG. PMID- 10710240 TI - Does hyperbaric oxygen (HBO) delivery rescue retinal photoreceptors in retinitis pigmentosa? AB - As previously reported in the literature, hyperbaric oxygen delivery seems to modify the natural course of retinitis pigmentosa. In order to evaluate these first encouraging data, 48 affected subjects were separately studied in two subgroups (cases and controls). All patients underwent yearly an ophthalmological examination completed by a maximum amplitude electroretinogram, conducted according to our 'differential derivation' system, a new recording technique specifically designed to enhance the signal-to-noise ratio. Oxygen delivery was provided regularly for 90 min daily (2.2 Absolute Atmosphere) in three cycles according to a standard protocol. In the cases, electroretinographic mean values were as follows: at T0 (basal) 4.68 +/- 3.81 microV; after one year (T1) 8.46 +/- 5.71 microV; at two years (T2) 10.7 +/- 7.6 microV; at the end of the study (T3) 14.4 +/- 11.7 microV. In the controls, electroretinographic mean values were as follows: at T0 4.92 +/- 3.05 microV; at T1 5.04 +/- 3.07 microV; at T2 3.46 +/- 2.77 microV: at T3 2.97 +/- 3.61 microV. Amplitudes showed a remarkable (p<0.001) increase in the cases, while a slightly significant (p<0.02) decrease was evident at the end of the study in the controls. In our opinion, retinal oxygen availability may be critical in retinal degeneration and hyperbaric oxygen delivery, inducing hyperoxia, seems to be able to bring about the rescue of the retinal photoreceptors helping them in their metabolic requirements. Unfortunately, our study demonstrates an increase in electroretinographic responses only, which may not necessarily also mean an evident change in visual acuity. PMID- 10710241 TI - Light potentiation of horizontal cells in the isolated rabbit retina. AB - When the retinas of some fishes and amphibians are dark-adapted the hyperpolarising response of horizontal cells (HCs) to a light stimulus is suppressed (= dark suppression) but is restored to that of light adaptation when a light stimulus is rapidly repeated (= light potentiation (LP). This phenomenon, which had not been previously demonstrated in a mammal, has been recorded from isolated rabbit retinas. The steady amount of LP of HCs after several light stimuli was expressed as the percentage of induced hyperpolarisation when that of the dark-adapted retina is taken as 100%. The LP was small (113+/-13%, n=14), when the HCs had stable and large responses (>15 mV), but varied greatly between HCs. Those HCs with slight LP (106+/-3% n=8) designated as x-HCs, mostly exhibited larger overshoots than did those HCs with stronger LP (123+/-14, n=6) and designated y-HCs. These mostly had a smaller overshoot. Other HCs (z-HCs) were unstable and were only slightly hyperpolarised by the first light stimulus after dark adaptation but showed the strongest LP (193+/-48%, n=7). These results could indicate a variable degree of light potentiation in normally-functioning HCs, which can be classified accordingly. It is possible, however, that the degree of LP is small in all normally-functioning HCs, but as the HCs in isolated mammalian retinal preparations deteriorate, the phenomenon of LP is progressively exaggerated. LP is not peculiar to HCs and can also occur in cells which depolarise in response to a light stimulus, and can be evident in the PIII component of the ERG. PMID- 10710242 TI - Do horizontal cells contribute to the rabbit ERG? AB - An increase in the amplitude of horizontal cell (HC) potentials, a decrease in the extracellular potassium around the photoreceptors and an increase in the PIII component of the ERG that occur with increasing light stimulus intensity are shown to be concomitant occurrences, which could, therefore, be causally related. With high intensity light stimuli the PIII component is prominent and the HC potentials exhibit an afterpotential. Within the range of intensities tested the peak times of the HC afterpotentials coincide with the peak times of PIII. This indicates the likelihood that the HC potential contributes to the PIII, although these results do not allow a quantitative assessment of the contribution of HC potentials to the PIII-component. PMID- 10710243 TI - Similarities and dissimilarities between pattern VEPs and motion VEPs. AB - The contrast response functions (CRF) of pattern-appearance and motion-onset VEPs for periodic stimuli (gratings) were compared. The CRF for pattern-appearance is accelerative for the P100 component and compressive for the N200 component. Contrary to these results, the CRF for motion-onset shows an almost negligible slope for both components within the contrast range tested (0.5-64%). To better isolate the neural contributions to these different VEP components, we studied the effects of prior adaptation to stationary and moving gratings. Adaptation to stationary gratings has no effect on both VEP components for motion-onset and the P100 component for pattern-appearance, but did reduce the amplitude of the N200 for pattern-appearance. Adaptation to slow (1 deg/s) and fast (4 deg/s) gratings left the P100 amplitudes unaltered, while it significantly reduced the N200 amplitudes for both pattern-appearance and motion-onset. These results suggest that the N200 component of the motion-onset VEP is generated by motion-dependent neurons, whereas the same component for pattern-appearance arises from contrast dependent neurons. The observed differences between P100 and N200 components appear to reflect the activity of both transient and sustained neural mechanisms. PMID- 10710245 TI - Calcium supplement necessary to correct hypocalcemia after total parathyroidectomy for renal osteodystrophy. AB - BACKGROUND: Prediction of the extent of calcium supplement will facilitate safe and efficient management of hypocalcemia in the early postoperative stage of total parathyroidectomy with autotransplantation (PTXa) in patients with renal osteodystrophy. METHODS: The correlation between the extent of calcium deficiency, estimated by the amount of calcium supplement over 48 h after PTXa and using various parameters such as carboxy terminal parathyroid hormone (c PTH), intact PTH (i-PTH), alkaline phosphatase (ALP), serum calcium, serum phosphorus, duration of hemodialysis, total weight of resected parathyroid glands and degree of subperiosteal resorption of the middle phalanx was examined in 49 patients who underwent PTX with subcutaneous autotransplantation. Bone mineral density (BMD) was also determined before, 3 months and 1 year after PTXa with dual energy X-ray absorptiometry (DEXA) in 13 patients. RESULTS: There was a positive correlation between pre-operative i-PTH level (r=0.56, P<0.0005) or ALP level (r=0.50, P<0.0005) and the amount of calcium supplement over 48 h after PTXa in these patients. Furthermore, the degree of subperiosteal resorption, determined by Jensen's classification, was significantly correlated with the amount of calcium supplement after PTX (P<0.05). Bone mineral density 3 months after (P<0.0005) and 1 year after PTXa (P<0.001) significantly increased compared with BMD before PTXa in all patients examined. CONCLUSION: These findings suggest that the pre-operative determination of i-PTH, ALP levels and degree of subperiosteal resorption allow the management of hypocalcemia safely and efficiently in renal osteodystrophy patients after PTXa. PMID- 10710244 TI - Predictors of visual acuity and the relative afferent pupillary defect in optic neuropathy. AB - The relationships among visual acuity (log MAR), diagnostic category, age, the magnitude of a relative afferent pupillary defect (RAPD) in log units, photopic foveal thresholds to white and colored light (dB), and the mean deviation on the Humphrey visual field (dB) were studied in patients with various optic neuropathies. All acuity and dB values were expressed as interocular differences, the majority of cases having normal acuity in the fellow eye. In multiple regression analyses, acuity and RAPD were alternately chosen as the dependent or response variable with all remaining variables serving as the predictors or independent variables. The main finding was that the only significant predictor of a RAPD was the interocular mean deviation difference on the Humphrey field and the only significant predictor of acuity was the foveal threshold to white light. Redundant and insignificant variables were therefore identified with multiple regression analysis. Subsidiary findings include: (a) although diagnostic group was not a significant predictor in the above, simple linear regression line slopes relating RAPD magnitude to the Humphrey mean deviation were significantly different between optic neuritis and compression categories; (b) for a given level of acuity, foveal thresholds were substantially worse in these cases with neuronal damage than in strabismic amblyopia, refractive error, or corneal damage; and (c) sensitivity losses for red vs. blue light were similar. PMID- 10710246 TI - A bladder preservation regimen using intra-arterial chemotherapy and radiotherapy for invasive bladder cancer: a prospective study. AB - BACKGROUND: A prospective study was performed to investigate combined treatment with intra-arterial chemotherapy and radiation therapy for bladder preservation in locally invasive bladder cancer. METHODS: Patients with invasive bladder cancer, stage T2-3N0M0, were included in the study. Intra-arterial chemotherapy was performed with three injections of methotrexate and cisplatin at 3-week intervals. Simultaneously, the patients underwent X-ray irradiation (40 Gy) of the small pelvic space. Where a post-treatment transurethral resection (TUR) biopsy showed no residual tumor, the tumor site was irradiated by a 30 Gy proton beam and the bladder was preserved. Where tumors remained, radical cystectomy was performed. RESULTS: Between 1990 and 1996, 42 patients were treated according to this protocol. Post-treatment TUR biopsy and urine cytology showed no residual tumors in 39 of 42 cases (93%). The bladder was preserved in accordance with the study protocol in 36 cases. A median follow-up of 38 months showed 3-year non recurrence in 72% of bladder-preserved patients and the rate of bladder preservation was 84%. The nine recurrences included eight cases of superficial bladder recurrence. One cancer death occurred among the bladder-preservation patients, giving 3-year survival and cause-specific survival rates of 84% and 100%, respectively. Although bladder function decreased slightly in compliance, bladder capacity was retained in almost all cases. CONCLUSIONS: This regimen is useful for bladder preservation in T2-3 locally invasive bladder cancer. Information from more cases and the results of more long-term observations are needed, as is an evaluation of appropriate subject selection and factors associated with quality of life issues, particularly regarding bladder function. PMID- 10710247 TI - Effect of prostatic biopsy on free-to-total prostate-specific antigen ratio in patients with prostate cancer. AB - BACKGROUND: We determined the effect of prostatic biopsy on the changes in total and free prostate-specific antigen (PSA) and free-to-total PSA ratio (F/T ratio) and examined if there are differences in these parameters between patients with benign and malignant histologic findings. METHODS: The concentration of total and free PSA and the F/T ratio were determined in 35 men before and 1 h after prostatic biopsy. The level of PSA was measured with a chemiluminescent enzyme assay. Of 35 patients, nine were diagnosed as having prostate cancer. RESULTS: In patients whose biopsy revealed cancer, the F/T ratio was lower than those without cancer, although there were no differences in total and free PSA value before prostatic biopsy. One hour after prostatic biopsy, there was an increase in the level of total and free PSA and the F/T ratio in all men. The increase in the F/T ratio was greater in patients whose biopsies revealed no prostate cancer. In patients with stage B cancer, these parameters increased more than those with stage C/D cancer. CONCLUSION: Prostatic biopsy causes a dramatic increase in total and free PSA. The F/T ratio also increased after biopsy. The PSA response to prostatic biopsy might be different in patients with and without prostatic malignancy. The response might also be different according to stage of prostate cancer. PMID- 10710248 TI - A case of eosinophilic cystitis in a 5-year-old boy. AB - Eosinophilic cystitis (EC) is rather an uncommon disease in childhood. A case of EC in a 5-year-old boy, in which open biopsy was needed for final diagnosis, is reported. After diagnosis, he was treated with pemirolast potassium followed-up with eosinophil cationic protein (ECP) in serum and urine. Eosinophil cationic protein is an appropriate marker of EC. PMID- 10710249 TI - Sarcomatoid renal cell carcinoma with scant carcinomatous components. AB - A 30-year-old male underwent radical nephrectomy for a right renal tumor 15 cm in diameter. On microscopic examination of initial 17 sections, the tumor consisted of pleomorphic giant cells and spindle neoplastic cells. There was no carcinomatous component. Immunohistochemically, the neoplastic cells were negative for keratin and epithelial membrane antigen but positive for vimentin. The giant cells were also scatteringly, weakly positive for myoglobin. At that time a diagnosis of rhabdomyosarcoma of the kidney was made. However, further microscopic examination of another eight sections revealed small areas of clear cell-type renal cell carcinoma (RCC) which transited to sarcomatous components and led to a diagnosis of sarcomatoid RCC. The patient underwent three cycles of adjuvant chemotherapy. He has been free of the disease for 14 months after nephrectomy. PMID- 10710250 TI - A case of neuroleptic malignant syndrome in a patient with hemodialysis. AB - A 63-year-old man with chronic renal failure who had received hemodialysis three times per week for 4 years developed neuroleptic malignant syndrome 10 days after taking amoxapine. His condition was characterized by muscle rigidity, elevation of body temperature and altered consciousness. Although he was treated with dantrolen and supportive care as well as discontinuation of amoxapine, his condition rapidly deteriorated, resulting in death. Because the pharmacokinetics of drugs, especially those such as antidepressants, in patients with chronic renal failure has not been fully clarified, one should be careful about giving such patients these drugs. PMID- 10710251 TI - Acute pancreatitis caused by extracorporeal shock wave lithotripsy for bilateral renal pelvic calculi. AB - An elderly woman with a history of cholecystectomy and a re-operation for postoperative peritonitis underwent extracorporeal shock wave lithotripsy (ESWL) for right and left renal pelvic calculi, 11 x 6 and 12 x 5 mm in size, to which 2400 and 1400 shots at 20 kV were given, respectively, on the same day. During the evening after the operation, the patient started to complain of upper abdominal pain. Laboratory examination on the next day revealed elevations in blood and urine amylase levels and a diagnosis of pancreatitis was made. Conservative treatment, including administration of protease inhibitor, did not improve her symptoms; abdominal distension became marked and she underwent laparotomy. Necrosection and indwelling of several drain tubes in abdomen were performed with an operative diagnosis of acute necrotic pancreatitis. With daily irrigation of drain tubes and treatment for methicillin-resistant Staphyloococcus aureus infection of the lungs and abdominal cavity, septicemia and duodenal fistula, the patient gradually recovered and was discharged on postoperative day 151. It was suggested that ESWL was responsible for the acute pancreatitis. Either an obstruction of the pancreatic duct by fragments of common duct stone, or mechanical injury of the pancreas due to adhesion between the pancreas and surrounding tissue caused by the lapalotomy, was considered as a possible cause of pancreatitis. To our knowledge, there has been no previous report of severe acute pancreatitis and the present case suggests that ESWL may cause severe pancreatic even in cases without stone shadow in the bile, common duct or pancreatic duct. PMID- 10710252 TI - Acute renal failure occurring from urinary retention due to a mullerian duct cyst. AB - A patient with a mullerian duct cyst, which caused acute renal failure secondary to urinary retention, is reported. The case was treated successfully by transurethral unroofing of the cyst. PMID- 10710254 TI - The blue train PMID- 10710255 TI - Reconstruction of Achilles tendon defect with a free quadriceps bone-tendon graft without anastamosis. AB - We report a method of Achilles tendon reconstruction using a free quadriceps bone tendon graft. The patient had a prior repair of a re-ruptured Achilles tendon, following which he developed massive necrosis of his skin and Achilles tendon leaving a 10 cm defect. First stage reconstruction consisted of soft tissue coverage of the skin defect with a sural fasciocutaneous flap. Reconstruction of the Achilles tendon followed, with the patellar bone block fixed to the calcaneus and the quadriceps tendon sutured proximally. PMID- 10710253 TI - Simultaneous production of granulocyte colony-stimulating factor and parathyroid hormone-related protein in bladder cancer. AB - A case of bladder cancer with simultaneous production of granulocyte colony stimulating factor (G-CSF) and parathyroid hormone-related protein (PTHrP) is reported. An 81-year-old male patient was admitted to the Saitama Medical School for treatment of gross hematuria, leukocytosis and hypercalcemia and diagnosed as having advanced bladder cancer. Immediately after a cystectomy was carried out, his white cell count and serum calcium levels returned to normal. However, the tumors recurred locally and the recurrence was accompanied by an increase in the serum G-CSF and PTHrP levels with a recurrent elevation of white cell count and the serum calcium level. The production of G-CSF and PTHrP in the tumor cells was confirmed by immunohistochemistry. PMID- 10710256 TI - Achilles tendon rupture repair: biomechanical comparison of the triple bundle technique versus the Krakow locking loop technique. AB - This study was designed to compare the tensile strength of ruptured Achilles tendons repaired using either the triple bundle technique or the Krakow locking loop technique. Eight pairs of fresh frozen cadaveric Achilles tendons were harvested. A simulated "Achilles tendon rupture" was created 4 cm from the calcaneal insertion in all sixteen tendons by transversely cutting the tendon with a scalpel. One Achilles tendon "rupture" of a pair was repaired using the triple bundle technique, while the other tendon of the pair was repaired using the Krakow locking loop technique. Then, using a servohydraulic testing machine, each tendon was tested to failure in tension at a displacement rate of 2.54 cm/sec. The average rupture load for the triple bundle technique was 453 N (range 397 n 549 N), while the average rupture load for the Krakow locking loop technique was 161 N (range 121 n 216 N). This difference in averages represents a statistically significant superiority of 2.8 to 1 (p < 0.001) in favor of the triple bundle technique. PMID- 10710257 TI - The biomechanical relationship between the tendoachilles, plantar fascia and metatarsophalangeal joint dorsiflexion angle. AB - We carried out an experiment to measure the relationship between tensile force in the tendoachilles and plantar fascia strain, and how this relationship is affected by the metatarsophalangeal joint dorsiflexion angle. Eight cadaver lower extremity specimens underwent biomechanical testing. Using a servo-hydraulic testing machine, a tensile force up to 500 N was applied to the tendoachilles while the strain on the plantar fascia was measured using an extensometer. The experiment was repeated at four different metatarsophalangeal joint dorsiflexion angles (0 degrees, 5 degrees, 30 degrees, and 45 degrees). Measurements and calculations showed that dorsiflexion of the toes tightens the plantar fascia (the windlass effect) and increases the effect that a tensile force in the tendoachilles has on the tensile strain and tensile force in the plantar fascia. PMID- 10710258 TI - Haglund's syndrome: disappointing results following surgery -- a clinical and radiographic analysis. AB - Clinical and radiographic data were analyzed for 49 surgical interventions of painful heel syndrome with a mean follow-up of 4 years and 7 months. In all patients the surgical procedure consisted of a triangular shaped resection of the posterosuperior portion of the calcaneus. The clinical evaluation showed disappointing results with complete relief of complaints in only 34 procedures (69.4%), and 7 patients (14.3%) with even a worsening of their symptoms. The time course of rehabilitation was long with an average of 6 months of functionally significant pain after surgery, reducing the willingness to undergo this operation again. Formerly published angular thresholds of radiographic calcaneal angles could not be confirmed to be predictors for the preoperative symptoms or the postoperative outcome. There was no accumulation of pathologic clinical foot shapes, only a slight increased calcaneal pitch in patients with isolated pain on the posterosuperior lateral part of the calcaneus. According to Haglund's description of painful heel syndrome, the clinical picture of our patients included affections of bone, tendon, peritendon, bursae, soft tissue and skin in different combinations. Based on this evaluation, we advise to be cautious to indicate the operation. All possibilities of conservative treatment should be performed prior to surgery. PMID- 10710259 TI - Psychogenic equinovarus: the importance of recognition and non-operative treatment. AB - Although the potential for musculoskeletal symptoms in hysteric conversion disorder was recognized by Sigmund Freud, reports of it in the orthopaedic literature have been limited to upper extremity manifestations. This study reports 3 cases which illustrate hysteric conversion presenting as primary foot and ankle complaints. Given its relative rarity, it is a diagnosis that is easy to miss. Clinical clues to its diagnosis and accepted methods of treatment are discussed. It is important to realize that this condition arises from an unconscious conflict and does not represent a voluntary falsification of symptoms. As such, confrontational treatment is not generally successful. PMID- 10710260 TI - Popliteal fossa neural blockade as the sole anesthetic technique for outpatient foot and ankle surgery. AB - Foot and ankle operations are being performed with greater frequency as outpatient procedures. Although the surgical procedure remains the same whether the operation is done in an inpatient or an outpatient setting, the anesthesia and postoperative analgesia are greatly affected when patients must be discharged soon after their operation. We have evaluated a regional anesthetic technique which blocks the sciatic nerve in the popliteal fossa and the saphenous nerve block at the knee. This was the sole anesthetic technique for both the operation and the immediate postoperative period. This technique appears to have several advantages: 1) Excellent anesthesia during the operation and for about ten hours postoperatively; 2) Use of a proximal calf tourniquet, and 3) Absence of systemic or local complications as might be seen with general, spinal or epidural anesthesia. PMID- 10710261 TI - Progressive pedal macrodactyly surgical history with 15 year follow-up. AB - Macrodactyly can affect the fingers and/or toes1. Histopathologic examination will distinguish macrodactylia fibrolipomatosis or neural fibrolipoma with macrodactyly, from macrodactylia as a part of neurofibromatosis. Surgical repair is aimed at decreasing the size of the affected foot so it is as near in size and shape to the normal foot as possible. Surgical approaches have included reconstructive surgery (usually staged debulking procedures), epiphyseal plate arrest and amputation. Repeated reconstructive surgical procedures, as illustrated in this report covering patient care over a 15 year period, are usually necessary due to recurring soft tissue and boney enlargement. PMID- 10710262 TI - Chronic recurrent subluxation of the peroneal tendons in a pediatric patient. Surgical recommendations. AB - Numerous surgical and non-operative approaches have been used to treat chronic recurrent subluxation of the peroneal tendons in adult athletes. There have been no published reports of surgical repair in children. In this report on a skeletally immature patient a modification of the Chrisman-Snook procedure (previously described for lateral ligament reconstruction) is described to correct recurrent subluxation of the peroneal tendons, child. PMID- 10710263 TI - Growth disturbances after distal tibial physeal fractures. AB - Twenty-four patients with distal tibial growth disturbance were reviewed. Disturbances were classified as physeal bar (prior to deformity), angular, linear or combined deformities. Treatment consisted of osteotomy in fourteen, epiphyseodesis in seven, excision of bony bar in two, and observation in one patient. Follow up was an average 36.6 months (range 4-129 months) after treatment of growth disturbance. The age at time of injury was 10.4 years of age average (range 3-15 years). There were 12 SH2, 2 SH3, 7 SH4, and 3 SH5 distal tibial physeal fractures. Thirteen of 15 fractures considered high energy and only 1 of 9 fractures considered low energy resulted in angular deformity. Angular and linear deformities presented an average 46 months (range 12-120 months) and physeal bars at an average 14 months (range 6-25 months) after injury. Patients with a delay in presentation of growth disturbance greater than 24 months had angular deformities in 92% compared with 33% in children presenting less than or at 24 months. Treatment based on type of deformity, age at time of injury, and growth remaining was considered successful in 83%. Patients with angular or linear deformities were more likely to present late, have high energy injuries, be male patients and have Salter-Harris types IV and V. Early diagnosis and treatment of growth disturbance can prevent severe deformity. PMID- 10710264 TI - Painful idiopathic rigid flatfoot in children and adolescents. AB - Nine patients (13 feet) were identified whose primary complaints were of atraumatic-onset, chronic pain in the hindfoot exacerbated with increased activity and who had the diagnosis of idiopathic rigid flatfeet. Eight of 11 were greater than the 95th percentile in weight for their age. Exam under anesthesia showed moderate to significant improvement in hindfoot motion in 9 feet; 4 feet required fractional peroneal lengthenings. Only 5 of 11 patients have had sustained relief of pain and report unlimited activity level. Children and adolescents with painful idiopathic rigid flatfeet without known causation can have significant, persistent, disability and do not uniformly respond well to traditionally-described nonoperative Interventions. PMID- 10710265 TI - Intraoperative imaging of the tibial plafond fracture: a potential pitfall. AB - STUDY DESIGN: Human tibial plafond cadaveric specimens were coronally sectioned and imaged to assess the accuracy of evaluation of ankle joint line congruity using anteroposterior radiography. Two interesting representative clinical cases are discussed. OBJECTIVES: To evaluate the validity of the routine use of anteroposterior radiographs to evaluate intra-operative ankle joint line congruity in circumstances where lateral radiographs are infeasible due to obscuring internal or external hardware. METHODS: Eleven frozen human cadaveric lower extremity specimens were used in this study. At the level of the tibial plafond, the specimens were sequentially sliced into 0.5cm sections in the coronal plane. True anteroposterior radiographs were taken with the specimen en bloc. Sequentially, the posterior slices were removed one by one, with an image taken after removing each section. The process was then repeated by removing the anterior sections sequentially with intervening radiographs. Each series of anteroposterior radiographs was then evaluated to characterize which portion of the joint line on the whole specimen view had been contributed by each of the sections. This then allowed us to make inferences about the evaluation of the joint line if it had been derived solely by anteroposterior radiography. Two poignant clinical cases demonstrating the clinical relevance of this information are discussed. RESULTS: By sequentially imaging after removing coronal sections of the tibial plafond we were able to accurately characterize the contribution of each portion of the plafond to the overall anteroposterior view. By primarily imaging the anterior portions of the plafond, with the posterior portions removed, the joint line image was virtually unchanged from the en bloc anteroposterior radiograph. However, removal of the anterior coronal sections caused large variation in the joint line image. These observations demonstrate that the anteroposterior radiograph of the tibial plafond characterizes the anterior portion of the joint well, while it represents a poor assessment of the posterior portion of the joint. This was well illustrated in our clinical case presentations. CONCLUSION: In severe fractures of the tibial plafond multiple forms of internal and external devices are frequently used for fixation. In these circumstances hardware may obscure the lateral view making it impossible to obtain adequate lateral radiographs to assess fracture reduction and joint line congruity. In this scenario, the anteroposterior radiograph is frequently relied upon to confirm the anatomic relationship of the displaced fragments. However, this view fails to accurately characterize reduction in the entire joint line and, intra-operatively, may mislead the surgeon to accept a reduction as anatomic when intra-articular incongruity still exists. Strict attention to pre-operative radiographs and the use of additional rotated views may aid the surgeon in this setting to assess fracture reduction and joint line congruence. PMID- 10710266 TI - Containment bag for jet lavage irrigation of the foot and ankle. AB - A clear plastic surgical bag which has an incorporated closure string may be used to contain fluid during jet lavage irrigation of foot and ankle wounds. This method may prevent fluid from spraying about, improving barrier precautions for both patient and operating room personnel. PMID- 10710267 TI - Preventive foot care in people with diabetes. American Diabetes Association. PMID- 10710268 TI - Effect of resistance exercise (body building) training on serum leptin levels in young men. Implications for relationship between body mass index and serum leptin. AB - Available data about the influence of exercise on leptin level are controversial, and there are no studies concerning leptin levels in trained men with low fat mass plus large increase of muscle. 65 healthy young male matched for age were separated in three groups. 1) 25 non-professional body builders; 2) 21 mild overweight sedentary subjects; 3) 19 normal weight sedentary controls. Body composition was determined by bioelectrical impedance. Serum leptin was measured in duplicate by RIA. STATISTICS: Student's t and Pearson's test. Athletes showed similar BMI than overweight subjects: 26.98+/-0.49 vs 27.12+/-0.41 but lower fat mass: 12.53+/-0.96 vs 16.16+/-1.01 % (p=0.0064) and lower leptin: 4.66+/-0.51 vs 7.31+/-0.76 microg/l (p=0.014). Athletes showed higher BMI than controls: 26.98+/ 0.49 vs 23.08+/-0.30 (p<0.0001) but similar fat mass: 12.53+/-0.96 vs 12.48+/ 0.73% and leptin: 4.66+/-0.51 vs 4.79+0.58 microg/l. Overweight subjects showed higher BMI than controls: 27.12+/-0.41 vs 23.08+/-0.30 (p<0.0001), higher fat mass: 16.16+/-1.01 vs 12.48+/-0.73% (p=0.0064) and higher leptin: 7.31+/-0.76 vs 4.79+/-0.589 microg/l (p=0.014). When leptin was calculated by fat mass no differences were observed between the three groups. There was a significant correlation between leptin and fat mass in all groups. Leptin correlated with BMI in overweight subjects (r=0.438, p=0.0463), but this correlation was not observed either in athletes or in controls. In conclusion 1) regardless of the high BMI characteristic of body builders, no correlation was observed with leptin; 2) trained state induced by resistance exercise does not influence leptin production independently of variations in body composition. PMID- 10710269 TI - Spontaneously occurring anti-PTH autoantibodies must be considered in the differential diagnosis of patients with elevated serum PTH levels. AB - We investigated the cause of elevated immunoreactive circulating parathyroid hormone (PTH) levels in two females of 41 (case 1) and 39 (case 2) years of age with low/normal serum calcium levels and hypocalcemia, respectively, and, in the latter case, hyperprolactinemia. Serum samples from both patients were fractionated by Sephadex G-100 Superfine chromatography. Fractions were assayed for PTH and prolactin (PRL) by immunoradiometric assays (IRMA) and for immunoglobulin G (IgG) by radial immunodiffusion. Sera from both patients were incubated with protein A and protein G Sepharose, centrifugated and the supernatant was assayed for PTH by IRMA. Sera were also subjected to affinity chromatography with an anti-human-IgG-agarose column. IgG and PTH or PRL were measured in the fractions by radial immunodiffusion and IRMA, respectively. In both cases the majority of serum PTH immunoreactivity eluted in the same fractions of IgG after gel filtration and was precipitated by protein A (89% in patient 1 and 96% in patient 2) and protein G (83% in patient 1 and 100% in patient 2), thus, behaving as IgG. In case 1, 79% of PTH was also retained by an anti-hIgG agarose column. High PRL levels in patient 2 were due to macroprolactinemia since most of PRL eluted as big,big (40%) and big-PRL (45%) after gel filtration. Forty-eight percent of PRL from patient 2 was retained by the anti-hIgG column indicating the presence of an anti-PRL autoantibody. These data suggest that spontaneously occurring anti-PTH autoantibodies must be considered in the differential diagnosis of patients with elevated serum PTH levels. PMID- 10710270 TI - A rapid procedure for the quantitation of natriuretic peptide RNAs by competitive RT-PCR in congenital heart defects. AB - We report an original application of quantitative reverse transcription polymerase chain reaction (Q.RT-PCR) for the evaluation of atrial natriuretic factor (ANF), brain natriuretic peptide (BNP) and beta-actin mRNA in small atrial samples (10-15 mg). A fixed amount of total RNA was simultaneously reverse transcribed and amplified with dilutions of a competitor RNA fragment used as a control. We constructed a single synthetic RNA competitor for ANF, BNP and beta actin. The competitors and targets shared the same primer sequences but yielded PCR products of different sizes. We have investigated ANF, BNP and beta-actin gene expression in patients affected by congenital heart defects (CHD) during the surgical correction of the defect. We collected a right atrial sample from each patient before the start of cardiopulmonary bypass. We studied 14 patients affected by tetralogy of fallot (no. 6), complex CHD (no. 4), and ventricular septal defect (no. 4). The results indicate that the Q.RT-PCR represents a simple, highly specific, non radioactive procedure for the quantification of natriuretic peptide gene expression and is particularly suitable for evaluation of gene expression in small myocardial samples. Our data also suggest that: 1) there is a significant relationship between the levels of ANF, BNP and beta-actin mRNA and the type of CHD; 2) the levels of BNP mRNA are more variable than ANF; 3) the beta-actin seems to be unsuitable for normalising cardiac gene expression in CHD because mRNA basal levels of this protein vary greatly among patients with different type of CHD. PMID- 10710271 TI - Ontogeny and regulation of variant thyroid hormone receptor isoforms in developing rat testis. AB - High affinity-low capacity nuclear triiodothyronine (T3) receptors (TRs), identified as a product of c-erbAalpha proto-oncogene, are expressed in prepubertal rat Sertoli cell. At this age, exogenous T3 treatment as well as hypothyroidism affects Sertoli cell functions. We examined the ontogenetic expression pattern of TRs in the rat testis. Northern analysis confirms that TRs are expressed at high level from fetal development until prepubertal period. RNase protection analysis demonstrates that TRalpha2, the variant isoform of TRalpha1, is constitutively expressed at all ages, while TRalpha3 is absent in the adult gonad. While TRalpha1 and TRalpha2 expression declines during development, Rev-erbAalpha (Rev), the antisense mRNA encoded by the same c erbAalpha genomic locus, increases beginning 5 days after birth and maximizing in adulthood. TRalpha1, TRalpha2, and Rev mRNAs do not appear to be directly regulated by thyroid hormone in testis; however, short-term neonatal hypothyroidism leads to the expression of TRalpha1 and its variant in adult testis, which is absent in control coeval animals. Thus, during development of rat testis, the levels of messages of genes encoded in the c-erbAalpha. genomic locus have different ontogenetic control. The ontogenetic profile of TRalpha1 and its variant isoforms within the seminiferous epithelium suggests that these receptors are involved in the differentiation of the male gonad. PMID- 10710272 TI - Minimally invasive surgery for thyroid small nodules: preliminary report. AB - Cytological assessment of cold thyroid nodules cannot exclude malignancy in case of follicular tumors. Many follicular nodules undergo surgery although most of them later on prove to be benign. We report a new minimally invasive video assisted approach (MIVA) for the treatment of thyroid lesions with a diameter minor than 3 cm. Ten females and 2 males (mean age: 37 yr) with a cold thyroid nodule and a cytological diagnosis of microfollicular tumor were selected for MIVA hemythyroidectomy. The procedure was carried out through a 15 mm incision with needlescopic instruments and a 30 infinity 5-mm endoscope. Mean operative time was 87 min (range 60-120). No complications were registered. Cosmetical result was excellent in all patients. MIVA hemythyroidectomy is safe and effective; indications and limits of this new procedure require further studies. PMID- 10710273 TI - Importance of bioavailable calcium drinking water for the maintenance of bone mass in post-menopausal women. AB - The aim of this research was to establish the importance of calcium intake through mineral water on vertebral bone density in women. To this purpose, we examined 255 women divided into two groups: those regularly drinking a high calcium content mineral water (group A; no.=175) and those using different type of water with a lower calcium content (group B; no.=80). Their dietary daily calcium intake was determined by means of a validated questionnaire (N.I.H. Consensus statement) and vertebral bone density was measured by Dual-Energy X-ray absorptiometry (Unigamma-plus ACN densitometer). Women in group A ingested a significantly higher quantity of calcium in water than women in group B (mean difference 258 mg; 95% confidence limits: 147-370 mg). The average bone density values were slightly but significantly higher in group A as compared to group B (mean+/-SD: 1.044+0,15 vs 1.002+0,14; p=0.03). In addition to age, BMI and menopausal status, calcium intake was a significant predictor of spinal BMD. These 4 variables explained about 35% of the spinal BMD variance. When the analysis was repeated separately for pre- and post-menopausal subjects, calcium remained a significant predictor in post-menopausal women (t=2.28; p=0.02), but not in premenopausal women. These results underline the importance of a lifelong daily calcium intake, resulting by the regular drinking of high bioavailable calcium water, in order to maintain bone mass after the menopause, in comparison to the use of a lower content calcium water. PMID- 10710274 TI - Persistent hypokalemia after successful adrenalectomy in a patient with Cushing's syndrome due to ectopic ACTH secretion: possible role of 11beta-hydroxysteroid dehydrogenase inhibition. AB - Ectopic ACTH secretion is characterized by a high incidence of hypokalemia. The pathophysiology of hypokalemia has not been totally clarified, although it has been postulated that excessive amounts of adrenal steroids may play a role, as well as a possible role of the inhibition of the enzyme 11beta-hydroxysteroid dehydrogenase (11beta-OHSD). This enzyme normally converts cortisol to cortisone avoiding the mineralocorticoid action of cortisol. We present a patient with ectopic ACTH secretion due to a metastatic carcinoid tumor. The clinical picture was characterized by maintained hypokalemia (1.4 mmol/l) resistant to potassium, spironolactone and ketoconazole administration. A bilateral adrenalectomy was performed but the hypokalemia persisted while he was receiving a physiological dose of cortisol. Eight days after adrenalectomy cortisol was replaced by an equivalent dose of dexamethasone. This change was followed by a rapid and persistent normalization of hypokalemia suggesting a mineralocorticoid effect of cortisol. In conclusion, the origin of hypokalemia in our patient with ectopic ACTH secretion was secondary to cortisol. We postulate that this peculiar effect of cortisol could have happened if an inhibition of 11beta-OHSD occurred. PMID- 10710275 TI - Complete remission of Nelson's syndrome after 1-year treatment with cabergoline. AB - In this case report we demonstrated that treatment with the long-acting D2 receptor agonist cabergoline for 1 year induced normalization of plasma ACTH levels and disappearance of the pituitary tumor in a patient with Nelson's syndrome. A young man underwent bilateral adrenalectomy and subsequent pituitary irradiation for Cushing's disease after unsuccessful neurosurgical treatment. Thereafter, he was given cortisone acetate replacement at the dose of 62.5 mg a day. Fifteen months after pituitary irradiation, he developed Nelson's syndrome, having skin hyperpigmentation, high plasma ACTH levels (376 ng/l) and a pituitary microadenoma (5 mm) documented at magnetic resonance imaging (MRI) of the pituitary region. After 6 months of cabergoline treatment, given at the dose of 1 mg a week, plasma ACTH levels were significantly decreased (from 376 to 113 ng/l) but they were not normalized. Cabergoline dose was then increased up to 2 mg a week. Six months later plasma ACTH levels were normalized (22 ng/l) and MRI demonstrated the disappearance of the pituitary adenoma. In order to investigate on the direct effect played by cabergoline treatment on the remission of Nelson's syndrome, the treatment was withdrawn. Plasma ACTH levels significantly increased (119 ng/l) after 3 months of treatment withdrawal. At the last follow-up, during cabergoline treatment at the dose of 2 mg/week plasma ACTH levels were normalized (40.4 ng/l). This case demonstrated that cabergoline treatment is able to induce the remission of Nelson's syndrome and may be a valid therapeutic alternative in this syndrome. PMID- 10710276 TI - Cold thyroid nodule as the sole manifestation of Rosai-Dorfman disease with mild lymphadenopathy, coexisting with chronic autoimmune thyroiditis. AB - A case of thyroid Rosai-Dorfman disease (RDD) without apparent lymphadenopathy in a 49-year-old woman with underlying euthyroid chronic autoimmune thyroiditis, as indicated by high thyroid autoantibodies titers, is presented. The initial presentation was that of a cold, hypoechogenic nodule of left thyroid lobe which increased in size during the two years of follow up, together with new ultrasonographic findings of the right lobe. No biochemical abnormalities were found apart from mild hypercalcemia. A near total thyroidectomy was performed. Histologically, the left lobe nodule as well as the right lobe lesions consisted of typical RDD cellular population, with the pathognomonic phenomenon of emperipolesis. Infiltration to the periphery of the gland was observed and three adjacent lymph nodes were also involved. The uninvolved thyroid parenchyma showed changes compatible with chronic autoimmune thyroiditis. No other localizations or systemic manifestations of RDD were revealed. Normocalcemia was restored promptly and the patient remains free of clinically overt disease one year post operatively. PMID- 10710277 TI - Is presurgical treatment with somatostatin analogs necessary in acromegalic patients? PMID- 10710279 TI - Growth hormone deficiency in APECED. PMID- 10710280 TI - Endocrinology and art. "The Baptism of the Princes of Marseille" (detail). PMID- 10710278 TI - Light, blindness and endocrine secretions. PMID- 10710281 TI - Canine vagus nerve stores cholecystokinin-58 and -8 but releases only cholecystokinin-8 upon electrical vagal stimulation. AB - Cholecystokinin-58 has been shown to be the major form of cholecystokinin (CCK) released to the circulation upon lumenal stimulation of the small intestine in humans and dogs. In anesthetized dogs, electrical vagal stimulation evokes pancreatic exocrine secretion that is in part mediated through the release of CCK. We studied the molecular form of CCK stored in canine vagus nerves and that released into circulation upon electrical vagal stimulation. Gel filtration and radioimmunoassay of the water and acid extracts of canine vagus nerves indicated CCK-8 (35%) and CCK-58 (65%) as the major molecular forms in the vagus nerve. Both forms of CCK isolated from the vagal extracts were equally bioactive as the standard CCK-8 and CCK-58, respectively, in stimulation of amylase release from isolated rat pancreatic acini. Analysis of plasma collected after electrical vagal stimulation indicated that CCK-8 is the only form released into the circulation. The release of CCK-8 upon electrical vagal stimulation was not affected by application of lidocaine to the upper small intestinal mucosa, suggesting that it was released from vagal nerve terminals. PMID- 10710282 TI - Identification of an NPY-Y1 receptor subtype in bovine chromaffin cells. AB - Bovine chromaffin cells have been used in a variety of studies designed to reveal different aspects of neuropeptide Y (NPY) action. Pharmacological data have defined five NPY receptor subtypes, only one of which (Y3) has not been cloned. Some studies with bovine chromaffin cells have concluded that the effects of NPY on this cell type are mediated by the Y3 subtype. Previous work from our laboratory demonstrates that a Y1 subtype mediates the effect of NPY in this tissue. In the current studies we provide further evidence for the existence of the Y1 subtype in bovine chromaffin cells. BIBP3226, the selective Y1 antagonist, potently displaces [125I]NPY from its binding site IC50 = 1.91 x 10(-9) M. Moreover, [125I]BIBP3226 binds to bovine chromaffin cell membranes with high affinity (IC50 = 5.9 x 10(-8) M). Examination of BIBP3226 antagonism of NPY inhibition of forskolin stimulated cyclic AMP accumulation reveals that it is a competitive antagonist with a K(B) similar to the IC50 for [125I]BIBP3226 binding. Northern blot analysis using a porcine cDNA clone for the Y1 subtype demonstrates a 3.5-kb mRNA species in chromaffin cells. These data identify the bovine chromaffin cell NPY receptor as a Y1 subtype. PMID- 10710283 TI - Changes in plasma adrenomedullin levels during the menstrual cycle. AB - We investigated whether the levels of adrenomedullin, a novel peptide produced by several tissues, including the pituitary gland, change during the ovarian cycle. We studied 13 healthy women with regular menstrual cycles. Plasma samples were collected at 7, 14, 21 and 28 days of the ovarian cycle and assayed for adrenomedullin 1-52 using a specific RIA. LH, FSH, 17beta-estradiol, and progesterone concentrations were also determined. The adrenomedullin profile during ovarian cycle was similar to that of LH; plasma adrenomedullin increased from 10.9 pg/ml at the 7th day to 15.1 pg/ml at the 14th, and decreased to 8.5 pg/ml in the subsequent menses. The changes in plasma adrenomedullin were related to changes in LH and 17beta-estradiol. The cause of the increase in adrenomedullin levels during the late follicular phase of the menstrual cycle is not clear. Since it has been demonstrated that adrenomedullin is involved in the regulation of hypothalamus-pituitary-adrenal gland and its secretion is regulated by sex hormones we speculate that adrenomedullin could also play a role in regulating the hypothalamus-pituitary-ovary feedback. Alternatively it may be involved in the regulation of fluid and electrolyte homeostasis during the menstrual cycle. PMID- 10710284 TI - Evidence that NPY Y1 receptors are involved in stimulation of feeding by orexins (hypocretins) in sated rats. AB - Neuropeptide Y (NPY) produced in the arcuate nucleus (ARC) of the hypothalamus stimulates feeding both directly by activating NPY receptors and indirectly through release of the orexigenic peptides, galanin and beta-endorphin (beta END), in the paraventricular nucleus (PVN) and surrounding neural sites. Orexin A and orexin B, produced outside the ARC in the lateral hypothalamic area (LH), have recently been shown to stimulate feeding. In the present studies we tested the hypothesis that NPYergic signaling may mediate feeding stimulated by orexins. In adult male rats injected intracerebroventricularly (i.c.v.) with orexin A (3, 10, 15 nmol) or orexin B (3, 10, 30 nmol) feeding was stimulated in a dose dependent manner; maximal feeding was seen after 15 nmol orexin A and 30 nmol orexin B. To determine whether NPY may mediate this orexin stimulated feeding, we used 1229U91, a selective NPY Y1 receptor antagonist (NPY-A). Whereas NPY-A on its own was ineffective, it suppressed NPY-induced feeding. Furthermore, NPY-A completely blocked the feeding evoked by either orexin A (15 nmol) or orexin B (30 nmol). These results show that orexin A and B stimulate feeding and further suggest that these excitatory effects may be mediated by NPYergic signaling through Y1 receptors. These findings are in accord with the view that the orexin NPY pathway may comprise a functional link upstream from NPY within the hypothalamic appetite regulating network. PMID- 10710285 TI - Differential glomerular response to continuous infusion of vasopressin in spontaneously hypertensive rats and Wistar-Kyoto rats. AB - To determine the difference of glomerular response to exogenous vasopressin (VP) in vivo between normotensive and hypertensive rats, we examined the effects of 14 day continuous infusion of VP (1.0 ng/kg/min) on the physiological and histological aspects in 7-week-old spontaneously hypertensive rats (SHR) and Wistar-Kyoto (WKY) rats. VP infusion did not result in significant changes in systolic blood pressure, heart rate, serum electrolytes, serum creatinine, urinary protein and N-acetyl-beta-glucosaminidase levels in both strains of rats. VP infusion significantly reduced daily urine volume associated with significant concentration of the urine in WKY rats but not SHR. Kidney and heart weights did not differ significantly after VP infusion between both strains. Glomerular mesangial expansion was significantly enhanced in VP infused SHR, but glomerular cellularity was not different between both strains following treatment. Competitive reverse transcription-polymerase chain reaction revealed that the level of glomerular transforming growth factor (TGF)-beta1 mRNA was significantly higher in SHR than WKY rats, and that this difference was significantly augmented after VP infusion in SHR. VP infusion, however, did not change the level of glomerular mRNAs of platelet-derived growth factor (PDGF) B-chain in both strains. Then, exogenous VP infusion contributes to the glomerular mesangial expansion in SHR, which involved overexpression of glomerular TGF-beta1 without any pressor effect. In contrast, the significant changes of glomerular expansion and TGF-beta1 level were not shown in WKY rats. These findings suggest that the glomerular response to the exogenous VP is preferentially enhanced in SHR. PMID- 10710286 TI - Chromogranin-B regulation of IAPP and insulin secretion. AB - In the endocrine pancreas, chromogranin-A and pancreastatin have been suggested to inhibit islet insulin secretion, whereas chromogranin-B has not been studied in this context. Furthermore, a putative effect by chromogranins on IAPP secretion is unknown. We aimed to elucidate the endogenous effect of chromogranin A, pancreastatin and chromogranin-B on islet insulin and IAPP secretion from pancreatic NMRI mouse islets. In acute experiments, there was a tendency towards an increase in insulin release, which became more manifest after a 48-h exposure. Moreover, 48 h exposure to chromogranin-B antiserum resulted in a significant increase in (pro)insulin synthesis. Neither antibodies against chromogranin-A nor pancreastatin had any effect on islet hormone secretion. None of the antibodies tested had any effect on islet IAPP or insulin content. We suggest that chromogranin-B released from islets may have an autocrine inhibitory effect on islet IAPP and insulin secretion. Our data imply a regulatory role of chromogranin-B in islet IAPP and insulin secretion. PMID- 10710287 TI - Thyrotropin-releasing hormone interacts with vasoactive intestinal peptide specific receptor in guinea pig cecal circular smooth muscle cells. AB - The relationship between thyrotropin-releasing hormone (TRH) binding sites and vasoactive intestinal peptide (VIP) receptors in circular muscle cells obtained from the guinea pig cecum was investigated using antagonists of VIP receptors and a selective receptor protection method. Both VIP10-28, a VIP antagonist, and atrial natriuretic peptide1-11 (ANP1-11), a VIP-specific receptor antagonist, completely inhibited 10(-5) M TRH-induced relaxation in a concentration-dependent manner. The muscle cells where cholecystokinin octapeptide (CCK-8) and TRH binding sites were protected completely preserved the inhibitory responses to TRH and ANP (a VIP-specific receptor agonist), and partially the inhibitory response to VIP. Peptide histidine isoleucine (PHI: a VIP-preferring receptor agonist) had no inhibitory effect on these cells. The muscle cells where CCK-8 and ANP (VIP specific) receptors were protected completely preserved the inhibitory responses to TRH and ANP and partially the inhibitory response to VIP. PHI had no inhibitory effect on these cells. The muscle cells where CCK-8 and VIP receptors (both VIP-specific and VIP-preferring receptors) were protected preserved completely the inhibitory responses to TRH, VIP, ANP, and PHI. The muscle cells where CCK-8 and PHI (VIP-preferring) receptors were protected completely preserved the inhibitory response to PHI and partially the inhibitory response to VIP. TRH and ANP had no inhibitory effect on these cells. This study first demonstrates that TRH interacts with VIP-specific receptor in guinea pig cecal circular smooth muscle cells. PMID- 10710288 TI - 2-36[K4,RYYSA(19-23)]PP a novel Y5-receptor preferring ligand with strong stimulatory effect on food intake. AB - Members of the neuropeptide Y (NPY) family regulate many physiological processes via interaction with at least four functional, pharmacologically distinct Y receptors. However, selective antagonists developed for several subtypes have not been useful in defining particular Y-receptor functions in vivo. To identify critical residues within members of the NPY family required for Y-receptor subtype-selectivity we have determined the contribution of each residue within NPY to receptor binding by replacing them with L-alanine. In a second study, chimeric peptides where single or stretches of residues were interchanged between members of the NPY family were generated and tested in radioligand binding studies. Overall, substituted alanine analogues exhibited similar orders of affinities at each Y-receptor subtype with no obvious subtype-selectivity. Residues of particular interest are Leu30 which exhibited selectivity for the Y4 receptor, whereas Asp16 does not appear to play any role in ligand binding. Several chimeric peptides, e.g., [K4]pancreatic polypeptide ([K4]PP) and [RYYSA(19-23)]PP clearly showed higher affinity at the Y4 and Y5 subtypes compared to the Y1 and Y2 subtypes. In addition, the transfer of a proline residue from position 14 to 13 in peptide YY decreases its affinity at the Y1-, Y4- and Y5-receptors but is unchanged at the Y2 subtype. Combining these results, and with the help of molecular modelling, second generation chimeras were designed. The most significant improvement was achieved in chimera 2 36[K4,RYYSA(19-23)]PP where the affinity for the Y5 subtype increased by ninefold over that from NPY. Several of these compounds were also tested for their ability to stimulate food intake in a rat model. Interestingly, again 2-36[K4,RYYSA(19 23)]PP showed the most dramatic effect with a major increase on food intake over a range of doses compared to NPY suggesting a possible synergistic effect of several Y-receptors on feeding behaviour. PMID- 10710289 TI - Ligand association with the rabbit kidney and brain Y1, Y2 and Y5-like neuropeptide Y (NPY) receptors shows large subtype-related differences in sensitivity to chaotropic and alkylating agents. AB - The binding to rabbit kidney or hypothalamic particulates of the subtype selective neuropeptide Y (NPY) receptor ligands [125I](Leu31,Pro34)hPYY (as Y1 site label at 2 nM human pancreatic polypeptide (hPP)), [125I]-hPYY(3-36) (Y2 label), and [125I]-hPP (Y5 label) displayed great differences in sensitivity to alkylators and chaotropic agents. Sensitivity to a nonionic chaotrope, urea, was much higher for the Y1 binding than for the Y5-like binding or the Y2 binding. The non-selective alkylator N-ethylmaleimide (NEM) and several alkylators selective for aminergic receptors were much more efficacious against the Y1 relative to the Y2 binding. Similar differences could be confirmed with the attachment of Y1 and Y2-selective tracers to CHO cells expressing the cloned guinea-pig Y1 or Y2 receptors. The Y5-like binding was quite insensitive to NEM, but sensitive to chloroethylclonidine (CEC) and prazobind, which were less potent at the Y1, and especially at the Y2 site. The unrestricted-access alkylator 2 aminoethyl methanethiosulfonate inhibited the binding to all subtypes, while the restricted-access agent 2-(trimethylammonium)ethylmethanethiosulfonate poorly inhibited the Y5-like binding, or the guanine nucleotide-insensitive Y2 binding. These results are compatible with an active conformation of the Y5-like site dependent on maintenance of a shared hydrophobic cavity. The Y2 sites resistant to guanosine polyphosphates and restricted-access alkylators were detected mainly in particulates slowly solubilized by cholate at 0-5 degrees C; these sites could be clustered. PMID- 10710290 TI - Identification of an interaction between the angiotensin II receptor sub-type AT2 and the ErbB3 receptor, a member of the epidermal growth factor receptor family. AB - To identify the proteins that interact and mediate angiotensin II receptor AT2 specific signaling, a random peptide library was screened by yeast-based Two Hybrid protein-protein interaction assay technique. A peptide that shared significant homology with the amino acids located between the residues Gly-Xaa Gly-Xaa-Xaa-Gly721 and Lys742, the residues predicted to be important for ATP binding of the ErbB3 and ErbB2 receptors, was identified to be interacting with the AT2 receptor. The interaction between the human ErbB3 receptor and the AT2 receptor was further confirmed using the cytoplasmic domain (amino acids 671-782) of the human ErbB3 receptor. Moreover, an AT2 receptor peptide that spans the amino acids 226-363, (spans the third ICL and carboxy terminal domain) could also interact with the AT2 receptor in a yeast Two-Hybrid protein-protein interaction assay. Studies using mutated and chimeric AT2 receptors showed that replacing the third intracellular loop (ICL) of the AT2 receptor with that of the AT1 abolishes the interaction between the ErbB3 and the AT2 in yeast Two-Hybrid protein-protein interaction assay. Thus the interaction between the AT2 receptor and the ErbB3 receptor seems to require the region spanning the third ICL and carboxy terminus of the AT2 receptor. Since the third ICL of the AT2 receptor is essential for exerting its inhibitory effects on cell growth, possible involvement of this region in the interaction with the cytoplasmic domain of the ErbB3 receptor suggests a novel signaling mechanism for the AT2 receptor mediated inhibition of cell growth. Furthermore, since both the AT2 and the ErbB3 receptors are expressed during fetal development, we propose that the existence of direct interaction between these two receptors may play a role in the regulation of growth during the initial stages of development. PMID- 10710292 TI - Incorporating a statistically based shape model into a system for computer assisted anterior cruciate ligament surgery. AB - This paper addresses the problem of extrapolating very sparse three-dimensional (3-D) data to obtain a complete surface representation. A new method that uses statistical shape models is proposed and its application to computer-assisted anterior cruciate ligament (ACL) reconstruction is detailed. The rupture of the ACL has become one of the most common knee injuries. One problem during reconstruction is to find the optimal attachment points for the graft. Therefore a system for computer-assisted reconstruction of the ACL has been proposed by TIMC laboratory. During surgery the surgeon collects several data points on the tibial and femoral joint surface with a 3-D localizer system. These 3-D data are used to find those attachment points resulting in a low anisometry of the graft, while preventing impingement between the graft and the femoral notch. As the collected data points only cover a small surface patch of the femur, it is desirable to extrapolate these data to also have a visualization in those areas where no data points are available. A sufficiently good approximation of the actual femur by the model would further allow us to better deal with the notch impingement problem of the graft. The chosen approach is to fit a deformable model to the data points, it can be subdivided into two steps, constructing the model and fitting this model to the data. To incorporate a priori knowledge into the model, the allowed deformations are determined by the statistics of the shape variation of a set of training objects. Matching the training objects together is obtained by elastic registration of surface points using octree splines. The fitting process of the sparse intra-operative data with the statistical model results in a non-linear multi-dimensional function minimization. A hybrid search strategy combining local and global methods is used to avoid local minima. First experimental results with a model generated from 10 femurs are presented, including fitting of the model with both simulated and real intra-operative data. PMID- 10710291 TI - Does the response of the intestinal epithelium to keratinocyte growth factor vary according to the method of administration? AB - BACKGROUND: Keratinocyte growth factor (KGF) is a potent mitogen and may be of value for the treatment of conditions such as short bowel syndrome and chemotherapy-induced mucositis. However the most efficacious route and method of administration is unclear. METHODS: Rats maintained by total parenteral nutrition (TPN) were given KGF (1 mg/kg/rat/day, i.v.) infused continuously or as a once daily injection. The same dose was also given s.c. to chow-fed rats. Changes in gut growth were assessed by measurement of wet weight, proliferation (vincristine induced metaphase arrest) and crypt branching index. Changes in gut hormone profile were also determined to examine if any trophic effects were mediated via this mechanism. RESULTS: KGF caused a 70-100% increase in wet weight of the stomach, small and large intestine of TPN-fed rats (P < 0.01) with no significant differences seen between the two methods of administration. The increase in metaphase counts was greatest in the stomach (about seven-fold P < 0.01), but was less pronounced in the distal small intestine and colon (about 50% increase). The trophic effect of KGF was much less prominent in orally-fed rats. Crypt branching index was significantly reduced by KGF in the proximal small intestine of TPN, but not orally-fed rats. Plasma gastrin, PYY, total glucagon, enteroglucagon and GLP-1 all increased by two-three-fold (all P < 0.01) in response to KGF whereas insulin levels fell by about 25% in the TPN group. CONCLUSIONS: The mitogenic action of KGF occurred predominantly in the stomach and proximal small intestine. Its efficacy was less pronounced in orally-fed animals, suggesting KGF may be of greatest benefit in conditions associated with lowered intestinal proliferation. Clinical trials of KGF can probably use single daily i.v. injections without reduction in efficacy. PMID- 10710293 TI - Brachytherapy optimal planning with application to intravascular radiation therapy. AB - We have been studying brachytherapy planning with the objective of minimizing the maximum deviation of the delivered dose from prescribed dose bounds for treatment volumes. A general framework for optimal treatment planning is presented and the minmax optimization is formulated as a linear program. Dose rate calculations are based on the dosimetry formulation of the American Association of Physicists in Medicine, Task Group 43. We apply the technique to optimal planning for intravascular brachytherapy of intimal hyperplasia using ultrasound data and 192Ir seeds. The planning includes determination of an optimal dwell-time sequence for a train of seeds that deliver radiation while stepping through the vessel lesion. The results illustrate the advantage of this strategy over the common approach of delivering radiation by positioning a single train of seeds along the whole lesion. PMID- 10710294 TI - CARABEAMER: a treatment planner for a robotic radiosurgical system with general kinematics. AB - Stereotactic radiosurgery is a minimally invasive procedure that uses a focused beam of radiation as an ablative instrument to destroy brain tumors. To deposit a high dose of radiation in a tumor, while reducing the dose to healthy tissue, a large number of beams are crossfired at the tumor from multiple directions. The treatment planning problem (also called the inverse dosimetry problem) is to compute a set of beams that produces the desired dose distribution. So far its investigation has focused on the generation of isocenter-based treatments in which the beam axes intersect at a common point, the isocenter. However this restriction limits the applicability of the treatments to tumors which have simple shapes. This paper describes CARABEAMER, a new treatment planner for a radiosurgical system in which the radiation source can be arbitrarily positioned and oriented by a six-degree-of-freedom manipulator. This planner uses randomized techniques to guess a promising initial set of beams. It then applies space partitioning and linear programming techniques to compute the energy to be delivered along each beam. Finally, it exploits the results of the linear program to iteratively adapt and improve the beam set. Experimental results obtained with CARABEAMER on both patient and synthetic cases are presented and discussed. These results demonstrate that a radiosurgical system with general kinematics can deliver treatments in which the region receiving a high dose closely matches the shape of the tumor, even in complicated cases. They also suggest new research directions which are discussed at the end of the paper. PMID- 10710295 TI - Evaluation of robot-based registration for subtraction radiography. AB - Digital subtraction of (plane film) radiographs of an area of interest taken over time is a powerful diagnostic tool. In dentistry for example, this is used to detect and treat periodontal disease (i.e. the sequence of diseases that leads to the loss of bone supporting the teeth). A precondition of subtraction radiography is correct three-dimensional registration of the film and X-ray source, without which the subtraction images are meaningless. A small misalignment can be compensated for by post-processing of the digitized films (by correspondence and rectification) to remove artifacts due to positioning errors. This paper presents a novel approach to subtraction radiography which replaces customary mechanical alignment methods (such as a stent or cephalostat) with a robot. A mechanical stent is a short rod attached at one end to the X-ray source and at the other to a mechanical appliance protruding from the patient's mouth. An experimental system was constructed which creates a 'sensorized stent' by integrating a plastic mouth appliance (with the impression of the patient's teeth), a robot carrying an X-ray source and a host computer. Results showed that the robotic system was superior to the mechanical alignment approach, due to its excellent accuracy and repeatability. This resulted in much less variation in the non registered X-ray images, and in a smaller standard deviation in the intensities of subtracted images overall. The results suggest that in the future, diagnostic studies including subtraction radiography will not need either mechanical alignment (which is imprecise) or post-processing registration (which is time consuming). PMID- 10710296 TI - The effects of physical constraints in laparoscopic surgery. AB - In order to provide guidelines for the development and evaluation of advanced laparoscopic instrumentation (including teleoperated devices), we assessed the impact of constrained motion on surgeons' ability to perform standardized pick and-place and suturing tasks when using an emulation of a perfectly transparent teleoperator under direct, binocular vision. The surgeons' performance when using the emulator represents an upper bound on performance using any conceivable teleoperator with one-to-one force and motion scaling. Our analysis examines the mean differences in task completion time between three open tool configurations and two laparoscopic tool configurations with various degrees of freedom (DOFs). Fifteen laparoscopic surgeons participated in the study. We show that avoiding reversed hand and tool motions and adding DOFs significantly improves suturing performance. In the pick-and-place task, avoiding the reversed motions also improves performance, but adding DOFs to an open tool configuration does not. For both tasks, subjects who use open tools constrained to four DOF complete their tasks in approximately 38% less time than when using standard four-DOF laparoscopic tools. The marginal benefit to overall surgical time of adding two additional degrees of freedom is likely to be small (approximately 2%), although surgeons may then feel confident in attempting more difficult procedures. PMID- 10710297 TI - Hippocrate: a safe robot arm for medical applications with force feedback. AB - We have developed a robotic system to assist doctors when they are moving ultrasonic probes on a patient's skin while exerting a given effort. The probes are used to monitor arteries for cardiovascular disease prevention, namely to reconstruct the three-dimensional profile of arteries. A preliminary feasibility study making use of an industrial robot has been made to validate the force control scheme. It has proven the interest of the robotized approach for such medical applications where force control is needed. In order to comply with safety constraints, a dedicated robotic system 'Hippocrate' has been designed. This paper describes the arm and the controller architectures, with emphasis on design strategies selected to meet safety requirements. Preliminary in vivo results are presented as well as a possible new application of Hippocrate as a tool for reconstructive surgery. PMID- 10710298 TI - Computer-integrated revision total hip replacement surgery: concept and preliminary results. AB - This paper describes an ongoing project to develop a computer-integrated system to assist surgeons in revision total hip replacement (RTHR) surgery. In RTHR surgery, a failing orthopedic hip implant, typically cemented, is replaced with a new one by removing the old implant, removing the cement and fitting a new implant into an enlarged canal broached in the femur. RTHR surgery is a difficult procedure fraught with technical challenges and a high incidence of complications. The goals of the computer-based system are the significant reduction of cement removal labor and time, the elimination of cortical wall penetration and femur fracture, the improved positioning and fit of the new implant resulting from precise, high-quality canal milling and the reduction of bone sacrificed to fit the new implant. Our starting points are the ROBODOC system for primary hip replacement surgery and the manual RTHR surgical protocol. We first discuss the main difficulties of computer-integrated RTHR surgery and identify key issues and possible solutions. We then describe possible system architectures and protocols for preoperative planning and intraoperative execution. We present a summary of methods and preliminary results in CT image metal artifact removal, interactive cement cut-volume definition and cement machining, anatomy-based registration using fluoroscopic X-ray images and clinical trials using an extended RTHR version of ROBODOC. We conclude with a summary of lessons learned and a discussion of current and future work. PMID- 10710299 TI - Global biodiversity scenarios for the year 2100. AB - Scenarios of changes in biodiversity for the year 2100 can now be developed based on scenarios of changes in atmospheric carbon dioxide, climate, vegetation, and land use and the known sensitivity of biodiversity to these changes. This study identified a ranking of the importance of drivers of change, a ranking of the biomes with respect to expected changes, and the major sources of uncertainties. For terrestrial ecosystems, land-use change probably will have the largest effect, followed by climate change, nitrogen deposition, biotic exchange, and elevated carbon dioxide concentration. For freshwater ecosystems, biotic exchange is much more important. Mediterranean climate and grassland ecosystems likely will experience the greatest proportional change in biodiversity because of the substantial influence of all drivers of biodiversity change. Northern temperate ecosystems are estimated to experience the least biodiversity change because major land-use change has already occurred. Plausible changes in biodiversity in other biomes depend on interactions among the causes of biodiversity change. These interactions represent one of the largest uncertainties in projections of future biodiversity change. PMID- 10710300 TI - Lunar impact history from (40)Ar/(39)Ar dating of glass spherules AB - Lunar spherules are small glass beads that are formed mainly as a result of small impacts on the lunar surface; the ages of these impacts can be determined by the (40)Ar/(39)Ar isochron technique. Here, 155 spherules separated from 1 gram of Apollo 14 soil were analyzed using this technique. The data show that over the last approximately 3.5 billion years, the cratering rate decreased by a factor of 2 to 3 to a low about 500 to 600 million years ago, then increased by a factor of 3.7 +/- 1.2 in the last 400 million years. This latter period coincided with rapid biotic evolutionary radiation on Earth. PMID- 10710301 TI - Internal structure and early thermal evolution of Mars from Mars Global Surveyor topography and gravity. AB - Topography and gravity measured by the Mars Global Surveyor have enabled determination of the global crust and upper mantle structure of Mars. The planet displays two distinct crustal zones that do not correlate globally with the geologic dichotomy: a region of crust that thins progressively from south to north and encompasses much of the southern highlands and Tharsis province and a region of approximately uniform crustal thickness that includes the northern lowlands and Arabia Terra. The strength of the lithosphere beneath the ancient southern highlands suggests that the northern hemisphere was a locus of high heat flow early in martian history. The thickness of the elastic lithosphere increases with time of loading in the northern plains and Tharsis. The northern lowlands contain structures interpreted as large buried channels that are consistent with northward transport of water and sediment to the lowlands before the end of northern hemisphere resurfacing. PMID- 10710302 TI - Suppression of rain and snow by urban and industrial air pollution AB - Direct evidence demonstrates that urban and industrial air pollution can completely shut off precipitation from clouds that have temperatures at their tops of about -10 degrees C over large areas. Satellite data reveal plumes of reduced cloud particle size and suppressed precipitation originating from major urban areas and from industrial facilities such as power plants. Measurements obtained by the Tropical Rainfall Measuring Mission satellite reveal that both cloud droplet coalescence and ice precipitation formation are inhibited in polluted clouds. PMID- 10710304 TI - Seismic images of the far side of the Sun AB - Images of an active region on the far side of the sun were derived by applying seismic holography to recent helioseismic observations from space. Active regions are the centers of energetic phenomena such as solar flares and coronal mass ejections, whose resulting electromagnetic and particle radiation interfere with telecommunications and power transmissions on Earth and can pose significant hazards to astronauts and spacecraft. Synoptic seismic imaging of far-side solar activity will now allow anticipation of the appearance of large active regions more than a week ahead of their arrival on the east solar limb. PMID- 10710303 TI - An archaeal iron-oxidizing extreme acidophile important in acid mine drainage. AB - A new species of Archaea grows at pH approximately 0.5 and approximately 40 degrees C in slime streamers and attached to pyrite surfaces at a sulfide ore body, Iron Mountain, California. This iron-oxidizing Archaeon is capable of growth at pH 0. This species represents a dominant prokaryote in the environment studied (slimes and sediments) and constituted up to 85% of the microbial community when solution concentrations were high (conductivity of 100 to 160 millisiemens per centimeter). The presence of this and other closely related Thermoplasmales suggests that these acidophiles are important contributors to acid mine drainage and may substantially impact iron and sulfur cycles. PMID- 10710305 TI - Extreme oxygen sensitivity of electronic properties of carbon nanotubes AB - The electronic properties of single-walled carbon nanotubes are shown here to be extremely sensitive to the chemical environment. Exposure to air or oxygen dramatically influences the nanotubes' electrical resistance, thermoelectric power, and local density of states, as determined by transport measurements and scanning tunneling spectroscopy. These electronic parameters can be reversibly "tuned" by surprisingly small concentrations of adsorbed gases, and an apparently semiconducting nanotube can be converted into an apparent metal through such exposure. These results, although demonstrating that nanotubes could find use as sensitive chemical gas sensors, likewise indicate that many supposedly intrinsic properties measured on as-prepared nanotubes may be severely compromised by extrinsic air exposure effects. PMID- 10710306 TI - Hematopoietic stem cell quiescence maintained by p21cip1/waf1. AB - Relative quiescence is a defining characteristic of hematopoietic stem cells, while their progeny have dramatic proliferative ability and inexorably move toward terminal differentiation. The quiescence of stem cells has been conjectured to be of critical biologic importance in protecting the stem cell compartment, which we directly assessed using mice engineered to be deficient in the G1 checkpoint regulator, cyclin-dependent kinase inhibitor, p21cip1/waf1 (p21). In the absence of p21, hematopoietic stem cell proliferation and absolute number were increased under normal homeostatic conditions. Exposing the animals to cell cycle-specific myelotoxic injury resulted in premature death due to hematopoietic cell depletion. Further, self-renewal of primitive cells was impaired in serially transplanted bone marrow from p21-/- mice, leading to hematopoietic failure. Therefore, p21 is the molecular switch governing the entry of stem cells into the cell cycle, and in its absence, increased cell cycling leads to stem cell exhaustion. Under conditions of stress, restricted cell cycling is crucial to prevent premature stem cell depletion and hematopoietic death. PMID- 10710307 TI - Complete genome sequence of Neisseria meningitidis serogroup B strain MC58. AB - The 2,272,351-base pair genome of Neisseria meningitidis strain MC58 (serogroup B), a causative agent of meningitis and septicemia, contains 2158 predicted coding regions, 1158 (53.7%) of which were assigned a biological role. Three major islands of horizontal DNA transfer were identified; two of these contain genes encoding proteins involved in pathogenicity, and the third island contains coding sequences only for hypothetical proteins. Insights into the commensal and virulence behavior of N. meningitidis can be gleaned from the genome, in which sequences for structural proteins of the pilus are clustered and several coding regions unique to serogroup B capsular polysaccharide synthesis can be identified. Finally, N. meningitidis contains more genes that undergo phase variation than any pathogen studied to date, a mechanism that controls their expression and contributes to the evasion of the host immune system. PMID- 10710308 TI - Identification of vaccine candidates against serogroup B meningococcus by whole genome sequencing. AB - Neisseria meningitidis is a major cause of bacterial septicemia and meningitis. Sequence variation of surface-exposed proteins and cross-reactivity of the serogroup B capsular polysaccharide with human tissues have hampered efforts to develop a successful vaccine. To overcome these obstacles, the entire genome sequence of a virulent serogroup B strain (MC58) was used to identify vaccine candidates. A total of 350 candidate antigens were expressed in Escherichia coli, purified, and used to immunize mice. The sera allowed the identification of proteins that are surface exposed, that are conserved in sequence across a range of strains, and that induce a bactericidal antibody response, a property known to correlate with vaccine efficacy in humans. PMID- 10710309 TI - gridlock, an HLH gene required for assembly of the aorta in zebrafish. AB - The first artery and vein of the vertebrate embryo assemble in the trunk by migration and coalescence of angioblasts to form endothelial tubes. The gridlock (grl) mutation in zebrafish selectively perturbs assembly of the artery (the aorta). Here it is shown that grl encodes a basic helix-loop-helix (bHLH) protein belonging to the Hairy/Enhancer of the split family of bHLH proteins. The grl gene is expressed in lateral plate mesoderm before vessel formation, and thereafter in the aorta and not in the vein. These results suggest that the arterial endothelial identity is established even before the onset of blood flow and implicate the grl gene in assignment of vessel-specific cell fate. PMID- 10710310 TI - DNA damage-induced activation of p53 by the checkpoint kinase Chk2. AB - Chk2 is a protein kinase that is activated in response to DNA damage and may regulate cell cycle arrest. We generated Chk2-deficient mouse cells by gene targeting. Chk2-/- embryonic stem cells failed to maintain gamma-irradiation induced arrest in the G2 phase of the cell cycle. Chk2-/- thymocytes were resistant to DNA damage-induced apoptosis. Chk2-/- cells were defective for p53 stabilization and for induction of p53-dependent transcripts such as p21 in response to gamma irradiation. Reintroduction of the Chk2 gene restored p53 dependent transcription in response to gamma irradiation. Chk2 directly phosphorylated p53 on serine 20, which is known to interfere with Mdm2 binding. This provides a mechanism for increased stability of p53 by prevention of ubiquitination in response to DNA damage. PMID- 10710311 TI - Identification of a cellular cofactor required for infection by feline leukemia virus. AB - Retroviral infection involves continued genetic variation, leading to phenotypic and immunological selection for more fit virus variants in the host. For retroviruses that cause immunodeficiency, pathogenesis is linked to the emergence of T cell-tropic, cytopathic viruses. Here we show that an immunodeficiency inducing, T cell-tropic feline leukemia virus (FeLV) has evolved such that it cannot infect cells unless both a classic multiple membrane-spanning receptor molecule (Pit1) and a second coreceptor or entry factor are present. This second receptor component, which we call FeLIX, was identified as an endogenously expressed protein that is similar to a portion of the FeLV envelope protein. This cellular protein can function either as a transmembrane protein or as a soluble component to facilitate infection. PMID- 10710312 TI - Candidate taste receptors in Drosophila. AB - Little is known about the molecular mechanisms of taste perception in animals, particularly the initial events of taste signaling. A large and diverse family of seven transmembrane domain proteins was identified from the Drosophila genome database with a computer algorithm that identifies proteins on the basis of structure. Eighteen of 19 genes examined were expressed in the Drosophila labellum, a gustatory organ of the proboscis. Expression was not detected in a variety of other tissues. The genes were not expressed in the labellum of a Drosophila mutant, pox-neuro70, in which taste neurons are eliminated. Tissue specificity of expression of these genes, along with their structural similarity, supports the possibility that the family encodes a large and divergent family of taste receptors. PMID- 10710313 TI - Correlates of sleep and waking in Drosophila melanogaster. AB - Drosophila exhibits a circadian rest-activity cycle, but it is not known whether fly rest constitutes sleep or is mere inactivity. It is shown here that, like mammalian sleep, rest in Drosophila is characterized by an increased arousal threshold and is homeostatically regulated independently of the circadian clock. As in mammals, rest is abundant in young flies, is reduced in older flies, and is modulated by stimulants and hypnotics. Several molecular markers modulated by sleep and waking in mammals are modulated by rest and activity in Drosophila, including cytochrome oxidase C, the endoplasmic reticulum chaperone protein BiP, and enzymes implicated in the catabolism of monoamines. Flies lacking one such enzyme, arylalkylamine N-acetyltransferase, show increased rest after rest deprivation. These results implicate the catabolism of monoamines in the regulation of sleep and waking in the fly and suggest that Drosophila may serve as a model system for the genetic dissection of sleep. PMID- 10710314 TI - Genetic suppression of polyglutamine toxicity in Drosophila. AB - A Drosophila model for Huntington's and other polyglutamine diseases was used to screen for genetic factors modifying the degeneration caused by expression of polyglutamine in the eye. Among 7000 P-element insertions, several suppressor strains were isolated, two of which led to the discovery of the suppressor genes described here. The predicted product of one, dHDJ1, is homologous to human heat shock protein 40/HDJ1. That of the second, dTPR2, is homologous to the human tetratricopeptide repeat protein 2. Each of these molecules contains a chaperone related J domain. Their suppression of polyglutamine toxicity was verified in transgenic flies. PMID- 10710334 TI - Beat-to-beat repolarization variability in ventricular myocytes and its suppression by electrical coupling. AB - Single ventricular myocytes paced at a constant rate and held at a constant temperature exhibit beat-to-beat variations in action potential duration (APD). In this study we sought to quantify this variability, assess its mechanism, and determine its responsiveness to electrotonic interactions with another myocyte. Interbeat APD(90) (90% repolarization) of single cells was normally distributed. We thus quantified APD(90) variability as the coefficient of variability, CV = (SD/mean APD(90)) x 100. The mean +/- SD of the CV in normal solution was 2.3 +/- 0.9 (132 cells). Extracellular TTX (13 microM) and intracellular EGTA (14 mM) both significantly reduced the CV by 44 and 26%, respectively. When applied in combination the CV fell by 54%. In contrast, inhibition of the rapid delayed rectifier current with L-691,121 (100 nM) increased the CV by 300%. The CV was also significantly reduced by 35% when two normal myocytes were electrically connected with a junctional resistance (R(j)) of 100 MOmega. Electrical coupling (R(j) = 100 MOmega) of a normal myocyte to one producing early afterdepolarization (EAD) completely blocked EAD formation. These results indicate that beat-to-beat APD variability is likely mediated by stochastic behavior of ion channels and that electrotonic interactions act to limit temporal dispersion of refractoriness, a major contributor to arrhythmogenesis. PMID- 10710335 TI - Nonlinear changes of transmembrane potential caused by defibrillation shocks in strands of cultured myocytes. AB - Organization of cardiac tissue into cell strands and layers has been implicated in changes of transmembrane potential (DeltaV(m)) during defibrillation. To determine the shock-induced DeltaV(m) in such structures, cell strands of variable width [strand width (SW) = 0.15-2 mm] were grown in culture. Uniform field shocks with variable strength [shock strength (SS) = 2-50 V/cm] were applied across strands during the action potential (AP) plateau, and DeltaV(m) were measured optically. Three different types of DeltaV(m) were observed. Small DeltaV(m) [<40%AP amplitude (APA)] were linearly dependent on SS and SW and were symmetrically distributed about a strand centerline with maximal positive and negative DeltaV(m) on opposite strand sides being equal. Intermediate DeltaV(m) (<200%APA) were strongly asymmetric with negative DeltaV(m) > positive DeltaV(m) because of a negative time-dependent shift of V(m) at the depolarized side of the strands. For large DeltaV(m) (>200%APA), a second time-dependent shift of V(m) to more positive levels was observed in the hyperpolarized portions of strands, causing reduction of the DeltaV(m) asymmetry. We conclude that during application of shocks to cell strands during the AP plateau, passive changes of V(m) were followed by two voltage- and time-dependent shifts of V(m), possibly reflecting changes of ionic currents or membrane electroporation. PMID- 10710336 TI - Altered LV inotropic reserve and mechanoenergetics early in the development of heart failure. AB - To test the hypothesis that alterations in left ventricular (LV) mechanoenergetics and the LV inotropic response to afterload manifest early in the evolution of heart failure, we examined six anesthetized dogs instrumented with LV micromanometers, piezoelectric crystals, and coronary sinus catheters before and after 24 h of rapid ventricular pacing (RVP). After autonomic blockade, the end-systolic pressure-volume relation (ESPVR), myocardial O(2) consumption (MVO(2)), and LV pressure-volume area (PVA) were defined at several different afterloads produced by graded infusions of phenylephrine. Short-term RVP resulted in reduced preload with proportionate reductions in stroke work and the maximum first derivative of LV pressure but with no significant reduction in baseline LV contractile state. In response to increased afterload, the baseline ESPVR shifted to the left with maintained end-systolic elastance (E(es)). In contrast, after short-term RVP, in response to comparable increases in afterload, the ESPVR displayed reduced E(es) (P < 0.05) and significantly less leftward shift compared with control (P < 0.05). Compared with the control MVO(2)-PVA relation, short-term RVP significantly increased the MVO(2) intercept (P < 0.05) with no change in slope. These results indicate that short-term RVP produces attenuation of afterload-induced enhancement of LV performance and increases energy consumption for nonmechanical processes with maintenance of contractile efficiency, suggesting that early in the development of tachycardia heart failure, there is blunting of length-dependent activation and increased O(2) requirements for excitation-contraction coupling, basal metabolism, or both. Rather than being adaptive mechanisms, these abnormalities may be primary defects involved in the progression of the heart failure phenotype. PMID- 10710337 TI - Alterations in relaxation to lactate and H(2)O(2) in human placental vessels from gestational diabetic pregnancies. AB - We determined whether alterations in the mechanism of relaxation to H(2)O(2) potentially contribute to the enhanced prostaglandin-mediated contractile response to H(2)O(2) and posthypoxic reoxygenation seen in human placental vessels of pregnancies with gestational diabetes mellitus (GDM). Isolated placental arteries and veins from GDM and uncomplicated full-term pregnancies were precontracted with prostaglandin F(2alpha) (PO(2) 35-38 Torr) and then exposed to lactate (1-10 mM), arachidonic acid (0.01-10 microM), nitroglycerin (1 nM-1 microM), forskolin (0.01-10 microM), or H(2)O(2) (1 microM-1 mM + 10 microM indomethacin). The rates of tissue H(2)O(2) metabolism by catalase and nitrite production were measured. The relaxation to lactate was reduced in GDM placental arteries and veins by 54-85 and 66-80%, and the relaxation to H(2)O(2) was inhibited by 80-94% in GDM placental veins compared with vessels from uncomplicated full-term pregnancies. H(2)O(2) caused only minimal relaxation of placental arteries. Responses to other relaxing agents were not altered in the GDM placental vessels. Diabetic vessels showed rates of nitrite production that were increased by 113-195% and rates of H(2)O(2) metabolism by catalase that were decreased by 44-61%. The loss of relaxation to H(2)O(2) and lactate (mediated via H(2)O(2)), perhaps as a result of the inhibition of catalase by nitric oxide, may explain the previously reported enhancement of prostaglandin-mediated contractile responses to H(2)O(2) and posthypoxic reoxygenation seen in GDM placental vessels. PMID- 10710338 TI - Altered L-type Ca(2+) channel currents in vascular smooth muscle cells from experimental diabetic rats. AB - Vascular complications of diabetes are associated with abnormal Ca(2+) handling by vascular smooth muscle cells (SMCs) in which the alteration in L-type voltage dependent Ca(2+) channel (VDCC) currents may play an important role. In the present study, the characteristics of L-type VDCC currents in tail artery SMCs from streptozotocin-induced diabetic rats were examined. The densities, but not the voltage dependence, of L-type VDCC currents were reduced as diabetes progressed from 1 wk to 3 mo. The inhibitory effect of dibutyryl-cAMP on L-type VDCC currents was greater in diabetic SMCs than in age-matched control cells (P < 0.01). Both the stimulatory effect of BAY K 8644 and the inhibitory effect of nifedipine on L-type VDCC currents were significantly enhanced in diabetic cells. The diabetes-related abnormalities in L-type VDCC currents were mimicked by culturing SMCs with a high concentration of glucose. Our results suggest that the properties of L-type VDCC in diabetic vascular SMCs were significantly altered, partially related to the increased L-type VDCC sensitivity to cAMP and hyperglycemia. PMID- 10710339 TI - Single-channel activity and expression of atrial L-type Ca(2+) channels in patients with latent hyperthyroidism. AB - Patients with "latent hyperthyroidism" (suppressed thyroid-stimulating hormone and normal circulating thyroid hormones) are at risk to develop atrial fibrillation. In animal models, hyperthyroidism is associated with increased cardiac L-type Ca(2+) current. Therefore, we assessed L-type channel function and expression in right atria from patients undergoing cardiac surgery. Single L-type channels were studied in the cell-attached condition. Voltage dependence of gating was similar in patients with and without latent hyperthyroidism. With use of a pulse protocol leading to maximum channel availability, single-channel activity was further analyzed. Average peak current was significantly enhanced in latent hyperthyroidism, mainly because of an increased channel availability (P < 0.05). Protein expression was analyzed by Western blot. In latent hyperthyroidism, expression of Ca(2+) channel alpha(1)-subunits was increased more than threefold (P < 0.01). In contrast, sarco(endo)plasmic reticulum Ca(2+) ATPase and phospholamban levels were not significantly changed. We only observed a trend toward increased sarco(endo)plasmic reticulum Ca(2+)-ATPase expression (P = 0.085). Function and expression of human atrial L-type Ca(2+) channels are increased in latent hyperthyroidism. These endocrine effects on the heart may be clinically relevant. PMID- 10710340 TI - Neurohormones in an ovine model of compensated postinfarction left ventricular dysfunction. AB - Clinical heart failure, often the result of myocardial infarction, may be preceded by a period of compensated left ventricular impairment. There is substantial need for an experimental model that reflects this human condition. In sheep, coronary artery ligation produced consistent left ventricular anteroapical myocardial infarctions resulting in chronic (5 wk), stable hemodynamic changes compared with sham controls, including reductions in ejection fraction (51 +/- 2 vs. 30 +/- 5%, P < 0.001), cardiac output (6.3 +/- 0.2 vs. 5.1 +/- 0.2 l/min, P < 0.01), and arterial pressure (93 +/- 2 vs. 79 +/- 3 mmHg, P < 0.001), and increases in cardiac preload (left atrial pressure, 3.3 +/- 0.1 vs. 8.3 +/- 1.3 mmHg, P < 0.001). These changes were associated with acute and sustained increases in plasma concentrations of atrial natriuretic peptide (ANP; 5 wk, 11 +/- 2 vs. 27 +/- 5 pmol/l, P < 0.001), brain natriuretic peptide (BNP; 3 +/- 0.2 vs. 11 +/- 2 pmol/l, P < 0.001), and amino-terminal pro-brain natriuretic peptide (NT-BNP; 17 +/- 3 vs. 42 +/- 12 pmol/l, P < 0.001). Significant correlations were observed between plasma levels of the natriuretic peptides (ANP, day 7 to week 5 samples; BNP and NT-BNP, day 1 to week 5 samples) and changes in left ventricular volumes and ejection fraction. In contrast, renin activity, aldosterone, catecholamines, and endothelin were not chronically elevated postinfarction and were not related to indexes of ventricular function. Coronary artery ligation in sheep produces the pathological, hemodynamic, and neurohormonal characteristics of compensated left ventricular impairment secondary to myocardial infarction. Plasma concentrations of the cardiac natriuretic peptides are sensitive markers of left ventricular dysfunction. This is a reproducible model that reflects the clinical condition and should prove suitable for investigating the pathophysiology of, and experimental therapies in, early left ventricular dysfunction. PMID- 10710341 TI - Comparison of local and systemic effects of insulin on myocardial glucose extraction in ischemic heart disease. AB - Physiological increases in circulating insulin level significantly increase myocardial glucose uptake in vivo. To what extent this represents a direct insulin action on the heart or results indirectly from reduction in circulating concentrations of free fatty acids (FFA) is uncertain. To examine this, we measured myocardial glucose, lactate, and FFA extraction in 10 fasting men (ages 49-76 yr) with stable coronary artery disease during sequential intracoronary (10 mU/min, coronary plasma insulin = 140 +/- 20 microU/ml) and intravenous (100 mU/min, systemic plasma insulin = 168 +/- 26 microU/ml) insulin infusion. Basally, hearts extracted 2 +/- 2% of arterial glucose and extracted 27 +/- 6% of FFA. Coronary insulin infusion increased glucose extraction to 5 +/- 3% (P < 0.01 vs. basal) without changing plasma FFA or heart FFA extraction. Conversion to intravenous infusion lowered plasma FFA by approximately 50% and heart FFA extraction by approximately 75%, increasing heart glucose extraction still further to 8 +/- 3% (P < 0. 01 vs. intracoronary). This suggests the increase in myocardial glucose extraction observed in response to an increment in systemic insulin concentration is mediated equally by a reduction in circulating FFA and by direct insulin action on the heart itself. Coronary insulin infusion increased myocardial lactate extraction as well (from 20 +/- 10% to 29 +/- 9%, P < 0.05), suggesting the local action may include stimulation of a metabolic step distal to glucose transport and glycolysis. PMID- 10710342 TI - Physiological cyclic stretch causes cell cycle arrest in cultured vascular smooth muscle cells. AB - Smooth muscle cells (SMC) are the major cellular component of the blood vessel wall and are continuously exposed to cyclic stretch due to pulsatile blood flow. This study examined the effects of a physiologically relevant level of cyclic stretch on rat aortic vascular SMC proliferation. Treatment of static SMC with serum, platelet-derived growth factor, or thrombin stimulated SMC proliferation, whereas exposure of SMC to cyclic stretch blocked the proliferative effect of these growth factors. The stretch-mediated inhibition in SMC growth was not due to cell detachment or increased cell death. Flow cytometry analysis revealed that cyclic stretch increased the fraction of SMC in the G(0)/G(1) phase of the cell cycle. Stretch-inhibited G(1)/S phase transition was associated with a decrease in retinoblastoma protein phosphorylation and with a selective increase in the cyclin-dependent kinase inhibitor p21, but not p27. These results demonstrate that cyclic stretch inhibits SMC growth by blocking cell cycle progression and suggest that physiological levels of cyclic stretch contribute to vascular homeostasis by inhibiting the proliferative pathway of SMC. PMID- 10710343 TI - Protective effects of leukopenia and tissue plasminogen activator in microvascular ischemia-reperfusion injury. AB - Ischemia shifts the anticoaugulant/procoagulant balance of the endothelium in favor of activation of coagulation. We studied whether cheek pouch microcirculation of leukopenic hamsters was protected by tissue plasminogen activator (tPA) (50 microg/100 g body wt) against ischemia-reperfusion injury. Adherent leukocytes, total perfused capillary length (PCL), permeability increase, and arteriolar and venular red blood cell (RBC) velocity were investigated by fluorescence microscopy. Measurements were made at control, 30 or 60 min of ischemia, and at 30 or 60 min of reperfusion. Hamsters were made leukopenic by treatment with cyclophosphamide (20 mg/100 g body wt ip, 4 days before the experiment), which decreased circulating leukocyte count by 85-90%. Leukopenic hamsters undergoing 30 min of ischemia followed by 30 min of reperfusion showed no significant decrease in PCL or increased permeability. Leukopenic hamsters undergoing 60 min of ischemia followed by 60 min of reperfusion presented a significant decrease in microvascular perfusion where PCL was 28 +/- 7% of baseline, low-flow conditions, and increased permeability. In leukopenic hamsters treated with tPA there was complete protection of capillary perfusion with no significant changes in permeability or arteriolar and venular RBC velocity. In conclusion, thrombus formation may be an additional and independent factor that with leukocyte-mediated mechanisms determines ischemia reperfusion injury. PMID- 10710344 TI - In eNOS knockout mice skeletal muscle arteriolar dilation to acetylcholine is mediated by EDHF. AB - The mechanisms that account for acetylcholine (ACh)-induced responses of skeletal muscle arterioles of mice lacking endothelial nitric oxide (NO) synthase (eNOS KO) were investigated. Isolated, cannulated, and pressurized arterioles of gracilis muscle from male eNOS-KO (74.1 +/- 2.3 microm) and wild-type (WT, 87.2 +/- 2.1 microm) mice developed spontaneous tone accounting for 63 and 61% of their passive diameter (116.8 +/- 3.4 vs. 143.2 +/- 2.8 microm, respectively) and dilated dose-dependently to ACh (10(-9)-10(-7) M). These dilations were significantly smaller in vessels of eNOS-KO compared with WT mice (29.2 +/- 2.0 microm vs. 46.3 +/- 2.1 microm, at maximum concentration) but responses to the NO donor, sodium nitrite (NaNO(2), 10(-6)-3 x 10(-5) M), were comparable in the vessels of the two strains. N(G)-nitro-L-arginine (L-NNA, 10(-4) M), an inhibitor of eNOS, inhibited ACh-induced dilations by 60-90% in arterioles of WT mice but did not affect responses in those of eNOS-KO mice. In arterioles of eNOS-KO mice, dilations to ACh were not affected by indomethacin but were essentially abolished by inhibitors of cytochrome P-450, clotrimazole (CTZ, 2 x 10(-6) M) or miconazole (MCZ, 2 x 10(-6) M), as well as by either high K(+) (40 mM) or iberiotoxin [10( 7) M, a blocker of Ca(2+)-dependent K(+) channels (K(Ca) channels)]. On the other hand, in WT arterioles CTZ or MCZ inhibited ACh-induced dilations only by approximately 10% and only in the presence of L-NNA. These results indicate that in arterioles of eNOS-KO mice, endothelium-derived hyperpolarizing factor (EDHF), synthesized via cytochrome P-450, accounts entirely for the mediation of ACh induced dilation via an increase in K(Ca)-channel activity. In contrast, in arterioles of WT mice, endothelium-derived NO predominantly mediates ACh-induced dilation in which participation of EDHF becomes apparent only after inhibition of NO synthesis. PMID- 10710345 TI - Phosphorylation of phospholamban and troponin I in beta-adrenergic-induced acceleration of cardiac relaxation. AB - Activation of cAMP-dependent protein kinase A (PKA) in ventricular myocytes by isoproterenol (Iso) causes phosphorylation of both phospholamban (PLB) and troponin I (TnI) and accelerates relaxation by up to twofold. Because PLB phosphorylation increases sarcoplasmic reticulum (SR) Ca pumping and TnI phosphorylation increases the rate of Ca dissociation from the myofilaments, both factors could contribute to the acceleration of relaxation seen with PKA activation. To compare quantitatively the role of TnI versus PLB phosphorylation, we measured relaxation rates before and after maximal Iso treatment for twitches of matched amplitudes in ventricular myocytes and muscle from wild-type (WT) mice and from mice in which the PLB gene was knocked out (PLB-KO). Because Iso increases contractions, even in the PLB-KO mouse, extracellular [Ca] or sarcomere length was adjusted to obtain matching twitch amplitudes (in the presence and absence of Iso). In PLB-KO myocytes and muscles (which were allowed to shorten), Iso did not alter the time constant (tau) of relaxation ( approximately 29 ms). However, with increasing isometric force development in the PLB-KO muscles, Iso progressively but modestly accelerated relaxation (by 17%). These results contrast with WT myocytes and muscles where Iso greatly reduced tau of cell relaxation and intracellular Ca concentration decline (by 30-50%), independent of mechanical load. The Iso treatment used produced comparable increases in phosphorylation of TnI and PLB in WT. We conclude that the effect of beta adrenergic activation on relaxation is mediated entirely by PLB phosphorylation in the absence of external load. However, TnI phosphorylation could contribute up to 14-18% of this lusitropic effect in the WT mouse during maximal isometric contractions. PMID- 10710346 TI - Developmental changes in endothelium-derived vasorelaxant factors in cerebral circulation. AB - Endothelium-derived prostanoids are predominant vasorelaxant factors in the cerebral circulation of newborn pigs in vivo, whereas in older pigs nitric oxide (NO)-mediated responses also contribute to the regulation of cerebral vascular tone. We compared the expression and activities of NO synthase and cyclooxygenase in the cerebral microcirculation of newborn and adult pigs. In adult animals, expression and activity of endothelial NO synthase in cerebral microvessels and in cultured cerebral endothelial cells is two- to threefold higher than in newborn pigs; acetylcholine and bradykinin cause a greater increase in NO production in adult pigs. Expression and activity of cyclooxygenase in cerebral microvascular endothelial cells is similar in newborn and adult pigs; acetylcholine and bradykinin stimulated dilator prostanoid production to the same degree in both age groups. Endothelial prostanoid synthesis in cerebral microvessels and cultured endothelial cells was inhibited 30-70% by NS-398, reflecting a large contribution of COX-2 in both newborn and adult animals. These data indicate that in the cerebral circulation of pigs, NO synthase is age dependently upregulated, whereas endothelial cyclooxygenase is not altered during postnatal development. PMID- 10710347 TI - Age-related changes in A(1)-adenosine receptor-mediated bradycardia. AB - The impact of age on functional sensitivity to A(1)-adenosine receptor activation was studied in Langendorff-perfused hearts from young (1-2 mo) and old (12-18 mo) male Wistar rats. Adenosine mediated bradycardia in young and old hearts, with sensitivity enhanced approximately 10-fold in old [negative logarithm of EC(50) (pEC(50)) = 4.56 +/- 0.11] versus young hearts (pEC(50) = 3.70 +/- 0. 09). Alternatively, the nonmetabolized A(1) agonists N(6)-cyclohexyladenosine and (R) N(6)-phenylisopropyladenosine were equipotent in young (pEC(50) = 7.43 +/- 0.12 and 6.61 +/- 0.19, respectively) and old hearts (pEC(50) = 7.07 +/- 0.10 and 6.80 +/- 0. 11, respectively), suggesting a role for uptake and/or catabolism in age related changes in adenosine sensitivity. In support of this suggestion, [(3)H] adenosine uptake was approximately twofold greater in young than in old hearts (from 3-100 microM adenosine). However, although inhibition of adenosine deaminase and adenosine transport with 10 microM erythro-9-(2-hydroxy-3 nonyl)adenine hydrochloride and 10 microM S-(4-nitrobenzyl)-6-thioinosine increased adenosine sensitivity three- to fourfold, it failed to abolish the sensitivity difference in old (pEC(50) = 4.95 +/- 0.08) versus young (pEC(50) = 4.29 +/- 0.13) hearts. Data indicate that 1) age increases functional A(1) receptor sensitivity to adenosine without altering the sensitivity of the A(1) receptor itself, and 2) age impairs adenosine transport and/or catabolism, but this does not explain differing functional sensitivity to adenosine. This increased functional sensitivity to adenosine may have physiological significance in the older heart. PMID- 10710348 TI - Altered molecular response to adrenoreceptor-induced cardiac hypertrophy in Egr-1 deficient mice. AB - Unmanipulated early growth response-1 (Egr-1)-deficient -/- mice have similar heart-to-body weight ratios but express lower amounts of atrial natriuretic factor (ANF), beta-myosin heavy chain (beta-MHC), skeletal actin, NGF1-A binding protein (NAB)-2, Sp1, c-fos, c-jun, GATA-4, and Nkx2.5 than +/+ or +/- mice. alpha-MHC, tubulin, and NAB-1 expression was similar. Isoproterenol (Iso) and phenylephrine (PE) infusion into +/+ and -/- mice increased heart weight, ANF, beta-MHC, skeletal actin, Sp1, NAB-2, c-fos, and c-jun expression, but induction in -/- mice was lower. Only Iso + PE-treated +/+ mice showed induction of NAB-1, GATA-4, and Nkx2.5. Foci of fibrosis were found in Iso + PE-treated -/- and +/+ mice. Surprisingly, vehicle-treated -/- mice displayed fibrosis and increased Sp1, skeletal actin, Nkx2.5, and GATA-4 expression without hypertrophy. Minipump removal caused the agonist-treated hearts and gene expression to regress to control or near-control levels. Thus Egr-1 deficiency caused a blunted catecholamine-induced hypertrophy response and increased sensitivity to stress. PMID- 10710349 TI - Characterization and functional consequences of delayed rectifier current transient in ventricular repolarization. AB - Although inactivation of the rapidly activating delayed rectifier current (I(Kr)) limits outward current on depolarization, the role of I(Kr) (and recovery from inactivation) during repolarization is uncertain. To characterize I(Kr) during ventricular repolarization (and compare with the inward rectifier current, I(K1)), voltage-clamp waveforms simulating the action potential were applied to canine ventricular, atrial, and Purkinje myocytes. In ventricular myocytes, I(Kr) was minimal at plateau potentials but transiently increased during repolarizing ramps. The I(Kr) transient was unaffected by repolarization rate and maximal after 150-ms depolarizations (+25 mV). Action potential clamps revealed the I(Kr) transient terminating the plateau. Although peak I(Kr) transient density was relatively uniform among myocytes, potentials characterizing the peak transients were widely dispersed. In contrast, peak inward rectifier current (I(K1)) density during repolarization was dispersed, whereas potentials characterizing I(K1) defined a narrower (more negative) voltage range. In summary, rapidly activating I(Kr) provides a delayed voltage-dependent (and functionally time-independent) outward transient during ventricular repolarization, consistent with rapid recovery from inactivation. The heterogeneous voltage dependence of I(Kr) provides a novel means for modulating the contribution of this current during repolarization. PMID- 10710350 TI - Heart failure alters the strength and mechanisms of the muscle metaboreflex. AB - We hypothesized that excessive sympathoactivation observed during strenuous exercise in subjects with heart failure (HF) may result from tonic activation of the muscle metaboreflex (MMR) via hypoperfusion of active skeletal muscle. We studied MMR responses in dogs during treadmill exercise by graded reduction of terminal aortic blood flow (TAQ) before and after induction of HF by rapid ventricular pacing. At a low workload, in both control and HF experiments, large decreases in TAQ were required to elicit the MMR pressor response. During control experiments, this pressor response resulted from increased cardiac output (CO), whereas in HF CO did not increase; thus the pressor response was solely due to peripheral vasoconstriction. In HF, MMR activation also induced higher plasma levels of vasopressin, norepinephrine (NE), and renin. At a higher workload, in control experiments any reduction of TAQ elicited MMR pressor responses. In HF, before any vascular occlusion, TAQ was already below MMR control threshold levels and reductions in TAQ again did not result in higher CO; thus SAP increased via peripheral vasoconstriction. NE rose markedly, indicating intense sympathetic activation. We conclude that in HF, the MMR is likely tonically active at moderate workloads and contributes to the tonic sympathoactivation. PMID- 10710351 TI - Meta-analysis of the age-associated decline in maximal aerobic capacity in men: relation to training status. AB - Based on cross-sectional data, we recently reported that, in contrast to the prevailing view, the rate of decline in maximal oxygen consumption (VO(2 max)) with age is greater in physically active compared with sedentary healthy women. We tested this hypothesis in men using a meta-analytic study of VO(2 max) values in the published literature. A total of 242 studies (538 subject groups and 13,828 subjects) met the inclusion criteria and were arbitrarily separated into sedentary (214 groups, 6,231 subjects), active (159 groups, 5,621 subjects), and endurance-trained (165 groups, 1,976 subjects) populations. Body fat percent increased with age in sedentary and active men (P < 0.001), whereas no change was observed in endurance-trained men. VO(2 max) was inversely and strongly related to age within each population (r = -0.80 to -0.88, all P < 0. 001) and was highest in endurance-trained and lowest in sedentary populations at any age. Absolute rates of decline in VO(2 max) with age were not different (P > 0.05) in sedentary (-4.0 ml. kg(-1). min(-1). decade(-1)), active (-4.0), and endurance trained (-4.6) populations. Similarly, there were no group differences (P > 0.05) in the relative (%) rates of decline in VO(2 max) with advancing age (-8.7, -7.3, and -6.8%/decade, respectively). Maximal heart rate was inversely related to age within each population (r = -0.88 to -0.93, all P < 0.001), but the rate of age related reduction was not different among the populations. There was a significant decline in running mileage and speed with advancing age in the endurance-trained men. The present cross-sectional meta-analytic findings do not support the hypothesis that the rate of decline in VO(2 max) with age is related to habitual aerobic exercise status in men. PMID- 10710352 TI - Hydrogen peroxide induces LFA-1-dependent neutrophil adherence to cardiac myocytes. AB - Adult cardiac myocytes express intercellular adhesion molecule (ICAM)-1 in response to cytokine stimulation. This allows stable adhesion of chemotactically stimulated but not unstimulated neutrophils. In the current study, we demonstrated that brief exposure of ICAM-1-expressing cardiac myocytes to H(2)O(2) promoted transient adhesive interactions between myocytes and neutrophils without added chemotactic factors. This transient adhesion differed in two ways from the stable adhesion promoted by exogenous chemotactic factors. It occurred more rapidly, peaking within 15 min, and it was dependent on leukocyte function-associated antigen (LFA)-1 (CD11a/CD18) on the neutrophil interacting with ICAM-1 on the myocyte. In contrast, chemotactic factor-induced adhesion peaked at 60 min and was dependent on Mac-1 (CD11b/CD18). The transient adhesion could be completely inhibited by platelet-activating factor (PAF) receptor antagonists WEB-2086 and SDZ-64-412. These results indicate that canine neutrophils may utilize both LFA-1 and Mac-1 to adhere to adult cardiac myocytes, with LFA-1 triggered by a PAF-like activity induced in myocytes by H(2)O(2). PMID- 10710353 TI - Role of cAMP in activation of ischemically sensitive abdominal visceral afferents. AB - A number of metabolites produced during abdominal ischemia can stimulate and/or sensitize visceral afferents. The precise mechanisms whereby these metabolites act are uncertain. Other studies have shown that the adenylate cyclase-cAMP system may be involved in the activation of sensory neurons. Therefore, we hypothesized that cAMP contributes to the activation of ischemically sensitive abdominal visceral afferents. Single-unit activity of abdominal visceral C fibers was recorded from the right thoracic sympathetic chain in anesthetized cats before and during 7 min of abdominal ischemia. Forty-six percent of ischemically sensitive C fibers responded to intra-arterial injection of 8-bromo-cAMP (0.35-1. 0 mg/kg), an analog of cAMP, with responses during ischemia increasing from 0.50 +/- 0.06 to 0.84 +/- 0.08 impulses/s (P < 0.05, n = 11 C fibers). Conversely, an inhibitor of adenylate cyclase, 2', 5'-dideoxyadenosine (DDA; 0.1 mg/kg iv), attenuated ischemia-induced increase in activity of afferents from 0.66 +/- 0.10 to 0.34 +/- 0. 09 impulses/s (P < 0.05; n = 8). Furthermore, whereas exogenous PGE(2) (3-4 microg/kg ia) augmented the ischemia-induced increase in activity of afferents (P < 0.05, n = 10), treatment with DDA (0.1 mg/kg iv) substantially reduced the increase in discharge activity of afferents during ischemia, which was augmented by PGE(2) (1.45 +/- 0.24 vs. 0.70 +/- 0.09 impulses/s, -DDA vs. +DDA; P < 0.05) in six fibers. A time control group (n = 4), however, demonstrated similar increases in the activity of afferents with repeated administration of PGE(2). These data suggest that cAMP contributes to the activation of abdominal visceral afferents during ischemia, particularly to the action of PGs on activation and/or sensitization of these endings. PMID- 10710354 TI - LPS tolerance in human endothelial cells: reduced PMN adhesion, E-selectin expression, and NF-kappaB mobilization. AB - Cytokine release from inflammatory (CD14(+)) cells is reduced after repeated stimulation with lipopolysaccharide (LPS; LPS tolerance). However, it is not known whether LPS tolerance can be induced in CD14(-) cells. The aim of the present study was to determine whether endothelial cells [human umbilical vein endothelial cells (HUVEC)] could be rendered tolerant to LPS with respect to LPS induced polymorphonuclear neutrophil (PMN) adhesion. LPS stimulation (0.5 microg/ml; 4 h) of naive HUVEC increased PMN adhesion. Pretreatment of HUVEC with LPS (0.5 microg/ml) for 24 h resulted in a reduction in the proadhesive effects of a subsequent LPS challenge. The initial LPS stimulation increased 1) mobilization of the nuclear transcription factor NF-kappaB to the nucleus and 2) surface levels of the adhesion molecules intercellular adhesion molecule-1 (ICAM 1) and E-selectin. In LPS-tolerant HUVEC, a second LPS challenge resulted in 1) less accumulation of NF-kappaB in the nucleus, 2) a reduction in E-selectin expression, and 3) unchanged ICAM-1 expression. LPS-tolerant cells were still capable of mobilizing NF-kappaB in response to stimulation with either interleukin-1beta or tumor necrosis factor-alpha, resulting in elevated E selectin levels and increased PMN adhesion. These studies show for the first time that LPS tolerance can be induced in endothelial cells with respect to PMN adhesion. This tolerance is specific for LPS and is associated with an inability of LPS to mobilize NF-kappaB, resulting in less E-selectin expression. PMID- 10710355 TI - DITPA prevents the blunted contraction-frequency relationship in myocytes from infarcted hearts. AB - Loss of the positive force-frequency relationship is a characteristic finding in failing hearts. The mechanisms of this change are not well understood. Myocardial infarction (MI) was induced in rabbits to produce left ventricular (LV) dysfunction. Beginning 1 day after MI, a subgroup of rabbits received diiodothyropropionic acid (DITPA) (3.75 mg x kg(-1) x day(-1) sc) for 3 wk. We measured contractions, Ca(2+) transients, action potentials, and sarcoplasmic reticulum (SR) Ca(2+) content at different stimulation rates in single LV myocytes. The shortening-frequency relationship was markedly flattened in MI myocytes compared with control myocytes. In addition, Ca(2+) transients, action potentials, and contractions were prolonged. Myocytes from DITPA-treated MI rabbits had preserved inotropic responses to increased stimulation rate and normal duration of action potentials and Ca(2+) transients. SR Ca(2+) content increased significantly when stimulation rate was increased from 0.5 to 2.0 Hz in control myocytes but did not change significantly in MI myocytes. Myocytes from DITPA-treated MI rabbits had a greater frequency-dependent increase in SR Ca(2+) content compared with the untreated MI rabbits. Thus single myocytes from infarcted rabbit hearts have frequency-dependent abnormalities of contractility, Ca(2+) cycling, and action potential repolarization. The flattened contraction frequency relationship can be partially explained by an attenuation of the normal enhancement of SR Ca(2+) content that occurs when stimulation rate is increased. Chronic DITPA administration after MI largely prevents the development of these abnormalities. PMID- 10710356 TI - Group III muscle afferents evoke reflex depressor responses to repetitive muscle contractions in rabbits. AB - Repetitive-twitch contraction of the hindlimb muscles in anesthetized rabbits consistently evokes a reflex depressor response, whereas this type of contraction in anesthetized cats evokes a reflex pressor response in about one-half of the preparations tested. Rapidly conducting group III fibers appear to comprise the afferent arm of the reflex arc, evoking the depressor response to twitch contraction in rabbits because electrical stimulation of their axons reflexly decreases arterial pressure. In contrast, electrical stimulation of the axons of slowly conducting group III and group IV afferents reflexly increases arterial pressure in rabbits. In the present study, we examined the discharge properties of group III and IV muscle afferents and found that the former (i.e., 13 of 20), but not the latter (i.e., 0 of 10), were stimulated by 5 min of repetitive-twitch contraction (1 Hz) of the rabbit triceps surae muscles. Moreover, most of the group III afferents responding to contraction appeared to be mechanically sensitive, discharging in synchrony with the muscle twitch. On average, rapidly conducting group III afferents responded for the 5-min duration of 1-Hz repetitive-twitch contraction, whereas slowly conducting group III afferents responded only for the first 2 min of contraction. We conclude that rapidly conducting group III afferents, which are mechanically sensitive, are primarily responsible for evoking the reflex depressor response to repetitive-twitch contractions in anesthetized rabbits. PMID- 10710357 TI - ATP-sensitive potassium channels may participate in the coupling of neuronal activity and cerebrovascular tone. AB - K(+) dilate and constrict cerebral vessels in a dose-dependent fashion. Modest elevations of abluminal K(+) cause vasodilatation, whereas larger extracellular K(+) concentration ([K(+)](out)) changes decrease cerebral blood flow. These dilations are believed to be mediated by opening of inward-rectifier potassium channels sensitive to Ba(2+). Because BaCl(2) also blocks ATP-sensitive K(+) channels (K(ATP)), we challenged K(+) dilations in penetrating, resistance-size (<60 mmu) rat neocortical vessels with the K(ATP) channel blocker glibenclamide (1 microM). Glibenclamide reduced K(+) responses from 138 +/- 8 to 110 +/- 0.8%. K(+) constrictions were not affected by glibenclamide. The Na(+)-K(+)-pump inhibitor ouabain (200 microM) did not significantly change resting vessel diameter but decreased K(+) dilations (from 153 +/- 9 to 99 +/- 2%). BaCl(2) blocked K(+) dilations with a half-maximal dissociation constant of 2.9 microM and reduced dilations to the specific K(ATP) agonist pinacidil with equal potency. We conclude that, in resistance vessels, K(+) dilations are mediated by K(ATP); we hypothesize that [K(+)](out) causes activation of Na(+)-K(+) pumps, depletion of intracellular ATP concentration, and subsequent opening of K(ATP). This latter hypothesis is supported by the blocking effect of ouabain. PMID- 10710358 TI - K(+) currents responsible for repolarization in mouse ventricle and their modulation by FK-506 and rapamycin. AB - Modulation of mouse ventricular action potentials and K(+) currents was examined using the whole cell patch-clamp technique. The composite mouse ventricular K(+) current (consisted of an outward transient followed by a slowly decaying sustained component. Use of the K(+) channel blockers tetraethylammonium and 4 aminopyridine and a transgenic mouse model revealed three pharmacologically and kinetically distinct currents: I(to), which contributed to the transient component; I(K), which contributed to the sustained component; and a slowly activating current (I(slow)), which contributed to both components. The immunosuppressant FK-506 increased action potential duration at 90% repolarization by 66.7% by decreasing the sustained component (-48% at +60 mV) and prolonging recovery from inactivation (by 26% at 200 ms) of the transient component. These effects were isolated to I(K) and I(to), respectively. Rapamycin had strikingly similar effects on these currents. Both FK-506 and rapamycin are known to target the immunophilin FKBP12. Thus we conclude that FKBP12 modulates specific mouse K(+) channels, and thus the mouse ventricular action potential, by interacting directly with K(+) channel proteins or with other associated regulatory proteins. PMID- 10710359 TI - Regional dysfunction correlates with myofiber disarray in transgenic mice with ventricular expression of ras. AB - A hallmark of certain cardiac diseases such as familial hypertrophic cardiomyopathy is focal myofiber disarray. Regional ventricular dysfunction occurs in human subjects with hypertrophic cardiomyopathy; however, no direct evidence exists to correlate regional dysfunction with myofiber disarray. We used a transgenic mouse, which exhibits regional myofiber disarray via ventricular expression of the human oncogene ras, to investigate the relationship between myofiber disarray and septal surface strain. An isolated ejecting mouse heart preparation was used to record deformation of markers on the septal surface and to determine nonhomogeneous septal surface strain maps. Myofiber disarray made in histological tissue sections was correlated with gradients in surface systolic shortening. Significantly smaller maximum principal shortening was associated with disarray located near the right ventricle (RV) septal surface. There was also significantly smaller surface shear strain associated with disarray located either near the RV surface or at the midwall. Because surface shear is a local indicator of torsion, we conclude that myofiber disarray is associated with reduced septal torsion and reduced surface shortening. PMID- 10710360 TI - Serotonin 5-HT(7) receptors mediate relaxation of porcine pial veins. AB - Isolated porcine pial veins in the presence of active muscle tone have been shown to exhibit rhythmic contractions (RC) that are inhibited by serotonin (5-HT) in a concentration-dependent manner. The 5-HT inhibition of RC is mediated by an as yet unidentified 5-HT receptor subtype located on the vascular smooth muscle. 5 carboxamidotryptamine, which is a potent but nonselective agonist at 5-HT(7) receptors, has been shown to be the most potent inhibitor of RC in porcine pial veins. Therefore, the present study was designed to determine if the 5-HT mediated inhibition of RC in pial veins is mediated by 5-HT(7) receptors and if 5 HT(7) receptor mRNA is expressed in endothelium-denuded pial veins; the study was done with the use of an in vitro tissue bath and RT-PCR techniques. Our findings indicated that 5-HT inhibition of RC in porcine pial veins was prevented by 5 HT(7)-receptor antagonists (clozapine, pimozide, and LY-215840) in a concentration-dependent manner. Furthermore, a strong PCR signal for the 5-HT(7) receptor was consistently detected in endothelium-denuded pial veins. Sequence analysis of the amplified products confirmed their high degree of homology with the porcine and/or human 5-HT(7)-receptor gene. Taken together, these data suggest that the 5-HT-induced inhibition of RC in porcine pial veins is at least in part mediated by 5-HT(7) receptors located on the venous smooth muscle. PMID- 10710361 TI - Modeling short-term interval-force relations in cardiac muscle. AB - This study employs two modeling approaches to investigate short-term interval force relations. The first approach is to develop a low-order, discrete-time model of excitation-contraction coupling to determine which parameter combinations produce the degree of postextrasystolic potentiation seen experimentally. Potentiation is found to increase 1) for low recirculation fraction, 2) for high releasable fraction, i.e., the maximum fraction of Ca(2+) released from the sarcoplasmic reticulum (SR) given full restitution, and 3) for strong negative feedback of the SR release on sarcolemmal Ca(2+) influx. The second modeling approach is to develop a more detailed single ventricular cell model that simulates action potentials, Ca(2+)-handling mechanisms, and isometric force generation by the myofilaments. A slow transition from the adapted state of the ryanodine receptor produces a gradual recovery of the SR release and restitution behavior. For potentiation, a small extrasystolic release leaves more Ca(2+) in the SR but also increases the SR loading by two mechanisms: 1) less Ca(2+)-induced inactivation of L-type channels and 2) reduction of action potential height by residual activation of the time-dependent delayed rectifier K(+) current, which increases Ca(2+) influx. The cooperativity of the myofilaments amplifies the relatively small changes in the Ca(2+) transient amplitude to produce larger changes in isometric force. These findings suggest that short-term interval-force relations result mainly from the interplay of the ryanodine receptor adaptation and the SR Ca(2+) loading, with additional contributions from membrane currents and myofilament activation. PMID- 10710362 TI - Cardiorespiratory interactions during periodic breathing in awake chronic heart failure patients. AB - We applied spectral techniques to the analysis of cardiorespiratory signals [instantaneous lung volume (ILV), instantaneous tidal volume (ITV), arterial O(2) saturation (Sa(O(2))) at the ear, heart rate (HR), systolic (SAP), and diastolic (DAP) arterial pressure] during nonapneic periodic breathing (PB) in 29 awake chronic heart failure (CHF) patients and estimated the timing relationships between respiratory and slow cardiovascular (<0.04 Hz) oscillations. Our aim was 1) to elucidate major mechanisms involved in cardiorespiratory interactions during PB and 2) to test the hypothesis of a central vasomotor origin of PB. All cardiovascular signals were characterized by a dominant (>/=84% of total power) oscillation at the frequency of PB (mean +/- SE: 0.022 +/- 0.0008 Hz), highly coherent (>/=0.89), and delayed with respect to ITV (ITV-HR, 2.4 +/- 0.72 s; ITV SAP, 6.7 +/- 0.65 s; ITV-DAP, 3.2 +/- 0.61 s; P < 0.01). Sa(O(2)) was highly coherent with (coherence function = 0.96 +/- 0. 009) and almost opposite in phase to ITV. These findings demonstrate the existence of a generalized cardiorespiratory rhythm led by the ventilatory oscillation and suggest that 1) the cyclic increase in inspiratory drive and cardiopulmonary reflexes and 2) mechanical effects of PB-induced changes in intrathoracic pressure are the more likely sources of the HR and blood pressure oscillations, respectively. The timing relationship between ITV and blood pressure signals excludes the possibility that PB represents the effect of a central vasomotor rhythm. PMID- 10710363 TI - Role of TNF-alpha in myocardial dysfunction after hemorrhagic shock and lower torso ischemia. AB - Ruptured abdominal aortic aneurysm (RAAA) repair, a combination of hemorrhagic shock and lower-torso ischemia, is associated with a 50-70% mortality. Myocardial dysfunction may contribute to the high rate of mortality after aneurysm repair. We attempted to determine whether RAAA repair results in cardiac dysfunction mediated by tumor necrosis factor-alpha (TNF-alpha). We modeled aortic rupture and repair in the rat by inducing hemorrhagic shock to a mean blood pressure of 50 mmHg for 1 h, followed by supramesenteric clamping of the aorta for 45 min. After 90 min of reperfusion, cardiac contractile function was assessed with a Langendorff preparation. Myocardial TNF-alpha, ATP and creatine phosphate (CP) levels, and markers of oxidant stress (F(2)-isoprostanes) were measured. Cardiac function in the combined shock and clamp rats was significantly depressed compared with sham-operated control rats but was similar to that noted in animals subjected to shock alone. Myocardial TNF-alpha concentrations increased 10-fold in the combined shock and clamp rats compared with sham rats, although there was no difference in myocardial ATP, CP, or F(2)-isoprostanes. TNF-alpha neutralization improved cardiac function by 50% in the combined shock and clamp rats. Hemorrhagic shock is the primary insult inducing cardiac dysfunction in this model of RAAA repair. An improvement in cardiac contractile function after immunoneutralization of TNF-alpha indicates that TNF-alpha mediates a significant portion of the myocardial dysfunction in this model. PMID- 10710364 TI - Protective effect of lung inflation in reperfusion-induced lung microvascular injury. AB - We used the isolated-perfused rat lung model to study the influence of pulmonary ventilation and surfactant instillation on the development of postreperfusion lung microvascular injury. We hypothesized that the state of lung inflation during ischemia contributes to the development of the injury during reperfusion. Pulmonary microvascular injury was assessed by continuously monitoring the wet lung weight and measuring the vessel wall (125)I-labeled albumin ((125)I-albumin) permeability-surface area product (PS). Sprague-Dawley rats (n = 24) were divided into one control group and five experimental groups (n = 4 rats per group). Control lungs were continuously ventilated with 20% O(2) and perfused for 120 min. All lung preparations were ventilated with 20% O(2) before the ischemia period and during the reperfusion period. The various groups differed only in the ventilatory gas mixtures used during the flow cessation: group I, ventilated with 20% O(2); group II, ventilated with 100% N(2); group III, lungs remained collapsed and unventilated; group IV, same as group III but pretreated with surfactant (4 ml/kg) instilled into the airway; and group V, same as group III but saline (4 ml/kg) was instilled into the airway. Control lungs remained isogravimetric with baseline (125)I-albumin PS value of 4.9 +/- 0.3 x 10(-3) ml x min(-1) x g wet lung wt(-1). Lung wet weight in group III increased by 1.45 +/- 0.35 g and albumin PS increased to 17.7 +/- 2.3 x 10(-3), indicating development of vascular injury during the reperfusion period. Lung wet weight and albumin PS did not increase in groups I and II, indicating that ventilation by either 20% O(2) or 100% N(2) prevented vascular injury. Pretreatment of collapsed lungs with surfactant before cessation of flow also prevented the vascular injury, whereas pretreatment with saline vehicle had no effect. These results indicate that the state of lung inflation during ischemia (irrespective of gas mixture used) and supplementation of surfactant prevent reperfusion-induced lung microvascular injury. PMID- 10710365 TI - Influence of sarcoplasmic reticulum calcium loading on mechanical and relaxation restitution. AB - Mechanical and relaxation restitution represent the restoration of contractile force and relaxation, respectively, in premature beats having progressively longer extrasystolic intervals (ESI); these phenomena are related to intracellular activator Ca(2+) by poorly defined mechanisms. We tested the hypothesis that the level of phospholamban [which modulates the affinity of the sarcoplasmic reticulum (SR) Ca(2+)-ATPase for Ca(2+), and thus the SR Ca(2+) load] may be an important determinant of both mechanical and relaxation restitution. Five mice with ablation of the phospholamban (PLB) gene (PLBKO), eight isogenic wild-type controls (129SvJ), eleven mice with PLB overexpression (PLBOE), and nine isogenic wild-type (FVB/N) controls were anesthetized and instrumented with a 1.4-Fr Millar catheter in the left ventricle and a 1-Fr pacemaker in the right atrium. At a cycle length of 200 ms, extrastimuli with increasing ESI were introduced, and the peak rates of left ventricular isovolumic contraction (+/-dP/dt(max)) were normalized and fit to monoexponential equations. In a subset, the protocols were repeated after ryanodine (4 ng/g) was administered to deplete SR Ca(2+) stores. The time constant of mechanical restitution in PLBKO was significantly shorter [6.3 +/- 1.2 (SE) vs. 47.7 +/- 7.6 ms] and began earlier (50 +/- 10 vs. 70 +/- 19 ms) than in 129SvJ. In contrast, the time constant of mechnical restitution was significantly longer (80.3 +/- 7.6 vs. 54.1 +/- 9.2 ms) in PLBOE than in FVB/N. The time constant of relaxation restitution was less in PLBKO than in 129SvJ (26.2 +/- 9.9 vs. 44.6 +/- 3.3, P < 0.05) but was similar in PLBOE and FVB/N (21.1 +/- 6.3 vs. 20.5 +/- 5.7 ms). Intravenous ryanodine decreased significantly the time constants of mechanical restitution in PLBOE, 129SvJ, and FVB/N but was lethal in PLBKO. In contrast, ryanodine increased the time constant of relaxation restitution. Thus 1) the phospholamban level is a critical determinant of mechanical restitution and (to a lesser extent) relaxation restitution in these transgenic models, and 2) ryanodine differentially affects mechanical and relaxation restitution. Furthermore, our data suggest a dissociation of processes within the SR that govern contraction and relaxation. PMID- 10710366 TI - Differential ICP responses elicited by electrical stimulation of medial preoptic area. AB - Recent findings indicate a complex role for the medial preoptic area (MPOA) in modulating penile erection. To further investigate this important area we measured changes in intracavernous pressure (ICP) elicited by electrical stimulation of the MPOA and evaluated the contribution of the cavernous nerve to the ICP responses after bilateral transection of the cavernous nerve (CN). In all experiments electrical stimulation was performed unilaterally in anesthetized male rats. Two distinct patterns of ICP response were seen after electrical stimulation of the MPOA: 1) increases in ICP during electrical stimulation (pattern 1, n = 10 rats) and 2) increases in ICP after electrical stimulation was terminated (pattern 2, n = 10 rats). For pattern 1, increases in ICP during stimulation exhibited a stable plateau without contraction of striated penile muscles, and bilateral transection of the CN eliminated the ICP responses. For pattern 2, increases in ICP observed after stimulation were lower, more variable, and accompanied by significant amplitude variations ("peaks"), caused by contraction of striated penile muscles. Bilateral transection of the CN eliminated the pattern 2 ICP response but did not alter striated muscle contraction. Histological studies documented that pattern 1 and pattern 2 responses occurred via electrical stimulation of the anterior and posterior areas of the MPOA, respectively. Thus both responses appear to result from activation of the CN, but the pattern 2 response apparently involves contraction of the striated penile muscles as well. PMID- 10710367 TI - Modulation of mouse cardiac function in vivo by eNOS and ANP. AB - To study the role of endothelial nitric oxide synthase (eNOS) in cardiac function, we compared eNOS expression, contractility, and relaxation in the left ventricles of wild-type and eNOS-deficient mice. eNOS immunostaining is localized to the macro- and microvascular endothelium throughout the myocardium in wild type mice and is absent in eNOS-/- mice. Whereas blood pressure is elevated in eNOS-/- mice, baseline cardiac contractility (dP/dt(max)) is similar in wild-type and eNOS-/- mice (9,673 +/- 2, 447 and 9,928 +/- 1,566 mmHg/s, respectively). The beta-adrenergic agonist isoproterenol (Iso) at doses of >/=1 ng causes enhanced increases in dP/dt(max) in eNOS-/- mice compared with wild-type controls in vivo (P < 0.01) as well as in Langendorff isolated heart preparations (P < 0.02). beta Adrenergic receptor binding (B(max)) is not significantly different in the two groups of animals (B(max) = 41.4 +/- 9.4 and 36.1 +/- 5.1 fmol/mg for wild-type and eNOS-/-). Iso-stimulated ventricular relaxation is also enhanced in the eNOS /- mice, as measured by dP/dt(min) in the isolated heart. However, baseline ventricular relaxation is normal in eNOS-/- mice (tau = 5.2 +/- 1.0 and 5.6 +/- 1.5 ms for wild-type and eNOS-/-, respectively), whereas it is impaired in wild type mice after NOS inhibition (tau = 8.3 +/- 2.4 ms). cGMP levels in the left ventricle are unaffected by eNOS gene deletion (wild-type: 3.1 +/- 0.8 pmol/mg, eNOS-/-: 3.1 +/- 0.6 pmol/mg), leading us to examine the level of another physiological regulator of cGMP. Atrial natriuretic peptide (ANP) expression is markedly upregulated in the eNOS-/- mice, and exogenous ANP restores ventricular relaxation in wild-type mice treated with NOS inhibitors. These results suggest that eNOS attenuates both inotropic and lusitropic responses to beta-adrenergic stimulation, and it also appears to regulate baseline ventricular relaxation in conjunction with ANP. PMID- 10710368 TI - Localization of dichlorofluorescin in cardiac myocytes: implications for assessment of oxidative stress. AB - Localization and staining features of the oxidant-sensitive fluorescent probe 2'7'-dichlorofluorescin (DCFH) were evaluated in isolated cardiac muscle cells. Cardiomyocytes rapidly accumulated the probe and retained steady levels of DCFH and its highly fluorescent oxidized product dichlorofluorescein (DCF) in probe free medium for 1.5 h. DCF was associated with mitochondria and was released by the proton ionophore carbonyl cyanide m-chlorophenylhydrazone but not by saponin, which permeabilizes the plasma membrane. A mitochondrial distribution of DCF was also suggested by experiments with the mitochondrial marker MitoTracker Red, in which quenching was observed between DCF and MitoTracker Red in live cells. Isolated cardiac mitochondria rapidly accumulated DCF, and high micromolar concentrations of the probe inhibited ADP-stimulated respiration rate. The study provides an information base essential for the interpretation and design of experiments with DCF as a marker of oxidative stress in cardiac muscle and reveals preferential localization of the probe in mitochondria. PMID- 10710370 TI - A novel servo-control system that imposes desired aortic input impedance on in situ rat heart. AB - To clarify the pathophysiological role of dynamic arterial properties in cardiovascular diseases, we attempted to develop a new control system that imposes desired aortic impedance on in situ rat left ventricle. In 38 anesthetized open-chest rats, ascending aortic pressure and flow waveforms were continuously sampled (1,000 Hz). Desired flow waveforms were calculated from measured aortic pressure waveforms and target impedance. To minimize the difference between measured and desired aortic flow waveforms, the computer generated commands to the servo-pump, connected to a side branch of the aorta. By iterating the process, we could successfully control aortic impedance in such a way as to manipulate compliance and characteristic impedance between 60 and 160% of their respective native values. The error between desired and measured aortic flow waveforms was 70 +/- 34 microl/s (root mean square; 4.4 +/- 1.4% of peak flow), indicating reasonable accuracy in controlling aortic impedance. This system enables us to examine the importance of dynamic arterial properties independently of other hemodynamic and neurohumoral factors in physiological and clinical settings. PMID- 10710369 TI - Functional expression of NOS 1 in vascular smooth muscle. AB - Substances that increase intracellular calcium concentration ([Ca(2+)](i)), such as serotonin, are known to induce vascular smooth muscle (VSM) contraction. However, increases in [Ca(2+)](i) also activate Ca(2+)/calmodulin-dependent nitric oxide synthases (NOS), which leads to increases in cGMP and activation of cGMP-dependent protein kinase (PKG). One recently identified substrate protein of PKG is the small heat shock protein, HSP20. The purpose of this study was to determine if serotonin activates a Ca(2+)-dependent NOS in VSM. Strips of bovine carotid arterial smooth muscle denuded of endothelium were stimulated with serotonin in the presence and absence of the nonspecific NOS inhibitor N monomethyl-L-arginine (L-NMMA). Activation of NOS was determined by increases in cGMP and in the phosphorylation of HSP20. Immunohistochemical and Western blotting techniques were performed to identify specific NOS isoforms in bovine carotid arterial smooth muscle preparations. Serotonin stimulation led to significant increases in cGMP and in the phosphorylation of HSP20, which were inhibited by pretreatment with L-NMMA. Antibodies against NOS 1 stained the media of bovine carotid and human renal arteries, whereas antibodies against NOS 3 stained only the endothelium. Additionally, the conversion of radiolabeled L arginine to L-citrulline NOS activity demonstrated a consistent amount of activity present in the endothelium-denuded smooth muscle preparations that was reduced by 99% with an NOS 1 specific inhibitor. Finally, an NOS 1 specific inhibitor, 7-nitroindazole, augmented contractions induced by high extracellular KCl. This study demonstrates that NOS 1 is present in VSM and may effect physiological contractile responses. PMID- 10710371 TI - Mechanisms of ischemic preconditioning effects on Ca(2+) paradox-induced changes in heart. AB - The effects of ischemic preconditioning (IP) on changes in cardiac performance and sarcoplasmic reticulum (SR) function due to Ca(2+) paradox were investigated. Isolated perfused hearts were subjected to IP (three cycles of 3-min ischemia and 3-min reperfusion) followed by Ca(2+)-free perfusion and reperfusion (Ca(2+) paradox). Perfusion of hearts with Ca(2+)-free medium for 5 min followed by reperfusion with Ca(2+)-containing medium for 30 min resulted in a dramatic decrease in the left ventricular (LV) developed pressure and a marked increase in LV end-diastolic pressure. Alterations in cardiac contractile activity due to Ca(2+) paradox were associated with depressed SR Ca(2+)-uptake, Ca(2+)-pump ATPase, and Ca(2+)-release activities as well as decreased SR protein contents for Ca(2+)-pump and Ca(2+) channels. All these changes due to Ca(2+) paradox were significantly prevented in hearts subjected to IP. The protective effects of IP on Ca(2+) paradox changes in cardiac contractile activity as well as SR Ca(2+) pump and Ca(2+)-release activities were lost when the hearts were treated with 8 (p-sulfophenyl)-theophylline, an adenosine receptor antagonist; KN-93, a specific Ca(2+)/calmodulin-dependent protein kinase II (CaMK II) inhibitor; or chelerythrine chloride, a protein kinase C (PKC) inhibitor. These results indicate that IP rendered cardioprotection by preventing a depression in SR function in Ca(2+) paradox hearts. Furthermore, these beneficial effects of IP may partly be mediated by adenosine receptors, PKC, and CaMK II. PMID- 10710372 TI - Sprint performance is related to muscle fascicle length in male 100-m sprinters. AB - The purpose of this study was to investigate the relationship between muscle fascicle length and sprint running performance in 37 male 100-m sprinters. The sample was divided into two performance groups by the personal-best 100-m time: 10.00-10.90 s (S10; n = 22) and 11.00-11.70 s (S11; n = 15). Muscle thickness and fascicle pennation angle of the vastus lateralis and gastrocnemius medialis and lateralis muscles were measured by B-mode ultrasonography, and fascicle length was estimated. Standing height, body weight, and leg length were similar between groups. Muscle thickness was similar between groups for vastus lateralis and gastrocnemius medialis, but S10 had a significantly greater gastrocnemius lateralis muscle thickness. S10 also had a greater muscle thickness in the upper portion of the thigh, which, given similar limb lengths, demonstrates an altered "muscle shape." Pennation angle was always less in S10 than in S11. In all muscles, S10 had significantly greater fascicle length than did S11, which significantly correlated with 100-m best performance (r values from -0.40 to 0.57). It is concluded that longer fascicle length is associated with greater sprinting performance. PMID- 10710373 TI - Interaction between left and right intercostal muscles in airway pressure generation. AB - The interactions between the different rib cage inspiratory muscles in the generation of pleural pressure remain largely unknown. In the present study, we have assessed in dogs the interactions between the parasternal intercostals and the interosseous intercostals situated on the right and left sides of the sternum. For each set of muscles, the changes in airway opening pressure (DeltaPao) obtained during separate right and left activation were added, and the calculated values (predicted DeltaPao) were then compared with the DeltaPao values obtained during symmetric, bilateral activation (measured DeltaPao). When the parasternal intercostals in one or two interspaces were activated, the measured DeltaPao was commonly greater than the predicted value. The difference, however, was only 10%. When the interosseous intercostals were activated, the measured DeltaPao was nearly equal to the predicted value. These observations strengthen our previous conclusion that the pressure changes produced by the rib cage inspiratory muscles are essentially additive. As a corollary, the rib cage can be considered as a linear elastic structure over a wide range of distortion. PMID- 10710374 TI - Lung function and ventilation inhomogeneity in rat lungs after allergen challenge. AB - We studied the early response to ovalbumin challenge in sensitized Brown-Norway rats through its effect on N(2), He, and SF(6) phase III slopes of the single breath washout and on indexes of lung function. Sensitized rats showed varying degrees of response in terms of pulmonary pressure (PL), with increases ranging between 125 and 225% of baseline. The sensitized rats presented decreased quasistatic compliance, forced vital capacity, and end-expiratory flow, with all three lung function indexes showing a significant negative correlation with corresponding PL values. They also showed significant positive correlations of PL with the N(2), He, and SF(6) phase III slopes, reflecting diffusion-convection dependent inhomogeneities generated by conformation changes throughout the entire rat lung. In addition, the rats showing the most marked PL increases (>150% baseline PL) also revealed a reversal of the SF(6)-He slope difference because of a more marked SF(6) than He slope increase. This latter finding suggests that the degree of structural heterogeneity during early response is even more marked in the most peripheral rat lung generations. PMID- 10710375 TI - Endothelium-dependent relaxation differs in porcine pulmonary arteries from the left and right caudal lobes. AB - We hypothesized that pulmonary arteries (PA) from identical branch orders within left and right caudal lung lobes would exhibit similar vasomotor responses. Arterial rings from caudal lung lobes of female swine were examined in vitro. Vascular smooth muscle contraction to KCl and norepinephrine did not differ. Vascular relaxation to endothelium-dependent (bradykinin, acetylcholine, A-23187) and -independent (sodium nitroprusside, zero-calcium Krebs solution) vasodilators was assessed. Right PA exhibited less maximal relaxation to acetylcholine (50%) than did left PA (69%; P < 0.001). Maximal relaxation to sodium nitroprusside did not differ, although right PA had a lower drug concentration resulting in half maximal relaxation (6.26 x 10(-8) M) than did left PA (9.57 x 10(-8) M; P < 0.05). Nitric oxide synthase inhibition with an arginine analog (N(omega)-nitro-L arginine methyl ester) depressed acetylcholine-induced relaxation but the left vs. right difference persisted. Indomethacin enhanced relaxation to acetylcholine and abolished the difference between left and right. We conclude that endothelium dependent vasorelaxation is less in porcine right than in left PA because of greater release of one or more constricting prostanoids in arteries from the right caudal lobe. PMID- 10710376 TI - Oxidant-increased endothelial permeability: prevention with phosphodiesterase inhibition vs. cAMP production. AB - The present objective was to determine whether hydrogen peroxide (H(2)O(2)) increases transvascular albumin clearance and lung weight in an isolated rat lung and whether posttreatment with cAMP-enhancing agents can prevent these increases. Transvascular albumin clearance was assessed by (125)I-labeled albumin clearance ((125)I-albumin flux/perfusate concentration of (125)I-albumin) at a given fluid filtration. Nonlinear regression analysis of transvascular albumin clearance vs. fluid filtration yielded values for the permeability-surface area product (PS) and the reflection coefficient (sigma). H(2)O(2) decreased sigma from a control value of 0.93 to 0.38, did not change PS, and increased lung weight. Posttreatment with isoproterenol, a beta(2)-adrenergic-receptor agonist, reduced the H(2)O(2)-induced decrease in sigma to 0.65 and augmented the increase in lung weight. Posttreatment with CP-80633, a phosphodiesterase 4 inhibitor, further reduced the H(2)O(2)-induced decrease in sigma to 0.79 and blocked the rise in lung weight. In the presence of isoproterenol or CP-80633, H(2)O(2) increased PS. Therefore, H(2)O(2) increased the convective and diffusive clearances of albumin across an intact pulmonary vasculature. Furthermore, inhibition of cAMP metabolism more effectively attenuated the H(2)O(2)-induced increases in convective albumin clearance and lung weight as compared with stimulation of cAMP production. PMID- 10710377 TI - Altered gravity downregulates aquaporin-1 protein expression in choroid plexus. AB - Aquaporin-1 (AQP1) is a water channel expressed abundantly at the apical pole of choroidal epithelial cells. The protein expression was quantified by immunocytochemistry and confocal microscopy in adult rats adapted to altered gravity. AQP1 expression was decreased by 64% at the apical pole of choroidal cells in rats dissected 5.5-8 h after a 14-day spaceflight. AQP1 was significantly overexpressed in rats readapted for 2 days to Earth's gravity after an 11-day flight (48% overshoot, when compared with the value measured in control rats). In a ground-based model that simulates some effects of weightlessness and alters choroidal structures and functions, apical AQP1 expression was reduced by 44% in choroid plexus from rats suspended head down for 14 days and by 69% in rats suspended for 28 days. Apical AQP1 was rapidly enhanced in choroid plexus of rats dissected 6 h after a 14-day suspension (57% overshoot, in comparison with control rats) and restored to the control level when rats were dissected 2 days after the end of a 14-day suspension. Decreases in the apical expression of choroidal AQP1 were also noted in rats adapted to hypergravity in the NASA 24-ft centrifuge: AQP1 expression was reduced by 47% and 85% in rats adapted for 14 days to 2 G and 3 G, respectively. AQP1 is downregulated in the apical membrane of choroidal cells in response to altered gravity and is rapidly restored after readaptation to normal gravity. This suggests that water transport, which is partly involved in the choroidal production of cerebrospinal fluid, might be decreased during spaceflight and after chronic hypergravity. PMID- 10710378 TI - In vivo estimation of contraction velocity of human vastus lateralis muscle during "isokinetic" action. AB - To determine the shortening velocities of fascicles of the vastus lateralis muscle (VL) during isokinetic knee extension, six male subjects were requested to extend the knee with maximal effort at angular velocities of 30 and 150 degrees /s. By using an ultrasonic apparatus, longitudinal images of the VL were produced every 30 ms during knee extension, and the fascicle length and angle of pennation were obtained from these images. The shortening fascicle length with extension of the knee (from 98 to 13 degrees of knee angle; full extension = 0 degrees ) was greater (43 mm) at 30 degrees /s than at 150 degrees /s (35 mm). Even when the angular velocity remained constant during the isokinetic range of motion, the fascicle velocity was found to change from 39 to 77 mm/s at 150 degrees /s and from 6 to 19 mm/s at 30 degrees /s. The force exerted by a fascicle changed with the length of the fascicle at changing angular velocities. The peak values of fascicle force and velocity were observed at approximately 90 mm of fascicle length. In conclusion, even if the angular velocity of knee extension is kept constant, the shortening velocity of a fascicle is dependent on the force applied to the muscle-tendon complex, and the phenomenon is considered to be caused mainly by the elongation of the elastic element (tendinous tissue). PMID- 10710379 TI - Tyrosine kinase inhibitors modulate agonist-induced vasodilation in the hamster cheek pouch. AB - The purpose of this study was to determine whether inhibitors of tyrosine kinase attenuate vasodilation elicited by endogenously elaborated and exogenously applied nitric oxide in the in situ peripheral microcirculation. Using intravital microscopy, we found that pretreatment with genistein (1.0 microM) and tyrphostin 25 (10.0 microM), two structurally unrelated tyrosine kinase inhibitors, significantly attenuated acetylcholine-, bradykinin- and nitroglycerin-induced dilation of second-order arterioles (51 +/- 1 microm) in the in situ hamster cheek pouch (P < 0.05). Both inhibitors nearly abrogated acetylcholine-induced responses but only partially blocked bradykinin- and nitroglycerin-induced vasodilation. Genistein and tyrphostin 25 alone had no significant effects on resting arteriolar diameter and on adenosine-induced vasodilation in the cheek pouch. On balance, these data indicate that tyrosine kinase inhibitors attenuate endogenously elaborated and exogenously applied nitric oxide-induced vasodilation in the in situ hamster cheek pouch. However, the extent of tyrosine kinase inhibitor-sensitive pathway involvement in this response appears to be agonist dependent. PMID- 10710380 TI - Protective and defensive airway reflexes evoked by nasal exposure to wood smoke in anesthetized rats. AB - We investigated the airway responses evoked by nasal wood smoke in anesthetized Sprague-Dawley rats. Wood smoke (5 ml, 1.4 ml/s) was delivered into an isolated nasal cavity while animals breathed spontaneously. In study 1, nasal wood smoke triggered either an apneic response (n = 26) or a sniff-like response (n = 16) within 1 s after smoke exposure in 42 normal rats. Both airway responses were abolished by trigeminal nerve denervation and by nasal application of a local anesthetic or a hydroxyl radical scavenger, but they were not significantly affected by removal of smoke particulates or nasal application of a saline vehicle. In study 2, nasal wood smoke only triggered a mild apneic response in two rats neonatally treated with capsaicin and had no effect on breathing in the other six; the treatment is known to chronically ablate C fibers and some Adelta fibers. In contrast, nasal wood smoke evoked an apneic response in six rats neonatally treated with the vehicle of capsaicin and elicited a sniff-like response in the other two. These results suggest that the apneic and sniff-like responses evoked by nasal wood smoke result from the stimulation of trigeminal nasal C-fiber and Adelta-fiber afferents by the gas-phase smoke and that hydroxyl radical is the triggering chemical factor. PMID- 10710381 TI - Blunted pressor and intramuscular metabolic responses to voluntary isometric exercise in multiple sclerosis. AB - To test the hypothesis that a lower mean arterial pressure (MAP) response during voluntary isometric exercise in multiple sclerosis (MS) is related to a dampened muscle metabolic signal, 9 MS and 11 control subjects performed an isometric dorsiflexor contraction at 30% maximal voluntary contraction until target failure (endurance time). We made continuous and noninvasive measurements of heart rate and MAP (Finapres) and of intramuscular pH and P(i) (phosphorus magnetic resonance spectroscopy) in a subset of 6 MS and 10 control subjects. Endurance times and change in heart rate were similar in MS and control subjects. The decrease in pH and increase in P(i) were less throughout exercise in MS compared with control subjects, as was the change in MAP response. Differences in muscle strength accounted for some of the difference in MAP response between groups. Cardiovascular responses during Valsalva and cold pressor tests were similar in MS and control subjects, suggesting that the blunted MAP response during exercise in MS was not due to a generalized dysautonomia. The dampened metabolic response in MS subjects was not explained by inadequate central muscle activation. These data suggest that the blunted pressor response to exercise in MS subjects may be largely appropriate to a blunted muscle metabolic response and differences in contracting muscle mass. PMID- 10710382 TI - Cellular adaptations in fat tissue of exercise-trained miniature swine: role of excess energy intake. AB - This study examined the influence of energy expenditure and energy intake on cellular mechanisms regulating adipose tissue metabolism. Twenty-four swine were assigned to restricted-fed sedentary, restricted-fed exercise-trained, full-fed sedentary, or full-fed exercise-trained groups. After 3 mo of treatment, adipocytes were isolated and adipocyte size, adenosine A(1) receptor characteristics, and lipolytic sensitivity were measured. Swine were infused with epinephrine during which adipose tissue extracellular adenosine, plasma fatty acids, and plasma glycerol were measured. Results revealed that adipocytes isolated from restricted-fed exercised swine had a smaller diameter, a lower number of A(1) receptors, and a greater sensitivity to lipolytic stimulation, compared with adipocytes from full-fed exercised swine. Extracellular adenosine levels were transiently increased on infusion of epinephrine in adipose tissue of restricted-fed exercised but not full-fed exercised swine. These results suggest a role for adenosine in explaining the discrepancy between in vitro and in vivo lipolysis findings and underscore the notion that excess energy intake dampens the lipolytic sensitivity of adipocytes to beta-agonists and adenosine, even if accompanied by exercise training. PMID- 10710383 TI - Ligation of the bronchial artery in sheep attenuates early pulmonary changes following exposure to smoke. AB - Smoke inhalation can produce acute pulmonary edema. Previous studies have shown that the bronchial arteries are important in acute pulmonary edema occurring after inhalation of a synthetic smoke containing acrolein, a common smoke toxin. We hypothesized that inhalation of smoke from burning cotton, known to contain acrolein, would produce in sheep acute pulmonary edema that was mediated by the bronchial circulation. We reasoned that occluding the bronchial arteries would eliminate smoke-induced pulmonary edema, whereas occlusion of the pulmonary artery would not. Smoke inhalation increased lung lymph flow from baseline from 2.4 +/- 0.7 to 5.6 +/- 1.2 ml/0.5 h at 30 min (P < 0.05) to 9.1 +/- 1 ml/0.5 h at 4 h (P < 0.05). Bronchial artery ligation diminished and delayed the rise in lymph flow with baseline at 2.8 +/- 0.7 ml/0.5 h rising to 3.1 +/- 0. 8 ml/0.5 h at 30 min to 6.5 +/- 1.5 ml/0.5 h at 240 min (P < 0.05). Wet-to-dry ratio was 4.1 +/- 0.2 in control, 5.1 +/- 0.3 in smoke inhalation (P < 0.05), and 4.4 +/- 0.4 in bronchial artery ligation plus smoke-inhalation group. Smoke inhalation after occlusion of the right pulmonary artery resulted in a wet-to-dry ratio after 4 h in the right lung of 5.5 +/- 0.8 (P < 0.05 vs. control) and in the left nonoccluded lung of 5.01 +/- 0.7 (P < 0.05). Thus the bronchial arteries may be major contributors to acute pulmonary and airway edema following smoke inhalation because the edema occurs in the lung with the pulmonary artery occluded but not in the lungs with bronchial arteries ligated. PMID- 10710384 TI - Effects of spaceflight and thyroid deficiency on hindlimb development. I. Muscle mass and IGF-I expression. AB - Thyroid deficiency (TD) in neonatal rats causes reduced growth of skeletal muscle that is disproportionately greater than that for other tissues (G. R. Adams, S. A. McCue, M. Zeng, and K. M. Baldwin. Am. J. Physiol. Regulatory Integrative Comp. Physiol. 276: R954-R961, 1999). TD depresses plasma insulin-like growth factor I (IGF-I) levels, suggesting a mechanism for this effect. We hypothesized that TD and exposure to spaceflight (SF) would interact to reduce skeletal muscle growth via a reduction in IGF-I levels. Neonatal rats were flown in space for 16 days. There was a similar, nonadditive reduction in the growth of the body ( approximately 50%) and muscle weight (fast muscles, approximately 60%) with either TD or SF. In the soleus muscle, either SF or TD alone resulted in growth reductions that were augmented by SF-TD interactions. There were strong correlations between 1) muscle mass and muscle IGF-I levels and 2) circulating IGF-I and body weight. These results indicate that either hypothyroidism or exposure to SF will limit the somatic and muscle-specific growth of neonatal rats. The impact of these perturbations on skeletal muscle growth is relatively greater than the effect on somatic growth. The mechanisms by which either TD or SF impact growth appear to have a common pathway involving the control of plasma and muscle IGF-I concentrations. PMID- 10710385 TI - Effects of spaceflight and thyroid deficiency on rat hindlimb development. II. Expression of MHC isoforms. AB - Both slow-twitch and fast-twitch muscles are undifferentiated after birth as to their contractile protein phenotype. Thus we examined the separate and combined effects of spaceflight (SF) and thyroid deficiency (TD) on myosin heavy chain (MHC) gene expression (protein and mRNA) in muscles of neonatal rats (7 and 14 days of age at launch) exposed to SF for 16 days. Spaceflight markedly reduced expression of the slow, type I MHC gene by approximately 55%, whereas it augmented expression of the fast IIx and IIb MHCs in antigravity skeletal muscles. In fast muscles, SF caused subtle increases in the fast IIb MHC relative to the other adult MHCs. In contrast, TD prevented the normal expression of the fast MHC phenotype, particularly the IIb MHC, whereas TD maintained expression of the embryonic/neonatal MHC isoforms; this response occurred independently of gravity. Collectively, these results suggest that normal expression of the type I MHC gene requires signals associated with weight-bearing activity, whereas normal expression of the IIb MHC requires an intact thyroid state acting independently of the weight-bearing activities typically encountered during neonatal development of laboratory rodents. Finally, MHC expression in developing muscles is chiefly regulated by pretranslational processes based on the tight relationship between the MHC protein and mRNA data. PMID- 10710386 TI - Development of a mathematical model that predicts optimal muscle activation patterns by using brief trains. AB - Because muscles must be repetitively activated during functional electrical stimulation, it is desirable to identify the stimulation pattern that produces the most force. Previous experimental work has shown that the optimal pattern contains an initial high-frequency burst of pulses (i.e., an initial doublet or triplet) followed by a low, constant-frequency portion. Pattern optimization is particularly challenging, because a muscle's contractile characteristics and, therefore, the optimal pattern change under different physiological conditions and are different for each person. This work describes the continued development and testing of a mathematical model that predicts isometric forces from fresh and fatigued muscles in response to brief trains of electrical pulses. By use of this model and an optimization algorithm, stimulation patterns that produced maximum forces from each subject were identified. PMID- 10710387 TI - Influence of positive airway pressure on the pressure gradient for venous return in humans. AB - To study the effect of positive airway pressure (Paw) on the pressure gradient for venous return [the difference between mean systemic filling pressure (Pms) and right atrial pressure (Pra)], we investigated 10 patients during general anesthesia for implantation of defibrillator devices. Paw was varied under apnea from 0 to 15 cmH(2)O, which increased Pra from 7.3 +/- 3.1 to 10.0 +/- 2.3 mmHg and decreased left ventricular stroke volume by 23 +/- 22%. Episodes of ventricular fibrillation, induced for defibrillator testing, were performed during 0- and 15-cmH(2)O Paw to measure Pms (value of Pra 7.5 s after onset of circulatory arrest). Positive Paw increased Pms from 10.2 +/- 3.5 to 12.7 +/- 3.2 mmHg, and thus the pressure gradient for venous return (Pms - Pra) remained unchanged. Echocardiography did not reveal signs of vascular collapse of the inferior and superior vena cava due to lung expansion. In conclusion, we demonstrated that positive Paw equally increases Pra and Pms in humans and alters venous return without changes in the pressure gradient (Pms - Pra). PMID- 10710388 TI - Effects of loaded breathing and hypoxia on diaphragm metabolism as measured by (31)P-NMR spectroscopy. AB - Diaphragm fatigue may contribute to respiratory failure. (31)P-nuclear magnetic resonance spectroscopy is a useful tool to assess energetic changes within the diaphragm during fatigue, as indicated by P(i) accumulation and phosphocreatine (PCr) depletion. We hypothesized that loaded breathing during hypoxia would lead to diaphragm fatigue and inadequate aerobic metabolism. Seven piglets were anesthetized by using halothane inhalation. Diaphragmatic contractility was assessed by transdiaphragmatic pressure (Pdi) at end expiration with the airway occluded. A nuclear magnetic resonance surface coil placed under the right hemidiaphragm measured P(i) and PCr during four conditions: control, inspiratory resistive breathing (IRB), IRB with hypoxia, and recovery (IRB without hypoxia). IRB alone resulted in hypercarbia (32 +/- 7 to 61 +/- 21 Torr) and respiratory acidosis but no change in diaphragm force output or aerobic metabolism. Combined IRB and hypoxia resulted in decreased force output (Pdi decreased by 40%; from 30 +/- 17 to 19 +/- 11 mmHg) and evidence of metabolic stress (ratio of P(i) to PCr increased by 290%; from 0.19 +/- 0.09 to 0.74 +/- 0.27). We conclude that diaphragm fatigue associated with inadequate aerobic oxidative metabolism occurs in the setting of loaded breathing and hypoxia. Conversely, aerobic metabolism and force output of the diaphragm remain unchanged from control during loaded normoxic or hyperoxic breathing despite the onset of respiratory failure. PMID- 10710389 TI - Respiratory mechanics and maximal expiratory flow in the anesthetized mouse. AB - Mice have been widely used in immunologic and other research to study the influence of different diseases on the lungs. However, the respiratory mechanical properties of the mouse are not clear. This study extended the methodology of measuring respiratory mechanics of anesthetized rats and guinea pigs and applied it to the mouse. First, we performed static pressure-volume and maximal expiratory flow-volume curves in 10 anesthetized paralyzed C57BL/6 mice. Second, in 10 mice, we measured dynamic respiratory compliance, forced expiratory volume in 0.1 s, and maximal expiratory flow before and after methacholine challenge. Averaged total lung capacity and functional residual capacity were 1.05 +/- 0.04 and 0.25 +/- 0.01 ml, respectively, in 20 mice weighing 22.2 +/- 0.4 g. The chest wall was very compliant. In terms of vital capacity (VC) per second, maximal expiratory flow values were 13.5, 8.0, and 2.8 VC/s at 75, 50, and 25% VC, respectively. Maximal flow-static pressure curves were relatively linear up to pressure equal to 9 cm H(2)O. In addition, methacholine challenge caused significant decreases in respiratory compliance, forced expiratory volume in 0.1 s, and maximal expiratory flow, indicating marked airway constriction. We conclude that respiratory mechanical parameters of mice (after normalization with body weight) are similar to those of guinea pigs and rats and that forced expiratory maneuver is a useful technique to detect airway constriction in this species. PMID- 10710390 TI - A comparison of bioimpedance methods for detection of body cell mass change in HIV infection. AB - The maintenance of body cell mass (BCM) is critical for survival in human immunodeficiency virus (HIV) infection. Accuracy of bioimpedance for measuring change (Delta) in intracellular water (ICW), which defines BCM, is uncertain. To evaluate bioimpedance-estimated DeltaBCM, the ICW of 21 weight-losing HIV patients was measured before and after anabolic steroid therapy by dilution (total body water by deuterium - extracellular water by bromide) and bioimpedance. Multiple-frequency modeling- and dilution-determined DeltaICW did not differ. The DeltaICW was predicted poorly by 50-kHz parallel reactance, 50 kHz impedance, and 200 - 5-kHz impedance. The DeltaICW predicted by 500 - 5-kHz impedance was closer to, but statistically different from, dilution-determined DeltaICW. However, the effect of random error on the measurement of systematic error in the 500 - 5-kHz method was 12-13% of the average measured DeltaICW; this was nearly twice the percent difference between obtained and threshold statistics. Although the 500 - 5-kHz method cannot be fully rejected, these results support the conclusion that only the multiple-frequency modeling approach accurately monitors DeltaBCM in HIV infection. PMID- 10710391 TI - Carotid baroreflex control of heart rate and blood pressure during ES leg cycling in paraplegics. AB - This study investigated control of heart rate (HR) and mean arterial pressure (MAP) at rest and during electrical stimulation (ES) leg cycling exercise (LCE) in paraplegics (Para). Seven men with complete spinal lesions (T(5)-T(11)) and six able-bodied (AB) men participated in this study. Beat-to-beat changes in HR and MAP were recorded during carotid sinus perturbation. Carotid baroreflex function curves were derived at rest and during ES-LCE for Para and during voluntary cycling (Vol) for AB. From rest to ES-LCE, oxygen uptake (VO(2)) increased (by 0.43 l/min) and HR rose (by 11 beats/min), yet MAP remained unchanged. In AB, Vol increased VO(2) (by 0.53 l/min), HR (by 22 beats/min), and MAP (by 8 mmHg). ES-LCE did not alter the carotid sinus pressure (CSP)-MAP relationship, but it displaced the CSP-HR relationship upward relative to rest. No rightward shift was observed during ES-LCE. Vol by AB produced an upward and rightward displacement of the CSP-MAP and CSP-HR relationships relative to rest. These findings suggested that the carotid sinus baroreflex was not reset during ES-LCE in Para. PMID- 10710392 TI - Effect of sleep restriction on orthostatic cardiovascular control in humans. AB - We hypothesized that sleep restriction (4 consecutive nights, 4 h sleep/night) attenuates orthostatic tolerance. The effect of sleep restriction on cardiovascular responses to simulated orthostasis, arterial baroreflex gain, and heart rate variability was evaluated in 10 healthy volunteers. Arterial baroreflex gain was determined from heart rate responses to nitroprusside phenylephrine injections, and orthostatic tolerance was tested via lower body negative pressure (LBNP). A Finapres device measured finger arterial pressure. No difference in baroreflex function, heart rate variability, or LBNP tolerance was observed with sleep restriction (P > 0.3). Systolic pressure was greater at -60 mmHg LBNP after sleep restriction than before sleep restriction (110 +/- 6 and 124 +/- 3 mmHg before and after sleep restriction, respectively, P = 0.038), whereas heart rate decreased (108 +/- 8 and 99 +/- 8 beats/min before and after sleep restriction, respectively, P = 0.028). These data demonstrate that sleep restriction produces subtle changes in cardiovascular responses to simulated orthostasis, but these changes do not compromise orthostatic tolerance. PMID- 10710393 TI - Myofibrillar or mitochondrial creatine kinase deficiency alone does not impair mouse diaphragm isotonic function. AB - Creatine kinase (CK) provides ATP buffering in skeletal muscle and is expressed as 1) cytosolic myofibrillar CK (M-CK) and 2) sarcomeric mitochondrial CK (ScCKmit) isoforms that differ in their subcellular localization. The diaphragm (Dia) expresses both M-CK and ScCKmit in abundance. We compared the power and work output of 1) control CK-sufficient (Ctl), 2) M-CK-deficient [M-CK(-/-)], 3) ScCKmit-deficient [ScCKmit(-/-)], and 4) combined M-CK/ScCKmit-deficient null mutant [CK(-/-)] Dia during repetitive isotonic activations to determine the effect of CK phenotype on Dia function. Maximum power was obtained at approximately 0.4 tetanic force in all groups. M-CK(-/-) and ScCKmit(-/-) Dia were able to sustain power and work output at Ctl levels during repetitive isotonic activation (75 Hz, 330-ms duration repeated each second at 0.4 tetanic force load), and the duration of sustained Dia shortening was 67 +/- 4 s in M-CK( /-), 60 +/- 4 s in ScCKmit(-/-), and 62 +/- 5 s in Ctl Dia. In contrast, CK(-/-) Dia power and work declined acutely and failed to sustain shortening altogether by 40 +/- 6 s. We conclude that Dia power and work output are not absolutely dependent on the presence of either M-CK or ScCKmit, whereas the complete absence of CK acutely impairs Dia shortening capacity during repetitive activation. PMID- 10710394 TI - Pulmonary vascular response of the coati to chronic hypoxia. AB - The unusually muscular pulmonary arteries normally present in cattle and swine residing at low altitude are associated with a rapid development of severe pulmonary hypertension when those animals are moved to high altitude. Because these species lack collateral ventilation, they appear to have an increased dependence on hypoxic vasoconstriction to maintain normal ventilation-perfusion balance, which, in turn, maintains thickened arterial walls. The only other species known to lack collateral ventilation is the coati, which, similarly, has thick-walled pulmonary arteries. We tested the hypothesis that coatis will develop severe high-altitude pulmonary hypertension by exposing six of these animals (Nasua narica) to a simulated altitude of 4,900 m for 6 wk. After the exposure, pulmonary arterial pressures were hardly elevated, right ventricular hypertrophy was minimal, there was no muscularization of pulmonary arterioles, and, most surprising of all, there was a decrease in medial thickness of muscular pulmonary arteries. These unexpected results break a consistent cross-species pattern in which animals with thick muscular pulmonary arteries at low altitude develop severe pulmonary hypertension at high altitude. PMID- 10710395 TI - Differential facilitation of high- and low-output nerve terminals from a single motoneuron. AB - In the crayfish opener neuromuscular preparation, regional differences in synaptic transmission are observed among the terminals of a single motoneuron. With a single stimulus, the high-output terminals of the proximal region of the muscle produce a larger excitatory postsynaptic potential than do the low-output terminals of the central region of the muscle. We tested the hypothesis that the low-output terminals exhibit more facilitation than do high-output terminals for twin-pulse, train, and continuous-stimulation paradigms. Previous studies have not employed several stimulation paradigms to induce facilitation among high- and low-output terminals of a single motoneuron. We found that the high-output terminals on the proximal fibers facilitate more than the low-output terminals on the central muscle fibers, in contrast with previous studies on similar muscles. The difference in measured facilitation is dependent on the stimulation paradigm. These results are important because ultrastructural differences between these high- and low-output terminals are known and can be used for correlation with physiological measurements. Short-term facilitation is a form of short-term memory at the synaptic level, and the processes understood at the crayfish neuromuscular junction may well be applicable to all chemical synapses. PMID- 10710396 TI - Lung resistance and elastance in spontaneously breathing preterm infants: effects of breathing pattern and demographics. AB - Reported values of lung resistance (RL) and elastance (EL) in spontaneously breathing preterm neonates vary widely. We hypothesized that this variability in lung properties can be largely explained by both inter- and intrasubject variability in breathing pattern and demographics. Thirty-three neonates receiving nasal continuous positive airway pressure [weight 606-1,792 g, gestational age (GA) of 25-33 wk, 2-49 days old] were studied. Transpulmonary pressure was measured by esophageal manometry and airway flow by face mask pneumotachography. Breath-to-breath changes in RL and EL in each infant were estimated by Fourier analysis of impedance (Z) and by multiple linear regression (MLR). RL(MLR) (RL(MLR) = 0.85 x RL(Z) -0.43; r(2) = 0.95) and EL(MLR) (EL(MLR) = 0.97 x EL(Z) + 8.4; r(2) = 0.98) were highly correlated to RL(Z) and EL(Z), respectively. Both RL (mean +/- SD; RL(Z) = 70 +/- 38, RL(MLR) = 59 +/- 36 cm H(2)O x s x l(-1)) and EL (EL(Z) = 434 +/- 212, EL(MLR) = 436 +/- 210 cm H(2)O/l) exhibited wide intra- and intersubject variability. Regardless of computation method, RL was found to decrease as a function of weight, age, respiratory rate (RR), and tidal volume (VT) whereas it increased as a function of RR. VT and inspiratory-to-expiratory time ratio (TI/TE). EL decreased with increasing weight, age, VT and female gender and increased as RR and TI/TE increased. We conclude that accounting for the effects of breathing pattern variability and demographic parameters on estimates of RL and EL is essential if they are to be of clinical value. Multivariate statistical models of RL and EL may facilitate the interpretation of lung mechanics measurements in spontaneously breathing infants. PMID- 10710397 TI - Heat acclimation improves regulation of plasma volume and plasma Na(+) content during exercise in horses. AB - This study determined the plasma volume (PV) and ion responses to heat acclimation and exercise in six trained Thoroughbred horses during 21 days of exposure to heat and humidity (33 degrees C, 83% relative humidity) for 4 h/day. During the 2nd h on days 0, 3, 7, 14, and 21, horses performed a standardized treadmill test, running at 50% of peak O(2) uptake until pulmonary artery temperature reached 41.5 degrees C. Heat acclimation resulted in an increase in PV from 21.3 +/- 1.1 liters on day 0 to 24.3 +/- 1.0 liters on day 14, returning to 22.6 +/- 0.9 liters on day 21. The corresponding total plasma protein contents were 1,273 +/- 53, 1,455 +/- 81, and 1,377 +/- 57 g, respectively, and increases in total plasma Na(+) plus Cl(-) content were 5,145 +/- 126, 5,749 +/- 146, and 5,394 +/- 114 mmol, respectively. Thus changes in PV were accompanied by direct changes in plasma protein and osmolyte contents. With exercise on day 0, PV decreased by 7.1 +/- 0.7% at 5 min of exercise and remained decreased (-6.7 +/- 1.3%) at 5 min of recovery. By day 21, PV decreased significantly less than on day 0 (by 5.2 +/- 0.9% at 5 min of exercise), was decreased by only 2.0 +/- 1.6% at 5 min of recovery, and was fully restored at 15 min of recovery. Plasma Na(+) concentration increased 3 meq/l during the first 5 min of exercise and was normalized by 5 min of recovery on day 0 and by end exercise on day 21. It is concluded that improved ability to regulate PV during exercise in response to heat acclimatization is associated with an increased PV and an improved conservation of Na(+). PMID- 10710398 TI - Mechanical properties of the tracheal mucosal membrane in the rabbit. I. steady state stiffness as a function of age. AB - Airway responsiveness is exaggerated in infancy and declines with maturation. These age-related differences (R.S. Tepper, T. Du, A. Styhler, M. Ludwig, and J.G. Martin. Am. J. Respir. Crit. Care Med. 151: 836-840, 1995; R.S. Tepper, S.J. Gunst, C.M. Doerschuk, Y. Shen, and W. Bray. J. Appl. Physiol. 78: 505-512, 1995; R.S. Tepper, J. Stevens, and H. Eigen. Am. J. Respir. Crit. Care Med. 149: 678 681, 1994) could be due to changes in the smooth muscle, the lung, and/or the airway wall. Folding of the mucosal membrane can provide an elastic load (R.K. Lambert, J. Appl. Physiol. 71: 666-673, 1991), which impedes smooth muscle shortening. We hypothesized that increased stiffness of the mucosal membrane occurs during aging, causing an increased mechanical load on airway smooth muscle and a decrease in airway responsiveness. Forty female New Zealand White rabbits between 0.75 and 35 mo of age were studied. Rectangular mucosal membrane strips oriented both longitudinally and circumferentially to the long axis of the trachea were dissected, and the stress-strain relationships of each strip were tested. The results showed that the membrane was stiffer in the longitudinal than in the circumferential direction of the airway. However, there was no significant change with age in either orientation. We conclude that the mechanical properties of the airway mucosal membrane did not change during maturation and were not likely to influence age-related changes in airway responsiveness. PMID- 10710399 TI - Mechanical properties of the tracheal mucosal membrane in the rabbit. II. Morphometric analysis. AB - Folding of the airway mucosal membrane provides a mechanical load that impedes airway smooth muscle contraction. Mechanical testing of rabbit tracheal mucosal membrane showed that the membrane is stiffer in the longitudinal than in the circumferential direction of the airway. To explain this difference in the mechanical properties, we studied the morphological structure of the rabbit tracheal mucosal membrane in both longitudinal and circumferential directions. The collagen fibers were found to form a random meshwork, which would not account for differences in stiffness in the longitudinal and circumferential directions. The volume fraction of the elastic fibers was measured using a point-counting technique. The orientation of the elastic fibers in the tissue samples was measured using a new method based on simple geometry and probability. The results showed that the volume fraction of the elastic fibers in the rabbit tracheal mucosal membrane was approximately 5% and that the elastic fibers were mainly oriented in the longitudinal direction. Age had no statistically significant effect on either the volume fraction or the orientation of the elastic fibers. Linear correlations were found between the steady-state stiffness and the quantity of the elastic fibers oriented in the direction of testing. PMID- 10710400 TI - Angiotensin-converting enzyme ID polymorphism and fitness phenotype in the HERITAGE Family Study. AB - It has been suggested that genetic variation in the angiotensin-converting enzyme (ACE) gene is associated with physical performance. We studied the association between the ACE insertion (I)/deletion (D) polymorphism and several fitness phenotypes measured before and after 20 wk of a standardized endurance training program in sedentary Caucasian (n = 476) and black (n = 248) subjects. Phenotypes measured were oxygen uptake (VO(2)), work rate, heart rate, minute ventilation, tidal volume, and blood lactate levels during maximal and submaximal [50 W and at 60 and 80% of maximal VO(2) (VO(2 max))] exercise and stroke volume and cardiac output during submaximal exercise (50 W and at 60% VO(2 max)). The ACE ID polymorphism was typed with the three-primer PCR method. Out of 216 association tests performed on 54 phenotypes in 4 groups of participants, only 11 showed significant (P values from 0.042 to 0. 0001) associations with the ACE ID polymorphism. In contrast to previous claims, in Caucasian offspring, the DD homozygotes showed a 14-38% greater increase with training in VO(2 max), VO(2) at 80% of VO(2 max), and all work rate phenotypes and a 36% greater decrease in heart rate at 50 W than did the II homozygotes. No associations were evident in Caucasian parents or black parents or offspring. Thus these data do not support the hypothesis that the ACE ID polymorphism plays a major role in cardiorespiratory endurance. PMID- 10710401 TI - Eukaryotic initiation factors and protein synthesis after resistance exercise in rats. AB - Translational control of protein synthesis depends on numerous eukaryotic initiation factors (eIFs) and we have previously shown (Am. J. Physiol. Endocrinol. Metab. 276: E721-E727, 1999) that increases in one factor, eIF2B, are associated with increases in rates of protein synthesis after resistance exercise in rats. In the present study we investigated whether the eIF4E family of initiation factors is also involved with an anabolic response to exercise. Male Sprague-Dawley rats either remained sedentary (n = 6) or performed acute resistance exercise (n = 6), and rates of protein synthesis were assessed in vivo 16 h after the last session of resistance exercise. eIF4E complexed to eIF4G (eIF4E x eIF4G), eIF4E binding protein 1 (4E-BP1) complexed to eIF4E, and phosphorylation state of eIF4E and 4E-BP1 (gamma-form) were assessed in gastrocnemius. Rates of protein synthesis were higher in exercised rats compared with sedentary rats [205 +/- 8 (SE) vs. 164 +/- 5.5 nmol phenylalanine incorporated x g muscle(-1) x h(-1), respectively; P < 0.05]. Arterial plasma insulin concentrations were not different between the two groups. A trend (P = 0.09) for an increase in eIF4E x eIF4G with exercise was noted; however, no statistically significant differences were observed in any of the components of the eIF4E family in response to resistance exercise. These new data, along with our previous report on eIF2B, suggest that the regulation of peptide chain initiation after exercise is more dependent on eIF2B than on the eIF4E system. PMID- 10710402 TI - Integrated response of the upper and lower respiratory tract of asthmatic subjects to frigid air. AB - To evaluate the influence of cold air hyperpnea on integrated upper and lower airway behavior, 22 asthmatic volunteers hyperventilated through their mouths (OHV) and noses (NHV) while pulmonary and nasal function were determined individually and in combination. In the isolated studies, OHV at a minute ventilation of 65 +/- 3 l/min lowered the 1-s forced expiratory volume (FEV(1)) 24 +/- 2% (P < 0. 001) and NHV (40 l/min) induced a 31 +/- 9% (P < 0.001) increase in nasal resistance (NR). In the combined studies, oral hyperpnea reduced the FEV(1) (DeltaFEV(1) 26 +/- 2%, P < 0.001) and evoked a significant rise in NR (DeltaNR 26 +/- 9%, P = 0.01). In contrast, NHV only affected the upper airway. NR rose 33 +/- 9% (P = 0.01), but airway caliber did not change (DeltaFEV(1) 2%, P = 0.27). The results of this investigation demonstrate that increasing the transfer of heat and water in the lower respiratory tract alters bronchial and nasal function in a linked fashion. Forcing the nose to augment its heat-exchanging activity, however, reduces nasal caliber but has no effect on the intrathoracic airways. PMID- 10710403 TI - Hemodynamic effects of periodic obstructive apneas in sedated pigs with congestive heart failure. AB - Because of similar physiological changes such as increased left ventricular (LV) afterload and sympathetic tone, an exaggerated depression in cardiac output (CO) could be expected in patients with coexisting obstructive sleep apnea and congestive heart failure (CHF). To determine cardiovascular effects and mechanisms of periodic obstructive apnea in the presence of CHF, 11 sedated and chronically instrumented pigs with CHF (rapid pacing) were tested with upper airway occlusion under room air breathing (RA), O(2) breathing (O2), and room air breathing after hexamethonium (Hex). All conditions led to large negative swings in intrathoracic pressure (-30 to -39 Torr) and hypercapnia (PCO(2) approximately 60 Torr), and RA and Hex also caused hypoxia (to approximately 42 Torr). Relative to baseline, RA increased mean arterial pressure (from 97.5 +/- 5.0 to 107.3 +/- 5.7 Torr, P < 0.01), systemic vascular resistance, LV end-diastolic pressure, and LV end-systolic length while it decreased CO (from 2.17 +/- 0.27 to 1.52 +/- 0.31 l/min, P < 0.01), stroke volume (SV; from 23.5 +/- 2.4 to 16.0 +/- 4.0 ml, P < 0.01), and LV end-diastolic length (LVEDL). O2 and Hex decreased mean arterial pressure [from 102.3 +/- 4.1 to 16.0 +/- 4.0 Torr (P < 0.01) with O2 and from 86.0 +/- 8.5 to 78.1 +/- 8.7 Torr (P < 0.05) with Hex] and blunted the reduction in CO [from 2.09 +/- 0.15 to 1.78 +/- 0.18 l/ml for O2 and from 2.91 +/- 0.43 to 2.50 +/- 0.35 l/ml for Hex (both P < 0.05)] and SV. However, the reduction in LVEDL and LV end-diastolic pressure was the same as with RA. There was no change in systemic vascular resistance and LVEDL during O2 and Hex relative to baseline. In the CHF pigs during apnea, there was an exaggerated reduction in CO and SV relative to our previously published data from normal sedated pigs under similar conditions. The primary difference between CHF (present study) and the normal animals is that, in addition to increased LV afterload, there was a decrease in LV preload in CHF contributing to SV depression not seen in normal animals. The decrease in LV preload during apneas in CHF may be related to effects of ventricular interdependence. PMID- 10710404 TI - Alveolar environment influences the metabolic and biophysical properties of exogenous surfactants. AB - Several factors have been shown to influence the efficacy of exogenous surfactant therapy in the acute respiratory distress syndrome. We investigated the effects of four different alveolar environments (control, saline-lavaged, N-nitroso-N methylurethane, and hydrochloric acid) on the metabolic and functional properties of two exogenous surfactant preparations: bovine lipid extract surfactant and recombinant surfactant-associated protein (SP) C drug product (rSPC) administered to each of these groups. The main difference between these preparations was the lack of SP-B in the rSPC. Our results demonstrated differences in the large aggregate pool sizes recovered from each of the experimental groups. We also observed differences in SP-A content, surface area cycling characteristics, and biophysical activities of these large aggregate forms after the administration of the two exogenous surfactant preparations. We conclude that the alveolar environment plays a critical role, influencing the overall efficacy of exogenous surfactant therapy. Thus further preclinical studies are warranted to investigate the specific factors within the alveolar environment that lead to the differences observed in this study. PMID- 10710405 TI - Increased expression of GLUT-4 and hexokinase in rat epitrochlearis muscles exposed to AICAR in vitro. AB - Exercise acutely stimulates muscle glucose transport and also brings about an adaptive increase in the capacity of muscle for glucose uptake by inducing increases in GLUT-4 and hexokinase.(1) Recent studies have provided evidence that activation of AMP protein kinase (AMPK) is involved in the stimulation of glucose transport by exercise. The purpose of this study was to determine whether activation of AMPK is also involved in mediating the adaptive increases in GLUT-4 and hexokinase. To this end, we examined the effect of incubating rat epitrochlearis muscles in culture medium for 18 h in the presence or absence of 5 aminoimidazole-4-carboxamide ribonucleoside (AICAR), which enters cells and is converted to the AMP analog ZMP, thus activating AMPK. Exposure of muscles to 0.5 mM AICAR in vitro for 18 h resulted in an approximately 50% increase in GLUT-4 protein and an approximately 80% increase in hexokinase. This finding provides strong evidence in support of the hypothesis that the activation of AMPK that occurs in muscle during exercise is involved in mediating the adaptive increases in GLUT-4 and hexokinase. PMID- 10710406 TI - Regional ventilation-perfusion distribution is more uniform in the prone position. AB - The arterial blood PO(2) is increased in the prone position in animals and humans because of an improvement in ventilation (VA) and perfusion (Q) matching. However, the mechanism of improved VA/Q is unknown. This experiment measured regional VA/Q heterogeneity and the correlation between VA and Q in supine and prone positions in pigs. Eight ketamine-diazepam-anesthetized, mechanically ventilated pigs were studied in supine and prone positions in random order. Regional VA and Q were measured using fluorescent-labeled aerosols and radioactive-labeled microspheres, respectively. The lungs were dried at total lung capacity and cubed into 603-967 small ( approximately 1.7-cm(3)) pieces. In the prone position the homogeneity of the ventilation distribution increased (P = 0.030) and the correlation between VA and Q increased (correlation coefficient = 0.72 +/- 0.08 and 0.82 +/- 0.06 in supine and prone positions, respectively, P = 0.03). The homogeneity of the VA/Q distribution increased in the prone position (P = 0.028). We conclude that the improvement in VA/Q matching in the prone position is secondary to increased homogeneity of the VA distribution and increased correlation of regional VA and Q. PMID- 10710407 TI - Effects of arousal and sleep state on systemic and pulmonary hemodynamics in obstructive apnea. AB - During obstructive sleep apnea (OSA), systemic (Psa) and pulmonary (Ppa) arterial pressures acutely increase after apnea termination, whereas left and right ventricular stroke volumes (SV) reach a nadir. In a canine model (n = 6), we examined the effects of arousal, parasympathetic blockade (atropine 1 mg/kg iv), and sleep state on cardiovascular responses to OSA. In the absence of arousal, SV remained constant after apnea termination, compared with a 4.4 +/- 1.7% decrease after apnea with arousal (P < 0.025). The rise in transmural Ppa was independent of arousal (4.5 +/- 1.0 vs. 4.1 +/- 1.2 mmHg with and without arousal, respectively), whereas Psa increased more after apnea termination in apneas with arousal compared with apneas without arousal. Parasympathetic blockade abolished the arousal-induced increase in Psa, indicating that arousal is associated with a vagal withdrawal of the parasympathetic tone to the heart. Rapid-eye-movement (REM) sleep blunted the increase in Psa (pre- to end-apnea: 5.6 +/- 2.3 mmHg vs. 10.3 +/- 1.6 mmHg, REM vs. non-REM, respectively, P < 0.025), but not transmural Ppa, during an obstructive apnea. We conclude that arousal and sleep state both have differential effects on the systemic and pulmonary circulation in OSA, indicating that, in patients with underlying cardiovascular disease, the hemodynamic consequences of OSA may be different for the right or the left side of the circulation. PMID- 10710408 TI - Neural and local effects of hypoxia on cardiovascular responses to obstructive apnea. AB - Obstructive sleep apnea (OSA) acutely increases systemic (Psa) and pulmonary (Ppa) arterial pressures and decreases ventricular stroke volume (SV). In this study, we used a canine model of OSA (n = 6) to examine the role of hypoxia and the autonomic nervous system (ANS) in mediating these cardiovascular responses. Hyperoxia (40% oxygen) completely blocked any increase in Ppa in response to obstructive apnea but only attenuated the increase in Psa. In contrast, after blockade of the ANS (20 mg/kg iv hexamethonium), obstructive apnea produced a decrease in Psa (-5.9 mmHg; P < 0.05) but no change in Ppa, and the fall in SV was abolished. Both the fall in Psa and the rise in Ppa that persisted after ANS blockade were abolished when apneas were induced during hyperoxia. We conclude that 1) hypoxia can account for all of the Ppa and the majority of the Psa response to obstructive apnea, 2) the ANS increases Psa but not Ppa in obstructive apnea, 3) the local effects of hypoxia associated with obstructive apnea cause vasodilation in the systemic vasculature and vasoconstriction in the pulmonary vasculature, and 4) a rise in Psa acts as an afterload to the heart and decreases SV over the course of the apnea. PMID- 10710409 TI - Iron supplementation improves endurance after training in iron-depleted, nonanemic women. AB - Our objective was to investigate the effects of iron depletion on adaptation to aerobic exercise, assessed by time to complete a 15-km cycle ergometer test. Forty-two iron-depleted (serum ferritin <16 microg/l), nonanemic (Hb >12 g/dl) women (18-33 yr old) received 100 mg of ferrous sulfate (S) or placebo (P) per day for 6 wk in a randomized, double-blind trial. Subjects trained for 30 min/day, 5 days/wk at 75-85% of maximum heart rate for the final 4 wk of the study. There were no group differences in baseline iron status or in 15-km time. Iron supplementation increased serum ferritin and decreased transferrin receptors in the S compared with the P group. The S and P groups decreased 15-km time and respiratory exchange ratio and increased work rate during the 15-km time trial after training. The decrease in 15-km time was greater in the S than in the P group (P = 0.04) and could be partially attributed to increases in serum ferritin and Hb. These results indicate that iron deficiency without anemia impairs favorable adaptation to aerobic exercise. PMID- 10710410 TI - High-volume, heavy-resistance strength training and muscle damage in young and older women. AB - To determine possible age differences in muscle damage response to strength training, ultrastructural muscle damage was assessed in seven 20- to 30-yr-old and six 65- to 75-yr-old previously sedentary women after heavy-resistance strength training (HRST). Subjects performed unilateral knee-extension exercise 3 days/wk for 9 wk. Bilateral muscle biopsies from the vastus lateralis were assessed for muscle damage via electron microscopy. HRST resulted in a 38 and 25% increase in strength in the young and older women, respectively (P < 0.05), but there were no between-group differences. In the young women, 2-4% of muscle fibers exhibited damage before and after training in both the trained and untrained legs (P = not significant). In contrast, muscle damage increased significantly after HRST, from 5 to 17% of fibers damaged (P < 0.01), in the older women in the trained leg compared with only 2 and 5% of fibers damaged in the untrained leg before and after training, respectively. The present results indicate that older women exhibit higher levels of muscle damage after chronic HRST than do young women. PMID- 10710411 TI - Transgenic animals in integrative biology: approaches and interpretations of outcome. AB - Technological innovations in methods for genetic manipulation of laboratory animals and in techniques for assessment of cardiovascular, respiratory, behavioral, and metabolic physiology in mouse models afford unprecedented opportunities for research in integrative biology. We provide here an overview of basic and advanced techniques for generation of transgenic mice and a discussion of how transgenic technology can be most advantageously applied to important physiological questions that can be addressed only within the intact organism. PMID- 10710412 TI - Form follows function: how muscle shape is regulated by work. AB - What determines the shape, size, and force output of cardiac and skeletal muscle? Chicago architect Louis Sullivan (1856-1924), father of the skyscraper, observed that "form follows function." This is as true for the structural elements of a striated muscle cell as it is for the architectural features of a building. Function is a critical evolutionary determinant, not form. To survive, the animal has evolved muscles with the capacity for dynamic responses to altered functional demand. For example, work against an increased load leads to increased mass and cross-sectional area (hypertrophy), which is directly proportional to an increased potential for force production. Thus a cell has the capacity to alter its shape as well as its volume in response to a need for altered force production. Muscle function relies primarily on an organized assembly of contractile and other sarcomeric proteins. From analysis of homogenized cells and molecular and biochemical assays, we have learned about transcription, translation, and posttranslational processes that underlie protein synthesis but still have done little in addressing the important questions of shape or regional cell growth. Skeletal muscles only grow in length as the bones grow; therefore, most studies of adult hypertrophy really only involve increased cross-sectional area. The heart chamber, however, can extend in both longitudinal and transverse directions, and cardiac cells can grow in length and width. We know little about the regulation of these directional processes that appear as a cell gets larger with hypertrophy or smaller with atrophy. This review gives a brief overview of the regulation of cell shape and the composition and aggregation of contractile proteins into filaments, the sarcomere, and myofibrils. We examine how mechanical activity regulates the turnover and exchange of contraction proteins. Finally, we suggest what kinds of experiments are needed to answer these fundamental questions about the regulation of muscle cell shape. PMID- 10710413 TI - Chronic contractile activity upregulates the proteasome system in rabbit skeletal muscle. AB - Remodeling of skeletal muscle in response to altered patterns of contractile activity is achieved, in part, by the regulated degradation of cellular proteins. The ubiquitin-proteasome system is a dominant pathway for protein degradation in eukaryotic cells. To test the role of this pathway in contraction-induced remodeling of skeletal muscle, we used a well-established model of continuous motor nerve stimulation to activate tibialis anterior (TA) muscles of New Zealand White rabbits for periods up to 28 days. Western blot analysis revealed marked and coordinated increases in protein levels of the 20S proteasome and two of its regulatory proteins, PA700 and PA28. mRNA of a representative proteasome subunit also increased coordinately in contracting muscles. Chronic contractile activity of TA also increased total proteasome activity in extracts, as measured by the hydrolysis of a proteasome-specific peptide substrate, and the total capacity of the ubiquitin-proteasome pathway, as measured by the ATP-dependent hydrolysis of an exogenous protein substrate. These results support the potential role of the ubiquitin-proteasome pathway of protein degradation in the contraction-induced remodeling of skeletal muscle. PMID- 10710414 TI - Time course evaluation of protein synthesis and glucose uptake after acute resistance exercise in rats. AB - The temporal pattern for changes in rates of protein synthesis and glucose uptake after resistance exercise, especially relative to each other, is not known. Male Sprague-Dawley rats performed acute resistance exercise (n = 7) or remained sedentary (n = 7 per group), and the following were assessed in vivo 1, 3, 6, 12 and 24 h later: rates of protein synthesis, rates of glucose uptake, phosphatidylinositol 3-kinase (PI3-kinase) activity, and p70(S6k) activity. Rates of protein synthesis in mixed gastrocnemius muscle did not increase until 12 h after exercise (e.g., at 12 h, sedentary = 138 +/- 4 vs. exercised = 178 +/- 6 nmol phenylalanine incorporated x g muscle(-1) x h(-1), mean +/- SE, P < 0.05), whereas at 6 h after exercise rates of glucose uptake were significantly elevated (sedentary = 0.18 +/- 0.020 vs. exercised = 0.38 +/- 0.024 micromol glucose 6 phosphate incorporated x kg muscle(-1) x min(-1), P < 0.05). At 24 h after exercise, rates of protein synthesis were still elevated, whereas glucose uptake had returned to basal levels. Arterial insulin concentrations were not different between groups at any time. Non-insulin-stimulated activities of PI3-kinase and p70(S6k) were higher at 6, 12, and 24 h after exercise (P < 0.05), and, generally, these occurred when rates of protein synthesis (12 and 24 h) and glucose uptake were elevated (6 and 12 but not 24 h) by exercise. These data suggest that regulators of protein synthesis and glucose uptake may respond to the same contraction-generated signals with different kinetics or that they respond to different intra- or extracellular signals that are generated by exercise. PMID- 10710415 TI - SURVEY AND SUMMARY: Saccharomyces cerevisiae basic helix-loop-helix proteins regulate diverse biological processes. AB - Basic helix-loop-helix (bHLH) proteins are among the most well studied and functionally important regulatory proteins in all eukaryotes. The HLH domain dictates dimerization to create homo- and heterodimers. Dimerization juxtaposes the basic regions of the two monomers to create a DNA interaction surface that recognizes the consensus sequence called the E-box, 5'-CANNTG-3'. Several bHLH proteins have been identified in the yeast Saccharomyces cerevisiae using traditional genetic methodologies. These proteins regulate diverse biological pathways. The completed sequence of the yeast genome, combined with novel methodologies allowing whole-genome expression studies, now offers a unique opportunity to study the function of these bHLH proteins. It is the purpose of this review to summarize the current knowledge of bHLH protein function in yeast. PMID- 10710416 TI - The C-terminal conserved domain of DNA-PKcs, missing in the SCID mouse, is required for kinase activity. AB - DNA-PKcs, the catalytic subunit of DNA-dependent protein kinase (DNA-PK), has a phosphoinositol 3-kinase (PI 3-K) domain close to its C-terminus. Cell lines derived from the SCID mouse have been utilised as a model DNA-PKcs-defective system. The SCID mutation results in truncation of DNA-Pkcs at the extreme C terminus leaving the PI 3-K domain intact. The mutated protein is expressed at low levels in most SCID cell lines, leaving open the question of whether the mutation abolishes kinase activity. Here, we show that a SCID cell line that expresses the mutant protein normally has dramatically impaired kinase activity. We estimate that the residual kinase activity typically present in SCID fibroblast cell lines is at least two orders of magnitude less than that found in control cells. Our results substantiate evidence that DNA-PKcs kinase activity is required for DSB rejoining and V(D)J recombination and show that the extreme C terminal region of DNA-PKcs, present in PI 3-K-related protein kinases but absent in bona fide PI 3 lipid kinases, is required for DNA-PKcs to function as a protein kinase. We also show that expression of mutant DNA-PKcs protein confers a growth disadvantage, providing an explanation for the lack of DNA-PKcs expression in most SCID cell lines. PMID- 10710417 TI - Characterization of an unusual tRNA-like sequence found inserted in a Neurospora retroplasmid. AB - We characterized an unusual tRNA-like sequence that had been found inserted in suppressive variants of the mitochondrial retroplasmid of Neurospora intermedia strain Varkud. We previously identified two forms of the tRNA-like sequence, one of 64 nt (TRL-64) and the other of 78 nt (TRL-78) containing a 14-nt internal insertion in the anticodon stem at a position expected for a nuclear tRNA intron. Here, we show that TRL-78 is encoded in Varkud mitochondrial (mt)DNA within a 7 kb sequence that is not present in Neurospora crassa wild-type 74 A mtDNA. This 7 kb insertion also contains a perfectly duplicated tRNA(Trp)gene, segments of several mitochondrial plasmids and numerous GC-rich palindromic sequences that are repeated elsewhere in the mtDNA. The mtDNA-encoded copy of TRL-78 is transcribed and apparently undergoes 5'- and 3'-end processing and 3' nucleotide addition by tRNA nucleotidyl transferase to yield a discrete tRNA-sized molecule. However, the 14 nt intron-like sequence in TRL-78, which is missing in the TRL-64 form, is not spliced detectably in vivo or in vitro. Our results show that TRL-78 is an unusual tRNA-like species that could be incorporated into suppressive retroplasmids by the same reverse transcription mechanism used to incorporate mt tRNAs. The tRNA-like sequence may have been derived from an intron-containing nuclear tRNA gene or it may serve some function, like mtRNA. Our results suggest that mt tRNAs or tRNA-like species may be integrated into mtDNA via reverse transcription, analogous to SINE elements in animal cells. PMID- 10710418 TI - Cloning and characterization of the 5'-upstream sequence governing the cell cycle dependent transcription of mouse DNA polymerase alpha 68 kDa subunit gene. AB - We have isolated the genomic DNA fragment spanning the 5-end and the first four exons encoding the 68 kDa subunit (p68) of the mouse DNA polymerase alpha-primase complex [corrected]. The p68 promoter region lacks TATA and CAAT boxes, but contains a GC-rich sequence, two palindrome sequences and two putative E2F binding sites [corrected]. A series of transient expression assays using a luciferase reporter gene indicated that a region from nucleotide position -89 to 30 (-89/-30) with respect to the transcription initiation site is crucial for basal transcription of the p68 gene in proliferating NIH 3T3 cells. In particular, part of the GC-rich sequence (-57/-46) and the palindrome (-81/-62) elements were necessary for promoter activity, both of which share homology with the E-box sequence. Gel mobility shift assays using NIH 3T3 nuclear extracts revealed that the upstream stimulatory factor, known as an E-box-binding protein, binds to these sites. Moreover, we observed binding of E2F to two sites near the transcription initiation site (-11/-3 and +9/+16). A transient luciferase expression assay using synchronized NIH 3T3 cells in G(0)phase revealed that these E2F sites are essential for transcription induction of the p68 gene after serum stimulation, but are dispensable for basal transcription. These results indicate that growth-dependent regulation of transcription of the mouse p68 and p180 genes is mediated by a common factor, E2F; however, basal transcription of the genes, interestingly, is regulated by different transcription factors. PMID- 10710419 TI - Interruption of the fragile X syndrome expanded sequence d(CGG)(n) by interspersed d(AGG) trinucleotides diminishes the formation and stability of d(CGG)(n) tetrahelical structures. AB - Fragile X syndrome is caused by expansion of a d(CGG) trinucleotide repeat sequence in the 5' untranslated region of the first exon of the FMR1 gene. Repeat expansion is thought to be instigated by formation of d(CGG)(n)secondary structures. Stable FMR1 d(CGG)(n)runs in normal individuals consist of 6-52 d(CGG) repeats that are interrupted every 9-11 triplets by a single d(AGG) trinucleotide. By contrast, individuals having fragile X syndrome premutation or full mutation present >54-200 or >200-2000 monotonous d(CGG) repeats, respectively. Here we show that the presence of interspersed d(AGG) triplets diminished in vitro formation of bimolecular tetrahelical structures of d(CGG)(18)oligomers. Tetraplex structures formed by d(CGG)(n)oligomers containing d(AGG) interspersions had lower thermal stability. In addition, tetraplex structures of d(CGG)(18)oligomers interspersed by d(AGG) triplets were unwound by human Werner syndrome DNA helicase at rates and to an extent that exceeded the unwinding of tetraplex form consisting of monotonous d(CGG)(18). Diminished formation and stability of tetraplex structures of d(AGG)-containing FMR1 d(CGG)(2-50)tracts might restrict their expansion in normal individuals. PMID- 10710420 TI - A missense mutation in the nuclear gene coding for the mitochondrial aspartyl tRNA synthetase suppresses a mitochondrial tRNA(Asp) mutation. AB - The nuclear suppressor allele NSM3 in strain FF1210-6C/170-E22 (E22), which suppresses a mutation of the yeast mitochondrial tRNA(Asp)gene in Saccharomyces cerevisiae, was cloned and identified. To isolate the NSM3 allele, a genomic DNA library using the vector YEp13 was constructed from strain E22. Nine YEp13 recombinant plasmids were isolated and shown to suppress the mutation in the mitochondrial tRNA(Asp)gene. These nine plasmids carry a common 4. 5-kb chromosomal DNA fragment which contains an open reading frame coding for yeast mitochondrial aspartyl-tRNA synthetase (AspRS) on the basis of its sequence identity to the MSD1 gene. The comparison of NSM3 DNA sequences between the suppressor and the wild-type version, cloned from the parental strain FF1210 6C/170, revealed a G to A transition that causes the replacement of amino acid serine (AGU) by an asparagine (AAU) at position 388. In experiments switching restriction fragments between the wild type and suppressor versions of the NSM3 gene, the rescue of respiratory deficiency was demonstrated only when the substitution was present in the construct. We conclude that the base substitution causes the respiratory rescue and discuss the possible mechanism as one which enhances interaction between the mutated tRNA(Asp)and the suppressor version of AspRS. PMID- 10710421 TI - Molecular characterisation of two paralogous SPO11 homologues in Arabidopsis thaliana. AB - The Spo11 protein of yeast has been found to be covalently bound to double-strand breaks in meiosis, demonstrating a unique role of the protein in the formation of these breaks. Homologues of the SPO11 gene have been found in various eukaryotes, indicating that the machinery involved in meiotic recombination is conserved in eukaryotes. Here we report on SPO11 homologues in plants. In contrast to what is known from other eukaryotes, Arabidopsis thaliana carries in its genome at least two SPO11 homologues, AtSPO11-1 and AtSPO11-2. Both genes are not more closely related to each other than to other eukaryotic SPO11 homologues, indicating that they did not arise via a recent duplication event during higher plant evolution. For both genes three different poly-adenylation sites were found. AtSPO11-1 is expressed not only in generative but also to a lesser extent in somatic tissues. We were able to detect in different organs various AtSPO11-1 cDNAs in which introns were differently spliced-a surprising phenomenon also reported for SPO11 homologues in mammals. In the case of AtSPO11-2 we found that the 3' end of the mRNA is overlapping with a mRNA produced by a gene located in inverse orientation next to it. This points to a possible antisense regulation mechanism. Our findings hint to the intriguing possibility that, at least for plants, Spo11-like proteins might have more and possibly other biological functions than originally anticipated for yeast. PMID- 10710422 TI - In vitro DNA synthesis opposite oxazolone and repair of this DNA damage using modified oligonucleotides. AB - Emphasis was placed in this work on the assessment of biological features of 2,2,4-triaminooxazolone, a major one-electron and(. )OH-mediated oxidation product of guanine. For this purpose, two oligonucleotides that contain a unique oxazolone residue were synthesized. Herein we report the mutagenic potential of oxazolone during in vitro DNA synthesis and its behavior towards DNA repair enzymes. Nucleotide insertion opposite oxazolone, catalyzed by Klenow fragment exo(-)and Taq polymerase indicates that the oxazolone lesion induces mainly dAMP insertion. This suggests that the formation of oxazolone in DNA may lead to G-->T transversions. On the other hand, oxazolone represents a blocking lesion when DNA synthesis is performed with DNA polymerase beta. Interestingly, DNA repair experiments carried out with formamidopyrimidine DNA N -glycosylase (Fpg) and endonuclease III (endo III) show that oxazolone is a substrate for both enzymes. Values of k (cat)/ K (m)for the Fpg-mediated removal of oxidative guanine lesions revealed that 8-oxo-7,8-dihydroguanine is only a slightly better substrate than oxazolone. In the case of endo III-mediated cleavage of modified bases, the present results suggest that oxazolone is a better substrate than 5-OHC, an oxidized pyrimidine base. Finally, MALDI-TOF-MS analysis of the DNA fragments released upon digestion of an oxazolone-containing oligonucleotide by Fpg gave insights into the enzymatic mechanism of oligonucleotide cleavage. PMID- 10710423 TI - Identification of two human nuclear proteins that recognise the cytosine-rich strand of human telomeres in vitro. AB - Most studies on the structure of DNA in telomeres have been dedicated to the double-stranded region or the guanosine-rich strand and consequently little is known about the factors that may bind to the telomere cytosine-rich (C-rich) strand. This led us to investigate whether proteins exist that can recognise C rich sequences. We have isolated several nuclear factors from human cell extracts that specifically bind the C-rich strand of vertebrate telomeres [namely a d(CCCTAA)(n)repeat] with high affinity and bind double-stranded telomeric DNA with a 100xreduced affinity. A biochemical assay allowed us to characterise four proteins of apparent molecular weights 66-64, 45 and 35 kDa, respectively. To identify these polypeptides we screened alambdagt11-based cDNA expression library, obtained from human HeLa cells using a radiolabelled telomeric oligonucleotide as a probe. Two clones were purified and sequenced: the first corresponded to the hnRNP K protein and the second to the ASF/SF2 splicing factor. Confirmation of the screening results was obtained with recombinant proteins, both of which bind to the human telomeric C-rich strand in vitro. PMID- 10710424 TI - Scp160p, a multiple KH-domain protein, is a component of mRNP complexes in yeast. AB - Scp160p is a 160 kDa protein in the yeast Saccharomyces cerevisiae that contains 14 repeats of the hnRNP K-homology (KH) domain, and demonstrates significant sequence homology to a family of proteins collectively known as vigilins. As a first step towards defining the function of Scp160p, we have characterized the subcellular distribution and in vivo interactions of this protein. Using sucrose gradient fractionation studies we have demonstrated that Scp160p in cytoplasmic lysates is predominantly associated with polyribosomes. Furthermore, we have found that Scp160p is released from polyribosomes by EDTA in the form of a large complex of> or =1300 kDa that is sensitive both to RNase and NaCl. Using affinity chromatography to isolate these complexes, we have identified two protein components other than Scp160p: poly(A) binding protein, Pab1p, and Bfr1p. The presence of Pab1p confirms these complexes to be mRNPs. The presence of Bfr1p is intriguing because the null phenotype for this gene is essentially the same as that reported for scp160 -null cells: increased cell size and aberrant DNA content. These results demonstrate that Scp160p associates with polyribosome bound mRNP complexes in vivo, implicating a role for this protein in one or more levels of mRNA metabolism in yeast. PMID- 10710425 TI - Detection of N-H...N hydrogen bonding in RNA via scalar couplings in the absence of observable imino proton resonances. AB - Hydrogen bond networks stabilize RNA secondary and tertiary structure and are thus essentially important for protein recognition. During structure refinements using either NMR or X-ray techniques, hydrogen bonds were usually inferred indirectly from the proximity of donor and acceptor functional groups. Recently, quantitative heteronuclear J(N,N)-HNN COSY NMR experiments were introduced that allowed the direct identification of donor and acceptor nitrogen atoms involved in hydrogen bonds. However, protons involved in base pairing interactions in nucleic acids are often not observable due to exchange processes. The application of a modified quantitative J(N,N)-HNN COSY pulse scheme permits observation of(2h)J(N,N) couplings via non-exchangeable protons. This approach allowed the unambiguous identification of the A27.U23 reverse Hoogsteen base pair involved in a U-A.U base triple in the HIV-2 transactivation response element-argininamide complex. Despite a wealth of NOE information, direct evidence for this interaction was lacking due to the rapid exchange of the U23 imino proton. The ability to directly observe hydrogen bonds, even in D(2)O and in the presence of rapid exchange, should facilitate structural studies of RNA. PMID- 10710426 TI - Positive and negative mutant selection in the human histone hairpin-binding protein using the yeast three-hybrid system. AB - We have used the yeast three-hybrid system in a positive selection for mutants of the human histone hairpin-binding protein (HBP) capable of interacting with non canonical hairpins and in a negative selection for loss-of-binding mutants. Interestingly, all mutations from the positive selection are located in the N- and C-terminal regions flanking a minimal RNA-binding domain (RBD) previously defined between amino acids 126 and 198. Further, in vitro binding studies demonstrate that the RBD, which shows no obvious similarity to other RNA-binding motifs, has a relaxed sequence specificity compared to full-length HBP, allowing it to bind to mutant hairpin RNAs not normally found in histone genes. These findings indicate that the sequences flanking the RBD are important for restricting binding to the highly conserved histone hairpin structure. Among the loss-of-binding mutations, about half are nonsense mutations distributed throughout the N-terminal part and the RBD whereas the other half are missense mutations restricted to the RBD. Whereas the nonsense mutations permit a more precise definition of the C-terminal border of the RBD, the missense mutations identify critical residues for RNA binding within the RBD. PMID- 10710427 TI - Computational identification of cis-acting elements affecting post transcriptional control of gene expression in Saccharomyces cerevisiae. AB - Understanding the regulation of gene expression requires the identification of cis -acting control elements that modulate gene function. The recent availability of complete genome sequences and profiles of mRNA expression has facilitated the development and utilization of computational methods to identify discrete regulatory elements. We have developed an oligomer counting method that identifies sequences that occur significantly more often in a group of interest relative to other genes in the genome. The use of a second parameter, which measures the frequency of oligomers within the group of interest, allows the detection of false positive signals caused by very infrequent oligomers that would otherwise appear as significant. Applying this method to gene groups that have a common expression pattern or shared function should identify oligomers that comprise cis -acting control elements. As a test of this method, we applied this approach to a set of intron-containing yeast genes, where we easily identified the known splicing signals as control elements. We have used this training set to examine how this method is affected by the length of the oligomer examined, as well as the size and composition of the gene group. These simulations allowed us to identify rules for selecting groups of genes to analyze. Finally, application of this method to nuclear genes encoding proteins targeted to the mitochondria identified a new putative cis -acting sequence in the 3'-untranslated region of this family of genes, which may play a role in mRNA localization or the regulation of mRNA stability or translation. PMID- 10710428 TI - Kinetic analysis of high-mobility-group proteins HMG-1 and HMG-I/Y binding to cholesterol-tagged DNA on a supported lipid monolayer. AB - High-mobility-group proteins HMG-1 and HMG-I/Y bind to multiple sites within a 268 bp A/T-rich enhancer element of the pea plastocyanin gene ( PetE ). Within a 31 bp region of the enhancer, the binding site for HMG-1 overlaps with the binding site for HMG-I/Y. The kinetics of binding and the affinities of HMG-1 and HMG-I/Y for the 31 bp DNA were determined using surface plasmon resonance. Due to very high non-specific interactions of the HMG proteins with a carboxymethyl dextran matrix, a novel method using a cholesterol tag to anchor the DNA in a supported lipid monolayer on a thin gold film was devised. The phosphatidylcholine monolayer produced a surface that reduced background interactions to a minimum and permitted the measurement of highly reproducible protein-DNA interactions. The association rate constant ( k (a)) of HMG-I/Y with the 31 bp DNA was approximately 5-fold higher than the rate constant for HMG-1, whereas the dissociation constant ( K (D)) for HMG-I/Y (3.1 nM) was approximately 7-fold lower than that for HMG-1 (20.1 nM). This suggests that HMG-I/Y should bind preferentially at the overlapping binding site within this region of the PetE enhancer. PMID- 10710429 TI - Initiation of translation by non-AUG codons in human T-cell lymphotropic virus type I mRNA encoding both Rex and Tax regulatory proteins. AB - Human T-cell lymphotropic virus type I (HTLV-I) double-spliced mRNA exhibits two GUG and two CUG codons upstream to, and in frame with, the sequences encoding Rex and Tax regulatory proteins, respectively. To verify whether these GUG and CUG codons could be used as additional initiation codons of translation, two chimeric constructs were built for directing the synthesis of either Rex-CAT or Tax-CAT fusion proteins. In both cases, the CAT reporter sequence was inserted after the Tax AUG codon and in frame with either the Rex or Tax AUG codon. Under transient expression of these constructs, other proteins of higher molecular mass were synthesized in addition to the expected Rex-CAT and Tax-CAT proteins. The potential non-AUG initiation codons were exchanged for either an AUG codon or a non-initiation codon. This allowed us to demonstrate that the two GUG codons in frame with the Rex coding sequence, and only the second CUG in frame with the Tax coding sequence, were used as additional initiation codons. In HTLV-I infected cells, two Rex and one Tax additional proteins were detected that exhibited molecular mass compatible with the use of the two GUG and the second CUG as additional initiation codons of translation. Comparison of the HTLV-I proviral DNA sequence with that of other HTLV-related retroviruses revealed a striking conservation of the three non-AUG initiation codons, strongly suggesting their use for the synthesis of additional Rex and Tax proteins. PMID- 10710430 TI - NotI clones in the analysis of the human genome. AB - Not I linking clones contain sequences flanking Not I recognition sites and were previously shown to be tightly associated with CpG islands and genes. To directly assess the value of Not I clones in genome research, high density grids with 50 000 Not I linking clones originating from six representative Not I linking libraries were constructed. Altogether, these libraries contained nearly 100 times the total number of Not I sites in the human genome. A total of 3437 sequences flanking Not I sites were generated. Analysis of 3265 unique sequences demonstrated that 51% of the clones displayed significant protein similarity to SWISSPROT and TREMBL database proteins based on MSPcrunch filtering with stringent parameters. Of the 3265 sequences, 1868 (57.2%) were new sequences, not present in the EMBL and EST databases (similarity < or =90%). Among these new sequences, 795 (24.3%) showed similarity to known proteins and 712 (21.8%) displayed an identity of >75% at the nucleotide level to sequences from EMBL or EST databases. The remaining 361 (11.1%) sequences were completely new, i.e. <75% identical. The work also showed tight, specific association of Not I sites with the first exon and suggest that the so-called 3' ESTs can actually be generated from 5'-ends of genes that contain Not I sites in their first exon. PMID- 10710431 TI - 2-Hydroxy-dATP is incorporated opposite G by Escherichia coli DNA polymerase III resulting in high mutagenicity. AB - Four kinds of oxidatively damaged DNA precursors, 8-hydroxydeoxyguanosine 5' triphosphate (8-OH-dGTP), 2-hydroxydeoxyadenosine 5'-triphosphate (2-OH-dATP), 5 hydroxydeoxycytidine 5'-triphosphate (5-OH-dCTP) and 5-formyldeoxyuridine 5' triphosphate (5-CHO-dUTP), were employed in in vitro gap-filling reactions of the supF gene conducted by the Escherichia coli DNA polymerase III holoenzyme, and these treated DNAs were transfected into various E.coli strains. When the manipulated DNAs were transfected into the repair-proficient strain, supF mutants were obtained much more frequently by the purine nucleotides than by the pyrimidine nucleotides (2-OH-dATP > 8-OH-dGTP >> 5-OH-dCTP approximately 5-CHO dUTP). This result is in contrast to our previous observation that these four oxidatively damaged nucleotides induce chromosomal gene mutations with similar frequencies when incorporated directly into E.coli. 2-OH-dATP elicited G-->T transversions, indicating the formation of G*2-OH-dATP pairs. These results demonstrate that 2-OH-dATP was highly mutagenic in this assay system containing the in vitro DNA synthesis by the E.coli replicative DNA polymerase, in addition to in the in vivo assay system reported previously. Slight increases in the mutant frequencies were observed when alkA (for 8-OH-dGTP and 2-OH-dATP) and mutY (for 2-OH-dATP) strains were used as hosts. This is the first report that clearly shows the formation of G*2-OH-dATP pairs. PMID- 10710432 TI - Human RecQ5beta, a large isomer of RecQ5 DNA helicase, localizes in the nucleoplasm and interacts with topoisomerases 3alpha and 3beta. AB - The RecQ helicase superfamily has been implicated in DNA repair and recombination. At least five human RecQ-related genes exist: RecQ1, BLM, WRN, RecQ4 and RecQ5. Mutations in BLM, WRN and RecQ4 are associated with Bloom, Werner and Rothmund-Thomson syndromes, respectively, involving a predisposition to malignancies and a cellular phenotype that includes increased chromosome instability. RecQ5 is small, containing only a core part of the RecQ helicase, but three isomer transcripts code for small RecQ5alpha (corresponding to the original RecQ5 with 410 amino acids), new large RecQ5beta (991 amino acids) and small RecQ5gamma (435 amino acids) proteins that contain the core helicase motifs. By determining the genomic structure, we found that the three isoforms are generated by differential splicing from the RecQ5 gene that contains at least 19 exons. Northern blot analysis using a RecQ5beta-specific probe indicates that RecQ5beta mRNA is expressed strongly in the testis. Immunocytochemical staining of three N-terminally tagged RecQ5 isomers expressed in 293EBNA cells showed that RecQ5beta migrates to the nucleus and exists exclusively in the nucleoplasm, while the small RecQ5alpha and RecQ5gamma proteins stay in the cytoplasm. Immunoprecipitation and an extended cytochemical experiment suggested that the nucleoplasmic RecQ5beta, like yeast Sgs1 DNA helicase, binds to topoisomerases 3alpha and 3beta, but not to topoisomerase 1. These results predict that RecQ5beta may have an important role in DNA metabolism and may also be related to a distinct genetic disease. PMID- 10710433 TI - In vivo, high-resolution analysis of yeast and mammalian RNA-protein interactions, RNA structure, RNA splicing and ribozyme cleavage by use of terminal transferase-dependent PCR. AB - We have investigated the analysis of RNA by use of terminal transferase-dependent PCR (TDPCR), a procedure previously used for the analysis of DNA and chromatin [J. Komura and A.D.Riggs, Nucleic Acids Res.,26, 1807-1811 (1998)]. When preceded by reverse transcription (RT), TDPCR provides an extremely sensitive, versatile, quantitative and nucleotide-level assay for detecting RNA lesions or structures that block primer extension during the RT step. The procedure is: (i) RT using a gene-specific oligonucleotide; (ii) ribo-tailing of the single-stranded cDNA product by use of terminal deoxy-nucleotidyl transferase; (iii) ligation of a DNA linker to the tailed cDNA by use of T4 DNA ligase; and (iv) PCR using a nested, gene-specific primer and a linker-specific primer. This procedure combines the versatility of a primer extension assay with nucleotide-level resolution, the specificity of nested primers and the sensitivity of PCR. Band patterns obtained are reproducible and quantifiable. We successfully used the technique for the study of yeast RNA structure, splicing intermediates and ribozyme cleavage. Also, in vivo footprint experiments, using mammalian cells and RNase T1, revealed the binding of iron-responsive element binding protein to iron responsive elements in the mRNAs of transferrin receptor and ferritin H-chain. PMID- 10710434 TI - Rapid, high fidelity analysis of simple sequence repeats on an electronically active DNA microchip. AB - We describe a method for the discrimination of short tandem repeat (STR) alleles based on active microarray hybridization. An essential factor in this method is electronic hybridization of the target DNA, at high stringency, in <5 min. High stringency is critical to avoid slippage of hybrids along repeat tracts at allele specific test sites in the array. These conditions are attainable only with hybridization kinetics realized by electronic concentration of DNA. A sandwich hybrid is assembled, in which proper base stacking of juxtaposed terminal nucleotides results in a thermodynamically favored complex. The increased stability of this complex relative to non-stacked termini and/or base pair mismatches is used to determine the identification of STR alleles. This method is capable of simultaneous and precise identification of alleles containing different numbers of repeats, as well as mutations within these repeats. Given the throughput capabilities of microarrays our system has the potential to enhance the use of microsatellites in forensic criminology, diagnostics and genetic mapping. PMID- 10710435 TI - A method for quantification of absolute amounts of nucleic acids by (RT)-PCR and a new mathematical model for data analysis. AB - Accurate quantification of nucleic acids by competitive (RT)-PCR requires a valid internal standard, a reference for data normalization and an adequate mathematical model for data analysis. We report here an effective procedure for the generation of homologous RNA internal standards and a strategy for synthesizing and using a reference target RNA in quantification of absolute amounts of nucleic acids. Further, a new mathematical model describing the general kinetic features of competitive PCR was developed. The model extends the validity of quantitative competitive (RT)-PCR beyond the exponential phase. The new method eliminates the errors arising from different amplification efficiencies of the co-amplified sequences and from heteroduplex formation in the system. The high accuracy (relative error <2%) is comparable to the recently developed real time detection 5'-nuclease PCR. Also, corresponding computer software has been devised for practical data analysis. PMID- 10710436 TI - A new approach for the identification and cloning of genes: the pBACwich system using Cre/lox site-specific recombination. AB - With current plant transformation methods ( Agrobacterium, biolistics and protoplast fusion), insertion of DNA into the genome occurs randomly and in many instances at multiple sites. Associated position effects, copy number differences and multigene interactions can make gene expression experiments difficult to interpret and plant phenotypes less predictable. An alternative approach to random integration of large DNA fragments into plants is to utilize one of several site-specific recombination (SSR) systems, such as Cre/ lox. Cre has been shown in numerous instances to mediate lox site-specific recombination in animal and plant cells. By incorporating the Cre/ lox SSR system into a bacterial artificial chromosome (BAC) vector, a more precise evaluation of large DNA inserts for genetic complementation should be possible. Site-specific insertion of DNA into predefined sites in the genome may eliminate unwanted 'position effects' caused by the random integration of exogenously introduced DNA. In an effort to make the Cre/ lox system an effective tool for site-directed integration of large DNAs, we constructed and tested a new vector potentially capable of integrating large DNA inserts into plant and fungal genomes. In this study, we present the construction of a new BAC vector, pBACwich, for the system and the use of this vector to demonstrate SSR of large DNA inserts (up to 230 kb) into plant and fungal genomes. PMID- 10710437 TI - RNA-binding analyses of HuC and HuD with the VEGF and c-myc 3'-untranslated regions using a novel ELISA-based assay. AB - Human members of the ELAV family, referred to as ELAV-like proteins (ELPs), include HuC, HuD, Hel-N1 and HuR. These proteins bind to AU-rich elements in the 3'-untranslated regions (3'-UTRs) of many growth-related mRNAs, including c-myc and VEGF, and may participate in regulating the stability of these transcripts. Here, I have developed an enzyme-linked immunosorbent assay (ELISA) which can rapidly assess the RNA-protein-binding properties of ELPs. With this assay, I demonstrate that HuC and HuD bind to the VEGF 3'-UTR regulatory segment (VRS) and to the c- myc 3'-UTR in a specific and concentration-dependent pattern, with both proteins showing a greater affinity for the VRS. Further analysis of the VRS indicated that the binding affinity was greater for the 3'-end where the majority of AU motifs reside. Binding to the VRS could be competed by both proteins as well as a poly(U) ribohomopolymer. The binding could not be competed by other ribohomopolymers or serum from patients with high titer anti-HuD antibodies. In summary, this assay provides a rapid analysis of ELP-RNA binding which can be utilized for further characterization of RNA-binding properties and for identification of competitor molecules for in vivo functional analysis of ELPs. PMID- 10710438 TI - Signal amplification through nucleotide extension and excision on a dendritic DNA platform. AB - Techniques that provide strong signal amplification are useful in diagnostic applications, especially in detecting low concentrations of non-amplifiable target molecules. A versatile and strong signal amplification method based on activities of a DNA polymerase to generate high concentrations of pyrophosphate (PPi) is described. The generation of PPi is catalyzed by nucleotide extension and excision activities of a DNA polymerase on an oligonucleotide cassette. The signal is generated upon enzymatic conversion of PPi to ATP and ATP levels subsequently detected with firefly luciferase. Bioluminescence produced by an oligonucleotide cassette consisting of just two polymerase reaction sites is sufficient to detect them at low attomole levels. The attachment of a large number of these oligonucleotide cassettes to DNA dendrimers enabled the detection of such poly-valent substrate molecules at low zeptomole (10(-21)mol) concentrations. The extent of signal amplification obtained with dendrimer substrates is comparable to exponential target amplifications provided by nucleic acid amplification methods. The attachment of such PPi-generating dendritic DNA platforms to ligands that mediate target recognition would potentially permit detection of extremely low concentrations of analytes in diagnostic assays. PMID- 10710439 TI - Octamer-primed sequencing technology: development of primer identification software. AB - Octamer sequencing technology (OST) is a primer-directed sequencing strategy in which an individual octamer primer is selected from a pre-synthesized octamer primer library and used to sequence a DNA fragment. However, selecting candidate primers from such a library is time consuming and can be a bottleneck in the sequencing process. To accelerate the sequencing process and to obtain high quality sequencing data, a computer program, electronic OST or eOST, was developed to automatically identify candidate primers from an octamer primer library. eOST integrates the base calling software PHRED to provide a quality assessment for target sequences and identifies potential primer binding sites located within a high quality target region. To increase the sequencing success rate, eOST includes a simple dynamic folding algorithm to automatically calculate the free energy and predict the secondary structure within the template in the vicinity of the octamer-binding site. Several parameters were found to be important, including base quality threshold, the window size of the template sequence segment, and the threshold [Delta] G value. OST, coupled with the eOST software, can be used to sequence short DNA fragments or in the finishing assembly stage of large-scale sequencing of genomic DNA. PMID- 10710440 TI - Locus-specific contig assembly in highly-duplicated genomes, using the BAC-RF method. AB - Polyploidy, the presence of multiple sets of chromosomes that are similar but not identical, complicates both chromosome walking and assembly of sequence-ready contigs for many plant taxa including a large number of economically-significant crops. Traditional 'dot-blot hybridization' or PCR-based assays for identifying BAC clones corresponding to a mapped DNA landmark usually do not provide sufficient information to distinguish between allelic and non-allelic loci. A restriction fragment matching method using pools of BAC DNA in combination with dot-blots reveals the locus specificity of individual BACs that correspond to multi-locus DNA probes, in a manner that can efficiently be applied on a large scale. This approach also provides an alternative means of mapping DNA loci that exploits many advantages of 'radiation hybrid' mapping in taxa for which such hybrids are not available. The BAC-RF method is a practical and reliable approach for using high-density RFLP maps to anchor sequence-ready BAC contigs in highly duplicated genomes, provides an alternative to high-density robotic gridding for screening BAC libraries when the necessary equipment is not available, and permits the expedient isolation of individual members of multigene or repetitive DNA families for a wide range of genetic and evolutionary investigations. PMID- 10710441 TI - Phage display of ScFv peptides recognizing the thymidine(6-4)thymidine photoproduct. AB - Solar ultraviolet (UV) radiation induces DNA photoproducts in skin cells and is the predominant cause of human skin cancers. To understand human susceptibility to skin cancer and to facilitate the development of prevention measures, highly specific reagents to detect and quantitate UV-induced DNA adducts in human skin will be needed. One approach towards this end is the use of monoclonal antibody based molecular dosimetry methods. To facilitate the development of photoproduct specific antibody reagents we have: (i) cloned and sequenced a single chain variable fragment (ScFv) gene coding for one such high affinity monoclonal antibody, [alpha]UVssDNA-1 (mAb C3B6), recognizing the thymidine(6-4)thymidine photoproduct; (ii) expressed and displayed the cloned ScFv gene on the surface of phage; (iii) selected functional recombinant phage by panning; (iv) purified the ScFv peptide; (v) shown that the purified ScFv peptide binds to UV-irradiated polythymidylic acid but not unirradiated polythymidylic acid. This is the first demonstration of the use of phage display to select a ScFv recognizing DNA damage. In addition, this is the initial step towards immortalizing the antibody gene for genetic manipulation, structure-function studies and application to human investigations. PMID- 10710442 TI - Use of terminal transferase-dependent antisense RNA amplification to determine the transcription start site of the Snrpn gene in individual neurons. AB - We describe here a very sensitive technique for RNA structure analysis and the determination of transcription start sites and demonstrate its use for mapping the start site of the imprinted Snrpn gene in individual hippocampal neurons. The method is adapted from reverse transcription-terminal transferase-dependent PCR (RT-TDPCR) to include amplification of the antisense sequence by in vitro transcription just prior to the final PCR step. The method should be useful for analysis of all genes for which variation in promoter usage and/or differences in RNA secondary structure may be specific to a given cell type or developmental stage. PMID- 10710443 TI - Signal-exon trap: a novel method for the identification of signal sequences from genomic DNA. AB - We describe a genomic DNA-based signal sequence trap method, signal-exon trap (SET), for the identification of genes encoding secreted and membrane-bound proteins. SET is based on the coupling of an exon trap to the translation of captured exons, which allows screening of the exon-encoded polypeptides for signal peptide function. Since most signal sequences are expected to be located in the 5'-terminal exons of genes, we first demonstrate that trapping of these exons is feasible. To test the applicability of SET for the screening of complex genomic DNA, we evaluated two critical features of the method. Specificity was assessed by the analysis of random genomic DNA and efficiency was demonstrated by screening a 425 kb YAC known to contain the genes of four secretory or membrane bound proteins. All trapped clones contained a translation initiation signal followed by a hydrophobic stretch of amino acids representing either a known signal peptide, transmembrane domain or novel sequence. Our results suggest that SET is a potentially useful method for the isolation of signal sequence containing genes and may find application in the discovery of novel members of known secretory gene clusters, as well as in other positional cloning approaches. PMID- 10710444 TI - Identification of differentially expressed genes from limited amounts of RNA. AB - The identification of cellular RNA expression profiles by differential display (DD) involves the visualization of RT-PCR products from the RNA. Traditionally, DD protocols require 200-500 ng RNA for each RT reaction. Thus, the limiting factor in DD is the amount of RNA available and the sensitivity of the RT reaction. By replacing the type of reverse transcriptase in our method, the sensitivity of DD increased up to 100-fold. Very significantly, the cDNA species obtained are higher in molecular weight, increasing the chances of detection of differential display genes with less background bands. The false positives and background in general also decreased due to the utilization of Taq polymerase antibody to facilitated DNA synthesis in the PCR reaction step. The reverse transcriptases described here may have a greater priming capacity as well as strong processivity which would explain the higher sensitivity accomplished in comparison to more standard reverse transcriptases. Additionally, the application of a more sensitive DD to samples when the amount of RNA is limited would be highly recommended. PMID- 10710445 TI - Oral lichen planus and oral cancer: is there enough epidemiologic evidence? PMID- 10710447 TI - Let us endeavor to say what we mean. PMID- 10710448 TI - Alveolar ridge augmentation with titanium mesh and autogenous bone. PMID- 10710449 TI - Effect of arthrocentesis and hydraulic distension on the temporomandibular joint disk position. AB - OBJECTIVE: Recent studies have suggested arthrocentesis and hydraulic distension as an effective treatment modality in patients demonstrating clinical findings consistent with the diagnosis of disk displacement without reduction, normal range of motion thereby being restored and pain of the temporomandibular joint reduced. In view of the fact that only a few studies have been performed to verify the biologic concept of disk displacement without reduction as a diagnostic and therapeutic approach in patients with "closed-lock" symptoms, the purpose of this study was to investigate whether temporomandibular joint-related variable disk position might be linked to cessation of related signs and symptoms associated with the performance of arthrocentesis and hydraulic distension. STUDY DESIGN: The study compared 15 patients, each of whom was assigned a clinical unilateral temporomandibular joint-related diagnosis of internal derangement (ID) type III (disk displacement without reduction) in combination with capsulitis/synovitis. Clinical diagnoses were made according to the Clinical Diagnostic Criteria for Temporomandibular Disorders. Bilateral sagittal and coronal magnetic resonance images were obtained immediately preoperatively and at 2-month follow-up to establish the presence or absence of associated types of ID. Temporomandibular joint-related pain, level of function, and mandibular range of motion were assessed preoperatively and the data were compared with the respective 2-month follow-up findings. RESULTS: Comparison of the pretreatment temporomandibular joint side-related data revealed the temporomandibular joint side with an ID-III in combination with capsulitis/synovitis to be associated with significantly more magnetic resonance imaging diagnoses of ID than of an absence of ID (P <.001) and with significantly more disk displacement without reduction than disk displacement with reduction (P <.001). At the 2-month follow up, clinical evaluation showed a significant reduction in temporomandibular joint related pain during function (P <.001), a significant reduction in the prevalence of temporomandibular joint-related diagnoses of capsulitis/synovitis (P <.001) and ID-III (P <.01), and a significant increase in mandibular range of motion (P <.01). There was no change in the prevalence rates of associated temporomandibular joint side-related IDs. CONCLUSIONS: The results confirm the concept of disk displacement as a diagnostic approach but not as a therapeutic approach for patients presenting with signs and symptoms of unilateral ID-III in combination with capsulitis/synovitis. In terms of clinical decision-making in temporomandibular disorder-related instances of ID, magnetic resonance imaging may be used as a diagnostic method for identifying the diagnostic validity of the variable "disk-condyle relationship." PMID- 10710450 TI - Three-dimensional computed tomographic evaluation of morphologic airway changes after mandibular setback osteotomy for prognathism. AB - OBJECTIVE: To observe changes in the pharyngeal airway and the hyoid bone position after mandibular setback osteotomy in 30 patients with mandibular prognathism by means of 3-dimensional computed tomography (3DCT). STUDY DESIGN: Preoperative and postoperative computed tomography (CT) examinations were performed on 17 patients treated by sagittal split ramus osteotomy with rigid osteosynthesis and on 13 patients treated by intraoral vertical ramus osteotomy without osteosynthesis. The amount of mandibular setback was measured by the preoperative to postoperative difference of the mandibular position in axial CT images. The sizes of the preoperative and postoperative pharyngeal airway were evaluated from semitransparent and crosscut 3DCT images. Postoperative displacement of the hyoid bone was evaluated by a technique to superimpose a postoperative hard tissue 3DCT image on the preoperative image. The helical scan technique was used in the CT examination. The volume rendering technique was used to create 3DCT images. RESULTS: The mean mandibular setback was 7.8 +/- 2.1 mm with a range of 5 to 11 mm. Three months after surgery, the lateral and frontal widths of the pharyngeal airway had decreased significantly in comparison with the preoperative width. The mean reduction rates of the lateral and frontal width were 23.6% and 11.4%, respectively. The diminished airway did not recover by either 6 months or 1 year after surgery in most cases. Downward and posterior displacement of the hyoid bone was seen postoperatively. There were positive correlations between the amount of mandibular setback and reduction of the lateral width of the pharyngeal airway (r = 0.54) and the amount of hyoid bone displacement (r = 0.42). There were no significant differences between the two surgical techniques. CONCLUSION: Three-dimensional computed tomography was a practical imaging technique to evaluate the morphologic airway changes. The pharyngeal airway may have irreversible narrowing after mandibular setback surgery. PMID- 10710451 TI - Endoscopic and ultrasonographic evaluation of the maxillary sinus after combined sinus floor augmentation and implant insertion. AB - OBJECTIVE: The aim of our study was a radiographic, endoscopic, and ultrasound follow-up of the maxillary sinus comparing 2 techniques of sinus floor augmentation. STUDY DESIGN: Sonograms, radiographs (Waters' view) of the sinuses, and endoscopy served before and during surgery to evaluate the maxillary sinus. One week after the operation, ultrasound and radiograph follow-up (Waters' view) were carried out. Six months after the operation, we performed an ultrasound follow-up along with uncovering the implants. If any pathologic condition was found, we took another x-ray film of the sinuses, performed another endoscopic examination, or both. RESULTS: In 23 of 63 patients, healing was uneventful. Waters' view revealed opacification of the maxillary sinus 1 week after surgery in 40 cases when the "window technique" was used. Sinusitis occurred 3 times, as a result of migration of bone chips in 2 patients. We lost 11 of 132 inserted implants during the healing and loading periods. CONCLUSION: Endoscope-controlled sinus floor augmentation may lower the complication rate in a remaining height of the jaws between 4 and 8 mm. In our group of patients, we proved by endoscopic examination that migration of cancellous bone sequestra was the reason for sinusitis. In case of infected bone grafts with antral symptoms, sinoscopy allowed debridement and removal of a sequestrum. PMID- 10710452 TI - Expanded polytetrafluoroethylene entubulation of the rabbit inferior alveolar nerve. AB - OBJECTIVE: The purpose of this study was to evaluate the use of an expanded polytetrafluoroethylene conduit in the treatment of a 6. 0-mm gap in the rabbit inferior alveolar nerve and compare the results with those of an autogenous interpositional tibial nerve graft. STUDY DESIGN: The inferior alveolar nerves of 5 adult New Zealand White female rabbits (10 nerves) were exposed bilaterally, and a 6-mm segment of each nerve was resected. On one side, chosen at random, the gap was immediately bridged through use of an 8.0 x 2. 0-mm expanded polytetrafluoroethylene conduit; on the other side, the gap was grafted with an autogenous tibial nerve graft. Two randomly selected nerves served as sham dissected controls. At 15 weeks after surgery, the animals were killed and the entire nerve segments were harvested and prepared according to standard fixation and embedding techniques. The sections were examined histomorphometrically to quantify the degree of axonal regeneration through definition of fascicular number, total fascicular surface area, axonal density, and mean axonal diameter at 3 locations along the repair site. RESULTS: Light microscopic examination revealed the presence of disorganized neural tissue in both groups, with slightly more fibrovascular interfascicular tissue in the expanded polytetrafluoroethylene group. Histomorphometric analysis revealed no significant differences between groups for most of the measured variables. The mean axonal diameter varied between groups, and the fascicular number was greater in the expanded polytetrafluoroethylene group at the middle site. CONCLUSIONS: This study demonstrates that regeneration of the inferior alveolar nerve can occur across a 6.0-mm gap through an expanded polytetrafluoroethylene tube with results comparable to those of an autogenous nerve graft, significant donor site morbidity being avoided. The significant differences between groups were probably due to greater containment of regenerating axonal fibers in the expanded polytetrafluoroethylene group. PMID- 10710453 TI - Changing prevalence of oral manifestations of human immuno-deficiency virus in the era of protease inhibitor therapy. AB - OBJECTIVE: The purpose of this study was to determine temporal trends in the prevalence of oral manifestations of human immunodeficiency virus (HIV). STUDY DESIGN: Five hundred seventy HIV-infected adults recruited consecutively were examined by using established presumptive clinical criteria for HIV-associated oral lesions. Prevalence of oral lesions before the widespread use of HIV protease inhibitors (February 1995 through August 1996, 8% of the early sample, n = 271) was compared with lesion prevalence in a more recent period of greater protease inhibitor use (December 1996 through February 1999, 42% of the late sample, n = 299). RESULTS: Overall prevalence of oral lesions significantly decreased from early to late periods, 47.6% to 37.5%, respectively (P =.01), with some variation by lesion type. Prevalence of hairy leukoplakia (25. 8% to 11.4%; P <.01) and necrotizing periodontal diseases (4.8% to 1. 7%; P =.03) decreased, whereas HIV salivary gland disease increased (1.8% to 5.0%; P =.04). Changes in prevalence of oral candidiasis (20.3% to 16.7%), aphthous ulcers (3.7% to 3.0%), oral warts (2.2% to 4.0%), herpes simplex virus lesions (1.8% to 2.0%), and Kaposi's sarcoma (1.1% to 0.3%) were not statistically significant (P >.20 for all comparisons). CONCLUSION: The pattern of oral opportunistic infections is changing in the era of protease inhibitor use. PMID- 10710454 TI - Salivary function and glycemic control in older persons with diabetes. AB - OBJECTIVE: There is no consensus on the possible association between diabetes and salivary dysfunction in older persons with diabetes. This study's purpose was to investigate the effect of diabetes and glycemic control on salivary function in an older population. STUDY DESIGN: Twenty nine persons with type 2 diabetes and 23 nondiabetic control subjects participated (age range, 54-90 years). Diabetic status was determined by a glycosylated hemoglobin (HbA(1c)) test and a 2-hour glucose tolerance test. Poor glycemic control was defined as HbA(1c) >9%. Unstimulated whole saliva, unstimulated parotid, and stimulated parotid flow rates were measured, and subjects completed a standardized xerostomia questionnaire. RESULTS: Persons with poorly controlled diabetes had lower (P =.01) stimulated parotid flow rates than persons with well-controlled diabetes and nondiabetic control subjects. There were no significant differences in xerostomic complaints based on diabetic or glycemic control status or salivary flow rates. CONCLUSIONS: These results provide some evidence that poorly controlled diabetes may be associated with salivary dysfunction in older adults who have no concomitant complaints of xerostomia. PMID- 10710455 TI - The role of the esophagus in dental erosion. AB - OBJECTIVE: The aim of this study was to measure lower esophageal sphincter pressure, sphincter length, and esophageal motility in patients with dental erosion and compare the results with measurements made in patients without gastroesophageal reflux or dental erosion. STUDY DESIGN: Lower esophageal sphincter length and esophageal motility were measured in 39 patients (age range, 15-74 years) with dental erosion through use of static esophageal pressure monitoring; the data were compared with those from 10 control subjects (age range, 26-46 years) with nonparametric statistical tests. RESULTS: Median lower esophageal sphincter pressure was 9 mmHg (range, 0-26 mmHg) in the patients with erosion and 9.5 mmHg (range, 0 -14 mmHg) in the controls; there was no statistically significant difference between the two subject groups. Similarly, there was no statistically significant difference in esophageal length between the subject groups. There was a statistically significant difference between the groups (P =.01) in the measurement of esophageal motility; the median value was 8% (range, 0% to 100%) in the patients with erosion and 0% (range, 0% to 18%) in the controls. CONCLUSIONS: It appears that esophageal motility in patients with dental erosion is more likely to be associated with low amplitude changes than with sphincter pressure alone. Poor esophageal motility may therefore be a risk factor in regurgitation erosion. PMID- 10710456 TI - Necrosis of the tongue after transient ischemic attack. AB - We report an unusual case of necrosis of the tongue after transient ischemic attack in a 67-year-old man. Angiography revealed occlusion of the right external carotid artery at the bifurcation of the common carotid artery. Debridement of the wound and removal of the necrotic tissue resulted in good healing. PMID- 10710457 TI - Multiple maxillary and mandibular exostoses associated with multiple dermatofibromas: a case report. AB - Exostoses of the maxilla and mandible are nodular protuberances of mature bone that need to be accurately distinguished from other more diagnostically significant lesions, notably exosteal osteomas. Multiple dermatofibromas are rare and may be associated with altered immune function. We report the case of an otherwise healthy 47-year-old woman who was first seen with multiple maxillary and mandibular exostoses associated with multiple dermatofibromas. This association has not been previously reported. PMID- 10710458 TI - Necrotizing ulcerative stomatitis in human immunodeficiency virus-seropositive individuals: a review of the histopathologic, immunohistochemical, and virologic characteristics of 18 cases. AB - OBJECTIVE: The purpose of this retrospective study was to delineate the histopathologic, immunohistochemical, and virologic characteristics of 18 cases of necrotizing ulcerative stomatitis. STUDY DESIGN: Eighteen examples or oral ulcerations in human immunodeficiency virus-seropositive individuals were identified that displayed unique histopathologic features. Immunohistochemic staining for CD1a, CD3, CD23, CD68, HLA-DR, p24, cytomegalovirus, HSV-1, and HSV 2 was performed, along with in situ hybridization for Epstein-Barr virus RNA and special staining for bacteria and fungi. RESULTS: The lesions demonstrated ulceration, extensive necrosis, leukocytoclasia, histiocytic vasculitis with luminal fibrin clots, and a prominent infiltrate of large atypical cells with amphophilic cytoplasm, vesicular nuclei, and prominent nucleoli, interspersed with crescentic histiocytes, a histologic picture resembling extranodal Kikuchi's disease. Immunohistochemical findings suggested that the large atypical cells were histiocytes. Fifty-six percent (10/18) of the cases were immunoreactive for human immunodeficiency virus p24 within focal histiocytes, whereas Epstein-Barr virus RNA was identified in 1 (6%) of 17 cases. CONCLUSIONS: Necrotizing ulcerative stomatitis is an inflammatory disease characterized by specific, reproducible microscopic features. We postulate that the histopathologic resemblance of necrotizing ulcerative stomatitis to extranodal Kikuchi's disease reflects a similar immune response to differing pathogens. PMID- 10710459 TI - Comparison of clinical and histologic diagnoses in periapical lesions. AB - OBJECTIVE: To determine the frequency with which histopathologic examination of periapical biopsy specimens contributed information not anticipated clinically. STUDY DESIGN: Clinical and histopathologic information from 805 sequentially submitted periapical biopsy specimens over a 2-year period was compared. Clinical data included endodontic status, age and sex of patient, location of lesion, and submitting clinician. Histopathologic diagnoses were categorized as 1) sequelae of pulpal necrosis (SPN), 2) complicated SPN (CSPN) with infection or antral involvement, or 3) periapical lesions unrelated to pulpal necrosis (PLUPN). RESULTS: Of the 805 cases, 788 (97.9%) were SPN, 9 (1.1%) were CSPN, and 8 (1%) were PLUPN, representing a range of locally aggressive but benign lesions and 1 malignancy. Comparison of clinical and histologic diagnoses indicated that the clinical interpretation was inaccurate in 4.1% of cases (suggesting SPN in PLUPN cases or PLUPN in SPN cases). In another 0.9% of cases, the histologic analysis (indicating CSPN) contributed additional information to the clinical diagnosis. CONCLUSIONS: A histopathologic examination contributed clinically relevant information in 5.0% of submitted cases. General extrapolation of this figure is not possible. Theoretical considerations, which could positively or negatively bias this figure, are discussed. PMID- 10710460 TI - Maxillofacial hydatid cysts. AB - We report 2 cases of hydatid cysts occurring in the submandibular gland and buccal submucosa, respectively. Our first case occurred in the submandibular salivary gland of a 20-year-old woman and the second involved the buccal submucosa of a 6-year-old boy. Both diagnoses were made after the excision of the lesions. Both patients were evaluated after surgery, and both were followed up, but no other organs were involved. PMID- 10710461 TI - Myofibromas presenting in the oral cavity: a series of 9 cases. AB - OBJECTIVES: Solitary myofibromas are well described in the head and neck, but oral examples are less well known, and jaw lesions are rare. We studied the clinicopathologic features of a series of such lesions. DESIGN: Nine cases of oral myofibromas were retrieved from archives and studied. RESULTS: Two cases involved the mandible (intraosseous), 3 involved the gingiva, 2 involved the tongue, and 2, the hard palate. There were 4 men and 5 women, aged 9 months to 50 years (mean, 24 years; median, 27 years). Deep lesions showed typical histology, with paucicellular lobules and intervening hemangiopericytoma-like zones. In ulcerated submucosal lesions, these features blended superficially with cellular fascicles. The tumors expressed smooth muscle actin but lacked desmin and S100 protein. None of the tumors recurred or metastasized. CONCLUSION: Myofibromas appear in osseous, intramuscular, and submucosal aspects of the oral cavity. Ulceration imparts a fascicular appearance that makes superficial biopsy specimens difficult to interpret. PMID- 10710462 TI - Five methods of calcium hydroxide intracanal placement: an in vitro evaluation. AB - OBJECTIVE: The purpose of this study was to evaluate root canal dressing with calcium hydroxide paste using 5 techniques of placement. STUDY DESIGN: After endodontic preparation, each of 50 single-rooted premolars was filled with calcium hydroxide paste through use of one of the following techniques: Gutta Condensor, MecaShaper, K-type ultrasonic file, Lentulo, and Pastinject. Radiographs of all 3 third root zones were made after cleaning and shaping and obturation. Density measurements (in pixels) were registered by means of an image analyzer and then compared through use of analysis of variance and Newman-Keuls multiple means tests. RESULTS: With respect to average filling density, the techniques ranked as follows, in ascending order: Gutta-Condensor, MecaShaper, K type ultrasonic file, Lentulo, and Pastinject. There was a significant difference (P <.0009) between the Gutta-Condensor and Pastinject techniques, between the MecaShaper and Pastinject techniques, and between the ultrasonic file and Pastinject techniques; all favored the Pastinject. There were no significant differences between the Gutta-Condensor, MecaShaper, ultrasonic file, and Lentulo techniques or between the Lentulo and Pastinject techniques. CONCLUSIONS: The use of a specially designed paste carrier (the Pastinject) improved calcium hydroxide placement in root canals. PMID- 10710463 TI - Comparison of diagnostic accuracy of digital imaging by using CCD and CMOS-APS sensors with E-speed film in the detection of periapical bony lesions. AB - OBJECTIVE: The purpose of this study was to compare E-speed Plus film and digital imaging with a charge-coupled device (CCD) sensor and a complementary metal-oxide semiconductor active pixel sensor (CMOS-APS) in the detection of periapical bone lesions. STUDY DESIGN: Periapical lesions were created in the cortical and trabecular bone of 10 dried human mandibles. Seventy radio-graphic images and 140 digital images were evaluated by 6 endodontists and 2 radiologists. Receiver operating characteristics curves and analysis of variance were used for statistical analysis.Results. No statistically significant differences were found between film, CCD, and CMOS-APS systems. Lesion detection occurred with significantly greater accuracy in cortical bone than in trabecular bone, as well as when the cortical plate was involved. CONCLUSIONS: That no differences were found between the two sensors lends support for the use of CMOS-APS sensors, which require less system power and may have longer life spans than CCD sensors. Digital imaging required 50% less radiation than film to obtain the same diagnostic information. PMID- 10710464 TI - The effect of surface treatments of nickel-titanium files on wear and cutting efficiency. AB - OBJECTIVES: This study was designed to verify whether nitridation treatment of the cutting surfaces resulted in surface or subsurface changes that produced an increase in the resistance to wear in nickel titanium (NiTi) endodontic files. STUDY DESIGN: Some experimental samples were exposed to ionic implantation by using 150 keV of nitrogen ions and doses of 1 x 10(17) ions per cm(2). Other samples were exposed to thermal nitridation processes performed for 480 minutes at 500 degrees C temperature. Control samples were not exposed to any process. The chemical composition of the surface layers of each sample was determined by means of x-ray photoelectron spectroscopy. The cutting efficiency was tested on an endotraining bloc. RESULTS: The experimental instruments showed in-depth distributions of chemical composition that were different from those seen in the control group; thermal-nitridated instruments demonstrated a surface ratio of nickel to titanium of 0.5. Implanted samples had a higher N/Ti ratio (1.2); this ratio may be due to the presence of a layer of titanium nitride. Samples in the experimental groups showed an increase in cutting ability as compared with the controls. CONCLUSIONS: Thermal nitridation and nitrogen-ionic implantation treatment of nickel-titanium files produced a higher wear resistance and an increased cutting capacity. PMID- 10710465 TI - Updated quality assurance self-assessment exercise in intraoral and panoramic radiography. American Academy of Oral and Maxillofacial Radiology, Radiology Practice Committee. AB - This updated self-assessment exercise for the dental team by the Radiology Practice Committee of the American Academy of Oral and Maxillofacial Radiology is intended to produce the highest quality diagnostic radiographs while keeping patient exposure as low as is reasonably achievable. To continue to provide the best radiographic services to patients, those involved in dental radiography need to be aware of the latest changes and advances in dental radiography and need to use them in their practice. PMID- 10710466 TI - Radiographic absorptiometry method in measurement of localized alveolar bone density changes. AB - OBJECTIVE: The objective of this study was to measure the accuracy and precision of a radiographic absorptiometry method by using an occlusal density reference wedge in quantification of localized alveolar bone density changes. STUDY DESIGN: Twenty-two volunteer subjects had baseline and follow-up radiographs taken of mandibular premolar-molar regions with an occlusal density reference wedge in both films and added bone chips in the baseline films. The absolute bone equivalent densities were calculated in the areas that contained bone chips from the baseline and follow-up radiographs. The differences in densities described the masses of the added bone chips that were then compared with the true masses by using regression analysis. RESULTS: The correlation between the estimated and true bone-chip masses ranged from R = 0.82 to 0.94, depending on the background bone density. There was an average 22% overestimation of the mass of the bone chips when they were in low-density background, and up to 69% overestimation when in high-density background. The precision error of the method, which was calculated from duplicate bone density measurements of non-changing areas in both films, was 4.5%. CONCLUSIONS: The accuracy of the intraoral radiographic absorptiometry method is low when used for absolute quantification of bone density. However, the precision of the method is good and the correlation is linear, indicating that the method can be used for serial assessment of bone density changes at individual sites. PMID- 10710467 TI - Suitability of the general-purpose graphic printer as an image output device for digital dental x-ray images. AB - OBJECTIVES: The purpose of this study was to evaluate the suitability of the general-purpose graphic printer as an image output device for digital dental x ray images. METHODS: The image quality as obtained by a thermal printer and by a dye sublimation printer was investigated. A grid pattern image was used to check parallelism and verticality of lines in each hard copy. A step-wedge image was printed with each printer, and the optical density, gradient, and root mean square granularity were compared. Depiction ability was also compared by using test images, including small signals. RESULTS: All of the lines were parallel and vertical on hard copies of both printers. The dye sublimation printer showed better results on optical density, gradient, root mean square granularity, and depiction ability. CONCLUSION: The dye sublimation printer produces images of the depiction ability comparable to the cathode ray tube display and seems suitable as an image output device for digital dental x-ray images. PMID- 10710468 TI - Enlargement of mandibular canal without hypesthesia caused by extranodal non Hodgkin's lymphoma: a case report. AB - A rare condition of enlargement of the mandibular canal caused by an extra-nodal non-Hodgkin's lymphoma in a 59-year-old Japanese woman was reported. The patient had a swelling of the hard palate and protrusion of both ocular bulbs, which had been present for 10 years. A panoramic radiograph revealed that the right mandibular canal was widely enlarged, extending from the mandibular foramen to the mental foramen, without bone destruction. The continuous dilation of the mandibular canal to an approximate 15-mm width was associated with peripheral bony sclerosis. Computed tomography and magnetic resonance imaging showed a soft tissue tumor inside the mandibular canal. The lesion demonstrated expansive growth in the orbits, extending to the skull base through the superior orbital fissures and cavernous sinus. The lymphoma was suspected to have grown so slowly that the adjacent mandibular canal and ocular bulbs enlarged without destroying the normal bone and nervous tissue. PMID- 10710489 TI - The angiotensin II type 2 receptor: an enigma with multiple variations. AB - Since it was discovered ten years ago, the angiotensin II (ANG II) type 2 (AT(2)) receptor has been an enigma. This receptor binds ANG II with a high affinity but is not responsible for mediating any of the classical physiological actions of this peptide, all of which involve the ANG II type 1 (AT(1)) receptor. Furthermore, the AT(2) receptor exhibits dramatic differences in biochemical and functional properties and in patterns of expression compared with the AT(1) receptor. During the past decade, much information has been gathered about the AT(2) receptor, and the steadily increasing number of publications indicates a growing interest in this new and independent area of research. A number of studies suggest a role of AT(2) receptors in brain, renal, and cardiovascular functions and in the processes of apoptosis and tissue regeneration. Despite these advances, nothing stands out as the major singular function of these receptors. The study of AT(2) receptors has reached a crossroads, and innovative approaches must be considered so that unifying mechanisms as to the function of these unique receptors can be put forward. In this review we will discuss the advances that have been made in understanding the biology of the AT(2) receptor. Furthermore, we will consider how these discoveries, along with newer experimental approaches, may eventually lead to the elusive physiological and pathophysiological functions of these receptors. PMID- 10710490 TI - Oxidant damage during and after spaceflight. AB - The objectives of this study were to assess oxidant damage during and after spaceflight and to compare the results against bed rest with 6 degrees head-down tilt. We measured the urinary excretion of the F(2) isoprostane, 8-iso prostaglandin (PG) F(2alpha), and 8-oxo-7,8-dihydro-2 deoxyguanosine (8-OH DG) before, during, and after long-duration spaceflight (4-9 mo) on the Russian space station MIR, short-duration spaceflight on the shuttle, and 17 days of bed rest. Sample collections on MIR were obtained between 88 and 186 days in orbit. 8-iso PGF(2alpha) and 8-OH DG are markers for oxidative damage to membrane lipids and DNA, respectively. Data are mean +/- SE. On MIR, isoprostane levels were decreased inflight (96. 9 +/- 11.6 vs. 76.7 +/- 14.9 ng. kg(-1). day(-1), P < 0.05, n = 6) due to decreased dietary intake secondary to impaired thermoregulation. Isoprostane excretion was increased postflight (245.7 +/- 55.8 ng. kg(-1). day(-1), P < 0.01). 8-OH DG excretion was unchanged with spaceflight and increased postflight (269 +/- 84 vs 442 +/- 180 ng. kg(-1). day(-1), P < 0.05). On the shuttle, 8-OH DG excretion was unchanged in- and postflight, but 8 iso-PGF(2alpha) excretion was decreased inflight (15.6 +/- 4.3 vs 8.0 +/- 2.7 ng. kg(-1). day(-1), P < 0.05). No changes were found with bed rest, but 8-iso PGF(2alpha) was increased during the recovery phase (48.9 +/- 23.0 vs 65.4 +/- 28.3 ng. kg(-1). day(-1), P < 0.05). The changes in isoprostane production were attributed to decreased production of oxygen radicals from the electron transport chain due to the reduced energy intake inflight. The postflight increases in the excretion of the products of oxidative damage were attributed to a combination of an increase in metabolic activity and the loss of some host antioxidant defenses inflight. We conclude that 1) oxidative damage was decreased inflight, and 2) oxidative damage was increased postflight. PMID- 10710491 TI - Estimation of doubly labeled water energy expenditure with confidence intervals. AB - Bivariate regression is used to estimate energy expenditure from doubly labeled water data. Two straight lines are fitted to the logarithms of the enrichments of oxygen-18 and deuterium simultaneously as a bivariate regression, so that the correlations between the oxygen and deuterium regression coefficients can be estimated. Maximum likelihood methods are used to extend bivariate regression to unbalanced situations caused by missing observations and to include replicate laboratory determination from the same urine samples, even if one of the replicates is missing. Use of maximum likelihood allows the determination of a confidence interval for the energy expenditure based on the log likelihood surface rather than use of the propagation of variance methods for nonlinear transformations. The model is extended to include the subject's deviations from the two lines as a bivariate continuous-time first-order autoregression to allow for serial correlation in the observations. The analysis of data from two subjects, one without apparent serial correlation and one with serial correlation, is presented. PMID- 10710492 TI - Differential action of hepatic sympathetic neuropeptides: metabolic action of galanin, vascular action of NPY. AB - Activation of hepatic nerves increases both hepatic glucose production (HGP) and hepatic arterial vasoconstriction, the latter best described by a decrease of hepatic arterial conductance (HAC). Because activation of canine hepatic nerves releases the neuropeptides galanin and neuropeptide Y (NPY) as well as the classical neurotransmitter norepinephrine (NE), we sought to determine the relative role of these neuropeptides vs. norepinephrine in mediating metabolic and vascular responses of the liver. We studied the effects of local exogenous infusions of galanin and NPY on HGP and HAC to predict the metabolic and vascular function of endogenously released neuropeptide. Galanin (n = 8) or NPY (n = 4) was infused with and without NE directly into the common hepatic artery of halothane-anesthetized dogs, and we measured changes in HGP and HAC. A low dose of exogenous galanin infused directly into the hepatic artery potentiated the HGP response to NE yet had little effect on HGP when infused alone. The same dose of galanin infused into a peripheral vein (n = 8) did not potentiate the HGP response to NE, suggesting that the locally infused galanin acted directly on the liver to modulate NE's metabolic action. In contrast, a large dose of exogenous NPY failed to influence HGP when infused either alone or in combination with NE. Finally, NPY, but not galanin, tended to decrease HAC when infused alone; neither neuropeptide potentiated the HAC response to NE. Therefore, both hepatic neuropeptides may contribute to the action of sympathetic nerves on liver metabolism and blood flow. It is likely that endogenous hepatic galanin acts directly on the liver to selectively modulate norepinephrine's metabolic action, whereas endogenous hepatic NPY acts independently of NE to cause vasoconstriction. PMID- 10710493 TI - No limiting role for glycogenin in determining maximal attainable glycogen levels in rat skeletal muscle. AB - We examined whether the protein level and/or activity of glycogenin, the protein core upon which glycogen is synthesized, is limiting for maximal attainable glycogen levels in rat skeletal muscle. Glycogenin activity was 27.5 +/- 1.4, 34.7 +/- 1.7, and 39.7 +/- 1.3 mU/mg protein in white gastrocnemius, red gastrocnemius, and soleus muscles, respectively. A similar fiber type dependency of glycogenin protein levels was seen. Neither glycogenin protein level nor the activity of glycogenin correlated with previously determined maximal attainable glycogen levels, which were 69.3 +/- 5.8, 137.4 +/- 10.1, and 80.0 +/- 5.4 micromol/g wet wt in white gastrocnemius, red gastrocnemius, and soleus muscles, respectively. In additional experiments, rats were exercise trained by swimming, which resulted in a significant increase in the maximal attainable glycogen levels in soleus muscles ( approximately 25%). This increase in maximal glycogen levels was not accompanied by an increase in glycogenin protein level or activity. Furthermore, even in the presence of very high glycogen levels ( approximately 170 micromol/g wet wt), approximately 30% of the total glycogen pool continued to be present as unsaturated glycogen molecules (proglycogen). Therefore, it is concluded that glycogenin plays no limiting role for maximal attainable glycogen levels in rat skeletal muscle. PMID- 10710494 TI - In vivo rates of erythrocyte glutathione synthesis in children with severe protein-energy malnutrition. AB - Although the compromised GSH status of children with edematous protein-energy malnutrition (PEM) has been documented, the in vivo kinetic mechanism(s) responsible for this is not known. To determine if decreased synthesis contributes to the alteration of GSH homeostasis, the fractional and absolute rates of synthesis of erythrocyte GSH were determined shortly after admission (study 1), approximately 9 days postadmission (study 2), and at recovery (study 3) in seven children with edematous PEM and seven children with nonedematous PEM. Children with edematous PEM had significantly lower erythrocyte GSH and slower absolute rates of GSH synthesis than children with nonedematous PEM both shortly after admission, when they were both malnourished and infected, and approximately 9 days later, when the infection had resolved but they were still malnourished. At these times, the edematous group also had significantly lower erythrocyte GSH concentrations and absolute rates of synthesis than at recovery. Plasma and erythrocyte-free cysteine concentrations of the edematous group were significantly lower at studies 1 and 2 than at recovery. In contrast, erythrocyte GSH concentrations, rates of GSH synthesis, and plasma and erythrocyte free cysteine concentrations of the nonedematous group were similar at all three time points and greater at studies 1 and 2 than in the edematous group. These results confirm that GSH deficiency is characteristic of edematous PEM and suggest that this is due to a reduced rate of synthesis secondary to a shortage in cysteine. PMID- 10710495 TI - Twenty-four-hour rhythms of plasma glucose and insulin secretion rate in regular night workers. AB - To determine whether the ultradian and circadian rhythms of glucose and insulin secretion rate (ISR) are adapted to their permanent nocturnal schedule, eight night workers were studied during their usual 24-h cycle with continuous enteral nutrition and a 10-min blood sampling procedure and were compared with 8 day active subjects studied once with nocturnal sleep and once with an acute 8-h shifted sleep. The mean 24-h glucose and ISR levels were similar in the three experiments. The duration and the number of the ultradian oscillations were influenced neither by the time of day nor by the sleep condition or its shift, but their mean amplitude increased during sleep whenever it occurred. In day active subjects, glucose and ISR levels were high during nighttime sleep and then decreased to a minimum in the afternoon. After the acute sleep shift, the glucose and ISR rhythms were split in a biphasic pattern with a slight increase during the night of deprivation and another during daytime sleep. In night workers, the glucose and ISR peak levels exhibited an 8-h shift in accordance with the sleep shift, but the onset of the glucose rise underwent a shift of only 6 h and the sleep-related amplification of the glucose and ISR oscillations did not occur simultaneously. These results demonstrate that despite a predominant influence of sleep, the 24-h glucose and ISR rhythms are only partially adapted in permanent night workers. PMID- 10710496 TI - Hormone-independent activation of EGP during hypoglycemia is absent in type 1 diabetes mellitus. AB - It has been suggested that insulin-induced suppression of endogenous glucose production (EGP) may be counteracted independently of increased epinephrine (Epi) or glucagon during moderate hypoglycemia. We examined EGP in nondiabetic (n = 12) and type 1 diabetic (DM1, n = 8) subjects while lowering plasma glucose (PG) from clamped euglycemia (5.6 mmol/l) to values just above the threshold for Epi and glucagon secretion (3.9 mmol/l). Individualized doses of insulin were infused to maintain euglycemia during pancreatic clamps by use of somatostatin (250 microg/h), glucagon (1.0 ng. kg(-1). min(-1)), and growth hormone (GH) (3.0 ng. kg(-1). min(-1)) infusions without need for exogenous glucose. Then, to achieve physiological hyperinsulinemia (HIns), insulin infusions were fixed at 20% above the rate previously determined for each subject. In nondiabetic subjects, PG was reduced from 5.4 +/- 0.1 mmol/l to 3.9 +/- 0.1 mmol/l in the experimental protocol, whereas it was held constant (5. 3 +/- 0.2 mmol/l and 5.5 mmol/l) in control studies. In the latter, EGP (estimated by [3-(3)H]glucose) fell to values 40% of basal (P < 0.01). In contrast, in the experimental protocol, at comparable HIns but with PG at 3.9 +/- 0.1 mmol/l, EGP was activated to values about twofold higher than in the euglycemic control (P < 0.01). In DM1 subjects, EGP failed to increase in the face of HIns and PG = 3.9 +/- 0.1 mmol/l. The decrease from basal EGP in DM1 subjects (4.4 +/- 1.0 micromol. kg(-1). min(-1)) was nearly twofold that in nondiabetics (2.5 +/- 0.8 micromol. kg(-1). min(-1), P < 0.02). When PG was lowered further to frank hypoglycemia ( approximately 3.1 mmol/l), the failure of EGP activation in DM1 subjects was even more profound but associated with a 50% lower plasma Epi response (P < 0. 02) compared with nondiabetics. We conclude that glucagon- or epinephrine-independent activation of EGP may accompany other counterregulatory mechanisms during mild hypoglycemia in humans and is impaired or absent in DM1. PMID- 10710497 TI - Contribution of changes in the chloride driving force to the fading of I(GABA) in frog melanotrophs. AB - Chloride redistribution during type A gamma-aminobutyric acid (GABA(A)) currents (I(GABA)) has been investigated in cultured frog pituitary melanotrophs with imposed intracellular chloride concentration ([Cl(-)](i)) in the whole cell configuration or with unaltered [Cl(-)](i) using the gramicidin-perforated patch approach. Prolonged GABA exposures elicited reproducible decaying currents. The decay of I(GABA) was associated with both a transient fall of conductance (g(GABA)) and shift of current reversal potential (E(GABA)). The shift of E(GABA) appeared to be time and driving force dependent. In the gramicidin-perforated patch configuration, repeated GABA exposures induced currents that gradually vanished. The fading of I(GABA) was due to persistent shifts of E(GABA) as a result of g(GABA) recovering from one GABA application to another. In cells alternatively clamped at potentials closely flanking resting potential and submitted to a train of brief GABA pulses, a reversal of I(GABA) was observed after 150 s recording. It is demonstrated that, in intact frog melanotrophs, shifts of E(GABA) combine with genuine receptor desensitization to depress I(GABA). These findings strongly suggest that shifts of E(GABA) may act as a negative feedback, reducing the bioelectrical and secretory responses induced by an intense release of GABA in vivo. PMID- 10710498 TI - Hepatic alpha- and beta-adrenergic receptors are not essential for the increase in R(a) during exercise in diabetes. AB - The purpose of this study was to determine the role of direct hepatic adrenergic stimulation in the control of endogenous glucose production (R(a)) during moderate exercise in poorly controlled alloxan-diabetic dogs. Chronically catheterized and instrumented (flow probes on hepatic artery and portal vein) dogs were made diabetic by administration of alloxan. Each study consisted of a 120-min equilibration, 30-min basal, 150-min moderate exercise, 30-min recovery, and 30-min blockade test period. Either vehicle (control; n = 6) or alpha (phentolamine)- and beta (propranolol)-adrenergic blockers (HAB; n = 6) were infused in the portal vein. In both groups, epinephrine (Epi) and norepinephrine (NE) were infused in the portal vein during the blockade test period to create suprapharmacological levels at the liver. Isotopic ([3-(3)H]glucose, [U (14)C]alanine) and arteriovenous difference methods were used to assess hepatic function. Arterial plasma glucose was similar in controls (345 +/- 24 mg/dl) and HAB (336 +/- 23 mg/dl) and was unchanged by exercise. Basal arterial insulin was 5 +/- 1 mU/ml in controls and 4 +/- 1 mU/ml in HAB and fell by approximately 50% during exercise in both groups. Basal arterial glucagon was similar in controls (56 +/- 10 pg/ml) and HAB (55 +/- 7 pg/ml) and rose similarly, by approximately 1.4-fold, with exercise in both groups. Despite greater arterial Epi and NE levels in HAB compared with controls during the basal and exercise periods, exercise-induced increases in catecholamines from basal were similar in both groups. Gluconeogenic conversion from alanine and lactate and the intrahepatic efficiency of this process were increased by twofold during exercise in both groups. R(a) rose similarly by 2.9 +/- 0.7 and 2.7 +/- 1.0 mg. kg(-1). min(-1) at time = 150 min during exercise in controls and HAB. During the blockade test period, arterial plasma glucose and R(a) rose to 454 +/- 43 mg/dl and 11.3 mg. kg(-1). min(-1) in controls, respectively, but were essentially unchanged in HAB. The attenuated response to the blockade test in HAB substantiates the effectiveness of the hepatic adrenergic blockade. In conclusion, these results demonstrate that direct hepatic adrenergic stimulation does not play a role in the stimulation of R(a) during exercise in poorly controlled diabetes. PMID- 10710499 TI - Methysergide reduces nonnutritive blood flow in normal and scalded skin. AB - Methysergide is a serotonin antagonist and has been demonstrated to reduce wound blood flow and edema formation. We have determined the effect of methysergide on protein kinetics in normal and scalded skin of anesthetized rabbits. L-[ring (13)C(6)]- or L-[ring-(2)H(5)]phenylalanine was used to reflect skin protein kinetics by use of an ear model, and L-[1-(13)C]leucine was used to reflect whole body protein kinetics. The results were that infusion of methysergide (2-3 mg. kg(-1). h(-1)) reduced the blood flow rate in normal skin by 50% without changing skin or whole body protein kinetics. After scald injury on the ear, administration of methysergide for 48 h reduced the weight of scalded ears (43 +/ 4 vs. 30 +/- 5 g, P < 0.01) and ear blood flow rate (42.6 +/- 4.9 vs. 5.8 +/- 1.0 ml. 100 g(-1). min(-1), P < 0.0001) and did not change wound protein kinetics. Methysergide reduced arteriovenous shunting and maintained inward phenylalanine transport from the blood to the skin pool. Using the microsphere technique, we found that the infusion of methysergide decreased blood perfusion by 33-36% in both normal and scalded ear skin. We conclude that methysergide administration reduces nonnutritive, as opposed to nutritive, blood flow in normal and scalded skin. PMID- 10710500 TI - Sensitivity of CPT I to malonyl-CoA in trained and untrained human skeletal muscle. AB - The present study examined the sensitivity of carnitine palmitoyltransferase I (CPT I) activity to its inhibitor malonyl-CoA (M-CoA), and simulated metabolic conditions of rest and exercise, in aerobically trained and untrained humans. Maximal CPT I activity was measured in mitochondria isolated from resting human skeletal muscle. Mean CPT I activity was 492.8 +/- 72.8 and 260.8 +/- 33.6 micromol. min(-1). kg wet muscle(-1) in trained and untrained subjects, respectively (pH 7.0, 37 degrees C). The sensitivity to M-CoA was greater in trained muscle; the IC(50) for M-CoA was 0.17 +/- 0.04 and 0.49 +/- 0.17 microM in trained and untrained muscle, respectively. The presence of acetyl-CoA, free coenzyme A (CoASH), and acetylcarnitine, in concentrations simulating rest and exercise conditions did not release the M-CoA-induced inhibition of CPT I activity. However, CPT I activity was reduced at pH 6.8 vs. pH 7.0 in both trained and untrained muscle in the presence of physiological concentrations of M CoA. The results of this study indicate that aerobic training is associated with an increase in the sensitivity of CPT I to M-CoA. Accumulations of acetyl-CoA, CoASH, and acetylcarnitine do not counteract the M-CoA-induced inhibition of CPT I activity. However, small decreases in pH produce large reductions in the activity of CPT I and may contribute to the decrease in fat metabolism that occurs during moderate and intense aerobic exercise intensities. PMID- 10710501 TI - Integrative physiology of splanchnic glutamine and ammonium metabolism. AB - The substrates for hepatic ureagenesis are equimolar amounts of ammonium and aspartate. The study design mimics conditions in which the liver receives more NH(+)(4) than aspartate precursors (very low-protein diet). Fasted dogs, fitted acutely with transhepatic catheters, were infused with a tracer amount of (15)NH(4)Cl. From arteriovenous differences, the major NH(+)(4) precursor for hepatic ureagenesis was via deamidation of glutamine in the portal drainage system (rather than in the liver), because there was a 1:1 stoichiometry between glutamine disappearance and NH(+)(4) appearance, and the amide (but not the amine) nitrogen of glutamine supplied the (15)N added to the portal venous NH(+)(4) pool. The liver extracted all this NH(+)(4) from glutamine deamidation plus an additional amount in a single pass, suggesting that there was an activator of hepatic ureagenesis. The other major source of nitrogen extracted by the liver was [(14)N]alanine. Because alanine was not produced in the portal venous system, we speculate that it was derived ultimately from proteins in peripheral tissues. PMID- 10710502 TI - Somatotropin increases protein balance by lowering body protein degradation in fed, growing pigs. AB - Somatotropin (ST) administration enhances protein deposition in well-nourished, growing animals. To determine whether the anabolic effect is due to an increase in protein synthesis or a decrease in proteolysis, pair-fed, weight-matched ( approximately 20 kg) growing swine were treated with porcine ST (150 microg. kg( 1). day(-1), n = 6) or diluent (n = 6) for 7 days. Whole body leucine appearance (R(a)), nonoxidative leucine disposal (NOLD), urea production, and leucine oxidation, as well as tissue protein synthesis (K(s)), were determined in the fed steady state using primed continuous infusions of [(13)C]leucine, [(13)C]bicarbonate, and [(15)N(2)]urea. ST treatment increased the efficiency with which the diet was used for growth. ST treatment also increased plasma insulin-like growth factor I (+100%) and insulin (+125%) concentrations and decreased plasma urea nitrogen concentrations (-53%). ST-treated pigs had lower leucine R(a) (-33%), leucine oxidation (-63%), and urea production (-70%). However, ST treatment altered neither NOLD nor K(s) in the longissimus dorsi, semitendinosus, or gastrocnemius muscles, liver, or jejunum. The results suggest that in the fed state, ST treatment of growing swine increases protein deposition primarily through a suppression of protein degradation and amino acid catabolism rather than a stimulation of protein synthesis. PMID- 10710503 TI - High-frequency oscillations in circulating amylin concentrations in healthy humans. AB - Amylin is stored in the pancreatic beta-cell granules and cosecreted with insulin in response to nutrient stimuli. To gain further insight into control of hormonal release in beta-cell physiology, we examined whether amylin, like insulin, circulates in a high-frequency oscillatory pattern, and if it does, to compare the secretory patterns of the two hormones. Eight overnight-fasted healthy individuals were studied during intravenous glucose infusion (2.0 mg. kg(-1). min(-1)). Blood was collected every minute for 90 min and analyzed in triplicate for amylin, total amylin immunoreactivity (TAI), and insulin. Mean plasma concentrations of amylin (nonglycosylated), TAI (nonglycosylated plus glycosylated), insulin, and glucose were 2.77 +/- 1.21 pmol/l, 7.60 +/- 1.73 pmol/l, 50.4 +/- 17.5 pmol/l, and 5.9 +/- 0.3 mmol/l, respectively. The 90-min time series of amylin, TAI, and insulin were analyzed for periodicity (by spectral analysis, autocorrelation analysis, and deconvolution analysis) and regularity [by approximate entropy (ApEn)]. Significant spectral density peaks were demonstrated by a random shuffling technique in 7 (out of 7), 8 (out of 8), and 8 (out of 8) time series, respectively, whereas autocorrelation analysis revealed significant pulsatility in 5 (out of 7), 7 (out of 8), and 5 (out of 8), respectively. The dominant periodicity of oscillations determined by spectral analysis was 4.6 +/- 0.3, 4.6 +/- 0.4, and 6. 5 +/- 1.1 min/pulse, respectively (amylin vs. insulin, P = 0.017, TAI vs. insulin, P = 0.018). By deconvolution analysis, amylin and insulin periodicities were, respectively, 6.3 +/- 1.0 and 5.5 +/- 0. 6 min. By application of the regularity statistic, ApEn, 6 (out of 7), 7 (out of 8), and 6 (out of 8), respectively, were found to be significantly different from random. In conclusion, like several other hormones, circulating amylin concentrations exhibit oscillations in the secretory patterns for nonglycosylated as well as glycosylated forms. Whether the high-frequency pulsatile release of amylin is disturbed in diabetes is not known. PMID- 10710504 TI - Cod and soy proteins compared with casein improve glucose tolerance and insulin sensitivity in rats. AB - The aim of the present study was to determine the effects of feeding various dietary proteins on insulin sensitivity and glucose tolerance in rats. Male Wistar rats were fed for 28 days with isoenergetic diets containing either casein, soy protein, or cod protein. Cod protein-fed and soy protein-fed rats had lower fasting plasma glucose and insulin concentrations compared with casein-fed animals. After intravenous glucose bolus, cod protein- and soy protein-fed rats induced lower incremental areas under glucose curves compared with casein-fed animals. Improved peripheral insulin sensitivity was confirmed by higher glucose disposal rates in cod protein- and soy protein-fed rats (15.2 +/- 0.3 and 13.9 +/ 0.6 mg. kg(-1). min(-1), respectively) compared with casein-fed animals (6.5 +/- 0.7 mg. kg(-1). min(-1), P < 0.05). Moreover, test meal experiments revealed that, in the postprandial state, the lower plasma insulin concentrations in cod protein- and soy protein-fed animals could be also due to decreased pancreatic insulin release and increased hepatic insulin removal. In conclusion, the metabolic responses to three common dietary proteins indicate that cod and soy proteins, when compared with casein, improve fasting glucose tolerance and peripheral insulin sensitivity in rats. PMID- 10710505 TI - Roles of insulin resistance and obesity in regulation of plasma insulin concentrations. AB - Plasma glucose, insulin, and C-peptide concentrations were determined in response to graded infusions of glucose, and insulin secretion rates were calculated over each sampling period. Measurements were also made of insulin clearance, resistance to insulin-mediated glucose, uptake, and the plasma glucose, insulin, and C-peptide concentrations at hourly intervals from 8:00 AM to 4:00 PM in response to breakfast and lunch. Plasma glucose, insulin, and C-peptide concentrations were significantly (P < 0.01) higher in obese women in response to the graded intravenous glucose infusion, associated with a 40% (P < 0.005) greater insulin secretory response. Degree of insulin resistance correlated positively (P < 0.05) with the increase in insulin secretion rate in both nonobese (r = 0.52) and obese (r = 0.58) groups and inversely (P < 0.05) with the decrease in insulin clearance in obese (r = -0.46) and nonobese (r = -0.39) individuals. Weight loss was associated with significantly lower plasma glucose, insulin, and C-peptide concentrations in response to graded glucose infusions and in day-long insulin concentrations. Neither insulin resistance nor the insulin secretory response changed after weight loss, whereas there was a significant increase in the rate of insulin clearance during the glucose infusion. It is concluded that 1) obesity is associated with a shift to the left in the glucose stimulated insulin secretory dose-response curve as well as a decrease in insulin clearance and 2) changes in insulin secretion and insulin clearance in obese women are more a function of insulin resistance than obesity. PMID- 10710506 TI - Cortisol inhibits hepatocyte growth factor/scatter factor expression and induces c-met transcripts in osteoblasts. AB - Hepatocyte growth factor/scatter factor (HGF/SF) is expressed by osteoblasts and has important effects on repair and bone remodeling. Because glucocorticoids regulate these two functions, we tested the effects of cortisol on the expression of HGF/SF and c-met, the protooncogene encoding the HGF/SF receptor, in cultures of osteoblast-enriched cells from 22-day fetal rat calvariae (Ob cells). Cortisol decreased HGF/SF mRNA levels and diminished the induction of HGF/SF transcripts by fibroblast growth factor-2 (FGF-2) and platelet-derived growth factor BB (PDGF BB). Cortisol also decreased FGF-2 and PDGF BB-induced HGF/SF mRNA and polypeptide levels in MC3T3 cells. In contrast, cortisol enhanced the expression of c-met transcripts in Ob cells. Cortisol did not modify the half-life of HGF/SF or of c-met mRNA in transcriptionally arrested cells, and it increased the rate of transcription of c-met. In conclusion, cortisol decreases HGF/SF transcripts in Ob cells and enhances c-met expression transcriptionally. The effects of cortisol on HGF/SF could be relevant to its inhibitory actions on bone formation and repair. PMID- 10710507 TI - Catabolism of arginine and ornithine in the perfused rat liver: effect of dietary protein and of glucagon. AB - The rates of oxidation of arginine and ornithine that occurred through a reaction pathway involving the enzyme ornithine aminotransferase (EC 2.6.1.13) were determined using (14)C-labeled amino acids in the isolated nonrecirculating perfused rat liver. At physiological concentrations of these amino acids, their catabolism is subject to chronic regulation by the level of protein consumed in the diet. (14)CO(2) production from [U-(14)C]ornithine (0.1 mM) and from [U (14)C]arginine (0.2 mM) was increased about fourfold in livers from rats fed 60% casein diets for 3-4 days. The catabolism of arginine in the perfused rat liver, but not that of ornithine, is subject to acute regulation by glucagon (10(-7) M), which stimulated arginine catabolism by approximately 40%. Dibutyryl cAMP (0.1 mM) activated arginine catabolism to a similar extent. In retrograde perfusions, glucagon caused a twofold increase in the rate of arginine catabolism, suggesting an effect of glucagon on arginase in the perivenous cells. PMID- 10710508 TI - Regulation of glycogen phosphorylase and PDH during exercise in human skeletal muscle during hypoxia. AB - The present study examined the acute effects of hypoxia on the regulation of skeletal muscle metabolism at rest and during 15 min of submaximal exercise. Subjects exercised on two occasions for 15 min at 55% of their normoxic maximal oxygen uptake while breathing 11% O(2) (hypoxia) or room air (normoxia). Muscle biopsies were taken at rest and after 1 and 15 min of exercise. At rest, no effects on muscle metabolism were observed in response to hypoxia. In the 1st min of exercise, glycogenolysis was significantly greater in hypoxia compared with normoxia. This small difference in glycogenolysis was associated with a tendency toward a greater concentration of substrate, free P(i), in hypoxia compared with normoxia. Pyruvate dehydrogenase activity (PDH(a)) was lower in hypoxia at 1 min compared with normoxia, resulting in a reduced rate of pyruvate oxidation and a greater lactate accumulation. During the last 14 min of exercise, glycogenolysis was greater in hypoxia despite a lower mole fraction of phosphorylase a. The greater glycogenolytic rate was maintained posttransformationally through significantly higher free [AMP] and [P(i)]. At the end of exercise, PDH(a) was greater in hypoxia compared with normoxia, contributing to a greater rate of pyruvate oxidation. Because of the higher glycogenolytic rate in hypoxia, the rate of pyruvate production continued to exceed the rate of pyruvate oxidation, resulting in significant lactate accumulation in hypoxia compared with no further lactate accumulation in normoxia. Hence, the elevated lactate production associated with hypoxia at the same absolute workload could in part be explained by the effects of hypoxia on the activities of the rate-limiting enzymes, phosphorylase and PDH, which regulate the rates of pyruvate production and pyruvate oxidation, respectively. PMID- 10710509 TI - Hyperglycemia contributes insulin resistance in hepatic and adipose tissue but not skeletal muscle of ZDF rats. AB - To determine the contribution of hyperglycemia to the insulin resistance in various insulin-sensitive tissues of Zucker diabetic fatty (ZDF) rats, T-1095, an oral sodium-dependent glucose transporter (SGLT) inhibitor, was administered by being mixed into food. Long-term treatment with T-1095 lowered both fed and fasting blood glucose levels to near normal ranges. A hyperinsulinemic euglycemic clamp study that was performed after 4 wk of T-1095 treatment demonstrated partial recovery of the reduced glucose infusion rate (GIR) in the T-1095-treated group. In the livers of T-1095-treated ZDF rats, hepatic glucose production rate (HGP) and glucose utilization rate (GUR) showed marked recovery, with almost complete normalization of reduced glucokinase/glucose-6-phosphatase (G-6-Pase) activities ratio. In adipose tissues, decreased GUR was also shown to be significantly improved with a normalization of insulin-induced GLUT-4 translocation. In contrast, in skeletal muscles, the reduced GUR was not significantly improved in response to amelioration of hyperglycemia by T-1095 treatment. These results suggest that the contribution of hyperglycemia to insulin resistance in ZDF rats is very high in the liver and considerably elevated in adipose tissues, although it is very low in skeletal muscle. PMID- 10710510 TI - Combined intraportal infusion of acetylcholine and adrenergic blockers augments net hepatic glucose uptake. AB - Portal glucose delivery in the conscious dog augments net hepatic glucose uptake (NHGU). To investigate the possible role of altered autonomic nervous activity in the effect of portal glucose delivery, the effects of adrenergic blockade and acetylcholine (ACh) on hepatic glucose metabolism were examined in 42-h-fasted conscious dogs. Each study consisted of an equilibration (-120 to -20 min), a control (-20 to 0 min), and a hyperglycemic-hyperinsulinemic period (0 to 300 min). During the last period, somatostatin (0.8 microg. kg(-1). min(-1)) was infused along with intraportal insulin (1.2 mU. kg(-1). min(-1)) and glucagon (0.5 ng. kg(-1). min(-1)). Hepatic sinusoidal insulin was four times basal (73 +/ 7 microU/ml) and glucagon was basal (55 +/- 7 pg/ml). Glucose was infused peripherally (0-300 min) to create hyperglycemia (220 mg/dl). In test protocol, phentolamine and propranolol were infused intraportally at 0.2 microg and 0.1 microg. kg(-1). min(-1) from 120 min on. ACh was infused intraportally at 3 microg. kg(-1). min(-1) from 210 min on. In control protocol, saline was given in place of the blockers and ACh. Hyperglycemia-hyperinsulinemia switched the net hepatic glucose balance (mg. kg(-1). min(-1)) from output (2.1 +/- 0.3 and 1.1 +/ 0.2) to uptake (2.8 +/- 0.9 and 2.6 +/- 0.6) and lactate balance (micromol. kg( 1). min(-1)) from uptake (7.5 +/- 2.2 and 6.7 +/- 1.6) to output (3.7 +/- 2.6 and 3.9 +/- 1.6) by 120 min in the control and test protocols, respectively. Thereafter, in the control protocol, NHGU tended to increase slightly (3.0 +/- 0.6 mg. kg(-1). min(-1) by 300 min). In the test protocol, adrenergic blockade did not alter NHGU, but ACh infusion increased it to 4.4 +/- 0.6 and 4.6 +/- 0.6 mg. kg(-1). min(-1) by 220 and 300 min, respectively. These data are consistent with the hypothesis that alterations in nerve activity contribute to the increase in NHGU seen after portal glucose delivery. PMID- 10710511 TI - Protein kinase C modulates insulin action in human skeletal muscle. AB - There is good evidence from cell lines and rodents that elevated protein kinase C (PKC) overexpression/activity causes insulin resistance. Therefore, the present study determined the effects of PKC activation/inhibition on insulin-mediated glucose transport in incubated human skeletal muscle and primary adipocytes to discern a potential role for PKC in insulin action. Rectus abdominus muscle strips or adipocytes from obese, insulin-resistant, and insulin-sensitive patients were incubated in vitro under basal and insulin (100 nM)-stimulated conditions in the presence of GF 109203X (GF), a PKC inhibitor, or 12 deoxyphorbol 13-phenylacetate 20-acetate (dPPA), a PKC activator. PKC inhibition had no effect on basal glucose transport. GF increased (P < 0.05) insulin stimulated 2-deoxyglucose (2-DOG) transport approximately twofold above basal. GF plus insulin also increased (P < 0.05) insulin receptor tyrosine phosphorylation 48% and phosphatidylinositol 3-kinase (PI 3-kinase) activity approximately 50% (P < 0.05) vs. insulin treatment alone. Similar results for GF on glucose uptake were observed in human primary adipocytes. Further support for the hypothesis that elevated PKC activity is related to insulin resistance comes from the finding that PKC activation by dPPA was associated with a 40% decrease (P < 0.05) in insulin-stimulated 2-DOG transport. Incubation of insulin-sensitive muscles with GF also resulted in enhanced insulin action ( approximately 3-fold above basal). These data demonstrate that certain PKC inhibitors augment insulin mediated glucose uptake and suggest that PKC may modulate insulin action in human skeletal muscle. PMID- 10710512 TI - Correction of diet-induced hyperglycemia, hyperinsulinemia, and skeletal muscle insulin resistance by moderate hyperleptinemia. AB - Human obesity and high fat feeding in rats are associated with the development of insulin resistance and perturbed carbohydrate and lipid metabolism. It has been proposed that these metabolic abnormalities may be reversible by interventions that increase plasma leptin. Up to now, studies in nongenetic animal models of obesity and in human obesity have concentrated on multiple injection therapy with mixed results. Our study sought to determine whether a sustained, moderate increase in plasma leptin, achieved by administration of a recombinant adenovirus containing the leptin cDNA (AdCMV-leptin) would be effective in reversing the metabolic abnormalities of the obese phenotype. Wistar rats fed a high-fat diet (HF) were heavier (P < 0.05), had increased fat mass and intramuscular triglycerides (mTG), and had elevated plasma glucose, insulin, triglyceride, and free fatty acids compared with standard chow-fed (SC) control animals (all P < 0.01). HF rats also had impaired glucose tolerance and were markedly insulin resistant, as demonstrated by a 40% reduction in insulin-stimulated muscle glucose uptake (P < 0.001). Increasing plasma leptin levels to 29.0 +/- 1.5 ng/ml (from 7.0 +/- 1.4 ng/ml, P < 0.001) for a period of 6 days decreased adipose mass by 40% and normalized plasma glucose and insulin levels. In addition, insulin stimulated skeletal muscle glucose uptake was normalized in hyperleptinemic rats, an effect that correlated closely with a 60% (P < 0.001) decrease in mTG. Importantly, HF rats that received a control adenovirus containing the beta galactosidase cDNA and were calorically matched to AdCMV-leptin-treated animals remained hyperglycemic, hyperinsulinemic, insulin resistant, and maintained elevated mTG. We conclude that a gene-therapeutic intervention that elevates plasma leptin moderately for a sustained period reverses diet-induced hyperglycemia, hyperinsulinemia, and skeletal muscle insulin resistance, and that these improvements are tightly linked to leptin-induced reductions in mTG. PMID- 10710513 TI - Polyamines in the lung: polyamine uptake and polyamine-linked pathological or toxicological conditions. AB - The natural polyamines putrescine, cadaverine, spermidine, and spermine are found in all cells. These (poly)cations exert interactions with anions, e.g., DNA and RNA. This feature represents their best-known direct physiological role in cellular functions: cell growth, division, and differentiation. The lung and, more specifically, alveolar epithelial cells appear to be endowed with a much higher polyamine uptake system than any other major organ. In the lung, the active accumulation of natural polyamines in the epithelium has been studied in various mammalian species including rat, hamster, rabbit, and human. The kinetic parameters (Michaelis-Menten constant and maximal uptake) of the uptake system are the same order of magnitude regardless of the polyamine or species studied and the in vitro system used. Also, other pulmonary cells accumulate polyamines but never to the same extent as the epithelium. Although different uptake systems exist for putrescine, spermidine, and spermine in the lung, neither the nature of the carrier protein nor the reason for its existence is known. Some pulmonary toxicological and/or pathological conditions have been related to polyamine metabolism and/or polyamine content in the lung. Polyamines possess an important intrinsic toxicity. From in vitro studies with nonpulmonary cells, it has been shown that spermidine and spermine can be metabolized to hydrogen peroxide, ammonium, and acrolein, which can all cause cellular toxicity. In hyperoxia or after ozone exposure, the increased polyamine synthesis and polyamine content of the rat lung is correlated with survival of the animals. Pulmonary hypertension induced by monocrotaline or hypoxia has also been linked to the increased polyamine metabolism and polyamine content of the lung. In a small number of studies, it has been shown that polyamines can contribute to the suppression of immunologic reactions in the lung. PMID- 10710514 TI - Mechanism of hypoxic pulmonary vasoconstriction involves ET(A) receptor-mediated inhibition of K(ATP) channel. AB - There is controversy on the role of endothelin (ET)-1 in the mechanism of hypoxic pulmonary vasoconstriction (HPV). Although HPV is inhibited by ET-1 subtype A (ET(A))-receptor antagonists in animals, it has been reported that ET(A)-receptor blockade does not affect HPV in isolated lungs. Thus we reassessed the role of ET 1 in HPV in both rats and isolated blood- and physiological salt solution (PSS) perfused rat lungs. In rats, the ET(A)-receptor antagonist BQ-123 and the nonselective ET(A)- and ET(B)-receptor antagonist PD-145065, but not the ET(B) receptor antagonist BQ-788, inhibited HPV. Similarly, BQ-123, but not BQ-788, attenuated HPV in blood-perfused lungs. In PSS-perfused lungs, either BQ-123, BQ 788, or the combination of both attenuated HPV equally. Inhibition of HPV by combined BQ-123 and BQ-788 in PSS-perfused lungs was prevented by costimulation with angiotensin II. The ATP-sensitive K(+) (K(ATP))-channel blocker glibenclamide also prevented inhibition of HPV by BQ-123 in both lungs and rats. These results suggest that ET-1 contributes to HPV in both isolated lungs and intact animals through ET(A) receptor-mediated suppression of K(ATP)-channel activity. PMID- 10710515 TI - Lamellar body membrane turnover is stimulated by secretagogues. AB - Lamellar bodies are specialized cellular organelles used for storage of surfactant by alveolar type II cells of the lung. We utilized monoclonal antibody (MAb) 3C9, which recognizes an integral lamellar body-limiting membrane protein of 180 kDa, to follow lamellar body trafficking. (125)I-labeled MAb 3C9 bound to the surface of type II cells and was internalized by the cells in a time- and concentration-dependent manner that was inhibitable by excess unlabeled antibody. The internalized antibody remained undegraded over a 4-h time period. The L2 rat lung cell line that does not have lamellar bodies did not bind iodinated 3C9. Exposure of type II cells to the secretagogues ATP, phorbol 12-myristate 13 acetate, and cAMP resulted in a 1.5- to 2-fold enhancement of binding and uptake of MAb 3C9. Calphostin C inhibited phorbol 12-myristate 13-acetate-stimulated phospholipid secretion and also reduced binding and uptake of MAb 3C9 by type II cells. Treatment of type II cells with phenylarsine oxide to obstruct clathrin mediated endocytosis had no effect on the internalization of MAb 3C9 while markedly blocking the uptake of surfactant protein A and transferrin. An actin mediated process was important for lamellar body membrane uptake because incubation with cytochalasin D partially inhibited MAb 3C9 incorporation by type II cells. These studies are compatible with enhanced lamellar body membrane turnover associated with surfactant secretion and indicate that this process can be monitored by the trafficking of the antigen reporter MAb 3C9. PMID- 10710516 TI - Influence of growth hormone on the lung growth response to tracheal obstruction in fetal sheep. AB - Obstructing the fetal trachea is a potent stimulus for fetal lung growth, but little is known about the factors that regulate this process. Our aim was to determine the role of growth hormone (GH) in regulating the increase in lung growth induced by obstruction of the trachea in fetal sheep. Twenty chronically catheterized fetal sheep, nine of which were hypophysectomized, were divided into four experimental groups: 1) control group (n = 4), 2) a group in which the fetal trachea was obstructed for 3 days (3-day obstructed; n = 6), 3) a 3-day obstructed group in which the pituitary was removed [hypophysectomized (HX)] and the fetus was given maintenance infusions of ACTH, thyroxine, and human GH (hGH; HX hGH 3-day obstructed; n = 5), and 4) a HX 3-day obstructed group in which the fetus was given maintenance infusions of ACTH and thyroxine (n = 5). Tracheal obstruction significantly increased fetal lung liquid volumes from 37.2 +/- 3.2 ml/kg in control fetuses to 75.6 +/- 9.0 ml/kg in 3-day obstructed fetuses, and the presence or absence of GH did not affect this increase. Similarly, the presence or absence of GH did not affect the increase in lung weight or protein content induced by 3 days of tracheal obstruction. However, in the absence of GH, 3 days of tracheal obstruction failed to increase total lung DNA content above unobstructed control values (107.9 +/- 5.3 and 94. 1 +/- 7.0 mg/kg for control and HX 3-day obstructed groups, respectively). In contrast, 3 days of tracheal obstruction increased total lung DNA content to a similar extent in fetuses with an intact pituitary and HX fetuses that received GH replacement (126.0 +/- 4.4 and 126.7 +/- 4.0 mg/kg for 3-day obstructed and HX hGH 3-day obstructed groups, respectively). These data indicate that the absence of GH either abolishes or delays the acceleration in cell division caused by an increase in fetal lung expansion. PMID- 10710517 TI - Pulmonary arterial smooth muscle cells modulate cytokine- and LPS-induced cytotoxicity in endothelial cells. AB - Cytokines and lipopolysaccharide (LPS) are known to be injurious to vascular endothelial cells (ECs), but the influence of adjacent vascular smooth muscle cells (SMCs) on this injury is unknown. Exposure of cultured rat (RPMECs) or human (HPMECs) pulmonary microvascular ECs on tissue culture plastic to a mixture of cytokines (interleukin-1beta, tumor necrosis factor-alpha, and interferon gamma) and LPS (cytomix) resulted in a significant increase in (51)Cr release to 35-40%. When unstimulated RPMECs were cocultured with cytomix-pretreated rat pulmonary microvascular SMCs (RPMSMCs) there was an increase in (51)Cr release to 8.4%, which was nitric oxide dependent. However, when RPMECs or HPMECs were stimulated in direct contact with their respective SMCs, rather than a further increase in cytomix-induced injury (e.g., >35-40%), (51)Cr release decreased to <10%. This cytoprotection was fully reproduced with fixed RPMSMCs, and partially reproduced by plating HPMECs on gelatin. These data show that the direct toxicity of a cytokine and endotoxin mixture on cultured ECs can be reduced by contact with vascular smooth muscle. PMID- 10710518 TI - Interstrain variation in murine susceptibility to inhaled acid-coated particles. AB - Epidemiologic studies have demonstrated a positive correlation between concentration of acid aerosol and increased morbidity and mortality in many urban environments. To determine whether genetic background is an important risk factor for susceptibility to the toxic effects of inhaled particles, we studied the interstrain (genetic) and intrastrain (environmental) variance of lung responses to acid-coated particle (ACP) aerosol in nine strains of inbred mice. A flow-past nose-only inhalation system was used to expose mice to ACPs produced by the cogeneration of a carbon black aerosol-sulfur dioxide (SO(2)) mixture at high humidity. Three days after a single 4-h exposure to ACPs or filtered air, mice underwent bronchoalveolar lavage, and cell differentials and total protein were determined as indexes of inflammation and epithelial permeability, respectively. To determine the effect of ACPs on alveolar macrophage (AM) function, lavaged AMs were isolated from exposed animals and Fc receptor-mediated phagocytosis was evaluated. Compared with air-exposed animals, there was a slight but significant exposure effect of ACPs on the mean number of lavageable polymorphonuclear leukocytes in C3H/HeJ and C3H/HeOuJ mice. ACP exposure also caused a significant decrease in AM phagocytosis. Relative to respective air-exposed animals, Fc receptor-mediated phagocytosis was suppressed in eight of nine strains. The order of strain-specific effect of ACPs on phagocytosis was C57BL/6J > 129/J > SJL/J > BALB/cJ > C3H/HeOuJ > A/J > SWR/J > AKR/J. There was no effect of ACP exposure on AM phagocytosis in C3H/HeJ mice. The significant interstrain variation in AM response to particle challenge indicates that genetic background has an important role in susceptibility. The effects of ACPs on AM function, inflammation, and epithelial hyperpermeability were not correlated (i.e., no cosegregation). This model may have important implications concerning interindividual variation in particle-induced compromise of host defense. PMID- 10710519 TI - Identification of functional TTF-1 and Sp1/Sp3 sites in the upstream promoter region of rabbit SP-B gene. AB - Surfactant protein B (SP-B) is essential for the maintenance of biophysical properties and physiological function of pulmonary surfactant. SP-B mRNA is expressed in a cell type-restricted manner in alveolar type II and bronchiolar (Clara) epithelial cells of the lung and is developmentally induced. In NCI-H441 cells, a lung cell line with characteristics of Clara cells, a minimal promoter region comprising -236 to +39 nucleotides supports high-level expression of chloramphenicol acetyltransferase reporter activity. In the present investigation, we characterized the upstream promoter region, -236 to -140 nucleotides, that is essential for promoter activity. Deletion mapping identified two segments, -236 to -170 and -170 to -140 nucleotides, that are important for promoter activity. Mutational analysis and gel mobility shift experiments identified thyroid transcription factor-1, Sp1, and Sp3 as important trans-acting factors that bind to sequences in the upstream promoter region. Our data suggest that SP-B promoter activity is dependent on interactions between factors bound to upstream and downstream regions of the promoter. PMID- 10710520 TI - Effect of bradykinin on membrane properties of guinea pig bronchial parasympathetic ganglion neurons. AB - The effect of bradykinin on membrane properties of parasympathetic ganglion neurons in isolated guinea pig bronchial tissue was studied using intracellular recording techniques. Bradykinin (1-100 nM) caused a reversible membrane potential depolarization of ganglion neurons that was not associated with a change in input resistance. The selective bradykinin B(2) receptor antagonist HOE 140 inhibited bradykinin-induced membrane depolarizations. Furthermore, the cyclooxygenase inhibitor indomethacin attenuated bradykinin-induced membrane depolarizations to a similar magnitude ( approximately 70%) as HOE-140. However, neurokinin-1 and -3 receptor antagonists did not have similar inhibitory effects. The ability of bradykinin to directly alter active properties of parasympathetic ganglion neurons was also examined. Bradykinin (100 nM) significantly reduced the duration of the afterhyperpolarization (AHP) that followed four consecutive action potentials. The inhibitory effect of bradykinin on the AHP response was reversed by HOE-140 but not by indomethacin. These results indicate that bradykinin can stimulate airway parasympathetic ganglion neurons independent of sensory nerve activation and provide an alternative mechanism for regulating airway parasympathetic tone. PMID- 10710521 TI - O(2)-evoked regulation of HIF-1alpha and NF-kappaB in perinatal lung epithelium requires glutathione biosynthesis. AB - To test the genetic capacity of the perinatal lung to respond to O(2) shifts that coincide with the first respiratory movements, rat fetal alveolar type II (fATII) epithelial cells were cultured at fetal distal lung PO(2) (23 Torr) and then exposed to postnatal (23 --> 76 Torr; mild hyperoxic shift), moderate (23 --> 152 Torr; moderate hyperoxic shift), or severe (23 --> 722 Torr; severe hyperoxic shift) oxygenation. Nuclear abundance and consensus binding characteristics of hypoxia-inducible factor (HIF)-1alpha and nuclear factor (NF)-kappaB (Rel A/p65) plus glutathione biosynthetic capacity were determined. Maximal HIF-1alpha activation at 23 Torr was sustained over the postnatal shift in (Delta) PO(2) and was elevated in vivo throughout late gestation. NF-kappaB was activated by the acute postnatal DeltaPO(2) in fATII cells, becoming maximal with moderate and severe oxygenation in vitro and within 6 h of birth in vivo, declining thereafter. fATII cell and whole lung glutathione and GSH-to-GSSG ratio increased fourfold with a postnatal DeltaPO(2) and were matched by threefold activity increases in gamma-glutamylcysteine synthetase and glutathione synthase. GSH concentration depletion by L-buthionine-(S, R)-sulfoximine abrogated both HIF 1alpha and NF-kappaB activation, with HIF-1alpha showing a heightened sensitivity to GSH concentration. We conclude that O(2)-linked genetic regulation in perinatal lung epithelium is responsive to developmental changes in glutathione biosynthetic capacity. PMID- 10710522 TI - Targeted gene delivery to pulmonary endothelium by anti-PECAM antibody. AB - To achieve efficient systemic gene delivery to the lung with minimal toxicity, a vector was developed by chemically conjugating a cationic polymer, polyethylenimine (PEI), with anti-platelet endothelial cell adhesion molecule (PECAM) antibody (Ab). Transfection of mouse lung endothelial cells with a plasmid expression vector with cDNA to luciferase (pCMVL) complexed with anti PECAM Ab-PEI conjugate was more efficient than that with PEI-pCMVL complexes. Furthermore, the anti-PECAM Ab-PEI conjugate mediated efficient transfection at lower charge plus-to-minus ratios. Conjugation of PEI with a control IgG (hamster IgG) did not enhance transfection of mouse lung endothelial cells, suggesting that the cellular uptake of anti-PECAM Ab-PEI-DNA complexes and subsequent gene expression were governed by a receptor-mediated process rather than by a nonspecific charge interaction. Conjugation of PEI with anti-PECAM Ab also led to significant improvement in lung gene transfer to intact mice after intravenous administration. The increase in lung transfection was associated with a decrease compared with PEI-pCMVL with respect to circulating proinflammatory cytokine (tumor necrosis factor-alpha) levels. These results indicate that targeted gene delivery to the lung endothelium is an effective strategy to enhance gene delivery to the pulmonary circulation while simultaneously reducing toxicity. PMID- 10710523 TI - Immunopathology of a two-hit murine model of acid aspiration lung injury. AB - In a two-hit model of acid aspiration lung injury, mice were subjected to nonlethal cecal ligation and puncture (CLP). After 48 h, intratracheal (IT) acid was administered, and mice were killed at several time points. Recruitment of neutrophils in response to acid was documented by myeloperoxidase assay and neutrophil counts in bronchoalveolar lavage (BAL) fluid and peaked at 8 h post-IT injection. Albumin in BAL fluid, an indicator of lung injury, also peaked at 8 h. When the contributions of the two hits were compared, neutrophil recruitment and lung injury occurred in response to acid but were not greatly influenced by addition of another hit. Neutrophil sequestration was preceded by elevations in KC and macrophage inflammatory protein-2alpha in plasma and BAL fluid. KC levels in BAL fluid were higher and peaked earlier than macrophage inflammatory protein 2alpha levels. When KC was blocked with specific antiserum, neutrophil recruitment was significantly reduced, whereas albumin in BAL fluid was not affected. In conclusion, murine KC mediated neutrophil recruitment but not lung injury in a two-hit model of aspiration lung injury. PMID- 10710524 TI - S-nitrosoglutathione-induced decrease in calcium sensitivity of airway smooth muscle. AB - The purpose of this study was to examine whether the nitric oxide donor S nitrosoglutathione (GSNO) relaxes canine tracheal smooth muscle (CTSM) strips by decreasing Ca(2+) sensitivity [i.e., the amount of force for a given intracellular Ca(2+) concentration ([Ca(2+)](i))]. We further investigated whether GSNO decreases Ca(2+) sensitivity by altering the relationship between regulatory myosin light chain (rMLC) phosphorylation and [Ca(2+)](i) and the relationship between force and rMLC phosphorylation. GSNO (100 microM) relaxed intact CTSM strips contracted with 45 mM KCl by decreasing Ca(2+) sensitivity in comparison to control strips without significantly decreasing [Ca(2+)](i). GSNO reduced the amount of rMLC phosphorylation for a given [Ca(2+)](i) but did not affect the relationship between isometric force and rMLC phosphorylation. These results show that in CTSM strips contracted with KCl, GSNO decreases Ca(2+) sensitivity by affecting the level of rMLC phosphorylation for a given [Ca(2+)](i), suggesting that myosin light chain kinase is inhibited or that smooth muscle protein phosphatases are activated by GSNO. PMID- 10710525 TI - Mesothelial cell apoptosis is confirmed in vivo by morphological change in cytokeratin distribution. AB - Apoptosis of mesothelial cells has been demonstrated in vitro but not in vivo. To identify apoptotic pleural cells as mesothelial, we used cytokeratin as a marker and found a striking spheroid, aggregated appearance of cytokeratin in apparently apoptotic mesothelial cells. In in vitro studies, we found that the aggregated cytokeratin pattern correlated with apoptosis in primary mesothelial cells from mice, rabbits, and humans and was not seen with necrosis. In in vivo studies in mice, we then used this cytokeratin pattern to identify and quantitate apoptotic mesothelial cells. Apoptotic mesothelial cells were best harvested by pleural lavage, indicating that they were loosely adherent or nonadherent. Instillation of RPMI 1640 medium or wollastonite for 24 h induced apoptosis in 0.1 +/- 0. 1 (SE) and 1.0 +/- 0.7%, respectively, of all mesothelial cells recovered, whereas instillation of known apoptotic stimuli, crocidolite asbestos (25 microg) for 24 h or actinomycin D plus murine tumor necrosis factor-alpha for 12 h, induced apoptosis in 5. 1 +/- 0.5 and 22.4 +/- 4.5%, respectively (significantly greater than in control experiments, P < 0.05). By analysis of cytokeratin staining, mesothelial cell apoptosis has been confirmed in vivo. PMID- 10710526 TI - Role of the Na/H antiport in pH-dependent cell death in pulmonary artery endothelial cells. AB - We investigated the role of intracellular pH (pH(i)) and Na/H exchange in cell death in human pulmonary artery endothelial cells (HPAEC) following a metabolic insult (inhibition-oxidative phosphorylation, glycolysis). Metabolic inhibition in medium at pH 7. 4 decreased viability (0-15% live cells) over 6 h. Cell death was attenuated by maneuvers that decreased pH(i) and inhibited Na/H exchange (acidosis, Na/H antiport inhibitors). In contrast, cell death was potentiated by maneuvers that elevated pH(i) or increased Na/H exchange (monensin, phorbol ester treatment) before the insult. HPAEC demonstrated a biphasic pH(i) response following a metabolic insult. An initial decrease in pH(i) was followed by a return to baseline over 60 min. Maneuvers that protected HPAEC and inhibited Na/H exchange (acidosis, Na(+)-free medium, antiport inhibitors) altered this pattern. pH(i) decreased, but no recovery was observed, suggesting that the return of pH(i) to normal was mediated by antiport activation. Although Na/H antiport activity was reduced (55-60% of control) following a metabolic insult, the cells still demonstrated active Na/H exchange despite significant ATP depletion. Phorbol ester pretreatment, which potentiated cell death, increased Na/H antiport activity above the level observed in monolayers subjected to a metabolic insult alone. These results demonstrate that HPAEC undergo a pH-dependent loss of viability linked to active Na/H exchange following a metabolic insult. Potentiation of cell death with phorbol ester treatment suggests that this cell death pathway involves protein kinase C-mediated phosphorylation events. PMID- 10710527 TI - IGFBP-3 mediates TGF-beta1-induced cell growth in human airway smooth muscle cells. AB - Both insulin-like growth factor binding protein-3 (IGFBP-3) and transforming growth factor-beta (TGF-beta) have been separately shown to have cell-specific growth-inhibiting or growth-potentiating effects. TGF-beta stimulates IGFBP-3 mRNA and peptide expression in several cell types, and TGF-beta-induced growth inhibition and apoptosis have been shown to be mediated through the induction of IGFBP-3. However, a link between the growth stimulatory effects of TGF-beta and IGFBP-3-induction has not been shown. In this study, we investigated the role of IGFBP-3 in mediating TGF-beta1-induced cell growth using human airway smooth muscle (ASM) cells as our model. TGF-beta1 (1 ng/ml) treatment induced a 10- to 20-fold increase in the levels of expression of IGFBP-3 mRNA and protein. Addition of either IGFBP-3 or TGF-beta1 to the growth medium resulted in an approximately twofold increase in cell proliferation. Coincubation of ASM cells with IGFBP-3 antisense (but not sense) oligomers as well as with an IGFBP-3 neutralizing antibody (but not with control IgG) blocked the growth induced by TGF-beta1 (P < 0.001). Several IGFBP-3-associated proteins were observed in ASM cell lysates, which may have a role in the cellular responses to IGFBP-3. These findings demonstrate that IGFBP-3 is capable of mediating the growth stimulatory effect of TGF-beta in ASM cells. PMID- 10710529 TI - Effects of dexamethasone on rhinovirus infection in cultured human tracheal epithelial cells. AB - To examine the effects of glucocorticoid on rhinovirus (RV) infection, primary cultures of human tracheal epithelial cells were infected with either RV2 or RV14. Viral infection was confirmed by demonstrating that viral RNA in infected cells and viral titers of supernatants and lysates from infected cells increased with time. RV14 infection upregulated the expression of mRNA and protein of intercellular adhesion molecule-1 (ICAM-1), the major RV receptor, on epithelial cells, and it increased the production of interleukin (IL)-1beta, IL-6, IL-8, and tumor necrosis factor-alpha in supernatants. Dexamethasone reduced the viral titers of supernatants and cell lysates, viral RNA of infected cells, and susceptibility of RV14 infection in association with inhibition of cytokine production and ICAM-1 induction. In contrast to RV14 infection, dexamethasone did not alter RV2 infection, a minor group of RVs. These results suggest that dexamethasone may inhibit RV14 infection by reducing the surface expression of ICAM-1 in cultured human tracheal epithelial cells. Glucocorticoid may modulate airway inflammation via reducing the production of proinflammatory cytokines and ICAM-1 induced by rhinovirus infection. PMID- 10710528 TI - p53-independent induction of GADD45 and GADD153 in mouse lungs exposed to hyperoxia. AB - Previous studies have shown that lungs of adult mice exposed to >95% oxygen have increased terminal deoxyribonucleotidyltransferase dUTP nick end-label staining and accumulate p53, the expression of which increases in cells exposed to DNA damaging agents. The present study was designed to determine whether hyperoxia also increased expression of the growth arrest and DNA damage (GADD) gene 45 and GADD153, which are induced by genotoxic stress through p53-dependent and independent pathways. GADD proteins have been shown to inhibit proliferation and stimulate DNA repair and/or apoptosis. GADD45 and GADD153 mRNAs were not detected in lungs exposed to room air but were detected after 48 and 72 h of exposure to hyperoxia. In situ hybridization and immunohistochemistry revealed that hyperoxia increased GADD45 and GADD153 expression in the bronchiolar epithelium and GADD45 expression predominantly in alveolar cells that were morphologically consistent with type II cells. Hyperoxia also increased GADD expression in p53-deficient mice. Terminal deoxyribonucleotidyltransferase dUTP nick end-label staining of lung cells from p53 wild-type and p53-null mice exposed to hyperoxia for 48 h revealed that hyperoxia-induced DNA fragmentation was not modified by p53 deficiency. These studies are consistent with the hypothesis that hyperoxia induced DNA fragmentation is associated with the expression of GADD genes that may participate in DNA repair and/or apoptosis. PMID- 10710530 TI - Lipopolysaccharide induces functional ICAM-1 expression in rat alveolar epithelial cells in vitro. AB - Lipopolysaccharide (LPS)-induced lung inflammation is known to increase pulmonary intercellular adhesion molecule-1 (ICAM-1) expression. In the present study, L2 cells, a cell line of alveolar epithelial cells, were stimulated with LPS, and ICAM-1 expression was studied. ICAM-1 protein on L2 cells peaked at 6 (38% increase; P < 0.01) and 10 (48% increase; P < 0.001) h after stimulation with Escherichia coli and Pseudomonas aeruginosa LPS, respectively. ICAM-1 mRNA expression was markedly increased, with a peak at 2-4 (E. coli) and 4-6 (P. aeruginosa) h. Adherence assays of neutrophils to LPS-stimulated L2 cells showed a threefold increase in adherence (P < 0.001). Pretreatment of the neutrophils with anti-lymphocyte function-associated antigen-1 and anti-Mac-1 antibodies reduced adherence by 54% (P < 0.001). Analysis of immunofluorescence staining for ICAM-1 showed an exclusive apical expression of ICAM-1. These results indicate that LPS upregulates functional active ICAM-1 on the apical part of the membrane in rat pneumocytes. PMID- 10710531 TI - Internalized SP-A and lipid are differentially resecreted by type II pneumocytes. AB - Biochemical and morphological assays were developed to study surfactant protein A (SP-A) and lipid resecretion kinetics by isolated type II cells in vitro. After a 10-min uptake period with SP-A (3 microg/10(6) cells) in combination with liposomes (60 microg/10(6) cells), the cells were allowed to resecrete. After 5 min of resecretion, only 21.7 +/- 4.6% of the internalized SP-A remained intracellularly compared with 54 +/- 2.9% of the lipids. Extracellular SP-A present during the resecretion period partially inhibited resecretion (SP-A, 36% at 5 min; lipid, approximately 16% at 5 min). Lipid resecretion was also dependent on the SP-A concentration present during the uptake period. Although, as shown by confocal laser scanning microscopy, after a 10-min uptake period at 37 degrees C, most of the fluorescein isothiocyanate-labeled SP-A and rhodamine phosphatidylethanolamine-labeled lipids colocalized within the cells, after an additional 10 min of resecretion, both the strength of the fluorescence signals and the extent of colocalization had markedly decreased. These data indicate that internalized lipid and SP-A can be resecreted rapidly by type II cells, likely via different pathways. PMID- 10710532 TI - MCP-1 in pleural injury: CCR2 mediates haptotaxis of pleural mesothelial cells. AB - Pleural injury results in the death of mesothelial cells and denudation of the mesothelial basement membrane. Repair of the mesothelium without fibrosis requires proliferation and migration of mesothelial cells into the injured area. We hypothesized that monocyte chemoattractant protein-1 (MCP-1) induces proliferative and haptotactic responses in pleural mesothelial cells (PMCs) and that the MCP-1 binding receptor CCR2 mediates the pleural repair process. We demonstrate that PMCs exhibited MCP-1-specific immunostaining on injury. MCP-1 induced proliferative and haptotactic responses in PMCs. PMCs express CCR2 in a time-dependent manner. Fluorescence-activated cell sorting analysis demonstrated that interleukin (IL)-2 upregulated CCR2 protein expression in PMCs, whereas lipopolysaccharide (LPS) downregulated the response at the initial period compared with that in resting PMCs. However, the inhibitory potential of LPS was lost after 12 h and showed a similar response at 24 and 48 h. Haptotactic migration was upregulated in PMCs that were cultured in the presence of IL-2. The increased haptotactic capacity of mesothelial cells in the presence of IL-2 correlated with increased CCR2 mRNA expression. PMCs cultured in the presence of LPS showed decreased haptotactic activity to MCP-1. Blocking the CCR2 with neutralizing antibodies decreased the haptotactic response of PMCs to MCP-1. These results suggest that the haptotactic migration of mesothelial cells in response to MCP-1 are mediated through CCR2, which may play a crucial role in reepithelialization of the denuded basement membrane at the site of pleural injury and may thus contribute to the regeneration of the mesothelium during the process of pleural repair. PMID- 10710533 TI - P. carinii induces selective alterations in component expression and biophysical activity of lung surfactant. AB - Studies of Pneumocystis carinii pneumonia (PCP) suggest an important role for the surfactant system in the pathogenesis of the hypoxemic respiratory insufficiency associated with this infection. We hypothesized that PCP induces selective alterations in alveolar surfactant component expression and resultant biophysical properties. PCP was induced by intratracheal inoculation of 2 x 10(5) P. carinii organisms into C.B-17 scid/scid mice. Six weeks after inoculation, large (LA)- and small (SA)-aggregate surfactant fractions were prepared from bronchoalveolar lavage fluids and analyzed for expression of surfactant components and for biophysical activity. Total phospholipid content was significantly reduced in LA surfactant fractions from mice infected with PCP (53 +/- 15% of uninfected mice; P < 0.05). Quantitation of hydrophobic surfactant protein (SP) content demonstrated significant reductions of alveolar SP-B and SP-C protein levels in mice with PCP compared with those in uninfected mice (46 +/- 7 and 19 +/- 6%, respectively; P < 0.05 for both). The reductions in phospholipid, SP-B, and SP-C in LA fractions measured during PCP were associated with an increase in the minimum surface tension of LAs as measured by pulsating bubble surfactometer (13.1 +/- 1.1 vs. 5.4 +/- 1.8 mN/m; P < 0.05). In contrast to decreases in the hydrophobic SPs, SP-D content in the SA fraction was markedly increased (343 +/- 30% of control value; P < 0. 05) and SP-A levels in LA surfactant were maintained (93 +/- 26% of control value) during P. carinii infection. In all cases, the changes in SP content were reflected by commensurate changes in the levels of mRNA. We conclude that PCP induces selective alterations in surfactant component expression, including profound decreases in hydrophobic protein contents and resultant increases in surface tension. These changes, demonstrated in an immunologically relevant animal model, suggest that alterations in surfactant could contribute to the hypoxemic respiratory insufficiency observed in PCP. PMID- 10710534 TI - Cellular disposition of transported polyamines in hypoxic rat lung and pulmonary arteries. AB - The polyamines putrescine, spermidine (SPD), and spermine are a family of low molecular-weight organic cations essential for cell growth and differentiation and other aspects of signal transduction. Hypoxic pulmonary vascular remodeling is accompanied by depressed lung polyamine synthesis and markedly augmented polyamine uptake. Cell types in which hypoxia induces polyamine transport in intact lung have not been delineated. Accordingly, rat lung and rat main pulmonary arterial explants were incubated with [(14)C]SPD in either normoxic (21% O(2)) or hypoxic (2% O(2)) environments for 24 h. Autoradiographic evaluation confirmed previous studies showing that, in normoxia, alveolar epithelial cells are dominant sites of polyamine uptake. In contrast, hypoxia was accompanied by prominent localization of [(14)C]SPD in conduit, muscularized, and partially muscularized pulmonary arteries, which was not evident in normoxic lung tissue. Hypoxic main pulmonary arterial explants also exhibited substantial increases in [(14)C]SPD uptake relative to control explants, and autoradiography revealed that enhanced uptake was most evident in the medial layer. Main pulmonary arterial explants denuded of endothelium failed to increase polyamine transport in hypoxia. Conversely, medium conditioned by endothelial cells cultured in hypoxic, but not in normoxic, environments enabled hypoxic transport induction in denuded arterial explants. These findings in arterial explants were recapitulated in rat cultured main pulmonary artery cells, including the enhancing effect of a soluble endothelium-derived factor(s) on hypoxic induction of [(14)C]SPD uptake in smooth muscle cells. Viewed collectively, these results show in intact lung tissue that hypoxia enhances polyamine transport in pulmonary artery smooth muscle by a mechanism requiring elaboration of an unknown factor(s) from endothelial cells. PMID- 10710535 TI - Fas cross-linking induces apoptosis in human airway smooth muscle cells. AB - Hypertrophy and hyperplasia lead to excess accumulation of smooth muscle in the airways of human asthmatic subjects. However, little is known about mechanisms that might counterbalance these processes, thereby limiting the quantity of smooth muscle in airways. Ligation of Fas on the surface of vascular smooth muscle cells and nonmuscle airway cells can lead to apoptotic cell death. We therefore tested the hypotheses that 1) human airway smooth muscle (HASM) expresses Fas, 2) Fas cross-linking induces apoptosis in these cells, and 3) tumor necrosis factor (TNF)-alpha potentiates Fas-mediated airway myocyte killing. Immunohistochemistry using CH-11 anti-Fas monoclonal IgM antibody revealed Fas expression in normal human bronchial smooth muscle in vivo. Flow cytometry using DX2 anti-Fas monoclonal IgG antibody revealed that passage 4 cultured HASM cells express surface Fas. Surface Fas decreased partially during prolonged serum deprivation of cultured HASM cells and was upregulated by TNF alpha stimulation. Fas cross-linking with CH-11 antibody induced apoptosis in cultured HASM cells, and this effect was reduced by long-term serum deprivation and synergistically potentiated by concomitant TNF-alpha exposure. TNF-alpha did not induce substantial apoptosis in the absence of Fas cross-linking. These data represent the first demonstration that Fas is expressed on HASM and suggest a mechanism by which Fas-mediated apoptosis could act to oppose excess smooth muscle accumulation during airway remodeling in asthma. PMID- 10710536 TI - Construction and characterization of a chimeric receptor containing the cytoplasmic domain of MUC1 mucin. AB - MUC1 mucin is a transmembrane glycoprotein that is highly expressed in various cancer cell lines and is also present in most of the glandular epithelial cells including the airway. Although the presence of numerous phosphorylation sites in its cytoplasmic domain suggests its potential role as a receptor, the unavailability of a ligand for MUC1 mucin has limited our understanding of its function. In this paper, we tried to circumvent this problem by constructing a chimeric receptor containing the cytoplasmic domain of MUC1 mucin, which can be phosphorylated on activation. To this end, we constructed a chimeric plasmid vector (pCD8/MUC1) by replacing the extracellular and transmembrane domains of human MUC1 mucin with those of human CD8. Transient transfection of the vector into COS-7 cells resulted in expression of the chimeric receptor on the surface of the COS-7 cells as judged by immunologic assays with various antibodies as well as by fluorescence-activated cell-sorting analysis. Treatment of the transfected COS-7 cells with an anti-CD8 antibody resulted in a significant increase in phosphorylation of tyrosine moieties of the chimeric receptor. This chimeric receptor will serve as a powerful tool in elucidating the signaling mechanism as well as the functional role of MUC1 mucin in the airway. PMID- 10710537 TI - Renal physiology of the mouse. AB - As the transgenic and gene-targeting technology has become an invaluable experimental approach to study the function of gene products, the need has been expanded to assess the physiology in the mouse, which is virtually the only animal species to which that new genetic technology can apply. In this regard, renal physiologists have also received fruits of success from modern technology in several key areas, and areas are expanding in both depth and scope. PMID- 10710538 TI - Toward a comprehensive molecular model of active calcium reabsorption. AB - The fine tuning of Ca(2+) excretion in the kidney takes place in the distal nephron, which consists of the distal convoluted tubule, connecting tubule, and initial portion of the cortical collecting duct. In these segments, Ca(2+) is reabsorbed through an active transcellular pathway. The apical influx of Ca(2+) into the distal renal cell is presumably the rate-limiting step in this process, and its molecular identity has remained obscure so far. The recently discovered epithelial Ca(2+) channel (ECaC) exhibits the expected properties for being the gatekeeper in transcellular Ca(2+) reabsorption. The characteristics and potential physiological role of ECaC will be discussed in this review. Our knowledge of the mechanisms involved in the regulation of transcellular Ca(2+) transport has advanced rapidly since the development of cell models originating from distal tubular cells. Studies using these models indicate that hormones including arginine vasopressin, PGE(2), adenosine, ATP, and atrial natriuretic peptide should be considered as calciotropic hormones controlling renal Ca(2+) handling. Evidence is now beginning to emerge that the stimulating calciotropic hormones utilize new cAMP-independent pathways to stimulate Ca(2+) reabsorption. These new findings allow the development of a comprehensive and detailed model of the process of transcellular calcium transport in the kidney whereby the individual contribution of the participating transporters can now be fully appreciated. PMID- 10710539 TI - Asymmetrical targeting of type II Na-P(i) cotransporters in renal and intestinal epithelial cell lines. AB - Targeting of newly synthesized transporters to either the apical or basolateral domains of polarized cells is crucial for the function of epithelia, such as in the renal proximal tubule or in the small intestine. Recently, different sodium phosphate cotransporters have been identified. Type II cotransporters can be subdivided into two groups: type IIa and type IIb. Type IIa is predominantly expressed in renal proximal tubules, whereas type IIb is located on the intestinal and lung epithelia. To gain some insights into the polarized targeting of the type II cotransporters, we have transiently expressed type IIa and type IIb cotransporters in several epithelial cell lines: two lines derived from renal proximal cells (opossum kidney and LLC-PK(1)), one from renal distal cells (Madin Darby canine kidney), and one from colonic epithelium (CaCo-2). We studied the expression of the transporters fused to the enhanced green fluorescent protein. Our data indicate that the polarized targeting is dependent on molecular determinants most probably located at the COOH terminus of the cotransporters as well as on the cellular context. PMID- 10710540 TI - Vasopressin regulates endothelin-B receptor in rat inner medullary collecting duct. AB - Previous studies have shown that endothelin (ET) antagonizes the actions of arginine vasopressin (AVP) in the renal collecting ducts. On the other hand, the effects of AVP on ET function within the collecting ducts of the kidney have not been investigated extensively. Using isolated inner medullary collecting ducts (IMCD), we examined the possibility that a decrease in ET(B) receptor mRNA accompanied AVP-induced downregulation of ET(B) receptors. Binding studies revealed that overnight incubation of rat IMCD cells with AVP significantly reduced the maximal binding capacity (B(max)) of ET. Activation of adenylate cyclase by forskolin decreased the total ET(B) receptor density by approximately 42% but did not affect the density of ET(A) receptors. The Rp diastereoisomer of adenosine 3', 5'-cyclic monophosphothionate, Rp-cAMPS (a specific inhibitor of protein kinase A), blocked the AVP-induced reduction in ET receptor density. Using competitive PCR method, we also observed downregulation of ET(B) receptor mRNA in IMCD treated with AVP. Additional studies were done with IMCD to determine whether AVP inhibited the ET-induced accumulation of cGMP. We saw a reduction in ET-induced cGMP accumulation when IMCD was incubated overnight with AVP. This inhibition of ET-induced accumulation of cGMP was blocked by Rp-cAMPS. These results suggest that AVP regulates ET(B) receptor expression in IMCD. PMID- 10710541 TI - Role of Na(+)/H(+) exchanger NHE3 in nephron function: micropuncture studies with S3226, an inhibitor of NHE3. AB - Na(+)/H(+) exchanger NHE3 is expressed in the luminal membrane of proximal tubule and thin and thick ascending limb of Henle's loop. To further define its role, the novel NHE3 inhibitor S3226 was employed in micropuncture experiments in nephrons with superficial glomeruli of anesthetized rats. Microperfusion of proximal convoluted tubule with S3226 revealed a dose-dependent inhibition of reabsorption (IC(50) of 4-5 microM) with a maximum inhibition of 30% for fluid and Na(+). During microperfusion of Henle's loop (last superficial proximal to first superficial distal tubular loop), no effect of S3226 (10 or 30 microM) on the reabsorption of fluid or Na(+) was observed. Finally, S3226 (30 microM) left the tubuloglomerular feedback response unaltered as determined by the fall in proximal tubular stop-flow pressure in response to increasing loop of Henle perfusion rate. These studies indicate that NHE3 significantly contributes to fluid and Na(+) reabsorption in proximal convoluted tubule. NHE3 appears not to significantly contribute to fluid or Na(+) reabsorption in the loop of Henle (including the S3 segment of proximal tubule) or macula densa control of nephron filtration. PMID- 10710542 TI - Protective effects of renal ischemic preconditioning and adenosine pretreatment: role of A(1) and A(3) receptors. AB - Renal ischemia and reperfusion during aortic and renal transplant surgery result in ischemic-reperfusion injury. Ischemic preconditioning and adenosine infusion before ischemia protect against ischemic-reperfusion injury in cardiac and skeletal muscle, but these protective phenomena have not been demonstrated in the kidney. Rats were randomized to sham operation, 45-min renal ischemia, ischemic preconditioning with four cycles of 8-min renal ischemia and 5-min reperfusion followed by 45-min renal ischemia, systemic adenosine pretreatment before 45-min renal ischemia, or pretreatments with selective adenosine receptor subtype agonists or antagonists before 45-min renal ischemia. Forty-five minutes of renal ischemia followed by 24 h of reperfusion resulted in marked rises in blood urea nitrogen and creatinine. Ischemic preconditioning and adenosine pretreatment protected renal function and improved renal morphology. A(1) adenosine receptor activation mimics and A(1) adenosine antagonism blocks adenosine-induced protection. In addition, A(3) adenosine receptor activation before renal ischemia worsens renal ischemic-reperfusion injury, and A(3) adenosine receptor antagonism protects renal function. We demonstrate for the first time that rat kidneys can be preconditioned to attenuate ischemic-reperfusion injury and adenosine infusion before ischemic insult protects renal function via A(1) adenosine receptor activation. Our data suggest that an A(1) adenosine agonist and A(3) adenosine antagonist may have clinically beneficial implications where renal ischemia is unavoidable. PMID- 10710543 TI - Prostaglandin E(2) interaction with AVP: effects on AQP2 phosphorylation and distribution. AB - Prostaglandin E(2) (PGE(2)) antagonizes the action of arginine vasopressin (AVP) on collecting duct water permeability. To investigate the mechanism of this antagonism, rat renal inner medulla (IM) was incubated with the two hormones, and the phosphorylation and subcellular distribution of the water channel, aquaporin 2 (AQP2) were studied. Using a phosphorylation state-specific AQP2 antibody, we demonstrated that AVP stimulates AQP2 phosphorylation at the Ser(256) protein kinase A consensus site in a time- and dose-dependent manner. In parallel studies using a differential centrifugation technique, we demonstrated that AVP induced translocation of AQP2 from an intracellular vesicle-enriched fraction to a plasma membrane-enriched fraction. PGE(2) (10(-7) M) added after AVP (10(-8) M) did not decrease AQP2 phosphorylation but reversed AVP-induced translocation of AQP2 to the plasma membrane. Preincubation of IM with PGE(2) did not prevent the effects of AVP on AQP2 phosphorylation and trafficking. PGE(2) alone did not influence AQP2 phosphorylation and subcellular distribution. Our data indicate that 1) recruitment of AQP2 to the plasma membrane and its retrieval to a pool of intracellular vesicles may be regulated independently, 2) PGE(2) may counteract AVP action by activation of AQP2 retrieval, 3) dephosphorylation of AQP2 is not a prerequisite for its internalization. PMID- 10710544 TI - Mutations in sixth transmembrane domain of AQP2 inhibit its translocation induced by vasopression. AB - Vasopression-induced phosphorylation of serine 256 of the aquaporin-2 (AQP2) water channel triggers translocation of the protein from cystolic reservoir vesicles to the apical membrane of collecting duct principal cells. Dileucine motifs are located in the sixth transmembrane domain (6TM) of AQP2 and are known as the signal sequence for internalization, sorting from the trans-Golgi network to endosomes/lysosomes, and basolateral sorting. In this study, involvement of 6TM in vasopressin-induced translocation of the protein was investigated. A series of mutations in 6TM of AQP2 was introduced to rat cDNA and expressed in LLC-PK(1) cells. Immunofluorescence microscopy indicated that the mutant AQP2 proteins were retained in the cytoplasm after vasopressin stimulation, which actually promoted the plasma membrane expression of wild-type protein. Immunoelectron microscopy showed that the mutant AQP2 proteins reached the endosomes but did not reach the plasma membrane. These results demonstrate that 6TM has essential domains for vasopressin-induced translocation from endosomes to the plasma membrane. PMID- 10710545 TI - Role of AP2 consensus sites in regulation of rat Npt2 (sodium-phosphate cotransporter) promoter. AB - Expression of the Npt2 gene, encoding the type II sodium-dependent phosphate cotransporter, is restricted to renal proximal tubule epithelium. We have isolated a 4,740-bp fragment of the 5'-flanking sequence of the rat Npt2 gene, identified the transcription initiation site, and demonstrated that this 5' flanking sequence drives luciferase-reporter gene expression, following transfection in the proximal tubule cell-derived opossum kidney (OK) cell line but not in unrelated cell lines. Analysis of the promoter sequence revealed the presence of 10 consensus binding motifs for the AP2 transcription factor. Transient transfection assays revealed an important effect of the number of tandemly repeated AP2 sites in enhancing promoter activity. The promoter sequence also revealed a pair of inverted repeats enclosing 1,324 bp of intervening sequence and containing 8 of the total 10 AP2 consensus sites in the promoter sequence. Deletion or reversal of orientation of the distal inverted repeat resulted in marked enhancement of promoter activity. Electrophoretic mobility shift analysis revealed a distinct pattern of transcription factor binding to oligonucleotides containing AP2 sites, using nuclear extracts from OK cells, compared with unrelated cell lines. Taken together, these results suggest an important role for AP2 consensus binding sites in regulating Npt2 gene expression and suggest a mechanism of regulation mediated by the interaction of inverted repeats enclosing these sites. PMID- 10710546 TI - Regulation of stanniocalcin in MDCK cells by hypertonicity and extracellular calcium. AB - Differential display RT-PCR cloning method was applied to poly(A)(+) RNA isolated from Madin-Darby canine kidney (MDCK) cells in isotonic or hypertonic medium. A differentially expressed 360-bp PCR fragment was isolated, subcloned, sequenced, and used to screen an MDCK cDNA library constructed in lambdaZapII. A composite sequence of two overlapping cDNA clones provided 1,053 bp of sequence that was 93% identical to human stanniocalcin and corresponded to the 3'-end of the mRNA. Although the fish homolog of this hormone inhibits calcium uptake by the gill and intestine, the function of mammalian stanniocalcin remains unknown. Stanniocalcin cDNA probe hybridizes to a 4.4-kb mRNA that is induced eightfold by hypertonicity, in a manner that is dependent on medium organic osmolytes. The mRNA induction correlates with increased total cellular content of the protein and its concomitant release to the medium, consistent with secretion for autocrine or paracrine activity. Furthermore, induction of the mRNA by hypertonicity is dependent on extracellular calcium and displays a threshold phenomenon. The data suggest that kidney stanniocalcin may have a role in the adaptation of kidney cells to osmotic stress, in a manner that is extracellular calcium dependent. PMID- 10710547 TI - Mechanisms of inactivation of the action of aldosterone on collecting duct by TGF beta. AB - The purpose of these experiments was to investigate the mechanisms whereby transforming growth factor-beta (TGF-beta) antagonizes the action of adrenocorticoid hormones on Na(+) transport by the rat inner medullary collecting duct in primary culture. Steroid hormones 1) increased Na(+) transport by three- to fourfold, 2) increased the maximum capacity of the Na(+)-K(+) pump by 30-50%, 3) increased the steady-state levels of the alpha(1)-subunit of the Na(+)-K(+) ATPase by approximately 30%, and 4) increased the steady-state levels of the alpha-subunit of the rat epithelial Na(+) channel (alpha-rENaC) by nearly fourfold. TGF-beta blocked the effects of steroids on the increase in Na(+) transport and the stimulation of the Na(+)-K(+)-ATPase and pump capacity. However, there was no effect of TGF-beta on the steroid-induced increase in mRNA levels of alpha-rENaC. The effects of TGF-beta were not secondary to the decrease in Na(+) transport per se, inasmuch as benzamil inhibited the increase in Na(+) transport but did not block the increase in pump capacity or Na(+)-K(+)-ATPase mRNA. The results indicate that TGF-beta does not inactivate the steroid receptor or its translocation to the nucleus. Rather, they indicate complex pathways involving interruption of the enhancement of pump activity and activation/inactivation of pathways distal to the steroid-induced increase in the transcription of alpha-rENaC. PMID- 10710548 TI - Reabsorption of betaine in Henle's loops of rat kidney in vivo. AB - This study was designed 1) to localize and 2) to characterize betaine reabsorption from the tubular lumen in rat kidney in vivo, and 3) to test whether reabsorption is modulated by the diuretic state. [(14)C]betaine (+ [(3)H]inulin) was microperfused through the proximal convoluted tubule (PCT) and microinfused into late proximal (LP) and early distal (ED) tubules, long loops of Henle (LLH), and vasa recta of the rat in vivo et situ, and the fractional recovery of the (14)C label was determined end proximally (PCT) and in the final urine, respectively. [(14)C]betaine was not reabsorbed during ED microinfusion, whereas fractional reabsorption during LP microinfusion was 82% at 0.06 mM betaine and decreased gradually to 4.8% at 60 mM. L-Proline had lower Michaelis-Menten constant (K(m)) and sarcosine a higher K(m) than betaine. Chronic, but not acute, diuresis inhibited betaine reabsorption in Henle's loops. Fractional [(14)C]betaine reabsorption in PCT was much smaller than that during LP microinfusion. [(14)C]betaine (7.28 mM) microinfused 1) into LLH was reabsorbed to 30% and 2) into vasa recta appeared in the ipsilateral urine to a much higher extent than contralaterally. In both cases, no saturation was detected at 70 mM. We conclude that betaine is reabsorbed by mediated transport from descending limbs of short Henle's loops by a proline-preferring carrier in a diuresis modulated manner. In the deep medulla, bidirectional blood/urine betaine transport exists. PMID- 10710549 TI - Substance P dependence of endosomal fusion during bladder inflammation. AB - Urinary bladder instillation of ovalbumin into presensitized guinea pigs stimulates rapid development of local bladder inflammation. Substance P is an important mediator of this inflammatory response, as substance P antagonists largely reverse the process. Vacuolization of the subapical endosomal compartment of the transitional epithelial cells lining the bladder suggests that changes in endosomal trafficking and fusion are also part of the inflammatory response. To test directly for substance P mediation of changes in endosomal fusion, we reconstituted fusion of transitional cell endosomes in vitro using both cuvette based and flow cytometry energy transfer assays. Bladders were loaded with fluorescent dyes by a hypotonic withdrawal protocol before endosomal isolation by gradient centrifugation. Endosomal fusion assayed by energy transfer during in vitro reconstitution was both cytosol and ATP dependent. Fusion was confirmed by the increase in vesicle size on electron micrographs of fused endosomal preparations compared with controls. In inflamed bladders, dye uptake was inhibited 20% and endosomal fusion was inhibited 50%. These changes are partly mediated by the neurokinin-1 (NK1) receptor (NK1R), as 4 mg/kg of CP-96,345, a highly selective NK1 antagonist, increased fusion in inflamed bladders but had no effect on control bladders. The receptor-mediated nature of this effect was demonstrated by the expression of substance P receptor mRNA in rat bladder lumen scrapings and by the detection of the NK1R message in guinea pig subapical endosomes by Western blot analysis. The NK1Rs were significantly upregulated following induction of an inflammatory response in the bladder. These results demonstrate that 1) in ovalbumin-induced inflammation in the guinea pig bladder, in vitro fusion of apical endosomes is inhibited, showing endocytotic processes are altered in inflammation; 2) pretreatment in vivo with an NK1R antagonist blocks this inhibition of in vitro fusion, demonstrating a role for NK1R in this process; and 3) the NK1R is present in higher amounts in apical endosomes of inflamed bladder, suggesting changes in translation or trafficking of the NK1R during the inflammatory process. This suggests that NK1R can change the fusion properties of membranes in which it resides. PMID- 10710550 TI - Macula densa Na(+)/H(+) exchange activities mediated by apical NHE2 and basolateral NHE4 isoforms. AB - Functional and immunohistochemical studies were performed to localize and identify Na(+)/H(+) exchanger (NHE) isoforms in macula densa cells. By using the isolated perfused thick ascending limb with attached glomerulus preparation dissected from rabbit kidney, intracellular pH (pH(i)) was measured with fluorescence microscopy by using 2',7'-bis-(2-carboxyethyl)-5-(and -6) carboxyfluorescein. NHE activity was assayed by measuring the initial rate of Na(+)-dependent pH(i) recovery from an acid load imposed by prior lumen and bath Na(+) removal. Removal of Na(+) from the bath resulted in a significant, DIDS insensitive, ethylisopropyl amiloride (EIPA)-inhibitable decrease in pH(i). This basolateral transporter showed very low affinity for EIPA and Hoechst 694 (IC(50) = 9.0 and 247 microM, respectively, consistent with NHE4). The recently reported apical NHE was more sensitive to inhibition by these drugs (IC(50) = 0.86 and 7.6 microM, respectively, consistent with NHE2). Increasing osmolality, a known activator of NHE4, greatly stimulated basolateral NHE. Immunohistochemical studies using antibodies against NHE1-4 peptides demonstrated expression of NHE2 along the apical and NHE4 along the basolateral, membrane, whereas NHE1 and NHE3 were not detected. These results suggest that macula densa cells functionally and immunologically express NHE2 at the apical membrane and NHE4 at the basolateral membrane. These two isoforms likely participate in Na(+) transport, pH(i), and cell volume regulation and may be involved in tubuloglomerular feedback signaling by these cells. PMID- 10710551 TI - mLin-7 is localized to the basolateral surface of renal epithelia via its NH(2) terminus. AB - In Caenorhabditis elegans, the basolateral localization of the Let-23 growth factor receptor tyrosine kinase requires the expression of three genes: lin-2, lin-7, and lin-10. Mammalian homologs of these three genes have been identified, and a complex of their protein products exists in mammalian neurons. In this paper, we examine the interaction of these mammalian proteins in renal epithelia. Coprecipitation experiments demonstrated that mLin-2/CASK binds to mLin-7, and immunofluorescent labeling showed that these proteins colocalized at the basolateral surface of Madin-Darby canine kidney cells and renal epithelia. Although labeling intensity varied markedly among different renal epithelial cells, those cells strongly expressing mLin-7 also showed intense mLin-2/CASK labeling. We have also demonstrated that mLin-2/CASK binding requires amino acids 12-32 of mLin-7 and have shown that this region of mLin-7 is also necessary for the targeting of mLin-7 to the basolateral surface. Furthermore, the overexpression of mLin-2/CASK mutants in Madin-Darby canine kidney cells caused endogenous mLin-7 to mislocalize. In summary, the NH(2) terminus of mLin-7 is crucial for its basolateral localization, likely through its interaction with mLin-2/CASK. PMID- 10710552 TI - Surviving rat distal tubule bicarbonate reabsorption: effects of chronic AT(1) blockade. AB - To determine the in vivo effects of chronic ANG II type 1 (AT(1))-receptor blockade by losartan (Los) on enhanced unidirectional bicarbonate reabsorption (J(HCO(3))) of surviving distal tubules, nephrectomized rats drank either water or a solution of Los, 7 days before microperfusion. J(HCO(3)) was suppressed by 50% after Los without further reduction by 5 nM concanamycin A (Conc), suggesting that Los suppresses all Conc-sensitive H(+)-ATPase pumping. Indeed, ultrastructural analysis of A-type intercalated cells revealed a 50% reduction of H(+)-ATPase immunogold labeling of the apical plasma membrane, whereas Western blotting showed that H(+)-ATPase protein levels were also reduced by one-half by Los treatment. To identify other transporters sustaining J(HCO(3)), we perfused three inhibitors simultaneously [5-(N, N-dimethyl) amiloride hydrochloride, Conc, Schering 28080] with or without prior Los treatment: J(HCO(3)) was unchanged despite marked reduction of water reabsorption. We conclude enhanced distal tubule J(HCO(3)) of surviving nephrons is largely mediated by AT(1) receptor dependent synthesis and insertion of apical H(+)-ATPase pumps in A-type intercalated cells. PMID- 10710553 TI - Enhanced renal function in bradykinin B(2) receptor transgenic mice. AB - The tissue kallikrein-kinin system has been recognized as a paracrine and/or autocrine hormonal system that regulates arterial pressure, renal hemodynamics, and electrolyte excretion. We have created a transgenic mouse model overexpressing human bradykinin B(2) receptor, and the mice developed lifetime hypotension. With this animal model, we further analyzed the potential role of B(2) receptors in regulation of renal function. Baseline urinary excretion, urinary potassium excretion, and pH were significantly increased in transgenic mice, whereas urinary sodium excretion and serum sodium concentration were unaltered. Transgenic mice exhibited increased renal blood flow, glomerular filtration rate, and urine flow. Enhanced renal function was accompanied by significant increases in urinary nitrate/nitrite, cGMP, and cAMP levels with unaltered urinary kinin levels in transgenic mice compared with control siblings. Renal cGMP and cAMP content was also significantly increased in transgenic mice. Because the renin-angiotensin system exerts vasoconstriction buffering vasodilation of the kallikrein-kinin system, expression of renin-angiotensin components was examined by Northern blot analysis. We found a significant increase in hepatic angiotensinogen expression with no changes in renal renin and pulmonary angiotensin-converting enzyme mRNA levels in B(2) receptor transgenic mice. These studies showed that overexpression of B(2) receptors in transgenic mice resulted in hypotension and enhanced renal function through activation of nitric oxide-cGMP and cAMP signal transduction pathways. PMID- 10710554 TI - Citrate and succinate transport in proximal tubule cells. AB - Urinary citrate, which inhibits calcium nephrolithiasis, is determined by proximal reabsorption via an apical dicarboxylate transporter. Citrate is predominantly trivalent at physiological pH, but citrate(-2) is transported at the apical membrane. We now demonstrate that low-Ca solutions induce transport of citrate(-2) and succinate in opossum kidney cells. With 1.2 mM extracellular Ca, citrate uptake was pH insensitive and not competed by succinate(-2). In contrast, with low extracellular Ca, citrate uptake increased twofold, was inhibited by succinate (and other dicarboxylates), was stimulated by lowering extracellular pH (consistent with citrate(-2) transport), and increased further by lowering extracellular Mg. The effect of Ca was incrementally concentration dependent, between 0 and 1.2 mM. The effect of Ca was not simply complexation with citrate because succinate (which is complexed significantly less) was affected by Ca similarly. Incubation of cells for 48 h in a low-pH media increased citrate transport (studied at control pH) more than twofold, suggesting induction of transporters. PMID- 10710555 TI - Effects of insulin and atrial natriuretic peptide on renal tubular sodium handling in sickle cell disease. AB - We assessed the effect of insulin and atrial natriuretic peptide (ANP) on renal sodium handling in eight patients with sickle cell disease (SCD), who are characterized by loss of vasa recta and long loops of Henle, and matched control subjects. During insulin infusion (50 mU. kg(-1). h(-1)), fractional sodium excretion decreased by 0.44 +/- 0.72% (P = 0.13) in patients with SCD and by 0. 57 +/- 0.34% (P = 0.002) in control subjects, whereas fractional distal sodium reabsorption increased by 4.1 +/- 1.5% (P < 0.001) and 3.0 +/- 1.5% (P < 0.001), respectively. Low-dose (0.3 pmol. kg(-1). h(-1)) ANP infusion did not affect renal sodium handling in patients with SCD but increased fractional sodium excretion by 0.34 +/- 0.22% (P = 0.003) in control subjects. High-dose (2 microg/min) ANP increased natriuresis to a similar extent in both groups. Insulin's antinatriuretic effects predominated over the natriuretic effects of low-dose, but not high-dose, ANP. These data suggest that insulin's antinatriuretic effect is localized at a distal tubular site other than the long loops of Henle and that the long loops are involved in the natriuretic effect of low-dose ANP, possibly mediated by changes in medullary blood flow. PMID- 10710556 TI - Guidelines on preventing cardiovascular disease in clinical practice. PMID- 10710557 TI - Promoting the health of looked after children. Government proposals demand leadership and a culture change. PMID- 10710558 TI - Good practice in sterilisation. PMID- 10710560 TI - Heart disease framework aims to cut deaths in england PMID- 10710559 TI - Asthma drug delivery devices for children. PMID- 10710561 TI - Doctors fail to follow advice in giving tPA PMID- 10710562 TI - Cannabinoids might reduce spasticity in multiple sclerosis PMID- 10710563 TI - In brief PMID- 10710564 TI - Australian government attacked for tobacco funding. PMID- 10710566 TI - US launches new clinical trials database. PMID- 10710565 TI - UN condemns Australian plans for "safe injecting rooms". PMID- 10710567 TI - Patients must disclose infectious diseases, Israeli doctors say. PMID- 10710568 TI - India eradicates guinea worm disease. PMID- 10710569 TI - Malaria epidemic expected in Mozambique. PMID- 10710570 TI - Surgeon in kidney inquiry himself faces surgery suit. PMID- 10710571 TI - Scientists find a new way around blood clotting disorders PMID- 10710572 TI - Romanian national insurance cannot cover GPs' prescriptions. PMID- 10710573 TI - Coronary and cardiovascular risk estimation for primary prevention: validation of a new Sheffield table in the 1995 Scottish health survey population. AB - OBJECTIVE: To examine the accuracy of a new version of the Sheffield table designed to aid decisions on lipids screening and detect thresholds for risk of coronary heart disease needed to implement current guidelines for primary prevention of cardiovascular disease. DESIGN: Comparison of decisions made on the basis of the table with absolute risk of coronary heart disease or cardiovascular disease calculated by the Framingham risk function. The decisions related to statin treatment when coronary risk is >/=30% over 10 years; aspirin treatment when the risk is >/=15% over 10 years; and the treatment of mild hypertension when the cardiovascular risk is >/=20% over 10 years. SETTING: The table is designed for use in general practice. SUBJECTS: Random sample of 1000 people aged 35-64 years from the 1995 Scottish health survey. MAIN OUTCOME MEASURES: Sensitivity, specificity, and positive and negative predictive values of the table. RESULTS: 13% of people had a coronary risk of >/=15%, and 2. 2% a risk of >/=30%, over 10 years. 22% had mild hypertension (systolic blood pressure 140-159 mm Hg). The table indicated lipids screening for everyone with a coronary risk of >/=15% over 10 years, for 95% of people with a ratio of total cholesterol to high density lipoprotein cholesterol of >/=8.0, but for <50% with a coronary risk of <5% over 10 years. Sensitivity and specificity were 97% and 95% respectively for a coronary risk of >/=15% over 10 years; 82% and 99% for a coronary risk of >/=30% over 10 years; and 88% and 90% for a cardiovascular risk of >/=20% over 10 years in mild hypertension. CONCLUSION: The table identifies all high risk people for lipids screening, reduces screening of low risk people by more than half, and ensures that treatments are prescribed appropriately to those at high risk, while avoiding inappropriate treatment of people at low risk. PMID- 10710574 TI - Using the Framingham model to predict heart disease in the United Kingdom: retrospective study. PMID- 10710575 TI - Should treatment recommendations for lipid lowering drugs be based on absolute coronary risk or risk reduction? PMID- 10710576 TI - Allergy associated with ciprofloxacin. PMID- 10710577 TI - Using thresholds based on risk of cardiovascular disease to target treatment for hypertension: modelling events averted and number treated. AB - OBJECTIVE: To estimate the impact of using thresholds based on absolute risk of cardiovascular disease to target drug treatment to lower blood pressure in the community. DESIGN: Modelling of three thresholds of treatment for hypertension based on the absolute risk of cardiovascular disease. 5 year risk of disease was estimated for each participant using an equation to predict risk. Net predicted impact of the thresholds on the number of people treated and the number of disease events averted over 5 years was calculated assuming a relative treatment benefit of one quarter. SETTING: Auckland, New Zealand. PARTICIPANTS: 2158 men and women aged 35-79 years randomly sampled from the general electoral rolls. MAIN OUTCOME MEASURES: Predicted 5 year risk of cardiovascular disease event, estimated number of people for whom treatment would be recommended, and disease events averted over 5 years at different treatment thresholds. RESULTS: 46 374 (12%) Auckland residents aged 35-79 receive drug treatment to lower their blood pressure, averting an estimated 1689 disease events over 5 years. Restricting treatment to individuals with blood pressure >/=170/100 mm Hg and those with blood pressure between 150/90-169/99 mm Hg who have a predicted 5 year risk of disease >/=10% would increase the net number for whom treatment would be recommended by 19 401. This 42% relative increase is predicted to avert 1139/1689 (68%) additional disease events overall over 5 years compared with current treatment. If the threshold for 5 year risk of disease is set at 15% the number recommended for treatment increases by <10% but about 620/1689 (37%) additional events can be averted. A 20% threshold decreases the net number of patients recommended for treatment by about 10% but averts 204/1689 (12%) more disease events than current treatment. CONCLUSIONS: Implementing treatment guidelines that use treatment thresholds based on absolute risk could significantly improve the efficiency of drug treatment to lower blood pressure in primary care. PMID- 10710579 TI - Risk assessment in primary prevention of coronary heart disease: randomised comparison of three scoring methods. PMID- 10710578 TI - Evaluation of computer based clinical decision support system and risk chart for management of hypertension in primary care: randomised controlled trial. AB - OBJECTIVES: To investigate the effect of a computer based clinical decision support system and a risk chart on absolute cardiovascular risk, blood pressure, and prescribing of cardiovascular drugs in hypertensive patients. DESIGN: Cluster randomised controlled trial. SETTING: 27 general practices in Avon. PARTICIPANTS: 614 patients aged between 60 and 79 years with high blood pressure. INTERVENTIONS: Patients were randomised to computer based clinical decision support system plus cardiovascular risk chart; cardiovascular risk chart alone; or usual care. MAIN OUTCOME MEASURES: Percentage of patients in each group with a five year cardiovascular risk >/=10%, systolic blood pressure, diastolic blood pressure, prescribing of cardiovascular drugs. RESULTS: Patients in the computer based clinical decision support system and chart only groups were no more likely to have cardiovascular risk reduced to below 10% than patients receiving usual care. Patients in the computer based clinical decision support group were more likely to have a cardiovascular risk >/=10% than chart only patients, odds ratio 2.3 (95% confidence interval 1.1 to 4.8). The chart only group had significantly lower systolic blood pressure compared with the usual care group (difference in means -4.6 mm Hg (95% confidence interval -8.4 to -0.8)). Reduction of diastolic blood pressure did not differ between the three groups. The chart only group were twice as likely to be prescribed two classes of cardiovascular drugs and over three times as likely to be prescribed three or more classes of drugs compared with the other groups. CONCLUSIONS: The computer based clinical decision support system did not confer any benefit in absolute risk reduction or blood pressure control and requires further development and evaluation before use in clinical care can be recommended. Use of chart guidelines are associated with a potentially important reduction in systolic blood pressure. PMID- 10710581 TI - Mother's milk PMID- 10710580 TI - Discovery of a temple of AEsculapius PMID- 10710582 TI - Acute ischaemic stroke. PMID- 10710583 TI - Unnecessary surgical treatment of fungal kerions in children. PMID- 10710584 TI - ABC of arterial and venous disease. Non-invasive methods of arterial and venous assessment. PMID- 10710585 TI - Estimating cardiovascular risk for primary prevention: outstanding questions for primary care. PMID- 10710586 TI - Joint British recommendations on prevention of coronary heart disease in clinical practice: summary. British Cardiac Society, British Hyperlipidaemia Association, British Hypertension Society, British Diabetic Association. PMID- 10710587 TI - A very peculiar dream PMID- 10710588 TI - Updated New Zealand cardiovascular disease risk-benefit prediction guide. PMID- 10710590 TI - Health minister clarifies care standards bill. PMID- 10710589 TI - Managing self poisoning. Gastric lavage is perhaps more important in developing countries. PMID- 10710591 TI - Differentiating between audit and research. Undue protection of patient confidentiality jeopardises both research and audit. PMID- 10710592 TI - Authors take issue with commentary on their paper. PMID- 10710593 TI - Mortality prediction model is preferable to APACHE. PMID- 10710594 TI - Development and evaluation committees' methods for appraising new drugs. Committee failed to meet aims in producing report on low molecular weight heparins. PMID- 10710595 TI - Osteoporosis and coeliac disease. Screening all patients with osteoporosis would be inappropriate. PMID- 10710596 TI - Hospital of the future. Health economics may be misleading. PMID- 10710597 TI - Seasonality of birth in children with diabetes. Results of various studies differ. PMID- 10710598 TI - Rationing certainly exists in treatment for cancer. PMID- 10710599 TI - Distress symptoms may be easy to miss. PMID- 10710600 TI - Centralisation of cancer services in rural areas has disadvantages. PMID- 10710601 TI - Obituaries PMID- 10710602 TI - BMA wants open debate on regulation of doctors PMID- 10710604 TI - Oxford handbook of tropical medicine PMID- 10710603 TI - The nazi war on cancer PMID- 10710606 TI - Risk of cardiovascular disease PMID- 10710605 TI - Some talk of genocide PMID- 10710607 TI - "Absolute" is inappropriate for quantitative risk estimation PMID- 10710609 TI - Being smarter about preventing heart disease PMID- 10710608 TI - Seize the day PMID- 10710610 TI - New table for assessing coronary risk has high specificity and sensitivity PMID- 10710611 TI - Framingham model for risk of heart disease fits a british population well PMID- 10710613 TI - Risk based guidelines may improve hypertension treatment PMID- 10710612 TI - Hypertensive patients don't gain from computerised risk assessment PMID- 10710614 TI - New zealand and british risk charts are easier to use in primary care than sheffield table PMID- 10710615 TI - NHS needs to implement coronary risk reduction for the 10% of population at high risk PMID- 10710616 TI - [Ultrastructural characteristics of the collagen resorption in the cirrhotic liver]. PMID- 10710618 TI - [Correlation between blood pressure and parameters of systemic hemodynamics during pressor and depressor effects of ventral regions of the medulla oblongata surface]. PMID- 10710617 TI - [Quantitative evaluation of the role of cholinergic, gastrin, and histamine regulation of pepsinogen production in the stomach of anesthesized rats]. PMID- 10710619 TI - [Formation of SiO2-induced granulomas in mice of various strains]. PMID- 10710620 TI - [Changes in gluco- and mineralocorticoid adrenal functions in rats with inherited arterial hypertension during experimental myocardial infarction]. PMID- 10710621 TI - [Effect of lead administration to pregnant rats on the brain of their offspring (delayed effect)]. PMID- 10710622 TI - [Changes in phospholipase A2 activity and lipid peroxidation in endotoxicosis during experimental peritonitis]. PMID- 10710623 TI - [Role of the sympathetic nervous system in the protective effect of the selective kappa-opiate agonist dynorphin A(1-13) on frequency of cardiac arrhythmias during myocardial ischemia]. PMID- 10710624 TI - [Effect of the extremely low dose of thyroliberin on microviscosity of the lipid component of biological membranes]. PMID- 10710625 TI - [Effect of the isoosmotic solution with decreased sodium level on mitochondria of cultured granule cells from the cerebellum]. PMID- 10710626 TI - [Stabilization of ion homeostasis by gangliosides during exposure of the cerebral cortex synaptosomes to toxic concentrations of glutamate]. PMID- 10710627 TI - [Comparative study of the antioxidant effect of solasulfone and alpha tocopherol]. PMID- 10710628 TI - [Quantitative analysis of synaptophysin (p38) in the brain of the second generation offspring from male rats with long-term morphine intoxication]. PMID- 10710629 TI - [Changes in lactate and succinate dehydrogenase activity in blood lymphocytes in male mice with aggressive and submissive types of behavior]. PMID- 10710630 TI - [Disturbed functioning of enzyme systems of the microsomal oxidation, glucuro- and glutathione conjugation of xenobiotics in the liver of rats intoxicated by deoxycholate and their correction]. PMID- 10710631 TI - [Correction by verapamil of the disturbed neuroregulatory systems during early atherogenesis]. PMID- 10710632 TI - [Effect of glucocorticoids and gestagens on the glutathione redox system in the rat skin during dermatitis]. PMID- 10710633 TI - [Mechanism of the cardioprotective effect of estradiol]. PMID- 10710634 TI - [Phospholipase A2 activity in the rat skin tissues in the norm and under hormone exposure]. PMID- 10710635 TI - [Kinetic characteristics of the energy production system in the rat brain during posthypoxic encephalopathy and its correction by Bergenia cressifolia extract]. PMID- 10710636 TI - [Fast clearance of the rhesus-positive erythrocytes by monoclonal anti-rhesus antibodies--an insufficient condition for effective prophylaxis of rhesus sensitization]. PMID- 10710637 TI - [Mutations in codon 61 in H-ras gene in liver tumors induced by strong and week carcinogens in mice]. PMID- 10710638 TI - [Comparative study of the reproduction of Sidnbis virus sensitive and resistant to adamantane derivatives]. PMID- 10710639 TI - [Androgen metabolism in malignant and benign bone tumors]. PMID- 10710640 TI - [Effect of the intensive physical load on immediate type allergy development]. PMID- 10710641 TI - [Characteristics of biological rhythms of tissue basophils in various regions of the rat cerebral cortex]. PMID- 10710642 TI - [[Effects of fractions of the cerebrospinal fluid from patients with drug addiction treated by liquor sorption on the behavior of rats-recipients]. PMID- 10710643 TI - [Changes in Purkinje cells of the cerebellum during postresuscitation period: morphometric and ultrastructural analysis]. PMID- 10710644 TI - [Dependence of responses of the mouth mucosa on physico-chemical characteristics of the surface of the plate denture made of acrylic resins]. PMID- 10710645 TI - [Cytologic analysis of the mouth mucosa surface]. PMID- 10710647 TI - Rb4Zr3Te16, a one-dimensional zirconium telluride synthesized from molten salt PMID- 10710648 TI - Potassium oxoaluminate antimonate(III), K2 PMID- 10710646 TI - [Cell ultrastructure in fish and amphibia livers during catabolism of dying erythrocytes]. PMID- 10710649 TI - Dimeric copper(II) 3,3-dimethylbutyrate adducts with ethanol, 2-methylpyridine, 3 methylpyridine, 4-methylpyridine and 3,3-dimethylbutyric acid PMID- 10710650 TI - Dimeric copper(II) trimethylsilylacetate adducts with pyridine, 2-methylpyridine, 3-methylpyridine and quinoline, and a polymeric copper(II) trimethylsilylacetate PMID- 10710651 TI - H...Br hydrogen bonding in trans-bis(acetonitrile-N)tetraaquacobalt(II) dibromide PMID- 10710653 TI - [Fe] PMID- 10710652 TI - Bis(mu-succinato-O,O':O",O"')bis- PMID- 10710654 TI - Disorder in the lactato group of (lactato-O,O')bis(triphenylphosphine-P)silver(I) corroborated by 31P two-dimensional CPCOSY NMR PMID- 10710655 TI - Bis(N,N'-dimethylimidazol-2-ylidene)mercury chlorotriiodomercury dimethyl sulfoxide solvate PMID- 10710656 TI - delta-1,4,7,10-tetrakis PMID- 10710657 TI - Bis PMID- 10710659 TI - fac-tricarbonyl PMID- 10710658 TI - The cyclic vanadylacetate (C24H20P) PMID- 10710660 TI - Copper(I) and copper(II) complexes of an ethylene cross-bridged cyclam PMID- 10710661 TI - Potassium bis(ethylenediamine)copper(II) ferricyanide PMID- 10710662 TI - mu-(4,4'-bipyridine)-N:N'-bis PMID- 10710663 TI - Bis(thiosemicarbazide-S,N)zinc(II) dinitrate. PMID- 10710664 TI - 7-diphenyl PMID- 10710665 TI - Bis(eta 5-isopropyltetramethylcyclopentadienyl)(tetrahydroborato-kappa 3 H,H',H")(tetrahydrofuran-O)-samarium(III): a lanthanidocene complex with a tridentate tetrahydroborato ligand PMID- 10710666 TI - Compositional disorder in trans- PMID- 10710667 TI - [] PMID- 10710668 TI - Polysulfonylamines. CXX. Bis(tetrahydrothiophene-S)gold(I) benzene-1,2 disulfonimidate PMID- 10710669 TI - Bis(1,2-diaminobenzene-N)bis(1,1,1,5,5,5-hexafluoropentane-2,4-dionato- O,O')iron(II), -cobalt(II) and -nickel(II) at low temperature PMID- 10710670 TI - A 1:2 complex of mercury(II) chloride with 1,3-imidazole-2-thione. PMID- 10710672 TI - (-)-sparteine copper(II) diacetate PMID- 10710671 TI - cis- and trans-dichloro(3,6-dihydro-1,2-oxazine-N)(dimethyl sulfoxide-S)- platinum(II). PMID- 10710673 TI - Two 9-alkylthiophenanthrenes at 193 K (alkyl = dodecyl and tetradecyl) PMID- 10710674 TI - Di-9-anthryl disulfide at 193 K PMID- 10710675 TI - (-)-(1R,2S,2'R,5R)-2-(1-hydroxyprop-2-yl)-5-methylcyclohexanol. PMID- 10710676 TI - (1R,2R)-2-(4-methylenecyclohex-2-enyl)propyl (1R,4S)-camphanate PMID- 10710677 TI - 3 beta-acetoxy-5 alpha,6 beta-dihydroxybisnorcholanic acid 22-->16 lactone PMID- 10710678 TI - 5-amino-4-(4-diethylaminophenyl)-2-phenyl-7-(pyrrolidin-1-yl)-1,6 -naphthyridine 8-carbonitrile. PMID- 10710679 TI - Nimonol and 6-oxonimonol. PMID- 10710680 TI - Two (Z)-N-aryl-3-benzylideneisoindolin-1-ones. PMID- 10710681 TI - trans,trans-2-cyano-5-(4-methoxyphenyl)penta-2,4-dienethioamide PMID- 10710682 TI - 2,4- PMID- 10710683 TI - 4,4,4',4',7,7'-hexamethyl-2,2'-spirobichroman PMID- 10710684 TI - 5'-allyl-2'-benzoyloxy-3'-methoxy-4-nitroazobenzene PMID- 10710686 TI - 9-phenyl-3,4,4a,9a-tetrahydrotriptycene and 9-phenyl-1,2,3,4,4a,9a hexahydrotriptycene PMID- 10710685 TI - 2-trichloromethylthio-1H-isoindole-1,3(2H)-dione (Folpet). PMID- 10710687 TI - 11-methylpyrido[2,3-b]acridine-5,12-dione. PMID- 10710688 TI - Very short F-H...F hydrogen bond in L-argininium fluoride hydrogen fluoride PMID- 10710689 TI - Racemic isopropyl 1,4-dihydro-2,6-dimethyl-4-(6-methyl-2-pyridyl)-3- nitropyridine-5-carboxylate hemibenzene solvate. PMID- 10710690 TI - Ammonium hydrogen squarate-water (3/2) PMID- 10710691 TI - Cyclic lipoundecapeptide tensin from Pseudomonas fluorescens strain 96.578. PMID- 10710692 TI - 5-ethyl-2'-deoxy-4'-thiouridine (R)-S-oxide monohydrate. PMID- 10710693 TI - alpha-haloketal (1'R,4S,5S)-dimethyl 2-(1'-bromoethyl)-2-(3,4-dimethoxyphenyl) 1,3-dioxolane-4,5-dicarboxylate PMID- 10710694 TI - Chlorotrimethylsilane PMID- 10710695 TI - 2,2'-diselenobis(4,4-diphenylcyclohexa-2,5-dienone) PMID- 10710696 TI - Definitions in forensics and radiology. PMID- 10710697 TI - Forensic radiology in historical perspective. PMID- 10710699 TI - The medicine goes down PMID- 10710698 TI - Scope of forensic radiology. PMID- 10710700 TI - Jailed youths have HIV smarts PMID- 10710701 TI - The impact of Mississippi's mandatory delay law on the timing of abortion. AB - CONTEXT: Mississippi mandates that a woman seeking an abortion must first receive, in person, information about the fetus and alternatives to abortion. She must then wait at least 24 hours before having an abortion. It is not clear how such mandatory delay requirements affect the timing during pregnancy at which abortion occurs. METHODS: The data for analysis, from the Mississippi Department of Health, are 34,748 abortions obtained by residents in the six-year period surrounding the law's enactment in August 1992 (i.e., from August 1989 through July 1995). The records were stratified by location of the nearest provider, so abortions to women whose nearest provider is in-state comprised the "treatment group" (N = 28,975), while abortions to women whose nearest provider is in a neighboring state with no such law comprised the "control group" (N = 5,773). Probit regressions were used to assess effects on the likelihood of a second trimester abortion, and ordinary least-squares regressions were used to determine effects on gestational age at the time of the abortion. RESULTS: After enactment of the law, the proportion of second-trimester procedures increased by 53% (from 7.5% of abortions to 11.5%) among women whose closest provider is in-state, but it increased by only 8% (from 10.5% to 11.3%) among women whose closest provider is out-of-state. And although the overall abortion rate declined among women in the treatment group over the period (from 11.3 procedures per 1,000 women aged 15 44 to 9.9), the rate of second-trimester procedures increased among these women (from 0.8 per 1,000 women aged 15-44 to 1.1). The law was independently associated with delays in obtaining an abortion: Once the law went into effect and net of all covariates, the proportion of second-trimester abortions increased by nearly three percentage points more among women living closest to an in-state provider than among those living closest to an out-of-state provider. The law increased the mean gestational age of the fetus at the time of the procedure by approximately four days. Women who live closest to abortion providers in other states were relatively unaffected by the law. CONCLUSIONS: The proportion of abortions performed later in pregnancy will probably increase if more states impose mandatory delay laws with in-person counseling requirements. PMID- 10710702 TI - Adolescent pregnancy and childbearing: levels and trends in developed countries. AB - CONTEXT: Adolescent pregnancy occurs in all societies, but the level of teenage pregnancy and childbearing varies from country to country. A cross-country analysis of birth and abortion measures is valuable for understanding trends, for identifying countries that are exceptional and for seeing where further in-depth studies are needed to understand observed patterns. METHODS: Birth, abortion and population data were obtained from various sources, such as national vital statistics reports, official statistics, published national and international sources, and government statistical offices. Trend data on adolescent birthrates were compiled for 46 countries over the period 1970-1995. Abortion rates for a recent year were available for 33 of the 46 countries, and data on trends in abortion rates could be gathered for 25 of the 46 countries. RESULTS: The level of adolescent pregnancy varies by a factor of almost 10 across the developed countries, from a very low rate in the Netherlands (12 pregnancies per 1,000 adolescents per year) to an extremely high rate in the Russian Federation (more than 100 per 1,000). Japan and most western European countries have very low or low pregnancy rates (under 40 per 1,000); moderate rates (40-69 per 1,000) occur in Australia, Canada, New Zealand and a number of European countries. A group of five countries--Belarus, Bulgaria, Romania, the Russian Federation and the United States--have pregnancy rates of 70 or more per 1,000. The adolescent birthrate has declined in the majority of industrialized countries over the past 25 years, and in some cases has been more than halved. Similarly, pregnancy rates in 12 of the 18 countries with accurate abortion reporting showed declines. Decreases in the adolescent abortion rate, however, were less prevalent. CONCLUSIONS: The trend toward lower adolescent birthrates and pregnancy rates over the past 25 years is widespread and is occurring across the industrialized world, suggesting that the reasons for this general trend are broader than factors limited to any one country: increased importance of education, increased motivation of young people to achieve higher levels of education and training, and greater centrality of goals other than motherhood and family formation for young women. PMID- 10710703 TI - Sexually transmitted diseases among adolescents in developed countries. AB - CONTEXT: Sexually transmitted diseases (STDs) are responsible for a variety of health problems, and can have especially serious consequences for adolescents and young adults. An international comparison of levels and trends in STDs would be useful to identify countries that are relatively successful in controlling the incidence of STDs, as a first step toward improving policies and programs in countries with high or growing STD incidence. METHODS: Incidence data for the past decade on three common bacterial STDs--syphilis, gonorrhea and chlamydia- were obtained for as many as 16 developed countries from official statistics, published national sources or scientific articles, and unpublished government data. Rates of incidence per 100,000 were calculated for adolescents, for young adults and for the total population. (These estimates should be considered conservative, because STDs commonly are underreported.) RESULTS: The incidence of these three STDs has generally decreased over the last decade, both in the general population and among adolescents. However, the Russian Federation is an important exception: Syphilis has risen dramatically in the 1990s. Except in the Russian Federation and Romania, the syphilis rate in the mid-1990s was quite low, with rates of less than seven reported cases per 100,000 teenagers in most developed countries. Gonorrhea incidence is many times higher than that of syphilis in several countries, and this disease disproportionately affects adolescents and young adults. Gonorrhea rates among adolescents can be as high as 600 per 100,000 (in the Russian Federation and the United States), although in many countries the reported rate among teenagers is below 10 per 100,000. In all countries with good reporting, chlamydia incidence is extremely high among adolescents (between 563 and 1,081 cases per 100,000). The reported incidence of all three STDs is generally higher among female teenagers than among males of the same age; this is especially true for chlamydia. CONCLUSION: Prevention programs, active screening strategies and better access to STD diagnosis and treatment services, especially for adolescents and young adults, are necessary to reduce the incidence and the burden of STDs among young people. PMID- 10710704 TI - Older, but not wiser: how men get information about AIDS and sexually transmitted diseases after high school. AB - CONTEXT: As they reach adulthood, young men are less likely to use condoms and are at increased risk for exposure to AIDS and other sexually transmitted diseases (STDs). Little is known about which prevention efforts reach men in their 20s. METHODS: Longitudinal data from the 1988, 1990-1991 and 1995 waves of the National Survey of Adolescent Males are used to identify sources of information about AIDS and STDs among 1,290 young men aged 22-26. Information receipt from four main sources, the topics covered by each source and the personal characteristics associated with getting more information are all explored. RESULTS: Twenty-two percent of men surveyed discussed disease prevention topics with a health provider in the last year, 48% attended a lecture or read a brochure, 51% spoke to a partner, friend or family member, and 96% heard about AIDS or STDs from the media (e.g., television advertisements, radio or magazine). Excluding media sources, 30% of young men reported getting no STD or AIDS prevention messages in the last year. Being black or Hispanic, having had a physical exam or an AIDS test in the last year, and having discussions about AIDS or STDs with parents or a health care provider in the past were associated with receiving more information. CONCLUSIONS: Although young men who are at higher risk for STD or HIV infection are more likely than other young men to get information about disease prevention, young adult men are much less likely than adolescents to receive AIDS or STD prevention education. More prevention efforts need to be aimed at young adults. PMID- 10710705 TI - Partner effects on a woman's intention to conceive: 'not with this partner'. AB - CONTEXT: Current definitions of pregnancy intention that are useful at aggregate levels are weak at the individual level. This is especially true in social contexts where childbearing and pregnancy often occur within casual or transient relationships. METHODS: Extensive data on lifetime partnerships and sexual behaviors, including pregnancies and births, from 250 low-income women who had experienced a total of 839 pregnancies are used to explore correlates of intention to conceive, as well as the extent to which women attribute their intentions to a current partnership. RESULTS: Some 57% of reported pregnancies were unintended. Overall, 21% of the women had not wished to conceive at least one of their pregnancies with the partner who impregnated them; that proportion rose to 33% among women who had had only unintended pregnancies. Even among women who had had no unintended pregnancies, 18% had had at least one conception that they had not wanted with their partner at the time of conception. Women were less likely to say they had not wanted to conceive with a particular partner if they were living with that partner than if they were not. The likelihood of not having wanted a pregnancy with a given partner rose with the lifetime number of serious partners. Pregnancies that were not wanted with a particular partner were more than twice as likely to end in abortion as were those that were (33% vs. 14%). CONCLUSIONS: Among these women, the desire to avoid childbearing relates more to the couple involved in the conception than to abstract notions of completed family size. It would therefore be useful to include items pertaining to partner relationships in future studies of pregnancy intention. PMID- 10710706 TI - The skeletal function of non-genic nuclear DNA: new evidence from ancient cell chimaeras. AB - DNA can be divided functionally into three categories: (1) genes--which code for proteins or specify non-messenger RNAs; (2) semons--short specific sequences involved in the replication, segregation, recombination or specific attachments of chromosomes, or chromosome regions (e.g. loops or domains) or selfish genetic elements; (3) secondary DNA--which does not function by means of specific sequences. Probably more than 90% of DNA in the biosphere is secondary DNA present in the nuclei of plants and phytoplankton. The amount of genic DNA is related to the complexity of the organism, whereas the amount of secondary DNA increases proportionally with cell volume, and not with complexity. This correlation is most simply explained by the skeletal DNA hypothesis, according to which nuclear DNA functions as the basic framework for the assembly of the nucleus and the total genomic DNA content functions (together with relatively invariant folding rules) in determining nuclear volumes. Balanced growth during the cell cycle requires the cytonuclear ratio to be basically constant, irrespective of cell volume; thus nuclear volumes, and therefore the overall genome size, have to be evolutionarily adjusted to changing cell volumes for optimal function. Bacteria, mitochondria, chloroplasts and viruses have no nuclear envelope; and the skeletal DNA hypothesis simply explains why secondary DNA is essentially absent from them but present in large cell nuclei. Hitherto it has been difficult to refute the alternative hypothesis that nuclear secondary DNA (whether 'junk' or selfish DNA) accumulates merely by mutation pressure, and that selection for economy is not strong enough to eliminate it, whereas accumulation in mitochondria and plastids is prevented by intracellular replicative competition between their multiple genomes. New data that discriminate clearly between these explanations for secondary DNA come from cryptomonads and chlorarachneans, two groups of algae that originated independently by secondary symbiogenesis (i.e., the merger of two radically different eukaryote cells) several hundred million years ago. In both groups the nucleus and plasma membrane of the former algal symbiont persist as the nucleomorphs and periplastid membrane, respectively. The fact that nucleomorphs have undergone a 200- to 1000-fold reduction in genome size and have virtually no secondary DNA shows that selection against non-functional nuclear DNA is strong enough to eliminate it very efficiently; therefore, the large amounts of secondary DNA in the former host nuclei of these chimaeras, and in nuclei generally, must be being maintained by positive selection. The divergent selection for secondary DNA in the nucleus and against it in nucleomorphs is readily explicable by the skeletal DNA hypothesis, given the different spectrum of gene functions that it encodes. PMID- 10710707 TI - DNA repeats in the human genome. AB - Repetitive DNA sequences, interspersed throughout the human genome, are capable of forming a wide variety of unusual DNA structures with simple and complex loopfolding patterns. The hairpin formed by the fragile X repeat, (CCG)n, and the bipartite triplex formed by the Friedreich's ataxia repeat, (GAA)n/(TTC)n, show simple loopfolding. On the other hand, the doubly folded hairpin formed by the human centromeric repeat, (AATGG)n, the hairpin G-quartet formed by (TTAGGG)n at the 3' telomere overhang, and the hairpin G-quartet, and hairpin C+.C paired i motif formed by the insulin minisatellite, [formula: see text] show multiple and complex loopfolding. We have performed high resolution nuclear magnetic resonance (NMR) spectroscopy and in vitro replication to show that unique base-pairing and loopfolding render stability to these unusual structures under physiological conditions. The formation of such stable structures offers a mechanism of unwinding which is advantageous during transcription. For example, the formation of the hairpin G-quartet, and hairpin C+.C paired i-motif upstream of the insulin gene may facilitate transcription. These unusual DNA structures also provide unique 'protein recognition motifs' quite different from a Watson-Crick double helix. For example, the hairpin G-quartet formed by (TTAGGG)n at the 3' telomere overhang is specifically recognized and stabilized by the human repair protein, Ku70/Ku80 hetero-dimer, which may be important in the stability of the telomere. However, the formation of the same unusual DNA structures during replication is likely to cause instability in the lengths of the DNA repeats. If the altered (generally expanded) length enhances the probability of the unusual structure during the next cycle of replication, it further increases the instability of the repeat causing a 'dynamic mutation'. In fact, NMR and in vitro replication studies show that the longer the repeat length the higher is the probability of hairpin formation by the fragile X repeat, (CCG)n. In addition, the hairpin of the fragile X repeat, upstream of the FMR-1 gene, is more susceptible to CpG methylation than its duplex thereby leading to methyl-directed suppression of transcription. Thus, the selective advantage of the unusual structures formed by the DNA repeats in the regulation of gene expression may be offset by the genomic instability caused by the same structures during replication. The repeat number is a critical parameter that helps maintain a balance between the advantage gained from an unusual structure during gene expression and the disadvantage posed by the same structure during replication. PMID- 10710709 TI - Superhelical DNA studied by solution scattering and computer models. AB - We present here recent results on the structure of superhelical DNA and its changes with salt concentration between 0.01 and 1.5 M NaCl. Scattering curves of two different superhelical DNAs were determined by static light scattering. The measured radii of gyration do not change significantly with salt concentration. Small-angle neutron scattering, together with calculations from a Monte Carlo model, allows to determine the superhelix diameter. Measured and simulated scattering curves agreed almost quantitatively. Experimentally we find that the diameter decreases from 16.0 +/- 0.9 nm at 10 mM to 9.0 +/- 0.7 nm at 100 mM NaCl. The superhelix diameter from the simulated conformations decreased from 18.0 +/- 1.5 nm at 10 mM to 9.4 +/- 1.5 nm at 100 mM NaCl. At higher salt concentrations up to 1.5 M NaCl, the diameter stays constant at 9 nm. PMID- 10710708 TI - Left-handed Z-DNA: structure and function. AB - Z-DNA is a high energy conformer of B-DNA that forms in vivo during transcription as a result of torsional strain generated by a moving polymerase. An understanding of the biological role of Z-DNA has advanced with the discovery that the RNA editing enzyme double-stranded RNA adenosine deaminase type I (ADAR1) has motifs specific for the Z-DNA conformation. Editing by ADAR1 requires a double-stranded RNA substrate. In the cases known, the substrate is formed by folding an intron back onto the exon that is targeted for modification. The use of introns to direct processing of exons requires that editing occurs before splicing. Recognition of Z-DNA by ADAR1 may allow editing of nascent transcripts to be initiated immediately after transcription, ensuring that editing and splicing are performed in the correct sequence. Structural characterization of the Z-DNA binding domain indicates that it belongs to the winged helix-turn-helix class of proteins and is similar to the globular domain of histone-H5. PMID- 10710710 TI - Micro-mechanical measurement of the torsional modulus of DNA. AB - The torsional modulus C of DNA is determined from the difference between the work of stretching a single overwound molecule and the work done in stretching one underwound by the same number of turns. The value obtained C/kBT = 86 +/- 10 nm is within the range (75 +/- 25 nm) estimated by more indirect methods. PMID- 10710711 TI - Sequence-dependent modelling of local DNA bending phenomena: curvature prediction and vibrational analysis. AB - Bending is a local conformational micropolymorphism of DNA in which the original B-DNA structure is only distorted but not extensively modified. Bending can be predicted by simple static geometry models as well as by a recently developed elastic model that incorporate sequence dependent anisotropic bendability (SDAB). The SDAB model qualitatively explains phenomena including affinity of protein binding, kinking, as well as sequence-dependent vibrational properties of DNA. The vibrational properties of DNA segments can be studied by finite element analysis of a model subjected to an initial bending moment. The frequency spectrum is obtained by applying Fourier analysis to the displacement values in the time domain. This analysis shows that the spectrum of the bending vibrations quite sensitively depends on the sequence, for example the spectrum of a curved sequence is characteristically different from the spectrum of straight sequence motifs of identical basepair composition. Curvature distributions are genome specific, and pronounced differences are found between protein-coding and regulatory regions, respectively, that is, sites of extreme curvature and/or bendability are less frequent in protein-coding regions. A WWW server is set up for the prediction of curvature and generation of 3D models from DNA sequences (http:@www.icgeb.trieste.it/dna). PMID- 10710712 TI - Modelling DNA stretching for physics and biology. AB - We have used internal coordinate molecular mechanics calculations to study how the DNA double helix deforms upon stretching. Results obtained for polymeric DNA under helical symmetry constraints suggest that two distinct forms, an unwound ribbon and a narrow fibre, can be formed as a function of which ends of the duplex are pulled. Similar results are also obtained with DNA oligomers. These experiments lead to force curves which exhibit a plateau as the conformational transition occurs. This behaviour is confirmed by applying an increasing force to DNA and observing a sudden length increase at a critical force value. It is finally shown some DNA binding proteins can also stretch DNA locally, to conformations related to those created by nanomanipulation. PMID- 10710714 TI - Fast combinatorial cartography by FISH on combed genomic DNA. AB - A novel combinatorial method combining FISH on combed genomic DNA is presented for a fast high-resolution (1 Kbps) gene cartography. PMID- 10710713 TI - Single-chain 434 repressors with altered DNA-binding specificities. Isolation of mutant single-chain repressors by phenotypic screening of combinatorial mutant libraries. AB - Combinatorial mutant libraries of the single-chain 434 repressor were used to discover novel DNA-binding specificities. Members of the library contain one wild type domain and one mutant domain which are connected by a recombinant peptide linker. The mutant domain contains randomized amino acids in place of the DNA contacting residues. The single-chain derivatives are expected to recognize artificial operators containing the DNA sequence of ACAA--6 base-pairs--NNNN, where ACAA is bound by the wild-type and NNNN by the mutant domain. An in vivo library screening method was used to isolate mutant DNA-binding domains which recognize the TTAA site of an asymmetric operator. Several mutants showed high affinity binding to the selection target and also strong (up to 80 fold) preference for TTAA over the wild type TTGT sequence. Some of the isolated mutants bound with very high affinities (10-50 pM) to operators containing the TTAC sequence, a close homologue of the TTAA selection target. PMID- 10710715 TI - RNA folding and catalysis. AB - Catalysis in RNA is intimately connected to the folding. The small nucleolytic ribozymes function by a nucleophilic attack of the 2'-oxygen on the 3'-phosphate, in an SN2 mechanism. This requires an alignment of the 2'-O, 3'-P and 5'-O, that does not occur in normal A-form RNA. It is therefore likely that structural distortion plays a major role in the enhancement of the reaction rate, facilitating the trajectory into the in-line transition state. Given the polyelectrolyte nature of nucleic acids, metal ions are critical to folding processes in RNA. We have shown that two small nucleolytic ribozymes, the hammerhead and hairpin ribozymes, undergo metal ion-induced folding processes. The hammerhead ribozyme folds in two stages, each of which is induced by the binding of a single structural ion. The first corresponds to the formation of the ribozyme scaffold, while the second is the formation of the catalytic core of the ribozyme. By contrast, the hairpin ribozyme undergoes a single folding event induced by the binding of at least two metal ions, and involves the close interaction between two internal loops to form the active ribozyme. PMID- 10710716 TI - Histone-DNA contacts in structure/function relationships of nucleosomes as revealed by crosslinking. AB - We describe studies of histone-DNA contacts in the nucleosome using the method of covalent zero length protein-DNA crosslinking. These studies show that in intact nuclei isolated from different sources the linear sequential arrangement of histone-DNA contacts in the nucleosomal core is essentially the same. However, the relative strength of certain contacts varies and correlates with the level of chromatin activity and condensation. These altered contacts are located in the sharply bent regions of the nucleosomal DNA and are supposed to be sensitive to the structural changes that may occur during nucleosome functions. Studies of the mechanism of these alterations revealed that the difference in strength of these contacts is attributed to the different conformational state of the nucleosomal core and is caused by stretching of the nucleosomal DNA upon chromatin decondensation during its activation. Histone-terminal domains may be involved in this process through posttranslational modifications affecting chromatin condensation. The described localization of the histone H2A C-terminal domain in the nucleosome by crosslinking demonstrates the ability of this methodology to determine the location of histone-terminal domains and thereby elucidate their role in nucleosome function. Results of the described experiments suggest that chromatin decondensation may alter the nucleosomal DNA conformation and affect the histone-DNA contacts resulting in a structural transition that may play a role in rendering the nucleosome competent for transcription and/or replication. PMID- 10710717 TI - The role of histone H1 in chromatin condensation and transcriptional repression. AB - Linker histones are a major determinant of chromatin condensation. We discuss here the nature and position of the interaction of the globular domain of histone H5 with the core nucleosome and the relevance of this positioning to chromatin structure and the regulation of transcription of the Xenopus borealis 5S rRNA genes. PMID- 10710718 TI - Chromatin control of HIV-1 gene expression. AB - Upon infection of susceptible cells, the RNA genome of the human immunodeficiency virus type 1 (HIV-1) is reverse transcribed into double-stranded DNA, which can be subsequently integrated into the cellular genome. After integration, the viral long terminal repeat (LTR) promoter is present in a nucleosome-bound conformation and is transcriptionally silent in the absence of stimulation. Activation of HIV 1 gene expression is concomitant with an acetylation-dependent rearrangement of the nucleosome positioned at the viral transcription start site. Thus, similar to most cellular genes, the transcriptional state of the integrated HIV-1 provirus is closely linked to histone acetylation. This enzymatic activity results from the function of histone-specific nuclear acetyltransferase (HAT) enzymes. Efficient viral transcription is strongly dependent on the virally-encoded Tat protein. the mechanism by which Tat increases the rate of transcriptional initiation has been recently demonstrated and involves the interaction of Tat with the transcriptional coactivator p300 and the closely related CREB-binding protein (CBP), having histone acetyltransferase activity. PMID- 10710719 TI - Mechanisms of separation of the complementary strands of DNA during replication. AB - This article is a perspective on the separation of the complementary strands of DNA during replication. Given the challenges of DNA strand separation and its vital importance, it is not surprising that cells have developed many strategies for promoting unlinking. We summarize seven different factors that contribute to strand separation and chromosome segregation. These are: (1) supercoiling promotes unlinking by condensation of DNA; (2) unlinking takes place throughout a replicating domain by the complementary action of topoisomerases on precatenanes and supercoils; (3) topological domains isolate the events near the replication fork and permit the supercoiling-dependent condensation of partially replicated DNA; (4) type-II topoisomerases use ATP to actively unlink DNA past the equilibrium position; (5) the effective DNA concentration in vivo is less than the global DNA concentration; (6) mechanical forces help unlink chromosomes; and (7) site-specific recombination promotes unlinking at the termination of replication by resolving circular dimeric chromosomes. PMID- 10710720 TI - Exploring structure space. A protein structure initiative. AB - The genome projects are changing biology by providing the genetic blueprints of entire organisms. The blueprints are tantalizing but we cannot deduce everything we need to know from them, including the structures and detailed functions of proteins. In this paper we describe an approach for obtaining structural information about proteins on a genomic scale. We describe how structural and functional information might eventually be put together to form a basis for describing life at many levels. We then describe how structural information fits into this picture and classes of proteins for which structural information would be useful in a genomic context. We conclude with a proposal for an initiative to determine protein structures on a very large scale. PMID- 10710721 TI - The protein data bank. Bridging the gap between the sequence and 3D structure world. AB - The protein data bank (PDB), at Brookhaven National Laboratory, is a database containing information on experimentally determined three-dimensional structures of proteins, nucleic acids, and other biological macromolecules, with approximately 9000 entries. The PDB has a 27-year history of service to a global community of researchers, educators, and students in a wide variety of scientific disciplines. Data are easily submitted via PDB's WWW-based tool AutoDep, in either PDB or mmCIF format, and are most conveniently examined via PDB's WWW based tool 3DB Browser. Collaborative centers have been, and continue to be, established worldwide to assist in data deposition, archiving, and distribution. PMID- 10710722 TI - Functional genomics and enzyme evolution. Homologous and analogous enzymes encoded in microbial genomes. AB - Computational analysis of complete genomes, followed by experimental testing of emerging hypotheses--the area of research often referred to as 'functional genomics'--aims at deciphering the wealth of information contained in genome sequences and at using it to improve our understanding of the mechanisms of cell function. This review centers on the recent progress in the genome analysis with special emphasis on the new insights in enzyme evolution. Standard methods of predicting functions for new proteins are listed and the common errors in their application are discussed. A new method of improving the functional predictions is introduced, based on a phylogenetic approach to functional prediction, as implemented in the recently constructed Clusters of Orthologous Groups (COG) database (available at http:@www.ncbi.nlm.nih.gov/COG). This approach provides a convenient way to characterize the protein families (and metabolic pathways) that are present or absent in any given organism. Comparative analysis of microbial genomes based on this approach shows that metabolic diversity generally correlates with the genome size-parasitic bacteria code for fewer enzymes and lesser number of metabolic pathways than their free-living relatives. Comparison of different genomes reveals another evolutionary trend, the non-orthologous gene displacement of some enzymes by unrelated proteins with the same cellular function. An examination of the phylogenetic distribution of such cases provides new clues to the problems of biochemical evolution, including evolution of glycolysis and the TCA cycle. PMID- 10710723 TI - The beamlines of ELETTRA and their application to structural biology. AB - Protein crystallographers are nowadays regular users of synchrotron radiation (SR) facilities for several applications. The goal of majority of users is simply to extract more accurate, higher resolution data from existing crystals; they use monochromatic radiation and the rotation method, in order to get a complete survey of the reciprocal space in a short time. In fact the brilliance of SR is essential, due to the weak scattering power of the samples, and because of their sensibility to radiation damage. Over the last few years, however, a general increase of interest for measurements at multiple wavelengths, which exploit the anomalous dispersion for the phase problem (multiwavelength anomalous diffraction -MAD), has generated the need of intense tuneable sources. For these applications, the emphasis is on accurate measurements of the small differences between the intensities of Bragg reflections at various energies across the absorption edge of an element present in the sample. The macromolecular diffraction beamline at ELETTRA, which is now running routinely since spring 1995, has been designed to provide a high flux--highly collimated tuneable X-rays source in the spectral range between 4 and 25 keV. The radiation source is the 57 pole wiggler, which delivers a very intense radiation up to 25 keV, and is shared and used simultaneously with the small angle X-ray scattering (SAXS) beamline. The front-end filter system has a cut-off energy at about 4 keV. The beamline optics consists of a pseudo-channel-cut double-crystal monochromator followed by a double focusing toroidal mirror. The tunability and the stability of the monochromator allows the user to perform MAD experiments, and for this purpose, a fluorescence probe for the exact calibration of the absorption edge is available on-line. The experimental station is based on an imaging plate area detector from MarResearch, with a sensible area of 345 mm in diameter. A cooled N2-stream is available to cool the sample crystal in order to reduce the radiation damage. SAXS is an experimental technique used to derive structural information about supra-molecular assemblies, amorphous materials and partly ordered systems (e.g. size and shape of large molecules). The high-flux SAXS beamline at ELETTRA is mainly intended for time-resolved studies on fast structural transitions in the sub-millisecond time region in solutions and in partly ordered systems, triggered by external or process parameters, with a SAXS resolution between 10 and 1400 A in real space. The source is the already mentioned 57-pole and the SAXS beamline accepts three discrete energies of its spectrum, namely 5.4, 8 and 16 keV. The beamline optics consists of a flat double-crystal monochromator and a double focusing toroidal mirror. A multi-purpose sample stage, movable along an optical table in order to optimise the sample to detect distance, allows to perform fast time-resolved relaxation studies based on temperature- or pressure-jumps as well as stopped flow experiments. Moreover, the users have option to install their own specialised sample surrounding equipment. The optimisation of the beamline with respect to high-flux and consequently high-flux density, allows to perform the following experiments: low contrast solution scattering, grazing incidence surface diffraction, micro-spot scanning, X-ray fluorescence analysis, time resolved studies > or = 11 microseconds, simultaneous small- and wide-angle measurements on gels, liquid crystals, biopolymers, amorphous materials, muscles. PMID- 10710724 TI - The paradox of physicians and administrators in health care organizations. AB - Rapidly changing times in health care challenge both physicians and health care administrators to manage the paradox of providing orderly, high quality, and efficient care while bringing forth innovations to address present unmet problems and surprises that emerge. Health care has grown throughout the past several centuries through differentiation and integration, becoming a highly complex biological system with the hospital as the central attractive force--or "strange attractor"--during this century. The theoretical model of complex adaptive systems promises more effective strategic direction in addressing these chaotic times where the new strange attractor moves beyond the hospital. PMID- 10710725 TI - Commentary: organizational directions for the millennium: what needs to be done! PMID- 10710726 TI - Commentary: the potential of chaos theory and complexity theory for health services management. PMID- 10710727 TI - Response to the commentaries of Kaluzny, Arndt, and Bigelow: personal reflections on health care organization as viewed through the lens of complexity theory. PMID- 10710728 TI - The paradox prescription: leading the medical group of the future. AB - Leaders of health care organizations must constantly deal with paradox in and around their organizations. The paradox profile is a modeling technique for examining how organizations prioritize their dealings with the eight competing issues from which the four paradoxes are derived. This article demonstrates the leadership challenges that exist when a paradox is present and how these tensions differ when the integrative status of the medical group--as either part of an integrated system or not--is considered. PMID- 10710729 TI - Integrated health care systems governance: prevention of illness and care for the sick and injured. AB - The public health aspects of promotion of health and prevention of illness take on a different vision when one moves from biological concerns to social and societal dyscrasias. Social illness is as much the concern of a hospital system board and its medical staff as the biomedical dyscrasias which are the traditional concerns of public health agencies. An organizational entity under a board of trustee's auspices must become involved because all social illnesses end up requiring physician and hospital care. PMID- 10710730 TI - The more things change, the more they stay the same. AB - For decades, the hospital environment has been described as turbulent and hostile. At the same time, the transfer of business practices into hospitals has been advocated, accompanied by the largely untested assumption that these practices are crucial to performance and even survival. As this pattern became entrenched, the accumulated knowledge gained within the industry of managing in a hostile and turbulent environment has been overlooked. We argue that it is time to question the pattern. PMID- 10710731 TI - Building client centered systems of care: choosing a process direction for the next century. AB - Forecasting the future of health care is difficult. However, we argue that this future will include the movement of health care through process improvement (enhancement) toward the objective of mass customization. This article discusses how mass customization might apply to specific portions of client-centered health care. PMID- 10710732 TI - Managing health care organizations: where professionalism meets complexity science. AB - This article examines the intersection of professionalism and complexity science as a source of new insights for improving the health care industry from both a clinical and business point of view. Viewing health care organizations as professional complex adaptive systems suggests eight leadership tasks for addressing the circumstances that engulf health care. Managers who adopt this view will be able to create new levers for positive movement in their organizations. PMID- 10710733 TI - Beyond managed costs. AB - Managed care organizations (MCOs) face an uncertain future. While consolidation and price competition have expanded their market share, health care expenditures are expected to rise in the near future, and the cost containment premise--and promise--of MCOs is being threatened by mixed blessing and nonsupportive stakeholders. To shed light on MCOs' situation, we discuss four drivers for change in health management in the U.S.: technology, regulation, consumerism, and demographics. Using those four drivers, we then assess the various stakeholders in the industry through a competitive analysis and a stakeholder analysis. These analyses suggest that the munificence of the MCO business environment has significantly declined, especially among supplier and buyer stakeholders. Hence, MCOs cannot continue to manage health care costs alone as this will no longer generate sufficient support among buyer and supplier stakeholders. Instead, MCOs must tackle five critical health care issues by working closely with other stakeholders and also by learning what they can from innovative health care initiatives both inside and outside the United States. PMID- 10710734 TI - Physicians and decisions: a simple rule for increasing connections in hospitals. AB - When hospitals are viewed as complex adaptive systems, simple rules can lead to behavior that emerges as complex and that enables creative, adaptive organizational responses. Based on empirical findings from a decade of research on hospital strategic decision making this article offers the simple rule of letting physicians help decide strategic issues. Seven managerial guidelines for implementing this rule are presented. PMID- 10710735 TI - Shoulder dystocia. Rotational maneuvers revisited. AB - Shoulder dystocia is an acute obstetric emergency that necessitates prompt, skillful intervention in order to prevent serious fetal trauma or death. Of the maneuvers described to deal with this difficult problem, rotational maneuvers are among the most ingenious. In spite of the effectiveness of these techniques, various technical deviations have led to the incorrect description and implementation of these maneuvers. This review of the rotational maneuvers used to counter shoulder dystocia gives particular attention to the techniques described originally. PMID- 10710736 TI - Pregnancy rates after laparoscopic treatment. Differences related to tubal status and presence of endometriosis. AB - OBJECTIVE: To examine how preexisting tubal adhesions and endometriosis affect pregnancy outcome after laparoscopic treatment in infertile women with no apparent causes of infertility other than tubal factors. STUDY DESIGN: Pregnancy outcomes in 186 infertile women for a follow-up period of 18 months after laparoscopy were analyzed. Laparoscopic manipulations consisted of adhesiolysis of tubes and removal of endometriotic lesions. RESULTS: The patients were classified into three groups, those with no tubal adhesions (group A, n = 83), unilateral tubal adhesions (group B, n = 46) and bilateral tubal adhesions with at least one tube patent (group C, n = 57). The cumulative pregnancy rate in group C (13.2%) was lower than in groups A (41.8%) and B (45.7%) 18 months after laparoscopy. The average time to conception in group A (6.7 +/- 0.8 months) tended to be shorter than that in group B (10.6 +/- 1.2 months). In group A, pregnancy rates were essentially the same between minimal/mild endometriosis and moderate/severe endometriosis. Regarding group B, women with minimal/mild endometriosis exhibited significantly higher pregnancy rates than those with moderate/severe endometriosis, while pregnancy rates in women without endometriosis fell in between. CONCLUSION: Pregnancy rates after laparoscopic treatment are different in relation to tubal status and the presence of endometriosis. PMID- 10710737 TI - CSF/serum beta-hCG ratio in patients with brain metastases of gestational trophoblastic tumor. AB - OBJECTIVE: To assess the accuracy of cerebrospinal fluid (CSF)/serum ratio of beta-subunit of human chorionic gonadotropin (beta-hCG) in detecting brain metastasis of gestational trophoblastic tumor (GTT). STUDY DESIGN: The subjects were ten patients with GTT and brain metastases. Spinal puncture and veni puncture were performed for measurement of beta-hCG titer in CSF and serum to determine the CSF/serum ratio before starting multiagent chemotherapy and/or brain irradiation. RESULTS: Five patients manifested a CSF/serum beta-hCG ratio > 1/60, and five manifested a ratio < 1/60. CONCLUSION: The CSF/serum beta-hCG ratio is not accurate enough to be routinely considered in the workup, management, and/or surveillance of GTT with brain metastases. Hence, it is perhaps not necessary to perform spinal puncture if the only purpose is to determine the CSF/serum ratio. PMID- 10710738 TI - Cultural elements of postpartum depression. A study of 327 Jewish Jerusalem women. AB - OBJECTIVE: To examine the social, cultural and religious factors underlying postpartum depression within a cultural cross-section of Jewish Jerusalem women. STUDY DESIGN: A prospective, repeated-measures study of 327 women. The Edinburgh Postpartum Depression Scale (EPDS) was administered immediately postpartum and 6 10 weeks later. Detailed sociodemographic information included perceptions of the pregnancy, community supports and religious affiliation. Odds ratios, 95% confidence interval and P values were calculated for all covariates. Multiple logistic regression was performed to estimate the degree of independent association between religiosity and postpartum depression. RESULTS: Postpartum depressive symptoms significantly associated with secular affiliation (odds ratio [OR] 2.9 [1.3-6.3] and tended toward an inverse association with orthodox affiliation (OR 0.6 [0.3-1.3]). Across secular, traditional, religious and orthodox groups, there was a decreasing trend in EPDS mean scores. Other predictors of depressive symptoms were psychiatric history, immigrant status and poor support with newborn care. CONCLUSION: Our study sample was particularly suitable for the assessment of cultural and religious elements of postpartum depression. We found religiosity, with its associated social and community structuring and well-defined social roles, to be significantly associated with self-reported postpartum depressive symptoms. These findings suggest that cultural factors, including role definitions, community support and rituals, may explain discrepancies found in the incidence of postpartum depression. PMID- 10710739 TI - Fetal heart rate baselines in twins. Interobserver agreement in antepartum estimation. AB - OBJECTIVE: To assess interobserver agreement in antepartum estimation of fetal heart rate (FHR) baselines in twins. STUDY DESIGN: Two residents and one specialist in obstetrics and gynecology, all with special interest in FHR monitoring, independently estimated baselines in 162 consecutive antepartum FHR tracings recorded in 24 twins. Tracings were obtained with a dual-channel fetal monitor for the simultaneous recording of both twins' heart rates. Baselines were estimated, as single values corresponding to the mean of the lowest stable FHR segment, in the absence of fetal movements and uterine contractions, within physiologic limits (110-150 beats per minute [bpm]). If these criteria were not met, the possibility of persistent bradycardia or tachycardia was considered, and if this was confirmed in a tracing with at least 40 minutes, a baseline < 110 or > 150 bpm was chosen. Interobserver agreement was assessed by the proportions of agreement (PA), kappa statistic (K) and intraclass correlation coefficient (ICCC), with 95% confidence intervals (CIs). RESULTS: Interobserver agreement was excellent, with a PA of 0.90 (95% CI: 0.89-0.91), K of 0.88 (95% CI: 0.84-0.92) and ICCC of 0.91 (95% CI: 0.88-0.94). CONCLUSION: Interobserver agreement in antepartum estimation of fetal heart rate baselines in twins was excellent with the baseline concept used in this study. PMID- 10710740 TI - Endometrial biopsy using the Tao Brush method. A study of 50 women in a general gynecologic practice. AB - OBJECTIVE: To test whether the Tao Brush can retrieve sufficient endometrium for diagnosis and also to observe patients' tolerance of its use. STUDY DESIGN: Fifty women underwent Tao Brush sampling: 25 were sampled by Tao Brush alone, and the remainder were sampled by Pipelle immediately following. Patients' reactions to each method were observed. RESULTS: Pipelle currettes larger pieces of endometrium; the Tao Brush obtains smaller pieces. There was no discrepancy between Tao Brush and Pipelle except that Pipelle sampled two of five endometrial polyps, while the Tao Brush sampled none. There was less tissue insufficient for diagnosis with the Tao Brush (2%) than Pipelle (12%). Most patients did not show signs of distress during Tao Brush sampling but grimaced during Pipelle suction curettage. For each of the 16 patients in the second group, the Tao Brush was significantly less painful than Pipelle (P < .01). CONCLUSION: Our data suggest that the Tao Brush is an effective alternative endometrial sampler, causes less pain and produces less tissue insufficient for diagnosis than does the Pipelle. The CPT billing codes (58100) are the same for both the Pipelle and Tao Brush. PMID- 10710741 TI - Influence of polyvinylpyrrolidone on the outcome of intracytoplasmic sperm injection. AB - OBJECTIVE: To assess the effectiveness of a procedure for intracytoplasmic sperm injection (ICSI) modified so as not to use polyvinylpyrrolidone (PVP) and to examine clinical outcome. STUDY DESIGN: Seventy-seven cycles of ICSI were performed over a one-year period. PVP was used for sperm immobilization in 39 of these cycles and was eliminated from the other 38 cycles. Difference in fertilization rate, cleavage rate, parthenogenetic activity, clinical pregnancy rate, ongoing pregnancy rate and grading of preembryos between the two groups was compared. RESULTS: The non-PVP group had a higher fertilization rate (57.63% vs. 84.43%, P < .001) and better preembryo quality (chi 2 = 6.80, P = .009) than the PVP group. There was no significant difference in cleavage rate, parthenogenetic activity, clinical pregnancy rate and ongoing pregnancy rate between the two groups. CONCLUSION: Performing ICSI without PVP may improve the fertilization rate and preembryo grading. However, further study with a larger cohort is necessary to determine whether the modified procedure can increase the pregnancy rate. PMID- 10710742 TI - Serum leptin concentrations in women during gonadotropin stimulation cycles. AB - OBJECTIVE: To determine whether elevated follicular steroid levels during gonadotropin stimulation cycles are associated with altered circulating leptin concentrations. STUDY DESIGN: Sequential serum samples were collected from women (N = 37) undergoing luteal phase GnRH agonist + FSH treatment cycles prior to oocyte retrieval for in vitro fertilization. Leptin concentrations in serum were measured by radioimmunoassay and compared with serum estradiol, testosterone and dehydroepiandrosterone sulfate (DHEAS) levels. RESULTS: Serum leptin concentrations during stimulated cycles were variable between patients and correlated positively (r = .556, P < .01) with body mass index. Serum leptin levels correlated positively (r = .185, P < .05) with estradiol concentrations across all days. Mean serum leptin concentrations on the day gonadotropin treatment began (baseline, 12.9 +/- 2.0 ng/mL) were lower (P < .0001) than on the day peak estradiol levels were reached (18.4 +/- 2.3 ng/mL). Serum leptin concentrations also correlated with DHEAS levels (r = .214, P < .05) but did not correlate with testosterone or the estradiol:testosterone ratio. CONCLUSION: Gonadotropin stimulation in women is associated with elevated leptin levels, consistent with an interaction between the reproductive axis and leptin secretion and/or clearance. PMID- 10710743 TI - Role of GnRH agonists in managing proximal fallopian tube obstruction. AB - OBJECTIVE: To assess the response of proximal fallopian tube obstruction to a medically induced hypoestrogenic state after control for spasm at the uterotubal ostium. STUDY DESIGN: This was a prospective, randomized, placebo-controlled, pilot study in a tertiary care, university-affiliated infertility practice. Twenty-one infertile women with unilateral or bilateral proximal tubal obstruction previously diagnosed by standard hysterosalpingography or laparoscopic chromotubation were randomized into two groups in a 2:1 design. Group I, 14 patients (27 occluded tubes), was administered a depot preparation of the gonadotropin-releasing hormone (GnRH) agonist leuprolide acetate, 3.75 mg intramuscularly every 28 days. Group II, seven patients (11 occluded tubes), was administered a placebo. Follow-up hysterosalpingography was performed after administration of the antispasmodic glucagon within four weeks of completion of the protocol. RESULTS: Evidence of ovarian suppression was confirmed within four weeks in group I. A trend toward higher posttherapy patency rates was noted in group I (74.7% vs. 40%). The lack of statistical significance may have been a function of sample size. Similarly, spontaneous intrauterine pregnancy rates were also higher in GnRH agonist-treated patients (35.1% vs. 16.6%, P < .05). Tubal patency rates were significantly greater in patients with documented estrogen sensitive disorders who received the agonist (75% vs. 20%, P < .05) CONCLUSION: Proximal tubal obstruction may be an estrogen-sensitive phenomenon in a subset of infertile patients. The administration of GnRH agonists may successfully overcome this state and result in enhanced conception rates. PMID- 10710744 TI - Trichomonas vaginalis and bacterial vaginosis. Coexistence in vaginal wet mount preparations from pregnant women. AB - OBJECTIVE: To identify how frequently trichomoniasis and characteristics of bacterial vaginosis (BV) occur concomitantly in wet mount preparations from pregnant women. STUDY DESIGN: Diagnosis of trichomoniasis was predicted on visualization of the organism. Diagnosis of BV required a positive volatile (whiff) test, presence of "clue cells" and one of two minor criteria: (1) absence of lactobacilli, or (2) a pH > 4.5. Pregnant women from January 1995 to July 1997 at our clinic had wet mount/KOH preparations performed as standard prenatal care. Corresponding medical charts were analyzed for symptoms, race, BV, sexually transmitted diseases, urinary tract infections and other infections. RESULTS: Of 191 pregnant women identified, 69 had trichomoniasis. Seventy-nine percent of the 69 were African American. Fifteen percent of pregnant women (17) had concomitant trichomoniasis and BV. Irrespective of race, 35-38% of pregnant women with trichomoniasis had another sexually transmitted disease or a urinary tract infection diagnosed in that pregnancy. CONCLUSION: BV, or bacteria excess syndrome, is a frequent coinfection in pregnant women harboring Trichomonas vaginalis. PMID- 10710745 TI - Comparing two embryo transfer catheters. Use of a trial transfer to determine the catheter applied. AB - OBJECTIVE: To analyze the performance of two different embryo transfer catheters (Wallace and Frydman) in an in vitro fertilization (IVF)-intracytoplasmic sperm injection (ICSI) program. STUDY DESIGN: Four hundred twenty-eight IVF or ICSI embryo transfer cycles were analyzed. A trial transfer was performed before the initiation of controlled ovarian hyperstimulation to determine the choice of embryo transfer catheter, Wallace or Frydman. Actual transfer was undertaken with the catheter chosen from the trial transfer. RESULTS: During actual embryo transfer, 214 (93.5%) of the intended 229 Wallace transfers were successful, and in 15 transfers the Frydman catheter was used. Of the intended 199 Frydman transfers, all were successful. Clinical pregnancy rate, implantation rate per embryo and ectopic pregnancy rate per transfer for the Wallace catheter were 41.6%, 16% and 0.9%, respectively. Respective rates for the Frydman catheter were 36.0%, 14.4% and 0.9% (P > .05 for all variables). Trial catheterization prevented most of the unanticipated procedural difficulties during the actual transfer. CONCLUSION: Both Wallace and Frydman catheters performed similarly, although there was a slight but nonsignificant increase in clinical pregnancy rates with the Wallace catheter. PMID- 10710746 TI - Spontaneous uterine rupture in the early third trimester after laparoscopically assisted myomectomy. A case report. AB - BACKGROUND: The development of new and innovative laparoscopic instruments has allowed a greater number of gynecologic surgeons to laparoscopically remove large, intramural leiomyomata. Cases of both successful pregnancy and uterine rupture following laparoscopic myomectomy have been reported. This is the first report of uterine rupture in pregnancy following a laparoscopically assisted myomectomy. CASE: A 26-year-old, nulligravid woman underwent a laparoscopically assisted myomectomy. While the myomectomy had been performed laparoscopically, the uterine incision had been repaired in layers through a minilaparotomy incision. Two years later she became pregnant and, at 29 weeks' gestation, presented to labor and delivery with contractions and uterine tenderness. Over the next several hours, a nonreassuring fetal heart rate developed, and a cesarean section was performed, revealing hemoperitoneum and uterine rupture at the site of the prior myomectomy. CONCLUSION: The ultimate integrity of a uterine incision may depend not only on how the incision is repaired but also on how it is made. Laparoscopically created uterine incisions may not be as strong as those made at laparotomy, regardless of the method of closure. PMID- 10710747 TI - Low-dose methotrexate treatment for interstitial pregnancy. A case report. AB - BACKGROUND: Only a small number of case reports have described medical treatment of interstitial ectopic pregnancies. Almost all of the reported patients were treated with repeated high doses (1 mg/kg) of methotrexate. CASE: At 6 weeks of gestation, a 31-year-old woman, gravida 5, para 4, was diagnosed with a 0.96 x 1.36-cm right cornual pregnancy. As the patient desired future fertility, she received 100 mg of intramuscular methotrexate (50 mg/m2). She was then followed on an outpatient basis, with serum human chorionic gonadotropin values appropriately declining. Serial ultrasound also showed decreasing size of the gestational sac. Twenty-one days after the methotrexate dose, the patient experienced rupture of the right posterior cornu, necessitating exploratory laparotomy. CONCLUSION: Extreme caution should be used when treating interstitial gestations with single-dose methotrexate. All patients should be extensively counseled regarding the significantly increased risk of failure, possibility of rupture and need for emergency surgery. PMID- 10710748 TI - Unilateral chronic tuboovarian abscess secondary to ruptured colonic diverticulum presenting as a brain abscess. A case report. AB - BACKGROUND: Tuboovarian abscesses (TOAs) are a somewhat unusual finding in postmenopausal patients without risk factors. We present a rare case of unilateral TOA initially presenting as a brain abscess in a postmenopausal woman. CASE: A 61-year-old woman presented with a complaint of forgetfulness, nausea and vomiting, with lower abdominal pain and diarrhea. She was found to have a brain abscess, which was treated by craniotomy, with drainage of the abscess, and intravenous antibiotics. The patient was subsequently found to have a pelvic mass, which, on laparotomy, was a unilateral TOA. Pathology demonstrated that the abscess contained vegetable matter consistent with origin in a ruptured diverticulum. CONCLUSION: Diagnosis of a brain abscess should prompt a thorough investigation for a primary infectious source, including the gastrointestinal and genitourinary tracts. PMID- 10710750 TI - Hysterotomy for retained placenta in a term angular pregnancy. A case report. AB - BACKGROUND: Angular placentation may be a cause of retained placenta and may require hysterotomy. CASE: A 33-year-old woman with a prior cesarean section underwent an uncomplicated vaginal delivery, had a retained placenta with postpartum hemorrhage and required hysterotomy because the placenta was inaccessible due to its angular location. CONCLUSION: Antepartum ultrasound diagnosis may be possible in some cases of angular placentation. PMID- 10710749 TI - Twin pregnancy in a woman on long-term epoprostenol therapy for primary pulmonary hypertension. A case report. AB - BACKGROUND: Pregnancy associated with primary pulmonary hypertension is an uncommon observation, with maternal mortality > 50%. Experience treating this condition is limited. Past reports have emphasized the need for pregnancy termination. In the last few years there has been considerable interest in long term intravenous use of epoprostenol (prostacyclin) in patients with primary pulmonary hypertension. CASE: A woman with severe primary pulmonary hypertension who was on long-term epoprostenol therapy became pregnant with twins and was treated with high doses of epoprostenol and nitric oxide during delivery and the postpartum period. She was well six months later on continuous epoprostenol therapy. The one viable infant was alive and still hospitalized at this writing. CONCLUSION: Epoprostenol therapy may be continued during pregnancy in patients with severe primary pulmonary hypertension for long-term pulmonary vasodilatation. PMID- 10710751 TI - Midcycle administration of single-dose GnRHa for luteal phase failure in women with ovarian hyperstimulation. A report of five cases. AB - BACKGROUND: Exogenous administration of gonadotropin-releasing hormone agonist (GnRHa) induces an endogenous midcycle gonadotropin surge. However, its use to induce ovulation and maintain luteal function in non-in vitro fertilization patients who receive ovarian stimulation is unknown. CASES: Five infertile women who underwent controlled ovarian hyperstimulation with human menotropin developed multiple ovarian follicles. In an attempt to circumvent the potential ovarian hyperstimulation syndrome, 1 mg of leuprolide acetate was administered subcutaneously to three patients in an attempt to induce the endogenous luteinizing hormone surge. All three patients began menstruation six to seven days after GnRHa administration with serum progesterone levels between 0.2 and 0.5 ng/mL. Similar ovarian stimulation cycles with ovulation induced by human chorionic gonadotropin in these individuals revealed a normal luteal phase length and midluteal progesterone levels. When double doses of leuprolide acetate were used on two patients, normal luteal length and midluteal serum progesterone levels occurred. CONCLUSION: A single bolus of GnRHa during the late follicular phase may be inadequate to initiate normal luteal function in cycles with ovarian hyperstimulation. PMID- 10710752 TI - Early form of ovarian cancer originating in inclusion cysts. A case report. AB - BACKGROUND: Most surface epithelial-stromal tumors of the ovary are thought to arise from epithelial inclusion cysts; thus, those cysts and the original surface epithelium are precursor lesions of ovarian carcinoma. CASE: A 46-year-old woman underwent a hysterectomy, right salpingo-oophorectomy and wedge biopsy of the left ovary because of a uterine myoma and right dermoid cyst. Histologic examination incidentally showed a tiny focus of well-differentiated serous carcinoma originating in inclusion cysts in the biopsied left ovary. Relaparotomy was performed, and only metaplastic epithelium in the inclusion cysts was found in the remnant of the left ovary. This finding was thought to be the origin of an ovarian cancer. CONCLUSION: Carcinoma precursors should occur in epithelial inclusion cysts of the ovary, as in the cervix and endometrium, but have been reported only rarely. Their incidental finding may help identify patients at risk of developing ovarian carcinoma. PMID- 10710753 TI - Factors to consider when designing phase III clinical trials involving economic evaluations. AB - Given the recognized need to examine cost-effectiveness data in addition to clinical data when making decisions relating to choice of clinical interventions, there is a growing interest and experience in undertaking economic evaluations alongside clinical trials. It has been argued, when an economic evaluation is necessary for assessing the cost-effectiveness of a medical intervention, the integration of both the medical and clinical issues need to be included at the start of designing a clinical trial. A proposal has been made for how clinical researchers and health economists may cooperate successfully at the successive phases in designing a clinical trial. Therefore, discussion points for clinical researchers and health economists are given, and the possible methodological consequences of adding an economic evaluation to a trial are addressed. PMID- 10710754 TI - Re-examining the temporal locus of knowledge of results (KR): a self-paced approach to learning. AB - Using a self-paced procedure, the effects of unconfounded temporal locus of KR in the acquisition of a simple linear-positioning task was examined. Changes in the chronological profile of KR delivery were evaluated when participants manipulated the time course of the experiment at their own discretion. 29 participants (18 to 32 years) practiced finding an 8-in. line with no fixed starting and ending points. One-way repeated-measures analyses of variance, simple regression analyses across blocks of practice (30), and Pearson product-moment correlations between the KR-time intervals and the performance scores indicated that (a) under self-paced procedures both the KR-delay and post-KR interval decreased congruently with performance error scores, while the temporal component of the task (movement time) and the ratio between the KR-delay and the post-KR interval remained unchanged, (b) any effect on intertrial interval and interstimulus interval in motor skill acquisition should be examined in terms of the KR-delay and post-KR interval, and (c) the relationship between the performance scores and post-KR interval may be used to indicate the point at which KR is no longer required. PMID- 10710755 TI - The running horse stops: the hypothetical role of the eyes in imagery of movement. AB - To examine the hypothetical role of the eyes in visual mental imagery of movement 72 undergraduate women students in psychology were asked to imagine a running horse and then to produce the same mental image without moving the eyes and the head. In 59% of the subjects interesting modifications of the imagined movement appeared: 37% observed an inhibition of the movement and 19% an evident slowing up of the moving figure. The interpretation of this result was made by hypothesizing that the eyes are concretely involved in visual imagery processes. PMID- 10710756 TI - Changes in Shotokan karate black-belt Heian kata performance times: a longitudinal study. AB - 10 experienced black-belt subjects were individually timed on each of the five Heian kata and then timed again four years later. Performance time increased significantly in four of the kata, and this was interpreted as a positive progression. PMID- 10710757 TI - Speaking speed effects on delayed auditory feedback disruption of speech fluency. AB - 24 Italian medical students performed a task of verbal fluency. 12 students (the control group) receiving Normal Auditory Feedback and 12 students receiving Delayed Auditory Feedback (delay of 200 msec.) performed six trials in six different experimental settings: normal or increased speaking rate, and, for each condition, once with bilateral input of the auditory feedback, once to the right ear, and once to the left ear. At the normal speaking rate, the disruptive effect of delayed feedback was confirmed. As the speaking rate increased, the total number of errors increased within the control group but decreased within the group given delayed feedback, although the total number of errors was always greater for the latter. In addition, speech was more disrupted when the auditory input was returned to the right ear (left hemisphere) for all the different conditions: Normal and Delayed Auditory Feedback, normal and increased speaking rate. In particular, the left hemisphere was less resistant to the disruptive effect of the delayed feedback than the right hemisphere. From these results, we suggest that, when speaking more quickly, one uses more central mechanisms of movement programming (cortical-cerebellum-thalamus-cortical, cortical-corpus striatum-thalamus-cortical, and cortical-thalamus-cortical circuits), or attentional control (cortico-reticular-cortical circuits) than peripheral mechanisms (tactile, proprioceptive, and acoustic circuits). This may explain the decreased disruptive influence of delayed auditory feedback on speed, fluency, and quality at increased speaking rates. Hemispheric specialization processes, however, may explain the more pronounced susceptibility of the left hemisphere or the less pronounced susceptibility of the right hemisphere during the delayed feedback condition. In fact, the former processes phonemic, grammatical, and lexical features of words whilst the latter is competent in using metaphors and prosody in controlling the emotional aspects of language. Moreover, the right hemisphere is more active on attentional tasks. PMID- 10710758 TI - Time taken as a determinant of advancement in Shotokan karate black-belt Heian kata performance. AB - 30 experienced black-belt subjects were individually timed on each of the five Heian kata. Significant relationships were found between years of training and increased performance time on all the kata and for four with age controlled. The multiple correlation was significant. PMID- 10710759 TI - KR-withdrawal and self-paced motor performance. AB - The purpose of this study was to examine the effects of KR-withdrawal on performance and the chronological profile of KR-delivery following the self-paced procedure and the blank trial technique. 120 participants were randomly separated into 12 groups and practiced finding an 8-in. line with no fixed starting and ending points. Appropriate 12 x 3 (groups by blocks of trials) analyses of variance and a prior contrasts were conducted to analyze all dependent variables (absolute error, variable error, movement time, KR-delay, post-KR interval, and KR-delay by post-KR interval ratio). The results indicated that (a) accuracy decreased significantly when KR was withdrawn early in practice, while the effects of KR-withdrawal later in practice affected accuracy differently. (b) Early in practice, participants performed at the same level of consistency even when KR was withdrawn; however, later in practice, the participants were less variable. (c) Movement time, KR-delay, and post-KR-interval were affected differently during early and later stages of practice. (d) Moderate to high correlations between absolute error and post-KR intervals may indicate the development of mechanisms for error detection. PMID- 10710760 TI - Time taken as a determinant of advancement in Shotokan karate black-belt Tekki kata performance. AB - 18 experienced black-belt subjects were individually timed on the kata Tekki Shodan. A significant relationship was found between years training and increased performance time and also between increased performance time and grade, both independent of age. PMID- 10710761 TI - Self-monitoring, and individual expectation of performance-norms in sport teams. AB - The main purpose of this study was to examine the relationship between self monitoring and individuals' expectation of performance-norms, the attitudes shared among team members about how high a performance the group should achieve in team sports. A secondary purpose was to assess whether the relationship between self-monitoring and individual expectation of performance-norms would be moderated by the type of group selected. Analysis suggests that for an elite sport team there is no difference between the performance-norm for the team and individuals' expectations in terms of self-monitoring. For recreational sport teams, however, those high on self-monitoring had higher individual expectations of performance-norms than the low self-monitors. PMID- 10710762 TI - Comparisons of magnitude estimation scaling of rock music by children, young adults, and older people. AB - The present study concerned the perceptual processing of complex auditory stimuli in 10 children (M age = 8.1) as compared to 10 young adults (M age = 19.3) and 10 older adult subjects (M age = 54.2). The auditory stimulus used was 10 sec. of rock music (Led Zeppelin, 1969). All three groups provided numerical responses to nine intensities of the rock music stimulus (10, 20, 30, 40, 50, 60, 70, 80, 90 dB above threshold). Analysis showed that the children reported a wider range of numerical responses than both adult groups. The mean numerical responses for the children ranged from .54 to 54.24. For the young adults the range was .76 to 11.37, and for the older subjects it was 1.6 to 23.31. Results suggest that the children were not bound by the same set of rules as the adults with regard to magnitude estimation scaling of the loudness of the rock music stimulus. Their internal scaling mechanisms appeared to be more flexible and broader based than those of the adults who participated in this study. PMID- 10710763 TI - Dynamic capture of sound motion by light stimuli moving in three-dimensional space. AB - A moving light stimulus produced a sensation of motion for a stationary sound stimulus presented simultaneously, which was called the dynamic visual capture by Mateeff, Hohnsbein, and Noack in 1985. The present study examined whether the moving light stimulus might induce a perceptual shift in the velocity, that is, the speed and the direction, of a moving sound stimulus. This type of the visual capture was explored in the directions where the sound stimulus moved along the motion tracks of the light stimulus: first in a horizontal and a vertical orientation and secondly in a depth orientation. Moreover, perceptual distortion of the direction of movement of the sound stimulus was investigated when the direction from which a sound stimulus moved deviated from the motion tracks of a light stimulus. It was found that the dynamic visual capture was inducible in all of three orientations, appearing more strongly in the vertical and the depth orientations than in the horizontal orientation. The perceived direction of the moving sound stimulus was greatly influenced by the direction where the light stimulus moved simultaneously and was often the same as the physical direction of the light stimulus, even when the physical motion directions of those two stimuli were perpendicular or opposite each other. PMID- 10710764 TI - Conflicting activities for exercise. AB - 190 nonexercisers foresaw regular exercise (> or = 3 times a week) as interfering with day-to-day activities. Most frequently mentioned were various social activities, doing household chores, and watching TV. The impediment of all activities, except watching TV, was expected to be at least rather bothersome. Interventions should focus on these particular barriers to action. PMID- 10710765 TI - More practice does not necessarily enhance transfer of learning: evidence and interpretations. AB - The purpose of this study was to investigate what was learned and transferred of a criterion task using a transfer task and a self-paced procedure. Based on the results of two previous studies, five durations of practice were specifically selected to examine what can be transferred to a new task after different amounts of practice (40, 75, 100, 125, and 150 acquisition trials) on a criterion task. The criterion task required subjects to practice finding an 8-in. line with no fixed starting and ending points from left to right. The transfer task was different from the criterion task in two dimensions, distance (10-in. line) and direction (right to left). 50 volunteers were randomly assigned into 5 groups. Appropriate 5 x 2 (groups by blocks of practice) analyses of variance and a priori contrasts were conducted for all dependent variables (absolute error, variable error, movement time, KR-delay, post-KR interval, intertrial interval, interstimulus interval, and KR-delay by post-KR interval ratio) to examine the effects of terminal acquisition of the criterion task on the initial performance of the transfer task. Analysis indicated that, during practice, participants developed a functional interaction between performance characteristics and chronological profile of KR-delivery of the criterion task that was transferable to the transfer task. Also, practice beyond a certain point was detrimental to the ability to transfer what was learned. PMID- 10710766 TI - Relationship between years training and performance time on the three Taikyoku kata for Shotokan karate black-belts. AB - 27 experienced black-belt subjects were individually timed on each of the three Taikyoku kata and correlations calculated between performance time and years training. Correlations were nonsignificant, more clearly clarifying the reasons why performance time increases as years of training increases in more complex forms. PMID- 10710767 TI - Geophysical variables and behavior: LXXXVIII. Atmospheric electromagnetism: the possible disturbing influence of natural sferics on ESP. AB - Sferics are electromagnetic impulses generated by electrical discharges during thunderstorms (lightning). One category is comprised of very low frequency electromagnetic waves, traveling over distances up to a thousand kilometers. Sferics have been shown to affect biological responses such as pain syndromes, reaction times, and power in the alpha band of the EEG. In the present study, in which 100 subjects took part, sferics have been studied in their relation to performance on a forced-choice extrasensory perception (ESP) task and to several secondary variables. The general finding is a negative correlation between ESP performance and sferics activity around the time of the session, most notably 24 48 hours prior to the session. Secondary variables appear to modulate this correlation, as has been found in previous research on sferics: the correlation tended to be stronger for persons who scored lower on Neuroticism and higher on the Openness scale of a Five-Factor Personality Questionnaire. PMID- 10710768 TI - Acoustic characteristics of vocal emotions simulated by actors. AB - This paper reports on a set of acoustic measures of vocal emotions simulated by a skilled actor and actress who read 12 short sentences. The emotions simulated were Neutral, Cold Anger, Hot Anger, Happiness, Sadness, Interest, and Elation. Acoustic measures were examined for digitised samples of the 168 sentences (2 speakers x 12 sentences x 7 emotions). These measures included the over-all mean energy, standard deviations of energy, mean fundamental frequency, mean values of the first six formant frequencies and their corresponding frequency bandwidths, mean durations of utterance, and mean articulation rate. Results displayed both similarities and differences in the acoustic measures of the two actors' simulations of the different emotions. These are presented and discussed. PMID- 10710769 TI - Perception of time: delay of estimation under auditory and visual tasks with type A measures. AB - The perception of time is likely to be lengthened when estimates of duration are delayed rather than given immediately as reported by Vitulli and Crimmins in 1999. The present study sought boundary conditions for this robust effect. A multivariate analysis of variance showed a reliable main effect for delay of estimation, yet comparisons between auditory versus visual short-term memory tasks showed no differences in time estimations. The analysis showed weak interactions between sex and delay of estimation and between sex and delay of estimation on Jenkins Activity Survey-Job Involvement scores. PMID- 10710770 TI - Classroom management programs for deaf children in state residential and large public schools. AB - Personnel in 4 randomly selected state residential schools for the deaf and 3 randomly selected large public schools with programs for the deaf were surveyed to assess the types of management or disciplinary programs and strategies currently in use with deaf students and the rated effectiveness of such programs. Several behavioral management programs were identified by respondents, with Assertive Discipline most often listed. Ratings of program effectiveness were generally above average on a number of qualitative criteria. PMID- 10710771 TI - A follow-up study of seven siblings with unusual sound preferences. AB - This is a follow-up descriptive study of seven siblings, ranging in age from 6 to 14 years who had never attended school or received speech therapy until these ages. Five of the seven siblings, along with their parents, and paternal grandmother were located in the Spring of 1997. All siblings have achieved normal weight and stature. They also achieved intelligible speech with no idiosyncratic sound patterns. PMID- 10710772 TI - Laboratory studies on the effects of temperature variations on drowsiness. AB - 20 subjects were analysed in a laboratory study investigating the inhibition of drowsiness by altering air temperature. The EEGs of the subjects were used to measure the effect of sequences of temperature lowerings of 10 degrees for 2- and 4-min. periods. The effect upon wakefulness was analysed through EEG recordings and subjective ratings. Results indicated that reductions of the air temperature of 10 degrees during repeated 2- and 4-min. periods significantly increased wakefulness. PMID- 10710773 TI - Dysfunction in smooth pursuit eye movements and history of childhood trauma. AB - Several commentators recently have advocated the view that a deficit in the performance of a smooth pursuit eye-movement task is a biological marker of the genetic predisposition to schizophrenia. This study considered the possibility that such an impairment is due in part to experiential or acquired characteristics, and specifically, to a history of childhood trauma. A sample of 100 Australian adults performed a visual tracking task and completed a self report measure of childhood trauma. Although the effect size was small, a relationship was found between eye-tracking performance and a childhood history of physical and emotional abuse. This finding suggests that eye-tracking performance may not be governed entirely by genetic factors, a possibility that has implications for the use of indices of smooth pursuit eye movement as a purely genetic marker of proneness to schizophrenia. Further investigation is needed to clarify the basis of the association between these deficits and childhood abuse. PMID- 10710774 TI - Synchronization, anticipation, and consistency in motor timing of children with dimensionally defined attention deficit hyperactivity behaviour. AB - We tested the hypothesis that children with hyperactive behaviour are impaired in the temporal organization of their motor output. The performance of 11 boys, scoring above a cut-off on standard scales of overactivity and inattention, was compared to that of controls in progressively more complex Motor-timing tasks. The tasks administered required self-paced and externally paced Sensorimotor Synchronization and Sensorimotor Anticipation. Deficits at a perceptual level were investigated with a Time-discrimination task. As hypothesized, we found that hyperactive children had no deficits in their perception of time but were impaired in timing their motor output. Hyperactive children were more inconsistent than controls in maintaining a freely chosen tapping rhythm, in synchronizing and in anticipating their motor response to external visual stimulation. PMID- 10710776 TI - Europa's complexion. PMID- 10710775 TI - Is there more than one source for the temporal binding factor for human consciousness? AB - If the emergence of higher order consciousness is produced by transcerebral electromagnetic processes recreated every 20 msec. to 25 msec., then any frequency band in addition to the 40-Hz range could serve as a binding factor for this process. Second order differences or "derivatives," obtained by lagging differences in real time between adjacent frequency integers, indicate that 20 msec. to 25 msec. intervals could be continuously recreated by shifts in frequency within approximately 6.3 and 7.8 Hz. Higher order differences indicate that variations within very narrow band widths between 5 Hz and 6 Hz would generate continuous recreations of 20-msec. to 25-msec. intervals. PMID- 10710777 TI - Ancient atomists. PMID- 10710778 TI - NASA's not shining moments. PMID- 10710779 TI - An elemental mystery. PMID- 10710780 TI - The nonnegligible lightness of gravity. PMID- 10710781 TI - Methane fever. PMID- 10710782 TI - Violent opposition. PMID- 10710783 TI - Asbestos in the air. PMID- 10710784 TI - Gene therapy setback. PMID- 10710785 TI - The Galileo mission to Jupiter and its moons. PMID- 10710786 TI - Melting below zero. PMID- 10710788 TI - Digital materials and virtual weathering. PMID- 10710787 TI - The early origins of autism. PMID- 10710789 TI - Capturing greenhouse gases. PMID- 10710790 TI - Transparent animals. PMID- 10710791 TI - Uprooting the tree of life. PMID- 10710792 TI - Gamma-ray bursts come home. PMID- 10710793 TI - Real and virtual sculptures. PMID- 10710794 TI - Managed behavioral health services: a bibliography of empirical studies, articles of interest, and books. PMID- 10710795 TI - Risk factors for cervical intraepithelial neoplasm in Alaska Native women: a pilot study. AB - Although rates for invasive cervical cancer have declined over the past twenty years among Alaska Native women, they continue to show high rates of pre-invasive cervical lesions (cervical intraepithelial neoplasia, or CIN). We investigated risk factors for CIN II/III among Alaska Native women in a pilot case-control study. Cases (n = 26) included women with biopsy-proven CIN II/III, while controls (n = 52) had normal cervical epithelium. The strongest risks associated with CIN II/III were HPV infection of any type (Crude Odds Ratio [OR] 8.4, 95% Confidence Interval [CI]: 2.9-29.4), HPV 16 infection (OR 40.8, 95% CI: 9.4 176.4), and a family history of cervical dysplasia (OR 3.9, 95% CI: 1.3-11.3). We also found that use of depot-medroxy progesterone acetate was associated with CIN (OR 3.0, 95% CI: 1.1-8.5). A larger investigation would be necessary to allow adequate evaluation of these, and other, risk factors for CIN among Alaska Native women. PMID- 10710796 TI - Respiratory syncytial virus: current status and hope for the future. PMID- 10710797 TI - Frontier medicine in SE Alaska. Harry Carlos DeVighne, MD. 9/3/1867 - 8/7/1957. PMID- 10710798 TI - Hypnosis: a window into the soul of healing. PMID- 10710799 TI - Era III medicine. PMID- 10710800 TI - Era III medicine. PMID- 10710801 TI - The mind/body continuum. PMID- 10710802 TI - The mind/body link in essential hypertension: time for a new paradigm. AB - The origin of essential hypertension is believed by many to be at least partially emotion-related. A widely held paradigm is that perceived emotional distress raises blood pressure and leads eventually to sustained hypertension. However, decades of research have not provided strong or consistent support for this view. The purpose of this article is to briefly review this research, and to present a very different view of the mind-body link of hypertension. This view focuses on the role of emotions that are not consciously perceived, emotions that are unknowingly kept from conscious awareness, and largely ignored by patients, physicians and research. It suggests that the mind/body connection is often operative when we least suspect it. The evidence for this understanding, and the important implications regarding treatment of hypertension and other unexplained medical conditions with a suspected mind/body link, are discussed. PMID- 10710803 TI - A review of mind/body therapies in the treatment of musculoskeletal disorders with implications for the elderly. AB - BACKGROUND: A comprehensive, but not systematic, review of the research on complementary and alternative treatments, specifically mind/body techniques, on musculoskeletal disease was conducted at Stanford University. The goals of the review were to establish a comprehensive literature review and provide a rationale for future research carrying the theme of "successful aging." METHODS: Computerized searches were conducted using MEDLINE, PsychInfo, Stanford Library, Dissertation Abstracts, Lexus-Nexus, the Internet as well as interviews conducted with practitioners and the elderly. Mind/body practices evaluated were: social support, cognitive-behavioral therapy, meditation, the placebo effect, imagery, visualization, spiritual/energy healing, music therapy, hypnosis, yoga, tai chi, and qigong. Studies published after 1990 were the priority, but when more recent literature was scarce, other controlled studies were included. RESULTS: Mind/body techniques were found to be efficacious primarily as complementary treatments for musculoskeletal disease and related disorders. Studies provided evidence for treatment efficacy but most apparent was the need for further controlled research. CONCLUSIONS: Reviewers found a dearth of randomized controlled research conducted in the US. There is a lack of studies with which to determine appropriate dosage and understand the mechanisms by which many of the practices work. Anecdotal evidence, some controlled research, clinical observation, as well as the cost effectiveness and lack of side effects of the mind/body treatments make further investigation a high priority. PMID- 10710804 TI - A homeopathic origin for placebo controls: 'an invaluable gift of God'. AB - The acknowledged early adoption of placebo controls in drug trials by homeopaths is currently thought to have been derived from prior external attempts to discredit the system. This claim is reexamined in the light of a comprehensive literature search for 19th-century homeopathic therapeutic trials and provings using placebo. Single-blind placebo controls, still used today, are shown to have originated independently within homeopathy's own disciplinary matrix before the first external evaluations. They are the most likely source for later placebo controlled crossover and parallel group experiments by homeopaths. PMID- 10710805 TI - Randomized prospective double-blind placebo-controlled study of dextrose prolotherapy for knee osteoarthritis with or without ACL laxity. AB - CONTEXT: Use of prolotherapy (injection of growth factors or growth factor stimulators). OBJECTIVE: Determine the effects of dextrose prolotherapy on knee osteoarthritis with or without anterior cruciate ligament (ACL) laxity. DESIGN: Prospective randomized double-blind placebo-controlled trial. SETTING: Outpatient physical medicine clinic. PATIENTS OR OTHER PARTICIPANTS: Six months or more of pain along with either grade 2 or more joint narrowing or grade 2 or more osteophytic change in any knee compartment. A total of 38 knees were completely void of cartilage radiographically in at least 1 compartment. INTERVENTION: Three bimonthly injections of 9 cc of either 10% dextrose and .075% lidocaine in bacteriostatic water (active solution) versus an identical control solution absent 10% dextrose. The dextrose-treated joints then received 3 further bimonthly injections of 10% dextrose in open-label fashion. MAIN OUTCOME MEASURES: Visual analogue scale for pain and swelling, frequency of leg buckling, goniometrically measured flexion, radiographic measures of joint narrowing and osteophytosis, and KT1000-measured anterior displacement difference (ADD). RESULTS: All knees: Hotelling multivariate analysis of paired observations between 0 and 6 months for pain, swelling, buckling episodes, and knee flexion range revealed significantly more benefit from the dextrose injection (P = .015). By 12 months (6 injections) the dextrose-treated knees improved in pain (44% decrease), swelling complaints (63% decrease), knee buckling frequency (85% decrease), and in flexion range (14 degree increase). Analysis of blinded radiographic readings of 0- and 12-month films revealed stability of all radiographic variables except for 2 variables which improved with statistical significance. (Lateral patellofemoral cartilage thickness [P = .019] and distal femur width in mm [P = .021]. Knees with ACL laxity: 6-month (3 injection) data revealed no significant improvement. However, Hotelling multivariate analysis of paired values at 0 and 12 months for pain, swelling, joint flexion, and joint laxity in the dextrose-treated knees, revealed a statistically significant improvement (P = .021). Individual paired t tests indicated that blinded measurement of goniometric knee flexion range improved by 12.8 degrees (P = .005), and ADD improved by 57% (P = .025). Eight out of 13 dextrose-treated knees with ACL laxity were no longer lax at the conclusion of 1 year. CONCLUSION: Prolotherapy injection with 10% dextrose resulted in clinically and statistically significant improvements in knee osteoarthritis. Preliminary blinded radiographic readings (1-year films, with 3-year total follow-up period planned) demonstrated improvement in several measures of osteoarthritis severity. ACL laxity, when present in these osteoarthritic patients, improved. PMID- 10710807 TI - Healing journey spans high-tech, high-touch at Hawaiian hospital. PMID- 10710806 TI - Earl E. Bakken. Building a healing hospital. Interview by Bonnie Horrigan. PMID- 10710808 TI - Integrating manual and movement therapy with philosophical counseling for treatment of a patient with amyotrophic lateral sclerosis: a case study that explores the principles of holistic intervention. AB - A patient (Travis) suffering from ALS received a holistic principle-based protocol that combined manual-movement techniques with philosophical counseling. After 4 sessions, he exhibited a remarkable improvement in head-neck alignment, balance/mobility, autonomic activity, and worldview for a 2-month span. These changes occurred only after his worldview underwent a shift from a dualistic split of mind and body to a nondualistic orientation. After this 2-month period of improvement, Travis' structural alignment and balance/mobility suddenly deteriorated rapidly. Yet, his enhanced worldview and autonomic tone continued through a final follow-up taken 17 weeks after the initial evaluation. PMID- 10710809 TI - In search of a deep psychobiology of hypnosis: visionary hypotheses for a new millennium. AB - This search for the deep psychobiological foundations of hypnosis begins with a review of some of the paradoxes of historical hypnosis and the impasse of current theory. It is proposed that further progress requires a deeper investigation of how psychosocial cues can modulate the mechanisms of healing at the CNS, autonomic, neuroendocrine and cellular-genetic levels. The dynamics of hypnotic communication and healing from the cognitive-behavior level to the cellular genetic are outlined in four stages: (1) Information transduction between the experiences of consciousness and the limbic-hypothalamic-pituitary system; (2) The psychosomatic network of messenger molecules and their receptors; (3) The immediate early gene protein cascade; and (4) State dependent memory, learning and behavior. Neuroscience research is outlined for its contributions to a mathematical model of how a psychobiological approach to the therapeutic applications of hypnosis and the placebo response could facilitate neurogenesis in the human hippocampus and healing at the cellular-genetic-protein level throughout the body. A series of ten hypotheses is proposed as a guide for theory and research in therapeutic hypnosis utilizing DNA chip technology in the new millennium. PMID- 10710810 TI - A deeper understanding of hypnosis: its secrets, its nature, its essence. AB - Numerous research projects converge on the conclusion that there are three major types of very good or highly responsive hypnotic subjects: (a) fantasy-prone individuals who have secretly spent much of their time since childhood fantasizing vividly and realistically; (b) amnesia-prone individuals who have developed special abilities for mentally repressing or compartmentalizing undesired memories, thoughts, and emotions; and (c) positively-set individuals who are maximally ready to cooperate, think-with, and imagine what is suggested to the best of their ability while letting go of contrary thoughts. The major principle that provides a deep understanding of hypnosis and hypnotic phenomena is that all hypnotic subjects are affected, albeit in different ways for different types of subjects, by four powerful behavior-determining factors that can be potentially maximized in hypnotic situations: (a) social factors that obligate the socialized subject to cooperate and try to actualize or realize the hypnotist's expectations and explicit suggestions; (b) the hypnotist's unique skills and personal characteristics (including creative ideas, communicative ability, and interpersonal efficacy) and the nature of the hypnotist-subject interpersonal relationship; (c) the effectiveness of the induction procedure in guiding the subject to think-with the suggestions; and (d) the depth of meaning, creativity, and "force" or "power" of the suggested ideas. PMID- 10710811 TI - The response set theory of hypnosis. AB - The response set theory of hypnosis (Kirsch & Lynn, 1997) is an extension of response expectancy theory (Kirsch, 1985), which is rooted in social cognitive approach to understanding human experience and behavior. Although the idea of a uniquely hypnotic altered state of consciousness is rejected, so too are compliance-based explanations of hypnotic behavior. Suggestions, in or out of the social context of hypnosis, can produce profound alterations in experience, including dissociative experiences, and these can be verified through corresponding changes in brain physiology. Response expectancies play a major, but not exclusive, role in the production of subjective experience in general (Kinsbourne, 1998) and therefore of these suggested experiences as well. Because of widespread cultural beliefs about hypnosis, the hypnotic context intensifies the effects of suggestion for most people, and this is the primary source of its therapeutic effects. PMID- 10710813 TI - Chronic alcohol abuse and nutritional status: recent acquisitions. PMID- 10710812 TI - The clinical importance of sociocognitive models of hypnosis: response set theory and Milton Erickson's strategic interventions. AB - This article documents the contributions and clinical relevance of influential sociocognitive models of hypnosis. We argue that an appreciation of the influence and interplay of sociocognitive constructs, combined with a knowledge of basic research findings, can contribute to sound clinical practice. This article extends previous statements of response set theory (e.g., Kirsch & Lynn, 1998, 1999; Lynn, 1998) by further elucidating the social and cognitive underpinnings of how response sets are established, maintained, and strengthened. It does so by providing a scientific rationale for Milton H. Erickson's most prominent strategic interventions. PMID- 10710815 TI - Effects of insulin-oral hypoglycemic agents combined therapy in outpatients with type 2 diabetes. AB - To evaluate the efficacy of combined insulin-OHAs therapy in subjects with NIDDM who received treatment with OHAs and insulin alone, we selected 60 outpatients divided in two groups: Group A: 36 subjects treated with OHAs therapy that received insulin treatment for secondary failure; Group B: 24 subjects in which OHAs therapy was added to insulin regimen to avoid the effects of hyperinsulinization. In the group A body weight increased significantly (+1.94 +/ 2.80 kg, p < 0.001 vs baseline), while in group B no gain of body weight was observed. Both groups showed a similar improvement of glycemic control. For the group A, the FPG and HbA1c decreased, respectively, from 14.64 +/- 3.76 to 8.72 +/- 2.92 mmol/l and from 9.10 +/- 0.30 to 7.20 +/- 0.53% at 6 months (p < 0.001). For the group B FPG and HbA1c decreased, respectively, from 12.05 +/- 3.49 to 8.24 +/- 3.01 mmol/l and from 8.3 +/- 0.1 to 6.8 +/- 0.13% (p < 0.001). Plasma cholesterol, triglycerides and uric acid concentrations did not show significant changes in either group. Insulin requirement in group A was 0.21 +/- 0.13 U/Kg/day. Despite of improvement of glycemia, total insulin requirement decreased in Group B from 0.53 +/- 0.25 to 0.34 +/- 0.2 U/Kg/day after OHAs therapy (p < 0.001). In the group A the bedtime insulin administration was prevalent (52.68%), while the most patients of group B needed a second or a third daily insulin injection (83.33%). In conclusion, in type 2 diabetic patients, therapy with combination of OHAs and insulin was associated with lower insulin doses and less weight gain. PMID- 10710814 TI - Atopic diseases among adults in Sor-Varanger community, northern Norway. An epidemiological study in an arctic area influenced by Russian industrial pollution. AB - BACKGROUND: Atopic diseases are common in western industrialized countries and their prevalence appears to be increasing. The prevalence seems to be higher in the cold, northerly regions of Europe. AIM: The aim of the study was to assess the prevalence of atopic diseases among adults in Sor-Varanger community, northern Norway, an arctic area at 70 degrees latitude north influenced by industrial pollution from smelting plants on the Kola peninsula. PATIENTS AND METHODS: The parents of all 575 schoolchildren aged 7-12 years in Sor-Varanger community (northern Norway) received a four page questionnaire concerning home environment and symptoms of allergy. A total of 1102 adults filled in and returned the questionnaire. RESULTS: 25.2% of the adults reported past and/or present symptoms (prevalence) of atopic diseases, whereas 38.7% of the children reported atopic symptoms. The prevalence of eczema occurred in 15.9%, followed by allergic rhinoconjunctivitis (10.3%) and asthma (5.6%), however, a significant difference between sexes was only found for eczema (p < 0.0005). Women smoked more frequently (45.6%) than men (39.0%). Keeping of furred pets occurred in 54.3% and dampness in 3.6% of the homes. CONCLUSION: This study indicates that the frequency of atopic diseases among adults is only two thirds of that reported in schoolchildren. Thus, the increased prevalence of atopic diseases over one generation could point towards factors associated with western lifestyle and living conditions (allergens and adjuvant factors). PMID- 10710816 TI - Pathophysiology, clinical features and management of hepatorenal syndrome. AB - Hepatorenal syndrome (HRS) is a form of functional renal failure occurring in patients with advanced liver disease. Hypoperfusion of the kidney, due to renal vasoconstriction, is the main feature of HRS. Conversely, the extrarenal circulation is characterized by low systemic resistance, especially occurring in splanchnic vessels, and arterial hypotension. It has been postulated that renal vasoconstriction is induced either by a hepatorenal reflex related to the diseased liver or by arterial vasodilation and the subsequent baroreceptor mediator activation of systemic vasoconstrictor factors. The diagnosis of HRS requires the exclusion of other causes of renal failure in patients with liver disease. On the basis of clinical and prognostic differences, two types of HRS have been defined. The prognosis of HRS is poor and, to date, the only effective treatment is the liver transplantation. PMID- 10710817 TI - Is apoptosis cause of pre-eclampsia? AB - Pre-eclampsia is the main cause of fetal and maternal morbidity and death associated with hypertension during complicating pregnancy. During physiological pregnancy, the immunological system undergoes secondary modifications, with an "exchange" between mother and fetus. Cytokines play an important role in the complex condition of partial fetal "rejection". It has suggested that the condition depend on immunological factors. In line with this hypothesis, apoptosis appear to play a key role in the pathophysiology of placental ischemia and the mechanism underlying this condition may be influenced by substances such as Bcl-2 which inhibits apoptosis. Neither aspirin nor calcium appear to improve maternal hypertension and proteinuria, although late ongoing trials may alter this view. At present, the condition can be resolved only by the end of pregnancy. Further studies are required in order to improve our understanding of these immunological mechanisms underlying hypertension during pregnancy, as the key to effective therapy may be their ability to "manipulate" them in an appropriate way. PMID- 10710818 TI - Estrogens and euosteogenesis in men. AB - The bone mineral density (BMD) has been analyzed in 200 male patients divided in 4 groups of age as follows: (A) 40-49, (B) 50-59, (C) 60-69, and (D) 70 years and above. BMD was measured by using the DEXA technique both in the ultradistal and mediodistal region of radius of the non-dominant side. In addition, the serum levels of testosterone (Ts), dihydrotestosterone (DHT) and 17-beta estradiol (E 2) have also been measured. The data obtained have shown that bone mineral density values are decreasing also in the males with advancing age, and the positive correlation (p < 0.05) of BMD with the E-2 levels also tend to decrease. These results suggest the hypothesis that the true sexual hormones regulating the rhythm of osteogenesis may be the estrogens in the males, too. PMID- 10710819 TI - The effect of a carbohydrate loading on running performance during a 25-km treadmill time trial by level of aerobic capacity in athletes. AB - The aim of the present study was to examine the influence of a high carbohydrate diet and the level of aerobic capacity on running performance during a 25-km treadmill time trial. The study used a 2*2 design with the factors being training and diet composition. We divided the athletes in 4 groups: 1. Trained athletes with carbohydrate loading (CHO1); 2. Trained athletes without carbohydrate loading (C1); 3. Untrained athletes with carbohydrate loading (CHO2); 4. Untrained athletes without carbohydrate loading (C2). The carbohydrate loading was effected with confectionery. Performance time, running speed, blood glucose and blood lactate concentrations were evaluated during two 25-km treadmill time trial (trial 1 and trial 2) separated by 7 days in which two groups (CHO1 and CHO2) had a carbohydrate loading. The results showed that the athletes with lower level of aerobic capacity had better performance time after carbohydrate loading. They ran faster and had a higher glucose and lactate concentrations in the last 5 km during trial 2. There were no significant differences in the other groups. In conclusion, we can assert that dietary carbohydrate loading can improve running performance and that confectionery can be used as an effective means of supplementing the normal carbohydrate intake in preparation for endurance competitions. But the improvement depends on some factors such as the distance and the level of aerobic capacity. PMID- 10710820 TI - Anaphylaxis from peanut oil in infant feedings and medications. AB - Peanut is a very dangerous allergen. Too often one is unaware that peanut allergens can be found everywhere, hidden in a myriad of foods, and without any indication of their presence. It is stressed that not even infant feedings and medications can be exempt, thus leading to anaphylaxis. PMID- 10710821 TI - Local anaesthesia with eutectic cream of lidocaine and prilocaine for treatment of cicatrizial phimosis in outpatients. AB - The authors report their experience in ambulatory treatment of cicatrizial phimosis in outpatients, by using local anaesthesia with eutectic cream of lidocaine and prilocaine. This preliminary study shows that preparation seems to be effective and well tolerated. To the aim of getting a good analgesic result the most important aspects are a careful application of the cream all over the concerned area and an occlusive bandage for 60-90 minutes at least. PMID- 10710822 TI - Helicobacter pylori and ischemic heart disease: current evidences. PMID- 10710823 TI - Comparison of two microemulsion of cyclosporine A in healthy volunteers. AB - This study evaluated the bioequivalence of a new Cyclosporine A microemulsion formulation in comparison to the reference market standard. Twenty-four adult healthy volunteers were randomised to receive the two Cyclosporin A microemulsion formulations, at a dose of 2.5 mg/kg, according to a cross-over design. Blood samples were taken before drug administration and at 12 points within 24 hours. Cyclosporine A whole blood concentrations were determined by HPLC. The pharmacokinetic parameters AUC0-t and AUC0-infinity were calculated by the trapezoidal rule, Cmax and Tmax were obtained directly from blood data. AUCs and Cmax were tested for bioequivalence after log transformation of data, differences for Tmax were evaluated by the rank test of Wilcoxon for paired data. The 90% confidence interval ratio between tested/reference drug was 0.98 for AUC0-t, 0.96 for AUC0-infinity and 1.01 for Cmax. All of them were within the range of bioequivalence. Tmax was 1.60 +/- 0.44 hours after test drug and 1.67 +/- 0.48 after reference drug (p = 0.27, Wilcoxon test). According to these results the two Cyclosporine A microemulsion formulations can be considered bioequivalent. PMID- 10710824 TI - New cyclosporine microemulsion randomized, cross-over bioequivalence steady-state study in renal transplanted patients. AB - A new microemulsion formulation of cyclosporine was compared with the marketed formulation in 18 stable renal transplanted patients. Aim of the study was not only to determine the bioequivalence between the two pharmaceutical preparations, but also to ascertain whether tested drug could maintain stable blood concentrations of cyclosporine. Renal transplanted patients under cyclosporine treatment from at least 12 months at a well individualized dosage (resulting in 90-200 ng/mL of blood level drug) have been selected. Patients received the same preceding dose of cyclosporine through both the two preparations according to a cross-over, randomized schedule during 4 weeks in two equally divided daily administrations. Serial blood samples were obtained over a 24-hour period at steady-state of each formulation. Cyclosporine concentrations were determined by a specific immunoassay method (FPIA) n whole blood taken in the last day of each cycle of treatment. Statistical comparisons of cyclosporine levels (using pharmacokinetic parameters) were cross-performed between formulations and days of blood test. Tested drug resulted bioequivalent with the reference marketed formulation. Furthermore, the study showed that tested drug maintained satisfactory stable blood concentrations of cyclosporine. PMID- 10710825 TI - Neurinomas of the brachial plexus: case report. AB - Neurinomas, also referred to as neurilemmomas and schwannomas, are rare benign tumours of the peripheral nerves, a low proportion of which arise from the brachial plexus. Authors report a case of an ancient schwannoma arising from the brachial plexus. The tumour, usually asymptomatic, may cause sensory radicular symptoms, or rarely motor deficits in the involved arm. Enucleation of the tumour from the nerve without damage to any of the fascicles is the correct treatment. PMID- 10710826 TI - Malignant tumours of the small intestine: a case of jejunal adenocarcinoma. AB - The Authors analyse the main problems concerning malignant tumours of the small intestine. They report a case of jejunal adenocarcinoma recently observed. They emphasise the importance of surgery, both diagnostic and therapeutic, even in advanced stages, for the treatment of this neoplasm. PMID- 10710827 TI - Review--antibiotic treatment in inflammatory bowel disease: rifaximin, a new possible approach. AB - The etiology of inflammatory disease is still unknown, but a body of evidence from clinical and experimental observation indicates a role for intestinal microflora in the pathogenesis of this disease. Reduction of microflora using antibiotics, bowel rest and fecal diversion decreases activity in Crohn's disease and in ulcerative colitis. Several trials have been carried out on the use of antibiotic treatment in patients with active ulcerative colitis with contrasting results. A number of trials have been carried out using Rifaximin, a non absorbable broad-spectrum antibiotic, confirming the absence of systemic bioavalaibility of the drug even when administered at very high doses and for prolonged periods. It may therefore be useful in treatment of ulcerative colitis and pouchitis, since its absorption through inflamed mucosa is negligible, it maintains a topical action without systemic effects and the lack of resistant bacterial strains may allow prolonged and repeated treatments. PMID- 10710828 TI - Sleep disorders. AB - Sleep disorders occupay an important position among the pathologies which may precede the appearance of headache. Some authors consider sleep disorders an expression of a functional, biochemical and/or neurotransmission alteration at central nervous system level. Sleep disorders may be distinguished, according to the Association Sleep Disorders classification in: alteration of the sleep-awake cycle, hypersomnia, parasomnia, insomnia. We observed 1876 normal children ranging from 3 to 14 years of age, 1073 (60.4%) of whom presented sleep disorders. Few studies have been carried out on the incidence of sleep disorders on casistics of healthy children. Date reported in literature state that sleep disorders do not exceed 25% of cases that is not more than one child out of four presents sleep disorders. This percentage is much lower than the 60.4% rate observed by us in children suffering from primary headache. Our results stress the importance of sleep disorders as a cephalalgic risk factor. PMID- 10710829 TI - Evaluation of anxiety and depression in childhood migraine. AB - Childhood migraine can be the expression of an unconscious attempt of the small patient to show a discomfort which is denied through the defence of somatization. We considered a sample of 73 children, 39 males and 34 females suffering from migraine. We evaluated the presence of emotional disorders through diagnostic interviews consisting of one by one submission of the Anxiety Scale Questionnaire for Evolutive Age and the Children Depression Scale Test. Within our sample we are able to distinguish three groups: a first group negative for both anxiety and depressive disorders, thus defined as control group; a second group presenting anxiety depressive disorders and a third one presenting a mostly depressive symptomatology. We found a significantly higher incidence of migraine in male firstborn children belonging to the group with a condition associated to anxiety and depression. PMID- 10710830 TI - Undergraduate medical examination an aid or a penalty. PMID- 10710831 TI - Allergy diagnosis by Pharmacia Cap System. AB - From September 1995 to September 1998, sera from 959 suspected allergy patients have been tested by the new Pharmacia Cap System. Of these, 80 per cent were diagnosed to suffer from some allergy while 20 per cent reacted negative. It was found that the CAP system gave accurate and clear cut results to the satisfaction of the patient and referring physician. PMID- 10710832 TI - Newer approaches in increasing life span. AB - Based on ideal conditions technical life span of human kind is approximately 110 120 years. Although number of studies including calorie restriction and antiparkinsonism drug (deprenyl) have indicated increased life span in animals, it is premature to expect them to increase life span in man. However, current studies like activation of immune system with DHEA in man and anticipation of antioxidant therapy contributing to increased life span are encouraging. Practice of meditation particularly TM and balanced diet might be contributory. PMID- 10710833 TI - Tetanus: study of 8,697 cases. AB - Records of 8,697 cases of tetanus seen over a period of 14 years are analysed. Overall mortality was 48.0%. Mortality in neonatal group was 86.38% whereas that in non-neonatal group it was 40.18%. Disease was seen more frequently in male than in female. Mortality in male was lower than in female. Incidence was highest in the first decade of life. Mortality was lowest (about 33%) in first two decades (excluding neonatal group). Mortality in neonatal group was highest (86.38%). Mortality was inversely related to length of incubation period. In cases with incubation period of 7 days or less, mortality was 58.26% in non neonates and 94.15% in neonates. Mortality was very low (2.14%) in 2,100 cases who did not develop spasms. In cases with spasms mortality was inversely related to the length of period of onset. Temperature of 100 degrees F within first 24 hours of admission was an adverse factor and these cases had higher mortality. Cases were divided into five grades according to the severity. Mortality in each grade was significantly different from that in the other. Mortality was lower in otogenic tetanus while it was higher in post-abortion and post-injection tetanus. Tetanus following penetrating injury carried higher mortality whereas tetanus following abrasions had lower mortality. With head and face as the site of infection, mortality was low while it was high when the site of infection was a trunk. Results were similar with dose of A.T.S. ranging between 5,000 and 60,000 i.u. and tetanus immune globulin, whereas mortality was high with higher and lower dose of A.T.S. or with no A.T.S. Respiratory spasms, respiratory failure, respiratory complications and circulatory failure were the common causes of death. PMID- 10710834 TI - Estrogen therapy: advantage and disadvantages. PMID- 10710835 TI - Sequential interactions in the marital communication of depressed men and women. AB - Despite studies showing patterns of sequential interaction between depressed wives and their husbands, no published research has contrasted sequential interactions of depressed husbands and their wives. This study compared problem solving interactions of 49 couples with a depressed husband, 41 with a depressed wife, and 50 normal controls. Interactions were coded using the Marital Interaction Coding System. Although no clear patterns of sequential interaction distinguished couples with a depressed wife from normal control couples, results suggested a unique pattern of interaction between depressed husbands and their spouses, whereby positive communications from the husband resulted in decreased positivity and increased negativity from their wives. Given the importance of positivity for promoting effective problem solving, this pattern appears to have important implications for couples' long-term marital satisfaction and husbands' mood regulation. PMID- 10710836 TI - The effects of treatment compliance on outcome in cognitive-behavioral therapy for panic disorder: quality versus quantity. AB - Cognitive-behavioral therapy (CBT) is skill based and assumes active patient participation in regard to treatment-related assignments. The effects of patient compliance in CBT outcome studies are equivocal, however, and 1 gap in the literature concerns the need to account for the quality versus the quantity of assigned work. In this study, both quality and quantity of home-based practice were assessed to better evaluate the effects of treatment compliance in patients with panic disorder (N = 48) who participated in a 12-session CBT protocol. Patient estimates of compliance were not significantly associated with most outcome measures. On the other hand, therapist ratings of compliance significantly predicted positive changes on most outcome measures. Moreover, therapist and independent rater estimates of the quality of the participant's work, relative to the quantity of the work, were relatively better predictors of outcome. PMID- 10710837 TI - Risk factors for adolescent substance abuse and dependence: data from a national sample. AB - A national household probability sample of 4,023 adolescents aged 12 to 17 years was interviewed by telephone about substance use, victimization experiences, familial substance use, and posttraumatic reactions to identify risk factors for Diagnostic and Statistical Manual of Mental Disorders--(4th ed.; American Psychiatric Association, 1994) defined substance abuse/dependence. Age and ethnicity data were available for 3,907 participants. Major findings were (a) adolescents who had been physically assaulted, who had been sexually assaulted, who had witnessed violence, or who had family members with alcohol or drug use problems had increased risk for current substance abuse/dependence; (b) posttraumatic stress disorder independently increased risk of marijuana and hard drug abuse/dependence; and (c) when effects of other variables were controlled, African Americans, but not Hispanics or Native Americans, were at approximately 1/3 the risk of substance abuse/dependence as Caucasians. PMID- 10710838 TI - Cognitive-behavioral stress management intervention effects on anxiety, 24-hr urinary norepinephrine output, and T-cytotoxic/suppressor cells over time among symptomatic HIV-infected gay men. AB - The present study tested the effects of a multimodal cognitive-behavioral stress management (CBSM) intervention on anxious mood, perceived stress, 24-hr urinary catecholamine levels, and changes in T-lymphocyte subpopulations over time in symptomatic HIV+ gay men. Seventy-three men were randomized to either a group based CBSM intervention (n = 47) or a wait-list control (WLC) condition (n = 26). Men assigned to CBSM showed significantly lower posttreatment levels of self reported anxiety, anger, total mood disturbance, and perceived stress and less norepinephrine (NE) output as compared with men in the WLC group. At the individual level, anxiety decreases paralleled NE reductions. Significantly greater numbers of T-cytotoxic/suppressor (CD3+CD8+) lymphocytes were found 6 to 12 months later in those assigned to CBSM. Moreover, greater decreases in NE output and a greater frequency of relaxation home practice during the 10-week CBSM intervention period predicted higher CD3+CD8+ cell counts at follow-up. PMID- 10710839 TI - Does psychotherapy homework lead to improvements in depression in cognitive behavioral therapy or does improvement lead to increased homework compliance? AB - The bidirectional causal relationships between psychotherapy homework (HW) compliance and changes in depression were assessed in 2 groups of depressed outpatients treated with cognitive-behavioral therapy using nonrecursive structural equation modeling techniques. The data were consistent with the hypothesis that HW compliance had a causal effect on changes in depression, and the magnitude of this effect was large. Patients who did the most HW improved much more than patients who did little or no HW. In contrast, depression severity did not appear to influence HW compliance. HW compliance did not appear to be a proxy for any other, unobserved (3rd) variable, such as motivation. Although the causal effect of HW on depression was large, the correlation between HW and depression was small. Some possible explanations, along with suggestions for future studies, are proposed. PMID- 10710840 TI - Predicting the 2 1/2-year outcome of dysthymic disorder: the roles of childhood adversity and family history of psychopathology. AB - Follow-up studies of dysthymic disorder (DD) indicate that demographic and clinical variables are not strong predictors of its outcome. The present study extended this literature by examining the relationship between the early home environment and family history of psychopathology and outcome in DD. Eighty-six outpatients with DD were followed up over a 30-month period using structured clinical interviews. A number of measures of childhood adversity and familial psychopathology assessed at baseline predicted outcome, even after controlling for baseline severity and clinical variables. The best predictors included a history of sexual abuse, quality of the patient's relationship with both parents, and higher familial loadings for drug abuse and Cluster A personality disorders. These findings indicated that childhood adversity and familial psychopathology have greater predictive utility for DD than for demographic and clinical variables. PMID- 10710841 TI - Contingent reinforcement increases cocaine abstinence during outpatient treatment and 1 year of follow-up. AB - This study assessed whether contingent incentives can be used to reinforce cocaine abstinence in dependent outpatients. Seventy cocaine-dependent outpatients were randomized into 2 conditions. All participants received 24 weeks of treatment and 1 year of follow-up. The treatment provided to all participants combined counseling based on the community reinforcement approach with incentives in the form of vouchers exchangeable for retail items. In 1 condition, incentives were delivered contingent on cocaine-free urinalysis results, whereas in the other condition incentives were delivered independent of urinalysis results. Abstinence-contingent incentives significantly increased cocaine abstinence during treatment and 1 year of follow-up compared with noncontingent incentives. PMID- 10710842 TI - Responsiveness of children with attention deficit-hyperactivity disorder to reward and response cost: differential impact on performance and motivation. AB - Using a within-subject design and both high- and low-interest tasks, this study examined the effects of reward (R), response cost (RC), and no contingency (NR) on performance and motivation of 22 children with attention deficit-hyperactivity disorder (ADHD) and 22 controls. Dependent variables included performance measures, self-rated performance and motivation, and a new measure of behavioral motivation based on a 2-min postcontingency task. Both contingencies benefited some aspects of the performance of ADHD children; relative to R, RC showed stronger effects but at the expense of decreased self-rated motivation on the low interest task. The performance of controls did not differ across tasks, whereas ADHD children performed relatively better on the high-interest task. Neither contingency decreased motivation measures relative to NR for either group. For ADHD children, motivational effects appeared to be influenced by self-perceptions of performance. PMID- 10710843 TI - Patterns of adjustment among children of battered women. AB - Children exposed to interparental violence have been characterized by an array of psychological problems, but findings regarding the precise nature of these problems have been inconsistent. This study used cluster analysis to determine whether distinct patterns of adjustment could be identified in 228 8- to 14-year old children residing in battered women's shelters. Five such patterns emerged: multiproblem-externalizing, multiproblem-internalizing, externalizing, mild distress, and no problems reported. This solution was cross-validated in independent halves of the sample and was similar for boys and girls. Differences among the clusters on relevant family and demographic variables were examined, and it was found that the clusters could be distinguished on the basis of the frequency of children's exposure to interparental violence, parent-child aggression, and children's appraisals of interparental conflict. PMID- 10710844 TI - Binge antecedents in obese women with and without binge eating disorder. AB - In this study, women with binge eating disorder (BED; n = 41) and weight- and age matched comparison women without BED (NBED; n = 38) monitored their eating for 6 days, using handheld computers to measure mood, appetite, and setting at all eating episodes and comparison noneating episodes. Poor mood, low alertness, feelings of poor eating control, and craving sweets all preceded binge episodes for the BED group. An unanticipated finding was the frequent report of binge episodes in the comparison group; only feelings of poor eating control and craving sweets predicted binge episodes in this group. Binge eating NBED women tended to experience worse mood, less control, and more craving than other NBED women, contributing to evidence of the close relationship of binge eating and decrements in emotional and appetitive functioning. PMID- 10710845 TI - Preventing relapse among former smokers: a comparison of minimal interventions through telephone and mail. AB - This study compared 2 minimal interventions for reducing relapse in ex-smokers. One intervention involved 12-month access to a telephone hot line. In the other intervention, 8 relapse-prevention booklets were mailed to participants over 1 year. The 2 interventions were crossed in a 2 x 2 factorial design, yielding control, hot-line-only, mailings-only, and combined conditions. The criterion of at least 1 week of abstinence at baseline was met by 584 participants, 446 of whom also completed a 12-month assessment. Repeated mailings, but not the hot line, reduced relapse for those participants who had been abstinent for less than 3 months at baseline. At follow-up, 12% of those in the mailings conditions were smoking again compared with 35% in the nonmailing conditions. As predicted, both interventions were effective at attenuating the association between depressive symptoms and poor outcome found in the control condition. PMID- 10710846 TI - Relation of therapeutic alliance and perfectionism to outcome in brief outpatient treatment of depression. AB - Prior analyses of the National Institute of Mental Health Treatment of Depression Collaborative Research Program demonstrated that perfectionism was negatively related to outcome, whereas both the patient's perception of the quality of the therapeutic relationship and the patient contribution to the therapeutic alliance were positively related to outcome across treatment conditions (S. J. Blatt, D. C. Zuroff, D. M. Quinlan, & P. A. Pilkonis, 1996; J. L. Krupnick et al., 1996). New analyses examining the relations among perfectionism, perceived relationship quality, and the therapeutic alliance demonstrated that (a) the patient contribution to the alliance and the perceived quality of the therapeutic relationship were independent predictors of outcome, (b) perfectionistic patients showed smaller increases in the Patient Alliance factor over the course of treatment, and (c) the negative relation between perfectionism and outcome was explained (mediated) by perfectionistic patients' failure to develop stronger therapeutic alliances. PMID- 10710847 TI - An interpersonal model of psychotherapy: linking patient and therapist developmental history, therapeutic process, and types of outcome. AB - This study empirically evaluated a 3-stage causal model based on interpersonal theory that relates patient and therapist early parental relations, the therapeutic alliance, and outcome. Data were from the Vanderbilt II database and encompassed 64 psychodynamic psychotherapies. Interpersonal variables were assessed using the Structural Analysis of Social Behavior. Initial support for the model was found, suggesting a direct effect of patient early parental relations on process and outcome, a direct effect of therapist early parental relations on process, and a direct effect of process on outcome--and thus indirect effects of both patient and therapist early parental relations on outcome mediated by the process. The psychotherapy process was assessed from 3 perspectives: patient and therapist self-report and reports by independent observers. Little convergence was found between the 3 perspectives. PMID- 10710849 TI - The role of complex thought in clinical prediction: social accountability and the need for cognition. AB - Research shows that clinical predictions are less accurate than statistical predictions and are held with unreasonable confidence. Because there are obstacles to the implementation of statistical prediction, factors that improve clinical judgment must be identified. One hundred twelve individuals participated in an experiment investigating the role of complex thought in clinical prediction. Results revealed marked performance differences related to the amount of available clinical information. Participants' assessed need for cognition was associated with their consistency, accuracy, and cue-weighting strategies. Social accountability improved confidence performance under certain task conditions but was unrelated to accuracy. Theoretical and practical implications of these results are discussed, with emphasis on the restructuring of tasks and the selection and training of human forecasters to promote accurate and appropriately confident clinical predictions. PMID- 10710848 TI - The timeline followback reports of psychoactive substance use by drug-abusing patients: psychometric properties. AB - The Timeline Followback (TLFB; L. C. Sobell & M. B. Sobell, 1996) interview, which uses a calendar method developed to evaluate daily patterns and frequency of drinking behavior over a specified time period, has well-established reliability and validity for assessing alcohol consumption. Although several investigators have used the TLFB to evaluate drug-using behavior, few studies have examined the psychometric properties of the interview for this purpose. The authors conducted TLFB interviews with a sample of adult drug-abusing patients seeking treatment for substance abuse (n = 113) at baseline, posttreatment, and quarterly thereafter for 12 months. It was found that the patients' reports about their drug consumption using this method generally had high (a) retest reliability, (b) convergent and discriminant validity with other measures, (c) agreement with collateral informants' reports of patients' substance use, and (d) agreement with results from patients' urine assays. PMID- 10710850 TI - Heterogeneity of variance in clinical data. AB - Traditional parametric (t, F) and nonparametric (Mann-Whitney-Wilcoxon U, Kruskal Wallis H) statistics are sensitive to heterogeneity of variance (heteroscedasticity). Moreover, there are theoretical reasons to expect, and empirical results to document, the existence of heteroscedasticity in clinical data. Transformations to reduce heteroscedasticity are problematic. This article reviews the literature on robust methods that are available and that should be widely used to control rate of Type I error and maintain power. No one robust method is ideal for all situations, but such methods are superior to the traditional tests. Specific recommendations are made for application under various conditions of heteroscedasticity. PMID- 10710851 TI - Power to detect homework effects in psychotherapy outcome research. AB - Studies of homework effects in psychotherapy outcome have produced inconsistent results. Although these findings may reflect the comparability of psychotherapy with and without homework assignments, many of these studies may not have been sensitive enough to detect the effects sizes (ESs) likely to be found when examining homework effects. The present study evaluated the power of homework research and showed that, on average, current power levels are relatively weak in controlled studies ranging from 0.58 for large ESs to 0.09 for small ESs. Thus, inconsistent findings between studies may very well be due to low statistical power. PMID- 10710852 TI - Clinician attributions associated with the diagnosis of schizophrenia in African American and non-African American patients. AB - The authors examined the schizophrenia diagnosis in 292 psychiatric inpatients in a largely African American community. Clinicians completed a free-response questionnaire that described their diagnostic decisions. Psychotic symptoms such as hallucinations, which were attributed to African American and non-African American patients at different rates, did not necessarily correspond to differences in diagnostic rates. Rather, symptoms not differentially attributed between groups often corresponded with higher rates of schizophrenia for African American patients. Attributions of negative symptoms showed the largest differences between African American and non-African American patients in rates of schizophrenia diagnosis; thought disorder equalized rates of the diagnosis between the 2 groups of patients. Logistic regression analyses suggested that different aggregate decision models were applied to patients of differing race. PMID- 10710853 TI - Tobacco withdrawal in women and menstrual cycle phase. AB - Because negative mood is a characteristic of both tobacco withdrawal and menstrual discomfort, withdrawal may vary by menstrual cycle phase. Tobacco withdrawal, mood, and menstrual discomfort were assessed in premenopausal women who quit smoking during either the follicular (Days 1-14 postmenstrual onset; n = 41) or luteal (Day 15 or longer postmenstrual onset; n = 37) phase of the menstrual cycle and maintained biochemically verified smoking abstinence during the postquit week. Women quitting during the luteal phase reported significantly greater increases in tobacco withdrawal and self-reported depressive symptoms than women quitting during the follicular phase. These results indicate that selecting a quit-smoking day early in the follicular phase may attenuate withdrawal and negative affect in premenopausal female smokers. PMID- 10710854 TI - Generality of Psychopathy Checklist-Revised factors over prisoners and substance dependent patients. AB - The Psychopathy Checklist-Revised (PCL-R; R. D. Hare, 1991) is an often-used device for assessment of adult antisociality. This research examined generalizability by replicating the 2-factor model for a sample of 326 male prisoners and assessing its congruence and relative reliability and specificity among 620 substance-dependent patients. Generality was assessed also across addiction subtypes (opioid, cocaine, and alcohol), age, gender, and ethnicity. The 2-factor model was found inappropriate for the substance-dependent samples, whereas a unidimensional model represented by the PCL-R total score was found generalizable across prison and substance-dependent samples. PMID- 10710855 TI - Selecting instruments for construct validity assessment. PMID- 10710856 TI - The Children's Cognitive Triad Inventory: reliability, validity, and congruence with Beck's cognitive triad theory of depression. AB - Depression, once thought rare in children, is now more widely recognized and believed to arise from negative views of self, world, and future, according to Beck's cognitive theory of depression. The Cognitive Triad Inventory for children measures the three negative views, and although reported as psychometrically adequate, this study extended previous analyses with confirmatory factor analysis in a sample of 122 school-aged children. Internal consistency was .82 (total scale) but ranged from .54 to .76 for subscales reflecting the views of self, world, and future. Confirmatory factor analysis revealed factors reflecting three aspects of the self rather than the three negative views. The findings suggest that Beck's theory about the negative cognitive triad may be less suitable for children than adults. PMID- 10710857 TI - Psychometric properties of the Index of Homophobia Scale in registered nurses. AB - The psychometric properties of the Index of Homophobia Scale (IHS; Bouton et al., 1987) were examined in a sample of registered nurses (n = 95). Scores on the IHS may range from 0 (no homophobia) to 28 (highly homophobic). This sample had relatively low total scores on the IHS (M = 7.99, SD = 5.41), indicating that subjects were predominantly not homophobic. Internal consistency as estimated by Cronbach's alpha was .88. The average item mean was 1.14 (SD = 0.29) ranging from .77 to 1.48. Corrected item-to-total scale correlations were satisfactory, ranging from .58 to .75. Inter-item correlations ranged from .35 to .67 indicating some redundancy in content sampling. The 7-item IHS indicates promise for future use with registered nurses, and perhaps also with other health care professionals in adequately and accurately measuring homophobia. PMID- 10710858 TI - Comparison of pain measures in surgical patients. AB - Numerous instruments have been developed for the measurement of pain with various clinical populations. This study was designed to compare pain measures for research in a sample of postoperative patients. The Brief Pain Inventory--Short Form (BPI-SF) was administered along with the Short-Form McGill Pain Questionnaire (SF-MPQ) and two visual analogue scales, one for pain while at rest (VAS-R) and one for pain upon movement (VAS-M), in random order, to 115 hospitalized patients twice following their surgery. An additional 29 patients completed the instruments once. Correlations between the visual analogue scales, BPI-SF, and SF-MPQ ranged from .33 to .76 (p < .01), suggesting that the instruments measure different aspects of pain and that instructions can influence the results. Recommendations for the selection of pain measures in patients experiencing acute pain and for future research are described. PMID- 10710859 TI - Preliminary testing of the Episode-Specific Interpretations of Exercise Inventory. AB - The purpose of this study was to develop the Episode-Specific Interpretations of Exercise Inventory (ESIE). Through a qualitative analysis of women's exercise experiences, episode-specific interpretations were defined operationally as consisting of five themes (Somatic Sensations, Affirmations, Connectedness, Explanations, and Reflections), each containing several categories. Nine-point adjectival scales were generated to represent these five qualitative themes. Older women (N = 364; mean age 69.5 +/- 6.7) completed the ESIE, the Exercise Benefits and Barriers Scale (EBBS), and three counterbalanced exercise behavior measures. Common factor analyses of the ESIE items resulted in nine factors. This measurement model was trimmed to six factors using structural equation modeling. Correlations with EBBS subscales and differences between high and low exercisers supported construct validity of the ESIE subscales. PMID- 10710860 TI - The Multidimensional Health Behavior Inventory. AB - Gaps in knowledge about what constitutes healthy and risky behaviors for young people hinder successful health promotion intervention strategies. With the development of appropriate instruments, behaviors can be measured and interventions can be implemented to improve health outcomes. The structure of a new health behavior instrument, the Multidimensional Health Behavior Inventory (MHBI), was explored with data from 1,077 college students, ages 18 to 24 years. Factor analysis of 116 health behavior questions yielded 7 factor-based scales with 57 items: diet (13 items), substance use (10 items), safety (9 items), checkup (9 items), social (6 items), stress (6 items), and exercise (4 items). Evaluation of the 7 behavior scales of the MHBI using subgroups defined by age, gender, and race will contribute to an understanding of health behaviors of older adolescents and young adults and will provide directions for research and clinical interventions. PMID- 10710861 TI - An instrument to measure the attentional demands of community-dwelling elders. AB - The capacity to direct attention (CDA) is a pivotal facet of cognitive functioning that allows people to focus on trains of thought, complex tasks, and the daily business of life. According to a theoretical framework of directed attentional fatigue (DAF) and restoration, excessive demand for attention depletes CDA, a condition called DAF. Attentional demands are factors such as feelings of loss and worries that require intense or prolonged use of CDA and thus can lead to DAF. The purposes of this study were to determine reliability and validity for an instrument to measure attentional demands, the Attentional Demands Survey (ADS). The ADS was administered to 197 (142 females, 50 males) community-dwelling elderly (ages 65-98 years, M = 77). A factor analysis revealed 4 factors/subscales consistent with theorized domains. Internal consistency for each subscale ranged from .87 to .90 and test-retest reliability was .91. The ADS can be used to explore the relationships among attentional demands, CDA, and interventions to support and restore attentional functioning for elders. PMID- 10710862 TI - Stroke epidemiological data of nine Asian countries. Asian Acute Stroke Advisory Panel (AASAP). AB - OBJECTIVE: To study the existing stroke epidemiology of nine Asian countries. METHOD: Stroke epidemiological data of nine Asian countries were collected and updated by the Asian Acute Stroke Advisory Panel (AASAP). The data included demographic information, health care resources and stroke epidemiology of individual countries. RESULT: Countries with good epidemiological data of stroke are Hong Kong, Taiwan, South Korea and Singapore. In these countries strokes rank as the second or third leading causes of death in the population. Malaysia, Thailand, Phillippines and Indonesia are countries with moderate epidemiological data of stroke. The available epidemiological data of stroke in these countries are mainly hospital-based information. Community-based studies are in the planning stage. India is the only country with fair epidemiological data of stroke available. The limitations of epidemiological study of stroke in India are due to their huge population size, poor income and limited health care resources. CONCLUSION: Combating strokes worldwide will never be truly accomplished unless basic information of developing countries is analysed and understood. The strength and weakness of information concerning strokes in developing countries need to be documented before the worldwide implementation of a stroke prevention programme. PMID- 10710863 TI - Efficacy of rapid infusion of streptokinase in patients with acute myocardial infarction. AB - The early use of thrombolytic agents is now the most important treatment in acute myocardial infarction (AMI). The earlier reperfusion should result in a higher survival rate. To determine whether the faster infusion of streptokinase (SK) will produce earlier reperfusion, 40 patients were enrolled to the trial. Half of them received 1.5 mu. of SK in one hour while the others received 1.5 mu. in half an hour. The rapid infusion group tended to have earlier reperfusion but there was no statistically significant difference in the reperfusion time. Hypotension was observed in both groups but more in the conventional group and responded to intravenous fluid replacement. Bleeding complication was low in both groups. Four patients died, one from re-occlusion and developed severe bradycardia, the remainder had cardiogenic shock which did not respond to treatment. It can be concluded that SK infusion in half an hour is safe but the beneficial effect remains to be seen in a large scale study. PMID- 10710864 TI - Abdominopelvic vascular injuries. AB - The clinical records of 25 patients with 32 abdominopelvic vascular injuries were reviewed. Sixty per cent of patients sustained blunt trauma and 40 per cent sustained penetrating trauma. Nineteen patients (76%) were in shock on arrival, 2 of them underwent ER thoracotomy when they first arrived in the emergency room. Nine patients (36%) had signs of lower extremity ischemia. The Injury Severity Score (ISS) ranged from 16-50, mean 29 +/- 10.0. Nineteen patients (76%) had 35 associated injuries. Of the 32 injured vessels; 8 were external iliac artery, 5 were renal vein, 4 were abdominal aorta, 3 were common iliac artery, common iliac vein, external iliac vein and inferior vena cava, and 1 was superior mesenteric artery, superior mesenteric vein and median sacral artery. Treatments included: 13 lateral repair, 4 prosthetic grafting, 4 nephrectomy, 3 ligation, 3 reversed saphenous vein grafting, 2 end to end anastomosis, 1 internal iliac artery grafting, 1 intravascular shunt and packing and 1 perihepatic packing. Nine patients (36%) died. High mortality was observed in injuries to the abdominal aorta (75%), inferior vena cava (66.7%), common iliac vein (66.7%) and associated major pelvic fractures (50%). Factors significantly associated with mortality were the presence of shock on arrival, associated injuries and high Injury Severity Score. The author concludes that short prehospital time, effective resuscitation and proper surgical decision making are important for survival in these critically injured patients. PMID- 10710865 TI - Personal respiratory protective devices: efficacy of intranasal stent with filters. AB - Intranasal, hollow, cylindrical, medical grade and silicone stent with two outer layers face mask filters at both ends was proposed for atmospheric suspended particulate matter prevention. The personal respiratory protective device efficacy was done at the Otolaryngology Department, Ramathibodi Hospital from April 1996 to October 1997. Single pulse mode of carbon dioxide laser smoke particle was the suitable source of atmospheric suspended particulate matter. A laser plume evacuator removed laser smoke particles with 5 Millipore filters of 0.22 um pore size or 5 intranasal stent with filters attached at the inlet end. A Millipore filter got the same laser smoke particle amount that passed through each intranasal stent filter with an air flow rate of 15 l/min controlled by a rotameter. Laser smoke particle deposition in filter materials was counted under a high power optical microscope. Laser smoke particle amount in each layer of a four-layer filter of intranasal stent with 7.5, 15.0 and 30.0 l/min air flow rates is shown. The filtration efficacy of four, three and two layers of intranasal stent with a filter for laser smoke particle retention were compared. An intranasal stent with filter could be applied in a human nasal vestibule with acceptable air flow resistant during public transportation in a traffic congested area. PMID- 10710866 TI - The urological complications of renal transplantation: an 11-year-experience at Ramathibodi Hospital. AB - From February 1986 to December 1996, renal transplantation was performed on 344 patients at Ramathibodi Hospital. The urological complications were retrospectively analyzed in 335 patients (338 renal transplants), 9 patients were lost to follow-up. There were 227 males and 108 females with age ranging from 15 to 62 years (mean age 40.28 years). There were 207 cadaveric and 131 living related graft donors. The ureteroneocystostomy was performed either by modified Politano-Leadbetter (93 cases) or extravesical technique (245 cases). There were 23 cases of urological complications: ureterovesical anastomotic leakage 6, ureteric obstruction 6, vesicoureteric reflux 4, significant bleeding from ureterovesical anastomosis 3, renal infarction with fistulas 2, hydronephrosis due to blood clot retention and swelling of the anastomosis, requiring temporary double J stenting 2. The analysis was done by dividing the patients into 3 groups, the first and second groups consisted of 100 cases each and the third group consisted of 138 cases. The urological complications in the groups were 10 per cent, 9 per cent and 2.89 per cent respectively. There was a statistically significant difference between the first two groups combined and the third group in terms of complications (p < 0.025). The urological complications of living related cases were 9 (6.87%), and of cadaveric cases were 14 (6.76%). There was no significant difference of the complications between living-related and cadaveric transplants (p < 0.05). The comparative results of the ureteric complications of the extravesical technique were significantly less than the modified Politano-Leadbetter technique (4.49% vs 10.75%), (p < 0.05). In conclusion, the extravesical technique of ureterovesical anastomosis was superior than the modified Leadbetter-Politano technique in terms of post-operative ureteral complications. PMID- 10710867 TI - Ileal interposition for the treatment of a long gap ureteral loss. AB - We retrospectively reviewed 10 patients (7 males and 3 females) who were treated with ileal interposition for long gap ureteral loss between 1989-1995. The mean patient age was 42 years old (35-52), mean ureteral gap was 18 cms (10-25). The etiology of ureteral loss included: 4 retroperitoneal fibrosis, 2 recurrent stone, 2 after pancreatitis and its complication and 2 after ureteral injury. The mean follow-up was 4 years (2-7). The post operative course was uneventful with no immediate and long term complications detected and there was no metabolic problem. Only asymptomatic bacteriuria in 5 cases (50%) was noted but it was not clinically significant. PMID- 10710868 TI - Reproductive outcome following hysteroscopic lysis of intrauterine adhesions: a result of 65 cases at Ramathibodi Hospital. AB - We reported the reproductive outcome of 65 patients with varying degrees of IUAs who underwent hysteroscopic adhesiolysis between August 1994 and December 1996 at Ramathibodi Hospital. Of the 65 patients treated, 29 had mild adhesions, 26 had moderate adhesions, and 10 had severe adhesions. Adhesions were lysed with hysteroscopic scissors in 25, with biopsy forceps through hysteroscope in 10, with electrosurgery using a monopolar probe in 22 patients, and with resectoscope in 8 patients. The mean duration of the procedure was 15 +/- 2.1 minutes. The mean follow-up was 12 +/- 1.4 months. Of the 44 patients who originally presented with secondary amenorrhea, 40 (90.9%) have normal menses, 4 (9.1%) have hypomenorrhea. Of the 6 patients who had hypomenorrhea, 5 (83.3%) have normal menses. Cyclic abdominal pain disappeared after treatment in all patients. Of the 45 patients with IUAs and infertility, 16 (35.6%) conceived. Two (20%) of the infertile patients with initially severe adhesions conceived. Of the 5 patients with RPL treated, delivered a full term baby and the other delivered a premature baby at 29 weeks of gestation. All 18 patients who delivered, had live births. Adhesion reformation was absent in patients with initially mild and moderate adhesion but occurred in 2 out of 10 (20%) patients with severe adhesions. These two patients initially suffered from secondary amenorrhea but reported hypomenorrhea after surgery. Both of them had tuberculosis of the genital tract. There were no serious complications occurring in all 65 procedures. All 65 patients were discharged a few hours after the operation. PMID- 10710869 TI - Computed X-ray densitometry measurement of mBMD & MCI in normal Thai and osteoporotic patients. AB - To establish the reference values of age-related change of metacarpal bone mineral density (mBMD) and metacarpal index (MCI) in screening for osteoporosis, both postero-anterior (PA) hands and lateral thoraco-lumbar radiography were done on 1,182 normal volunteers aged 17-83. From PA hands radiographs, mBMD and MCI were measured by computed X-ray densitometry (CXD) (Bonalyzer, Teijin Ltd., Tokyo). Exclusion of the surgical menopause condition and the causes of affected bone loss, the results show that mean mBMD and MCI in various age groups were significantly different (p-value < 0.005 for both) in females. Both values increased gradually from age under 20 and peaked in the 30-39 years age group, then decreased gradually until age 50 and decreased markedly after age 50. The yearly rate of bone loss from the peak density detected by mBMD and MCI was 1.3 per cent and 1.6 per cent between aged 50-59, 1.6 per cent and 2.7 per cent in subjects aged 60-69, 1.3 per cent and 3.2 per cent in those aged 70-79. However, mBMD and MCI in males did not show a downward trend with age. It indicated that a screening program for early prevention of osteoporosis may be necessary only in females before, during and after menopause. Because 92.3 per cent of 39 osteoporotic subjects had abnormal CXD measurements lower than -2 standard deviations (SD) limit of mean mBMD in young healthy women (aged 20-40 years), this value appeared to constitute a satisfactory definition of "high risk of developing osteoporosis". The incidence rate of high risk of developing osteoporosis was 3.03 per cent in a normal young population, while the risk rate occurred 4.76, 13.14, 34.28, 47.30 and 47.00 per cent in subjects aged 40-49, 50 59, 60-69, 70-79 and > 80, respectively. Results confirmed the necessity of early prevention of osteoporosis in postmenopausal women. These measurements may be appropriate for mass screening to separate patients who have a greater risk for development of osteoporosis from those at lesser risk. PMID- 10710870 TI - Antifungal drug combinations for Cryptococcus neoformans and Prototheca spp. AB - Seventy-one isolates of Cryptococcus neoformans and 5 isolates of Prototheca spp. were tested for in vitro susceptibility against amphotericin B alone and against the combination of amphotericin B with each clinically relevant concentration of flucytosine (5-FC) and rifampin by broth dilution methods. The combinations of amphotericin B and rifampin produced greater effect on reduction of the minimal inhibition concentration (MIC) of amphotericin B than did either drug used individually. Flucytosine combined with amphotericin B produced little or no reduction of the MIC compared with amphotericin B alone. PMID- 10710871 TI - Comparison of ofloxacin otic solution with oral amoxycillin plus chloramphenicol ear drop in treatment of chronic suppurative otitis media with acute exacerbation. AB - The efficacy and safety of 0.3 per cent Ofloxacin otic solution (OFLX) 6 drops twice daily was compared with those of oral Amoxycillin 500 mg three times daily plus 1 per cent Chloramphenicol ear drop at 3 drops three times daily (AMOX + CRP) in a two-week treatment of chronic suppurative otitis media (CSOM) with acute exacerbation. 80 adult patients were enrolled in a prospective, randomized, investigator-blind study at the outpatient ENT service of Chulalongkorn University Hospital. The most common pathogens isolated at the pretreatment visit were Staphylococcus aureus (30.3%) and Pseudomonas aeruginosa (24.7%). The susceptibility of all the pathogenic isolates to ofloxacin, amoxycillin and chloramphenicol were 96.4, 57.1 and 51.8 per cent respectively. The overall response expressed as an improvement or cure of otalgia, otorrhea and middle ear mucosal inflammation was recorded. It revealed that the improvement rate of the OFLX-treated patients was better than that of AMOX + CRP-treated, but was not statistically significant. However, the cure rate was significantly better in OFLX-treated than in AMOX + CRP-treated groups in terms of painless (p = 0.05) and dry (p < 0.001) ears. Ototoxicity was assessed by an elevation in bone conduction threshold (BC) and/or speech reception threshold (SRT) of greater than 5 dB or a presence of high tone hearing loss resulting from treatments. A significant decrease in BC and SRT was revealed in OFLX-treated ears (p < 0.0001; p = 0.002 respectively) but a significant elevation of BC was found in AMOX + CRP treated ears (p = 0.007). The ototoxic rate was significantly higher in AMOX + CRP-treated than in OFLX-treated ears whether assessed by BC (p < 0.001) or SRT (p = 0.03). In conclusion, OFLX was more effective and safer than AMOX + CRP in the treatment of CSOM with acute exacerbation. PMID- 10710872 TI - Prevalence and clinical characteristics of fragile X syndrome at child development clinic, Ramathibodi Hospital. AB - Fragile X syndrome, the most common cause of inherited mental retardation, is an X-linked genetic disorder caused by an expanded CGG repeat in the fragile X mental retardation 1 gene. It is characterized by mental retardation, behavioral features, and physical features, such as a long face with large protruding ears and macro-orchidism. A screening for the syndrome was conducted in a representative sample of pediatric patients, who had developmental delay or mental retardation with unknown cause, at the Child Development Clinic, Ramathibodi Hospital. The DNA test was performed on all patients using PCR and southern blot techniques. Five positive cases were detected from 114 screened subjects, and more four cases confirmed among other family members. Two of five positive families initially denied a family history of mental retardation. Among 9 cases of fragile X syndrome, four had hyperactivity and two had autistic like behavior. More than half had rather a long face or prominent ears. Three boys had macro-orchidism. PMID- 10710873 TI - Microsurgical toe to thumb transplantation for traumatic thumb loss. AB - Toe-to-thumb transfer was performed on 13 patients with traumatic thumb loss, using microvascular technique. Of those, 9 patients had industrial injury, 3 patients had a sharp cut injury and the remaining one had animal bite injury. The toes used for the transfer were the great toe(5), and the second toe(8). There were twelve successes, and one partial success. Ten patients have been followed up for more than 12 months, and they are reviewed in detail. Total active motion after great toe and second toe transfer was 45 degrees and 68 degrees respectively. Static two-point discrimination was 10-15 mm in 10 patients. The donor foot did not suffer functionally. All of the patients returned to gainful employment postoperatively. PMID- 10710875 TI - Multiple cranial neuropathy in Cogan's syndrome. AB - A 20 year old woman presented with recurrent alternative keratitis for four months. One month before admission, she developed progressive hearing loss, visual impairment, facial diparesis and bilateral trigeminal neuropathy. Cogan's syndrome was diagnosed. Prompt treatment with corticosteroid resulted in dramatic improvement of the ocular, otological and neurological dysfunctions. PMID- 10710874 TI - Phase II study of concurrent chemoradiotherapy for inoperable (bulky) stage III (A/B) non-small cell lung cancer (NSCLC): a preliminary report. AB - We designed a phase II study to determine the feasibility and toxicity of concomitant radiotherapy and Paclitaxel/Carboplatin followed by adjuvant chemotherapy of the same regimen in patients with newly diagnosed inoperable stage III A/B non-small cell lung cancer. Patients were irradiated with a total dose of 66 Gy. Weekly courses of Paclitaxel 45 mg/m2 and Carboplatin AUC 2 were administered intravenously during the irradiation period. After completion of concurrent chemoradiotherapy, adjuvant chemotherapy with Paclitaxel 175 mg/m2 and Carboplatin AUC 6 intravenously every 3 weeks for 4 cycles were given. Since March 1998, 15 patients have been enrolled. All patients were assessable for efficacy and toxicity after concurrent chemoradiotherapy. Eleven patients were assessable for efficacy and toxicity after adjuvant chemotherapy. After concomitant chemoradiotherapy, complete response (CR) was documented in 2 of 15 (13%). Partial response (PR) was documented in 9 of 15 (60%). After completion of adjuvant chemotherapy in 11 patients, the overall response rate was 91 per cent. (18% CR, 73% PR). There were 8 per cent gr. 3-4 neutropenia which occurred during adjuvant chemotherapy. Concomitant Paclitaxel/Carboplatin and radiotherapy are promising modalities in the treatment of inoperable stage III A/B non-small cell lung cancer. PMID- 10710876 TI - Malignant carcinoid tumor of the appendix with liver and lung metastasis: report of a case with a high level of serum carcinoembryonic antigen. AB - We report elevated serum carcinoembryonic antigen (CEA) in a case of malignant carcinoid tumor of the appendix with liver and lung metastasis. A 55-year-old Thai man was found to have multiple nodules in the liver by ultrasonography. Serum CEA was 7,387.9 ng/mL (normal 0-4.1 ng/mL) leading to a clinical impression of colonic carcinoma with liver metastasis. During the investigation, he developed acute abdomen caused by ruptured acute appendicitis. Malignant carcinoid tumor of the appendix, 1 cm in diameter and located proximal to the ruptured acute appendicitis, was identified. The tumor cells showed trabecular or insular growth pattern, some nuclear pleomorphism but typically fine nuclear chromatin, frequent mitoses and focal necrosis. They were immunoreactive for antibody to chromogranin, neuron-specific enolase, CEA, and cytokeratin. Tumor metastases were discovered in the liver, right lung, mediastinal and right supraclavicular lymph nodes. Electron microscopic study demonstrated pleomorphic neurosecretory granules of the midgut type of carcinoid tumor. PMID- 10710877 TI - Medical procedure training, is it the problem? PMID- 10710878 TI - Ophthalmology in Thailand 1997. AB - This article is a brief history of ophthalmology in Thailand after World War II and the National Programmes for the Prevention of Blindness in the past two decades from the late 1970's. Presentation illustrates the efforts of the eye sector in Thailand not only modernizing ophthalmology. However, eye care services in public places has also been made available to all assured with easy accessibility and acceptable quality. The National Blindness Prevention programmes have been forwarded through adoption of primary eye care approach integrating into the national health scheme of primary health care. Avoidable blindness once prevalent in 1980's, has been reduced from 1.1 per cent to 0.3 per cent in twenty years. Eye care networks that came into existence in that period covering the whole country are now once again under realignment for new challenges from unavoidable types of blindness. PMID- 10710879 TI - Personal respiratory protective devices: efficacy of Millipore and Whatman filters. AB - A Millipore filter with 0.22 micron pore size and a Whatman grade 1 filter with > 11 microns particle retention were used to capture laser smoke particle mimic atmospheric suspended particulate matter. The experiment was conducted at the Department of Otolaryngology in Ramathibodi Hospital from April 1996 to October 1997. The laser smoke particle evacuator with rotameter created an air flow rate of 15 l/min through the filters. The mean and standard deviation of the laser smoke particle count under high power optical microscope in a 10 Millipore filter and a 10 Whatman filter were 411,327.6 +/- 13,325.0 and 290,453.0 +/- 28,409.8 respectively, 29.4 per cent different. Laser smoke particle size distribution in both filters under eyepiece micrometer was: 1 to 10 microns in Millipore (99.0%) and in Whatman (96.2%), 1 to 5 microns in Millipore (77.1%) and in Whatman (77.6%), no laser smoke particle larger than 17 microns was detected. The Millipore filter ruptured when the air flow rate was greater than 15 l/min. The Whatman filter was suitable for evaluating filtration efficacy of various personal respiratory protective devices in a high air flow rate condition. PMID- 10710880 TI - Recombinant interferon-alpha 2b and megestrol acetate in patients with advanced renal cell carcinoma. AB - Fifteen patients with histologically proven metastatic or unresectable renal cell carcinoma were enrolled for a phase II trial of Interferon-alpha 2b (> or = 6 x 10(6) U/m2) plus megestrol acetate (160 mg/day). A response rate of 14.3 per cent was achieved in this study. We observed weight gain (median 3.2 kilogram; range 1.1 to 6.9) in 5 patients, and stable weight in 5 of the 14 patients who completed the protocol. Weight gain occurred regardless of extent of metastasis or response to interferon. Our data suggest a possible role for megestrol acetate in alleviating anorexia and weight loss in patients with renal cell carcinoma undergoing interferon treatment. Further clinical trials are clearly warranted. PMID- 10710881 TI - The use of historical and anthropometric data as risk factors for screening of low mBMD & MCI. AB - To evaluate the risk factors which affect bone loss in screening for osteoporosis, interview of anamnestic data (age, marriage status, pregnancies, menopausal age, intake of calcium, vegetables, protein and coffee, excessive use of alcohol and smoking, sedentary habits, family history), medical data, surgical data, followed by measurement of anthropometric variables [weight, height, antero posterior (AP) thickness at xiphoid level], blood examination (calcium, inorganic phosphorus, alkaline phosphatase), both postero-anterior (PA) hands and lateral thoraco-lumbar radiography were done in 1,182 normal volunteers aged 17-83. From PA hands radiographs, metacarpal bone mineral density (mBMD) and metacarpal index (MCI) were measured by computed X-ray densitometry (CXD) (Bonalyzer, Teijin Ltd., Tokyo). The results showed that the mean of menopausal age in Thai females was 48.86 +/- 3.09 years ranging from 39 to 55 years. The average number of children in their family was 2.10. Correlation among anthropometric variables, AP thickness was positive linear correlation to weight/height ratio (r = 0.7878, p value < 0.005). Weight, AP thickness and body mass index (BMI) significantly increased with aging (r = 0.2456, 0.4489 and 0.3484, p-value < 0.005, 0.001 and < 0.005), but decreased with height (r = -0.1030, p-value = 0.001). Lower mBMD and MCI were associated with increased age, married female, increased pregnancies, increased AP thickness, decreased vegetable intake, increased protein intake and increased years after menopause. From a multiple regression analysis, the significant factors that can predict the MCI were years after menopause, sex, daily vegetable intake and hormonal replacement. The incidence rate of high risk of developing osteoporosis in females, no vegetable intake and no hormonal replacement subjects occurred 7.50, 2.22 and 2.63 times greater than in males, vegetable intake and hormonal replacement subjects, respectively. In postmenopausal women since 1-2, 3-5, 6-10, 11-15 and > 15 years, the incidence rate were 5.24, 14.51, 17.01, 20.86 and 29.76 times greater than the rate of premenopausal women. Concerning perimenopausal women, only 2 of all factors influenced the measured mBMD and MCI. The incidence rate of high risk of developing osteoporosis in women who intake protein > 30 g/d and intake medicine (corticosteroid) was 2.96 and 6.16 times greater than < 30 g/d protein intake and no medicine intake subjects. PMID- 10710882 TI - An identical neo-mutation in the thyroid hormone receptor beta gene (A317T) in 2 unrelated Thai families with resistance to thyroid hormone. AB - We reported two unrelated Thai girls with resistance to thyroid hormone. The affected patients presented with goiter and no other stigmata of hyperthyroidism. Their serum T4, T3, free T4 and free T3 concentrations were high and they had normal levels of TSH. The affected girl in family 1 was treated with an antithyroid drug for 1-9/12 years. The affected girl in family 2 was only observed her thyroid function tests. TRH test showed normal TSH response in both girls. Analysis of the thyroid hormone receptor beta gene of both affected girls revealed the same missense mutation, changing the guanine in nucleotide 1234 to an adenine which results in the replacement of the normal alanine (GCT) with a threonine (ACT) at codon 317. Two proposita were heterozygous, and this mutation was not present in their parents compatible with a neo-mutation. PMID- 10710883 TI - Treatment of adolescent idiopathic scoliosis using Cotrel-Dubousset spinal instrumentation. AB - Eighteen patients with idiopathic scoliosis who underwent posterior spinal correction and fusion using Cotrel-Dubousset instrumentation between 1991 and 1996, were evaluated for curve correction and complications. Age at surgery averaged 14.7 years. Follow-up averaged 3.7 years. Thoracic curve correction averaged 65 per cent in those with King type III/IV curves and 51 per cent in those with King type II curves. At the recent follow-up, correction loss averaged 12 per cent and 8 per cent, respectively. Lumbar curve correction averaged 31 per cent after instrumentation in type II curves, with a loss of approximately 3 per cent correction at follow-up. Thoracic sagittal contour improved 14 degrees for hypokyphotic patients. Apical vertebral rotation improved an average of 37 per cent after derotation maneuver of the left side rod. No neurologic complications or deep infection occurred. In conclusion, frontal and sagittal thoracic curve correction can be satisfactorily achieved using Cotrel-Dubousset instrumentation. PMID- 10710884 TI - Primary gastrostomy button: a means of long-term enteral feeding in children. AB - Between June 1992 and December 1997, forty-two patients (M 19, F 23) received 94 primary gastrostomy buttons due to 22 intellectual handicap, 7 cystic fibrosis, 4 severe gastrooesophageal reflux, 2 bronchopulmonary dysplasia, 2 tumours in the neck region and 5 miscellaneous causes. Open fundoplication concomitant with primary button, primary open button and laparoscopic fundoplication concomitant with primary button were performed in 20, 15 and 7 patients respectively. The average longevity +/- standard deviation of all buttons was 388.36 +/- 360.35 days. The average longevity of the buttons of the laparoscopic fundoplication group was significantly lower than the others. The major causes of removal of Bard buttons were valve incompetence and flap damage, whereas, balloon leakage was the major cause of removal of the Mic-key button. There were merely minor stomal complications and no gastric separation and peritonitis. Because of the acceptable longevity of the buttons and minimal complications, we concluded that the primary gastrostomy button was the preferable method of long term enteral feeding in children. PMID- 10710885 TI - Intravenous gamma globulin for treatment of chronic idiopathic thrombocytopenic purpura in children. AB - A prospective and descriptive study was carried out in 17 children with chronic ITP. Five-day course of Intraglobin (400 mg/kg/d x 5) was given intravenously to 10 children with the age of 4-16 years (5 males and 5 females). Two-day course of Venoglobulin-I (1 g/kg/d x 2) was given intravenously to 7 children with the age of 3-15 years (3 males and 4 females). Intraglobin and Venoglobulin-I were effective in treating children with chronic ITP. All of the patients had transient increased in their platelet counts during the first 2 weeks. The two day course of Venoglobulin-I was superior to the five-day course of Intraglobin. Mild adverse effects were observed in a greater percentage of patients treated by Venoglobulin-I than in patients treated by Intraglobin. Intravenous immunoglobulin was one of the choices of treatment in children with chronic ITP, but the cost of immunoglobulin or gamma globulin is quiet high. PMID- 10710886 TI - Modified single step ultrasound guided percutaneous transhepatic biliary drainage (MSSUGPTBD): experience with 102 patients. AB - Modified single step ultrasound guided percutaneous transheptic biliary drainage was performed in 102 patients between 1993 and 1998 at the National Cancer Institute, Bangkok with successfully placed drainage tubes in the dilated bile ducts. The advantages of this technique are single step puncture without major complication or bleeding, reduction of radiation exposure, capability for bile duct selection and time saving. PMID- 10710887 TI - Effect of national seminar on AIDS and anesthesia upon knowledge, attitude and practice concerning HIV among Thai anesthesia personnel. AB - In the national seminar of AIDS and Anesthesia which was a short course educational program in all aspects of HIV medicine, 195 questionnaires about knowledge, attitude and practice concerning HIV were distributed among the participants (anesthesiologists and nurse anesthetists) in 3 periods, pretest, post test (at the end of 2 days seminar) and post test 2 (at 4 months after the seminar). There were 177 (90.76%) respondents who completed both pretest and post test 1 questionnaires. About 12 questions of knowledge; mean scores were statistically significantly increased; 7.95 (0.98) vs 9.5 (0.78), P < 0.001. Two thirds (8 out of 12 questions) were answered correctly in post test 1 more than in the pretest by Mc Nemar Chi-square test; P < 0.05. About attitude; 2 out of 5 answers changed significantly by Mc Nemar Chi-square test; P < 0.05. The post test 2 questionnaires were mailed to all 177 participants twice asking to reply only once. All questionnaires were to be completed anonymously. The post test 2 with a response rate of 65.5 per cent revealed that universal precautions were frequently used among Thai anesthesia personnel but not universally followed. At least one-third of the respondents admitted recapping before disposal of used needles. Fifty six per cent of respondents (vs 22.8% in pretest) admitted re using one syringe for more than one patient. In conclusion, this study showed that a short course educational program may improve knowledge about HIV and partly change attitude, but can not change behaviour. Changing the practice of anesthesia health care workers needs continual education and appropriate training. PMID- 10710888 TI - Excimer laser photorefractive keratectomy and laser in situ keratomileusis for myopia and astigmatism. AB - The efficacy, predictability, safety, and short-term stability of excimer laser photorefractive keratectomy (PRK) and laser in situ keratomileusis (LASIK) for treatment of myopia and astigmatism were determined. The preoperative myopia ranged from -1.50 to -15.75 D and the astigmatism was less than 4.0 D. Of the 147 eyes, 73 and 74 underwent PRK and LASIK, respectively. Mean preoperative spherical equivalent refraction (SE) was -3.72 +/- 1.69 D in the PRK group and 7.66 +/- 2.30 D in the LASIK group. Mean postoperative SE at the last examination (3 to 6 months) was -0.13 +/- 0.82 D and -0.38 +/- 1.19 D in the PRK and LASIK groups, respectively. Eighty six percent in the PRK group and 77 per cent in the LASIK group achieved a SE within +/- 1.0 D and the refractions were stable between 1 month and 3-6 months. Uncorrected visual acuity of 20/40 or better was noted in 91 per cent in the PRK group and 97 per cent in the LASIK group. No eyes lost one or more lines of best spectacle-corrected visual acuity in both groups. PRK and LASIK appear to be effective, safe, predictable, and short-term stable in treating myopia and astigmatism. Longer follow-up studies will help evaluate the long-term stability of the procedure and possibility of later complications. PMID- 10710889 TI - Fine needle aspiration cytology of metastatic transitional cell carcinoma to the liver. AB - A 63-year-old man presented with a three-month history of painless hematuria. A cystoscopic examination revealed a diffuse small nodulopapillary growth of the bladder mucosa. Biopsy resulted in the diagnosis of a transitional cell carcinoma (TCC), grade II. Therefore, total cystectomy with an ileal conduit was performed and the pathologic examination demonstrated a TCC grade II/III apparently confined to the mucosa. However, an ultrasonographic study carried out one year later revealed tumor masses in the pelvic cavity and the liver. FNA and needle biopsy of the liver were carried out and the diagnosis of a metastatic TCC was made from the former. Needle biopsy results pointed to a metastatic undifferentiated carcinoma, most likely originating from the TCC. The advantage of FNA is discussed. It is being used with increasing frequency to diagnose mass lesions in the liver and can identify metastatic tumors which have specific cytologic features that are different from primary liver tumor. PMID- 10710890 TI - Primary mucoepidermoid carcinoma of the intrahepatic bile duct: a case report with review of literature. AB - A 64-year-old Thai man had a primary mucoepidermoid carcinoma, a variant of the cholangiocarcinoma, of the left lobe of the liver. A 5 x 4 x 3 cm tumor metastasized within the liver and to the porta hepatic and parapancreatic lymph nodes with compression of the head of pancreas and distal part of the common bile duct resulting in obstructive janudice. The patient died within 7 months from massive hemorrhage of a chronic peptic ulcer of the duodenum. Review of the medical literature in the English language disclosed 11 primary mucoepidermoid carcinomas of the bile ducts, including the present instance. Nine tumors were intrahepatic; three were in the right lobe and six in the left lobe. Two neoplasms were extrahepatic; they arose in the common hepatic ducts. There were 6 men and 5 women of 44 to 78 years old; the mean age was 60 years. The size of the tumor ranged from 1.5 to 18 cm in greatest dimension; the average size was 8.4 cm. The tumor metastasized frequently to the regional lymph nodes. Invasion of the portal vein and hepatic artery has also occurred. Ten patients having primary mucoepidermoid carcinoma of the bile duct died within 11 months regardless of treatments. Only one patient was well, 10 months after extensive resection of a 1.5 cm tumor of the common hepatic duct. It is concluded that the prognosis is generally poor for the patient having primary mucoepidermoid carcinoma of the bile duct. PMID- 10710891 TI - Colonic carcinoma: a case report in a child and review literature. AB - This article reports one case of child colonic carcinoma. This is a rare disease in children which usually occurs in predisposing conditions, e.g. ulcerative colitis, familial polyposis coli, Gardner's syndrome, Turcot's syndrome and Peutz Jegher's syndrome. The patient in this report was 12 years old. He presented with chronic intermittent colicky abdominal pain and uncorrectable iron deficiency anemia for 7 months prior to definite diagnosis. This report also reviews the literature about colorectal carcinoma in children. Physicians can make an early diagnosis with a high index of suspicion if they cannot explain clearly what causes abdominal pain. Further investigations should be performed, thereby, avoiding delayed diagnosis and improving survival rate. PMID- 10710892 TI - Traumatic testicular dislocation a review of 36 cases. AB - We retrospectively reviewed 36 patients who were treated in our institutes with traumatic testicular dislocation from 1975 to 1997. The mean patient age was 25 years old (18-38). Average time to present at the emergency room was 1 hour (0.5 6). Bilateral dislocation was found in thirty cases and unilateral dislocation was found in six cases. The sites of dislocation included: 34 cases (64 testes) at superficial inguinal area, one case (one testis) at acetabular area, and one case (one testis) at the perineal area. Closed reduction under general anesthesia was successful in 14 cases, open reduction after failed closed reduction in 10 cases, open exploration and repaired testis with reposition in 11 cases and orchiectomy only in one case. The overall results after treatment showed the normal size and position of the testis. PMID- 10710893 TI - Disclosure of author contributions in scientific publications. PMID- 10710894 TI - The impact of zidovudine use in HIV-infected pregnant women on vertical transmission of HIV in Mississippi. AB - In February 1994, results of a large placebo-controlled trial of zidovudine (ZDV) use during pregnancy (ACTG 076) showed a dramatic reduction in vertical transmission of HIV. In August 1994, the Public Health Service (PHS) recommended routine ZDV use in HIV infected pregnant women and their neonates for the prevention of vertical transmission. We retrospectively reviewed vertical transmission rates of HIV in Mississippi from 1/1/90 to 8/30/94 before the PHS guidelines were released and from 9/1/94 to 12/31/97 after the PHS guidelines were released. We also reviewed data on ZDV use in HIV infected pregnant women and their neonates from 9/1/94 to 12/31/97. Antenatal, intrapartum and neonatal ZDV use increased from 61%, 59% and 73% respectively to 79%, 77% and 92% respectively. After 9/1/94, vertical transmission rates fell by 44%. Zidovudine use during pregnancy has increased in Mississippi since release of the PHS guidelines resulting in a dramatic decline in vertical transmission rates. PMID- 10710895 TI - Unrestricted riding in pickup truck cargo beds poses significant hazards. AB - Individuals riding in the beds of pickup trucks face significant risks of debilitating injury or death, yet Mississippi currently has no legislation restricting ridership in truck beds. Data collection on accidents involving truck bed passengers indicates that children make up the majority of victims. Such accidents impose a heavy burden on society in terms of both medical expenses and impaired quality of life for the victims. PMID- 10710896 TI - Back to the AMA. PMID- 10710897 TI - "To err is human"--or is it process? PMID- 10710898 TI - Prostate cancer: an overview. PMID- 10710899 TI - Detection and diagnosis of prostate cancer. PMID- 10710900 TI - Management of localized prostate cancer. PMID- 10710901 TI - Surgical management of localized prostate cancer. PMID- 10710902 TI - Radiotherapy in prostate cancer: improvements in an effective treatment and future prospects of further gains. AB - Radiation therapy has useful applications in the management of all stages of prostate cancer. Modern advances have improved the efficacy of radiotherapy, and have lowered the toxicity. Radiotherapy offers patients with early stage, clinically localized disease a non-invasive curative option that has low toxicity. Patients with metastatic prostate cancer can be palliated with judicious use of radiotherapy. PMID- 10710903 TI - Trans rectal ultrasound guided radioactive seed implant: the Sumter experience. AB - 108 patients were treated between 1/06/97 and 7/19/99. Ages ranged from 53 years to 82 years. Pretreatment PSA ranged from 4.0 to 34.6 and post treatment PSA from 0 to 7.3. Given the fact that carcinoma of the prostate is a relatively slow growing tumor and that long survival times are not uncommon even in the face of relatively advanced disease, our treatment team believes that the preferred method of treatment in otherwise healthy patients who have a 20 year life expectancy is radical surgery. We do believe that brachytherapy has a role in the treatment of organ-confined disease and may possibly be proven to be as effective as surgery; however, until there is at least 15-year data, it cannot be considered as the primary treatment of choice in young men. The senior author has stressed the term "treatment team" in this paper for a reason. We sincerely believe that the best results for our patients are obtained when we approach the treatment of the patient as a unified team that makes use of our individual talents and training for the good of the patient as a whole. Each member of the team performs the procedures that he is best trained to do, and we are consistent in our approach and advice to our patients regardless of our disciplines. PMID- 10710904 TI - Cryosurgical ablation of the prostate in South Carolina. PMID- 10710905 TI - Cryosurgical ablation of the prostate in South Carolina. PMID- 10710907 TI - Prostate cancer and nutrition. PMID- 10710906 TI - Medical therapy of prostate cancer: 1999. AB - Prostate cancer is the most commonly diagnosed male malignancy and the second most common cause of male cancer death in the U.S. The principles of management of newly diagnosed metastatic prostate cancer have changed little in the last 50 years. Medical therapy continues to have no standard role in the management of localized disease. The various hormonal monotherapy approaches targeting androgen deprivation yield comparable results in the treatment of advanced disease. Supplementing an LHRH agonist (but not orchiectomy) with an antiandrogen may improve survival in men with minimal disease, but the economic cost and the risk for significant impairment of quality of life are quite high. The benefit of combined androgen blockade for patients with more extensive disease remains unclear, with support for this approach waning in recent years. New chemotherapeutic agents and combinations of such, as well as agents with entirely new mechanisms of action, recently have shown encouraging results in the treatment of HRPC. Much additional research is needed to improve our armamentarium against this epidemic disease. PMID- 10710908 TI - Special issue--prostate cancer. PMID- 10710909 TI - Counseling the patient with prostate cancer. PMID- 10710910 TI - [Surgical treatment of carotid chemodectomas]. AB - 49 patients have been examined and 38 of them operated in RRCS RAMS for carotid hemadectomas (CH). CH is a rather rare tumor developing from the carotid gland locating in bifurcation of the common carotid artery. It is slowly growing tumor which is scantily active and rarely metastasizes. In 35 of the 38 operated patients radical ablation of the tumor was carried out. In the rest 3 patients radical removal of the tumor was impossible due to its dissemination and malignancy. The classification was used according to which all the tumors were distributed in 3 groups in dependence on the character of their growth. The basic principle of their division was the relation of CH to the carotid arteries. The use of this classification allowed to define individual surgical tactics. As a result of the treatment 97.4% of the patients were discharged with improvement. During the follow-up period (94 +/- 5 months) there were no relapses detected in these patients. PMID- 10710911 TI - [Long-term result of hemicorporectomy]. AB - A 12-year follow up result of hemicorporectomy in a patient is represented. The patient was operated several times since 1983 for massive perianal condylomas. In 1985 the ulcer with hard edges was revealed in the perianal region, spreading to the perineum and root of the scrotum. Biopsy data evidenced for epidermoid carcinoma Abdominoperineal extirpation of the rectum was carried out with broad dissection of the skin of the perineum and with resection of the seminal follicle. Postoperative period was complicated by prolonged pyogenous infection of the perineal wound which prevented from radiation treatment. 9 months later the relapse of the tumor was detected in the perineum with deep pyogenic fistulas formation. 6 courses of chemotherapy by 5-fluorouracyl were carried out. During the process of examination in September 1987 in the perineal area a large massive tumor occupying the whole pelvic cavity and growing into the posterior wall of the urine bladder and left ishial bone, spreading to the scrotal root and surrounded by the net of fistulous tracts was revealed. Hemicorporectomy was carried out with previously layed one-stem sygmostomy keeping intact, and retroperitoneal Y-shaped uretero-ureter anastomosis being formed and right ureter being fixed at the skin of the right abdominal wall. A special prosthesis--"a glass"--was made for the patient, in which he could move from the bed to the chair or the wheeled chair, to move at home or in the street and to drive his own car. Later, evacuation of the uroliths through the uretherocutaneous stoma was observed. Gradually urolithiasis progressed, mainly in the right kidney, and in 1995 the development of purulent rightsided paranephritis was detected which demanded right-sided nephrectomy. Thus, in spite of a number of complications we can state, that hemicorporectomy has cured the patient of advanced, cancer and he feels satisfied with this treatment and saving 12 years of life. PMID- 10710912 TI - [Subtotal colectomy as surgical treatment of acute colonic ileus obturation of tumor genesis]. AB - The experience of subtotal colectomy in 20 patients as a method of treatment of left-sided obturation colonic ileus has been summarised. In 18 patients the operation was performed as an urgent procedure and in 2 patients it was less urgent. In 4 patients urgent subtotal colectomy was combined with resection of the other organs due to local spread of the tumor. In 4 patients complications developed in the early postoperative period, in 2 patients they were followed by urgent operation. There were 2 lethal outcomes due to decompensation of associated chronic cardio-vascular and pulmonary diseases. The authors suggest that subtotal colectomy in treatment of acute neoplastic colorectal obstruction could be used by surgeons of nonspecialized departments of urgent surgery provided that the competence of the surgeons is high enough and preoperative treatment is adequate. PMID- 10710913 TI - [Estimation of stomach surgery results]. AB - Operations on the stomach make up about 2-5% of all operations performed in clinics of general surgery. In evaluation of the efficacy and quality of treatment the quantitative indices are used: intraoperative and postoperative complications, frequency of relapses of the basic disease, frequency and degree of severity of developing functional manifestations of the diseases of postoperative stomach and metabolic disturbances. However, contemporary trends to standardization of surgical technique and to high rank of technological equipment of the operation unit predetermine related or identical results after various operations on the stomach. Differences in outcomes of surgical treatment are best of all revealed in the study of qualitative results. The qualitative evaluation of various operative procedures with the use of integrale scales of Johnston and Visick has some substantial disadvantages. WHO recommends to use "adequacy of treatment", i.e. achievement of the adequate quality of life for each patient as a criteria for evaluation of the efficacy of treatment. The authors suggest that, until the influence of particular operation on the level of the quality of life of patients in postoperative period (in comparison with the initial ones before the operation and in healthy people) is not studied, it is impossible to judge about its advantages and shortcomings. The quality of surgical operation--the choice of the procedure and its technical realisation--predetermines quality of life of patients in postoperative period. PMID- 10710914 TI - [Plastic surgery in patients with obesity late after horizontal banded gastroplasty]. AB - The results of 361 plastic operations in 296 patients with morbid obesity late after horizontal gastroplasty were analyzed. Plastic and corrective operations aimed at removal of redundant lipocutaneous "aprons" at the anterior abdominal wall, thighs, thoracic wall, gluteal region and the arms, represent a final stage of surgical treatment of patients with morbid obesity. The indications, technique and the results of plastic operations performed from 1985 to 1998, are thoroughly elucidated. The analysis of early postoperative complications has established, that it a reasonable to perform such operations 1-3 years after gastroplasty when body weight stabilizes and there are no vitamin deficiency, iron deficient anemia, hypoproteinemia, hydroionic disturbances or other complications of the later period. Complex prophylactic measures for prevention of pyoseptic and thromboembolic complications in patients with obesity late after gastroplasty permits to avoid severe complications and lethal outcomes in patients after plastic operations. PMID- 10710915 TI - [Laparoscopic and standard cholecystectomy: comparison of immediate results]. AB - The advantages of laparoscopic cholecystectomy (LChE) are undoubtable in comparison with traditional one. However the experience showed, that LChE is not devoid of some shortcomings. It is characterized by the same typical complications as in ChE, besides it may be followed by some specific complications. The literature concerning the rate of complications in LChE are controversial. The comparative analysis has been carried out concerning the rate of complications after ChE (6800 operations) and LchE (900). The mean age of patients with choledocholythiasis--61.4 years. 40% of the patients had severe accompanying diseases coronary artery disease, complicated forms of arrythmia, arterial hypertension, diabetes mellitus, obesity. There were no significant differences between patients who underwent ChE (group 1) and LChE (group 2) by their age and the rate of accompanying diseases. Intraoperative cholangiography was performed in 3.5% of cases of group 1 and in 1.1%--in group 2. The average rate of the operations on extrahepatic bile ducts in group 1 was also lower- choledocholithotomy was carried out in 2.7%, transduodenal papillosphyncterotomy- in 1.7% of cases, drainage of the choledochal duct--in 1.9%. Combined operations were carried out in 10% of patients of group 1 and in 9.1% cases of group 2. The rate of intraoperative bleedings which demanded repeated operations made up in patients of group 1 0.1%, in patients of group 2 0.5%. Intraoperative damage of the choledochal duct in group 1 were detected in 0.14% and in group 2--in 0.11% of cases (the only complication of LChE in a patient was assessed as a endogenous wall clipping). PMID- 10710916 TI - [Intraoperative ultrasonography in surgery on organs of abdominal cavity and retroperitoneal space]. AB - Since 1984 to 1999 intraoperative US examination (IOUSE) was made in 863 patients. 236 of them had diseases of the liver, 137 diseases of the gall bladder and biliary tract, 158 pancreatic disorders, 151 diseases of gastro-intestinal tract, 72 diseases of the adrenal glands, 101 constitutional obesity, 3 splenic disorders and 5 gunshot wounds of the organs of the abdominal cavity and the retroperitoneal soft tissues. In masses of the liver IOUSE showed limits for resection of the liver, location of the diseased area with the vessels and biliary ducts, unpalpablefoci. OUSE allowed to perform US-guided puncture and sclerosizing small metastatic nodes as well as a puncture and antiparasitic treatment of inaccessible echinococcal cysts. In cholelithiasis IOUSE allows to reveal or to rule out the presence of the concrements in bile ducts. In cases of diseases of the pancreas IOUSE helps to localize pathologic area, defines its interrelations with adjacent major vessels and ducts, reveals unpalpable masses and enables US-guided puncture of the major pancreatic duct followed by its further opening. In gastric tumors it is possible under US control to determine precisely the limits of the infiltration of gastric wall which defines the volume for resection. In operation on the adrenals from thoracophrenolumbar approach IOUSE through the diaphragmatic cupula determines the place and the length of phrenotomy and elucidates interrelations of the tumor with adjacent organs. Sensibility of IOUSE made up from 87 to 100%, and specificity--92-100% depending on the disease. PMID- 10710917 TI - [Mode of surgery completion in peritonitis]. AB - 1310 patients with various forms of peritonitis were operated during 1989-1998. The tactics of the treatment was determined depending on bacterial contamination of the abdominal cavity. In abscesses of the abdominal cavity with massive bacterial contamination (6-7 CFU/g) drainage procedure was used. Mortality rate made up 4.8%. In local extended and diffuse peritonitis with a slight bacterial contamination of the abdominal cavity (3-5 CFU/g) and in absence of fibrinous deposition fixed on peritoneum, the drainage of the abdominal cavity was not used, and laparoscopy was performed in postoperative period for the control of the course of infectious process. Mortality rate was 0.6%. In extended peritonitis with massive bacterial contamination (6-8 CFU/g) the method of repeated explorations and sanitations of the abdominal cavity was used, mortality rate being 17.8%. The overall lethality made up 7.8%. Postoperative wound infection occurred in 6.7%, intraabdominal infection as abscesses or progressing peritonitis--in 2.1% of cases. PMID- 10710918 TI - [Endobronchial laser therapy in complex preoperative preparation of patients with lung diseases]. AB - The method of endobronchial laser therapy with the use of photosensitiser photosense (phthalocyanine aluminum)--was used since 1998 for the first time in faculty surgical clinic, as preoperative preparation in 36 patients with surgical diseases of the lungs (malignant and benign tumors and chronic inflammatory diseases). The method aimed at elimination of postoperative pyogenous complications and improvement of the respiratory system functional. The method consists in introduction into the organism by various ways the photosensitizing preparation photosence, followed by irradiation of the right and left parts of bronchial tree by low intensity laser light, (wave length 675 mm) through dispersing light guide, introduced through the flexible bronchoscope to the areas of the ostia of the lobar bronchi. 3 ways of introduction of the photosensitizer were used: endobronchial (9 patients); aerosol (22 patients); combination of the endobronchial lavage and intravenous injection (5 patients). The duration of the procedure of irradiation made up 5 min. at each side of the bronchial tree, the power density being 85 mV/cm2. The procedure was repeated twice for 2 days. The following results were obtained: antibacterial effect; reduction of endoscopic and morphological features of inflammation of the bronchial mucosa; stimulation of local immunity; decrease of the tumor aggression (according to histological examination); the appearance of the tumor destruction areas; lavering of Ki-67 and bcl-2, increase of c-bax; the improvement of the respiratory system functional; positive dynamics of clinical status (a decrease of cough, dispnea, quantity of mucus discharge and haemoptysis). The perspectiveness of the method is stressed as a preoperative preparation in patients with surgical diseases of the lungs. PMID- 10710919 TI - [Application of ultrasound in differential diagnosis of pleurisy]. AB - According to literature data, pleural exudate is diagnosed approximately in 10% of pulmological patients. The importance of differential diagnosis of pleuriesy is expained by variability of the diseases accompanied by pleural exudate. 146 patients with various diseases accompanied by pleural exudate were examined: 39 from them had pleural empyema, 53--diffuse pleurisy, 42--incapsulated pleurisy, and 12--tumors of the pleura. All the patients underwent ultrasound examination (USE) of the chest together with a routine roentgenography, as well as computed tomography. USE was performed on the. Toshiba and Hitachi egurpment, which work in real time regime with sectorial sensing element (3.5-5.0 mHz). The obtained data show, that USE is most effective diagnostic tool in incapsulated and diffuse liquor accumulations in pleural cavity. In 6 patients the diagnosis of pleuritis was possible only at USE of the pleural cavity. Evaluation of the structure of the liquor and detection of tumor conglomerates on parietal pleura have a substantial diagnostic value. It is noted that echotomographic picture might be insufficiently informative in some situations with a small length of the contact of the lesion with the thoracic wall (less than 3 cm) or thickness of pleural layers up to 4-5 mm, in the presence of artefacts due to pulmonary tissues and osteal structures of the thoracic wall (ribs, scapula, sternum). PMID- 10710920 TI - [Postoperative thromboembolism of pulmonary artery]. AB - Pathologo-morphological and clinical records of 803 surgical patients have been analysed, in whom the thromboembolism of pulmonary artery (TEPA) was the only or the main cause of the death. Intravital diagnosis of TEPA was established in 32% of all the deceased. 87% of patients with evident symptoms of the embolism died during the first 2 hours, 52.3% died during 15-30 min. 67% of all TEPA were revealed in postoperative weak 1, 15%--in week 2, and 18%--on later terms (15-45 days). 82.6% of patients who died of TEPA underwent elective operations, and 17.4%--urgent operations. The indications were absolute in 67.3% of cases and relative--in 32.7%. 3/4 of all embolisms developed during the smooth p/o period, and 1/4--on the background of various complications. The rate of TEPA with lethal outcomes made up: 4.3% after the amputation of the lower limb for acute and chronic arterial circulatory disturbances, 2.4%--after transvesical adenomectomy of the prostate, 0.7% and 0.2%--after the operations on the biliary system and the stomach, respectively, and 0.14%--after appendectomy. 410 random purposeful autopsies detected that the sources of the TEPA in 99.3% of cases were thromboses in the system of inferior caval vein which developed silently in 88.3% of cases. In 90.9% of patients thrombus came from the deep veins of the shin, and in 9.1%- from iliac and femoral veins. The rate of postoperative TEPA as a cause of death, according to the autopsies data, has increased from 1.4% in 1972-1973 to 2.1% in 1990-1991. Up to 1997 the rate of this index decreased to 1.3%, the quantity of the autopsies (in operated patients--100%) being equal, and in conditions of increased surgical activity. The positive tendency in the dynamics of lethality of TEPA in conditions of a large industrial city, according to authors' view, is referred chiefly to implementation of drug prophylaxis of postoperative thrombosis of the deep veins of the lower extremities using mini-doses of heparine. PMID- 10710921 TI - [Clinico-diagnostic significance of fibronectin level determination in severely burnt]. AB - Fibronectin content was determined in plasma and granulation tissues of burnt wounds in burnt patients as well as in plasma and wound exudate of patients operated on for large postoperative abdominal hernia and in whom for several days after surgery the continuous discharge of the wound exudate through the drainage tubes was observed. The same data were observed in fibroblast cultures and nutritional medium in which they have been cultured. It was established that dynamic determination of plasma fibronectin in burn disease allows to estimate the adequacy of the treatment to timely predict probability of development of complications and the outcome of the disease, while determination of fibronectin content in the granulation tissues of burn wounds indicates optimal terms for transplantation of fibroblasts and autotransplants as well as a significant increase of the effectiveness of surgical treatment for victims with spacious burns. PMID- 10710922 TI - [Syndrome of systemic response to inflammation (lecture)]. PMID- 10710923 TI - [Combination of perforative duodenal ulcer and acute myocardial infarction]. PMID- 10710924 TI - [Polyps of choledochus]. PMID- 10710925 TI - [Intraintestinal endometriosis complicated by intestinal obstruction]. PMID- 10710926 TI - [False acute abdomen as a mask of some endocrine diseases]. PMID- 10710927 TI - Security update. WEDI summit prepares for HIPAA implementation. PMID- 10710928 TI - Behavioral health tomorrow. Stepping up to the challenge. PMID- 10710929 TI - Culture counts: how institutional values affect computer use. PMID- 10710930 TI - Resolution 2000: create an inviting E-practice. PMID- 10710932 TI - E-healthcare. Urging providers to embrace the Web. PMID- 10710931 TI - Aggressive collaboration: the Pittsburgh Center for Biomedical Informatics. PMID- 10710933 TI - Complying with the new coding regulations. PMID- 10710934 TI - Clinical decision support systems come of age. PMID- 10710935 TI - MGMA '99. Better medical group management through technology. PMID- 10710936 TI - An educational tool to illustrate cost-effectiveness in diagnostic pathways for coronary artery disease. AB - Rapid increases in healthcare costs have led to increased interest in the cost effectiveness of medical interventions. Coronary artery disease is responsible for a significant share of total healthcare spending, and therefore economic evaluations of medical procedures to treat the condition are potentially very important. We have developed a spreadsheet model as an educational tool that can be used to illustrate cost-effectiveness in the selection of diagnostic pathways (a "work-up" strategy of tests designed to reach a final diagnosis) for coronary artery disease. The model, in Microsoft Excel, is easy to use, requiring no specialist computer knowledge. It is menu-driven and the user navigates the model via a number of on-screen buttons. A data entry screen allows the user to customize the data for the key model parameters, making it possible to take into account location-specific features. The data entry screen also allows the user to undertake sensitivity analysis and rate "what if" scenarios. The model demonstrates how sensitive the cost-effectiveness of different diagnostic pathways is to the pretest probability of disease. This package could also be used as a decision support tool, although it is important to recognize some of its limitations for this purpose. PMID- 10710937 TI - Idiopathic retroperitoneal fibrosis: idiopathic or secondary to atherosclerosis? PMID- 10710938 TI - The role of advanced atherosclerosis in idiopathic retroperitoneal fibrosis. Analysis of nine cases. AB - Idiopathic retroperitoneal fibrosis is an uncommon inflammatory disease characterized by a periaortic fibrous mass throughout the retroperitoneum that often surrounds the ureters, leading to ureteral obstruction. An immune-mediated process has been suggested in which the antigen is derived from atherosclerotic plaques. The optimal treatment of patients with idiopathic retroperitoneal fibrosis is unknown because of its rare occurrence. Surgery, medical treatment with immunosuppressive drugs, or a combination of both are proposed. In this retrospective study we report nine cases of idiopathic retroperitoneal fibrosis treated in our hospital. We quantitated the severity of atherosclerosis by measuring the degree of calcification in the aorta and compared this with a group of 19 age- and sex-matched controls without idiopathic retroperitoneal fibrosis. The idiopathic retroperitoneal fibrosis patients had a higher percentage of aneurysms of the abdominal aorta. However, abdominal aortic calcifications are not more prominent in idiopathic retroperitoneal fibrosis patients than in age matched controls. These findings suggest that atherosclerosis as determined by abdominal aortic calcifications is not enough to provoke idiopathic retroperitoneal fibrosis. Furthermore, we evaluated the relapse rate of surgery versus medical treatment. In contrast to recent publications, no superiority of corticosteroids over surgical treatment could be demonstrated. However, our study is too small and retrospective. We estimate that a study which could find the most optimal treatment for idiopathic retroperitoneal fibrosis would need the participation of 40-50 medical centers each with a catchment population of about 200,000. PMID- 10710939 TI - Central diabetes insipidus preceding acute myeloid leukemia with t(3;12)(q26;p12). AB - A 52-year-old woman presented with polyuria and polydipsia. A diagnosis of central diabetes insipidus (DI) was made, which turned out to be the first sign of acute myeloid leukemia (AML). Cytogenetic analysis revealed a balanced translocation between chromosome 3 and 12 t(3;12)(q26;p12). The patient was treated with standard induction chemotherapy and vasopressin. Before consolidation chemotherapy could be administered, deep venous thrombosis was diagnosed and leukemia relapsed. Rescue chemotherapy was started. This is the first report of an association between AML with t(3;12) and DI. Its possible pathogenesis is discussed with a review of the literature. PMID- 10710940 TI - Fibromuscular dysplasia in a 63-year-old woman. PMID- 10710941 TI - Extrapulmonary small-cell carcinoma: report of three cases and update of therapy and prognosis. AB - Three male patients with extrapulmonary small-cell carcinoma originating from esophagus, pancreas and prostate are described. The patient with the esophagus tumor had a combined small-cell and undifferentiated carcinoma. The other two patients had a pure small-cell carcinoma. All patients were treated with primary combination chemotherapy consisting of etoposide and cisplatin followed in one patient by locoregional radiotherapy. The patients with the esophagus and the pancreas tumor showed a partial response; the patient with the prostate tumor achieved a complete remission but relapsed with brain metastasis. All patients are alive 7, 13 and 19 months, respectively after initiation of the therapy. As in pulmonary small-cell carcinoma, primary chemotherapy is the treatment of choice in extrapulmonary small-cell carcinoma. PMID- 10710942 TI - Relationship between pernicious anaemia and gastric neuroendocrine cell disorders. AB - The incidence of gastric carcinoid tumours is increasing. This rise is probably due to the number of gastroscopies and improved histological techniques. The majority (65%) of these gastric tumours is associated with chronic atrophic gastritis and pernicious anaemia. In this article two patients are presented, one with pernicious anaemia and gastric neuroendocrine cell hyperplasia and one with pernicious anaemia and multiple gastric carcinoids. These neuroendocrine cell disorders have a relatively favourable prognosis. Therefore, a wait-and-see policy was preferred. The pathogenesis, clinical symptoms, diagnosis, prognosis and treatment of these different neuroendocrine cell manifestations are discussed. We recommend performing a gastroscopy at the time of diagnosis for young patients with pernicious anaemia, and whenever abdominal problems, unexplained weight loss or aggravation of the anaemia arise. PMID- 10710944 TI - [Cardiac rupture in acute myocardial infarction]. AB - Free wall rupture of the heart is the most common cause of death following pump failure. The incidence of death is 10-16% of all deaths because of acute myocardial infarction (AMI). In respect of time between the onset of AMI to Cardiac Rupture (CR), early (80%) and late CR are distinguished. Other clinical classification distinguishes acute and subacute CR. CR is considered subacute if the time between the onset of typical symptoms of CR and irreversible shock is longer as 30 min. There are three problems to solve: 1) selection of patient particularly threatened with CR, 2) defining the prodroms of CR and early diagnosis, 3) advancing the methods of surgical treatment. CR occurs more often in women, hypertensive patients and patient > 60 years old sustaining the first infarction. Thrombolytic agents diminish overall mortality in AMI, but do not influence frequency of CR. There are three mechanisms of CR incidence: 1) blood effusion into the ischemic zone resulting in the loss of tissue strength, 2) influence of thrombolytic therapy on degradation and inhibition collagen synthesis, 3) absorption of collagen by lymphocyte infiltration in infarction zone. Cardiac insufficiency with cardiogenic shock and rapid increase of pericardial effusion in echo examination and electro-mechanical discordance are considered to be clinical signs of CR and tamponade. CRP is an independent marker of subacute CR. Surgical treatment is possible only in case of subacute CR. Pericardiocentesis and bloodletting could temporary diminish cardiac tamponade and allow transfer to the operating room. PMID- 10710943 TI - Tuberculosis screening among immigrants in The Netherlands: what is its contribution to public health? AB - Understanding the epidemiology of tuberculosis in migrant communities and designing adequate and comprehensive control strategies is a major challenge facing public health authorities in many low-prevalence countries. In The Netherlands, screening immigrants from tuberculosis high prevalence countries has been conducted since 1966. In this paper, we review risk factors for tuberculosis in migrant populations, the public health importance of tuberculosis and the current screening policy in The Netherlands. TB treatment outcome in migrant populations and operational considerations that ought to be taken into account to optimize current screening practices are also reviewed. The article recommends the setting-up of an information system to evaluate the effectiveness of screening immigrants in The Netherlands, and adjustment of screening policies where needed. PMID- 10710945 TI - [Dihydrotestosterone therapy of men with coronary artery disease and processes of peroxidation]. AB - In 11 men after myocardial infarction (mean age 58.8 years) influence of 3 months transdermal dihydrotestosterone (DHT) treatment on malondialdehyde (MDA), total cholesterol, HDL cholesterol, triglycerides and platelets was estimated. Decrease in MDA concentration after DHT treatment only in men with low (< 13 nmol/l) testosterone (T) level (p = 0.06) was observed. MDA concentration after 3 months DHT treatment was significantly lower in hypotestosteronemic in comparison to normotestosteronemic men (p < 0.05). Plasma lipid levels and platelet count were not affected by DHT administration. Decrease serum MDA concentration in low testosterone men with coronary artery disease may suggest that DHT treatment in this group has beneficial effects on peroxidation processes. PMID- 10710946 TI - [Diagnostic usefulness of +procalcitonin in internal medicine]. AB - Procalcitonin (PCT), a 116 amino acid prohormon of calcitonin, is a novel diagnostic marker of severe bacterial infection. The aim of the study was to evaluate the diagnostic usefulness of serum procalcitonin concentration in severely sick patients admitted to the department of internal diseases. Fifty one patients were included into the study and divided into two groups: group A--36 severely sick patients with different medical problems without signs of bacterial infection; group B--15 patients with severe bacterial infection (sepsis--7 patients, pneumonia--6, pyelonephritis--1, peritonitis--1). Twenty eight healthy controls were also included into the study. Serum PCT concentration measured by immunoluminometric assay was undetectable or low in control group (range 0.0-0.1 ng/ml) and group A (range 0.0-1.8 ng/ml). In group B (range 0.0-183 ng/ml) markedly elevated PCT levels were observed in all patients with sepsis, peritonitis and some patients with pneumonia. We conclude that high serum PCT level support the diagnosis of severe bacterial infection. PMID- 10710947 TI - [Diurnal variation of blood pressure in type II diabetic patients]. AB - The aim of the study was to evaluate diurnal variation of blood pressure in type II diabetic patients and to reveal important factors influencing 24-hour blood pressure profile in these patients. Investigations were carried out in 52 patients with type II diabetes mellitus including 28 patients with hypertension. Control group was 26 healthy men. In all subjects 24-hour ambulatory blood pressure monitoring was performed. In patients with diabetes mellitus both with and without concomitant hypertension significantly smaller night drop in blood pressure and heart rate was found out. There was no significant differences between subgroup of patients with diabetes mellitus with and without concomitant hypertension in night drop in blood pressure and heart rate. In patients with microalbuminuria where was smaller night drop in blood pressure and heart rate compared to patients with normal renal function. PMID- 10710948 TI - [Effect of hydroxylation polymorphism on the concentration of propranolol in the blood of humans with hyperlipidemia]. AB - An influence of hydroxylation phenotype on the concentration of propranolol [corrected] was examined in 52 subjects with hyperlipidemia divided into 4 groups: 1--control, normolipemic, 2--hypercholesterolemic, 3- hypertriglyceridemic, and 4--mixed-form hyperlipidemic. Each study group included extensive metabolizers and one subject characterized by a poor hydroxylation phenotype. Propranolol was given intragastrically at a single dose of 80 mg [corrected]. Blood was sampled within 24 hours following the drug administration. HPLC method was used for determining blood serum concentrations of propranolol. In each study group mean blood serum concentrations of propranolol in poor metabolizers were at maximum in subject with hypertriglyceridemia, at minimum in the normolipemic one, and intermediate in hypercholesterolemic (upper) and mixed form hyperlipidemic ones. Lipid metabolic disturbances also affected blood serum concentrations. They were the highest in hypertriglyceridemic patients, whereas in hypercholesterolemic were, in early stage of observation, even lower then in normolipemic subjects. Blood serum concentrations of propranolol [corrected] attained minimal values in patients with mixed form of hyperlipidemia. In the light of the present study we can state that hyperlipidemia modifies the blood serum concentrations of propranolol [corrected]. Although, the type of hyperlipidemia and lipophilic propranolol are not the only determinants affecting blood serum concentrations of propranolol, but also a genetic factor, i.e. hydroxylation phenotype may play an important role. PMID- 10710949 TI - [Polycystic kidney disease in newborns: analysis of coincidence of congenital abnormalities and other syndromes]. AB - The retrospective analysis of 45 cases of polycystic and multicystic kidney disease in newborns and fetuses was done, focused on the association with congenital anomalies in the another organs, regarding the classification of multicystic dysplasia, subcapsular (obstructive) dysplasia, autosomal dominant and autosomal recessive polycystic kidney disease. The analysis was based on the results of 1961 autopsies of newborns and stillborns. PMID- 10710950 TI - [Crystalline structure of urinary stones]. AB - After examining about 600 urinary calculus, their structure was characterised. Four kinds of renal stone structures were distinguished. A few particular structures were presented such as the ones whose centres were empty i.e. they had a cavity inside either completely empty or filled with loose or granulated powder (they amounted to 8 pieces i.e. 1.2% of all examined urinary stones). Another specific case was phosphate renal stone with a very complex layer structure. It was concluded that the establishing of so called nucleus is not decisive in the process of building up renal stones. There exist the renal stones which are built up of separate, big crystals, which cannot contain any nucleus or of solid mass of crystalline powder. The reasons of building up renal stones are only of physics-chemical nature i.e. they result from a simple phenomenon in which crystallization of chemical compounds from supersaturated solutions is involved. PMID- 10710951 TI - [The prolonged asystole during head down tilt test: prognostic significance and therapy]. AB - The head up tilt test is a worthy of acknowledgment and widely accepted tool used in diagnosing patients with syncope of unknown origin. In some cases tilting provokes asystole lasting even several tens of seconds. Not long ago this kind of vasovagal reaction was being associated with appearance of syncope combined with body injures. The prognostic significance of asystole during head-up tilt test and the safe and efficacious way of therapy in such cases hasn't been established yet. There were three cases of vasovagal reaction, resulting in asystole, described in order to show a large number of diagnostic and therapeutic problems related to the discussed syndrome. The presentation of actual views on diagnosing and therapy of patients with vasovagal syndrome, as a matter of fact is a synthesis of published results of clinical studies and its aim is to be more objective about so many obliging theories. PMID- 10710952 TI - [Carcinoid: unusual clinical course]. AB - Carcinoid is a slowly developing neuroendocrine tumour. It appears with frequency of 1.5/100,000 persons. Usually it is localized in appendix, small intestine, rectum and bronchi. Clinical sings. of carcinoid syndrome develop in only 10% cases of tumour. We present three cases of carcinoid: the first one with evidences of heart insufficiency, the second one with evidences of colon cancer, the third one coexisting with Graves-Basedow disease. PMID- 10710953 TI - [Ascher's syndrome: report of 2 cases]. AB - A fifteen year old boy and the forty two year old women were treated because of blepharochalasis. In both cases the goiter and cheilochalasis were observed. Ascher's syndrome was diagnosed on the basis of these three principal symptoms. The facial appearance and the eyelids functioning were improved by plastic surgical treatment. It also stopped the illness developing. PMID- 10710954 TI - [Difficulties in diagnosis of facial and perimaxillary tissue actinomycosis]. AB - The authors according to literature data, discuss some difficulties in diagnosis of three cases of facial and perimaxillary tissue actinomycosis. The histopathological examination of changed tissue is the most important point in diagnostic procedure in relation to clinical symptoms. PMID- 10710955 TI - [Obesity and cardiovascular diseases]. AB - Obesity is a chronic complex disorder, which requires long-term treatment. The aim of this study was to estimate on the basis of current literature the coexistence of the cardiovascular system diseases and overweight. It was concluded that obesity is an independent risk factor for coronary heart disease, hypertension and heart failure. PMID- 10710956 TI - [The assessment and clinical significance of heart rate variability]. AB - Heart rate variability (HRV) is a phenomenon to generation through the sinus node consecutive impulses in the different succession. HRV is regarded as a marker of autonomic nervous system tone of the heart. To assess HRV following methods: time domain, frequency domain and non-linear analysis are known. Time domain parameters correlate with frequency domain parameters. Some parameters can be used substitution, particularly reflect parasympathetic activity: rMSSD, pNN50 and HF. In clinical practice the most useful is time domain analysis based on 24 hours ecg Holter monitoring. Among time domain parameters the most significant prognostic value has SDNN. Decreased HRV following many diseases has been described. Significant prognostic value of decreased HRV after myocardial infarction (MI) and in patients with chronic heart failure (CHF) has been proved. Decreased HRV after MI is independent as well as ejection fraction (EF) sudden cardiac death risk factor. In patients with SDNN value below 50 ms high risk of cardiac death is observed. SDNN should be estimated on 7th day of MI to evaluate patients with high risk of sudden cardiac death. In patients after MI with ventricular tachycardia (VT) before VT decreased HRV is described. During MI beneficial influence of infarct-related artery patency on HRV is observed. HRV correlates with EF and infarct site too. HRV in patients with CHF correlates with EF and functional severity of CHF. Correlation between decreased HRV and increased mortality in CHF has been shown. In diabetic patients decreased HRV is observed. Following diabetes examination of HRV is useful to estimate early phase of autonomic neuropathy. Increase HRV parameters is observed in the course of beta-adrenolytic and converting enzyme inhibition treatment. In other diseases, including heart transplantation prognostic value of HRV and its clinical significance are still investigated. PMID- 10710957 TI - [Paracetamol prenatal toxicity]. AB - Paracetamol is common used over-the-counter (OTC) nonnarcotic analgesic and antipyretic drug during pregnancy. Several animal and human studies on reproductive effect of paracetamol have been done and the overall results do not implicate paracetamol as a teratogen. However, there are some dates that paracetamol administration during pregnancy may have an effect on lowering body birth weight and length. PMID- 10710958 TI - [Heliodor Swiecicki: precursor of anesthesia with nitrous oxide in obstetrics]. AB - Heliodor Swiecicki (1856-1926) gynaecologist and obstetrics had his own clinic in Poznan. He was one of the firsts who made anaesthesia with N2O and O2. Because of him it was possible to build by Ash & Sons machine with mixture of N2O and O2. He show his own constructions on X International Medical Congress in Berlin in 1890. It was a great success. PMID- 10710959 TI - Relationships among assisted suicide and religiousness, resources available, denial of dying, and autonomy. AB - A nonrandom sample of eligible voters in Michigan (N = 218; mean age 35.7 yr.) anonymously completed a questionnaire during the two weeks prior to voting on a ballot proposal endorsing physician-assisted suicide. Favoring assisted suicide correlated negatively with scores on religiousness, believing that only a troubled mind would favor assisted suicide and that vulnerable individuals would suffer were assisted suicide legal, with denial of dying, and with resources available during one's final years. Favoring assisted suicide correlated positively with indicating this to be a medical rather than moral issue, making one's own decisions on moral issues, and believing that people may have different opinions on assisted suicide. The role of experience with dying needs further study. PMID- 10710960 TI - Religiosity and mental health. AB - In a sample of 99 undergraduates, religiosity and psychoticism scores were negatively associated (r = -.34). PMID- 10710961 TI - Differences in cognitive complexity of adolescents with foreclosed and achieved identity status. AB - The purpose of this study was to assess whether the complexity of the self concept differs based on identity status in late adolescence. Adolescents in the achieved status have a strong sense of identity that has emerged following an intense period of exploration. Adolescents in the foreclosed status also have a strong sense of identity, but they have never been through a period of exploration. It was expected that adolescents in the achieved status would have more complex self-concepts than those in the foreclosed status. 62 university students were classified into the achieved or foreclosed identity status based on their scores on the Extended Version of the Objective Measure of Ego Identity Status-2. They then completed a trait sort to measure the complexity of their self-concepts. Complexity scores were calculated based on the H statistic, an index of dispersion derived from information theory. The results were as expected. Possible structural changes underlying the process of developing identity are presented, and the usefulness of structural self-concept models for studying development of identity is examined. PMID- 10710962 TI - Manic-depression, suicidality, and obsessive-compulsive tendencies. AB - In a sample of 73 undergraduates, obsessional-compulsive tendencies (and especially checking) were associated with scores on measures of manic, depressive, and suicidal tendencies. PMID- 10710963 TI - Temporal stability of the Launay-Slade Hallucination Scale for high- and low scoring normal subjects. AB - It has been documented that many normal people report hallucinatory experiences. The Launay-Slade Hallucination Scale is widely used to investigate differences between subjects who score high or low in hallucinatory predisposition. In this study we addressed the question of whether scores remain stable over time. We selected 19 high-scoring subjects and 17 low-scoring subjects (upper and lower quartile) from a group of 243 undergraduate students who completed the scale. These two groups retook the scale after a period of 3 to 6 weeks. 81% received the same classification, which supports the view of hallucinatory predisposition as a stable trait; however, for 30% of the subjects the mean rating at the second time differed more than 3 SDs from the mean rating at the first time. In addition, the mean rating of 19% of subjects changed enough so they did not receive the same classification, indicating that state characteristics may also affect these scores. PMID- 10710964 TI - Use of the MMPI and MMPI-2 in patients being evaluated for cardiac transplant. AB - The MMPI and MMPI-2 have been used extensively in the study of personality correlates in various coronary populations. It has been hypothesized that personality variables are associated with various clinical outcomes, such as quality of life, morbidity, and mortality; however, no data are available in a cardiac transplant sample. This study presents descriptive MMPI and MMPI-2 data for male and female patients awaiting cardiac transplant. Analyses of differences in MMPI-2 depression scales between cardiomyopathy groups for 366 men and 99 women were performed. Significant differences were found between ischemic and nonischemic male patients. PMID- 10710965 TI - Criterion-related validity of the Arabic Obsessive-Compulsive Scale in Kuwaiti and American students. AB - The Arabic Obsessive-Compulsive Scale and the Maudsley Obsessional-Compulsive Inventory were administered to 159 Kuwaiti (Arabic version) and 72 American college students (English version). Pearson correlations for total score on the two scales were .68 and .64 for Kuwaiti and Americans, respectively, denoting a good convergent validity of the new Arabic Obsessive-Compulsive Scale against scores on the Maudsley Obsessional-Compulsive Inventory in the two different national groups. PMID- 10710966 TI - Autobiographical recollection and affects of cues in an amnesic patient. AB - The individual described herein (Y.K.) is a pure amnesic patient with severe anterograde amnesia and a selective loss of specific personal episodes in his remote memories. In this study, we examined whether more concrete cues could elicit specificity in his autobiographical memory. It was found that the richness of his autobiographical memory under the condition with probes was significantly higher than condition without probes, although Y.K.'s performance on recalling autobiographical memory was inferior to that of control subjects under both conditions. These results and our previous findings indicate that neither the disorder at the level of the framework's thematic retrieval nor the destruction of individual traces will account for the loss of Y.K.'s autobiographical memory. We suspect that Y.K.'s loss of autobiographical memory may be due to problems in the interface between thematic retrieval frameworks and individual traces. PMID- 10710967 TI - Depressive and manic tendencies in high school and college students. AB - A sample of 101 high school students obtained higher mania scores than, but did not differ in depression scores from, a sample of 504 college students. PMID- 10710968 TI - Structure of personality disorders from the perspective of the Revised Neo Personality Inventory domain scales and the Psychopathology-5 Scales. AB - This study tested the generality and comprehensiveness of the five-factor model of personality as applied to the Personality Adjective Checklist's (Strack, 1987) personality disorder scales. A sample of 258 undergraduates (113 men and 145 women) completed the Personality Adjective Checklist, the Revised NEO Personality Inventory, and the Psychopathology-5 Scales for partial course credit. A combined principal axis analysis with varimax rotation was performed for nonoverlapping scales of the Personality Adjective Checklist, the Revised NEO Personality Inventory domain scales and the Psychopathology-5 scales. The results indicated four factors which were identified as Neuroticism, Extraversion, Disagreebleness, and Conscientiousness. Openness did not emerge as a separate factor. These results supported the comprehensiveness but not the generality of the five-factor model as applied to personality disorders. PMID- 10710969 TI - Disorder concept scales and personality dimensions in a young adult sample. AB - Previous research has shown correlations between normal personality variables of the Sixteen Personality Factor Questionnaire (16PF) and Axis II personality disorder scales using the Morey, Waugh, and Blashfield Minnesota Multiphasic Personality Inventory (MMPI) and Millon Clinical Multiaxial Inventory. This study (N = 37) compared variables from the adolescent version of the 16PF, the High School Personality Questionnaire, Revised, including the new Clinical Supplement and the MMPI scales of Morey, et al. and yielded results similar to those from earlier studies with other inventories. Extraversion scores correlated positively with those on Narcissistic and Histrionic scales, negatively with scores on Schizoid, Avoidant, and Schizotypal scales; scores on Independence had a similar pattern. PMID- 10710970 TI - Effects of a small monetary incentive and follow-up mailings on return rates of a survey to nurse practitioners. AB - The purpose was to examine the effectiveness of a modest monetary incentive ($1) and none in increasing the response rate of a mail survey to 600 nurse practitioners. The response rate in the incentive group was 81% and 66% in the control group, significant by chi-square test. The most cost effective survey technique for increasing the response rate of nurse practitioners was to code the envelopes and eliminate the monetary incentive. PMID- 10710971 TI - Criterion validity of a new measure of self-actualization. AB - Many authors have questioned the validity of the major existing measures of self actualization (the Personal Orientation Inventory, the Personal Orientation Dimensions, and the Short Index of Self-actualization), given their serious theoretical and methodological limitations. A new inventory was developed, the Measure of Actualization of Potential, and various studies were conducted to assess its theoretical and empirical content as well as its construct validity, internal consistency, and temporal reliability. The present aim was to estimate the criterion validity of the new measure using humanistic clinical psychologists' assessments as the criterion. The results show that eight clinical psychologists' rankings of 73 individuals are positively and highly correlated with the scores obtained by the same individuals on the Measure of Actualization of Potential. PMID- 10710972 TI - Substance abuse and academic and career problems for three age groups of college students. AB - Scores on the Substance Abuse, the Academic Problems, and the Career Problems subscales of the College Adjustment Scales were compared for 30 college students in three age groups. No significant mean difference was found among age groups on any scale. PMID- 10710973 TI - Peer-based interventions to influence health-related behaviors and attitudes: a meta-analysis. AB - The effects of 47 peer-based health education programs described in 36 published studies were examined. The over-all effect size was small: the mean d was .190 when controls received no program and .020 when controls received an alternative program. Programs were divided into those focusing on preventing or reducing smoking and programs on other health issues; the latter were further divided into primary prevention and secondary prevention programs. Differences among studies suggested several biases which were likely to have influenced the effect sizes. Preventive interventions that produce only small effects can be valuable because many participants would not have developed the problem even without the program. This review suggested that, when health education programs are studied, (a) detailed statistical information should be provided to facilitate using the research findings in meta-analyses and (b) the costs of innovative programs should be presented to judge whether the results are worth the cost. PMID- 10710974 TI - Suicide-intervention trainees' perceptions of awareness for warning signs of suicide. AB - 55 trainees in suicide intervention were administered a revised version of the Suicide Opinion Questionnaire to examine the effects of suicide intervention training and experience in suicide prevention on their perceptions about the availability of suicide warning signs. Over-all, results indicated that most participants agreed with the ideas that warning signs of suicide are usually evident and that family members of those who have committed suicide may not be aware of these signs. Effects of intervention training on perceptions about the availability of suicide warning signs were not evident. Multivariate analysis indicated that participants with less than one year of experience in suicide prevention expressed significantly (p < .01) more agreement than participants with between one and five years of experience in suicide prevention for the idea that family members may not be aware of suicide warning signs in their suicidal relatives. PMID- 10710975 TI - An exploratory study of the hardy personality at work in the health care industry. AB - The concept of personality hardiness is tested for its contribution to perceptions of the nursing work environment. Hardiness has been traditionally linked to positive health outcomes and stress resistance, but the traits which lead to disease-resistance logically have other effects on the individual. Results of this exploratory study indicate that hardiness is negatively linked to perceptions of work pressure and role ambiguity. Positive relationships were found between hardiness and job clarity, organizational involvement, and peer cohesion. Suggestions for additional study are enumerated. PMID- 10710976 TI - Assessment of optimistic self-beliefs: further validation of the Chinese version of the General Self-Efficacy Scale. AB - The Chinese version of the General Self-efficacy Scale developed by Jerusalem and Schwarzer was tested in a sample of 74 Chinese adults with mild mental health symptoms. Analysis showed the scale was unidimensional and had good internal reliability (alpha = .92). The scale score also differentiated groups of different mental health status and correlated strongly with scores on the General Health Questionnaire, State-Trait Anxiety Inventory, and Center for Epidemiologic Studies--Depression Scale. Test-retest reliability over six months was also adequate. PMID- 10710977 TI - Sympathomimetic preference and ethnicity in male felons. AB - A disproportionate number of Euro-American inmates given either an amphetamine- or cocaine-abuse diagnosis upon admission to prison were given a diagnosis of amphetamine abuse. A disproportionate number of African-American inmates were given a cocaine-abuse diagnosis. Of the 800 consecutively admitted inmates who were given one diagnosis or the other, 68% of the cocaine abusers, but only 4% of the amphetamine abusers, were African American. PMID- 10710978 TI - Measurement of laboratory aggression: a new response-choice paradigm. AB - 40 undergraduate student volunteers were tested on a new Response-choice Aggression Paradigm. Men and women were provoked in a reaction time competition by receiving electric shocks and were allowed to respond to a confederate with similar shocks or to refrain from any retaliation. Analysis indicated positive association between a self-report measure of physical aggression and laboratory responses on the paradigm, and positive associations among aggression indices of the task. The results confirm earlier findings of sex differences in aggression and offer new measures of aggression "flashpoint" as a step closer to aggressive behavior in naturalistic settings. PMID- 10710979 TI - The visual analogue mood scale: can a single-item scale accurately classify depressive mood state? AB - Visual analogue mood scales provide extremely rapid, single-item assessment of affective states. This study examined discrimination on a single-item visual analogue depression scale between dysphoric or mildly depressed and nondepressed individuals in a sample of 284 college students. The visual analogue mood scale significantly discriminated subjects by mood category and achieved comparable hit rate, sensitivity, and specificity relative to other well validated mood scales. PMID- 10710980 TI - Perceived family environment and adjustment in American-born and immigrant Asian adolescents. AB - 57 immigrant Asian adolescents were compared with 44 American-born adolescents of Asian descent to investigate differences in perceived family environment and adjustment. Immigrant Asian adolescents were significantly less adjusted, perceived significantly less independence and achievement orientation and significantly more organization in their families than their American-born peers. The family environment differences, unlike adjustment differences, persisted over length of time in the host country (USA). PMID- 10710981 TI - Correlates of divorce rates in Caribbean nations. AB - Divorce rates in 14 Caribbean nations were associated with the size of the nation and the amount of inwardly-directed violence. PMID- 10710982 TI - Laboratory measurement of aggression in high school age athletes: provocation in a nonsporting context. AB - This study investigated the relationship between aggression and type of sports involvement in high school age boys. Athletes (16 boys), ages 15 to 18 years, were separated into two groups, one of 8 athletes who participated in sports with high physical contact, e.g., football and basketball, and the other of 8 athletes who participated in low contact sports, e.g., track and baseball. Students participated in six 25-min. Point Subtraction Aggression Paradigm sessions. The paradigm is an established laboratory model of aggression with three response options: (1) a point-maintained response, (2) an aggressive response, and (3) an escape response. Analysis indicated that the only difference between the groups was that individuals who participated in high contact sports emitted significantly more aggressive responses than individuals who participated in low contact sports. Similarly, psychometric measures of aggression indicated that individuals in the former group self-reported more behavioral incidents of aggression than those in the latter group. PMID- 10710983 TI - Noticing. PMID- 10710984 TI - A critique of the Model State Social Work Practice Act. AB - This article reviews the Model State Social Work Practice Act approved by the American Association of State Social Work Boards in 1998. The limitations of the definition of social work practice included in the model stature, inclusion of BSWs and social workers employed by public and nonprofit agencies, the promulgation of dual codes of ethics, and lack of assurance of effective services are discussed. Strategies for political action to benefit consumers, the public, and social workers are provided. PMID- 10710985 TI - Valuing families: social work practice with families from a strengths perspective. AB - Social work and social workers have long been concerned with families. Historically, most approaches to social work with families have focused on individual pathology and problem solving or have considered problems of a family member to be symptoms of family dysfunction. In contrast, other approaches to social work have focused on growth, function, and healing. This article describes both problem-focused and growth-focused approaches to practice and presents a strengths approach to practice that values families and builds resilience. Assumptions of the strengths approach are discussed and applied to work with families through a case example. PMID- 10710986 TI - Effects of court-ordered substance abuse treatment in child protective services cases. AB - Courts often play active roles in the lives of families supervised by child protective services (CPS). Judges adjudicate dependency, mandate services, determine placements of children, and order continued supervision or termination of parental rights or services. This study examined the effects of court orders in preventing recurrence of substance abuse in the cases of 447 children in kinship care while under CPS supervision. In addition, the effects of court orders on duration of service and on numbers of placements were studied. Results suggested that court interventions had mixed outcomes. Levels of compliance with mandated substance abuse and mental health treatment did not appear to influence rates of reabuse or duration of service. Court orders appeared to affect both the number of caretakers and placements the children experienced. Children adjudicated dependent were more likely to have multiple caretakers than those under voluntary supervision. PMID- 10710987 TI - Social workers employed in substance abuse treatment agencies: a training needs assessment. AB - This article describes the results of an assessment of the substance abuse treatment training needs of social workers working in randomly selected substance abuse treatment facilities in New England. This assessment revealed that clinical supervision related to substance abuse treatment had not been available to a significant percentage of the respondents throughout their careers. Despite limited previous training experience and considerable barriers to current training, social workers surveyed in this study reported significantly higher levels of knowledge and skill than other substance abuse treatment providers in 10 of 12 substance abuse treatment areas investigated. Despite these high levels of knowledge and skill, respondents reported considerable need for and interest in additional substance abuse treatment training. This study identified the areas of assessment, advanced clinical techniques, and dual diagnosis as priorities for future training among social workers working in substance abuse treatment facilities. PMID- 10710988 TI - A topography of self-help groups: an empirical analysis. AB - The current managed health care system creates an environment in which social workers need to be knowledgeable about low-cost interventions. Self-help groups have the potential to be beneficial to social workers' clients. Surprisingly, little is known about the characteristics and activities of many groups and the extent to which groups receive guidance and support from professionals and established national and local organizations. Whereas many social workers are aware of Alcoholics Anonymous (AA), there are thousands of other types of groups that could be helpful to their clients. This study examines the member and group characteristics, professional involvement, and local and national affiliations of 253 self-help groups. Results suggested that many groups have shared leadership, recruit group members, receive assistance from professionals, and receive guidance from national and local organizations. Results are discussed in terms of how social workers can assist and use self-help groups in the current managed health care system. PMID- 10710989 TI - Use and support of multicultural and antiracist education: research-informed interdisciplinary social work practice. AB - Multicultural education (MCE) and antiracist education (ARE) are the primary curricula through which school educators are combating the effects of racism and bigotry. To assist in the efforts of the educators, social workers need an understanding of MCE and ARE objectives, assumptions, and current research to guide their practice. This article discusses the differences between MCE and ARE and presents the findings of a study conducted in five northwestern United States school districts. It examines curricula, policies, and practices used to address racism and bigotry among elementary, middle, and high school students; attempts to discover underlying barriers to implementing antiracist and antibigotry curricula; and reveals the perspectives of teachers, administrators, counselors, and social workers. Implications for social work practice, policy development, and research are discussed. PMID- 10710990 TI - Design evaluation: illuminating social work practice for better outcomes. AB - One of the difficulties in social work lies in finding forms of evaluation that suit the various kinds of work people actually do. There is a need for methods that allow a valid and rigorous evaluation of process as well as outcomes, particularly in new areas of work. A valuable example of such evaluation evolved at St. Luke's, a voluntary agency in Victoria, Australia, that used design evaluation to articulate a new model of practice. Design evaluation involved having the evaluator work with staff to describe work practices as a series of stages with associated principles. This process of documenting, clarifying, and illuminating the model led to its progressive refinement and concurrent improved services delivery to clients. Testing the program model with clients provided some valuable feedback about its effectiveness. This form of evaluation is complementary to outcomes evaluation and can also provide the information needed for a subsequent outcomes evaluation. PMID- 10710991 TI - Adoption policy in the United States: a word of caution. PMID- 10710992 TI - Wanted: knowledge and wisdom. PMID- 10710993 TI - Recovered memory therapy: a dubious practice technique. PMID- 10710995 TI - Constructing the future. PMID- 10710994 TI - Recovered memory therapy: a dubious practice technique. PMID- 10710996 TI - Progress towards poliomyelitis eradication, Chad, 1996-1999. PMID- 10710997 TI - NHS executive tells nurses to be 'public spirited'. PMID- 10710998 TI - Hospitals need specialist inpatient adolescent units. PMID- 10710999 TI - Case 4: neighbourliness (a). Nurses committing offences outside the work environment. PMID- 10711000 TI - Confidentiality. 1: Nurses' duty to respect patient confidentiality. AB - Trust is at the heart of the nurse/patient relationship, yet many nurses find themselves in situations where they are tom between their duty of confidentiality to the patient and other duties. This series of articles seeks to analyse the source of the duty of confidentiality and discusses individual situations giving rise to conflict within the statutory law context. PMID- 10711001 TI - Urinary catheter management: minimizing the risk of infection. AB - With catheterization comes the risk of infection and therefore people should not be catheterized unless their clinical condition dictates that it is absolutely necessary. Nurses are responsible for both inserting catheters and the subsequent management of the catheterized patient. A high level of nursing knowledge and skill is required to achieve effective and safe management. This article continues the infection control series by reviewing the principles of catheter management with regard to controlling infection. PMID- 10711002 TI - Catheter valves: a special focus on the Bard Flip-Flo catheter. AB - This article examines catheter valves with a special focus on the Bard Flip-Flo. Catheter valves were first introduced into the UK in 1986. Ten years later, seven catheter valves were available of which three, including Flip-Flo, were available on prescription. Catheter valves are suitable for both males and females and both short- and long-term urethral or suprapubic catheters. Catheter valves can combine with urinary drainage bags to form link systems. The benefits of catheter valve usage, many of which are perceived but not well researched, include reduced urinary tract infection and maintenance of bladder tone and capacity. Current opinion indicates that patients with detrusor instability, impaired bladder sensation or confusion may find catheter valves unsuitable. Catheter care, including health education and monitoring, are required for catheter valve patients as well as conventional leg bag users. The Bard Flip-Flo catheter valve offers many benefits and is an established product within the UK. PMID- 10711003 TI - Family carers. 1: Difficulties and levels of support in Sweden. AB - Sweden has a well developed welfare system and until fairly recently the needs of family carers have received limited attention. However, there is now a growing acceptance of the need to provide better and more sensitive support services. Using questionnaires validated in the UK and translated into Swedish, this series of four articles considers the difficulties, satisfactions and coping mechanisms of a sample of Swedish carers, identifying the implications of the results for the design and delivery of healthcare interventions. This article focuses on carers' perceived difficulties and their satisfaction with existing levels of help. PMID- 10711004 TI - The placebo effect and the hidden benefits of oral medications. AB - The effectiveness of any drug involves many factors not just the pharmacological ingredients. The hidden benefits of a drug are termed the 'placebo effect', but because many of these factors cannot be explained scientifically they are often dismissed as irrelevant. However, factors including the attributes of the drug itself (shape, colour, taste), the relationship between the nurse dispensing the drug and the patient receiving it, the route of administration, the setting in which the drug is given and wider sociocultural beliefs surrounding the ingestion of medicines can all have a major influence on a drug's efficacy. This article explains the term 'placebo' and explores some of the factors contributing to the placebo effect in relation to oral medications. Nurses need to appreciate the influence of such factors in order to provide appropriate and holistic care to patients with different social and cultural beliefs and expectations. PMID- 10711005 TI - Stoma care: empowering patients through teaching practical skills. AB - Teaching patients practical skills in stoma care is a complex process and although, arguably, at the very heart of stoma care nursing practice, has been largely ignored in the literature. Teaching principles are based upon social learning theory and educationalists provide guidelines on the most effective way to teach a practical skill. These guidelines have been utilized by nurses when teaching patients with newly formed stomas how to change a pouch. The process of adapting to a stoma and its daily management takes time. Psychologically, however, some patients will adapt more easily than others and researchers have attempted to identify factors which may account for this. Studies have demonstrated that patients who are satisfied with the amount of preoperative information they receive are less likely to develop psychological problems. Psychological adjustment may be affected if patients feel that they have developed insufficient pouch changing skills or have problems with leakage from their pouch or sore skin around their stoma. Studies have also demonstrated that cognitive factors, such as patients feeling in control of their illness and stoma, have been found to play a role in psychological adaptation. Clinical nurse specialists in stoma care are in an ideal position to target these cognitive factors using a variety of strategies including effective practical teaching to empower patients, thus facilitating psychological adaptation following stoma surgery. PMID- 10711006 TI - Sexual health. 1: Sexuality and nurses' role in sexual health. AB - This article, the first of two, looks at the issue of sexuality in health care. Sexuality underpins much of what a person is and has significance in everyone's life. Through sexuality, people express their most intimate feelings of individuality and their need for emotional closeness with other human beings. Sexuality is not just about sexual intercourse, it is about the concept of people as men and women--about their manliness or femininity. Sexuality also affects the way people see themselves or would like to be seen, their appearance and behaviour, and their desire to attract those who matter to them. It is about the fears and fantasies people have about themselves and others. Nurses need to have an understanding of sexuality as they may have a crucial part to play in its dynamic progress. PMID- 10711008 TI - Development of a meaningful learning environment in theatres. AB - During the 1990s, geographical distancing of clinical areas and nurse education establishments led to breakdowns in communication. This article focuses on how Good Hope Hospital NHS Trust and the University of Central England in Birmingham created a meaningful learning environment for student nurses in the operating theatre using a revised version of Crofts and Taylor's (1996) perioperative placement model. The student evaluation has demonstrated that the quality of placement, teaching and supervision were of a high standard and students felt that their nursing skills had been enriched as a direct result of the placement. This article will also examine the background to student placements in the operating theatre. PMID- 10711007 TI - Trident T2: a new development in urine bag tap design. AB - This article looks at Seton Scholl Continence Care's Simpla Trident T2 leg bag with the new advanced lever-action tap which has recently won the Independent Living Design Award. This easy-to-use lever-action tap has a simple, positive action which allows more independence and less intervention from carers. The lever-action tap enables the patient with limited hand coordination or poor finger/thumb dexterity to operate it effectively. The tap is smooth to the touch and the corners have been rounded to reduce any sharpness. The Simpla Trident T1 and T2 bags have the protected sample port to allow safe urine sampling without the need to break the system. PMID- 10711009 TI - Health and safety in the NHS. AB - With the increase in the number of employer liability claims there is a need for all healthcare organizations to demonstrate that they can meet the requirements of NHS health and safety issues (HSG(97)6). The NHS is the second biggest employer in Europe and is currently paying out in excess of 30 million Pounds on employer liability cases. Health and safety strategy should aim to remedy the current situation, enabling healthcare providers to meet legal and common law duties of care. A clear and focused programme for action can be established, building on the current momentum for change and development. PMID- 10711010 TI - The qualities needed to become an expert nurse. PMID- 10711011 TI - More clinical skills but not at the expense of theory. PMID- 10711012 TI - Consumers not confident about self-regulation. PMID- 10711013 TI - Case 9: Failure to take appropriate action. Staff nurse who failed to care adequately for a patient after a fall. PMID- 10711014 TI - The selection of female urinals: results of a multicentre evaluation. AB - Female urinals are designed to enable women to empty their bladders while not on the toilet and are therefore potentially useful in preventing incontinence. However, there is little published information to guide product selection. Therefore, an evaluation of these products was undertaken by the Continence Products Evaluation Network (funded by the Medical Devices Agency). All 13 reusable female urinals available in the UK in March 1997 were evaluated. Each urinal was evaluated by 28-32 community-based women. Preliminarily, each subject tested all urinals by trying to place them in one or two of their preferred positions, to establish if the urinals were suitable for full testing. Each of the urinals that were selected for full testing were then used for 1 week each. During this week the subjects kept a diary to record leakage or spillage when using the urinal. At the end of the week a product evaluation form was filled in to record product performance. The results from full testing indicate that all urinals were successful for some subjects. However, some urinals were found to be successful for all four main positions (e.g. Petal Female Urinal) while others were successful mainly in one or two positions (e.g. Bridge Saddle Pan and Subaseal). Many urinals were successful in the standing/crouching and sitting on the edge (of chair or bed) positions, while comparatively few urinals were successful in the lying position. It was found that the chances of finding a suitable urinal increased with levels of independence. This means that subjects with higher levels of dependency found fewer urinals to be suitable for their needs when used without assistance. The results of this evaluation provide guidance for product selection. However, it is recommended that continence specialists keep samples of the full range of female urinals to enable women to experiment with urinals in order to find one that best suits their needs. PMID- 10711015 TI - Inflammatory bowel disease. 2: Medical and surgical treatment. AB - The first article in this three-part series reviewed the aetiology, treatment and concept of participation in inflammatory bowel disease. This article provides a brief overview of current trends in the medical and surgical treatment of inflammatory bowel disease. The discussion of medical management will include the most common drugs utilized for symptom control, their modes of action, routes of administration and side-effects. In addition, the role of nutritional support, both as an adjunctive and a primary therapy, will be considered. The options for, and consequences of, definitive surgical intervention in the event of failed medical management of both ulcerative colitis and Crohn's disease will be examined. PMID- 10711016 TI - Mediating factors in the grieving process of the suddenly bereaved. AB - This article explores the role of the staff of accident and emergency (A&E) departments in managing events surrounding sudden death. Key aspects, such as the way in which initial contact with relatives is made, reception and resuscitation procedures, viewing of the deceased, and the bureaucratic elements of these events, are discussed in terms of the impact that they may have on the subsequent adjustment of the bereaved to the sudden death of their relative. The management of death may be problematic, particularly if it involves children or if the body of the dead person is mutilated. This article suggests ways to support health professionals in facilitating the processes of grieving, and raises awareness of the potential influence the experiences in the A&E department at the time of bereavement may have on the grieving process. PMID- 10711017 TI - Teaching communication with the dying across cultural boundaries. AB - This article is based upon the author's personal experience, gained as a teacher of palliative care, when he was invited to be part of a small humanitarian aid team of experienced British nurse educators which worked for 2 weeks in the newly opened First Moscow Hospice in October 1997. The team was to act as both teachers and clinicians assisting the Russian health professionals to both understand and use the palliative care approach in a workable context for their culture. A case study is used to illustrate the ethical and practice-based nursing issues that arose which were related to communication difficulties, challenging myths and cultural norms, and the use of 'self' to demonstrate good practice. This article highlights how it is possible to overcome some of these difficulties, and shows what an honest and sensitive approach to communicating with the dying can achieve. PMID- 10711018 TI - Infectious diseases: statutory notification and surveillance. AB - The notification of infectious diseases has evolved over a number years and will continue to do as practices change or are modified. This article gives an insight into how the system for notification has developed. It outlines the process and purpose for notification and how the accumulation of data gathered can help nurses, midwives and health visitors make strategic plans for intervention and prevention of further outbreaks of disease. The nurse, midwife or health visitor has a central role to play in ensuring that diseases are notified to the 'proper officer'. Furthermore, if nurses, midwives and health visitors were given the power to notify diseases, the number of diseases notified may increase. This would result in a clearer and truer account of the number of notifiable diseases. PMID- 10711019 TI - Job satisfaction among Swedish nurses and laboratory technologists. AB - Concerns about recruitment and retention of nursing staff are currently increasing in a number of countries. This article describes a study which identifies sources of job satisfaction among nurses and medical laboratory technologists in Sweden. A previously validated questionnaire was translated into Swedish and sent to all relevant staff working at a large hospital in southern Sweden. Analysis of responses suggests that while staff still found their work interesting, rewards are potentially decreasing and sources of stress increasing. Comparisons are made with literature from the UK and USA and the need for action to address a number of concerns is highlighted. PMID- 10711020 TI - The Pentaflex pressure-reducing mattress replacement. AB - Support surfaces play a crucial role in pressure sore prevention. This article outlines the use of polyurethane foam as a versatile and effective support surface. It briefly describes a number of tests which are used to determine foam quality and summarizes the characteristics and clinical evidence supporting the use of the Pentaflex pressure-reducing foam mattress. PMID- 10711021 TI - Ethics and nursing research. 2: Examination of the research process. AB - In this article, the second in a series on ethics in nursing research, the author explores the relationship between the guiding ethical principles and the steps of the research process. In the first article (Vol 8(13): 888-92) the two dominant theories of ethics, utilitarianism and deontology, along with the guiding principles of beneficence/non-maleficence and respect for human dignity, justice, informed consent and vulnerable subjects were discussed as they relate to the rights of individuals undergoing the research. In this article, the author describes the association between these principles and the elemental steps of the research process which are: the selection of the research problem; data collection; sampling; informed consent; data analysis; and research presentation. The ethical conduct of many of these research steps is guided by ethics committees but for those that are not nurses need to rely on their own integrity, honesty and committment to the current prevailing ethical principles. PMID- 10711022 TI - Confidentiality. 6: HIV/AIDS patients and the duty of confidentiality. PMID- 10711023 TI - Nursing must be the core of nurse consultants' work. PMID- 10711024 TI - Public and private sectors need the same standards. PMID- 10711025 TI - No patient should be considered untreatable. PMID- 10711026 TI - Case 10: Inappropriate therapeutic relationships. Psychiatric nurse who formed an improper relationship with a client. PMID- 10711027 TI - Central venous catheters: preventing infection and occlusion. AB - As central venous catheters (CVCs) become more widely used in today's healthcare environment, nurses require expert knowledge in relation to CVC maintenance to prevent complications and maximize efforts to optimize the individual's health status. This is especially so since CVCs have begun to be used outside intensive care units, e.g. in general wards, and can be associated with high incidences of infection, occlusion and subsequent compromise in patient health. Nurses are responsible for the maintenance and use of central access devices, such as CVCs, resulting in a need for literature specific to the nursing aspects of CVC management. This article addresses many nursing issues pertaining to care of the central line, focusing on evidence- and research-based literature, and also reviews the literature to make recommendations for practice. PMID- 10711028 TI - Efficacy and cost-effectiveness of the Thermo contour mattress. AB - Use of air mattresses in reduction of pressure sore incidence is an important part of quality patient care. However, there will never be enough air mattresses to match the demand as increased education and an increase in the general provision of air mattresses can lead to an unrealistic expectation of obtaining air mattresses when required for patients. This raises the demand for air mattresses and increases costs within hospital trusts. This study examined a way to redress the balance through use of an alternative, cost-effective type of mattress (thermoelastic polymer) in the prevention of pressure sores. Prime consideration was given to the comfort of the patient. The claims of the manufacturers, Barrington Healthcare, that 'patients with Waterlow scores of 23 can be nursed safely on this product' were explored as part of the study. A total of 407 patients took part over a 6-month period. Twenty-four Thermo contour mattresses were loaned to one ward for the study and patients were allocated to the mattress through admission to the experimental ward. Patients were then split into two groups on two wards. Group B were allocated to a Thermo contour mattress and group A were nursed on air mattresses and foam mattresses generally used throughout the trust. Results showed that more patients were comfortable on the Thermo contour mattresses than on all other mattresses. Patients with Waterlow scores under 25 did not develop pressure sores on the Thermo contour mattress. However, the sample of patients with a Waterlow score of between 20 and 25 was small and therefore further research is required. PMID- 10711029 TI - Psychological methods of treating hallucinations and delusions: 1. AB - This article, the first in a two-part series, describes how the neuro-biological, social and environmental factors involved in the course and treatment of schizophrenia have helped to establish a pathway to recovery or remission that does not embrace pharmacological therapy alone. Research into drug-resistant symptoms and poor adherence rates (Curson et al, 1985) demands effective alternatives and additions to more traditional approaches to treatment such as medication. The second part of the article describes how evidence-based psychological interventions were used to help a client suffering from treatment resistant delusions, allied to a chaotic lifestyle and substance misuse, that had previously presented a substantial challenge to the mental health services. PMID- 10711030 TI - Seasonal affective disorder: its recognition and treatment. AB - This article provides an introduction to seasonal affective disorder (SAD) and outlines various therapies, including phototherapy (light therapy), used in its treatment. SAD, colloquially termed 'winter blues', is a common condition that is thought to be caused by reduced levels of daylight in winter. During this period sufferers generally feel low and may experience clinical depression. The Department of Psychiatry at the University of Southampton has an established SAD service as part of its mood disorders clinic, which was developed from a research based clinical investigation unit set up in the early 1990s. SAD is described as a mood disorder with a seasonal pattern and has a greater prevalence in countries with greater northern latitude. The aetiology of SAD is unclear, although the most promising theory suggests the role of the neurotransmitter serotonin. SAD is difficult to treat with conventional antidepressants although there is evidence that serotonin selective reuptake inhibitors may be useful for some patients. Phototherapy (light therapy) has been used successfully by many patients although it remains controversial and difficult to obtain on the NHS. PMID- 10711031 TI - Evaluating clinical supervision. PMID- 10711032 TI - Inflammatory bowel disease. 3: importance of partnership in care. AB - In the preceding articles (Vol 8(13): 858-62; Vol 8(14): 926-30), the pathogenesis and aetiology of inflammatory bowel disease (IBD) and its medical and surgical treatment options were considered. In this final part of the series, the concept of an alliance between the patient with IBD and healthcare professionals in terms of healthcare choices is examined. The article explores the role of the clinical nurse specialist in IBD in relation to education, psychological support and compliance with treatment regimens. There is also a discussion of the social and psychological impact of IBD on the patient, and an exploration of the positive effects such a participatory relationship may have on health and quality of life. PMID- 10711033 TI - Examining the range of medicated and paste-impregnated bandages. AB - Medicated and paste-impregnated bandages have been used by healthcare professionals for many years, especially in the management of problem skin surrounding leg ulcers. This article examines the range of bandages and the indications for use as well as the role of zinc in wound healing. PMID- 10711034 TI - Drawtex: a unique dressing that can be tailor-made to fit wounds. AB - Drawtex is a new and innovative dressing using dispersion technology which works on capillary action. This promotes moist wound healing and provides the optimum environment at the wound interface. This dressing has the ability to absorb exudate 30 times it own weight. The unique capillary action draws the exudate away from the wound bed and into the core of the dressing from where it disperses into a second layer of Drawtex. The practitioner tailors the Drawtex to conform to the wound bed. Drawtex's non-adherence reduces the frequency of dressing change after the first week. This is not the only benefit as it is also very cost effective at half the price of other modern dressing products. PMID- 10711035 TI - Improving the nutrition of patients using Entera Fibre Plus. AB - This article examines the role of fibre in the diet with a special focus on Entera Fibre Plus, a new high energy, fibre-containing sip feed. Early enteral feeds were all fibre-free or 'low residue'. In the last 10 years fibre-containing feeds have become available, mostly designed for tube feeding and containing a single fibre source. Entera Fibre Plus contains a mixed fibre source, providing both soluble and insoluble fibre types, including inulin. This helps provide the full range of fibre benefits, which include reduction of constipation and diarrhoea, maintaining the health of the gut and encouraging beneficial gut bacteria to assist the body's defence systems. Inulin has a specific role in the latter. Entera Fibre Plus provides 300 kcal and 5 g of fibre per 200 ml pack and is available in six flavours. It is a convenient and palatable way of improving the nutritional status of patients who are malnourished and may be of particular benefit to elderly people, those on long-term supplements and those who suffer from chronic constipation or diarrhoea. Entera Fibre Plus is approved by the Advisory Committee on Borderline Substances (ACBS) and is available at retall pharmacles or on prescription. PMID- 10711036 TI - Prison health care: what is it that makes prison nursing unique? AB - Nursing in the prison service is an emerging area of practice and is becoming increasingly recognized for the benefits nurses provide to patient/prisoner care. This article, the second in a series of articles on prison nursing, highlights some of the current issues that face prison nurses. Also one person's experience of shadowing a prison nurse is described. PMID- 10711037 TI - Valuing diversity and ethnicity: UKCC initiative. PMID- 10711038 TI - Should children be able to request euthanasia? PMID- 10711039 TI - Nurses have an important role in consent issues. PMID- 10711040 TI - Case 11: Sexual harassment at work. Senior staff nurse who took advantage of his position. PMID- 10711041 TI - Managing and maintaining a safe environment in the hospital setting. AB - On entering hospital, patients and visitors assume that they are in a safe environment. Maintaining a safe environment in hospitals depends on not only the infrastructure, but also the equipment and materials that are used on the premises. Complaints about hospitals often include comments on the environment, its lack of cleanliness, poor food and the general look of debilitation. Key legislation for managing a safe environment is the Health and Safety at Work Act 1974. Complementary guidance includes the Control of Substances Hazardous to Health Regulations 1999 and the Environmental Protection Act 1990. The Incorporation of such legislation into local policies and guidelines ensures that healthcare staff can set standards to maintain the integrity of the patient's environment. This article will consider aspects of hospital life involved in maintaining a safe environment. PMID- 10711042 TI - Carbon monoxide poisoning and hyperbaric oxygen therapy. AB - This article describes the treatment of carbon monoxide (CO) poisoning with hyperbaric oxygen therapy (HBO). Carbon monoxide poisoning is the commonest cause of fatal poisoning in the UK. Despite this, HBO is an underused treatment modality. Current criteria for hyperbaric treatment include any patient with neurological deficit and any episode of depressed consciousness, cardiovascular disturbance, patients initially treated with surface oxygen and who developed recurrent symptomatology and minor symptoms unresponsive to oxygen. During a 5 year period 82 patients have been treated from a wide geographical area. Of these patients 57% suffered carbon monoxide poisoning as a result of self-poisoning. Other causes of poisoning were: house fire; faulty gas appliances; industrial furnaces; and petrol generators. Of the 82 patients treated, 13 required mechanical ventilation and full haemodynamic monitoring, while the remainder were able to walk in and a few patients received intravenous sedation. In recent years the trend has been to re-treat patients more than once in the first 24 hours to increase efficacy and hopefully decrease the serious sequelae that can occur following CO poisoning. PMID- 10711043 TI - A survey of specialist and advanced nursing practice in the UK. AB - This article is the result of a 1-year research project which aimed to develop a contemporary account of the expectations of senior personnel in NHS trusts throughout the UK with regard to the roles of specialist nurses (SN) and advanced nurses (AN). A questionnaire was sent to the chief nurses in 490 NHS trusts throughout the UK. Findings highlight the distinctions made by senior personnel regarding the roles of SNs and ANs, employment issues, fields of practice, work activities, advantages and disadvantages of employing SNs and ANs and ways in which all of these factors have changed since the last survey in 1996. Differences were highlighted between the roles of SNs and ANs in terms of fields of practice and the strategies used to evaluate the effectiveness of their posts. PMID- 10711044 TI - Running a nurse-led nebulizer clinic in a district general hospital. AB - The widespread home use of nebulizers for delivering bronchodilating medication in patients with a variety of respiratory conditions is well documented, as is the lack of appropriate assessment before this form of treatment is prescribed (Cochrane et al, 1985). When it is considered that the use of nebulizers at home is increasing (Drug Information Bulletin, 1994), it is essential that appropriate assessment is undertaken in line with British Thoracic Society recommendations in order to ensure that the degree of benefit is sufficient to justify the inconvenience, potential dangers and high cost of nebulizer therapy in the home. This article examines a nurse-led nebulizer assessment clinic that was set up as part of a service for providing home nebulization for long-term use in patients with chronic obstructive pulmonary disease or chronic severe asthma. PMID- 10711045 TI - Reasons for prolonged hospital stays following heart surgery. AB - Improvements in cardiac surgery techniques and after care have resulted in a reduction in postoperative stay. Ten years ago the average length of stay following surgery was 13-15 days (Sanchez et al, 1994). Today it is more likely to be 4-7 days (Bemat, 1997). A recent audit provided information on postoperative hospitalization in a cardiac population that was deemed suitable for immediate high dependency care rather than intensive care. The authors carried out retrospective examination of patients' notes in order to detect the possible causes for delayed discharge. The audit was conducted over a 3-month period and information was collected on 210 postoperative cardiac patients. The study population was restricted to all cardiac patients transferred directly to the hospital's 'overnight intensive recovery' unit. These patients are regarded as low- to medium-risk cardiac patients. Thirty seven per cent of the study population experienced a prolonged hospital stay, i.e. greater than 7 days. In the majority of cases the reasons for delayed discharge were non-cardiac in origin. The authors reviewed the literature to identify strategies that may reduce the incidence of preventable complications leading to prolonged hospitalization. They concluded that nurses have a fundamental role to play in reducing the incidence and severity of postoperative complications through patient education, motivation and early identification of potential problems. PMID- 10711046 TI - Hallucinations and delusions. 2: A dual diagnosis case study. AB - This article, the second of two parts, describes how evidence-based psychological interventions were used to help a client suffering from treatment-resistant delusions and substance misuse, allied to a chaotic lifestyle, that had previously presented a substantial challenge to services. The first part (Vol 8(15): 998-1002) Investigated how the neurobiological, social and environmental factors involved in the course and treatment of schizophrenia have helped to establish a pathway to recovery or remission that does not involve pharmacological therapy alone. PMID- 10711047 TI - System 4: the four-layer bandage system from SSL International. AB - The System 4 multi-layer system (SSL International) is a compression regime indicated for the treatment of venous leg ulceration. Compression bandaging has become the main treatment for effective healing rates in this type of leg ulcer. System 4 is simple to use and cost-effective. PMID- 10711048 TI - Confidentiality. 7: Human fertilization and embryology issues. AB - A woman who was having in-vitro fertilization (IVF) treatment was involved in a road accident. Her sister, a nurse, who was with her at the time, knew the drugs which she was taking as part of the IVF treatment and passed this information on to those caring for her in the accident and emergency department. Subsequently, the patient complained that this information should not have been passed on. She did not want anyone else to know that she was receiving fertility treatment. What is the law? PMID- 10711049 TI - The NHS is losing its personal touch with clients. PMID- 10711050 TI - Winnipeg's pediatric cardiac inquest--a nursing perspective. PMID- 10711051 TI - Winnipeg's pediatric cardiac inquest: the ethical issues. PMID- 10711052 TI - What makes your day? A study of the quality of worklife of OR nurses. AB - From data obtained in this study, the weak or negligible influences on the quality of worklife of OR nurses are: Organizational Structure Leadership, and Organizational Learning. Things that matter to OR nurses and that influence their quality of worklife are: Collaborative Decision-Making, Multiskilled Workers, Change, Organizational Culture, Locus of Control; and the most important influence of all- Teamwork. So, now when the question is asked "What Makes Your Day?" The answer is clear--at the end of the day, it all boils down to the most fundamental of all answers--People! People are involved in collaborative decision making, multiskilled workers, change, organizational culture, locus of control, and teamwork. These People include the OR nurse, who is involved in collaborative decision-making and all the other items that affect your daily work environment. The people you collaborate with, the people you interact with in the organization, and the people that make up the team--you and everyone around you are responsible for that magic ingredient--Teamwork. What you say, what you do, and how you behave makes all the difference in the daily worklife of your colleagues. Each and every comment and interaction contributes to the efficiency and effectiveness of the team. Each and every day from this day forward, remember this--remember how important your role is in building a strong and effective team. At the recent AORN Congress, Joan Rivers shared a favourite saying with us: "The past is history, The future a mystery, Today is a gift from God, that is why it is called the present." Today is all that we really have--let us make the best of each and every day as we continue to respect and value each member of the team. Teamwork--our building block of the future--yours, mine, and every other member of the surgical team. Each member can make an enormous contribution--we only need to believe that the "best is yet to be--the best resides in me". Yes, we can do our part in making our workplace a good one. But we can only do so much. It's time for governments, administrators, and managers to examine the work environment, to identify the kinds of things that motivate nurses to get up in the morning and go to work, and what makes that workplace pleasant enough that they are happy to stay there. We are hearing about nursing shortages across Canada and the United States. Recruitment and retention strategies are returning to hospitals. Besides "sign-on" bonuses, it is time for governments and administrators to examine the culture of the workplace. Questions that must be answered: Are nurses included in decisions made? Do nurses have what they need to work with? Are they given a reasonable workload? Are nurses part of a team that values them and their unique contribution to patient care? In the words of Senator Lucie Pepin (1999): "We must turn our anger first, into passion, then into action. A hostile or unpleasant workplace must not be tolerated!" With confidence we must be assertive as we look to improve our work environment. Yes, we can do our part, but now it is time for the other stakeholders to pay attention! PMID- 10711053 TI - Canadian post basic OR education programs. PMID- 10711054 TI - Shouldice Hospital: dedicated to the repair of hernias. AB - The Shouldice Hospital has been dedicated to the repair of hernias for over fifty four years, processing over 250,000 cases in that time. The procedure used has been continually refined over the years, and the recurrence rate is less than one percent (1%). Qualified medical personnel from all over the world visit, frequently to observe our method of hernia repair. PMID- 10711056 TI - Straddling two worlds: what NPs do best. PMID- 10711055 TI - Winnipeg's pediatric cardiac inquest: the patient's and parent's advocate. PMID- 10711057 TI - Care of the patient with chronic pain: Part I. AB - Chronic nonmalignant pain is estimated to affect over 50 million Americans. It frequently results in significant physical, behavioral, psychological, social, and spiritual problems for patients and their families. In spite of its prevalence and consequences, chronic pain is often misunderstood and inadequately managed by healthcare professionals. Advanced practice nurses who are knowledgeable about chronic pain and the complex biopsychosocial-spiritual needs of this patient population serve an important role in recognizing these patients and intervening appropriately in their care. The purpose of this two-part article is to provide that information. Part I outlines the pathophysiology, assessment, biopsychosocial-spiritual aspects, and pharmacological treatment of chronic pain. Part II addresses a variety of nonpharmacologic and self-management interventions one can use in the primary care setting to treat these difficult health problems. PMID- 10711058 TI - Chronic mesenteric ischemia: role of the nurse practitioner in diagnosis and management. AB - Mesenteric ischemia can be a very subtle chronic disease state that may be hard to diagnose. The early symptoms are mild and may be mistaken for other disease entities in the elderly patient with multiple medical problems. Mesenteric ischemia can be fatal secondary to complications such as malnutrition, infection, and compounded comorbid conditions. The risk factors for this problem are the same as for any vascular disease: advanced age, hypertension, smoking, hyperlipidemia, and diabetes. Because of the subtle nature of symptoms, diagnosis may be delayed. The practitioner must be vigilant in evaluating patients at risk and refer these patients to the appropriate vascular specialist. PMID- 10711059 TI - Promotion of foot health in diabetes. AB - Foot ulcers are a complication of diabetes mellitus. Patient education, foot care, and diligent assessment by the practitioner are necessary to ensure adequate foot health. Patients should be instructed to inspect the feet daily for ulcers and signs of infection, and instructed on daily cleansing of the feet with a mild soap and application of moisturizers to the feet. Practitioners should include assessments for neuropathy, peripheral vascular disease, foot deformities, condition of the toenails, and footwear. This proactive approach is important in the prevention of foot ulcers in diabetes mellitus. PMID- 10711060 TI - Human babesiosis: a case study. AB - Babesiosis is an intraerythrocytic parasitic infection caused by protozoa of the genus Babesia and transmitted by the Ixodes dammini tick, which also transmits Lyme disease. Babesiosis is emerging as an illness of public health significance in the United States. Occurrences of Babesiosis infections have been reported during spring, summer, and fall in coastal areas in the northeastern United States. Asymptomatic patients may need only supportive care, whereas asplenic, elderly, and immunocompromised patients are at greatest risk for severe disease. However, overall mortality rates for symptomatic cases are less than 10%. This article presents a case report on a white male in his 70s diagnosed with human Babesiosis and emphasizes the need for early detection and prompt interventions to minimize the sequelae related to this tick-borne disease. PMID- 10711061 TI - Abdominal pain in an adolescent male: a case analysis. PMID- 10711062 TI - The relationship between characteristics of women with mental retardation and outcomes of the gynecologic examination. AB - This study examines the relationships among client characteristics and the outcomes of gynecologic examination success, sedation use, and cooperation with the examination in a sample of 99 women with mental retardation who received services from a nurse-managed women's health clinic in a large county medical center. Client characteristics that were associated with a successful gynecologic examination for this population of women were isolated. The findings from this study suggest that the presence of behavioral problems, profound mental retardation, and expressive language difficulties are important signals to providers. Special approaches are necessary when performing gynecologic examinations for women with mental retardation. PMID- 10711063 TI - Evaluating clinical decision-making skills of nurse practitioner students. AB - The evaluation of clinical competencies of nurse practitioner (NP) students has traditionally been accomplished by direct observation of student-patient interactions. Adult and family NP faculty at the Medical University of South Carolina directly observe students at their clinical site at least twice each semester. Recently, faculty recognized the need for additional validation of clinical decision-making skill development and added the Clinical Competency Evaluation (CCE), a standardized simulated patient encounter, to the evaluation process. The purpose of the CCE is to (a) maximize use of available evaluation technology, (b) evaluate student clinical skills in a controlled, standardized environment using a criterion-referenced format, (c) give students an additional performance feedback mechanism, and (d) identify benchmarks to validate student advancement and completion of the NP program. This article discusses how the CCE process was developed, current methods of conducting and grading the examination, faculty and student evaluation of the outcomes, and recommendations. PMID- 10711064 TI - Evolution of the nursing process. AB - The nursing process, long considered the basic intellectual tool for nursing, has continued to evolve through use by advanced practice nurses (APNs). Traditionally, the nursing process has been comprehensive in guiding the assessment and intervention phases but has left unspecified the cognitive processes involved in clinical decision making. In an effort to compensate for that weakness, nurses in advanced practice have typically integrated medical diagnostic reasoning into the nursing process. The work of Benner and colleagues (1984, 1996) in explicating the unique reasoning process used by APNs has helped further our understanding of clinical decision making in advanced practice. The evolved nursing process, as described in this article, includes the array of clinical decisions that must be made during a patient encounter, including assessment decisions, diagnosis, choice of interventions, and identification of outcomes. This article reviews the nursing and medical literature related to both clinical decision making and diagnostic reasoning, and applies that literature to the evolution of the nursing process. PMID- 10711065 TI - Enhancing your patient communications with e-mail. PMID- 10711066 TI - Profiles of editorial board members: North American neighbors. PMID- 10711067 TI - Recognition of diabetes educators. PMID- 10711068 TI - Management of children with diabetes in the school setting. American Association of Diabetes Educators. PMID- 10711069 TI - Special considerations for the education and management of older adults with diabetes. American Association of Diabetes Educators. PMID- 10711070 TI - Meal planning self-efficacy index for adolescents with diabetes. PMID- 10711071 TI - Comparing outpatient and inpatient diabetes education for newly diagnosed pediatric patients. AB - PURPOSE: The purpose of this study was to compare the efficacy of outpatient vs inpatient programs on medical, cognitive, behavioral, and psychosocial outcomes. METHODS: Using three large, tertiary medical centers in the United States, the sample of 32 children newly diagnosed with diabetes and their parents were recruited. Children and parents who received outpatient education were compared with those who received inpatient education. The following outcome variables were compared: (1) rates of hospital readmissions and/or emergency room visits for either severe hypoglycemia or ketoacidosis, (2) knowledge, (3) sharing of responsibilities, (4) adherence, (5) family functioning, (6) coping, and (7) quality of life. RESULTS: In general, no statistically significant differences were found between the groups. A trend was noted in the outpatient group with regard to improved use of emergency precautions on the adherence measure, roles on the family functioning measure, maintaining family integration on the parental coping measure, and disposition on the children's coping instrument. CONCLUSIONS: Findings support the safety and efficacy of the outpatient program method. PMID- 10711072 TI - Diabetes educators' perspectives on barriers for patients and educators in diabetes education. AB - PURPOSE: This study assessed diabetes educators' perspectives on barriers that potentially affect patient access to and utilization of diabetes education. METHODS: A 40-item questionnaire was developed to collect needs assessment data on diabetes education programs. The questionnaire was mailed to a Washington State professional practice group in diabetes education (N = 143). RESULTS: Most respondents were registered nurses (61%) or registered dietitians (27%); 74% were certified diabetes educators. The educators' perceptions of the difficulty that type 1 versus type 2 patients experience in different areas of self-management after diabetes education underscored the importance of learning effective long term skills for self-care. Some of the reasons given for type 2 patients dropping out of diabetes education programs were also cited as barriers to conducting follow-up, such as education being a low priority for the patient and the patient not being able to afford diabetes care services. CONCLUSIONS: Educators appeared to relate many patient barriers to a lack of patient understanding of the evolving nature of diabetes and the subsequent need for educational support. The role of continuing education for all patients needs to be emphasized to the patient during the initial education, as well as to the healthcare community and the patient's support network. PMID- 10711073 TI - Using a focus group to design a diabetes education program for an African American population. AB - PURPOSE: The purpose of this project was to determine what factors need to be considered in planning a diabetes education program to better meet the needs of African Americans with diabetes in a community served by a suburban community hospital. METHODS: Two focus group sessions were conducted. The sessions were recorded, transcribed, and analyzed by members of the research team. RESULTS: Four themes emerged that had bearing on program development: (1) a sense of personal powerlessness, (2) fear related to complications, (3) recognition of knowledge deficits accompanied by an inability to link behavior to outcomes, and (4) a clear vision of what kind of educational setting would interest and benefit the group. CONCLUSIONS: There were significant differences between what was being offered for diabetes education at the facility and what was desired according to the focus group, including factors of cost and leadership. Recommendations for future program planning are given. PMID- 10711074 TI - A multiethnic study of the predictors of macrosomia. AB - PURPOSE: This study examined predictors of macrosomia in a multiethnic sample of 213 low-income women diagnosed with gestational diabetes mellitus (GDM) after week 24 of their pregnancy. METHODS: Medical records were reviewed retrospectively. Variables examined were mother's height, weight history, educational level, age at diagnosis, weeks at diagnosis and delivery, type of diabetes, mean fasting blood glucose (FBG), and infant's weight, sex, and Apgar scores. RESULTS: Fifty-one percent of babies were macrosomic. Weight gain, nonpregnant weight, weight at delivery, FBG, and Apgar scores at 1 minute were associated with macrosomia, especially in Hispanic women. Logistic regression revealed that nonpregnant weight was the strongest predictor of macrosomia. CONCLUSIONS: Nonobese GDM mothers with optimal weight gain but with high FBG levels > 90 mg/dL may be at risk for macrosomia. The major concerns with obese GDM mothers are nonpregnant weight and high blood glucose levels, in this order. Education for women with GDM should target these risk factors to decrease macrosomia. PMID- 10711075 TI - Use of lispro insulin and quality of life in adolescents on intensive therapy. AB - PURPOSE: This study examined the metabolic and quality-of-life effects of using lispro insulin in teenagers. METHODS: Teenagers on multiple daily injections who had not reached metabolic treatment goals were offered the opportunity to use lispro insulin as part of a larger ongoing study of intensive management in youth. Of the 51 who were eligible, 35 used lispro and were followed for 12 months; the remaining 16 had reached treatment goals, were not offered lispro, and comprised the control group. RESULTS: After 12 months, the teens who received lispro insulin achieved equivalent levels of metabolic control to those achieved by teens in the control group, without differences in total daily dose, insulin regimen, or adverse events. Those who received lispro found coping with diabetes less difficult than those who continued on regular insulin, and they reported less negative impact of diabetes on quality of life and fewer worries about diabetes. Both groups were equally satisfied with their diabetes treatment. CONCLUSIONS: Lispro insulin is a safe alternative for youth on intensive regimens, may assist youth in coping with diabetes, and may improve their quality of life. PMID- 10711076 TI - The use of alternative therapies by diabetes educators. PMID- 10711078 TI - The educator's guide to diabetes resources. American Association of Diabetes Educators. PMID- 10711077 TI - Getting referrals for diabetes education and self-management training. PMID- 10711079 TI - Diabetes education research. PMID- 10711080 TI - Diabetes patient education research: an integrative literature review. AB - PURPOSE: The purpose of this study is to summarize the accumulated state of knowledge in the area of diabetes patient education research and highlight important issues that research has left unanswered. METHODS: An integrative literature review was conducted on the topic of diabetes patient education between the years 1985 and 1998. Keywords used in the computerized search were diabetes mellitus, patient education, health education, research, and behavior change. The databases searched were MEDLINE, CINAHL, HealthSTAR, EMBASE, and CHID HE. A total of 78 papers were reviewed. RESULTS: Most studies lacked a theoretical framework and the majority of studies were conducted in an outpatient setting. HbA1c was the most frequently employed outcome measure, with little, if any, description of the interventions. CONCLUSIONS: Much has been learned in terms of the effectiveness of diabetes education on improving knowledge. However, other topic areas and outcomes need further exploration. PMID- 10711081 TI - Characteristics of the adult learner. AB - PURPOSE: This article provides an overview of adult learner characteristics, with an emphasis on those characteristics studied in diabetes patient education research. METHODS: A selected review of the conceptual and research literature on general adult education and adult learning was conducted, with particular attention to diabetes patient education studies. RESULTS: Characteristics reviewed included learning styles, literacy level, age/aging, ethnicity or culture, gender, and knowledge. Studies of the learning style of group vs individual education indicated some positive benefits for group learning; questions remain about optimal size or periodicity. Studies evaluating the benefits of culturally specific interventions for diabetes management have yielded some information. Characteristics related to gender and age have been studied, but often in pilot or feasibility studies without the power to answer the study questions. CONCLUSIONS: There continue to be many gaps in knowledge related to adult learner characteristics in diabetes education. Lessons from both general adult learning literature and patient education literature from other chronic diseases should be evaluated and incorporated. The complexities of these learner characteristics create challenges in designing studies. However, evidence to support the need for effective educational interventions is of great importance for implementing change in health care. PMID- 10711082 TI - Characteristics of the learner: children and adolescents. AB - PURPOSE: The purposes of this paper are to (1) review the literature on educational interventions for children and adolescents; (2) determine what kinds of interventions have been studied, how effective they are, and their outcomes; and (3) develop recommendations for further research in the field. METHODS: An integrative literature review approach was used. Articles were included in this review if they met the criteria of being an empirical study reporting results of an intervention whose primary subjects were children with type 1 diabetes and/or their families, and published between 1980 and January 1, 1999. Of the 59 articles identified, 41 met these criteria. RESULTS: The majority of the studies focused on adolescents. Results suggested that traditional educational interventions are successful in increasing knowledge but less successful in increasing quality of life or improving metabolic control. Psychosocial and family interventions (coping skills training) have been more successful in both quality-of-life and metabolic outcomes. CONCLUSIONS: There is much work to be done to strengthen our understanding of what works under what conditions. More studies of younger children, minority youth, and families are needed using well controlled experimental designs with adequate samples. PMID- 10711083 TI - Provider impact in diabetes education. What we know, what we would like to know, paradigms for asking. AB - PURPOSE: A literature search from 1985 to the present was performed using Web based search engines to identify evidence-based studies of diabetes education. METHODS: Twelve studies were identified in which a provider characteristic was defined as a discriminate variable associated with impact or efficacy of the education intervention. Provider was defined as all those participating in diabetes care and education (e.g., patient, education and care teams, funding and policy agencies). The Certified Diabetes Educator credential also was described. RESULTS: Specialty or discipline of the provider/care team, technology when used to assist providers, and physician practice patterns have been assessed. There is a paucity of research that assesses provider impact or the specific impact of provider characteristics on diabetes outcomes in a controlled and scientifically rigorous fashion. CONCLUSIONS: Suggestions for evaluating provider impact based on literature concerning psychotherapy and healthcare education outcomes are: identify provider characteristics/attitudes/skills and link them to outcomes, define therapeutic alliance in diabetes care/education and assess its contribution to outcomes; and use performance measures as provider characteristics. PMID- 10711084 TI - Theory is the cart, vision is the horse: reflections on research in diabetes patient education. AB - PURPOSE: In this paper, we examine the nature of vision and the role it plays in helping educators identify and use theories productively. We also discuss the role of theory in diabetes education and provide criteria for selecting appropriate theories. METHODS: The vision of diabetes education developed at the Michigan Diabetes Research and Training Center was used to illustrate how our vision has influenced our use of educational and behavioral theories. RESULTS: Both our vision and our theoretical assumptions should be articulated, discussed, debated, and studied. CONCLUSIONS: Diabetes patient education research can systematically contribute to the development of a sound, coherent, and progressive body of knowledge that will truly serve diabetes patient education. PMID- 10711085 TI - Interventions to promote diabetes self-management: state of the science. AB - PURPOSE: The purpose of this paper is to review the diabetes education literature that has emerged over the past 20 years to determine what we currently know about diabetes self-management interventions and their effectiveness in producing improved health outcomes. METHODS: Findings of studies that were reported prior to 1990 were compared with findings of studies that have been conducted since 1990 to determine what recent changes and patterns in diabetes self-management education have occurred. Future directions in diabetes self-management research and practice were projected from these findings. RESULTS: Most studies lacked adequate descriptions of the interventions tested, which precludes replication or application of the most effective strategies to clinical practice. Trends in interventions have evolved from education only to education plus behavioral models, with more attention given to interventions specifically for minority populations. The interventions that have been designed and tested seem to be longer, with more emphasis on simple, practical approaches to diabetes self management. CONCLUSIONS: The literature supports the effectiveness of diabetes education and behavioral interventions in improving psychosocial and health outcomes. The question of how to best achieve these improved outcomes continues to need further exploration. PMID- 10711086 TI - Behavior change in diabetes education. AB - PURPOSE: The purpose of this paper is to explore how diabetes education produces change in self-care behavior. METHODS: Published research on diabetes education relevant to behavior change was examined and a framework was formulated for the study of behavior change. RESULTS: Research indicates that education improves patient self-management, which in turn improves glycemic control and health status. Yet, there is relatively little information on what types of education produce what particular benefits for which types of patients. Moreover, we do not know the benefits of various forms of education (for selected groups) relative to their costs. Empirical studies of how education produces behavior change are few, but much preliminary work has been done to identify potential behavioral determinants that can be targeted by interventions. Theoretical models of behavior change have been advanced (e.g., stages of change) but they have yet to be rigorously tested. CONCLUSIONS: Initial answers to the research questions can be generated by conducting more sophisticated analyses of the type of data already being collected. However, obtaining complete answers to some of these questions will require more extensive data collection, including large-scale studies of multiple interventions in multiple patient groups. PMID- 10711087 TI - Outcomes of and for diabetes education research. AB - PURPOSE: The purpose of this paper is to review outcome measures used to evaluate diabetes self-management education and make recommendations for future research. METHODS: Three perspectives were used: (1) the frequency with which different measures were collected prior to 1990 was compared with a sample of the 1997 to 1999 literature, (2) a multilevel pyramid model of psychosocial-environmental factors was used to evaluate the level of outcomes assessed, and (3) the RE-AIM evaluation framework was used to assess the public health impact of studies reported in the literature. RESULTS: Knowledge and HbA1c measures are often collected to the exclusion of other, possibly more appropriate outcomes. Research has focused almost exclusively on individual or family level outcomes and paid little attention to effects at systems levels, such as neighborhoods, communities, or healthcare systems. More recent studies have been evaluating the reach of interventions, but more practice-oriented research needs to be conducted with representative patients, providers, and settings. CONCLUSIONS: Much has been learned about the efficacy of diabetes self-management and about measurement issues. Future research should now focus on effectiveness and generalization issues. PMID- 10711088 TI - Congestive heart failure in the elderly. AB - CHF is a complex disease that is complicated by the normal changes that accompany the aging process. As nurses we must understand not only the medical management but also the quality-of-life issues that are critical for these patients and their caregivers. PMID- 10711089 TI - Atrial fibrillation in the older adult. Presentation and management issues. AB - Atrial fibrillation (AF) is a common cardiac arrhythmia in older adults and the most common cause of embolic stroke. Diagnosis of this illness is a challenge in that the older adult may be asymptomatic and/or may present with such atypical symptoms as a change in function, mood, or cognition. This challenge is demonstrated in three vignettes of older individuals who presented with acute onset AF. Nurses need to evaluate older patients for AF so that prompt treatment can be initiated. This proactive approach can have a major impact on preventing strokes and improving the quality of life in these individuals. PMID- 10711090 TI - Fostering hope in the elderly congestive heart failure patient in critical care. AB - This article presents a patient-centered framework applied to the elderly patient with congestive heart failure (CHF). Although the elderly have been the focus of numerous articles, the needs of the elderly CHF patient in the critical care setting, especially concerning hope versus hopelessness, have been neglected. Guidelines for the clinical management of patients experiencing hopelessness are explored. The four dimensions of hopelessness discussed herein are experiential, spiritual/transcendence, irrational, and relational processes. Nursing diagnosis, interventions, goal-setting, and family support also are discussed. Helping the elderly CHF patient maintain hope when confronted with repeated hospitalizations is a challenge for the critical care nurse. PMID- 10711091 TI - A joint effort to affect lives. The COPD Wellness Program. AB - Health care providers can work toward the goal of maintaining quality, dignity, and independence in the elderly when they combine resources with the community to promote wellness and help manage chronic problems. The Chronic Obstructive Pulmonary Disease (COPD) Wellness Program was undertaken for this purpose. The target participants were elderly individuals and their care partners who met the program qualifications: a diagnosis or symptoms of COPD and residency in a certain independent housing facility. The program was designed to help the participants maximize their independence through knowledge and improved management of their disease process. PMID- 10711092 TI - Improving nursing assistant turnover and stability rates in a long-term care facility. AB - In this study, we developed formulas to calculate nursing assistant turnover and stability rates, identified reasons for termination and facility-specific strategies to reduce turnover, and evaluated the effectiveness of implemented strategies. Although turnover remained relatively unchanged (23% in year 1, 28% in year 2), the stability rate remained high (76% in year 1, 75% in year 2). Tracking turnover rates without tracking stability yields an incomplete picture of a facility's efforts to attract and retain qualified employees. Achieving high stability rates in addition to low turnover rates are important goals, and we have included some recommendations. PMID- 10711093 TI - Using bone density measurements in the detection and treatment of osteoporosis. PMID- 10711094 TI - The Geriatric Depression Scale (GDS). PMID- 10711095 TI - Interdisciplinary health care as the link between nursing homes and educational institutions. PMID- 10711096 TI - Ann Schmidt Luggen, PhD, RN, CS, CNAA. A force to reckon with. Interview by Priscilla Ebersole. PMID- 10711097 TI - Update on hypertension management. PMID- 10711098 TI - Teaching strategies for special patient groups encountered in home care. PMID- 10711099 TI - Diet lessons learned from aging hearts. PMID- 10711100 TI - Attending to more than just physical needs. PMID- 10711101 TI - An intervention study to enhance medication compliance in community-dwelling elderly individuals. AB - OBJECTIVE: To determine whether daily videotelephone or regular telephone reminders would increase the proportion of prescribed cardiac medications taken in a sample of elderly individuals who have congestive heart failure (CHF). METHODS: The authors recruited community-dwelling individuals age 65 and older who had the primary or secondary diagnosis of CHF into a randomized controlled trial of reminder calls designed to enhance medication compliance. There were three arms: a control group that received usual care; a group that received regular daily telephone call reminders; and a group that received daily videotelephone call reminders. Compliance was defined as the percent of therapeutic coverage as recorded by Medication Event Monitoring System (MEMS) caps. Subjects were recruited from 2 sources: a large urban home health care agency and a large urban ambulatory clinic of a major teaching hospital. Baseline and post-intervention MOS 36-Item Short-Form Health Survey (SF-36) scores and Minnesota Living with Heart Failure (MLHF) scores were obtained. RESULTS: There was a significant time effect during the course of the study from baseline to post-intervention (F[2,34] = 4.08, p < .05). Over time the elderly individuals who were called, either by telephone or videotelephone, showed enhanced medication compliance relative to the control group. There was a trend, but no significant difference between the two intervention groups. Both SF-36 and MLHF scores improved from baseline to post-intervention for all groups. There was no significant change in the SF-36 scores for the sample, but there was a significant change for the MLHF scores (p < .001). The control group had a significant fall off in the medication compliance rate during the course of the study, dropping from 81% to 57%. CONCLUSIONS: Telephone interventions are effective in enhancing medication compliance and may prove more cost effective than clinic visits or preparation of pre-poured pill boxes in the home. Technologic advances which enable clinicians to monitor and enhance patient medication compliance may reduce costly and distressing hospitalization for elderly individuals with CHF. PMID- 10711102 TI - Spiritual well-being among older adults. AB - 1. Older adults are a highly spiritual and religious group of individuals. 2. Nurses neglect spiritual interventions for older adults because of a fear of imposing their own philosophies, and lack of knowledge about the abstract. 3. Nurses need to be aware of spiritual needs not related to Judeo-Christian concepts. 4. The profound benefits to humankind of religious practice and spiritual well-being among most life forms has been reported. PMID- 10711103 TI - Nursing unit meal management maintenance program. Continuation of safe-swallowing and feeding beyond skilled therapeutic intervention. AB - Feeding, deglutition, and swallowing are the most natural means of alimentation. These means provide nutrition and hydration to the body. Management of dysphagia has become a widespread clinical practice in acute care, rehabilitation, and long term skilled nursing facilities from rehabilitation and nursing perspectives. Caring for chronic dysphagic and feeding-impaired patients on nursing units is a central concern of this article. The purpose of this descriptive article is to report on a practical meal supervision program that may be used on nursing units to facilitate the continuum of dysphagia care after skilled therapeutic intervention. This program stresses the importance of: (a) identifying patient help level and corresponding nursing care; (b) centralized, supervised dining; (c) organized seating assignments; and (d) an effective, integrated communication system. This article addresses staff training and continuous quality improvement monitoring, and discusses potential investigations concerning compliance. PMID- 10711104 TI - Their only privacy is between their sheets. Privacy and the sexuality of elderly nursing home residents. AB - This study which was based on hermeneutic phenomenology, investigated how paid caregivers in two nursing homes experienced older residents' sexuality. Caregivers acknowledged there were obstacles to residents fulfilling their sexual desires. The medical model of care still largely determines the norms and values within nursing homes and governs the actions of staff. Sexuality within the current medical model is not considered to be part of the primary caregiving role because sexuality is not deemed to play a vital role in the maintenance of bodily functions. Because they are geared toward the demands of basic body care, nursing homes are not inclined to create an environment conductive to the fulfillment of sexual needs. PMID- 10711105 TI - Development and implementation of a case management model for long-term care. PMID- 10711106 TI - Care decisions in irreversible dementia: who speaks for the patient? PMID- 10711107 TI - A tribute to our oldest citizens. PMID- 10711108 TI - The Pittsburgh Sleep Quality Index (PSQI). PMID- 10711109 TI - Urban older adults and the forfeiture of a driver's license. AB - Frequently, it is assumed that urban older adults are not vulnerable to the consequences of forfeiting a driver's license because of the availability of public transportation. Although all of the study participants were involved in accidents prior to forfeiting their driver's licenses, 221 (78%) of them stated they had been safe drivers. The majority of participants indicated they had been given no choice by either their families or the bureaucracy in forfeiting their driver's licenses. Regret, loneliness, and immobility were the results of no longer being allowed to drive. PMID- 10711110 TI - Options, outcomes, values, likelihoods decision-making guide for patients and their families. AB - When family caregivers are faced with making daily decisions, they may feel burdened, frustrated, and even in conflict with other family members. The OOVL Guide is a tool to help people think about the various aspects of a decision situation and combine them to make a decision. The OOVL Guide supports nurses in their advocacy role because it affirms the importance of the clients' values and preferences. The OOVL Guide provides decision-makers, both professionals and lay people, with a structure and procedure for making choices for themselves or others. PMID- 10711111 TI - Older women and the meaning of health. AB - Health promotion has become an essential component of comprehensive health care, especially considering the growing female population. Older women, in particular, have demonstrated an increasing need for preventive services or health care maintenance and promotion. These needs consist of services provided by health care professionals that can help women meet their own self-care demands. However, before the health care needs of women can be met, they first must be identified. Using Orem's Self-Care Theory, a three-part qualitative phenomenological study was conducted in an attempt to understand what health actually means to older women and what types of health behaviors they feel are important to maintain or promote health. Three different groups of women age 55 and older with a variety of education, socioeconomic, and racial backgrounds were interviewed. Their responses were analyzed carefully, and each participant was interviewed a second time to validate recurrent themes regarding their statements about health. Results of the interviews with the three different groups of women were surprisingly similar. Although each participant had a unique application of their definition of health, there were five recurrent themes identified by 75% to 80% of the women. The themes were interactions with a being greater than themselves, acceptance of self, humor, flexibility, and being other-centered. Numerous examples supported the five themes. In addition, several "healthy behaviors" also were identified. Based on the findings of this 3-year study, older women seem to have their own perspectives on what health actually means and how they can best monitor or maintain their health. While the list is slightly different than the traditional one prescribed by health care professionals, perhaps it is time to encourage older adults to become more of a partner in their care. These healthy behaviors could be incorporated easily into the treatment and management plans and thereby reflect both parties' definitions of what it means to be healthy. Health care professionals must continue to ask older adults about their view of health and encourage their partnership in health-related issues. All individuals, no matter what age, are being encouraged to assume more responsibility for their own personal health, and by including older adults in the decision-making process, this partnership can be strengthened. PMID- 10711112 TI - The world it is a-aging. PMID- 10711113 TI - Caring for elderly individuals in nursing homes. PMID- 10711114 TI - What adaptations in nursing care do you make for your oldest old clients? PMID- 10711115 TI - Behavioral vs drug treatment for urge urinary incontinence in older women: a randomized controlled trial. PMID- 10711116 TI - Prevalent urinary incontinence as a correlate of pregnancy, vaginal childbirth and obstetric techniques. PMID- 10711117 TI - International Year of Older Persons. PMID- 10711118 TI - A looming crisis for the written journal? PMID- 10711119 TI - How to get started or "what is the research question?". PMID- 10711120 TI - Description of pressure ulcer pain at rest and at dressing change. AB - PURPOSE: To describe and compare the pain experienced by patients with stage II, III, and IV pressure ulcers both at rest and during dressing change. PATIENTS AND SETTING: Thirty-two subjects were recruited from acute, extended, and home care settings in the Midwest. The sample population ranged in age from 47 to 95 years (mean age 74.7 years, SD = 12.8), were white, and evenly divided between male and female patients. METHODS: Patients used the McGill Pain Questionnaire to rate their pressure ulcer pain at rest and again at dressing change. RESULTS: Twenty eight subjects (87.5%) reported pain at dressing change and 27 (84.4%) experienced pain at rest, compared with 4 (12.5%) subjects who reported no pressure ulcer-related pain. Of the 28 reporting pain, 21 (75%) rated their pain as mild, discomforting, or distressing, and 5 (18%) described their pain as horrible or excruciating. Twelve (42%) reported their pain as continuous, occurring both at rest and during dressing changes. Despite these reports, only 2 (6%) subjects had received medication for their pressure ulcer pain. Although none of the hypotheses were statistically significant, we observed that patients with stage II, III, and IV pressure ulcers experienced pain. CONCLUSIONS: Within this study sample, the majority of patients had pressure ulcer-related pain, which for some was severe and constant. Therefore we conclude that the potential for pressure ulcer-related pain should be anticipated and assessed on a regular basis. In addition, further research is needed to increase our understanding of pressure ulcer pain and to find effective interventions for its relief. PMID- 10711121 TI - Prevalence, incidence, and prediction of pressure ulcers on a rehabilitation unit. AB - OBJECTIVE: This retrospective chart review study was conducted to determine the prevalence and incidence of pressure ulcers, and the contribution of known risk factors toward the predicted occurrence of pressure ulcers in a long-term rehabilitation setting. SUBJECTS AND SETTING: A continuous series of 170 adult men with a mean age of 69.2 years were studied during a 1-year period. METHODS: Patient charts were reviewed retrospectively for risk factors and documentation of pressure ulcer development by 1 researcher on a data recording form. RESULTS: The pressure ulcer prevalence was 12% and the incidence over the 1-year observation period 6%. Using the odds ratio test, significant risk factors in the sample were identified as hypoalbuminemia (odds ratio = 11:1), low diastolic blood pressure (odds ratio = 4.6:1), stool and urine incontinence (odds ratio = 1.5:1), and peripheral edema (odds ratio = 3.5:1). CONCLUSION: Specific characteristics in this sample of patients in a long-term rehabilitation center contributed to the increased risk for pressure ulcer development. Risk assessment based on knowledge of specific risk factors, prevention, and early intervention is crucial to lowering the prevalence and incidence of pressure ulcers in this setting. PMID- 10711122 TI - Intraoperatively acquired pressure ulcer prevalence: a national study. AB - OBJECTIVE: This study was completed to determine the prevalence and identify comorbid conditions for intraoperatively acquired pressure ulcers. DESIGN: A multisite, descriptive study was conducted. SETTING AND SUBJECTS: A multisite sample population of patients from 33 of 50 states undergoing a surgical procedure of at least 3 hours' duration was studied. INSTRUMENTS: A Hospital Background Data Form and a Patient Data Form were constructed to collect demographic data and information pertinent to surgically acquired pressure ulcerations. The Weighted Index Comorbidity Scale was incorporated into the Patient Data Form. METHODS: Institutional and patient data forms were mailed to 1543 members of WOCN who practiced in an acute care facility. Each member was asked to collect data on those patients who had surgeries of 3 hours or longer during a period of 1 week. MAIN OUTCOME MEASURES: Patient and facility characteristics, visual observations of the patient's skin over a 72-hour period after surgery, and the Weighted Index Comorbidity Scale were used to determine the prevalence of surgically acquired pressure ulcers and the presence of relevant comorbid conditions. RESULTS: An analysis of 104 returned facility surveys including 1128 patients was completed. The prevalence of pressure ulceration among this group was 8.5%. Forty percent of those surveyed underwent a procedure lasting approximately 3 hours and 33% underwent surgery lasting more than 5 hours. As the length of surgery increased, so did the percentage of patients with pressure ulcers. Most patients had at least 1 comorbid condition (78%). CONCLUSIONS: The risk of intraoperative ulcerations increases as surgical time increases. Although patients with comorbid conditions known to affect the risk of ulceration under normal circumstances experienced pressure ulcers in this investigation, no significant relationship was found to link the presence of these conditions to an increased risk of intraoperatively acquired ulcers. Therefore all surgical patients undergoing prolonged procedures should be considered at risk for intraoperative ulceration. PMID- 10711123 TI - Comparison of subjective classification of stool consistency and stool water content. AB - OBJECTIVE: The purpose of this study was to compare the stool consistency categorizations made by 39 adults with fecal incontinence and the percentage of water in their stools determined by lyophilization. METHODS: Subjects collected all stools daily for 8 days during a baseline period and at the end of a fiber treatment period. Stool consistency was recorded as hard and formed, soft but formed, loose and unformed, or liquid. Aliquots of the stools were lyophilized to constant weight. MAIN OUTCOME MEASURES: The main outcome measures were the percentage of stool water among stools in each consistency category and the correlation between subjects' stool consistency categorizations and the percentage of stool water. RESULTS: The subjects were 8 men and 21 women, ranging in age from 30 to 89 years, who were participating in a study of the effectiveness of dietary fiber for treating fecal incontinence. A total of 1023 stool samples were analyzed. Significant differences in the mean percentage of water were found among the 4 stool consistency categories (hard and formed = 68% +/- 0.9%, soft but formed = 74% +/- 0.3%, loose and unformed = 80% +/- 0.4%, and liquid = 85% +/- 0.3%; P < .001). Ninety-six percent of the stools had a percentage of water within 2 SDs of the mean percentage of water of other stools in their consistency category. CONCLUSION: This classification system of stool consistency is a valid and practical measure for clinical studies. It may be useful for clinicians and patients to evaluate outcomes of treatments directed at improving stool consistency. PMID- 10711124 TI - Nursing interventions for urinary incontinence in home health. AB - Urinary incontinence (UI) is a significant health care issue among older patients in the acute care, extended care, and home care settings. With the shift toward health care delivery in the home setting, it is becoming increasingly necessary for home health care professionals to become knowledgeable about the causes, assessment, and treatment. This article will review the epidemiology, causes, assessment, and management of UI in the home care setting. Emphasis is placed on the role of the WOC nurse as coordinator of a multidisciplinary team providing care of the home bound patient with UI. PMID- 10711125 TI - Calciphylaxis in a patient with end-stage renal disease. PMID- 10711126 TI - Treatment of migraine headache. PMID- 10711127 TI - Preventing and recognizing drug eruptions of the skin. PMID- 10711128 TI - Cholesterol and your health. PMID- 10711129 TI - Streamlining drug therapy. PMID- 10711130 TI - Outpatient management of patients on warfarin. AB - The recent publication of the American College of Chest Physicians (ACCP) Fifth Consensus Conference on Antithrombotic Therapy provides the primary care clinician with a thorough guideline for the treatment and prevention of thromboembolic diseases. These recommendations are reviewed in this article as they relate to the outpatient management of patients receiving warfarin. Common drug-drug, drug-food, and drug-disease interactions also are reviewed along with practical dosing recommendations. PMID- 10711131 TI - Dietary supplements used in the treatment of depression, anxiety, and sleep disorders. AB - Dietary supplement use has increased during the past decade. Epidemiologic studies suggest that patients turn to dietary supplements because of a reluctance to take prescription medications or a lack of satisfaction with the results. They often perceive dietary supplements to be a safer or more natural alternative. Patients with mental health conditions, including depression, anxiety, and sleep disorders, are among those who use dietary supplements. St. John's Wort is used to treat depression. Clinical studies comparing dietary supplements with low-dose antidepressants (maprotiline, amitriptyline, or imipramine at 75 mg/day) or high dose antidepressants (imipramine at 150 mg/day) find no significant difference between treatments. Kava kava is used to treat anxiety. Clinical trials demonstrate it to be superior to placebo, and roughly equivalent to oxazepam 15 mg/day or bromazepam 9 mg/day. Agents discussed for use in sleep disorders include melatonin, valerian, 5-hydroxytryptamine, catnip, chamomile, gotu kola, hops, L-tryptophan, lavender, passionflower, skullcap, and valerian. Familiarity with the evidence for use and the possible resulting risks can help health professionals to guide patient decisions regarding use of dietary supplements. PMID- 10711132 TI - A review of NSAID complications: gastrointestinal and more. AB - Nonsteroidal anti-inflammatory drugs (NSAIDs) are among the most widely prescribed medications in the world. Their use is associated with significant morbidity, commonly including gastrointestinal, renal, and hepatic effects. Other less common effects include otic, central nervous system (CNS), ophthalmic, allergic, and dermatologic effects. Recognizing and diagnosing potential complications are imperative to obtain favorable outcomes in patients on NSAIDs. Among the NSAIDs currently on the market, there exists a wide variability of incidence of organ-specific side effects. Some of these side effects are class specific, but most are not. Much research also is being performed in the field of COX-2-specific inhibitory agents, which would theoretically reduce the gastrointestinal insult of these drugs. Patient awareness through education is crucial in decreasing morbidity and preventing the adverse effects of these drugs. PMID- 10711133 TI - Clinical and pharmacologic management of chronic heart failure associated with left ventricular systolic dysfunction. AB - Despite many recent advances, heart failure continues to be a major cause of morbidity and mortality in the United States. Once a patient is identified and evaluated as having heart failure, nonpharmacologic therapies such as a dietary restrictions, lifestyle changes, exercise strategies, and patient education are initiated as well as pharmacologic therapy. Angiotensin-converting-enzyme inhibitors, diuretics, and digoxin all are considered traditional therapy in the management of chronic heart failure associated with left ventricular systolic dysfunction. The role of newer therapies such as beta-blockers and angiotensin II receptor antagonists in the treatment of heart failure is in the process of being defined. Furthermore, new data is appearing concerning calcium channel blockers, spironolactone, and combinations of the various agents. Appropriate use of the traditional agents accompanied by a through understanding of the newer therapies and their roles is essential to the continued reduction of the morbidity and mortality associated with heart failure. PMID- 10711134 TI - Therapeutic issues in oral glucocorticoid use. AB - The therapeutic use of glucocorticoids are extensive despite numerous adverse effects related to their use. The fear of hypothalamic-pituitary-adrenal (HPA) axis suppression and adrenal crisis often prolongs glucocorticoid therapy unnecessarily. Glucocorticoids are the most common cause of drug-induced osteoporosis and cataract formation. Other effects of concern to the primary care provider include increased blood pressure, blood glucose, and cholesterol. Glucocorticoid therapy also may lead to avascular necrosis, growth retardation, and myopathy. Practical dosing strategies such as alternate-day therapy reduce the risk of developing most adverse effects. Several tapering regimens exist that lessen the potential for steroid withdrawal syndrome. Identifying common glucocorticoid drug interactions averts drug toxicity. By appropriately monitoring patients on long-term glucocorticoid therapy, adverse outcomes are minimized. PMID- 10711135 TI - Hepatitis C infection: a review. AB - Hepatitis C, transmitted through body fluid exchange, affects approximately 1.8% of the U.S. population, roughly 3.9 million persons. Transfusion of blood and blood products was once an important source of hepatitis C transmission. Since the initiation of the hepatitis C screening program in 1985, however, injection drug use has become a major route. Hepatitis C is a leading cause of chronic liver disease. In 80% to 85% of those infected with the virus, chronic hepatitis C eventually develops, which can lead to cirrhosis and hepatocellular carcinoma, with alcohol abuse and coinfection with hepatitis B as additional risk factors. Screening for hepatitis C can be achieved with serologic assays. Molecular assays are helpful in confirming the diagnosis, assessing viral load, and characterizing the genetic nature of the viruses. Interferon alpha (IFN-alpha) and a combination of IFN-alpha 2B and ribavirin are therapies available in treatment of hepatitis C, but sustained response to the treatment has been unsatisfactory. Further studies are indicated to obtain more effective therapies for eradication of the disease. Hepatitis C is preventable, and clinicians should use every opportunity possible in their practice to assess those at risk and actively engage them in risk factor reductions. PMID- 10711136 TI - HMOs will be rewarded for taking sick patients. PMID- 10711137 TI - A lexicon of the financial environment. PMID- 10711138 TI - The history and evolution of hospital payment systems: how did we get here? AB - After many years of testing and experimentation, most insurance companies, managed care plans, and self-insured employers now have effective programs in place to manage their health care expenditures. More important, the recent efforts to balance the federal budget have led to a series of changes in the payment systems for both Medicare and Medicaid that are designed to reduce the amount of money the federal government will spend on health care services in the future. It appears that all the loopholes have been closed. Many hospitals, as well as other types of providers, are now projecting flat revenue growth, while some are actually anticipating declines in revenues. To fully understand health care finance and be effective in reducing costs, hospital managers need to understand the history and evolution of the payment system and how they have influenced both cost measurement and cost control efforts. PMID- 10711139 TI - Financial and operational skills for nurse managers. AB - The nurse manager role has changed from managing one inpatient unit to responsibility for multiple units, more employees, larger operating budgets, and clinical services outside the traditional nursing inpatient setting. The increasing complexity of the role requires the manager to function at a higher level of financial and operational performance. The article describes a leadership development project that utilized "partnerships" to coach managers in financial and operational management skills. It describes the analytic tools used to evaluate unit performance and shows how the utilization of these tools provides managers with the data they need to develop appropriate solutions to problems on their units. PMID- 10711141 TI - Managing nursing assets: a primer on maximizing investment in people. AB - Nurse managers require critical competencies and skills to thrive in today's turbulent health care environment. A nurse manager's relationship with nursing staff is the primary variable that can reduce nursing turnover and improve unit morale in a variety of measurable ways. Self-confidence is the cornerstone competency required for the nurse manager's success. Nurse managers set the context for the delivery of patient services. To do so effectively, a working partnership with finance and human resources is required. Both functions can provide essential information for real time decision making, including variance analysis, flexible budgets, absenteeism rating, turnover rate, and innovative compensation programs. PMID- 10711140 TI - Transformational management style positively affects financial outcomes. AB - Two specific examples from the author's experience demonstrate the central theme that positive financial outcomes are a function of a transformational leadership style of management. The article focuses on participation competencies utilizing involvement, empowerment, and accountability. Developing staff and empowering them to make decisions about their work and outcomes are necessary to achieve a high-quality, cost-effective outcome, the key to financial success. PMID- 10711142 TI - Understanding and championing the merger process: key leadership roles for successful outcomes. AB - Mergers can effectively achieve both financial results and enhancements of quality. The nurse executive plays a vital role in the success of merger activities and has the opportunity to emerge as one of the leaders of the process. A systems approach to cost, quality, and service must be captured for successful merger outcomes to occur. But, in order for the nurse executive to be effective, he or she must understand and champion the merger process, critical roles, and challenges during the merger as well as use efficient communication methods to work effectively with physicians, reporting staff, and hired consultants. PMID- 10711143 TI - The managed care contract: survival or closure? AB - The managed health care contract can be considered the most powerful tool in the health care environment today. Providers of care as well as insurers need to fully comprehend the legal and financial impact of these contracts. Too many organizations (providers and insurers) are getting caught in financial failure, resulting in bottom line-driven health care. It seems that the days of providing the most ethically appropriate health care are gone. Too much emphasis has been put on providers and administrators to provide the care that will result in positive income. The only way to "protect" oneself is in the creation, negotiation, and administration of the managed health care contract. PMID- 10711144 TI - Career development for nurses in today's health care environment and the value of nontraditional roles. AB - Roles that were essential in the health care setting of the past are rapidly disappearing in today's era of redesign. Career development and advancement are tumultuous processes in the current health care environment. The article illustrates what nontraditional roles can offer nurses through a case study on the contract negotiation process. It is hoped that the article will encourage nurses to take risk in choosing career experiences. Such choices will facilitate the learning that is necessary to lead the complex health care organizations of today. PMID- 10711145 TI - Strategic decision making on shifting sands. AB - Strategic decision making in today's health care marketplace is very challenging. In the past, strategic plans were written for five to ten years. With the rapidly changing environment, planning for more than two years is difficult. It is critical for organizations to set a pathway for success. However, the organization needs to be able to respond quickly to environmental changes and take advantage of opportunities. The article presents a case study on a relatively new organization that has ambitious strategic goals. It discusses the decision-making process on the ways to enter a new marketplace, including the key factors assessed by the organization and their implications. PMID- 10711146 TI - Financial management for entrepreneurs. AB - Thousands of nurses are leaving corporate life and the bedside for the world of entrepreneurship. It is a healthy sign for the profession. Being in business is one of the best ways to learn about ultimate risk-taking, an experience woefully lacking in the world of hospital- or corporate-based nursing. There seems to be no way to soften the ultimate risk of embarking on a business, although each person can certainly learn what worked for others and what to avoid. In this article, I outline some of the key things that worked and did not work for me during my 20-year stint as a nurse entrepreneur. PMID- 10711147 TI - Multidisciplinary collaboration: a method for measurement. PMID- 10711148 TI - An important theme for the financial environment. PMID- 10711149 TI - Nursing information systems: who needs them? AB - This article examines some of the reasons why--both financial and perceptual- organizations are becoming skeptical of nursing information systems. Then it delineates some key benefits and explains how nursing can and should be prepared to head off such skepticism. PMID- 10711150 TI - I didn't think I'd live this long. PMID- 10711151 TI - Leadership effectiveness: how do you measure up? AB - The purpose of this article is to provide leaders with a tool to collect accurate data of subordinate's perceptions. Leaders in the nursing profession must feel comfortable and be encouraged to seek the opinion of the staff they serve. Without actively seeking the feedback of subordinates there is no opportunity for personal growth and insight. So, while asking for staff feedback can be a "daring adventure" for any leader, leading staff who do not want to follow will result in an organization of "nothing" in today's health care arena. PMID- 10711152 TI - Grants management skills keep funded projects on target. AB - An often neglected but equally relevant aspect of program development and nursing research is the management of funded projects from the time an idea is generated to the final submission of project report. When effective grants management skills are used to oversee the development of a program proposal and the oversight of the funded project, the likelihood for successful outcome is increased. In this article several steps have been described in the grants management process that can be used to assure that projects are developed and proceed according to plan. PMID- 10711153 TI - Cost analysis of emergency room use by low-income patients. PMID- 10711154 TI - Constant observation in medical-surgical settings: a multihospital study. PMID- 10711155 TI - Handling office politics. PMID- 10711156 TI - The development of a shared governance model in the ambulatory setting. AB - Many individuals within the ambulatory division have benefited both personally and professionally as a result of this shared governance model and its concepts. Staff who rarely volunteered for projects became involved, and others developed their leadership skills. Although the ambulatory division's shared governance model did not reach maturity, many tasks were accomplished through collaborative efforts. As a result, there were many positive staff outcomes such as a greater understanding of their workplace, and a greater appreciation of their peers. PMID- 10711157 TI - Integrated delivery systems: mergers and acquisitions. AB - Mergers and acquisitions are usually the way an IDS is built. The CNO and/or CNOs/DONs have an integral role in the resolution of the M/A process. During this time of significant change, during which there may even be chaos, the CNOs work to maintain stability so there is as little impact as possible on patient outcomes, a core responsibility of the CNOs. The CNOs should focus on identifying and working with the highly skilled individuals in the organization to get to the recovery stage of the M/A process, at which time a high-performing organization is achieved. To build this new organization or IDS, the old organizations of the M/A must be changed (Moss Kanter, 1994). The successful CNOs will manage the trade-offs and will become experts in collaboration. The CNO's goals are to maximize the quality of patient care, the professional satisfaction of the nurse, and the goals of achieving cost effectiveness for the system (Clifford, 1998), and keeping this focus through the M/A process will yield success. PMID- 10711158 TI - Taking your performance measurements? Information technology can help. PMID- 10711159 TI - Political activities for rainy days. PMID- 10711160 TI - The art of leading with grace. AB - Uncivilized environments take their toll on people who work there. Cultures of distrust are created and there is no warmth, reverence, or love available for the healing work of health care. We can blame the staff, or we can look at ourselves and recognize that the staff is merely a reflection of ourselves. Gracious leaders create a gracious and loving staff who care for patients and their families in extraordinary ways. PMID- 10711161 TI - Preparing tomorrow's nurses for the no margin-no mission world of health care. PMID- 10711162 TI - The use of unlicensed assistive personnel and selected outcome indications. AB - This pilot study examine the satisfaction levels of patients, RNs, and UAPs after implementing a patient care delivery system using UAPs as nurse extenders on a 41 bed short-stay medical-surgical observation unit. It also compared patient fall statistics prior to and after implementing the new care model. The convenience study sample included 40 patients, 15 RNs, and 9 UAPs and covered 2 months of unit experience in mid 1997. Risk management statistics from a comparable period 2 years prior showed no significant difference in the number of patient falls. Patient satisfaction scores on five of the seven regularly collected questions was higher in 1997 than the comparable earlier period. RN satisfaction with the care model using UAPs was above a neutral score of 3 on a 5-point Likert scale on two out of three items, but additional attention to the RN's role in effective delegation to UAPs was needed. PMID- 10711163 TI - Self-scheduling with Microsoft Excel. AB - Excessive time was being spent by the emergency department (ED) staff, head nurse, and unit secretary on a complex 6-week manual self-scheduling system. This issue, plus inevitable errors and staff dissatisfaction, resulted in a manager lead initiative to automate elements of the scheduling process using Microsoft Excel. The implementation of this initiative included: common coding of all 8 hour and 12-hour shifts, with each 4-hour period represented by a cell; the creation of a 6-week master schedule using the "count-if" function of Excel based on current staffing guidelines; staff time-off requests then entered by the department secretary; the head nurse, with staff input, then fine-tuned the schedule to provide even unit coverage. Outcomes of these changes included an increase in staff satisfaction, time saved by the head nurse, and staff work time saved because there was less arguing about the schedule. Ultimately, the automated self-scheduling method was expanded to the entire 700-bed hospital. PMID- 10711165 TI - Marketing service guarantees for health care. AB - The author introduces the concept of service guarantees for application in health care and differentiates between explicit, implicit, and conditional vs. unconditional types of guarantees. An example of an unconditional guarantee of satisfaction is provided by the hospitality industry. Firms conveying an implicit guarantee are those with outstanding reputations for products such as luxury automobiles, or ultimate customer service, like Nordstrom. Federal Express and Domino's Pizza offer explicit guarantees of on-time delivery. Taking this concept into efforts to improve health care delivery involves a number of caveats. Customers invited to use exceptional service cards may use these to record either satisfaction or dissatisfaction. The cards need to provide enough specific information about issues so that "immediate action could be taken to improve processes." Front-line employees should be empowered to respond to complaints in a meaningful way to resolve the problem before the client leaves the premises. PMID- 10711164 TI - Analysis of chronic emergency department use. AB - This retrospective emergency department (ED) chart review study examined the relationship between acuity level and the type of insurance in a patient population who used the ED on a chronic basis (seven or more times in the calendar year 1996). Of 1,185 patients seen in the ED in 1996, 122 had between 7 and 29 visits. In the study population: 62.5% of their visits were classified as nonurgent; 42.6% of the nonurgent visits were made by those with insurance; the highest frequency of visits took place between 8:00 am and 4:00 pm when most alternative nonemergency facilities are open. Some of the factors seen as influencing overuse of the ED include: the fact that some chronic use appeared to be associated with psychiatric conditions including substance abuse and patients with recurrent chest pain; and patient's perceptions that access to lab and X-ray facilities are readily available within the ED. PMID- 10711166 TI - Integrated delivery systems: the chief nursing officer. PMID- 10711167 TI - Legislating patient privacy. PMID- 10711168 TI - Adding value by expanding RN roles in ambulatory care. PMID- 10711169 TI - Using administrative data for practice and management. PMID- 10711170 TI - The culture of courage. AB - We all want health care to be delivered in an atmosphere of utmost integrity. However, integrity only occurs where there is courage to do the right thing in very difficult situations. Leaders must be very courageous to be successful. They must foster and build courage among their staff to create cultures in which the staff feel safe and supported to always act with integrity. We are not born with courage. We learn courage by eventually mastering situations in which we can act with less and less fear. The leader's obligation is to assess the environment and build opportunities for staff to work courageously with less and less fear as they master more difficult situations in their everyday lives. PMID- 10711171 TI - Oh Lord, won't you buy me a Mercedes Benz. PMID- 10711172 TI - The silent health care revolution: the rising demand for complementary medicine. PMID- 10711173 TI - Nurse education: is it responding to the forces of supply and demand? PMID- 10711174 TI - A tale of two nursing centers: a cautionary study of profitability. PMID- 10711175 TI - Hospital-acquired pressure ulcers: a comparison of costs in medical vs. surgical patients. PMID- 10711176 TI - Making good oral presentations. PMID- 10711177 TI - The image of a nurse--myth vs. reality. PMID- 10711178 TI - Preventing adverse drug reactions through automation. PMID- 10711179 TI - Balanced Budget Act of 1997 impact on a nurse-managed academic nursing practice for frail elders. PMID- 10711180 TI - Delays in patient transfer: postanesthesia care nursing. PMID- 10711181 TI - The past as prologue. PMID- 10711182 TI - Partnership economics: nursing's challenge in a quantum age. AB - While the task of ascribing economic value to human caring is incredibly complex, Eisler (1998) reminds us that economic systems can and do change--and all our economic institutions are human creations. Humans can create new economic rules that recognize caring as the most foundational economically productive work (Eisler, 1998). In selected situations, the work of nursing has enormous economic value and new research continues to emerge which support our intuitive beliefs about the economic value of nursing's work (Levinson, Roter, & Frankel, 1997; Revans, 1964; Salmond, 1972). The future of health care requires that this work continues. The value of developing conscious economic inventions for nursing has the potential to assist nursing in its cost-effectiveness analysis, document the economic value of caring behaviors, inspire recruitment, and decrease the number of nurses leaving the profession. Nursing is challenged to shift its consciousness to one that is based on the value equation of quality, cost, and service. We are challenged to recognize the need to build a more equitable and humane economic system that gives real value to the work of caring, and to partner with other health care leaders, policy analysts, and economists in the process. Ultimately, developing economic measurements for the redefined work in nursing will require significant changes in economic measurements, institutions, rules as well as social norms, customs, and laws to quantify those goods not traditionally valued. The focus and energy of the nursing profession must move beyond overwhelming notions of loss and change; to challenging leadership to actively write and produce a better script for health in America beyond what we have developed over the past century. Indeed, this work is the requisite for the profession if what our children receive is the next level of enhancement from that which this generation obtained. It is no longer an option in creating a preferred future, it is nursing's mandate. PMID- 10711183 TI - Staff nurse empowerment and collective accountability: effect on perceived productivity and self-rated work effectiveness. PMID- 10711185 TI - International credentialing and immigration of nurses: CGFNS. PMID- 10711184 TI - Caregiver-patient ratio: capturing census and staffing variability. PMID- 10711186 TI - Evaluating cost center productivity. AB - The monthly and yearly productivity summaries were developed and applied to a computer spreadsheet to aid the nurse manager in better understanding and communicating budget issues for diverse ambulatory care departments. A computerized spreadsheet using a commercially available personal computer program, such as Lotus, Quattro Pro, or Excel, can be used to more quickly and accurately track and summarize monthly budget reports. The data can be entered into the spreadsheet either manually or imported by query from the financial mainframe system. Contact your agency's finance or information department for information on how to accomplish this. Periodically acuity and resources should be measured and compared with quality monitors to maintain standards. For the past 10 years, our facility has successfully used this tool to make more informed decisions by identifying trouble spots early, and taking corrective action to avoid crisis management. PMID- 10711187 TI - The new millennium and leadership: evolution or entropy? AB - Health care in the new millennium will be different. We have a wonderful opportunity to live in a new paradigm of leadership and management that opens up myriad possibilities for innovations in health care and leadership. If leaders stand still and don't quickly adapt to the new demands, they will be killed as leaders in this new millennium. The next 20 years are only for those who can act and adapt quickly. PMID- 10711188 TI - Post-millennium breather nowhere in sight. PMID- 10711189 TI - Integrated delivery systems: sharing resources. PMID- 10711190 TI - What is the issue?: pseudoaddiction or undertreatment of pain. PMID- 10711191 TI - Consumer Bill of Rights legislation: a study in controversy. PMID- 10711192 TI - Advancing safety. PMID- 10711193 TI - The "boys in the back room": nurses are stuck with a bad deal. PMID- 10711194 TI - The need for universal health care is now. PMID- 10711195 TI - All aboard--riding on the fast track. PMID- 10711196 TI - The RN in independent practice you! PMID- 10711197 TI - Gender traps in today's work place. PMID- 10711198 TI - Visionary nursing empowers nurses. PMID- 10711199 TI - Student perceptions of empowerment in their graduate program. AB - Empowerment is a critical aspect of the education of the Advanced Practice Nurse (APN). This paper reports a study which asked for feedback from APN students about their experience of feelings of empowerment and powerlessness in their graduate program. Learning which was driven by the student's interest, which respected and valued their experience and which guided them in defining and pursuing individual goals was empowering. Powerlessness was experienced when faculty controlled the process, were demeaning about students' abilities and when learning was not connected to their goals. Advanced Practice Nurses (APN) are in great demand for the emerging health care system. It is critical that students graduate well prepared to face the challenge of today's health care environment. An overarching objective of advanced practice is to enable patients to take control and be responsible for their own health needs, to empower them to care for themselves. The connection between the empowerment of APNs and their ability to empower their patients is unclear. The relationship between the way APN students are treated and how they treat their patients when they graduate is also not known. This investigation looked at what elicited feelings of powerlessness and empowerment in students as a first step in understanding the process. This paper describes a pilot study which elicited student experiences in their graduate program which engendered feelings of empowerment or powerlessness. The study was a replication of Chandler's previous work with staff nurses. It is hoped that this information will provide a framework for a knowledge base to develop educational strategies which will empower the APN student. PMID- 10711200 TI - Has society in the USA really given informed consent to managed health care? AB - It is the assertion of this paper that the current environment of applied managed care in the United States amounts in effect to bio-medical experimentation with humans as the subjects. The ethical dilemma of using humans as subjects of experimentation with unsure outcomes has been addressed historically. Perhaps the most famous attempt at delineating permissible behavior is the Nuremberg code of 1947. While stopping short of the logical extreme--accusing the managed care drivers of being war criminals--this text illustrates the unsatisfactory nature of the current health-care status quo, and proposes alternative means by which economic savings may be realized for the benefit of society, without violating the social obligation to provide health-care. Among the strongest proposals is that ethical or moral constraints may need the protection of regulation (legislation and possibly federal regulation) by and for the common good of the United States' society as a whole. PMID- 10711201 TI - How to protect yourself against malpractice. PMID- 10711202 TI - The Woodhull Study on nursing and the media: health care's invisible partner. PMID- 10711203 TI - The view askew. PMID- 10711204 TI - 24 hour visitation. PMID- 10711205 TI - Success secrets ... 7 golden rules for working on a shoestring budget. PMID- 10711206 TI - It's still who you know. PMID- 10711207 TI - How managed care effects current labor law: the case of nurses and physicians. PMID- 10711208 TI - Differentiated practice--the recycled professional vs technical debate. PMID- 10711209 TI - Going from novice to expert. PMID- 10711210 TI - Managing malpractice insurance. PMID- 10711211 TI - Creating confidence in the midst of chaos. PMID- 10711212 TI - Taking care of you and not me! PMID- 10711213 TI - We're not in Kansas anymore: shaping a new and better health care system. PMID- 10711214 TI - Therapeutic touch. A retrospective on therapeutic touch. PMID- 10711215 TI - Telling our stories: what are nurses angry about? PMID- 10711216 TI - To kill a nurse. PMID- 10711218 TI - Oppression--how nurses can overcome it. PMID- 10711217 TI - Pulling from Peter to save Paul: is "floating" administratively or professionally sound? PMID- 10711219 TI - Burnout or exploitation? PMID- 10711221 TI - Capitol connections: a network in action. PMID- 10711220 TI - Market driven health care & its impact on nurses. PMID- 10711222 TI - The nurse entrepreneur redefined. Are you one? PMID- 10711223 TI - Success secrets ... 5 ways to weather a cash flow drought--and still pay your bills. PMID- 10711224 TI - Financial planning for nurse entrepreneurs; "the most frequently asked question". Interview by Carolyn Zagury. PMID- 10711225 TI - RNs in commercials: do you have what it takes? PMID- 10711226 TI - Welcome to the independent practice of the profession of nursing! PMID- 10711227 TI - Winning profits with Guerrilla Marketing. How science is helping marketing. PMID- 10711228 TI - Biological Therapy of Cancer: From Basic Research to Clinical Applications. Munich, Germany, 27-30 October 1999. Proceedings. PMID- 10711229 TI - Developing a global strategy for cancer. AB - Over the next 25 years there will be a dramatic increase in the number of people developing cancer. Globally, 10 million new cancer patients are diagnosed each year and this will be 20 million by the year 2020. Cancer is now the public's most feared disease. Billions of dollars are spent annually on cancer research by the drug industry, cancer charities and governments, but a cure for cancer appears elusive. And yet, we are in the midst of a revolution in our ability to image parts of the body, painlessly and in fine detail. We also now understand the intricate workings of the human genome--ultimately responsible for controlling all biological processes in health and disease. By the year 2003 the entire DNA sequence of the human genome will be determined. Powerful computer networks will allow detailed comparisons of genetic structure, so identifying new risk factors. Gene chips will detect minute code changes of considerable relevance. Novel screening technologies will allow us to detect just a few cancer cells in a patient. Robotically guided destructive processes will target abnormal cells in patients long before any cancer-related symptoms develop. And all this is likely by the first quarter of the next century. How are people, society and healthcare systems going to deal with these tremendous technological advances for cancer? Detailed information will be available in every home through easily understandable computer links. Choices now made by professionals will be equally understandable to all. Public education on health will be strengthened allowing a more critical and realistic assessment of media reports on new technologies. But as technology becomes more complex, the gap between the global rich and poor could widen. The export of unhealthy lifestyles--cigarette smoking, dietary habits and sedentary occupations will disproportionately increase cancer in many developing countries, which can least afford the treatment costs. The WHO Cancer Programme is developing a strategy to identify priorities in cancer prevention, detection and treatment in a wide range of epidemiological and economic settings. PMID- 10711230 TI - Viruses in human cancers. PMID- 10711231 TI - Control of the cell cycle and apoptosis. PMID- 10711232 TI - Molecular pathology and future developments. AB - There has already been a 'molecular' revolution in pathology. Demonstrating transcription of specific single genes or small gene sets and their protein products by in situ hybridisation and immunocytochemistry is routine in diagnostic and experimental pathology. A perhaps-greater revolution is imminent with the application of more recently established and emergent technologies in pathology. These include new approaches to polymerase chain reaction (PCR); simultaneous studies of multiple genes and their expression using oligonucleotide and cDNA arrays; serial analysis of gene expression (SAGE); expressed sequence tag (EST) sequencing, subtractive cloning and differential display; high throughput sequencing; comparative genomic hybridization, multiplex fluorescence in situ hybridisation (FISH) (spectral karyotyping); reverse chromosome painting; knockout and transgenic organisms; laser microdissection and micro-machining; and new methods in bio-informatics, 'data mining' and data visualisation. Molecular methods will profoundly change diagnosis, prognosis and treatment targeting in oncology and elucidate fundamental mechanisms of neoplastic transformation. Individual susceptibility to specific diseases will become assessable and screening will be refined. The new molecular biology will be most fruitful in partnership with classical approaches to pathology: the expectation that molecular methods alone will answer all pathological questions is unrealistic. A further challenge for the biomedical community in the 'genome era' will be to ensure that the benefits of these sophisticated technologies are enjoyed globally. PMID- 10711233 TI - Rho-like GTPases: their role in epithelial cell-cell adhesion and invasion. AB - The family members of small Rho-like GTPases, RhoA, Rac1 and Cdc42Hs, are regulators of diverse cellular signalling pathways, including cytoskeletal organisation, transcription and cell-cycle progression. Recent research has given insight into the complex regulation of cell-cell adhesion and migratory responses of epithelial cells. The Rho-like GTPases RhoA, Rac1 and Cdc42Hs as major determinants of cytoskeletal organisation have been identified as key regulators of epithelial architecture, as well as of cell migration. These findings highlight the complex regulation and cross-talk of GTPase-dependent signalling pathways arising from cell-cell and cell-matrix interactions. The molecular mechanism of how Rho-like GTPases couple to molecules mediating either cell-cell adhesion or cell migration will be of particular interest to understand the invasive phenotype of epithelial tumours. PMID- 10711234 TI - Cancer prevention: epidemiology and perspectives. AB - Following increases up until the late 1980s, some decline in cancer mortality has been observed in North America and in Western Europe. Approximately half the decline can be attributed to the levelling off in lung and other tobacco-related cancer epidemics and the rest to several factors, including reduced exposure to occupational carcinogens, prevention and early diagnosis, and improved treatment. Between 25 and 30% of all cancer deaths in Europe are due to tobacco smoking. In this review the effect of tobacco smoking on cancer incidence and mortality is examined, together with other important aetiological factors including alcohol, diet and environmental and occupational carcinogens. The effect of new treatments and the potential for prevention of cancer are also discussed. PMID- 10711235 TI - Screening for cancer: future potential. AB - Much progress has been made in cancer screening over the past decade, but a great deal more needs to be done if screening is to make a major impact on worldwide cancer mortality. Where fully implemented, cytological screening for cervical precursor lesions has had a major impact on mortality. However, the cost and required infrastructure levels are high, and new approaches are needed if screening is to be effective in the developing world. Testing for the human papillomavirus and automated liquid based cytology offer great promise to improve quality, reduce overall cost and make screening more viable generally. Breast screening has been less successful, although useful mortality benefits have been achieved in women aged over 50 years. Full implementation in countries that can afford it will save lives, but radical new approaches will be needed to conquer breast cancer. Colorectal cancer screening offers the best hope of a major reduction in cancer mortality over the next decade. Less certainty exists about screening for other major cancers such as lung, prostate and ovary, but a range of potential approaches merit investigation. PMID- 10711236 TI - Why can't we convince the young not to smoke? AB - Tobacco is the largest cause of preventable death and morbidity in the world. Significant progress has been made in national tobacco control programmes, followed by a significant reduction in smoking-associated diseases. However, other populations have taken up the habit and the worldwide surge in cigarette smoking by young people is particularly worrisome. Based on our own experience as well as reported data, we examined determinants of tobacco use, at the familial, peer and societal levels as well as various prevention strategies based on legislation, health promotion and society awareness. Reasons for failures include under-enforcement of legislation, uniform approach to diverse populations and too limited means. Recommendations for future actions should include integrated policies and health programmes. Most importantly, the society outlook on tobacco should be changed, making non-smoking the norm and the objective. PMID- 10711237 TI - Molecular genetics, natural history and the demise of childhood leukaemia. AB - The patterns of genetic change, clonal evolution, natural history and latency are very different in the paediatric leukaemias compared with adult epithelial cancers but are similar to those in other childhood cancers of mesenchymal stem cell origin. This distinction has a biological logic in the context of the selective pressures for clonal emergence in different developmental and cellular contexts and has a major impact on curability. Most childhood leukaemias and some other mesenchymal stem cell tumours are of fetal origin and can metastasize without corruption of restraints on cell proliferation or bypassing apoptosis. In marked contrast to most invasive or metastatic epithelial carcinomas in adults, these former cancers then retain sensitivity to therapeutic apoptosis. Moreover, their abbreviated and less complex evolutionary status is associated with less genetic diversity and instability, minimising opportunity for clonal selection for resistance. A minority of leukaemias in children and a higher fraction in adults do, however, have genetic alterations that bypass cell cycle controls and apoptosis imposition. These are the 'bad news' genotypes. The cellular and molecular diversity of acute leukaemia impacts also on aetiology. Paediatric acute leukaemias can be initiated prenatally by illegitimate recombination and fusion gene formation in fetal haemopoiesis. For acute lymphoblastic leukaemia (ALL) in children, twin studies suggest that a secondary postnatal molecular event is also required. This may be promoted by an abnormal or delayed response to common infections. Even for a classic case of a cancer that is intrinsically curable by systematic chemotherapy i.e. childhood ALL, prevention may turn out to be the preferred option. PMID- 10711238 TI - Screening for hereditary cancer and genetic testing, epitomized by breast cancer. AB - The new genetics is having an impact on many areas of healthcare. Diversity in the genetic code accounts for differences in phenotypes between populations and it is becoming apparent that genetic differences may have a role in predisposition to and behaviour of disease. Genetic models suggest that there are two types of genetic predisposition to disease: the so-called high and low penetrance genes. At present, most of the impact on medicine has been from highly penetrant genes, and genetic testing for disease predisposition, particularly for diseases of late onset (e.g. certain cancers) is in its infancy. As a general statement, approximately 5-10% of common cancers are due to such highly penetrant genes. The category of genes that will become of increasing interest is that of the low penetrance genes. Often these are normal variations in genes that result in a slightly increased risk of disease. These are analogous to high blood pressure carrying an increased risk of cardiovascular disease. Once rapid genetic analysis is available for these types of genes, such analysis would be analogous to taking someone's blood pressure in a general practitioner's (GP's) surgery to identify individuals at increased risk of cardiovascular disease. This will produce a revolutionary change in the way we practise medicine. Genetic analysis will become faster and may therefore be more commonplace. It is possible to envisage an era when genetic analysis will become a routine part of primary care to identify changes in low penetrance genes that will confer a 'risk profile' for patients. This will then enable their primary care physicians to advise about primary prevention and even prescribe certain preventive drugs to decrease the risk of certain diseases occurring. This proactive rather than reactive style of practising medicine is potentially exciting, however it carries with it ethical, legal and social implications for how we deal with this new knowledge. PMID- 10711239 TI - From Halsted to prevention and beyond: advances in the management of breast cancer during the twentieth century. AB - This commentary evaluates progress made in the treatment of breast cancer during the twentieth century. Most of the period from 1900 to 1970 was governed by the 'non-science' of anecdotalism and classical inductivism and was marked by the absence of a scientific gestalt. In keeping with the Halstedian concept that breast cancer was a local disease that spread throughout the body by contiguous extension and could be cured by more expansive surgery, the disease was treated with radical surgery. In 1950, however, a new era of enlightenment began to emerge. The awareness that there was a scientific process in which hypotheses generated from laboratory and clinical investigation could be tested by means of randomised clinical trials was a seminal advance, as were findings from studies that laid the groundwork for the modern era of steroid hormone action, including identification of oestrogen receptors. Expanding knowledge regarding tumour cell kinetics, tumour heterogeneity, and technological advances related to mammography and radiation therapy were also to play a role in making possible the advances in therapy that were subsequently to occur. In the past 30 years, as a result of laboratory and clinical investigation, the Halstedian thesis of cancer surgery was displaced by an alternative hypothesis that was supported by findings from subsequent clinical trials. A new paradigm governed surgery for breast cancer, and lumpectomy followed by radiation therapy became accepted practice. A second paradigm that governed the use of adjuvant systemic therapy arose as a result of laboratory and clinical investigation. Treating patients who were free of identifiable metastatic disease with systemic adjuvant therapy because some of them might develop distant disease in the future was a revolutionary departure from prior treatment strategy and became a new exemplar. Not only did the chemotherapy favourably alter the outcome of breast cancer patients, but the anti oestrogen tamoxifen benefited patients with all stages of the disease. Tamoxifen also reduced the incidence of contralateral breast cancer, as well as tumour in the ipsilateral breast following lumpectomy. The use of preoperative therapy was also found to enhance breast-conserving surgery in women with large tumours, although its value in other circumstances is still being defined. The observation that, as a result of tamoxifen administration, invasive and non-invasive breast cancers can be prevented in women who are at increased risk for such tumours, and the finding that pathological entities such as atypical hyperplasia, lobular carcinoma in situ (LCIS) and intraductal carcinoma (DCIS) can identify women who should be considered candidates for tamoxifen serve as a fitting capstone to the accomplishments of the twentieth century. Breast cancer prevention has now become a reality. Unfortunately, a variety of circumstances have arisen as the result of advances in the understanding and treatment of breast cancer over the last 30 years that threaten to nullify the progress that has been achieved. This distressing phenomenon may be reviewed as a 'paradox of accomplishment'. The numerous uncertainties, issues and questions that have arisen following the report of each advance in treatment, the surfeit of new information that has not yet been integrated into treatment strategies, the undesirable consequences of enhanced tumour detection, a reversion to Halstedianism and anecdotalism, and the uncertainty of therapeutic decision making resulting from the demonstration of small but statistically significant benefits, particularly in patients with good prognosis, need to be addressed. Inappropriate interpretation of those circumstances threatens to deny women with breast cancer and those at high risk for the disease the opportunity to benefit from treatments that have been proven to be of worth. Perhaps the most important accomplishment of the twentieth century relates to the change in the pro PMID- 10711240 TI - Selective oestrogen receptor modulation: molecular pharmacology for the millennium. AB - Knowledge of the mechanism of action and pharmacology of tamoxifen and raloxifene, for the prevention of breast cancer and osteoporosis respectively, has opened the door for the discovery of multifunctional medicines. There is now the potential to prevent osteoporosis, coronary heart disease, breast and endometrial cancer in postmenopausal women with elevated risk factors. PMID- 10711241 TI - Genetic pathways in colorectal and other cancers. AB - Cells from cancers show aberrant behaviour such as unrestrained growth, invasion into adjacent tissue and metastasis. All these features of cancer cell behaviour can be explained in terms of genetic changes and the functional impact of these changes. In this review, colorectal cancer (CRC) is examined as a classical example of multistep carcinogenesis. First there is an overview which shows that cancers develop by a process of somatic evolution. This gives rise to preferred genetic pathways of tumorigenesis. The factors which may influence the development and ultimate choice of genetic pathways are then examined. Next, CRC is studied as a specific disease and the putative genetic pathways are described. The mutations that comprise these pathways and the possible functional sequelae of these are explored. The review concludes with a look at those avenues which may further elucidate the natural history of CRC and lead to improved therapy. PMID- 10711242 TI - Skin cancer and solar UV radiation. AB - Ultraviolet (UV) radiation in sunlight is the most prominent and ubiquitous physical carcinogen in our natural environment. It is highly genotoxic but does not penetrate the body any deeper than the skin. Like all organisms regularly exposed to sunlight, the human skin is extremely well adapted to continuous UV stress. Well-pigmented skin is clearly better protected than white Caucasian skin. The sun-seeking habits of white Caucasians in developed countries are likely to have contributed strongly to the increase in skin cancer observed over the last century. Skin cancer is by far the most common type of cancer in the U.S.A. and Australia, which appears to be the result of an 'unnatural displacement' of people with sun-sensitive skin to sub-tropical regions. Although campaigns have been successful in informing people about the risks of sun exposure, general attitudes and behaviour do not yet appear to have changed to the extent that trends in skin cancer morbidity and the corresponding burden on public healthcare will be reversed. The relationship between skin cancer and regular sun exposure was suspected by physicians in the late 19th century, and subsequently substantiated in animal experiments in the early part of the 20th century. UV radiation was found to be highly genotoxic, and DNA repair proved to be crucial in fending off detrimental effects such as mutagenesis and cell death. In fact, around 1940 it was shown that the wavelength dependence of mutagenicity paralleled the UV absorption by DNA. In the 1970s research on UV carcinogenesis received a new impetus from the arising concern about a possible future depletion of the stratospheric ozone layer: the resulting increases in ambient UV loads were expected to raise skin cancer incidences. Epidemiological studies in the last decades of the 20th century have greatly refined our knowledge on the aetiology of skin cancers. Analyses of gene mutations in skin carcinomas have identified UV radiation as the cause. The relationship between the most fatal skin cancer, i.e. malignant melanoma and solar UV exposure is, however, still unclear and needs to be clarified to optimise preventive measures and minimise mortality from skin cancers. PMID- 10711243 TI - Discovering novel chemotherapeutic drugs for the third millennium. AB - There is enormous potential for the discovery of innovative cancer drugs with improved efficacy and selectivity for the third millennium. In this review we show how novel mechanism-based agents are being discovered by focusing on the molecular targets and pathways that are causally involved in cancer formation, maintenance and progression. We also show how new technologies, from genomics through high through-put bioscience, combinatorial chemistry, rational drug design and molecular pharmacodynamic and imaging techniques, are accelerating the pace of cancer drug discovery. The process of contemporary small molecule drug discovery is described and progress and current issues are reviewed. New and potential targets and pathways for therapeutic intervention are illustrated. The first examples of a new generation of molecular therapeutics are now entering hypothesis-testing clinical trials and showing activity. The early years of the new millennium will see a range of exciting new agents moving from bench to bedside and beginning to impact on the management and cure of cancer. PMID- 10711245 TI - Gene therapy for cancer. PMID- 10711244 TI - Cancer chemoprevention: progress and promise. AB - Cancer chemoprevention is the use of agents to inhibit, delay or reverse carcinogenesis. The focus of chemoprevention research in the next millennium will include defining the genotypic and phenotypic (functional and histological) changes during carcinogenesis, the cancer risk conferred by these changes, their modulation in preclinical experimentation and randomised clinical trials by chemopreventive drugs, dietary agents and regimens and treatments resulting from early detection. The key elements of this research effort will be basic and translational risk evaluation programmes; chemopreventive and dietary agent drug discovery and development; development of transgenic animal models; required safety and pharmacology studies; well-designed phase I, II and III chemoprevention studies; and much expanded early detection programmes. The large number of chemoprevention research programmes now ongoing ensures that the promise of chemoprevention will continue to be realised in the next decade. PMID- 10711246 TI - Improving communication with cancer patients. AB - If doctors and nurses involved in cancer care are to help patients and their families achieve an optimal level of quality of life and psychological adjustment they must be able to carry out key communication tasks successfully. Yet, objective scrutiny of their consultations confirms that deficiencies in their ability to conduct these tasks remain. The reasons for this are discussed before important innovations in training and their impact are described. PMID- 10711247 TI - Histopathological workup of sentinel lymph nodes: how much is enough? PMID- 10711248 TI - Antimicrobial prescribing. PMID- 10711249 TI - Prognostic evaluation of metallothionein expression in human colorectal neoplasms. AB - AIM: To investigate the role of metallothionein in colorectal tumours and the possible relation with other factors associated with tumour progression: expression of cathepsin D (CD), CD44, p53, Rb, bcl-2, c-erbB-2, epidermal growth factor receptor (EGFR), proliferation indices (Ki-67, proliferating cell nuclear antigen (PCNA)), and conventional clinicopathological variables. METHODS: The immunohistochemical expression of metallothionein was investigated in 23 cases of colorectal adenoma and 94 adenocarcinomas. Metallothionein expression was examined by the avidinbiotin peroxidase immunoperoxidase (ABC) using the monoclonal mouse antibody E9, on formalin fixed, paraffin embedded tissue. RESULTS: Positive metallothionein expression (> 5% of neoplastic cells) was observed in 30.4% of adenomas and 25.5% of adenocarcinomas, while 8.7% of adenomas and 14.9% carcinomas showed focal metallothionein positivity. In contrast, 60.9% of adenomas and 59.6% of carcinomas almost completely lacked metallothionein expression. In the series of adenocarcinomas, metallothionein expression was inversely correlated with CD44 in neoplastic cells (p = 0.01). There was no statistically significant difference of metallothionein expression, or the other variables examined, between adenocarcinomas and adenomas. CONCLUSIONS: Metallothionein expression does not seem to indicate aggressive biological behaviour in colorectal adenocarcinomas, in comparison with the other types of carcinoma. The inverse correlation with CD44 could suggest that the decreased metallothionein expression may contribute to the metastatic spread of the lymph node involvement in colorectal cancer. Metallothionein expression does not seem to represent an independent prognostic marker in colorectal cancer. PMID- 10711250 TI - Cyclin dependent kinase inhibitor p27Kip1 expression in normal and neoplastic cervical epithelium. AB - AIM: To investigate whether there is loss of the p27Kip1 protein in developing cervical cancer and whether p27Kip1 immunoreactivity has any relation to the proliferative indicator Ki-67. METHODS: The expression of p27Kip1 and Ki-67 was assessed by immunohistochemistry in serial sections from normal epithelium (13), low grade (27) and high grade (19) squamous intraepithelial lesions (LSIL, HSIL), and invasive cervical cancer (23). In the SIL cases the presence of human papillomavirus (HPV) genomic sequences was assessed by in situ hybridisation. The results were evaluated by image analysis, and reported as mean score of the percentage of p27Kip1 and of Ki-67 positive cells in each histological group. RESULTS: In general, p27Kip1 immunostaining was related to squamous differentation, and was intense in normal epithelium (47%), while it was reduced in SIL lesions as an effect of the decreased number of differentiating cells. However, decrease in the p27Kip1 expression was more evident in LSIL (36%) than in HSIL (39%); in the latter, p27Kip1 had a different intraepithelial distribution in that the staining extended to the basal cells. The average levels of p27Kip1 were similar in SIL lesions associated to low, intermediate, and high risk HPV types. Compared with normal epithelium and dysplasia, invasive cancer showed significantly lower p27Kip1 levels (23%). There was no relation between p27Kip1 and Ki-67 labelling indices in any of the histological groups examined. CONCLUSIONS: A reduction in p27Kip1 protein occurs in cervical cancer independently of the proliferative status. The changes in p27Kip1 expression may be related to the unregulated kinetics of developing cervical cancer. PMID- 10711251 TI - Decidual T lymphocyte activation in hydatidiform mole. AB - AIM: To quantify and compare decidual leucocyte subpopulations in complete and partial hydatidiform molar pregnancy with those in normal early pregnancy. METHODS: Decidual leucocytes were characterised using an avidin-biotin technique and a panel of monoclonal antibodies in formalin fixed, paraffin embedded decidual tissues from 10 normal first trimester pregnancy terminations and from 13 partial and 13 complete hydatidiform moles. Immunostained cells were fully quantitated and differences between subject groups were analysed using the Mann Whitney test. T lymphocyte populations were further characterised using double immunohistochemical labelling. RESULTS: The numbers and percentages of CD3+ and CD4+ T cells were significantly increased in complete hydatidiform moles compared with partial moles and normal early pregnancy decidua. No differences were found in the number or percentage of CD8+ T cells. The CD8+ to CD4+ T cell ratio decreased significantly in complete mole compared with partial mole and normal decidua. The numbers and percentages of CD45RO+ cells increased significantly in both partial and complete hydatidiform mole compared with normal first trimester decidua. Double labelling confirmed that 50% of CD3+ T cells in complete and partial molar pregnancy coexpressed CD45RO, compared with 30% in normal pregnancy. Other leucocyte populations (eGLs, macrophages, B cells, and classical natural killer cells) did not differ between complete and partial mole and normal pregnancy decidua. CONCLUSIONS: The presence of an increased population of activated decidual CD45RO+ T cells in complete and partial molar pregnancy suggests altered maternal immune responses against molar trophoblast. PMID- 10711252 TI - Immunohistochemical study of thrombospondin and its receptors alpha root of beta 3 and CD36 in normal thyroid and in thyroid tumours. AB - AIM: To describe the pattern of distribution of thrombospondin (TSP1) and its receptors, alpha root of beta 3 integrin and CD36, in normal human thyroid tissue and to compare their expression in different benign and malignant thyroid conditions. METHODS: Immunohistochemistry was used to study TSP1 and its receptors in 40 surgical thyroidectomy specimens (normal parenchyma, 7; follicular adenoma, 4; multinodular goitre, 13; papillary carcinoma, 6; follicular carcinoma, 8; anaplastic carcinoma, 2). RESULTS: In the normal thyroid parenchyma, there was weak expression of TSP1 limited to the vessels with no staining of the extracellular matrix. In goitres, the expression of TSP1 was more pronounced in areas of fibrosis, with staining of alpha root of beta 3 on thyrocytes located in the vicinity. In thyroid adenomas, expression of TSP1 was slightly enhanced compared with normal tissue, located in the basement membrane of vessels. In papillary carcinomas, TSP1 was abundant in the desmoplastic stroma with a cytoplasmic distribution of alpha root of beta 3 integrin in thyrocytes. In follicular carcinomas, TSP1 was less abundant in the extracellular matrix, limited to the vessels of the stroma with a weaker expression of alpha root of beta 3 on thyrocytes than in papillary carcinomas. In anaplastic carcinomas, TSP1 was only present in the numerous capillaries of the stroma, with a marked positivity for alpha root of beta 3 in one case. No immunostaining of thyrocytes is observed with CD36. CONCLUSIONS: These results suggest the importance of the interaction between alpha root of beta 3 integrin and TSP1 during remodelling of the matrix in fibrous goitres with areas of early sclerosis comparable with wound healing. In papillary carcinomas, the overexpression of TSP1 restricted to the stroma suggests protective effects against tumour progression. PMID- 10711253 TI - Mononuclear leucocyte function tests in the assessment of the biocompatibility of peritoneal dialysis fluids. AB - BACKGROUND: Previous studies showed that the currently used dextrose based peritoneal dialysis fluids impair several leucocyte functions. AIMS: To determine which in vitro mononuclear leucocyte (monocyte) function tests most clearly reflect the biocompatibility of peritoneal dialysis fluid. METHODS: Monocytes were tested for phagocytic capacity, bactericidal activity, Fc and C3 receptor expression, and chemiluminescence response, and by analysis of the release of interleukin 8 (IL-8) and tumour necrosis factor alpha (TNF alpha) in the presence of test fluids. Cytokine release was studied in an alternative dynamic in vitro peritoneal dialysis model in which monocytes were exposed to test fluid that was continuously equilibrated with an interstitial fluid-like medium through a microporous membrane. The chemiluminescence response by stressed monocytes was also tested after an 18 h recovery period. All tests were performed during or after exposure to different degrees of glycerol induced osmotic stress and after exposure to a 1% milk-whey derived, polypeptide enriched test fluid. Cells incubated in 0.1% gel Hanks buffer (GH) served as control. RESULTS: Osmotic stress induced impairment of leucocyte function was found by the chemiluminescence assay (mean (SEM): 179 (20)% v 138 (23)% after 30 minutes in 0.5% and 1.5% glycerol, respectively) and by the analysis of IL-8 released by monocytes (44 (9) ng in 0.7% glycerol v 40 (7) ng in 2.0% glycerol). Only the chemiluminescence assay showed a protective effect of polypeptides on leucocyte function (after > or = 60 minutes). If monocytes were allowed to recover in culture medium after exposure to test fluids, the changes in chemiluminescence response appeared to be reversible after a 30 minute exposure, but became more pronounced after 60 and 120 minutes. The phagocytosis and bacterial killing assays were less sensitive. The observations carried out with the phagocytosis assay did not correspond with the Fc or C3 receptor density data. CONCLUSIONS: The release of IL-8 by peripheral blood monocytes in a two compartment model and their chemiluminescence response are appropriate assays for the assessment of changes in leucocyte function in response to different peritoneal dialysis fluids. PMID- 10711254 TI - A human skin explant model for predicting graft-versus-host disease following bone marrow transplantation. AB - Graft-versus-host disease (GVHD) is the most serious complication following bone marrow transplantation, with an incidence of 40-60%. The disease can be fatal in 50% of cases, even in patients receiving marrow from an HLA identical sibling. Several assays have been developed to try to predict the development of GVHD, including mixed lymphocyte culture reaction, cytotoxic T lymphocyte precursor, and helper T lymphocyte precursor frequency assays. This review describes an in vitro skin explant model which has been used since 1988 for both predicting acute GVHD in HLA identical sibling bone marrow transplantation and studying the pathophysiology of the disease. The model involves sensitising donor lymphocytes in vitro in a primary mixed lymphocyte reaction and then evaluating the secondary response on patient skin biopsies by grading the graft-versus-host reactivity (grades I-IV) histopathologically. From analysis of collective data the model is a clear predictor of GVHD and superior to the other assays widely used, with a correlation of 82% with clinical outcome. The skin explant model allows the investigator to study the pathogenesis of GVHD. The cytokines TNF alpha and IFN gamma are shown to be important mediators of cellular damage in graft-versus-host reactions. Recent work has also involved using the model to study the alloreactivity of cord blood. The model is currently being assessed in several bone marrow transplantation centres in Europe on different patient groups including those who receive marrow from haploidentical and matched unrelated donors. PMID- 10711255 TI - A reliable method for the simultaneous identification of H pylori and gastric metaplasia in the duodenum. AB - While an association of Helicobacter pylori infection with duodenal mucosa gastric metaplasia has been described, the details and extent of the interaction are lacking. One of the limiting factors has been the lack of a staining technique that allows simultaneous visualisation of the bacteria and gastric metaplasia in the duodenum. This report describes a new stain that allows the simultaneous visualisation of duodenal gastric metaplasia and H pylori and compares the new stain with the component stains. PMID- 10711256 TI - Pseudo-Gaucher cells in myelodysplasia. AB - A case of myelodysplastic syndrome is reported, in which the bone marrow contained many cells with the typical light microscopic morphology of Gaucher cells. In the absence of any evidence of inherited Gaucher's disease, these cells are considered to be pseudo-Gaucher cells, which have been described previously in association with other haematological diseases. This is the first report of their occurrence in myelodysplastic syndrome. PMID- 10711257 TI - Immunohistochemical identification of erythroid precursors in paraffin embedded bone marrow sections: spectrin is a superior marker to glycophorin. AB - AIM: To investigate whether spectrin can be used as an immunohistochemical marker for erythroid precursors in routinely processed paraffin embedded bone marrow sections. METHODS: Bone marrow biopsies and clot sections were stained with rabbit antihuman erythrocyte spectrin antibodies, specific for erythroid cells as shown by western blotting and bone marrow smears, and compared to sections stained with antiglycophorin monoclonal antibodies (JC159 and Ret49f). RESULTS: Antispectrin antibodies resulted in diffuse cytoplasmic staining of early erythroblasts and membranous staining of late erythroblasts as well as erythrocytes. In haematopathological samples, immature erythroid cell clusters were clearly identified. In contrast, antiglycophorin monoclonal antibodies resulted in only membranous staining of late erythroblasts, and faint staining of early erythroblasts. CONCLUSIONS: Spectrin may be a superior marker to glycophorin for the identification of erythroid precursors in paraffin embedded sections. PMID- 10711258 TI - Metastases in axillary sentinel lymph nodes in breast cancer as detected by intensive histopathological work up. AB - AIM: To assess the value of the intensive histological work up of axillary sentinel lymph nodes (SLN) to demonstrate regional metastatic disease. METHODS: From a series of 58 successful lymphatic mapping procedures, 78 SLN were analysed by serial sections (mean of 49 levels/SLN) and by immunostaining to cytokeratin and epithelial membrane antigen, and the results compared with those obtained by assessing the central cross section. RESULTS: The central cross section would have failed to detect metastases in eight of 26 lymph nodes (31%) in patients with breast cancer metastasising to the SLN only. This would have led to a false negative node status in six of 21 patients (29%). Two micrometastases were detected with the aid of immunostains. CONCLUSIONS: The results suggest the need to examine SLN at multiple levels and to use immunohistochemistry in negative cases. Serial sections are also useful in the case of micrometastases, as some of these may convert to macrometastases at deeper levels. Multiple level investigation of SLN and immunohistochemistry in the event of the negativity of standard stains would result in improved staging and an increase in the proportion of node positive disease detected. PMID- 10711259 TI - Accuracy and cost of intraoperative lymph node frozen sections at radical prostatectomy. AB - AIM: To assess the value of intraoperative diagnostic examination of frozen sections of lymph nodes removed during radical prostatectomy. METHODS: Pelvic lymph nodes from patients with prostatic carcinoma were obtained (1) as frozen sections during radical prostatectomy, to exclude patients from non-curative surgery, and (2) as paraffin sections postoperatively from lymphadenectomy performed at radical prostatectomy, to stage the tumour and assess need for adjuvant treatment. Findings from the two approaches were used to assess the accuracy and cost of frozen section diagnosis, and to judge the results of omitting intraoperative diagnosis. RESULTS: In 82 patients frozen section revealed metastasis in six (7.3%), and metastases were found in a further four (4.9%) on paraffin sections (false negatives). Of the 195 patients undergoing staging lymphadenectomy (without frozen section), metastatic cancer was seen in nine cases (4.6%). The frozen section cost of metastatic cancer detection per patient was calculated as 7516 Pounds (550 Pounds x 82/6), with an associated false negative rate of 33%. CONCLUSIONS: Frozen section diagnosis of metastatic carcinoma in pelvic lymph nodes before radical prostatectomy has a high false negative rate and is costly. It may not be justified with the observed low incidence of lymph node metastasis. PMID- 10711260 TI - Cryptococcal meningitis in an HIV negative patient with systemic sarcoidosis. AB - A case of Cryptococcus neoformans meningitis is described in an HIV negative patient with undiagnosed systemic sarcoidosis. The patient presented with signs of meningitis together with generalised lymphadenopathy and hepatosplenomegaly. Cryptococcal meningitis was diagnosed on lumbar puncture. She was treated with intravenous amphotericin B but died within two weeks of admission. Necropsy revealed lesions in the lungs, liver, spleen, lymph nodes, small intestine, and bone marrow consistent with sarcoidosis. Microscopically the lesions contained non-caseating epithelioid cell granulomas typical of sarcoidosis. No Schaumann or Hamazaki-Wesenberg bodies were identified. Cryptococcus neoformans meningitis is generally associated with immunosuppressive disorders. As T cell abnormalities have been described in sarcoidosis, this could have been a case of opportunistic infection. Although rare, sarcoidosis merits consideration in patients with cryptococcal disease in the absence of HIV infection. PMID- 10711261 TI - L26 (CD20) staining of Bouin fixed bone marrow biopsies. PMID- 10711262 TI - Calcium oxalate (Weddellite) crystals within ductal carcinoma in situ. PMID- 10711263 TI - Reporting of occupational and environmental research: use and misuse of statistical and epidemiological methods. AB - OBJECTIVES: To report some of the most serious omissions and errors which may occur in papers submitted to Occupational and Environmental Medicine, and to give guidelines on the essential components that should be included in papers reporting results from studies of occupational and environmental health. METHODS: Since 1994 Occupational and Environmental Medicine has used a panel of medical statisticians to review submitted papers which have a substantial statistical content. Although some studies may have genuine errors in their design, execution, and analysis, many of the problems identified during the reviewing process are due to inadequate and incomplete reporting of essential aspects of a study. This paper outlines some of the most important errors and omissions that may occur. Observational studies are often the preferred choice of design in occupational and environmental medicine. Some of the issues relating to design, execution, and analysis which should be considered when reporting three of the most common observational study designs, cross sectional, case-control, and cohort are described. An illustration of good reporting practice is given for each. Various mathematical modelling techniques are often used in the analysis of these studies, the reporting of which causes a major problem to some authors. Suggestions for the presentation of results from modelling are made. CONCLUSIONS: There is increasing interest in the development and application of formal "good epidemiology practices". These not only consider issues of data quality, study design, and study conduct, but through their structured approach to the documentation of the study procedures, provide the potential for more rigorous reporting of the results in the scientific literature. PMID- 10711264 TI - Occupational exposure to carcinogens in the European Union. AB - OBJECTIVES: To construct a computer assisted information system for the estimation of the numbers of workers exposed to established and suspected human carcinogens in the member states of the European Union (EU). METHODS: A database called CAREX (carcinogen exposure) was designed to provide selected exposure data and documented estimates of the number of workers exposed to carcinogens by country, carcinogen, and industry. CAREX includes data on agents evaluated by the International Agency for Research on Cancer (IARC) (all agents in groups 1 and 2A as of February 1995, and selected agents in group 2B) and on ionising radiation, displayed across the 55 industrial classes. The 1990-3 occupational exposure was estimated in two phases. Firstly, estimates were generated by the CAREX system on the basis of national labour force data and exposure prevalence estimates from two reference countries (Finland and the United States) which had the most comprehensive data available on exposures to these agents. For selected countries, these estimates were then refined by national experts in view of the perceived exposure patterns in their own countries compared with those of the reference countries. RESULTS: About 32 million workers (23% of those employed) in the EU were exposed to agents covered by CAREX. At least 22 million workers were exposed to IARC group 1 carcinogens. The exposed workers had altogether 42 million exposures (1.3 mean exposures for each exposed worker). The most common exposures were solar radiation (9.1 million workers exposed at least 75% of working time), environmental tobacco smoke (7.5 million workers exposed at least 75% of working time), crystalline silica (3.2 million exposed), diesel exhaust (3.0 million), radon (2.7 million), and wood dust (2.6 million). CONCLUSION: These preliminary estimates indicate that in the early 1990s, a substantial proportion of workers in the EU were exposed to carcinogens. PMID- 10711266 TI - Cohort study of occupational risk factors of low back pain in construction workers. AB - OBJECTIVES: To identify work related risk factors of future low back pain (LBP) in a cohort of construction workers free of LBP at the start of follow up. METHODS: The Hamburg construction worker study comprises 571 male construction workers who have undergone two comprehensive interview and physical examination surveys. A cohort of 285 subjects without LBP at baseline was identified. After a follow up of 3 years, the 1 year prevalence of self reported LBP was determined in the 230 men followed up (80.7%). Prevalence ratios (PRs) with 95% confidence intervals (95% CIs) of LBP at follow up according to self reported work tasks of construction workers measured at baseline were estimated from Cox's regression models which were adjusted for age, and anthropometric measures. RESULTS: At follow up 71 out of 230 workers (30.9%) reported LBP during the preceding 12 months. Four work tasks (scaffolding, erecting roof structures, sawing wood, laying large sandstones) with an increased risk of 1 year prevalence of LBP at follow up were further evaluated. After further adjustment for occupation the relative risk was increased for workers who had reported > or = 2 hour/shifts laying large sandstones (PR = 2.6; 95% CI 1.1 to 6.5). Work load of bricklayers was additionally estimated by an index on stone load (high exposure: PR = 4.0; 95% CI 0.8 to 19.8), and an index for laying huge bricks/blocks (yes/no: PR = 1.7; 95% CI 0.5 to 5.7). CONCLUSIONS: The results suggest that self reported differences in brick characteristics (size and type of stone) and temporal aspects of the work of bricklayers (average hours per shift laying specified stones) can predict the future prevalence of LBP. The data have to be interpreted with caution because multiple risk factors were tested. PMID- 10711265 TI - Neurobehavioural effects of occupational exposure to cadmium: a cross sectional epidemiological study. AB - BACKGROUND: A patient with unexplained minor behavioural changes associated with an axonal sensorimotor polyneuropathy had a history of chronic occupational exposure to cadmium (Cd). Although animal studies have shown that Cd is a potent neurotoxicant, little is known about its toxicity for the human central nervous system. The aim of this study was to investigate the toxic potential of chronic occupational exposure to Cd on neurobehavioural functions. METHODS: A cross sectional epidemiological study was conducted ina group of Cd workers and an age matched control group. Eighty nine adult men (42 exposed to Cd and 47 control workers) were given a blinded standardised examination that consisted of computer assisted neurobehavioural tests (neurobehavioural examination system), a validated questionnaire to assess neurotoxic complaints (neurotoxicity symptom checklist--60, NSC-60), and a standardised self administered questionnaire to detect complaints consistent with peripheral neuropathy and dysfunction of the autonomic nervous system. Historical and current data on biomonitoring of exposure to Cd, either the highest value of Cd in urine (CdU in microgram Cd/g creatinine) of each Cd worker during work (CdUmax) or the current value (CdUcurrent) of each control, were available as well as data on microproteinuria. RESULTS: Cd workers (CdUmax: mean (range), 12.6 (0.4-38.4)) performed worse than the controls (CdUcurrent: mean (range), 0.7 (0.1-2.0)) on visuomotor tasks, symbol digit substitution (p = 0.008), and simple reaction time to direction (p = 0.058) or location (p = 0.042) of a stimulus. In multiple linear regression analysis, symbol digit substitution, simple direction reaction time test, and simple location reaction time test were significantly related to CdUmax, (beta = 0.35 (p < 0.001), beta = 0.25 (p = 0.012), and beta = 0.23 (p = 0.021) respectively). More complaints consistent with peripheral neuropathy (p = 0.004), complaints about equilibrium (p = 0.015), and complaints about concentration ability (p = 0.053) were found in the group exposed to Cd than in the control group, and these variables correlated positively with CdUmax (peripheral neuropathy: beta = 0.38, p < 0.001; equilibrium: beta = 0.22, p = 0.057; concentration ability: beta = 0.27, p = 0.020). CONCLUSION: Slowing of visuomotor functioning on neurobehavioural testing and increase in complaints consistent with peripheral neuropathy, complaints about equilibrium, and complaints about concentration ability were dose dependently associated with CdU. Age, exposure to other neurotoxicants, or status of renal function could not explain these findings. The present study also indicates that an excess of complaints may be detected in Cd workers before signs of microproteinuria induced by Cd occur. PMID- 10711267 TI - Cross sectional study of a workforce exposed to hand-arm vibration: with objective tests and the Stockholm workshop scales. AB - OBJECTIVES: Medical surveillance of workforces exposed to vibration has been recommended with the Stockholm workshop scales. The aims of this study were (a) to evaluate how the results of the objective tests individually and jointly associated with the final Stockholm workshop staging, (b) how this staging related to the history of exposure to vibration, and (c) how different trades were affected by the hazards from vibrating tools. METHODS: All workers exposed to vibration in a heavy engineering company were examined with a questionnaire and a battery of tests. An assessment of staging by the Stockholm workshop scales was made. Estimates of the daily exposure and lifetime dosage of vibration of the various trades were reached. RESULTS: The average years of tool use was 23.3 years (range 3-47 years) and the mean lifetime exposure was 11,022 (range 1012 46,125) hours. The individual neurological tests were all strongly associated with the Stockholm neurological staging but the cold provocation test was not associated with the Stockholm vascular staging. Neurological staging was significantly associated with age, years of tool use, and total hours of exposure to vibration, but not with trade or smoking. Vascular staging was significantly associated with age, years of tool use, total hours of exposure to vibration, and trade, but not with smoking. The mean neurological latent period was 19.7 (range 2-40) years and for the vascular component 19.1 (range 2-40) years. These means varied significantly by trade. The overall prevalence of neurological findings of 62% was greater than the overall prevalence of vascular findings, which was 33%. CONCLUSIONS: (1) The neurological objective tests were found to be of use in neurological staging. The cold provocation test was not associated with the vascular staging and therefore was of little value. (2) Years of tool use was the exposure variable most significantly associated with evidence of damage to neurological component while years of tool use and trade were the variables most associated with vascular damage. (3) The prevalence of neurological symptoms (62%) was greater than the prevalence of vascular symptoms (33%). (4) Dressers and welders have shorter latent periods than platers and fitters. PMID- 10711268 TI - Respiratory health surveillance in a toluene di-isocyanate production unit, 1967 97: clinical observations and lung function analyses. AB - OBJECTIVES: To characterise irritant and allergic airway responses and assess changes in forced vital capacity (FVC) and forced expiratory volume in 1 second (FEV1) relative to exposure to toluene di-isocyanate (TDI). METHODS: Employees (n = 313) ever assigned to a TDI production unit for > or = 3 months (1967-92) were identified from personnel records along with 158 frequency matched referents without known exposure to TDI. Reports made during visits to the occupational clinic of incidents related to exposure to TDI and annual periodic examination results (questionnaire, physical findings, and spirometry) were abstracted and assessed relative to industrial hygiene estimates of exposure to TDI. RESULTS: Mean 8 hour time weighted average estimates of TDI concentrations ranged from 9.9 ppb in jobs with potentially high exposure during the early years of plant operations to 0.5 ppb in jobs with potentially low exposure in more recent years. The corresponding rates of visits to the clinic due to incidents of exposure to TDI (including both irritant and allergic airway responses) declined from 20.5 to 1.0 visits per 100 years of employment at the unit. The annual incidence of asthma induced by TDI declined from 1.8% before 1980 to 0.7% afterwards. Neither cross sectional nor longitudinal analyses of FVC and FEV1 showed significant dose response findings relative to exposure to TDI across the total exposed population. Among cases of occupational asthma there was an apparent initial decline in FEV1 within 2 years of first reporting symptoms, but not an accelerated rate of decline in follow up tests from 4-30 years after induction of asthma. CONCLUSIONS: Occurrences of both asthma induced by TDI and irritant airway responses due to exposure to TDI were found in this cohort, but there was no relation between cumulative exposure to TDI and irreversible airflow obstruction as assessed by spirometry. PMID- 10711269 TI - Investigation of factors which might indicate susceptibility to particulate air pollution. AB - OBJECTIVES: To determine whether previous symptoms or recognized risk factors of cardiovascular ill health, are associated with an increased likelihood of adverse health effects related to particulate air pollution. METHODS: Cardiovascular event rates were studied relative to urban concentrations of particulate air pollution and baseline risk factors. The Edinburgh artery study consisted of a cohort of 1592 subjects aged 55-74 and was followed up to the end of March 1998 for a median of 10 years resulting in about 5 million person-days of observation. Baseline measurements included plasma fibrinogen and blood and plasma viscosity. A nested case-control approach was used to investigate a possible interaction between effects of these selected baseline risk factors and particulate air pollution, on subsequent event rates. RESULTS: During the follow up period there were 343 fatal and non-fatal myocardial infarctions or strokes. Trends in adverse cardiovascular outcomes related to pollution were identified among subjects belonging to the highest baseline quintile of plasma fibrinogen. Evidence for interactions between concentrations of particulate pollution and fibrinogen was not established at conventional levels of significance. CONCLUSIONS: People with high concentrations of plasma fibrinogen might be more susceptible to adverse cardiovascular effects of particulate air pollution, but limitations of power mean that evidence relating to such an interaction is not conclusive. A range of cardiopulmonary risk factors warrant investigation in relation to possible susceptibility to air pollution. PMID- 10711270 TI - Prospective study of work related respiratory symptoms in trainee bakers. AB - OBJECTIVES: To investigate the occurrence of work related respiratory symptoms and to assess the effect of atopy in a group of trainee bakers. METHODS: A prospective study of work related respiratory symptoms among 125 trainee bakers who were investigated with a questionnaire plus skin prick test with wheat flour and alpha-amylase allergens at baseline and then after 6, 18, and 30 months. RESULTS: At the baseline examination, four students (3.2%) complained of respiratory symptoms (cough and rhinitis) when working with flours and four were skin positive to wheat flour or alpha-amylase. The incidence of work related respiratory symptoms was 3.4% at 6 months, and the cumulative incidence was 4.8% and 9.0% at 18 and 30 months, respectively. The incidence of skin sensitisation to occupational allergens was 4.6% at 6 months and the cumulative incidence was 4.6% at 18 months and 10.1% at 30 months. The generalised estimating equation approach to longitudinal data showed that work related respiratory symptoms in the study population was significantly associated with a personal history of allergic disease (odds ratio (OR) 5.8, 95% confidence interval (95% CI) 1.8 to 18.2) and skin sensitisation to wheat flour or alpha-amylase (OR 4.3, 95% CI 1.2 to 14.9). Atopy based on prick test was not related to the occurrence of work related respiratory symptoms over time (OR 1.1, 95% CI 0.3 to 3.8). CONCLUSIONS: Personal history of allergic disease is a predisposing factor for the development of symptoms caused by exposure to wheat flour and may be a criterion of unsuitability for starting a career as a baker. Atopy based on the skin prick test is useful for identifying subjects with allergic disease, but should not be used to exclude non-symptomatic atopic people from bakery work. PMID- 10711271 TI - Two patients with occupational asthma who returned to work with dust respirators. AB - OBJECTIVES: To assess the efficacy of dust respirators in preventing asthma attacks in patients with occupational asthma (asthma induced by buckwheat flour or wheat flour). METHODS: The effect of the work environment was examined in two patients with occupational asthma with and without the use of a commercially available mask or a dust respirator. Pulmonary function tests were performed immediately before and after work and at 1 hourly intervals for 14 hours after returning to the hospital. RESULTS: In patient 1, environmental exposure resulted in no symptoms during and immediately after work, but coughing, wheezing, and dyspnoea developed after 6 hours. Peak expiratory flow rate (PEFR) decreased by 44% 7 hours after leaving the work environment, showing only a positive late asthmatic reaction (LAR). In patient 2, environmental exposure resulted in coughing and wheezing 10 minutes after initiation during bread making, and PEFR decreased by 39%. After 7 hours, PEFR decreased by 34%. The environmental provocation tests in both patients were repeated after wearing a commercial respiratory. This resulted in a complete suppression of LAR in patient 1 and of immediate asthmatic reaction (IAR) and LAR in patient 2. CONCLUSIONS: Two patients with asthma induced by buckwheat flour or wheat flour in whom asthmatic attacks could be prevented with a dust respirator are reported. Dust respirators are effective in preventing asthma attacks induced by buckwheat flour and wheat flour. PMID- 10711272 TI - Incidence of cancer among workers exposed to vinyl chloride in polyvinyl chloride manufacture. AB - Based on results from two previous studies where an excess of melanomas was found in a cohort of workers exposed to vinyl chloride (VCM), a follow up of the incidence of cancer in the same cohort of 428 workers was carried out to scrutinize whether or not the excess could be confirmed by new cases. The total number of deaths in the study group from 1953 to the end of 1993 was 132 v 141 expected, and the total number of incident cancer cases was 56 v 57 expected. There were 11 cases of lung cancer v eight expected, seven cases of melanomas v 2.07 expected, and two cases of thyroid cancer v 0.34 expected. Five of the seven melanoma cases had occurred in the group that had been most heavily exposed to VCM v 0.7 expected. In the present follow up we also found five cases of the spinocellular cancer of the skin v 1.7 expected. Out of these five cases four were diagnosed after 1984. Two of the five cases v 0.7 expected had occurred in the most heavily exposed group. The total number of skin cancers (melanomas and spinocellular cancers) were 12 v 3.7 expected. There was one new case of melanoma between 1985 and 1993 v 0.7 expected. Hence, the strength of the relation between the observed and expected number of cases was reduced compared with the last follow up, and does not strengthen the previously indicated causal relation between exposure to VCM and development of malignant melanoma. There was no excess of testicular cancers in this study. The present results may indicate that occurrence of spinocellular skin cancer could bear some relation to work in the manufacture of polyvinyl chloride (PVC). Confirmation is needed from studies on other cohorts exposed to VCM. PMID- 10711273 TI - Pilots' fitness, a time for questions. PMID- 10711274 TI - Chlorination disinfection byproducts in water and their association with adverse reproductive outcomes: a review. AB - OBJECTIVES AND METHODS: Chlorination has been the major disinfectant process for domestic drinking water for many years. Concern about the potential health effects of the byproducts of chlorination has prompted the investigation of the possible association between exposure to these byproducts and incidence of human cancer, and more recently, with adverse reproductive outcomes. This paper evaluates both the toxicological and epidemiological data involving chlorination disinfection byproducts (DBPs) and adverse reproductive outcomes, and makes recommendations for future research. RESULTS AND CONCLUSIONS: Relatively few toxicological and epidemiological studies have been carried out examining the effects of DBPs on reproductive health outcomes. The main outcomes of interest so far have been low birth weight, preterm delivery, spontaneous abortions, stillbirth, and birth defects--in particular central nervous system, major cardiac defects, oral cleft, and respiratory, and neural tube defects. Various toxicological and epidemiological studies point towards an association between trihalomethanes (THMs), one of the main DBPs and marker for total DBP load, and (low) birth weight, although the evidence is not conclusive. Administered doses in toxicological studies have been high and even though epidemiological studies have mostly shown excess risks, these were often not significant and the assessment of exposure was often limited. Some studies have shown associations for DBPs and other outcomes such as spontaneous abortions, stillbirth and birth defects, and although the evidence for these associations is weaker it is gaining weight. There is no evidence for an association between THMs and preterm delivery. The main limitation of most studies so far has been the relatively crude methodology, in particular for assessment of exposure. RECOMMENDATIONS: Large, well designed epidemiological studies focusing on well defined end points taking into account relevant confounders and with particular emphasis on exposure characterisation are ideally needed to confirm or refute these preliminary findings. In practice, these studies may be impracticable, partly due to the cost involved, but this is an issue that can be put right--for example, by use of subsets of the population in the design of exposure models. The studies should also reflect differences of culture and water treatment in different parts of the world. To identify the specific components that may be of aetiological concern and hence to fit the most appropriate exposure model with which to investigate human exposure to chlorinated DBPs, further detailed toxicological assessments of the mixture of byproducts commonly found in drinking water are also needed. PMID- 10711275 TI - Associations between ambient ozone, hydrocarbons, and childhood wheezy episodes: a prospective observational study in south east London. AB - OBJECTIVES: To explore the hypothesis that hydrocarbon species and other air pollutants which accumulate at low and high concentrations of ozone are more directly associated with childhood wheezy episodes than ozone. METHODS: Prospective observational study over 1 year set in the Lewisham district of south east London. The daily attendance rate of children with acute wheeze at the accident and emergency department of Lewisham Hospital was related to local measurements of ozone, hydrocarbon species, nitrogen dioxide (NO2), sulphur dioxide (SO2) and small particulate matter with diameter < 10 microns (PM10). RESULTS: An inverse relation was found between the air pollutants and ozone. After seasonal and meteorological adjustment a non-linear U shaped trend was found between incidence of wheeze and ozone. The trend was significant in children < 2 years of age but not in older children. In the younger age group, after adjustment for season, temperature, wind speed, and respiratory infection, the incidence relative to that at the mean daily ozone concentration of 32.7 micrograms/m3, was estimated to increase by 65% (95% confidence interval (95% CI) 22% to 122%) at an ozone concentration of 5 micrograms/m3 (1.5 SDs below the mean) and by 63% (95% CI -6% to 184%) at 80 micrograms/m3 (2.5 SDs above the mean). For several hydrocarbons there were significant positive linear relations found, again in children < 2 years of age but not older children. For benzene, the incidence increased by 8% (95% CI 2 to 13%) per SD (SD 2.8 micrograms/m3) increase in benzene concentration. A same day association between incidence and ozone was found to be the most significant but for other pollutants a lag of 2 days gave the most significant associations. No significant association was found for the non-hydrocarbon pollutants including SO2, NO2, and PM10. CONCLUSIONS: A U shaped relation was found between ozone and the incidence of wheezy episodes in young children. Certain hydrocarbon pollutants accumulate in the atmosphere when ozone concentrations are low, and are associated with childhood wheezy episodes. However, the U shaped association of ozone on incidence cannot be explained by these other pollutants. The finding supports an earlier finding that incidences of wheeze are at a minimum when ozone concentrations are 30-40 micrograms/m3. PMID- 10711276 TI - Risk of mortality, cancer incidence, and stroke in a population potentially exposed to cadmium. AB - OBJECTIVES: To follow up mortality and cancer incidence in a cohort potentially exposed to cadmium and to perform a geographical (ecological) analysis to further assess the health effects of potential exposure to cadmium. METHODS: The English village of Shipham has very high concentrations of cadmium in the soil. A previous cohort study of residents of Shipham in 1939 showed overall mortality below that expected, but a 40% excess of mortality from stroke. This study extends the follow up of the cohort for mortality to 1997, and includes an analysis of cancer incidence from 1971 to 1992, and a geographical study of mortality and cancer incidence. Standardised mortality and incidence ratios (SMRs and SIRs) were estimated with regional reference rates. Comparisons were made with the nearby village of Hutton. RESULTS: All cause cohort mortality was lower than expected in both villages, although there was excess cancer incidence in both Shipham (SIR 167, 95% confidence interval (95% CI) 106 to 250) and Hutton (SIR 167, 95% CI 105 to 253). There was an excess of mortality from hypertension, cerebrovascular disease, and nephritis and nephrosis, of borderline significance, in Shipham (SMR 128, 95% CI 99 to 162). In the geographical study, all cause mortality in Shipham was also lower than expected (SMR 84, 95% CI 71 to 100). There was an excess in genitourinary cancers in both Shipham (SIR 160, 95% CI 107 to 239) and Hutton (SIR 153, 95% CI 122 to 192). CONCLUSION: No clear evidence of health effects from possible exposure to cadmium in Shipham was found despite the extremely high concentrations of cadmium in the soil. PMID- 10711277 TI - Requirements for occupational medicine training in Europe: a Delphi study. AB - OBJECTIVES: To identify the common core competencies required for occupational physicians in Europe. METHOD: A modified Delphi survey was conducted among members of the European Association of Schools of Occupational Medicine (EASOM), the Occupational Medicine Section of the Union of European Medical Specialities (UEMS), and of the European Network of Societies of Occupational Physicians (ENSOP). An initial questionnaire based on the training syllabus of the United Kingdom Faculty of Occupational Medicine was circulated and respondents were asked to rate the importance of each item. The results were discussed at a conference on the subject of competencies. A further questionnaire was developed and circulated which asked respondents to rank items within each section. RESULTS: There was a 74% response in the first round and an 80% response in the second. Respondents' ratings from most important to least important were; occupational hazards to health, research methods, health promotion, occupational health law and ethics, communications, assessment of disability, environmental medicine, and management. In the second round, among those topics ranked most highly were; hazards to health and the illnesses which they cause, control of risks, and diagnoses of work related ill health. Topics such as principles of occupational safety and selection of personal protection equipment were of least importance. Although the assessment of fitness was regarded as important, monitoring and advising on sickness absence were not highly rated. Management competency was regarded as of low importance. CONCLUSION: This survey identified that respondents had traditional disease focused views of the competencies required of occupational physicians and that competencies were lagging behind the evolving definition of occupational health. PMID- 10711278 TI - A further cohort study of workers employed at a factory manufacturing chemicals for the rubber industry, with special reference to the chemicals 2 mercaptobenzothiazole (MBT), aniline, phenyl-beta-naphthylamine and o-toluidine. AB - OBJECTIVES: To investigate mortality and cancer morbidity in workers from a factory manufacturing chemicals for the rubber industry. METHODS: The mortality (1955-96) and cancer morbidity experience (1971-92) of a cohort of 2160 male production workers from a chemical factory in north Wales were investigated. All subjects had at least 6 months employment at the factory and some employment in the period 1955-84. Detailed job histories were abstracted from company computerised records and estimates of individual cumulative exposure to 2 mercaptobenzothiazole (MBT) and its derivatives were obtained, with a job exposure matrix derived by a former factory hygienist. Durations of employment in the aniline, phenyl-beta-naphthylamine (PBN) and o-toluidine departments were also calculated. Two analytical approaches were used, indirect standardisation and Poisson regression. RESULTS: Based on serial rates for the general population of England and Wales, observed mortality for the total cohort was close to expectation for all causes (observed (obs) deaths 1131, expected (exp) deaths 1114.5, standardised mortality ratio (SMR) 101), and for all cancers (obs 305, exp 300.2, SMR 102). There was a significant (p < 0.05) excess mortality from cancer of the bladder in the 605 study subjects potentially exposed to one or more of the four chemicals being investigated (obs 9, exp 3.25, SMR 277, 95% confidence interval (95% CI) 127 to 526). This excess was dependent primarily on deaths occurring > 20 years after first exposure in those who started employment before 1955 (obs 7, exp 1.25, SMR 560, 95% CI 225 to 1154, p < 0.001). There were 30 subjects in the total study cohort who, on the basis of death certificates or cancer registration particulars, had had malignant bladder cancer. In separate analyses of the four exposure history variables (after adjustment for age), Poisson regression showed significant positive trends for risk of notification of bladder cancer increasing with cumulative duration of employment in the PBN (p < 0.001) and o-toluidine departments (p < 0.01); similar findings were not obtained for cumulative exposure to MBT or for duration of employment in the aniline department. In a simultaneous analysis of all four chemical exposure variables, a significant positive trend remained for duration of employment with exposure to PBN (p < 0.05). Further analyses of all cases of bladder cancer (malignant and benign diagnoses) used employment histories lagged by 15 years; similar findings were obtained. CONCLUSIONS: It seems likely that some members of this cohort have had occupational bladder cancer. Confident interpretation is difficult because of small numbers in the exposed subcohorts, relatively crude measures of exposure assessment for the four chemicals under study, and presence of unconsidered potential chemical confounders. The simplest interpretation of the findings about bladder cancer may be that PBN (or a chemical reagent or chemical intermediate associated with its production at this factory in the 1930s and 1940s) is a bladder carcinogen. Priority should be given, however, to obtaining information on the cancer experience of other working populations exposed to PBN or to o toluidine. PMID- 10711279 TI - The role of physical and psychological factors in occupational low back pain: a prospective cohort study. AB - OBJECTIVE: To examine risk factors for onset of low back pain (LBP) in healthcare workers. METHODS: Nursing students, during their 3 year training period, and 1 year after training were studied in a prospective cohort study, with repeated self reported measurements of determinants of LBP at 6 monthly intervals for 3 years during training, and after a 12 month interval there was an additional final follow up. RESULTS: During training, increased risk of new episodes of LBP was associated with having had LBP at baseline, with part time work, and with a high score on the general health questionnaire (GHQ). A high GHQ score preceded the onset of LBP, in such a way that a high score at the immediately previous follow up increased risk of LBP at the next follow up. 12 Months after training, a history of recurring LBP during training increased the risk of a new episode as did having obtained work as a nurse. A high GHQ score at this follow up was also associated with a concurrently increased risk. Pre-existing GHQ score, either at the end of training or at baseline, had no effect on risk of LBP 12 months after training. CONCLUSIONS: Other than a history of LBP, pre-existing psychological distress was the only factor found to have a pre-existing influence on new episodes of LBP. Increased levels of psychological distress (as measured by the GHQ) preceded the occurrence of new episodes of pain by only short intervening periods, implying a role for acute distress in the onset of the disorder. This finding suggests that management of the onset of occupational LBP may be improved by management of psychological distress. PMID- 10711280 TI - Risk of enzyme allergy in the detergent industry. AB - OBJECTIVES: To assess the prevalence of enzyme sensitisation in the detergent industry. METHODS: A cross sectional study was conducted in a detergent factory. Sensitisation to enzymes was examined by skin prick and radioallergosorbent (RAST) tests. 76 Workers were tested; 40 in manufacturing, packing, and maintenance, and 36 non-exposed people in management and sales departments. The workers were interviewed for work related respiratory and skin symptoms. Total dust concentrations were measured by a gravimetric method, and the concentration of protease in air by a catalytic method. RESULTS: Nine workers (22%) were sensitised to enzymes in the exposed group of 40, whereas none were sensitised in the non-exposed group. All the sensitised people had symptoms at work; all had rhinitis and one had asthma. Protease concentrations were generally < 20 ng/m3, but occasional peak values up to 80 ng/m3 were detected in the packing and maintenance tasks, and high values of > 1 microgram/m3 in the mixing area. CONCLUSION: Despite the use of encapsulated enzyme preparations, high enzyme concentrations in workplace air are possible, resulting in a higher risk of sensitisation than expected. PMID- 10711281 TI - Performance of population specific job exposure matrices (JEMs): European collaborative analyses on occupational risk factors for chronic obstructive pulmonary disease with job exposure matrices (ECOJEM). AB - OBJECTIVES: To compare the performance of population specific job exposure matrices (JEMs) and self reported occupational exposure with data on exposure and lung function from three European general populations. METHODS: Self reported occupational exposure (yes or no) and present occupation were recorded in the three general population surveys conducted in France, The Netherlands, and Norway. Analysis was performed on subjects, aged 25-64, who provided good forced expiratory volume in 1 second (FEV1) tracings and whose occupations were performed by at least two people, in the French (6217 men and 5571 women), the Dutch (men from urban (854) and rural (780) areas), and the Norwegian (395 men) surveys. Two population specific JEMs, based on the percentage of subjects who reported themselves exposed in each job, were constructed for each survey and each sex. The first matrix classified jobs into three categories of exposure according to the proportion of subjects who reported themselves exposed in each job (P10-50 JEM, low < 10%, moderate 10-49%, high > or = 50%). For the second matrix, a dichotomous variable was constructed to have the same statistical power as the self reported exposure--that is, the exposure prevalence (p) was the same with both exposure assessment methods (Pp JEM). Relations between occupational exposure, as estimated by the two JEMs and self reported exposure, and age, height, city, and smoking adjusted FEV1 score were compared. RESULTS: Significant associations between occupational exposure estimated by the population specific JEM and lung function were found in the French and the rural Dutch surveys, whereas no significant relation was found with self reported exposure. In populations with few subjects in most jobs, exposure cannot be estimated with sufficient precision by a population specific JEM, which may explain the lack of relation in the Norwegian and the Dutch (urban area) surveys. CONCLUSION: The population specific JEM, which was easy to construct and cost little, seemed to perform better than crude self reported exposures, in populations with sufficient numbers of subjects per job. PMID- 10711282 TI - Lung function, biological monitoring, and biological effect monitoring of gemstone cutters exposed to beryls. AB - OBJECTIVES: Gemstone cutters are potentially exposed to various carcinogenic and fibrogenic metals such as chromium, nickel, aluminium, and beryllium, as well as to lead. Increased beryllium concentrations had been reported in the air of workplaces of beryl cutters in Idar-Oberstein, Germany. The aim of the survey was to study the excretion of beryllium in cutters and grinders with occupational exposure to beryls--for example, aquamarines and emeralds--to examine the prevalence of beryllium sensitisation with the beryllium lymphocyte transformation test (BeLT), to examine the prevalence of lung disease induced by beryllium, to describe the internal load of the respective metals relative to work process, and to screen for genotoxic effects in this particular profession. METHODS: In a cross sectional investigation, 57 out of 100 gemstone cutters working in 12 factories in Idar-Oberstein with occupational exposure to beryls underwent medical examinations, a chest radiograph, lung function testing (spirometry, airway resistance with the interrupter technique), and biological monitoring, including measurements of aluminium, chromium, and nickel in urine as well as lead in blood. Beryllium in urine was measured with a newly developed direct electrothermal atomic absorption spectroscopy technique with a measurement limit of 0.06 microgram/l. Also, cytogenetic tests (rates of micronuclei and sister chromatid exchange), and a BeLT were performed. Airborne concentrations of beryllium were measured in three factories. As no adequate local control group was available, the cutters were categorised into those with an exposure to beryls of > 4 hours/week (group A) and < or = 4 hours/week (group B). RESULTS: Clinical, radiological, or spirometric abnormalities indicating pneumoconiosis were detected in none of the gemstone cutters. Metal concentrations in biological material were far below the respective biological limit values, and beryllium in urine was only measurable in subjects of group A. Cytogenetic investigations showed normal values which were independent of the duration of beryllium exposure. In one subject, the BeLT was positive. Beryllium stimulation indices were significantly higher in subjects with detectable beryllium in the urine than in those with beryllium concentrations below the detection limit (p < 0.05). In one factory, two out of four measurements of airborne beryllium concentrations were well above the German threshold limit value of 2 micrograms/m3 (twofold and 10-fold), and all gemstone cutters working in this factory had measurable beryllium concentrations in urine. CONCLUSION: No adverse clinical health effects were found in this cross sectional investigation of gemstone cutters working with beryls. However, an improvement in workplace hygiene is recommended, accompanied by biological monitoring of beryllium in urine. PMID- 10711283 TI - Half life of chromium in serum and urine in a former plasma cutter of stainless steel. AB - For 8 years chromium in serum and urine has been followed up in a former plasma cutter of stainless steel who was exposed to airborne dust and fumes containing chromium during this work. After the first examination for serum chromium the exposure ended. Serum chromium concentration has been measured seven times during the period and was initially very high and has subsequently dropped slowly. The half life was 40 months in serum. Urinary chromium has been measured five times. The half life was 129 months in urine. The study shows that exposure to airborne dust and fumes containing chromium may cause accumulation of chromium in the body, and that when exposure ends, elimination of chromium is very slow. Previous studies suggest that chromium mainly accumulates in the lungs. PMID- 10711284 TI - Update of the Texaco mortality study 1947-93: Part II. PMID- 10711285 TI - Childhood leukaemia, population mixing, and paternal occupation. PMID- 10711286 TI - CARJ in the new millennium. PMID- 10711287 TI - Canadian versus US radiology certification examinations: the 1999 experience. PMID- 10711288 TI - Stereotaxic core needle biopsy of breast microcalcifications obtained using a standard mammography table with an add-on unit. AB - OBJECTIVE: To demonstrate the reliability of stereotaxic biopsy of indeterminate microcalcifications using a standard mammography table with an add-on unit. METHODS: In 121 cases of indeterminate microcalcifications, core biopsy was performed using a standard mammography table with an add-on stereotaxic unit. Microcalcifications were identified on radiography of core specimens. RESULTS: Microcalcifications and a definitive histologic diagnosis were obtained in 112 core biopsies (92.6%), with no significant complications. In 23 lesions frank malignancy was diagnosed, and all of these diagnoses were confirmed on surgery. Pathologic examination suggested carcinoma in 4 lesions, and open biopsy confirmed malignancy in 3 of these cases. Four lesions showed atypical ductal hyperplasia. Benign disease was diagnosed in 81 lesions, of which 78 remained stable on mammographic follow-up (mean 16 months later) and 3 were subjected to surgical biopsy (of which 1 was malignant and 2 were benign). Nine cases were technically unsatisfactory because microcalcifications were not sampled. CONCLUSION: Stereotaxic core biopsy performed with an add-on unit is a safe and reliable technique for biopsy of indeterminate microcalcifications. For successful biopsy, microcalcifications must be harvested. Pathologic results should be correlated with mammographic findings. The accuracy rate compares favourably with results reported using prone biopsy tables. In an era of cost containment, this alternative to prone biopsy tables could result in significant savings in terms of capital investment and use of hospital rooms. In this study, surgical biopsy could have been avoided in 64.5% of cases. PMID- 10711289 TI - Biventricular heart failure secondary to a pericardial cystic mass: case report. PMID- 10711290 TI - Cecocutaneous fistula after remote appendectomy: case report. PMID- 10711291 TI - Pelvic varices diagnosed with endorectal surface coil magnetic resonance imaging: case report. PMID- 10711292 TI - Magnetic resonance imaging for differentiating delayed-onset sterile abscess complicating vaccination from soft-tissue neoplasm: case report. PMID- 10711293 TI - Percutaneous retrieval of a vena cava filter from the right atrium: case report. PMID- 10711294 TI - Residents' corner. Answer to case of the month #69. Wernicke encephalopathy: an unusual presentation, with classic magnetic resonance imaging findings. PMID- 10711295 TI - Residents' corner. Answer to case of the month #70. Ovarian vein thrombosis. PMID- 10711296 TI - Residents' corner. Answer to case of the month #71. Pulmonary alveolar proteinosis. PMID- 10711297 TI - Residents' corner. Answer to case of the month #72. Congenital bronchial atresia. PMID- 10711298 TI - Residents' corner. Answer to case of the month #73. Hodgkin's disease with pulmonary parenchymal involvement. PMID- 10711299 TI - Residents' corner. Answer to case of the month #74. Crohn's disease: an unusual presentation. PMID- 10711300 TI - Prevention of coronary heart disease. Part I. Primary prevention. AB - The first concern in primary prevention is the physician's belief that primary prevention is important for all adults and that intervention can significantly affect risk. Given the coronary plaque burden over many years and the importance of the development of healthy lifestyles early in adulthood to decrease coronary plaque burden, there are excellent reasons to begin prevention even with young adults. At the very least, a patient seen for any reason should provide a smoking history, have knowledge of the presence of early CHD in first-degree relatives and measurements of blood pressure, height, and weight, provide evidence for a cholesterol level within 5 years (after age 20 according to NCEP guidelines or in middle age according to ACP guidelines), and be given an assessment of glucose tolerance or diabetes. Information about alcohol intake and physical activity status are also of some importance. Other than height, weight, and blood pressure, during the physical examination, the physician should initially assess the strength of pulses in the lower extremities, evidence for carotid or femoral bruits, and eyegrounds for retinal arterial changes, and the skin and subcutaneous tissue should be examined for xanthomas and the eyes should be examined for corneal arcus and xanthelesma. These elements should be part of any initial examination by a primary care physician and are not extraordinary. In addition to lipid and blood sugar analyses, other evaluations may include blood urea nitrogen and creatinine and electrolytes in patients with hypertension or diabetes or in patients who are on antihypertensive agents. It may be prudent to obtain an ECG for patients who are older than 40 years. The elements mentioned above are the elements of the history, physical examination, and laboratory examination in subjects without a past history of CHD and with no clinical evidence for CHD. Primary prevention management begins with a discussion of risk factors with the patient. The key interventions aim at the lowering of blood pressure to at least less than 140/90 mm Hg, the complete cessation of smoking, the lowering of lipid levels to less than 130 mg/dL, the lowering of triglycerides to less than 200 mg/dL (or, some would argue, < 150 mg/dL), and the attempt to keep HDL cholesterol above 35 mg/dL (more than 40 to 45 mg/dL is a better goal) with the use of lifestyle modification. For patients with diabetes, strict control of glucose levels is essential to minimize disease of the microvasculature and possibly to minimize progressive renal disease. There are several lifestyle modifications for lipids. For patients with elevated LDL cholesterol, modifications include a less than 30% fat calorie diet and less than 300 mg of cholesterol intake daily, with fat calories approximately equally distributed among saturated fats, polyunsaturated fats, and monounsaturated fats (1/3, 1/3, 1/3; rule of 3s). The assistance of a dietician is extremely helpful in this regard. For patients with a low HDL cholesterol, weight reduction (for overweight patients) by calorie control and increased physical activity and smoking cessation will have some modest effect. For patients with elevated triglycerides, a diet similar to that for lowering of LDL cholesterol with the addition of stricter calorie limitation, avoidance of refined sugars, increase in complex carbohydrates, and avoidance of alcohol will be helpful. A decrease in the percent of fat calories to 20% to 25% will be of assistance to those patients with particularly high triglycerides. The treatment of underlying conditions such as diabetes mellitus, hypothyroidism, liver disease, and some renal conditions may also significantly modify high triglycerides. For patients with hypertension, limitation of sodium to 2 gm/d (6 gm sodium chloride), limitation of alcohol to 1 to 2 drinks a day, increased physical activity, and weight reduction are the key lifestyle modifications. (ABSTRACT TRUNCATED) PMID- 10711301 TI - Nursing home 'abuse' litigation is instructive. PMID- 10711302 TI - Valproic acid dosing recommendations questioned. PMID- 10711303 TI - Fighting frailty. Prescription for healthier aging includes exercise, nutrition, safety, and research. PMID- 10711304 TI - Geriatric photo quiz. Aneurysm. Cardiovascular problem can present in various locations and requires diligent management. PMID- 10711305 TI - Acute MI. Age-related presentations and treatment options. AB - At least 60% of all acute myocardial infarctions (AMIs) occur in patients age 65 and older and about one-third in patients older than age 75. The presentation of AMI is modified by age-related changes in endothelial function, smooth muscle cell activity, diastolic function, and responses to circulating catecholamines. Atypical presentations are common in the older patient and require the physician to maintain a high index of suspicion for AMI. ECG findings of ST-segment depression make many older patients ineligible for reperfusion by thrombolytic therapy. Acute and post-MI medical therapies are underused in the older population and should be individualized, based on the presence of comorbid conditions. PMID- 10711306 TI - Elder abuse. Using clinical tools to identify clues of mistreatment. AB - Elder abuse occurs most commonly in residential rather than institutional settings, and the most likely perpetrators are known by the victim. Although a defined set of risk factors has not been developed, careful questioning and assessment can help determine whether a patient is at increased risk. The common types of elder maltreatment include caregiver and self-neglect, emotional and psychological abuse, fiduciary exploitation, and physical abuse. Assessment consists of comprehensive physical examination, including scrutiny of the musculoskelatal and genitourinary systems, neurologic and cognitive testing, and detailed social and sexual histories. Clues that cannot be explained medically may signal elder abuse. To properly intervene, clinicians should be familiar with state laws governing reporting procedures and patient privacy. PMID- 10711307 TI - Avoiding adverse reactions. Effective lower-dose drug therapies for older patients. AB - Adverse drug reactions (ADRs) are a leading cause of morbidity and mortality, with the highest incidence occurring in patients age 60 and older. Prescribing the lowest effective doses of medications to older patients can help avoid ADRs, minimize bothersome side effects, and increase rates of compliance. For many drugs, clinical experience and published studies demonstrate the effectiveness of doses substantially lower than those recommended in standard references. This article examines the problem of ADRs in older patients, discusses pharmacokinetic data regarding older versus younger adults, and provides effective lower doses for many common drugs. PMID- 10711308 TI - Clinical practice guidelines. Review of the recommendations for colorectal screening. AB - Data suggest that vigilant screening and polyp removal can help reduce mortality caused by colorectal cancer (CRC), the incidence of which increases with age. Although several screening methods are available, wide variation exists among them in accuracy, effectiveness of identifying cancerous lesions, potential complications, costs, ease of administration, and patient compliance. Moreover, a lack of direct evidence demonstrating the long-term benefits (compared with the costs and risks) of the individual screening methods complicates the decision of when and whether to screen. In 1994, an expert, multidisciplinary panel was convened to review the evidence and produce recommendations to help guide clinicians and patients with regard to CRC screening and surveillance. The resulting clinical practice guidelines, released in 1997, are examined. PMID- 10711309 TI - Itchy papules on the arms and neck. Lesions develop on sites that are easy for the patient to reach and scratch. PMID- 10711310 TI - Resolution of drug-induced agranulocytosis. PMID- 10711311 TI - What money can't buy: the bad policy of substituting fixed payments for employer sponsored health insurance. PMID- 10711312 TI - Do HMOs make a difference? Introduction. AB - The study presented in this and the following five papers analyzes how health maintenance organizations (HMOs) affect privately insured individuals' access to health care, use of services, and assessments of care. Using a common data source and methodology, the study examines differences in a broad range of measures between HMOs and other types of insurance, controlling for health status and an extensive set of other individual characteristics and market location. HMO/non HMO differences also are examined across population subgroups defined by health status, income, race, and age. Data come from the Community Tracking Study Household Survey, a recent, large national survey. Findings show that a person's type of health insurance coverage has little effect on the likelihood of unmet or delayed needs for medical care in the aggregate, but the types of access problems faced by HMO and non-HMO enrollees differ. HMO enrollees are less likely to face financial barriers to care, but more likely to face barriers related to the organization of care delivery. HMO enrollees use more ambulatory and preventive care, but results show no differences in hospital, surgery, and emergency room use. Compared with other types of insurance, physician visits under HMOs are more likely to be to primary care physicians than to specialists. Finally, across nearly all measures of patients' satisfaction, ratings of their last doctor's visit, and trust in their physicians, HMO enrollees' assessments of care are lower than those of people not in HMOs. Across all measures, the study finds few subgroup differences. PMID- 10711313 TI - Do HMOs make a difference? Data and methods. AB - This paper describes the common data source and methods used in this study. Data come from the Community Tracking Study Household Survey, a nationally representative survey of individuals conducted in 1996-1997. Focusing on the privately insured, nonelderly population, the study examines the effect of health maintenance organizations (HMOs) on access, service use, and consumer assessments, as well as how these effects differ across population subgroups. Multivariate models control for population characteristics and location differences between HMO and non-HMO enrollees. Tests for endogeneity of plan type (selection bias) indicated that this did not pose a threat to the analysis. PMID- 10711314 TI - Do HMOs make a difference? Access to health care. AB - This analysis examines the effects of health maintenance organizations (HMOs) on access to care among the privately insured, nonelderly population. After controlling for population and location differences, HMO and non-HMO enrollees differ little in reports of unmet or delayed care needs. Yet type of insurance affects the source of access problems. HMO enrollees face lower financial barriers to care and are more likely to report a regular source of care than those enrolled in other types of insurance, but they are more likely to report access problems related to the organization of care delivery. PMID- 10711315 TI - Do HMOs make a difference? Use of health services. AB - This study analyzes the effects of health maintenance organizations (HMOs) on the use of health services by the privately insured, nonelderly population. After controlling for population and location differences, HMOs increase physician visits, nonphysician practitioner visits, and total ambulatory visits by modest but significant margins, while shifting the mix of physician care from specialists to primary care physicians. HMOs also increase use of two preventive services: mammography screening and flu shots. Contrary to expectation, however, the study finds no significant differences between HMO and non-HMO enrollees in the use of hospital, surgery, and emergency room services. PMID- 10711316 TI - Do HMOs make a difference? Consumer assessments of health care. AB - This study examines the effects of health maintenance organizations (HMOs) on consumer assessments of health care among the privately insured, nonelderly population. After controlling for population and location differences, the study finds that HMO enrollees are less likely than those in non-HMOs to be satisfied with their care, to rate their last medical visit highly, and to express trust in their physicians. One exception is a finding of little or no statistically significant difference between HMO and non-HMO enrollees in the likelihood of distrust that a physician may provide unnecessary services. PMID- 10711317 TI - Do HMOs make a difference? Summary and implications. AB - The findings of this study of the effects of health maintenance organizations (HMOs) have implications for consumers' choice between HMOs and other types of insurance: consumers face a trade-off that flows in part from the design of HMOs. HMO enrollees get more primary and preventive care and face lower out-of-pocket costs, but they get less specialist care, experience more provider access and organizational barriers to care, and report less satisfaction, lower ratings of care, and less trust in their physicians. Policymakers should recognize that this trade-off will be attractive to some people but not to others. PMID- 10711318 TI - Capitated contracting of integrated health provider organizations. AB - This paper examines global capitation of integrated health provider organizations that link physicians and hospitals, such as physician-hospital organizations and management service organizations. These organizations have proliferated in recent years, but their contracting activity has not been studied. We develop a conceptual model to understand the capitated contracting bargaining process. Exploratory multivariate analysis suggests that global capitation of these organizations is more common in markets with high health maintenance organization (HMO) market share, greater numbers of HMOs, and fewer physician group practices. Additionally, health provider organizations with more complex case mix, nonprofit status, more affiliated physicians, health system affiliations, and diversity in physician organizational arrangements are more likely to have global capitation. Finally, state regulation of provider contracting with self-insured employers appears to have spillover effects on health plan risk contracting with health providers. PMID- 10711319 TI - Access to physicians' services for vulnerable Medicare beneficiaries. AB - This article examines whether changes in physician reimbursement under the Medicare Fee Schedule (MFS) had differential impacts on access to care for vulnerable and nonvulnerable Medicare beneficiaries. The quasi-experimental research design takes advantage of cross-sectional differences in the magnitude of the MFS impact on payments. We selected a stratified random sample to ensure adequate representation of vulnerable group members and constructed service specific measures of the MFS payment change. While we found few effects on access attributable to the MFS, we did find substantial utilization gaps between vulnerable and nonvulnerable subpopulations for primary care services, as well as for high-cost procedures during episodes of care for acute myocardial infarctions. PMID- 10711320 TI - A management tool for controlling the rate of non-acceptable inpatient hospital claims. AB - This paper demonstrates a tool for substantially improved monitoring of the validity of health insurance claims. Using a Bayesian regression model, we predict the probability of a non-acceptable claim (NAC) for each claim record and the expected number of NACs for any set of claims. When applied to a large set of hospital discharge claims, the tool shows a substantial improvement in the ability to estimate the actual number of NACs in the set. The tool permits ongoing monitoring of claims, more precise control, and a substantial reduction in audit cost in claims administration. It is conceptually applicable to other ongoing quality control systems, where inexpensively obtained information can be used to predict events that are costly to measure directly. PMID- 10711321 TI - Sliding-scale premium health insurance programs: four states' experiences. AB - As publicly funded health insurance shifts more toward coverage of working families of low and moderate incomes, there has been growing interest in beneficiary cost sharing, in the form of sliding-scale premiums. In the 1990s, Hawaii, Minnesota, Tennessee, and Washington initiated expansion programs that used sliding-scale premiums for working-class families. The experience in these states indicates that it is feasible to require cost sharing of premiums, but there are a number of design and operational complexities. A preliminary analysis indicates that, as expected, higher out-of-pocket premium shares were associated with lower participation rates. PMID- 10711322 TI - The effect of a closed formulary on prescription drug use and costs. AB - This study examines the effect of a closed formulary on pharmaceutical use and spending. We compared an employer plan that implemented a closed formulary in July 1997 to a control group with an open formulary, for the nine months preceding and following implementation of the formulary. When controlling for age, gender, and chronic disease score, the closed formulary was associated with significantly lower increases in utilization and expenditures, a higher prior authorization rate, and a reduced rate of continuation with chronic medications in the nine months following its implementation. These findings have implications for the design of prescription drug benefits. PMID- 10711323 TI - Health risk of low-dose pesticides mixtures: a review of the 1985-1998 literature on combination toxicology and health risk assessment. AB - A literature review covering the last 14 yr has been performed in the field of combination toxicology and human risk assessment from exposure to chemical mixtures, with special emphasis on mixtures of pesticides at low doses, that is, at levels likely to occur in human diet and environment. Despite a large body of knowledge in the field of risk assessment methodologies for exposure to chemical and pesticide mixtures, there is no single methodological approach in "combination toxicology" and health risk assessment of chemical mixtures, and therefore professional judgment is still required. Generally, the dose or response additivity approach that may be applied to evaluate potential risk for chemical mixtures in human toxicology overestimates the risk of a combination of chemicals. The recent endocrine disrupter issue demonstrated the difficulty of reproducibility of data when testing environmental toxicants at very low levels, and the need for more basic work in this field. The use of integrated methodological approaches may provide more reliable predictive data in the risk assessment of chemical mixtures in future. Yet data have demonstrated that exposure to a combination of compounds does not cause effects stronger than the ones of their most active component, provided components are present at low concentration levels, like acceptable daily intake (ADI) or reference dose (RfD) levels, well below their respective no-observed-adverse-effect levels (NOAELs). Although it has been demonstrated that a combination of compounds with the same target organ and the same or very similar mechanisms of action may cause additive or synergistic effects, the chance of such effects will most likely diminish with decreasing exposure levels to such combinations. Synergism and antagonism may both occur at the same time at different organs or targets in the same organism. However, and despite some exceptions, it has been demonstrated that interaction between components is not a common event at low levels of human exposure such as those that may occur through pesticides residues in food or drinking water. The introduction of a special safety factor as a standard for mixtures in addition to those normally used for deriving ADIs, RfDs, or minimal risk levels is not supported by data. It can be concluded from our review that, as a general rule, exposure to mixtures of pesticides at low doses of the individual constituents does not represent a potential source of concern to human health. PMID- 10711324 TI - Molecular mechanisms regulating iNOS expression in various cell types. AB - Inducible nitric oxide synthase (iNOS) has been shown to be present in a variety of cell types, and nitric oxide (NO) has been implicated in a multitude of biological functions. The purpose of this review is twofold: (1) to provide a comprehensive table of cell types that produce NO together with the effects of agents used to study iNOS regulation, as a ready reference for the investigators in the field; and (2) to summarize recent observations dealing with iNOS signal transduction mechanisms. Initially, the major regulation of NO production was believed to occur at the transcription step, but now it is recognized that NO regulation can occur at the transcriptional, posttranscriptional, translational, and posttranslational level. There have been a number of studies of the regulation of iNOS in various cell types, often yielding conflicting results. The major emphasis of this review is on iNOS signal transduction mechanisms. For example, the role of JAK kinases and mitogen-activated protein (MAP) kinases in iNOS regulation is elaborated. In addition, species differences in the iNOS promoter region and the role of RNA structure in iNOS expression is discussed. The role MAP kinases play in translational regulation in addition to transcriptional regulation is emphasized. An analysis of the current data and suggestions for future studies are also presented. PMID- 10711325 TI - Perceptual and physiologic effects of histamine challenge on nasal breathing. AB - The purpose of this study is to determine the effect of histamine-induced nasal congestion on nasal airflow and the perception of externally applied resistance to nasal breathing. Nasal cross-sectional area and nasal airflow during free breathing were measured in 15 adult subjects before and after histamine challenge. The threshold for perception of resistance to nasal breathing was determined using a dynamic perturbator device, with both free breathing and controlled nasal air-flow. The average threshold for perception of nasal resistance was 0.383 Pa/cm3/s at baseline. After histamine application, there was a significant decrease in nasal cross-sectional area (p = 0.0001), associated with a decrease in nasal airflow (r = 0.6). The average threshold of perception increased to 1.373 Pa/cm3/s (p < 0.0001). When nasal airflow was controlled at the baseline rate, the threshold of perception improved to 0.638 Pa/cm3/s (p = 0.024). These findings indicate that nasal congestion causes a reduction in both nasal airflow and the perception of resistance to nasal breathing. The ability to detect nasal airway impairment is improved with increased nasal airflow. An improved understanding of the physiology of the subjective perception of nasal patency may lead to innovative methods for the treatment of nasal obstruction. PMID- 10711327 TI - Isolated sphenoid sinus lesions. AB - Pathologic conditions involving the sphenoid sinus alone are rare. A retrospective chart review was performed of 182 cases of isolated sphenoid sinus lesions seen at the Mayo Clinic between 1935 and 1998. There were 53 cases of sinusitis, 44 mucoceles, and 15 fungus-related cases (61.5%), and the rest of the cases were divided among numerous other pathologic entities. Symptoms, differential diagnosis, and various therapeutic modalities are discussed. We believe that these data will be useful to clinicians considering multiple pathologic possibilities when faced with a lesion involving the sphenoid sinus alone. PMID- 10711326 TI - Maxillary sinus involution after endoscopic sinus surgery in a child: a case report. AB - Studies in animal models have suggested that functional endoscopic sinus surgery may affect facial skeletal growth in children, although reviews of large clinical series do not support this observation. This is a case report of a 12 year old male referred to the senior author (SBL) several months after undergoing bilateral functional endoscopic sinus surgery. The preoperative computed tomograms of the paranasal sinuses were normal with symmetrical well-developed paranasal sinuses. Postoperative computed tomography revealed nearly total involution of the osseous skeleton of the left maxillary sinus. This is the first clinical report of alterations in the facial skeleton of a child secondary to iatrogenic trauma directed at the osteomeatal complex. The case and related literature are reviewed in detail. PMID- 10711328 TI - Osteoplastic flap versus modified endoscopic Lothrop procedure in patients with frontal sinus disease. AB - The aim of the present study was to review the clinical results of osteoplastic flap procedure with abdominal fat obliteration and modified endoscopic Lothrop procedure. Charts of patients with frontal sinus disease who underwent osteoplastic flap procedure with abdominal fat obliteration or modified endoscopic Lothrop procedure were retrospectively reviewed. Forty-three patients with frontal sinus disease underwent osteoplastic flap procedure with abdominal fat obliteration. Frontal sinus disease was chronic sinusitis in 21, mucocele in 18, and papilloma in four. None of the patients had recurrence within 3 to 12 years follow-up period. Six patients had decreased forehead sensation, one had a CSF leak, and one had loss of the fat graft. Fifteen patients with chronic frontal sinusitis underwent modified endoscopic Lothrop procedure. The follow-up period ranged from 0.5 to 2.5 years. Two patients had recurrence of disease 2 and 6 months after surgery and required osteoplastic flap. In patients with chronic frontal sinusitis, both procedures achieved good relief of symptoms; however, follow-up time of modified endoscopic Lothrop procedure was smaller than that of osteoplastic flap procedure. In conclusion, osteoplastic flap procedure with abdominal fat obliteration provides successful treatment in patients with frontal chronic sinusitis, mucocele, or papilloma. Modified endoscopic Lothrop procedure achieves the relief of symptoms in patients with chronic frontal sinusitis. With the future availability of long term follow-up results, modified endoscopic Lothrop procedure may reduce the number of osteoplastic flap procedures in patients with chronic frontal sinusitis. PMID- 10711329 TI - Histopathology of tissue samples removed using the microdebrider technique: implications for endoscopic sinus surgery. AB - Microdebriders are being used with increasing frequency in endoscopic sinus surgery to provide precise removal of soft tissue and bone with simultaneous suction and irrigation. To date, no one has analyzed whether histopathology of tissue processed through a microdebrider is maintained. Fifteen tissue samples (squamous cell cancer, esthesioneuroblastoma, lymphoma, sarcoma, inverted papilloma, benign thyroid, and nasal mucosa) were processed though a microdebrider in various modes and speeds. Pathology slides were developed, coded, and presented as unknowns to the pathologist for diagnosis. Specimens taken from routine biopsy techniques were then compared to those passed through the microdebrider. The study found no significant loss of morphologic features in the tissue passed through the microdebrider. The microdebrider may be used for both routine and oncologic procedures without altering the histopathology necessary for diagnosis. PMID- 10711330 TI - Preoperative sagittal CT evaluation of the frontal recess. AB - Endoscopic surgical approaches for chronic frontal sinusitis require the reestablishment of adequate frontal sinus ventilation and drainage for relief of symptoms. After the resection of anterior ethmoid mucosal disease and cellular structure, the anterior to posterior depth of the nasofrontal beak to the base of skull at the insertion of the ethmoidal bulla (frontal sinus ostium) often represents a critical margin for functional success. However, little information concerning this dimension is available. Depending on intraoperative surgical judgment of this distance, extended endoscopic surgical procedures involving additional bone resection may be indicated. These approaches may be hazardous due to the proximity of the cranial cavity and orbit. In addition, secondary stenosis can result from the subsequent inflammatory response. Improved CT imaging, high resolution sagittal reformatting, and computer workstations provide the ability to obtain direct preoperative measurements of the frontal recess. We used a paramedian sagittal section and recorded the maximal anterior to posterior depth from the nasofrontal beak to the base of skull at the insertion of the ethmoidal bulla in 20 patients, 31 sides, undergoing primary endoscopic frontoethmoidectomy. In addition, we found a positive correlation between this distance and agger nasi air cell size measured in the same 31 sides. PMID- 10711331 TI - Nontraumatic nasal septal abscesses in the immunocompromised: etiology, recognition, treatment, and sequelae. AB - Proper management of a nasal septal abscess requires prompt diagnosis, adequate surgical drainage, and antibiotics to prevent the potentially dangerous spread of infection and the development of severe functional and cosmetic sequelae. Most septal abscesses are the result of trauma to the nose with septal hematoma and subsequent infection. We present our experience with nasal septal abscesses in five immunocompromised patients without history of nasal trauma. All patients were treated with surgical drainage and antibiotics. The infections in four patients resolved, whereas in the fifth, the infection led to death. We report these cases to depict alternate etiologies of nasal septal abscess, particularly in the immunocompromised patient. Our review illustrates the wide spectrum of disease presentation, provides treatment strategies, and emphasizes the potentially catastrophic sequelae of this disease when unrecognized. With the growing number of immunocompromised individuals, it is important to recognize the potential for immunocompromise to influence the development of septal abscess. PMID- 10711332 TI - Chondromyxoid fibroma of the nasal septum: a case report emphasizing clinical correlation. AB - Chondromyxoid fibromas are uncommon tumors most often seen in long bones of adolescent and young males. Involvement of craniofacial bones is extremely unusual, with sporadic case reports described in the literature. We describe the first case of chondromyxoid fibroma arising in the nasal septum with local destruction and expansile growth into the ethmoid bone and inferior turbinate in a 60-year-old female. The fortuitous discovery of this otherwise asymptomatic lesion and its follow-up are detailed. The literature is reviewed and salient clinical, radiographic, and pathologic correlative findings are emphasized. PMID- 10711333 TI - Treatment of the obstructive nose by CO2-laser reduction of the inferior turbinates: technique and results. AB - In our department we treat chronically enlarged inferior turbinates (not responding to adequate medical therapy) with the CO2-laser or by partial inferior turbinoplasty. In this work the technique is described and short- and long-term results are evaluated. We divided our study group into two populations: one group with a follow-up of 18 months for which we could obtain objective measurements of the nasal patency by means of a preoperative and postoperative active anterior rhinomanometry, and another group with a follow-up of more than 15 years, for which (because of the absence of a pre-operative rhinomanometry) we could only obtain a subjective evaluation of the results by means of a questionnaire. Relief of nasal obstruction was reported by 88.5% of the patients until 5 years after laser surgery on the inferior turbinates. There seems to be no functional impairment on the nasal mucosa after this kind of surgery. The advantages and drawbacks of this technique are discussed. PMID- 10711334 TI - Olfactory function evaluated by SPECT. AB - Few articles on neuroimaging techniques in the study of central and peripheral olfactory pathways are present in the literature. By Single Photon Emission Computed Tomography (SPECT), cortical perfusion increment after sensorial stimulation can be evaluated objectively. In the present research, 10 healthy adults underwent SPECT by CER.TO.96 cerebral tomograph, before and after olfactory stimulation with lavender-water. A variable degree of cortical activation was detected in all patients. Gyrus rectus (+24.5%), orbito-frontal cortex (right +26.6%, left +25.6%), and superior temporal (right +9.9%, left +5.5%) cortical areas were always activated. A slight perfusion increase was present in middle temporal (right +3.2%, left +2.1%) and parieto-occipital (right +0.4%, left +2%) regions. Five patients affected by posttraumatic anosmia were also investigated: they showed a perfusion increment markedly inferior to 0.5% in every olfactory area. SPECT is a rather diffused, easily performed technique which yields objective semi-quantitative information on brain perfusion. Hence, it can be regarded as a promising contribution in the fields of smell neurophysiology, clinical olfactometry, and medicolegal queries. PMID- 10711335 TI - Electron microscopic and functional aspects of the human vomeronasal organ. AB - The vomeronasal organ or Jacobson's organ is essential for pheromone detection and reproductive behavior in most mammals. The purpose of this article is to describe the fine structure of the adult human vomeronasal organ in 14 specimens and to discuss functional aspects. Our studies show a duct-like invagination of the epithelium, surrounded by numerous exocrine glands with short ducts; their fine structure suggests serous secretion. In the depth of the invagination, pseudostratified columnar epithelial cells are seen, with plump processes, kinocilia, and microvilli at the apical cell membrane. There are several cell types that differ regarding their organelles and electron density; the light sensory cells exhibit neurofilaments. Underneath the typical basement membrane, in the very vascular lamina propria, numerous myelinated and unmyelinated axons are present. These morphologic findings, which are unique in the human body, suggest that a chemosensory epithelium corresponding to a vomeronasal organ may exist. Its central connections and the possible functional significance for pheromone detection are unknown. Preservation of the vomeronasal organ in endonasal surgery could become important both clinically and medicolegally, should function be demonstrated in humans. PMID- 10711336 TI - Acellular dermal allograft for sellar reconstruction after transsphenoidal hypophysectomy. AB - Obliteration of the sphenoid sinus using fat is often used after transsphenoidal hypophysectomy. The morbidity of this approach includes donor site complications, fat necrosis, and delayed mucocele formation. As obliteration with fat is intended to prevent cerebrospinal fluid (CSF) leakage, an alternative for this technique would be techniques used for CSF rhinorrhea repair. Instead of sinus obliteration, these defects are repaired with fascial autografts, which are unfortunately associated with donor site complications. To avoid sinus obliteration and donor site complications, we have reconstructed the sella with acellular dermal allograft in lieu of sinus obliteration. Transsphenoidal hypophysectomy was performed under combined microscopic and endoscopic visualization. For closure, the sellar anterior wall was reconstructed with acellular dermal allograft, septal cartilage/bone autograft, and fibrin glue. The sinus mucosa was then draped over the reconstruction and held in place with microfibrillar collagen hemostat slurry. The sphenoid sinus was not obliterated. Postoperatively, all patients underwent serial nasal endoscopy. Thirteen patients underwent the procedure as described for removal of pituitary adenoma. Postoperative discomfort and pain were minimal. Intraoperative CSF leaks were identified in five patients; none of these patients experienced a postoperative CSF leak. The microfibrillar collagen hemostat was cleared by sphenoid mucociliary clearance. One patient developed acute sphenoid sinusitis several weeks after surgery; this patient did not develop meningitis. One postoperative CSF leak occurred in an obese patient, in whom an intraoperative CSF leak was not identified; this leak resolved with bedrest and delayed lumbar drainage alone. Sellar reconstruction with acellular dermal allograft may eliminate the need for sphenoid sinus obliteration after transsphenoidal hypophysectomy. Acellular dermal allograft sellar reconstruction ultimately provides for an aerated, functioning sphenoid sinus without increased CSF leak risk or potential donor site morbidity. PMID- 10711337 TI - Differential tetraethylammonium sensitivity of KCNQ1-4 potassium channels. AB - In Shaker-group potassium channels the presence of a tyrosine residue, just downstream of the pore signature sequence GYG, determines sensitivity to tetraethylammonium (TEA). The KCNQ family of channels has a variety of amino acid residues in the equivalent position. We studied the effect of TEA on currents generated by KCNQ homomers and heteromers expressed in CHO cells. We used wild type KCNQ1-4 channels and heteromeric KCNQ2/3 channels incorporating either wild type KCNQ3 subunits or a mutated KCNQ3 in which tyrosine replaced threonine at position 323 (mutant T323Y). IC50 values were (mM): KCNQ1, 5.0; KCNQ2, 0.3; KCNQ3, > 30; KCNQ4, 3.0; KCNQ2 + KCNQ3, 3.8; and KCNQ2 + KCNQ3(T323Y), 0.5. While the high TEA sensitivity of KCNQ2 may be conferred by a tyrosine residue lacking in the other channels, the intermediate TEA sensitivity of KCNQ1 and KCNQ4 implies that other residues are also important in determining TEA block of the KCNQ channels. PMID- 10711338 TI - Urothelium-derived inhibitory factor(s) influences on detrusor muscle contractility in vitro. AB - The function of the bladder urothelium in modulating contractile responses of the underlying detrusor smooth muscle to muscarinic stimulation has been examined in the pig bladder. Saturation curves for [3H]-QNB binding demonstrated a greater muscarinic receptor density in the urothelium than in the detrusor smooth muscle. The presence of an intact urothelium on isolated bladder strips inhibited contractions induced by carbachol but not KCl. Contractions of a urothelium denuded muscle strip were inhibited in the presence of a second bladder strip with an intact urothelium, but not if the second strip was denuded. The urothelium-induced inhibition of contractions was not prevented in the presence of L-NOARG, methylene blue, indomethacin, propranolol, suramin, TEA or apamin. The data suggest the presence of a diffusable, urothelium-derived inhibitory factor, which could not be identified but appears to be neither nitric oxide, a cyclo-oxygenase product, a catecholamine, adenosine, GABA nor an EDHF sensitive to apamin. PMID- 10711339 TI - Pharmacological profile of BIBN4096BS, the first selective small molecule CGRP antagonist. AB - Calcitonin gene-related peptide (CGRP) is one of the most potent endogenous vasodilators known. This peptide is increased during migraine attacks and has been implicated in the pathogenesis of migraine headache. Here we report on the first small molecule selective CGRP antagonist: BIBN4096BS. In vitro, this compound is extremely potent at primate CGRP receptors exhibiting an affinity (Ki) for human CGRP receptors of 14.4 +/- 6.3 (n = 4) pM. In an in vivo model, BIBN4096BS in doses between 1 and 30 micrograms kg-1 (i.v.) inhibited the effects of CGRP, released by stimulation of the trigeminal ganglion, on facial blood flow in marmoset monkeys. It is concluded that BIBN4096BS is a potent and selective CGRP antagonist. PMID- 10711340 TI - Changes in the cytosolic Ca2+ concentration and Ca(2+)-sensitivity of the contractile apparatus during angiotensin II-induced desensitization in the rabbit femoral artery. AB - 1. To investigate the underlying mechanism for the angiotensin II-induced desensitization of the contractile response during the prolonged stimulation of the vascular smooth muscle, we determined the effects of angiotensin-II on (1) cytosolic Ca2+ concentration ([Ca2+]i) and tension using fura-2-loaded medial strips of the rabbit femoral artery, (2) 45Ca2+ influx in ring preparations, and (3) Ca(2+)-sensitivity of the contractile apparatus in alpha-toxin permeabilized preparations. 2. In the presence of extracellular Ca2+, high concentrations of angiotensin-II elicited biphasic increases in [Ca2+]i and tension, which consisted of initial transient and subsequent lower and sustained phases. 3. The 45Ca2+ influx initially increased after the application of 10(-6) M angiotensin II, and thereafter gradually decreased. At 20 min after the application, there was a discrepancy between the level of [Ca2+]i and the extent of 45Ca2+ influx. 4. The relationships between [Ca2+]i and tension suggested that the angiotensin II-induced increase in the Ca(2+)-sensitivity of the contractile apparatus was maintained during the desensitization of smooth muscle contraction. 5. When 10( 6) M angiotensin-II was applied during the sustained phase of contraction induced by 118 mm K(+)-depolarization, at 10 min after the application, the [Ca2+]i levels were significantly lower and the tension levels were significantly higher than those prior to the application of angiotensin-II. 6. In conclusion, the decrease in [Ca2+]i, which is partially due to the inhibition of the Ca2+ influx, is mainly responsible for the desensitization evoked by high concentrations of angiotensin-II, and angiotensin-II seems to activate additional mechanisms which inhibit Ca2+ signaling during prolonged stimulation. PMID- 10711341 TI - The mechanism of the decrease in cytosolic Ca2+ concentrations induced by angiotensin II in the high K(+)-depolarized rabbit femoral artery. AB - 1. Using front-surface fluorometry of fura-2-loaded strips, and measuring the transmembrane 45Ca2+ fluxes of ring preparations of the rabbit femoral artery, the mechanism underlying a sustained decrease in the cytosolic Ca2+ concentration ([Ca2+]i) induced by angiotensin II (AT-II) was investigated. 2. The application of AT-II during steady-state 118 mM K(+)-induced contractions caused a sustained decrease in [Ca2+]i following a rapid and transient increase in [Ca2+]i, while the tension was transiently enhanced. 3. When the intracellular Ca2+ stores were depleted by thapsigargin, the initial rapid and transient increase in [Ca2+]i was abolished, however, neither the sustained decrease in [Ca2+]i nor the enhancement of tension were affected. 4. Depolarization with 118 mM K+ physiological salt solution containing 1.25 mM Ba2+ induced a sustained increase in both the cytosolic Ba2+ concentration ([Ba2+]i) level and tension. However, the application of 10(-6) M AT-II during sustained Ba(2+)-contractions was found to have no effect on [Ba2+]i, but it did enhance tension. 5. After thapsigargin treatment, AT-II neither decreased nor increased the enhanced Ca2+ efflux rate induced by 118 mM K(+)-depolarization, whereas AT-II did increase the enhanced 45Ca2+ influx and the 45Ca2+ net uptake induced by 118 mM K(+)-depolarization. 6. Pretreatment with calphostin-C, partially, but significantly inhibited the decrease in [Ca2+]i induced by AT-II. 7. These findings therefore suggest that AT II stimulates Ca2+ sequestration into the thapsigargin-insensitive Ca2+ stores, and thus induces a decrease in [Ca2+]i in the high external K(+)-stimulated rabbit femoral artery. PMID- 10711343 TI - Gamma-Aminobutyric acid (GABA) transport across human intestinal epithelial (Caco 2) cell monolayers. AB - 1. Transintestinal absorption of gamma-aminobutyric acid (GABA) via a pH dependent mechanism is demonstrated in the model human intestinal epithelial cell line Caco-2. 2. Experiments with BCECF [2',7',-bis(2-carboxyethyl)-5(6)- carboxyfluorescein]-loaded Caco-2 cells demonstrate that GABA transport across the apical membrane is coupled to proton flow into the cell. 3. Short-circuit current (ISC) measurements using Caco-2 cell monolayers under voltage-clamped conditions demonstrate that pH-dependent GABA transport is a rheogenic process even in the absence of extracellular Na+, consistent with H+/GABA symport. 4. A range of GABA analogues were tested for their abilities to: (a) inhibit pH dependent [3H]GABA uptake across the apical membrane; (b) stimulate H+ flow across the apical surface of BCECF-loaded Caco-2 cell monolayers; (c) increase inward ISC across voltage-clamped Caco-2 cell monolayers. 5. Nipecotic acid, isonipecotic acid, D,L-beta-aminobutyric acid, and 3-amino-1-propanesulphonic acid each caused a marked acidification of intracellular pH and an increase in ISC when superfused at the apical surface of Caco-2 cell monolayers. In contrast L-alpha-amino-n-butyric acid failed to induce proton flow or ISC. The ability of these compounds to induce proton or current flow across the apical surface of this intestinal epithelium was closely related to the relative inhibitory effects on [3H]GABA uptake. 6. These observations demonstrate H+/GABA symport and suggest that this transport mechanism may be accessible as a route for oral absorption of therapeutically-useful GABA analogues. PMID- 10711342 TI - Regulation of beta 1- and beta 3-adrenergic agonist-stimulated lipolytic response in hyperthyroid and hypothyroid rat white adipocytes. AB - 1. This study examined the effects of thyroid status on the lipolytic responses of rat white adipocytes to beta-adrenoceptor (beta-AR) stimulation. The beta 1- and beta 3-AR mRNAs and proteins were measured by Northern and saturation analyses, respectively. Glycerol production and adenyl cyclase (AC) activity induced by various non-selective and selective beta 1/beta 3-AR agonists and drugs which act distal to the receptor in the signalling cascade were measured in cells from untreated, triiodothyronine (T3)-treated and thyroidectomized rats. 2. The beta 3-AR density was enhanced (72%) by T3-treatment and reduced (50%) by introduction of a hypothyroid state while beta 1-AR number remained unaffected. The beta 1- and beta 3-AR density was correlated with the specific mRNA level in all thyroid status. 3. The lipolytic responses to isoprenaline, noradrenaline (beta 1/beta 3/beta 3-AR agonists) and BRL 37344 (beta 3-AR agonist) were potentiated by 48, 58 and 48%, respectively in hyperthyroidism and reduced by about 80% in hypothyroidism. 4. T3-treatment increased the maximal lipolytic response to the partial beta 3-AR (CGP 12177) and beta 1-AR (xamoterol) agonists by 234 and 260%, respectively, increasing their efficacy (intrinsic activity: 0.95 versus 0.43 and 1.02 versus 0.42). The maximal AC response to these agonists was increased by 84 and 58%, respectively, without changing their efficacy. 5. In the hypothyroid state, the maximal lipolytic and AC responses were decreased with CGP (0.17 +/- 0.03 versus 0.41 +/- 0.08 mumol glycerol/10(6) adipocytes; 0.048 +/ 0.005 versus 0.114 +/- 0.006 pmol cyclic AMP min-1 mg-1) but not changed with xamoterol. 6. The changes in lipolytic responses to postreceptor-acting agents (forskolin, enprofylline and dibutenyl cyclic AMP, (Bu)2cAMP) suggest the modifications on receptor coupling and phosphodiesterase levels in both thyroid states. 7. Thyroid status affects lipolysis by modifying beta 3-AR density and postreceptor events without changes in the beta 1-AR functionality. PMID- 10711344 TI - Differential regulation of mammalian brain-specific proline transporter by calcium and calcium-dependent protein kinases. AB - 1. This study examined the role of [Ca2+]I and Ca(2+)-dependent kinases in the modulation of high-affinity, mammalian brain-specific L-proline transporter (PROT). 2. beta-PMA (phorbol 12-myristate 13-acetate), an activator of protein kinase C (PKC), inhibits PRO uptake, and bisindolymalemide I (BIM), a potent PKC inhibitor, prevents beta-PMA inhibition. Down-regulation of PKC by chronic treatment with beta-PMA enhances PROT function indicating PROT regulation by tonic activity of PKC. 3. Thapsigargin, which increases [Ca2+]I levels by inhibiting Ca(2+)-ATPase, inhibits PROT and exhibits additive inhibition when co treated with beta-PMA. KN-62, a Ca2+/calmodulin-dependent kinase II (CaMK II) inhibitor, but not BIM (a PKC inhibitor) prevents the inhibition by thapsigargin. These data suggest that PKC and CaMK II modulate PROT and that thapsigargin mediates its effect via CaMK II. 4. Thapsigargin raises [Ca2+]I and increases PRO induced current on a second time scale, whereas the inhibitory effect of thapsigargin occurs only after 10 min of treatment. These data suggest that Ca2+ differentially regulate PROT: Ca2+ initially enhances PRO transport but eventually inhibits transport function through CaMK II pathway. 5. Ca(2+)-induced stimulation exemplifies the acute regulation of a neurotransmitter transporter, which may play a critical role in the profile of neurotransmitters during synaptic transmission. PMID- 10711345 TI - Somatostatin-induced control of cytosolic free calcium in pituitary tumour cells. AB - 1. In rat pituitary tumour cells (GC cells), spontaneous oscillations of the intracellular concentration of Ca2+ ([Ca2+]i) induce growth hormone (GH) secretion that is inhibited by octreotide, a somatostatin (SRIF) agonist which binds to SRIF subtype (sst) receptor 2. The effects of its functional activation on the control of [Ca2+]i were investigated using fluorimetric measurements of [Ca2+]i. 2. SRIF decreases the basal [Ca2+]i and the [Ca2+]i rise in response to forskolin (FSK) through the inhibition of L-type voltage-dependent Ca2+ channels. 3. Pretreatment with octreotide or with L-Tyr8++ Cyanamid 154806, a sst2 receptor antagonist, abolishes the SRIF-induced inhibition of [Ca2+]i. Octreotide is known to operate through agonist-induced desensitization, while the antagonist operates through receptor blockade. 4. sst1 and sst2 receptor-immunoreactivities (-IRs) are localized to cell membranes. sst2, but not sst1 receptor-IR, internalizes after cell exposure to octreotide. 5. SRIF-induced inhibition of basal [Ca2+]i or FSK-induced Ca2+ entry is blocked by pertussis toxin (PTX). 6. FSK-induced cyclic AMP accumulation is only partially decreased by SRIF or octreotide, indicating that sst2 receptors are coupled to intracellular pathways other than adenylyl cyclase (AC) inhibition. 7. In the presence of H-89, an inhibitor of cyclic AMP dependent protein kinase (PKA), SRIF-induced inhibition of basal [Ca2+]i is still present, although reduced in amplitude. 8. SRIF inhibits [Ca2+]i by activating sst2 receptors. Inhibition of AC activity is only partly responsible for this effect, and other transduction pathways may be involved. PMID- 10711346 TI - The novel immunosuppressant SDZ-RAD protects rat brain slices from cyclosporine induced reduction of high-energy phosphates. AB - 1. SDZ-RAD, 40-O-(2-hydroxyethyl)-rapamycin, is a novel macrolide immunosuppressant. Because of its synergistic interaction, SDZ-RAD is under clinical investigation as immunosuppressant in combination with cyclosporine after organ transplantation. Neurotoxicity is a critical side-effect of cyclosporine. 2. We studied the effect of SDZ-RAD and its combination with cyclosporine on high-energy phosphates, phosphocreatine (PCr) and nucleoside triphosphates (NTP), in brain slices using 31P-magnetic resonance spectroscopy (MRS). 3. Cyclosporine significantly reduced high-energy phosphates after 2 h in a dose-dependent manner (100 micrograms l-1: 93 +/- 3% of control (NTP), 91 +/- 3% (PCr); 500 micrograms l-1: 84 +/- 2% (NTP), 73 +/- 2 (PCr); 5000 micrograms l 1: 68 +/- 3% (NTP), 55 +/- 5% (PCr); n = 6; P < 0.02). 4. In contrast, after perfusion for 2 h, SDZ-RAD (500 micrograms l-1 and 5000 micrograms l-1) significantly increased high-energy phosphate concentrations in the brain slices (P < 0.02). Even at the lowest concentration, SDZ-RAD protected brain energy metabolism against cyclosporine toxicity: 100 micrograms l-1 SDZ-RAD + 5000 micrograms l-1 cyclosporine: 86 +/- 3% (NTP), 83 +/- 7% (PCr), n = 3, P < 0.03 compared to cyclosporine alone. 5. As evaluated using an algorithm based on Loewe isobolograms, the effects of SDZ-RAD/cyclosporine combinations on brain energy reduction were antagonistic. Both drugs were found in mitochondria using h.p.l.c MS analysis. 6. We conclude that cyclosporine inhibits mitochondrial high-energy phosphate metabolism, which can be antagonized by SDZ-RAD. PMID- 10711347 TI - Prejunctional muscarinic inhibitory control of acetylcholine release in the human isolated detrusor: involvement of the M4 receptor subtype. AB - 1. Experiments were carried out in human detrusor strips to characterize muscarinic receptor subtypes involved in the prejunctional regulation of acetylcholine (ACh) release from cholinergic nerve terminals, and in the postjunctional smooth muscle contractile response. 2. In detrusor strips preincubated with [3H]-choline, electrical field stimulation (600 pulses) delivered in six trains at 10 Hz produced a tritium outflow and a contractile response. In the presence of 10 microM paraoxon (to prevent ACh degradation) the tritium outflow was characterized by HPLC analysis as [3H]-ACh (76%) and [3H] choline (24%). 3. Electrically-evoked [3H]-ACh release was abolished by tetrodotoxin (TTX: 300 nM) and unaffected by hexamethonium (10 microM), indicating a postganglionic event. It was reduced by physostigmine (100 nM) and the muscarinic receptor agonist, muscarone (10 nM-1 microM), and enhanced by atropine (0.1-100 nM). These findings indicate the presence of a muscarinic negative feedback mechanism controlling ACh release. 4. The effects of various subtype-preferring muscarinic receptor antagonists were evaluated on [3H]-ACh release and muscle contraction. The rank potency (-log EC50) orders at pre- and postjunctional level were: atropine > or = 4-diphenyl-acetoxy-N-piperidine (4 DAMP) > mamba toxin 3 (MT-3) > tripitramine > para-fluorohexahydrosiladiphenidol (pF-HHSiD) > or = methoctramine > or = pirenzepine > tripinamide, and atropine > or = 4-DAMP > pF-HHSiD >> pirenzepine = tripitramine > tripinamide > methoctramine >> MT-3, respectively. 5. The comparison of pre- and post junctional potencies and the relationship analysis with the affinity constants at human cloned muscarinic receptor subtypes indicates that the muscarinic autoreceptor inhibiting ACh release in human detrusor is an M4 receptor, while the receptor involved in muscular contraction belongs to the M3 subtype. PMID- 10711349 TI - Effects of some isoprostanes on the human umbilical artery in vitro. AB - 1. Cumulative concentration-effect curves for the selective prostanoid TP receptor agonist U46619 and six isoprostanes were constructed in the human isolated umbilical artery. 2. All compounds except 8-iso-PGF3 alpha produced concentration-dependent contractions. The contractile response to the isoprostanes increased with each cumulative addition up to a point, after which subsequent addition reduced the contraction below the previous level. This 'downturn' in the concentration-effect curve did not occur with U46619. 3. The potencies of the compounds tested were as follows (pEC50 +/- s.e.mean): U46619, 6.7 +/- 0.2; 8-iso-PGE2, 6.5 +/- 0.1; 8-iso-PGF2 alpha, 5.8 +/- 0.2; 8-iso-PGE1, 5.4 +/- 0.1; 8-iso-PGF1 alpha, 5.0 +/- 0.1; 8-iso-PGF2 beta > 4.8; 8-iso-PGF3 alpha >> 4.8 (n = 4-17). Neither 8-iso-PGF2 beta nor 8-iso-PGF3 alpha at 44 microM had a significant effect on cumulative concentration-effect curves to U46619. 4. The selective TP receptor antagonist GR32191 (0.1 microM) caused rightward shifts in the concentration-effect curves to all the active compounds. pA2 values for GR32191 against U46619, 8-iso-PGE2, 8-iso-PGF2 alpha, 8-iso-PGE1 were 7.6 +/- 0.2, 9 +/- 1, 8.2 +/- 0.3 and 7.7 +/- 0.3, respectively (n = 4). 5. Neither N omega-nitro-L-arginine methyl ester (100 microM) nor the selective DP receptor antagonist BW A868C (50 nM) affected the complex concentration-effect curve to 8-iso-PGE2 (n = 3). 6. Stable contractions to U46619 (1-3 microM) were unaffected by anandamide at concentrations up to 60 microM. PMID- 10711348 TI - No contractile effect for 5-HT1D and 5-HT1F receptor agonists in human and bovine cerebral arteries: similarity with human coronary artery. AB - 1. Using subtype-selective 5-HT1 receptor agonists and/or the 5-HT1 receptor antagonist GR127935, we characterized in vitro the 5-HT receptor that mediates the contraction of human and bovine cerebral arteries. Further, we investigated which sumatriptan-sensitive receptors are present in human coronary artery by reverse-transcriptase polymerase chain reaction (RT-PCR). 2. Agonists with affinity at the 5-HT1B receptor, such as sumatriptan, alniditan and/or IS-159, elicited dose-dependent contraction in both human and bovine cerebral arteries. They behaved as full agonists at the sumatriptan-sensitive 5-HT1 receptors in both species. In contrast, PNU-109291 and LY344864, selective agonists at 5-HT1D and 5-HT1F receptors, respectively, were devoid of any significant vasocontractile activity in cerebral arteries, or did not affect the sumatriptan induced vasocontraction. The rank order of agonist potency was similar in both species and could be summarized as 5-HT = alniditan > sumatriptan = IS-159 >>> PNU-109291 = LY344864. 3. In bovine cerebral arteries, the 5-HT1 receptor antagonist GR127935 dose-dependently inhibited the vasoconstrictions elicited by both 5-HT and sumatriptan, with respective pA2 values of 8.0 and 8.6. 4. RT-PCR studies in human coronary arteries showed a strong signal for the 5-HT1B receptor while message for the 5-HT1F receptor was weak and less frequently detected. Expression of 5-HT1D receptor mRNA was not detected in any sample. 5. The present results demonstrate that the triptan-induced contraction in brain vessels is mediated exclusively by the 5-HT1B receptor, which is also present in a majority of human coronary arteries. These results suggest that selective 5-HT1D and 5 HT1F receptor agonists might represent new antimigraine drugs devoid of cerebro- and cardiovascular effects. PMID- 10711350 TI - Participation of mitogen-activated protein kinase in thapsigargin- and TPA induced histamine production in murine macrophage RAW 264.7 cells. AB - 1. Stimulation of the murine macrophage cell line RAW 264.7 with thapsigargin, an endomembrane Ca(2+)-ATPase inhibitor, induced histamine production in a time- and concentration-dependent manner. 2. The protein kinase C activator, 12-O tetradecanoylphorbol 13-acetate (TPA), also enhanced histamine production. 3. alpha-Fluoromethylhistidine, a suicide substrate of L-histidine decarboxylase (HDC), suppressed the thapsigargin (30 nM)- and TPA (30 nM)-induced histamine production. 4. Both thapsigargin (30 nM) and TPA (30 nM) induced phosphorylation of p44/p42 MAP kinase and p38 MAP kinase. 5. PD98059, a specific inhibitor of MEK 1 which phosphorylates p44/p42 MAP kinase, strongly suppressed both the thapsigargin (30 nM)- and TPA (30 nM)-induced histamine production, whereas SB203580, a specific inhibitor of p38 MAP kinase, inhibited them only partially. 6. The other MEK-1 inhibitor, U-0126, also inhibited both the thapsigargin- and TPA-induced histamine production in a concentration-dependent manner. 7. Thapsigargin (30 nM) and TPA (30 nM) increased the levels of HDC mRNA at 4 h, but PD98059 suppressed both the thapsigargin- and TPA-induced increases in the HDC mRNA level. 8. These findings indicate that thapsigargin and TPA induce histamine production in RAW 264.7 cells by increasing the level of HDC mRNA, and that both the thapsigargin- and TPA-induced histamine production are regulated largely by p44/p42 MAP kinase and partially by p38 MAP kinase. PMID- 10711351 TI - Involvement of kinins in hyperresponsiveness induced by platelet activating factor in the human nasal airway. AB - 1. The aim of this study was to investigate the role of kinins in the development of nasal hyperresponsiveness induced by platelet activating factor (PAF) in normal human subjects. 2. Intranasal administration of PAF, 60 micrograms, induced an increased responsiveness to histamine, 200 micrograms per nostril, 6 h later. This effect was abolished by pretreatment with the bradykinin B2 receptor antagonists icatibant and [1-adamantaneacetyl-D-Arg0,Hyp3,beta-(2-thienyl)-Al a5,8,D-Phe7]-bradykinin ([Ad]-BK), both at 200 micrograms, every 2 h following PAF administration. 3. In a separate experiment, utilizing the same protocol, nasal lavage was used to measure the release of mediators into the nasal cavity following treatment with PAF. PAF increased the levels of eosinophil cationic protein (ECP) and kinin detected in the lavage samples, compared with a saline control. The levels of these mediators were reduced by pretreatment with either icatibant or [Ad]-BK. 4. Administration of lyso-PAF, 60 micrograms intranasally, did not cause a rise in kinin or ECP levels in nasal lavage fluid. 5. Exogenous bradykinin, 500 micrograms, or a saline control, applied topically to the nasal mucosa every 30 min for 2 h, failed to cause hyperresponsiveness to histamine. 6. We conclude that bradykinin itself does not cause hyperresponsiveness, but is involved in the hyperresponsiveness induced by PAF in the human nasal airway. PMID- 10711352 TI - The effects of heparin and related molecules upon the adhesion of human polymorphonuclear leucocytes to vascular endothelium in vitro. AB - 1. The effects of an unfractionated heparin preparation (Multiparin), a low molecular weight heparin preparation (Fragmin) and a selectively O-desulphated derivative of heparin lacking anticoagulant activity, have been investigated for their effects on the adhesion of human polymorphonuclear leucocytes (PMNs) to cultured human umbilical vein endothelial cells (HUVECs) in vitro. The effect of poly-L-glutamic acid, a large, polyanionic molecule was also studied. 2. Unfractionated heparin (50-1000 U ml-1), the O-desulphated derivative (0.3-6 mg ml-1) and the low molecular weight heparin (50 U-1000 U ml-1) all inhibited significantly the adhesion of 51Cr labelled PMNs to HUVECs stimulated with interleukin-1 beta (IL-1 beta; 10 U ml-1), bacterial lipopolysaccharide (LPS; 2.5 micrograms ml-1) or tumour necrosis factor-alpha (TNF-alpha; 125 U ml-1) for 6 h, whereas poly-L-glutamic acid had no effect. In addition, the three heparin preparations in the same concentration range inhibited significantly the adhesion of f-met-leu-phe-stimulated PMNs to resting HUVECs. 3. The effects of unfractionated heparin upon the expression of adhesion molecules intercellular adhesion molecule-1 (ICAM-1) and E-selection were also investigated, as were the effects of unfractionated heparin upon adhesion of human PMNs to previously stimulated HUVECs. Heparin had little effect upon levels of expression of these adhesion molecules on stimulated HUVECs. However, a profound effect upon PMN adhesion to previously stimulated HUVECs was demonstrated using the same preparation, suggesting that inhibition of adhesion molecule expression is not a major component of the described inhibitory effects of heparin. 4. Pre-incubation of PMNs with heparin followed by washing inhibited their adhesion to HUVECs, under different conditions of cellular activation, implying that heparin can bind to these cells and exert its anti-adhesive effects even when not directly present in the system. 5. These observations would suggest that both heparin and a low molecular weight heparin are capable of inhibiting adhesion of human PMNs to endothelial cells, an effect not dependent solely upon the polyanionic nature of these molecules, nor dependent upon their ability to act as anticoagulants. PMID- 10711353 TI - The group II metabotropic glutamate receptor agonist, DCG-IV, alleviates akinesia following intranigral or intraventricular administration in the reserpine-treated rat. AB - 1. This study examined whether activation of group II metabotropic glutamate (mGlu) receptors in the substantia nigra pars reticulata (SNr) could reverse akinesia in a rodent model of Parkinson's disease (PD). 2. Male Sprague Dawley rats, stereotaxically cannulated above either the SNr or third ventricle, were rendered akinetic by injection of reserpine (5 mg kg-1 s.c.). Eighteen hours later, the rotational behaviour induced by unilateral injection of the group II mGlu receptor agonist, (2S,2'R,3'R)-2-(2',3'-dicarboxycyclopropyl)glycine (DCG IV), was examined. 3. Following intranigral injection, DCG-IV (0.125-0.75 nmol in 0.1 microliter) produced a dose-dependent increase in net contraversive rotations (n = 6-8 animals per dose), reaching a maximum of 395 +/- 51 rotations 60 min-1 after 0.75 nmol. The effects of DCG-IV (0.5 nmol) were inhibited by 63.0 +/- 9.0% following 30 min pre-treatment with the group II mGlu receptor antagonist, (2S) alpha-ethylglutamic acid (EGLU; 100 nmol in 0.2 microliter; n = 6). 4. Following intraventricular injection, DCG-IV (0.125-1.5 nmol in 2 microliters) produced a dose-dependent increase in bilateral locomotor activity (n = 6-7 animals per dose), reaching a maximum of 180 +/- 21 locomotor units 30 min-1 after 0.5 nmol. Pre-treatment with EGLU (200 nmol in 2 microliters) inhibited the effects of DCG IV (0.5 nmol) by 68.2 +/- 12.3% (n = 5). 5. These data show that activation of group II mGlu receptors in the SNr provides relief of akinesia in the reserpinized rat model of PD. The reversal seen following intraventricular administration supports the likely therapeutic benefit of systemically-active group II mGlu receptor agonists in PD. PMID- 10711354 TI - Clotrimazole inhibits the recombinant human cardiac L-type Ca2+ channel alpha 1C subunit. AB - 1. Clotrimazole (CLT) is an antimycotic agent with a potential role in the treatment of cancer. Whole-cell patch clamp recordings and Fura-2 AM fluorescence measurements were used to investigate the inhibition by CLT of recombinant human cardiac L-type Ca2+ channel alpha 1C subunits, stably expressed in human embryonic kidney (HEK 293) cells. 2. CLT (100 nmol l-1 to 25 mumol l-1) reduced Ca2+ channel currents in a concentration-dependent manner. Inhibition was neither use- or voltage-dependent. The effects of CLT were rapid and maximal effects were attained within 3 min. Application of CLT also caused an acceleration of apparent Ca2+ channel current inactivation. 3. Basal current density and the degree of inhibition due to CLT were not significantly altered by pretreating cells with 3 mmol l-1 1-aminobenzotriazole for 1 h, or by dialysing cells for 10 min with 2 mmol l-1 alpha-napthoflavone via the patch pipette, suggesting that the inhibitory action of CLT was not due to inhibition of cytochrome P-450. 4. CLT (10 mumol l-1) did not influence [Ca2+]i, as determined by Fura-2 AM fluorescence measurements. 5. Dialysing cells for 10 min with the non-specific serine/threonine kinase inhibitor H-7 (10 mumol l-1) was without effect on basal current density or on the inhibitory response to 10 mumol l-1 CLT, indicating that CLT is not acting via an indirect effect on these kinases. 6. These data suggest that CLT exerts a direct blocking effect on the alpha 1C subunit at therapeutic concentrations. This effect may explain the abbreviation of the action potential duration by CLT observed in cardiac myocytes. PMID- 10711356 TI - Effects of local delivery of trapidil on neointima formation in a rabbit angioplasty model. AB - 1. Smooth muscle cell (SMC) proliferation can result in luminal reduction of a vessel following balloon angioplasty. This study was designed (i) to determine if local administration of trapidil (triazolopyrimidine) into a vessel wall reduces neointima formation, and (ii) to explore the mechanism involved in the subsequent reduction in cell proliferation. 2. Following balloon angioplasty in 40 anaesthetized New Zealand White rabbits, trapidil (50-200 mg) or its vehicle (saline) was injected into the dilated vessel wall of the right femoral artery. Experimental groups and time of investigation: (I) vehicle (2 weeks, n = 3), (II) trapidil-100 mg (2 weeks, n = 3), (III) vehicle (3 weeks, n = 8), (IV) trapidil 50 mg (3 weeks, n = 5); (V) trapidil-100 mg (3 weeks, n = 9) or (V) trapidil-200 mg (3 weeks, n = 7). 3. After 2 weeks, there was a significant reduction of intimal hyperplasia (expressed as intima to media area ratio) in the trapidil group compared with vehicle (0.44 +/- 0.04 vs 0.93 +/- 0.04, *P < 0.05) and also a significant reduction in cell proliferation (% ratio of BrdU-positive cells to total cell number: vehicle 14 +/- 2% vs trapidil 6 +/- 1%, *P < 0.05). 4. After 3 weeks, there was a dose-dependent reduction of intimal hyperplasia in the trapidil groups compared with vehicle (trapidil 50 mg 1.14 +/- 0.04; trapidil 100 mg 0.91 +/- 0.09*; trapidil 200 mg 0.77 +/- 0.09* vs vehicle 1.67 +/- 0.23, *P < 0.05). 5. Thus, the local administration of trapidil to the rabbit femoral artery reduces the neointima formation, which occurs 2 or 3 weeks after balloon angioplasty via a mechanism, which is dependent on inhibition of cell proliferation. PMID- 10711355 TI - Mechanisms of 17 beta-oestradiol induced vasodilatation in isolated pressurized rat small arteries. AB - 1. The influence of 17 beta-oestradiol on pressurized isolated rat mesenteric and coronary small arteries was investigated. 2. 17 beta-oestradiol caused rapid (t1.0 < 5 mins) concentration-dependent relaxations of pre-contracted pressurized (50 mmHg) isolated rat mesenteric and coronary arteries. Similar responses were observed in both vessel types. Significant relaxations were only observed at concentrations exceeding 3 microM. 3. The vasodilatory responses in both types of artery were unaffected by 10 microM L-nitro arginine (L-NNA) alone or in the presence of 10 microM indomethacin, inhibitors of nitric oxide and prostaglandin synthesis respectively. They were also unaffected by the pre-contracting agent used i.e. high K+ or U46619 (a thromboxane analogue). 4. Neither the oestrogen receptor antagonist ICI 182,780 (10 microM) nor the protein synthesis inhibitor cycloheximide (100 microM) had any effect on the responses of mesenteric arteries to 17 beta-oestradiol. 5. 17 alpha-oestradiol had only a minor effect on mesenteric arterial diameter over a concentration range similar to the effective vasodilatory range for 17 beta-oestradiol. 6. Membrane impermeant 17 beta oestradiol conjugated to bovine serum albumin (beta-oestradiol-17-hemisuccinate BSA) (E-H-BSA) resulted in a vasodilatation of pressurized arteries. 7. Wortmannin, an inhibitor of myosin light chain kinase, near maximally relaxed pressurized mesenteric arteries although the time course for the response was significantly slower than that for 17 beta-oestradiol. 8. These results taken together suggest that the acute effects of 17 beta-oestradiol on isolated pressurized arterial tone may be due to effects directly on the vascular smooth muscle via non-genomic mechanisms that involve a stereospecific interaction at the plasma membrane. PMID- 10711357 TI - Ontogenic aspects of D1 receptor coupling to G proteins and regulation of rat jejunal Na+, K+ ATPase activity and electrolyte transport. AB - 1. The present study examined the effect of dopamine on rat jejunal electrolyte transport (rheogenic transport and Na+, K(+)-ATPase activity) in adult (60-day old) and young (20-day old) animals. 2. In young rats, dopamine, in the presence of phentolamine, produced an increase in jejunal Isc, this being completely abolished by SKF 83566, and not changed by S-sulpiride. SKF 38393, but not quinerolane, also increased Isc; this effect was abolished by SKF 83566 and ouabain, but not by furosemide. In adult rats, dopamine in the presence of phentolamine (0.2 microM) decreased Isc. 3. Na+, K(+)-ATPase activity in isolated jejunal epithelial cells from adult rats was 2.4 fold that in young rats. In the presence of phentolamine, both dopamine and SKF 38393, but not quinerolane, significantly decreased jejunal Na+, K(+)-ATPase activity in young animals but not in adult animals. 4. Binding [3H]-Sch 23390 to membranes of jejunal mucosa revealed the presence of a single class of receptors in both young and adult rats, with similar KD and Bmax values. 5. GTP gamma S and cholera toxin inhibited jejunal Na+, K(+)-ATPase activity in young, but not in adult rats. Co-incubation of pertussis toxin with dopamine was found to potentiate the inhibitory effects of dopamine upon the enzyme in both young and adult rats. 6. Regulation of Na+, K(+)-ATPase activity by cholera toxin-sensitive G proteins is absent in adult animals, and such difference may explain the failure of dopamine to inhibit intestinal Na+, K(+)-ATPase activity in adult rats. PMID- 10711358 TI - On the mechanism of d-amphetamine-induced changes in glutamate, ascorbic acid and uric acid release in the striatum of freely moving rats. AB - 1. The effects of systemic, intrastriatal or intranigral administration of d amphetamine on glutamate, aspartate, ascorbic acid (AA), uric acid, dopamine (DA), dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA) and 5 hydroxyindoleacetic acid (5-HIAA) concentrations in dialysates from the striatum of freely-moving rats were evaluated using microdialysis. 2. d-Amphetamine (2 mg kg-1) given subcutaneously (s.c.) increased DA, AA and uric acid and decreased DOPAC + HVA, glutamate and aspartate dialysate concentrations over a 3 h period after d-amphetamine. 5-HIAA concentrations were unaffected. Individual changes in glutamate and AA dialysate concentrations were negatively correlated. 3. d Amphetamine (0.2 mM), given intrastriatally, increased DA and decreased DOPAC + HVA and aspartate dialysate concentrations, but failed to change those of glutamate, AA uric acid or 5-HIAA, over a 2 h period after d-amphetamine. Haloperidol (0.1 mM), given intrastriatally, increased aspartate concentrations without affecting those of glutamate or AA. 4. d-Amphetamine (0.2 mM), given intranigrally, increased AA and uric acid dialysate concentrations and decreased those of glutamate, aspartate and DA; DOPAC + HVA and 5-HIAA concentrations were unaffected. 5. These results suggest that d-amphetamine-induced increases in AA and uric acid and decreases in glutamate concentrations are triggered at nigral sites. The changes in aspartate levels may be evoked by at least two mechanisms: striatal (mediated by inhibitory dopaminergic receptors) and nigral (activation of amino acid carrier-mediated uptake). PMID- 10711359 TI - Characterization of prostanoid receptors mediating inhibition of histamine release from anti-IgE-activated rat peritoneal mast cells. AB - 1. Prostanoid receptors mediating inhibition of anti-IgE induced histamine release from rat peritoneal mast cells have been characterized pharmacologically. PGD2 and the specific DP receptor agonists BW 245C and ZK 118182 were the most potent inhibitors with half-maximal concentrations of 0.26, 0.06 and 0.02 microM respectively. The maximum inhibition attainable was 60-65% with 10(-5) M BW 245C and ZK 118182. 2. Among several EP receptor agonists investigated, only PGE2 and the EP2/EP3 receptor agonist misoprostol induced significant inhibition (46.8 +/- 4.7% at 10(-4) M and 18.7 +/- 6.8% at 10(-5) M respectively). The IP receptor agonists cicaprost and iloprost were both less potent than the DP agonists in inhibiting histamine release (45.2 +/- 3.3% and 35.1 +/- 2.5% inhibition respectively at 10(-5) M), whereas PGF2 alpha and the TP receptor agonist U-46619 were only marginally effective. 3. The EP4/TP receptor antagonist AH 23848 failed to affect the inhibitory actions of PGD2 or PGE2 even at 10(-5) M, whereas the DP/EP1/EP2 receptor antagonist AH 6809 slightly enhanced the effect of PGD2 at 10(-6) M. 4. At concentrations of 3 x 10(-6) to 10(-5) M, the putative DP receptor antagonist ZK 138357 dose-dependently suppressed the inhibitory activities of the DP agonists, PGE2 and cicaprost. The antagonism of ZK 138357 against the DP receptor agonists appeared to be competitive with pA2 values of around six. 5. In conclusion, these data support our earlier proposal that an inhibitory DP receptor is the predominant prostanoid receptor in rat peritoneal mast cell. The properties of this receptor in relation to putative DP receptor subtypes reported in the literature are discussed. PMID- 10711360 TI - The gastric H,K-ATPase blocker lansoprazole is an inhibitor of chloride channels. AB - 1. It was postulated that swelling dependent chloride channels are involved in the proton secretion of parietal cells. Since omeprazole, lansoprazole and its acid activated sulphenamide form AG2000 are structurally related to phenol derivatives known to block swelling dependent chloride channels, we set out to test, whether these substances--which are known to block the H,K-ATPase--could also lead to an inhibition of swelling-dependent chloride channels. Swelling dependent chloride channels--characterized in many different cell types--show highly conserved biophysical and pharmacological features, therefore we investigated the effect of omeprazole, lansoprazole and its acid activated sulphenamide form AG2000 on swelling-dependent chloride channels elicited in fibroblasts, after the reduction of the extracellular osmolarity. 2. Omeprazole, lansoprazole and its acid activated sulphenamide form AG2000 are able to block swelling-dependent chloride channels (IClswell). 3. Lansoprazole and its protonated metabolite AG2000 act on at least two different sites of the IClswell protein: on an extracellular site which seems to be in a functional proximity to the nucleotide binding site, and on an intracellular site which allows the formation of disulfide-bridges. 4. The inhibition of the proton pump and the simultaneous blocking of chloride channels by omeprazole, lansoprazole and its acid activated sulphenamide form AG2000, as described here could be an effective mode to restrict proton secretion in parietal cells. PMID- 10711361 TI - Evidence against potassium as an endothelium-derived hyperpolarizing factor in rat mesenteric small arteries. AB - 1. Endothelium-derived hyperpolarizing factor (EDHF) has recently been identified as potassium released from endothelial cells into the myo-endothelial space. The present study was designed to test this hypothesis. 2. In rat small mesenteric arteries, mounted in a wire myograph, relaxation to acetylcholine or potassium was not significantly changed following incubation with oxadiazolo-quinoxalin-1 one (ODQ, 4 microM) and indomethacin (10 microM, n = 9). 3. Maximal relaxations to acetylcholine occurred in all arteries, were maintained and were significantly greater (P < 0.01, n = 9) than the transient relaxations to potassium, which only occurred in 30-40% of vessels. 4. Removal of the vascular endothelium abolished relaxant responses both to potassium and acetylcholine (P < 0.005, n = 9). 5. Compared with responses in 5.5 mM potassium PSS, relaxation responses to added potassium in arteries maintained in 1.5 mM potassium PSS were more marked and were not dependent on the presence of an intact endothelium (n = 8). 6. Incubation with BaCl2 (50 microM) significantly inhibited the maximal relaxant response to potassium in the presence of an intact endothelium in 5.5 mM potassium PSS (P < 0.05, n = 4), but had no effect on relaxation of de endothelialized preparations in 1.5 mM potassium PSS (n = 5). 7. Treatment with ouabain (0.1 mM) abolished the relaxant response to potassium in 1.5 mM potassium PSS (P < 0.001, n = 9), but only partly inhibited the maximal relaxant response to acetylcholine in 5.5 mM potassium PSS (P < 0.01, n = 5). 8. These data show that at physiological concentrations of potassium an intact endothelium is necessary for potassium-induced relaxation in rat mesenteric arteries. Furthermore, the response to potassium is clearly different to that from acetylcholine, indicating that potassium does not mimic EDHF released by acetylcholine in these arteries. PMID- 10711364 TI - The self-compressing tibial intramedullary nail. AB - Laboratory evaluation of a self-compressing tibial nail demonstrated that significant, initial compression of a simulated fracture or nonunion can be obtained and controlled. However, when this nail was tested in cyclic loading, loss of its initial static compression occurred. PMID- 10711362 TI - Extracellular nucleotides activate the p38-stress-activated protein kinase cascade in glomerular mesangial cells. AB - 1. Extracellular ATP and UTP have been reported to activate a nucleotide receptor (P2Y2-receptor) that mediates arachidonic acid release with subsequent prostaglandin formation, a reaction critically depending on the activity of a cytosolic phospholipase A2. In addition, extracellular nucleotides trigger activation of the classical mitogen-activated protein kinase (MAPK) cascade and cell proliferation as well as of the stress-activated protein kinase (SAPK) cascade. 2. In this study, we report that ATP and UTP are also able to activate the p38-MAPK pathway as measured by phosphorylation of the p38-MAPK and its upstream activators MKK3/6, as well as phosphorylation of the transcription factor ATF2 in a immunocomplex-kinase assay. 3. Time courses reveal that ATP and UTP induce a rapid and transient activation of the p38-MAPK activity with a maximal activation after 5 min of stimulation which declined to control levels over the next 20 min. 4. A series of ATP and UPT analogues were tested for their ability to stimulate p38-MAPK activity. UTP and ATP were very effective analogues to activate p38-MAPK, whereas ADP and gamma-thio-ATP had only moderate activating effects. 2-Methyl-thio-ATP, beta gamma-imido-ATP, AMP, adenosine and UDP had no significant effects of p38-MAPK activity. In addition, the extracellular nucleotide-mediated effect on p38-MAPK was almost completely blocked by 1 mM of suramin, a putative P2-purinoceptor antagonist. 5. In summary, these results demonstrate for the first time that extracellular nucleotides are able to activate the MKK3/6- p38-MAPK cascade most likely via the P2Y2-receptor. Moreover, this finding implies that all three MAPK subtypes are signalling candidates for extracellular nucleotide-stimulated cell responses. PMID- 10711363 TI - Characterization of MPP+ secretion across human intestinal Caco-2 cell monolayers: role of P-glycoprotein and a novel Na(+)-dependent organic cation transport mechanism. AB - 1. In the kidney, a number of transport proteins involved in the secretion of permanently charged organic cations have recently been cloned. To evaluate the possible similarities between intestine and kidney in the handling of organic cations we investigated the transport of 1-methyl-4-phenylpyridinium (MPP+) across monolayers of intestinal Caco-2 cells. MPP+ is a prototypic substrate of the cloned organic cation transporters hOCT1 and hOCT2. 2. In Caco-2 cell monolayers, the basolateral to apical flux of MPP+ was significantly greater than the apical to basolateral flux, consistent with net secretion of MPP+. 3. Net secretion of MPP+ was abolished by addition of either 10 microM cyclosporin A or 100 microM verapamil to the apical membrane. In contrast, secretion of MPP+ was unaffected by addition of either TEA (2 mM) or decynium-22 (2 microM) to either apical or basolateral membranes. These results suggest that MPP+ secretion is mediated primarily by P-glycoprotein located at the apical membrane. We found no evidence of a role for hOCT1 or hOCT2 in the secretion of MPP+. 4. In addition to net secretion of MPP+, we found evidence of a Na(+)-dependent MPP+ uptake mechanism at the apical membrane of Caco-2 cells. 5. Na(+)-dependent MPP+ uptake was sensitive to inhibition by the organic cations; decynium-22 (2 microM), TEA (2 mM) and cimetidine (5 mM) but not by carnitine, guanidine or proline. 6. These results suggest that net secretion of MPP+ across the apical membrane of Caco-2 cells is a function of the relative contributions of MPP+ secretion mediated by P glycoprotein and MPP+ absorption mediated by a novel Na(+)-dependent transport mechanism. PMID- 10711365 TI - The results of intraoperative autotransfusion in orthopaedic surgery. AB - Perioperative hemorrhage associated with major orthopaedic surgery can become life threatening. Homologous bank blood transfusion can replace the volume of blood lost but it has serious disadvantages such as the transmission of viral agents, it has an insufficient platelet count, and transfusion reactions are possible. Hypotensive anesthesia, predeposited autologous blood transfusion and intraoperative autotransfusion are used to reduce these disadvantages. This study evaluates the results of 700 patients who underwent major orthopaedic intervention in our clinic between June 1991 and April 1998. Ninety-nine patients had hip surgery while 601 patients had spinal surgery. The autotransfusion unit saved an average of 858.9 +/- 136.8 cc of blood and an average of 1.9 +/- 1.2 units of saved blood was transfused. None of these patients needed homologous blood transfusion. One hundred patients who had spinal surgery during the same period were used as a control group. The control group required an average of 3.2 +/- 2.1 units of bank blood. Preoperative and postoperative hematocrit values revealed a statistically significant difference between the autotransfusion group and the homologous transfusion group (p < 0.05). The results of this study suggest that intraoperative autotransfusion prevents the decrease in hematocrit values while reducing the need for bank blood transfusion and hence avoiding the risk of transmission of viral infections. PMID- 10711367 TI - Lumbosacral transitional vertebrae and low back pain. AB - The correlation between lumbosacral transitional vertebrae and low back pain was analyzed in a study of plain radiographs from normal subjects and patients with chronic low back pain. Of 184 normal subjects, only 29 (15.8%) had transitional vertebrae, while 97 (35.1%) of 276 patients were noted to have transitional vertebrae. The difference was highly significant (p < 0.01). The incidence of Castellvi type II transitional vertebrae was significantly higher in patients with low back pain than in controls (p < 0.01). This study indicates the etiologic significance of lumbosacral transitional vertebra in low back pain. PMID- 10711366 TI - Increased operative bleeding during orthopaedic surgery in patients with type I Gaucher disease and bone involvement. AB - To aid clinicians in identifying patients with type I Gaucher disease who are at risk of excessive bleeding, we reviewed the coagulation parameters of six affected patients with bone involvement who underwent orthopaedic surgery at two centers, and of 22 patients under treatment at another, seven of whom had total splenectomy. All patients were of Jewish Ashkenazi origin. Among the latter group, prolonged prothrombin time was noted in 81%. Incidence of clotting factor deficiency were as follows: factor XI, 36.3%; V, 31.8%; VIII, 27.2%; IX, 13.6%; and XII, 27.2%. Most of the abnormalities occurred in the non-splenectomized patients. Two of the six orthopaedic surgery patients had excessive intraoperative and postoperative bleeding. One, who underwent spinal decompression had prolonged prothrombin time, and the other, who had total hip replacement, showed a deficiency of factor XI. The second patient's hemoglobin level was maintained with transfusion of fresh frozen plasma during contralateral hip arthroplasty five months later. We suggest that preoperative evaluation of clotting factors and replacement therapy may prevent excessive bleeding in patients with type I Gaucher disease. PMID- 10711368 TI - Intraobserver and interobserver reliability of Kalamchi and Macewen's classification system for evaluation of avascular necrosis of the femoral head in developmental hip dysplasia. AB - This study evaluates the intraobserver and interobserver reliability of the Kalamchi and MacEwen's classification system of avascular necrosis of the femoral head. Radiographs of 48 developmentally dysplastic hips that had an average follow-up of 40.5 months (range: 36 to 52 months) and that had been treated by the same operative technique were interpreted twice by four experienced pediatric orthopaedic surgeons. When the absence or presence of avascular necrosis was taken into consideration the average intraobserver agreement percentage and kappa coefficient were 86% and 0.71, respectively. The average interobserver agreement percentage and kappa coefficient were 83% and 0.66, respectively. When the agreement on the type of avascular necrosis was analyzed, the average intraobserver agreement percentage and kappa coefficient were 85% and 0.74, respectively. The average interobserver agreement percentage and kappa coefficient were 81% and 0.66, respectively. No statistically significant difference was found between the rates of avascular necrosis of four observers. The Kalamchi and MacEwen's classification system was found to be reliable and reproducible. PMID- 10711369 TI - Hip fractures. An epidemiological review. AB - A computerized literature search was performed to identify articles on the epidemiology of hip fractures. Information from a total of 56 papers was gathered, mainly from developed Western countries. Inclusion criteria were the method of randomization, sample size, and relevance for the topic. As the proportion of elderly in the general population has increased, the incidence of hip fractures has risen. Mortality is high: a third of patients do not survive beyond one year after fracturing their hip. There are two main determinants for hip fractures: falls and increased bone fragility. The most important risk factors identified are neuromuscular and visual impairment, pre-existing medical conditions, dementia, low bone density, alcohol intake, smoking, and immobilization. Also, thinner people are at higher risk because of the reduced energy dissipation following a fall. There is a significantly higher risk for institutionalized patients. Primary prevention focuses on eliminating the two main determinants by decreasing falls and their effects, by strengthening the bone either through elimination of risk factors or through medication. Surgical management and early rehabilitation should be favored. Prevention has been shown to be the single most important factor to slow down the increasing incidence of such fractures. More research into prevention is required. PMID- 10711370 TI - The surgical treatment of spinal deformities in the Department of Orthopaedic Surgery, University Medical School of Pecs, Hungary: history and development. AB - Since its foundation in 1966, a great deal of attention has been paid to the treatment of spinal deformities, especially idiopathic scoliosis, in the Department of Orthopaedic Surgery, Medical School of University, Pecs. The authors compared the results of three different surgical methods (modified Hibbs, Harrington, Cotrel-Dubousset) on 171 patients between 1966 and 1992. The average follow-up period was between 2 and 13.2 years. In cases in which the modified Hibbs procedure was used we achieved a 26% correction, in the Harrington cases we achieved a 43.9% correction, and the loss of correction was 100% after the Hibbs operation and 18% after the Harrington operation in the follow-up period. In cases of Cotrel-Dubousset Instrumentation we achieved about 70% correction, but the follow-up period was too short (2 years) to evaluate the loss of correction in these cases. PMID- 10711371 TI - Lectin histochemistry of pathological bones. AB - Our present knowledge of the structure and function of glycoproteins in bone tissue is very limited. The introduction of lectins into histology offered principally a new approach for studying the presence and chemical structure of glycoproteins in tissue sections. In this paper these highly specific carbohydrate binding molecules have been used to characterize glycoproteins in the cellular elements of normal bone and benign bone lesions. We retrospectively examined 35 benign bone lesions (7 fibrous dysplasias, 5 foreign body granulomas, 5 epulis, 8 osteoid osteomas, 10 giant cell tumors) together with 25 normal bone samples. In normal bone samples and all cases of benign bone lesions, two characteristic types of PNA binding were found after neuraminidase digestion in osteoclasts. In osteoclasts which did not adhere to bone surface, diffuse intracytoplasmic PNA binding was seen, and following the adherence to the bone surface, it disappeared, and the resorption zone became stainable. We assume that this PNA binding glycoprotein is formed in the cytoplasm of osteoclasts, then, after the activation of osteoclasts, the glycoprotein gets accumulated at the resorption zone of the cytomembrane where it plays a significant role in the bone resorption. PMID- 10711372 TI - Lumbar spinal synovial cyst presenting with neurological deficit. A case report and review of the literature. AB - A spinal synovial cyst is a rare condition. We reported a case of lumbar spinal synovial cyst presenting neurological deficit. A 78-year-old woman was admitted to our hospital with the low back pain radiating to the left buttock. A myelography with computed tomography and magnetic resonance imaging revealed an extradural cystic lesion at the L5 and S1 level. At the time of surgery, a standard posterior approach was used to expose the posterior elements from L5 to S1. An en bloc laminectomy and total removal of the cyst was performed at the L5 to S1 level. The postoperative recovery was uneventful except for a slight pain in the left leg which persisted for some time. PMID- 10711373 TI - Treatment of posttraumatic midshaft clavicular pseudarthrosis with the Herbert cannulated bone screw and autologous bone grafting. A case report. AB - Pseudarthrosis of the midshaft of the clavicle can be treated successfully using the Herbert cannulated bone screw with no need for a second operation to remove the implant after bone union. PMID- 10711374 TI - Entrapment of the median nerve in a greenstick forearm fracture. A case report and review of the literature. AB - We report a case of low median nerve palsy occurring as a complication of a closed both-bone forearm fracture in a child. Following delayed diagnosis, surgical exploration was performed and it was observed that the median nerve was entrapped in the callus of the radius fracture. PMID- 10711375 TI - Osteonecrosis of the resurfaced patella following bilateral total knee arthroplasty. A case report and review of the literature. AB - A 77-year-old woman with severe varus and flexion deformity of both knee joints, hypertension, and arteriosclerosis underwent bilateral total knee arthroplasty. Six weeks after the operation on the right knee and four months after the operation on the left knee, osteolysis of the patellas was seen on the radiographs. Three years after the operations, the lateral and inferior poles of both patellas were sclerotic and displaced. There was no knee pain and no extension lag throughout the follow-up period. This case suggests that the prognosis of patellar osteonecrosis after total knee arthroplasty is good if the continuity of the retinaculum and the extensor mechanism are preserved, and if instability of the knee does not occur. PMID- 10711376 TI - Exposure of the hip by anterior osteotomy of the greater trochanter. PMID- 10711377 TI - [Morphoscopic perception at variable contrast and luminance levels in 97 subjects]. AB - BACKGROUND: Morphoscopic perception at variable contrast and luminance levels has been assessed in various diseases and in occupational medicine. To our knowledge, this function has not been studied in young adults of similar age. We performed a prospective study to assess morphoscopic perception at variable contrast and luminance levels in a group of young servicemen. METHODS: A total of 97 subjects (194 eyes) with a mean age of 22.4 (standard deviation 1.16) years were assessed with the Gradual monitor. After visual acuity was measured, morphoscopic perception was assessed at a mean light level of 85 cd/m2, at low, medium and high spatial frequencies, with myopic correction. All subjects whose visual acuity was not correctable to 10/10 in both eyes or whose score was statistically too different from the mean underwent a complete ophthalmologic examination. RESULTS: Overall, 87 subjects had a visual acuity correctable to 10/10 or better in both eyes. For 34 of the 87, at least one of the six scores was more than 1 standard deviation below the mean; however, for 16 subjects the result was within normal limits after minor adjustment of the correction, often of a small degree of astigmatism. Of the 18 remaining subjects 7 had microstrabismus, 8 had anatomic lesions and 3 had lesions of unknown origin. Of the 13 subjects with a visual acuity of less than 10/10, 5 had relative amblyopia, 2 had major ametropia, 2 had unilateral and 2 bilateral ophthalmologic lesions, and 2 had reduced acuity of unknown cause. INTERPRETATION: In this young population the most frequent cause of reduced contrast sensitivity (40%) was associated with minor uncorrected or badly corrected refractive problems. The next most frequent cause was problems with binocular vision, with or without amblyopia (11%). In these subjects contrast sensitivity was reduced in the nondominant eye and, in five cases, in the dominant eye also. PMID- 10711378 TI - The genetic aspects of adult-onset glaucoma: a perspective from the Greater Toronto area. AB - BACKGROUND: The myocilin gene is the first glaucoma gene to be associated with primary open-angle glaucoma (POAG). The hereditary subset of POAG and the role of the myocilin gene in our population are not clearly defined. Identification of cases of hereditary glaucoma and a better appreciation of the role of the myocilin gene may allow earlier diagnosis of the disease and optimize management of those at risk for glaucoma. METHODS: Patients were recruited from university glaucoma practices in the Greater Toronto area from 1996 to 1998. Pedigree analysis and DNA banking were performed for each participant. Mutational analysis of the myocilin gene by means of single-strand conformation polymorphism analysis and direct sequencing was completed for 140 probands with POAG of diverse ethnic background. RESULTS: A total of 103 patients (55.7%) had a family history of glaucoma. Disease-causing mutations of the myocilin gene were observed in 7 (5.0%) of the 140 probands, which accounted for 6.5% (5/77) of the familial cases. Most mutations were associated with familial disease, which implies a 50% risk of transmission of a high-risk factor for glaucoma. INTERPRETATION: The hereditary subset of POAG is significant, and heritable glaucoma should always be suspected. In spite of the diversity of the ethnic background of our subjects, the observed prevalence of myocilin gene mutations was comparable to that previously reported, and such mutations do not appear to spare any ethnic group. PMID- 10711379 TI - Outcome of lacrimal surgery in older patients. AB - BACKGROUND: Performance of lacrimal surgery under neuroleptic (local) anesthesia has greatly facilitated the procedure and decreased the associated morbidity. We reviewed the outcome of lacrimal surgery in older patients to determine whether such surgery can be performed safely in the outpatient setting in this group. METHODS: Review of the office and hospital charts and the surgical and anesthetic records of 120 patients (84 women and 36 men) aged 70 to 90 years who underwent lacrimal drainage procedures (dacryocystorhinostomy [DCR], canaliculodacryocystorhinostomy, DCR with insertion of a Jones tube, or a revision endonasal procedure with probing and tube insertion) at a university affiliated hospital in Toronto in 1996. The interval between surgery and data collection ranged from 10 to 22 months. RESULTS: Of the 120 patients 65 were aged 70 to 75 years, 38 were 76 to 80 years, 11 were 81 to 85 years, and 6 were 86 to 90 years. Ninety-six patients had a unilateral procedure, and 24 (22 of whom were aged 70 to 80) had a bilateral procedure. Concomitant conditions, such as hypertension and cardiac disorders, were found in 104 patients (87%). Of the 120 patients 98 (82%) (including all those aged 81 to 90) had local anesthesia, and 22 (18%) had general anesthesia. In one case anesthesia had to be changed from local to general during the procedure because of noncompliance. A total of 112 patients (93%) whose surgery was planned as a day procedure were able to leave the hospital the same day. Three additional patients were admitted to hospital for an overnight stay because of increased bleeding at the time of surgery (one patient) or a history of cardiac problems (two patients). Five patients who had planned overnight stays because of cardiac problems did well during surgery and were discharged the same day, without consequence. None of the patients had to be readmitted at a later date for bleeding or health problems. In 109 patients (91%) the presenting symptom(s) was completely relieved. Overall, 116 patients (97%) had a totally open system with no reflux on syringing. INTERPRETATION: The surgical goals and techniques of lacrimal surgery in older patients were not compromised by performing the surgery in the outpatient setting and under neuroleptic anesthesia in most cases. PMID- 10711380 TI - Comparison of the efficacy of argon green and krypton yellow laser photocoagulation by sectorial treatment of a circumscribed choroidal hemangioma. PMID- 10711381 TI - Cataract extraction by phacoemulsification in a sympathizing eye. PMID- 10711382 TI - Tobramycin-responsive Fusarium oxysporum keratitis. PMID- 10711383 TI - Ocular factitious disorder presenting as endophthalmitis. PMID- 10711384 TI - Intraocular gnathostomiasis: a rare Canadian case. PMID- 10711385 TI - Mechanisms of nephrotoxicity from metal combinations: a review. AB - The common environmental pollutants arsenic, lead, and cadmium are each known to induce chronic renal disease and the molecular mechanisms of such toxic events are being clarified. Nephrotoxicity of these metals is due to the fact that urinary elimination is a main route of excretion, and the proximal tubules are especially sensitive due to their high reabsorptive activity. Renal pathological effects of these metals vary with the chemical form of the metal, the dose, and whether the exposure is acute or chronic in nature. The few isolated studies of combined metal exposures indicate that these pathological effects may be altered due to unknown interactions of these metals within the kidney. Biological factors within the cell such as metal binding proteins and inclusion bodies may also influence metal-metal interactions. Further research is needed to specify the parameters or criteria by which metal interactions is to be assessed for unique biological response patterns to aid in the risk assessment analysis of environmental and occupational metal exposures. PMID- 10711386 TI - Recent developments in the understanding of benzene toxicity and leukemogenesis. PMID- 10711387 TI - Toxicological evaluations of alternative fluorocarbons. PMID- 10711388 TI - Derivation of U.S. EPA's oral Reference Dose (RfD) for methylmercury. AB - Mercury (Hg) cycles in the environment through a series of complex chemical and physical transformations that occur in air, soils, and water bodies. One component of the environmental mercury cycle is the formation of methylmercury (MHg) primarily by aquatic and marine microorganisms and the accumulation of MHg in foodwebs, particularly in piscivorous species. Human consumption of piscivorous fish and other piscivorus animals is the most common pathway of exposure to MHg. For non-carcinogenic toxic endpoints, the U.S. EPA typically develops a Reference Dose (RfD). This is generally interpreted to be a concentration of a chemical which can be consumed on a daily basis over a lifetime without expectation of adverse effect. There is substantial evidence in both animal and humans that MHg is a neurotoxicant in the adult and the child as well as a developmental neurotoxicant for the fetus. Epidemics of MHg poisoning in Japan and Iraq have resulted from high-dose exposures to MHg. In these epidemics adults, children, nursing infants and fetuses were affected by MHg. The epidemics demonstrate that neurotoxicity is the health effect of greatest concern and that effects on the developing human nervous system apparently occur at lower exposures than those affecting the adult nervous system. We describe how the data from the Iraqi MHg epidemic were used to derive the current RfD of 0.1 microgram/Kgbw/day (U.S. EPA, 1995;). PMID- 10711390 TI - Case history review--demilitarization combustion permits. AB - In May 1993, Administrative Browner of the U.S. Environmental Protection Agency (USEPA) announced that an indirect exposure health risk assessment was required for all hazardous waste combustion facilities seeking a Resource Conservation and Recovery Act permit. These types of risk assessments evaluate the health and environmental effects from inhalation of emissions (direct exposure) and from contact with environmental media and consumption of food products impacted by the emissions (indirect exposure). Completion of an indirect exposure risk assessment is often complicated by the various methodologies available for generating results and by the requirements of the regulating community. To minimize this complexity and to maximize consistency between risk assessments, the USEPA developed a number of detailed guidance documents. Site-specific conditions and toxicological data gaps, however, continue to present challenges not addressed by these guidance documents. This paper presents some of the specific challenges encountered by the U.S. Army Center for Health Promotion and Preventive Medicine when performing indirect exposure health risk assessments for several demilitarization combustion facilities. PMID- 10711389 TI - Preliminary assessment of health impacts for the Newport Chemical Agent Disposal Facility. AB - A Preliminary Assessment of Health Impacts (PAHI) study was conducted to look at potential human and environmental health impacts due to the air and water emissions generated from the proposed Newport Chemical Agent Disposal Facility (NECDF) in Newport, Indiana. As an alternative to incineration, the NECDF will use a neutralization-based treatment process followed by supercritical water oxidation to destroy the VX nerve agent stored in ton containers at the Newport Chemical Depot. There is no regulatory guidance on conducting an assessment of health impacts for this type of facility. Therefore, The U.S. Army Center for Health Promotion and Preventive Medicine (USACHPPM) designed a PAHI study based on bench-scale data and best engineering estimates that conservatively evaluate possible health effects from the projected air and water emissions. The air portion of the PAHI focused primarily on estimating carcinogenic risks and noncarcinogenic hazards from direct and indirect exposures to the subsistence farmer, subsistence fisher, adult resident, and child resident. The water portion of the PAHI evaluated potential human and environmental impacts using two different procedures individual compound analysis and whole effluent toxicity analysis. The individual compounds analysis compared constituent concentrations in the water emission to Indiana State Water Quality Standards and U.S. Environmental Protection Agency Ambient Water Quality Criteria to evaluate the potential health impacts to human, terrestrial, and aquatic life. The whole effluent toxicity tests were conducted on mammalian (mouse, rat, and rabbit) and aquatic (water flea, algae, and minnows) species to test for potential additive, synergistic, and antagonistic effects. The results of the air and water portion of the study showed that the operation of NECDF would be safe to exposed human and environmental receptors. A Final Assessment of Health Impacts (FAHI) will also be conducted by USACHPPM, after NECDF is constructed and before full-scale operations, to validate the results of the PAHI. PMID- 10711391 TI - Military deployment toxicology: a program manager's perspective. AB - The Persian Gulf War drew attention to the potential hazards of chemicals that personnel may encounter during military operations and deployments overseas. During the War, the oil well fires of Kuwait highlighted the military threat of industrial chemicals in the area of operations. Following the War, the occurrence of Gulf War Illnesses brought home concerns and suspicions regarding "low level" and "mixed" exposures to chemicals. The public's concern and attention resulted in numerous institutional responses to the real and perceived problems of health risks during military deployments. These institutional responses ranged in scope from a Presidential Review Directive to the initiative known as the Deployment Toxicology Research, Development, Testing and Evaluation (RDT&E) Program. Most institutions, however, seem to agree that additional research is needed to assess the health risks from chemical exposures during military deployments. Establishing and managing an effective RDT&E program in risk assessment for deployed forces is a challenging enterprise. The Deployment Toxicology RDT&E Program was conceived utilizing the military's acquisition framework, an effective methodology with a proven record of fielding of new technologies. Based on a series of structured meetings with military representatives that would utilize new risk assessment tools, a hierarchical set of plans was developed to identify and prioritize end products. The challenge ahead for the Deployment Toxicology RDT&E Program is to execute these plans, provide the necessary oversight, and transition the results into successful product development. PMID- 10711392 TI - Assessing the feasibility of an in vitro cytotoxicity method to detect harmful ubiquitous chemicals (detection of non-warfare hazardous chemicals in the operational theater). AB - The objective of this work was to assess the feasibility of accomplishing aqueous extracts of soil samples and determining if the extracted solution induced adverse effects in the human myelomonocytic cell line, HL60. Dosing of HL60 cells was accomplished over a 24-hour period using 100% of extracted media from standard soil samples containing known contaminants. Assessments of viability, apoptosis, reduced thiols, and mitochondrial membrane integrity were accomplished by argon-ion laser flow cytometric analysis, using chemical labels specific for each end-point. The in vitro cytotoxicity data was compared with the results of Microtox and Mutatox tests as well as earthworm and plant toxicity tests. In vitro cytotoxicity tests' results exhibited good correlation with other tests' results. PMID- 10711393 TI - Application of neurobehavioral toxicology methods to the military deployment toxicology assessment program. AB - The military Tri-Service (Army, Navy & Marines, Air Force) Deployment Toxicology Assessment Program (DTAP) represents a 30-year (1996-2026) planning effort to implement comprehensive systems for the protection of internationally deployed troops against toxicant exposures. A major objective of DTAP is the implementation of a global surveillance system to identify chemicals with the potential to reduce human performance capacity. Implementation requires prior development of complex human risk assessment models, known collectively as the Neurobehavioral Toxicity Evaluation Instrument (NTEI), based on mathematical interpolation of results from tissue-based and in vivo animal studies validated by human performance assessment research. The Neurobehavioral Toxicity Assessment Group (NTAG) at the Naval Health Research Center Detachment-Toxicology (NHRC-TD), Dayton, OH, and associated academic institutions are developing and cross validating cellular-level (NTAS), laboratory small animal (NTAB), nonhuman primate (GASP), and human-based (GASH) toxicity assessment batteries. These batteries will be utilized to develop and evaluate mathematical predictors of human neurobehavioral toxicity, as a function of laboratory performance deficits predicted by quantitative structural analysis relationship (QSAR-like) properties of potential toxicants identified by international surveillance systems. Finally, physiologically-based pharmacokinetic (PBPK) and pharmacodynamic (PBPD) modeling of NTAS, NTAB, GASP, GASH data will support multi-organizational development and validation of the NTEI. The validated NTEI tool will represent a complex database management system, integrating global satellite surveillance input to provide real-time decision-making support for deployed military personnel. PMID- 10711394 TI - Chemical exposure guidelines for deployed military personnel. AB - In light of the absence of guidelines and standards applicable to deployed military personnel, the U.S. Army Center for Health Promotion and Preventive Medicine (USACHPPM) has completed Technical Guide 230A, Short-term Chemical Exposure Guidelines for Deployed Military Personnel. This guide provides estimated concentration levels associated with various types of effects for short term exposures from 1 hour up to 2 weeks. A second TG (TG230B, Long-term Chemical Exposure Guidelines for Deployed Military Personnel) is being developed to address potentially longer deployment related exposures (i.e. greater than 2 weeks up to 1 year). This article focuses on TG230A which describes varying severity levels and health effects associated with short-term chemical exposures in a format consistent with the existing doctrinal military risk management paradigm. It is a consolidated reference tool for trained military medical staff to evaluate different chemical hazards, and will ensure more expedient risk management decisions during deployments. The TG also establishes the standard reference for pre- and post-deployment evaluations and risk management decisions, including determinations of resource requirements and equipment specifications. PMID- 10711396 TI - Using GIS to study the health impact of air emissions. AB - Geographical Information Systems (GIS) is a fast-developing technology with an ever-increasing number of applications. Air dispersion modeling is a well established discipline that can produce results in a spatial context. The marriage of these two applications is optimal because it leverages the predictive capacity of modeling with the data management, analysis, and display capabilities of GIS. In the public health arena, exposure estimation techniques are invaluable. The utilization of air emission data, such as U.S. EPA Toxic Release Inventory (TRI) data, and air dispersion modeling with GIS enable public health professionals to identify and define the potentially exposed population, estimate the health risk burden of that population, and determine correlations between point-based health outcome results with estimated health risk. PMID- 10711395 TI - Rapid separation of nitroaromatic compounds by solvating gas chromatography. AB - Nitroaromatic compounds such as 2,4,6-trinitrotoluene are used in the production of explosive and munitions, and are environmental contaminants as a result of such use. Conventional methods to detect these compounds have relied on liquid and gas chromatography to isolate the individual compounds which may be present at low levels in complex environmental matrices before final detection and identification. A new method, solvating gas chromatography, was used to rapidly separate 8 nitroaromatic compounds, showing improved speed relative to conventional liquid and gas chromatography methods. Solvating gas chromatography allows near real-time detection for these energetic compounds, providing improvements in their detection as environmental contaminants, and as compounds of interest to law enforcement and military organizations. PMID- 10711397 TI - Genetic polymorphisms in ethanol metabolism: issues and goals for physiologically based pharmacokinetic modeling. AB - Chronic exposure to excessive ethanol consumption has adverse effects on virtually all organs and tissues in the body, including but not limited to the liver, pancreas, reproductive organs, central nervous system, and the fetus. Exposure to ethanol can also enhance the toxicity of other chemicals. Not all persons exposed to the same amount of ethanol experience the same degree of adverse effects. For example, only 12% to 13% of alcohol abusers develop cirrhosis. Possible factors which may alter susceptibility include age, sex, nutritional status, health status (i.e., smokers) and race. Some of these factors affect susceptibility because they alter ethanol metabolism, which occurs primarily in the liver by alcohol dehydrogenase (ADH). Genetic polymorphisms for ADH partially account for the observed differences in ethanol elimination rates among various populations but the relative contribution to susceptibility is not completely understood. Incorporation of the kinetic parameters associated with ADH polymorphisms into a physiologically based pharmacokinetic (PBPK) model for ethanol will aid in assessing the relative contribution to susceptibility. The specific information required to develop this model includes Km and Kcat values for each ADH isoform and the amount of each isoform present in the liver. Blood ethanol concentrations (BEC) from various populations with known ADH phenotypes are also necessary to validate the model. The impact of inclusion of these data on PBPK model predictions was examined using available information from adult white and African American males. PMID- 10711398 TI - Comparison of human and rodent lung dosimetry models for particle clearance and retention. AB - Interspecies differences in the kinetics and/or mechanisms of particle retention can influence the amount and location of particle retention in the lungs, which can also influence the tissue response to a given particle burden. Dosimetric models may be used to adjust for differences in the exposure-dose relationships in different species, thus allowing for comparison of lung responses at equivalent doses. Although the rat is one of the most frequently used animal models for assessing the risk of exposures to hazardous substances in humans, few data are available for comparison of human and animal responses to inhaled particles. A biologically-based human dosimetric lung model was developed to describe the fate of respirable particles in the lungs of humans, using data from U.S. coal miners and assumptions about the overloading of alveolar clearance from studies in rats. This model includes alveolar, interstitial, and hilar lymph node compartments. The form of the model that provides the best fit to the lung dust burden data in these coal miners includes a first-order interstitialization process and either no dose-dependent decline in alveolar clearance or much less decline than expected from rodent studies. These findings are consistent with the particle retention patterns observed previously in the lungs of primates. This human lung dosimetry model is useful for investigating the factors that may influence the relationships between the airborne particle exposure, lung dust burden, and fibrotic lung disease. PMID- 10711399 TI - Lung cancer risk associated with exposure to man-made fibers. AB - We show that available experimental data from long-term experiments are consistent with the hypothesis that the oncogenic potential of man-made fibers is determined completely by their biopersistence. We present an analysis of these data within the initiation-promotion-progression paradigm of carcinogenesis. Our method of analysis takes explicit account of the temporal pattern of fiber burden in the rat lung, and suggests that fibers act as initiators in the lung. We estimate a dose-dependent initiation parameter and show how it can be transported to human populations for assessment of the risk of lung cancer following exposure to man-made fibers. PMID- 10711400 TI - Preliminary development of a physiological model for perchlorate in the adult male rat: a framework for further studies. PMID- 10711401 TI - Effects of TCDD on thyroid hormone homeostasis in the rat. AB - A physiological dosimetric model was constructed to describe the effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on circulating thyroid hormones in the rat and to test the hypothesis that these hormonal changes cause chronically elevated serum thyrotropin (thyroid stimulating hormone, TSH), which mediates growth promotion and may lead to thyroid tumors in TCDD-treated rats. The model included diffusion restricted distribution of TCDD among compartments for liver, kidney, white fat, slowly and rapidly perfused tissues, and the thyroxine sensitive tissues brown fat, pituitary, and thyroid. Blood was distributed among major vessels and the capillary beds of the tissues. Metabolism of TCDD was limited to the liver. Secretion of 3,5,3'-triiodothyronine (T3) and thyroxine (3,5,3',5'-tetraiodothyronine, T4) from the thyroid was modeled as stimulated by circulating TSH, whose release from the pituitary was regulated by the hypothalamic peptides thyrotropin releasing hormone (activating) and somatostatin (inhibiting). Release of these peptides was represented as inhibited and activated, respectively, by circulating T4. Binding proteins for T3 and T4 and metabolism of the hormones by deiodination were included in thyroxine-sensitive tissues. Induction of hepatic UDP-glucuronosyltransferase-1*6 (UGT), the enzyme which glucuronidates T4, was modeled as induced by the complex formed between TCDD and the aryl hydrocarbon receptor. The computed extent of deiodination, primacy of the thyroid in generating T3 from T4, dependence of liver and kidney on locally produced T3, and export of T3 formed in the pituitary agreed with experimental observations. The model reproduced the observed decrease in circulating T4 and elevated serum TSH following chronic administration of TCDD. The altered levels were attributed to the increased clearance of T4 by the induced UGT and the consequent modification of feedback control of hormone releases. These results are consistent with the hypothesis of growth stimulation by elevated TSH, but measured values of this hormone in blood of rats vary over a large range, and the change induced by TCDD is often small. Measured UGT levels are less variable and the increase in this protein is much greater, suggesting that this response may be a more reliable biomarker for effects of TCDD on the thyroid. PMID- 10711402 TI - Investigation of in vitro toxicity of jet fuels JP-8 and Jet A. AB - The in vitro cytotoxicity and electrophysiological toxicity of Jet Propulsion-8 (JP-8 jet fuel) on four cell types: H4IIE liver cell line, NIH Swiss 3T3 cell line, neuroblastoma x glioma NG108-15 cells, and embryonic hippocampal neurons were investigated. H4IIE cells exposed to Jet A (a commercial fuel) and JP-8 demonstrated identical toxicity with an IC50 of 12.6 +/- 0.4 micrograms/ml for the two fuels. Comparison of H4IIE and NIH/3T3 toxicity to JP-8 revealed that NIH/3T3 cells were more sensitive to JP-8 than H4IIE cells, with an IC50 8.5 +/- 0.1 micrograms/ml. JP-8 exposure for the hippocampal neurons proved to be highly toxic (IC50 of < 2 micrograms/ml), while in contrast, the NG108-15 cells were much less sensitive. Electrophysiological examination of NG108-15 cells showed that administration of JP-8 at 1 microgram/ml did not alter significantly any of the electrophysiological properties. However, exposure to JP-8 at 10 micrograms/ml during a current stimulus of +46 pA decreased the amplitude of the action potential to 83 +/- 7% (n = 4), the rate of rise, dV/dtMAX to 50 +/- 8% (n = 4), and the spiking rate to 25 +/- 11% (n = 4) of the corresponding control levels. These results demonstrate JP-8 induced cytotoxic varies among cell types. The possible mechanisms underlying these observations are presented. PMID- 10711403 TI - Uncertainty analysis of the estimated ingestion rates used to derive the methylmercury reference dose. AB - The U.S. EPA's Reference Dose (RfD) for methylmercury (MeHg) uses a simple one compartment toxicokinetic model to estimate ingestion doses in mg/kg-day from measured concentrations of mercury in hair. The model includes a number of input variables for which point estimates are made. Uncertainty in the inputs is addressed by the use of a 3-fold uncertainty factor. There are, however, published ranges for each variable which are used to develop distributions for each of the inputs. Monte Carlo output of the model is generated. The 90% confidence interval spans a 3-fold to 5-fold range of ingestion doses for any given concentration of mercury in hair. The hair:blood mercury concentration ratio contributes most to the variance of the output. The results indicate that the uncertainty factor of three is appropriate. PMID- 10711404 TI - A multiple-purpose design approach to the evaluation of risks from mixtures of disinfection by-products. AB - Drinking water disinfection has effectively eliminated much of the morbidity and mortality associated with waterborne infectious diseases in the United States. Various disinfection processes, however, produce certain types and amounts of disinfection by-products (DBPs), including trihalomethanes (THM), haloacetic acids, haloacetonitriles, and bromate, among others. Human health risks from the ubiquitous exposure to complex mixtures of DBPs are of concern because existing epidemiologic and toxicologic studies suggest the existence of systemic or carcinogenic effects. Researchers from several organizations have developed a multiple-purpose design approach to this problem that combines efficient laboratory experimental designs with statistical models to provide data on critical research issues (e.g., estimation of human health risk from low-level DBP exposures, evaluation of additivity assumptions as useful for risk characterization, estimation of health risks from different drinking water treatment options). A series of THM experiments have been designed to study embryonic development, mortality and cancer in Japanese medaka (Oryzias latipes) and liver and kidney endpoints in female CD-1 mice. The studies are to provide dose-response data for specific mixtures of the 4 THMs, for the single chemicals, and for binary combinations. The dose-levels and mixing ratios for these experiments were selected to be useful for development and refinement of three different statistical methods: testing for departures from dose-additivity; development of an interactions-based hazard index; and use of proportional response addition as a risk characterization method. Preliminary results suggest that dose-additivity is a reasonable risk assessment assumption for DBPs. The future of mixtures research will depend on such collaborative efforts that maximize the use of resources and focus on issues of high relevance to the risk assessment of human health. PMID- 10711405 TI - Development and utilization of primary hepatocyte culture systems to evaluate metabolism, DNA binding, and DNA repair of xenobiotics. AB - The use of isolated hepatocytes as an approach to evaluate hepatotoxic and hepatocarcinogenic compounds and investigate mechanisms by which chemicals induce liver lesions is well established. This review discusses techniques developed in the author's laboratory describing (1) isolation and primary culture of rodent hepatocytes detailing methods which are optimal for obtaining large numbers of viable cells, (2) DNA damage induced by physical and chemical agents in rodent hepatocytes measured as unscheduled DNA synthesis, and (3) metabolic activation of model hepatocarcinogens, their binding to DNA, and identification of individual adducts thought to be responsible for induction of DNA repair. PMID- 10711406 TI - Illegal or legitimate use? Precursor compounds to amphetamine and methamphetamine. AB - The interpretation of methamphetamine and amphetamine positive test results in biological samples is a challenge to clinical and forensic toxicology for several reasons. The effects of pH and dilution of urine samples and the knowledge about legitimate and illicit sources have to be taken into account. Besides a potentially legal prescription of amphetamines, many substances metabolize to methamphetamine or amphetamine in the body: amphetaminil, benzphetamine, clobenzorex, deprenyl, dimethylamphetamine, ethylamphetamine, famprofazone, fencamine, fenethylline, fenproporex, furfenorex, mefenorex, mesocarb, and prenylamine. Especially the knowledge of potential origins of methamphetamine and amphetamine turns out to be very important to prevent a misinterpretation of the surrounding circumstances and to prove illegal drug abuse. In this review, potential precursor compounds are described, including their medical use and major clinical effects and their metabolic profiles, as well as some clues which help to identify the sources. PMID- 10711407 TI - History of drug metabolism research in Japan. PMID- 10711408 TI - Cryopreserved primary hepatocytes as a constantly available in vitro model for the evaluation of human and animal drug metabolism and enzyme induction. AB - The use of primary hepatocytes is now well established for both studies of drug metabolism and enzyme induction. Cryopreservation of primary hepatocytes decreases the need for fresh liver tissue. This is especially important for research with human hepatocytes because availability of human liver tissue is limited. In this review, we summarize our research on optimization and validation of cryopreservation techniques. The critical elements for successful cryopreservation of hepatocytes are (1) the freezing protocol, (2) the concentration of the cryoprotectant [10% dimethyl-sulfoxide (DMSO)], (3) slow addition and removal of DMSO, (4) carbogen equilibration during isolation of hepatocytes and before cryopreservation, and (5) removal of unvital hepatocytes by Percoll centrifugation after thawing. Hepatocytes of human, monkey, dog, rat, and mouse isolated and cryopreserved by our standard procedure have a viability > or = 80%. Metabolic capacity of cryopreserved hepatocytes determined by testosterone hydroxylation, 7-ethoxyresorufin-O-de-ethylase (EROD), 7 ethoxycoumarin-O-deethylase (ECOD), glutathione S-transferase, UDP-glucuronosyl transferase, sulfotransferase, and epoxide hydrolase activities is > or = 60% of freshly isolated cells. Cryopreserved hepatocytes in suspension were successfully applied in short-term metabolism studies and as a metabolizing system in mutagenicity investigations. For instance, the complex pattern of benzo[a]pyrene metabolites including phase II metabolites formed by freshly isolated and cryopreserved hepatocytes was almost identical. For the study of enzyme induction, a longer time period and therefore cryopreserved hepatocyte cultures are required. We present a technique with cryopreserved hepatocytes that allows the induction of testosterone metabolism with similar induction factors as for fresh cultures. However, enzyme activities of induced hepatocytes and solvent controls were smaller in the cryopreserved cells. In conclusion, cryopreserved hepatocytes held in suspension can be recommended for short-term metabolism or toxicity studies. Systems with cryopreserved hepatocyte cultures that could be applied for studies of enzyme induction are already in a state allowing practical application, but may be further optimized. PMID- 10711409 TI - Detection of Hantaan and Seoul viruses by reverse transcriptase-polymerase chain reaction (RT-PCR) and restriction fragment length polymorphism (RFLP) in renal syndrome patients with hemorrhagic fever. AB - BACKGROUND: Hemorrhagic fever with renal syndrome (HFRS), also called Korean hemorrhagic fever (KHF), is the most common cause of acute renal failure in the Far East. Two serotypes of hantavirus, Hantaan and Seoul viruses are known pathogens for HFRS in Korea. PURPOSE: To elucidate the diagnostic applicability for the serotype diagnosis in HFRS patients, we used nested reverse transcriptase PCR and restriction fragment length polymorphism (nRT-PCR/RFLP) to screen 2 prototype viruses, 11 virus isolates from HFRS patients, and 69 specimens obtained from 31 HFRS patients. METHODS: The nRT-PCR was performed using primers specific for the G1 segments of the Hantaan (HF3 1140-1163, HB14 1363-1342) and Seoul (SF2 809-832, SB3 1200-1177) viruses. The initial PCR products were then further amplified using nested primers for the Hantaan (HF4 1141-1164, HB13 1360 1339) and Seoul (SF7 863-884, SB1 1165-1142 ) viruses. Amplified segments were then digested with restriction enzymes specific for either Hantaan (C1a I) or Seoul (Sac I) virus sequences. RESULTS: In all cultured viruses, the serotypes identified by nRT-PCR/RFLP were consistent with those of PRNT. nRT-PCR/RFLP results indicated the presence of Hantaan virus in 10 patients and of Seoul virus in 15 patients. In 3 patients, both Hantaan- and Seoul-specific amplified bands were visualized in serially collected samples, and in 4 patients no amplicon was detected. Among 69 specimens, 55 were positive; these positive specimens were obtained between days 3 and days 33 of illness. The positive rate was not affected by the clinical phase, day of illness, or severity of HFRS. CONCLUSIONS: nRT-PCR/RFLP is a rapid and convenient method for serotype diagnosis in most HFRS patients. It could also allow detection of genetic variation of hantavirus within the same serotype. PMID- 10711410 TI - Use of cultured tubular cells isolated from human urine for investigation of renal transporter. AB - AIM: To facilitate the understanding of the transporter function of human renal tubular cells, we have developed a simple method using primary cultured proximal tubule (PT) cells isolated from voided urine. METHODS: PT cells grown to confluence on glass coverslips could be identified by parallel arrays of spindle cells and hemicyst formation. Brush-border gamma-glutamyl transpeptidase (gammaGTP) activity was histochemically identified. Apical membrane Na+/H+ exchanger (NHE) activity was measured by monitoring changes in intracellular pH (pHi) after an acid load in a single cell level using the pH-sensitive dye 2'7' bis-(2-carboxyethyl)-5.6'carboxyfluorescein (BCECF). RESULTS: Amiloride and 5-(N ethyl-N-isopropyl) amiloride (EIPA) inhibited the NHE activity with half-maximal inhibition values (IC50) of 15.3 and 4.0 microM, respectively. NHE-3 mRNA was detected by the RT-PCR technique in clonally proliferated PT cells. CONCLUSION: These results suggest that cultured PT cells isolated from human urine express amiloride-resistant NHE-3 activity on the apical membranes, which can be compared to functional properties of PT in vivo. Our experimental strategy offers a useful experimental approach to investigating human renal tubular transport function in vitro. PMID- 10711411 TI - Influence of growth hormone on whole body and regional soft tissue composition in adult patients on hemodialysis. A double-blind, randomized, placebo-controlled study. AB - BACKGROUND: Many adult patients in chronic hemodialysis exhibit malnourishment and muscle wasting, which in some may be due partly to blockage of the biological action of growth hormone and the somatomedines. Growth hormone (GH) promotes protein synthesis, and long-term treatment with growth hormone has induced an augmentation in lean body-mass (LBM) in normal elderly persons, in persons with GH deficiency as well as growth improvement in uremic children. The purpose of this study was to evaluate the effect of long-term GH treatment on soft tissues in hemodialyzed patients by dual-energy X-ray absorptiometry (DXA) with special regard to the improvement in lean body mass and fat mass (FM). DESIGN: The study was double-blinded, randomized, and placebo-controlled. Twenty enfeebled patients in chronic hemodialysis were treated by subcutaneous injections of biosynthetic human GH (4 IU/m2 per day) or placebo, given every evening for 6 months. Soft tissues as LBM and FM, were measured by DXA scan, and height, and weight were recorded before, and after 6 months treatment. Serum concentration of insulin like growth factor (IGF-I) and type III collagen N-terminal propeptide (PIIINP) were analyzed at baseline and after 2, 4 and 6 months. RESULTS: Six months of GH therapy induced a total FM reduction of 3.05 +/- 0.75 kg (mean +/- SEM) (p < 0.001) (n = 9) corresponding to 25% of the total fat mass. The reduction in fat was most marked at the trunk, i.e. 1.39 +/- 0.41 kg (p < 0.001) corresponding to 40% of the total FM reduction. Total LBM increased by 3.14 +/-0.41 kg (p < 0.001) in the GH group. Regional changes for arm, truncus and leg in GH group amounted to 0.22 +/- 0.06 kg (p < 0.001), 1.64 +/- 0.37 kg (p < 0.001) and 0.51 +/- 0.06 kg (p < 0.001), respectively. In contrast, total body weight remained unchanged. Serum IGF-I increased from 199 +/- 14.8 microg/l to 527 +/- 111 microg/l (p < 0.0001) at month 6, and the serum PIIINP from 7.8 +/- 1.3/microg/l to 14.3 +/- 2.1 microg/l (p < 0.001) in the GH-treated group. In the placebo group (n = 11) there were no significant changes in FM, LBM or PIIINP while serum IGF-I decreased from 285 +/- 36 microg/l to 219 +/- 35 microg/l (p < 0.01) after 6 months treatment. CONCLUSIONS: Six months of GH therapy to patients with chronic renal failure resulted in marked changes of the soft tissue with an increase in LBM, and reduction of FM particularly at the trunk. The data imply that GH induced changes in body composition are maintained with long-term therapy. Very few side-effects of GH treatment were observed, and no serious ones were encountered, though the dosage were 2 to 3 times higher than the one given to GH insufficient, non-uremic persons, and the serum IGF-I concentrations during treatment equalized those seen in acromegalia. This indicates the existence of a reduced biological effect of GH and IGF-I in uremic persons. PMID- 10711412 TI - Bioimpedance resistance ratios for the evaluation of dry weight in hemodialysis. AB - MATERIAL: Restoration of body water compartments to normal by ultrafiltration is a major goal of hemodialysis. Dry weight is the term used to define normal body water in dialysis patients, but it is limited, as it is based solely on clinical observations. Bioimpedance spectroscopy can accurately measure the resistance of body fluid compartments. The ratio of the resistances of the intracellular to extracellular water should reflect the relative volume of these compartments. As dialysis patients accumulate excess fluid in their extracellular compartment, this ratio may prove useful in the evaluation of dry weight. METHODS: We measured the resistances of the intracellular and extracellular fluid compartments in normal subjects to define the normal ratio of the resistances of these compartments. Women had a slightly higher ratio than men (women: 2.41 +/- 0.23 vs. men: 2.08 +/- 0.23 vs. p < 0.0001). The ratios determined in the normal population were taken as the normal physiologic ratio and were used to define physiologic dry weight. We then compared dialysis patients both pre- and post dialysis to this normal population. RESULTS: We found that most patients (67%, n = 18) had an elevated ratio pre-dialysis suggesting excess extracellular fluid. Of the 38 treatments in which patients achieved their clinical dry weight, 19 (50%) had persistently elevated Ri/Re ratios, suggesting they had not reached physiologic dry weight. CONCLUSION: These data suggest that many dialysis patients carry excess extracellular fluid post dialysis despite achieving their clinical dry weight. Furthermore, the resistance ratio derived from bioimpedance spectroscopy may be a useful clinical tool in determining dry weight. PMID- 10711413 TI - TNF-alpha: mRNA, plasma protein levels and soluble receptors in patients on chronic hemodialysis, on CAPD and with end-stage renal failure. AB - BACKGROUND: Patients on hemodialysis suffer from an impaired immunity against infectious agents, hyporesponsiveness to vaccination and are prone to develop malignancies. This clinical state of immunoincompetence may be due to a disbalance in their defense mechanisms in which TNF-alpha and its soluble receptors 1 and 2 play a central role. PATIENTS AND METHODS: We measured, with double-sandwich ELISA, the levels of TNF-alpha and the soluble TNF-receptors in peripheral blood of patients on chronic intermittent hemodialysis (CIHD), on peritoneal dialysis (CAPD) and pre-dialysis end-stage renal failure (ESRF). Using reverse transcriptase polymerase chain reaction (RT-PCR) analysis, we quantified the amount of TNF-alpha mRNA in peripheral blood mononuclear cells (PBMC) obtained from these patient groups. RESULTS: In none of the patient groups, elevated levels of TNF-alpha were detected with ELISA, while high levels of soluble TNF receptors were present in ESRF, CAPD and CIHD patients. This may be the result of an activated TNF-alpha system or due to their impaired renal clearance. TNF-alpha mRNA level was elevated in CIHD patients compared to ESRF and CAPD patients or healthy controls. CONCLUSION: This suggests that only during chronic HD is the TNF-alpha system activated. High levels of sTNFR, found in ESRF or CAPD patients do not reflect activation of TNF-alpha system, but are the result of impaired renal clearance of the receptors. Indeed, we found a strong linear correlation between the levels of sTNF receptors and renal function. Nevertheless, these high levels of sTNF receptors are biological active, as they were able to bind active TNF-alpha up to 75% (range 46 - 83%) and thus inhibit the bioactivity and bioavailability of produced TNF-alpha. This may play a role in the immunoincompetence of these patients. PMID- 10711414 TI - Metabolic acidosis and composite nutritional index (CNI) in CAPD patients. AB - BACKGROUND: Metabolic acidosis (MA) has been recognized as an important stimulus for net protein catabolism. Continuous ambulatory peritoneal dialysis (CAPD) is regarded to have advantage in normalizing the acid-base homeostasis over hemodialysis due to continuous supply of buffer. However, many CAPD patients still remain acidotic and the clinical impact of this MA is uncertain. Recent studies revealed no specific correlation between a certain degree of MA and protein nutritional status while others showed that mild alkalosis is nutritionally beneficial to CAPD patients. A cross-sectional study evaluating acid-base and nutritional status was performed to examine the relationship between acid-base status and overall nutritional status assessed by composite nutritional index (CNI) and to get a basic information about the optimal pH/HCO3 in CAPD patients. PATIENTS AND METHODS: Total 198 clinically stable patients maintained on CAPD more than 6 months were evaluated. Each of 10 nutritional parameters of CNI consisting of clinical (subjective global assessment), biochemical (total lymphocyte count, albumin, prealbumin, insulin-like growth factor-1, transferrin) and anthropometric parameters (body mass index, % lean body mass, triceps skinfold thickness, midarm muscle circumference) was graded from 0 to 3 point (0; normal, 1; mildly decreased, 2; moderately decreased, 3; severely decreased). RESULTS: Mean CNI score was 8.2 +/- 5.2 with a range of 0 to 25. CNI was positively correlated with age, duration of peritoneal dialysis, incidence of peritonitis, C-reactive protein (CRP), HCO3 and dialytic protein loss whereas it was inversely correlated with hemoglobin and nPNA. In patients with MA (n = 25, mean arterial HCO3 19.5 +/- 1.9 mM/l), CNI score was significantly lower (6.3 +/- 3.5) compared to patients with normal acid-base status (n = 63, 9.5 +/- 5.9, p < 0.01) or metabolic alkalosis (n = 47, 10.1 +/- 4.6, p < 0.001). Multiple regression analysis revealed that the incidence of peritonitis, duration of dialysis, CRP and dialytic protein loss were the independent predictors of CNI. CONCLUSIONS: CAPD patients with mild to moderate degree of MA appear to be associated with more favorable overall nutritional status expressed as CNI. Prospective studies evaluating changes in the nutritional parameters with the correction of acid-base abnormality are needed to understand the real impact of acid-base status on nutritional status and to know the optimal pH/HCO3 in CAPD patients. PMID- 10711415 TI - Epstein-Barr virus in pediatric patients after renal transplantation. AB - BACKGROUND: Pediatric allograft recipients are at an increased risk for lymphotropic virus-associated disorders, particularly in association with primary EBV infection. PATIENTS AND METHODS: Twenty-nine children, adolescents and young adults after renal transplantation were studied in comparison with a healthy young adult control group for evidence of primary, reactivated or chronic active EBV infection at two different time points. RESULTS: Prevalence of antibodies against viral capsid antigen (VCA) was > or = 90% in both groups, whereas anti Epstein-Barr nuclear antigen (EBNA) was detected only in 19 of 26 seropositive patients compared with seropositive controls (p = 0.01). Persistence of EBV DNA in leukocytes for > or = 6 months was observed in 11 seropositive patients (38%) and one control patient (p < 0.007) using nested polymerase chain reaction. In the transplant recipients, 3 cases of primary EBV infection and 3 cases of chronic active EBV infection were identified. One of these cases developed a non Hodgkin lymphoma one year later. CONCLUSION: The results suggest that determination of pretransplant antibody status in recipients, rapid detection of EBV infection in seronegative symptomatic recipients, and regular screening for persistent EBV DNA in patients at risk to develop post-transplantation lymphoproliferative disease should be performed. PMID- 10711416 TI - A randomized placebo-controlled double-blind trial of lipid-lowering strategies in patients with renal insufficiency: diet modification with or without fenofibrate. AB - BACKGROUND: Renal insufficiency is characterized by lipoprotein abnormalities including elevated triglyceride levels. PATIENTS AND METHODS: The safety and efficacy of micronized fenofibrate as a treatment for dyslipidemia in patients with progressive renal insufficiency was evaluated in a randomized, placebo controlled double-blind study comparing fenofibrate and dietary modification to dietary modification alone. Patients were evaluated following a 3-month pre randomization period of dietary counseling. Twenty-eight patients with moderate renal insufficiency and triglyceride levels 2.3 mmol/l or LDL/HDL ratio 5 were randomized to placebo (n = 12) or fenofibrate (n = 16) therapy. Treatment and dietary counseling continued for 6 months. RESULTS: Ten of 16 patients (63%) treated with fenofibrate achieved a 30% reduction in triglyceride levels or LDL/HDL ratio reduction < 5 compared to 2 of 17% in the placebo group (p = 0.015). Triglyceride levels were significantly reduced in the fenofibrate group ( 31%) versus placebo (+1.3%, p = 0.003). In compliant patients (n = 25) there was also a significantly greater increase in HDL cholesterol levels in the fenofibrate group (+19.9%) compared to placebo (-4.7%, p = 0.001). Changes in measured creatinine clearance were not significantly different between the groups and there were no serious adverse effects of treatment. CONCLUSION: Fenofibrate therapy combined with dietary modification effectively reduced triglyceride levels in renal insufficiency patients without serious adverse effects. PMID- 10711417 TI - Severe interstitial nephritis in a patient with renal amyloidosis and exacerbation of Crohn's disease. AB - A 57-year-old man with long-term untreated Crohn's disease presented with exacerbation of his bowel disease, volume depletion, nephrotic syndrome and rapid decline in renal function. Renal biopsy revealed amyloidosis and extensive interstitial infiltration. Initiation of steroid therapy was associated with improvement in renal function and postponement of dialysis, suggesting that control of interstitial inflammation might have a therapeutic role in renal amyloidosis. We hypothesize that volume depletion could magnify toxicity of proteinuria, thus augmenting interstitial inflammation and accelerating the deterioration in renal function. PMID- 10711418 TI - Minimal change glomerulonephritis associated with hydatid disease. AB - A 63-year-old man presented to our department with dyspnea and peripheral edema. A cystic mass in the right upper abdomen, consistent with echinococcal disease was discovered. Proteinuria was also present, and a nephrotic syndrome was diagnosed. The kidney biopsy revealed minimal change glomerulonephritis. Treatment with the antiechinococcal drug albendazole induced complete remission of the nephrotic syndrome, suggesting an etiopathogenic role for a hydatid antigen in the development of an immune-mediated glomerulonephritis. PMID- 10711419 TI - Urinary activity of beta-glucuronidase and excretion of glycosaminoglycans in the diagnosis of diabetic nephropathy. PMID- 10711420 TI - Uncoupling of apoptosis and Jun/AP-1 activity in human promonocytic cells treated with DNA-damaging and stress-inducing agents. AB - Earlier studies have indicated that Jun/AP-1 activity is associated with, and probably required for apoptosis induction by DNA-damaging and stress-inducing agents in human myeloid cells. To investigate this possibility, we examined the capacity of continuous treatments with etoposide (10 microM) and camptothecin (0.4 microM), and pulse treatments with X-rays (20 Gy), heat (2 h at 42.5 C) and cadmium chloride (2 h at 200 microM) followed by recovery, to provoke apoptosis and to simulate c-jun and c-fos expression and AP-1 binding in U-937 human promonocytic cells. All these treatments generated apoptosis with similar efficacy (50-60% apoptotic cells at 6 h of treatment or recovery). However, the capacity to increase c-jun and c-fos mRNA levels and to stimulate AP-1 binding was very different, ranging from more than a twelve-fold increase in the case of cadmium, to almost no increase in the case of heat-shock and etoposide. When the cells were pre-conditioned with a soft heat shock (1 h at 42 degrees C) the cadmium-provoked apoptosis was greatly inhibited, but the stimulation of AP-1 binding was not affected. The administration of cAMP-increasing agents also reduced the etoposide- and cadmium-provoked apoptosis. However, cAMP greatly stimulated c-jun and c-fos expression and AP-1 binding when applied together with etoposide (which itself was ineffective), and potentiated the cadmium-induced AP 1 binding. Conversely, retinoic acid abrogated the cadmium-provoked stimulation of AP-1 binding and transactivation capacity, and greatly inhibited the stimulation of binding caused by camptothecin and X-rays. However, retinoic acid did not inhibit the induction of apoptosis by these agents. These results indicate that Jun/AP-1 activity is not necessarily coupled with apoptosis, nor required for apoptosis induction by DNA-damaging and stress-inducing agents in human promonocytic cells. PMID- 10711421 TI - Rearrangements of sea urchin egg cytoplasmic membrane domains at fertilization. AB - Fertilization in the sea urchin is accompanied by rapid reorganization of the egg endoplasmic reticulum (ER). ER-derived vesicles contribute to one of three classes of membranes used in assembling the male pronuclear envelope in vitro. We provide here biochemical evidence for the rearrangement of sea urchin egg cytoplasmic membrane domains at fertilization up to the first mitosis, with respect to two nuclear envelope markers, lamin B and lamin B receptor (LBR), using purified vesicles prepared from homogenates fractionated by floatation on sucrose gradients. In unfertilized eggs, immunoprecipitation data indicate that most of lamin B and LBR are localized in the same vesicles but do not interact. By 3 min post-fertilization, both proteins are more widely distributed across the gradients and by 12 min most of lamin B and LBR are localized in vesicles of different densities. This partitioning is maintained throughout S phase. At mitosis, most lamin B and LBR remain in distinct vesicles, while a small proportion of lamin B and LBR, likely derived from the disassembled nuclear envelope, associate in a minor subset of vesicles. The results illustrate a dynamic reorganization of egg cytoplasmic membranes at fertilization, and the establishment of distinct membrane domains enriched in specific nuclear envelope markers during the first cell cycle of sea urchin development. Additionally, we demonstrate that male pro-nuclear membrane assembly occurs only when both cytosol and membranes originate from fertilized but not unfertilized eggs, suggesting that fertilization-induced membrane rearrangements contribute to the ability of the egg to assemble the male pronuclear envelope. PMID- 10711422 TI - Characterisation and tissue-specific expression of the two keratin subfamilies of intermediate filament proteins in the cephalochordate Branchiostoma. AB - The cloning of three intermediate filament proteins expressed at the gastrula stage (kl, Y1, X1) extends the size of the IF multigene family of Branchiostoma to at least 13 members. This is one of the largest protein families established for the lancelet. Sequence comparisons indicate five keratin orthologs, three of type I (E1, k1, Y1) and two of type II (E2, D1). This assignment is confirmed by the obligatory heteropolymeric polymerisation behaviour of the recombinant proteins. In line with the hetero-coiled-coil principle IF are formed by any stoichiometric mixture of type I and II keratin orthologs. In spite of the strong sequence drift chimeric IF are formed between K8, a human keratin II, and two of the lancelet type I keratins. We discuss whether the remaining 8 IF proteins reflect three additional and potentially cephalochordate-specific subfamilies. The tissue-specific expression patterns of the 5 keratins and some other IF proteins were analysed by immunofluorescence in the adult. Keratins are primarily present in ectodermally derived tissues. Developmental control of the expression of some IF proteins is observed, but three keratins (k1, Y1, D1) and an additional IF protein (X1) detected at the gastrula stage are expressed throughout the life cycle. PMID- 10711423 TI - Intracisternal granules in the adipokinetic cells of locusts are not degraded and apparently function as supplementary stores of secretory material. AB - The intracisternal granules in locust adipokinetic cells appear to represent accumulations of secretory material within cisternae of the rough endoplasmic reticulum. An important question is whether these granules are destined for degradation or represent stores of (pro)hormones. Two strategies were used to answer this question. First, cytochemistry was applied to elucidate the properties of intracisternal granules. The endocytic tracers horseradish peroxidase and wheat-germ agglutinin-conjugated horseradish peroxidase were used to facilitate the identification of endocytic, autophagic, and lysosomal organelles, which may be involved in the degradation of intracisternal granules. No intracisternal granules could be found within autophagosomes, and granules fused with endocytic and lysosomal organelles were not observed, nor could tracer be found within the granules. The lysosomal enzyme acid phosphatase was absent from the granules. Second, biochemical analysis of the content of intracisternal granules revealed that these granules contain prohormones as well as hormones. Prohormones were present in relatively higher amounts compared with ordinary secretory granules. Since the intracisternal granules in locust adipokinetic cells are not degraded and contain intact (pro)hormones it is concluded that they function as supplementary stores of secretory material. PMID- 10711424 TI - Immunocytochemical localization of a urate oxidase immunoreactive protein in the plasma membranes and membranes of the secretory/endocytic compartments of digestive gland cells of the mussel Mytilus galloprovincialis. AB - The subcellular compartmentalization of urate oxidase (UOX) in the digestive glands of mussels, Mytilus galloprovincialis Lmk, was studied by means of immunoblotting and immunocytochemistry, using an antibody raised in rabbit against rat liver UOX. Western blot analysis of subcellular fractions revealed an immunoreactive polypeptide with a molecular weight similar to the corresponding mammalian hepatic protein. This crossreactive polypeptide of 32 kDa was particle bound yet not peroxisome-associated. In paraffin sections the antiserum specifically labeled the plasma membrane of the digestive gland epithelial cells and discrete regions within the perinuclear and apical portions of the digestive tubules and duct cells. By electron microscopy gold particles representing antigenic sites were found on the microvilli and the lateral plasma membrane as well as the membranes of the secretory/ endocytic compartments, that is, the Golgi complex, secretory and some endocytic vesicle membranes. Since the peroxisomal UOX-antibody exhibits a comparable immunoreactivity towards a urate transporter channel protein in rat kidney proximal tubules and has been used for its molecular cloning (Leal-Pinto et al., 1997, J. Biol. Chem. 272, 617-625), we suggest that the membrane protein identified in mussel digestive glands could represent a homologous urate-transporter protein. PMID- 10711425 TI - The tetraspanin CD9 associates with the integrin alpha6beta4 in cultured human epidermal keratinocytes and is involved in cell motility. AB - Integrins are involved in several ways in keratinocyte physiology, including cell motility. CD9 is a member of the tetraspanin protein family which is found in association with other transmembrane proteins like the integrins. CD9 is expressed in the epidermal tissue, but this expression is not regulated by differentiation. The present work focuses on association of CD9 with the integrin alpha6beta4 in keratinocytes. In vivo, CD9 does not co-localize with alpha6beta4, and is not internalized with the integrin upon basal detachment with dispase. In vitro, CD9 is found partly in co-localization with alpha6beta4 and is internalized with the integrin after keratinocyte detachment with dispase. Using blocking antibodies in a phagokinetic tracks assay, we show that CD9, and to a lesser extent alpha6beta4, but not the tetraspanin CD82, promote motility of subconfluent keratinocytes on collagen I. Our observations also suggest that CD9 is involved in the formation of lamellipodia. We also report that the phorbol ester TPA has no effect on CD9 expression and association with alpha6beta4, but increases keratinocyte motility, possibly through modulation of integrin subunits expression, or through upregulation of collagenase-1 expression. Together, these results confirm that CD9 associates with alpha6beta4 in cultured keratinocytes, possibly in order to regulate the function of the integrin, and that CD9 is involved in keratinocyte motility on collagen. The data suggest that regulation of adhesion characteristics by CD9 in keratinocytes may play a role in epidermal repair. PMID- 10711426 TI - Processing and localisation of a GPI-anchored Plasmodium falciparum surface protein expressed by the baculovirus system. AB - We describe the expression, in insect cells using the baculovirus system, of two protein fragments derived from the C-terminus of merozoite surface protein 1(MSP 1) of the human malaria parasite Plasmodium falciparum, and their glycosylation and intracellular location. The transport and intracellular localisation of the intact C-terminal MSP-1 fragment, modified by addition of a signal sequence for secretion, was compared with that of a similar control protein in which translation of the GPI-cleavage/attachment site was abolished by insertion of a stop codon into the DNA sequence. Both proteins could only be detected intracellularly, most likely in the endoplasmic reticulum. This lack of transport to the cell surface or beyond, was confirmed for both proteins by immunofluorescence with a specific antibody and characterisation of their N glycans. The N-glycans had not been processed by enzymes localised in post endoplasmic reticulum compartments. In contrast to MSP-1, the surface antigen SAG 1 of Toxoplasma gondii was efficiently transported out of the endoplasmic reticulum of insect cells and was located, at least in part, on the cell surface. No GPI-anchor could be detected for either of the MSP-1 constructs or SAG-1, showing that the difference in transport is a property of the individual proteins and cannot be attributed to the lack of a GPI-anchor. The different intracellular location and post-translational modification of recombinant proteins expressed in insect cells, as compared to the native proteins expressed in parasites, and the possible implications for vaccine development are discussed. PMID- 10711427 TI - Alterations in the glycoconjugates of pancreatic cell membrane induced by acute pancreatitis. AB - The alterations that progressively appear in plasma membrane glycoconjugates of rat pancreatic cells at different stages of acute pancreatitis induced by duct obstruction have been analyzed on individual cells by flow cytometry using the fluoresceinated lectins, wheat germ agglutinin (WGA), Tetragonolobus purpureus agglutinin (TP) and Concanavalin A (Con A), which specifically bind to N-acetyl D glucosamine, L-fucose and D-mannose, respectively. Two populations of pancreatic cells were differentiated according to the forward scatter (size), which showed different density of saccharidic terminals located at external positions in the glycoconjugates of the plasma membrane. A significant increase in WGA and TP binding was found 1.5 h after pancreatic obstruction, which could be due to the fusion of zymogen granules with the plasma membrane as suggested by the basolateral exocytosis observed by electron microscopy at this stage. The most external sugar residues of membrane glycoconjugates are removed 12 h after pancreatic duct obstruction as a consequence of an advanced state of pancreatitis. The hydrolytic process reaches greater depths in the membrane 48 h after obstruction. At this stage a significant decrease in WGA, TP and ConA binding was found in all pancreatic cells, indicating the loss of N-acetyl D glucosamine and/or sialic acid, L-fucose and even D-mannose which is located in the core of the glycan. The results provide information about the progressive degradation induced by acute pancreatitis in pancreatic cell membrane glycoconjugates. PMID- 10711428 TI - Esophageal histology does not provide additional useful information over clinical assessment in identifying reflux patients presenting for esophagogastroduodenoscopy. AB - We prospectively evaluated the value of histology in identifying gastroesophageal reflux disease (GERD) in consecutive patients enrolled for upper endoscopy. GERD was defined as heartburn occurring at least weekly. Macroscopic esophagitis was graded and an esophageal biopsy was taken 2 cm above the gastroesophageal junction. Histological esophagitis was identified by: (1) basal cell hyperplasia >15%, (2) increased papillary length >66%, and (3) infiltration by leukocytes/eosinophils. The sensitivity, specificity, and positive and negative predictive value of histological esophagitis in patients with and without typical reflux symptoms, with and without endoscopic changes, or both were evaluated. Of 178 patients, reflux symptoms were present in 59% (N = 105) and esophageal erosions in 19% (N = 34); 75 patients had reflux symptoms but no erosions. While the specificity of histology was adequate (78%), it was insensitive (30%). The positive and negative predictive values were 67% and 44%, respectively. No single individual parameter was better than any other. Thus, histology appears to be of no additional value in identifying GERD. PMID- 10711429 TI - Methylene blue staining and endoscopic ultrasound evaluation of Barrett's esophagus with low-grade dysplasia. AB - This study was performed to determine if either methylene blue staining or endoscopic ultrasound helped direct biopsies in patients with a history of Barrett's esophagus with low-grade dysplasia. Patients underwent radial endoscopic ultrasound scanning to measure esophageal wall thickness, followed by endoscopy with methylene blue staining and biopsies. Mean esophageal wall thickness for squamous mucosa (2.3 +/- 0.2 mm), nondysplastic Barrett's (2.6 +/- 0.2 mm), and Barrett's with dysplasia (2.9 +/- 0.3 mm) were similar. With staining, Barrett's mucosa stained blue more often than gastric epithelium (68% vs 15%, respectively; P < 0.001). The sensitivity and specificity for strong staining detecting Barrett's were 68% and 85%, respectively. Barrett's with low grade dysplasia stained blue less frequently (52%) than nondysplastic Barrett's (74%; P < 0.05), but the positive predictive value for poor staining indicating dysplasia was 41%. Endoscopic ultrasound was not helpful in directing biopsies in these patients. The utility of methylene blue for detecting dysplasia needs further investigation. PMID- 10711430 TI - Effect of bile and pancreatic juice on adenoviral-mediated gene delivery: implications on the feasibility of gene delivery through ERCP. AB - Current research in gene delivery to the liver is focused on the intravenous, intraarterial, intraportal, or intratumoral route. Another possible route for gene delivery is via the common bile duct through endoscopic retrograde cholangiopancreatography (ERCP). Whether bile and pancreatic juice have any effect on gene delivery is not established. To evaluate the effect of bile and pancreatic juice on adenoviral-mediated gene delivery, liver and pancreatic cell lines were infected with a recombinant adenovirus expressing an E. coli beta galactosidase gene under the control of a cytomegalovirus promoter (rAdCMVpLacZ) in the absence or presence of various concentrations of bile and pancreatic juice. The proportion of cells infected was evaluated through X-gal staining. The toxicity of bile and pancreatic juice was also evaluated through cell morphology and detachment. Bile appeared to induce significant cytotoxicity in HepG2 and Huh7 cells (50% viability with 15 min of incubation). Neither bile nor pancreatic juice affected transgene expression. In the absence of bile/pancreatic juice, HepG2 (15-25%) and PANC-1 cells (10-18%) were less susceptible to rAdCMVpLacZ compared to Huh7 cells (75-84%, vs HepG2, P < 0.001) and BxPc-3 (82-95%, vs PANC 1, P < 0.001) at a multiplicity of infection (MOI) of 5. Bile reduced the transduction efficiency, but 5-10% HepG2 and 5-42% of Huh7 cells were still transduced in the presence of 80% bile for up to 10 min. Adenoviral-mediated gene delivery was reduced in the presence of pancreatic juice with a low multiplicity of infection (MOI of 5), but this effect was negated with an MOI of 50. These data provide encouragement to develop adenoviral-mediated gene delivery through ERCP. PMID- 10711431 TI - Endoscopic transpapillary drainage of an infected pancreatic fluid collection in pancreas divisum. PMID- 10711432 TI - Measurement and evaluation of gastric emptying using radiopaque barium markers. AB - To simplify assessment of gastric emptying, we have developed a radiopaque barium marker method. The subjects were 11 healthy volunteers, 30 patients with progressive systemic sclerosis, 16 patients with dysmotility-like dyspepsia, 7 patients with irritable bowel syndrome, and 6 patients with diabetes mellitus. We tested three types of radiopaque markers with manometry by a three-channel strain gauge transducer. The 4.5-mm ring-shaped markers with a specific gravity of 1.2 were emptied from the stomach in correlation with the number of high-amplitude (>50 mm Hg) antral contractions. The percentage of residual markers at 2 hr was significantly (P < 0.05) lower in patients with irritable bowed syndrome than in normal controls, and at 5 hr it was significantly (P < 0.05) higher in patients with systemic sclerosis with esophageal dysmotility, dysmotility-like dyspepsia, or diabetes than in normal controls. This radiopaque barium marker method may be useful as a screening test for determining whether gastric emptying is rapid or delayed. PMID- 10711433 TI - Role of anger in antral motor activity in irritable bowel syndrome. AB - There is considerable evidence indicating that patients with irritable bowel syndrome respond to emotional and environmental stimulation with increased colon motor activity. It has been suggested also that increased colon motor activity is not confined to the colon and may be representative of a broader disorder affecting the rest of the gastrointestinal tract in this population. The results of our current study suggest that anger may have a significant, although differential effect on antral motor activity in IBS patients compared to normal controls. We found that while antral motor activity did not differ significantly in our groups during rest, anger decreased antral motor activity in IBS patients and increased antral motor activity in normal controls. The difference was not attributable to a difference in anger levels since the groups did not differ in their response to the standardized anger stressor. Rather, the difference in the antral motor response appears to be qualitative and a possible marker for irritable bowel syndrome. Our data further suggest that increased colon motor activity in IBS patients during emotional stress is not a result of a rise in motor activity throughout the gastrointestinal tract, but a phenomenon that may be unique to the colon in this patient population. PMID- 10711434 TI - Gastric emptying and dyspeptic symptoms in patients with nonautoimmune fundic atrophic gastritis. AB - Our aim was to evaluate the relationship between gastric emptying and demographic, clinical, histological, and secretory features in patients with nonautoimmune fundic atrophic gastritis. Only 31% of 45 patients with fundic atrophic gastritis presented with achlorhydria. Scintigraphic gastric emptying of solids was delayed compared to healthy controls. Patients with achlorhydria showed gastric emptying rates lower than those with preserved acid secretion. Significant, but weak, correlations were observed between emptying rates and both peak acid output (Rs = 0.33) and serum gastrin levels (Rs = -0.36), but not with grading of mucosal atrophy. No symptom differences were observed between patients with or without achlorhydria, but a weak correlation was detected between peak acid output and the severity of epigastric pain (Rs = 0.40). In conclusion, patients with fundic atrophic gastritis present delayed gastric emptying that is weakly related to the reduction of the acid secretion and the raising of serum gastrin levels rather than to the severity of the atrophy. PMID- 10711435 TI - A normal gastrointestinal motility excludes chronic intestinal pseudoobstruction in children. AB - Gastrointestinal manometry has gained wide acceptance in the approach to patients with suspected enteric neuromuscular disorders. However, performing gastrointestinal manometry in these subjects without a previous exhaustive diagnostic evaluation is unjustified. Twelve children (median age: 7.0 years; range: 8 months-13 years), with clinical and x-ray features suggesting chronic intestinal pseudoobstruction, were referred to our unit for gastrointestinal manometry. The latter was performed with a perfused catheter for 5 hr in the fasting state and for 90 min after feeding. Data were compared with those recorded in eight age-matched controls. In all patients and controls, interdigestive motor complexes with propagated phases III were detected; a regular postprandial antroduodenal motor activity was also recorded. Patients and controls did not differ for fed antral and duodenal motility indexes, fed antroduodenal coordination, and length of duodenal phase III. Most of the patients showed short or prolonged bursts of nonpropagated activity in the fasting and/or fed states; in four cases fasting and/or fed sustained phasic activity was recorded. Manometric evidence of migrating motor complexes and postfeeding activity did not support the diagnosis of intestinal pseudoobstruction and suggested redirecting the diagnostic evaluation. Final diagnoses were: Munchausen syndrome-by-proxy (four cases), celiac disease (two cases), intestinal malrotation (two cases), Crohn's disease (two cases), multiple food intolerance (one case), and congenital chloride-losing diarrhea (one case). It is concluded that in children with suspected chronic intestinal pseudoobstruction manometric evidence of migrating motor complexes and fed motor activity excludes an enteric neuromuscular disorder and suggests a reassessment of the diagnostic work-up. Furthermore, if gastrointestinal manometry shows migrating motor complexes and postfeeding motor activity, qualitative abnormalities of the manometric tracings do not indicate an underlying enteric neuromuscular disorder and must not be overemphasized. Patients referred for gastrointestinal manometry should previously undergo an extensive diagnostic investigation to exclude disorders mimicking chronic intestinal pseudoobstruction. PMID- 10711436 TI - Treatment of Helicobacter pylori infection in clinical practice in the United States: results from 224 patients. AB - Our objectives were to define treatment success, compliance, and side effects for treatment of Helicobacter pylori in clinical practice. In all, 224 consecutive patients received Helicobacter pylori treatment: 97 received two weeks of bismuth subsalicylate, metronidazole, tetracycline four times a day with a H2-receptor antagonist twice a day (BMT); 89 received one week of metronidazole, lansoprazole, and clarithromycin twice a day (MLC); and 38 received one week of BMT with lansoprazole twice a day (BMT-PPI). Cure rates were: BMT 81% (95% CI 74 89%), MLC 90% (95% CI 84-96%) BMT-PPI 87% (95% CI 81-92%). More patients prescribed a bismuth-based regimen discontinued medications due to side effects compared to MLC (P = 0.049). Nausea was more common for BMT compared to MLC (P = 0.04). In conclusion, treatment of Helicobacter pylori infection with a one-week course of MLC achieves a high rate of cure in clinical practice. Significantly fewer patients prescribed PPI-based therapy discontinue medications due to side effects as compared to bismuth-based triple therapy. PMID- 10711437 TI - Isolation of bacteria other than Helicobacter pylori from stomachs of squirrel monkeys (Saimiri spp.) with gastritis. AB - Gastric biopsy specimens obtained from 12 squirrel monkeys (Saimiri spp.) were investigated by culture for the presence of bacteria. The stomachs of two monkeys with gastritis were colonized with gram-negative, urease-positive bacteria, identified as Ochrobactrum anthropi by the Vitek and API NFT methods (BioMerieux). A third monkey with gastritis was positive for Aeromonas salmonicida and Pseudomonas vesicularis (both urease-negative). No Helicobacter pylori was isolated from squirrel monkeys. Light microscopic and transmission electron microscopic examination revealed that the O. anthropi isolates were covered by extracellular material, indicating a capsule. Characterization of the O. anthropi urease revealed Michaelis-Menten constants (Km values) of 6.2 and 4.0 mM urea for the ureases of O. anthropi isolates S664 and S1835, respectively, and 3.7 for type strain 49188. Western blot analysis using H. pylori- and H. felis specific antibodies detected shared antigenic epitopes between the ureases of H. pylori, H. felis, and O. anthropi. The apparent molecular mass of the urease enzymes of the O. anthropi isolates was determined on 6% nondenaturing gels to be approximately 82 kDa. Antimicrobial susceptibility tests, using the MicroScan method (Dade International), revealed multidrug resistance for the O. anthropi isolates with susceptibilities for the antibiotics amikacin, ciprofloxacin, gentamicin, cefoperazone, tobramycin, imipenem, and trimethoprim/sulfamethoxazole. PMID- 10711438 TI - Effectiveness of single dilation with Maloney dilator versus endoscopic rupture of Schatzki's ring using biopsy forceps. AB - Current recommendations for treatment of patients with symptomatic Schatzki's ring are based on anecdotal experience or uncontrolled studies. Maloney dilation is the gold standard. We performed a randomized controlled trial to compare the use of a single 52-Fr Maloney dilation versus four quadrant biopsy of Schatzki's ring for relief of dysphagia. The subjects answered standardized dysphagia related questions on a scale of 0-5 (0 = no dysphagia; 5 = cannot handle secretions). To account for modifications in diet and eating habits, subjects answered 11 question to arrive at a eating/diet score. Patients with Schatzki's ring were randomized into one of the two protocols. Group 1 underwent endoscopic biopsies of the ring, one biopsy in each quadrant. In group 2, the endoscope was taken out, and a single 52-Fr Maloney dilation was performed. Twenty-six patients participated in the study and were followed for up to 15 months. There was no significant difference in age, sex, race, smoking, alcohol abuse, or medication intake between the two groups. Dysphagia score improved by 91% in both groups at three months and 84% and 85% at 12 months in groups 1 and 2, respectively. The eating/diet habit score improved by 78% in both groups. There was one failure in each group, and one recurrence at six months in the dilation group. Fifty-five percent of dilation group and 100% of biopsy group described the procedure as easy. There was no difference in the amount of sedatives used during the procedure or the acid blockers after the procedure. In patients undergoing endoscopy, the superior cost/safety profile of endoscopic biopsy makes it a preferred choice for treatment of Schatzki's ring over bougienage. PMID- 10711439 TI - Characterization of esophageal striated muscle in patients with achalasia. AB - Many studies have been conducted analyzing the manometric properties of patients with achalasia, but the striated portion of the esophagus has never been analyzed and is often overlooked. We retrospectively reviewed 120 manometric tracings (20 achalasia, 100 controls) performed between 1994 and 1997 and excluded tracings from patients with chronic cough and nutcracker esophagus. The data were assessed for age, sex, symptoms, duration of symptoms, lower esophageal sphincter pressure, gastroesophageal gradient, upper esophageal sphincter pressure, smooth muscle contraction amplitude and duration, striated muscle contraction amplitude and duration, length from upper esophageal sphincter to maximal striated muscle contraction, and esophageal length. The maximum striated muscle contraction amplitude was significantly decreased in achalasia patients with a median amplitude of 45 mm Hg (range 12-95) vs 76 mm Hg (range 30-210) in the control group (P = 0.002). Although the wave forms were similar, the maximum striated muscle contraction duration and the distance from the upper esophageal sphincter in achalasia patients was not significantly different from controls. The length of the esophagus was significantly longer in achalasia patients with a median value of 25 cm (range 21-30) vs 21 cm (range 17-26) in the control group (P < 0.001). Patients with achalasia have significantly lower maximum striated muscle contraction amplitudes and longer esophagi, but the duration of the contractions and the configuration of the wave forms are not different. PMID- 10711440 TI - Effects of pro-inflammatory cytokines on acid secretion. PMID- 10711441 TI - Spontaneous apoptotic DNA fragmentation in cultured guinea pig gastric mucosal cells. AB - The purpose of this study was to elucidate the mechanism of spontaneous and rapid cell death of cultured gastric pit cells. Gastric pit cells have a rapid cell turnover rate in vivo. We here show that guinea pig gastric pit cells in culture undergo spontaneous and rapid apoptotic DNA fragmentation, which may represent the rapid cell turnover cycle of gastric pit cells in vivo. This spontaneous apoptotic DNA fragmentation required the presence of fetal calf serum in the culture media. Furthermore, the spontaneous apoptotic DNA fragmentation was prevented by protein synthesis and caspase inhibitors. PMID- 10711442 TI - A splice variant of the transcript for guanylyl cyclase C is expressed in human colon and colorectal cancer cells. AB - Guanylyl cyclase C is a sensitive and specific biomarker for metastatic colorectal cancer. A variant of the guanylyl cyclase C transcript was identified that possesses a 142-bp deletion at the 3' end of exon 1 reflecting alternative splicing of mRNA, introducing a shift in the open reading frame that prevents translation of a guanylyl cyclase C-related product. This variant was identified in human intestine and colon carcinomas, but not in extraintestinal tissues or tumors. These studies demonstrate that GCC and the splice variant contribute to the pool of GCC transcripts detected by RT-PCR in human tissues. They indicate that primers for RT-PCR that amplify regions downstream from the deletion are required to assess the full complement of GCC transcripts (GCC + GCC(var)) in human tissues and body fluids for staging and postoperative surveillance of patients with colorectal cancer. PMID- 10711443 TI - Allelic imbalance on 16q in small, unifocal hepatocellular carcinoma: correlation with HBV and HCV infections and cellular proliferation rate. AB - In advanced hepatocellular carcinoma (HCC), allelic loss on chromosome 16q may occur. To better define the frequency of this alteration in small HCC and to more closely identify the affected region for further positional cloning of the putative tumor suppressor gene contained in this region, microsatellite polymorphism analysis was conducted on small, unifocal HCC, without signs of intrahepatic or systemic spread. We also tried to assess its possible correlation with hepatitis virus infections (HBV and HCV) and cellular proliferation rate. DNA from 35 small (<4 cm), unifocal HCC and from the corresponding nontumorous surrounding tissue was analyzed by 10 sets of microsatellite polymorphic markers. Serologic markers for hepatitis virus B and C infections were investigated in all cases. AgNOR protein quantity was assessed by image analysis on cryostatic sections stained with silver. The percentage of tumours with allelic imbalance ranged from 11.1 to 37%. The minimal involved region was assessed at 16q24.3, corresponding to the D16S413 marker, which was also the most commonly affected locus (10 of 27 informative cases, 37%). Allelic imbalance on chromosome 16q was significantly associated with HBV infection: 8 of 10 cases showed an actual or previous HBV infection in the group showing allelic imbalance, versus 6 with a previous HBV infection out of 25 in the control group (P < 0.01). No difference was found as far as HCV infection is concerned. The mean (+/-SE) AgNOR protein value in six cases showing allelic imbalance was 8.36 +/- 1.2 microm2, compared to 6.45 +/- 0.68 microm2 in 13 cases retaining both the alleles at 16q but the difference proved not statistically significant. In conclusion, in this series of small, unifocal HCC the minimal region of allelic imbalance on 16q was restricted to 16q24.3. It was found to be associated with HBV infection but not with increased cellular proliferation rate. PMID- 10711444 TI - Impact of concomitant hepatolithiasis on patients with peripheral cholangiocarcinoma. AB - The association of hepatolithiasis (HL) and peripheral cholangiocarcinoma (PCC) has been well recognized. However, information concerning the impact of hepatolithiasis on patients with peripheral cholangiocarcinoma is sparse and therefore difficult to assess. A total of 162 consecutive patients with histologically proven peripheral cholangiocarcinoma were treated surgically at Chang-Gung Memorial Hospital between 1977 and 1994. Among them, 106 patients (65.4%) had associated hepatolithiasis (PCC + HL group), and the remaining 56 patients (34.6%) did not (the PCC - HL group). The differences in demographics, symptomatology, laboratory data, tumor staging, histological pattern, resectability rates, and long-term survival of these two groups were compared. The male to female ratio was 0.7 in the PCC + HL group and 1.3 in the PCC - HL group (P < 0.05). Two thirds of the PCC + HL group presented with acute cholangitis, whereas two thirds of the PCC - HL group presented with hepatomegaly (P < 0.01). Those patients in the PCC + HL group were in earlier stages than those of the PCC - HL group at the time of the initial diagnosis (P < 0.05). The resectability rate for the PCC + HL group was 31.1% and for the PCC - HL group, 26.8% (P > 0.05). Surgical mortality rates were 3.8% in the PCC + HL group and 3.6% in the PCC - HL group (P > 0.05). The morbidity rate was much higher in the PCC + HL group than in the PCC - HL group (P < 0.01). The 1-, 3-, and 5-year survival rates were 35.5%, 20.5%, and 16.5% in the PCC + HL group and 27.2%, 8.8%, and 7.8% in the PCC - HL group (P > 0.05). In conclusion, the presence of hepatolithiasis hindered an exact diagnosis of underlying cholangiocarcinoma preoperatively, precipitated biliary sepsis which affected resectability, and increased postoperative morbidity. Hepatolithiasis per se, however, did not influence the long-term survival. PMID- 10711445 TI - Mutations of p53 tumor suppressor gene, apoptosis, and proliferation in intrahepatic cholangiocellular carcinoma of the liver. AB - This study was performed to examine the correlation between mutations of the p53 tumor suppressor gene, the occurrence of apoptosis, and proliferation in cholangiocellular carcinoma of the liver. The results obtained were compared with pathohistological stage (according to UICC) and grade and with disease related survival rate. In 41 curatively (R0-) resected intrahepatic cholangiocellular carcinomas, the status of the p53 gene was determined by direct sequencing of exons 4-9 and immunohistochemically. Apoptosis was assessed using the in situ end labeling (ISEL) technique in combination with morphological criteria. Proliferation was analyzed by immunohistochemistry of MIB-1 (Ki-67), Proliferating cell nuclear antigen (PCNA), and silver-stained nucleolar organizer regions (AgNOR). The results obtained were compared with pathohistological stage (according to UICC), grade, several other histopathological factors, and survival rate. Mutations of p53 were detected in 15/41 carcinomas examined (37%). The most common change was a G-->C and C-->T transition, changing the hot spot amino acid determined by exons 4-8. Of these 15 tumors, 14 were also p53-positive by immunohistochemistry. In each carcinoma examined, we could demonstrate MIB-1, PCNA, and AgNOR dots and also apoptotic cells in variable proportions. The proliferation markers showed a significant correlation among themselves. In univariate survival analysis, the extent of the primary tumor, lymph node status, grade, and p53 were significant factors influencing patient survival. Performing multivariate Cox regression survival analysis, however, only the extent of primary tumor and lymph node status had an independent prognostic impact. Apoptosis was not related to patient prognosis or to other parameters examined. In conclusion, these results indicated that p53 could serve as an additional prognostic parameter that could provide auxiliary information for patient outcome. However, tumor stage and lymph node involvement were the strongest prognostic factors. We failed to establish apoptosis or other pathological parameters as factors predicting the prognosis of patients with cholangiocellular carcinoma. PMID- 10711446 TI - Loss of inhibitory growth regulation by TGF-beta1 in preneoplastic lesions in rat liver. AB - Injection of pig serum into rats twice a week for eight weeks induced transforming growth factor-beta1 (TGF-beta1) mRNA expression and protein production resulting in liver fibrosis without parenchymal cell injury. Eight week treatment with pig serum reduced bromodeoxyuridine (BrdU) -positive hepatocytes 24 hr after 70% partial hepatectomy compared to that in the livers of rats treated with saline for eight weeks. Administration of a choline-deficient L amino acid-defined (CDAA) diet for six weeks with pig serum coadministration, after pretreatment with pig serum for eight weeks, led to the development of preneoplastic lesions that were positive for the placental form of glutathione S transferase (GSTP). Eight-week pretreatment with pig serum induced more GSTP positive lesions and TGF-beta1 mRNA expression and protein concentration in the livers of rats subsequently fed a CDAA diet for six weeks than in rats fed the CDAA diet with saline treatment. These results indicate that TGF-beta1 induced by pig serum treatment inhibited hepatocyte proliferation but failed to prevent the development of preneoplastic lesions in a CDAA diet model. PMID- 10711447 TI - Prognostic value of abdominal CT scanning and hepatic histopathology in patients with acute liver failure. AB - Acute liver failure has extremely high mortality without liver transplantation. We attempted to determine the value of abdominal CT scanning and liver biopsy in its management. A retrospective analysis of patients with acute liver failure was performed; demographic, clinical, radiologic and histopathologic features were noted. Over a period of 13 years, 177 patients were evaluated. The mean age was 39 years and 63% were females. The patients were divided into three groups. Fourteen percent survived with medical management (group I), 37% died (group II), and 49% had liver transplantation (group III). Most patients showed diffuse low density of the liver on CT scanning and the proportions were similar in the three groups. Moderate to large ascites was not present in group I but occurred in 31% of patients in group II and in 15% in group III. Mean hepatic volumes were similar in the three groups; however, 97% of the patients with a liver volume of less than 1000 ml either died or required liver transplantation. Liver biopsies among patients with spontaneous recovery (group I) were distinguished by the presence of regenerative changes and a hepatic parenchymal necrosis of less than 50%. These results suggest that in patients with acute liver failure a liver volume of less than 1000 ml and/or hepatic parenchymal necrosis of greater than 50% is indicative of a poor prognosis. This information may assist decision making in such patients, in particular, regarding the need for liver transplantation. PMID- 10711448 TI - Heart rate variability as a predictor of autonomic dysfunction in patients awaiting liver transplantation. AB - Chronic liver disease, both alcoholic and nonalcoholic, has been shown to be associated with autonomic neuropathy, as well as other hemodynamic and circulatory disturbances. In a longitudinal study, the presence of autonomic neuropathy and the severity of liver disease were independent risk factors for mortality. The aim of this study was to determine whether the severity of liver disease correlated with measures of heart rate variability. We studied 21 patients being evaluated for liver transplantation to determine if severity of disease correlated with heart rate variability and compared them to seven healthy controls. Heart rate variability was determined for a series of 500 consecutive R R intervals during quiet breathing. Standard deviation, pNN50, a marker of parasympathetic function, and approximate entropy (ApEn), a recently described measure of regularity, were calculated. Four standard tests of autonomic function were also performed. pNN50 was significantly reduced in all liver disease patients compared to controls (P < 0.05). Both standard deviation and ApEn were significantly reduced in Child's class C patients suggesting a generalized dysfunction in cardiovascular homeostasis. ApEn was significantly lower in the nonsurvivors during follow-up than the survivors (P < 0.05). In conclusion, increasing severity of liver failure is associated with a reduction in total heart rate variability and regularity. Measurement of heart rate variability offers a simple, noninvasive means of assessing the cardiovascular and autonomic effects of liver disease, particularly in those awaiting liver transplantation. PMID- 10711449 TI - Health profile preferences of hepatitis C patients. AB - The side effects of interferon-alpha for chronic hepatitis C are well-known. Patients may differ with respect to their tolerance of these side effects and also with respect to their individual preferences. We administered a brief questionnaire to 67 outpatients with hepatitis C virus infection. Patients were asked to make hypothetical choices between six-month profiles of health. The results were as follows: (1) patients preferred to expedite rather than postpone intervals of poor health; (2) preferences of patients with low quality-of-life were quite similar to preferences of healthier patients; (3) patients' choices satisfied transitivity; (4) patients' choices satisfied preferential independence; and (5) patients gave a variety of reasons for their choices. These results corroborate other investigations of health preferences, and serve to introduce the field of preference elicitation to gastroenterologists. PMID- 10711450 TI - Hepatitis C virus infection in institutionalized psychiatric patients: possible role of transmission by razor sharing. AB - The objective of this study was to determine if HCV can be transmitted from patient to patient in psychiatric institutions and to determine possible routes of infection. We did a cross-sectional survey of 196 Japanese psychiatric patients tested for HCV and HBV markers and 400 age- and sex-matched controls. Anti-HCV was detected in 10.2% and antibody to hepatitis B core antigen was detected in 44.4% of the patients, a significantly higher prevalence than found among matched controls. A multiple regression logistic analysis was used to identify risk factors that could indicate the route of infection by HCV. Duration of hospitalization, age, razor sharing, and history of surgery proved to be statistically significant independent risk factors associated with positive anti HCV results [odds ratio (OR), 4.00; 95% confidence interval (CI), CI, 1.74-9.19; OR, 2.19; 95% CI, 1.27-1.3.77; OR, 4.90; 95% CI, 1.29-18.86; OR, 3.35; 95% CI, 0.997-11.3, respectively]. These observations suggest that razor sharing played an important role in the spread of the HCV infection in the institutionalized psychiatric patients we studied. PMID- 10711451 TI - Delayed hemoperitoneum following large-volume paracentesis in a patient with cirrhosis and ascites. AB - The diagnosis of early or late hemoperitoneum after large-volume paracentesis can be reached easily by a repeat tap, but gastrointestinal bleeding and other common causes of hypotension in cirrhotics must be ruled out first. When the hemoperitoneum is confirmed, imaging studies are often inconclusive and laparotomy should be considered when hemodynamic instability persists despite adequate fluid resuscitation. However, in instances of delayed hemoperitoneum, it must be anticipated that operation may not identify the bleeding site and result in further decompensation of the liver. OLT may well be the best therapeutic option in this rare, high-risk situation. PMID- 10711452 TI - Interpretation of simultaneous measurements of hepatic extraction fractions of indocyanine green and sorbitol: evidence of hepatic shunts and capillarization? AB - Sorbitol and indocyanine green (ICG) have high hepatic extraction fractions (E(sorb) and E(ICG)) in normal subjects. A curved relationship has been observed between E(sorb) and E(ICG) in liver disease. According to one interpretation, the decrease of E(sorb) is a result of intrahepatic shunting and 1 - E(sorb) is the fraction of shunted flow (the shunt hypothesis). Under the further assumption that capillarization of functioning sinusoids prevents hepatic uptake of plasma protein-bound ICG and allows uptake of water-soluble sorbitol, the difference E(sorb) - E(ICG) has been suggested as a measure of capillarization. We propose an alternative hypothesis: that the sinusoidal permeability-surface area products for sorbitol and ICG are reduced in proportion by liver disease (proportional reduction hypothesis). Based on the sinusoidal perfusion model, predictions were produced from both hypotheses for the relation between E(sorb) and E(ICG) and the additional effects of capillarization were described. By use of liver vein catheterization, E(sorb) and E(ICG) were simultaneously measured during continuous infusions in 53 human subjects with varying degrees of liver disease. The data were in better agreement with the predictions of the proportional reduction hypothesis than with the shunt hypothesis. Even though both intrahepatic portosystemic shunts and sinusoidal capillarization are known to occur in cirrhosis and also may have influenced our data, they appeared to be of minor importance from a kinetic point of view. These findings favor the proportional reduction hypothesis and do not support the use of systemic nonrenal clearance of sorbitol as a measure of "functional liver blood flow." PMID- 10711453 TI - Immunological similarities between primary sclerosing cholangitis and chronic sclerosing sialadenitis: report of the overlapping of these two autoimmune diseases. AB - Primary sclerosing cholangitis (PSC) is characterized by destructive inflammation and fibrosis affecting the bile ducts. The etiology of PSC is still unknown, although lymphocytic infiltration in the portal areas suggests an immune-mediated destruction of the bile ducts. Patients with one autoimmune disease often suffer from one or more other autoimmune diseases. It is well known that there is a close relationship between PSC and inflammatory bowel disease, particularly ulcerative colitis(UC). However, the pathological findings in UC and other overlap diseases do not resemble those of PSC. In the present study, we report a patient with chronic sclerosing sialadenitis (Kuttner's tumor) and PSC. It is compared the sclerosing changes in both salivary glands and bile ducts histologically. In addition, the expression pattern of mast cell tryptase, b-FGF, and HLA-DR were examined in both tissues immunohistochemically. Histological features of sclerosing change in both salivary and bile ducts were quite similar. Marked mast cell infiltration and b-FGF expression were seen in the sclerosing areas in both tissues. In active inflammatory areas of the salivary glands, HLA DR expression was also seen. We hypothesized that similar immune reactions occur in both the salivary gland and bile ducts and are responsible for the fibrosis that follows. PMID- 10711454 TI - Bleeding jejunal varices and portal thrombosis in a splenectomized patient with hereditary spherocytosis. PMID- 10711455 TI - Microsatellite instability is uncommon in intestinal mucosa of patients with Crohn's disease. AB - Patients with long-standing inflammatory bowel disease have an increased risk for colorectal carcinoma. Microsatellite instability occurs in colonic neoplasms and has been reported in colonic tissues from patients with ulcerative colitis. Patients with Crohn's disease also have an increased risk for colorectal cancer, although it is lower than that associated with ulcerative colitis. This study was designed to determine whether microsatellite instability occurs in Crohn's disease, and whether it occurs with similar frequency to that observed in ulcerative colitis. In all, 177 tissue samples from 33 patients with Crohn's disease were evaluated for microsatellite alterations. Microsatellite instability occurred in five different tissue samples from one of 33 Crohn's disease patients. Four of the five tissue samples showed microsatellite instability at more than one locus. We conclude that microsatellite instability is less common in Crohn's disease than ulcerative colitis and may reflect differences in cancer risk between these two forms of inflammatory bowel disease. PMID- 10711457 TI - Iron supplementation may aggravate inflammatory status of colitis in a rat model. AB - Iron supplementation is one of the principal therapies in inflammatory bowel disease. Iron is a major prooxidative agent; therefore therapeutic iron as well as heme iron from chronic mucosal bleeding can increase the iron-mediated oxidative stress in colitis by facilitating the Fenton reaction, namely production of hydroxyl radicals. In the present study colitis was induced in the iodoacetamide rat model. Forty male Whistar rats were divided into four groups, each group receiving a different diet regimen in parallel with colitis induction: Malondialdehyde was measured to assess the degree of tissue oxidative stress. There were microscopic changes, and significantly more severe colitis was seen in colonic biopsies when iron was supplemented. It was concluded that iron supplementation can amplify the inflammatory response and enhance the subsequent mucosal damage in a rat model of colitis. We suggest that the resultant oxidative stress generated by iron supplementation leads to the extension and propagation of crypt abscesses. PMID- 10711456 TI - Evidence that colitis is initiated by environmental stress and sustained by fecal factors in the cotton-top tamarin (Saguinus oedipus). AB - Parallel changes in spontaneously occurring inflammation in colonic Thiry-Vella loops and the in-line colon of cotton-top tamarins were studied in a colitis inducing environment at 8 and 15 months following surgical preparation of the loops. Gross disease severity and numbers of inflammatory/immune cells per unit area of lamina propria in histological sections from endoscopic biopsies were analyzed. Cell counts and severity of colitis declined over time in the Thiry Villa loops while the disease followed its characteristic course in the remaining large bowel and in the colons of controls. Perfusion of the loops with the animals' feces increased the density of the cellular infiltrate in the lamina propria in parallel with increased severity of inflammation. Electron micrographs of the colonic mucosa showed invasion by microorganisms. The predominant microorganism had characteristics of Helicobacter sp. The results implicate the fecal stream as a factor in the persistence of colitis in the tamarin model. Nevertheless, fecal factors appear not to be the primary trigger, as evidenced by findings that the disease is not expressed in wild-living tamarins and that it enters remission when affected animals are transferred to natural conditions from a colitis-inducing environment. Both an adverse environment and the fecal contents appear to be required for expression of the disease. PMID- 10711458 TI - Increased levels of circulating ICAM-1, E-selectin, and IL-2 receptors in celiac disease. AB - Adhesive interactions between endothelium and circulating cells are crucial for the development of inflammatory reactions. We found significantly higher serum levels of soluble intracellular adhesion molecule-1 (sICAM-1, 492.5 +/- 22.1 ng/ml) in patients with active celiac disease (including IgA-deficient patients) than in patients on a gluten-free diet (335.7 +/- 20.0 ng/ml) (P < 0.001) and healthy controls (207.4 +/- 11.2 ng/ml) (P < 0.001). The concentration of soluble E-selectin in sera from celiac patients (37.2 +/- 3.4 ng/ml) was also higher (P < 0.001) than in sera from healthy controls (15.5 +/- 0.7 ng/ml) but, in contrast to sICAM-1, it remained high in the patients after treatment (30.2 +/- 2.7 ng/ml). Interestingly, the concentration of circulating soluble interleukin-2 receptors, molecules indicating lymphocyte activation, was only increased in sera from patients with active celiac disease (2943.0 +/- 214.1 pg/ml), and the level in sera from treated patients and healthy controls was comparable (1936 +/- 349 and 1416 +/- 111.7 pg/ml). The elevated serum level of soluble cell adhesion molecules could be used as a supplementary, noninvasive procedure for monitoring intestinal immune reactions. PMID- 10711459 TI - Usefulness of screening program for celiac disease in autoimmune thyroiditis. AB - We determined the prevalence of celiac disease in subjects with autoimmune thyroiditis compared with sick and healthy subjects. The screening was performed with IgA-class endomysium antibody, by indirect immunofluorescence using human umbilical cord as the antigenic substrate. Six of the 172 patients with autoimmune thyroiditis were found to be anti-endomysium positive (3.4%) and five of these underwent intestinal biopsy, which showed total villous atrophy. By contrast, 3 (0.75%) of 396 patients with nongastroenterologic malignancies and 10 (0.25%) of 4000 blood donors were found to have celiac disease. The prevalence of autoimmune diseases was significantly higher in patients with both celiac disease and autoimmune thyroiditis than in patients with autoimmune thyroiditis alone (P = 0.01). This study confirms that celiac disease is increased among patients with autoimmune thyroiditis. We suggest that these patients may benefit from screening for celiac disease so as to eliminate symptoms and limit the risk of developing other autoimmune disorders. PMID- 10711460 TI - Serum unconjugated bile acids as a test for intestinal bacterial overgrowth in dogs. AB - Small intestinal bacterial overgrowth (SIBO) has a high incidence in dogs and, as in humans, is difficult to diagnose. The aim of this study was to determine the diagnostic significance of serum unconjugated bile acid concentrations in dogs with bacterial overgrowth. Fasting sera were obtained from 23 dogs: 10 with culture-proven SIBO, 8 with indirectly diagnosed SIBO (normal pancreatic function but small intestinal disease associated with subnormal serum cobalamin and supranormal folate concentrations), and 5 healthy controls. Unconjugated bile acids were determined using gas chromatography-mass spectrometry after isolation by liquid-solid extraction and anion-exchange chromatography. Mean serum unconjugated bile acid concentrations were significantly elevated in dogs with SIBO (mean +/- SD: 0.91 +/- 1.03 micromol/liter), and in dogs with indirectly diagnosed SIBO (2.11 +/- 2.20 micromol/liter) compared to clinically healthy dogs (0.015 +/- 0.015 micromol/liter, P < 0.005). Cholic acid was the predominant unconjugated bile acid in the serum of dogs with SIBO. In conclusion serum unconjugated bile acid concentrations of healthy dogs are significantly lower than reported values for humans, and this fraction represents a relatively small proportion (0-2.3%; mean 0.8%) of the total bile acids in dogs. Unconjugated bile acids increased 10- to 20-fold in dogs with SIBO indicating the clinical utility of serum unconjugated bile acids for diagnosis of intestinal bacterial overgrowth in dogs. PMID- 10711461 TI - Burn and starvation increase programmed cell death in small bowel epithelial cells. AB - Maintenance of gut mucosal homeostasis depends on a balance between cell proliferation and cell death. Gut mucosal integrity is impaired after severe burn and during starvation. We determined the effect of burn, starvation, and the combination of both on small bowel epithelial apoptosis and proliferation. Fifty adult male Fischer 344 rats (260-300 g) received a 60% full-thickness scald burn and were randomly divided into fed and starved groups. Small intestine was taken at 12, 24, and 48 hr after injury. All animals in the 12-hr group were starved while recovering from anesthesia. Apoptosis was quantified by immunohistochemical staining (TUNEL) and mucosal proliferation was determined by bromodeoxyuridine (BrdU) incorporation. The apoptotic index was higher in burned rats compared to controls at 12 hr after burn; both these groups were starved (P < 0.05). At 24 and 48 hr after burn, apoptosis was highest in the starved groups, with no additional effects of burn (P < 0.05). Mucosal epithelial cell proliferation was not different between groups at any time point. In conclusion, burn and starvation both increase apoptosis in the small bowel mucosa; however, these effects are not additive. Apoptosis could be attenuated by enteral feeding, which delineates the importance of early enteral feeding initiation after injury to maintain mucosal integrity. PMID- 10711464 TI - Paraganglioma of the pancreas: literature review and case report. AB - Extraadrenal paragangliomas are very rare tumors arising from cells derived from the neural crest. These tumors are encountered only as case reports, and as a result, little is known of their natural history. We present a case of pancreatic paraganglioma and review all previously reported cases. PMID- 10711463 TI - Ascites of severe acute pancreatitis in rats transcriptionally up-regulates expression of interleukin-6 and -8 in vascular endothelium and mononuclear leukocytes. AB - The molecular mechanisms that link acute pancreatitis and multiple organ failure remain unknown. We examined the effect of ascitic fluids prepared from rats with experimental necrotizing pancreatitis on the expression of interleukin (IL) -6 and IL-8 in human umbilical vein endothelial cells (HUVEC) and human monocytic THP-1 cells. Incubation of HUVEC or THP-1 cells with the ascitic fluids resulted in a concentration-dependent up-regulation of the cytokine expression with comparable mRNA induction. Electrophoretic mobility shift assay revealed that the ascitic fluids increased the nuclear factor-kappaB (NF-kappaB) and NF-IL6 binding activities. Intraperitoneal injection of ascitic fluids into healthy rats induced the activation of NF-kappaB in the infiltrating leukocytes in the lung. Our results suggested that ascitic fluids may play a role in the pathophysiology of severe acute pancreatitis through the activation of transcription factors and consequent cytokine productions in distant organs. PMID- 10711462 TI - Gallbladder motility and lithogenesis in obese patients during diet-induced weight loss. AB - Obesity and weight loss are important risk factors for gallstone development. The mechanisms involved are unknown. We prospectively studied changes in gallbladder (GB) emptying and bile composition during weight loss. We studied 12 alithiasic obese subjects who entered a six-month diet program (800-1200 kcal/day, 26 g fat/day). As controls we evaluated 12 healthy nulliparous nonobese young women. GB volumes were studied by ultrasonography (fasting volume, GBFV; residual volume after a liquid meal, GBRV) at entry and after 4 and 20 weeks of dieting. Bile acid pool size, biliary lipid composition, presence of cholesterol crystals, and nucleation time were also studied. Of 12 obese subjects studied (mean BMI 35.1 kg/m2), 10 remained in the program for six months, but only six completed the entire study protocol, obtaining a significant weight loss (BMI: 31.2 kg/m2, P < 0.001). GBFV was greater in obese subjects than in nonobese controls (27.5 +/- 10.7 vs 11.7 +/- 6 ml; P < 0.05). GBRV and GB emptying curves were similar in both groups and did not change during weight loss. The obese subject who developed gallstones (1/10) was the only one who had cholesterol crystals in bile and a sluggish initial GB emptying. IN CONCLUSION: (1) obese subjects had a greater GBFV than controls; however, the GB emptying was adequate. (2) During weight loss we did not observe significant changes in GB kinetics or the bile parameters studied. (3) We observed a relatively low frequency of gallstone formation, which can be explained by a high fat content of the diet (26 g/day) and by the adequate GB emptying of our group of patients. (4) An abnormal GB contractility and cholesterol crystals in bile could be considered premonitory to gallstone formation. PMID- 10711466 TI - Evidence-based surgery: a progenitor in the literature of cholecystectomy. PMID- 10711465 TI - Current status of sentinel node biopsy in breast cancer. PMID- 10711467 TI - The leaking anterior resection and the management of SIRS, MODS and CHAOS. PMID- 10711468 TI - Inflammatory conditions of the common bile duct. AB - BACKGROUND: Inflammatory conditions of the biliary tree are infrequently reported with the exception of sclerosing cholangitis. These conditions, especially when affecting the common bile duct (CBD), can be mistaken for tumours; their existence should be considered in the differential diagnosis of biliary lesions and, in particular, biliary strictures. Few descriptions of inflammatory conditions exist in the published literature. METHODS: A comprehensive search was undertaken of the last 20 years of published literature. RESULTS: Potential aetiological factors include infection, infestation, traumatic bile duct injury, foreign body or gallstone reaction and the host inflammatory response. CONCLUSIONS: These conditions typically mimic malignant tumours of the CBD but knowledge of the aforementioned processes might alter the surgical approach to biliary strictures (especially after previous cholecystectomy), with greater emphasis on surgical exploration. PMID- 10711469 TI - Sentinel node biopsy in breast cancer: results of 103 cases. AB - BACKGROUND: In early breast cancer the status of the axillary nodes has been shown to be one of the primary prognostic indicators. Biopsy of the sentinel node, or first draining lymph node, of a tumour has been investigated as an alternative to axillary dissection in early breast cancer. A series of sentinel node biopsies in 103 patients is reported here. METHODS: Both pre-operative lymphoscintigraphy and intra-operative blue dye were used to map the sentinel nodes. RESULTS: Mapping was successful in 87 (84.4%) cases and sentinel nodes were retrieved in 94.2% of these patients. Where lymphoscintigraphic mapping was unsuccessful, sentinel nodes were found in 37.5%. When sentinel nodes were retrieved, correlation of the sentinel node status with the axillary nodes was accurate in 97.5%. There were two false negatives, both in large tumours. The sentinel node status was an accurate predictor of axillary status in 95.7% of the node positive patients. CONCLUSIONS: If only the 86 patients with invasive carcinoma and four or more axillary nodes removed at surgery are considered, the sentinel node was accurate in assessing the axillary status in 97.7% of the total patient group (2.3% false negative rate), 97.2% of those in whom sentinel nodes were successfully retrieved (2.8% false negative rate) and 94.9% of the patients with positive axillary nodes (5.1% false negative rate). Sentinel node biopsy is a valid technique providing an accurate reflection of the axillary node status and having a low false negative rate. PMID- 10711470 TI - The value of cytology in granulomatous mastitis: a report of 16 cases from Malaysia. AB - BACKGROUND: Granulomatous mastitis is a rare condition of the breast that can mimic a carcinoma. There are characteristic histological features, the most important of which is a predominantly lobular inflammatory process. It must be differentiated from known causes of granulomatous inflammation, such as tuberculosis. METHODS: In the present paper, the clinical and pathological features of 16 patients with granulomatous mastitis seen over a 3-year period in the University Hospital, Kuala Lumpur, are described. RESULTS: A clinical suspicion of malignancy was present in 10 cases. One of the patients was nulliparous. One had an associated hyperprolactinaemia, while two had systemic lupus erythromatosis. One of the patients was pregnant at the time of presentation. Four patients had localized lumps excised, five were treated conservatively because the lesion was too extensive to resect, and seven patients required drainage procedures for abscess formation. CONCLUSION: Awareness of this condition is important because it mimics a carcinoma, and surgery may not be the best treatment for recurrent disease. PMID- 10711471 TI - Role of selective intra-operative cholangiography during cholecystectomy. AB - BACKGROUND: The use of routine intra-operative cholangiography (IOC) remains controversial. This prospective study was carried out to determine whether to perform selective or routine IOC in patients undergoing cholecystectomy for gallstones. METHODS: All consecutive patients undergoing open cholecystectomy over a 16-month period were included in the present study. They were divided into two groups based on the absence (n = 79) or presence (n = 55) of indicators of choledocholithiasis. All patients were subjected to cholangiography. Each indicator, subsets of indicators and all indicators combined were evaluated for their ability to predict choledocholithiasis. RESULTS: There would be only two missed stones (1.5%) if selective cholangiography was to be practised. Intra operative cholangiography had a positive predictive value of 100%. The sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) and accuracy of all the indicators combined were 93.5%, 84.6%, 74.5%, 97.5% and 88.0%, respectively. The best indicators in each subset were jaundice, common bile duct diameter as assessed by ultrasonography, and a palpable stone during surgery with NPV of 82.7%, 91.1% and 96.8%, respectively. CONCLUSION: Routine IOC during cholecystectomy is not essential for the prevention of retained stones. A combination of the various indicators of choledocholithiasis can be used to select patients for cholangiography. PMID- 10711472 TI - Intermittent pump versus compression bandages in the treatment of venous leg ulcers. AB - BACKGROUND: The purpose of the present paper was to compare healing rate and leg swelling with an intermittent compression pump versus compression bandages in the treatment of venous leg ulcers, and to also compare patient compliance and satisfaction with the two techniques. METHODS: A randomized cross-over study of patients attending an outpatient wound clinic (n = 16) was undertaken. A regular monthly follow-up with measurement of ulcer size and leg volume was carried out, as well as completion of a questionnaire. RESULTS: Assessment was possible in 11 of the 16 patients. There was no significant difference between treatment types with regards to ulcer healing rates or control of leg oedema. The survey revealed that patients found the pump easier and more comfortable to use, with a trend towards increased compliance. CONCLUSIONS: Although the present study was too small for generalizable conclusions, compression pumps and bandages are comparable in efficacy for the healing of venous leg ulcers. The compression pump is reported as being easier and more comfortable to use than bandages. PMID- 10711473 TI - Australasian survey of split skin graft donor site dressings. AB - BACKGROUND: There is an ever increasing array of products available for wound dressings. The aim of the present study was to establish which dressings should be used as standard controls for future studies; what factors are regarded as most important in assessing a dressing; what the level of satisfaction is with the available products; what the strengths and weaknesses of the commonly used dressings are; and what dressings would be preferred if cost were no issue. METHODS: A postal survey was sent to every plastic and reconstructive surgeon registered in Australasia (n = 217). A total of 53% responded. RESULTS: The most commonly used dressing type overall is the calcium alginates, despite the fact that they were not the highest performing dressings. This is also the most commonly used in Australia. In contrast scarlet red is still used most commonly in New Zealand. The level of satisfaction with the most commonly used dressing varied very little. The factor regarded most important was patient comfort level. A profile of the commonly used dressing was constructed. Calcium alginates and or scarlet red should be used as the control for new product comparisons. CONCLUSIONS: Most of the respondents were satisfied with their preferred dressing and were not interested in trying alternative dressings. PMID- 10711474 TI - Hip fracture rates in South Australia: into the next century. AB - BACKGROUND: Fractures of the femoral neck already represent a major public health problem in Australia. This situation is set to worsen as the population ages. The present study estimates the number of patients over 50 years of age with femoral neck fractures that is expected to impact on the South Australian healthcare service into the next century. METHODS: Population projections from the Australian Bureau of Statistics 1996 census were combined with age- and gender specific incidence rates for fractures of the femoral neck for persons over the age of 50 in South Australia. Projections for the expected number of hip fractures in this State were then calculated. RESULTS: Assuming there are no changes in the age- and gender-specific incidence of fracture rates, the number of fractures in South Australia is estimated to increase by approximately 66% by the year 2021 and 190% by 2051. CONCLUSION: Based on the population projections and the assumption that conditions contributing to hip fractures remain constant, the number of fractured neck of femurs will increase in far greater proportion than the overall population in the next century. The results of the present study indicate the serious implications for the South Australian healthcare system if there is no reduction in incidence rates. PMID- 10711475 TI - Mandibular reconstruction following ablative tumour surgery: an overview of treatment planning. AB - BACKGROUND: In the last half century the evolution of mandibular reconstruction has resulted in a multitude of surgical procedures that have been brought about by advances made in bone science, grafting techniques and materials technology. With the ever increasing number of surgical options currently available it has become essential to take stock of these achievements and reconsider the fundamental principles of reconstructive surgery. METHODS: The choice of reconstructive techniques is only part of a wider number of variables that must be considered which will ultimately determine the success or failure of the procedure. RESULTS: Variables likely to influence the outcome of mandibular reconstruction are the site and extent of the defect, the needs and tolerance of the patient, the timing of reconstruction and the surgical skill and techniques available. CONCLUSION: By careful consideration of these cardinal variables it may be possible to formulate a treatment plan that will not only satisfy the essential needs of the patient but also help increase the likelihood of a successful outcome. PMID- 10711476 TI - A biodialysis system for liver support tested in a porcine hepatic failure model. AB - BACKGROUND: A practical liver support system for patients in fulminant hepatic failure (FHF) remains a needed therapeutic modality. A new method of bioartificial liver support, the liver biodialysis system (LBDS), is described. METHODS: Porcine hepatocytes, removed from direct contact with the treated subject's circulation, are in culture in a bioreactor which is combined in a dialysis circuit for patient treatment. The LBDS was tested in a porcine ischaemic hepatic failure model. RESULTS: The viable hepatocyte content of the bioreactor was 2.49 +/- 0.72 x 10(10). Cells remained viable in culture throughout the experiments (30 +/- 3 h) without evidence of immunological damage. A decrease in the degree of accumulation in the blood of ammonia (P < 0.02) and of 14 amino acids (P < 0.001) was achieved by the LBDS. Cerebral perfusion pressure was maintained at significantly higher levels in LBDS-treated animals (P < 0.05). CONCLUSIONS: In the LBDS, hepatocytes in large numbers and satisfactory culture conditions in a bioreactor have sustained viability and function. When combined in a dialysis circuit for the treatment of FHF pigs, immune reactions between the blood and hepatocytes were prevented and beneficial metabolic effects were observed. PMID- 10711477 TI - Sentinel node biopsy in breast cancer: recommendations for surgeons, pathologists, nuclear physicians and radiologists in Australia and New Zealand. AB - BACKGROUND: Assessment of axillary lymph node status is necessary for patients with invasive breast cancer. Sentinel node biopsy is a new minimally invasive technique that may provide accurate assessment of regional lymph node status while limiting the morbidity associated with axillary clearance. METHODS: A workshop conducted in Adelaide in November 1998 aimed to assess current sentinel node mapping and biopsy techniques, and make recommendations regarding its application in the surgical management of early breast cancer in Australia and New Zealand. RESULTS: At the conclusion of the workshop, a consensus was reached regarding indications, exclusions, sentinel node mapping/biopsy technique, nuclear medicine requirements, pathology and safety of sentinel node biopsy in breast cancer. It was agreed that a feasibility study according to an agreed prospective protocol was necessary to validate the technique by breast surgeons. Surgeons that satisfied validation criteria for the feasibility study could then consider a prospective randomized study comparing sentinel node biopsy with standard axillary dissection. CONCLUSIONS: Sentinel node biopsy in breast cancer involves close cooperation between members of a multidisciplinary team including surgeons, nuclear physicians, pathologists and radiologists. Although the technique has the potential to reduce morbidity associated with axillary surgery, surgical performance in this area will need to be closely monitored to ensure that the technique does not fall into disrepute by adversely affecting breast cancer prognosis. PMID- 10711478 TI - Sir Edward Hughes and colorectal surgery. PMID- 10711479 TI - A cerebrospinal fluid collection presenting as an abdominal mass following lumbar vertebrectomy for trauma. PMID- 10711480 TI - Primary pancreatic tuberculosis: presentation and diagnosis. PMID- 10711481 TI - Secretory breast carcinoma in a 9-year-old boy. PMID- 10711482 TI - Clostridium difficile splenic abscess. PMID- 10711483 TI - Vaccinia necrosum: a forgotten disease. PMID- 10711484 TI - Simply add oxygen: why isn't oxygen administration taught in all resuscitation training? AB - Resuscitation training is all about procedures, yet the overwhelming need is to get sufficient oxygen to hypoxic and endangered tissues. Neither the Australian Resuscitation Council nor the American Heart Association make mention in any basic courses of the need to add oxygen as soon as possible. The Australian Resuscitation Council has a separate policy on oxygen, and the American Heart Association mentions the need, but not until page 2274 of the 1992 'Guidelines'. Major first aid training organisations do not mention the use of oxygen resuscitation in basic courses. There seems to be a belief in first aid training that oxygen administration is potentially dangerous and is an 'Advanced' skill. Oxygen should be used as soon as possible, in as near 100% as possible in all resuscitation situations, and for the early management of injury and illness. Its use will never disadvantage a patient under these circumstances. This skill should be added into all resuscitation training. In these days of high technology for all, we can easily add the simple skill of administration of oxygen, as soon as available, to basic life support and first aid. An oxygen supply should be as easily available as a fire extinguisher, and as simply used. PMID- 10711485 TI - Witnessed resuscitation by relatives. AB - Witnessed resuscitation is the process of active 'medical' resuscitation in the presence of family members. Witnessed resuscitation though not as yet wide spread in practice is becoming established. Early reports of programs designed to promote such a process first appeared in the early 1980s. More recent work appears to show both public support and a desire for inclusion in the resuscitation process. Some research has been produced that indicates both satisfaction and psychological benefit for those relatives enabled to witness. Limited work only, exists pertaining to the effects on health care providers and these reports currently do not show any significant deleterious effects. Approval of witnessed resuscitation programs is not universal amongst all groups of health care workers. Concerns about the ethics of witnessed resuscitation and its medico legal implications have been raised. The quality of the initial witnessed resuscitation reports is however variable and there is a great need for further work to validate the initial findings particularly in the areas of psychological stressors in staff and risk management implications. PMID- 10711486 TI - Out-of-hospital cardiac arrest in north-east Germany: increased resuscitation efforts and improved survival. AB - In the years after 1989 major political and socioeconomic changes have taken place in East Germany. In parallel, emergency medical services (EMS) were restructured according to western standards. In Stralsund the EMS was restructured from a single to a two tier system with implementation of a second ambulance base in 1990. The number of household telephone extensions more than doubled. To analyze the effects of these changes, patients receiving advanced life support (ALS) for out-of-hospital cardiac arrest of cardiac origin (OHCA) between 1984 and 1988, and from 1991 to 1997 were studied. Adjusted per 100,000 inhabitants, the number of OHCA patients receiving ALS increased from 11 per year before 1989 to 52 per year after 1990 (P < 0.01). Survival without relevant neurologic defects was achieved in 3.7% (2/53) of patients before 1989 and in 8.1% (22/273) after 1990. Response time of the ALS unit shortened from 11.0 +/- 1.4 to 9.0 +/- 0.4 min (n.s.), while response time of any EMS shortened from 11.0 +/- 1.4 to 6.1 +/- 0.3 min (P < 0.005). Adjusted for observation period and population served, there was a 10-fold increase in the number of resuscitations attempted at home and an 8-fold increase in the absolute number of OHCA survivors without relevant neurological defects. In parallel to socioeconomic changes, the restructuring of the EMS in Stralsund and the rapid expansion of the telephone network led to a significant increase in the number of patients successfully resuscitated from OHCA. If the present results can be transferred to other former socialist countries of East and Middle Europe, they may have important implications for the EMS in these regions. PMID- 10711487 TI - Abdominal haemorrhage--a preventable cause of death after field stabilization? AB - The causes of preventable death vary in different operational settings, and the topic has not previously been explored in a fully developed central European rescue system. The factors associated with potentially preventable death were studied in a retrospective study of 430 fatal traffic accident victims (1980-96) in Lorrach County, Germany. Mission protocols could be retrieved for detailed analysis in 239 of the cases. These were studied in order to identify factors associated with preventable death. At the scene of the accident, 38% of the patients died without cardiopulmonary resuscitation (CPR) and 18% after CPR. Four patients died after a certain delay without CPR before reaching hospital. A total of 43% of the victims were admitted to hospital, 5% had received prehospital CPR and the remaining 38% had not. In a subgroup representing the experience of a single emergency physician 60 fatalities were studied. Of these, 27 (45%) patients died within the hospital; almost half of these cases (13/27) had been conscious at some time after the accident and of these, seven (7/13) died from intra-abdominal bleeding within 4 h after admission. The same cause of death was found in 3 of the 14 comatose patients. Pleural drainage was carried out in four patients and unrecognized pneumothoraces or spinal injuries did not occur. Tracheal intubation was employed in 24/27. Medical antishock trousers (MAST) were not available. The data indicate that intra-abdominal haemorrhage is an underestimated cause of death in a comprehensive rescue system, possibly as a consequence of field stabilization. The use of MAST may be a relevant therapeutic option to prevent these fatalities. The method offers the possibility of intra abdominal compression and haemostasis after tracheal intubation has been performed. Previous controlled studies on MAST may have been biased by faulty methodology (e.g. absence of tracheal intubation) and inappropriate indications (e.g. other causes of shock). The value of MAST in comprehensive rescue systems should therefore be reassessed. The difficulties in identifying factors leading to preventable death in a retrospective analysis, are discussed and it is recommended that a permanent prospective quality control be performed in all cases of fatal accidents in order to ensure the continued improvement of prehospital emergency medical systems. PMID- 10711488 TI - Optimisation of tidal volumes given with self-inflatable bags without additional oxygen. AB - The European Resuscitation Council has recommended smaller tidal volumes of 500 ml during basic life support ventilation in order to minimise gastric inflation. One method of delivering these tidal volumes may be to use paediatric instead of adult self-inflatable bags; however, we have demonstrated in other studies that only 350 ml may be delivered, using this technique. The reduced risk of gastric inflation was offset by oxygenation problems, rendering the strategy of attempting to deliver tidal volumes of 500 ml with a paediatric self-inflatable bag questionable, at least when using room-air. In this report, we assessed the effects of a self-inflatable bag with a size between the maximum size of a paediatric (700 ml) and an adult (1500 ml) self-inflatable bag on respiratory variables and blood gases during bag-valve-mask ventilation. After induction of anaesthesia, 50 patients were block-randomised into two groups of 25 each. They were ventilated with room-air with either an adult (maximum volume, 1500 ml) or a newly developed medium-size (maximum volume, 1100 ml; Drager, Lubeck, Germany) self-inflatable bag for 5 min before intubation. When compared with the adult self-inflatable bag, the medium-size bag resulted in significantly lower exhaled tidal volumes and tidal volumes per kg bodyweight (624 + 24 versus 738 +/- 20 ml, and 8.5 +/- 0.3 versus 10.7 +/- 0.3 ml kg(-1), respectively; P < 0.001), oxygen saturation (95 +/- 0.4 versus 96 +/- 0.3%; P < 0.05), and partial pressure of oxygen (78 +/- 3 versus 87 +/- 3 mmHg; P < 0.05). Carbon dioxide levels were comparable (37 +/- 1 versus 37 +/- 1 mmHg). Our results indicate that smaller tidal volumes of about 8 ml x kg(-1) (approximately 600 ml), given with a new medium-size self-inflatable bag and room-air, maintained adequate carbon dioxide elimination and oxygenation during bag-valve-mask ventilation. Accordingly, the new medium-size self-inflatable bag may combine both adequate ventilatory support and reduced risk of gastric inflation during bag-valve-mask ventilation. PMID- 10711489 TI - Is hospital care of major importance for outcome after out-of-hospital cardiac arrest? Experience acquired from patients with out-of-hospital cardiac arrest resuscitated by the same Emergency Medical Service and admitted to one of two hospitals over a 16-year period in the municipality of Goteborg. AB - AIM: To describe patient characteristics, hospital investigations and interventions and early mortality among patients being hospitalized after out-of hospital cardiac arrest in two hospitals. SETTING: Municipality of Goteborg, Sweden. PATIENTS: All patients suffering an out-of-hospital cardiac arrest who were successfully resuscitated and admitted to hospital between 1 October 1980 and 31 December 1996. All patients were resuscitated by the same Emergency Medical Service and admitted alive to one of the two city hospitals in Goteborg. RESULTS: Of 579 patients admitted to Sahlgrenska Hospital, 253 (44%) were discharged alive and of 459 patients admitted to Ostra Hospital, 152 (33%) were discharged alive (P < 0.001). More patients in Sahlgrenska Hospital were still receiving cardiopulmonary resuscitation (CPR) treatment (P = 0.03), but patients in Ostra had a lower systolic blood pressure and higher heart rate on admission. A larger percentage of patients admitted to Sahlgrenska Hospital underwent coronary angiography (P < 0.001), electrophysiological testing (P < 0.001), Holter recording (P < 0.001), echocardiography (P = 0.004), percutaneous transluminal coronary angioplasty (PTCA, P = 0.009), implantation of automatic implantable cardioverter defibrillator (AICD, P = 0.03) and exercise stress tests (P = 0.003). Inhabitants in the catchment area of Ostra Hospital had a less favourable socio-economic profile. CONCLUSION: Survival after out-of-hospital cardiac arrest may be affected by the course of hospital management. Other variables that might influence survival are socio-economic factors and cardiorespiratory status on admission to hospital. Further investigation is called for as more patients are being hospitalised alive after out-of-hospital cardiac arrest. PMID- 10711490 TI - Comparison of a two-finger versus two-thumb method for chest compressions by healthcare providers in an infant mechanical model. AB - OBJECTIVE: To compare the two-finger versus the two-thumb method of chest compression on an infant model. METHOD: STUDY: an unblinded, prospective, cross over experimental study. SETTING: the metropolitan area of a city with a population of greater than 260,000. PARTICIPANTS: pediatric medical personnel and emergency workers. Anyone unable to complete the study was excluded. INTERVENTIONS: participants performed chest compressions on an infant mannikin for 2 min. PARTICIPANTS were randomized to use the two-finger method or the two thumb method for the first minute. The investigators recorded the skillguide readings of green (correct), green and orange (too deep), red (wrong placement), or no light (too shallow). Sixty or more correct compressions were judged to be adequate. RESULTS: Two hundred and nine participants completed the study. PARTICIPANTS included: 66 nurses, 45 EMTs, 38 physicians, 27 paramedics, 14 nurse's assistants/emergency department technicians, 10 firefighters, five respiratory therapists, and four students. Seventy-one percent (149/209) of participants failed to give adequate compressions by either method. Only 40 participants performed adequate compressions using the two-thumb method (95% confidence interval. 14-25%). Thirty-eight participants gave adequate compressions using the two-finger method (95% confidence interval, 13-24%). No statistically significant difference existed between the two groups (P = 0.877; the McNemar test). A statistically significant difference was found in the number of shallow compressions for each method. Forty participants (19.1%) had more than 40 compressions that were too shallow versus 15 (7.2%) using the two-thumb method (P < 0.005). CONCLUSIONS: Medical personnel often fail to give adequate compressions. The two-thumb method was as adequate as the two-finger method. Overall, more compressions were measured as shallow with the two-finger method. PMID- 10711491 TI - Massive postoperative swelling of the tongue: manual decompression and tactile intubation as a life-saving measure. AB - Massive swelling of the tongue due to haemorrhage is a rare but potentially fatal complication secondary to trauma, surgery, tumour invasion or uncontrolled anticoagulant therapy. This article presents a report of bleeding from the left lingual artery secondary to elective excision of a lipoma of the floor of the mouth and subsequent life-threatening upper airway obstruction. In this case, the upper airway obstruction was managed by manual decompression of the tongue and tactile nasal intubation. To our knowledge this case provides the first description of using this method in life-threatening upper airway obstruction caused by massive haemorrhagic swelling of the tongue. PMID- 10711492 TI - Recalling the causes of pulseless electrical activity (PEA) PMID- 10711493 TI - About being cited in tables. PMID- 10711494 TI - Forced air surface rewarming in patients with severe accidental hypothermia. PMID- 10711495 TI - Expression and function of EGF-related peptides and their receptors in gynecological cancer--from basic science to therapy. AB - EGF-related peptides and their receptors play an important, but not fully understood role, both, in epithelial physiology and pathophysiology but also in human tumor carcinogenesis and tumor behavior, respectively. Overexpression of EGF-related growth factors from normal epithelium to carcinomas has been demonstrated for several human tissues such as breast, endometrium, cervix and ovary. Additionally, the differential overexpression of EGFR or erb B-2 in various malignancies has already proven to be efficacious in stratifying patients with respect to a poor prognosis. These data suggest that EGF-related growth factors, erb B receptors or signaling proteins that function either upstream or downstream from these receptors may represent novel targets for selective tumor therapy. In the future, conventional chemotherapy regimes will ultimately be wedded to more biologically-oriented therapies. One important target for these novel therapeutic approaches in solid tumors will be the EGF-related growth factors and their receptors. PMID- 10711496 TI - Species-dependent pharmacological properties of the melanocortin-5 receptor. AB - The genes encoding the melanocortin-3 receptor and melanocortin-5 receptor have been cloned from rhesus monkey. Heterologous expression in CHO cells indicated species dependent in vitro pharmacological properties for the human and rhesus melanocortin-5 receptors. Several peptides including NDP-alpha-MSH, alpha-MSH, MT II and ACTH1-24 are more potent at the rhesus melanocortin-5 receptor than the human melanocortin-5 receptor by more than 10-fold. In contrast, we found no species difference in pharmacological properties between the human and rhesus melanocortin-3 receptors. Such a species-dependent pharmacological difference for melanocortin-5 receptor appears to be an exception compared to other G protein coupled receptors from human and rhesus monkey. PMID- 10711497 TI - Golf complements a GPA1 null mutation in Saccharomyces cerevisiae and functionally couples to the STE2 pheromone receptor. AB - We have produced a plasmid designed for the expression of heterologous G protein alpha subunits in the yeast Saccharomyces cerevisiae. Introduction of these genes is by simple cassette replacement using unique restriction sites, and their expression is controlled by the regulatory sequences of the S. cerevisiae GPA1 gene. Levels of expression are therefore suitable for interaction of these heterologous proteins with elements of the yeast pheromone response pathway. We believe that this plasmid will facilitate the coupling of more members of the seven transmembrane domain superfamily of receptors, through their native G protein alpha subunit, to the yeast pheromone response pathway. The plasmid pRGP, is a stable centromeric shuttle vector with a HIS3-selectable marker. We have demonstrated that production of GPA1 from this plasmid functionally complements a gpal1- null mutation. A similar response is obtained when an alternative G protein alpha subunit, G(olf), is introduced using pRGP. We believe that this is the first example of a heterologous G protein shown to couple to a yeast pheromone receptor. PMID- 10711498 TI - Functional receptor coupling to Gi is a mechanism of agonist-promoted desensitization of the beta2-adrenergic receptor. AB - The beta2-adrenergic receptor (beta2AR) couples to Gs activating adenylyl cyclase (AC) and increasing cAMP. Such signaling undergoes desensitization with continued agonist exposure. Beta2AR also couple to Gi after receptor phosphorylation by the cAMP dependent protein kinase A, but the efficiency of such coupling is not known. Given the PKA dependence of beta2AR-Gi coupling, we explored whether this may be a mechanism of agonist-promoted desensitization. HEK293 cells were transfected to express beta2AR or beta2AR and Gialpha2, and then treated with vehicle or the agonist isoproterenol to evoke agonist-promoted beta2AR desensitization. Membrane AC activities showed that Gialpha2 overexpression decreased basal levels, but the fold-stimulation of the AC over basal by agonist was not altered. However, with treatment of the cells with isoproterenol prior to membrane preparation, a marked decrease in agonist-stimulated AC was observed with the cells overexpressing Gialpha2. In the absence of such overexpression, beta2AR desensitization was 23+/-7%, while with 5-fold Gialpha2 overexpression desensitization was 58+/-5% (p<0.01, n=4). The effect of Gi on desensitization was receptor-specific, in that forskolin responses were not altered by G(i)alpha2 overexpression. Thus, acquired beta2AR coupling to Gi is an important mechanism of agonist-promoted desensitization, and pathologic conditions that increase Gi levels contribute to beta2AR dysfunction. PMID- 10711499 TI - Expression of OB receptor splice variants during prenatal development of the mouse. AB - In adult animals, signaling through the leptin receptor (OB-R) has been shown to play a critical role in fat metabolism. However, it is not known when these receptors are first expressed and what their role may be during embryonic development. To date, at least 6 splice variants of the OB-R have been identified. Although the function of each of these individual splice variants are unknown, only one of them, ob-rL ,encodes a receptor with a long intracellular domain that is implicated in OB-R signaling. In this study we have used in situ hybridization to examine the localization of OB-R splice variants during embryonic development of C57B1/6J mice. Using a probe, ob-r, that recognizes all of the splice variants, ob-r mRNA was found to be distributed in developing bone, mesenchyme, notochord and liver. In addition, epithelial structures including leptomeninges, choroid plexi and hair follicles also expressed ob-r. No ob-r mRNA was detected in the CNS. ob-rL, expression was only detected in notochord, bone and mesenchyme. The differential expression of these two mRNA isoforms suggests that the extracellular and intracellular domains of the OB receptor perform different biological functions. PMID- 10711500 TI - Screening of ligand binding on melatonin receptor using non-peptide combinatorial libraries. AB - The screening of combinatorial libraries requires a deconvolution procedure to obtain, in fine, the most active compound of the starting library. The standard screening assays used in regular molecular pharmacology, have been poorly assessed when transposed to combinatorial chemistry-related experiments, particularly those involving large numbers of chemicals in a single assay. One key issue is the effect of the inactive analogs on the identification of the active ligand in mixtures. We chose melatonin receptors to measure the apparent affinity of a single ligand when tested alone or in mixtures of non-peptide low molecular weight compounds. Using ligands with IC50 from the micro- to the picomolar range, mixed with increasingly complex mixtures of 5 to 20 or 25 inactive compounds, we analyzed the displacements from the mt1 and MT2 melatonin receptor subtypes of the radioligand 2-iodomelatonin (KD= 25 pmol/l and 200 pmol/l, respectively) . The behavior of equimolar mixtures in displacement curves led to the conclusion that the observed binding affinity reflects the dilution effect of mixing the active component with inactive compounds but does not reveal noticeable interactions which would interfere with the binding process. From the practical point of view, the concentrations of the active species in the binding assay should be large enough to displace significantly the radioligand, a requirement which may be limited by the solubility of the ligand mixtures. In contrast, previous observations with peptide libraries report that the dilution effect is often compensated by additive or synergic action of structurally related analogs, thus making possible the deconvolution of very large (typically up to 10(7) compounds) peptide libraries. PMID- 10711501 TI - Selective staining by the fluorochrome, 5,5-diphenyl-9-ethyl-DiOC2(3). I. Physicochemical studies of dye-dye and dye-tissue interactions. AB - The 5,5'-diphenyl-9-ethyl-oxacarbocyanine (5,5'-diphenyl-9-ethyl-DiOC2(3); CD) has properties suitable for histological investigations including the spectral range for absorption and fluorescence emission, the values of the corresponding molar coefficients and fluorescence quantum yield. Furthermore, CD remains relatively unchanged over the entire pH range and interacts with protein beta sheets. The latter fact is detectable spectroscopically as a bathochromic shift. In water-containing media such as the histological stain and washes, CD exists as a monomer, a dimer and in two aggregated states. These differ in their binding affinity to tissue sections, in their solubility in water, alcohol and water/alcohol mixtures, and in their UV/VIS absorption and fluorescence emission. The ratio of the various CD states and the contrast of selectively stained tissue areas can be controlled via the staining conditions and the sequence of the washes. Furthermore, mounting in a xylene-based medium produces a solvatochromic spectral shift of the CD monomer, which leads to a marked elevation in phase contrast. PMID- 10711502 TI - Selective staining by the fluorochrome 5,5-diphenyl-9-ethyl-oxacarbocyanine. II. Application to paraffin embedded nervous tissue. AB - We present a simple and rapid method to label specific structures of neural tissue in paraffin sections. After incubation of deparaffinized sections in the cyanine dye 5,5'-diphenyl-9-ethyl-oxacarbocyanine (DEOC), three different washing and mounting procedures were performed. Incubation in DEOC followed by washes in ethanol and water and mounting in glycerol-gelatin resulted in selective labeling of myelin in the central and peripheral nervous systems. Weak labeling of myelin and axons and staining of nuclei of neurons was seen after incubation in DEOC followed by washes in ethanol and xylene, and mounting in Eukitt. Nuclear proteins, the endoplasmic reticulum and the Golgi apparatus were stained strongly and exclusively after incubation in DEOC, washes in water, ethanol and xylene, and mounting in Eukitt. Thus the absorption pattern of DEOC is changed significantly by solvents applied after the fluorochrome. In any case, fluorescence did not fade even after repeated and intense fluorescence microscopy over a period of 6 months. PMID- 10711503 TI - Diaminobenzidine induces fluorescence in nervous tissue and provides intrinsic counterstaining of sections prepared for peroxidase histochemistry. AB - 3,3'-Diaminobenzidine (DAB) is widely used as a chromogen for visualization of horseradish peroxidase activity in neuroanatomical tracing experiments and in immunohistochemistry. The product of the enzymatically catalyzed oxidation of DAB by hydrogen peroxide is brown and nonfluorescent. In frozen sections of formaldehyde fixed rat and mouse brain that had been exposed to DAB either alone or with hydrogen peroxide, we observed strong greenish fluorescence in myelinated nerve fibers and in the somata of some neurons. This fluorescence was not associated with brown coloration and was not due to endogenous peroxidase activity. Extractions, blocking reactions, and other histochemical tests indicate that the fluorescence resulted from the combination of DAB with aldehyde groups that were formed by oxidation of unsaturated linkages in lipids. DAB induced fluorescence provides a simple and useful demonstration of background anatomy in sections that also contain specifically localized deposits of peroxidase activity. PMID- 10711504 TI - A simple drying method for field emission scanning electron microscopy for chromosomes. AB - Hexamethyldisilazane treatment and subsequent air drying of spread plant chromosomes is compared with critical point drying. The two procedures are equivalent for preparing chromosomes for examination by field emission scanning electron microscopy at low voltage. PMID- 10711505 TI - Fixative-dependent increase in immunogold labeling following antigen retrieval on acrylic and epoxy sections. AB - We examined the increase in immunogold labeling of variably fixed, resin embedded tissue sections following antigen retrieval by heating in citrate solution. Fibrin clots and porcine renal tissue were fixed in glutaraldehyde, paraformaldehyde or ethanol, and specimens were embedded in LR-White or epoxy resin. Immunogold labeling was performed on ultra-thin sections with anti fibrinogen for the fibrin clots and anti-IgG for the porcine renal tissue. Immunogold labeling increased greatly after heating epoxy sections regardless of the fixative used. The ratio labeling(retrieved)/labeling(nonretrieved) (Lr/Ln) was 2.8 or higher, and the largest increases were obtained for anti-IgG. Heating induced a large increase of immunolabeling for LR-White sections only when the specimens had been fixed in paraformaldehyde (Lr/Ln = 2.2 for anti-IgG and 1.4 for antifibrinogen). LR-White sections showed decreased, insignificant or weakly increased immunolabeling of ethanol or glutaraldehyde fixed tissues following antigen retrieval. Disruption of aldehyde cross-links is not the only mechanism for antigen retrieval when epoxy sections are heated in citrate solution since large increases in immunolabeling were obtained on ethanol fixed tissue. The large heat-induced increases in immunolabeling on epoxy sections are probably caused by the disruption of chemical bonds between the epoxy resin and side groups of proteins. PMID- 10711506 TI - Flow cytometric method using fluorescent microspheres to measure reticuloendothelial function or particulate translocation. AB - The reticuloendothelial system (RES) influences the outcome of vascular shock and environmental stress. We describe a procedure that employs flow cytometry and 1 microm fluorescent microspheres (FM) to study RES function. FM (2 x 10(10) beads/kg) were administered via a jugular cannula in Sprague-Dawley rats. After 15 min, blood and tissues were collected and digested in 15% KOH. Phycoerythrin 1 microm beads were added to each sample as an internal standard and analyzed by flow cytometry. FM were preferentially cleared by the spleen, liver and lung. Clearance was confirmed by fluorescent photomicroscopy. Addition of the internal standard to determine accurately aspiration volume enhanced precision. This procedure offers advantages over other RES clearance methods including bacterial, radioactive or carbon clearance assays. Moreover, this method could enhance accuracy, reproducibility and speed of data collection in particulate transport studies that are based on manual microscopic scanning and FM counting. PMID- 10711507 TI - Morphological study of the role of calcium in the assembly of the rotavirus outer capsid protein VP7. AB - Maturation of rotavirus occurs in the endoplasmic reticulum (ER), a site of intracellular calcium storage. It was demonstrated previously that calcium plays an important role in the maturation of bovine rotavirus. We used protein A colloidal gold conjugated to an antibody to localize VP7, the outer capsid protein of the simian rotavius SA11, in permeabilized infected cells in the presence and absence of calcium in the culture medium. In medium containing calcium, VP7 was associated with nonenveloped double-shelled particles and membranous structures of the ER. In calcium-free medium, gold particles were not associated with the ER or with virus particles. Gold particles were distributed through the cytoplasm and were mainly associated with granular structures, but did not assemble onto virus particles. Our data suggest that in calcium-free medium, VP7 is synthesized, but does not remain incorporated, in the ER. PMID- 10711508 TI - Overview of the radiology of connective tissue disorders in children. AB - This review article describes the imaging finding of the connective tissue disorders in children. The radiological features of the following conditions are described; the spondyloarthropathics, systemic lupus erythematosus (SLE), dermatomyositis, scleroderma, the vasculitides, Kawasaki disease, synovitis, acne, pustulosis, hyperostosis, osteitis (SAPHO), and focal myositis. The features on several integrated imaging techniques are described. PMID- 10711509 TI - MRI and ultrasound in children with juvenile chronic arthritis. AB - In this era of advancing imaging technology, a knowledge of the relative values of available imaging techniques is necessary to optimize the management of children with juvenile chronic arthritis (JCA). After clinical examination, plain films remain the initial investigation. The need for radiation protection must be a priority in children with JCA. Conventional radiographs allow grouping of the various arthritides (on the base of the distribution and pattern of joint space changes) and staging of disease progression. Ultrasound (US) is very sensitive in the detection of joint effusions, especially in the hip, and guides fluid aspiration. US and Doppler can be used for the evaluation of synovial hypertrophy and activity. Arthrography and to a certain extent nuclear studies have been replaced by magnetic resonance imaging (MRI). MRI can demonstrate articular cartilage, joint effusion, synovial hypertrophy, cortical and medullary bone, cartilage and bone perfusion, and fibrocartilaginous structures (menisci and ligaments). Contrast enhanced MRI is the most sensitive modality to determine whether an arthritic condition is present. However, it does not assist in establishing a specific diagnosis. MRI determines accurately the activity and the extent of the disease and is particularly useful in the early detection of articular damage. Finally, MRI is of major importance in the evaluation of response to local therapy (especially steroids) and the detection of complications. PMID- 10711510 TI - Overview of the radiology of juvenile idiopathic arthritis (JIA). AB - Plain films remain the basic tool for diagnosis and follow-up evaluation of juvenile idiopathic arthritis (JIA). In this paper, we review the new classification of JIA: systemic arthritis. oligoarthritis (persistent), oligoarthritis (extended), polyarticular arthritis (rheumatoid factor negative), polyarticular arthritis (rheumatoid factor positive), enthesitis-related arthritis, psoriatic arthritis and unclassified arthritis. We will also review regional abnormalities of three stages: an early stage, an intermediate stage, a late stage, as well as the differential diagnosis. PMID- 10711511 TI - Joint disease in children of Asiatic origin. AB - Joint disorders in Asian children are varied due to the diversity of the Asian population and show some ethnic trends. The ethnic diversity, socio-economic and geographic factors in Asia have limited the availability of data from some of the ethnic groups, many of whom live in remote and relatively underdeveloped areas, are not subjected to epidemiological surveillance and have little awareness of these diseases and their consequences. Geographic and socio-economic factors also play a significant role in some of the joint diseases peculiar to Asian children. In general, the current available data suggests that there are no large differences in the epidemiology and clinical features between the Western and Asian children. This article reviews the available literature on joint diseases in Asian children. PMID- 10711512 TI - Overview of the clinical presentation of connective tissue diseases in children. PMID- 10711514 TI - Juvenile idiopathic arthritis: a clinical overview. AB - The chronic arthritides in childhood remain a poorly understood group of conditions. Their classification has been a source of much confusion over the years with differences in terminology between Europe and North America. A significant step forward in paediatric rheumatology has been the recent development of an internationally agreed classification system which uses the overall term juvenile idiopathic arthritis (JIA). The various subtypes of JIA and their clinical features are described, together with an overview of their differential diagnosis, complications and outcomes. An outline of current management strategies is given and potential future developments highlighted. PMID- 10711513 TI - Rheumatological presentation of developmental bone diseases. AB - Developmental bone disease may be present, with rheumatological disorders as the major symptoms, even in children. The major lesions encountered are early osteo arthritis, osteo chondromatosis and vertebral involvement with two leading types, pseudo Scheuermann's disease or pseudo ankylosing spondylitis. This paper presents the different features and lists the rheumatological problems in bone dysplasia. PMID- 10711515 TI - The persistently irritable joint in childhood an orthopaedic perspective. AB - Persistent joint or periarticular irritability and pain in children can have numerous explanations. This overview explores the diverse range of orthopaedic conditions, both acquired and congenital, that may lead to such a presentation. PMID- 10711516 TI - FDA, Janssen bolster cardiac risk warnings for cisapride. PMID- 10711517 TI - Oxcarbazepine approved for partial seizures. PMID- 10711518 TI - HHS accuses infusion companies of overcharging. PMID- 10711519 TI - Recap of FDA product approvals--1999. PMID- 10711520 TI - Managed care may not have edge in preventive medicine. Greater use of behavior change strategies advocated. PMID- 10711521 TI - Patient outcomes comparable for primary care physicians, independent nurse practitioners. PMID- 10711522 TI - Zaleplon. PMID- 10711523 TI - Pharmacists and patient care: stop talking, start doing. PMID- 10711524 TI - Sirolimus: a new agent for prevention of renal allograft rejection. AB - The pharmacology, pharmacokinetics, clinical efficacy, adverse effects, and dosage and administration of sirolimus are reviewed. Sirolimus can be used as an adjunct to cyclosporine and corticosteroids for preventing organ rejection in renal allograft recipients. It appears to block cell-cycle progression in the mid to-late G1 phase; it also inhibits interleukin-2-dependent and independent proliferation of B-lymphocytes and production of immunoglobulins A, M, and G. Sirolimus is extensively metabolized by the liver and undergoes O-demethylation and hydroxylation. The mean oral clearance is 208 +/- 95 mL/hr/kg with an elimination half-life of 57 to 63 hours in kidney transplant recipients. Sirolimus is poorly absorbed from the gastrointestinal tract; its apparent oral bioavailability is about 15%. Two pivotal Phase III, multicenter, randomized, double-blind trials evaluating the efficacy of sirolimus in the prophylaxis of acute renal allograft rejection after transplantation have been conducted. A significant reduction in the occurrence of acute rejection was noted in both trials compared with azathioprine- or placebo-treated patients. Also, both sirolimus groups required significantly less antibody therapy for treatment of rejection. The primary adverse effects associated with sirolimus therapy include hypercholesterolemia and hypertriglyceridemia. Hypertension, diarrhea, increased creatinine, hypokalemia, arthralgia, rash, acne, leukopenia, and thrombocytopenia have also been observed. The labeled recommendations are a 6-mg loading dose as soon as possible after transplantation and a maintenance dose of 2 mg per day. Sirolimus is a potent novel immunosuppressive agent with proven efficacy in the prevention of acute rejection after renal transplantation. Use of sirolimus may lead to effective therapeutic strategies for sparing the use of cyclosporine and corticosteroids. PMID- 10711525 TI - Observational cohort study of switching warfarin sodium products in a managed care organization. AB - The effectiveness and safety of switching warfarin sodium products in a cohort of patients at a managed care organization were studied. Consecutive patients seen during routine visits at two anticoagulation clinics were requested to voluntarily use a generic warfarin sodium product (Barr Laboratories) instead of Coumadin (Dupont Pharma) for eight weeks. All patients who had received anticoagulation therapy with Coumadin for more than three months, who had a target International Normalized Ratio (INR) of 2.0-3.0 or 2.5-3.5, and who had an INR in the target range at the baseline visit were eligible for the study. The control group was composed of patients who were eligible for the study but were not asked to switch to the generic product or declined to switch products. There were 105 patients in each study group. The before-and-after differences in INR between the conversion group and the control group were small and of negligible clinical importance. Compared with the period before the product switch, the variability of the INR in patients in the conversion group was not significantly different after the switch. Of the 210 patients in the study, only 24 had a change in INR of >1.0 after the baseline visit; in each case, a factor not related to the product switch was associated with the INR change. Use of a generic warfarin sodium product (Barr Laboratories) in patients previously receiving the innovator product (Dupont Pharma) did not change the INRs more than did continued use of the innovator product by another group of patients. PMID- 10711526 TI - Relationship between selected personality traits and citation for violating pharmacy board regulations. AB - The relationship between certain personality traits and citation for pharmacy board violations was studied. The Gordon Personal Profile-Inventory (GPP-I) was mailed to three samples of pharmacists licensed in North Carolina (95 pharmacist leaders, 199 pharmacists who had been cited for violating one or more board of pharmacy regulations, and a random sample of 494 pharmacists licensed in the state). The pharmacists were asked to provide demographic information and to complete the 38-item GPP-I, which measures eight different personality traits (ascendancy, responsibility, emotional stability, sociability, cautiousness, original thinking, personal relations, and vigor). The response rates for the three samples were 78.9%, 23.6%, and 58.3%, respectively. Pharmacists who had been cited for one or more board of pharmacy violations had significantly lower scores on the GPP-I for the personality trait vigor than general pharmacists. They also had lower scores on the GPP-I traits of ascendancy, original thinking, and vigor than pharmacy leaders. They were less likely to have advanced degrees or belong to any pharmacy organizations. They tended to be male, older, and out of school longer than those pharmacists who had never been cited for violating one or more board of pharmacy regulations. Significant differences in personality traits were found between pharmacists who had been cited for violating board of pharmacy regulations and general pharmacists and pharmacy leaders. PMID- 10711527 TI - Rhabdomyolysis in a patient receiving the combination of cerivastatin and gemfibrozil. AB - Straightforward reports of unusual drug experiences are included in this section. While selected references may be cited, the purpose of a Drug Experience report is not to present an extensive review of the literature. A related section, Grand Rounds, includes papers that are well-documented patient case reports with a thorough review of the important literature to help put the case in perspective. Authors should report serious adverse drug reactions to the FDA medical products reporting program (MedWatch). Such reporting will not jeopardize the chances that AJHP will publish manuscripts on the same drug reactions. PMID- 10711528 TI - Focus on the future: ASHP 2000. American Society of Health-System Pharmacists. PMID- 10711529 TI - Developing a strategic plan for quality in pharmacy practice. PMID- 10711530 TI - Pharmacy in the American Civil War. AB - The role of pharmacists and the process of military drug supply in the American Civil War are described. Most raw drugs used in the United States in the mid 1800s were imported. During the Civil War, imports into the North continued, but the Union blockade forced the Confederacy to obtain medicines through means such as smuggling, capture of enemy supplies, and processing of indigenous medicinal plants. Medical supplies for Civil War troops were typically purchased by military physicians called medical purveyors and sometimes by pharmacists serving as acting medical purveyors. In the latter half of the war, U.S. Army medical laboratories, in which many pharmacists were employed, inspected purchases, repackaged supplies bought in bulk, and manufactured medicines from raw materials. The Confederacy also had medical laboratories, which were primarily responsible for manufacturing medicines from indigenous plant material but also inspected drugs that had been smuggled into the South. At a few large Union medical depots, pharmacists called medical storekeepers assumed many of the responsibilities of medical purveyors by receiving, storing, issuing, and accounting for supplies. Noncommissioned officers called hospital stewards assumed diverse duties that included dispensing drugs prescribed by military physicians. Although many hospital stewards were pharmacists or physicians, others had no previous pharmaceutical experience. Civilian pharmacists were employed in the medical laboratories and in military general hospitals. Pharmacists participated in nearly every aspect of military drug supply during the Civil War. PMID- 10711531 TI - Tricks of the trade in starting efavirenz therapy. PMID- 10711532 TI - Habitat alteration and the conservation of African primates: case study of Kibale National Park, Uganda. AB - Tropical forests and the animals they support are being threatened by accelerating rates of forest conversion and degradation. In a continually fluctuating sociopolitical world, it is often impossible to protect areas from such conversion until the political environment is suitable to pursue conservation goals, by which time, the forests have often been converted to other uses. This reality suggests a need for inquiry into which primate species can persist after different types of disturbances and how quickly primate communities can recover from disturbance. Here we examine the persistence of primate populations in disturbed habitats by providing a case study of patterns of primate abundance in areas of Kibale National Park (766 km2), Uganda, that have been modified by different types and intensities of human activities, primarily commercial logging and agricultural clearing. Distributional surveys at 24 sites and detailed line-transect censuses at six sites demonstrate that primate populations in Kibale are often high and suggest that patterns of population change associated with disturbance are complex. Analysis of the land use coverage of Kibale reveals that abandoned farms (10.3%) and degraded forest (8.7%) now cover 146 km2. Unfortunately, we do not know what proportion of the farms were established on areas that were forest versus grassland. However, if the areas that are now abandoned farms were all once forested, this means that 79 km2 of forest has been lost. Based on density estimates from nearby sites, this would represent a loss of 52,612 monkeys and 200 chimpanzees. Populations would also have been affected by the degradation of the 66 km2 (8.7%) of forest. These estimates of the potential reductions in the primate populations that could have resulted from forest clearing and degradation illustrate the importance of protecting land. A review of the literature illustrates that the biomass of primates found within Kibale is very high in comparison to other locations and thus illustrates the importance of Kibale to regional conservation. PMID- 10711533 TI - Intra-troop affiliative relationships of females with newborn infants in wild ring-tailed lemurs (Lemur catta). AB - To determine how the birth and development of infants influence their mothers' social relationships with other adult troop members, we observed two free-ranging troops of ring-tailed lemurs (Lemur catta) at the Berenty Reserve, Madagascar. The number of acts of affiliative contact that the mothers received from other adult troop members during the first and second months of infant life were significantly higher than those before they gave birth, and the values during the third month were as low as that before giving birth. Two mothers received acts of affiliative contact less frequently after their infants died, compared with the values while the infants were alive. On the other hand, more than 95% of all acts of licking of infants by adult troop members other than their mothers occurred when the infants were in contact with their mothers. These findings suggest that infants per se and mothers per se were not attractive, but rather the mother infant pair was attractive to other troop members. Acts of infant-licking were observed in almost all mother-mother pairs and mother-non-mother adult female pairs, and in two thirds of mother-adult male pairs. Moreover, the frequency of infant-licking was not affected by female parity, female and male dominance rank, or infant sex. Therefore, acts of infant-licking, which are widespread among troop members, may function to make or maintain stable social relationships. PMID- 10711534 TI - Conservation and evolution of microsatellite loci in primate taxa. AB - Microsatellites are promising genetic markers for the study of demographic structure and phylogenetic history in populations. However, little information exists on the molecular nature of the repeats and their flanking sequences of a same microsatellite in a large range of species. In this study, we report polymorphism and consensus sequences of eight microsatellite loci using human primers in 20 primate species. The results show size polymorphism in almost all species and microsatellites. These loci are therefore useful markers for population genetic studies between populations of the same species. Insertion/deletion events are frequent in the flanking regions, the majority concerning several contiguous bases. This is in contrast with the more usual single base pair events in non-coding regions. The ranges of allele lengths in non-human primates often show no overlap with that of human, usually due to the deletion/insertion events in the flanking sequences, producing smaller allele lengths rather than smaller numbers of repeats. The use of length of PCR product will bias the inter-species interpretation reducing the number of observable alleles and treating as the same allele very divergent molecular sequences. Caution should be used when employing microsatellites in cross-species comparisons in which the species under study are separated by significant amounts of evolutionary time: in such cases allele comparison cannot be based on lengths alone. PMID- 10711535 TI - Habitat use by Chiropotes satanas utahicki and syntopic platyrrhines in eastern Amazonia. AB - Primates were surveyed at two sites in the Xingu-Tocantins interfluvium, in eastern Amazonia, where at least eight platyrrhines are known to occur, including the endemic Chiropotes satanas utahicki, vulnerable to extinction. Only three other forms; Alouatta belzebul belzebul, Cebus apella apella, and Saguinus midas niger; were recorded at both sites. Data on habitat use (forest type and strata) were collected in standard line transect surveys and analyzed with relation to the availability of forest types, as well as between sites and species. The smallest- (S. midas) and largest-bodied (A. belzebul) species were relatively common at the continuous forest site, where they exhibited a significant preference for primary terra firme forest. At this site, Cebus demonstrated a significant preference for liana and flooded forest in contrast with primary or secondary terra firme forests. The medium-sized Cebus and Chiropotes were more common in the isolated forest fragment (where they were also observed together frequently), but no clear habitat preferences were found at this site for any species. A. belzebul occupied significantly higher forest strata than other species, which all used relatively similar levels. C.s. utahicki was active in much lower forest strata than other bearded sakis, whereas S. midas was observed at much higher levels than at other sites in eastern Amazonia. It remains unclear whether and to what extent observed patterns are determined by differences between taxa, populations, or ecosystems, but the data indicate that C.s. utahicki is relatively tolerant of habitat disturbance. PMID- 10711536 TI - Effectiveness of hysterectomy. AB - OBJECTIVE: To measure the effectiveness of hysterectomy in relieving adverse symptoms and to identify factors associated with lack of symptom relief. METHODS: In a 2-year prospective study, data were collected before and at 3, 6, 12, 18, and 24 months after hysterectomy in 1,299 women who had hysterectomies for benign conditions at 28 hospitals across Maryland. Effectiveness was measured in terms of relief of symptoms such as problematic vaginal bleeding, pelvic pain, and urinary incontinence. Psychologic function and quality of life before and after surgery also were assessed. RESULTS: Symptom severity, depression, and anxiety levels decreased significantly after hysterectomy and quality of life improved, particularly in the area of social function. However, 8% of women had at least as many symptoms at problematic-severe levels 1 and 2 years after hysterectomy as before. In multiple logistic regression, several presurgical patient characteristics predicted lack of symptom relief, including therapy for emotional or psychologic problems, depression, and household income of $35,000 or less. Bilateral oophorectomy predicted lack of symptom relief at 24 months but not at 12 months after hysterectomy. CONCLUSION: Significant improvements were seen after hysterectomy for all three aspects of health status (symptoms, psychologic function, and quality of life), which persisted or continued to improve throughout the 2 years of follow-up. However, hysterectomy did not relieve symptoms for some women, particularly those who had low incomes or were in therapy at the time of hysterectomy. PMID- 10711537 TI - Periurethral collagen injection for stress incontinence with and without urethral hypermobility. AB - OBJECTIVE: To compare the use of periurethral collagen injection in the treatment of female stress urinary incontinence due to intrinsic sphincter deficiency in women with and without urethral hypermobility. METHODS: A retrospective review was performed of 60 periurethral collagen injections performed on 40 consecutive women from January 1996 to December 1997. A review of the office chart and operative notes was performed to obtain demographic, urodynamic, and procedural data. Outcome data were obtained by personal or telephone interview, using patients' subjective assessments including an analog satisfaction scale. RESULTS: Nine of 40 patients (23%) had urethral hypermobility. Compared with patients without hypermobility, patients with hypermobility required a similar number of procedures (a mean of 1.9 compared with 1.4, P = .13) and required similar amounts of collagen on the first injection (5.6 mL compared with 5.3 mL, P = .69). Preoperative urodynamic parameters were similar in both groups. Rates of subjective dryness were equivalent in patients with and without hypermobility at 1 month (76% and 46%, P = .24) and at 6 months (71% and 32%, P = .09) following initial injection. A post hoc power analysis was performed to evaluate the primary study measures of continence at 1 and 6 months, and number of collagen injections. This revealed that a sample size of 40 patients would be sufficient to detect a 2.5-fold difference in number of injections and a 3-fold difference in subjective dryness. CONCLUSION: Coexisting urethral hypermobility should not preclude the use of collagen injections in women with stress urinary incontinence. PMID- 10711538 TI - Lumbosacral spine and pelvic inlet changes associated with pelvic organ prolapse. AB - OBJECTIVE: To determine the association between advanced pelvic organ prolapse and changes in lumbar lordosis and/or pelvic inlet orientation. METHODS: Lateral lumbosacral spine/pelvic x-rays were taken of women with grade 2 or greater uterovaginal prolapse and women with grade 1 or less prolapse standing in their usual upright posture. The angles of lumbar lordosis and the pelvic inlet were measured by a radiologist who was masked to the pelvic examination findings. RESULTS: Twenty women with prolapse were matched with 20 women without significant prolapse. There were no significant differences in the mean (+/- standard deviation [SDI) age (55.3 +/- 9.0 years compared with 53.4 +/- 9.5 years), body mass index (BMI) (28.9 +/- 5.6 compared with 28.4 +/- 5.2), gravidity (5.6 +/- 3.5 compared with 5.0 +/- 2.7), and vaginal parity (4.65 +/- 3.3 compared with 4.5 +/- 2.9) between the prolapse and nonprolapse groups, respectively. All participants were vaginally parous. The mean lumbar lordotic angle in women with pelvic organ prolapse (32.0 degrees +/- 9.8 degrees) was significantly lower than that of controls (42.4 degrees +/- 10.9 degrees) (P < .003). The mean angle of the pelvic inlet in women with pelvic organ prolapse (37.5 degrees +/- 7.0 degrees) was significantly larger than that of controls (29.5 degrees +/- 7.3 degrees) (P < .001). The differences in the mean angles of lumbar lordosis and the pelvic inlet, between the case and control groups, remained significant after multivariable logistic regression was performed. CONCLUSION: Women with advanced uterovaginal prolapse have less lumbar lordosis and a pelvic inlet that is oriented less vertically than women without prolapse. PMID- 10711539 TI - Late presentation of ureteral injury after laparoscopic surgery. AB - OBJECTIVE: To describe clinical presentation, etiology, and treatment of ureteral injuries recognized late in women who had gynecologic laparoscopies. METHODS: We reviewed the charts of 12 women who had delayed recognition of ureteral injuries between January 1991 and December 1998. RESULTS: Patients presented with fever, hematuria, flank pain, or peritonitis between 3 and 33 days postoperatively. The mechanism of ureteral injuries was electrocoagulation in seven women, laser ablation in one, and stapler ligation in four. The sites of injury were near the inferior margin of the sacroiliac joint on excretory urogram in eight women and near the ureterovesical junction in four. Three women initially treated with internal ureteral stents were subsequently treated with ureteroneocystostomy because of progression of urinary ascites in two and a delayed ureteral stricture in one. In nine patients, attempts at ureteral stenting were unsuccessful and immediate ureteral reconstruction was done. Outcomes were good in all cases. CONCLUSION: Delayed recognition of ureteral injury after gynecologic laparoscopy was associated with serious complications, and initial treatment with ureteral stenting was not useful. We advocate early open repair for those injuries. PMID- 10711540 TI - Comparison of immediate and deferred colposcopy in a cervical screening program. AB - OBJECTIVE: To evaluate the effect of delaying colposcopy in women with negative Papanicolaou smears and positive speculoscopy results. METHODS: This was a prospective study of asymptomatic women ages 13-60 years, regularly scheduled for pelvic examinations. All women had Papanicolaou smears and magnified visual examinations with speculoscopy. Women with negative Papanicolaou smears and positive speculoscopy results were quasirandomized to immediate or deferred colposcopy groups. RESULTS: A total of 800 women completed all phases of the study, 124 of whom had negative Papanicolaou smears and positive speculoscopy results. Among 57 women who had immediate colposcopies, 64.9% had histologic evidence of neoplasia. Sixty-seven women had their scheduled colposcopies deferred for 6 months. During this period, 21% (14) were lost to follow-up and 29% (13) of those evaluated converted from speculoscopy positive to speculoscopy negative. Among the 32 (71%) women who remained speculoscopy positive, 90% were found to have histologic evidence of neoplasia on colposcopic biopsy. CONCLUSION: In women with normal Papanicolaou smears and positive speculoscopy results, the diagnostic yield can be improved by deferring colposcopy for 6 months. Deferral should be considered only for women who are reliable for follow-up. PMID- 10711541 TI - Anesthetic efficacy of intrauterine lidocaine for endometrial biopsy: a randomized double-masked trial. AB - OBJECTIVE: To determine the efficacy of intrauterine lidocaine for decreasing pain associated with endometrial biopsy using the Pipelle instrument (Unimar; Wilton, CT). METHODS: Forty-one premenopausal and postmenopausal women had 5 mL of either 2% lidocaine or saline instilled in their uteri before endometrial biopsies. Subsequently, each woman completed a 20-cm visual analogue scale for subjective pain experience. We compared histologic findings in endometrial specimens. RESULTS: Before the study, analysis of specimens (n = 6) found no histologic effect on ability to interpret endometrial biopsies by pathologists who were masked to lidocaine or saline. There was no statistically significant difference in age, parity, race, history of chronic pelvic pain, menopausal status, use of tenaculum, or prior endometrial biopsy. The procedure was easy, with no clinically significant side effects. On the visual pain scale, the median (range) score for the lidocaine group (4.7, 0-19.7) compared with the placebo group (9.9, 1.6-20) showed significant reduction in pain corresponding with a decrease from moderate to mild (P < .01). CONCLUSION: Intrauterine lidocaine is simple and effective for decreasing pain associated with the Pipelle endometrial biopsy. PMID- 10711542 TI - Establishing a new technique of laparoscopic pelvic and para-aortic lymphadenectomy. AB - OBJECTIVE: To assess the number of operations necessary to develop and standardize a laparoscopic approach to pelvic and para-aortic lymphadenectomy, with radicality and number of complications as quality markers. METHODS: Over 4 years, 108 women had complete laparoscopic pelvic and para-aortic lymphadenectomies combined with laparoscopy-assisted radical vaginal hysterectomies for primary therapy of cervical cancer. Complete data and videotapes were available for 99 women. Operating time and radicality for specific anatomic subareas were measured by review of video documentation and histologic lymph node counts. Intra- and postoperative complications were recorded prospectively. To analyze the progress of surgery, we compared two groups of women, one operated on at the beginning of our study (early group, subjects 6-35) and one operated on in the final period of the study (late group, subjects 79-108). RESULTS: The operating time for pelvic and para-aortic lymphadenectomy increased constantly. Comparing the early and late groups for para-aortic lymphadenectomy, there was an increase in mean operating time (34.8 versus 73.2 minutes; P < .001) and mean histologic lymph node yield (5.1 versus 10.6; P < .001). For pelvic lymphadenectomy, mean operating time increased slightly (60.7 versus 69.7 minutes; not significant) but mean histologic lymph node count decreased over time (24.3 versus 21.0; not significant). Retrospective evaluation of videotapes showed that the radicality of lymphadenectomy improved continuously in all evaluated subareas. CONCLUSION: Establishment of a protocol for para-aortic and pelvic lymphadenectomy took 100 operations. Video documentation was a more reliable indicator of progress in technical performance than were histologic lymph node counts. PMID- 10711543 TI - Amniotic fluid--soluble vascular endothelial growth factor receptor-1 in preeclampsia. AB - OBJECTIVE: To measure the levels of the soluble receptor for the potent angiogenic agent vascular endothelial growth factor (VEGF) in amniotic fluid (AF) in healthy and complicated pregnancies, and compare them with levels of erythropoietin, another factor upregulated by hypoxia. METHODS: We assessed amniotic fluid from the second (n = 35, gestational weeks 14-19) and third (n = 29) trimesters of healthy women, and from the third trimesters of preeclamptic (n = 22) and diabetic women with (n = 11) or without preeclampsia (n = 34) and from women with fetal growth restriction (FGR) (n = 14) for soluble VEGF receptor-1 (VEGFR-1) by enzyme-linked immunosorbent assay. RESULTS: In early normal pregnancy, AF-soluble VEGFR-1 levels were higher (median 22 ng/mL, range 2.3-29.5 ng/mL) than in the third trimester (median 13 ng/mL, range 0.5-32 ng/mL; P < .05). In preeclamptic women during the third trimester, levels were higher (median 20 ng/mL, range 10.5-37 ng/mL; P < .05) than healthy controls. The lowest third-trimester levels were in diabetic women (median 11 ng/mL, range 0.5-27 ng/mL). In women with preeclampsia and diabetes, AF-soluble VEGFR-1 levels remained lower (median 13, range 6-32 ng/mL; P < .05) than in women with preeclampsia alone. Amniotic fluid levels of soluble VEGFR-1 in women with FGR (median 19.5 ng/mL, range 5-40 ng/mL) did not statistically differ from those of controls. The AF levels of soluble VEGFR-1 did not correlate with those of erythropoietin. Soluble VEGFR-1 was clearly detectable (median 14 ng/mL, range 9 22 ng/mL) in culture media from placental biopsies (n = 20). CONCLUSION: Preeclampsia is associated with increased levels of soluble VEGFR-1, which are independent of erythropoietin, another hypoxia-inducible factor. PMID- 10711544 TI - Tocolysis with advanced cervical dilatation. AB - OBJECTIVE: To assess the feasibility and potential benefit of delaying delivery in women with advanced preterm labor. METHODS: Two hundred fifty-seven gravidas with intact membranes and preterm labor at cervical dilatations of at least 3 cm were studied. Women were excluded if they had premature rupture of membranes, gestational age less than 24 or more than 35.9 weeks, complete cervical dilatation, severe hemorrhage, chorioamnionitis, and triplets or higher-order gestations. Management consisted of tocolysis with intravenous magnesium sulfate as the primary agent, antenatal steroids, antibiotics, and amniocentesis. The primary endpoint was delay to delivery interval. Statistical analyses by cervical dilatation were performed using the Pearson chi2 test and a nonparametric test of trend. RESULTS: Eighty-one percent of pregnancies were referrals in utero from outlying hospitals. Delivery was delayed 24 hours or longer in 74% and beyond 48 hours in 60% of cases. Among 229 women who delivered at our center, 21% remained undelivered after 1 week. Evaluating delay as a function of cervical dilatation, trend analysis found a highly significant inverse relationship (P < .001). Among women dilated 5 cm, 46% delivered beyond 48 hours. Among those dilated 6 cm or more, 19% delivered beyond 48 hours. Mild pulmonary edema developed in five percent, and all responded promptly to medical interventions. Chorioamnionitis developed in eight percent. CONCLUSION: Delaying delivery 24-48 hours to allow antenatal steroid use or other interventions is possible in women with advanced preterm labor. PMID- 10711545 TI - Tissue concentrations of endothelial cell adhesion molecules in the lower uterine segment during term parturition. AB - OBJECTIVE: To determine the concentration of endothelial cell adhesion molecules in the lower uterine segment during parturition at term. METHODS: We analyzed protein extracts from the lower uterine segments of 38 women who had nonelective cesareans at term. We measured concentrations of intercellular adhesion molecule 1, endothelial leukocyte adhesion molecule-1, vascular cell adhesion molecule-1, and platelet endothelial cell adhesion molecule-1 by enzyme-linked immunosorbent assay. Subjects were grouped according to cervical dilatation (less than 2 cm, n = 10; 2 to less than 4 cm, n = 9; 4-6 cm, n = 9; more than 6 cm, n = 10) and duration of labor (up to 6 hours, n = 14; 6-12 hours, n = 10; 12-24 hours, n = 9; longer than 24 hours, n = 5) at the time of cesarean. RESULTS: The median concentration of intercellular adhesion molecule-1 increased significantly with increasing dilatation (from 2.24 ng/mg total protein at less than 2 cm to 6.73 ng/mg at 4-6 cm) and increasing duration of labor (from 2.53 ng/mg up to 6 hours to 5.90 ng/mg at 12-24 hours). However, this study did not have adequate statistical power to identify differences in concentrations of the other endothelial adhesion molecules. CONCLUSION: The results indicate that parturition at term is associated with expression of intercellular adhesion molecule-1. PMID- 10711546 TI - Risk factors for postcesarean surgical site infection. AB - OBJECTIVE: To determine postcesarean complications and identify independent risk factors for surgical site infection. METHODS: We studied a cohort of 969 women delivered by cesarean between May and August 1997. Infections were determined by examinations during ward rounds, reviews of laboratory results, and follow-up for 30 days after discharge. Risk factors were identified by multiple logistic regression. RESULTS: Surgical complications were rare. There were febrile morbidity and infection complications in 16.2% and 12.4% of subjects, respectively. Eighty-five subjects had 95 surgical site infections (9.8%), and seven risk factors were independently associated with infection. Risk factors included preoperative remote infection (adjusted odd ratio [OR] 16.5, 95% confidence interval [CI] 2.1, 128.3); chorioamnionitis (OR 10.6, 95% CI 2.1, 54.2); maternal preoperative condition (OR 5.3 for those with severe systemic disease [American Society of Anesthesiologists score > or =31, 95% CI 1.2, 24.0); preeclampsia (OR 2.3, 95% CI 1.1, 4.9); higher body mass index (OR 2.0 for every five-unit increment, 95% CI 1.3, 3.0); nulliparity (OR 1.8, 95% CI 1.1, 3.2); and increased surgical blood loss (OR 1.3 for every 100-mL increment, 95% CI 1.1, 1.5). CONCLUSION: Host susceptibility and existing infections were important predictors of surgical site infection after cesarean delivery. Further intervention should target this high-risk group to reduce the clinical effect of surgical site infection. PMID- 10711547 TI - Estrogens in intrahepatic cholestasis of pregnancy. AB - OBJECTIVE: To determine whether estrogen production and excretion are impaired in gravidas with intrahepatic cholestasis. METHODS: Plasma and urine samples were collected from 13 women from the United States and Chile at 35-38 weeks' gestation with mild (n = 9) or severe (n = 4) intrahepatic cholestasis of pregnancy. Urinary and plasma steroid levels from women with cholestasis were compared with levels from 27 normal pregnant women within the same gestational age range. Urinary concentrations of dehydroepiandrosterone (DHEA), estrone (E1), estradiol (E2), estriol (E3), estetrol, progesterone, and 16-hydroxy-pregnenolone were measured by gas chromatography mass spectrometry, and plasma concentrations of DHEA sulfate, progesterone, unconjugated E1, unconjugated E2, unconjugated E3, sulfated E3 derivatives, glucuronidated E3 derivatives, and total E3 were measured by radioimmunoassay. RESULTS: Compared with normal pregnant women, women with cholestasis had significantly lower plasma levels of estrogens and DHEA sulfate, the precursor to placental estrogen production synthesized by the fetal adrenal gland (Hotelling-Lawley trace = 0.81; F4,19 = 3.9; P = .02). The mean plasma DHEA sulfate, unconjugated E2, unconjugated E3, and total E3 concentrations were 0.271, 10.21, 9.80, and 99.53 ng/mL, respectively, in women with cholestasis compared with 0.802, 18.98, 16.28, and 145.07 ng/mL for controls. CONCLUSION: Fetal adrenal production of DHEA sulfate, and in response, downstream placental production of estrogens, was compromised by intrahepatic cholestasis of pregnancy. PMID- 10711548 TI - Effects of hospital policies based on 1996 group B streptococcal disease consensus guidelines. The Active Bacterial Core Surveillance Team. AB - OBJECTIVE: To determine whether the 1996 consensus guidelines for prevention of early-onset group B streptococcal disease developed by the Centers for Disease Control and Prevention, ACOG, and the American Academy of Pediatrics are affecting obstetric practice and disease occurrence. METHODS: Personnel in hospitals with obstetric services in seven surveillance areas completed surveys about their programs, patient populations, and group B streptococcal disease prevention policies. Survey results were linked to group B streptococcal disease cases identified by active surveillance in 1996 and 1997. An early onset case was defined as a case in which group B streptococci were isolated from a sterile site in the 1st 6 days of life. The number of cases in 1996 and 1997 were compared using a paired t test. Linear regression was used to assess hospital characteristics associated with group B streptococcal disease cases. RESULTS: Of 177 hospitals, 165 (93%) responded, and 96 (58%) of those had group B streptococcal disease prevention policies. Hospitals that established or revised their policies in 1996 had a lower mean number of cases in 1997 than in 1996 (0.58 versus 1.29, P = .006). Linear regression analysis, controlling for number of births, indicated that a hospital's having more black mothers and location in particular states were associated with more cases of disease. Citing the 1996 ACOG reference as the source for hospital group B streptococcal disease prevention policy was associated with fewer cases of group B streptococcal disease (P = .038). CONCLUSION: The publication and adoption of the guidelines were associated with decreasing occurrence of group B streptococcal disease. PMID- 10711549 TI - Interpregnancy interval and the risk of premature infants. AB - OBJECTIVE: Interpregnancy intervals are associated with the risk of low birth weight (LBW) infants, but the association between interpregnancy interval and prematurity is unknown. Our objective was to determine whether interpregnancy intervals were associated with the risk of premature infants and to define the degree of risk according to interpregnancy interval. METHODS: We analyzed 289,842 singleton infants born to parous Mexican-origin Hispanic and non-Hispanic white women in the United States who resided in the same county and delivered between January 1, 1991 and September 30, 1991. Interpregnancy interval was defined as the number of months between the previous live birth and conception of the index pregnancy. Multivariate logistic regression analysis was used to estimate odds ratios and 95% confidence intervals for the risk of interpregnancy interval on very premature (23-32 weeks), moderately premature (33-37 weeks), and term gestation (38-42 weeks). RESULTS: Nearly 37% of women had interpregnancy intervals less than 18 months, 45.5% of women had intervals of 18-59 months, and 17.6% of women had intervals over 59 months. After adjusting for confounding variables, women with intervals less than 18 months were 14-47% more likely to have very premature and moderately premature infants than women with intervals of 18-59 months. Women with intervals over 59 months were 12-45% more likely to have very premature and moderately premature infants than women with intervals of 18 59 months. CONCLUSION: Women with interpregnancy intervals from 18-59 months had the lowest risk of very premature and moderately premature infants. Further study is needed to define the mechanisms through which interpregnancy interval influences pregnancy outcome. PMID- 10711550 TI - Mastocytosis complicating pregnancy. AB - OBJECTIVE: To review the experience of women who conceived after developing mastocytosis and who were observed at the National Institutes of Health. METHODS: We reviewed our patient database for the years 1984-1998 to identify women with mastocytosis who had conceived. We then reviewed each woman's record, asked each woman to complete a questionnaire, and with permission wrote outside hospitals to obtain records of each labor and delivery. RESULTS: We identified eight women who had become pregnant. These women delivered a total of 11 live infants. In approximately a third of the pregnancies, patients experienced worsening of symptoms. They often used fewer medications during pregnancy because of safety concerns, and no greater incidence of adverse reactions was noted. Antihistamines were used most commonly, followed by oral prednisone. Medications used during delivery were well tolerated and included epidural analgesics. Neonates were generally healthy. None to date have developed urticaria pigmentosa or systemic mastocytosis. CONCLUSION: A subset of women with mastocytosis might have had exacerbated mastocytosis during and after pregnancy, but labor and delivery progressed normally. Infants were born generally healthy and were without mastocytosis. Thus there appears to be no absolute contraindication to pregnancy for women with mastocytosis, although women should be aware that the choice to have a child is not without some added risk. PMID- 10711551 TI - Direct medical cost of pelvic inflammatory disease and its sequelae: decreasing, but still substantial. AB - OBJECTIVE: To estimate direct medical costs and average lifetime cost per case of pelvic inflammatory disease (PID). METHODS: We estimated the direct medical expenditures for PID and its three major sequelae (chronic pelvic pain, ectopic pregnancy, and infertility) and determined the average lifetime cost of a case of PID and its sequelae. We analyzed 3 years of claims data of privately insured individuals to determine costs, and 3 years of national survey data to determine number of cases of PID, chronic pelvic pain, and ectopic pregnancy. We developed a probability model to determine the average lifetime cost of a case of PID. RESULTS: Direct medical expenditures for PID and its sequelae were estimated at $1.88 billion in 1998: $1.06 billion for PID, $166 million for chronic pelvic pain, $295 million for ectopic pregnancy, and $360 million for infertility associated with PID. The expected lifetime cost of a case of PID was $1167 in 1998 dollars. The majority of those costs ($843 per case) represent care for acute PID rather than diagnosis and treatment of sequelae. Approximately 73% of cases will not accrue costs beyond the treatment of acute PID. CONCLUSION: The direct medical cost of PID is still substantial. The majority of PID related costs are incurred in the treatment of acute PID. Because most PID-related costs arise in the first year from treatment of acute PID infection, strategies that prevent PID are likely to be cost-effective within a single year. PMID- 10711552 TI - Direct transport of progesterone from vagina to uterus. AB - OBJECTIVE: To compare progesterone concentrations in serum and endometrial tissue from hysterectomy specimens after vaginal or intramuscular (IM) administration of progesterone gel. METHODS: This was a randomized open study of 14 post-menopausal women undergoing transabdominal hysterectomies. Participants received either vaginal progesterone gel, 90 mg, or IM progesterone, 50 mg, at 8:00 AM and 8:00 PM on the day before surgery and at 6:00 AM on the day of surgery. Venous blood samples for progesterone measurement were collected at 8:00 AM on the day before surgery (baseline) and during surgery. After removal of the uterus, the endometrium was sampled from the anterior and posterior walls. Results were expressed as ratios of endometrial to serum progesterone concentrations x 100. RESULTS: Ratios of endometrial to serum progesterone concentrations were markedly higher in women who received vaginal progesterone (14.1 median, 8.5-59.4 range; 95% confidence interval [CI] 9.89, 38.79) compared with IM injections (1.2 median, 0.5-13.1 range; 95% CI -0.48, 7.39) (P < .005). CONCLUSION: Ratios of endometrial to serum progesterone concentrations were higher after vaginal administration of progesterone than after IM injections. Our findings in endometrial tissue specimens from hysterectomies excluded the possibility of contamination by progesterone that remained in the vagina. PMID- 10711553 TI - Cost-effectiveness of single-dose methotrexate compared with laparoscopic treatment of ectopic pregnancy. AB - OBJECTIVE: To evaluate the cost-effectiveness of treatment with intramuscular (IM) methotrexate compared with fallopian tube-sparing laparoscopy for small unruptured ectopic pregnancy. METHODS: A decision-analytic model accounting for varying resolution rates, complication rates, and cost estimates was built to compare the use of methotrexate with laparoscopy. Meta-analysis results of studies identified by a MEDLINE search for IM methotrexate resolution rates and tube-sparing laparoscopy resolution rates were used in model estimation. A similar process was used to generate model complication rates. Data on associated resource use were derived from established clinical guidelines. Estimates of 1998 costs incurred by provider organizations were calculated using data from a large managed care organization. RESULTS: The average methotrexate resolution rate among the studies included was 87% (range 75-90%). The average laparoscopy resolution rate was 91% (range 72-100%). Complication rates for methotrexate ranged from 0% to 22%, with an average of 10% for minor complications, and from 0% to 11% for serious complications, with an average of 7%. Complication rates for laparoscopy ranged from 0% to 8% for intraoperative complications, with an average of 2%, and from 0% to 15% for postoperative complications, with an average of 9%. Baseline model estimates indicated an average cost saving of more than $3000 per resolved ectopic pregnancy with methotrexate treatment compared with laparoscopy. Results of extensive sensitivity analyses supported the finding of a cost saving with methotrexate treatment. CONCLUSION: Single-dose methotrexate is a cost-saving, nonsurgical, fallopian tube-sparing treatment for ectopic pregnancy. PMID- 10711554 TI - Vaginal colonization by Candida in asymptomatic women with and without a history of recurrent vulvovaginal candidiasis. AB - OBJECTIVE: The asymptomatic carriage of Candida in the vagina of women with a history of recurrent vulvovaginal candidiasis was compared with that of women with no such history. METHODS: Vaginal swabs from 50 women with a history of recurrent vulvovaginal candidiasis and 45 women with one or fewer episodes of candidal vaginitis within the past 12 months were evaluated for Candida by wet mount/Gram stain, culture, and polymerase chain reaction (PCR). All women were asymptomatic for at least 30 days. RESULTS: Candida was identified in 28 women by PCR, in 14 women by culture, and in 13 women by wet mount/Gram stain. Candida was identified by PCR in a similar proportion of patients with previous recurrent vulvovaginal candidiasis (30%) and in controls (28.8%). However, Candida was identified by culture in more women with previous recurrent vulvovaginal candidiasis (22%) than in controls (6.6%, P = .04); it also was identified by wet mount/Gram stain in more women with recurrent vulvovaginal candidiasis (22%) than in controls (4.4%, P = .01). For the recurrent vulvovaginal candidiasis patients, culture and wet mount/Gram stain had a sensitivity of 66.6% compared with PCR. For the controls, the sensitivity of the two former assays relative to PCR was only 15.3%. CONCLUSION: Women with a history of recurrent vulvovaginal candidiasis have more easily detectable Candida in their vagina, even when asymptomatic, than do other women. A relative inefficiency in regulating the proliferation of Candida in the vagina may increase susceptibility to periodic symptomatic recurrences. PMID- 10711555 TI - A urinary control device for management of female stress incontinence. AB - OBJECTIVE: To examine the performance of a silicon urinary control device for nonsurgical management of women with genuine stress incontinence. METHODS: A 3 month prospective study involved 41 women with genuine stress incontinence. They completed urinary diaries of voiding, incontinence, and severity of incontinence on a 4-point scale over a week. Subjects were taught how to apply the device and used it as required from the second week. Visual analogue scales were used to record aspects of use (such as acceptability, comfort, and ease of application), and 2-hour perineal pad tests were completed at recruitment, after 2 weeks, and after 3 months. Data were compared by Mann-Whitney U test, or Wilcoxon test. RESULTS: Ten women (24.4%) declined to participate and six (14.6%) withdrew before 2 weeks. Ten (24.4%) failed to attend for 2-week follow-up and 11 (26.8%) did not continue for 3 months. Two (4.9%) did not attend 3-month follow-up. Only two women (4.9%) completed the study. There was no difference in pad test results or in results from voiding diaries. CONCLUSION: The urinary incontinence device had low acceptability and was ineffective, and we cannot recommend it for nonsurgical management of genuine stress incontinence. PMID- 10711556 TI - Gynecologists' training, knowledge, and experiences in genetics: a survey. AB - OBJECTIVE: To explore gynecologists' knowledge, training, and practice experience with genetic screening and DNA-based testing. METHODS: A questionnaire survey was sent to 1,248 ACOG Fellows, of whom 564 (45%) responded. One hundred thirty-four respondents (24%) reported that they do not order DNA-based tests or take family histories to screen for heritable diseases or disorders. Results from the 428 respondents who provide genetic screening services are reported. RESULTS: Most physicians (90%) knew that genetic tests are most informative when used in conjunction with family histories. Gynecologists gave more correct responses regarding genetic testing for breast and ovarian cancers than for colon cancer and other adult-onset diseases. Sixty-five percent of the respondents had not received formal training in DNA-based testing in gynecologic practice. Older physicians were less likely to have had training. Younger physicians generally gave more correct responses on the knowledge portion of the survey (r = -.165, P < .01). Physicians who had formal training in genetics gave more correct answers. Physicians who order DNA-based tests scored higher than those who do not and had no formal training, but not higher than those who had formal training and do not order DNA-based tests. CONCLUSION: Gynecologists were more knowledgeable about genetic issues pertaining to breast and ovarian cancer than to other cancers or certain adult-onset disorders. Training appeared to increase knowledge. Increased training and affiliation with genetic specialists and others could improve gynecologists' ability to use genetic screening in clinical practice. PMID- 10711557 TI - Early motor development of breech- and cephalic-presenting infants. AB - OBJECTIVE: This study was conducted to determine whether breech-presenting infants have a different pattern of early neuromotor development than cephalic presenting infants--regardless of mode of delivery-thus explaining both the failure to assume cephalic version at the end of gestation and the higher rates of childhood motor impairments associated with breech presentation. METHODS: Ninety morphologically normal, term, breech-presenting singletons with birthweights greater than 2,500 g were paired with a similar cephalic-presenting infant, matched for gender and mode of delivery (n = 180; 100 delivered abdominally and 80 delivered vaginally). Data on neurological status (Neurological Assessment of the Preterm and Full-term Newborn Infant) and motor performance (Alberta Infant Motor Scale, Peabody Developmental Motor Scales, and age of walking) were collected prospectively over the first 18 months of life. This study was designed with a power of .80 to detect a "medium" effect size for motor development using the Alberta Infant Motor Scale. The data were analyzed using analysis of variance techniques. RESULTS: Breech-presenting infants had minor transient differences compared with cephalic-presenting infants. First, they had more open popliteal angles at birth (P < .001). Second, they had significantly lower motor scores at 6 weeks than the normative sample (P < .001). At 18 months, three infants were diagnosed with neurological problems, all of whom were delivered electively in the cesarean-breech group. CONCLUSION: As a group, breech-presenting infants do not have a persistent, inherently different pattern of motor development than cephalic-presenting infants. Mode of delivery did not explain the excess neuromotor impairment detected in the subgroup of breech infants. PMID- 10711558 TI - Persistent intrahepatic right umbilical vein in the fetus: a benign anatomic variant. AB - OBJECTIVE: To evaluate outcomes of fetuses with antepartum sonographic diagnoses of persistent intrahepatic right umbilical veins. METHODS: A detailed fetal sonographic examination was done in 30,240 consecutive pregnancies at 14-26 weeks' gestation. High- and low-risk pregnancies were included and persistent right umbilical veins specifically were recorded. RESULTS: Sixty-nine fetuses had persistent intrahepatic right umbilical veins, of which 60 had no additional sonographic abnormalities, four had transient nuchal findings, and four had minor anomalies or anatomic variants. Only one of the 69 fetuses had a major anomaly (diaphragmatic hernia), and died after surgery. The remaining 68 fetuses were normal and healthy after birth. CONCLUSION: Persistent intrahepatic right umbilical vein is a fetal anatomic variant that is not rare and usually associated with a favorable outcome. PMID- 10711559 TI - Isolated clubfoot diagnosed prenatally: is karyotyping indicated? AB - OBJECTIVE: To evaluate the appropriateness of fetal karyotyping after prenatal sonographic diagnosis of isolated unilateral or bilateral clubfoot. METHODS: We retrospectively reviewed a database of fetal abnormalities diagnosed by ultrasound at a single tertiary referral center from July 1994 to March 1999 for cases of unilateral or bilateral clubfoot. Fetuses who had additional anomalies diagnosed prenatally, after targeted sonographic fetal anatomy surveys, were excluded. Outcome results included fetal karyotype diagnosed by amniocentesis, or newborn physical examination by a pediatrician. RESULTS: During the 5-year period, 5,731 fetal abnormalities were diagnosed from more than 27,000 targeted prenatal ultrasound examinations. There were 51 cases of isolated clubfoot. The mean maternal age at diagnosis was 30.5 years. The mean gestational age at diagnosis was 21.6 weeks. Twenty-three of the women (45%) were at increased risk of fetal aneuploidy, on the basis of advanced maternal age or abnormal maternal serum screening. Six women (12%) had positive family histories of clubfoot; however, no cases of aneuploidy were found by fetal karyotype evaluation or newborn physical examination. All cases of clubfoot diagnosed prenatally were confirmed at newborn physical examination, and no additional malformations were detected. CONCLUSION: After prenatal diagnosis of isolated unilateral or bilateral clubfoot, there appeared to be no indication to offer karyotyping, provided that a detailed sonographic fetal anatomy survey was normal and there were no additional indications for invasive prenatal diagnoses. PMID- 10711560 TI - Thrombocytopenia in term infants: a population-based study. AB - OBJECTIVE: To assess the prevalence and causes of thrombocytopenia among full term infants. METHODS: We conducted a 1-year, population-based surveillance study involving all full-term infants (at least 37 weeks' gestation) born to native Finnish women in Helsinki. In cases of thrombocytopenia (cord platelet count less than 150 x 10(9)/L) clinical risk factors were evaluated and immunologic studies were performed on both parents and on the infant; 95% confidence intervals (CIs) were calculated on the basis of binomial distribution. RESULTS: Platelet counts were done in cord blood from 4,489 infants, 84.9% of the study population. Eighty nine infants had platelet counts below 150 x 10(9)/L (2.0%; 95% CI 1.5, 2.3) in cord blood and 11 were less than 50 x 10(9)/L (0.24%; 95% CI 0.10, 0.38). All causes of clinically important thrombocytopenia, those presenting with bleeding and requiring treatment, were related to fetomaternal alloimmune thrombocytopenia. The incidence of severe alloimmune thrombocytopenia was one in 1500 live births and one in 900 of all thrombocytopenia. An immunologic mechanism was involved in ten of 65 (15.4%; 95% CI 6.6, 24.2) infants studied and in four of 15 (26.7%; 95% CI 4.3, 49.1) cases of severe thrombocytopenia. CONCLUSION: Immunologic studies should be considered in all cases of severe neonatal thrombocytopenia for careful monitoring and prevention of potentially severe complications in subsequent pregnancies. PMID- 10711561 TI - Maternal serum ferritin and fetal growth. AB - OBJECTIVE: To determine the relationship between maternal serum ferritin and concentrations and specific types of fetal growth restriction (FGR). METHODS: Serum ferritin concentrations were measured at approximately 25 and 36 weeks' gestation in 480 multiparas with singleton fetuses who participated in a study of risk factors for repeated FGR. Asymmetric FGR was defined by low birth weight for gestational age criteria and a ponderal index less than 2.32, and symmetric FGR was defined by the same birth weight for gestational age criteria and a ponderal index of at least 2.32. RESULTS: Among 480 infants, 370 were appropriate for gestational age (AGA), 58 had asymmetric FGR, and 52 had symmetric FGR. Higher ferritin concentrations were associated with black race, maternal age 25 years or older, and smoking. Mothers of asymmetric-FGR infants had higher mean ferritin levels than mothers of AGA infants at 25 weeks' (38.0 versus 20.2 microg/L, P < .01) and 36 weeks' gestation (21.0 versus 13.3 microg/L, P < .01), whereas mothers of symmetric-FGR infants had significantly lower ferritin levels at 36 weeks (8.3 microg/L). For mothers with serum ferritin levels of at least 26 microg/L (highest quartile at 25 weeks), the adjusted odds ratio (OR) for asymmetric-FGR infants was 3.4, 95% confidence interval (CI) 1.6, 7.2. There was a similar association between the highest quartile of serum ferritin at 36 weeks (at least 20 microg/L) and asymmetric FGR (adjusted OR 2.7, 95% CI 1.3, 5.8). Women with serum ferritin levels less than 3 microg/L (lowest quartile at 36 weeks) had an adjusted OR for symmetric-FGR infants of 2.2, 95% CI 1.01, 4.6. CONCLUSION: High maternal serum ferritin levels are associated with asymmetric FGR, whereas low serum ferritin levels are associated with symmetric FGR. PMID- 10711562 TI - Preterm delivery in mice with renal abscess. AB - OBJECTIVE: Our purpose was to develop a mouse model of renal abscess to study the effect of extrauterine infection on preterm delivery. METHODS: Escherichia coli or sterile medium was injected into the left kidney of 70 pregnant mice that had completed approximately 75% of gestation. Preterm delivery rates were recorded for various inocula. Kidney specimens were obtained and examined grossly and histologically for abscess formation. RESULTS: Thirty-one of 51 animals (60.8%) infected with 1 x 10(5)-9 x 10(6) bacteria and none of 19 uninfected animals delivered prematurely (P < .001). Renal abscess was induced in 100% of mice receiving bacterial inoculation but in none receiving sterile medium. CONCLUSION: Kidney injection provides a reliable method for inducing renal abscess in pregnant mice. Renal abscess induces preterm delivery at a stable rate across a wide range of bacterial inocula. This model of extrauterine infection may be particularly useful in investigations of infection-induced preterm delivery. PMID- 10711563 TI - Magnetic resonance spectroscopy to detect lecithin in amniotic fluid and fetal lung. AB - BACKGROUND: Proton magnetic resonance spectroscopy is a noninvasive technique that detects molecules within a specified region in vivo. Lecithin, the major component of surfactant, has a characteristic magnetic resonance signal, but to our knowledge, it has never been reported in fetal lung or amniotic fluid (AF). The objective of this study was to characterize the lecithin signal in utero, which could lead to a noninvasive fetal lung maturity test. METHOD: Human fetal lung and AF pockets can be identified and studied with magnetic resonance spectroscopy with the use of a 1.5-tesla Vision whole-body magnetic resonance scanner (Siemens Medical Systems; Erlangen, Germany). Spectroscopy data are collected with a single-voxel-point-resolved spectroscopy sequence. After identification of fetal anatomy with the use of scout magnetic resonance images, magnetic resonance spectroscopy of human fetal lung and AF identifies a lecithin peak. EXPERIENCE: Three healthy gravidas near term were studied and lecithin peaks were identified in all. CONCLUSION: Lecithin can be identified in vivo with the use of volume-selected proton magnetic resonance spectroscopy. Patient comfort and extremely short scan times suggest that refined magnetic resonance spectroscopy might be a safe, quick, and comfortable test of fetal lung maturity. PMID- 10711564 TI - Videoteleconferencing for administration of a multisite obstetrics and gynecology core clerkship. AB - Recent changes and trends in health care delivery have required medical schools to use multiple sites to obtain adequate patient exposure for their students. Decentralization of clinical undergraduate medical education may lead to a lack of continuity in curricula, evaluation, and feedback. We describe the use of interactive videoteleconferencing as a tool to link and improve a multi-site undergraduate core clerkship in obstetrics and gynecology. The Uniformed Services University of the Health Sciences, Bethesda, Maryland, currently utilizes five geographically separate sites for its 6-week core clerkship in obstetrics and gynecology. The site coordinators, clerkship director, and administrative personnel from the parent institution meet approximately 3 weeks after the completion of each core clerkship for live, real-time, and interactive broadcast to complete student evaluations, review curricula, and discuss problems with current students and other pertinent educational issues. Videoteleconferencing provides a mechanism to ensure consistency in curriculum and student evaluations and provides administrative support to distant sites. Furthermore, it enables site coordinators to keep the clerkship director abreast of students and clerkship issues. PMID- 10711565 TI - Preventing perineal trauma during childbirth: a systematic review. AB - OBJECTIVE: To review systematically techniques proposed to prevent perineal trauma during childbirth and meta-analyze the evidence of their efficacy from randomized controlled trials. DATA SOURCES: MEDLINE (1966-1999), the Cochrane Library (1999 Issue 1), and the Cochrane Collaboration: Pregnancy and Childbirth Database (1995); and reference lists from articles identified. Search terms included childbirth or pregnancy or delivery, and perineum, episiotomy, perineal massage, obstetric forceps, vacuum extraction, labor stage-second. No language or study-type constraints were imposed. STUDY SELECTION: Randomized controlled trials (RCTs) of interventions affecting perineal trauma were reviewed. If no RCTs were available, nonrandomized research designs such as cohort studies were included. Studies were selected by examination of titles and abstracts of more than 1,500 articles, followed by analysis of the methods sections of studies that appeared to be RCTs. INTEGRATION AND RESULTS: Eligible studies used random or quasirandom allocation of an intervention of interest and reported perineal outcomes. Further exclusions were based on failure to report results by intention to treat, or incomplete or internally inconsistent reporting of perineal outcomes. Final selection of studies and data extraction was by consensus of the first two authors. Data from trials that evaluated similar interventions were combined using a random effects model to determine weighted estimate of risk difference and number needed to treat. Effects of sensitivity analysis and quality scoring were examined. Results indicated good evidence that avoiding episiotomy decreased perineal trauma (absolute risk difference -0.23, 95% confidence interval [CI] -0.35, -0.11). In nulliparas, perineal massage during the weeks before giving birth also protected against perineal trauma (risk difference -0.08, CI -0.12, -0.04). Vacuum extraction (risk difference -0.06, CI 0.10, -0.02) and spontaneous birth (-0.11, 95% CI -0.18, -0.04) caused less anal sphincter trauma than forceps delivery. The mother's position during the second stage has little influence on perineal trauma (supported upright versus recumbent: risk difference 0.02, 95% CI -0.05, 0.09). CONCLUSION: Factors shown to increase perineal integrity include avoiding episiotomy, spontaneous or vacuum assisted rather than forceps birth, and in nulliparas, perineal massage during the weeks before childbirth. Second-stage position has little effect. Further information on techniques to protect the perineum during spontaneous delivery is sorely needed. PMID- 10711566 TI - Randomized, double-masked comparison of oxytocin dosage in induction and augmentation of labor. PMID- 10711567 TI - Randomized, double-masked comparison of oxytocin dosage in induction and augmentation of labor. PMID- 10711568 TI - The great douching debate: to douche, or not to douche. PMID- 10711569 TI - Cervical ripening and labor induction with a controlled-release dinoprostone vaginal insert: a meta-analysis. PMID- 10711570 TI - Diagnostic accuracy of ultrasound above and below the beta-hCG discriminatory zone. PMID- 10711571 TI - Can diabetic neuropathy be prevented by angiotensin-converting enzyme inhibitors? AB - The incidence of diabetes and its complications is increasing to staggering proportions. Presently the WHO estimates an overall prevalence of 130 million, but by 2025 there will be 300 million individuals with diabetes mellitus. The incidence of diabetic neuropathy approaches 50% in most diabetic populations; there is no treatment, and its consequences in the form of foot ulceration and amputation are financially punishing for health care providers. Attempts to develop treatments have faltered for want of an understanding of the aetiology of diabetic neuropathy. As a consequence, 1999 saw the demise of two further compounds: recombinant growth factor by Roche-Genentech and the aldose reductase inhibitor zopolrestat, by Pfizer, both had reached phase III clinical trials. They joined an impressive list of at least 30 other compounds which have reached phase III clinical trials and failed to establish efficacy. The need to establish a viable treatment for human diabetic neuropathy is absolutely paramount. To provide a rational answer as to whether angiotensin-converting enzyme (ACE) inhibitors can prevent human diabetic neuropathy, two major issues need addressing: 1) Does vascular dysfunction cause human diabetic neuropathy? 2) Can ACE inhibitors ameliorate diabetic vascular dysfunction and hence neuropathy? Epidemiological studies support a strong association between neuropathy, retinopathy and nephropathy. Microangiopathy is deemed as the root cause of both nephropathy, and retinopathy and mounting evidence provides support for a vascular basis of diabetic neuropathy. ACE inhibitors appear to correct many of the abnormalities associated with the vascular dysfunction found in diabetes. Thus effective ACE inhibition impacts very positively on cardiovascular outcomes in patients with ischaemic heart disease, particularly in diabetic patients. ACE inhibition also prevents the development and progression of incipient and established diabetic nephropathy and delays progression of background retinopathy. Quinapril improves measures of diabetic autonomic neuropathy. Our recent study has demonstrated a significant improvement in peripheral neuropathy following 12 months of treatment with the ACE inhibitor trandolapril. PMID- 10711572 TI - Age-related cognitive decline, mild cognitive impairment or preclinical Alzheimer's disease? AB - With the promising development of effective treatment, significant improvement in the very early diagnosis of Alzheimer's disease (AD) is required. There is vast agreement that a decline in memory, especially in verbal episodic memory, is the earliest and perhaps the most sensitive sign of incipient AD at the preclinical stage. However, this review offers evidence that impairment in episodic memory can be observed in normal elderly people as well as in aged subjects with mild cognitive impairment (MCI), a large proportion of whom will, however, not convert to dementia. Quantitative measurement of atrophy and brain activation in the hippocampal-parahippocampal formation by using structural and functional magnetic resonance imaging may help to distinguish the MCI decliners from the nondecliners. Cerebrospinal fluid levels of tau protein and Abet1-42 peptide, together with the presence of an apolipoprotein (apo)E epsilon4 allele may also increase our confidence in the early positive diagnosis of AD. This review concludes, however, that while adequate for constituting groups of patients in a research perspective, the extensive diagnostic procedure based on specific cognitive testing, neuroimaging and biological investigations is still out of reach for the practitioner. PMID- 10711573 TI - Molecular basis of androgen receptor diseases. AB - androgens act through a single intracellular androgen receptor (AR) which is encoded by a single-copy gene in the X chromosome. Disruption of the AR by genetic mutation results in complete androgen insensitivity syndrome (CAIS) and the female phenotype in otherwise healthy 46XY individuals. Although CAIS is the best known phenotype, recent studies from our laboratory and elsewhere show that malfunction of the AR is associated with many androgen-regulated diseases or conditions that cross traditional clinical disciplines ranging from paediatrics (ambiguous genitalia), gynaecology (primary amenorrhoea), urology (prostate cancer), neurology (spinal bulbar muscular atrophy), reproductive medicine (male infertility), orthopedics (rheumatoid arthritis), oncology (breast cancer) and dermatology (hirsutism, baldness and acne). Of particular interest are the roles that polymorphic CAG trinucleotide repeat tracts and subtle mutations in the AR ligand-binding domain have in the aetiology of male infertility and prostate cancer, two conditions affecting large numbers of patients. Novel mechanisms of pathogenesis have been uncovered in these cases, and they involve defective protein-protein interactions with coregulator molecules such as TIF2 (transcriptional intermediary factor 2). Knowledge of the critical role that the AR plays in the pathogenesis of these diverse conditions has led to improved diagnostic methods and successful therapy. PMID- 10711574 TI - Medical significance of Caenorhabditis elegans. AB - Caenorhabditis elegans is now the model organism of choice for a growing number of researchers. A combination of its apparent simplicity, exquisite genetics, the existence of a full molecular toolkit and a complete genome sequence makes it ideal for rapid and effective study of gene function. A survey of the C. elegans genome indicates that this 'simple' worm contains many genes with a high degree of similarity to human disease genes. For many human disease genes it has proven, and will continue to prove, difficult to elucidate their function by direct study. In such cases simpler model organisms may prove to be a more productive starting point. The basic function of a human disease gene may be studied in the background of C. elegans, in which the most important interactions are likely to be conserved, providing an insight into disease process in humans. Here we consider the significance of this modality for human disease processes and discuss how C. elegans may, in some cases, be ideal in the study of the function of human disease genes and act as a model for groups of parasitic nematodes that have a severe impact on world health. PMID- 10711575 TI - Novel cancer therapies: more efficacy, less toxicity and improved organ preservation. AB - Novel approaches to the treatment of cancer include techniques such as gene therapy, antiangiogenic therapy, monoclonal antibodies either alone or linked with radioactive isotopes or cytotoxins, cancer immunotherapy and vaccines, oligonucleotides and antisense technologies as well as anticancer drugs targeting single metabolic processes, enzymes or oncoproteins. However, substantial improvements are also being made in more conventional cancer treatment modalities. These comprise radiotherapy given concomitantly with chemotherapy, which appears to improve treatment results in a number of common types of human cancer. Other important advances include conformal and intensity-modulated radiation therapy, which may allow for higher target doses with little or no increase in toxicity. Stereotactic radiation therapy for extracranial targets is also being developed, as well as biologically targeted radiation therapy, in which targeting is based on metabolic pathways or carrier molecules, such as boronated compounds in boron neutron capture therapy or monoclonal antibodies in radioimmunotherapy. Sentinel node biopsy and neoadjuvant chemotherapy for breast cancer represent advances in surgery and cancer chemotherapy, which may also allow for a greater chance for organ and tissue preservation without a loss in treatment efficacy. PMID- 10711576 TI - Organ-conserving approaches to muscle-invasive bladder cancer: future alternatives to radical cystectomy. AB - In the USA radical surgery remains the golden standard for invasive bladder cancer. Yet in most other areas of surgical oncology the trend of the 1990s has been towards organ conservation with chemoradiation with or without limited local surgery. Patients with breast, oesophageal, anal, lung and larynx cancer are routinely offered conservative therapies as valid options in the management of their diseases but bladder stands apart from the crowd. Evidence is presented here to show that this need not be the case. Four older randomized trials failed to show a survival advantage when immediate cystectomy was compared with radiation followed by salvage cystectomy, if required. Five and 8-year survival rates for clinically staged patients treated by transurethral resection and chemoradiation (trimodality therapy) in several modern, large and mature series show survival rates comparable to those reported in contemporary radical cystectomy series. Eighty per cent of those alive 5 years after chemoradiation still retain their native bladder. Although superficial relapse occurs in 20% of cases, it remains responsive to BCG (Bacilles bilie de Calmette-Guerin) in the manner of de novo superficial disease. Quality-of-life studies show that the retained bladder functions well. At the Massachusetts General Hospital and in the multicentre prospective trials, less than 1% of patients needed cystectomy for bladder morbidity. It is of note that continent diversions may be performed as salvage after contemporary radiation therapy. Trimodality therapy is a novel and contemporary approach that owes little to the radiation treatment offered in the 1970s. While it will never entirely take the place of radical cystectomy, it should be offered as a reasonable alternative to patients with a new diagnosis of bladder cancer. This multidisciplinary approach will allow uro-oncology to keep in step with the oncological vanguard. PMID- 10711577 TI - Neoadjuvant chemotherapy for breast cancer. AB - Primary or neoadjuvant chemotherapy in early breast cancer offers the chance to use the tumour as an in vivo measure of response, with the additional possibility of downstaging and avoidance of mastectomy. Tumour response to preoperative chemotherapy correlates with the outcome and could be a surrogate for evaluating the effect of chemotherapy on micrometastases. Randomized studies have shown that preoperative chemotherapy is as effective as postoperative chemotherapy, but there has not been a significant increase in the disease-free survival or overall survival in the groups studied. The overall response rates reported have varied between 60% and 100% with complete clinical responses from 10% to almost 50%, avoiding mastectomy in most cases. Clinical responders have a better prognosis than nonresponders; pathological complete remissions at present offer the best prediction of good long-term outcome, but occur in less than 20% of patients. Biological predictors reflecting changes in apoptosis and/or proliferation may in the future offer the best surrogate markers for long-term outcome, and trials have recently begun in this area. PMID- 10711578 TI - Role of sentinel lymph node dissection in breast cancer. AB - The sentinel node concept is valid for penile cancer, melanoma, breast cancer and is probably also applicable to other solid malignancies. Sentinel nodes are the one or two initial nodes in the regional nodal drainage basin encountered by the lymphatic effluent from a tumour, which can be identified with an injection of vital dye or other lymphogogue. Sentinel lymph node dissection (SLND), a minimally invasive procedure with negligible morbidity, has therefore been utilized as an alternative to complete axillary lymph node dissection (ALND) for staging breast cancer. Examination of sentinel nodes provides a focused histopathological assessment of tissue most likely to harbour metastases, providing enhanced staging accuracy with a low false-negative rate. Tumour-free sentinel nodes are predictive of a tumour-free axilla, thereby allowing for the possibility of SLND without ALND and sparing patients the morbidity of ALND. Most of the experience from SLND has been obtained for axillary sentinel nodes. However, sentinel nodes have been identified in nonaxillary sites, such as the internal mammary nodes, but data on SLND for these regions is scarce. The ultimate role of SLND in breast cancer, which may be to identify sentinel-node negative patients or even those with sentinel node metastases who can safely avoid ALND without sacrificing regional control and possibly gain a therapeutic benefit, cannot be defined before we have the results of large trials that are currently in progress. PMID- 10711579 TI - Conformal radiotherapy for prostate cancer. AB - This review presents the contribution conformal treatment has made to the external beam treatment of prostate cancer drawing on 10 years of experience with three-dimensional conformal radiation treatment in 1500 patients with prostate cancer at our institution. Major contributions from other institutions and the Radiation Therapy Oncology Group (RTOG) clinical trials group are also noted. Patient immobilization, computed tomography (CT) scan identification of the target in three dimensions and beams-eye-view conformal treatment portals are critical to the process. Higher doses of radiation are associated with a marked increase in cure rates as the dose-response curve is extremely steep. The advantage given by the high dose depending on prognostic group ranges from 14% to 35% in 5-year cures. Conformal techniques protect normal tissues so that late complications are rare and in fact fewer than those observed with standard radiation technique at standard radiation doses. Educational efforts are underway in the USA to assist practising radiation oncologists to deliver the required radiation doses of 75-80 Gy safely. Conformal therapy cures more prostate cancers causing fewer complications than standard treatment technique and standard dose. It must therefore be disseminated into the practice of radiation oncology worldwide. PMID- 10711580 TI - Linear accelerator radiosurgery in the management of brain tumours. AB - Radiosurgery is an increasingly popular method for treating a variety of intracranial tumours. A great deal of treatment data has been accumulated suggesting that radiosurgery may be the treatment of choice for small acoustic schwannomas. Moreover, radiosurgery promises excellent tumour control and minimal risk in the treatment of small meningiomas in risky surgical locations such as the cavernous sinus. Radiosurgery offers superior local control rates for many metastatic neoplasms and has promise as an adjuvant 'boost' technique in certain malignant gliomas. This article presents a brief description of the linear accelerator, LINAC, radiosurgical technique, followed by a review of the more common applications of stereotactic radiosurgery in the treatment of intracranial neoplastic disease. PMID- 10711581 TI - Boron neutron capture therapy for malignant gliomas. AB - Boron neutron capture therapy (BNCT) represents a promising modality for a relatively selective radiation dose delivery to the tumour tissue. Boron-10 nuclei capture slow 'thermal' neutrons preferentially and, upon capture, promptly undergo 10B(n,alpha)7Li reaction. The ionization tracks of energetic and heavy lithium and helium ions resulting from this reaction are only about one cell diameter in length (approximately 14 microm). Because of their high linear energy transfer (LET) these ions have a high relative biological effectiveness (RBE) for controlling tumour growth. The key to effective BNCT of tumours, such as glioblastoma multiforme (GBM), is the preferential accumulation of boron-10 in the tumour, including the infiltrating GBM cells, as compared with that in the vital structures of the normal brain. Provided that a sufficiently high tumour boron-10 concentration (approximately 10(9) boron-10 atoms/cell) and an adequate thermal neutron fluence (approximately 10(12) neutrons/cm2) are achieved, it is the ratio of the boron-10 concentration in tumour cells to that in the normal brain cells that will largely determine the therapeutic gain of BNCT. PMID- 10711582 TI - Using information, motivational enhancement, and skills training to reduce the risk of HIV infection for low-income urban women: a second randomized clinical trial. AB - This randomized clinical trial evaluated an HIV-risk reduction (HIV-RR) intervention based on the information-motivation-behavioral skills model. At baseline, 102 women (M age = 29 years; 88% African American) completed a survey regarding HIV-related knowledge, risk perceptions, behavioral intentions, and risk behavior. Participants were then assigned to either the HIV-RR intervention or a health-promotion control group. Postintervention and follow-up data indicated that women in the HIV-RR program enhanced their knowledge and strengthened their risk reduction intentions relative to controls. Moreover, HIV RR women who expressed "imperfect" intentions also increased their condom use, talked more with partners about condom use and HIV testing, and were more likely to have refused unprotected sex. PMID- 10711583 TI - Cognitive-behavioral stress management increases free testosterone and decreases psychological distress in HIV-seropositive men. AB - The effects of a 10-week group-based cognitive-behavioral stress management (CBSM) intervention on psychological distress and plasma free testosterone in symptomatic, HIV-seropositive men were examined. Participants were randomized to either CBSM (n = 42) or a wait-list control group (n = 23). Men in the CBSM intervention showed significant increases in testosterone, whereas control participants showed significant decreases. Those participating in CBSM had significant distress reductions, whereas controls showed no such change. Alterations in free testosterone were inversely related to changes in distress states over time, independent of any changes in cortisol. These findings demonstrate that a short-term CBSM intervention increases free testosterone levels among symptomatic, HIV-seropositive men, and alterations in free testosterone are associated with changes in psychological distress observed during CBSM. PMID- 10711584 TI - Stress, depressive symptoms, and smoking cessation among pregnant women. AB - Perceived stress and depressive symptoms were examined as correlates and predictors of smoking cessation during pregnancy in a sample of 819 pregnant smokers (454 baseline smokers and 365 baseline quitters). Women who quit early in pregnancy had lower levels of stress and depressive symptoms than baseline smokers. Adjusting for level of addiction and other demographic factors related to stress and depressive symptoms eliminated the significant association between depressive symptoms and smoking cessation. Lower levels of stress and depressive symptoms were not predictive of cessation in later pregnancy. Prenatal healthcare providers should continue to assess level of addiction and provide targeted intensive cessation interventions. Interventions that reduce stress and depression may also be of benefit to women who are continuing smokers in early pregnancy. PMID- 10711585 TI - A longitudinal model of social contact, social support, depression, and alcohol use. AB - The longitudinal relations among contact with one's social network (social contact), perceived social support, depression, and alcohol use were examined. An integrative model was developed from affect regulation theory and theories of social support and dysfunctional drinking. Data were obtained from a random sample of 1,192 adults. The 3-wave panel model was tested using structural equation modeling analysis. Results revealed that (a) social contact was positively related to perceived social support; (b) perceived social support was, in turn, negatively related to depression; and (c) depression was, in turn, positively related to alcohol use for 1 of 2 longitudinal lags. There was partial support for the feedback hypothesis that increased alcohol use leads to decreased contact with family and friends. Although the results generally supported the authors' hypotheses, the significant coefficients in the model were generally small in size. PMID- 10711586 TI - Time-of-day effects on response of natural killer cells to acute stress in men and women. AB - Diurnal influences on natural killer (NK) cell changes to acute stress were assessed in 21 men and 21 women assigned to either an acute stress (mental arithmetic) or control task condition. Sessions began at either 8 a.m. or 2 p.m. Number of NK (CD3-CD56+) cells and NK activity were measured at baseline, during the 5-min task, and 60 and 90 min after the task. Both morning and afternoon stress participants had elevated NK cell numbers during the task. After the task, number of NK cells decreased in morning stress participants but remained significantly above baseline levels 60 and 90 min posttask. NK cell numbers in afternoon stress participants decreased to below baseline levels 60 and 90 min after the task. Changes in NK activity were driven primarily by diurnal influences. NK activity increased in all morning participants and stayed increased 60 and 90 min posttask. NK activity of all afternoon participants also increased during the task but dropped below baseline 60 and 90 min later. Greater increases in NK levels and activity during the task were associated with greater heart rate changes. PMID- 10711587 TI - Task demands and the pressures of everyday life: associations between cardiovascular reactivity and work blood pressure and heart rate. AB - Associations between cardiovascular stress reactivity and blood pressure and heart rate recorded in everyday life were hypothesized to depend on the stressfulness of the ambulatory monitoring period relative to standardized tasks and on activity levels at the time of measurement. One hundred two female and 60 male school teachers carried out high- and low-demand tasks under standardized conditions and ambulatory monitoring during the working day. Stress ratings during the day were close to those recorded during the low-demand task. Reactions to the low-demand task were significant predictors of ambulatory blood pressure and heart rate independent of baseline, age, gender, and body mass. Associations were more consistent for ambulatory recordings taken when participants were seated than when they were standing and when the ambulatory monitoring day was considered to be as stressful as usual or more stressful than usual, and not less stressful than usual. Laboratory-field associations of cardiovascular activity depend in part on the congruence of stressfulness and physical activity level in the 2 situations. PMID- 10711588 TI - Constructive anger verbal behavior predicts blood pressure in a population-based sample. AB - The creation of an observational Constructive Anger Behavior-Verbal Style Scale (CAB-V) and its relation to resting blood pressure (BP) in an age- and sex stratified, population-based sample is examined. Participants (N = 1,862) provided hypertension risk factor information, had resting BP assessed multiple times, and completed a videotaped interview, which was later coded for CAB-V and Hostile Style. High CAB-V scores remained a significant predictor of lower resting BP when controlling for the effects of standard hypertension risk factors (age, sex, body mass index, physical activity, alcohol use, smoking status, parental myocardial infarction history, education, and diabetic status) and psychosocial measures (anxiety, depression, hostility, social support, and Hostile Style). This relation also remained when excluding known hypertensive persons. Results suggest that constructive anger expression may have an independent beneficial association with resting BP. PMID- 10711589 TI - Perceived probability, perceived severity, and health-protective behavior. AB - It seems obvious that 2 key attributes of health hazards, their perceived probability and perceived severity, do not act independently on the motivation to engage in protective behavior. If a health problem is perceived to have no chance of occurring, there should be no interest in acting against it, regardless of how serious it might be. Nevertheless, researchers seldom observe the expected interaction between probability and severity. A case study approach was used to examine how probability and severity combine to influence interest in protection. Ratings of motivation to act, probability, and severity for 201 hazards were collected from 12 participants, and data were analyzed for each person separately. Analyses revealed the expected Probability x Severity interaction. Additional calculations showed why it is difficult to detect this interaction using between-subjects designs. The data also revealed that people are surprisingly insensitive to variations in hazard probability when probabilities are in the moderate to high range. PMID- 10711590 TI - Repression predicts outcome following multidisciplinary treatment of chronic pain. AB - This study examined whether repression predicts outcome following multidisciplinary treatment for chronic pain and whether links between anxiety and outcome are obscured by repressors. Ninety-three chronic pain patients completed a 4-week pain program. Lifting capacity, walking endurance, depression, pain severity, and activity were measured at pre- and posttreatment. Low-anxious, repressor, high-anxious, and defensive/high-anxious groups were formed from median splits of Anxiety Content (ACS) and Lie scales of the Minnesota Multiphasic Personality Inventory-2 (MMPI-2; Butcher, Dahlstrom, Graham, Tellegen, & Kaemmer, 1989). Significant ACS x Lie interactions were found for lifting capacity, depression, and pain severity changes. Planned comparisons showed that both repressors and high-anxious patients performed poorly on lifting capacity; repressors alone recovered poorly on depression and pain severity. Results imply that repression may interfere with the process and outcome of pain programs. PMID- 10711592 TI - Physician-diagnosed medical disorders in relation to PTSD symptoms in older male military veterans. AB - The association between physician-diagnosed medical disorders and combat-related posttraumatic stress disorder (PTSD) symptoms was examined in 605 male combat veterans of World War II and the Korean conflict. Physician exams were performed at periodic intervals beginning in the 1960s. PTSD symptoms were assessed in 1990. Cox regression was used to examine the onset of each of 12 disorder categories as a function of PTSD symptoms, controlling for age, smoking, alcohol use, and body weight at study entry. Even with control for these factors, PTSD symptoms were associated with increased onset of arterial, lower gastrointestinal, dermatologic, and musculoskeletal disorders. There was only weak evidence that PTSD mediated the effects of combat exposure on morbidity. Possible mediators of the relationship between combat exposure, PTSD, and physical morbidity are discussed. PMID- 10711591 TI - Follow-up of coping skills training in adults with sickle cell disease: analysis of daily pain and coping practice diaries. AB - This study examined the 3-month follow-up effects of a pain coping skills intervention in African American adults with sickle cell disease. Sixty-seven participants were randomly assigned to either a coping skills condition or a disease-education control condition. Multivariate analyses applied to summary measures of coping, laboratory pain perception, and clinical measures indicated that participants in the coping intervention reported significantly lower laboratory pain and significantly higher coping attempts at 3-month follow-up in comparison with the control condition. Multilevel random effects models applied to prospective daily diaries of daily pain, health care contacts, and coping practice indicated that on pain days when participants practiced their strategies, they had less major health care contacts in comparison with days when they did not use strategies. PMID- 10711593 TI - Genetic and biochemical properties of a hemolysin (pyolysin) produced by a swine isolate of Arcanobacterium (Actinomyces) pyogenes. AB - Arcanobacterium (Actinomyces) pyogenes, a causative agent of various pyogenic diseases in domestic animals, produces a hemolysin which is thought to be an important virulence factor. This hemolysin was purified from the culture supernatant of A. pyogenes swine isolate. The purified hemolysin showed a single band with a molecular mass of 56 kDa on SDS-polyacrylamide gel electrophoresis, and its isoelectric point was 9.2. The activity of this hemolysin was not enhanced by the addition of L-cysteine or sodium thioglycolate, but it was inhibited by cholesterol. The gene encoding the hemolysin was cloned, sequenced and expressed in Escherichia coli by means of ZAP Express vector. Analysis by SDS polyacrylamide gel electrophoresis with immunoblotting showed that the molecular weight of the hemolysin expressed in E. coli is the same as that of the hemolysin purified from A. pyogenes. Nucleotide sequence analysis revealed an open reading frame of 1,605 bp encoding a 534 amino acid protein of 57,989 Da. The nucleotide sequence of the hemolysin gene from A. pyogenes swine isolate differed only slightly (97.6% identity) from the sequence of plo gene from A. pyogenes strain BBR1 reported by Billington et al (J. Bacteriol. 179: 6100-6106, 1997). The cysteine residue existed in the undecapeptide region of the hemolysin, which is highly conserved in thiol-activated cytolysins (cholesterol-binding cytolysins), and is replaced with alanine. Therefore, the hemolysin of A. pyogenes seems to be a novel member of the thiol-activated cytolysin family. PMID- 10711594 TI - Highly selective antibacterial activity of novel alkyl quinolone alkaloids from a Chinese herbal medicine, Gosyuyu (Wu-Chu-Yu), against Helicobacter pylori in vitro. AB - To elucidate the antibacterial activity of Gosyuyu, the crude extract from the fruit of Evodia rutaecarpa, a Chinese herbal medicine, has been fractionated chromatographically, and each fraction was assayed for antibacterial activity against Helicobacterpylori (H. pylori) in vitro. As the result, a single spot having marked antibacterial activity against H. pylori was obtained and the chemical structure was analyzed. The isolated compound was revealed to be a novel alkyl quinolone alkaloid based on the solubility, IR spectra, NMR analysis and mass spectrometric data after purification by TLC of silica. We compared the antimicrobial activity of this compound with that of other antimicrobial agents and examined susceptibility of various intestinal pathogens. As the result, the new quinolone compounds obtained from Gosyuyu extracts were found to be a mixture of two quinolone alkaloids, 1-methyl-2-[(Z)-8-tridecenyl]-4-(1H)-quinolone and 1 methyl-2-[(Z)-7-tridecenyl]-4-(1H)-quinolone (MW: 339), reported previously. The minimum inhibitory concentration (MIC) of these compounds against reference strains and clinically isolated H. pylori strains were less than 0.05 microg/ml, which was similar to the MIC of amoxicillin and clarithromycin that are used worldwide for the eradication of H. pylori, clinically. Furthermore, it was noted that the antimicrobial activity of these compounds was highly selective against H. pylori and almost non-active against other intestinal pathogens. The above results showed that these alkyl methyl quinolone (AM quinolones) alkaloids were useful for the eradication of H. pylori without affecting other intestinal flora. PMID- 10711595 TI - Porphyromonas gingivalis fimbriae induce adhesion of monocytic cell line U937 to endothelial cells. AB - This study used the human monocytic cell line U937 to examine whether or not Porphyromonas gingivalis fimbriae could induce the adhesion of monocytes to endothelial cells. An in vitro adhesion assay was used to investigate the effects of the fimbriae on U937 cell adhesion to human umbilical vein endothelial cells (HUVEC). The fimbriae enhanced U937 cell adhesion to HUVEC in a dose-dependent manner. U937 cells adhered better to HUVEC pretreated with the fimbriae for a minimum of 2 hr than to untreated HUVEC. The enhanced adhesion was inhibited by a monoclonal antibody against P. gingivalis 381 fimbriae. Pretreatment of U937 cells with the fimbriae for 24 hr enhanced U937 cell adhesion to HUVEC approximately 4-fold. This phenomenon was inhibited by an anti-CD11b antibody, suggesting the involvement of CD11b. These results indicate that P. gingivalis fimbriae can induce monocyte adhesion to the endothelial cell surface. They also suggest that the fimbriae may be involved in the initial event for infiltration of monocytes into the periodontal tissues of individuals with adult periodontitis. PMID- 10711596 TI - Coprobacillus catenaformis gen. nov., sp. nov., a new genus and species isolated from human feces. AB - Three strains of Eubacterium-like isolates from human feces were characterized by biochemical tests and 16S rDNA analysis. The phenotypic characteristics of the three strains resembled those of the genus Collinsella transferred from the genus Eubacterium recently. However, Eubacterium-like strains were phylogenetically members of the Clostridium subphylum of gram-positive bacteria, and these showed a specific phylogenetic association with Clostridium ramosum and C. spiroforme. C. ramosum and C. spiroforme are gram-positive, anaerobic, spore-forming bacteria that belong to the genus Clostridium, and the G + C contents are 26.0 and 27.4 mol%, respectively. However, the three Eubacterium-like strains had G + C contents of 32.1 to 33.1 mol% and were non-spore-forming rods. Based on phenotypic characteristics, we can differentiate these species, and furthermore, a 16S rDNA sequence divergence of greater than 9% with a new related genus, Coprobacillus, is proposed for the three strains, with one species, Coprobacillus catenaformis. The type strain of C. catenaformis is JCM 10604T. PMID- 10711597 TI - Therapeutic oral vaccination induces mucosal immune response sufficient to eliminate long-term Helicobacter pylori infection. AB - We examined the efficacy of therapeutic oral vaccination using Helicobacter pylori-whole cell sonicate and cholera toxin (CT) in mice persistently infected with H. pylori. Efficacy was determined by bacterial culture and microscopic examination of gastric tissues for the persistence of bacteria at 6 weeks after the last vaccination. Vaccination of H. pylori-whole cell sonicate combined with CT eradicated bacteria in 10/16 mice (62.5%). Interestingly, oral vaccination with CT alone also eliminated the bacteria in 8/17 mice (47.1%). However, a therapeutic intraperitoneally administered vaccine failed to eradicate H. pylori from the stomach (1/17 mice, 5.9%). Identification of the type of immunity involved in the eradication process showed that oral vaccination enhanced the antigen-specific IgA in the feces and saliva. The efficacy of eradication of H. pylori correlated well with increases in IgA secretion in mucosal tissue and a higher labeling index of IgA-positive lumina of pyloric glands. Moreover, the expression of IL-4 mRNA in the stomach of mice with eradicated bacteria was higher than in the uneradicated group. Our results suggest that the efficacy of vaccination depends on the mucosal IgA response in the gastrointestinal tract against H. pylori via Th2 cell activation and that therapeutic oral vaccination induces a mucosal immune response sufficient to eradicate long-term infection with H. pylori. PMID- 10711598 TI - The dsbA-dsbB disulfide bond formation system of Burkholderia cepacia is involved in the production of protease and alkaline phosphatase, motility, metal resistance, and multi-drug resistance. AB - In a previous study, we isolated a dsbB mutant of Burkholderia cepacia KF1 and showed that phenotypes of protease production and motility are dependent on DsbB, a membrane-bound disulfide bond oxidoreductase. We have now isolated a dsbA mutant by transposon mutagenesis, cloned the dsbA gene encoding a periplasmic disulfide bond oxidoreductase, and characterized the function of the DsbA-DsbB disulfide bond formation system in B. cepacia. The complementing DNA fragment had an open reading frame for a 212-amino acid polypeptide with a potential redox active site sequence of Cys-Pro-His-Cys that is homologous to Escherichia coli DsbA. The dsbA mutant, as well as the previously isolated dsbB mutant, was defective in the production of extracellular protease and alkaline phosphatase, as well as in motility. In addition, mutation in the DsbA-DsbB system resulted in an increase in sensitivity to Cd2+ and Zn2+ as well as a variety of antibiotics including beta-lactams, kanamycin, erythromycin, novobiocin, ofloxacin and sodium dodecyl sulfate. These results suggested that the DsbA-DsbB system might be involved in the formation of a metal efflux system as well as a multi-drug resistance system. PMID- 10711599 TI - Characterization of the cyclophilin of Trichophyton mentagrophytes. AB - A genetic approach to cyclophilins in a dermatophyte, Trichophyton mentagrophytes, was carried out. The nucleotide and deduced amino acid sequences of the cyclophilin of T. mentagrophytes shared about 70% sequence similarity with those of Schizosaccharomyces pombe, Saccharomyces cerevisiae and Candida albicans. However, the first 21 amino acid and the C-terminal amino acid regions of 188 to 226 of the T. mentagrophytes cyclophilin were distinct from those of the other fungal cyclophilins. The recombinant glutathione S-transferase (GST)-T. mentagrophytes cyclophilin fusion protein produced by Escherichia coli was purified. The protease digest of the fusion protein had a molecular weight of about 13 kDa and peptidyl-prolyl cis-trans isomerase (PPI) activity. This digest protein from T. mentagrophytes was confirmed to be cyclophilin by proving PPI activity. PMID- 10711600 TI - Leukocyte integrin-dependent and antibody-independent cytotoxicity of macrophage against allografts. AB - Macrophages (Mphis), but not T cells, infiltrating into the rejection site of either i.p. allografted Meth A (H-2d) fibrosarcoma cells in C57BL/6 (B6) (H-2b) mice or BALB/c (H-2d) skin onto B6 mice are cytotoxic against allografts with H 2d specificity. To determine the mechanisms of specific killing of allografts by allograft-induced Mphi (AIM), we raised approximately 5,000 rat monoclonal antibodies (mAbs) against AIM and selected three of them (R1-73, R2-40 and R1 34), each of which inhibited cytotoxic activity against allografts in a dose dependent manner. The antigens recognized by R1-73, R2-40 and R1-34 mAbs were defined by immunoprecipitation and Western blot analyses as CD11a, CD18 and CD11b, respectively; and the allografts expressed CD54, a ligand of CD11a or CD11b, suggesting leukocyte integrin-dependent killing. Although Ab-dependent cellular cytotoxicity has been recognized as a mechanism of specific killing by Mphis, the infiltration of AIM into the rejection site of allografts far (approximately 6 days) preceded the appearance of serum IgG Ab specific for the allograft. AIM exhibiting full cytotoxic activity against allografts was also induced in the transplantation site of Fcgamma receptor knockout [(B6x129) F1] mice as well as B10.D2 (H-2 compatible with allograft) and B6-xid (X-linked immunodeficiency with B cell-specific defect) strains of mice. In the latter two strains of mice, the levels of serum IgG Ab to the allograft were negligible. Moreover, the cytotoxic activity of AIM against allografts was not affected by pretreatment of the cells with anti-mouse IgG serum, suggesting Ab-independent cytotoxicity. PMID- 10711601 TI - Simple preparation of parapoxvirus genome DNA for endonuclease analysis. AB - We compared five methods for improved extraction of very-large parapoxvirus DNA from infected cells: (i) alkaline-lysis procedure followed by phenol extraction; (ii) modified Hirt procedure, which was a neutral lysis procedure followed by phenol extraction; (iii) Hirt procedure; (iv) method used for extraction of vaccinia virus DNA; and (v) standard procedure using virus purification with an ultracentrifuge and protease-sodium dodecyl sulfate-phenol treatment. The alkaline-lysis procedure was more rapid, inexpensive and simpler than the other methods. Moreover, with this method it is not necessary to prepare any special facilities, reagents and kits. Although the extracted DNA was still crude, we could reproducibly prepare viral DNA from 2 X 10(6) infected cells in less than 2 hr and it could be readily digested by restriction endonuclease. This method will aid rapid genetic classification of parapoxvirus. PMID- 10711602 TI - Survey on antibody against parapoxvirus among cattle in Japan. AB - A seroepidemiological survey was performed on antibody against parapoxvirus among cattle in Japan using the agar gel immunodiffusion test and enzyme-linked immunosorbent assay. A total 1,819 sera were collected from cattle in various parts of Japan for the survey. The positive rates were in the range of 40 to 98%, and the reactors increased gradually in number with advancing age. These results indicate that parapoxvirus infection is already prevalent among cattle in Japan. It remains to be elucidated, however, whether the antibodies detected have been produced by infection of the same virus or other viruses that belong to the genus parapoxvirus. PMID- 10711603 TI - CPR-Total (TAFI and activated TAFI) levels in plasma/serum of hemophiliacs. AB - Arginine carboxypeptidase (CPR) is a single-chain plasma protein generated during coagulation from a precursor (proCPR). proCPR is the same molecule as thrombin activable fibrinolysis inhibitor (TAFI), which retards fibrin clot lysis in vitro and most likely modulates fibrinolysis in vivo. In this study, the amount of CPR total, which includes proCPR (TAFI) and CPR (activated TAFI), in hemophiliac patients was evaluated using a newly developed enzyme linked immunosorbent assay (ELISA). The amount of CPR-total in plasma or serum of most of the hemophiliac patients was in the range of healthy individuals. There was no significant difference in hemophiliac patients with or without HIV-1 infection. However, two out of the 74 hemophiliac patients showed a significantly high level. The upregulation of CPR-total might contribute to compensate for inefficient coagulation in some hemophiliac individuals. PMID- 10711604 TI - Root substance removal with Er:YAG laser radiation at different parameters using a new delivery system. AB - BACKGROUND: The recently introduced Er:YAG laser radiation appears to be a promising alternative in treating dental hard tissue due to its thermo-mechanical ablation properties and the lack of thermal side effects. The present in vitro study attempted to evaluate the use of Er:YAG laser radiation in combination with a specially developed delivery system in removing calculus from root surfaces. METHODS: Fifty extracted anterior teeth, premolars and molars, were divided into 2 groups of 25 each with (group A) and without (group B) subgingival calculus. Source of radiation was an Er:YAG laser device with a wavelength of 2.94 microm, in the infrared optical spectrum, a pulse duration of 250 ns, and a pulse repetition rate of 15 pps. In each group, 6 teeth were irradiated with 300 laser pulses either at 60 mJ, 80 mJ, 100 mJ, or 150 mJ. The samples were continually moved linearly using a computer numeric controlled device. The volumetric evaluation of root substance removal was performed with a 3-dimensional laser scanning system (100,000 surface points per sample, accuracy: 5 microm) and special image analyzing software. A scanning electron microscopic (SEM) observation was performed to assess the laser induced ultrastructural changes on the root surfaces. Statistical analysis was carried out with ANOVA followed by the Scheff*e test and with regression analysis according to Pearson-Bravais at a level of significance of 5% (P <0.05). RESULTS: The linear measurement of substance removal on calculus samples (group A) revealed average depths between 174.38 (+/-16.13) microm and 501.85 (+/-111.01) microm. Defect depths on the teeth without calculus (group B) ranged from 37.78 (+/-14.03) microm to 484.44 (+/-80.63) microm. The SEM observation of laser-treated root surfaces revealed no signs of thermal damage; e.g., melting, fusion, or cracking. CONCLUSIONS: The results of the present study showed that a substance removal with Er:YAG laser radiation at lower energy densities is comparable, in effect, to that after conventional root surface instrumentation with curets. The results seem to indicate that calculus removal can be selectively done using lower radiation energies. Considering the favorable results of the SEM investigation, the use of the Er:YAG laser in periodontal therapy may be possible in the future. PMID- 10711605 TI - Interleukin-1 genetic association with periodontitis in clinical practice. AB - BACKGROUND: Periodontitis is a bacterial disease modified by multiple risk factors. The pro-inflammatory cytokine interleukin- (IL-1) is a key regulator of the host responses to microbial infection and a major modulator of extracellular matrix catabolism and bone resorption. It has been reported that variations in the IL-1 gene cluster on chromosome 2 are associated with increased susceptibility to severe adult periodontitis. METHODS: The present study evaluated the association between a composite IL-1 genotype, including allele 2 at each of two loci (IL-1A +4845 plus IL- B +3954), and a broad spectrum of periodontally healthy to diseased patients in a population that is typically encountered in a dental practice setting. Ninety patients, non-smokers or former smokers with less than 10 pack-year (pk/yr) history, were recruited from a private dental practice. The major outcome variable was bone loss determined by computerized linear measurements of radiographs. Genotypes were analyzed from finger-stick blood samples using previously reported methods. RESULTS: Multivariate logistic regression models demonstrated that patient age, former smoking history, and the IL-1 genotype were significantly associated with severity of adult periodontitis. For non-smokers or former light smokers (<5 pk/yr), IL-1 genotype positives were at increased odds ratio of having moderate to severe periodontal disease of 3.75 (95% CI: 1.04-13.50) to 5.27 (95% CI: 1.23 22.70), depending on ethnicity, compared to IL-1 genotype negatives. Former moderate smokers (>5 pk/yr and <10 pk/yr) who were IL-1 genotype negative were at increased odds ratio of having moderate to severe periodontal disease of 7.43 (95% CI: 1.20-46.20) compared to non-smokers or former light smokers who were IL 1 genotype negative. In addition, past smoking history was also a significant effect modifier as demonstrated by the statistically significant interaction between past smoking history status and IL-1 genotype status. CONCLUSIONS: This study demonstrates that the composite IL-1 genotype is significantly associated with the severity of adult periodontitis. It also confirmed that both IL-1 genotyping and smoking history provide objective risk factors for periodontal disease in a private practice environment. PMID- 10711606 TI - Low prevalence of a periodontitis-associated interleukin-1 composite genotype in individuals of Chinese heritage. AB - BACKGROUND: Polymorphisms in the interleukin-1 (IL-1) gene cluster have been associated with an increased risk of developing certain diseases. A specific composite genotype of IL-1A and IL-1B polymorphisms, consisting of allele 2 of both IL-1A +4845 and IL-1B +3954 (formerly +3953) has been associated with an increased risk of severe adult periodontitis. Approximately 30% of the European population carry this genotype. The prevalence of the above IL-1A and IL-1B composite genotype in populations of different ethnic origins is unknown. Therefore, the primary aim of this study was to determine the prevalence of the IL-1 composite genotype in individuals of Chinese heritage, since epidemiologic studies indicate that periodontitis is widespread among ethnic Chinese. An additional aim was to evaluate if there was an association between the composite genotype and the severity of periodontal disease. METHODS: A convenience sample of 300 volunteers of Chinese heritage (ages 21 to 69 years) received a periodontal examination including full-mouth clinical attachment loss measurements, probing depths, plaque index scores, and bleeding on probing. Blood was collected from a fingerstick and placed on a blotting paper card. The blood samples were analyzed for IL-1A +4845 and IL-1B +3954 polymorphisms using polymerase chain reaction (PCR)-based methods. RESULTS: Only 7 of the 300 subjects (2.3%) carried the composite IL- 1 genotype consisting of allele 2 of both IL-1A +4845 and IL-1B +3954. Allele 2 of the IL-1A +4845 polymorphism was carried by 17.0% (51/300) of the subjects; of these, only 2 were homozygous. Allele 2 of the IL-1B +3954 polymorphism was much rarer with only 3.3% (10/300) of the study population carrying this marker. All of the people who carried the IL-1B polymorphism were heterozygous. Too few of the subjects were positive for the IL-1 composite genotype to establish any relationship with the susceptibility to periodontitis. CONCLUSIONS: It was concluded that the prevalences of both IL 1A and IL-1B polymorphisms are dramatically lower in Chinese than those reported for Europeans. Findings from this study bring into question the usefulness of the composite genotype of allele 2 of both IL-1A +4845 and IL-1B +3954 as a method for determining the susceptibility of Chinese patients to adult periodontitis. PMID- 10711607 TI - Mucogingival interceptive surgery of buccally-erupted premolars in patients scheduled for orthodontic treatment. I. A 7-year longitudinal study. AB - BACKGROUND: Mucogingival interceptive therapy in patients with buccally erupting teeth is performed to prevent the ectopic permanent tooth from developing periodontal lesions. The keratinized tissue entrapped between the erupting tooth and the deciduous tooth is retained to maintain a satisfactory width of the gingiva for the permanent tooth. The aim of the present study on buccally-erupted premolars scheduled for orthodontics was to evaluate the keratinized tissue width 3 months, 2 years, and 7 years subsequent to mucogingival interceptive therapy and orthodontic treatment. METHODS: Twenty-nine patients participated. Three different surgical techniques were used according to specific indications. Eight patients were treated with double pedicle flaps (DPF), 10 patients with apically positioned flaps (APF), and 11 with free gingival grafts (FGG). RESULTS: The amount of keratinized tissue on the treated (test) sites was not significantly less than on the control (untreated) sites showing normally erupting premolars at all observation periods. All 3 surgical procedures appeared to be effective in saving the keratinized tissue for the permanent tooth. Preoperative periodontal parameters such as gingival width, probing depth, and bleeding on probing significantly influenced the outcome 3 months after surgery (P <0.01). CONCLUSIONS: Mucogingival interceptive surgery is an effective approach to conserve the keratinized buccal gingiva of ectopically erupting premolars. PMID- 10711608 TI - Mucogingival interceptive surgery of buccally-erupted premolars in patients scheduled for orthodontic treatment. II. Surgically treated versus nonsurgically treated cases. AB - BACKGROUND: The aim of this 2-year longitudinal study was to compare the width of keratinized gingiva after orthodontic therapy for buccally erupting premolars that had been pretreated by extraction of deciduous teeth alone versus interceptive mucogingival surgery. METHODS: In 8 patients (aged 9 to 12 years) who presented with bilateral buccal eruption of homologous teeth (premolars), one side was randomly treated with extraction of the deciduous molar and mucogingival surgery (test site), while the other side was treated only by extraction of the deciduous molar (control site). All of the subjects underwent orthodontic treatment with fixed appliances. RESULTS: At the baseline visit prior to any treatment, there was no significant difference between the mean amount of keratinized gingiva at test sites (3.06 mm) and control sites (2.93 mm). Two years later, upon completion of orthodontic treatment, there was a significant difference between test (2.93 mm) and control (1.37 mm) sites in the mean width of keratinized tissue. In the control (untreated) group, 2 sites exhibited 1 mm of gingival recession after orthodontic treatment. CONCLUSIONS: Mucogingival interceptive surgery is an effective technique to maintain keratinized tissue in correspondence with buccally-erupted teeth. PMID- 10711609 TI - Coronally advanced flap procedure for root coverage. Flap with tension versus flap without tension: a randomized controlled clinical study. AB - BACKGROUND: This clinical controlled study was designed to measure the tension of coronally advanced flaps (CAF) performed to treat shallow gingival recessions and to compare the recession reduction (Rec Red) achieved in a test group (flaps with tension) and in a control group (flaps without tension) 3 months after surgery. METHODS: Eleven patients, aged 22 to 41 years, with high levels of oral hygiene (full mouth plaque score <20%) were selected for the study. Each patient showed 2 bilateral Miller Class I maxillary or mandibular gingival recessions located on homologous teeth. A total of 22 recessions were treated. The recession depth at the right site was similar to that at the left site (difference < or =1 mm). For each patient, the 2 recessions underwent CAF procedure in the same surgical session. Before suturing, the residual tension (FTens) of both right and left flaps was measured with a dynamometer. Then, one site was randomly assigned to the test group and the contralateral site to the control group. In the test site the flap was sutured. In the control site the flap was further relaxed, the tension was measured again, and the flap was sutured. RESULTS: In the test group (with tension) the initial mean recession depth was 2.82 +/- 0.64 mm and mean FTens was 6.5 g, while in the control group (without tension) the initial mean recession depth was 2.68 +/- 0.81 mm and mean FTens was 0.4 g. Three months later, the test group showed a mean recession reduction of 2.18 +/- 0.60 mm, a mean percent root coverage of 78 +/- 15%, and complete root coverage was achieved on 2 teeth (18%). In the control group the mean recession reduction was 2.32 +/- 0.81 mm and mean percent root coverage was 87 +/- 13%. Complete root coverage was obtained on 5 teeth (45%). The difference of recession reduction between the test and control group was not statistically significant (P = 0.3911). In the test group, linear regression analysis showed a statistically significant association between recession reduction and both recession depth at baseline (P= 0.0001) and mean of the 3 tensions recorded on the test side (MFTens) (P = 0.0009). CONCLUSIONS: This study shows that minimal flap tension does not influence recession reduction after 3 months when shallow recessions are treated by means of CAF. In the test group (with tension), the statistical analysis suggests that the higher the flap tension, the lower the recession reduction. PMID- 10711610 TI - Fluoroquinolones in the treatment of Actinobacillus actinomycetemcomitans associated periodontitis. AB - BACKGROUND: Periodontitis patients harboring Actinobacillus actinmycetemcomitans (Aa) are prime candidates for systemic antibiotic therapy. Besides tetracycline and the combination of metronidazole and amoxicillin the fluoroquinolones are also believed to have antibacterial activity against Aa. The aim of the present study was to evaluate systemic ofloxacin therapy as adjunct to flap surgery. METHODS: Twenty-five adult periodontitis patients with subgingival detection of Aa were treated with 2x200 mg/d ofloxacin for 5 days as adjunct to open flap surgery (test). Another 10 patients received only flap surgery (control). Probing depth (PD) and clinical attachment level (CAL) was recorded and subgingival plaque samples were cultivated on TSBV agar for detection of Aa at baseline as well as 3 and 12 months following therapy. RESULTS: At 3 and 12 months following therapy mean PD at monitored sites in the test group changed from 6.8 mm (+/-1.3) to 3.6 mm (+/-1.0), 3.8 mm (+/-1.1) and CAL from 7.5 mm (+/-1.4) to 5.4 mm (+/ 1.4), 5.5 mm (+/-1.3). In the control group PD changed from 6.5 mm (+/-0.7) to 4.0 mm (+/-1.7), 4.1 mm (+/-1.6) and CAL from 7.5 mm (+/-1.0) to 6.3 mm (+/-1.7), 6.4 mm (+/-1.8). P was <0.05 for CAL between groups. Three and 12 months following adjunctive systemic ofloxacin therapy, Aa was suppressed below detectable levels in 22 of 22, test patients, whereas Aa could not be recovered in only 2 of the 10 controls. (P<0.0001). CONCLUSIONS: Systemic ofloxacin as adjunct to open flap surgery is able to suppress A. actinomycetemcomitans below detectable level in patients harboring this organism at baseline. PMID- 10711611 TI - Peptostreptococcus micros smooth and rough genotypes in periodontitis and gingivitis. AB - BACKGROUND: Two genotypes can be distinguished within the species Peptostreptococcus micros: a smooth (Sm) and a rough (Rg) type. To date no systematic study has been performed on the prevalence and proportion of both types in untreated periodontitis patients and subjects without destructive periodontal disease. Therefore, the present study was performed to investigate: 1) the relative importance of the Sm and the Rg genotype of P micros in periodontitis and gingivitis; 2) the correlation between smoking and the 2 genotypes of P micros; and 3) the systemic antibody response against the 2 genotypes in relation to the periodontal condition and smoking. METHODS: A total of 104 untreated periodontitis patients and 41 individuals with gingivitis underwent clinical examination and microbiological sampling. Pocket samples were cultured anaerobically on blood agar plates to determine the prevalence and proportion of the Sm and Rg types of P micros. Serum antibody titers against both types of P micros were determined in all subjects by enzyme-linked immunosorbent assay (ELISA) using whole bacterial cells as antigen. Additionally, in a representative group of subjects, the antigen specificity of the serum antibodies was assessed by immunoblotting experiments. RESULTS: The prevalence of the Sm genotype was higher in subjects with periodontitis (94%) compared to subjects with gingivitis (59%), whereas the prevalence of the Rg type was not significantly different (38% versus 29%). Similar analyses were performed for subgroups of smokers and non-smokers; within the periodontitis group, the prevalence of the Sm type was not different between smokers and non-smokers (96% and 92%, respectively), whereas the prevalence of the Rg type was higher in smokers (48%) compared to non-smokers (19%). No difference in prevalence of both types was observed between smokers and non-smokers within the gingivitis group. The titers and specificity of P micros-specific immunoglobulins in periodontitis patients were not different from those in gingivitis subjects, nor were they related to smoking status or culture-positivity. CONCLUSIONS: The results of this study suggest that both the Sm and the Rg genotypes of P micros are part of the normal oral microbiota. However, the elevated prevalence of the Sm genotype in periodontitis and the elevated prevalence of the Rg type in periodontitis patients who smoke implies that both types can behave as opportunistic pathogens in destructive periodontal disease. PMID- 10711612 TI - A comparative scanning electron microscopic study on the characteristics of demineralized dentin root surface using different tetracycline HCl concentrations and application times. AB - BACKGROUND: The use of demineralizing agents has been reported to enhance the degree of connective tissue attachment to denuded roots. Of the agents used, tetracycline HCI has received the most attention. METHODS: The purpose of this study was to evaluate and compare the SEM surface morphology of human root dentin following various concentrations and application times of tetracycline HCl. The surface characteristics were compared and the width of dentin tubule orifices were measured and analyzed. Twelve human teeth were collected and stored in distilled water after soft tissue removal using hand instruments. The apical third of each root was removed and remaining mid-root region was sectioned longitudinally through the root canal following removal of its cementum with a fine diamond bur. A total of 48 dentin specimens were randomly divided into 8 groups for conditioning at different tetracycline HCl concentrations, at 0, 10, 25, 50, 75, 100, 125, and 150 mg/ml. Application times for each group were 1, 3, and 5 minutes. Immediately following treatment procedures, the specimens were rinsed, fixed in GTA-PBF, dehydrated, and prepared for SEM. Each specimen was examined at 2 magnification values, representative fields photographed, and data tested by one-way ANOVA and paired t test statistical analysis. RESULTS: Our results indicate that the use of tetracycline HCl solution between 50 mg/ml and 150 mg/ml showed a statistically significant opening of dentin tubules. All tetracycline HCl groups at 1, 3, and 5 minutes show smear layer removal from the dentin surface. CONCLUSIONS: Dentin demineralization is not time dependent at all concentrations of tetracycline HCI. PMID- 10711613 TI - Development of an in vitro wound healing model for periodontal cells. AB - BACKGROUND: Periodontal wound healing and regeneration are influenced by a multitude of factors. While many in vitro investigations have compared the proliferation of periodontal ligament (PDL) cells and gingival fibroblasts (GF), there are no reports directly comparing the abilities of these 2 cell types to fill a wound site. As such, the goals of this research were: 1) to develop an in vitro model of wound healing which would allow for the investigation of the biologic basis of periodontal wound healing and regeneration and 2) to compare the rates of PDL cells and GF to fill an in vitro wound site. METHODS: Using both human PDL cells and GF confluent cultures, in vitro wounds were mechanically created, removing a 3 mm wide band of the cell layer. Wounded cultures were then incubated for time periods up to 12 days in media containing fetal bovine serum (FBS) concentrations (0, 0.1, 1, 5, 10, and 20%) as appropriate for each experiment. Slides were fixed, stained, and cells quantified within the wound boundaries by computer-assisted histomorphometry. The effect of wounding a cell layer was determined by comparing wounded cells as described above with a cell layer margin created without physically disrupting the cell layer. RESULTS: The in vitro model for periodontal wound healing established in this study showed that GF fill in the wound site at a significantly (P <0.0025) faster rate than PDL cells over 12 days of healing. In addition, PDL cells and GF were found to have unique concentration-dependent responses to FBS (P<0.0025). It was also shown that wounding resulted in a significant delay (P <0.01) in the initial healing response of an in vitro wound. CONCLUSION: This in vitro model demonstrated that the characteristics of wound healing are dependent on cell type, disruption (wounding) of the cell layer, and serum concentration. In addition, this model has incorporated both proliferation and migration to provide the first direct evidence demonstrating GF has a significantly greater ability to fill a wound site than PDL cells. This in vitro model may be utilized in future investigations of the biologic basis of periodontal wound healing. PMID- 10711615 TI - Periodontopathic bacteria in children with Down syndrome. AB - BACKGROUND: It is widely known that individuals with Down syndrome (DS) often develop severe early-onset periodontal diseases. In this study, we examined the prevalence of periodontopathic bacteria in DS children to determine if specific pathogens are acquired in their childhood. METHODS: The subjects were 60 DS children (2 to 13 years old, 5 in each age bracket) and 60 age-matched controls. Ten pathogens, Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Bacteroides forsythus, Treponema denticola, Prevotella intermedia, P nigrescens, Capnocytophaga ochracea, C. sputigena, Campyrobacter rectus, and Eikenella corrodens were surveyed in subgingival plaque samples using a polymerase chain reaction. Periodontal status was evaluated by probing depth, bleeding on probing, and gingival index. RESULTS: No significant difference in periodontal status was observed between the DS and control groups, however, all of the pathogens were detected with greater frequency in the DS children. B. forsythus, T. denticola, P. nigrescens, and C. rectus were significantly prevalent throughout all age brackets of the DS children (P <0.01 or 0.05). The occurrence of P. gingivalis was also significant in the DS subjects over 5 years old. A cluster analysis of the microbial profiles of the DS subjects showed that gingivitis severity was associated with increased varieties of the harboring pathogens and the distribution of P. gingivalis. CONCLUSIONS: These results suggest that various periodontopathogens can colonize in the very early childhood of DS patients and maturation of subgingival components, including P. gingivalis, plays an important role in the initiation of gingival inflammation. PMID- 10711614 TI - Guided tissue regeneration with a bioabsorbable polylactic acid membrane in gingival recessions. A histometric study in dogs. AB - BACKGROUND: The goal of this investigation was to histologically and histometrically evaluate the healing process of gingival recessions treated by guided tissue regeneration with bioabsorbable polylactic acid membranes (GTR group) and to compare it to that obtained with coronally positioned flaps (CPF group). METHODS: Gingival recessions were surgically created on the buccal aspect of the upper cuspids of 5 mongrel dogs. The defects (5x7 mm) were exposed to plaque accumulation for 3 months. The contralateral defects were then randomly assigned to each group. After 3 months of healing, the dogs were sacrificed and the blocks were processed. The histometric parameters evaluated included length of sulcular and junctional epithelium, connective tissue adaptation, new cementum, new bone, and defect coverage. RESULTS: The extension of the epithelium was 1.9 +/- 0.8 mm for the GTR-group and 3.0 +/- 0.9 mm for the CPF-group (P = 0.16). The connective tissue adaptation was 0.1 +/- 0.1 and 0.8 +/- 0.5 mm in the GTR group and CPF group, respectively (P = 0.051). The new cementum was 3.8 +/- 1.5 mm and 2.4 +/- 0.3 mm in the GTR group and CPF group, respectively (P= 0.16). Bone formation was 1.1 +/- 0.5 mm in the GTR group and 1.4 +/- 0.2 mm in the CPF group (P = 0.53). Histologically, the defect coverage observed was similar, 90.5% and 91.9% for the GTR group and the CPF group, respectively. No statistical differences in any of the parameters could be detected. CONCLUSIONS: Within the limits of this study, it can be concluded that both procedures resulted in a favorable healing response with no significant difference between the treatments. PMID- 10711616 TI - Use of a biodegradable chlorhexidine chip in the treatment of adult periodontitis: clinical and radiographic findings. AB - BACKGROUND: Previous multi-center trials demonstrated the efficacy of a biodegradable chlorhexidine-gelatin chip (CHX) in reducing probing depth in patients with periodontitis. The present study utilized a subset of subjects from the parent study to determine if the CHX chip was effective in maintaining alveolar bone over a 9-month period. METHODS: Forty-five subjects with at least four 5 to 8 mm pockets, stratified by smoking status, were enrolled in this double-blind controlled, placebo-controlled trial. Control groups received either placebo chip plus scaling and root planing (SRP) or SRP alone. Test group subjects received active CHX chip or SRP alone (to maintain the blind). Standardized radiographs were taken for quantitative digital subtraction radiography at baseline and 9 months. RESULTS: At 9 months, 15% of SRP treated subjects experienced loss of bone in 1 or more sites, no subject treated with active chip plus SRP lost bone (P <0.01). At 9 months, significant differences in the change in probing depth and clinical attachment levels favoring the active chip over SRP alone or SRP plus CHX chip were also observed (P <0.05). CONCLUSIONS: These data indicate that the CHX chip, when used as an adjunct to scaling and root planing, significantly reduces loss of alveolar bone. PMID- 10711617 TI - The effect of chlorhexidine mouthrinses on early bacterial colonization of guided tissue regeneration membranes. An in vivo study. AB - BACKGROUND: Different membrane materials accumulate varying amounts of bacteria when exposed in the oral cavity, due to their textural and structural surface characteristics. The aim of the study was to evaluate the effect of chlorhexidine mouthrinses on the in vivo early bacterial colonization of 3 different guided tissue regeneration membrane materials. METHODS: Rectangular-shaped strips cut from 3 periodontal membranes (expanded polytetrafluoroethylene, polyglactin 910, and polylactic acid) were glued to removable devices adapted to the 2 upper quadrants in 8 dental students. In each student 1 quadrant was randomly selected as test side while the other served as control side. The experiment was divided in 2 phases: in the first phase plaque accumulation was followed for 4 hours while the second accumulation was followed for 24 hours. During the 4-hour experiment, students rinsed the test device twice (immediately following device application and after 2 hours) with 0. 12% chlorhexidine solution. The control device was rinsed with saline. In the second phase, students rinsed the test device with chlorhexidine and the control devices with saline 3 times (after device application and at 8 and 16 hours). Both the 4-hour and the 24-hour specimens were processed for scanning electron microscopy analysis. Fifty-four fields (at 200x magnification) were randomly selected and analyzed on each strip. Magnification was increased to determine the presence and morphotype of bacteria. The presence or absence of bacteria was assessed in a binomial fashion: the field was bacteria-positive when bacteria constituted the deposits covering the membrane surface. The microscopic field was negative (bacteria-negative) when no bacteria were observed. Bacteria-positive fields showing rods and filaments as prevalent morphotypes were recorded as rod-positive fields. RESULTS: The results of data analysis suggest that bacterial contamination of membrane materials is significantly reduced by treatment with chlorhexidine. They also suggest that other variables affect plaque accumulation as well; i.e., the time allowed (4 versus 24 hours) and the different membrane materials. The interaction between these 2 variables is also highly significant, thereby indicating a different rate of plaque accumulation on different materials, irrespective of the treatment with chlorhexidine. CONCLUSIONS: It was concluded that chlorhexidine mouthrinses may be effective in reducing and delaying the early bacterial accumulation on membrane materials although they are not able to fully prevent it. Membrane surface characteristics seem to be a more critical factor than the use of chlorhexidine, in influencing bacterial adhesion and colonization of barrier materials. PMID- 10711618 TI - Normal osteoconduction and repair in and around submerged highly bisphosphonate complexed hydroxyapatite implants in rat tibiae. AB - BACKGROUND: Ceramic hydroxyapatite implants have been used in dentistry for their unique compatibility with alveolar bone. Recently it was reported that bisphosphonates may be beneficial in preventing alveolar bone destruction associated with natural and experimental periodontal disease. Furthermore, bisphosphonate does prevent resorption of alveolar bone following mucoperiosteal flap surgery. We undertook a preliminary study evaluating the effects of highly bisphosphonate-complexed hydroxyapatite implants on osteoconduction and repair in rat tibiae. METHODS: Porous hydroxyapatite implants were pre-incubated in 10(-2)M bisphosphonate solutions at pH 3.49 and pH 7.32. The implants had a diameter of 2.1 mm and a height of 2 mm and adsorbed 115 microg bisphosphonate in vitro. Bisphosphonate/hydroxyapatite implants and plain hydroxyapatite implants were inserted in opposite tibial metaphyses of 35 rats. The measurement errors for the mineral density (MD) of the implants and the proximal trabecular mineral bone density (TD) were estimated by peripheral computed tomography and the bone mineral density (BMD) measurement error by dual x-ray absorptiometry. RESULTS: The measurement errors for the MD of the implants and the TD by peripheral computed tomography were 0.81% and 1.96%, respectively ex vivo. The BMD measurement error estimated by dual x-ray absorptiometry was 0.51% ex vivo. TD and BMD for bisphosphonate/hydroxyapatite implants were insignificantly higher compared to plain hydroxyapatite implants. Bisphosphonate/hydroxyapatite pre incubated at pH 7.32 were found to be nondegradable implants, while bisphosphonate/hydroxyapatite (pH 3.49) implants were slowly degradable and lost a significant 5% of their density. Histologically, all bisphosphonate/hydroxyapatite implants appeared to be fully integrated and effective as bone replacement material in rat tibial bone. They exhibited vascularization and osteoconduction of tibial bone growth along and inside their porous structure. CONCLUSIONS: Our study suggests that normal osteoconduction and repair occurred in and around the highly bisphosphonate-complexed hydroxyapatite implants in rat tibiae. PMID- 10711619 TI - Alveolar bone response to submerged bisphosphonate-complexed hydroxyapatite implants. AB - BACKGROUND: In most studies using submerged hydroxyapatite implants, maintenance of alveolar bone after tooth extraction was attempted with plain hydroxyapatite materials. However, clinical results have shown that hydroxyapatite may require biological modification with a bone resorption-inhibiting agent which may be beneficial for maintenance of alveolar bone. We conducted experimental and clinical studies to evaluate the effect of highly bisphosphonate-complexed hydroxyapatite implants on osteoconduction and repair in alveolar bone. METHODS: Porous hydroxyapatite implants were pre-incubated in 10(-2)M bisphosphonate solutions at pH 3.49. The implants had a diameter of 2.1 mm and a height of 2 mm and adsorbed 115 microg bisphosphonate. Five goats were implanted with 4 plain hydroxyapatite implants on each side of the mandible in root extraction sockets for the precision analysis of dual x-ray absorptiometry (DEXA) measurements. Ten goats were implanted with 4 bisphosphonate/hydroxyapatite implants on one side of the mandible and 4 plain hydroxyapatite implants on the opposite mandible. In a clinical study, 23 bisphosphonate/hydroxyapatite implants were placed in periodontally destroyed tooth root sockets and followed up during one year. RESULTS: The range for the bone mineral density (BMD) measurement errors for goat histologic sections was 0.48% to 1.03%. There were large differences in peri implant BMDs in the left and right mandible of the same goat, irrespective as to whether hydroxyapatite or bisphosphonate/hydroxyapatite implants were present. This was due to local anatomical differences typical of alveolar bone. These differences were not significant. Histologically, all bisphosphonate/hydroxyapatite as well as hydroxyapatite controls appeared to be fully integrated and effective as bone replacement material in goat alveolar bone. They exhibited vascularization and osteoconduction of alveolar bone growth along and inside their porous structure. In patients peri-implant healing was clinically and radiographically comparable to plain hydroxyapatite implants. All implants were retained and no dehiscences developed. Radiographically, peri implant radiolucencies disappeared and alveolar bone was deposited in close proximity to the implants. CONCLUSIONS: This study contributes to the understanding of the biological properties of hydroxyapatite implants as carriers for the bone-modulating agent bisphosphonate. Our study suggests that normal osteoconduction and repair occurred in alveolar bone around the highly bisphosphonate-complexed hydroxyapatite implants. PMID- 10711620 TI - Tissue response to titanium implants in the rat maxilla: ultrastructural and histochemical observations of the bone-titanium interface. AB - BACKGROUND: The detailed mechanism of osseointegration, the most appropriate implant-bone interface, remains unclear in jaw tissues at the ultrastructural level in contrast to the many reports using long bones. The present study reports on tissue response to titanium-implantation on an animal model using rat maxilla. METHODS: Animals were sacrificed at 1 to 28 days post-implantation and prepared tissue specimens, freed from implants by a cryofracture technique, were processed for transmission electron microscopy and histochemistry for tartrate resistant acid phosphatase activity (TRAPase). RESULTS: Different patterns in bone formation were recognized between lateral and base areas of implant cavities. In the lateral area with narrow gaps, bone deposition took place from the pre existing bone towards the implant after active bone resorption by osteoclasts reactive to TRAPase. However, no distinct bone formation appeared in the lateral area where the implant had been installed close to the osteotomy margin. On the other hand, new bone formation was found at the base area without any apparent bone resorption. Interestingly, mononuclear cells reactive to TRAPase, presumably preosteoclasts, frequently occurred near preosteoblasts. Osseointegration around the implants was obtained in this model by 28 days post-implantation except for the lateral area with complete contact with implants, where the thin layer remained in contact with the implant surface. CONCLUSIONS: These findings indicate that ossification proceeds at different modes around the titanium implant in rat maxilla, depending on the nature of the recipient bones and the dimension of the gap between the implant and osteotomy margin. PMID- 10711622 TI - Interproximal papillae reconstruction in maxillary implants. AB - BACKGROUND: Gingival esthetics has become an important factor in the overall success of most maxillary implant-supported restorations. Periodontal plastic surgery procedures may be used to enhance esthetics in the maxillary anterior region. The purpose of the present study was to evaluate a new surgical approach, performed at implant exposure, to reconstruct interdental papillae around maxillary implant-supported restorations. METHODS: The surgical procedure was performed on 32 patients, in which 36 consecutive single tooth osseointegrated implants were exposed in the anterior and premolar maxillary region. Previous to implant exposure and 6 months postoperatively, once the implant-supported restoration was in place, mesial and distal papilla contour measurements were calculated, based on a modification of the papillary index score (PIS). Statistical analysis consisted of paired t test, Pearson's correlation, and ANOVA with repeated measures. RESULTS: Preoperative PIS ranged from 0 to 3 and from 1 to 3 at the 6 months follow-up control. A mean of mesial and distal papilla, within the same tooth, was used for paired t test statistical analysis. A mean increase of 1.07 (SD 0.43) in PIS was statistically significant (P<0.001). At the second measurement, in no site was PIS smaller (0%) while in 64 sites PIS was higher (89%). In 51 papilla (71%) there was an increase of 1 PIS unit and 13 (18%) of 2 PIS units between both measurements. CONCLUSIONS: The presented surgical technique performed at second stage implant surgery was useful for partial or total interproximal papilla reconstruction adjacent to maxillary single-implant restorations. PMID- 10711621 TI - Gingival recession treatment: guided tissue regeneration with bioabsorbable membrane versus connective tissue graft. AB - BACKGROUND: Gingival recession represents a significant concern for patients and a therapeutic problem for clinicians. Several techniques have been proposed to achieve root coverage. The purpose of this randomized clinical trial was to evaluate the effect of a guided tissue regeneration (GTR) procedure in comparison to connective tissue graft (CTG) in the treatment of gingival recession defects. METHODS: Twelve patients, each contributing a pair of Miller Class I or II buccal gingival recessions, were treated. In each patient one randomly chosen defect received a poly(lactic acid)-based bioabsorbable membrane, while the paired defect received a CTG. Clinical recordings included oral hygiene standards and gingival health, recession depth (RD), recession width (RW), probing depth (PD), clinical attachment level (CAL), and keratinized tissue width (KT). RESULTS: Mean RD statistically significantly decreased from 2.5 mm presurgery to 0.5 mm with GTR (81% root coverage), and from 2.5 mm to 0.1 mm with CTG (96% root coverage), at 6 months postsurgery. Prevalence of complete root coverage was 58% for the GTR group and 83% for the CTG group. Mean CAL gain was 2.0 mm for the GTR group and 2.2 mm for the CTG group. No statistically significant differences between treatment groups were observed for changes in RD, RW, PD, CAL, and KT. CONCLUSIONS: Treatment of human gingival recession defects by means of either GTR or CTG results in clinically and statistically significant improvement of the soft tissue conditions of the defect when pre- and post-treatment measurements were compared. Although differences between CTG and GTR in mean root coverage and prevalence of complete coverage consistently favored the CTG procedure, the differences in measurements were not statistically significant. PMID- 10711623 TI - Treatment of gingival hyperpigmentation for esthetic purposes by Nd:YAG laser: report of 4 cases. AB - Gingival hyperpigmentation may cause esthetic problems and embarrassment, especially in patients with a gummy smile. This report presents the use of the Nd:YAG laser for gingival depigmentation. Four cases, 3 females and 1 male, ages between 24 to 28 years old, presented with the same chief complaint of unesthetic gingiva caused by melanin hyperpigmentation. The Nd:YAG laser was set at 6 watts, 60 millijoules per pulse, and 100 pulses per second. The procedures were performed with contact mode in all pigmented areas by using a handpiece with a 320 microm diameter fiber optic. Ablation of the gingival hyperpigmented areas were accomplished without any bleeding complications or significant postoperative pain. Three to 4 weeks after the procedures, the hyperpigmented gingiva appeared healthy, pink, and firm. No recurrence of hyperpigmentation had been found in 11 to 13 months of follow-up. However, in delicate areas such as the marginal gingiva, the Nd:YAG laser should be used cautiously. PMID- 10711625 TI - Periosteal osteosarcoma of the jaws: report of 2 cases. AB - Osteosarcoma (OS) occurs most often in the long bones. OS of the jaws has clinical and biologic aspects different from those of the long bones. They tend to occur at an older mean age, pain and swelling are more typical, and prognosis is more favorable. Nearly all OS shows a very prominent central intramedullary bone component. Only rarely are juxtacortical (peripheral) OS located in the jaws. There are 2 main types of juxtacortical OS, periosteal and parosteal. We present 2 cases of OS of the jaws where the clinical, radiologic, and histologic findings pointed to a diagnosis of periosteal OS. Both patients presented, in fact, with lesions located superficially on the bone surface with no marrow involvement. Both tumors were characterized by the presence of a moderately differentiated chondroblastic tumor with foci of osteoid and bone formation. Periosteal OS should be differentiated microscopically from periosteal chondrosarcoma, intramedullary OS with periosteal extension, high-grade surface OS, and parosteal OS. The clinical differential diagnosis was done, in these cases, for epulis, gingival tumors, peripheral odontogenic fibroma, peripheral ossifying fibroma, pyogenic granuloma, peripheral giant cell granuloma, and mesenchymal malignant tumors. PMID- 10711624 TI - Liposarcoma involving the periodontal tissues. A case report. AB - Liposarcomas constitute 15 to 20% of all soft tissue tumors. They are extremely rare in the head and neck and in the oral cavity. A 30-year-old patient was seen for a soft, painless mass in the right palate. Through panoramic radiography it was possible to observe a radiolucent area with sharp margins in the right upper quadrant. The lesion, after an incisional biopsy, was diagnosed as a "myxoid liposarcoma." The patient underwent a wide excision of the lesion with bone laminectomy and he is well at a 4-year follow-up. The differential diagnosis included salivary gland tumors and palatal abscess. PMID- 10711626 TI - Periodontics and patients with special needs. PMID- 10711627 TI - Effect of in utero and postnatal exposure to environmental tobacco smoke on the developmental expression of pulmonary cytochrome P450 monooxygenases. AB - Pulmonary cytochrome P450 monooxygenases metabolize xenobiotic chemicals, including those found in environmental tobacco smoke (ETS). Exposure to ETS beginning at birth has been shown to induce the P450 CYP1A1 by seven days of life. The effects of perinatal exposure to ETS of the rat lung on the expression of CYP1A1, 1B1, 2B1, and NADPH cytochrome P450 reductase were measured using semi quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). Timed pregnant dams and their pups were exposed to aged and diluted sidestream cigarette smoke (ADSS) as a surrogate for ETS for four hours/ day from gestational day 5 through postnatal day 21. For all genes analyzed, mRNA could be detected in the fetal lung beginning at gestational day 17 but were not altered by ADSS. In contrast, intraperitoneal injection of dams with beta-naphthoflavone significantly elevated both CYP1A1 and 1B1 at gestational day 21, indicating that these genes are inducible. Continued exposure to ADSS significantly induced CYP1A1 but not other P450 genes as early as one day after birth.. We conclude that (1) ADSS induces pulmonary CYP1A1 in the first day of life; (2) fetal cytochrome P450 genes are not induced by maternal exposure to ADSS; and (3) in the fetal lung, CYP1A1 and 1B1 can be induced by beta-naphthoflavone. PMID- 10711628 TI - Modulation of xenobiotic metabolism and oxidative stress in chronic streptozotocin-induced diabetic rats fed with Momordica charantia fruit extract. AB - We studied the long-term effects of streptozotocin-induced diabetes on tissue specific cytochrome P450 (CYP) and glutathione-dependent (GSH-dependent) xenobiotic metabolism in rats. In addition, we also studied the effect of antidiabetic Momordica charantia (karela) fruit-extract feeding on the modulation of xenobiotic metabolism and oxidative stress in rats with diabetes. Our results have indicated an increase (35-50%) in CYP4A-dependent lauric acid hydroxylation in liver, kidney, and brain of diabetic rats. About a two-fold increase in CYP2E dependent hepatic aniline hydroxylation and a 90-100% increase in CYP1A-dependent ethoxycoumarin-O-deethylase activities in kidney and brain were also observed. A significant increase (80%) in aminopyrene N-demethylase activity was observed only in rat kidney, and a decrease was observed in the liver and brain of diabetic rats. A significant increase (77%) in NADPH-dependent lipid peroxidation (LPO) in kidney of diabetic rats was also observed. On the other hand, a decrease in hepatic LPO was seen during chronic diabetes. During diabetes an increased expression of CYP1A1, CYP2E1, and CYP4A1 isoenzymes was also seen by Western blot analysis. Karela-juice feeding modulates the enzyme expression and catalytic activities in a tissue- and isoenzyme-specific manner. A marked decrease (65%) in hepatic GSH content and glutathione S-transferase (GST) activity and an increase (about two-fold) in brain GSH and GST activity was observed in diabetic rats. On the other hand, renal GST was markedly reduced, and GSH content was moderately higher than that of control rats. Western blot analyses using specific antibodies have confirmed the tissue-specific alterations in the expression of GST isoenzymes. Karela-juice feeding, in general, reversed the effect of chronic diabetes on the modulation of both P450-dependent monooxygenase activities and GSH-dependent oxidative stress related LPO and GST activities. These results have suggested that the modulation of xenobiotic metabolism and oxidative stress in various tissues may be related to altered metabolism of endogenous substrates and hormonal status during diabetes. The findings may have significant implications in elucidating the therapeutic use of antidiabetic drugs and management of Type 1 diabetes in chronic diabetic patients. PMID- 10711629 TI - Metabolism of thalidomide in human microsomes, cloned human cytochrome P-450 isozymes, and Hansen's disease patients. AB - Previous in vitro studies in rat microsomal preparations suggested that thalidomide is metabolized by the cytochrome P450 system (CYP). In this study, we examined the extent of thalidomide metabolism by preparations of pooled human microsomes, microsomes containing cloned human CYP isozymes (CYPIA2, CYP2A6, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1, CYP3A4), and Hansen's disease patients. Results indicated that thalidomide was a poor substrate for CYP isozymes. Alteration of incubation buffer, pH, incubation time, and microsome and thalidomide concentrations did not increase the production of any metabolites. Thalidomide also did not inhibit metabolism of CYP-specific substrates and therefore any interactions with other drugs that are metabolized by the same enzyme system are unlikely. Hansen's patients were given a single oral dose of thalidomide (400 mg), and their blood and urine were collected at time points up to 72 hours, processed, and analyzed by tandem mass spectrometry. Although thalidomide was present in the plasma and urine, no metabolites were found in the plasma and very low amounts of the 5-OH thalidomide metabolite were present in the urine. These results suggest that thalidomide does not undergo significant metabolism by human CYP and that clinically important interactions between thalidomide and drugs that are also metabolized by this enzyme system are unlikely. The major route of thalidomide breakdown in humans and animals is through spontaneous hydrolysis with subsequent elimination in the urine. PMID- 10711630 TI - The influence of diet on the regional distribution of glutathione S-transferase activity in channel catfish intestine. AB - There is evidence that glutathione conjugates are the major metabolites formed following systemic uptake of carcinogenic contaminants from the intestine. The effect of commercial diet versus a semi-purified diet on the distribution of glutathione S-transferase (GST) activity was examined in proximal, medial, and distal sections of catfish intestine. The bulk of GST activity with 1-chloro-2,4 dinitrobenzene, ethacrynic acid, and 3H-benzo[a]pyrene-4,5-oxide, and the percent cytosolic protein cross-reacting with anti-catfish GST-pi were in the more proximal segments and dropped off distally in the two diet groups. However, the total GST-pi cross-reacting protein in the proximal section was significantly higher in fish fed a chow diet. Western blot analysis revealed pi-class GST to be expressed principally in the proximal intestine. Cytosol samples cross-reacted with antibodies to human GST-alpha, -mu, and -pi, but not -theta, classes. Alpha like GST isoforms of MW 26,200 and 24,600, absent in sections from fish fed a purified diet, were differentially expressed only in the distal section of chow fed fish. These results indicate that diet significantly elicits regional differences in GST protein levels, that components of the commercial chow affect GST protein expression in the distal intestine, and that maintenance diet should be taken into consideration during dietary exposure studies. PMID- 10711631 TI - Aryl hydrocarbon hydroxylase activity in F-344 rats subchronically exposed to benzo(a)pyrene and fluoranthene through diet. AB - In order to investigate the relationship between aryl hydrocarbon hydroxylase (AHH) activity and exposure to benzo[a]pyrene [B(a)p] and fluoranthene (FLA), AHH activities in liver tissues of male and female F-344 rats were determined. Based on a range-finding study, doses of 0, 5, 50, and 100 mg/kg B(a)p or 0, 150, 750, and 1500 mg/kg FLA were administered in the animal diet over a 90-day period. After dosing, animals were sacrificed, liver tissues were removed, and microsomes were isolated. AHH activities were determined by reverse-phase HPLC coupled with fluorescence detection using 3-hydroxy B(a)p, and trans-2,3-dihydroxy-1,10-epoxy 1,2,3,10b tetrahydrofluoranthene as the standards. A dose-dependent increase in enzyme activity was observed with increased B(a)p or FLA exposure in both males and females. Our results also demonstrate that B(a)p-exposed females possess a higher AHH activity than males, but there is no significant sex difference with regard to enzyme activity in the case of FLA at higher doses. Overall, our findings suggest that long-term exposure to the parent compound results in elevated levels of AHH activity, which may contribute to the formation of toxic reactive metabolites and subsequent symptoms in target organs. PMID- 10711632 TI - Monovalent cation effects on the activity of the xenobiotic/medium-chain fatty acid:CoA ligases are substrate specific. AB - The effect of monovalent cation on the activity of the XL-I and XL-III forms of xenobiotic/medium-chain fatty acid:CoA ligase (XM-ligase) was investigated using a variety of different carboxylic acid substrates. With benzoate or p hydroxybenzoate as substrate, the XL-I ligase was essentially inactive in the absence of monovalent cation. However, with phenylacetic acid and medium-chain fatty acids as substrate, the enzyme retained 3 to 10% activity upon removal of monovalent cation. Further, while Na+ was ineffective with benzoate and p hydroxybenzoate as substrates, it was effective with other substrates, although still less effective than K+. For XL-III, activity toward benzoate, hydroxybenzoate, and salicylate was insignificant in the absence of monovalent cation, but this rate was 10% of the K(+)-supported rate for hexanoate and 20% for decanoate. Also, with decanoate as substrate, XL-III was activated more by Na+ than by K+. Thus, the nature of the dependence on monovalent cation for activity is substrate-selective. Kinetic analysis of the effect of K+ on the activity of XL-I and XL-III revealed that activation by K+ was not the result of alteration of the affinity of the enzymes for either ATP or the carboxylic acid. For both forms of XM-ligase, K+ was found to enhance the affinity of the enzyme for CoA, regardless of the substrate, although the extent of the enhancement was substrate-specific. In almost all cases there was further activation, even at saturating concentrations of CoA, which indicates an additional effect of monovalent cation on the catalytic rate constant for the reaction. The exception was activation of XL-III activity toward decanoate, which was solely the result of enhanced binding affinity for CoA. PMID- 10711633 TI - Effects of trivalent antimony on human erythrocyte glutathione-S-transferases. AB - Trivalent antimony (SB3+) in the form of potassium antimony tartrate was found to be an inhibitor of glutathione-S-transferases (GST) from human erythrocytes with a 50% inhibition concentration (IC50) of 0.05 mM. The inhibition was, however, incomplete with 15-20% of the GST activity remaining unaffected. In comparison, ethacrynic acid, a known inhibitor of GST, was tenfold more potent and affected close to 100% inhibition. Pentavalent antimony (SB5+) in the form of sodium stibogluconate had no effect on GST. Group V metalloids such as arsenite was slightly inhibitory, and arsenate was noninhibitory. When compared with five heavy metals, the inhibitory potency followed the order of SB3+ > Hg2+, Cu2+ > Cd 2+ > Cr3+ > Fe2+ x SB3+ inhibition of GST was competitive against the substrate 1 chloro-2,4-dinitrobenzene (CDNB) with an apparent Ki of 0.018 mM. Increasing the glutathione (GSH) concentration, however, produced a biphasic response: at concentrations below 1 mM, GSH was noncompetitive against SB3+, but at 1 mM and higher it was apparently competitive. A concurrent study of interactions between GSH, CDNB, and SB3+ showed that there was a significant nonenzymatic conjugation of CDNB at high GSH concentrations, which was suppressed by SB3+. The presence of albumin (500 mg/dL), or up to 5 mM N-acetylcysteine, cysteine, or ethylenediamine tetraacetic acid (EDTA) did not protect GST from the inhibitory effect of SB3+. The ability of erythrocyte GST to conjugate CDNB, which was measured directly by the formation of dinitrophenyl-glutathione (DNP-glutathione), was reduced by approximately 20 and 33%, respectively, in the presence of 2 and 10 mM SB3+, and nearly abolished with the addition of 0.2 mM ethacrynic acid. Based on these inhibition characteristics and the preferential accumulation of SB3+ in mammalian erythrocytes, it may be deduced that in the case of high antimonial intake, for example, during therapeutic treatment of Leishmaniasis, SB3+ levels in erythrocytes may be high enough to depress GST activity, which might compromise the ability of erythrocytes to detoxify electrophilic xenotbiotics. PMID- 10711634 TI - A new technique for implantation of tissue culture melanoma cells in a murine model of metastatic ocular melanoma. AB - The aim of this study was to compare the transcorneal and transconjunctival techniques for the implantation of intraocular melanoma cells and development of metastasis in a murine model. Groups of C57BL/6 mice were given either transconjunctival or transcorneal inoculations of 2.5 x 10(5)/2.5 microl tissue culture B16-LS9 melanoma cells into the intraocular posterior compartment (PC). The eyes were enucleated at 4-11 days post-inoculation and histologically examined. The mice were sacrificed 14 days after enucleation and necropsies were performed with histological evaluation for visceral metastases. Intraocular and extraocular tumour growth was present in all of the eyes inoculated via the transconjunctival route. Pulmonary metastases were found in this group if the eye was enucleated 7 or more days post-inoculation. The melanoma remained confined to the inside of the eye in the transcorneal group until day 7. Haematogenous metastases to the lung and liver developed from the intraocular melanoma in this group. Transcorneal inoculation of tissue culture melanoma cells into the murine PC provides a useful animal model for visceral metastasis of ocular melanoma. PMID- 10711635 TI - Comparison of in vitro cytotoxicity of N-acetyl and N-propionyl derivatives of phenolic thioether amines in melanoma and neuroblastoma cells and the relationship to tyrosinase and tyrosine hydroxylase enzyme activity. AB - Our laboratory has synthesized two new phenolic thioether amines, N-propionyl-4-S cysteaminylphenol (N-Pr-4-S-CAP) and N[2-[(4-propionyloxyphenyl)thio]ethyl] propionamide (N,O-diPr-4-S-CAP). These compounds, along with the previously described phenolic thioether amine N-acetyl-4-S-cysteaminylphenol (N-Ac-4-S-CAP) and its acetyl form (N,O-diAc-4-S-CAP), are tyrosine-amine derivative analogues. The cytotoxicity of these compounds is thought to be tyrosinase dependent, which may make them suitable for targeted anti-melanoma therapy since only melanocytes and their malignant counterparts contain this active enzyme. To further investigate this hypothesis, we performed MTT [3-(4,5-dimethylthiazol-2-yl)2,5 diphenyltetrazolium bromide] assays to determine the cytotoxicity of these compounds in 10 different cell lines. Specifically, we examined to what extent cytotoxicity is related to tyrosinase and tyrosine hydroxylase activity using melanoma and neuroblastoma cells, which have a common metabolic pathway using tyrosinase and tyrosine hydroxylase, respectively. The most sensitive cell line was the highly pigmented SK-MEL-23 melanoma cell line, which shows a very high tyrosinase activity with the highest melanin pigmentation. KAN and SK-NSH (two neuroblastoma cell lines), which have no tyrosinase activity but high tyrosine hydroxylase, were also sensitive. However, C32 (a non-pigmented melanoma with a lower tyrosinase activity) was also sensitive, and MeWo (a moderately pigmented melanoma with a high tyrosinase activity) was less sensitive. This in vitro study may indicate that there is a non-tyrosinase-mediated mechanism of cytotoxicity for phenolic thioether amines in addition to the tyrosinase-mediated one described previously. PMID- 10711636 TI - A direct comparison of cytolytic T-lymphocyte responses to Melan-A peptides in vitro: differential immunogenicity of Melan-A27-35 and Melan-A26-35. AB - In this study we directly compared the in vitro responses of T-cells from normal donors and melanoma patients to Melan-A27-35 and Melan-A26-35. These peptides have been previously used in peptide-based vaccination studies. Following three stimulations with peptide-pulsed antigen-presenting cells in vitro, Melan-A specific cytolytic T-lymphocytes (CTLs) were generated from seven of 20 subjects; two of the seven subjects responded reproducibly to both Melan-A27-35 and Melan A26-35, three to only Melan-A27-35 and two to only Melan-A26-35. However, CTLs generated with either Melan-A27-35 or Melan-A26-35 showed cross recognition, and both types of CTL could recognize naturally processed antigen displayed on HLA A2+ tumour cells. Furthermore, Melan-A-specific CTLs could also be generated by stimulating peripheral blood mononuclear cells with autologous melanoma cells. Our results suggest that some subjects may have a bias in their CTL repertoire which favours the generation of Melan-A27-35 specific CTLs, while others may favour Melan-A26-35 specific CTLs. It is also likely that CTL precursors capable of detecting both peptides may have different affinities to the two Melan-A peptides. Since it is difficult to predict the CTL responses to Melan-A peptide in a given individual, we suggest vaccinating with both Melan-A27-35 and Melan A26-35 peptides in clinical trials. PMID- 10711637 TI - Experimental ruthenium plaque therapy of amelanotic and melanotic melanomas in the hamster eye. AB - The effects of beta-radiation on melanoma implanted into the hamster's eye were investigated. Two Bomirski hamster melanomas (BHMs), differing in their melanin content, were compared with regard to their radiosensitivity to ruthenium-106 (106Ru) radiation. Tumours growing in the iris were irradiated with 3, 6 or 10 Gy of 106Ru given as a single dose or in four fractions at 24 h Intervals. Tumour growth kinetics and distant metastases were studied, and the eyeballs were examined histologically. Dose-dependent delay of tumour growth was observed in both melanomas. After treatment with a dose of 6 Gy, the Ab amelanotic tumours grew 2.6 times slower, and the Ma melanotic tumours 1.4 times slower than untreated ones. The location of metastases differed in the two tested lines- pigmented metastases were found mainly in the lungs, while unpigmented metastases were found mainly in the kidneys. Histopathological analysis showed signs of blood vessel damage such as endothelial cells swelling, erythrocyte extravasation and tumour necrosis. This last finding increased with the rising dose of beta radiation. Pigmented tumours were found to be two times more resistant to beta radiation than amelanotic ones. The pattern of metastases of BHMs is determined by the type of melanoma (Ab or Ma). Exposure to beta-radiation from 106Ru did not significantly affect either the number or size of metastases except at a dose of 10 Gy. This dose caused a statistically significant decrease in the number of metastases in the Ma melanotic subline. PMID- 10711638 TI - Cutaneous dysplastic naevi in uveal melanoma patients: markers for prognosis? AB - We have previously reported that the presence of cutaneous dysplastic naevi is a risk factor for uveal melanoma. In the present study our goal was to determine the incidence of different histopathological features of uveal melanoma among 91 patients with or without cutaneous dysplastic naevi. Statistical analysis revealed that the presence of cutaneous dysplastic naevi in uveal melanoma patients is associated with an increased incidence of the prognostically worst forms of uveal melanoma (epithelioid or mixed cell type melanomas). The relative risk was 5.97 (95% confidence interval 1.61-22.14). Our results suggest that the presence of cutaneous dysplastic naevi is not only a risk factor but also a prognostic factor for uveal melanoma. PMID- 10711639 TI - Contrast-enhanced high resolution magnetic resonance imaging of pigmented malignant melanoma using Mn-TPPS4 and Gd-DTPA: experimental results. AB - The aim of this study was to evaluate the potential of the paramagnetic metalloporphyrin Mn-TPPS4 (using Gd-DTPA as the reference) for magnetic resonance imaging (MRI) of pigmented malignant melanoma in an animal model. High resolution MRI (2.0 T, 2.0 cm surface coil, T1-weighted FLASH two-dimensional sequence) was performed on 15 mice (C57bl6) with intracutaneous implanted melanoma (B16F1) before and after intravenous administration of Gd-DTPA (Magnevist, Schering AG, Berlin, Germany) and Mn-TPPS4 (Porphyrin Products, Logan, Utah, USA). The images were evaluated quantitatively by calculating the percentage enhancement, the slope of the signal intensity-to-time curves, the percentage increase in the signal intensity, and the signal-to-noise and contrast-to-noise ratios. The qualitative evaluation was accomplished by visual assessment of the enhancement, the demarcation of the tumours from the surrounding tissue, and the homogeneity of the tumours. Contrast medium-enhanced images showed an increase in signal intensity for all the tumours, with no significant difference between the contrast media. Specific accumulation of the contrast media in the melanoma could not be proved. Demarcation of tumours from the surrounding tissue is better after administration of contrast media; regressive changed areas were better depicted. PMID- 10711640 TI - Can tissue drug concentrations be monitored by microdialysis during or after isolated limb perfusion for melanoma treatment? AB - Isolated limb perfusion (ILP) with melphalan is used to treat recurrent melanoma. This study aimed to develop a microdialysis technique for melphalan tissue concentration measurement during ILP. The effects of melphalan concentration (50 600 microg/ml), microdIalysis flow rate (0.55-17.5 microl/min), probe length (5 50 mm) and temperature (25-41.5 degrees C) on in vitro recovery were studied. In addition, in vivo recovery was measured in rat hindlimbs perfused with melphalan using 50 mm microdialysis probes implanted subcutaneously and into muscle. Both dialysate and tissue sample melphalan concentrations were determined by high performance liquid chromatography. The in vitro recovery of melphalan was not affected by melphalan concentration or temperature, but increased with probe length and decreased with flow rate. The melphalan concentrations in subcutaneous and muscle dialysates were not significantly different. A linear relationship was found between tissue dialysate concentrations and actual tissue concentrations of melphalan (r2 = 0.97). Microdialysis is a potential method for tissue drug monitoring which may assist in the efficacious use of cytotoxics in human ILP. PMID- 10711641 TI - Macrophage-mediated immunostimulation modulates therapeutic efficacy of interleukin-2 based chemoimmunotherapy in advanced metastatic melanoma patients. AB - The biological mechanisms of chemoimmunotherapy efficacy in vivo have not been fully clarified; furthermore, few data are available to predict its efficacy on the basis of clinical and immunological pretreatment factors. In this paper, pre- and post-treatment serum levels of cytokines (interleukin [IL]-6, IL-10, IL-12 and neopterin) and soluble IL-2 receptors (sIL-2R), as well as circulating levels of T-cell and NK subpopulations, were analysed according to clinical outcome in 66 advanced metastatic melanoma (MM) patients treated with subcutaneous IL-2 in association with interferon-alpha, cisplatin and tamoxifen. Our purpose was to correlate the immune modifications during treatment with the clinical response and to define pretreatment factors with predictive value for clinical outcome. The overall response rate was 35%, with a median overall survival of 11.3 months. During treatment, responding patients showed a common marked increase in IL-12 (mainly released by activated macrophages), sIL-2R and neopterin serum levels, associated with high levels of total lymphocytes and circulating natural killer lymphocytes; progressing patients were characterized by an increase in IL-6 serum levels (directly related to the increase in tumour burden). Multivariate analysis showed that high pretreatment IL-12 levels (P = 0.05) and, to a lesser extent, lactate dehydrogenase levels in the normal range (< or = 450 U/I; P = 0.061) are independent favourable prognostic factors for survival. Our results show that macrophage activation in an immunostimulating way either before or during treatment is associated with a better clinical response and improved survival in advanced MM patients treated with IL-2-based chemoimmunotherapy. PMID- 10711642 TI - Subcutaneous interleukin-2 and interferon-alpha plus cisplatin with and without prophylactic cimetidine in patients with metastatic malignant melanoma: a phase II study. AB - A phase II study was performed to evaluate the efficacy of cisplatin combined with interleukin-2 and interferon-alpha2b administered subcutaneously to patients with metastatic malignant melanoma (MMM). Between April 1994 and January 1999, 87 patients with MMM and a WHO performance status of < or = 2 were entered into the study. The first 42 patients had prophylactic cimetidine; the other 45 patients did not. An overall response rate of 27% was achieved in the 82 patients evaluable for response. The median response duration was 7.0 months (range 4.4 29.0 months). The median survival for all patients was 10.1 months (range 0.4 64.9+ months). Toxicity was substantial but generally manageable and usually reversed on dose reduction or temporary interruption of treatment. Two patients (2%) died of treatment-related toxicity. No difference in response or survival was seen in the patients treated with or without cimetidine. In multivariate analysis, lactate dehydrogenase level (P < 0.001), number of metastatic sites (P = 0.014) and performance status (P = 0.035) was shown to be independent prognostic factors for survival. This high dose interleukin-2 subcutaneous regimen resulted in a small fraction of long-term survivors. The response and survival results were not superior to other studies using lower and less toxic interleukin-2 doses. PMID- 10711643 TI - Melanoma metastatic to the spine: a review of 133 cases. AB - Although spine metastasis from melanoma is an uncommon event, it can pose a complex management problem. The presentation and natural history of melanoma metastatic to the spine has not been described in the medical literature. We have conducted a review of the records of 133 patients with melanoma metastatic to the spine in order to obtain retrospective data on demographic information, clinical presentation, disease course and survival. Patients with cutaneous, ocular and mucosal melanoma were all represented, but those with primary cutaneous tumours of the trunk were more prevalent than expected. Other sites of metastatic disease were present in nearly all patients and metastases to other skeletal sites were not unusual. Pain was the most common presenting symptom. The radiographic diagnosis was generally made easily by plain radiographs, computed tomography or magnetic resonance imaging, with the most frequent finding being a destructive lesion. Bone scan gave false-negative results 15% of the time. The median survival for the group was 4 months. It is concluded that melanoma metastatic to the spine represents a late event in the evolution of this illness. Palliation should be the goal of treatment, but symptom management should be individualized, bearing in mind the short anticipated survival of these patients. PMID- 10711644 TI - Duration of survival for disseminated malignant melanoma: results of a meta analysis. AB - The aim of this study was to estimate the survival experience of patients with disseminated malignant (stage IV) melanoma regardless of the treatment received. Articles, including abstracts, published in the English language medical literature between 1985 and 1999 were identified through an electronic key word literature search using MEDLINE and PUB-MED together with the reference lists of review articles. Some literature published prior to 1985 was identified through these searches and was also included. Median survivals were calculated across all the data, for the post- and pre-1985 periods, and for the larger studies alone (n > or = 20) using a weighting scheme based on sample size and study characteristics. An attempt was also made to estimate the frequency of long-term survival to 2, 3 and 5 years in this patient population (also stratified according to the study publication time). A total of 83 studies were identified comprising a total of 6322 patients. The overall median survival from diagnosis of stage IV melanoma was estimated to be 8.1 months (approximate 95% confidence interval [CI] 7.3-8.9 months). For the 59 studies published since 1985 (covering 3715 patients), the median survival was 8.9 months (approximate 95% CI 7.9-9.9 months). Prior to 1985, the median survival was 5.8 months (approximate 95% CI 4.5-7.1 months). For the 67 studies with at least 20 patients (a total of 6024 patients), the median survival was 8.1 months (approximate 95% CI 7.2-9.0 months). Long-term survival for the post- and pre-1985 periods over 2, 3 and 5 years was estimated to be 13.6%, 9.7% and 2.3%, respectively, and was consistent across the two time periods. In conclusion, while the survival experience in stage IV melanoma patients has improved since 1985, the use of varied approaches to treat this disease has not produced a favourable long-term prognosis. This meta-analysis will allow the survival results from current and future trials to be compared with the expected survival based on cumulative findings to date. PMID- 10711645 TI - Beyond ethical individualism. AB - Contemporary ethical debate about clinical practice centres primarily on the individual resolution of dilemmas, an approach which is incompatible with the social constructionist focus on human interdependence. Many constructionists argue that virtue ethics (VE) offers a more useful perspective on ethics than either consequentialism or deontology. From this perspective, the purpose of ethics is not to specify the right act in a particular situation, but to understand ethical and unethical practices conceptually, i.e. how these are learned, and how these contribute to and develop the ethical life in an ethical environment. Criticisms of VE are considered alongside discussion of its implications for clinical practice with people who have intellectual disability. PMID- 10711646 TI - Changes in skills for people with intellectual disability: a follow-up of the Camberwell Cohort. AB - The skills of a total population of children with severe intellectual disability and/or autism from Camberwell, South London, UK, and the initial follow-up data, taken when the subjects were adolescents and young adults (Shah 1986), are described in the present study. Changes in skills over time are presented within the categories of communication, self-care, and educational and cognitive skills, as assessed by the Handicaps, Behaviours and Skills schedule. The results indicated that skills had improved in many areas between times 1 and 2, but that this improvement was more noticeable for the children who had been youngest at time 1. The implications of these results and predictions for a further follow-up study are discussed. PMID- 10711647 TI - Intellectual characteristics of Prader-Willi syndrome: comparison of genetic subtypes. AB - Advances in genetics have led to an increased understanding of the role of the genotype on behavioural functioning. The purpose of the present study was to examine differences in intellectual functioning in individuals with Prader-Willi syndrome (PWS) with a paternal 15q11-q13 deletion versus maternal uniparental disomy (UPD) of chromosome 15. Measures of intelligence and academic achievement were administered to 38 individuals with PWS (24 with deletion and 14 with UPD). The subjects with UPD had significantly higher verbal IQ scores than those with deletion (P< 0.01). The magnitude of the difference in verbal IQ was 9.1 points (69.9 versus 60.8 for UPD and deletion PWS subjects, respectively). Only 17% of subjects with the 15q11-q13 deletion had a verbal IQ > or = 70, while 50% of those with UPD had a verbal IQ > or = 70. Performance IQ scores did not differ between the two PWS genetic subtype groups. This is the first report to document the difference between verbal and performance IQ score patterns among subjects with PWS of the deletion versus the UPD subtype. PMID- 10711648 TI - Strategic planning and progress under the All Wales Strategy: reflecting the perceptions of stakeholders. AB - Nominated representatives from the various stakeholder interests, i.e. social services, health, education, voluntary organizations, parent groups and self advocacy groups, involved in the implementation of the All Wales Strategy for the development of services for people with intellectual disability were interviewed 2 years after the end of the initial 10-year phase. Interviewees were asked to reflect on the strengths and weaknesses of policy implementation, including: changing priorities, planning arrangements, agency roles, central guidance and financial mechanisms, consumer participation, and the impact of more recent policy or structural developments. Despite recognition of the leadership of the Welsh Office, the shift in thinking achieved, the developments made in joint agency collaboration and in consumer participation in planning, and an increasing competence to plan effectively over time, the overriding perception was that more could have been made of the opportunity afforded by the clearest and best resourced central government policy within the UK in this area. At the heart of this judgement lay concerns about pragmatic rather than strategic planning, a failure to link annual service developments to a final comprehensive end point and a related failure to integrate planning to meet community needs with hospital resettlement Factors which may have contributed to these weaknesses are discussed, as are lessons for subsequent community care policy. PMID- 10711649 TI - Needs for oral care among people with intellectual disability not in contact with Community Dental Services. AB - Previous research has found an unmet need for oral care among people with intellectual disability. The key factors which have been indicated are low expectations, fear of treatment, lack of awareness among carers and problems in accessing dental services. The withdrawal of many general dental practitioners (GDPs) from the National Health Service (NHS) may have exacerbated the latter problem in the UK. The aims of the present study were: (1) to assess the extent of unmet clinical needs in a group of adults with intellectual disability living in the community who were not in contact with the Community Dental Service (CDS); and (2) to explore their perceptions of teeth and contact with dentists to identify how oral care can be improved. Interviews were completed with subjects and/or carers and a dental examination was completed. There were higher levels of untreated caries (decay), and gingival or periodontal (gum) problems among the sample than in either the general population, or in a previous survey of CDS users at day centres and residential facilities. The subjects were largely unaware of dental problems, and used the appearance and absence of pain to judge the condition of their teeth. They depended greatly on their carers for decision making and support with regard to visiting the dentist and tooth-brushing. Carers requested training in oral care and the use of dental services, and support in dealing with clients who have problems tolerating tooth-brushing. The subjects had experienced a wide variation in the treatment provided by dentists, but had not found it difficult to access a dentist despite recent reductions in the availability of NHS dental care. They expressed a particular need for a good relationship with their dentist and for their dentist to have personal skills in relating to people with an intellectual disability. Dental screening checks and oral care training for carers should be made easily available. Care plans should include tooth-brushing and dietary issues for all clients who have their own natural teeth. There are significant training issues for dentists in developing personal skills in total communication, disability awareness and attitudes which value people with intellectual disability. PMID- 10711650 TI - An investigation into diet treatment for adults with previously untreated phenylketonuria and severe intellectual disability. AB - There is evidence in the literature which suggests that adults with previously untreated phenylketonuria (PKU) benefit from a low phenylalanine diet. A prospective study providing a phenylalanine-restricted diet to five subjects with severe intellectual disability arising from untreated PKU is reported. Physical, social and behavioural measures were used to monitor the effects of the diet Four out of the five subjects derived considerable benefit. It is concluded that the restricted diet is worth trying in most individuals with previously untreated PKU, and that possible benefits are in the areas of concentration, alertness, mood, irritability and adaptive behaviour. PMID- 10711651 TI - Comorbid Down's syndrome, Tourette syndrome and intellectual disability: registry prevalence and developmental course. AB - The co-occurrence of Tourette syndrome (TS) and Down's syndrome (DS) has been previously reported in the literature. In the present study, a retrospective record review was conducted using the North Dakota TS registry in order to ascertain the number of cases of TS and DS, and to develop case descriptions. We identified five cases from North Dakota. Two of these patients were simply comorbid for TS and DS. One was additionally comorbid for bipolar disorder, another for childhood disintegrative disorder and a third had a D/G group translocation. The association between DS and TS occured in 2% of TS patients. Contrary to the situation in patients with pervasive developmental disorders, the presence of TS in DS may be a negative prognostic indicator. PMID- 10711652 TI - Continuity and change in the use of residential services by adults with intellectual disability: the Aberdeen cohort at mid-life. AB - The present paper reports a follow-up study of a representative cohort of people with intellectual disability, now in middle age, who entered adult services on leaving school in the late 1960s, and whose adult years have coincided with a period of radical policy change and considerable service expansion. The present authors focus on the changes which have taken place in their use of residential services over this period. Firstly, the situation in early adulthood when they were last studied is examined. At this time, 60% of the cohort were still living at home, almost all with both parents. The remaining 40% were in institutions, mainly long-stay mental handicap hospitals. Out-of-home placement is associated with gender (maleness), challenging behaviour, and the absence of one or both parents. The present authors compare this with the situation 20 years later. The most significant change in this time is the expansion of the community sector, accounting for about half of the surviving cohort, with numbers still at home or in hospital correspondingly reduced. Almost half of the cohort had experienced little or no change in residential situation. On average, they were less intellectually and socially able than those who had moved into community provision. PMID- 10711653 TI - Pathological and neuropathological findings in two males with fragile-X syndrome. AB - The present paper addresses post mortem pathological and neuropathological findings in two males with fragile-X syndrome, aged 67 and 87 years. Both subjects died from sudden, unexpected cardiovascular causes, and both showed abnormalities of the mitral valve, ventricular hypertrophy and cardiomegaly. Both cases demonstrated macrocephaly characteristic of the classical Martin-Bell phenotype in FRAXA. There was increased brain weight in both cases: macroscopically, both cerebral and cerebellar hemispheres appeared normal, but dilated lateral ventricles were seen; and microscopic examination of the brain in case 2 showed normal hexalaminar architecture and no gross neuronal dropout. The hippocampus showed mild CA4 pyramidal cell loss and associated gliosis. The cerebellum showed focal Purkinje cell loss and corresponding Bergmann gliosis. Whilst there is a need to delineate the microscopic features of fragile-X syndrome from those of the ageing process, there is an urgent need for more systematic neuropathological studies of fragile-X syndrome; the increased brain weight and Purkinje cell loss in autism and fragile-X syndrome reopens the debate on these two conditions. The case for further research into the cardiac anomalies in fragile-X syndrome is also strengthened by the findings. Finally, the present report confirms the role of interstitial cell hyperplasia as the major cause of megalo-testes in this condition. PMID- 10711654 TI - Oral self-mutilation in a patient with rhombencephalosynapsys. AB - Rhombencephalosynapsis (RS) is a rare cerebellar malformation. Its essential features are the absence of the incisura cerebelli posterior, fusion of the cerebellar hemispheres, the absence of the velum medullare anterius and nuclei fastigii, and fusion of the dentate nuclei, which are shifted towards the mid line. Clinically, affected patients present with signs of cerebellar and motor disturbances. The present report describes a new patient affected by RS. The subject first presented at the age of 22 years because of a psychiatric symptomatology which was characterized by obsessive oral self-mutilation associated with an intellectual disability. Objective evaluation documented dysmorphic features, while neurological examination showed only a slight truncal ataxia. The subject's IQ was 74 on the Wechsler Scale (verbal IQ = 79, performance IQ = 74). Psychiatric evaluation with DSM-IV criteria documented an obsessive-compulsive personality disorder associated with emotional instability and oral self-mutilation. The typical picture of rhombencephalosynapsis was evident on magnetic resonance imaging. Both chromosomal analysis and routine biochemical investigations were normal. The relationship between oral self injurious behaviour and cerebellar malformations is discussed with particular regard to the behavioural aspects of cerebellar congenital pathology in affective disorders and in autism. PMID- 10711655 TI - A case of mosaic trisomy 21 with Down's syndrome signs and normal intellectual development. AB - The present case study describes an adult male with clinical signs of mild Down's syndrome (DS), who performed well at school and reached university level. A karyotype was done on a lymphocyte culture and mosaic trisomy 21 was found in 3% of the 437 cells analysed. Eleven signs from Jackson's checklist were found in the clinical evaluation, which along with the analysis of the subject's dermatoglyphic traits, confirmed the DS diagnosis. Cognitive evaluation was done with several psychological tests and the results were within the average range. This rare phenotypic association shows that normal intellectual development may be possible in DS. This finding could be explained by the low trisomic cell frequency, which may have little effect on the critical tissues for intellectual development, and it might also reflect the wide phenotypic variation in mosaic trisomy 21. Other factors, such as strong family support, early and continued intervention programmes for both physical and speech therapy, and a thorough educational process, also provided opportunities for the development of the cognitive potential of the subject. PMID- 10711656 TI - Regulation of glucose transporters--implications for insulin resistance states. AB - Altered glucose homeostasis in the different diabetic states often results from a combination of insulin deficiency (absolute or relative), and impaired hormone action. The latter involves alterations in the expression and/or function of glucose transporters in insulin responsive peripheral tissues - skeletal muscle and adipose tissue. Since whole body glucose utilization depends mainly on controlled changes in glucose transport in these tissues, this review focuses on the role of glucose transporters in the regulation of insulin-stimulated glucose transport activity. The molecular mechanisms by which several inducers of insulin resistance inhibit insulin action on glucose uptake are also discussed. Better understanding of the complex regulation of glucose transport and transporters will hopefully shed light on potential sites for new pharmaceutical interventions. Several excellent reviews have been published in the past 2 years detailing various aspects which are discussed only briefly in this review. They are mentioned in the text to allow further reading. PMID- 10711657 TI - A twelve year study of the incidence of childhood type 1 diabetes mellitus in the Eastern Province of Saudi Arabia. AB - OBJECTIVE: Study of the incidence of childhood type 1 diabetes in the Eastern Province of Saudi Arabia. METHODS: Analysis included all children eligible for care in our hospital who had type 1 diabetes diagnosed before their 15th birthday between 1986 and 1997. RESULTS: A total of 46 children (27 girls and 19 boys) were identified, with a median age at diagnosis of 10.3 yr. The overall age adjusted incidence rate was 12.3 x 10(5)/yr; it was 9.9 x 10(5)/yr for males and 14.8 x 10(5)/yr for females. The number of patients increased significantly (relative risk of 46) over the study period (p=0.01), more significantly in females (p=0.007) and in the 10-14 year age group (p=0.01). CONCLUSIONS: The incidence of type 1 diabetes increased markedly over the past 12 years, mainly in females and children over 10 years of age. The data confirm the need to develop a national registry and the need for further epidemiological research. PMID- 10711658 TI - Leptin binding activity (LBA) in plasma of nondiabetic and diabetic adolescents and obese children: relation to auxologic and hormonal data. AB - Leptin circulates in serum bound to high molecular weight proteins. Hypothesizing that leptin binding proteins may regulate the functional efficiency of leptin, we characterized auxologic and hormonal factors that influence leptin binding in three disparate groups: normal adolescents, obese children, and teenagers with type I diabetes mellitus (IDDM). Specific leptin binding activity (sLBA) was assessed by column chromatography after incubation of serum with 125I-leptin in the presence and absence of excess unlabeled leptin. Mean sLBA was 17.0 +/- 7% (SD) in the healthy adolescents (n=41), 6.6 +/-4.3% in the obese children (n=26), and 14.9 +/-7.3% in the diabetic teenagers (n=17). At any value of sLBA, obese children had higher serum leptin levels than non-obese adolescents or diabetic teenagers, consistent with "leptin resistance" in the obese group. sLBA was higher in males than in females only in those with diabetes (18.6 +/- 7.3 vs 10.9 +/- 5.1%, p<0.05). sLBA correlated inversely with serum insulin-like growth factor-I values in the normal group (r= -0.45, p<0.01) and with insulin in the obese children (r= -0.53, p<0.01). There was no correlation between sLBA or serum leptin values and HbA1c, in the diabetic group. The serum leptin concentration was the principal determinant explaining the total variability of sLBA in all three cohorts. However, body mass index (BMI = weight/ height2) accounted for more of the total variability of percent specific binding in the healthy adolescents than in the other groups. We conclude that sLBA reflects circulating leptin levels, body composition, and hormonal milieu. Thus, in addition to leptin, qualitative and quantitative characteristics of leptin binding may play a physiological role in the regulation of appetite and in the "leptin resistance" of obesity. PMID- 10711659 TI - Breast milk leptin concentrations in initial and terminal milk samples: relationships to maternal and infant plasma leptin concentrations, adiposity, serum glucose, insulin, lipid and lipoprotein levels. AB - Leptin has recently been shown to be present in human milk and is produced by mammary epithelial cells. We studied leptin concentrations in human milk and its relationships with maternal and infant plasma leptin concentrations, adiposity, serum glucose, insulin, lipid and lipoprotein levels. We also compared the initial and terminal milk leptin concentrations to investigate whether leptin acts as a satiety factor. Venous blood samples were obtained from 18 healthy lactating women aged from 17-42 years and their 3-120 day-old infants. Breast milk samples were collected just before and immediately after suckling, when the infant had self-terminated sucking. Leptin mean values in breast milk were lower than in maternal plasma (p<0.001). Breast milk log leptin concentrations positively correlated with both maternal and infant plasma log leptin concentrations (p<0.001 and p=0.001, respectively) and negatively correlated with maternal serum total cholesterol and low-density lipoprotein cholesterol levels (p<0.001 and p<0.01, respectively), but did not correlate with maternal and infant adiposity, serum glucose and insulin levels, maternal serum HDL-C, triglyceride levels and infants' lipid and lipoprotein concentrations (p>0.05). Using stepwise multiple regression analysis, maternal plasma log leptin and serum HDL-C concentrations were related to breast milk log leptin concentration (R2=0.82; p<0.0001 and p<0.001, respectively). There was no significant difference between initial and terminal milk leptin levels (p>0.05). We concluded that maternal leptin may be transferred to the infant via milk and may exert biological effects; there may be factors other than adiposity affecting breast milk leptin levels, and that leptin might not contribute to the development of satiation at the end of suckling. PMID- 10711660 TI - M-mode echocardiographic evaluation of systolic function, LV volume and mass in children on growth hormone therapy. AB - Since abnormal endogenous growth hormone (GH) secretion in adults is associated with cardiac dysfunction, it is important to ensure that GH therapy in children and adolescents does not cause similar effects. Forty-two growth hormone deficient children (Group 1) (19 girls, 23 boys) were evaluated. Six girls and seven boys were prepubertal with a mean age of 6.65 yr (range 4.37-9.73 yr). Twenty-nine were pubertal (13 girls, 16 boys), mean age 13.57 yr (range 10.08 16.76 yr). The patients had been on long-term GH therapy for 34.97 +/- 18.78 months with an average weekly dose of 17.61 IU/m2/wk. The mean height SDS was 2.85 +/- 1.22 for boys and -2.5 +/- 0.64 for girls at the onset of therapy, and at the time of examination -1.8 +/- 1.32 for the boys and 1.87 +/- 0.94 for the girls. Thirty-four normal control subjects (Group 2) matched for age, sex and body size were also studied. Left ventricular volume (LV), mass and systolic function [shortening fraction (FS)] were evaluated by two-dimensional guided M mode echocardiography. Blood pressure was also measured. No differences in blood pressure were observed between patients and controls. There was no correlation of GH dose and duration of therapy with LV measurements. No significant differences were found between Group 1 and Group 2. These observations suggest that long term administration of GH does not produce adverse cardiac effects in GH deficient children. Nevertheless, longer follow-up studies are still needed to confirm the safety of long-term rhGH treatment. PMID- 10711661 TI - Growth hormone treatment in short children with beta-thalassemia major. AB - The effect of one year recombinant human growth hormone (rhGH) treatment on growth rate and bone age was studied in ten short prepubertal children with beta thalassemia major (age range 7.10-12.03 yr) with normal GH response to provocative stimuli. rhGH was given subcutaneously every day in a dose of 28 IU/m2/week. In the 10 children who completed 12 months of treatment the growth velocity increased from 4.22+/-0.81 cm/yr (-1.38+/-0.80 SDS for CA) to 7.61+/ 1.16 cm/yr (+2.27+/-1.64 SDS for CA). IGF-I was low before treatment, 138.3 +/ 38.9 ng/ml, and rose significantly to 232.2+/-122.1, 243.2+/-98.4 and 227.5+/ 86.2 at 3, 6 and 12 months post-treatment, respectively (p<0.01). Bone maturation was accelerated in proportion to the increase in chronological age. The mean pre treatment bone age in the ten children was 8.20+/-1.97 and increased to 9.55+/ 1.80 yr after one year of treatment. Our data demonstrate that GH treatment of thalassemic children with normal GH reserve and low serum IGF-I concentrations with supraphysiological doses of rhGH for one year can cause a significant increase in serum IGF-I levels and growth velocity, but it remains to be elucidated whether long-term administration will affect the final height. PMID- 10711662 TI - Magnetic resonance imaging in growth hormone deficiency: relationship between endocrine function and morphological findings. AB - Magnetic resonance imaging (MRI) using gadopentetate dimeglumine (Gd-DTPA) improves the delineation of hypothalamic-pituitary structures and facilitates the detection of anatomical abnormalities which are indicators of permanent growth hormone deficiency (GHD). The aim of this study was to determine the frequency of neuroradiological abnormalities in 85 (52 M, 33 F) patients with hereditary or idiopathic forms of isolated GHD (IGHD) or multiple pituitary hormone deficiency (MPHD) and also to investigate the relationship between anatomical findings and hormonal status. Pituitary hypoplasia with absent or thin infundibulum and ectopic posterior pituitary (EPP) were the most frequent findings in 39 patients with MPHD, whereas in 46 patients with IGHD the most frequent finding was pituitary hypoplasia without neuroradiological abnormalities. All patients whose infundibulum was not visualized after Gd-DTPA injection belonged to the MPHD group; therefore, absence of pituitary stalk can be a good indicator of the severity of hormonal deficiencies. Pituitary hypoplasia was found in all patients with familial IGHD. Among patients with abnormalities of the hypothalamic pituitary area on MRI, normal or breech delivery frequency distributed equally. Therefore it seems that mechanical or hypoxic prenatal events cannot be the primary etiological factor in all patients with neuroradiological abnormalities since half of these patients had normal delivery and birth history. The localization of the bright spot of the posterior pituitary at the level of the median eminence, midstalk position or at the end of the infundibulum may suggest a neuronal migration defect which may occur during early embryogenesis. In conclusion, in children with GHD a careful examination of the hypothalamic pituitary area by MRI after enhancement helps to establish the diagnosis and predicts the prognosis. PMID- 10711663 TI - Iron deposition in the anterior pituitary in homozygous beta-thalassemia: MRI evaluation and correlation with gonadal function. AB - OBJECTIVE: Iron deposition in the anterior pituitary continues to pose a serious problem in older patients with homozygous beta-thalassemia particularly in terms of gonadal function. This study aimed to investigate whether iron loading within the pituitary correlated with endocrine function. PATIENTS: 33 patients above 15 years of age, with transfusion-dependent homozygous beta-thalassemia and iron overload were studied. All had been receiving deferoxamine since 1978. DESIGN AND MEASUREMENTS: The endocrine status of the patients was assessed on clinical examination by an endocrinologist, and by a gonadotropin releasing hormone stimulation test. MRI of the pituitary was carried out for each patient. RESULTS: Anterior pituitary function (GnRH stimulation test) correlated well with MRI results. However, no correlation was found between the MRI measurements, the GnRH stimulation test and the clinical status of the patients, as 28 out of the 33 patients achieved normal puberty. CONCLUSIONS: MRI in conjunction with a GnRH stimulation test may be useful in predicting future impairment of pituitary function; however, further studies are needed to assess the effect of chelation therapy on the iron overload in the gland. PMID- 10711664 TI - Geographic variation in groundwater iodine and iodine deficiency in Israel, The West Bank and Gaza. AB - BACKGROUND: Iodine deficiency during pregnancy and infancy is the world's most common preventable cause of mental retardation. Previous studies have shown a high incidence of goiter and low groundwater iodine concentrations in northern Israel. OBJECTIVE: We examined the relationship between low groundwater iodine and iodine deficiency in pregnant women and schoolchildren. SUBJECTS AND METHODS: We measured the urinary iodine excretion of school-children in the West Bank and Gaza and rural and urban pregnant women in Western Galilee (an area known to have low groundwater iodine concentrations). We also measured iodine concentrations in groundwater in various locations in the West Bank and Gaza. RESULTS: Lower urinary iodine excretion was found among pregnant Arab women living in rural Western Galilee (101+/-7 microg iodine/g creatinine). 20% of them excreted <50 microg I/g creatinine. This is relatively less than found among pregnant Jewish women living in cities in the same area (154+/-13 microg I/g creatinine). Low iodine concentrations (<5 microg/l) were found in groundwater in the Nablus, Ramallah, Bethlehem highlands, as compared to normal concentrations in the lowland districts of the West Bank and Gaza. In a cohort of 728 schoolchildren aged 8-10, 10% (range 8-13%) of children from areas of low groundwater iodine had low levels of urinary iodine excretion, as compared to only <5% of those from districts with groundwater iodine concentrations >10 microg/l. CONCLUSIONS: Lower concentrations of groundwater iodine are related to low urinary iodine excretion in Israel, the West Bank and Gaza. PMID- 10711665 TI - The effect of abnormal intrauterine thyroid hormone economies on infant cognitive abilities. AB - OBJECTIVE: To evaluate how intrauterine and neonatal thyroid hormone deficiencies affect infant cognitive abilities. METHOD: 26 infants with intrauterine or neonatal thyroid hormone deficiency and 20 full-term infants with normal thyroid economies were studied at 6 months of age or corrected age. Reasons for thyroid hormone deficiency were maternal hypothyroidism, maternal hyperthyroidism treated with antithyroid medication, congenital hypothyroidism, and low-risk prematurity. A computer-generated task during which infants' eye-movements were videotaped was used to assess attention, memory, and learning abilities RESULTS: Data from transcribed videotapes showed the study group was significantly less attentive and had longer reaction times than controls but did not differ on indices of sustaining attention or learning. Within thyroid-deficient groups, offspring of treated hyperthyroid mothers showed an atypical profile suggestive of hypervigilance. CONCLUSION: A decreased fetal or maternal thyroid hormone supply in pregnancy is associated with infants' poorer attention and altered rates of information processing. PMID- 10711666 TI - Neuroendocrinological response to standardized mixed meal in female anorectic patients during active and refeeding phases. AB - To evaluate the neuroendocrinological dysfunction in anorexia nervosa, plasma somatostatin, glucose, insulin, and growth hormone were monitored in ten patients with anorexia nervosa in the active and refeeding (remission) phases of the disorder and in nine age-matched healthy control subjects. Somatostatin levels were significantly higher in the anorectic patients in both the active and refeeding phases than in the controls at baseline (mean+/-SD 27.4 +/-5.5 and 31.1+/-2.6 vs 21.3+/-1.9 pg/ml; p<0.001), and significantly higher in the anorectic patients in the active phase compared to the refeeding phase and to the controls in response to a mixed meal (p<0.05). Insulin levels were significantly lower in the anorectic patients in both the active and refeeding phases compared to the controls at baseline (9.3+/-1.1, 7.6+/-1.0 vs 14.7+/-3.5 microU/ml; p<0.0001) and after a mixed meal (p<0.05). An attenuated glucose response discriminated the anorectic patients in the active state from the same patients in the refeeding state and the controls (p<0.0001). There was no significant difference in growth hormone response between the anorectic patients and the controls. These findings suggest that there is an augmented response of somatostatin and an attenuated response of insulin to mixed meal stimulation in active anorexia. The diminished insulin response persists during the refeeding phase. It seems that central and peripheral alterations in endocrine function occur in anorexia nervosa. PMID- 10711667 TI - Pre-operative control of arterial hypertension using ketoconazole in pediatric patients with adrenocortical tumors. AB - Adrenocortical tumors are rare in childhood, appearing more frequently in some regions such as South and South-eastern regions of Brazil and India. Common clinical signs include virilization, Cushing's syndrome, feminization and hypertension, either isolated or in association. The aim of this report is to present our experience with the pre-operative use of ketoconazole in children with an adrenocortical tumor to control elevated blood pressure levels non responsive to the usual treatment. Over the last 16 years, of 46 children diagnosed as having adrenocortical tumor, 17 developed hypertension (diastolic pressure greater than the 95th percentile for age and sex according to data from the Task Force on Blood Pressure Control in Children), associated with virilization and/or Cushing's syndrome. In three of these 17 patients, conventional antihypertensive therapy failed, and they were treated with ketoconazole (200-300 mg/day). This resulted in rapid control of the blood pressure. It is concluded that in selected patients, ketoconazole may be useful adjuvant therapy for the palliative control of the arterial hypertension secondary to adrenocortical tumors, without side effects. PMID- 10711668 TI - Correlation of blood-spot 17-hydroxyprogesterone daily profiles and urinary steroid profiles in congenital adrenal hyperplasia. AB - OBJECTIVE: To compare the value of blood-spot 17-hydroxyprogesterone (17-OHP) daily profiles and urinary steroid excretion in untreated and treated patients with congenital adrenal hyperplasia (CAH). PATIENTS: Ten patients with CAH were investigated during steroid replacement therapy (Group 1), and 11 patients were investigated without treatment (Group 2). METHODS: Capillary blood samples were collected for measurement of blood-spot 17-OHP values by non-chromatographic radioimmunoassay. Steroid profiles of 24-h urine samples were analyzed by gas chromatography. RESULTS: There was a close correlation between the individual daily means of blood-spot 17-OHP measurements and the pregnanetriol/ tetrahydrocortisone ratio in both groups of patients (Group 2: r=0.839, p<0.001; Group 1: r=0.686, p<0.001). Almost the same correlation was found between the blood-spot 17-OHP value and the sum of three 17-hydroxyprogesterone metabolites/the sum of three cortisol/cortisone metabolites ratio (Group 2: r=0.918, p<0.001; Group 1: r=0.741, p<0.001). CONCLUSIONS: Blood-spot 17-OHP measurements and 24-h urinary steroid profile have the same impact in identification and monitoring therapy of children with CAH. PMID- 10711669 TI - Patient with Weismann-Netter and Stuhl (toxopachyosteosis) syndrome with communicant hydrocephalus and arachnoid cyst. AB - Weismann-Netter and Stuhl (toxopachyosteosis) syndrome is a rare bone disease, manifesting with anterior bowing of the tibiae, short stature and mild mental retardation. We report a patient with Weismann-Netter and Stuhl syndrome with an unusual manifestation communicant hydrocephalus and arachnoid cyst. PMID- 10711670 TI - Effect of alendronate treatment on the clinical picture and bone turnover markers in chronic idiopathic hyperphosphatasia. AB - Chronic idiopathic hyperphosphatasia (CIH), also known as juvenile Paget's disease, is characterized by increased bone turnover, persistently elevated serum alkaline phosphatase concentrations and progressive bone deformities. The pathogenesis of the disease is unknown. Currently, there is no specific treatment and agents that reduce bone turnover have been tried in some cases with limited success. In this report, we present our experience with alendronate treatment in a 17 year-old boy with CIH. Ten weeks of treatment with alendronate resulted in marked clinical improvement and normalization of serum alkaline phosphatase activity. Serum osteocalcin and urinary deoxypyridinoline levels were decreased approximately 50% compared to pretreatment values, indicating decreased bone turnover rate. Alendronate seems to be a promising and safe agent for treatment of CIH. PMID- 10711671 TI - Retesting of patients with isolated growth hormone deficiency (IGHD) using provocative tests and reevaluation of them after termination of therapy. PMID- 10711672 TI - Cytoskeletal involvement in the regulation of aqueous humor outflow. PMID- 10711673 TI - Photodynamic therapy with verteporfin (Visudyne): impact on ophthalmology and visual sciences. PMID- 10711674 TI - A novel locus for Leber congenital amaurosis (LCA4) with anterior keratoconus mapping to chromosome 17p13. AB - PURPOSE: A two-generation consanguineous Pakistani family with autosomal recessive Leber congenital amaurosis (LCA, MIM 204,000) and keratoconus was identified. All affected individuals have bilateral keratoconus and congenital pigmentary retinopathy. The goal of this study was to link the disease phenotype in this family. METHODS: Genomic DNA was amplified across the polymorphic microsatellite poly-CA regions identified by markers. Polymerase chain reaction (PCR) products were separated by nondenaturing polyacrylamide gel electrophoresis. Alleles were assigned to individuals, which allowed calculation of LOD scores using the Cyrillic and MLINK software program. The retinal guanylate cyclase (RETGC-1, GDB symbol GUC2D) and pigment epithelium-derived factor (PEDF) genes were analyzed by heteroduplex analysis and direct sequencing for mutations in diseased individuals. RESULTS: Based on a whole genome linkage analysis the first locus for this combined phenotype has been mapped to chromosome 17p13. Linkage analysis gave a two point LOD score of 3.21 for marker D17S829. Surrounding this marker is a region of homozygosity of 15.77 cM, between the markers D17S1866 and D17S960; however, the crossover for the marker D17S1529 refines the region to 10.77 cM within which the disease gene is predicted to lie. Mutation screening of the nearby RETGC-1 gene, which has been shown to be associated with LCA1, revealed no mutations in the affected individuals of this family. Similarly, another prime candidate in the region PEDF was also screened for mutations. The factor has been shown to be involved in the photoreceptor differentiation and neuronal survival. No mutations were found in this gene either. Furthermore, RETGC-1 was physically excluded from the critical disease region based on the existing physical map. CONCLUSIONS: It is therefore suggested that this combined phenotype maps to a new locus and is due to an as yet uncharacterized gene within the 17p13 chromosomal region. PMID- 10711675 TI - Comparison of PCR detection methods for B1, P30, and 18S rDNA genes of T. gondii in aqueous humor. AB - PURPOSE: Comparison of polymerase chain reaction (PCR) amplification of three Toxoplasma gondii genes in aqueous humor. METHODS: Nested PCRs carried out using published methods were optimized for maximum sensitivity and specificity. Five pairs of oligonucleotide primers, directed against the B1, P30, and ribosomal genes, were used and compared to determine which sequences were most effective in detecting T. gondii DNA. Methods were developed with DNA templates in water and were subsequently applied to both normal and inflamed aqueous. RESULTS: After one round of PCR amplification, P30 and ribosomal primers were able to detect 1 pg genomic T. gondii DNA. However, those directed against the B1 gene were able to detect 50 fg (approximately single tachyzoite). This level of sensitivity was also achieved using the P30 primers after a second round of PCR; however, only primers based on the B1 gene maintained this level of sensitivity in both normal and inflamed aqueous. B1-specific primers did not amplify sequences from fungal, bacterial, or human lymphocyte DNA. The sensitivity of T. gondii detection using B1 gene-specific primers was not compromised when large amounts of human lymphocyte DNA were present, and application to an ocular sample or retinal section from patients with toxoplasma chorioretinitis was successful in confirming the presence of T. gondii DNA. CONCLUSIONS: The B1 PCR protocol appears to be the most sensitive protocol in the detection of T. gondii DNA and has been successful in identification of T. gondii DNA in ocular fluids and retinal sections. This provides direct evidence of the presence of T. gondii within the eye and may therefore help in the management of toxoplasma retinochoroiditis. PMID- 10711676 TI - Fas mediates apoptosis and oxidant-induced cell death in cultured hRPE cells. AB - PURPOSE: To investigate whether Fas ligand (FasL) and the Fas receptor system mediates apoptosis in cultured human retinal pigment epithelial (hRPE) cells and contributes to oxidant-induced death of hRPE cells. METHODS: Expression of FasL and Fas in cultured hRPE cells was examined by Western blot analysis and flow cytometry. The susceptibility of hRPE cells to Fas-mediated apoptosis was determined by incubating cells with recombinant soluble Fas ligand (sFasL). Characteristics of apoptosis assessed included chromatin condensation, DNA cleavage, and phosphatidylserine exposure. To investigate the possible involvement of Fas-mediated apoptosis in oxidative killing of hRPE cells, the effects of the oxidant tert-butylhydroperoxide (tBH) on the expression of FasL and Fas were studied. The specificity of effects of oxidant was examined using the antioxidants glutathione and N-acetyl-L-cysteine (NAC). The requirement for the Fas pathway in tBH-induced apoptosis was investigated using an antagonistic anti-Fas antibody ZB4 that blocks the interaction between FasL and Fas. RESULTS: Cultured hRPE cells constitutively expressed FasL and Fas. Ligation of Fas receptor with recombinant sFasL triggered apoptosis in hRPE cells. tBH treatment of hRPE cells resulted in increased expression of FasL and Fas. Glutathione and NAC completely abrogated tBH-induced increase in FasL and Fas expression and apoptosis. Blocking FasL and Fas interaction by ZB4 inhibited tBH-induced apoptosis, but only partially. CONCLUSIONS: A functional Fas-mediated apoptotic pathway is present in cultured hRPE cells and can be activated with sFasL or by upregulation of FasL and Fas expression with an oxidant. The incomplete inhibition by blocking antibody indicates that the Fas pathway is involved in oxidant-induced apoptosis, but other triggering mechanisms are also important. PMID- 10711677 TI - Novel mutations in the TULP1 gene causing autosomal recessive retinitis pigmentosa. AB - PURPOSE: To assess the contribution of TULP1 to autosomal recessive retinitis pigmentosa (arRP). METHODS: Fifteen exons of the gene were screened by single strand conformation polymorphism analysis of 7 (of 49) arRP pedigrees showing cosegregation with TULP1 locus markers. RESULTS: In one of the seven families two allelic mutations, IVS4-2delAGA and c.937delC, were found in exons 5 and 10, respectively. CONCLUSIONS: Two novel mutations in TULP1 were found to be associated with arRP. That they both compromise the gene product supports their pathogenicity. This gene was present in no more than 2% of a panel of 49 Spanish families affected by arRP. PMID- 10711678 TI - Cell cycle kinetics in corneal endothelium from old and young donors. AB - PURPOSE: To compare cell cycle kinetics in corneal endothelial cells from young and old donors. METHODS: Human corneas were obtained from the eye bank and separated into two groups: young (19 corneas, <30 years of age) and old (40 corneas, >50 years of age). Corneas were cut in quarters, and the endothelium was released from contact inhibition by producing a 2-mm scrape wound. Unwounded endothelium acted as a negative control. Corneal pieces were exposed for 24, 36, 48, 60, 72, and 84 hours to medium containing 10% fetal bovine serum, 20 ng/ml fibroblast growth factor, and 50 mg/ml gentamicin or the same medium supplemented with 10 ng/ml epidermal growth factor (EGF). Tissue was fixed, immunostained for Ki67 (a marker for the late G1-through M-phase) or for 5-bromo-2'-deoxyuridine (BrdU; a marker for the S-phase), and mounted in medium containing propidium iodide (PI) to visualize all nuclei. Confocal images were evaluated using an image analysis program to count Ki67-positive and PI-stained cells and to evaluate cell cycle position. Cells were counted in 15x100 microm2 areas randomly selected from each wound, and the mean was used for subsequent calculations. RESULTS: Human corneal endothelial cells could be reliably scored for their position within the cell cycle using Ki67 staining patterns. In both age groups, cells repopulating the wound area stained positively for Ki67, whereas no Ki67 staining was observed in unwounded areas under any condition tested. Cells from old donors treated with fetal bovine serum and FGF stained positively for Ki67, indicating that these cells were actively cycling. Compared with cells from young donors, old cells entered the cell cycle more slowly (48 versus 36 hours), the peak of Ki67 staining occurred later (72 versus 60 hours), and fewer cells proliferated (23% versus 47%) or exhibited mitotic figures (4% versus 7%). Addition of EGF to the culture medium increased Ki67 staining in both groups, but the effect on old cells was more dramatic. More cells from old donors entered the cell cycle by 36 hours after wounding, the number of proliferating cells increased 1.6-fold, and the relative number of mitotic figures increased 2.5-fold over cells treated in the absence of EGF. CONCLUSIONS: Regardless of donor age, corneal endothelial cells can enter and complete the cell cycle. In the presence of fetal bovine serum and FGF, cells from old donors can proliferate but respond more slowly and to a lesser extent than cells from young donors. EGF added to the medium stimulates cells from old donors to enter the cell cycle faster, increases the relative number of actively cycling cells, and increases the number of cells exhibiting mitotic figures. The resultant hypothesis is that it is possible to stimulate a significant proliferative response in corneal endothelial cells from old individuals. Administration of an optimal combination of stimulatory growth factors is required under conditions in which cells have been transiently released from contact inhibition. PMID- 10711679 TI - Expression of estrogen receptor alpha and beta in the mouse cornea. AB - PURPOSE: To test the possibility that estrogen has a direct effect on corneal cells, the possible occurrence of estrogen receptor alpha (ERalpha) and beta (ERbeta) in the cornea of mice was examined. METHODS: To test for the occurrence of ER proteins in the cornea of mice, an immunocytochemical method was used. To test for the occurrence of ER mRNAs in the cornea of mice, reverse transcription polymerase chain reaction (RT-PCR) was used. RESULTS: Immunocytochemical examination revealed that both ERalpha and ERbeta exist in the cell nuclei of corneal epithelial, stromal, and endothelial cells of both male and female mice. RT-PCR revealed that RNAs of ERs occur in the cornea of both male and female mice. CONCLUSIONS: Because ERalpha and ERbeta occur in corneal cells of mice, estrogen may exert biological functions in corneal cells through direct interaction with these ERs. PMID- 10711680 TI - Expression of MMPs and TIMPs in human pterygia and cultured pterygium epithelial cells. AB - PURPOSE: Pterygia are a common, benign, fibrovascular, and infiltrative process of the corneal-conjunctival junction of unknown pathogenesis. Matrix metalloproteinases (MMPs) are a family of proteolytic enzymes active against all components of the extracellular matrix, whose activity is specifically neutralized by tissue inhibitors of MMPs (TIMPs). In the current study the hypothesis was that MMPs and TIMPs may actively participate in the formation and progression of pterygia. METHODS: In this study, 25 pterygium specimens and 15 normal conjunctival biopsies obtained from subjects undergoing surgery for glaucoma and cataract, were processed for immunohistochemistry or in situ hybridization. Pterygium epithelial cells (PECs) were cultured under serum-free conditions and exposed to proinflammatory cytokines to determine both the mRNA and protein expression profiles of MMPs and TIMPs. RESULTS: Collagenase-1 and gelatinase A were expressed in all pterygia examined, specifically localized to the epithelium (directly adjacent to collagen type III), with gelatinase B expression exclusively associated with neutrophils. No collagenase-1 or gelatinase A was detected in normal conjunctiva. TIMP-1 and -3 were localized to epithelial cells with additional TIMP-3 immunoreactivity detected in the extracellular matrix, endothelial cells and leukocytes of all diseased tissue. TIMP-3 protein was evident in 4 of 15 normal conjunctiva. Induction of collagenase-1, gelatinase A, and TIMP-1 mRNA and protein was demonstrated in epithelial cells treated with tumor necrosis factor-alpha and interleukin-1alpha, whereas TIMP-3 expression was unaltered. CONCLUSIONS: This is the first study to document the cellular expression of MMPs and TIMPs in pterygia and cultured human PECs. MMPs and TIMPs may contribute to the inflammation, tissue remodeling, and angiogenesis that characterize pterygia. Understanding the role these proteins play may lead to novel therapies intended to reduce the progressive nature of pterygia. PMID- 10711681 TI - The use of atomic force microscopy for the observation of corneal epithelium surface. AB - PURPOSE: To evaluate the feasibility of imaging normal corneal epithelium by means of atomic force microscopy (AFM). METHODS: Twelve normal corneas from six albino rabbits were examined using a commercial atomic force microscope. Six corneas were examined in balanced salt solution after fixation in glutaraldehyde 2.5% and six without any fixation. Rectangular silicon nitride cantilevers with a spring constant of 10 to 20 mN/m were used. The measured forces after imaging were less than 100 pN. All reported images were made with 512x512-pixel definition with typical scan rates ranging from 1 to 5 Hz. RESULTS: High-quality images of corneal epithelium surface were obtained from fixed and unfixed specimens in magnifications ranging from x2000 to x2,000,000. Imaging of fixed specimens was always easier. In unfixed specimens fuzzy images were very common, probably because of the presence of the cell glycocalyx. AFM revealed the typical polygonal corneal epithelial cells. The cell surface was covered by microprojections; at cell borders the microprojections were arranged in two characteristic parallel rows. Craterlike formations were revealed in several specimens. The microprojections' morphology and their surface details were revealed using magnifications up to x2,000,000. Three-dimensional representation of the images facilitated better understanding of the surface topography. Measurements in horizontal and vertical plane were made using the section analysis tool. CONCLUSIONS: In this work the AFM parameters appropriate for corneal epithelium imaging in physiological medium were defined. AFM represents a new powerful tool for corneal epithelium imaging, and its application in this field warrants further investigation. PMID- 10711682 TI - Key factors in the subjective and objective assessment of conjunctival erythema. AB - PURPOSE: To establish objectively measurable characteristics of the conjunctival vasculature that correspond with the judgment of erythema by human observers. METHODS: Color images of bulbar conjunctiva from 21 subjects were digitally analyzed to extract the following variables characteristic of the scene: vessel width (W), number of vessels (V), proportion of area occupied by vessels (PA), relative redness both in vessels (RRV) and in the whole image (RRI), red-green difference both in vessels (RGV) and in the whole image (RGI), red-blue difference both in vessels (RBV) and in the whole image (RBI), and red hue value (RHV). These data were compared with subjective judgments by a panel of seven trained observers who independently rated erythema in the same images, using a 0 to 4 scale with decimal interpolation between grades. RESULTS: Correlation analysis indicated significant associations (P<0.05) between the mean response of the human observers and all the objective variables except RHV. Associations with the morphometric variables PA (R2 = 0.93) and V(R2 = 0.90) were markedly stronger than for the best colorimetric variable RBV (R2 = 0.62). CONCLUSIONS: Judgments of erythema made by human observers do not rely primarily on color but can be closely approximated by a univariate, linear model involving only the proportion of the scene occupied by vessels. Under the conditions of this study, grading of erythema by trained observers can be considered to constitute measurement to at least an interval level. PMID- 10711683 TI - Neurotrophic factors in the human cornea. AB - PURPOSE: To investigate neurotrophic growth factors and corresponding receptors in human and rabbit corneal epithelium and stroma. METHODS: Transcription of nerve growth factor (NGF), neurotrophin 3 (NT-3), NT-4, brain-derived neurotrophic factor (BDNF), glial cell line- derived neurotrophic factor (GDNF), and receptors Trk A-E, was investigated by reverse transcription-polymerase chain reaction. DNA dot blot analysis allowed to estimate transcription levels. Single cell proliferation assays were performed using recombinant NGF, BDNF, and GDNF. Mitogen-activated protein kinase signal transduction was investigated with Western blot analysis using antibodies against activated and total extracellular signal-regulated kinase (ERK) 1/2 and the jun N-terminal protein kinase (JNK) 1/2. RESULTS: Transcription of NGF, NT-3, BDNF, and Trk A, Trk B, Trk C, and Trk E receptors was detected in both ex vivo and cultured epithelium and stroma. Transcription of NT-4 was only detected in epithelium and transcription of GDNF only in stroma. Levels of transcription were higher for NT-3, NT-4, and the Trk receptors and lower for NGF, BDNF, and GDNF. NGF and GDNF stimulated both epithelial colony formation and proliferation, whereas BDNF only enhanced colony formation. Stromal proliferation was enhanced in serum-free medium. In epithelium, predominantly ERK 1 was activated by NGF, GDNF, and BDNF. In stromal cells NGF and GDNF stimulated phosphorylation of ERK 1 and JNK 1. CONCLUSIONS: Neurotrophic factors and tyrosine kinase receptors are transcribed in the human cornea. GDNF and NGF stimulate corneal epithelial proliferation, and the effect of the latter might be mediated by activation of ERK 1. Neurotrophic factors have very specific effects on phosphorylation of ERK and JNK in epithelial and stromal cells. The differential expression of NT-4 and GDNF suggests a regulatory function within the cytokine network of the cornea. PMID- 10711684 TI - Quantification of MUC2 and MUC5AC transcripts in human conjunctiva. AB - PURPOSE: Transcripts of mucins 1, 4, and 5AC have been identified in human conjunctival tissue. Of these, only MUC5AC has been localized to goblet cells. MUC2 is a goblet cell mucin originally identified in the intestinal mucosa. The presence of MUC2 transcripts and levels of MUC2 and MUC5AC transcripts in normal human conjunctiva, as determined by quantitative polymerase chain reaction (PCR), is reported. METHODS: RNA from conjunctivae of six donors (3 men, 3 women, 44 to 69 years, all white) was isolated and subjected to competitive reverse transcription-PCR. Internal standards, which are dsDNA molecules with ends complementary to a given primer pair but containing nonhomologous central sequences, were prepared for each gene assayed. Titration of a constant amount of cDNA against serial dilutions of the internal standard allowed quantification of the template cDNA. MUC2 and MUC5AC levels were compared to levels of the "housekeeping" gene, beta2-microglobulin (beta2M). The identity of PCR products was confirmed by sequencing. RESULTS: In the six individual samples tested, beta2M mRNA is expressed, on average, at approximately 10(-20) moles per sample (1 microg RNA) or approximately 63.5x10(4) molecules. The mRNA encoding MUC5AC, a relatively abundant ocular mucin, exists at levels 10-fold lower than beta2M. In contrast to previous reports of MUC2 mRNA being absent at the ocular surface, these results show that MUC2 transcripts are present and expressed at levels 5900 fold lower than for MUC5AC. Apparently, MUC2 transcripts exist on the order of only approximately 100 to 1000 molecules/microg of RNA in the analyzed samples. CONCLUSIONS: MUC2 transcripts are present in human conjunctival tissue and their abundance is much lower than that of MUC5AC. This is the first application of competitive PCR to the quantitative analysis of mucin expression in human ocular tissue. The sensitivity of this method allows the detection of MUC2 transcripts that were not detected by Northern blot analysis or in situ hybridization in previous studies. It also makes possible the comparison of relative levels of expression for ocular mucins. PMID- 10711685 TI - Reflex lacrimation in patients with glaucoma and healthy control subjects by fluorophotometry. AB - PURPOSE: Steady state tear turnover (TTO), defined as TTO under normal physiological conditions, is significantly lower in patients with untreated glaucoma than in healthy control subjects. To obtain more information on the effect of glaucoma on lacrimation, a method for quantification of reflex lacrimation was developed and applied to patients with glaucoma or ocular hypertension and healthy control subjects. METHODS: After instillation of 2 microl of fluorescein (2%), the decay of fluorescein concentration in tears was measured by fluorophotometry over 10 minutes to determine steady state TTO. Then, reflex lacrimation was induced by stimulating the trigeminal nerve with ethanol vapor via the nostrils. Thereafter, the decay of fluorescein and corresponding steady state TTO were determined again. An index of reflex lacrimation, defined as the percentage decrease in fluorescein concentration as a result of stimulation, was calculated by forward and backward extrapolation of the steady state decay of the fluorescein concentration in tears, relative to the time of stimulation. RESULTS: The index of reflex lacrimation was determined in 16 patients with newly discovered but not yet treated glaucoma, 16 patients with untreated ocular hypertension, and 16 healthy control subjects. The values did not differ between groups (mean +/- SD, 67.0%+/-17.7%, 63.5%+/-21.3%, and 70.4%+/ 19.6%, respectively; ANOVA, P>0.25). Surprisingly, the steady state TTO after stimulation was lower than that before stimulation in each group (ratio, 0.62+/ 0.46; paired t-test, P<0.04). CONCLUSIONS: The method developed is appropriate for the quantification of reflex lacrimation. Reflex lacrimation is not influenced significantly by glaucoma or ocular hypertension. The decreased steady state TTO after reflex stimulation may be caused by exhaustion of the lacrimal glands after excessive reflex lacrimation, indicating that normal lacrimation probably also contains reflex tears. PMID- 10711686 TI - Rotation of Listing's plane with convergence: independence from eye position. AB - PURPOSE: To determine whether asymmetrical vergence results in a rotation of Listing's plane independent of vergence-associated changes of eye position in the orbit. METHODS: Six normal subjects were required to fixate on a 3x3 array (40 degrees on a side) of light-emitting diodes arranged on a flat screen 124 cm from the subject. Disparity-induced vergence was elicited with a horizontal Fresnel prism (30 cm/m, approximately 17 degrees) placed in front of one eye. In four subjects accommodative vergence (10 degrees to 15 degrees) was produced by placing a minus spherical lens in front of one eye while the other eye was covered. Eye position was measured binocularly using three-axis search coils. Control data were collected without prisms during monocular and binocular viewing. For all data a planar regression was used to fit torsional eye position as a function of horizontal and vertical position to calculate the horizontal and vertical primary positions that define the orientation of Listing's plane. RESULTS: In the prism experiment, the horizontal primary position of the eye not wearing the prism rotated temporally by 3.9 degrees +/-1.7 degrees compared with the both eyes viewing control condition. The rotation of the prism eye was in a similar range (3.4 degrees +/-2.0 degrees). With accommodation, the horizontal primary position of the viewing eye rotated temporally by 4.4 degrees +/-1.4 degrees compared with the monocular viewing control. In both the accommodation and the prism paradigms there was usually a rotation of vertical primary position downward. CONCLUSIONS: Vergence-induced changes in Listing's plane can be independent of changes in orbital position associated with vergence. This finding supports a role for changes in central innervation in the elaboration of Listing's law. PMID- 10711687 TI - Effects of betaxolol on light responses and membrane conductance in retinal ganglion cells. AB - PURPOSE: To examine the physiological effects of betaxolol, a beta1-adrenergic receptor blocker commonly used in the treatment of glaucoma, on retinal ganglion cells and to evaluate its potential to elicit responses consistent with a neuroprotective agent against ganglion cell degeneration. METHODS: Single-unit extracellular recording, electroretinogram (ERG), intracellular and whole-cell patch-clamp recording techniques were made from flatmounted, isolated retina, superfused eyecup, and living retinal slice preparations of the larval tiger salamander. RESULTS: Bath application of 20 microM betaxolol reduced the glutamate-induced increase of spontaneous spike rate in retinal ganglion cell by approximately 30%. The glutamate-induced postsynaptic current recorded under voltage-clamp conditions was reduced by 50 microM betaxolol, and the difference current-voltage (I-V) relation (I(Control)-I(betaxolol)) was N-shaped and AP5 sensitive, characteristic of N-methyl-D-aspartate receptor-mediated current. Application of 50 microM betaxolol reversibly reduced the voltage-gated sodium and calcium currents by approximately one third of their peak amplitudes. The times-to-action of betaxolol on ganglion cells are long (15-35 minutes for 20-50 microM betaxolol), indicative of modulation through slow biochemical cascades. Betaxolol, up to 100 microM, exerted no effects on horizontal cells or the ERG, suggesting that the primary actions of this beta1 blocker are restricted to retinal ganglion cells. CONCLUSIONS: These physiological experiments provide supporting evidence that betaxolol acts in a manner consistent with preventing retinal ganglion cell death induced by elevated extracellular glutamate or by increased spontaneous spike rates under pathologic conditions. The physiological actions of betaxolol lead to reducing neurotoxic effects in ganglion cells, which are the most susceptible retinal neurons to glutamate-induced damages under ischemic and glaucomatous conditions. Therefore, betaxolol has the potential to be a neuroprotective agent against retinal degeneration in patients with disorders mediated by such mechanisms. PMID- 10711688 TI - Localization of myocilin/trabecular meshwork--inducible glucocorticoid response protein in the human eye. AB - PURPOSE: To study distribution and cellular localization of myocilin/trabecular meshwork-inducible glucocorticoid response protein (TIGR) in the human eye. METHODS: A peptide antibody against a portion of the myosin-like domain of myocilin/TIGR was developed. Different ocular tissues from three human donors were investigated by one- and two-dimensional gel electrophoresis and Western blot analysis. Immunohistochemistry was performed on 25 human eyes enucleated because of posterior choroidal melanoma and on 7 normal human donor eyes. RESULTS: By Western blot analysis, a band at approximately 57 kDa was visualized in cornea, trabecular meshwork, lamina cribrosa, optic nerve, retina, iris, ciliary body, and vitreous humor. By immunohistochemistry, immunoreactivity for myocilin/TIGR was observed in cells of the corneal epi- and endothelium and extracellularly in the corneal stroma and sclera. In the trabecular meshwork, cells of the uveal and corneoscleral meshwork were stained, as was the cribriform area directly adjacent to Schlemm's canal. Positive staining was seen in cells of the ciliary epithelium, ciliary muscle, lens epithelium, and in stromal and smooth muscle cells of the iris. Throughout the entire vitreous body, fine filamentous material was positively labeled. In the retina, staining was seen along the outer surface of rods and cones, in neurons of the inner and outer nuclear layer, and in the axons of optic nerve ganglion cells. Optic nerve axons were stained in the prelaminar, laminar, and postlaminar parts of the nerve. In the region of the lamina cribrosa, astrocytes in the glial columns and cribriform plates were positively labeled. CONCLUSIONS: Myocilin TIGR is expressed in almost every ocular tissue. Depending on the respective tissue, it is observed extra- or intracellularly. The presence of myocilin/TIGR in optic nerve axons and lamina cribrosa astrocytes indicates that the trabecular meshwork might not be the only target of abnormal myocilin/TIGR in GLC1A-linked open-angle glaucoma. PMID- 10711689 TI - Number of ganglion cells in glaucoma eyes compared with threshold visual field tests in the same persons. AB - PURPOSE: To compare the number of retinal ganglion cells (RGCs) topographically mapped with specific visual field threshold test data in the same eyes among glaucoma patients. METHODS: Seventeen eyes of 13 persons with well-documented glaucoma histories and Humphrey threshold visual field tests (San Leandro, CA) were obtained from eye banks. RGC number was estimated by histologic counts of retinal sections and by counts of remaining axons in the optic nerves. The locations of the retinal samples corresponded to specific test points in the visual field. The data for glaucoma patients were compared with 17 eyes of 17 persons who were group matched for age, had no ocular history, and had normal eyes by histologic examination. RESULTS: The mean RGC loss for the entire retina averaged 10.2%, indicating that many eyes had early glaucoma damage. RGC body loss averaged 35.7% in eyes with corrected pattern SD probability less than 0.5%. When upper to lower retina RGC counts were compared with their corresponding visual field data within each eye, a 5-dB loss in sensitivity was associated with 25% RGC loss. For individual points that were abnormal at a probability less than 0.5%, the mean RGC loss was 29%. In control eyes, the loss of RGCs with age was estimated as 7205 cells per year in persons between 55 and 95 years of age. In optic nerves from glaucoma subjects, smaller axons were significantly more likely to be present than larger axons (R2 = 0.78, P<0.001). CONCLUSIONS: At least 25% to 35% RGC loss is associated with statistical abnormalities in automated visual field testing. In addition, these data corroborate previous findings that RGCs with larger diameter axons preferentially die in glaucoma. PMID- 10711690 TI - S-nitrosoglutathione photolysis as a novel therapy for antifibrosis in filtration surgery. AB - PURPOSE: To determine whether a novel peroxynitrite-based photosensitizer S nitrosoglutathione (GSNO) can produce specific in vitro light-induced cell death of both standard animal lung and human Tenon's capsule (TC) fibroblasts and to compare this effect with that produced by the established photodynamic porphyrin precursor 5-aminolevulinic acid (ALA). METHODS: V79-4 Chinese hamster lung and human TC fibroblasts were established in tissue culture. GSNO, together with its radioactive tritiated and fluorescent dansylated derivatives, were synthesized. The labeled molecules were prepared to determine the time course of uptake into the fibroblasts. Uptake was monitored by scintillation counting for the tritiated GSNO and confocal fluorescence microscopy for the dansylated GSNO. The uptake of ALA and biosynthesis of its photosensitive product were determined by fluorescence emission spectroscopy of a separate set of fibroblasts. Once uptake was established, both cell lines were incubated with varying concentrations of GSNO or ALA as a function of time (0, 4, or 24 hours) before light exposure (200 msec pulsed visible light, 0.068 W per pulse, for 10 minutes at a distance of 10 cm). After 10 minutes of irradiation, the cells were washed and exposed to fresh tissue culture medium. The effect of the treatment was determined 24 hours later by measuring cell viability. RESULTS: A 2-minute drug treatment time (0 hours incubation) with GSNO, followed by 10 minutes of irradiation, resulted in approximately 78% of fibroblast cell death at the lowest concentration of GSNO used compared with the control, which was exposed to light, but no GSNO. The higher concentrations of GSNO, or longer drug treatment times before irradiation, did not statistically increase cell death. Maximal cell death was thus obtained using the lowest GSNO concentration (50 mM) and drug treatment time (2 minutes). In contrast, the well-established photosensitizer ALA killed only approximately 4% of cells at the lowest concentration and drug treatment time tested. At drug treatment times of 4 hours and less, increased concentrations of ALA did not produce cell death of more statistical significance. It was not until 24 hours of drug treatment that comparable amounts of cell death were produced by ALA and GSNO. In all experiments similar results were obtained with the animal lung and human TC fibroblasts, suggesting that the source of the fibroblast had no effect on the outcome. The differences in treatment effects between GSNO and ALA were statistically significant under all conditions tested. CONCLUSIONS: GSNO is able to cause light-specific cell death of human TC fibroblasts at drug treatment times (2 minutes) and irradiation times (10 minutes) that would be compatible with its use in glaucoma filtering surgery. This in vitro performance was superior to that of the well-established photosensitizer ALA, which required treatment times longer than 4 hours to approach the light-specific cell death produced by only 2 minutes of GSNO treatment. PMID- 10711691 TI - TGF-beta1, -beta2, and -beta3 in vitro: biphasic effects on Tenon's fibroblast contraction, proliferation, and migration. AB - PURPOSE: To compare the effects of the three human transforming growth factor beta (TGF-beta) isoforms and different concentrations of TGF-beta on human Tenon's capsule fibroblasts (HTF), with a view to delineating the role of this growth factor in the subconjunctival scarring response after glaucoma filtration surgery. METHODS: Application of recombinant human TGF-beta1, -beta2, and -beta3 (range 0-10(-8) M) was assessed using several assays of HTF function: fibroblast mediated collagen contraction, proliferation, and migration. RESULTS: All three isoforms of TGF-beta behaved in a similar manner in vitro. They each stimulated HTF-mediated collagen contraction, proliferation, and migration with a characteristic concentration-dependent response, with peak activities at 10(-9), 10(-12), and 10(-9) M, respectively, that were significantly different from control (P<0.05). At concentrations above and below peak activities, HTF activity was reduced, demonstrating biphasic effects of TGF-beta. CONCLUSIONS: TGF-beta1, beta2, and -beta3 have similar actions in vitro; this is demonstrated by their effects on several HTF-mediated functions. TGF-beta induces a response in HTF that is concentration-dependent, with different functions being maximally stimulated at different concentrations. This biphasic response highlights the significance of the concentration profile of TGF-beta at the wound site. These findings are important in filtration surgery, where constant changes in the local environment occur due to the passage of aqueous and the wound healing process. The varying levels of TGF-beta in the aqueous and subconjunctival tissues may thus significantly modify the conjunctival scarring response. PMID- 10711692 TI - Obstructed axonal transport of BDNF and its receptor TrkB in experimental glaucoma. AB - PURPOSE: In both animal model system and in human glaucoma, retinal ganglion cells (RGCs) die by apoptosis. To understand how RGC apoptosis is initiated in these systems, the authors studied RGC neurotrophin transport in experimental glaucoma using acute intraocular pressure (IOP) elevations in rats and chronic IOP elevation and unilateral optic nerve transections in monkeys. METHODS: Eyes were studied in masked fashion by light and electron microscopy and by immunohistochemistry with antibodies directed against the tyrosine kinase receptors (TrkA, B, and C) and against brain-derived neurotrophic factor (BDNF), as well as by autoradiography to identify retrograde axonal transport of 125I BDNF injected into the superior colliculus. RESULTS: With acute glaucoma in the rat, RGC axons became abnormally dilated, accumulating vesicles presumed to be moving in axonal transport at the optic nerve head. Label for TrkB, but not TrkA, was relatively increased at and behind the optic nerve head with IOP elevation. Abnormal, focal labeling for TrkB and BDNF was identified in axons of monkey optic nerve heads with chronic glaucoma. With acute IOP elevation in rats, radiolabeled BDNF arrived at cells in the RGC layer at less than half the level of control eyes. CONCLUSIONS: Interruption of BDNF retrograde transport and accumulation of TrkB at the optic nerve head in acute and chronic glaucoma models suggest a role for neurotrophin deprivation in the pathogenesis of RGC death in glaucoma. PMID- 10711693 TI - Technique for detecting serial topographic changes in the optic disc and peripapillary retina using scanning laser tomography. AB - PURPOSE: To describe and evaluate a new statistical technique for detecting topographic changes in the optic disc and peripapillary retina measured with confocal scanning laser tomography. METHODS: The 256x256-pixel array of topographic height values obtained with each image from the Heidelberg Retina Tomograph (Heidelberg Engineering, Heidelberg, Germany) was divided into an array of 64x64 superpixels, where each superpixel contained 16 (i.e., 4x4) pixels. An analysis of variance technique was developed to analyze each superpixel with three baseline and three follow-up images. The performance of the technique was tested with and without adjustment for spatial correlation of topographic values using computer simulations and with real data from a normal control subject and a patient with progressive glaucomatous disc change. RESULTS: Computer simulation with fixed population means and variance, and varying spatial correlation showed a monotonically increasing number of superpixels with significant test results (false positives), with 20% false-positives when the spatial correlation was 0.8 (the approximate median value in real patient data). The number of false-positive results was similar (17%) in serial images of a normal subject. When corrected for spatial correlation, the number of false-positives was independent of the level of spatial correlation and remained at the expected value of less than 5% in both simulations and real data. Although the number of significant test results in the patient with progressive glaucoma decreased after correction for spatial correlation, the change was readily apparent. Statistical power to detect mean differences in topographic values ranging from 0.5 to 4.0 SDs in computer simulation showed low power for changes of 1 SD or less, but increased dramatically with larger changes. CONCLUSIONS: This technique has a high level of sensitivity to detect changes in the optic disc while maintaining a high level of specificity. PMID- 10711694 TI - Adenovirus keratitis: a role for interleukin-8. AB - PURPOSE: Adenovirus type 19 (Ad19) infection of the human cornea results in a chronic, multifocal, subepithelial keratitis. Existing evidence suggests that early subepithelial corneal infiltrates are composed of polymorphonuclear neutrophils. In this study, the capacity of Ad19-infected human corneal stromal fibroblasts (HCFs) to produce neutrophil chemotactants (chemokines) was tested. METHODS: HCFs grown from human donor corneas and passaged thrice were infected with a corneal isolate of Ad19 or mock-infected with virus-free media. Bioactivity of the cell supernatants was tested by a neutrophil chemotaxis assay. Supernatants were assayed by enzyme-linked immunosorbent assay for the neutrophil chemotactants interleukin-8 (IL-8) and GRO-alpha. Corneal facsimiles were generated with HCFs and collagen type I, infected with Ad19, and assayed by immunohistochemistry. RESULTS: Ad19 infection of HCFs increased neutrophil chemotaxis from a baseline of 0.4+/-0.7 cells/high-powered field (hpf; mock infected) to 21.8+/-2.3 cells/hpf (Ad19-infected). Chemotaxis was reduced by the addition of neutralizing antibodies against IL-8 and GRO-alpha. Infection of HCFs induced quantities of IL-8 protein 300- and 1000-fold over mock-infected controls at 4 and 24 hours, respectively (33 versus 11,813 pg/mL at 4 hours, and 57 versus 76,376 pg/mL at 24 hours, P< or = 0.001 for both). In contrast, GRO-alpha protein levels were only sevenfold higher at 24 hours postinfection (118 pg/mL in mock infected controls versus 880 pg/mL in Ad19-infected cell supernatants). Neither chemokine was induced by infection of an immortalized human corneal epithelial cell line. Immunohistochemistry of infected corneal facsimiles demonstrated IL-8 in the extracellular matrix within 3 days after infection. CONCLUSIONS: Production of chemokines in infected tissues facilitates an early innate immune response to infection, and in the infected corneal stroma represents an elementary defense mechanism. Interleukin-8 may play a role in the development of subepithelial infiltrates in adenovirus keratitis. PMID- 10711695 TI - ACAID induced by allogeneic corneal tissue promotes subsequent survival of orthotopic corneal grafts. AB - PURPOSE: To determine whether immune deviation is induced by allogeneic corneal tissue implanted in the anterior chamber and whether survival of subsequent orthotopic corneal allografts is thereby enhanced. METHODS: Corneal tissue from C57BL/6 mice was implanted in the anterior chamber of eyes of BALB/c mice. The fate of these implants was assessed histologically, and the donor-specific immune response of recipient mice was tested for donor-specific delayed hypersensitivity and the capacity to accept or reject C57BL/6 corneas grafted orthotopically into the fellow eye. RESULTS: C57BL/6 cornea implants in the anterior chamber failed to induce donor-specific delayed hypersensitivity but impaired donor-specific delayed hypersensitivity in a proportion of recipients with implants in place for 2 weeks. Mice with donor-specific delayed hypersensitivity rejected the intraocular implants. Mice bearing C57BL/6 cornea implants in the anterior chamber for 2 (but not 4) weeks accepted the C57BL/6 corneas grafted orthotopically into the fellow eye at a high rate and with few rejection reactions. CONCLUSIONS: Implantation of allogeneic corneal tissue in the anterior chamber of the eye has the transient capacity to alter the recipient alloimmune response in a manner that promotes survival of subsequent orthotopic corneal allografts. PMID- 10711696 TI - Kinetics of apoptotic cells in experimental autoimmune uveoretinitis. AB - PURPOSE: To investigate the role of apoptosis in immunopathogenic mechanisms of experimental autoimmune uveoretinitis (EAU), the kinetics of apoptotic cells and expression of Fas and Fas ligand (FasL) in the eye with EAU were studied. METHODS: Male inbred Lewis rats were immunized with S-antigen (40 microg/rat), and eyes were examined to detect apoptotic cells on days 1, 4, 8, and 10 post immunization and days 0, 2, 4, 6, and 8 after the onset of EAU. The clinical and pathologic scores were used for estimating EAU. Apoptotic cells were analyzed by TdT-mediated dUTP nick-end labeling, electron microscopic and immunohistologic examinations, and agarose gel electrophoresis. The anti-rat Fas and anti-rat FasL antibodies were used to examine the expression of Fas and FasL. RESULTS: Apoptotic cells were detected in the infiltrating cells in the aqueous humor, the vitreous body, the iris-ciliary body, and the retina. Apoptotic cells were observed as early as the day of EAU onset and reached a peak on day 2 after the disease onset. Fas and FasL were expressed on the infiltrating cells in the aqueous humor and the vitreous. FasL was expressed on resident cells of the ciliary body. The kinetics of the expression of FasL corresponded with the kinetics of apoptotic cells. CONCLUSIONS: Fas-FasL-mediated apoptosis is considered to occur in the eye with EAU and plays a role in the immunopathogenic mechanisms to eliminate ocular infiltrating cells, thereby down-regulating the inflammatory processes. PMID- 10711697 TI - Increased severity of Pseudomonas aeruginosa corneal infection in strains of mice designated as Th1 versus Th2 responsive. AB - PURPOSE: Mice favoring Th1 (C57BL/6, C57BL/10, and B10.D2/nSn) versus Th2 (BALB/c, BALB/cBy, BALB.B, and BALB.K) response development were evaluated for their response to infection with Pseudomonas aeruginosa. This study addresses the question of whether Th1 versus Th2 response propensity affects the pathogenesis of bacterial keratitis in mice. METHODS: Ocular disease was determined by mean clinical score, slit lamp, plate counts, and histopathology, and antigen-specific cellular responses were assessed by immunostaining and measurement of delayed type hypersensitivity (DTH). RESULTS: Strains of mice favoring Th1 (B6, BL10, and B10.D2) versus Th2 (BALB/c, BALB/cBy, BALB.B, and BALB.K) responsiveness were infected with P. aeruginosa. Mice favoring Th1 response development exhibited a similar course of disease and the infected eyes of all mice perforated by 7 days postinfection (p.i.). Strains (BALB/c, BALB/cBy, BALB.B, and BALB.K) favoring Th2 response development exhibited a milder course of disease, and none of the infected corneas perforated at 7 days p.i. In a Th1-responsive strain (B10.D2), positive immunostaining for CD4+ and CD8+ T cells was observed in the cornea by 3 days p.i. and by 5 days p.i., respectively, some cells stained positively for IL2 R, indicating that the cells were activated. In contrast, in a Th2 responder strain (BALB/c), there was no detectable positive immunostaining in cornea for any of the T-cell markers tested and DTH was significantly elevated in B10.D2 versus BALB/c mice. CONCLUSIONS: These studies are the first to provide evidence that in P. aeruginosa ocular infection, mouse strains favoring development of a Th1-type response are susceptible (cornea perforates), whereas strains favoring Th2 response development are resistant (no corneal perforation). PMID- 10711698 TI - Participation of pigment epithelium of iris and ciliary body in ocular immune privilege. 1. Inhibition of T-cell activation in vitro by direct cell-to-cell contact. AB - PURPOSE: To determine by what mechanism(s) iris and ciliary body (I/CB) pigment epithelial (PE) cells inhibit T-cell activation in vitro. METHODS: Pure cultured I/CB PE cells were obtained from eyes of normal and CD95 ligand (CD95L)deficient mice and tested for their capacity to suppress T-cell activation in three different T-cell receptor (Tcr) ligand systems: mixed lymphocyte reactions, stimulation of Tcr transgenic T cells (D011.10) by specific antigen (ovalbumin), and ligation of the Tcr-associated CD3 molecule by anti-CD3 antibodies. Proliferation and secretion of cytokines (interferon [IFN]-gamma, interleukin [IL]-2, IL-4, and IL-10) were assessed as measures of T-cell activation. Suppressive influences of I/CB PE cells were determined on the basis of RT-PCR- detected cytokine genes expressed by I/CB PE cells, immunosuppression mediated by supernatants of cultured I/CB PE cells, direct contact between I/CB PE cells and T lymphocytes, and promotion of apoptosis among responding T cells. Attempts to reverse I/CB PE-dependent suppression of T-cell activation included the use of neutralizing antibodies to IL-10, tumor necrosis factor (TNF)-alpha and transforming growth factor (TGF)-beta, and the addition of exogenous IL-2 and IL 12. RESULTS: Cultured mouse I/CB PE cells (including CD95L-deficient cells), which were more than 95% keratin positive, suppressed T-cell proliferation and secretion of IFN-gamma, IL-2, IL-4, and IL-10 in a dose-dependent fashion in all three Tcr ligand systems. Supernatants of cultured I/CB PE cells displayed little suppression activity, whereas cultures in which I/CB PE cells contacted responding T cells directly were profoundly immunosuppressive. Cultured I/CB PE cells expressed mRNA for TGF-beta1, TGF-beta2, IL-6, IL-10, and TNF-alpha, but not IL-4, IFN-gamma, proopiomelanocortin (POMC), and CD95L (Fas L). Antibodies to TGF-beta, IL-10, and TNF-alpha failed to reverse suppression mediated by I/CB PE cells. Moreover, neither exogenous IL-2 or IL-12 relieved the suppression. CONCLUSIONS: Cultured I/CB PE cells, through direct cell-to-cell contact, prevent T cells from proliferating and secreting cytokines when stimulated through the Tcr for antigen by a mechanism that does not involve CD95 or apoptosis. PMID- 10711699 TI - Polymerase chain reaction for detection of Mycobacterium tuberculosis in epiretinal membrane in Eales' disease. AB - PURPOSE: Tuberculous etiology has been suggested in Eales' disease. Because epiretinal membrane (ERM) is formed on the inner surface of the retina in Eales' disease, it could be the most appropriate intraocular specimen for investigation. Therefore, a nested polymerase chain reaction (nPCR), which detects MPB64 gene of Mycobacterium tuberculosis on the archival specimens of ERM of well-documented Eales' and non-Eales' patients, was applied and the results compared. METHODS: nPCR technique was standardized, and the sensitivity and specificity of the primers were determined. nPCR technique was applied to tissue sections obtained from formalin-fixed and paraffin-embedded tissues of ERM from 23 patients with Eales' disease and 27 noninfective and non-Eales' disease patients as controls. RESULTS: nPCR technique was specific for M. tuberculosis genome and sensitive enough to detect 0.25 fg (corresponding to the presence of a single bacillus). Eleven (47.8%) ERM of 23 Eales' disease and 3 (11.1%) of 27 controls were positive for M. tuberculosis genome. The difference between the two groups was statistically significant (P = 0.001), indicating association of this bacterium with Eales' disease. CONCLUSIONS: The demonstration of the presence of M. tuberculosis DNA by nPCR technique in significant number of ERM of Eales' disease compared with the controls further emphasizes the probable role of this bacterium in the pathogenesis of this enigmatic clinical condition. PMID- 10711700 TI - Th1 versus Th2 immune responses in autoimmune lacrimal gland disease in MRL/Mp mice. AB - PURPOSE: In MRL/Mp-lpr/lpr (MRL/lpr) and MRL/Mp-+/+ (MRL/+) mice, a T-cell-driven lacrimal gland inflammation spontaneously develops that is a model for Sjogren's syndrome. The lacrimal gland lesions in these mice were evaluated by immunohistochemistry for the relative contributions of T-helper (Th)1 versus Th2 immune responses. METHODS: Frozen sections of lacrimal glands from MRL/lpr and MRL/+ mice ages 1 through 5 months were stained with monoclonal antibodies to the cytokines interferon (IFN)-gamma and interleukin (IL)4 and to the cell surface costimulatory molecules B7-1 and B7-2, which are associated with Th1 and Th2 responses, respectively. RESULTS: The median proportion of cells staining for IL 4 ranged from 30% to 67% over time for MRL/lpr mice and from 30% to 55% for MRL/+ mice. The median proportion of cells staining for IFN-gamma ranged from 1% to 5% for MRL/lpr mice and from 0% to 3% for MRL/+ mice. The proportion of cells staining positively for IL-4 was significantly greater than for IFN-gamma in both MRL/lpr (mean difference, 33%; P = 0.0001) and MRL/+ mice (mean difference, 42%; P = 0.0002). The median proportion of cells staining positively for B7-2 ranged from 20% to 38% for MRL/lpr mice and from 16% to 34% for MRL/+ mice. The median proportion of cells staining for B7-1 ranged from 2% to 10% for MRL/lpr mice and from 2% to 5% for MRL/+ mice. The proportion of cells staining positively for B7 2 was significantly greater than for B7-1 for both MRL/lpr mice (mean difference, 15%; P = 0.001) and for MRL/+ mice (mean difference, 19%; P = 0.006). CONCLUSIONS: On the basis of immunohistochemistry for cytokines and costimulatory molecules, inflammatory lacrimal gland lesions in MRL/lpr and MRL/+ mice appear to be a largely Th2 phenomenon. PMID- 10711701 TI - Development and characterization of an H2O2-resistant immortal lens epithelial cell line. AB - PURPOSE: To determine how nature would protect lens epithelial cells from H2O2 stress, an immortal murine lens epithelial cell line, alphaTN4-1, was subjected to gradually increasing H2O2 levels over a period of approximately 8 months. The resultant conditioned cells grew normally when exposed daily to 125 microM H2O2, whereas normal cells died within 9 hours. This communication describes changes in the cell biology of the conditioned cells that allowed them to remain viable. The manner in which critical biochemical parameters were affected in both conditioned and normal cells is also reported. METHODS: Conditioned cells were obtained by gradually increasing the concentration of H2O2 over a period of approximately 8 months, introducing an aliquot of H2O2 every 24 hours. A wide spectra of biological parameters were evaluated, including catalase, GSH peroxidase and other antioxidative enzyme activities, cell number and cell viability, non protein thiol, ATP, transport systems, thymidine incorporation, and DNA cleavage. RESULTS: Surprisingly, the conditioned cells did not degrade the medium H2O2 more rapidly than normal cells. However, analyses of the antioxidative defenses indicated that catalase activity was increased 60-fold and glutathione peroxidase (GSH Px) approximately 2.7-fold. Glucose-6-phosphate dehydrogenase, GSH S transferase, and GSSG reductase also had increased activity. Using one dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis, in situ trypsin digestion and matrix-assisted laser desorption/ionization mass spectrometry, a highly amplified doublet in the conditioned cell preparation was shown to be GSH S-transferase alpha-1 and alpha-2 isomers. Examination of key biochemical parameters indicated that while most such parameters in the conditioned cells showed marked decay in the first hour or so after stress, recovery was then observed and within a few hours, these parameters were back in the normal range. In contrast, damage in the normal cells was not repaired. The damage to DNA was shown to involve Fenton chemistry. In the presence of a metal ion chelator, normal cells survive H2O2 stress. CONCLUSIONS: The overall conclusion from this investigation is that nature has chosen to respond to the H2O2 stress by not only increasing the activity of enzymes degrading H2O2 but also the systems involved in repair, generation of reducing potential, and detoxification. All but one system of those evaluated appears to be permanently modified. PMID- 10711702 TI - Phosphatidylinositol 3-kinase in bovine lens and its stimulation by insulin and IGF-1. AB - PURPOSE: To identify and characterize phosphatidylinositol 3-kinase (PI-3K) in the lens and to study its involvement as a signal mediator in lens epithelial cells exposed to insulin and insulin-like growth factor (IGF)-1, which are known to induce lens epithelial cell proliferation and differentiation into fiber cells. METHODS: Concentric fiber cell layers from single bovine lens were prepared by dissolution in buffer. PI-3K activity in capsule-epithelium and fiber cell layers was determined after immunoprecipitation with antibodies against p85, the regulatory subunit of PI-3K. High-performance liquid chromatography on an ion exchange column (Partisil-SAX; Whatman, Maidstone, United Kingdom) was used to identify PI-3K reaction products. Cultured bovine lens epithelial cells were stimulated with insulin or IGF-1, and PI-3K activity was determined after immunoprecipitation with antibody against phosphotyrosine. Association of p85 with other proteins after stimulation was determined in anti-p85 immunoprecipitates by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western immunoblot analysis using anti-phosphotyrosine antibody. RESULTS: PI-3K activity was found in both lens epithelial cells and fiber cells. The highest specific activity was found in the capsule-epithelium, but there was considerable activity in other fiber cell layers. Insulin and IGF-1 stimulated the PI-3K activity in epithelial cells in culture by more than 100%, and activation of the enzyme resulted in tyrosine phosphorylation of the p85 subunit. After stimulation, the p85 subunit of PI-3K was associated with 100- and 180-kDa tyrosine phosphorylated proteins. CONCLUSIONS: The activation of PI-3K and its association with specific tyrosine-phosphorylated proteins may be important in insulin and IGF-1 signal transduction in lens epithelial cells. The presence of significant PI-3K activity throughout the lens further suggests that this signal transduction enzyme is sustained in fiber cells. PMID- 10711703 TI - PKC isoenzymes in the chicken lens and TPA-induced effects on intercellular communication. AB - PURPOSE: Because lens connexins are phosphoproteins and intercellular communication between lens cells may be modulated by connexin phosphorylation, experiments were designed to characterize the expression of protein kinase C (PKC) isoenzymes in the chicken lens and in lentoid-containing cultures and to study the effects of 12-O-tetradecanoylphorbol-13-acetate (TPA) treatment on the distribution of PKC isoenzymes and intercellular communication. METHODS: The presence and distribution of PKC isoenzymes were studied by immunoblot analysis and immunofluorescence in chicken lens sections and in cell cultures under control conditions and after treatment with TPA. Intercellular communication was assessed by transfer of microinjected Lucifer yellow. RESULTS: PKC alpha, gamma, iota, epsilon, and mu were detected in lens homogenates by immunoblot analysis. The levels of PKC alpha, gamma, iota, and mu decreased between the 7th and the 18th embryonic days. Levels of PKC epsilon remained relatively constant during the period of study. Similarly, lens cells in culture expressed isoenzymes alpha, gamma, epsilon, iota, and mu. PKC beta was not detected in lens or culture homogenates. In lens sections, all PKC isoenzymes analyzed were present in epithelial cells, in the annular pad region, and in the posterior aspect of fiber cells. The anti-PKC gamma antibody also stained fiber cell membranes. Analysis of lentoid cultures by immunofluorescence revealed that PKC gamma, epsilon, and iota and minimal amounts of PKC alpha were present in lentoid cells. Treatment with 200 nM TPA for 15 to 30 minutes induced translocation of PKC gamma to the plasma membrane of lentoid cells and significantly reduced the transfer of microinjected Lucifer yellow. CONCLUSIONS: Several PKC isoenzymes are expressed by lens cells in situ and in culture. The gamma isoenzyme, present in lens fibers, was activated in lentoid cells by TPA, a known activator of PKC. We have previously demonstrated TPA-induced phosphorylation of the gap junction protein connexin56 (Cx56). The new data presented in the current study demonstrate that TPA treatment also decreased intercellular communication. Taken together, the results suggest that differential phosphorylation of Cx56 by PKCgamma may induce a conformational change in the protein which, in turn, might lead to channel closure. PMID- 10711704 TI - Three-dimensional organization of primary lens fiber cells. AB - PURPOSE: To visualize the three-dimensional organization of primary lens fiber cells. METHODS: The gene for Green Fluorescent Protein (GFP) was introduced into the lens vesicle using two different vector systems: a replication deficient adenovirus or an expression plasmid. Injected embryos were allowed to develop for several days and then were examined by confocal microscopy. RESULTS: Injection of either vector resulted in GFP expression in primary fiber cells. GFP-expressing cells were heterogeneous in shape and length. Some regions of the fibers were varicose, with diameters >10 microm; regions between the varicosities were often extremely thin, with diameters of <2 microm. No differences in the morphologies of GFP-expressing cells were noted between adenovirus and plasmid-injected lenses, suggesting that the irregular, undulating, appearance of the primary fibers was not the result of viral infection. Three-dimensional reconstruction of primary fiber cells revealed that, by E6, the posterior tips of the fibers had detached from the lens capsule. The anterior fiber tips remained in contact with the overlying epithelium for 1 to 2 additional days, demonstrating that the formation of the anterior and posterior sutures was asynchronous. CONCLUSIONS: The three-dimensional cellular organization of GFP-expressing cells is consistent with previous analyses of fiber cell morphology in the embryonic nucleus of adult human and bovine lenses. The present data confirm that the disorganized appearance of primary fiber cells observed in adult lenses is largely a reflection of developmental processes rather than a consequence of aging. PMID- 10711705 TI - Endothelin-1 contributes to hyperoxia-induced vasoconstriction in the human retina. AB - PURPOSE: There is evidence that ocular blood flow strongly depends on arterial oxygen tension. Results from recent animal studies indicate that the vasoconstrictor response to hyperoxia may be mediated in part by an increased production of endothelin (ET)-1. In an effort to answer the question whether the retinal vasoconstrictive response to hyperoxia in humans is mediated through ET 1, changes in ocular hemodynamics induced by 100% O2 breathing were studied in the absence and presence of an ET(A) receptor antagonist (BQ-123). METHODS: The study was a randomized, placebo-controlled, double-masked, balanced, three-way crossover design. On separate study days 15 healthy male subjects received infusions of BQ-123 (either 60 microg/min or 120 microg/min) or placebo. The effects of BQ-123 or placebo on hyperoxia-induced (100% O2 breathing) changes in retinal and pulsatile choroidal blood flow were assessed with the blue-field entoptic technique and with laser interferometric measurement of fundus pulsation, respectively. RESULTS: During baseline conditions, hyperoxia caused a decrease in retinal blood flow between -29% and -34% (P<0.001) and a decrease in fundus pulsation amplitude between -7% and -8% (P<0.001). BQ-123 dose dependently blunted the response to hyperoxia in the retina (60 microg/min: -25%, 120 microg/min: -20%; P = 0.003), but not in the choroid. CONCLUSIONS: These results indicate that ET-1 contributes to hyperoxia-induced retinal vasoconstriction in the human retina. PMID- 10711706 TI - Functional characterization of organic cation drug transport in the pigmented rabbit conjunctiva. AB - PURPOSE: To characterize carrier-mediated organic cation drug transport in the rabbit conjunctiva. METHODS: The transport of [14C]guanidine, the model substrate, in the excised pigmented rabbit conjunctiva was evaluated in the modified Ussing chamber. Tetraethylammonium (TEA) transport also was investigated to determine substrate specificity. RESULTS: The apparent permeability coefficient for guanidine and TEA in the mucosal-to-serosal (ms) direction was 5.4 and 49.6 times greater than that in the serosal-to-mucosal (sm) direction, respectively. Guanidine transport in the ms (but not sm) direction revealed temperature and concentration dependency over 0.02 to 10 mM with an apparent Michaelis-Menten constant of 3.1 mM and a maximal flux of 11.4 nmol/(cm2 x h). Net guanidine transport measured at 0.1 mM across the conjunctiva was decreased by 71% or 82%, respectively, on the addition of 1 microM valinomycin (a K+ ionophore) in both bathing fluids or in a high K+ buffer in the mucosal fluid. Interestingly, net guanidine transport was reduced, rather than enhanced, by 63% upon acidifying the mucosal bathing fluid. By contrast, net guanidine transport was not affected by the serosal presence of 0.5 mM ouabain (a Na+, K+-ATPase inhibitor), by the mucosal and serosal presence of 0.1 microM monensin (a Na+ ionophore) or 0.3 microM carbonyl cyanide p-(trifluoromethoxy)phenyl-hydrazone (FCCP, a H+ ionophore). Guanidine transport in the ms direction was polyspecific, as indicated by the 48% to 82% inhibition by structurally diverse amines. In particular, guanidine ms transport was inhibited by the antiglaucoma drugs dipivefrine (72%), brimonidine (70%), and carbachol (78%). CONCLUSIONS: A carrier mediated organic cation transport process appears to exist in the conjunctiva, mediating the absorption of organic amines, including certain amine-type ophthalmic drugs. This process may be driven by an inside-negative apical membrane potential difference. PMID- 10711707 TI - Laser treatment and the mechanism of edema reduction in branch retinal vein occlusion. AB - PURPOSE: To test a hypothesis on the physiological mechanism of the disappearance of macular edema after laser treatment. The hypothesis is based on the effect grid laser treatment has on retinal oxygenation and hemodynamics. It predicts that laser-induced reduction of macular edema is associated with shortening and narrowing of retinal vessels in patients with branch retinal vein occlusion (BRVO). METHODS: The study included 12 subjects, treated with argon laser photocoagulation for BRVO and macular edema. Fundus photographs taken at the time of diagnosis and again after laser treatment, were digitized, and diameter and segment length of retinal vessels was measured using NIH-Image program. RESULTS: Macular edema disappeared or was dramatically reduced in all cases after laser treatment. The diameter of occluded venules constricted to 0.81+/-0.02 (mean +/- SD, P = 0.019) of the prelaser diameter and adjacent arterioles constricted to 0.78+/-0.01 (P = 0.008). The laser treatment also led to shortening of the affected vessels. The final segment length of the occluded venules was 0.95+/ 0.17 (P = 0.005) of the length before treatment. The corresponding value for the adjacent arterioles is 0.95+/-0.14 (P = 0.008). Control arterioles and venules in the same fundus did not change in either length or width. CONCLUSIONS: These results do not reject the authors' hypothesis that the disappearance of macular edema in BRVO can be explained by the effect the laser photocoagulation has on retinal oxygenation. Increased oxygenation causes vessel constriction and shortening and lower intravascular pressure, which reduces edema formation according to Starling's law. PMID- 10711708 TI - Nonadrenergic, noncholinergic relaxation of bovine iris sphincter: role of endogenous nitric oxide. AB - PURPOSE: To investigate the role of endogenously generated nitric oxide (NO) in the relaxation of bovine iris sphincter. METHODS: Isolated bovine sphincters were mounted on an isometric tension apparatus. Contraction-relaxation response was elicited by electrical field stimulation (ES; 12 Hz, 50-msec duration, 70-80 V). Relaxation was arbitrarily defined as maximal decrease of tension below prestimulation baseline after cessation of ES. We also determined the tissue levels of cyclic guanosine monophosphate (cGMP) by radioimmunoassay. RESULTS: ES produced a biphasic response: contraction followed by relaxation. After cessation of ES, the muscle relaxed to below the initial baseline tension. Tetrodotoxin (TTX) abolished most of the contraction and all the relaxation response. Atropine blocked most of the contraction component, leaving the relaxation component unchanged. Prazosin and bupranolol (alpha1-adrenergic and beta-adrenergic antagonists, respectively) also did not affect the relaxation component of the response. Neither substance P nor its antagonist (N-acetyl-L-tryptophane 3,5-bis (trifluoromethyl)-benzyl ester; ATTB) inhibited or mimicked the response. The nitric oxide synthase (NOS) inhibitors Nomega-nitro-L-arginine methyl ester (L NAME), Nomega-nitro-L-arginine (L-NNA), and aminoguanidine dose-dependently inhibited the relaxation response by 50% to 70%. The free radical scavenger 2-(4 carboxyphenyl) 4,4,5,5-tetramethyl-imidazoline-1-oxyl-3-oxide (carboxy-PTIO) and the guanylyl cyclase inhibitor methylene blue also abrogated 70% and 45% of the relaxation response, respectively. ES caused an increase in muscle cGMP from 2.3+/-0.3 to 3.9+/-0.5 picomoles per muscle. L-NNA or L-NAME significantly decreased the tissue cGMP content (to 1.2+/-0.1 picomoles per muscle) and prevented the increase caused by ES. CONCLUSIONS: The relaxation component of the iris sphincter response to ES is a distinct nonadrenergic, noncholinergic, ES induced event. Most of the relaxation is mediated by the endogenously generated NO-guanylyl cyclase-cGMP cascade. PMID- 10711709 TI - Neovascularization grading methods in a rat model of retinopathy of prematurity. AB - PURPOSE: The method of counting cell nuclei above the internal limiting membrane in histologic sections is considered the standard when quantifying neovascularization (NV) in rodent oxygen-induced retinopathy (OIR). An alternative, more rapid method of counting clock hours in flatmounted adenosine diphosphatase (ADPase)-stained rat retinas is analogous to clinically scoring retinopathy of prematurity (ROP). In the present study, the validity of counting clock hours was evaluated by a direct comparison of these techniques. The intereye correlation of NV score and retinal vascular area were also studied. METHODS: Newborn Sprague-Dawley rats were exposed to cycles of O2 (80-10%) for 7 days, followed by 5 days of room air recovery. Preretinal NV was quantified by three masked observers counting clock hours in flatmounted ADPase-stained retinas of both eyes. Retinal vascular and total retinal areas were calculated using computer-assisted analysis. Representative retinas that had been scored positive (n = 10) and negative (n = 3) for NV and room air control retinas (n = 3) were embedded in paraffin. Each entire peripheral retinal quadrant was serially sectioned at 6 microm and stained with hematoxylin and eosin. Nuclei above the internal limiting membrane were then counted in a masked manner. The total number of nuclei counted per retina was defined as the nucleus count (704-938 sections per retina; 12,900 sections). Correlations were evaluated using Spearman rank coefficients. RESULTS: The nucleus count was 0 to 44 in room air control retinas, 0 to 40 in negative OIR retinas, and 250 to 5634 in positive OIR retinas. The nucleus count was highly correlated with the clock hour score (r(s) = 0.95, P = 0.0001). For the paired retinas, there was a significant correlation between right and left eyes in the severity of NV (clock hours; r(s) = 0.76, P = 0.0001) and the ratio of retinal vascular area to total retinal area (r(s) = 0.81, P = 0.0001). CONCLUSIONS: The more rapid method of counting clock hours in flatmounted ADPase-stained retinas is valid for quantifying NV in rat models of ROP. Incidence and severity of NV and vascularized areas were similar between left and right eyes, which permits the use of paired retinas for complementary research techniques. PMID- 10711710 TI - New ABCR mutations and clinical phenotype in Italian patients with Stargardt disease. AB - PURPOSE: To assess the mutation spectrum in the ABCR gene and clinical phenotypes in Italian families with autosomal recessive Stargardt disease (STGD1) and fundus flavimaculatus (FFM). METHODS: Eleven families from southern Italy, including 18 patients with diagnoses of STGD1, were clinically examined. Ophthalmologic examination included kinetic perimetry, electrophysiological studies, and fluorescein angiography. DNA samples of the affected individuals and their family members were analyzed for variants in all 50 exons of the ABCR gene by a combination of single-strand conformation polymorphism analysis and direct sequencing techniques. RESULTS: TenABCR variants were identified in 16 (73%) of 22 mutant alleles of patients with STGD1. Five mutations of 10 that were found had not been previously described. The majority of variants represent missense amino acid substitutions, and all mutant alleles cosegregate with the disease in the respective families. These ABCR variants were not detected in 170 unaffected control individuals (340 chromosomes) of Italian origin. Clinical evaluation of these families affected by STGD1 showed an unusually high frequency of early age related macular degeneration (AMD) in parents of patients with STGD1 (8/22; 36%), consistent with the hypothesis that some heterozygous ABCR mutations enhance susceptibility to AMD. CONCLUSIONS: Patients from southern Italy with Stargardt disease show extensive allelic heterogeneity of the ABCR gene, concordant with previous observations in patients with STGD1 from different ethnic groups. Half the mutations identified in this study had not been previously described in patients with STGD1. Screening of increasingly large numbers of patients would help to determine whether this can be explained by ethnic differences, or is an indicator of extensive allelic heterogeneity of ABCR in STGD1 and other eye diseases. In 6 (55%) of 11 families, the first-degree relatives of patients with STGD1 were diagnosed with early AMD, supporting the previous observation that some STGD1 alleles are also associated with AMD. PMID- 10711711 TI - TIMP-3, collagen, and elastin immunohistochemistry and histopathology of Sorsby's fundus dystrophy. AB - PURPOSE: Mutations in the tissue inhibitor of metalloproteinases-3 (TIMP-3) gene have previously been identified in patients with Sorsby's fundus dystrophy (SFD). We evaluated the ocular distribution of TIMP-3 and other extracellular constituents in SFD. METHODS: The eyes of an SFD donor with a confirmed TIMP-3 mutation were examined using histologic techniques demonstrating connective tissue, calcium, and lipid. Immunohistochemical analyses were performed using antibodies against TIMP-3, collagen type IV, V, and VI, laminin, fibronectin, elastin, and fibrillin. Electron microscopy also was used. RESULTS: A subretinal pigment epithelium (sub-RPE) deposit similar to that previously described was seen. A morphologically similar but different deposit was present internal to the nonpigmented ciliary epithelium (NPCE). Both deposits contained collagens, elastin, glycosaminoglycans, lipids, and calcium. Immunolabeling of TIMP-3 was found in the basement membrane of the NPCE, Bruch's membrane, and choroidal vessels in normal control subjects. In SFD, immunolabeling of TIMP-3 also was present in the sub-RPE deposit and in the inner portion of the ciliary body deposit. TIMP-3 immunoreactivity was more extensive in the SFD eye. The pattern of elastin immunoreactivity was remarkably similar to that of TIMP-3. Electron microscopy revealed a morphologically altered elastic layer of the Bruch's membrane. CONCLUSIONS: Sub-RPE TIMP-3 immunoreactivity appears more extensive in SFD than in control subjects. There is also a correspondence between TIMP-3 and elastin immunoreactivies, which invites speculation as to a link between the SFD TIMP-3 mutation and altered elastin processing. The accumulation of abnormal material in SFD is more widespread than previously reported. In view of this, SFD might be better termed Sorsby's ocular epitheliopathy. PMID- 10711712 TI - Dorsal retinal pigment epithelium differentiates as neural retina in the microphthalmia (mi/mi) mouse. AB - PURPOSE: Microphthalmia, a bHLH-zip transcription factor associated with the onset and maintenance of pigmentation, identifies the retinal pigment epithelial (RPE) compartment during optic vesicle and optic cup development. To determine a role for microphthalmia (mi) during eye development, the effects of an mi loss of function mutation on RPE and neural retinal were investigated in the mi/mi mouse. METHODS: A series of embryonic and postnatal mi/mi and wild-type eyes were sectioned and labeled with neural retina- and RPE cell type-specific antibodies. Photoreceptor loss was quantified by counting the number of photoreceptor nuclei spanning the outer nuclear layer throughout postnatal retinal development. RESULTS: Early neural retinal differentiation is not affected in the mi/mi mouse. The mi/mi ventral retinal pigment epithelial layer begins to develop normally, but does not pigment or attain a differentiated cuboidal morphology. The dorsal region of mi/mi retinal pigment epithelium expands and forms an ectopic retina, which develops all major retinal cell types along a similar time course as the wild type. After birth, mi/mi photoreceptors begin to form rosettes, outer segments fail to elongate, and over an extended time period, the retina degenerates. CONCLUSIONS: Together these results suggest that early retinal development can proceed normally in the mi/mi mutant, but later retinal histogenesis is dependent on the presence of a differentiated retinal pigment epithelium. Most importantly, loss of mi function permits a change in cell fate from RPE to retina in the dorsal eye. PMID- 10711713 TI - The retina of c-fos-/- mice: electrophysiologic, morphologic and biochemical aspects. AB - PURPOSE: Mice without a functional c-Fos protein (c-fos-/- mice) do not exhibit light-induced apoptotic cell death of rods in contrast to their wild-type littermates (c-fos+/+ mice). To analyze the consequences of the absence of c-fos in the retina, we investigated whether the retinas of c-fos-/- mice have a reduced capacity to absorb and transduce light compared with c-fos+/+ mice. METHODS: Retinal function was evaluated in dark-adapted mice by full-field electroretinograms (ERGs) over more than 6 log units of intensity. Retinal morphology was studied by light- and electron microscopy. Arrestin and the heat shock protein 70 (Hsp70) were detected by Western blot analysis. The rhodopsin content and the kinetics of rhodopsin regeneration were determined in retinal extracts. RESULTS: Although the configuration of the ERGs was comparable in both groups of mice, c-fos-/- mice showed a marked variability in all quantitative ERG measures with lower mean amplitudes, longer latencies, and a 0.9-log-unit lower b wave sensitivity on average. Morphometry showed that c-fos-/- mice have 23% fewer rods on average, whereas the number of cones was comparable among c-fos+/+ and c fos-/- mice. Arrestin levels appeared slightly reduced in c-fos-/- mice when compared with c-fos+/+ mice, whereas Hsp70 levels were comparable in both genotypes. The kinetics of rhodopsin regeneration were similar, but c-fos-/- mice had a 25% lower rhodopsin content on average. CONCLUSIONS: Compared with c-fos+/+ mice, retinal function in c-fos-/- mice is attenuated to a variable but marked degree, which may be, at least in part, related to the reduced number of rods and the reduced rhodopsin content. However, c-fos does not appear to be essential for the ability to absorb photons, nor for phototransduction or the function of second-order neurons. The resistance to light-induced apoptosis of photoreceptor cells in c-fos-/- mice may result from the acute deficit of c-fos in the apoptotic cascade rather than from developmental deficits affecting rod photoreceptor function. PMID- 10711714 TI - Impairment of rod cGMP-gated channel alpha-subunit expression leads to photoreceptor and bipolar cell degeneration. AB - PURPOSE: To determine whether alterations in rod cGMP-gated channel function mediate retinal degeneration, a transgenic approach in which the alpha subunit of the rod cGMP-gated channel is reduced by the expression of an antisense RNA was used. METHODS: A 890-bp fragment of the 5' mouse rod cGMP-gated channel cDNA was cloned in the antisense orientation under the control of the strong bacterial cytomegalovirus promoter. This antisense construct was used to generate transgenic mice in which the expression of the antisense transgene was measured by reverse transcription-polymerase chain reaction. Histologic, immunocytochemical, and TdT-dUTP terminal nick-end labeling (TUNEL) analyses were performed on control and transgenic retina sections to determine the effects of antisense expression on specific cell types. RESULTS: The antisense RNA was able to inhibit the translation of the rod channel protein in an in vitro assay. Three transgenic mouse lines expressing the antisense construct were obtained. Molecular analyses showed that the antisense is expressed in the eye of each transgenic mouse line, where it reduces rod cGMP-gated channel RNA expression. Histologic and immunocytochemical data showed that transgenic retinas have a reduced number of photoreceptors and bipolar cells. TUNEL staining confirmed that photoreceptor and bipolar cells degenerate along an apoptotic pathway. CONCLUSIONS: Impairment of rod cGMP-gated channel alpha subunit expression leads to photoreceptor and bipolar cell degeneration. These transgenic mice are the first model of retinal degeneration caused by an alteration in the expression of the rod cGMP-gated channel. This model system can be used to test therapies designed to slow or stalled retinal degenerations. PMID- 10711715 TI - Neurotrophic factors cause activation of intracellular signaling pathways in Muller cells and other cells of the inner retina, but not photoreceptors. AB - PURPOSE: Intravitreal injection of brain-derived neurotrophic factor (BDNF), ciliary neurotrophic factor (CNTF), or basic fibroblast growth factor (FGF2) promotes survival of photoreceptors exposed to various types of insults, but it is not known if these survival-promoting effects occur by direct action of the factors on photoreceptors or indirectly through the activation of other cells. In this study, the authors have sought to address this issue by determining which cells in the retina show evidence of activated intracellular signaling pathways acutely and at longer time points after intravitreal injection of these agents. METHODS: Retinas were removed from C57BL/6J mice at 1, 6, or 24 hours after intravitreal injection of 1 microg of human BDNF, rat CNTF, human FGF2, or human transforming growth factor-alpha (TGFalpha), and immunohistochemically stained for phosphorylated extracellular signal-regulated kinase (pERK), phosphorylated cAMP responsive element binding protein (pCREB), or c-fos. Retinal organ cultures were incubated with 10 ng/ml of BDNF, CNTF, FGF2, or TGFalpha for 10 or 30 minutes or 1, 3, or 6 hours and then immunohistochemically stained for pERK, pCREB, or c-fos. RESULTS: Intravitreal injection of BDNF, CNTF, or FGF2 resulted in a rapid increase in pERK immunoreactivity in Muller cells and a rapid increase in c-fos immunoreactivity in Muller, amacrine, and ganglion cells. Immunoreactivity for pERK and c-fos returned to baseline in all retinal cells at 6 or 24 hours after injection, but there was increased staining for glial fibrillary acidic protein (GFAP) in Muller cells at these time points. At no time after injection was there any staining for pERK or c-fos in photoreceptors. Similarly, retinal explants treated with FGF2, BDNF, or CNTF showed increased staining for pCREB, pERK, and c-fos in cells of the inner retina, but not photoreceptors. CONCLUSIONS: These data support the hypothesis that BDNF, CNTF, and FGF2 exert their effects on photoreceptors by acting indirectly through activation of Muller cells and perhaps other nonphotoreceptor cells. PMID- 10711716 TI - Retinoic acid produces rod photoreceptor selective apoptosis in developing mammalian retina. AB - PURPOSE: All-trans retinoic acid (ATRA) or 9-cis retinoic acid (9CRA), added to dissociated developing neural retinal cells, induces progenitor cells to adopt the rod cell's fate. Retinoic acid (RA) also produces apoptotic cell death in developing tissues. The effects of retinoids on mouse retinal development were examined. METHODS: Retinas were explanted on postnatal day (PN)1 and cultured with or without the retinal pigment epithelium (RPE) attached. Retinas were cultured for 3 weeks in the absence or presence of 100 or 500 nM ATRA or 9CRA. Morphologic development and apoptotic cell death were examined using cell specific immunocytochemical markers, the TdT-dUTP terminal nick-end labeling (TUNEL) method, and a caspase assay. RESULTS: Retinal explants, with and without RPE, had similar age-dependent increases in opsin expression. In contrast, explants with RPE had less apoptosis during the first week than retinas without RPE. In explants with RPE, ATRA or 9CRA produced rod-selective apoptotic cell death in which 20% to 25% were lost by PN7 with no further loss by PN21. 9CRA treated explants without RPE had a decreased number of apoptotic cells and a higher number of (rhod)opsin-positive cells at PN3. CONCLUSIONS: Factors in RPE appear to regulate rod apoptosis in developing retina. Retinoids produce rod selective apoptotic cell death during normal rod differentiation. In contrast, retinoids accelerate the expression of opsin in retinas without RPE. These differential effects of RA on rod photoreceptors-apoptosis and differentiation are similar to those observed in other developing tissues and play an important role in both normal and pathologic development. PMID- 10711718 TI - Effects of superior colliculus inhibition on visual motion processing in the lateral suprasylvian visual area of the cat. AB - PURPOSE: To clarify whether visual inputs of the tectothalamocortical pathway influence motion processing within the lateral suprasylvian (LS) area of the cat. METHODS: This study was conducted in five cats. Tungsten microelectrodes were used for recording visual evoked potentials. The electrodes were introduced into the LS area. An array of 120 randomly located dots was projected onto the stimulus field (40 degrees x 40 degrees) in front of the animal by a slide projector. The dots were moved rightward and leftward alternatively with interstimulus intervals by a mirror attached to a galvanometer, the movements of which were controlled by a microcomputer. Each motion sequence consisted of an abrupt onset of motion that continued for 100 msec followed by an abrupt offset and a stationary phase of 900 msec; the total duration of each sequence was thus 1000 msec. The velocity of the motion was varied in 12 steps. The onset of motion was used as the trigger for recording evoked potentials. Single or multiple injections (two to three) of muscimol were made, mainly into the rostral superior colliculus (SC). The amplitudes of evoked potentials before and after the muscimol injection were compared. RESULTS: A large negative wave (N1) with the peak latency of 89.80+/-16.39 msec (mean +/- SD, n = 191) was recorded consistently. The amplitude of N1 was not altered by the muscimol injection into the SC when the velocity of motion was 50 deg/sec or less. When the velocity of motion was 75 deg/sec or more, however, the amplitude of N1 was reduced to 62% to 72% of that noted before the muscimol injection. CONCLUSIONS: These findings suggest that the LS area processes the visual motion inputs reaching through the two parallel pathways, the geniculostriate pathway and the tectothalamocortical pathway, when the velocity of visual motion is 75 deg/sec or more. PMID- 10711717 TI - Effect of onset age of strabismus on the binocular responses of neurons in the monkey visual cortex. AB - PURPOSE: By 6 weeks of age, neurons in the monkey's primary visual cortex acquire qualitatively adult-like binocular response properties and behaviorally stereopsis emerges. In this study, it was determined whether the onset of strabismus has a more severe impact on cortical binocularity before or after this critical developmental age. METHODS: Infant monkeys were fit with a light-weight helmet which held a total of 27 diopters of base-in prisms in front of their two eyes for a fixed period of two weeks. For one group of infant monkeys, prism rearing began at 2 weeks of age and for a second group, the onset was at 6 weeks of age. Immediately after the rearing period, i.e., at 4 weeks and 8 weeks of age, respectively, extracellular single-unit recording methods were used to determine the nature and severity of alterations in the binocular response properties of V1 neurons. Dichoptic sinewave gratings were used as visual stimuli. RESULTS: In comparison to normal age-matched infants, V1 neurons in both strabismic groups exhibited reductions in sensitivity to interocular spatial phase disparities (disparity sensitivity) and a higher prevalence of binocular inhibitory interactions (binocular suppression). However, the reduction in disparity sensitivity and the magnitude of binocular suppression were much greater in the late (6-8 weeks) than the early (2- 4 weeks) onset group. CONCLUSIONS: Discordant binocular signals due to brief periods of early strabismus have more serious effects on the development of binocular properties of V1 neurons if they occur shortly after rather than before the emergence of stereopsis (i.e., when the binocular connections are relatively more mature but the visual cortex still shows a high degree of plasticity). PMID- 10711719 TI - The regulatory effects of thyroid hormone on the activity of 3-hydroxy-3 methylglutaryl coenzyme A reductase. AB - The effects of 3, 3', 5-triiodothyronine (T3) on 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase activity were evaluated in the C100 baby hamster kidney cell line. Cells cultured in Minimal Essential Medium (MEM) were supplemented with 10% thyroid hormone-depleted fetal bovine serum (THDS-MEM) and had a 70.1% lower level of HMG-CoA reductase activity than the cells grown in a medium supplemented with fetal bovine serum (FBS). When T3 was added to THDS-MEM, the reduction of the reductase activity was blocked in a dose-dependent manner. In the cells grown in THDS-MEM for 48 hours, T3 (10(-6) M) treatment rapidly increased HMG-CoA reductase activity, achieving the control level six hours after treatment. Such effects of T3 were blocked by actinomycin D (5 microg/ml) or cycloheximide (10 microg/ml). The transcriptional rate of the HMG-CoA reductase gene did not change significantly regardless of the presence of T3, while T3 inhibited the 25-hydroxycholesterol-mediated decay of the reductase mRNA significantly. Our results show that T3-dependent regulation of HMG-CoA reductase activity, via the de novo synthesis of the reductase enzyme, seems to be mediated at least partially by the stabilization of HMG-CoA reductase mRNA. PMID- 10711720 TI - The pyruvate dehydrogenase E1 alpha gene is testosterone and prolactin regulated in prostate epithelial cells. AB - The prostate gland of humans and other animals has the unique function of accumulating and secreting extraordinarily high levels of citrate. The prostate secretory epithelial cells synthesize citrate which, due to a limiting mitochondrial (m-) aconitase, accumulates rather than being oxidized. Thus citrate is essentially an end product of metabolism in prostate. For continued net citrate production, a continual source of oxaloacetate (OAA) and acetyl CoA is required. Glucose via pyruvate oxidation provides the source of Acetyl CoA. In prostate cells, citrate production is regulated by testosterone and/or by prolactin. Both hormones selectively regulate the level and activity of pyruvate dehydrogenase E1 alpha (E1a) in animal prostate cells; thereby regulating the availability of acetyl CoA for citrate synthesis. Studies were conducted to determine if testosterone and prolactin might regulate the expression of the E1a gene in prostate epithelial cells. Prolactin treatment of rat ventral and lateral prostate cells and human PC3 cells increased the levels of E1a mRNA and the rates of transcription of the E1a gene. Testosterone also increased the mRNA level and transcription of E1a in rat ventral prostate cells, and in PC3 cells transfected with androgen receptor. However, testosterone treatment resulted in a repression of E1a gene expression in lateral prostate cells. Evidence is presented which supports the view that prolactin regulation of E1a is mediated via PKC. The rapidity of the effects of both hormones is representative of an immediate-early gene response. To our knowledge this represents the first report in any mammalian cells that, in addition to its constitutive expression in all mammalian cells, the E1a gene is a hormonally-regulated gene in specifically targeted prostate epithelial cells. PMID- 10711721 TI - Seasonal pattern of leutinizing, follicle-stimulating hormone, testosterone and progesterone in adult population of both sexes in the Jordan Valley. AB - Differences were observed in hormonal levels between in both sexes of people living in Irbid City (620 meters above sea level) and in the Jordan Valley (360 meters below sea level). In addition, exercise was shown to differentially affect hormonal levels of both sexes at the above and below sea level locations. Serum levels of leutinizing hormone (LH) and testosterone (T) in adult male and serum levels of follicle-stimulating hormone (FSH) and progesterone (P) in adult female people were investigated in Irbid City and in the Jordan Valley during the years 1997 and 1998. The levels of these hormones were followed each month during this period. In males living in Irbid City, LH and T peaked from March through June, and in females at the same site, FSH and P also peaked from March through June. These data confirm the seasonal variation in sex hormones reported elsewhere in (wo)man and in other species. In males and females of the Jordan Valley, serum levels of LH, FSH, T and P were all higher than those of Irbid City throughout the year. Additionally, peaks of LH and T in male and FSH and P in female subjects in the Jordan Valley were observed from March through September. The high levels of these hormones and the extension of their peaks are suggested to be due to effects of the environmental factors of the Jordan Valley (high temperature, high barometric pressure) compared to those in Irbid City and other areas located at above sea level altitude. PMID- 10711722 TI - Water deprivation reveals early neuromyopathy in diabetic mice. AB - The effects of water deprivation on peripheral nerve and muscle function were investigated in flexor digitorum superficialis muscle of control and diabetic mice. Twenty mice (30 g average body weight) were injected once with streptozotocin solution (200 mg/kg) to induce experimental diabetes and another 20 mice of similar body weight served as controls. Two weeks later, comparative analyses of in situ muscle isometric contractile characteristics were performed by direct muscle stimulation and indirect nerve stimulation (at 1, 5 and 30 Hz) in urethane-anesthetized (2 mg/g, i.p.) control and diabetic mice. One day prior to the experiments, 10 control and 10 diabetic mice were deprived of water. The study contained four groups: hydrated (H) control, dehydrated (DH) control, H diabetic and DH diabetic. There were no significant differences in synaptic delay or twitch tension between H control and DH control. Comparing H control and H diabetic groups, no differences were noticed in synaptic delay or twitch tension; except at 30 Hz where twitch tension was reduced in H diabetic mice. Significant differences were observed when comparing DH control and DH diabetic mice. DH diabetic showed a significant increase in synaptic delay (from 7.4 to 9.3 ms) and a significant decrease in twitch tension evoked either by indirect nerve or by direct muscle stimulation (from 4.4 g to 1.9 g and from 4.4 g to 2.3 g, respectively). These results revealed that water deprivation enhances diabetes effects at the neuromuscular junction and at the muscle leading to further complication of neuromyopathy. PMID- 10711723 TI - Inhibition of apoptosis in cultured immature rat Leydig cells by human chorionic gonadotropin associated with Bcl-2 mRNA expression. AB - Bcl-2 is the first identified negative regulator of apoptotic cell death. When the level of Bcl-2 mRNA in rat whole testes was examined in the present study, it gradually decreased in rats from 2.5 to 9 weeks old. We also examined the effect of human chorionic gonadotropin (hCG) on apoptosis and the level of Bcl-2 mRNA expression in immature Leydig cells in vitro. When the cells were cultured with serum free media (SFM), Bcl-2 mRNA levels gradually decreased. On the other hand, the level of Bcl-2 mRNA in cells treated with 50 ng/ml of hCG decreased at 6 h, but increased after 12 h. At 24 h, the level of Bcl-2 mRNA in the treated cells was almost the same as that of control cells (time = 0). At 12 h after the addition of various concentrations (from 0.1-1000 ng/ml) of hCG, Bcl-2 mRNA levels increased in a dose-dependent manner. An analysis of DNA fragmentation showed that treatment with hCG prevents the apoptosis of immature Leydig cells. Our findings suggest that Bcl-2 mRNA may be related to the programmed cell death of immature rat Leydig cells in vitro, which are inhibited by hCG. PMID- 10711724 TI - Molecular forms of alpha-melanocyte-stimulating hormone in pheochromocytoma tissue. AB - The molecular forms of alpha-melanocyte-stimulating hormone (MSH) in pheochromocytoma tissues have been characterized using reversed-phase HPLC and radioimmunoassay. Six alpha-MSH-related peptides were detected. Three of the six peaks had elution times identical to those of synthetic desacetyl-alpha-MSH, alpha-MSH and diacetyl-alpha-MSH. The remaining three forms of the alpha-MSH-like immunoreactivity suggest a different processing of pro-opiomelanocortin in this tissue than in the intermediate lobe of the pituitary gland. Current results confirm that alpha-MSH are present in human pheochromocytoma and suggest that alpha-MSH has role in the pathophysiology of this tissue. PMID- 10711725 TI - Regulation of 11beta-hydroxysteroid dehydrogenase enzymes by dietary sodium in the rat. AB - 11Beta-hydroxysterold dehydrogenase enzymes (11beta-HSD1, 11beta-HSD2) regulate access of adrenocorticosteroids to receptors. 11Beta-HSD2 is a dehydrogenase that protects mineralocorticoid receptors from circulating glucocorticoid hormones, 11beta-HSD1 is a reductase that promotes formation of active hormone in glucocorticoid-sensitive tissues. Here we investigate whether low or high sodium diets affect 11beta-HSD enzyme activities and mRNA expression in liver and kidney tissues. 11Beta-HSD activity was measured as dehydrogenation of 3H-corticosterone by microsomes in the presence of NAD or NADP. In situ hybridisation techniques were used to assess expression of 11beta-HSD1 mRNA (liver and kidney) and 11beta HSD2 mRNA (kidney). Dietary sodium did not affect 11beta-HSD2 mRNA expression in collecting tubules of the medulla: 11beta-HSD1 mRNA in proximal tubules of the inner cortex/outer medulla was lower after a high sodium diet. 11Beta-HSD1 mRNA in liver was unaffected by treatment. Renal enzyme activity with NAD (11beta-HSD2 cofactor) was lower following a high sodium diet (P < 0.05). In the presence of NADP (11beta-HSD1 co-factor), neither renal nor hepatic activities were affected. Dietary sodium restriction appears to increase 11beta-HSD activity by a non genomic mechanism; this should enhance aldosterone specificity for mineralocorticoid receptors. 11Beta-HSD1 mRNA expression varies independent of enzyme activity and is not clearly related to altered glucocorticoid activity. PMID- 10711726 TI - Effect of bilateral ovariectomy and ovarian steroid hormones on the antioxidant systems and plasma malondialdehyde levels in Wistar rats. AB - We studied in the Wistar rat the effect of bilateral ovariectomy and of the administration of estradiol (E2) alone or together with medroxyprogesterone (MPA) on the lipid peroxidation (MDA) and the enzymatic (SOD and catalase) and non enzymatic (vitamins A and E) antioxidant systems. At 15 days after castration, we observed no variations in the lipid peroxidation levels (MDA), with there being a slight increase in vitamins A and E (p<0.05), catalase (p<0.01), and SOD (n.s.) activity. The addition of estradiol led to a decline (n.s.) in the MDA values and in the catalase and SOD activity, with no modification of the vit. A and E concentrations. The combined (E2+MPA) therapy did not modify the results obtained with E2 alone, although the catalase decline did now reach significance (p<0.05). PMID- 10711727 TI - Effects of recombinant murine leptin[1-147] and leptin fragment 116-130 on steroid secretion and proliferative activity of the regenerating rat adrenal cortex. AB - Leptin, the product of the ob gene, is a hormone mainly secreted by the adipose tissue, which acts through specific receptors (Ob-R) widely distributed in the body tissues. Ob-Rs are present in the mammalian hypothalamo-pituitary-adrenal axis, and evidence indicates that leptin regulates adrenocortical secretion. Moreover, leptin is known to act as a growth promoting factor in some tissues, including the endocrine ovary. We have investigated the effects of three subcutaneous injections of 2 nmol/100 g of native murine leptin[1-147] and of its biologically active fragment 116-130 on the secretory and proliferative activity of the regenerating rat adrenal cortex. Leptin[1-147] increased plasma aldosterone concentration at day 8 and plasma corticosterone concentration (PBC) at day 5 of regeneration, without affecting mitotic index. In contrast, leptin[116-130] lowered PBC and mitotic index at both times of adrenal regeneration. In light of the fact that adrenal regeneration is at least in part dependent on the pituitary ACTH, we conclude that: (i) native leptin moderately stimulates steroid secretion, acting directly on the adrenal cortex, through signaling mechanisms other than those involved in the ACTH action; (ii) native leptin is unable to enhance the proliferative activity of regenerating adrenals, which conceivably is maximally stimulated by ACTH; (iii)leptin[1-147] and leptin[116-130] differently interact with Ob-Rs or interact with different receptors; and (iv) leptin[116-130] inhibits the signaling pathways mediating both the secretagogue effect of native leptin and the proliferogenic effect of ACTH. PMID- 10711728 TI - The effect of prostaglandin E1 on human bone metabolism: evaluation by biochemical markers for bone turnover. AB - Prostaglandins (PGs) have complex and multiple effects on bone metabolism. Although the osteogenic effect of PGE in humans was initially found in an infant with a congenital heart disease, there have been few reports on the effect of PGE in human in vivo. The aim of this study was to investigate the effect of PGE1 on human bone metabolism, using biochemical bone markers. A total of 18 subjects were treated with PGE1 in lipid microspheres. Six subjects were given 10 microg of lipo-PGE, intravenously daily for 14 days, and twelve subjects were given the same dose twice a week for 7 weeks. Before and after the administration of PGE1, blood and a spot urine was obtained in the morning. Bone formation markers (alkaline phosphatase, osteocalcin, procollagen I carboxy-terminal peptide) did not change. In the subjects with daily administration for 2 weeks, type I collagen pyridinolines crosslinked C-telopeptide (ICTP) increased significantly. In the subjects treated twice a week, free deoxypyridinoline (Dpd) increased significantly. When all subjects were analyzed, bone resorption markers (ICTP and Dpd) increased, but not significantly (p=0.055 for ICTP, p=0.055 for Dpd). Therefore, PGE1 at the dosage used in this study did not increase bone formation but increased bone resorption in humans. PMID- 10711729 TI - Inter-relationships between endothelin and prostaglandin F2alpha in corpus luteum function. AB - In cattle and other species, the corpus luteum plays a central role in the regulation of cyclicity and maintenance of pregnancy. In the absence of fertilization and implantation, the corpus luteum undergoes functional and morphological regression or luteolysis. Luteal regression is initiated in domestic ruminants by surges of prostaglandin F2alpha (PGF2alpha) from the uterus. Despite intensive investigation, the mechanisms by which PGF2alpha causes luteal regression remain undetermined. Recent studies from several laboratories have demonstrated that endothelial cells and their product, endothelin 1, are required for the manifestation of the luteolytic effects of PGF2alpha. Experimental evidence strongly supports the concept that luteal endothelin 1 inhibits luteal steroidogenesis and mediates the effects of PGF2alpha. Endothelin 1 caused a dose-dependent reduction in both basal and luteinizing hormone stimulated biosynthesis of progesterone and prostacyclin, and an increase in PGF2alpha by ovine and bovine luteal cells. Specific receptors for endothelin 1 were identified on large and small bovine luteal cells, and the addition of specific endothelin receptor antagonists abolished the inhibitory effects of endothelin 1. Luteal endothelin 1 content increased as the cyclic corpus luteum aged, and the highest concentrations were observed during luteolysis. The amount of mRNA encoding endothelin 1 was greatly increased during the period of luteolysis. Gene expression for endothelin 1 was increased, in a time-dependent manner, in corpora lutea collected from heifers and ewes after exogenous administration of PGF2alpha. In heifers, exogenous PGF2alpha resulted in increased luteal output of endothelin 1. In ewes, the luteolytic effects of PGF2alpha were mitigated by pretreatment with a specific endothelin receptor antagonist. Administration of endothelin 1 or a sub-luteolytic dose of PGF2alpha to ewes reduced concentrations of jugular venous progesterone but did not shorten luteal lifespan. However, a combination of endothelin 1 and PGF2alpha acted synergistically to bring about complete luteolysis and reduced lifespan of the corpus luteum. In summary, endothelin 1 appears to have a direct effect on luteal cells in cattle and sheep, and it plays an essential role in mediating the luteolytic effects of PGF2alpha. PMID- 10711730 TI - Maternal recognition of pregnancy in marsupials. AB - Pregnancy in kangaroos and wallabies (macropodid marsupials) induces multiple unilateral responses in the reproductive system that override those related to proximity to the single corpus luteum on one ovary or to the follicle on the contralateral ovary. This situation is in contrast to most other non-macropodid marsupials, in which the responses are dependent on the corpus luteum. There is now good evidence that these unilateral responses in macropodids are controlled by the feto-placental unit acting locally to stimulate the endometrium and myometrium. Pregnancy also influences the duration of the oestrous cycle and maternal behaviour. The stimuli responsible for these effects probably include paracrine, endocrine and mechanical stimuli resulting from uterine stretch. Taken together, these unilateral responses demonstrate that there is a refined maternal recognition of pregnancy in at least the macropodid marsupials. PMID- 10711731 TI - The human corpus luteum: remodelling during luteolysis and maternal recognition of pregnancy. AB - The marked tissue remodelling associated with luteolysis involves increased expression and activity of matrix metalloproteinases (MMPs) and an influx of immune cells, notably macrophages. Since the corpus luteum expresses high concentrations of specific tissue inhibitors of MMPs, it is clear that it is not only the increased activity of MMPs that is important, but also their tissue localization. Human chorionic gonadotrophin inhibits both MMP expression and macrophage influx in the rescued corpus luteum of early pregnancy. However, macrophages and the main cellular sources of MMPs in the corpus luteum do not express LH-hCG receptors. Therefore, it is likely that products of the steroidogenic cells, which do express LH-hCG receptors, are involved in the differential paracrine regulation of MMP expression and macrophage influx during luteolysis and maternal recognition of pregnancy. PMID- 10711732 TI - Placental leptin. AB - Placental tissues from humans, rodents and farm animals contain leptin and its receptor. Leptin produced by the human placenta has the same size, charge and immunoreactivity as leptin produced by adipose tissue. However, the expression of human placental leptin appears to be regulated by a placenta-specific upstream enhancer. In this review the occurrence of leptin and its receptor in a range of species and placental types is described, and its significance during pregnancy discussed. Placental leptin contributes to the increase in maternal circulating concentrations of leptin during late pregnancy when it is likely to have an endocrine role in regulating maternal energy balance. Placental leptin may have angiogenic and immunomodulatory activities, which affect the placenta in an autocrine or paracrine manner. It also appears to affect fetal growth and development by binding to leptin receptors present in fetal organs. PMID- 10711733 TI - Genome mapping in ruminants and map locations for genes influencing reproduction. AB - Genetic maps provide a critical link between genes and phenotypes and are essential tools in the search for the genetic basis of variation in reproductive traits. Genes coding for hormones, growth factors, receptors, binding proteins, transcription factors and enzymes that influence the development and function of the reproductive axis have been assigned to genetic maps of ruminants and locations can be found in the respective genome databases. In addition, comparative information on gene structure and map location will help define the functions of essential genes. Gene locations from other species can be used because of extensive comparative links among mammalian gene maps. Large-scale projects to sequence genes and the ability to map these genes in parallel in radiation hybrid panels of different species will greatly improve the maps and our ability to translate between them. Cloning the genes responsible for genetic differences in fertility and fecundity in ruminants is likely to provide valuable clues to understanding ovarian function and germ cell development. PMID- 10711734 TI - Effects of dietary fatty acids on reproduction in ruminants. AB - Fats in the diet can influence reproduction positively by altering both ovarian follicle and corpus luteum function via improved energy status and by increasing precursors for the synthesis of reproductive hormones such as steroids and prostaglandins. Dietary fatty acids of the n-3 family reduce ovarian and endometrial synthesis of prostaglandin F2alpha, decrease ovulation rate in rats and delay parturition in sheep and humans. Polyunsaturated fatty acids such as linoleic, linolenic, eicosapentaenoic and docosahexaenoic acids may inhibit prostaglandin F2alpha synthesis through mechanisms such as decreased availability of its precursor arachidonic acid, an increased competition by these fatty acids with arachidonic acid for binding to prostaglandin H synthase, and inhibition of prostaglandin H synthase synthesis and activity. It is not known whether polyunsaturated fatty acids regulate expression of candidate genes such as phospholipase A2 and prostaglandin H synthase via activation of nuclear transcription factors such as peroxisome proliferator-activated receptors. Manipulation of the fatty acid profile of the diet can be used potentially to amplify suppression of uterine synthesis of prostaglandin F2alpha during early pregnancy in cattle, which may contribute to a reduction in embryonic mortality. Feeding fats and targeting of fatty acids to reproductive tissues may be a potential strategy to integrate nutrition and reproductive management to improve animal productivity. PMID- 10711735 TI - Cryopreservation of semen from domestic livestock. AB - Fifty years after the first successful cryopreservation of spermatozoa, the technique is an integral part of the cattle breeding industry but has failed to establish itself commercially in the production of other breeds of domestic livestock. New assessment techniques have shown that the ejaculate consists of a heterogeneous population of cells, which achieve their full fertility potential at different rates within the female tract and thus maximize the chances of a fertile spermatozoon successfully combining with an egg. It is becoming apparent that the freeze-thaw process results in a more homogeneous cell population, which may be functionally compromised. One aspect of sperm function that has been demonstrated to be affected by cryopreservation is the process of capacitation. Chlortetracycline staining has shown that frozen-thawed spermatozoa undergo an accelerated 'capacitation-like' process which has implications for their interaction with the female tract, ability to establish sperm reservoirs in vivo and hence for their life expectancy after insemination. In addition to heterogeneity within the ejaculate, there is increasing evidence for variation between individuals in the success of sperm freezing. Post-thaw sperm survival may be consistently poor for certain individual animals even though pre-freeze parameters appear normal. The mechanisms that may underlie such differences in cryosensitivity remain unclear. A greater role for the use of frozen semen in livestock production can come only from an improvement in the preservation of the functional competence of the cryopreserved spermatozoon after insemination into the female tract. PMID- 10711736 TI - Cadherins: crucial regulators of structure and function in reproductive tissues. AB - Cadherins are cell surface proteins that are directly involved in a wide variety of processes such as cell adhesion, cell sorting, cell survival, morphogenesis, formation of intercellular junctions, maintenance of tissue integrity and tumourigenesis. This review discusses the multiple functions of cadherins in reproductive tissues. Furthermore, the role of the intracellular signalling protein beta-catenin in regulating cadherin function is reviewed. Finally, the findings that cadherin concentrations in reproductive tissues are responsive to steroid hormones is discussed. The modulation of cadherin expression by hormones is in agreement with the hypothesis that these proteins are dynamically involved in the maintenance of structure and function in reproductive tissues. PMID- 10711737 TI - Arterial hypertension and hemorheology. What is the relationship? AB - The strong relationship between arterial hypertension and hemorheological alteration present in all the vascular segments can be affirmed. These hemorheological changes are: an increase in the whole blood and plasma viscosity, in red blood cell (RBC) rigidity and aggregability and subsequently a decrease in oxygen delivery. Anyway, there are very interesting, still now unclear questions to be resolved: 1. Can arterial hypertension (with its vascular and cardiac remodeling) be the most relevant factor inducing alterations in the macro and the microrheology? 2. Can the altered hemorheology, above all in the microcircle, be one of the numerous causes of hypertension? 3. What is the influence of tissue oxygenation changes in these situations: are these changes or effects of alterations during hypertension? In the next years using new technologies we hope that these problems will be resolved indicating new ways to treat hypertensives and to present its related complications. PMID- 10711738 TI - Microvascular oxidative stress, immune reaction and apoptosis in hypertensives. AB - The mechanisms that lead to organ injury in hypertension are incompletely understood. In particular, there is a lack of evidence that serves to link the elevation of arterial blood pressure with end organ damage. Experimental models of hypertension have a range of microvascular abnormalities in addition to a shift in blood pressure. There is evidence for an oxidative stress in microvascular endothelium derived from xanthine and NADPH oxidase. Furthermore, there exists an immune suppression accompanied by abnormally elevated circulating leukocyte counts, depression of selectin membrane adhesion to the endothelium and enhanced cell apoptosis. Many of the deficiencies in the spontaneously hypertensive rats can be corrected by adrenalectomy, suggesting a contribution of glucocorticoids to the abnormalities in this model. These observations suggest a significantly enhanced vascular oxidative stress which is accompanied by a frustrated inflammatory response due to a glucocorticoid dependent deficiency of leukocyte adhesion to vascular endothelium. PMID- 10711739 TI - Red blood cell (RBC) deformability, RBC aggregability and tissue oxygenation in hypertension. AB - Arterial hypertension could be considered a progressive ischaemic syndrome interesting the macro and the microcirculation. In order to improve the clinical and therapeutic approach to the treatment of arterial hypertension, research has centered on blood flow to evaluate the different components and their very intricate relationships influencing the micro- and the macrocirculation. Of course the main problem is to study the link between the blood flow and the peripheral tissue oxygenation. During hypertension very important alterations in rheological, mechanical and biochemical characteristics of erythrocytes and of blood flow have been shown. It is very relevant the increase in blood viscosity, the decrease in red blood cell (RBC) deformability, the formation of RBC "rouleaux" and RBC aggregates. These hemorheological determinants can favour an increase of peripheral resistances and of arterial blood pressure, causing or worsening hypertension, a decrease in oxygen transport to tissue and peripheral perfusion, a decrease of the active exchange surface area in the microvasculature, especially in complicated hypertension. We have studied 320 patients: 123 with Essential Hypertension (EH) (M 59, F 64 aged 50 +/- 25 years); 81 with Secondary Hypertension (SH) without associated other pathologies influencing hemorheology (M 42, F 39 aged 48 +/- 20 years); 116 SH with other pathologies or conditions associated influencing hemorheology such as: diabetes, lipoidoproteinosis, obesity, smoking, HD, elderly, etc. (M 48, F 68 aged 46 +/- 20 years). Using a Laser-assisted Optical Rotational Red Cell Analyzer (LORCA) acc. to Hardeman (1994) we studied Elongation Index (EI) and aggregation kinetics of red blood cells in these patients. We also evaluated TcpO2 and TcpCO2 using a transcutaneous oxymeter (Microgas 7650, Kontron Instruments). In hypertensives we found a decrease in erythrocyte deformability (evaluated with EI), in erythrocyte aggregation time, a fibrinogenaemia increase, an increase of shear rate to disaggregate erythrocytes, a decrease in cellular oxygen delivery and tissue oxygenation, an impairment of microcirculation. These changes may be involved in the development of arterial hypertension and in its pathogenesis. These patterns also are more impaired in hypertensives with diabetes, lipoidoproteinosis, etc. These patterns are not related with the age of the patients but they are significantly and directly related (p < 0.01) with the patient hypertension-age. This could be a new way to realize a better treatment in hypertensives and a prevention of cardiovascular complications (i.e.: myocardial infarction, TIA, etc.). PMID- 10711740 TI - Red cell fluidity in hypertension. AB - Rheological findings in essential hypertension are increases in haematocrit, plasma fibrinogen, plasma and whole blood viscosity, and erythrocyte aggregability as well as impaired erythrocyte deformability. Are these abnormalities secondary effects of an increased blood pressure via an increased filtration pressure rendering haemoconcentration or is the initial pressure increase the result of a deterioration of any of the rheologic variables? Since the diameter of the red cell is about 8.5 microm, and that of the smallest capillaries about 3 microm, the ability of the cell to deform is of vital importance for capillary flow, and a decreased erythrocyte deformability could cause an increased microvascular flow resistance. We found a negative correlation between erythrocyte deformability and fasting insulin and also a decreased erythrocyte deformability in hypertensive patients during a 2 h euglycaemic insulin clamp. Associations between increased intracellular Ca2+ and decreased erythrocyte deformability on one hand and between in vitro insulin and an accumulation of Ca2+ in red blood cells on the other have earlier been shown. Hence, a decreased insulin sensitivity might be one important factor in the development of hypertension acting via an impaired erythrocyte deformability and an increased flow resistance in the microcirculation. PMID- 10711741 TI - Haemorheological disturbances in hypertension: the influence of diabetes and smoking. AB - Patients suffering from diabetes or hypertension commonly exhibit increased blood viscosity compared with healthy controls. This is primarily the result of elevated plasma fibrinogen concentration. Cigarette smokers also exhibit raised blood viscosity but in their case the main cause is a raised haematocrit. In this paper the effects of concurrent hypertension, diabetes and cigarette smoking on blood viscosity is reviewed. Evidence is presented that the haemorheological disturbances associated with each of these modalities are additive when present together in a subject. PMID- 10711742 TI - Red blood cell intercellular interactions in oxidative stress states. AB - Red blood cell (RBC) intercellular interactions, i.e., self-aggregation and adherence to endothelial cells (EC), play important roles in microcirculation. These RBC flow properties are determined by cell membrane components, which are prone to damaging reactive oxygen species (ROS) produced in oxidative stress (OS) states. Alterations in RBC aggregability and adherence have been linked to the pathophysiology in numerous diseases associated with OS. We investigated RBC intercellular interactions in four OS states--thalassemia, treatment of RBC with phenyl-hydrazine or H2O2, and photodynamic virus inactivation of blood units. All these OS states increased RBC adherence to EC, but only part of them elevated their aggregability, while others abolished it. It is proposed that (1) different OS states might induce disparate effects on RBC intercellular interactions; (2) RBC aggregability and adherence to EC, although both intercellular interactions, are controlled by different cell surface factors. PMID- 10711743 TI - Hemorheologic alterations in hypertension: chicken or egg? AB - The serious pathophysiologic consequences of systemic hypertension have prompted numerous basic science and clinical studies related to cardiac or vascular abnormalities. More recently, hemorheologic alterations in hypertension have been explored in an attempt to determine the import of rheologic factors vis-a-vis the elevated total peripheral resistance observed in hypertensive patients. To date, various studies have documented several abnormalities (e.g., increased blood and plasma viscosity, elevated hematocrit), with some suggesting altered RBC or WBC rigidity. However, there is much less information relevant to the "chicken or egg" problems: (1) are reported hemorheological abnormalities the cause or the result of hypertension; (2) does normalization of blood pressure also normalize hemorheological parameters? In overview, the current literature indicates that hemorheology and hypertension may be linked, but that the details of this association and its cause-effect relations remain unclear. PMID- 10711744 TI - Influence of a calcium antagonist on blood rheology and arterial compliance in hypertension: comparison with a thiazide diuretic. AB - Beyond their effects on blood pressure, antihypertensive agents may produce additional effects on blood rheology and arterial compliance abnormalities which may play a role in the target organ damage. These effects may depend only on their specific pharmacological properties. We compared the effects of nitrendipine to hydrochlorothiazide in 33 mild to moderate hypertensives in a double blind parallel group trial. Blood rheology and radial artery diameter and compliance were measured at t=0 and t=2 months. Both drugs produced blood pressure lowering. Blood viscosity decreased in the nitrendipine group and increased in the hydrochlorothiazide patients. Red blood cells deformability increased only in the nitrendipine group. Radial artery diameter and compliance were not different between the two groups but there was a trend to an increase of the cross-sectional compliance in hydrochlorothiazide group and to a decrease in nitrendipine group. Our data show that a calcium antagonist (Nitrendipine) could improve rheological parameters. PMID- 10711745 TI - Hemorheological and hemodynamic parameters in patients with essential hypertension and their modification by alpha-1 inhibitor drug treatment. AB - Hemorheological factors play an important role in the pathogenesis of different cardiovascular diseases. The hemorheological and hemodynamic parameters in essential hypertension and their possible modification by antihypertensive treatment were examined in the following two studies. In the first study the fundus appearance and hemorheological parameters (plasma and whole blood viscosity (WBV), fibrinogen level) of 33 hypertensive patients (mean age: 55 years) were examined. The fundus appearance showed retinopathy in all the cases between stages I-III. All the measured hemorheological parameters of the examined patients were in the pathological range (WBV at 90 s(-1): 5.18 mPa s) and were significantly (p < 0.01) higher than in healthy controls (WBV at 90 s(-1): 4.18 mPa s). The hemorheological factors showed a parallel deterioration with the fundus appearance, namely their values were significantly (p < 0.01) higher in patients with a fundus appearance stage III (WBV at 90 s(-1): 6.02 mPa s) than stage I (WBV at 90 s(-1): 4.51 mPa s). These results show that there is a correlation between hemorheological parameters and fundus appearance in hypertensives, and this suggests that hemorheological factors may play a role in the development of hypertensive retinopathy. In the second study the hemorheological and hemodynamical effects of Doxazosin, a selective alpha-1 adrenoreceptor blocker agent, was examined in twenty patients (mean age: 54 years) with essential hypertension. Hemorheologic (hematocrit, fibrinogen, plasma and whole blood viscosity) and hemodynamic (cardiac output and index, total peripheral resistance) parameters and plasma lipids were determined. The measurements were carried out before the beginning of the treatment, after 1 week and after 12 weeks treatment periods. Besides significant reduction of blood pressure and total peripheral resistance (p < 0.001), a decrease in cholesterol (p < 0.001) and triglycerides (p < 0.01) levels and a beneficial effect on hemorheological parameters was detected. Fibrinogen and plasma viscosity decreased significantly (p < 0.01). Hematocrit value was also lower after one week (p < 0.001), then an increase could be seen. Whole blood viscosity showed similar changes as hematocrit, but the degree of its final increase was slighter, which was supported by the significantly lower value of corrected blood viscosity (p < 0.05). All these findings indicate that hemorheological factors may play a role in the pathogenesis and in the development of organ damages in hypertension. PMID- 10711746 TI - Principal results of hypertension optimal treatment (HOT) study and their clinical impact. HOT cooperative group. PMID- 10711747 TI - Microvascular changes during laboratory stimuli and structural haemodynamic indices: the role of pulse pressure. AB - Office and ambulatory pulse pressure have been recognized as independent predictors of cardiovascular mortality and atherosclerosis in hypertensives as well as in normotensives. On the other hand, the vascular reactivity, in subjects with high pulsatile component of blood pressure, has not been studied yet. The purpose of our study was to identify the regional muscular hemodynamics and the cutaneous microvascular changes during laboratory stimuli in young adult very mild hypertensives with high pulse pressure. The cardiovascular (Finapres), the forearm vascular (plethysmography) and the microvascular cutaneous (laser-Doppler flowmetry and transcutaneous oximetry) responses to psychophysiological stimuli were measured. In addition, the hyperemic forearm vascular response to the ischaemic test was measured as haemodynamic index of vascular damage. We studied 15 very mild hypertensives with higher office pulse pressure and 15 patients with similar age, history of hypertension, metabolic parameters and systodiastolic blood pressure but lower pulse pressure values. Patients with high pulse pressure demonstrated reduced hyperemic response and increased residual vascular resistance at the forearm ischaemic test. They did not vary for all the parameters, except pulse pressure, during the baseline period but the total stress response, as residualized area-under-the-curve, was notably different. Patients with higher office pulse pressure demonstrated a significant increased heart rate, systolic and pulsatile blood pressure reactivity. On the contrary, they showed a reduced forearm and cutaneous blood flow response combined to a reduced transcutaneous tissutal oxygenation. The findings suggest that the increased pulsatile component of blood pressure might be associated to structural and functional vascular impairments since the very early stages of hypertension in young adults without metabolic disorders. PMID- 10711748 TI - Arterial distensibility in hypercholesterolemia and diabetes. PMID- 10711749 TI - Identification of patients at high risk for cerebral stroke: the role of transesophageal echocardiography. PMID- 10711750 TI - Microcirculation and tissue metabolism in peripheral arterial disease. AB - It is now clear that different pathophysiologic mechanisms have a profound influence on the extent of the functional impairment in intermittent claudication. In particular, metabolic derangements, including impaired oxygen delivery and/or extraction, reduced nitric oxide synthesis, reduced glucose oxidation, accumulation of toxic metabolites and reduction in carnitine availability are correlated with disease severity. Therefore, metabolic interventions aimed at counteracting these alterations may represent a valid therapeutic approach to the treatment of this condition. To date, verapamil and L arginine efficacy has been proven in few patients; a large scale clinical trial, conversely, reports that propionyl-L-carnitine appears to be an effective and well tolerated drug for the treatment of intermittent claudication. PMID- 10711751 TI - Membrane fluidity and oxygen diffusion in cholesterol-enriched endothelial cells. AB - In this study, we measured the influence of cholesterol rigidification on oxygen permeability in human endothelial cell monolayer membranes (ECs). Cholesterol induced membrane rigidification was assessed at different membrane depths by a fluorescence polarization method with diphenyl-hexatriene (DPH) and 1-(4 trimethylamino)-6-phenylhexatriene (TMA-DPH). Fluorescence quenching by oxygen was probed in preferentially labelled membrane with pyrene butyric acid (PyC4) and pyrene dodecanoic acid (PyC12), as shown with a 3D fluorescence microscope (CellScan System). With both probes the experiments revealed a decrease in oxygen diffusion as the cholesterol concentration increased in the medium culture (from 3.42 microM to 17.11 microM). We showed that very low concentrations of cholesterol (about 1000 times below normal value, 6.2 mM) particularly decrease oxygen levels or diffusion rate in the middle region of the membrane. In conclusion, these findings prove in a direct manner that cholesterol significantly affect the endothelial barrier function and molecular oxygen transfer to underlying tissues. Risk factors (cholesterol) directly would contribute to tissue ischemia. PMID- 10711752 TI - Invasive and non-invasive PO2 measurements in clinical practice. PMID- 10711753 TI - The arteriolar structural changes and their evolution in hypertension. PMID- 10711754 TI - Endothelin, microcirculation and hemorheology. AB - Although a large amount of data concerning microcirculation and cardiovascular disease exists, little is known about microcirculation and hypertension. This is largely due to the difficulty in selectively examining capillaries and metarterioles, independently from small arteries or large vessels. The physiological role of capillaries and metarterioles, the two elements that make up the microcycle, is peculiar and closely related to metabolic exchange. During the hypertensive state, several factors can alter these mechanisms. These include elevated plasma viscosity, abnormal membrane properties of red blood cells, and an increase in fibrinogen, LTL and hematocrit levels. The question of whether an abnormal release of endothelium derived vasoactive factors from capillaries, or an abnormal production of chemical factors by blood cells running through the vasculature area is present in hypertensives is fascinating, but unfortunately neither experimental nor clinical data has yet been able to answer it. Recently, evidence of the formation of endothelin by red blood cells from endogenous precursors was given, suggesting that red blood cells may modulate the vascular tone both directly, through the release of ATP or endothelin-1, and indirectly, when hemolysis occurs and hemoglobin is released. The pathological significance of these findings has not been clearly demonstrated in hypertension thus far, although it is reasonable to hypothesise that there are clinical implications for the pathogenesis and the progression of vascular damage during the hypertensive state. PMID- 10711755 TI - Clinical potential of in vitro measured red cell deformability, a myth? AB - For many years the study of Red Blood Cell (RBC) deformability has been limited to specialised hematological research institutes and this has hampered a widespread clinical testing of this dynamic RBC property. Consequently, the clinical relevance of such in vitro measurements has remained questionable now for a considerable time. The recent availability of the LORCA, a routinely applicable and computer assisted instrument for this purpose, opens now the possibility to evaluate RBC deformability on a large scale in various pathological situations associated with impaired microcirculatory flow. In this communication we present our clinical experience obtained thusfar with this instrument. Besides the effect of physiological aging of normal RBC, the results of a clinical study on malaria tropica, case studies of hereditary elliptocytosis, Smith-Lemli-Opitz syndrome (a cholesterol biosynthesis defect), the treatment of sickle cell crisis with hydroxy-urea as well as the clinical intervention with Cyclosporin, are collected. In conclusion, it can be stated that the limited clinical experience with the LORCA as is reported here, yields sufficient evidence about the clinical potential of this technique. PMID- 10711756 TI - Evaluation of hemorheological parameters and red cell morphology in hypertension. AB - The aim of our study was to evaluate hemorheological parameters in two groups of patients both suffering from essential hypertension compared with an homologous group of subjects not suffering from hypertension; the 1st group was composed of 16 patients (8 females and 8 males, mean age 65.6 +/- 16.5) suffering from essential hypertension; the 2nd one of 26 patients (8 females and 18 males, mean age 74.3 +/- 11.7) suffering from essential hypertension and cerebral or cardiac ischemia in a chronic phase. The group of controls was composed of 20 subjects (mean age 50.5 +/- 11.5). In all these subjects we evaluated: blood viscosity, hematocrit, plasmatic fibrinogen, red cell morphology according to Zipursky Forconi method and blood pressure. Our results show that blood viscosity and fibrinogen were statistically increased relative to controls. Comparison between these groups leads us to observe that blood viscosity and fibrinogen are slightly higher in the first group in a statistically significant way (7.5 +/- 1.1 cPs 10 s(-1) 1st group, 7.9 +/- 1.05 cPs 10 s(-1) 2nd group; 362.2 +/- 167.3 mg% 1st group, 384.6 +/- 106.9 mg% 2nd group). Blood pressure was higher in the second group. The study of red cell morphology in all the patients showed a prevalence in the percentage of discocytes, cells which have less deformability compared to bowls. The study demonstrated EMI (Erythrocyte Morphology Index) decreased in a statistically significant way compared to controls (0.70 +/- 0.03 1st group; 0.65 +/- 0.02 2nd group; 1.2 +/- 0.09 control group). In subjects suffering from essential hypertension with end-organ damage EMI further decreases. So we observed that hypertension is also delineated by the increase in the discocytes percentage which results in reduction of the red cell deformability. In conclusion, when hypertension causes end-organ damage, the hemorheological alterations, which implicate a worsening in microcirculation, are more evident. PMID- 10711757 TI - Measurement of erythrocyte deformability by two laser diffraction methods. AB - The aim of this work is to study the deformability of red blood cells (RBC) by two laser diffraction methods: the Laser-assisted Optical Rotational Cell Analyser (LORCA, Mechatronics, Amsterdam, Netherlands) and a Shear Stress Diffractometer (RHEODYN SSD, Myrenne, Roetgen, Germany). Experiments were carried out on 46 healthy human subjects. The elongation index EI of normal and hardened RBCs (obtained by heating blood at 49 degrees C or by incubating RBCs in solutions of diamide) was measured. The results showed that the standard deviations of the experimental data for normal RBCs were relatively small, especially at high shear stresses (more than 3.0 Pa), but higher than those reported before. Some correlations between the results given by the two instruments were also found. It should be noted that for hardened RBCs, the standard deviations of the measurements were important compared with the mean values in the two instruments. PMID- 10711758 TI - Comparison of three optical methods to study erythrocyte aggregation. AB - The aim of this work was to evaluate three optical methods designed to determine erythrocyte aggregation: Erythroaggregometer (EA; Regulest, France), Laser assisted Optical Rotational Cell Analyzer (LORCA; Mechatronics, Netherlands) and Fully Automatic Erythrocyte Aggregometer (FAEA; Myrenne, GmbH, Germany). Blood samples were taken from fifty donors (26 males and 24 females). The aggregation of normal red blood cell (RBC) and RBCs suspended in three normo- and hyperaggregating suspending media was studied. The results revealed some significant correlations between parameters measured by these instruments, in particular, between the indexes of aggregation of EA and LORCA. Further, RBC aggregation of multiple myeloma patients was also studied and a hyper erythrocyte aggregation state was found by EA and LORCA. PMID- 10711759 TI - Flow motion. PMID- 10711760 TI - Current approaches to non-invasive optical oximetry. AB - The optical oxymetry methods based on light transmission or diffuse reflectance of living tissues are discussed. The experimental techniques now under test are highlighted. PMID- 10711761 TI - A non invasive optical oximetry in humans: preliminary data. AB - In order to evaluate the oxygen saturation of hemoglobin in living tissues we use a new experimental instrument: the optical oximeter (LOX). The LOX uses 2 LEDs, like the pulse oximeter (PO). One LED operates at 670 nm (Hb absorption), the other one operates at 830 nm, i.e., near the isosbestic point for which Hb and HbO2 show the same extinction. The oxygen saturation of Hb can be expressed as a linear function of the ratio of the absorption coefficients at the wavelengths of the 2 LEDs. Indeed the LOX is based on diffused reflectance measurements and not on light transmission such as the POs. We studied in standard conditions 10 healthy volunteer subjects (6 M, 4 F aged 40 +/- 4 years) non smokers. We used TO (Combi Sensor) at the subclavicular standard area and at the right ankle, PO with the probe always at 2nd finger of the right hand and LOX at right wrist and at right ankle. We obtained a significant relationship between values of TO, PO and LOX (p < 0.01). Our preliminary data suggest that this could be a new methodology to evaluate tissular oxygenation, also exploring living tissues several centimeters deep. PMID- 10711762 TI - Hemorheology in complicated hypertension. AB - During essential and secondary arterial hypertension it is possible to observe changes in microcirculation perfusion associated with a reduction in tissue oxygenation due in part to hemorheological changes such as an increase in blood viscosity or the formation of the red blood cell "rouleaux" which favour an increase in peripheral resistance and can cause or worsen arterial hypertension. We studied 21 healthy subjects (11 male and 10 female aged 42 +/- 4) and 26 hypertensive subjects (14 male and 12 female aged 49 +/- 3). The patients were non smokers and non suffering from respiratory or haemathological pathologies. They were not undergoing antihypertensive or vasodilatory pharmaceutical treatment. The patients suffered from mild hypertension (II WHO) with Peripheral Occlusive Arterial Disease (POAD II "a" acc. to Leriche-Fontaine class.). The patients showed an increase in cholesterolaemia (6.42 +/- 0.81 mmol/l) and trygliceridaemia (2.73 +/- 0.09 mmol/l) at an average level. The patients were studied in standard conditions with a constant temperature of 22 degrees C. We measured SBP, DBP, MBP, and the HR. We also measured the elongation index (EI) (with shear stress range 0.30 to 30 pascals) using LORCA, acc. to Hardeman method (1994), in order to study the erythrocyte deformability and aggregation kinetics in dynamic condition. To evaluate deformability in static conditions we calculated the Erythrocyte Morphologic Index (EMI), acc. to Forconi method, via the bowl/discocyte ratio (for 100 red blood cells fixed in glutaraldehyde at 0.3% and observed with an optical microscope under immersion in glycerol). Peripheral oxygenation was taken transcutaneously (TcpO2). To establish the level of vascular disease we used the Regional Perfusion Index (RPI = TcpO2 foot/TcpO2 subclavean) and doppler guided Winsor Index (WI). The Student "t" test and linear regression were used for the statistical analysis. Our data confirm a reduction in peripheral tissue oxygenation in hypertensives especially if suffering from vascular disease which correlates significantly (p < 0.01) with a reduction in red blood cell deformability. This itself can increase peripheral resistances and favour the onset of hemorheological complications, at a cerebral-vascular level, which are frequent in hypertensives. PMID- 10711763 TI - Arterial hypertension decreases fibrinogen molecules contribution to the inter red cells connections in stroke patients. AB - In a group of 36 patients with acute ischemic stroke, the study of influence of arterial hypertension on the red cells and fibrinogen interaction was carried out by the aspect of fibrinogen molecules contribution to this phenomenon, respecting the role of other plasma biochemical factors. Patients were divided into two clinical groups: with and without arterial hypertension. In the blood samples with stable haematocrit ratio the following rheological properties were estimated: plasma viscosity, yield shear stress (YSS), fibrinogen level and parameter R (the difference between twice measurements of fibrinogen concentration). There were also measured the levels of albumin, IgG, IgA, IgM, cholesterol, HDL and LDL. The value of parameter R in patients with arterial hypertension was lower than the group without accompanying disease (statistical significance was indicated for 80% and 60% of plasma dilution). The first group of patients was also characterized by significant increase of IgA level (in comparison with the control group), whereas the red cells and fibrinogen interaction measured as a value of YSS was similar in both analyzed clinical groups. This study indicated a specific character some of hemorehological changes in relation to a certain vascular pathology. PMID- 10711764 TI - Effect of controlled ethanol intake on arterial blood pressure, heart rate and red blood cells deformability. AB - Although blood pressure is in most cases determined genetically, environmental factors such as obesity or alcohol consumption are implicated in the development of hypertension. On the other hand it is well known that alcohol drinking and hypertension are associated with reduction of red blood cells deformability. AIM: To compare the effect of ethanol on arterial pressure and red cell deformability in well controlled experimental conditions. METHODS: Five healthy volunteers, men with age range between 26-38 underwent ethanol consumption 1.0 g per kg of body weight during 10 min period. Before and 90 min, 3 h and 5 h after ethanol consumption arterial pressure, heart rate, blood morphology, alcohol and acetaldehyde blood level and deformability index by Rheodyn SSD were measured. RESULTS: After ethanol consumption rise in ethanol and acetaldehyde blood levels and no changes in blood pressure, but significant increase in heart rate were observed. Elongation index of red blood cells at low shear rates has not been changed. At shear stresses of 12.0, 30.0 and 60 Pa significant increase in elongation index of erythrocytes after 1.5 hour of ethanol intake had occurred. PMID- 10711765 TI - Tissular oxygenation and venoarteriolar reflex disturbances in diabetes mellitus: vasoregulator effect of Buflomedil. AB - The present study was designed to investigate whether the Veno-Arteriolar Reflex (VAR) mediated via a local nervous reflex mechanism may be used as a microvascular approach to predict the effect of vasoactive drugs in diabetic patients. The vasoactive drug we studied here was Buflomedil. The effect of a single infusion of 400 mg of Buflomedil was examined on VAR and on transcutaneous oxygen pressure (TcPO2). Investigations were performed in 42 diabetic patients. The VAR was assessed on dorsal foot and dorsal big toe by measuring changes in skin blood flux induced by lowering the leg. TcPO2 was measured on dorsal foot. Before Buflomedil infusion, patients were characterized by a loss of the VAR in comparison to healthy volunteers. The loss of the VAR was associated to significant decreases in TcPO2 values. Buflomedil infusion led to significant increases in VAR at the two sites of measurement and also in TcPO2 values. These findings indicate that the VAR can be used as a sensitive microvascular test, as it allows to detect the effect of Buflomedil. Furthermore, our findings demonstrate that the Buflomedil-induced improvement in VAR is identical in the two diabetic groups with or without complications. This result emphasizes the benefit of Buflomedil not only in diabetics with microangiopathy or those suffering from a peripheral vascular disease for reducing pain or healing trophic lesions, but also in those patients without any clinically detectable macro- or microangiopathy in order to prevent or to reduce as long as possible the risk of developing diabetes related complications by the normalization of functional microangiopathy. PMID- 10711766 TI - Erythrocytes stored in CPD SAG-mannitol: evaluation of their deformability. AB - The erythrocytes deformability of three blood samples coming from healthy donors has been evaluated by Laser-assisted Optical Rotational Cell Analyzer (LORCA). Blood samples were stored in CPD SAG-mannitol. The study of progressive reduction in the Elongation Index (EI) during the preservation may be used as way of evaluation of erythrocyte vitality in stored blood samples. PMID- 10711767 TI - Oxidative stress is an early marker of endothelial dysfunction? PMID- 10711768 TI - Hemorheological aspects in hypertensive menopausal smoker women treated with female hormones. AB - In postmenopausal hypertensive women (PostMHW) the erythrocyte deformability (ED) is reduced if compared with premenopausal hypertensive women (PreMHW). This might partially explain the increased incidence of cardiovascular diseases (CD) in hypertensive women after menopause. Moreover a positive correlation exists between estradiol and rheological patterns in women. If PostMHW smoke cigarettes, there is an important decrease in hemorheological parameters. On the other hand if PostMHW are submitted to an hormonal replacement therapy (HRT) they can show controversial results with an impairment if hemorheological parameters. The aim of this study was to evaluate the influence of smoking and HRT on PostMHW. We studied four groups of subjects: Group 1: PreMHW (10 F aged 35 +/- 3 years) non smokers; Group 2: PostMHW (8 F aged 45 +/- 2 years) non smokers; Group 3: PostMHW (14 F aged 48 +/- 4 years) smokers (20 cigarettes per day); Group 4: PostMHW (16 F aged 50 +/- 2 years) smokers (20 cigarettes per day) submitted to HRT. We evaluated Elongation Index of erythrocytes under torsion force of 30 pascals (EI- 30 Pa) using a new computerized instrument Laser assisted Optical rotational Red Cell Analyzer (LORCA) (Mechatronics, Hoorn, NL) acc. to Hardeman (1994) and, also with the same LORCA, Aggregation Index (AI), t(1/2). We measured the transcutaneous oxygen partial pressure (TcpO2) in subclavicular standard area using a Transcutaneous Oximeter (Microgas 7650 Kontron Instruments with Combi Sensor) and total cholesterolaemia. In PostMHW our data showed a significant (p < 0.01) impairment of hemorheological patterns and tissue oxygenation if compared with PreMHW (Group 1). In Group 3 there is a significant (p < 0.01) decrease in EI, a significant (p < 0.01) increase in AI, a significant (p < 0.01) decrease in t(1/2) and TcpO2 if compared with Control Group 1 and Group 2. Finally a further significant (p < 0.01) impairment in hemorheology and tissue oxygenation showed Group 4. In conclusion, it seems necessary, that many studies will be performed to understand really protective action of HRT on cardiovascular diseases in PostMHW and it is necessary to suppress cigarette smoking to prevent cardiovascular complications in these patients. PMID- 10711769 TI - The choroidal circulation after retinal detachment surgery. AB - PURPOSE: to assess the effects on choroidal perfusion of scleral buckling and vitrectomy procedures for retinal detachment. METHODS: 17 patients were included in the study: 7 underwent scleral buckling and 10 vitrectomy, for rhegmatogenous retinal detachment. pOBF Langham system evaluation was performed before and after surgery (the fellow eye served as control). Statistical analysis was based on paired t-test. RESULTS: in the scleral buckling group was found a mean change (SD) of 470 (355.32) microl/min; in the vitrectomy group the mean difference (SD) was 254.2 (132.53) microl/min. In both groups a statistically significant difference with the control was found. CONCLUSIONS: our results show that both surgical procedures affect choroidal perfusion and could impair optic nerve head feeding. PMID- 10711770 TI - The role of erythrocytapheresis in secondary erythrocytosis therapy. AB - In chronic respiratory insufficiency secondary erythrocytosis (SPC), causing pulmonary hypertension and dx ventricular insufficiency, is often noticed. An alternative therapy to phlebotomy for SPC is isovolemic large volume erythrocytapheresis performed with cell separator (CSE) in order to quickly remove a large volume of red blood cells (RBC) while saving plasma proteins and clotting factors. In order to evaluate the efficiency and safety of CSE in SPC we reported a retrospective analysis of our experience with 61 SPC patients: from April 1996 to May 1998 we performed 208 CSE using Haemonetics MCS3P (TAE protocol). Before every apheresis procedure we verified Hb (in median 18.8 g/dl), Ht (in median 58.4%), viscometry, coagulation test, EGA, PFR and ECG. 11 patients were treated with 1 CSE, 12 with 3, 29 with 4 and 9 with 5. The mean volume of RBC removed was 576 ml (range 426-800); Hb post CSE averaged 14.4 g/dl and Ht post CSE averaged 42.7%; hematic viscosity post CSE was significantly reduced while tissue oxygen tension increased: the improvement of symptomatology and hematochemical parameters was maintained on the average for 6.5 months. All the procedures were well tolerated and light side effects (paresthesias citrate depending in 27 apheresis) were easily controlled. CSE, compared to phlebotomy, has the advantage of selectively removing RBC without loss of clotting factors, platelets and plasma proteins. Although CSE has relatively high costs we noticed a decrease of hospital recurrence (about 50-65%) in SPC patients treated with apheresis. PMID- 10711771 TI - Gender difference in rheologic properties of blood and risk of cardiovascular diseases. AB - According to official statistical data there is a significant difference between pre-menopausal women and age-matched men in morbidity and mortality from cardiac diseases and especially from myocardial infarction. There are several speculations regarding the nature of this phenomenon which have both supporting and refuting evidence. Our hypothesis was that due to regular physiologic bleeding, rheological properties of blood of pre-menopausal women are superior to those of men, and place such women at a lower risk of cardiovascular diseases than men in any age group. We believe that this difference in hemorheological properties is due to the reduced concentration of red blood cells (RBCs) and due to greater population of younger and less population of older RBCs in female blood. We studied mechanical properties of blood from 47 pre-menopausal women and 50 age-matched men. Compared to female blood, male blood had higher viscosity and RBC aggregation and lower RBC deformability. Oxygen Delivery Index, calculated as a ratio of hematocrit to blood viscosity, was found to be significantly lower in male blood. Decreased oxygen delivery along with increased RBC aggregation and decreased RBC deformability may contribute to the higher risk for the development of cardiovascular diseases. Regular blood donation may reduce hematocrit and blood viscosity, improve rheological properties of blood, and increase oxygen delivery in men. PMID- 10711772 TI - Microcirculatory damage of common carotid artery wall in obese and non obese subjects. AB - The objective of the present study was to determine whether the intima-media thickness (IMT) is independently related with obesity, and central fat accumulation in healthy subjects. Common carotid artery IMT, parameters of body fat accumulation and distribution (body mass index, waist circumference, waist-to hip ratio), blood pressure levels, and circulating fasting insulin, glucose, and lipid (cholesterol, HDL-cholesterol, triglycerides, LDL-cholesterol) levels were determined in a population of non-diabetic normal weight and obese subjects. Smoking habits (packs-years) were also taken into account. 239 healthy subjects (143 women and 96 men), with age ranging between 18 and 45 years, were enrolled into the study. They were divided indo two groups according to the body mass index (BMI), obese (132 subjects, 77 woman and 55 men, with BMI greater than 27.0) and controls (107 subjects: 66 women and 41 men, with BMI lower than 27.0). Common carotid artery intima-media thickness was measured by B-mode ultrasound imaging. Fasting plasma metabolic parameters (glucose and lipids) and insulin levels were determined by enzymatic and radioimmunological assays, respectively. Insulin sensitivity was estimated by insulin tolerance test (ITT) and the rate constant for plasma glucose disappearance (KITT) during the 3- to 15-min period following the regular insulin injection was taken as a measure of in vivo insulin action. Obese patients showed higher IMT than controls, and IMT was significantly associated with BMI in the whole population (r = 0.316, p < 0.001). Age (r = 0.327, p < 0.001), KITT (r = -0.201, p < 0.01), fasting blood glucose (r = 0.187, p < 0.01), LDL-chol (r = 0.201, p < 0.01), smoking (r = 0.147, p < 0.05), MBP levels (r = 0.154, p < 0.05), cholesterol (r = 0.152, p < 0.05) and HDL-chol (r = -0.159, p < 0.05) were also significantly associated with IMT. Age (r = 0.330, p < 0.05), BMI (r = 0.299, p < 0.01), waist (r = 0.312, p < 0.001), WHR (r = 0.266, p < 0.001) and KITT (r = -0.259, p < 0.01) were the parameters most strongly correlated with IMT in women, and age (r = 0.324, p < 0.001), BMI (r = 0.338, p < 0.001) waist (r = 0.325, p < 0.001) and LDL-chol (r = 0.283, p < 0.01) where the parameters most strongly correlated with IMT in men. When a stepwise multiple regression analysis was performed for the whole population, only age (p < 0.001) and BMI (p < 0.001) maintained a significant positive relationship with IMT. When a stepwise multiple regression analysis was performed separately for men and women, BMI or waist circumference or WHR were alternatively entered into the model; interestingly, only age, BMI and waist were still significantly correlated with IMT, whereas WHR did not maintain a significant correlation with IMT. In conclusion, BMI and waist circumference, but not WHR, are strongly and independently associated with the IMT of common carotid artery. These results suggests that central fat accumulation may accelerate the development of earlier clinically silent stages of atherosclerosis, thus possibly explaining the higher prevalence of cardiovascular diseases in patients with abdominal obesity. PMID- 10711773 TI - Modelisation of leukocyte adhesion on a fibrinogen coated surface in static conditions. AB - The adhesion of polymorphonuclear leukocytes (PMNs) on the vascular endothelium is a complex process that occurs during biological and pathological events and involves a large family of molecules. This phenomenom could be approached by a modelisation study of the adhesion of PMNs on a biological substrate, fibrinogen. Two different physiological conditions were tested such as the activated state of PMNs with a synthetic pro-inflammatory activator (N-Formyl-Methionyl-Leucyl Phenylalanine, FMLP). The activated state of PMNs was both quantified by flow cytometry and controlled by fluorescence microscopy. The results suggest that quiescent PMNs deposit in accordance with the ballistic deposition model. The preliminary results obtained with FMLP-stimulated PMNs show a different deposit process compared to quiescent PMNs but do not allow to determine exactly a deposition model. PMID- 10711774 TI - Morphometric characteristics of red blood cells as diagnostic factors for coronary artery disease. AB - Digital imaging microscopy plays an important role in computer aided cytological and histological diagnosis. In this study, digital imaging microscopy was used to measure Red Blood Cells (RBCs) morphometric characteristics such as area, perimeter, major and minor axis length, elongation, compactness and roughness in both groups: healthy individuals and Coronary Artery Disease (CAD) patients. All the measured characteristics, except elongation, showed a statistically significant difference between the two groups. In particular, patients with CAD presented greater values than healthy ones. Finally multivariate analysis was used in order to take into account the whole measured profile of each patient. Compactness, perimeter and elongation improved the diagnostic ability, whereas all the others did not show any significant improvement. These three characteristics correctly classify the 84.5% of the patients. In conclusion, RBCs morphometric characteristics could be considered as diagnostic factors for CAD. PMID- 10711775 TI - Application of near-infrared tissue oxymetry to the diagnosis of peripheral vascular disease. AB - Near-infrared spectroscopy (NIRS) is a noninvasive technique to measure the tissue oxygenation in real time. This optical method has many advantages over the invasive analysis currently used for clinical tests. Among the possible applications of near-infrared oxymetry, we report three protocols (exercise, venous occlusion and tilting table) in conjunction with NIRS, and discuss their applicability in the diagnosis of peripheral vascular disease (PVD). PMID- 10711776 TI - Investigation on cerebral hemodynamics in patients with carotid disease receiving carotid endarterectomy. AB - 100 patients (pts) receiving CEA (carotid endarterectomy) were evaluated in this study. In some of them postoperative complications were observed, characterized by TIA (transient ischemic attack) and, mostly, by cerebral hyperperfusion. In only two of the pts investigated CEA needed the implantation of a shunt, due to the emergence of intolerance signs at carotid Clamping (C) evaluated by TCD (transcranial Doppler); the preoperative cerebral angiography in the two subjects in question did not show malformations of the circle of Willis. The hyperperfusive phenomenon and the absence of intracranial compensation flows during CEA seemed to be ascribed to a more or less severe impairment of cerebral reserve. Such impairment of the autoregulatory capacity seems to be crucial to the pathogenesis of hemodynamic stroke. Thus the indication to CEA, in pts with severe carotid disease, should take into account also the cerebral reserve (CR) impairment to prevent both thromboembolic and hemodynamic stroke. The predictive and diagnostic role of TCD turns out to be crucial in assessing and selecting pts candidate to CEA. PMID- 10711777 TI - Hemorheological disturbances and characteristic parameters in patients with cerebrovascular disease. AB - Whole blood and plasma viscosity were measured of 90 patients with chronic cerebrovascular disease (CVD) and 56 healthy individuals by Couette rotational viscometer Contraves Low Shear 30 at a steady flow. Plasma viscosity was measured with capillary viscometer of Ubbelohde type. Two subgroups of patients were investigated: 23 patients with transient ischemic attacks (TIAs) and 67 patients with chronic cerebral infarctions (CCI). They were compared with two control groups: 56 healthy individuals without risk factors for stroke and 37 randomly selected subjects with risk factors for stroke. It is established significant elevation of plasma viscosity in the patients with cerebral ischemia. The elevation of blood viscosity was most pronounced at shear rate of 94.5 s(-1). This comparison is confirmed by the criterion, using blood rigidity number h defined by the formula of Whittington and Harkness. Conclusion is drawn from our study that chronic ischemic cerebrovascular disorders are characterized with chronic hyperviscosity. PMID- 10711778 TI - Effects of carotid thromboendarterectomy on circulating endothelin-1. AB - The aim of this study was to investigate the behaviour of circulating ET-1 in patients with carotid atherosclerosis, before and after carotid thromboendarterectomy (TEA), to test the hypothesis that plasma ET-1 decreases after removal of atherosclerotic lesion. Plasma immunoreactive ET-1 levels were determined in 17 patients with symptomatic and/or hemodynamically significant carotid atherosclerosis on the day before TEA, 48 h and 72 h after surgery and, in 11 of them, also after 8 h and 24 h. Compared to controls, ET-1 levels were significantly higher both before and after TEA; after carotid revascularisation (8 h) ET-1 increased; then, from the 24th h, ET-1 gradually decreased and at the 48th h and 72th h the decrease was significant in front of basal values. The increase of plasma ET-1 in the acute postoperative phase may reflect the degree of surgical stress and manipulation in diseased blood vessels; the following decrease may indicate the improvement of vascular dysfunction in the involved carotid site; the persistence of high ET-1 levels 72 h after surgery could suggest the presence of residual ischemia in the involved district and/or the involvement of other sites in ET-1 production. PMID- 10711779 TI - Haemorheological and fibrinolytic activity in hypertensive Nigerians. AB - Hypertension is the most significant cardiovascular risk factor to emerge in developing countries. 64 hypertensives (40 old hypertensives and 18 newly diagnosed hypertensives) and 40 age and sex matched normotensives controls were studied. Plasma fibrinogen concentration (PFC), euglobulin lysis time (ELT) and relative plasma viscosity (RPV) and relative whole blood viscosity (RWBV) were estimated. The PFC, RPV, RWBV and ELT were significantly higher in new and old hypertensives (P < 0.005), respectively, compared with controls, while the RPV was significantly higher (P < 0.005) in new hypertensives than old hypertensives. The significantly higher PFC, RPV and ELT in new hypertensives correspond to higher BP in the new hypertensives than old hypertensives. There were consistence significance levels in PFC, RPV and ELT in both female hypertensives and controls (P < 0.005), respectively, with higher BP in male than female. Though there were no significant mean differences between the hypertensive patients in the first two years of the disease, there were cumulative consistent increase in fibrinogen levels and euglobulin lysis time from two years; while RPV shows inconsistence variation until the 5th year. We conclude that a defective rheology and fibrin clearing mechanism may contribute to aetiology of vascular complications in hypertensive patients especially in the long term. PMID- 10711781 TI - Reduced deformability of erythrocytes as feature of congenital dyserythropoietic anaemia type II (HEMPAS). AB - In this study we report results regarding erythrocytes deformability in congenital dyserythropoietic anemia type II (HEMPAS) by the use of LORCA (Laser assisted Optical Rotational Cell Analyzer). The reduced erythrocytes deformability observed in seven case of CDA II is caused by changes in the structure of glycoproteins due to the incomplete glycosylation of erythrocytic and erythroblastic membrane. Erythrocytes deformability (EI) was shown to be inversely related with mean corpuscular volume (MCV) and mean haemoglobin concentration (MCH). PMID- 10711780 TI - Tissular oxygenation and erythrocyte deformability in haemodialyzed patients using different dialytic membranes. AB - We studied the effect of some dialytic membrane on tissular oxygenation (TO) and erythrocyte deformability (ED). Sixteen patients (10 M and 6 F, aged 59 +/- 12 years) have been submitted to bicarbonate dialysis (BD) and subdivided into four groups (GR) of 4 patients each: GR 1 (hemophan membrane, 35 BD), GR 2 (polyacrylonitrile, PAN AN 69, 42 BD), GR 3 (polysulphone, 38 BD) and GR 4 (polycarbonate, 37 BD). The TO has been detected with the transcutaneous oxygen pressure (Tc pO2) using a transcutaneous oxymeter and the ED has been evaluated with the EMI (Erythrocyte Morphometric Index), which results from the ratio between deformable erythrocytes (bowl shape) and rigid erythrocytes (discocyte shape), for every 100 red cells fixed in vitro with 0.3% glutaraldehyde. The ED was also evaluated using a laser instrument: Laser Optical Rotational Cell Analyser. During BD was observed a significant decrease of Tc pO2 in the 1st hour only in the 1st and 2nd GR and, in contrast with results obtained in the 3rd and 4th GR, in the same GRs the EMI showed a significant reduction of ED at the end of BD. Finally the LORCA results, showing a significant decrease of ED only in the 1st and 2nd GR, confirmed the data obtained with EMI. In conclusion, our study has suggested that hemophan and PAN AN 69 are less biocompatible than polysulphone and polycarbonate membranes according to effects on ED and TO. PMID- 10711782 TI - LDH-C4: a unique target of mammalian spermatozoa. PMID- 10711783 TI - Chemical modulation of activity in steroidal estrogens. PMID- 10711784 TI - Use of sterol mutants as probes for sterol functions in the yeast, Saccharomyces cerevisiae. PMID- 10711785 TI - Integration of the metabolic pathways of steroids, carotenoids, and retinoids. PMID- 10711786 TI - Plasma coenzyme Q10 concentrations are not decreased in male patients with coronary atherosclerosis. AB - Coenzyme Q10 (CoQ10) is an important mitochondrial electron transfer component and has been postulated to function as a powerful antioxidant protecting LDL from oxidative damage. It could thus reduce the risk of cardiovascular disease. Thus far, beneficial effects of supplementation with CoQ10 have been reported. To study the relation between unsupplemented concentrations of plasma CoQ10 and coronary atherosclerosis, we performed a case-control study among 71 male cases with angiographically documented severe coronary atherosclerosis and 69 healthy male controls free from symptomatic cardiovascular disease and without atherosclerotic plaques in the carotid artery. Plasma CoQ10 concentrations (mean +/- SE) were 0.86+/-0.04 vs. 0.83+/-0.04 micromol/l for cases and controls, respectively. The CoQ10/LDL-cholesterol ratio (micromol/ mmol) was slightly lower in cases than in controls (0.22+/-0.01 vs. 0.26+/-0.03). Differences in CoQ10 concentrations and CoQ10/LDL-cholesterol ratio did not reach significance. The odds ratios (95% confidence interval) for the risk of coronary atherosclerosis calculated per micromol/l increase of CoQ10 was 1.12 (0.28-4.43) after adjustment for age, smoking habits, total cholesterol and diastolic blood pressure. We conclude that an unsupplemented plasma CoQ10 concentration is not related to risk of coronary atherosclerosis. PMID- 10711787 TI - Urinary 8-oxo-7,8-dihydro-2'-deoxyguanosine and 5-(hydroxymethyl) uracil in smokers. AB - Cigarette smoke is known to generate free radicals by various mechanisms. In this study involving 30 non-smokers and 30 smokers, we show that urinary excretion of 5-(hydroxymethyl) uracil (HMUra) was not different in the two groups (6.54+/-2.07 vs. 6.70+/-1.68 nmol/mmol creatinine). In contrast, 8-oxo-7,8-dihydro-2' deoxyguanosine (8-oxo-dGuo) excretion increased by 16% (1.16+/-0.35 vs. 1.35+/ 0.50 nmol/mmol creatinine, p = 0.039). Results concerning 8-oxo-dGuo are in agreement with those of previous studies. We observed significant multiple correlations between HMUra and creatinine (r(p) = 0.44), BMI (r(p) = -0.27) and nicotine derivatives (r(p) = 0.26). Multiple correlation analysis showed relations between 8-oxo-dGuo on the one hand, and: creatinine (r(p) = 0.36), nicotine derivatives (r(p) = 0.29), BMI (r(p) = -0.24) on the other. PMID- 10711788 TI - Prooxidant and antioxidant properties of Trolox C, analogue of vitamin E, in oxidation of low-density lipoprotein. AB - Trolox C (Trolox), a water-soluble analogue of vitamin E lacking the phytyl chain, was investigated with respect to its effect on the oxidation of low density lipoprotein (LDL). Trolox was added at different time points of LDL oxidation induced by Cu2+ and aqueous peroxyl radicals. In the case of Cu2+ induced LDL oxidation, the effect of Trolox changed from antioxidant to prooxidant when added at later time points during oxidation; this transition occurred whenever alpha-tocopherol was just consumed in oxidizing LDL. Thus, in the case of Cu2+ -dependent LDL oxidation, the presence of lipophilic antioxidants in the LDL particle is likely to be a prerequisite for the antioxidant activity of Trolox. When oxidation was induced by peroxyl radicals, as a model of metal-independent oxidation, the effect of Trolox was always antioxidant, suggesting the importance of Cu2+ /Cu+ redox-cycling in the prooxidant mechanism of Trolox. Our data suggest that, in the absence of significant amounts of lipophilic antioxidants, LDL becomes highly susceptible to oxidation induced by transition metals in the presence of aqueous reductants. PMID- 10711789 TI - Plasma chain-breaking antioxidants in preterm infants with good and poor short term outcome. AB - Many complications of prematurity have been suggested to result from free radical generation and an inadequacy of antioxidative capacity. We measured the plasma total peroxyl radical-trapping capability (TRAP) and concentrations of the main chain-breaking antioxidants contributing to it, i.e. uric acid, ascorbic acid, alpha-tocopherol, protein sulfhydryl groups and bilirubin, in 21 preterm infants with a mean birth weight of 1440 g and gestational age of 30 wk. The infants were divided into two groups according to their short-term outcome; the good outcome group (GOG) (N = 11) with no signs of morbidity and the poor outcome group (POG) (N = 10) with intraventricular haemorrhage and/or bronchopulmonary dysplasia and/or retinopathy. Arterial blood samples were obtained 3 and 10 days postpartum. TRAP was measured with a chemiluminescent method. As a comparison, venous blood samples from 13 adults (aged from 18 to 34) were used. At day 3 the poor outcome group had significantly higher TRAP than the good outcome or control group, mainly because of elevated uric acid concentration. Also the concentration of unidentified antioxidants was significantly lower in GOG. By day 10 the TRAP decreased substantially in both groups. However, from the components of TRAP, both ascorbate and the unidentified fraction decreased more in POG (p = 0.017 and 0.021, respectively). Furthermore in POG on day 10 urate concentration did not significantly differ from day 3 values. In conclusion, in preterm infants high TRAP was associated with high plasma uric acid concentration and a poor short term prognosis. PMID- 10711790 TI - Dietary docosahexaenoic acid enhances ferric nitrilotriacetate-induced oxidative damage in mice but not when additional alpha-tocopherol is supplemented. AB - Weaning mice were fed a diet supplemented with beef tallow (BT) or BT plus docosahexaenoic acid (DHA) containing 100 mg alpha-tocopherol/kg (alpha-Toc100) or 500 mg alpha-tocopherol/kg (alpha-Toc500) for 4 wk to modify membrane fatty acid unsaturation, and then were administered ferric nitrilotriacetate (Fe-NTA). The mortality caused by Fe-NTA was higher in the group fed the DHA (alpha-Toc100) diet than in the BT diet groups but the DHA (alpha-Toc500) diet suppressed this increase. Serum and kidney alpha-tocopherol contents were slightly influenced by the dietary fatty acids but not significantly. These results indicate that the increased unsaturation of tissue lipids enhances oxidative damage induced by Fe NTA in mice fed DHA (alpha-Toc100) but not when additional alpha-tocopherol is supplemented. The apparent discrepancy between the observed enhancement by dietary DHA of oxidative damage and the beneficial effects of dietary DHA on the so-called free radical diseases is discussed in terms of strong bolus oxidative stress and moderate chronic oxidative stress. PMID- 10711791 TI - Regeneration of phenolic antioxidants from phenoxyl radicals: an ESR and electrochemical study of antioxidant hierarchy. AB - Radicals from the flavonoids quercetin, (+)-catechin, (+/-)-taxifolin and luteolin, and from all-rac-alpha-tocopherol have been generated electrochemically by one-electron oxidation in deaerated dimethylformamide (DMF), and characterised by electron spin resonance spectroscopy (ESR) after spin-trapping by 5,5-dimethyl 1-pyrroline-N-oxide (DMPO). Simulations of the ESR spectrum based on estimated coupling constants of the spin-trapped quercetin radical, confirmed that this antioxidant radical is oxygen-centered. The complex mixture of radicals, quinoid intermediates and stable two-electron oxidation products, were for each antioxidant allowed to react with each of the four other antioxidants, and the progression of reaction followed by ESR after addition of DMPO, and the product solution further analysed by HPLC. All-rac-alpha-tocopherol was found to be most efficient in regenerating each of the other antioxidants from their oxidation products with a regeneration index (defined as moles regenerated of the oxidised phenolic antioxidant divided with moles of all-rac-alpha-tocopherol consumed) of 0.90+/-0.16 for quercetin, 0.48+/-0.11 for (+)-catechin, 0.48+/-0.06 for (+/-) taxifolin and 0.50+/-0.10 for luteolin in equimolar 1.00 mM solution. Quercetin was found to have the highest regeneration index among the flavonoids: 0.88+/ 0.13 for (+/-)-catechin, 0.41+/-0.03 for (+/-)-taxifolin and 0.41+/-0.02 for luteolin. The antioxidant hierarchy based on the reduction potentials determined by cyclic voltammetry under similar conditions (0.93 V for all-rac-alpha tocopherol, 1.07 V for quercetin, 1.15 V for luteolin, 1.16V for (+)-catechin and 1.20 V for (+/-)-taxifolin) is compared with the observed over-all regeneration (34% for quercetin, 34% for (+)-catechin, 52% for (+/-)-taxifolin and 43% for luteolin by all-rac-alpha-tocopherol). PMID- 10711792 TI - Apoptotic macrophage-derived foam cells of human atheromas are rich in iron and ferritin, suggesting iron-catalysed reactions to be involved in apoptosis. AB - We investigated the presence of low-molecular-weight iron and ferritin in human atheromas, and their possible relation to the apoptotic process. Arterial wall segments with fatty streaks were collected from coronary arteries and thoracic aortas of 12 clinical autopsy cases with general atherosclerosis. Normal appearing regions from the same cases together with normal coronary arteries from seven young forensic autopsy cases, without any sign of atherosclerosis, were used for comparison. Anti-CD68 (macrophage marker) and anti-ferritin antibodies were applied to serial sections of the arterial wall segments, fixed in formadehyde and embedded in paraffin wax, using an avidin-biotin complex (ABC) technique. Similarly, apoptotic cells were assayed by the TUNEL technique, while low-molecular-weight iron was cytochemically detected by autometallography. Cell counting and computerised image analysis were performed to compare the distribution of macrophages, ferritin- and iron-rich cells, and apoptotic cells in the intima, media, and adventitia of the arteries. Pronounced ferritin accumulation, occurrence of lysosomal low-molecular-weight iron, and apoptosis mainly concerned CD68-positive cells (macrophages) in the atherosclerotic lesions. No ferritin- or CD68-positivity was found in normal coronary arteries from the young forensic-autopsy cases, while a moderate number of such cells were observed in the intima of normal looking vessel areas from the control cases. In the intima, cytosolic ferritin and low-molecular-weight iron with a lysosomal type distribution were found in many CD68-positive macrophages which frequently were surrounded by erythrocytes. A substantial number of apoptotic cells within the intima, media, and adventitia were registered in all atherosclerotic lesions examined, although mainly in the vulnerable macrophage-enriched areas of the atheroma shoulder. We suggest that iron may occur within the cytosol, mainly bound in ferritin, but also in low-molecular weight, redox-active form within the acidic vacuolar apparatus of macrophages and macrophage-derived foam cells following erythrophagocytosis or phagocytosis of apoptotic cells. Low-molecular weight iron within lysosomes, present due to degradation of iron-containing structures, such as ferritin, may partially become exocytosed and contribute to cell-mediated LDL-oxidation. Moreover, such lysosomal iron may also sensitise lysosomes to oxidative stress and induce apoptosis of macrophage/foam-cells that may result in instability and rupture of atherosclerotic plaques. PMID- 10711793 TI - Antioxidant activities of carotenoids: quantitative relationships between theoretical calculations and experimental literature data. AB - Quantitative structure activity relationships (QSARs) are described for the antioxidant activity of series of all-trans carotenoids. The antioxidant activity of the carotenoids is characterised by literature data for (i) their relative ability to scavenge the ABTS*+ radical cation, reflected by the so-called trolox equivalent antioxidant capacity (TEAC) value, (ii) their relative rate of oxidation by a range of free radicals, or (iii) their capacity to inhibit lipid peroxidation in multilamellar liposomes, leading to a decrease in formation of thiobarbituric acid reactive substances (TBARS). All these antioxidant values for radical scavenging action correlate quantitatively with computer-calculated ionisation potentials of the carotenoids. These correlations are observed both when the ionisation potential is calculated as the negative of the energy of the highest occupied molecular orbital (-E(HOMO)) of the molecule, or as the relative change in heat of formation (deltadeltaHF) upon the one-electron oxidation of the carotenoids. The calculations provide a theoretical assay able to characterise the intrinsic electron donating capacity of an antioxidant, in hydrophilic, hydrophobic or artificial membrane environment. PMID- 10711794 TI - Interaction of hydroxycinnamic acid derivatives with the Cl3COO radical: a pulse radiolysis study. AB - The electron transfer reactions between the trichloromethylperoxyl radical (Cl3COO*) and hydroxycinnamic acid derivatives, including chlorogenic acid, sinapic acid, caffeic acid, ferulic acid and 3,4-(methylenedioxy)cinnamic acid, have been studied by pulse radiolysis. The hydroxycinnamic acid derivatives, especially sinapic acid, are identified as good antioxidants for reduction of Cl3COO* via electron transfer reactions. From buildup kinetic analysis of phenoxyl radical, the rate constant for reaction of Cl3COO* with sinapic acid has been determined to be 8.2x10(7) dm3 mol(-1) s(-1), while the rate constants of electron transfer from other hydroxycinnamic acid derivatives to Cl3COO* were obtained to be about 2x10(7) dm3 mol(-1) s(-1). The reaction of 3,4 (methylenedioxy) cinnamic acid with Cl3COO* was investigated as an evidence for the electron transfer mechanism. PMID- 10711795 TI - Seasonality of birth and ventricular enlargement in chronic schizophrenia. AB - Many studies have established that birth dates during the winter and early spring months are more common in schizophrenic patients than in the general population. It has been hypothesized that children born in winter are more likely to be exposed to environmental factors which could lead to the development of schizophrenia later in life. Another finding of interest has been the demonstration in brain-imaging studies that mild ventricular enlargement is more often found in schizophrenic patients than in healthy control subjects. In the present report, an increased incidence of ventricular enlargement was found in schizophrenic patients born in the winter months. Although the relationship between seasonality of birth and brain abnormalities is unclear, these phenomena could be partly linked. PMID- 10711796 TI - Echogenicity of the brainstem raphe in patients with major depression. AB - A novel transcranial ultrasound technique was used to detect differences in the echogenicity and echotexture of the brainstem dorsal raphe nucleus in 20 patients with major depression compared with 20 age- and sex-matched healthy adults. Transcranial color-coded real-time sonography visualized the mesencephalic and pontine brainstem and its midline structure. The echogenicity of the raphe was classified in a four-point scale. Compared with healthy subjects, the depressed patients were characterized by a significant decrease in the echogenicity of the brainstem raphe. The echogenicity score was not correlated, however, with measures of psychopathology such as the Hamilton Rating Scale for Depression, the Clinical Global Impression Scale, the Global Assessment Scale, or the Depression Scale of von Zerssen. These preliminary findings suggest that the brainstem raphe may be involved in the etiopathogenesis of major depression. The echogenicity score should be further evaluated as a possible trait marker in different types of affective disorders. PMID- 10711797 TI - Corpus callosum morphology from magnetic resonance images in Tourette's syndrome. AB - We measured the midline cross-sectional area and other morphologic features of the corpus callosum (CC) from magnetic resonance (MR) images in 14 unmedicated patients with Tourette's syndrome (TS) and 14 normal control subjects matched for age, sex, handedness, and socioeconomic status. Each CC was manually circumscribed on midline images from a T1-weighted sagittal series, and the area of the entire CC and five anatomic subdivisions were measured. CC circumference, regional width, and mean callosal curvature were also measured. CC cross sectional area correlated positively with brain size and basal ganglia volumes. The magnitude of reduction (17.7%) in total CC area in TS patients compared with control subjects was similar to the reductions seen in all CC subdivision areas. Analyses of covariance with total midsagittal cross-sectional head area as a covariate revealed the reductions to be statistically significant for the overall CC area and all subregion areas. CC width tended to be nonsignificantly thinner in all subdivisions (from 5% to 11%), and the overall length of the center line measured from rostrum to splenium was significantly reduced in the TS group (by 5.3%). Measures of mean callosal curvature suggested that CCs in TS patients are less rounded than those of normal control subjects. Worst-ever motor tic symptoms showed the strongest significant correlation with the length of the CC center line in TS patients (r = 0.88). These findings suggest that structural interhemispheric connectivity may be aberrant in the central nervous systems of TS patients, and they provide indirect supportive evidence for the presence of altered cerebral lateralization in the disorder. PMID- 10711798 TI - Increased limbic blood flow and total sleep deprivation in major depression with melancholia. AB - Single photon emission computed tomography (SPECT) with technetium-99m-d,l hexamethyl-propylene amine oxime (99Tcm-HMPAO) was carried out in 20 melancholic patients before and after total sleep deprivation. Findings in 11 responders to total sleep deprivation (defined by > or = 40% improvement on the Hamilton Rating Scale for Depression) were compared with findings in nine nonresponders. On the basis of a semiquantitative evaluation of SPECT findings, responders showed relative hyperperfusion before sleep deprivation in the right anterior cingulate cortex and in the right and left fronto-orbital cortex and basal cingulate gyrus. Responders who showed > or = 50% improvement also showed hippocampal overactivation before sleep deprivation. It is possible that limbic overactivation may characterize depressed responders to total sleep deprivation as a distinct subtype. Another possibility is that the pattern of limbic hyperactivation reflects the increased number of bipolar patients in the responder group, with response to total sleep deprivation being only a covariate of this bipolar-unipolar distinction. PMID- 10711799 TI - Dopamine D2 receptor occupancy measured by single photon emission computed tomography with 123I-Iodobenzamide in chronic schizophrenia. AB - Single photon emission computed tomography (SPECT) with 123I-iodobenzamide (123I IBZM) was used to study 22 chronic schizophrenic patients. The patients, who were receiving maintenance therapy with typical neuroleptics, had not shown any significant improvement since their admission to the hospital. Basal ganglia/frontal cortex ratios of the uptake of 123I-IBZM did not show significant differences on the basis of neuroleptic dosage in chlorpromazine equivalents. There were, however, significant differences in 123I-IBZM uptake in the basal ganglia among patients characterized by negative, mixed, and positive symptoms of schizophrenia. Although only a small number of patients had shown a positive response to treatment by the time of discharge, D2 receptor blockade was significantly higher in responders than in nonresponders. In addition, there was an inverse correlation between reduced activation as measured by the Brief Psychiatric Rating Scale and the basal ganglia/frontal cortex ratio. These findings suggest a complex pathogenetic link between the blockade of dopamine D2 receptors and psychopathology in chronic schizophrenic patients. SPECT studies with 123I-IBZM appear to have prognostic value in identifying chronic schizophrenic patients who respond poorly to neuroleptic treatment. PMID- 10711800 TI - Healing fear and hatred. PMID- 10711801 TI - Caring through relation and dialogue: a nursing perspective for patient education. AB - Nursing's recognition of the importance of patient education is evident in nursing's history and in the work of early nurse theorists. However, to date, there has been no clear articulation of a nursing framework specific to this essential aspect of clinical care. A nursing perspective for patient education based on a conceptualization of caring is presented. This perspective includes explication of philosophic views for nursing and patient education. A middle range theory for patient education is generated from this philosophic premise. PMID- 10711802 TI - Healing as appreciating wholeness. AB - The "clinicalization" of human experience by the health care disciplines has been instrumental in denying important facets of human life and not fully accounting for the essence and wholeness of experience. A unitary conceptualization of healing as appreciating wholeness is proposed. Appreciating is actualized through a praxis approach used for research and practice--unitary pattern appreciation- created to bring the theoretical principles of unitary science into practical reality. It is essentially a praxis for exploration of the wholeness within human/environmental pattern. Healing is conceptualized as the realization, knowledge, and appreciation of the inherent wholeness in life that elucidates prospects of clarified understanding and opportunities for action. The essential features of the process of pattern appreciation are a synoptic stance toward pattern information, a participatory engagement with people in the exploration of wholeness, and the transformative nature of the process that illuminates the possibilities in wholeness. A case demonstrates the nature of the unitary pattern appreciation process and its healing qualities. PMID- 10711803 TI - Womanist ways of knowing: theoretical considerations for research with African American women. AB - Research designs that are congruent with theoretical frameworks of African American women are important. However, many researchers remain unfamiliar with womanist thought or are unsure of how it can be used to inform specific aspects of research design. The article explicates a womanist epistemologic framework that can undergird the development of intervention designs aimed at assisting African American women incorporate health-promoting behaviors into their lives. PMID- 10711804 TI - The practices of mothering in caregiving an adult son with AIDS. AB - One invisible and silent phenomenon associated with the acquired immune deficiency syndrome (AIDS) epidemic is the return of mothers to care for their adult sons who are dying of the disease. This article presents an emergent fit of data from an interpretative study with 14 such mothers into Leonard's practices of mothering framework. Conceptualizing mothering as a practice rather than a technical skill provides a context for understanding nurture and care. The mothers' stories reveal moral content of mothering that is centrally important to cultural life, as well as implications for nursing practice. PMID- 10711805 TI - Investing in self-care: a midrange theory of self-care grounded in the lived experience of low-income HIV-positive white women. AB - Little is known about the types of interventions that invite low-income women into partnerships that motivate self-care practices when living with human immunodeficiency virus/acquired immune deficiency syndrome (HIV/AIDS). The increasing incidence of HIV infection in low-income women with histories of inattention to self-care calls for nursing theories that address self-care practices. The purpose of this article is to describe a midrange theory developed from grounded theory research and to discuss implications of theoretical construction for future knowledge development. For the 12 women in this study, self-care practices developed over time and through four categories: focusing self, fitting resources, feeling emotions, and finding meaning. The core category, investing in self-care, linked the categories and carried explanatory power for developing midrange theory. Implications for nursing knowledge development through partnerships with low-income women are discussed. PMID- 10711806 TI - Commonalities in women's spirituality and women's health. AB - Both women's spirituality and women's health movements have grown dramatically in recent years. If clinicians understood in greater depth the commonalities between these two perspectives, then they would be better positioned to foster the health of women more fully. In this article, concepts of feminism, religion, spirituality, and women's health are described briefly. After identifying some assumptions, themes, and characteristics of both women's spirituality and women's health, the commonalities between these two perspectives are delineated. Next, processes critical to women's spirituality and women's health are proposed. Finally, implications for clinical practice are offered. PMID- 10711807 TI - Relationships between nurses and family caregivers: partners in care? AB - Increasing reliance on family care of elderly people at home calls for a critical analysis of the relationship between formal and informal caregivers. Although much has been written about how health professionals and family caregivers should relate to one another, we know very little about the relationships that develop between them. Using data from a qualitative study, this article illustrates that relationships between community nurses and family members caring for frail elders are complex, dynamic, and multifaceted. Shifting boundaries in caring work leads to changes in nurse-family caregiver relationships, which can be categorized as four distinct, yet interconnected, types: (1) nurse-helper, (2) worker-worker, (3) manager-worker, and (4) nurse-patient. Each type is described, and implications for nursing practice and research are discussed. PMID- 10711808 TI - Opioid modulation of magnocellular neurosecretory cell activity. AB - Magnocellular neurosecretory cells of the hypothalamic supraoptic and paraventricular nuclei secrete the hormones, oxytocin and vasopressin, into the systemic circulation from the posterior pituitary gland. Oxytocin is important for parturition and is essential for lactation. Vasopressin regulates body fluid homeostasis. The secretion of these hormones is altered in response to peripheral stimuli that are conveyed via projections from other parts of the brain. Endogenous opioid peptide systems interact with the magnocellular neurosecretory system at several levels to restrain the basal secretion of these hormones as well as their secretory responses to various physiological stimuli. The inhibition of basal secretion can occur at the level of the neurosecretory terminals where endogenous opioids inhibit the release of oxytocin, and at the cell bodies of magnocellular cells to modulate the activity pattern of vasopressin cells. The responses of the magnocellular neurosecretory system to physiological stimuli are also regulated by these mechanisms but in addition probably also by pre-synaptic inhibition of afferent inputs to magnocellular cells as well as direct effects on the cell bodies of afferent input cells to modulate their activity. Here, we review the mechanisms and functional consequences of opioid interactions with oxytocin and vasopressin cells. PMID- 10711809 TI - Varied duration of congenital hypothyroidism potentiates perseveration in a response alternation discrimination task. AB - The behavior of five groups of rats (seven rats per group) made hypothyroid for varying lengths of time and one group of seven normal control rats was assessed under forced alternation fixed-ratio (FR1, FR3, FR5 and FR10), alternating lever cyclic-ratio (ALCR) and progressive-ratio (PR3) schedules of reinforcement. Hypothyroidism was produced by adding methimazole (MMI) to the drinking water of pregnant dams from embryonic day E16 to postnatal day P25. Four groups were given replacement thyroxine (T4) injections beginning at specific time points (P1, P7, P13, and P19). There were no differences in behavioral performance between control and experimental groups under the FR schedule, which indicates that the animals' sensorimotor abilities were intact. Under the forced ALCR schedule, all groups reached criteria similarly. However, under the choice lever ALCR schedule, control animals and those which received T4 replacement from early on (P1, P7, P13 groups) performed well and all had reached criteria by 11 sessions. In contrast, animals which did not receive any T4 replacement or received it late (P19 group) took longer to reach criteria and 5/14 animals had not reached criteria at all by 20 sessions. This deterioration in performance was paralleled by an increase in perseverative behavior as evidenced by an increased frequency of pressing the wrong lever when alternation of lever was required. This suggests that congenital hypothyroidism results in increased perseveration leading to a decrease in learning when a discrimination between correct and incorrect operanda is made available. PMID- 10711810 TI - Na+/Ca2+ exchanger activity induces a slow DC potential after in vitro ischemia in rat hippocampal CA1 region. AB - In rat hippocampal CA1 neurons recorded intracellularly from tissue slices, a rapid depolarization occurred approximately 5 min after application of ischemia simulating medium. In extracellular recordings obtained from CA1 region, a rapid negative-going DC potential (rapid DC potential) was recorded, corresponding to a rapid depolarization. When oxygen and glucose were reintroduced after generating the rapid depolarization, the membrane further depolarized and the potential became 0 mV after 5 min. Contrary, the DC potential began to repolarize slowly and subsequently a slow negative-going DC potential (slow DC potential) occurred within 1 min. A prolonged application of ischemia-simulating medium suppressed the slow DC potential. Addition of a high concentration of ouabain in normoxic medium reproduced a rapid but not a slow DC potential. The slow DC potential was reduced in low Na+- or Co2+-containing medium, but was not affected in low Cl-, high K+ or K+-free medium, suggesting that the slow DC potential is Na+-and Ca2+ dependent. Ni2+ (Ca2+ channel blocker as well as the Na+/Ca2+ exchanger blocker) and benzamil hydrochloride (Na+/Ca2+ exchanger blocker) reduced the slow DC potential dose-dependently. These results suggest that the slow DC potential is mediated by forward mode operation of Na+/Ca2+ exchangers in non-neuronal cells, and that reactivation of Na+, K+-ATPase is necessary to the Na+/Ca2 +exchanger activity. PMID- 10711811 TI - Functional and pharmacological properties of GABArho1delta51 receptors. AB - Gamma-aminobutyrate is the main inhibitory neurotransmitter in the vertebrate brain, and the gamma-aminobutyric acid (GABA) receptor subunit GABArho1delta51 is an alternatively spliced form of the GABArho1 receptor that was recently isolated from human retina cDNA libraries. The rho1delta51 receptor subunit lacks 17 amino acids in the extracellular N-terminal domain and, when expressed in Xenopus oocytes, forms functional homomeric GABA receptors. Unexpectedly, even after a such a big deletion, the fundamental properties of the deleted variant receptors are very similar to those of the complete GABArho1 receptors. For example, both types of receptors are bicuculline resistant, desensitize very little, and are negatively modulated by Zn2+ and positively modulated by La3+. In spite of such similarities, the GABArho1delta51 receptors are more sensitive to GABA, to the specific GABA(C) antagonist (1,2,5,6-tetrahydropyridine-4-yl)methylphosphinic acid and to Zn2+, than the complete GABArho1 receptors. The GABArho1delta51 receptors extend the variety of inhibitory receptors in the retina. Their functional significance still remains to be determined. PMID- 10711812 TI - Modulation of extracellular glutamate and pressor response to muscle contraction during NMDA-receptor blockade in the rostral ventrolateral medulla. AB - Recently our laboratory demonstrated increases in extracellular glutamate concentrations within the rostral ventrolateral medulla (RVLM) during static muscle contraction (Caringi, D.C., Maher, T., Chaiyakul, P., Asmundsson, G., Ishide, T., Ally, A. Pflugers Arch. Eur. J. Physiol., 435:465-471, 1998). In this study, we determined effects of microdialyzing D(-)2-amino-7-phosphonohepatanoic acid (AP-7), an NMDA-receptor antagonist, into the RVLM on changes in mean arterial pressure (MAP), heart rate (HR), and extracellular glutamate levels during muscle contraction in anesthetized rats. Bilateral placements of microdialysis probes into the RVLM were verified by perfusing L-glutamate and obtaining a pressor response. Muscle contraction for 2 min, increased MAP and HR by 22+/-4 mmHg and 28+/-5 bpm, respectively. Extracellular glutamate as determined by microdialysis increased from 0.8+/-0.2 to 6.3+/-1.2 ng/5 microl. Microdialysis of AP-7 (1.0 microM) for 30 min inhibited contraction-evoked MAP and HR responses (10+/-3 mmHg and 13+/-3 bpm) and attenuated increases in glutamate during muscle contraction. Developed tensions did not differ during contractions before and after AP-7. Results demonstrate that NMDA-receptor blockade in the RVLM inhibits cardiovascular responses during static muscle contraction via a reduction in extracellular glutamate levels. PMID- 10711813 TI - Lithium plus pilocarpine induced status epilepticus--biochemical changes. AB - The aim of the study was to investigate the changes in biochemical mechanisms facilitating cellular damages in the lithium plus pilocarpine treatment and the resulting status epilepticus. The whole brain free fatty acid (FFA) level as well as the activities of superoxide dismutase (SOD), glutathione peroxidase (GPX), glutamate dehydrogenase, aspartate-aminotransferase (AST), alanine aminotransferase, gamma-glutamoyl transferase, alkaline phosphatase (ALP), lactate dehydrogenase (LDH) and creatine kinase in the frontal cortex, cerebellum, hippocampus and pons-medulla region of Hannover-Wistar rats were determined. The control group was intact with no previous experimental history. LiCl (125 mg/kg i.p.) was injected 20 h prior to pilocarpine (30 mg/kg i.p.) and the treated rats were sacrificed 1 or 2 1/2 h after pilocarpine administration. The results show that lithium plus pilocarpine administration and the resulting status epilepticus produced the significant increase of the brain FFA content. Decreased GPX activities were detected in the frontal cortex, cerebellum and hippocampus of the treated rats without the accompanying decrease of SOD activity. Increased AST and LDH activities were observed in the frontal cortex, increased soluble ALP activities in the frontal cortex and pons-medulla region whereas the increased activity of membrane bound ALP was detected in the hippocampus of the rats with status epilepticus. Activities of the other analysed enzymes did not change in the examined brain regions. The presented data indicate clear regional differences of biochemical changes caused by lithium plus pilocarpine treatment and the resulting status epilepticus, frontal cortex being the most affected site. PMID- 10711814 TI - Non-muscle myosin IIB-like immunoreactivity is present at the drebrin-binding cytoskeleton in neurons. AB - Dendritic spines are extremely motile, providing a structural mechanism for synaptic plasticity. Actin-myosin interaction is thought to be responsible for the change in the shape of spine. We have already reported that drebrin, an actin binding protein, inhibits actin-myosin interaction and is enriched in the dendritic spine of mature neurons. In this study, we prepared the actin cytoskeleton of dendritic spines as an immunoprecipitate with anti-drebrin antibody from adult guinea-pig brain, immunized mice with the cytoskeleton, and obtained a monoclonal antibody (MAb) called MAb G650. MAb G650 reacted with non muscle myosin IIB, but it did not react with muscle myosin II or non-muscle myosin IIA. Immunoblotting with this antibody revealed that drebrin-binding cytoskeleton contains this myosin IIB-like immunreactivity. Immunohistochemistry using MAb G650 demonstrated that this myosin IIB-like immunreactivity can be detected in the neuronal cell bodies and their apical dendrites, where drebrin is hardly detected. These data demonstrate that a myosin subtype associated with drebrin-binding actin filaments in the dendritic spines is myosin IIB, although this myosin is widely distributed in somato-dendritic subdomains of neurons. Furthermore, it is indicated that the cytoskeletons in dendritic spine were uniquely characterized with actin-binding proteins such as drebrin, but not with myosins. PMID- 10711815 TI - Relationship of high-density lipoprotein cholesterol and red blood cell filterability: cross-sectional study of healthy subjects. AB - The deformability of red blood cells (RBCs) is an important rheologic factor in the maintenance of normal blood flow in the microcirculation. Contrary to the well-known relationship between hyperlipidemia and atherosclerosis, the relationship between RBC rheology and the serum lipid profile has remained controversial and obscure. Moreover, the correlation of high-density lipoprotein (HDL)-cholesterol and RBC deformability has not been fully understood. In the present cross-sectional study of 139 apparently healthy subjects, we investigated whole-cell deformability (filterability) of RBCs in relation to the lipid profile, using a nickel mesh filter with 3.2-microm pores. RBC filterability was independent of gender, age and serum levels of low-density lipoprotein (LDL) cholesterol. The filterability was significantly proportional to the HDL cholesterol values (r = 0.382, p < 0.01), whereas it was inversely proportional to the triglyceride levels (r = -0.259, p < 0.01). These findings may provide new insight into the role of HDL-cholesterol not only in preventing atherosclerotic progression but also in improving RBC filterability. PMID- 10711816 TI - Comparative values of erythrocyte aggregability versus other indices of hemorheological disorders in patients with ischemic brain infarcts. AB - The aim of the present study is comparison of changes of principal hemorheological factors responsible for blood flow disorders in the microcirculation in patients with ischemic brain infarcts. It was found that in venous blood samples the erythrocyte aggregability (examined with a direct, highly sensitive, quantitative technique) was considerably more increased (by mean of 120%) than the blood plasma fibrinogen contents, blood plasma viscosity, and hematocrit which increased only by 23.7%, 7.4% and 3.6%, respectively, as compared to the same hemorheological factors in the healthy controls. These results led us to the following conclusion: among the other tests the enhanced erythrocyte aggregability, when measured with an appropriate technique, is the best diagnostic indicator of hemorheological derangements during development of the ischemic brain infarct. PMID- 10711817 TI - Erythrocyte deformability in Waldenstrom's macroglobulinemia. AB - The deformability of erythrocytes in Waldenstrom's Macroglobulinemia (WM) patients are analysed by shear stress diffractometer in vitro. Erythrocytes obtained from blood samples of WM patients are subjected to well defined shear forces, varying from 0.3 to 60 Pa, and their corresponding elongation indexes generated by diffraction pattern of a laser beam are recorded. For comparison, equal number of samples from normal adult volunteers is also studied. The results show a shear dependent decrease in deformability index in WM patient samples. The decrease in deformability index of WM patients is significant at high shear forces (>12 Pa) when compared to those of normal samples whereas it is not significant at low shear forces. As the deformability of erythrocytes together with elevated plasma viscosity and enhanced erythrocyte aggregation may cause potential flow stagnation at microvasculature, these studies are clinically relevant. PMID- 10711818 TI - Morning increase of whole blood viscosity in obstructive sleep apnea syndrome. AB - OBJECTIVE: Patients affected by obstructive sleep apnea syndrome (OSAS) have an increased risk of cardiovascular diseases such as stroke and myocardial infarction. The pathophysiological mechanisms leading to increased vascular risk are still matter of debate. A relative morning hyperviscosity could be one of the leading mechanisms of cardiovascular morbidity which is actually known to be especially high in the morning hours. METHODS: Whole blood viscosity (WBV) at seven shear rates, ranging from 0.47 to 118 sec(-1), haematocrit (Hct), and plasma fibrinogen (F) concentration, were measured on venous blood samples in 12 patients with OSAS and in 8 healthy controls at 8-9 p.m. and at 7-8 a.m. the morning after. WBV values were normalized on Hct by the computation of the standardised normal deviate z on the normal database of the laboratory. RESULTS: No changes were observed in controls. Hct, F and normalized WBV (independently from Hct changes) significantly increased in the morning hours in OSAS patients. CONCLUSIONS: Viscosity of whole blood increases in the morning in OSAS patients but not in healthy controls. This condition may be related to the increased susceptibility to cerebral ischemia in patients affected by OSAS, particularly evident in the early morning. PMID- 10711819 TI - Leukocyte flotation during gravity sedimentation of the whole blood. AB - The original Westergren blood sedimentation technique was modified to assess leukocyte sedimentation properties. The relative change of leukocyte and erythrocyte counts was measured in the upper half section of blood column in vertically positioned sedimentation tubes in 10-minute-intervals for 60 minutes. During the first 20 minutes of gravity sedimentation, the leukocytes taken from critically ill patients showed upward flotation, however, healthy individuals' leukocytes demonstrated slight sedimentation. The upward flotation rate of leukocytes seemed less dependent on erythrocyte sedimentation during the first 15 minutes of sedimentation time than after it. Based on this observation, the sedimentation properties of leukocytes were characterized by the leukocyte antisedimentation rate taken at the 15th minute of sedimentation time (LAR15). Erythrocyte aggregability index, plasma fibrinogen concentration and native leukocyte count did not correlate to LAR15 in healthy volunteers (n = 25). However, LAR15 was correlated to leukocyte adherence (p < 0.01), to whole blood viscosity (p < 0.05), to hematocrit (p < 0.05) and to the conventional erythrocyte sedimentation rate (p < 0.05). PMID- 10711820 TI - Deformability and filterability of white blood cell subpopulations. Evaluation of these parameters in the cell line HL-60 and in type II diabetes mellitus. AB - The rheological properties of human leukocytes (WBCs) have been studied using the micropipette aspiration and the filtration technique. Partial micropipette (i.d. 2.8-4.5 microm) aspiration of individual leukocytes under constant aspiration pressure of 8 mm H20 and measurement of the aspirated length as a function of time (creep experiments) according to the Evans model have been carried out and the apparent viscosity mu(app) was estimated. In the filtration experiments, using the Hemorheometer, the Index Rigidity of Leukocytes, ILR, was also estimated. The apparent viscosity mu(app) of normal PMN and MNC was significantly different p < 0.05), while the LYM and PMN had no statistical difference (p < 0.5). The leukocytes of the cell line HL-60 were more rigid than the normal PMN (p < 0.01), while the PMN from patients with type II diabetes mellitus were more rigid than the normal PMN (p < 0.005). The results of IRL showed similar differences among all of the leukocyte subpopulations. Comparison of these findings suggests a possible relationship between ILR and mu(app) which in this case is: ILR = 598 + 0.54 mu(app) (r = 0.986, p-value 0.0003). PMID- 10711821 TI - Effect of creatine on erythrocyte rheology in vitro. AB - To evaluate how creatine influences erythrocyte deformability, we determined its effect on erythrocyte filterability in 9 subjects with insulin dependent diabetes (IDDM) without complications, 14 diabetics with uremia and 10 non-diabetic controls. The short-term incubation (15 min at 37 degrees C) of diabetic erythrocytes with 3 mM creatine improved cell filterability (assessed according to the Reid method) from IDDM subjects without complications by 28.4% and that from diabetics with uremia by 18.9%. No rheological effect of creatine was found in erythrocytes from non-diabetic controls. However, a significant protective effect against erythrocyte filterability impairment induced by treatment of red blood cells from non-diabetic controls with hydrogen peroxide was observed with 3 mM (p < 0.04) and 5 mM (p < 0.01) creatine, respectively. Measurement of the thiobarbituric acid (TBA) reactivity was used to assess hydrogen peroxide induced formation of malondialdehyde (MDA). We found that creatine inhibits hydrogen peroxide-induced erythrocyte MDA-formation in a dose dependent manner by 20.4%, 22.3% and 41.4% for 1, 3 and 5 mM creatine, respectively. These results suggest that creatine by its ability to inhibit erythrocyte lipid peroxidation may contribute to the maintenance of normal cell deformability. PMID- 10711822 TI - Hypervolumetric hemodilution with HES 100/0.5 10% in patients with peripheral arterial occlusive disease (Fontaine, stage II): an open clinical and pharmacological phase IV study. AB - The efficacy of three weekly interventions with hypervolumetric hemodilution of a new preparation of hydroxyethyl starch (HES 100/0.5, 10%, C2/C6 substitution ratio of 6.5) on pain-free walking distance of patients with peripheral arterial occlusive disease (PAOD) stage IIb on the Fontaine classification was investigated. In addition quantitative data on the pharmacokinetic properties of this HES preparation, and it's impact on hemorheology, hemostasis and homeostasis were shown. Ten patients were included according to a predefined protocol, and treated openly with 500 ml HES 100/0.5 10% on nine occasions over 18 days. Pain free walking distance, the main outcome measure, showed a mean increase of 82 m (+60%). Hematocrit decreased 4 percentage points on average (5.5 percentage points one hour after interventions). Plasma viscosity dropped 5% on average with significant changes immediately after interventions only in patients whose baseline values had been equal to or above the 2 s reference area. Erythrocyte aggregation decreased by 16% in the course of treatment (8% immediately after interventions), systolic blood pressure by 13%, and total protein by 7%. Complement showed a trend towards lower values (-20%), and creatinine, pH and urine viscosity remained unchanged. Apart from complement changes, all reductions mirrored the dilution effects. As to pharmacokinetics, serum mean molecular weight distribution was very similar to that of the infusion. A minor adverse drug reaction (light, spontaneously disappearing pruritus) was observed in one case. PMID- 10711823 TI - Decreased mechanical stability of neonatal red cell membrane quantified by measurement of the elastic area compressibility modulus. AB - Previous studies suggest that early loss of membrane might account for the shortened life-span of neonatal red blood cells (RBC). The elastic area compressibility modulus describes the force required to achieve a defined expansion of the membrane and is thus a suitable intrinsic material property to describe mechanical stability of the RBC membrane. We studied RBC in eight cord blood samples (representing fetal blood), ten 5-day old full-term neonates and seven healthy adults. The RBC were suspended in hypotonic buffer solution and investigated using a modification of the micropipet technique by Evans et al. Cord blood RBC (204+/-33 dyn/cm) and neonatal RBC (209+/-11 dyn/cm) showed a 25% lower elastic area compressibility modulus than adult cells (278+/-26 dyn/cm; p < 0.05). Thus, less force was necessary for expansion of neonatal RBC membrane. We conclude that neonatal RBC are more susceptible to mechanical damage of their membrane than adult cells. PMID- 10711824 TI - Deliberation on the scientific triumvirate of clinical hemorheology, biorheology and microcirculation. An essay. PMID- 10711825 TI - Leukocytes in ovarian function. AB - It has become apparent that in the ovary, the immune system contributes to the regulation of gonadal function. Leukocytes present within the ovary may constitute potential in-situ modulators of ovarian function that act through local secretion of regulatory soluble factors. These factors include numerous cytokines that largely originate by the action of immune cells within the ovary. Actual rupture of the follicle during ovulation may be dependent on tissue remodelling that is characteristic of an acute inflammatory reaction and includes mobilization of thecal fibroblasts, increased leukocyte migration, release of various mediators and loosening of connective tissue elements in the follicle wall. Both corpus luteum formation and luteal regression also involve progressive infiltration of lymphocytes and macrophages, release of chemokines and cytokines, and communication through cell adhesion molecules. In this review, we examine the evidence for the leukocytes and their products in regulation of ovarian function and relate the potential significance of these cells and substances to some ovarian disorders. PMID- 10711826 TI - Endometrial leukocytes and menstruation. AB - This review examines evidence supporting the concept that menstruation occurs as a result of an inflammatory process. In the endometrium, leukocyte numbers rise in the late secretory phase following the fall in serum progesterone concentrations. It is postulated that products released following activation of these leukocytes are critically important for menstruation. Mast cells, eosinophils, neutrophils and macrophages in particular are involved. Endometrial granular lymphocytes may also play a role, although their increase in numbers is somewhat earlier during the menstrual cycle than that of the others, suggesting perhaps a primary role in embryo implantation. Leukocyte products include a range of proteases, chemokines and cytokines which in concert result in focal production and activation of matrix metalloproteinases by endometrial cells and the subsequent breakdown of tissue that characterizes menstruation. Regulation of leukocyte entry, proliferation, differentiation and activation within the endometrium is not yet well understood, although both chemokines and cytokines produced locally by endometrial cells are clearly implicated. The role of progesterone in regulating these events is still not understood although the lack of progesterone receptors on endometrial leukocytes suggests indirect actions. PMID- 10711827 TI - Uterine leukocytes and decidualization. AB - The transformation of endometrium into decidua is an essential feature of normal implantation and pregnancy. There is a close association with an unusual leukocyte population, uterine natural killer (NK) cells, and onset of decidualization. These uterine NK cells are seen in close contact with stromal cells ultrastructurally and are also seen encircling vessels and glands. The possibility that uterine NK cells in the late secretory phase and in early decidua may be important in initiating and maintaining decidualization is raised. In contrast, the death of uterine NK cells could be an early event in the onset of endometrial breakdown at menstruation. The period between implantation and menstruation (7-14 days after luteinizing hormone surge) is the time when implantation is known to be particularly vulnerable. In this review, the possibility that uterine leukocytes might influence the critical decision that the mid- to late secretory endometrium must make either to decidualize or to undergo menstruation is explored. PMID- 10711828 TI - Heritability and molecular genetic studies of endometriosis. AB - Endometriosis is a common disease defined as the growth of endometrial tissue outside the uterine cavity that often results in a vast array of gynaecological problems including dyspareunia, dysmenorrhoea, pelvic pain and infertility. Despite the increasing evidence that supports a genetic component to this common gynaecological condition, the basic aetiology and pathogenesis of endometriosis remain unknown. It is likely that endometriosis is a common polygenic/multifactorial disease caused by an interaction between multiple genes as well as the environment. Such conditions do not have a clear Mendelian pattern of inheritance. Recent molecular cytogenetic studies on endometriotic tissue and an established endometriosis-derived cell line provide novel evidence that acquired chromosome-specific alterations may be involved in endometriosis, possibly reflecting clonal expansion of chromosomally abnormal cells. Molecular DNA studies examining the role of loss of heterozygosity in endometriotic lesions has identified candidate tumour suppressor gene loci, including 5q, 6q, 9p, 11q and 22q, that may play a role in the malignant transformation of endometriotic implants to endometrioid ovarian cancers. Evidence of mutations in the tumour suppressor PTEN gene in the endometrioid subtype of epithelial ovarian cancer further suggests that somatic genetic alterations represent early events in the transformation of benign endometriotic cells. Genetic factors are also likely to influence individual susceptibility to endometriosis. There is now evidence that heritable allelic differences in drug-metabolizing enzymes play an important role in the development of endometriosis. Further studies are warranted to identify major susceptibility gene(s) and the mechanism involved in endometriosis to assist in the development of better methods for early detection, diagnosis and prevention. PMID- 10711829 TI - Vascular endothelial growth factor and endometriotic angiogenesis. AB - Peritoneal endometriosis is a significant debilitating gynaecological problem of widespread prevalence. It is now generally accepted that the pathogenesis of peritoneal endometriosis involves the implantation of exfoliated endometrium. Essential for its survival is the generation and maintenance of an extensive blood supply both within and surrounding the ectopic tissue. The vascular endothelial growth factor (VEGF) family of angiogenic molecules is involved in both physiological angiogenesis, and a number of pathological conditions that are characterized by excessive angiogenesis. Increasing evidence suggests that the VEGF family may also be involved with both the aetiology and maintenance of peritoneal endometriosis. Sources of this factor include the eutopic endometrium, ectopic endometriotic tissue and peritoneal fluid macrophages. Important to its aetiology is the correct peritoneal environment in which the exfoliated endometrium is seeded and implants. Established ectopic tissue is then dependent on the peritoneal environment for its survival, an environment that supports angiogenesis. Our increasing knowledge of the involvement of the VEGF family in endometriotic angiogenesis raises the possibility of novel approaches to its medical management, with particular focus on the anti-angiogenic control of the action of VEGF. PMID- 10711830 TI - Pituitary-ovarian dysfunction and endometriosis. AB - Significant association between endometriosis, including minor endometriosis, and infertility (or strictly subfertility) is shown by prevalence studies, but a causal relationship has not been established. This review focuses on evidence for pituitary-ovarian dysfunction as a cause for the subfertility. A methodological problem is lack of fertile controls with endometriosis. Group comparison with tubal infertility cases as controls have demonstrated: impaired follicular growth, reduction in circulating oestradiol concentrations during the pre ovulatory phase and of oestradiol and progesterone during the early luteal phase, and disturbed luteinizing hormone (LH) surge patterns. Reduction in LH concentrations in pre-ovulatory follicular fluid has also been found, and granulosa cells collected at the same time have demonstrated impaired steroidogenic capacity in vitro, but these were not consistently proven findings. Pooled data from published studies show significantly reduced oocyte fertilization rates (49%) compared with controls (69%), even after maximal stimulation with exogenous follicle stimulating hormone (FSH) and human chorionic gonadotrophin (HCG) (54 versus 69%). The implantation rate is also slightly (though significantly) reduced (11 versus 13%). The findings suggest an inherent disorder of follicular function, with LH surge impairment probably being a secondary phenomenon. The resulting reduction in the chance of fertilization of the oocyte would contribute substantially to the subfertility associated with endometriosis. It seems that the benefit of in-vitro fertilization is gained through the excessive number of oocytes obtained by stimulation. PMID- 10711831 TI - Follicular hormonal environment and embryo quality in women with endometriosis. AB - The use of in-vitro fertilization (IVF) as a therapeutic tool in patients with endometriosis has provided information about the disease and, in particular, aspects of the reproductive process in humans, particularly folliculogenesis, fertilization, embryo development and implantation. Retrospective analyses of IVF and oocyte donation programmes showed impaired implantation in patients with endometriosis. Otherwise, the observation that embryo development was blocked more frequently in cases of endometriosis, suggested that impaired embryo quality may be responsible for the reduced implantation rates. Similarly, women with the disease undergoing oocyte donation had the same chance of implantation as patients without endometriosis, suggesting that the endometrial milieu is not affected in patients with endometriosis. The quality of the oocyte may, therefore, be altered in patients with endometriosis. To investigate this, we studied steroid secretion in women undergoing IVF. Progesterone concentrations in follicular fluid increased with the severity of the disease and an increase in progesterone accumulation in vitro was observed in basal and human chorionic gonadotrophin (HCG)-induced granulosa cell cultures. We postulated that the pattern of progesterone secretion may be related to the release of cytokines by ovarian and/or immune cells. To test this, we measured interleukin (IL)-1, IL-6 and vascular endothelial growth factor (VEGF) concentrations in serum, follicular fluid and granulosa cell cultures. IL-6 concentrations in serum were increased in natural cycles in women with endometriosis and showed a significant decrease in stimulated cycles in IVF. Also, IL-6 concentrations were increased in the follicular fluid of women with endometriosis and released in higher amounts by granulosa-luteal cells from patients with endometriosis. VEGF was accumulated in lower concentrations in the follicular fluid of endometriosis patients. These observations show that the follicular environment is different in cases with endometriosis and suggest that infertility in patients with endometriosis may be related to alterations within the oocyte which, in turn, result in embryos of lower quality, and with a reduced ability to implant. PMID- 10711832 TI - The role of HOX genes in human implantation. AB - HOX genes are transcriptional regulators that play an essential role in determining tissue identity during embryonic development. HOX genes are involved in the development of the Mullerian system and then continue to be expressed in the adult uterus. Two HOX genes have been demonstrated necessary for uterine receptivity in knock-out mice. We have recently elucidated the expression patterns and regulation of HOX genes in the development of the human endometrium during the menstrual cycle. HOX genes are likely key regulators of human implantation. This paper will review the role of HOX genes in the reproductive tract, specifically the evidence that HOX genes are important for human endometrial development and receptivity. PMID- 10711833 TI - Extra-uterine pregnancy following assisted conception treatment. AB - Ectopic pregnancy may be the only life-threatening disease in which prevalence has increased as mortality has declined. The most prominent theory to explain this phenomenon involves increased sensitivity of serum beta-human chorionic gonadotrophin (HCG) immunoassay and improved quality of transvaginal ultrasound, combined with a heightened awareness and increased suspicion of the condition among clinicians which has allowed early detection of ectopic pregnancy. Laparotomy, once the standard treatment of ectopic pregnancy, has been replaced almost entirely by operative laparoscopy. This is associated with a shorter hospital stay, fewer post-operative analgesic requirements, reduced costs and lower risk of adhesion formation. Laparotomy, however, remains necessary in cases with haemodynamic instability and with exceptional locations, e.g. cervical, abdominal and interstitial implantation. In selected cases, non-surgical management has also obtained high success rates. Among medical therapies, the most common is systemic or local administration of methotrexate. The other option is expectant management involving follow-up using serial serum HCG measurements and ultrasound scans. Thus, life-threatening ectopic pregnancy is now evolving into a medical disease, with the possibility of lower-cost treatment, faster recovery and higher subsequent fertility. In this review we assess the risk of extra-uterine implantation after assisted conception treatment, the accuracy of various diagnostic tools and focus on the efficacy, safety and the fertility outcomes of surgical and nonsurgical management of ectopic pregnancy. PMID- 10711834 TI - Human male infertility: chromosome anomalies, meiotic disorders, abnormal spermatozoa and recurrent abortion. AB - Human male infertility is often related to chromosome abnormalities. In chromosomally normal infertile males, the rates of chromosome 21 and sex chromosome disomy in spermatozoa are increased. Higher incidences of trisomy 21 (seldom of paternal origin) and sex chromosome aneuploidy are also found. XXY and XYY patients produce increased numbers of XY, XX and YY spermatozoa, indicating an increased risk of production of XXY, XYY and XXX individuals. Since XXYs can reproduce using intracytoplasmic sperm injection (ICSI), this could explain the slight increase of sex chromosome anomalies in ICSI series. Carriers of structural reorganizations produce unbalanced spermatozoa, and risk having children with duplications and/or deficiencies. In some cases, this risk is considerably lower or higher than average. These patients also show increased diploidy, and a higher risk of producing diandric triploids. Meiotic disorders are frequent in infertile males, and increase with severe oligoasthenozoospemia (OA) and/or high follicle stimulating hormone (FSH) concentrations. These patients produce spermatozoa with autosomal and sex chromosome disomies, and diploid spermatozoa. Their contribution to recurrent abortion depends on the production of trisomies, monosomies and of triploids. The most frequent sperm chromosome anomaly in infertile males is diploidy, originated by either meiotic mutations or by a compromised testicular environment. PMID- 10711835 TI - Oral alkylating agents for breast cancer therapy. AB - Oral cyclophosphamide is well tolerated and effective. Published data support its use as part of adjuvant and metastatic breast cancer treatment regimens. Cyclophosphamide has generally been administered at a higher dose intensity when given orally compared with intravenous infusion. However, there is currently no evidence that oral cyclophosphamide is either more toxic or more or less effective than an equivalent dose of intravenous cyclophosphamide. There is evidence in both the adjuvant and metastatic settings that classical oral cyclophosphamide-methotrexate-fluorouracil (CMF) is more effective than intravenous CMF, possibly because of the greater dose intensity of classical CMF. Prolonged administration of oral cyclophosphamide up to high cumulative doses is associated with an elevated risk of a secondary leukaemia. The rates of chemotherapy-related amenorrhoea with oral cyclophosphamide are directly related to the total dose of cyclophosphamide administered and the patient's age. With the growing availability of other oral cytotoxic agents with demonstrated effectiveness in breast cancer, it is likely that oral cyclophosphamide will be incorporated once again into regimens for both metastatic and adjuvant treatment. PMID- 10711836 TI - Cyclophosphamide and etoposide for non-small cell and small cell lung cancer. AB - Oral chemotherapeutic regimens with limited toxicity are desirable in that quality of life can be maintained and clinic/hospital visits minimised during therapy. We have investigated the use of extended courses of oral cyclophosphamide and oral etoposide for the treatment of non-small cell lung cancer (NSCLC) and small cell lung cancer (SCLC). A 14-day course of oral combination chemotherapy every 28 days resulted in a 12% response rate and a median survival of 6 months (1-year survival, 26%) in stage IV NSCLC. This regimen could not be intensified with carboplatin because of synergistic granulocytopenia. A 14-day course every 28 days resulted in a 40% response rate and a median survival of 7 months in poor-prognosis extensive-disease SCLC. Pharmacodynamic modelling revealed that the granulocyte nadir could be predicted from a single plasma etoposide level drawn on the second day of therapy, potentially allowing dose adjustment during the treatment cycle. Oral chemotherapy remains a promising route for the treatment of lung cancer. PMID- 10711837 TI - Current role of oral etoposide in the management of small cell lung cancer. AB - Etoposide is part of first-line chemotherapy for patients with small cell lung cancer (SCLC) and is one of the few cytotoxic drugs available in an oral formulation. There have been 3 major avenues of investigation of the role of oral etoposide in SCLC: the optimal schedule of administration, its potential as single agent therapy, and its incorporation into combination chemotherapy. Current evidence suggests that, in SCLC, the optimal schedule of administration is over 3 to 5 days. Its use as a single agent for palliation cannot be supported, but it has a continued use within combination chemotherapy for most patients. In addition, it is likely to be incorporated into regimens with new agents that may offer significant advantages over current combinations. PMID- 10711838 TI - The role of oral etoposide in non-small cell lung cancer. AB - Oral etoposide has been tested alone and in combination in a number of tumour types since the late 1980s because of its mild toxicity, high response rates, ease of administration, and comparatively low cost. Encouraging early results with protracted oral etoposide therapy in small cell lung cancer have not been borne out in non-small cell lung cancer (NSCLC), particularly in the advanced disease setting. However, in stage IV NSCLC, oral etoposide does appear to be as compatible with most of the newer agents as it has been with platinum compounds; these combinations continue to be explored, although they have not penetrated into standard usage. In stage III NSCLC, large combined-modality studies are ongoing. Other investigations examining protracted administration in combination with radiation 'sensitisers' are planned. It is possible that by exploiting the 'radiosensitising effect' of prolonged low dose oral etoposide, combined with that of other proven radiosensitisers such as paclitaxel, gemcitabine, and topotecan, we may identify a niche for oral etoposide in the future. PMID- 10711839 TI - Oral etoposide in germ cell tumours. AB - Patients with germ cell tumours who relapse or fail to achieve disease-free status after first-line chemotherapy have a poor prognosis. When administered orally, etoposide produces responses in approximately 25% of patients whose disease is refractory to therapy and is a reasonable choice for palliative treatment in patients who are otherwise incurable. Oral etoposide has also been studied as maintenance therapy in patients who have been treated with salvage chemotherapy or surgery, with results that compare favourably with historical data. We recommend 3 months of maintenance oral etoposide for patients who achieve a complete response to any type of salvage therapy. PMID- 10711840 TI - Oral etoposide in lymphoma. AB - Etoposide is one of the most active agents for the therapy of lymphomas. Oral etoposide has proven to be active in and clearly beneficial for patients with previously treated lymphomas. The optimal dose and schedule of oral etoposide for use in combination chemotherapy are still uncertain, but low daily doses (50 to 100 mg) for 10 to 14 days may be near optimal. Studies in previously untreated patients using combination chemotherapy that includes oral etoposide are needed, since preliminary data suggest that this agent has excellent activity and tolerability when combined or alternated with methotrexate, calcium folinate (calcium leucovorin), cyclophosphamide, vincristine, and prednisone in the elderly and medically unfit patient. Combination therapy approaches may also be helpful in HIV-related lymphomas. Additional studies are warranted. PMID- 10711841 TI - Oral etoposide for the treatment of recurrent ovarian cancer. AB - Oral etoposide has been studied in numerous clinical trials for the treatment of recurrent ovarian cancer. In different studies there has been a varied response rate, and it appears that the activity of this drug is dependent to a large part on the extent of prior therapy. On the basis of data from more than 270 patients in 9 different studies, the overall response rate is 20.4%. However, in the largest study performed by the Gynecologic Oncology Group (GOG), in which 82 previously treated patients received oral etoposide, the response rate was 30.5%. The GOG study categorised patients who received oral etoposide according to their response to initial therapy: patients were deemed either platinum sensitive or platinum resistant. No patients had received more than one prior regimen at the time they were entered into the oral etoposide trial. In 41 platinum-resistant patients, the overall response rate was 26.8%, including a 7.3% clinical complete remission rate. In patients who were platinum sensitive, the overall response rate was 34.1%, with a 14.6% clinical complete remission rate. Toxicity was acceptable, with myelosuppression being the dose-limiting toxicity. This GOG study confirms the activity of oral etoposide as second-line therapy both for platinum-resistant and platinum-sensitive ovarian cancer patients. Additional studies are in progress to determine how oral etoposide can be combined with paclitaxel and a platinum compound for use as initial therapy for previously untreated patients with advanced disease. PMID- 10711842 TI - Extended-schedule oral etoposide in selected neoplasms and overview of administration and scheduling issues. AB - Extended schedules of oral etoposide have been evaluated in many types of advanced cancer. In addition to their use in the common solid tumours, extended schedules have been employed in Kaposi's sarcoma (both AIDS-related and endemic types), medulloblastoma, glioma, and hepatocellular carcinoma. Single agent activity was demonstrated in all of these tumour subtypes. For patients with carcinoma of unknown primary site, we have recently incorporated a 10-day oral etoposide schedule into a combination regimen that also includes paclitaxel and carboplatin. With this regimen we achieved a 47% response rate in a group of 53 evaluable patients, with a median survival of 13.4 months. Patients with adenocarcinoma and poorly differentiated carcinoma of unknown primary site had comparable response rates and survival. According to a large number of clinical trials and pharmacokinetic data, a daily oral etoposide dose of 50 mg/m2 consistently produces serum concentrations >1 mg/L for several hours each day. Lower doses fail to consistently produce this serum concentration, which is considered necessary for optimum tumoricidal activity. Optimal dose duration is 10 to 14 days, particularly when combination regimens are being employed. Oral etoposide has an established role as a single agent in patients with low grade non-Hodgkin's lymphoma, Kaposi's sarcoma, and testicular cancer (if residual carcinoma is resected after first-line treatment). The optimal use of extended schedule etoposide in combination regimens is not defined but is being evaluated in a number of etoposide-sensitive malignancies. PMID- 10711844 TI - The role of tegafur/uracil in pulmonary malignancy. AB - Tegafur/uracil (UFT) is an oral fluorinated pyrimidine composed of a combination of uracil and tegafur in a 4:1 molar ratio. When given on a continuous basis, it yields a fluorouracil area under the concentration-time curve identical to that achieved with isomolar concentrations of fluorouracil administered by continuous infusion. Its 8.7% response rate in advanced non-small cell lung cancer (NSCLC) is modest, but it has demonstrated synergy with cisplatin in human solid tumour xenografts. In combination with cisplatin in patients with advanced NSCLC, response rates of 29 to 43% were observed with acceptable toxicity. Researchers in Japan have demonstrated the utility of this agent as adjuvant therapy after surgical resection, either alone or in combination with cisplatin and vindesine versus observation; long term survival rates exceed 60%, compared with 49% for the control group. Efforts in the US will assess the role of this agent as non cross-resistant consolidation therapy after a plateau in response has been obtained with platinum-based therapy in advanced disease. They will also recapitulate the Japanese studies, assessing their utility as adjuvant therapy in NSCLC patients who have undergone resection. Finally, ongoing phase I studies are combining this agent with paclitaxel and other systemic agents. PMID- 10711843 TI - New oral chemotherapeutic agents for lung cancer. AB - Anticancer treatment has recently shifted to include a broad range of antineoplastic therapies. Old agents are continuously being re-evaluated, and new mechanisms of treatment are rapidly being explored and developed. At the same time, the patient's perceived quality of life, adverse effects of therapy, time demands, and healthcare costs have become paramount in the treatment process. Lung cancer is the most common cause of cancer death in the USA, and because many of the patients are older or debilitated, these issues become all the more important. The oral administration of anticancer therapy offers both quality-of life and healthcare cost advantages. Oral forms of 3 new cytotoxic agents and 2 novel oral therapies are discussed. Vinorelbine, a vinca alkaloid, has well documented activity in non-small cell lung cancer. Myelosuppression is dose limiting; neurotoxicity is rare. Satraplatin (JM-216), an oral platinum derivative, shows activity in lung cancer with a favourable adverse effect profile, with no neurotoxicity or nephrotoxicity. The oral topoisomerase I inhibitor topotecan may be ideal for obtaining long term low plasma drug concentrations, which appears to maximise efficacy. LGD-1069 is a retinoid X receptor agonist that modulates cell proliferation, and BAY-129566, a matrix metalloproteinase inhibitor, appears to interrupt both the processes of angiogenesis and metastasis. LGD-1069 and BAY-129566 are nontraditional anticancer agents which may be used in conjunction with chemotherapy, other modalities, or in prevention. These 5 agents will be discussed with particular reference to recent developments in the treatment of lung cancer. PMID- 10711845 TI - Tegafur/uracil + calcium folinate in colorectal cancer: double modulation of fluorouracil. AB - The oral chemotherapeutic agent tegafur/uracil (UFT) is the first of a new class of anticancer drugs called dihydropyrimidine dehydrogenase inhibitory fluoropyrimidines. Tegafur/uracil combines uracil with the fluorouracil prodrug tegafur in a 4:1 molar ratio. Uracil competitively inhibits the degradation of fluorouracil, which results in the concentration of fluorouracil remaining at sustained levels in both plasma and tumour. Tegafur/uracil has been commercially available in Japan since 1983 and examined extensively in various tumours. Trials conducted in the US have focused on the combination of tegafur/uracil plus calcium folinate (calcium leucovorin) [ORZEL]. Several phase I and II trials have evaluated the maximum tolerated dose, pharmacokinetics, efficacy, and safety of this combination in the treatment of colorectal cancer. Results have shown that tegafur/uracil at 300 mg/m2/day in divided doses given every 8 hours for 28 days provides prolonged exposure to fluorouracil. Furthermore, tegafur/uracil + calcium folinate is well tolerated, with dose-limiting toxicity manifesting as diarrhoea. Compared with intravenous fluorouracil plus folinic acid (leucovorin) regimens, tegafur/uracil + calcium folinate has similar efficacy with less toxicity and is more convenient because it is an oral regimen. Early studies have also shown potential cost savings because of fewer complications. PMID- 10711846 TI - Recent developments in oral chemotherapy options for gastric carcinoma. AB - The incidence of carcinoma of the stomach is low in the United States, Canada, and Australia but is a significant health problem in Asia, South America, Eastern Europe, and countries of the previous Soviet Union. For patients with advanced disease, chemotherapy remains palliative. With the increasing emphasis on patients' quality of life, convenience, and cost containment, oral chemotherapy has come into increasing focus. We review oral chemotherapy agents for use in patients with advanced gastric carcinoma. Etoposide, given intravenously, has modest activity in gastric carcinoma. We studied oral etoposide, which was administered to 28 patients at the starting dose of 50 mg/m2/day for 21 days followed by a 7-day rest period. Five patients achieved a partial response and 4 patients achieved a minor response. The drug was well tolerated. Common toxicities included myelosuppression, alopecia, and nausea. Oral etoposide thus shows evidence of modest activity against gastric carcinoma. In Japan, considerable advances have been made in the oral chemotherapy of gastric carcinoma. The second generation fluorouracil prodrug tegafur/uracil (UFT) has been extensively evaluated in Japan, Korea, and Spain. Data predominantly from Japan indicate that tegafur/uracil has a response rate of approximately 20% in treatment naive patients with advanced gastric carcinoma. When combined with other active agents, tegafur/uracil has a response rate of more than 30% in these patients. The available data also suggest that tegafur/uracil is well tolerated and that patient acceptance is high. In conclusion, future clinical research is likely to focus on the development of convenient outpatient regimens with efficacy equal to that of intravenous regimens. PMID- 10711847 TI - Oral chemotherapy in head and neck cancer. AB - Chemotherapy plays an important role in the palliative treatment of head and neck cancer and in the neoadjuvant setting for larynx preservation. Together with concomitant radiotherapy, chemotherapy is also important for the curative and palliative therapy of unresectable head and neck cancer. Although issues relating to anatomical and pharmacological constraints exist, new orally administered drugs, as well as oral substitutes for the currently utilised intravenous drugs, would be extremely desirable in each of these situations. Of the oral fluorinated pyrimidines, tegafur/uracil (UFT) alone produced a complete response rate of 19%, and combination therapy of tegafur/uracil or tegafur with cisplatin or carboplatin has produced response rates comparable to those seen with intravenous fluorouracil (5-FU) plus cisplatin or carboplatin. An initial dose-finding study of 5-FU plus eniluracil indicates that further studies are warranted. The ribonuclease reductase inhibitor hydroxycarbamide (hydroxyurea) has been extensively studied in combination with 5-FU and radiotherapy (the FHX regimen) in patients with head and neck cancer, with high rates of local control. Improvement in locoregional and distant control rates may occur when FHX is combined with additional systemically active agents (cisplatin then paclitaxel) and hyperfractionated radiotherapy is used. Good candidate drugs for head and neck cancer include BMS-182751, an oral platinum complex, and capecitabine and S 1, other oral fluoropyrimidines. In addition, methotrexate and cyclophosphamide both have some activity in head and neck cancer and deserve further investigation. PMID- 10711848 TI - Oral combination chemotherapy in the management of AIDS-related lymphoproliferative malignancies. AB - An oral combination chemotherapy regimen initially developed for AIDS-related non Hodgkin's lymphoma includes lomustine (CCNU), etoposide, cyclophosphamide, and procarbazine. This regimen takes advantage of oral administration, the in vitro synergy of these drugs and their first-line efficacy in lymphoma, and the ability of lomustine and procarbazine to cross the blood-brain barrier. This regimen was used to treat 38 patients with AIDS-related non-Hodgkin's lymphoma. The overall objective response rate was 66% (34% complete response rate) with a 5% CNS relapse rate, and a median survival duration of 7.0 months. One-third of the patients survived for 1 year, 11% for 2 years, and half of the patients survived free from progression of their lymphoma. On the basis of these results, this oral regimen was modified and administered to 5 patients with AIDS-related primary CNS lymphoma as part of a sequential combined-modality chemotherapy and radiation regimen. Rapid progression of CNS disease was observed in this group of patients, with a median survival duration of 1.0 month. The identical regimen was administered to 7 patients with AIDS-related Hodgkin's disease: we observed a 71% partial remission rate and a median survival duration of 7.0 months. Myelosuppression remains the most significant clinical toxicity. Our results with this oral regimen appear comparable to those of standard intravenous combination chemotherapy regimens in patients with AIDS-related non-Hodgkin's lymphoma. PMID- 10711849 TI - Oral chemotherapy agents in the treatment of leukaemia. AB - Oral chemotherapy agents have been an important component of the treatment of leukaemia for many years. Obstacles such as poor or erratic bioavailability and noncompliance have often limited the utility of oral agents in the treatment of leukaemia. However, recent evaluations of new or existing oral agents have expanded the clinician's options and understanding of the use of these drugs in the treatment of leukaemia. One major advance is the use of tretinoin (all-trans retinoic acid) in the treatment of acute promyelocytic leukaemia (APL). Tretinoin, an oral vitamin A derivative that reverses abnormal differentiation in APL is now an essential component of first-line therapy for APL, replacing standard intravenous chemotherapy induction regimens. Other advances include an increased understanding of the pharmacokinetic and pharmacodynamic profile of oral chemotherapy agents such as etoposide and high dose busulfan, allowing for modifications or individualisation of administration regimens to enhance efficacy or minimise toxicity. Evaluations of noncompliance with oral agents in the treatment of leukaemia have also provided the clinician with important information on how this obstacle to oral therapy may be overcome or minimised. PMID- 10711850 TI - Improving fluorouracil chemotherapy with novel orally administered fluoropyrimidines. AB - During the 40 years since the initial synthesis of fluorouracil, there have been many attempts to improve fluoropyrimidine chemotherapy. These have included the utilisation of different schedules of fluorouracil administration, modulation of the metabolism of fluorouracil with other drugs to increase its therapeutic benefit, and the synthesis of prodrugs of fluorouracil that are potentially more effective and less toxic. Of particular interest at present is the clinical evaluation of several new fluoropyrimidine drugs that can be orally administered. These include capecitabine, tegafur/uracil (UFT), eniluracil (GW-776C85; 5 ethynyluracil), S-1, and emitefur (BOF-A2). The pharmacological principles that have influenced the development of these new drugs are initially presented. This is followed by a review of our current knowledge of the clinical pharmacology of each of these new agents, focusing on antitumour activity and toxicity from studies conducted in the US. Studies of capecitabine, tegafur/uracil, and early studies with eniluracil indicate that these drugs have at least similar activity to protracted fluorouracil infusion but with additional quality-of-life and economic benefits. PMID- 10711851 TI - Oral chemotherapy in the treatment of hormone-refractory prostate cancer. AB - Oral chemotherapy has become a major component of the treatment of advanced prostate cancer. The recognition that prostate cancer grows very slowly and must be treated continuously with active agents has led to the development of several therapeutic regimens. These regimens employ oral agents such as estramustine, cyclophosphamide, and etoposide, as they can be taken on a daily basis at home by the patients. These regimens have demonstrated activity in patients with hormone refractory prostate cancer; declines in both prostate specific antigen and soft tissue lesions have been demonstrated. PMID- 10711852 TI - Oral cancer chemotherapy in paediatric patients: obstacles and potential for development and utilisation. AB - Although most cancer chemotherapy in children is administered parenterally, oral formulations are becoming increasingly available for use in patients as young as infants. Incentives for this approach include safety, flexibility, reduced financial cost, improved quality of life, and the potential for improved efficacy. The ability to deliver chemotherapy at home and apply schedules of administration that increase the agent's efficacy because patients do not require hospitalisation or visits to the clinic renders oral chemotherapy particularly attractive. Obstacles to oral chemotherapy in paediatric patients include restrictions in dose size and schedule, uncertain or low bioavailability, adverse effects of malabsorption, and adherence (noncompliance and refusal to take oral chemotherapy). Techniques to overcome most of these limitations are available or can be developed, and lack of an oral formulation can be solved in many instances by the pharmaceutical industry. Future developments in oral chemotherapy should not be limited to adult patient applications. PMID- 10711853 TI - Rationale for once-daily therapy in asthma: compliance issues. AB - Compliance in asthma is known to be poor. Once-daily treatment provides an additional therapeutic option for the clinician whose treatment aims include maximising treatment satisfaction and compliance. Once-daily treatment is preferred by some but not all patients and may lead to greater compliance in patients who are concerned about the effect of corticosteroids on their health. When switching to once-daily treatment, patients should be given a choice as to the time of treatment to minimise the impact of forgetting a dose. PMID- 10711854 TI - Pharmacological factors that influence the choice of inhaled corticosteroids. AB - Local therapeutic effect relative to the risk of adverse effects of inhaled drugs, i.e. airway selectivity, is determined by the efficiency of the delivery system and the physicochemical and pharmacokinetic properties of the drug molecule. For the inhaled corticosteroid formulations, many of the pharmacokinetic prerequisites for airway selectivity have been fulfilled, but there are still differences that may influence the choice of treatment regimens. This choice should be based on disease severity, age, inhalation technique, preference and expected compliance, together with a knowledge of individual features of different corticosteroid formulations. Simple to use, hand-held pressurised or breath-actuated inhalers have favourable lung deposition properties and are appropriate for most patients. For small children or severely ill patients, nebulised treatment or spacers may be advocated. A corticosteroid formulation with a high intrinsic activity and long duration of action allows for once-daily administration in some patient groups. These properties may also partly compensate for noncompliance when more frequent administration schemes are used. The risk of adverse effects is reduced if systemic exposure is held to a minimum by rapid elimination and low tissue distribution. PMID- 10711855 TI - Inhaled corticosteroid therapy in newly detected mild asthma. AB - Inhaled corticosteroids are the most effective medications currently available to treat symptomatic asthma, and are free of clinically relevant unwanted effects, when used at the doses needed to provide optimal control in most patients with asthma. Inhaled corticosteroids also improve the physiological abnormalities of variable airflow obstruction and airway hyperresponsiveness that characterise asthma. Inhaled corticosteroids are also cost-beneficial when compared with other treatments, even in patients with milder asthma who are treated in primary care. For these reasons, inhaled corticosteroids are now being considered as first-line therapy for patients with regular daily asthma symptoms. Inhaled corticosteroids are, however, often not utilised until other treatments have been demonstrated not to provide optimal asthma control. Available evidence from both children and adults with asthma suggests that the benefits achieved from inhaled corticosteroids are reduced when their introduction as therapy is delayed for several years after persistent symptoms develop. For this reason, corticosteroids should be started soon after a diagnosis is made and persistent symptoms develop. It is not yet known, however, whether inhaled corticosteroids should be used in asthmatic patients who have very mild and infrequent symptoms and who have normal airway calibre most of the time. The current consensus statements do not recommend regular treatment in such patients. Airway biopsies from these asthmatic patients do show evidence of airway inflammation and structural changes; however, we do not yet know whether they lose lung function more rapidly than individuals without asthma, or whether the morbidity associated with very mild asthma warrants the use of regular treatment. This issue is being addressed in a large, multinational, placebo-controlled trial. The results of this study (available in 3 more years) will resolve this persisting question about inhaled corticosteroid use in mild asthma. PMID- 10711856 TI - Once-daily inhaled corticosteroids in mild to moderate asthma: improving acceptance of treatment. AB - Despite the established efficacy of inhaled corticosteroids in improving lung function in asthma, there has not been a corresponding improvement in morbidity and mortality associated with the disease, which, in part, may result from non compliance with the prescribed regimen. The reasons for this are many and varied, but an important measure in improving the level of compliance in asthma patients is simplification of the treatment regimen, which may be achieved by reducing the dose frequency and improving the ease of administration. In clinical trials designed to determine whether a reduction in dose frequency to once daily is associated with similar efficacy to that with more frequent administration, a number of studies have shown that once-daily administration of inhaled corticosteroids in both adults and children is as effective in controlling asthma as twice-daily administration of the same dosage, both when given as initial therapy in corticosteroid-naive patients and in patients already receiving an inhaled corticosteroid. The drug for which most evidence to support a dosage change from twice-daily to once-daily therapy currently exists is budesonide, though limited evidence with other inhaled corticosteroids such as beclomethasone dipropionate, fluticasone propionate and flunisolide also supports once-daily use. Despite the larger single dosage with once-daily budesonide therapy, there has been no evidence in clinical trials of a greater incidence of local adverse effects such as hoarseness, throat irritation or oropharyngeal candidosis, and no evidence of adrenal suppression or growth retardation. Since compliance is an important factor that can affect the success or failure of asthma therapy, a reduction in the frequency of administration to once daily offers the potential advantage of improved compliance with treatment and hence better control of asthma. In the short term clinical trials conducted to date, patient preferences have favoured the once-daily regimen over twice-daily administration. When combined with other (e.g. educational) measures to improve patient compliance, a switch from twice-daily (or more frequent) administration to once-daily inhaled corticosteroid therapy seems likely to be beneficial in improving the long term outcome of treatment. PMID- 10711857 TI - Once-daily inhaled corticosteroids in children with asthma: dry powder inhalers. AB - The cornerstone of pharmacological management of asthma in childhood is inhaled corticosteroids. These drugs are intended for long term treatment and, consequently, compliance is a major issue. Once-daily administration of maintenance medication would simplify treatment and it is likely that it would lead to better compliance. Moreover, the excellent safety profile of inhaled corticosteroid treatment tailored to disease severity may, theoretically, be further improved with once-daily administration. Studies comparing inhaled corticosteroids given once or twice daily to patients with asthma indicate that unstable asthma is best treated with at least 2 daily doses. On the other hand, it has been demonstrated that, if the asthma is stabilised, most children can be treated with inhaled corticosteroids once daily without loss of efficacy. Thus, the data suggest that newly diagnosed asthma, or asthma after deterioration, should first be reliably controlled with inhaled corticosteroids divided into at least 2 daily doses. Once-daily maintenance treatment should then be tried with the aim of improving compliance and quality of life. A dry powder inhalation device is probably the best choice for children from the age of 5 years. PMID- 10711858 TI - Once-daily inhaled corticosteroids in children with asthma: nebulisation. AB - Current guidelines on the management of childhood asthma have emphasised the important preventive role of inhaled corticosteroids, which should be used at the lowest possible doses that are compatible with good disease control. However, some children do not respond to inhaled corticosteroids, the most common reasons for which are inability to use conventional hand-held inhalers (plus spacers and face masks) effectively or lack of cooperation with them, particularly among infants and young children. In these patients, nebulisers have proved effective in administering corticosteroids, and this form of delivery is often preferred by both the children and their parents, despite their longer administration times (commonly around 10 minutes). Compliance with these devices may therefore be better than with a conventional pressurised metered-dose inhaler plus spacer and face mask. Recent studies with nebulised budesonide have demonstrated that once daily administration is as effective in maintaining control of asthma symptoms in children as the usual twice-daily administration. In children with moderately severe persistent asthma, the improvement provided by once-daily nebulised doses of 1.0 mg budesonide has been found to be equivalent to that with twice-daily doses of 0.25 or 0.5 mg, indicating that once-daily therapy is an effective option that can be considered in many patients. In view of the time-consuming nature of nebuliser administration, reduction of the frequency of corticosteroid administration from twice to once daily may be useful in simplifying the treatment programme and improving compliance with it. This may be beneficial in reducing under-utilisation of inhaled corticosteroids in children with asthma and improving long term control of the disease. PMID- 10711859 TI - Clinical implications of antibiotic resistance for management of acute otitis media. PMID- 10711860 TI - Persistence of Staphylococcus aureus on mucosal membranes: superantigens and internalization by host cells. PMID- 10711861 TI - Endocrine and lipid effects of oral L-arginine treatment in healthy postmenopausal women. AB - As a substrate for nitric oxide synthesis, L-arginine may give the same protection as estrogen, but its other biologic effects may adversely affect atherogenesis. Therefore, possible endocrine and lipid effects of L-arginine were investigated in a double-blind, placebo-controlled, single crossover study. After randomization, oral L-arginine (9 g) or placebo was given daily for 1 month, with crossover to the alternate therapy after a 1-month washout period, to 10 postmenopausal women receiving no estrogen. Compared with placebo, L-arginine increased growth hormone (1.5+/-1.8 mg/L vs. 0.6+/-0.6 mg/L, P = .04) but had no effect on insulin and catecholamines. Total cholesterol, triglyceride, apolipoprotein E, and low-, very-low-, and high-density lipoprotein cholesterol levels were also unaffected. Lipoprotein(a) measured by an immunoturbidimetric method was increased by L-arginine in 9 of 10 women relative to placebo (0.46+/ 0.35 g/L vs. 0.38+/-0.30 g/L, P = .053), and the changes in lipoprotein(a) levels significantly correlated with the relative increase in growth hormone (r = 0.85, P = .03). However, lipoprotein(a) measured by an enzyme-linked immunosorbent assay failed to demonstrate significant changes. Lack of an increase by L arginine in lipoprotein(a) with a verifiable apolipoprotein(a) isoform independent method, despite an increase in growth hormone, questions the validity of previous observations for growth hormone-induced increases in lipoprotein(a). The observed lack of effect on major endocrine hormones and lipid profile support the safety of oral L-arginine administration. PMID- 10711862 TI - Filtration of activated granulocytes during cardiopulmonary bypass surgery: a morphologic and immunologic study to characterize the trapped leukocytes. AB - Cardiopulmonary bypass surgery induces an inflammatory reaction among others by activation of granulocytes. Leukocyte filtration has been shown to reduce the postoperative morbidity mediated by activated granulocytes. However, little is known about the mechanism of filter-leukocyte interaction. This study examines whether a leukocyte filter removes activated granulocytes or a general leukocyte population. Eleven patients undergoing cardiopulmonary bypass surgery were included in this study. Leukocyte filtration was achieved before the reperfusion phase with a Pall non-woven polyester filter located at the venous side of the heart-lung machine. After filtration, the trapped granulocytes inside the filter were examined morphologically with light and scanning electron microscopy and immunologically by CD45RO antigen binding to the filter material. Furthermore, leukocyte release markers were measured to determine whether cells were activated during filtration. Microscopic evaluation revealed 84% granulocytes and 14% lymphocytes trapped in the filter, compared with 78% granulocytes and 22% lymphocytes in the blood before filtration. Granulocytes were trapped significantly more in the first blood contact layer of the filter material than in the middle layer and last layer, whereas lymphocytes trapped slightly more in the middle layer. The near maximum level of CD45RO expression was measured on granulocytes trapped inside the filter material, whereas CD2 and CD19 measured on lymphocytes were bound to a minor extent. Beta-glucuronidase concentration did not increase after filtration, suggesting the absence of activation of granulocytes by filtration. A leukocyte filter made of non-woven polyester material removes the activated granulocytes rather than leukocytes at random. This implies that this particular type of leukocyte removal filter is suitable for use in cardiopulmonary bypass patients whose granulocytes in the circulation are activated. Furthermore, measurement of activated granulocytes instead of total leukocyte count is likely preferable for functional assessment of leukocyte removal devices. PMID- 10711863 TI - Effect of glycoprotein IIb/IIIa inhibitors on CD62p expression, platelet aggregates, and microparticles in vitro. AB - Flow cytometry can detect platelet activation (CD62p), aggregate formation, microparticle formation, and glycoprotein IIb/IIIa (GP IIb/IIIa) receptor occupancy in one sample at the level of single particles. We studied the effect of GP IIb/IIIa inhibitors on platelet activation with flow cytometry in vitro. Citrated whole blood was incubated with increasing concentrations of three different GP IIb/IIIa inhibitors (c7E3, DMP728, XJ757), then thrombin or adenosine diphosphate (ADP) was added, and after 1 minute the sample was fixed. Samples with thrombin but without c7E3 had a decrease in platelet count, from a mean of 260,000 platelets/microl to 56,000 platelets/microL, and aggregates increased. Samples with concentrations of c7E3 that resulted in 80% or more receptor blockade had no decrease in platelet count, and no aggregates were formed, but the number of CD62p-positive single platelets increased from 1200 to 7400 platelets/microL. The two other inhibitors (DMP 725, XJ757) or ADP instead of thrombin gave similar results. Microparticle formation did not change with platelet activation in the presence of a GP IIb/IIIa inhibitor. With small inhibitor doses resulting in <80% receptor blockade, the number of aggregates did not change or was even higher than that in samples without inhibitor. GP IIb/IIIa inhibitors do prevent aggregate formation but they do not prevent activation of platelets. With GP IIb/IIIa inhibition, more activated single platelets remain in the blood. One may expect an increasing number of circulating, activated platelets with the use of GP IIb/IIIa inhibitors. PMID- 10711864 TI - Relationship between red cell mean corpuscular volume and 6-thioguanine nucleotides in patients treated with azathioprine. AB - Azathioprine (AZA) is characterized by high interindividual differences in bioavailability and metabolization. The aim of the present study was to analyze, in patients treated with AZA for various immune system disorders, whether the variation in red blood cell mean corpuscular volume (deltaMCV) could be used as an indirect estimation of the level of the active immune modifier metabolite 6 thioguanine nucleotides (6-TGN). In 43 consecutive patients treated with a stable dose of AZA for at least 6 months who were not initially anemic, the erythrocyte 6-TGN levels with routine hematologic parameters were determined two to four times at 1-month intervals. In most patients MCV significantly increased after 3 months of therapy and stabilized after 6 months. The correlation between the daily dose of AZA and the 6-TGN level was mild (r = 0.51; P<.001). A weak correlation was also found between the dose of AZA and the deltaMCV after at least 6 months of therapy (r = 0.36; P<.05). The correlation between deltaMCV and 6-TGN level, however, was much better (r = 0.74; P<.001). The lack of a significant increase in MCV after 3 to 4 months of AZA therapy reflects low 6-TGN levels, sometimes a result of undertreatment. A determination of the 6-TGN level during the first months after AZA therapy is begun will allow more accurate adaptation of the effective dose. We observed that deltaMCV could be used as an indicator of 6-TGN levels after 6 months of AZA treatment. An increase in MCV of at least 6 fL is expected to reflect a 6-TGN level of about 175 pmol/8x10(8) red blood cells (probably being within a therapeutic value). PMID- 10711865 TI - Hemoglobin stimulates the release of proinflammatory cytokines from leukocytes in whole blood. AB - Isolated mononuclear leukocytes, when incubated with purified hemoglobin Ao (HbAo), release the proinflammatory cytokines interleukin-8 (IL-8) and tumor necrosis factor-alpha (TNF-alpha). In this study we examined whether leukocytes in whole blood, when incubated with HbAo, release IL-8, TNF-alpha, and IL-6. Leukocytes in whole blood incubated with HbAo for 4 hours at 37 degrees C, 5% CO2, and 95% humidity released 187, 1313, and 50 pg/mL of IL-6, IL-8, and TNF alpha, respectively, as compared with 6, 192, and 2 pg/mL released by leukocytes in blood incubated with human serum albumin (HSA). Furthermore, plasma from blood incubated with HbAo exhibited chemotactic activity and stimulated human umbilical vein endothelial cells to become adherent to neutrophils. These activities were 3.3 and 2.6 times those measured in plasma from blood incubated with HSA. Hydrocortisone (0.05 micromol/L to 50 micromol/L) inhibited cytokine release in a dose-dependent manner with ED50 values of 0.23 micromol/L, 0.19 micromol/L, and 0.10 micromol/L for IL-6, IL-8, and TNF-alpha, respectively. The release of proinflammatory cytokines in whole blood after exposure to hemoglobin solutions is consistent with the possibility that an inflammatory reaction could develop on infusion of hemoglobin, whereas inhibition of cytokine release by hydrocortisone suggests that the inclusion of anti-inflammatory compounds in hemoglobin solutions may prevent undesirable effects caused by inflammation after infusion. PMID- 10711866 TI - Dexamethasone lowers circulating E-selectin and ICAM-1 in healthy men. AB - Plasma levels of circulating adhesion molecules (AMs) are increased in a number of inflammatory and cardiovascular disorders. Yet the mechanisms regulating the physiologic levels of soluble AMs are largely unknown. It has recently been postulated that glucocorticoids may exert their anti-inflammatory actions partially through the inhibition of cytokine-stimulated expression of E-selectin and intercellular adhesion molecule (ICAM-1). However, it remains controversial whether glucocorticoids affect the basal expression of AMs on resting cells. We have thus evaluated the effects of glucocorticoids by infusing therapeutic doses of dexamethasone (0.04 mg/kg and 1.0 mg/kg twice a day for 2 days) or placebo on plasma levels of circulating E-selectin (cE-selectin), soluble thrombomodulin (sTM), circulating ICAM-1 (cICAM-1), and circulating vascular cell adhesion molecule (cVCAM-1) in 9 healthy men. Plasma was obtained before infusion at 24 and 48 hours. Compared with baseline, levels of cE-selectin decreased by 16% and 22% with the lower and the higher doses, respectively, at 48 hours (P = .007), whereas sTM was unchanged. Both doses of dexamethasone reduced cICAM-1 by about 15% at 48 hours (P = .007), but there were no changes in cVCAM. Dexamethasone time-dependently decreases plasma levels of cE-selectin and cICAM-1 in healthy men. This demonstrates that a glucocorticoid-sensitive mechanism specifically down-regulates normal plasma levels of cE-selectin and cICAM-1 in healthy subjects, which could thus reflect minor baseline inflammation. PMID- 10711867 TI - Effects of dietary polyunsaturated fatty acid supplementation in early renal insufficiency in dogs. AB - Dietary supplementation with polyunsaturated fatty acids (PUFAs) alters the course of experimental kidney disease in dogs. In particular, supplementation with omega-6 PUFAs hastens the decline of kidney function, and omega-3 PUFAs are renoprotective. We investigated the early stages of renal insufficiency to determine whether PUFA supplementation altered the magnitude of hypercholesterolemia or glomerular hemodynamics. Two months after 11/12 nephrectomy, dogs were randomly divided into three groups of 6 animals each. Each group of dogs was then fed a low-fat basal diet supplemented with one of three sources of lipid to achieve a final concentration of 15% added fat. Fat sources were rich in omega-3 PUFAs (menhaden fish oil, group FO), omega-6 PUFAs (safflower oil, group SO), or saturated fatty acids (beef tallow, group C). Early in renal insufficiency, before significant kidney damage, group FO had a lower (P<.05) serum cholesterol concentration and tended to have a lower urinary prostaglandin E2 (PGE2) and thromboxane A2 (TxA2) excretion than group C. In contrast, group SO had a higher mean glomerular capillary pressure (P<.05) and more glomerular enlargement (P<.05) and tended to have higher eicosanoid excretion rates than group C. These differences in lipid metabolism, glomerular hypertension and hypertrophy, and urinary eicosanoid metabolism could explain, in part, the beneficial effects of omega-3 PUFAs and the detrimental effects of omega-6 PUFAs when administered on a long-term basis in this model of renal insufficiency. PMID- 10711869 TI - Longitudinal changes in optic disc topography of adult patients after trabeculectomy. PMID- 10711868 TI - Further observations and characterization of monoclonal antibodies reacting with B cell alloantigens associated with rheumatic fever and rheumatic heart disease. AB - Elevated levels of B lymphocytes with a unique surface alloantigen have been reported to be characteristic of patients with acute rheumatic fever or rheumatic heart disease. Mouse monoclonal antibodies (mAbs) to this alloantigen have been proposed as being useful in identifying individuals at risk for the development of these sequelae of group A streptococcal infection. However, previous studies have suggested that the discriminating ability of the mAbs was highest when the mAbs were made by using lymphocytes from the same ethnic population. To confirm and extend this observation, additional mouse mAbs were developed and their properties defined. These three mAbs-PG-12A, PG-13A, and PG-20A-reacted with B cells from more than 90% of North Indian patients with acute rheumatic fever or rheumatic heart disease. Each of these three new mAbs identified the highest levels of reactive B cells in patients with active acute rheumatic fever. Lower levels of positive reacting lymphocytes were found in individuals with quiescent chronic rheumatic heart disease, and markedly reduced percentages of reactive cells were observed in normal control subjects. The proportion of reactive lymphocytes in individual patients varied according to which of the three was tested, suggesting the possibility of a spectrum of "rheumatic" epitopes in susceptible individuals. The data further suggested that enhanced discriminatory ability for identifying "at-risk" susceptible patients could be obtained by testing with a combination of mAbs. If reduction in the incidence of acute rheumatic fever can be facilitated by early identification of susceptible individuals, accurate and sensitive detection of a marker antigen would result in more cost-effective public health measures. Additional population studies are required to more precisely define and confirm these detection techniques. PMID- 10711870 TI - Anterior ischemic optic neuropathy: trouble waiting to happen. PMID- 10711871 TI - MRI for metallic foreign bodies? PMID- 10711872 TI - Limbal stem cell allografting from related live donors for corneal surface reconstruction. PMID- 10711873 TI - Harold Ridley knighted. PMID- 10711874 TI - Eleven commandments for teaching ophthalmic surgery. PMID- 10711875 TI - Effect of trabecular aspiration on intraocular pressure in pigment dispersion syndrome and pigmentary glaucoma. AB - OBJECTIVE: Recently, we described a new form of nonfiltering glaucoma surgery trabecular aspiration-designed to increase trabecular outflow in pseudoexfoliation glaucoma. This study was carried out to investigate whether trabecular aspiration is equally safe and effective in the treatment of pigment dispersion syndrome (PDS) and pigmentary glaucoma (PG). STUDY DESIGN: Prospective, nonrandomized comparative trial with historical control. PARTICIPANTS: Twenty eyes of 20 patients with medically uncontrolled intraocular pressure (IOP) caused by PDS or PG were treated by trabecular aspiration. INTERVENTION: Trabecular pigment particles were cleared with a pressure of 100 to 200 mmHg using a specially designed aspiration probe. MAIN OUTCOME MEASURES: The IOP and number of medications before and after surgery were measured. Intraoperative and postoperative complications were analyzed. Surgical success was defined as IOP < or = 21 mmHg with no more than one topical medication. Results were compared with those previously reported with similar treatment of pseudoexfoliative glaucoma. RESULTS: Mean pretreatment IOP averaged 27.0 (standard deviation [SD], 3.3) mmHg and was significantly reduced to 23.7 (SD, 3.9) mmHg at last follow-up (20.1 +/- 8.6 months). However, the cumulative life table success rates were only 42% and 15% at 3 and 12 months, respectively. Considering both groups separately, the success rates in the PDS group were 63% and 18% at 3 and 12 months compared with a success rate of 12% in the PG group as early as 1 month after surgery. CONCLUSIONS: Eyes with PDS responded better to trabecular aspiration than do those with PG, indicating that PDS and PG are two successive stages of the same disease process. Altogether, trabecular aspiration failed to achieve long-term pressure control in either of the two groups. PMID- 10711876 TI - Trabeculectomy with mitomycin-C in the treatment of pediatric glaucomas. AB - PURPOSE: To evaluate the effectiveness and safety of trabeculectomy with mitomycin-C (MMC) in the management of childhood glaucomas. DESIGN: Retrospective, noncomparative case series. PARTICIPANTS: All patients less than 18 years of age who underwent trabeculectomy with MMC from June 1991 through October 1997 were included. METHODS: The medical records of 29 consecutive patients (29 eyes) were reviewed. Data collected during routine patient follow-up were analyzed. Surgical outcome was evaluated using Kaplan-Meier life-table analysis. MAIN OUTCOME MEASURES: Clinical outcome assessment included intraocular pressure (IOP) control, visual acuity, and identification of complications. Successful IOP control was defined as 5 mmHg < or = final IOP < or = 21 mmHg, with or without antiglaucoma medications and without further glaucoma surgery or loss of light perception. Outcomes for the group of patients with primary infantile glaucoma were compared with those for the group with secondary glaucomas. RESULTS: Mean patient age was 6.4 +/- 4.4 years (range, 0.2-15.3 years). A variety of primary and secondary glaucomas were represented. Mitomycin C (0.5 mg/ml) on a surgical sponge was applied to the episcleral surface for an average of 3.8 +/- 1.0 minutes (range, 1.5-5.0 minutes). Mean follow-up time for patients categorized as successes was 25.1 +/- 16.0 months (range, 5.5-59.7 months). The 12-, 24-, and 36-month life-table success rates for IOP control were 82%, 59%, and 59% respectively. There was no difference between the primary infantile glaucoma group and the secondary glaucoma group with respect to length of follow-up, rate of successful IOP control, and incidence of complications. Five patients (17%) experienced late bleb-related infection (BRI) at an average of 27.9 +/- 18.1 months (range, 5.4-55.5 months) after surgery. Other complications included hyphema, retinal detachment, late-onset bleb leak, flat anterior chamber, chronic hypotony, decompression retinopathy, suture abscess, and phthisis. CONCLUSIONS: Trabeculectomy with MMC may be useful in the management of childhood glaucomas in which goniotomy, trabeculotomy, or both have failed. However, the high incidence of BRI in this series over an extended follow up interval dictates caution in using MMC as an adjunct in pediatric trabeculectomy. PMID- 10711878 TI - Quantification of aqueous melanin granules, intraocular pressure and glaucomatous damage in primary pigment dispersion syndrome. AB - OBJECTIVE: Aqueous melanin granules may be accurately quantified with the laser flare-cell meter and have been demonstrated to be increased in primary pigment dispersion syndrome (PDS). It was the aim of this study to correlate intraocular pressure, glaucomatous damage of the optic nerve head, and visual field defects with the number of aqueous melanin granules in PDS. DESIGN: Cross-sectional study. PARTICIPANTS: Thirty-nine eyes of 21 patients with PDS and either ocular hypertension or pigmentary glaucoma. MAIN OUTCOME MEASURES: A 24-hour intraocular pressure (IOP) profile, automated perimetry (Octopus G1), and analysis of photostereographs and HRT (Heidelberg Retina Tomograph) images of the optic disc were performed. Aqueous melanin granules were quantified using the cell count mode of the laser flare-cell meter (KOWA FC-1000) with undilated and dilated pupils. Granule counts were correlated with maximum and mean IOP, maximum range (amplitude) of IOP, mean defect of automated perimetry (G1-program), and damage to the optic disc was measured with the HRT. RESULTS: The number of aqueous melanin granules showed a strong correlation with maximum IOP in both undilated (r = 0.72, P < 0.001) and dilated eyes (r = 0.5, P = 0.02). A marginal correlation was found with the IOP range (r = 0.43, P = 0.04) and the mean defect of automated perimetry (r = 0.41, P = 0.06) in undilated eyes. The mean IOP and HRT measurements of the optic disc (area, volume of the neuroretinal rim, third moment in contour) showed no statistically significant correlation with the number of aqueous melanin granules (r < 0.4, P > 0.2). CONCLUSIONS: A larger number of aqueous melanin granules is strongly associated with high IOP and also with visual field loss, providing additional evidence of the relation between aqueous melanin dispersion and development of pigmentary glaucoma. Quantification of aqueous melanin granules with the laser flare-cell meter might be useful for evaluation of treatment effects, including laser iridotomy, in patients with PDS. PMID- 10711877 TI - A comparative study of betaxolol and dorzolamide effect on ocular circulation in normal-tension glaucoma patients. AB - OBJECTIVE: To determine whether dosages of a selective beta-blocking agent (betaxolol) and a topical carbonic anhydrase inhibitor (dorzolamide), sufficient to significantly lower intraocular pressure (IOP), have similar or disparate impact on the retinal and retrobulbar circulation. DESIGN: Counterbalanced crossover, with open-label use of medications. PARTICIPANTS: Nine persons with normal-tension glaucoma (NTG). INTERVENTION: After a 3-week drug washout, NTG patients were studied after 1 month of treatment with either dorzolamide or betaxolol, with determinations of IOP and retinal and retrobulbar hemodynamics. MAIN OUTCOME MEASURES: At baseline and after treatment with each drug, retinal arteriovenous passage time was determined by scanning laser ophthalmoscopy after fluorescein dye injection, and flow velocities in the central retinal and ophthalmic arteries were measured with color Doppler ultrasonography imaging. RESULTS: Betaxolol and dorzolamide each lowered IOP significantly, with these changes apparent and maximal after 2 weeks (each P < 0.05). In contrast, dorzolamide (but not betaxolol) accelerated arteriovenous passage of fluorescein dye in the inferior temporal quadrant of the retina (P < 0.05). Neither drug affected arteriovenous passage in the superotemporal retina or any aspect of central retinal or ophthalmic artery flow velocity after either 2 or 4 weeks. CONCLUSIONS: Although both dorzolamide and betaxolol are effective ocular hypotensive agents and their topical instillation leaves retrobulbar hemodynamics unaltered, dorzolamide alone accelerates inferotemporal retinal dye transit. PMID- 10711879 TI - Uveal effusion syndrome: clinical features, surgical treatment, histologic examination of the sclera, and pathophysiology. AB - PURPOSE: To clarify clinical features and pathophysiology and to evaluate surgical outcome of subscleral sclerectomy for primary uveal effusion syndrome. DESIGN: Prospective, consecutive noncomparative case series. PARTICIPANTS: Nineteen eyes of 16 patients diagnosed with uveal effusion syndrome treated in our clinic between 1989 and 1998. METHODS: Patients were examined by routine ophthalmologic examinations, fluorescein and indocyanine green angiography; measurement of the axial length of the eyeball; magnetic resonance imaging; and echography. Subscleral sclerectomy (sclerectomy under the scleral flap) was performed at the equator on all patients. Histologic examination of excised sclera was carried out on all samples. Patients were followed for outcome over time. MAIN OUTCOME MEASURES: Reattachment of the choroid and retina with resolution of the serous fluid. RESULTS: Three subgroups were identified: In type 1, nanophthalmic eye; the eyeball is small (average axial length 16 mm) and high hypermetropic (average +16 diopters); in type 2, the eyeball size is normal (average axial length 21 mm) with small refractive error; and in type 3, the eyeball size is normal. Histologically, types 1 and 2 demonstrated abnormal sclera with disorganization of collagen fiber bundles and deposits of proteoglycans in the matrix, whereas type 3 showed normal sclera. Subscleral sclerectomy was effective for types 1 and 2, inducing postoperative resolution of the subretinal fluid. However, type 3 eyes were not helped by this technique. CONCLUSIONS: Primary uveal effusion syndrome is caused by abnormalities of the sclera and increased resistance to transscleral fluid outflow as subscleral sclerectomy is an effective treatment in types 1 and 2 only, correct preoperative classification is essential for early surgical management. PMID- 10711880 TI - Diet and cataract: the Blue Mountains Eye Study. AB - PURPOSE: To investigate relationships between a wide range of macro- and micronutrients, including antioxidant vitamins, and the three main types of cataract in older people. DESIGN: Population-based cross-sectional study. PARTICIPANTS: Two thousand nine hundred people aged 49 to 97 years living in an urban community near Sydney, Australia. TESTING: Food frequency questionnaires and lens photography. MAIN OUTCOME MEASURE: Lens photographs were graded for presence and severity of cortical, nuclear, and posterior subcapsular cataracts. RESULTS: Higher intakes of protein, vitamin A, niacin, thiamin, and riboflavin were associated with reduced prevalence of nuclear cataract. After adjusting for multiple known cataract risk factors, the odds ratios for those in the highest intake quintile groups compared to those in the lowest intake quintiles were 0.5 (95% confidence interval [CI], 0.3-0.8) for protein, 0.5 (95% CI, 0.3-0.9) for vitamin A, 0.6 (95% CI, 0.4-0.9) for niacin, 0.6 (95% CI, 0.4-0.9) for thiamin, and 0.5 (95% CI, 0.3-0.9) for riboflavin. Intake of polyunsaturated fats was associated with reduced prevalence of cortical cataract. No nutrients were associated with posterior subcapsular cataract. CONCLUSIONS: The nucleus of the lens is particularly sensitive to nutrient deficiencies. Protein, vitamin A, niacin, thiamin, and riboflavin protected against nuclear cataract in this study. PMID- 10711882 TI - Anterior capsule opacification: a histopathological study comparing different IOL styles. AB - OBJECTIVE: To compare the degree of anterior capsule opacification (ACO) in human eyes obtained post-mortem containing various rigid and foldable posterior chamber intraocular lens (PC-IOL) designs. DESIGN: Comparative autopsy tissue study with clinicopathologic correlations. MATERIALS: Four hundred sixty human globes containing the following PC-IOL styles were analyzed: (1) one-piece polymethylmethacrylate (PMMA) optic-PMMA haptic (n = 50), (2) one-piece silicone plate IOL, large hole (n = 40), (3) one-piece silicone-plate IOL, small hole (n = 67), (4) three-piece PMMA optic-PMMA/Prolene haptic (n = 51), (5) three-piece acrylic optic-PMMA haptic (n = 96), (6) three-piece silicone optic-PMMA haptic (n = 24), (7) three-piece silicone optic-polyimide haptic (n = 40), and (8) three piece silicone optic-prolene haptic (n = 92). TESTING: The globes were sectioned in the equatorial plane for gross examination and then processed through paraffin; sectioned, and stained with hematoxylin-eosin, periodic acid-Schiff, and Masson's trichrome stains; and examined by light microscopy. MAIN OUTCOME MEASURES: Anterior capsule opacification was scored in each eye by grading the histologic sections from 0 to III, according to the amount (thickness) of proliferative tissue and cells measured in sagittal sections on the inner surface of the anterior capsule at the capsulorhexis margin. RESULTS: The difference among the eight groups was significant (P < 0.0001). Mean ACO scores were highest with the large and small hole one-piece silicone-plate lenses (1.77 +/- 0.86 and 1.28 +/- 0.77, respectively). The lowest mean score was observed in the group of three-piece acrylic optic-PMMA haptics lenses (0.51 +/- 0.52). CONCLUSIONS: Our results confirm previous clinical observations that the rate of ACO is relatively high with plate-haptic silicone IOLs. The lowest rate was noted with the three piece acrylic optic-PMMA haptic IOL. The IOL design and IOL material are significant factors in the development of ACO. PMID- 10711881 TI - Phacoemulsification versus extracapsular cataract extraction in patients with diabetes. AB - OBJECTIVE: To compare phacoemulsification with extracapsular cataract surgery in patients with diabetes and to identify determinants of postoperative visual acuity. DESIGN: Prospective, randomized, paired-eye trial. PARTICIPANTS: Forty six patients with diabetes and bilateral cataract. INTERVENTION: Patients were allocated to phacoemulsification surgery with silicone intraocular lens to one randomly determined eye, and extracapsular cataract surgery with 7-mm polymethylmethacrylate intraocular lens to the other. MAIN OUTCOME MEASURES: Logarithm of minimum angle of resolution visual acuity (logMAR VA), incidence of clinically significant macular edema (CSME), retinopathy progression, indices of anterior segment inflammation, and incidence of capsulotomy. RESULTS: Compared with eyes undergoing phacoemulsification, eyes managed with extracapsular surgery had more anterior chamber cells (P = 0.0004) and flare (P = 0.007) 1 week after surgery and a higher incidence of posterior synechiae (P = 0.04) and intraocular lens deposits (P < 0.0005) in the first postoperative year. The need for posterior capsulotomy was greater in eyes undergoing extracapsular surgery (16 of 46 vs. 5 of 46, P = 0.01). No difference in incidence of postoperative CSME, progression of retinopathy, or development of high-risk proliferative retinopathy was identified between techniques (P = 1.0, 0.8, and 0.2). Median 1-year logMAR VA was worse in eyes undergoing extracapsular surgery (0.08 vs. 0.06, P = 0.02), especially in those with retinopathy (0.14 vs. 0.08, respectively; P = 0.01). The presence or absence of CSME at the time of surgery was the most significant determinant of 1-year logMAR VA in regression models for both extracapsular (P = 0.0004, R2 = 0.45) and phacoemulsification groups (P < 0.00005, R2 = 0.46). CONCLUSIONS: Phacoemulsification is associated with better postoperative VA, less postoperative inflammation, and less need for capsulotomy than extracapsular cataract surgery in patients with diabetes. However, with both techniques, the principal determinant of postoperative VA appears to be the presence or absence of CSME at the time of surgery. Early intervention, reducing the risk that unrecognized CSME is present at the time of surgery, may be more critical to outcome than choice of surgical technique. PMID- 10711883 TI - Results of bilateral photorefractive keratectomy. AB - OBJECTIVE: To study the refraction and potential risks of bilateral photorefractive keratectomy for myopia. DESIGN: Retrospective, noncomparative case series. PARTICIPANTS: Four hundred eighteen consecutive patients (836 eyes) with myopia from -18 to -0.50 diopters (D) had bilateral simultaneous refractive surgery. INTERVENTION: Refractive surgery was performed with the Nidek EC-5000 slit-scanning excimer laser (Nidek Co., Tokyo, Japan). MAIN OUTCOME MEASURES: Manifest refraction, Snellen best spectacle-corrected visual acuity (BSCVA) and uncorrected visual acuity, haze, and halos were evaluated for 12 months after surgery. Both eyes of each patient were examined at the same follow-up after initial treatment. RESULTS: At 12 months after surgery, 95% of eyes were within +/- 1D of emmetropia, and 0.4% of patients had residual anisometropia more than 2 D. Thirty-three eyes (3.9%) were retreated, whereas 5 patients (1.2%) had retreatments in both eyes. Odds ratios between unilateral versus bilateral postoperative events were evaluated for the retreatments (1:0.22; P = 0.001), undercorrections more than 1 D (1:0.17; P < 0.0001), overcorrections more than 1 D (1:0.23; P < 0.0001), loss of 1 line of BSCVA (1:0.44; P < or = 0.002), and loss of 2 lines of BSCVA (1:0.18; P = 0.013). At 12 months after surgery, one eye with a loss of 2 lines of BSCVA was treated for haze more than 1; the fellow eye had haze 0.75. No patient had bilateral haze more than 1. At 12 months after surgery, no patient lost 2 lines of BSCVA in both eyes. There was a higher incidence of halos in both eyes rather than in one eye only (odds ratio: 4.17; P < 0.0001). No postoperative infections occurred. CONCLUSIONS: Bilateral events occurred approximately 2.5 to 4 times less often than unilateral events, except for the incidence of halos. The calculated odds ratio did not show an increased risk for the fellow eye while performing bilateral surgery, although not exempted from the risk of infection. It may be difficult to predict a complication in the fellow eye based on the results of the first operated eye. PMID- 10711884 TI - Outcome of sulcus fixation of dislocated posterior chamber intraocular lenses using temporary externalization of the haptics. AB - OBJECTIVE: This study evaluated the visual outcome and complications of repositioning and sulcus fixation of a dislocated posterior chamber intraocular lens (PC IOL) using a technique in which the haptics of the IOL are temporarily externalized for suture placement. DESIGN: Retrospective, noncomparative case series. PARTICIPANTS: Seventy-eight patients with a dislocated PC IOL. INTERVENTION: All patients underwent surgery to fixate the PC IOL using this technique. MAIN OUTCOME MEASURES: Patients were evaluated for visual acuity, refractive error, and surgical complications associated with the procedure. RESULTS: The average visual acuity before surgery was 20/205 (range, 20/20 to light perception), with a median refractive error of -1.00 diopters (D; range, 7.25-+15.00 D). After surgery, the average visual acuity improved to 20/72 (range, 20/20 to no light perception), with a median refractive error of -0.75 D (range, -5.50-+3.50 D). Patients were observed for a median of 15.5 months (range, 6-57 months). Twenty patients had postoperative cystoid macular edema (26%), 7 patients had an epiretinal membrane (ERM) (9%), and 5 patients had a retinal detachment (6%). Eight patients (10%) experienced iris capture of the sutured IOL, and in three patients (4%) the PC IOL dislocated again after surgery. CONCLUSIONS: This technique is an effective method for securing a dislocated PC IOL. PMID- 10711885 TI - Visual outcomes following the use of intravitreal steroids in the treatment of postoperative endophthalmitis. AB - OBJECTIVE: To compare visual outcomes between cases of acute postoperative endophthalmitis that did or did not receive intravitreal steroids. DESIGN: Retrospective nonrandomized comparative trial. PARTICIPANTS: Fifty-seven patients with postoperative endophthalmitis. INTERVENTION: Thirty-one patients with postoperative endophthalmitis resulting from cataract extraction received both intravitreal antibiotics and steroids, whereas the remaining 26 received only intravitreal antibiotics. MAIN OUTCOME MEASURES: Improvement in visual acuity. RESULTS: Multivariate logistic regression was used to analyze the variables that potentially influence a three-line visual acuity improvement. The mean baseline visual acuities of both groups were comparable. The use of intravitreal steroids reduced the probability of developing a three-line improvement in visual acuity (odds ratio [OR] = 0.287; 95% confidence interval [CI] [0.072-0.852]). On the basis of logistic regression analysis using our multivariate model, gender, baseline visual acuity, and pars plana vitrectomy were not significantly associated with visual outcome differences between the two groups. CONCLUSIONS: Patients who received intravitreal steroids had a significantly reduced likelihood of obtaining a three-line improvement in visual acuity. At a minimum our study provides no support for their use and, therefore, steroids may not be efficacious for acute endophthalmitis related to cataract extraction. PMID- 10711886 TI - The deep plane facelift: a 20-year evolution of technique. AB - PURPOSE: This report reviews the unique technical and conceptual oculoplastic innovations in the discipline of facelift surgery by analyzing the evolution of facelift technique at a university-based oculoplastic program. DESIGNED: Retrospective, noncomparative case series. PARTICIPANTS: We analyzed 313 patients undergoing a facelift from 1980 through 1997. Most procedures were performed by the senior author. METHODS: Three primary eras of surgical technique were identified: limited skin flap with superficial musculo-aponeurotic system plication (25 patients), extended skin flap with neck dissection and superficial musculo-aponeurotic system plication (210 patients), and deep plane facelift with robust superficial musculo-aponeurotic system flap (78 patients). RESULTS: The steps in the evolution were designed to improve the results of the surgery regarding rejuvenation of the neck, jowls, and nasolabial fold, and to reduce the "tattletale signs" of facelift surgery including postauricular scarring, change in the position of the sideburn and temporal hairline, and unnatural results caused by pulling the tissues posteriorly, rather than repositioning them vertically. There were no complications in the skin flap only group. In the extended skin flap and superficial musculo-aponeurotic system plication group, there was one mandibular paresis which partially resolved. In the deep plane facelift (n = 78), there was one laceration of the parotid duct, successfully stented during surgery. CONCLUSIONS: The deep plane facelift, with vertical elevation of the midface, jowls, and neck, is a logical extension of the mid facelifting techniques that have been used by oculoplastic surgeons. Compared with cutaneous undermining with superficial musculo-aponeurotic system plication, we found patient and physician acceptance higher using the deep plane technique. PMID- 10711887 TI - Postherpetic lacrimal obstruction. AB - OBJECTIVE: The purpose of the study was to review the surgical management of postherpetic lacrimal obstruction. DESIGN: Retrospective noncomparative case series. PARTICIPANTS: One hundred sixty patients (111 female, 49 male) with a mean age at presentation of 31 years. All had a history typical of primary herpes simplex blepharoconjunctivitis. INTERVENTION: Open lacrimal surgery was performed on 158 patients (171 eyes), of whom 99 patients (108 eyes) had undergone no surgery before being seen at Moorfields. The most common primary procedure was dacryocystorhinostomy (DCR) with anterograde or retrograde intubation (94 eyes, 54%), and primary placement of a Lester Jones tube (conjunctivo-DCR) was performed in 56 eyes (32%). A secondary procedure was required in 43 eyes (26%), the most common being closed placement of a Lester Jones tube (40 eyes). MAIN OUTCOME MEASURES: Persistent symptoms of impaired lacrimal drainage and need for additional surgery. RESULTS: Reduction of epiphora was good or complete in 171/173 eyes (98%). CONCLUSIONS: This study shows that there is a justification for procedures that use any remaining unaffected portion of canaliculi, such as DCR, with anterograde or retrograde intubation, as well as the more commonly used Lester Jones tube. PMID- 10711888 TI - Clinical diversity of hereditary Duane's retraction syndrome. AB - OBJECTIVE: To define the spectrum of ophthalmic manifestations of Duane's retraction syndrome (DRS) in a large family. DESIGN: Cross-sectional study of 110 among 114 living relatives in an extended family. METHODS: History and ophthalmic examination obtained on all participants. MAIN OUTCOME MEASURES: Ocular motility, strabismus, visual acuity, binocularity, associated neurologic problems. RESULTS: Twenty-five individuals were affected with DRS. Twenty-four subjects (96%) had bilateral DRS, but there was a broad spectrum of severity. Strabismus occurred in 76% and amblyopia in 48%. Associated findings included fourth cranial nerve palsy, partial third cranial nerve palsy, nystagmus, seizures, and deafness. Fourth cranial nerve palsies and manifest strabismus tended to cluster within single family units. CONCLUSIONS: Strabismus and amblyopia are much more common with bilateral DRS than with unilateral DRS. There is much phenotypic variability among individuals within families with hereditary Duane's syndrome. The responsible gene(s) may affect the development of many cranial nerves. Genetic compounding may play a role in the phenotypic segregation seen within this large family. PMID- 10711889 TI - The natural history and ophthalmic involvement in childhood myasthenia gravis at the hospital for sick children. AB - OBJECTIVE: To characterize signs, symptoms, and the natural history of myasthenic syndromes in pediatric patients. DESIGN: Retrospective noncomparative case series. PARTICIPANTS: Thirty-four patients with a diagnosis of myasthenia were identified from either the hospital's or treating physician's database. METHODS: Retrospective chart review, clinical examination, and telephone interview. MAIN OUTCOME MEASURES: Information pertaining to the ophthalmologic and neurologic examination, diagnostic interventions, and treatment was noted. Patients with active disease, attending during the study period, were examined at their outpatient visits. Those who no longer attended the hospital were contacted by means of a telephone interview to complete their follow-up. RESULTS: Thirty-four children were found to have myasthenia. Two had transient neonatal myasthenia, which resolved quickly. Seven (20.6%) patients had congenital myasthenic syndromes (CMS) and 25 (73.5%, 19 females) were affected with autoimmune myasthenia gravis (AMG). In those patients with severe CMS, three showed signs of generalized weakness, including failure to thrive, frequent apneas, and aspirations. In four patients with mild CMS, eye signs were relatively more prominent. In all patients with CMS, strabismus, ophthalmoplegia, and ptosis were the main ophthalmologic signs and remained relatively constant. Fourteen (56%) patients with AMG had ocular signs and symptoms, and five of them progressed to systemic involvement in 7.8 months on average (range, 1-23). The remaining nine patients with ocular AMG had either strabismus or ptosis and were treated with pyridostigmine (nine patients) and prednisone (two patients). Patients with ocular AMG were seen at 78 months on average, those with systemic AMG at 85.6 months. Systemic AMG was seen in 16 patients. No thymomas were found in 14 patients who underwent thymectomy. Of the 25 patients with AMG, 8 are still being treated, 8 are in remission for an average of 65.2 months and are asymptomatic, 4 patients are receiving long-term immunosuppressants (1 has likely sustained permanent damage to her extraocular muscles with complete ophthalmoplegia and ptosis), and 4 have been lost to follow-up. Finally, one patient died after aspiration because of bulbar weakness. CONCLUSIONS: Patients with CMS varied in the degree of severity. Apneic attacks, aspiration, and failure to thrive may obscure the diagnosis. Compared with AMG, their ophthalmologic signs and symptoms were usually permanent. Visual signs and symptoms were usually prominent in those patients with active AMG, but those in remission were asymptomatic. More than half of the patients with juvenile AMG had ocular symptoms. Generalization occurred in a minority in an average of 7.8 months. Patients entered remission after approximately 2 years of treatment and were visually asymptomatic. This study suggests that long-term permanent damage to the extraocular muscles as a result of juvenile AMG is rare. Myasthenia gravis is a life-threatening disease as evidenced by the death of one of our patients. Many of these patients are first seen by the ophthalmologist who can aid the diagnosis, screen for amblyopia, and monitor the patient's response to therapy. PMID- 10711890 TI - Visual acuity in children with coloboma: clinical features and a new phenotypic classification system. AB - OBJECTIVE: The aims of this study were to describe the clinical features and biometric findings in the eyes of children with coloboma and to develop a classification of coloboma that correlates with visual function. DESIGN: Retrospective observational case series. PARTICIPANTS: One hundred thirteen children and young adults (48 female, 65 male) aged 0 to 20 years with 196 eyes having coloboma. METHODS: Children with coloboma were recruited from schools for the blind, integrated education programs, schools for the mentally handicapped, community-based rehabilitation services, and hospital clinics in Andhra Pradesh, India, between January 1998 and January 1999. Visual function was assessed, including distance and near visual acuity (VA), and navigational vision. The corneal diameter and axial length of eyes were measured wherever possible. MAIN OUTCOME MEASURES: Anatomic site of coloboma, association with microcornea and/or microphthalmos, VA, presence of navigational vision and reading vision. RESULTS: Of 196 eyes with colobomatous malformations, 11 had microphthalmos with cyst, and 185 eyes had coloboma (associated with microcornea in 155 eyes and with a normal corneal diameter in 30). Microphthalmos was present in 72 of the 185 eyes with coloboma, of which 71 of 72 also had microcornea. The prognosis for vision depended on the phenotype of the better eye. Microphthalmos with cyst had the worst prognosis (VA < 3/60, 100%; reading and navigational vision, 0%). Microcornea with microphthalmos had a worse prognosis than microcornea without microphthalmos (VA < 3/60: 66.7% vs. 23.3%; unable to read N10: 66.7% vs. 34.1 %; no navigational vision: 30.6% vs. 6.73%). Simple coloboma (no microcornea or microphthalmos) had the best prognosis (VA < 3/60: 6.7%; able to read N10: 93.3%; navigational vision: 100%). A corneal diameter <6 mm had a poor visual prognosis, whereas a corneal diameter >10 mm had a good prognosis. CONCLUSIONS: A phenotypic classification of coloboma is proposed, which in this study showed a good correlation with visual acuity, reading, and navigational vision. Microphthalmos with cyst had the worst prognosis, coloboma with microcornea and microphthalmos a poor prognosis, coloboma with only microcornea had an intermediate prognosis, and simple coloboma had the best prognosis. PMID- 10711891 TI - Levodopa may improve vision loss in recent-onset, nonarteritic anterior ischemic optic neuropathy. AB - OBJECTIVE: To study the effect of levodopa in improving visual function in patients treated within 45 days of onset of nonarteritic anterior ischemic optic neuropathy (NAION). DESIGN: Nonrandomized, retrospective, comparative trial. PARTICIPANTS: The study involved 37 patients with NAION of less than 45 days duration. METHODS: Eighteen patients who had been treated with levodopa were assigned to the case group, and 19 untreated patients were assigned to the control group. Snellen visual acuity converted to logMAR and mean deviation on Humphrey automated perimetry (Program 24-2, Humphrey Instruments, San Leardro, CA) were evaluated at the initial and 6-month visits. INTERVENTION: The 18 patients in the case group were administered a capsule of 100 mg levodopa/25 mg carbidopa (Sinemet 25-100) three times daily for 3 weeks. MAIN OUTCOME MEASURES: The primary outcome measures were changes in visual acuity and visual field at 6 months from baseline. Improvement in visual acuity was defined as a difference of -0.3 logMAR or less between the 6-month and initial visual acuities, whereas worsened visual acuity was a difference of +0.3 logMAR or more. Each 0.3 LogMar represented a doubling of the visual angle, i.e., a change by three lines on the eye chart. Improvement in visual field was defined as a difference in mean deviation of +3.0 dB or more between the 6-month and initial visual field tests, whereas worsened visual field was a difference in mean deviation of -3.0 dB or less. RESULTS: The proportions of patients with worsened, unchanged, and improved visual acuity at 6 months were compared for the levodopa and control groups. There was a significant difference (P = 0.012) between the groups. Examination of the proportions showed that a higher proportion of patients who received levodopa had improved visual acuity with a corresponding lower proportion having worsened visual acuity as compared with the control patients. Ten of 13 patients (76.9%) in the levodopa group with 20/40 visual acuity or worse at baseline had improved visual acuity at 6 months, and none of the 18 patients had worsened visual acuity. In contrast, 3 of 10 control patients (30%) with 20/40 visual acuity or worse at baseline had improved visual acuity at 6 months, and 3 of 19 control patients (16.3%) had worsened visual acuity. The proportions of patients with worsened, unchanged, and improved visual fields at 6 months were compared for the levodopa and control groups. There was no significant difference between the groups (P = 0.25). CONCLUSIONS: Patients treated with levodopa within 45 days of onset of NAION were more likely to experience improvement and less likely to have worsened visual acuity than untreated patients. Levodopa appears to be beneficial in the treatment of recent-onset NAION. PMID- 10711892 TI - A comparison of tangent screen, goldmann, and humphrey perimetry in the detection and localization of occipital lesions. AB - OBJECTIVE: To compare manual kinetic perimetry with tangent screen and Goldmann techniques and automated static perimetry with the Humphrey Field Analyzer in the detection and localization of occipital lobe lesions. DESIGN: Prospective consecutive comparative case series. PARTICIPANTS: Twelve patients with well defined occipital lobe infarcts on magnetic resonance (MR) imaging were studied. MAIN OUTCOME MEASURES: The patients were tested by tangent screen, Goldmann, and Humphrey perimetry (central 30-2 threshold program). The three visual fields were compared and correlated with MR images. RESULTS: All three perimetric techniques detected the presence of postchiasmal lesions. However, localization of lesions differed with perimetric technique. Visual fields obtained from tangent screen and Goldmann perimetry were similar and corresponded well with the location of lesions on MR images in all 12 patients. Humphrey perimetry inaccurately localized the lesion to the proximal part of the postchiasmal pathway by revealing incongruous fields in two patients, failed to detect sparing of the posterior occipital cortex or occipital pole in four patients, and estimated a larger extent of damage in one patient when compared with MR images and manual perimetry. CONCLUSIONS: All three perimetric techniques are satisfactory screening tests to detect occipital lesions. However, tangent screen and Goldmann perimetry provide information about the location and extent of lesions that is more consistent with prevailing knowledge of the effects of the lesion in the postgeniculate visual pathway. PMID- 10711893 TI - The role of restricted motility in determining outcomes for vertical strabismus surgery in Graves' ophthalmology. AB - OBJECTIVE: To identify factors predictive of operative success or failure for vertical muscle surgery performed in patients with Graves' ophthalmopathy. DESIGN: Prospective noncomparative case series. PARTICIPANTS: Thirty-one consecutive patients with Graves' ophthalmopathy who demonstrated vertical ocular motor imbalance, with or without simultaneous horizontal muscle imbalance. INTERVENTION: Vertical extraocular muscle surgery performed either in isolation or in association with horizontal muscle surgery. MAIN OUTCOME MEASUREMENTS: Vertical limitations of extraocular muscles were correlated with preoperative hypertropia. Stepwise linear regression was used to determine the significant predictors of postoperative hypertropia in primary gaze. Logistic analysis was used to estimate the probability of surgical failure (>5 diopters) on the basis of preoperative parameters. RESULTS: The amount of preoperative hypertropia was negatively correlated with total restriction of vertical ductions (r = -0.52, P < 0.01). Preoperative hypertropia was positively correlated with asymmetry in muscle restriction between the two eyes (r = 0.67, P < 0.0001). The best predictor of preoperative hypertropia was the difference between restriction of the contralateral opposing recti, namely the right superior rectus, and the left inferior rectus, as well as the right inferior rectus and the left superior rectus (r = 0.74, P < 0.0001). Restriction of the contralateral opposing recti was also the most significant predictor of surgical success (postoperative hypertropia < 5 prism diopters). CONCLUSIONS: Surgery tailored to address restriction of ductions, specifically the difference between contralateral opposing recti, is likely to improve the success of initial surgery beyond that based primarily on the magnitude of the vertical deviation. PMID- 10711894 TI - Malignant granular cell tumor metastatic to the orbit. AB - OBJECTIVE: Malignant granular cell tumor is a rare type of soft tissue sarcoma. To our knowledge, ocular (eyelid) involvement has been described in only two cases. Herein, we report the clinicopathologic features of an unusual case of malignant granular cell tumor metastatic to the orbit. DESIGN: Observational case report. METHODS: Retrospective review of the medical record and the histopathologic and electron microscopic findings and review of the literature. RESULTS: A 72-year-old man with biopsy-proven granular cell tumor in the cervical region was initially seen with proptosis and motility disturbance. A magnetic resonance imaging scan showed a large intraconal mass, and biopsy of the orbital mass revealed granular cell tumor. Histopathologic examination of the primary neck tumor and the orbital mass revealed increased nuclear atypia and pleomorphism in the consecutive lesions. The morphologic impression of granular cell tumor was also supported by the immunohistochemical demonstration of S-100 protein expression and ultrastructural findings typical of granular cell tumor. Six months after the orbital involvement, systemic workup revealed multiple apparent bony and lung metastases. CONCLUSIONS: We report the first malignant granular cell tumor metastatic to the orbit and suggest the inclusion of this tumor in the differential diagnosis of metastatic orbital lesions. PMID- 10711895 TI - Survival after rhino-orbital-cerebral mucormycosis in an immunocompetent patient. AB - OBJECTIVE: Rhino-orbital-cerebral mucormycosis is usually associated with a poor prognosis and is almost exclusively seen in immunocompromised patients. We report the third documented case of rhino-orbital-cerebral mucormycosis caused by Apophysomyces elegans (a new genus of the family Mucoraceae first isolated in 1979) in an immunocompetent individual. Orbital exenteration and radical debridement of involved adjacent structures combined with intravenous liposomal amphotericin resulted in patient survival. DESIGN: Interventional case report. METHOD: A 59-year-old immunocompetent white man sustained a high-pressure water jet injury to the right inner canthus while cleaning an air conditioner filter. He later had "orbital cellulitis" develop that did not respond to antibiotics and progressed to orbital infarction. Imaging studies and biopsy results led to a diagnosis of mucormycosis. Tissue culture grew Apophysomyces elegans, a new genus of the family Mucoraceae first isolated in 1979. Orbital exenteration and radical debridement of involved adjacent structures, combined with intravenous liposomal amphotericin, resulted in patient survival. RESULTS: After orbital exenteration and debridement of involved adjacent structures along with intravenous liposomal amphotericin, our patient has remained free from relapse with long-term follow up. CONCLUSIONS: The agent causing this case of rhino-orbital-cerebral mucormycosis (Apophysomyces elegans) contrasts with the three genera most commonly responsible for mucormycosis (Rhizopus, Mucor, and Absidia) in that infections with this agent tend to occur in warm climates, by means of traumatic inoculation, and in immunocompetent patients. Rhino-orbital-cerebral mucormycosis should be considered in all patients with orbital inflammation associated with multiple cranial nerve palsies and retinal or orbital infarction, regardless of their immunologic status. A team approach to management is recommended for early, appropriate surgery and systemic antifungal agents. PMID- 10711896 TI - Comparison of topographic indices that correlate with visual acuity in videokeratography. AB - OBJECTIVE: To evaluate the relationship between the best spectacle-corrected visual acuity (BSCVA) and two quantitative indices of the anterior corneal surface obtained by videokeratography. DESIGN: Prospective, single center, comparative, observational study. PARTICIPANTS: Eighty-nine normal eyes and 52 eyes with keratoconus with contact lens-corrected visual acuity of 20/20 or better. INTERVENTION: Videokeratography was performed with the TMS-2 and the CAS system 2000. MAIN OUTCOME MEASURES: The relationship between the BSCVA recorded in log minimal angle of resolution (logMAR) units, the surface regularity index (SRI), and the predicted corneal acuity (PCA) were assessed by linear regression analysis. RESULTS: The BSCVAs for all eyes ranged from 0.82 to -0.30 logMAR units. BSCVA was highly correlated with the SRI (r = 0.70, P < 0.0001) and the PCA (r = -0.61, P < 0.0001). There was no statistical difference in the regression slopes and the intercepts for the estimated BSCVA using the SRI and measured BSCVA, and the estimated BSCVA using PCA and measured BSCVA. CONCLUSIONS: Two quantitative topographic indices, SRI and PCA, are useful for estimating the effect of irregular astigmatism on visual acuity even though both indices quantify different aspects of the anterior surface of the cornea. PMID- 10711897 TI - In vivo confocal microscopy of patients with corneal recurrent erosion syndrome or epithelial basement membrane dystrophy. AB - OBJECTIVE: To characterize morphologic changes in corneas of patients with recurrent erosion syndrome or epithelial basement membrane dystrophy using in vivo confocal microscopy. DESIGN: Observational case series PARTICIPANTS: Fourteen eyes of eight patients with diagnosed epithelial basement membrane dystrophy and 13 eyes of seven patients with recurrent erosion syndrome were examined. METHODS: Slit-lamp examination and in vivo confocal microscopy. The pathologic findings are presented as digitized images obtained from video tape recorded during the confocal microscopy. MAIN OUTCOME MEASURES: The morphology of corneal surface epithelial cells, basal epithelial cells, subbasal nerve plexus, Bowman's layer, stromal keratocytes, and endothelium was analyzed. RESULTS: The surface epithelium was intact in all but two eyes. One cornea (a basement membrane disorder with clinically visible dots) had multinucleate surface epithelial cells, and one eye with recurrent corneal erosions showed a freely floating surface epithelium sheet in the tear fluid. Patients in both groups showed islets of highly reflective cells with presumed intracellular deposits surrounded by normal cells in the basal epithelial cell layer. The basal epithelial cell area also showed other pathologic changes, including drop-shaped configurations, streaks, or ridges. Folding of the Bowman's layer was also observed in both groups. Anterior keratocytes showed signs of activation (highly reflective nuclei with visible processes) in some of the patients regardless of the clinical diagnosis, and in recurrent erosions even increased deposition of abnormal extracellular matrix in the anterior stroma was suspected. Posterior corneal keratocytes and endothelium appeared normal when examined. The subbasal nerve plexus showed various pathologic changes, such as short or strangely shaped nerve fiber bundles, decreased numbers of long nerve fiber bundles, only faintly visible long nerve fiber bundles (instead of the normally observed long parallel running interconnected bundles), or increased amounts of Langerhans cells, but only one patient (with recurrent erosion syndrome) lacked the subbasal nerve plexus. CONCLUSIONS: In vivo confocal microscopy of corneas with recurrent erosions or epithelial basement membrane dystrophy showed deposits in basal epithelial cells, subbasal microfolds and streaks, damaged subbasal nerves, or altered morphology of the anterior stroma. Confocal microscopy cannot replace biomicroscopy in making a specific diagnosis, but it sometimes helps the diagnosis in corneas that appear normal under a biomicroscope. PMID- 10711898 TI - Keratomycosis: clinical and microbiologic experience with dematiaceous fungi. AB - OBJECTIVE: To assess the significance of dematiaceous fungi in the causation of keratomycosis. DESIGN: Retrospective, noncomparative case series. PARTICIPANTS: Eighty-eight cases of dematiaceous fungal keratitis seen at the L. V. Prasad Eye Institute, Hyderabad, India from January 1991 through December 1996. INTERVENTION: Only culture-proven cases were analyzed. MAIN OUTCOME MEASURES: Predisposing factors, clinical characteristics, microbiology, treatment methods, and outcome. RESULTS: Of 557 cases of fungal keratitis seen during the study period, dematiaceous fungi were the etiologic agents in 88 (15.7%), after Fusarium in 210 (37.6%) and Aspergillus species in 170 cases (30.4%), respectively. Trauma was the most common predisposing factor (47.7%). Fifty-three eyes (61.3%) had the classical clinical picture of yellow-white, dry raised infiltrate with feathery hyphate edges at initial examination. The characteristic macroscopic pigmentation was seen in only 24 eyes (27.27%). Septate branching fungal filaments were identified in 78 smears (88.63%) on light microscopy, of which 5 (5.7%) also showed the presence of bacteria. Curvularia species dominated the spectrum (22.7%). Treatment was started in 48 eyes with topical antifungal agents, whereas 37 received both oral and topical antifungal agents. Outcome data were available for 68 cases. Forty-nine (72%) responded to medical therapy, whereas 13 eyes required therapeutic penetrating keratoplasty and 6 eyes had to be eviscerated. CONCLUSIONS: This is the largest series of keratitis caused by dematiaceous fungi reported to date. It clearly brings out the clinical importance of this group of corneal infections. PMID- 10711899 TI - The expanding clinical spectrum of ocular lyme borreliosis. AB - OBJECTIVE: To delineate the clinical manifestations of ocular Lyme borreliosis, while concentrating on new symptoms and findings and the phase of appearance of ophthalmologic disorders. DESIGN: Observational case series. PARTICIPANTS: Ten patients with Lyme borreliosis-associated ophthalmologic findings previously reported from the Helsinki University Central Hospital in addition to 10 new cases that have since been diagnosed. INTERVENTION/TESTING: The patients underwent medical and ophthalmologic evaluation. The diagnosis of Lyme borreliosis was based on medical history, clinical ocular and systemic findings, determinations of antibodies to Borrelia burgdorferi by enzyme-linked immunosorbent assay and immunoblot analysis, the detection of DNA of B. burgdorferi by polymerase chain reaction, and exclusion of other infectious and inflammatory causes. MAIN OUTCOME MEASURES: Ocular complaints, presenting ophthalmologic findings, and the stage of Lyme borreliosis were recorded. RESULTS: Four patients presented with a neuro-ophthalmologic disorder, five had external ocular inflammation, 10 patients had uveitis, and one had branch retinal vein occlusion. One patient developed episcleritis and one patient developed abducens palsy within 2 months of the infection incident. In the remaining 14 patients in whom the time of infection was traced, the ocular manifestations appeared in the late stage of Lyme borreliosis. Two patients with a neuro ophthalmologic disorder and one with external ocular inflammation experienced severe photophobia, whereas the main reported symptom of the patients with uveitis was decreased visual acuity. Four patients with external ocular disease and one with a neuro-ophthalmologic disorder experienced severe periodic ocular or facial pain. Retinal vasculitis developed in seven patients with uveitis. CONCLUSIONS: Lyme borreliosis can cause a variety of ocular manifestations, which develop mainly in the late stage of the disease. Photophobia and severe periodic ocular pain can be characteristic symptoms of Lyme borreliosis. In the differential diagnosis of retinal vasculitis, Lyme borreliosis should be taken into account, especially in endemic areas. PMID- 10711900 TI - The use of topical aqueous suppressants in the prevention of postoperative intraocular pressure elevation after pars plana vitrectomy with long-acting gas tamponade. AB - OBJECTIVE: To determine whether topical aqueous suppressant therapy applied after pars plana vitrectomy with gas tamponade prevents postoperative intraocular pressure (IOP) elevation. DESIGN: Prospective, nonrandomized comparative study. PARTICIPANTS: Forty-one patients who met inclusion criteria and underwent pars plana vitrectomy with gas tamponade (SF6 18%-20% or C3F8 12%-16%) over a 1-year period. INTERVENTION: Treatment eyes received topical aqueous suppressants at the end of surgery. MAIN OUTCOME MEASURES: Postoperative IOP at 4 to 6 hours, 1 day, and 1 week. RESULTS: Twenty-one control and 20 treatment eyes met the inclusion criteria. The IOP (in mmHg) measured at 4 to 6 hours (23.05 [control, 14.73 [treatment]) and 1 day (23.24 [control], 17.28 [treatment]) postoperatively showed a statistically significant difference between the groups (P = 0.0038) at 4 to 6 hours and a trend toward significance (P = 0.057) at 1 day. Eleven control and three treatment eyes had an IOP spike above 25 mmHg at 4 to 6 hours or 1 day postoperatively (P = 0.02), and six control eyes and one treatment eye had postoperative IOP greater than 30 mmHg. A pressure rise greater than 40 mmHg was seen in two control eyes and no treatment eyes. CONCLUSIONS: Use of topical aqueous suppressants after pars plana vitrectomy with long-acting gas tamponade is effective in preventing significant postoperative IOP elevation in most cases. PMID- 10711901 TI - Comparison between optical coherence tomography and fundus fluorescein angiography for the detection of cystoid macular edema in patients with uveitis. AB - PURPOSE: To compare optical coherence tomography (OCT) with fundus fluorescein angiography (FFA) for the detection of cystoid macular edema (CME) in patients with uveitis. DESIGN: Prospective comparative observational series. PARTICIPANTS: One hundred twenty-one eyes of 58 patients with uveitis of varied causes (seven patients were studied twice). TESTING: Patients with suspected CME underwent OCT scanning followed by FFA at the same visit. MAIN OUTCOME MEASURES: Detection and distribution of macular edema. RESULTS: One hundred eight eyes had similar results on both OCT and FFA in that 67 eyes had CME and 41 eyes had no CME. In 10 eyes subretinal fluid was detected on OCT but not FFA. Five of these eyes had CME on FFA but not OCT. Three other eyes had CME that was detected by FFA but not by OCT. Compared with FFA, the OCT sensitivity for detecting CME was 96% (including the eyes with subretinal fluid), and the OCT specificity was 100%. CONCLUSIONS: OCT is as effective at detecting CME as is FFA but is superior in demonstrating axial distribution of fluid. PMID- 10711902 TI - Identification of ingrowth site of idiopathic subfoveal choroidal neovascularization by indocyanine green angiography. AB - PURPOSE: This study aimed to determine whether indocyanine green (ICG) angiography is useful to identify the ingrowth site of idiopathic choroidal neovascularization (CNV), which can predict visual outcomes after surgical removal of idiopathic CNV. DESIGN: Consecutive, observational case series. PARTICIPANTS: Twenty-six patients with idiopathic subfoveal CNV, of whom six underwent submacular surgery. INTERVENTION: Indocyanine green videoangiography with a scanning laser ophthalmoscope. MAIN OUTCOME MEASURES: We studied ICG videoangiographic images of choroidal neovascular membranes from the early phase to the late phase with special attention to abnormal findings, which can indicate the ingrowth site of CNV. RESULTS: Early ICG angiography demonstrated distinct neovascular vessels in 24 of the 26 patients (92%). Hypofluorescent rims continuously or intermittently surrounded neovascular membranes on late ICG angiograms in 21 of the 26 patients (81%). In 22 of the 26 patients (85%), ICG angiography demonstrated hypofluorescent areas within the CNV. These hypofluorescent areas frequently became ring shaped in the middle to late phase of the ICG angiography. In 14 of 16 patients (88%) with CNV larger than half a disc area, the filling of neovascular vessels appeared from the inside of the hypofluorescent areas and branched out toward the surrounding hyperfluorescent membrane in the early phase. In all six patients who underwent surgical removal of CNV, ICG videoangiography showed these hypofluorescent areas from which neovascular vessels emanated. Three of the four surgical patients, in whom hypofluorescent areas or central fluorescent areas surrounded by ring-shaped hypofluorescence were extrafoveal or juxtafoveal, had a best postoperative visual acuity of 20/60 or better. In contrast, both surgical patients with subfoveal hypofluorescent areas had a best postoperative visual acuity of 20/70 or worse. CONCLUSIONS: Although further observations are needed, ICG angiography may be a useful adjunct in the identification of the ingrowth site of idiopathic CNV, which can predict visual outcomes after surgery. PMID- 10711903 TI - Intraocular foreign bodies: management, prognostic factors, and visual outcomes. AB - OBJECTIVE: To determine prognostic factors and visual outcomes in patients with intraocular foreign bodies (IOFBs). DESIGN: Retrospective, noncomparative, interventional case series. PARTICIPANTS: Fifty-nine consecutive patients undergoing removal of an IOFB with a minimum of 6 months of follow-up. INTERVENTION: Surgical removal of the IOFB and repair of associated ocular trauma. MAIN OUTCOME MEASURES: Final best corrected visual acuity was the main outcome measured. Ocular findings at presentation were compared with final visual acuity to determine prognostic factors for visual outcome. RESULTS: Final best corrected visual acuity of 20/40 or more was obtained in 42 patients (71%) and ambulatory vision (>5/200) was achieved in 50 patients (85%). Presenting visual acuity was predictive of final visual outcome (P < 0.01). Prognostic factors for a better visual outcome (P < 0.05) included better presenting visual acuity and hammering metal on metal as the mechanism of injury. Prognostic factors for a poor outcome (P < 0.05) included poor presenting visual acuity, the presence of an afferent pupillary defect, and vitreous hemorrhage. CONCLUSIONS: Final visual outcomes were excellent in 71% of patients. Presenting visual acuity was the strongest predictor of final visual outcome in this series. Additional predictive factors included the mechanism of injury, the presence of an afferent pupillary defect, and vitreous hemorrhage. PMID- 10711904 TI - Vitrectomy for macular hemorrhage associated with retinal arterial macroaneurysm. AB - OBJECTIVE: To evaluate the safety and efficacy of pars plana vitrectomy to treat massive macular hemorrhage caused by retinal arterial macroaneurysm. DESIGN: Retrospective case series. PARTICIPANTS: Eight eyes of eight patients. METHODS: We retrospectively reviewed the charts for eight eyes of eight patients in which pars plana vitrectomy had been performed to remove a massive macular hemorrhage secondary to a ruptured retinal arterial macroaneurysm. In each case, the preretinal/intralamellar hemorrhage was removed, and in three of the eight eyes a subretinal hemorrhage was removed via a retinotomy after clot lysis using tissue plasminogen activator. MAIN OUTCOME MEASURE: Pars plana vitrectomy to treat macular hemorrhage secondary to retinal arterial macroaneurysm. RESULTS: The duration of symptoms ranged from 10 to 80 days (average, 31 days). The preoperative visual acuities ranged from counting fingers to 0.09. Follow-up ranged from 3 to 36 months (average, 19 months). The postoperative visual acuities improved in seven eyes and remained unchanged in one; vision was better than 0.1 in six eyes and better than 0.4 in five. Postoperative complications included a mild vitreous hemorrhage in two eyes, a macular hole in one, and a cataract in two. CONCLUSIONS: Pars plana vitrectomy appears to be relatively safe and effective in treating massive macular hemorrhage caused by a retinal arterial macroaneurysm. PMID- 10711905 TI - A 40-year perspective on the prevalence of depression: the Stirling County Study. AB - BACKGROUND: According to epidemiologic studies that use recall of lifetime episodes, the prevalence of depression is increasing. This report from the Stirling County Study compares rates of current depression among representative samples of adults from a population in Atlantic Canada. METHODS: Sample sizes were 1003, 1201, and 1396 in 1952, 1970, and 1992, respectively. The depression component of the study's method, the DPAX (DP for depression and AX for anxiety), was employed. The original procedure (DPAX-1) was applied in all years. A revision (DPAX-2) was used in 1970 and 1992. The Diagnostic Interview Schedule (DIS) was also used in 1992. RESULTS: With the DPAX-1, the overall prevalence of current depression was steady at 5% over the 2 early samples but declined in 1992 because of vernacular changes referring to dysphoria. The DPAX-2 gave a stable overall prevalence of 5% in the 2 recent samples, but indicated that women and younger people were at greater risk in 1992 than in 1970. The DIS, like the DPAX 2, found a current 1992 rate of 5% for major depressive episodes combined with dysthymia. Recalled lifetime rates using the DIS showed the same profile interpreted in other studies as suggesting an increase in depression over time. CONCLUSIONS: Three samples over a 40-year period showed a stable current prevalence of depression using the DPAX methods that was comparable in 1992 with the current rates using the DIS. This casts doubt on the interpretation that depression is generally increasing. Within the overall steady rate observed in this study, historical change was a matter of redistribution by sex and age, with a higher rate among younger women being of recent origin. PMID- 10711906 TI - A comparison of self-report and clinical diagnostic interviews for depression: diagnostic interview schedule and schedules for clinical assessment in neuropsychiatry in the Baltimore epidemiologic catchment area follow-up. AB - BACKGROUND: The field of psychiatric epidemiology continues to employ self-report instruments, but the low degree of agreement between diagnoses achieved using these instruments vs. that achieved by psychiatrists in the clinical modality threatens the credibility of the results. METHODS: In the Baltimore Epidemiologic Catchment Area follow-up, 349 individuals who had a Diagnostic Interview Schedule (DIS) interview were blindly examined by psychiatrists using the Schedules for Clinical Assessment in Neuropsychiatry (SCAN). Comparisons were made at the level of diagnosis, syndrome, and DSM-IV symptom group. Indexes of agreement were computed and characteristics of discrepant cases were identified. RESULTS: Agreement on diagnosis of major depressive disorder was only fair (kappa = 0.20), with the DIS missing many cases judged to meet criteria for diagnosis using the SCAN (29% sensitivity). A major source of discrepancy was respondents with false negative diagnoses who repeatedly failed to report DIS symptoms attributed to life crises or medical conditions. Older age, male sex, and lower impairment were associated with underdetection by the DIS, using logistic regression analysis. In spite of the diagnostic discrepancy, there was substantial correlation in numbers of symptom groups in the 2 modalities (r = 0.49). Agreement was highest (about 55% sensitivity and 90% specificity) when both the SCAN and DIS thresholds were set at the level of depression syndrome instead of diagnosis. CONCLUSIONS: Weak agreement at the level of diagnosis continues to threaten the credibility of estimates of prevalence of specific disorders. A bias toward underreporting, as well as stronger agreement at the level of the depression syndrome and on ordinal measures of depressive symptoms, suggests that associations with risk factors are conservative. PMID- 10711907 TI - Community diagnosis counts. AB - Because of such validity-research deficits and the ceiling on agreement between instruments imposed by less-than-perfect reliability characteristics of each instrument, it is not appropriate to assume that the semistructured clinician interview is more valid than the epidemiologic interview. The Baltimore ECA site is uniquely situated to address this issue by comparing the outcome of subjects identified with current depression in the 1982 clinical reappraisal interview with those identified by the DIS at the same time to see if the 13-year follow-up is similar to that found over 16 years by Murphy et al. Where do we go from here in improving our diagnostic criteria for DSM-V, constructing better diagnostic instruments, and conducting the next generation of epidemiologic studies? Certainly the categorical diagnostic criteria themselves, without a dimensional symptom level, are never used in clinical treatment trials. Hence the "clinical significance" criteria of significant distress or disability added to DSM-IV should be further refined, with the possible addition of "staging" of disorders. The objective would be to provide a better indication of treatment need and clinical prognosis as in current cancer diagnostic assessments. For epidemiologic studies, the addition of symptom scales and disability assessments to the traditional categorical diagnoses should be helpful in developing community measures of treatment need. Different methods of assessment may be useful for diagnoses in which an impaired perception of reality occurs, such as schizophrenia. With some of these adjustments, it should be feasible to "count" those with clinically significant diagnoses in the community, and thus improve the validity and clinical utility of our diagnoses for predicting clinical course and responsiveness to specific treatments. PMID- 10711908 TI - Controversies in community-based psychiatric epidemiology: let the data speak for themselves. PMID- 10711909 TI - A comparison of diagnostic interviews for depression in the Stirling County study: challenges for psychiatric epidemiology. AB - BACKGROUND: High prevalence rates in psychiatric epidemiologic studies raise questions about whether data-gathering procedures identify transient responses rather than clinical disorders. This issue is explored relevant to depression using data from the Stirling County Study. METHODS: The study's customary method, the DPAX (DP for depression and AX for anxiety) was compared with the Diagnostic Interview Schedule (DIS), both of which were administered to a sample of 1396 subjects selected in 1992. Reasons for discordance were analyzed, and demographic correlates of responses to questions about dysphoria were examined. These lay administered interviews were then compared with clinician-administered interviews that used the Structured Clinical Interview for DSM-III-R (SCID) with 139 subjects. The kappa statistic and logistic regression were used for statistical assessment. RESULTS: For the level of agreement between the DPAX and the DIS for current and lifetime depression, kappa = 0.40 and kappa = 0.33, respectively. Subjects diagnosed only by the DPAX tended to have less education than those diagnosed only by the DIS. Some idioms for dysphoria seemed to work better than others. Using SCID interviews as a clinical standard, the DPAX had 15% sensitivity and 96% specificity and the DIS had 25% sensitivity and 98% specificity. CONCLUSIONS: Comprehension of an interview can be improved by using multiple questions for dysphoria and a simpler mode of inquiry. Clinician administered interviews tend to corroborate disorders identified in lay administered interviews but suggest that survey methods underestimate prevalence. Further research is needed to evaluate the validity of both types of interviews, but evidence from a 16-year follow-up evaluation indicates that depression diagnosed by the DPAX is a serious disorder in terms of morbidity and mortality. PMID- 10711910 TI - Decreased glutamic acid decarboxylase67 messenger RNA expression in a subset of prefrontal cortical gamma-aminobutyric acid neurons in subjects with schizophrenia. AB - BACKGROUND: Markers of gamma-aminobutyric acid (GABA) neurotransmission seem to be altered in the prefrontal cortex (PFC) of subjects with schizophrenia. We sought to determine whether the expression of the messenger RNA (mRNA) for the synthesizing enzyme of GABA, glutamic acid decarboxylase67 (GAD67), is decreased in the PFC of subjects with schizophrenia, whether this change is present in all or only some GABA neurons, and whether long-term treatment with haloperidol decanoate contributes to altered GAD67 mRNA expression. METHODS: Tissue sections from 10 pairs of subjects with schizophrenia and control subjects and 4 pairs of haloperidol-treated and control monkeys were processed for in situ hybridization histochemical analysis with sulfur-35-labeled oligonucleotide probes for GAD67 mRNA and exposed to nuclear emulsion. Within each layer of PFC area 9, neurons expressing a detectable level of GAD67 mRNA were quantified for cell density and the relative level of mRNA expression per cell (grain density per neuron). RESULTS: In subjects with schizophrenia, the density of labeled neurons was significantly (P<.05) decreased by 25% to 35% in cortical layers 3 to 5. In contrast, the mean grain density per labeled neuron did not differ across subject groups. Similar analyses in monkeys revealed no effect of long-term haloperidol treatment on either the density of the labeled neurons or the grain density per labeled neuron. CONCLUSIONS: These findings indicate that in subjects with schizophrenia, GAD67 mRNA expression is relatively unaltered in most PFC GABA neurons but is reduced below a detectable level in a subset of GABA neurons. Altered GABA neurotransmission in this subset may contribute to PFC dysfunction in subjects with schizophrenia. PMID- 10711911 TI - Neuropsychological change in early phase schizophrenia during 12 months of treatment with olanzapine, risperidone, or haloperidol. The Canadian Collaborative Group for research in schizophrenia. AB - BACKGROUND: The purpose of this investigation was to test the efficacy of novel antipsychotic medications in the treatment of cognitive impairment in early phase schizophrenia. METHODS: Sixty-five patients in this multicenter double-blind study were randomly assigned to olanzapine (5-20 mg), risperidone (4-10 mg), or haloperidol (5-20 mg). Standard measures of clinical and motor syndromes were administered, as well as a comprehensive battery of tests to assess (1) motor skills, (2) attention span, (3) verbal fluency and reasoning, (4) nonverbal fluency and construction, (5) executive skills, and (6) immediate recall at baseline and after 6, 30, and 54 weeks of treatment. RESULTS: The general cognitive index derived from the 6 domain scores revealed a significantly greater benefit from treatment with olanzapine relative to haloperidol and olanzapine relative to risperidone, but no significant difference was shown between risperidone and haloperidol. The improvement related to olanzapine was apparent after 6 weeks and enhanced after 30 and 54 weeks of treatment. Exploratory within group analyses of the 6 cognitive domains after a conservative Bonferroni adjustment revealed a significant improvement with olanzapine only on the immediate recall domain, and similar analyses of the 17 individual tests revealed a significant improvement with olanzapine only on the Hooper Visual Organization Test. CONCLUSIONS: These data suggest that olanzapine has some superior cognitive benefits relative to haloperidol and risperidone. A larger sample replication study is necessary to confirm and generalize the observations of this study and begin evaluation of the implications of this change to cerebral function and quality of life for people with schizophrenia. PMID- 10711912 TI - Illicit psychoactive substance use, heavy use, abuse, and dependence in a US population-based sample of male twins. AB - BACKGROUND: In order to develop informed approaches to prevention and treatment of illicit psychoactive substance use, abuse, and dependence, we need to understand the sources of individual differences in risk. METHODS: In personal interviews with 1198 male-male twin pairs (708 monozygotic and 490 dizygotic) ascertained from a population-based registry, we assessed lifetime use, heavy use, and abuse of and dependence on cannabis, sedatives, stimulants, cocaine, opiates, and hallucinogens. Twin resemblance was assessed by probandwise concordance, odds ratio, tetrachoric correlations, and biometrical model fitting. RESULTS: Twin resemblance for substance use, heavy use, abuse, and dependence was substantial, and consistently greater in monozygotic than in dizygotic twins. For any drug use and for cannabis and hallucinogen use, model fitting suggested that twin resemblance was due to both genetic and familial-environmental factors. Twin resemblance for sedative, stimulant, cocaine, and opiate use, however, was caused solely by genetic factors. With 2 exceptions (cocaine abuse and stimulant dependence), twin resemblance for heavy use, abuse, and dependence resulted from only genetic factors, with heritability of liability usually ranging from 60% to 80%. No consistent evidence was found for violations of the equal environment assumption. CONCLUSIONS: In accord with prior results in studies of women, the family environment plays a role in twin resemblance for some forms of substance use in men. However, twin resemblance for heavy use, abuse, and dependence in men is largely caused by genetic factors, and heritability estimates are high. PMID- 10711913 TI - Attenuation of the neuropsychiatric effects of ketamine with lamotrigine: support for hyperglutamatergic effects of N-methyl-D-aspartate receptor antagonists. AB - BACKGROUND: The cognitive, behavioral, and mood effects of N-methyl-D-aspartate (NMDA) receptor antagonists, such as phencyclidine and ketamine, have been used to study the effects of NMDA receptor dysfunction. Pharmacological modulation of the effects of NMDA receptor antagonists, such as ketamine, may lead to development of novel therapeutic agents for psychiatric illnesses such as schizophrenia. Preclinical studies indicate that some ketamine effects may be mediated through increased glutamate release. In this study, we tested the hypothesis that lamotrigine, a drug reported to inhibit glutamate release, will reduce the neuropsychiatric effects of ketamine in humans. METHOD: Healthy subjects (n = 16) completed 4 test days involving the administration of lamotrigine, 300 mg by mouth, or placebo 2 hours prior to administration of ketamine (0.26 mg/kg by intravenous bolus and 0.65 mg/kg per hour by intravenous infusion) or placebo in a randomized order under double-blind conditions. Behavioral and cognitive assessments were performed at baseline and after administration of the medications. RESULTS: Lamotrigine significantly decreased ketamine-induced perceptual abnormalities as assessed by the Clinician Administered Dissociative States Scale (P<.001); positive symptoms of schizophrenia as assessed by the Brief Psychiatric Rating Scale positive symptoms subscale (P<.001); negative symptoms as assessed by the Brief Psychiatric Rating Scale negative symptoms subscale (P<.05); and learning and memory impairment as assessed by the Hopkins Verbal Learning Test (P<.05). However, lamotrigine increased the immediate mood-elevating effects of ketamine (P<.05). CONCLUSIONS: Glutamate release-inhibiting drugs may reduce the hyperglutamatergic consequences of NMDA receptor dysfunction implicated in the pathophysiologic processes of neuropsychiatric illnesses such as schizophrenia. Further study is needed. PMID- 10711914 TI - A program for relapse prevention in schizophrenia: a controlled study. AB - BACKGROUND: This study examined whether a program for relapse prevention (PRP) is more effective than treatment as usual (TAU) in reducing relapse and rehospitalization rates among outpatients with schizophrenia. METHODS: Eighty-two outpatients with DSM-III-R schizophrenia or schizoaffective disorder were randomly assigned to receive either PRP (experimental group, n = 41) or TAU (control group, n = 41) and were followed up for an 18-month prospective controlled study. Patients in both groups were prescribed standard doses of maintenance antipsychotic medication. Treatment with PRP consisted of a combination of psychoeducation, active monitoring for prodromal symptoms with clinical intervention when such symptoms occurred, weekly group therapy for patients, and multifamily groups. The TAU consisted of biweekly individual supportive therapy and medication management. RESULTS: Outcome rates over 18 months were 17% for relapse (7 patients) and 22% for rehospitalization (9 patients) in the PRP group, compared with 34% for relapse (14 patients) and 39% for rehospitalization (16 patients) in the TAU group (P = .01 and P = .03, respectively). Addition of age, sex, baseline Global Assessment Scale score, Positive and Negative Syndrome Scale scores (3 measures), and substance abuse to the proportional hazards regression models all yielded nonsignificant effects. The PRP teams were much more likely than the TAU psychiatrists to identify prodromal episodes before patients met objective relapse criteria or needed hospitalization. CONCLUSIONS: The PRP was effective in detecting prodromal symptoms of relapse early in an episode. Crisis intervention including increased antipsychotic medication use during the prodromal phase reduced relapse and rehospitalization rates. PMID- 10711915 TI - Executive dysfunction and long-term outcomes of geriatric depression. AB - BACKGROUND: This study investigated the relationship of executive and memory impairment to relapse, recurrence, and course of residual depressive symptoms and signs after remission of geriatric major depression. METHODS: Fifty-eight elderly subjects remitted from major depression received continuation nortriptyline treatment (plasma levels 60-150 ng/mL) for 16 weeks and then were randomly assigned to either nortriptyline maintenance therapy or placebo for up to 2 years. Diagnosis was made using the Research Diagnostic Criteria and the DSM-IV criteria after an interview using the Schedule for Affective Disorders and Schizophrenia. Executive dysfunction and memory were assessed with the Dementia Rating Scale, disability and social support were rated with the Philadelphia Multiphasic Instrument, and medical burden was assessed with the Cumulative Illness Rating Scale. RESULTS: Abnormal initiation and perseveration scores, but not memory impairment, were associated with relapse and recurrence of geriatric depression and with fluctuations of depressive symptoms in the whole group and in subjects who never met criteria for relapse or recurrence during the follow-up period. Memory impairment, disability, medical burden, social support, and history of previous episodes did not significantly influence the outcome of depression in this sample. CONCLUSIONS: Executive dysfunction was found to be associated with relapse and recurrence of geriatric major depression and with residual depressive symptoms. These observations, if confirmed, will aid clinicians in identifying patients in need of vigilant follow-up. The findings of this study provide the rationale for investigation of the role of specific prefrontal pathways in predisposing or perpetuating depressive syndromes or symptoms in elderly patients. PMID- 10711916 TI - Dextromethorphan challenge in alcohol-dependent patients and controls. PMID- 10711917 TI - Altruism and confidentiality in organ donation. PMID- 10711918 TI - Eradication of Helicobacter pylori and non-ulcer dyspepsia. PMID- 10711919 TI - Early promise of stapling technique for haemorrhoidectomy. PMID- 10711920 TI - Treatment of chronic bulimic symptoms: new answers, more questions. PMID- 10711921 TI - Renal conservation for gas-forming infections. PMID- 10711922 TI - P300, a state and a trait marker in schizophrenia. PMID- 10711923 TI - Stress hyperglycaemia and increased risk of death after myocardial infarction in patients with and without diabetes: a systematic overview. AB - BACKGROUND: High blood glucose concentration may increase risk of death and poor outcome after acute myocardial infarction. We did a systematic review and meta analysis to assess the risk of in-hospital mortality or congestive heart failure after myocardial infarction in patients with and without diabetes who had stress hyperglycaemia on admission. METHODS: We did two searches of MEDLINE for English language articles published from 1966 to October, 1998, a computerised search of Science Citation Index from 1980 to September, 1998, and manual searches of bibliographies. Two searchers identified all cohort studies or clinical trials reporting in-hospital mortality or rates of congestive heart failure after myocardial infarction in relation to glucose concentration on admission. We compared the relative risks of in-hospital mortality and congestive heart failure in hyperglycaemic and normoglycaemic patients with and without diabetes. FINDINGS: 14 articles describing 15 studies were identified. Patients without diabetes who had glucose concentrations more than or equal to range 6.1-8.0 mmol/L had a 3.9-fold (95% CI 2.9-5.4) higher risk of death than patients without diabetes who had lower glucose concentrations. Glucose concentrations higher than values in the range of 8.0-10.0 mmol/L on admission were associated with increased risk of congestive heart failure or cardiogenic shock in patients without diabetes. In patients with diabetes who had glucose concentrations more than or equal to range 10.0-11.0 mmol/L the risk of death was moderately increased (relative risk 1.7 [1.2-2.4]). INTERPRETATION: Stress hyperglycaemia with myocardial infarction is associated with an increased risk of in-hospital mortality in patients with and without diabetes; the risk of congestive heart failure or cardiogenic shock is also increased in patients without diabetes. PMID- 10711924 TI - Circumferential mucosectomy (stapled haemorrhoidectomy) versus conventional haemorrhoidectomy: randomised controlled trial. AB - BACKGROUND: Haemorrhoidectomy is commonly an inpatient procedure because it is frequently associated with postoperative pain. Day case haemorrhoidectomy is a similar operation to that used on inpatients but with different strategies for managing postoperative pain. Circumferential mucosectomy (stapled haemorrhoidectomy) may be associated with less postoperative pain than conventional haemorrhoidectomy. We compared stapled haemorrhoidectomy with conventional haemorrhoidectomy in patients with third degree haemorrhoids. METHODS: We randomly assigned 22 patients to conventional haemorrhoidectomy by the diathermy dissection or to stapled haemorrhoidectomy with the use of an intraluminal stapling device. Patients were discharged when free of pain, took co codamol as required, completed visual analogue charts each day, and were assessed at 1 and 6 weeks postoperatively for symptom control. FINDINGS: All patients received the assigned treatment. Mean inpatient stay was lower in the group assigned to stapled as opposed to conventional haemorrhoidectomy (1.09 [0.3] vs 2.82 [0.09] nights, p<0.001), experienced less pain overall (p=0.003), and returned to normal activities sooner (8.1 [1.53] vs 16.9 [2.33] days, p<0.005). Stapled haemorrhoidectomy controlled symptoms of prolapse, discharge and bleeding in all patients. INTERPRETATION: Stapled haemorrhoidectomy is an effective treatment for third degree haemorrhoids with significant advantages for patients compared with conventional haemorrhoidectomy. PMID- 10711925 TI - Stapling procedure for haemorrhoids versus Milligan-Morgan haemorrhoidectomy: randomised controlled trial. AB - BACKGROUND: Surgical haemorrhoidectomy has a reputation for being a painful procedure for a fairly benign disorder. The circular transanal stapled technique for the treatment of haemorrhoids has the potential to offer a less painful rectal procedure in place of ablative perianal surgery. We compared the short term outcome of the circular stapled procedure for haemorrhoids with current standard surgery in a randomised controlled trial. METHODS: 40 patients admitted for surgical treatment of prolapsing haemorrhoids were randomly assigned to Milligan-Morgan haemorrhoidectomy (n=20) or the circular stapled procedure. Under general anaesthesia patients underwent standardised diathermy excision haemorrhoidectomy or had a circumferential doughnut of rectal mucosa and submucosa above the dentate line excised and closed with a standard circular end to-end stapling device. All patients received standardised preoperative and postoperative analgesic and laxative regimens. Patients completed linear analogue pain charts each day and were interviewed at 1, 3, and 6-10 weeks postoperatively. Summary measures of average pain experience were calculated from 10 cm linear analogue pain scores and were used as the primary outcome measure. FINDINGS: The stapled group had shorter anaesthesia time (median 18 [range 9-25] vs 22 [15-35] mins). Average pain in the stapled group was significantly lower than it was in the Milligan-Morgan group (2.1 [0.2-7.6] vs 6.5 [3.1-8.5], 95.1% CI difference medians 1.9-4.7, p<0.0001. Mann-Whitney U test). Average pain relative to what the patient expected was also significantly less in the stapled group (-2.8 [-4.4 to 1.3] vs 0.7 [-1.8 to 3.4]. Hospital stay and time to first bowel motion were not significantly different between groups. Return to normal activity was significantly shorter in the stapled group (17 [3-60] vs 34 [14-90]. Early and late complications, patient-assessed symptom control, and functional outcome appear similar after short-term follow-up. INTERPRETATION: The circular stapled technique offers a significantly less painful alternative to Milligan Morgan haemorrhoidectomy and is associated with an earlier return to normal activity. Early symptom control and functional outcome appear similar. However, long-term symptomatic and functional outcome need further study. PMID- 10711926 TI - Thalassaemia in Sri Lanka: implications for the future health burden of Asian populations. Sri Lanka Thalassaemia Study Group. AB - BACKGROUND: Thalassaemias pose an increasing problem for the Indian subcontinent and many Asian countries. We analysed the different types of thalassaemia in the Sri Lankan population, surveyed gene frequencies in schoolchildren, and estimated the burden of disease and requirements for its control. METHODS: We analysed blood samples from patients attending clinics in nine hospitals and defined the different types of beta thalassaemia by high-performance liquid chromatography (HPLC) and DNA analysis. The range of mutations was obtained by analysis of beta globin genes. Capillary blood was obtained from schoolchildren from different parts of the island and analysed by HPLC to provide an approximate assessment of the carrier frequency of beta thalassaemia and haemoglobin E (HbE). To estimate the frequency of alpha thalassaemia the alpha-globin genotypes were also analysed when it was possible. FINDINGS: Blood samples were obtained from 703 patients with beta thalassaemia and from 1600 schoolchildren. The thalassaemia mutations were unevenly spread. Although 23 different beta-thalassaemia mutations were found, three accounted for the thalassaemia phenotype in about 70% of the patients, most whom are homozygotes or compound heterozygotes for IVS1-5 (G-->C) or IVS1-1 (G-->A). The third common mutation, codon 26 (G-->A), which produces HbE, interacts with one or other of these mutations to produce HbE/beta thalassaemia; this comprises 13.0-30.9% of cases in the main centres. Samples from 472 patients were analysed to determine the alpha-globin genotype. Overall, 15.5% patients were carriers for deletion forms of alpha+ thalassaemia. Average gene frequencies showed that there will be more than 2000 patients requiring treatment at any one time, in the future, of whom those with HbE/beta thalassaemia will account for about 40%. INTERPRETATION: In Sri Lanka, interactions of the two common beta-thalassaemia alleles will nearly always result in a transfusion-dependent disorder. However, about 40% of patients will have HbE/beta thalassaemia, which has a variable course. The management of these disorders could require about 5% of the total health budget. We need to learn more about the natural history and appropriate management of HbE/beta thalassaemia if resources are to be used effectively. PMID- 10711927 TI - Effect of decreasing afferent vagal activity with ondansetron on symptoms of bulimia nervosa: a randomised, double-blind trial. AB - BACKGROUND: Several lines of evidence have led us to postulate that afferent vagal hyperactivity could be an important factor in the pathophysiology of the eating disorder bulimia nervosa. Ondansetron is a peripherally active antagonist of the serotonin receptor 5-HT3, and is marketed for prevention of vagally mediated emesis caused by cancer chemotherapeutic agents. We investigated the effects of ondansetron on bulimic behaviours in patients with severe and chronic bulimia nervosa in a randomised, double-blind, placebo-controlled study. METHODS: We enrolled patients with severe bulimia nervosa (at least seven coupled binge/vomit episodes per week). The patients were otherwise healthy, their weight was normal, and they were not receiving medical or psychiatric treatment. During the first week of the study, patients recorded all eating-behaviour events to establish a baseline. In the second week, all patients received placebo, but were told that they were receiving either placebo or active drug. At the end of this single-blind phase, patients were randomly assigned placebo or ondansetron (24 mg daily) for a further 4 weeks. The primary outcome measure was the number of binge/vomit episodes per week. Data were analysed by intention to treat. FINDINGS: 29 patients met the inclusion criteria, of whom 28 completed the baseline study, and 26 completed the single-blind placebo week. 12 patients were assigned placebo, and 14 ondansetron; one patient in the ondansetron group dropped out owing to accidental injury. During the 4th week of double-blind treatment, mean binge/vomit frequencies were 13.2 per week (SD 11.6) in the placebo group, versus 6.5 per week (3.9) in the ondansetron group (estimated difference 6.8 [95% CI 4.0-9.5]; p<0.0001). The ondansetron group also showed significant improvement, compared with the placebo group, in two secondary indicators of disease severity. The amount of time spent engaging in bulimic behaviours was decreased on average by 7.6 h per week in the ondansetron group, compared with 2.3 h in the placebo group (estimated difference 5.1 [0.6-9.7]). Similarly, the number of normal meals and snacks increased on average by 4.3 normal eating episodes without vomiting per week in the ondansetron group, compared with 0.2 in the placebo group (estimated difference 4.1 [1.0-7.2]). INTERPRETATION: The decrease in binge-eating and vomiting under ondansetron treatment was not achieved by compensatory changes in eating behaviour such as by a smaller number of binges of longer duration, or by not eating, or by binge eating without vomiting. Instead, our findings indicate a normalisation of the physiological mechanism(s) controlling meal termination and satiation. Since meal termination and satiety are mainly vagally mediated functions, since binge-eating and vomiting produce intense stimulation of vagal afferent fibres, and since ondansetron and other 5-HT3 antagonists decrease afferent vagal activity, the symptom improvement may result from a pharmacological correction of abnormal vagal neurotransmission. PMID- 10711928 TI - Response to different measles vaccine strains given by aerosol and subcutaneous routes to schoolchildren: a randomised trial. AB - BACKGROUND: More than one dose of measles vaccine is necessary for the sustained control of measles. The aerosol route is thought to be more immunogenic for booster doses than traditional subcutaneous injections, so we did a randomised comparative trial of aerosol and subcutaneous measles vaccines in South African schoolchildren. METHODS: 4327 schoolchildren (aged 5-14 years), assigned by block randomisation of classrooms, received standard titre doses of either Schwarz or Edmonston-Zagreb measles vaccines subcutaneously or by aerosol. Blood samples for antibody assay were collected before vaccination, at 1 month, and 1 year after vaccination. The main endpoints (antibody titres at 1 month and 1 year) were compared between groups. FINDINGS: 992 children had antibody titre data available for all timepoints. 14 (3.6%) of 385 children who received Edmonston-Zagreb vaccine by aerosol were seronegative 1 year after vaccination, compared with 28 (8.6%) of 326 children who received Edmonston-Zagreb subcutaneous vaccine and 39 (13.9%) of 281 children who received Schwarz subcutaneous vaccine. At 1 month, 326 (84.7%) children who received aerosol Edmonston-Zagreb vaccine had seroconverted, compared with 257 (78.8%) who received subcutaneous Edmonston Zagreb vaccine and 176 (62.6%) who received subcutaneous Schwarz vaccine. At 1 month, only 116 (22.7%) of 511 children in the Schwarz aerosol group had seroconverted; this aerosol vaccine had no detectable potency after 2 min of nebulisation. There were no serious side-effects: about 5% of children in each group had a rash within 2 weeks of vaccination. INTERPRETATION: An aerosol vaccination method that uses currently available devices and a suitably stable vaccine is effective and acceptable. This form of delivery is adaptable to mass campaigns, avoids the risks associated with injections, and could help measles eradication. PMID- 10711929 TI - A coughing policeman. PMID- 10711930 TI - QT dispersion as a predictor of acute heart failure after high-dose cyclophosphamide. AB - No useful predictor of risk of acute heart failure in peripheral-blood stem-cell transplantation (PBSCT) regimens, Including high-dose cyclophosphamide, has previously been available. Corrected QT dispersion can predict acute heart failure after high-dose cyclophosphamide chemotherapy used in PBSCT. PMID- 10711931 TI - Severe osteopenia in symptom-free adults with a childhood diagnosis of coeliac disease. AB - The frequency of osteopenia in symptom-free adults diagnosed with coeliac disease during childhood and who resumed a normal diet during adolescence is unknown. Severe osteopenia (a bone mineral density below two standard deviations of the mean) was found in up to a third of symptom-free young adults on a normal diet. These patients should not be thought to be disease-free but should receive long term follow-up and most of them should be advised to resume a gluten-free diet. PMID- 10711932 TI - Pulse-inversion mode imaging of liver specific microbubbles: improved detection of subcentimetre metastases. AB - Pulse-inversion mode (a new ultrasound mode) can be used to image the late liver specific parenchymal phase of the microbubble contrast-agent Levovist. Scanning in pulse-inversion mode after Levovist improves the detection of liver metastases and reveals more lesions of smaller size than conventional ultrasonography and computed tomography. PMID- 10711933 TI - Brugada syndrome and sudden cardiac death in children. AB - In five children from the same family who died after unexplained cardiac arrest, Brugada syndrome syndrome was suspected based on the transient manifestation of the typical electrocardiogram pattern in one of them. A mutation in the cardiac sodium-channel confirmed the diagnosis of Brugada syndrome, which suggests that this disease may cause sudden death in children. PMID- 10711934 TI - Life threatening pelvic sepsis after stapled haemorrhoidectomy. AB - We describe a case of severe retroperitoneal sepsis after stapled haemorrhoidectomy. The operation seemed to be technically satisfactory, and we suggest that routine antibiotic prophylaxis may be indicated with this procedure. PMID- 10711935 TI - So just how did life start on Earth? PMID- 10711936 TI - A new National Health Service revolution causes trouble. PMID- 10711937 TI - No clear victor in battle between fertility expert and UK university. PMID- 10711938 TI - Newspaper apportions blame in Spanish hepatitis C scandal. PMID- 10711939 TI - Scabies and pediculosis. AB - Scabies and pediculosis are ubiquitous, contagious, and debilitating parasitic dermatoses. They have been known since antiquity and are distributed worldwide. Clusters of infestation occur-for example, scabies affecting immunocompromised individuals or patients and staff in hospitals and nursing homes for the elderly, and pediculosis affecting schoolchildren or homeless people. Associations with other disorders are common: infections with human T-cell leukaemia/lymphoma virus I (HTLV-I) and HIV are associated with scabies, and trench fever and exanthematous typhus with pediculosis. Specific forms of scabies, including bullous scabies or localised crusted scabies, may be misdiagnosed. Moreover, definitive parasitic diagnosis can be difficult to obtain, and the value of new techniques remains to be confirmed. Difficulties in management have returned scabies and pediculosis to the limelight. PMID- 10711940 TI - Influenza virus neuraminidase inhibitors. AB - Neuraminidase promotes influenza virus release from infected cells and facilitates virus spread within the respiratory tract. Several potent and specific inhibitors of this enzyme have been developed, and two (zanamivir and oseltamivir) have been approved for human use. Unlike amantadine and rimantadine that target the M2 protein of influenza A viruses, these drugs inhibit replication of both influenza A and B viruses. Zanamivir is delivered by inhalation because of its low oral bioavailability whereas oseltamivir is administered by mouth. Early treatment with either drug reduces the severity and duration of influenza symptoms and associated complications. Both agents are effective for chemoprophylaxis. Because of a broader antiviral spectrum, better tolerance, and less potential for emergence of resistance than is seen with the M2 inhibitors, the neuraminidase inhibitors represent an important advance in the treatment of influenza. PMID- 10711941 TI - Need to focus research in stroke rehabilitation. PMID- 10711942 TI - Will South African physicians build a culture of human rights? PMID- 10711943 TI - Paediatrician Flora Brovina remains imprisoned in Kosovo. PMID- 10711944 TI - Death penalty suspended in Illinois. PMID- 10711945 TI - Clozapine and sudden death. PMID- 10711946 TI - Clozapine and sudden death. PMID- 10711947 TI - Clozapine and sudden death. PMID- 10711948 TI - Clozapine and sudden death. PMID- 10711949 TI - Parkinson's disease and motor-neuron disease in former prisoners-of-war. PMID- 10711950 TI - Enforced bedrest. PMID- 10711951 TI - Swedish in-vitro fertilisation study. PMID- 10711952 TI - Swedish in-vitro fertilisation study. PMID- 10711953 TI - Swedish in-vitro fertilisation study. PMID- 10711954 TI - Swedish in-vitro fertilisation study. PMID- 10711955 TI - Role of orexins in sleep and arousal mechanisms. PMID- 10711956 TI - Fighting spirit in patients with cancer. PMID- 10711957 TI - Occurrence of stroke with tamoxifen in NSABP B-24. PMID- 10711958 TI - Dubious benefits and potential risk of soy phyto-oestrogens. PMID- 10711959 TI - Publishing surgery. PMID- 10711960 TI - A data-donor scheme for brain researchers. PMID- 10711961 TI - Allergy to latex. PMID- 10711962 TI - Philoctetes' foot. PMID- 10711963 TI - Biomaterials used in the posterior segment of the eye. AB - The treatment of posterior segment eye disease and related conditions has improved greatly in recent years with the advent of new therapies, materials and devices. Vitreoretinal conditions, however, remain significant causes of blindness in the developed world. Biomaterials play a major role in the treatment of many of these disorders and the success rate of vitreoretinal surgery, especially in the repair of retinal detachment and related conditions, would increase with the introduction of new and improved materials. This review, which focuses on disorders that feature retinal detachment, briefly describes the anatomy of the eye and the nature and treatment of posterior segment eye disorders. The roles, required properties and suitability of the materials used in vitreoretinal surgery as scleral buckles, tamponade agents or drug delivery devices, are reviewed. Experimental approaches are discussed, along with the methods used for their evaluation, and future directions for biomaterial research in the posterior segment of the eye are considered. PMID- 10711964 TI - Osteoblast adhesion on biomaterials. AB - The development of tissue engineering in the field of orthopaedic surgery is now booming. Two fields of research in particular are emerging: the association of osteo-inductive factors with implantable materials; and the association of osteogenic stem cells with these materials (hybrid materials). In both cases, an understanding of the phenomena of cell adhesion and, in particular, understanding of the proteins involved in osteoblast adhesion on contact with the materials is of crucial importance. The proteins involved in osteoblast adhesion are described in this review (extracellular matrix proteins, cytoskeletal proteins, integrins, cadherins, etc.). During osteoblast/material interactions, their expression is modified according to the surface characteristics of materials. Their involvement in osteoblastic response to mechanical stimulation highlights the significance of taking them into consideration during development of future biomaterials. Finally, an understanding of the proteins involved in osteoblast adhesion opens up new possibilities for the grafting of these proteins (or synthesized peptide) onto vector materials, to increase their in vivo bioactivity or to promote cell integration within the vector material during the development of hybrid materials. PMID- 10711965 TI - Platelet adhesion onto segmented polyurethane film surfaces modified by addition and crosslinking of PEO-containing block copolymers. AB - Polyethylene oxide (PEO) surfaces were prepared by the addition of PEO-containing amphiphilic block copolymers as surface modifying additives and of dicumyl peroxide (DCP) as a crosslinking agent in segmented polyurethane (PU). PEO polypropylene oxide-PEO triblock copolymers (Pluronics) with different PEO chain length (from 0 to 98) were used as the surface modifying additives. The PEO additives in the PU film were then crosslinked to be stably entrapped in the PU matrix. The crosslinking was done by free radicals produced from the decomposition of DCP in the film through heating (120 degrees C) or ultraviolet irradiation (254 nm). The surface properties of the PEO additive-entrapped PU films were investigated by the measurement of water contact angles and electron spectroscopy for chemical analysis. The bulk properties such as water absorption, long-term film stability, and tensile strength and elongation at break, were also investigated. It was observed that addition of a small amount (5 wt% based on PU) of the PEO additives resulted in a considerable change of surface characteristics. The PEO additives were stably entrapped in the PU films by crosslinking of them, without significant changes of bulk properties of the films. From the platelet adhesion test on the prepared PEO additive-containing film surfaces, it was observed that the platelet adhesion on the surfaces decreases with increase in PEO chain length of PEO additives. The film surface containing additive with long PEO chains (chain length of 98) was particularly effective in preventing platelet adhesion. The crosslinking of the PEO additives in PU films did not affect the behavior of platelet adhesion on the surfaces; the films with crosslinked PEO additives showed similar platelet adhesion on the surfaces to the films with uncrosslinked ones. PMID- 10711966 TI - Influence of glass composition on the properties of glass polyalkenoate cements. Part IV: influence of fluorine content. AB - The influence of fluorine content of the glass in a series of glasses based on: 4.5SiO2-3.0Al2O3-1.5P2O5-(5.0-X)CaO-XCaF2 was investigated on their cement formation with poly(acrylic acid). Increasing the fluorine content of the glass was found to reduce the glass transition temperature, as a result of fluorine replacing bridging oxygens by non-bridging fluorines. Working and setting times of the cement pastes reduced with increasing fluorine content of the glass. Compressive strength and Young's moduli increased with fluorine content initially and then remained approximately constant. Fracture toughness, toughness and un notched fracture strength were not significantly influenced by fluorine content. The results are consistent with fluorine simultaneously disrupting the glass network and reducing the basicity of the glass. PMID- 10711967 TI - Use of plasma glow for surface-engineering biomolecules to enhance bloodcompatibility of Dacron and PTFE vascular prosthesis. AB - The search for a nonthrombogenic material having patency to be used for small diameter vascular graft applications continues to be a field of extensive investigation. The purpose of the present study was to examine whether surface modification of polytetra fluoroethylene (PTFE, Teflon) and polyethylene terephthalate (Dacron) vascular grafts might extend graft biocompatibility without modifying the graft structure. A series of surface coatings were prepared by modifying the argon plasma-treated PTFE and Dacron grafts with collagen IV and laminin and subsequently immobilizing bioactive molecules like PGE1, heparin or phosphatidyl choline via the carbodiimide functionalities. Surface analysis by Fourier transform infrared spectroscopy-attenuated total reflectance revealed the presence of new functional groups on the modified graft surfaces. In vitro studies showed that fibrinogen adsorption and platelet adhesion on modified grafts were significantly reduced. This study proposes that surface grafting of matrix components (collagen-type IV and laminin) and subsequent immobilization of bioactive molecules (PGE1, heparin or phosphatidyl choline) changed the surface conditioning of vascular grafts and subsequently improved their biocompatibility. However, more detailed in vivo studies are needed to confirm these observations. PMID- 10711968 TI - Physical and biocompatibility properties of poly-epsilon-caprolactone produced using in situ polymerisation: a novel manufacturing technique for long-fibre composite materials. AB - Preliminary investigations into a novel process for the production of poly epsilon-caprolactone (PCL) to be used as a matrix material in a bioabsorbable composite material are detailed. This material is primarily being developed as a bone substitute for use in maxillofacial reconstructive surgery, however, the technique described could be adapted to other areas where bioabsorbable composite materials may be used. The development of a totally bioabsorbable long-fibre composite material would allow a two-stage degradation to occur with the matrix material degrading first leaving a scaffold structure of degradable fibres which would be absorbed at a later stage. Caprolactone monomer was polymerised in situ within a tool cavity to produce a net shape moulding. Inclusion of a fibre preform within the tool cavity which was impregnated by the liquid monomer produces a long-fibre composite material. PCL with a range of molecular weights has been produced using this liquid moulding technique to assess the physical and biocompatibility properties compared to commercially available PCL. Osteoblast like cells derived from human craniofacial bone (CFC) have been used to assess the in vitro biocompatibility of the PCL. The results show that high-quality PCL with a narrow molecular weight distribution and properties similar to commercially available PCL can be produced using this technique. Polymerisation of the monomer around a woven fibre preform made of a poly(lactic acid) (PLA)/poly(glycolic acid) (PGA) copolymer (vicryl mesh) produced a bioabsorbable long-fibre composite material. Further work is ongoing to develop this system towards a method for improving craniofacial bone reconstruction. PMID- 10711969 TI - Hydrophilic, semipermeable membranes fabricated with poly(ethylene oxide) polysulfone block copolymer. AB - Semipermeable membranes may be fabricated from mixtures of poly(ethylene oxide)/polysulfone block copolymer (PEO-b-PSF) and polysulfone. Membranes fabricated with PEO-b-PSF possess a hydrophilic surface. PEO-b-PSF segregates to the membrane surface during phase inversion fabrication of the membrane rendering the surface hydrophilic. Changes in surface hydrophilicity were demonstrated by a dramatic reduction in the dynamic contact angle in water. With regard to the similar microporous hollow fiber membranes, a PEO-b-PSF membrane had a dynamic water contact angle of 33 degrees +/- 2 compared to a 111 degrees +/- 3 for a polysulfone membrane. Studies on porcine platelet-rich plasma in vitro demonstrated that the hydrophilic PEO-b-PSF membrane was resistant to platelet adhesion compared to a polysulfone membrane. An order of magnitude fewer adherent platelets were observed on a PEO-b-PSF membrane compared to a polysulfone membrane. The hydrophilicity of PEO-b-PSF makes it a unique material for the fabrication of membranes for medical devices. PMID- 10711970 TI - Synthesis and properties of amphiphilic networks. 1: the effect of hydration and polymer composition on the adhesion of immunoglobulin-G to poly(laurylmethacrylate-stat-glycerolmonomethacrylate-stat-ethylene-gly col dimethacrylate) networks. AB - A series of hydrogels composed of varying fractions of dodecyl methacrylate (DM) and 2,3-dihydroxypropyl methacrylate (GM) were prepared using ethylene glycol dimethacrylate (EGDMA) as the cross-linking agent. The study found that for a series of gels with the same monomer ratio, bulk hydration could be controlled by adjusting the cross-link density. The ability to control cross-link density allowed the preparation of gels with the same bulk hydration but different ratios of the two monomers. The adsorption of IgG to the gels was investigated using ELISA. The aim of the project was to investigate the effect of the bulk hydration and polymer composition on IgG adsorption. The results show that for a series of gels with the same monomer ratio, there is a clear trend towards a reduction in protein adsorption as the bulk hydration and accompanying chain mobility of the gel increases. Studies on gels of the same bulk hydration but differing ratios of monomer show higher protein adsorption as the proportion of GM increases. PMID- 10711971 TI - Incorporation of morsellized bone graft under controlled loading conditions. A new animal model in the goat. AB - The aim of this study was to develop a new animal model in which we could assess the in vivo effects of mechanical stimuli in the incorporation process of impacted morsellized bone grafts. The subcutaneous pressure implant SPI was developed for use in the goat. This device can generate controlled loading conditions onto a fixed amount of bone graft in the distal femur. Twenty goats were divided into three groups: non-loaded, 2 or 4 MPa loads (1 Hz, 1 h/day). The goats were sacrificed after 3, 6 or 12 weeks. The results were documented by clinical observations, quantitative bone density from QCT-scanning and histomorphometry. Nine post-mortem knee specimens were prepared in a similar manner to the experimental knees to determine the reproducibility and mechanical stability of the grafting method. Three goats were lost due to complications, the others functioned clinically well. Histology showed invasion of the bone graft by a front of vascular fibrous tissue after which osteoclasts resorbed the dead bone graft, followed by woven bone apposition on the graft remnants. At 12 weeks the loaded grafts had transformed into a vital trabecular structure. QCT bone density measurements revealed persistently high densities in the 12-weeks 4 MPa specimens, but reduced densities in the 2 MPa and non-loaded specimens. Morphometrically, the mineralising surface was larger in the 4 MPa group (P = 0.02) and the incorporation and remodelling processes had advanced more rapidly in the 2 MPa specimens (P = 0.04). Although the numbers investigated in this study in each group were low, statistical differences were found in the amount of graft left after incorporation and in the apposition rate of the new bone. In the future this model will be used to study the incorporation potential of different types of bone graft and bone graft substitutes. PMID- 10711972 TI - Microstructural dependence of Young's and shear moduli of P2O5 glass reinforced hydroxyapatite for biomedical applications. AB - P2O5 glass reinforced hydroxyapatite composite materials were prepared through a liquid-phase sintering process. Secondary phases, beta- and alpha-tricalcium phosphates (beta-TCP and alpha-TCP), were formed in the microstructure of the composites, due to the reaction between the liquid glassy phase and the hydroxyapatite matrix. The dynamic Young's modulus (E) and shear modulus (G) of these composites were determined using an impulse excitation method. By applying the Duckworth-Knudsen equation, the elastic property results were correlated with the relative proportion of beta-TCP and alpha-TCP phases and with the porosity percentage present in the microstructure. Glass reinforced hydroxyapatite composites showed lower Young's and shear moduli than unmodified hydroxyapatite, mainly due to the presence of beta-TCP phase. The Duckworth-Knudsen model demonstrated an exponential dependence of E and G modulus with porosity and mathematical equations were derived for composite materials with porosity correction factors (b) of 4.04 and 4.11, respectively, indicating that porosity largely decreased both E and G moduli. PMID- 10711973 TI - Three evaluations of task-surface heights in elderly people's homes. AB - The aim of the study was to compare subjective evaluations of the heights of some kitchen facilities and furniture with an expert's opinion of the same heights and with the height recommendations derived from the subjects' anthropometric data. Experiments were conducted in a mock-up simulator, where the subjects, a group of the Finnish elderly (N = 55), performed small tasks similar to the typical daily living activities at home. The mock-up room consisted of "task-surfaces" whose heights were adjustable to various fixed positions. For example, three identical chairs with different heights were available. In their subjective assessments, the elderly systematically rated the 450 mm-high chair as the most suitable. The lowest (350 mm) and the highest (550 mm) chairs were not liked. The other two evaluations supported this conclusion. The height of the lowest kitchen shelf should not be lower than 300 cm. A work surface height of 850 mm seems preferable for most of the Finnish elderly. The different evaluation procedures gave relatively consistent results, but some important differences were also noticed and are discussed. PMID- 10711974 TI - Ergonomics investigation of retail ice cream operations. AB - A comprehensive ergonomics evaluation of retail ice cream shops, including field and laboratory data collection, was conducted using a human:workplace model approach to ergonomics practice. The goal of the evaluation was to provide recommendations to enhance the health, safety, and productivity of shop employees. Active and passive surveillance and facility walk-throughs were used to guide the selection of analyses. A primary focus of the investigation was quantifying the task demands of scooping ice cream, which have not been documented in the literature. This goal was accomplished through the use of a custom-designed instrumented ice cream scoop. Data were collected at an ice cream shop under typical conditions, while the laboratory experiment investigated task demands of ice cream scooping over a range of realistic temperatures. Manual materials handling task analyses and anthropometric evaluations comprised the majority of other analyses performed. Recommendations are presented that are applicable to the operation of retail ice cream shops that serve hard (i.e., scooped) ice cream. PMID- 10711975 TI - Prospective memory: a secondary task with promise. AB - This paper examines the utility of prospective memory as a secondary task. For this purpose, the results of seven experiments were analysed to evaluate the sensitivity of prospective memory, making a direct comparison with another secondary task (reaction time). Both tasks were used as subsidiary elements in a complex task environment, the design of which was based on the micro-world research paradigm. The results indicated that prospective memory was slightly more vulnerable to variations in experimental conditions than was reaction time. The paper concludes that, overall, prospective memory appears to be a suitable secondary task. This is partly attributed to the element of self-initiation, which is a feature of some prospective memory tasks. It is recommended to use time-based and pulse-based prospective memory tasks since they seem to be the more sensitive. PMID- 10711976 TI - An observation instrument for assessment of work technique in patient transfer tasks. AB - The aim of the study was to construct an observation instrument for description and assessment of nursing personnel's work technique in patient transfer tasks with regard to musculoskeletal health and safety, and to evaluate the validity and reliability of the instrument. The instrument consists of 24 items arranged in three phases of a transfer: the preparation phase, the starting position and the actual performance. Observations are made from video recordings. A detailed description of the individual's work technique, including actions taken to prepare the transfer, the interaction with the patient and any assistant co worker, and the motor performance of the nurse, is provided. An attempt was made to quantify the assessments, by calculating an overall score of the work technique with regard to the level of musculoskeletal hazard and safety. The validity and reliability of the instrument were evaluated on 35 video-recorded patient transfers from hospital wards. The validity and reliability were mostly satisfactory, both when evaluating the agreements between the observations of each item (i.e. kappa values > 0.40), and when evaluating the agreements between the overall scores (i.e. intraclass correlation coefficients 0.71-0.90). Further improvements to enhance the agreements are suggested. PMID- 10711977 TI - Scenario building as an ergonomics method in consumer product design. AB - The role of human factors in design appears to have broadened from data analysis and interpretation into application of discovery and "user experience" design. The human factors practitioner is continually in search of ways to enhance and to better communicate their contributions, as well as to raise the prominence of the user at all stages of the design process. In work with design teams on the development of many consumer products, scenario building has proved to be a valuable addition to the repertoire of more traditional human factors methods. It is a powerful exploration, prototyping and communication tool, and is particularly useful early on in the product design process. This paper describes some advantages and potential pitfalls in using scenarios, and provides examples of how and where they can be usefully applied. PMID- 10711978 TI - Kinetics and energetics during uphill and downhill carrying of different weights. AB - During physically heavy work tasks the musculoskeletal tissues are exposed to both mechanical and metabolic loading. The aim of the present study was to test a biomechanical model for prediction of whole-body energy turnover from kinematic and anthropometric data during load carrying. Total loads of 0, 10 and 20 kg were carried symmetrically or asymmetrically in the hands, while walking on a treadmill (4.5 km h(-1)) horizontally, uphill, or downhill the slopes being 8%. Mean values for the directly measured oxygen uptake ranged for all trials from 0.5 to 2.1 l O2 min(-1), and analysis of variance showed significant differences regarding slope, load carried, and symmetry. The calculated values of oxygen uptake based on the biomechanical model correlated significantly with the directly measured values, fitting to the line Y = 0.990 X + 0.144, where Y is the estimated and X is the measured oxygen uptake in l min(-1). The close relationship between energy turnover rate measured directly and estimated based on a biomechanical model justifies the assessment of the metabolic load from kinematic data. PMID- 10711979 TI - Biomechanical analyses of paramedics simulating frequently performed strenuous work tasks. AB - Firefighters performing emergency rescue functions are at an elevated risk of musculoskeletal injuries. The objective of the current study was to analyze the biomechanical stresses placed on the body based on simulations of the following strenuous and frequently performed emergency rescue tasks: (1) transferring a patient from a bed to a stretcher using bedsheets, (2) transferring a patient from the ambulance stretcher to a hospital gurney, (3) carrying a victim down a set of stairs and through a landing using a stairchair, (4) carrying a victim down a set of stairs and through a landing using a backboard, and (5) carrying a victim down a straight set of stairs using a stretcher. Postural data were analyzed using the University of Michigan's Three-Dimensional Static Strength Prediction Program and the relative risk of low back disorder (LBD) was quantified using the trunk motion model published by Marras et al. (1993, spine 18, 617-628). Peak compression values and the probabilities from the Marras et al. (1993) model indicated that the most hazardous tasks performed as part of this simulation included pulling a victim from a bed to a stretcher, the initial descent of a set of stairs when using the stretcher, and lifting a victim on a backboard from the floor. Overall, the two models were well correlated in their assessment of the task components modelled (r = 0.78). These data indicate where engineering changes to equipment regularly used by emergency rescue personnel would have the greatest impact in reducing the risk of musculoskeletal injury. PMID- 10711980 TI - An ergonomics study on compatibility of controls of overhead cranes in a heavy engineering factory in West Bengal. AB - Ergonomics studies, on the machine control and the resultant movements of the cabins and the hooks in 51 electric overhead travelling cranes in a heavy engineering factory, showed that control-movement compatibility is absent in most of the cranes. Also, the layout of the groups of controls and the orientations of each of the individual controls with respect to the operators' seats varied from one crane to another. As the operators were shifted from one crane to another every week, there was a high chance of making mistakes during moving the controls, which might have resulted in severe accidents, especially during periods of high workload. A number of low-cost ergonomics solutions have been recommended to minimize these problems. PMID- 10711981 TI - Patient handling with and without slings: an analysis of the risk of injury to the lumbar spine. AB - Health professionals handling patients are known to be at risk of sustaining work related low back injuries. It is not known whether the use of lifting slings reduces the risk of injury to the lumbar spine for patient handlers. This study used kinematic variables and subjective ratings of body part stress and lifter preference as measures of relative risk for three two-person techniques for carrying a patient from one chair to another chair. The techniques used no slings, one and two slings respectively. Twenty-two nurses performed five trials each of the three techniques. Kinematic measures of angular displacement, velocity and acceleration were obtained using the lumbar motion monitor and visual analogue scales were used to obtain measures of body part stress for seven body parts. Angular displacement, velocity and acceleration were significantly greater (p < 0.05) in the frontal, sagittal and transverse planes for the no sling technique compared to techniques using slings. Comparatively small yet significant differences between techniques using slings were recorded for sagittal flexion and rotation. There was no significant difference between one and two sling techniques for other dependent variables. Mean total body stress rating was higher for the no sling technique and all subjects indicated that their first preference was for slings. Although all three measures of risk rated the no sling technique as carrying a higher level of risk than the techniques using slings. No single measure adequately captured all aspects of relative risk. The elimination of manual patient handling is thought to be the best option for the reduction of work related back injuries in patient handlers. Where resources or technology are not yet adequate to provide practical alternatives and where the use of manual technique for a seat to seat task is unavoidable, the use of patient handling slings will reduce the risk. PMID- 10711982 TI - Rapid entire body assessment (REBA). AB - This technical note details the preliminary stage in the development of a postural analysis tool, Rapid Entire Body Assessment (REBA). REBA has been developed to fill a perceived need for a practitioner's field tool, specifically designed to be sensitive to the type of unpredictable working postures found in health care and other service industries. A team of ergonomists, physiotherapists, occupational therapists and nurses collected and individually coded over 600 postural examples to produce a new tool incorporating dynamic and static postural loading factors, human-load interface (coupling), and a new concept of a gravity-assisted upper limb position. Initial reliability for inter observer coding shows promise but further work is needed to establish the validity of the tool. PMID- 10711983 TI - Some observations regarding the vibrational environment in child safety seats. AB - A growing issue in the area of vehicular ride comfort is that of child safety seats. Postural, thermal and vibrational comfort considerations are finding their way into child seat design. This paper makes some observations regarding the current state of child safety seat design, then goes on to present the results of vibration tests performed over two road surfaces using two child seats and two children. The vibration levels measured at the interfaces between the children and their seats were found to be higher than the vibration levels between the driver and the driver's seat. Calculated power spectral densities and acceleration transmissibility functions showed that the vibration transmission characteristics of the coupled system consisting of the automobile seat, child seat and child were different from those of the driver/seat system. Whereas, automobile seats normally reduce vibrational disturbances at most frequencies, the child seats tested amplified vibration at most frequencies up to 60 Hz. PMID- 10711984 TI - Muscular dystrophy: historical overview and classification in the genetic era. AB - Despite recent advances in molecular genetics, it has proven very difficult to arrive at an accurate and clinically useful classification of the muscular dystrophies. Much of this difficulty arises from confusion related to the term "muscular dystrophy" itself, as well as a general reluctance on the part of the neuromuscular community to abandon traditional, clinically based classifications. Nevertheless, advances in the understanding of the molecular defects of these disorders have permitted a foundation for classification based on molecular biology. This review presents a historical perspective on the classification of the muscular dystrophies, and furnishes the underpinnings of a genetic classification that can be used both at the bedside and in the research laboratory. PMID- 10711985 TI - The childhood muscular dystrophies: making order out of chaos. AB - New discoveries have dramatically changed the way we approach and think about patients with childhood muscular dystrophies. An aura of order and organization seems to be at hand for a group of diseases which previously seemed endlessly heterogeneous. We have learned that young boys and girls with proximal muscle weakness, large calves and elevated serum CK may have any one of a number of closely connected disorders which affect a complex of interacting proteins of the dystrophin-glycoprotein complex. This complex links the intracellular cytoskeleton to the extracellular matrix. Patients with Duchenne and Becker dystrophies lack dystrophin, while some of the limb girdle muscular dystrophies (an archaic term) are deficient in sarcoglycans and other proteins. The concept of interrelated disorders extends to the previously orphaned distal muscular dystrophies, or distal myopathies, as they are often called. A surprise finding is that the C. elegans protein, dysferlin, is conserved and expressed in man. We know little of the function of this protein in human primates, but its loss in muscle has brought seemingly disparate disorders together, since both a form of LGMD (2B) and distal myopathy (Miyoshi myopathy) are deficient in this same gene product. The congenital muscular dystrophies are also well-entrenched in our expanding concepts of orderliness of disease. The defect in the laminin-alpha2 chain, a direct ligand to the dystrophin-glycoprotein complex, causes a form of muscular dystrophy which affects infants. Another variant of congenital muscular dystrophy is deficient the integrin alpha7, an important laminin receptor. Finally, in Fukuyama congenital muscular dystrophy, the deficient fukutin gene product may also be linked to the basal lamina, permitting overmigration of neuronal cells which lead to micropolygyria in the brain, and at the same time cause basal lamina defects in the extracellular matrix of skeletal muscle, which leads to muscular dystrophy. As we approach the millennium, those of us who have seen the transition from the pre-molecular to the molecular era of myology know that we leave behind a great legacy of chaos (no great loss), replaced by a foundation for conceptual organization which will serve to establish new roots for research as well as for the enriched practice of medicine. The future looks bright for our field and our patients! PMID- 10711986 TI - The myotonic dystrophies. AB - Myotonic dystrophy, or dystrophia myotonica (DM), is the most common inherited muscle disorder in adults. DM is a multisystem disease in which the most disabling feature is muscle wasting that begins in the distal limb and cranial muscles. The genetic basis for DM is an expanded CTG repeat in the DMPK gene on chromosome 19. The size of the expanded repeat, and the severity of the disease, tend to increase in successive generations. The mechanism by which this unusual mutation leads to muscle wasting, myotonia, cataracts, heart block, and neurobehavioral abnormalities has not been clearly defined. Identification of the DM gene has made it easier to delineate other DM-like disorders that are clinically and genetically distinct. The most common of these is proximal myotonic myopathy (PROMM), which is characterized by early involvement of proximal limb muscles. The genetic locus for another DM-like disorder, called DM type 2, was recently mapped to chromosome 3. PMID- 10711987 TI - Facioscapulohumeral dystrophy. AB - Facioscapulohumeral dystrophy (FSHD) is one of the most common inherited neuromuscular disorders, with an estimated prevalence of 1:20,000. The disease is autosomal dominant, although 10-30% of cases appear to arise from a de novo mutation. The disease presents with a characteristic pattern of weakness which affects predominantly the face and scapular stabilizer muscles. Symptoms usually begin in childhood, and >90% of patients have some evidence of disease on examination by age 20. The course of the disease is slowly progressive, although many patients have long periods of relatively stable function. The cause of the disease is unknown, but recent studies have demonstrated genetic linkage to a locus on the long arm of chromosome 4 (4q35). Probes from this region detect an EcoR1 "short fragment" that cosegregates with FSHD in familial cases and appears de novo in most sporadic cases. Although the size of the small fragment correlates inversely with disease severity, the exact relationship of the fragment to the pathogenesis of the clinical disease is unclear, and a specific FSHD gene has not been identified. FSHD is currently untreatable. Few therapeutic trials of the disorder, have been conducted, largely because little is known about the underlying mechanisms of muscle injury in this disease. PMID- 10711988 TI - Distal myopathies. AB - Although muscle disease classically presents with proximal extremity weakness, some myopathic disorders, including several types of muscular dystrophy, result in predominantly, or exclusively, distal muscle involvement. Accurate diagnosis of these relatively uncommon conditions can be challenging for the clinician, because of both the unusual phenotype and the significant overlap in the clinical features of many of these entities. Advances in molecular genetics have permitted a tentative classification of these disorders and have led to the identification of the responsible gene lesion for several of these entities. This review summarizes current understanding of this interesting group of muscular dystrophies and briefly summarizes other myopathies that can present with distal weakness. PMID- 10711989 TI - Oculopharyngeal muscular dystrophy. AB - Autosomal dominant oculopharyngeal muscular dystrophy (OPMD) is an adult-onset disease with worldwide distribution. It usually presents in the fifth or sixth decades with progressive dysphagia, eyelid ptosis, and proximal limb weakness. Unique intranuclear filament inclusions in skeletal muscle fibers are its morphological hallmark. Surgical correction of the ptosis and cricopharyngeal myotomy are the only therapies available. Autosomal dominant OPMD is caused by short (GCG)8-13 riplet-repeat expansions in the polyadenylation binding protein 2 (PABP2) gene, which is localized in chromosome 14q11. Autosomal recessive OPMD is caused by a double dose of a (GCG)7 PABP2 allele. The GCG expansions cause lengthening of a predicted polyalanine tract in the protein. The expanded polyalanine domains may cause polyalanine nuclear toxicity by accumulating as nondegradable nuclear filaments. PMID- 10711990 TI - Emery-Dreifuss muscular dystrophy. AB - Emery-Dreifuss muscular dystrophy (EDMD) is the third most common X-linked muscular dystrophy. This disorder is characterized by childhood onset of early contractures, humeroperoneal muscle atrophy, and cardiac conduction abnormalities. Weakness is slowly progressive, but there is a broad spectrum of clinical severity. Patients and carriers are at risk of sudden death. Regular cardiac evaluation is mandatory to assess the risk of cardiac arrhythmias. Unique atrial pathology is seen at autopsy. The mutated gene in EDMD is localized to the long arm of the X chromosome. Mutations in the gene lead to abolished synthesis of the gene product, emerin. Emerin is localized to the nuclear membrane of skeletal, cardiac, and smooth muscle. The term Emery-Dreifuss syndrome describes patients who have the EDMD phenotype without X-linked inheritance. There is no treatment for the underlying disease, but early placement of pacemakers may be lifesaving. PMID- 10711991 TI - Outlook for therapy in the muscular dystrophies. AB - Despite recent advances in our understanding of the muscular dystrophies, current therapy for these disorders remains chiefly supportive. With the exception of prednisone for Duchenne dystrophy, no effective treatment for these disorders has been found that will improve strength or even slow the progression of weakness. This review highlights current strategies for treating muscular dystrophies and discusses recent trials of various pharmacologic agents, including corticosteroids, anabolic steroids, growth factors and beta-adrenergic agonists. The outlook for effective gene therapy is also presented. PMID- 10711992 TI - Early observations on muscular dystrophy: Gowers' textbook revisited. AB - Early clinical observations on Duchenne muscular dystrophy can be traced through the works of Meryon, Little, Duchenne, Gowers, and Erb. Gowers sites Sir Charles Bell with its earliest clinical description. Gowers' phenomenal textbook provides vivid descriptions of Duchenne dystrophy, clinical features which are herein revisited. PMID- 10711993 TI - Registration and difference analysis of corresponding mammogram images. AB - An automated technique is proposed for identifying differences between corresponding mammogram images. The technique recovers an approximate deformation between a pair of mammograms based on identifying corresponding features across the two images. The registration process is completed using an unwarping technique for transforming one image into the coordinate system of the other. A difference image is generated using intensity-weighted subtraction in order to identify regions of large difference. Evaluation of the technique is performed using 124 bilateral image pairs which contain a total of 77 abnormalities of different types. The purpose of this paper is to measure the extent to which the mammogram registration technique is able to provide useful information for identifying abnormalities in mammograms. PMID- 10711994 TI - Bone labelling on micro-magnetic resonance images. AB - The study of trabecular bone is receiving increasing interest within the medical community working in the field of skeletal diseases, such as osteoporosis. Quantification of trabecular bone structure usually requires as a starting point a correct segmentation of the trabecular network. We have developed a probabilistic relaxation labelling technique, which uses local features of the trabecular bone images to improve segmentation. Tests on synthetic images show that bone labelling performs a more accurate segmentation than conventional techniques such as thresholding, especially by preserving the connectivity of the trabecular network. Tests on acquired data show that porosity values obtained after segmentation are in good agreement with experimental values obtained by weighing the bone samples. PMID- 10711995 TI - Stradx: real-time acquisition and visualization of freehand three-dimensional ultrasound. AB - Conventional freehand three-dimensional (3-D) ultrasound is a multi-stage process. First, the clinician scans the area of interest. Next, the ultrasound data is used to construct a 3-D voxel array, which can then be visualized by, for example, any-plane slicing. The strict separation of data acquisition and visualization disturbs the interactive nature of the ultrasound examination. Furthermore, some systems require the clinician to wait for an unacceptable amount of time while the voxel array is constructed. In this paper, we describe a novel freehand 3-D ultrasound system which allows accurate acquisition of the raw data and immediate visualization of arbitrary slices through the data. Minimal processing separates the acquisition and visualization processes: in particular, at no stage is a voxel array constructed. Instead, the standard graphics hardware found inside most desktop computers is exploited to synthesize arbitrary slices directly from the raw B-scans. PMID- 10711996 TI - Fast surface and volume estimation from non-parallel cross-sections, for freehand three-dimensional ultrasound. AB - Volume measurements from ultrasound B-scans are useful in many clinical areas. It has been demonstrated previously that using three-dimensional (3-D) ultrasound can greatly increase the accuracy of these measurements. Freehand 3-D ultrasound allows freedom of movement in scanning, but the processing is complicated by having non-parallel scan planes. Two techniques are proposed for volume measurement from such data, which also improve surface and volume estimation from data acquired on parallel planes. Cubic planimetry is a more accurate extension of a volume measurement technique involving vector areas and centroids of cross sections. Maximal-disc shape-based interpolation is an extension of shape-based interpolation which uses maximal disc representations to adjust the interpolation direction locally and hence improve the quality of the surface generated. Both methods are tested in simulation and in vivo. Volumes estimated using cubic planimetry are more accurate than step-section planimetry, and require fewer cross-sections, even for complex objects. Maximal-disc shape-based interpolation provides a reliable means of reconstructing surfaces from a handful of cross sections, and can therefore be used to give confidence in the segmentation and hence also the cubic planimetry volume. PMID- 10711997 TI - Alignment of magnetic-resonance brain datasets with the stereotactical coordinate system. AB - Neuroanatomical and neurofunctional studies are often referenced to high resolution magnetic-resonance brain datasets. For the analysis of the cortical surface, mapping of functional information on to the cortex or visualization, it is necessary to remove the outer surfaces of the brain. For intersubject comparison, it is useful to align the dataset with a coordinate system and introduce a spatial normalization. We describe an image processing chain that combines all of these steps in an interaction-free procedure. We report on a period of 2 years of routine application of this procedure, with >250 successfully processed datasets from healthy subjects and patients with various forms of brain damage. PMID- 10711998 TI - Deformable meshes with automated topology changes for coarse-to-fine three dimensional surface extraction. AB - This work presents a generic deformable model for extracting objects from volumetric data with a coarse-to-fine approach. This model is based on a dynamic triangulated surface which alters its geometry according to internal and external constraints to perform shape recovery. A new framework for topology changes is proposed to extract complex objects: within this framework, the model dynamically adapts its topology to the geometry of its vertices according to simple distance constraints. In order to speed up the process, an algorithm of pyramid construction with any reduction factor transforms the image into a set of images with progressively higher resolutions. This organization into a hierarchy, combined with a model which can adapt its sampling to the resolution of the workspace, enables a fast estimation of the shapes included in the image. After that, the model searches for finer and finer details while relying successively on the different levels of the pyramid. PMID- 10711999 TI - In vivo veritas. PMID- 10712000 TI - Engulfment of prions in the germinal centre. PMID- 10712001 TI - How much longer will tumour cells fool the immune system? PMID- 10712002 TI - Antigen recognition. PMID- 10712003 TI - Itraconazole verus fluconazole in neutropenic patients. PMID- 10712004 TI - Kaposi's sarcoma-associated herpesvirus (KSHV) in bone marrow biopsy from patients with multiple myeloma: PCR amplification of orf26 but not orf72 and orf75 sequences. PMID- 10712005 TI - Antenatal testing for haemoglobinopathies. PMID- 10712006 TI - The retinoic acid syndrome in non-M3 acute myeloid leukaemia: a case report. PMID- 10712007 TI - Some women pioneers in haematology. PMID- 10712008 TI - Photo quiz. Cutaneous alternariosis. PMID- 10712009 TI - Proceedings of the 3rd Single Chromosome 6 Workshop. Cambridge, United Kingdom, 2 5 March 1997. PMID- 10712010 TI - Quality improvement guidelines for adult diagnostic neuroangiography. Cooperative study between ASITN, ASNR and SCVIR. American Society of Interventional and Therapeutic Neuroradiology. American Society of Neuroradiology. Society of Cardiovascular and Interventional Radiology. PMID- 10712012 TI - Distal myopathies. 25th ENMC International Workshop, 18-20 November 1994, Naarden, The Netherlands. PMID- 10712011 TI - Close linkage of Mdm-1, a gene amplified and overexpressed in a transformed 3T3 cell line, with gamma interferon (Ifg) on chromosome 10 of the mouse. AB - The Mdm-1 gene was mapped to the distal end of Chromosome (Chr) 10. An extensive series of restriction fragment variants was identified among conventional and exotic inbred strains of mice. Mapping was carried out with recombinant inbred strains and an intersubspecific testcross. No recombinants were observed between Mdm-1 and the gamma interferon locus (Ifg). These two loci appear to be in linkage disequilibrium among inbred strains. Data from the testcross place Mdm-1 approximately 11 centimorgans distal to the steel (Sl) locus. Because of its extensive polymorphism, Mdm-1 is a useful genetic marker for distal Chr 10. PMID- 10712013 TI - Proceedings of the 27th ENMC sponsored workshop on congenital muscular dystrophy. 22-24 April 1994, The Netherlands. PMID- 10712014 TI - Research database for dystrophin gene and protein expression. 35th ENMC Workshop, 6-8 October 1995, Naarden, The Netherlands. PMID- 10712015 TI - 38th ENMC International Workshop. Spinal muscular atrophy trial group 10-12 December 1995, Naarden, The Netherlands. PMID- 10712016 TI - 50th ENMC International Workshop: congenital muscular dystrophy. 28 February 1997 to 2 March 1997, Naarden, The Netherlands. PMID- 10712017 TI - Commentary: applying the BMJ's guidelines on educational interventions. PMID- 10712018 TI - Meningococcal disease in healthcare workers. Prophylaxis is not necessary. PMID- 10712019 TI - Meningococcal disease in healthcare workers. Recommendation will cause unease among healthcare staff. PMID- 10712020 TI - Meningococcal disease in healthcare workers. Long term effects and costs are unclear. PMID- 10712021 TI - Meningococcal disease in healthcare workers. Ceftriaxone may be helpful. PMID- 10712022 TI - Why mortality from heart disease is low in France. High cholesterol may not have same effect on cardiovascular risk in southern Europe as elsewhere. PMID- 10712023 TI - Why mortality from heart disease is low in France. Wine consumption clearly correlates with residual differences in mortality. PMID- 10712024 TI - Private finance initiative. Series did not address real planning issues. PMID- 10712025 TI - Private finance initiative. The initiative puts strain on primary care groups in east London. PMID- 10712026 TI - Commentary: oral exams--get them right or don't bother. PMID- 10712027 TI - Zanamivir, influenza, and meningococcal disease. NHS regulations are of questionable legality. PMID- 10712028 TI - Managing patients with lung cancer. Special palliative care is needed. PMID- 10712029 TI - Managing patients with lung cancer. Biomedical literature does not support routine use of laboratory variables as prognostic factors. PMID- 10712030 TI - Managing patients with lung cancer. Common international guidelines must be developed. PMID- 10712031 TI - Competing interests and controversy about third generation oral contraceptives. Science is not a dispassionate activity. PMID- 10712032 TI - Not such distant mirrors. Coffee enemas may be effective shock treatment. PMID- 10712033 TI - Human population growth. China's one child family policy is changing. PMID- 10712034 TI - Human population growth. Population issue is not entirely satisfactory. PMID- 10712035 TI - Coping with winter bed crises. Answer is not data showing crises will happen. PMID- 10712036 TI - Coping with winter bed crises. Essence of problem is insufficient resources. PMID- 10712037 TI - Prospective risk of stillbirth. Randomised trials of earlier induction of labour are needed. PMID- 10712038 TI - Prospective risk of stillbirth. Impending fetal death must be identified and pre empted. PMID- 10712040 TI - Peer reviewed papers selected from presentations at the International Conference on Nitric Oxide held at the National University of Singapore. PMID- 10712039 TI - Medicine to serve an ageing society. Profile of elderly people in society needs to be raised. PMID- 10712041 TI - Phospholipases A2 and Wnts are unlikely to share a common ancestor. PMID- 10712042 TI - First Surgeon General's report on mental health stresses effectiveness of treatment, notes many fail to seek help. PMID- 10712043 TI - NAMI review finds only a third NIMH funding goes to support research on severe mental illnesses. PMID- 10712044 TI - Special issue on the Henry Kempe's Center: past, present, future. PMID- 10712045 TI - Denver's Kempe Children's Center. PMID- 10712046 TI - Modafinil approved for narcolepsy. PMID- 10712047 TI - The management of extreme hypernatraemia secondary to salt poisoning in an infant. PMID- 10712048 TI - Unilateral pulmonary cystic enlargement in a newborn: remember the one-sided blind intubation. PMID- 10712049 TI - Reflections on sevoflurane. PMID- 10712050 TI - Exchange of tracheostomy tubes in an infant. PMID- 10712051 TI - Some hints to make neonatal epidural anaesthesia less difficult. PMID- 10712052 TI - The Hyderabad Chloroform Commission. PMID- 10712053 TI - Proceedings of the Canadian Conference on Shared Responsibility and Health Impact Assessment: Advancing the Population Health Agenda. Vancouver, British Columbia, Canada. 2-3 May 1999. PMID- 10712054 TI - Smoking in enclosed shopping centres: employee and public responses to simulated violation. PMID- 10712055 TI - Tobacco education in Cairo, Egypt: is there an effect on adolescent smoking? PMID- 10712056 TI - Motel kids syndrome. PMID- 10712057 TI - [Dissertations in orthodontics in 1999]. PMID- 10712058 TI - [Intense uptake of 99mTc HMPAO leukocytes in a patient with a non-infected pseudoaneurysm of the left femoral artery]. PMID- 10712059 TI - [Cortical renal scintigraphy and urinary tract infection]. PMID- 10712061 TI - [Nomenclature for nuclear medicine procedures. 1999 update]. PMID- 10712060 TI - [PET Radiopharmacy Unit]. PMID- 10712062 TI - [Informed consent in the radioisotopic treatment of hyperthyroidism. Procedures Committee of the Spanish Nuclear Medicine Society]. PMID- 10712063 TI - 30 years later: the relation between structure and function in the brain from a contemporary point of view (1966), part I. PMID- 10712064 TI - On the problem of the relation between structure and function in the brain from a contemporary point of view. 1966. PMID- 10712065 TI - Could a nasal spray protect people against human immunodeficiency virus? PMID- 10712066 TI - Theoretical investigation of the interrelationships between stationary and personal sampling in exposure estimation. AB - In exposure estimation, personal sampling is the method of choice as it is a nearby representative of the contaminant concentration in the breathing zone. Due to the versatility of the stationary sampling in obtaining much higher sensitivity, in its adaptability to telemetering observations, it may also be an attractive sampling method for many circumstances. However, the two sampling methods differ in many theoretically important ways that go beyond the obvious differences. The theoretical investigation of the stationary and personal sampling methods vis-a-vis sampling for exposure estimation shows that the area sampling can be used to represent personal sampling under restricted conditions. Under the restricted conditions, an area of concentration within specified bounds may be determined in relation to a reasonably well-defined source. The extension of the theory to multiple or ill-defined sources pose potential complications that may be intractable through a theoretical analysis. These limitations and restrictions are inherent to the underlying premises of the two methods; therefore they are not amenable to easy correction. Even though these restrictions may suggest only a limited role for area sampling in exposure assessment, the theory shown also suggests areas of further applied and theoretical research to extend the proper use of area sampling in exposure assessment. PMID- 10712067 TI - Assessment of occupational exposure patterns by frequency-domain analysis of time series data. AB - Laboratory evidence increasingly points to exposure pattern characteristics, including the duration, frequency, and timing of the exposure during the day, as important factors influencing the biological response to extremely low-frequency magnetic fields. An exploratory analysis of exposure patterns was conducted in 113 electric utility workers employed as electricians, cable splicers, line workers, and power plant operators. The purpose of the study was to describe extremely low-frequency magnetic field exposure pattern characteristics of electric utility workers and evaluate grouping strategies for classifying occupational exposures based on their exposure pattern characteristics. Exposure patterns describe the cyclic fluctuation in exposures over time, and were evaluated by partitioning the variation of the time series into frequency components using frequency-domain analysis of the transformed and processed time series. The study samples were classified using traditional grouping strategies based on occupation and time-weighted average (TWA), and non-traditional grouping strategies based on cluster analysis of the standardized, low-frequency exposure pattern components. Rules for classifying samples into each group were developed using linear discriminant analysis, with the performance of each grouping strategy evaluated using a crossvalidation study design to estimate the rate of misclassification. Exposure patterns appeared unrelated to grouping strategies based on quartiles of the workday TWA, but were related to pattern clusters and occupation. The linear discriminant function produced very low misclassification error rates for the cluster grouping strategy (10%) compared to occupation (50%) and TWA quartile (69%) grouping strategies. Significant differences in the exposure patterns occurring between clusters and between occupational groups were observed, indicating that at least one of the spectral estimates in two of the groups were significantly different. However, patterns clusters produced the greatest contrast in exposure patterns of all grouping strategies, explaining 99 percent of the total variation compared to 58 percent of the total variation by occupation. PMID- 10712069 TI - A tiered approach to deterministic models for indoor air exposures. AB - There are a number of deterministic mathematical approaches available for modeling indoor air pollution concentrations. These models range in complexity from simple saturation vapor pressure models to models using computational fluid dynamics, with many in between these extremes. This range reflects the variety of ways pollutant generation, transport, and mixing are treated in the different models. In selecting which model to use, a tiered approach is useful. The tiered approach considers the goal of the modeling, the availability of model inputs, and the degree of uncertainty that is acceptable. The simpler models are easy to use and the inputs are often readily available. However, they usually have an inherently high degree of uncertainty which requires conservative assumptions that may result in overestimates of concentrations. The more complex models are more difficult to use and require more precise inputs. However, they have the potential to allow the modeler to reduce or quantify uncertainty. This ability to address uncertainty may be limited by the understanding of and the availability of the model inputs. An application example of the tiered approach using a completely mixed space model, a two-zone model, and a turbulent diffusion model illustrates the selection criteria for model use and the model performances. Ultimately, an understanding of the principles behind the models and their inherent strengths and weaknesses is required for their appropriate application to exposure assessment. PMID- 10712068 TI - Comparison of mathematical models for exposure assessment with computational fluid dynamic simulation. AB - For many years exposure to airborne contaminants has been estimated by air or biological monitoring. In occupational settings, mathematical models increasingly are employed as adjuncts to monitoring, for instance, during process design or in retrospective epidemiological studies. Models can make predictions in a wide variety of scenarios, can be used for rapid screening, and may reduce the need for monitoring in exposure assessment. However, models make simplifying assumptions regarding air flow and contaminant transport. The errors resulting from these assumptions have not been systematically evaluated. Here we compare exposure estimates from the single-zone completely mixed (CM-1), two-zone completely mixed (CM-2), and uniform diffusivity (UD) models with workroom concentration fields predicted by computational fluid dynamics (CFD). The room air flow, concentration fields, and the breathing zone concentration of a stationary worker were computed using Fluent V4.3 for factorial combinations of three source locations, three dilution air flow rates and two emission rate profiles, constant and time-varying. These numerical experiments were used to generate plausible concentration fields, not to simulate exactly the processes in a real workroom. Thus, "error" is defined here as difference between model and CFD predictions. For both constant and time-varying emission sources, exposure estimates depended on receptor and source location. For the constant source case, ventilation rate was shown to be inconsequential to CM-1 model error. CM-1, CM-2, and UD models differed in their agreement with CFD. UD was closest to CFD for estimating concentration in the simulated breathing zone (BZ) near the source, although large errors resulted when the model was applied to the plane of possible breathing zones. CM-1 performed better for this plane but underestimated the near-source BZ exposure. For the near-source BZ location, CM-2 replicated CFD predictions more closely than CM-1 did, but less closely than UD did. Error in CM 1 model estimation of short-term average exposure to a time-varying source was highly dependent on ventilation rate. Error decreased as ventilation rate increased. PMID- 10712071 TI - Laboratory evaluation of pressure differential-based respirable dust detector tube. AB - Assessment of exposure to occupational dusts is a first step in reducing exposures to harmful dust concentrations. A new type of respirable dust sampler was developed and compared side-by-side to personal gravimetric samplers in the laboratory. The new sampler correlates filter back pressure with mass accumulation to provide mid-shift- and end-of-shift determinations of cumulative exposure. The sampler uses a small low flow rate pump to draw dust through a small detector tube that contains a porous urethane foam respirable classification section and glass fiber filter that collects respirable dust. Six different coal dusts were aerosolized in a laboratory dust chamber and a total of 119 triplicate observations were obtained. For individual coal types, the correlation coefficients were between 0.87 and 0.97. The precision of the two methods was similar, with the percent relative standard deviation of the personal samplers of 12 percent and the new detector method of 14 percent. For all coal types tested th data were best described by a power function where delta P = 1.43 mass (0.85), with a correlation coefficient of 0.73. The method becomes more accurate at higher dust loadings such that all laboratory data with mass loadings greater than an equivalent concentration of 2 mg/m3 fall within +/- 25 percent of the power function. Assessment of the method under field conditions is in progress. PMID- 10712070 TI - Preliminary report on the results of the second phase of a round- robin endotoxin assay study using cotton dust. AB - In an on-going endotoxin assay study, a two-part interlaboratory endotoxin assay study has been completed. The purpose of the study was to compare the variation in assay results between different laboratories, and, if the variation was high, to see if a common protocol would reduce the variation. In both parts of the study, membrane filters laden with the same approximate amount and type of cotton dust were sent for analysis to laboratories that "routinely" perform endotoxin analyses. First, each of these laboratories performed the analysis using the methodology common to its laboratory. In the second part of the study, membrane filters with cotton dust were again sent to the same laboratories where the analyses were performed as before but with a common extraction protocol. The preliminary results from the first phase of the study have been collected and showed that intra-laboratory variations were small, but large and significant interlaboratory variation was observed. The results were reported elsewhere. The preliminary results from the second part of the study consisting of the data currently collected are presented here. Again, intra-laboratory variations were small, but, also again, large and significant inter-laboratory variation was observed. However, in this part of the study, the range between the highest and lowest average results was narrower than in the first part of the study. Influence of the assay kit type was examined. The variation within assay kit type was small but significant differences in results were observed between assay kit types. The findings suggest that endotoxin concentration in samples can be ranked within laboratories, but not necessarily between laboratories. However, some of the variation between laboratories has been reduced by a common extraction protocol which suggests the possibility of further standardization that may lead to better comparability between laboratories. PMID- 10712072 TI - Validation of parachlorobenzotrifluoride, benzotrifluoride, and monochlorotoluene on diffusive samplers. AB - Three solvents (OXSOL 10, monochlorotoluene or mixed isomers of 1- chloro-2 methyl benzene and 1-chloro-4-methyl benzene; OXSOL 100, parachlorobenzotrifluoride or 1-chloro-4-(trifluoromethyl) benzene; and OXSOL 2000, benzotrifluoride or trifluoromethyl benzene) produced by Occidental Chemical Corporation (Niagara Falls, NY) were considered as candidates for SKC, Inc.'s on-going diffusive sampler validation program. The 575-series diffusive sampler contains coconut-shell charcoal (575-001) or Anasorb 747 (575-002). Both samplers were used in this study. Desorption efficiency was tested at loadings equivalent to eight-hour time-weighted average (TWA) exposures to 0.01-2 times the Occidental Chemical Corporation in-house limit values (respectively: 50 ppm, 25 ppm, and 100 ppm,. All results met the National Institute for Occupational Safety and Health (NIOSH) criteria of > 75 percent, and, except for the lower loadings of parachlorobenzotrifluoride, the results were in the range of 90-110 percent. The calculated uptake rates were verified for different periods of exposure, up to eight hours, and found to be within 5 percent of the calculated for all three compounds on both samplers. A detailed comparison of the results from different time periods indicated no significant reverse diffusion effects for any combination of sampler and analyte. Samplers exposed to standard atmospheres of each compound were stored for three weeks at ambient temperatures and reanalyzed with results between 94 and 107 percent of expected. Based on full validation of samplers for the lower homologue (benzene), the bi-level theory of sample validation as endorsed by international validation protocols establishes this as a complete validation of the featured samplers for sampling vapors of these chemicals in air. PMID- 10712073 TI - but quantitation of probiotics, cytokines and organ substrate flux is more problematical. PMID- 10712074 TI - Functional foods for athletes. PMID- 10712075 TI - Special dialogue: Herbal remedies for depression. PMID- 10712077 TI - Proceedings of a conference on public/private-sector collaboration in contraceptive research and development: a new partnership. Italy, 29 March-2 April 1999. PMID- 10712076 TI - Cloning, high level-expression and characterization of human lens thioltransferase. PMID- 10712078 TI - Ultrasonographic findings in Crohn's disease. PMID- 10712079 TI - Association between colon cancer and adenocarcinoma of the oesophagus. PMID- 10712080 TI - Diversion colitis as a trigger for ulcerative colitis. PMID- 10712081 TI - Quality of life of parents of children on home parenteral nutrition. PMID- 10712082 TI - Helicobacter pylori infection and autoimmune pathogenesis of gastric neoplasias. PMID- 10712083 TI - [Symposium "Emerging Approaches to Cardiovascular Diseases: Detailed results and implications of the HOPE Study". Atlanta, 10. November 1999]. PMID- 10712084 TI - [Vasopeptidase inhibition. Intense and long-lasting blood pressure lowering]. PMID- 10712085 TI - [Long-term administration of Dalteparin in unstable coronary syndrome. FRISC II study opens novel therapeutic perspectives. Fragmin and Fast Revascularization during Instability in Coronary Artery Disease]. PMID- 10712086 TI - Care for life. Central is the patient. PMID- 10712087 TI - ELITE II. AT1-blockers not better, but more tolerable than ACE-inhibitors. PMID- 10712088 TI - [Approval for recombinant growth factor. Since January 2000, patients with diabetic foot ulcers can be treated with the growth factor becaplermin (Regranex), manufactured with genetic technology]. PMID- 10712089 TI - [Health care rationing or flexibility of health care]. PMID- 10712090 TI - [Global budget taxation]. PMID- 10712091 TI - [Prenatal diagnosis or prenatal selection. A grave step of grade for the physician]. PMID- 10712092 TI - [Pharmacology of NSAIDS]. PMID- 10712093 TI - [Ovarian insufficiency induced by cytotoxic drugs]. PMID- 10712094 TI - [Endocarditis prophylaxis]. PMID- 10712095 TI - [Acute ischemia of the legs and rapidly progressing kidney failure in a 39 year old woman. Contribution to N. Buchner et al. (1999) Internist 40:555-560]. PMID- 10712096 TI - Nucleolus organizer regions in Physalaemus cuvievi (Anura, Leptodactylidae), with evidence of a unique case of Ag-NOR variability. AB - We studied ten specimens of Physalaemus cuvieri collected at different localities in Brazil using conventional staining and banding techniques. All specimens had 2n = 22. There were karyotypic variants: distinct patterns in the number and chromosome localization of Ag-NORs as well as in the corresponding secondary constrictions. Preliminary C-banding patterns obtained for specimens from two localities are also suggestive of karyotypic differentiation in P. cuvieri. PMID- 10712097 TI - Cross-species amplification of salmonid microsatellites which reveal polymorphism in European and Arctic grayling, Salmonidae: Thymallus spp. PMID- 10712098 TI - Centers for Disease Control and Prevention. 4th Decennial International Conference on Nosocomial and Healthcare-Associated Infections. Atlanta, Georgia, March 5-9, 2000. Abstracts. PMID- 10712099 TI - Freeze-fracture cytochemistry: a new method combining immunocytochemistry and enzyme cytochemistry on replicas. AB - We describe a new freeze-fracture cytochemical technique consisting of combined immunocytochemistry and enzyme cytochemistry. This technique reveals the relationship between molecules in biological membranes by double labeling with two different cytochemical markers (i.e., immunogold probes and cerium). In this method, antigens were detected with specific primary antibodies and appropriate secondary immunoprobes. Subsequently, alkaline phosphates activity was detected with cerium as the capture agent on the same replicas. Octyl-glucoside (OG) digestion before the cytochemical reactions was crucial to the success of this combined method. OG is an efficient detergent and OG digestion can preserve both immunocytochemical antigenicity and enzyme activity on replicas. As an initial examination, we applied this technique to the study of glycosyl-phosphatidyl inositol-anchored proteins and adhesion molecules in human neutrophils. The method described here should serve as a unique additional approach for the study of topology and dynamics of molecules in biomembranes. PMID- 10712100 TI - SPARC is expressed by ganglion cells and astrocytes in bovine retina. AB - SPARC (secreted protein, acidic and rich in cysteine)/osteonectin is a matricellular, counteradhesive glycoprotein that disrupts cell-matrix interactions, interacts with growth factors and components of extracellular matrix, and modulates the cell cycle, but appears to subserve only minor structural roles. SPARC is expressed in a variety of tissues during embryogenesis and remodeling and is believed to regulate vascular morphogenesis and cellular differentiation. Although usually limited in normal adult tissues, SPARC is expressed at significant levels in the adult central nervous system. Using a monoclonal antibody against bovine bone osteonectin, we have determined the localization of SPARC in newborn (3-day-old) and adult (4-8-year-old) normal bovine retinas. SPARC was present in the soma of ganglion cells and strong reactivity was found in ganglion cell axons. Muller cells displayed no immunoreactivity, but SPARC was present in retinal astrocytes that were identified by the astrocyte marker glial fibrillary acidic protein (GFAP). Newborn calf retina showed a staining pattern similar to that of adult retina but exhibited significantly reduced levels of SPARC. Minimal levels of SPARC protein were also detected in some capillaries of the inner retina of both newborn and adult animals, whereas large vessels were negative. The presence of SPARC in the retina was confirmed by Western blotting of retinal extracts. These data indicate that SPARC originating from bot h neurons and glia of the inner retina may be an important modulator of retinal angiogenesis. The increased expression of SPARC in adult relative to newborn retinal tissue also indicates that SPARC has an ongoing role in the maintenance of retinal functions. PMID- 10712101 TI - The abuse of the English language. PMID- 10712102 TI - Festschrift to L. Joseph Butterfield, MD. PMID- 10712103 TI - Honey: is it worth rubbing it in? PMID- 10712104 TI - The CAG repeat within the androgen receptor gene in male breast cancer patients. PMID- 10712105 TI - Deletion and duplication of the adenomatous polyposis coli gene resulting from an interchromosomal insertion involving 5(q22q23.3) in the father. PMID- 10712106 TI - Hypoparathyroidism, retarded growth and development, and dysmorphism or Sanjad Sakati syndrome: an Arab disease reminiscent of Kenny-Caffey syndrome. PMID- 10712107 TI - Tandem duplication within the neurofibromatosis type 1 gene (NF1) and reciprocal t(15;16)(q26.3;q12.1) translocation in familial association of NF1 with intestinal neuronal dysplasia type B (IND B) PMID- 10712108 TI - Identification of novel alleles at a polymorphic microsatellite repeat region in the human NRAMP1 gene promoter: analysis of allele frequencies in primary biliary cirrhosis. PMID- 10712109 TI - No evidence for imprinting in distal 18q. PMID- 10712111 TI - Clinical geneticists' attitudes and practice towards testing for breast cancer susceptibility genes. PMID- 10712110 TI - Renal angiomyolipomata and learning difficulty in tuberous sclerosis complex. PMID- 10712112 TI - Ultrasound detects occult colorectal tumors. PMID- 10712113 TI - Urokinase and dialysis therapy. PMID- 10712114 TI - Histologic characteristics of sternoclavicular beta2-microglobulin amyloidosis. PMID- 10712115 TI - Arteriovenous fistula adequacy. PMID- 10712116 TI - Dipyridamole and low-dose warfarin without cyclophosphamide in the management of IgA nephropathy. PMID- 10712117 TI - Detection of fragments of beta2-microglobulin in amyloid fibrils. PMID- 10712118 TI - Lower extremity amputations in ESRD patients. PMID- 10712119 TI - Need for a weight-based hemodialysis prescription rather than changing the hemodialysis-dose-measure from Kt/V to Kt. PMID- 10712120 TI - Therapy for cardiovascular disease in dialysis patients. PMID- 10712121 TI - Virtual colonoscopy. PMID- 10712122 TI - Virtual colonoscopy. PMID- 10712123 TI - Virtual colonoscopy. PMID- 10712124 TI - Virtual colonoscopy. PMID- 10712125 TI - Risks and benefits of screening for intracranial aneurysms. PMID- 10712126 TI - Medical mystery--the answer. PMID- 10712127 TI - Direct sale of sildenafil (Viagra) to consumers over the Internet. PMID- 10712128 TI - Direct sale of sildenafil (Viagra) to consumers over the Internet. PMID- 10712129 TI - The use of physical restraints in medical emergencies. PMID- 10712130 TI - The use of physical restraints in medical emergencies. PMID- 10712133 TI - The use of physical restraints in medical emergencies. PMID- 10712132 TI - The use of physical restraints in medical emergencies. PMID- 10712131 TI - The use of physical restraints in medical emergencies. PMID- 10712134 TI - Electric razors as a potential vector for viral hepatitis. PMID- 10712135 TI - Brainstem tuberculoma micmicking glaucoma. PMID- 10712136 TI - Proceedings from the International Symposium on Primordial Prevention of Cardiovascular Disease Risk Factors. International symposium honoring the career and research of Henry Blackburn, M.D. PMID- 10712137 TI - Changing the course of rheumatoid arthritis. Introduction. PMID- 10712138 TI - Were Spaniards among the first Americans? PMID- 10712139 TI - Genomics. Mouse sequencers take up the shotgun. PMID- 10712140 TI - Cell biology. New clue to age control in yeast. PMID- 10712141 TI - Molecular computing. RNA works out knight moves. PMID- 10712142 TI - Scientific misconduct. Fired researcher is rehired and refired. PMID- 10712143 TI - Epidemiology. U.K. plans major medical DNA database. PMID- 10712144 TI - Biomedicine. New genetic tricks to rejuvenate ailing livers. PMID- 10712145 TI - Scientific community. Controversy claims CDC lab chief. PMID- 10712146 TI - Ecologists on a mission to save the world. PMID- 10712147 TI - Transforming a discipline. A new breed of scientist-advocate emerges. PMID- 10712148 TI - Biomedical patents. Patent office may raise the bar on gene claims. PMID- 10712149 TI - Politics, misinformation, and biotechnology. PMID- 10712150 TI - "Restoration"--a misnomer? PMID- 10712151 TI - Names for cash. PMID- 10712152 TI - "Science wars". PMID- 10712153 TI - Clarificatioin of AstraZeneca's R&D strategies. PMID- 10712154 TI - Cooperating on childhood cancer. PMID- 10712155 TI - Policy forum: public health. The health and wealth of nations. PMID- 10712156 TI - Perspectives: structural biology. SRP--where the RNA and membrane worlds meet. PMID- 10712157 TI - Perspectives: evolutionary biology. A powerhouse divided. PMID- 10712158 TI - Techview: computers and biology. Bioinformatics in the information age. PMID- 10712159 TI - Suicide statistics. PMID- 10712160 TI - The hypothesis is there is no hypothesis. The Microarray Meeting, Scottsdale, Arizona, USA, 22-25 September 1999. PMID- 10712161 TI - Trans-Siberian X press report. International Symposium on X Chromosome Inactivation in Mammals, Institute of Cytology and Genetics, Novosibirsk, Russia, 6-12 September, 1999. PMID- 10712162 TI - P2Y receptors: in the middle of the road. PMID- 10712163 TI - Immunophilin ligands as a novel treatment of neurological disorders. PMID- 10712164 TI - Nurses and models of practice...then & now. PMID- 10712165 TI - DVT prevention. Low-molecular-weight heparin is a viable option. PMID- 10712166 TI - The rising tide of health care errors. PMID- 10712167 TI - Preventing needlesticks in your facility. PMID- 10712168 TI - Re-enforcing hygiene practices of anaesthetists. PMID- 10712169 TI - Emergency management of previously undiagnosed anterior mediastinal tumour. PMID- 10712170 TI - Laryngoscopy grades. PMID- 10712171 TI - Ultrasound and percutaneous tracheostomy. PMID- 10712172 TI - Airway control and percutaneous tracheostomy. PMID- 10712173 TI - 'Ventouse'-aided intubation. PMID- 10712174 TI - The LMA and difficulty with internal jugular vein cannulation. PMID- 10712175 TI - Another 'whoosh' test. PMID- 10712176 TI - Ambulation in labour. PMID- 10712177 TI - An unexpectedly shallow epidural space. PMID- 10712178 TI - Spinal anaesthesia in three patients with Paget's disease of bone. PMID- 10712179 TI - Paediatric intensive care. PMID- 10712180 TI - Visual estimation of children's weights. PMID- 10712181 TI - Pain and opiates following elective craniotomy. PMID- 10712182 TI - Capnography readings. PMID- 10712183 TI - Ophthalmic draping and carbon dioxide retention. PMID- 10712184 TI - Subcutaneous emphysema in a patient with bronchial asthma after general anaesthesia for transurethral resection. PMID- 10712185 TI - No laughing matter--a failure of pipeline nitrous oxide supply. PMID- 10712186 TI - Are anaesthetic trainees a high-risk group for road accidents? PMID- 10712187 TI - Palacos and peanut oil. PMID- 10712188 TI - Dystrophica myotonia and suxamethonium. PMID- 10712189 TI - Food and agriculture security: guarding against natural threats and terrorist attacks affecting health, national food supplies, and agricultural economics. Proceedings of an international conference. Washington DC, USA. September 28-30, 1998. PMID- 10712190 TI - [Meeting of the National Academy of Pharmacy. Proceedings of the meeting of 20 October 1999]. PMID- 10712191 TI - [Proceedings of the meeting 10 November 1999]. PMID- 10712192 TI - [In memoriam Sergei Mikhailovich Navashin]. PMID- 10712193 TI - James V. Neel, M.D., Ph.D. (March 22, 1915-January 31, 2000): founder effect. PMID- 10712194 TI - Genetic testing in adoption. The American Society of Human Genetics Social Issues Committee and The American College of Medical Genetics Social, Ethical, and Legal Issues Committee. PMID- 10712195 TI - Paternal origin of FGFR2 mutations in sporadic cases of Crouzon syndrome and Pfeiffer syndrome. AB - Crouzon syndrome and Pfeiffer syndrome are both autosomal dominant craniosynostotic disorders that can be caused by mutations in the fibroblast growth factor receptor 2 (FGFR2) gene. To determine the parental origin of these FGFR2 mutations, the amplification refractory mutation system (ARMS) was used. ARMS PCR primers were developed to recognize polymorphisms that could distinguish maternal and paternal alleles. A total of 4,374 bases between introns IIIa and 11 of the FGFR2 gene were sequenced and were assayed by heteroduplex analysis, to identify polymorphisms. Two polymorphisms (1333TA/TATA and 2710 C/T) were found and were used with two previously described polymorphisms, to screen a total of 41 families. Twenty-two of these families were shown to be informative (11 for Crouzon syndrome and 11 for Pfeiffer syndrome). Eleven different mutations in the 22 families were detected by either restriction digest or allele-specific oligonucleotide hybridization of ARMS PCR products. We molecularly proved the origin of these different mutations to be paternal for all informative cases analyzed (P=2. 4x10-7; 95% confidence limits 87%-100%). Advanced paternal age was noted for the fathers of patients with Crouzon syndrome or Pfeiffer syndrome, compared with the fathers of control individuals (34. 50+/-7.65 years vs. 30.45+/ 1.28 years, P<.01). Our data on advanced paternal age corroborates and extends previous clinical evidence based on statistical analyses as well as additional reports of advanced paternal age associated with paternal origin of three sporadic mutations causing Apert syndrome (FGFR2) and achondroplasia (FGFR3). Our results suggest that older men either have accumulated or are more susceptible to a variety of germline mutations. PMID- 10712196 TI - Characterization of the NPHP1 locus: mutational mechanism involved in deletions in familial juvenile nephronophthisis. AB - Familial juvenile nephronophthisis is an autosomal recessive, genetically heterogeneous kidney disorder representing the most frequent inherited cause of chronic renal failure in children. A gene, NPHP1, responsible for approximately 85% of the purely renal form of nephronophthisis, has been mapped to 2q13 and characterized. The major NPHP1 gene defect is a large homozygous deletion found in approximately 80% of the patients. In this study, by large-scale genomic sequencing and pulsed-field gel electrophoresis analysis, we characterized the complex organization of the NPHP1 locus and determined the mutational mechanism that results in the large deletion observed in most patients. We showed that the deletion is 290 kb in size and that NPHP1 is flanked by two large inverted repeats of approximately 330 kb. In addition, a second sequence of 45 kb located adjacent to the proximal 330-kb repeat was shown to be directly repeated 250 kb away within the distal 330-kb repeat deleting the sequence tag site (STS) 804H10R present in the proximal copy. The patients' deletion breakpoints appear to be located within the 45-kb repeat, suggesting an unequal recombination between the two homologous copies of this smaller repeat. Moreover, we demonstrated a nonpathologic rearrangement involving the two 330-kb inverted repeats found in 11 patients and, in the homozygous state, in 2 (1.3%) control individuals. This could be explained by interchromosomal mispairing of the 330-kb inverted repeat, followed by double recombination or by a prior intrachromosomal mispairing of these repeats, leading to an inversion of the NPHP1 region, followed by an interchromosomal unequal crossover event. This complex rearrangement, as well as the common deletion found in most patients, illustrates the high level of rearrangements occurring in the centromeric region of chromosome 2. PMID- 10712197 TI - Minor lesion mutational spectrum of the entire NF1 gene does not explain its high mutability but points to a functional domain upstream of the GAP-related domain. AB - More than 500 unrelated patients with neurofibromatosis type 1 (NF1) were screened for mutations in the NF1 gene. For each patient, the whole coding sequence and all splice sites were studied for aberrations, either by the protein truncation test (PTT), temperature-gradient gel electrophoresis (TGGE) of genomic PCR products, or, most often, by direct genomic sequencing (DGS) of all individual exons. A total of 301 sequence variants, including 278 bona fide pathogenic mutations, were identified. As many as 216 or 183 of the genuine mutations, comprising 179 or 161 different ones, can be considered novel when compared to the recent findings of Upadhyaya and Cooper, or to the NNFF mutation database. Mutation-detection efficiencies of the various screening methods were similar: 47.1% for PTT, 53.7% for TGGE, and 54.9% for DGS. Some 224 mutations (80.2%) yielded directly or indirectly premature termination codons. These mutations showed even distribution over the whole gene from exon 1 to exon 47. Of all sequence variants determined in our study, <20% represent C-->T or G-->A transitions within a CpG dinucleotide, and only six different mutations also occur in NF1 pseudogenes, with five being typical C-->T transitions in a CpG. Thus, neither frequent deamination of 5-methylcytosines nor interchromosomal gene conversion may account for the high mutation rate of the NF1 gene. As opposed to the truncating mutations, the 28 (10.1%) missense or single-amino-acid-deletion mutations identified clustered in two distinct regions, the GAP-related domain (GRD) and an upstream gene segment comprising exons 11-17. The latter forms a so called cysteine/serine-rich domain with three cysteine pairs suggestive of ATP binding, as well as three potential cAMP-dependent protein kinase (PKA) recognition sites obviously phosphorylated by PKA. Coincidence of mutated amino acids and those conserved between human and Drosophila strongly suggest significant functional relevance of this region, with major roles played by exons 12a and 15 and part of exon 16. PMID- 10712198 TI - An unstable trinucleotide-repeat region on chromosome 13 implicated in spinocerebellar ataxia: a common expansion locus. AB - Larger CAG/CTG trinucleotide-repeat tracts in individuals affected with schizophrenia (SCZ) and bipolar affective disorder (BPAD) in comparison with control individuals have previously been reported, implying a possible etiological role for trinucleotide repeats in these diseases. Two unstable CAG/CTG repeats, SEF2-1B and ERDA1, have recently been cloned, and studies indicate that the majority of individuals with large repeats as detected by repeat-expansion detection (RED) have large repeat alleles at these loci. These repeats do not show association of large alleles with either BPAD or SCZ. Using RED, we have identified a BPAD individual with a very large CAG/CTG repeat that is not due to expansion at SEF2-1B or ERDA1. From this individual's DNA, we have cloned a highly polymorphic trinucleotide repeat consisting of (CTA)n (CTG)n, which is very long ( approximately 1,800 bp) in this patient. The repeat region localizes to chromosome 13q21, within 1.2 cM of fragile site FRA13C. Repeat alleles in our sample were unstable in 13 (5.6%) of 231 meioses. Large alleles (>100 repeats) were observed in 14 (1. 25%) of 1,120 patients with psychosis, borderline personality disorder, or juvenile-onset depression and in 5 (.7%) of 710 healthy controls. Very large alleles were also detected for Centre d'Etude Polymorphisme Humaine (CEPH) reference family 1334. This triplet expansion has recently been reported to be the cause of spinocerebellar ataxia type 8 (SCA8); however, none of our large alleles above the disease threshold occurred in individuals either affected by SCA or with known family history of SCA. The high frequency of large alleles at this locus is inconsistent with the much rarer occurrence of SCA8. Thus, it seems unlikely that expansion alone causes SCA8; other genetic mechanisms may be necessary to explain SCA8 etiology. PMID- 10712199 TI - High germinal instability of the (CTG)n at the SCA8 locus of both expanded and normal alleles. AB - The autosomal dominant spinocerebellar ataxias (SCAs) are a group of late-onset, neurodegenerative disorders for which 10 loci have been mapped (SCA1, SCA2, SCA4 SCA8, SCA10, MJD, and DRPLA). The mutant proteins have shown an expanded polyglutamine tract in SCA1, SCA2, MJD/SCA3, SCA6, SCA7, and DRPLA; a glycine-to arginine substitution was found in SCA6 as well. Recently, an untranslated (CTG)n expansion on chromosome 13q was described as being the cause of SCA8. We have now (1) assessed the repeat size in a group of patients with ataxia and a large number of controls, (2) examined the intergenerational transmission of the repeat, and (3) estimated the instability of repeat size in the sperm of one patient and two healthy controls. Normal SCA8 chromosomes showed an apparently trimodal distribution, with classes of small (15-21 CTGs), intermediate (22-37 CTGs), and large (40-91 CTGs) alleles; large alleles accounted for only0.7% of all normal-size alleles. No expanded alleles (>/=100 CTGs) were found in controls. Expansion of the CTG tract was found in five families with ataxia; expanded alleles (all paternally transmitted) were characterized mostly by repeat size contraction. There was a high germinal instability of both expanded and normal alleles: in one patient, the expanded allele (152 CTGs) had mostly contraction in size (often into the normal range); in the sperm of two normal controls, contractions were also more frequent, but occasional expansions into the upper limit of the normal size range were also seen. In conclusion, our results show (1) no overlapping between control (15-91) and pathogenic (100-152) alleles and (2) a high instability in spermatogenesis (both for expanded and normal alleles), suggesting a high mutational rate at the SCA8 locus. PMID- 10712200 TI - Relaxation of insulin-like growth factor 2 imprinting and discordant methylation at KvDMR1 in two first cousins affected by Beckwith-Wiedemann and Klippel Trenaunay-Weber syndromes. AB - Beckwith-Wiedeman syndrome (BWS) and Klippel-Trenaunay-Weber syndrome (KTWS) are different human disorders characterized, among other features, by tissue overgrowth. Deregulation of one or more imprinted genes located at chromosome 11p15.5, of which insulin-like growth factor 2 (IGF2) is the most likely candidate, is believed to cause BWS, whereas the etiology of KTWS is completely obscure. We report a case of BWS and a case of KTWS in a single family. The probands, sons of two sisters, showed relaxation of the maternal IGF2 imprinting, although they inherited different 11p15.5 alleles from their mothers and did not show any chromosome rearrangement. The patient with BWS also displayed hypomethylation at KvDMR1, a maternally methylated CpG island within an intron of the KvLQT1 gene. The unaffected brother of the BWS proband shared the same maternal and paternal 11p15.5 haplotype with his brother, but the KvDMR1 locus was normally methylated. Methylation of the H19 gene was normal in both the BWS and KTWS probands. Linkage between the insulin-like growth factor 2 receptor (IGF2R) gene and the tissue overgrowth was also excluded. These results raise the possibility that a defective modifier or regulatory gene unlinked to 11p15.5 caused a spectrum of epigenetic alterations in the germ line or early development of both cousins, ranging from the relaxation of IGF2 imprinting in the KTWS proband to disruption of both the imprinted expression of IGF2 and the imprinted methylation of KvDMR1 in the BWS proband. Analysis of these data also indicates that loss of IGF2 imprinting is not necessarily linked to alteration of methylation at the KvDMR1 or H19 loci and supports the notion that IGF2 overexpression is involved in the etiology of the tissue hypertrophy observed in different overgrowth disorders, including KTWS. PMID- 10712201 TI - Identification of novel imprinted transcripts in the Prader-Willi syndrome and Angelman syndrome deletion region: further evidence for regional imprinting control. AB - Deletions and other abnormalities of human chromosome 15q11-q13 are associated with two developmental disorders, Prader-Willi syndrome (PWS) and Angelman syndrome (AS). Loss of expression of imprinted, paternally expressed genes has been implicated in PWS. However, the number of imprinted genes that contribute to PWS, and the range over which the imprinting signal acts to silence one copy of the gene in a parent-of-origin-specific manner, are unknown. To identify additional imprinted genes that could contribute to the PWS phenotype and to understand the regional control of imprinting in 15q11-q13, we have constructed an imprinted transcript map of the PWS-AS deletion interval. The imprinting status of 22 expressed sequence tags derived from the radiation-hybrid human transcript maps or physical maps was determined in a reverse transcriptase-PCR assay and correlated with the position of the transcripts on the physical map. Seven new paternally expressed transcripts localize to an approximately 1.5-Mb domain surrounding the SNRPN-associated imprinting center, which already includes four imprinted, paternally expressed genes. All other tested new transcripts in the deletion region were expressed from both alleles. A domain of exclusive paternal expression surrounding the imprinting center suggests strong regional control of the imprinting process. This study provides the means for further investigation of additional genes that cause or modify the phenotypes associated with rearrangements of 15q11-q13. PMID- 10712202 TI - Decreased elastin deposition and high proliferation of fibroblasts from Costello syndrome are related to functional deficiency in the 67-kD elastin-binding protein. AB - Costello syndrome is characterized by mental retardation, loose skin, coarse face, skeletal deformations, cardiomyopathy, and predisposition to numerous malignancies. The genetic origin of Costello syndrome has not yet been defined. Using immunohistochemistry and metabolic labeling with [3H]-valine, we have established that cultured skin fibroblasts obtained from patients with Costello syndrome did not assemble elastic fibers, despite an adequate synthesis of tropoelastin and normal deposition of the microfibrillar scaffold. We found that impaired production of elastic fibers by these fibroblasts is associated with a functional deficiency of the 67-kD elastin-binding protein (EBP), which is normally required to chaperone tropoelastin through the secretory pathways and to its extracellular assembly. Metabolic pulse labeling of the 67-kD EBP with radioactive serine and further chase of this tracer indicated that both normal fibroblasts and fibroblasts from patients with Costello syndrome initially synthesized comparable amounts of this protein; however, the fibroblasts from Costello syndrome patients quickly lost it into the conditioned media. Because the normal association between EBP and tropoelastin can be disrupted on contact with galactosugar-bearing moieties, and the fibroblasts from patients with Costello syndrome revealed an unusual accumulation of chondroitin sulfate-bearing proteoglycans (CD44 and biglycan), we postulate that a chondroitin sulfate may be responsible for shedding EBP from Costello cells and in turn for their impaired elastogenesis. This was further supported by the fact that exposure to chondroitinase ABC, an enzyme capable of chondroitin sulfate degradation, restored normal production of elastic fibers by fibroblasts from patients with Costello syndrome. We also present evidence that loss of EBP from fibroblasts of Costello syndrome patients is associated with an unusually high rate of cellular proliferation. PMID- 10712204 TI - A locus for brachydactyly type A-1 maps to chromosome 2q35-q36. AB - Brachydactyly type A-1 (BDA1) was, in 1903, the first recorded example of a human anomaly with Mendelian autosomal dominant inheritance. Two large families, the affected members of which were radiographed, were recruited in the study we describe here. Two-point linkage analysis for pedigree 1 (maximum LOD score [Zmax] 6.59 at recombination fraction [theta] 0.00) and for pedigree 2 (Zmax=5.53 at straight theta=0.00) mapped the locus for BDA1 in the two families to chromosome 2q. Haplotype analysis of pedigree 1 confined the locus for family 1 within an interval of <8.1 cM flanked by markers D2S2248 and D2S360, which was mapped to chromosome 2q35-q36 on the cytogenetic map. Haplotype analysis of pedigree 2 confined the locus for family 2 within an interval of <28. 8 cM flanked by markers GATA30E06 and D2S427, which was localized to chromosome 2q35 q37. The two families had no identical haplotype within the defined region, which suggests that the two families were not related. PMID- 10712203 TI - Functional analysis of the neurofibromatosis type 2 protein by means of disease causing point mutations. AB - Despite intense study of the neurofibromatosis type 2 (NF2) tumor-suppressor protein merlin, the biological properties and tumor-suppressor functions of merlin are still largely unknown. In this study, we examined the molecular activities of NF2-causing mutant merlin proteins in transfected mammalian cells, to elucidate the merlin properties that are critical for tumor-suppressor function. Most important, we found that 80% of the merlin mutants studied significantly altered cell adhesion, causing cells to detach from the substratum. This finding implies a function for merlin in regulating cell-matrix attachment, and changes in cell adhesion caused by mutant protein expression may be an initial step in the pathogenesis of NF2. In addition, five different mutations in merlin caused a significant increase in detergent solubility of merlin compared to wild type, indicating a decreased ability to interact with the cytoskeleton. Although not correlated to the cell-adhesion phenotype, four missense mutations decreased the binding of merlin to the ERM-interacting protein EBP-50, implicating this interaction in merlin inhibition of cell growth. Last, we found that some NF2 point mutations in merlin most closely resembled gain-of-function alleles in their cellular phenotype, which suggests that mutant NF2 alleles may not always act in a loss-of-function manner, as had been assumed, but may include a spectrum of allelic types with different phenotypic effects on the function of the protein. In aggregate, these cellular phenotypes provide a useful assay for identifying the functional domains and molecular partners necessary for merlin tumor-suppressor activity. PMID- 10712205 TI - Two new loci for autosomal recessive ichthyosis on chromosomes 3p21 and 19p12-q12 and evidence for further genetic heterogeneity. AB - Autosomal recessive ichthyosis (ARI) includes a heterogeneous group of disorders of keratinization characterized by desquamation over the whole body. Two forms largely limited to the skin have been defined: lamellar ichthyosis (LI) and nonbullous congenital ichthyosiform erythroderma (NCIE). A first gene for LI, transglutaminase TGM1, has been identified on chromosome 14, and a second one has been localized on chromosome 2. In a genomewide scan of nine large consanguineous families, using homozygosity mapping, two new loci for ARI were found, one for a lamellar form in a 6-cM interval on chromosome 19 and a second for an erythrodermic form in a 7.7-cM interval on chromosome 3. Linkage to one of the four loci could be demonstrated in more than half of 51 consanguineous families, most of them from the Mediterranean basin. All four loci could be excluded in the others, implying further genetic heterogeneity in this disorder. Multipoint linkage analysis gave maximal LOD scores of 11.25 at locus D19S566 and 8.53 at locus D3S3564. PMID- 10712206 TI - Localization of the Netherton syndrome gene to chromosome 5q32, by linkage analysis and homozygosity mapping. AB - Netherton syndrome (NS [MIM 256500]) is a rare and severe autosomal recessive disorder characterized by congenital ichthyosis, a specific hair-shaft defect (trichorrhexis invaginata), and atopic manifestations. Infants with this syndrome often fail to thrive; life-threatening complications result in high postnatal mortality. We report the assignment of the NS gene to chromosome 5q32, by linkage analysis and homozygosity mapping in 20 families affected with NS. Significant evidence for linkage (maximum multipoint LOD score 10.11) between markers D5S2017 and D5S413 was obtained, with no evidence for locus heterogeneity. Analysis of critical recombinants mapped the NS locus between markers D5S463 and D5S2013, within an <3.5-cM genetic interval. The NS locus is telomeric to the cytokine gene cluster in 5q31. The five known genes encoding casein kinase Ialpha, the alpha subunit of retinal rod cGMP phosphodiesterase, the regulator of mitotic spindle assembly, adrenergic receptor beta2, and the diastrophic dysplasia sulfate-transporter gene, as well as the 38 expressed-sequence tags mapped within the critical region, are not obvious candidates. Our study is the first step toward the positional cloning of the NS gene. This finding promises a better understanding of the molecular mechanisms that control epidermal differentiation and immunity. PMID- 10712207 TI - Fine mapping of the chromosome 12 late-onset Alzheimer disease locus: potential genetic and phenotypic heterogeneity. AB - Apolipoprotein E (APOE) is the only confirmed susceptibility gene for late-onset Alzheimer disease (AD). In a recent genomic screen of 54 families with late-onset AD, we detected significant evidence for a second late-onset AD locus located on chromosome 12 between D12S373 and D12S390. Linkage to this region was strongest in 27 large families with at least one affected individual without an APOE-4 allele, suggesting that APOE and the chromosome 12 locus might have independent effects. We have since genotyped several additional markers across the region, to refine the linkage results. In analyzing these additional data, we have addressed the issue of heterogeneity in the data set by weighting results by clinical and neuropathologic features, sibship size, and APOE genotype. When considering all possible affected sib pairs (ASPs) per nuclear family, we obtained a peak maximum LOD score between D12S1057 and D12S1042. The magnitude and location of the maximum LOD score changed when different weighting schemes were used to control for the number of ASPs contributed by each nuclear family. Using the affected relative-pair method implemented in GENEHUNTER-PLUS, we obtained a maximum LOD score between D12S398 and D12S1632, 25 cM from the original maximum LOD score. These results indicate that family size influences the location estimate for the chromosome 12 AD gene. The results of conditional linkage analysis by use of GENEHUNTER-PLUS indicated that evidence for linkage to chromosome 12 was stronger in families with affected individuals lacking an APOE-4 allele; much of this evidence came from families with affected individuals with neuropathologic diagnosis of dementia with Lewy bodies (DLB). Taken together, these results indicate that the chromosome 12 locus acts independently of APOE to increase the risk of late-onset familial AD and that it may be associated with the DLB variant of AD. PMID- 10712209 TI - Combined analysis of hereditary prostate cancer linkage to 1q24-25: results from 772 hereditary prostate cancer families from the International Consortium for Prostate Cancer Genetics. AB - A previous linkage study provided evidence for a prostate cancer-susceptibility locus at 1q24-25. Subsequent reports in additional collections of families have yielded conflicting results. In addition, evidence for locus heterogeneity has been provided by the identification of other putative hereditary prostate cancer loci on Xq27-28, 1q42-43, and 1p36. The present study describes a combined analysis for six markers in the 1q24-25 region in 772 families affected by hereditary prostate cancer and ascertained by the members of the International Consortium for Prostate Cancer Genetics (ICPCG) from North America, Australia, Finland, Norway, Sweden, and the United Kingdom. Overall, there was some evidence for linkage, with a peak parametric multipoint LOD score assuming heterogeneity (HLOD) of 1.40 (P=.01) at D1S212. The estimated proportion of families (alpha) linked to the locus was.06 (1-LOD support interval.01-.12). This evidence was not observed by a nonparametric approach, presumably because of the extensive heterogeneity. Further parametric analysis revealed a significant effect of the presence of male-to-male disease transmission within the families. In the subset of 491 such families, the peak HLOD was 2.56 (P=.0006) and alpha =.11 (1-LOD support interval.04-.19), compared with HLODs of 0 in the remaining 281 families. Within the families with male-to-male disease transmission, alpha increased with the early mean age at diagnosis (<65 years, alpha =.19, with 1-LOD support interval.06-.34) and the number of affected family members (five or more family members, alpha =.15, with 1-LOD support interval.04-.28). The highest value of alpha was observed for the 48 families that met all three criteria (peak HLOD = 2.25, P=.001, alpha=.29, with 1-LOD support interval.08-.53). These results support the finding of a prostate cancer-susceptibility gene linked to 1q24-25, albeit in a defined subset of prostate cancer families. Although HPC1 accounts for only a small proportion of all families affected by hereditary prostate cancer, it appears to play a more prominent role in the subset of families with several members affected at an early age and with male-to-male disease transmission. PMID- 10712208 TI - A genome screen of multiplex sibships with prostate cancer. AB - Analysis of a genome screen of 504 brothers with prostate cancer (CaP) who were from 230 multiplex sibships identified five regions with nominally positive linkage signals, on chromosomes 2q, 12p, 15q, 16p, and 16q. The strongest signal in these data is found on chromosome 16q, between markers D16S515 and D16S3040, a region suspected to contain a tumor-suppressor gene. On the basis of findings from previous genome screens of families with CaP, three preplanned subanalyses were carried out, in the hope of increasing the subgroup homogeneity. Subgroups were formed by dividing the sibships into a group with a positive family history (FH+) that met criteria for "hereditary" CaP (n=111) versus those which did not meet the criteria (n=119) and by dividing the families into those with a mean onset age below the median (n=115) versus those with a mean onset age above the median (n=115). A separate subanalysis was carried out for families with a history of breast cancer (CaB+ [n=53]). Analyses of these subgroups revealed a number of potentially important differences in regions that were nonsignificant when all the families were analyzed together. In particular, the subgroup without a positive family history (FH-) had a signal in a region that is proximal to the putative site of the HPC1 locus on chromosome 1, whereas the late-age-at-onset group had a signal on 4q. The CaB+ subgroup revealed a strong linkage signal at 1p35.1. PMID- 10712210 TI - Multipoint estimation of genetic maps for human trisomies with one parent or other partial data. AB - Centromeric-mapping methods have been used to investigate the association between altered recombination and meiotic nondisjunction in humans. For trisomies, current methods are based on the genotypes from a trisomic offspring and both parents. Because it is sometimes difficult to obtain samples from both parents and because the ability to use sources of DNA previously not available (e.g., stored paraffin-embedded pathological samples) has increased, we have been interested in creating similar maps for trisomic populations in which one of the parents of the trisomic individual is unavailable for genotyping. In this paper, we derive multipoint likelihoods for both missing-parent data and conventional two-parent data. We find that likelihoods for two-parent data and for data generated without a sample from the correctly disjoining parent can be maximized in exactly the same way but also that missing-parent data has a high frequency of partial data of the same sort produced by intercross matings. Previously published centromeric-mapping methods use incorrect likelihoods for intercross matings and thus can perform poorly on missing-parent data. We wrote a FORTRAN program to maximize our multipoint likelihoods and used it in simulation studies to demonstrate the biases in the previous methods. PMID- 10712211 TI - Significant admixture linkage disequilibrium across 30 cM around the FY locus in African Americans. AB - Scientists, to understand the importance of allelic polymorphisms on phenotypes that are quantitative and environmentally interacting, are now turning to population-association screens, especially in instances in which pedigree analysis is difficult. Because association screens require linkage disequilibrium between markers and disease loci, maximizing the degree of linkage disequilibrium increases the chances of discovering functional gene-marker associations. One theoretically valid approach-mapping by admixture linkage disequilibrium (MALD), using recently admixed African Americans-is empirically evaluated here by measurement of marker associations with 15 short tandem repeats (STRs) and an insertion/deletion polymorphism of the AT3 locus in a 70-cM segment at 1q22-23, around the FY (Duffy) locus. The FY polymorphism (-46T-->C) disrupts the GATA promoter motif, specifically blocking FY erythroid expression and has a nearly fixed allele-frequency difference between European Americans and native Africans that is likely a consequence of a selective advantage of FY-/- in malaria infections. Analysis of linkage disequilibrium around the FY gene has indicated that there is strong and consistent linkage disequilibrium between FY and three flanking loci (D1S303, SPTA1, and D1S484) spanning 8 cM. We observed significant linkage-disequilibrium signals over a 30-cM region from -4.4 to 16.3 cM (from D1S2777 to D1S196) for STRs and at 26.4 cM (AT3), which provided quantitative estimates of centimorgan limits, by MALD assessment in African American population-association analyses, of 5-10 cM. PMID- 10712213 TI - mtDNA affinities of the peoples of North-Central Mexico. AB - mtDNA haplotypes of representatives of the cosmopolitan peoples of north-central Mexico were studied. Two hundred twenty-three samples from individuals residing in vicinities of two localities in north-central Mexico were analyzed. A combination of strategies was employed to identify the origin of each haplotype, including length variation analysis of the COII and tRNALYS intergenic region, nucleotide sequence analysis of control region hypervariable segment 1, and RFLP analysis of PCR products spanning diagnostic sites. Analysis of these data revealed that the majority of the mtDNA haplotypes were of Native American origin, belonging to one of four primary Native American haplogroups. Others were of European or African origin, and the frequency of African haplotypes was equivalent to that of haplotypes of European derivation. These results provide diagnostic, discrete character, molecular genetic evidence that, together with results of previous studies of classical genetic systems, is informative with regard to both the magnitude of African admixture and the relative maternal contribution of African, European, and Native American peoples to the genetic heritage of Mexico. Phylogenetic analysis revealed that African sequences formed a basal, paraphyletic group. PMID- 10712212 TI - The distribution of human genetic diversity: a comparison of mitochondrial, autosomal, and Y-chromosome data. AB - We report a comparison of worldwide genetic variation among 255 individuals by using autosomal, mitochondrial, and Y-chromosome polymorphisms. Variation is assessed by use of 30 autosomal restriction-site polymorphisms (RSPs), 60 autosomal short-tandem-repeat polymorphisms (STRPs), 13 Alu-insertion polymorphisms and one LINE-1 element, 611 bp of mitochondrial control-region sequence, and 10 Y-chromosome polymorphisms. Analysis of these data reveals substantial congruity among this diverse array of genetic systems. With the exception of the autosomal RSPs, in which an ascertainment bias exists, all systems show greater gene diversity in Africans than in either Europeans or Asians. Africans also have the largest total number of alleles, as well as the largest number of unique alleles, for most systems. GST values are 11%-18% for the autosomal systems and are two to three times higher for the mtDNA sequence and Y-chromosome RSPs. This difference is expected because of the lower effective population size of mtDNA and Y chromosomes. A lower value is seen for Y chromosome STRs, reflecting a relative lack of continental population structure, as a result of rapid mutation and genetic drift. Africa has higher GST values than does either Europe or Asia for all systems except the Y-chromosome STRs and Alus. All systems except the Y-chromosome STRs show less variation between populations within continents than between continents. These results are reassuring in their consistency and offer broad support for an African origin of modern human populations. PMID- 10712214 TI - mtDNA and the origin of the Icelanders: deciphering signals of recent population history. AB - Previous attempts to investigate the origin of the Icelanders have provided estimates of ancestry ranging from a 98% British Isles contribution to an 86% Scandinavian contribution. We generated mitochondrial sequence data for 401 Icelandic individuals and compared these data with >2,500 other European sequences from published sources, to determine the probable origins of women who contributed to Iceland's settlement. Although the mean number of base-pair differences is high in the Icelandic sequences and they are widely distributed in the overall European mtDNA phylogeny, we find a smaller number of distinct mitochondrial lineages, compared with most other European populations. The frequencies of a number of mtDNA lineages in the Icelanders deviate noticeably from those in neighboring populations, suggesting that founder effects and genetic drift may have had a considerable influence on the Icelandic gene pool. This is in accordance with available demographic evidence about Icelandic population history. A comparison with published mtDNA lineages from European populations indicates that, whereas most founding females probably originated from Scandinavia and the British Isles, lesser contributions from other populations may also have taken place. We present a highly resolved phylogenetic network for the Icelandic data, identifying a number of previously unreported mtDNA lineage clusters and providing a detailed depiction of the evolutionary relationships between European mtDNA clusters. Our findings indicate that European populations contain a large number of closely related mitochondrial lineages, many of which have not yet been sampled in the current comparative data set. Consequently, substantial increases in sample sizes that use mtDNA data will be needed to obtain valid estimates of the diverse ancestral mixtures that ultimately gave rise to contemporary populations. PMID- 10712215 TI - Phylogenetic network of the mtDNA haplogroup U in Northern Finland based on sequence analysis of the complete coding region by conformation-sensitive gel electrophoresis. AB - Mutations in mtDNA have accumulated sequentially, and maternal lineages have diverged to form population-specific genotypes. Classification of the genotypes has been made based on differences found in restriction fragment analysis of the coding region or in the sequence of the hypervariable segment I. Both methods have shortcomings, as the former may not detect all the important polymorphisms and the latter makes use of a segment containing hypervariable nucleotide positions. Here, we have used conformation-sensitive gel electrophoresis (CSGE) to detect polymorphisms within the coding region of mtDNA from 22 Finns belonging to haplogroup U. Sixty-three overlapping PCR fragments covering the entire coding region were analyzed by CSGE, and the fragments that differed in their migration pattern were sequenced. CSGE proved to be a sensitive and specific method for identifying mtDNA substitutions. The phylogenetic network of the 22 coding-region sequences constituted a perfect tree, free of homoplasy, and provided several previously unidentified common polymorphisms characterizing subgroups of U. After contrasting this data with that of hypervariable segment I, we concluded that position 16192 seems to be prone to recurrent mutations and that position 16270 has experienced a back mutation. Interestingly, all 22 samples were found to belong to subcluster U5, suggesting that this subcluster is more frequent in Finns than in other European populations. Complete sequence data of the mtDNA yield a more reliable phylogenetic network and a more accurate classification of the haplogroups than previous ones. In medical genetics, such networks may help to decide between a rare polymorphism and a pathogenic mutation; in population genetics, the networks may enable more detailed analyses of population history and mtDNA evolution. PMID- 10712216 TI - QTL fine mapping by measuring and testing for Hardy-Weinberg and linkage disequilibrium at a series of linked marker loci in extreme samples of populations. AB - It has recently been demonstrated that fine-scale mapping of a susceptibility locus for a complex disease can be accomplished on the basis of deviations from Hardy-Weinberg (HW) equilibrium at closely linked marker loci among affected individuals. We extend this theory to fine-scale localization of a quantitative trait locus (QTL) from extreme individuals in populations, by means of HW and linkage-disequilibrium (LD) analyses. QTL mapping and/or linkage analyses can establish a large genomic region ( approximately 30 cM) that contains a QTL. The QTL can be fine mapped by examination of the degree of deviation from HW and LD at a series of closely linked marker loci. The tests can be performed for samples of individuals belonging to either high or low percentiles of the phenotype distribution or for combined samples of these extreme individuals. The statistical properties (the power and the size) of the tests of this fine-mapping approach are investigated and are compared extensively, under various genetic models and parameters for the QTL and marker loci. On the basis of the results, a two-stage procedure that uses extreme samples and different tests (for HW and LD) is suggested for QTL fine mapping. This two-step procedure is economic and powerful and can accurately narrow a genomic region containing a QTL from approximately 30-1 cM, a range that renders physical mapping feasible for identification of the QTL. In addition, the relationship between parameterizations of complex diseases, by means of penetrance, and those of complex quantitative traits, by means of genotypic values, is outlined. This means that many statistical genetic methods developed for searching for susceptibility loci of complex diseases can be directly adopted and/or extended to QTL mapping for quantitative traits. PMID- 10712217 TI - A two-stage variable-stringency semiparametric method for mapping quantitative trait loci with the use of genomewide-scan data on sib pairs. AB - Genomewide scans for mapping loci have proved to be extremely powerful and popular. We present a semiparametric method of mapping a quantitative-trait locus (QTL) or QTLs with the use of sib-pair data generated from a two-stage genomic scan. In a two-stage genomic scan, either the entire genome or a large portion of the genome is saturated with low-density markers at the first stage. At the second stage, the intervals that are identified as probable locations of the trait loci, by means of analysis of data from the first stage, are then saturated with higher-density markers. These data are then analyzed for fine mapping of the loci. Our statistical strategy for analysis of data from the first stage is a low stringency method based on the rank correlation of squared trait-difference values of the sib pairs and the estimated identity-by-descent scores at the marker loci. We suggest the use of a low-stringency method at the first stage, to save on computational time and to avoid missing any marker interval that may contain the trait loci. For analysis of data from the second stage, we have developed a high-stringency nonparametric-regression approach, using the kernel smoothing technique. Through extensive simulations, we show that this approach is more powerful than is a currently used method for mapping QTLs by use of sib pairs, particularly in the presence of dominance and epistatic effects at the trait loci. PMID- 10712218 TI - The trimmed-haplotype test for linkage disequilibrium. AB - Single-marker linkage-disequilibrium (LD) methods cannot fully describe disequilibrium in an entire chromosomal region surrounding a disease allele. With the advent of myriad tightly linked microsatellite markers, we have an opportunity to extend LD analysis from single markers to multiple-marker haplotypes. Haplotype analysis has increased statistical power to disclose the presence of a disease locus in situations where it correctly reflects the historical process involved. For maximum efficiency, evidence of LD ought to come not just from a single haplotype, which may well be rare, but in addition from many similar haplotypes that could have descended from the same ancestral founder but have been trimmed in succeeding generations. We present such an analysis, called the "trimmed-haplotype method." We focus on chromosomal regions that are small enough that disequilibrium in significant portions of them may have been preserved in some pedigrees and yet that contain enough markers to minimize coincidental occurrence of the haplotype in the absence of a disease allele: perhaps regions 1-2 cM in length. In general, we could have no idea what haplotype an ancestral founder carried generations ago, nor do we usually have a precise chromosomal location for the disease-susceptibility locus. Therefore, we must search through all possible haplotypes surrounding multiple locations. Since such repeated testing obliterates the sampling distribution of the test, we employ bootstrap methods to calculate significance levels. Trimmed-haplotype analysis is performed on family data in which genotypes have been assembled into haplotypes. It can be applied either to conventional parent-affected-offspring triads or to multiplex pedigrees. We present a method for summarizing the LD evidence, in any pedigree, that can be employed in trimmed-haplotype analysis as well as in other methods. PMID- 10712219 TI - Statistical tests for detection of misspecified relationships by use of genome screen data. AB - Misspecified relationships can have serious consequences for linkage studies, resulting in either reduced power or false-positive evidence for linkage. If some individuals in the pedigree are untyped, then Mendelian errors may not be observed. Previous approaches to detection of misspecified relationships by use of genotype data were developed for sib and half-sib pairs. We extend the likelihood calculations of Goring and Ott and Boehnke and Cox to more-general relative pairs, for which identity-by-descent (IBD) status is no longer a Markov chain, and we propose a likelihood-ratio test. We also extend the identity-by state (IBS)-based test of Ehm and Wagner to nonsib relative pairs. The likelihood ratio test has high power, but its drawbacks include the need to construct and apply a separate Markov chain for each possible alternative relationship and the need for simulation to assess significance. The IBS-based test is simpler but has lower power. We propose two new test statistics-conditional expected IBD (EIBD) and adjusted IBS (AIBS)-designed to retain the simplicity of IBS while increasing power by taking into account chance sharing. In simulations, the power of EIBD is generally close to that of the likelihood-ratio test. The power of AIBS is higher than that of IBS, in all cases considered. We suggest a strategy of initial screening by use of EIBD and AIBS, followed by application of the likelihood ratio test to only a subset of relative pairs, identified by use of EIBD and AIBS. We apply the methods to a Genetic Analysis Workshop 11 data set from the Collaborative Study on the Genetics of Alcoholism. PMID- 10712220 TI - Linkage analysis in the presence of errors I: complex-valued recombination fractions and complex phenotypes. AB - Linkage is a phenomenon that correlates the genotypes of loci, rather than the phenotypes of one locus to the genotypes of another. It is therefore necessary to convert the observed trait phenotypes into trait-locus genotypes, which can then be analyzed for coinheritance with marker-locus genotypes. However, if the mode of inheritance of the trait is not known accurately, this conversion can often result in errors in the inferred trait-locus genotypes, which, in turn, can lead to the misclassification of the recombination status of meioses. As a result, the recombination fraction can be overestimated in two-point analysis, and false exclusions of the true trait locus can occur in multipoint analysis. We propose a method that increases the robustness of multipoint analysis to errors in the mode of inheritance assumptions of the trait, by explicitly allowing for misclassification of trait-locus genotypes. To this end, the definition of the recombination fraction is extended to the complex plane, as Theta=straight theta+straightepsiloni; theta is the recombination fraction between actual ("real") genotypes of marker and trait loci, and straightepsilon is the probability of apparent but false ("imaginary") recombinations between the actual and inferred trait-locus genotypes. "Complex" multipoint LOD scores are proven to be stochastically equivalent to conventional two-point LOD scores. The greater robustness to modeling errors normally associated with two-point analysis can thus be extended to multiple two-point analysis and multipoint analysis. The use of complex-valued recombination fractions also allows the stochastic equivalence of "model-based" and "model-free" methods to be extended to multipoint analysis. PMID- 10712221 TI - Linkage analysis in the presence of errors II: marker-locus genotyping errors modeled with hypercomplex recombination fractions. AB - It is well known that genotyping errors lead to loss of power in gene-mapping studies and underestimation of the strength of correlations between trait- and marker-locus genotypes. In two-point linkage analysis, these errors can be absorbed in an inflated recombination-fraction estimate, leaving the test statistic quite robust. In multipoint analysis, however, genotyping errors can easily result in false exclusion of the true location of a disease-predisposing gene. In a companion article, we described a "complex-valued" extension of the recombination fraction to accommodate errors in the assignment of trait-locus genotypes, leading to a multipoint LOD score with the same robustness to errors in trait-locus genotypes that is seen with the conventional two-point LOD score. Here, a further extension of this model to "hypercomplex-valued" recombination fractions (hereafter referred to as "hypercomplex recombination fractions") is presented, to handle random and systematic sources of marker-locus genotyping errors. This leads to a multipoint method (either "model-based" or "model-free") with the same robustness to marker-locus genotyping errors that is seen with conventional two-point analysis but with the advantage that multiple marker loci can be used jointly to increase meiotic informativeness. The cost of this increased robustness is a decrease in fine-scale resolution of the estimated map location of the trait locus, in comparison with traditional multipoint analysis. This probability model further leads to algorithms for the estimation of the lower bounds for the error rates for genomewide and locus-specific genotyping, based on the null-hypothesis distribution of the LOD-score statistic in the presence of such errors. It is argued that those genome scans in which the LOD score is 0 for >50% of the genome are likely to be characterized by high rates of genotyping errors in general. PMID- 10712222 TI - Bias and efficiency in family-based gene-characterization studies: conditional, prospective, retrospective, and joint likelihoods. AB - We revisit the usual conditional likelihood for stratum-matched case-control studies and consider three alternatives that may be more appropriate for family based gene-characterization studies: First, the prospective likelihood, that is, Pr(D/G,A second, the retrospective likelihood, Pr(G/D); and third, the ascertainment-corrected joint likelihood, Pr(D,G/A). These likelihoods provide unbiased estimators of genetic relative risk parameters, as well as population allele frequencies and baseline risks. The parameter estimates based on the retrospective likelihood remain unbiased even when the ascertainment scheme cannot be modeled, as long as ascertainment only depends on families' phenotypes. Despite the need to estimate additional parameters, the prospective, retrospective, and joint likelihoods can lead to considerable gains in efficiency, relative to the conditional likelihood, when estimating genetic relative risk. This is true if baseline risks and allele frequencies can be assumed to be homogeneous. In the presence of heterogeneity, however, the parameter estimates assuming homogeneity can be seriously biased. We discuss the extent of this problem and present a mixed models approach for providing consistent parameter estimates when baseline risks and allele frequencies are heterogeneous. The efficiency gains of the mixed-model prospective, retrospective, and joint likelihoods relative to the efficiency of conditional likelihood are small in the situations presented here. PMID- 10712223 TI - Assignment of a novel locus for autosomal recessive congenital ichthyosis to chromosome 19p13.1-p13.2. AB - Autosomal recessive congenital ichthyosis (ARCI) is a rare, clinically and genetically heterogeneous genodermatosis. One gene (transglutaminase 1, on 14q11) and one additional locus (on 2q33-35, with an unidentified gene) have been shown to be associated with a lamellar, nonerythrodermic type of ARCI. We performed a genomewide scan, with 370 highly polymorphic microsatellite markers, on five affected individuals from one large Finnish family with nonerythrodermic, nonlamellar ARCI. The only evidence for linkage emerged from markers in a 6.0-cM region on chromosome 19p13.1-2. The maximum two-point LOD score of 7.33 was obtained with the locus D19S252, and multipoint likelihood calculations gave a maximum location score of 5.2. The affected individuals share two common core haplotypes, which makes compound heterozygosity possible. The novel disease locus is the third locus linked to ARCI, supporting previous evidence for genetic heterogeneity of ARCI. This is also the first locus for a nonlamellar, nonerythrodermic phenotype of ARCI. PMID- 10712224 TI - Cloning, sequencing, and analysis of inv8 chromosome breakpoints associated with recombinant 8 syndrome. AB - Rec8 syndrome (also known as "recombinant 8 syndrome" and "San Luis Valley syndrome") is a chromosomal disorder found in individuals of Hispanic descent with ancestry from the San Luis Valley of southern Colorado and northern New Mexico. Affected individuals typically have mental retardation, congenital heart defects, seizures, a characteristic facial appearance, and other manifestations. The recombinant chromosome is rec(8)dup(8q)inv(8)(p23.1q22.1), and is derived from a parental pericentric inversion, inv(8)(p23.1q22.1). Here we report on the cloning, sequencing, and characterization of the 8p23.1 and 8q22 breakpoints from the inversion 8 chromosome associated with Rec8 syndrome. Analysis of the breakpoint regions indicates that they are highly repetitive. Of 6 kb surrounding the 8p23.1 breakpoint, 75% consists of repetitive gene family members-including Alu, LINE, and LTR elements-and the inversion took place in a small single-copy region flanked by repetitive elements. Analysis of 3.7 kb surrounding the 8q22 breakpoint region reveals that it is 99% repetitive and contains multiple LTR elements, and that the 8q inversion site is within one of the LTR elements. PMID- 10712225 TI - A recurrent expansion of a maternal allele with 36 CAG repeats causes Huntington disease in two sisters. AB - Large intergenerational repeat expansions of the CAG trinucleotide repeat in the HD gene have been well documented for the male germline. We describe a recurrent large expansion of a maternal allele with 36 CAG repeats (to 66 and 57 repeats, respectively, in two daughters) associated with onset of Huntington disease (HD) in the second and third decade in a family without history of HD. Our findings give evidence of a gonadal mosaicism in the unaffected mother. We hypothesize that large expansions also occur in the female germline and that a negative selection of oocytes with long repeats might explain the different instability behavior of the male and the female germlines. PMID- 10712226 TI - Analysis of aneuploidy frequencies in sperm from patients with hereditary nonpolyposis colon cancer and an hMSH2 mutation. AB - Hereditary nonpolyposis colon cancer (HNPCC) has been shown to be caused by mutations in the mismatch repair genes hMSH2, hMLH1, hPMS1, and hPMS2. Recent evidence has demonstrated that mutations in mismatch repair genes disrupt meiosis in mice. A large HNPCC kindred in Newfoundland, Canada, has an hMSH2 mutation-an A-->T transversion at the +3 position of the splice-donor site of exon 5. We have studied sperm from men with this hMSH2 mutation, since it is possible that mismatch repair mutations in humans might also have an effect on meiosis and normal segregation of chromosomes. The frequencies of aneuploid and diploid sperm were determined in 10 men with the hMSH2 mutation, by use of multicolor FISH analysis for chromosomes 13, 21, X, and Y. A minimum of 10,000 sperm per man was studied per chromosome probe. Control individuals consisted of men in the same kindred with HNPCC who did not carry the mutation and of other normal men from Newfoundland. A total of 321,663 sperm were analyzed: 200,905 sperm were from men carrying the hMSH2 mutation and 120,758 sperm were from control men. There was a significantly increased frequency of disomy 13, disomy 21, XX, and diploidy in mutation carriers compared with control men. These results suggest that the hMSH2 mutation may affect meiosis in humans. PMID- 10712227 TI - Exact multipoint quantitative-trait linkage analysis in pedigrees by variance components. AB - Methods based on variance components are powerful tools for linkage analysis of quantitative traits, because they allow simultaneous consideration of all pedigree members. The central idea is to identify loci making a significant contribution to the population variance of a trait, by use of allele-sharing probabilities derived from genotyped marker loci. The technique is only as powerful as the methods used to infer these probabilities, but, to date, no implementation has made full use of the inheritance information in mapping data. Here we present a new implementation that uses an exact multipoint algorithm to extract the full probability distribution of allele sharing at every point in a mapped region. At each locus in the region, the program fits a model that partitions total phenotypic variance into components due to environmental factors, a major gene at the locus, and other unlinked genes. Numerical methods are used to derive maximum-likelihood estimates of the variance components, under the assumption of multivariate normality. A likelihood-ratio test is then applied to detect any significant effect of the hypothesized major gene. Simulations show the method to have greater power than does traditional sib-pair analysis. The method is freely available in a new release of the software package GENEHUNTER. PMID- 10712228 TI - The transmission/disequilibrium test for linkage on the X chromosome. AB - The transmission/disequilibrium test (TDT), which detects linkage between a marker and disease loci in the presence of linkage disequilibrium, was introduced by Spielman et al. The original TDT requires families in which the genotypes are known for both parents and for at least one affected offspring, and this limits its applicability to diseases with late onset. The sib-TDT, or S-TDT, which utilizes families with affected and unaffected siblings, was introduced as an alternative method, by Spielman and Ewens, and the TDT and S-TDT can be combined in an overall test (i.e., a combined-TDT, or C-TDT). The TDT statistics described so far are for autosomal chromosomes. We have extended these TDT methods to test for linkage between X-linked markers and diseases that affect either males only or both sexes. For diseases of late onset, when parental genotypes are often unavailable, the X-linkage C-TDT may allow for more power than is provided by the X-linkage TDT alone. PMID- 10712229 TI - The transmission/disequilibrium test and parental-genotype reconstruction for X chromosomal markers. AB - Family-based association methods have recently been introduced that allow testing for linkage in the presence of linkage disequilibrium between a marker and a disease even if there is only incomplete parental-marker information. No such tests are currently available for X-linked markers. This report fills this methodological gap by presenting the X-linked sibling transmission/disequilibrium test (XS-TDT) and the X-linked reconstruction-combination transmission/disequilibrium test (XRC-TDT). As do their autosomal counterparts (S TDT and RC-TDT), these tests make no assumption about the mode of inheritance of the disease and the ascertainment of the sample. They protect against spurious association due to population stratification. The two tests were compared by simulations, which show that (1) the X-linked RC-TDT is, in general, considerably more powerful than the X-linked S-TDT and (2) the lack of parental-genotype information can be offset by the typing of a sufficient number of sibling controls. A freely available SAS implementation of these tests allows the calculation of exact P values. PMID- 10712231 TI - mtDNA haplogroups and frequency patterns in Europe. PMID- 10712230 TI - Gamete-competition models. AB - The gamete-competition model is an application of the Bradley-Terry model for ranking of sports teams. If allele i of a marker locus is assigned parameter taui>0, then the probability that a parent with heterozygous genotype i/j transmits allele i is Pr(i/j-->)=tau(i)/(tau(i) + tau(j). Mendelian segregation corresponds to the choice tau(i)=1 for all i. To test whether Mendelian segregation is true, one can estimate the tau(i) from pedigree data and perform a likelihood-ratio test under the constraint that one tau(i) equals 1. Although this procedure generates an interesting method for performance of segregation analysis with a marker locus, its real promise lies in generalization of the transmission/disequilibrium test. Quantitative as well as qualitative outcomes can be considered. The gamete-competition model uses full pedigree data and gives an estimate of the strength of transmission distortion to affected children for each allele. Covariates are incorporated by rewriting of tau(i)=exp(beta(t)x(k)), where beta is a parameter vector and xk is a covariate vector for the kth transmitted gamete. Examples of covariates include disease-severity indicators for the child, sex of the child, or repeat number for tandem-repeat alleles. PMID- 10712232 TI - Reconstruction of prehistory on the basis of genetic data. PMID- 10712233 TI - The I kappa B/NF-kappa B system: a key determinant of mucosalinflammation and protection. AB - The ubiquitous transcription factor NF-kappa B is a central regulator of the transcriptional activation of a number of genes involved in cell adhesion, immune and proinflammatory responses, apoptosis, differentiation, and growth. Induction of these genes in intestinal epithelial cells (IECs) by activated NF-kappa B profoundly influences mucosal inflammation and repair. NF-kappa B activation requires the removal of I kappa B from NF-kappa B by inducible proteolysis, which liberates this transcription factor for migration to the nucleus, where it binds to kappa B-regulatory elements and induces transcription. I kappa B alpha degradation is incomplete and delayed in IECs, resulting in buffered responses to luminal stimuli. The stimulatory environment partially determines whether the effect of NF-kappa B is protective or deleterious for the host. kappa B-dependent proinflammatory gene expression, particularly chemokines, major histocompatibility complex class II antigens, and adhesion molecules may be extremely important in early protective responses to mucosal pathogens but, when dysregulated, could lead to the development of chronic inflammation, as seen in inflammatory bowel diseases. The key role of NF-kappa B in regulating expression of a number of proinflammatory genes makes this protein an attractive target for selective therapeutic intervention. PMID- 10712234 TI - TNF-alpha potentiates the ion secretion induced by muscarinic receptor activation in HT29cl.19A cells. AB - Chronic gastrointestinal diseases such as ulcerative colitis and Crohn's disease are characterized by severe diarrhea. Mucosal biopsies of these patients show enhanced levels of cytokines, secreted by infiltrated inflammatory cells. In this study, we investigated the effect of the cytokine tumor necrosis factor-alpha (TNF-alpha) on ion secretion in human intestinal epithelial cells. The conventional microelectrode technique in the cell line HT29cl. 19A was used, which allows for simultaneous measurements of transepithelial potential difference and intracellular potential difference across the apical membrane. Preincubation (2-78 h) with 10 ng/ml TNF-alpha did not change basal secretory activity. However, the secretory response to the muscarinic receptor agonist carbachol was strongly increased after exposure to TNF-alpha. Application of the protein kinase C (PKC) inhibitor GF 109203X (bisindolylmaleimide I) inhibited the response to carbachol as well as the TNF-alpha-potentiated response, indicating that PKC mediates the effect of carbachol in this cell line. Propranolol, a substance that inhibits the phospholipase D (PLD) pathway, strongly reduced the response to muscarinic stimulation and its potentiation by TNF-alpha. The results indicate that activation of PLD is involved in ion secretion induced by muscarinic receptor activation and that TNF-alpha can potentiate this pathway. PMID- 10712235 TI - Stretch-induced activation of Ca(2+)-activated K(+) channels in mouse skeletal muscle fibers. AB - High-conductance Ca(2+)-activated K(+) (K(Ca)) channels were studied in mouse skeletal muscle fibers using the patch-clamp technique. In inside-out patches, application of negative pressure to the patch induced a dose-dependent and reversible activation of K(Ca) channels. Stretch-induced increase in channel activity was found to be of the same magnitude in the presence and in the absence of Ca(2+) in the pipette. The dose-response relationships between K(Ca) channel activity and intracellular Ca(2+) and between K(Ca) channel activity and membrane potential revealed that voltage and Ca(2+) sensitivity were not altered by membrane stretch. In cell-attached patches, in the presence of high external Ca(2+) concentration, stretch-induced activation was also observed. We conclude that membrane stretch is a potential mode of regulation of skeletal muscle K(Ca) channel activity and could be involved in the regulation of muscle excitability during contraction-relaxation cycles. PMID- 10712236 TI - Polyamine depletion delays apoptosis of rat intestinal epithelial cells. AB - The polyamines spermidine, spermine, and their precursor putrescine are essential for cell growth and the regulation of the cell cycle. Recent studies suggest that excessive accumulation of polyamines favors either malignant transformation or apoptosis, depending on the cell type and the stimulus. This study examines the involvement of polyamines in the induction of apoptosis by the DNA topoisomerase I inhibitor, camptothecin. In IEC-6 cells, camptothecin induced apoptosis within 6 h, accompanied by detachment of cells. Detached cells showed DNA laddering and caspase 3 induction, characteristic features of apoptosis. Depletion of putrescine, spermidine, and spermine by DL-alpha-difluoromethylornithine (DFMO), a specific inhibitor of ornithine decarboxylase (ODC) that is the first rate limiting enzyme for polyamine biosynthesis, decreased the apoptotic index. Delayed apoptosis was accompanied by a decrease in caspase 3 activity in polyamine-depleted cells. Addition of putrescine restored the induction of apoptosis as indicated by an increase in the number of detached cells and caspase 3 activity. Polyamine depletion did not change the level of caspase 3 protein. Inhibition of S-adenosylmethionine decarboxylase by a specific inhibitor [diethylglyoxal bis-(guanylhydrazone); DEGBG] led to depletion of spermidine and spermine with a significant accumulation of putrescine and induction of ODC. The DEGBG-treated cells showed an increase in apoptosis, suggesting the importance of putrescine in the apoptotic process. Addition of putrescine to DFMO-treated cell extracts did not increase caspase 3 activity. The above results indicate that polyamine depletion delays the onset of apoptosis in IEC-6 cells and confers protection against DNA damaging agents, suggesting that polyamines might be involved in the caspase activating signal cascade. PMID- 10712237 TI - Characterization of the synthesis and release of catecholamine in the rat carotid body in vitro. AB - The aim of this work was to determine contents and turnover rates for dopamine (DA) and norepinephrine (NE) and to identify the catecholamine (CA) released during stimulation of the rat carotid body (CB). Turnover rates and the release of CA were measured in an in vitro preparation using a combination of HPLC and radioisotopic methods. Mean rat CB levels of DA and NE were 209 and 45 pmol/mg tissue, respectively. With [(3)H]tyrosine as precursor, rat CB synthesized [(3)H]CA in a time- and concentration-dependent manner; calculated turnover times for DA and NE were 5.77 and 11.4 h, respectively. Hypoxia and dibutyryl adenosine 3',5'-cyclic monophosphate significantly increased [(3)H]CA synthesis. In normoxia, rat CB released [(3)H]DA and [(3)H]NE in a ratio of 5:1, comparable to that of the endogenous tissue CA. Hypoxia and high K(+) preferentially released [(3)H]DA, nicotine preferentially released [(3)H]NE, and acidic stimuli released both amines in proportion to tissue content. Release of [(3)H]CA induced by hypoxia and high K(+) was nearly fully dependent on extracellular Ca(2+), whereas basal normoxic release was not altered by removal of Ca(2+) from the incubating solution. We conclude that the rat CB is an organ with higher levels of DA than NE that preferentially releases DA or NE in a stimulus-specific manner. PMID- 10712238 TI - ERK signaling mediates the induction of inflammatory cytokines by bufalin in human monocytic cells. AB - Treatment of human leukemia THP-1 cells with bufalin, a specific inhibitor of Na(+)-K(+)-ATPase, sequentially induces c-fos and inflammatory cytokines interleukin-1 beta (IL-1 beta) and tumor necrosis factor-alpha (TNF-alpha) gene expressions before the appearance of mature phenotypes of monocytic cells. In this study we examined the signal transduction leading to bufalin-induced gene expressions. Bufalin selectively activated extracellular signal-regulated kinase (ERK), compared with other mitogen-activated protein (MAP) kinase family members. Pretreatment of THP-1 cells with PD-98059, an inhibitor of the ERK-kinase cascade, abolished bufalin-induced c-fos and IL-1 beta gene expressions, indicating that the ERK-kinase cascade mediates the induction of inflammatory cytokines by bufalin. Inhibition of the Na(+)/Ca(2+) exchanger by KB-R7943 and of protein kinase C (PKC) by Ro-31-8220 suppressed ERK activation and gene expressions of c-fos and IL-1 beta. These findings suggest that Na(+)-K(+)-ATPase inhibition by bufalin induces calcium influx and thereby activates PKC and ERK. In cells treated with an inhibitor of p38 MAP kinases, SB-203580, bufalin mediated ERK activation became persistent and the induction of IL-1 beta and TNF alpha expressions was significantly augmented. These results suggest that cross talk in bufalin-mediated ERK activation is negatively regulated by endogenous p38 MAP kinase activations. PMID- 10712239 TI - Reversal of sexual dimorphism in splenic T lymphocyte responses after trauma hemorrhage with aging. AB - Previous studies have demonstrated that hemorrhagic shock produces immunodepression in young male mice, whereas the immunoresponsiveness in young proestrus female mice is enhanced under such conditions. This sexually dimorphic immune response to hemorrhage appears to be related to high estrogen and testosterone levels in females and males, respectively. Nonetheless, it is unknown what impact the age-related decline in the sex steroid levels has on the immune response after hemorrhage. To study this, young (2-3 mo) and aged (18-19 mo) male and female CBA/J NIA mice were subjected to laparotomy (i.e., soft tissue trauma) and hemorrhage (35 +/- 5 mmHg for 90 min and fluid resuscitation) or sham operation. Twenty-four hours later, splenocyte responses were assessed in vitro. Splenic T lymphocyte responses [i.e., proliferation, interleukin-2 (IL-2) and interferon-gamma (IFN-gamma) release] were depressed in young males and enhanced in young females after trauma-hemorrhage. In contrast, in the aged male and female groups these parameters of splenocyte function were reversed after trauma-hemorrhage (i.e., increased proliferation and IL-2 release in aged males compared with suppressed proliferation and IFN-gamma release in aged females). Furthermore, the release of the immunosuppressive cytokine IL-10 inversely correlated with the age- and gender-related changes in splenocyte responses after trauma-hemorrhage. Thus the sexually dimorphic immune response in young males and females to trauma-hemorrhage appears to reverse as sex hormone levels decline with age. PMID- 10712240 TI - Effect of lactate on depolarization-induced Ca(2+) release in mechanically skinned skeletal muscle fibers. AB - It is unclear whether accumulation of lactate in skeletal muscle fibers during intense activity contributes to muscle fatigue. Using mechanically skinned fibers from rat and toad muscle, we were able to examine the effect of L(+)-lactate on excitation-contraction coupling independently of other metabolic changes. We investigated the effects of lactate on the contractile apparatus, caffeine induced Ca(2+) release from the sarcoplasmic reticulum, and depolarization induced Ca(2+) release. Lactate (15 or 30 mM) had only a small inhibitory effect directly on the contractile apparatus and caused appreciable (20-35%) inhibition of caffeine-induced Ca(2+) release, seemingly by a direct effect on the Ca(2+) release channels. However, 15 mM lactate had no detectable effect on Ca(2+) release when it was triggered by the normal voltage sensor mechanism, and 30 mM lactate reduced such release by only <10%. These results indicate that lactate has only a relatively small inhibitory effect on normal excitation-contraction coupling, indicating that lactate accumulation per se is not a major factor in muscle fatigue. PMID- 10712241 TI - Functional significance of human trp1 and trp3 in store-operated Ca(2+) entry in HEK-293 cells. AB - The Drosophila trp (transient receptor potential) gene appears to encode the Drosophila store-operated channel (SOC), and some mammalian trp homologues have been proposed to encode mammalian SOCs. This study provides evidence for the expression of three trp homologues (Mtrp2, Mtrp3, and Mtrp4) in fibroblasts from wild-type and src knockout mice, and four trp homologues (Htrp1, Htrp3, Htrp4, and Htrp6) in human embryonic kidney (HEK-293) cells based on RT-PCR techniques. In HEK-293 cells stably transfected with a 323-bp Htrp3 antisense construct (Htrp3AS), Northern blot analysis revealed that the expression of a 4-kb transcript was dramatically suppressed in comparison to that observed in cells stably transfected with a short Htrp3 sense construct (Htrp3S). Activity of SOCs, monitored as Ba(2+) influx following Ca(2+) store depletion with thapsigargin, was reduced by 32% in Htrp3AS cells in comparison with Htrp3S cells. Transient transfection of a 369-bp Htrp1 antisense construct in cells stably expressing Htrp3AS induced a higher level of inhibition (55%) of store-operated Ca(2+) entry. These data suggest that Htrp1 and Htrp3 may be functional subunits of SOCs. PMID- 10712242 TI - Inhibition of CaM kinase II activation and force maintenance by KN-93 in arterial smooth muscle. AB - Ca(+)/calmodulin-dependent protein kinase II (CaM kinase II) has been implicated in the regulation of smooth muscle contractility. The goals of this study were to determine: 1) to what extent CaM kinase II is activated by contractile stimuli in intact arterial smooth muscle, and 2) the effect of a CaM kinase II inhibitor (KN 93) on CaM kinase II activation, phosphorylation of myosin regulatory light chains (MLC(20)), and force. Both histamine (1 microM) and KCl depolarization activated CaM kinase II with a time course preceding maximal force development, and suprabasal CaM kinase II activation was sustained during tonic contractions. CaM kinase II activation was inhibited by KN-93 pretreatment (IC(50) approximately 1 microM). KN-93 inhibited histamine-induced tonic force maintenance, whereas early force development and MLC(20) phosphorylation responses during the entire time course were unaffected. Both force development and maintenance in response to KCl were inhibited by KN-93. Rapid increases in KCl-induced MLC(20) phosphorylation were also inhibited by KN-93, whereas steady state MLC(20) phosphorylation responses were unaffected. In contrast, phorbol 12,13-dibutyrate (PDBu) did not activate CaM kinase II and PDBu-stimulated force development was unaffected by KN-93. Thus KN-93 appears to target a step(s) essential for force maintenance in response to physiological stimuli, suggesting a role for CaM kinase II in regulating tonic contractile responses in arterial smooth muscle. Pharmacological activation of protein kinase C bypasses the KN-93 sensitive step. PMID- 10712243 TI - Voltage-dependent stimulation of the Na(+)-K(+) pump by insulin in rabbit cardiac myocytes. AB - Insulin enhances Na(+)-K(+) pump activity in various noncardiac tissues. We examined whether insulin exposure in vitro regulates Na(+)-K(+) pump function in rabbit ventricular myocytes. Pump current (I(p)) was measured using the whole cell patch-clamp technique at test potentials (V(m)s) from -100 to +60 mV. When the Na(+) concentration in the patch pipette ([Na](pip)) was 10 mM, insulin caused a V(m)-dependent increase in I(p). The increase was approximately 70% when V(m) was at near physiological diastolic potentials. This effect persisted after elimination of extracellular voltage-dependent steps and when K(+) and K(+) congeners were excluded from the patch pipettes. When [Na](pip) was 80 mM, causing near-maximal pump stimulation, insulin had no effect, suggesting that it did not cause an increase in membrane pump density. Effects of tyrphostin A25, wortmannin, okadaic acid, or bisindolylmaleimide I in pipette solutions suggested that the insulin-induced increase in I(p) involved activation of tyrosine kinase, phosphatidylinositol 3-kinase, and protein phosphatase 1, whereas protein phosphatase 2A and protein kinase C were not involved. PMID- 10712244 TI - Phosphatidylcholine participates in the interaction between macrophages and lymphocytes. AB - The role of phosphatidylcholine molecules as mediator for the control of lymphocyte proliferation by macrophages was investigated. Phosphatidylcholine added to the culture medium inhibited the concanavalin A-stimulated lymphocyte proliferation in a concentration-dependent manner. The potency of this effect was dependent on the presence of arachidonic acid in the phosphatidylcholine molecules. The phosphatidylcholine transfer from macrophages to lymphocytes was then investigated. Macrophages incorporated phosphatidylcholine at a much higher rate than lymphocytes and exported phosphatidylcholine to the culture medium. When cocultured, a significant amount of phosphatidylcholine incorporated by macrophages was transferred to lymphocytes. To examine the possible physiological importance of the transfer process, the lymphocyte proliferation was measured in coculture conditions. Macrophages were treated with phosphatidylcholine and washed, and then these cells were cocultured with concanavalin A-stimulated lymphocytes. The effect observed in coculture was an inhibition of lymphocyte proliferation, which was also dependent on the molecular species of the phosphatidylcholine. Therefore, phosphatidylcholine may act as a mediator of the macrophage effect on lymphocyte proliferation. PMID- 10712245 TI - Sensitivity of myometrium to CGRP varies during mouse estrous cycle and in response to progesterone. AB - Calcitonin gene-related peptide (CGRP) inhibits contractions of the myometrium. Isometric force measurements on myometrial strips were carried out to monitor the inhibitory capacity of CGRP in the myometrium during the estrous cycle and in response to estrogen and progesterone in ovariectomized mice. CGRP inhibition of KCl-induced contractions was lowest at estrus and significantly increased during metestrus and diestrus. Progesterone treatment of ovariectomized mice resulted in a significant increase in the responsiveness of the myometrium to CGRP. Expression of CGRP-receptor component protein (CGRP-RCP), a marker of CGRP receptor expression, was quantitated by Western and Northern blot analyses. The levels of inhibition exerted by CGRP during the various stages of the estrous cycle and in response to steroid hormone treatment correlated with the protein levels of CGRP-RCP. The mRNA levels did not change significantly during the estrous cycle or in response to hormone treatment, indicating that the regulation of CGRP-RCP protein does not occur at the transcriptional level. CGRP had an inhibitory effect both when applied before the stimulus for contraction and when applied during a sustained contracture induced by KCl. This suggests that CGRP induced generation of second messengers can influence late events in electro /chemomechanical coupling and/or the contractile machinery directly. PMID- 10712246 TI - Pharmacological activation of cloned intermediate- and small-conductance Ca(2+) activated K(+) channels. AB - We previously characterized 1-ethyl-2-benzimidazolinone (1-EBIO), as well as the clinically useful benzoxazoles, chlorzoxazone (CZ), and zoxazolamine (ZOX), as pharmacological activators of the intermediate-conductance Ca(2+)-activated K(+) channel, hIK1. The mechanism of activation of hIK1, as well as the highly homologous small-conductance, Ca(2+)-dependent K(+) channel, rSK2, was determined following heterologous expression in Xenopus oocytes using two-electrode voltage clamp (TEVC) and excised, inside-out patch-clamp techniques. 1-EBIO, CZ, and ZOX activated both hIK1 and rSK2 in TEVC and excised inside-out patch-clamp experiments. In excised, inside-out patches, 1-EBIO and CZ induced a concentration-dependent activation of hIK1, with half-maximal (K(1/2)) values of 84 microM and 98 microM, respectively. Similarly, CZ activated rSK2 with a K(1/2) of 87 microM. In the absence of CZ, the Ca(2+)-dependent activation of hIK1 was best fit with a K(1/2) of 700 nM and a Hill coefficient (n) of 2.0. rSK2 was activated by Ca(2+) with a K(1/2) of 700 nM and an n of 2.5. Addition of CZ had no effect on either the K(1/2) or n for Ca(2+)-dependent activation of either hIK1 or rSK2. Rather, CZ increased channel activity at all Ca(2+) concentrations (V(max)). Event-duration analysis revealed hIK1 was minimally described by two open and three closed times. Activation by 1-EBIO had no effect on tau(o1), tau(o2), or tau(c1), whereas tau(c2) and tau(c3) were reduced from 9.0 and 92.6 ms to 5.0 and 44.1 ms, respectively. In conclusion, we define 1-EBIO, CZ, and ZOX as the first known activators of hIK1 and rSK2. Openers of IK and SK channels may be therapeutically beneficial in cystic fibrosis and vascular diseases. PMID- 10712247 TI - Effects of beta(2)-agonist clenbuterol on biochemical and contractile properties of unloaded soleus fibers of rat. AB - The effects of clenbuterol beta(2)-agonist administration were investigated in normal and atrophied [15-day hindlimb-unloaded (HU)] rat soleus muscles. We showed that clenbuterol had a specific effect on muscle tissue, since it reduces soleus atrophy induced by HU. The study of Ca(2+) activation properties of single skinned fibers revealed that clenbuterol partly prevented the decrease in maximal tension after HU, with a preferential effect on fast-twitch fibers. Clenbuterol improved the Ca(2+) sensitivity in slow- and fast-twitch fibers by shifting the tension-pCa relationship toward lower Ca(2+) concentrations, but this effect was more marked after HU than in normal conditions. Whole muscle electrophoresis indicated slow-to-fast transitions of the myosin heavy chain isoforms for unloaded and for clenbuterol-treated soleus. The coupling of the two latter conditions did not, however, increase these phenotypical transformations. Our findings indicated that clenbuterol had an anabolic action and a beta(2) adrenergic effect on muscle fibers and appeared to counteract some effects of unloading disuse conditions. PMID- 10712248 TI - A myosin phosphatase targeting subunit isoform transition defines a smooth muscle developmental phenotypic switch. AB - Smooth muscle myosin phosphatase dephosphorylates the regulatory myosin light chain and thus mediates smooth muscle relaxation. The activity of this myosin phosphatase is dependent upon its myosin-targeting subunit (MYPT1). Isoforms of MYPT1 have been identified, but how they are generated and their relationship to smooth muscle phenotypes is not clear. Cloning of the middle section of chicken and rat MYPT1 genes revealed that each gene gave rise to isoforms by cassette type alternative splicing of exons. In chicken, a 123-nucleotide exon was included or excluded from the mature mRNA, whereas in rat two exons immediately downstream were alternative. MYPT1 isoforms lacking the alternative exon were only detected in mature chicken smooth muscle tissues that display phasic contractile properties, but the isoform ratios were variable. The patterns of expression of rat MYPT1 mRNA isoforms were more complex, with three major and two minor isoforms present in all smooth muscle tissues at varying stoichiometries. Isoform switching was identified in the developing chicken gizzard, in which the exon-skipped isoform replaced the exon-included isoform around the time of hatching. This isoform switch occurred after transitions in myosin heavy chain and myosin light chain (MLC(17)) isoforms and correlated with a severalfold increase in the rate of relaxation. The developmental switch of MYPT1 isoforms is a good model for determining the mechanisms and significance of alternative splicing in smooth muscle. PMID- 10712249 TI - Mechanisms of P(i) regulation of the skeletal muscle SR Ca(2+) release channel. AB - Inorganic phosphate (P(i)) accumulates in the fibers of actively working muscle where it acts at various sites to modulate contraction. To characterize the role of P(i) as a regulator of the sarcoplasmic reticulum (SR) calcium (Ca(2+)) release channel, we examined the action of P(i) on purified SR Ca(2+) release channels, isolated SR vesicles, and skinned skeletal muscle fibers. In single channel studies, addition of P(i) to the cis chamber increased single channel open probability (P(o); 0.079 +/- 0.020 in 0 P(i), 0. 157 +/- 0.034 in 20 mM P(i)) by decreasing mean channel closed time; mean channel open times were unaffected. In contrast, the ATP analog, beta,gamma-methyleneadenosine 5' triphosphate (AMP-PCP), enhanced P(o) by increasing single channel open time and decreasing channel closed time. P(i) stimulation of [(3)H]ryanodine binding by SR vesicles was similar at all concentrations of AMP-PCP, suggesting P(i) and adenine nucleotides act via independent sites. In skinned muscle fibers, 40 mM P(i) enhanced Ca(2+)-induced Ca(2+) release, suggesting an in situ stimulation of the release channel by high concentrations of P(i). Our results support the hypothesis that P(i) may be an important endogenous modulator of the skeletal muscle SR Ca(2+) release channel under fatiguing conditions in vivo, acting via a mechanism distinct from adenine nucleotides. PMID- 10712251 TI - HSV-1 amplicon vectors are a highly efficient gene delivery system for skeletal muscle myoblasts and myotubes. AB - Analysis of RyR1 structure function in muscle cells is made difficult by the low (<5%) transfection efficiencies of myoblasts or myotubes using calcium phosphate or cationic lipid techniques. We inserted the full-length 15.3-kb RyR1 cDNA into a herpes simplex virus type 1 (HSV-1) amplicon vector, pHSVPrPUC between the ori/IE 4/5 promoter sequence and the HSV-1 DNA cleavage/packaging signal (pac). pHSVGN and pHSVGRyR1, two amplicons that expressed green fluorescent protein, were used for fluorescence-activated cell sorter analysis of transduction efficiency. All amplicons were packaged into HSV-1 virus particles using a helper virus-free packaging system and yielded 10(6) transducing vector units/ml. HSVRyR1, HSVGRyR1, and HSVGN virions efficiently transduced mouse myoblasts and myotubes, expressing the desired product in 70-90% of the cells at multiplicity of infection 5. The transduced cells appeared healthy and RyR1 produced by this method was targeted properly and restored skeletal excitation-contraction coupling in dyspedic myotubes. The myotubes produced sufficient protein to allow single-channel analyses from as few as 10 100-mm dishes. In most cases this method could preclude the need for permanent transfectants for the study of RyR1 structure function. PMID- 10712250 TI - Lysophosphatidic acid and sphingosine 1-phosphate stimulate endothelial cell wound healing. AB - Lysophosphatidic acid (LPA) and sphingosine 1-phosphate (S1P) are potent lipid growth factors with similar abilities to stimulate cytoskeleton-based cellular functions. Their effects are mediated by a subfamily of G protein-coupled receptors (GPCRs) encoded by endothelial differentiation genes (edgs). We hypothesize that large quantities of LPA and S1P generated by activated platelets may influence endothelial cell functions. Using an in vitro wound healing assay, we observed that LPA and S1P stimulated closure of wounded monolayers of human umbilical vein endothelial cells and adult bovine aortic endothelial cells, which express LPA receptor Edg2, and S1P receptors Edg1 and Edg3. The two major components of wound healing, cell migration and proliferation, were stimulated individually by both lipids. LPA and S1P also stimulated intracellular Ca(2+) mobilization and mitogen-activated protein kinase (MAPK) phosphorylation. Pertussis toxin partially blocked the effects of both lipids on endothelial cell migration, MAPK phosphorylation, and Ca(2+) mobilization, implicating G(i)/(o) coupled Edg receptor signaling in endothelial cells. LPA and S1P did not cross desensitize each other in Ca(2+) responses, suggesting involvement of distinct receptors. Thus LPA and S1P affect endothelial cell functions through signaling pathways activated by distinct GPCRs and may contribute to the healing of wounded vasculatures. PMID- 10712252 TI - IX. Mast cell-deficient mice and intestinal biology. AB - Mutant mice that express abnormalities in mast cell development represent a powerful tool for the investigation of multiple aspects of mast cell biology. In addition, the identification of the genes affected by these mutations has not only increased our knowledge about the mast cell but has opened new areas of investigation as to the role of these gene products in gastrointestinal pathology, immunology, and physiology. PMID- 10712253 TI - I. Cellular and molecular biology of sodium channel beta-subunits: therapeutic implications for pain? AB - Voltage-gated sodium channel alpha-subunits have been shown to be key mediators of the pathophysiology of pain. The present review considers the role of sodium channel auxiliary beta-subunits in channel modulation, channel protein expression levels, and interactions with extracellular matrix and cytoskeletal signaling molecules. Although beta-subunits have not yet been directly implicated in pain mechanisms, their intimate association with and ability to regulate alpha subunits predicts that they may be a viable target for therapeutic intervention in the future. It is proposed that multifunctional sodium channel beta-subunits provide a critical link between extracellular and intracellular signaling molecules and thus have the ability to fine tune channel activity and electrical excitability. PMID- 10712254 TI - Fas and Fas ligand in gut and liver. AB - Apoptosis (programmed cell death) has been shown to play a major role in development and in the pathogenesis of numerous diseases. A principal mechanism of apoptosis is molecular interaction between surface molecules known as the "death receptors" and their ligands. Perhaps the best-studied death receptor and ligand system is the Fas/Fas ligand (FasL) system, in which FasL, a member of the tumor necrosis factor (TNF) family of death-inducing ligands, signals death through the death receptor Fas, thereby resulting in the apoptotic death of the cell. Numerous cells in the liver and gastrointestinal tract have been shown to express Fas/FasL, and there is a growing body of evidence that the Fas/FasL system plays a major role in the pathogenesis of many liver and gastrointestinal diseases, such as inflammatory bowel disease, graft vs. host disease, and hepatitis. Here we review the Fas/FasL system and the evidence that it is involved in the pathogenesis of liver and gastrointestinal diseases. PMID- 10712255 TI - Role of endothelin-1 in regulation of the postnatal intestinal circulation. AB - Newborn intestine is uniquely prone to vasoconstriction in response to a wide variety of perturbations. To test the hypothesis that endothelin (ET)-1 is an important factor in this process, we determined the effects of exogenous ET-1 administration and blockade of endogenous ET-1 in vivo and in vitro in 3- and 35 day-old swine. Intramesenteric artery administration of exogenous ET-1 to vascularly isolated in vivo gut loops (10(-9) M/kg bolus) caused vasoconstriction and reduced gut O(2) uptake similarly in these age groups. Selective blockade of ET(A) or ET(B) receptors with BQ-610 or BQ-788, respectively, in vascularly isolated in vivo gut loops had no effect on gut vascular resistance or O(2) uptake in either age group; within in vitro gut loops, BQ-610 significantly increased vasoconstriction when perfusion pressure was reduced below baseline, but only in 3-day-old animals; i.e., it impaired the autoregulatory response to perfusion pressure reduction. Exogenous ET-1 significantly decreased capillary perfusion within in vitro gut loops, as evidenced by a decrease in capillary filtration coefficient, but only in 3-day-old animals; furthermore, blockade of endogenous ET-1 activity with BQ-610 significantly enhanced capillary filtration coefficient in 3-day-old animals and increased O(2) extraction ratio. ET-1 did not depress intestinal metabolic rate, as evidenced by its effect on the O(2) uptake-blood flow relationship; it did compromise tissue oxygenation because of its effects on intestinal O(2) transport. ET-1 concentration in mesenteric venous effluent exceeded arterial concentration, but only in 3-day-old intestine, suggesting production of ET-1 by newborn intestine. We conclude that ET-1 exerts an age-dependent effect on intestinal hemodynamics in postnatal intestine, having a greater impact in 3- than in 35-day-old intestine. PMID- 10712256 TI - Complete lower esophageal sphincter relaxation observed in some achalasia patients is functionally inadequate. AB - Generally accepted manometric criteria for the diagnosis of achalasia are absent peristalsis and incomplete lower esophageal sphincter (LES) relaxation. However, in some patients with otherwise typical features of achalasia, esophageal manometry shows complete LES relaxation during swallowing. To establish whether such apparently complete LES relaxations are functionally adequate, we quantified changes in resistance to flow at the esophagogastric junction (EGJ) during wet swallowing. We studied seven achalasia patients with manometrically complete (>80%) LES relaxation, eight achalasia patients with incomplete (<40%) LES relaxation, and eight healthy volunteers. Complete LES relaxation on standard manometry (open-tip catheters) was confirmed in five of the seven achalasia patients by a Dentsleeve. Changes in EGJ resistance to flow were quantified using a pneumatic resistometer. Manometrically, the relaxation time span was significantly longer in patients with complete LES relaxation than in those with incomplete relaxation (7. 3 +/- 0.5 vs. 4.4 +/- 0.7 s; P < 0.05). The fall in EGJ resistance from basal values during swallowing was markedly reduced in both achalasia groups (21 +/- 8% in those with manometrically complete relaxation and 4 +/- 2% in those with incomplete relaxation) by comparison with healthy individuals, in whom resistance fell by 90 +/- 3% (P < 0.05 vs. both achalasia groups). The duration of EGJ resistance drop was also much shorter in achalasia with (0.7 +/- 0.2 s) and without (0.2 +/- 0.1 s) complete LES relaxation compared with healthy control values (6.6 +/- 1.2 s). Our results reveal that the apparently complete LES relaxation observed manometrically in some patients with achalasia is functionally inadequate since it is not associated with the normal profound fall in EGJ resistance to flow. PMID- 10712257 TI - Increased insulin-like growth factor binding protein-4 expression after partial hepatectomy in the rat. AB - The insulin-like growth factor (IGF) binding proteins (IGFBPs) are important regulators of cell growth produced by different tissues. The IGFBPs regulate cell growth by modulating the activity and bioavailability of IGFs. The evidence that IGFBP-1 is a liver-specific immediate-early gene highly induced after 70% partial hepatectomy (PHx) suggests a role for the IGF-IGFBP system in hepatic regeneration. In this work we analyzed the effect of PHx on the expression of IGFBP-4, which is highly produced by the liver and very abundant in rat serum. Our results show a marked increase in hepatic IGFBP-4 mRNA levels 6-12 h after PHx and no significant change in sham-operated control animals. A parallel rise in IGFBP-4 transcript abundance was observed in the kidneys of PHx rats but not in sham-operated animals. Moreover, ligand blot analysis demonstrated that serum IGFBP-4 levels began to increase 12-24 h after surgery, consistent with the rise in the corresponding mRNA. This enhancement in IGFBP-4 production after PHx could be part of a fine regulatory mechanism to modulate IGF activity during liver regeneration. PMID- 10712259 TI - Mechanisms of Mg(2+) transport in cultured ruminal epithelial cells. AB - Net Mg(2+) absorption from the rumen is mainly mediated by a transcellular pathway, with the greater part (62%) being electrically silent. To investigate this component of Mg(2+) transport, experiments were performed with isolated ruminal epithelial cells (REC). Using the fluorescent indicators mag-fura 2, sodium-binding benzofuran isophthalate, and 2', 7'-bis(2-carboxyethyl)-5(6) carboxyfluorescein, we measured the intracellular free Mg(2+) concentration ([Mg(2+)](i)), the intracellular Na(+) concentration ([Na(+)](i)), and the intracellular pH (pH(i)) of REC under basal conditions, after stimulation with butyrate and HCO(-)(3), and after changing the transmembrane chemical gradients for Mg(2+), H(+), and Na(+). REC had a mean resting pH(i) of 6.83 +/- 0.1, [Mg(2+)](i) was 0.56 +/- 0. 14 mM, and [Na(+)](i) was 18.95 +/- 3.9 mM. Exposure to both HCO(-)(3) and HCO(-)(3)/butyrate led to a stimulation of Mg(2+) influx that amounted to 27.7 +/- 5 and 29 +/- 10.6 microM/min, respectively, compared with 15 +/- 1 microM/min in control solution. The increase of [Mg(2+)](i) was dependent on extracellular Mg(2+) concentration ([Mg(2+)](e)). Regulation of pH(i) has been demonstrated to be Na(+) dependent and is performed, for the most part, by a Na(+)/H(+) exchanger. The recovery of pH(i) was fully blocked in nominally Na(+)-free media, even if [Mg(2+)](e) was stepwise increased from 0 to 7.5 mM. However, an increase of [Mg(2+)](i) was observed after reversing the transmembrane Na(+) gradient. This rise in [Mg(2+)](i) was pH independent, K(+) insensitive, dependent on [Mg(2+)](e), imipramine and quinidine sensitive, and accompanied by a decrease of [Na(+)](i). The results are consistent with the existence of a Na(+)/Mg(2+) exchanger in the cell membrane of REC. The coupling between butyrate, CO(2)/HCO(-)(3), and Mg(2+) transport may be mediated by another mechanism, perhaps by cotransport of Mg(2+) and HCO(-)(3). PMID- 10712258 TI - Mice overexpressing progastrin are predisposed for developing aberrant colonic crypt foci in response to AOM. AB - Recent studies show that nonamidated gastrins (Gly-gastrin and progastrin) stimulate colonic proliferation. However, the role of nonamidated vs. amidated gastrins in colon carcinogenesis has not been defined. We measured intermediate markers of carcinogenesis in transgenic mice overexpressing either progastrin (hGAS) or amidated gastrin (INS-GAS) in response to azoxymethane (AOM). The hGAS mice showed significantly higher numbers of aberrant crypt foci (140-200% increase) compared with that in wild-type (WT) and INS-GAS mice (P < 0.05) after AOM treatment. The bromodeoxyuridine-labeling index of colonic crypts also was significantly elevated in hGAS mice vs. that in WT and INS-GAS mice. The results therefore provide evidence for a mitogenic and cocarcinogenic role of nonamidated gastrins (progastrin), which is apparently not shared by the amidated gastrins. Although nonamidated gastrins are now believed to mediate mitogenic effects via novel receptors, amidated gastrins mediate biological effects via different receptor subtypes, which may explain the difference in the cocarcinogenic potential of nonamidated vs. amidated gastrins. In conclusion, our results provide strong support for a cocarcinogenic role for nonamidated gastrins in colon carcinogenesis. PMID- 10712260 TI - Acid-base effects on electrolyte transport in CA II-deficient mouse colon. AB - To determine the role of carbonic anhydrase (CA) in colonic electrolyte transport, we studied Car-2(0) mice, mutants deficient in cytosolic CA II. Ion fluxes were measured under short-circuit conditions in an Ussing chamber. CA was analyzed by assay and Western blots. In Car-2(0) mouse colonic mucosa, total CA activity was reduced 80% and cytosolic CA I and membrane-bound CA IV activities were not increased. Western blots confirmed the absence of CA II in Car-2(0) mice. Normal mouse distal colon exhibited net Na(+) and Cl(-) absorption, a serosa-positive PD, and was specifically sensitive to pH. Decrease in pH stimulated active Na(+) and Cl(-) absorption whether it was caused by increasing solution PCO(2), reducing HCO(-)(3) concentration, or reducing pH in CO(2)/HCO( )(3)-free HEPES-Ringer solution. Membrane-permeant methazolamide, but not impermeant benzolamide, at 0.1 mM prevented the effects of pH. Car-2(0) mice exhibited similar basal transport rates and responses to pH and CA inhibitors. We conclude that basal and pH-stimulated colonic electrolyte absorption in mice requires CA I. CA II and IV may have accessory roles. PMID- 10712261 TI - Minor involvement of nitric oxide during chronic endotoxemia in anesthetized pigs. AB - To study the modifications of hepatic blood flow and hepatic function over time during endotoxemia, 10 pigs received a continuous intravenous infusion of endotoxin (Endo, 160 ng. kg(-1). h(-1)) over 18 h and 7 control (Ctrl) animals received a saline infusion. The involvement of nitric oxide (NO) in this endotoxic model was assessed by measuring plasma concentrations of NO(-)(2), NO( )(3), and cGMP, by testing vascular reactivity to ACh, and by evaluating inducible NO synthase (NOS 2) expression in hepatic biopsies. Endotoxin induced hypotensive and normokinetic shock in association with few modifications of hepatic blood flow, and hepatic injury was observed in both groups. Endotoxin did not increase plasma concentrations of NO(-)(2), NO(-)(3), and cGMP. The ACh dependent decrease of mean arterial pressure was reduced in Endo pigs, whereas a minor difference was observed between Ctrl and Endo pigs for ACh-dependent modification of hepatic perfusion. Hepatic NOS 2 mRNA was not detected in Ctrl pigs. In Endo pigs, NOS 2 protein expression was detected only in tissues surrounding the portal vein and the inferior vena cava, whereas NOS 2 mRNA was expressed in all hepatic biopsies. Thus, although endotoxemia induces NOS 2 expression in the liver, our findings show that NO involvement is lower in pigs than in rodents during endotoxemia. PMID- 10712262 TI - Glucagon-like peptide-2 increases sucrase-isomaltase but not caudal-related homeobox protein-2 gene expression. AB - To determine the effect of glucagon-like peptide-2 (GLP-2) on sucrase-isomaltase and caudal-related homeobox protein-2 (Cdx-2) gene expression, male Wistar rats were divided into total parenteral nutrition (TPN)-fed and GLP-2-treated, TPN-fed groups. TPN was given via a jugular line, inserted under anesthesia, for 7 days. The treatment group received 40 microg/day of GLP-2 intravenously with the TPN diet. The small intestine and colon were weighed and measured. Tissue was obtained from the jejunum, terminal ileum, and midcolon. RNA analysis, morphometry, and microdissection were performed. The weight of the small intestine of GLP-2-treated rats was greater than that of TPN-fed rats (P < 0.001). GLP-2 increased the mean metaphase arrests/crypt in both the jejunum and ileum (P < 0.001). Ileal expression of sucrase-isomaltase was increased by 1. 6 fold (P < 0.05). Jejunal expression was increased by a similar amount, although not significantly (P = 0.08). There was no change in Cdx-2 gene expression. Thus GLP-2 can maintain small intestinal morphology and function, but effects on gene expression are not mediated by gross changes in the level of the mRNA for the homeobox protein Cdx-2. PMID- 10712263 TI - Coupling of M(2) muscarinic receptors to ERK MAP kinases and caldesmon phosphorylation in colonic smooth muscle. AB - Coupling of M(2) and M(3) muscarinic receptors to activation of mitogen-activated protein (MAP) kinases and phosphorylation of caldesmon was studied in canine colonic smooth muscle strips in which M(3) receptors were selectively inactivated by N, N-dimethyl-4-piperidinyl diphenylacetate (4-DAMP) mustard (40 nM). ACh elicited activation of extracellular signal-regulated kinase (ERK) 1, ERK2, and p38 MAP kinases in control muscles and increased phosphorylation of caldesmon (Ser(789)), a putative downstream target of MAP kinases. Alkylation of M(3) receptors with 4-DAMP had only a modest inhibitory effect on ERK activation, p38 MAP kinase activation, and caldesmon phosphorylation. Subsequent treatment with 1 microM AF-DX 116 completely prevented activation of ERK and p38 MAP kinase and prevented caldesmon phosphorylation. Caldesmon phosphorylation was blocked by the MAP kinase/ERK kinase inhibitor PD-98509 but not by the p38 MAP kinase inhibitor SB-203580. These results indicate that colonic smooth muscle M(2) receptors are coupled to ERK and p38 MAP kinases. Activation of ERK, but not p38 MAP kinases, results in phosphorylation of caldesmon in vivo, which is a novel function for M(2) receptor activation in smooth muscle. PMID- 10712264 TI - Characterization of inducible nature of MRP3 in rat liver. AB - We found previously that expression of multidrug resistance-associated protein (MRP) 3 is induced in a mutant rat strain (Eisai hyperbilirubinemic rats) whose canalicular multispecific organic anion transporter (cMOAT/MRP2) function is hereditarily defective and in normal Sprague-Dawley (SD) rats after ligation of the common bile duct. In the present study, the inducible nature of MRP3 was examined, using Northern and Western blot analyses, in comparison with that of other secondary active [Na(+)-taurocholic acid cotransporting polypeptide (Ntcp), organic anion transporting polypeptide 1 (oatp1), and organic cation transporter (OCT1)] and primary active [P-glycoprotein (P-gp), cMOAT/MRP2, and MRP6] transporters. alpha-Naphthylisothiocyanate treatment and common bile duct ligation induced expression of P-gp and MRP3, whereas expression of Ntcp, oatp1, and OCT1 was reduced by the same treatment. Although expression of MRP3 was also induced by administration of phenobarbital, that of cMOAT/MRP2, MRP1, and MRP6 was not affected by any of these treatments. Moreover, the mRNA level of MRP3, but not that of P-gp, was increased in SD rats after administration of bilirubin and in Gunn rats whose hepatic bilirubin concentration is elevated because of a defect in the expression of UDP-glucuronosyl transferase. However, the MRP3 protein level was not affected by bilirubin administration. Although the increased MRP3 mRNA level was associated with the increased concentration of bilirubin and/or its glucuronides in mutant rats and in SD rats that had undergone common bile duct ligation or alpha-naphthylisothiocyanate treatment, we must assume that factor(s) other than these physiological substances are also involved in the increased protein level of MRP3. PMID- 10712265 TI - Cell-specific localization of insulin-like growth factor binding protein mRNAs in rat liver. AB - The liver is a major source of circulating insulin-like growth factor I (IGF-I), and it also synthesizes several classes of IGF binding proteins (IGFBPs). Synthesis of IGF-I and IGFBPs is regulated by hormones, growth factors, and cytokines. They are nutritionally regulated and expressed in developmentally specific patterns. To gain insight into cellular regulatory mechanisms that determine hepatic synthesis of IGF-I and IGFBPs and to identify potential target cells for IGF-I within the liver, we studied the cellular sites of synthesis of IGF-I, IGF receptor, growth hormone (GH) receptor, and IGFBPs in freshly isolated rat hepatocytes, endothelial cells, and Kupffer cells. We also localized cellular sites of IGFBP synthesis by in situ hybridization histochemistry. Western ligand and immunoblot analyses were used to determine IGFBP secretion by isolated cells. Two IGF-I mRNA subtypes with different 5' ends (class 1 and class 2) were detected in all isolated liver cell preparations. Type 1 IGF receptor mRNA was detected in endothelial cells, indicating that these cells are a local target for IGF actions in liver. GH receptor was expressed in all cell preparations, consistent with GH regulation of IGF-I and IGFBP synthesis in multiple liver cell types. The IGFBPs expressed striking cell-specific expression. IGFBP-1 was synthesized only in hepatocytes, and IGFBP-3 was expressed in Kupffer and endothelial cells. IGFBP-4 was expressed at high levels in hepatocytes and at low levels in Kupffer and endothelial cells. Cell-specific expression of distinct IGFBPs in the liver provides the potential for cell-specific regulation of hepatic and endocrine actions of IGF-I. PMID- 10712266 TI - Antroduodenal motility in chronic pancreatitis: are abnormalities related to exocrine insufficiency? AB - In patients with chronic pancreatitis (CP) the relation among exocrine pancreatic secretion, gastrointestinal hormone release, and motility is disturbed. We studied digestive and interdigestive antroduodenal motility and postprandial gut hormone release in 26 patients with CP. Fifteen of these patients had pancreatic insufficiency (PI) established by urinary para-aminobenzoic acid test and fecal fat excretion. Antroduodenal motility was recorded after ingestion of a mixed liquid meal. The effect of pancreatic enzyme supplementation was studied in 8 of the 15 CP patients with PI. The duration of the postprandial antroduodenal motor pattern was significantly (P < 0.01) prolonged in CP patients (324 +/- 20 min) compared with controls (215 +/- 19 min). Antral motility indexes in the first hour after meal ingestion were significantly reduced in CP patients. The interdigestive migrating motor complex cycle length was significantly (P < 0.01) shorter in CP patients (90 +/- 8 min) compared with controls (129 +/- 8 min). These abnormalities were more pronounced in CP patients with exocrine PI. After supplementation of pancreatic enzymes, these alterations in motility reverted toward normal. Digestive and interdigestive antroduodenal motility are abnormal in patients with CP but significantly different from controls only in those with exocrine PI. These abnormalities in antroduodenal motility in CP are related to maldigestion. PMID- 10712267 TI - Medium-chain triglycerides inhibit free radical formation and TNF-alpha production in rats given enteral ethanol. AB - This study determined whether free radical formation by the liver, tumor necrosis factor (TNF)-alpha production by isolated Kupffer cells, and plasma endotoxin are affected by dietary saturated fat. Rats were fed enteral ethanol and corn oil (E CO) or medium-chain triglycerides (E-MCT) and control rats received corn oil (C CO) or medium-chain triglycerides (C-MCT) for 2 wk. E-CO rats developed moderate fatty infiltration and slight inflammation; however, E-MCT prevented liver injury. Serum aspartate aminotransferase levels, gut permeability, and plasma endotoxin doubled with E-CO but were blunted approximately 50% with E-MCT. In Kupffer cells from E-CO rats, intracellular calcium was elevated by lipopolysaccharide (LPS) in a dose-dependent manner. In cells from E-MCT rats, increases were blunted by approximately 40-50% at all concentrations of LPS. The LPS-induced increase in TNF-alpha production by Kupffer cells was dose dependent and was blunted by 40% by MCT. E-CO increased radical adducts and was reduced approximately 50% by MCT. MCT prevent early alcohol-induced liver injury, in part, by inhibition of free radical formation and TNF-alpha production by inhibition of endotoxin-mediated activation of Kupffer cells. PMID- 10712268 TI - A novel in vitro model of Brunner's gland secretion in the guinea pig duodenum. AB - A novel in vitro model that combined functional and morphological techniques was employed to directly examine pathways regulating Brunner's gland secretion in isolation from epithelium. In vitro submucosal preparations were dissected from guinea pig duodenum. A videomicroscopy technique was used to measure changes in luminal diameter of glandular acini as an index of activation of secretion. Carbachol elicited concentration-dependent dilations of the lumen (EC(50) = 2 microM) by activating muscarinic receptors on acinar cells. Ultrastructural and histological analyses demonstrated that dilation was accompanied by single and compound exocytosis of mucin-containing granules and the accumulation of mucoid material within the lumen. Inflammatory mediators (histamine, PGE(1), PGE(2)) and intestinal hormones (CCK, gastrin, vasoactive intestinal polypeptide, secretin) also stimulated glandular secretion, whereas activation of submucosal secretomotor neurons by 5-hydroxytryptamine did not. This study directly demonstrates that multiple hormonal, inflammatory, and neurocrine agents activate Brunner's glands, whereas many have dissimilar effects on the epithelium. This suggests that Brunner's glands are regulated by pathways that act both in parallel to and in isolation from those controlling epithelial secretion. PMID- 10712269 TI - Regulation of adherens junction protein p120(ctn) by 10 nM CCK precedes actin breakdown in rat pancreatic acini. AB - The initial pathophysiological events that characterize CCK-hyperstimulation pancreatitis include the breakdown of the actin filament system and disruption of cadherin-catenin protein complexes. Cadherins and catenins are part of adherens junctions, which may act as anchor for the cellular actin filament system. We examined the composition and regulation of adherens junctions during CCK-induced acinar cell damage. Freshly isolated CCK-stimulated rat pancreatic acini were examined for actin filaments and functional adherens junctions by immunocytology and laser confocal scanning microscopy or by coprecipitation and immunoblotting for E-cadherin, beta- and alpha-catenin, p120(ctn), and phosphotyrosine. In addition to E-cadherin and beta-catenin, acinar cells express the cadherin regulatory protein p120(ctn) and the attachment protein alpha-catenin. Both colocalize and coimmunoprecipitate with E-cadherin in one complex, and all colocalize with the terminal actin web. Supramaximal secretory CCK concentrations (10 nM) initiated tyrosine phosphorylation of p120(ctn) but not of beta-catenin within 2 min, preceding the breakdown of the terminal actin web by several minutes. Under these conditions, the cadherin-catenin association within the adherens junction complex remained intact. We describe for the first time supramaximal CCK-dependent tyrosine phosphorylation of the adherens junction protein p120(ctn) and demonstrate the presence of an intact adherens junction protein complex in acinar cells. p120(ctn) may participate in the actin filament breakdown during experimental conditions mimicking pancreatitis. PMID- 10712270 TI - Domain-specific purinergic signaling in polarized rat cholangiocytes. AB - In cholangiocytes, adenine nucleotides function as autocrine/paracrine signals that modulate ductular ion transport by activation of purinergic receptors. The purpose of these studies was to identify cellular signals that modulate ATP release and nucleotide processing in polarized normal rat cholangiocytes. In Ussing chamber studies, selective exposure of the apical and basolateral membranes to ATP or adenosine 5'-O-(3-thiotriphosphate) (ATPgammaS) stimulated increases in short-circuit current. Apical purinergic receptor agonist preference was consistent with the P2Y(2) subtype. In contrast, basolateral ADP was more potent in stimulating transepithelial currents, consistent with the expression of different basolateral P2 receptor(s). Luminometric analysis revealed that both membranes exhibited constitutive ATP efflux. Hypotonic exposure enhanced ATP release in both compartments, whereas decreases in ATP efflux during hypertonicity were more prominent at the apical membrane. Increases in intracellular cAMP, cGMP, and Ca(2+) also increased ATP permeability, but selective effects on apical and basolateral ATP release differed. Finally, the kinetics of ATP degradation in apical and basolateral compartments were distinct. These findings suggest that there are domain-specific signaling pathways that contribute to purinergic responses in polarized cholangiocytes. PMID- 10712271 TI - Effects of hypoxia, anoxia, and endogenous ethanol on thermoregulation in goldfish, Carassius auratus. AB - Effects of hypoxia, anoxia, and endogenous ethanol (EtOH) on selected temperature (T(sel)) and activity in goldfish were evaluated. Blood and brain EtOH concentrations ([EtOH]) and brain oxygen partial pressure (PO(2)) were quantified at crucial ambient oxygen pressures. Below a threshold value near 31 Torr, T(sel) decreased as a function of environmental PO(2). T(sel) of 15 degrees C-acclimated fish was approximately 10 degrees C at the onset of anoxia and changed little over 2 h. Activity showed a similar response pattern. Brain [EtOH] was significantly elevated above control levels after 1 h anoxia. In normoxic water, T(sel) remained different in previously anoxic and normoxic control fish for approximately 20 min. Blood [EtOH] of previously anoxic fish remained significantly elevated ([EtOH] >4.0 micromol/g blood), and activity was significantly depressed at 20 min. Brain PO(2) reached normal levels in <3 min. We conclude that [EtOH] (brain or blood) and brain PO(2) are not proximal causes of either behavioral anapyrexia (hypothermia) or inactivity in goldfish exposed to oxygen-depleted environments. PMID- 10712272 TI - Norepinephrine turnover in peripheral tissues of rats with heart failure. AB - Experiments were performed to determine if there is regional heterogeneity in sympathetic neural activation of peripheral tissues in rats with chronic heart failure (HF; 6-8 wk after coronary artery ligation). Norepinephrine (NE) turnover, an index of sympathetic activation, was determined on the basis of the decline in tissue NE levels that occurs during the 8-h after tyrosine hydroxylase inhibition (alpha-methyl-DL-p-tyrosine, 300 mg/kg ip at 4-h intervals). Compared with sham-operated rats, NE turnover was increased in the cardiac left ventricle, skeletal muscle, duodenum, and kidney of rats with HF, but was unaltered in liver and spleen. The increased renal NE turnover in HF was largely a reflection of increased turnover in the cortex, with no change evident in the medulla. Blockade of sympathetic ganglionic traffic (hexamethonium, 2 mg/kg sc at 2-h intervals) eliminated the tissue-specific effects of HF on tissue NE levels measured 8-h after tyrosine hydroxylase inhibition. These data support the contention that chronic HF evokes a central nervous system-mediated increase in basal sympathetic tone that exhibits regional heterogeneity (both between and within organs), a phenomenon that likely contributes to the functional consequences of this pathophysiological state. PMID- 10712273 TI - Interstitial pH, K(+), lactate, and phosphate determined with MSNA during exercise in humans. AB - The purpose of the present study was to use the microdialysis technique to simultaneously measure the interstitial concentrations of several putative stimulators of the exercise pressor reflex during 5 min of intermittent static quadriceps exercise in humans (n = 7). Exercise resulted in approximately a threefold (P < 0.05) increase in muscle sympathetic nerve activity (MSNA) and 13 +/- 3 beats/min (P < 0.05) and 20 +/- 2 mmHg (P < 0.05) increases in heart rate and blood pressure, respectively. During recovery, all reflex responses quickly returned to baseline. Interstitial lactate levels were increased (P < 0.05) from rest (1.1 +/- 0.1 mM) to exercise (1. 6 +/- 0.2 mM) and were further increased (P < 0.05) during recovery (2.0 +/- 0.2 mM). Dialysate phosphate concentrations were 0.55 +/- 0. 04, 0.71 +/- 0.05, and 0.48 +/- 0.03 mM during rest, exercise, and recovery, respectively, and were significantly elevated during exercise. At the onset of exercise, dialysate K(+) levels rose rapidly above resting values (4.2 +/- 0.1 meq/l) and continued to increase during the exercise bout. After 5 min of contractions, dialysate K(+) levels had peaked with an increase (P < 0.05) of 0.6 +/- 0.1 meq/l and subsequently decreased during recovery, not being different from rest after 3 min. In contrast, H(+) concentrations rapidly decreased (P < 0.05) from resting levels (69.4 +/- 3.7 nM) during quadriceps exercise and continued to decrease with a mean decline (P < 0.05) of 16.7 +/- 3.8 nM being achieved after 5 min. During recovery, H(+) concentrations rapidly increased and were not significantly different from baseline after 1 min. This study represents the first time that skeletal muscle interstitial pH, K(+), lactate, and phosphate have been measured in conjunction with MSNA, heart rate, and blood pressure during intermittent static quadriceps exercise in humans. These data suggest that interstitial K(+) and phosphate, but not lactate and H(+), may contribute to the stimulation of the exercise pressor reflex. PMID- 10712274 TI - Enalapril and captopril enhance glutathione-dependent antioxidant defenses in mouse tissues. AB - The effect of enalapril and captopril on total glutathione content (GSSG + GSH) and selenium-dependent glutathione peroxidase (Se-GPx) and glutathione reductase (GSSG-Rd) activities was investigated in mouse tissues. CF-1 mice (4-mo-old females) received water containing enalapril (20 mg/l) or captopril (50 mg/l) for 11 wk. Enalapril increased GSSG + GSH content (P < 0.05) in erythrocytes (147%), brain (112%), and lung (67%), and captopril increased GSSG + GSH content in erythrocytes (190%) and brain (132%). Enalapril enhanced Se-GPx activity in kidney cortex (42%) and kidney medulla (23%) and captopril in kidney cortex (30%). GSSG-Rd activity was enhanced by enalapril in erythrocytes (21%), brain (21%), liver (18%), and kidney cortex (53%) and by captopril in erythrocytes (25%), brain (19%), and liver (34%). In vitro erythrocyte oxidant stress was evaluated by thiobarbituric acid-reactive substances (TBARS) production (control 365 +/- 11, enalapril 221 +/- 26, captopril 206 +/- 17 nmol TBARS x g Hb(-1) x h( 1); both P < 0.05 vs. control) and phenylhydrazine-induced methemoglobin (MetHb) formation (control 66.5 +/- 3.5, enalapril 52.9 +/- 0.4, captopril: 56.4 +/- 2.9 micromol MetHb/g Hb; both P < 0.05 vs. control). Both angiotensin-converting enzyme inhibitor treatments were associated with increased nitric oxide production, as assessed by plasma NO-(3) + NO-(2) level determination (control 9.22 +/- 0.64, enalapril 13.7 +/- 1.9, captopril 17.3 +/- 3.0 micromol NO-(3) + NO-(2)/l plasma; both P < 0.05 vs. control). These findings support our previous reports on the enalapril- and captopril-induced enhancement of endogenous antioxidant defenses and include new data on glutathione-dependent defenses, thus furthering current knowledge on the association of ACE inhibition and antioxidants. PMID- 10712275 TI - Myocardial renin is neither necessary nor sufficient to initiate or maintain ventricular hypertrophy. AB - We tested the hypothesis that the myocardial renin-angiotensin system (RAS) is both necessary and sufficient to initiate and maintain all classes of ventricular hypertrophy. Myocardial and plasma renin and angiotensinogen were measured in rats during initiation and maintenance of ventricular hypertrophy associated with DOCA implants and 1% NaCl drinking water, with and without the AT(1) ANG II receptor blocker losartan. Additional groups of rats were given a low-sodium diet (0.04%) for 3 wk. Ventricular hypertrophy was initiated within 7 days and maintained for 35 days in DOCA-treated rats despite significantly low myocardial and plasma renin, normal or low myocardial and plasma angiotensinogen, or the presence of losartan. Furthermore, there was no ventricular hypertrophy in low salt diet-fed animals despite increased myocardial and plasma renin levels and normal angiotensinogen levels. Therefore, the myocardial RAS is not necessary to initiate or maintain cardiac hypertrophy in DOCA-treated rats and is not sufficient to initiate cardiac hypertrophy in low-salt diet-fed rats. Additionally, myocardial renin and angiotensinogen were significantly correlated with corresponding plasma levels. PMID- 10712276 TI - Phosphorylating pathways and fatigue development in contracting Xenopus single skeletal muscle fibers. AB - To investigate the differential contribution of oxidative and substrate-level phosphorylation to force production during repetitive, maximal tetanic contractions, single skeletal muscle fiber performance was examined under conditions of high-oxygen availability and anoxia. Tetanic force development (P) was measured in isolated, single type-1 muscle fibers (fast twitch; n = 6) dissected from Xenopus lumbrical muscle while being stimulated at increasing frequencies (0.25, 0.33, and 0.5 Hz), with each frequency lasting 2 min. Two separate work bouts were conducted, with the perfusate PO(2) being either 0 or 159 mmHg. No significant (P < 0. 05) difference was found in the initial peak tensions (P(0)) between the high (334 +/- 57 kPa) and the low (325 +/- 41 kPa) PO(2) treatment. No significant difference in P was observed between the treatments during the first 50 s. However, a significant difference in force production was observed between the high (P/P(0) = 0.96 +/- 0.02) and the low PO(2) condition (P/P(0) = 0.92 +/- 0.02) by 60 s of work. After 60 s, steady state force production was maintained during the high compared with the low PO(2) condition until stimulation frequency was increased, at which point developed tension during the high PO(2) condition began to decline. Time to fatigue (P/P(0) = 0.3) was reached significantly sooner during the low (250 +/- 16 s) than the high PO(2) condition (367 +/- 28 s). These results demonstrate that during the first 50 s of 0.25-Hz contractions, substrate-level phosphorylation has the capacity to maintain force and ATP hydrolysis when oxidative phosphorylation is absent. This period was followed by an oxygen-dependent phase in which force generation was maintained during the high PO(2) condition (but not during the low PO(2) condition) until the onset of a final fatiguing phase, at which a calculated maximal rate of oxidative phosphorylation was reached. PMID- 10712277 TI - Nociceptin inhibits rat sympathetic preganglionic neurons in situ and in vitro. AB - In vitro and in situ experiments were conducted to evaluate the hypothesis that the nonclassical opioid peptide nociceptin acting on sympathetic preganglionic neurons (SPNs) inhibits spinal sympathetic outflow. First, whole cell patch recordings were made from antidromically identified SPNs from immature (12-16 day old) rat spinal cord slices. Nociceptin (0.1, 0.3, and 1 microM) concentration dependently suppressed the excitatory postsynaptic potentials (EPSPs) evoked by focal stimulation and hyperpolarized a population of SPNs; these effects were naloxone insensitive. L-Glutamate-induced depolarizations were not significantly changed by nociceptin. Results from this series of experiments indicate that nociceptin inhibits the activity of SPNs by either a presynaptic or postsynaptic site of action, whereby the peptide reduces, respectively, the amplitude of EPSPs or the excitability of SPNs. Second, intrathecal injection of nociceptin (3, 10, and 30 nmol) to urethan-anesthetized rats dose dependently reduced the mean arterial pressure and heart rate; these effects were not prevented by prior intravenous administration of naloxone (1 mg/kg). Physiological saline given intrathecally was without appreciable effects. These results, together with earlier observations of the detection of nociceptin-immunoreactive nerve fibers and nociceptin receptor immunoreactivity in the rat intermediolateral cell column, raise the possibility that the opioid peptide, which may be released endogenously, reduces spinal sympathetic outflow by depressing the activity of SPNs. PMID- 10712279 TI - Suppression of food intake, body weight, and body fat by jejunal fatty acid infusions. AB - Three experiments investigated effects of jejunal lipid infusions given on 4 or 21 consecutive days in adult, male Sprague-Dawley rats. In experiment 1, 7-h infusions of linoleic or oleic acid (0.2 ml/h for 7 h; total load = 11.5 kcal) on 4 consecutive days reduced total intake (ad libitum consumption of the liquid diet Boost, Mead Johnson, plus load) by approximately 15% and decreased weight gain compared with 4-day tests with saline administration. In experiment 2, linoleic acid at 0.1 ml/h for 7 h (5.7 kcal) was ineffective, whereas the same load delivered in 3.5 h produced effects similar in magnitude to those in the first experiment. In experiment 3, jejunal infusions of linoleic acid (0.2 ml/h for 7 h) on 21 consecutive days reduced mean total intake by 16%, body weight by 10%, and carcass fat by 48% compared with controls receiving saline. The net decrease in caloric intake may reflect the combined activation of pre- and postabsorptive mechanisms, and it suggests a possible treatment for obesity. PMID- 10712278 TI - Isometric force and endurance in soleus muscle of thyroid hormone receptor alpha(1)- or -beta-deficient mice. AB - The specific role of each subtype of thyroid hormone receptor (TR) on skeletal muscle function is unclear. We have therefore studied kinetics of isometric twitches and tetani as well as fatigue resistance in isolated soleus muscles of R alpha(1)- or -beta-deficient mice. The results show 20-40% longer contraction and relaxation times of twitches and tetani in soleus muscles from TR-alpha(1) deficient mice compared with their wild-type controls. TR-beta-deficient mice, which have high thyroid hormone levels, were less fatigue resistant than their wild-type controls, but contraction and relaxation times were not different. Western blot analyses showed a reduced concentration of the fast-type sarcoplasmic reticulum Ca(2+)-ATPase (SERCa1) in TR-alpha(1)-deficient mice, but no changes were observed in TR-beta-deficient mice compared with their respective controls. We conclude that in skeletal muscle, both TR-alpha(1) and TR-beta are required to get a normal thyroid hormone response. PMID- 10712280 TI - Control of pulmonary surfactant secretion: an evolutionary perspective. AB - Pulmonary surfactant, a mixture consisting of phospholipids (PL) and proteins, is secreted by type II cells in the lungs of all air-breathing vertebrates. Virtually nothing is known about the factors that control the secretion of pulmonary surfactant in nonmammalian vertebrates. With the use of type II cell cultures from Australian lungfish, North American bullfrogs, and fat-tailed dunnarts, we describe the autonomic regulation of surfactant secretion among the vertebrates. ACh, but not epinephrine (Epi), stimulated total PL and disaturated PL (DSP) secretion from type II cells isolated from Australian lungfish. Both Epi and ACh stimulated PL and DSP secretion from type II cells of bullfrogs and fat tailed dunnarts. Neither Epi nor ACh affected the secretion of cholesterol from type II cell cultures of bullfrogs or dunnarts. Pulmonary surfactant secretion may be predominantly controlled by the autonomic nervous system in nonmammalian vertebrates. The parasympathetic nervous system may predominate at lower body temperatures, stimulating surfactant secretion without elevating metabolic rate. Adrenergic influences on the surfactant system may have developed subsequent to the radiation of the tetrapods. Furthermore, ventilatory influences on the surfactant system may have arisen at the time of the evolution of the mammalian bronchoalveolar lung. Further studies using other carefully chosen species from each of the vertebrate groups are required to confirm this hypothesis. PMID- 10712281 TI - Lipopolysaccharide effects on neuronal activity in rat basal forebrain and hypothalamus during sleep and waking. AB - Peripheral administration of lipopolysaccharide (LPS) is associated with alterations in sleep and the electroencephalogram. To evaluate potential neuronal mechanisms for the somnogenic effects of LPS administration, we used unanesthetized rats to survey the firing patterns of neurons in various regions of rat basal forebrain (BF) and hypothalamus during spontaneous sleep and waking and during the epochs of sleep and waking that occurred after the intraperitoneal administration of LPS. In the brain regions studied, LPS administration was associated with altered firing rates in 39% of the neurons examined. A larger proportion of LPS-responsive units showed vigilance-related alterations in firing rates compared with nonresponsive units. Approximately equal proportions of LPS responsive neurons showed increased and decreased firing rates after LPS administration, with some units in the lateral preoptic area of the hypothalamus showing particularly robust increases. These findings are consistent with other studies showing vigilance-related changes in neuronal activity in various regions of BF and hypothalamus and further demonstrate that peripheral LPS administration alters neuronal firing rates in these structures during both sleep and waking. PMID- 10712282 TI - Mg(2+)-induced endothelium-dependent relaxation of blood vessels and blood pressure lowering: role of NO. AB - In vitro extracellular Mg(2+) concentration ([Mg(2+)](0)) produces endothelium dependent and endothelium-independent relaxations in rat aorta in a concentration dependent manner. These relaxant effects of Mg(2+) on intact rat aortic rings, but not denuded rat aortic rings, were suppressed by either N(G)-monomethyl-L arginine (L-NMMA), N(omega)-nitro-L-arginine methyl ester (L-NAME), or methylene blue. The inhibitory effects of L-NMMA and L-NAME could be reversed partly by L arginine. [Mg(2+)](0)-induced dilatation in vivo in rat mesenteric arterioles and venules was almost completely inhibited by N(G)-nitro-L-arginine and L-NMMA. Removal of extracellular Ca(2+) concentration ([Ca(2+)](0)) or buffering intracellular Ca(2+) concentration in endothelial cells, with 10 microM 1, 2 bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-AM, markedly attenuated the relaxant effects of Mg(2+). Mg(2+) produced nitric oxide (NO) release from the intact aortic rings in a concentration-dependent manner. Removal of [Ca(2+)](0) diminished the increased NO release induced by elevated levels of [Mg(2+)](0). In vivo infusion of increasing doses (1-30 microM/min) of MgSO(4), directly into the femoral veins of anesthetized rats, elicited significant concentration-dependent sustained increases in serum total Mg and concomitant decreases in arterial blood pressure. Before and after employment of various doses of MgSO(4), intravenous administration of either L-NMMA (10 mg/kg) or L-NAME (10 mg/kg) increased (i.e., reversed) the MgSO(4)-lowered blood pressure markedly, and intravenous injection of L-arginine restored partially the increased blood pressure effects of both L NMMA and L-NAME. Our results suggest that 1) small blood vessels are very dependent on NO release for Mg(2+) dilatations and 2) the endothelium-dependent relaxation induced by extracellular Mg(2+) is mediated by release of endothelium derived relaxing factor-NO from the endothelium, and requires Ca(2+) and formation of guanosine 3',5'-cyclic monophosphate. PMID- 10712283 TI - Angiotensin stimulates TGF-beta1 and clusterin in the hydronephrotic neonatal rat kidney. AB - Unilateral ureteral obstruction (UUO) induces activation of the renin-angiotensin system and upregulation of transforming growth factor-beta1 (TGF-beta1; a cytokine modulating cellular adhesion and fibrogenesis) and clusterin (a glycoprotein produced in response to cellular injury). This study was designed to examine the regulation of renal TGF-beta1 and clusterin by ANG II in the neonatal rat. Animals were subjected to UUO in the first 2 days of life, and renal TGF beta1 and clusterin mRNA were measured 3 days later. Rats were divided into treatment groups receiving saline vehicle, ANG, losartan (AT(1) receptor inhibitor), or PD-123319 (AT(2) receptor inhibitor). ANG stimulated renal TGF beta1 expression via AT(1) receptors, a response similar to that in the adult. In contrast, clusterin expression was stimulated via AT(2) receptors, a response differing from that in the adult, in which ANG inhibits clusterin expression via AT(1) receptors. We speculate that the unique response of the neonatal hydronephrotic kidney to ANG II is due to the preponderance of AT(2) receptors in the developing kidney. PMID- 10712284 TI - Prandial lactate infusion inhibits spontaneous feeding in rats. AB - To investigate the acute effects of lactate on spontaneous feeding, we infused lactate in the hepatic portal vein (0.5, 1.0, and 1.5 mmol lactate/meal) or in the vena cava (1.0 and 1.5 mmol lactate/meal) of ad libitum-fed rats during their first spontaneous nocturnal meal. Infusions (5 min, 0.1 ml/min) were remotely controlled, and a computerized feeding system recorded meal patterns. In separate crossover tests, meal size decreased independent of the infusion route after 1.0 and 1.5 mmol but not after 0.5 mmol lactate. The subsequent intermeal interval (IMI) tended to decrease only after vena cava infusion of 1.0 mmol lactate. The size of the second nocturnal meal increased after the 1.0 mmol lactate infusion. Hepatic portal infusion of 1.5 mmol lactate increased the satiety ratio [subsequent IMI (min)/meal size (g)] by 175%, which was higher than the insignificant 43% increase after vena cava infusion. Hepatic portal infusion of 1.5 mmol lactate also increased systemic plasma lactate but not glucose concentration at 1 min after the end of infusion. The results are consistent with the idea that meal-induced increases in circulating lactate play a role in the control of meal size (satiation). Moreover, the results suggest that lactate also contributes to postprandial satiety and that the liver is involved in this effect. The exact mechanisms of lactate's inhibitory effects on feeding and the site(s) where lactate acts to terminate meals remain to be identified. PMID- 10712285 TI - Insulin-like growth factor I mediates high-fat diet-induced adipogenesis in Osborne-Mendel rats. AB - To determine if high-fat (HF) diet-induced changes in adipose tissue cellularity are associated with the presence of paracrine growth factor(s) that alter preadipocyte proliferation, Osborne-Mendel rats were fed either a HF (76% energy) or a low-fat (LF, 12% energy) diet for 85 days. HF-fed rats had greater (P < 0.05) fat pad size, total fat cell number, number of small (30-70 microm) and large (80-140 microm) adipocytes, and percentage of 100- to 140-microm adipocytes compared with LF-fed rats. Preadipocytes in primary cell culture treated with inguinal adipose tissue conditioned medium (ATCM) prepared from HF-fed rats had greater (P < 0.05) proliferation compared with cultures treated with ATCM from LF fed rats. Proliferative capacity of ATCM prepared from HF-fed rats was attenuated after the stripping of the medium of insulin-like growth factor I using an immunomagnetic bead separation system. These data are consistent with the concept that insulin-like growth factor I is involved in the paracrine regulation of adipogenesis. PMID- 10712286 TI - Neonatal hypoxic hyperlipidemia in the rat: effects on aldosterone and corticosterone synthesis in vitro. AB - Neonatal hypoxia increases aldosterone production and plasma lipids. Because fatty acids can inhibit aldosterone synthesis, we hypothesized that increases in plasma lipids restrain aldosteronogenesis in the hypoxic neonate. We exposed rats to 7 days of hypoxia from birth to 7 days of age (suckling) or from 28 to 35 days of age (weaned at day 21). Plasma was analyzed for lipid content, and steroidogenesis was studied in dispersed whole adrenal glands untreated and treated to wash away lipids. Hypoxia increased plasma cholesterol, triglycerides, and nonesterified fatty acids in the suckling neonatal rat only. Washing away lipids increased aldosterone production in cells from 7-day-old rats exposed to hypoxia, but not in cells from normoxic 7-day-old rats or from normoxic or hypoxic 35-day-old rats. Addition of oleic or linolenic acid to washed cells inhibited both aldosterone and corticosterone production, although cells from hypoxic 7-day-old rats were less sensitive. We conclude that hypoxia induces hyperlipidemia in the suckling neonate and that elevated nonesterified fatty acids inhibit aldosteronogenesis. PMID- 10712287 TI - Isotope recycling in lactating dogs (Canis familiaris). AB - Isotope-based techniques for the measurement of water turnover, energy expenditure, and milk intake often assume that there is no recycling of isotopes once they have left the labeled animal. In experiments involving lactating females or their suckling offspring, there are several possible routes of isotope recycling. These include the consumption of labeled milk by offspring, the ingestion of labeled excreta, and the rebreathing of exhaled labeled CO(2) or water vapor by both mother and offspring. Isotope recycling might be especially important during lactation because the offspring are in close contact with each other and their mother for prolonged periods. We show here in 24- to 30-day-old domestic dog Canis familiaris puppies that there was no detectable transfer of (18)O or (2)H from labeled to unlabeled pups in two litters (16 pups, 8 labeled, 8 unlabeled) that were weaned early and independent of their mother. However, there was a significant transfer of both isotopes from labeled to unlabeled pups and from labeled pups to their mothers in nine equivalent nursing litters of the same age (27 labeled, 26 unlabeled pups). The increases in enrichment of isotopes in unlabeled offspring were greater than the increases in enrichment of the mothers. This indicates that maternal ingestion of offspring excreta and subsequent transfer of isotope in milk is not the sole pathway of recycling. Additional routes must also be important, such as exchange of isotope between pups on saliva-coated nipples and perhaps direct ingestion of excreta by unweaned young. Recycling is unlikely to be an important factor when determining maternal metabolic rate during peak lactation in domestic dogs. However, experiments that are designed to assess the energy demands of pups and isotope-based estimates of water turnover in offspring may need to take into account any effects of isotope recycling. In a theoretical example, removing the effects of recycling increased the measured energy expenditure in pups by up to 7% and increased the calculated elimination rates of both isotopes by up to 11.1% in (18)oxygen and 10.9% in (2)hydrogen. PMID- 10712288 TI - Denudations as paracellular routes for alphafetoprotein and creatinine across the human syncytiotrophoblast. AB - We tested two hypotheses: 1) that fibrin-containing fibrinoid-filled denudations of the syncytiotrophoblast may provide a route for paracellular diffusion and 2) that placentas from women who had elevated maternal serum alphafetoprotein (MSAFP) in midgestation had raised permeability to AFP and greater denudation than in normal pregnancy. We measured AFP and creatinine clearance across term placental cotyledons from the above groups and used light microscope morphometric analysis to determine the volume density of fibrin-containing fibrinoid deposits. There was no significant difference between the two groups in terms of AFP and creatinine clearance or volume density of fibrin-containing fibrinoid deposits. The combined data showed a significant (P < 0.05) positive correlation between creatinine clearance, but not AFP clearance, and volume density of fibrin containing fibrinoid. We conclude that syncytiotrophoblast denudations, with associated fibrinoid, do provide a route for diffusion of small hydrophilic solutes, but that other anatomic features of the placenta are rate limiting for transfer of AFP and similarly sized molecules. PMID- 10712289 TI - The neurosteroid allopregnanolone modulates oxytocin expression in the hypothalamic paraventricular nucleus. AB - Virgin, ovariectomized rats exposed to 2 wk of sequential estradiol (E(2)) and progesterone (P) followed by P withdrawal have increased hypothalamic oxytocin (OT) mRNA and peptide levels relative to sham-treated animals. This increase is prevented if P is sustained. In the central nervous system, P is metabolized to the neurosteroid allopregnanolone (3alpha-hydroxy-5alpha-pregnan-20-one), which exerts effects by acting as a positive allosteric modulator of GABA(A) receptor/Cl(-)-channel complexes. In the present study, ovariectomized rats that received sequential E(2) and P for 2 wk followed by P withdrawal were administered allopregnanolone at the time of P withdrawal. Hypothalamic and plasma allopregnanolone concentrations, serum E(2) and P concentrations, and hypothalamic OT mRNA levels were measured at death. Steroid-induced increases in OT mRNA were attenuated in animals treated with allopregnanolone at the time of P withdrawal. The results suggest that allopregnanolone plays an important modulatory role in steroid-mediated increases in hypothalamic OT. PMID- 10712290 TI - Pressor effects of orexins injected intracisternally and to rostral ventrolateral medulla of anesthetized rats. AB - Orexin A and B, two recently isolated hypothalamic peptides, have been reported to increase food consumption upon intracerebroventricular injections in rats. In addition to the hypothalamus, orexin A-immunoreactive fibers have been observed in several areas of the medulla that are associated with cardiovascular functions. The present study was undertaken to evaluate the hypothesis that orexins may influence cardiovascular response by interacting with neurons in the medulla. Intracisternal injections of orexins A (0.0056-7.0 nmol) or B (0.028 0.28 nmol) dose dependently increased mean arterial pressure (MAP) by 4-27 mmHg and heart rate (HR) by 26-80 beats/min in urethan-anesthetized rats, with orexin A being more effective in this regard. MAP and HR were not changed by intravenous injection of orexins at higher concentrations. Microinjection of orexin A (14 pmol/50.6 nl) to the rostral ventrolateral medulla, which was confirmed by histological examination, increased MAP and HR. Our results indicate that, in addition to a role in positive feeding behavior, orexins may enhance cardiovascular response via an action on medullary neurons. PMID- 10712291 TI - Regulation of body temperature and energy requirements of hibernating alpine marmots (Marmota marmota). AB - Body temperature and metabolic rate were recorded continuously in two groups of marmots either exposed to seasonally decreasing ambient temperature (15 to 0 degrees C) over the entire hibernation season or to short-duration temperature changes during midwinter. Hibernation bouts were characterized by an initial 95% reduction of metabolic rate facilitating the drop in body temperature and by rhythmic fluctuations during continued hibernation. During midwinter, we observed a constant minimal metabolic rate of 13.6 ml O(2) x kg(-1) x h(-1) between 5 and 15 degrees C ambient temperature, although body temperature increased from 7.8 to 17.6 degrees C, and a proportional increase of metabolic rate below 5 degrees C ambient temperature. This apparent lack of a Q(10) effect shows that energy expenditure is actively downregulated and controlled at a minimum level despite changes in body temperature. However, thermal conductance stayed minimal (7.65 +/ 1.95 ml O(2) x kg(-1) x h(-1) x degrees C(-1)) at all temperatures, thus slowing down cooling velocity when entering hibernation. Basal metabolic rate of summer active marmots was double that of winter-fasting marmots (370 vs. 190 ml O(2) x kg(-1) x h(-1)). In summary, we provide strong evidence that hibernation is not only a voluntary but a well-regulated strategy to counter food shortage and increased energy demands during winter. PMID- 10712292 TI - Skeletal and cardiac muscle protein turnover during cold acclimation in young rats. AB - The effect of long-term cold exposure on skeletal and cardiac muscle protein turnover was investigated in young growing animals. Two groups of 36 male 28-day old rats were maintained at either 5 degrees C (cold) or 25 degrees C (control). Rates of protein synthesis and degradation were measured in vivo on days 5, 10, 15, and 20. Protein mass by day 20 was approximately 28% lower in skeletal muscle (gastrocnemius and soleus) and approximately 24% higher in heart in cold compared with control rats (P < 0.05). In skeletal muscle, the fractional rates of protein synthesis (k(syn)) and degradation (k(deg)) were not significantly different between cold and control rats, although k(syn) was lower (approximately -26%) in cold rats on day 5; consequent to the lower protein mass, the absolute rates of protein synthesis (approximately -21%; P < 0. 05) and degradation (approximately 13%; P < 0.1) were lower in cold compared with control rats. In heart, overall, k(syn) (approximately +12%; P < 0.1) and k(deg) (approximately +22%; P < 0.05) were higher in cold compared with control rats; consequently, the absolute rates of synthesis (approximately +44%) and degradation (approximately +54%) were higher in cold compared with control rats (P < 0.05). Plasma triiodothyronine concentration was higher (P < 0.05) in cold compared with control rats. These data indicate that long-term cold acclimation in skeletal muscle is associated with the establishment of a new homeostasis in protein turnover with decreased protein mass and normal fractional rates of protein turnover. In heart, unlike skeletal muscle, rates of protein turnover did not appear to immediately return to normal as increased rates of protein turnover were observed beyond day 5. These data also indicate that increased rates of protein turnover in skeletal muscle are unlikely to contribute to increased metabolic heat production during cold acclimation. PMID- 10712294 TI - Functional status of the regenerated chorda tympani nerve as assessed in a salt taste discrimination task. AB - We tested whether the recovered ability of rats to discriminate NaCl from KCl after chorda tympani nerve transection (CTX) is causally linked to nerve regeneration or some other compensatory process. Rats were presurgically trained in an operant NaCl vs. KCl discrimination task. Rats with regenerated nerves, histologically confirmed by anterior tongue taste pore counts and tested 62 days after CTX (CTX-62R; n = 5), performed as well as those tested 62 days after sham surgery (Sham-62; n = 5), but both of these groups initially performed slightly worse than animals tested 7 days after sham surgery (Sham-7; n = 4). Performance of rats tested either 7 (CTX-7P; n = 5) or 62 (CTX-62P; n = 4) days after CTX in which nerve regeneration was prevented was severely disrupted. Adulteration of the stimuli with amiloride, an epithelial sodium channel blocker, impaired discrimination performance in a similar dose-dependent manner in the Sham-7 (n = 2), Sham-62 (n = 5), and CTX-62R (n = 5) groups, suggesting that the functional status of the amiloride-sensitive transduction pathway returns to normal in rats with regenerated chorda tympani nerves. Performance of CTX rats without regenerated nerves (CTX-7P, n = 2; CTX-62P, n = 4) was further degraded by amiloride treatment, suggesting that taste receptors innervated by other nerves are sensitive to amiloride. In conclusion, nerve regeneration is an essential component underlying full recovery of salt discrimination function after CTX. PMID- 10712293 TI - Sucrose consumption increases naloxone-induced c-Fos immunoreactivity in limbic forebrain. AB - Opioids have long been known to have an important role in feeding behavior, particularly related to the rewarding aspects of food. Considerable behavioral evidence suggests that sucrose consumption induces endogenous opioid release, affecting feeding behavior as well as other opioid-mediated behaviors, such as analgesia, dependence, and withdrawal. In the present study, rats were given access to a 10% sucrose solution or water for 3 wk, then they were injected with 10 mg/kg naloxone or saline. Brains were subsequently analyzed for c-Fos immunoreactivity (c-Fos-IR) in limbic and autonomic regions in the forebrain and hindbrain. Main effects of sucrose consumption or naloxone injection were seen in several areas, but a significant interaction was seen only in the central nucleus of the amygdala and in the lateral division of the periaqueductal gray. In the central nucleus of the amygdala, naloxone administration to those rats drinking water significantly increased c-Fos-IR, an effect that was significantly enhanced by sucrose consumption, suggesting an upregulation of endogenous opioid tone in this area. The data from this study indicate that the central nucleus of the amygdala has a key role in the integration of gustatory, hedonic, and autonomic signals as they relate to sucrose consumption, if not to food intake regulation in general. Furthermore, the data from this study lend further support to the hypothesis that sucrose consumption induces the release of endogenous opioids. PMID- 10712295 TI - Vasopressin and oxytocin release evoked by NaCl loads are selectively blunted by area postrema lesions. AB - The present study investigated the effect of area postrema lesions (APX) on stimulated neurohypophysial secretion of vasopressin (VP) and oxytocin (OT) in conscious rats. Blunted increases in plasma levels of both pituitary hormones were observed when rats with APX were infused intravenously with 1 M NaCl solution (2 ml/h for 6 h). In contrast, plasma VP and OT increased normally in rats with APX when equivalent increases in plasma osmolality (but not plasma Na(+)) resulted from intravenous infusion of an equiosmotic solution of 1 M mannitol and 0.5 M NaCl. Furthermore, APX did not affect increases in plasma VP and OT stimulated by plasma volume deficits, nor did APX disrupt OT secretion stimulated by intravenous injection of cholecystokinin. These findings suggest that the area postrema plays an important role in mediating secretion of VP and OT in response to an NaCl load, but not in response to an equiosmotic load that does not cause substantial hypernatremia, and not in response to other stimuli of neurohypophysial hormone secretion. Together with previous reports, these results suggest that APX impairs Na(+) regulation in rats. PMID- 10712296 TI - Functional link between distal vasodilation and sleep-onset latency? AB - Thermoregulatory processes have long been implicated in initiation of human sleep. The purpose of this study was to evaluate the role of heat loss in sleep initiation, under the controlled conditions of a constant-routine protocol modified to permit nocturnal sleep. Heat loss was indirectly measured by means of the distal-to-proximal skin temperature gradient (DPG). A stepwise regression analysis revealed that the DPG was the best predictor variable for sleep-onset latency (compared with core body temperature or its rate of change, heart rate, melatonin onset, and subjective sleepiness ratings). This study provides evidence that selective vasodilation of distal skin regions (and hence heat loss) promotes the rapid onset of sleep. PMID- 10712297 TI - Hyperthermia stimulates energy-proteasome-dependent protein degradation in cultured myotubes. AB - Previous studies suggest that elevated temperature stimulates protein degradation in skeletal muscle, but the intracellular mechanisms are not fully understood. We tested the role of different proteolytic pathways in temperature-dependent degradation of long- and short-lived proteins in cultured L6 myotubes. When cells were cultured at different temperatures from 37 to 43 degrees C, the degradation of both classes of proteins increased, with a maximal effect noted at 41 degrees C. The effect of high temperature was more pronounced on long-lived than on short lived protein degradation. By using blockers of individual proteolytic pathways, we found evidence that the increased degradation of both long-lived and short lived proteins at high temperature was independent of lysosomal and calcium mediated mechanisms but reflected energy-proteasome-dependent degradation. mRNA levels for enzymes and other components of different proteolytic pathways were not influenced by high temperature. The results suggest that hyperthermia stimulates the degradation of muscle proteins and that this effect of temperature is regulated by similar mechanisms for short- and long-lived proteins. Elevated temperature may contribute to the catabolic response in skeletal muscle typically seen in sepsis and severe infection. PMID- 10712298 TI - Siberian hamsters that fail to reentrain to the photocycle have suppressed melatonin levels. AB - Siberian hamsters readily reentrain to a 3-h phase delay of the photocycle (16 h light/day) but fail to reentrain to a 5-h phase delay. This study tested whether melatonin production was suppressed in animals that failed to reentrain. Melatonin was measured on the day before, day of, or several days after each phase shift. Melatonin levels measured 4 h after dark onset were approximately 83 microg/ml on the day before each phase delay and undetectable (<6 microg/ml) during the light phase on the day of the phase shift. Activity onsets regained their prior phase relationship to the photocycle 4 (3 h) or 5 (5 h) days after the phase shift; on that day, melatonin levels were measured 4 h after dark onset. Melatonin levels were unaffected by the 3-h phase delay (>57.6 microg/ml) but were undetectable after a 5-h phase delay (<8 microg/ml). Thus melatonin remained suppressed only after the phase delay to which hamsters also fail to reentrain. This relationship suggests that the propensity for reentrainment may be influenced by changes in melatonin production following a phase shift of the photocycle. PMID- 10712299 TI - Apoptosis in polycystic kidney disease: involvement of caspases. AB - Polycystic kidney disease (PKD) is characterized by the development of large renal cysts and progressive loss of renal function. Although the cause of the development of renal cysts is unknown, recent evidence suggests that excessive apoptosis occurs in PKD. With the use of terminal deoxynucleotidyl transferase dUTP nick-end labeling staining, we have confirmed the presence of apoptotic bodies in cystic kidneys of congenital polycystic kidney (cpk) disease mice carrying a homozygous mutation at 3 wk of age. Apoptosis was localized primarily to the interstitium with little evidence of cell death in cyst epithelium or noncystic tubules. In addition, we observed that the expression of various caspases, bax and bcl-2, was upregulated in cystic kidneys. With the use of various substrates in enzyme activity assays, we have demonstrated a greater than sevenfold increase in caspase 4 activity and a sixfold increase in caspase 3 activity. These data suggest that there is a caspase-dependent apoptosis pathway associated with PKD and support the hypothesis that apoptotic cell death contributes to cyst formation in PKD. PMID- 10712300 TI - Regulation of atrial natriuretic peptide gene expression in gastric antrum by fasting. AB - Atrial natriuretic peptide (ANP) gene expression was localized in the rat gastric antrum using immunohistochemistry and in situ hybridization to mucosal cells in the lower portion of the antropyloric glands. Colocalization of immunoreactive ANP, long-acting natriuretic peptide, i.e., proANP-(1-30), and serotonin in these cells identified them to be enterochromaffin cells. Fasting for 72 h in 8-mo-old (adult) rats produced a significant (P < 0.05) decrease in the levels of ANP prohormone mRNA, immunoreactive proANP-(1-30) and ANP to approximately 33% of that of fed rats. Fasting in 1-mo-old rats had no effect on these parameters. Transcripts for natriuretic peptide receptor subtypes NPR-A, NPR-B, and NPR-C were found in both mucosa and muscle tissues of the antrum. ANP, brain natriuretic peptide (BNP), and C-type natriuretic peptide (CNP) stimulated the production of cGMP in antral mucosa in vitro with a potency of ANP > BNP >> CNP, suggesting that these receptors were functional. We conclude that fasting decreases ANP prohormone mRNA and its gene products, long-acting natriuretic peptide, and ANP in the antrum of adult rats. PMID- 10712301 TI - Expression and localization of angiotensin subtype receptor proteins in the hypertensive rat heart. AB - The cellular localization of the AT(2) receptor and the regulation of its expression in hypertrophied left ventricle are not well known. We compared the expression of the cardiac AT(1) and AT(2) receptor in spontaneously hypertensive rats/Izumo strain (SHR/Izm) and Wistar Kyoto rats/Izumo strain (WKY/Izm), ages 4, 12, and 20 wk, by means of immunohistochemistry and Western blot analysis. In SHR/Izm, compared with WKY/Izm, blood pressure (161 +/- 2 vs. 120 +/- 2 mmHg at 12 wk, P /= 80% of predicted values yielded 84% sensitivity and 73% specificity. Post-bronchodilator changes in R(0)(SD) [DeltaR(0)(SD)] were mostly correlated to changes in MEF(50). The optimal DeltaR(0)(SD) cutoff value to discriminate between children with the presence or absence of significant reversibility in FEV(1) yielded 69% sensitivity and 78% specificity. In children unable to perform forced expiratory maneuvers (n = 132), this DeltaR(0)(SD) cutoff clearly identified a subgroup of young children with high R(0) values at baseline, that returned to normal after bronchodilation. We conclude that FOT measurements allow reliable evaluation of bronchial obstruction and its reversibility in asthmatic children over 3 yr old. PMID- 10712316 TI - Psychological profile and ventilatory response to inspiratory resistive loading. AB - The purpose of this study was to explore the contribution of psychological state to both the ventilatory response and the intensity of dyspnea experienced after the addition of small inspiratory loads to breathing. We hypothesized that patients with either a specific psychiatric diagnosis or a specific psychological trait will associate a greater degree of dyspnea with a loaded breathing task than will control subjects. To insure the inclusion of persons with relevant psychological profiles, we recruited both subjects enrolled in the Chronic Fatigue Center and normal control subjects. In all, 52 subjects inspired first through a small (1.34 cm H(2)O/L/s) and second through a moderate (3.54 cm H(2)O/L/s) inspiratory resistive load (IRL). Ventilation was monitored throughout the 5-min sessions. Dyspnea was quantified with the Borg scale at specified times during the protocol. Standard psychological tests were administered. We found that subjects could be divided into two groups. One, the "responders," reported Borg scores higher than those of the second, or "nonresponder" group, at all times during the protocol. By contrast, there was no difference between groups with respect to ventilation. Responders had higher scores on tests of depression (the Center for Epidemiological Study depression scale) than did nonresponders. We conclude that the variability observed in subjective responses to IRL is explained, in part, by differences in psychological state. PMID- 10712317 TI - Characterization of nonresponse to high caloric oral nutritional therapy in depleted patients with chronic obstructive pulmonary disease. AB - Nutritional support can increase body weight and physiologic function in COPD, but there are some patients who do not respond to nutritional therapy. The aim of this prospective study was to describe the nonresponse to 8 wk of oral nutritional supplementation therapy (500 to 750 kcal/d extra), implemented in an inpatient pulmonary rehabilitation program, with respect to lung function, body composition, energy balance, and systemic inflammatory profile in 24 (16 male) depleted patients with COPD. On the basis of the weight change after 8 wk, patients were divided into three groups (Group 1: weight gain < 2% of baseline body weight, n = 5; Group 2: weight gain 2 to 5%, n = 9; Group 3: weight gain >/= 5%, n = 10). Although no differences were seen in lung function and body composition, Group 1 was characterized by older age, a lower baseline dietary intake/resting energy expenditure (REE) ratio, and a greater number of users of continuous supplemental oxygen when compared with Group 3. In addition, Group 1 exhibited higher baseline concentrations of fasting glucose and LPS-binding protein than did Groups 2 and 3. The concentrations of the soluble TNF- receptors 55 and 75 were elevated in Groups 1 and 2 when compared with Group 3. Furthermore, a significant, inverse correlation coefficient between baseline dietary intake and soluble intercellular adhesion molecule was revealed (r = 0.50, p = 0.016). On linear regression analysis, age, baseline intake/REE ratio, sTNF-receptor 55, and extracellular/intracellular water (ECW/ICW) ratio were selected as independent, significant parameters contributing to a total explained variation of 78% in weight change after nutritional therapy. In conclusion, nonresponse to nutritional therapy in COPD is associated with ageing, relative anorexia, and an elevated systemic inflammatory response. Further research is needed to investigate whether these factors contribute to eventual disturbances in intermediary metabolism as reflected by the increased glucose concentration and ECW/ICW ratio. PMID- 10712318 TI - Treatment of gram-positive nosocomial pneumonia. Prospective randomized comparison of quinupristin/dalfopristin versus vancomycin. Nosocomial Pneumonia Group. AB - Nosocomial pneumonia is a frequent complication in hospitalized patients. Gram positive pathogens, particularly Staphylococcus aureus, are responsible for the increasing frequency of nosocomial pneumonia. To evaluate the efficacy and safety of intravenous quinupristin/dalfopristin (Synercid) in the treatment of nosocomial pneumonia caused by gram-positive pathogens we conducted a prospective, randomized, open-label, international, multicenter, comparative clinical trial. Two hundred ninety-eight patients with nosocomial pneumonia were enrolled in 74 active centers in five countries: a subgroup of 171 (87 quinupristin/dalfopristin-treated and 84 vancomycin-treated patients) were evaluable for the major efficacy end points. One hundred fifty received 7.5 mg/kg of quinupristin/dalfopristin every 8 h; 148 patients received 1 g of vancomycin every 12 h. Aztreonam at a dose of 2 g every 8 h could be administered in both groups for coverage of gram-negative organisms, and tobramycin was added for coverage against Pseudomonas aeruginosa. The primary efficacy end point was the clinical response between the seventh and the thirteenth day after the end of treatment in clinically evaluable patients with documented causative pathogen(s) at baseline (bacteriologically evaluable population). Therapy was clinically successful (cure or improvement) in 49 (56.3%) of patients receiving quinupristin/dalfopristin and 49 (58.3%) patients receiving vancomycin (difference, -2.0% [95% CI, -16.8% to 12.8%]) in the bacteriologically evaluable population. Equivalent clinical success rates were also observed in the all treated population (n = 298), and in the bacteriologically evaluable patients intubated in baseline (39/72 [54%] compared with 36/67 [54%]). The by-pathogen bacteriologic response was similar in both treatment groups, with equivalent clinical success rates for Streptococcus pneumoniae, Staphylococcus aureus, and methicillin-resistant Staphylococcus aureus. Adverse events (venous and nonvenous) were equally distributed between groups; 15.3% of quinupristin/dalfopristin patients and 9.5% of vancomycin patients discontinued therapy because of an adverse clinical event. In this study quinupristin/dalfopristin was shown to be equivalent to vancomycin in the treatment of nosocomial pneumonia caused by gram-positive pathogens. Quinupristin/dalfopristin merits further evaluation for the treatment of nosocomial pneumonia caused by methicillin-resistant S. aureus. PMID- 10712319 TI - Assessment of bronchodilator responsiveness in infants with bronchiolitis. A comparison of the tidal and the raised volume rapid thoracoabdominal compression technique. AB - Whether bronchodilators should be used for the treatment of infants with bronchiolitis is subject to debate, partly because of the low sensitivity of the methods for assessing lung function changes in infants. In the present study, we compared the recently introduced raised volume (RVRTC) with the conventional end tidal rapid thoracoabdominal compression (ETRTC) technique in infants with acute viral bronchiolitis. In 17 infants lung function was assessed by both methods, at baseline values and after salbutamol inhalation. Forced expiratory volumes (FEV(0.5), FEV(0.75), FEV(1.0)) were used for the quantification of RVRTC measurement, and maximal expiratory flow at functional residual capacity (Vmax (FRC)) for ETRTC measurements. A significant individual change was defined by a mean postbronchodilator value that differed from baseline value by more than twice the within-subject coefficient of variation (CV). Group mean intrasubject CVs ranged from 4.7% to 5.3% for FEV parameters; it was 14.0% for Vmax (FRC). For the group, post-bronchodilator measurements did not differ significantly from baseline measurements. For the majority of infants, however, the within-subject comparison of responses revealed substantial differences between both techniques; while no infant demonstrated a significant increase in Vmax (FRC), eight (47%) infants responded with significantly improved timed volumes. The RVRTC technique provides the investigator with a more sensitive diagnostic tool for documenting the effectiveness of therapeutic interventions on an individual basis. Furthermore, the findings of the present study provide a rationale for the application of bronchodilators in a subgroup of infants with acute bronchiolitis. PMID- 10712320 TI - Interleukin-8 secretion and neutrophil recruitment accompanies induced sputum eosinophil activation in children with acute asthma. AB - Although airway inflammation is recognized as a key feature of asthma, the characteristics of airway inflammation in children with acute severe asthma are not well defined. The aim of this study was to describe the characteristics of airway inflammation in children with an acute exacerbation of asthma using sputum cell counts and fluid-phase measurements and to examine the changes in these parameters upon resolution of the exacerbation. Children (n = 38) presenting to the Emergency Department with acute asthma underwent successful sputum induction using ultrasonically nebulized normal saline (n = 22), or expectorated sputum spontaneously (n = 16). Sputum induction was repeated at least 2 wk later when the children had recovered (n = 28). Sputum portions were selected, dispersed and total and differential cell counts performed. Neutrophil elastase and EG2 positive eosinophils were assessed and fluid-phase eosinophil cationic protein (ECP), myeloperoxidase (MPO), interleukin-8 (IL-8), and IL-5 were measured. During the acute exacerbation the median (range) total cell count was 8.4 x 10(6)/ml (0.5 to 190.3), and fell significantly at resolution to 1.3 x 10(6)/ml (p < 0.01). The inflammatory cell infiltrate was mixed and included eosinophils (0.8 x 10(6)/ml), neutrophils (3.3 x 10(6)/ml), and mast cells. EG2(+) cells were high and correlated with the degree of airflow obstruction (r = -0.5, p = 0.02). They decreased significantly at resolution as did supernatant ECP (1,078 versus 272 ng/ml), suggesting that eosinophils were activated during the exacerbation. MPO was 220 ng/ ml at exacerbation and fell significantly to 1 ng/ml at resolution. Levels of IL-8 and IL-5 were elevated during the acute exacerbation and IL-8 concentrations decreased at resolution. In conclusion, airway inflammation can be studied in children with acute asthma by sputum induction. Airway inflammation is present during an acute exacerbation of asthma, and is characterized by infiltration and activation of both eosinophils and neutrophils. The heterogeneity of airway inflammation in acute asthma may influence response to corticosteroid therapy. PMID- 10712321 TI - Effect of a dopexamine-induced increase in cardiac index on splanchnic hemodynamics in septic shock. AB - In 12 patients with hyperdynamic septic shock we studied the effect of dopexamine, a combined dopamine and beta-adrenergic agonist, on hepatosplanchnic hemodynamics and O(2) exchange. All patients required noradrenaline to maintain mean arterial pressure > 60 mm Hg (noradrenaline >/= 0.04 microg x kg(-1) x min( 1)) with a cardiac index >/= 3.0 L/min/m(2). Splanchnic blood flow (Qspl) was measured using primed continuous infusion of indocyanine green dye with hepatic venous sampling. In addition tonometric gastric mucosal-arterial and gastric mucosal-hepatic venous P CO(2) gradients were assessed as indicators of regional energy balance. After 90 min of stable hemodynamics a first measurement was obtained. Then dopexamine infusion was titrated (1-4 microg x kg(-1) x min(-1)) to increase cardiac output by approximately 25% (20-30%). After 90 min all measurements were repeated, and dopexamine was withdrawn followed by a third measurement. Median Qspl (0.86/1.23-0. 66 versus 0.96/1.42-0.85 L/min/m (2) [median value/25th-75th percentiles], p < 0.05) increased whereas the fractional contribution of Qspl to total blood flow decreased (21/28-13 versus 19/28-12%, p < 0.05). Although both global and regional oxygen delivery (DO(2)) consistently increased, neither global or regional V O(2) nor PCO(2) gradients were significantly affected. In patients with septic shock and ongoing noradrenaline treatment dopexamine seems to have no preferential effects on hepatosplanchnic hemodynamics, O(2) exchange, or energy balance. PMID- 10712322 TI - Tuberculosis screening of immigrants to low-prevalence countries. A cost effectiveness analysis. AB - All adult immigrant applicants to Canada undergo chest radiographic screening for tuberculosis (TB). Tuberculin skin testing could reduce the number of chest X rays, and identify more candidates for prophylaxis. We modeled the cost effectiveness of chest radiography and tuberculin skin testing for TB prevention over a 20-yr time frame, among three simulated cohorts of 20-yr-old immigrants. Compared with no screening, radiographic screening prevented 4.3% of expected active TB cases in the highest risk cohort (50% TB-infected, 10% human immunodeficiency virus [HIV] seroprevalence), and 8.0% in the lowest risk cohort (5% TB-infected, 1% HIV seroprevalence). Tuberculin skin testing further reduced the expected incidence 8.0% and 4.0%, respectively. Compared with no screening, radiographic screening cost $3,943 Canadian per active TB case prevented in the highest risk cohort, and $236,496 per case prevented in the lowest risk group. Compared with radiographic screening, mass tuberculin skin testing cost $32,601 per additional case prevented in the highest risk group, and $68,799 per additional case prevented in the lowest risk group. Chest radiographic screening of young immigrants from countries with a high prevalence of TB is a relatively inexpensive means of TB prevention. Tuberculin skin testing is considerably less cost-effective. For immigrants from low-prevalence countries, both interventions are extremely costly with negligible impact. The cost-effectiveness of either strategy would be greatly enhanced by increased adherence to chemoprophylaxis recommendations. Radiographic screening of groups with a high prevalence of tuberculous infection will then likely save money. PMID- 10712323 TI - Serum carotenoids, alpha-tocopherol, and lung function among Dutch elderly. AB - Antioxidant vitamins (provitamins) may protect against loss of lung function over time. We studied the association between serum carotenoids (alpha-carotene, beta carotene, lycopene, beta-cryptoxanthin, zeaxanthin, and lutein), alpha tocopherol, and lung function among noninstitutionalized Dutch elderly age 65 to 85 yr (n = 528). Multiple linear regression analysis was performed with FEV(1) or FVC as dependent variables and serum levels of antioxidants in quintiles as independent variables. We adjusted for age, gender, height, and pack-years of smoking. Subjects in the fifth quintile of serum beta-carotene had a 195 ml (95% confidence interval [95% CI]: 40 to 351 ml) higher and those in the fifth quintile of alpha-carotene had a 257 ml (95% CI: 99 to 414 ml) higher FEV(1) compared with subjects in the first quintile of these carotenoids. Significant (p < 0.05) positive trends were observed between alpha-carotene, beta-carotene, lycopene, and FEV(1) and between alpha-carotene, beta-carotene, and FVC. Subjects in the highest quintile of the other carotenoids or alpha-tocopherol did not have significantly higher FEV(1) or FVC compared with subjects in the first quintile of these antioxidants. In conclusion, this study shows that from the six major serum carotenoids and alpha-tocopherol studied, particularly alpha-carotene, beta carotene, and lycopene were positively associated with lung function in the elderly and may be considered as candidates for further investigations. PMID- 10712324 TI - Feasibility of a clinical trial of augmentation therapy for alpha(1)-antitrypsin deficiency. The Alpha 1-Antitrypsin Deficiency Registry Study Group. AB - We examined the feasibility of a randomized clinical trial of intravenous augmentation therapy for individuals with alpha 1-antitrypsin (alpha1AT) deficiency, basing calculations on newly available data obtained from the NHLBI Registry of Patients with Severe Deficiency of Alpha 1-Antitrypsin. Using rate of FEV(1) decline as the primary outcome and adjusting for noncompliance, a study of subjects with Stage II chronic obstructive pulmonary disease (COPD) (initial FEV(1) 35 to 49% predicted) with biannual spirometry measures obtained over 4 yr of follow-up would require 147 subjects per treatment arm to detect a difference in FEV(1) decline of 23 ml/yr (i.e., a 28% reduction), the difference observed in the NHLBI Registry (1-sided test, alpha = 0.05, 90% power). To detect a 40% reduction in mortality in a 5-year study of subjects with baseline FEV(1) 35 to 49% predicted, recruited over the first 2 yr and then followed an additional 3 yr, 342 subjects per treatment arm would be needed. Though significant impediments to carrying out a clinical trial exist, including the cost of such a trial and the potential difficulties in recruiting patients for a placebo controlled trial, we recommend a randomized controlled trial as the best method to evaluate the efficacy of intravenous augmentation therapy and of possible future treatments. PMID- 10712325 TI - Respiratory effects of environmental tobacco smoke in a panel study of asthmatic and symptomatic children. AB - The effect of environmental tobacco smoke (ETS) on respiratory health was investigated among 7- to 12-yr-old children with asthmatic (n = 74) or cough (n = 95) symptoms. For 3 mo the children measured their peak expiratory flow rate (PEFR) every morning and evening, and kept a daily diary of respiratory symptoms. They also noted daily whether they had used respiratory medication and whether someone had smoked inside their home. Eleven percent of the asthmatic children and 14% of the children with cough had exposure to ETS at home during the study. In multiple regression and analyses controlling for potential confounders, any exposure to ETS during the study was associated with a reduction of 42 L/min (95% confidence interval [CI]: 10 to 74 L/min) in morning and 41 L/min (95% CI: 8 to 74 L/min) in evening PEFR among asthmatic children. Among these children, a dose dependent increase in the effect of ETS was also seen. Daily variation in ETS exposure was only weakly (-9.2 L/min; 95% CI: 2.9 to 21.2 L/min) associated with PEFR, but the previous day's ETS exposure was a risk factor for bronchodilator use (relative risk [RR]: 10.3; 95% CI: 1.3 to 83.7), as well as for cough (RR: 12.4; 95% CI: 2.4 to 63.3) and phlegm production (RR: 7. 8; 95% CI: 1.4 to 41.7), on any given day. Among children with cough only, there was only a weak suggestion of any possible ETS effect. In conclusion, we found that exposure to ETS was associated with a decline in peak flow and increases in respiratory symptoms and use of bronchodilator drugs among asthmatic children. PMID- 10712326 TI - A randomized, prospective evaluation of noninvasive ventilation for acute respiratory failure. AB - We compared noninvasive positive-pressure ventilation (NPPV), using bilevel positive airway pressure, with usual medical care (UMC) in the therapy of patients with acute respiratory failure (ARF) in a prospective, randomized trial. Patients were subgrouped according to the disease leading to ARF (chronic obstructive pulmonary disease [COPD], a non-COPD-related pulmonary process, neuromuscular disease, and status postextubation), and were then randomized to NPPV or UMC. Thirty-two patients were evaluated in the NPPV group and 29 in the UMC group. The rate of endotracheal intubation (ETI) was significantly lower in the NPPV than in the UMC group (6.38 intubations versus 21.25 intubations per 100 ICU days, p = 0.002). Mortality rates in the intensive care unit (ICU) were similar for the two treatment groups (2.39 deaths versus 4.27 deaths per 100 ICU days, p = 0.21, NPPV versus UMC, respectively). Patients with hypoxemic ARF in the NPPV group had a significantly lower ETI rate than those in the UMC group (7.46 intubations versus 22.64 intubations per 100 ICU days, p = 0.026); a similar trend was noted for patients with hypercapnic ARF (5.41 intubations versus 18.52 intubations per 100 ICU days, p = 0.064, NPPV versus UMC, respectively). Patients with ARF in the non-COPD category had a lower rate of ETI with NPPV than with UMC (8.45 intubations versus 30.30 intubations per 100 ICU days, p = 0.01). Although the rate of ETI was lower among COPD patients receiving NPPV, this trend did not reach statistical significance (5.26 intubations versus 15.63 intubations per 100 ICU days, p = 0.12, NPPV versus UMC, respectively). In conclusion, NPPV with bilevel positive airway pressure reduces the rate of ETI in patients with ARF of various etiologies. PMID- 10712327 TI - Lung T-helper cells expressing T-cell receptor AV2S3 associate with clinical features of pulmonary sarcoidosis. AB - In previous reports of studies of Scandinavian sarcoidosis patients, we have described a strong association between lung-restricted expansions of T cells expressing T-cell receptor (TCR) AV2S3 and the human leukocyte antigen (HLA) DRB1*0301 (DR17) and -DRB3*0101 alleles, suggesting the presence of a specific antigen in sarcoidosis. In the present study, the degree of lung-accumulated TCR AV2S3(+) T cells was related to clinical data in 51 HLA-DRB1*0301/DRB3*0101 positive Scandinavian patients with pulmonary sarcoidosis. Significantly more AV2S3(+) lung T cells (median: 30.0% of CD4(+) cells in bronchoalveolar lavage fluid [BALF]) were found accumulated in patients with a short (< 2 yr) than with a long (> 2 yr) (median: 18.6%) disease duration (p = 0.003). A strong positive association was also found between lung-restricted AV2S3(+) T cells and both the CD4(+)-to-CD8(+) cell ratio (p = 7 x 10(-6)) in BALF and with an acute disease onset (p = 0.018). Negative associations were found between both the interval from disease onset to bronchoalveolar lavage (p = 0.0001) and the age of the patient (p = 0.002). Our findings strongly link lung-accumulated AV2S3(+) T cells to the acute inflammatory response in sarcoidosis. Moreover, the association of these cells with a good prognosis indicates that AV2S3(+) T cells may have a protective role against a presumed sarcoidosis antigen. PMID- 10712328 TI - Compensation for increase in respiratory workload during mechanical ventilation. Pressure-support versus proportional-assist ventilation. AB - Variation in respiratory impedance may occur in mechanically ventilated patients. During pressure-targeted ventilatory support, this may lead to patient-ventilator asynchrony. We assessed the hypothesis that during pressure-support ventilation (PSV), preservation of minute ventilation (V E) consequent to added mechanical loads would result in an increase in respiratory rate (RR) due to the large reduction in tidal volume (VT). WITH proportional-assist ventilation (PAV), preservation of V E would occur through the preservation of VT, with a smaller effect on RR. We anticipated that this compensatory strategy would result in greater patient comfort and a reduce work of breathing. An increase in respiratory impedance was obtained by chest and abdominal binding in 10 patients during weaning from mechanical ventilation. V E remained constant in both ventilatory modes after chest and abdominal compression. During PSV, this maintenance of VE was obtained through a 58 +/- 3% increase in RR that compensated for a 29 +/- 2% reduction in VT. The magnitudes of the reduction in VT (10 +/- 3%) and of the increase in RR (14 +/- 2%) were smaller (p < 0. 001) during PAV. During both PSV and PAV, chest and abdominal compression caused increases in both the pressure-time product (PTP) of the diaphragm per minute (142.9 +/- 26.9 cm H(2)O. s/min, PSV, and 117.6 +/- 16.4 cm H(2)O. s/min, PAV) and per liter (13.4 +/- 2.5 cm H(2)O. s/L, PSV, and 9.6 +/- 0.7 cm H(2)O. s/L, PAV). These increments were greater (p < 0.001) during PSV than during PAV. The capability of keeping VT and V E constant through increases in inspiratory effort after increases in mechanical loads is relatively preserved only during PAV. The ventilatory response to an added respiratory load during PSV required greater muscle effort than during PAV. PMID- 10712329 TI - The risk of myocardial infarction associated with inhaled beta-adrenoceptor agonists. AB - Beta-adrenoceptor agonists (beta-agonists), in widespread clinical use for obstructive lung disease, have been associated with an increased risk of cardiovascular mortality. The objective of this study was to assess the association between incident myocardial infarction and the use of inhaled beta agonists. We performed a case-control study within the Group Health Cooperative of Puget Sound (GHC). Between 1989 and 1994, we identified 1,444 cases with an incident myocardial infarction and 4,094 control subjects frequency-matched on age, sex, hypertension, and index date. The computerized pharmacy database of the health maintenance organization (HMO) was used to assess the use of beta-agonists by metered dose inhaler (MDI). Cardiovascular risk factor information was obtained from medical record review. In comparison to subjects who did not fill a beta-agonist prescription, subjects who had filled one beta-agonist MDI prescription in the 3 mo prior to their index date had an elevated estimated risk of myocardial infarction (adjusted odds ratio [OR]: 1.67 [95% CI, 1.07 to 2.60]). The elevated risk was limited to those subjects who had a history of cardiovascular disease (adjusted OR: 3.22 [95% CI, 1.63 to 6.35]) and among those with cardiovascular disease, to new users of beta-agonists (adjusted OR: 7.32 [95% CI, 2.34 to 22.8]). There was no dose-response relationship between beta agonists use and risk of myocardial infarction. In this study, new use of beta agonists was associated with an increased risk of myocardial infarction, although we cannot determine if the association is causal. Our study suggests that clinicians should exercise caution when giving an initial beta-adrenoceptor agonist prescription to patients with cardiovascular disease. PMID- 10712330 TI - Pivotal role of anionic phospholipids in determining dynamic behavior of lung surfactant. AB - Phosphatidylglycerol (PG) and phosphatidylinositol (PI) are anionic phospholipids (APLs) present in lung surfactant of virtually all species studied, although their specific contribution to function is unknown. This study examines how APLs influence surfactant monolayer stability and adsorption under static and dynamic conditions. Interfacial properties of surfactants reconstituted with native phospholipids (PL), and phospholipids devoid of anionic species (DAPL), were characterized by pulsating bubble surfactometry. Measurements were made for PL and DAPL alone; with 3% surfactant proteins B and C (SP-B/C); with SP-B/C and 5% surfactant protein A (SP-A); and with SP-B/C, SP-A, and 8% neutral lipids (NL). Equilibrium and dynamic properties of PL and DAPL were similar. However, whereas (DAPL + SP-B/C) and (DAPL + SP-B/C + SP-A) mixtures were similar to corresponding PL mixtures with respect to gamma(equil), they displayed markedly different dynamic behavior. In particular, the degree of film compression required to reach gamma(min) was significantly increased in DAPL mixtures (80 to 90% area reduction) compared with PL, although both samples reached gamma(min) < 3.0 dynes/cm. The addition of NL to (DAPL + SP-B/C + SP-A) produced an increase in gamma(min) to 15 to 20 dynes/cm during dynamic compression, whereas NL had no significant impact on the behavior of (PL + SP-B/C + SP-A). Purified PG (5% wt/wt) restored nearly normal dynamic properties to (DAPL + SP-B/C + SP-A + NL), whereas phosphatidylcholine (PC) (5% wt/wt) had no beneficial effect. These results suggest that APLs play a critical role in promoting surface film stability during dynamic compression through interactions with nonlipid surfactant components, and prevent destabilization of the surface film by cholesterol and other NL. PMID- 10712331 TI - Bronchial responsiveness among inbred mouse strains. Role of airway smooth-muscle shortening velocity. AB - To investigate the relationship between bronchial responsiveness and airway smooth-muscle (ASM) contractile properties, we studied inbred mice with known interstrain differences in airway responsiveness. Using oscillatory mechanics, we confirmed that A/J mice were hyperresponsive to methacholine (MCh) as compared with mice of the C3H/HeJ and C57BL/6J strains. Analysis of respiratory system resistance and elastance at different flow oscillation frequencies indicated that interstrain differences in responsiveness are present in both central and peripheral airways of these mice. We used video microscopy to measure the rate of contraction of explanted airways, and found that the airways of A/J mice contracted more rapidly than those of C3H/HeJ or C57BL/6J mice. In studies of a fourth strain (Balb/C) of mice, we found both bronchial hyperresponsiveness and increased ASM shortening velocity. The rank order of responsiveness among strains was the same as that for shortening velocity (A/J > Balb/C > C3H/HeJ > C57BL/6J). Furthermore, in each strain of mice, shortening velocity correlated with the achieved degree of airway narrowing and with a greater likelihood of airway closure in individual airways. In contrast, generation of isometric tension in trachealis, morphometric measurements of tracheal ASM, tracheal myosin content, and dose-response curves for MCh of explanted intraparenchymal bronchi failed to correspond to the in vivo phenotype of airway reactivity. These results indicate that bronchial responsiveness is related to ASM shortening velocity, and underscore the importance of smooth-muscle dynamics in understanding the mechanisms of bronchial responsiveness. PMID- 10712332 TI - Sleep disorders and diaphragmatic function in patients with amyotrophic lateral sclerosis. AB - In amyotrophic lateral sclerosis (ALS), the progressive loss of upper and lower motor neurons leads to respiratory failure, often with predominant diaphragm dysfunction, and death. Because the diaphragm is the only active inspiratory muscle during rapid eye movement (REM) sleep, there is a high theoretical risk of respiratory disorders during REM sleep in patients with ALS. To assess this hypothesis, we studied sleep characteristics (polysomnography) in 21 patients with ALS, stratified according to the presence or absence of diaphragmatic dysfunction. Diaphragmatic dysfunction was defined as an absent or delayed diaphragm response to cervical or cortical magnetic stimulation, abdominal paradox, or respiratory pulse (Group 1, 13 patients). These patients did not differ in age, clinical course, or form (bulbar or spinal) from the eight others, who did not have diaphragmatic dysfunction (Group 2). REM sleep was reduced in Group 1 (7 +/- 7% of total sleep time; mean +/- SD) and normal in Group 2 (18 +/- 6%, p = 0.004). Apneas or hypopneas were rare in both groups. In Group 1, REM sleep was absent or minimal (less than 3 min) in five patients. An unusual and remarkable preservation of phasic inspiratory sternomastoid activation during REM was associated with longer REM sleep duration in six of the other patients with diaphragmatic dysfunction. Median survival time was dramatically shorter (217 d) in Group 1 than in Group 2 (619 d, p = 0.015). PMID- 10712333 TI - Treatment with nasal CPAP decreases automobile accidents in patients with sleep apnea. AB - We studied 50 consecutive patients to test the hypothesis that successful treatment of obstructive sleep apnea with nasal continuous positive airway pressure (nasal CPAP) will decrease automobile accidents in patients with sleep apnea. Thirty-six (72%) of the patients reported using nasal CPAP regularly during 2 yr. Fourteen patients reported they had not used CPAP during 2 yr. The patients with sleep apnea in this study had a higher automobile crash rate than all drivers in the state of Colorado (0.07 versus 0. 01 crash per driver per year, p < 0.02). Patients who were treated with nasal CPAP had a lower crash rate while being treated than before treatment (0.07 versus 0 crash per driver per year, p < 0.03). Untreated patients with sleep apnea continued to have a high crash rate (0.07 crash per driver before and after diagnosis). Drivers with sleep apnea were reluctant to report their automobile crashes, for the drivers in this study reported only one-third of the crashes in which they were involved. This is the first study to confirm with traffic records that patients with sleep apnea have fewer automobile crashes while being treated with nasal CPAP. PMID- 10712334 TI - A pilot study of the effect of gentamicin on nasal potential difference measurements in cystic fibrosis patients carrying stop mutations. AB - Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene containing a premature termination signal are expected to produce little or no CFTR chloride channels. It has been shown in vitro, that aminoglycoside antibiotics can increase the frequency of erroneous insertion of nonsense codons hence permitting the translation of CFTR alleles carrying missense mutations to continue reading to the end of the gene. This led to the appearance of functional CFTR channels at the apical plasma membrane. The aim of this research was to determine if topical application of gentamicin to the nasal epithelium of patients with cystic fibrosis (CF) carrying stop mutations can express, in vivo, functional CFTR channels. Nine CF patients carrying stop mutations (mean age 23 +/- 11 yr, range 12 to 46 yr) received gentamicin drops (0.3%, 3 mg/ml) three times daily intranasally for a total of 14 d. Nasal potential difference (PD) was measured before and after the treatment. Before gentamicin application all the patients had abnormal nasal PD typical of CF. After gentamicin treatment, significant repolarization of the nasal epithelium representing chloride transport was increased from -1 +/- 1 mV to -10 +/- 11 mV (p < 0. 001). In conclusion, gentamicin may influence the underlying chloride transport abnormality in patients with CF carrying stop mutations. PMID- 10712335 TI - Predictors of improvements in daytime function outcomes with CPAP therapy. AB - Continuous positive airway pressure (CPAP) therapy improves daytime function in the sleep apnea/ hypopnea syndrome (SAHS) but it is unclear which patients benefit and what factors predict this improvement. To test the hypothesis that brief arousals from sleep predict improvements in daytime functioning with CPAP therapy, we prospectively studied 62 patients with polysomnography-defined SAHS. Each underwent daytime function assessments at baseline and after 6 mo of CPAP therapy to measure objective sleepiness, psychological well-being, quality of life, and cognitive performance. The microarousal frequency and AHI were poor predictors of improvements in daytime function with CPAP. Measures of hypoxemia predicted improvements in the mean sleep latency on the maintenance of wakefulness test, SAHS symptoms, quality of life, and reaction time, but such correlations were weak or moderate only explaining between 7% and 22% of variance. Significant relationships were found between CPAP use and improvements in self-ratings of daytime function. Results suggest that standard polysomnographic baseline variables are poor predictors of the response to CPAP therapy. PMID- 10712336 TI - Initial versus delayed acute renal failure in the intensive care unit. A multicenter prospective epidemiological study. Rhone-Alpes Area Study Group on Acute Renal Failure. AB - We performed a prospective study in the 28 multidisciplinary intensive care units (ICUs) in the Rhone-Alpes area in France to investigate the role of initial versus delayed occurrence of acute renal failure (ARF) in patient outcome. ARF was defined as a serum creatinine concentration > 300 micromol/L, urine output < 500 ml/24 h (or < 180 ml/8 h), or hemodialysis requirement. Over the 1-yr study period, 1,086 patients presented with ARF on ICU admission or during the first 2 d of ICU stay (Group A; 736 patients), from Day 3 to Day 6 (Group B; 202 patients), or from Day 7 (Group C; 148 patients). The overall hospital mortality rate was 66% (61% in Group A, 71% in Group B, and 81% in Group C; p < 0.0001). Logistic regression analysis of a random sample of 510 patients showed that SAPS II score on ICU admission, number of ARF episodes, previous health status, absence of oliguria, absence of hemodialysis, and absence of ischemic acute tubular necrosis were predictive of patient survival. This model was tested and validated on the basis of the remaining patients. Thus, in this population, late ARF was not a predictive factor for patient outcome. PMID- 10712337 TI - Respiratory muscle recruitment and exercise performance in eucapnic and hypercapnic severe chronic obstructive pulmonary disease. AB - If chronic hypercapnia in patients with severe COPD occurs as a consequence of respiratory muscle (RM) weakness or fatigue, we would expect that ventilatory muscle recruitment (VMR) and exercise performance in stable hypercapnic patients would differ from those in eucapnic patients. We evaluated exercise performance and RM function at rest and during exercise in 19 eucapnic (PCO(2) 40 +/- 3 mm Hg), and 13 hypercapnic (PCO(2) 52 +/- 10 mm Hg) patients with severe COPD. A metabolic cart was used to determine V E, V O(2), V CO(2), and HR. Gastric (Pg) and esophageal (Ppl) balloons were used to measure Pg, Ppl, and Pdi. Ventilatory muscle recruitment pattern (VMR) was partitioned using end-inspiratory and end expiratory Pg and Ppl. Hypercapnic patients had lower FEV(1) (0.60 +/- 0.24 versus 0.95 +/- 0.31 L, p < 0.001), MVV (28 +/- 11 versus 41 +/- 13 L, p < 0.001), resting PO(2) (61 +/- 11 versus 70 +/- 11 mm Hg, p < 0.001), peak PO(2) (60 +/- 20 versus 75 +/- 22 mm Hg, p < 0.005), and V E(max) (24 +/- 10 versus 32 +/- 12 L/min, p < 0.001). Patients in both groups had similar FRC (5.7 +/- 1.6 versus 5.0 +/- 1.5 L), V O(2)max (0.58 +/- 0.30 versus 0.76 +/- 0.32 L/min), Watts (45 +/- 48 versus 71 +/- 59), V E/MVV (88 +/- 33 versus 79 +/- 14), and HRmax (117 +/- 17 versus 128 +/- 18 beats/min). PI(max) (67 +/- 28 versus 65 +/- 32 cm H(2)O) and PE(max) (98 +/- 34 versus 96 +/- 40 cm H(2)O) were also similar in both groups. VMR (DeltaPg/DeltaPpl) at rest (-0.28 +/- 0.51 versus 0 +/- 0.35) and during exercise (0.4 +/- 0.2 versus 0.39 +/- 0.15) was equally affected in both groups. We conclude that exercise capacity and ventilatory muscle recruitment are similarly impaired in eucapnic and hypercapnic patients with severe COPD. These findings make inability of the lung to increase ventilation and not respiratory muscle dysfunction a more attractive explanation for CO(2) retention in stable hypercapnic patients. PMID- 10712338 TI - Influence of body weight on the severity of dyspnea in chronic obstructive pulmonary disease. AB - A substantial number of patients with COPD are underweight (UW); they comprise the clinical subtype of "dyspneic" or emphysematous. To determine whether these patients are more dyspneic than normal weight (NW) patients with COPD, we quantitated the severity of dyspnea, using a modified Medical Research Council (MRC) dyspnea scale, in 33 UW and 57 NW patients and compared their pulmonary function tests (PFTs), arterial blood gases (ABGs), and respiratory muscle strength as estimated by maximum static inspiratory (PI(max)) and expiratory (PE(max)) mouth pressures (all as means +/- SEM). Body mass index was 18.7 +/- 1.2 and 24.5 +/- 1.8 kg/m(2) in UW and NW patients, respectively (p < 0.0001). The MRC dyspnea scale was 3. 1 +/- 0.9 in UW and 2.5 +/- 1.2 in NW groups (p = 0.035). All PFT and ABG parameters were similar in the two groups except for DCO (36 +/- 11% in UW and 57 +/- 17% in NW, p < 0.001) and PI(max) (55 +/- 18 mm Hg in UW and 66 +/- 19 mm Hg in NW, p = 0.020). In a stepwise multiple regression model, %DCO and %MVV combined were the best predictors of dyspnea severity (R(2) = 0.30, p = 0.001). We conclude that UW patients with COPD are more dyspneic than NW patients. Although the origin of dyspnea in COPD is multifactorial, changes in DCO and respiratory muscle strength may contribute to its intensity. PMID- 10712339 TI - Modulation of release of reactive oxygen species by the contracting diaphragm. AB - Recent reports have demonstrated that superoxide is released by the contracting diaphragm. Moreover, extracellular scavengers of superoxide (i.e., exogenously administered superoxide dismutase) reduce diaphragm fatigue rate, arguing that superoxide released from contracting muscles may have functionally significant effects. The mechanism by which free radical formation and release occurs has not, however, been determined, and all past studies of this phenomenon have been conducted at a single muscle length (the length of maximum force generation, Lo) and at a single level of carbon dioxide. The purpose of the present study was twofold: (1) to examine the effect of blockade of two free radical-generating pathways (i.e., to block cyclooxygenase with indomethacin and xanthine oxidase with oxypurinol) on superoxide release by the contracting diaphragm, and (2) to examine the effect of altering muscle length, carbon dioxide levels, and stimulation frequency on superoxide release during contraction. Studies were performed using an isolated, arterially perfused, rat diaphragm preparation in which superoxide release was assessed in real time by measuring arteriovenous cytochrome c reduction gradients across this muscle. We found that superoxide release during contraction was: (1) not altered by indomethacin administration, (2) partially reduced by oxypurinol administration, (3) reduced by decreasing muscle length, (4) reduced by increasing carbon dioxide concentrations, and (5) reduced by decreasing stimulation frequency. The first two findings indicate that xanthine oxidase pathways contribute to free radical formation under these circumstances but cyclooxygenase does not. The last three findings suggest that these common physiologic alterations have significant effects on free radical release by contracting muscle. PMID- 10712340 TI - Smooth reference equations for slow vital capacity and flow-volume curve indexes. AB - We derived reference values for slow vital capacity (VC) and flow-volume curve indexes (FVC, FEV(1), and flows) from the 1,185 tracings provided by 1,039 "normal" subjects who participated in one or both cross-sectional surveys of the Po River Delta study in 1980-1982 and in 1988-1991. Definition of "normal" was based on negative answers to questions on respiratory symptoms/diseases or recent infections, current/past tobacco smoking, and work exposure to noxious agents. Reference equations were derived separately by sex as linear regressions of body mass index (BMI = weight/height(2)), BMI-squared, height, height-squared, and age. Age entered all the models by natural cubic splines using two break points, except for the ratios FEV(1)/VC and FEV(1)/FVC. Random effects models were applied to adjust for the potential intrasubject correlation. BMI, along with height and age, appeared to be an important predictor, which was significantly associated with VC, FEV(1), FVC, FEV(1)/FVC, and PEF in both sexes, and with FEV(1)/VC and FEF(25-75) in females. Natural cubic splines provided smooth reference equation curves (no "jumps" or "angled points") over the entire age span, differently from the conventional reference equations. Thus, we recommend the use of smooth continuous equations for predicting lung function indexes, along with the inclusion of BMI in the equations. PMID- 10712341 TI - A common FCER1B gene promoter polymorphism influences total serum IgE levels in a Japanese population. AB - Genetic factors are important in defining total serum IgE levels. Linkage analyses have localized a gene or genes that influence atopic phenotype at chromosome 11q13. Variants of the FCER1B gene have been identified, which are associated with an increased risk of developing atopy and bronchial asthma. Given uncertain functional consequences and low frequencies of these coding variants of FCER1B, we screened for new mutations using 24 subjects with atopic asthma. A common -109C/T polymorphism at the promoter region of FCER1B was identified, although no variant was found in the entire coding region. We genotyped this promoter polymorphism in 226 healthy control subjects and 226 asthmatic subjects using polymerase chain reaction-restriction fragment length polymorphism (PCR RFLP) analysis. Allele frequencies were 0.697 for -109T and 0.303 for -109C in 226 healthy control subjects. No significant difference in the distribution of 109C/T polymorphism was found between asthmatic subjects and healthy control subjects. A homozygosity for the -109T allele, however, was associated with increased total serum IgE levels in 226 subjects with asthma (p = 0.0015). The strongest evidence for an association between total serum IgE levels and -109C/T polymorphism (p = 0.0004) was obtained when age at onset of asthma was incorporated into the analysis. Our findings may represent genetic heterogeneity and complex interactions between genetic and environmental components involved in the regulation of total IgE levels, providing evidence that the -109C/T polymorphism of the FCER1B promoter region is one of the genetic factors identified thus far, which affects total serum IgE levels in a Japanese population. PMID- 10712342 TI - Peripheral airway smooth muscle mechanics in obstructive airways disease. AB - The purpose of this study was to determine whether altered airway smooth muscle (ASM) contractility contributes to the pathogenesis of obstructive airways diseases such as chronic obstructive pulmonary disease (COPD) and asthma. The passive and active mechanical properties of isolated human peripheral airways were measured in vitro by myography. The amount of ASM was measured by morphometry. Pulmonary function was assessed before surgery by the FEV(1) (%pred) and the FEV(1)/ FVC (%). Fifteen airways were studied from nonobstructed (NOB) patients, and 15 from obstructed (OB, FEV(1)/FVC < 70%) patients (62 +/- 10 yr, mean +/- SD). The maximal isometric force (Fmax), stress (Fmax/ASM), airway diameter at Lmax (Dmax), maximal isotonic shortening (%Lmax), and normalized airway smooth muscle (ASM/Dmax) were determined in all patients. There was a significant correlation between Fmax and FEV(1) (%pred) (r = -0.579, p < 0.004), between Fmax and FEV(1)/FVC (%) (r = -0.720, p < 0.003), and between stress and FEV(1)/FVC (%) (-0.611, p < 0.002). There was no correlation between isotonic shortening and either measure of pulmonary function. A positive correlation was found between force and shortening (r = 0.442, p < 0.05), and stress and shortening (r = 0.538, p < 0.01). Both force and stress were significantly increased (p < 0.05) in OB (Fmax = 0.87 +/- 0.8 g, stress = 76 +/- 47 mN/mm(2)) versus NOB (Fmax = 0.42 +/- 0.18 g, stress = 51 +/- 21 mN/mm(2)) patients, while isotonic shortening was not different between the two groups. ASM and ASM/Dmax were both significantly increased in the OB patient group (p < 0.05). These results suggest that obstructive airways disease is associated with an increase in the ability of the ASM to generate force. (Values represent means +/- SD.) PMID- 10712344 TI - PAF-induced RANTES production by human airway smooth muscle cells requires both p38 MAP kinase and Erk. AB - Airway smooth muscle (ASM) cells, which have been regarded as having contractile properties in response to contractile inflammatory mediators, may also participate in airway inflammatory response by expressing various cytokines, including RANTES. However, the intracellular signal that regulates cytokine expression in ASM cells has not been determined. In the present study, we examined the role of p38 mitogen-activated protein (MAP) kinase and extracellular signal-regulated kinase (Erk) in RANTES production by ASM cells stimulated by platelet-activating factor (PAF) and tumor necrosis factor (TNF)-alpha. The results showed that PAF induced the threonine and tyrosine phosphorylation of p38 MAP kinase and Erk, and p38 MAP kinase and Erk activity. SB 203580 and PD 98059 almost completely inhibited p38 MAP kinase and Erk activity, respectively. SB 203580 and PD 98059 partially inhibited and acted additively to inhibit PAF induced RANTES production. PAF also induced c-Jun-NH(2)-terminal kinase ( JNK) phosphorylation. TNF-alpha induced p38 MAP kinase and Erk phosphorylation, but neither SB 203580 nor PD 98059 inhibited RANTES production. These results indicate that both p38 MAP kinase and Erk involve RANTES production by ASM cells stimulated with PAF, but not TNF-alpha, and that the role of p38 MAP kinase and Erk in RANTES production by ASM cells appears to be stimulus-dependent. PMID- 10712343 TI - Transient effect of inhaled fluticasone on airway mucosal blood flow in subjects with and without asthma. AB - Topically applied glucocorticosteroids (GS) have been shown to cause local vasoconstriction in normal skin and this phenomenon is commonly used to assess the potency of topical GS (McKenzie skin blanching test). The purpose of the present study was to determine if an inhaled GS, fluticasone propionate (FP), similarly leads to vasoconstriction in the airway mucosa and if subjects with and without asthma have differential vascular responsiveness to GS. In 10 nonsmokers with stable asthma and 10 nonasthmatic nonsmokers, airway mucosal blood flow (Qaw) expressed per milliliter of anatomical dead space and the forced expiratory volume in 1 s (FEV (1)) were determined before and serially after inhalation of FP (88 to 1,760 microg) or placebo. Baseline mean (+/- SE) Qaw was 55.1 +/- 1.0 and 44.2 +/- 1.1 microl x min(-1) x ml(-1) in subjects with and without asthma, respectively (p < 0.001). The corresponding mean FEV(1) values were 2.34 +/- 0.13 and 3.22 +/- 0.12 L (p < 0.001). FP at 880 microg but not placebo produced a transient decrease in mean Qaw with a nadir at 30 min and return toward baseline at 90 min post-inhalation; the maximum mean decrease was 37% in subjects with asthma and 21% in unaffected subjects (p < 0.01); 880 microg of FP was the lowest effective dose. FEV(1) did not change after FP administration in either group. These results demonstrate a transient vasoconstrictive action of inhaled FP in the airway mucosa, with a greater vascular responsiveness in subjects with asthma than in unaffected subjects. The measurement of Qaw may provide a more relevant means of assessing the potency of inhaled GS than the McKenzie skin blanching test. In addition, our observation suggests that inhaled GS have potentially beneficial effects in asthma that is not related to their antiinflammatory action. PMID- 10712345 TI - Sensitive identification of mycobacterial species using PCR-RFLP on bronchial washings. AB - In 98 patients (24 with active pulmonary tuberculosis [TB] lesions, 28 with cured TB lesions, and 46 with nontuberculous opacities [control group] in chest CT scans), we examined whether washing the bronchus after brushing the lesion, then applying polymerase chain reaction-restriction fragment length polymorphism (PCR RFLP) to the bronchial washings might be useful for diagnosing TB and nontuberculous mycobacteriosis (NTMosis). After biopsy and brushing with a bronchoscope, the bronchus connecting to the lesion was washed with 20 ml saline. The saline used for washing the brushes (5 ml; brushing sample), and 3 to 10 ml saline aspirated through the forceps channel (washing sample) were examined by PCR-RFLP, which proved able to identify Mycobacterium tuberculosis and seven species of nontuberculous mycobacteria (NTM). The values obtained for the sensitivity of the PCR-RFLP with respect to the brushing sample, the washing sample, and both samples mixed together were 70, 76, and 91%, respectively, when only patients who were culture-positive or radiologically improved after antituberculous therapy were considered as showing true infection. A mixture of brushing and washing samples provides useful material for PCR and culture, and the PCR-RFLP used here is a good method for the simultaneous identification of several species of mycobacterium (including M. tuberculosis). PMID- 10712346 TI - Cytoprotective effects of nitroglycerin in ischemia-reperfusion-induced lung injury. AB - Prevention of ischemia-reperfusion (IR) injury is crucial for successful lung transplantation. We investigated whether a nitric oxide donor, nitroglycerin (NTG), could suppress the oxidative stress of IR injury and improve pulmonary function after reperfusion in an ex vivo rat lung perfusion model. In Fresh group of animals, the lungs were flushed with perfusate, followed immediately by reperfusion, and no lung injury was observed. In NTG- and NTG+ groups of animals, the lungs were flushed with perfusate alone or perfusate containing NTG, respectively. Harvested lung and heart blocks from these latter two groups were immersed in the corresponding perfusate at 4 degrees C for 15 h, and were then reperfused for 60 min. Reperfusion induced pulmonary edema in the NTG- group, but not in the NTG+ group. Shunt fractions in NTG+ group were significantly lower than in the NTG- group throughout reperfusion. NTG had no effect on pulmonary arterial pressure or myeloperoxidase activity. In contrast, oxidative DNA damage assessed immunohistochemically with a monoclonal antibody against 8-hydroxy-2' deoxyguanosine (8-OHdG) was significantly increased in the NTG- group, in the order alveolar epithelium > pulmonary endothelium > bronchial epithelium. NTG treatment significantly decreased staining with the anti-8-OHdG antibody in all three areas of tissue. Therefore, administration of NTG attenuates the oxidative stress of IR injury, and may improve pulmonary function after reperfusion. PMID- 10712347 TI - Transport of bifunctional proteins across respiratory epithelial cells via the polymeric immunoglobulin receptor. AB - Neutrophil elastase (NE) contributes to progression of the lung disease characteristic of cystic fibrosis (CF). We developed a strategy that permits the delivery of alpha(1)-antitrypsin (alpha(1)-AT) to inaccessible CF airways by targeting the respiratory epithelium via the polymeric immunoglobulin receptor (pIgR). A fusion protein consisting of a single-chain Fv directed against human secretory component (SC) and linked to human alpha(1)-AT was effectively transported in a basolateral-to-apical direction across in vitro model systems of polarized respiratory epithelium consisting of 16HBEo cells transfected with human pIgR complementary DNA, which overexpress the receptor, and human respiratory epithelial cells grown in primary culture at an air-liquid interface. When applied to the basolateral surface, the anti-SC Fv/alpha(1)-AT fusion protein penetrated the respiratory epithelia, with transcytosis of the fusion protein being related to the amount of SC detected at the apical surface. Significantly less fusion protein crossed the cells in the opposite direction. In addition, because the antihuman SC Fv/alpha(1)-AT fusion protein was transported vectorially and deposited into the small volume of apical surface fluid, the antiprotease component of this protein was concentrated atop the epithelium. Thus, in cell models, this system is capable of concentrating the antiprotease of the fusion protein, in the thin film of epithelial surface fluid to a level expected to be therapeutic in the airways of many patients with CF. PMID- 10712349 TI - Cryptococcus albidus-induced summer-type hypersensitivity pneumonitis. AB - We studied summer-type hypersensitivity pneumonitis believed to be induced by Cryptococcus albidus in the home environments of the patients. All patients had antibodies that were reactive to Cryptococcus neoformans and Trichosporon cutaneum in sera and bronchoalveolar lavage (BAL) fluids. Cryptococcus albidus strains were isolated from 62.5% of the patient home environments. Trichosporon cutaneum was found in none of the patient homes. To study local antibody production in the lung, we cultured BAL cells to measure anti-C. neoformans and anti-T. cutaneum antibodies in the culture supernatants by the ELISA method. IgG, IgA, and IgM anti-Cryptococcus and anti-Trichosporon antibodies were found in all culture supernatants. A significant correlation was observed in antibody binding activity between Cryptococcus and Trichosporon antigen. However, the amount of IgA and IgM antibody bound to C. neoformans was significantly higher than was bound to T. cutaneum. Most anti-Cryptococcus and anti-Trichosporon antibody was absorbed by C. albidus. Our results suggest that C. albidus may be an etiologic agent in most of the cases we studied, and that IgA and IgM antibody in BAL fluid may be secreted by plasma cells in the lung. PMID- 10712348 TI - CD23 exhibits negative regulatory effects on allergic sensitization and airway hyperresponsiveness. AB - The effects of an anti-CD23 monoclonal antibody (B3B4) in CD23-deficient and CD23 overexpressing mice were compared in a murine model of allergic sensitization. After sensitization and challenge with OA, mice developed increased serum levels of OA-specific IgE and IgG(1) with airway eosinophilia and AHR when compared with nonsensitized animals. Anti-CD23 treatment was studied under two protocols: 10-d OA aerosol exposure and intraperitoneal sensitization followed by aerosol challenge. In both protocols anti-CD23 significantly reduced IgE and IgG(1) levels, abolished eosinophilia, and normalized AHR in BALB/c and wild-type CD23+/+ mice but not in CD23-/- mice. These changes were associated with increases in IFN-gamma and decreases in IL-4 production, suggesting that CD23 binding may affect not only IgE production but also the Th1/Th2 imbalance during the development of allergic AHR. Absence of CD23 in gene-deficient mice significantly enhanced OA-specific IgE and IgG(1) levels, airway eosinophilia, and AHR when compared with CD23+/+ wild-type littermates after sensitization and airway challenge. Sensitized and challenged CD23 transgenic mice also developed eosinophilic airway inflammation and methacholine hyperresponsiveness. However, the extent of AHR, BAL, and tissue eosinophilia in these animals showed a significant negative correlation with levels of CD23 expression on splenic T and B cells, demonstrating a limiting role of CD23 in the development of allergic AHR. PMID- 10712350 TI - Tenascin mRNA expression at the foci of recent injury in usual interstitial pneumonia. AB - To elucidate which cells are synthesizing tenascin in usual interstitial pneumonia (UIP) we have analyzed thoracoscopic or open lung biopsies from 30 patients with UIP by mRNA in situ hybridization, using (35)S-labeled tenascin RNA probes. The phenotype of the cells expressing tenascin mRNA was confirmed by immunohistochemical stainings of serial sections with antibodies against alpha smooth muscle actin and human cytokeratin. The results demonstrate that tenascin is expressed at the foci of recent lesions consisting of intralumenal or incorporating loose fibrotic buds. The cells expressing tenascin mRNA were located in and underneath the newly formed epithelium. Immunohistochemical stainings showed that the cells in the newly formed epithelium were strongly cytokeratin positive, and thus evidently regenerating type 2 pneumocytes, while the cells underneath the newly formed epithelium were alpha-smooth muscle actin positive and apparently myofibroblasts. Tenascin mRNA expression was clearly stronger and more frequent in myofibroblasts than in type 2 pneumocytes, however. Weak tenascin mRNA expression was also found in metaplastic bronchiolar-type epithelium and alveolar macrophages. Our results are thus in good agreement with the previous studies showing that tenascin is actively synthesized at the early fibrotic lesions in UIP. Furthermore, results demonstrate that the interaction between the epithelium and the underlying connective tissue plays a significant role in tenascin synthesis and that myofibroblasts are mainly responsible for its synthesis in fibroblastic foci of UIP. PMID- 10712351 TI - Allelic heterogeneity in hereditary surfactant protein B (SP-B) deficiency. AB - Inability to produce surfactant protein B (SP-B) causes fatal neonatal respiratory disease. A frame-shift mutation (121ins2) is the predominant but not exclusive cause of disease. To determine the range of mechanisms responsible for SP-B deficiency, both alleles from 32 affected infants were characterized. Sixteen infants were homozygous for the 121ins2 mutation, 10 infants were heterozygous for the 121ins2 and another mutation, and six infants were homozygous for other mutations. Thirteen novel SP-B gene mutations were identified, which were not found in a control population. One novel mutation was found in two unrelated families. Surfactant protein expression was evaluated by immunohistochemistry and/or protein blotting. Absence of proSP-B and mature SP-B was associated with nonsense and frame-shift mutations. In contrast, proSP-B expression was associated with missense mutations, or mutations causing in-frame deletions or insertions, and low levels of mature SP-B expression were associated with four mutations. Extracellular staining for proSP-C and/or aberrantly processed SP-C was observed in lungs of all infants with SP-B gene mutations. Hereditary SP-B deficiency is caused by a variety of distinct mutations in the SP B gene and may be associated with reduced, as well as absent, levels of mature SP B, likely caused by impaired processing of proSP-B. PMID- 10712352 TI - Endothelin receptor blockade attenuates lipopolysaccharide-induced pulmonary nitric oxide production. AB - Increased nitric oxide (NO) synthesis by the inducible nitric oxide synthase (iNOS) has been shown to contribute to the development of acute lung injury and delayed hypotension in animals injected with bacterial lipopolysaccharides (LPS). Recent evidence indicates that endothelin-1 (ET-1) is also elevated in septic humans and in animals. To assess the contribution of ETs to LPS-induced pulmonary NO production and iNOS expression, we used P1/fl, a 22 amino acid peptide, to selectively antagonize endothelin-A receptors. Anesthetized, mechanically ventilated rats were injected with either saline or LPS (E. coli endotoxin, 20 mg/kg) and studied for 5 h. Two other groups of rats were pretreated 15 min earlier with P1/fl peptide (20 microg/kg). Unlike saline-treated rats, rats injected with LPS showed a progressive decline in arterial pressure and a significant rise in plasma ET concentration and serum nitrite-nitrate level. In the lungs, LPS injection elicited a several-fold rise in lung iNOS activity and exhaled NO concentration and increased lung wet/dry ratio significantly. Pretreatment with P1/fl peptide eliminated the decline in arterial pressure, the rise in lung wet/dry ratio, lung NOS activity, and iNOS protein expression and significantly attenuated the increase in pulmonary exhaled NO production but had no effect on plasma ET concentration. We conclude that activation of ET-A receptors by rising ET-1 concentration enhances NO production and iNOS expression in the respiratory and vascular systems and contributes to both LPS-induced hypotension and acute lung injury. PMID- 10712353 TI - A novel alveolar type I cell-specific biochemical marker of human acute lung injury. AB - Currently there is no recognized biochemical or molecular marker for human parenchymal lung injury analogous to markers for acute myocardial injury. Injury to the alveolar epithelial barrier is of central importance in the pathogenesis of and recovery from acute lung injury. In animal models, an alveolar type I cell specific protein, RTI(40), has been shown to be an accurate marker of alveolar epithelial damage. We now report that HTI(56), a novel apical plasma membrane protein specific to the human type I cell, is a biochemical marker for lung injury. Using a sensitive, quantitative, light-based ELISA, we measured HTI(56) in pulmonary edema fluid from 15 patients with a clinical diagnosis of acute lung injury and 12 control patients with hydrostatic (cardiogenic) pulmonary edema. HTI(56) was also measured in plasma from these two groups and from 11 normal volunteers. The amount of HTI(56) was 4. 3-fold higher (p < 0.0001) in alveolar edema fluid and 1.4-fold higher (p < 0.05) in plasma from the patients with acute lung injury, compared with patients with hydrostatic pulmonary edema. To our knowledge, this study is the first to utilize a specific marker of alveolar epithelial damage in human disease and demonstrates the feasibility of using a blood test to detect lung parenchymal damage. PMID- 10712354 TI - A long-term study of the antiinflammatory effect of low-dose budesonide plus formoterol versus high-dose budesonide in asthma. AB - Adding inhaled long-acting beta(2)-agonists to a low dose of inhaled corticosteroids (ICS), results in better clinical asthma control than increasing the dose of ICS. However, this approach may mask underlying airway inflammation. In a double-blind parallel-group study, we evaluated the effect of adding formoterol to a low dose of budesonide, compared with a higher dose of budesonide, on the composition of induced sputum. After a 4-wk run-in period of treatment with budesonide (800 microg, twice daily), 60 patients with moderate asthma were randomly assigned to a 1-yr treatment with 400 microg of budesonide plus placebo, twice daily (BUD800), or 100 microg of budesonide plus 12 microg of formoterol, twice daily (BUD200+F). All drugs were administered via Turbuhaler. Budesonide (800 microg, twice daily) during run-in significantly reduced median sputum eosinophils from 4.5 to 0.68%. No significant changes in the proportion of eosinophils, EG2(+) cells, other inflammatory cells, or ECP levels in sputum were observed over the ensuing 1-yr treatment with BUD200+F or BUD800. Clinical asthma control was not significantly different between both groups. In conclusion, no significant differences in sputum markers of airway inflammation were observed during a 1-yr treatment with a low dose of inhaled budesonide plus formoterol compared with a higher dose of budesonide. PMID- 10712355 TI - Downregulation of estrogen and progesterone receptors in the abnormal smooth muscle cells in pulmonary lymphangioleiomyomatosis following therapy. An immunohistochemical study. AB - Immunohistochemical and confocal microscopic studies were made on lung tissue from 10 women with lymphangioleiomyomatosis (LAM) to evaluate the distribution of estrogen receptors (ER) and progesterone receptors (PR) in the abnormal smooth muscle cells (LAM cells) that characterize this disorder. PR and ER were localized mainly in the nuclei of large, epithelioid LAM cells, in five patients in whom tissues were obtained before treatment. However, the reaction for PR and ER was essentially negative in similarly processed tissues from five patients studied after receiving hormonal therapy (progesterone and tamoxifen). In the untreated group, staining for ER and PR colocalized with that for HMB-45, but not with that for membrane type-1 matrix metalloproteinase (MT-1-MMP), which we have shown to be localized in proliferating LAM cells. These observations demonstrate that PR and ER are selectively expressed in a subpopulation of LAM cells that are larger in size, have a limited ability to proliferate, and do not produce MT-1 MMP, the enzyme that activates MMP-2 (which is secreted by LAM cells and is capable of lysing elastin and collagens). ER and PR in LAM cells appear to be downregulated by hormonal therapy. PMID- 10712356 TI - Spiral computed tomography is comparable to angiography for the diagnosis of pulmonary embolism. AB - The use of spiral computed tomography (CT) for the diagnosis of pulmonary embolism has been compared to angiography, the current gold standard. However, the accuracy of pulmonary angiography has never been evaluated against an independent gold standard. The aim of this study was to compare contrast-enhanced spiral CT to pulmonary angiography for the detection of subsegmental-sized pulmonary emboli by using a methacrylate cast of porcine pulmonary vessels as an independent gold standard. We studied 16 anesthetized, juvenile pigs and injected colored methacrylate beads (3.8 mm, small; 4.2 mm, large) via the jugular vein. After embolization spiral CT (3 mm and 1 mm collimation), and pulmonary angiography were performed. Pigs were killed and the pulmonary arterial tree was cast using methacrylate. Spiral CT and angiography were interpreted independently by two radiologists. Sensitivity and 95% confidence intervals for 3 mm and 1 mm collimation CT and angiography, respectively, were: 82% (73 to 88%), 87% (79 to 93%), 87% (79 to 93%) (p = 0.42). Positive predictive values and 95% confidence intervals for 3 mm and 1 mm collimation CT and angiography, respectively, were: 94% (86 to 94%), 81% (73 to 88%), and 88% (80 to 93%). There was no difference between spiral CT and angiography for detection of subsegmental-sized pulmonary emboli. We conclude that spiral CT is comparable to angiography for detection of pulmonary emboli. PMID- 10712357 TI - Goblet cell hyperplasia and epithelial inflammation in peripheral airways of smokers with both symptoms of chronic bronchitis and chronic airflow limitation. AB - To quantify the number of goblet cells and inflammatory cells in the epithelium of peripheral airways in smokers with both symptoms of chronic bronchitis and chronic airflow limitation, we examined surgical specimens obtained from 25 subjects undergoing lung resection for localized pulmonary lesions: 10 smokers with symptoms of chronic bronchitis and chronic airflow limitation, six asymptomatic smokers with normal lung function, and nine nonsmoking control subjects. Peripheral airways were examined with histochemical methods to identify goblet cells and with immunohistochemical methods to identify total leukocytes (CD45(+) cells), neutrophils, macrophages, CD4(+) and CD8(+) cells in the epithelium. When compared with nonsmokers, smokers with both symptoms of chronic bronchitis and chronic airflow limitation had an increased number of goblet cells (p < 0.01), CD45(+) cells (p < 0. 01), macrophages (p < 0.05), and CD8(+) cells (p < 0.01) in the epithelium of peripheral airways. When all the smokers were grouped together, they showed an increased number of neutrophils (p < 0.05) along with an increased number of goblet cells, CD45(+) cells, macrophages and CD8(+) cells (p < 0.05) compared with nonsmokers. In conclusion, smokers with both symptoms of chronic bronchitis and chronic airflow limitation have an increased number of goblet cells and inflammatory cells in the epithelium of peripheral airways. PMID- 10712358 TI - Inhibition of CD11-CD18 complex prevents acute lung injury and reduces mortality after peritonitis in rabbits. AB - Acute lung injury is frequent after severe peritonitis. The aim of this study was to investigate whether inhibition of the adhesion molecule CD11-CD18 on polymorphonuclear leukocytes (PMNs) would have any beneficial effects on pulmonary function and mortality in an animal model reproducing these clinical conditions. Acute peritonitis was induced in 36 rabbits by intraperitoneal injection of zymosan (0.6 g/kg) suspended in mineral oil; 20 were pretreated with a murine-specific IgG2a anti-CD18 monoclonal antibody, 16 (controls) with nonspecific purified murine IgG (1 mg/kg). The animals were followed for 10 d, then killed for histologic examination of the lungs. Blood samples were taken on Days 0, 1, 3, 7, and 10 for red blood cell (RBC), white blood cell (WBC), and platelet counts, pH, PO(2), PCO(2), carbon dioxide content (HCO(3)(-)) measurements, and renal and liver tests. Treatment with the anti-CD18 monoclonal antibody reduced mortality by approximately 40% (p < 0.05). PO(2) was higher in these treated animals than in the control animals throughout the study (p < 0.05 on Day 1, 3, and 10). On Day 1 control animals had significant leukopenia, whereas anti-CD18-treated animals had a moderate increase of the number of circulating WBC compared with baseline values (p < 0.05 between groups). The lungs of the anti-CD18-treated animals showed minor signs of inflammation and PMN infiltration whereas controls had interstitial and intra-alveolar edema and a large number of granulocytes. Quantification of PMNs by morphometry showed that there were constantly less granulocytes in the lungs of the animals treated with the anti-CD18 antibody (p < 0.001). PMN infiltration correlated with the levels of PO(2) (p < 0.001). Lung tissue of anti-CD18-treated rabbits contained less malonyldialdehyde, a by-product of membrane lipid peroxidation by PMN oxygen radicals (950 +/- 120 versus 1,710 +/- 450 pM/mg of protein) and, conversely, more of the antioxidant alpha-tocopherol (136 +/- 22 versus 40 +/- 9 ng/mg of protein), than the control rabbits (p < 0.01). In this particular model of ARDS the monoclonal antibody against the CD11-CD18 complex had a beneficial effect, reducing PMN infiltration and oxygen radical release in the lungs, preventing alveolocapillary membrane damage, improving gas exchange and, finally, significantly reducing mortality. PMID- 10712359 TI - Importance of interleukin-8 in the development of reexpansion lung injury in rabbits. AB - Reexpansion of a collapsed lung induces increased microvascular permeability leading to reexpansion pulmonary edema (REPE). This study was designed to prove the hypothesis that local overproduction of interleukin-8 (IL-8) induces inflammatory cell accumulation which leads to the induction of REPE. Initially, we examined the detailed characteristics of a rabbit model of REPE in association with IL-8 production and its mRNA expression. The lung tissue to plasma ratio of radiolabeled albumin (T/P ratio), the lung wet to dry ratio, and bronchoalveolar lavage (BAL) neutrophil counts were significantly increased in the reexpanded lung. IL-8 concentrations and mRNA expression were significantly increased in the reexpanded lung homogenate. Immunohistochemically, alveolar macrophages (AMs) and epithelial cells in the reexpanded lung and AMs in the collapsed lung were positive for IL-8. Second, we examined the effect of pretreatment with a specific monoclonal anti-IL-8 antibody (Ab) or control IgG on the development of REPE. The T/P ratio and BAL neutrophil counts were conspicuously decreased by pretreatment with anti-IL-8 Ab, but not with control IgG. On a histopathological study, lung injury and leukocyte infiltration were attenuated by the pretreatment with anti IL-8 Ab. In conclusion, IL-8 production is enhanced in the reexpanded lung, and contributes to the development of REPE. The pretreatment with anti-IL-8 antibody may be useful as a novel protective therapy for this disease. PMID- 10712360 TI - Thoracic lymphangiomas, lymphangiectasis, lymphangiomatosis, and lymphatic dysplasia syndrome. PMID- 10712361 TI - Exhaled nitric oxide and exercise-induced bronchoconstriction in asthmatic children. AB - It is known that exhaled nitric oxide (ENO) is increased in asthmatic individuals, probably as an expression of airway inflammation, but no studies have been reported of ENO and exercise-induced bronchoconstriction (EIB). We assessed the effect of a treadmill exercise challenge on ENO concentration in 24 asthmatic children aged 11.2 +/- 0.4 yr (mean +/- SEM). According to the presence or absence of EIB, the children were divided into an EIB group (n = 10) and a non EIB group (n = 14). ENO was measured with a single-breath reservoir technique. FEV(1), ENO, and heart rate were measured at baseline and 1, 6, 12, and 18 min after the end of exercise. We also measured ENO in 18 healthy control children aged 10.8 +/- 0.6 yr, of whom nine underwent an exercise challenge identical to that of the asthmatic children. After the exercise test, the mean decrease in FEV(1) was 34% in the EIB group and 5% in the non-EIB group. The EIB group had higher baseline ENO values (12.3 +/- 1.6 ppb) than the healthy children (6.1 +/- 0.2 ppb) (p < 0.01). The time course of ENO was similar in the EIB, non-EIB, and control groups, with no significant changes after exercise (p = NS). In the overall group of asthmatic children there was a significant correlation (r = 0.61, p < 0.01) between baseline (preexercise) ENO and magnitude of the maximal decrease in FEV(1) after exercise. In conclusion, our study shows that ENO levels do not change during acute airway obstruction induced by exercise challenge in asthmatic children. In addition, baseline ENO values correlate with the magnitude of postexercise bronchoconstriction, suggesting that NO may be a predictor of airway hyperresponsiveness to exercise. PMID- 10712362 TI - Bronchial hyperresponsiveness in children and adolescents with Crohn's disease. AB - Pulmonary manifestations have been described in Crohn's disease (CD). Bronchial responsiveness to methacholine (MCh) was evaluated in 14 children with CD with no evidence of airway disease, 10 asthmatics, and 10 healthy subjects. In patients with CD total blood eosinophils and serum IgE were 0.20 x 10(9) x L(-1) (95% CI 1.68 to 2.08) and 138.4 kU x L(-)(1) (95% CI 18.84 to 257.96), respectively. Three patients with CD had positive prick tests. Bronchial hyperresponsiveness (BHR) was demonstrated in 10 patients with CD (71%) and in the asthmatics, but not in control subjects. In patients with CD PD(20) appeared significantly greater than in asthmatics (699 microg [95% CI 238 to 1,115] versus 104 microg [95% CI 37.35 to 293]; p < 0.05), and was not related either to baseline FEV(1) or IgE or eosinophils (r = 0.32; r = -0.5; r = -0.15, p = NS, respectively). Neither activity nor treatment or duration of CD affected BHR. Five nonatopic CD patients underwent a second MCh challenge over a 25-mo period: the PD(20) appeared significantly greater than basal PD(20) (1,941 microg versus 575 microg, p < 0.05, respectively), in the absence of significant changes of disease activity. BHR might be the expression of subclinical airway inflammation, a phenomenon which can be responsible for the development of various pulmonary manifestations in CD. PMID- 10712363 TI - Fine mapping of PPH1, a gene for familial primary pulmonary hypertension, to a 3 cM region on chromosome 2q33. AB - Familial primary pulmonary hypertension (PPH) is a rare autosomal dominant disease characterized by distinctive changes in pulmonary arterioles that lead to increased pulmonary artery pressures, right ventricular failure, and death. Our previous studies had mapped the disease locus, PPH1, to a 27-cM region on chromosome 2q31-q33, with a maximum multipoint logarithm of the odds favoring genetic linkage score of 3.87 with markers D2S350 and D2S364. To narrow the minimal genetic region for PPH, we physically mapped 33 highly polymorphic microsatellite markers and used them to genotype 44 affected individuals and 133 unaffected individuals from 17 families with PPH. We observed recombination events that substantially reduced the interval for PPH1 to the approximately 3-cM region that separates D2S311 and D2S1384. This entire region lies within chromosome 2q33. A maximum two-point lod score of 7.23 at a recombination fraction of zero was obtained for marker D2S307. A maximum multipoint lod score of 7.41 was observed close to marker D2S1367. The current minimal genetic region contains multiple candidate genes for PPH, including a locus thought to play a role in lung cancer. PMID- 10712365 TI - Airway smooth muscle in asthma. Perturbed equilibria of myosin binding. PMID- 10712366 TI - Deciphering the homeokinetic code of airway smooth muscle. PMID- 10712367 TI - What makes the airways contract abnormally? Is it inflammation? PMID- 10712368 TI - Airway remodeling in asthma. Unanswered questions. PMID- 10712369 TI - Key factors in the development of asthma: atopy. PMID- 10712370 TI - Endogenous inhibitory mechanisms in asthma. PMID- 10712371 TI - Why does airway inflammation persist? Is it failure to treat early? PMID- 10712372 TI - Why does airway inflammation persist? Is it leukotrienes? PMID- 10712373 TI - Interactions between corticosteroids and beta-adrenergic agonists in asthma disease induction, progression, and exacerbation. PMID- 10712374 TI - beta(2)-adrenergic receptor pharmacogenetics. PMID- 10712375 TI - The genetics of asthma. The important questions. PMID- 10712376 TI - What determines asthma phenotype? Is it the interaction between allergy and the smooth muscle? PMID- 10712377 TI - What determines asthma phenotype? Respiratory infections and asthma. PMID- 10712379 TI - The fick principle and the steady state PMID- 10712378 TI - What is the relationship between airway hyperresponsiveness and atopy? PMID- 10712380 TI - Platelet glycoprotein IIb/IIIa receptors and Glanzmann's thrombasthenia. PMID- 10712381 TI - Altered flow-induced arterial remodeling in vimentin-deficient mice. AB - The endothelial cytoskeleton plays a key role in arterial responses to acute changes in shear stress. We evaluated whether the intermediate filament protein vimentin is involved in the structural responses of arteries to chronic changes in blood flow (BF). In wild-type mice (V+/+) and in vimentin-deficient mice (V-/ ), the left common carotid artery (LCA) was ligated near its bifurcation, and 4 weeks later, the structures of the occluded and of the contralateral arteries were evaluated and compared with the structures of arteries from sham-operated mice. Body weight and mean carotid artery BF did not differ between the strains, but LCA and right carotid artery (RCA) diameter (737+/-14 microm [LCA] and 723+/ 14 microm [RCA] for V-/- versus 808+/-20 microm [LCA] and 796+/-20 microm [RCA] for V+/+) and medial cross-sectional area (CSAm) were significantly smaller in V /- (21+/-1 and 22+/-2 x 10(3) microm(2) for LCA and RCA, respectively) than in V+/+ (28+/-2 and 28+/-3 x 10(3) microm(2) for LCA and RCA, respectively). In V+/+, LCA ligation eliminated BF in the occluded vessel (before ligation, 0. 35+/ 0.02 mL/min) and increased BF from 0.34+/-0.02 to 0.68+/-0.04 mL/min in the RCA. In V-/-, the BF change in the occluded LCA was comparable (from 0.38+/-0.05 mL/min to zero-flow rates), but the BF increase in the RCA was less pronounced (from 0.33+/-0.02 to 0. 50+/-0.05 mL/min). In the occluded LCA of V+/+, arterial diameter was markedly reduced (-162 microm), and CSAm was significantly increased (5 x 10(3) microm(2)), whereas in the high-flow RCA of V+/+, carotid artery diameter and CSAm were not significantly modified. In the occluded LCA of V-/-, arterial diameter was reduced to a lesser extent (-77 microm) and CSAm was increased to a larger extent (10 x 10(3) microm(2)) than in V+/+. In contrast to V+/+, the high-flow RCA of V-/- displayed a significant increase in diameter (52 microm) and a significant increase in CSAm (5 x 10(3) microm(2)). These observations provide the first direct evidence for a role of the cytoskeleton in flow-induced arterial remodeling. Furthermore, they dissociate (1) between acute and chronic arterial responses to altered BF, (2) between alterations of lumen diameter and wall mass during arterial remodeling, and (3) between developmental and imposed flow-induced arterial remodeling. PMID- 10712382 TI - Fluid shear stress induces heat shock protein 60 expression in endothelial cells in vitro and in vivo. AB - Recent investigations indicate that the initial event in the pathogenesis of atherosclerosis involves an (auto)immunologic injury to the vessel wall. Heat shock proteins (hsps), which are expressed on the endothelial cell surface, constitute possible autoantigens. After being exposed to shear stress of 30 dyne/cm(2) in vitro by means of a rotational viscometer, human umbilical vein endothelial cells were immunohistochemically stained for hsp 60 by the monoclonal antibody ML-30; static control cells were negative. Maximal hsp 60 induction was observed after 12 hours of hemodynamic stress. In Northern blots, the level of hsp 60 mRNA was markedly increased after only 1 hour of shear stress in human umbilical vein endothelial cells compared with static control cells. In vivo investigations in Lewis rats confirmed these in vitro findings: the intima and media of frozen sections of the right common carotid artery exposed to increased wall shear stress (after ligation of the left common carotid artery) were stained for hsp 60. The vessel wall of the left low-shear-stress-exposed side was negative. These findings demonstrate that shear stress results in hsp 60 induction in endothelial cells in vivo and in vitro, providing the prerequisite for humoral and cellular reactions to endothelial hsp in the earliest stages of atherosclerosis. PMID- 10712383 TI - Enhanced expression of osteopontin in human diabetic artery and analysis of its functional role in accelerated atherogenesis. AB - We have previously reported that high glucose stimulates osteopontin (OPN) expression through protein kinase C-dependent pathways as well as hexosamine pathways in cultured rat aortic smooth muscle cells. The finding prompted us to study in vivo expression of OPN in diabetes mellitus. In the present study, we found by immunohistochemistry that medial layers of the carotid arteries of streptozotocin-induced diabetic rats and the forearm arteries of diabetic patients stained positively for OPN antibodies, whereas the staining from arteries of control rats and nondiabetic patients was negative. We also found that OPN stimulated the migration and enhanced platelet-derived growth factor (PDGF)-mediated DNA synthesis of cultured rat aortic smooth muscle cells. OPN and PDGF synergistically activated focal adhesion kinase as well as extracellular signal-regulated kinase; this finding seems to explain the OPN-induced enhancement of PDGF-mediated DNA synthesis. Taken together, our present results raise a possibility that OPN plays a role in the development of diabetic vascular complications. PMID- 10712384 TI - Regulation of cdk2 activity in endothelial cells that are inhibited from growth by cell contact. AB - Endothelial cells (ECs) are quiescent in normal blood vessels but undergo rapid bursts of proliferation after vascular injury and during angiogenesis. Here we show that the activity of cyclin-dependent kinase-2 (cdk2), a key regulator of the G1 and S phases of the cell cycle, is expressed at high levels in proliferating ECs but at low levels in ECs that are contact-inhibited for growth. Despite these differences in kinase activity, the protein levels of cdk2 and 1 of its activating subunits, cyclin E, are not modulated by these different growth conditions. The cdk inhibitor p27 is highly expressed in contact-inhibited but not proliferating ECs, whereas the level of cyclin A protein is preferentially expressed in proliferating ECs. p27 protein was detected in immunoprecipitable complexes with cdk2 or cyclin E in cultures that were contact-inhibited for growth. The functional significance of the p27 induction was indicated by the detection of a heat-stable cdk2 inhibitory activity that was induced by endothelial cell-cell contact and could be immunodepleted with anti-p27 antibodies. In a confluent EC monolayer, cdk2 kinase activity was activated by a scraping injury that led to cell migration and proliferation. The injury-induced activation of cdk2 coincided with the downregulation of p27 and the induction of cyclin A. These data demonstrate that p27 is induced in confluent cultures of ECs. They also indicate that both p27 induction and cyclin A downregulation contribute to the inhibition of cdk2 and cell proliferation by cell-cell contact in ECs. PMID- 10712385 TI - p53, p21(WAF1/CIP1), and MDM2 involvement in proliferation and apoptosis in an in vitro model of conditionally immortalized human vascular smooth muscle cells. AB - Using an in vitro model of a conditionally immortalized cell line, we investigated how human vascular smooth muscle cells (VSMCs) are affected by the expression of simian virus 40 (SV40) large T antigen (LT antigen), which binds to cell cycle regulators, such as the tumor suppressor protein p53. Cells were obtained after infection of saphenous vein-derived VSMCs with a nonreplicative retroviral vector containing a temperature-sensitive (ts) mutant of SV40 LT antigen and were shown to have maintained some characteristics and responses of VSMCs. Under permissive temperature conditions (36 degrees C), the increased rate of cell proliferation was shown to be associated with expression of LT antigen and with LT-antigen binding to and inactivation of p53. p53 inactivation failed to block apoptosis induced by serum withdrawal or by UV irradiation. Downregulation of LT-antigen expression at the nonpermissive temperature (39 degrees C) was shown to be associated with growth arrest, increased expression of the cell cycle inhibitor p21(WAF1/CIP1), increased MDM2-promoter activity, and differential expression of MDM2 gene products, suggesting that p53-induced transcription/transactivation may be involved in VSMC cell cycle control but not necessarily apoptosis. The established SMC line HVTs-SM1 may be a useful model for the study of processes involved in myointimal hyperplasia and cellular aging, as well as for the study of cell cycle control in general. PMID- 10712386 TI - Angiotensin II stimulates endothelial vascular cell adhesion molecule-1 via nuclear factor-kappaB activation induced by intracellular oxidative stress. AB - The recruitment of monocytes via the endothelial expression of vascular cell adhesion molecule-1 (VCAM-1) is a key step in the formation of the initial lesion in atherosclerosis. Because angiotensin (Ang) II may be involved in this process, we investigated its role on the signaling cascade leading to VCAM-1 expression in endothelial cells. Ang II stimulates mRNA and protein expression of VCAM-1 in these cells via the AT(1) receptor. This effect was enhanced by N(G)-nitro-L arginine methyl ester, a nitric oxide synthase inhibitor, and blocked by pyrrolidinedithiocarbamate, an antioxidant molecule. Ang II activated the redox sensitive transcription factor nuclear factor-kappaB and stimulated the degradation of both inhibitor of kappaB (IkappaB)alpha and IkappaBbeta with different kinetics. The degradation of IkappaBs induced by Ang II was not modified by incubation with exogenous superoxide dismutase and catalase, suggesting that this effect was not mediated by the extracellular production of O(2)(-). In contrast, rotenone and antimycin, 2 inhibitors of the mitochondrial respiratory chain, inhibited the Ang II-induced IkappaB degradation, showing that generation of reactive oxygen species in the mitochondria is involved on Ang II action. BXT-51702, a glutathione peroxidase mimic, inhibited the effect of Ang II, and aminotriazole, an inhibitor of catalase, enhanced it, suggesting a role for H(2)O(2) in IkappaB degradation. This is confirmed by experiments showing that Ang II stimulates the intracellular production of H(2)O(2) in endothelial cells. These results demonstrate that Ang II induced an intracellular oxidative stress in endothelial cells, which stimulates IkappaB degradation and nuclear factor-kappaB activation. This activation enhances the expression of VCAM-1 and probably other genes involved in the early stages of atherosclerosis. PMID- 10712387 TI - Regulation by fibrinogen and its products of intercellular adhesion molecule-1 expression in human saphenous vein endothelial cells. AB - It has been reported that fibrinogen may act as a bridging ligand, binding to intercellular adhesion molecule-1 (ICAM-1) on human umbilical vein endothelial cells and to Mac-1 on THP-1 cells (a monocytic cell line) to increase adhesion. In this study, we investigated whether fibrinogen altered the expression of ICAM 1 and, thus, increased the adhesion of THP-1 cells to cultured human saphenous vein endothelial cells (HSVECs). Incubation of HSVECs with 0.3 to 4 micromol/L fibrinogen caused a time- and concentration-dependent increase in ICAM-1, as determined by ELISA. The 4- to 5-fold increase in ICAM-1 protein concentration in HSVECs stimulated by 4 micromol/L fibrinogen for 6 hours was concomitant with a 4 to 5-fold increase in ICAM-1 mRNA. This fibrinogen-stimulated ICAM-1 upregulation was associated with a 2-fold increase in THP-1 cell adhesion to HSVECs. The fibrinogen-derived peptide Bbeta15-42 bound to HSVECs (K(d) 0.18 micromol/L). Preincubation of HSVECs with Bbeta15-42, a neutralizing antibody to urokinase plasminogen activator (uPA), or the F(ab)(1) fragment of a monoclonal antibody to vascular endothelial cadherin significantly attenuated the increase in ICAM-1 stimulated by fibrinogen. Capillary electrophoretic analysis indicated that anti-uPA prevented the release of any fibrinopeptide B (Bbeta1-14) in cultures of HSVECs incubated with 4 micromol/L fibrinogen for 6 hours. Moreover, incubation of HSVECs with either fibrin monomer (1 micromol/L) or monoclonal antibodies to vascular endothelial cadherin (25 microg/mL) increased ICAM-1 protein concentration 3- to 4-fold. These findings indicate that cleavage of fibrinopeptide B from fibrinogen by endothelial uPA permits the exposed Bbeta15 42 sequence of fibrinogen to bind to vascular endothelial cadherin on HSVECs and to upregulate the expression of ICAM-1. PMID- 10712388 TI - Nitric oxide induces the synthesis of vascular endothelial growth factor by rat vascular smooth muscle cells. AB - Vascular endothelial growth factor (VEGF) is known to induce the release of nitric oxide (NO) from endothelial cells. However, the effect of NO on VEGF synthesis is not clear. Accordingly, the effect of endogenous and exogenous NO on VEGF synthesis by rat vascular smooth muscle cells (VSMCs) was investigated. Two in vitro models were used: (1) VSMCs stimulated to produce NO by treatment with interleukin (IL)-1beta (10 ng/mL) and (2) VSMCs lipotransfected with pKecNOS plasmid, containing the endothelial constitutive NO synthase (ecNOS) cDNA. The synthesis of NO was inhibited by N(omega)-nitro-L-arginine methyl ester (L-NAME, 2 to 5 mmol/L) or diaminohydroxypyrimidine (DAHP, 2.5 to 5 mmol/L), inhibitors of NOS and GTP cyclohydrolase I, respectively. Some cells treated with L-NAME or DAHP were supplemented with L-arginine (10 mmol/L) or tetrahydrobiopterin (BH(4); 100 micromol/L), respectively. In addition, we studied the effect of sodium nitroprusside (SNP; 10 and 100 micromol/L) and chemically related compounds, potassium ferrocyanide and ferricyanide, on VEGF generation. IL-1beta induced iNOS expression and NO generation and significantly upregulated VEGF mRNA expression and protein synthesis. L-NAME and DAHP totally inhibited NO generation and decreased the IL-1beta-upregulated VEGF synthesis by 30% to 40%. Supplementation with L-arginine or BH(4) increased NO generation by L-NAME- or DAHP-treated cells, and VEGF synthesis was augmented by addition of BH(4). The cells generating NO after pKecNOS transfection released significantly higher amounts of VEGF than cells transfected with control plasmids. Inhibition of NO generation by L-NAME decreased VEGF synthesis. In contrast to the effect of endogenous NO, we observed the inhibition of VEGF synthesis in the presence of high (10 or 100 micromol/L) concentrations of SNP. This effect was mimicked by chemically related ferricyanide and ferrocyanide compounds, suggesting that the inhibitory effect of sodium nitroprusside may be mediated by an NO-independent mechanism. The results indicate that endogenous NO enhances VEGF synthesis. The positive interaction between endogenous NO and VEGF may have implications for endothelial regeneration after balloon angioplasty and for angiogenesis. PMID- 10712389 TI - Local delivery of platelet-derived growth factor receptor-specific tyrphostin inhibits neointimal formation in rats. AB - Signal transduction through the platelet-derived growth factor (PDGF)/PDGF receptor (PDGFR) system is involved in the process of postangioplasty restenosis. Tyrphostins are low molecular weight inhibitors of protein tyrosine kinases. We assessed the antiproliferative effects of PDGFRbeta-specific tyrphostin AG-1295 in vitro and in vivo. AG-1295 significantly inhibited rat smooth muscle cell growth stimulated by PDGF-BB or FCS. This antiproliferative effect was paralleled by reversible reduction of the total phosphotyrosine level and the degree of PDGFRbeta phosphorylation by the drug in vitro. Local sustained delivery of the drug from perivascularly implanted polymeric matrices resulted in focal AG-1295 levels of 711 and 29.1 ng/mg of dry arterial tissue 1 and 14 days after implantation in rats. AG-1295 delivered from polymeric matrices resulted in a 35% reduction of neointimal formation on day 14 after balloon injury in the rat carotid model. Tyrosine phosphorylation of certain transduction proteins in arterial tissue extracts was significantly upregulated by balloon injury on day 3 but was essentially returned to or below basal levels 14 days after injury. Tyrphostin treatment decreased tyrosine phosphorylation at both time points below the basal levels. Moreover, the enhancement of PDGFRbeta expression 3 and 14 days after arterial injury was strongly inhibited by AG-1295 treatment. It can be concluded that AG-1295 reduces neointimal formation by inhibiting PDGFbeta triggered tyrosine phosphorylation. PMID- 10712390 TI - Cyclooxygenase-2 regulates granulocyte-macrophage colony-stimulating factor, but not interleukin-8, production by human vascular cells: role of cAMP. AB - Vascular smooth muscle is now recognized as an important site of mediator generation under inflammatory conditions. Indeed, the release of leukocyte activators, such as granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin (IL)-8, by human arterial smooth muscle cells has recently been demonstrated. However, the potential for venous cells to release GM-CSF has not been addressed. We have shown that human vascular smooth muscle cells express the "inflammatory" form of cyclooxygenase (COX), cyclooxygenase-2 (COX-2), when stimulated with cytokines. In some nonvascular cell types, the COX activity has been shown to regulate the release of GM-CSF and IL-8, although the nature of the isoform responsible was not addressed. We show that human venous smooth muscle cells, like their arterial counterparts, release GM-CSF after stimulation with IL 1beta. Similarly, both cell types released IL-8. Under the same conditions, we found that COX-2 activity suppressed GM-CSF, but not IL-8, release by both types of human vascular cells. Moreover, the prostacyclin mimetic, cicaprost, and the cAMP analogue, dibutyryl cAMP, inhibited GM-CSF release from these cells. These observations suggest that COX-2 activity suppresses GM-CSF release via a cAMP dependent pathway in human vascular cells and illustrates a novel mechanism by which this enzyme can modulate immune and inflammatory events. PMID- 10712391 TI - Plasma levels of extracellular superoxide dismutase in an Australian population: genetic contribution to normal variation and correlations with plasma nitric oxide and apolipoprotein A-I levels. AB - Extracellular superoxide dismutase (EC-SOD) is a major superoxide scavenger and may be important to normal vascular function and cardiovascular health. We analyzed family data from 610 healthy Australians to detect and quantify the effects of genes on normal variation in plasma levels of EC-SOD and to test for pleiotropy with plasma nitric oxide (NO) and apolipoprotein A-I (apoA-I). Using maximum-likelihood-based variance decomposition methods, we determined that sex, age, and plasma levels of HDL cholesterol, apoA-I, and creatinine accounted for 38.6% of the variance in plasma EC-SOD levels and that additive genes accounted for 35% (P<0.00002). Multivariate analyses of plasma levels of EC-SOD, NO(x) (a measure of basal NO production), and apoA-I detected significant genetic correlations, indicating pleiotropy between EC-SOD and apoA-I (genetic correlation [rho(G)]=-0.45) and between NO(x) and apoA-I (rho(G)=0.58) but not between EC-SOD and NO(x). Genes shared by EC-SOD and apoA-I account for 20% of the genetic variance and, respectively, 7% and 9% of the phenotypic variance in both traits. Shared genes also account for >33% of the genetic variance and 5% and 15% of the respective phenotypic variance in NO(x) and apoA-I. In healthy individuals, over a third of the variance in EC-SOD plasma levels is due to the additive effects of genes. Some genes influence EC-SOD and apoA-I levels. The same is true of NO(x) and apoA-I but not of EC-SOD and NO(x). These patterns of pleiotropy can guide subsequent attempts to identify the genes and physiological mechanisms underlying them. PMID- 10712393 TI - Oxidized low density lipoprotein is a prognostic marker of transplant-associated coronary artery disease. AB - Retrospective studies identified oxidized low density lipoprotein (LDL) in the blood as a diagnostic marker of coronary artery disease (CAD). This prospective study sought to determine the prognostic value of oxidized LDL for CAD in cardiac transplant patients. Oxidized LDL was measured in 99 cardiac transplant patients with normal coronary angiograms at baseline and was measured again after a median follow-up of 2 years at the time of a second angiogram. Twenty-one patients developed angiographically detectable cardiac transplant vasculopathy (cases), and 78 individuals did not (controls). Cases had significantly higher baseline plasma levels of oxidized LDL than did controls: 1.18+/-0.70 versus 0.57+/-0.20 mg/dL (mean+/-SD, P<0.0001). The increase of oxidized LDL at the end of the follow-up was significantly higher in cases than in controls: 0. 75+/-0.73 mg/dL versus 0.14+/-0.27 mg/dL (P<0.0001). Baseline levels of oxidized LDL predicted cardiac transplant vasculopathy (chi(2)=16, P<0.0001) independent of pretransplant ischemic cardiomyopathy, time after transplantation, age, and serum levels of LDL and high density lipoprotein cholesterol. The development of transplant CAD was associated with a further increase of plasma levels of oxidized LDL (chi(2)=14, P=0.0002). Oxidized LDL is a prognostic marker of transplant CAD. PMID- 10712392 TI - In vivo uptake of radiolabeled MDA2, an oxidation-specific monoclonal antibody, provides an accurate measure of atherosclerotic lesions rich in oxidized LDL and is highly sensitive to their regression. AB - To determine whether labeled antibodies against oxidized LDL (OxLDL) offer advantages for quantifying atherosclerosis, we compared in vivo aortic uptake of (125)I-labeled MDA2, a monoclonal antibody against malondialdehyde-lysine epitopes), atherosclerotic surface area, and aortic weight in Watanabe heritable hyperlipidemic and New Zealand White rabbits and in low density lipoprotein receptor-deficient (LDLR(-/-)) and apolipoprotein E-deficient (apoE(-/-)) mice. Absolute and specific uptakes of (125)I-MDA2 were significantly greater in plaque than in normal aortas. Uptake of (125)I-MDA2 significantly correlated with aortic weight and percent atherosclerotic surface area in rabbits and mice. To assess whether (125)I-MDA2 uptake reflects changes in lesion content of OxLDL, in a separate study, extensive atherosclerosis was induced in 4 groups of LDLR(-/-) mice by feeding them a high fat/cholesterol diet for 6 months. A baseline group was euthanized at this time. The remaining groups were fed "regression" diets (chow or chow+1% vitamin E+0.05% vitamin C) or the high fat/cholesterol diet for 6 more months. When atherosclerosis was measured as percent surface area or aortic weight, there was strong progression in the high fat/cholesterol group, moderate progression in the chow group, and no progression in the chow+vitamin E+vitamin C group compared with the baseline group. The (125)I-MDA2 method also yielded a significant increase in atherosclerosis in the high fat/cholesterol group but significant decreases in the chow and chow+vitamin E+vitamin C groups. Immunocytochemistry showed fewer oxidation-specific epitopes in lesions from the chow and chow+vitamin E+vitamin C groups. Thus, the uptake of (125)I-MDA2 correlates well with traditional measures of atherosclerosis but also reflects reduced plaque OxLDL content after hypocholesterolemic intervention. PMID- 10712394 TI - Antibodies against oxidized LDL and carotid artery intima-media thickness in a healthy population. AB - Oxidation of LDLs plays an important role in atherosclerosis, and immune response to oxidized LDL (oxLDL) may modulate atherogenesis. Although immunization with oxLDL is shown to suppress atherogenesis in animal models, the role of the immune response to oxLDL is not well established in humans. We investigated the relationship between the titer of anti-oxLDL antibody (oxLDL Ab) and arterial wall thickness in a healthy population with no clinical signs of atherosclerosis. Intima-media thickness of the carotid arteries (CA-IMT) was measured by high resolution B-mode ultrasonography in 446 healthy subjects. The titer of IgG-class oxLDL Ab was measured by a solid-phase ELISA. In univariate analysis, CA-IMT correlated positively with age, systolic blood pressure, total cholesterol, triglyceride, LDL cholesterol, body mass index, and waist-to-hip ratio, whereas it correlated negatively with HDL cholesterol and oxLDL Ab titer. The inverse association between oxLDL Ab titer and CA-IMT remained significant in multiple regression analysis, which took other confounding variables into account. These results indicate an independent inverse relationship between oxLDL Ab titer and CA-IMT in healthy subjects, supporting the hypothesis that immune response to oxLDL may have a protective role at an early stage of human atherosclerosis. PMID- 10712395 TI - Oxidized cholesterol in the diet accelerates the development of atherosclerosis in LDL receptor- and apolipoprotein E-deficient mice. AB - The aim of the current study was to determine whether oxidized cholesterol in the diet accelerates atherosclerosis in low density lipoprotein receptor- (LDLR) and apolipoprotein E- (apo E) deficient mice. Mice were fed either a control diet or a diet containing oxidized cholesterol. For LDLR-deficient mice, the control diet consisted of regular mouse chow to which 1.0% cholesterol was added. The oxidized diet was identical to the control diet except that 5% of the added cholesterol was oxidized. In apo E-deficient mice, the control diet contained 0.15% cholesterol, whereas in the oxidized diet, 5% of the added cholesterol was oxidized. LDLR-deficient and apo E-deficient mice were fed the experimental diets for 7 and 4 months, respectively. In mice fed the oxidized-cholesterol diets, the levels of oxidized cholesterol in sera were increased. At the end of the experiment, aortas were removed and atherosclerosis was assessed. We found that in LDLR-deficient mice, feeding of an oxidized-cholesterol diet resulted in a 32% increase in fatty streak lesions (15.93+/-1.59% versus 21.00+/-1.38%, P<0.03). Similarly, in apo E-deficient mice, feeding of an oxidized-cholesterol diet increased fatty streak lesions by 38% (15.01+/-0.92% versus 20. 70+/-0.86%, P<0.001). The results of the current study thus demonstrate that oxidized cholesterol in the diet accelerates fatty streak lesion formation in both LDLR- and apo E-deficient mice. PMID- 10712396 TI - Identification of soluble forms of lectin-like oxidized LDL receptor-1. AB - Lectin-like oxidized LDL receptor-1 (LOX-1) is a type II membrane protein belonging to the C-type lectin family molecules, which can act as a cell-surface endocytosis receptor for atherogenic oxidized LDL. In this study, we show that soluble forms of LOX-1 are present in conditioned media of cultured bovine aortic endothelial cells (BAECs) and CHO-K1 cells stably transfected with LOX-1 cDNA. Immunoblot analysis of conditioned media from TNF-alpha-activated BAECs and CHO K1 cells stably expressing LOX-1 revealed that soluble LOX-1 has an approximate molecular mass of 35 kDa. In TNF-alpha-activated BAECs, cell-surface expression of LOX-1 precedes soluble LOX-1 production. Cell-surface biotinylation followed by immunoprecipitation and immunoblotting showed that soluble LOX-1 in cell conditioned media is derived from LOX-1 expressed on the cell surface. Production of soluble LOX-1 was inhibited by PMSF, suggesting that PMSF-sensitive proteases may be involved in this process. Purification of soluble LOX-1 by high performance liquid chromatography and N-terminal amino acid sequencing of soluble LOX-1 identified the 2 cleavage sites between Arg(86)-Ser(87) and Lys(89) Ser(90), which were located in the membrane proximal extracellular domain of LOX 1. The data demonstrate that cell-surface LOX-1 can be cleaved at 2 different sites and transformed into soluble forms. Further studies may explore therapeutic and diagnostic applications of soluble LOX-1 in atherosclerotic diseases. PMID- 10712397 TI - Gene transfer and hepatic overexpression of the HDL receptor SR-BI reduces atherosclerosis in the cholesterol-fed LDL receptor-deficient mouse. AB - HDL cholesterol levels in humans are inversely correlated with the risk of atherosclerosis. The class B scavenger receptor type I (SR-BI) is the first molecularly well-defined HDL receptor, and hepatic overexpression of SR-BI in normal mice has been shown to result in decreased plasma HDL cholesterol levels. To determine whether SR-BI overexpression is proatherogenic or is protective against atherosclerosis, LDL receptor-deficient mice were placed on a high fat/high-cholesterol diet for 2 or 12 weeks to induce atherosclerotic lesions of different stages and then were injected with a recombinant adenovirus encoding murine SR-BI. Transient hepatic overexpression of SR-BI in mice with both early and advanced lesions significantly decreased atherosclerosis. SR-BI expression was associated with markedly decreased HDL cholesterol and either unchanged or only modestly reduced non-HDL cholesterol levels; in all experiments, the mean HDL cholesterol levels were significantly correlated with atherosclerotic lesion size. These data suggest that interventions that promote HDL cholesterol transport and lower plasma HDL cholesterol levels can suppress atherosclerosis, even when initiated after significant lesion development. Thus, stimulation of hepatic SR-BI activity may provide a novel target for therapeutic intervention in atherosclerotic cardiovascular disease. PMID- 10712398 TI - Hypercholesterolemia increases coronary endothelial dysfunction, lipid content, and accelerated atherosclerosis after heart transplantation. AB - Hyperlipidemia may increase endothelial damage and promote accelerated atherogenesis in graft coronary vasculopathy. To study the effects of hypercholesterolemia on coronary endothelial dysfunction, intimal hyperplasia, and lipid content, a porcine model of heterotopic heart transplantation, allowing nonacute rejection without immunosuppressive drugs, was used. A high cholesterol diet was fed to donor and recipient swine 1 month before and after transplantation. The endothelial function of coronary arteries of native and transplanted hearts from cholesterol-fed animals was studied in organ chambers 30 days after implantation and compared with endothelial function in arteries from animals fed a normal diet. The total serum cholesterol increased 3-fold in donors and recipients. Endothelium-dependent relaxations to serotonin, to the alpha(2) adrenergic agonist UK14,304, and to the direct G-protein activator sodium fluoride were decreased significantly in allografted hearts compared with native hearts from both groups. Relaxations to the calcium ionophore A23187 and bradykinin were decreased significantly in allografts from animals fed the high cholesterol diet. The prevalence of intimal hyperplasia was significantly increased in coronary arteries from hypercholesterolemic swine. There was a significant increase in the lipid content of allograft arteries of hypercholesterolemic recipients. Hypercholesterolemia causes a general coronary endothelial dysfunction, increases the prevalence of intimal hyperplasia, and augments the incorporation of lipids in the vascular wall after heart transplantation. Hyperlipidemia accelerates graft coronary atherosclerosis through its effects on the endothelium. PMID- 10712399 TI - Cholesterol-lowering treatment is associated with improvement in coronary vascular remodeling and endothelial function in patients with normal or mildly diseased coronary arteries. AB - Coronary vascular remodeling and altered endothelial function have been described in the early stages of native atherosclerosis. The purpose of this study was to evaluate the association between cholesterol-lowering therapy and coronary vascular remodeling and endothelial function in patients with normal or mildly diseases coronary arteries. Patients (N=101) with normal or mildly diseased coronary arteries by coronary angiography underwent intravascular ultrasound examination of the left anterior descending coronary artery. Vessel and lumen area, atherosclerotic plaque area, and plaque morphology were evaluated. Vascular reactivity was examined with the use of intracoronary adenosine, acetylcholine, and nitroglycerin. Patients were divided into 3 groups based on the total cholesterol levels: group 1 (n=25), patients with a history of hypercholesterolemia adequately treated (total cholesterol <240 mg/dL); group 2 (n=26), patients with hypercholesterolemia not adequately controlled (total cholesterol >/=240 mg/dL); and group 3 (n=50), patients without hypercholesterolemia. Vessel area and lumen area were significantly greater in groups 1 and 3 than in group 2 (for respective values in groups 1, 2, and 3: vessel area 11.9+/-0.5, 10.6+/-0.4, and 11.8+/-0.4 mm(2), both P<0.05; lumen area 8.3+/-0.4, 6.9+/-0.3, and 8.9+/-0.3 mm(2), both P<0.01). However, plaque areas in groups 1 and 2 were similar. Furthermore, acetylcholine-induced percent increases in coronary blood flow were significantly greater in groups 1 and 3 than in group 2 (for respective values in groups 1, 2, and 3: 70.5+/-20.1%, 22.8+/-13.7%, and 68.6+/-14.8%, both P<0. 05). Cholesterol-lowering treatment is associated with an improvement in coronary lumen area that results not from a decrease in plaque area but from an increase in vessel area, reflecting vascular remodeling. Additionally, this adaptive process may occur in association with an improvement of endothelium-dependent vasodilation of the resistance coronary artery. PMID- 10712400 TI - Not acute but chronic hypertriglyceridemia is associated with impaired endothelium-dependent vasodilation: reversal after lipid-lowering therapy by atorvastatin. AB - There is controversy regarding the relation between hypertriglyceridemia (HTG) and endothelial function. This study was designed to investigate endothelial function in a patient group with chronic HTG, before and during lipid-lowering therapy by atorvastatin. In addition, the effects of acute HTG on endothelial function were studied in normolipidemic individuals. Eight male patients with chronic HTG were studied before and after 6 weeks of lipid-lowering treatment with 80 mg atorvastatin once daily. Ten age-matched control subjects were studied at baseline and immediately after a high-dose infusion of artificial triglycerides. The endothelium-dependent response to serotonin was attenuated in the HTG group, whereas the response to acetylcholine was comparable to the response in the control group. The response to the endothelium-independent vasodilator nitroprusside was comparable in both groups. In response to atorvastatin therapy, serum triglyceride and cholesterol levels decreased significantly by 43% (paired t test, P=0.017) and 38% (paired t test, P=0.012), respectively. After 6 weeks of treatment, the forearm blood flow response to serotonin improved from 63% to 106% (ANOVA, P<0.001). Induction of acute HTG in the control subjects did not affect the forearm blood flow responses to serotonin and nitroprusside; however, the response to acetylcholine was paradoxically increased. In conclusion, patients with chronic HTG have an impaired endothelium dependent vasodilation to serotonin that is normalized after 6 weeks of lipid lowering therapy by atorvastatin. PMID- 10712401 TI - Expression of secretory group IIA phospholipase A(2) in relation to the presence of microbial agents, macrophage infiltrates, and transcripts of proinflammatory cytokines in human aortic tissues. AB - Recent seroepidemiological and immunohistochemical studies have demonstrated an association between microbial infections and atherosclerosis. However, the mechanisms underlying this association are widely unknown. In the present study, arterial specimens obtained at autopsy after sudden death were analyzed concerning (1) the presence of Chlamydia pneumoniae, cytomegalovirus, herpes simplex virus, and Helicobacter pylori; (2) the expression of secretory group IIA phospholipase A(2) (sPLA(2)-IIA) and of proinflammatory cytokines; and (3) the stage of atherosclerosis. Genomic DNA of microbial pathogens was determined by the polymerase chain reaction technique. The expression of sPLA(2)-IIA was studied immunohistochemically by using monoclonal antibodies against human sPLA(2)-IIA. Transcripts specific for sPLA(2)-IIA, interleukin-1beta, tumor necrosis factor-alpha, and interferon-gamma were identified by reverse transcription-polymerase chain reaction. In 18 of 102 analyzed specimens, DNA of microbial pathogens was found. Thirteen sections were positive for C pneumoniae, whereas 2 specimens were positive either for cytomegalovirus or for herpes simplex virus. One section contained genomic DNA of all 3 pathogens simultaneously. None of the analyzed tissues exhibited nucleic acids specific for H pylori. In addition to macrophage infiltrates, the presence of microbial DNA was closely associated with the occurrence of transcripts specific for proinflammatory cytokines and sPLA(2)-IIA. Pathogens as well as sPLA(2)-IIA and cytokines were found to be present not only in advanced but also in early stages of atherosclerosis. In tissues negative for sPLA(2)-IIA and cytokine expression, none of the pathogens could be identified. Because macrophages exposed to phospholipase A(2)-treated lipoproteins are transformed into foam cells in vitro, the results of this study suggest an alternative mechanism by which microbial infections may act in a proatherogenic fashion in vessel walls. PMID- 10712402 TI - Role of serum amyloid A during metabolism of acute-phase HDL by macrophages. AB - The serum amyloid A (SAA) family of proteins is encoded by multiple genes that display allelic variation and a high degree of homology in mammals. Triggered by inflammation after stimulation of hepatocytes by lymphokine-mediated processes, the concentrations of SAA may increase during the acute-phase reaction to levels 1000-fold greater than those found in the noninflammatory state. In addition to its role as an acute-phase reactant, SAA (104 amino acids, 12 kDa) is considered to be the precursor protein of secondary reactive amyloidosis, in which the N terminal portion is incorporated into the bulk of amyloid fibrils. However, the association with lipoproteins of the high-density range and subsequent modulation of the metabolic properties of its physiological carrier appear to be the principal role of SAA. Because SAA may displace apolipoprotein A-I, the major protein component of native high density lipoprotein (HDL), during the acute phase reaction, the present study was aimed at (1) investigating binding properties of native and acute-phase (SAA-enriched) HDL by J774 macrophages, (2) elucidating whether the presence of SAA on HDL particles affects selective uptake of HDL-associated cholesteryl esters, and (3) comparing cellular cholesterol efflux mediated by native and acute-phase HDL. Both the total and the specific binding at 4 degrees C of rabbit acute-phase HDL were approximately 2-fold higher than for native HDL. Nonlinear regression analysis revealed K(d) values of 7.0 x 10(-7) mol/L (native HDL) and 3.1 x 10(-7) mol/L (acute-phase HDL), respectively. The corresponding B(max) values were 203 ng of total lipoprotein per milligram of cell protein (native HDL) and 250 ng of total lipoprotein per milligram of cell protein (acute-phase HDL). At 37 degrees C, holoparticle turnover was slightly enhanced for acute-phase HDL, a fact reflected by 2-fold higher degradation rates. In contrast, the presence of SAA on HDL specifically increased (1. 7-fold) the selective uptake of HDL cholesteryl esters from acute-phase HDL by J774 macrophages, a widely used in vitro model to study foam cell formation and cholesterol efflux properties. Although ligand blotting experiments with solubilized J774 membrane proteins failed to identify the scavenger receptor-BI as a binding protein for both native and acute-phase HDL, 2 binding proteins with molecular masses of 100 and 72 kDa, the latter comigrating with CD55 (also termed decay-accelerating factor), were identified. During cholesterol efflux studies, it became apparent that the ability of acute-phase HDL with regard to cellular cholesterol removal was considerably lower than that for native HDL. This was reflected by a 1.7-fold increase in tau/2 values (22 versus 36 hours; native versus acute-phase HDL). Our observations of increased HDL cholesteryl ester uptake and reduced cellular cholesterol efflux (acute-phase versus native HDL) suggest that displacement of apolipoprotein A-I by SAA results in considerable altered metabolic properties of its main physiological carrier. These changes in the apolipoprotein moieties appear (at least in the in vitro system tested) to transform an originally antiatherogenic into a proatherogenic lipoprotein particle. PMID- 10712403 TI - The pathogenesis of foam cell formation: modified LDL stimulates uptake of co incubated LDL via macropinocytosis. AB - Previously, modified LDLs were shown to stimulate macropinocytosis in pigeon macrophages. Simultaneous intracellular trafficking of LDL and AcLDL, differentially labeled with colloidal gold, was done to determine whether uptake of LDL, which does not cause foam cell formation, was internalized via a separate route from AcLDL, which stimulates foam cell formation. AcLDL and LDL were followed at either low (12 microg/mL) concentrations near the saturation of high affinity binding sites or high (50 to 150 microg/mL) lipoprotein concentrations used to induce foam cell formation. The colloidal gold distribution and percentage of co-labeling as observed by transmission electron microscopy were determined for organelles involved with coated-pit endocytosis or macropinocytosis. LDL simultaneously incubated with AcLDL on macrophages at the low concentration was predominately internalized via coated-pit endocytosis. AcLDL was internalized via both coated-pit endocytosis and macropinocytosis at low concentration. At higher lipoprotein concentrations (50 to 150 microg/mL), AcLDL continued to be internalized via macropinocytosis. Interestingly, a significant portion of the co-incubated LDL, at high concentrations, also trafficked via macropinocytosis. LDL internalized by macropinosomes at high lipoprotein concentrations suggests that AcLDL-stimulated macropinocytosis might increase uptake of co-incubated lipoproteins. When (125)I-LDL was incubated with cold AcLDL, LDL degradation at 37 degrees C doubled, without a corresponding increase in cell association or total binding of LDL at 4 degrees C. These studies suggest that modified LDL-stimulated macropinocytosis is a mechanism for increased degradation of co-incubated LDL potentially leading to foam cell formation. PMID- 10712404 TI - Dehydroepiandrosterone retards atherosclerosis formation through its conversion to estrogen: the possible role of nitric oxide. AB - Dehydroepiandrosterone (DHEA) is speculated to have an antiatherosclerotic effect, although the mechanism of action remains unclear. The objective of the current study was to determine whether the antiatherosclerotic effect of DHEA is related to its conversion to estrogen and to define the role of nitric oxide (NO) in the antiatherosclerotic effect of DHEA. Forty-eight oophorectomized rabbits were divided into 5 groups and fed the following diets for 10 weeks: group 1, a regular rabbit diet plus 1% cholesterol (a high-cholesterol diet [HCD]); group 2, an HCD plus 0.3% DHEA; group 3, an HCD plus 0.3% DHEA and fadrozole (2.0 mg x kg( 1) x d(-1)), a specific aromatase inhibitor; group 4, an HCD plus 17beta estradiol (20 microg x kg(-1) x d(-1)); and group 5, a regular diet. Atherosclerotic lesions, lipid deposition in aortic vessels, and basal and stimulated NO release were measured in the aforementioned groups of rabbits. NO release was measured by using an NO-selective electrode as well as by measuring vascular responses and the plasma NO metabolites nitrite and nitrate. The plasma total cholesterol level was increased, but there were no significant differences in lipid profile in the 4 groups of rabbits that were fed the HCD. The area occupied by atherosclerosis in the thoracic aorta was diminished by approximately 60% in the DHEA-treated rabbits (group 2) compared with the HCD group of rabbits (group 1); there was a corresponding 80% decrease in the estradiol group (group 4) but only a 30% decrease in the DHEA plus fadrozole group (group 3). In the aortas of rabbits from groups 1 and 3, the acetylcholine-induced and tone-related basal NO-mediated relaxations were diminished compared with those of the controls (group 5). However, these relaxations were restored in the aortas of group 2 and 4 rabbits, and an increase in NO release was observed in groups 2 and 4 compared with groups 1 and 3, as measured by an NO-selective electrode. Injection of neither solvent (20% ethanol/distilled water) nor fadrozole significantly affected the atherosclerotic area or the NO-related responses described above. We conclude that approximately 50% of the total antiatherosclerotic effect of DHEA was achieved through the conversion of DHEA to estrogen. NO may also play a role in the antiatherosclerotic effect of DHEA and 17beta-estradiol. PMID- 10712405 TI - In vivo evidence for both lipolytic and nonlipolytic function of hepatic lipase in the metabolism of HDL. AB - To investigate the in vivo role that hepatic lipase (HL) plays in HDL metabolism independently of its lipolytic function, recombinant adenovirus (rAdV) expressing native HL, catalytically inactive HL (HL-145G), and luciferase control was injected in HL-deficient mice. At day 4 after infusion of 2 x 10(8) plaque forming units of rHL-AdV and rHL-145G-AdV, similar plasma concentrations were detected in postheparin plasma (HL=8.4+/-0.8 microg/mL and HL-145G=8.3+/-0.8 microg/mL). Mice expressing HL had significant reductions of cholesterol (-76%), phospholipids (PL; -68%), HDL cholesterol (-79%), apolipoprotein (apo) A-I ( 45%), and apoA-II (-59%; P<0.05 for all), whereas mice expressing HL-145G decreased their cholesterol (-49%), PL (-40%), HDL cholesterol (-42%), and apoA II (-89%; P<0.005 for all) but had no changes in apoA-I. The plasma kinetics of (125)I-labeled apoA-I HDL, (131)I-labeled apoA-II HDL, and [(3)H]cholesteryl ester (CE) HDL revealed that compared with mice expressing luciferase control (fractional catabolic rate [FCR] in d(-1): apoA-I HDL=1.3+/-0.1; apoA-II HDL=2.1+/-0; CE HDL=4.1+/-0.7), both HL and HL-145G enhanced the plasma clearance of CEs and apoA-II present in HDL (apoA-II HDL=5.6+/-0.5 and 4.4+/-0.2; CE HDL=9.3+/-0. 0 and 8.3+/-1.1, respectively), whereas the clearance of apoA-I HDL was enhanced in mice expressing HL (FCR=4.6+/-0.3) but not HL-145G (FCR=1.4+/ 0.4). These combined findings demonstrate that both lipolytic and nonlipolytic functions of HL are important for HDL metabolism in vivo. Our study provides, for the first time, in vivo evidence for a role of HL in HDL metabolism independent of lipolysis and provides new insights into the role of HL in facilitating distinct metabolic pathways involved in the catabolism of apoA-I- versus apoA-II containing HDL. PMID- 10712406 TI - Apolipoprotein A-I and A-II kinetic parameters as assessed by endogenous labeling with [(2)H(3)]leucine in middle-aged and elderly men and women. AB - The purpose of our study was to investigate high density lipoprotein (HDL) apolipoprotein (apo) A-I and apoA-II kinetics in a state of constant feeding after a primed-constant infusion of [5,5, 5-(2)H(3)]L-leucine in 32 normolipidemic older men and postmenopausal women (aged 41 to 79 years). ApoA-I and apoA-II were isolated from plasma HDL, and enrichment was determined by gas chromatography/mass spectrometry. The fractional secretion rate was obtained by using a monoexponential equation calculated with the SAAM II program (Department of Bioengineering, University of Washington, Seattle). Mean HDL cholesterol (HDL C) and total triglyceride levels were 23% higher and 27% lower, respectively, in women than in men. Mean plasma apoA-I levels were 10% greater in women than in men, whereas mean apoA-II levels were similar. HDL size, estimated by gradient sizing gels and by the HDL-C/apoA-I+apoA-II ratio, was significantly higher in women than in men. Mean apoA-I secretion rates (SRs) were similar in men and women (12.28+/-3.64 versus 11.96+/-2.92 mg/kg per day), whereas there was a trend toward a lower (-13%) apoA-I fractional catabolic rate (FCR) in women compared with men (0.199+/-0.037 versus 0. 225+/-0.062 pools per day, P=0.11). Mean apoA II SRs (2.21+/-0.57 versus 2.27+/-0.91 mg/kg per day) and FCRs (0.179+/-0.034 versus 0. 181+/-0.068 pools per day) were similar in men and women. For the group as a whole, there was an inverse association between the HDL-C/apoA-I+apoA-II ratio and apoA-I FCR and between the ratio and triglyceride levels. Plasma levels of apoA-I and apoA-II were correlated with their respective SRs but not FCRs. These data suggest a major role for apoA-I and apoA-II SRs in regulating the plasma levels of these proteins, whereas apoA-I FCR might be an important factor contributing to the differences in apoA-I levels between men and postmenopausal women. Moreover, plasma triglyceride levels are important determinants of HDL size and apoA-I catabolism. PMID- 10712407 TI - Segregation analysis of apolipoproteins A-1 and B-100 measured before and after an exercise training program: the HERITAGE Family Study. AB - Complex segregation analyses of apolipoproteins (apo) A-1 and B-100 were performed in a sample of 520 individuals from 99 white families who participated in the HERITAGE Family Study. In these sedentary families, plasma apo A-1 and B 100 concentrations were measured before and after a 20-week endurance exercise training program. Baseline apo A-1 and B-100 were adjusted for the effects of age (age-adjusted baseline apo A-1 and B-100) and for the effects of age and BMI (age BMI-adjusted baseline apo A-1 and B-100). The change in response to training was computed as a simple Delta (posttraining minus baseline) and was adjusted for age and the baseline (age-baseline-adjusted apo A-1 and B-100 responses to training). In the present study, a major gene could not be inferred for baseline apo A-1. Rather, we found a major effect along with a multifactorial effect accounting for 8% to 9% and 51% to 56% of the variance, respectively. In addition, no clear evidence supported a major-gene effect for its response to training, whereas the transmission of a major effect from parents to offspring was ambiguous, ie, genetic in nature or familial environmental in origin. The major effect accounted for 15% of the variance, with an additional 21% and 58% of the variance being accounted for by a multifactorial effect in parents and offspring, respectively. It is interesting to have obtained evidence of a putative recessive major locus for baseline apo B-100, which accounted for 50% to 56% of the variance, with an additional 25% to 29% of the variance due to a multifactorial effect. In contrast, no major effect for its response to training was identified, although a multifactorial effect was found that accounted for 27% of the variance. The novel findings arising from the present study are summarized as follows. Baseline apo A 1 and its response to training were influenced by a major effect and a multifactorial effect. Baseline apo B-100 was influenced by a putative major recessive gene with a multifactorial component, but its response to training was influenced solely by a multifactorial component in these sedentary families. PMID- 10712408 TI - Association of the C-514T polymorphism in the hepatic lipase gene with variations in lipoprotein subclass profiles: The Framingham Offspring Study. AB - Hepatic lipase is involved in the metabolism of several lipoproteins and has a key role in reverse cholesterol transport. A common C-to-T substitution at position -514 of the hepatic lipase promoter has been associated with variations in plasma high density lipoprotein cholesterol (HDL-C) levels and hepatic lipase activity. The aim of the current study was to investigate the association of this polymorphism to lipoprotein levels in a population-based sample of 1314 male and 1353 female Framingham Offspring Study participants. In men and women, carriers of the -514T allele had higher HDL-C and apolipoprotein A-I (apoAI) concentrations compared with noncarriers. The higher HDL-C levels associated with the -514T allele was due to an increase in the HDL(2)-C subfraction, and this association was stronger in women compared with men (P=0.0043 versus 0.0517). To gain further understanding about the metabolic basis of these effects, HDL and low density lipoprotein (LDL) subclass profiles were measured by using automated nuclear magnetic resonance spectroscopy and gradient gel electrophoresis, respectively. The association of the -514T allele with higher HDL-C levels seen in men and women was primarily due to significant increases in the large HDL subfractions (size range 8.8 to 13.0 nm). In contrast, there was no relationship between the hepatic lipase polymorphism at position -514 and the LDL particle size distribution after adjustment for familial relationships, age, body mass index, smoking, alcohol intake, use of beta-blockers, apoE genotype, and menopausal status and estrogen therapy in women. Moreover, multiple regression analyses suggested that the C-514T polymorphism contributed significantly to the variability of HDL particle size in men and women (P<0.04). Thus, our results show that the C-514T polymorphism in the hepatic lipase gene is associated with significant variations in the lipoprotein profile in men and women. PMID- 10712409 TI - Diabetes mellitus: subclinical cardiovascular disease and risk of incident cardiovascular disease and all-cause mortality. AB - Previously diagnosed diabetes mellitus, newly diagnosed diabetes mellitus, and impaired glucose tolerance are important determinants of the risk of clinical cardiovascular disease (CVD). We have evaluated the relation of patients with subclinical CVD, diabetes, and impaired glucose tolerance and "normal" subjects and the risk of clinical CVD in the Cardiovascular Health Study. Diabetes (1343), impaired glucose tolerance (1433), and normal (2421) were defined by World Health Organization criteria at baseline in 1989 to 1990. The average follow-up was 6.4 years (mean age 73 years). Diabetics had a higher prevalence of clinical and subclinical CVD at baseline. Compared with diabetes in the absence of subclinical disease, the presence of subclinical CVD and diabetes was associated with significant increased adjusted relative risk of death (1.5, CI 0.93 to 2.41), relative risk of incident coronary heart disease (1.99, CI 1.25 to 3.19), and incident myocardial infarction (1.93, CI 0.96 to 3.91). The risk of clinical events was greater for participants with a history of diabetes compared with newly diagnosed diabetics at baseline. Compared with nondiabetic nonhypertensive subjects without subclinical disease, patients with a combination of diabetes, hypertension, and subclinical disease had a 12-fold increased risk of stroke. Fasting blood glucose levels were a weak predictor of incident coronary heart disease as were most other risk factors. Subclinical CVD was the primary determinant of clinical CVD among diabetics in the Cardiovascular Health Study. PMID- 10712411 TI - Effects of coronary heart disease risk factors on atherosclerosis of selected regions of the aorta and right coronary artery. PDAY Research Group. Pathobiological Determinants of Atherosclerosis in Youth. AB - We examined topographic distributions of atherosclerosis and their relation to risk factors for adult coronary heart disease in right coronary arteries and abdominal aortas of more than 2000 autopsied persons 15 through 34 years of age. We digitized images of Sudan IV-stained fatty streaks and of manually outlined raised lesions and computed the percent surface area involved by each lesion in each of 6 regions of each artery. In abdominal aortas of 15- to 24-year-old persons, fatty streaks involve an elongated oval area on the dorsolateral intimal surface and another oval area in the middle third of the ventral surface. Raised lesions in 25- to 34-year-old persons involve an oval area in the distal third of the dorsolateral intimal surface. In other areas of the abdominal aortas of older persons, fatty streaks occur but raised lesions are rare. In the right coronary arteries of 15- to 24-year-old persons, fatty streaks are most frequent on the myocardial aspect of the first 2 cm. Raised lesions follow a similar pattern in 25- to 34-year-old persons. High non-HDL cholesterol and low HDL cholesterol concentrations are associated with more extensive fatty streaks and raised lesions in all regions of both arteries. Smoking is associated with more extensive fatty streaks and raised lesions of the abdominal aorta, particularly in the dorsolateral region of the distal third of the abdominal aorta. Hypertension is not associated with fatty streaks in whites or blacks but is associated with more extensive raised lesions in blacks. Risk factor effects on arterial regions that are vulnerable to lesions are approximately 25% greater than risk factor effects assessed over entire arterial segments. These risk factor effects on vulnerable sites emphasize the need for risk factor control during adolescence and young adulthood to prevent or delay the progression of atherosclerosis. PMID- 10712410 TI - LDL cholesterol as a strong predictor of coronary heart disease in diabetic individuals with insulin resistance and low LDL: The Strong Heart Study. AB - Diabetes has been shown to increase the risk of coronary heart disease in all populations studied. However, there is a lack of information on the relative importance of diabetes-associated risk factors for cardiovascular disease (CVD), especially the role of lipid levels, because low density lipoprotein (LDL) cholesterol often is not elevated in diabetic individuals. The objective of this analysis was to evaluate CVD risk factors in a large cohort of diabetic individuals and to compare the importance of dyslipidemia (ie, elevated triglycerides and low levels of high density lipoprotein [HDL] cholesterol) and LDL cholesterol in determining CVD risk in diabetic individuals. The Strong Heart Study assesses coronary heart disease and its risk factors in American Indians in Arizona, Oklahoma, and South/North Dakota. The baseline clinical examinations (July 1989 to January 1992) consisted of a personal interview, physical examination, and drawing of blood samples for 4549 study participants (2034 with diabetes), 45 to 74 years of age. Follow-up averaged 4.8 years. Fatal and nonfatal CVD events were confirmed by standardized record review. Participants with diabetes, compared with those with normal glucose tolerance, had lower LDL cholesterol levels but significantly elevated triglyceride levels, lower HDL cholesterol levels, and smaller LDL particle size. Significant independent predictors of CVD in those with diabetes included age, albuminuria, LDL cholesterol, HDL cholesterol (inverse), fibrinogen, and percent body fat (inverse). A 10-mg/dL increase in LDL cholesterol was associated with a 12% increase in CVD risk. Thus, even at concentrations well below the National Cholesterol Education Program target of 130 mg/dL, LDL cholesterol is a strong independent predictor of coronary heart disease in individuals with diabetes, even when components of diabetic dyslipidemia are present. These results support recent recommendations for aggressive control of LDL cholesterol in diabetic individuals, with a target level of <100 mg/dL. PMID- 10712412 TI - Atherosclerosis progression in LDL receptor-deficient and apolipoprotein E deficient mice is independent of genetic alterations in plasminogen activator inhibitor-1. AB - Impaired fibrinolysis has been linked to atherosclerosis in a number of experimental and clinical studies. Plasminogen activator inhibitor type 1 (PAI-1) is the primary inhibitor of plasminogen activation and has been proposed to promote atherosclerosis by facilitating fibrin deposition within developing lesions. We examined the contribution of PAI-1 to disease progression in 2 established mouse models of atherosclerosis. Mice lacking apolipoprotein E (apoE /-) and mice lacking the low density lipoprotein receptor (LDLR-/-) were crossbred with transgenic mice overexpressing PAI-1 (resulting in PAI-1 Tg(+)/apoE-/- and PAI-1 Tg(+)/LDLR-/-, respectively) or were crossbred with mice completely deficient in PAI-1 gene expression (resulting in PAI-1-/-/apoE-/- and PAI-1-/-/LDLR-/-, respectively). All animals were placed on a western diet (21% fat and 0.15% cholesterol) at 4 weeks of age and analyzed for the extent of atherosclerosis after an additional 6, 15, or 30 weeks. Intimal and medial areas were determined by computer-assisted morphometric analysis of standardized microscopic sections from the base of the aorta. Atherosclerotic lesions were also characterized by histochemical analyses with the use of markers for smooth muscle cells, macrophages, and fibrin deposition. Typical atherosclerotic lesions were observed in all experimental animals, with greater severity at the later time points and generally more extensive lesions in apoE-/- than in comparable LDLR-/- mice. No significant differences in lesion size or histological appearance were observed among PAI-1-/-, PAI-1 Tg(+), or PAI-1 wild-type mice at any of the time points on either the apoE-/- or LDLR-/- genetic background. We conclude that genetic modification of PAI-1 expression does not significantly alter the progression of atherosclerosis in either of these well-established mouse models. These results suggest that fibrinolytic balance (as well as the potential contribution of PAI-1 to the regulation of cell migration) plays only a limited role in the pathogenesis of the simple atherosclerotic lesions observed in the mouse. PMID- 10712413 TI - Local plasminogen activator inhibitor type 1 overexpression in rat carotid artery enhances thrombosis and endothelial regeneration while inhibiting intimal thickening. AB - Elevated levels of plasminogen activator inhibitor type 1 (PAI-1) are found in advanced atherosclerotic plaque compared with normal vessel and may contribute to plaque progression and complications associated with plaque rupture. Increased expression of PAI-1 probably contributes to the thrombotic properties of advanced atherosclerotic plaque by impeding plasmin generation and degradation of fibrin. To test this hypothesis, we have deliberately created synthetic neointimas by seeding onto the denuded luminal surface of rat carotid arteries smooth muscle cells transduced with replication-defective retrovirus encoding rat PAI-1. This cell-based gene transfer method results in stable, long-term, and localized gene expression. PAI-1 overexpression increases mural thrombus accumulation at 4 days but decreases neointimal area by 30% and 25% at 1 week and 2 weeks, respectively. PAI-1 overexpression accelerates reendothelialization of injured arteries compared with control arteries at 1 week, 2 weeks, and 1 month. PAI-1 overexpression does not alter matrix accumulation at 1 week. Increased PAI-1 expression in the rat carotid artery enhances thrombosis and endothelial regeneration while inhibiting intimal thickening. These results suggest that PAI 1 could play a direct role in the development of advanced atherosclerotic plaque and in the repair of the diseased vessel after fibrous cap disruption. PMID- 10712414 TI - In vivo dynamic real-time monitoring and quantification of platelet-thrombus formation: use of a local isotope detector. AB - Current methods for monitoring thrombosis and thrombus growth are invasive and provide only single-time-point data. Animal models rely mainly on flow changes as a surrogate of thrombus formation. Our aim was to validate a unique potentially noninvasive system to detect and quantify dynamic thrombus formation in vivo by using a porcine model of carotid artery injury. Thrombus growth was monitored by deposition of autologous (111)In-labeled platelet activity over the injured artery by use of miniaturized gamma detectors and Doppler blood flow. Counts were recorded at 2-minute intervals for 2 hours. The technique was validated by comparing standard antithrombotic agents against controls. Platelet recruitment was detected before significant change in flow. Thrombus formation, calculated as the area under the curve (platelets x minutes x 10(6)), was greatest for control animals (11.7+/-1.28), followed by animals treated with aspirin (6.13+/-0.91, P<0.05), heparin (2.45+/-0.34, P<0.05), and hirudin (0.2+/-0.01, P<0.01 compared with heparin). The rate of platelet deposition was assessed as the slope of the curve in the first 30 minutes (platelets x 10(6) per minute) for the following treatment groups of animals: control, 3.53+/-0.34; aspirin, 1.67+/-0. 34 (P<0.01); heparin, 1.55+/-0.3 (P<0.01); and hirudin, 0.25+/-0.03 (P<0.001). There was no statistical difference between heparin and aspirin treatments. Change in flow was assessed as reduction from baseline: control, >99+/-0.34%; aspirin, 39+/ 9.1%; heparin, 36+/-12. 5%; and hirudin, 17+/-5.4%. There was no statistical difference between the aspirin- and heparin-treated groups. Morphometric analysis revealed >99+/-0.63% occlusion of the luminal area with thrombus for the control group, 43+/-14.3% for the aspirin-treated group, 30+/-5.6% for the heparin treated group, and <10+/-1.8% for the hirudin-treated group. Assessment of platelet-thrombus formation with this technique was more sensitive than change in flow in determining antithrombotic efficacy, and thrombus formation was detected earlier. This study validates a new quantitative, sensitive, potentially noninvasive, portable, in vivo monitoring of dynamic thrombus growth, which appears applicable to phase II studies in humans. PMID- 10712415 TI - P2Y receptor regulation of PAI-1 expression in vascular smooth muscle cells. AB - P2Y-type purine and pyrimidine nucleotide receptors play important roles in the regulation of vascular hemostasis. In this article, the regulation of plasminogen activator inhibitor-1 (PAI-1) expression in rat aortic smooth muscle cells (RASMCs) by adenine and uridine nucleotides was examined and compared. Northern analysis revealed that RASMCs express multiple P2Y receptor subtypes, including P2Y(1), P2Y(2), and P2Y(6). Treatment of RASMCs with UTP increased PAI-1 mRNA expression and extracellular PAI-1 protein levels by 21-fold (P<0.001) and 7-fold (P<0.001), respectively. The ED(50) for the effect of UTP on PAI-1 expression was approximately 1 micromol/L, and its maximal effect occurred at 3 hours. UDP stimulated a 5-fold increase (P<0.005) in PAI-1 expression. In contrast to these potent stimulatory effects of uridine nucleotides, ATP and 2-methylthioadenosine triphosphate (2-MeSATP) caused a small and transient increase in PAI-1 mRNA at 1 hour, followed by a rapid decrease to baseline levels. ADP produced only an inhibitory effect, reducing PAI-1 mRNA levels by 63% (P<0.05) at 3 hours. The relative nucleotide potency in stimulating PAI-1 expression is UTP>UDP>ATP=2 MeSATP, consistent with a predominant role of the P2Y(6) receptor. Further studies revealed that exposure of RASMCs to either ATP or ADP for 3 hours inhibited both UTP- and angiotensin II-stimulated PAI-1 expression by up to 90% (P<0.001). Thus, ATP induced a small and transient upregulation of PAI-1 that was followed by a strong inhibition of PAI-1 expression. These results show that extracellular adenine and uridine nucleotides exert potent and opposing effects on vascular PAI-1 expression. PMID- 10712416 TI - Reversible regulation of tissue factor-induced coagulation by glycosyl phosphatidylinositol-anchored tissue factor pathway inhibitor. AB - Endothelial and tumor cells synthesize tissue factor pathway inhibitor (TFPI-1), which regulates tissue factor (TF) function by TF. VIIa. Xa. TFPI-1 quaternary complex formation (where VIIa and Xa are coagulation factors) and by translocation of these complexes into glycosphingolipid-rich microdomains of the cell membrane. Recombinant TFPI-1 added exogenously to cells is targeted to a degradation pathway. This study analyzes whether quaternary complex formation with endogenous TFPI-1 results also in internalization and degradation. We demonstrate that endogenous TFPI-1 and recombinant TFPI-1 differ in their distribution on the cell surface. Recombinant TFPI-1 is found in phospholipid- and glycosphingolipid-rich membrane domains, whereas endogenous TFPI-1 preferentially localizes to glycosphingolipid-rich microdomains. On quaternary complex formation, endogenous TFPI-1 remains protease sensitive and accessible for antibodies on intact cells, demonstrating that it is not appreciably internalized. Rather, regulation of TF by TFPI-1 is restored within 12 hours, consistent with dissociation of quaternary complexes on the cell surface. Endogenous TFPI-1 can be released from the cell surface by phospholipase treatment, indicating that TFPI-1 either is a glycosyl phosphatidylinositol (GPI) anchored protein or binds to a GPI-linked receptor. We demonstrate that expression of a recombinant GPI-anchored form of TFPI-1 targets TF. VIIa complexes to glycosphingolipid-rich membrane fractions. Thus, GPI anchoring of TFPI-1 is sufficient for regulation of TF. VIIa complex function by a pathway of reversible inhibition rather than internalization and degradation. PMID- 10712417 TI - Regulated von Willebrand factor secretion is associated with agonist-specific patterns of cytoskeletal remodeling in cultured endothelial cells. AB - von Willebrand factor (vWF), an adhesive glycoprotein involved in primary hemostasis, is stored and released from endothelial secretory granules called Weibel-Palade bodies. Regulated secretion occurs in reaction either to [Ca(2+)](i)-raising agents (histamine or thrombin) or to cAMP-raising agents (epinephrine, adenosine, or forskolin). We investigated the pattern of release and the cytoskeletal requirements for secretion in response to these 2 classes of agonists. Secretion induced by [Ca(2+)](i)-raising agents involves peripheral and central granules and is inhibited by colchicine-induced microtubule disruption. It is accompanied by Rho-dependent stress fiber formation and cell retraction. Secretion and remodeling occur in the same individual cells. However, secretion is potentiated by cytochalasin E and C3 toxin, indicating that stress fiber formation antagonizes vWF secretion. In contrast, vWF secretion induced by cAMP raising agents involves the release of only peripheral granules (implying less vWF release on a per cell basis) and is not inhibited by microtubule disruption. cAMP-mediated secretion is accompanied by disruption of stress fibers, strengthening of the cortical actin rim, and preservation of cell-cell contacts. It is unaffected by cytochalasins or C3 toxin. In contrast to [Ca(2+)](i)-raising agents, cAMP-raising agents induce secretion without cell retraction/intercellular gap formation. Thus, they are likely to play a physiological role in the regulation of endothelial vWF secretion and, therefore, of plasma vWF levels. PMID- 10712419 TI - ATVB online only : march 2000 PMID- 10712418 TI - Polymorphisms in the 5' regulatory region of the tissue factor gene and the risk of myocardial infarction and venous thromboembolism: the ECTIM and PATHROS studies. Etude Cas-Temoins de l'Infarctus du Myocarde. Paris Thrombosis case control Study. AB - Tissue factor (TF) is a transmembrane protein considered to be responsible for the initiation of coagulation. TF gene expression may be induced in monocytes and endothelial cells and is present in atherosclerotic plaque to initiate thrombus formation. To investigate whether individual differences in TF gene expression could predispose subjects to thrombosis, we sequenced the 5' domain of the gene up to nucleotide 2732 and found 6 different polymorphisms: 4 of them were completely concordant and defined 2 haplotypes with similar frequencies, designated as 1208 D and 1208 I. Genotyping of patients with myocardial infarction in a case-control study involving 2354 subjects showed no association between the polymorphisms and nonfatal coronary thrombosis. In another study involving 255 patients with venous thromboembolism and 1204 controls, allele D was less common in the cases (P=0.022). The odds ratio associated with the presence of at least 1 D allele was 0.72 (P=0. 031). Comparison of subgroups of control subjects who were homozygous for the D or I allele demonstrated a lower plasma TF concentration in DD homozygotes. These results indicate that the TF gene promoter exists in 2 major forms differing at 4 sites. The 1208 D haplotype is not associated with coronary thrombosis but is associated with reduced plasma TF levels and a lower risk of venous thrombosis. PMID- 10712420 TI - Ras/Rac-Dependent activation of p38 mitogen-activated protein kinases in smooth muscle cells stimulated by cyclic strain stress AB - p38, a subfamily of the mitogen-activated protein kinases (MAPKs), is a crucial signal transducer between a variety of extracellular stimuli and gene expression in mammalian cells. This kinase is activated in cultured cells stimulated by heat shock, osmotic stress, and proinflammatory cytokines, but a similar activation of p38 MAPKs in vascular smooth muscle cells (SMCs) stimulated by mechanical stress has yet to be studied. We studied signal pathways leading to time- and strength dependent p38 activation in rat SMCs in response to cyclic strain stress. p38 phosphorylation in stressed SMCs showed maximal activation at 10 minutes. This activation was significantly inhibited by pretreatment of the SMCs with pertussis toxin, a G-protein antagonist, and enhanced by treatment with suramin, a growth factor receptor antagonist, but opposite effects in the activation of extracellular signal-regulated kinases stimulated by mechanical forces were found. p38 activation was markedly reduced in stressed SMCs after protein kinase C depletion. Interestingly, SMC lines stably expressing dominant-negative ras (ras N17) or rac1 (rac1 N17) almost abolished p38 phosphorylation induced by cyclic strain stress. When p38 activation was inhibited by the specific inhibitor SB 202190, SMC migration, determined in a Boyden chamber in response to stimulation with platelet-derived growth factor-BB, and SMC proliferation, stimulated by cyclic strain stress, were abrogated. Thus, we provide the first evidence that cyclic strain stress rapidly activates p38 MAPKs via activation of protein kinase C ras/rac signal pathways, suggesting that p38 MAPKs are important signal transducers mediating the mechanical stress-induced cell responses essential for SMC migration and proliferation. PMID- 10712422 TI - Egr-1: is it always immediate and early? PMID- 10712423 TI - RANKing the importance of measles virus in Paget's disease. PMID- 10712424 TI - Hypoxia and human placental development. PMID- 10712425 TI - CADASIL: Notch signaling defect or protein accumulation problem? PMID- 10712426 TI - The missing link in lysosomal enzyme targeting. PMID- 10712427 TI - Normal physiology and HIV pathophysiology of human T-cell dynamics. PMID- 10712428 TI - Hepatoma-derived growth factor stimulates smooth muscle cell growth and is expressed in vascular development. AB - Hepatoma-derived growth factor (HDGF) is the first member identified of a new family of secreted heparin-binding growth factors highly expressed in the fetal aorta. The biologic role of HDGF in vascular growth is unknown. Here, we demonstrate that HDGF mRNA is expressed in smooth muscle cells (SMCs), most prominently in proliferating SMCs, 8-24 hours after serum stimulation. Exogenous HDGF and endogenous overexpression of HDGF stimulated a significant increase in SMC number and DNA synthesis. Rat aortic SMCs transfected with a hemagglutinin epitope-tagged rat HDGF cDNA contain HA-HDGF in their nuclei during S-phase. We also detected native HDGF in nuclei of cultured SMCs, of SMCs and endothelial cells from 19-day fetal (but not in the adult) rat aorta, of SMCs proximal to abdominal aortic constriction in adult rats, and of SMCs in the neointima formed after endothelial denudation of the rat common carotid artery. Moreover, HDGF colocalizes with the proliferating cell nuclear antigen (PCNA) in SMCs in human atherosclerotic carotid arteries, suggesting that HDGF helps regulate SMC growth during development and in response to vascular injury. PMID- 10712429 TI - Hypoxia-inducible factor-1 mediates the biological effects of oxygen on human trophoblast differentiation through TGFbeta(3) AB - During early pregnancy, placentation occurs in a relatively hypoxic environment that is essential for appropriate embryonic development. Intervillous blood flow increases around 10 to 12 weeks of gestation and results in exposure of trophoblast cells to increased oxygen tension. Before this time, low oxygen appears to prevent trophoblast differentiation toward an invasive phenotype. Using human villous explants of 5-8 weeks' gestation, we found that low oxygen tension triggered trophoblast proliferation, fibronectin synthesis, alpha(5) integrin expression, and gelatinase A activity. These biochemical markers were barely detectable under oxic conditions. We therefore examined the placental expression of hypoxia-inducible factor-1 (HIF-1), a master regulator of oxygen homeostasis, and determined that expression of HIF-1alpha subunit during the first trimester of gestation parallels that of TGFbeta(3), an inhibitor of extravillous trophoblast differentiation. Expression of both molecules is high in early pregnancy and falls around 9 weeks of gestation, when placental pO(2) levels are believed to increase. Increasing oxygen tension induced a similar decrease in expression in cultured explants. Moreover, antisense inhibition of HIF-1alpha expression in hypoxic explants inhibited expression of TGFbeta(3), arrested cell proliferation, decreased alpha(5) expression and gelatinase A activity, and triggered biochemical markers of an invasive trophoblast phenotype such as alpha(1) integrin and gelatinase B expression. These data suggest that the oxygen-regulated early events of trophoblast differentiation are in part mediated by TGFbeta(3) through HIF-1 transcription factors. PMID- 10712430 TI - Targeting the urokinase plasminogen activator receptor enhances gene transfer to human airway epithelia. AB - Developing gene therapy for cystic fibrosis has been hindered by limited binding and endocytosis of vectors by human airway epithelia. Here we show that the apical membrane of airway epithelia express the urokinase plasminogen activator receptor (uPAR). Urokinase plasminogen activator (uPA), or a 7-residue peptide derived from this protein (u7-peptide), bound the receptor and stimulated apical endocytosis. Both ligands enhanced gene transfer by nonspecifically bound adenovirus and adeno-associated virus vectors and by a modified adenovirus vector that had been coupled to the u7-peptide. These data provide the first evidence that targeting an apical receptor can circumvent the two most important barriers to gene transfer in airway epithelia. Thus, the uPA/uPAR system may offer significant advantages for delivering genes and other pharmaceuticals to airway epithelia. PMID- 10712431 TI - The ectodomain of the Notch3 receptor accumulates within the cerebrovasculature of CADASIL patients. AB - Mutations in Notch3 cause CADASIL (cerebral autosomal dominant adult onset arteriopathy), which leads to stroke and dementia in humans. CADASIL arteriopathy is characterized by major alterations of vascular smooth muscle cells and the presence of specific granular osmiophilic deposits. Patients carry highly stereotyped mutations that lead to an odd number of cysteine residues within EGF like repeats of the Notch3 receptor extracellular domain. Such mutations may alter the processing or the trafficking of this receptor, or may favor its oligomerization. In this study, we examined the Notch3 expression pattern in normal tissues and investigated the consequences of mutations on Notch3 expression in transfected cells and CADASIL brains. In normal tissues, Notch3 expression is restricted to vascular smooth muscle cells. Notch3 undergoes a proteolytic cleavage leading to a 210-kDa extracellular fragment and a 97-kDa intracellular fragment. In CADASIL brains, we found evidence of a dramatic and selective accumulation of the 210-kDa Notch3 cleavage product. Notch3 accumulates at the cytoplasmic membrane of vascular smooth muscle cells, in close vicinity to but not within the granular osmiophilic material. These results strongly suggest that CADASIL mutations specifically impair the clearance of the Notch3 ectodomain, but not the cytosolic domain, from the cell surface. PMID- 10712432 TI - Osteoclasts expressing the measles virus nucleocapsid gene display a pagetic phenotype. AB - Osteoclasts (OCLs) in Paget's disease are markedly increased in number and size, have increased numbers of nuclei per multinucleated cell, and demonstrate increased resorption capacity and increased sensitivity to 1,25-(OH)(2)D(3), the active form of vitamin D. These cells also contain nuclear inclusions, reminiscent of those seen in paramyxovirus-infected cells, which cross-react with antibodies to measles virus nucleocapsid (MVNP) antigen. To elucidate the role of MV in the abnormal OCL phenotype of Paget's disease, we transduced normal OCL precursors with retroviral vectors expressing MVNP and the MV matrix (MVM) genes. The transduced cells were then cultured with 1,25-(OH)(2)D(3) for14 or 21 days to induce formation of OCL-like multinucleated cells. The MVNP-transduced cells formed increased numbers of multinucleated cells, which contained many more nuclei and had increased resorption capacity compared with multinucleated cells derived from empty vector-transduced (EV-transduced) and MVM-transduced or normal bone marrow cells. Furthermore, MVNP-transduced cells showed increased sensitivity to 1, 25-(OH)(2)D(3), and formed OCLs at concentrations of 1, 25 (OH)(2)D(3) that were 1 log lower than that required for normal, EV-transduced, or MVM-transduced cells. These results demonstrate that expression of the MVNP gene in normal OCL precursors stimulates OCL formation and induces OCLs that express a phenotype similar to that of pagetic OCLs. These results support a potential pathophysiologic role for MV infection in the abnormal OCL activity and morphology that are characteristic of pagetic OCLs. PMID- 10712433 TI - Paternal versus maternal transmission of a stimulatory G-protein alpha subunit knockout produces opposite effects on energy metabolism. AB - Heterozygous disruption of Gnas, the gene encoding the stimulatory G-protein alpha subunit (G(s)alpha), leads to distinct phenotypes depending on whether the maternal (m-/+) or paternal (+/p-) allele is disrupted. G(s)alpha is imprinted, with the maternal allele preferentially expressed in adipose tissue. Hence, expression is decreased in m-/+ mice but normal in +/p- mice. M-/+ mice become obese, with increased lipid per cell in white and brown adipose tissue, whereas +/p- mice are thin, with decreased lipid in adipose tissue. These effects are not due to abnormalities in thyroid hormone status, food intake, or leptin secretion. +/p- mice are hypermetabolic at both ambient temperature (21 degrees C) and thermoneutrality (30 degrees C). In contrast, m-/+ mice are hypometabolic at ambient temperature and eumetabolic at thermoneutrality M-/+ and wild-type mice have similar dose-response curves for metabolic response to a beta(3)-adrenergic agonist, CL316243, indicating normal sensitivity of adipose tissue to sympathetic stimulation. Measurement of urinary catecholamines suggests that +/p- and m-/+ mice have increased and decreased activation of the sympathetic nervous system, respectively. This is to our knowledge the first animal model in which a single genetic defect leads to opposite effects on energy metabolism depending on parental inheritance. This probably results from deficiency of maternal- and paternal-specific Gnas gene products, respectively. PMID- 10712434 TI - Use of autoantigen-knockout mice in developing an active autoimmune disease model for pemphigus. AB - The development of experimental models of active autoimmune diseases can be difficult due to tolerance of autoantigens, but knockout mice, which fail to acquire tolerance to the defective gene product, provide a useful tool for this purpose. Using knockout mice lacking desmoglein 3 (Dsg3), the target antigen of pemphigus vulgaris (PV), we have generated an active disease model for this autoantibody-mediated disease. Dsg3(-/-) mice, but not Dsg3(+/-) littermates, produced anti-Dsg3 IgG that binds native Dsg3, when immunized with recombinant mouse Dsg3. Splenocytes from the immunized Dsg3(-/-) mice were then adoptively transferred into Rag-2(-/-) immunodeficient mice expressing Dsg3. Anti-Dsg3 IgG was stably produced in the recipient mice for more than 6 months without further boosting. This IgG bound to Dsg3 in vivo and disrupted the cell-cell adhesion of keratinocytes. Consequently, the recipient mice developed erosions in their oral mucous membranes with typical histologic findings of PV. In addition, the recipient mice showed telogen hair loss, as found in Dsg3(-/-) mice. Collectively, the recipient mice developed the phenotype of PV due to the pathogenic anti-Dsg3 IgG. This model will be valuable for developing novel therapeutic strategies. Furthermore, our approach can be applied broadly for the development of various autoimmune disease models. PMID- 10712435 TI - Elevated luteinizing hormone induces expression of its receptor and promotes steroidogenesis in the adrenal cortex. AB - Transgenic (TG) female mice expressing bLHbeta-CTP (a chimeric protein derived from the beta-subunit of bovine luteinizing hormone [LH] and a fragment of the beta-subunit of human chorionic gonadotropin [hCG]) exhibit elevated serum LH, infertility, polycystic ovaries, and ovarian tumors. In humans, increased LH secretion also occurs in infertility and polycystic ovarian syndrome, often concomitant with adrenocortical dysfunction. We therefore investigated adrenal function in LH overexpressing bLHbeta-CTP female mice. The size of their adrenals was increased by 80% with histological signs of cortical stimulation. Furthermore, adrenal steroid production was increased, with up to 14-fold elevated serum corticosterone. Primary adrenal cells from TG and control females responded similarly to ACTH stimulation, but, surprisingly, the TG adrenals responded to hCG with significantly increased cAMP, progesterone, and corticosterone production. LH receptor (LHR) expression and activity were also elevated in adrenals from female TG mice, but gonadectomized TG females showed no increase in corticosterone, suggesting that the dysfunctional ovaries of the intact TG females promote adrenocortical hyperfunction. We suggest that, in intact TG females, enhanced ovarian estrogen synthesis causes increased secretion of prolactin (PRL), which elevates LHR expression. Chronically elevated serum LH, augmented by enhanced PRL production, induces functional LHR expression in mouse adrenal cortex, leading to elevated, LH-dependent, corticosterone production. Thus, besides polycystic ovaries, the bLHbeta-CTP mice provide a useful model for studying human disorders related to elevated LH secretion and adrenocortical hyperfunction. PMID- 10712436 TI - CD28-B7 blockade prevents the development of experimental autoimmune glomerulonephritis. AB - Experimental autoimmune glomerulonephritis (EAG), an animal model of Goodpasture's disease, can be induced in Wistar Kyoto (WKY) rats by a single injection of rat glomerular basement membrane (GBM) in adjuvant. EAG is characterized by circulating and deposited anti-GBM antibodies, accompanied by focal necrotizing glomerulonephritis with crescent formation. The role of T cells in the pathogenesis of EAG remains unclear. T-cell costimulation is provided by ligation of CD28 with either B7.1 (CD80) or B7.2 (CD86) on antigen-presenting cells, and can be inhibited by a soluble form of CTLA4 (CTLA4-Ig) that binds to both B7.1 and B7.2. We examined the effect of CD28-B7 blockade on the development of EAG using native CTLA4-Ig or mutant CTLA4-Ig (Y100F-Ig), which selectively blocks B7.1. Native CTLA4-Ig treatment ameliorated EAG by several measures, including the levels of circulating anti-GBM antibodies, albuminuria, the deposition of IgG and fibrin in the glomeruli, the severity of glomerular abnormalities, and the numbers of infiltrating T cells and macrophages. Y100F-Ig resulted in a similar reduction in the severity of nephritis, but produced no overall reduction in circulating anti-GBM antibodies, although there was a reduction in IgG2a antibodies. We concluded that CD28-B7 blockade reduced autoantibody production and cellular infiltration of glomeruli, and prevented target organ injury. Our results suggest a key role for B7. 1 in costimulation of Th1-like autoimmune responses in the rat, and show that glomerular injury in EAG is largely dependent on cell-mediated mechanisms. PMID- 10712437 TI - High-level expression of Egr-1 and Egr-1-inducible genes in mouse and human atherosclerosis. AB - To understand the mRNA transcript profile in the human atherosclerotic lesion, RNA was prepared from the fibrous cap versus adjacent media of 13 patients undergoing carotid endarterectomy. cDNA expression arrays bearing 588 known genes indicated that lesions express unexpectedly high levels of the early growth response gene, Egr-1 (NGFI-A), a zinc-finger transcription factor that modulates a cluster of stress-responsive genes including PDGF and TGF-beta. Expression of Egr-1 was an average of 5-fold higher in the lesion than in the adjacent media, a result confirmed by RT-PCR, and many Egr-1-inducible genes were also strongly elevated in the lesion. Time-course analyses revealed that Egr-1 was not induced ex vivo. Immunocytochemistry indicated that Egr-1 was expressed prominently in the smooth muscle-actin positive cells, particularly in areas of macrophage infiltration, and in other cell types, including endothelial cells. Induction of atherosclerosis in LDL receptor-null mice by feeding them a high-fat diet resulted in a progressive increase in Egr-1 expression in the aorta. Thus, induction of Egr-1 by atherogenic factors may be a key step in coordinating the cellular events that result in vascular lesions. PMID- 10712438 TI - Gene dosage affects the cardiac and brain phenotype in nonmuscle myosin II-B depleted mice. AB - Complete ablation of nonmuscle myosin heavy chain II-B (NMHC-B) in mice resulted in cardiac and brain defects that were lethal during embryonic development or on the day of birth. In this paper, we report on the generation of mice with decreased amounts of NMHC-B. First, we generated B(DeltaI)/B(DeltaI) mice by replacing a neural-specific alternative exon with the PGK-Neo cassette. This resulted in decreased amounts of NMHC-B in all tissues, including a decrease of 88% in the heart and 65% in the brain compared with B(+)/B(+) tissues. B(DeltaI)/B(DeltaI) mice developed cardiac myocyte hypertrophy between 7 months and 11 months of age, at which time they reexpressed the cardiac beta-MHC. Serial sections of B(DeltaI)/B(DeltaI) brains showed abnormalities in neural cell migration and adhesion in the ventricular wall. Crossing B(DeltaI)/B(DeltaI) with B(+)/B(-) mice generated B(DeltaI)/B(-) mice, which showed a further decrease of approximately 55% in NMHC-B in the heart and brain compared with B(DeltaI)/B(DeltaI) mice. Five of 8 B(DeltaI)/B(-) mice were born with a membranous ventricular septal defect. Moreover, 5 of 5 B(DeltaI)/B(-) mice developed myocyte hypertrophy by 1 month; B(DeltaI)/B(-) mice also reexpressed the cardiac beta-MHC. More than 60% of B(DeltaI)/B(-) mice developed overt hydrocephalus and showed more severe defects in neural cell migration and adhesion than did B(DeltaI)/B(DeltaI) mice. These data on B(DeltaI)/B(DeltaI) and B(DeltaI)/B(-) mice demonstrate a gene dosage effect of the amount of NMHC-B on the severity and time of onset of the defects in the heart and brain. PMID- 10712439 TI - Molecular basis of variant pseudo-hurler polydystrophy (mucolipidosis IIIC) AB - Mucolipidosis IIIC, or variant pseudo-Hurler polydystrophy, is an autosomal recessive disease of lysosomal hydrolase trafficking. Unlike the related diseases, mucolipidosis II and IIIA, the enzyme affected in mucolipidosis IIIC (N Acetylglucosamine-1-phosphotransferase [GlcNAc-phosphotransferase]) retains full transferase activity on synthetic substrates but lacks activity on lysosomal hydrolases. Bovine GlcNAc-phosphotransferase has recently been isolated as a multisubunit enzyme with the subunit structure alpha(2)beta(2)gamma(2). We cloned the cDNA for the human gamma-subunit and localized its gene to chromosome 16p. We also showed, in a large multiplex Druze family that exhibits this disorder, that MLIIIC also maps to this chromosomal region. Sequence analysis of the gamma subunit cDNA in patients from 3 families identified a frameshift mutation, in codon 167 of the gamma subunit, that segregated with the disease, indicating MLIIIC results from mutations in the phosphotransferase gamma-subunit gene. This is to our knowledge the first description of the molecular basis for a human mucolipidosis and suggests that the gamma subunit functions in lysosomal hydrolase recognition. PMID- 10712440 TI - The CD44-initiated pathway of T-cell extravasation uses VLA-4 but not LFA-1 for firm adhesion. AB - Leukocytes extravasate from the blood in response to physiologic or pathologic demands by means of complementary ligand interactions between leukocytes and endothelial cells. The multistep model of leukocyte extravasation involves an initial transient interaction ("rolling" adhesion), followed by secondary (firm) adhesion. We recently showed that binding of CD44 on activated T lymphocytes to endothelial hyaluronan (HA) mediates a primary adhesive interaction under shear stress, permitting extravasation at sites of inflammation. The mechanism for subsequent firm adhesion has not been elucidated. Here we demonstrate that the integrin VLA-4 is used in secondary adhesion after CD44-mediated primary adhesion of human and mouse T cells in vitro, and by mouse T cells in an in vivo model. We show that clonal cell lines and polyclonally activated normal T cells roll under physiologic shear forces on hyaluronate and require VCAM-1, but not ICAM-1, as ligand for subsequent firm adhesion. This firm adhesion is also VLA-4 dependent, as shown by antibody inhibition. Moreover, in vivo short-term homing experiments in a model dependent on CD44 and HA demonstrate that superantigen-activated T cells require VLA-4, but not LFA-1, for entry into an inflamed peritoneal site. Thus, extravasation of activated T cells initiated by CD44 binding to HA depends upon VLA-4-mediated firm adhesion, which may explain the frequent association of these adhesion receptors with diverse chronic inflammatory processes. PMID- 10712441 TI - Factors influencing T-cell turnover in HIV-1-seropositive patients. AB - HIV-1 disease is associated with pathological effects on T-cell production, destruction, and distribution. Using the deuterated (2H) glucose method for endogenous labeling, we have analyzed host factors that influence T-cell turnover in HIV-1-uninfected and -infected humans. In untreated HIV-1 disease, the average half life of circulating T cells was diminished without compensatory increases in cell production. Within 12 weeks of the initiation of highly active antiretroviral therapy (HAART), the absolute production rates of circulating T cells increased, and normal half-lives and production rates were restored by 12 36 months. Interpatient heterogeneity in the absolute degree of turnover correlated with the relative proportion of naive- and memory/effector-phenotype T cells in each of the CD4+ and CD8+ populations. The half-lives of naive-phenotype T cells ranged from 116-365 days (fractional replacement rates of 0.19-0.60% per day), whereas memory/effector-phenotype T cells persisted with half-lives from 22 79 days (fractional replacement rates of 0.87-3.14% per day). Naive-phenotype T cells were more abundant, and the half-life of total T cells was prolonged in individuals with abundant thymic tissue, as assessed by computed tomography. Such interpatient variation in T-cell kinetics may be reflective of differences in functional immune reconstitution after treatment for HIV-1 disease. PMID- 10712442 TI - CA3-released entorhinal seizures disclose dentate gyrus epileptogenicity and unmask a temporoammonic pathway. AB - We have investigated the propagation of epileptiform discharges induced by 4 aminopyridine (4-AP, 50 microM) in adult mouse hippocampus-entorhinal cortex slices, before and after Schaffer collateral cut. 4-AP application induced 1) ictal epileptiform activity that disappeared over time and 2) interictal epileptiform discharges, which continued throughout the experiment. Using simultaneous field potential and [K(+)](o) recordings, we found that entorhinal and dentate ictal epileptiform discharges were accompanied by comparable elevations in [K(+)](o) (up to 12 mM from a baseline value of 3.2 mM), whereas smaller rises in [K(+)](o) (up to 6 mM) were associated with ictal activity in CA3. Cutting the Schaffer collaterals disclosed the occurrence of ictal discharges that were associated with larger rises in [K(+)](o) as compared with the intact slice. Further lesion of the perforant path blocked ictal activity and the associated [K(+)](o) increases in the dentate gyrus, indicating synaptic propagation to this area. Time delay measurements demonstrated that ictal epileptiform activity in the intact hippocampal-entorhinal cortex slice propagated via the trisynaptic path. However, after Schaffer collateral cut, ictal discharges continued to occur in CA1 and subiculum and spread to these areas directly from the entorhinal cortex. Thus our data indicate that the increased epileptogenicity of the dentate gyrus (a prominent feature of temporal lobe epilepsy as well), may depend on perforant path propagation of entorhinal ictal discharges, irrespective of mossy fiber reorganization. Moreover, hippocampal neuronal damage that is acutely mimicked in our model by Schaffer collateral cut, may contribute to "short-circuit" propagation of activity by pathways that are masked when the hippocampus is intact. PMID- 10712443 TI - Three-dimensional kinematics of ocular drift in humans with cerebellar atrophy. AB - One of the signs of the cerebellar ocular motor syndrome is the inability to maintain horizontal and vertical fixation. Typically, in the presence of cerebellar atrophy, the eyes show horizontal gaze-evoked and vertical downbeat nystagmus. We investigated whether or not the cerebellar ocular motor syndrome also includes a torsional drift and, specifically, if it is independent from the drift in the horizontal-vertical plane. The existence of such a torsional drift would suggest that the cerebellum is critically involved in maintaining the eyes in Listing's plane. Eighteen patients with cerebellar atrophy (diagnosis confirmed by magnetic resonance imaging) were tested and compared with a group of normal subjects. Three-dimensional eye movements (horizontal, vertical, and torsional) during attempted fixations of targets at different horizontal and vertical eccentricities were recorded by dual search coils in a three-field magnetic frame. The overall ocular drift was composed of an upward drift that increased during lateral gaze, a horizontal centripetal drift that appeared during lateral gaze, and a torsional drift that depended on horizontal eye position. The vertical drift consisted of two subcomponents: a vertical gaze evoked drift and a constant vertical velocity bias. The increase of upward drift velocity with eccentric horizontal gaze was caused by an increase of the vertical velocity bias; this component did not comply with Listing's law. The horizontal eye-position-dependent torsional drift was intorsional in abduction and extorsional in adduction, which led to an additional violation of Listing's law. The existence of torsional drift that is eye-position-dependent suggests that the cerebellum is critically involved in the implementation of Listing's law, perhaps by mapping a tonic torsional signal that depends on the direction of the line of sight. The magnitude of this signal might reflect the difference in torsional eye position between the torsional resting position determined by the mechanics of the eye plant and the torsional position required by Listing's law. PMID- 10712444 TI - Activity-dependent activation of presynaptic metabotropic glutamate receptors in locus coeruleus. AB - Synaptic activation of metabotropic glutamate receptors (mGluRs) in the locus coeruleus (LC) was investigated in adult rat brain slice preparations. Evoked excitatory postsynaptic potentials (EPSPs) resulting from stimulation of LC afferents were measured with current clamp from intracellularly recorded LC neurons. In this preparation, mGluR agonists (+/-)-1-aminocyclopentane-trans-1, 3 dicarboxylic acid (t-ACPD) and L(+)-2-amino-4-phosphonobutyric acid (L-AP4) activate distinct presynaptic mGluRs, resulting in an inhibition of EPSPs. When two stimuli were applied to afferents at intervals >200 ms, the amplitude of the second [test (T)] EPSP was identical in amplitude to the first [control(C)]. However, when a stimulation volley was delivered before T, the amplitude of the latter EPSP was consistently smaller than C. The activity-dependent depression (ADD) was dependent on the frequency and duration of the train and the interval between the train and T. ADD was potentiated in the presence of an excitatory amino acid (EAA) uptake inhibitor L-trans-pyrrolidine-2,4-dicarboxylic acid (t PDC, 100 microM), changing the T/C ratio from 0.84 +/- 0.05 (mean +/- SE) in control to 0.69 +/- 0.04 in t-PDC (n = 9). In the presence of t-PDC, the depolarizing response of LC neurons to focally applied glutamate was also increased. Together, these results suggest that accumulation of EAA after synaptic stimulation may be responsible for ADD. To test if ADD is a result of the activation of presynaptic mGluRs, the effect of selective mGluR antagonists on ADD was assessed. In the presence of t-PDC, bath applied (S)-amino-2-methyl-4 phosphonobutanoic acid (MAP4, 500 microM), a mGluR group III antagonist, significantly reversed the decrease in T/C ratio after a train stimulation [from 0.66 +/- 0.04 to 0.81 +/- 0.02 (mean +/- SE), n = 5]. The T/C ratio in the presence of MAP4 was not different from that measured in the absence of a stimulation volley. Conversely, ethyl glutamic acid (EGLU, 500 microM), a mGluR group II antagonist, failed to alter the T/C ratio. Together, these results suggest that, in LC, group III presynaptic mGluR activation provides a feedback mechanism by which excitatory synaptic transmission can be negatively modulated during high-frequency synaptic activity. Furthermore, this study provides functional differentiation between presynaptic groups II and III mGluR in LC and suggests that the group II mGluR may be involved in functions distinct from those of group III mGluRs. PMID- 10712445 TI - HERG-Like potassium current regulates the resting membrane potential in glomus cells of the rabbit carotid body. AB - Direct evidence for a specific K(+) channel underlying the resting membrane potential in glomus cells of the carotid body has been absent. The product of the human ether-a-go-go-related gene (HERG) produces inward rectifier currents that are known to contribute to the resting membrane potential in other neuronal cells. The goal of the present study was to determine whether carotid body glomus cells express HERG-like K(+) current, and if so, to determine whether a HERG-like current regulates the resting membrane potential. Freshly dissociated rabbit glomus cells under whole cell voltage clamp exhibited slowly decaying outward currents that activated 20-30 mV positive to the resting membrane potential. Raising extracellular K(+) revealed a slowly deactivating inward tail current indicative of HERG-like K(+) current. HERG-like currents were not found in cells resembling type II cells. The HERG-like current was blocked by dofetilide (DOF) in a concentration-dependent manner (IC(50) = 13 +/- 4 nM, mean +/- SE) and high concentrations of Ba(2+) (1 and 10 mM). The biophysical and pharmacological characteristics of this inward tail current suggest that it is conducted by a HERG-like channel. The steady-state activation properties of the HERG-like current (V(h) = -44 +/- 2 mV) suggest that it is active at the resting membrane potential in glomus cells. In whole cell, current-clamped glomus cells (average resting membrane potential, - 48 +/- 4 mV), DOF, but not tetraethylammonium, caused a significant (13 mV) depolarizing shift in the resting membrane potential. Using fluorescence imaging, DOF increased [Ca(2+)](i) in isolated glomus cells. In an in-vitro carotid body preparation, DOF increased basal sensory discharge in the carotid sinus nerve in a concentration-dependent manner. These results demonstrate that glomus cells express a HERG-like current that is active at, and responsible for controlling the resting membrane potential. PMID- 10712446 TI - Circuit dynamics and coding strategies in rodent somatosensory cortex. AB - Previous experimental studies of both cortical barrel and thalamic barreloid neuron responses in rodent somatosensory cortex have indicated an active role for barrel circuitry in processing thalamic signals. Previous modeling studies of the same system have suggested that a major function of the barrel circuit is to render the response magnitude of barrel neurons particularly sensitive to the temporal distribution of thalamic input. Specifically, thalamic inputs that are initially synchronous strongly engage recurrent excitatory connections in the barrel and generate a response that briefly withstands the strong damping effects of inhibitory circuitry. To test this experimentally, we recorded responses from 40 cortical barrel neurons and 63 thalamic barreloid neurons evoked by whisker deflections varying in velocity and amplitude. This stimulus evoked thalamic response profiles that varied in terms of both their magnitude and timing. The magnitude of the thalamic population response, measured as the average number of evoked spikes per stimulus, increased with both deflection velocity and amplitude. On the other hand, the degree of initial synchrony, measured from population peristimulus time histograms, was highly correlated with the velocity of whisker deflection, deflection amplitude having little or no effect on thalamic synchrony. Consistent with the predictions of the model, the cortical population response was determined largely by whisker velocity and was highly correlated with the degree of initial synchrony among thalamic neurons (R(2) = 0.91), as compared with the average number of evoked thalamic spikes (R(2) = 0.38). Individually, the response of nearly all cortical cells displayed a positive correlation with deflection velocity; this homogeneity is consistent with the dependence of the cortical response on local circuit interactions as proposed by the model. By contrast, the response of individual thalamic neurons varied widely. These findings validate the predictions of the modeling studies and, more importantly, demonstrate that the mechanism by which the cortex processes an afferent signal is inextricably linked with, and in fact determines, the saliency of neural codes embedded in the thalamic response. PMID- 10712448 TI - Modification of synaptic transmission and sodium channel inactivation by the insect-selective scorpion toxin LqhalphaIT. AB - The peptide LqhalphaIT is an alpha-scorpion toxin that shows significant selectivity for insect sodium channels over mammalian channels. We examined the symptoms of LqhalphaIT-induced paralysis and its neurophysiological correlates in the house fly (Musca domestica). Injection of LqhalphaIT into fly larvae produced hyperactivity characterized by continuous, irregular muscle twitching throughout the body. These symptoms were correlated with elevated excitability in motor units caused by two physiological effects of the toxin: 1) increased transmitter release and 2) repetitive action potentials in motor nerves. Increased transmitter release was evident as augmentation of neurally evoked synaptic current, and this was correlated with an increased duration of action potential associated current (APAC) in loose patch recordings from nerve terminals. Repetitive APACs were observed to invade nerve endings. The toxin produced marked inhibition of sodium current inactivation in fly central neurons, which can account for increased duration of the APAC and elevated neurotransmitter release at the neuromuscular junction. Steady-state inactivation was shifted significantly to more positive potentials, whereas voltage-dependent activation of the channels was not affected. The shift in steady-state inactivation provides a mechanism for inducing repetitive activity in motoneurons. The effects of LqhalphaIT on sodium channel inactivation in motor nerve endings can account both for increased transmitter release and repetitive activity leading to hyperactivity in affected insects. PMID- 10712447 TI - Cannabinoid receptor modulation of synapses received by cerebellar Purkinje cells. AB - The high density of cannabinoid receptors in the cerebellum and the degradation of motor coordination produced by cannabinoid intoxication suggest that synaptic transmission in the cerebellum may be strongly regulated by cannabinoid receptors. Therefore the effects of exogenous cannabinoids on synapses received by Purkinje cells were investigated in rat cerebellar slices. Parallel fiber evoked (PF) excitatory postsynaptic currents (EPSCs) were strongly inhibited by bath application of the cannabinoid receptor agonist WIN 55212-2 (5 microM, 12% of baseline EPSC amplitude). This effect was completely blocked by the cannabinoid CB1 receptor antagonist SR 141716. It is unlikely that this was the result of alterations in axonal excitability because fiber volley velocity and kinetics were unchanged and a cannabinoid-induced decrease in fiber volley amplitude was very minor (93% of baseline). WIN 55212-2 had no effect on the amplitude or frequency of spontaneously occurring miniature EPSCs (mEPSCs), suggesting that the effect of CB1 receptor activation on PF EPSCs was presynaptically expressed, but giving no evidence for modulation of release processes after Ca(2+) influx. EPSCs evoked by climbing fiber (CF) stimulation were less powerfully attenuated by WIN 55212-2 (5 microM, 74% of baseline). Large, action potential-dependent, spontaneously occurring inhibitory postsynaptic currents (sIPSCs) were either severely reduced in amplitude (<25% of baseline) or eliminated. Miniature IPSCs (mIPSCs) were reduced in frequency (52% of baseline) but not in amplitude, demonstrating suppression of presynaptic vesicle release processes after Ca(2+) influx and suggesting an absence of postsynaptic modulation. The decrease in mIPSC frequency was not large enough to account for the decrease in sIPSC amplitude, suggesting that presynaptic voltage gated channel modulation was also involved. Thus, while CB1 receptor activation reduced neurotransmitter release at all major classes of Purkinje cell synapses, this was not accomplished by a single molecular mechanism. At excitatory synapses, cannabinoid suppression of neurotransmitter release was mediated by modulation of voltage-gated channels in the presynaptic axon terminal. At inhibitory synapses, in addition to modulation of presynaptic voltage-gated channels, suppression of the downstream vesicle release machinery also played a large role. PMID- 10712449 TI - Group I, II, and III mGluR compounds affect rhythm generation in the gastric circuit of the crustacean stomatogastric ganglion. AB - We have studied the effects of group I, II, and III metabotropic glutamate receptor (mGluR) agonists on rhythm generation by the gastric circuit of the stomatogastric ganglion (STG) of the Caribbean spiny lobster Panulirus argus. All mGluR agonists and some antagonists we tested in this study had clear and distinct effects on gastric rhythm generation when superfused over combined oscillating or blocked silent STG preparations. A consistent difference between group I agonists and group II and III agonists was that group I agonists acted excitatory. The group I-specific agonists L-quisqualic acid and (S)-3,5 dihydroxyphenylglycine, as well as the nonspecific agonist (1S,3R)-1 aminocyclopentane-1, 3-dicarboxylic acid accelerated ongoing rhythms and could induce gastric rhythms in silent preparations. The group II agonist (2S,1'S, 2'S) 2-(carboxycyclopropyl)glycine (L-CCG-I) and the group III agonist L(+)-2-amino-4 phosphonobutyric acid (L-AP4) slowed down or completely blocked ongoing gastric rhythms and were without detectable effect on silent preparations. The action of L-CCG-I was blocked partially by the group-II-specific antagonist, (RS)-1-amino-5 phosphonoindan-1-carboxylic acid [(RS)APICA], and the group-III-specific antagonist (RS)-alpha-methyl-4-phosphonophenylglycine completely blocked the action of L-AP4. Besides its antagonistic action, the group-II-specific antagonist (RS)APICA had a remarkably strong apparent inverse agonist action when applied alone on oscillating preparations. The action of all drugs was dose dependent and reversible, although recovery was not always complete. In our experiments, the effects of none of the mGluR-specific agonists were antagonized or amplified by the N-methyl-D-aspartate (NMDA)-receptor-specific antagonist D(-) 2-amino-5-phosphonopentanoic acid, excluding the contamination of responses to mGluR agonists by nonspecific cross-reactivity with NMDA receptors. Picrotoxin did not prevent the inhibitory action of L-CCG-I and L-AP4. We conclude that mGluRs, probably similar to those belonging to groups I, II, and III described in mammals, may play a role as modulators of gastric circuit rhythm generation in vivo. PMID- 10712450 TI - Morphological identification of physiologically characterized afferents innervating the turtle posterior crista. AB - The turtle posterior crista consists of two hemicristae. Each hemicrista extends from the planum semilunatum to the nonsensory torus and includes a central zone (CZ) surrounded by a peripheral zone (PZ). Type I and type II hair cells are found in the CZ and are innervated by calyx, dimorphic and bouton afferents. Only type II hair cells and bouton fibers are found in the PZ. Units were intraaxonally labeled in a half-head preparation. Bouton (B) units could be near the planum (BP), near the torus (BT), or in midportions of a hemicrista, including the PZ and CZ. Discharge properties of B units vary with longitudinal position in a hemicrista but not with morphological features of their peripheral terminations. BP units are regularly discharging and have small gains and small phase leads re angular head velocity. BT units are irregular and have large gains and large phase leads. BM units have intermediate properties. Calyx (C) and dimorphic (D) units have similar discharge properties and were placed into a single calyx-bearing (CD) category. While having an irregular discharge resembling BT units, CD units have gains and phases similar to those of BM units. Rather than any single discharge property, it is the relation between discharge regularity and either gain or phase that makes CD units distinctive. Multivariate statistical formulas were developed to infer a unit's morphological class (B or CD) and longitudinal position solely from its discharge properties. To verify the use of the formulas, discharge properties were compared for units recorded intraaxonally or extracellularly in the half-head or extracellularly in intact animals. Most B units have background rates of 10-30 spikes/s. The CD category was separated into CD-high and CD-low units with background rates above or below 5 spikes/s, respectively. CD-low units have lower gains and phases and are located nearer the planum than CD-high units. In their response dynamics over a frequency range from 0.01-3 Hz, BP units conform to an overdamped torsion pendulum model. Other units show departures from the model, including high frequency gain increases and phase leads. The longitudinal gradient in the physiology of turtle B units resembles a similar gradient in the anamniote crista. In many respects, turtle CD units have discharge properties resembling those of calyx-bearing units in the mammalian central zone. PMID- 10712451 TI - Responses to efferent activation and excitatory response-intensity relations of turtle posterior-crista afferents. AB - Multivariate statistical formulas were used to infer the morphological type and longitudinal position of extracellularly recorded afferents. Efferent fibers were stimulated electrically in the nerve branch interconnecting the anterior and posterior VIIIth nerves. Responses of bouton (B) units depended on their inferred position: BP units (near the planum semilunatum) showed small excitatory responses; BT units (near the torus) were inhibited; BM units (in an intermediate position) had a mixed response, including an initial inhibition and a delayed excitation. Calyx-bearing (CD-high) units with an appreciable background discharge showed large per-train excitatory responses followed by smaller post train responses that could outlast the shock train by 100 s. Excitatory responses were smaller in calyx-bearing (CD-low) units having little or no background activity than in CD-high units. Excitatory response-intensity functions, derived from the discharge during 2-s angular-velocity ramps varying in intensity, were fit by empirical functions that gave estimates of the maximal response (r(MAX)), a threshold velocity (v(T)), and the velocity producing a half-maximal response (v(1/2)). Linear gain is equal to r(MAX)/v(S), v(S) = v(1/2) - v(T). v(S) provides a measure of the velocity range over which the response is nearly linear. For B units, r(MAX) declines by as much as twofold over the 2-s ramp, whereas for CD units, r(MAX) increases by 15% during the same time period. At the end of the ramp, r(MAX) is on average twice as high in CD as in B units. Thresholds are negligible in most spontaneously active units, including almost all B and CD-high units. Silent CD-low units typically have thresholds of 10-100 deg/s. BT units have very high linear gains and v(S) < 10 deg/s. Linear gains are considerably lower in BP units and v(S) > 150 deg/s. CD-high units have intermediate gains and near 100 deg/s v(S) values. CD-low units have low gains and v(S) values ranging from 150 to more than 300 deg/s. The results suggest that BT units are designed to measure the small head movements involved in postural control, whereas BP and CD units are more appropriate for monitoring large volitional head movements. The former units are silenced by efferent activation, whereas the latter units are excited. This suggests that the efferent system switches the turtle posterior crista from a "postural" to a "volitional" mode. PMID- 10712453 TI - Diverse ionic currents and electrical activity of cultured myenteric neurons from the guinea pig proximal colon. AB - The aim of this study was to perform a patch-clamp analysis of myenteric neurons from the guinea pig proximal colon. Neurons were enzymatically dispersed, cultured for 2-7 days, and recorded from using whole cell patch clamp. The majority of cells fired phasically, whereas about one-quarter of the neurons fired in a tonic manner. Neurons were divided into three types based on the currents activated. The majority of tonically firing neurons lacked an A-type current, but generated a large fast transient outward current that was associated with the rapid repolarizing phase of an action potential. The fast transient outward current was dependent on calcium entry and was blocked by tetraethylammonium. Cells that expressed both an A-type current and a fast transient outward current were mostly phasic. Depolarization of these cells to suprathreshold potentials from less than -60 mV failed to trigger action potentials, or action potentials were only triggered after a delay of >50 ms. However, depolarizations from more positive potentials triggered action potentials with minimal latency. Neurons that expressed neither the A-type current or the fast transient outward current were all phasic. Sixteen percent of neurons were similar to AH/type II neurons in that they generated a prolonged afterhyperpolarization following an action potential. The current underlying the prolonged afterhyperpolarization showed weak inward rectification and had a reversal potential near the potassium equilibrium potential. Thus cultured isolated myenteric neurons of the guinea pig proximal colon retain many of the diverse properties of intact neurons. This preparation is suitable for further biophysical and molecular characterization of channels expressed in colonic myenteric neurons. PMID- 10712452 TI - Acetylcholine modulates respiratory pattern: effects mediated by M3-like receptors in preBotzinger complex inspiratory neurons. AB - Perturbations of cholinergic neurotransmission in the brain stem affect respiratory motor pattern both in vivo and in vitro; the underlying cellular mechanisms are unclear. Using a medullary slice preparation from neonatal rat that spontaneously generates respiratory rhythm, we patch-clamped inspiratory neurons in the preBotzinger complex (preBotC), the hypothesized site for respiratory rhythm generation, and simultaneously recorded respiratory-related motor output from the hypoglossal nerve (XIIn). Most (88%) of the inspiratory neurons tested responded to local application of acetylcholine (ACh) or carbachol (CCh) or bath application of muscarine. Bath application of 50 microM muscarine increased the frequency, amplitude, and duration of XIIn inspiratory bursts. At the cellular level, muscarine induced a tonic inward current, increased the duration, and decreased the amplitude of the phasic inspiratory inward currents in preBotC inspiratory neurons recorded under voltage clamp at -60 mV. Muscarine also induced seizure-like activity evident during expiratory periods in XIIn activity; these effects were blocked by atropine. In the presence of tetrodotoxin (TTX), local ejection of 2 mM CCh or ACh onto preBotC inspiratory neurons induced an inward current along with an increase in membrane conductance under voltage clamp and induced a depolarization under current clamp. This response was blocked by atropine in a concentration-dependent manner. Bath application of 1 microM pirenzepine, 10 microM gallamine, or 10 microM himbacine had little effect on the CCh-induced current, whereas 10 microM 4-diphenylacetoxy-N-methylpiperidine methiodide blocked the current. The current-voltage (I-V) relationship of the CCh induced response was linear in the range of -110 to -20 mV and reversed at -11.4 mV. Similar responses were found in both pacemaker and nonpacemaker inspiratory neurons. The response to CCh was unaffected when patch electrodes contained a high concentration of EGTA (11 mM) or bis-(o-aminophenoxy)-N,N,N',N'-tetraacetic acid (10 mM). The response to CCh was reduced greatly by substitution of 128 mM Tris-Cl for NaCl in the bath solution; the I-V curve shifted to the left and the reversal potential shifted to -47 mV. Lowering extracellular Cl(-) concentration from 140 to 70 mM had no effect on the reversal potential. These results suggest that in preBotC inspiratory neurons, ACh acts on M3-like ACh receptors on the postsynaptic neurons to open a channel permeable to Na(+) and K(+) that is not Ca(2+) dependent. This inward cation current plays a major role in depolarizing preBotC inspiratory neurons, including pacemakers, that may account for the ACh induced increase in the frequency of respiratory motor output observed at the systems/behavioral level. PMID- 10712454 TI - Modulation of laryngeal responses to superior laryngeal nerve stimulation by volitional swallowing in awake humans. AB - Laryngeal sensori-motor closure reflexes are important for the protection of the airway and prevent the entry of foreign substances into the trachea and lungs. The purpose of this study was to determine how such reflexes might be modulated during volitional swallowing in awake humans, when persons are at risk of entry of food or liquids into the airway. The frequency and the amplitude of laryngeal adductor responses evoked by electrical stimulation of the internal branch of the superior laryngeal nerve (ISLN) were studied during different phases of volitional swallowing. Subjects swallowed water on command while electrical stimuli were presented to the ISLN at various intervals from 500 ms to 5 s following the command. Laryngeal adductor responses to unilateral ISLN stimulation were recorded bilaterally in the thyroarytenoid muscles using hooked wire electrodes. Early ipsilateral R1 responses occurred at 17 ms, and later bilateral R2 began around 65 ms. The muscle responses to stimuli occurring during expiration without swallowing were quantified as control trials. Responses to stimulation presented before swallowing, during the swallow, within 3 s after swallowing, and between 3 and 5 s after a swallow were measured. The frequency and amplitude of three responses (ipsilateral R1 and bilateral R2) relative to the control responses were compared across the different phases relative to the occurrence of swallowing. Results demonstrated that a reduction occurred in both the frequency and amplitude of the later bilateral R2 laryngeal responses to electrical stimulation for up to 3 s after swallowing (P = 0.005). The amplitude and frequency of ipsilateral R1 laryngeal responses, however, did not show a significant main effect following the swallow (P = 0.28), although there was a significant time by measure interaction (P = 0.006) related to reduced R1 response amplitude up to 3 s after swallowing (P = 0.021). Therefore, the more rapid and shorter unilateral R1 responses continued to provide some, albeit reduced, laryngeal protective functions after swallowing, whereas the later bilateral R2 responses were suppressed both in occurrence and amplitude for up to 3 s after swallowing. The results suggest that R2 laryngeal adductor responses are suppressed following swallowing when residues may remain in the laryngeal vestibule putting persons at increased risk for the entry of foreign substances into the airway. PMID- 10712455 TI - Competition between internal AlF(4)(-) and receptor-mediated stimulation of dorsal raphe neuron G-proteins coupled to calcium current inhibition. AB - Intracellular aluminum fluoride (AlF(4)(-)), placed in a patch pipette, activated a G-protein, resulting in a "tonic" inhibition of the Ca(2+) current of isolated serotonergic neurons of the rat dorsal raphe nucleus. Serotonin (5-HT) also inhibits the Ca(2+) current of these cells. After external bath application and quick removal of 5-HT to an AlF(4)(-) containing cell, there was a reversal or transient disinhibition (TD) of the inhibitory effect of AlF(4)(-) on Ca(2+) current. A short predepolarization of the membrane potential to +70 mV, a condition that is known to reverse G-protein-mediated inhibition, reversed the inhibitory effect of AlF(4)(-) on Ca(2+) current and brought the Ca(2+) current to the same level as that seen at the peak of the TD current. With AlF(4)(-) in the pipette, the TD phenomenon could be eliminated by lowering pipette MgATP, or by totally chelating pipette Al(3+). In the presence of AlF(4)(-), but with either lowered MgATP or extreme efforts to eliminate pipette Al(3+), the rate of recovery from 5-HT on wash was slowed, a condition opposite to that where a TD occurred. The putative complex of AlF(4)(-)-bound G-protein (Galpha.GDP. AlF(4)( )) appeared to free G-betagamma-subunits, mimicking the effect on Ca(2+) channels of the G.GTP complex. The ON-rate of the inhibition of Ca(2+) current, after a depolarizing pulse, by betagamma-subunits released by AlF(4)(-) in the pipette was significantly slower than that of the agonist-activated G-protein. The OFF rate of the AlF(4)(-)-mediated inhibition in response to a depolarizing pulse, a measure of the affinity of the free G-betagamma-subunit for the Ca(2+) channel, was slightly slower than that of the agonist stimulated G-protein. In summary, AlF(4)(-) modified the OFF-rate kinetics of G-protein activation by agonists, but had little effect on the kinetics of the interaction of the betagamma-subunit with Ca(2+) channels. Agonist application temporarily reversed the effects of AlF(4)(-), making it a complementary tool to GTP-gamma-S for the study of G protein interactions. PMID- 10712456 TI - Neurons in the primate superior colliculus coding for arm movements in gaze related coordinates. AB - In the intermediate and deep layers of the superior colliculus (SC), a well established oculomotor structure, a substantial population of cells is involved in the control of arm movements. To examine the reference frame of these neurons, we recorded in two rhesus monkeys (Macaca mulatta) the discharges of 331 neurons in the SC and the underlying mesencephalic reticular formation (MRF) while monkeys reached to the same target location during different gaze orientations. For 65 reach-related cells with sufficient data and for simultaneously recorded electromyograms (EMGs) of 11 arm muscles, we calculated an ANOVA (factors: target position, gaze angle) and a gaze-dependency (GD) index. EMGs and the activity of many (60%) of the reach-related neurons were not influenced by the target representation on the retina or eye position. We refer to these as "gaze independent" reach neurons. For 40%, however, the GD fell outside the range of the muscle modulation, and the ANOVA showed a significant influence of gaze. These "gaze-related" reach neurons discharge only when the monkey reaches for targets having specific coordinates in relation to the gaze axis, i.e., for targets in a gaze-related "reach movement field" (RMF). Neuronal activity was not modulated by the specific path of the arm movement, the muscle pattern that is necessary for its realization or the arm that was used for the reach. In each SC we found gaze-related neurons with RMFs both in the contralateral and in the ipsilateral hemifield. The topographical organization of the gaze-related reach neurons in the SC could not be matched with the well-known visual and oculomotor maps. Gaze-related neurons were more modulated in their strength of activity with different directions of arm movements than were gaze-independent reach neurons. Gaze-related reach neurons were recorded at a median depth of 2.03 mm below SC surface in the intermediate layers, where they overlap with saccade-related burst neurons (median depth: 1.55 mm). Most of the gaze-independent reach cells were found in a median depth of 4.01 mm below the SC surface in the deep layers and in the underlying MRF. The gaze-related reach neurons operating in a gaze-centered coordinate system could signal either the desired target position with respect to gaze direction or the motor error between gaze axis and reach target. The gaze independent reach neurons, possibly operating in a shoulder- or arm-centered reference frame, might carry signals closer to motor output. Together these two types of reach neurons add evidence to our hypothesis that the SC is involved in the sensorimotor transformation for eye-hand coordination in primates. PMID- 10712457 TI - Exon 5 and spermine regulate deactivation of NMDA receptor subtypes. AB - Deactivation of N-methyl-D-aspartate (NMDA) channels after brief agonist exposure determines the duration of their synaptic activation during excitatory neurotransmission. We performed patch-clamp recordings of L-glutamate responses from human embryonic kidney tumoral cells (HEK293) expressing NR1 subunit variants lacking exon 5 together with the NR2B subunit. These responses had deactivation components that lasted several seconds. The presence of exon 5 or spermine greatly accelerated deactivation of L-glutamate responses through alterations in desensitization. These effects were also observed at positive holding potentials and in the presence of physiological Mg(2+). Thus NR1 splicing and polyamines may have profound effects on the kinetics of NMDA receptor mediated synaptic transmission. PMID- 10712458 TI - Selective inhibition by adenosine of mGluR IPSPs in dopamine neurons after cocaine treatment. AB - With repeated exposure to psychostimulants such as cocaine and amphetamine, long lasting changes occur in the mesolimbic dopamine system that are thought to underlie continued drug-seeking and relapse. One consequence of repeated cocaine treatment is an increase in extracellular adenosine in the ventral tegmental area (VTA), which results in tonic inhibition of synaptic input to dopamine neurons. The synapse specificity of this increased adenosine tone was examined on glutamate- and GABA-mediated responses using the selective A1 receptor antagonist 1, 3-dipropyl-8-cyclopentylxanthine (DPCPX). The slow, metabotropic glutamate receptor (mGluR)-mediated inhibitory postsynaptic potential (IPSP) was enhanced by DPCPX only in slices from psychostimulant-treated animals. Under resting conditions, DPCPX was without effect on fast excitatory postsynaptic currents (EPSCs) in slices from saline- or cocaine-treated animals. However, in the presence of amphetamine, DPCPX did augment fast EPSCs in slices from cocaine treated rats. Although DPCPX increased GABA(B) IPSPs, the magnitude of the increase was not altered by cocaine pretreatment, even in the presence of amphetamine. This suggests that the elevated adenosine tone induced by cocaine treatment acts preferentially on glutamate terminals. Furthermore, the inhibition of the mGluR IPSP by endogenous adenosine may result in more effective burst firing mediated by glutamate afferents in cocaine-treated rats, a phenomenon known to enhance dopamine release. PMID- 10712459 TI - Neuromuscular system of the flexible arm of the octopus: physiological characterization. AB - The octopus arm is an outstanding example of an efficient boneless and highly flexible appendage. We have begun characterizing the neuromuscular system of the octopus arm in both innervated muscle preparations and dissociated muscle cells. Functionally antagonistic longitudinal and transverse muscle fibers showed no differences in membrane properties and mode of innervation. The muscle cells are excitable but have a broad range of linear membrane properties. They are electrotonically very compact so that localized synaptic inputs can control the membrane potential of the entire muscle cell. Three distinct excitatory neuronal inputs to each arm muscle cell were identified; their reversal potentials were extrapolated to be about -10 mV. These appear to be cholinergic as they are blocked by hexamethonium, D-tubocurarine, and atropine. Two inputs have low quantal amplitude (1-7 mV) and slow rise times (4-15 ms), whereas the third has a large size (5-25 mV) and fast rise time (2-4 ms). This large synaptic input is most likely due to exceptionally large quantal events. The probability of release is rather low, suggesting a stochastic activation of muscle cells. All inputs demonstrated a modest activity-dependent plasticity typical of fast neuromuscular systems. The pre- and postsynaptic properties suggest a rather direct relation between neuronal activity and muscle action. The lack of significant electrical coupling between muscle fibers and the indications for the small size of the motor units suggest that the neuromuscular system of the octopus arm has evolved to ensure a high level of precise localization in the neural control of arm function. PMID- 10712460 TI - Dendritic spikes and their influence on extracellular calcium signaling. AB - Extracellular calcium is critical for many neural functions, including neurotransmission, cell adhesion, and neural plasticity. Experiments have shown that normal neural activity is associated with changes in extracellular calcium, which has motivated recent computational work that employs such fluctuations in an information-bearing role. This possibility suggests that a new style of computing is taking place in the mammalian brain in addition to current 'circuit' models that use only neurons and connections. Previous computational models of rapid external calcium changes used only rough approximations of calcium channel dynamics to compute the expected calcium decrements in the extracellular space. Using realistic calcium channel models, experimentally measured back-propagating action potentials, and a model of the extracellular space, we computed the fluctuations in external calcium that accrue during neural activity. In this realistic setting, we showed that rapid, significant changes in local external calcium can occur when dendrites are invaded by back-propagating spikes, even in the presence of an extracellular calcium buffer. We further showed how different geometric arrangements of calcium channels or dendrites prolong or amplify these fluctuations. Finally, we computed the influence of experimentally measured synaptic input on peridendritic calcium fluctuations. Remarkably, appropriately timed synaptic input can amplify significantly the decrement in external calcium. The model shows that the extracellular space and the calcium channels that access it provide a medium that naturally integrates coincident spike activity from different dendrites that intersect the same tissue volume. PMID- 10712462 TI - Escape swim network interneurons have diverse roles in behavioral switching and putative arousal in Pleurobranchaea. AB - Escape swimming in the predatory sea slug Pleurobranchaea is a dominant behavior that overrides feeding, a behavioral switch caused by swim-induced inhibition of feeding command neurons. We have now found distinct roles for the different swim interneurons in acute suppression of feeding during the swim and in a longer-term stimulation of excitability in the feeding network. The identified pattern generating swim neurons A1, A3, A10, and their follower interneuron A-ci1, suppress feeding motor output partly by excitation of the I1 feeding interneurons, which monosynaptically inhibit both the feeding command neurons, PC(P), PSE, and other major interneurons, the I2s. This mechanism exerts broad inhibition of the feeding network suitable to an escape response; broader than feeding suppression in learned and satiation-induced food avoidance and acting through a different presynaptic pathway. Four intrinsic neuromodulatory neurons of the swim network, the serotonergic As1-4, add little to direct suppression of feeding. Rather, they monosynaptically excite the serotonergic metacerebral giant (MCG) neurons of the feeding network, themselves intrinsic neuromodulators of feeding, as well as a cluster of adjacent serotonergic feeding neurons, with both fast and slow EPSPs. They also provide mild neuromodulatory excitation of the PC(P)/PSE feeding command neurons, and I1 and I2 feeding interneurons, which is masked by inhibition during the swim. As1-4 also excite the serotonergic pedal ganglion G neurons for creeping locomotion. These observations further delineate the nature of the putative serotonergic arousal system of gastropods and suggest a central coordinating role to As1-4. PMID- 10712461 TI - Extracellular pH changes and accompanying cation shifts during ouabain-induced spreading depression. AB - Interstitial ionic shifts that accompany ouabain-induced spreading depression (SD) were studied in rat hippocampal and cortical slices in the presence and absence of extracellular Ca(2+). A double-barreled ion-selective microelectrode specific for H(+), K(+), Na(+), or Ca(2+) was placed in the CA1 stratum radiatum or midcortical layer. Superfusion of 100 microM ouabain caused a rapid, negative, interstitial voltage shift (2-10 mV) after 3-5 min. The negativity was accompanied by a rapid alkaline transient followed by prolonged acidosis. In media containing 3 mM Ca(2+), the alkalosis induced by ouabain averaged 0.07 +/- 0.01 unit pH. In media with no added Ca(2+) and 2 mM EGTA, the alkaline shift was not significantly different (0.09 +/- 0.02 unit pH). The alkaline transient was unaffected by inhibiting Na(+)-H(+) exchange with ethylisopropylamiloride (EIPA) or by blocking endoplasmic reticulum Ca(2+) uptake with thapsigargin or cyclopiazonic acid. Alkaline transients were also observed in Ca(2+)-free media when SD was induced by microinjecting high K(+). The late acidification accompanying ouabain-induced SD was significantly reduced in Ca(2+)-free media and in solutions containing EIPA. The ouabain-induced SD was associated with a rapid but relatively modest increase in [K(+)](o). In the presence of 3 mM external Ca(2+), the mean peak elevation of [K(+)](o) was 12 +/- 0.62 mM. In Ca(2+)-free media, the elevation of [K(+)](o) had a more gradual onset and reached a significantly larger peak value, which averaged 22 +/- 1.1 mM. The decrease in [Na(+)](o) that accompanied ouabain-induced SD was somewhat greater. The [Na(+)](o) decreased by averages of 40 +/- 7 and 33 +/- 3 mM in Ca(2+) and Ca(2+)-free media, respectively. In media containing 1.2 mM Ca(2+), ouabain induced SD was associated with a substantial decrease in [Ca(2+)](o) that averaged 0.73 +/- 0. 07 mM. These data demonstrate that in comparison with conventional SD, ouabain-induced SD exhibits ion shifts that are qualitatively similar but quantitatively diminished. The presence of external Ca(2+) can modulate the phenomenon but is irrelevant to the generation of the SD and its accompanying alkaline pH transient. Significance of these results is discussed in reference to the propagation of SD and the generation of interstitial pH changes. PMID- 10712463 TI - Responses of neurons in the inferior colliculus to dynamic interaural phase cues: evidence for a mechanism of binaural adaptation. AB - Responses to sound stimuli that humans perceive as moving were obtained for 89 neurons in the inferior colliculus (IC) of urethan-anesthetized guinea pigs. Triangular and sinusoidal interaural phase modulation (IPM), which produced dynamically varying interaural phase disparities (IPDs), was used to present stimuli with different depths, directions, centers, and rates of apparent motion. Many neurons appeared sensitive to dynamic IPDs, with responses at any given IPD depending strongly on the IPDs the stimulus had just passed through. However, it was the temporal pattern of the response, rather than the motion cues in the IPM, that determined sensitivity to features such as motion depth, direction, and center locus. IPM restricted only to the center of the IPD responsive area, evoked lower discharge rates than when the stimulus either moved through the IPD responsive area from outside, or up and down its flanks. When the stimulus was moved through the response area first in one direction and then back in the other, and the same IPDs evoked different responses, the response to the motion away from the center of the IPD responsive area was always lower than the response to the motion toward the center. When the IPD was closer at which the direction of motion reversed was to the center, the response to the following motion was lower. In no case did we find any evidence for neurons that under all conditions preferred one direction of motion to the other. We conclude that responses of IC neurons to IPM stimuli depend not on the history of stimulation, per se, but on the history of their response to stimulation, irrespective of the specific motion cues that evoke those responses. These data are consistent with the involvement of an adaptation mechanism that resides at or above the level of binaural integration. We conclude that our data provide no evidence for specialized motion detection involving dynamic IPD cues in the auditory midbrain of the mammal. PMID- 10712464 TI - Firing properties and electrotonic structure of Xenopus larval spinal neurons. AB - Whole cell voltage- and current-clamp measurements were done on intact Xenopus laevis larval spinal neurons at developmental stages 42-47. Firing patterns and electrotonic properties of putative interneurons from the dorsal and ventral medial regions of the spinal cord at myotome levels 4-6 were measured in isolated spinal cord preparations. Passive electrotonic parameters were determined with internal cesium sulfate solutions as well as in the presence of active potassium conductances. Step-clamp stimuli were combined with white-noise frequency domain measurements to determine both linear and nonlinear responses at different membrane potential levels. Comparison of analytic and compartmental dendritic models provided a way to determine the number of compartments needed to describe the dendritic structure. The electrotonic structure of putative interneurons was correlated with their firing behavior such that highly accommodating neurons (Type B) had relatively larger dendritic areas and lower electrotonic lengths compared with neurons that showed sustained action potential firing in response to a constant current (Type A). Type A neurons had a wide range of dendritic areas and potassium conductances that were activated at membrane potentials more negative than observed in Type B neurons. The differences in the potassium conductances were in part responsible for a much greater rectification in the steady-state current voltage (I-V curve) of the strongly accommodating neurons compared with repetitively firing cells. The average values of the passive electrotonic parameters found for Rall Type A and B neurons were c(soma) = 3.3 and 2.6 pF, g(soma) = 187 and 38 pS, L = 0.36 and 0.21, and A = 3.3 and 6.5 for soma capacitance, soma conductance, electrotonic length, and the ratio of the dendritic to somatic areas, respectively. Thus these experiments suggest that there is a correlation between the electrotonic structure and the excitability properties elicited from the somatic region. PMID- 10712465 TI - Active dendritic membrane properties of Xenopus larval spinal neurons analyzed with a whole cell soma voltage clamp. AB - Voltage- and current-clamp measurements of inwardly directed currents were made from the somatic regions of Xenopus laevis spinal neurons. Current-voltage (I-V) curves determined under voltage clamp, but not current clamp, were able to indicate a negative slope conductance in neurons that showed strong accommodating action potential responses to a constant current stimulation. Voltage-clamp I-V curves from repetitive firing neurons did not have a net negative slope conductance and had identical I-V plots under current clamp. Frequency domain responses indicate negative slope conductances with different properties with or without tetrodotoxin, suggesting that both sodium and calcium currents are present in these spinal neurons. The currents obtained from a voltage clamp of the somatic region were analyzed in terms of spatially controlled soma membrane currents and additional currents from dendritic potential responses. Linearized frequency domain analysis in combination with both voltage- and current-clamp responses over a range of membrane potentials was essential for an accurate determination of consistent neuronal model behavior. In essence, the data obtained at resting or hyperpolarized membrane potentials in the frequency domain were used to determine the electrotonic structure, while both the frequency and time domain data at depolarized potentials were required to characterize the voltage-dependent channels. Finally, the dendritic and somatic membrane properties were used to reconstruct the action potential behavior and quantitatively predict the dependence of neuronal firing properties on electrotonic structure. The reconstructed action potentials reproduced the behavior of two broad distributions of interneurons characterized by their degree of accommodation. These studies suggest that in addition to the ionic conductances, electrotonic structure is correlated with the action potential behavior of larval neurons. PMID- 10712466 TI - Stochastic resonance improves signal detection in hippocampal CA1 neurons. AB - Stochastic resonance (SR) is a phenomenon observed in nonlinear systems whereby the introduction of noise enhances the detection of a subthreshold signal for a certain range of noise intensity. The nonlinear threshold detection mechanism that neurons employ and the noisy environment in which they reside makes it likely that SR plays a role in neural signal detection. Although the role of SR in sensory neural systems has been studied extensively, its role in central neurons is unknown. In many central neurons, such as the hippocampal CA1 cell, very large dendritic trees are responsible for detecting neural input in a noisy environment. Attenuation due to the electrotonic length of these trees is significant, suggesting that a method other than passive summation is necessary if signals at the distal ends of the tree are to be detected. The hypothesis that SR plays an important role in the detection of distal synaptic inputs first was tested in a computer simulation of a CA1 cell and then verified with in vitro rat hippocampal slices. The results clearly showed that SR can enhance signal detection in CA1 hippocampal cells. Moreover, high levels of noise were found to equalize detection of synaptic signals received at varying positions on the dendritic tree. The amount of noise needed to evoke the effect is compared with physiological noise in slices and in vivo. PMID- 10712467 TI - NMDA and AMPA receptors in the dorsal nucleus of the lateral lemniscus shape binaural responses in rat inferior colliculus. AB - Binaural responses of single neurons in the rat's central nucleus of the inferior colliculus (ICC) were recorded before and after local injection of excitatory amino acid receptor antagonists (either 1,2, 3,4-tetrahydro-6-nitro-2,3-dioxo benzo[f]quinoxaline-7-sulfonamide disodium [NBQX], (+/-)-3-(2-carboxypiperazin-4 yl)-propyl-1-phosphonic acid [CPP], 6-cyano-7-nitroquinoxaline-2,3-dione [CNQX], or (+/-)-2amino-5-phosphonovaleric acid [APV]) into the dorsal nucleus of the lateral lemniscus (DNLL). Responses were evoked by clicks delivered separately to the two ears at interaural time delays between -1.0 and +30 ms (positive values referring to ipsilateral leading contralateral click pairs). The neurons in our sample were excited by contralateral stimulation and inhibited by ipsilateral stimulation, and the probability of action potentials was reduced as the ipsilateral stimulus was advanced. Binaural inhibition resulted in response suppression that lasted up to 30 ms. Injection of excitatory amino acid antagonists into the DNLL contralateral to the recording site reduced the strength of binaural inhibition in the ICC. The alpha-amino-3-hydroxy-5-methyl-4 isoxazolepropionic acid (AMPA) receptor antagonist NBQX preferentially affected responses at small interaural time intervals (0-1.0 ms), whereas the N-methyl-D aspartate (NMDA) antagonist CPP preferentially affected responses at longer intervals (1-30 ms). Both CNQX and APV produced a release from binaural inhibition, but neither drug was selective for specific intervals. The data support the idea that binaural inhibition in the rat ICC is influenced by both AMPA and NMDA receptor-mediated excitatory events in the contralateral DNLL. The results suggest that the AMPA receptors contribute selectively to the initial component of binaural inhibition and the NMDA receptors to a longer lasting component. PMID- 10712468 TI - Synchronized fast rhythms in inspiratory and expiratory nerve discharges during fictive vocalization. AB - In precollicular decerebrate and paralyzed cats, respiratory nerve activities were recorded during fictive vocalization (FV), which consisted of a distinctive pattern of 1) decreased inspiratory (I) and expiratory (E) phase durations, 2) marked increase of phrenic activity and moderate changes of recurrent laryngeal (RL) and superior laryngeal (SL) I activities, and 3) massive recruitment of laryngeal and abdominal (ABD; lumbar) E activities. FV was produced by electrical stimulation (100 Hz) in the midbrain periaqueductal gray (PAG) or its putative descending pathways in the ventrolateral pons (VLP). Spectral and correlation analyses revealed three types of effect on fast rhythms during FV. 1) I activities: the coherent high-frequency oscillations in I (I-HFO, 60-90 Hz) present in phrenic and RL discharges during the control state did not change qualitatively, but there was an increase of power and a moderate increase (4-10 Hz) of frequency. Sometimes a distinct relatively weak stimulus-locked rhythm appeared. 2) RL and SL activities during E: in recruited discharges, a prominent intrinsic rhythm (coherent E-HFOs at 50-70 Hz) appeared; sometimes a distinct relatively strong stimulus-locked rhythm appeared. 3) ABD activities during E: this recruited activity had no intrinsic rhythm but had an evoked oscillation locked to the stimulus frequency. Thus FV is characterized by 1) appearance of prominent coherent intrinsic rhythms in RL and SL E discharges, which presumably arise as a result of excitation and increased interactions in laryngeal networks; 2) modification of intrinsic rhythmic interactions in inspiratory networks; and 3) evoked rhythms in augmenting-E neuron networks without occurrence of intrinsic rhythms. PMID- 10712469 TI - Facilitatory I wave interaction in proximal arm and lower limb muscle representations of the human motor cortex. AB - Transcranial magnetic stimulation (TMS) of the human motor cortex elicits direct and indirect (I) waves in the corticospinal tract. Facilitatory I wave interaction has been demonstrated with a suprathreshold first stimulus (S1) followed by a subthreshold to threshold second stimulus (S2). Intracortical inhibition (ICI) and intracortical facilitation (ICF) can be studied by another paired TMS paradigm with a subthreshold conditioning stimulus (CS) followed by a suprathreshold test stimulus. Facilitatory I wave interaction in motor representations other than the hand area and its relationship to ICI and ICF has not been studied. We studied I wave interaction, ICI and ICF in an intrinsic hand muscle (abductor pollicis brevis, APB), in a proximal arm muscle (biceps brachii, BB) and in a lower limb muscle (tibialis anterior, TA) in 11 normal subjects. I wave facilitation was studied by paired TMS at 24 interstimulus intervals (ISIs) from 0.5 to 5.1 ms. For APB and TA, facilitation occurred in three distinct peaks at ISIs of 0.9-1.7, 2. 5-3.5, and 4.1-5.1 ms. For BB, facilitation was significant for the first two peaks. The latencies of the peaks were similar among different muscles, but the magnitude of facilitation was much greater for APB and TA compared with BB. For all three muscles, changing the S2 to transcranial electrical stimulation (TES) resulted in much less facilitation of the first peak. For APB, there was significant I wave facilitation with S2 at 72% motor threshold (MT). The same stimulus used as the CS did not elicit ICF at ISI of 15 ms, suggesting that the threshold for eliciting I wave facilitation is lower than that for ICF. For BB and TA, there was no I wave facilitation with S2 at 90% of APB MT, and the same stimulus used as CS led to ICI. Thus in BB and TA the threshold for eliciting ICI is lower than that for I wave facilitation. We conclude that the circuits that mediate I wave interactions are present in the proximal arm and lower limb representations of the motor cortex. I wave facilitation occurs predominately in the cortex and may be primarily related to the monosynaptic corticomotoneuronal (CM) system. The reduced I wave facilitation for BB compared with APB and TA may be related to less extensive CM projection and involvement of other polysynaptic descending pathways. I wave facilitation, ICI, and ICF appears to be mediated by different neuronal circuits. PMID- 10712470 TI - Voltage-dependent, pertussis toxin insensitive inhibition of calcium currents by histamine in bovine adrenal chromaffin cells. AB - Histamine is a known secretagogue in adrenal chromaffin cells. Activation of G protein linked H(1) receptors stimulates phospholipase C, which generates inositol trisphosphate leading to release of intracellular calcium stores and stimulation of calcium influx through store operated and other channels. This calcium leads to the release of catecholamines. In chromaffin cells, the main physiological trigger for catecholamine release is calcium influx through voltage gated calcium channels (I(Ca)). Therefore, these channels are important targets for the regulation of secretion. In particular N- and P/Q-type I(Ca) are subject to inhibition by transmitter/hormone receptor activation of heterotrimeric G proteins. However, the direct effect of histamine on I(Ca) in chromaffin cells is unknown. This paper reports that histamine inhibited I(Ca) in cultured bovine adrenal chromaffin cells and this response was blocked by the H(1) antagonist mepyramine. With high levels of calcium buffering in the patch pipette solution (10 mM EGTA), histamine slowed the activation kinetics and inhibited the amplitude of I(Ca). A conditioning prepulse to +100 mV reversed the kinetic slowing and partially relieved the inhibition. These features are characteristic of a membrane delimited, voltage-dependent pathway which is thought to involve direct binding of G-protein betagamma subunits to the Ca channels. However, unlike virtually every other example of this type of inhibition, the response to histamine was not blocked by pretreating the cells with pertussis toxin (PTX). The voltage-dependent, PTX insensitive inhibition produced by histamine was modest compared with the PTX sensitive inhibition produced by ATP (28% vs. 53%). When histamine and ATP were applied concomitantly there was no additivity of the inhibition beyond that produced by ATP alone (even though the agonists appear to activate distinct G-proteins) suggesting that the inhibition produced by ATP is maximal. When experiments were carried out under conditions of low levels of calcium buffering in the patch pipette solution (0.1 mM EGTA), histamine inhibited I(Ca) in some cells using an entirely voltage insensitive pathway. We demonstrate that activation of PTX insensitive G-proteins (most likely Gq) by H(1) receptors inhibits I(Ca). This may represent a mechanism by which histamine exerts inhibitory (in addition to previously identified stimulatory) effects on catecholamine release. PMID- 10712471 TI - Sodium pump activity, not glial spatial buffering, clears potassium after epileptiform activity induced in the dentate gyrus. AB - A number of mechanisms have been proposed to play a role in the regulation of activity-dependent variations in extracellular potassium concentration ([K(+)](o)). We tested possible regulatory mechanisms for [K(+)](o) during spontaneous recurrent epileptiform activity induced in the dentate gyrus of hippocampal slices from adult rats by perfusion with 8 mM potassium and 0-added calcium medium in an interface chamber. Local application of tetrodotoxin blocked local [K(+)](o) changes, suggesting that potassium is released and taken up locally. Perfusion with barium or cesium, blockers of the inward rectifying potassium channel, did not alter the baseline [K(+)](o), the ceiling level of [K(+)](o) reached during the burst, or the rate of [K(+)](o) recovery after termination of the bursts. Decreasing gap junctional conductance did not change the baseline [K(+)](o) or the half-time of recovery of the [K(+)](o) after the bursts but did cause a decrease in the ceiling level of [K(+)](o). Perfusion with furosemide, which will block cation/chloride cotransporters, or perfusion with low chloride did not change the baseline [K(+)](o) or the half-time of recovery of the [K(+)](o) after the bursts but did increase the ceiling level of [K(+)](o). Bath or local application of ouabain, a Na(+)/K(+)-ATPase inhibitor, increased the baseline [K(+)](o), slowed the rate of [K(+)](o) recovery, and induced spreading depression. These findings suggest that potassium redistribution by glia only plays a minor role in the regulation of [K(+)](o) in this model. The major regulator of [K(+)](o) in this model appears to be uptake via a Na(+)/K(+)-ATPase, most likely neuronal. PMID- 10712472 TI - Spontaneous activity in the statoacoustic ganglion of the chicken embryo. AB - Statoacoustic ganglion cells in the mature bird include neurons that are responsive to sound (auditory) and those that are not (nonauditory). Those that are nonauditory have been shown to innervate an otolith organ, the macula lagena, whereas auditory neurons innervate the basilar papilla. In the present study, single-unit recordings of statoacoustic ganglion cells were made in embryonic (E19, mean = 19.2 days of incubation) and hatchling (P6-P14, mean = 8.6 days posthatch) chickens. Spontaneous activity from the two age groups was compared with developmental changes. Activity was evaluated for 47 auditory, 11 nonauditory, and 6 undefined eighth nerve neurons in embryos and 29 auditory, 26 nonauditory, and 1 undefined neurons in hatchlings. For auditory neurons, spontaneous activity displayed an irregular pattern [discharge interval coefficient of variation (CV) was >0.5, mean CV for embryos was 1.46 +/- 0.58 and for hatchlings was 1.02 +/- 0.25; means +/- SD]. Embryonic discharge rates ranged from 0.05 to 97.6 spikes per second (sp/s) for all neurons (mean 18.6 +/- 16.9 sp/s). Hatchling spontaneous rates ranged from 1.2 to 185.2 sp/s (mean 66.5 +/- 39.6 sp/s). Discharge rates were significantly higher for hatchlings (P < 0.001). Many embryonic auditory neurons displayed long silent periods between irregular bursts of neural activity, a feature not seen posthatch. All regular bursting discharge patterns were correlated with heart rate in both embryos and hatchlings. Preferred intervals were visible in the time interval histograms (TIHs) of only one embryonic neuron in contrast to 55% of the neurons in posthatch animals. Generally, the embryonic auditory TIH displayed a modified quasi-Poisson distribution. Nonauditory units generally displayed regular (CV <0.5) or irregular (CV >0.5) activity and Gaussian and modified-Gaussian TIHs. Long silent periods or bursting patterns were not a characteristic of embryonic nonauditory neurons. CV varied systematically as a function of discharge rate in nonauditory but not auditory primary afferents. Minimum spike intervals (dead time) and interval modes for auditory neurons were longer in embryos (dead time: embryos 2.88 +/- 6.85 ms; hatchlings 1.50 +/- 1.76 ms; modal intervals: embryo 10.09 +/- 22.50 ms, hatchling 3.54 +/- 3.29 ms). The results show that significant developmental changes occur in spontaneous activity between E19 and posthatch. It is likely that both presynaptic and postsynaptic changes in the neuroepithelium contribute to maturational refinements during this period of development. PMID- 10712473 TI - Predictive modulation of muscle coordination pattern magnitude scales fingertip force magnitude over the voluntary range. AB - Human fingers have sufficiently more muscles than joints such that every fingertip force of submaximal magnitude can be produced by an infinite number of muscle coordination patterns. Nevertheless, the nervous system seems to effortlessly select muscle coordination patterns when sequentially producing fingertip forces of low, moderate, and maximal magnitude. The hypothesis of this study is that the selection of coordination patterns to produce submaximal forces is simplified by the appropriate modulation of the magnitude of a muscle coordination pattern capable of producing the largest expected fingertip force. In each of three directions, eight subjects were asked to sequentially produce fingertip forces of low, moderate, and maximal magnitude with their dominant forefinger. Muscle activity was described by fine-wire electromyograms (EMGs) simultaneously collected from all muscles of the forefinger. A muscle coordination pattern was defined as the vector list of the EMG activity of each muscle. For all force directions, statistically significant muscle coordination patterns similar to those previously reported for 100% of maximal fingertip forces were found for 50% of maximal voluntary force. Furthermore the coordination pattern and fingertip force vector magnitudes were highly correlated (r > 0.88). Average coordination pattern vectors at 50 and 100% of maximal force were highly correlated with each other, as well as with individual coordination pattern vectors in the ramp transitions preceding them. In contrast to this consistency of EMG coordination patterns, predictions using a musculoskeletal computer model of the forefinger show that force magnitudes ATP > alpha,beta -MeATP for all three expressed splice variants. The ATP receptor antagonists suramin and PPADS reduced the effects of ATP on all three variants. Results demonstrate that three P2X(2) splice variants from guinea pig cochlea, P2X(2-1), P2X(2-2), and P2X(2-3), can individually form nonselective cation receptor channels when these subunits are expressed in HEK293 cells. The distinct properties of these P2X(2) receptor splice variants may contribute to the differences in the response to ATP observed in native cochlear cells. PMID- 10712476 TI - Regionally selective blockade of GABAergic inhibition by zinc in the thalamocortical system: functional significance. AB - The thalamocortical (TC) system is a tightly coupled synaptic circuit in which GABAergic inhibition originating from the nucleus reticularis thalami (NRT) serves to synchronize oscillatory TC rhythmic behavior. Zinc is colocalized within nerve terminals throughout the TC system with dense staining for zinc observed in NRT, neocortex, and thalamus. Whole cell voltage-clamp recordings of GABA-evoked responses were conducted in neurons isolated from ventrobasal thalamus, NRT, and somatosensory cortex to investigate modulation of the GABA mediated chloride conductance by zinc. Zinc blocked GABA responses in a regionally specific, noncompetitive manner within the TC system. The regional levels of GABA blockade efficacy by zinc were: thalamus > NRT > cortex. The relationship between clonazepam and zinc sensitivity of GABA(A)-mediated responses was examined to investigate possible presence or absence of specific GABA(A) receptor (GABAR) subunits. These properties of GABARs have been hypothesized previously to be dependent on presence or absence of the gamma2 subunit and seem to display an inverse relationship. In cross-correlation plots, thalamic and NRT neurons did not show a statistically significant relationship between clonazepam and zinc sensitivity; however, a statistically significant correlation was observed in cortical neurons. Spontaneous epileptic TC oscillations can be induced in vitro by perfusion of TC slices with an extracellular medium containing no added Mg(2+). Multiple varieties of oscillations are generated, including simple TC burst complexes (sTBCs), which resemble spike-wave discharge activity. A second variant was termed a complex TC burst complex (cTBC), which resembled generalized tonic clonic seizure activity. sTBCs were exacerbated by zinc, whereas cTBCs were blocked completely by zinc. This supported the concept that zinc release may modulate TC rhythms in vivo. Zinc interacts with a variety of ionic conductances, including GABAR currents, N methyl-D-aspartate (NMDA) receptor currents, and transient potassium (A) currents. D-2-amino-5-phosphonovaleric acid and 4-aminopyridine blocked both s- and cTBCs in TC slices. Therefore NMDA and A current-blocking effects of zinc are insufficient to explain differential zinc sensitivity of these rhythms. This supports a significant role of zinc-induced GABAR modulation in differential TC rhythm effects. Zinc is localized in high levels within the TC system and appears to be released during TC activity. Furthermore application of exogenous zinc modulates TC rhythms and differentially blocks GABARs within the TC system. These data are consistent with the hypothesis that endogenously released zinc may have important neuromodulatory actions impacting generation of TC rhythms, mediated at least in part by effects on GABARs. PMID- 10712477 TI - Canal-otolith interactions after off-vertical axis rotations I. Spatial reorientation of horizontal vestibuloocular reflex. AB - We examined the three-dimensional (3-D) spatial orientation of postrotatory eye velocity after horizontal off-vertical axis rotations by varying the final body orientation with respect to gravity. Three rhesus monkeys were oriented in one of two positions before the onset of rotation: pitched 24 degrees nose-up or 90 degrees nose-up (supine) relative to the earth-horizontal plane and rotated at +/ 60 degrees /s around the body-longitudinal axis. After 10 turns, the animals were stopped in 1 of 12 final positions separated by 30 degrees. An empirical analysis of the postrotatory responses showed that the resultant response plane remained space-invariant, i.e., accurately represented the actual head tilt plane at rotation stop. The alignment of the response vector with the spatial vertical was less complete. A complementary analysis, based on a 3-D model that implemented the spatial transformation and dynamic interaction of otolith and lateral semicircular canal signals, confirmed the empirical description of the spatial response. In addition, it allowed an estimation of the low-pass filter time constants in central otolith and semicircular canal pathways as well as the weighting ratio between direct and inertially transformed canal signals in the output. Our results support the hypothesis that the central vestibular system represents head velocity in gravity-centered coordinates by sensory integration of otolith and semicircular canal signals. PMID- 10712478 TI - Local infusion of scopolamine into intraparietal cortex slows covert orienting in rhesus monkeys. AB - There is accumulating evidence to suggest that cholinergic neurotransmission may play an important role in visuospatial attention, but the brain sites at which acetylcholine modulates attention are not well understood. The present work tested the hypothesis that the cholinergic influences within the intraparietal cortex are necessary for normal attentional shifting (covert orienting) in nonhuman primates. Two rhesus monkeys were trained to perform a visual, cued target detection task for liquid reinforcement. The animals pressed a lever to produce a visual display in which a central fixation point was flanked by two circles. Shortly after fixation was established, one of the circles brightened (cue), and a target appeared subsequently within one of the circles. Detection was signaled by a manual response and the reaction time to the appearance of the target was recorded. Four types of trials were presented. For valid cue trials, the cue and target were at the same spatial location; for invalid cues, cue and target were in opposite hemifields; for double cues, both cues were brightened but the target appeared in either the left or right circle; in no-cue trials, the cue was omitted. We localized the intraparietal region by recording attention related, cellular activity with intracerebral microelectrodes. Among visually responsive cells in this area, valid cues presented to the receptive fields of visual neurons enhanced the responses to target stimuli in about half the cells and inhibited those responses in the remainder. In addition, some cells showed longer response latencies to invalid cues than to valid cues. We then infused scopolamine into attention-related activity sites and assessed its effect on performance. Scopolamine produced a dose-dependent increase in reaction times and decrease in performance accuracy that lasted more than 1 h. Neither vehicle injections in the same locations nor scopolamine outside the physiologically defined area produced any significant change in behavior. Under our conditions of measurement, we conclude that activity mediated by muscarinic cholinergic receptors within the intraparietal cortex is necessary for normal covert orienting. PMID- 10712479 TI - Inactivation of parietal and prefrontal cortex reveals interdependence of neural activity during memory-guided saccades. AB - Dorsolateral prefrontal and posterior parietal cortex share reciprocal projections. They also share nearly identical patterns of neuronal activation during performance of memory-guided saccades. To test the hypothesis that the reciprocal projections between parietal and prefrontal neurons may entrain their parallel activation, the present experiments have combined cortical cooling in one cortical area with single-unit recording in the other to more precisely determine the physiological interactions between the two during working memory performance. The activity of 105 cortical neurons during the performance of an oculomotor delayed response (ODR) task (43 parietal neurons during prefrontal cooling, 62 prefrontal neurons during parietal cooling) was compared across two blocks of trials collected while the distant cortical area either was maintained at normal body temperature or cooled. The mean firing rates of 71% of the prefrontal neurons during ODR performance changed significantly when parietal cortex was cooled. Prefrontal neurons the activity of which was modulated during the cue, delay, or saccade periods of the task were equally vulnerable to parietal inactivation. Further, both lower and higher firing rates relative to the precool period were seen with comparable frequency. Similar results were obtained from the converse experiment, in which the mean firing rates of 76% of the parietal neurons were significantly different while prefrontal cortex was cooled, specifically in those task epochs when the activity of each neuron was modulated during ODR performance. These effects again were seen equally in all epochs of the ODR task in the form of augmented or suppressed activity. Significant effects on the latency of neuronal activation during cue and saccade periods of the task were absent irrespective of the area cooled. Cooling was associated in some cases with a shift in the best direction of Gaussian tuning functions fit to neuronal activity, and these shifts were on average larger during parietal than prefrontal cooling. In view of the parallel between the similarity in activity patterns previously reported and the largely symmetrical cooling effects presently obtained, the data suggest that prefrontal and parietal neurons achieve matched activation during ODR performance through a symmetrical exchange of neuronal signals between them; in both cortical areas, neurons activated during the cue, delay, and also saccade epochs of the ODR task participate in reciprocal neurotransmission; and the output of each cortical area produces a mixture of excitatory and inhibitory drives within its target. PMID- 10712480 TI - Role of cAMP in the short-term modulation of a neuromuscular system in aplysia. AB - Neuromuscular synapses in buccal muscle I3a of Aplysia are modulated by the small cardioactive peptide (SCP), a peptide cotransmitter that is intrinsic to the motor neurons, and by serotonin (5-HT) released from modulatory neurons that are extrinsic to the motor circuit. Although the modulation of excitatory junction potentials (EJPs) and contractions by 5-HT and SCP has been studied extensively in this muscle, little is known about the mechanisms that underlie the modulation. 5-HT and SCP, at 1 microM, were found to potently increase the level of cAMP in I3a. Therefore we investigated whether the activation of the cAMP pathway was sufficient to modulate EJPs and contractions. The direct activation of adenylyl cyclase with forskolin increased the level of cAMP, facilitated EJPs, and potentiated contractions. Indeed, the short-term effects of forskolin were very similar to all aspects of the short-term effects of 5-HT and SCP. Membrane permeable cAMP analogues also mimicked the effects of 5-HT and SCP on EJPs and contractions. However, it seems likely that some effects of 5-HT are also mediated through other second-messenger pathways because low concentrations of 5 HT modulate EJPs and contractions but do not significantly increase cAMP levels in I3a. It is possible that lower concentrations of 5-HT function through receptors linked to protein kinase C (PKC) because phorbol, an activator of PKC, modulated EJPs and contractions without increasing the levels of cAMP. In conclusion, we provide evidence that pharmacological agents that activate the cAMP pathway mimicked most of the effects of 5-HT or SCP and that more than one second-messenger system appears to be involved in the modulation of the I3a neuromuscular system. PMID- 10712481 TI - Chronic NMDA exposure accelerates development of GABAergic inhibition in the superior colliculus. AB - Maturation of excitatory synaptic connections depends on the amount and pattern of their activity, and activity can affect development of inhibitory synapses as well. In the superficial visual layers of the superior colliculus (sSC), developmental increases in the effectiveness of gamma-aminobutyric acid (GABA(A)) receptor-mediated inhibition may be driven by the maturation of visual inputs. In the rat sSC, GABA(A) receptor currents significantly jump in amplitude between postnatal days 17 and 18 (P17 and P18), approximately when the effects of cortical inputs are first detected in collicular neurons. We manipulated the development of these currents in vivo by implanting a drug-infused slice of the ethylene-vinyl acetate copolymer Elvax over the superior colliculus of P8 rats to chronically release from this plastic low levels of N-methyl-D-aspartate (NMDA). Sham-treated control animals received a similar implant containing only the solvent for NMDA. To examine the effects of this treatment on the development of GABA-mediated neurotransmission, we used whole cell voltage-clamp recording of spontaneous synaptic currents (sPSCs) from sSC neurons in untreated, NMDA treated, and sham-treated superior colliculus slices ranging in age from 10 to 20 days postnatal. Both amplitude and frequency of sPSCs were studied at holding potentials of +50 mV in the presence and absence of the GABA(A) receptor antagonist, bicuculline methiodide (BMI). The normal developmental increase in GABA(A) receptor currents occurred on schedule (P18) in sham-treated sSC, but NMDA treatment caused premature up-regulation (P12). The average sPSCs in early NMDA-treated neurons were significantly larger than in age-matched sham controls or in age-matched, untreated neurons. No differences in average sPSC amplitudes across treatments or ages were present in BMI-insensitive, predominantly glutamatergic synaptic currents of the same neurons. NMDA treatment also significantly increased levels of glutamate decarboxylase (GAD), measured by quantitative western blotting with staining at P13 and P19. Cell counting using the dissector method for MAP 2 and GAD(67) at P13 and P19 indicated that the differences in GABAergic transmission were not due to increases in the proportion of inhibitory to excitatory neurons after NMDA treatment. However, chronic treatments begun at P8 with Elvax containing both NMDA and BMI significantly decreased total neuron density at P19 ( approximately 15%), suggesting that the NMDA-induced increase in GABA(A) receptor currents may protect against excitotoxicity. PMID- 10712482 TI - Rapid activation of GABAergic interneurons and possible calcium independent GABA release in the mormyrid electrosensory lobe. AB - The primary afferent fibers from the electroreceptors of mormyrid electric fish terminate centrally in the granular layer of the electrosensory lobe (ELL). This study examines the excitatory and inhibitory processes that take place in this layer using an in vitro slice preparation and field potentials evoked by stimulation of primary afferent fibers in the deep fiber layer of ELL. The postsynaptic response to stimulation of the afferent fibers was still present after blocking chemical transmission in three different ways: by adding glutamate receptor antagonists to the medium, by substituting a nominally calcium-free medium for normal medium, and by blocking calcium channels with cadmium. Blockade of chemical transmission was demonstrated by disappearance of control responses to parallel fiber stimulation. The continued presence of a postsynaptic response in the absence of chemical excitation is consistent with previous anatomic and physiological evidence for electrical synapses between afferent fibers and granular cells in ELL. Granular cell activation by primary afferent fibers was followed by a powerful, short-latency inhibition mediated by GABA and GABA(A) receptors, as indicated by a large increase in the postsynaptic response to afferent fiber stimulation following application of the GABA(A) receptor antagonist, bicuculline. Bicuculline caused a marked increase of the postsynaptic response even after chemical synaptic excitation had been blocked by glutamate receptor antagonists, by a calcium-free medium, or by cadmium. Thus activation of the inhibitory interneurons responsible for GABA release did not require chemical excitation. Nonchemical excitation of the inhibitory interneurons could be mediated either by electrical synapses between afferent fibers and inhibitory interneurons, or by nonsynaptic activation of the large GABAergic terminals that are known to be present on granular cells. The marked increase of the postsynaptic response caused by bicuculline in a calcium-free medium or in the presence of cadmium suggests that the release of GABA by inhibitory terminals was not entirely dependent on calcium influx. This effect of bicuculline on the postsynaptic response in a calcium-free medium or in the presence of cadmium was markedly reduced by prior addition of the GABA transporter antagonist, nipecotic acid. Thus calcium-independent release of GABA may occur in ELL and may be partly dependent on reversal of a GABA transporter. Rapid and powerful inhibition at the first stage in the processing of electrosensory information could serve to enhance the small differences in latency among afferent fibers that appear to encode small differences in stimulus intensity. PMID- 10712483 TI - Diverse synaptic connections between peptidergic radula mechanoafferent neurons and neurons in the feeding system of Aplysia. AB - The buccal ganglion of Aplysia contains a heterogeneous population of peptidergic, radula mechanoafferent (RM) neurons. To investigate their function, two of the larger RM cells (B21, B22) were identified by morphological and electrophysiological criteria. Both are low-threshold, rapidly adapting, mechanoafferent neurons that responded to touch of the radula, the structure that grasps food during ingestive and egestive feeding movements. Sensory responses of the cells consisted of spike bursts at frequencies of 8-35 Hz. Each cell was found to make chemical, electrical, or combined synapses with other sensory neurons, motor neurons and interneurons involved in radula closure and/or protraction-retraction movements of the odontophore. Motor neurons receiving input included the following: B8a/b, B15, and B16, which innervate muscles contributing to radula closing; and B82, a newly identified neuron that innervates the anterodorsal region of the I1/I3 muscles of the buccal mass. B21 and B22 can affect buccal motor programs by way of their connections to interneurons such as B19 and B64. Fast, chemical, excitatory postsynaptic potentials (EPSPs) produced by RM neurons, such as B21, exhibited strong, frequency-dependent facilitation, a form of homosynaptic plasticity. Firing B21 also produced a slow EPSP in B15 that increased the excitability of the cell. Thus a sensory neuron mediating a behavioral response may have modulatory effects. The data suggest multiple functions for RM neurons including 1) triggering of phase transitions in rhythmic motor programs, 2) adjusting the force of radula closure, 3) switching from biting to swallowing or swallowing to rejection, and 4) enhancing food-induced arousal. PMID- 10712484 TI - Outputs of radula mechanoafferent neurons in Aplysia are modulated by motor neurons, interneurons, and sensory neurons. AB - The gain of sensory inputs into the nervous system can be modulated so that the nature and intensity of afferent input is variable. Sometimes the variability is a function of other sensory inputs or of the state of motor systems that generate behavior. A form of sensory modulation was investigated in the Aplysia feeding system at the level of a radula mechanoafferent neuron (B21) that provides chemical synaptic input to a group of motor neurons (B8a/b, B15) that control closure and retraction movements of the radula, a food grasping structure. B21 has been shown to receive both excitatory and inhibitory synaptic inputs from a variety of neuron types. The current study investigated the morphological basis of these heterosynaptic inputs, whether the inputs could serve to modulate the chemical synaptic outputs of B21, and whether the neurons producing the heterosynaptic inputs were periodically active during feeding motor programs that might modulate B21 outputs in a phase-specific manner. Four cell types making monosynaptic connections to B21 were found capable of heterosynaptically modulating the chemical synaptic output of B21 to motor neurons B8a and B15. These included the following: 1) other sensory neurons, e.g. , B22; 2) interneurons, e.g., B19; 3) motor neurons, e.g., B82; and 4) multifunction neurons that have sensory, motor, and interneuronal functions, e.g., B4/5. Each cell type was phasically active in one or more feeding motor programs driven by command-like interneurons, including an egestive motor program driven by CBI-1 and an ingestive motor program driven by CBI-2. Moreover, the phase of activity differed for each of the modulator cells. During the motor programs, shifts in B21 membrane potential were related to the activity patterns of some of the modulator cells. Inhibitory chemical synapses mediated the modulation produced by B4/5, whereas excitatory and/or electrical synapses were involved in the other instances. The data indicate that modulation is due to block of action potential invasion into synaptic release regions or to alterations of transmitter release as a function of the presynaptic membrane potential. The results indicate that just as the motor system of Aplysia can be modulated by intrinsic mechanisms that can enhance its efficiency, the properties of primary sensory cells can be modified by diverse inputs from mediating circuitry. Such modulation could serve to optimize sensory cells for the different roles they might play. PMID- 10712485 TI - Primate translational vestibuloocular reflexes. I. High-frequency dynamics and three-dimensional properties during lateral motion. AB - The dynamics and three-dimensional (3-D) properties of the primate translational vestibuloocular reflex (trVOR) for high-frequency (4-12 Hz, +/-0.3-0.4 g) lateral motion were investigated during near-target viewing at center and eccentric targets. Horizontal response gains increased with frequency and depended on target eccentricity. The larger the horizontal and vertical target eccentricity, the steeper the dependence of horizontal response gain on frequency. In addition to horizontal eye movements, robust torsional response components also were present at all frequencies. During center-target fixation, torsional response phase was opposite (anticompensatory) to that expected for an "apparent" tilt response. Instead torsional response components depended systematically on vertical-target eccentricity, increasing in amplitude when looking down and reversing phase when looking up. As a result the trVOR eye velocity vector systematically tilted away from a purely horizontal direction, through an angle that increased with vertical eccentricity with a slope of approximately 0.7. This systematic dependence of torsional eye velocity tilt on vertical eye position suggests that the trVOR might follow the 3-D kinematic requirements that have been shown to govern visually guided eye movements and near-target fixation. PMID- 10712486 TI - Primate translational vestibuloocular reflexes. II. Version and vergence responses to fore-aft motion. AB - To maintain binocular fixation on near targets during fore-aft translational disturbances, largely disjunctive eye movements are elicited the amplitude and direction of which should be tuned to the horizontal and vertical eccentricities of the target. The eye movements generated during this task have been investigated here as trained rhesus monkeys fixated isovergence targets at different horizontal and vertical eccentricities during 10 Hz fore-aft oscillations. The elicited eye movements complied with the geometric requirements for binocular fixation, although not ideally. First, the corresponding vergence angle for which the movement of each eye would be compensatory was consistently less than that dictated by the actual fixation parameters. Second, the eye position with zero sensitivity to translation was not straight ahead, as geometrically required, but rather exhibited a systematic dependence on viewing distance and vergence angle. Third, responses were asymmetric, with gains being larger for abducting and downward compared with adducting and upward gaze directions, respectively. As frequency was varied between 4 and 12 Hz, responses exhibited high-pass filter properties with significant differences between abduction and adduction responses. As a result of these differences, vergence sensitivity increased as a function of frequency with a steeper slope than that of version. Despite largely undercompensatory version responses, vergence sensitivity was closer to ideal. Moreover, the observed dependence of vergence sensitivity on vergence angle, which was varied between 2.5 and 10 MA, was largely linear rather than quadratic (as geometrically predicted). We conclude that the spatial tuning of eye velocity sensitivity as a function of gaze and viewing distance follows the general geometric dependencies required for the maintenance of foveal visual acuity. However, systematic deviations from ideal behavior exist that might reflect asymmetric processing of abduction/adduction responses perhaps because of different functional dependencies of version and vergence eye movement components during translation. PMID- 10712487 TI - Primate translational vestibuloocular reflexes. III. Effects of bilateral labyrinthine electrical stimulation. AB - The effects of functional, reversible ablation and potential recruitment of the most irregular otolith afferents on the dynamics and sensitivity of the translational vestibuloocular reflexes (trVORs) were investigated in rhesus monkeys trained to fixate near and far targets. Translational motion stimuli consisted of either steady-state lateral and fore-aft sinusoidal oscillations or short-lasting transient lateral head displacements. Short-duration (usually <2 s) anodal (inhibitory) and cathodal (excitatory) currents (50-100 microA) were delivered bilaterally during motion. In the presence of anodal labyrinthine stimulation, trVOR sensitivity and its dependence on viewing distance were significantly decreased. In addition, anodal currents significantly increased phase lags. During transient motion, anodal stimulation resulted in significantly lower initial eye acceleration and more sluggish responses. Cathodal currents tended to have opposite effects. The main characteristics of these results were simulated by a simple model where both regularly and irregularly discharging afferents contribute to the trVORs. Anodal labyrinthine currents also were found to decrease eye velocity during long-duration, constant velocity rotations, although results were generally more variable compared with those during translational motion. PMID- 10712488 TI - Response differences in monkey TE and perirhinal cortex: stimulus association related to reward schedules. AB - Anatomic and behavioral evidence shows that TE and perirhinal cortices are two directly connected but distinct inferior temporal areas. Despite this distinctness, physiological properties of neurons in these two areas generally have been similar with neurons in both areas showing selectivity for complex visual patterns and showing response modulations related to behavioral context in the sequential delayed match-to-sample (DMS) trials, attention, and stimulus familiarity. Here we identify physiological differences in the neuronal activity of these two areas. We recorded single neurons from area TE and perirhinal cortex while the monkeys performed a simple behavioral task using randomly interleaved visually cued reward schedules of one, two, or three DMS trials. The monkeys used the cue's relation to the reward schedule (indicated by the brightness) to adjust their behavioral performance. They performed most quickly and most accurately in trials in which reward was immediately forthcoming and progressively less well as more intermediate trials remained. Thus the monkeys appeared more motivated as they progressed through the trial schedule. Neurons in both TE and perirhinal cortex responded to both the visual cues related to the reward schedules and the stimulus patterns used in the DMS trials. As expected, neurons in both areas showed response selectivity to the DMS patterns, and significant, but small, modulations related to the behavioral context in the DMS trial. However, TE and perirhinal neurons showed strikingly different response properties. The latency distribution of perirhinal responses was centered 66 ms later than the distribution of TE responses, a larger difference than the 10-15 ms usually found in sequentially connected visual cortical areas. In TE, cue-related responses were related to the cue's brightness. In perirhinal cortex, cue-related responses were related to the trial schedules independently of the cue's brightness. For example, some perirhinal neurons responded in the first trial of any reward schedule including the one trial schedule, whereas other neurons failed to respond in the first trial but respond in the last trial of any schedule. The majority of perirhinal neurons had more complicated relations to the schedule. The cue-related activity of TE neurons is interpreted most parsimoniously as a response to the stimulus brightness, whereas the cue-related activity of perirhinal neurons is interpreted most parsimoniously as carrying associative information about the animal's progress through the reward schedule. Perirhinal cortex may be part of a system gauging the relation between work schedules and rewards. PMID- 10712489 TI - Action potential reflection and failure at axon branch points cause stepwise changes in EPSPs in a neuron essential for learning. AB - In leech mechanosensory neurons, action potentials reverse direction, or reflect, at central branch points. This process enhances synaptic transmission from individual axon branches by rapidly activating synapses twice, thereby producing facilitation. At the same branch points action potentials may fail to propagate, which can reduce transmission. It is now shown that presynaptic action potential reflection and failure under physiological conditions influence transmission to the same postsynaptic neuron, the S cell. The S cell is an interneuron essential for a form of nonassociative learning, sensitization of the whole body shortening reflex. The P to S synapse has components that appear monosynaptic (termed "direct") and polysynaptic, both with glutamatergic pharmacology. Reflection at P cell branch points on average doubled transmission to the S cell, whereas action potential failure, or conduction block, at the same branch points decreased it by one-half. Each of two different branch points affected transmission, indicating that the P to S connection is spatially distributed around these branch points. This was confirmed by examining the locations of individual contacts made by the P cell with the S cell and its electrically coupled partner C cells. These results show that presynaptic neuronal morphology produces a range of transmission states at a set of synapses onto a neuron necessary for a form of learning. Reflection and conduction block are activity-dependent and are basic properties of action potential propagation that have been seen in other systems, including axons and dendrites in the mammalian brain. Individual branch points and the distribution of synapses around those branch points can substantially influence neuronal transmission and plasticity. PMID- 10712490 TI - Fast reaction to different sensory modalities activates common fields in the motor areas, but the anterior cingulate cortex is involved in the speed of reaction. AB - We examined which motor areas would participate in the coding of a simple opposition of the thumb triggered by auditory, somatosensory and visual signals. We tested which motor areas might be active in response to all three modalities, which motor structures would be activated specifically in response to each modality, and which neural populations would be involved in the speed of the reaction. The subjects were required to press a button with their right thumb as soon as they detected a change in the sensory signal. The regional cerebral blood flow (rCBF) was measured quantitatively with (15)O-butanol and positron emission tomography (PET) in nine normal male subjects. Cytoarchitectural areas were delimited in 10 post mortem brains by objective and quantitative methods. The images of the post mortem brains subsequently were transformed into standard anatomic format. One PET scanning for each of the sensory modalities was done. The control condition was rest with the subjects having their eyes closed. The rCBF images were anatomically standardized, and clusters of significant changes in rCBF were identified. These were localized to motor areas delimited on a preliminary basis, such as supplementary motor area (SMA), dorsal premotor zone (PMD), rostral cingulate motor area (CMAr), and within areas delimited by using microstructural i.e., cytoarchitectonic criteria, such as areas 4a, 4p, 3a, 3b, and 1. Fields of activation observed as a main effect for all three modalities were located bilaterally in the SMA, CMAr, contralateral PMD, primary motor (M1), and primary somatosensory cortex (SI). The activation in M1 engaged areas 4a and 4p and expanded into area 6. The activation in SI engaged areas 3b, 1, and extended into somatosensory association areas and the supramarginal gyrus posteriorly. We identified significant activations that were specific for each modality in the respective sensory association cortices, though no modality specific regions were found in the motor areas. Fields in the anterior cingulate cortex, rostral to the CMAr, consistently showed significant negative correlation with mean reaction time (RT) in all three tasks. These results show that simple reaction time tasks activate many subdivisions of the motor cortices. The information from different sensory modalities converge onto the common structures: the contralateral areas 4a, 4p, 3b, 1, the PMD, and bilaterally on the SMA and the CMAr. The anterior cingulate cortex might be a key structure which determine the speed of reaction in simple RT tasks. PMID- 10712491 TI - Ca(2+)- and metabolism-related changes of mitochondrial potential in voltage clamped CA1 pyramidal neurons in situ. AB - In hippocampal slices from rats, dialysis with rhodamine-123 (Rh-123) and/or fura 2 via the patch electrode allowed monitoring of mitochondrial potential (DeltaPsi) changes and intracellular Ca(2+) ([Ca(2+)](i)) of CA1 pyramidal neurons. Plasmalemmal depolarization to 0 mV caused a mean [Ca(2+)](i) rise of 300 nM and increased Rh-123 fluorescence signal (RFS) by /=20 microm from the soma had little effect on the amplitude of the sAHP recorded in cortical pyramidal cells. By this process of elimination, it is suggested that sAHP channels may be concentrated in the basal dendrites of CA1 pyramids. PMID- 10712496 TI - Ratio of inhibited-to-activated pallidal neurons decreases dramatically during passive limb movement in the MPTP-treated monkey. AB - Mink advanced the hypothesis in 1996 that the role of the basal ganglia (BG) is primarily one of focused selection; the encouragement of motor mechanisms inducing a desired movement and the inhibition of competing mechanisms. This would imply, in normal subjects, a ratio of inhibited-to-activated (I/A) movement related globus pallidus pars internalis (GPi) neurons <1 and a drastic decrease of this ratio in the parkinsonian state. 1-Methyl-4-phenyl-1,2,3,6 tetrahydropyridine (MPTP) intoxication should therefore decrease the specificity of the response of this neuronal population. To test this working hypothesis we studied the activity of GPi neurons in response to passive limb movement in the normal and the parkinsonian monkey. Extracellular unit recordings monitored any correlation between passive limb movements and eventual modifications of the neuronal activity of the GPi in two calm, awake, and drug naive monkeys (Macaca fascicularis) before and after MPTP intoxication. In the normal animal, arm- and leg-related neurons were located in clusters in the medial part of the GPi. The I/A ratio was 0.22. Most GPi cells were linked to a single joint. In the MPTP treated monkey, the number of movement-related neurons increased, the I/A ratio dropped significantly to 0.03, and most responding cells were linked to several joints. These data, which cannot be explained by the classic "box" model, endorse Mink's hypothesis. PMID- 10712497 TI - Rostrocaudal distinction of the dorsal premotor area based on oculomotor involvement. AB - To investigate functional differences between the rostral and caudal parts of the dorsal premotor cortex (PMd), we first examined the effects of intracortical microstimulation (ICMS) while monkeys were performing oculomotor and limb motor tasks or while they were at rest. We found that saccades were evoked from the rostral part (PMdr) whereas ICMS in the caudal part (PMdc) predominantly produced forelimb or body movements. Subsequently, we examined neuronal activity in relation to the performance of visually cued and memorized saccades while monkeys reached an arm toward a visual target. We found that roughly equal numbers of PMdr neurons were active during performance of the oculomotor and limb motor tasks. In contrast, the majority of PMdc neurons were related preferentially to arm movements and not to saccades. In the subsequent analysis, we found that the oculomotor effects evoked in the PMdr differ from the effects evoked in either the frontal eye field (FEF) or supplementary eye field (SEF). These findings suggest that the PMdr is involved in oculomotor as well as limb motor behavior. However, the oculomotor involvement of the PMdr seems to have a functional aspect different from that operating in the FEF and SEF. PMID- 10712498 TI - Differential organization of touch and pain in human primary somatosensory cortex. AB - Processing of tactile stimuli within somatosensory cortices has been shown to be complex and hierarchically organized. However, the precise organization of nociceptive processing within these cortices has remained largely unknown. We used whole-head magnetoencephalography to directly compare cortical responses to stimulation of tactile and nociceptive afferents of the dorsum of the hand in humans. Within the primary somatosensory cortex (SI), nociceptive stimuli activated a single source whereas tactile stimuli activated two sequentially peaking sources. Along the postcentral gyrus, the nociceptive SI source was located 10 mm more medially than the early tactile SI response arising from cytoarchitectonical area 3b and corresponded spatially to the later tactile SI response. Considering a mediolateral location difference between the hand representations of cytoarchitectonical areas 3b and 1, the present results suggest generation of the single nociceptive response in area 1, whereas tactile stimuli activate sequentially peaking sources in areas 3b and 1. Thus nociceptive processing apparently does not share the complex and hierarchical organization of tactile processing subserving elaborated sensory capacities. This difference in the organization of both modalities may reflect that pain perception rather requires reactions to and avoidance of harmful stimuli than sophisticated sensory capacities. PMID- 10712499 TI - Ernest everett just lecture, 1999. The value of mentoring in the career of a young scientist PMID- 10712500 TI - Half a century of "the nuclear matrix". AB - A cell fraction that would today be termed "the nuclear matrix" was first described and patented in 1948 by Russian investigators. In 1974 this fraction was rediscovered and promoted as a fundamental organizing principle of eukaryotic gene expression. Yet, convincing evidence for this functional role of the nuclear matrix has been elusive and has recently been further challenged. What do we really know about the nonchromatin elements (if any) of internal nuclear structure? Are there objective reasons (as opposed to thinly veiled disdain) to question experiments that use harsh nuclear extraction steps and precipitation prone conditions? Are the known biophysical properties of the nucleoplasm in vivo consistent with the existence of an extensive network of anastomosing filaments coursing dendritically throughout the interchromatin space? To what extent may the genome itself contribute information for its own quarternary structure in the interphase nucleus? These questions and recent work that bears on the mystique of the nuclear matrix are addressed in this essay. The degree to which gene expression literally depends on nonchromatin nuclear structure as a facilitating organizational format remains an intriguing but unsolved issue in eukaryotic cell biology, and considerable skepticism continues to surround the nuclear matrix fraction as an accurate representation of the in vivo situation. PMID- 10712501 TI - Phosphatidylinositol 4,5-bisphosphate regulates two steps of homotypic vacuole fusion. AB - Yeast vacuoles undergo cycles of fragmentation and fusion as part of their transmission to the daughter cell and in response to changes of nutrients and the environment. Vacuole fusion can be reconstituted in a cell free system. We now show that the vacuoles synthesize phosphoinositides during in vitro fusion. Of these phosphoinositides, phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate (PI(4,5)P(2)) are important for fusion. Monoclonal antibodies to PI(4,5)P(2), neomycin (a phosphoinositide ligand), and phosphatidylinositol-specific phospholipase C interfere with the reaction. Readdition of PI(4, 5)P(2) restores fusion in each case. Phosphatidylinositol 3 phosphate and PI(3,5)P(2) synthesis are not required. PI(4,5)P(2) is necessary for priming, i.e., for the Sec18p (NSF)-driven release of Sec17p (alpha-SNAP), which activates the vacuoles for subsequent tethering and docking. Therefore, it represents the kinetically earliest requirement identified for vacuole fusion so far. Furthermore, PI(4,5)P(2) is required at a step that can only occur after docking but before the BAPTA sensitive step in the latest stage of the reaction. We hence propose that PI(4,5)P(2) controls two steps of vacuole fusion. PMID- 10712502 TI - Clathrin-mediated endocytosis of MUC1 is modulated by its glycosylation state. AB - MUC1 is a mucin-like type 1 transmembrane protein associated with the apical surface of epithelial cells. In human tumors of epithelial origin MUC1 is overexpressed in an underglycosylated form with truncated O-glycans and accumulates in intracellular compartments. To understand the basis for this altered subcellular localization, we compared the synthesis and trafficking of various glycosylated forms of MUC1 in normal (Chinese hamster ovary) cells and glycosylation-defective (ldlD) cells that lack the epimerase to make UDP Gal/GalNAc from UDP-Glc/GlcNAc. Although the MUC1 synthesized in ldlD cells was rapidly degraded, addition of GalNAc alone to the culture media resulted in stabilization and near normal surface expression of MUC1 with truncated but sialylated O-glycans. Interestingly, the initial rate of endocytosis of this underglycosylated MUC1 was stimulated by twofold compared with fully glycosylated MUC1. However, the half-lives of the two forms were not different, indicating that trafficking to lysosomes was not affected. Both the normal and stimulated internalization of MUC1 could be blocked by hypertonic media, a hallmark of clathrin-mediated endocytosis. MUC1 endocytosis was also blocked by expression of a dominant-negative mutant of dynamin-1 (K44A), and MUC1 was observed in both clathrin-coated pits and vesicles by immunoelectron microscopy of ultrathin cryosections. Our data suggest that the subcellular redistribution of MUC1 in tumor cells could be a direct result of altered endocytic trafficking induced by its aberrant glycosylation; potential models are discussed. These results also implicate a new role for O-glycans on mucin-like membrane proteins entering the endocytic pathway through clathrin-coated pits. PMID- 10712503 TI - Glucose depletion rapidly inhibits translation initiation in yeast. AB - Glucose performs key functions as a signaling molecule in the yeast Saccharomyces cerevisiae. Glucose depletion is known to regulate gene expression via pathways that lead to derepression of genes at the transcriptional level. In this study, we have investigated the effect of glucose depletion on protein synthesis. We discovered that glucose withdrawal from the growth medium led to a rapid inhibition of protein synthesis and that this effect was readily reversed upon readdition of glucose. Neither the inhibition nor the reactivation of translation required new transcription. This inhibition also did not require activation of the amino acid starvation pathway or inactivation of the TOR kinase pathway. However, mutants in the glucose repression (reg1, glc7, hxk2, and ssn6), hexose transporter induction (snf3 rgt2), and cAMP-dependent protein kinase (tpk1(w) and tpk2(w)) pathways were resistant to the inhibitory effects of glucose withdrawal on translation. These findings highlight the intimate connection between the nutrient status of the cell and its translational capacity. They also help to define a new area of posttranscriptional regulation in yeast. PMID- 10712504 TI - Restoration of tight junction structure and barrier function by down-regulation of the mitogen-activated protein kinase pathway in ras-transformed Madin-Darby canine kidney cells. AB - In the Madin-Darby canine kidney epithelial cell line, the proteins occludin and ZO-1 are structural components of the tight junctions that seal the paracellular spaces between the cells and contribute to the epithelial barrier function. In Ras-transformed Madin-Darby canine kidney cells, occludin, claudin-1, and ZO-1 were absent from cell-cell contacts but were present in the cytoplasm, and the adherens junction protein E-cadherin was weakly expressed. After treatment of the Ras-transformed cells with the mitogen-activated protein kinase kinase (MEK1) inhibitor PD98059, which blocks the activation of mitogen-activated protein kinase (MAPK), occludin, claudin-1, and ZO-1 were recruited to the cell membrane, tight junctions were assembled, and E-cadherin protein expression was induced. Although it is generally believed that E-cadherin-mediated cell-cell adhesion is required for tight junction assembly, the recruitment of occludin to the cell cell contact area and the restoration of epithelial cell morphology preceded the appearance of E-cadherin at cell-cell contacts. Both electron microscopy and a fourfold increase in the transepithelial electrical resistance indicated the formation of functional tight junctions after MEK1 inhibition. Moreover, inhibition of MAPK activity stabilized occludin and ZO-1 by differentially increasing their half-lives. We also found that during the process of tight junction assembly after MEK1 inhibition, tyrosine phosphorylation of occludin and ZO-1, but not claudin-1, increased significantly. Our study demonstrates that down-regulation of the MAPK signaling pathway causes the restoration of epithelial cell morphology and the assembly of tight junctions in Ras-transformed epithelial cells and that tyrosine phosphorylation of occludin and ZO-1 may play a role in some aspects of tight junction formation. PMID- 10712505 TI - Dynein-dependent movements of the mitotic spindle in Saccharomyces cerevisiae Do not require filamentous actin. AB - In budding yeast, the mitotic spindle is positioned in the neck between the mother and the bud so that both cells inherit one nucleus. The movement of the mitotic spindle into the neck can be divided into two phases: (1) Kip3p-dependent movement of the nucleus to the neck and alignment of the short spindle, followed by (2) dynein-dependent movement of the spindle into the neck and oscillation of the elongating spindle within the neck. Actin has been hypothesized to be involved in all these movements. To test this hypothesis, we disrupted the actin cytoskeleton with the use of mutations and latrunculin A (latrunculin). We assayed nuclear segregation in synchronized cell populations and observed spindle movements in individual living cells. In synchronized cell populations, no actin cytoskeletal mutant segregated nuclei as poorly as cells lacking dynein function. Furthermore, nuclei segregated efficiently in latrunculin-treated cells. Individual living cell analysis revealed that the preanaphase spindle was mispositioned and misaligned in latrunculin-treated cells and that astral microtubules were misoriented, confirming a role for filamentous actin in the early, Kip3p-dependent phase of spindle positioning. Surprisingly, mispositioned and misaligned mitotic spindles moved into the neck in the absence of filamentous actin, albeit less efficiently. Finally, dynein-dependent sliding of astral microtubules along the cortex and oscillation of the elongating mitotic spindle in the neck occurred in the absence of filamentous actin. PMID- 10712506 TI - Mechanisms of sod2 gene amplification in Schizosaccharomyces pombe. AB - Gene amplification in eukaryotes plays an important role in drug resistance, tumorigenesis, and evolution. The Schizosaccharomyces pombe sod2 gene provides a useful model system to analyze this process. sod2 is near the telomere of chromosome I and encodes a plasma membrane Na(+)(Li(+))/H(+) antiporter. When sod2 is amplified, S. pombe survives otherwise lethal concentrations of LiCl, and >90% of the amplified sod2 genes are found in 180- and 225-kilobase (kb) linear amplicons. The sequence of the novel joint of the 180-kb amplicon indicates that it is formed by recombination between homologous regions near the telomeres of the long arm of chromosome I and the short arm of chromosome II. The 225-kb amplicon, isolated three times more frequently than the 180-kb amplicon, is a palindrome derived from a region near the telomere of chromosome I. The center of symmetry of this palindrome contains an inverted repeat consisting of two identical 134-base pair sequences separated by a 290-base pair spacer. LiCl resistant mutants arise 200-600 times more frequently in strains deficient for topoisomerases or DNA ligase activity than in wild-type strains, but the mutant cells contain the same amplicons. These data suggest that amplicon formation may begin with DNA lesions such as breaks. In the case of the 225-kb amplicon, the breaks may lead to a hairpin structure, which is then replicated to form a double stranded linear amplicon, or to a cruciform structure, which is then resolved to yield the same amplicon. PMID- 10712507 TI - Activation of Wee1 by p42 MAPK in vitro and in cycling xenopus egg extracts. AB - Xenopus oocytes and eggs provide a dramatic example of how the consequences of p42 mitogen-activated protein kinase (p42 MAPK) activation depend on the particular context in which the activation occurs. In oocytes, the activation of Mos, MEK, and p42 MAPK is required for progesterone-induced Cdc2 activation, and activated forms of any of these proteins can bring about Cdc2 activation in the absence of progesterone. However, in fertilized eggs, activation of the Mos/MEK/p42 MAPK pathway has the opposite effect, inhibiting Cdc2 activation and causing a G2 phase delay or arrest. In the present study, we have investigated the mechanism and physiological significance of the p42 MAPK-induced G2 phase arrest, using Xenopus egg extracts as a model system. We found that Wee1-depleted extracts were unable to arrest in G2 phase in response to Mos, and adding back Wee1 to the extracts restored their ability to arrest. This finding formally places Wee1 downstream of Mos/MEK/p42 MAPK. Purified recombinant p42 MAPK was found to phosphorylate recombinant Wee1 in vitro at sites that are phosphorylated in extracts. Phosphorylation by p42 MAPK resulted in a modest ( approximately 2 fold) increase in the kinase activity of Wee1 toward Cdc2. Titration experiments in extracts demonstrated that a twofold increase in Wee1 activity is sufficient to cause the delay in mitotic entry seen in Mos-treated extracts. Finally, we present evidence that the negative regulation of Cdc2 by Mos/MEK/p42 MAPK contributes to the presence of an unusually long G2 phase in the first mitotic cell cycle. Prematurely inactivating p42 MAPK in egg extracts resulted in a corresponding hastening of the first mitosis. The negative effect of p42 MAPK on Cdc2 activation may help ensure that the first mitotic cell cycle is long enough to allow karyogamy to be accomplished successfully. PMID- 10712508 TI - Dynamics of the endoplasmic reticulum and golgi apparatus during early sea urchin development. AB - The endoplasmic reticulum (ER) and Golgi were labeled by green fluorescent protein chimeras and observed by time-lapse confocal microscopy during the rapid cell cycles of sea urchin embryos. The ER undergoes a cyclical microtubule dependent accumulation at the mitotic poles and by photobleaching experiments remains continuous through the cell cycle. Finger-like indentations of the nuclear envelope near the mitotic poles appear 2-3 min before the permeability barrier of the nuclear envelope begins to change. This permeability change in turn is approximately 30 s before nuclear envelope breakdown. During interphase, there are many scattered, disconnected Golgi stacks throughout the cytoplasm, which appear as 1- to 2-microm fluorescent spots. The number of Golgi spots begins to decline soon after nuclear envelope breakdown, reaches a minimum soon after cytokinesis, and then rapidly increases. At higher magnification, smaller spots are seen, along with increased fluorescence in the ER. Quantitative measurements, along with nocodazole and photobleaching experiments, are consistent with a redistribution of some of the Golgi to the ER during mitosis. The scattered Golgi coalesce into a single large aggregate during the interphase after the ninth embryonic cleavage; this is likely to be preparatory for secretion of the hatching enzyme during the following cleavage cycle. PMID- 10712509 TI - Degradation of the transcription factor Gcn4 requires the kinase Pho85 and the SCF(CDC4) ubiquitin-ligase complex. AB - Gcn4, a yeast transcriptional activator that promotes the expression of amino acid and purine biosynthesis genes, is rapidly degraded in rich medium. Here we report that SCF(CDC4), a recently characterized protein complex that acts in conjunction with the ubiquitin-conjugating enzyme Cdc34 to degrade cell cycle regulators, is also necessary for the degradation of the transcription factor Gcn4. Degradation of Gcn4 occurs throughout the cell cycle, whereas degradation of the known cell cycle substrates of Cdc34/SCF(CDC4) is cell cycle regulated. Gcn4 ubiquitination and degradation are regulated by starvation for amino acids, whereas the degradation of the cell cycle substrates of Cdc34/SCF(CDC4) is unaffected by starvation. We further show that unlike the cell cycle substrates of Cdc34/SCF(CDC4), which require phosphorylation by the kinase Cdc28, Gcn4 degradation requires the kinase Pho85. We identify the critical target site of Pho85 on Gcn4; a mutation of this site stabilizes the protein. A specific Pho85 Pcl complex that is able to phosphorylate Gcn4 on that site is inactive under conditions under which Gcn4 is stable. Thus, Cdc34/SCF(CDC4) activity is constitutive, and regulation of the stability of its various substrates occurs at the level of their phosphorylation. PMID- 10712510 TI - Caspase-mediated cleavage of p130cas in etoposide-induced apoptotic Rat-1 cells. AB - Apoptosis causes characteristic morphological changes in cells, including membrane blebbing, cell detachment from the extracellular matrix, and loss of cell-cell contacts. We investigated the changes in focal adhesion proteins during etoposide-induced apoptosis in Rat-1 cells and found that during apoptosis, p130cas (Crk-associated substrate [Cas]) is cleaved by caspase-3. Sequence analysis showed that Cas contains 10 DXXD consensus sites preferred by caspase-3. We identified two of these sites (DVPD(416)G and DSPD(748)G) in vitro, and point mutations substituting the Asp of DVPD(416)G and DSPD(748)G with Glu blocked caspase-3-mediated cleavage. Cleavage at DVPD(416)G generated a 74-kDa fragment, which was in turn cleaved at DSPD(748)G, yielding 47- and 31-kDa fragments. Immunofluorescence microscopy revealed well-developed focal adhesion sites in control cells that dramatically declined in number in etoposide-treated cells. Cas cleavage correlated temporally with the onset of apoptosis and coincided with the loss of p125FAK (focal adhesion kinase [FAK]) from focal adhesion sites and the attenuation of Cas-paxillin interactions. Considering that Cas associates with FAK, paxillin, and other molecules involved in the integrin signaling pathway, these results suggest that caspase-mediated cleavage of Cas contributes to the disassembly of focal adhesion complexes and interrupts survival signals from the extracellular matrix. PMID- 10712511 TI - Brefeldin A-dependent membrane tubule formation reconstituted in vitro is driven by a cell cycle-regulated microtubule motor. AB - Treatment of cultured cells with brefeldin A (BFA) induces the formation of extensive membrane tubules from the Golgi apparatus, trans-Golgi network, and early endosomes in a microtubule-dependent manner. We have reconstituted this transport process in vitro using Xenopus egg cytosol and a rat liver Golgi enriched membrane fraction. The presence of BFA results in the formation of an intricate, interconnected tubular membrane network, a process that, as in vivo, is inhibited by nocodazole, the H1 anti-kinesin monoclonal antibody, and by membrane pretreatment with guanosine 5'-O-(3-thiotriphosphate). Surprisingly, membrane tubule formation is not due to the action of conventional kinesin or any of the other motors implicated in Golgi membrane dynamics. Two candidate motors of approximately 100 and approximately 130 kDa have been identified using the H1 antibody, both of which exhibit motor properties in a biochemical assay. Finally, BFA-induced membrane tubule formation does not occur in metaphase cytosol, and because membrane binding of both candidate motors is not altered after incubation in metaphase compared with interphase cytosol, these results suggest that either the ATPase or microtubule-binding activity of the relevant motor is cell cycle regulated. PMID- 10712512 TI - Dual lipid modification motifs in G(alpha) and G(gamma) subunits are required for full activity of the pheromone response pathway in Saccharomyces cerevisiae. AB - To establish the biological function of thioacylation (palmitoylation), we have studied the heterotrimeric guanine nucleotide-binding protein (G protein) subunits of the pheromone response pathway of Saccharomyces cerevisiae. The yeast G protein gamma subunit (Ste18p) is unusual among G(gamma) subunits because it is farnesylated at cysteine 107 and has the potential to be thioacylated at cysteine 106. Substitution of either cysteine results in a strong signaling defect. In this study, we found that Ste18p is thioacylated at cysteine 106, which depended on prenylation of cysteine 107. Ste18p was targeted to the plasma membrane even in the absence of prenylation or thioacylation. However, G protein activation released prenylation- or thioacylation-defective Ste18p into the cytoplasm. Hence, lipid modifications of the G(gamma) subunit are dispensable for G protein activation by receptor, but they are required to maintain the plasma membrane association of G(betagamma) after receptor-stimulated release from G(alpha). The G protein alpha subunit (Gpa1p) is tandemly modified at its N terminus with amide and thioester-linked fatty acids. Here we show that Gpa1p was thioacylated in vivo with a mixture of radioactive myristate and palmitate. Mutation of the thioacylation site in Gpa1p resulted in yeast cells that displayed partial activation of the pathway in the absence of pheromone. Thus, dual lipidation motifs on Gpa1p and Ste18p are required for a fully functional pheromone response pathway. PMID- 10712513 TI - Apg5p functions in the sequestration step in the cytoplasm-to-vacuole targeting and macroautophagy pathways. AB - The cytoplasm-to-vacuole targeting (Cvt) pathway and macroautophagy are dynamic events involving the rearrangement of membrane to form a sequestering vesicle in the cytosol, which subsequently delivers its cargo to the vacuole. This process requires the concerted action of various proteins, including Apg5p. Recently, it was shown that another protein required for the import of aminopeptidase I (API) and autophagy, Apg12p, is covalently attached to Apg5p through the action of an E1-like enzyme, Apg7p. We have undertaken an analysis of Apg5p function to gain a better understanding of the role of this novel nonubiquitin conjugation reaction in these import pathways. We have generated the first temperature-sensitive mutant in the Cvt pathway, designated apg5(ts). Biochemical analysis of API import in the apg5(ts) strain confirmed that Apg5p is directly required for the import of API via the Cvt pathway. By analyzing the stage of API import that is blocked in the apg5(ts) mutant, we have determined that Apg5p is involved in the sequestration step and is required for vesicle formation and/or completion. PMID- 10712514 TI - Sec24p and Iss1p function interchangeably in transport vesicle formation from the endoplasmic reticulum in Saccharomyces cerevisiae. AB - The Sec23p/Sec24p complex functions as a component of the COPII coat in vesicle transport from the endoplasmic reticulum. Here we characterize Saccharomyces cerevisiae SEC24, which encodes a protein of 926 amino acids (YIL109C), and a close homologue, ISS1 (YNL049C), which is 55% identical to SEC24. SEC24 is essential for vesicular transport in vivo because depletion of Sec24p is lethal, causing exaggeration of the endoplasmic reticulum and a block in the maturation of carboxypeptidase Y. Overproduction of Sec24p suppressed the temperature sensitivity of sec23-2, and overproduction of both Sec24p and Sec23p suppressed the temperature sensitivity of sec16-2. SEC24 gene disruption could be complemented by overexpression of ISS1, indicating functional redundancy between the two homologous proteins. Deletion of ISS1 had no significant effect on growth or secretion; however, iss1Delta mutants were found to be synthetically lethal with mutations in the v-SNARE genes SEC22 and BET1. Moreover, overexpression of ISS1 could suppress mutations in SEC22. These genetic interactions suggest that Iss1p may be specialized for the packaging or the function of COPII v-SNAREs. Iss1p tagged with His(6) at its C terminus copurified with Sec23p. Pure Sec23p/Iss1p could replace Sec23p/Sec24p in the packaging of a soluble cargo molecule (alpha-factor) and v-SNAREs (Sec22p and Bet1p) into COPII vesicles. Abundant proteins in the purified vesicles produced with Sec23p/Iss1p were indistinguishable from those in the regular COPII vesicles produced with Sec23p/Sec24p. PMID- 10712515 TI - Identification of two RNA-binding proteins associated with human telomerase RNA. AB - Telomerase plays a crucial role in telomere maintenance in vivo. To understand telomerase regulation, we have been characterizing components of the enzyme. To date several components of the mammalian telomerase holoenzyme have been identified: the essential RNA component (human telomerase RNA [hTR]), the catalytic subunit human telomerase reverse transcriptase (hTERT), and telomerase associated protein 1. Here we describe the identification of two new proteins that interact with hTR: hStau and L22. Antisera against both proteins immunoprecipitated hTR, hTERT, and telomerase activity from cell extracts, suggesting that the proteins are associated with telomerase. Both proteins localized to the nucleolus and cytoplasm. Although these proteins are associated with telomerase, we found no evidence of their association with each other or with telomerase-associated protein 1. Both hStau and L22 are more abundant than TERT. This, together with their localization, suggests that they may be associated with other ribonucleoprotein complexes in cells. We propose that these two hTR-associated proteins may play a role in hTR processing, telomerase assembly, or localization in vivo. PMID- 10712516 TI - Discontinuous actin hexagon, a protein essential for cortical furrow formation in Drosophila, is membrane associated and hyperphosphorylated. AB - discontinuous actin hexagon (dah) is a maternal-effect gene essential for the formation of cortical furrows during Drosophila embryogenesis, and DAH protein colocalizes with actin in these furrows. Biochemical fractionation experiments presented here demonstrate that DAH is highly enriched in the membrane fraction and that its membrane association is resistant to high-salt and alkaline washes. Furthermore, it partitions into the detergent phase of the Triton X-114 solution, indicating its tight binding to the membranes. DAH can also interact with the actin cytoskeleton, because a fraction of DAH remains insoluble to nonionic detergent along with actin. These biochemical characterizations suggest that DAH may play a role in the linkage of the actin cytoskeleton to membranes. Using phosphatase inhibitors, we detected multiple phosphorylated forms of DAH in embryonic extracts. The DAH phosphorylation peaks during cellularization, a stage at which DAH function is critical. A kinase activity is coimmunoprecipitated with the DAH complex and hyperphosphorylates DAH in vitro. Purified casein kinase I can also hyperphosphorylate DAH in the immune complex. Both DAH localization and phosphorylation are disrupted in another maternal-effect mutant, nuclear-fallout. It is possible that nuclear-fallout collaborates with dah and directs DAH protein localization to the cortical furrows. PMID- 10712517 TI - Bone morphogenetic protein receptor complexes on the surface of live cells: a new oligomerization mode for serine/threonine kinase receptors. AB - The bone morphogenetic proteins (BMPs) play important roles in embryogenesis and normal cell growth. The BMP receptors belong to the family of serine/threonine kinase receptors, whose activation has been investigated intensively for the transforming growth factor-beta (TGF-beta) receptor subfamily. However, the interactions between the BMP receptors, the composition of the active receptor complex, and the role of the ligand in its formation have not yet been investigated and were usually assumed to follow the same pattern as the TGF-beta receptors. Here we demonstrate that the oligomerization pattern of the BMP receptors is different and is more flexible and susceptible to modulation by ligand. Using several complementary approaches, we investigated the formation of homomeric and heteromeric complexes between the two known BMP type I receptors (BR-Ia and BR-Ib) and the BMP type II receptor (BR-II). Coimmunoprecipitation studies detected the formation of heteromeric and homomeric complexes among all the BMP receptor types even in the absence of ligand. These complexes were also detected at the cell surface after BMP-2 binding and cross-linking. Using antibody-mediated immunofluorescence copatching of epitope-tagged receptors, we provide evidence in live cells for preexisting heteromeric (BR-II/BR-Ia and BR II/BR-Ib) and homomeric (BR-II/BR-II, BR-Ia/ BR-Ia, BR-Ib/ BR-Ib, and also BR-Ia/ BR-Ib) oligomers in the absence of ligand. BMP-2 binding significantly increased hetero- and homo-oligomerization (except for the BR-II homo-oligomer, which binds ligand poorly in the absence of BR-I). In contrast to previous observations on TGF-beta receptors, which were found to be fully homodimeric in the absence of ligand, the BMP receptors show a much more flexible oligomerization pattern. This novel feature in the oligomerization mode of the BMP receptors allows higher variety and flexibility in their responses to various ligands as compared with the TGF-beta receptors. PMID- 10712518 TI - Periodic expression of the cyclin-dependent kinase inhibitor p57(Kip2) in trophoblast giant cells defines a G2-like gap phase of the endocycle. AB - Endoreduplication is an unusual form of cell cycle in which rounds of DNA synthesis repeat in the absence of intervening mitoses. How G1/S cyclin-dependent kinase (Cdk) activity is regulated during the mammalian endocycle is poorly understood. We show here that expression of the G1/S Cdk inhibitor p57(Kip2) is induced coincidentally with the transition to the endocycle in trophoblast giant cells. Kip2 mRNA is constitutively expressed during subsequent endocycles, but the protein level fluctuates. In trophoblast giant cells synchronized for the first few endocycles, the p57(Kip2) protein accumulates only at the end of S phase and then rapidly disappears a few hours before the onset of the next S phase. The protein becomes stabilized by mutation of a C-terminal Cdk phosphorylation site. As a consequence, introduction of this stable form of p57(Kip2) into giant cells blocks S-phase entry. These data imply that p57(Kip2) is subject to phosphorylation-dependent turnover. Surprisingly, although this occurs in endoreduplicating giant cells, p57(Kip2) is stable when ectopically expressed in proliferating trophoblast cells, indicating that these cells lack the mechanism for protein targeting and/or degradation. These data show that the appearance of p57(Kip2) punctuates the completion of DNA replication, whereas its turnover is subsequently required to initiate the next round of endoreduplication in trophoblast giant cells. Cyclical expression of a Cdk inhibitor, by terminating G1/S Cdk activity, may help promote the resetting of DNA replication machinery. PMID- 10712519 TI - Physical state of the extracellular matrix regulates the structure and molecular composition of cell-matrix adhesions. AB - This study establishes that the physical state of the extracellular matrix can regulate integrin-mediated cytoskeletal assembly and tyrosine phosphorylation to generate two distinct types of cell-matrix adhesions. In primary fibroblasts, alpha(5)beta(1) integrin associates mainly with fibronectin fibrils and forms adhesions structurally distinct from focal contacts, independent of actomyosin mediated cell contractility. These "fibrillar adhesions" are enriched in tensin, but contain low levels of the typical focal contact components paxillin, vinculin, and tyrosine-phosphorylated proteins. However, when the fibronectin is covalently linked to the substrate, alpha(5)beta(1) integrin forms highly tyrosine-phosphorylated, "classical" focal contacts containing high levels of paxillin and vinculin. These experiments indicate that the physical state of the matrix, not just its molecular composition, is a critical factor in defining cytoskeletal organization and phosphorylation at adhesion sites. We propose that molecular organization of adhesion sites is controlled by at least two mechanisms: 1) specific integrins associate with their ligands in transmembrane complexes with appropriate cytoplasmic anchor proteins (e.g., fibronectin alpha(5)beta(1) integrin-tensin complexes), and 2) physical properties (e.g., rigidity) of the extracellular matrix regulate local tension at adhesion sites and activate local tyrosine phosphorylation, recruiting a variety of plaque molecules to these sites. These mechanisms generate structurally and functionally distinct types of matrix adhesions in fibroblasts. PMID- 10712520 TI - Transforming growth factor beta(1) selectively inhibits the cyclic AMP-dependent proliferation of primary thyroid epithelial cells by preventing the association of cyclin D3-cdk4 with nuclear p27(kip1). AB - Dog thyroid epithelial cells in primary culture constitute a physiologically relevant model of positive control of DNA synthesis initiation and G0-S prereplicative phase progression by cAMP as a second messenger for thyrotropin (thyroid-stimulating hormone [TSH]). As previously shown in this system, the cAMP dependent mitogenic pathway differs from growth factor cascades as it stimulates the accumulation of p27(kip1) but not cyclins D. Nevertheless, TSH induces the nuclear translocations and assembly of cyclin D3 and cdk4, which are essential in cAMP-dependent mitogenesis. Here we demonstrate that transforming growth factor beta(1) (TGFbeta(1)) selectively inhibits the cAMP-dependent cell cycle in mid-G1 and various cell cycle regulatory events, but it weakly affects the stimulation of DNA synthesis by epidermal growth factor (EGF), hepatocyte growth factor, serum, and phorbol esters. EGF+serum and TSH did not interfere importantly with TGFbeta receptor signaling, because they did not affect the TGFbeta-induced nuclear translocation of Smad 2 and 3. TGFbeta inhibited the phosphorylation of Rb, p107, and p130 induced by TSH, but it weakly affected the phosphorylation state of Rb-related proteins in EGF+serum-treated cells. TGFbeta did not inhibit c-myc expression. In TSH-stimulated cells, TGFbeta did not affect the expression of cyclin D3, cdk4, and p27(kip1), nor the induced formation of cyclin D3-cdk4 complexes, but it prevented the TSH-induced relocalization of p27(kip1) from cdk2 to cyclin D3-cdk4. It prevented the nuclear translocations of cdk4 and cyclin D3 without altering the assembly of cyclin D3-cdk4 complexes probably formed in the cytoplasm, where they were prevented from sequestering nuclear p27(kip1) away from cdk2. This study dissociates the assembly of cyclin D3-cdk4 complexes from their nuclear localization and association with p27(kip1). It provides a new mechanism of regulation of proliferation by TGFbeta, which points out the subcellular location of cyclin D-cdk4 complexes as a crucial factor integrating mitogenic and antimitogenic regulations in an epithelial cell in primary culture. PMID- 10712521 TI - The alpha isoform of protein kinase C is involved in signaling the response of desmosomes to wounding in cultured epithelial cells. AB - Initiation of reepithelialization upon wounding is still poorly understood. To enhance this understanding, we focus here on changes in the adhesive state of desmosomes of cultured Madin-Darby canine kidney cells in response to wounding of confluent cell sheets. Previous results show that desmosomal adhesion in Madin Darby canine kidney cells changes from a calcium-dependent state to calcium independence in confluent cell sheets. We show that this change, which requires culture confluence to develop, is rapidly reversed upon wounding of confluent cell sheets. Moreover, the change to calcium dependence in wound edge cells is propagated to cells hundreds of micrometers away from the wound edge. Rapid transition from calcium independence to calcium dependence also occurs when cells are treated with phorbol esters that activate PKC. PKC inhibitors, including the conventional isoform inhibitor Go6976, cause rapid transition from calcium dependence to calcium independence, even in subconfluent cells. The cellular location of the alpha isoform of PKC correlates with the calcium dependence of desmosomes. Upon monolayer wounding, PKCalpha translocates rapidly to the cell periphery, becomes Triton X-100 insoluble, and also becomes concentrated in lamellipodia. The PKCalpha translocation upon wounding precedes both the increase in PKC activity in the membrane fraction and the reversion of desmosomes to calcium dependence. Specific depletion of PKCalpha with an antisense oligonucleotide increases the number of cells with calcium-independent desmosomes. These results show that PKCalpha participates in a novel signaling pathway that modulates desmosomal adhesion in response to wounding. PMID- 10712522 TI - Polarized sphingolipid transport from the subapical compartment changes during cell polarity development. AB - The subapical compartment (SAC) plays an important role in the polarized transport of proteins and lipids. In hepatoma-derived HepG2 cells, fluorescent analogues of sphingomyelin and glucosylceramide are sorted in the SAC. Here, evidence is provided that shows that polarity development is regulated by a transient activation of endogenous protein kinase A and involves a transient activation of a specific membrane transport pathway, marked by the trafficking of the labeled sphingomyelin, from the SAC to the apical membrane. This protein kinase A-regulated pathway differs from the apical recycling pathway, which also traverses SAC. After reaching optimal polarity, the direction of the apically activated pathway switches to one in the basolateral direction, without affecting the apical recycling pathway. PMID- 10712523 TI - The p42/p44 MAP kinase pathway prevents apoptosis induced by anchorage and serum removal. AB - Anchorage removal like growth factor removal induces apoptosis. In the present study we have characterized signaling pathways that can prevent this cell death using a highly growth factor- and anchorage-dependent line of lung fibroblasts (CCL39). After anchorage removal from exponentially growing cells, annexin V-FITC labeling can be detected after 8 h. Apoptosis was confirmed by analysis of sub-G1 DNA content and Western blotting of the caspase substrate poly (ADP-ribose) polymerase. Growth factor withdrawal accelerates and potentiates suspension induced cell death. Activation of Raf-1 kinase in suspension cultures of CCL39 or Madin-Darby canine kidney cells stably expressing an estrogen-inducible activated Raf-1 construct (DeltaRaf-1:ER) suppresses apoptosis induced by growth factor and/or anchorage removal. This protective effect appears to be mediated by the Raf, mitogen- or extracellular signal-regulated kinase kinase (MEK), and mitogen activated protein kinase module because it is sensitive to pharmacological inhibition of MEK-1 and it can be mimicked by expression of constitutively active MEK-1 in CCL39 cells. Finally, apoptosis induced by disruption of the actin cytoskeleton with the Rho-directed toxin B (Clostridium difficile) is prevented by activation of the DeltaRaf-1:ER chimeric construct. These findings highlight the ability of p42/p44 mitogen-activated protein kinase to generate survival signals that counteract cell death induced by loss of matrix contact, cytoskeletal integrity, and extracellular mitogenic factors. PMID- 10712524 TI - Shade avoidance responses. Driving auxin along lateral routes. PMID- 10712525 TI - Biochemical and molecular genetic aspects of floral scents. PMID- 10712526 TI - Purification of a jojoba embryo fatty acyl-coenzyme A reductase and expression of its cDNA in high erucic acid rapeseed. AB - The jojoba (Simmondsia chinensis) plant produces esters of long-chain alcohols and fatty acids (waxes) as a seed lipid energy reserve. This is in contrast to the triglycerides found in seeds of other plants. We purified an alcohol-forming fatty acyl-coenzyme A reductase (FAR) from developing embryos and cloned the cDNA encoding the enzyme. Expression of a cDNA in Escherichia coli confers FAR activity upon those cells and results in the accumulation of fatty alcohols. The FAR sequence shows significant homology to an Arabidopsis protein of unknown function that is essential for pollen development. When the jojoba FAR cDNA is expressed in embryos of Brassica napus, long-chain alcohols can be detected in transmethylated seed oils. Resynthesis of the gene to reduce its A plus T content resulted in increased levels of alcohol production. In addition to free alcohols, novel wax esters were detected in the transgenic seed oils. In vitro assays revealed that B. napus embryos have an endogenous fatty acyl-coenzyme A: fatty alcohol acyl-transferase activity that could account for this wax synthesis. Thus, introduction of a single cDNA into B. napus results in a redirection of a portion of seed oil synthesis from triglycerides to waxes. PMID- 10712527 TI - Purification of a jojoba embryo wax synthase, cloning of its cDNA, and production of high levels of wax in seeds of transgenic arabidopsis. AB - Wax synthase (WS, fatty acyl-coenzyme A [coA]: fatty alcohol acyltransferase) catalyzes the final step in the synthesis of linear esters (waxes) that accumulate in seeds of jojoba (Simmondsia chinensis). We have characterized and partially purified this enzyme from developing jojoba embryos. A protein whose presence correlated with WS activity during chromatographic fractionation was identified and a cDNA encoding that protein was cloned. Seed-specific expression of the cDNA in transgenic Arabidopsis conferred high levels of WS activity on developing embryos from those plants. The WS sequence has significant homology with several Arabidopsis open reading frames of unknown function. Wax production in jojoba requires, in addition to WS, a fatty acyl-CoA reductase (FAR) and an efficient fatty acid elongase system that forms the substrates preferred by the FAR. We have expressed the jojoba WS cDNA in Arabidopsis in combination with cDNAs encoding the jojoba FAR and a beta-ketoacyl-CoA synthase (a component of fatty acid elongase) from Lunaria annua. (13)C-Nuclear magnetic resonance analysis of pooled whole seeds from transgenic plants indicated that as many as 49% of the oil molecules in the seeds were waxes. Gas chromatography analysis of transmethylated oil from individual seeds suggested that wax levels may represent up to 70% (by weight) of the oil present in those seeds. PMID- 10712528 TI - Expression of aluminum-induced genes in transgenic arabidopsis plants can ameliorate aluminum stress and/or oxidative stress. AB - To examine the biological role of Al-stress-induced genes, nine genes derived from Arabidopsis, tobacco (Nicotiana tabacum L.), wheat (Triticum aestivum L.), and yeast (Saccharomyces cerevisiae) were expressed in Arabidopsis ecotype Landsberg. Lines containing eight of these genes were phenotypically normal and were tested in root elongation assays for their sensitivity to Al, Cd, Cu, Na, Zn, and to oxidative stresses. An Arabidopsis blue-copper-binding protein gene (AtBCB), a tobacco glutathione S-transferase gene (parB), a tobacco peroxidase gene (NtPox), and a tobacco GDP-dissociation inhibitor gene (NtGDI1) conferred a degree of resistance to Al. Two of these genes, AtBCB and parB, and a peroxidase gene from Arabidopsis (AtPox) also showed increased resistance to oxidative stress induced by diamide, while parB conferred resistance to Cu and Na. Al content of Al-treated root tips was reduced in the four Al-resistant plant lines compared with wild-type Ler-0, as judged by morin staining. All four Al-resistant lines also showed reduced staining of roots with 2',7'-dichloro fluorescein diacetate (H(2)DCFDA), an indicator of oxidative stress. We conclude that Al induced genes can serve to protect against Al toxicity, and also provide genetic evidence for a link between Al stress and oxidative stress in plants. PMID- 10712529 TI - Post-transcriptional regulation prevents accumulation of glutathione reductase protein and activity in the bundle sheath cells of maize. AB - Glutathione reductase (GR; EC 1.6.4.2) activity was assayed in bundle sheath and mesophyll cells of maize (Zea mays L. var H99) from plants grown at 20 degrees C, 18 degrees C, and 15 degrees C. The purity of each fraction was determined by measuring the associated activity of the compartment-specific marker enzymes, Rubisco and phosphoenolpyruvate carboxylase, respectively. GR activity and the abundance of GR protein and mRNA increased in plants grown at 15 degrees C and 18 degrees C compared with those grown at 20 degrees C. In all cases GR activity was found only in mesophyll fractions of the leaves, with no GR activity being detectable in bundle sheath extracts. Immunogold labeling with GR-specific antibodies showed that the GR protein was exclusively localized in the mesophyll cells of leaves at all growth temperatures, whereas GR transcripts (as determined by in situ hybridization techniques) were observed in both cell types. These results indicate that post-transcriptional regulation prevents GR accumulation in the bundle sheath cells of maize leaves. The resulting limitation on the capacity for regeneration of reduced glutathione in this compartment may contribute to the extreme chilling sensitivity of maize leaves. PMID- 10712530 TI - StGCPRP, a potato gene strongly expressed in stomatal guard cells, defines a novel type of repetitive proline-rich proteins. AB - Guard cells represent a highly differentiated cell type within the epidermis of plant leaves and stems. They respond to many endogenous and environmental signals and thereby modify the size of the stomatal pore they surround. We identified a novel gene that is highly expressed in guard cells of potato (Solanum tuberosum). It encodes a repetitive proline (Pro)-rich protein of 54 kD (491 amino acids) and was named StGCPRP (S. tuberosum guard cell Pro-rich protein). StGCPRP has a bipartite structure. The C-terminal part of StGCPRP contains a high percentage (46%) of Pro residues organized in distinct repetitive sequence motifs, whereas its extended N terminus is essentially free of Pros. StGCPRP represents the first member of a novel class of hybrid Pro-rich proteins that we designated NHyPRPs. In young but not in mature leaves, StGCPRP transcripts were also present at high levels in mesophyll cells (in addition to guard cells), indicating developmental regulation of StGCPRP gene expression. In addition, StGCPRP expression is regulated by environmental factors, as shown by a decrease in StGCPRP transcript levels under drought stress. Two proteins similar to StGCPRP were found to be encoded by the Arabidopsis genome, indicating that NHyPRPs are more widely distributed in higher plants. PMID- 10712531 TI - Aluminum tolerance genes on the short arm of chromosome 3R are linked to organic acid release in triticale. AB - Triticale, a hybrid between wheat and rye, shows a high degree of Al tolerance that is inherited from rye, but the mechanisms of high Al tolerance in both rye and triticale are unknown. We found that the short arm of chromosome 3R carries genes necessary for Al tolerance in triticale (x Triticosecale Wittmark cv Currency). Detailed comparative studies with a 3DS.3RL translocation line (ST22) and a non-substitution line (ST2) were conducted. Root elongation was similarly inhibited by Al in ST2 and ST22 during the first 12 h of Al treatment, but more strongly in ST22 than in ST2 at 18 h and thereafter. The root inhibition induced by other metals (Cu, Cd, and La) was similar between ST2 and ST22, suggesting that the action of the genes for Al tolerance on the short arm of triticale chromosome 3R is highly specific to Al. A 2-fold larger amount of malate and citrate was released from the roots of ST2 than from ST22 at 12 and 18 h after Al treatment, respectively. The marked lag phase in the inhibition of root elongation and the release of organic acids implies that the expression of genes on the short arm of triticale chromosome 3R is induced by Al, and that these genes are necessary for the release of organic acids. PMID- 10712532 TI - A novel gibberellin-induced gene from rice and its potential regulatory role in stem growth. AB - Os-GRF1 (Oryza sativa-GROWTH-REGULATING FACTOR1) was identified in a search for genes that are differentially expressed in the intercalary meristem of deepwater rice (Oryza sativa L.) internodes in response to gibberellin (GA). Os-GRF1 displays general features of transcription factors, contains a functional nuclear localization signal, and has three regions with similarities to sequences in the database. One of these regions is similar to a protein interaction domain of SWI2/SNF2, which is a subunit of a chromatin-remodeling complex in yeast. The two other domains are novel and found only in plant proteins of unknown function. To study its role in plant growth, Os-GRF1 was expressed in Arabidopsis. Stem elongation of transformed plants was severely inhibited, and normal growth could not be recovered by the application of GA. Our results indicate that Os-GRF1 belongs to a novel class of plant proteins and may play a regulatory role in GA induced stem elongation. PMID- 10712533 TI - Expression of AtPRP3, a proline-rich structural cell wall protein from Arabidopsis, is regulated by cell-type-specific developmental pathways involved in root hair formation. AB - The tightly regulated expression patterns of structural cell wall proteins in several plant species indicate that they play a crucial role in determining the extracellular matrix structure for specific cell types. We demonstrate that AtPRP3, a proline-rich cell wall protein in Arabidopsis, is expressed in root hair-bearing epidermal cells at the root/shoot junction and within the root differentiation zone of light-grown seedlings. Several lines of evidence support a direct relationship between AtPRP3 expression and root hair development. AtPRP3/beta-glucuronidase (GUS) expression increased in roots of transgenic seedlings treated with either 1-aminocyclopropane-1-carboxylic acid (ACC) or alpha-naphthaleneacetic acid (alpha-NAA), compounds known to promote root hair formation. In the presence of 1-alpha-(2-aminoethoxyvinyl)glycine (AVG), an inhibitor of ethylene biosynthesis, AtPRP3/GUS expression was strongly reduced, but could be rescued by co-addition of ACC or alpha-NAA to the growth medium. In addition, AtPRP3/GUS activity was enhanced in ttg and gl2 mutant backgrounds that exhibit ectopic root hairs, but was reduced in rhd6 and 35S-R root-hair-less mutant seedlings. These results indicate that AtPRP3 is regulated by developmental pathways involved in root hair formation, and are consistent with AtPRP3's contributing to cell wall structure in Arabidopsis root hairs. PMID- 10712534 TI - Characterization of the Brassica napus extraplastidial linoleate desaturase by expression in Saccharomyces cerevisiae. AB - The substrate specificity and regioselectivity of the Brassica napus extraplastidial linoleate desaturase (FAD3) was investigated in vivo in a heterologous expression system. A strain of the yeast Saccharomyces cerevisiae producing the plant enzyme was constructed and cultured in media containing a variety of fatty acids. The products of desaturation of these potential substrates were determined by gas chromatographic and mass spectrometric analysis of the yeast cultures. The results indicate that the enzyme has: (a) omega-3, as opposed to Delta-15 or double-bond-related regioselectivity, (b) the ability to desaturate substrates in the 16 to 22 carbon range, (c) a preference for substrates with omega-6 double bonds, but the ability to desaturate substrates with omega-6 hydroxyl groups or omega-9 or omega-5 double bonds, and (d) a relative insensitivity to double bonds proximal to the carboxyl end of the substrate. PMID- 10712535 TI - Hormone autotrophic growth and differentiation identifies mutant lines of Arabidopsis with altered cytokinin and auxin content or signaling. AB - We describe mutant tissue lines of Arabidopsis that are able to grow in vitro as callus on hormone-free medium. The 14 lines presented here show different hormone autotrophic differentiation behaviors that can be classified into three categories: (a) forming roots (rooty callus), (b) forming shoots or shoot-like structures (shooty callus), or (c) growing without organ formation (callus). Three fast-growing lines showed altered steady-state mRNA levels of the Cdc2 and CycD3 cell cycle genes. Three of the six rooty callus lines contained about 20- to 30-fold higher levels of auxins than wild-type callus. These and two other lines with normal auxin content showed an increased steady-state level of IAA1 and IAA2 transcripts in the absence of exogenous auxin. Five of the six shooty callus lines had increased steady-state mRNA levels of the CKI1 gene and/or of the homeobox genes KNAT1 and STM, suggesting that the phenotype is linked to altered cytokinin signaling. Also, one cytokinin-overproducing line with only 5% of wild-type cytokinin oxidase activity was identified. These results indicate that screening for hormone-autonomous growth identifies mutants with altered hormone content or signaling. PMID- 10712536 TI - Differential expression of photosynthesis and nitrogen fixation genes in the cyanobacterium Plectonema boryanum. AB - The filamentous non-heterocystous cyanobacterium Plectonema boryanum fixes dinitrogen at a high rate during microaerobic growth in continuous illumination by temporal separation of oxygen-evolving photosynthesis and oxygen-sensitive dinitrogen fixation. The onset of nitrogen fixation is preceded by a depression in photosynthesis that establishes a sufficiently low level of dissolved oxygen in the growth medium. A several-fold reduction in the level of transcripts coding for phycocyanin (cpcBA) and the chlorophyll a binding protein of photosystem II (psbC) and psbA accompanied the depression in photosynthetic oxygen evolution. Unlike most of the other organisms examined to date, in P. boryanum, psbC and psbD do not appear to be co-transcribed. The psbC transcripts were down-regulated several fold, while the psbD transcript declined marginally during the nitrogen fixation phase. A decrease in dissolved oxygen and a dramatic increase in the level of nifH transcripts and the enzyme activity of nitrogenase were characteristic of the nitrogen fixation phase. The level of transcript for glnA, which encodes glutamine synthetase, was not altered. Reciprocal regulation of gene expression was well orchestrated with the alternating cycles of photosynthesis and nitrogen fixation in P. boryanum. PMID- 10712537 TI - Regulation of sulfate assimilation by nitrogen in Arabidopsis. AB - Using Arabidopsis, we analyzed the effect of omission of a nitrogen source and of the addition of different nitrogen-containing compounds on the extractable activity and the enzyme and mRNA accumulation of adenosine 5'-phosphosulfate reductase (APR). During 72 h without a nitrogen source, the APR activity decreased to 70% and 50% of controls in leaves and roots, respectively, while cysteine (Cys) and glutathione contents were not affected. Northern and western analysis revealed that the decrease of APR activity was correlated with decreased mRNA and enzyme levels. The reduced APR activity in roots could be fully restored within 24 h by the addition of 4 mM each of NO(3)(-), NH(4)(+), or glutamine (Gln), or 1 mM O-acetylserine (OAS). (35)SO(4)(2-) feeding showed that after addition of NH(4)(+), Gln, or OAS to nitrogen-starved plants, incorporation of (35)S into proteins significantly increased in roots; however, glutathione and Cys labeling was higher only with Gln and OAS or with OAS alone, respectively. OAS strongly increased mRNA levels of all three APR isoforms in roots and also those of sulfite reductase, Cys synthase, and serine acetyltransferase. Our data demonstrate that sulfate reduction is regulated by nitrogen nutrition at the transcriptional level and that OAS plays a major role in this regulation. PMID- 10712538 TI - Genetic engineering of glycinebetaine production toward enhancing stress tolerance in plants: metabolic limitations. AB - Glycinebetaine (betaine) affords osmoprotection in bacteria, plants and animals, and protects cell components against harsh conditions in vitro. This and a compelling body of other evidence have encouraged the engineering of betaine production in plants lacking it. We have installed the metabolic step for oxidation of choline, a ubiquitous substance, to betaine in three diverse species, Arabidopsis, Brassica napus, and tobacco (Nicotiana tabacum), by constitutive expression of a bacterial choline oxidase gene. The highest levels of betaine in independent transgenics were 18.6, 12.8, and 13 micromol g(-1) dry weight, respectively, values 10- to 20-fold lower than the levels found in natural betaine producers. However, choline-fed transgenic plants synthesized substantially more betaine. Increasing the choline supplementation further enhanced betaine synthesis, up to 613 micromol g(-1) dry weight in Arabidopsis, 250 micromol g(-1) dry weight in B. napus, and 80 micromol g(-1) dry weight in tobacco. These studies demonstrate the need to enhance the endogenous choline supply to support accumulation of physiologically relevant amounts of betaine. A moderate stress tolerance was noted in some but not all betaine-producing transgenic lines based on relative shoot growth. Furthermore, the responses to stresses such as salinity, drought, and freezing were variable among the three species. PMID- 10712539 TI - Salicylic acid mediated by the oxidative burst is a key molecule in local and systemic responses of cotton challenged by an avirulent race of Xanthomonas campestris pv malvacearum. AB - We analyzed the production of reactive oxygen species, the accumulation of salicylic acid (SA), and peroxidase activity during the incompatible interaction between cotyledons of the cotton (Gossypium hirsutum) cv Reba B50/Xanthomonas campestris pv malvacearum (Xcm) race 18. SA was detected in petioles of cotyledons 6 h after infection and 24 h post inoculation in cotyledons and untreated leaves. The first peak of SA occurred 3 h after generation of superoxide (O(2)(.-)), and was inhibited by infiltration of catalase. Peroxidase activity and accumulation of SA increased in petioles of cotyledons and leaves following H(2)O(2) infiltration of cotyledons from 0.85 to 1 mM. Infiltration of 2 mM SA increased peroxidase activity in treated cotyledons and in the first leaves, but most of the infiltrated SA was rapidly conjugated within the cotyledons. When increasing concentrations of SA were infiltrated 2. 5 h post inoculation at the beginning of the oxidative burst, the activity of the apoplastic cationic O(2)(.-)-generating peroxidase decreased in a dose-dependent manner. We have shown that during the cotton hypersensitive response to Xcm, H(2)O(2) is required for local and systemic accumulation of SA, which may locally control the generation of O(2)(.-). Detaching cotyledons at intervals after inoculation demonstrated that the signal leading to systemic accumulation of SA was emitted around 3 h post inoculation, and was associated with the oxidative burst. SA produced 6 h post infection at HR sites was not the primary mobile signal diffusing systemically from infected cotyledons. PMID- 10712540 TI - Coordinate changes in carbon partitioning and plastidial metabolism during the development of oilseed rape embryos. AB - Measurements of metabolic fluxes in whole embryos and isolated plastids have revealed major changes in the pathways of carbon utilization during cotyledon filling by oilseed rape (Brassica napus L.) embryos. In the early cotyledon stage (stage A), embryos used sucrose (Suc) predominantly for starch synthesis. Plastids isolated from these embryos imported glucose-6-phosphate (Glc-6-P) and partitioned it to starch and fatty acids synthesis and to the oxidative pentose phosphate pathway in the ratio of 2:1:1 on a hexose basis. Of the substrates tested, Glc-6-P gave the highest rates of fatty acid synthesis by the plastids and pyruvate was used weakly. By the mid- to late-cotyledon stage (stage C), oil accumulation by the embryos was rapid, as was their utilization of Suc for oil synthesis in vitro. Plastids from C-stage embryos differed markedly from those of stage-A embryos: (a) pyruvate uptake and utilization for fatty acid synthesis increased by respectively 18- and 25-fold; (b) Glc-6-P partitioning was predominantly to the oxidative pentose phosphate pathway (respective ratios of 1:1:3); and (c) the rate of plastidial fatty acid synthesis more than doubled. This increased rate of fatty synthesis was dependent upon the increase in pyruvate uptake and was mediated through the induction of a saturable transporter activity. PMID- 10712541 TI - Spatial regulation of pectic polysaccharides in relation to pit fields in cell walls of tomato fruit pericarp. AB - Scanning electron microscopic examination of intact tomato (Lycopersicon esculentum) pericarp and isolated pericarp cell walls revealed pit fields and associated radiating ridges on the inner face of cell walls. In regions of the cell wall away from pit fields, equivalent ridges occurred in parallel arrays. Treatment of isolated cell walls with a calcium chelator resulted in the loss of these ridges, indicating that they contain homogalacturonan-rich pectic polysaccharides. Immunolabeling procedures confirmed that pit fields and associated radiating ridges contained homogalacturonan. Epitopes of the side chains of pectic polysaccharides were not located in the same regions as homogalacturonan and were spatially regulated in relation to pit fields. A (1- >4)-beta-galactan epitope was absent from cell walls in regions of pit fields. A (1-->5)-alpha-arabinan epitope occurred most abundantly at the inner face of cell walls in regions surrounding the pit fields. PMID- 10712542 TI - Isolation and characterization of HvNRT2.3 and HvNRT2.4, cDNAs encoding high affinity nitrate transporters from roots of barley. AB - Two full-length cDNAs, HvNRT2.3 and HvNRT2.4, were isolated from roots of barley (Hordeum vulgare), using reverse transcriptase-PCR and RACE-PCR. The corresponding polypeptides, consisting of 507 amino acids (molecular masses of 54.6 kD), belong to the major facilitator superfamily (MFS), and are closely related (>87% identity) to those encoded by HvNRT2.1 and HvNRT2.2 (formerly BCH1 and BCH2, respectively) from roots of barley. The latter are considered to encode inducible high-affinity NO(3)(-) transporters (Trueman et al., 1996). HvNRT2 transcripts were undetectable in NO(3)(-)-deprived plants. Following exposure to either NO(3)(-) or NO(2)(-), transcript abundance and (13)NO(3)(-) influx increased to a maximum by 6 to 12 h, then declined in HvNRT2.1, HvNRT2.2, and HvNRT2.3. The pattern of HvNRT2.4 transcript abundance was different, remaining high after achieving peak abundance. When external NO(3)(-) concentrations were varied from 0 to 500 microM under steady-state conditions of NO(3)(-) supply, HvNRT2 transcript accumulation and (13)NO(3)(-) influx were highest in 50 microM NO(3)(-) -grown plants. When NH(4)(+) was provided together with NO(3)(-), transcript accumulation during the first 2 h was similar to that due to NO(3)(-) alone, but by 4 h the transcript level was significantly reduced. HvNRT2 transcript was undetectable in leaf tissues. PMID- 10712543 TI - Detoxification of arsenic by phytochelatins in plants. AB - As is a ubiquitous element present in the atmosphere as well as in the aquatic and terrestrial environments. Arsenite and arsenate are the major forms of As intoxication, and these anions are readily taken up by plants. Both anions efficiently induce the biosynthesis of phytochelatins (PCs) ([gamma-glutamate cysteine](n)-glycine) in vivo and in vitro. The rapid induction of the metal binding PCs has been observed in cell suspension cultures of Rauvolfia serpentina, in seedlings of Arabidopsis, and in enzyme preparations of Silene vulgaris upon challenge to arsenicals. The rate of PC formation in enzyme preparations was lower compared with Cd-induced biosynthesis, but was accompanied by a prolonged induction phase that resulted finally in higher peptide levels. An approximately 3:1 ratio of the sulfhydryl groups from PCs to As is compatible with reported As-glutathione complexes. The identity of the As-induced PCs and of reconstituted metal-peptide complexes has unequivocally been demonstrated by electrospray ionization mass spectroscopy. Gel filtration experiments and inhibitor studies also indicate a complexation and detoxification of As by the induced PCs. PMID- 10712544 TI - Differential screening indicates a dramatic change in mRNA profiles during grape berry ripening. Cloning and characterization of cDNAs encoding putative cell wall and stress response proteins. AB - We used differential screening to isolate ripening-associated cDNAs from a Shiraz grape (Vitis vinifera L.) berry cDNA library. A rapid increase in the mRNA levels of a number of cDNAs not present in unripe fruit occurred in grape berries at the onset of ripening. The putative translation products of some of these clones had homologs in other species that are involved in cell wall structure. These included four proline-rich proteins, a small protein that is similar to the non catalytic, N-terminal domain of some pectin methylesterases, and two other glutamate-rich proteins. The remainder of the clones encoded putative stress response proteins. These included two thaumatin-like proteins, a metallothionein, a transcription factor, a cytochrome P450 enzyme, and proteins induced by water, sugar, and/or cold stress in other species. Many of the homologs of the grape cDNAs thought to be involved in cell wall structure or stress-related responses also accumulate in a developmental manner in other plants. This may indicate that the grape mRNAs accumulate in response to stresses such as the storage of high concentrations of sugars and rapid cell expansion, or they may accumulate as part of the ripening developmental program. PMID- 10712545 TI - Initial binding of preproteins involving the Toc159 receptor can be bypassed during protein import into chloroplasts. AB - Two integral outer envelope GTPases, Toc34 and Toc86, are proposed to regulate the recognition and translocation of nuclear-encoded preproteins during the early stages of protein import into chloroplasts. Defining the precise roles of Toc86 and Toc34 has been complicated by the inability to distinguish their GTPase activities. Furthermore, the assignment of Toc86 function is rendered equivocal by recent reports suggesting that the standard protocol for the isolation of chloroplasts results in significant proteolysis of Toc86 (B. Bolter, T. May, J. Soll [1998] FEBS Lett 441: 59-62; G. Schatz [1998] Nature 395: 439-440). We demonstrate that Toc86 corresponds to a native protein of 159 kD in pea (Pisum sativum), designated Toc159. We take advantage of the proteolytic sensitivity of Toc159 to selectively remove its 100-kD cytoplasmic GTPase domain and thereby distinguish its activities from other import components. Proteolysis eliminates detectable binding of preproteins at the chloroplast surface, which is consistent with the proposed role of Toc159 as a receptor component. Remarkably, preprotein translocation across the outer membrane can occur in the absence of the Toc159 cytoplasmic domain, suggesting that binding can be bypassed. Translocation remains sensitive to GTP analogs in the absence of the Toc159 GTP-binding domain, providing evidence that Toc34 plays a key role in the regulation of translocation by GTP. PMID- 10712546 TI - A weakly voltage-dependent, nonselective cation channel mediates toxic sodium influx in wheat. AB - To determine the transporters responsible for toxic Na(+) influx in wheat (Triticum aestivum), root plasma membrane preparations were screened using the planar lipid bilayer technique as an assay for Na(+)-permeable ion channel activity. The predominant channel in the bilayer was a 44-pS channel that we called the nonselective cation (NSC) channel, which was nonselective for monovalent cations and weakly voltage dependent. Single channel characteristics of the NSC channel were compared with (22)Na(+) influx into excised root segments. Na(+) influx through the NSC channel resembled (22)Na(+) influx in its partial sensitivity to inhibition by Ca(2+), Mg(2+), and Gd(3+), and its insensitivity to all other inhibitors tested (tetraethylammonium, quinine, Cs(+), tetrodotoxin, verapamil, amiloride, and flufenamate). Na(+) influx through the NSC channel also closely resembled an instantaneous current in wheat root protoplasts (S.D. Tyerman, M. Skerrett, A. Garill, G.P. Findlay, R. Leigh [1997] J Exp Bot 48: 459-480) in its permeability sequence, selectivity for K(+) over Na(+) (approximately 1.25), insensitivity to tetraethylammonium, voltage independence, and partial sensitivity to Ca(2+). Comparison of tissue, protoplast (S.D. Tyerman, M. Skerrett, A. Garill, G.P. Findlay, R. Leigh [1997] J Exp Bot 48: 459-480), and single- channel data indicate that toxic Na(+) influx is catalyzed by a single transporter, and this is likely to be the NSC channel identified in planar lipid bilayers. PMID- 10712547 TI - Differential ion accumulation and ion fluxes in the mesophyll and epidermis of barley. AB - In barley (Hordeum vulgare L.) leaves, differential ion accumulation commonly results in inorganic phosphate (Pi) being confined to the mesophyll and Ca(2+) to the epidermis, with preferential epidermal accumulation of Cl(-), Na(+), and some other ions. The pattern was confirmed in this study for major inorganic anions and cations by analysis of barley leaf protoplasts. The work focused on the extent to which differences in plasma membrane ion transport processes underlie these observations. Ion transport across the plasma membrane of barley epidermal and mesophyll protoplasts was investigated electrophysiologically (by microelectrode impalement and patch clamping) and radiometrically. Data from both approaches suggested that similar types of ion-selective channels and membrane transporters, which catalyze the transport of Ca(2+), K(+), Na(+), and Pi, exist in the plasma membrane of the two cell types. In general, the simple presence or absence of ion transporters could not explain cell-type-specific differences in ion accumulation. However, patch-clamp data suggested that differential regulation of instantaneously activating ion channels in the plasma membrane could explain the preferential accumulation of Na(+) in the epidermis. PMID- 10712548 TI - Import of lyso-phosphatidylcholine into chloroplasts likely at the origin of eukaryotic plastidial lipids. AB - Plastids rely on the import of extraplastidial precursor for the synthesis of their own lipids. This key phenomenon in the formation of plastidial phosphatidylcholine (PC) and of the most abundant lipids on earth, namely galactolipids, is poorly understood. Various suggestions have been made on the nature of the precursor molecule(s) transferred to plastids, but despite general agreement that PC or a close metabolite plays a central role, there is no clear cut answer to this question because of a lack of conclusive experimental data. We therefore designed experiments to discriminate between a transfer of PC, 1 acylglycero phosphorylcholine (lyso-PC), or glycerophosphorylcholine. After pulse chase experiments with glycerol and acetate, plastids of leek (Allium porrum L.) seedlings were purified. The labels of the glycerol moiety and the sn-1- and sn-2 bound fatty acids of plastidial lipids were determined and compared with those associated with the extraplastidial PC. After import, plastid lipids contained the glycerol moiety and the fatty acids esterified to the sn-1 position originating from the extraplastidial PC; no import of sn-2-bound fatty acid was detected. These results rule out a transfer of PC or glycerophosphorylcholine, and are totally explained by an import of lyso-PC molecules used subsequently as precursor for the synthesis of eukaryotic plastid lipids. PMID- 10712549 TI - Characterization of XET-related genes of rice. AB - To elucidate the mechanism of internodal elongation in rice (Oryza sativa L.), we analyzed genes encoding xyloglucan endotransglycosylase (XET), a cell wall loosening enzyme essential for cell elongation. Four rice XET-related (XTR) genes, OsXTR1, OsXTR2, OsXTR3, and OsXTR4, were isolated and their expression patterns in rice plants determined. The expression of the four XTR genes showed different patterns of organ specificity and responses to several plant hormones. OsXTR1 and OsXTR3 were up-regulated by gibberellin and brassinosteroids, whereas OsXTR2 and OsXTR4 showed no clear response to these hormones. Expression of the four XTR genes was also investigated in elongating internodes at different developmental stages. OsXTR1 and OsXTR3 were preferentially expressed in the elongating zone of internodes, while OsXTR2 and OsXTR4 were expressed in nodes and in the divisional and elongating zones of internodes. In three genetic mutants with abnormal heights, the expression of OsXTR1 and OsXTR3 correlated with the height of the mutants, whereas no such correlation was observed for OsXTR2 and OsXTR4. Based on these observations, we discuss the roles that OsXTR1 and OsXTR3 may play in internodal elongation in rice. PMID- 10712550 TI - Thermal effect of CO(2) on apoplastic ice in rye and oat during freezing. AB - Meristematic tissues from rye (Secale cereale) and oat (Avena sativa) were studied in an isothermal calorimeter at -3 degrees C. When the frozen tissue was placed in the calorimeter, the pressure increased within 4 d to 25 and 9 kPa above ambient pressure in the sample vessels containing crowns of rye and oat, respectively. Concurrently, the thermal output went down to -194 microW in rye over the 4-d period; this negative thermal activity could be accounted for by ice melting in the plants. When the pressure was released, the output from the calorimeter went from -194 to 229 microW within 1 h, suggesting that water had frozen in the plants. We propose that CO(2) from respiration had dissolved in the water in the plants and caused melting of ice (heat absorption) due to the colligative properties of solutions. When the pressure was released, the CO(2) came out of solution and the water froze (heat evolution). These thermal observations were duplicated in a simplified, non-biological system using a glycol/water mixture that was partially frozen at -3 degrees C. PMID- 10712551 TI - GDP-fucose uptake into the Golgi apparatus during xyloglucan biosynthesis requires the activity of a transporter-like protein other than the UDP-glucose transporter. AB - The molecular mechanisms regulating hemicelluloses and pectin biosynthesis are poorly understood. An important question in this regard is how glycosyltransferases are oriented in the Golgi cisternae, and how nucleotide sugars are made available for the synthesis of the polymers. Here we show that the branching enzyme xyloglucan alpha,1-2 fucosyltransferase (XG-FucTase) from growing pea (Pisum sativum) epicotyls was latent and protected against proteolytic inactivation on intact, right-side-in pea stem Golgi vesicles. Moreover, much of the XG-FucTase activity was membrane associated. These data indicate that XG-FucTase is a membrane-bound luminal enzyme. GDP-Fuc uptake studies demonstrated that GDP-Fuc was taken up into Golgi vesicles in a protein mediated process, and that this uptake was not competed by UDP-Glc, suggesting that a specific GDP-Fuc transporter is involved in xyloglucan biosynthesis. Once in the lumen, Fuc was transferred onto endogenous acceptors, including xyloglucan. GDPase activity was detected in the lumen of the vesicles, suggesting than the GDP produced upon transfer of Fuc was hydrolyzed to GMP and inorganic phosphate. We suggest than the GDP-Fuc transporter and GDPase may be regulators of xyloglucan fucosylation in the Golgi apparatus from pea epicotyls. PMID- 10712552 TI - Sodium-dependent nitrate transport at the plasma membrane of leaf cells of the marine higher plant Zostera marina L. AB - NO(3)(-) is present at micromolar concentrations in seawater and must be absorbed by marine plants against a steep electrochemical potential difference across the plasma membrane. We studied NO(3)(-) transport in the marine angiosperm Zostera marina L. to address the question of how NO(3)(-) uptake is energized. Electrophysiological studies demonstrated that micromolar concentrations of NO(3)(-) induced depolarizations of the plasma membrane of leaf cells. Depolarizations showed saturation kinetics (K(m) = 2.31 +/- 0.78 microM NO(3)(-)) and were enhanced in alkaline conditions. The addition of NO(3)(-) did not affect the membrane potential in the absence of Na(+), but depolarizations were restored when Na(+) was resupplied. NO(3)(-)-induced depolarizations at increasing Na(+) concentrations showed saturation kinetics (K(m) = 0.72 +/- 0.18 mM Na(+)). Monensin, an ionophore that dissipates the Na(+) electrochemical potential, inhibited NO(3)(-)-evoked depolarizations by 85%, and NO(3)(-) uptake (measured by depletion from the external medium) was stimulated by Na(+) ions and by light. Our results strongly suggest that NO(3)(-) uptake in Z. marina is mediated by a high-affinity Na(+)-symport system, which is described here (for the first time to our knowledge) in an angiosperm. Coupling the uptake of NO(3)(-) to that of Na(+) enables the steep inwardly-directed electrochemical potential for Na(+) to drive net accumulation of NO(3)(-) within leaf cells. PMID- 10712553 TI - Analysis of reductant supply systems for ferredoxin-dependent sulfite reductase in photosynthetic and nonphotosynthetic organs of maize. AB - Sulfite reductase (SiR) catalyzes the reduction of sulfite to sulfide in chloroplasts and root plastids using ferredoxin (Fd) as an electron donor. Using purified maize (Zea mays L.) SiR and isoproteins of Fd and Fd-NADP(+) reductase (FNR), we reconstituted illuminated thylakoid membrane- and NADPH-dependent sulfite reduction systems. Fd I and L-FNR were distributed in leaves and Fd III and R-FNR in roots. The stromal concentrations of SiR and Fd I were estimated at 1.2 and 37 microM, respectively. The molar ratio of Fd III to SiR in root plastids was approximately 3:1. Photoreduced Fd I and Fd III showed a comparable ability to donate electrons to SiR. In contrast, when being reduced with NADPH via FNRs, Fd III showed a several-fold higher activity than Fd I. Fd III and R FNR showed the highest rate of sulfite reduction among all combinations tested. NADP(+) decreased the rate of sulfite reduction in a dose-dependent manner. These results demonstrate that the participation of Fd III and high NADPH/NADP(+) ratio are crucial for non-photosynthetic sulfite reduction. In accordance with this view, a cysteine-auxotrophic Escherichia coli mutant defective for NADPH dependent SiR was rescued by co-expression of maize SiR with Fd III but not with Fd I. PMID- 10712554 TI - Salicylates of intact Salix myrsinifolia plantlets do not undergo rapid metabolic turnover. AB - Salicylates, the main phenolic glucosides of northern willow (Salix spp.), play an important role in plant-herbivore interactions. Salicylates are labile metabolites that are thought to undergo metabolic turnover. Salicylates are synthesized from phenylalanine (Phe) via the shikimate pathway. 2-Aminoindan-2 phosphonic acid (AIP), a strong inhibitor of Phe ammonia-lyase (EC 4.3.1.5), was used to block the biosynthesis of salicylates. The aim of this study was to investigate long-term turnover of salicylates in intact micropropagated plantlets of Salix myrsinifolia Salisb. The biosynthesis of salicylates was inhibited efficiently but not completely by 30 microM 2-aminoindan-2-phosphonic acid. Inhibitor treatment, aside from leading to a high accumulation of Phe, also led to an increase in tyrosine and tryptophan, indicating that 2-aminoindan-2 phosphonic acid may also inhibit enzymes other than Phe ammonia-lyase. Salicylates were shown to be unexpectedly stable metabolites that did not undergo marked metabolic turnover in intact plants; in leaves no significant turnover occurred, and in the stems the five salicylates studied were turned over slowly, with half-lives of 11 to 25 d. The total amount of salicylate in mature shoots decreased only 0.6% per day. PMID- 10712555 TI - Molecular cloning and characterization of ATP-phosphoribosyl transferase from Arabidopsis, a key enzyme in the histidine biosynthetic pathway. AB - We have characterized two isoforms of ATP-phosphoribosyl transferase (ATP-PRT) from Arabidopsis (AtATP-PRT1 [accession no. AB025251] and AtATP-PRT2), catalyzing the first step of the pathway of hisidine (His) biosynthesis. The primary structures deduced from AtATP-PRT1 and AtATP-PRT2 cDNAs share an overall amino acid identity of 74.6% and contain N-terminal chloroplast transit peptide sequences. DNA-blot analyses indicated that the ATP-PRTs in Arabidopsis are encoded by two separate genes with a closely similar gene structural organization. Both gene transcripts were detected throughout development, and protein-blot analysis revealed predominant accumulation of the AtATP-PRT proteins in Arabidopsis leaves. The His auxotrophy of a his1 mutant of Saccharomyces cerevisiae was suppressed by the transformation with AtATP-PRT1 and AtATP-PRT2 cDNAs, indicating that both isoforms are functionally active ATP-PRT enzymes. The K(m) values for ATP and phosphoribosyl pyrophosphate of the recombinant AtATP-PRT proteins were comparable to those of the native ATP-PRTs from higher plants and bacteria. It was demonstrated that the recombinant AtATP-PRTs were inhibited by L His (50% inhibition of initial activity = 40-320 microM), suggesting that His biosynthesis was regulated in plants through feedback inhibition by L-His. PMID- 10712556 TI - Leaf respiration of snow gum in the light and dark. Interactions between temperature and irradiance. AB - We investigated the effect of temperature and irradiance on leaf respiration (R, non-photorespiratory mitochondrial CO(2) release) of snow gum (Eucalyptus pauciflora Sieb. ex Spreng). Seedlings were hydroponically grown under constant 20 degrees C, controlled-environment conditions. Measurements of R (using the Laisk method) and photosynthesis (at 37 Pa CO(2)) were made at several irradiances (0-2,000 micromol photons m(-2) s(-1)) and temperatures (6 degrees C 30 degrees C). At 15 degrees C to 30 degrees C, substantial inhibition of R occurred at 12 micromol photons m(-2) s(-1), with maximum inhibition occurring at 100 to 200 micromol photons m(-2) s(-1). Higher irradiance had little additional effect on R at these moderate temperatures. The irradiance necessary to maximally inhibit R at 6 degrees C to 10 degrees C was lower than that at 15 degrees C to 30 degrees C. Moreover, although R was inhibited by low irradiance at 6 degrees C to 10 degrees C, it recovered with progressive increases in irradiance. The temperature sensitivity of R was greater in darkness than under bright light. At 30 degrees C and high irradiance, light-inhibited rates of R represented 2% of gross CO(2) uptake (v(c)), whereas photorespiratory CO(2) release was approximately 20% of v(c). If light had not inhibited leaf respiration at 30 degrees C and high irradiance, R would have represented 11% of v(c). Variations in light inhibition of R can therefore have a substantial impact on the proportion of photosynthesis that is respired. We conclude that the rate of R in the light is highly variable, being dependent on irradiance and temperature. PMID- 10712557 TI - Auxin metabolism in the root apical meristem. AB - Within the root meristem of flowering plants is a group of mitotically inactive cells designated the quiescent center (QC). Recent work links the quiescent state to high levels of the growth regulator auxin that accumulates in the QC via polar transport. This in turn results in elevated levels of the enzyme ascorbic acid oxidase (AAO), resulting in a reduction of ascorbic acid (AA) within the QC and mitotic quiescence. We present evidence for additional interactions between auxin, AAO, and AA, and report that, in vitro, AAO oxidatively decarboxylates auxin, suggesting a mechanism for regulating auxin levels within the QC. We also report that oxidative decarboxylation occurs at the root tip and that an intact root cap must be present for this metabolic event to occur. Finally, we consider how interaction between auxin and AAO may influence root development by regulating the formation of the QC. PMID- 10712558 TI - Expression of tryptophan decarboxylase and tyrosine decarboxylase genes in tobacco results in altered biochemical and physiological phenotypes. AB - The substrate specificity of tryptophan (Trp) decarboxylase (TDC) for Trp and tyrosine (Tyr) decarboxylase (TYDC) for Tyr was used to modify the in vivo pools of these amino acids in transgenic tobacco. Expression of TDC and TYDC was shown to deplete the levels of Trp and Tyr, respectively, during seedling development. The creation of artificial metabolic sinks for Trp and Tyr also drastically affected the levels of phenylalanine, as well as those of the non-aromatic amino acids methionine, valine, and leucine. Transgenic seedlings also displayed a root curling phenotype that directly correlated with the depletion of the Trp pool. Non-transformed control seedlings could be induced to display this phenotype after treatment with inhibitors of auxin translocation such as 2,3,5 triiodobenzoic acid or N-1-naphthylphthalamic acid. The depletion of aromatic amino acids was also correlated with increases in the activities of the shikimate and phenylpropanoid pathways in older, light-treated transgenic seedlings expressing TDC, TYDC, or both. These results provide in vivo confirmation that aromatic amino acids exert regulatory feedback control over carbon flux through the shikimate pathway, as well as affecting pathways outside of aromatic amino acid biosynthesis. PMID- 10712559 TI - Genotypical differences in aluminum resistance of maize are expressed in the distal part of the transition zone. Is reduced basipetal auxin flow involved in inhibition of root elongation by aluminum? AB - Short-term Al treatment (90 microM Al at pH 4.5 for 1 h) of the distal transition zone (DTZ; 1-2 mm from the root tip), which does not contribute significantly to root elongation, inhibited root elongation in the main elongation zone (EZ; 2.5-5 mm from the root tip) to the same extent as treatment of the entire maize (Zea mays) root apex. Application of Al to the EZ had no effect on root elongation. Higher genotypical resistance to Al applied to the entire root apex, and specifically to the DTZ, was expressed by less inhibition of root elongation, Al accumulation, and Al-induced callose formation, primarily in the DTZ. A characteristic pH profile along the surface of the root apex with a maximum of pH 5.3 in the DTZ was demonstrated. Al application induced a substantial flattening of the pH profile moreso in the Al-sensitive than in the Al-resistant cultivar. Application of indole-3-acetic acid to the EZ but not to the meristematic zone significantly alleviated the inhibition of root elongation induced by the application of Al to the DTZ. Basipetal transport of exogenously applied [(3)H]indole-3-acetic acid to the meristematic zone was significantly inhibited by Al application to the DTZ in the Al-sensitive maize cv Lixis. Our results provide evidence that the primary mechanisms of genotypical differences in Al resistance are located within the DTZ, and suggest a signaling pathway in the root apex mediating the Al signal between the DTZ and the EZ through basipetal auxin transport. PMID- 10712560 TI - Calcium-regulated proteolysis of eEF1A. AB - Eukaryotic elongation factor 1alpha (eEF1A) can be post-translationally modified by the addition of phosphorylglycerylethanolamine (PGE). [(14)C]Ethanolamine was incorporated into the PGE modification, and with carrot (Daucus carota L.) suspension culture cells, eEF1A was the only protein that incorporated detectable quantities of [(14)C]ethanolamine (Ransom et al., 1998). When 1 mM CaCl(2) was added to microsomes containing [(14)C]ethanolamine-labeled eEF1A ([(14)C]et eEF1A), there was a 60% decrease in the amount of [(14)C]et-eEF1A recovered after 10 min. The loss of endogenous [(14)C]et-eEF1A was prevented by adding EGTA. Recombinant eEF1A, which did not contain the PGE modification, also was degraded by microsomes in a Ca(2+)-regulated manner, indicating that PGE modification was not necessary for proteolysis; however, it enabled us to quantify enodgenous eEF1A. By monitoring [(14)C]et-eEF1A, we found that treatment with phospholipase D or C, but not phospholipase A(2), resulted in a decrease in [(14)C]et-eEF1A from carrot microsomes. The fact that there was no loss of [(14)C]et-eEF1A with phospholipase A(2) treatment even in the presence of 1 mM Ca(2+) suggested that the loss of membrane lipids was not essential for eEF1A proteolysis and that lysolipids or fatty acids decreased proteolysis. At micromolar Ca(2+) concentrations, proteolysis of eEF1A was pH sensitive. When 1 microM CaCl(2) was added at pH 7.2, 35% of [(14)C]et-eEF1A was lost; while at pH 6.8, 10 microM CaCl(2) was required to give a similar loss of protein. These data suggest that eEF1A may be an important downstream target for Ca(2+) and lipid-mediated signal transduction cascades. PMID- 10712561 TI - Abscisic acid accumulation maintains maize primary root elongation at low water potentials by restricting ethylene production. AB - Previous work showed that primary root elongation in maize (Zea mays L.) seedlings at low water potentials (psi(w)) requires the accumulation of abscisic acid (ABA) (R.E. Sharp, Y. Wu, G.S. Voetberg, I.N. Saab, M.E. LeNoble [1994] J Exp Bot 45: 1743-1751). The objective of the present study was to determine whether the inhibition of elongation in ABA-deficient roots is attributable to ethylene. At a psi(w) of -1.6 MPa, inhibition of root elongation in dark-grown seedlings treated with fluridone to impose ABA deficiency was largely prevented with two inhibitors of ethylene synthesis (aminooxyacetic acid and aminoethoxyvinylglycine) and one inhibitor of ethylene action (silver thiosulfate). The fluridone treatment caused an increase in the rate of ethylene evolution from intact seedlings. This effect was completely prevented with aminooxyacetic acid and also when ABA was supplied at a concentration that restored the ABA content of the root elongation zone and the root elongation rate. Consistent results were obtained when ABA deficiency was imposed using the vp5 mutant. Both fluridone-treated and vp5 roots exhibited additional morphological symptoms of excess ethylene. The results demonstrate that an important role of ABA accumulation in the maintenance of root elongation at low psi(w) is to restrict ethylene production. PMID- 10712563 TI - The electronic plant gene register. PMID- 10712562 TI - Characterization of ripening-regulated cDNAs and their expression in ethylene suppressed charentais melon fruit. AB - Charentais melons (Cucumis melo cv Reticulatus) are climacteric and undergo extremely rapid ripening. Sixteen cDNAs corresponding to mRNAs whose abundance is ripening regulated were isolated to characterize the changes in gene expression that accompany this very rapid ripening process. Sequence comparisons indicated that eight of these cDNA clones encoded proteins that have been previously characterized, with one corresponding to ACC (1-aminocyclopropane-1-carboxylic acid) oxidase, three to proteins associated with pathogen responses, two to proteins involved in sulfur amino acid biosynthesis, and two having significant homology to a seed storage protein or a yeast secretory protein. The remaining eight cDNA sequences did not reveal significant sequence similarities to previously characterized proteins. The majority of the 16 ripening-regulated cDNAs corresponded to mRNAs that were fruit specific, although three were expressed at low levels in vegetative tissues. When examined in transgenic antisense ACC oxidase melon fruit, three distinct patterns of mRNA accumulation were observed. One group of cDNAs corresponded to mRNAs whose abundance was reduced in transgenic fruit but inducible by ethylene treatment, indicating that these genes are directly regulated by ethylene. A second group of mRNAs was not significantly altered in the transgenic fruit and was unaffected by treatment with ethylene, indicating that these genes are regulated by ethylene-independent developmental cues. The third and largest group of cDNAs showed an unexpected pattern of expression, with levels of mRNA reduced in transgenic fruit and remaining low after exposure to ethylene. Regulation of this third group of genes thus appears to ethylene independent, but may be regulated by developmental cues that require ethylene at a certain stage in fruit development. The results confirm that both ethylene-dependent and ethylene-independent pathways of gene regulation coexist in climacteric fruit. PMID- 10712564 TI - On the direction and velocity of blood flow in the extradural intravertebral vein of harp seals (Phoca groenlandica) during simulated diving. AB - Ronald et al. (1977) suggested that blood flow in the caudal/lumbar sections of the extradural intravertebral vein (EIV) of seals changes direction from running towards the head before diving, to the opposite during diving. The possible advantage would be that the oxygen-depleted venous effluent from the brain is routed via the EIV to the posterior parts of the hepatic sinuses and the inferior caval vein and, hence, is prevented from mixing with the more oxygen-rich venous blood in their anterior parts. We have re-examined this hypothesis by use of Doppler flowmetry. A catheter-tip flow probe was introduced into the EIV of two similar-sized juvenile harp seals, and flow direction and rate determined before, during and after simulated dives lasting for 5 min, at three positions (caudal, lumbar and thoracic) along the EIV. Regardless of probe position, blood was mainly flowing towards the head in 11 of 13 experiments prior to diving, in 8 of 13 experiments during diving and in 11 of 13 experiments during recovery after diving (and away from the head in the remaining experiments). Flow direction was most variable in the caudal position. Mean blood velocity in the EIV was substantially lower during diving (0.10 +/- 0.22 cm s-1 (n=5) in thoracic position) than in the pre-dive (3.98 +/- 3.32 cm s-1 [n=5]) and post-dive (5.75 +/- 4.07 cm s-1 [n=5]) situations. Thus, the direction and rate of flow in the EIV was variable, particularly during diving, as is to be expected in a system of anastomosing, valveless veins. We conclude that the hypothesis of Ronald et al. (1977) most likely is false. PMID- 10712565 TI - The effect of alpha-phenyl-tert-butyl nitrone (PBN) on free radical formation in transient focal ischaemia measured by microdialysis and 3,4-dihydroxybenzoate formation. AB - alpha-phenyl-tert-butyl nitrone (PBN) reduces infarct size, improves recovery of brain energy metabolism and delays the secondary increase in extracellular potassium after focal ischaemia, presumably by trapping OH radicals. We investigated the effect of PBN on the formation of 3,4-dihydroxybenzoic acid (3,4 DHBA) as a measure of OH radical formation, during and following middle cerebral artery occlusion (MCAO). Rats, subjected to 2 h of ischaemia followed by 3 h of recirculation, were injected with either vehicle or PBN (100 mg kg-1 i.p.) prior to MCAO or immediately after recirculation, respectively. The in vivo microdialysis technique was used to collect samples for analysis of 3,4-DHBA by HPLC. The basal levels of 3,4-DHBA were 56-77 nmol L-1 in the four groups. During ischaemia, the formation of 3,4-DHBA decreased by about 50% in all groups. Upon recirculation, a 3-fold rise in 3,4-DHBA formation was seen. At 2 h of recirculation the mean value of 3,4-DHBA in the pretreated, vehicle-injected animals was 125 +/- 18 nmol L-1 and in the PBN-injected 145 +/- 48 nmol L-1, respectively. When the animals were treated after MCAO either with vehicle or PBN the values at 2 h recirculation were 155 +/- 148 and 189 +/- 145 nmol L-1, respectively. No statistically significant difference between vehicle- and PBN injected groups was seen. We conclude that during reperfusion following MCAO, hydroxyl radical formation increases. The increase is not ameliorated by PBN which suggests that PBN does not protect the brain by a general scavenging of OH radicals, although tissue specific actions cannot be excluded. PMID- 10712566 TI - Tyramine-induced endogenous noradrenaline efflux from in situ cardiac sympathetic nerve ending in cats. AB - With the use of dialysis technique, the effects of tyramine on in situ cardiac sympathetic nerve endings were examined in anaesthetized cats. Dialysis probes were implanted in the left ventricular myocardium, and the concentration of dialysate noradrenaline (NA) served as an indicator of NA output at the cardiac sympathetic nerve ending. Locally applied tyramine (600 microM) increased dialysate NA levels from 17 +/- 1 (pg mL-1) to 3466 +/- 209 (pg mL-1). Pretreatment with reserpine (vesicle transport NA blocker 1 microM) did not affect tyramine-induced NA efflux. The tyramine-induced NA efflux was augmented by pretreatment with pargyline (1 mM) but suppressed by pargyline (10 mM). Pretreatment with alpha-methyl-tyrosine suppressed NA efflux evoked by tyramine. These pretreatments did not affect the time course of NA efflux but only altered peak height of NA efflux. The efflux of NA evoked by tyramine was not associated with any reduction of dihydroxyphenylglycol (DHPG). In contrast, in the pretreatment with reserpine, the efflux of NA was associated with a reduction of DHPG. This result suggests that NA graduation between axoplasm and stored vesicle contributes to maintaining the axoplasmic NA level during carrier-mediated outward NA transport. The tyramine-induced NA efflux provides a close reflection of the NA content at the nerve ending. With the use of dialysis, this experimental model is suitable for studying the mechanism of sympathomimetic amine-induced neurotransmitter efflux. PMID- 10712567 TI - Further evidence for the existence of a volume-activated taurine efflux pathway in rat mammary tissue independent from volume-sensitive Cl- channels. AB - The effect of cell-swelling, induced by a hyposmotic shock, on the fractional loss of taurine, D-aspartate and iodide from lactating rat mammary tissue explants has been examined. In paired experiments, cell-swelling markedly increased the fractional efflux of taurine but not that of D-aspartate. Similarly, in paired experiments, a hyposmotic challenge stimulated taurine release but not iodide efflux. The results suggest that volume-activated taurine efflux from mammary tissue explants is via a pathway independent from volume sensitive anion channels. It is apparent that the volume-activated taurine efflux pathway in mammary tissue is not the volume-sensitive organic osmolyte-anion channel which has been described in other cells. Therefore, the results of this study together with others in the literature constitute prima facie evidence for the existence of more than one type of swelling-activated pathway which accepts taurine as a substrate. PMID- 10712568 TI - Angiotensin II type 1 receptors stimulate protein synthesis in human cardiac fibroblasts via a Ca2+-sensitive PKC-dependent tyrosine kinase pathway. AB - The aim of the present study was to investigate the proliferative effects of Ang II in human cardiac fibroblasts. The effects of Ang II in human cardiac fibroblasts on the 3H-thymidine incorporation, the cell number, the 3H-leucine incorporation and the total protein content were measured. The expression of receptor mRNA was performed by reverse transcription-polymerase chain reaction (RT-PCR). Ang II increased 3H-leucine incorporation in a concentration-dependent manner but not 3H-thymidine incorporation in primary cultures of human cardiac fibroblasts. The maximum effect (24 +/- 3% over control) was obtained at a concentration of 10 nM. There were no significant alterations of cell number or total protein content, suggesting that Ang II stimulated protein synthesis but did not induce hypertrophy. The accumulation of 3H-leucine was blocked by the AT1 receptor antagonist candesartan but not by the AT2 receptor antagonist PD123319. By using RT-PCR, both AT1 and AT2 receptors mRNA were found to be expressed in human cardiac fibroblasts. The selective MAPKK inhibitor PD098059, the protein kinase C inhibitor K252a or the phospholipase C inhibitor U73122 did not significantly inhibit Ang II augmented 3H-leucine incorporation. However, this was significantly blocked by the Ca2+-dependent protein kinase C inhibitor GO6976, the non-selective protein kinase inhibitor staurosporine and the tyrosine kinase inhibitor tyrphostin 25. The effects of Ang II were unaffected by the Gi protein blocker pertussis toxin, indicating a Gi-protein-independent pathway. Ang II was synergistic with insulin but showed no significant increase on 3H-leucine incorporation when combined with PDGF or EGF. In summary, Ang II stimulates protein synthesis through AT1 receptors in human cardiac fibroblasts, but has no hypertrophic or hyperplastic effect. The response is mediated by a MAPKK independent and Ca2+-sensitive PKC-dependent pathway. PMID- 10712569 TI - Arterio-venous differences of blood acid-base status and plasma sodium caused by intense bicycling. AB - After intense exercise muscle may give off hydrogen ions independently of lactate, perhaps by a mechanism involving sodium ions. To examine this possibility further five healthy young men cycled for 2 min to exhaustion. Blood was drawn from catheters in the femoral artery and vein during exercise and at 1 h intervals after exercise. The blood samples were analysed for pH, blood gases, lactate, haemoglobin, and plasma proteins and electrolytes. Base deficit was calculated directly without using common approximations. The leg blood flow was also measured, thus allowing calculations of the leg's exchange of metabolites. The arterial blood lactate concentration rose to 14.2 +/- 1.0 mmol L-1, the plasma pH fell to 7. 18 +/- 0.02, and the base deficit rose 22% more than the blood lactate concentration did. The femoral-venous minus arterial differences peaked at 1.8 +/- 0.2 mmol L-1 (lactate), -0.24 +/- 0.01 (pH), and 4.5 +/- 0.4 mmol L-1 (base deficit), and -2.5 +/- 0.7 mmol L-1 (plasma sodium concentration corrected for volume changes). Thus, near the end of the exercise and for the first 10 min of the recovery period the leg gave off more hydrogen ions than lactate ions to the blood, and sodium left plasma in proportion to the extra hydrogen ions appearing. The leg's integrated excess release of hydrogen ions of 0.88 +/- 0.45 mmol kg-1 body mass was 67% of the integrated lactate release. Base deficit calculated by the traditional approximate equations underestimated the true value, but the error was less than 10%. We conclude that intense exercise and lactic acidosis may lead to a muscle release of hydrogen ions independent of lactate release, possibly by a Na+,H+ exchange. Hydrogen ions were largely buffered in the red blood cells. PMID- 10712570 TI - Elasticity of tendon structures of the lower limbs in sprinters. AB - The present study aims to investigate the elasticity of tendon structures of the lower limbs in sprinters and its relation with sprint performance. Subjects were 10 male sprinters and 14 controls whose anthropometric variables and isometric maximum strength were similar. The elongation (L) of the tendon and aponeurosis of vastus lateralis (VL) and medial gastrocnemius muscles (MG) during isometric knee extension and planter flexion, respectively, were determined using a real time ultrasonic apparatus in vivo, while the subjects developed a gradually increasing torque from zero (relax) to maximal effort (MVC) within 5 s. While sprinters compared with controls showed significantly greater L above 500 N (about 50% of MVC) and higher dL/dF for VL at less than 20% of MVC during knee extension, there were no significant differences between the two groups in L and dL/dF for MG at every 10% of MVC during plantar flexion. Moreover, the average value of dL/dF above 50% of MVC, proposed as the compliance of tendon structures, did not significantly differ between sprinters and controls in either VL or MG. In a regression analysis within sprinters, the compliance of VL was negatively correlated to 100-m sprint time, r=-0.757 (P < 0.05), but that of MG was not, r=0.228 (P > 0.05). Thus the present results indicate that the elasticity of tendon structures of VL and MG at high force production levels, which might be assumed to associate with the storage and subsequent release of energy during exercises involving the stretch-shortening cycle, are similar in both sprinters and controls. For sprinters, however, the tendon structures of VL are more compliant than that for controls at low force production levels, and its elasticity at high force production levels may influence sprint performance. PMID- 10712571 TI - The effects of a prolonged undernutrition on serum lipids and fatty acid composition of reindeer calves during winter and spring. AB - We examined the effects of undernutrition on lipid metabolism in reindeer (<1 year) during mid-winter and spring, with particular focus on the proportions of polyunsaturated fatty acids (PUFAs) in major serum lipids. The reindeer (n=8) were fed their winter feed, lichen, ad libitum for 5 weeks, followed by 40% restriction of energy for 8 weeks and refeeding to normal for 6 weeks. The concentrations of major serum lipids, cholesterol and phospholipids decreased significantly during the ad libitum period (by 50 and 44%, respectively). The proportion of major PUFA, linoleic acid in serum cholesteryl esters, decreased from 48.2 to 38.4% during the ad libitum period (P < 0.01), and to 29.2% during the restriction period (P < 0.001). The proportion of linoleic acid in phospholipids decreased from 27.9 to 15.6% during the ad libitum period (P < 0. 001), and to 13.0% during the restriction (P < 0.01). Also alpha-linolenic acid in the major lipids decreased significantly during the ad libitum and restriction periods. The decreases in the major lipids and linoleic acid were reversed during the refeeding. The control group (n=8) which was fed high-quality concentrates ad libitum gained weight most of the spring but showed similar although slower decreases in the major serum lipids and PUFAs than the lichen group. Our results indicate that feeding reindeer on lichen during winter leads to the retardation of growth and reductions in major serum lipids and their principal C18-PUFA proportions. The decreased proportions of the principal PUFAs most probably reflect their low dietary intake but may have been modified also by seasonal factors. PMID- 10712573 TI - No apparent effect of nitric oxide on the local matching of pulmonary perfusion and ventilation in awake sheep. AB - The respiratory tissue in the lung receives nitric oxide (NO) from two sources; NO produced in upper airways, and NO produced in lung parenchyma. It has been hypothesized that optimal local matching of ventilation and perfusion (which is necessary for effective gas exchange) is ensured because well-ventilated lung tissue has a higher concentration of NO and thereby higher blood flow owing to the vasodilatory effect of NO. To test this hypothesis, we simultaneously measured the distributions of local (regions of approximately 1.5 cm3) blood flow (radioactive microspheres) and local ventilation (fluorescent aerosol) in five tracheostomized, awake and standing sheep. Tracers for perfusion and ventilation were administered (1) at baseline, (2) during endogenous NO production blockage (L-NAME 25 mg kg-1) and administration of NO free air, and (3) when the sheep received exogenous NO ( approximately 30 p.p.m.), but having its endogenous NO production blocked. The intrapulmonary distribution of ventilation was similar in all three situations. Within horizontal levels of the lung, distribution of perfusion was not affected by variable access to NO, but along the gravitational axis perfusion was more evenly distributed when the sheep had no access to NO. Exogenous NO tended to restore the baseline vertical profile. These changes in vertical distribution of perfusion can be explained by the effect of variable NO concentrations on pulmonary arterial pressure and cardiac output. Variable access to NO had no effect on arterial blood gases. We conclude that NO is important for the vertical distribution of pulmonary perfusion, but has no apparent effect on the local matching of ventilation and perfusion within horizontal layers of the lung. PMID- 10712572 TI - The effect of AVP-V2 receptor stimulation on local GFR in the rat kidney. AB - The effect of AVP-V2 receptor agonist desmopressin, dDAVP, its non-peptide antagonist OPC-31260 and vehicle infusion on glomerular filtration rate (GFR) in the outer, middle and inner cortex was studied in both hydropenic and water diuretic Inactin anaesthetized female Sprague-Dawley rats using the aprotinin method. Two subsequent GFR measurements were carried out in the same kidney by injection of 125I- and 131I-labelled aprotinin before and after i.v. infusion of dDAVP, OPC-31260 or the vehicle. Acute infusion of dDAVP in hydropenic rats increased total GFR by 14% relative to vehicle infusion, whereas in water diuretic rats it had no effect relative to vehicle. No significant changes in arterial pressure (Pa) or renal blood flow (RBF) were recorded. Infusion of OPC 31260 reduced total GFR by 11% compared with vehicle. These results are consistent with the findings that a presensitization of the vasculature by high plasma levels of AVP is necessary for the renal vascular effects mediated by the V2 or V2-like receptors to occur. The ratio between inner and outer cortex GFR remained unchanged from control to experimental condition as follows: dDAVP infusion in hydropenic rats, 0.504 vs. 0.494 in control; vehicle infusion in hydropenic rats, 0. 393 vs. 0.392; OPC-31260 infusion in hydropenic rats, 0.517 vs. 0. 523; dDAVP in water diuretic rats, 0.547 vs. 0.543; vehicle in water diuretic rats, 0.413 vs. 0.417. Thus no significant difference in the GFR response was observed between superficial and deep cortical layers of the rat kidney. PMID- 10712574 TI - Effects of peritoneal hyaluronidase treatment on transperitoneal solute and fluid transport in the rat. AB - The importance of the interstitium and its major ground substance component, hyaluronan (HA), for solute and fluid transport across the peritoneal membrane has been debated during the last few years. We therefore partly removed HA from the peritoneal membrane using enzymatic digestion with hyaluronidase for 2 h, after which the transport properties of the peritoneal membrane were studied in peritoneal dialysis dwells. A dialysis fluid containing 3.86% glucose was used. As a marker of macromolecular transport, the total peritoneal clearance of radiolabelled albumin out of the peritoneal cavity and its clearance to plasma were measured, as well as the albumin clearance from plasma to dialysate. Transport of small solutes between plasma and dialysate was measured by assessing the mass transfer area coefficient of 51Cr-EDTA and glucose. Hyaluronidase preincubation yielded a 78% reduction of HA in the superficial layer of the peritoneal membrane, without alterations in the transport of either small or large solutes compared with the situation in preincubated controls. The only changes observed were between rats incubated with either hyaluronidase or vehicle alone compared to non-incubated controls. In conclusion, despite a large reduction of the HA content of the tissues surrounding the peritoneal cavity, hyaluronidase incubation did not produce any significant changes in solute and fluid transport across the peritoneal membrane. Our data indicate that peritoneal membrane HA in physiological concentrations plays a rather subordinate role in the overall transport of small solutes and water across the capillary interstitial peritoneal barrier. PMID- 10712575 TI - Effects of acute hypobaric hypoxia on regional cerebral blood flow distribution: a single photon emission computed tomography study in humans. AB - Single Photon Emission Computed Tomography (SPECT) and radiopharmaceutical stabilizing agents allowed us to investigate regional cerebral blood flow (CBF) distribution in six resting healthy subjects during acute laboratory hypobaric hypoxic conditions. In the hypobaric experiment stabilized 99mTc-D, L-hexamethyl propylene amine oxime was injected 40 min after reaching hypoxic conditions corresponding to an altitude of 5500 m above sea level. Arterial blood sample was taken after five additional minutes. Mean arterial oxygen pressure and haemoglobin saturation were 28 mmHg and 56%, respectively. The control experiment was performed similarly, apart from barometric pressure and blood gas analysis. We analysed CBF distribution in 12 regions of functional interest bilaterally in frontal, parietal, temporal, occipital cortex, in the hippocampus, in the basal ganglia and other central structures of brain. No overall effect of hypoxia on normalized regional CBF distribution in the considered regions was found. Motor cortex (Brodmann 4) and basal ganglia were the only regions in which hypobaric hypoxia significantly increased relative distribution of the radiopharmaceutical [F(1,5)=18.30; P < 0.008 and F(1,5)=10.85; P < 0.022, respectively]. Despite severe hypoxia, we did not observe any major regional CBF redistribution. We found a small relative increase in blood flow to the motor cortex and the basal ganglia, at rest after 40 min of hypobaric hypoxia, suggesting a preferential compensatory mechanism of these functional regions of brain. PMID- 10712576 TI - Impact of oestradiol and progesterone on the glycosaminoglycans and their depolymerizing enzymes of the rat mammary gland. AB - The influence of oestradiol and progesterone either singly or in combination with each other on the levels of hyaluronic acid, heparan sulphate, chondroitin sulphate, and on the activity of hyaluronidase and chondroitinase were investigated in the mammary gland of ovary-intact and in ovariectomized rats, administered oestradiol and/or progesterone. Administration of oestradiol to ovary-intact rats elevated the levels of hyaluronic acid and decreased the levels of heparan sulphate while progesterone, when administered alone, could elevate only chondroitin sulphate when compared with controls. The steroids when administered in combination, however, increased the levels of all glycosaminoglycans studied. Ovariectomized animals showed a decrease in heparan sulphate alone as compared with controls while administration of oestradiol to these rats elevated the levels of both heparan sulphate and chondroitin sulphate as compared with ovariectomized rats. Also the administration of progesterone either singly or in combination increased the levels of heparan sulphate and also decreased the levels of hyaluronic acid with no impact on the levels of chondoritin sulphate. In ovary-intact animals administration of oestradiol alone had no effect on hyaluronidase activity. Progesterone either singly or in combination with oestradiol reduced the activity of hyaluronidase, whereas it had no influence on the activity of chondroitinase. The activities of both the enzymes were decreased in ovariectomized animals and administration of oestradiol and/or progesterone to the above groups resulted in an increase. This study demonstrates that oestradiol anzd progesterone play an important role in modulating glycosaminoglycans and their depolymerizing enzymes, thereby influencing the activities of the mammary epithelium. PMID- 10712577 TI - Na+-Ca2+ exchange current from rabbit isolated atrioventricular nodal and ventricular myocytes compared using action potential and ramp waveforms. AB - We measured and compared Na-Ca exchanger current (INa-Ca) from rabbit isolated ventricular and atrioventricular (AV) nodal myocytes, using action potential (AP) and ramp voltage commands. Whole cell patch-clamp recordings were made at 35-37 degrees C; INa-Ca was measured as 5 mM nickel (Ni)- sensitive current with major interfering voltage and calcium-activated currents blocked. In ventricular cells a 2-s descending ramp elicited INa-Ca showing outward rectification and a reversal potential (Erev) of -13.1 +/- 1. 2 mV (n = 12; mean +/- SEM). With a ventricular AP as the voltage command, the profile of INa-Ca followed the applied waveform closely. The current-voltage relation during AP repolarization was almost linear and showed an Erev of -38.3 +/- 5.3 mV (n = 6). As INa-Ca depended on the applied voltage waveform, comparisons between the two cell types utilized the same command waveform (a series of AV nodal APs). In ventricular myocytes this elicited INa-Ca that reversed near -38 mV and was inwardly directed during the pacemaker potential. This command was also applied to AV node cells; mean INa Ca density at all voltages encompassed by the AP (-70 to +30 mV) did not differ significantly from that in ventricular myocytes (P > 0.05, ANOVA). This finding was confirmed using brief (250 ms) voltage ramp protocols (P > 0.1 ANOVA). These data represent the first direct measurements of AV nodal INa-Ca and suggest that the exchanger may be functionally expressed to similar levels in the two cell types. They may also suggest a possible role for INa-Ca during the pacemaker potential in AV node as inward INa-Ca was observed over the pacemaker potential range even with bulk internal Ca buffered to a low level. PMID- 10712578 TI - Effect of hyperoxia on aerobic and anaerobic performances and muscle metabolism during maximal cycling exercise. AB - The hyperoxia-improved tolerance to maximal aerobic performance was studied in relation to exercising muscle metabolic state. Five students were submitted to four different tests on a cycle ergometer, each being conducted under normoxia and hyperoxia (60% FiO2) on separate days: Test 1, a progressive exercise until exhaustion to determine the maximal work load (Wmax) which was unchanged by hyperoxia; Test 2, an exercise at Wmax (287 +/- 12 W) until exhaustion to determine the performance time (texh) which was elevated by 38% under hyperoxia but exhaustion occurred at the same arterial proton and lactate concentrations; Test 3 (S-Exercise test) consisted of cycling at Wmax for 90% normoxic-texh (4.8 +/- 0.5 min under both O2 conditions) then followed by a 10-s sprint bout during which the total work output (Wtot) was determined; Wtot was elevated by 15% when exercising under hyperoxia; Test 4 (M-Exercise test) consisted also of cycling at Wmax for 4.8 +/- 0.5 min with blood and muscle samples taken at rest and at the end of the exercise to compare the level of different metabolites. During hyperoxic M-Exercise test, glycogen was twice more depleted whereas glucose-6 phosphate and lactate were less accumulated when compared with normoxia. No significant differences were observed for pyruvate, phosphocreatine and muscle/blood lactate ratio between the two conditions. Conversely to normoxia, levels of ATP, ADP and total NADH were maintained at their resting level under 60% FiO2. These data lead us to suppose a higher oxidation rate for pyruvate and NADH in mitochondria, thereby lowering the metabolic acidosis and allowing a better functioning of the glycolytic and contractile processes to delay the time to exhaustion. PMID- 10712579 TI - Immunomodulation by 8-week voluntary exercise in mice. AB - This study was designed to evaluate the effects of voluntary exercise on macrophage and lymphocyte functions in mice. Male A/He inbred mice aged 19 weeks were divided into two groups: a group given voluntary exercise and a control group (n = 10 in each group). Exercise consisted of spontaneous running in wheels for 8 weeks (3 days week-1). Glucose consumption of peritoneal macrophages in the exercise group during incubation up to 72 h was significantly higher than that in the control group (70 and 13%, respectively). Also, activities of acid phosphatase (APH) (10.75 +/- 0.37 IU), beta-glucuronidase (GLU) (1.55 +/- 0.07 IU) and lactate dehydrogenase (LDH) (43.3 +/- 0.7 IU) in the peritoneal macrophages in the exercise group was significantly increased (P < 0.01). Compared with the control group, the exercise group had a significant increase of about twofold in macrophage production of nitric oxide (NO2-) stimulated by lipopolysaccharide (LPS) (11.1 +/- 0.1 vs. 5.9 +/- 0.1 microM mL-1 in exercise and control groups, respectively; P < 0.01). Stimulation indices both by concanavalin A (Con A) and phytohaemagglutinin were also significantly higher in the exercise group (P < 0.01). A significant increase in the splenocyte production of interleukin-2 (IL-2) stimulated by Con A was noticed in the exercise group (354.1 +/- 28.8 vs. 218.9 +/- 23.5 pg mL-1 in exercise and control groups, respectively; P < 0.01). These findings suggest that voluntary exercise enhances not only macrophage function but also lymphocyte responsiveness in mice. In the studies of voluntary exercise, evaluation of NO2- production, as an indicator of macrophage function, is recommended. PMID- 10712580 TI - Corticotropic and serotonergic responses to acute stress with/without prior exercise training in different rat strains. AB - The ability to cope with exercise training depends both on environmental and genetic background; however, whether the genetic status may affect (i) the hormonal status of trained subjects and, (ii) its responses to a heterotypic stressor is unknown. Herein, we have used Spontaneously Hypertensive Rats (SHR) and Lewis rats, that differ with regard to their psychoneuroendocrine profiles, to study the influences of an 8-week training programme and/or a 1-h immobilization stress on plasma adrenocorticotropin (ACTH) and corticosterone levels. In addition, brain serotonin metabolism was also measured as an index of neurochemical reactivity to stress. The amplitude of immobilization-elicited increases in ACTH levels which differed with the rat strain (Lewis > SHR), was amplified by prior training; besides, training decreased the strain difference in basal corticosterone (SHR > Lewis) and affected corticosterone response to immobilization in a strain-dependent manner. Thus, immobilization, which increased corticosterone levels in sedentary Lewis but not in SHRs, did not reveal interstrain differences in trained rats. Taken with the observation of a stimulatory effect of training on adrenal weights in SHRs, but not in Lewis, it is concluded that the effects of training on the corticotropic axis depend on the genetic profile of the individual. Lastly, training amplified the response of midbrain (but not striatum or hippocampus) serotonin metabolism to immobilization in a strain-independent manner although the levels of serotonin precursor, namely tryptophan, varied with training and immobilization in a strain-dependent manner. This study shows that some neuroendocrine and neurochemical effects of training undergo interindividual variability. PMID- 10712581 TI - Hypothermia affects the phagocytic activity of rat peritoneal macrophages. AB - To examine the effect of hypothermia on the phagocytic capacity of rat peritoneal macrophages for latex particles, male Wistar rats were exposed to 4 degrees C for 8 and 72 h. While the shorter exposure to cold did not affect body temperature and macrophage function, animals exposed to 4 degrees C for 72 h showed a mean decrease of their body temperature by 1.5 degrees C. The superoxide anion production was significantly increased whereas the number of phagocytic cells decreased. In addition, the mean number of latex particles engulfed by each individual cell was lower than that of controls. Peripheral blood mononuclear cells (PBMC) of these animals showed lower mitogen response to phytohaemagglutinin (PHA), while that for concanavalin A (Con-A) remained unchanged. Peritoneal macrophages exposed in vitro to 24 degrees C for 60 min showed a decreased phagocytic capacity in comparison with macrophages kept at 37 degrees C, an observation suggesting the development of an indigenous cell defect for phagocytosis at lower temperatures. On the other hand, the effect of additional humoral factor(s) on macrophage activity, such as an increase in serum level of catecholamines and corticosterone, cannot be excluded. The results of the study may contribute to understanding the predisposition to infections during exposure to cold. PMID- 10712582 TI - Effect of nervous excitation on acid secretion in horses. AB - Nervous excitation was induced by various means in horses provided with a gastric cannula. Insulin hypoglycaemia profoundly inhibited the basal acid output and volume secreted from the stomach. No clear effect on acid secretion was noted after administration of bethanechol, as the acid output was covered by the copious secretion of saliva. Atropine almost abolished the basal acid output. Sensoric stimulation by teasing caused a slight but not significant increase in the total acid output. These data suggest that cholinergic excitation might play a role in the stimulation of both volume and acid secretion in the horse. The inhibitory effect seen on these two parameters after insulin hypoglycaemia may hypothetically be ascribed to inhibitory impulses carried in peptide neurones of the vagal nerves or to inhibitory impulses in adrenergic nerves acting directly or indirectly on the parietal cells. PMID- 10712583 TI - New lessons in the regulation of glucose metabolism taught by the glucose 6 phosphatase system. AB - The operation of glucose 6-phosphatase (EC 3.1.3.9) (Glc6Pase) stems from the interaction of at least two highly hydrophobic proteins embedded in the ER membrane, a heavily glycosylated catalytic subunit of m 36 kDa (P36) and a 46-kDa putative glucose 6-phosphate (Glc6P) translocase (P46). Topology studies of P36 and P46 predict, respectively, nine and ten transmembrane domains with the N terminal end of P36 oriented towards the lumen of the ER and both termini of P46 oriented towards the cytoplasm. P36 gene expression is increased by glucose, fructose 2,6-bisphosphate (Fru-2,6-P2) and free fatty acids, as well as by glucocorticoids and cyclic AMP; the latter are counteracted by insulin. P46 gene expression is affected by glucose, insulin and cyclic AMP in a manner similar to P36. Accordingly, several response elements for glucocorticoids, cyclic AMP and insulin regulated by hepatocyte nuclear factors were found in the Glc6Pase promoter. Mutations in P36 and P46 lead to glycogen storage disease (GSD) type-1a and type-1 non a (formerly 1b and 1c), respectively. Adenovirus-mediated overexpression of P36 in hepatocytes and in vivo impairs glycogen metabolism and glycolysis and increases glucose production; P36 overexpression in INS-1 cells results in decreased glycolysis and glucose-induced insulin secretion. The nature of the interaction between P36 and P46 in controling Glc6Pase activity remains to be defined. The latter might also have functions other than Glc6P transport that are related to Glc6P metabolism. PMID- 10712584 TI - New functions of a long-known molecule. Emerging roles of NAD in cellular signaling. AB - Over the past decades, the pyridine nucleotides have been established as important molecules in signaling pathways, besides their well known function in energy transduction. Similarly to another molecule carrying such dual functions, ATP, NAD(P)+ may serve as substrate for covalent protein modification or as precursor of biologically active compounds. Protein modification is catalyzed by ADP-ribosyl transferases that attach the ADP-ribose moiety of NAD+ to specific amino-acid residues of the acceptor proteins. For a number of ADP ribosylation reactions the specific transferases and their target proteins have been identified. As a result of the modification, the biological activity of the acceptor proteins may be severely changed. The cell nucleus contains enzymes catalyzing the transfer of ADP-ribose polymers (polyADP-ribose) onto the acceptor proteins. The best known enzyme of this type is poly(ADP-ribose) polymerase 1 (PARP1), which has been implicated in the regulation of several important processes including DNA repair, transcription, apoptosis, neoplastic transformation and others. The second group of reactions leads to the synthesis of an unusual cyclic nucleotide, cyclic ADP-ribose (cADPR). Moreover, the enzymes catalyzing this reaction may also replace the nicotinamide of NADP+ by nicotinic acid resulting in the synthesis of nicotinic acid adenine dinucleotide phosphate (NAADP+). Both cADPR and NAADP+ have been reported to be potent intracellular calcium-mobilizing agents. In concert with inositol 1,4,5-trisphosphate, they participate in cytosolic calcium regulation by releasing calcium from intracellular stores. PMID- 10712585 TI - Quantitative analysis of gene expression with an improved green fluorescent protein. p6. AB - The fast and easy in vivo detection predestines the green fluorescent protein (GFP) for its use as a reporter to quantify promoter activities. We have increased the sensitivity of GFP detection 320-fold compared to the wild-type by constructing gfp+, which contains mutations improving the folding efficiency and the fluorescence yield of GFP+. Twelve expression levels were measured using fusions of the gfp+ and lacZ genes with the tetA promoter in Escherichia coli. The agreement of GFP+ fluorescence with beta-galactosidase activities was excellent, demonstrating that the gfp+ gene can be used to accurately quantify gene expression in vivo. However, expression of the gfp+ gene from the stronger hsp60 promoter revealed that high cellular concentrations of GFP+ caused an inner filter effect reducing the fluorescence by 50%, thus underestimating promoter activity. This effect is probably due to the higher absorbance of cells containing GFP+. Thus promoters with activities differing by about two orders of magnitude can be correctly quantified using the gfp+ gene. Possibilities of using GFP variants beyond this range are discussed. PMID- 10712586 TI - The proteome of Mycoplasma genitalium. Chaps-soluble component. AB - Mycoplasma genitalium is the smallest member of the class Mollicutes, with a genome size of 580 kb. It has the potential to express 480 gene products, and is therefore considered to be an excellent model to assess: (a) the minimum metabolism required by a free living cell; and (b) proteomic technologies and the information obtained by proteome analysis. Here, we report on the most complete proteome observed at 73% (expected proteome), and analysed at 33% (reported proteome). The use of four overlapping pH windows in conjunction with SDS/PAGE has allowed 427 distinct proteins to be resolved in association with the exponential growth of M. genitalium. Proof of expression for 201 proteins of sufficient abundance on silver stained two-dimensional gels was obtained using peptide mass fingerprinting (PMF) of which 158 were identified. The potential for gene product modification in even the simplest known self-replicating organism was quantified at a ratio of 1.22 : 1, more proteins than genes. A reduction in protein expression of 42% was observed for post-exponentially-grown cells. DnaK, GroEL, DNA gyrase, and a cytadherence accessory protein were significantly elevated, while some ribosomal proteins were reduced in relative abundance. The strengths and weaknesses of techniques employed were assessed with respect to the observed and predicted proteome derived from DNA sequence information. Proteomics was shown to provide a perspective into the biochemical and metabolic activities of this organism, beyond that obtainable by sequencing alone. PMID- 10712587 TI - Negative feedback and ultrasensitivity can bring about oscillations in the mitogen-activated protein kinase cascades. AB - Functional organization of signal transduction into protein phosphorylation cascades, such as the mitogen-activated protein kinase (MAPK) cascades, greatly enhances the sensitivity of cellular targets to external stimuli. The sensitivity increases multiplicatively with the number of cascade levels, so that a tiny change in a stimulus results in a large change in the response, the phenomenon referred to as ultrasensitivity. In a variety of cell types, the MAPK cascades are imbedded in long feedback loops, positive or negative, depending on whether the terminal kinase stimulates or inhibits the activation of the initial level. Here we demonstrate that a negative feedback loop combined with intrinsic ultrasensitivity of the MAPK cascade can bring about sustained oscillations in MAPK phosphorylation. Based on recent kinetic data on the MAPK cascades, we predict that the period of oscillations can range from minutes to hours. The phosphorylation level can vary between the base level and almost 100% of the total protein. The oscillations of the phosphorylation cascades and slow protein diffusion in the cytoplasm can lead to intracellular waves of phospho-proteins. PMID- 10712588 TI - A protein kinase C-independent pathway leading to c-Jun-dependent expression of 100-kDa Ras GTPase-activating protein in JEG-3 human choriocarcinoma cells. AB - Although the 100-kDa Ras GTPase-activating protein (p100 RasGAP) has been reported to exist specifically in human placental trophoblasts, the molecular mechanisms responsible for regulating its expression remain unclear. In this study we used okadaic acid, an inhibitor of serine/threonine phosphatase 1 and 2 A, as a probe to explore the signaling pathway regulating the expression of p100 RasGAP in JEG-3 human placental choriocarcinoma cells. Treatment of JEG-3 cells with okadaic acid provoked dose- and time-dependent stimulation of p100 RasGAP expression without marked modification of expression of p120 RasGAP, another isoform of RasGAP. Co-treatment of cells with okadaic acid and the protein kinase C activator, phorbol 12-myristate 13-acetate, exerted an additive effect on p100 RasGAP induction. Moreover, the response of the p100 RasGAP de novo synthesis to okadaic acid was not affected by the selective inhibitor of protein kinase C, GF 109203X. Thus this study identified a novel signaling pathway regulating p100 RasGAP expression, which is independent of protein kinase C. In addition, okadaic acid treatment resulted in the activation of ERK2 (p42 MAP kinase) and the induction of both c-Jun and c-Fos proteins without activating JNK (c-Jun NH2 terminal kinase). Significantly, blockade of c-Jun expression with antisense c jun oligonucleotides suppressed p100 RasGAP expression. Taken together, it is concluded that okadaic acid induces the expression of p100 RasGAP protein in JEG 3 cells preceded by activation of ERK and AP-1 cascade, and that this okadaic acid-induced p100 RasGAP expression is independent of protein kinase C-mediated pathway but requires c-Jun/AP-1 function. PMID- 10712589 TI - A diminished activation capacity of the interferon-inducible protein kinase PKR in human T lymphocytes. AB - The double-stranded (ds) RNA activated protein kinase PKR is an interferon (IFN) inducible serine/threonine protein that regulates protein synthesis through the phosphorylation of the alpha subunit of translation initiation factor 2 (eIF 2alpha). PKR activation in cells is induced by virus infection or treatment with dsRNA and is modulated by a number of viral and cellular factors. To better understand the mechanisms of PKR action we have analyzed and compared the mode of PKR activation in a number of cell lines of different histological origin. Here we show that PKR activation and phosphorylation of eIF-2alpha are both diminished in various virus-transformed and nontransformed human T cells. Priming of T cells with IFN does not restore PKR activation. In vitro kinase assays show that the diminished PKR activation in T cells correlates with the presence of a 60-kDa (p60) phosphoprotein coimmunoprecipitated with PKR. P60 is absent from PKR immunoprecipitates from non T cells. Incubation of active PKR with T cell extracts results in inhibition of PKR autophosphorylation, which is proportional to the amount of phosphorylated p60 in the kinase reactions. Treatment of T cells with proteasome inhibitors or incubation of PKR immunoprecipitates with phosphatase inhibitors does not restore PKR activation. However, phosphorylation of p60 is enhanced upon treatment with the phosphatase inhibitor microcystin. These data show that the impaired activation capacity of PKR in human T cells is exerted at the post-translational levels in a manner that is independent of cell transformation or virus infection. PMID- 10712590 TI - Effects of singlet oxygen on membrane sterols in the yeast Saccharomyces cerevisiae. AB - Photodynamic treatment of the yeast Saccharomyces cerevisiae with the singlet oxygen sensitizer toluidine blue and visible light leads to rapid oxidation of ergosterol and accumulation of oxidized ergosterol derivatives in the plasma membrane. The predominant oxidation product accumulated was identified as 5alpha, 6alpha-epoxy-(22E)-ergosta-8,22-dien-3beta,7a lpha-diol (8-DED). 9(11) dehydroergosterol (DHE) was identified as a minor oxidation product. In heat inactivated cells ergosterol is photooxidized to ergosterol epidioxide (EEP) and DHE. Disrupted cell preparations of S. cerevisiae convert EEP to 8-DED, and this activity is abolished in a boiled control indicating the presence of a membrane associated enzyme with an EEP isomerase activity. Yeast selectively mobilizes ergosterol from the intracellular sterol ester pool to replenish the level of free ergosterol in the plasma membrane during singlet oxygen oxidation. The following reaction pathway is proposed: singlet oxygen-mediated oxidation of ergosterol leads to mainly the formation of EEP, which is enzymatically rearranged to 8-DED. Ergosterol 7-hydroperoxide, a known minor product of the reaction of singlet oxygen with ergosterol, is formed at a much lower rate and decomposes to give DHE. Changes of physical properties of the plasma membrane are induced by depletion of ergosterol and accumulation of polar derivatives. Subsequent permeation of photosensitizer through the plasma membrane into the cell leads to events including impairment of mitochondrial function and cell inactivation. PMID- 10712592 TI - Favourable side-chain orientation of cleavage site dibasic residues of prohormone in proteolytic processing by prohormone convertase 1/3. AB - Previous studies using selectively modified pro-ocytocin/neurophysin substrate analogues and the purified metalloprotease, pro-ocytocin/neurophysin convertase (magnolysin; EC 3.4 24.62), have shown that dibasic cleavage site processing is associated with a prohormone sequence organized in a beta-turn structure. We have used various peptide analogues of the pro-ocytocin-neurophysin processing domain, and recombinant prohormone convertase 1/3, to test the validity of this property towards this member of the family of prohormone convertases (PCs). The enzymatic cleavage analysis and kinetics showed that: (a) with methyl amide (N-Met) modification, a secondary structure beta-turn breaker, the enzyme substrate interaction was abolished; (b) cleavage was favoured when the dibasic substrate side-chains were oriented in opposite directions; (c) the amino acid present at the P'1 position is important in the enzyme-substrate interaction; (d) the flexibility of the peptide substrate is necessary for the interaction; (e) Addition of dimethylsulfoxide to the cleavage assay favoured the cleavage of the pro-ocytocin/neurophysin large substrate over that of the smaller one pGlu-Arg Thr-Lys-Arg-methyl coumarin amide. These data allowed us to conclude that proteolytic processing of pro-ocytocin-related peptide substrates by PC1/3 as well as by the metalloenzyme, magnolysin, involves selective recognition of precise cleavage site local secondary structure by the processing enzyme. It is hypothesized that this may represent a general property of peptide precursor proteolytic processing systems. PMID- 10712591 TI - Molecular cloning of the cDNA encoding laccase from Pycnoporus cinnabarinus I-937 and expression in Pichia pastoris. AB - Laccases are multicopper-containing enzymes which catalyse the oxidation of phenolic and nonphenolic compounds with the concomitant reduction of molecular oxygen. In this study, a full-length cDNA coding for laccase (lac1) from Pycnoporus cinnabarinus I-937 was isolated and characterized. The corresponding open reading frame is 1557 nucleotides long and encodes a protein of 518 amino acids. The cDNA encodes a precursor protein containing a 21 amino-acid signal sequence corresponding to a putative signal peptide. The deduced amino-acid sequence of the encoded protein was similar to that of other laccase proteins, with the residues involved in copper coordination sharing the greatest extent of similarity. The cDNA encoding for laccase was placed under the control of the alcohol oxidase (Aox 1) promoter and expressed in the methylotropic yeast Pichia pastoris. The laccase leader peptide, as well as the Saccharomyces cerevisiae alpha-factor signal peptide, efficiently directed the secretion into the culture medium of laccase in an active form. Moreover, the laccase activity was directly detected in plates. The identity of the recombinant product was further confirmed by protein immunoblotting. The expected molecular mass of the mature protein is 81 kDa. However, the apparent molecular mass of the recombinant protein is 110 k Da, thus suggesting that the protein expressed in P. pastoris may be hyperglycosylated. PMID- 10712593 TI - Roles of his205, his296, his303 and Asp259 in catalysis by NAD+-specific D lactate dehydrogenase. AB - The role of three histidine residues (His205, His296 and His303) and Asp259, important for the catalysis of NAD+-specific D-lactate dehydrogenase, was investigated using site-directed mutagenesis. None of these residues is presumed to be involved in coenzyme binding because Km for NADH remained essentially unchanged for all the mutant enzymes. Replacement of His205 with lysine resulted in a 125-fold reduction in kcat and a slight lowering of the Km value for pyruvate. D259N mutant showed a 56-fold reduction in kcat and a fivefold lowering of Km. The enzymatic activity profile shifted towards acidic pH by approximately 2 units. The H303K mutation produced no significant change in kcat values, although Km for pyruvate increased fourfold. Substitution of His296 with lysine produced no significant change in kcat values or in Km for substrate. The results obtained suggest that His205 and Asp259 play an important role in catalysis, whereas His303 does not. This corroborates structural information available for some members of the D-specific dehydrogenases family. The catalytic His296, proposed from structural studies to be the active site acid/base catalyst, is not invariant. Its function can be accomplished by lysine and this has significant implications for the enzymatic mechanism. PMID- 10712594 TI - Structural characterization of the outer core and the O-chain linkage region of lipopolysaccharide from Pseudomonas aeruginosa serotype O5. AB - The point of attachment of the O-chain in the outer core region of Pseudomonas aeruginosa serotype O5 lipopolysaccharide (LPS) was determined following a detailed analysis of the extended core oligosaccharide, containing one trisaccharide O-chain repeating unit, present in both the wild-type strain PAO1 and O-chain deficient mutant strains AK1401 and PAO-rfc. The structure of the extended core oligosaccharide was determined by various mass spectrometric methods as well as one-dimensional and two-dimensional NMR spectroscopy. Furthermore, the one-dimensional analogues of NOESY and TOCSY experiments were applied to confirm the structure of the outer core region in the O-chain polysaccharide. In both the extended core oligosaccharide and the core of the smooth LPS, a loss of one of the beta-glucosyl residues and the translocation of the alpha-rhamnosyl residue, followed by the attachment of the first O-chain repeating unit was observed. This process is complicated and could involve two distinct rhamnosyltransferases, one with alpha-1, 6-linkage specificity and another with alpha-1,3-linkage specificity. It is also plausible that an alpha 1,3 rhamnosyltransferase facilitates the addition of the 'new' alpha-rhamnosyl residue that will act as a receptor for the attachment of the single O-antigen repeating unit in the LPS of the semi-rough mutant. The 2-amino-2-deoxy-fucosyl residue of the first O-chain repeating unit directly attached to the core was found to have a beta-anomeric configuration instead of an alpha configuration, characteristic for this residue as a component of the O-chain polysaccharide. The results of this study provide the first example of the mechanistic implications of the structure of the outer core region in a fully assembled O-chain containing LPS, differing from the O-chain deficient rough LPS. PMID- 10712595 TI - Salmon antithrombin has only three carbohydrate side chains, and shows functional similarities to human beta-antithrombin. AB - Antithrombin, a major coagulation inhibitor in mammals, has for the first time been cDNA cloned from a fish species. The predicted mature liver antithrombin of Atlantic salmon (Salmo salar) consists of 430 amino acids and shows about 67% sequence identity to mammalian and chicken antithrombins. Due to a single nucleotide replacement, Asn135 of the antithrombin in higher vertebrates is substituted by Asp in the salmon homolog. Hence, in contrast to the vertebrate antithrombins known so far, salmon antithrombin lacks the potential glycosylation site located close to the heparin binding site. The existence of only three N linked side chains is evidenced by the sequential removal of three carbohydrate chains from salmon antithrombin during timed-digestion with N-glycosidase F. The high heparin binding affinity of the salmon inhibitor, Kd of 2.2 and 48 nM at I = 0.15 and 0.3, respectively, is very similar to that of the minor human isoform beta-antithrombin, which is not glycosylated at Asn135. Furthermore, the invariant third-position Ser137 at this glycosylation site of mammalian and chicken antithrombins is substituted by Thr in the salmon, a replacement that has been shown to induce full glycosylation in human antithrombin. Thus a rapidly reacting pool of antithrombin may have evolved in two different ways: absence of a glycosylation site in lower vertebrates vs. incomplete glycosylation of a part of the circulating antithrombin in higher vertebrates. Salmon antithrombin appears to have three complex oligosaccharide side chains containing sialic acid terminally linked alpha(2-3) to galactose, while trace amounts of Galbeta(1 4)GlcNAc suggest microheterogeneity due to partial loss of sialic acid. PMID- 10712596 TI - Upregulation of cytosolic chaperonin CCT subunits during recovery from chemical stress that causes accumulation of unfolded proteins. AB - The chaperonin containing TCP-1 (CCT) is a molecular chaperone consisting of eight subunit species and assists in the folding of actin, tubulin and some other cytosolic proteins. We examined the stress response of CCT subunit proteins in mammalian cultured cells using chemical stressors that cause accumulation of unfolded proteins. Levels of CCT subunit proteins in HeLa cells were coordinately and transiently upregulated under continuous chemical stress with sodium arsenite. CCT subunit levels in several mammalian cell lines were also upregulated during recovery from chemical stress caused by sodium arsenite or a proline analogue, L-azetidine-2-carboxylic acid. Several unidentified proteins that were newly synthesized and associated with CCT were found to increase concomitantly with CCT subunits themselves and known substrates during recovery from the stress. These results suggest that CCT plays important roles in the recovery of cells from protein damage by assisting in the folding of proteins that are actively synthesized and/or renatured during this period. PMID- 10712597 TI - High expression of the human hepatocarcinoma-intestine-pancreas/pancreatic associated protein (HIP/PAP) gene in the mammary gland of lactating transgenic mice. Secretion into the milk and purification of the HIP/PAP lectin. AB - The human hepatocarcinoma-intestine-pancreas/pancreatic-associated protein (HIP/PAP) gene was previously identified because of its increased expression in primary liver cancers and during the acute phase of pancreatitis. In normal tissues, HIP/PAP is expressed both in endocrine and exocrine cells of the intestine and pancreas. HIP/PAP is a lactose binding C-type lectin which acts as an adhesion molecule for rat hepatocytes. The aim of the work was to study the HIP/PAP secretory pathway and to produce high levels of HIP/PAP in the milk of lactating transgenic mice. In view of its lactose C-type lectin properties, we have studied the consequences of the expression of HIP/PAP on mammary epithelial cells. In homozygous mice, production reached 11.2 mg.mL-1 of milk. High levels of soluble and pure HIP/PAP (18.6 mg) were purified from 29 mL of milk. The purified protein was sequenced and the N-terminal amino acid of the mature HIP/PAP was identified as Glu27, thus localizing the site of cleavage of the signal peptide. The HIP/PAP transgene was only expressed in the mammary gland of lactating transgenic mice. HIP/PAP was detected by immunofluorescence in the whole gland, but labelling was heterogeneous between alveolar clusters, with strongly positive sparse cells. Using immuno electron microscopy, HIP/PAP was observed in all the compartments of the secretory pathway within the mammary epithelial cells. We provide evidence that HIP/PAP is secreted through the Golgi pathway. However, the number of distended Golgi saccules was increased when compared to that found in wild-type mouse mammary cells. These modifications could be related to HIP/PAP C-type lectin specific properties. PMID- 10712598 TI - Structural studies of the O-specific polysaccharide from Plesiomonas shigelloides strain CNCTC 113/92. AB - The structure of the O-specific side chain of the lipopolysaccharide (LPS) of Plesiomonas shigelloides, strain CNCTC 113/92 has been investigated by NMR spectroscopy, matrix-assisted laser desorption/ionization time of flight mass spectrometry and sugar and methylation analysis. It was concluded that the polysaccharide is composed of a hexasaccharide repeating unit with the following structure: in which D-beta-D-Hepp is Dglycero-beta-Dmanno-heptopyranose and 6d beta-D-Hep is 6-deoxy-beta-Dmanno-heptopyranose. This structure represents a novel hexasaccharide repeating unit of bacterial O-antigen that is characteristic and unique to the Plesiomonas shigelloides strain. Using the high-resolution magic angle spinning technique, 1H-NMR spectra were also obtained for the O polysaccharide components of isolated LPS and in their original form directly on the surface of bacterial cells. PMID- 10712599 TI - Degradation of transcription factor IRF-1 by the ubiquitin-proteasome pathway. The C-terminal region governs the protein stability. AB - Interferon regulatory factor-1(IRF-1) is a transcriptional activator of interferon genes and interferon-inducible genes. It has been shown that IRF-1 functions not only as a regulator of the interferon-responsive system but also as a regulator of cell growth and apoptosis. In addition, it is known that IRF-1 is a short-lived protein, but the mechanism that regulates its stability has not yet been clarified. Here, we show that IRF-1 is degraded via the ubiquitin-proteasome pathway. IRF-1 protein degradation in HeLa and NIH3T3 cells was inhibited by treatment with proteasome-specific inhibitors. Overexpression of IRF-1 protein and ubiquitin in COS7 cells revealed specific multiubiquitination of IRF-1. Although the full-length IRF-1 was unstable, IRF-1 mutants with C-terminal truncations larger than 39 amino acids were found to be almost stable, suggesting that the 39-residue C-terminal region controls the stability of IRF-1. Further analysis of the stability of a green fluorescent protein-fusion protein containing the 39-residue C-terminal region of IRF-1 showed that this C-terminal region confers instability on green fluorescent protein, a normally stable protein, suggesting that this region functions as a protein-degradation signal. Taking the results together, it can be concluded that the 39-residue C-terminal region is necessary and sufficient to control the stability of the IRF-1 protein. PMID- 10712600 TI - Study of the subunit interactions in myosin phosphatase by surface plasmon resonance. AB - The interactions of the catalytic subunit of type 1 protein phosphatase (PP1c) and the N-terminal half (residues 1-511) of myosin phosphatase target subunit 1 (MYPT1) were studied. Biotinylated MYPT1 derivatives were immobilized on streptavidin-biosensor chips, and binding parameters with PP1c were determined by surface plasmon resonance (SPR). The affinity of binding of PP1c was: MYPT11-296 > MYPT11-38 > MYPT123-38. No binding was detected with MYPT11-34, suggesting a critical role for residues 35-38, i.e. the PP1c binding motif. Binding of residues 1-22 was inferred from: a higher affinity binding to PP1c for MYPT11-38 compared to MYPT123-38, as deduced from SPR kinetic data and ligand competition assays; and an activation of the myosin light chain phosphatase activity of PP1c by MYPT11-38, but not by MYPT123-38. Residues 40-296 (ankyrin repeats) in MYPT11 296 inhibited the phosphorylase phosphatase activity of PP1c (IC50 = 0.2 nM), whereas MYPT11-38, MYPT123-38 or MYPT11-34 were without effect. MYPT140-511, which alone did not bind to PP1c, showed facilitated binding to the complexes of PP1c-MYPT11-38 and PP1c-MYPT123-38. The inhibitory effect of MYPT140-511 on the phosphorylase phosphatase activity of PP1c also was increased in the presence of MYPT11-38. The binding of MYPT1304-511 to complexes of PP1c and MYPT11-38, or MYPT11-296, was detected by SPR. These results suggest that within the N-terminal half of MYPT1 there are at least four binding sites for PP1c. The essential interaction is with the PP1c-binding motif and the other interactions are facilitated in an ordered and cooperative manner. PMID- 10712601 TI - Molecular characterization of two genes from Streptomyces tendae Tu901 required for the formation of the 4-formyl-4-imidazolin-2-one-containing nucleoside moiety of the peptidyl nucleoside antibiotic nikkomycin. AB - The genes nikQ and nikR were identified by sequencing DNA of the nikkomycin biosynthetic gene cluster from Streptomyces tendae Tu901/8c. The nikQ gene encodes a P450 cytochrome, and the predicted NikR gene product shows 48-56% sequence identity with uracil phosphoribosyltransferases from eukaryotic organisms. The nikQ and nikR genes were inactivated separately by insertion of a kanamycin-resistance cassette. Inactivation of the nikQ gene abolished synthesis of nikkomycins containing 4-formyl-4-imidazolin-2-one as the base (nikkomycins X and I), whereas production of nikkomycins containing uracil (nikkomycins Z and J) was not affected. Nikkomycin X and I production could be restored by feeding 4 formyl-4-imidazolin-2-one to the nikQ mutants, indicating that NikQ is responsible for its formation from L-histidine. Disruption of the nikR gene resulted in formation of decreased amounts of nikkomycins X and I, whereas nikkomycins Z and J were synthesized at wild-type levels. A fluorouracil resistant nikR mutant lacking uracil phosphoribosyltransferase (UPRTase) activity did not synthesize nikkomycins X and I and accumulated 4-formyl-4-imidazolin-2 one in its culture filtrate, whereas formation of nikkomycins Z and J was unimpaired. The mutant was complemented to nikkomycin X and I production by nikR expressed from the mel promoter of plasmid pIJ702. The nikR gene expressed in Escherichia coli led to the production of UPRTase activity. Our results indicate that NikR converts 4-formyl-4-imidazolin-2-one to yield 5'-phosphoribosyl-4 formyl-4-imidazolin-2-one, the precursor of nikkomycins containing this base. PMID- 10712602 TI - Several dinucleoside polyphosphates are acceptor substrates in the T4 RNA ligase catalyzed reaction. AB - Several dinucleoside polyphosphates accept cytidine-3', 5'-bisphosphate from the adenylylated donor 5'-adenylylated cytidine 5',3'-bisphosphate in the T4 RNA ligase catalyzed reaction. The 5'-adenylylated cytidine 5',3'-bisphosphate synthesized in a first step, from ATP and cytidine-3',5'-bisphosphate, is used as a substrate to transfer the cytidine-3',5'-bisphosphate residue to the 3'-OH group(s) of diguanosine tetraphosphate (Gp4G) giving rise to Gp4GpCp and pCpGp4GpCp in a ratio of approximately 10 : 1, respectively. The synthesized Gp4GpCp was characterized by treatment with snake venom phosphodiesterase and alkaline phosphatase and analysis (chromatographic position and UV spectra) of the reaction products by HPLC. The apparent Km values measured for Gp4G and 5' adenylylated cytidine 5',3'-bisphosphate in this reaction were approximately 4 mM and 0.4 mM, respectively. In the presence of 0.5 mM ATP and 0.5 mM cytidine-3',5' bisphosphate, the relative efficiencies of the following nucleoside(5')oligophospho(5')nucleosides as acceptors of cytidine-3',5' bisphosphate from 5'-adenylylated cytidine 5', 3'-bisphosphate are indicated in parentheses: Gp4G (100); Gp5G (101); Ap4G (47); Ap4A (39). Gp2G, Gp3G and Xp4X were not substrates of the reaction. Dinucleotides containing two guanines and at least four inner phosphates were the preferred acceptors of cytidine-3', 5' bisphosphate at their 3'-OH group(s). PMID- 10712603 TI - Aspartic protease inhibitors. An integrated approach for the design andsynthesis of diaminodiol-based peptidomimetics. AB - Aspartic proteases play key roles in a variety of pathologies, including acquired immunodeficiency syndrome. Peptidomimetic inhibitors can act as drugs to combat these pathologies. We have developed an integrated methodology for preparing human immunodeficiency virus (HIV)-1 aspartic protease diaminodiol inhibitors, based on a computational method that predicts the potential inhibitory activity of the designed structures in terms of calculated enzyme-inhibitor complexation energies. This is combined with a versatile synthetic strategy that couples a high degree of stereochemical control in the central diaminodiol module with complete flexibility in the choice of side chains in the core and in flanking residues. A series of 23 tetrameric, pentameric and hexameric inhibitors, with a wide range of calculated relative complexation energies (-47.2 to +117 kJ.mol-1) and predicted hydrophobicities (logPo/w = 1.8-8.4) was thus assembled from readily available amino acids and carboxylic acids. The IC50 values for these compounds ranged from 3.2 nM to 90 microM, allowing study of correlations between structure and activity, and individuation of factors other than calculated complexation energies that determine the inhibition potency. Multivariable regression analysis revealed the importance of side-chain bulkiness and rigidity at the P2, P2' positions, suggesting possible improvements for the prediction process used to select candidate structures. PMID- 10712605 TI - The role of amino-acid residues in the hydrophobic patch surrounding the haem group of cytochrome f in the interaction with plastocyanin. AB - Soluble turnip cytochrome f has been purified from the periplasmic fraction of Escherichia coli expressing a truncated petA gene encoding the precursor protein lacking the C-terminal 33 amino-acid residues. The protein is identical [as judged by 1H-NMR spectroscopy, midpoint redox potential (+ 365 mV) and electron transfer reactions with plastocyanin] to cytochrome f purified from turnip leaves. Several residues in the hydrophobic patch surrounding the haem group have been changed by site-directed mutagenesis, and the proteins purified from E. coli. The Y1F and Q7N mutants showed only minor changes in the plastocyanin binding constant Ka and the second-order rate constant for electron transfer to plastocyanin, whereas the Y160S mutant showed a 30% decrease in the overall rate of electron transfer caused in part by a 60% decrease in binding constant and partially compensated by an increased driving force due to a 27-mV decrease in redox potential. In contrast, the F4Y mutant showed increased rates of electron transfer which may be ascribed to an increased binding constant and a 14-mV decrease in midpoint redox potential. This indicates that subtle changes in the hydrophobic patch can influence rates of electron transfer to plastocyanin by changing the binding constants and altering the midpoint redox potential of the cytochrome haem group. PMID- 10712604 TI - Suramin blocks nucleotide triphosphate binding to ribosomal protein L3 from Trypanoplasma borreli. AB - Ribosomal protein L3 (L3) has been demonstrated to participate in formation of the peptidyltransferase center and is essential for its catalytic activity. In the present study we show that L3 is able to bind nucleotide triphosphates with high and specific affinity in vitro. L3 was serendipitously identified by screening of a genomic phage library from a primitive kinetoplastid flagellate Trypanoplasma borreli with the ATPase domain of the topoisomerase II gene as a probe. The cloned gene was overexpressed and purified as a his-tag fusion protein in E. coli. Radioligand binding experiments, using [gamma-35S]ATP, showed that L3 is able to bind ATP but also GTP and UTP with similar high affinity (IC50 50-100 nM), while it has no ATPase activity. Furthermore, we showed that L3 has more than 500-fold higher affinity for nucleotide triphosphates compared to the corresponding nucleotide monophosphates and diphosphates. Molecular genetic and biochemical analyses allowed us to localize the NTP binding domain of L3 to the N terminal 296 residues. Suramin, a polysulfonated naphthylamine derivative of urea, known for its chemotherapeutic effects completely inhibited the binding of [gamma-35S]ATP at subclinical levels. Results obtained with surface plasmon resonance technology showed that suramin both forms weak multimolecular complexes with L3 and binds strongly to L3 in nearly stoichiometric amounts. PMID- 10712607 TI - Interaction and coordination geometries for Ag(I) in the two metal sites of hemocyanin. AB - 111Ag(I) perturbed angular correlations of gamma-rays (PAC) has been used to investigate the binuclear metal site of 111Ag(I)-substituted Carcinus aestuarii deoxyhemocyanin. The studies have shown that apo-hemocyanin is able to bind 2 mol of Ag(I) per mol of protein and that the binding is specific for the metal ion sites. The PAC spectra show pronounced changes when the stoichiometry of Ag(I) to protein is increased from 0.1 to 2.0. These changes have been interpreted as evidence of interactions between the two sites in terms of a structural destabilization of the first occupied site caused by the occupation of the second site. The experimental data for the Ag(I)-substituted metal sites do not agree well with the three-coordinated structure found in the Cu(I) holo-protein. However, if a water molecule is included as a coordinating ligand in the Ag(I) metal site a successful interpretation of the experimental data can be obtained. PMID- 10712606 TI - Characterization of cis-acting elements in the promoter of the mouse metallothionein-3 gene. Activation of gene expression during neuronal differentiation of P19 embryonal carcinoma cells. AB - The metallothionein (MT)3 gene is expressed predominantly in the brain and the organs of the reproductive system, and fails to respond to metal ions in vivo. A CTG repeat was proposed to function as a potential repressor element in nonpermissive cells, and a sequence similar to the JC virus silencer element was found to function as a negative element in permissive primary astrocytes. The objective of this study was to characterize further the mechanisms governing cell type specific MT-3 gene transcription. We searched for a suitable cell line expressing the MT-3 gene to be used for determination of MT-3 promoter tissue specificity, and showed that MT-3 expression is activated during neuroectodermal differentiation of P19 cells induced by retinoic acid to levels similar to those found in whole brain. Deletion of the CTG repeat or of the JC virus silencer did not promote MT-3 promoter activity in nonpermissive cells, or enhance expression in permissive cells. We identified MT-3 promoter sequences interacting with liver and brain nuclear proteins, as assayed by DNase I footprinting analyses and electrophoretic mobility shift assay, and assessed the role of these sequences in the regulation of MT-3 expression by cotransfection experiments. We generated stable transfectants in permissive C6 and nonpermissive NIH-3T3 cells, and analysed the methylation status of the MT-3 gene. These studies show that regulation of tissue-specific MT-3 gene expression does not appear to involve a repressor, and suggest that other mechanisms such as chromatin organization and epigenetic modifications could account for the absence of MT-3 gene transcription in nonpermissive cells. PMID- 10712608 TI - Effects of cadmium on manganese peroxidase competitive inhibition of MnII oxidation and thermal stabilization of the enzyme. AB - Inhibition of manganese peroxidase by cadmium was studied under steady-state and transient-state kinetic conditions. CdII is a reversible competitive inhibitor of MnII in the steady state with Ki approximately 10 microM. CdII also inhibits enzyme-generated MnIII-chelate-mediated oxidation of 2,6-dimethoxyphenol with Ki approximately 4 microM. CdII does not inhibit direct oxidation of phenols such as 2,6-dimethoxyphenol or guaiacol (2-methoxyphenol) in the absence of MnII. CdII alters the heme Soret on binding manganese peroxidase and exhibits a Kd approximately 8 microM, similar to Mn (Kd approximately 10 microM). Under transient-state conditions, CdII inhibits reduction of compound I and compound II by MnII at pH 4.5. However, CdII does not inhibit formation of compound I nor does it inhibit reduction of the enzyme intermediates by phenols in the absence of MnII. Kinetic analysis suggests that CdII binds at the MnII-binding site, preventing oxidation of MnII, but does not impair oxidation of substrates, such as phenols, which do not bind at the MnII-binding site. Finally, at pH 4.5 and 55 degrees C, MnII and CdII both protect manganese peroxidase from thermal denaturation more efficiently than CaII, extending the half-life of the enzyme by more than twofold. Furthermore, the combination of half MnII and half CdII nearly quadruples the enzyme half-life over either metal alone or either metal in combination with CaII. PMID- 10712609 TI - A hydrophobic sequence at position 313-316 (Leu-Ala-Phe-Trp) in the fifth domain of apolipoprotein H (beta2-glycoprotein I) is crucial for cardiolipin binding. AB - Apolipoprotein H (apoH, protein; APOH, gene) binds to negatively charged phospholipids, which triggers the production of a subset of autoantibodies against phospholipid in patients with autoimmune diseases. We have demonstrated that two naturally occurring missense mutations in the fifth domain of apoH, Trp316Ser and Cys306Gly, disrupt the binding of native apoH to phosphatidylserine [Sanghera, D. K., Wagenknecht, D. R., McIntyre, J. A. & Kamboh, M. I. (1997) Hum. Mol. Genet. 6, 311-316]. To confirm whether these are functional mutations, we mutagenized APOH cDNAs and transiently expressed them in COS-1 cells. The cardiolipin ELISA of wild-type and mutant recombinant apoH confirmed that the Gly306 and Ser316 mutations are responsible for abolishing the binding of recombinant apoH to cardiolipin. These mutations, however, had no effect on the levels of expression or secretion of recombinant apoH in transfected COS-1 cells. While the Cys306Gly mutation disrupts a disulfide bond between Cys306 and Cys281, which appears to be critical for clustering positively charged amino acids, the Trp316Ser mutation affects the integrity of an evolutionarily conserved hydrophobic sequence at position 313-316 (Leu-Ala-Phe-Trp), which is hypothesized to interact with anionic phospholipid. To test this hypothesis, we exchanged the remaining three hydrophobic amino acids with neutral amino acids by site-directed mutagenesis (Leu313Gly, Ala314Ser and Phe315Ser). Binding of the Leu313Gly and Phe315Ser mutants to cardiolipin was significantly reduced to 25% and 13%, respectively, of that of the wild-type. On the other hand, the Ala314Ser mutation showed normal cardiolipin binding. Taken together with our previous findings, these results strongly suggest that the configuration of the fifth domain of apoH, as well as the integrity of the highly conserved hydrophobic amino acids at positions 313-316, is essential for the binding of apoH to anionic phospholipid. PMID- 10712610 TI - Localization of the catalytic activity in restrictocin molecule by deletion mutagenesis. AB - Restrictocin, produced by the fungus Aspergillus restrictus, is a highly specific ribonucleolytic toxin which cleaves a single phosphodiester bond between G4325 and A4326 in the 28S rRNA. It is a nonglycosylated, single-chain, basic protein of 149 amino acids. The putative catalytic site of restrictocin includes Tyr47, His49, Glu95, Arg120 and His136. To map the catalytic activity in the restrictocin molecule, and to study the role of N- and C-terminus in its activity, we have systematically deleted amino-acid residues from both the termini. Three N-terminal deletions removing 8, 15 and 30 amino acids, and three C-terminal deletions lacking 4, 6, and 11 amino acids were constructed. The deletion mutants were expressed in Escherichia coli, purified to homogeneity and functionally characterized. Removal of eight N-terminal or four C-terminal amino acids rendered restrictocin partially inactive, whereas any further deletions from either end resulted in the complete inactivation of the toxin. The study demonstrates that intact N- and C-termini are required for the optimum functional activity of restrictocin. PMID- 10712611 TI - The NMR solution structure of the ion channel peptaibol chrysospermin C bound to dodecylphosphocholine micelles. AB - Chrysospermin C is a 19-residue peptaibol capable of forming transmembrane ion channels in phospholipid bilayers. The conformation of chrysospermin C bound to dodecylphosphocholine micelles has been solved using heteronuclear NMR spectroscopy. Selective 15N-labeling and 13C-labeling of specific alpha aminoisobutyric acid residues was used to obtain complete stereospecific assignments for all eight alpha-aminoisobutyric acid residues. Structures were calculated using 339 distance constraints and 40 angle constraints obtained from NMR data. The NMR structures superimpose with mean global rmsd values to the mean structure of 0. 27 A (backbone heavy atoms) and 0.42 A (all heavy atoms). Chrysospermin C bound to decylphosphocholine micelles displays two well-defined helices at the N-terminus (residues Phe1-Aib9) and C-terminus (Aib13-Trp-ol19). A slight bend preceding Pro14, i.e. encompassing residues 10-12, results in an angle of approximately 38 degrees between the mean axes of the two helical regions. The bend structure observed for chrysospermin C is compatible with the sequences of all 18 long peptaibols and may represent a common 'active' conformation. The structure of chrysospermin C shows clear hydrophobic and hydrophilic surfaces which would be appropriate for the formation of oligomeric ion channels. PMID- 10712612 TI - Fluorescent neoglycolipids. Improved probes for oligosaccharide ligand discovery. AB - A second generation of lipid-linked oligosaccharide probes, fluorescent neoglycolipids, has been designed and synthesized for ligand discovery within highly complex mixtures of oligosaccharides. The aminolipid 1,2-dihexadecyl-sn glycero-3-phosphoethanolamine (DHPE), which has been used extensively to generate neoglycolipids for biological and structural studies, has been modified to incorporate a fluorescent label, anthracene. This new lipid reagent, N aminoacetyl-N-(9-anthracenylmethyl)-1, 2-dihexadecyl-sn-glycero-3 phosphoethanolamine (ADHP), synthesized from anthracenaldehyde and DHPE gives an intense fluorescence under UV light. Fluorescent neoglycolipids derived from a variety of neutral and acidic oligosaccharides by conjugation to ADHP, by reductive amination, can be detected and quantified by spectrophotometry and scanning densitometry, and resolved by TLC and HPLC with subpicomole detection. Antigenicities of the ADHP-neoglycolipids are well retained, and picomole levels can be detected using monoclonal carbohydrate sequence-specific antibodies. Among O-glycans from an ovarian cystadenoma mucin, isomeric oligosaccharide sequences, sialyl-Lea- and sialyl-Lex-active, could be resolved by HPLC as fluorescent neoglycolipids, and sequenced by liquid secondary-ion mass spectrometry. Thus the neoglycolipid technology now uniquely combines high sensitivity of immuno detection with a comparable sensitivity of chemical detection. Principles are thus established for a streamlined technology whereby an oligosaccharide population is carried through ligand detection and ligand isolation steps, and sequence determination by mass spectrometry, enzymatic sequencing and other state of-the-art technologies for carbohydrate analysis. PMID- 10712613 TI - Molecular cloning of the human and murine 2-amino-3-ketobutyrate coenzyme A ligase cDNAs. AB - The conversion of L-threonine to glycine in both prokaryotes and eukaryotes takes place through a two-step biochemical pathway involving the enzymes L-threonine dehydrogenase (EC 1.1.1103) and 2-amino-3-ketobutyrate coenzyme A ligase (KBL; EC 2.3.1.29). The genes encoding these enzymes have been described in prokaryotes but not in eukaryotes. We report the cloning of transcripts for KBL, the second enzyme in the pathway, from human and murine lung and a partial transcript from bovine liver. Two peptide sequences from the purified bovine KBL protein, one from the N-terminus and the other from the peptide containing the pyridoxal 5' phosphate-binding lysine residue [Tong, H. & Davis, L. (1994) J. Biol. Chem. 269, 4057-4064], are identical with regions of the conceptual translation of the transcript obtained from bovine liver. The partial transcript from bovine liver was very similar to the human transcript, being 91% and 92% identical at the nucleotide and amino-acid levels, respectively. The human and murine KBL transcripts are 1.5 kb long, with ORFs encoding proteins of 419 and 416 residues, respectively. The mouse protein has 90% identity with the human protein. The human transcript is strongly expressed in heart, brain, liver and pancreas compared with the lung. The N-termini of both human and mouse proteins have characteristics of mitochondrial import sequences. Both human and murine proteins have 54% identity with the well-characterised prokaryote KLB protein from Escherichia coli. Database searches with the human cDNA sequence enabled us to identify the human KBL gene on chromosome 22q12-13, consisting of nine exons over 9 kb, and a hypothetical Caenorhabditis elegans KLB gene on chromosome IV, consisting of five exons over 2 kb. PMID- 10712614 TI - A second functional delta5 fatty acid desaturase in the cellular slime mould Dictyostelium discoideum. AB - A cDNA with homology to fatty acid desaturases was selected by searching the cDNA data bank of Dictyostelium discoideum (http://www. csm.biol.tsukuba.ac.jp/cDNAproject.html) with conserved histidine box motifs. Using this sequence, genomic DNA encoding the Delta5 desaturase was amplified from the genomic DNA of D. discoideum, and its desaturase activity was confirmed by the overexpression mutation in D. discoideum and the gain-of-function mutation in yeast. The cloned cDNA is 1565 nucleotides in length, and the deduced amino acid sequence comprised 467 amino-acid residues containing an N-terminal cytochrome b5 domain that shared 43% identity with cytochrome b5 of Oryza sativa. The whole sequence was 42% identical to the Delta5 desaturase of Mortierella alpina. This desaturase is a novel member of the cytochrome b5-containing Delta5 fatty acid desaturase. As we have already reported one other Delta5 desaturase in Dictyostelium, this organism is the first to be confirmed as having two functional Delta5 fatty acid desaturase genes. The substrate specificities of the two functional Delta5 desaturases of D. discoideum were also examined. PMID- 10712615 TI - Affinity for the cognate monoclonal antibody of synthetic peptides derived from selection by phage display. Role of sequences flanking thebinding motif. AB - Randomized peptide sequences displayed at the surface of filamentous phages are often used to select antibody ligands. The selected sequences are generally further used in the form of synthetic peptides; however, as such, their affinity for the selecting antibody is extremely variable and factors influencing this affinity have not been fully deciphered. We have used an f88.4 phage-displayed peptide library to identify ligands of mAb 11E12, an antibody reactive to human cardiac troponin I. A majority of the sequences thus selected showed a (T/A/I/L) EP(K/R/H) motif, homologous to the Y-TEPH motif identified by multiple peptide synthesis as the critical motif recognized by mAb 11E12 in the peptide epitope. A set of 15-mer synthetic peptides derived from the phage-selected sequences was used in BIACORE to characterize their interaction with mAb 11E12. Most peptides exhibited affinities in the 7-26 nM range. These affinities represented, however, only 1.9-7. 5% of the affinity of the 15-mer peptide epitope. In circular dichroism experiments, the peptide epitope showed a propensity to have some stabilized conformation, whereas a low-affinity peptide selected by phage-display did not. To try to decipher the molecular basis of this difference in affinity, new peptides were prepared by grafting the N- or the C-terminal sequence of the peptide epitope to the Y-TEPK motif of a low-affinity peptide selected by phage display. These hybrid peptides showed marked increases both in affinity (as assessed using BIACORE) and in inhibitory potency (as assessed in competition ELISA), compared with the parent sequence. Thus, the sequences flanking the motif, although not containing critical residues, convey some determinants necessary for high affinity. The affinity of a given peptide strongly depends on its capacity to maintain the antigenically reactive structure it has on the phage, implying that it is impossible to predict whether high- or low-affinity peptides will be obtained from phage display. PMID- 10712616 TI - Pyridoxal 5'-phoshate schiff base in Citrobacter freundii tyrosinephenol-lyase. Ionic and tautomeric equilibria. AB - Spectral properties of the internal Schiff base in tyrosine phenol-lyase have been investigated in the presence of an activating cation K+ and a cation inhibitor Na+. The holoenzyme absorption spectra in the pH range 6.5-8.7 were recorded in the presence of K+. No apparent pKa value of the coenzyme chromophore was found in this pH range, indicating that the internal Schiff base does not change its ionic form on going from pH 6.5 to 8.7. To determine the ionic state and tautomeric composition of the Schiff base in tyrosine phenol-lyase, the absorption and circular dichroism spectra were analyzed using lognormal distribution curves. The predominant form of the internal Schiff base is that with protonated pyridinium and aldimine nitrogen atoms and deprotonated 3' hydroxy group, i.e. the ketoenamine. This form is in prototropic equilibrium with its enolimine tautomer. The internal aldimine ionic form is changed upon replacement of K+ with Na+. This replacement leads to a significant decrease in the pKa value of pyridinium nitrogen of the pyridoxal-P. PMID- 10712617 TI - Structural analysis of two glycosphingolipids from the lipopolysaccharide-lacking bacterium Sphingomonas capsulata. AB - Two glycosphingolipids, GSL-1 and GSL-3, were isolated from Sphingomonas capsulata and studied by methylation analysis, laser desorption mass spectrometry, and 1H and 13C NMR spectroscopy, including two-dimensional 1H,1H COSY and heteronuclear 13C,1H COSY experiments. GSL-1 and GSL-3 differ in their carbohydrate part, their structures being alpha-D-GlcpA-(1-->1)-Cer and alpha-D Galp-(1-->6)-alpha-D-GlcpN-(1-->4)-alpha-D-GlcpA(1-- >1)Cer, respectively. Variations occur in the ceramide of GSL-1 and GSL-3, both having the same long chain bases, erythro-2-amino-1, 3-octadecanediol (sphinganine), (13Z)-erythro-2 amino-13-eicosene-1, 3-diol and (13Z)-erythro-2-amino-13,14-methylene-1,3 eicosanediol, in the ratios 2.6 : 1 : 3.5 in GSL-1 and 1 : 1.2 : 1.5 in GSL-3. All bases are quantitatively substituted by amide-linked (S)-2-hydroxymyristic acid. PMID- 10712618 TI - Cloning and over-expression of thermostable Bacillus sp. YM55-1 aspartase and site-directed mutagenesis for probing a catalytic residue. AB - A thermostable aspartase gene (aspB) from Bacillus sp. YM55-1 was cloned and the gene sequenced. The aspB gene (1407 bp ORF) encodes a protein with a molecular mass of 51 627 Da, consisting of 468 amino-acid residues. An amino-acid sequence comparison revealed that Bacillus YM55-1 aspartase shared 71% homology with Bacillus subtilis aspartase and 49% with Escherichia coli and Pseudomonas fluorescens aspartases. The E. coli TK237/pUCASPB strain, which was obtained by transforming E. coli TK237 (aspartase-null strain) with a vector plasmid (pUCASPB) containing the cloned aspB gene, produced a large amount of the enzyme corresponding to > 10% of the total soluble protein. The over-expressed recombinant enzyme (native molecular mass: 200 kDa) was purified effectively and rapidly using heat treatment and affinity chromatography. In order to probe the catalytic residues of this enzyme, two conserved amino-acid residues, Lys183 and His134, were individually mutated to alanine. Although the tertiary structure of each mutant was estimated to be the same as that of wild-type aspartase in CD and fluorescence measurements, the Lys183Ala mutant lost its activity completely, whereas His134Ala retained full activity. This finding suggests that Lys183 may be involved in the catalytic activity of this thermostable Bacillus YM55-1 aspartase. PMID- 10712619 TI - Exploring substrate binding and discrimination in fructose1, 6-bisphosphate and tagatose 1,6-bisphosphate aldolases. AB - Fructose 1,6-bisphosphate aldolase catalyses the reversible condensation of glycerone-P and glyceraldehyde 3-phosphate into fructose 1,6-bisphosphate. A recent structure of the Escherichia coli Class II fructose 1,6-bisphosphate aldolase [Hall, D.R., Leonard, G.A., Reed, C.D., Watt, C.I., Berry, A. & Hunter, W.N. (1999) J. Mol. Biol. 287, 383-394] in the presence of the transition state analogue phosphoglycolohydroxamate delineated the roles of individual amino acids in binding glycerone-P and in the initial proton abstraction steps of the mechanism. The X-ray structure has now been used, together with sequence alignments, site-directed mutagenesis and steady-state enzyme kinetics to extend these studies to map important residues in the binding of glyceraldehyde 3 phosphate. From these studies three residues (Asn35, Ser61 and Lys325) have been identified as important in catalysis. We show that mutation of Ser61 to alanine increases the Km value for fructose 1, 6-bisphosphate 16-fold and product inhibition studies indicate that this effect is manifested most strongly in the glyceraldehyde 3-phosphate binding pocket of the active site, demonstrating that Ser61 is involved in binding glyceraldehyde 3-phosphate. In contrast a S61T mutant had no effect on catalysis emphasizing the importance of an hydroxyl group for this role. Mutation of Asn35 (N35A) resulted in an enzyme with only 1.5% of the activity of the wild-type enzyme and different partial reactions indicate that this residue effects the binding of both triose substrates. Finally, mutation of Lys325 has a greater effect on catalysis than on binding, however, given the magnitude of the effects it is likely that it plays an indirect role in maintaining other critical residues in a catalytically competent conformation. Interestingly, despite its proximity to the active site and high sequence conservation, replacement of a fourth residue, Gln59 (Q59A) had no significant effect on the function of the enzyme. In a separate study to characterize the molecular basis of aldolase specificity, the agaY-encoded tagatose 1,6 bisphosphate aldolase of E. coli was cloned, expressed and kinetically characterized. Our studies showed that the two aldolases are highly discriminating between the diastereoisomers fructose bisphosphate and tagatose bisphosphate, each enzyme preferring its cognate substrate by a factor of 300 1500-fold. This produces an overall discrimination factor of almost 5 x 105 between the two enzymes. Using the X-ray structure of the fructose 1,6 bisphosphate aldolase and multiple sequence alignments, several residues were identified, which are highly conserved and are in the vicinity of the active site. These residues might potentially be important in substrate recognition. As a consequence, nine mutations were made in attempts to switch the specificity of the fructose 1,6-bisphosphate aldolase to that of the tagatose 1,6-bisphosphate aldolase and the effect on substrate discrimination was evaluated. Surprisingly, despite making multiple changes in the active site, many of which abolished fructose 1, 6-bisphosphate aldolase activity, no switch in specificity was observed. This highlights the complexity of enzyme catalysis in this family of enzymes, and points to the need for further structural studies before we fully understand the subtleties of the shaping of the active site for complementarity to the cognate substrate. PMID- 10712620 TI - Complexes of smooth muscle tropomyosin with F-actin studied by differential scanning calorimetry. AB - Differential scanning calorimetry (DSC) and light scattering were used to analyze the interaction of duck gizzard tropomyosin (tropomyosin) with rabbit skeletal muscle F-actin. In the absence of F-actin, tropomyosin, represented mainly by heterodimers, unfolds at 41 degrees C with a sharp thermal transition. Interaction of tropomyosin heterodimers with F-actin causes a 2-6 degrees C shift in the tropomyosin thermal transition to higher temperature, depending on the tropomyosin/actin molar ratio and protein concentration. A pronounced shift of the tropomyosin thermal transition was observed only for tropomyosin heterodimers, and not for homodimers. The most pronounced effect was observed after complete saturation of F-actin with tropomyosin molecules, at tropomyosin/actin molar ratios > 1 : 7. Under these conditions, two well separated peaks of tropomyosin were observed on the thermogram besides the peak of F-actin, the peak characteristic of free tropomyosin heterodimer, and the peak with a maximum at 45-47 degrees C corresponding to tropomyosin bound to F-actin. By measuring the temperature-dependence of light scattering, we found that thermal unfolding of tropomyosin is accompanied by its dissociation from F-actin. Thermal unfolding of tropomyosin is almost completely reversible, whereas F-actin denatures irreversibly. The addition of tropomyosin has no effect on thermal unfolding of F-actin, which denatures with a maximum at 64 degrees C in the absence and at 78 degrees C in the presence of a twofold molar excess of phalloidin. After the F-actin-tropomyosin complex had been heated to 90 degrees C and then cooled (i.e. after complete irreversible denaturation of F-actin), only the peak characteristic of free tropomyosin was observed on the thermogram during reheating, whereas the thermal transitions of F-actin and actin-bound tropomyosin completely disappeared. Therefore, the DSC method allows changes in thermal unfolding of tropomyosin resulting from its interaction with F-actin to be probed very precisely. PMID- 10712621 TI - Pig MAP/ITIH4 and haptoglobin are interleukin-6-dependent acute-phase plasma proteins in porcine primary cultured hepatocytes. AB - The acute-phase expression of pig MAP (major acute-phase protein)/ITIH4 (inter alpha-trypsin inhibitor heavy chain 4) and haptoglobin were analysed in primary cultures of isolated pig hepatocytes in response to recombinant human (rh) cytokines: tumor necrosis factor-alpha (TNF-alpha), interleukin-1 (IL-1), interleukin-6 (IL-6), as well as to bacterial lipopolysaccharide (LPS). Analysis of pig MAP/ITIH4 and haptoglobin mRNAs was carried out by RT-PCR amplification. Secreted proteins from the cytokine-treated hepatocytes were quantified by immunochemical techniques. Time-course and dose-response experiments show that pig MAP/ITIH4 and haptoglobin belong to the type II acute-phase proteins, as they are specifically induced by rhIL-6 and not by rhTNF-alpha or rhIL-1. Stimulation of cultured pig hepatocytes with rhIL-6 for 48 h at doses of 1000 U.mL-1 showed a fourfold to fivefold increase in pig MAP/ITIH4 concentration in the medium, while the concentration of haptoglobin only increased twofold. A similar increase in the concentration of pig MAP/ITIH4 was also observed in media of LPS-treated hepatocytes with the simultaneous generation of IL-6 by the Kupffer cells present in the cultures. Albumin secretion decreased after stimulation with doses of 100 or 1000 U.mL-1 rhTNF-alpha, rhIL-1 or rhIL-6. Therefore, it can be concluded that pig MAP/ITIH4 behaves as a major acute-phase protein produced by porcine hepatocytes under the effect of inflammatory cytokines. PMID- 10712622 TI - Neuronal and vascular localization of histamine N-methyltransferase in the bovine central nervous system. AB - Histamine N-methyltransferase (HMT) (EC 2.1.1.8) plays a crucial role in the inactivation of the neurotransmitter histamine in the CNS. However, the localization of HMT remains to be determined. In the present study, we investigated immunohistochemical localization of HMT in the bovine CNS using a polyclonal antibody against bovine HMT. The HMT-like immunoreactivity was observed mainly in neurons. Strongly immunoreactive neurons were present in the oculomotor nucleus and ruber nucleus in the midbrain, the facial nucleus in the pons, the dorsal vagal nucleus and hypoglossal nucleus in the medulla oblongata and in the anterior horn as well as intermediolateral zone of the spinal cord. Intermediately immunoreactive neurons were present in the piriform cortex and the inferior olivary nucleus. The grey matter of the forebrain regions was diffusely and faintly stained. In the cerebellum and the striatum, the nerve fibres in the white matter were positive. The tuberomammillary nucleus, where histaminergic neurons are present, were weakly positive. The other immunoreactive structures in the CNS were blood vessels. Almost all of the blood vessel walls, irrespective of whether they were arterial or venous, were variably stained. The glial fibrillary acidic protein- (GFAP-) immunoreactive astrocytes were not stained. These findings indicated that histamine released from histaminergic nerve terminals or varicose fibres is methylated mainly in postsynaptic or extrasynaptic neurons rather than in astrocytes. The localization of HMT in the blood vessel wall may mean that blood-borne histamine and histamine released from mast cells associated with the blood vessels are catabolized in this structure. PMID- 10712623 TI - Differential sensitivity to Zolpidem of IPSPs activated by morphologically identified CA1 interneurons in slices of rat hippocampus. AB - Hippocampal pyramidal cells express several alpha-subunits, which determine the affinity of GABAA (gamma-aminobutyric acid) receptors for benzodiazepine site ligands. This study asked whether inhibitory postsynaptic potentials (IPSPs) elicited by specific interneuronal subclasses were differentially sensitive to the alpha1-preferring agonist Zolpidem, i.e. whether different receptors mediate different inhibitory connections. Paired intracellular recordings in which the presynaptic cell was an interneuron and the postsynaptic cell a CA1 pyramid were performed in slices of adult rat hippocampus. Resultant IPSPs were challenged with Zolpidem, cells filled with biocytin and identified morphologically. IPSPs elicited by fast spiking (FS) basket cells (n = 9) were enhanced more than IPSPs elicited by regular spiking (RS) basket cells (n = 10). At FS basket cell synapses the efficacy of Zolpidem was equivalent to that of Diazepam, while RS basket cell IPSPs are enhanced 50% less by Zolpidem than by Diazepam. Thus, while alpha1 subunits may dominate at synapses supplied by FS basket cells, RS basket cell synapses also involve alpha2/3 subunits. Two bistratified cell IPSPs tested with Zolpidem did not increase in amplitude, despite powerful enhancements of bistratified cell IPSPs by Diazepam, consistent with previous indications that these synapses utilize alpha5-containing receptors. Enhancements of basket cell IPSPs by Zolpidem and Diazepam were bi- or triphasic with steep amplitude increases separated by plateaux, occurring 10-15, 25-30 and 45-55 min after adding the drug to the bath. The entire enhancement was, however, blocked by the antagonist Flumazenil (n = 7). Flumazenil, either alone (n = 3), or after Zolpidem, reduced IPSP amplitude to approximately 90% of control, suggesting that alpha4-containing receptors were not involved. PMID- 10712624 TI - Light-adaptive effects of retinoic acid on receptive field properties of retinal horizontal cells. AB - Besides its role in ocular development, retinoic acid (RA), which is a light correlated byproduct of the phototransduction cycle, was recently shown to affect light-driven synaptic plasticity in the outer plexiform layer of the adult fish retina. Tuning by ambient light conditions of the retinal network properties is very prominent in outer plexiform layer circuits, and we therefore examined whether RA could affect cone horizontal cell physiology similar to ambient light. Performing intracellular recordings and dye injections in the dark-adapted inverted eyecup preparation of the carp, we found that RA reduced the receptive fields of horizontal cell somata and impaired gap junctional communication. This action was not observed among coupled axon terminals of horizontal cells and appeared to be stereospecific because it could only be attributed to all-trans and 13-cis RA but not to the 9-cis isomer and photoisomerized all-trans RA. Modulation of receptive field size occurred independently of the dopaminergic system. Furthermore, RA affected the light responsiveness of cone horizontal cells. Compared to the dark-adapted condition, responsiveness to intense light stimulation was enhanced but decreased when low intensities were used. Moreover, following RA treatment H2-type horizontal cells of dark-adapted retinae which do not give rise to colour-opponent light properties became colour-opponent and performed depolarizing responses to long-wavelength stimulation. In all these cases RA perfectly matched the effects of light adaptation, supporting the notion that RA acts as an endogenous neuromodulator. PMID- 10712626 TI - Leukaemia inhibitory factor is required for normal inflammatory responses to injury in the peripheral and central nervous systems in vivo and is chemotactic for macrophages in vitro. AB - The cytokine leukaemia inhibitory factor (LIF) is up-regulated in glial cells after injury to the peripheral and central nervous systems. In addition, LIF is required for the changes in neuropeptide expression that normally occur when the axons of sympathetic and sensory neurons are transected. We investigated whether LIF is also necessary for the initial inflammatory response that follows mechanical injury to the sciatic nerve and cerebral cortex of adult mice. We find that inflammatory cell infiltration into crushed sciatic nerve is significantly slower in LIF knock-out (KO) mice compared with wild-type (WT) mice. Similarly, the microglial and astroglial responses to surgical injury of the cortex are significantly slower in LIF KO mice compared with WT mice. Consistent with these in vivo results, LIF is chemotactic for peritoneal macrophages in a microchamber culture assay. Thus, LIF is a key regulator of neural injury in vivo, where it is produced by glia and can act directly on neurons, glia and inflammatory cells. We also find that the initial inflammatory response to cortical injury is diminished in interleukin (IL)-6 KO mice. Surprisingly, however, the inflammatory response in LIF-IL-6 double KO mice is very similar to that of the single KO mice, suggesting that these cytokines may act in series rather than in parallel in this response. PMID- 10712625 TI - A dynamic regulation of GDNF-family receptors correlates with a specific trophic dependency of cranial motor neuron subpopulations during development. AB - Glial cell line-derived neurotrophic factor (GDNF) family ligands promote the survival of developing motor neurons in vivo and in vitro. However, not all neurons survive with any single ligand in culture and GDNF null mutant mice display only a partial motor neuron loss. An interesting possibility is that subpopulations of motor neurons based on their function and/or their myotopic organization require distinct members of GDNF family ligands. Because responsiveness to the different ligands depends on the expression of their cognate ligand-binding receptor we have herein addressed this issue by examining the expression of GDNF-family receptors (gfr) during development and in the adult in cranial motor nuclei subpopulations. We have furthermore examined the in vivo role of GDNF for cranial motor neuron subpopulations. The shared ret receptor was expressed in all somatic, branchial and visceral cranial embryonic motor nuclei examined, showing that they are all competent to respond to GDNF family ligands during development. At early stages of development both the GDNF receptor, gfralpha1, and the neurturin (NTN) receptor, gfralpha2, were expressed in the oculomotor, facial and spinal accessory, and only gfralpha1 in the trochlear, superior salivatory, trigeminal, hypoglossal and weakly in the dorsal motor nucleus of the vagus and the ambiguous nucleus. The abducens nucleus was negative for both gfralpha1 and gfralpha2. The artemin (ART) receptor, gfralpha3, was expressed only in the superior salivatory nucleus. A motor neuron subnuclei specific expression of gfralpha1 and gfralpha2 was seen in the facial and trigeminal nuclei which corresponded to their dependence on GDNF in null mutant mice. We found that the expression was dynamic in these nuclei, which may reflect developmental changes in their trophic factor dependency. Analysis of GDNF null mutant mice revealed that the dynamic receptor expression is regulated by the ligand in vivo, indicating that the attainment of changes in dependency could be ligand induced. Our results indicate that specific GDNF family ligands support selective muscle-motor neuron circuits during development. PMID- 10712627 TI - Acetylcholine receptors are required for postsynaptic aggregation driven by the agrin signalling pathway. AB - To investigate the role of acetylcholine receptors (AChRs) in the aggregation of postsynaptic molecules on muscle cells, we utilized the 1R- genetic variant of C2 muscle cells which has very little expression of AChRs in its cell membrane. On C2 myotubes, AChRs cluster spontaneously, with the frequency of clustering greatly enhanced by motor neuron-derived agrin. Signal transduction events driven by agrin, including the tyrosine phosphorylation of muscle-specific kinase (MuSK) and the AChR beta subunit, have been implicated as requirements of postsynaptic scaffold assembly. We show here that some molecules of the postsynaptic scaffold spontaneously aggregate and colocalize on 1R- myotubes at very low frequency, including an as yet unidentified agrin binding molecule, beta-dystroglycan and MuSK. Agrin is unable to increase the frequency of these aggregations, but does cause tyrosine phosphorylation of MuSK. We conclude that free molecules can associate into aggregates independently of AChRs, but AChRs are required for high frequency molecular aggregation driven by the agrin signalling pathway. MuSK tyrosine phosphorylation appears to precede a requisite event involving AChRs that aggregates postsynaptic molecules. PMID- 10712628 TI - NOS inhibition during postnatal development leads to increased ipsilateral retinocollicular and retinogeniculate projections in rats. AB - Synthesis of nitric oxide (NO) occurs downstream from activation of N-methyl-D aspartate (NMDA) receptors; NO reportedly acts as a retrograde messenger, influencing the refinement and stabilization of coactive afferent terminals. Cells and neuropil in the rat superior colliculus (SC) and lateral geniculate body (LGB) show intense, developmentally regulated activity for NO synthase (NOS). To study the role of NO in the development of retinogeniculate and retinotectal axon arbors, we examined primary visual projections of rats that had received intraperitoneal injections of Nomega-nitro-L-arginine (L-NoArg, an NOS inhibitor) on postnatal day 0, and daily thereafter for 4-6 weeks. Treated rats showed significant alterations in ipsilateral retinotectal projections, in the mediolateral and anteroposterior axes; there was an increase in the density of fibres entering the SC, in branch length, and in the numbers of boutons on retinotectal arbors in the treated group. Ipsilaterally projecting retinal axons also showed an increase in density and distribution in the dorsal nucleus of the LGB. If animals were allowed to survive for several months after stopping treatment, similar changes were also noted, but these were much less striking. Our results support the hypothesis that, in the mammalian visual system, NO released from target neurons in the SC and LGB serves as a retrograde signal which feeds back on retinal afferents, influencing their growth. The effects of NOS inhibition are partially reversed after treatment is stopped, indicating that lack of NO synthesis delays the maturation of retinofugal connections, and also that NO plays a constitutive role in their development. PMID- 10712629 TI - Abnormal axonal physiology is associated with altered expression and distribution of Kv1.1 and Kv1.2 K+ channels after chronic spinal cord injury. AB - Dysfunction of surviving axons which traverse the site of spinal cord injury (SCI) has been linked to altered sensitivity to the K+ channel blocker 4 aminopyridine (4-AP) and appears to contribute to post-traumatic neurological deficits although the underlying mechanisms remain unclear. In this study, sucrose gap electrophysiology in isolated dorsal column strips, Western blotting and confocal immunofluorescence microscopy were used to identify the K+ channels associated with axonal dysfunction after chronic (6-8 weeks postinjury) clip compresssion SCI of the thoracic cord at T7 in rats. The K+ channel blockers 4-AP (200 microM, 1 mM and 10 mM) and alpha-dendrotoxin (alpha-DTX, 500 nM) resulted in a significant relative increase in the amplitude and area of compound action potentials (CAP) recorded from chronically injured dorsal column axons in comparison with control noninjured preparations. In contrast, TEA (10 mM) and CsCl (2 mM) had similar effects on injured and control spinal cord axons. Western blotting and quantitative immunofluorescence microscopy showed increased expression of Kv1.1 and Kv1.2 K+ channel proteins on spinal cord axons following injury. In addition, Kv1.1 and Kv1.2 showed a dispersed staining pattern along injured axons in contrast to a paired juxtaparanodal localization in uninjured spinal cord axons. Furthermore, labelled alpha-DTX colocalized with Kv1.1 and Kv1.2 along axons. These findings suggest a novel mechanism of axonal dysfunction after SCI whereby an increased 4-AP- and alpha-DTX-sensitive K+ conductance, mediated in part by increased Kv1.1 and Kv1.2 K+ channel expression, contributes to abnormal axonal physiology in surviving axons. PMID- 10712631 TI - Hyperpolarization-activated current, Ih, in inspiratory brainstem neurons and its inhibition by hypoxia. AB - A hyperpolarization-activated current, Ih, is often implied in pacemaker-like depolarizations during rhythmic oscillatory activity. We describe Ih in the isolated respiratory centre of immature mice (P6-P11). Ih was recorded in 15% (22/146) of all inspiratory neurons examined. The mean half-maximal Ih activation occurred at -78 mV and the reversal potential was -40 mV. Ih was inhibited by Cs+ (1-5 mM) and by organic blockers N-ethyl-1,6-dihydro-1, 2-dimethyl-6 (methylimino)-N-phenyl-4-pyrimidinamine (ZD 7288; 0.3-3 microM) and N,N'-bis-(3,4 dimethylphenylethyl)-N-methylamine (YS 035, 3-30 microM), but not by Ba2+ (0.5 mM). The organic Ih blockers did not change the inspiratory bursts recorded from the XIIth nerve and synaptic drives in inspiratory neurons. Hypoxia reversibly inhibited Ih but, in the presence of organic blockers, the hypoxic reaction remained unchanged. We conclude that although Ih channels are functional in a minority of inspiratory neurons, Ih does not contribute to respiratory rhythm generation or its modulation by hypoxia. PMID- 10712630 TI - NO-mediated cGMP synthesis in cholinergic neurons in the rat forebrain: effects of lesioning dopaminergic or serotonergic pathways on nNOS and cGMP synthesis. AB - Nitric oxide synthase (NOS) activity and NO-mediated cGMP synthesis were studied in the rat forebrain of control animals and animals which had received a unilateral lesioning of dopaminergic or serotonergic pathways. Lesioning of the dopaminergic innervation using 6-hydroxydopamine resulted in a 50% decrease in NOS activity in the lesioned frontal cortex and caudate putamen. Lesioning of the serotonergic innervation using 5,7-dihydroxytryptamine had no effect on NOS activity. NO-mediated cGMP accumulation in rat forebrain slices was not affected by 6-hydroxydopamine or 5,7, -dihydroxytryptamine lesioning. Using cGMP immunocytochemistry, it was demonstrated that NO-mediated cGMP synthesis was absent from dopaminergic, serotonergic, GABA-ergic and neuronal NOS-containing nerve fibres. A minor colocalization of cGMP immunoreactivity was found in parvalbumin-containing fibres in the cortex. Extensive colocalization between cGMP immunoreactivity and the acetylcholine transporter was found in all cortical areas and in the caudate putamen. There was no effect of the lesions on this colocalization. These results demonstrate NO-mediated cGMP accumulation in cholinergic fibres in the forebrain of the rat and suggest an anterograde signalling function of NO in cholinergic neuronal systems in the cortex and caudate putamen of the rat. PMID- 10712632 TI - Reduction of stress-induced analgesia but not of exogenous opioid effects in mice lacking CB1 receptors. AB - CB1 cannabinoid receptors are widely distributed in the central nervous system where they mediate most of the cannabinoid-induced responses. Here we have evaluated the interactions between the CB1 cannabinoid receptors and the endogenous opioid system by assaying a number of well-characterized opioid responses, e.g. antinociception and stress-mediated effects, on mutant mice in which the CB1 receptor gene was invalidated. The spontaneous responses to various nociceptive stimuli (thermal, mechanical and visceral pain) were not changed in mutant CB1 mice. Furthermore, the absence of the CB1 cannabinoid receptor did not modify the antinociceptive effects induced by different opioid agonists: morphine (preferential mu opioid agonist), D-Pen2-D-Pen5-enkephalin (DPDPE) and deltorphin II (selective delta opioid agonists), and U-50,488H (selective kappa opioid agonist) in the hot-plate and tail-immersion tests. In contrast, the stress induced opioid mediated responses were modified in CB1 mutants. Indeed, these mutants did not exhibit antinociception following a forced swim in water at 34 degrees C and presented a decrease in the immobility induced by the previous exposure to electric footshock. However, the antinociception induced by a forced swim in water at 10 degrees C was preserved in CB1 mutants. These results indicate that CB1 receptors are not involved in the antinociceptive responses to exogenous opioids, but that a physiological interaction between the opioid and cannabinoid systems is necessary to allow the development of opioid-mediated responses to stress. PMID- 10712634 TI - Responses to linear and logarithmic frequency-modulated sweeps in ferret primary auditory cortex. AB - Multi-unit responses to frequency-modulated (FM) sweeps were studied in the primary auditory cortex of ferrets using six different stimulation paradigms. In particular, the differences between the responses to linear FM sweeps (where frequency changes linearly with time) and logarithmic FM sweeps (where frequency changes exponentially with time) were emphasized. Some general features of the responses to FM sweeps are independent of the exact details of the frequency trajectory. Both for linear and for logarithmic FM sweeps, a short burst of spikes occurred when the sweep reached a triggering frequency close to the best frequency of the cluster. The neuronal preference for FM velocity was also independent of frequency trajectory. Thus, clusters that responded best to slow logarithmic FM also preferred slow linear FM and vice versa. Consequently, topographic distributions of velocity preference were roughly independent of the stimulation paradigm. Other characteristics of the responses, however, depended on the exact details of the frequency trajectory. A significant number of clusters showed large differences in directional sensitivity between linear and logarithmic FM sweeps; these differences depended on the velocity preference of the clusters in some paradigms but not in others. Consequently, topographic distributions of directional sensitivity differed between linear and logarithmic paradigms. In conclusion, some characteristics of cluster responses to FM sweeps depend on the exact details of the stimulation paradigm and are not 'invariants' of the cluster. PMID- 10712633 TI - Overexpression of spermidine/spermine N-acetyltransferase in transgenic mice protects the animals from kainate-induced toxicity. AB - We recently generated a transgenic mouse line with activated polyamine catabolism through overexpression of spermidine/spermine N1-acetyltransferase (SSAT). A detailed analysis of brain polyamine concentrations indicated that all brain regions of these animals showed distinct signs of activated polyamine catabolism, e.g. overaccumulation of putrescine (three- to 17-fold), appearance of N1 acetylspermidine and decreases in spermidine concentrations. In situ hybridization analyses revealed a marked overexpression of SSAT-specific mRNA all over the brain tissue of the transgenic animals. The transgenic animals appeared to tolerate subcutaneous injections of high-dose kainate substantially better as their overall mortality was less than 50% of that of their syngenic littermates. We used the expression of glial fibrillary acidic protein (GFAP) as a marker of brain injury in response to kainate. In situ hybridization analysis with GFAP oligonucleotide up to 7 days after the administration of sublethal kainate doses showed reduced GFAP expression in transgenic animals in comparison with their non transgenic littermates. This difference was especially striking in the cerebral cortex of the transgenic mice where the exposure to kainate hardly induced GFAP expression. The treatment with kainate likewise resulted in loss of the hippocampal (CA3) neurons in non-transgenic but not transgenic animals. These results support our earlier findings indicating that elevated concentrations of brain putrescine, irrespective whether derived from an overexpression of ornithine decarboxylase, or as shown here, from an overexpression of SSAT, play in all likelihood a neuroprotective role in brain injury. PMID- 10712635 TI - Clustering of delta glutamate receptors is regulated by the actin cytoskeleton in the dendritic spines of cultured rat Purkinje cells. AB - The interaction of neurotransmitter receptors with the cytoskeleton is an important mechanism for the targeting of receptors to the postsynaptic membrane. Using cytoskeleton-perturbing agents, it was demonstrated that delta glutamate receptors, predominantly expressed on the dendritic spines of cerebellar Purkinje cells, are anchored to the actin cytoskeleton. The number of delta glutamate receptor-immunoreactive clusters was dramatically decreased following treatment of the Purkinje cells with the actin-disrupting agents, cytochalasin D or latrunculin A, without any significant effect on the number of presynaptic contacts of the granule cell axons. The clusters disrupted by latrunculin A were re-established 24 h after removal of the drug. These results suggest that morphological changes in the actin cytoskeleton regulate the delta glutamate receptor clustering on the dendritic spines, and may affect synaptic efficacy and plasticity. PMID- 10712636 TI - Circadian modulation of calcium levels in cells in the suprachiasmatic nucleus. AB - There is reason to believe that resting free calcium concentration [Ca2+]i in neurons in the suprachiasmatic nucleus (SCN) may vary with the circadian cycle. In order to start to examine this hypothesis, optical techniques were utilized to estimate resting Ca2+ levels in SCN cells in a rat brain slice preparation. [Ca2+]i measured from the soma was significantly higher in the day than in the night. Animals from a reversed light-dark cycle were used to confirm that the phase of the rhythm was determined by the prior light-dark cycle. The rhythm in Ca2+ levels continued to be expressed in tissue collected from animals maintained in constant darkness, thus confirming the endogenous nature of this variation. Interestingly, the rhythm in Ca2+ levels was not observed when animals were housed in constant light. Finally, the rhythm in Ca2+ levels was prevented when slices were exposed to tetrodotoxin (TTX), a blocker of voltage-sensitive sodium channels. Similar results were obtained with the voltage-sensitive Ca2+ channel blocker methoxyverapamil. These observations suggest a critical role for membrane events in driving the observed rhythm in Ca2+. Conceptually, this rhythm can be thought of as an output of the circadian oscillator. Because [Ca2+]i is known to play a critical role in many cellular processes, the presence of this rhythm is likely to have many implications for the cell biology of SCN neurons. PMID- 10712637 TI - Specific activation of the mu opioid receptor (MOR) by endomorphin 1 and endomorphin 2. AB - The recently discovered endomorphin 1 (Tyr-Pro-Trp-Phe-NH2) and endomorphin 2 (Tyr-Pro-Phe-Phe-NH2) were investigated with respect to their direct receptor binding properties, and to their ability to activate G proteins and to inhibit adenylyl cyclase in both cellular and animal models. Both tetrapeptides activated G proteins and inhibited adenylyl cyclase activity in membrane preparations from cells stably expressing the mu opioid receptor, an effect reversed by the mu receptor antagonist CTAP (D-Phe-Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH2), but they had no influence on cells stably expressing the delta opioid receptor. To further establish the selectivity of these peptides for the mu opioid receptor, brain preparations of mice lacking the mu opioid receptor gene were used to study their binding and signalling properties. Endomorphin 2, tritiated by a dehalotritiation method resulting in a specific radioactivity of 1.98 TBq/mmol (53.4 Ci/mmol), labelled the brain membranes of wild-type mice with a Kd value of 1.77 nM and a Bmax of 63.33 fmol/mg protein. In membranes of mice lacking the mu receptor gene, no binding was observed, and both endomorphins failed to stimulate [35S]guanosine 5'-O-(3-thio)triphosphate ([35S]GTPgammaS) binding and to inhibit adenylyl cyclase. These data show that endomorphins are capable of activating G proteins and inhibiting adenylyl cyclase activity, and all these effects are mediated by the mu opioid receptors. PMID- 10712638 TI - Facilitation of cholinergic transmission by substance P methyl ester in the mouse hippocampal slice preparation. AB - Using sharp microelectrode recording from CA1 pyramidal neurons of the adult mouse hippocampal slice preparation, we studied the modulatory action of the selective neurokinin 1 (NK1) receptor agonist substance P methyl ester (SPME), a peptidase-resistant analogue of the peptide substance P (SP), on cholinergic responses. While SPME (0.1-1 microM) had only slight effects on membrane potential and input resistance of CA1 neurons, it largely and reversibly enhanced the membrane depolarization and oscillatory activity induced by the cholinergic agonist carbachol (CCh; 0.1-100 microM). This effect of SPME was prevented by the selective NK1 receptor antagonist SR 140333 (4 microM). In about half of the tested neurons the action of SPME was preserved in tetrodotoxin (TTX) solution, suggesting that it partly occurred at the level of pyramidal cells. Cholinergic slow excitatory postsynaptic potentials (sEPSPs) were reversibly enhanced by SPME which increased their amplitude and prolonged any associated bursting activity. This action was also blocked by SR 140333. The present results suggest that SPME largely enhances cholinergic activity in the mouse hippocampus, an effect which can help to explain, in this brain area, the recently reported facilitation of seizures by SP. PMID- 10712639 TI - Overexpression of neuropeptide Y induced by brain-derived neurotrophic factor in the rat hippocampus is long lasting. AB - Brain-derived neurotrophic factor (BDNF) plays an important role in hippocampal neuroplasticity. In particular, BDNF upregulation in the hippocampus by epileptic seizures suggests its involvement in the neuronal rearrangements accompanying epileptogenesis. We have shown previously that chronic infusion of BDNF in the hippocampus induces a long-term delay in hippocampal kindling progression. Although BDNF has been shown to enhance the excitability of this structure upon acute application, long-term transcriptional regulations leading to increased inhibition within the hippocampus may account for its suppressive effects on epileptogenesis. Therefore, the long-term consequences of a 7-day chronic intrahippocampal infusion of BDNF (12 microg/day) were investigated up to 2 weeks after the end of the infusion, on the expression of neurotransmitters contained in inhibitory hippocampal interneurons and which display anti-epileptic properties. Our results show that BDNF does not modify levels of immunostaining for glutamic acid decarboxylase, the rate-limiting enzyme for gamma-aminobutyric acid (GABA) synthesis, and somatostatin. Conversely, BDNF induces a long-lasting increase of neuropeptide Y (NPY) in the hippocampus, measured by immunohistochemistry and radioimmunoassay, outlasting the end of the infusion by at least 7 days. The distribution of BDNF-induced neuropeptide Y immunoreactivity is similar to the pattern observed in animals submitted to hippocampal kindling, with the exception of mossy fibres which only become immunoreactive following seizure activity. The enduring increase of neuropeptide Y expression induced by BDNF in the hippocampus suggests that this neurotrophin can trigger long-term genomic effects, which may contribute to the neuroplasticity of this structure, in particular during epileptogenesis. PMID- 10712640 TI - Inositol 1,4,5-trisphosphate and adenophostin analogues induce responses in turtle olfactory sensory neurons. AB - Using the whole-cell mode of the patch-clamp technique, we recorded inward currents in response to inositol-1,4,5-trisphosphate (IP3) and adenophostin analogues in turtle olfactory sensory neurons. Dialysis of IP3 into the neurons induced inward currents with an increase in membrane conductance in a dose dependent manner under the voltage-clamp conditions (holding potential -70 mV). The application of Ca2+-free Ringer solution to neurons previously dialysed with IP3 induced inward currents that were reversibly inhibited by application of Na+, Ca2+-free Ringer solution, normal Ringer solution or 10 microM ruthenium red. Dialysis of the adenophostin analogues, novel IP3 receptor ligands, also induced inward currents with an increase in membrane conductance. The magnitude of the responses to the adenophostin analogues varied among these derivatives. The application of Ca2+-free Ringer solution to neurons previously dialysed with the adenophostin analogues induced inward currents that were inhibited by the application of normal Ringer solution. The reversal potential of inward currents induced by an adenophostin analogue was similar to that induced by IP3, suggesting that inward currents induced by the adenophostin analogue were generated by a similar ionic mechanism to that induced by IP3. The present study demonstrated that IP3-mediated transduction pathways exist in turtle olfactory receptor neurons and that adenophostin analogues act as agonists of IP3. PMID- 10712641 TI - Retinal ganglion cells with NADPH-diaphorase activity in the chick form a regular mosaic with a strong dorsoventral asymmetry that can be modelled by a minimal spacing rule. AB - We have identified a class of retinal ganglion cells in the chick retina that can be labelled by NADPH-diaphorase histochemistry. These cells have a remarkable topographic distribution, being restricted to the dorsal hemiretina, and form a highly regular mosaic, as revealed by the analysis of nearest neighbour distribution and Delaunay triangulation. Autocorrelation analysis of the mosaic of NADPH-diaphorase-positive retinal ganglion cells shows that the mosaic spatial organization could be generated with the single constraint that two elements cannot be closer than a given minimal distance (d(min)), which we confirmed by computer simulations. In contrast with what has been observed in other mosaics, here d(min) varies with cell density. However, the observed variation of the exclusion area is consistent with an original assembly of the mosaic with a constant d(min) (as is the case in other mosaics), followed by differential expansion of the retina during development. PMID- 10712643 TI - Dual effect of ecdysone on adult cricket mushroom bodies. AB - Mushroom bodies, which are the main integrative centre for insect sensorial information, play a critical role in associative olfactory learning and memory. This paired brain structure contains interneurons grouped in a cortex, sending their axons into organized neuropiles. In the house cricket (Acheta domesticus) brain, persistent neuroblasts proliferate throughout adult life. Juvenile hormone (JH) has been shown to stimulate this proliferation [Cayre, M., Strambi, C. & Strambi, A. (1994) Nature, 368, 57-59]. In the present study, the effect of morphogenetic hormones on mushroom body cells maintained in primary culture was examined. Whereas JH did not significantly affect neurite growth, ecdysone significantly stimulated neurite elongation. Moreover, ecdysone also acted on neuroblast proliferation, as demonstrated by the reduced number of cells labelled with 5-bromodeoxyuridine following ecdysone application. Heterospecific antibodies raised against ecdysone receptor protein and ultraspiracle protein, the two heterodimers of ecdysteroid receptors, showed positive immunoreactivity in nervous tissue extracts and in nuclei of mushroom body cells, indicating the occurrence of putative ecdysteroid receptors in cricket mushroom body cells. These data indicate a dual role for ecdysone in adult cricket mushroom bodies: this hormone inhibits neuroblast proliferation and stimulates interneuron differentiation. These results suggest that a constant remodelling of mushroom body structure could result from physiological changes in hormone titres during adult life. PMID- 10712642 TI - Spatial, temporal and subcellular localization of islet-brain 1 (IB1), a homologue of JIP-1, in mouse brain. AB - Islet-brain 1 (IB1) was recently identified as a DNA-binding protein of the GLUT2 gene promoter. The mouse IB1 is the rat and human homologue of the Jun interacting protein 1 (JIP-1) which has been recognized as a key player in the regulation of c-Jun amino-terminal kinase (JNK) mitogen-activated protein kinase (MAPK) pathways. JIP-1 is involved in the control of apoptosis and may play a role in brain development and aging. Here, IB1 was studied in adult and developing mouse brain tissue by in situ hybridization, Northern and Western blot analysis at cellular and subcellular levels, as well as by immunocytochemistry in brain sections and cell cultures. IB1 expression was localized in the synaptic regions of the olfactory bulb, retina, cerebral and cerebellar cortex and hippocampus in the adult mouse brain. IB1 was also detected in a restricted number of axons, as in the mossy fibres from dentate gyrus in the hippocampus, and was found in soma, dendrites and axons of cerebellar Purkinje cells. After birth, IB1 expression peaks at postnatal day 15. IB1 was located in axonal and dendritic growth cones in primary telencephalon cells. By biochemical and subcellular fractionation of neuronal cells, IB1 was detected both in the cytosolic and membrane fractions. Taken together with previous data, the restricted neuronal expression of IB1 in developing and adult brain and its prominent localization in synapses suggest that the protein may be critical for cell signalling in developing and mature nerve terminals. PMID- 10712644 TI - The seasonal pattern of cell proliferation and neuron number in the dentate gyrus of wild adult eastern grey squirrels. AB - The dentate gyrus is one of two areas in the mammalian brain that produces neurons in adulthood. Neurogenesis (proliferation, survival, and differentiation of new neurons) is regulated by experience, and increased neurogenesis appears to be correlated with improved spatial learning in mammals and birds. We tested the hypothesis that in long-lived mammals that scatter-hoard food, seasonal variations in spatial memory processing (i.e. increased processing during caching season in the autumn) might correlate with changes in neurogenesis and neuron number in the granule cell layer of the dentate gyrus (gcl DG). We investigated the rate of cell proliferation and the total number of neurons in the granule cell layer of wild adult eastern grey squirrels (Sciurus carolinensis) at three different times of the year (October, January and June). We found no seasonal differences in cell proliferation rate or in total neuron number in the granule cell layer. Our findings are in agreement with those of previous studies in laboratory mice and rats, and in free-ranging, food-caching, black-capped chickadees, as well as with current hypotheses regarding the relationship between neurogenesis and learning. Our results, however, are also in agreement with the hypothesis that neurogenesis in the dentate gyrus represents a maintenance system that may be regulated by environmental factors, and that changes in total neuron number previously reported in rodents represent developmental changes rather than adult plasticity. The patterns observed in mature wild rodents, such as free ranging squirrels, may represent more accurately the extent of hippocampal plasticity in adult mammals. PMID- 10712645 TI - Endothelial and neuronal nitric oxide synthases are present in the suprachiasmatic nuclei of Syrian hamsters and rats. AB - Nitric oxide (NO) is involved in the transmission of light information to suprachiasmatic nuclei (SCN). By immunocytochemistry, we showed that both neuronal and endothelial NO synthase isoforms (nNOS and eNOS) were present in the SCN of rats and hamsters. nNOS-immunoreactive neurons were located mainly around the SCN with only a few nNOS neurons within the nucleus. By double-label immunocytochemistry, we also found, within the population of SCN glial fibrillary acidic protein (GFAP)-immunoreactive astrocytes, a subpopulation of eNOS immunoreactive astrocytes. Using Western blot analysis, we detected in SCN protein extracts eNOS and nNOS proteins having the expected 140 and 150 kDa molecular weights, respectively. By in situ hybridization of a 2.4-kb murine eNOS probe, mRNA for eNOS was located in the SCN of rats and hamsters. The transcript was further identified by detection of a RT-PCR product of the predicted size, after amplification of total RNA with primers specific for eNOS. In the SCN and cerebellum, the size of the mRNA for nNOS, detected with a rat probe on Northern blot, was approximately 10.5 kb, corresponding to that previously published. In the same tissues, we found two transcripts, one weakly expressed at approximately 4.0 kb and another more strongly expressed at approximately 2.6 kb, both hybridizing with two non-overlapping murine and rat eNOS probes. These results suggested the existence in the SCN of alternate transcripts for eNOS. We propose that two pathways could link light stimuli and NO release in the SCN: one involving N-methyl-D-aspartate (NMDA) receptors and nNOS in neurons; the other linking alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors and eNOS in astrocytes. PMID- 10712646 TI - Afferent-specific modulation of short-term synaptic plasticity by neurotrophins in dentate gyrus. AB - Neurotrophins modulate synaptic transmission and plasticity in the adult brain. We here show a novel feature of this synaptic modulation, i.e. that two populations of excitatory synaptic connections to granule cells in the dentate gyrus, lateral perforant path (LPP) and medial perforant path (MPP), are differentially influenced by the neurotrophins BDNF and NT-3. Using field recordings and whole-cell patch-clamp recordings in hippocampal slices, we found that paired-pulse (PP) depression at MPP-granule cell synapses was impaired in BDNF knock-out (+/-) mice, but PP facilitation at LPP synapses to the same cells was not impaired. In accordance, scavenging of endogenous BDNF with TrkB-IgG fusion protein also impaired PP depression at MPP-granule cell synapses, but not PP facilitation at LPP-granule cell synapses. Conversely, in NT-3+/- mice, PP facilitation was impaired at LPP-granule cell synapses whilst PP depression at MPP-granule cell synapses was unaffected. These deficits could be reversed by application of exogenous neurotrophins in an afferent-specific manner. Our data suggest that BDNF and NT-3 differentially regulate the synaptic impact of different afferent inputs onto single target neurons in the CNS. PMID- 10712647 TI - In vivo survival requirement of a subset of nodose ganglion neurons for nerve growth factor. AB - The sensory neurons of the nodose ganglion are the classic example of a population of peripheral nervous system neurons that do not require nerve growth factor (NGF) for survival during development but are dependent on other neurotrophins. We have re-examined this assertion by studying the development of the nodose ganglion of mice that have a null mutation in the NGF gene. Compared with wild-type embryos, the number of neurons undergoing apoptosis was elevated in NGF -/- mice, resulting in a significant reduction in the total number of neurons in the ganglion by the end of embryonic development. TrkA, the NGF receptor tyrosine kinase, was expressed in the nodose ganglion throughout development and there was a marked decrease in TrkA mRNA expression in the nodose ganglion of NGF -/- embryos. Although the in vitro survival of the majority of nodose neurons was promoted by brain-derived neurotrophic factor (BDNF), a minor proportion was supported by NGF in cultures established over a range of embryonic stages. These results clearly demonstrate that a subset of nodose ganglion neurons depends on NGF for survival during development. The finding that the expression of tyrosine hydroxylase (TH) mRNA was unaffected in the nodose ganglia of NGF-deficient embryos indicates that this NGF-dependent subset is distinct from the subset of catacholaminergic neurons in the nodose ganglion. PMID- 10712648 TI - Hippocampal stem cells differentiate into excitatory and inhibitory neurons. AB - Stem cell technology promises new and rapid advances in cell therapy and drug discovery. Clearly, the value of this approach will be limited by the differentiated functions displayed by the progeny of stem cells. The foetal and adult central nervous system (CNS) harbour stem cells that can be expanded in vitro and differentiate into immature neurons and glia. Surprisingly, we do not know if neurons derived from stem cells form synapses, a definitive feature of neuronal function. Neuronal differentiation is a complex process and in this paper we establish conditions that permit extensive maturation of neurons in the presence of neurotrophins. These conditions permit the differentiation of rat hippocampal stem cells into both excitatory (glutamatergic) and inhibitory (GABAergic) neurons. The proportion of excitatory and inhibitory synapses was strongly influenced by specific neurotrophins, and these responses reflect the region of origin of the stem cells in the brain. These data show that stem cells can be used to study mechanisms of excitation and inhibition in the nervous system. PMID- 10712649 TI - Neurokinin 1 receptor expression by neurons in laminae I, III and IV of the rat spinal dorsal horn that project to the brainstem. AB - Large neurons in laminae III and IV of the spinal cord which express the neurokinin 1 receptor and have dendrites that enter the superficial laminae are a major target for substance P (SP)-containing (nociceptive) primary afferents. Although some of these neurons project to the thalamus, we know little about other possible projection targets. The main aim of this study was to determine whether all cells of this type are projection neurons and to provide information about brainstem sites to which they project. Injections of cholera toxin B subunit were made into four brainstem areas that receive input from the spinal cord, and the proportion of cells of this type in the L4 spinal segment that were retrogradely labelled was determined in each case. The results suggest that most of these cells (>90%) project to the contralateral lateral reticular nucleus (or to a nearby region), while many (>60%) send axons to the lateral parabrachial area and some to the dorsal part of the caudal medulla. However, few of these cells project to the periaqueductal grey matter. As lamina I neurons with the neurokinin 1 receptor appear to be important in the generation of hyperalgesia, we also examined projection neurons in this lamina and found that for each injection site the great majority possessed the receptor. These results demonstrate that dorsal horn neurons which express the neurokinin 1 receptor contribute to several ascending pathways that are thought to be important in pain mechanisms. PMID- 10712650 TI - Effects of monoamines on interneurons in four spinal reflex pathways from group I and/or group II muscle afferents. AB - Effects of locally applied serotonin (5-HT) and noradrenaline (NA) were tested on extracellularly recorded responses of single spinal interneurons in deeply anaesthetized cats. These effects were tested on: (i) interneurons mediating reciprocal inhibition from group Ia afferents; (ii) interneurons mediating non reciprocal inhibition from group Ia and Ib afferents; (iii) intermediate zone interneurons co-excited by group I and II afferents; and (iv) dorsal horn interneurons excited by group II afferents. Effects of monoamines were tested on responses evoked at latencies compatible with monosynaptic coupling. Responses evoked by group Ia and/or Ib muscle afferents were facilitated in all of the tested interneurons both by NA and 5-HT. Responses evoked by group II muscle afferents were depressed in the majority of the interneurons but were facilitated in some of them. 5-HT depressed these responses in all dorsal horn interneurons and in one subpopulation of intermediate zone interneurons, while it facilitated them in another subpopulation of intermediate zone interneurons. NA depressed them in all intermediate zone interneurons and in one subpopulation of dorsal horn interneurons, while it facilitated them in another subpopulation of dorsal horn interneurons. The results of this study lead to the conclusions that: (i) modulation of synaptic actions of muscle spindle and tendon organ afferents on spinal interneurons by 5-HT and NA is related to both the type of the afferent and the functional type of the interneuron; and that (ii) 5-HT and NA counteract each others' actions on some interneuronal types but mutually enhance them on the others. PMID- 10712651 TI - Cerebral oligaemia episode triggers free radical formation and late cognitive deficiencies. AB - Sixty minutes of cerebral oligaemic hypoxia, induced by bilateral clamping of the carotid arteries (BCCA) in pentobarbital-anaesthetized normotensive rats, induces a late progressive cognitive decline when compared with sham-operated controls. Analysis at BCCA of hippocampal metabolism using microdialysis showed increased release of glutamate, aspartate and gamma-aminobutyric acid, followed by a progressive rise in the formation of hydroxyl free radicals measured as 2,3 dihydroxybenzoic acid (2,3-DHBA), their reaction product with salicylate, though only in the re-perfusion phase. In the striatum increased dopamine release occurred during BCCA, whereas glutamate and aspartate showed an increase only during the late re-perfusion phase. gamma-Aminobutyric acid (GABA) concentration increased during BCCA and early re-perfusion. An increase in 2,3-DHBA was seen during BCCA, and persisted over 2 h of re-perfusion. Six and 13 months after surgery, though not as early as 3 months, BCCA-treated rats perform worse than sham-operated controls in a water-maze, where decreased swimming speed reveals striatal dysfunction, while hippocampal dysfunction manifested as diminished spatial bias. These results show that cerebral oligaemia, similarly to cerebral ischaemia, leads to increased extracellular dopamine, aspartate and glutamate, and the production of hydroxyl radicals in structures associated with learning and memory processes. Unlike cerebral ischaemia, in cerebral oligaemia the appearance of spatial memory deficits is delayed. PMID- 10712652 TI - The novel brain neuropeptide, orexin-A, modulates the sleep-wake cycle of rats. AB - Orexin-A is a novel neuropeptide initially isolated from hypothalamic extracts but now known to be present in fibres distributed throughout the rat CNS including many regions associated with sleep-wake regulation. The recognition of a particularly dense innervation of orexinergic nerves in the locus coeruleus, together with the observed increase in firing rate of locus coeruleus neurons following application of orexin-A in vitro, further highlighted a potential involvement of the peptide in modulating the arousal state. The present study was undertaken to determine the effects of intracerebroventricularly (ICV) administered orexin-A on the sleep-wake cycle of conscious rats using electroencephalographic and electromyographic recordings. When administered at the onset of the normal sleep period, orexin-A (1, 10 or 30 microg/rat ICV) produced a dose-dependent increase in the time rats spent awake during the second and third hours after dosing. The enhancement of arousal was accompanied by a marked reduction in paradoxical sleep and deep slow wave sleep at the highest dose. The latency to the first occurrence of paradoxical sleep was also prolonged. This overall profile of increased arousal and decreased paradoxical sleep is consistent with a high rate of firing of locus coeruleus neurons as would be expected to occur following ICV administration of orexin-A. It is concluded that orexin-A may play an important physiological role in regulating the sleep-wake state, a hypothesis considerably strengthened by the recently reported narcoleptic phenotype of prepro-orexin (the precursor for orexin-A) knockout mice. PMID- 10712653 TI - Is right hemisphere specialization for face discrimination specific to humans? AB - Patterns of neural activation during face recognition were investigated in sheep by quantifying altered c-fos mRNA expression in situations where faces (sheep vs. human) can (faces upright) and cannot (faces inverted) be discriminated. Exposure to upright faces selectively increased expression significantly more in the right inferior temporal cortex than in the left, and active choice between upright faces additionally increased expression bilaterally in basal amygdala and hippocampus (CA1-4). Exposure to inverted faces did not lead to enhanced activation in the right inferior temporal cortex, amygdala or hippocampus but instead increased expression levels in the diagonal band of Broca, parietal and cingulate cortices. These results show that discrimination of upright faces in sheep preferentially engages the right temporal cortex, as it does in humans, and that performance of active choices between such faces may additionally involve the basal amygdala and hippocampus. PMID- 10712654 TI - Hippocampal participation in navigational map learning in young homing pigeons is dependent on training experience. AB - The homing pigeon navigational map is perhaps one of the most striking examples of a naturally occurring spatial representation of the environment used to guide navigation. In a previous study, it was found that hippocampal lesions thoroughly disrupt the ability of young homing pigeons held in an outdoor aviary to learn a navigational map. However, since that study an accumulation of anecdotal data has hinted that hippocampal-lesioned young pigeons allowed to fly during their first summer could learn a navigational map. In the present study, young control and hippocampal-lesioned homing pigeons were either held in an outdoor aviary or allowed to fly during the time of navigational map learning. At the end of their first summer, the birds were experimentally released to test for navigational map learning. Independent of training experience, control pigeons oriented homeward during the experimental releases demonstrating that they learned a navigational map. Surprisingly, while the aviary-held hippocampal-lesioned pigeons failed to learn a navigational map as reported previously, hippocampal-lesioned birds allowed flight experience learned a navigational map indistinguishable from the two control groups. A subsequent experiment revealed that the navigational map learned by the three groups was based on atmospheric odours. The results demonstrate that hippocampal participation in navigational map learning depends on the type of experience a young bird pigeon has, and presumably, the type of navigational map learned. PMID- 10712655 TI - Eye position-sensitive units in hippocampal formation and in inferotemporal cortex of the macaque monkey. AB - The activity of 330 hippocampal and inferotemporal cells was recorded while seated monkeys with fixed heads worked in a visual discrimination task. Monkeys had to move their eyes to one of five different positions to maintain gaze on an image. The image was then extinguished and the monkeys maintained a fixed gaze on the target position in darkness to obtain a reward. The five positions of image presentation were on a horizontal line, consisting of a centre position and lateral positions which were 10 and 20 degrees right and left of it. Twenty-two per cent of single units recorded from the hippocampus showed statistically significant sensitivity to target position in complete darkness. A similar fraction (23%) was significantly affected by target position in the light. Position sensitivity was also found among cells recorded from the inferotemporal cortex. Eye position significantly influenced the activity of 19% of inferotemporal units in darkness and 28% of inferotemporal units in the light. Interestingly, the populations of cells showing position effect in the light and in darkness were independent. PMID- 10712656 TI - Oxytocin induces preservation of social recognition in male rats by activating alpha-adrenoceptors of the olfactory bulb. AB - In this report, a series of four experiments was performed to evaluate the relationship between the olfactory bulb norepinephrine system and intra-olfactory bulb infusion of oxytocin in the preservation of social memory responses. The present data indicate that oxytocin exerts this preservation of social recognition through a specific, receptor-mediated mechanism within the olfactory bulb (experiment 1). The involvement of the olfactory bulb norepinephrine system is revealed by the demonstration that retrodialysis of oxytocin into the olfactory bulb increases norepinephrine release (experiment 4). Our data suggest that the increased output of olfactory bulb norepinephrine resulting from oxytocin appears to activate alpha-adrenoceptors to produce this preservation in recognition because infusions of clonidine into the olfactory bulb preserve recognition responses in a manner similar to that observed with oxytocin (experiment 2). In addition, a co-infusion of oxytocin with phentolamine abolishes recognition responses (experiment 3). Accordingly, this model affords the opportunity to study neuropeptide-catecholamine interactions, link these interactions with a specific behavioural outcome and identify a novel function/site of action for oxytocin in the male. PMID- 10712657 TI - Does the migraine aura reflect cortical organization? AB - Individuals suffering from classical migraine report an astonishing diversity of migraine auras. A frequently reported symptom is a visual hallucination known as fortification illusion (FI). Here we demonstrate that the typical zig-zag pattern of the FI can be reproduced using experimental data of orientation maps of the primary visual cortex (V1) assuming that a continuous excitation front propagates across V1. We put forward a model in which the cortical neurons within this excitation wave are activated sufficiently to contribute to conscious perception. It is shown that the discontinuous repetitive nature of the zig-zag pattern of the FI can reflect the specific layout of visual cortical orientation maps. Additionally, dynamic features of the FI are predicted based on our model. PMID- 10712658 TI - Basal expression of c-Fos and Zif268 in the rat basal ganglia: immunohistochemical characterization of striatal Zif268-positive neurons. AB - Basal expression of the protein products of the inducible immediate early genes (IEGs), c-Fos and Zif268, was investigated in five regions of the rat basal ganglia using immunohistochemistry. In particular, high basal levels of Zif268 but very low levels of c-Fos were seen in the caudate-putamen (CPu). Double immunostaining revealed that many of the constitutively expressed Zif268-positive neurons were GABAergic but very few were cholinergic or neuronal nitric oxide synthase (nNOS)-positive, and some of the Zif268-positive neurons were also immunopositive for a glutamate NMDA receptor subunit NR1 or NR2A. No regional difference between the medial and lateral parts of the CPu was observed in the cellular phenotypes of Zif268-positive neurons. Almost no basal levels of Zif268 were seen in the other four regions: the globus pallidus, entopeduncular nucleus, subthalamic nucleus and substantia nigra pars reticulata. As in the CPu, negligible levels of c-Fos were seen in these four regions. Differential expression of these two IEGs may suggest gene-specific and region-specific functions of c-Fos and Zif268 in the basal ganglia. Constitutive expression of Zif268 existing mainly in the GABAergic neurons in the CPu may at least in part be maintained by glutamatergic afferents. PMID- 10712659 TI - Survival effects of BDNF and NT-3 on axotomized rubrospinal neurons depend on the temporal pattern of neurotrophin administration. AB - This study shows that both BDNF and NT-3 can prevent cell death in axotomized adult rat rubrospinal neurons (RSNs), but that the efficacy of neuroprotection depends on the temporal pattern of treatment. At 8 weeks after cervical spinal cord injury, 51% of the RSNs had died. Subarachnoidal BDNF infusion into the cisterna magna for 4 weeks resulted in neuronal hypertrophy and 71% survival. Continuous infusion for 8 weeks into the lumbar subarachnoidal space with either BDNF or NT-3 gave similar survival rates, while a combination of BDNF and NT-3 resulted in 96% survival, although the cells were atrophic. When administration of either BDNF or NT-3 was delayed and performed during postoperative weeks 5-8, the number of surviving neurons was increased compared to early treatment. Delayed treatment with a combination of BDNF and NT-3 resulted in complete survival and a reduction in neuronal atrophy. A decreased expression of TrkB receptors and microtubule-associated protein-2 in the RSNs after axotomy was counteracted by BDNF and NT-3. Microglial activity remained increased even when complete cell survival was achieved. Thus, the combination of neurotrophins as well as the temporal pattern of treatment need to be adequately defined to optimize survival of injured spinal tract neurons. PMID- 10712660 TI - Editorial PMID- 10712661 TI - Heat shock proteins refine the danger theory. PMID- 10712662 TI - The route of administration of an immunodominant peptide derived from heat-shock protein 65 dramatically affects disease outcome in pristane-induced arthritis. AB - Previous studies have shown that immunization of mice with an immunodominant epitope from heat-shock protein 65 (hsp 65) (amino acids 261-271) can protect from the development of pristane-induced arthritis (PIA) and this protection is mediated by an antigen-specific T helper type 2 (Th2) cytokine response. Here we confirm these findings and show that frequent intranasal administration of this peptide exacerbates disease. In naive mice given peptide intranasally an antigen specific T-cell population is systemically activated similar to that induced by peptide immunization in incomplete Freund's adjuvant. Thus, a paradox exists whereby apparently similar peptide-specific populations are either associated with protection from, or exacerbation of, PIA. However, comparison of cytokine profiles reveals differences between these two cell populations. Peptide inhalation induces the production of Th1-type cytokines (interferon-gamma) whereas intraperitoneal immunization leads to the production of Th2-type cytokines (interleukin-4, interleukin-5 and interleukin-10) by splenic T cells upon stimulation with peptide. Thus, for the application of nasal 'tolerance' in clinical medicine, it is important to identify antigens and dosing regimes that counteract but do not activate adverse immune responses. PMID- 10712663 TI - T-cell tolerance induction is normal in the (NZB x NZW)F1 murine model of systemic lupus erythematosus. AB - The (New Zealand black (NZB) x New Zealand white (NZW))F1 (NZB/W) mouse strain spontaneously develops an autoimmune disease characterized by anti-dsDNA antibody production and glomerulonephritis. Although evidence suggests that production of pathogenic autoantibodies is T-cell dependent, the immunological defects that lead to activation of these autoreactive T cells are unknown. In particular, it has not been resolved whether autoreactive T cells become activated in these mice because of a generalized defect in T-cell tolerance induction. Previous work has demonstrated that thymic and peripheral tolerance to strongly deleting antigens are intact in NZB/W mice. In this study we investigate whether these mice possess a more subtle T-cell tolerance defect. To this end, we have produced NZB/W mice carrying a transgene encoding beef insulin (BI) which is expressed at levels close to the threshold for T-cell tolerance induction. In BALB/c mice this transgene produces a profound but incomplete state of BI-specific T-cell tolerance, mediated predominantly by clonal anergy. Comparison of BI-specific tolerance in NZB/W, major histocompatibility complex (MHC)-matched (BALB/c x NZW)F1, and BALB/c BI-transgenic mice clearly demonstrates that T-cell tolerance induction is normal in NZB/W mice. The data suggest that the loss of T-cell tolerance that ultimately supports nephritogenic autoantibody production in NZB/W mice does not result from a generalized defect in T-cell tolerance, and by extension likely results from aberrant activation of specific autoreactive T cells. PMID- 10712664 TI - Monoclonal anti-double-stranded DNA autoantibody stimulates the expression and release of IL-1beta, IL-6, IL-8, IL-10 and TNF-alpha from normal human mononuclear cells involving in the lupus pathogenesis. AB - In our previous reports, we found polyclonal anti-double-stranded DNA antibodies (anti-dsDNA) purified from patients with active systemic lupus erythematosus (SLE) exerted inhibitory effect on [3H]thymidine incorporation of human mononuclear cells (MNC). However, the other immunological effects of anti-dsDNA on the functions of MNC have not yet been reported. In this study, two monoclonal antibodies, 12B3 and 9D7, with different anti-dsDNA activity were evaluated for their effects on the expression and release of different cytokines from human MNC. We confirmed absence of endotoxin in the two monoclonal antibody preparations and the used medium as detected by Limulus amoebocyte lysate test. The mRNA expression and release of different cytokines including interleukin (IL) 1beta, IL-2, IL-4, IL-6, IL-8, IL-10, tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) were measured. We found the two monoclonal anti dsDNA not only dose-responsively suppressed the phytohaemagglutinin (PHA)-induced thymidine uptake of human MNC but stimulated the mRNA expression of IL-1beta, IL 6 and IL-8 in normal human MNC detected by reverse transcription-polymerase chain reaction (RT-PCR). Enzyme-linked immunosorbent assay (ELISA) measurement of cytokines in MNC culture supernatants revealed that anti-dsDNA enhanced IL-1beta, IL-8, TNF-alpha and IL-10 release from resting MNC. These effects of anti-dsDNA antibodies were not affected by polymyxin B, a potent binder and neutralizer of lipopolysaccharide (LPS). These in vitro studies suggest that anti-dsDNA possess a dual effect on normal human MNC: (a) to enhance the release of proinflammatory cytokines (IL-1beta, IL-8 and TNF-alpha) from MNC to augment inflammatory reaction; and (b) to polarize the immune reaction towards the T helper 2 (Th2) (increased IL-10 production) pathway. This unique effect of anti-dsDNA may play a role in lupus pathogenesis by augmenting inflammatory reactions and autoantibody production which are commonly found in patients with active SLE. PMID- 10712665 TI - The effects of DNA containing CpG motif on dendritic cells. AB - Dendritic cells (DC) are specialized antigen-presenting cells. DC can acquire and process antigens in the periphery before maturing and migrating to secondary lymphoid tissues where they present the antigens and deliver co-stimulatory signals to T cells. We describe an immunostimulatory oligonucleotide containing a CpG motif that stimulated murine DC to up-regulate co-stimulatory molecules, induce T-cell proliferative responses and secrete interleukin-12 in vitro. Administration of this oligonucleotide, but not of a control oligonucleotide lacking this motif, to mice led to the disappearance of DC from the marginal zone and T-cell areas of spleen, but not from heart or kidney. The same CpG did not cause maturation of monocyte-derived human DC in vitro, but lipopolysaccharide treated monocyte-derived DC showed enhanced functional activity and up-regulated co-stimulatory molecules. PMID- 10712666 TI - Effective formation of major histocompatibility complex class II-peptide complexes from endogenous antigen by thyroid epithelial cells. AB - In autoimmune thyroid disease, thyroid epithelial cells (TEC) express major histocompatibility complex (MHC) class II molecules, potentially enabling them to present thyroid self-antigens to CD4-positive T cells. However, despite this, TEC may fail to present endogenous antigen as a result of limited processing or MHC class II loading capacity, or inadequate MHC class II levels. We addressed these issues using the cloned rat TEC line, Fischer rat thyroid cell line (FRTL5), which was transfected using an adenoviral expression vector that expressed ovalbumin (OVA) as an integral membrane protein. OVA-expressing FRTL5 cells very efficiently activated a panel of OVA-specific, class II-restricted T-cell hybridomas. This response was dependent on induction of MHC class II molecules by interferon-gamma (IFN-gamma) and was blocked by anti-MHC class II antibodies. Poor responses were seen to exogenously added OVA or OVA peptides. These results provide the most direct evidence to date that TEC can form MHC class II-peptide complexes derived from self-antigen in sufficient quantities to activate T cells. PMID- 10712667 TI - An acidic microenvironment impairs the generation of non-major histocompatibility complex-restricted killer cells. AB - The microenvironment within solid tumours has often been shown to exhibit an acidic local pH. In recent studies we could demonstrate that an acidic extracellular pH (pHe) inhibits the non-major histocompatibility complex (MHC) restricted cytotoxicity of immunocompetent effector cells. However, within tumours the activation of cytotoxic cells may already be impaired by low pHe. Therefore, we investigated the influence of acidic conditions on the generation of active killer cells. The cytotoxic activity of natural killer (NK) as well as lymphokine-activated killer (LAK) cells against K562, Daudi and Raji cells was analysed after an activation period of 3 days at pHe 7.2-6.5. A minor reduction of pHe from 7.2 to 7.0 during the culture period resulted in a strong inhibition of the natural cytotoxicity of NK cells. Furthermore, acidic pHe below 7.2 prevented the generation of activated LAK cells by interleukin-2 (IL-2). The cytotoxic capacity could not be reconstituted if cells cultured at a pHe of 6.5 were returned to physiological pH for another 24 hr. Analysis of the cellular subtypes within the various cultures did not reveal differences regarding the frequencies of NK cells, CD8+ T cells, or CD4+ T cells. However, an acidic pHe clearly inhibited the activation-induced increase of relevant adhesion molecules. The production of cytokines which are involved in the regulation of the cytotoxic process (tumour necrosis factor-alpha, interferon-gamma, IL-10, IL-12 and transforming growth factor-beta1) was also affected by pHe, as their release was strongly inhibited at pHe 7.0. Furthermore, we observed a considerable decrease in the metabolic activity of effector cells at acidic pHe. In summary, our findings suggest that an acidic microenvironment impairs the induction of an anti tumoral immune response within solid tumours. PMID- 10712668 TI - Antigen receptor signalling in apoptosis-resistant mutants of WEHI 231 cells. AB - Ligation of membrane immunoglobulin M (mIgM) induces cell cycle arrest and apoptosis in the WEHI 231 B-lymphoma cell line. The molecular mechanisms which link receptor ligation and the nuclear events that underlie this response, have yet to be fully elucidated. Here we have examined the signals induced following mIgM cross-linking in variants of WEHI 231 that no longer undergo apoptosis in response to this stimulus. Tyrosine phosphorylation of cellular substrates in two of the variants is identical to that seen in wild-type cells but in one of the mutants, VS2.12, a restricted set of substrates becomes tyrosine phosphorylated. In a second variant (E8), mIgM cross-linking does not induce elevation of intracellular Ca2+, although tyrosine phosphorylation of PLCgamma2 is induced to an equivalent extent to that seen in WEHI 231 cells. A third variant, 2E10.F9, is resistant to apoptosis despite the fact that all signals analysed appear to be similar to those induced in wild-type cells. Our findings show that resistance to apoptosis can arise as a result of mutations affecting discrete stages of the mIgM signalling pathway. The mutant lines reported here show defects that have not yet been identified in previous studies and are likely to be useful tools in dissecting the signalling of cell death in B lymphocytes. PMID- 10712669 TI - Differential effects of CD4 and CD8 engagement on the development of cytokine profiles of murine CD4+ and CD8+ T lymphocytes. AB - A simple culture system devoid of antigen-presenting cells was used to examine the ability of immobilized antibodies to lymphocyte function-associated antigen-1 (LFA-1) (CD11a), CD28 and CD4 or CD8 to modulate the responses of normal murine CD4+ and CD8+ lymph node T cells to immobilized anti-CD3 antibody and interleukin 2 (IL-2). All the antibodies enhanced proliferative responses to limiting anti CD3 antibody. Both CD4+ and CD8+ cells produced substantial titres of IL-3 and interferon-gamma (IFN-gamma) in primary and secondary cultures regardless of the coactivating antibodies used for priming. By contrast, the combination of anti CD4 with anti-CD3 antibody stimulated significantly higher titres of IL-4 than any other antibody combination in cultures of CD4+ cells. This CD4-dependent IL-4 response was induced in CD4+ T cells of naive (CD44low) phenotype and was similar in magnitude to the response induced by exogenous IL-4 but, unlike the latter, was not associated with elevated IL-3 synthesis. A comparable effect of anti-CD8 antibodies on CD8+ cells was not observed: although IL-4 production by CD8+ cells was induced by exogenous IL-4, it was not detected following coactivation with anti-CD8 or any other antibodies. We conclude that anti-CD4 antibody is a potent inducer of IL-4-secreting CD4+ T cells whose effects can be distinguished from those of anti-CD8 antibody on CD8+ T cells and from those of IL-4 on either subset. PMID- 10712670 TI - CXC chemokine receptor 4 expression and stromal cell-derived factor-1alpha induced chemotaxis in CD4+ T lymphocytes are regulated by interleukin-4 and interleukin-10. AB - We report that interleukin (IL)-4 and IL-10 can significantly up- or down regulate CXC chemokine receptor 4 (CXCR4) expression on CD4+ T lymphocytes, respectively. Stromal cell-derived factor-1alpha (SDF-1alpha)-induced CD4+ T lymphocyte chemotaxis was also correspondingly regulated by IL-4 and IL-10. IL-4 and IL-10 up- or down-regulated CXCR4 mRNA expression in CD4+ T lymphocytes, respectively, as detected by real-time quantitative reverse transcription polymerase chain reaction (RT-PCR). Scatchard analysis revealed a type of CXCR4 with affinity (Kd approximately 6.3 nM), and approximately 70,000 SDF-1alpha binding sites per cell, among freshly isolated CD4+ T lymphocytes, and two types of CXCR4 with different affinities (Kd1 approximately 4.4 nM and Kd2 approximately 14.6 nM), and a total of approximately 130,000 SDF-1alpha-binding sites per cell, among IL-4-stimulated CD4+ T lymphocytes. The regulation of CXCR4 expression in CD4+ T lymphocytes by IL-4 and IL-10 could be blocked by a selective inhibitor of protein kinase (staurosporine) or by a selective inhibitor of cAMP- and cGMP-dependent protein kinase (H-8), indicating that these cytokines regulate CXCR4 on CD4+ T lymphocytes via both cAMP and cGMP signalling pathways. The fact that cyclosporin A or ionomycin were able to independently change the CXCR4 expression and block the effects of IL-4 and IL-10 on CXCR4 expression implied that the capacity of IL-4 and IL-10 to regulate CXCR4 on CD4+ T lymphocytes is not linked to calcium-mobilization stimulation. These results indicate that the effects of IL-4 and IL-10 on the CXCR4-SDF-1 receptor-ligand pair may be of particular importance in the cytokine/chemokine environment concerning the inflammatory processes and in the progression of human immunodeficiency virus (HIV) infection. PMID- 10712671 TI - Human amniotic fluid lacks interleukin-2 and interleukin-15 but can interact with the beta-chain of the interleukin-2 receptor. AB - The present study investigated whether an explanation for the conflicting reports on the interleukin-2 (IL-2) status of amniotic fluid is due to the presence of IL 15 which shares biological activities with IL-2 and utilizes the IL-2 receptor beta-chain. Amniotic fluids from 45 normally progressing pregnancies between 14 and 16 weeks after the last menstrual period were assayed for IL-2 and IL-15 by bioassay and enzyme-linked immunosorbent assay (ELISA). The ability of amniotic fluids to induce cytotoxic T lymphoblastoid line-2 (CTLL-2) cell proliferation was demonstrated to be dependent upon bioassay culture conditions. In serum-free medium each amniotic fluid stimulated CTLL-2 proliferation with a mean level of IL-2-like bioactivity of 14.7 +/- 2.3 ng/ml but amniotic fluids failed to induce CTLL-2 proliferation in serum-supplemented medium. Treatment with neutralizing anti-IL-2 or anti-IL-15 antibodies failed to inhibit amniotic fluid-induced CTLL cell proliferation in serum-free medium, indicating a lack of IL-2 and IL-15 bioactivity. In contrast, treatment with anti-IL-2 receptor beta-chain antibody significantly reduced amniotic fluid-induced proliferation. The lack of IL-2 and IL-15 activity in amniotic fluids was confirmed using ELISA. Although high levels of IL-15 immunoactivity were detected in all samples, specificity controls showed a lack of specific IL-15 immunoactivity in amniotic fluid. Pretreatment of amniotic fluids with 100-500 ng/ml mouse immunoglobulin G abrogated IL-15 immunoactivity, indicating that amniotic fluid contains molecules binding to Fc regions of immunoglobulins and responsible for false ELISA positivity. These studies unequivocally show that amniotic fluid lacks IL-2 and IL-15 but can stimulate CTLL-2 cell proliferation via the IL-2 receptor beta-chain. The absence of IL-2 and IL-15 in normal mid-trimester amniotic fluid suggests that the cytokine profile of human pregnancy appears to be associated with a bias against type 1 cytokines within the feto-placental unit. PMID- 10712673 TI - Redundancy or cell-type-specific regulation? Tumour necrosis factor in alveolar macrophages and mast cells. AB - Tumour necrosis factor (TNF) is an important inflammatory cytokine produced by several cell types. To test the hypothesis that there is cell-type-specific regulation and not redundancy of TNF production, we investigated its production by alveolar macrophages (AM) and peritoneal mast cells (PMC). Cell lysates of freshly isolated AM and PMC contained 9 +/- 3 pg and 57 +/- 17 pg of TNF/10(6) cells, respectively. Furthermore, unstimulated PMC expressed 4 x 10(3)-fold more attomols of TNF mRNA/microg total RNA compared with AM. These data may explain in part the greater TNF-dependent cytotoxicity of PMC. Furthermore, fixed PMC showed significantly higher TNF-dependent cytotoxic activity than AM (sevenfold), suggesting that PMC express more membrane TNF than AM. Although AM and PMC contain different amounts of TNF, antigen stimulation caused a similar release of TNF from sensitized rats. Interferon (IFN)-gamma, respectively, stimulated and inhibited AM and PMC TNF-dependent cytotoxicity whereas lipopolysaccharide (LPS) significantly stimulated TNF-dependent cytotoxicity in both cell types. However, TNF released (AM 400-fold and PMC threefold) and TNF mRNA expression, as measured by competitive reverse transcription-polymerase chain reaction (AM 7 x 10(3)-fold and PMC twofold), were considerably greater in LPS-stimulated AM than PMC. Our data indicate that TNF is differentially expressed in these two cell types and that its production is dependent on the nature of the stimulus. These data provide vital basis in experimental approaches aimed at modulating the effect of TNF in airway disease conditions involving both AM and mast cells. PMID- 10712672 TI - Nerve growth factor-beta induces mast-cell marker expression during in vitro culture of human umbilical cord blood cells. AB - Nerve growth factor-beta (NGF) is known as a growth factor for human basophils and murine mast cells and has recently been shown to also up-regulate mast cell characteristics in human leukaemic mast cells. We have examined here the effect of NGF on the differentiation of normal human mast cells from cord blood progenitors during culture with stem cell factor (SCF), NGF alone or in combination, or fibroblast supernatants. All these supplements induced mast cell immunoreactivity against tryptase, c-Kit and FcepsilonRIalpha, but none of the cells reacted against the basophil specific antibody 2D7 before or during culture. Intracellular tryptase activity increased as well, with maximal levels on combined culture with SCF and NGF. On reverse transcription-polymerase chain reaction (RT-PCR), cells lacked tryptase and chymase and expressed low levels of FcepsilonRI and c-Kit mRNA prior to culture, with marked up-regulation of FcepsilonRI and c-Kit, and with de novo expression of mast-cell specific alpha- and beta-tryptase by week 3, and of chymase by week 5. Only the TrkA and not the p75 NGF receptor was detected at m-RNA and protein level, and only the TrkA NGF receptor was up-regulated during NGF-driven culture. These findings show therefore that, like SCF, NGF is another growth factor that can induce and regulate human mast-cell development and differentiation. PMID- 10712674 TI - Mast cells enhance contraction of three-dimensional collagen lattices by fibroblasts by cell-cell interaction: role of stem cell factor/c-kit. AB - Reorganization of the extracellular matrix is important in many biological and pathophysiological processes, including tissue remodelling, wound healing, or cancer metastasis. The ability of cultured fibroblasts to reorganize and contract three-dimensional type I collagen gels is regarded as an in vitro model for this process. In tissue fibrosis, complex interactions among fibroblasts, inflammatory cells and the extracellular matrix are taking place. Mast cells have often been discussed to play a role in several fibrotic conditions including scleroderma, scar formation, or wound healing. In this study, we examined the effects of mast cells on contraction of collagen lattices. The results demonstrate that co culture of dermal fibroblasts with a human mast cell line (HMC-1) significantly enhanced contraction of the three-dimensional collagen lattices, whereas mast cells alone failed to contract the gel. Addition of culture supernatants of mast cells did not enhance the speed of gel contraction, indicating the importance of cell-cell contact. Morphological analysis showed that mast cells were incorporated into the lattices. Histological examination also demonstrated that within the lattices, mast cells were localized in close contact to, or attached to, fibroblasts. As fibroblasts and mast cells are known to attach via stem cell factor (SCF)/c-kit interaction when co-cultured in monolayers, we also examined the effect of antibodies against SCF and c-kit in this system. Addition of both antibodies inhibited gel contraction up to 70%. In contrast, antibodies against interleukin-4 (IL-4) and IL-4 receptor did not affect gel contraction. These results indicate that mast cells enhance fibroblast-mediated contraction of collagen lattices via direct cell-cell contact, mediated in part by SCF/c-kit interactions. PMID- 10712676 TI - Evidence for multiple CD95-CD95 ligand interactions in anteriorchamber-associated immune deviation induced by soluble protein antigen. AB - We have investigated whether CD95-CD95 ligand interactions are important in anterior chamber-associated immune deviation (ACAID) induced by soluble protein antigen, and if so, to identify the participating cells on which these molecules are expressed. Peritoneal exudate cells as antigen-presenting cells (APC) obtained from B6.lpr/lpr, B6.gld/gld and C57BL/6 mice were cultured with ovalbumin (OVA) and transforming growth factor-beta2 (TGF-beta2) overnight, then injected intravenously into C57BL/6 or B6.lpr/lpr recipients. Some B6.lpr/lpr mice were reconstituted with naive T cells from wild-type C57BL/6 donors. In other experiments, B6. lpr/lpr and B6.gld/gld mice received an anterior chamber injection of OVA followed 7 days later by subcutaneous immunization with OVA plus adjuvant. Delayed hypersensitivity (DH) was assessed with an ear swelling assay. T cells activated in vitro with OVA-pulsed, TGF-beta-treated APC were tested in vivo for their capacity to suppress DH expression in a local adoptive transfer assay. The results indicate that when ACAID was induced by in-vitro generated ACAID-inducing cells, the APC expressed CD95L, and recipient T cells expressed CD95. The capacity of in vitro generated regulatory T cells to suppress DH expression to OVA in vivo was not governed by CD95-CD95L interactions. When OVA was injected into the anterior chamber of naive mice, CD95 expression was required for ACAID induction, although ACAID was readily induced in CD95L deficient mice. We conclude that CD95-CD95L interactions are required in ACAID for the initial stage of APC presentation of eye-derived antigens to T cells, and that CD95-CD95L interactions participate at one or more additional step in the process by which ACAID is induced by soluble protein antigens. PMID- 10712675 TI - A novel CD18 genomic deletion in a patient with severe leucocyte adhesion deficiency: a possible CD2/lymphocyte function-associated antigen-1 functional association in humans. AB - Leucocyte adhesion deficiency (LAD) is an autosomal-recessive genetic disease that is characterized clinically by severe bacterial infections and caused by mutations in the CD18 gene that codes for the beta2 integrin subunit. A patient with a severe LAD phenotype was studied and the molecular basis of the disease was identified as a single homozygous defect in a Herpes virus saimiri (HVS) transformed T-cell line. The defect identified involves a deletion of 171 bp in the cDNA that encodes part of the proteic extracellular domain. This genetic abnormality was further studied at the genomic DNA level and found to consist of a deletion of 169 bp (from -37 of intron 4 to +132 of exon 5), which abolishes the normal splicing and results in the total skipping of exon 5. The 171-bp shortened 'in-frame' mRNA not only resulted in the absence of CD18 expression on the cell surface but also in its absence in the cytoplasm of HVS T-cell lines. Functionally, the LAD-derived HVS T-cell lines showed a severe, selective T-cell activation impairment in the CD2 (but not in the CD3) pathway. This defect was not reversible when exogenous interleukin-2 (IL-2) was added, suggesting that there is also a functional interaction of the lymphocyte function-associated antigen-1 (LFA-1) protein in the CD2 signal transduction pathway in human T cells, as has been previously reported in mice and in the human Papillon-Lefevre syndrome. Thus, HVS transformation is not only a suitable model for T-cell immunodeficiency studies and characterization, but is also a good system for investigating the immune system in pathological conditions. It may also be used in the future in cellular models for in vitro gene-therapy trials. PMID- 10712677 TI - Induction of T helper 1- and T helper 2-type immune responses during Haemonchus contortus infection in sheep. AB - The production of cytokines by lymphoid cells, isolated from non-infected and Haemonchus contortus-infected lambs, was investigated. Particular attention was paid to differences in T helper 1- (Th1) and Th2-type immune profiles between genetically resistant and random-bred animal groups. Non-infected resistant and random-bred lambs produced equivalent levels of interferon-gamma (IFN-gamma) and interleukin-5 (IL-5), from isolated abomasal lymph node cells (ALN), mesenteric lymph node cells (MLN) and spleen cells (SC), in response to in vitro stimulation with T-cell mitogen (concanavalin A) or larval parasite antigen. ALN and MLN cells derived from infected resistant and random-bred lambs produced relatively lower levels of IFN-gamma, following in vitro stimulation with parasite antigen, when compared with their uninfected counterparts. In contrast, infected lambs of both groups showed enhanced mitogen- and antigen-stimulated production of IL-5, in comparison with uninfected controls, at days 5 and 28 postinfection (p.i.). Mitogen- and antigen-stimulated IL-5 responses were higher among resistant lambs compared with random-bred lambs, with the highest overall production of IL-5 by parasite antigen-stimulated ALN and MLN cells. Among day 28 p.i. lambs, levels of cell culture-derived parasite-specific immunoglobulin G1 (IgG1) and IgE antibodies were higher in resistant lambs than in random-bred lambs, following in vitro stimulation of SC or ALN cells with parasite antigen. Finally, after 28 days p.i., histological examination of abomasal tissue revealed higher densities of mast cells and eosinophils in the mucosa of resistant lambs than in random bred lambs. Taken together, these data support the notion of a strong Th2-type immune response to Haemonchus infection in genetically resistant sheep, and support the claim for a Th1/Th2 dichotomy in ruminants. PMID- 10712678 TI - Interferon-gamma plays a critical role in intestinal immunity against Salmonella typhimurium infection. AB - Salmonella bacteria are a major cause of food-borne infectious diarrhoea and there is great interest in understanding the pathogenesis of Salmonella infection and in vaccine development. Potential vaccines include the aromatic mutants of S. typhimurium. Such non-lethal Aro mutants have also been useful for studying Salmonella infections in mouse models. Studies of systemic infection, using these Aro mutants, in both normal and cytokine gene knockout mice, indicate that interferon-gamma (IFN-gamma) plays a key role in the resolution of Salmonella infection. The present studies have investigated the outcome of oral infection in mice with attenuated Salmonella because this infection route mimics natural infection in humans. In IFN-gamma gene knockout (IFN-gamma-/-) mice, intestinal immunity was impaired and oral challenge resulted in disseminated septicaemia 2 weeks later. No dissemination of infection was seen in wild-type mice. In wild type mice, both CD4 and CD8 cell numbers increased in the gut following Salmonella challenge, together with increased expression of major histocompatibility complex (MHC) II and vascular cell adhesion molecule-1 (VCAM 1). No such changes were seen in IFNgamma-/- mice. Following oral challenge, antilipopolysaccharide (LPS) and antiphosphoryl choline antibodies increased by more than 100-fold in both serum and faecal pellet extracts of IFNgamma-/- mice compared with wild-type mice. Our data show that IFN-gamma production is essential for resolution of enteric Salmonella infection and that antibody has little effect on this process. PMID- 10712679 TI - Mycobacterium tuberculosis-activated dendritic cells induce protective immunity in mice. AB - Activated dendritic cells are critically important in the priming of T-cell responses. In this report we show that the infection of a conditionally immortalized dendritic cell line (tsDC) with Mycobacterium tuberculosis resulted in the up-regulation of B7-1 and B7-2 co-stimulatory molecules and the induction of several inflammatory cytokines, including tumour necrosis factor-alpha and interleukin-6, -1beta and -12. In addition, we show that these activated dendritic cells were capable of eliciting antigen-specific T-cell responses and potent anti-mycobacterial protective immunity in a murine model of experimental tuberculosis infection. PMID- 10712680 TI - Methylcitrate synthase from Aspergillus nidulans: implications for propionate as an antifungal agent. AB - Aspergillus nidulans was used as a model organism to investigate the fungal propionate metabolism and the mechanism of growth inhibition by propionate. The fungus is able to grow slowly on propionate as sole carbon and energy source. Propionate is oxidized to pyruvate via the methylcitrate cycle. The key enzyme methylcitrate synthase was purified and the corresponding gene mcsA, which contains two introns, was cloned, sequenced and overexpressed in A. nidulans. The derived amino acid sequence of the enzyme shows more than 50% identity to those of most eukaryotic citrate synthases, but only 14% identity to the sequence of the recently detected bacterial methylcitrate synthase from Escherichia coli. A mcsA deletion strain was unable to grow on propionate. The inhibitory growth effect of propionate on glucose medium was enhanced in this strain, which led to the assumption that trapping of the available CoA as propionyl-CoA and/or the accumulating propionyl-CoA itself interferes with other biosynthetic pathways such as fatty acid and polyketide syntheses. In the wild-type strain, however, the predominant inhibitor may be methylcitrate. Propionate (100 mM) not only impaired hyphal growth of A. nidulans but also synthesis of the green polyketide derived pigment of the conidia, whereas in the mutant pigmentation was abolished with 20 mM propionate. PMID- 10712681 TI - The development of a FACS-based strategy for the isolation of Shigella flexneri mutants that are deficient in intercellular spread. AB - In the disease course of bacillary dysentery, pathogenic Shigella flexneri invade colonic epithelial cells and spread both within and between host cells. The ability to spread intercellularly allows the organism to infect an entire epithelial layer without significant contact with the extracellular milieu. Using fluorescence activated cell sorter (FACS)-based technology, we developed a rapid and powerful selection strategy for the isolation of S. flexneri mutants that are unable to spread from cell to cell. The majority of mutants identified using this strategy harbour mutations that affect the structure of their lipopolysaccharide or the ability of the bacteria to move intracellularly via actin-based motility; both factors have previously been shown to be essential for cell-to-cell spread. However, using a modified strategy that eliminated both of these types of mutants, we identified several mutants that provide us with evidence that bacterial proteins of the type III secretion system, which are essential for bacterial entry into host cells, also play a role in cell-to-cell spread. PMID- 10712682 TI - Modulation of host immune responses, induction of apoptosis and inhibition of NF kappaB activation by the Bordetella type III secretion system. AB - Bordetella bronchiseptica establishes respiratory tract infections in laboratory animals with high efficiency. Colonization persists for the life of the animal and infection is usually asymptomatic in immunocompetent hosts. We hypothesize that this reflects a balance between immunostimulatory events associated with infection and immunomodulatory events mediated by the bacteria. We have identified 15 loci that are part of a type III secretion apparatus in B. bronchiseptica and three secreted proteins. The functions of the type III secretion system were investigated by comparing the phenotypes of wild-type bacteria with two strains that are defective in type III secretion using in vivo and in vitro infection models. Type III secretion mutants were defective in long term colonization of the trachea in immunocompetent mice. The mutants also elicited higher titres of anti-Bordetella antibodies upon infection compared with wild-type bacteria. Type III secretion mutants also showed increased lethal virulence in immunodeficient SCID-beige mice. These observations suggest that type III-secreted products of B. bronchiseptica interact with components of both innate and adaptive immune systems of the host. B. bronchiseptica induced apoptosis in macrophages in vitro and inflammatory cells in vivo and type III secretion was required for this process. Infection of an epithelial cell line with high numbers of wild type, but not type III deficient B. bronchiseptica resulted in rapid aggregation of NF-kappaB into large complexes in the cytoplasm. NF-kappaB aggregation was dependent on type III secretion and aggregated NF kappaB did not respond to TNFalpha activation, suggesting B. bronchiseptica may modulate host immunity by inactivating NF-kappaB. Based on these in vivo and in vitro results, we hypothesize that the Bordetella type III secretion system functions to modulate host immune responses during infection. PMID- 10712683 TI - A member of the Plasmodium falciparum Pf60 multigene family codes for a nuclear protein expressed by readthrough of an internal stop codon. AB - Four large multigene families have been described in Plasmodium falciparum malaria parasites (var, rif, stevor and Pf60). var and rif genes code for erythrocyte surface proteins and undergo clonal antigenic variation. We report here the characterization of the first Pf60 gene. The 6.1 gene is constitutively expressed by all mature blood stages and codes for a protein located within the nucleus. It has a single copy, 7-exon, 5' domain, separated by an internal stop codon from a 3' domain that presents a high homology with var exon II. Double site immunoassay and P. falciparum transient transfection using the reporter luciferase gene demonstrated translation through the internal ochre codon. The 6.1 N-terminal domain has no homology with any protein described to date. Sequence analysis identified a leucine zipper and a putative nuclear localization signal and showed a high probability for coiled coils. Evidence for N-terminal coiled coil-mediated protein interactions was obtained. This identifies the 6.1 protein as a novel nuclear protein. These data show that the Pf60 and var genes form a superfamily with a common 3' domain, possibly involved in regulating homo- or heteromeric interactions. PMID- 10712684 TI - Anaerobic nitrate reductase (narGHJI) activity of Mycobacterium bovis BCG in vitro and its contribution to virulence in immunodeficient mice. AB - Mycobacterium tuberculosis and Mycobacterium bovis cause tuberculosis, which is responsible for the deaths of more people each year than any other bacterial infectious disease. Disseminated disease with Mycobacterium bovis BCG, the only currently available vaccine against tuberculosis, occurs in immunocompetent and immunodeficient individuals. Although mycobacteria are obligate aerobes, they are thought to face an anaerobic environment during infection, notably inside abscesses and granulomas. The purpose of this study was to define a metabolic pathway that could allow mycobacteria to exist under these conditions. Recently, the complete genome of M. tuberculosis has been sequenced, and genes homologous to an anaerobic nitrate reductase (narGHJI), an enzyme allowing nitrate respiration when oxygen is absent, were found. Here, we show that the narGHJI cluster of M. tuberculosis is functional as it conferred anaerobic nitrate reductase activity to Mycobacterium smegmatis. A narG mutant of M. bovis BCG was generated by targeted gene deletion. The mutant lacked the ability to reduce nitrate under anaerobic conditions. Both mutant and M. bovis BCG wild type grew equally well under aerobic conditions in vitro. Histology of immunodeficient mice (SCID) infected with M. bovis BCG wild type revealed large granulomas teeming with acid-fast bacilli; all mice showed signs of clinical disease after 50 days and succumbed after 80 days. In contrast, mice infected with the mutant had smaller granulomas containing fewer bacteria; these mice showed no signs of clinical disease after more than 200 days. Thus, it seems that nitrate respiration contributes significantly to virulence of M. bovis BCG in immunodeficient SCID mice. PMID- 10712685 TI - Characterization of the in vivo acceptors of the mycoloyl residues transferred by the corynebacterial PS1 and the related mycobacterial antigens 85. AB - Mycolic acids, long-chain (C70-C90) alpha-alkyl, beta-hydroxy fatty acids, are characteristic cell envelope components of mycobacteria; similar but shorter chain substances occur in corynebacteria and related taxa. These compounds apparently play an important role in the physiology of these bacteria. The deduced N-terminal region of PS1, one of the two major secreted proteins of Corynebacterium glutamicum encoded by the csp1 gene, is similar to the antigens 85 complex of Mycobacterium tuberculosis which has been shown to be associated in vitro with a mycoloyltransferase activity onto trehalose. Overexpression of PS1 in the wild-type strain of C. glutamicum suggested the implication of the protein in the transfer of corynomycolates, evidenced by an increase esterification of the cell wall arabinogalactan with corynomycolic acid residues and an accumulation of trehalose dicorynomycolates. Overexpression of truncated forms of PS1 demonstrated that the crucial region for transfer activity of the protein involves all the region of homology with antigens 85. To establish the putative mycoloyltransferase activity of PS1, a csp1-inactivated mutant of C. glutamicum was biochemically characterized. Inactivation of the gene resulted in: (i) a 50% decrease in the cell wall corynomycolate content; (ii) the alteration of the permeability of the C. glutamicum cell envelope; (iii) the decrease of the trehalose dicorynomycolate content; (iv) the accumulation of trehalose monocorynomycolate; and (v) the appearance of a glycolipid identified as 6 corynomycoloylglucose. Complementation of the mutant by the csp1 gene fully restored the wild-type phenotype. Finally, a mycoloyltransferase assay established that PS1 possesses a trehalose mycoloyltransferase activity. To define the in vivo function of antigens 85, the csp1-inactivated mutant was complemented with the fbpA, fbpB or fbpC genes. Complementation with the different fbp genes restored the normal cell wall corynomycolate content and permeability, but did not affect either the fate of trehalose corynomycolates or the occurrence of glucose corynomycolate. Thus, PS1 is one of the enzymes that transfer corynomycoloyl residues onto both the cell wall arabinogalactan and trehalose monocorynomycolate, whereas in the whole bacterium the mycobacterial antigens 85A, 85B and 85C can transfer mycolates only onto the cell wall acceptor in C. glutamicum. PMID- 10712686 TI - HtrA is the unique surface housekeeping protease in Lactococcus lactis and is required for natural protein processing. AB - We identified an exported protease in Lactococcus lactis ssp. lactis strain IL1403 belonging to the HtrA/DegP family. Inactivation of the chromosomal gene (htrALl) encoding this protease (HtrALl) results in growth thermo-sensitivity at very high temperatures (above 37 degrees C for L. lactis). The role of htrALl in extracellular proteolysis under normal growth conditions was examined by testing the stability of different exported proteins (i.e. fusions, a heterologous pre pro-protein or a native protein containing repeats), having different locations. In the wild-type (wt) strain, degradation products, including the C-terminal protein ends, were present in the medium, indicating that proteolysis occurs during or after export to the cell surface; in one case, degradation was nearly total. In contrast, proteolysis was totally abolished in the htrA strain for all five proteins tested, and the yield of full-length products was significantly increased. These results suggest that HtrALl is the sole extracellular protease that degrades abnormal exported proteins. In addition, our results reveal that HtrALl is needed for the pro-peptide processing of a natural pro-protein and for maturation of a native protein. We propose that in lactococci, and possibly in other Gram-positive organisms with small sized-genomes, a single surface protease, HtrA, is totally responsible for the housekeeping of exported proteins. PMID- 10712687 TI - Interactions among components of the Salmonella flagellar export apparatus and its substrates. AB - We have examined the cytoplasmic components (FliH, FliI and FliJ) of the type III flagellar protein export apparatus, plus the cytoplasmic domains (FlhAC and FlhBC) of two of its six membrane components. FliH, FlhAC and FliJ, when overproduced, caused inhibition of motility of wild-type cells and inhibition of the export of substrates such as the hook protein FlgE. Co-overproduction of FliH and FliI substantially relieved the inhibition caused by FliH, suggesting that it is excess free FliH that is inhibitory and that FliH and FliI form a complex. We purified His-FLAG-tagged versions of: (i) export components FliH, FliI, FliJ, FlhAC and FlhBC; (ii) rod/hook-type export substrates FlgB (rod protein), FlgE (hook protein), FlgD (hook capping protein) and FliE (basal body protein); and (iii) filament-type export substrates FlgK and FlgL (hook-filament junction proteins) and FliC (flagellin). We tested for protein-protein interactions by affinity blotting. In many cases, a given protein interacted with more than one other component, indicating that there are likely to be multiple dynamic interactions or interactions that involve more than two components. Interactions of FlhBC with rod/hook-type substrates were strong, whereas those with filament type substrates were very weak; this may reflect the role of FlhB in substrate specificity switching. We propose a model for the flagellar export apparatus in which FlhA and FlhB and the other four integral membrane proteins of the apparatus form a complex at the base of the flagellar motor. A soluble complex of at least three proteins (FliH, FliI and FliJ) bind the protein to be exported and then interact with the complex at the motor to deliver the protein, which is then exported in an ATP-dependent process mediated by FliI. PMID- 10712688 TI - Identification of the Escherichia coli K-12 Nramp orthologue (MntH) as a selective divalent metal ion transporter. AB - The Escherichia coli mntH (formerly yfeP) gene encodes a putative membrane protein (MntH) highly similar to members of the eukaryotic Nramp family of divalent metal ion transporters. To determine the function of E. coli MntH, a null mutant was created and MntH was overexpressed both in wild-type E. coli and in the metal-dependent mutant hflB1(Ts). At the restrictive temperature 42 degrees C, the mntH null mutation reduces the suppression of hflB1(Ts) thermosensitivity by exogenous divalent metals. Conversely, overexpression of MntH restores growth at 42 degrees C, increases suppression of the ts phenotype by Fe(II) and Ni(II) and renders hflB1(Ts) cells hypersensitive to Mn(II). Transport studies in intact cells show that MntH selectively facilitates uptake of 54Mn(II) and 55Fe(II) in a temperature-, time- and proton-dependent manner. Competition studies in uptake assays and growth inhibition experiments in hflB1(Ts) mutants together indicate that MntH is a divalent metal cation transporter of broad substrate specificity. The functional characteristics of MntH suggest that it corresponds to the previously described manganese transporter of E. coli. This study indicates that proton-dependent divalent metal ion uptake has been preserved in the Nramp family from bacteria to humans. PMID- 10712689 TI - Molecular cloning of the calcium and sodium ATPases in Neurospora crassa. AB - Using PCR, reverse transcription-PCR (RT-PCR) and colony hybridization in a genomic library, we isolated six genes which encode type II P-type ATPases in Neurospora crassa. The six full-length cDNAs were cloned in a yeast expression vector and transformed into Saccharomyces cerevisiae null Ca2+- or Na+-ATPase mutants. Three cDNAs suppressed the defect of the Ca2+ mutant and two of these protected from Mn2+ toxicity. One cDNA suppressed the defect of the Na+ mutant and two cDNAs were not functional in S. cerevisiae. The expression of the transcripts of the six genes in the presence of Ca2+, Na+, high pH or supporting an osmotic shock indicated that, with the exception of one of the Ca2+-ATPases, the main function of the cloned ATPases is the adaptation to stress conditions. The relationship between the cloned fungal Ca2+- and Na+-ATPases and plant type II P-ATPases is discussed. PMID- 10712690 TI - Regulation of competence for genetic transformation in Streptococcus pneumoniae: expression of dpnA, a late competence gene encoding a DNA methyltransferase of the DpnII restriction system. AB - The chromosomal DpnII gene cassette of Streptococcus pneumoniae encodes two methyltransferases and an endonuclease. One methyltransferase acts on double stranded and the other on single-stranded DNA. Two mRNAs are transcribed from the cassette. One, a SigA promoter transcript, includes all three genes; the other includes a truncated form of the second methyltransferase gene (dpnA) and the endonuclease gene. The truncated dpnA, which is translated from the second start codon in the full gene, was shown to produce active enzyme. A promoter reporter plasmid for S. pneumoniae was devised to characterize the promoter for the second mRNA. This transcript was found to depend on a promoter that responded to the induction of competence for genetic transformation. The promoter contains the combox sequence recognized by a SigH-containing RNA polymerase. As part of the competence regulon, the dpnA gene makes a product able to methylate incoming plasmid strands to protect them from the endonuclease and allow plasmid establishment. Its function differs from most genes in the regulon, which are involved in DNA uptake. Comparison of R6 and Rx strains of S. pneumoniae showed the temperature dependence of transformation in R6 to result from temperature sensitivity of the uptake apparatus and not the development of competence. PMID- 10712691 TI - Transfer of electrons across the cytoplasmic membrane by DsbD, a membrane protein involved in thiol-disulphide exchange and protein folding in the bacterial periplasm. AB - Reduction of non-native protein disulphides in the periplasm of Escherichia coli is catalysed by three enzymes, DsbC, DsbG and DsbE, each of which harbours a catalytic Cys-X-X-Cys dithiol motif. This dithiol motif requires continuous reduction for activity. Genetic evidence suggests that the source of periplasmic reducing power resides within the cytoplasm, provided by thioredoxin (trxA) and thioredoxin reductase (trxB). Cytoplasmic electrons donated by thioredoxin are thought to be transferred into the periplasm via the DsbD membrane protein. To understand the molecular nature of electron transfer, we have analysed the membrane topology of DsbD. DsbD is exported by an N-terminal signal peptide. The N- and C-terminal domains are positioned in the periplasmic space, connected by eight transmembrane segments. Electron transfer was shown to require five cysteine sulphydryl of DsbD. Trans complementation of mutant DsbD molecules revealed intermolecular electron transfer. We discuss a model whereby the membrane-embedded disulphides of DsbD accept electrons from cytoplasmic thioredoxin and transfer them to the C-terminal periplasmic dithiol motif of DsbD. PMID- 10712692 TI - Characterization of ylbF, a new gene involved in competence development and sporulation in Bacillus subtilis. AB - We used mini Tn10 transposition to generate a library of Bacillus subtilis insertion mutants, with the goal of identifying and characterizing new competence genes. Two new regulatory genes were identified in our screen: ypuN (also known as rsiX, the anti-sigmaX factor) and ylbF. The disruption of ylbF leads to a dramatic decrease in the expression of comK, encoding the competence transcription factor. Our data show that ylbF positively controls ComK at a post transcriptional level. It has been reported previously that ComK is degraded in vivo and in vitro by a multimeric protein complex composed of ClpP, ClpC and MecA. This proteolysis is inhibited by the ComS peptide. We show that both the overexpression of comS and the inactivation of mecA individually suffice to bypass the competence phenotype of the ylbF mutation. This mutation does not seem to alter the cellular concentrations of MecA or ClpP, and we propose a role for YlbF in modulating the translation, stability or activity of ComS. In addition to its role in competence, ylbF also appears to regulate sporulation by acting before stage II. PMID- 10712693 TI - Multiple N-acetyl neuraminic acid synthetase (neuB) genes in Campylobacter jejuni: identification and characterization of the gene involved in sialylation of lipo-oligosaccharide. AB - N-acetyl neuraminic acid (NANA) is a common constituent of Campylobacter jejuni lipo-oligosaccharide (LOS). Such structures often mimic human gangliosides and are thought to be involved in the triggering of Guillain-Barre syndrome (GBS) and Miller-Fisher syndrome (MFS) following C. jejuni infection. Analysis of the C. jejuni NCTC 11168 genome sequence identified three putative NANA synthetase genes termed neuB1, neuB2 and neuB3. The NANA synthetase activity of all three C. jejuni neuB gene products was confirmed by complementation experiments in an Escherichia coli neuB-deficient strain. Isogenic mutants were created in all three neuB genes, and for one such mutant (neuB1) LOS was shown to have increased mobility. C. jejuni NCTC 11168 wild-type LOS bound cholera toxin, indicating the presence of NANA in a LOS structure mimicking the ganglioside GM1. This property was lost in the neuB1 mutant. Gas chromatography-mass spectrometry and fast atom bombardment-mass spectrometry analysis of LOS from wild-type and the neuB1 mutant strain demonstrated the lack of NANA in the latter. Expression of the neuB1 gene in E. coli confirmed that NeuB1 was capable of in vitro NANA biosynthesis through condensation of N-acetyl-D-mannosamine and phosphoenolpyruvate. Southern analysis demonstrated that the neuB1 gene was confined to strains of C. jejuni with LOS containing a single NANA residue. Mutagenesis of neuB2 and neuB3 did not affect LOS, but neuB3 mutants were aflagellate and non-motile. No phenotype was evident for neuB2 mutants in strain NCTC 11168, but for strain G1 the flagellin protein from the neuB2 mutant showed an apparent reduction in molecular size relative to the wild type. Thus, the neuB genes of C. jejuni appear to be involved in the biosynthesis of at least two distinct surface structures: LOS and flagella. PMID- 10712694 TI - The respiratory gene OXA1 has two fission yeast orthologues which together encode a function essential for cellular viability. AB - The Saccharomyces cerevisiae nuclear gene OXA1, which is conserved from prokaryotes to human, was shown to be essential for cytochrome c oxidase and F1F0 ATP synthase biogenesis. We have searched for an orthologue of OXA1 in Schizosaccharomyces pombe, another yeast that is highly diverged from S. cerevisiae and which could more closely model higher eukaryotes. In particular, S. pombe exhibits a limited growth under anaerobic conditions and is petite negative, that is it does not tolerate large deletions of its mitochondrial DNA. Surprisingly, two S. pombe cDNAs able to complement an S. cerevisiae oxa1 mutation were isolated. The corresponding genes have different chromosomal locations and intron contents. They encode distinct proteins, both sharing a weak sequence identity one with the other and with Oxa1p. A phenotypic analysis of both single inactivations demonstrates that only one gene is essential for respiration in S. pombe. However, the double inactivation is lethal. This work gives new insight into the dependence of S. pombe viability upon oxa1 function, providing evidence of a connection between petite negativity, a functional respiratory chain and F1F0-ATP synthase complex in S. pombe. PMID- 10712696 TI - Functional genomics of Helicobacter pylori: identification of a beta-1,4 galactosyltransferase and generation of mutants with altered lipopolysaccharide. AB - A previously annotated open reading frame (ORF) (HP0826) from Helicobacter pylori was cloned and expressed in Escherichia coli cells and determined to be a beta 1,4-galactosyltransferase that used GlcNAc as an acceptor. Mutational analysis in H. pylori strains demonstrated that this enzyme plays a key role in the biosynthesis of the type 2 N-acetyl-lactosamine (LacNAc) polysaccharide O-chain backbone, by catalysing the addition of Gal to GlcNAc. To examine the potential role of this O-chain structure in bacterial colonization of the host stomach, the mutation was introduced into H. pylori strain SS1 which is known to be capable of colonizing the gastric mucosa of mice. Compared with the parental strain, mutated SS1 was less efficient at colonizing the murine stomach. PMID- 10712695 TI - The putative iron transport system SitABCD encoded on SPI1 is required for full virulence of Salmonella typhimurium. AB - Salmonella typhimurium is an invasive pathogen that causes diseases ranging from mild gastroenteritis to enteric fever. During the infection process, S. typhimurium induces a number of virulence genes required to circumvent host defences and/or acquire nutrients in the host. We have used the in vivo expression technology (IVET) system to select for S. typhimurium genes that are induced after invasion of a murine cultured cell line. We have characterized a putative iron transporter in Salmonella pathogenicity island 1, termed sitABCD. The sitABCD operon is induced under iron-deficient conditions in vitro and is repressed by Fur. This locus is induced in the animal specifically after invasion of the intestinal epithelium. We show that a sit null mutant is significantly attenuated in BALB/c mice, suggesting that SitABCD plays an important role in iron acquisition in the animal. PMID- 10712697 TI - The archaeal halophilic virus-encoded Dam-like methyltransferase M. phiCh1-I methylates adenine residues and complements dam mutants in the low salt environment of Escherichia coli. AB - The genome of the archaeal virus phiCh1, infecting Natrialba magadii (formerly Natronobacterium magadii), is composed of 58.5 kbp linear ds DNA. Virus particles contain several RNA species in sizes of 100-800 nucleotides. A fraction of phiCh1 genomes is modified within 5'-GATC-3' and related sequences, as determined by various restriction enzyme digestion analyses. High performance liquid chromatography revealed a fifth base, in addition to the four nucleosides, which was identified as N6-methyladenosine. Genetic analyses and subsequent sequencing led to the identification of a DNA (N6-adenine) methyltransferase (mtase) gene. The protein product was designated M.phiCh1-I. By the localization of the most conserved motifs (a DPPY motif occurring before FxGxG), the enzyme was placed within the beta-subgroup of the (N6-adenine) methyltransferase class. The mtase gene of phiCh1 was classified as a 'late' gene, as determined by measuring the kinetics of mRNA and protein expression in N. magadii during the lytic cycle of phiCh1. After infection of cells, M.phiCh1-I mRNA and protein could be detected in lower amounts than in the situation of virus induction from lysogenic cells. Consequently, only about 5% of the phiCh1 progeny genomes after infection of N. magadii carry the M.phiCh1-I methylation in contrast to 50% of virus genomes generated by induction of phiCh1-lysogenic N. magadii cells. Heterologous expression of the mtase from a halophile with 3 M cytoplasmic salt concentration showed an unexpected feature: the protein was active in the low environment of Escherichia coli and was able to methylate DNA in vivo. Interestingly, it seemed to exhibit a higher sequence specificity in E. coli that resulted in adenine methylation exclusively in the sequence 5'-GATC-3'. Additionally, expression of M.phiCh1-I in dam- E. coli cells led to a complete substitution of the function of M. Dam in DNA mismatch repair. PMID- 10712698 TI - The bacteriophage T4 anti-sigma factor AsiA is not necessary for the inhibition of early promoters in vivo. AB - Bacteriophage T4 early promoters are utilized immediately after infection and are abruptly turned off 2-3 min later (at 30 degrees C) when the middle promoters are activated. The viral early protein AsiA has been suspected to bring about this transcriptional switch: not only does it activate transcription at middle promoters in vivo and in vitro but it also shows potent anti-sigma70 activity in vitro, suggesting that it is responsible for the shut-off of early transcription. We show here that after infection with a phage deleted for the asiA gene the inhibition of early transcription occurs to the same extent and with the same kinetics as in a wild-type infection. Thus, another AsiA-independent circuit efficiently turns off early transcription. The association of a mutation in asiA with a mutation in mod, rpbA, motA or motB has no effect on the inhibition of early promoters, showing that none of these phage-encoded transcriptional regulators is necessary for AsiA-independent shut-off. It is not known whether AsiA is able to inhibit early promoters in vivo, but host transcription is strongly inhibited in vivo upon induction of AsiA from a multicopy plasmid. PMID- 10712699 TI - NtcA represses transcription of gifA and gifB, genes that encode inhibitors of glutamine synthetase type I from Synechocystis sp. PCC 6803. AB - Synechocystis sp. PCC 6803 glutamine synthetase type I (GS) activity is controlled by direct interaction with two inactivating factors (IF7 and IF17). IF7 and IF17 are homologous polypeptides encoded by the gifA and gifB genes respectively. We investigated the transcriptional regulation of these genes. Expression of both genes is maximum in the presence of ammonium, when GS is inactivated. Nitrogen starvation attenuates the ammonium-mediated induction of gifA and gifB as well as the ammonium-mediated inactivation of GS. Putative binding sites for the transcription factor NtcA were identified at -7.5 and -30.5 bp upstream of gifB and gifA transcription start points respectively. Synechocystis NtcA protein binding to both promoters was demonstrated by gel electrophoresis mobility shift assays. Constitutive high expression levels of both genes were found in a Synechocystis NtcA non-segregated mutant (SNC1), which showed a fourfold reduction in the ntcA expression. These experiments indicate a repressive role for NtcA on the transcription of gifA and gifB genes. Our results demonstrate that NtcA plays a central role in GS regulation in cyanobacteria, stimulating transcription of the glnA gene (GS structural gene) and suppressing transcription of the GS inactivating factor genes gifA and gifB. PMID- 10712700 TI - Two types of cold sensitivity associated with the A184-->V change in the DnaA protein. AB - Multicopy dnaA(Ts) strains carrying the dnaA5 or dnaA46 allele are high temperature resistant but are cold sensitive for colony formation. The DnaA5 and DnaA46 proteins both have an A184-->V change in the ATP binding motif of the protein, but they also have one additional mutation. The mutations were separated, and it was found that a plasmid carrying exclusively the A184-->V mutation conferred a phenotype virtually identical to that of the dnaA5 plasmid. Strains carrying plasmids with either of the additional mutations behaved like a strain carrying the dnaA+ plasmid. In temperature downshifts from 42 degrees C to 30 degrees C, chromosome replication was stimulated in the multicopy dnaA46 strain. The DNA per mass ratio increased threefold, and exponential growth was maintained for more than four mass doublings. Strains carrying plasmids with the dnaA(A184-->V) or the dnaA5 gene behaved differently. The temperature downshift resulted in run out of DNA synthesis and the strains eventually ceased growth. The arrest of DNA synthesis was not due to the inability to initiate chromosome replication because marker frequency analysis showed high initiation activity after temperature downshift. However, the marker frequencies indicated that most, if not all, of the newly initiated replication forks were stalled soon after the onset of chromosome replication. Thus, it appears that the multicopy dnaA(A184- >V) strains are cold sensitive because of an inability to elongate replication at low temperature. The multicopy dnaA46 strains, on the contrary, exhibit productive initiation and normal fork movement. In this case, the cold-sensitive phenotype may be due to DNA overproduction. PMID- 10712701 TI - Non-hydrolysable GTP-gamma-S stabilizes the FtsZ polymer in a GDP-bound state. AB - FtsZ, a tubulin homologue, forms a cytokinetic ring at the site of cell division in prokaryotes. The ring is thought to consist of polymers that assemble in a strictly GTP-dependent way. GTP, but not guanosine-5'-O-(3-thiotriphosphate) (GTP gamma-S), has been shown to induce polymerization of FtsZ, whereas in vitro Ca2+ is known to inhibit the GTP hydrolysis activity of FtsZ. We have studied FtsZ dynamics at limiting GTP concentrations in the presence of 10 mM Ca2+. GTP and its non-hydrolysable analogue GTP-gamma-S bind FtsZ with similar affinity, whereas the non-hydrolysable analogue guanylyl-imidodiphosphate (GMP-PNP) is a poor substrate. Preformed FtsZ polymers can be stabilized by GTP-gamma-S and are destabilized by GDP. As more than 95% of the nucleotide associated with the FtsZ polymer is in the GDP form, it is concluded that GTP hydrolysis by itself does not trigger FtsZ polymer disassembly. Strikingly, GTP-gamma-S exchanges only a small portion of the FtsZ polymer-bound GDP. These data suggest that FtsZ polymers are stabilized by a small fraction of GTP-containing FtsZ subunits. These subunits may be located either throughout the polymer or at the polymer ends, forming a GTP cap similar to tubulin. PMID- 10712702 TI - Identification of a candidate glycosaminoglycan-binding adhesin of the Lyme disease spirochete Borrelia burgdorferi. AB - Binding of glycosaminoglycans (GAGs) by Borrelia burgdorferi, the Lyme disease spirochete, has the potential to promote the colonization of diverse tissues. GAG binding by B. burgdorferi is associated with haemagglutination and we have identified a 26 kDa protein, which we have termed Bgp (Borrelia GAG-binding protein), on the basis of its ability to bind to heparin and erythrocytes. Bgp was found in outer membrane fractions of B. burgdorferi and on the surface of intact bacteria, as assayed by labelling with a membrane-impermeable biotinylating agent or anti-Bgp antibodies. Purified recombinant Bgp agglutinated erythrocytes, binds to the same spectrum of GAGs as the B. burgdorferi strain from which the cloned bgp sequence was obtained, and inhibited B. burgdorferi binding to purified GAGs and to cultured mammalian cells. Thus, Bgp is a strong candidate for a GAG-binding adhesin of B. burgdorferi. PMID- 10712703 TI - Rapid inactivation of the Escherichia coli Kdp K+ uptake system by high potassium concentrations. AB - The Kdp K+ uptake system of Escherichia coli is induced by limitation for K+ and/or high osmolarity. In the present study, the regulation of the activity of the Kdp system has been investigated in E. coli mutants possessing only the Kdp system as the mechanism of K+ accumulation. Cells grown in the presence of low K+ (0.1-1 mM) exhibit normal growth. However, growth inhibition results from exposure of cells to moderate levels of external K+ (> 5 mM). Measurement of the cytoplasmic pH, of K+ pools and of transport via the Kdp system demonstrates that the Kdp system is rapidly and irreversibly inhibited by moderate external K+. Concentrations of K+ greater than 2 mM are sufficient to cause inhibition of Kdp. At pH 6, this results in rapid lowering of the capacity for pH homeostasis, but at pH 7 the intracellular pH is unaffected. Parallel analysis of the expression of the Kdp system in a Kdp+/kdpFABC-lacZ strain shows that levels of K+ that are sufficient to inhibit Kdp activity also repress expression. As a result, growth inhibition of strains solely possessing Kdp arises jointly from inhibition of Kdp activity and repression of Kdp gene expression. These data identify an important aspect of the regulation of potassium transport via the Kdp system and also provide support for a model of regulation of Kdp expression via at least two mechanisms: sensing of both turgor and external K+ concentration. PMID- 10712704 TI - cis-acting regulatory sequences for antitermination in the transcript of the Bacillus subtilis hut operon and histidine-dependent binding of HutP to the transcript containing the regulatory sequences. AB - The location of the cis-acting regulatory region for histidine-dependent antitermination of the Bacillus subtilis hut operon was determined. A secondary structure, whose sequences partially overlap with the downstream terminator, was found in the regulatory region of the hut transcript. Mutational analysis of the regulatory region showed that the secondary structure was required for histidine dependent antitermination. An electrophoretic mobility-shift assay demonstrated that, in response to the presence of histidine and Mg2+, purified HutP bound hut RNA bearing putative secondary structure but not RNA lacking the potential to form putative secondary structure. Native gel electrophoresis showed that HutP existed as a hexamer. A filter-binding assay revealed that the concentration of histidine required for half-maximal binding of HutP to RNA was 3.1 mM and that the Kd for binding of HutP to RNA was approximately 0.56 microM in the presence of histidine. These results suggested that putative secondary structure in the regulatory region of hut mRNA could function as an antiterminator to inhibit the formation of the terminator structure and that HutP causes expression of the hut structural genes by binding to the putative antiterminator structure in response to the presence of histidine. PMID- 10712705 TI - The transmembrane domain 10 of the yeast Pdr5p ABC antifungal efflux pump determines both substrate specificity and inhibitor susceptibility. AB - We have previously shown that a S1360F mutation in transmembrane domain 10 (TMD10) of the Pdr5p ABC transporter modulates substrate specificity and simultaneously leads to a loss of FK506 inhibition. In this study, we have constructed and characterized the S1360F/A/T and T1364F/A/S mutations located in the hydrophilic face of the amphipatic Pdr5p TMD10. A T1364F mutation leads to a reduction in Pdr5p-mediated azole and rhodamine 6G resistance. Like S1360F, the T1364F and T1364A mutants were nearly non-responsive to FK506 inhibition. Most remarkably, however, the S1360A mutation increases FK506 inhibitor susceptibility, because Pdr5p-S1360A is hypersensitive to FK506 inhibition when compared with either wild-type Pdr5p or the non-responsive S1360F variant. Hence, the Pdr5p TMD10 determines both azole substrate specificity and susceptibility to reversal agents. This is the first demonstration of a eukaryotic ABC transporter where a single residue change causes either a loss or a gain in inhibitor susceptibility, depending on the nature of the mutational change. These results have important implications for the design of efficient reversal agents that could be used to overcome multidrug resistance mediated by ABC transporter overexpression. PMID- 10712706 TI - Rad6p represses yeast-hypha morphogenesis in the human fungal pathogen Candida albicans. AB - Rad6p plays important roles in post-replication DNA repair, chromatin organization, gene silencing and meiosis. In this study, we show that Rad6p also regulates yeast-hypha morphogenesis in the human pathogen Candida albicans. CaRAD6 gene and cDNAs were isolated and characterized revealing that the gene carries two 5'-proximal introns. CaRad6p shows a high degree of sequence similarity to Rad6 proteins from fungi to man (60-83% identity), and it suppresses the UV sensitivity and lack of induced mutagenesis displayed by a Saccharomyces cerevisiae rad6 mutant. In C. albicans, CaRAD6 expression is induced in response to UV, and CaRad6p depletion confers UV sensitivity, confirming that Rad6p serves a role in protecting this fungus against UV damage. CaRAD6 overexpression inhibits hyphal development, whereas CaRad6p depletion enhances hyphal growth. Also, CaRAD6 mRNA levels decrease during the yeast-hypha transition. These effects are dependent on Efg1p, but not Cph1p, indicating that CaRad6p acts specifically through the Efg1p morphogenetic signalling pathway to repress yeast-hypha morphogenesis. PMID- 10712707 TI - Modified ultrafiltration and open heart surgery in children. PMID- 10712709 TI - Historical abstract PMID- 10712708 TI - Pain, nociception and the developing infant. AB - The study of paediatric pain and the provision of safe but reliable analgesia for all age groups has become a central issue in paediatric anaesthesia. For the practising paediatric anaesthetist, this represents a major challenge: the developing infant is not a single discrete entity but one that changes constantly with increasing maturity of individual organs. Recent developments in both basic and clinical sciences has given valuable insights into this process, giving the clinician a firm basis to provide analgesia at all ages. This review looks at some of the most interesting recent developments in this field. PMID- 10712710 TI - Comparison of three techniques for induction of anaesthesia with sevoflurane in children. AB - This study was designed to evaluate the clinical characteristics of three induction techniques using sevoflurane in children scheduled for tonsillectomy: incremental induction with sevoflurane(2,4,6,7%) in 100% O2 (group IC-O2; n=23); induction with high concentration of sevoflurane in 100% O2 (group HC-O2; n=22); and induction with high concentration of sevoflurane in a mixture of O2:N2O(50:50) (group HC-N2O; n=20). Induction was well accepted and well tolerated in most children. The addition of nitrous oxide resulted in faster loss of consciousness (P< 0.001) compared to the other induction techniques and in a tendency for reduced excitement compared with the same rapid technique without nitrous oxide (P=0.053). Time to tracheal intubation was identical in the three groups and intubation conditions were scored as good in most children. During the early induction phase, an increase in SAP and HR was observed in the three groups. Changes were maximal at two min after the beginning of induction in the three groups. SAP and HR values were back to baseline values at the time of tracheal intubation. In conclusion, the addition of nitrous oxide to a high sevoflurane concentration decreases the time to loss of eyelash reflex, tends to reduce the incidence of excitement and is not associated with an increased incidence of respiratory complications even in patients with obstructive airway. PMID- 10712711 TI - A comparison of halothane and isoflurane for gaseous induction of anaesthesia in infants. AB - Sixty-four ASA 1 and 2 infants between the ages of 44 weeks postconceptual age and one year presenting for routine, elective surgery were randomly anaesthetized with either 3% halothane in oxygen (Group H) or 5% isoflurane in oxygen (Group I). Patients in Group I took a mean (SD) time of 70.1(13.6) s to loss of eyelash reflex and 80.0 (13.5) s to tolerating the face mask, compared with 80.2 (17.7) s and 93.4 (20.5) s in Group H (P=0.028 and 0.0072, respectively). There were no significant differences between the groups for preinduction or induction state, lowest oxygen saturation, or the incidence of airway related complications or interventions. This study demonstrates that 5% isoflurane in oxygen induces anaesthesia in infants more quickly than 3% halothane in oxygen, without any increase in the incidence or severity of airway-related complications. PMID- 10712712 TI - The effect of ketamine on 0.25% and 0.125% bupivacaine for caudal epidural blockade in children. AB - Forty boys aged from one to five years undergoing orchidopexy were randomly allocated to receive one of two solutions for caudal epidural injection. Group A received 1 ml.kg-1 of 0.125% bupivacaine with ketamine 0.5 mg.kg-1 and Group B received 1 ml.kg-1 of bupivacaine 0.25% with ketamine 0.5 mg.kg-1. Postoperative pain was assessed by means of a modified Objective Pain Score and analgesia was administered if this score exceeded four. The median duration of caudal analgesia was 8 h in Group A compared with 9.5 h in Group B (P<0.05). The time taken to recover the ability to walk was a median of two h in Group A and three h in Group B (P<0.05). There were no differences between the groups in the incidence of urinary retention or postoperative sedation. PMID- 10712713 TI - The influence of breathing system filters on paediatric capnography. AB - Breathing system filters are in common use during paediatric anaesthesia. Expired gas sampling from the patient side of these filters may contaminate and saturate the sampling line, while sampling from the machine side may cause underestimation of end-tidal carbon dioxide (PECO 2). The aim of this investigation was to elucidate the degree of underestimation of PECO 2 induced by sampling from the machine side of a breathing system filter. Ten spontaneously breathing children and ten children receiving mechanical ventilation under general anaesthesia were studied. PECO 2 was higher at the patient side of the filter in both ventilated and spontaneously breathing groups (P<0.002 for each). The bias in measuring at the machine side of the filter was significantly greater in the spontaneously breathing children as compared with the mechanically ventilated children (-1.8 vs -0.7 kPa; P<0.004). PMID- 10712714 TI - Evaluation of the efficiency of heat and moisture exchangers during paediatric anaesthesia. AB - This study evaluates the efficiency of heat and moisture exchangers (HMEs) in allowing adequate humidification and warming during anaesthesia in children. Eighteen paediatric patients undergoing anaesthesia were divided into two groups: group A ten patients: infants up to 10 kg-->Hygrobaby HME; group B 8 patients: children above 10 kg-->Hygroboy HME. The following parameters were evaluated: body temperature (bT), room temperature (rT), fresh gas temperature, HME warm-up time, inspired and expired gases temperature and humidity, conserving efficiency, and duration of anaesthesia. Gas temperatures were recorded by means of a recorder fitted with four thermal probes. Humidity values were mathematically derived. The correlation between efficiency and rT, bT, and fresh gas temperature was computed. In both groups the inspired gases temperatures were below 30 degrees C. Inspired absolute humidity was never more than 28 mgH2O.l(-1). The conserving efficiency was good (0.93 in both groups). A positive correlation was found between efficiency and fresh gas temperature. HMEs did not meet the minimum standards for humidity and heating during anaesthesia in children, although their conserving efficiency was found to be satisfactory. PMID- 10712715 TI - Fibreoptic bronchoscopy in sedated infants facilitated by an airway endoscopy mask. AB - Fibreoptic bronchoscopy (FB) is frequently associated with a decline in PaO2, whose degree and duration can be substantial especially in infants. The effect of a face mask, which allows the administration of 100% oxygen and continuous positive airway pressure during FB, on the incidence and severity of hypoxaemia was studied in thirty-one consecutive infants. Sedation was provided by intravenous propofol titrated to allow patient comfort. A transient fall in SpO2 <95% was recorded in 6/31 patients during endoscopy of the upper airway (lasting 1.6+/-1.1 min) and in 11/31 patients during endoscopy of the lower airways (lasting 1.4+/-1.1 min). Capillary blood gas analysis before and after endoscopy of the lower airways demonstrated an increase in the PCO2 6.4+/-1.3 to 7.3+/-1.4 kPa (49+/-10 to 56+/-11 mmHg). The risk of hypoxaemia in sedated infants breathing spontaneously is low when 100% oxygen and continuous positive airway pressure are administered during FB PMID- 10712716 TI - Open heart surgery; pump prime effects and cerebral arteriovenous differences in glucose, lactate and ketones. AB - This study included 17 young children, who were operated with cardioplumonary bypass for congenital heart defects and were cooled to 20 degrees C or 25 degrees C. No glucose, except for the pump prime solution, was administered during surgery. Samples of arterial blood, cerebral venous blood from the jugular bulb and mixed venous blood from the bypass circuit were obtained and analysed for concentrations of glucose, lactate and ketones as well as oxygen saturation. The prime content of lactate significantly contributed to the arterial lactate concentrations, which together with the cerebral arteriovenous (A-V) lactate differences remained elevated throughout the bypass period. The prime content of glucose had less influence on the arterial concentrations and these did not increase until the rewarming period, when indications of gluconeogenesis from lactate were found. Arterial ketone concentrations also increased during rewarming in parallel with significant cerebral uptake of ketones. The lowest cerebral A-V glucose, lactate and oxygen saturation differences were found at the target minimum temperature and this effect was significantly more pronounced in the patients cooled to 20 degrees C. PMID- 10712717 TI - Use of intravenous ketamine-midazolam association for pain procedures in children with cancer. A prospective study. AB - We evaluated the safety and efficacy of midazolam-ketamine association to control pain induced by diagnostic procedures in paediatric oncology patients. 226 procedures were carried out in 92 patients aged three days to 18 years. Drugs were given i.v. by an anaesthesiologist. Midazolam dose was 25 microg.kg-1 and ketamine 0. 5 to 2 mg.kg-1, depending on number and invasiveness of procedures. The mean dose of ketamine was 1 mg.kg-1. Mean duration of sedation was ten min. No complication was observed and analgesia was considered satisfactory in 89 out of 92 patients. These results indicate that midazolam-ketamine is a safe and effective association in pain management for paediatric oncology patients and efficiently induces brief unconscious sedation with analgesia. PMID- 10712718 TI - An epidural catheter introducer in children. AB - Usefulness of an epidural catheter introducer was tested in paediatric epidural anaesthesia. We tried to place an epidural catheter in 100 infants and children. When catheter insertion was difficult, an epidural catheter introducer, which was made of a piece of 6-Fr suction tubing, was utilized. The introducer is threaded over the epidural catheter down to the hub orifice of the epidural needle. The catheter can then be advanced into the epidural space through this introducer. In this way, we were able to place the catheter in 94 percent of patients. Even a simple introducer is effective in passing the epidural catheter into the epidural space in infants and children. Perhaps manufacturers should provide threading devices with catheters or epidural needles. PMID- 10712719 TI - A rare case of upper airway obstruction in an infant caused by basal encephalocele complicating facial midline deformity. AB - A four-month-old male infant with basal encephalocele of the transsphenoidal type presented with upper airway obstruction and facial midline deformity, including cleft lip, cleft palate, hypertelorism and exophthalmos. Basal encephalocele is a rare disease, and usually not detectable from the outside. In this case, initially the cause of an upper airway obstruction was considered to be posterior rhinostenosis, and posterior rhinoplasty with inferior nasal conchectomy was scheduled. However, in preoperative examination, computed tomography (CT) and magnetic resonance imaging (MRI) revealed a bony defect in the sphenoidal bone and a cystic mass in communication with cerebrospinal fluid, herniating into the nasal cavity through the bony defect. The mass was diagnosed as a transsphenoidal encephalocele, the scheduled operation cancelled, and tracheostomy performed for airway management. The possibility of basal encephalocele should be considered in the case of upper airway obstruction with facial midline deformity. PMID- 10712720 TI - A wandering nasal prong-a thing of risks and problems. AB - We describe an unusual complication of nasal continuous positive airway pressure (nCPAP) ventilation in a preterm low birth weight neonate being weaned from respiratory support. The tube used to administer nasal CPAP became dislodged from its metal connector whilst in the nasopharynx and slipped into the stomach. After waiting eight days the tube showed no signs of passing spontaneously through the gastrointestinal tract and retrieval was then successfully achieved by means of a 3.5 mm paediatric fibreoptic bronchoscope without complication. PMID- 10712721 TI - Can a dose of 2microg.kg(-1) caudal clonidine cause respiratory depression in neonates? AB - A case of multiple life-threatening postoperative apnoeas in a term neonate undergoing inguinal herniorrhaphy and orchidopexy who received light inhalation anaesthesia combined with caudal block with 1 ml.kg-1 ropivacaine 0.2% plus 2 microg.kg-1 clonidine is reported. The patient showed no apparent risk factors for postanaesthetic apnoea. Oxycardiorespirography five days after surgery only showed minor abnormalities. Clonidine though administered caudally in the usual dose of 2 microg.kg-1 appeared to be the most likely cause for postanaesthetic apnoea in this neonate. PMID- 10712722 TI - One lung anaesthesia for video assisted thoracoscopic lung biopsy in a paediatric patient. PMID- 10712723 TI - Use of intravenous immunoglobulins as an adjunct in the conservative management of chylothorax. AB - Two children who developed chylothorax after surgery for congenital heart disease are presented. The conservative management of chylothorax is reviewed and the use of immunoglobulins in the treatment of sepsis is discussed. One patient survived. PMID- 10712724 TI - Tracheal tube insertion through laryngeal mask airway in paediatric patients. PMID- 10712725 TI - Resistance to vecuronium after immobilization. PMID- 10712726 TI - Tubeless anaesthesia for microlaryngoscopy. PMID- 10712727 TI - Filters and Ayre's T-piece. PMID- 10712728 TI - Mathematical modeling as a tool to investigate the design and operation of the zymotis packed-bed bioreactor for solid-state fermentation. AB - Zymotis bioreactors for solid-state fermentation (SSF) are packed-bed bioreactors with internal cooling plates. This design has potential to overcome the problem of heat removal, which is one of the main challenges in SSF. In ordinary packed bed bioreactors, which lack internal plates, large axial temperature gradients arise, leading to poor microbial growth in the end of the bed near the air outlet. The Zymotis design is suitable for SSF processes in which the substrate bed must be maintained static, but little is known about how to design and operate Zymotis bioreactors. We use a two-dimensional heat transfer model, describing the growth of Aspergillus niger on a starchy substrate, to provide guidelines for the optimum design and operation of Zymotis bioreactors. As for ordinary packed-beds, the superficial velocity of the process air is a key variable. However, the Zymotis design introduces other important variables, namely, the spacing between the internal cooling plates and the temperature of the cooling water. High productivities can be achieved at large scale, but only if small spacings between the cooling plates are used, and if the cooling water temperature is varied during the fermentation in response to bed temperatures. PMID- 10712729 TI - Production of Sm37-GAPDH, a major therapeutical target in human schistosomiasis. AB - Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a key enzyme in the glycolytic metabolism and the production of energy. This probably explains why GAPDH was evidenced as a major therapeutical target in several parasitic diseases; either as a vaccine candidate or as a target for chemotherapeutic treatments. Schistosoma mansoni GAPDH (Sm37-GAPDH) is one of the main schistosome vaccine candidates. The production of recombinant Sm37-GAPDH is essential to evaluate the ability of this molecule to induce protective immunity in animals and possibly in humans. The cDNA encoding Sm37-GAPDH has been cloned and sequenced. In addition, five B cell (including the major B-cell epitope Sm35-5) and two T cell epitopes have been localized on the molecule. Different expression systems have been evaluated in respect with the production yield and the GAPDH enzymatic activity. Some of them have led to either a high production of insoluble material (E. coli) or to an inactive enzyme (Pischia pastoris). The present article describes the production setting of rSm37-GAPDH using the baculovirus-insect cell system. Large amounts of soluble rSm37-GAPDH with enzymatic activity were obtained. Most sera from individuals living in an area endemic for S. mansoni recognised the rSm37 molecule and inhibited its catalytic activity. PMID- 10712730 TI - Bioartificial kidney. I. Theoretical analysis of convective flow in hollow fiber modules: application to a bioartificial hemofilter. AB - Analytical expressions describing convective flow in a continuous arteriovenous hollow fiber hemofilter were developed. In the lumen of the hollow fiber membrane, existing analytical expressions were applied to describe velocity profiles and pressure. For flow in the shell (the extracapillary space separating the fibers), analytical expressions for the radial and axial velocity profiles and pressure distribution were derived by first finding the stream function. The expressions are based on a similarity solution. Previous analyses of ultrafiltration have either ignored osmotic pressure or assumed constant shell pressure. In this paper, the axial variation in lumen pressure, shell pressure, and osmotic pressure were accounted for. The predicted filtration rates agree well with the experimental results. This flow model is general enough to describe flow in hollow fiber membrane systems employed as bioreactors (e.g., for cell cultures and as bioartificial organs) and as separators (e.g., ultrafiltration and microfiltration) operating in the open-shell mode. The results were applied to determine the design of an optimally functioning bioartificial hemofilter for use ex vivo or in vivo. PMID- 10712731 TI - Bioartificial kidney. II. A convective flow model of a hollow fiber bioartificial renal tubule. AB - An existing model for volume transport in the rat proximal tubule was modified and applied to a bioartificial renal tubule. The predicted volume transport agrees well with experimental data. The volume transport model was coupled to the analytic solutions of flow in the bioartificial renal tubule bioreactor, operated in the open-shell mode with perfusion in both the lumen and surrounding shell. The results suggest that the performance of a multifiber bioreactor can be improved by controlling shell inlet conditions and fiber spacing. These results have important implications for the design and analysis not only for the bioartificial renal tubule bioreactor but also for the general case of hollow fiber bioreactors operated in the open-shell mode with perfusion in both the lumen and surrounding shell. PMID- 10712732 TI - Effect of biomass concentration and mycelial morphology on fermentation broth rheology. AB - The effect of biomass concentration and mycelial morphology on fungal fermentation broth rheological properties has been investigated. In previous work it had been shown that commonly used rheological parameters, such as the power law consistency and flow behavior indices, could be correlated successfully with the broth biomass concentration and clump morphological parameters of roughness and compactness. More recent work on a broader range of data showed a correlation between roughness and compactness; consequently, it was not correct to use both of these morphological variables simultaneously in rheological parameter correlations. Furthermore, earlier correlations were only made using clump morphological parameters, as clumps were found to be around 90% of the biomass in batch fermentations. In the present work it was found that the percentage of clumps fell to around 30% to 40% of a sample during the later stages of fed-batch fermentations. No clear relationship between the flow behavior index and biomass concentration was found, at least for those phases of the fermentation in which the viscosities were high enough for the rheology to be characterized by a disk turbine rheometer. The mean value of the flow behavior index was found to be 0.35 +/- 0.1 (standard deviation) throughout both batch and fed-batch fermentations, although some significant deviations from this value were observed early and very late in the fermentations. Correlations for the consistency index, measured using a disk turbine rheometer, were based on the biomass concentration and the mycelial size (represented by the mean projected area or the mean maximum dimension of all the mycelia). These correlations were reasonably successful for both fed-batch and batch fermentations. The correlation using the mean maximum dimension was preferred to that using the mean projected area, as the former is independent of magnification. The proposed correlation is: where K is the consistency index (Pa. s(n>)), C(m) is the biomass concentration as dry cell weight (g L(-1)), and D is the mean maximum dimension (microm). It should be noted that small changes in the exponent on the biomass concentration (alpha) may dramatically affect any predictions. Consequently, caution in the use of this correlation (and that based on mean projected area) is advocated, although its accuracy may be suitable for operational or design purposes. PMID- 10712733 TI - Modeling of eicosapentaenoic acid (EPA) production from Phaeodactylum tricornutum cultures in tubular photobioreactors. Effects of dilution rate, tube diameter, and solar irradiance. AB - A model for the prediction of eicosapentaenoic acid (EPA) productivity from Phaeodactylum tricornutum cultures that takes into account the existence of photolimitation and photoinhibition of growth under outdoor conditions is presented. The effects of the external irradiance on the culture surface, the average irradiance inside the culture, and the light regime at which the cells are exposed on pigments and EPA content are studied. The chlorophyll content decreases exponentially with the average irradiance, whereas the carotenoids content increases linearly with the external irradiance due to a higher extension of photoinhibition. A decrease in the fatty acid content of the biomass with irradiance on reactor surface is observed when photoinhibition becomes relevant. The average irradiance within the culture mainly influenced the fatty acid profile of the biomass. As the average irradiance becomes higher, percentages of saturated and monounsaturated fatty acids decrease, increasing the portion of EPA. By taking into account the different relationships among pigment and EPA content with the irradiance, the variation in EPA productivity over the year can be simulated as a function of average and external irradiance. For the two photobioreactors employed the maximum EPA productivity is attained in spring and fall (30 mg L(-1) day(-1) for tube diameter 0. 06 m and 50 mg L(-1) day(-1) for tube diameter 0.03 m). In winter, the biomass productivity is limited by low light availability although the EPA content is maximum. In summer, the biomass productivity is higher although the EPA content diminished by photoinhibition; the higher the dilution rate, the lower the minimum. Thus, the conditions that increase the biomass productivity and the polyunsaturated fatty acids content are in opposition, the optimum being reached by operating under photolimitation with high growth rates in order to produce a high proportion of polyunsaturated fatty acids. PMID- 10712734 TI - Preparation of Thermus thermophilus holo-chaperonin-immobilized microspheres with high ability to facilitate protein refolding. AB - Carboxylated poly(styrene/acrylamide) (P(St/AAm)-H) microspheres with different acrylamide contents were prepared by emulsifier-free emulsion polymerization. Thermus thermophilus holo-chaperonin (cpn) was covalently immobilized onto these microspheres with high yield. The T. thermophilus holo-cpn-immobilized microspheres were used for refolding of guanidine hydrochloride (Gdn-HCl) denatured enzymes and showed sufficiently high ability to facilitate refolding of Leuconostoc mesenteroides glucose-6-phosphate dehydrogenase (G6PD) and pig heart lactate dehydrogenase (LDH) at 30 degrees C and Bacillus stearothermophilus LDH at 60 degrees C. The specific ability of T. thermophilus holo-cpn-immobilized microspheres increased with increasing immobilization amount and reached plateau at around 10-15 mg/m(2). When the immobilization amounts of T. thermophilus holo cpn were approximately 10 mg/m(2), the microspheres retained sufficiently high ability to facilitate protein refolding during repeated use. Therefore, the P(St/AAm)-H microspheres on which approximately 10 mg/m(2) of T. thermophilus holo-cpn is immobilized are very effective for refolding of various (Gdn-HCl) denatured enzymes over a wide temperature range. PMID- 10712735 TI - Purification of lysozyme using ultrafiltration. AB - This article examines the separation of lysozyme from chicken egg white by ultrafiltration with 25 kDa and 50 kDa MWCO polysulfone membranes. The effects of pH, system hydrodynamics, feed concentration, and transmembrane pressure on permeate flux, lysozyme transmission, purification factor, and productivity have been discussed. With both types of membranes, higher permeate flux and lysozyme transmission were observed at higher pH. Higher lysozyme purity was generally obtained with the 25 kDa MWCO membrane. Purity of lysozyme decreased when the feed concentration was increased. With the 50 kDa MWCO membrane permeate flux, productivity and the purity of lysozyme were found to increase with increase in transmembrane pressure. The possibility of using a two-step ultrafiltration process for achieving high productivity along with high purity of lysozyme was also investigated. PMID- 10712736 TI - Effect of substrate and cellulase concentration on simultaneous saccharification and fermentation of steam-pretreated softwood for ethanol production. AB - Economic optimization of the production of ethanol by simultaneous saccharification and fermentation (SSF) requires knowledge about the influence of substrate and enzyme concentration on yield and productivity. Although SSF has been investigated extensively, the optimal conditions for SSF of softwoods have yet not been determined. In this study, SO2-impregnated and steam-pretreated spruce was used as substrate for the production of ethanol by SSF. Commercial enzymes were used in combination with the yeast Saccharomyces cerevisiae. The effects of the concentration of substrate (2% to 10% w/w) and of cellulases (5 to 32 FPU/g cellulose) were investigated. SSF was found to be sensitive to contamination because lactic acid was produced. The ethanol yield increased with increasing cellulase loading. The highest ethanol yield, 68% of the theoretical based on the glucose and mannose present in the original wood, was obtained at 5% substrate concentration. This yield corresponds to 82% of the theoretical based on the cellulose and soluble glucose and mannose present at the start of SSF. A higher substrate concentration caused inefficient fermentation, whereas a lower substrate concentration, 2%, resulted in increased formation of lactic acid, which lowered the yield. Compared with separate hydrolysis and fermentation, SSF gave a higher yield and doubled the productivity. PMID- 10712737 TI - Directed evolution of a novel N-carbamylase/D-hydantoinase fusion enzyme for functional expression with enhanced stability. AB - Bifunctional enzymes find a wide application as a monitoring facility and a potential biocatalyst in molecular biology and biotechnology. Recombination of natural enzymes to a bifunctional fusion offers valuable tools, but the functional and structural instability of artificial fusion enzymes remains to be solved. Based on structural traits of microbial D-hydantoinase, we attempted to construct a bifunctional N-carbamylase/D-hydantoinase fusion enzyme that would be useful for the synthesis of nonnatural D-amino acids in a concerted fashion. The bifunctional ability of D-hydantoinase, as a fusion partner, was noticeable, but the resulting fusion enzyme was subjected to serious proteolysis in vivo, as generally encountered in the expression of large the multidomain polypeptide in E. coli. In an effort to improve the structural instability imposed by artificial linear fusion, directed evolution of the fusion enzyme was performed using DNA shuffling with a consensus primer to maintain a crucial domain for the enzyme activity. The evolved fusion enzyme, F11, was selected after repeated rounds, and this enzyme was found to show sixfold increased performance in the production of D-amino acid compared with the parent fusion enzyme, which was mainly due to the enhanced structural stability of the evolved fusion enzyme. This result is an example showing that directed evolution of the linearly fused polypeptide may broaden the opportunity to generate a fusion enzyme with greater potential. PMID- 10712738 TI - Synthesis of bioprocesses using physical properties data. AB - The aim of this article is to illustrate and evaluate a synthesis procedure which has been extended to tackle bioprocesses. Physical property information is used to screen candidate units thereby reducing the size of the synthesis problem. In this way, only units which exploit large property differences between components in a stream are selected. This is important for bioprocesses because of the large number of components and wide range of unit operations which are available. The screening technique and bioprocess-unit-design methodologies have been incorporated within an implicit enumeration algorithm which was developed for chemical process synthesis and is implemented in Java programming language. An important advantage is the ability of the bioprocess synthesis software to generate a ranked list of flowsheets which may subsequently be analyzed in more detail. Two case studies are used to evaluate the bioprocess-synthesis technique. The first system involves a product which is secreted from the host organism. The second has significantly different characteristics in that the product is intracellular and forms inclusion bodies. The latter case study, in particular, is a large synthesis problem with 12 unit operations and 20 contaminant compounds. The results show that the synthesis methodology identifies a set of economically optimal flowsheets in a reasonable computational time which demonstrates its ability to deal with large synthesis problems. Using the synthesis methodology we can generate bioprocesses which are optimal in a system wide, rather than unit-by-unit, sense. PMID- 10712739 TI - Characterization of kinetics and thermostability of Acremonium strictum glucooligosaccharide oxidase. AB - The kinetic and thermostability properties of a glucooligosaccharide oxidase from Acremonium strictum were determined. This enzyme produces only maltobionic acid from maltose. It is most active at pH 9 to 10.5, and is most stable at pH 6.5. Values of both K(M) and V(max) on maltose are highest at pH 10. The highest values of K(M) and V(max) occur with glucose, maltopentaose, and maltoheptaose, whereas the lowest values of K(M) are with maltotriose and of V(max) are with maltohexaose. Values of K(M) with any substrate and at any pH are always substantially above 1 mM. Activation energies for catalysis and thermoinactivation are 23 kJ/mol and 421 kJ/mol, respectively. The N-terminal sequence is not homologous with any other oxidase, but has some homology with other proteins having different functions. These unusual properties suggest that glucooligosaccharides may not be the primary substrates of this enzyme. PMID- 10712740 TI - DNA repair and gene therapy: implications for translational uses. AB - Gene therapy has been proposed to have implications in the treatment of cancer. By genetically manipulating the hematopoietic stem cell compartment with genes that confer resistance to chemotherapeutic agents, the dose escalation that is necessary to effectively treat the cancers could potentially be achieved. DNA repair genes are some of the potential candidates to confer increased resistance to chemotherapeutic agents. Although initial focus in this area has been on the direct reversal protein (MGMT), its protective ability is limited to those agents that produce O(6)-methylGuanine cross-links-agents that are not extensively used clinically (e.g., nitrosoureas). Furthermore, most alkylating agents attack more sites in DNA other than O(6)-methylGuanine, such that the protections afforded by MGMT may prevent the initial cytotoxicity, but at a price of increased mutational burden and potential secondary leukemias. Therefore, some of the genes that are being tested as candidates for gene transfer are base excision repair (BER) genes. We and others have found that overexpression of selective BER genes confers resistance to chemotherapeutic agents such as thiotepa, ionizing radiation, bleomycin, and other agents. As these "proof of concept" analyses mature, many more clinically relevant chemotherapeutic agents can be tested for BER protective ability. PMID- 10712741 TI - An interesting state of affairs in genetic toxicology. PMID- 10712743 TI - Polymorphisms of the GSTP1 and GSTM1 genes and PAH-DNA adducts in human mononuclear white blood cells. AB - Glutathione S-transferases (GSTs) are an important part of the protection system against a wide range of potentially harmful chemical compounds. GSTP1 and GSTM1 are mainly involved in detoxification reactions of PAH carcinogenic intermediates produced by cytochrome P450 (CYP). Polymorphism of the GST genes may influence the level of carcinogen-DNA adducts in human tissues and be associated with individual susceptibility to carcinogens. In this study, we examined the effect of common polymorphism in exon 5 (105Ile --> Val) of the GSTP1 gene, alone and in combination with GSTM1-deletion polymorphism, on the level of PAH-DNA adducts measured by (32)P-postlabeling assay in mononuclear white blood cells collected in winter and in summer from a total of 170 healthy volunteers. When GSTP1 genotypes alone were compared, no statistically significant differences in adduct levels were found. However, smokers with GSTM1(null)/GSTP1-AG or -GG combined genotype showed significantly higher adduct levels in summer than carriers of other GSTM1/GSTP1 combinations (5.60 +/- 5.10 vs. 3.45 +/- 4. 28/10(8) nucleotides, P = 0.015). Among smokers carrying GSTP1-AG or -GG genotype, individuals with GSTM1(null) genotype had a significantly higher level of adducts in summer than subjects with GSTM1(+) genotype (5.60 +/- 5.10 vs. 1.82 +/- 1.98/10(8), P = 0.002) and GSTM1(null)/GSTP1-AA genotype carriers (5.60 +/- 5.10 vs. 4.13 +/- 5.84/10(8), P = 0.03). When adduct levels measured either in winter or in the nonsmoker group were considered, no influence of GSTM1/GSTP1 genotypes was found. Our data show that the combined GSTM1 and GSTP1 genetic polymorphisms may modulate PAH-DNA adduct levels in mononuclear WBCs from individuals exposed to specific carcinogenic compounds, e.g., tobacco smoke, in relatively lower exposure environmental conditions (i.e., in summer). PMID- 10712742 TI - Differential modulation of the genotoxicity of food carcinogens by naturally occurring monomeric and dimeric polyphenolics. AB - Naturally occurring dimeric polyphenolics and their gallate esters were isolated from grape seeds, almond fruits, and apple skin, and their ability to modulate the mutagenicity of food carcinogens was studied in the Ames test, and compared to that of the monomeric green tea flavonols, (+)-catechin and (-)-epicatechin. Neither the monomeric nor the dimeric polyphenols and their galloylated derivatives influenced the mutagenic activity elicited by the indirectly acting food carcinogens benzo[a]pyrene and 2-amino-3-methylimidazo-[4,5-f]quinoline (IQ), in the presence of a hepatic activation system derived from Aroclor 1254 treated rats; the only exception was the B7 dimer, which, at concentrations above 1 microM, suppressed the mutagenicity of IQ. None of the polyphenolics modulated the mutagenic activity elicited by the directly acting carcinogen N'-methyl-N' nitro-nitrosoguanidine (MNNG). In contrast, all the dimeric polyphenols and the galloylated metabolites, at concentrations over 1 microM, potentiated the mutagenic activity induced by the indirectly acting carcinogen N nitrosopyrrolidine, in the presence of an activation system derived from isoniazid-treated rats. In conclusion, dimeric polyphenols and galloylated derivatives of plant origin are unlikely to influence the initiation stage of the carcinogenicity of chemicals through mechanisms that involve inhibition of their cytochrome P450-mediated bioactivation or scavenging of the reactive, genotoxic intermediates. PMID- 10712744 TI - Mutation spectra of the drinking water mutagen 3-chloro-4-methyl-5-hydroxy-2(5H) furanone (MCF) in Salmonella TA100 and TA104: comparison to MX. AB - The chlorinated drinking water mutagen 3-chloro-4-methyl-5-hydroxy-2(5H)-furanone (MCF) occurs at concentrations similar to or greater than that of the related furanone 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX). MCF and MX differ structurally only by replacement of a 3-methyl in MCF with a 3 dichloromethyl in MX; yet, MCF is significantly less mutagenic than MX and produces different adducts when reacted with nucleosides or DNA. To explore further the effects that these structural differences might have on the biological activity of MCF and MX, we determined the mutation spectra of MCF in Salmonella strains TA100 and TA104 and of MX in strain TA104; the spectrum of MX in TA100 had been determined previously. In TA100, which presents only GC targets for mutagenesis, MCF induced primarily (75%) GC --> TA transversions, with most of the remaining revertants (20%) being GC --> AT transitions. This spectrum was not significantly different from that of MX in TA100 (P = 0.07). In TA104, which presents both GC and AT targets, MCF induced a lower percentage (57%) of GC --> TA transversions, with most of the remaining revertants (33%) being AT --> TA transversions. In contrast, MX induced almost only (98%) GC --> TA transversions in TA104, with the remaining revertants (2%) being AT --> TA transversions. Thus, almost all (98%) of the MX mutations were targeted at GC sites in TA104, whereas only 63% of the MCF mutations were so targeted. These results are consistent with the published findings that MX: (1) forms an adduct on guanosine when reacted with guanosine, (2) induces apurinic sites in DNA, and (3) forms a minor adduct on adenosine when reacted with adenosine or DNA. The results are also consistent with evidence that MCF forms adenosine adducts when reacted with adenosine. Our results show that the replacement of the 4-methyl in MCF with a 4-dichloromethyl to form MX not only increases dramatically the mutagenic potency but also shifts significantly the mutagenic specificity from almost equal targeting of GC and AT sites by MCF to almost exclusive targeting of GC sites by MX. Environ. Mol. Mutagen. 35:106-113, 2000 Published 2000 Wiley-Liss, Inc. PMID- 10712745 TI - Arsenic[III] and heavy metal ions induce intrachromosomal homologous recombination in the hprt gene of V79 Chinese hamster cells. AB - In the present study the carcinogenic metal ions Cd[II], Co[II], Cr[VI], Ni[II], and Pb[II], as well as As[III], were examined for their ability to induce intrachromosomal homologous and nonhomologous recombination in the hprt gene of two V79 Chinese hamster cell lines, SPD8 and Sp5, respectively. With the exception of Pb[II], all of these ions enhanced homologous recombination, the order of potency being Cr>Cd>As>Co>Ni. In contrast, Cr[VI] was the only ion to enhance recombination of the nonhomologous type. In order to obtain additional information on the mechanism of recombination in the SPD8 cell line, individual clones exhibiting metal-induced recombination were isolated, and the sequence of their hprt gene determined. These findings confirmed that all recombinogenic events in this cell line were of the homologous type, involving predominantly a chromatid exchange mechanism. The mechanisms underlying the recombination induced by these ions are discussed in relationship to their genotoxicity, as well as to DNA repair and replication. Induced recombination may constitute a novel mechanism for induction of neoplastic disease. PMID- 10712746 TI - Phenobarbital increases DNA adduct and metabolites formed by ochratoxin A: role of CYP 2C9 and microsomal glutathione-S-transferase. AB - Ochratoxin A (OTA), a mycotoxin that induces nephrotoxicity and urinary tract tumors, is genotoxic and can be metabolized not only by different cytochromes P450 (CYP) but also by peroxidases involved in the arachidonic cascade, although the exact nature of the metabolites involved in the genotoxic process is still unknown. In order to establish the relation between OTA genotoxicity and the formation of metabolites, we chose three experimental models: kidney microsomes from rabbit, human bronchial epithelial cells, and microsomes from yeast that specifically express the human cytochrome P450 2C9 or 2B6 genes. OTA-DNA adducts were analyzed by (32)P postlabeling and the OTA derivatives formed were isolated by HPLC after incubation of OTA in the presence of: (1) kidney microsomes from rabbit pretreated or not with phenobarbital (PB); (2) human pulmonary epithelial cells simultaneously pretreated (or not) with PB alone or in the presence of ethacrynic acid (EA); (3) microsomes expressing CYP 2B6 and 2C9. PB pretreatment significantly increased DNA adducts formed after OTA treatment, both in the presence of kidney microsomes and bronchial epithelial cells, and induced the formation of new adducts. Ethacrynic acid, which inhibits microsomal glutathione S-transferase, reduced DNA adduct level. DNA adducts were detected when OTA were incubated with microsomes expressing human CYP 2C9 but not with those expressing CYP 2B6. Several metabolites detected by HPLC were increased after PB treatment. Some of them could be related to DNA-adduct formation. In conclusion, OTA biotransformation, enhanced by PB pretreatment, increased DNA-adduct formation through pathways involving microsomal glutathion-S-transferase and CYP 2C9. PMID- 10712747 TI - Detection of specific DNA lesions by a combination of comet assay and FISH in plants. AB - We have studied comet formation on Vicia faba nuclei embedded in agarose and treated with the endonucleases DNase I (to produce SSBs or DSBs at random sites), FokI (to produce DSBs preferentially within FokI repeats), or EcoRI (to produce DSBs at random sites but not within FokI elements). DNase I-induced SSBs were detected when enzyme treatment was followed by alkaline denaturation. DSBs efficiently mediated comet formation using neutral conditions. FISH with DNA probes, detecting specific chromosomal domains such as FokI element-containing heterochromatin, NORs, or telomeres, was done on comets. The distribution of FISH signals between the head and tail of comets indicated to which degree these domains were damaged and reflected the distribution of cleavage sites for the applied restriction endonucleases within these domains. The data confirmed the expectation that the observed comet formation was based on enzyme-specific DNA breakage. PMID- 10712748 TI - Tissue-specific activities of methylated dibenzo[c,g]carbazoles in mice: carcinogenicity, DNA adduct formation, and CYP1A induction in liver and skin. AB - The potent multitissue carcinogen 7H-dibenzo[c,g]carbazole and nine methylated derivatives, synthesized on the basis of the positions in the parent compound that are involved in metabolism, were tested for their ability to induce sarcomas and hepatic tumors in XVIInc/Z mice. In addition, the capacity of these compounds to induce DNA adducts in skin and liver was investigated by (32)P-postlabeling analysis after their topical administration. Induction by these compounds of cytochromes P450 of the 1A family in liver and skin was investigated and correlated to their carcinogenic potential. A clear correlation was found between the tissue specificity of DNA binding and the capacity of each compound to induce skin or liver tumors. In contrast, no direct relationship was observed between the capacity of the compounds to induce cytochromes 1A1/1A2 and the tissue specificity of carcinogenesis or DNA binding in liver or skin. The results are discussed with respect to the positions of methyl groups in the 7H dibenzo[c,g]carbazole molecule. PMID- 10712749 TI - In vivo loss of heterozygosity in T-cells of B6C3F1 Aprt(+/-) mice. AB - We have used B6C3F1 mice heterozygous at Aprt (adenine phosphoribosyltransferase) as a model to study in vivo loss of heterozygosity (LOH) in normal splenic T lymphocytes. APRT-deficient T-cells were selected in medium containing 50 microg/ml 2, 6-diaminopurine (DAP), an adenine analog that is toxic only to cells with APRT enzyme activity. DAP-resistant (DAP(r)) T-cell variants were recovered at an average frequency of 3 x 10(-5) from 21 B6C3F1 Aprt(+/-) mice. Allele specific PCR of Aprt showed that about 70% of 122 DAP(r) colonies were caused by loss of the nontargeted Aprt allele (Aprt(+)). Analysis of microsatellite markers along the length of chromosome 8 suggested that mitotic recombination, or chromosome loss, with or without duplication of the remaining chromosome are the predominant mechanisms resulting in loss of Aprt(+). DNA sequencing of Aprt RT PCR products from the DAP(r) variants that retained Aprt(+) indicated that point mutation as well as other mechanisms could cause this second class of variants. The high spontaneous frequency of in vivo Aprt LOH in mouse T-cells, mediated by LOH mechanisms that are also known to produce human cancers, suggests that the Aprt heterozygous mouse is a valid model for studying the diversity of mechanisms for in vivo somatic mutagenesis. PMID- 10712750 TI - Shooting holes in the heart: an update on TMR. PMID- 10712751 TI - Stages of change: getting to where you want to be. PMID- 10712752 TI - High blood pressure and blood clots. PMID- 10712753 TI - New safety recommendations for use of cisapride (Propulsid). PMID- 10712754 TI - Ask the doctor. I had an EKG done a few weeks ago, and the reading came back "probable inferior myocardial infarction of indeterminate age." I am, to the best of my knowledge, perfectly healthy at the age of 72. Now my doctor wants to do a bunch of tests on me. Should I go along? PMID- 10712755 TI - Stroke. Preemptive strikes. PMID- 10712756 TI - Research. Studies at Harvard. PMID- 10712757 TI - Cancer. Staying fit for survival. PMID- 10712758 TI - Lyme disease vaccine. Safe. Effective. But necessary? PMID- 10712759 TI - By the way, doctor... I've started drinking soy milk instead of cow's milk because I've developed an intolerance to lactose. I know there are so-called phytoestrogens in soy products and that they are probably of some benefit to me. However, is there a problem for my husband? PMID- 10712760 TI - Personality disorders - part II. PMID- 10712761 TI - Gender bias in the diagnosis of personality disorders. PMID- 10712762 TI - Expressed emotion and borderline personality. PMID- 10712763 TI - What is the psychiatric significance of loneliness? PMID- 10712765 TI - Germ welfare. PMID- 10712764 TI - Kidney stones. PMID- 10712766 TI - Exercise and older men: it's never too late. PMID- 10712767 TI - On call. Last week I received a mailing from a Canadian company that did not support American drugs like Hytrin and Proscar in dealing with enlarged prostates. There were about 30 testimonials in favor of its product. The company says that zinc is an essential ingredient to shrink the prostate. It further states the need for pyridoxine and certain amino acids, Serenoa, repens (saw palmetto), serrulata, Panax extract, and hydrangea extract. Comments from various European magazines are also cited... PMID- 10712768 TI - Medical memo. Declining male births in industrial countries. PMID- 10712769 TI - HRT 2000: pause for thought. PMID- 10712770 TI - Intestinal disorders. Relieving irritable bowel syndrome. PMID- 10712771 TI - Herbal medicine. Black cohosh: the woman's herb? PMID- 10712772 TI - Vaccinations: not just for children. PMID- 10712773 TI - By the way, doctor. I never know whether to use heat or ice on a strained muscle or my aching back. Are there any guidelines I should follow? PMID- 10712774 TI - [Construction of prokaryotic expression vector of hALR and its expression in E.Coli]. AB - OBJECTIVE: To construct prokaryotic expression vector of hALR and express rhALR in E.Coli. through PCR and recombinant gene technology. METHODS: The coding region with Nde I and BamH I sites of hALR was obtained from pUC19-hALR constructed with PCR method. PCR product and plasmid pET11a were digested by corresponding endonucleases, respectively. The fragments cut were ligated by T(4) DNA ligase to gain recombinant expression vector. The recombinant plasmid pET11a hALR was electrotransformed into BL(21) (DE(3)) strain. The rhALR was expressed in the bacteria under induction of IPTG. RESULTS: Endonucleases digesting and DNA sequening confirmed that the coding region of hALR was correctly inserted into the vector. The rhALR was successfully expressed in E.Coli. Its MW. With 1. 5A10(4) was in correspondence with theoretic value and its amount is 30% of total bacteria protein. It existed not only in supernatant but also in precipitation of broken bacteria. CONCLUSION: The successes in construction of expressive vector of hALR and in expression of rhALR in E.Coli. make it possible to study further on its biological functions and antibody preparation. PMID- 10712775 TI - [Cloning and sequence analysis of human genomic DNA of augmenter of liver regeneration hepatitis]. AB - OBJECTIVE: To clone the human genomic DNA of augmenter of liver regeneration (ALR) and specify the intron-exon structure. METHODS: Using human ALR cDNA sequence as a reference and BLAST path as a nucleotide homology search tool, GenBank has been searched for ALR homologous genomic DNA sequence. The intron exon sequences were defined by the Breathnath-Chambon rule. RESULTS: The coding sequence of human ALR consists of 3 exons, and is similar to murine ALR genomic DNA structure. The human ALR genomic DNA is 1813 nt long and codes a protein of 125 amino acid residues. CONCLUSION: Human genomic DNA of ALR consists of 3 exons and 2 introns. PMID- 10712776 TI - [Study of HBV X protein and RMP, an RPB5 mediate protein competitively interacting with general transcription factor TF2B]. AB - OBJECTIVE: HBX is an essential viral protein that is important for HBV replication and might be a cofactor in the development of hepatocellular carcinoma. So many researchers are trying to find out the antagonistic factors of HBX. RMP, a newly cloned RNA polymerase II subunit RPB5 mediate protein, represses the transcription transactivation of HBV X protein. The purpose of this study was to elucidate the mechanism how RMP represses the transactivation of HBX. METHODS: In this report, DNA recombinant technique in vitro protein-protein binding assay (Pull-Down assay) and chloramphenicol acetyltransferase assay were used. RESULTS: The d10 domain of TF2B are responsible for binding to RMP, which is same as TF2B binding to the HBX protein. RMP specifically disrupts the binding between TF2B and HBX protein, vice versa HBX protein also disrupt the interaction of TF2B-RMP as demonstrated by the in vitro protein binding competition assay. Overexpression of RMP represses HBX transactivation on the reporters with HBX responsive cis-elements in transient transfected COS-1 cells. The repression can be rescued by overexpression of TF2B. CONCLUSION: The results clearly suggested that transcription co-activator HBX protein and corepressor RMP competitively associated with TF2B. Disrupting the interaction between transcription co repressor RMP with TF2B is probably one of the mechanisms to account for the transcriptional transactivation of HBX protein. PMID- 10712777 TI - [Expression and immunological reactivity of recombinant HCV-core protein]. AB - OBJECTIVE: To express HCV-core proteins in E.coli and to develop effective HCV core DNA-based vaccine. METHODS: The vector that expresses the highly conserved HCV core genes were constructed. The pGEX-3X HCVCore constructs contained the 1 201 ncls (1-67aa, C201), 1-402 ncls (1-134aa, C402) and 1-591ncls ( 1-197aa, C591), then expressed in E.coli cells. RESULTS: The products of HCV C201 and C402 genes were expressed as a fusion protein with glutathione-S-transferase (GST, 26kDa) whose molecular weight were 3.1 x 10(4) and 3.9 x 10(4) separately. C591 gene was not effectively expressed in E.coli. The expressed proteins were sequestered within inclusion bodies (IB) and a variety of procedures designed to minimize IB formation proved unsuccessful. The method finally adopted involved the purification of inclusion bodies followed by the solubilization, purification, and refolding of the expressed protein. The purified C402 protein was antigenically reactive with serum from chronically infected HCV patients. BALB/C mice were immunized by a subcutaneous injection of C402 protein together with Freund's complete adjuvant which produced strong anti-HCV core humoral immune responses. CONCLUSION: It is important for the study of gene vaccine to construct a certain length of HCV core gene. PMID- 10712778 TI - [Effects of sandostatin on gastric mucosal perfusion in rats with portal hypertensive gastropathy]. AB - OBJECTIVE: To observe the effects of sandostatin on gastric mucosal blood flow (GMBF) in rats with (portal hypertensive gastropathy, PHG) and its mechanisms. METHODS: Two weeks after portal vein ligation, the changes of systemic hemodynamics, GMBF, (portal vein pressure, PVP) were observed and plasma glucagon, plasma and gastric mucosal NO2-/NO3-, 6-Keto-PGF(1alpha) were measured after intravenous sandostatin, propranolol and normal saline. RESULTS: GMBF and PVP were reduced after intravenous sandostatin or propranolol. Sandostatin decreased plasma glucagon, plasma and gastric mucosal NO2-/NO3-, and had no effect on MAP and heart rate of PHG rats as compared with control group of PHG. Propranolol had not these effects, but it might decrease MAP and heart rate. There were no differences of plasma and gastric mucosal 6-Keto-PGF(1alpha) between all PHG groups. CONCLUSION: Sandostatin reduced GMBF of rats with PHG by hindering glucagon, NO, and had no effect on systemic hemodynamics. PMID- 10712779 TI - [Therapeutic efficacy of microsphere-entrapped curcuma aromatica oil infused via hepatic artery against transplanted hepatoma in rats]. AB - OBJECTIVE: To observe the therapeutic efficacy of microsphere-entrapped Curcuma aromatica oil (MS-CAO) infused via hepatic artery against transplanted hepatoma in rats. METHODS: One hundred duplicated rats bearing transplanted hepatoma were randomly divided into control group, CAO group, blank microsphere (B-MS) group, high-dose and low-dose MS-CAO groups. Each group included twenty rats. Normal saline (0.2-0.3 ml), CAO (10mg/kg), B-MS (10mg/kg) and MS-CAO (10mg/kg, 5mg/kg) were infused from gastroduodenal artery into hepatic artery, in five groups of rats respectively. The survival periods and the tumor growth rates and necrosis grades were compared in rats among five groups. RESULTS: Compared with the control group of rats, the tumor growth rates were significantly inhibited (1.23% +/- 0.66%, 4.86% +/- 1.47% vs 22.44% +/- 17.81%, F=10.21, P<0.01), the tumor necrosis grades were more extensive (Hc=23.63, P<0.01) and the survival periods were also prolonged (25.50 d +/- 3.89 d, 22.70 d +/- 3.92 d vs 11.70 d +/- 1.89 d, F=36.53, P<0.01) in either high-dose or low-dose MS-CAO group. The therapeutic effects of MS-CAO were superior to either CAO or B-MS. CONCLUSION: The therapeutic efficacy of MS-CAO infused via hepatic artery against hepatoma was much better than that of CAO or B-MS in the rats bearing transplanted hepatoma. PMID- 10712780 TI - [Antitumor effect of arsenic trioxide on mice experimental liver cancer]. AB - OBJECTIVE: To investigate the antitumor activity of arsenic trioxide on mice experimental liver cancer. METHODS: Mice bearing HepA solid and ascitic liver tumor were used in vivo experiments. RESULTS: The tumor-bearing mice were treated with arsenic trioxide 2.0mg/kg/d and 3.5mg/kg/d intravenously for 7 days. After the administration of arsenic trioxide, the growth of solid tumor were obviously inhibited, the inhibitory rate were 31.63% and 42.13% respectively. Under the inspection of transmission electron microscope, some cells of the solid tumor showed the typically morphological characteristics of apoptosis. The apoptotic cells were detected by in situ TdT-mediated dUTP nick end labeling (TUNEL). The immunohistochemical staining showed that the number of bcl-2 protein positive cells decreased, but the number of bax protein positive cells increased. It also showed that arsenic trioxide could significantly prolong the mean survival time in ascitic-tumor-bearing mice, the prolonging rate of life span was 59.29% and 76.69% respectively. The sub-G1 peaks were observed by flow cytometry on the ascitic specimen and the apoptotic rate were 25.98% and 53.17% respectively. CONCLUSION: arsenic trioxide has obvious antitumor activity on HepA liver tumor bearing mice. The mechanism of arsenic trioxide may mainly be inducing liver cancer cells to undergo apoptosis, which may be related to downregulate the expression of bcl-2 genes and upregulate the expression of bax genes. PMID- 10712781 TI - [Experimental study on value of carcino-embryonic gama-glutamyl transferase isoenzymes for monitoring carcinogenesis of hepatocytes]. AB - OBJECTIVE: To explore the value of gama-glutamyl transferase (GGT) abnormal expression in monitoring carcinogenesis of hepatocytes. METHODS: The kinetic alterations of rat (SD) liver pathology, enzymatic histochemistry, total RNA levels, different molecular form GGT and its isoenzymes were investigated on experimental hepatomas inducing with 0.05% 2-fluoenylacetamide (2-FAA). RESULTS: The GGTs of livers were overexpressed during canceration, liver GGT specific activities (U/g) including soluble and membrane-combined GGT activities of each group in experiment rats were significantly higher (q=4.90, P<0.01) than those of control group. The total RNA concentration was highly positively correlated with liver GGT specific activities or serum GGT activities (r=0.90, r=0.79, P<0.01). The mixed isoenzyme patterns of control rats' and embryo rats' GGT were found in sera and livers of experimental rats, the carcino-embryonic GGT bands can be detected at early stages of rat hepatocyte canceration. CONCLUSIONS: The present data suggest that GGT is a sensitive enzyme for carcinogenesis of hepatocytes, the analysis of carcino-embryonic GGT is a useful marker for early diagnosis of rat hepatoma. PMID- 10712782 TI - [Transfection and expression of HCV-NS(5)B gene in Huh-7 cells]. AB - OBJECTIVE: Hepatitis C virus (HCV) is the major cause of blood-borne non-A, non-B hepatitis. An RNA-dependent RNA polymerase is encoded by HCV non-structural protein 5B (NS(5)B) gene. A full-length HCV RNA has been transfected into human hepatoma cells, and HCV NS(5)B has been expressed in E.Coli purified. But human hepatocyte line with high expression of NS(5)B has not been established yet. METHODS: The NS(5)B gene has been transfected into Huh-7 cells by Lipofectamine. The results of transfection were confirmed by PCR and Southern blot analysis, and the level of the NS(5)B protein in Huh-7 cells was detected by Western blot analysis. RESULTS: There were the NS(5)B sequence and the expression of HCV-RNA polymerase in Huh-7 cells transfected NS(5)B plasmids. CONCLUSIONS: We have established a HCV RNA polymerase expression system in Huh-7 cells that can be used to study the mechanism of HCV replication and to test gene therapy of hepatitis C. PMID- 10712783 TI - [Study on biliary motility in cirrhotic patients with portal hypertension]. AB - OBJECTIVE: To study biliary motility in cirrhotic patients with portal hypertension and analyze the relationship between these biliary motility and the degree of liver function. METHODS: Biliary motility function was detected in 40 patients with portal hypertension and 30 normal controls by using quantitative hepatobiliary scintigraphy. RESULTS: The rate of dysfunction of gallbladder in cirrhotic patients was 52.5%, and it was significantly higher than that in controls (chi(2)=11.92, P<0.01). Compared with the normal controls, the bile's transit in biliary duct of cirrhotic patients was slower (t=11.09, P<0.01), the time of gallbladder visualization became longer (t=5.65, P<0.01), and the gallbladder emptying time was markedly prolonged (t=2.10, P<0.05). A correlation was also seen between these biliary motility and the degree of liver function. CONCLUSION: Cirrhotic patients with portal hypertension often have abnormal motility of gallbladder and biliary duct. PMID- 10712784 TI - [Expression of FADD in primary hepatocellular carcinoma and its relationship with hepatic apoptosis]. AB - OBJECTIVE: To investigate the features of Fas-associated death domain (FADD) expression in HCC and to determine its relationship with hepatic apoptosis. METHODS: Immunohistochemistry method was used to detect FADD expression in HCC. In situ end-labeling (ISEL) was employed to determine apoptotic cells. RESULTS: Ten cases (25. 6%) of HCC were detected to express FADD protein. The positive rate in HCC was lower than that in non-cancerous adjacent liver tissues which showed as high as 62.5% cases harbored FADD protein (P<0.05). Six out of 18 (33.3%) cases of mid to well differentiated HCC had detectable FADD protein. It is a little higher, but with no statistic significance, than that in poorly differentiated HCC which showed only 19% cases were positive for FADD. In those of grade I/II, 8 cases (38.1%) were FADD positive, while only 11.1% (2/18) cases of grade III/IV had FADD protein (P<0.05). No relationship was found between FADD expression and other clinical features, such as sex, age, tumor size or metastasis. ISEL positive cells could be seen in all cases of HCC. The hepatic apoptosis was associated with FADD expression because more apoptotic cells were detected in those cases which showed moderate to strong FADD positive, as compared with negative or weak FADD positive cases (P<0.05). While no relationship was found between FADD expression and hepatic apoptosis in non cancerous adjacent liver tissues. CONCLUSION: Loss of FADD expression plays an important role in HCC carcinogenesis. The apoptosis of cells is associated with FADD expression in HCC. PMID- 10712785 TI - [Study of expression of hepatitis C virus antigens and viral replication in extrahepatic tissues]. AB - OBJECTIVE: To explore the state of hepatitis C virus (HCV) infection and replication in extrahepatic tissues and its significance in hepatitis C patients. METHODS: Immunohistochemical assay and reverse transcriptase polymerase chain reaction (RT-PCR) were used respectively to detect HCV antigens (NS3, NS5 and CP10) and HCV RNA positive-and negative-strand in kidney, heart, pancreas, and intestine from 9 hepatitis C patients. RESULTS: Positive staining for HCV NS3, NS5 and CP10 in extrahepatic tissues was found in 6 of the 9 (66.7%) patients. The expression of HCV antigens was observed only in plasma of cells. The amount of positive-staining cells was less and there were a few differences between the kind, amounts and distribution of HCV antigen-positive-staining cells in different extrahepatic tissues. HCV RNA was detected in extrahepatic tissues from all the patients by RT-PCR. The detection rates of HCV RNA positive-and negative strand were 64.5% and 35.5% in 31 extrahepatic tissues specimens, respectively. The positive of HCV RNA positive-and negative-strand detection was coincident with that of HCV antigens detection in these tissues (Kappa>0.40). CONCLUSIONS: These results suggest a possible presence of HCV infection and replication in extrahepatic tissues. Low-level of HCV extrahepatic infection and replication is possibly a major cause of a low positive rate and inconsistent results of HCV detection in extrahepatic tissues. PMID- 10712786 TI - [Clinical manifestation and pathological change of autoimmune hepatitis]. AB - OBJECTIVE: To determine clinical manifestation and pathological changes in autoimmune hepatitis (AIH) by liver biopsy. METHODS: The clinical manifestation and pathological changes by liver biopsy in 14 AIH patients were investigated retrospectively. RESULTS: Clinically, AIH affects females with a females/male ratio of 13/1, the age range was 28-66 year; there was extended elevation of ALT and AST, hyperglobulinemia, hypergammaglobulinemia with predominant elevation of IgG and different kinds of antibodies. Patients usually had autoimmune diseases with pathological change of typical chronic hepatitis. The levels of globulin (33 approximately 60 g/L), gammaglobulin (23.9% approximately 60.5%), IgG (8.2 approximately 39. 0 g/L), ds-DNA (1.5 approximately 58.0 g/L) in AIH were significantly higher than those of normal (t=9.7, 9.3, 6.2, 3.5, P<0.01). CONCLUSION: AIH has severe liver impairment with many other symptoms due to multi systemic damage out of liver. AIH is a distinct autoimmune disease. PMID- 10712787 TI - [Clinical pathology of Dubin-Johnson syndrome]. AB - OBJECTIVE: To investigate the property of pigment granules in the hepatocytes in patients with Dubin-Johnson syndrome. METHOD: Light microscopy, histochemical, immnohistochemical and electron microscopy techniques were used to study the pigment glanules and the expression of S-100 protein and HMB45 in hepatocytes. RESULTS: Histological examination revealed normal lobular architecture and the abundont brown pigments which were chiefly seen in the centrilobular zone hepatocytes. The pigment granules were evidenced to have the characterization of both lipofuscin and melanin by histochemical staining and ultrastructural studies and to have the featurts of the melanin by immnohistochemical staining. CONCLUSION: The results suggest that the pigment graunles are lipfuscin-melanin complex in the hepatocytes in patients with Dubin-Johnson syndrome. PMID- 10712788 TI - [Molecular design and immunoreactivity studies of multiple antigen peptide corresponding to hypervariable region 1 sequence of hepatitis C virus]. AB - OBJECTIVE: To get more new information about the antigen epitopes of the hypervariable region 1 (HVR1) within the putative envelope glycoprotein of hepatitis C virus (HCV). METHODS: We designed and synthesized linear epitope peptides (LP) and multiple antigen peptide (MAP, symmetric 8 branches) derived from 22 amino acids (position 390-411aa) of HCV-BJ (HCV 1b genotype) HVR1. We compared the binding-reactivity of LP with MAP to native anti-HVR1 antibody in HCV-infected patients sera. In addition, the HCV genotype was tested to assess the prevalence of anti-HVR1 by cleaving PCR products with restriction enzyme. RESULTS: (1)There was a significant difference (P< 0.0001) of mean A450 between anti-MAP (mean +/- s =0.461 +/- 0.590) and anti-LP (mean +/- s =0.145 +/- 0.166), and the immunoreactivity of MAP and LP to the native antibody had a significant association (P < 0.001). (2)The rate of anti-LP and anti-MAP detection in 59 anti HCV2.0 positive sera was 29% and 58% respectively. (3)The binding of MAP to antibody protein in sera from patients infected with other genotypes besides HCV 1b genotype was observed. CONCLUSION: The binding-reactivity of MAP to native antibody was obviously higher than that of LP, and there is a cross-reaction between them. It was possible to analyse conformational antigen epitopes in the region and develop HCV molecular vaccine by synthesizing MAP corresponding to HCV HVR1 sequences. PMID- 10712789 TI - Ask the doctor. I've been taking vitamin E for years in hopes that it will prevent heart problems. Now the news is that it is of no help at all. Can you shed some light on this? PMID- 10712791 TI - Psychiatric treatments and patients in 1997. PMID- 10712790 TI - By the way, doctor... My problem is the opposite of everyone else I know - I have been losing, not gaining, weight. I am a 56-year-old man, and a year ago I weighed 166. Today, I'm down to 152. I feel pretty good, and my appetite is fine, but I am concerned. I did start to smoke again last year. PMID- 10712792 TI - New hope for early Alzheimer's disease. PMID- 10712793 TI - By the way, doctor. I am 68 years old and healthy, but over the past year, I've found myself less and less able to get enough sleep. My doctor is reluctant to prescribe sleeping pills, but I feel desperate to get my eight hours, and I'm tired most days. Can you recommend a sedative? PMID- 10712795 TI - Editor's acknowledgements PMID- 10712796 TI - Farm animal research: identifying all the customers. PMID- 10712794 TI - Anisomycin uses multiple mechanisms to stimulate mitogen-activated protein kinases and gene expression and to inhibit neuronal differentiation in PC12 phaeochromocytoma cells. AB - Treatment of PC12 cells with nerve growth factor (NGF) stimulates extracellular signal-regulated kinases (ERKs), as well as stress-activated c-Jun N-terminal kinases (JNKs) and p38 kinase, and induces neuronal differentiation. While the pivotal role of ERKs in NGF-induced morphological differentiation is well established, the contribution of JNK- and p38-pathways is less clear. The role of the JNK- and p38-pathway in PC12 cells was analysed by using anisomycin, a protein synthesis inhibitor that activates JNKs and p38. Non-toxic concentrations of anisomycin were found to stimulate these enzyme activities as well as the expression of the early response genes c-jun, c-fos and zif268, and to inhibit NGF-induced neurite formation. These effects of anisomycin appear to be mediated by the generation of reactive oxygen species (ROS), which in turn act through both TrkA/Ras-dependent and -independent signalling pathways. In addition, cross talk between the p38- and ERK-pathways appears to play a role in the action of anisomycin. PMID- 10712797 TI - Competitive exclusion--an alternative to antibiotics. PMID- 10712798 TI - Researching risk factors associated with cattle lameness. PMID- 10712799 TI - Prospects for 'competitive exclusion' treatment to control salmonellas and other foodborne pathogens in poultry. AB - In newly hatched chicks, the rapid establishment of an adult-type intestinal microflora, via the oral route, produces almost immediate resistance to colonization by any food poisoning salmonellas that gain access to the rearing environment. Exploitation of the 'competitive exclusion' (CE) effect is now an accepted part of the overall strategy by which poultry-associated salmonellas are being controlled in some countries. This review covers practical aspects of CE treatment and factors affecting efficacy in both laboratory-scale trials and field studies. It also considers possible applications in preventing colonization of poultry with Escherichia coli O157 and Campylobacter jejuni. For the latter, evidence suggests that the 'protective' organisms are different from those involved in salmonella control. PMID- 10712800 TI - Infections and intoxications associated with animal feed and forage which may present a hazard to human health. AB - Animal feed or forage may be the source of a limited number of infections for farm animals that could lead to human illness. Likely organisms include Salmonella enterica, Toxoplasma gondii, Trichinella spiralis and possibly the agent of bovine spongiform encephalopathy. The risk to human health from other infectious agents which may contaminate either feed or forage appear to be either negligible, e.g. Bacillus anthracis and Mycobacterium bovis, or non-existent, e.g. Clostridium botulinum toxin and Listeria monocytogenes. Mycotoxins present in animal feed can result in foods of animal origin also containing them. This risk is well recognized but has yet to be quantified accurately and in some instances the risk may be of theoretical rather than practical importance. Pesticides, agricultural and industrial chemicals, heavy metals and radionuclides may pollute animal feed and forages. The methods available for controlling pollution from these sources are well understood from a technical point of view although the effective implementation of controls can be difficult. PMID- 10712801 TI - Behavioural and physiological responses of sheep to 16 h transport and a novel environment post-transport. AB - The effect of a novel lairage environment on the ability of sheep to recover from 16 h of transport was investigated. Sheep were transported from grass paddocks to either novel outside paddocks or inside pens, and housed groups were transported to either familiar or novel inside pens. During transport, sheep from outside paddocks lay down less than those from inside pens. In sheep transported to inside pens, those from outside paddocks spent more time lying and spent less time eating; hay and water intakes during the first 12 h post-transport were lower than those previously kept inside. There was no obvious effect of a novel environment post-transport on blood biochemistry, suggesting that the lower post transport feed and water intakes in a novel environment did not have a significant effect on the ability of the sheep to recover from the feed and water deprivation associated with transport. PMID- 10712802 TI - A comparison of hoof lesions and behaviour in pregnant and early lactation heifers at housing. AB - Lameness, hoof lesion development and behaviour were compared for two groups of 10 heifers: one in early pregnancy (PH), the other in early lactation (LH). Both groups were housed in the summer in cubicles under identical conditions. Behavioural observations commenced immediately after housing, and then at 2, 4 and 6 weeks post-housing. Locomotion scores were assessed weekly, and feet were examined for lesions at approximately -1, 0, 1 and 2 months after housing. All four feet were photographed, lesions were scored subjectively for severity, and lesion size and position were estimated using image analysis techniques. LH already had greater total lesion scores before housing. More severe linear lesions in the LH group were associated with reduced lying, and less idling, increased standing in cubicles and more disturbed lying behaviour. PMID- 10712803 TI - Canine serum protein patterns using high-resolution electrophoresis (HRE). AB - Serum protein values were determined in 26 healthy dogs using agarose gel electrophoresis (SPE), splitting the electrophoretic separation into six regions: albumin, alpha(1), alpha(2), beta(1), beta(2)and gamma globulins. High-resolution electrophoresis (HRE) was used to separate single proteins. Serum proteins from dogs (26 healthy and 20 affected by various diseases) were then characterized by electrophoretic immunofixation (IFE) and Sudan black staining on HRE film. Haemoglobin and normal canine plasma and serum were used to identify haptoglobin and fibrinogen, respectively. In the standard pattern, determined by HRE, the following proteins were identified: albumin, alpha(1)-lipoprotein (alpha(1) region), haptoglobin and alpha(2)-macroglobulin (alpha(2)-region), beta lipoprotein and C3 (beta(1)-region), transferrin and IgM (beta(2)-region), IgG (mostly in gamma -region and partly in beta(2)-region). The HRE pattern shown by healthy dogs could be compared with those of dogs affected by various diseases to obtain clinical information. PMID- 10712804 TI - Relative accuracy of the identification of ovarian structures in the cow by ultrasonography and palpation per rectum. AB - Manual palpation or ultrasonographic examination of the cow's genital tract are currently used by veterinarians involved in reproductive management, but knowledge of the potential and the limitations of both methods is important to obtain an optimal accuracy in the diagnosis of physiological and pathological ovarian structures. This review presents the main features of manual and ultrasonographic characteristics of follicles, corpora lutea (with or without a cavity), follicular and lutenized cysts and the reliability of the two methods is compared. Manual diagnosis of follicles <10 mm is rather inaccurate, but ultrasound offers the possibility to diagnose follicles <5 mm and to measure their inner diameter. The predictive values of the presence or absence of a corpus luteum as determined by palpation are similar (78 vs. 75%). Manual or ultrasonographic diagnosis of the growing or regressing corpus luteum is rather difficult. The positive predictive value of a mature corpus luteum diagnosed by ultrasonography is lower (87%) than the negative predictive value (92%). Compared to manual palpation, ultrasonography permits a better estimation of the number and to determine more precisely the size of the mature corpus luteum. The positive values for follicular cysts diagnosed by palpation or by ultrasonography are 66 and 74% respectively, and for luteal cysts, the values are 66 and 85%, respectively.Manual palpation or ultrasonography are useful tools to diagnose ovarian structures in the cow. The accuracy of such methods can be enhanced by securing information on the reproductive history of the animal, palpation of uterine horns, vaginal examination or progesterone determinations. PMID- 10712805 TI - Infectious agents associated with rabbit pneumonia: isolation of amyxomatous myxoma virus strains. AB - Sixty-six rabbits, with no history of vaccination against myxomatosis and which had died of pulmonary lesions, were submitted for virological and serological tests for Myxoma virus (MV) infection and for bacteriological examinations. At post mortem, the diagnoses based on observed lesions were as follows: acute haemorrhagic pneumonia (38%); acute suppurative bronchopneumonia (35%); and fibrinohaemorrhagic bronchopneumonia with fibrinous pleuritis (27%). MV was isolated from 10% of the rabbits, mainly from those with acute haemorrhagic pneumonia. Serological evidence of MV infection was demonstrated in 44% of rabbits. Pathogenic bacteria species isolated from lungs were Pasteurella (spp. and multocida), Escherichia coli, Bordetella bronchiseptica and Pseudomonas aeruginosa, respectively, from 41, 11, 7 and 6% of samples. No relationship could be established between the presence of specific antibodies to MV and the observed pulmonary lesions or the results of the bacteriological examinations. A significant trend was established between the severity of the lesions and the results of the bacteriological examinations. PMID- 10712806 TI - Changes in interstitial cells during development of buffalo testis. AB - Interstitial cells were identified and counted in the testis of Murrah buffalo calves and bulls at the age of 1, 3, 6, 9, 12, 15, 18, 21, 24, 30, 36, 42, 48 and 72 months and older. Six types of cells were identified in the testicular interstitium of 1-month-old calves. These were mesenchymal cells, fetal type Leydig cells, fibroblasts, myoid cells, pericytes and endothelial cells. Adult Leydig cells were visible in 3-month-old calves, but mesenchymal cells were not seen from 18 months onwards. The percentage of mesenchymal cells reached a maximum in 1 month, fetal type Leydig cells in 3 months, adult Leydig cells in 72 months and beyond, fibroblasts in 36 months, myoid cells in 18 months, pericytes in 21 months and endothelial cells after 15 months. Changing percentages of various interstitial cells revealed that myoid cells may have differentiated into fibroblasts and mesenchymal cells, which then differentiated into adult Leydig cells. PMID- 10712807 TI - Influence of vitamin D and retinoids on the induction of functional differentiation in vitro of canine osteosarcoma clonal cells. AB - The efficacy of 22-oxacalcitriol (OCT), calcitriol, cholecalciferol, all-trans retinoic acid (ATRA) and 9-cis retinoic acid (9-cis RA) to differentiate in vitro four clonal cells of the canine osteosarcoma cell line POS into cells having properties of a functionally mature osteoblast bone cell were investigated. The induction of intracellular alkaline phosphatase (ALP) activity, osteocalcin (GLA OC) and type I collagen (PIP) production after 72 h treatment were used as markers of differentiation. At a concentration of 10(-8)M, OCT and calcitriol significantly induced all markers, and ATRA only the ALP of osteoblast, chondroblast and undifferentiated clonal cells. At the same concentration, 9-cis RA and cholecalciferol induced ALP of chondroblast and osteoblast cells, respectively; ATRA, 9-cis RA and cholecalciferol induced PIP of chondroblast and undifferentiated cells. None of the drugs significantly differentiated fibroblast cells. The ability of these agents to differentiate osteosarcoma cells into cells that exhibit properties of functionally mature osteoblastic bone cells may promote normal osteogenesis and reverse the loss of control of their differentiation. PMID- 10712808 TI - Efficacy evaluations of the use of oral tilmicosin in pneumonic calves. AB - The therapeutic effect of oral tilmicosin was compared with enrofloxacin, and the efficacy of three doses of the drug was examined in two fully randomized blinded field trials. Pneumonic milk-fed calves between 3 days and 2.5 months of age were allocated into two groups in trial 1 (50 animals) and into three groups in trial 2 (69 calves). In the first trial, the animals were treated with 25 mg/kg/day tilmicosin or 2.5 mg/kg/day enrofloxacin in milk for 5 days. In the second trial, the calves received either 25 mg/kg/day tilmicosin for 5 days or 3 days, or else 12.5 mg/kg tilmicosin for 5 days. All calves were clinically examined for 10 days. In the first trial, oral tilmicosin at a dose of 25 mg/kg/day for 5 days proved to be effective for the treatment of endemic pasteurellosis of milk-fed calves. The efficacy was the same as that of enrofloxacin. All three doses in the second trial were effective and were statistically equivalent to the original dose tested. PMID- 10712809 TI - Neospora caninum and bovine virus diarrhoea virus infections in Swedish dairy cows in relation to abortion. AB - The protozoan parasite Neospora caninum and bovine virus diarrhoea virus (BVDV) are recognized as important causes of bovine abortion and congenital disease worldwide. In this study, serological investigations were performed to estimate the prevalence of N. caninum infection in Swedish dairy cattle, to assess to what extent it may affect abortion rates, and to determine possible effects of coinfection with BVDV. The overall N. caninum seroprevalence in Swedish dairy cows was estimated at 2% (16/780), and the BVDV seroprevalence was 32% (249/780). Among aborting cows from herds with abortion problems, 7% (26/378) had antibodies to N. caninum and 42% (153/378) to BVDV. Seventeen of the N. caninum positive animals also had antibodies to BVDV. There was a statistically significant (P = 0.013) association between presence of antibodies to N. caninum and BVDV. In a case-control study comprising sera from cows in herds without recognized abortion problems, 6% (5/89) and 1% (1/89) of sera from aborting and non-aborting cows, respectively, had antibodies to N. caninum. Two of the N. caninum seropositive aborting cows also had antibodies to BVDV. These results confirm that N. caninum infection is associated with bovine abortion in Sweden and also indicate that there might be concurrent effects of N. caninum and BVDV. It is concluded that Swedish dairy cows have a low prevalence of N. caninum infection which is favourable in relation to possible future control programmes. PMID- 10712810 TI - Nuclear vacuolation as a cause of sterility in an angus bull. PMID- 10712811 TI - Contact lens wear alters the production of certain inflammatory mediators in tears. AB - Contact lens wear has been associated with an increased risk of corneal infection and/or inflammation. We studied the hypothesis that contact lens wear alters the number of polymorphonuclear leukocytes (PMNs) on the cornea during sleep and the levels of inflammatory mediators in the tear film. Three groups of subjects were analysed: a non-contact lens wearing group (NCLW), non-adapted (neophyte) contact lens wearers (NACLW) who wore lenses during sleep for the first time in this study and adapted contact lens wearers (ACLW) who normally wore lenses on a daily wear schedule. Ocular PMNs were collected by a non-contact irrigation technique and their numbers counted after staining. Tears were collected from each group and analysed using ELISAs for the presence of the PMN chemoattractants IL-8 and LTB(4)and the cytokines IL-1beta, IL-6 and GM-CSF. Corneal irrigation data demonstrated significantly higher numbers of PMNs from NACLW (P<0.05) compared to the other groups. ACLW showed significantly fewer PMNs (P =0.03) compared to NCLW group. The NACLW group had significantly lower concentrations (P<0. 05) of IL-8, LTB(4)and IL-6 in their tears after 8 hr of sleep compared to the other groups. The ACLW group had significantly (P<0. 05) higher levels of IL-8 at most time points compared to the other two groups. The levels of the chemoattractants IL-8 and LTB(4)in tears were inversely related to the numbers of PMNs from the corneal surface and the chemotaxis of PMNs in vitro. During one night sleep in contact lenses the numbers of PMNs and the concentration of certain inflammatory mediators are significantly altered compared to no lens wear. However, this alteration changes from NACLW to ACLW. This may have effects on the ability of the eye to defend itself during contact lens wear. PMID- 10712812 TI - Epidermal growth factor stimulates phospholipase cgamma1 in cultured rabbit corneal epithelial cells. AB - Phospholipase Cgamma1 (PLCgamma1) catalyses hydrolysis of phosphatidylinositol 4,5-bisphosphate to generate diacylglycerol and inositol 1,4,5-trisphosphate (IP(3)), two second messengers which play important roles in cell proliferation and differentiation. The purpose of the current study was to identify PLCgamma1 in corneal epithelial cells and investigate whether epidermal growth factor (EGF) stimulates the activity of this enzyme. Addition of EGF to [(3)H]myo-inositol labeled, cultured corneal epithelial cells stimulated production of IP(3), indicating activation of PLC. Western immunoblot analysis and an in vitro assay of PLC activity revealed that EGF activates gamma1 isoform of PLC, which is localized predominantly in the cytosolic fraction of the epithelial cells. EGF receptors were detected in the epithelial cells by EGF receptor antibody. Addition of EGF to the cells caused tyrosine phosphorylation of the receptors, translocation of PLCgamma1 from cytosol to plasma membrane, and phosphorylation of the enzyme at tyrosine residues. Addition of tyrphostin A-25, an inhibitor of receptor tyrosine kinase, attenuated the tyrosine phosphorylation of PLCgamma1 as well as its enzyme activity. These findings suggest that EGF stimulates PLCgamma1 in rabbit corneal epithelial cells, and that this effect is probably mediated by tyrosine phosphorylation of the enzyme. PMID- 10712813 TI - Rabbit pigmented ciliary epithelium produces interleukin-6 in response to inflammatory cytokines. AB - Interleukin-6 is a multifunctional cytokine that is found in high concentrations in intraocular fluids during the uveitic response. Although monocytic cells are a major source of interleukin-6, resident intraocular cells may also contribute to its accumulation in intraocular fluids during uveitis. The purpose of this study was to determine whether interleukin-6 is produced by pigmented ciliary epithelial cells and whether agents known to stimulate interleukin-6 production, such as interleukin-1beta, tumor necrosis factor-alpha, bacterial endotoxin, and stimulators of the adenylyl cyclase/adenosine 3',5'-cyclic monophosphate system, increase interleukin-6 production by these cells. Primary and first-passage cultures of nontransformed rabbit pigmented ciliary epithelial cells were incubated with the test agents for varying periods of time in serum-free medium and interleukin-6 levels in the cell-conditioned medium were measured by bioassay.Little, if any interleukin-6 was released from pigmented ciliary epithelial cells incubated for up to 18 hr in serum-free medium. Interleukin 1betastimulated interleukin-6 release in a time- and concentration-dependent manner. Tumor necrosis factor-alpha, although ineffective alone, increased interleukin-1beta-induced interleukin-6 release in a concentration-dependent manner when co-incubated with interleukin-1betafor 18 hr. However, tumor necrosis factor-alphadid not enhance interleukin-1beta-induced interleukin-6 release if co incubated with interleukin-1betafor a shorter time (6 hr). A 6 hr exposure to bacterial endotoxin did not stimulate interleukin-6 release from pigmented ciliary epithelial cells. Co-incubation of pigmented ciliary epithelial cells with interleukin-1betaand agents that stimulate the adenyl cyclase/adenosine 3',5'-cyclic monophosphate system through cell surface G-protein transduced receptors, i.e. isoproterenol, vasoactive intestinal peptide or prostaglandin E(2), significantly enhanced the ability of interleukin-1betato stimulate interleukin-6 release. However, neither the adenyl cyclase activator, forskolin or the adenosine 3', 5'-cyclic monophosphate-mimetic, dibutyryl 3',5'-cyclic monophosphate enhanced interleukin-1beta-induced release of interleukin-6. These results indicate that the pigmented ciliary epithelium is one potential source of interleukin-6 and may contribute to the elevation in intraocular fluid interleukin-6 levels observed during various intraocular inflammatory episodes. Although agents that activate the adenyl cyclase/adenosine 3', 5'-cyclic monophosphate system through cell surface G-protein transduced receptors increased interleukin-1beta-induced release of interleukin-6, the ineffectiveness of forskolin and dibutryl 3', 5'-cyclic monophosphate suggest that simply increasing intracellular 3',5'-cyclic monophosphate is not sufficient to augment interleukin-1beta-induced release of interleukin-6. The significance of interleukin-6 in the intraocular inflammatory response is discussed in terms of its proposed role in an endogenous antiinflammatory system acting through induction of interleukin-1 receptor antagonist, soluble tumor necrosis factor receptor, acute-phase proteins and corticosteroids. PMID- 10712814 TI - Acetylsalicylic acid does not reduce the intraocular pressure variation in ocular hypertension or glaucoma. AB - The purpose of this study was to measure if intraocular pressure (IOP) and IOP variations in patients with ocular hypertension and glaucoma are decreased by acetylsalicylic acid (ASA). The hypothesis to be tested was that short-term fluctuations in the IOP are caused by breaks of the inner wall of Schlemm's canal that are repaired by platelets inducing a cycle of breaks and repair. Furthermore, prostaglandins affect uveoscleral outflow and ASA inhibits prostaglandin biosynthesis and platelet aggregation. This implies that ASA may have complex effects on the IOP and its variations.In 28 patients with ocular hypertension or glaucoma the IOP was measured seven times during 2 hr on two succeeding days. Five hundred mg ASA or placebo was administrated orally in a masked fashion 15 hr prior to the second session. After wash-out, this procedure was repeated with a cross-over design. The same study outline was used in 28 glaucoma patients except for the cross-over design. There were no statistically significant differences in the mean IOP or in the IOP variations between the placebo treated and the ASA treated eyes in either group, and there were no significant differences between the day before and after treatment in any group. The results suggest that ASA does not affect IOP variations in a clinically significant way and that a single dose of ASA has no significant effect on mean IOP. PMID- 10712815 TI - Influence of muscarinic agonists and tyrosine kinase inhibitors on L-type Ca(2+)Channels in human and bovine trabecular meshwork cells. AB - Trabecularmeshwork (TM), a smooth muscle-like tissue with contractile properties, is involved in the regulation of aqueous humor outflow. However, little is known about the regulation of Ca(2+)influx in trabecular meshwork cells. We investigated the influence of acetylcholine and tyrosine kinases on Ca(2+)conductances of bovine TM (BTM) and human TM (HTM) cells using the perforated-patch configuration of the patch-clamp technique and measurements of intracellular free Ca(2+)([Ca(2+)](i)). Depolarization of the cells in the presence of 10 m m Ba(2+)or Ca(2+)led to an activation of inward currents at potentials positive to -30 mV with characteristics typical of L-type Ca(2+)currents: when using 10 m m Ba(2+), maximal inward current and inactivation time constant (tau) increased; the L-type Ca(2+)channel blocker nifedipine (1 microm) reduced and the L-type Ca(2+)channel agonist BayK8644 (5 microm) enhanced maximal inward current. Acetylcholine (100 microm) and carbachol (1 microm) led to an increase in inward Ba(2+)current whereas application of the tyrosine kinase inhibitors genistein (50 microm) and lavendustin A (20 microm) resulted in a decrease in inward current. The application of daidzein (10 microm), an inactive analog of genistein had no effect. Depolarization of the cells with 135 m m K(+)or direct stimulation of L-type channels by application of BayK 8644 led to an increase in [Ca(2+)](i). Carbachol (1 microm) induced an increase in [Ca(2+)](i)which was decreased by application of the tyrosine kinase inhibitor genistein (50 microm). We conclude that HTM and BTM cells express voltage dependent L-type Ca(2+)channels that influence intracellular Ca(2+)concentration and thus may modulate TM contractility. The activity of L-type Ca(2+)currents is influenced by muscarinic agonists and tyrosine kinases. PMID- 10712816 TI - Cyclic AMP-dependent stimulation of basolateral K(+)conductance in the rabbit conjunctival epithelium. AB - The regulation of Na(+)transport by cAMP in freshly isolated rabbit conjunctival epithelium, a tissue exhibiting both Cl(-)secretion and Na(+)absorption, was examined. Bulbar-palpebral segments of conjunctiva were mounted between Ussing type hemichambers under short-circuit conditions in Cl(-)free media. In this situation, the short-circuit current (I(sc)) measures an amiloride-resistant Na(+)absorptive process in the apical-to-basolateral direction. Apical additions (each at 10 microm) of cAMP-elevating compounds, forskolin, rolipram, IBMX and epinephrine all stimulated the Na(+)-dependent I(sc)by approximately 3.5-4.5 microA cm(-2)(minimal 40% increase) and reduced transepithelial resistance (R(t)) by at least 7% (P<0.05). Pre-exposure (1 hr) to the protein kinase A (PKA) inhibitor H-89 (10 microm), which in itself inhibited the I(sc)by 0. 5 microA cm( 2), attenuated the I(sc)responses of the cAMP-elevating agents (P< 0.05, unpaired data). In reverse, H-89 promptly decreased the I(sc)by 1.5-2.5 microA cm(-2)and increased R(t)by 5% (P<0.05) in tissues pre-stimulated with either forskolin or an epinephrine plus IBMX combination. Additions of epinephrine or rolipram to apically permeabilized preparations using amphotericin B, increased the I(sc)by 12 and 22% respectively over baseline and reduced R(t)by 6% (P<0.05). Similarly, in the presence of a transepithelial K(+)gradient (apical to basolateral) and amphotericin B, cAMP elevation stimulated K diffusion across the preparation by at least 1.8 microA cm(-2)and decreased the R(t)by 4% (P<0.05), changes that were reversed by subsequent H-89 addition. Under Cl(-)rich conditions, pretreatment with 5 m m Ba(2+)reduced the basal I(sc)by 59% and blocked the cAMP-induced I(sc)stimulations typically seen in the presence of the anion. The results provide evidence for a PKA-regulated, Ba(2+)-inhibitable (voltage insensitive) basolateral K(+)conductance in rabbit conjunctival epithelial cells. The action of Cl(-)secretogogues acting via cAMP on basolateral K(+)channel activity indicates that endogenous levels of cAMP may play a role in the regulation of Cl( )secretion and Na(+)absorption in the conjunctiva. PMID- 10712817 TI - Effect of latrunculin-B on outflow facility in monkeys. AB - Latrunculin-B (LAT-B), a macrolide derived from the marine sponge Latrunculia magnifica, sequesters monomeric G-actin, leading to the disassembly of actin filaments in cultured cells. In this study, we determined the effect of LAT-B on outflow facility in living monkeys. Total outflow facility was measured by 2 level constant pressure perfusion of the anterior chamber (AC) before and immediately after AC exchange infusion or 2 hr after topical application of LAT-B or vehicle. Both AC exchange infusion and topical application of LAT-B dose- and time-dependently increased outflow facility by two- to four-fold. Those findings suggest that pharmacological disorganization of the actin cytoskeleton in the trabecular meshwork by specific actin inhibitors such as LAT-B may be a useful anti-glaucoma strategy. PMID- 10712818 TI - Role of muscarinic cholinergic transmission in Edinger-Westphal nucleus-induced choroidal vasodilation in pigeon. AB - Activation of the parasympathetic ciliary ganglion input to the choroid causes increases in choroidal blood flow. We examined the role and the type of muscarinic receptors within the choroid that are involved in these increases in choroidal blood flow, using electrical stimulation of the nucleus of Edinger Westphal (EW) to activate the ciliary ganglion input to choroid in ketamine anesthetized pigeons. Baseline choroidal blood flow and its EW-evoked increases measured as peak and total (area under the curve) responses were determined using laser Doppler flowmetry. The EW-evoked responses were reduced dose-dependently after administration of 4-diphenyl-acetoxy-N-methylpiperedine (4-DAMP), a relatively selective antagonist of M3 type muscarinic receptors, with a maximal mean decrease of 86% (peak response) and 93% (total response) at a dose of 10 microg kg(-1)i.v. without a significant effect on baseline choroidal blood flow, heart rate or systemic arterial blood pressure. Atropine, a non-selective antagonist of muscarinic receptors, decreased the EW-evoked responses to a lesser extent than 4-DAMP after intravenous administration of 1 mg kg(-1)(by 67% for peak response and by 53% for total response) or topical administration of a 5% solution (by 41% for peak response and by 62% for total response), both of which increased heart rate and systemic arterial blood pressure without a consistent effect on baseline choroidal blood flow. In contrast, himbacine (i.p. 10 microg kg(-1)), a relatively selective antagonist of M2 type muscarinic receptors, increased the EW-evoked parasympathetic cholinergic vasodilation (by 93% for the peak response and by 142% for the total response) without a significant effect on heart rate, systemic arterial blood pressure or baseline choroidal blood flow. The results of our study suggest a major role of M3 type muscarinic receptors in the EW-evoked increases in choroidal blood flow. Based on findings that the ciliary ganglion input to choroid does not synthesize nitric oxide but inhibitors of NO production do block EW-evoked choroidal vasodilation, it seems likely that the M3 receptors acted on by 4-DAMP are present on choroidal endothelial cells and mediate choroidal vasodilation via stimulation of endothelial release of nitric oxide. In contrast, M2 muscarinic receptors may play a presynaptic role in downregulating EW-evoked parasympathetic cholinergic vasodilation in avian choroid. PMID- 10712819 TI - Amniotic membrane patching promotes healing and inhibits proteinase activity on wound healing following acute corneal alkali burn. AB - Amniotic membrane (AM) contains basement membrane components and various proteinase inhibitors. Furthermore, when used as a graft, the basement membrane of AM could block inflammatory insults to a damaged corneal surface. Thus, we evaluated whether amniotic membrane patching could promote the healing process by inhibiting proteolytic damage. Alkali wounds were inflicted on the central corneas of rabbits by applying a round filter paper, 6.0 mm in diameter, soaked in 1 N NaOH for 30 sec. Amniotic membrane patching was performed over the perilimbal sclera immediately after wounding. A total of 115 rabbits were divided into four groups: (1) immediately covered by AM with the amnion cell side down up to the perilimbal sclera (n =26); (2) covered by AM with the stromal side down up to the perilimbal sclera (n =19); (3) anchored to the fornix (n =29); and (4) uncovered as a control (n =41). AM was removed 3 days postoperatively. During follow-ups, epithelial defects, corneal thickness and its opacity of each eye were measured. Some corneas were removed for histopathologic studies and for proteinase activity assay and zymography. The epithelial healing was faster and the corneal thickness was thicker in all three AM-covered groups than in the control (P<0.05). No significant difference was found between covered and anchored groups (P>0.05). Corneal opacity was least in the amnion cell side down group. Infiltration of polymorphonuclear leukocytes (PMNs) was much less in AM covered groups than in the control. Pathological results were associated with zymographic findings, which revealed much higher proteinase activity in uncovered group than AM-covered groups. Immediate intervention for acute alkali burns with AM as a temporary patch promotes wound healing by inhibiting proteinase activity and PMNs infiltration. PMID- 10712820 TI - Kv3.3 potassium channels in lens epithelium and corneal endothelium. AB - Human Kv3.3/KCNC3 is a Shaw-type potassium channel that has been mapped to chromosome 19q13.3-13.4. Complete mouse and rat Kv3.3 cDNA coding sequences have been published, yet the human Kv3.3 cDNA has remained incomplete for years. We report here for the first time the amino acid sequence for hKv3.3 and the electrophysiological behavior of the encoded channel in transiently transfected mammalian cells. In addition, we report the occurrence of Kv3.3 message in rabbit corneal endothelial cells and the properties of the currents when the corneal channel is expressed. The hKv3.3 gene is highly GC-rich (69%) and contains numerous GC runs which made DNA sequencing and PCR amplification especially problematic. The full-length sequence contains two possible start codons. The encoded 757 amino acid hKv3. 3 protein is about 93% identical to mouse and rat Kv3.3 in the first 659 amino acids before the C-terminal domains diverge greatly as a result of alternative splicing. The rabbit cornea Kv3.3 is a close sequence match for hKv3.3 even in the C-terminal domain. However, we have not yet found a cornea sequence which contains the first potential start codon from hKv3.3. Electrophysiologically, the hKv3. 3 channel produces an A-current although expression of constructs which lack the 5' region of the first start codon inactivate much more slowly than full-length constructs. This short hKv3.3 construct also shows changes in activation. PMID- 10712821 TI - Expression of the keratan sulfate proteoglycans lumican, keratocan and osteoglycin/mimecan during chick corneal development. AB - The corneal proteoglycans belong to the Leu-rich proteoglycan (LRP) gene family and contain chondroitin/dermatan (CS/DS) or keratan sulfate (KS) chains. These proteoglycans play a critical role in generating and maintaining a transparent matrix within the corneal stroma. Decorin which has CS/DS chains and lumican which has KS chains, were first to be identified in the cornea. Two other corneal KS proteoglycans (KSPGs), keratocan and osteoglycin/mimecan were recently identified in bovine corneas. We cloned and sequenced chick osteoglycin/mimecan and found it to contain a stretch of 60 amino acids that showed no identity to the presumed mammalian homolog. The 177 base pair DNA coding for this unique sequence shows 47% identity to an 189 base pair sequence between exons 4 and 5 of the bovine osteoglycin/mimecan gene. This indicates that this cDNA represents an alternatively spliced form of osteoglycin/mimecan containing a unique N-terminal sequence. The expression of each of the three corneal KSPGs in the developing and mature chick cornea was investigated by competitive PCR and immuno-biochemical analysis of corneal extracts. Competitive PCR was used to determine the message levels for chick lumican, keratocan and osteoglycin in embryonic day 9, 12, 15, 18 and adult corneas. Results showed that lumican mRNA fluctuated during development but remained at a relatively high level while keratocan and osteoglycin message levels declined steadily from day 9 to adult. Additionally, lumican mRNA was present at higher levels, during all stages of corneal development, than keratocan and at much higher levels than osteoglycin. Antibodies shown to be specific for each KSPG were used to characterize proteoglycans isolated from embryonic and adult chick corneas. KSPGs from embryonic corneas eluted 1-2 fractions earlier on Q-Sepharose than KSPG from adult corneas. Additionally, Western blot analysis showed that embryonic KSPGs were more keratanase-resistant, endo-beta-galactosidase sensitive than adult KSPGs. The results of this study indicate an alteration in sulfation or the fine structure of the glycosaminoglycan chains occurs during corneal maturation for the 3 KSPGs. PMID- 10712822 TI - Atomic force microscopic study of the human cornea following excimer laser keratectomy. AB - The aim of this study was to examine the corneal surface structures with a new investigative method, the atomic force microscope following 193 nm excimer laser photoablation. Fresh human corneas were irradiated in vitro with an increasing number of impulses emitted by a 193 nm ArF laboratory excimer laser in order to produce either smooth flat surfaces or stair-like formations within the cornea. The corneas were investigated in a Topometrix(R) atomic force microscope in their native state. For comparison, three corneas were fixed with glutaraldehyde and processed for scanning electron microscopy. Atomic force microscopy and scanning electron microscopy revealed the same surface characteristics of photoablated corneas, though the preparation for scanning electron microscopy induced considerable shrinkage of the tissues. The layers of the cornea could be distinguished from each other and deeper ablations of the stroma produced a rougher surface. On the lateral walls of ablated stairs small droplets of ejected material could be seen with scanning electron microscope. Atomic force microscope produces three-dimensional images of the scanned native corneal surfaces and it could be a valuable tool to investigate the corneal smoothness. Our investigations have provided similar results as those obtained with scanning electron microscopy showing that the laser-ablated corneal surface remains relatively smooth. We suggest that the formation of condense droplets of ejected materials is based on hydrodynamic motions induced by boiling water solutions. PMID- 10712825 TI - Volume contents and index. PMID- 10712824 TI - Survival of the retinal pigment epithelium in vitro: comparison of freshly isolated and subcultured cells. AB - Cells of the retinal pigment epithelium (RPE) are generated prenatally and generally survive the lifetime of the individual without undergoing proliferation or replacement. Therefore, the mechanisms promoting individual RPE cell survival and longevity in vivo may be distinct from, or a limited subset of, the mechanisms known to promote survival in proliferative cells in culture. To identify specific factors that sustain cell viability independent of effects on cell division, we studied RPE cells in low-density suspension culture, in which cell proliferation is inhibited. Single cells from Xenopus laevis eyes were plated onto a non-adhesive surface in protein-free medium, then assayed for survival using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Cell viability in these cultures was essentially undiminished over the initial 2 days. However, by approximately 1 week in culture, only an average of 53% of the cells remained alive. Plating cells on a fibronectin-coated substratum significantly enhanced survival, such that the number of cells alive at 1 week was 80-90% of the initial level. Essentially identical results were obtained with laminin- or collagen IV-coated substrata, or with insulin (5 microg ml(-1)) in the medium. The absence of cell division in these cultures was confirmed by cell counting and BrdU incorporation experiments. Interestingly, in suspension cultures derived from monolayers previously established on microporous membrane filters, cells lost viability much faster (average of 80% dead at 3 days), and showed a relatively greater response to extracellular matrix proteins (five-fold increase in cell survival at 3 days). Enhanced RPE survival in response to fibronectin required spreading of the cell on a substratum, rather than mere adherence, as there was a high correlation between the percentage of spread cells and the percentage that were MTT-positive (r=0.940). Cell spreading apparently enhanced survival by preventing the initiation of programmed cell death: unattached non-viable cells in culture exhibited morphological features expected of apoptosis, as well as positive staining by the TUNEL reaction. These studies demonstrate that, of several factors shown to maintain or increase cell number in proliferating cultures, some have their effect, at least in part, by promoting the survival of individual cells. The increased susceptibility of subcultured RPE to cell death has implications for clinical transplantation applications that may require manipulation of RPE in vitro. PMID- 10712823 TI - Maillard reactions in lens proteins: methylglyoxal-mediated modifications in the rat lens. AB - The nonenzymatic Maillard reaction is thought to contribute to aging and cataract formation in the lens. As levels of methylglyoxal (MG) and glutathione (GSH) affect the reaction, we examined the relationship of these factors and determined the effect of a glyoxalase I inhibitor on the Maillard reaction. Rat lens cultures were maintained for up to 3 days in TC-199 medium with or without 20 m m glyceraldehyde (GLD) and 250 microm S-[N-hydroxy-N-(4-chlorophenyl) carbamoyl] glutathione diethyl ester (HCCG diester). We measured GSH, MG, D-lactate, glyoxalase I activity, immunoreactive MG-derived advanced glycation endproducts (MG-AGEs) and imidazolysine in organ cultured rat lenses. In vitro experiments with isolated rat lens proteins revealed that HCCG alone inhibited glyoxalase I activity in a dose-dependent manner. In organ cultured rat lens protein, GLD increased MG levels 24-fold, and the addition of HCCG diester further increased it by about two-fold. GSH levels fell sharply in the presence of GLD and this was prevented to some extent by the presence of HCCG diester. D-lactate production in the lens was suppressed by HCCG diester treatment. Dialysed lens proteins retained glyoxalase I activity, indicating that the enzyme was unaltered during incubation. MG-AGEs and imidazolysine levels were significantly higher (P<0.05) in GLD-treated lenses, but a combination of HCCG diester and GLD lowered immunoreactive MG-AGEs and imidazolysine levels compared to GLD alone. HCCG had no significant effect on MG-AGE formation in lens proteins incubated with GLD or MG. We conclude that exogenous GLD enhances MG and MG-AGE levels in the rat lens and that this increase is accompanied by a loss in GSH. In addition, inhibition of glyoxalase I promotes MG accumulation. PMID- 10712826 TI - THE CANNABINOID AGONIST HU 210 MODIFIES RAT BEHAVIOURAL RESPONSES TO NOVELTY AND STRESS. AB - Experiments were performed on groups of rats after acute and sub-chronic treatment (once daily for 9 days) with the cannabinoid agonist HU 210 (25-100 ug kg(-1), i.p.) as well as 24 h and 7 days after the last drug injection. The animals underwent three behavioural tests in novel environments. In the observation cages (Test 1), rat locomotor activity was found to be dose dependently reduced after acute and sub-chronic treatment at all doses and virtually unchanged during abstinence; grooming was potently inhibited by acute treatment but potentiated by the sub-chronic one at doses of 50 and 100 ug kg( 1), the effect of the higher dose persisting after 24 h and 7 days abstinence. Vocalization in animals in response to a tactile stimulus was highest after HU 210 at 100 ug kg(-1)in all experimental modes except after 7 days abstinence. In the X-maze (Test 2), sub-chronic HU 210 dose- dependently enhanced rat natural aversion for open arms, and this behaviour persisted during abstinence after the highest dose. Grooming in the X-maze was completely absent in rats acutely injected with HU 210 but potentiated in those sub-chronically treated or abstinent. In the swimming test (Test 3) rats sub-chronically treated at 50 and 100 pg kg(-1)displayed relevant wall-hugging and the same occurred 24 h after last injection. On the whole, our results are indicative of an anxiogenic-like effect of sub-chronic HU 210 at high doses and reflect the persistence of enhanced emotional response to novel environments when the treatment is discontinued. 2000 Academic Press@p$hr Copyright 2000 Academic Press. PMID- 10712827 TI - DISTRIBUTION OF LIPID-SOLUBLE ANTIOXIDANTS IN LIPOPROTEINS FROM HEALTHY SUBJECTS. I. CORRELATION WITH PLASMA ANTIOXIDANT LEVELS AND COMPOSITION OF LIPOPROTEINS. AB - The concentration of five lipid-soluble antioxidants (gamma- and alpha tocopherol, lycopene, beta-carotene and ubiquinol-10) was measured in plasma and very low-density, low-density and high-density lipoproteins (VLDL, LDL and HDL) isolated from young healthy normo- cholesterolemic subjects. Alpha-tocopherol was the exclusive antioxidant whose plasma concentration significantly correlated with the absolute concentration of total cholesterol (r =0.541, P<0.001). No correlation was found between plasma concentration and lipoprotein content of alpha-tocopherol and ubiquinol-10, whereas it reached statistically significant values for gamma-tocopherol, lycopene and beta-carotene. The alpha-tocopherol content in VLDL and HDL, but not in LDL, was strictly associated with the relative abundance of cholesterol and phospholipids in the lipoprotein particles. Moreover, the difference between alpha-tocopherol concentration in VLDL and LDL appeared to be strictly related to the differences in cholesterol, phospholipids and triglycerides. The percent distribution of the total plasma pool of antioxidant in each lipoprotein class revealed that gamma- and alpha-tocopherol were roughly equally distributed in LDL and HDL. On the other hand, lycopene, beta-carotene and ubiquinol-10 were preferentially sequestered in LDL. Finally, the absolute and relative concentration of alpha-tocopherol, but not that of other antioxidants, in HDL exhibited a statistically significant correlation with plasma HDL/LDL cholesterol ratio. These findings indicate that: (i) plasma concentrations of major lipid-soluble antioxidants are not always predictive of their levels in lipoproteins and that, within individual lipoprotein classes, (ii) the lipid composition, metabolism and relative plasma concentration may significantly affect their abundance. 2000 Academic Press@p$hr Copyright 2000 Academic Press. PMID- 10712828 TI - DISTRIBUTION OF LIPID-SOLUBLE ANTIOXIDANTS IN LIPOPROTEINS FROM HEALTHY SUBJECTS. II. EFFECTS OF IN VIVO SUPPLEMENTATION WITH alpha-TOCOPHEROL. AB - The effects of orally supplemented dl -alpha-tocopherol on the plasma concentration of lipid-soluble antioxidants and their distribution in very-low density, low-density and high-density lipoproteins (VLDL, LDL and HDL) was investigated in a cohort of control normocholesterolemic adult subjects receiving 600 mg alpha-tocopherol daily for 2 weeks. This regimen did not modify the plasma lipid profile (total, LDL and HDL cholesterol and triglycerides) and chemical composition of VLDL, LDL and HDL. Plasma concentration of alpha-tocopherol increased from 19.44+/-4.77 to 38.03+/-9.06 um and this was associated with slight decrease in the concentration of gamma-tocopherol from 1.27+/-0.97 to 0.99+/-1.17 um, without any significant changes of either lycopene and beta carotene. Qualitatively similar changes were found in VLDL, LDL and HDL but the net increase of alpha-tocopherol in plasma did not correlate with the increase in alpha-tocopherol content in any of the lipoprotein types. Following supplementation, the percentage of total plasma alpha-tocopherol pool carried by VLDL increased from 20.97+/-6.07% to 33.57+/-6.97%, whereas it decreased from 41.85+/-7.02% to 36.36+/-5.69% in the case of LDL and from 37.17+/-6.04% to 30.05+/-4.88% in the case of HDL. The absolute and relative enrichment of alpha tocopherol in either VLDL and LDL did not exhibit any statistically relevant correlation with the chemical composition of these lipoproteins in the different subjects investigated. On the other hand, the amount of alpha-tocopherol enriching the HDL particles was inversely related to the relative abundance of protein (r =0.449;P<0.05) and directly to the phospholipid/protein ratio (r =0.480, P<0.05). 2000 Academic Press@p$hr Copyright 2000 Academic Press. PMID- 10712829 TI - INFLUENCE OF BASELINE VALUES ON LIPIDS, LIPOPROTEINS AND FIBRINOLYTIC PARAMETERS DURING AMLODIPINE TREATMENT OF HYPERTENSION IN JAPANESE PATIENTS. AB - Twenty-four Japanese hypertensive patients of both sexes, grouped as having 'medium' and 'high' baseline total lipid values, had their serum lipids, lipoproteins and plasma fibrinolytic parameters, renin and noradrenaline levels determined after 3 months of amlodipine treatment. For the patients with 'medium baseline values', total plasminogen activator inhibitor-1 (PAI-1) and t-PA-PAI-1 complex levels decreased, while the changes in lipids and lipoproteins were not significant after amlodipine treatment. For the patients with 'high baseline values', the mean triglyceride and very low density lipoprotein cholesterol (VLDLC) levels were reduced while the reductions in total and free PAI-1 and the increase in tissue plasminogen (t-PA) levels were not significant after amlodipine treatment. Negative correlations were observed between t-PA and high density lipoprotein cholesterol (HDLC) and HDLC/total cholesterol (TC) ratio in the patients with 'medium baseline values' while t-PA positively correlated with HDLC/TC ratio in patients with 'high baseline values'. The mean levels of renin and noradrenaline remained unchanged before and after amlodipine treatment in the two baseline groups. These findings show that baseline lipid levels of the hypertensive patients could influence lipids and fibrinolytic parameters differently during amlodipine treatment. The baseline lipid levels also influenced the metabolic association between lipids and fibrinolytic function in hypertensive patients during amlodipine treatment. The baseline total lipid values could therefore provide explanations for the complex metabolic interaction between lipids and fibrinolytic function as well as for the antiatherogenic actions of amlodipine treatment in hypertensive patients. 2000 Academic Press@p$hr Copyright 2000 Academic Press. PMID- 10712830 TI - INFLUENCE OF BASELINE VALUES ON LIPIDS, LIPOPROTEINS AND FIBRINOLYTIC PARAMETERS DURING TREATMENT OF HYPERTENSION WITH CILNIDIPINE. AB - Sixteen adult hypertensive patients of both sexes, classified as having 'medium' (total lipid profile 240-300 mg dl(-1)), and 'high' (total lipid profile >300 mg dl(-1)) baseline values, underwent serum lipids, lipoproteins and plasma fibrinolytic parameters evaluations after 3 months of cilnidipine treatment. Patients with 'medium baseline values' did not have any change in lipids, lipoproteins and fibrinolytic parameters while patients with 'high baseline values' had beneficial lipid and lipoprotein changes [decreases in total cholesterol (TC), triglycerides (TG), very low density lipoprotein-cholesterol (VLDLC) and increases in high density lipoprotein-cholesterol (HDLC), and HDLC/TC ratio] after cilnidipine treatment. Changes in lipids were negatively associated with fibrinolysis for the patients with 'medium baseline values' and positively associated in patients with 'high baseline values' after cilnidipine treatment. Reduction in blood pressure was related to fibrinolysis and reduced risk of coronary heart disease in the patients with 'high baseline values' after cilnidipine therapy. These results show that during cilnidipine treatment, the baseline lipid profile levels of the patients may influence the lipid altering actions as well as the interaction between lipids and fibrinolysis. 2000 Academic Press@p$hr Copyright 2000 Academic Press. PMID- 10712831 TI - CYTOTOXIC ACTIVITY OF KAEMPFEROL GLYCOSIDES AGAINST HUMAN LEUKAEMIC CELL LINES IN VITRO. AB - Two kaempferol coumaroyl glycosides (i.e. platanoside and tiliroside) isolated from the methanolic extract of Platanus orientalis L. buds, were examined for their in vitro cytotoxic activity against a panel of human leukaemic cell lines. Platanoside (1) exhibited cytotoxic activity against most of the cell lines tested, while tiliroside (2) was active against two of the nine tested cell lines. Compound 1, was examined for its effect on the uptake of [(3)H]thymidine as a marker of DNA synthesis. Kaempferol was used as a control. 2000 Academic Press@p$hr Copyright 2000 Academic Press. PMID- 10712832 TI - GRANULOCYTE CHEMILUMINESCENCE ACTIVITY, ANTIBODY PRODUCTION AND PERCENTAGE OF CD4(+)AND CD8(+)LYMPHOCYTES IN PERIPHERAL BLOOD OF OFFSPRING OF SALBUTAMOL TREATED PREGNANT C3H MICE. AB - Preterm delivery is one of the most important problems in obstetric care. One of commonly used treatment of such high risk cases is salbutamol-beta(2)adrenoceptor agonist. The aim of present study was to determine if such treatment causes any changes in neonatal immune system and therefore should be considered in newborn care. The experiments were performed in 4-5- and 6-7-week-old female and male offspring of salbutamol treated C3H inbred mice. In the present study chemiluminescent activity of peripheral blood granulocytes, percentage of CD4(+)and CD8(+)lymphocytes and antibody production were evaluated. A lower number of peripheral blood granulocytes in 6-7-week-old offspring of salbutamol treated mothers was observed, while in the case of younger mice's lymphocytes count in both groups, the differences were not signtificant as compared to control group. In 4-5-week-old mice a lower percentage of CD4(+), CD3(+)and CD8(+)was evaluated, while in older offspring the percentage of CD4(+)and CD3(+)was higher in the case of the progeny of salbutamol treated mothers. As far as chemiluminescent activity was concerned no differences were found in any of experimental groups. We showed higher IgM production both in male and female offspring of the experimental group and no changes in IgG levels in mice sera. Alterations observed in progeny of salbutamol treated mice might have influence on their further immune system development and function. 2000 Academic Press@p$hr Copyright 2000 Academic Press. PMID- 10712833 TI - THYMUS AND LYMPH NODE CELL CD4(+)AND CD8(+)MARKER EXPRESSION AND THEIR ANGIOGENIC ACTIVITY OF OFFSPRING OF SALBUTAMOL-TREATED PREGNANT C3H MICE. AB - Salbutamol, beta(2)-adrenoceptor agonist is a first choice drug in preterm delivery treatment. The aim of the present study was to determine whether salbutamol can cause any alterations in neonatal immune systems and therefore should be considered in newborn care. The experiments were performed on 4-5- and 6-7-week-old female and male offspring of salbutamol-treated C3H inbred mice. Thymus and lymph node weight, cellularity and lymphatic organs, lymphocytes phenotypes and their angiogenic activity were evaluated. We observed lower thymus weight in 6-7-week-old progeny of salbutamol-treated mothers and in the same time lower thymus cell number in both age groups. Lower lymph node weight was developed in older progeny while cellularity was diminished both in 4-5- and 6-7 week-old offspring of salbutamol-treated mothers. We have not detected any changes in percentage of CD4, CD8, CD3 and CD4CD8 positive lymphocytes in progeny of salbutamol-treated mothers. As far as lymph node lymphocytes phenotype is considered we showed in both age groups lowering of CD4 and CD3 positive cells in experimental groups. In the LIA test (lymphocyte-induced angiogenesis) we showed lower lymph node cell angiogenic activity of salbutamol-treated mothers' progeny in both age groups. In the case of thymus lymphocytes we have not observed any alterations in their angiogenic activity. The differences in histological examination of thymus and lymph nodes were not detected in experimental and control groups. 2000 Academic Press@p$hr Copyright 2000 Academic Press. PMID- 10712834 TI - IMMUNOSUPPRESSION AND RECOVERY OF DRUG-IMPAIRED HOST RESISTANCE AGAINST CANDIDA ALBICANS INFECTION BY OXOGLAUCINE. AB - The immunosuppressive action of aporphinoid alkaloid oxoglaucine was studied in experimental Candida albicans (C. albicans) infection in mice. The alkaloid augmented host resistance to pathogen applied to mice (6-8 weeks of age) at a low dose of 2 mg kg(-1)in 3 days and impaired it at a high dose of 10 mg kg(-1). The suppressive activity observed under the latter schedule correlated with the inhibited proliferative response of splenic cells to mitogens and with decreased popliteal lymph node (PLN) reaction to C. albicans. Treatment of mice with oxoglaucine (at the age of 5 days) at a dose of 5 mg kg(-1)in 3 consecutive days increased the susceptibility to Candida inoculation at the age of 6 weeks. Delayed type hypersensitivity (DTH) response to C. albicans was enhanced after pretreatment of adult mice and was suppressed after administration to newborn mice. Long-time treatment (10 days) with oxoglaucine, cyclophoshamide or prednisolone at a dose of 10 mg kg(-1)increased the rate of mortality of Candida infected mice. Combined pretreatment of mice with cyclophosphamide or prednisolone (5 days at a dose of 5 mg kg(-1)) followed by oxoglaucine (5 days at a dose of 5 mg kg(-1)), prolonged the survival of infected mice. 2000 Academic Press@p$hr Copyright 2000 Academic Press. PMID- 10712835 TI - EFFECTS OF SOME ANTIBIOTICS ON ENZYME ACTIVITY OF GLUCOSE-6-PHOSPHATE DEHYDROGENASE FROM HUMAN ERYTHROCYTES. AB - Inhibitory effects of some antibiotics on glucose-6-phosphate dehydrogenase from the erythrocytes of human have been investigated. For this purpose, at the beginning, erythrocyte glucose-6-phosphate dehydrogenase was purified 13.654 times in a yield of 28% by using ammonium sulphate precipitation and 2',5'-ADP Sepharose 4B affinity gel. Temperature of +4 degrees C was maintained during the purification process. Enzyme activity was determined with the Beutler method by using a spectrophotometer at 340 nm. This method was utilized for all kinetic studies. Sodium ceftizoxime, sodium ampicillin, sodium cefuroxime, sodium cefazolin, sodium cefoperazone, streptomycin sulphate, gentamicin sulphate, and netilmicin sulphate were used as antibiotics. All the antibiotics indicated the inhibitory effects on the enzyme. K(i)constants for glucose-6-phosphate dehydrogenase were found by means of Lineweaver-Burk graphs. While sodium cefoperazone, gentamicin sulphate, and netilmicin sulphate showed competitive inhibition, the others displayed non-competitive inhibition. In addition, I(50)values of the antibiotics were determined by plotting activity percent vs [I]. In addition, in vivo studies were done for sodium sefuroxime in Sprague Dawley type rats. It was found that G6PD in erythrocyte was more inhibited by the drug in 2.5 h. 2000 Academic Press@p$hr Copyright 2000 Academic Press. PMID- 10712836 TI - PREVENTION OF CISPLATIN-INDUCED NEPHROTOXICITY BY METHIMAZOLE. AB - Nephrotoxicity is a dose-limiting factor in the use of cisplatin against solid tumours. Methimazole, an antithyroid drug containing a free SH group, has a nephroprotective potential against chemically-induced nephrotoxicity. We tried to explore the nephrotoxic effect of the experimentally therapeutic dose of cisplatin (7 mg kg(-1), i.p.), particularly on the nuclear level of kidney cells in male albino rats, as well as the possible protective effect of methimazole. Furthermore, the drug interaction regarding the oncolytic effect of cisplatin was examined in Ehrlich ascites carcinoma (EAC)-bearing mice. A single dose of cisplatin caused kidney damage, 6 days after injection, manifested by 219% increase in serum creatinine, 384% increase in blood urea nitrogen and 170% increase in kidney content of lipid peroxides. Kidney DNA showed clear fragmentations detected by gel electrophoresis. However, kidney reduced glutathione was unchanged at that time period. Histological examination of kidney confirmed the toxic effect of cisplatin. Methimazole (40 mg kg(-1), i.p., 30 min before cisplatin injection) significantly protected the kidney from the nephrotoxic effect of cisplatin as judged from the biochemical parameters investigated as well as the histopathological examination. On the other hand, the survival data in EAC-bearing mice treated with both drugs indicated the persistence of an effective cytotoxic action. This study points to a promising use of this combination and necessitates further experimental and clinical studies. 2000 Academic Press@p$hr Copyright 2000 Academic Press. PMID- 10712837 TI - EFFECTS OF NITRIC OXIDE SYNTHESIS INHIBITION ON THE Na,K-ATPase ACTIVITY IN THE KIDNEY. AB - The present study was aimed at investigating the role of endogenous nitric oxide (NO) in regulating Na,K-ATPase activity in the kidney. The expression of alpha-1 and beta-1 subunits; and the enzymatic activity of Na,K-ATPase were determined in the kidney of rats treated with an NO synthase inhibitor, N(G)-nitro- l -arginine methyl ester (l -NAME). Following the treatment with l -NAME in the drinking water for 4 weeks, Na,K-ATPase activity was increased while tissue nitrite/nitrate levels were decreased in the kidney. Supplementation with l arginine prevented the l -NAME-induced changes. The expression of either alpha-1 or beta-1 subunit protein of Na,K-ATPase, assessed by Western blot analysis, was not affected by l -NAME-treatment. An acute in vitro treatment of the kidney with l -NAME also caused an increase of Na,K-ATPase activity; which was again prevented by cotreatment with l -arginine. On the contrary, treatment with sodium nitroprusside significantly decreased Na,K-ATPase activity. These results suggest that the endogenous NO plays a direct inhibitory role on Na,K-ATPase activity in the kidney. 2000 Academic Press@p$hr Copyright 2000 Academic Press. PMID- 10712838 TI - Evolutionary analysis of gamma-carbonic anhydrase and structurally related proteins. AB - We studied the evolutionary relationships between gamma-carbonic anhydrase (gamma CA) and a very diverse group of proteins that share the sequence motif characteristic of the left-handed parallel beta-helix (LbetaH) fold. This sequence motif is characterized by the imperfect tandem repetition of short hexapeptide units, which makes it difficult to obtain a reliable alignment based on sequence information alone. To solve this problem, we used a structural alignment of three members of the group with known crystallographic structures as a seed to obtain a reliable sequence alignment. Then, we applied protein maximum parsimony and maximum-likelihood phylogenetic inference methods to this alignment. We found that gamma-CA belongs to a diverse superfamily of proteins that share the LbetaH domain. This superfamily is composed mainly of acyltransferases. The most remarkable feature of the phylogenetic tree obtained is that its main branches group together functionally related proteins, so that the coarse topology can be rather easily explained in terms of functional diversification. Regarding the main branch of the tree containing gamma-CA, we found that, in addition to the group of its closest relatives that had already been studied, gamma-CA is closely related to the tetrahydrodipicolinate N succinyltransferases. PMID- 10712839 TI - Phylogenetic relationships among bumble bees (Bombus, latreille) inferred from mitochondrial cytochrome b and cytochrome oxidase I sequences. AB - We conducted a molecular study intending to derive an estimate of the relationships within the genus Bombus (bumble bees) by comparing the mitochondrial cytochrome b and cytochrome oxidase I (COI) genes from 19 species, spanning 10 of approximately 16 European subgenera and 3 subgenera from North and South America. Our trees differ from the most recent classifications of bumble bees. Although bootstrap values for deep branches are low, our sequences show significant data structure and low homoplasy, and all trees share some groups and patterns. In all cases, the subgenus Bombus s. str. clusters among the most derived bumble bees, contrary to other molecular studies. In all trees, B. funebris is the sister taxon of B. robustus, and in five of the six trees, B. wurflenii is the sister taxon to this clade. B. nevadensis is basal to the other species in the analysis of the cytochrome b gene, but appears to be among the most derived according to the analysis of the COI region. The species representing the subgenera Thoracobombus and Fervidobombus are consistently among the earliest diverged. Species that appear in very different positions in different trees are B. nevadensis, B. mesomelas, B. balteatus, and B. hyperboreus. All subgenera with two representatives in our analysis are apparently monophyletic except Fervidobombus, Melanobombus, and Pyrobombus. The groups formed by pocket makers and non-pocket makers within Bombus also appear to be paraphyletic, and therefore some subgenera may not accurately reflect phylogeny. PMID- 10712840 TI - The distribution of group I introns in lichen algae suggests that lichenization facilitates intron lateral transfer. AB - The nuclear-encoded small subunit ribosomal DNA gene of many lichen-forming green algae in the genus Trebouxia contains a group I intron at Escherichia coli genic position 1512. We studied the evolutionary history of the 1512 intron in Trebouxia spp. (Trebouxiophyceae) by analyzing intron and "host" cell phylogenies. The host trees were constructed by comparing internal transcribed spacer regions of rDNA. Maximum-likelihood, maximum-parsimony, and distance analyses suggest that the 1512 intron was present in the common ancestor of the green algal classes Trebouxiophyceae, Chlorophyceae, and Ulvophyceae. The 1512 intron, however, was laterally transferred at least three times among later diverging Trebouxia spp. that form lichen partnerships. Intron secondary structure analyses are consistent with this result. Our results support the hypothesis that lichenization may facilitate 1512 group I intron lateral transfer through the close cell-to-cell contact that occurs between the lichen algal and fungal symbionts in the developing lichen thallus. PMID- 10712841 TI - Phylogeny of taxaceae and cephalotaxaceae genera inferred from chloroplast matK gene and nuclear rDNA ITS region. AB - Phylogeny of the Taxaceae genera and the monotypic family Cephalotaxaceae has been extraordinarily controversial. In this paper chloroplast matK genes and nuclear ITS sequences were determined for all six genera of the two families and representatives of other conifer families. Analysis using either the nonsynonymous sites or the deduced amino acid sequences of matK genes strongly indicates that taxad genera and Cephalotaxaceae are monophyletic, with the Taxodiaceae/Cupressaceae clade as their sister group. Cephalotaxus is basal to the taxad genera, among which two clades, Torreya/Amentotaxus and Taxus/Pseudotaxus/Austrotaxus, are resolved. They correspond to Janchen's two tribes, Torreyeae and Taxeae. In Taxeae, Austrotaxus is the first to branch off. Analyses of the nuclear ITS sequence data corroborated the topology of the matK gene tree. These results refute the views that Cephalotaxaceae has no alliance with Taxaceae and that Austrotaxus and Amentotaxus should be excluded from the Taxaceae. We estimated the divergence time between the Taxodiaceae/Cupressaceae and the Cephalotaxaceae/Taxaceae clades to be 192-230 Myr ago and the divergence time between taxads and Cephalotaxus to be 149-179 Myr ago. Soon after the latter divergence event, within 6-8 Myr, the two taxad tribes originated. In conclusion, our data do not support Florin's claim that taxads could be traced to Devonian psilophytes (359-395 Myr ago). PMID- 10712842 TI - How slippage-derived sequences are incorporated into rRNA variable-region secondary structure: implications for phylogeny reconstruction. AB - We analyzed the type and frequency of mutational changes in hypervariable rRNA regions, using the highly length-variable region V4 of the small subunit rRNA locus of tiger beetles (Cicindelidae) as an example. Phylogenetic analysis of indels in closely related species showed that (1) most indels are single nucleotides (usually A or T and sometimes G) or di-nucleotides of A and T. These occur at numerous foci, and they exhibit a strong bias for duplication of 5' single and di-nucleotide motifs but not 3' motifs. (2) Insertions/deletions in stem-forming regions affected paired and unpaired bases with about equal frequency but they did not disrupt the secondary structure. (3) Recurring mutations involving short repeats of the same bases caused parallel evolution of similar sequence motifs in the rRNA of different lineages. The observed types of change are consistent with the propostion that slippage is the main mutational mechanism. Slippage-derived sequences tend to be self-complementary, and therefore the stem-loop structure could be self-organizing as a consequence of the underlying mutational mechanism. Thus, the secondary structure in the cicindelid V4 region may be conserved due to the dynamics of the mutational mechanism rather than to functional constraints. These processes may also have a tendency to produce similar primary sequences irrespective of phylogenetic associations. The findings have implications for sequence alignment in phylogenetic analysis and should caution against the use of secondary structure to improve the determination of positional homology in hypervariable regions. PMID- 10712843 TI - PCR survey of Hox genes in the crinoid and ophiuroid: evidence for anterior conservation and posterior expansion in the echinoderm Hox gene cluster. AB - To help elucidate the cluster organization of Hox genes in echinoderms, we amplified a homeobox region by polymerase chain reaction (PCR) and cloned and sequenced the PCR products for the comatulid crinoid Oxycomanthus japonicus and the ophiuroid Stegophiura sladeni. The crinoid had at least three anterior, four medial, and four posterior genes, and the ophiuroid had at least one anterior, three medial, and six (one of which being a possible trans-paralog) posterior genes. The survey of the crinoid detected all three anterior complements (PG1-3 genes). It was inferred that the Hox genes of each species are organized into a single cluster and that a novel cognate group of echinoderm posterior genes (tentatively termed HboxP9) exists among echinoderms in addition to the known posterior genes Hbox4, Hbox7, and Hbox10. The results, combined with the data of other echinoderm classes, strongly suggest that the presence of a single Hox gene cluster is a common feature among echinoderms and that the cluster has the general features of the deuterostome Hox gene cluster, i.e., the anterior conservation and posterior expansion. The results of the ophiuroid imply that the posterior genes in this class diversified after the phylum Echinodermata had been established. PMID- 10712844 TI - Phylogenetic relationships of xenodontine snakes inferred from 12S and 16S ribosomal RNA sequences. AB - The phylogenetic relationships of xenodontine snakes are inferred from sequence analyses of portions of two mitochondrial genes (12S and 16S ribosomal RNA) in 85 species. Although support values for most of the basal nodes are low, the general pattern of cladogenesis observed is congruent with many independent molecular, morphological, and geographical data. The monophyly of xenodontines and the basal position of North American xenodontines in comparison with Neotropical xenodontines are favored, suggesting an Asian-North American origin of xenodontines. West Indian xenodontines (including endemic genera and members of the genus Alsophis) appear to form a monophyletic group belonging to the South American clade. Their mid-Cenozoic origin by dispersal using ocean currents is supported. Within South American mainland xenodontines, the tribes Hydropsini, Pseudoboini, and Xenodontini are monophyletic. Finally, our results suggest that some morphological and ecological traits concerning maxillary dentition, macrohabitat use, and foraging strategy have appeared multiple times during the evolution of xenodontine snakes. PMID- 10712845 TI - Primate genus Miopithecus: evidence for the existence of species and subspecies of dwarf guenons based on cellular and endogenous viral sequences. AB - Sequence data from the mitochondrial 12S rRNA gene were combined with endogenous retrovirus sequences to study the position of the genus Miopithecus in the primate tree. The mitochondrial sequences indicated that Miopithecus is a true genus distinct from Cercopithecus, although talapoin monkeys are commonly referred to as dwarf guenons. The existence of two species of dwarf guenons, suggested by differences in coat color, pigmentation, and geographic location, was supported by substantial mitochondrial 12S rRNA gene divergence. In line with the informal proposal of J. Kingdon (1997, "The Kingdon Field Guide to African Mammals," Academic Press, London), we use the names Miopithecus talapoin for the southern, darker species and Miopithecus ougouensis for the northern, lighter colored monkeys. Different 12S rRNA gene haplotypes found in M. ougouensis individuals suggest the possible existence of additional subspecies. Simian endogenous retrovirus (SERV) strain 23. 1 proviruses were introduced in the primate germ-line after the Cercopithecinae split from the Colobinae, estimated at around 9-14 million years ago. SERV sequences were used for timing of divergence events in Cercopithecinae and confirmed the close relationship between the genera Cercopithecus and Miopithecus, which was only weakly supported by the more variable mtDNA sequences in a distance analysis, demonstrating the utility of these pseudogenes in phylogenetic grouping. PMID- 10712846 TI - Pre-pleistocene refugia and differentiation between populations of the caucasian salamander (Mertensiella caucasica). AB - A 350-bp fragment of the mitochondrial cytochrome-b gene was sequenced in the Caucasian salamander, Mertensiella caucasica, representing 10 populations from across its range along the Black Sea coast. Five haplotypes were discovered among 65 fragments analyzed, differing at 2-50 positions. The highest differentiation between haplotypes was observed in animals from the eastern part of the species' range (Borjomi) compared to those from the remainder of the species' range. Randomly amplified nuclear DNA revealed a pattern of spatial genetic variation similar to that of the mitochondrial genome. M. caucasica, as currently known, represents two evolutionary lineages that evolved independently, perhaps since the lower Pliocene. These lineages represent taxa, possibly to be described as species, distributed in the Borjomi area in central Georgia and in southwestern Georgia and northeastern Turkey. The multivariate analysis of morphological data did not reveal significant differences between the taxa. However, substantial morphological differentiation was observed within both lineages, showing parallel patterns in body proportions and coloration patterns. This variation is possibly associated with extant ecological conditions. Salamanders with reduced pigmentation from southwestern Georgia were not genetically distinguishable from neighboring populations. PMID- 10712847 TI - Phylogeny of east asian bufonids inferred from mitochondrial DNA sequences (Anura: amphibia). AB - We investigated the relationships of Asian bufonids using partial sequences of mitochondrial DNA genes. Twenty-six samples representing 14 species of Bufo from China and Vietnam and 2 species of Torrentophryne from China were examined. Three samples of Bufo viridis from Armenia and Georgia were also sequenced to make a comparison to its sibling tetraploid species B. danatensis. Bufo americanus, from Canada, was used as the outgroup. Sequences from the 12S ribosomal RNA, 16S ribosomal RNA, cytochrome b, and the control region were analyzed using parsimony. East Asian bufonids were grouped into two major clades. One clade included B. andrewsi, B. bankorensis, B. gargarizans, B. tibetanus, B. tuberculatus, its sister clade B. cryptotympanicus, and the 2 species of Torrentophryne. The second clade consisted of B. galeatus, B. himalayanus, B. melanostictus, and a new species from Vietnam. The placement of three taxa (B. raddei, B. viridis, and its sister species, B. danatensis) was problematic. The genus Torrentophryne should be synonymized with Bufo to remove paraphyly. Because B. raddei does not belong to the clade that includes B. viridis and B. danatensis, it was removed from the viridis species group. The species status of B. bankorensis from Taiwan is evaluated. PMID- 10712848 TI - A phylogenetic study of the malagasy couas with insights into cuckoo relationships. AB - The avian family Cuculidae (cuckoos) is a diverse group of birds that vary considerably in behaviors of interest to behavioral ecologists, e.g., obligate brood parasitism and cooperative breeding. The taxonomy of this group has historically been relatively stable but has not been extensively evaluated using molecular methods. The goal of this study was to evaluate phylogenetic relationships within the ecologically diverse genus Coua and the placement of Coua among major cuckoo lineages. We sequenced 429 bp of cytochrome b (cyt b) and 522 bp of ND2, both mitochondrial genes, for 26 species of cuckoos spanning 13 genera. We also included the enigmatic hoatzin (Opisthocomus hoazin) and used two Tauraco species as outgroups. ND2 exhibited higher rates of DNA sequence and amino acid substitution than cyt b; however, this did not greatly affect the overall levels of phylogenetic resolution and support provided by these two genes. Combined analyses produced two alternative phylogenies, depending on weighting scheme, both of which were fully resolved and were characterized by high bootstrap support. These phylogenies recovered monophyly for all of the traditional cuckoo subfamilies and indicated, with strong support, that the hoatzin is outside of Cuculidae. Within Coua, an arboreal and a terrestrial clade were identified. In contrast, habitat choice of Coua species did not greatly reflect the phylogeny. PMID- 10712849 TI - Phylogeny of ips DeGeer species (Coleoptera: scolytidae) inferred from mitochondrial cytochrome oxidase I DNA sequence. AB - We used 766 bp of DNA sequence data from the mitochondrial cytochrome oxidase I gene to reconstruct a phylogeny for 39 of 43 Ips species, many of which are economically important bark beetles. The phylogeny was reconstructed using equally weighted and weighted parsimony. In both analyses, peripheral clades were well supported while internal clades were poorly supported. Phylogenetic analysis of translated amino acids produced a poorly resolved tree that was discordant with trees reconstructed with nucleotide sequence data. Two main conclusions are drawn about the monophyly of Ips and traditional systematic groups within Ips. First, Ips is monophyletic only when I. mannsfeldi, I. nobilis, and the concinnus and latidens species groups are excluded. The latidens group, I. mannsfeldi, and I. nobilis form a monophyletic group with 3 Orthotomicus species, while the concinnus group has a more basal position. Second, the majority of the species groups in the current classification for Ips are not monophyletic. European Ips species do not form a monophyletic group, contrary to common usage, and are dispersed on the phylogenetic tree among North American species. These results indicate that a formal systematic revision of Ips is needed. PMID- 10712850 TI - Characterization of histone H3/H4 gene region and phylogenetic affinity of Ichthyophthirius multifiliis based on H4 DNA sequence variation. AB - We sequenced the amino-terminal third of the histone H3 and H4 genes and the intergenic region from Ichthyophthirius multifiliis. Fourteen recombinant clones of 646 bp were sequenced and the level of sequence variation detected among these clones was similar to that reported among closely related species of Tetrahymena and to levels of sequence variation detected within other ciliates. The intergenic region is 417 bp and approximately 92% AT rich, making it the longest and most AT-rich ciliate H3/H4 intergenic region yet identified. Similar to Tetrahymena, the intergenic region of Ichthyophthirius contains two CCAAT regions arranged in a complementary orientation. A neighbor-joining tree was constructed based on nucleotide sequence variation among H4 genes to evaluate evolutionary relationships within and among six classes of Ciliophora. The single shortest neighbor-joining tree depicted a sister-group relationship of Ichthyophthirius with taxa of Tetrahymenina, thereby supporting monophyly of Oligohymenophorea. PMID- 10712851 TI - Modes of evolution in the protease and kringle domains of the plasminogen prothrombin family. AB - Phylogenetic analysis of protease domains of the vertebrate plasminogen prothrombin family revealed two major subfamilies: (1) a subfamily containing macrophage-stimulating protein (MSP), hepatocyte growth factor (HGF), plasminogen, and apolipoprotein(a) (APOA); and (2) a subfamily containing prothrombin, HGF activator, and plasminogen activators. There was evidence that these two subfamilies diverged prior to the divergence of amphibians and amniotes. The phylogeny indicated a close relationship of APOA from the European hedgehog, rhesus monkey, and human with plasminogen. Phylogenetic analysis of repeated kringle domains supported the hypothesis that APOA evolved independently in hedgehog and primates through numerous duplications of different kringle domains of the ancestral plasminogen. Phylogenies of kringle domains revealed two modes of evolution: (1) a conservative mode, whereby duplication of kringle domains occurred prior to cladogenesis and the same kringle structure has been maintained in different lineages (exemplified by plasminogen and prothrombin); and (2) a concerted mode, whereby kringle domains have duplicated since cladogenesis and thus orthologous relationships do not exist between kringles of different lineages (exemplified by APOA). PMID- 10712852 TI - Testing hypotheses of vicariance in the agamid lizard Laudakia caucasia from mountain ranges on the northern Iranian Plateau. PMID- 10712853 TI - Juvenile hormone paces behavioral development in the adult worker honey bee. AB - Behavioral development in the adult worker honey bee (Apis mellifera), from performing tasks inside the hive to foraging, is associated with an increase in the blood titer of juvenile hormone III (JH), and hormone treatment results in precocious foraging. To study behavioral development in the absence of JH we removed its glandular source, the corpora allata, in 1-day-old adult bees. The age at onset of foraging for allatectomized bees in typical colonies was significantly older compared with that of sham-operated bees in 3 out of 4 colonies; this delay was eliminated by hormone replacement in 3 out of 3 colonies. To determine the effects of corpora allata removal on sensitivity to changes in conditions that influence the rate of behavioral development, we used "single-cohort" colonies (composed of only young bees) in which some colony members initiate foraging precociously. The age at onset of foraging for allatectomized bees was significantly older compared with that of sham-operated bees in 2 out of 3 colonies, and this delay was eliminated by hormone replacement. Allatectomized bees initiated foraging at significantly younger ages in single-cohort colonies than in typical colonies. These results demonstrate that JH influences the pace of behavioral development in honey bees, but is not essential for either foraging or altering behavioral development in response to changes in conditions. PMID- 10712854 TI - Effect of thyroid hormone administration on estrogen-induced sex behavior in female mice. AB - Effects of thyroid hormones on reproductive processes have been described in sheep, birds, and rats. To extend this subject to mice for eventual analysis with genetically modified animals, we looked for effects of thyroid hormone treatment on lordosis behavior of ovariectomized estrogen-treated female mice. High doses of thyroid hormones reduced lordosis behavior. Since we could not explain this result by pharmacokinetic or peripheral effects, we infer that it worked by a central mechanism. Future investigations must determine whether endogenous fluctuations within the thyroid's normal physiological range have any behavioral effects. PMID- 10712855 TI - Rapid behavioral response to corticosterone varies with photoperiod and dose. AB - The magnitude of the adrenocortical response to stress can be modulated by a variety of environmental and physiological factors, such as daylength and body condition. To determine the consequences of this modulation for the organism, one also needs to investigate behavioral sensitivity to glucocorticoids. We examined the behavioral response of Gambel's white-crowned sparrows (Zonotrichia leucophrys gambelii) to elevated glucocorticoids. Using a behavioral assay in which a rapid and transient dose of corticosterone (CORT: the avian glucocorticoid) rapidly increases perch hopping, we first investigated the photoperiodic regulation of this behavioral response. Intermediate levels of CORT ( approximately 24 ng/ml), which increased activity in sparrows exposed to a long day (breeding) photoperiod, had no behavioral effect in sparrows exposed to a short-day (winter) photoperiod. Hence, the neural mechanisms which regulate the behavioral response to CORT appear to be less sensitive under a winter photoperiod. Using the same behavioral assay, we also measured a dose-response curve for CORT's effects on activity in sparrows exposed to the long-day photoperiod. Intermediate levels (24 and 40 ng/ml) increased activity to threefold background levels, whereas high physiological levels (65 and 97 ng/ml) had no effect. Given that the behavioral response does not increase linearly with CORT, we can no longer assume that modulation of the adrenocortical response to stress will result in linear changes in behavior. PMID- 10712856 TI - Corticosterone treatment has no effect on reproductive hormones or aggressive behavior in free-living male tree sparrows, Spizella arborea. AB - We examined the effect of corticosterone on plasma levels of reproductive hormones (testosterone, dihydrotestosterone, and luteinizing hormone) and territorial defense behavior in male tree sparrows, Spizella arborea. Birds receiving Silastic implants filled with corticosterone (B) had significantly higher plasma levels of B than control birds, which received empty implants, and exhibited pectoral muscle wastage and a decrease in body mass. We evaluated the hormonal and agonistic responses of the two implanted groups of birds using a simulated territorial intrusion (STI) 2 to 4 days after they were implanted. Corticosterone-treated and control birds did not differ in their circulating levels of reproductive hormones or in their behavioral responses to STI (latency to respond to intrusion, number of songs, and closest approach to a decoy and tape recording). Unlike previous studies of north temperate passerines, high physiological levels of exogenous B had no effect either on circulating levels of reproductive hormones or on territorial behaviors associated with breeding. Nonetheless, untreated tree sparrows do mount a robust adrenocortical response, having a two- to fourfold increase in plasma B levels during a 1-h period of capture. Thus, adrenocortical responsiveness is maintained in these birds, but elevated levels of glucocorticoids do not suppress reproductive hormones or associated behaviors. We believe that this hormonal and behavioral refractoriness to glucocorticoids-or uncoupling of the stress response from the reproductive axis-may be advantageous for species having extreme temporal constraints on their breeding schedules. PMID- 10712857 TI - Paternal mouthbrooding in the black-chinned tilapia, Sarotherodon melanotheron (Pisces: cichlidae): changes in gonadal steroids and potential for vitellogenin transfer to larvae. AB - The black-chinned tilapia (Sarotherodon melanotheron) is a paternal mouthbrooder. Pairs of adult black-chinned tilapia were raised in freshwater and the males were sampled during the mouthbrooding cycle. Sampling also occurred 10 days after release of the free-swimming fry for comparison. During the first week of incubation of the eggs, total androgens and estradiol were low (<5 and <0.3 ng/ml, respectively). During the second week of brooding, when the eggs have hatched and they are called newly hatched embryos, plasma levels of gonadal steroids increased (13-38 ng androgen/ml and >0.6 ng estradiol/ml). The plasma concentrations of vitellogenin (VTG) in male parents changed during mouthbrooding, with decreases occurring between egg pickup and hatching of the embryo (Day 6 of mouthbrooding). The pattern of change in concentrations of VTG in surface mucus of male parents differed from the pattern in plasma, with peak concentrations occurring at the time of hatching. The amount of VTG in mucus was similar to that measured in the female Oreochromis mossambicus during mouthbrooding of embryos. The appearance of peak VTG levels in the mucus at the time of hatching when plasma levels have declined and the availability of comparable amounts of mucus VTG in both maternal and paternal mouthbrooding tilapia, despite unequivalent plasma levels, support the possibility that parental provisioning of the young occurs during mouthbrooding in tilapia. PMID- 10712858 TI - Peripheral pulses of oxytocin increase partner preferences in female, but not male, prairie voles. AB - Centrally administered oxytocin (OT) facilitates social behaviors including the partner preferences that characterize the monogamous social system of prairie voles. In contrast peripherally administered OT generally has been ineffective in influencing central processes including behavior. OT from the posterior pituitary gland is released in pulses into the peripheral circulation. We hypothesized that peripherally administered OT, if delivered in repeated injections mimicking these pulses, would influence behavior. Male and female prairie voles received three subcutaneous injections of OT, a single injection of OT, or isotonic saline. Animals then were placed with an adult member of the opposite sex, designated as a "partner," for a 1-h period of cohabitation, and subsequently tested for preference for the familiar partner versus a comparable stranger. Females treated with pulses of peripheral OT (1, 5, or 20 microg) displayed a significant preference for the partner compared to control females, while females receiving a lower dose of OT (0.1 microg) or a single injection (20 microg) did not. There was also a significant within-group effect as pulsed OT-treated females spent more time with the partner when compared to the stranger, while control females spent equal amounts of time with the partner and stranger. Peripheral pulses of OT were no longer effective in inducing partner preferences when females were pretreated with a selective OT receptor antagonist, administered either peripherally or centrally. In contrast to females, peripheral treatment with OT did not facilitate the formation of partner preferences in males. PMID- 10712859 TI - Psychological state and mood effects of steroidal chemosignals in women and men. AB - We tested the hypothesis that isolated steroids, claimed to act like pheromones, affect human psychological state or mood. In the first experiment, we established that two steroids, Delta4, 16-androstadien-3-one and 1,3,5(10)16-estratetraen-3 ol, modulated emotional states within 6 min of exposure. In men and women, neither steroid had specific effects on states of alertness or negative-confused mood. However, both steroids increased positive stimulated mood state in women but decreased it in men. These psychological findings do not parallel the reported sexually specific effects of these two steroids on the surface potential activity of putative vomeronasal epithelium. In a second experiment on women, we replicated that Delta4,16-androstadien-3-one modulated their general mood state, even when women were not aware of its odor and gave identical olfactory descriptions for the steroid and the control carrier solutions. In this within subjects, repeated-measures experiment, androstadienone prevented the deterioration in general mood which occurred during exposure to the clove oil carrier solution in the laboratory environment. Thus, androstadienone appears to modulate affect, rather than releasing stereotyped behaviors or emotions. It is premature to call these steroids human pheromones. They are nonetheless psychologically potent, mandating future work delineating their function-i.e., whether these steroids are communicative chemosignals, context specific, or related to unconscious associations. In light of these modulatory effects and the complexity of human behavior, the function of chemosignals and pheromonal systems in a variety of species may need to be expanded to include the concept of modulators, as well as the traditional releasers, primers, and signaling compounds. PMID- 10712860 TI - Perinatal exposure to estradiol masculinizes aspects of sexually dimorphic behavior and morphology in gray short-tailed opossums (Monodelphis domestica). AB - The effects on adult sexually dimorphic behavior of perinatal exposure to estrogen were examined by treating male and female gray opossums with estradiol (EST), an estrogen receptor antagonist (tamoxifen:TX) or oil control (OIL) during the first week of life, a time period corresponding in this marsupial to late gestation in rodent species. Following gonadectomy and replacement therapy with testosterone in adulthood, males showed more scent-marking behavior than females and EST animals showed more scent marking than TX or OIL animals. Also, phalluses were longer and body weight was higher in males than in females and in EST treated animals than in TX-treated animals; OIL animals were intermediate in these morphological measures. EST animals of both sexes showed less female typical screeching threat behavior than OIL or TX animals. Because these hormone manipulations were conducted on the "fetus" directly in this marsupial (rather than via the maternal circulation as in previously studied eutherian species), these findings provide unique confirming evidence for masculinization of aspects of behavior and morphology by early exposure to estradiol in mammals. PMID- 10712862 TI - Appetitive and consummatory sexual behaviors of female rats in bilevel chambers. II. Patterns of estrus termination following vaginocervical stimulation. AB - Copulation with intromission or manual vaginocervical stimulation (VCS) shortens the duration that intact female rats maintain lordosis responding during estrus. The present study examined whether VCS could shorten the duration of both appetitive and consummatory measures of female sexual behavior, and whether these effects occur differentially in time and across different hormone priming intervals. Ovariectomized, sexually experienced female rats were administered subcutaneous injections of estradiol benzoate 48 h and progesterone 4 h, before receiving 50 manual VCSs with a lubricated glass rod distributed over 1 h. Control females received sham VCSs distributed over the same time. The females were then tested for sexual behavior in bilevel chambers with two sexually vigorous males (to one ejaculatory series or 10 min with each male, separated by 5 min) 12, 16, and 20 h after VCS. Prior to the final hormone treatment, different groups of females had been given the same hormone treatment either 28, 14, 7, or 4 days before. In females tested at 28- and 14-day hormone intervals, VCS induced both active and passive rejection responses at 12, 16, and 20 h. In contrast, females that received sham VCS displayed relatively normal sexual behavior at 12 h, although by 16 and 20 h these females displayed active and passive rejection. Females tested at 7- or 4-day intervals displayed normal levels of lordosis at all testing times, regardless of VCS treatment. These data indicate that VCS facilitates rejection responses that precede the decrease in lordosis responsiveness. However, the effects of VCS are dependent on the frequency of hormone priming, suggesting that hormone treatment may block some of the long-term inhibitory effects of VCS on female sexual behavior. PMID- 10712861 TI - Spatial working memory and hippocampal size across pregnancy in rats. AB - The present experiments investigated the effects of pregnancy on performance in the Morris water maze and on hippocampal volume. In the first study, pregnant rats (in between the first and second trimester) outperformed nonpregnant rats on the Morris water maze on 1 day of testing. In the second study, rats were tested in a working memory variation of the maze in which the spatial location of the platform varied. Pregnant females traveled shorter distances than nonpregnant females during the first two trimesters, but performed worse than nonpregnant females during the third trimester. Latency measures showed a similar profile. Group differences in performance were not related to changes in swim speed. However, changes in performance in pregnant females may be related to estrogen, progesterone, and/or corticosterone levels during pregnancy, with low levels of estradiol and high levels of progesterone being associated with better performance. There were no significant differences between pregnant and nonpregnant animals on any of the brain measures, although pregnant animals tended to have a smaller hippocampus than nonpregnant animals. These results indicate that pregnancy can affect performance, possibly related to the hormonal changes that accompany pregnancy. PMID- 10712863 TI - The VUV Absorption Spectrum of Lead Monoxide. AB - The VUV absorption spectrum of PbO vapor down to 300 A has been observed for the first time. Four ionization energies have been determined from Rydberg series at 8.73, 8.83, 12.81, and 26.28 eV corresponding to the X(2)Pi, A(2)Sigma(+), B(2)X(+), and C(2)Sigma(+) electronic states of the PbO(+) ion. Copyright 2000 Academic Press. PMID- 10712864 TI - The angleICI Bending Satellites in the Millimeter-Wave Rotational Spectra of CH(2)I(2) and CD(2)I(2). AB - Rotational transitions in the first four excited states of the low-frequency angleICI bending mode, nu(4), have been assigned in the mm-wave rotational spectra of CH(2)I(2) and of CD(2)I(2). Measurements of transition frequencies, made over the frequency region 167-326 GHz and for J" up to 190, allowed determination of sextic level spectroscopic constants for all states. The changes in spectroscopic constants with vibrational excitation show very small anharmonicity, in spite of the very low frequency of this mode (121 cm(-1)). Vibrational excitation affects the moments of inertia in such a way that the planar moment P(b), about the plane perpendicular to both angleICI and angleHCH, is practically invariant. Vibrational change in P(c), the moment along the principal axis in the HCH plane and perpendicular to the angleHCH bisector, has been successfully reproduced with an ab initio harmonic force field so that there is no discernible vibrational change in angleHCH on excitation of angleICI. Finally, the change in P(a) leads to estimated vibrational change of +0.12 degrees in the value of angleICI itself. Copyright 2000 Academic Press. PMID- 10712865 TI - The nu(2), nu(4) + nu(5), nu(6) + nu(9), and nu(8) Band System in 1,1,1 Trifluoroethane. AB - High-resolution FTIR spectra of 1,1,1-trifluoroethane (HFC-143a) have been recorded in the region from 1370 to 1470 cm(-1) with an unapodized resolution of 0.0016 cm(-1) at room temperature and of 0.004 cm(-1) at 183 and 100 K. The two main infrared active bands of A(1) symmetry have been shown to be nu(2) at 1407.5 cm(-1) and nu(4) + nu(5) at 1440.5 cm(-1). With the aid of Raman spectra, the two infrared inactive bands of E symmetry in this spectra region have been shown to be nu(8) at 1457.5 cm(-1) and nu(6) + nu(9) at 1446.2 cm(-1). The nu(2) band was analyzed as an isolated band, whereas the nu(4) + nu(5) band was analyzed as part of the triad nu(4) + nu(5), nu(6) + nu(9), and nu(8). Copyright 2000 Academic Press. PMID- 10712866 TI - Electronic Spectroscopy of Rhodium Mononitride. AB - We report the first observation of gas-phase electronic spectra for rhodium mononitride. The RhN molecules have been produced in the reaction of laser ablated rhodium metal with ammonia. Many vibronic bands have been studied in the 400-700 nm region using laser-induced fluorescence. Rotational analyses of the stronger of these, together with excited state lifetime measurements and Rh(14)N Rh(15)N isotopic shifts, identify three electronic systems in the region: [15.1]1 X(1)Sigma(+), [19.5]0(+)-X(1)Sigma(+), and [22.4]0(+)-X(1)Sigma(+) with (0, 0) bands near 15 071, 19 489, and 22 385 cm(-1), respectively. The (1)Sigma(+) symmetry for the ground state agrees with theoretical predictions. Dispersed fluorescence spectra have been recorded which reveal the presence of electronic states at T = 553, 1740, and 3920 cm(-1). Copyright 2000 Academic Press. PMID- 10712867 TI - Millimeter-Wave Spectrum of CF(2)Cl(2), Taking into Account the Hyperfine Structure. AB - A new investigation of the CF(2)(35)Cl(2) ground state rotational spectrum in the 50-100 GHz frequency range gave an opportunity to improve the accuracy of rotational and quartic distortion parameters by an order of magnitude and to determine values of six sextic centrifugal distortion parameters. A special technique was used to process line profiles of unresolved or partially resolved hyperfine structures of rotational transitions. It allowed us to improve significantly values for the diagonal parameters of quadrupole coupling chi(aa), (chi(bb)-chi(cc)) and to determine the off-diagonal parameter chi(ab). Copyright 2000 Academic Press. PMID- 10712868 TI - Microwave Torsional-Rotational Spectra of gauche CH(3)CD(2)OH and CH(3)CD(2)OD. AB - The microwave torsional-rotational spectra of gauche CH(3)CD(2)OH and CH(3)CD(2)OD have been identified, assigned, and analyzed up to 70 GHz. From the observed a- and c-dipole transitions, it has been possible to determine the effective rotational coefficients and the gauche tunneling energy of the hydroxyl internal rotation. The product of inertia terms I(xy) and I(xz) were included in the analyses using the framework fixed axis method (FFAM) approach to the hydroxyl internal rotation. Further, the analyses were sensitive to selected effective centrifugal distortion coefficients. For CH(3)CD(2)OH, a-dipole lines were assigned for the first excited gauche state. As for CH(3)CH(2)OH, these lines were highly nonrigid rotor in behavior more than likely due to the resonance with the first excited state of the methyl torsion. Copyright 2000 Academic Press. PMID- 10712869 TI - Electronic Spectrum of the Antimony Monoxide Radical. AB - The 0-1 band (4271.3 A) of the B(2)Sigma(+)-X(2)(2)Pi(3/2) transition, the 0-6 band (4275 A) of the H(2)Pi(3/2)-X(2)(2)Pi(3/2) transition, the 0-0 band (3776.2 A) of the B(2)Sigma(+)-X(1)(2)Pi(1/2) transition, and the 2-0 (3251.3 A) as well as the 3-0 (3194.2 A) bands of the C(2)Sigma(-)-X(1)(2)Pi(1/2) transition of SbO have been photographed at high resolution using the single isotope of antimony (121)Sb. An unambiguous analysis of the rotational structure of all these bands has been carried out for the first time. The magnitude and sign of the Lambda doubling constant p(0) of the X(1)(2)Pi(1/2) state have been determined and as a result of that, the symmetry of the C state has been uniquely identified as C(2)Sigma(-). Accurate rotational constants of all the states involved have been determined by a global linear least-squares fit. Copyright 2000 Academic Press. PMID- 10712870 TI - Rovibrational Spectroscopy of the v(5) = 1 Level of (28)SiDF(3). AB - The nu(5) fundamental band of trifluorosilane-d (SiDF(3)) at 627 cm(-1) was studied for the first time by high-resolution FTIR spectroscopy at a resolution of 2.4 x 10(-3) cm(-1). The analysis was performed simultaneously with available microwave and newly measured submillimeter-wave data in the approximation of an isolated degenerate fundamental level of a C(3 Kv) symmetric top molecule leading to a standard deviation of 0.22 x 10(-3) cm(-1) for the reproduction of the infrared wavenumbers, 36 kHz for the microwave, and 198 kHz for the submillimeter wave frequencies, respectively. The unitary equivalence between the two reductions (Q and D) of the effective Hamiltonian applied in the analysis is demonstrated. Copyright 2000 Academic Press. PMID- 10712871 TI - High-Order Resonance Interactions in HDO: Analysis of the Absorption Spectrum in the 14 980-15 350 cm(-1) Spectral Region. AB - The absorption spectrum of the HDO molecule recorded by intracavity laser absorption spectroscopy in the 14 980-15 350 cm(-1) spectral region was assigned and modeled in the frame of the effective Hamiltonian approach. The spectrum (496 lines) results, mainly, from transitions to the rotational sublevels of the (014) bright state. An important number of transitions involving the (142) and (0 12 0) highly excited bending states could be identified, borrowing their intensities through high-order resonance interactions with the (014) state. An original feature shown by the present analysis is that all the transitions involving unperturbed energy levels of the (014) state are exclusively of A type, while both A- and B-type transitions are observed when the upper states are perturbed by the resonance interactions. One hundred forty-five energy levels of the three interacting states were derived from the spectrum and fitted to the effective rotational Hamiltonian in Pade-Borel approximants form with 29 varied parameters yielding an rms deviation of 0.038 cm(-1). A few energy levels are affected by additional local resonances with perturbers which have been identified. Finally, 48 transitions of the very weak 6nu(1) band were assigned and fitted as an isolated band. Copyright 2000 Academic Press. PMID- 10712872 TI - Rotational Spectra of the Thiosulfeno Radical, HSS and DSS, between 0.3 and 0.9 THz. AB - The rotational spectra of the HSS and DSS radicals were studied in selected regions between 331 and 883 GHz. The radicals were produced by discharging a gaseous mixture of hydrogen (or deuterium) and hydrogen sulfide in the cell. The observation of the b-type Q-branch and R-branch lines with K(a) = 2-1 and 3-2 for HSS and DSS, respectively, as well as the a-type R-branch lines allowed the improvement and the determination of the molecular constants among them (in MHz) We have reevaluated the harmonic force field of HSS and the ground state average and approximate equilibrium structural parameters. For the latter, we obtained r(HS) = 135.23 pm, r(SS) = 196.03 pm, and angle = 101.74 degrees. These results are compared with those from previous and own quantum chemical calculations as well as with results of related molecules. Copyright 2000 Academic Press. PMID- 10712873 TI - First Observation of Hyperfine Structure in K(2). AB - The hyperfine structure of the (v'-v") = (27-0) band of the b(3)Pi(u0(+)) <-- X(1)Sigma(+)(g) transition has been observed by laser excitation spectroscopy in a highly collimated molecular beam. Hyperfine parameters for magnetic dipole and electric quadrupole interaction are derived from least-squares fitting of the hyperfine pattern and deperturbation between A(1)Sigma(+)(u) and b(3)Pi(u0(+)). Copyright 2000 Academic Press. PMID- 10712874 TI - The High-Resolution FTIR Spectrum of the nu(1) Band of DOBr. AB - Fourier transform infrared spectra of the nu(1) bands of DO(79)Br and DO(81)Br have been recorded with a resolution of 0.0055 cm(-1) in the frequency range of 2510-2800 cm(-1). A total of 1901 lines of the A/B hybrid-type bands of both isotopic species have been assigned and fitted to upper state rovibrational constants employing a Watson's S-reduced Hamiltonian up to sextic terms. The K(a) >/= 4 subband transitions were found to be perturbed and were not included in the fit. The unperturbed band centers for nu(1) of DO(79)Br and DO(81)Br are 2668.79211 +/- 0.00006 and 2668.78842 +/- 0.00005 cm(-1), respectively. The ratio of the vibrational dipole transition moments (u(a):u(b)) was found to be 1.30 +/- 0.04 for the band. Copyright 2000 Academic Press. PMID- 10712875 TI - Uniform Supersonic Expansion for FTIR Absorption Spectroscopy: The nu(5) Band of (NO)(2) at 26 K. AB - A high-resolution Fourier transform interferometer (Bruker IFS 120 HR) was combined with a uniform supersonic expansion produced by means of axisymmetric Laval nozzles. The geometry profile of the nozzle enabled us to work under precise thermodynamic and kinetic conditions. The effect of the cooling rate of different nozzles on cluster nucleation is illustrated. The experimental sensitivity was tested by recording the nu(5) band of (NO)(2) at 26 K. Copyright 2000 Academic Press. PMID- 10712876 TI - The Near-Infrared Transition of CuCl Observed by Intracavity Laser Spectroscopy. AB - The near-infrared electronic transition of CuCl, occurring in the region of 745 nm, was recorded using intracavity laser absorption spectroscopy. The (0, 0), (1, 1), and (2, 2) vibronic bands were analyzed, and from this the molecular constants for the two electronic states were derived. Originally assigned as A' (3)Sigma(+)-X(1)Sigma(+), we have confirmed that this transition does not connect to the ground state, but occurs between two unknown excited states. The excited CuCl molecules were produced in a copper hollow cathode, operated using argon and a small amount of CCl(4). Line positions were referenced to iodine spectra observed from a heated extracavity cell using the broadband spectral output of the intracavity laser as the light source. Copyright 2000 Academic Press. PMID- 10712877 TI - High-Resolution Infrared Spectrum of BrCN in the nu(2) and nu(1)/2nu(2) Regions. AB - FT infrared spectra of BrCN have been recorded in the region of the nu(2) band near 340 cm(-1), the nu(1) band near 580 cm(-1), and the 2nu(2) band near 690 cm( 1) with a resolution between 2.9 and 4.7 x 10(-3) cm(-1). The vibrational levels (01(1)0), (10(0)0), (02(0)0), (02(2)0), (11(1)0), and (20(0)0) have been analyzed employing cold bands, hot bands, and new millimeter-wave transitions. Band-by band polynomial analyses and a combined fit of all data relevant to the 2v(1) + v(2) = 2 polyad levels have been performed. The latter fit considered l-resonance interactions between the (02(0)0), e and (02(2)0), e levels and Fermi resonance between the two Sigma states (10(0)0) and (02(0)0). Altogether about 1000 pieces of data up to J = 100 were fitted for each of the two isotopic species with rms of the residuals of 2-8 x 10(-4) cm(-1) for the infrared and 10-120 kHz for the pure rotational data. Copyright 2000 Academic Press. PMID- 10712878 TI - The Millimeter- and Submillimeter-Wave Spectrum and the Dipole Moment of Ethylenimine. AB - The rotational spectrum of ethylenimine (aziridine, c-C(2)H(4)NH) has been investigated in selected regions from 118 to 950 GHz using the Cologne terahertz spectrometer. About 320 lines have been measured spanning the quantum numbers 2 85% identical to each other and have indistinguishable activities in vitro, we show here that the Drosophila beta isoform has a distinct biological role. We show that PP1beta9C corresponds to flapwing (flw), previously identified mutants of which are viable but flightless because of defects in indirect flight muscles (IFMs) [8]. We have isolated a new, semi-lethal flw allele that shows a range of defects, especially in muscles, which break away from their attachment sites and degenerate. PMID- 10712909 TI - Exclusion of CD45 from the T-cell receptor signaling area in antigen-stimulated T lymphocytes. AB - T lymphocytes are activated by the engagement of their antigen receptors (TCRs) with complexes of peptide and major histocompatibility complex (MHC) molecules displayed on the cell surface of antigen-presenting cells (APCs) [1]. An unresolved question of antigen recognition by T cells is how TCR triggering actually occurs at the cell-cell contact area. We visualized T-cell-APC contact sites using confocal microscopy and three-dimensional reconstruction of z sections. We show the rapid formation of a specialized signaling domain at the T cell-APC contact site that is characterized by a broad and sustained area of tyrosine phosphorylation. The T-lymphocyte cell-surface molecule CD2 is rapidly recruited into this signaling domain, whereas TCRs progressively percolate from the entire T-cell surface into the phosphorylation area. Remarkably, the highly expressed phosphatase CD45 is excluded from the signaling domain. Our results indicate that physiological TCR triggering at the T-cell-APC contact site is the result of a localized alteration in the balance between cellular kinases and phosphatases. We therefore provide experimental evidence to support current models of T-cell activation based on CD45 exclusion from the TCR signaling area [2] [3] [4]. PMID- 10712911 TI - Keep the motor running PMID- 10712910 TI - Double-stranded RNA interference shows that Engrailed controls the synaptic specificity of identified sensory neurons. AB - The transcription factor Engrailed (En) controls the topography of axonal projections by regulating the expression of cell-adhesion molecules [1] [2] [3] [4] but it is not known whether it also controls the choice of individual synaptic target cells. In the cercal sensory system of the larval cockroach (Periplaneta americana), small numbers of identified wind-sensitive sensory neurons form highly specific synaptic connections with 14 identified giant interneurons [5] [6], and target-cell choice is independent of the pattern of axonal projections [6]. En is a putative positional determinant in the array of cercal sensory neurons [7]. In the present study, double-stranded RNA (dsRNA) interference [8] was used to abolish En expression. This treatment changed the axonal arborisation and synaptic outputs of an identified En-positive sensory neuron so that it came to resemble a nearby En-negative cell, which was itself unaffected. We thus demonstrate directly that En controls synaptic choice, as well as axon projections. PMID- 10712914 TI - The road less travelled PMID- 10712912 TI - Molecular motors. PMID- 10712915 TI - Protein phosphatase 2A: a panoply of enzymes. AB - Protein phosphatase 2A describes an extended family of intracellular protein serine/threonine phosphatases sharing a common catalytic subunit that regulates a variety of processes by means of diverse regulatory subunits. During the past year, studies have shown that protein phosphatase 2A influences events ranging from the initiation of DNA replication to vertebrate axis formation to apoptosis. PMID- 10712916 TI - Regulation of cAMP and cGMP signaling: new phosphodiesterases and new functions. AB - The past eighteen months have provided much progress in the cyclic nucleotide phosphodiesterase (PDE) field. Six new phosphodiesterase genes have been discovered and characterized. In addition, several new highly specific PDE inhibitors have been developed and approved for clinical use. Finally, new strategies have been employed to determine PDE function in model systems including the use of antisense oligonucleotide and disruption techniques. PMID- 10712917 TI - Farnesyltransferase inhibitors: antineoplastic mechanism and clinical prospects. AB - Recent work suggests that farnesyltransferase inhibitors suppress cancer cell proliferation through mechanisms other than inhibiting Ras isoprenylation, which is not a crucial event. Recent evidence also suggests that the antineoplastic properties of farnesyltransferase inhibitors are due to alterations in the isoprenylation of RhoB, an endosomal Rho protein that functions in receptor trafficking. A shift in conceptual focus from Ras to Rho to understand how farnesyltransferase inhibitors act provides a new vantage to address old questions in the field and suggests strategies to improve and potentially widen clinical applications. PMID- 10712918 TI - A-kinase anchoring proteins: protein kinase A and beyond. AB - Compartmentalization of kinases and phosphatases is a key determinant in the specificity of second messenger mediated signaling events. Localization of the cAMP-dependent protein kinase (PKA) and other signaling enzymes is mediated by interaction with A-kinase anchoring proteins (AKAPs). In the past year there have been many advances in our understanding of AKAPs, particularly in the field of the functional consequences of PKA anchoring. PMID- 10712919 TI - Regulation of signal transduction by endocytosis. AB - Endocytosis of ligand-activated receptors has generally been considered a mechanism to attenuate signaling. There is now a growing body of evidence suggesting that this process is much more sophisticated and that endocytic membrane trafficking regulates both the intensity of signaling and the co localization of activated receptors with downstream signaling molecules. PMID- 10712920 TI - Signalling through the insulin receptor. AB - The insulin receptor substrates function at the heart of the insulin signalling network. It has recently become apparent that the intracellular localisation of these molecules is regulated in a precise manner that is critical for both the generation and the termination of the insulin signal. Some insulin receptor substrate isoforms appear to be associated with an insoluble matrix that resembles the cytoskeleton. When inappropriately dissociated from this matrix the signalling network collapses concomitant with loss of insulin sensitivity. PMID- 10712921 TI - Signal transduction: hanging on a scaffold. AB - Recent data concerning scaffolding proteins profoundly challenge our conceptions of multicomponent signal transduction systems. Recent studies of the phototransduction system in Drosophila suggest two points. First, scaffolding markedly limits the possibilities for signal amplification. Second, the methods generally available to study signal transduction may be too crude to assess the in vivo roles of scaffolds. Studies of the mitogen-activated protein kinase pathway scaffold, Ste5, indicate functions beyond that of a passive structural element. Finally, the identification of new mitogen-activated protein kinase pathway scaffolds suggests the existence of multiple 'signalosomes' or 'transducisomes'. PMID- 10712922 TI - Cell-substrate interactions and signaling through ILK. AB - Interactions between cells and the extracellular matrix (ECM) result in the regulation of cell growth, cell differentiation and cell migration. These interactions are mediated by integrins and growth factor receptors and intracellular effectors that couple these receptors to downstream components are key to the transduction of ECM signals. This review summarizes recent advances in our understanding of signal transduction via integrins, focusing on the role of integrin-linked kinase in some of these pathways. Research into this interesting protein is uncovering novel aspects of coordinated signaling by the ECM and growth factors. PMID- 10712923 TI - The Ras branch of small GTPases: Ras family members don't fall far from the tree. AB - The Ras branch of the Ras superfamily consists of small GTPases most closely related to Ras and include the R-Ras, Rap, Ral, Rheb, Rin and Rit proteins. Although our understanding of Ras signaling and biology is now considerable, recent observations suggest that Ras function is more complex than previously believed. First, the three Ras proteins may not be functionally identical. Second, Ras function involves functional cross-talk with their close relatives. PMID- 10712924 TI - The genetics of the protein 4.1 family: organizers of the membrane and cytoskeleton. AB - Protein 4.1 (also called band 4.1 or simply 4.1) was originally identified as an abundant protein of the human erythrocyte, in which it stabilizes the spectrin/actin cytoskeleton. The protein and its relatives have since been found in many cell types of metazoan organisms and they are often concentrated in the nucleus, as well as in cell-cell junctions. They form multimolecular complexes with transmembrane and membrane-associated proteins, and these complexes may be important for both structural stability and signal transduction at sites of cell contact. PMID- 10712925 TI - Smads as transcriptional co-modulators. AB - The Smad signalling pathway is critical for transmitting transforming growth factor-beta (TGF-beta) superfamily signals from the cell surface to the nucleus. In the nucleus, Smads regulate transcriptional responses by recruiting co activators and co-repressors to a wide array of DNA-binding partners. Thus, Smads function as transcriptional co-modulators to regulate TGFbeta-dependent gene expression. PMID- 10712926 TI - Cholesterol in signal transduction. AB - Membrane cholesterol impinges on signal transduction in several ways, which is highlighted in particular by the Hedgehog signaling pathway. In Hedgehog signaling, cholesterol is important for ligand biogenesis, as well as for signal transduction in receiving cells. Hedgehog ligands are post-translationally modified by cholesterol, and the Hedgehog receptor, Patched, is structurally similar to the Niemann-Pick C1 protein, which functions in intracellular lipid transport. Although the exact role of cholesterol in Hedgehog signal transduction remains elusive and is probably multifaceted, studies over the past year have implicated raft membrane subdomains, cholesterol transport and a link between protein and lipid trafficking in endocytic compartments. PMID- 10712927 TI - Protein phosphatases and the regulation of mitogen-activated protein kinase signalling. AB - The magnitude and duration of signalling through mitogen- and stress-activated kinases are critical determinants of biological effect. This reflects a balance between the activities of upstream activators and a complex regulatory network of protein phosphatases. These mitogen-activated protein kinase phosphatases include both dual-specificity (threonine/tyrosine) and tyrosine-specific enzymes, and recent evidence suggests that a single mitogen-activated protein kinase isoform may be acted upon by both classes of protein phosphatase. In both cases, substrate selectivity is determined by specific protein-protein interactions mediated through noncatalytic amino-terminal mitogen-activated protein kinase binding domains. Future challenges include the determination of exactly how this network of protein phosphatases interacts selectively with mitogen-activated protein kinase signalling complexes to achieve precise regulation of these key pathways in mammalian cells. PMID- 10712928 TI - Predictive, structure-based model of amino acid recognition by nonribosomal peptide synthetase adenylation domains. AB - BACKGROUND: Nonribosomal peptide synthetases (NRPSs) are large modular proteins that selectively bind, activate and condense amino acids in an ordered manner. Substrate recognition and activation occurs by reaction with ATP within the adenylation (A) domain of each module. Recently, the crystal structure of the A domain from the gramicidin synthetase (GrsA) with L-phenylalanine and adenosine monophosphate bound has been determined. RESULTS: Critical residues in all known NRPS A domains have been identified that align with eight binding-pocket residues in the GrsA A domain and define sets of remarkably conserved recognition templates. Phylogenetic relationships among these sets and the likely specificity determinants for polar and nonpolar amino acids were determined in light of extensive published biochemical data for these enzymes. The binding specificity of greater than 80% of the known NRPS A domains has been correlated with more than 30 amino acid substrates. CONCLUSIONS: The analysis presented allows the specificity of A domains of unknown function (e.g. from polymerase chain reaction amplification or genome sequencing) to be predicted. Furthermore, it provides a rational framework for altering of A domain specificity by site-directed mutagenesis, which has significant potential for engineering the biosynthesis of novel natural products. PMID- 10712929 TI - Organelle pH studies using targeted avidin and fluorescein-biotin. AB - BACKGROUND: Mammalian organelles of the secretory pathway are of differing pH. The pH values form a decreasing gradient: the endoplasmic reticulum (ER) is nearly neutral, the Golgi is mildly acidic and the secretory granules are more acidic still ( approximately pH 5). The mechanisms that regulate pH in these organelles are still unknown. RESULTS: Using a novel method, we tested whether differences in H(+) 'leak' and/or counterion conductances contributed to the pH difference between two secretory pathway organelles. A pH-sensitive, membrane permeable fluorescein-biotin was targeted to endoplasmic-reticulum- and Golgi localized avidin-chimera proteins in HeLa cells. In live, intact cells, ER pH (pH(ER)) was 7.2 +/- 0.2 and Golgi pH (pH(G)) was 6.4 +/- 0.3 and was dissipated by bafilomycin. Buffer capacities of the cytosol, ER and Golgi were all similar (6-10 mM/pH). ER membranes had an apparent H(+) permeability three times greater than that of Golgi membranes. Removal of either K(+) or Cl(-) did not affect ER and Golgi H(+) leak rates, or steady-state pH(G) and pH(ER). CONCLUSIONS: The Golgi is more acidic than the ER because it has an active H(+) pump and fewer or smaller H(+) leaks. Neither buffer capacity nor counterion permeabilities were key determinants of pH(G), pH(ER) or ER/Golgi H(+) leak rates. PMID- 10712930 TI - A Src SH2 selective binding compound inhibits osteoclast-mediated resorption. AB - BACKGROUND: The observations that Src(-/-) mice develop osteopetrosis and Src family tyrosine kinase inhibitors decrease osteoclast-mediated resorption of bone have implicated Src in the regulation of osteoclast-resorptive activity. We have designed and synthesized a compound, AP22161, that binds selectively to the Src SH2 domain and demonstrated that it inhibits Src-dependent cellular activity and inhibits osteoclast-mediated resorption. RESULTS: AP22161 was designed to bind selectively to the Src SH2 domain by targeting a cysteine residue within the highly conserved phosphotyrosine-binding pocket. AP22161 was tested in vitro for binding to SH2 domains and was found to bind selectively and with high affinity to the Src SH2 domain. AP22161 was further tested in mechanism-based cellular assays and found to block Src SH2 binding to peptide ligands, inhibit Src dependent cellular activity and diminish osteoclast resorptive activity. CONCLUSIONS: These results indicate that a compound that selectively inhibits Src SH2 binding can be used to inhibit osteoclast resorption. Furthermore, AP22161 has the potential to be further developed for treating osteoporosis. PMID- 10712931 TI - Sequence specific alkylation of DNA by hairpin pyrrole-imidazole polyamide conjugates. AB - BACKGROUND: Pyrrole-imidazole polyamides are synthetic ligands that recognize predetermined sequences in the minor groove of DNA with affinities and specificities comparable to those of DNA-binding proteins. As a result of their DNA-binding properties, polyamides could deliver reactive moieties for covalent reaction at specific DNA sequences and thereby inhibit DNA-protein interactions. Site-specific alkylation of DNA could be a useful tool for regulating gene expression. As a minimal first step, we set out to design and synthesize a class of hairpin polyamides equipped with DNA alkylating agents and characterize the specificity and yield of covalent modification. RESULTS: Bis(dichloroethylamino)benzene derivatives of the well-characterized chlorambucil (CHL) were attached to the gamma turn of an eight-ring hairpin polyamide targeted to the HIV-1 promoter. We found that a hairpin polyamide-CHL conjugate binds and selectively alkylates predetermined sites in the HIV promoter at subnanomolar concentrations. Cleavage sites were determined on both strands of a restriction fragment containing the HIV-1 promoter, revealing good specificity and a high yield of alkylation. CONCLUSIONS: The ability of polyamide-CHL conjugates to sequence specifically alkylate double-stranded DNA in high yield and at low concentrations sets the stage for testing their use as regulators of gene expression in cell culture and ultimately in complex organisms. PMID- 10712932 TI - Inhibition of Escherichia coli porphobilinogen synthase using analogs of postulated intermediates. AB - BACKGROUND: Porphobilinogen synthase is the second enzyme involved in the biosynthesis of natural tetrapyrrolic compounds, and condenses two molecules of 5 aminolevulinic acid (ALA) through a nonsymmetrical pathway to form porphobilinogen. Each substrate is recognized individually at two different active site positions to be regioselectively introduced into the product. According to pulse-labeling experiments, the substrate forming the propionic acid sidechain of porphobilinogen is recognized first. Two different mechanisms for the first bond-forming step between the two substrates have been proposed. The first involves carbon-carbon bond formation (an aldol-type reaction) and the second carbon-nitrogen bond formation, leading to an iminium ion. RESULTS: With the help of kinetic studies, we determined the Michaelis constants for each substrate recognition site. These results explain the Michaelis-Menten behavior of substrate analog inhibitors - they act as competitive inhibitors. Under standard conditions, however, another set of inhibitors demonstrates uncompetitive, mixed, pure irreversible, slow-binding or even quasi-irreversible inhibition behavior. CONCLUSIONS: Analysis of the different classes of inhibition behavior allowed us to make a correlation between the type of inhibition and a specific site of interaction. Analyzing the inhibition behavior of analogs of postulated intermediates strongly suggests that carbon-nitrogen bond formation occurs first. PMID- 10712933 TI - Active-site variants of Streptomyces griseus protease B with peptide-ligation activity. AB - BACKGROUND: Peptide-ligating technologies facilitate a range of manipulations for the study of protein structure and function that are not possible using conventional genetic or mutagenic methods. To different extents, the currently available enzymatic and nonenzymatic methodologies are synthetically demanding, sequence-dependent and/or sensitive to denaturants. No single coupling method is universally applicable. Accordingly, new strategies for peptide ligation are sought. RESULTS: Site-specific variants (Ser195-->Gly, S195G, and Ser195-->Ala, S195A) of Streptomyces griseus protease B (SGPB) were generated that efficiently catalyze peptide ligation (i.e., aminolysis of ester-, thioester- and para nitroanilide-activated peptides). The variants also showed reduced hydrolytic activity relative to the wild-type enzyme. The ratio of aminolysis to hydrolysis was greater for the S195A variant, which was also capable of catalyzing ligation in concentrations of urea as high as 2 M. CONCLUSIONS: Mutagenic substitution of the active-site serine residue of SGPB by either glycine or alanine has created a unique class of peptide-ligating catalysts that are useful for coupling relatively stable ester- and para-nitroanilide-activated substrates. Ligation proceeds through an acyl-enzyme intermediate involving His57. Serine to alanine mutations may provide a general strategy for converting proteases with chymotrypsin-like protein folds into peptide-coupling enzymes. PMID- 10712934 TI - Catalytic asymmetric syntheses and biological activities of singly dehydroxylated 19-nor-1alpha,25-dihydroxyvitamin D(3) A-ring analogs in cancer cell differentiation and apoptosis. AB - BACKGROUND: 1alpha,25-Dihydroxyvitamin D(3) (1alpha,25(OH)(2)D(3)) has been shown to modulate not only proliferation and differentiation, but also apoptosis in malignant cells, indicating that it could be useful for treating cancer. Little information is available concerning the structural motifs of the 1alpha, 25(OH)(2)D(3) molecule responsible for modulation of differentiation and apoptosis, however. We set out to synthesize singly dehydroxylated A-ring analogs of 19-nor-1alpha,25(OH)(2)D(3) in a catalytic asymmetric fashion, and to investigate their biological activities in leukemia HL-60 cells. RESULTS: A series of singly dehydroxylated 19-nor-1alpha,25-dihydroxyvitamin D(3) A-ring analogs were synthesized using a combinatiorial sequence of regioselective propiolate-ene reaction and catalytic asymmetric carbonyl-ene cyclization. Surprisingly, the analogs could be clearly divided into two categories; one group, bearing 1alpha-hydroxy or 3beta-hydroxy groups in the A-ring, were potent differentiators and the second group, bearing 1beta-hydroxy or 3alpha-hydroxy groups, were potent stimulators of apoptosis. CONCLUSIONS: We have clearly identified the structural motifs of 19-nor-1alpha,25(OH)(2)D(3) analogs responsible for differentiation and apoptosis in HL-60 cells. These findings will provide useful information not only for development of therapeutic agents for treatment of leukemia and other cancers, but also for structure-function studies of 1alpha,25(OH)(2)D(3). PMID- 10712935 TI - DNA replication at high resolution. AB - Several decades of research have delineated the roles of many proteins central to DNA replication. Here we present a structural perspective of this work spanning the past 15 years and highlight several recent advances in the field. PMID- 10712936 TI - Tyrosine sulfation: a modulator of extracellular protein-protein interactions. AB - Tyrosine sulfation is a post-translational modification of many secreted and membrane-bound proteins. Its biological roles have been unclear. Recent work has implicated tyrosine sulfate as a determinant of protein-protein interactions involved in leukocyte adhesion, hemostasis and chemokine signaling. PMID- 10712937 TI - Genomic-scale analysis of gene expression in resting and activated T cells. AB - Recent advances in gene array technology and isolation of lymphocytes now allow comprehensive analysis of gene expression in many different types of T cells. So far only a few sets of results have been published. However it is already clear that these analyses provide accurate measurements of gene expression in T cells. This technology offers the first opportunity to examine global and subtle changes in gene expression in response to specific stimuli. PMID- 10712938 TI - The role of SLP-76 and LAT in lymphocyte development. AB - SLP-76 and LAT are two recently identified adapter proteins that are involved in the signal transduction cascade initiated by engagement of the TCR. The role of these two molecules in thymocyte development has become clearer following studies of gene targeted mice. The data indicate that SLP-76 and LAT are each critical for the expansion and differentiation of double-negative thymocytes and that SLP 76 is essential for allelic exclusion at the TCRbeta locus. PMID- 10712939 TI - Notch signaling in T cell development. AB - Notch signaling regulates cell fate decisions during development. Recent experiments suggest that Notch signaling is essential for initial commitment to the T cell lineage and may function together with signals from the pre-TCR and the TCR to regulate subsequent steps of T cell development. PMID- 10712940 TI - Self-antigen presentation by thymic stromal cells: a subtle division of labor. AB - Self-antigen-MHC complexes expressed by thymic stromal cells serve as ligands for TCR-mediated positive and negative selection, resulting in a self-MHC-restricted, self-tolerant T cell repertoire. It has recently become apparent that thymic stromal cells differ in their accessibility to antigen as well as their ability to process and present antigen. These differences result in the sampling by thymic stromal cells of largely nonoverlapping self-antigen pools and the display of self-peptide profiles specific for each cell type. In conjunction with single or serial cell-cell interactions between thymocytes and stromal cells, such differences in self-antigen display allow for maximal (re)presentation of 'self' in the thymus and optimize the efficacy of positive and negative selection. PMID- 10712941 TI - Unique and unprecedented recombination mechanisms in class switching. PMID- 10712942 TI - To be or not to be a pro-T? AB - Recent studies have begun to unravel some of the molecular pathways that appear to control the processes of T cell determination in the earliest thymocyte precursors. In addition, the analyses of mouse mutants with an entirely alymphoid thymus have shed light on the developmental relationship of pro-T cells and thymic dendritic cells, revealing that development of thymocytes and thymic dendritic cells can be dissociated. PMID- 10712943 TI - Regulation of RAG expression in developing lymphocytes. AB - Proper expression of products of the recombination-activating genes (RAGs) is essential for the development of the adaptive immune system. A major advance in the past year toward understanding RAG regulation is the establishment of green fluorescent protein (GFP)-RAG indicator mouse strains. In vivo visualization of RAG expression in single cells has helped to define the cells that express RAGs in secondary lymphoid organs and revealed differential cis requirements for stage and lineage-specific RAG expression. PMID- 10712944 TI - Lymphoid precursors. AB - Lymphopoiesis of mature and diverse populations of T, B and NK (natural killer) cells from multipotent hematopoietic stem cells is an ideal model of tissue generation and regeneration. Identification and isolation of hematolymphoid stem and progenitor cells in several laboratories over the past several years have provided populations that can be studied biologically for lineage commitment and biochemically for receptor function, signal transduction and selective gene expression. These studies may ultimately provide candidate genes involved in lineage commitment, cell death or survival, self-renewal and migratory capacities of progenitors. PMID- 10712945 TI - B-1 cells: orthodox or conformist? AB - B-1 cells differ from conventional peripheral B cells (B-2) by anatomical location, surface marker expression, antibody repertoire and growth properties. The lineage hypothesis of B-1 cell development attributes the properties of B-1 cells to their unique origin. The induced differentiation hypothesis suggests the surface-immunoglobulin-driven development of B-1 cells from common B-1/B-2 cell progenitors. In both models self-antigen-induced signalling plays the central role in positive selection of B-1 cells. The ability of B-1 cells to be positively selected by self-antigens raises questions about the mechanism of this phenomenon. PMID- 10712947 TI - Analysis of large-scale gene expression data. AB - The advent of cDNA and oligonucleotide microarray technologies has led to a paradigm shift in biological investigation, such that the bottleneck in research is shifting from data generation to data analysis. Hierarchical clustering, divisive clustering, self-organizing maps and k-means clustering have all been recently used to make sense of this mass of data. PMID- 10712946 TI - Lineage commitment in lymphopoiesis. AB - The mechanisms controlling the commitment of hematopoietic progenitor cells to the lymphoid lineages are still mostly unknown. Recent findings indicate that the earliest phase of B cell development may proceed in two steps. At the onset of B lymphopoiesis, the transcription factors E2A and EBF coordinately activate the B cell-specific gene expression program. Subsequently, Pax5 appears to repress the promiscuous transcription of lineage-inappropriate genes and thus commits progenitor cells to the B-lymphoid pathway by suppressing alternative cell fates. B-lineage commitment by Pax5 seems to occur in a stochastic manner in the bone marrow, as indicated by the random activation of only one of the two Pax5 alleles in early pro-B cells. In contrast, loss- and gain-of-function analyses have implicated the Notch1 receptor in the specification of the T cell fate, which may thus be controlled by instructive signals in the thymus. PMID- 10712948 TI - Genomic-scale gene expression analysis of lymphocyte growth, tolerance and malignancy. AB - Immunologists are already comfortable with the need for monitoring many different gene products simultaneously. It is a common challenge to remember what CD-one hundred-and-something is, and an ever-increasing number of colours are required for identification on the flow cytometer. Gene expression arrays now offer the possibility of extending this approach beyond the cell surface and expanding it dramatically to survey the entire catalogue of gene transcripts in a lymphoid cell. PMID- 10712949 TI - How the host 'sees' pathogens: global gene expression responses to infection. AB - Innate immune responses to pathogens are believed to be patterned and stereotyped. Adaptive responses display variety but in relatively few types of products and with limited numbers of mechanisms. Is this apparent disparity between microbial pathogen diversity and a restricted set of host responses an accurate picture of infection or is it the result of a limited collection of analytic tools? DNA microarray technology permits one to address simple descriptive questions about global gene expression inside cells. In particular, it offers an opportunity to examine the relationship between host and pathogen in much greater detail than has been possible previously. One can now ask, firstly, how a host cell or organism 'sees' a microbial pathogen from the viewpoint of gene expression responses and, secondly, at what level it is able to discriminate between different agents. Other potential insights to be reaped include the identification of microbial determinants of the host response, the temporal features of the 'conversation' between host and pathogen, novel strategies for therapeutic and prophylactic intervention and prognostic markers of outcome. PMID- 10712950 TI - Genomic-scale gene expression profiling of normal and malignant immune cells. AB - Gene expression variation is critical for the normal development and physiology of immune cells. Using cDNA microarrays, a systematic, genomic-scale view of gene expression in immune cells at many stages of differentiation and activation can be obtained. From the high vantagepoint provided by this technology, the gene expression physiology of immune cells appears remarkably ordered and logical. Each stage of lymphocyte differentiation can be defined by a characteristic gene expression signature. Genes that are co-regulated over hundreds of experimental conditions often encode functionally related proteins. Gene expression profiles also provide unprecedented ability to define the molecular and functional relationships between normal and malignant lymphocyte cell populations. PMID- 10712951 TI - International Rice Genome Sequencing Project: the effort to completely sequence the rice genome. AB - The International Rice Genome Sequencing Project (IRGSP) involves researchers from ten countries who are working to completely and accurately sequence the rice genome within a short period. Sequencing uses a map-based clone-by-clone shotgun strategy; shared bacterial artificial chromosome/P1-derived artificial chromosome libraries have been constructed from Oryza sativa ssp. japonica variety 'Nipponbare'. End-sequencing, fingerprinting and marker-aided PCR screening are being used to make sequence-ready contigs. Annotated sequences are immediately released for public use and are made available with supplemental information at each IRGSP member's website. The IRGSP works to promote the development of rice and cereal genomics in addition to producing genome sequence data. PMID- 10712952 TI - Synteny: recent advances and future prospects. AB - Their small sizes have meant that the Arabidopsis and rice genomes are the best studied of all plant genomes. Although even closely related plant species can show large variations in genome size, extensive genome colinearity has been established at the genetic level and recently also at the gene level. This allows the transfer of information and resources assembled for rice and Arabidopsis to be used in the genome analysis of many other plants. PMID- 10712953 TI - Chasing the dream: plant EST microarrays. AB - DNA microarray technology is poised to make an important contribution to the field of plant biology. Stimulated by recent funding programs, expressed sequence tag sequencing and microarray production either has begun or is being contemplated for most economically important plant species. Although the DNA microarray technology is still being refined, the basic methods are well established. The real challenges lie in data analysis and data management. To fully realize the value of this technology, centralized databases that are capable of storing microarray expression data and managing information from a variety of sources will be needed. These information resources are under development and will help usher in a new era in plant functional genomics. PMID- 10712954 TI - Genomic approaches to plant disease resistance. AB - Genomic approaches are beginning to revolutionize our understanding of plant disease resistance. Large-scale sequencing will reveal the detailed organization of resistance-gene clusters and the genetic mechanisms involved in generating new resistance specificities. Global functional analyses will elucidate the complex regulatory networks and the diversity of proteins involved in resistance and susceptibility. PMID- 10712955 TI - Saturation mutagenesis using maize transposons. AB - Transposon mutagenesis facilitates gene discovery by tagging genes for cloning. New genomics projects are now cataloging transposon insertion sites to define all maize genes. Once identified, transposon insertions are 'hot spots' for generating new alleles that are useful in functional studies. PMID- 10712956 TI - Genomic approaches to plant stress tolerance. AB - Past efforts to improve plant tolerance to drought, high salinity and low temperature through breeding and genetic engineering have had limited success owing to the genetic complexity of stress responses. Progress is now anticipated through comparative genomics studies of an evolutionarily diverse set of model organisms, and through the use of techniques such as high-throughput analysis of expressed sequence tags, large-scale parallel analysis of gene expression, targeted or random mutagenesis, and gain-of-function or mutant complementation. The discovery of novel genes, determination of their expression patterns in response to abiotic stress, and an improved understanding of their roles in stress adaptation (obtained by the use of functional genomics) will provide the basis of effective engineering strategies leading to greater stress tolerance. PMID- 10712957 TI - Chlamydomonas reinhardtii and photosynthesis: genetics to genomics. AB - Genetic and physiological features of the green alga Chlamydomonas reinhardtii have provided a useful model for elucidating the function, biogenesis and regulation of the photosynthetic apparatus. Combining these characteristics with newly developed molecular technologies for engineering Chlamydomonas and the promise of global analyses of nuclear and chloroplast gene expression will add a new perspective to views on photosynthetic function and regulation. PMID- 10712958 TI - Phytoremediation of toxic elemental and organic pollutants. AB - Phytoremediation is the use of plants to extract, sequester, and/or detoxify pollutants. Phytoremediation is widely viewed as the ecologically responsible alternative to the environmentally destructive physical remediation methods currently practiced. Plants have many endogenous genetic, biochemical, and physiological properties that make them ideal agents for soil and water remediation. Significant progress has been made in recent years in developing native or genetically modified plants for the remediation of environmental contaminants. Because elements are immutable, phytoremediation strategies for radionuclide and heavy metal pollutants focus on hyperaccumulation above-ground. In contrast, organic pollutants can potentially be completely mineralized by plants. PMID- 10712959 TI - Novel genes for disease-resistance breeding. AB - Plant disease control is entering an exciting period during which transgenic plants showing improved resistance to pathogenic viruses, bacteria, fungi and insects are being developed. This review summarizes the first successful attempts to engineer fungal resistance in crops, and highlights two promising approaches. Biotechnology provides the promise of new integrated disease management strategies that combine modern fungicides and transgenic crops to provide effective disease control for modern agriculture. PMID- 10712961 TI - [Extracorporeal circulation is the only technic that assures perfect and complete myocardial revascularization. The arguments pro]. PMID- 10712960 TI - Moral and ethical issues in plant biotechnology. AB - Plant biotechnology has recently become the focus of heated controversy and media attention, particularly in the UK. The most obvious concerns have centred upon the possible effects of the technology on the environment and on human health, but a broader and more fundamental set of considerations has also been evident in much of the debate, these are usually referred to as 'moral' or 'ethical' concerns. PMID- 10712962 TI - [Is coronary artery bypass grafting under extracorporeal circulation the only technic that assures complete myocardial revascularization? Pros and cons]. AB - Like other technical advances in medicine, the initial phase of rejection of off pump revascularization has given way to its acceptance with some limitations. Today's main concern is whether the technique can be safely applied to access all coronary arteries, specially the postero-lateral vessels of the heart. We believe that off-pump CABG has proven to be an excellent alternative to the standard revascularization techniques avoiding the risks and complications of cardiopulmonary bypass (CPB). It can be applied to any case with minimal incidence of conversions thus avoiding the complications of CPB. In this debate we describe our technique, discuss our experience with complete myocardial revascularization, and suggest that the CPB machine should be a readily available tool for more complex cases rather than the current concept that it is an indispensable element for myocardial revascularization. PMID- 10712963 TI - [Myocardial revascularization of the anterior descending artery with the classical technic of mammary anastomosis]. AB - INTRODUCTION AND OBJECTIVES: To establish the results obtained with the classical technique of mammary anastomosis of the anterior descending artery. MATERIALS AND METHODS: Between January 1982 and July 1997, 154 patients received an anastomosis of the left internal mammary artery to the left anterior descending coronary artery with use of the classical technique (sternotomy and extracorporeal circulation). RESULTS: There was no operative mortality in our group, nor paraoperational myocardial infarction, nor cephalic vascular accidents. One (0.6%) patient had sternal wound infection, and another (0.6%) required another reoperation for postoperational bleeding. All (100%) were followed-up from 3-183 months (average 64. 4 months). Actuarial global survival at 5, 10 and 15 years was 95.6%+/-2.1; 92.1%+/-4 and 85.5+/- 7.5 respectively, and the actuarial probability of being free from cardiac death was 99%+/-0.9; 99% and 99%. The actuarial probability of being free from myocardial infarction was 99%+/-0.9; 99%+/-0.9 and 99%; and from angina was 95%+/-2.2; 86.9%+/-4.9 and 74.5%+/-12.2 at 5, 10 and 15 years.Finally, the actuarial probability of being free from reoperation was 99%+/-0.9; 99%; 99% and from angioplasty 96.9%+/-1.7; 91.4%+/ 4.1; 91.4%+/-4.1 at 5, 10 and 15 years, respectively. The average hospital charges in the last 10% of the patients was U$ 6.200. CONCLUSIONS: Revascularization of the left anterior descending with the left internal mammary artery and the classical technique (sternotomy and extracorporeal circulation) is a safe, minimal risk, effective, long lasting and cost efficient procedure with excellent results at 10 and 15 years. PMID- 10712964 TI - [Refractory angina treated by spinal cord stimulation. The results of a long-term follow-up]. AB - INTRODUCTION AND OBJECTIVES: The quality of life in patients with refractory angina has been shown to improve dramatically with spinal cord stimulation because of its beneficial results. The aim of this study was to assess the long term morbimortality of this technique of spinal cord stimulation in the long term. PATIENTS AND METHODS: 41 patients with refractory angina and treated with spinal cord stimulation were included. Median follow-up was 31.0 [12.0-42.5] months, and total follow-up was 1,236 months. RESULTS: Annual number of admissions per patient year were dramatically reduced after spinal cord stimulation (2.31 vs. 0.28). Patients that died during follow-up had a 3-fold increase rate of admissions than patients that survived (0.37 vs. 0.19). However, patients that died during follow-up also had a lower admission rate after spinal cord stimulation (2.03 vs. 0.37). Overall mortality was 9.7%/year; cardiac mortality was 7.7%/year. Both figures are not different from those of other groups of patients with similar anatomical characteristics of coronary artery disease severity without spinal cord stimulation. Complications of this treatment were minimal (we only observed an early post implantation infection and a battery extrusion, without any complications with electrodes). The outcome was similar in patients with subacute unstable refractory angina or stable angina. CONCLUSIONS: Spinal cord stimulation can be considered a safe and effective alternative treatment of refractory angina. Long-term morbidity is low, and mortality is not higher than the expected in this group of patients. PMID- 10712965 TI - [Balloon pulmonary valvuloplasty in the neonatal period. The clinical and echocardiographic effects]. AB - AIM: To analyze the efficacy of balloon pulmonary valvuloplasty (BPV) as the elective treatment for neonatal critical pulmonary valvar stenosis (PVS). MATERIALS AND METHODS: The results of clinical and echocardiographic features before and after the BPV were reviewed in 29 neonates (18+/-12 days of life). Different hemodynamic and 2-D color Doppler echocardiographic were evaluated. The BPV result was classified as favourable if no other balloon or surgical therapy was required to normalise pulmonary flow and achieve a sustained right ventricle pulmonary artery (RV-PA) Doppler gradient below 40 mmHg. It was considered unfavourable if the neonate died, needed surgery or redilation and/or the RV-PA Doppler gradient was > or =40 mm Hg. The study developed in three phases: pre BPV immediate post BPV until the hospital discharge (14+/-11 days), and in the mid term follow-up of between 8 and 96 months (51+/-31 months). RESULTS: Mortality was not registered with BPV. The RV/left ventricular systolic pressure decreased from 1.4+/-0.3 to 0.8+/-0.3 (p<0.01) as a consequence of the dilation, and the the systemic oxygen saturation increased from 85 +/-12 to 92+/-6% (p<0.01). The RV-PA Doppler gradient diminished from 86+/-18 to 28+/-16 mm Hg immediately after BPV (p<0.01) and was registered at 13+/-6 mm Hg in the follow-up (n = 24). The RV PA junction Z value grew from -1.25+/-0.9 before valvuloplasty to -0.51 +/-0.7 at the final echocardiogram (p<0.01). No changes in the tricuspid diameter were detected between both periods of time. Five neonates obtained unsatisfactory results: 4 in the immediate post BPV (systemic-pulmonary artery shunt 2, transannular patch 2), and 1 in the mid-term follow-up (valvectomy + transannular patch). The actuarial curve reflects that 82,7% of the patients were free form reinterventions at 8 years. CONCLUSIONS: BPV is safe and effective to relief PVS in the neonate. The balloon promotes advantageous changes in both, pulmonary annulus and the right ventricle. In addition, the RV-PA Doppler gradient observations in the follow-up, support the expectation that the BPV is a "curative" therapy. PMID- 10712966 TI - [Biphasic low-energy internal cardioversion in atrial fibrillation induced during electrophysiologic study]. AB - INTRODUCTION: Atrial fibrillation is observed in 10% of electrophysiological studies. Previous studies have shown the efficacy of biphasic low energy internal cardioversion to restore sinus rhythm. We studied the efficacy and safety of low energy internal cardioversion and the biphasic curve, in sustained atrial fibrillation (>15 min) during electrophysiologic procedures. MATERIALS AND METHODS: From January 1997 to August 1998, 320 patients underwent an electrophysiological study. An internal cardioversion was done on those patients who developed sustained atrial fibrillation. We delivered biphasic shocks between electrodes-catheters positioned in the right atrium and the coronary sinus. Increasing energy was applied until restoration of sinus rhythm or a maximum of 10 joules were achieved with no result. A right ventricle electrode was used to synchronize the V wave and for temporary pacing. RESULTS: Thirty one episodes of sustained atrial fibrillation were observed in 26 patients (1,23 episodes/patient) and a mean of 2,58 internal cardioversions were applied per every patient. Sinus rhythm was restored in twenty three patients. The mean energy delivered was 4.1 joules. The mean time for the recovery was 3,200 ms. Temporary pacing was used in 16% of the patients for up to 1 minute. No AV blocks were observed. CONCLUSIONS: Internal cardioversion successfully restored sinus rhythm in 88.5% of the patients who presented sustained atrial fibrillation (88.5%). Temporary pacing was necessary for the treatment of postsinus shock pauses. PMID- 10712967 TI - [The general characteristics and short- and long-term results of infective endocarditis in non-drug addicts]. AB - INTRODUCTION AND OBJECTIVES: Infective endocarditis is a disease with a high morbimortality during the active phase and a considerable risk of complications during follow-up. The aim of our study is to describe the clinical and prognostic features of infective endocarditis in non-drug addict patients in short and long terms. PATIENTS AND METHODS: A prospective study of 138 cases of infective endocarditis in non-drug addict patients through the parenteral pathway treated in our institution from 1987 to 1997. RESULTS: The mean age was 44 +/- 20 years. Ninety-five patients (69%) had native valve infective endocarditis and forty three (31%) had prosthetic valve endocarditis. Streptococci were the causal microorganism in 34% and staphylococci in 33%. 83% of patients developed some type of complications during hospital stay. 51% of patients were operated on during the active phase (22% were urgent). The in-hospital mortality rate was 21%. 10 patients (9%) needed late cardiac surgery and seven patients (5%) died during follow-up. Global survival at 10 years was 71%. There were no statistical differences in survival in as much as the type of treatment received during the hospital stay in the active phase (medical alone or combined medical-surgical). CONCLUSIONS: A high early surgery rate in the active phase related to good long term results and does not increase early in-hospital mortality. Medical treatment also offers good long-term results in cases of infectious endocarditis with absence of bad prognostic factors and good clinical outcome. PMID- 10712968 TI - [Ischemic heart disease risk in the west Valladolid health area]. AB - INTRODUCTION AND OBJECTIVES: Cardiovascular diseases, especially ischemic heart disease present a high morbidity and mortality rate in our country. The aim of this study is to estimate the average coronary risk of people living in the west Valladolid Health District. METHODS: Cross-sectional study in a random sample of 369 people between 35 and 64 years of age from the general population, of systolic blood pressure, total serum cholesterol and cigarette smoking. With these three factors, plus sex and age, individual coronary risk was calculated through the Dundee Coronary Risk-Disk method. RESULTS: The coronary risk in the studied district, which is to say the individual probability of suffering a coronary event within five years of life, was estimated in 5.22 (95% CL: 4.75 5.69), higher among men, 5.66 (95% CL: 4.95-6.36), than among women, 4.63 (95% CL: 4.15-5.11). A descendent trend in coronary risk as age increases was found. CONCLUSIONS: This method is relatively easy to obtain for community studies and simple to use for individual risk. The coronary risk of a person from the studied population has similar levels to figures found in other studies from our settings. The community levels of isolated coronary risk factors do not permit the establishment the best option in coronary risk control, and only a multicausal approach will allow us to evaluate the most efficient interventions for each age group and sex. PMID- 10712969 TI - [The clinical practice guidelines of the Sociedad Espanola de Cardiologia on cardiomyopathies and myocarditis]. AB - Myocardial diseases are a extraordinarily heterogeneous group of processes that only have in common the fact that they involve heart muscle and that they cause a wide spectrum of myocardial dysfunction. The approach of the management and treatment of the cardiomyopathies is a continuous matter of discussion because the vast majority of alternatives in this field have not been based on the best scientific possible evidence and, since except for the case of heart failure associated with dilated cardiomyopathy. The majority of different options have not been studied by means of large (or even small) randomized trials. Nevertheless, this chapter has tried to provide the reader with different approaches on how to deal with important clinical problems in dilated, hypertrophic and restrictive cardiomyopathies, and in myocarditis as well. For this, we have utilized the most relevant information found coupled with our best clinical judgment, although we admit that many of the clinical recommendations can be controversial. PMID- 10712971 TI - [ [In Process Citation] PMID- 10712970 TI - [The clinical practice guidelines of the Sociedad Espanola de Cardiologia on pericardial pathology]. AB - The pericardium is a serous membrane consisting of two layers (parietal and visceral), which may be involved by different infectious, physical, traumatic, or inflammatory agents as well as in metabolic or systemic diseases. The reactions of the pericardium to these insults result in rather nonspecific clinical features, such as the characteristic inflammatory findings in acute pericarditis, the development of pericardial effusion with the possible complication of cardiac tamponade, and a fibrous retractile reaction that may lead to constrictive pericarditis. These phenomena are not mutually exclusive and can be simultaneous or consecutive in the same patient; however, for the sake of clarity they are independently discussed. The aim of the present guidelines is to provide orientation about the management of patients with pericardial disease. Such management should basically rest on the knowledge of the clinical and epidemiological features (such as disease frequency) of the different types of pericardial disease that determine the diagnostic and therapeutic yield of the different invasive pericardial procedures (pericardiocentesis, pericardial biopsy and pericardiectomy), and, therefore, their respective indications. In addition, the indication of the different types of medical therapy are discussed. On the other hand, emphasis is made on the possible limitation of the validity of these guidelines for patients belonging to geographical areas or socioeconomic contexts with different etiologic spectra. PMID- 10712972 TI - [The efficacy of platelet IIb/IIIa receptor blockers in acute coronary syndromes]. AB - The knowledge of the central role of platelets in the pathogenesis of acute coronary syndromes, on the one hand, and the fact that aspirin is a weak antiplatelet agent on the other, have led to an intensive investigational activity in antiplatelet drugs in the last years. Actually, the literature in the last two years is inundated with studies on the use of platelet IIb/IIIa receptor blockers in different clinical settings. Agents that block the IIb/IIIa platelet receptor have shown to be useful in improving prognosis of patients with acute coronary syndromes, especially in those undergoing percutaneous coronary revascularization procedures. However, their potential risk of bleeding and their high cost have prevented them from being applied universally and routinely. Furthermore there are still some unclear issues regarding the use of these drugs such as their correct dosage, the optimal duration of treatment and the direct comparison of the efficacy of different types of IIb/IIIa blockers available. On the other hand, oral IIb/IIIa antagonists have not improved the efficacy of aspirin to date and, moreover, they have been proven to be unsafe. Finally, it is necessary to identify those patients who will obtain the greatest benefit from the treatment in order to avoid the unnecessary risks and costs that would be derived from their universal use. PMID- 10712973 TI - [Predictors of sudden death in coronary artery disease]. AB - Although advances in the management of acute myocardial infarction have resulted in a decline in long-term risk of sudden death, it continues to be high in certain subsets of patients. Thus, it is important to identify and treat these patients. Left ventricular ejection fraction less than 0.40, frequent premature ventricular ectopy on Holter monitoring, late potentials on signal-averaged electrocardiogram, impaired heart rate variability, abnormal baroreflex sensitivity and inducible sustained monomorphic ventricular tachycardia during electrophysiological study are predictors of sudden death and arrhythmic events. Although the negative predictive value of each factor is high, the positive predictive accuracy is low. Several tests can be combined to obtain higher positive predictive values. In fact, in some studies combined noninvasive tests have been used to select patients for ventricular stimulation study. Some preventive treatment can be applied in these patients. Available data do not justify prophylactic therapy with amiodarone in high-risk survivors of acute myocardial infarction. Sudden death and total mortality have been significantly reduced in postinfarction patients by long-term beta blockade. Hence, beta blockers should be given to all patients with acute myocardial infarction who do not have contraindications to their use. The MADIT study has shown the beneficial effect of implantable cardioverter defibrillator in reducing mortality in patients with prior myocardial infarction, an ejection fraction less than 0.36, asymptomatic nonsustained ventricular tachycardia, and inducible sustained ventricular tachycardia, unsuppressable by procainamide. Besides, several studies are under way to evaluate the prophylactic use of implantable defibrillator for improving survival in high-risk patients. PMID- 10712974 TI - [Complete atrioventricular block and cardiac tumor in a newborn infant]. PMID- 10712975 TI - [Peripheral cholesterol embolism in a percutaneous coronary angioplasty procedure. A case report]. AB - Cholesterol embolism is a rare but potentially serious complication of cardiac catheterization. We report the case of a patient who presented a cholesterol embolism in the lower extremities after percutaneous angioplasty and the elective implantation of a stent. Clinical evolution was favourable. The appearance of cholesterol embolism could have been precipitated, in this case, by anticoagulation treatment with heparin and intense antiaggregation. PMID- 10712976 TI - [ST elevation and tension pneumothorax]. AB - We present a case of a sixty-nine-year-old male admitted to the hospital because of an acute respiratory failure that needed intubation and mechanical ventilation. Shortly after several attempts of right and left (the last one successful) subclavian vein cannulation (the last one successful) he developed a bilateral tension pneumothorax with important hemodynamic repercussion, a critical hypoxia and an ST elevation in inferior leads. Other more typical electrocardiographic changes could be observed: decrease in QRS amplitude and diminishing of precordial R voltage. After removing the air of the right pleural space, all the electrocardiographic signs disappeared returning to normal without electric or enzymatic assay of myocardial necrosis. PMID- 10712977 TI - [Severe digoxin poisoning. The successful use of the classic treatment]. AB - In our environment, the use of Fab antibodies for digoxin intoxication is often difficult due to the low availability of this drug in most centers. We present a case of massive digoxin intoxication that was successfully managed with the classic treatment. Later, we discuss the need to individualize the management of this kind of intoxications in order to reduce, when possible, sanitary costs. PMID- 10712978 TI - Back to basics: if it's dry, wet it : the case for humidification of nasal continuous positive airway pressure air. PMID- 10712979 TI - Why do girls use less oxygen during exercise than boys? Cause or effect of decreased work. PMID- 10712980 TI - Preventing multidrug-resistant tuberculosis and errors in tuberculosis treatment around the globe. PMID- 10712981 TI - Profiling drug resistance in immigrants with tuberculosis. PMID- 10712982 TI - Noninvasive positive pressure ventilation: testing the bridge. PMID- 10712983 TI - Toward a more thoughtful approach to fever in critically ill patients. PMID- 10712984 TI - Influence of cardiac functional capacity on gender differences in maximal oxygen uptake in children. AB - OBJECTIVE: To examine the role of gender differences in cardiac functional capacity in explaining higher mean values for maximal oxygen uptake (VO(2)max) in boys than in girls. DESIGN: Comparative group exercise testing. SETTING: Pediatric exercise testing laboratory. SUBJECTS: Twenty-five prepubertal boys (mean [+/- SD] age, 12 +/- 0.4 years) and 24 premenarcheal girls (mean age, 11.7 +/- 0.5 years). INTERVENTIONS: Maximal incremental upright cycle exercise. MEASUREMENTS AND RESULTS: Mean values for VO(2)max were the following: boys, 47.2 +/- 6.1 mL/kg/min; and girls, 40.4 +/- 5.8 mL/kg/min (16.8% difference; p < 0.05). The average maximal stroke index with Doppler echocardiography was 62 +/- 9 mL/m(2) for boys and 55 +/- 9 mL/m(2) for girls (12.7% difference; p < 0.05). No significant gender differences were seen in maximal heart rate or arterial venous oxygen difference. When VO(2)max and maximal stroke volume (SV) were expressed relative to lean body mass, gender differences declined but persisted, falling to 6.2% and 5.2%, respectively. CONCLUSIONS: These findings indicate that differences in SV as well as in body composition contribute to gender-related variations in VO(2)max during childhood. Whether this reflects small gender differences in relative heart size or dynamic factors influencing ventricular preload and contractility during exercise is unknown. PMID- 10712985 TI - Is mitral valve prolapse due to cardiac entrapment in the chest Cavity? A CT view. AB - BACKGROUND: Mitral valve prolapse (MVP) is the most frequently diagnosed valvular disease, but its pathophysiology remains elusive. Its complete absence in 1,734 neonatal echocardiographic studies suggests that this may be an acquired rather than a congenital disease. We observed several patients with distorted cardiac and valvular anatomies on electron beam CT (EBCT) images of the chest who reported symptoms reminiscent of MVP. In these patients, the heart is compressed between the spine and the anterior chest wall and it appears trapped in a chest cavity that is too small for its size. METHODS: We performed EBCT in 66 patients with echocardiographically proven MVP and no clinical pectus excavatum (group A; 80% were women; mean age, 48 +/- 12 years) and in 96 control patients without MVP by echocardiography (group B; 72% were women; mean age, 49 +/- 10 years). EBCT alone was also performed on 200 patients who had reported atypical chest discomfort and palpitations to their physicians (group C) and on 200 asymptomatic patients (group D). The EBCT measurements included the following: anteroposterior chest diameter (APD); the angle formed by the confluence of the mitral valve ring with the interatrial septum (ANGLE); and the contact area between the posterior surface of the anterior chest wall and the myocardium (CA). Entrapment was considered present if the individual patient's measurements varied by more than two SDs compared to measurements made in control subjects (group B). RESULTS: EBCT images demonstrated cardiac entrapment in 82% of group A patients and in 4.2% of group B patients (p < 0.001). ANGLE and CA were significantly larger in MVP patients than in group B patients (114 +/- 9 degrees vs 91 +/- 5 degrees and 6,230 +/- 2,020 mm(2) vs 476 +/- 1,009 mm(2), respectively; p < 0.001 for both comparisons), while APD was significantly smaller (91 +/- 16 mm vs 128 +/- 17 mm, respectively; p < 0.001). The prevalence of entrapment was significantly greater in group C patients than in group D patients (22% vs 6.5%; p < 0. 001). CONCLUSIONS: MVP may be an acquired condition caused by a growth disproportion between the heart and the chest cavity, with distortion of the mitral valve annulus and subsequent leaflet prolapse. A narrow APD, a wide ANGLE, and a large CA characterize this condition. Similar findings are found in a sizable proportion of patients with atypical chest pain symptoms and palpitations. PMID- 10712986 TI - Partial improvement in pulmonary function after successful percutaneous balloon mitral valvotomy. AB - STUDY OBJECTIVES: This study was performed to assess the changes in pulmonary function after a successful percutaneous balloon mitral valvotomy (PBMV) in 23 consecutive patients with symptomatic mitral stenosis. METHODS AND RESULTS: Lung function preprocedure and postprocedure were evaluated by spirometric flow, static pulmonary volumes, and diffusion capacity of the lung for carbon monoxide (DLCO). At baseline, a reduction in small airways flow (maximal expiratory flow at 50% of vital capacity, 70 +/- 29% of predicted value; maximal expiratory flow at 25% of vital capacity, 55 +/- 26% of predicted value) and an increase in DLCO (118 +/- 29%) and Krough Index (KCO; 123 +/- 29% of predicted value) were observed. PBMV caused an improvement in hemodynamic parameters with an increase in mitral valve area (from 1.0 +/- 0.3 to 1.9 +/- 0.5 cm(2); p < 0.001) and a decrease in left atrial pressure (from 17 +/- 3 to 12 +/- 5 mm Hg; p < 0.001). These changes were associated with a significant increase in FVC (from 2.8 +/- 0.84 to 2.9 +/- 0.80 L; p < 0.05) and in FEV(1) (from 2.2 +/- 0.72 to 2.3 +/- 0.68 L; p < 0.05). A decrease in DLCO was observed after PBMV (from 26.7 +/- 7 to 22.5 +/- 5.4 mL/min/mm Hg; p < 0.001; and KCO, from 6.2 +/- 1.4 to 5.2 +/- 1.2 mL/min/mm Hg/L; p < 0.001). No significant changes in small airways flow were detected, suggesting only a partial improvement in pulmonary congestion. CONCLUSION: We conclude that the initial impairment of lung function in patients with symptomatic mitral stenosis is only partially ameliorated by PBMV. PMID- 10712987 TI - Safety, hemodynamic profile, and feasibility of dobutamine stress technetium myocardial perfusion single-photon emission CT imaging for evaluation of coronary artery disease in the elderly. AB - OBJECTIVES: Cardiovascular disease is the leading cause of morbidity and mortality in the elderly. The evaluation of coronary artery disease by exercise stress testing is frequently limited by the patient's inability to exercise. Although pharmacologic stress testing with dobutamine is an alternative, the safety of dobutamine myocardial perfusion scintigraphy in the elderly has not been previously studied. PATIENTS AND METHODS: We studied the safety and feasibility of dobutamine (up to 40 microg/kg/min)-atropine (up to 1 mg) stress myocardial perfusion scintigraphy using technetium single-photon emission CT imaging in 227 patients > or = 70 years old (mean +/- SD age, 75 +/- 4 years). A control group of 227 patients < 70 years old (mean age, 55 +/- 11 years; matched for gender, prevalence of previous infarction, beta-blocker therapy, and severity of resting perfusion abnormalities) was studied to assess age-related differences in the safety and the hemodynamic response. A feasible test was defined as the achievement of the target heart rate and/or an ischemic end point (angina, ST segment depression, or reversible perfusion abnormalities). RESULTS: No myocardial infarction or death occurred during the test. The target heart rate was achieved more frequently in the elderly patients (87% vs 79%; p < 0.05). The elderly patients had a higher prevalence of supraventricular tachycardia (7% vs 1%; p < 0.005) and premature ventricular contraction (74% vs 32%; p < 0.005) during the test, as compared to the younger patients. There was a trend to a higher prevalence of ventricular tachycardia (5% vs 2%) and atrial fibrillation (3% vs 0.4%) in the elderly patients. Arrhythmias were terminated spontaneously by termination of dobutamine infusion or by administration of metoprolol. Independent predictors of supraventricular tachyarrhythmias and ventricular tachycardia were older age (p < 0.001; chi(2), 9.8) and myocardial perfusion defect score at rest (p < 0.01; chi(2), 6.8) respectively, by using a multivariate analysis of clinical and stress test variables. Elderly patients had a higher prevalence of systolic BP drop > 20 mm Hg during the test (37% vs 12%; p < 0.05). The test was terminated due to hypotension in 2% of the elderly patients and in 1% of the control group. Age was the most powerful predictor of hypotension (p < 0.005; chi(2), 10.3). The test was considered feasible in 216 elderly patients (95%) and in 209 patients of the control group (92%). CONCLUSION: Dobutamine-atropine stress myocardial perfusion scintigraphy is a highly feasible method for the evaluation of coronary artery disease in the elderly. Elderly patients have a higher risk for developing hypotension and supraventricular tachyarrhythmias during a dobutamine stress test. However, dobutamine-induced hypotension is often asymptomatic and rarely necessitates the termination of the test. PMID- 10712988 TI - Transesophageal dobutamine stress echocardiography in the evaluation of myocardial ischemia in morbidly obese subjects. AB - BACKGROUND: The evaluation of chest pain or suspected coronary artery disease (CAD) in morbidly obese subjects is limited by the inability of routine diagnostic techniques to adequately image these individuals. Morbidly obese subjects are therefore often inadequately treated or inappropriately treated for presumed CAD. METHODS AND RESULTS: We prospectively evaluated 23 morbidly obese patients with chest pain using transesophageal dobutamine stress echocardiography (TE-DSE). The mean (+/- SD) weight was 164 +/- 8 kg (range, 118 to 215 kg). We identified nine patients with abnormal TE-DSE findings. Five of these patients subsequently had cardiac catheterization with confirmation of CAD in the regions identified by TE-DSE. Over a follow-up period of 18 +/- 6 months, three cardiac events (non-Q-wave myocardial infarction) occurred in the same group, including two patients without confirmatory cardiac catheterization data. Thus, seven of nine patients with positive results of TE-DSE had objective confirmatory evidence of CAD. No cardiac events were observed in the group with normal TE-DSE over the same follow-up period. CONCLUSION: TE-DSE is a safe and potentially useful technique for the evaluation of suspected CAD in morbidly obese subjects. PMID- 10712989 TI - Acute exacerbation of COPD: factors associated with poor treatment outcome. AB - OBJECTIVES: To determine the effect of age, severity of lung disease, severity and frequency of exacerbation, steroid use, choice of an antibiotic, and the presence of comorbidity on the outcome of treatment for an acute exacerbation of COPD. DESIGN: A retrospective chart analysis over 24 months. SETTING: A university Veterans Affairs medical center. PATIENTS: Outpatients with COPD who were treated with an antibiotic over a period of 24 months for an acute exacerbation of COPD. METHODS: Severity of an acute exacerbation of COPD was defined using the criteria of Anthonisen et al: increased dyspnea, increased sputum volume, and increased sputum purulence. Severity of lung disease was stratified based on FEV(1) percent predicted using American Thoracic Society guidelines (stage I, FEV(1) > or = 50%; stage II, FEV(1) 35 to 49%; stage III, FEV(1) < 35%). Treatment outcome was judged successful when the patient had no return visit in 4 weeks for a respiratory problem. Failure was defined as a return visit for persistent respiratory symptoms that required a change of an antibiotic in < 4 weeks. RESULTS: One-hundred seven patients with COPD (mean age +/- SD, 66.9 +/- 9.5 years) experienced 232 exacerbations over 24 months. First line antibiotics (trimethoprim-sulfamethoxazole, ampicillin/amoxicillin, and erythromycin) were used to treat 78% of all exacerbations. Treatment failure was noted in 12.1% of first exacerbations and 14. 7% of all exacerbations, with more than half the failures requiring hospitalization. Host factors that were independently associated with treatment failure included the following: FEV(1) < 35% (46.4% vs 22.4%; p = 0.047), use of home oxygen (60.7% vs 15.6%; p < 0. 0001), frequency of exacerbation (3.8 +/- 2.0 vs 1.6 +/- 0.91; p < 0. 001), history of previous pneumonia (64.3% vs 35.1 p < 0.007), history of sinusitis (28.6% vs 8.8%; p < 0.009) and use of maintenance steroids (32.1% vs 15.2% p = 0.052). Using stepwise logistic regression analysis to identify the top independent variables, the use of home oxygen (p = 0.0002) and frequency of exacerbation (p < 0.0001) correctly classified failures in 83.3% of the patients. Surprisingly, age, the choice of an antibiotic, and the presence of any one or more comorbidity did not affect the treatment outcome. CONCLUSION: The results of our study suggest that patient host factors and not antibiotic choice may determine treatment outcome. Prospective studies in appropriately stratified patients are needed to validate these findings. PMID- 10712990 TI - Nutritional support for individuals with COPD: a meta-analysis. AB - RATIONALE: Malnutrition in patients with COPD is associated with an impaired pulmonary status, reduced diaphragmatic mass, lower exercise capacity, and higher mortality rate when compared to adequately nourished individuals with COPD. Nutritional support may therefore be a useful part of their comprehensive care. PURPOSE: To conduct a meta-analysis of randomized controlled trials (RCTs) to clarify whether nutritional supplementation (caloric supplementation for at least 2 weeks) improved anthropometric measures, pulmonary function, respiratory muscle strength, and functional exercise capacity in patients with stable COPD. METHODS: RCTs were identified from several sources, including the Cochrane Airways Group register of RCTs, a hand search of abstracts presented at international meetings, and consultation with experts. Two reviewers independently selected trials for inclusion, assessed quality, and extracted the data. Within each trial and for each outcome, we calculated an effect size. The effect sizes were then pooled by a random-effects model. Homogeneity among the effect sizes was also tested. RESULTS: From 272 references, nine RCTs were ultimately included. Six articles were considered as high quality. Only two studies were double blinded. For each of the outcomes studied, the effect of nutritional support was small: the 95% confidence intervals around the pooled effect sizes all included zero. The effect of nutritional support was homogeneous across studies. CONCLUSION: Nutritional support had no effect on improving anthropometric measures, lung function, or functional exercise capacity among patients with stable COPD. PMID- 10712991 TI - Incidence of nocturnal desaturation while breathing oxygen in COPD patients undergoing long-term oxygen therapy. AB - STUDY OBJECTIVE: It is suggested that oxygen flow be increased by 1 L/min during sleep in COPD patients undergoing long-term oxygen therapy (LTOT) in order to avoid nocturnal desaturations. The purpose of this study was to investigate the occurrence of nocturnal desaturations while breathing oxygen in COPD patients receiving LTOT. SETTING: Inpatient/university hospital. PATIENTS: We studied 82 consecutive COPD patients. Their functional characteristics were as follows (mean +/- SD): FVC, 2.15 +/- 0.69 L; FEV(1), 0.87 +/- 0.33 L; PaO(2), 51.6 +/- 5 mm Hg; and PaCO(2), 47 +/- 8 mm Hg. MEASUREMENTS: Overnight pulse oximetry (PO) was performed twice: (1) while breathing air and (2) while breathing supplemental oxygen assuring satisfactory diurnal resting oxygenation (mean PaO(2) during oxygen breathing, 67 +/- 6 mm Hg; mean arterial oxygen saturation [SaO(2)] during oxygen breathing, 93%). RESULTS: PO performed while patients were breathing air showed a mean overnight SaO(2) of 82.7 +/- 6.7%. Patients spent 90% of the recording time with an SaO(2) of < 90%. While breathing oxygen, 43 patients (52.4%) remained well oxygenated. Their mean overnight SaO(2) while breathing oxygen was 94.4 +/- 2.1%, and time spent with saturation < 90% was 6.9 +/- 8.6%. Thirty-nine patients (47.6%) spent > 30% of the night with an SaO(2) of < 90% while breathing supplemental oxygen. Their mean overnight SaO(2) while breathing oxygen was 87.1 +/- 4.5%, and time spent with an SaO(2) of < 90% was 66.1 +/- 24.7% of the recording time. Comparison of ventilatory variables and daytime blood gases between both groups revealed statistically significantly higher PaCO(2) on air (p < 0.001) and on oxygen (p < 0. 05), and lower PaO(2) on oxygen (p < 0.05) in the group of patients demonstrating significant nocturnal desaturation. CONCLUSIONS: We conclude that about half of COPD patients undergoing LTOT need increased oxygen flow during sleep. Patients with both hypercapnia (PaCO(2) > or = 45 mm Hg) and PaO(2) < 65 mm Hg while breathing oxygen are most likely to desaturate during sleep. PMID- 10712992 TI - Upregulation of gelatinases A and B, collagenases 1 and 2, and increased parenchymal cell death in COPD. AB - BACKGROUND: A central feature in the pathogenesis of COPD is the inflammation coexisting with an abnormal protease/antiprotease balance. However, the possible role of different serine and metalloproteinases remains controversial. PATIENTS AND MEASUREMENTS: We examined the expression of gelatinases A and B (matrix metalloproteinase [MMP]-2 and MMP-9); collagenases 1, 2, and 3 (MMP-1, MMP-8, and MMP-13); as well as the presence of apoptosis in lung tissues of 10 COPD patients and 5 control subjects. In addition, gelatinase-A and gelatinase-B activities were assessed in BAL obtained from eight COPD patients, and from six healthy nonsmokers and six healthy smoker control subjects. SETTING: Tertiary referral center and university laboratories of biochemistry, and lung cell kinetics. RESULTS: Immunohistochemical analysis of COPD lungs showed a markedly increased expression of collagenases 1 and 2, and gelatinases A and B, while collagenase 3 was not found. Neutrophils exhibited a positive signal for collagenase 2 and gelatinase B, whereas collagenase 1 and gelatinase A were revealed mainly in macrophages and epithelial cells. BAL gelatin zymography showed a moderate increase of progelatinase-A activity and intense bands corresponding to progelatinase B. In situ end labeling of fragmented DNA displayed foci of positive endothelial cells, although some alveolar epithelial, interstitial, and inflammatory cells also revealed intranuclear staining. CONCLUSION: These findings suggest that there is an upregulation of collagenase 1 and 2 and gelatinases A and B, and an increase in endothelial and epithelial cell death, which may contribute to the pathogenesis of COPD through the remodeling of airways and alveolar structures. PMID- 10712993 TI - Increased exhaled nitric oxide in chronic bronchitis: comparison with asthma and COPD. AB - STUDY OBJECTIVES: To test the hypothesis that exhaled nitric oxide (NO) is increased in patients with chronic bronchitis, and to compare the results with exhaled NO in patients with asthma and COPD. STUDY DESIGN: Cross-sectional survey. SETTING AND PATIENTS: Veterans Administration pulmonary function laboratory. Patients (n = 179) were recruited from 234 consecutive patients. Two nonsmoking control groups of similar age, with normal spirometry measurements and no lung disease, were used (18 patient control subjects and 20 volunteers). MEASUREMENTS: Participants completed questionnaires and spirometry testing. Exhaled NO was measured by chemiluminescence using a single-breath exhalation technique. RESULTS: Current smoking status was associated with reduced levels of exhaled NO (smokers, 9. 2 +/- 0.9 parts per billion [ppb]; never and ex-smokers, 14.3 +/- 0. 6 ppb; p < 0.0001). Current smokers (n = 57) were excluded from further analysis. Among nonsmokers, the levels of exhaled NO were significantly higher in patients with chronic bronchitis (17.0 +/- 1. 1 ppb; p = 0.035) and asthma (16.4 +/- 1.3 ppb; p = 0.05) but not in those with COPD (14.7 +/- 1.0 ppb; p = 0.17) when compared with either control group (patient control subjects, 11.1 +/- 1.6 ppb; outside control subjects, 11.5 +/- 1.5 ppb). The highest mean exhaled NO concentration occurred in patients with both chronic bronchitis and asthma (20.2 +/- 1.6 ppb; p = 0.005 vs control subjects). CONCLUSIONS: Exhaled NO is increased in patients with chronic bronchitis. The increase of exhaled NO in patients with chronic bronchitis was similar to that seen in patients with asthma. The highest mean exhaled NO occurred in patients with both chronic bronchitis and asthma. Exhaled NO was not increased in patients with COPD. Although chronic bronchitis and asthma have distinct histopathologic features, increased exhaled NO in patients with both diseases suggests common features of inflammation. PMID- 10712994 TI - Exhaled nitric oxide and exercise in stable COPD patients. AB - STUDY OBJECTIVE: To evaluate exhaled nitric oxide (eNO) during exercise in patients with stable COPD. SETTING: Outpatient evaluation in a rehabilitation center. PATIENTS: Eleven consecutive male patients with stable COPD (age, 65 +/- 6 years; FEV(1), 56 +/- 10% predicted). Eight healthy (six men; age, 51 +/- 16 years) nonsmoking, nonatopic volunteers served as control subjects. METHODS: In each subject, a symptom-limited cycle ergometry test was performed by monitoring eNO with the tidal-breath method to assess eNO concentration (FENO) and output (VNO) at rest, peak exercise, and recovery time. RESULTS: Resting FENO (9.8 +/- 5.1 and 14.1 +/- 6.3 parts per billion, respectively) and VNO (4.2 +/- 2.0 and 5.9 +/- 3.4 nmol/min, respectively) were lower, although not significantly, in COPD patients than in control subjects. In both groups, FENO significantly decreased whereas VNO significantly increased during exercise. Both variables returned to baseline during the recovery time. Peak exercise VNO, but not FENO, was significantly lower in COPD patients than in control subjects (7.9 +/- 5.4 and 12.7 +/- 6.0 nmol/min, respectively, p < 0.05). The rise in VNO was weakly correlated to oxygen consumption VO(2)) both in control subjects (r = 0.31, p = 0. 002) and in COPD patients (r = 0.22, p = 0.03). FENO showed an inverse correlation to VO(2) in both groups (r = -0.53, p = 0.000; r = -0.31, p = 0.003 in control subjects and COPD patients, respectively). CONCLUSIONS: In patients with mild and moderate COPD, eNO during exercise parallels that observed in normal control subjects. VNO, but not FENO, is significantly reduced at peak exercise in COPD patients as compared with control subjects. The long-term effects of exercise training on eNO has to be evaluated by further studies. PMID- 10712995 TI - Churg-Strauss syndrome in patients receiving montelukast as treatment for asthma. AB - STUDY OBJECTIVES: We previously reported eight patients who developed Churg Strauss syndrome in association with zafirlukast treatment for asthma and postulated that the syndrome resulted from unmasking of a previously existing condition due to corticosteroid withdrawal and not from a direct drug effect. The availability of montelukast, a new leukotriene receptor antagonist with a different molecular structure, permitted us to test this hypothesis. Our goals were to ascertain whether the Churg-Strauss syndrome developed in patients taking montelukast and other novel asthma medications, and to describe potential mechanisms for the syndrome. DESIGN: Case series. SETTING: Outpatient and hospital practices of pulmonologists in the United States and Belgium. PATIENTS: Four adults (one man, three women) who received montelukast as treatment for asthma; two women who received salmeterol/fluticasone therapy, but not montelukast. RESULTS: Churg-Strauss syndrome developed in the four asthmatic patients who received montelukast. In each case, there was a long history of difficult-to-control asthma characterized by multiple exacerbations that had required frequent courses of oral systemic corticosteroids or high doses of inhaled corticosteroids for control. Two other asthmatics who received fluticasone and salmeterol but not montelukast therapy developed the same syndrome with tapering doses of oral or high doses of inhaled corticosteroids. CONCLUSIONS: The occurrence of Churg-Strauss syndrome in asthmatic patients receiving leukotriene modifiers appears to be related to unmasking of an underlying vasculitic syndrome that is initially clinically recognized as moderate to severe asthma and treated with corticosteroids. Montelukast does not appear to directly cause the syndrome in these patients. PMID- 10712996 TI - Demonstration of in vivo bioequivalence of a generic albuterol metered-dose inhaler to Ventolin. AB - STUDY OBJECTIVE: To use histamine bronchoprovocation and bioassay statistical procedures to evaluate the in vivo bioequivalence of a generic albuterol metered dose inhaler (MDI). DESIGN: A randomized, double-blind, balanced, crossover design was used to determine the potency of each generic albuterol MDI actuation relative to Ventolin (Glaxo Wellcome; Research Triangle Park, NC) administration. One treatment was administered on each of 4 study days. A histamine bronchoprovocation procedure was initiated 1.25 h before and 15 min after administration of the study treatment. PATIENTS: Twenty-four nonsmoking subjects with mild-to-moderate asthma were studied (18 to 65 years of age; FEV(1), > 60% of predicted; and provocative concentration of histamine causing a 20% fall in FEV(1) [PC(20)], < or = 8 mg/mL at screening). INTERVENTIONS: One and four actuations (90 and 360 microg, respectively) of the generic MDI and of Ventolin MDI. Placebo inhalers were used to maintain blinding of inhaler and doses. MEASUREMENTS AND RESULTS: The primary outcome variable was histamine PC(20) measured after study treatment administration. A significant dose-effect relationship was present (p < 0.0001). Deviation from parallelism of the generic and Ventolin dose-response curves (p = 0.95) and differences in overall mean response between the two formulations (p = 0.68) were not significant. Using Finney 2 x 2 bioassay statistical procedures, we estimated that one actuation of the generic albuterol MDI was equivalent to 1.01 puffs of Ventolin (90% confidence interval, 0.69 to 1.50). CONCLUSION: The generic albuterol MDI delivers a quantity of albuterol to the beta(2)-receptor site in the lung that is the bioequivalent to Ventolin. Further, this study reinforces the validity of this statistical methodology for determining in vivo bioequivalence. PMID- 10712997 TI - Specific inspiratory muscle training in patients with mild asthma with high consumption of inhaled beta(2)-agonists. AB - BACKGROUND: It has been known for many years that there are variations between asthmatic patients in terms of their perception of breathlessness during airway obstruction. STUDY OBJECTIVE: To investigate the relationship between beta(2) agonist consumption and the score of perception of dyspnea, in mild asthmatics, and the relationship between the effect of specific inspiratory muscle training (SIMT) on the score of perception of dyspnea and beta(2)-agonist consumption in "high perceivers." METHODS: Daily beta(2)-agonist consumption was assessed during a 4-week run-in period in 82 patients with mild asthma. Patients with a mean beta(2)-agonist consumption of > 1 puff/d ("high consumers") then were randomized into two groups: one group of patients received SIMT for 3 months; the other group of patients was assigned as a control group and received sham training. Inspiratory muscle strength and perception of dyspnea were assessed before patients entered the study, following the 4-week run-in period, and after completing the training period. RESULTS: Following the 4-week run-in period, 23 high-consumer patients (mean [+/- SEM] beta(2)-agonist consumption, 2.7 +/- 0.4 puffs/d) were detected. The mean Borg score during breathing against resistance was significantly higher (p < 0.05) in the patients with high beta(2)-agonist consumption than in the subjects with low mean beta(2)-agonist consumption. Following SIMT, the mean maximal inspiratory pressure increased significantly from 94.1 +/- 5.1 to 109.7 +/- 5.2 cm H(2)O (p < 0.005) in the training group. The increase in inspiratory muscle strength was associated with a statistically significant decrease in the mean Borg score during breathing against resistance (p < 0.05) as well as in the mean daily beta(2)-agonist consumption. CONCLUSIONS: We have shown that patients with mild asthma, who have a high beta(2)-agonist consumption, have a higher perception of dyspnea than those with normal consumption. In addition, SIMT was associated with a decrease in perception of dyspnea and a decrease in beta(2)-agonist consumption. PMID- 10712998 TI - Uncontrolled oxygen administration and respiratory failure in acute asthma. AB - STUDY OBJECTIVES: To determine if 100% oxygen administration adversely influences gas exchange in acutely ill asthmatic subjects. DESIGN: Prospective preinterventional and postinterventional comparison. SETTING: University hospital emergency department. PATIENTS: Thirty-seven asthmatic subjects seeking care for symptomatic exacerbations. INTERVENTIONS: Twenty minutes of 100% oxygen administration by face mask. MEASUREMENTS AND RESULTS: Arterial blood gases and FEV(1) were measured before and during the last minute of oxygen administration. On presentation, the subjects had moderately severe airway obstruction (FEV(1), 49.1 +/- 3.6% of predicted); hypocarbia (PaCO(2), 36.8 +/- 1.1 mm Hg); hypoxemia (PaO(2), 70.2 +/- 2.5 mm Hg); and respiratory alkalosis (pH, 7.43 +/- 0.01). During oxygen breathing, 25 patients (67.6%) experienced elevations in PaCO(2) ranging from 1 to 10 mm Hg (mean, 4.1 +/- 0.6 mm Hg; p = 0.0003). The increase was considered to be a physiologic manifestation of the Haldane effect (ie, < or = 2 mm Hg) in 10 subjects, but in the remaining 15 subjects (40.5% of the total studied), the elevation represented worsening gas exchange. In seven of these patients (46.7%), hypercapnic respiratory failure developed (PaCO(2) before oxygen, 39.6 +/- 0.6; during oxygen, 44.7 +/- 0.7 mm Hg; p = 0.005), and in six patients (40%), it worsened (PaCO(2) before oxygen, 46.8 +/- 1.9; during oxygen, 52.0 +/- 3.1 mm Hg; p = 0.03). In general, the tendency toward hypercarbia was the greatest in the participants with the most severe airway obstructions. CONCLUSIONS: Our data demonstrate that the administration of 100% oxygen to acutely ill asthmatics may adversely influence carbon dioxide elimination. PMID- 10712999 TI - Errors in the treatment of tuberculosis in Baltimore. AB - BACKGROUND: Incomplete or incorrect antibiotic therapy, especially in the initial phase of antituberculosis (anti-TB) treatment, is a major cause of acquired drug resistance and treatment failure. We determined the extent of errors in anti-TB treatment regimens by way of nonadherence to recommended treatment protocols among patients with TB in Baltimore, MD, a city with declining rates of disease. An error was defined as using too few drugs or the wrong drugs, giving inadequate doses of drugs, or prescribing an inadequate duration of treatment. METHODS: We reviewed the records of all patients with culture-positive, pulmonary TB reported in the city of Baltimore from January 1, 1994, to December 31, 1995. We determined demographic information, initial anti-TB regimen, doses and duration of therapy, history or presence of resistance to anti-TB drugs, injecting-drug or alcohol abuse, HIV status, and whether treatment was given by a private physician or by the Tuberculosis Clinic of the Baltimore City Health Department (BCHD). RESULTS: Of the 110 cases of active pulmonary TB, 17 cases (15.4%) had errors in treatment for control of their current disease. Thirteen of 34 privately treated patients (38%) had some error in their initial anti-TB regimen, compared with 4 of 76 patients (5.2%) treated by the Tuberculosis Clinic of the BCHD (p < 0.0001). Patients were otherwise similar as determined by age, sex, HIV status, drug-resistance characteristics, and injecting-drug use, regardless of whether they had erroneous anti-TB regimens. CONCLUSION: In a low-prevalence area, private physicians make frequent errors in prescribing anti-TB therapy. Additional educational resources for physicians and increased use of expert consultation may contribute to improved TB control. PMID- 10713000 TI - Epidemiology and ethnic distribution of multidrug-resistant tuberculosis in southern Israel, 1992-1997: the impact of immigration. AB - STUDY OBJECTIVES: To assess the incidence of tuberculosis in the native and immigrant populations of southern Israel in the period between 1992 and 1997, and to study the prevalence of drug resistance overall and among these subpopulations in the region in order to create guidelines for empirical antituberculous treatment in this region. DESIGN: A retrospective population-based study. SETTING: The southern district of the country and its tertiary-care hospital. PATIENTS: All new culture-proven tuberculosis cases diagnosed in adults residing in the Negev region during the study period. Patients were classified into four groups according to ethnic origin and immigration date. RESULTS: During the study period, 249 new cases involving 249 patients were recorded. Immigrants from the former Soviet Union (IFSU) were significantly younger and of male gender, and the incidence among this group rose sharply. IFSU had higher rates of resistance to any drug or drug combination. Isoniazid resistance rates were 16% overall and 32% among IFSU. Resistance to any drug was observed in 29% overall and 50% of isolates among IFSU. Multidrug-resistant tuberculosis was observed in 8.5% and 17%, respectively. CONCLUSIONS: The population of southern Israel carries very high rates of drug-resistant tuberculosis, mandating quadruple empiric treatment. IFSU should be regarded as having multidrug-resistant tuberculosis until proven otherwise, and empiric therapy with at least five drugs should be considered. This report demonstrates the influence of immigration on the incidence of tuberculosis, and the great value of local surveillance of population-specific resistance rates in an immigrant society, in order to optimize drug treatment and prevent the dissemination of resistant strains. PMID- 10713001 TI - Outcomes of patients with multidrug-resistant pulmonary tuberculosis treated with ofloxacin/levofloxacin-containing regimens. AB - OBJECTIVE: To analyze outcomes of patients with multidrug-resistant tuberculosis (MDR-TB) treated with ofloxacin/levofloxacin-containing regimens. MATERIALS AND METHODS: From February 1990 through June 1997, 63 MDR-TB patients (with bacillary resistance to at least isoniazid and rifampin in vitro) were analyzed retrospectively. Twenty-two patients (34.9%) had had no previous antituberculosis chemotherapy. Each patient received either ofloxacin (53) or levofloxacin (10) even though 13 patients had bacilli resistant to ofloxacin in vitro. The other accompanying drugs mainly included aminoglycosides, cycloserine, ethionamide/prothionamide, and pyrazinamide. Sputum smear and culture examinations for acid-fast bacilli (AFB) were performed monthly for the initial 6 months and then at 2- to 3-month intervals until the end of treatment. Comparison was made between clinical successes and failures using univariate and multiple logistic regression analyses for the following variables: age, sex, presence of cavitation, extent of disease, sputum smear positivity, in vitro resistance to ofloxacin, in vitro resistance to streptomycin and/or ethambutol, treatment adherence, and the number of drugs per regimen. RESULTS: Fifty-one patients (81.0%) were cured, nine patients (14.3%) failed, and three patients (4.7%) died. For the entire group, the mean duration of treatment was 14.0 months, and the mean number of drugs was 4.7. Mean durations of chemotherapy in successful and failed patients were 14.5 and 14.2 months, respectively. Mean time for sputum smear and culture conversions were 1.7 and 2.1 months, respectively. Only cavitation, resistance to ofloxacin, and poor adherence were found to be variables independently associated with adverse outcomes (p < 0.05; odds ratios = 15.9, 13.5, 12.8, respectively). Negative sputum cultures after 2 and 3 months of therapy were 100% predictive of cure. Positive sputum cultures after 2 and 3 months were 52.3% and 84.6% predictive of failure, respectively. One patient (2.1%) relapsed after apparent cure. Twenty-five patients experienced adverse drug reactions, but only 12 of them needed drug modifications. CONCLUSION: Most MDR-TB patients can be treated effectively with ofloxacin/levofloxacin-containing regimens. Presence of cavitation, resistance to ofloxacin in vitro, and poor adherence to therapy portend treatment failure. Monitoring monthly sputum culture for AFB in the initial months of chemotherapy helps predict clinical outcomes. PMID- 10713002 TI - Evaluation of diffusing capacity in patients with a restrictive lung disease. AB - BACKGROUND: In healthy volunteers, the single-breath diffusing capacity of the lung for carbon monoxide (DLCO) decreases and DLCO normalized per liter alveolar volume (VA; DLCO/VA) increases if VA is decreased. We hypothesized that comparison of DLCO/VA with its predicted value at predicted total lung capacity (TLC) will result in an underestimation of the diffusion disorder in patients with a restrictive lung disease, if a similar relationship exists between DLCO/VA and lung volume as found in healthy volunteers. OBJECTIVE: To test this hypothesis, we studied total gas transfer DLCO and DLCO/VA as functions of VA in patients who developed a restrictive lung disease and a diffusion disorder in a short period of time. DESIGN: An observational survey. SETTING: Pulmonary function department. PATIENTS: Thirteen patients without any initial pulmonary pathology who developed the mentioned pulmonary pathology due to bleomycin treatment. INTERVENTIONS: Bleomycin treatment. MEASUREMENTS AND RESULTS: We performed the single-breath test at various VA levels before, during, and after bleomycin treatment. In the majority of the patients, the DLCO vs VA relationship remained parabolic, but shifted downwards during therapy. Therefore, the linear DLCO/VA vs VA relationship shifted downwards, while the negative slope was not changed, indicating the development of a decreased gas transfer. Six patients also developed a volume restriction. CONCLUSIONS: The agreement of the data with the hypothesis increased its probability. Consequently, to evaluate a diffusion disorder, DLCO/VA at a lower actual TLC of patients with a lung restriction should be compared to a reference DLCO/VA at a lung volume equal to the actual TLC. PMID- 10713003 TI - Breath carbon monoxide as an indication of smoking habit. AB - STUDY OBJECTIVE: To assess whether the breath carbon monoxide (CO) concentration can be used to determine a patient's smoking habits in a respiratory outpatient clinic. DESIGN: To provide a normal range for smokers and nonsmokers, 41 outpatients and 24 healthy subjects were questioned on their smoking habits and asked to provide two breaths into a CO monitor (EC50 Smokerlyser; Bedfont Instruments; Kent, UK). In a subsequent single-blind study, 51 different outpatients were not told of the purpose of the study and were assessed by extensive questionnaire, spirometry, and Smokerlyser estimation. SETTING: The Chest Clinic and Pulmonary Medicine Department at the Northern General Hospital, Sheffield, UK. PARTICIPANTS: Phase 1 involved 41 outpatients attending the Chest Clinic and 24 nonoutpatient colleagues. In phase 2, an additional 51 different outpatients were studied. MEASUREMENTS AND RESULTS: The mean (SD) breath CO levels were 17.4 (11.6) parts per million (ppm) for smokers and 1.8 (1.3) ppm for nonsmokers (p < 0.001). A level of 6 ppm was taken as the cutoff, as this gave a selectivity of 96% and a sensitivity of 94% for outpatients. Of the 51 study patients, 5 admitted to smoking in the administered questionnaire. Eight denied smoking but had a mean breath CO > 6 ppm (7.5 to 42 ppm). Of these, three admitted to smoking after being explained the implication of the reading. CONCLUSIONS: Breath CO concentration provides an easy, noninvasive, and immediate way of assessing a patient's smoking status. A reading > 6 ppm strongly suggests that an outpatient is a smoker. PMID- 10713005 TI - Comparative efficacy of positron emission tomography with fluorodeoxyglucose in evaluation of small (<1 cm), intermediate (1 to 3 cm), and large (>3 cm) lymph node lesions. AB - PURPOSE: Our objective was to determine if positron emission tomography (PET) with fluorodeoxyglucose (FDG; PET-FDG) imaging is equally efficacious in detection of metastases in small and large mediastinal lymph nodes as compared to CT scanning. MATERIALS AND METHODS: PET-FDG imaging, CT scanning, and histology results of sampled mediastinal lymph nodes were compared in 54 patients of total 118 patients studied. Efficacy of PET and CT was determined and compared in small (< 1 cm), intermediate (1 to 3 cm), and large (> 3 cm) mediastinal lesions. RESULTS: PET was accurate in 94% of patients in characterizing "N" disease as compared to 61% with CT. Overall, sensitivity, specificity, and accuracy of PET for staging mediastinal lymph nodes (n = 168 in 54 patients) was 96, 93, and 94%, as compared to 68, 65, and 66% with CT. Positive and negative predictive value of PET in detecting mediastinal adenopathy was 86% and 98%, as compared to 47% and 82% with CT, respectively. PET was also highly reliable and accurate for detecting lymph nodes < 1 cm, 1 to 3 cm, and > 3 cm in size with superior efficacy than CT. Sensitivity, specificity, and accuracy of PET for detecting malignancy in lymph node lesions < 1 cm in size was 97, 82, and 95%, respectively. CONCLUSION: PET-FDG imaging is equally reliable and accurate for detecting disease in small and large lymph node lesions in patients with suspected or proven lung cancer with better efficacy than CT. PMID- 10713004 TI - Race and gender differences in the effects of smoking on lung function. AB - STUDY OBJECTIVE: To assess the extent to which the relationship between smoking and lung function in adults varies by gender and race/ethnicity. DESIGN: A random effects metaregression analysis to synthesize results from common cross-sectional regression models fit to participants in each of 10 gender-race strata in each of eight large population-based observational studies or clinical trials. SETTING: Source data collected as part of the most recently completed examination cycle for each of the participating studies. PARTICIPANTS: Participants ranged in age from 30 to 85 years, although the age, race, gender, and general health characteristics of each of the populations varied greatly. INTERVENTIONS: Most of the studies were observational in nature, although some did involve lifestyle interventions. All treatment assignments were ignored in the analysis. MEASUREMENTS AND RESULTS: All studies measured lung function using standardized methods with centrally trained and certified technicians. Study findings confirm statistically significant, dose-related smoking effects in all race-gender groups studied. Significant gender differences in the effects of cigarette smoking were seen only for blacks; black men who smoked had greater smoking-related declines in FEV(1) than did black women. This effect was present in only one of two smoking models, however. Significant racial differences in the effects of smoking were seen only for men, with Asian/Pacific Islanders having smaller smoking related declines than white men in both models. CONCLUSIONS: In summary, this analysis generally failed to support the hypothesis of widespread differences in the effects of cigarette smoking on lung function between gender or racial subgroups. PMID- 10713006 TI - Synchronous roentgenographically occult lung carcinoma in patients with resectable primary lung cancer. AB - OBJECTIVE: To assess the prevalence of synchronous roentgenographically occult lung carcinoma (ROLC) in patients with resectable roentgenographically visible lung cancer (RVLC). METHODS: Patients undergoing surgery for RVLC in the same University Hospital were prospectively evaluated before surgery by fluorescence bronchoscopy under local anesthesia to detect synchronous ROLC. All abnormal areas, with the exception of the RVLC, had biopsies made. RESULTS: From June 1996 to January 1999, 43 patients (male/female ratio: 1.7/1.0) were evaluated before lobectomy (n = 34) or pneumonectomy (n = 10) for 44 primary RVLC. There were 10 T1N0, 19 T2N0, 1 T1N1, 9 T2N1, 1 T3N0, 3 T1N2, and 1 T3N1 lesions. The histologic type was mainly squamous carcinoma (n = 21) and adenocarcinoma (n = 14). All but two patients were smokers or ex-smokers (mean +/- SD, 48 +/- 28 pack-years). A total of 177 endobronchial biopsies were performed (4.1 +/- 2.5); 8 were too small to be informative, 43 showed non-preneoplastic alterations, and 50 were normal. There were 7 basal cell hyperplasias, 56 metaplasias, 9 dysplasias, and 4 carcinomas in situ (CIS). All the dysplasias and CIS lesions were observed in eight subjects. The synchronous CIS were treated by surgery (n = 1) or localized therapeutic modalities (n = 3). CONCLUSIONS: The high prevalence of synchronous early lung cancers (9.3%) as well as metaplasia and dysplasia in this series of patients with resectable RVLC suggests that fluorescence bronchoscopy may be a useful adjunct in the preoperative evaluation of lung cancer. PMID- 10713007 TI - Long-term effect of bilateral plication of the diaphragm. AB - STUDY OBJECTIVES: To assess the feasibility and clinical outcome of bilateral plication of the diaphragm in patients with bilateral diaphragmatic paralysis (BDP) caused by neuralgic amyotrophy (NA), a mononeuritis of the phrenic nerves. DESIGN: Prospective, case-control study over a 1-year period. SETTING: A university hospital in The Netherlands. PATIENTS: Six patients who presented with BDP caused by NA. METHODS: The diagnosis of BDP was based on the absence of muscle response after cervical magnetic stimulation of both phrenic nerves. Three patients did not undergo surgery but were observed for a period of 2 years, and the other three patients underwent a limited lateral thoracotomy at the eighth intercostal space. Plication was performed by U-stitches until the diaphragm was as tight as possible. Vital capacity (VC) and arterial blood gas was measured during follow-up. RESULTS: One month postoperatively, mean VC measured in the supine position was significantly improved by 17%, and this effect was sustained for 12 months. Arterial PO(2) increased by 45%. VC and blood gas levels did not improve in the three patients that were only observed during the 2-year period. All three surgical patients could sleep in the supine position after the operation. CONCLUSION: Bilateral plication of the diaphragm for NA-induced paralysis results in improvement of ventilation and blood gas exchange, allowing patients to sleep in the supine position without dyspnea. PMID- 10713008 TI - Intrapleural administration of a large amount of diluted fibrin glue for intractable pneumothorax. AB - OBJECTIVE: Pleurodesis using chemical agents has been applied to high-risk patients with pneumothorax. This treatment, however, is sometimes unsuccessful in patients with intractable pneumothorax or intrapleural dead space. We developed a technique for the intrapleural administration of diluted fibrin glue as a treatment for such patients. METHODS: Fibrin glue was diluted fourfold with saline solution and/or contrast medium. Pleurodesis with a large amount of the diluted fibrin glue was performed in 40 high-risk patients with intractable pneumothorax and in 13 postthoracotomy patients with persistent air leakage associated with an intrapleural dead space. RESULTS: The air leaks were stopped by administration of the glue in all patients of both groups. During the follow up period, a recurrence rate of 12.5% was observed in the former group. These recurrent pneumothoraces also were successfully treated by glue administration with no further recurrence. In the 13 postthoracotomy patients, there was no recurrence after the initial treatment. Pyrexia (12.5%) and chest discomfort (4.1%) were observed as side effects, but there were no findings of severe chest pain or thoracic empyema. CONCLUSIONS: These results suggest that intrapleural administration of a large amount of diluted fibrin glue is a useful treatment for intractable pneumothoraces in high-risk or postthoracotomy patients who have an intrapleural dead space. PMID- 10713009 TI - Mortality from primary pulmonary hypertension in the United States, 1979-1996. AB - STUDY OBJECTIVES: To determine whether primary pulmonary hypertension mortality in the United States increased since 1979 coincident with the introduction of anorexigens. DESIGN: Examination of annual age-adjusted and age-specific primary pulmonary hypertension mortality in the United States from 1979 through 1996 and in five selected states from 1992 through 1996. SETTING: The United States, from 1979 through 1996. PATIENTS OR PARTICIPANTS: Residents of the United States, from 1979 through 1996. INTERVENTIONS: None. MEASUREMENTS AND RESULTS: Annual age adjusted mortality increased at different rates among white men and women and black men and women. The greatest increase was among black women (who also had the highest rates). Age-specific mortality showed a high rate among infants < 1 year old, a low rate in childhood, and an ascending rate throughout the remainder of life. Similar patterns were identified at the state level. CONCLUSIONS: Primary pulmonary hypertension mortality in the United States has increased notably since 1979. Some portion of this increase may be related to the introduction of anorexigens. Improvements in diagnostic recognition may also explain part of the increase in mortality. These results need to be confirmed in a diagnosis validation study, particularly because the same mortality data suggest that the disease may be more common in the elderly than has been previously reported. PMID- 10713010 TI - Carboxyhemoglobin half-life in carbon monoxide-poisoned patients treated with 100% oxygen at atmospheric pressure. AB - STUDY OBJECTIVES: There are large reported differences for the carboxyhemoglobin (COHb) half-life (COHb t(1/2)) in humans breathing 100% atmospheric O(2) following CO inhalation in tightly controlled experiments compared to the COHb t(1/2) observed in clinical CO poisoning (range, 36 to 131 min, respectively). Other reports have suggested that the COHb t(1/2) may be affected by gender differences, age, and lung function. We wished to test the hypothesis that the COHb t(1/2) might also be influenced by CO poisoning vs experimental CO exposure, by a history of loss of consciousness (LOC), concurrent tobacco smoking, and by PaO(2). The purpose of the present study was to measure the COHb t(1/2) in a cohort of CO-poisoned patients and to determine if those listed factors influenced the COHb t(1/2). DESIGN: Retrospective chart review from 1985 to 1995. We calculated the COHb t(1/2) of CO-poisoned patients who were treated with high flow supplemental atmospheric pressure O(2) delivered by nonrebreather face mask or endotracheal tube. SETTING: Hyperbaric medicine department of a tertiary-care teaching hospital. PATIENTS: Of 240 CO-poisoned patients, 93 had at least two COHb measurements > 2% (upper limit of normal) with recorded times of the measurements, permitting calculation of the COHb t(1/2). RESULTS: The COHb t(1/2) was 74 +/- 25 min (mean +/- 1 SD) with a range from 26 to 148 min. By stepwise multiple linear regression analysis, the PaO(2) influenced the COHb t(1/2) (R(2) = 0.19; p < 0.001), whereas the COHb t(1/2) was not influenced by gender, age, smoke inhalation, history of LOC, concurrent tobacco smoking, degree of initial metabolic acidosis (base excess), or initial COHb level. CONCLUSIONS: The COHb t(1/2) of 93 CO-poisoned patients treated with 100% O(2) at atmospheric pressure was 74 +/- 25 min, considerably shorter than the COHb t(1/2) reported in prior clinical reports (approximately 130 +/- 130 min) and was influenced only by the patient's PaO(2). PMID- 10713011 TI - Sleep in critically ill patients requiring mechanical ventilation. AB - STUDY OBJECTIVES: To objectively measure sleep in critically ill patients requiring mechanical ventilation and to define selection criteria for future studies of sleep continuity in this population. DESIGN: Prospective cohort analysis. SETTING: University teaching hospital medical-surgical ICU. PATIENTS: Twenty critically ill (APACHE II [acute physiology and chronic health evaluation II] acute physiology score [APS], 10 +/- 5), mechanically ventilated adults (male 12, female 8, age 62 +/- 15 years) with mild to moderate acute lung injury (lung injury score, 1.8 +/- 0.9) 10 +/- 7 days after admission to the ICU. MEASUREMENTS AND RESULTS: Patients were divided into three groups based on 24-h polysomnography (PSG) findings. No patient demonstrated normal sleep. In the "disrupted sleep" group (n = 8), electrophysiologic sleep was identified and was distributed throughout the day (6:00 AM to 10:00 PM; 4.0 +/- 2.9 h) and night (10:00 PM to 6:00 AM; 3.0 +/- 1.9 h) with equivalent proportions of non-rapid eye movement (NREM) and rapid eye movement (REM) sleep. Nocturnal sleep efficiency was severely reduced (38 +/- 24%) with an increased proportion of stage 1 NREM sleep (40 +/- 28% total sleep time [TST]) and a reduced proportion of REM sleep (10 +/- 14% TST). Severe sleep fragmentation was reflected by a high frequency of arousals (20 +/- 17/h) and awakenings (22 +/- 25/h). Electrophysiologic sleep was not identifiable in the PSG recordings of the remaining patients. These were classified either as "atypical sleep" (n = 5), characterized by transitions from stage 1 NREM to slow wave sleep with a virtual absence of stage 2 NREM and reduced stage REM sleep, or "coma" (n = 7), characterized by > 50% delta or theta EEG activity with (n = 5) and without (n = 2) evidence of EEG activation either spontaneously or in response to deep painful stimuli. The combined atypical sleep and coma groups had a higher APS (13 +/- 4 vs 6 +/- 4) and higher doses of sedative medications than the disrupted sleep group. CONCLUSION: Sleep, as it is conventionally measured, was identified only in a subgroup of critically ill patients requiring mechanical ventilation and was severely disrupted. We have proposed specific criteria to select patients for future studies to evaluate potential causes of sleep disruption in this population. PMID- 10713012 TI - Dose-response to inhaled aerosolized prostacyclin for hypoxemia due to ARDS. AB - STUDY OBJECTIVES: This study was carried out to determine the efficacy of and dose-response relationships to inhaled aerosolized prostacyclin (IAP), when used as a selective pulmonary vasodilator (SPV) in patients with severe hypoxemia due to ARDS. DESIGN: Unblinded, interventional, prospective clinical study. SETTING: A general ICU in a university-affiliated, tertiary referral center. PATIENTS: Nine adult patients with severe ARDS (lung injury score, > or = 2.5). INTERVENTIONS: All patients received IAP over the dose range 0 to 50 ng/kg/min. The IAP was delivered via a jet nebulizer placed in the ventilator circuit. Dose increments were 10 ng/kg/min every 30 min. MEASUREMENTS AND RESULTS: Cardiovascular parameters (cardiac index and mean pulmonary and systemic pressures), indexes of oxygenation (PaO(2)/fraction of inspired oxygen [FIO(2)] ratio and alveolar-arterial oxygen partial pressure difference [P(A-a)O(2)]) and shunt fraction were measured or calculated at each dose interval, as were platelet aggregation and systemic levels of prostacyclin metabolite (6-keto prostaglandin F1(alpha)). A generalized linear regression model was used to determine a dose effect of IAP on these parameters. The Wilcoxon rank sum test for related measures was used to compare the effects of various doses of IAP. IAP acted as an SPV, with a statistically significant dose-related improvement in PaO(2)/FIO(2) ratio (p = 0.003) and P(A-a)O(2) (p = 0.01). Systemic prostacyclin metabolite levels increased significantly in response to delivered IAP (p = 0.001). There was no significant dose effect on systemic or pulmonary arterial pressures, or on platelet function, as determined by platelet aggregation in response to challenge with adenosine diphosphate. CONCLUSIONS: IAP is an efficacious SPV, with marked dose-related improvement in oxygenation and with no demonstrable effect on systemic arterial pressures over the dose range 0 to 50 ng/kg/min. Despite significant systemic levels of prostacyclin metabolite, there was no demonstrable platelet function defect. PMID- 10713013 TI - Predicting the result of noninvasive ventilation in severe acute exacerbations of patients with chronic airflow limitation. AB - OBJECTIVE: To analyze prospectively the factors related to the success of noninvasive ventilation (NIV) in the treatment of acute exacerbations of chronic airflow limitation (CAFL) and to generate a multiple regression model in order to detect which patients can be successfully treated by this method. SETTING: A respiratory medicine ward of a referral hospital. METHODS AND PRINCIPAL RESULTS: Initially, we examined 44 episodes of acute respiratory failure in 36 patients with CAFL in whom mechanical ventilation was advisable. In 34 of 44 episodes (77%), NIV was used successfully. Patients in whom NIV succeeded had a lower FEV(1) prior to admission, a higher level of consciousness (LC), and significant improvements in PaCO(2), pH, and LC after 1 h of NIV. A logistic regression model consisting of baseline FEV(1) and PaCO(2) values, initial PaCO(2), pH, and LC values on admission, and PaCO(2) values after 1 h of NIV allowed us to correctly classify > 95% of the 44 episodes in which the outcome was successful. In the second part of the study, we prospectively validated the equation in another 15 consecutive CAFL patients with acute hypercapnic respiratory failure. NIV successfully treated 12 patients (80%), and the model correctly classified 14 patients (93%). CONCLUSION: Good LC at the beginning of NIV and improvements in pH, PaCO(2), and LC values after 1 h of NIV are associated with successful responses to NIV in COPD patients with acute hypercapnic respiratory failure. Our validated multiple regression model confirms that these variables predict the result of NIV in acute hypercapnic failure in CAFL patients. PMID- 10713014 TI - Radionuclide imaging of acute lung transplant rejection with annexin V. AB - STUDY OBJECTIVES: Early detection and treatment of lung transplant rejection is critical for preservation of pulmonary graft function. Damage to pulmonary allografts is mediated by apoptotic cell death induced by the alloreactive T lymphocytes that infiltrate lung grafts. Previous studies demonstrate that acute cardiac allograft rejection can be visualized using radiolabeled annexin V. This study was done to determine whether this technique could visualize acute rejection in a rodent model of unilateral orthotopic lung transplantation. DESIGN: Eighteen Sprague-Dawley ACI rats underwent removal of their left lung followed by orthotopic transplant of either an allogeneic (PVG, immunologically mismatched; N = 10) or a syngeneic (ACI, immunologically matched) pulmonary graft (N = 8). Animals were imaged 1 h after IV injection of 1 mCi (37.0 MBq) of (99m)Tc-annexin V 1 to 7 days after transplantation. RESULTS: Lungs receiving the allograft demonstrated moderate to marked mononuclear infiltration of the perivascular, interstitial, and peribronchial tissues. No mononuclear infiltrates were noted in the native right lungs nor in the syngeneic transplants. Region of interest image analysis revealed significant (p < 0.0005) increases of transplant to normal lung activity ratios 3 to 7 days after allograft surgery. The increased annexin V uptake in these lungs was confirmed at biodistribution assay (allograft 151% greater than isograft activity, p < 0.005). CONCLUSIONS: Acute experimental lung transplant rejection can be noninvasively identified using (99m)Tc-annexin V. Radiolabeled annexin V may be a clinically useful noninvasive screening tool for acute rejection. PMID- 10713015 TI - Endothelial apoptosis: could it have a role in the pathogenesis and treatment of disease? AB - Endothelial apoptosis can be found in a number of diseases. This review summarizes the current knowledge about the causes and consequences of endothelial apoptosis, and analyzes its possible role in the pathogenesis and treatment of several diseases. Novel forms of therapy based on the proposed pathophysiologic mechanisms are discussed. PMID- 10713016 TI - Fever in the ICU. AB - Fever is a common problem in ICU patients. The presence of fever frequently results in the performance of diagnostic tests and procedures that significantly increase medical costs and expose the patient to unnecessary invasive diagnostic procedures and the inappropriate use of antibiotics. ICU patients frequently have multiple infectious and noninfectious causes of fever, necessitating a systematic and comprehensive diagnostic approach. Pneumonia, sinusitis, and blood stream infection are the most common infectious causes of fever. The urinary tract is unimportant in most ICU patients as a primary source of infection. Fever is a basic evolutionary response to infection, is an important host defense mechanism and, in the majority of patients, does not require treatment in itself. This article reviews the common infectious and noninfectious causes of fever in ICU patients and outlines a rational approach to the management of this problem. PMID- 10713017 TI - Anorexigens and pulmonary hypertension in the United States: results from the surveillance of North American pulmonary hypertension. AB - BACKGROUND: The use of appetite suppressants in Europe has been associated with the development of primary pulmonary hypertension (PPH). Recently, fenfluramine appetite suppressants became widely used in the United States but were withdrawn in September 1997 because of concerns over adverse effects. MATERIALS AND METHODS: We conducted a prospective surveillance study on patients diagnosed with pulmonary hypertension at 12 large referral centers in North America. Data collected on patients seen from September 1, 1996, to December 31, 1997, included the cause of the pulmonary hypertension and its severity. Patients with no identifiable cause of pulmonary hypertension were classed as PPH. A history of drug exposure also was taken with special attention on the use of antidepressants, anorexigens, and amphetamines. RESULTS: Five hundred seventy nine patients were studied, 205 with PPH and 374 with pulmonary hypertension from other causes (secondary pulmonary hypertension [SPH]). The use of anorexigens was common in both groups. However, of the medications surveyed, only the fenfluramines had a significant preferential association with PPH as compared with SPH (adjusted odds ratio for use > 6 months, 7.5; 95% confidence interval, 1.7 to 32.4). The association was stronger with longer duration of use when compared to shorter duration of use and was more pronounced in recent users than in remote users. An unexpectedly high (11.4%) number of patients with SPH had used anorexigens. CONCLUSION: The magnitude of the association with PPH, the increase of association with increasing duration of use, and the specificity for fenfluramines are consistent with previous studies indicating that fenfluramines are causally related to PPH. The high prevalence of anorexigen use in patients with SPH also raises the possibility that these drugs precipitate pulmonary hypertension in patients with underlying conditions associated with SPH. PMID- 10713018 TI - Should health-care systems pay for replacement therapy in patients with alpha(1) antitrypsin deficiency? A critical review and cost-effectiveness analysis. AB - STUDY OBJECTIVES: Assess cost effectiveness for providing alpha(1)-antitrypsin (alpha(1)-AT) replacement therapy to individuals with severe COPD and alpha(1)-AT deficiency. MATERIALS AND METHODS: The electronic databases MEDLINE and EMBASE were searched, and relevant bibliographies were reviewed. Effect size, defined as the absolute risk difference between treated and untreated groups, was taken from the highest level of supporting evidence. The cost for providing alpha(1)-AT replacement therapy was analyzed from a payer perspective and was based on Medicare reimbursement rates. Effect size and costs were varied. The year of life saved was discounted up to 7%. RESULTS: The incremental cost per year of life saved for alpha(1)-AT replacement therapy (60 mg/kg/wk IV) in a 70-kg subject with severe alpha(1)-AT deficiency and an FEV(1) < 50% of predicted based on the National Institutes of Health (NIH) Registry mortality rate data is $13,971. The incremental cost depends substantially on the mortality rate reduction. When the effect size is altered from 10 to 70%, with the cost fixed at $52,000, the incremental cost per year of life saved ranges from $152,941 to $7,330. When effect size is 55% (as in the NIH Registry) but costs are increased almost 300%, from $52,000 to $150,000 per year, then the incremental cost per year of life saved increases from $13,971 to $40,301. CONCLUSION: No randomized, placebo controlled trials are available to assess mortality rate reduction with alpha(1) AT replacement therapy. The best currently available data are observational, from the NIH Registry. Based on these data, alpha(1)-AT replacement therapy is cost effective in individuals who have severe alpha(1)-AT deficiency and severe COPD. PMID- 10713019 TI - Calculation of provocative concentration causing a 20% fall in FEV(1): comparison of lowest vs highest post-challenge FEV(1). AB - BACKGROUND: Considerable, unexamined controversy exists surrounding the use of the highest vs the lowest FEV(1) for calculating the provocative concentration causing a 20% fall in FEV(1) (PC(20)) during direct bronchoprovocation challenges. OBJECTIVE: To compare the PC(20) calculated using the lowest FEV(1) post-diluent and post-histamine/methacholine vs the PC(20) calculated using the highest FEV(1) post-diluent and post-histamine/methacholine. METHOD: Retrospective analysis of 225 challenges: 75 research methacholine challenges, 75 research histamine challenges, and 75 clinical methacholine challenges. For each test, the PC(20) was calculated twice, first using the lowest post-diluent FEV(1) to the lowest post-histamine/methacholine FEV(1), and then using the highest to the highest. RESULTS: The intraclass correlation coefficients for methacholine research, histamine research, and methacholine clinic challenges were 0.99, 0.98, and 0.95, respectively. The PC(20) calculated using the lowest to lowest FEV(1) was slightly and significantly lower in all three groups (paired t test p < 0.0001). CONCLUSIONS: The PC(20) values calculated using the highest FEV(1) are almost identical to the PC(20) values calculated using the lowest FEV(1). The difference, although clinically irrelevant, holds statistical significance. PMID- 10713020 TI - Video-assisted contralateral treatment for bronchial stump diastasis after left pneumonectomy. AB - Postoperative bronchial stump failure is a life-threatening complication, and several surgical approaches and procedures have been developed to close the stump. In this report, we describe a case of left mainstem bronchial stump diastasis after pneumonectomy for lung cancer, in which the bronchial stump was re-closed using a contralateral approach with video-assisted thoracic surgery, with good success. The left main bronchus was closed with an automatic stapler device, but the stump reopened and left pyothorax developed postoperatively. Emergent intratracheal intubation and ventilation was required due to rapid progression of right pyothorax. Under strict nutritional management by IV hyperalimentation, administration of antibiotics to which the organisms were sensitive, and drainage, the patient recovered from pneumonia. However, thoracic air leak increased daily, and reoperation for bronchial diastasis was performed. Using this approach, the left main bronchus near the carina was easily exposed extrapleurally, with only the azygos vein being incised. Video-assisted contralateral treatment was effective in avoiding sternal osteomyelitis due to a transpericardial approach via median sternotomy in the case of mainstem bronchial stump failure, only after left pneumonectomy. PMID- 10713021 TI - Tissue effects of bronchoscopic electrocautery: bronchoscopic appearance and histologic changes of bronchial wall after electrocautery. AB - STUDY OBJECTIVES: To study tissue effects of bronchoscopic electrocautery (BE). DESIGN: In six patients with non-small cell lung cancer, a BE procedure was performed immediately before surgery. After patients were placed on ventilation, normal mucosa on different carinae was treated with a cautery probe (2-mm(2) surface area) at a power setting of 30 W with a variable time of application of 1 to 5 s. Bronchoscopic appearance of the treated area was documented photographically, and histologic changes of the bronchial wall were examined. SETTING: Bronchoscopy unit of a university hospital. MEASUREMENTS AND RESULTS: BE resulted bronchoscopically in whitening of the bronchial mucosa with crater shaped lesions. After longer duration of BE application, deeper craters with more profound charring were seen. Histologic changes of the lesions showed craters containing a variable amount of necrotic tissue. In one case, thin subsegmental carinae were coagulated and measurements could not be performed. In the remaining five cases, microscopic findings revealed 0.2 +/- 0.1-mm necrosis after 1 s; 0.4 +/- 0.2-mm necrosis after 2 s; 0.9 +/- 0.5-mm necrosis after 3 s; and 1.9 +/- 0.8 mm necrosis after 5 s. A variable degree of tissue damage surrounding the necrotic tissue area was found. In one case, cartilage damage appeared after 3 s of coagulation, and extensive damage of the underlying cartilage was seen in four cases after 5 s of application. CONCLUSIONS: Superficial damage was obtained by short duration of BE (< or = 2 s), and longer duration of coagulation (3 s or 5 s) caused damage to the underlying cartilage. Bronchoscopic appearance after endobronchial electrocautery corresponded with the histologic changes. PMID- 10713022 TI - A 55-year-old man admitted to the medical ICU with the diagnosis of status asthmaticus responds poorly to intensive asthma therapy. PMID- 10713024 TI - A large thoracic mass in a 57-year-old patient. Solitary fibrous tumor of the pleura. PMID- 10713023 TI - A 52-year-old man with a lung mass and acute abdominal pain. PMID- 10713025 TI - Adult presentation of agenesis of the hemidiaphragm. AB - A 66-year-old woman presented with a 3-day history of classical features of large bowel obstruction. At emergency laparotomy, the transverse colon and splenic flexure were located in the left hemithorax. The entire left hemidiaphragm was absent, and there were no diaphragmatic remnants visible. This is the oldest reported case of an absent hemidiaphragm. Previous cases of "agenesis" of the hemidiaphragm in adults either reported diaphragmatic remnants intraoperatively or failed to rule them out radiologically when managed conservatively. We would suggest that this is the first reported case of an adult presenting with true agenesis of the hemidiaphragm. PMID- 10713026 TI - Conservative treatment of postsurgical lymphatic leaks with somatostatin-14. AB - Successful management of lymphatic leaks by continuous IV administration of somatostatin was first reported by Ulibarri and coworkers in Spain,(1) and more recently by authors from Italy(2) and Switzerland.(3) The present article reports the clinical history of two patients in whom postsurgical lymphatic leak was successfully treated after the administration of either somatostatin-14 alone (case 1) or combined somatostatin-14 and total parenteral nutrition (TPN; case 2). Although further pathophysiologic studies are needed for the elucidation of its mechanisms of action, somatostatin-14 seems to be an intriguing therapy against postsurgical lymphatic leaks that may make potentially risky transthoracic reoperation unnecessary. PMID- 10713027 TI - Giant cell myocarditis responding to immunosuppressive therapy. AB - An unusual case of giant cell myocarditis presenting with cardiogenic shock that dramatically responded to conventional dose of steroids and azathioprine is reported. Cardiac recovery was rapid, complete (left ventricular ejection fraction rose to 55% from 10%), and was accompanied by the disappearance of the inflammatory infiltrates including giant cells in the control endomyocardial biopsy. Maintenance of the recovery at 16 months of follow-up on a low dose of azathioprine suggests that giant cell myocarditis might be a heterogeneous disease having either a negative untreatable trend necessitating cardiac transplantation, or a curable substrate responding to immunosuppressive drugs. PMID- 10713028 TI - The surgical management of severe gastroparesis in heart/lung transplant recipients. AB - This article describes the use of gastric bypass surgery for severe gastroparesis in two lung transplant recipients. In addition to feeding intolerance, both our patients suffered from severe erosive esophagitis, transfusion-dependent upper GI hemorrhage, and recurrent aspiration pneumonia. They responded poorly to promotility agents and were eventually treated with Roux-en-Y esophagojejunostomy one patient with subtotal gastrectomy, and one with gastric bypass without distal gastric resection. Both cases were improved by surgery. Early surgical referral may be indicated in the management of lung transplant recipients with severe symptomatic gastroparesis in whom medical management has failed. On the basis of our experience, gastric bypass with esophagojejunostomy is a worthwhile option in lung transplant recipients with severe gastroparesis. PMID- 10713029 TI - Effect of pericardial pressure on human coronary circulation. AB - A 52-year-old patient underwent percutaneous balloon pericardiotomy because of rapid fluid accumulation. During the procedure, we calculated the amount of blood flow to the nondiseased left anterior descending coronary artery while pericardial pressure was gradually increased by the infusion of warmed normal saline solution. Coronary vasodilator reserve was assessed by intracoronary adenosine. With increasing pericardial pressure, there was a continuous decline in coronary blood flow, due to an increase in coronary vascular resistance, and an unaffected hyperemic response throughout. The maximal hyperemic flow was far less under increased pericardial pressure than at normal pressure, which implies an augmented susceptibility to myocardial ischemia. PMID- 10713030 TI - Pancreaticopleural fistula: diagnosis with magnetic resonance pancreatography. AB - Pancreaticopleural fistula secondary to chronic pancreatitis is a rare cause of recurrent pleural effusion. The demonstration of the fistula with endoscopic retrograde pancreatography and CT is invasive or limited. We report in two patients the use of magnetic resonance pancreatography as a noninvasive alternative to endoscopic retrograde pancreatography for the diagnosis of pancreaticopleural fistula. PMID- 10713031 TI - Treatment of obstructive airway disease with a cysteine donor protein supplement: a case report. AB - Oxidant/antioxidant imbalance can occur in obstructive airways disease as a result of ongoing inflammation. Glutathione (GSH) plays a major role in pulmonary antioxidant protection. As an alternative or complement to anti-inflammatory therapy, augmenting antioxidant protection could diminish the effects of inflammation. We describe a case of a patient who had obstructive lung disease responsive to corticosteroids, and low whole blood GSH levels. After 1 month of supplementation with a whey-based oral supplement designed to provide GSH precursors, whole blood GSH levels and pulmonary function increased significantly and dramatically. The potential for such supplementation in pulmonary inflammatory conditions deserves further study. PMID- 10713032 TI - An unreported risk in the use of home nasal continuous positive airway pressure and home nasal ventilation in children: mid-face hypoplasia. AB - We report the case of a 15-year-old boy with obstructive sleep apnea and obesity who was treated since the age of 5 with nasal continuous positive airway pressure. Due to the long-term use of a nasal mask, the child developed a mid face hypoplasia. Chronic use of a nasal mask for home ventilation in children should always be associated with regular evaluations of maxillomandibular growth. PMID- 10713033 TI - Hand hygiene in the ICU. PMID- 10713034 TI - Treatment of right heart thromboemboli with IV recombinant tissue-type plasminogen activator. PMID- 10713035 TI - Natural decrease of benign metastasizing leiomyoma. PMID- 10713036 TI - Spirometry in the diagnosis of small airways obstruction. PMID- 10713037 TI - Inflammatory pseudotumor of the lung with pleural thickening treated with corticosteroids. PMID- 10713038 TI - Pulmonary embolism in a patient with coagulopathy from end-stage liver disease. PMID- 10713039 TI - Home mechanical ventilation in amyotrophic lateral sclerosis patients is not always a problem. PMID- 10713040 TI - Humidification during continuous positive airway pressure therapy. PMID- 10713041 TI - Crystal structure of the human U4/U6 small nuclear ribonucleoprotein particle specific SnuCyp-20, a nuclear cyclophilin. AB - The cyclophilin SnuCyp-20 is a specific component of the human U4/U6 small nuclear ribonucleoprotein particle involved in the nuclear splicing of pre-mRNA. It stably associates with the U4/U6-60kD and -90kD proteins, the human orthologues of the Saccharomyces cerevisiae Prp4 and Prp3 splicing factors. We have determined the crystal structure of SnuCyp-20 at 2.0-A resolution by molecular replacement. The structure of SnuCyp-20 closely resembles that of human cyclophilin A (hCypA). In particular, the catalytic centers of SnuCyp-20 and hCypA superimpose perfectly, which is reflected by the observed peptidyl-prolyl cis/trans-isomerase activity of SnuCyp-20. The surface properties of both proteins, however, differ significantly. Apart from seven additional amino terminal residues, the insertion of five amino acids in the loop alpha1-beta3 and of one amino acid in the loop alpha2-beta8 changes the conformations of both loops. The enlarged loop alpha1-beta3 is involved in the formation of a wide cleft with predominantly hydrophobic character. We propose that this enlarged loop is required for the interaction with the U4/U6-60kD protein. PMID- 10713042 TI - Differential targeting of Shaker-like potassium channels to lipid rafts. AB - Ion channel targeting within neuronal and muscle membranes is an important determinant of electrical excitability. Recent evidence suggests that there exists within the membrane specialized microdomains commonly referred to as lipid rafts. These domains are enriched in cholesterol and sphingolipids and concentrate a number of signal transduction proteins such as nitric-oxide synthase, ligand-gated receptors, and multiple protein kinases. Here, we demonstrate that the voltage-gated K(+) channel Kv2.1, but not Kv4.2, targets to lipid rafts in both heterologous expression systems and rat brain. The Kv2.1 association with lipid rafts does not appear to involve caveolin. Depletion of cellular cholesterol alters the buoyancy of the Kv2.1 associated rafts and shifts the midpoint of Kv2.1 inactivation by nearly 40 mV without affecting peak current density or channel activation. The differential targeting of Kv channels to lipid rafts represents a novel mechanism both for the subcellular sorting of K(+) channels to regions of the membrane rich in signaling complexes and for modulating channel properties via alterations in lipid content. PMID- 10713043 TI - Fidelity of human DNA polymerase eta. AB - Xeroderma pigmentosum (XP) patients are highly sensitive to sunlight, and they suffer from a high incidence of skin cancers. The variant form of XP results from mutations in the hRAD30A gene, which encodes the DNA polymerase in humans, hPol(eta). Of the eukaryotic DNA polymerases, only human Pol(eta) and its yeast counterpart have the ability to replicate DNA containing a cis-syn thymine thymine (T-T) dimer. Here we measure the fidelity of hPol(eta) on all four nondamaged template bases and at each thymine residue of a cis-syn T-T dimer. Opposite all four nondamaged template bases, hPol(eta) misincorporates nucleotides with a frequency of approximately 10(-2)-10(-3), and importantly, hPol(eta) synthesizes DNA opposite the T-T dimer with the same accuracy and efficiency as opposite the nondamaged DNA. The low fidelity of hPol(eta) may derive from a flexible active site that renders the enzyme more tolerant of geometric distortions in DNA and enables it to synthesize DNA past a T-T dimer. PMID- 10713044 TI - Human DNA damage checkpoint protein hRAD9 is a 3' to 5' exonuclease. AB - Human RAD9 protein (hRAD9) is a homolog of the fission yeast Rad9 protein, one of the six so-called checkpoint Rad proteins involved in the early steps of DNA damage checkpoint response in Schizosaccharomyces pombe. It has been shown previously that, in vivo, a highly modified form of hRAD9 makes a ternary complex with two other checkpoint Rad proteins, hRAD1 and hHUS1 (Volkmer, E., and Karnitz, L. M. (1999) J. Biol. Chem. 274, 567-570; St. Onge, R. P., Udell, C. M., Casselman, R., and Davey, S. (1999) Mol. Biol. Cell. 10, 1985-1995). However, the function of this complex is not known at present. To help define the functions of checkpoint Rad proteins in humans, we expressed hRAD9 in Escherichia coli, purified the recombinant protein and characterized it. We found that hRAD9 is a 3' to 5' exonuclease and located the nuclease active site to the region between residues 51 and 91 of the 391-amino acid-long protein. Our results suggest that exonucleolytic processing of primary DNA lesion by hRAD9 may contribute to DNA damage checkpoint response in humans. PMID- 10713045 TI - Brain copper content and cuproenzyme activity do not vary with prion protein expression level. AB - Prion diseases are neurodegenerative disorders that result from conformational transformation of a normal cell surface glycoprotein, PrP(C), into a pathogenic isoform, PrP(Sc). Although the normal physiological function of PrP(C) has remained enigmatic, the recent observation that the protein binds copper ions with micromolar affinity suggests a possible role in brain copper metabolism. In this study, we have used mice that express 0, 1, and 10 times the normal level of PrP to assess the effect of PrP expression level on the amount of brain copper and on the properties of two brain cuproenzymes. Using mass spectrometry, we find that the amount of ionic copper in subcellular fractions from brain is similar in all three lines of mice. In addition, the enzymatic activities of Cu-Zn superoxide dismutase and cytochrome c oxidase in brain extracts are similar in these groups of animals, as is the incorporation of (64)Cu into Cu-Zn superoxide dismutase both in cultured cerebellar neurons and in vivo. Our results differ from those of another set of published studies, and they require a re-evaluation of the role of PrP(C) in copper metabolism. PMID- 10713046 TI - Pituitary tumor transforming gene (PTTG) transforming and transactivation activity. AB - Pituitary tumor transforming gene (PTTG) is a newly identified transforming gene, the functional mechanism of which is little understood. Computational analysis reveals a C terminus rich in Glu and Pro, a known characteristic of transcriptional activation domains. We report here that murine PTTG indeed possesses transactivation ability, which correlates highly with its transforming properties. Pro(139), Ser(159), Pro(157)-Pro(158)-Ser(159)-Pro(160) (PPXP motif), and Leu(120)-Asp(121)-Phe(122)-Asp(123)-Leu(124) were found to be important for transactivation. Mutation to Ala at a key Pro(139) residue not only disrupted the transactivation function but also resulted in the loss of transforming ability in NIH3T3 cells. A murine PTTG cDNA that encodes a variant C-terminal tail (Gly-Lys Gly-Val-Arg-Ser-Asn-Gly-Cys-Lys-Asp-Leu-Val-Thr) was cloned. This novel PTTG is devoid of transactivation and transforming ability; deletion of its variant C terminal tail restores both transactivation and transforming ability. These results show a high correlation between the transforming and transactivation functions of PTTG and also indicate that the novel PTTG variant may function as an endogenous competitor to wild-type PTTG. PMID- 10713047 TI - A protein conjugation system in yeast with homology to biosynthetic enzyme reaction of prokaryotes. AB - Protein conjugation, such as ubiquitination, is the process by which the C terminal glycine of a small modifier protein is covalently attached to target protein(s) through sequential reactions with an activating enzyme and conjugating enzymes. Here we report on a novel protein conjugation system in yeast. A newly identified ubiquitin related modifier, Urm1 is a 99-amino acid protein terminated with glycine-glycine. Urm1 is conjugated to target proteins, which requires the C terminal glycine of Urm1. At the first step of this reaction, Urm1 forms a thioester with a novel E1-like protein, Uba4. Deltaurm1 and Deltauba4 cells showed a temperature-sensitive growth phenotype. Urm1 and Uba4 show similarity to prokaryotic proteins essential for molybdopterin and thiamin biosynthesis, although the Urm1 system is not involved in these pathways. This is the fifth conjugation system in yeast, following ubiquitin, Smt3, Rub1, and Apg12, but it is unique in respect to relation to prokaryotic enzyme systems. This fact may provide an important clue regarding evolution of protein conjugation systems in eukaryotic cells. PMID- 10713048 TI - The pleckstrin homology domain of phospholipase C-beta(2) links the binding of gbetagamma to activation of the catalytic core. AB - Pleckstrin homology (PH) domains are membrane tethering devices found in many signal transducing proteins. These domains also couple to the betagamma subunits of GTP binding proteins (G proteins), but whether this association transmits allosteric information to the catalytic core is unclear. To address this question, we constructed protein chimeras in which the PH domain of phospholipase C-beta(2) (PLC-beta(2)), which is regulated by Gbetagamma, replaces the PH domain of PLC-delta(1) which binds to, but is not regulated by, Gbetagamma. We found that attachment of the PH domain of PLC-beta(2) onto PLC-delta(1) not only causes the membrane-binding properties of PLC-delta(1) to become similar to those of PLC beta(2), but also results in a Gbetagamma-regulated enzyme. Thus, PH domains are more than simple tethering devices and mediate regulatory signals to the host protein. PMID- 10713049 TI - Interaction between protein kinase C delta and the c-Abl tyrosine kinase in the cellular response to oxidative stress. AB - Protein kinase C (PKC) isoforms are phosphorylated on tyrosine in the response of cells to oxidative stress. The present studies demonstrate that treatment of cells with hydrogen peroxide (H(2)O(2)) induces binding of the PKCdelta isoform and the c-Abl protein-tyrosine kinase. The results show that c-Abl phosphorylates PKCdelta in the H(2)O(2) response. We also show that PKCdelta phosphorylates and activates c-Abl in vitro. In cells, induction of c-Abl activity by H(2)O(2) is attenuated by the PKCdelta inhibitor, rottlerin, and by overexpression of the regulatory domain of PKCdelta. These findings support a functional interaction between PKCdelta and c-Abl in the cellular response to oxidative stress. PMID- 10713050 TI - Involvement of focal adhesion kinase in hepatocyte growth factor-induced scatter of Madin-Darby canine kidney cells. AB - Focal adhesion kinase (FAK) has been implicated to play a critical role in integrin-mediated control of cell behavior. However, it is unclear whether FAK also participates in the regulation of growth factor-elicited cellular functions. In this study, we have demonstrated that although overexpression of FAK in Madin Dardy canine kidney cells did not alter their growth property or ability to form tubules within collagen gel upon hepatocyte growth factor (HGF) stimulation, it apparently enhanced HGF-induced cell scattering. This enhancement was largely because of an increase in the third phase (i.e. cell migration) of cell scattering rather than the first two phases (i.e. cell spreading and cell-cell dissociation). Conversely, the expression of FAK-related nonkinase significantly ( approximately 60%) inhibited HGF-induced cell migration. Moreover, we have found that the effect of FAK on promoting HGF-induced cell motility was greatly dependent on cell-matrix interactions. We showed that HGF treatment selectively increased the expression of integrins alpha(2) and, to a lesser extent, alpha(3) in Madin-Dardy canine kidney cells and that a monoclonal antibody against integrin alpha(2) efficiently blocked HGF-enhanced cell migration on collagen. In our efforts to determine the mechanism by which FAK promotes HGF-induced cell migration, we found that FAK mutants deficient in phosphatidylinositol 3-kinase or p130(Cas) binding failed to promote HGF-induced cell migration. Interestingly, cells expressing a FAK mutant defective in Grb2 binding exhibited a rate of migration approximately 50% lower than that of cells expressing wild type FAK in response to HGF stimulation. Taken together, our results suggest a link between HGF-increased integrin expression, FAK activation, and enhanced cell motility and implicate a role for FAK in the facilitation of growth factor-induced cell motility. PMID- 10713051 TI - Epidermal growth factor-stimulated tyrosine phosphorylation of caveolin-1. Enhanced caveolin-1 tyrosine phosphorylation following aberrant epidermal growth factor receptor status. AB - Caveolin-1 is the major coat protein of caveolae and has been reported to interact with various intracellular signaling molecules including the epidermal growth factor (EGF) receptor. To investigate the involvement of caveolin-1 in EGF receptor action, we used mouse B82L fibroblasts transfected with (a) wild type EGF receptor, (b) a C-terminally truncated EGF receptor at residue 1022, (c) a C terminally truncated EGF receptor at residue 973, or (d) a kinase-inactive EGF receptor (K721M). Following EGF treatment, there was a distinct electrophoretic mobility shift of the caveolin-1 present in cells expressing the truncated forms of the EGF receptor, but this shift was not detectable in cells bearing either normal levels of the wild type EGF receptor or a kinase-inactive receptor. This mobility shift was also not observed following the addition of other cell stimuli, such as platelet-derived growth factor, insulin, basic fibroblast growth factor, or phorbol 12-myristate 13-acetate. Analysis of caveolin-1 immunoprecipitates from EGF-stimulated or nonstimulated cells demonstrated that the EGF-induced mobility shift of caveolin-1 was associated with its tyrosine phosphorylation in cells expressing truncated EGF receptors. Maximal caveolin-1 phosphorylation was achieved within 5 min after exposure to 10 nM EGF and remained elevated for at least 2 h. Additionally, several distinct phosphotyrosine-containing proteins (60, 45, 29, 24, and 20 kDa) were co immunoprecipitated with caveolin-1 in an EGF-dependent manner. Furthermore, the Src family kinase inhibitor, PP1, does not affect autophosphorylation of the receptor, but it does inhibit the EGF-induced mobility shift and phosphorylation of caveolin-1. Conversely, the MEK inhibitors PD98059 and UO126 could attenuate EGF-induced mitogen-activated protein kinase activation, they do not affect the EGF-induced mobility shift of caveolin-1. Because truncation and overexpression of the EGF receptor have been linked to cell transformation, these results provide the first evidence that the tyrosine phosphorylation of caveolin-1 occurs via an EGF-sensitive signaling pathway that can be potentiated by an aberrant activity or expression of various forms of the EGF receptor. PMID- 10713052 TI - Novel human secreted phospholipase A(2) with homology to the group III bee venom enzyme. AB - Venom and mammalian secreted phospholipases A(2) (sPLA(2)s) have been associated with numerous physiological, pathological, and toxic processes. So far, structurally related group I and II sPLA(2)s have been found in vertebrates such as mammals and snakes, whereas group III sPLA(2)s have mainly been found in venom from invertebrates such as bees and scorpions. Here we report the cloning and expression of a cDNA coding for a human group III (hGIII) sPLA(2). The full length cDNA codes for a signal peptide of 19 residues followed by a protein of 490 amino acids made up of a central sPLA(2) domain (141 residues) flanked by large N- and C-terminal regions (130 and 219 residues, respectively). The sPLA(2) domain is 31% identical to bee venom sPLA(2) and displays all of the features of group III sPLA(2)s including 10 cysteines. The hGIII sPLA(2) gene consists of at least 7 exons and maps to chromosome 22q. By Northern blot analysis, a 4.4 kilobase hGIII transcript was found in kidney, heart, liver, and skeletal muscle. Transfection of hGIII sPLA(2) cDNA in COS cells led to accumulation of sPLA(2) activity in the culture medium, indicating that the cDNA codes for a secreted enzyme. Using small unilamellar vesicles as substrate, hGIII sPLA(2) was found to be a Ca(2+)-dependent enzyme showing an 11-fold preference for phosphatidylglycerol over phosphatidylcholine and optimal activity at pH 8. PMID- 10713053 TI - Characterization of the linkage between the type III capsular polysaccharide and the bacterial cell wall of group B Streptococcus. AB - The capsular polysaccharide of group B Streptococcus is a key virulence factor and an important target for protective immune responses. Until now, the nature of the attachment between the capsular polysaccharide and the bacterial cell has been poorly defined. We isolated insoluble cell wall fragments from lysates of type III group B Streptococcus and showed that the complexes contained both capsular polysaccharide and group B carbohydrate covalently bound to peptidoglycan. Treatment with the endo-N-acetylmuramidase mutanolysin released soluble complexes of capsular polysaccharide linked to group B carbohydrate by peptidoglycan fragments. Capsular polysaccharide could be enzymatically cleaved from group B carbohydrate by treatment of the soluble complexes with beta-N acetylglucosaminidase, which catalyzes hydrolysis of the beta-D-GlcNAc(1-->4)beta D-MurNAc subunit produced by mutanolysin digestion of peptidoglycan. Evidence from gas chromatography/mass spectrometry and (31)P NMR analysis of the separated polysaccharides supports a model of the group B Streptococcus cell surface in which the group B carbohydrate and the capsular polysaccharide are independently linked to the glycan backbone of cell wall peptidoglycan; group B carbohydrate is linked to N-acetylmuramic acid, and capsular polysaccharide is linked via a phosphodiester bond and an oligosaccharide linker to N-acetylglucosamine. PMID- 10713054 TI - A beta1,3-glucan recognition protein from an insect, Manduca sexta, agglutinates microorganisms and activates the phenoloxidase cascade. AB - Pattern recognition proteins function in innate immune responses by binding to molecules on the surface of invading pathogens and initiating host defense reactions. We report the purification and molecular cloning of a cDNA for a 53 kDa beta1,3-glucan-recognition protein from the tobacco hornworm, Manduca sexta. This protein is constitutively expressed in fat body and secreted into hemolymph. The protein contains a region with sequence similarity to several glucanases, but it lacks glucanase activity. It binds to the surface of and agglutinates yeast, as well as gram-negative and gram-positive bacteria. Beta1,3-glucan-recognition protein in the presence of laminarin, a soluble glucan, stimulated activation of prophenoloxidase in plasma, whereas laminarin alone did not. These results suggest that beta1,3-glucan-recognition protein serves as a pattern recognition molecule for beta1,3-glucan on the surface of fungal cell walls. After binding to beta1,3-glucan, the protein may interact with a serine protease, leading to the activation of the prophenoloxidase cascade, a pathway in insects for defense against microbial infection. PMID- 10713055 TI - A deficiency of microsomal triglyceride transfer protein reduces apolipoprotein B secretion. AB - Microsomal triglyceride transfer protein (MTP) transfers lipids to apolipoprotein B (apoB) within the endoplasmic reticulum, a process that involves direct interactions between apoB and the large subunit of MTP. Recent studies with heterozygous MTP knockout mice have suggested that half-normal levels of MTP in the liver reduce apoB secretion. We hypothesized that reduced apoB secretion in the setting of half-normal MTP levels might be caused by a reduced MTP:apoB ratio in the endoplasmic reticulum, which would reduce the number of apoB-MTP interactions. If this hypothesis were true, half-normal levels of MTP might have little impact on lipoprotein secretion in the setting of half-normal levels of apoB synthesis (since the ratio of MTP to apoB would not be abnormally low) and might cause an exaggerated reduction in lipoprotein secretion in the setting of apoB overexpression (since the MTP:apoB ratio would be even lower). To test this hypothesis, we examined the effects of heterozygous MTP deficiency on apoB metabolism in the setting of normal levels of apoB synthesis, half-normal levels of apoB synthesis (heterozygous Apob deficiency), and increased levels of apoB synthesis (transgenic overexpression of human apoB). Contrary to our expectations, half-normal levels of MTP reduced the plasma apoB100 levels to the same extent ( approximately 25-35%) at each level of apoB synthesis. In addition, apoB secretion from primary hepatocytes was reduced to a comparable extent at each level of apoB synthesis. Thus, these results indicate that the concentration of MTP within the endoplasmic reticulum rather than the MTP:apoB ratio is the critical determinant of lipoprotein secretion. Finally, we found that heterozygosity for an apoB knockout mutation lowered plasma apoB100 levels more than heterozygosity for an MTP knockout allele. Consistent with that result, hepatic triglyceride accumulation was greater in heterozygous apoB knockout mice than in heterozygous MTP knockout mice. PMID- 10713056 TI - Microbial elicitors induce activation and dual phosphorylation of the Arabidopsis thaliana MAPK 6. AB - Protein kinases related to the family of mitogen-activated kinases (MAPKs) have been established as signal transduction components in a variety of processes in plants. For Arabidopsis thaliana, however, although one of the genetically best studied plant species, biochemical data on activation of mitogen-activated protein kinases are lacking. A. thaliana MAPK 6 (AtMPK6) is the Arabidopsis orthologue of a tobacco MAPK termed salicylate-induced protein kinase, which is activated by general and race-specific elicitors as well as by physical stress. Using a C terminus-specific antibody, we show that AtMPK6 is activated in elicitor-treated cell cultures of A. thaliana. Four different elicitors from bacteria, fungi, and plants lead to a rapid and transient activation of AtMPK6, indicating a conserved signaling pathway. The induction was equally rapid as medium alkalinization, one of the earliest elicitor response observed in cell cultures. A similarly rapid activation of AtMPK6 was observed in elicitor-treated leaf strips, demonstrating that recognition of the elicitors and activation of the MAPK pathway occurs also in intact plants. We demonstrate by in vivo labeling that AtMPK6 is phosphorylated on threonine and tyrosine residues in elicited cells. PMID- 10713057 TI - A role for the actin cytoskeleton in the initiation and maintenance of store mediated calcium entry in human platelets. Evidence for conformational coupling. AB - The nature of the mechanism underlying store-mediated Ca(2+) entry has been investigated in human platelets through a combination of cytoskeletal modifications. Inhibition of actin polymerization by cytochalasin D or latrunculin A had a biphasic time-dependent effect on Ca(2+) entry, showing an initial potentiation followed by inhibition of Ca(2+) entry. Moreover, addition of these agents after induction of store-mediated Ca(2+) entry inhibited the Ca(2+) influx mechanism. Jasplakinolide, which reorganizes actin filaments into a tight cortical layer adjacent to the plasma membrane, prevented activation of store-mediated Ca(2+) entry but did not modify this process after its activation. In addition, jasplakinolide prevented cytochalasin D-induced inhibition of store mediated Ca(2+) entry. Calyculin A, an inhibitor of protein serine/threonine phosphatases 1 and 2 which activates translocation of existing F-actin to the cell periphery without inducing actin polymerization, also prevented activation of store-mediated Ca(2+) entry. Finally, inhibition of vesicular transport with brefeldin A inhibited activation of store-mediated Ca(2+) entry but did not alter this mechanism once initiated. These data suggest that store-mediated Ca(2+) entry in platelets may be mediated by a reversible trafficking and coupling of the endoplasmic reticulum with the plasma membrane, which shows close parallels to the events mediating secretion. PMID- 10713058 TI - Re-evaluation of the binding of ATP to metallothionein. AB - In a recent paper Jiang et al. (Jiang, L. J., Maret, W. & Vallee, B. L. (1998) Proc. Natl. Acad. Sci. U. S. A. 95, 9146-9149) reported that metallothionein interacts with adenosine triphosphate (ATP) to form a 1:1 complex with a dissociation constant of K(d) = 176 +/- 33 microM at pH 7.4. In an effort to characterize further this interaction using nuclear magnetic resonance spectroscopy, titration calorimetry, gel-filtration chromatography, affinity chromatography, and ultrafiltration, we were unable to find any evidence for the binding of ATP to metallothionein. PMID- 10713059 TI - Interaction of von Willebrand factor domain A1 with platelet glycoprotein Ibalpha (1-289). Slow intrinsic binding kinetics mediate rapid platelet adhesion. AB - We investigated the crucial hemostatic interaction between von Willebrand factor (VWF) and platelet glycoprotein (GP) Ibalpha. Recombinant VWF A1 domain (residues Glu(497)-Pro(705) of VWF) bound stoichiometrically to a GPIbalpha-calmodulin fusion protein (residues His(1)-Val(289) of GPIbalpha; GPIbalpha-CaM) immobilized on W-7-agarose with a K(d) of 3.3 microM. The variant VWF A1(R545A) bound to GPIbalpha-CaM 20-fold more tightly, mainly because the association rate constant k(on) increased from 1,100 to 8,800 M(-1) s(-1). The GPIbalpha mutations G233V and M239V cause platelet-type pseudo-von Willebrand disease, and VWF A1 bound to GPIbalpha(G233V)-CaM and GPIbalpha(M239V)-CaM with a K(d) of 1.0 and 0.63 microM, respectively. The increased affinity of VWF A1 for GPIbalpha(M239V)-CaM was explained by an increase in k(on) to 4,500 M(-1) s(-1). GPIbalpha-CaM bound with similar affinity to recombinant VWF A1, to multimeric plasma VWF, and to a fragment of dispase-digested plasma VWF (residues Leu(480)/Val(481)-Gly(718)). VWF A1 and A1(R545A) bound to platelets with affinities and rate constants similar to those for binding to GPIbalpha-CaM, and botrocetin had the expected positively cooperative effect on the binding of VWF A1 to GPIbalpha-CaM. Therefore, allosteric regulation by botrocetin of VWF A1 binding to GPIbalpha, and the increased binding affinity caused by mutations in VWF or GPIbalpha, are reproduced by isolated structural domains. The substantial increase in k(on) caused by mutations in either A1 or GPIbalpha suggests that productive interaction requires rate-limiting conformational changes in both binding sites. The exceptionally slow k(on) and k(off) provide important new constraints on models for rapid platelet tethering at high wall shear rates. PMID- 10713060 TI - Identification and functional characterization of thioredoxin from Trypanosoma brucei brucei. AB - Trypanosomes and Leishmania, the causative agents of several tropical diseases, lack the glutathione/glutathione reductase system but have trypanothione/trypanothione reductase instead. The uniqueness of this thiol metabolism and the failure to detect thioredoxin reductases in these parasites have led to the suggestion that these protozoa lack a thioredoxin system. As presented here, this is not the case. A gene encoding thioredoxin has been cloned from Trypanosoma brucei, the causative agent of African sleeping sickness. The single copy gene, which encodes a protein of 107 amino acid residues, is expressed in all developmental stages of the parasite. The deduced protein sequence is 56% identical with a putative thioredoxin revealed by the genome project of Leishmania major. The relationship to other thioredoxins is low. T. brucei thioredoxin is unusual in having a calculated pI value of 8.5. The gene has been overexpressed in Escherichia coli. The recombinant protein is a substrate of human thioredoxin reductase with a K(m) value of 6 microM but is not reduced by trypanothione reductase. T. brucei thioredoxin catalyzes the reduction of insulin by dithioerythritol, and functions as an electron donor for T. brucei ribonucleotide reductase. The parasite protein is the first classical thioredoxin of the order Kinetoplastida characterized so far. PMID- 10713061 TI - Cloning and characterization of two promoters for the human calcium-sensing receptor (CaSR) and changes of CaSR expression in parathyroid adenomas. AB - Histological analyses showed that expression of the parathyroid calcium-sensing receptor (CaSR) is decreased in parathyroid adenomas. Because reduced expression of CaSR may result in insufficient suppression of parathyroid hormone secretion, the elucidation of regulatory mechanisms of CaSR expression is indispensable for understanding the pathogenesis of parathyroid adenomas. Two cDNA clones for human CaSR with different 5'-untranslated regions have been isolated. However, the structure of the promoter region of human CaSR and the mechanisms of production of multiple CaSR mRNAs are unknown. We have cloned promoter regions of human CaSR by screening a genomic library. The human CaSR gene has two promoters and two 5' untranslated exons (exons 1A and 1B), and alternative usage of these exons leads to production of multiple CaSR mRNAs. The upstream promoter has TATA and CAAT boxes, and the downstream promoter is GC-rich. Northern blot analysis showed that expression levels of exon 1A in parathyroid adenomas are significantly less than those in normal glands. However, expression of exon 1B was not different between adenomas and normal glands. Thus, specific reduction of the transcript driven by the upstream promoter was observed in parathyroid adenomas. Further analyses of factors that modulate the activity of the upstream promoter are necessary to clarify the pathogenesis of parathyroid adenomas. PMID- 10713062 TI - Nerve growth factor activation of nuclear factor kappaB through its p75 receptor is an anti-apoptotic signal in RN22 schwannoma cells. AB - Recent evidence indicates that nerve growth factor (NGF) produces its effects through signaling contributions from both TrkA and the p75 receptor. In contrast to its trophic actions through TrkA, NGF binding to p75 has been shown to activate programmed cell death through a mechanism involving the stress kinase JNK. However, this receptor also activates nuclear factor kappaB (NF-kappaB), the role of which has yet to be determined. We investigated the function of p75 mediated NF-kappaB stimulation in regulating cell survival in the rat schwannoma cell line RN22, which expresses p75, but not TrkA. Gel shift assays demonstrated activation of NF-kappaB in response to NGF within 30 min and lasting at least 4 h. NGF also stimulated JNK in the cells (detected by in vitro kinase assays) with a similar time course. Preventing activation of NF-kappaB with the specific inhibitor SN50 resulted in NGF-induced cell loss. Similarly, transfection of the cells with a mutant form of the endogenous NF-kappaB inhibitor (IkappaBalphaDeltaN), which cannot be degraded and therefore remains bound to NF kappaB, preventing its activation, resulted in a significant increase in the number of apoptotic cells following NGF treatment. These results suggest that NGF activation of NF-kappaB through the p75 receptor promotes survival, counterbalancing the pro-apoptotic signal. PMID- 10713063 TI - Recombinant toxins that bind to the urokinase receptor are cytotoxic without requiring binding to the alpha(2)-macroglobulin receptor. AB - The alpha(2-)macroglobulin receptor (alpha(2)MR) has been reported to mediate the internalization of the urokinase plasminogen activator receptor (uPAR) via ligand binding to both receptors. To target malignant uPAR-expressing cells and to determine whether uPAR can internalize without ligand binding to alpha(2)MR, we engineered two recombinant toxins, ATF-PE38 and ATF-PE38KDEL. Each consists of the amino-terminal fragment (ATF) of human urokinase and a truncated form of Pseudomonas exotoxin (PE) devoid of domain Ia, which binds alpha(2)MR. ATF-PE38 and ATF-PE38KDEL were cytotoxic toward malignant uPAR-bearing cells, with IC(50) values as low as 0.02 ng/ml (0.3 pM). Cytotoxicity could be blocked using either recombinant urokinase or free ATF, indicating that the cytotoxicity of the recombinant toxins was specific. Radiolabeled ATF-PE38 had high affinity for uPAR (K(d) = 0.4-8 nM) on a variety of different malignant cell types and internalized at a rate similar to that of ATF. The cytotoxicity was not diminished by receptor associated protein, which binds and shields the alpha(2)MR from other proteins, or by incubation with phorbol myristate acetate, which is known to decrease the number of alpha(2)MRs in U937 cells or by antibodies to alpha(2)MR. Therefore, these recombinant toxins appear to internalize via uPAR without association with the alpha(2)MR. PMID- 10713064 TI - Involvement of protein kinase C delta (PKCdelta) in phorbol ester-induced apoptosis in LNCaP prostate cancer cells. Lack of proteolytic cleavage of PKCdelta. AB - Phorbol esters, the activators of protein kinase C (PKC), induce apoptosis in androgen-sensitive LNCaP prostate cancer cells. The role of individual PKC isozymes as mediators of this effect has not been thoroughly examined to date. To study the involvement of the novel isozyme PKCdelta, we used a replication deficient adenovirus (PKCdeltaAdV), which allowed for a tightly controlled expression of PKCdelta in LNCaP cells. A significant reduction in cell number was observed after infection of LNCaP cells with PKCdeltaAdV. Overexpression of PKCdelta markedly enhanced the apoptotic effect of phorbol 12-myristate 13 acetate in LNCaP cells. PKCdelta-mediated apoptosis was substantially reduced by the pan-caspase inhibitor z-VAD and by Bcl-2 overexpression. Importantly, and contrary to other cell types, PKCdelta-mediated apoptosis does not involve its proteolytic cleavage by caspase-3, suggesting that allosteric activation of PKCdelta is sufficient to trigger apoptosis in LNCaP cells. In addition, phorbol ester-induced apoptosis was blocked by a kinase-deficient mutant of PKCdelta, supporting the concept that PKCdelta plays an important role in the regulation of apoptotic cell death in LNCaP prostate cancer cells. PMID- 10713065 TI - Glycogen synthase kinase-3beta facilitates staurosporine- and heat shock-induced apoptosis. Protection by lithium. AB - The potential role of glycogen synthase kinase-3beta in modulating apoptosis was examined in human SH-SY5Y neuroblastoma cells. Staurosporine treatment caused time- and concentration-dependent increases in the activities of caspase-3 and caspase-9 but not caspase-1, increased proteolysis of poly(ADP-ribose) polymerase, and induced morphological changes consistent with apoptosis. Overexpression of glycogen synthase kinase-3beta to levels 3.5 times that in control cells did not alter basal indices of apoptosis but potentiated staurosporine-induced activation of caspase-3, caspase-9, proteolysis of poly(ADP ribose) polymerase, and morphological changes indicative of apoptosis. Inhibition of glycogen synthase kinase-3beta by lithium attenuated the enhanced staurosporine-induced activation of caspase-3 in cells overexpressing glycogen synthase kinase-3beta. In cells subjected to heat shock, caspase-3 activity was more than three times greater in glycogen synthase kinase-3beta-transfected than control cells, and this potentiated response was inhibited by lithium treatment. Thus, glycogen synthase kinase-3beta facilitated apoptosis induced by two experimental paradigms. These findings indicate that glycogen synthase kinase 3beta may contribute to pro-apoptotic-signaling activity, that inhibition of glycogen synthase kinase-3beta can contribute to anti-apoptotic-signaling mechanisms, and that the neuroprotective actions of lithium may be due in part to its inhibitory modulation of glycogen synthase kinase-3beta. PMID- 10713066 TI - Novel interaction between the transcription factor CHOP (GADD153) and the ribosomal protein FTE/S3a modulates erythropoiesis. AB - The transcription factor CHOP (GADD153) heterodimerizes with other C/EBP family members, especially C/EBPbeta, thus preventing their homodimerization and binding to DNA sequences specific for the homodimers. Some CHOP-C/EBP heterodimers apparently bind to alternative DNA sequence and thereby regulate the transcription of other genes. Recently, we demonstrated that CHOP is up-regulated during certain stages of erythroid differentiation and that ectopic overexpression of CHOP enhances this process (Coutts, M., Cui, K., Davis, K. L., Keutzer, J. C., and Sytkowski, A. J. (1999) Blood 93, 3369-3378). In the present study, we report that CHOP also interacts with another non-C/EBP protein designated v-fos transformation effector (FTE) (Kho, C. J., and Zarbl, H. (1992) Proc. Natl. Acad. Sci. U. S. A. 89, 2200-2204), which is identical to ribosomal protein S3a (Metspalu, A., Rebane, A., Hoth, S., Pooga, M., Stahl, J. , and Kruppa, J. (1992) Gene (Amst.) 119, 313-316). Bacterially expressed His-CHOP and in vitro translated (35)S-labeled FTE/S3a-Gal4 fusion protein co immunoprecipitated using anti-CHOP antibodies, and both anti-CHOP and anti FTE/S3a antibodies co-immunoprecipitated CHOP and FTE/S3a from lysates of Rauscher murine erythroleukemia cells overexpressing both proteins. The in vivo interaction of CHOP and FTE/S3a was also demonstrated in cells overexpressing FTE/S3a but with endogenous expression levels of CHOP. Western blot analysis demonstrated co-localization of CHOP and FTE/S3a in both the cytosol and the nuclei of non-transfected cells. Overexpression of FTE/S3a inhibited differentiation of Rauscher cells induced either by erythropoietin or by dimethyl sulfoxide. This inhibition was reversed partially by simultaneous overexpression of CHOP or of antisense fte/S3a. FTE/S3a appears to be a bifunctional ribosomal protein that regulates CHOP and, hence, C/EBP function during erythropoiesis. PMID- 10713067 TI - Tsc3p is an 80-amino acid protein associated with serine palmitoyltransferase and required for optimal enzyme activity. AB - Serine palmitoyltransferase catalyzes the first step of sphingolipid synthesis, condensation of serine and palmitoyl CoA to form the long chain base 3 ketosphinganine. The LCB1/TSC2 and LCB2/TSC1 genes encode homologous proteins of the alpha-oxoamine synthase family required for serine palmitoyltransferase activity. The other alpha-oxoamine synthases are soluble homodimers, but serine palmitoyltransferase is a membrane-associated enzyme composed of at least two subunits, Lcb1p and Lcb2p. Here, we report the characterization of a third gene, TSC3, required for optimal 3-ketosphinganine synthesis in Saccharomyces cerevisiae. S. cerevisiae cells lacking the TSC3 gene have a temperature sensitive lethal phenotype that is reversed by supplying 3-ketosphinganine, dihydrosphingosine, or phytosphingosine in the growth medium. The tsc3 mutant cells have severely reduced serine palmitoyltransferase activity. The TSC3 gene encodes a novel 80-amino acid protein with a predominantly hydrophilic amino terminal half and a hydrophobic carboxyl terminus that is membrane-associated. Tsc3p coimmunoprecipitates with Lcb1p and/or Lcb2p but does not bind as tightly as Lcb1p and Lcb2p bind to each other. Lcb1p and Lcb2p remain tightly associated with each other and localize to the membrane in cells lacking Tsc3p. However, Lcb2p is unstable in cells lacking Lcb1p and vice versa. PMID- 10713068 TI - Reconstitution of angiotensin receptor mRNA down-regulation in vascular smooth muscle. Post-transcriptional control by protein kinase a but not mitogenic signaling directed by the 5'-untranslated region. AB - Cell surface receptor activation generally leads to changes in mRNA abundance, which may involve regulatory targets in processes working at the post transcriptional level. Many types of agonists down-regulate vascular smooth muscle angiotensin receptor (AT(1)-R) gene expression, but it is unclear which of these activate post-transcriptional mechanisms. To reconstitute faithfully the normal AT(1)-R mRNA regulatory environment, tetracycline-suppressible promoters drive highly accurate recombinant AT(1)-R mRNA mimics in vascular smooth muscle cells that co-express an endogenous AT(1)-R mRNA. Down-regulation of the latter occurs shortly after stimulating mitogenic receptors or by using forskolin, but only cAMP signaling reduces expression of the recombinant AT(1)-R mRNA. Transcription of the recombinant mRNA is unaffected by cAMP signaling. Deletions of the AT(1)-R mRNA 3'-untranslated region do not impair cAMP-mediated down regulation. Both loss of function and gain of function mutants show the response is mediated by the 5'-untranslated region. These observations provide the first direct functional evidence for modulation of vascular AT(1)-R gene expression by a mechanism involving a protein kinase A-regulated post-transcriptional process. PMID- 10713069 TI - Two distinct nucleosome alterations characterize chromatin remodeling at the Saccharomyces cerevisiae ADH2 promoter. AB - Glucose depletion derepresses the Saccharomyces cerevisiae ADH2 gene; this metabolic change is accompanied by chromatin structural modifications in the promoter region. We show that the ADR6/SWI1 gene is not necessary for derepression of the wild type chromosomal ADH2, whereas the transcription factor Adr1p, which regulates several S. cerevisiae functions, plays a major role in driving nucleosome reconfiguration and ADH2 expression. When we tested the effect of individual domains of the regulatory protein Adr1p on the chromatin structure of ADH2, a remodeling consisting of at least two steps was observed. Adr1p derivatives were analyzed in derepressing conditions, showing that the Adr1p DNA binding domain alone causes an alteration in chromatin organization in the absence of transcription. This alteration differs from the remodeling observed in the presence of the Adr1p activation domain when the promoter is transcriptionally active. PMID- 10713070 TI - The structure of the nuclear factor-kappaB protein-DNA complex varies with DNA binding site sequence. AB - Transcriptional regulation of many immune responsive genes is under the control of the transcription factor NF-kappaB. This factor is found in cells as a dimer which can contain any two members of the Rel family of proteins (p50, p65, p52, c Rel, and RelB). The different dimers show distinct preferences for DNA-binding site sequences. To understand the relationship between the DNA binding properties of the dimer forms and transcriptional activation, the physical properties of the complexes of p50 and p65 with DNA have been analyzed. Comparison of apparent DNA binding affinity showed differences in selectivity of DNA-binding site sequence. The ionic strength dependence of apparent binding affinity has shown that the number of ionic interactions in the protein-DNA complex depends on the DNA binding site sequence and the dimer form, which are consistent with changes in the structure of the protein-DNA complex. Using a fluorescent technique to measure DNA structure changes, protein binding does not appear to alter the structure of the DNA-binding site within the limits of detection. These results are consistent with a change in protein structure that may result in activation differences due to alternative interactions with other transcription proteins. PMID- 10713071 TI - CCC1 suppresses mitochondrial damage in the yeast model of Friedreich's ataxia by limiting mitochondrial iron accumulation. AB - Deletion of YFH1 in Saccharomyces cerevisiae leads to a loss of respiratory competence due to excessive mitochondrial iron accumulation. A suppressor screen identified a gene, CCC1, that maintained respiratory function in a Deltayfh1 yeast strain regardless of extracellular iron concentration. CCC1 expression prevented excessive mitochondrial iron accumulation by limiting mitochondrial iron uptake rather than by increasing mitochondrial iron egress. Expression of CCC1 did not result in sequestration of iron in membranous compartments or cellular iron export. CCC1 expression in wild type cells resulted in increased expression of the high affinity iron transport system composed of FET3 and FTR1, suggesting that intracellular iron is not sensed by the iron-dependent transcription factor Aft1p. Introduction of AFT1(up), a constitutive allele of the iron transcription factor, AFT1, that also leads to increased high affinity iron transport did not prevent Deltayfh1 cells from becoming respiratory incompetent. Although the mechanism by which CCC1 expression affects cytosolic iron is not known, the data suggest that excessive mitochondrial iron accumulation only occurs when cytosolic free iron levels are high. PMID- 10713072 TI - G(i)-dependent activation of c-Jun N-terminal kinase in human embryonal kidney 293 cells. AB - Heterotrimeric G proteins stimulate the activities of two stress-activated protein kinases, c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase in mammalian cells. In this study, we examined whether alpha subunits of G(i) family activate JNK using transient expression system in human embryonal kidney 293 cells. Constitutively activated mutants of Galpha(i1), Galpha(i2), and Galpha(i3) increased JNK activity. In contrast, constitutively activated Galpha(o) and Galpha(z) mutants did not stimulate JNK activity. To examine the mechanism of JNK activation by Galpha(i), kinase-deficient mutants of mitogen activated protein kinase kinase 4 (MKK4) and 7 (MKK7), which are known to be JNK activators, were transfected into the cells. However, Galpha(i)-induced JNK activation was not blocked effectively by kinase-deficient MKK4 and MKK7. In addition, activated Galpha(i) mutant failed to stimulate MKK4 and MKK7 activities. Furthermore, JNK activation by Galpha(i) was inhibited by dominant negative Rho and Cdc42 and tyrosine kinase inhibitors, but not dominant-negative Rac and phosphatidylinositol 3-kinase inhibitors. These results indicate that Galpha(i) regulates JNK activity dependent on small GTPases Rho and Cdc42 and on tyrosine kinase but not on MKK4 and MKK7. PMID- 10713073 TI - Purification and characterization of a magnesium-dependent neutral sphingomyelinase from bovine brain. AB - The magnesium-dependent, plasma membrane-associated neutral sphingomyelinase (N SMase) catalyzes hydrolysis of membrane sphingomyelin to form ceramide, a lipid signaling molecule implied in intracellular signaling. We report here the biochemical purification to apparent homogeneity of N-SMase from bovine brain. Proteins from Nonidet P-40 extracts of brain membranes were subjected to four purification steps yielding a N-SMase preparation that exhibited a specific enzymatic activity 23,330-fold increased over the brain homogenate. When analyzed by two-dimensional gel electrophoresis, the purified enzyme presented as two major protein species of 46 and 97 kDa, respectively. Matrix-assisted laser desorption/ionization-mass spectrometry analysis of tryptic peptides revealed at least partial identity of these two proteins. Amino acid sequencing of tryptic peptides showed no apparent homologies of bovine N-SMase to any known protein. Peptide-specific antibodies recognized a single 97-kDa protein in Western blot analysis of cell lysates. The purified enzyme displayed a K(m) of 40 microM for sphingomyelin with an optimal activity at pH 7-8. Bovine brain N-SMase was strictly dependent on Mg(2+), whereas Zn(2+) and Ca(2+) proved inhibitory. The highly purified bovine N-SMase was effectively blocked by glutathione and scyphostatin. Scyphostatin proved to be a potent inhibitor of N-SMase with 95% inhibition observed at 20 microM scyphostatin. The results of this study define a N-SMase that fulfills the biochemical and functional criteria characteristic of the tumor necrosis factor-responsive membrane-bound N-SMase. PMID- 10713074 TI - A eukaryotic SWI2/SNF2 domain, an exquisite detector of double-stranded to single stranded DNA transition elements. AB - Many members of the SWI2/SNF2 family of adenosine triphosphatases participate in the assembly/disassembly of multiprotein complexes involved in the DNA metabolic processes of transcription, recombination, repair, and chromatin remodeling. The DNA molecule serves as an essential effector or catalyst for most of the members of this particular class of proteins, and the structure of the DNA may be more important than the nucleotide sequence. Inspection of the DNA structure at sites where multiprotein complexes are assembled/disassembled for these various DNA metabolic processes reveals the presence of a common element: a double-stranded to single-stranded transition region. We now show that this DNA element is crucial for the ATP hydrolytic function of an SWI2/SNF2 family member: DNA dependent ATPase A. We further demonstrate that a domain containing the seven helicase-related motifs that are common to the SWI2/SNF2 family of proteins mediates the interaction with the DNA, yielding specific DNA structural recognition. This study forms a primary step toward understanding the physico biochemical nature of the interaction between a particular class of DNA-dependent ATPase and their DNA effectors. Furthermore, this study provides a foundation for development of mechanisms to specifically target this class of DNA-dependent ATPases. PMID- 10713075 TI - pH-induced intramolecular electron transfer between the iron-sulfur protein and cytochrome c(1) in bovine cytochrome bc(1) complex. AB - Structural analysis of the bc(1) complex suggests that the extra membrane domain of iron-sulfur protein (ISP) undergoes substantial movement during the catalytic cycle. Binding of Qo site inhibitors to this complex affects the mobility of ISP. Taking advantage of the difference in the pH dependence of the redox midpoint potentials of cytochrome c(1) and ISP, we have measured electron transfer between the [2Fe-2S] cluster and heme c(1) in native and inhibitor-treated partially reduced cytochrome bc(1) complexes. The rate of the pH-induced cytochrome c(1) reduction can be estimated by conventional stopped-flow techniques (t1/2, 1-2 ms), whereas the rate of cytochrome c(1) oxidation is too high for stopped-flow measurement. These results suggest that oxidized ISP has a higher mobility than reduced ISP and that the movement of reduced ISP may require an energy input from another component. In the 5-n-undecyl-6-hydroxy-4,7-dioxobenzothiazole (UHDBT) inhibited complex, the rate of cytochrome c(1) reduction is greatly decreased to a t1/2 of approximately 2.8 s. An even lower rate is observed with the stigmatellin-treated complex. These results support the idea that UHDBT and stigmatellin arrest the [2Fe-2S] cluster at a fixed position, 31 A from heme c(1), making electron transfer very slow. PMID- 10713076 TI - Interaction of the corepressor Alien with DAX-1 is abrogated by mutations of DAX 1 involved in adrenal hypoplasia congenita. AB - DAX-1 is an unusual member of the nuclear hormone receptor (NHR) superfamily. Lack of DAX-1-mediated silencing leads to adrenal hypoplasia congenita and hypogonadotropic hypogonadism. Gene silencing through NHRs such as the thyroid hormone receptor (TR) is mediated by corepressors. We have previously characterized a novel corepressor, termed Alien, which interacts with TR and the ecdysone receptor but not with the retinoic acid receptors RAR or RXR. Here, we show that DAX-1 interacts with the corepressor Alien but not with the corepressor SMRT. This interaction is mediated by the DAX-1-silencing domain. Naturally occurring mutants of the DAX-1 gene fail to interact with Alien and have lost silencing function. Because the silencing domain of DAX-1 is unusual for NHRs, we mapped the interaction of Alien with DAX-1 and with TR. We show that Alien exhibits different binding characteristics to DAX-1 and TR. Furthermore, Northern experiments demonstrate that Alien is expressed in the adrenal gland and testis in tissues where DAX-1 is specifically expressed. Interestingly, a novel adrenal gland-specific mRNA of Alien was discovered. Thus, the impairment of Alien binding seems to play an important role in the pathogenesis mediated by DAX-1 mutants. PMID- 10713077 TI - Role of c-jun expression increased by heat shock- and ceramide-activated caspase 3 in HL-60 cell apoptosis. Possible involvement of ceramide in heat shock-induced apoptosis. AB - Ceramide has emerged as a lipid mediator in apoptosis induced by a variety of stresses. As we previously showed that the activation of AP-1, a nuclear transcription factor was indispensable to ceramide-induced apoptosis in human leukemia HL-60 cells (Sawai, H., Okazaki, T., Yamamoto, H., Okano, H., Takeda, Y., Tashima, M., Sawada, H., Okuma, M., Ishikura, H., Umehara, H., and Domae, N. (1995) J. Biol. Chem. 270, 27326-27331), the role and mechanism of heat shock (HS)-increased c-jun expression in apoptosis was here investigated. HS increased morphological changes compatible with apoptosis in human leukemia HL-60 cells, and induced ceramide generation and sphingomyelin hydrolysis with an increase of neutral magnesium-dependent sphingomyelinase activity. When HS failed to induce apoptosis in HS-resistant HL-60 cells, ceramide generation was not detected, suggesting that ceramide was involved in downstream signals required for HS induced apoptosis. Both HS and N-acetylsphingosine (C(2)-ceramide) increased the expression of c-jun/c-fos mRNAs with the peak 2 h after treatment. When we examined whether the inhibition of c-jun expression by its antisense oligodeoxynucleotides (AS) blocked HS- or C(2)-ceramide-induced apoptosis, AS of c-jun gene inhibited apoptotic morphological changes and DNA fragmentation whereas did not sense oligodeoxynucleotides. Moreover, a synthetic tetrapeptide, acetyl-Asp-Met-Gln-Asp-aldehyde (DMQD-CHO), which inhibited the formation of active form of caspase-3 more efficiently than those of caspase-4, -6, -7, and 8, blocked both caspase-3 like activity, c-jun expression and apoptosis induced by HS or C(2)-ceramide, although DMQD-CHO did not affect HS-induced ceramide generation. These results suggested that the ceramide was generated through sphingomyelin hydrolysis by HS-activated neutral, magnesium-dependent sphingomyelinase and that subsequent c-jun expression through activation of caspase-3 played a role in HS-induced HL-60 cell apoptosis. PMID- 10713078 TI - Identification of key functional amino acids of the mouse fertilin beta (ADAM2) disintegrin loop for cell-cell adhesion during fertilization. AB - Fertilin beta (also known as ADAM2) is a cell adhesion molecule on the surface of mammalian sperm that participates in sperm-egg membrane binding. Fertilin beta is a member of the molecular family known as ADAMs or MDCs. These proteins have a disintegrin domain with homology to integrin ligands found in snake venoms; several of these snake proteins have an RGD tripeptide presented on an extended "disintegrin loop." However, fertilin beta lacks an RGD tripeptide and instead has the consensus sequence X(D/E)ECD (QDECD in mouse fertilin beta) in its putative disintegrin loop, and there is controversy over which amino acids comprise the active site of the fertilin beta disintegrin loop. We have used point-mutated versions of the sequence AQDECDVT and two bioassays to identify the key functional amino acids of this sequence from the mouse fertilin beta disintegrin domain. Amino acid substitutions for the terminal aspartic acid residue of the QDECD sequence result in dramatically reduced activities in the two assays for protein function, implicating the terminal aspartic acid residue as critical for protein function. Substitutions for the glutamic acid and the cysteine residues in the QDECD sequence result in slight reductions in activity, whereas substitution of the first aspartic acid has virtually no effect. These data suggest that the conserved ECD sequence of the mouse fertilin beta disintegrin loop, especially the terminal D residue, contributes more to the protein's activity than does the QDE sequence that aligns with the RGD tripeptide in other disintegrins. PMID- 10713079 TI - O(2) sensing by airway chemoreceptor-derived cells. Protein kinase c activation reveals functional evidence for involvement of NADPH oxidase. AB - Accumulating evidence suggests that neuroepithelial bodies are airway O(2) sensors. Recently, we have established the H-146 small cell lung carcinoma line as a suitable model to study the biochemical basis of neuroepithelial body cell chemotransduction. Here we explore the possibility that hypoxic modulation of K(+) channels is intimately linked to activity of NADPH oxidase. Graded hypoxia caused graded inhibition of whole cell K(+) currents, which correlated well with membrane depolarization. Pretreatment with the phorbol ester, 12-O-tetradecanoyl (TPA), inhibited K(+) currents at all potentials. Although 4alpha-phorbol 12,13 didecanoate and TPA in the presence of bisindolylmaleimide were also able to depress K(+) currents, only TPA could significantly ameliorate hypoxic depression of these currents. Thus, protein kinase C (PKC) activation modulates the sensitivity of these cells to changes in pO(2). Furthermore, because the addition of H(2)O(2), a downstream product of NADPH oxidase, could only activate K(+) currents during hypoxia (when endogenous H(2)O(2) production is suppressed), it appears likely that PKC modulates the affinity of NADPH oxidase for O(2) potentially via phosphorylation of the p47(phox) subunit, which is present in these cells. These data show that PKC is an important regulator of the O(2) transduction pathway and suggests that NADPH oxidase represents a significant component of the airway O(2) sensor. PMID- 10713080 TI - Activation of primary human monocytes by the oxidized form of alpha1-antitrypsin. AB - The oxidation of methionine residues in many proteins, including the serine proteinase inhibitor alpha1-antitrypsin (AAT), can result in functional inactivation. In this study we investigated the pro-inflammatory properties of oxidized AAT (oxAAT), specifically its ability to activate human monocytes in culture. Monocytes stimulated with oxAAT at concentrations up to 0.2 mg/ml for 24 h showed significant elevation in monocyte chemoattractant protein-1, cytokine interleukin-6, and tumor necrosis factor-alpha expression and increased NADPH oxidase activity. Monocytes activated with oxAAT showed surprising effects on lipid metabolism. Expression of low density lipoprotein (LDL) receptors increased by up to 76% compared with controls but was not accompanied by any changes in (125)I-labeled LDL binding and, paradoxically, decreased LDL uptake, degradation, and intracellular cholesterol synthesis. oxAAT also down-regulated the scavenger receptor CD36, which takes up and is up-regulated by oxidized LDL and is down regulated by cholesterol efflux. As a by-product of oxidative events accompanying inflammation, oxAAT has multiple effects on cytokine expression, generation of reactive oxygen species, and on intracellular lipid metabolism. The up-regulation of monocyte-derived reactive oxygen by oxAAT could potentially result in self amplification of AAT oxidation and, thereby, the other effects deriving from it. This implies that there are as yet unidentified regulatory processes that control this cycle. PMID- 10713081 TI - Design of peptides with high affinities for heparin and endothelial cell proteoglycans. AB - Proteoglycan-binding peptides were designed based on consensus sequences in heparin-binding proteins: XBBXBX and XBBBXXBX, where X and B are hydropathic and basic residues, respectively. Initial peptide constructs included (AKKARA)(n) and (ARKKAAKA)(n) (n = 1-6). Affinity coelectrophoresis revealed that low M(r) peptides (600-1,300) had no affinities for low M(r) heparin, but higher M(r) peptides (2,000-3,500) exhibited significant affinities (K(d) congruent with 50 150 nM), which increased with peptide M(r). Affinity was strongest when sequence arrays were contiguous and alanines and arginines occupied hydropathic and basic positions, but inclusion of prolines was disruptive. A peptide including a single consensus sequence of the serglycin proteoglycan core protein bound heparin strongly (K(d) congruent with 200 nM), likely owing to dimerization through cysteine-cysteine linkages. Circular dichroism showed that high affinity heparin binding peptides converted from a charged coil to an alpha-helix upon heparin addition, whereas weak heparin-binding peptides did not. Higher M(r) peptides exhibited high affinities for total endothelial cell proteoglycans (K(d) congruent with 300 nM), and approximately 4-fold weaker affinities for their free glycosaminoglycan chains. Thus, peptides including concatamers of heparin-binding consensus sequences may exhibit strong affinities for heparin and proteoglycans. Such peptides may be applicable in promoting cell-substratum adhesion or in the design of drugs targeted to proteoglycan-containing cell surfaces and extracellular matrices. PMID- 10713082 TI - Structural studies of lacUV5-RNA polymerase interactions in vitro. Ethylation interference and missing nucleoside analysis. AB - Substantial effort has been made to investigate the interactions that the Escherichia coli RNA polymerase makes with promoter DNA during transcription initiation. The lacUV5 promoter has been the object of many of these studies, and to date, an incredible wealth of information exists on how RNA polymerase interacts with this promoter. We have sought to expand current knowledge by the use of two chemical interference protocols, phosphate ethylation and missing nucleoside. We have added to existing information with the identification of additional phosphates, for example, at the start site of the template strand that, when ethylated, perturb the binding of RNA polymerase. We have also discovered a number of positions, most remarkably -37 to -34 of the nontemplate strand, where nucleoside loss decreases binding. Finally, we have discovered positions of ethylation and/or nucleoside loss that can stimulate binding. In particular, missing nucleosides and phosphate ethylation near the transcription start site enhance RNA polymerase binding. PMID- 10713083 TI - The antiactivator TraM interferes with the autoinducer-dependent binding of TraR to DNA by interacting with the C-terminal region of the quorum-sensing activator. AB - Conjugal transfer of Agrobacterium tumefaciens Ti plasmids is regulated by quorum sensing via the transcriptional activator TraR and the acyl-homoserine lactone Agrobacterium autoinducer (AAI). Unique to this system, the activity of TraR is negatively modulated by an antiactivator called TraM. Analyses from yeast two hybrid studies suggest that TraM directly interacts with the activator, but the conditions under which these components interact and the region of TraR responsible for this interaction are not known. Induction of traM in a strain in which TraR was activating transcription of a reporter system led to rapid cessation of gene expression. As assessed by a genetic assay that measures AAI dependent DNA binding, TraM inhibited TraR function before and after the transcription factor had bound to its DNA recognition site. Consistent with this observation, in gel retardation assays, purified TraM abolished the DNA binding activity of TraR in a concentration-dependent manner. Such inhibition occurred independent of the order of addition of the reactants. As assessed by far Western analyses TraM interacts with TraR by directly binding the activator. TraM in its native form interacted with native TraR and also with heat-treated TraR but only when SDS was included with the denatured protein. TraM interacted with TraR on blots prepared with total lysates of cells grown in the presence and absence of AAI. Far Western analysis of N- and C-terminal deletion mutants localized a domain of TraR contributing to TraM binding to the C-terminal portion of the activator protein. Random mutagenesis by hydroxylamine treatment and error-prone polymerase chain reaction identified several residues in this region of TraR important for interacting with TraM as well as for transcriptional activation or/and DNA binding. We conclude that TraM inhibits TraR by binding to the activator at a domain within or close to the helix-turn-helix motif located at the C terminus of the protein. PMID- 10713084 TI - PRMT1 is the predominant type I protein arginine methyltransferase in mammalian cells. AB - Type I protein arginine methyltransferases catalyze the formation of asymmetric omega-N(G),N(G)-dimethylarginine residues by transferring methyl groups from S adenosyl-L-methionine to guanidino groups of arginine residues in a variety of eucaryotic proteins. The predominant type I enzyme activity is found in mammalian cells as a high molecular weight complex (300-400 kDa). In a previous study, this protein arginine methyltransferase activity was identified as an additional activity of 10-formyltetrahydrofolate dehydrogenase (FDH) protein. However, immunodepletion of FDH activity in RAT1 cells and in murine tissue extracts with antibody to FDH does not diminish type I methyltransferase activity toward the methyl-accepting substrates glutathione S-transferase fibrillarin glycine arginine domain fusion protein or heterogeneous nuclear ribonucleoprotein A1. Similarly, immunodepletion with anti-FDH antibody does not remove the endogenous methylating activity for hypomethylated proteins present in extracts from adenosine dialdehyde-treated RAT1 cells. In contrast, anti-PRMT1 antibody can remove PRMT1 activity from RAT1 extracts, murine tissue extracts, and purified rat liver FDH preparations. Tissue extracts from FDH(+/+), FDH(+/-), and FDH(-/-) mice have similar protein arginine methyltransferase activities but high, intermediate, and undetectable FDH activities, respectively. Recombinant glutathione S-transferase-PRMT1, but not purified FDH, can be cross-linked to the methyl-donor substrate S-adenosyl-L-methionine. We conclude that PRMT1 contributes the major type I protein arginine methyltransferase enzyme activity present in mammalian cells and tissues. PMID- 10713085 TI - Structural analysis of murine zona pellucida glycans. Evidence for the expression of core 2-type O-glycans and the Sd(a) antigen. AB - Murine sperm initiate fertilization by binding to specific oligosaccharides linked to the zona pellucida, the specialized matrix coating the egg. Biophysical analyses have revealed the presence of both high mannose and complex-type N glycans in murine zona pellucida. The predominant high mannose-type glycan had the composition Man(5)GlcNAc(2), but larger oligosaccharides of this type were also detected. Biantennary, triantennary, and tetraantennary complex-type N glycans were found to be terminated with the following antennae: Galbeta1 4GlcNAc, NeuAcalpha2-3Galbeta1-4GlcNAc, NeuGcalpha2-3Galbeta1-4GlcNAc, the Sd(a) antigen (NeuAcalpha2-3[GalNAcbeta1-4]Galbeta1-4GlcNAc, NeuGcalpha2-3[GalNAcbeta1 4]Galbeta1-4GlcNAc), and terminal GlcNAc. Polylactosamine-type sequence was also detected on a subset of the antennae. Analysis of the O-glycans indicated that the majority were core 2-type (Galbeta1-4GlcNAcbeta1-6[Galbeta1-3]GalNAc). The beta1-6-linked branches attached to these O-glycans were terminated with the same sequences as the N-glycans, except for terminal GlcNAc. Glycans bearing Galbeta1 4GlcNAcbeta1-6 branches have previously been suggested to mediate initial murine gamete binding. Oligosaccharides terminated with GalNAcbeta1-4Gal have been implicated in the secondary binding interaction that occurs following the acrosome reaction. The significant implications of these observations are discussed. PMID- 10713086 TI - N- and C-terminal domains direct cell type-specific sorting of chromogranin A to secretory granules. AB - Chromogranins are a family of regulated secretory proteins that are stored in secretory granules in endocrine and neuroendocrine cells and released in response to extracellular stimulation (regulated secretion). A conserved N-terminal disulfide bond is necessary for sorting of chromogranins in neuroendocrine PC12 cells. Surprisingly, this disulfide bond is not necessary for sorting of chromogranins in endocrine GH4C1 cells. To investigate the sorting mechanism in GH4C1 cells, we made several mutant forms removing highly conserved N- and C terminal regions of bovine chromogranin A. Removing the conserved N-terminal disulfide bond and the conserved C-terminal dimerization and tetramerization domain did not affect the sorting of chromogranin A to the regulated secretory pathway. In contrast, removing the C-terminal 90 amino acids of chromogranin A caused rerouting to the constitutive secretory pathway and impaired aggregation properties as compared with wild-type chromogranin A. Since this mutant was sorted to the regulated secretory pathway in PC12 cells, these results demonstrate that chromogranins contain independent N- and C-terminal sorting domains that function in a cell type-specific manner. Moreover, this is the first evidence that low pH/calcium-induced aggregation is necessary for sorting of a chromogranin to the regulated secretory pathway of endocrine cells. PMID- 10713087 TI - The alpha and beta subunits of the GA-binding protein form a stable heterodimer in solution. Revised model of heterotetrameric complex assembly. AB - We have studied the assembly of GA-binding protein (GABP) in solution and established the role of DNA in the assembly of the transcriptionally active GABPalpha(2)beta(2) heterotetrameric complex. GABP binds DNA containing a single PEA3/Ets-binding site (PEA3/EBS) exclusively as the alphabeta heterodimer complex, but readily binds as the GABPalpha(2)beta(2) heterotetramer complex on DNA containing two PEA3/EBSs. Positioning of the PEA3/EBSs on the same face of the DNA helix stabilizes heterotetramer complex binding. These observations suggest that GABPalphabeta heterodimers are the predominant molecular species in solution and that DNA containing two PEA3/EBSs promotes formation of the GABPalpha(2)beta(2) heterotetrameric complex. We analyzed the assembly of GABPalpha(2)beta(2) heteromeric complexes in solution by analytical ultracentrifugation. GABPalpha exists as a monomer in solution while GABPbeta exists in a monomer-dimer equilibrium (K(d) = 1.8 +/- 0.27 microM). In equimolar mixtures of the two subunits, GABPalpha and GABPbeta formed a stable heterodimer, with no heterotetramer complex detected. Thus, GABP exists in solution as the heterodimer previously shown to be a weak transcriptional activator. Assembly of the transcriptionally active GABPalpha(2)beta(2) heterotetramer complex requires the presence of specific DNA containing at least two PEA3/EBSs. PMID- 10713088 TI - Reduction of nitrite to nitric oxide catalyzed by xanthine oxidoreductase. AB - Xanthine oxidase (XO) was shown to catalyze the reduction of nitrite to nitric oxide (NO), under anaerobic conditions, in the presence of either NADH or xanthine as reducing substrate. NO production was directly demonstrated by ozone chemiluminescence and showed stoichiometry of approximately 2:1 versus NADH depletion. With xanthine as reducing substrate, the kinetics of NO production were complicated by enzyme inactivation, resulting from NO-induced conversion of XO to its relatively inactive desulfo-form. Steady-state kinetic parameters were determined spectrophotometrically for urate production and NADH oxidation catalyzed by XO and xanthine dehydrogenase in the presence of nitrite under anaerobic conditions. pH optima for anaerobic NO production catalyzed by XO in the presence of nitrite were 7.0 for NADH and 10(3)-fold slower than the rate of ADP/ATP exchange. Thus, in contrast to DnaK and eukaryotic forms of hsp70 that have been characterized to date, the R if T equilibrium balance for Hsc66 is shifted in favor of the low peptide affinity T state, and regulation of the reaction cycle is expected to occur at the ATP hydrolysis step rather than at nucleotide exchange. PMID- 10713092 TI - Differential activation of CC chemokine receptors by AOP-RANTES. AB - RANTES (regulated on activation normal T cell expressed) has been found at elevated levels in biological fluids from patients with a wide range of allergic and autoimmune diseases and is able to attract several subtypes of leukocytes including eosinophils and monocytes into inflamed tissue. Amino-terminal modifications of RANTES produce receptor antagonists which are candidates for blocking this cellular recruitment. Met-RANTES has been shown to modulate inflammation in vivo, while AOP-RANTES is a potent inhibitor of R5 human immunodeficiency virus type 1 (HIV-1) strains and has been shown to down-modulate CCR5 and prevent recycling of the receptor. We have studied the effect of AOP RANTES in eosinophil activation and have found that it is able to efficiently elicit eosinophil effector functions through CCR3, as measured by the release of reactive oxygen species and calcium mobilization, whereas Met-RANTES is inactive in these assays. AOP-RANTES is found to inhibit CCR3-mediated HIV-1 infection with moderate potency, in contrast to its potent inhibition of CCR5-mediated HIV 1 infection. Furthermore, we have investigated the abilities of these modified proteins to down-modulate CCR1 and CCR3 from the surface of monocytes and eosinophils. We show here that AOP-RANTES is much less effective than RANTES in down-modulation of CCR1. Surprisingly, recycling of CCR1 was minimal after incubation with RANTES while there was complete recycling with AOP-RANTES. In the case of CCR3, no significant difference was found between RANTES and AOP-RANTES in down-modulation and recycling. It therefore appears that trafficking of RANTES receptors follows different patterns, which opens up potential new targets for therapeutic intervention. PMID- 10713093 TI - Multiple discontinuous ligand-mimetic antibody binding sites define a ligand binding pocket in integrin alpha(IIb)beta(3). AB - Integrin alpha(IIb)beta(3), a platelet fibrinogen receptor, is critically involved in thrombosis and hemostasis. However, how ligands interact with alpha(IIb)beta(3) has been controversial. Ligand-mimetic anti-alpha(IIb)beta(3) antibodies (PAC-1, LJ-CP3, and OP-G2) contain the RGD-like RYD sequence in their CDR3 in the heavy chain and have structural and functional similarities to native ligands. We have located binding sites for ligand-mimetic antibodies in alpha(IIb) and beta(3) using human-to-mouse chimeras, which we expect to maintain functional integrity of alpha(IIb)beta(3). Here we report that these antibodies recognize several discontinuous binding sites in both the alpha(IIb) and beta(3) subunits; these binding sites are located in residues 156-162 and 229-230 of alpha(IIb) and residues 179-183 of beta(3). In contrast, several nonligand mimetic antibodies (e.g. 7E3) recognize single epitopes in either subunit. Thus, binding to several discontinuous sites in both subunits is unique to ligand mimetic antibodies. Interestingly, these binding sites overlap with several (but not all) of the sequences that have been reported to be critical for fibrinogen binding (e.g. N-terminal repeats 2-3 but not repeats 4-7, of alpha(IIb)). These results suggest that ligand-mimetic antibodies and probably native ligands may make direct contact with these discontinuous binding sites in both subunits, which may constitute a ligand-binding pocket. PMID- 10713094 TI - Purification and characterization of ATM from human placenta. A manganese dependent, wortmannin-sensitive serine/threonine protein kinase. AB - ATM is mutated in the human genetic disorder ataxia telangiectasia, which is characterized by ataxia, immune defects, and cancer predisposition. Cells that lack ATM exhibit delayed up-regulation of p53 in response to ionizing radiation. Serine 15 of p53 is phosphorylated in vivo in response to ionizing radiation, and antibodies to ATM immunoprecipitate a protein kinase activity that, in the presence of manganese, phosphorylates p53 at serine 15. Immunoprecipitates of ATM also phosphorylate PHAS-I in a manganese-dependent manner. Here we have purified ATM from human cells using nine chromatographic steps. Highly purified ATM phosphorylated PHAS-I, the 32-kDa subunit of RPA, serine 15 of p53, and Chk2 in vitro. The majority of the ATM phosphorylation sites in Chk2 were located in the amino-terminal 57 amino acids. In each case, phosphorylation was strictly dependent on manganese. ATM protein kinase activity was inhibited by wortmannin with an IC(50) of approximately 100 nM. Phosphorylation of RPA, but not p53, Chk2, or PHAS-I, was stimulated by DNA. The related protein, DNA-dependent protein kinase catalytic subunit, also phosphorylated PHAS-I, RPA, and Chk2 in the presence of manganese, suggesting that the requirement for manganese is a characteristic of this class of enzyme. PMID- 10713095 TI - Involvement of histidine residues in proton sensing of ROMK1 channel. AB - ROMK channels are inhibited by intracellular acidification. This pH sensitivity is related to several amino acid residues in the channel proteins such as Lys-61, Thr-51, and His-206 (in ROMK2). Unlike all other amino acids, histidine is titratable at pH 6-7 carrying a positive charge below pH 6. To test the hypothesis that certain histidine residues are engaged in CO(2) and pH sensing of ROMK1, we performed experiments by systematic mutations of all histidine residues in the channel using the site-directed mutagenesis. There are two histidine residues in the N terminus. Mutations of His-23, His-31, or both together did not affect channel sensitivity to CO(2). Six histidine residues are located in the C terminus. His-225, His-274, His-342, and His-354 were critical in CO(2) and pH sensing. Mutation of either of them reduced CO(2) and pH sensitivities by 20-50% and approximately 0.2 pH units, respectively. Simultaneous mutations of all of them eliminated the CO(2) sensitivity and caused this mutant channel to respond to only extremely acidic pH. Similar mutations of His-280 had no effect. The role of His-270 in CO(2) and pH sensing is unclear, because substitutions of this residue with either a neutral, negative, or positive amino acid did not produce any functional channel. These results therefore indicate that histidine residues contribute to the sensitivity of the ROMK1 channel to hypercapnia and intracellular acidosis. PMID- 10713096 TI - Divergent roles for Ras and Rap in the activation of p38 mitogen-activated protein kinase by interleukin-1. AB - We have found that lethal toxin from Clostridium sordellii, which specifically inactivates the low molecular weight G proteins Ras, Rap, and Rac, inhibits the activation of p38 mitogen-activated protein kinase (MAPK) by interleukin-1 (IL-1) in EL4.NOB-1 cells and primary fibroblasts. The target protein involved appeared to be Ras, because transient transfections with dominant negative RasN17 inhibited p38 MAPK activation by IL-1. Furthermore, transfections of cells with constitutively active RasVHa-activated p38 MAPK. Further evidence for Ras involvement came from the observation that IL-1 caused a rapid activation of Ras in the cells and from the inhibitory effects of the Ras inhibitors manumycin A and damnacanthal. Toxin B from Clostridium difficile, which inactivates Rac, Cdc42, and Rho, was without effect. Dominant negative versions of Rac (RacN17) or Rap (Rap1AN17) did not inhibit the response. Intriguingly, transfection of cells with dominant negative Rap1AN17 activated p38 MAPK. Furthermore, constitutively active Rap1AV12 inhibited p38 MAPK activation by IL-1, consistent with Rap antagonizing Ras function. IL-1 also activated Rap in the cells, but with slower kinetics than Ras. Our studies therefore provide clear evidence using multiple approaches for Ras as a signaling component in the activation of p38 MAPK by IL 1, with Rap having an inhibitory effect. PMID- 10713097 TI - Characterization of glucokinase regulatory protein-deficient mice. AB - The glucokinase regulatory protein (GKRP) inhibits glucokinase competitively with respect to glucose by forming a protein-protein complex with this enzyme. The physiological role of GKRP in controlling hepatic glucokinase activity was addressed using gene targeting to disrupt GKRP gene expression. Heterozygote and homozygote knockout mice have a substantial decrease in hepatic glucokinase expression and enzymatic activity as measured at saturating glucose concentrations when compared with wild-type mice, with no change in basal blood glucose levels. Interestingly, when assayed under conditions to promote the association between glucokinase and GKRP, liver glucokinase activity in wild-type and null mice displayed comparable glucose phosphorylation capacities at physiological glucose concentrations (5 mM). Thus, despite reduced hepatic glucokinase expression levels in the null mice, glucokinase activity in the liver homogenates was maintained at nearly normal levels due to the absence of the inhibitory effects of GKRP. However, following a glucose tolerance test, the homozygote knockout mice show impaired glucose clearance, indicating that they cannot recruit sufficient glucokinase due to the absence of a nuclear reserve. These data suggest both a regulatory and a stabilizing role for GKRP in maintaining adequate glucokinase in the liver. Furthermore, this study provides evidence for the important role GKRP plays in acutely regulating of hepatic glucose metabolism. PMID- 10713098 TI - Biochemical characterization of netrin-synergizing activity. AB - The netrin-1 protein elicits spinal commissural axon outgrowth and turning in vitro and has been shown to be required for commissural axon guidance in vivo in the developing spinal cord. Biochemical observations made during the purification of netrin-1 suggest that this ligand and its receptor, DCC, may not function alone in directing commissural axon guidance. Recombinant netrin-1 protein is approximately 10 times more active in eliciting axon outgrowth from embryonic day (E) 13 rat dorsal spinal cord explants than from E11 rat dorsal spinal cord explants (Serafini, T., Kennedy, T. E., Galko, M. J., Mirzayan, C., Jessell, T. M., and Tessier-Lavigne, M. (1994) Cell 78, 409-424) even though the starting material for the netrin purification, a high salt extract of E10 chicken brain membranes, is equally active on E13 and E11 explants. We previously reported an activity termed netrin-synergizing activity (NSA) that can potentiate the outgrowth-promoting activity of netrin-1 on E11 explants (Serafini et al.). Here we report a biochemical characterization of NSA in netrin-depleted high salt extracts of E10 chicken brain membranes. We provide evidence that NSA is composed of a denaturation-resistant basic protein(s) in the 25-35-kDa size range. We also provide evidence that the activity may be heterogeneous, splitting into three species that may be distinct or related. The results reported here should facilitate purification of this activity from a more abundant source or identification of the activity based on similarity to known proteins that share its distinctive biochemical properties. PMID- 10713099 TI - Noncovalent association of P-selectin glycoprotein ligand-1 and minimal determinants for binding to P-selectin. AB - P-selectin glycoprotein ligand-1 (PSGL-1) is a disulfide-bonded, homodimeric mucin ( approximately 250 kDa) on leukocytes that binds to P-selectin on platelets and endothelial cells during the initial steps in inflammation. Because it has been proposed that only covalently dimerized PSGL-1 can bind P-selectin, we investigated the factors controlling dimerization of PSGL-1 and re-examined whether covalent dimers are required for binding its P-selectin. Recombinant forms of PSGL-1 were created in which the single extracellular Cys (Cys(320)) was replaced with either Ser (C320S-PSGL-1) or Ala (C320A-PSGL-1). Both recombinants migrated as monomeric species of approximately 120 kDa under both nonreducing and reducing conditions on SDS-polyacrylamide gel electrophoresis. P-selectin bound similarly to cells expressing either wild type or mutated forms of PSGL-1 in both flow cytometric and rolling adhesion assays. Unexpectedly, chemical cross-linking studies revealed that both C320S- and C320A-PSGL-1 noncovalently associate in the plasma membrane and cross-linking generates dimeric species. Chimeric recombinants of PSGL-1 in which the transmembrane domain in PSGL-1 was replaced with the transmembrane domain of CD43 (CD43TMD-PSGL-1) could not be chemically cross-linked, suggesting that residues within the transmembrane domain of PSGL-1 are required for noncovalent association. Cells expressing CD43TMD-PSGL-1 bound P selectin. To further address the ability of P-selectin to bind monomeric derivatives of PSGL-1, intact HL-60 cells were trypsin-treated, which generated a soluble approximately 25-kDa NH(2)-terminal fragment of PSGL-1 that bound to immobilized P-selectin. Because N-glycosylation of PSGL-1 hinders trypsin cleavage, a recombinant form of PSGL-1 was generated in which all three potential N-glycosylation sites were mutated (DeltaN-PSGL-1). Cells expressing DeltaN-PSGL 1 bound P-selectin, and trypsin treatment of the cells generated NH(2)-terminal monomeric fragments (<10 kDa) of PSGL-1 that bound to P-selectin. These results demonstrate that Cys(320)-dependent dimerization of PSGL-1 is not required for binding to P-selectin and that a small monomeric fragment of PSGL-1 is sufficient for P-selectin recognition. PMID- 10713100 TI - Cdc42-interacting protein 4 mediates binding of the Wiskott-Aldrich syndrome protein to microtubules. AB - The Wiskott-Aldrich syndrome is an inherited X-linked immunodeficiency characterized by thrombocytopenia, eczema, and a tendency toward lymphoid malignancy. Lymphocytes from affected individuals have cytoskeletal abnormalities, and monocytes show impaired motility. The Wiskott-Aldrich syndrome protein (WASP) is a multi-domain protein involved in cytoskeletal organization. In a two-hybrid screen, we identified the protein Cdc42-interacting protein 4 (CIP4) as a WASP interactor. CIP4, like WASP, is a Cdc42 effector protein involved in cytoskeletal organization. We found that the WASP-CIP4 interaction is mediated by the binding of the Src homology 3 domain of CIP4 to the proline-rich segment of WASP. Cdc42 was not required for this interaction. Co-expression of CIP4 and green fluorescent protein-WASP in COS-7 cells led to the association of WASP with microtubules. In vitro experiments showed that CIP4 binds to microtubules via its NH(2) terminus. The region of CIP4 responsible for binding to active Cdc42 was localized to amino acids 383-417, and the mutation I398S abrogated binding. Deletion of the Cdc42-binding domain of CIP4 did not affect the colocalization of WASP with microtubules in vivo. We conclude that CIP4 can mediate the association of WASP with microtubules. This may facilitate transport of WASP to sites of substrate adhesion in hematopoietic cells. PMID- 10713101 TI - Subtypes of the somatostatin receptor assemble as functional homo- and heterodimers. AB - The existence of receptor dimers has been proposed for several G protein-coupled receptors. However, the question of whether G protein-coupled receptor dimers are necessary for activating or modulating normal receptor function is unclear. We address this question with somatostatin receptors (SSTRs) of which there are five distinct subtypes. By using transfected mutant and wild type receptors, as well as endogenous receptors, we provide pharmacological, biochemical, and physical evidence, based on fluorescence resonance energy transfer analysis, that activation by ligand induces SSTR dimerization, both homo- and heterodimerization with other members of the SSTR family, and that dimerization alters the functional properties of the receptor such as ligand binding affinity and agonist induced receptor internalization and up-regulation. Double label confocal fluorescence microscopy showed that when SSTR1 and SSTR5 subtypes were coexpressed in Chinese hamster ovary-K1 cells and treated with agonist they underwent internalization and were colocalized in cytoplasmic vesicles. SSTR5 formed heterodimers with SSTR1 but not with SSTR4 suggesting that heterodimerization is a specific process that is restricted to some but not all receptor subtype combinations. Direct protein interaction between different members of the SSTR subfamily defines a new level of molecular cross-talk between subtypes of the SSTR and possibly related receptor families. PMID- 10713102 TI - TrkA amino acids controlling specificity for nerve growth factor. AB - Neurotrophins are important for the development and maintenance of the vertebrate nervous system, mediating their signal into the cell by specific interaction with tyrosine kinase receptors of the Trk family. The extracellular portion of the Trk receptors has been previously proposed to consist of a cysteine-rich motif, a leucine-rich motif, a second cysteine-rich motif followed by two immunoglobulin like domains. Earlier studies have shown that a major neurotrophin-binding site in the Trk receptors resides in the second immunoglobulin-like domain. Although the individual amino acids in TrkA involved in binding to nerve growth factor (NGF) and those in TrkC involved in binding to neurotrophin-3 have been mapped in this domain, the Trk amino acids that provide specificity remained unclear. In this study, a minimum set of residues in the human TrkC second immunoglobulin like domain, which does not bind nerve growth factor (NGF), were substituted with those from human TrkA. The resulting Trk variant recruited binding of NGF equivalent to TrkA, maintained neurotrophin-3 binding equivalent to TrkC, and also bound brain-derived neurotrophin, although with lower affinity compared with TrkB. This implies that the amino acids in the second immunoglobulin-like domain that determine Trk specificity are distinct for each Trk. PMID- 10713103 TI - Targeted oncogenesis reveals a distinct tissue-specific utilization of alternative promoters of the human mineralocorticoid receptor gene in transgenic mice. AB - The human mineralocorticoid receptor (hMR) is a nuclear receptor mediating aldosterone action, whose expression is driven by two alternative promoters, P1 and P2, flanking the two first 5'-untranslated exons. In vivo characterization of hMR regulatory regions was performed by targeted oncogenesis in mice using P1 or P2 directing expression of the large T antigen of SV40 (TAg). While transgenic P1.TAg founders rapidly developed lethal hibernomas from brown fat, cerebral primitive neuroectodermal tumors and facial leiomyosarcomas occurred in P2.TAg mice. Quantitative analyses of mouse MR (mMR) and transgene expression indicate that P1 promoter was transcriptionally active in all MR-expressing tissues, directing strong TAg expression in testis and salivary glands, moderate in lung, brain, uterus, liver, and heart but, unlike mMR, rather low in colon and kidney. Importantly, the renal transgene expression colocalized with mMR in the distal nephron. In contrast, P2 promoter was approximately 10 times less potent than P1, with no activity in the brain and colon. Several immortalized cell lines were established from both neoplastic and normal tissues of transgenic mice. These cells exhibited differentiated characteristics and maintained MR expression, thus providing useful models for further studies exploring the widespread expression and functions of MR. Our results demonstrate that hMR gene expression in vivo is controlled by complex regulatory mechanisms involving distinct tissue-specific utilization of alternative promoters. PMID- 10713104 TI - HS1 interacts with Lyn and is critical for erythropoietin-induced differentiation of erythroid cells. AB - Erythroid cells terminally differentiate in response to erythropoietin binding its cognate receptor. Previously we have shown that the tyrosine kinase Lyn associates with the erythropoietin receptor and is essential for hemoglobin synthesis in three erythroleukemic cell lines. To understand Lyn signaling events in erythroid cells, the yeast two-hybrid system was used to analyze interactions with other proteins. Here we show that the hemopoietic-specific protein HS1 interacted directly with the SH3 domain of Lyn, via its proline-rich region. A truncated HS1, bearing the Lyn-binding domain, was introduced into J2E erythroleukemic cells to determine the impact upon responsiveness to erythropoietin. Truncated HS1 had a striking effect on the phenotype of the J2E line-the cells were smaller, more basophilic than the parental proerythoblastoid cells and had fewer surface erythropoietin receptors. Moreover, basal and erythropoietin-induced proliferation and differentiation were markedly suppressed. The inability of cells containing the truncated HS1 to differentiate may be a consequence of markedly reduced levels of Lyn and GATA-1. In addition, erythropoietin stimulation of these cells resulted in rapid, endosome-mediated degradation of endogenous HS1. The truncated HS1 also suppressed the development of erythroid colonies from fetal liver cells. These data show that disrupting HS1 has profoundly influenced the ability of erythroid cells to terminally differentiate. PMID- 10713105 TI - A novel member of the BTB/POZ family, PATZ, associates with the RNF4 RING finger protein and acts as a transcriptional repressor. AB - We have identified a novel human gene encoding a 59-kDa POZ-AT hook-zinc finger protein (PATZ) that interacts with RNF4, a mediator of androgen receptor activity, and acts as a transcriptional repressor. PATZ cDNA was isolated through a two-hybrid interaction screening using the RING finger protein RNF4 as a bait. In vitro and in vivo interaction between RNF4 and PATZ was demonstrated by protein-protein affinity chromatography and coimmunoprecipitation experiments. Such interaction occurred through a small region of PATZ containing an AT-hook DNA binding domain. Immunofluorescence staining and confocal microscopy showed that PATZ localizes in distinct punctate nuclear regions and colocalizes with RNF4. Functional analysis was performed by cotransfection assays: PATZ acted as a transcriptional repressor, whereas its partner RNF4 behaved as a transcriptional activator. When both proteins were overexpressed a strong repression of the basal transcription was observed, indicating that the association of PATZ with RNF4 switches activation to repression. In addition, RNF4 was also found to associate with HMGI(Y), a chromatin-modeling factor containing AT-hook domains. PMID- 10713106 TI - The human growth hormone gene cluster locus control region supports position independent pituitary- and placenta-specific expression in the transgenic mouse. AB - The human growth hormone (hGH) cluster contains five genes. The hGH-N gene is predominantly expressed in pituitary somatotropes, whereas the remaining four genes, the chorionic somatomammotropin genes (hCS-L, hCS-A, and hCS-B) and hGH-V, are expressed selectively in the placenta. In contrast, the mouse genome contains a single pituitary-specific GH gene and lacks any GH-related CS genes. Activation of the hGH transgene in the mouse is dependent on its linkage to a previously described locus control region (LCR) located -15 to -32 kilobases upstream of the hGH cluster. The sporadic, nonreproducible expression of hCS transgenes lacking the LCR suggests that they may be dependent on hGH LCR activity as well. To determine whether the hCS genes could be expressed with appropriate placental specificity, a series of five transgenic mouse lines carrying an 87-kilobase human genomic insert encompassing the majority of the hGH gene cluster and the entire contiguous LCR was established. All of the hGH cluster genes were appropriately expressed in each of these lines. High level expression of hGH was restricted to the pituitary and hCS to the labyrinthine layer of the placenta. The expression of the GH cluster genes in their respective tissues paralleled transgene copy numbers irrespective of the transgene insertion site in the host mouse genome. These studies have extended the utility of the transgenic mouse model for the analysis of the full spectrum of hGH gene cluster activation. Further, they support a role for the hGH LCR in placental hCS, as well as pituitary hGH gene activation, and expression. PMID- 10713107 TI - Cellubrevin is present in the basolateral endocytic compartment of hepatocytes and follows the transcytotic pathway after IgA internalization. AB - The endocytic compartment of polarized cells is organized in basolateral and apical endosomes plus those endocytic structures specialized in recycling and transcytosis, which are still poorly characterized. The complexity of the various populations of endosomes has been demonstrated by the exquisite repertoire of endogenous proteins. In this study we examined the distribution of cellubrevin in the endocytic compartment of hepatocytes, since its intracellular location and function in polarized cells are largely unknown. Highly purified rat liver endosomes were isolated from estradiol-treated rats, and the early/sorting endosomal fraction was further subfractionated in a multistep sucrose density gradient, and studied. Analysis of dissected endosomal fractions showed that cellubrevin was located in early/sorting endosomes, with Rab4, annexins II and VI, and transferrin receptor, but in a specific subpopulation of these early endosomes with the same density range as pIgA and Raf-1. Interestingly, only in those isolated endosomal fractions, endosomes enriched in transcytotic structures (of livers loaded with IgA), the polymeric immunoglobulin receptor specifically co-immunoprecipitated with cellubrevin. In addition, confocal and immuno-electron microscopy identification of cellubrevin in tubular structures underneath the sinusoidal plasma membrane together with the re-organization of cellubrevin, in the endocytic compartment, after the IgA loading, strongly suggest the involvement of cellubrevin in the transcytosis of pIgA. PMID- 10713108 TI - Beta-amyloid stimulation of inducible nitric-oxide synthase in astrocytes is interleukin-1beta- and tumor necrosis factor-alpha (TNFalpha)-dependent, and involves a TNFalpha receptor-associated factor- and NFkappaB-inducing kinase dependent signaling mechanism. AB - In Alzheimer's disease, beta-amyloid (Abeta) plaques are surrounded by activated astrocytes and microglia. A growing body of evidence suggests that these activated glia contribute to neurotoxicity through the induction of inflammatory cytokines such as interleukin (IL)-1beta and tumor necrosis factor-alpha (TNFalpha) and the production of neurotoxic free radicals, mediated in part by the expression of inducible nitric-oxide synthase (iNOS). Here, we address the possibility that Abeta-stimulated iNOS expression might result from an initial induction of IL-1beta and TNFalpha. We find that in Abeta-stimulated astrocyte cultures, IL-1beta and TNFalpha production occur before iNOS production, new protein synthesis is required for increased iNOS mRNA levels, and the IL-1 receptor antagonist IL-1ra can inhibit nitrite accumulation. Likewise, dominant negative mutants of tumor necrosis factor-alpha receptor-associated factor (TRAF) 6, TRAF2, and NFkappaB-inducing kinase (NIK), intracellular proteins involved in IL-1 and TNFalpha receptor signaling cascades, inhibit Abeta-stimulated iNOS promoter activity. Our data suggest that Abeta stimulation of astrocyte iNOS is mediated in part by IL-1beta and TNFalpha, and involves a TRAF6-, TRAF2-, and NIK dependent signaling mechanism. PMID- 10713109 TI - Cell wall biogenesis of Blastomyces dermatitidis. Evidence for a novel mechanism of cell surface localization of a virulence-associated adhesin via extracellular release and reassociation with cell wall chitin. AB - Pathogenic yeast of Blastomyces dermatitidis express a surface protein adhesin, WI-1. Due to the crucial role of WI-1 in adherence and disease pathogenesis, we investigated how the protein localizes to the surface of B. dermatitidis. WI-1 released extracellularly by wild-type yeast coated the surfaces of co-cultured knockout yeast within 3 h of incubation, implying that secreted WI-1 provides a pathway for loading the protein onto the yeast cell wall. In radioligand binding assays, purified WI-1 bound saturably, specifically, and with high affinity (K(d) = 8.3 x 10(-9)) to the cell surface of knockout yeast devoid of WI-1. WI-1 added exogenously, in vitro, to knockout yeast was indistinguishable from native cell surface WI-1 by fluorescence staining and restored adhesivity to the knockout yeast in macrophage binding and phagocytosis assays. Analysis of interactions between WI-1 and elements of the yeast cell wall identified chitin as the anchor point for WI-1. This interaction was shown to hinge on the 24-amino acid tandem repeat sequence of WI-1. Efforts to extract surface WI-1 from the yeast demonstrated that it is fastened to the wall by non-covalent interactions and covalent links between cysteine residues. We conclude that the yeast cell surface adhesin WI-1 localizes to the cell wall, in part, through extracellular release followed by high affinity binding back onto exposed chitin fibrils. These findings point to a novel pathway of cell wall biogenesis in yeast and an unanticipated role for chitin in anchoring and displaying a surface adhesin and virulence determinant. PMID- 10713110 TI - A novel aspartyl proteinase from apocrine epithelia and breast tumors. AB - GCDFP-15 (gross cystic disease fluid protein, 15 kDa) is a secretory marker of apocrine differentiation in breast carcinoma. In human breast cancer cell lines, gene expression is regulated by hormones, including androgens and prolactin. The protein is also known under different names in different body fluids such as gp17 in seminal plasma. GCDFP-15/gp17 is a ligand of CD4 and is a potent inhibitor of T-cell apoptosis induced by sequential CD4/T-cell receptor triggering. We now report that GCDFP-15/gp17 is a protease exhibiting structural properties relating it to the aspartyl proteinase superfamily. Unexpectedly, GCDFP-15/gp17 appears to be related to the retroviral members rather than to the known cellular members of this class. Site-specific mutagenesis of Asp(22) (predicted to be catalytically important for the active site) and pepstatin A inhibition confirmed that the protein is an aspartic-type protease. We also show that, among the substrates tested, GCDFP-15/gp17 is specific for fibronectin. The study of GCDFP-15/gp17 mediated proteolysis may provide a handle to understand phenomena as diverse as mammary tumor progression and fertilization. PMID- 10713111 TI - A novel topology model of the human Na(+)/H(+) exchanger isoform 1. AB - The membrane topology of the human Na(+)/H(+) exchanger isoform 1 (NHE1) was assessed by substituted cysteine accessibility analysis. Eighty-three cysteine residues were individually introduced into a functional cysteineless NHE1, and these mutants were expressed in the exchanger-deficient PS120 cells. The topological disposition of introduced cysteines was determined by labeling with a biotinylated maleimide in the presence or absence of preincubation with the membrane-impermeable sulfhydryl reagent, 2-trimethylammoniumethyl methanethiosulfonate in streptolysin O-permeabilized or nonpermeabilized cells. We proposed a new model for the topology of NHE1 that is significantly different from the model derived from hydropathy analysis. In this model, NHE1 is composed of 12 transmembrane segments (TMs) with the N and C termini located in the cytosol. The large, last extracellular loop in the membrane domain of the original model was suggested to comprise an intracellular loop, a new transmembrane segment (TM11), and an extracellular loop in the new model. Interestingly, cysteines at 183 and 184 and at 324 and 325 mapped to intracellular loops connecting TMs 4 and 5 (IL2) and TMs 8 and 9 (IL4), respectively, were accessible to sulfhydryl reagents from the outside. Furthermore, exchange activities of two mutants, R180C and Q181C, within IL2 were markedly inhibited by external MTSET. These data suggest that part of IL2 or IL4 may be located in a pore-lining region that is accessible from either side of the membrane and involved in ion transport. PMID- 10713112 TI - A human importin-beta family protein, transportin-SR2, interacts with the phosphorylated RS domain of SR proteins. AB - Serine/arginine-rich proteins (SR proteins) are mainly involved in the splicing of precursor mRNA. RS domains are also found in proteins that have influence on other aspects of gene expression. Proteins that contain an RS domain are often located in the speckled domains of the nucleus. Here we show that the RS domain derived from a human papillomavirus E2 transcriptional activator can target a heterologous protein to the nucleus, as it does in many other SR proteins, but insufficient for localization in speckles. By using E2 as a bait in a yeast two hybrid screen, we identified a human importin-beta family protein that is homologous to yeast Mtr10p and almost identical to human transportin-SR. This transportin-SR2 (TRN-SR2) protein can interact with several cellular SR proteins. More importantly, we demonstrated that TRN-SR2 can directly interact with phosphorylated, but not unphosphorylated, RS domains. Finally, an indirect immunofluoresence study revealed that a transiently expressed TRN-SR2 mutant lacking the N-terminal region becomes localized to the nucleus in a speckled pattern that coincides with the distribution of the SR protein SC35. Thus, our results likely reflect a role of TRN-SR2 in the cellular trafficking of phosphorylated SR proteins. PMID- 10713113 TI - Mitochondrial import and processing of wild type and type III mutant isovaleryl CoA dehydrogenase. AB - Isovaleric acidemia is a rare inborn error of metabolism caused by a deficiency of isovaleryl-CoA dehydrogenase (IVD), a nucleus-encoded, homotetrameric, mitochondrial flavoenzyme that catalyzes the conversion of isovaleryl-CoA to 3 methylcrotonyl-CoA. We have previously identified a nucleotide deletion in the gene for IVD in fibroblasts from a patient with isovaleric acidemia leading to a shift in reading frame and premature termination of translation. The mutant IVD precursor is imported and processed to mature size, but no active enzyme is detected in mutant fibroblasts or expressed in Escherichia coli. Examination of the crystal structure of human IVD reveals that the C terminus is involved in tetramer stability. In vitro mitochondrial import experiments show that wild type IVD protein rapidly and stably forms mature homotetramer following import, whereas Type III mutant protein never forms stable oligomers. An additional series of mutant proteins with truncations and/or alterations in the C-terminal sequence implicates the C terminus of IVD in both enzyme activity and tetramer stability. Importantly, a dimeric intermediate in the folding pathway for wild type IVD has been identified in the in vitro mitochondrial import experiments, the first report of such an intermediate in the biogenesis of an acyl-CoA dehydrogenase. PMID- 10713114 TI - Heparan sulfate proteoglycan isoforms of the CD44 hyaluronan receptor induced in human inflammatory macrophages can function as paracrine regulators of fibroblast growth factor action. AB - The CD44 glycoprotein is expressed in multiple isoforms on a variety of cell types where it functions as a receptor for hyaluronan-mediated motility. Recently, interest has centered on CD44 heparan sulfate proteoglycan (HSPG) isoforms because of their potential to sequester heparin-binding growth factors and chemokines. Expression of these isoforms on ectodermal cells has recently been shown to regulate limb morphogenesis via presentation of fibroblast growth factor (FGF) 4/FGF 8 while expression on tumor cells was shown to sequester hepatocyte growth factor and promote tumor dissemination. To date, however, CD44 HSPG expression in tissue macrophages and lymphocytes has not been adequately investigated, despite the fact these cells actively synthesize growth factors and chemokines and indirect evidence that monocyte CD44 sequesters macrophage inflammatory protein-1beta. Here we show primary human monocytes rather than lymphocytes express CD44 HSPGs, but only following in vitro differentiation to macrophages or activation with the proinflammatory cytokine interleukin-1alpha or bacterial lipopolysaccharide. Furthermore, we show these isoforms are preferentially modified with heparan rather than chondroitin sulfate, bind the macrophage-derived growth factors FGF-2, vascular endothelial growth factor, and heparin-binding epidermal growth factor with varying affinities (K(d) 25-330 nM) and in the case of FGF-2, can stimulate productive binding to the high affinity tyrosine kinase FGF receptor 1 (FGFR1). In contrast, we find no evidence for significant binding to C-C chemokines. Last, we confirm by immunofluorescent antibody staining that inflamed synovial membrane macrophages express CD44 HSPGs and that expression is greatest in cells containing high FGF-2 levels. These results suggest a paracrine role for macrophage CD44 HSPG isoforms in the regulation of growth factor action during inflammation. PMID- 10713115 TI - Dual role for the glutamine phosphoribosylpyrophosphate amidotransferase ammonia channel. Interdomain signaling and intermediate channeling. AB - Glutamine phosphoribosylpyrophosphate (PRPP) amidotransferase catalyzes the first reaction of de novo purine nucleotide synthesis in two steps at two sites. Glutamine is hydrolyzed to glutamate plus NH(3) at an N-terminal glutaminase site, and NH(3) is transferred through a 20-A hydrophobic channel to a distal PRPP site for synthesis of phosphoribosylamine. Binding of PRPP is required to activate the glutaminase site (termed interdomain signaling) to prevent the wasteful hydrolysis of glutamine in the absence of phosphoribosylamine synthesis. Mutations were constructed to analyze the function of the NH(3) channel. In the wild type enzyme, NH(3) derived from glutamine hydrolysis was transferred to the PRPP site, and little or none was released. Replacement of Leu-415 at the PRPP end of the channel with an alanine resulted in a leaky channel and release of NH(3) to the solvent. Mutations in five amino acids that line the channel and two other residues required for the reorganization of phosphoribosyltransferase domain "flexible loop" that leads to formation of the channel perturbed channel function as well as interdomain signaling. The data emphasize the role of the NH(3) channel in coupling interdomain signaling and NH(3) transfer. PMID- 10713116 TI - A mutation in yeast topoisomerase II that confers hypersensitivity to multiple classes of topoisomerase II poisons. AB - A mutation was constructed in the CAP homology domain of yeast topoisomerase II that resulted in hypersensitivity to the intercalating agent N-[4-(9 acridinylamino)-3-methoxy-phenyl]methanesulfonamide and the fluoroquinolone 6, 8 difluoro-7-(4'-hydroxyphenyl)-1-cyclopropyl-4-quinolone-3-carboxyli c acid, but not to etoposide. This mutation, which changes threonine at position 744 to proline, also confers hypersensitivity to anti-bacterial fluoroquinolones. The purified T744P mutant protein had wild type enzymatic activity in the absence of drugs, and no alteration in drug-independent DNA cleavage. Enhanced DNA cleavage in the presence of N-[4-(9-acridinylamino)-3-methoxy-phenyl]methanesulfonamide and fluoroquinolones was observed, in agreement with the results observed in vivo. DNA cleavage was also seen in the presence of norfloxacin and oxolinic acid, two quinolones that are inactive against eukaryotic topoisomerase II. The hypersensitivity was not associated with heat-stable covalent complexes, as was seen in another drug-hypersensitive mutant. Molecular modeling suggests that the mutation in the CAP homology domain may displace amino acids that play important roles in catalysis by topoisomerase II and may explain the drug-hypersensitive phenotype. PMID- 10713117 TI - A membrane-bound Fas decoy receptor expressed by human thymocytes. AB - Human thymocytes at several stages of maturation express Fas, yet resist apoptosis induction through its ligation. A proximal step in apoptotic signaling through Fas is implicated in this resistance, as these cells undergo normal levels of apoptosis induction after exposure to tumor necrosis factor-alpha. We studied the Fas receptors expressed in human thymocytes to search for mechanisms of receptor-mediated inhibition of Fas signaling in these cells. We describe here a unique, membrane-bound form of Fas receptor that contained a complete extracellular domain of Fas but that lacked a death domain due to alternative splicing of exon 7. This Fas decoy receptor (FDR) was shown to have nearly wild type ability to bind native human Fas ligand and was expressed predominantly at the plasma membrane. Unlike soluble forms of Fas receptor, FDR dominantly inhibited apoptosis induction by Fas ligand in transfected human embryonic kidney cells. Titration of FDR in Fas-expressing cells suggests that FDR may operate through the formation of mixed receptor complexes. FDR also dominantly inhibited Fas-induced apoptosis in Jurkat T cells. In mixing experiments with wild-type Fas, FDR was capable of inhibiting death signaling at molar ratios less than 0.5, and this relative level of FDR:wild type message was observed in at least some thymocytes tested. The data suggest that Fas signal pathways in primary human cells may be regulated by expression of a membrane-bound decoy receptor, analogous to the regulation of tumor necrosis factor-related apoptosis inducing ligand (TRAIL)-induced apoptosis by decoy receptors. PMID- 10713118 TI - Ca(2+)/calmodulin-dependent protein kinase IV is expressed in spermatids and targeted to chromatin and the nuclear matrix. AB - Ca(2+)/calmodulin-dependent protein kinase IV and calspermin are two proteins encoded by the Camk4 gene. Both are highly expressed in the testis, where in situ hybridization studies in rat testes have demonstrated that CaMKIV mRNA is localized to pachytene spermatocytes, while calspermin mRNA is restricted to spermatids. We have examined the expression patterns of both CaMKIV and calspermin in mouse testis and unexpectedly find that CaMKIV is expressed in spermatogonia and spermatids but excluded from spermatocytes, while calspermin is found only in spermatids. CaMKIV and calspermin expression in the testis are stage-dependent and appear to be coordinately regulated. In germ cells, we find that CaMKIV is associated with the chromatin. We further demonstrate that a fraction of CaMKIV in spermatids is hyperphosphorylated and specifically localized to the nuclear matrix. These novel findings may implicate CaMKIV in chromatin remodeling during nuclear condensation of spermatids. PMID- 10713119 TI - Trypsin sheds light on the singular case of seminal RNase, a dimer with two quaternary conformations. AB - Dimeric seminal RNase presents the singular case of a dimer with access at equilibrium to two conformations: one in which the subunits exchange, or swap, their NH(2)-terminal arms; the other with no exchange. Thus a continuous unfolding/refolding of structural elements into two alternative conformations takes place in the native protein at equilibrium. The phenomenon was investigated by kinetic and mass spectrometric analyses of the effects of trypsin on the native protein, on its isolated quaternary forms, as well as on a monomeric derivative of the protein and on homologous dimeric RNase A. The kinetics of tryptic action on the protein forms and on the protein derivatives, as well as the location of the tryptic cleavage sites, and their chronological sequence, led to the identification of relevant interconversion intermediates, to the description of a model for the interconversion process, and to a hypothesis for the unique phenomenon of the dual quaternary conformation of seminal RNase. PMID- 10713120 TI - Molecular cloning of mouse ganglioside sialidase and its increased expression in Neuro2a cell differentiation. AB - Ganglioside sialidases have been implicated in neuronal differentiation processes, including neurite outgrowth. To understand further the roles and regulation mechanisms of the sialidase in neuronal systems, we have cloned mouse ganglioside sialidase cDNA and observed its expression in Neuro2a cell differentiation. A 3339-base pair cDNA, cloned based on the sequence information of previously cloned enzymes, encodes 418 amino acids containing three Asp boxes characteristic of sialidases. Northern blot analysis revealed a 3.4-kilobase transcript expressed highly in heart but also in several other tissues including brain. In situ hybridization of mouse brain demonstrated the mRNA to be present in the cerebral cortex, as well as in the granule cell layer, Purkinje cells, and deep cerebellar nucleus of the cerebellum. Transient expression of the cDNA in COS-1 cells resulted in over 300-fold increase in sialidase activity toward gangliosides compared with the control level, with a preference for ganglioside substrate. During 5-bromodeoxyuridine-induced Neuro2a cell differentiation, the expression of the sialidase was increased as assessed by activity assays and quantitative reverse transcription-polymerase chain reaction analyses. Stable transfection of the sialidase in Neuro2a cells resulted in accelerated neurite arborization following 5-bromodeoxyuridine treatment, indicating the direct participation of this ganglioside sialidase in neuronal cell differentiation. PMID- 10713121 TI - Signal-transducing mechanisms involved in activation of the platelet collagen receptor integrin alpha(2)beta(1). AB - Evidence was obtained about the mechanism responsible for platelet integrin alpha(2)beta activation by determining effects of various inhibitors on soluble collagen binding, a parameter to assess integrin alpha(2)beta(1) activation, in stimulated platelets. Agonists that can also activate platelet glycoprotein IIb/IIIa are able to activate integrin alpha(2)beta(1), but those operating via glycoprotein Ib cannot. Activation of alpha(2)beta(1) induced by low thrombin or collagen-related peptide concentrations was almost completely inhibited by apyrase, and the inhibitors wortmannin, 4-amino-5-(chlorophenyl)-7-(t butyl)pyrazolo[3,4-d]pyrimidine, bisindolylmaleimide I, and SQ29548 significantly inhibited it. Activation induced by high thrombin or collagen-related peptide concentrations was far less sensitive to these inhibitors. However, only wortmannin markedly inhibited ADP-induced integrin alpha(2)beta(1) activation, and this was not ADP concentration-dependent. These results suggest that at the low agonist concentrations, the released ADP would be a primary inducer of integrin alpha(2)beta(1) activation, while at the high agonist concentrations, there would be several pathways through which integrin alpha(2)beta(1) activation can be induced. Kinetic analyses revealed that ADP-induced platelets had about the same number of binding sites (B(max)) as thrombin-induced platelets, but their affinity (K(d)) for soluble collagen was 3.7-12.7-fold lower, suggesting that activated integrin alpha(2)beta(1) induced by ADP is different from that induced by thrombin. The data are consistent with an activation mechanism involving released ADP and in which there exists two different states of activated integrin alpha(2)beta(1); these activated forms of integrin alpha(2)beta(1) would have different conformations that determine their ligand affinity. PMID- 10713122 TI - HER-2/neu blocks tumor necrosis factor-induced apoptosis via the Akt/NF-kappaB pathway. AB - Overexpression of HER-2/neu correlates with poor survival of breast and ovarian cancer patients and induces resistance to tumor necrosis factor (TNF), which causes cancer cells to escape from host immune defenses. The mechanism of HER 2/neu-induced TNF resistance is unknown. Here we report that HER-2/neu activates Akt and NF-kappaB without extracellular stimulation. Blocking of the Akt pathway by a dominant-negative Akt sensitizes the HER-2/neu-overexpressing cells to TNF induced apoptosis and inhibits IkappaB kinases, IkappaB phosphorylation, and NF kappaB activation. Our results suggested that HER-2/neu constitutively activates the Akt/NF-kappaB anti-apoptotic cascade to confer resistance to TNF on cancer cells and reduce host defenses against neoplasia. PMID- 10713124 TI - The effect of the DNA flanking the lesion on formation of the UvrB-DNA preincision complex. Mechanism for the UvrA-mediated loading of UvrB onto a DNA damaged site. AB - The UvrB-DNA preincision complex plays a key role in nucleotide excision repair in Escherichia coli. To study the formation of this complex, derivatives of a DNA substrate containing a cholesterol adduct were constructed. Introduction of a single strand nick into either the top or the bottom strand at the 3' side of the adduct stabilized the UvrB-DNA complex, most likely by the release of local stress in the DNA. Removal of both DNA strands up to the 3' incision site still allowed formation of the preincision complex. Similar modifications at the 5' side of the damage, however, gave different results. The introduction of a single strand nick at the 5' incision site completely abolished the UvrA-mediated formation of the UvrB-DNA complex. Deletion of both DNA strands up to the 5' incision site also prevented the UvrA-mediated loading of UvrB onto the damaged site, but UvrB by itself could bind very efficiently. This demonstrates that the UvrB protein is capable of recognizing damage without the matchmaker function of the UvrA protein. Our results also indicate that the UvrA-mediated loading of the UvrB protein is an asymmetric process, which starts at the 5' side of the damage. PMID- 10713123 TI - Biochemical characterization of endogenously formed eosinophilic crystals in the lungs of mice. AB - Crystals seldom form spontaneously within tissues of mammals, except in the urinary tract or in association with eosinophil-rich diseases in humans (Charcot Leyden crystals). Endogenously formed eosinophilic crystals have been reported in respiratory tract and other tissues of several strains of mice, but the biochemical characterization of these crystals has not been reported. In this study, eosinophilic crystal formation was examined in homozygous C57BL/6J viable motheaten mice, lung-specific surfactant apoprotein C promoter/soluble human tumor necrosis factor p75 receptor type II fusion protein transgenic mice (C57BL/6NTac x Sv/129), and CD40L-deficient mice with spontaneous Pneumocystis carinii infection. In viable motheaten but not wild type mice, rapidly developing crystals represented a major feature of the fatal lung injury induced by macrophage dysregulation. Conversely, eosinophilic crystals did not form until 4 8 months of age in transgenic and CD40L-deficient mice and were present in 10-30% of age-matched wild type controls. Mass spectrometry analysis of proteins from bronchoalveolar lavage fluid identified the crystals as Ym1, sometimes referred to as T-lymphocyte-derived eosinophil chemotactic factor. The Ym1 sequence was homologous to chitinase, and enzymatic assays indicated a 3-5-fold increase in chitinase activity compared with control mice. Intracellular and extracellular crystals associated with epithelial damage suggested that the crystals may contribute to lung inflammation through mechanical damage and enzymatic degradation. PMID- 10713125 TI - The role of ATP binding and hydrolysis by UvrB during nucleotide excision repair. AB - We have isolated UvrB-DNA complexes by capture of biotinylated damaged DNA substrates on streptavidin-coated magnetic beads. With this method the UvrB-DNA preincision complex remains stable even in the absence of ATP. For the binding of UvrC to the UvrB-DNA complex no cofactor is needed. The subsequent induction of 3' incision does require ATP binding by UvrB but not hydrolysis. This ATP binding induces a conformational change in the DNA, resulting in the appearance of the DNase I-hypersensitive site at the 5' side of the damage. In contrast, the 5' incision is not dependent on ATP binding because it occurs with the same efficiency with ADP. We show with competition experiments that both incision reactions are induced by the binding of the same UvrC molecule. A DNA substrate containing damage close to the 5' end of the damaged strand is specifically bound by UvrB in the absence of UvrA and ATP (Moolenaar, G. F., Monaco, V., van der Marel, G. A., van Boom, J. H., Visse, R., and Goosen, N. (2000) J. Biol. Chem. 275, 8038-8043). To initiate the formation of an active UvrBC-DNA incision complex, however, UvrB first needs to hydrolyze ATP, and subsequently a new ATP molecule must be bound. Implications of these findings for the mechanism of the UvrA-mediated formation of the UvrB-DNA preincision complex will be discussed. PMID- 10713126 TI - New functions for non-collagenous domains of human collagen type IV. Novel integrin ligands inhibiting angiogenesis and tumor growth in vivo. AB - Collagen type IV is a major component of the basal lamina of blood vessels. Six genetically distinct collagen type IV chains have been identified and are distributed in a tissue-specific manner. Here we define a novel function for soluble non-collagenous (NC1) domains of the alpha2(IV), alpha3(IV), and alpha6(IV) chains of human collagen type IV in the regulation of angiogenesis and tumor growth. These NC1 domains were shown to regulate endothelial cell adhesion and migration by distinct alpha(v) and beta(1) integrin-dependent mechanisms. Systemic administration of recombinant alpha2(IV), alpha3(IV), and alpha6(IV) NC1 domains potently inhibit angiogenesis and tumor growth, whereas alpha1(IV), alpha4(IV), and alpha5(IV) showed little if any effect. These findings suggest that specific NC1 domains of collagen type IV may represent an important new class of angiogenesis inhibitors. PMID- 10713127 TI - Identification and characterization of human SLP-2, a novel homologue of stomatin (band 7.2b) present in erythrocytes and other tissues. AB - Human stomatin (band 7.2b) is a 31-kDa erythrocyte membrane protein of unknown function but implicated in the control of ion channel permeability, mechanoreception, and lipid domain organization. Although absent in erythrocytes from patients with hereditary stomatocytosis, stomatin is not linked to this disorder. A second stomatin homologue, termed SLP-1, has been identified in nonerythroid tissues, and other stomatin related proteins are found in Drosophila, Caenorhabditis elegans, and plants. We now report the cloning and characterization of a new and unusual stomatin homologue, human SLP-2 (stomatin like protein 2). SLP-2 is encoded by an approximately 1.5-kilobase mRNA (GenBank(TM) accession no. AF190167). The gene for human SLP-2, HUSLP2, is present on chromosome 9p13. Its derived amino acid sequence predicts a 38,537-kDa protein that is overall approximately 20% similar to human stomatin. Northern and Western blots for SLP-1 and SLP-2 reveal a wide but incompletely overlapping tissue distribution. Unlike SLP-1, SLP-2 is also present in mature human erythrocytes ( approximately 4,000 +/- 5,600 (+/- 2 S.D.) copies/cell). SLP-2 lacks a characteristic NH(2)-terminal hydrophobic domain found in other stomatin homologues and (unlike stomatin) is fully extractable from erythrocyte membranes by NaOH, pH 11. SLP-2 partitions into both Triton X-100-soluble and -insoluble pools in erythrocyte ghost membranes or when expressed in cultured COS cells and migrates anomalously on SDS-polyacrylamide gel electrophoresis analysis with apparent mobilities of approximately 45,500, 44,600, and 34,300 M(r). The smallest of these protein bands is believed to represent the product of alternative translation initiated at AUGs beginning with nt 217 or 391, although this point has not been rigorously proven. Collectively, these findings identify a novel and unusual member of the stomatin gene superfamily that interacts with the peripheral erythrocyte cytoskeleton and presumably other integral membrane proteins but not directly with the membrane bilayer. We hypothesize that SLP-2 may link stomatin or other integral membrane proteins to the peripheral cytoskeleton and thereby play a role in regulating ion channel conductances or the organization of sphingolipid and cholesterol-rich lipid rafts. PMID- 10713129 TI - Fibroblast growth factor 9 secretion is mediated by a non-cleaved amino-terminal signal sequence. AB - Fibroblast growth factors are a family of intercellular signaling molecules with multiple and varied roles in animal development. Most are exported from cells by means of a classical amino-terminal signal sequence that is cleaved from the mature protein during its passage through the secretory pathway. Fibroblast growth factor-9 (Fgf-9) does not contain a recognizable signal sequence, although it is efficiently secreted. In this study, we show that Fgf-9 enters the endoplasmic reticulum and traverses the Golgi complex in a similar manner to other constitutively secreted proteins. Deletion and point mutation analysis has revealed an atypical non-cleaved signal sequence within the amino-terminal region of Fgf-9. Moreover, the first 28 amino acids of Fgf-9 can function as an efficient non-cleaved signal peptide when appended to the amino terminus of green fluorescent protein. PMID- 10713128 TI - Annexin 24 from Capsicum annuum. X-ray structure and biochemical characterization. AB - This work provides the first three-dimensional structure of a member of the plant annexin family and correlates these findings with biochemical properties of this protein. Annexin 24(Ca32) from Capsicum annuum was purified as a native protein from bell pepper and was also prepared by recombinant techniques. To overcome the problem of precipitation of the recombinant wild-type protein in crystallization trials, two mutants were designed. Whereas an N-terminal truncation mutant turned out to be an unstable protein, the N-terminal His-tagged annexin 24(Ca32) was crystallized, and the three-dimensional structure was determined by x-ray diffraction at 2. 8 A resolution. The structure refined to an R-factor of 0.216 adopts the typical annexin fold; the detailed structure, however, is different from non-plant annexins, especially in domains I and III and in the membrane binding loops on the convex side. Within the unit cell there are two molecules per asymmetric unit, which differ in conformation of the IAB-loop. Both conformers show Trp-35 on the surface. The loop-out conformation is stabilized by tight interactions of this tryptophan with residue side chains of a symmetry related molecule and enforced by a bound sulfate. Characterization of this plant annexin using biophysical methods revealed calcium-dependent binding to phospholipid vesicles with preference for phosphatidylcholine over phosphatidylserine and magnesium-dependent phosphodiesterase activity in vitro as shown with adenosine triphosphate as the substrate. A comparative unfolding study of recombinant annexin 24(Ca32) wild type and of the His-tag fusion protein indicates higher stability of the latter. The effect of this N-terminal modification is also visible from CD spectra. Both proteins were subjected to a FURA-2-based calcium influx assay, which gave high influx rates for the wild-type but greatly reduced influx rates for the fusion protein. We therefore conclude that the N-terminal domain is indeed a major regulatory element modulating different annexin properties by allosteric mechanisms. PMID- 10713130 TI - Specific chaperone-like activity of inhibitor of caspase-activated DNase for caspase-activated DNase. AB - Caspase-activated DNase (CAD) is the enzyme that causes DNA fragmentation during apoptosis. CAD forms aggregates when it is synthesized in the absence of an inhibitor of CAD (ICAD). Here, using renaturation systems of chemically denatured CAD, we report that ICAD-L, a long form of ICAD, has a chaperone-like activity specific for CAD. Murine CAD carries 14 cysteines, most of which were found to be in reduced form. Reducing agents enhanced the production of the functional CAD in an in vitro translation system. The denatured CAD could be efficiently renatured under highly reducing conditions only in the presence of ICAD-L. This process was ATP-independent. In contrast, reticulocyte lysates stimulated ICAD-L- and ATP dependent renaturation of denatured CAD without requiring a high concentration of reducing agents. These results indicate that ICAD-L works not only as a specific inhibitor but also as a specific chaperone for CAD. PMID- 10713131 TI - In the human malaria parasite Plasmodium falciparum, polyamines are synthesized by a bifunctional ornithine decarboxylase, S-adenosylmethionine decarboxylase. AB - The polyamines putrescine, spermidine, and spermine are crucial for cell differentiation and proliferation. Interference with polyamine biosynthesis by inhibition of the rate-limiting enzymes ornithine decarboxylase (ODC) and S adenosylmethionine decarboxylase (AdoMetDC) has been discussed as a potential chemotherapy of cancer and parasitic infections. Usually both enzymes are individually transcribed and highly regulated as monofunctional proteins. We have isolated a cDNA from the malaria parasite Plasmodium falciparum that encodes both proteins on a single open reading frame, with the AdoMetDC domain in the N terminal region connected to a C-terminal ODC domain by a hinge region. The predicted molecular mass of the entire transcript is 166 kDa. The ODC/AdoMetDC coding region was subcloned into the expression vector pASK IBA3 and transformed into the AdoMetDC- and ODC-deficient Escherichia coli cell line EWH331. The resulting recombinant protein exhibited both AdoMetDC and ODC activity and co eluted after gel filtration on Superdex S-200 at approximately 333 kDa, which is in good agreement with the molecular mass of approximately 326 kDa determined for the native protein from isolated P. falciparum. SDS-polyacrylamide gel electrophoresis analysis of the recombinant ODC/AdoMetDC revealed a heterotetrameric structure of the active enzyme indicating processing of the AdoMetDC domain. The data presented describe the occurrence of a unique bifunctional ODC/AdoMetDC in P. falciparum, an organization which is possibly exploitable for the design of new antimalarial drugs. PMID- 10713132 TI - Roles of topoisomerases in maintaining steady-state DNA supercoiling in Escherichia coli. AB - DNA supercoiling is essential for bacterial cell survival. We demonstrated that DNA topoisomerase IV, acting in concert with topoisomerase I and gyrase, makes an important contribution to the steady-state level of supercoiling in Escherichia coli. Following inhibition of gyrase, topoisomerase IV alone relaxed plasmid DNA to a final supercoiling density (sigma) of -0.015 at an initial rate of 0.8 links min(-1). Topoisomerase I relaxed DNA at a faster rate, 5 links min(-1), but only to a sigma of -0.05. Inhibition of topoisomerase IV in wild-type cells increased supercoiling to approximately the same level as in a mutant lacking topoisomerase I activity (to sigma = -0.08). The role of topoisomerase IV was revealed by two functional assays. Removal of both topoisomerase I and topoisomerase IV caused the DNA to become hyper-negatively supercoiled (sigma = -0.09), greatly stimulating transcription from the supercoiling sensitive leu-500 promoter and increasing the number of supercoils trapped by lambda integrase site-specific recombination. PMID- 10713134 TI - Inhibition of homodimerization of poly(ADP-ribose) polymerase by its C-terminal cleavage products produced during apoptosis. AB - The biochemical role of the C-terminal fragment of poly(ADP-ribose) polymerase (PARP) was investigated in HeLa cells undergoing UV-mediated apoptosis. During the course of apoptosis, the C-terminal cleavage product of PARP interacted with intact PARP and down-regulated PARP activity by blocking the homodimerization of PARP. The basic leucine zipper motif in the auto-modification domain of the C terminal fragment of PARP represented the site of association, and Leu(405) was critical to the ability of the basic leucine zipper motif to associate with intact PARP. The expression of the C-terminal fragment of PARP stimulated UV mediated apoptosis. These results suggest that the C-terminal cleavage product of PARP produced during apoptosis blocks the homodimerization of PARP and inhibits the cellular PARP activity. The inhibition of the cellular PARP activity might prevent cellular NAD(+) depletion and stimulate apoptosis by maintaining the basal cellular energy level required for the completion of apoptosis. PMID- 10713133 TI - Yin-yang 1 and glucocorticoid receptor participate in the Stat5-mediated growth hormone response of the serine protease inhibitor 2.1 gene. AB - A growth hormone-inducible nuclear factor complex (GHINF), affinity-purified using the growth hormone response element (GHRE) from the promoter of rat serine protease inhibitor 2.1, was found to contain Stat5a and -5b, as well as additional components. The ubiquitous transcription factor yin-yang 1 (YY1) is present in GHINF. An antibody to YY1 inhibited the formation of the GHINF.GHRE complex in an electrophoretic mobility shift assay. Furthermore, Stat5 was co immunoprecipitated from rat hepatic nuclear extracts with antibodies to YY1. An examination of the GHRE shows that, in addition to two gamma-activated sites, it contains a putative YY1 binding site between the two gamma-activated sites, overlapping them both. Mutation of this putative YY1 site results in a decrease of GHINF.GHRE complex formation in an electrophoretic mobility shift assay and a corresponding decrease in growth hormone (GH) response in functional assays. The glucocorticoid receptor was also present in GHINF, and Stat5 co immunoprecipitates with glucocorticoid receptor in hepatic nuclear extracts from rats treated with GH. GH activation of serine protease inhibitor 2.1 requires the unique sequence of the GHRE encompassing the recognition sites of several transcription factors, and the interaction of these factors enhances the assembly of the transcription complex. PMID- 10713135 TI - Cloning and function of rabbit peroxisome proliferator-activated receptor delta/beta in mature osteoclasts. AB - Osteoclasts modulate bone resorption under physiological and pathological conditions. Previously, we showed that both estrogens and retinoids regulated osteoclastic bone resorption and postulated that such regulation was directly mediated through their cognate receptors expressed in mature osteoclasts. In this study, we searched for expression of other members of the nuclear hormone receptor superfamily in osteoclasts. Using the low stringency homologous hybridization method, we isolated the peroxisome proliferator-activated receptor delta/beta (PPARdelta/beta) cDNA from mature rabbit osteoclasts. Northern blot analysis showed that PPARdelta/beta mRNA was highly expressed in highly enriched rabbit osteoclasts. Carbaprostacyclin, a prostacyclin analogue known to be a ligand for PPARdelta/beta, significantly induced both bone-resorbing activities of isolated mature rabbit osteoclasts and mRNA expression of the cathepsin K, carbonic anhydrase type II, and tartrate-resistant acid phosphatase genes in these cells. Moreover, the carbaprostacyclin-induced bone resorption was completely blocked by an antisense phosphothiorate oligodeoxynucleotide of PPARdelta/beta but not by the sense phosphothiorate oligodeoxynucleotide of the same DNA sequence. Our results suggest that PPARdelta/beta may be involved in direct modulation of osteoclastic bone resorption. PMID- 10713136 TI - Basic fibroblast growth factor stimulates surface expression and activity of Na(+)/H(+) exchanger NHE3 via mechanism involving phosphatidylinositol 3-kinase. AB - Na(+)/H(+) exchanger NHE3 is a plasma membrane (PM) protein, which contributes to Na(+) absorption in the intestine. Growth factors stimulate NHE3 via phosphatidylinositol 3-kinase (PI3-K), but mechanism of this process is not clear. To examine the hypothesis that growth factors stimulate NHE3 by modulating NHE3 recycling, and that PI3-K participates in this mechanism, we used PS120 fibroblasts expressing a fusion protein of NHE3 and green fluorescent protein. At steady state, approximately 25% of cellular NHE3 content was expressed at PM. Inhibition of PI3-K decreased PM expression of NHE3, which correlated with retention of the exchanger in recycling endosomal compartment. In contrast, basic fibroblast growth factor (bFGF) increased PM expression of NHE3, which was associated with a 2-fold increase in rate constant for exit of the exchanger from the recycling compartment. Qualitatively similar effects of bFGF were observed in cells pretreated with PI3-K inhibitors, but their magnitude was only approximately 50% of that in intact cells. These data suggest that: (i) bFGF stimulates NHE3 by increasing PM expression of the exchanger; (ii) PI3-K mediates PM expression of NHE3 in both basal and bFGF-stimulated conditions, and (iii) not all of the effects of bFGF on NHE3 expression are mediated by PI3-K, suggesting additional regulatory mechanisms. PMID- 10713137 TI - Ubiquitination of the PEST-like endocytosis signal of the yeast a-factor receptor. AB - A 58-residue-long, PEST-like sequence within the yeast a-factor receptor (Ste3p) specifies the ubiquitination, endocytosis, and consequent vacuolar degradation of the receptor protein (Roth, A. F., Sullivan, D. M., and Davis, N. G. (1998) J. Cell Biol. 142, 949-961). The present work investigates three lysyl residues that map within this sequence as the potential ubiquitin acceptor sites. Lys --> Arg substitution mutants were tested for effects on both ubiquitination and endocytosis. Results indicate that the three lysines function redundantly; a severe blockade to both ubiquitination and endocytosis is seen only for receptors having all three lysines replaced. Of the three, Lys(432) plays the predominant role; ubiquitination and turnover are significantly impaired for receptors having just the K432R mutation. CNBr fragmentation of the receptor protein, used for the physical mapping of the ubiquitin attachment sites, showed PEST-like sequence lysines to be modified both with single ubiquitin moieties as well with short multi-ubiquitin chains, two or three ubiquitins long. Thus, in addition to being the signal for ubiquitination, the Ste3p PEST-like sequence also provides the site for ubiquitin attachment. To test if this endocytosis signal functions solely for ubiquitination, we have asked if the requirement for the PEST-like sequence in endocytosis might be bypassed through pre-attachment of ubiquitin to the receptor protein. Indeed, Ste3-ubiquitin translational fusions that have a ubiquitin moiety fused to the receptor in place of the PEST-like signal do undergo rapid endocytosis and vacuolar turnover. We conclude that ubiquitin alone, with no required contribution from receptor sequences, provides the sufficient signal for initiating uptake. In addition, our results confirm conclusions originally drawn from studies with the alpha-factor receptor (Terrell, J., Shih, S., Dunn, R., and Hicke, L. (1998) Mol. Cell 1, 193-202), namely that mono-ubiquitin, and not multi-ubiquitin chains provide the primary recognition determinant for uptake. Although mono-ubiquitination suffices, our results indicate that multi-ubiquitination serves to augment the rate of uptake. PMID- 10713138 TI - ARF1 regulates pH-dependent COP functions in the early endocytic pathway. AB - Coat proteins of the COP family were recently shown by us and others to be involved in membrane transport in the endocytic pathway, in addition to their known functions in the biosynthetic pathway. We have also shown that membrane association of endosomal COPs depends on the acidic endosomal pH, in contrast to biosynthetic COPs. In this paper, we report that both membrane recruitment of endosomal COPs and in vitro biogenesis of transport intermediates destined for late endosomes, depend on a cytosolic factor, which we identified as the small GTP-binding protein ARF1. Our data indicate that ARF1 does not act via activation of an endosomal phospholipase D. We also find that ARF1 membrane association is regulated by the endosomal pH, and that this controls the pH-dependent association of endosomal COPs. These studies thus show that ARF1 regulates COP functions in the endocytic pathway, and indicate that ARF1 acts as the cytosolic component of a transmembrane pH-sensing mechanism. PMID- 10713139 TI - Novel dual repressor elements for neuronal cell-specific transcription of the rat 5-HT1A receptor gene. AB - The level of expression of the 5-HT1A receptor in the raphe and limbic systems is implicated in the etiology and treatment of major depression and anxiety disorders. The rat 5-HT1A receptor gene is regulated by a proximal TATA-driven promoter and by upstream repressors that inhibit gene expression. Deletion of a 71-base pair (bp) segment between -1590/-1519 bp of the 5-HT1A receptor gene induced over 10-fold enhancement of transcriptional activity in both 5-HT1A receptor-expressing (RN46A raphe and SN48 septal) cells and receptor-negative (L6 myoblast and C6 glioma) cells. A 31-bp segment of the repressor was protected from DNase I digestion by RN46A or L6 nuclear extracts. Within the 31-bp segment, a single protein complex was present in receptor-expressing cells that bound a novel 14-bp DNA element; in receptor-negative cells, an additional complex bound an adjacent 12-bp sequence. In receptor-positive but not receptor-negative cells, mutation of the 14-bp element to eliminate protein binding abrogated repression to nearly the same extent as deletion of the -1590/-1519 bp segment. Additional mutation of both 14-bp and 12-bp elements abolished protein binding and repressor activity in receptor-negative cells. Thus a single protein-DNA complex at the 14 bp element represses the 5-HT1A receptor gene in 5-HT1A receptor-positive neuronal cells, whereas adjacent DNA elements provide a dual repression mechanism in 5-HT1A receptor-negative cells. PMID- 10713140 TI - Mutations at critical N-glycosylation sites reduce tyrosinase activity by altering folding and quality control. AB - Tyrosinase is a copper-containing enzyme that regulates melanin biosynthesis in mammals. Mutations at a single N-glycosylation sequon of tyrosinase have been reported to be responsible for oculocutaneous albinism type IA in humans, characterized by inactive tyrosinase and the total absence of pigmentation. To probe the role that each N-glycosylation site plays in the synthesis of biologically active tyrosinase, we analyzed the calnexin mediated folding of tyrosinase N-glycosylation mutants. We have determined that four of the six potential N-glycosylation sites, including that associated with albinism, are occupied. Analysis of the folding pathway and activity of 15 tyrosinase mutants lacking one or more of the occupied N-glycosylation sites shows that glycans at any two N-glycosylation sites are sufficient to interact with calnexin and give partial activity, but a specific pair of sites (Asn(86) and Asn(371)) is required for full activity. The mutants with less than two N-glycosylation sites do not interact with calnexin and show a complete absence of enzyme activity. Copper analysis of selected mutants suggests that the observed partial activity is due to two populations with differential copper content. By correlating the degree of folding with the activity of tyrosinase, we propose a local folding mechanism for tyrosinase that can explain the mechanism of inactivation of tyrosinase N glycosylation mutants found in certain pigmentation disorders. PMID- 10713141 TI - Inhibition of laminin-5 production in breast epithelial cells by overexpression of p300. AB - The transcriptional coactivator p300 is essential for normal embryonic development and cellular differentiation. We have been studying the role of p300 in the transcription of a variety of genes, and we became interested in the role of this coactivator in the transcription of genes important in breast epithelial cell biology. From MCF-10A cells (spontaneously immortalized, nontransformed human breast epithelial cells), we developed cell lines that stably overexpress p300. These p300-overexpressing cells displayed reduced adhesion to culture dishes and were found to secrete an extracellular matrix deficient in laminin-5. Laminin-5 is the major extracellular matrix component produced by breast epithelium. Immunofluorescence studies, as well as experiments using normal matrix, confirmed that the decreased adhesion of p300-overexpressing cells is due to laminin-5-deficient extracellular matrix and not due to loss of laminin-5 receptors. Northern blots revealed markedly decreased levels of expression of two of the genes (designated LAMA3 and LAMC2) encoding the alpha3 and gamma2 chains of the laminin-5 heterotrimer in the cells that overexpress p300, whereas LAMB3 mRNA, encoding the third or beta3 chain of laminin-5, was not markedly reduced. Transient transfection experiments with a vector containing a murine LAMA3 promoter demonstrate that overexpressing p300 down-regulates the LAMA3 promoter. In summary, overexpression of p300 leads to down-regulation of laminin-5 production in breast epithelial cells, resulting in decreased adhesion. PMID- 10713142 TI - Molecular basis of cell-specific endothelial nitric-oxide synthase expression in airway epithelium. AB - Nitric oxide (NO) plays an important role in airway function, and endothelial NO synthase (eNOS) is expressed in airway epithelium. To determine the basis of cell specific eNOS expression in airway epithelium, studies were performed in NCI-H441 human bronchiolar epithelial cells transfected with the human eNOS promoter fused to luciferase. Transfection with 1624 base pairs of sequence 5' to the initiation ATG (position -1624) yielded a 19-fold increase in promoter activity versus vector alone. No activity was found in lung fibroblasts, which do not express eNOS. 5' deletions from -1624 to -279 had modest effects on promoter activity in H441 cells. Further deletion to -248 reduced activity by 65%, and activity was lost with deletion to -79. Point mutations revealed that the GATA binding motif at -254 is mandatory for promoter activity and that the positive regulatory element between -248 and -79 is the Sp1 binding motif at -125. Electrophoretic mobility shift assays yielded two complexes with the GATA site and three with the Sp1 site. Immunodepletion with antiserum to GATA-2 prevented formation of the slowest migrating GATA complex, and antiserum to Sp1 supershifted the slowest migrating Sp1 complex. An electrophoretic mobility shift assay with H441 versus fibroblast nuclei revealed that the slowest migrating GATA complex is unique to airway epithelium. Thus, cell-specific eNOS expression in airway epithelium is dependent on the interaction of GATA-2 with the core eNOS promoter, and the proximal Sp1 binding site is also an important positive regulatory element. PMID- 10713143 TI - Histone octamer dissociation is not required for in vitro replication of simian virus 40 minichromosomes. AB - Replication of chromosomal templates requires the passage of the replication machinery through nucleosomally organized DNA. To gain further insights into these processes we have used chromatin that was reconstituted with dimethyl suberimidate-cross-linked histone octamers as template in the SV40 in vitro replication system. By supercoiling analysis we found that cross-linked histone octamers were reconstituted with the same kinetic and efficiency as control octamers. Minichromosomes with cross-linked nucleosomes were completely replicated, although the efficiency of replication was lower compared with control chromatin. Analysis of the chromatin structure of the replicated DNA revealed that the cross-linked octamer is transferred to the daughter strands. Thus, our data imply that histone octamer dissociation is not a prerequisite for the passage of the replication machinery and the transfer of the parental nucleosomes. PMID- 10713144 TI - Purification and characterization of DnaC810, a primosomal protein capable of bypassing PriA function. AB - Escherichia coli strains lacking PriA are severely compromised in their ability to repair UV-damaged DNA and to perform homologous recombination. These phenotypes arise because of a lack of PriA-directed replication fork assembly at recombination intermediates such as D-loops. Naturally arising suppressor mutations in dnaC restore strains carrying the priA2::kan null allele to wild type function. We have cloned one such gene, dnaC810, and overexpressed, purified, and characterized the DnaC810 protein. DnaC810 can support a PriA independent synthesis of phiX174 complementary strand DNA. This can be attributed to its ability, unlike wild-type DnaC, to catalyze a SSB-insensitive general priming reaction with DnaB and DnaG on any SSB-coated single-stranded DNA. Gel mobility shift analysis revealed that DnaC810 could load DnaB directly to SSB coated single-stranded DNA as well as to D loop DNA. This explains the ability of DnaC810 to bypass the requirement for PriA, PriB, PriC, and DnaT during replication fork assembly at recombination intermediates. PMID- 10713145 TI - Cloning and characterization of a new isoform of skeletal muscle triadin. AB - We have shown that several isoforms of triadin, a protein involved in calcium release process through the ryanodine receptor, are expressed in rat skeletal muscle, and we have cloned two of these isoforms. One is the rat homolog of the 95-kDa triadin identified in rabbit skeletal muscle, and the second one, shorter, is a truncated form of the previous one, but with a new unique COOH-terminal end. We propose to name the two proteins identified here Trisk 95 and Trisk 51. We have produced antibodies specific to each isoform. Using these antibodies, we have shown that the newly identified protein, Trisk 51, is actually expressed in adult rat skeletal muscle and also in rat embryo skeletal muscle. Immunofluorescent labeling of rat skeletal muscle with anti-Trisk 95, anti-Trisk 51, or anti-ryanodine receptor antibodies shows a similar localization of these proteins, in the tissue. Transfection of L6 cells with cDNA of Trisk 51 or Trisk 95 leads to the expression of proteins with the expected molecular weight, identical to those detected in rat skeletal muscle. Both proteins appear during differentiation of satellite cells in myotubes which may indicate the involvement of these two isoforms in the building of a functional calcium release machinery. PMID- 10713146 TI - Stoichiometry of P1 plasmid partition complexes. AB - The P1 plasmid prophage is faithfully partitioned by a high affinity nucleoprotein complex assembled at the centromere-like parS site. This partition complex is composed of P1 ParB and Escherichia coli integration host factor (IHF), bound specifically to parS. We have investigated the assembly of ParB at parS and its stoichiometry of binding. Measured by gel mobility shift assays, ParB and IHF bind tightly to parS and form a specific complex, called I + B1. We observed that as ParB concentration was increased, a second, larger complex (I + B2) formed, followed by the formation of larger complexes, indicating that additional ParB molecules joined the initial complex. Shift Western blotting experiments indicated that the I + B2 complex contained twice as much ParB as the I + B1 complex. Using mixtures of ParB and a larger polyhistidine-tagged version of ParB (His-ParB) in DNA binding assays, we determined that the initial I + B1 complex contains one dimer of ParB. Therefore, one dimer of ParB binds to its recognition sequences that span an IHF-directed bend in parS. Once this complex forms, a second dimer can join the complex, but this assembly requires much higher ParB concentrations. PMID- 10713147 TI - Distinct mitochondrial and cytosolic enzymes mediate trypanothione-dependent peroxide metabolism in Trypanosoma cruzi. AB - The American trypanosome Trypanosoma cruzi is exposed to toxic oxygen metabolites that are generated by drug metabolism and immune responses in addition to those produced by endogenous processes. However, much remains to be resolved about the parasite oxidative defense system, including the mechanism(s) of peroxide reduction. Here we show that reduction of peroxides in T. cruzi is catalyzed by two distinct trypanothione-dependent enzymes. These were localized to the cytosol and mitochondrion. Both are members of the peroxiredoxin family of antioxidant proteins and are characterized by the presence of two conserved domains containing redox active cysteines. The role of these proteins in protecting T. cruzi from peroxide-mediated damage was demonstrated following overexpression of enzyme activity. The parasite-specific features of T. cruzi cytoplasmic peroxiredoxin and T. cruzi mitochondrial peroxiredoxin may be exploitable in terms of drug development. PMID- 10713148 TI - Cleavage preferences of the apoptotic endonuclease DFF40 (caspase-activated DNase or nuclease) on naked DNA and chromatin substrates. AB - Here we report the co-factor requirements for DNA fragmentation factor (DFF) endonuclease and characterize its cleavage sites on naked DNA and chromatin substrates. The endonuclease exhibits a pH optimum of 7.5, requires Mg(2+), not Ca(2+), and is inhibited by Zn(2+). The enzyme generates blunt ends or ends with 1-base 5'-overhangs possessing 5'-phosphate and 3'-hydroxyl groups and is specific for double- and not single-stranded DNA or RNA. DFF endonuclease has a moderately greater sequence preference than micrococcal nuclease or DNase I, and the sites attacked possess a dyad axis of symmetry with respect to purine and pyrimidine content. Using HeLa cell nuclei or chromatin reconstituted on a 5 S rRNA gene tandem array, we prove that the enzyme attacks chromatin in the internucleosomal linker, generating oligonucleosomal DNA ladders sharper than those created by micrococcal nuclease. Histone H1, high mobility group-1, and topoisomerase II activate DFF endonuclease activity on naked DNA substrates but much less so on chromatin substrates. We conclude that DFF is a useful reagent for chromatin research. PMID- 10713149 TI - Specificity of DNA lesion bypass by the yeast DNA polymerase eta. AB - DNA polymerase eta (Pol(eta), xeroderma pigmentosum variant, or Rad30) plays an important role in an error-free response to unrepaired UV damage during replication. It faithfully synthesizes DNA opposite a thymine-thymine cis-syn cyclobutane dimer. We have purified the yeast Pol(eta) and studied its lesion bypass activity in vitro with various types of DNA damage. The yeast Pol(eta) lacked a nuclease or a proofreading activity. It efficiently bypassed 8 oxoguanine, incorporating C, A, and G opposite the lesion with a relative efficiency of approximately 100:56:14, respectively. The yeast Pol(eta) efficiently incorporated a C opposite an acetylaminofluorene-modified G, and efficiently inserted a G or less frequently an A opposite an apurinic/apyrimidinic (AP) site but was unable to extend the DNA synthesis further in both cases. However, some continued DNA synthesis was observed in the presence of the yeast Pol(zeta) following the Pol(eta) action opposite an AP site, achieving true lesion bypass. In contrast, the yeast Pol(alpha) was able to bypass efficiently a template AP site, predominantly incorporating an A residue opposite the lesion. These results suggest that other than UV damage, Pol(eta) may also play a role in bypassing additional DNA lesions, some of which can be error-prone. PMID- 10713150 TI - Role of SNAP23 in insulin-induced translocation of GLUT4 in 3T3-L1 adipocytes. Mediation of complex formation between syntaxin4 and VAMP2. AB - Both syntaxin4 and VAMP2 are implicated in insulin regulation of glucose transporter-4 (GLUT4) trafficking in adipocytes as target (t) soluble N ethylmaleimide-sensitive factor attachment protein receptors (SNARE) and vesicle (v)-SNARE proteins, respectively, which mediate fusion of GLUT4-containing vesicles with the plasma membrane. Synaptosome-associated 23-kDa protein (SNAP23) is a widely expressed isoform of SNAP25, the principal t-SNARE of neuronal cells, and colocalizes with syntaxin4 in the plasma membrane of 3T3-L1 adipocytes. In the present study, two SNAP23 mutants, SNAP23-DeltaC8 (amino acids 1 to 202) and SNAP23-DeltaC49 (amino acids 1 to 161), were generated to determine whether SNAP23 is required for insulin-induced translocation of GLUT4 to the plasma membrane in 3T3-L1 adipocytes. Wild-type SNAP23 (SNAP23-WT) promoted the interaction between syntaxin4 and VAMP2 both in vitro and in vivo. Although SNAP23-DeltaC49 bound to neither syntaxin4 nor VAMP2, the SNAP23-DeltaC8 mutant bound to syntaxin4 but not to VAMP2. In addition, although SNAP23-DeltaC8 bound to syntaxin4, it did not mediate the interaction between syntaxin4 and VAMP2. Moreover, overexpression of SNAP23-DeltaC8 in 3T3-L1 adipocytes by adenovirus mediated gene transfer inhibited insulin-induced translocation of GLUT4 but not that of GLUT1. In contrast, overexpression of neither SNAP23-WT nor SNAP23 DeltaC49 in 3T3-L1 adipocytes affected the translocation of GLUT4 or GLUT1. Together, these results demonstrate that SNAP23 contributes to insulin-dependent trafficking of GLUT4 to the plasma membrane in 3T3-L1 adipocytes by mediating the interaction between t-SNARE (syntaxin4) and v-SNARE (VAMP2). PMID- 10713151 TI - A Ca(2+)-independent activation of a type IV cytosolic phospholipase A(2) underlies the receptor stimulation of arachidonic acid-dependent noncapacitative calcium entry. AB - The oscillatory [Ca(2+)](i) signals typically seen following physiologically relevant stimulation of phospholipase C-linked receptors are associated with a receptor-activated entry of Ca(2+), which plays a critical role in driving the oscillations and influencing their frequency. We have recently shown that this receptor-activated entry of Ca(2+) does not conform to the widely accepted "capacitative" model and, instead, reflects the activity of a distinct, novel Ca(2+) entry pathway regulated by arachidonic acid (Shuttleworth, T. J., and Thompson, J. L. (1998) J. Biol. Chem. 273, 32636-32643). We now show that the generation of arachidonic acid under these conditions results from the activity of a type IV cytosolic phospholipase A(2) (cPLA(2)). Although cPLA(2) activation commonly involves a Ca(2+)-dependent translocation to the membrane, at these low agonist concentrations cPLA(2) activation was independent of increases in [Ca(2+)](i), and no detectable translocation to the membrane occurs. Nevertheless, stimulation of cPLA(2) activity was confined to the membrane fraction, where an increase in phosphorylation of the enzyme was observed. We suggest that, at the low agonist concentrations associated with oscillatory [Ca(2+)](i) signals, cPLA(2) activation involves an increased phosphorylation of a discrete pool of the total cellular cPLA(2) that is already localized within the membrane fraction at resting [Ca(2+)](i). PMID- 10713152 TI - Distinct recognition of collagen subtypes by alpha(1)beta(1) and alpha(2)beta(1) integrins. Alpha(1)beta(1) mediates cell adhesion to type XIII collagen. AB - Two integrin-type collagen receptors, alpha(1)beta(1) and alpha(2)beta(1), are structurally very similar. However, cells can concomitantly express the both receptors and they might have independent functions. Here, Chinese hamster ovary (CHO) cells, which lack endogenous collagen receptors, were transfected with either alpha(1) or alpha(2) integrin cDNA. Cells were allowed to adhere to various collagen types and their integrin function was tested by observing the progression of cell spreading. The cells expressing alpha(1)beta(1) integrin could spread on collagen types I, III, IV, and V but not on type II, while alpha(2)beta(1) integrin could mediate cell spreading on collagen types I-V. Type XIII is a transmembrane collagen and its interaction with the integrins has not been previously studied. CHO-alpha1beta1 cells could spread on human recombinant type XIII collagen, unlike CHO-alpha2beta1 cells. Integrins alpha(1)beta(1) and alpha(2)beta(1) recognize collagens with the specific alphaI domains. The alpha(1)I and alpha(2)I domains were produced as recombinant proteins, labeled with europium and used in a sensitive solid-phase binding assay based on time resolved fluorescence. alpha(1)I domain, unlike the alpha(2)I domain, could attach to type XIII collagen. The results indicate, that alpha(1)beta(1) and alpha(2)beta(1) have different ligand binding specificity. Distinct recognition of different collagen subtypes by the alphaI domains can partially explain the differences seen in cell spreading. However, despite the fact that CHO alpha1beta1 cells could not spread on type II collagen alpha(1)I domain could bind to this collagen type. Thus, the cell spreading on collagens may also be regulated by factors other than the integrins. PMID- 10713154 TI - UTP-dependent and -independent pathways of mRNA turnover in Trypanosoma brucei mitochondria. AB - Although primary transcripts are polycistronic in the mitochondria of Trypanosoma brucei, steady-state levels of mature, monocistronic RNAs change throughout the parasitic life cycle. This indicates that steady-state RNA abundance is controlled by posttranscriptional mechanisms involving differential RNA stability. In this study, in organello pulse-chase labeling experiments were used to analyze the stability of different T. brucei mitochondrial RNA populations. In this system, total RNA and rRNA are stable for many hours. In contrast, mRNAs can be degraded by two biochemically distinct turnover pathways. The first pathway results in the rapid degradation of mRNA (half-life [t(1/2)] of 11 to 18 min) and is dependent upon the presence of an mRNA poly(A) tail. Remarkably, this pathway also requires the addition of UTP and therefore is termed UTP dependent. The second pathway results in slow turnover of mitochondrial mRNA (t(1/2) of approximately 3 h) and is not dependent upon the presence of an mRNA poly(A) tail or the addition of exogenous UTP. In summary, these results demonstrate the presence of a novel, UTP-dependent degradation pathway for T. brucei mitochondrial mRNAs and reveal an unprecedented role for both UTP and mRNA polyadenylation in T. brucei mitochondrial gene expression. PMID- 10713153 TI - Transcription-coupled translation control of AML1/RUNX1 is mediated by cap- and internal ribosome entry site-dependent mechanisms. AB - AML1/RUNX1 belongs to the runt domain transcription factors that are important regulators of hematopoiesis and osteogenesis. Expression of AML1 is regulated at the level of transcription by two promoters, distal (D) and proximal (P), that give rise to mRNAs bearing two distinct 5' untranslated regions (5'UTRs) (D-UTR and P-UTR). Here we show that these 5'UTRs act as translation regulators in vivo. AML1 mRNAs bearing the uncommonly long (1,631-bp) P-UTR are poorly translated, whereas those with the shorter (452-bp) D-UTR are readily translated. The low translational efficiency of the P-UTR is attributed to its length and the cis acting elements along it. Transfections and in vitro assays with bicistronic constructs demonstrate that the D-UTR mediates cap-dependent translation whereas the P-UTR mediates cap-independent translation and contains a functional internal ribosome entry site (IRES). The IRES-containing bicistronic constructs are more active in hematopoietic cell lines that normally express the P-UTR-containing mRNAs. Furthermore, we show that the IRES-dependent translation increases during megakaryocytic differentiation but not during erythroid differentiation, of K562 cells. These results strongly suggest that the function of the P-UTR IRES dependent translation in vivo is to tightly regulate the translation of AML1 mRNAs. The data show that AML1 expression is regulated through usage of alternative promoters coupled with IRES-mediated translation control. This IRES mediated translation regulation adds an important new dimension to the fine-tuned control of AML1 expression. PMID- 10713155 TI - A novel type of splicing enhancer regulating adenovirus pre-mRNA splicing. AB - Splicing of the adenovirus IIIa pre-mRNA is subjected to a temporal regulation, such that efficient IIIa 3' splice site usage is confined to the late phase of the infectious cycle. Here we show that IIIa pre-mRNA splicing is activated more than 200-fold in nuclear extracts prepared from late adenovirus-infected cells (Ad-NE) compared to uninfected HeLa cell nuclear extracts (HeLa-NE). In contrast, splicing of the beta-globin pre-mRNA is repressed in Ad-NE. We constructed hybrid pre-mRNAs between IIIa and beta-globin in order to identify the minimal IIIa sequence element conferring enhanced splicing in Ad-NE. Using this approach, we show that the IIIa branch site/pyrimidine tract functions as a Janus element: it blocks splicing in HeLa-NE and functions as a splicing enhancer in Ad-NE. Therefore, we named this sequence the IIIa virus infection-dependent splicing enhancer (3VDE). This element is essential for regulated IIIa pre-mRNA splicing in Ad-NE and sufficient to confer an enhanced splicing phenotype to the beta globin pre-mRNA in Ad-NE. We further show that the increase in IIIa splicing observed in Ad-NE is not accompanied by a similar increase in U2AF binding to the IIIa pyrimidine tract. This finding suggests that splicing activation by the 3VDE may operate without efficient U2AF interaction with the pre-mRNA. Importantly, this report represents the first description of a splicing enhancer that has evolved to function selectively in the context of a virus infection, a finding that adds a new level at which viruses may subvert the host cell RNA biosynthetic machinery to facilitate their own replication. PMID- 10713156 TI - Nedd8 modification of cul-1 activates SCF(beta(TrCP))-dependent ubiquitination of IkappaBalpha. AB - Regulation of NF-kappaB occurs through phosphorylation-dependent ubiquitination of IkappaBalpha, which is degraded by the 26S proteasome. Recent studies have shown that ubiquitination of IkappaBalpha is carried out by a ubiquitin-ligase enzyme complex called SCF(beta(TrCP)). Here we show that Nedd8 modification of the Cul-1 component of SCF(beta(TrCP)) is important for function of SCF(beta(TrCP)) in ubiquitination of IkappaBalpha. In cells, Nedd8-conjugated Cul 1 was complexed with two substrates of SCF(beta(TrCP)), phosphorylated IkappaBalpha and beta-catenin, indicating that Nedd8-Cul-1 conjugates are part of SCF(beta(TrCP)) in vivo. Although only a minute fraction of total cellular Cul-1 is modified by Nedd8, the Cul-1 associated with ectopically expressed betaTrCP was highly enriched for the Nedd8-conjugated form. Moreover, optimal ubiquitination of IkappaBalpha required Nedd8 and the Nedd8-conjugating enzyme, Ubc12. The site of Nedd8 ligation to Cul-1 is essential, as SCF(beta(TrCP)) containing a K720R mutant of Cul-1 only weakly supported IkappaBalpha ubiquitination compared to SCF(beta(TrCP)) containing WT Cul-1, suggesting that the Nedd8 ligation of Cul-1 affects the ubiquitination activity of SCF(beta(TrCP)). These observations provide a functional link between the highly related ubiquitin and Nedd8 pathways of protein modification and show how they operate together to selectively target the signal-dependent degradation of IkappaBalpha. PMID- 10713157 TI - Synergistic interaction of MEK kinase 2, c-Jun N-terminal kinase (JNK) kinase 2, and JNK1 results in efficient and specific JNK1 activation. AB - Mitogen-activated protein kinases (MAPKs) are activated through cascades or modules consisting of a MAPK, a MAPK kinase (MAPKK), and a MAPKK kinase (MAPKKK). Investigating the molecular basis of activation of the c-Jun N-terminal kinase (JNK) subgroup of MAPK by the MAPKKK MEKK2, we found that strong and specific JNK1 activation by MEKK2 was mediated by the MAPKK JNK kinase 2 (JNKK2) rather than by JNKK1 through formation of a tripartite complex consisting of MEKK2, JNKK2, and JNK1. No scaffold protein was required for the MEKK2-JNKK2-JNK1 tripartite-complex formation. Expression of JNK1, JNKK2, and MEKK2 significantly augmented the coprecipitation of, respectively, MEKK2-JNKK2, MEKK2-JNK1, and JNKK2-JNK1, indicating that the interaction of MEKK2, JNKK2, and JNK1 is synergistic. Finally, the JNK1 was activated more efficiently in the MEKK2-JNKK2 JNK1 complex than was the JNK1 excluded from the complex. Thus, formation of a signaling complex through synergistic interaction of a MAPKKK, a MAPKK, and a MAPK molecule like MEKK2-JNKK2-JNK1 is likely to be responsible for the efficient, specific flow of information via MAPK cascades. PMID- 10713158 TI - Modulation of DNA binding protein affinity directly affects target site demethylation. AB - It has recently been shown that in Xenopus, DNA demethylation at promoter regions may involve protein-DNA interactions, based on the specificity of the demethylated sites. Utilizing a stable episomal system in human cells, we recently mapped the sites and dissected the steps of demethylation at oriP sites bound by EBNA1 protein. Although it is clear that protein binding is required for demethylation of the oriP sites, it is uncertain whether this is a unique feature of the replication origin or whether it is a general phenomenon for all DNA sequences to which sequence-specific proteins are bound. In the present study, we utilize the well-defined Escherichia coli lac repressor/operator system in human cells to determine whether protein binding to methylated DNA, in a region that is neither a replication origin nor a promoter, can also lead to demethylation of the binding sites. We found that demethylation specified by protein binding is not unique to the replication origin or to the promoter. We also found that transcriptional activity does not influence demethylation of the lac operator. Isopropyl-beta-D-thiogalactopyranoside (IPTG), an inhibitor of the lac repressor, can prevent demethylation of the lac operator DNA sites and can modulate demethylation of the lac operator by affecting the binding affinity of the lac repressor. Using this system, a titration of protein binding can be done. This titration permits one to infer that protein binding site occupancy is the determinant of demethylation at DNA sites and permits a determination of how this process progresses over time. PMID- 10713159 TI - Architectural transcription factors and the SAGA complex function in parallel pathways to activate transcription. AB - Recent work has shown that transcription of the yeast HO gene involves the sequential recruitment of a series of transcription factors. We have performed a functional analysis of HO regulation by determining the ability of mutations in SIN1, SIN3, RPD3, and SIN4 negative regulators to permit HO expression in the absence of certain activators. Mutations in the SIN1 (=SPT2) gene do not affect HO regulation, in contrast to results of other studies using an HO:lacZ reporter, and our data show that the regulatory properties of an HO:lacZ reporter differ from that of the native HO gene. Mutations in SIN3 and RPD3, which encode components of a histone deacetylase complex, show the same pattern of genetic suppression, and this suppression pattern differs from that seen in a sin4 mutant. The Sin4 protein is present in two transcriptional regulatory complexes, the RNA polymerase II holoenzyme/mediator and the SAGA histone acetylase complex. Our genetic analysis allows us to conclude that Swi/Snf chromatin remodeling complex has multiple roles in HO activation, and the data suggest that the ability of the SBF transcription factor to bind to the HO promoter may be affected by the acetylation state of the HO promoter. We also demonstrate that the Nhp6 architectural transcription factor, encoded by the redundant NHP6A and NHP6B genes, is required for HO expression. Suppression analysis with sin3, rpd3, and sin4 mutations suggests that Nhp6 and Gcn5 have similar functions. A gcn5 nhp6a nhp6b triple mutant is extremely sick, suggesting that the SAGA complex and the Nhp6 architectural transcription factors function in parallel pathways to activate transcription. We find that disruption of SIN4 allows this strain to grow at a reasonable rate, indicating a critical role for Sin4 in detecting structural changes in chromatin mediated by Gcn5 and Nhp6. These studies underscore the critical role of chromatin structure in regulating HO gene expression. PMID- 10713160 TI - Repression of CDK1 and other genes with CDE and CHR promoter elements during DNA damage-induced G(2)/M arrest in human cells. AB - Entry into mitosis is controlled by the cyclin-dependent kinase CDK1 and can be delayed in response to DNA damage. In some systems, such G(2)/M arrest has been shown to reflect the stabilization of inhibitory phosphorylation sites on CDK1. In human cells, full G(2) arrest appears to involve additional mechanisms. We describe here the prolonged (>6 day) downregulation of CDK1 protein and mRNA levels following DNA damage in human cells. This silencing of gene expression is observed in primary human fibroblasts and in two cell lines with functional p53 but not in HeLa cells, where p53 is inactive. Silencing is accompanied by the accumulation of cells in G(2), when CDK1 expression is normally maximal. The response is impaired by mutations in cis-acting elements (CDE and CHR) in the CDK1 promoter, indicating that silencing occurs at the transcriptional level. These elements have previously been implicated in the repression of transcription during G(1) that is normally lifted as cells progress into S and G(2). Interestingly, we find that other genes, including those for CDC25C, cyclin A2, cyclin B1, CENP-A, and topoisomerase IIalpha, that are normally expressed preferentially in G(2) and whose promoter regions include putative CDE and CHR elements are also downregulated in response to DNA damage. These data, together with those of other groups, support the existence of a p53-dependent, DNA damage activated pathway leading to CHR- and CDE-mediated transcriptional repression of various G(2)-specific genes. This pathway may be required for sustained periods of G(2) arrest following DNA damage. PMID- 10713161 TI - The yeast ULP2 (SMT4) gene encodes a novel protease specific for the ubiquitin like Smt3 protein. AB - Yeast Smt3 and its vertebrate homolog SUMO-1 are ubiquitin-like proteins (Ubls) that are reversibly ligated to other proteins. Like SMT3, SMT4 was first isolated as a high-copy-number suppressor of a defective centromere-binding protein. We show here that SMT4 encodes an Smt3-deconjugating enzyme, Ulp2. In cells lacking Ulp2, specific Smt3-protein conjugates accumulate, and the conjugate pattern is distinct from that observed in a ulp1(ts) strain, which is defective for a distantly related Smt3-specific protease, Ulp1. The ulp2Delta mutant exhibits a pleiotropic phenotype that includes temperature-sensitive growth, abnormal cell morphology, decreased plasmid and chromosome stability, and a severe sporulation defect. The mutant is also hypersensitive to DNA-damaging agents, hydroxyurea, and benomyl. Although cell cycle checkpoint arrest in response to DNA damage, replication inhibition, or spindle defects occurs with normal kinetics, recovery from arrest is impaired. Surprisingly, either introduction of a ulp1(ts) mutation or overproduction of catalytically inactive Ulp1 can substantially overcome the ulp2Delta defects. Inactivation of Ulp2 also suppresses several ulp1(ts) defects, and the double mutant accumulates far fewer Smt3-protein conjugates than either single mutant. Our data suggest the existence of a feedback mechanism that limits Smt3-protein ligation when Smt3 deconjugation by both Ulp1 and Ulp2 is compromised, allowing a partial recovery of cell function. PMID- 10713162 TI - Involvement of the checkpoint protein Mec1p in silencing of gene expression at telomeres in Saccharomyces cerevisiae. AB - Yeast strains with a mutation in the MEC1 gene are deficient in the cellular checkpoint response to DNA-damaging agents and have short telomeres (K. B. Ritchie, J. C. Mallory, and T. D. Petes, Mol. Cell. Biol. 19:6065-6075, 1999; T. A. Weinert, G. L. Kiser, and L. H. Hartwell, Genes Dev. 8:652-665, 1994). In wild type yeast cells, genes inserted near the telomeres are transcriptionally silenced (D. E. Gottschling, O. M. Aparichio, B. L. Billington, and V. A. Zakian, Cell 63:751-762, 1990). We show that mec1 strains have reduced ability to silence gene expression near the telomere. This deficiency was alleviated by the sml1 mutation. Overexpression of Mec1p also resulted in a silencing defect, although this overexpression did not affect the checkpoint function of Mec1p. Telomeric silencing was not affected by mutations in several other genes in the Mec1p checkpoint pathway (null mutations in RAD9 and CHK1 or in several hypomorphic rad53 alleles) but was reduced by a null mutation of DUN1. In addition, the loss of telomeric silencing in mec1 strains was not a consequence of the slightly shortened telomeres observed in these strains. PMID- 10713163 TI - Impaired core promoter recognition caused by novel yeast TAF145 mutations can be restored by creating a canonical TATA element within the promoter region of the TUB2 gene. AB - The general transcription factor TFIID, which is composed of TATA-binding protein (TBP) and an array of TBP-associated factors (TAFs), has been shown to play a crucial role in recognition of the core promoters of eukaryotic genes. We isolated Saccharomyces cerevisiae yeast TAF145 (yTAF145) temperature-sensitive mutants in which transcription of a specific subset of genes was impaired at restrictive temperatures. The set of genes affected in these mutants overlapped with but was not identical to the set of genes affected by a previously reported yTAF145 mutant (W.-C. Shen and M. R. Green, Cell 90:615-624, 1997). To identify sequences which rendered transcription yTAF145 dependent, we conducted deletion analysis of the TUB2 promoter using a novel mini-CLN2 hybrid gene reporter system. The results showed that the yTAF145 mutations we isolated impaired core promoter recognition but did not affect activation by any of the transcriptional activators we tested. These observations are consistent with the reported yTAF145 dependence of the CLN2 core promoter in the mutant isolated by Shen and Green, although the CLN2 core promoter functioned normally in the mutants we report here. These results suggest that different promoters require different yTAF145 functions for efficient transcription. Interestingly, insertion of a canonical TATA element into the TATA-less TUB2 promoter rescued impaired transcription in the yTAF145 mutants we studied. It therefore appears that strong binding of TBP to the core promoter can alleviate the requirement for at least one yTAF145 function. PMID- 10713164 TI - SKIP, a CBF1-associated protein, interacts with the ankyrin repeat domain of NotchIC To facilitate NotchIC function. AB - Notch proteins are transmembrane receptors that mediate intercell communication and direct individual cell fate decisions. The activated intracellular form of Notch, NotchIC, translocates to the nucleus, where it targets the DNA binding protein CBF1. CBF1 mediates transcriptional repression through the recruitment of an SMRT-histone deacetylase-containing corepressor complex. We have examined the mechanism whereby NotchIC overcomes CBF1-mediated transcriptional repression. We identified SKIP (Ski-interacting protein) as a CBF1 binding protein in a yeast two-hybrid screen. Both CBF1 and SKIP are highly conserved evolutionarily, and the SKIP-CBF1 interaction is also conserved in assays using the Caenorhabditis elegans and Drosophila melanogaster SKIP homologs. Protein-protein interaction assays demonstrated interaction between SKIP and the corepressor SMRT. More surprisingly, SKIP also interacted with NotchIC. The SMRT and NotchIC interactions were mutually exclusive. In competition binding experiments SMRT displaced NotchIC from CBF1 and from SKIP. Contact with SKIP is required for biological activity of NotchIC. A mutation in the fourth ankyrin repeat that abolished Notch signal transduction did not affect interaction with CBF1 but abolished interaction with SKIP. Further, NotchIC was unable to block muscle cell differentiation in myoblasts expressing antisense SKIP. The results suggest a model in which NotchIC activates responsive promoters by competing with the SMRT corepressor complex for contacts on both CBF1 and SKIP. PMID- 10713165 TI - A tissue-specific coactivator of steroid receptors, identified in a functional genetic screen. AB - Steroid receptors mediate responses to lipophilic hormones in a tissue- and ligand-specific manner. To identify nonreceptor proteins that confer specificity or regulate steroid signaling, we screened a human cDNA library in a steroid responsive yeast strain. One of the identified cDNAs, isolated in the screen as ligand effect modulator 6, showed no homology to yeast or Caenorhabditis elegans proteins but high similarity to the recently described mouse coactivator PGC-1 and was accordingly termed hPGC-1. The hPGC-1 DNA encodes a nuclear protein that is expressed in a tissue-specific manner and carries novel motifs for transcriptional regulators. The expression of hPGC-1 in mammalian cells enhanced potently the transcriptional response to several steroids in a receptor-specific manner. hPGC-1-mediated enhancement required the receptor hormone-binding domain and was dependent on agonist ligands. Functional analysis of hPGC-1 revealed two domains that interact with steroid receptors in a hormone-dependent manner, a potent transcriptional activation function, and a putative dimerization domain. Our findings suggest a regulatory function for hPGC-1 as a tissue-specific coactivator for a subset of nuclear receptors. PMID- 10713166 TI - c-Myc proteolysis by the ubiquitin-proteasome pathway: stabilization of c-Myc in Burkitt's lymphoma cells. AB - The c-Myc oncoprotein is a transcription factor which is a critical regulator of cellular proliferation. Deregulated expression of c-Myc is associated with many human cancers, including Burkitt's lymphoma. The c-Myc protein is normally degraded very rapidly with a half-life of 20 to 30 min. Here we demonstrate that proteolysis of c-Myc in vivo is mediated by the ubiquitin-proteasome pathway. Inhibition of proteasome activity blocks c-Myc degradation, and c-Myc is a substrate for ubiquitination in vivo. Furthermore, an increase in c-Myc stability occurs in mitotic cells and is associated with inhibited c-Myc ubiquitination. Deletion analysis was used to identify regions of the c-Myc protein which are required for rapid proteolysis. We found that a centrally located PEST sequence, amino acids 226 to 270, is necessary for rapid c-Myc degradation, but not for ubiquitination. Also, N-terminal sequences, located within the first 158 amino acids of c-Myc, are necessary for both efficient c-Myc ubiquitination and subsequent degradation. We found that c-Myc is significantly stabilized (two- to sixfold) in many Burkitt's lymphoma-derived cell lines, suggesting that aberrant c-Myc proteolysis may play a role in the pathogenesis of Burkitt's lymphoma. Finally, mutation of Thr-58, a major phosphorylation site in c-Myc and a mutational hot spot in Burkitt's lymphoma, increases c-Myc stability; however, mutation of c-Myc is not essential for stabilization in Burkitt's lymphoma cells. PMID- 10713168 TI - DNA interstrand cross-links induce futile repair synthesis in mammalian cell extracts. AB - DNA interstrand cross-links are induced by many carcinogens and anticancer drugs. It was previously shown that mammalian DNA excision repair nuclease makes dual incisions 5' to the cross-linked base of a psoralen cross-link, generating a gap of 22 to 28 nucleotides adjacent to the cross-link. We wished to find the fates of the gap and the cross-link in this complex structure under conditions conducive to repair synthesis, using cell extracts from wild-type and cross linker-sensitive mutant cell lines. We found that the extracts from both types of strains filled in the gap but were severely defective in ligating the resulting nick and incapable of removing the cross-link. The net result was a futile damage induced DNA synthesis which converted a gap into a nick without removing the damage. In addition, in this study, we showed that the structure-specific endonuclease, the XPF-ERCC1 heterodimer, acted as a 3'-to-5' exonuclease on cross linked DNA in the presence of RPA. Collectively, these observations shed some light on the cellular processing of DNA cross-links and reveal that cross-links induce a futile DNA synthesis cycle that may constitute a signal for specific cellular responses to cross-linked DNA. PMID- 10713167 TI - Nucleotide excision repair/TFIIH helicases RAD3 and SSL2 inhibit short-sequence recombination and Ty1 retrotransposition by similar mechanisms. AB - Eukaryotic genomes contain potentially unstable sequences whose rearrangement threatens genome structure and function. Here we show that certain mutant alleles of the nucleotide excision repair (NER)/TFIIH helicase genes RAD3 and SSL2 (RAD25) confer synthetic lethality and destabilize the Saccharomyces cerevisiae genome by increasing both short-sequence recombination and Ty1 retrotransposition. The rad3-G595R and ssl2-rtt mutations do not markedly alter Ty1 RNA or protein levels or target site specificity. However, these mutations cause an increase in the physical stability of broken DNA molecules and unincorporated Ty1 cDNA, which leads to higher levels of short-sequence recombination and Ty1 retrotransposition. Our results link components of the core NER/TFIIH complex with genome stability, homologous recombination, and host defense against Ty1 retrotransposition via a mechanism that involves DNA degradation. PMID- 10713169 TI - Genetic analysis of the YDR1-BUR6 repressor complex reveals an intricate balance among transcriptional regulatory proteins in yeast. AB - A transcriptional repressor complex encoded by two essential genes, YDR1 and BUR6, was isolated from Saccharomyces cerevisiae and shown to be the functional counterpart of the human repressor complex Dr1-DRAP1. To elucidate the mechanism of repression by this complex, altered forms of Ydr1 and Bur6 were studied in vitro and in vivo. Deletion of the C-terminal 41 amino acids of Ydr1 resulted in loss of repressor activity and a growth defect, suggesting that the C-terminal domain of Ydr1 functions as a potent transcriptional repressor. A screen for extragenic suppressors of a cold-sensitive ydr1 (ydr1(cs)) mutant led to the identification of recessive mutations in the SIN4 gene, which encodes a component of the SRB-MED complex. The sin4 alleles suppressed not only ydr1(cs) mutations but also bur6(cs) mutations. In contrast, deletion of the gal11 gene, whose product is also a member of the SRB-MED complex, failed to suppress ydr1(cs) and bur6(cs) mutations, indicating that suppression is not due to general defects in the SRB-MED complex. Moreover, one of the sin4 alleles, but not the sin4 deletion, was found to specifically suppress the inviability of a ydr1 deletion, demonstrating that the essential function of Ydr1 becomes dispensable in a sin4 mutant background. Biochemical analysis of the SRB-MED complex from the sin4 suppressor strain revealed a structurally distinct form of the SRB-MED complex that lacks a subset of mediator subunits. These results define a delicate balance between positive and negative regulators of transcription operating through the Ydr1-Bur6 repressor complex. PMID- 10713170 TI - Placental failure in mice lacking the mammalian homolog of glial cells missing, GCMa. AB - The GCM family of transcription factors consists of Drosophila melanogaster GCM, an important regulator of gliogenesis in the fly, and its two mammalian homologs, GCMa and GCMb. To clarify the function of these mammalian homologs, we deleted GCMa in mice. Genetic ablation of murine GCMa (mGCMa) is embryonic lethal, with mice dying between 9.5 and 10 days postcoitum. At the time of death, no abnormalities were apparent in the embryo proper. Nervous system development, in particular, was not impaired, as might have been expected in analogy to Drosophila GCM. Instead, placental failure was the cause of death. In agreement with the selective expression of mGCMa in labyrinthine trophoblasts, mutant placentas did not develop a functional labyrinth layer, which is necessary for nutrient and gas exchange between maternal and fetal blood. Only a few fetal blood vessels entered the placenta, and these failed to thrive and branch normally. Labyrinthine trophoblasts did not differentiate. All other layers of the placenta, including spongiotrophoblast and giant cell layer, formed normally. Our results indicate that mGCMa plays a critical role in trophoblast differentiation and the signal transduction processes required for normal vascularization of the placenta. PMID- 10713171 TI - H-ras but not K-ras traffics to the plasma membrane through the exocytic pathway. AB - Ras proteins must be localized to the inner surface of the plasma membrane to be biologically active. The motifs that effect Ras plasma membrane targeting consist of a C-terminal CAAX motif plus a second signal comprising palmitoylation of adjacent cysteine residues or the presence of a polybasic domain. In this study, we examined how Ras proteins access the cell surface after processing of the CAAX motif is completed in the endoplasmic reticulum (ER). We show that palmitoylated CAAX proteins, in addition to being localized at the plasma membrane, are found throughout the exocytic pathway and accumulate in the Golgi region when cells are incubated at 15 degrees C. In contrast, polybasic CAAX proteins are found only at the cell surface and not in the exocytic pathway. CAAX proteins which lack a second signal for plasma membrane targeting accumulate in the ER and Golgi. Brefeldin A (BFA) significantly inhibits the plasma membrane accumulation of newly synthesized, palmitoylated CAAX proteins without inhibiting their palmitoylation. BFA has no effect on the trafficking of polybasic CAAX proteins. We conclude that H-ras and K-ras traffic to the cell surface through different routes and that the polybasic domain is a sorting signal diverting K-Ras out of the classical exocytic pathway proximal to the Golgi. Farnesylated Ras proteins that lack a polybasic domain reach the Golgi but require palmitoylation in order to traffic further to the cell surface. These data also indicate that a Ras palmitoyltransferase is present in an early compartment of the exocytic pathway. PMID- 10713172 TI - Presence of a member of the mitochondrial carrier family in hydrogenosomes: conservation of membrane-targeting pathways between hydrogenosomes and mitochondria. AB - A number of microaerophilic eukaryotes lack mitochondria but possess another organelle involved in energy metabolism, the hydrogenosome. Limited phylogenetic analyses of nuclear genes support a common origin for these two organelles. We have identified a protein of the mitochondrial carrier family in the hydrogenosome of Trichomonas vaginalis and have shown that this protein, Hmp31, is phylogenetically related to the mitochondrial ADP-ATP carrier (AAC). We demonstrate that the hydrogenosomal AAC can be targeted to the inner membrane of mitochondria isolated from Saccharomyces cerevisiae through the Tim9-Tim10 import pathway used for the assembly of mitochondrial carrier proteins. Conversely, yeast mitochondrial AAC can be targeted into the membranes of hydrogenosomes. The hydrogenosomal AAC contains a cleavable, N-terminal presequence; however, this sequence is not necessary for targeting the protein to the organelle. These data indicate that the membrane-targeting signal(s) for hydrogenosomal AAC is internal, similar to that found for mitochondrial carrier proteins. Our findings indicate that the membrane carriers and membrane protein-targeting machinery of hydrogenosomes and mitochondria have a common evolutionary origin. Together, they provide strong evidence that a single endosymbiont evolved into a progenitor organelle in early eukaryotic cells that ultimately give rise to these two distinct organelles and support the hydrogen hypothesis for the origin of the eukaryotic cell. PMID- 10713173 TI - Expression and functional analysis of Uch-L3 during mouse development. AB - Mice homozygous for the s(1Acrg) deletion at the Ednrb locus arrest at embryonic day 8.5. To determine the molecular basis of this defect, we initiated positional cloning of the s(1Acrg) minimal region. The mouse Uch-L3 (ubiquitin C-terminal hydrolase L3) gene was mapped within the s(1Acrg) minimal region. Because Uch-L3 transcripts were present in embryonic structures relevant to the s(1Acrg) phenotype, we created a targeted mutation in Uch-L3 to address its role during development and its possible contribution to the s(1Acrg) phenotype. Mice homozygous for the mutation Uch-L3(Delta3-7) were viable, with no obvious developmental or histological abnormalities. Although high levels of Uch-L3 RNA were detected in testes and thymus, Uch-L3(Delta3-7) homozygotes were fertile, and no defect in intrathymic T-cell differentiation was detected. We conclude that the s(1Acrg) phenotype is either complex and multigenic or due to the loss of another gene within the region. We propose that Uch-L3 may be functionally redundant with its homologue Uch-L1. PMID- 10713174 TI - Defects in tRNA processing and nuclear export induce GCN4 translation independently of phosphorylation of the alpha subunit of eukaryotic translation initiation factor 2. AB - Induction of GCN4 translation in amino acid-starved cells involves the inhibition of initiator tRNA(Met) binding to eukaryotic translation initiation factor 2 (eIF2) in response to eIF2 phosphorylation by protein kinase GCN2. It was shown previously that GCN4 translation could be induced independently of GCN2 by overexpressing a mutant tRNA(AAC)(Val) (tRNA(Val*)) or the RNA component of RNase MRP encoded by NME1. Here we show that overexpression of the tRNA pseudouridine 55 synthase encoded by PUS4 also leads to translational derepression of GCN4 (Gcd(-) phenotype) independently of eIF2 phosphorylation. Surprisingly, the Gcd( ) phenotype of high-copy-number PUS4 (hcPUS4) did not require PUS4 enzymatic activity, and several lines of evidence indicate that PUS4 overexpression did not diminish functional initiator tRNA(Met) levels. The presence of hcPUS4 or hcNME1 led to the accumulation of certain tRNA precursors, and their Gcd(-) phenotypes were reversed by overexpressing the RNA component of RNase P (RPR1), responsible for 5'-end processing of all tRNAs. Consistently, overexpression of a mutant pre tRNA(Tyr) that cannot be processed by RNase P had a Gcd(-) phenotype. Interestingly, the Gcd(-) phenotype of hcPUS4 also was reversed by overexpressing LOS1, required for efficient nuclear export of tRNA, and los1Delta cells have a Gcd(-) phenotype. Overproduced PUS4 appears to impede 5'-end processing or export of certain tRNAs in the nucleus in a manner remedied by increased expression of RNase P or LOS1, respectively. The mutant tRNA(Val*) showed nuclear accumulation in otherwise wild-type cells, suggesting a defect in export to the cytoplasm. We propose that yeast contains a nuclear surveillance system that perceives defects in processing or export of tRNA and evokes a reduction in translation initiation at the step of initiator tRNA(Met) binding to the ribosome. PMID- 10713175 TI - Cooperative signals governing ARF-mdm2 interaction and nucleolar localization of the complex. AB - The ARF tumor suppressor protein stabilizes p53 by antagonizing its negative regulator, Mdm2 (Hdm2 in humans). Both mouse p19(ARF) and human p14(ARF) bind to the central region of Mdm2 (residues 210 to 304), a segment that does not overlap with its N-terminal p53-binding domain, nuclear import or export signals, or C terminal RING domain required for Mdm2 E3 ubiquitin ligase activity. The N terminal 37 amino acids of mouse p19(ARF) are necessary and sufficient for binding to Mdm2, localization of Mdm2 to nucleoli, and p53-dependent cell cycle arrest. Although a nucleolar localization signal (NrLS) maps within a different segment (residues 82 to 101) of the human p14(ARF) protein, binding to Mdm2 and nucleolar import of ARF-Mdm2 complexes are both required for cell cycle arrest induced by either the mouse or human ARF proteins. Because many codons of mouse ARF mRNA are not recognized by the most abundant bacterial tRNAs, we synthesized ARF minigenes containing preferred bacterial codons. Using bacterially produced ARF polypeptides and chemically synthesized peptides conjugated to Sepharose, residues 1 to 14 and 26 to 37 of mouse p19(ARF) were found to interact independently and cooperatively with Mdm2, while residues 15 to 25 were dispensable for binding. Paradoxically, residues 26 to 37 of mouse p19(ARF) are also essential for ARF nucleolar localization in the absence of Mdm2. However, the mobilization of the p19(ARF)-Mdm2 complex into nucleoli also requires a cryptic NrLS within the Mdm2 C-terminal RING domain. The Mdm2 NrLS is unmasked upon ARF binding, and its deletion prevents import of the ARF-Mdm2 complex into nucleoli. Collectively, the results suggest that ARF binding to Mdm2 induces a conformational change that facilitates nucleolar import of the ARF-Mdm2 complex and p53-dependent cell cycle arrest. Hence, the ARF-Mdm2 interaction can be viewed as bidirectional, with each protein being capable of regulating the subnuclear localization of the other. PMID- 10713176 TI - v-Jun overrides the mitogen dependence of S-phase entry by deregulating retinoblastoma protein phosphorylation and E2F-pocket protein interactions as a consequence of enhanced cyclin E-cdk2 catalytic activity. AB - v-Jun accelerates G(1) progression and shares the capacity of the Myc, E2F, and E1A oncoproteins to sustain S-phase entry in the absence of mitogens; however, how it does so is unknown. To gain insight into the mechanism, we investigated how v-Jun affects mitogen-dependent processes which control the G(1)/S transition. We show that v-Jun enables cells to express cyclin A and cyclin A cdk2 kinase activity in the absence of growth factors and that deregulation of cdk2 is required for S-phase entry. Cyclin A expression is repressed in quiescent cells by E2F acting in conjunction with its pocket protein partners Rb, p107, and p130; however, v-Jun overrides this control, causing phosphorylated Rb and proliferation-specific E2F-p107 complexes to persist after mitogen withdrawal. Dephosphorylation of Rb and destruction of cyclin A nevertheless occur normally at mitosis, indicating that v-Jun enables cells to rephosphorylate Rb and reaccumulate cyclin A without exogenous mitogenic stimulation each time the mitotic "clock" is reset. D-cyclin-cdk activity is required for Rb phosphorylation in v-Jun-transformed cells, since ectopic expression of the cdk4- and cdk6-specific inhibitor p16(INK4A) inhibits both DNA synthesis and cell proliferation. Despite this, v-Jun does not stimulate D-cyclin-cdk activity but does induce a marked deregulation of cyclin E-cdk2. In particular, hormonal activation of a conditional v-Jun-estrogen receptor fusion protein in quiescent, growth factor-deprived cells stimulates cyclin E-cdk2 activity and triggers Rb phosphorylation and DNA synthesis. Thus, v-Jun overrides the mitogen dependence of S-phase entry by deregulating Rb phosphorylation, E2F-pocket protein interactions, and ultimately cyclin A-cdk2 activity. This is the first report, however, that cyclin E-cdk2, rather than D-cyclin-cdk, is likely to be the critical Rb kinase target of v-Jun. PMID- 10713177 TI - Defining the regulatory factors required for epidermal gene expression. AB - Keratins K5 and K14 are the hallmarks of mitotically active keratinocytes of stratified epithelia. They are transcribed at a high level and in a tissue specific manner, enabling us to use the K14 gene to elucidate the regulatory mechanism underlying epidermis-specific transcription. We have identified four DNase I-hypersensitive sites (HSs) present in the 5' regulatory sequences of the K14 gene under specific conditions where the gene is actively expressed. Two of these sites (HSsII and -III) are conserved in position and sequence within the human and mouse K14 genes. Using an in vivo transgenic approach and an in vitro keratinocyte culture approach, we have discovered that most of K14's transcriptional activity is restricted to a novel 700-bp regulatory domain encompassing these HSs. This enhancer is sufficient to confer epidermis-specific activity to a heterologous promoter in transfection assays in culture and in transgenic mice in vivo. A 125-bp DNA fragment encompassing HSsII harbors the majority of the transactivation activity in vitro, and electrophoretic mobility shift and mutational assays reveal a role for AP-1, ets, and AP-2 family members in orchestrating the keratinocyte-preferred expression of HSsII. The HSsII element also confers epidermal expressivity to a heterologous promoter in transgenic mice, although it is not sufficient on its own to fully restrict activity to keratinocytes. Within the HSsII element, the ets and AP-2 sites appear to be most critical in collaborating to regulate epidermal specificity in vivo. PMID- 10713178 TI - Mixed-lineage kinase 3 delivers CD3/CD28-derived signals into the IkappaB kinase complex. AB - The phosphorylation of IkappaB by the multiprotein IkappaB kinase complex (IKC) precedes the activation of transcription factor NF-kappaB, a key regulator of the inflammatory response. Here we identified the mixed-lineage group kinase 3 (MLK3) as an activator of NF-kappaB. Expression of the wild-type form of this mitogen activated protein kinase kinase kinase (MAPKKK) induced nuclear immigration, DNA binding, and transcriptional activity of NF-kappaB. MLK3 directly phosphorylated and thus activated IkappaB kinase alpha (IKKalpha) and IKKbeta, revealing its function as an IkappaB kinase kinase (IKKK). MLK3 cooperated with the other two IKKKs, MEKK1 and NF-kappaB-inducing kinase, in the induction of IKK activity. MLK3 bound to components of the IKC in vivo. This protein-protein interaction was dependent on the central leucine zipper region of MLK3. A kinase-deficient version of MLK3 strongly impaired NF-kappaB-dependent transcription induced by T cell costimulation but not in response to tumor necrosis factor alpha or interleukin-1. Accordingly, endogenous MLK3 was phosphorylated and activated by T cell costimulation but not by treatment of cells with tumor necrosis factor alpha or interleukin-1. A dominant negative version of MLK3 inhibited NF-kappaB- and CD28RE/AP-dependent transcription elicited by the Rho family GTPases Rac and Cdc42, thereby providing a novel link between these GTPases and the IKC. PMID- 10713179 TI - Promoter-proximal pausing on the hsp70 promoter in Drosophila melanogaster depends on the upstream regulator. AB - RNA polymerase II pauses in the promoter-proximal region of many genes during transcription. In the case of the hsp70 promoter from Drosophila melanogaster, this pause is long-lived and occurs even when the gene is not induced. Paused polymerase escapes during heat shock when the transcriptional activator heat shock factor associates with the promoter. However, pausing is still evident, especially when induction is at an intermediate level. Yeast Gal4 protein (Gal4p) will induce transcription of the hsp70 promoter in Drosophila when binding sites for Gal4p are positioned upstream from the hsp70 TATA element. To further our understanding of promoter-proximal pausing, we have analyzed the effect of Gal4p on promoter-proximal pausing in salivary glands of Drosophila larvae. Using permanganate genomic footprinting, we observed that various levels of Gal4p induction resulted in an even distribution of RNA polymerase throughout the first 76 nucleotides of the transcribed region. In contrast, promoter-proximal pausing still occurs on endogenous and transgenic hsp70 promoters in salivary glands when these promoters are induced by heat shock. We also determined that mutations introduced into the region where the polymerase pauses do not inhibit pausing in a cell-free system. Taken together, these results indicate that promoter-proximal pausing is dictated by the regulatory proteins interacting upstream from the core promoter region. PMID- 10713180 TI - Cooperation of p27(Kip1) and p18(INK4c) in progestin-mediated cell cycle arrest in T-47D breast cancer cells. AB - The steroid hormone progesterone regulates proliferation and differentiation in the mammary gland and uterus by cell cycle phase-specific actions. The long-term effect of progestins on T-47D breast cancer cells is inhibition of cellular proliferation. This is accompanied by decreased G(1) cyclin-dependent kinase (CDK) activities, redistribution of the CDK inhibitor p27(Kip1) among these CDK complexes, and alterations in the elution profile of cyclin E-Cdk2 upon gel filtration chromatography, such that high-molecular-weight complexes predominate. This study aimed to determine the relative contribution of CDK inhibitors to these events. Following progestin treatment, the majority of cyclin E- and D-CDK complexes were bound to p27(Kip1) and few were bound to p21(Cip1). In vitro, recombinant His(6)-p27 could quantitatively reproduce the effects on cyclin E Cdk2 kinase activity and the shift in molecular weight observed following progestin treatment. In contrast, cyclin D-Cdk4 was not inhibited by His(6)-p27 in vitro or p27(Kip1) in vivo. However, an increase in the expression of the Cdk4/6 inhibitor p18(INK4c) and its extensive association with Cdk4 and Cdk6 were apparent following progestin treatment. Recombinant p18(INK4c) led to the reassortment of cyclin-CDK-CDK inhibitor complexes in vitro, with consequent decrease in cyclin E-Cdk2 activity. These results suggest a concerted model of progestin action whereby p27(Kip1) and p18(INK4c) cooperate to inhibit cyclin E Cdk2 and Cdk4. Since similar models have been developed for growth inhibition by transforming growth factor beta and during adipogenesis, interaction between the Cip/Kip and INK4 families of inhibitors may be a common theme in physiological growth arrest and differentiation. PMID- 10713181 TI - Transcriptional repression by blimp-1 (PRDI-BF1) involves recruitment of histone deacetylase. AB - B-lymphocyte-induced maturation protein (Blimp-1) is a transcriptional repressor that is considered to be a master regulator of terminal B-cell development because it is sufficient to trigger differentiation in the BCL(1)-cell model. Transcription of the c-myc gene is repressed by Blimp-1 during B-cell differentiation. In this study, we have explored the mechanism by which Blimp-1 represses transcription by using Gal4-fusion protein assays and assays in which Blimp-1 represses the natural c-myc promoter. The results show that Blimp-1 represses the c-myc promoter by an active mechanism that is independent of the adjacently bound activator YY1. Blimp-1 contains two regions that independently associate with histone deacetylase (HDAC) and endogenous Blimp-1 in nuclear extracts binds in vitro to the c-myc Blimp-1 site in a complex containing HDAC. The functional importance of recruiting HDAC for Blimp-1-dependent repression of c-myc transcription is supported by two experiments. First, the HDAC inhibitor tricostatin A inhibits Blimp-1-dependent repression in cotransfection assays. Second, a chromatin immunoprecipitation assay shows that expression of Blimp-1 causes deacetylation of histone H3 associated with the c-myc promoter, and this deacetylation depends on the Blimp-1 binding site in the c-myc promoter. PMID- 10713182 TI - The orphan nuclear receptor Ear-2 is a negative coregulator for thyroid hormone nuclear receptor function. AB - Thyroid hormone (T3) nuclear receptors (TR) are ligand-dependent transcription factors which regulate growth, differentiation, and development. One emerging hypothesis suggests that TR mediate these diverse effects via a large network of coregulators. Recently, we found that TR-mediated transcriptional responses varied in six cell lines derived from different tissues. We therefore used human TR subtype beta1 (TRbeta1) as bait to search for coregulators in human colon carcinoma RKO cells with a yeast two-hybrid system. RKO cells exhibited T3 dependent and -independent transcriptional activation. One of the three positive clones was identified as Ear-2, which is a distant member of the chick ovalbumin upstream promoter-transcription factors of the orphan nuclear receptor family. The physical interaction between Ear-2 and TRbeta1 was further confirmed by specific binding of Ear-2 to glutathione S-transferase-TRbeta1. In addition, Ear 2 was found to associate with TRbeta1 in cells. As a result of this physical interaction, binding of TRbeta1 to the T3 response elements was inhibited. Using reporter systems, we found that both the basal activation and the T3-dependent activation mediated by TRbeta1 were repressed by Ear-2 in CV1 cells. In RKO cells, however, the T3-independent transcriptional activity was more sensitive to the repression effect of Ear-2 than the T3-dependent transcriptional activity. The repression effect of Ear-2 was reversed by steroid hormone receptor coactivator 1. These results suggest that TR-mediated responses reflect a balance of corepressors and coactivators in cells. These findings further strengthen the hypothesis that the diverse activities of TR are achieved via a large network of coregulators that includes Ear-2. PMID- 10713183 TI - p21-activated kinase 1 plays a critical role in cellular activation by Nef. AB - The activation of Nef-associated kinase (NAK) by Nef from human and simian immunodeficiency viruses is critical for efficient viral replication and pathogenesis. This induction occurs via the guanine nucleotide exchange factor Vav and the small GTPases Rac1 and Cdc42. In this study, we identified NAK as p21 activated kinase 1 (PAK1). PAK1 bound to Nef in vitro and in vivo. Moreover, the induction of cytoskeletal rearrangements such as the formation of trichopodia, the activation of Jun N-terminal kinase, and the increase of viral production were blocked by an inhibitory peptide that targets the kinase activity of PAK1 (PAK1 83-149). These results identify NAK as PAK1 and emphasize the central role its kinase activity plays in cytoskeletal rearrangements and cellular signaling by Nef. PMID- 10713185 TI - Lumbar herniated disc presenting with cauda equina syndrome. Long-term follow-up of four cases. AB - BACKGROUND: Cauda equina syndrome is a relatively rare presenting symptom of lumbar herniated discs. Early operative decompression is advocated, but it may not always restore the bladder function. In such cases, knowing the long-term outcome of bladder or sphincter disturbances is quite useful in planning the management of these patients. METHODS: Hospital records of patients operated on for lumbar herniated discs were reviewed. Charts and imaging studies of those patients who presented with classic cauda equina syndrome were analyzed. RESULTS: There were 4 patients (2.8%) of 144 consecutive surgical series of lumbar disc herniation, whose presenting symptom was classic cauda equina syndrome. Postoperatively, the patients had been followed at the clinic for a mean period of 6.4 years. Certain characteristic findings were noted on patients' neurological and radiological examinations. Although the recovery process of bladder function was very slow, taking months to years, all four patients achieved almost normal voiding with no major impairment of daily activities. CONCLUSIONS: Even if short-term recovery of bladder function is poor after lumbar disc surgery for cauda equina syndrome, the long-term outcome is not necessarily so. We should follow up on these patients with such measures as intermittent self catheterization and drug therapy, expecting slow but steady recovery of bladder and sphincter function. PMID- 10713186 TI - Os odontoideum: etiology, diagnosis, and management. AB - BACKGROUND: There have been few reports of os odontoideum since the initial description. METHODS: Forty-four patients with os odontoideum treated during the period 1980 through 1996 were reviewed. There were 33 males and 11 females. Their ages ranged from 7 to 56 years, with an average of 24.6 years. Five patients with no symptoms were treated conservatively. Thirty-nine patients underwent operative treatment including nine posterior atlantoaxial fusions and 33 occipitocervical fusions. RESULTS: The patients were followed up for one to 16 years, with an average of 6.5 years. Five patients treated conservatively have remained stable. All 39 treated patients achieved solid arthrodesis. The results were satisfactory. CONCLUSIONS: We conclude that fusion is indicated if atlantoaxial instability or clinical symptoms are significant, and that occipitocervical fusion should be considered in the operative management of os odontoideum if atlantoaxial arthrodesis is impossible. PMID- 10713187 TI - Cisternal irrigation therapy with urokinase and ascorbic acid for prevention of vasospasm after aneurysmal subarachnoid hemorrhage. Outcome in 217 patients. AB - BACKGROUND: Cisternal irrigation therapy with urokinase and ascorbic acid was introduced to prevent symptomatic vasospasm after aneurysmal subarachnoid hemorrhage (SAH). To dissolve and wash out the subarachnoid clot, cisternal irrigation with urokinase is used. Ascorbic acid is added to degenerate oxy hemoglobin, one of the strongest spasmogenic substances, into verdohemelike products, which are nonspasmogenic. The efficacy and safety of this therapy were evaluated. METHODS: This therapy was performed consecutively in 217 patients. The degree of SAH of the patients was classified as Fisher CT Group 3, and the highest CT number (Hounsfield number) exceeded 60 in the SAH, which suggested a significant risk for symptomatic vasospasm. All patients underwent surgery within 72 hours from the onset of SAH. After clipping the aneurysm, irrigation tubes were placed in the Sylvian fissure (inlet) unilaterally or bilaterally and in the prepontine or chiasmal cistern (outlet). Lactated Ringer's solution with urokinase (120 IU/mL) and ascorbic acid (4 mg/mL) was infused at a rate of 30 mL/hour/side for approximately 10 days. RESULTS: Of the 217 patients studied, symptomatic vasospasm was observed in 6 cases (2.8%), and two of these six cases (0.9%) demonstrated sequelae. The average total blood volume calculated from the drainage fluid was approximately 114 mL. Analysis of the absorption spectrum of the drainage fluid revealed disappearance of the oxy-hemoglobin-specific 576-nm peak. Complications occurred in eight patients during irrigation therapy; two patients experienced seizures, two patients developed meningitis, and four patients had an intracranial hemorrhage. However, all of these patients recovered without neurological deficits. CONCLUSIONS: These results suggest that cisternal irrigation therapy with urokinase and ascorbic acid is effective in preventing symptomatic vasospasm after aneurysmal SAH. PMID- 10713188 TI - Chronic hydrocephalus in elderly patients following subarachnoid hemorrhage. AB - BACKGROUND: With the aging of the population, surgery for ruptured intracranial aneurysms is increasing among the elderly. We sought to clarify the characteristics of chronic hydrocephalus following aneurysmal subarachnoid hemorrhage (SAH) in elderly patients. METHODS: Of the 576 surgically treated patients, 289 were aged 59 years or younger, 169 were 60 to 69, and 118 were 70 years or older. The relationship between chronic hydrocephalus and the causative factors was analyzed for each age group. RESULTS: Of the 576 patients, chronic hydrocephalus was observed in 215 (37%), with the incidence increasing significantly with age (p < 0.001) and being the highest in the oldest age group. In elderly patients, the incidence of chronic hydrocephalus was relatively high, even after mild SAH. The incidence of chronic hydrocephalus was high regardless of age in patients with severe SAH, such as in those with H&H grades III-IV, SAH grades III-IV, acute hydrocephalus, symptomatic vasospasm, and intraventricular hemorrhage, and in those with vertebro-basilar artery aneurysms. CONCLUSION: In the elderly, the incidence of chronic hydrocephalus following SAH was significantly higher than in younger patients, even after mild SAH. In elderly patients, careful observation and individualized treatment are necessary even if SAH is mild. PMID- 10713189 TI - Risk factors for the development of post-traumatic cerebral vasospasm. AB - BACKGROUND: Post-traumatic vasospasm is a well-recognized sequela of head injury. The risk factors associated with post-traumatic vasospasm have not been well defined. We studied 119 consecutive patients with head injury to determine the risk factors for post-traumatic vasospasm. METHODS: Twenty-nine (27.1%) patients were excluded from the study because of poor insonation (n = 12) or a hospital stay of less than 72 hours (n = 17). Seventy (77.8%) of 90 patients suffered severe head injury. Sixteen (17.8%) patients sustained moderate head injury and four (4.4%) patients sustained mild head injury. All patients were monitored with transcranial Doppler (TCD) ultrasonography daily. RESULTS: Post-traumatic vasospasm was detected in 32 (35.6%) of 90 patients. Among these patients, 29 (90.6%) had severe head injury, and three (9.4%) had moderate head injury. None of the patients with mild head injury suffered post-traumatic vasospasm. In most cases, the onset of post-traumatic vasospasm began on the fifth day and lasted 1 to 9 days. In 8 (25%) patients, post-traumatic vasospasm began within the first three days of the head injury. Among 32 patients with post-traumatic vasospasm, 10 (31.2%) patients had mild vasospasm, 20 (65.5%) had moderate vasospasm, and 2 (6.3%) had severe post-traumatic vasospasm. Clinical deterioration was documented in two (2.5%) patients. CONCLUSIONS: Development of post-traumatic vasospasm correlated only with severe subarachnoid hemorrhage on initial computed tomographic scan. There was an increased incidence of post-traumatic vasospasm in patients with epidural hematomas, subdural hematomas, and intracerebral hemorrhages. The Glasgow Coma Scale (GCS) score on admission was inversely related to the development of post-traumatic vasospasm. In most cases, the period of vasospasm was short and clinical deterioration was rare. Probably, two varieties of post-traumatic vasospasm exist, one that lasts a shorter time and does not correlate with the presence of SAH, and a second that correlates with the presence of SAH, lasts longer, and resembles aneurysmal vasospasm. PMID- 10713190 TI - Endothelin-1 concentration increases in the cerebrospinal fluid in cerebral vasospasm caused by subarachnoid hemorrhage. AB - BACKGROUND: Endothelin-1 (ET-1) was originally identified as a potent vasoconstrictor peptide. Numerous reports have suggested its roles in various disorders. Although there is a great deal of evidence establishing the relationship between ET-1 and cerebral vasospasm in animals, this relationship still remains to be clarified in humans. METHODS: The concentration of ET-1 in cerebrospinal fluid (CSF) was measured by radioimmunoassay in 23 subarachnoid hemorrhage patients. CSF samples were collected every 10 days after surgery from the cisternal drainage tube. RESULTS: Initial concentrations of ET-1 in the CSF collected the first day after operation were all increased compared with the control CSF. In seven of the eight vasospasm patients, the concentrations of ET-1 had increased before the observation of vasospasm and then decreased before the disappearance of the vasospasm. In 13 out of the 15 patients without vasospasm, the concentrations of ET-1 in CSF decreased with time. CONCLUSION: We confirmed that the concentration of ET-1 in CSF increased before the onset of cerebral vasospasm caused by subarachnoid hemorrhage. The ET-1 concentration in the CSF could be a useful marker to detect cerebral vasospasm after subarachnoid hemorrhage. PMID- 10713191 TI - Traumatic aneurysms and arteriovenous fistulas of the extracranial vessels in war injuries. AB - BACKGROUND: Extracranial vessel injuries are potentially devastating complications of penetrating head and neck wounds associated with war conflicts. These vasculopathies may be occlusive or they may lead to formation of traumatic aneurysms (TA) and arteriovenous fistulae (AVF). Even though these penetrating injuries are usually clinically silent and often appear only as small superficial wounds, they may lead to catastrophic hemorrhage or vascular insult. In this study, we attempted to elucidate signs, symptoms and circumstances present in these victims who are at risk of harboring an occult vasculopathy, excluding the occlusive ones and concentrating primarily on TAs and AVFs. MATERIALS AND METHODS: In a prospective study conducted during 8 years of war between Iran and Iraq, we encountered 13 cases of traumatic vasculopathies of the extracranial carotid and vertebral arteries. The type and number of injuries were: carotid jugular fistula (CJF) 3, carotid trunk or branch aneurysm (CA) 2, superficial temporal artery aneurysm (STA) 3, vertebral artery aneurysm (VA) 2, vertebral arteriovenous fistula (VAVF) 1, ophthalmic artery aneurysm (Oph. An.) 1, and lingual artery aneurysm (Lin. An.) 1. Angiography was performed between the 5th and 30th day after the injury and surgical intervention was performed in all cases. RESULTS: The Glasgow outcome scale (GOS) score was 13-15 in all victims at the time of discharge from the base hospital without any additional neurological deficit. The follow-up period varied from 5 to 8 years in all cases in whom no further morbidity or mortality occurred. Single photon emission computed tomography was the noninvasive tool used for measurement of cerebral blood flow in the cases in which a major vessel ligation was performed; no remarkable change in cerebral blood flow was noted. CONCLUSION: Early recognition of stigmas suggesting possible formation of extracranial traumatic vasculopathies such as TAs or AVFs in the difficult situation of war frontier hospitals should be highlighted for attending physicians or younger neurosurgeons. Performing angiography promptly in suspected cases can pick up such traumatic vascular lesions earlier. Using simpler surgical techniques in situations in which more sophisticated endovascular equipment is unavailable can be life-saving for these usually young victims. PMID- 10713192 TI - The rubber dam interposition technique during intracranial aneurysm clipping under deep hypothermic circulatory arrest. AB - OBJECTIVE: We describe a technique for isolating perforating arteries by use of a rubber sheet during the clipping of intracranial aneurysms under deep hypothermic circulatory arrest. METHODS: After meticulous dissection of the perforating arteries from the aneurysm, a rubber sheet, cut from a surgical glove to the proper shape, is inserted between the aneurysm and the arteries to prevent arterial reattachment to the aneurysmal neck. RESULTS: Even when perforators are invisible at the time of neck clipping because of a large aneurysmal body, the blade of the clip slides along the rubber sheet without injuring the vessels. CONCLUSION: With this method, an aneurysm, especially a large or giant one, can be clipped easily and safely even when the patient is in deep hypothermic circulatory arrest. PMID- 10713193 TI - Techniques of coiling cerebral aneurysms. AB - BACKGROUND: More than 200 aneurysms have been coiled at the UIC Medical Center within the last 5 years. We describe in detail the technical factors that increase the chance of complete occlusion of a cerebral aneurysm with coils. Aneurysms selected for coiling have good geometry or are in a location that is difficult to reach surgically. Patients with medical conditions that preclude surgical treatment may also undergo coiling. METHODS: Patients with aneurysms, either ruptured or unruptured, are treated under general anesthesia, fully anticoagulated and deeply paralyzed. Coiling is done under simultaneous biplane roadmapping. After the first coil has created a mesh, the aneurysm is densely packed with soft coils of decreasing diameter, until no more coils can be deployed into the aneurysm. RESULTS: The morbidity and mortality rates associated with the coiling procedure have continuously decreased over the last 5 years. The morphological outcomes have improved, due to extensive use of the remodeling technique and to advancements in materials, such as refinements in the coils themselves or the availability of over-the-wire balloon catheters in different sizes and hydrophilic wires with complex tip configurations. Twenty-one percent of the aneurysms were considered to be incompletely occluded immediately after coiling. Of this group, one-third of the aneurysms were found to be completely occluded on follow-up angiograms by 6 months; these have remained occluded. One third were more than 95% occluded after the coiling procedure; in these patients, the dome was completely occluded, but there was a small neck remnant, which has remained stable in all patients on control angiograms obtained at 6 months and 1, 2, and 4 years; none have rebled. These patients are followed medically. The remaining one-third of the aneurysms in this subgroup were less than 95% occluded, although the dome was completely thrombosed. None of them have rebled, but the neck remnant in most has regrown over a period ranging from 6 months to 2 years. These patients have undergone a second treatment-either surgical clipping, permanent occlusion of the parent vessel, or repeat coiling using the remodeling technique. The overall rebleeding rate of incompletely occluded aneurysms is extremely low (less than 1%). CONCLUSION: The low morbidity and mortality rates and the good morphological outcome obtained in most cases make coiling a reasonable alternative to surgical clipping in properly selected cases. PMID- 10713194 TI - Brain tumors in Mexico: characteristics and prognosis of glioblastoma. AB - BACKGROUND: Frequency and clinical characteristics of brain tumors have been studied in several populations from different genetic backgrounds; their peculiarities in the Mexican mestizo population shed light on the descriptive and comparative epidemiologic analysis of the genetic participation in brain tumors. METHODS: To analyze the frequency of intracranial neoplasms at the National Institute of Neurology and Neurosurgery of Mexico between 1987 and 1994, demographic, clinical, surgical, and neuropathological records were reviewed and compared with other reports. Glioblastoma cases were followed to investigate survival and prognostic factors. RESULTS: In a seven-year period 1,776 patients with brain tumors were treated; 419 (24%) had pituitary adenoma; 586 (33%) had glioma. Of the latter, 165 had glioblastoma multiforme, representing 28% of all gliomas and 9% of all neoplasms. Mean survival for glioblastoma was 16 months and the longest mean survival was obtained in patients with radical neurosurgical resection plus radiotherapy and chemotherapy. Cumulative analysis showed that 41% of patients survived less than one year, 39% from 1 to 2 years, 12% from 2 to 3 years and 8% more than three years. Factors that showed prognostic significance were age, therapeutic approach, tumor size, and pre- and postoperative clinical status (p < 0.05). CONCLUSIONS: This study comprises the largest series on the frequency of brain tumors in a Latin American population. When compared with other studies, the proportion of glioma and glioblastoma among brain neoplasms was low whereas pituitary adenoma was high. Mean survival for glioblastoma was similar to other reports; in these patients, the overall therapeutic response is still far from satisfactory. PMID- 10713195 TI - Characteristic features of malignant lymphoma with central nervous system involvement. AB - BACKGROUND: Cerebral lymphoma is becoming increasingly common. METHODS: We reviewed the records of all our patients with non-Hodgkin's lymphoma (NHL) seen from April 1987 to August 1996 in our institute. Our analysis of these patients with lymphomatous central nervous system (CNS) involvement documents the clinical features, histology, and prognostic factors in CNS lymphoma. RESULTS: A total of 351 cases of NHL were treated in our institute. CNS lymphoma was found in 58 of 351 (16.5%) patients in our series. Forty-nine of 58 (84.5%) patients with CNS involvement also had systemic disease. Primary CNS lymphomas were detected in nine patients. Leptomeningeal infiltration was seen in 31 of 58 patients, whereas intracerebral infiltration was detected in 28 patients. Initial symptoms of CNS involvement included severe headache, muscle weakness, and other neurological signs. Malignant cells were detected in 32 of 132 studies in the cerebrospinal fluid examination. In the nine patients with primary CNS lymphoma, the median survival time was 16.5 months (range, 4-28 months). The overall median survival of the 58 CNS lymphoma patients was only 13.4 months (range, 1 to 32 months). CONCLUSIONS: Because prophylactic treatment was only successful in systemically well-controlled patients, control of systemic lymphoma seems to be of great importance. PMID- 10713196 TI - Endoscopic-guided direct endonasal approach for pituitary surgery. AB - BACKGROUND: Submucosal dissection of the nasal septum is often performed as part of the transseptal approach to the sella. To evaluate whether this submucosal dissection is a necessary component of this operation, we compared the morbidity of a direct transmucosal endonasal approach to that of the transseptal approach in patients undergoing pituitary surgery. METHODS: Forty-one consecutive patients undergoing pituitary surgery from January 1996 to March 1999 were included in this study. The first 21 patients underwent the standard transseptal operation through either a sublabial or columellar incision. The latter 20 patients were operated on through an endoscopically guided, direct endonasal exposure, without any submucosal dissection of the nasal septum. The operative morbidity, the duration of surgery, and the length of hospitalization for each group were compared. RESULTS: The sphenoid sinus exposure obtained through the endonasal route was comparable with the transseptal approach and was adequate for resection of most pituitary tumors. Although the morbidity of the two approaches was similar, patients undergoing the endonasal operation had less postoperative facial pain. Furthermore, the endonasal approach significantly decreased the length of the operation (116 minutes vs. 161 minutes, p = 0.002) and the duration of hospitalization (3.6 vs. 5.1 days, p = 0.003) as compared with the transseptal route. CONCLUSIONS: Morbidity of the endonasal approach to the sphenoid sinus is comparable to that of a conventional transseptal approach. By eliminating the submucosal dissection, the endonasal approach reduces postoperative facial discomfort and decreases length of surgery and hospitalization. PMID- 10713197 TI - Isolated sphenoid sinus abscess: clinical and radiological failure in preoperative diagnosis. Case report and review of the literature. AB - BACKGROUND: Isolated sphenoid sinusitis and abscess formation is a rare entity, which can lead to misdiagnosed or improperly treated patients and an unfavorable outcome. Invasion of the skull base and cavernous sinus usually causes cranial nerve palsies, suggesting a neoplasm at the initial presentation. CASE DESCRIPTION: A case of isolated abscess in the sphenoid sinus is reported. The complete destruction of the clivus and its unexceptional radiological data, in addition to the absence of clinical and laboratory evidence of infection, led us to misdiagnose a possible clival chordoma during preoperative evaluation. The patient underwent an endonasal-transsphenoidal procedure for diagnosis and surgical removal. Surgical drainage and prolonged antimicrobial treatment resulted in complete clinical recovery. CONCLUSION: Its close proximity to vital structures and slender bony structures may allow the infection to disseminate, with serious neurological complications. On the other hand, the variable clinical presentations and radiological data usually cause delayed or missed diagnosis in these cases. This emphasizes the importance of documentation of this unusual entity and its radiological manifestations. PMID- 10713198 TI - Positioning of vagal nerve stimulators: technical note. AB - BACKGROUND: Vagal nerve stimulation has become an important treatment for patients with intractable seizure disorders. Many of these patients will require magnetic resonance imaging (MRIs) of the brain after the stimulator has been implanted to monitor underlying neurologic conditions. Functional MRI (fMRI) is also being used in the evaluation of epilepsy. With the current recommended implant techniques the magnetic field of the MRI will deactivate the pulse generator while the patient is in the supine position for the scan. A simple change in positioning of the pulse generator will help to avoid deactivating the device during an MRI. This will avoid exposing the patient to lengthy time periods with a deactivated stimulator and also allow for the performance of fMRIs and any other MRI scans needed to monitor underlying neurologic conditions. METHODS: A working model of the NeuroCybernetic Prosthesis (NCP) pulse generator was assessed with an oscilloscope and LED light connected to it that related activation of the generator while in the MRI. This simulation was performed with the device alone, in multiple positions. Then patients with implanted devices who could personally confirm the activation of their stimulators were also studied. RESULTS: A pulse generator placed with the electrode inputs parallel to the long axis of the body was not deactivated by the magnetic field of the MRI when the patient was in the supine position. CONCLUSION: Changing the implant position of a vagal nerve stimulator pulse generator will help to prevent deactivation of the device while in the MRI, allowing for the performance of fMRIs while not exposing the patient to lengthy time periods with a deactivated vagal nerve stimulator. PMID- 10713199 TI - Does autonomic neuropathy influence spinal cord stimulation therapy success in diabetic patients with critical lower limb ischemia? AB - BACKGROUND: Spinal cord stimulation (SCS) improves microcirculatory blood flow and relieves diabetic neuropathic and ischemic pain, reducing the amputation rate in patients with peripheral arterial occlusive disease (PAOD). The purpose of this study was to evaluate whether the presence of autonomic neuropathy in diabetic patients with PAOD influences the success of SCS therapy. METHODS: Sixty consecutive diabetic patients (15 with early and 13 with definite and/or combined autonomic neuropathy) with an ankle/brachial systolic pressure index (ABI) less than 0.20 +/- 0.08, underwent spinal cord stimulation after failed conservative or surgical treatment. The neuropathic status of the patients was evaluated before implantation and pedal TcpO2 measurements on the dorsum of the foot were performed. RESULTS: Limb salvage and pain relief >75%, evaluated with the visual analogue scale, were achieved in 35 patients, whereas in 12 a partial success with pain relief >50% and limb salvage for at least 6 months were obtained. In 13 patients the method failed and the ischemic limbs were amputated. Among the 28 diabetic patients with autonomic neuropathy the treatment failed or resulted in only partial success in 25, whereas in all 32 patients without neuropathy limb salvage and pain relief >75% were achieved (p < 0.0001). Partial success in 10 patients with early neuropathy and in two with definite was achieved (p = 0.008), whereas in 11 patients with definite neuropathy and in two with early the method failed (p < 0.001). The stage of the neuropathy was inversely related to the success of SCS therapy, independent of the stage of the disease. The method's success was related to the presence of adequate paraesthesias and warm feeling in the painful area with size reduction of the trophic lesions. CONCLUSIONS: Diabetic patients with peripheral arterial occlusive disease presenting with intractable pain may be successfully treated with spinal cord stimulation unless they have associated severe autonomic neuropathy. PMID- 10713200 TI - Good recovery after surgery in an extreme case of Guyon's canal syndrome. PMID- 10713201 TI - Evaded bioethics and the vocation of medicine--the future at stake. PMID- 10713202 TI - Polymerization of lignin fragments contained in a model effluent by polyphenoloxidases and horseradish peroxidase/hydrogen peroxide system* AB - An effluent containing soluble lignin fragments was treated with potato polyphenoloxidases (PPO) or horseradish peroxidase/hydrogen peroxide system (HRP/H(2)O(2)). In both cases the reaction was evidenced by the formation of a brown precipitate that was a consequence of the polymerization of lignin fragments. The effect of reaction time, pH, and amount of soluble lignin per unit of enzyme activity on the insolubilization yield was evaluated for PPO-initiated reactions. For HRP-initiated reactions, the amount of H(2)O(2) per unit of enzyme activity was also evaluated. Mathematical models were calculated to predict the insolubilization yield as a function of the significant variables. Based on these models, the insolubilization reaction was optimized and reached maximal values of ca. 50% in both reaction systems. Higher insolubilization yields were not achieved. Chemical characterization of the soluble lignin fragments indicated that the insolubilization yield could not be improved, because the lignin fragments had limited amounts of free phenolic substructures available for the initial steps of the polymerization reaction. PMID- 10713203 TI - Evidence of Pluronic F-68 direct interaction with insect cells: impact on shear protection, recombinant protein, and baculovirus production* AB - Pluronic F-68 has been widely used to protect animal cells from hydrodynamic stress, but its mechanism of action is still debatable. Published evidence indicates that Pluronic F-68 interacts with cells, yet scarce information exists of its effect on recombinant protein and virus production by insect cells. In this work, the effect of Pluronic F-68 on production of recombinant baculovirus and rotavirus protein VP7 was determined. Evidence of Pluronic F-68 direct interaction with Sf-9 insect cells also was obtained. Maximum recombinant VP7 concentration and yield increased 10x, whereas virus production decreased by 20x, in spinner flask cultures with 0.05% (w/v) Pluronic F-68 compared to controls lacking the additive. No differences were observed in media rheology, nor kinetics of growth and infection (as inferred from cell size) between both cultures. Hence, Pluronic F-68 influenced cell physiology independently of its shear protective effect. Cells subjected to a laminar shear rate of 3000 s(-1) for 15 min, without gas/liquid interfaces, were protected by Pluronic F-68 even after its removal from culture medium. Furthermore, the protective action was immediate in vortexed cells. The results shown here indicate that Pluronic F-68 physically interacts with cells in a direct, strong, and stable mode, not only protecting them from hydrodynamic damage, but also modifying their capacity for recombinant protein and virus production. PMID- 10713204 TI - Comparative study of thylakoid membranes sensitivity for herbicide detection after physical or chemical immobilization. AB - A micro-test using immobilized thylakoid membranes as sensing element in a micro electrochemical cell has been developed to assess impairment at the level of the light-driven transport of electrons. In this study, thylakoids isolated from spinach leaves were either immobilized by entrapment in poly(vinylalcohol) bearing styrylpyridinium groups or by chemical immobilization in an albumin glutaraldehyde crosslinked matrix. The two immobilization procedures were compared upon the sensitivity of the immobilized materials to detect nine herbicides targetting photosystem II. Despite the largely differing mode of immobilization, the procedures led to strikingly similar detection capabilities for herbicides. Inherent characteristics of both immobilization procedures are also discussed. PMID- 10713205 TI - Soybean peroxidase as an effective bromination catalyst* AB - Soybean peroxidase (SBP), an acidic peroxidase isolated from the seed coat, has been shown to be an effective catalyst for the oxidation of a variety of organic compounds. In the present study, we demonstrate that SBP can catalyze halogenation reactions. In the presence of H(2)O(2), SBP catalyzed the oxidation of bromide and iodide but not chloride. Veratryl alcohol (3,4-dimethoxybenzyl alcohol) served as a useful substrate for SBP-catalyzed halogenations yielding the 6-bromo derivative. Halogenation of veratryl alcohol was optimal at pHs below 2.5 with rates of 2.4 um/min, achieving complete conversions of 150-um veratryl alcohol in 24 h. The enzyme showed essentially no brominating activity at pHs above 5.5. SBP-catalyzed bromination of veratryl alcohol proceeded with a maximum reaction velocity, (V(max))(apparent), of 5.8 x 10(-1) um/min, a K(m) of 78 um and a catalytic efficiency (k(cat)/K(m) of 1.37 x 10(5) M/min at pH 4.0, assuming all of the enzyme's active sites participate in the reaction. SBP also catalyzed the bromination of several other organic substrates including pyrazole to produce a single product 1-bromopyrazole, indole to yield both 5-bromoindole and 5 hydroxyindole, and the decarboxylative bromination of 3,4 dimethoxy-trans cinnamic acid to trans-2-bromo-1-(3,4 dimethoxyphenyl)ethylene. A catalytic mechanism for SBP-catalyzed bromination has been proposed based on experimental results in this and related studies. PMID- 10713206 TI - Influence of water activity and aqueous solvent ordering on enzyme kinetics of alcohol dehydrogenase, lysozyme, and beta-galactosidase. AB - Effects of the water activity (a(w)) and the solvent ordering, as determined by the activity coefficient of water, were investigated on the enzyme kinetics of alcohol dehydrogenase, lysozyme, and beta-galactosidase in various aqueous solutions. The water activity and the solvent ordering were adjusted by addition of electrolytes (NaCl, KCl, CsCl, etc.) or nonelectrolytes (sugars, alcohols, urea, etc.) at various concentrations. Although the enzyme kinetics were strongly dependent on a(w), a(w) was not a complete determinant of the enzyme behavior in aqueous solutions. Enzyme kinetics were also dependent on the solvent ordering. At a fixed a(w), all the enzyme kinetic parameters tested had a good correlation with the solvent ordering parameter as represented by the parameter alpha, an index of the deviation of the water state from the ideal solution, determined from the activity coefficient of water in solutions. Solvent ordering was expected to affect the enzyme kinetics through its effect on the hydrophobic interaction between the enzyme and the substrate and also on the thermal fluctuation. PMID- 10713207 TI - Synthesis of allysine ethylene acetal using phenylalanine dehydrogenase from Thermoactinomyces intermedius. AB - Allysine ethylene acetal [(S)-2-amino-5-(1,3-dioxolan-2-yl)-pentanoic acid (2)] was prepared from the corresponding keto acid by reductive amination using phenylalanine dehydrogenase (PDH) from Thermoactinomyces intermedius ATCC 33205. Glutamate, alanine, and leucine dehydrogenases, and PDH from Sporosarcina species (listed in order of increasing effectiveness) also gave the desired amino acid but were less effective. The reaction requires ammonia and NADH. NAD produced during the reaction was recyled to NADH by the oxidation of formate to CO(2) using formate dehydrogenase (FDH). PDH was produced by growth of T. intermedius ATCC 33205 or by growth of recombinant Escherichia coli or Pichia pastoris expressing the Thermoactinomyces enzyme. Using heat-dried T. intermedius as a source of PDH and heat-dried Candida boidinii SC13822 as a source of FDH,98%, but production of T. intermedius could not be scaled up. Using heat-dried recombinant E. coli as a source of PDH and heat-dried Candida boidinii 98%. In a third generation process, heat-dried methanol-grown P. pastoris expressing endogenous FDH and recombinant Thermoactinomyces98% ee. PMID- 10713208 TI - A poly(N-isopropylacrylamide-co-N-acryloxysuccinimide-co-2-hydroxyethyl methacrylate) composite hydrogel membrane for urease immobilization to enhance urea hydrolysis rate by temperature swing* AB - A composite membrane made of cross-linked poly(N-isopropylacrylamide-co-N acryloxysuccinimide-co-2-hydroxyethyl methacrylate) (p(NIPAAm-NAS-HEMA)) hydrogel on polyester nonwoven support has been synthesized. The composite membrane shows temperature-responsive properties similar to conventional PNIPAAm hydrogels beads, which reversibly swells below and de-swells above the lower critical solution temperature of PNIPAAm (around 32 to 33 degrees C). Diffusion of urea through the membrane was temperature-dependent with the effective diffusion coefficient at 20 degrees C being 18 times that at 60 degrees C. Urease was immobilized directly to the membrane by forming covalent bonds between its amino groups and the succinimide ester groups of the membrane. Membrane prepared with NIPAAm to NAS molar ratio of 9, and then reacted in pH 7 buffer with 6 mg of urease gave the best immobilized enzyme, where 0.102 mg protein and 5.71 U activity per cm(2) membrane, and 55% relative specific activity could be obtained. There was negligible internal mass transfer resistance for this preparation judging from the calculated effectiveness factor. Urease shows enhanced thermal stability after immobilization with the first-order inactivation rate constant at 70 degrees C decreased to 1/8 of that of free urease. Membrane immobilized urease could be utilized in a two-compartment membrane reactor with temperature swing to substantially enhance urea hydrolysis rate. The best operating condition of the membrane reactor was with temperature cycling between 60 to 20 degrees C and with temperature change every 10 min, where concentration of product ammonia after 3 h reaction increased 3.8-folds when compared with isothermal operation at 60 degrees C. PMID- 10713209 TI - Nitrile biotransformations using free and immobilized cells of a thermophilic Bacillus spp. AB - A thermophilic Bacillus spp. capable of transforming aliphatic nitriles, cyclic nitriles and dinitriles was used as a free cell suspension and immobilized in alginate beads to study the utilization of acetonitrile and acrylonitrile in a buffered biotransformation medium. The cells grew optimally at 65 degrees C and contained a nitrile hydratase-amidase enzyme system that transformed nitrile compounds stoichiometrically to the corresponding carboxylic acids. In the presence of urea or chloroacetone, amidase activity was inhibited and the amide intermediate was accumulated. Mass transfer limitation of nitrile utilization rates was observed with immobilized cells, but the alginate afforded the cells some degree of additional thermal stability and potential advantage in re-use. In vitro inhibition of the partially purified amidase was confirmed and the use of whole cells of this organism in a continuous bioreactor to generate amide products from nitrile substrates was demonstrated. PMID- 10713210 TI - Characterization of specificity of subtilisin Carlsberg towards peptide T by high performance liquid chromatography and electrospray mass spectrometry. AB - Peptide T has a sequence (Ala-Ser-Thr-Thr-Thr-Asn-Tyr-Thr) belonging to HIV envelope that is involved in the interaction with CD(4) receptor of T lymphocytes. Research of protease activities towards this peptide is very relevant for AIDS therapy. Characterization of specificity of subtilisin Carlsberg towards this very hydrophilic peptide is proposed by using high performance liquid chromatography and mass spectrometry. Peptide T was totally hydrolysed by the protease after 24 h. Separation of hydrophilic fragments was perfected with an hydrophilic stationary phase and a reversed acetonitrile gradient. Peptide masses were determined by ion spray mass spectrometry. Four primary and one secondary hydrolysis products were found, corresponding to cleavage at the carboxylic side of threonine. Specifities of subtilisin Carlsberg towards the Segments 19 to 26 of bovine pancreatic ribonuclease A, an homologous fragment of peptide T, and peptide T were compared. PMID- 10713211 TI - Effect of environmental conditions on biological decolorization of textile dyestuff by C. versicolor. AB - Effects of environmental conditions such as pH, media composition, carbon and nitrogen sources, TOC/N ratio, and dyestuff concentrations on decolorization of reactive phytalocyanin type textile dyestuff Everzol Turquoise Blue G by white rot fungi, Coriolus versicolor 20) or low nitrogen concentration was essential for effective decolorization of the dyestuff. Dyestuff concentration should be lower than 500 mg/l for complete decolorization. Only partial decolorization was observed for dyestuff concentrations above 500 mg/l. Adsorption of the dyestuff on surfaces of the fungi was insignificant (<20%). PMID- 10713212 TI - Effect of protein hydrolysate on the degradation of diesel fuel in soil* AB - The addition of protein hydrolysate solution to soil contaminated with diesel fuel was investigated for effects on diesel degradation. The application of protein hydrolysate solution led to an increase in the removal of diesel from the soil. At the end of the 21d experimental period the amount of diesel removed from the soil was 21% greater with the addition of protein hydrolysate solution when compared to a control system. This increased removal was linked to increases in both the number of hydrocarbon degrading micro-organisms, and an extension to their period of activity. PMID- 10713213 TI - The influence of culture conditions on mycelial structure and cellulase production by Trichoderma reesei Rut C-30. AB - The morphology of Trichoderma reesei Rut C-30, during submerged cultivations in shake flask, was examined. The influence of the size inoculum and the composition of the fermentation medium on the morphology and cellulase production was studied. Different inoculum sizes were studied but the significative change in fungus morphology was observed for spores concentration between 10(5) and 10(7) spores/ml (i.e. 10(2) and 10(4) spores/ml in pre-culture medium). In the medium without Tween 80, at low inoculum size, the majority of the pellets were large and well individualized, in contrast, at higher inoculation densities small flocs were obtained, with higher production of soluble protein and higher filter paper activity. It was found that the average pellet size seems to be inversely proportional to the inoculum size. Medium composition, namely Tween 80, also influences the morphology of T. reesei Rut C-30 and enzyme production. The presence of Tween 80 in fermentation medium inhibited the pellet formation of this strain. PMID- 10713214 TI - Investigation of acid proteinase biosynthesis by the fungus Humicola lutea 120-5 in an airlift bioreactor. AB - Acid proteinase production using filamentous fungus Humicola lutea 120-5 was studied under batch and continuous fermentation conditions in an airlift bioreactor. A comparison with proteinase production by fungal cells, cultivated in stirred tank bioreactor was made. The process performance in both fermentation devices was similar with respect to substrate utilization, biomass, and enzyme concentration. Continuous acid proteinase production was achieved for 14 days at an optimal dilution rate of 0.05/h with maximum specific activity of 90 U/mg DW of mycelia and yield of 38 U/mg glucose. The volumetric productivity (50 U/ml. h) was approximately 3 times higher than this of the batch system. All continuous experiments were carried out without any bacterial contamination, due to the low pH (3.0-3.5) during the process. The "pellet" type growth of the fungus in the airlift reactor prevented the system from plugging with filaments. PMID- 10713215 TI - Production and purification of protease from a Bacillus subtilis that can deproteinize crustacean wastes* AB - A protease-producing microorganism was isolated in northern Taiwan and identified as a strain of Bacillus subtilis. B. subtilis Y-108 thus isolated can be used for deproteinization of crustacean wastes in the preparation of chitin. For deproteinization tests, liquid phase fermentation of untreated shrimp shell, crab shell, and lobster shell wastes with this microbe showed protein removal of 88, 67, and 83%, respectively. In contrast, the protein removal of the acid treated wastes was 76, 62, 56%, respectively. The optimized conditions for protease production was found when the culture was shaken at 30 degrees C for 3 days in 100 ml of medium (phosphate buffer adjusted to pH 6.0) containing 7% shrimp and crab shell powder (SCSP), 0.1% K(2)HPO(4), 0.05% MgSO(4), 1.0% arabinose, 1.5% NaNO(3), and 1.5% CaCl(2). Under such conditions, the protease of B. subtilis Y 108 attained the highest activity. It was as high as 20.2 U/ml. The protease was purified in a three-step procedure involving ammonium sulfate precipitation, DEAE Sepharose CL-6B ionic exchange chromatography, and Sephacryl S-200 gel permeation chromatography. The enzyme was shown to have a relative molecular weight of 44 kDa by SDS polyacrylamide gel electrophoresis. The protease was most active at pH 8.0 and 50 degrees C with casein as substrate. The protease was activated by Mn(+2), Fe(+2), Zn(+2), Mg(+2), Co(+2), but inhibited completely by Hg(+2). The protease was also inhibited by metal-chelating agent such as EDTA, sulfhydryl reagents as beta-mercaptoethanol, and by cysteine hydrochloride, histidine, glycerol. The EDTA was the most effective inhibitor that caused complete inhibition of protease. It was concluded that this enzyme is a metal-chelator sensitive neutral protease. PMID- 10713216 TI - Clues for the existence of two different epoxide hydrolase activities in the fungus Beauveria bassiana. AB - Epoxide hydrolase activity was produced during the exponential and stationary growth phases of the fungus Beauveria bassiana ATCC 7159. It was completely cell associated. After cell disruption epoxide hydrolase activity was recovered in both the cell debris (EH "A") and the soluble fraction (EH "B"), but not in the membrane fraction. Activity assays of these fractions with two different substrates indicated that their substrate specificity, as well as the corresponding E value and, to a lesser extent, their regioselectivity, were different. Also, we could observe that the absolute configuration of the residual epoxide was opposite. This indicates that these two epoxide hydrolase activities are substantially different and are, therefore, interestingly complementary biocatalysts for the preparation of the corresponding epoxides and/or vicinal diols in nearly enantiopure form. PMID- 10713217 TI - Purification and characterization of an extracellular lipase from a thermophilic Rhizopus oryzae strain isolated from palm fruit. AB - We have isolated a lipolytic strain from palm fruit that was identified as a Rhizopus oryzae. Culture conditions were optimized and highest lipase production amounting to 120 U/ml was achieved after 4 days of cultivation. The extracellular lipase was purified 1200-fold by ammonium sulfate precipitation, sulphopropyl Sepharose chromatography, Sephadex G 75 gel filtration and a second sulphopropyl Sepharose chromatography. The specific activity of the purified enzyme was 8800 U/mg. The lipolytic enzyme has a molecular mass of 32 kDa by SDS-polyacrylamide gel electrophoresis and gel filtration. The enzyme exhibited a single band in active polyacrylamide gel electrophoresis and its isoelectric point was 7.6. Analysis of Rhizopus oryzae lipase by RP-HPLC confirmed the homogeneity of the enzyme preparation. Determination of the N-terminal sequence over 19 amino acid residues showed a high homology with lipases of the same genus. The optimum pH for enzyme activity was 7.5. Lipase was stable in the pH range from 4.5 to 7.5. The optimum temperature for lipase activity was 35 degrees C and about 65% of its activity was retained after incubation at 45 degrees C for 30 min. The lipolytic enzyme was inhibited by Triton X100, SDS, and metal ions such as Fe(3+), Cu(2+), Hg(2+) and Fe(2+). Lipase activity against triolein was enhanced by sodium cholate or taurocholate. The purified lipase had a preference for the hydrolysis of saturated fatty acid chains (C(8)-C(18)) and a 1, 3-position specificity. It showed a good stability in organic solvents and especially in long chain-fatty alcohol. The enzyme poorly hydrolyzed triacylglycerols containing n-3 polyunsaturated fatty acids, and appeared as a suitable biocatalyst for selective esterification of sardine free fatty acids with hexanol as substrate. About 76% of sardine free fatty acids were esterified after 30 h reaction whereas 90% of docosahexaenoic acid (DHA) was recovered in the unesterified fatty acids. PMID- 10713218 TI - Comparison of simple neural networks and nonlinear regression models for descriptive modeling of Lactobacillus helveticus growth in pH-controlled batch cultures. AB - A set of 20 Lactobacillus helveticus growth curves was obtained from pH controlled batch cultures with different pH setpoints, whey permeate and yeast extract concentrations. To find the best descriptive model of the biomass concentration versus time (y = X(t)) growth curve, fitting results of a large number of models were compared with statistical and approximate methods. Models studied included simple neural networks, reparameterized Logistic, Gompertz, Richards, Schnute, Weibull, and Morgan-Mercier-Flodin models, Amrane-Prigent model, and four new models based on autonomous growth functions. Simple neural networks with only four weights were good descriptive models of the growth curves and fitting qualities were similar to those of the best existing four-parameter models, such as the Logistic model. However, meaningful parameters had to be calculated numerically and use of simple neural networks yielded no distinctive advantages over other models. A new five-parameter model, based on an autonomous growth function, yielded the best fitting results, even when the number of model parameters was accounted for in the comparisons. However, the maximum specific growth rate was not always well estimated. Therefore the five-parameter Richards model was chosen as the best descriptive model of the growth curve. PMID- 10713219 TI - Enzymatic esterification of ethanol by an immobilised Rhizomucor miehei lipase in a perforated rotating disc bioreactor. AB - A perforated rotating disc bioreactor was developed to perform the esterification of ethanol with oleic acid, catalyzed by a lipase from Rhizomucor miehei immobilized by adsorption on to a hydrophobic support-Accurel EP700. The bioreactor with total recirculation operated at an optimum agitation rate of 400 rev./min. The experimental results, in this condition, were predict by a kinetic model using the constants obtained in the batch (Erlenmeyer flasks) assays: a catalytic constant, k(cat) = 5.78 mmol/h. mg protein; a Michaelis constant for ethanol, K(m(Et)) = 1.20 M; a Michaelis constant for oleic acid, K(m(Ol)) = 1.16 x 10(-8) M, and a dissociation constant of the ethanol-lipase complex, K((Et)) = 9.46 x 10(7) M. The efficiency of conversion gradually decreased during continuous operation of the reactor. The enzymatic activity decayed according to a first order deactivation model and the integrated equations of a continuous stirred tank reactor (CSTR) and a plug flow reactor (PFR). A half-life time of the lipase of about 10 days and a deactivation constant of 0.003 h(-1) were obtained in the present system. PMID- 10713220 TI - Critical evaluation of p-nitrophenylphosphorylcholine (p-NPPC) as artificial substrate for the detection of phospholipase C* AB - Phospholipase C (PLC) activity secreted by bacteria as a virulence factor is commonly detected by use of the artificial substrate p nitrophenylphosphorylcholine (p-NPPC). We examined several commercially available enzymes (phosphodiesterases, phosphomonoesterases, phospholipase A, lipase, protease) for their hydrolytic activity towards p-NPPC and compared these results with those of PLC tests using phospholipid substrates. Our data indicate that, in addition to PLC, several other enzymes which can affect phosphate esters are able to hydrolyze p-NPPC. We therefore suggest to use lipid substrates for correct characterization of bacterial PLCs, especially when whole bacteria or crude enzyme preparations are investigated. PMID- 10713221 TI - Isolation and properties of a cellulosome-type multienzyme complex of the thermophilic Bacteroides sp. strain P-1. AB - The extracellular form of cellulosome-type multienzyme complex of thermophilic Bacteroides sp. strain P-1 which was isolated from the anaerobic digester, is described. Multienzyme complex was isolated from the culture supernatant by an adsorption-desorption affinity chromatography on microcrystalline cellulose. The isolated multienzyme complex was found to form a complex that exhibited a high molecular weight (estimated at more than 1400 kDa) and was quite stable, requiring strong denaturing condition for dissociation. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate resolved multienzyme complex into at least 12 subunits with the molecular weight range of 49 to 209 kDa, respectively. The isolated multienzyme complex showed cellulose-binding ability, cellulase and xylanase activities and effected the hydrolysis of crystalline cellulose and lignocellulosic materials in the form of corncob, corn hull, rice straw, and sugarcane bagasse. PMID- 10713222 TI - LASIK mania. PMID- 10713223 TI - Creating cataracts of varying hardness to practice extracapsular cataract extraction and phacoemulsification(2). AB - We describe a method to harden the nucleus of crystalline lenses in postmortem human globes by intralenticular injection of a mixture of paraformaldehyde and glutaraldehyde (Karnovsky's solution). Evaluation of the nuclear manipulations, using the Miyake-Apple posterior video/photographic technique, shows that the method provides suitable specimens for practicing extracapsular cataract extraction and phacoemulsification. PMID- 10713224 TI - Interlenticular opacification: clinicopathological correlation of a complication of posterior chamber piggyback intraocular lenses. AB - PURPOSE: To present a clinicopathological correlation of 2 pairs of piggyback posterior chamber intraocular lenses (PC IOLs) explanted because of opacification between the lens optics. SETTING: Gayton Health Center, Eyesight Associates of Middle Georgia, Warner Robins, Georgia, and Center for Research on Ocular Therapeutics and Biodevices, Storm Eye Institute, Medical University of South Carolina, Charleston, South Carolina, USA. METHODS: Two pairs of piggyback AcrySof lenses were explanted from 2 patients with significant visual loss related to opacification between the optics. They were submitted for pathological analysis. Gross and histopathological examinations were performed, and photomicroscopy was used to document the results. RESULTS: Gross examination showed accumulation of a membrane-like white material between the lenses. Histopathological examination revealed that the tissue consisted of retained/proliferative lens epithelial cells (bladder cells or pearls) mixed with lens cortical material. CONCLUSION: Piggyback PC IOLs were explanted in 2 cases because of a newly described complication, interlenticular opacification. Three surgical means may help prevent this complication: meticulous cortical cleanup, especially in the equatorial region; creation of a relatively large continuous curvilinear capsulorhexis to sequester retained cells peripheral to the IOL optic within the equatorial fornix; insertion of the posterior IOL in the capsular bag and the anterior IOL in the ciliary sulcus to isolate retained cells from the interlenticular space. PMID- 10713225 TI - Corneal sensation after laser in situ keratomileusis. AB - PURPOSE: To report the time course for the return of corneal sensation following laser in situ keratomileusis (LASIK). SETTING: University-based retractive surgery practice. METHODS: Twenty-eight eyes of 18 patients having LASIK were evaluated. Preoperative and postoperative corneal sensation at the nasal flap hinge, at the central cornea, and within the temporal flap edge were measured before and after LASIK for a 3 week period using the Cochet-Bonnet esthesiometer (Luneau). RESULTS: Corneal sensation initially decreased in all 3 positions of the flap measured after LASIK; the greatest decrease was in the central cornea. Near preoperative corneal sensation returned by 3 weeks. The degree of sensation loss did not appear to correlate with the ablation depth. CONCLUSION: Corneal sensation is significantly decreased for approximately 2 to 3 weeks after LASIK, centrally greater than nasally at the flap hinge or temporally within the flap edge, but it generally returns to near the preoperative level by 3 weeks postoperatively. PMID- 10713227 TI - Eye movement during laser in situ keratomileusis. AB - PURPOSE: To measure eye motion in patients having laser in situ keratomileusis (LASIK) using a video technique and determine centration and variance of the eye position during surgery. SETTING: Laser refractive surgery center. METHODS: The procedure was videotaped in 5 consecutive eyes having LASIK performed by a single surgeon with the VISX Star S2 excimer laser. Following surgery, video images of the eyes were digitized and stored in a computer for processing. Digitized images were obtained at a rate of 25 images per second during the laser procedure. The pupil margin and a visual landmark, such as a scleral blood vessel, were identified in the initial image of each eye. Custom software was used to track the location of the landmark and the pupil center in subsequent images. RESULTS: Three of the 5 eyes were well centered on average. The remaining 2 eyes were decentered inferiorly by approximately 0.25 mm. The standard deviation in all eyes was approximately 0.10 mm. CONCLUSIONS: With these techniques, the position of the entrance pupil center relative to the excimer laser axis could be determined. Although the system is not fast enough to be used during surgery, it does allow quantification of centration and intraoperative motion after surgery. PMID- 10713226 TI - Retinal detachment in myopic eyes after photorefractive keratectomy. AB - PURPOSE: To analyze the incidence and characteristics of retinal detachment (RD) in myopic patients who had photorefractive keratectomy (PRK). SETTING: Universidad Miguel Hernandez, Instituto Oftalmologico de Alicante, Alicante, Spain. METHODS: The incidence of RD in 5936 consecutive eyes that had PRK to correct myopia was studied. Mean follow-up was 38.5 months +/- 17.4 (SD). RESULTS: Retinal detachment occurred in 5 eyes (0.08%); 2 in women and 3 in men. The mean interval between PRK and RD was 21. 00 +/- 15.89 months (range 9 to 48 months). The mean best corrected visual acuity (BCVA) after PRK and before RD development was 20/81 (range 20/200 to 20/25). After RD repair, the mean BCVA was 20/460 (range 20/2000 to 20/29). In 4 of the 5 eyes, BCVA after RD was within 1 line of the preoperative value; in 1 eye, it decreased from 20/40 to 20/2000. The mean spherical equivalent (SE) before RD treatment was -1.35 +/- 1.08 diopters (D) (range 0 to -3.00 D) and after RD treatment, -2.95 +/- 0.83 D (range -2.00 to -4.00 D). Differences between SE before and after RD treatment were statistically significant (P =.01, paired Student t test). CONCLUSIONS: The incidence of RD after PRK to correct myopia was 0.08%. In 4 of 5 eyes, there was little or no visual loss; but in the group as a whole, there was a significant increase in myopic SE. PMID- 10713228 TI - Apical nodular subepithelial corneal scar after retreatment in hyperopic photorefractive keratectomy. AB - PURPOSE: To report a complication, apical nodular subepithelial corneal scar, that can occur after hyperopic photorefractive keratectomy (PRK) retreatment. SETTING: Istanbul University Eye Research Center, Istanbul, Turkey. METHODS: Twelve eyes of 6 patients with apical nodular subepithelial corneal scar were retrospectively studied. Mean age of the 5 men and 1 woman was 30.2 years +/- 5.4 (SD). All eyes had hyperopic PRK retreatment for regression 9.5 +/- 1.44 months after primary hyperopic PRK. Mean spherical equivalent refraction of the residual hyperopia before retreatment was +4.67 +/- 0.81 diopters (D). All patients had a corneal haze grade of less than 1+. Hyperopic PRK retreatment was performed with a 193 nm excimer laser (Chiron Technolas Keracor 116). RESULTS: Apical nodular subepithelial corneal scars developed within the first month of hyperopic PRK retreatment and progressed until the third month in 12 eyes of 6 patients. The lesion was round and symmetrical in both eyes, smaller than 2.0 mm, and centrally located. During the mean 40.66 +/- 2.46 month follow-up, the lesion did not change in size or density. Mean spherical equivalent refraction after retreatment was 2.88 +/- 0.88 D (range +1.50 to +4.00 D) at 1 month and +4.36 +/- 1.83 D at last follow-up. Refraction was unmeasurable in 3 eyes. Five eyes lost 1 line of best spectacle-corrected visual acuity and 7 eyes, 2 or more lines. The surface regularity indexes were higher than 2.00 in all the eyes. CONCLUSION: Hyperopic PRK retreatment may cause the sight-threatening complication of apical nodular subepithelial corneal scar. This complication behaves unlike corneal haze and does not resolve spontaneously over time. PMID- 10713229 TI - Pathogenesis and management of laser in situ keratomileusis flap buttonhole. AB - PURPOSE: To describe the clinical features and outcomes in patients who had a flap buttonhole during laser in situ keratomileusis (LASIK) and propose an etiopathogenic mechanism for this complication. SETTING: University Eye Center, Department of Ophthalmology and Visual Sciences, Prince of Wales Hospital, The Chinese University of Hong Kong, Hong Kong, China. METHODS: Retrospective review of case records of 6 patients (6 eyes) who had a flap buttonhole during LASIK. RESULTS: The mean patient age was 38.2 years +/- 4.1 (SD) and the mean preoperative spherical equivalent (SE) refraction, -8.13 +/- 4.04 diopters (D). Mean keratometry was 44.20 +/- 1.30 D. Retreatment was performed after a mean interval of 9.2 +/- 3.2 months. Final postoperative SE refraction was -0.44 +/- 0.58 D after a mean follow-up of 59.0 +/- 5.3 weeks. No patient experienced loss of best spectacle-corrected visual acuity. CONCLUSIONS: Retreatment of eyes that have a flap buttonhole during LASIK is associated with good visual outcomes. Flap buttonholes can produce alterations in refraction, so retreatment is best performed after the refractive error has stabilized. Microkeratome malfunction may be responsible for the occurrence of a flap buttonhole during LASIK in eyes that do not have significant corneal steepening. PMID- 10713230 TI - Photorefractive keratectomy for myopia with a 15 Hz repetition rate. AB - PURPOSE: To evaluate excimer laser photorefractive keratectomy (PRK) for myopia using a repetition rate of 15 Hz instead of 10 Hz. SETTING: The Cornea and Laser Eye Institute, Teaneck, and Department of Ophthalmology, UMDNJ-New Jersey Medical School, Newark, New Jersey, USA. METHODS: Photorefractive keratectomy using a 15 Hz repetition rate was performed in 23 eyes of 14 patients by a single surgeon at 1 center. The attempted corrections ranged from -2.8 diopters (D) to -5.5 D. Preoperative and postoperative uncorrected visual acuity (UCVA), best spectacle corrected visual acuity (BSCVA), predictability, corneal haze, and subjective glare/halo were evaluated over 6 months. RESULTS: At 6 months, UCVA was 20/32 or better in all eyes and at least 20/20 in 14 eyes (73.7%). Two eyes (10.5%) lost 2 or more Snellen lines of BSCVA; postoperative BSCVA was at least 20/25 in 100% of eyes and 20/20 or better in 95.0%. Fifteen eyes (78.9%) were within +/-0.5 D of attempted correction, and 19 (100%) were within +/-1.0 D. Mean spherical equivalent refraction was -4.62 D preoperatively, +0.15 D at 1 month, -0.09 D at 3 months, and -0.37 D at 6 months. At 6 months, 4 eyes (21.0%) had no corneal haze and 14 (73.7%) had trace subepithelial haze. Fifteen eyes (78.9%) had no glare/halo effect at 6 months, and 4 (21.0%) had minimal glare/halo effect. CONCLUSIONS: Clinical outcomes after excimer laser PRK for myopia using an increased repetition rate of 15 Hz were good and similar to those in studies conducted with a 10 Hz repetition rate. PMID- 10713231 TI - Phakic posterior chamber lenses for high myopia: functional and anatomical outcomes. AB - PURPOSE: To evaluate the functional and the anatomical outcomes after implantation of phakic posterior chamber intraocular lenses (IOLs) in highly myopic eyes. SETTING: Service d'Ophtalmologie, Hopital Purpan, Toulouse, France. METHODS: Fifty-eight eyes of 46 patients that had implantation of phakic posterior chamber lenses for high myopia were evaluated. Predictability, efficiency, safety, and subjective and objective quality of vision were evaluated preoperatively and at least 6 months postoperatively. The effect of the procedure on the cornea, aqueous humor, pupil, anterior chamber angle, crystalline lens, and retina were studied. RESULTS: Mean preoperative myopia was -13.85 diopters (D) +/- 3.1 (SD) (range -8.00 to -19.25 D). Mean postoperative spherical equivalent was -1.22 +/- 0.83 D (range +0.75 to -3.50 D); 56.9% of eyes were within +/-1.00 D of the predicted result, and 77.6% gained 1 or more lines of best corrected visual acuity. All contact-lens-intolerant patients had improved quality of vision for day and night driving, distance vision, and vision under dim illumination. The mean postoperative level of contrast sensitivity without correction was higher than the mean preoperative level with correction. Adverse events were 2 cases of crystalline lens opacification 16 and 18 months after surgery and 2 cases of pigment deposits in the angle with increased intraocular pressure, which was controlled by beta-blockers. CONCLUSION: Implantation of posterior chamber phakic IOLs is effective and predictable; however, long-term follow-up is needed. PMID- 10713232 TI - Black diaphragm aniridia intraocular lens for congenital aniridia: long-term follow-up. AB - PURPOSE: To present long-term results of implantation of a black diaphragm aniridia intraocular lens (IOL) in eyes with congenital aniridia. SETTING: Eye Hospital, Heinrich-Heine-University, Dusseldorf, Germany. METHODS: Cataract surgery was performed in 19 eyes of 14 patients with congenital aniridia. The black diaphragm aniridia IOL was implanted in front of the capsular bag in the ciliary sulcus. Mean patient age was 30 years (range 10 to 59 years) and mean follow-up, 46 months (range 12 to 84 months). Before surgery, corneal epithelial disorders; corneal pannus; cataract; hypoplasia of the macula, optic nerve, or both; and nystagmus were present in all 19 eyes. Clinically detectable glaucoma was present in 5 eyes. RESULTS: Despite the presence of amblyopia and nystagmus, visual acuity improved in 14 of the 19 eyes. The main postoperative problems were glaucoma deterioration (4 of 19 eyes) or development (4 of 19 eyes), cystoid macular edema (2 of 11 eyes), chronic endothelial cell loss (3 of 11 eyes), and progression of corneal epithelial disorders (4 of 19 eyes). Glaucoma was controlled by medical or surgical therapy in all patients. Intraocular lens explantation was performed in 2 eyes with glaucoma. CONCLUSION: Implantation of the black diaphragm aniridia IOL improved visual acuity in the majority of patients with a variety of endogenous problems in addition to aniridia. PMID- 10713233 TI - Effect of tropicamide on aqueous flare before and after cataract surgery. AB - PURPOSE: To examine the effect of tropicamide on flare intensity under phakic and pseudophakic conditions and to differentiate between the possible mechanisms of action of tropicamide on aqueous flare. SETTING: Department of Ophthalmology, Vienna General Hospital, University of Vienna, Vienna, Austria. METHODS: In this prospective study, aqueous flare was measured with the laser flare-cell meter in 20 eyes of 20 patients with age-related cataract enrolled for cataract surgery. Measurements were performed before and 30, 90, and 180 minutes after pupil dilation with tropicamide 0.5%. This measurement was performed in the phakic eye on the day before surgery and in the pseudophakic eye on postoperative days 1, 3, 7, and 28. RESULTS: After tropicamide instillation, aqueous flare decreased preoperatively and on all postoperative days. There was a continuous flare decrease until 3 hours after instillation, reaching a maximum decrease of about 30%. Pupil diameter reached its maximum after 30 minutes. CONCLUSION: Tropicamide significantly decreased aqueous flare, seemingly by pharmacological means, not volumetric changes. The time between drug application and measurement should be kept constant. PMID- 10713234 TI - Latanoprost versus timolol gel to prevent ocular hypertension after phacoemulsification and intraocular lens implantation. AB - PURPOSE: To evaluate the efficacy of latanoprost and timolol gel in preventing ocular hypertension in the early period after phacoemulsification and posterior chamber intraocular lens (PC IOL) implantation. SETTING: Department of Ophthalmology and Visual Sciences, Prince of Wales Hospital, Hong Kong, China. METHODS: This prospective randomized double-masked clinical trial comprised patients with uncomplicated cataract having phacoemulsification with PC IOL implantation. They were randomly assigned to 1 of 3 groups: postoperative application of timolol 0.5% gel-forming solution (Timoptol-XE(R)) (Group 1), latanoprost 0.005% (Group 2), and control (Group 3). Intraocular pressure (IOP) was measured 2, 4, and 24 hours postoperatively. The anterior chamber was examined for the levels of cells and flare using slitlamp biomicroscopy. RESULTS: Group 1 had a significantly greater reduction in mean IOP 2, 4, and 24 hours after phacoemulsification and PC IOL implantation than Group 3 (P <.05). There were no significant differences between Groups 2 and 3 at any interval (P. 05). No excessive postoperative anterior chamber inflammation was observed in any group. CONCLUSIONS: A single dose of latanoprost given after phacoemulsification and PC IOL implantation did not produce a significant IOP-lowering effect when compared with a control group in the first 24 hours postoperatively. A single dose of timolol gel produced a significant postoperative IOP decrease as soon as 2 hours and up to 24 hours after surgery. Timolol gel and latanoprost are safe, but timolol is more effective than latanoprost in preventing postoperative ocular hypertension. PMID- 10713235 TI - Effect of irrigating solution and irrigation temperature on the cornea and pupil during phacoemulsification. AB - PURPOSE: To evaluate the effect of irrigation solution and temperature on pupil diameter, corneal endothelium, and corneal pachymetry during and after phacoemulsification. SETTING: Klinik Dardenne, Bonn, Germany. METHODS: Eighty patients who had cataract surgery by phacoemulsification were assigned to 1 of 4 cross-classified groups and had intraoperative irrigation with room-temperature or refrigerated fortified balanced salt solution (BSS Plus) or modified Ringer's solution. Pupil diameters were recorded at different stages during the surgery. Epithelial cell counts and pachymetry were determined before and 1 day after surgery. RESULTS: The solution temperature did not affect any parameter. The type of solution did not influence endothelial cell loss; however, the solution had a significant effect on corneal pachymetry 1 day postoperatively. The corneas irrigated with BSS Plus were less swollen than the corneas irrigated with Ringer's solution. By day 14, corneal thickness was equal among all groups. CONCLUSION: Long-term results were equally favorable in all 4 groups. However BSS Plus induced less short-term corneal swelling than Ringer's solution. From these findings, it appears that BSS Plus may decrease corneal risk in cases with compromised corneas or prolonged surgery. PMID- 10713236 TI - Incidence of anterior intraocular lens precipitates after combined phacotrabeculectomy. AB - PURPOSE: To compare the incidence of anterior intraocular lens (IOL) precipitates on silicone and poly(methyl methacrylate) (PMMA) IOLs after phacotrabeculetomy. SETTING: District general hospital in the United Kingdom. METHODS: Ninety-five consecutive eyes of 77 patients who had combined phacotrabeculectomy between April 1992 and October 1996 were retrospectively studied. A slitlamp biomicroscope was used to look for precipitates. RESULTS: Mean patient age was 75 years (range 51 to 89 years). Preoperatively, mean intraocular pressure (IOP) was 26 mm Hg +/- 5.26 (SD). Thirty plate-haptic silicone IOLs and 65 PMMA IOLs were implanted. Six months postoperatively, median corrected Snellen acuity improved from 6/18 to 6/9. The mean number of antiglaucoma medications dropped from 1.46 preoperatively to 0.29 postoperatively; 73 eyes (76.8%) had an IOP of less than 22 mm Hg with no medication. In 1 surgeon's experience, lens precipitates appeared in 10 of 59 cases (16.9%), with 7 of 14 (50.0%) in the silicone IOL group and 3 of 45 (6.7%) in the PMMA IOL group. The difference was statistically significant (P <.001, chi-square). CONCLUSION: The incidence of anterior IOL precipitates was significantly higher in patients with a silicone IOL than in those with a PMMA IOL, suggesting that use of silicone IOLs may increase the incidence of postoperative IOL precipitates. PMID- 10713237 TI - 3M diffractive multifocal intraocular lens: eight year follow-up. AB - PURPOSE: To study the long-term results of implantation of the 3M diffractive multifocal intraocular lens (IOL). SETTING: Eye Department, ASA, Arendal, Norway. METHODS: The study comprised 97 eyes in 72 patients. Follow-up was 8 years. Distance and near visual acuities, refractive results, contrast sensitivity, IOL centration, and neodymium:YAG (Nd:YAG) capsulotomies to treat posterior capsule opacification were evaluated. RESULTS: All patients without ocular pathology achieved a best corrected visual acuity (BCVA) of 0.5 or better (i.e., 98.7% had a BCVA of 1.0 or better). Without correction, 73.8% of eyes had a Jaeger acuity of J3 or better and with distance correction, 92.1%. Emmetropia or within +/-0.25 diopter (D) of it was achieved in 58.8% of eyes. An astigmatic shift of 0.827 D cylinder correction was induced. This shift was mainly against the rule (0.717 D). Contrast sensitivity was reduced with spatial frequencies of 6 to 18 cycles per degree. No need for spectacles was reported by 54.2% of patients and by 68.0% of those with bilateral implantation. For near tasks, 63.9% of patients never used spectacles. The IOLs were well centered or minimally decentered in 99.0% of eyes. Posterior capsule opacification was treated by Nd:YAG laser capsulotomy in 55.7% of eyes, with a mean time between surgery and treatment of 34.0 months +/- 23.2 (SD). CONCLUSION: This long-term study proved the 3M diffractive IOL to be safe and effective despite some reduction in contrast sensitivity at higher spatial frequencies. More than half the patients never wore spectacles; 2 of 3 patients with IOLs in both eyes never wore spectacles. Proper patient selection is crucial. PMID- 10713238 TI - Cataract surgery in the very elderly. AB - OBJECTIVE: To analyze the outcome of cataract extraction in patients aged 85 years and older (>/=85 group) and to compare their outcome with that of patients younger than 85 years (/=85 group. In this group, visual acuity in the operative eye improved in 84.3% and a visual acuity of 0.5 or better was achieved in 71.4%. A no-benefit outcome, as defined by the Catquest questionnaire, was found in 12.7% of the >/=85 group and 8.4% of the 8 linked N-acetyl-neuraminic acid (sialic acid). The enzymes required for sialic acid biosynthesis and polymerization are encoded in region A of the capsule gene complex. We here describe the enzymatic activity of the siaA gene product as determined by biochemical analysis. siaA was overexpressed in Escherichia coli and the SiaA protein was purified to homogeneity. Enzymatic assays revealed that SiaA did not accept N-acetyl glucosamine as substrate, but only N-acetyl-glucosamine-6-phosphate (EC 5.1.3.9). SiaA catalyzes the isomerization of N-acetyl-glucosamine-6-phosphate to form N acetyl-mannosamine-6-phosphate. This reaction represents the first step in capsule biosynthesis of N. meningitidis B. PMID- 10713416 TI - Small heat shock proteins, IbpA and IbpB, are involved in resistances to heat and superoxide stresses in Escherichia coli. AB - To investigate the function of Escherichia coli small heat shock proteins, IbpA and IbpB, we constructed ibpA-, ibpB- and ibpAB-overexpressing strains and also an ibpAB-disrupted strain. The ibpA-, ibpB- and ibpAB-overexpressing strains were found to be resistant not only to heat but also to superoxide stress. However, the ibpAB-disrupted strain was not more sensitive to these stresses than the wild type strain. The heat sensitivity of a rpoH amber mutant was partially suppressed by the overexpression of plac::ibpAB. These results suggest that IbpA and IbpB may be involved in the resistances to heat and oxidative stress. PMID- 10713417 TI - Protein catabolism in growing Bacillus megaterium during adaptation to salt stress. AB - Elevated concentration of NaCl in liquid medium caused a concentration-dependent growth delay (adaptation lag) and decrease in the maximal growth rate of Bacillus megaterium. The adaptation to salt stress was accompanied by transformation of some otherwise stable (long-lived; LLP) cell proteins into quickly degraded (short-lived; SLP) ones. Exposure to the strongly growth-reducing 1 M NaCl increased the size of the SLP 'pool' of intracellular proteins from about 5 to about 15% of total protein. The major intracellular proteolytic capacity of B. megaterium is represented by intracellular serine proteinases (ISP). Paradoxically, their specific activity was lowered or masked during the adaptation phase marked by increased catabolism of short-lived and/or destabilized proteins by the stress. This documents that intracellular proteolytic activity cannot be a key regulator of protein catabolism during adaptation to stress. PMID- 10713418 TI - Functional analysis of heterologous holin proteins in a lambdaDeltaS genetic background. AB - Holins are small hydrophobic proteins causing non-specific membrane lesions at the end of bacteriophage multiplication, to promote access of the murein hydrolase to their substrate. We have established a lambdaDeltaS genetic system, which enables functional expression of holins from various phages in an isogenic phage lambda background, and allows qualitative evaluation of their ability to support lysis of Escherichia coli cells. Synthesis of Holins is under control of native lambda transcription and translation initiation signals, and the temperature-sensitive CIts857 repressor. A number of different holins were tested in this study. The opposing action of phage lambda S105 and S107 holin variants in lysis timing could be confirmed, whereas we found evidence for a functionally non-homologous dual translational start motif in the Listeria phage Hol500 holin, i.e., the Hol500-96 polypeptide starting at Met-1 revealed a more distinct lytic activity as compared to the shorter product Hol500-93. The largest holin known, HolTW from a Staphylococcus aureus phage, revealed an early lysis phenotype in the lambdaDeltaSthf background, which conferred a plaque forming defect due to premature lysis. Mutant analysis revealed that an altered C-terminus and/or a V52L substitution were sufficient to delay lysis and enable plaque formation. These results suggest that the extensively charged HolTW C-terminus may be important in regulation of lysis timing. The gene 17.5 product of E. coli phage T7 was found to support sudden, saltatory cell lysis in the lambdaDeltaSthf background, which clearly confirms its holin character. In conclusion, lambdaDeltaSthf offers a useful genetic tool for studying the structure-function relationship of the extremely heterogeneous group of holin protein orthologs. PMID- 10713419 TI - MsiK-dependent trehalose uptake in Streptomyces reticuli. AB - During cultivation in the presence of trehalose Streptomyces reticuli expresses an inducible, highly specific trehalose uptake system that is absent in Streptomyces lividans. A palmitated trehalose-binding protein was identified in the cytoplasmic membrane of mycelia, extracted with the detergent Triton X-100 and purified using a trehalose affinity matrix. Immunological studies showed that within S. reticuli the synthesis of the ATP-binding protein MsiK is induced by trehalose. The data suggest that MsiK assists the trehalose ABC transporter, like the previously described ABC transport systems for maltose and cellobiose/cellotriose, respectively. PMID- 10713420 TI - Molecular cloning of two (R)-specific enoyl-CoA hydratase genes from Pseudomonas aeruginosa and their use for polyhydroxyalkanoate synthesis. AB - Two Pseudomonas aeruginosa genes, termed phaJ1(Pa) and phaJ2(Pa), homologous to the Aeromonas caviae (R)-specific enoyl-CoA hydratase gene (phaJ(Ac)) were cloned using a PCR technique to investigate the monomer-supplying ability for polyhydroxyalkanoate (PHA) synthesis from beta-oxidation cycle. Two expression plasmids for phaJ1(Pa) and phaJ2(Pa) were constructed and introduced into Escherichia coli DH5alpha strain. The recombinants harboring phaJ1(Pa) or phaJ2(Pa) showed high (R)-specific enoyl-CoA hydratase activity with different substrate specificities, that is, specific for short chain-length enoyl-CoA or medium chain-length enoyl-CoA, respectively. In addition, co-expression of these two hydratase genes with PHA synthase gene in E. coli LS5218 resulted in the accumulation of PHA up to 14-29 wt% of cell dry weight from dodecanoate as a sole carbon source. It has been suggested that phaJ1(Pa) and phaJ2(Pa) products have the monomer-supplying ability for PHA synthesis from beta-oxidation cycle. PMID- 10713421 TI - Effect of pretreatment of rubber material on its biodegradability by various rubber degrading bacteria. AB - The effect of pretreatment of several cis-1,4-polyisoprene containing rubbers on their biodegradability was examined. Tests were carried out with six recently isolated and characterized rubber degrading bacteria belonging to the genera Gordonia (strains Kb2, Kd2 and VH2), Mycobacterium, Micromonospora and Pseudomonas. All strains were able to use natural rubber (NR) as well as NR latex gloves as sole carbon source. Extraction of NR latex gloves by organic solvents resulted in an enhancement of growth for three of the selected strains. On the other hand, growth of Gordonia sp. (strain Kb2 and Kd2), Mycobacterium fortuitum NF4 and Micromonospora aurantiaca W2b on synthetic cis-1,4-polyisoprene did only occur after removal of the antioxidants, that are usually added during manufacture to prevent aging of the materials. Detailed degradation studies performed with Gordonia sp. Kb2 revealed an enhanced mineralization of pretreated NR latex gloves and mineralization of purified natural rubber (NR), indicating the actual mineralization of cis-1,4-polyisoprene rubber constituent even after removal of non-rubber constituent that may act as co-metabolic substrate and support microbial growth. Further analysis by scanning electron microscopy (SEM) clearly demonstrated the enhanced colonization efficiency of these bacteria towards pretreated NR latex gloves. Colonization was additionally visualized by staining of overgrown NR latex gloves with Schiff's reagent, and the purple color produced in the area of degradation was an evidence for the accumulation of aldehydes containing oligomers. Further enhancement of latex gloves degradation could be achieved after successive replacement of mineral salts medium during cultivation. Thereby, a rapid disintegration of untreated NR latex gloves material was accomplished by Gordonia sp. strain VH2. PMID- 10713422 TI - Temperature-independent and -dependent expression of desaturase genes in filamentous cyanobacterium Spirulina platensis strain C1 (Arthrospira sp. PCC 9438) AB - The alteration of the degree of unsaturated fatty acids in membrane lipids has been shown to be a key mechanism in the tolerance to temperature stress of living organisms. The step that most influences the physiology of membranes has been proposed to be the amount of di-unsaturated fatty acids in membrane lipids. In this study, we found that the desaturation of fatty acid to yield the di unsaturated fatty acid 18:2(9,12), in Spirulina platensis strain C1, was not regulated by temperature. As shown by the fatty acid composition and gene expression patterns, the levels of 18:1(9) and 18:2(9,12) remained almost constant either when the cells were grown at 35 degrees C (normal growth temperature) or 22 and 40 degrees C. The expression of desC (Delta9) and desA (Delta12) genes, which are responsible for the introduction of first and second double bonds into fatty acids, respectively, was not affected by the temperature shift from 35 to 22 degrees C or to 40 degrees C. Only the expression and mRNA stability of the desD gene (Delta6) that is responsible for the introduction of a third double bond into fatty acids were enhanced by a temperature shift from 35 to 22 degrees C, but not the shift from 35 to 40 degrees C. The increase in the level of desD mRNA elevated the desaturation of fatty acid from 18:2(9,12) to 18:3(6,9,12) at 22 degrees C. However, the increased level of 18:3(6,9,12) was observed after 36 h of incubation at 22 degrees C, indicating a slow response to temperature of fatty acid desaturation in this cyanobacterium. These findings suggest that the desaturation of fatty acids might not be a key mechanism in the response to the temperature change of S. platensis strain C1. PMID- 10713423 TI - Detection of sequence variation in PCR-amplified fragments of omp2 gene from three species of the family Chlamydiaceae using agarose gel electrophoresis containing bisbenzimide-PEG. AB - A simple technique providing a means for rapid genetic differentiation of chlamydial strains is described. The technique is based on a single-step sequence specific separation of PCR-amplified DNA fragments by electrophoresis in an agarose gel containing a DNA ligand - bisbenzimide-PEG. A hypervariable region at the 5' end of the omp2 gene of Chlamydiaceae species encoding the 60-kDa cysteine rich outer membrane protein was selected as a target for PCR. The appropriate fragments were amplified from strains of Chlamydia trachomatis, Chlamydophila pneumoniae, and Chlamydophila psittaci, and the PCR products originating from different species were electrophoretically separated in the presence of the DNA ligand. We therefore demonstrated that PCR with a single pair of primers followed by simple agarose gel electrophoresis with bisbenzimide-PEG can be applied to the differentiation of three members of the family Chlamydiaceae which are commonly recognized as human pathogens. PMID- 10713424 TI - Constitutive expression of the UGA4 gene in Saccharomyces cerevisiae depends on two positive-acting proteins, Uga3p and Uga35p. AB - The first specific precursor of porphyrin biosynthesis is delta-aminolevulinic acid. delta-Aminolevulinic acid enters Saccharomyces cerevisiae cells through the gamma-aminobutyric acid specific permease Uga4p. It was described that this permease is inducible by gamma-aminobutyric acid and its regulation involves several specific and pleiotropic transcriptional factors. However, some studies showed that under certain growth conditions the synthesis of Uga4p was not dependent on the presence of gamma-aminobutyric acid. To study the effect of the trans-acting factors Uga43p, Uga3p, Uga35p, Ure2p and Gln3p on the expression of UGA4, we measured gamma-aminobutyric acid and delta-aminolevulinic acid uptake in yeast mutant cells, lacking one of these regulatory factors, grown under different conditions. Experiments analyzing the UGA4 promoter using a fusion construction UGA4::lacZ were also carried out. The results show that the constitutive expression of the UGA4 gene found in cells under certain growth conditions depends on the presence of Uga3p and Uga35p. In contrast, Gln3p and Ure2p do not seem to have any effect on this constitutive mechanism. PMID- 10713425 TI - Identification of iron-regulated genes of Helicobacter pylori by a modified fur titration assay (FURTA-Hp). AB - The Escherichia coli-based Fur titration assay (FURTA), although a powerful tool for identification of genes regulated by the ferric uptake regulator (Fur), was unsuccessful for the gastric pathogen Helicobacter pylori. The FURTA was modified by construction of an E. coli indicator strain producing H. pylori Fur only. The promoter regions of the ferric citrate receptor homolog fecA2 and the riboflavin synthesis gene ribBA were both positive in the modified FURTA, but negative in the original FURTA. Transcription of fecA2 and ribBA was demonstrated to be iron repressed in H. pylori. This type of modification should allow FURTA analysis for bacteria with Fur binding sequences poorly recognized by E. coli Fur. PMID- 10713426 TI - Evidence for the transport of zinc(II) ions via the pit inorganic phosphate transport system in Escherichia coli. AB - A locus involved in zinc(II) uptake in Escherichia coli K-12 was identified through the generation of a zinc(II)-resistant mutant by transposon (Tn10dCam) mutagenesis. The mutation was located within the pitA gene, which encodes the low affinity inorganic phosphate transport system (Pit). The pitA mutant accumulated reduced amounts of zinc(II) when exposed to 0.5-2.0 mM ZnSO(4) during growth in Luria-Bertani medium. PMID- 10713427 TI - The utilization of 4-aminobutylphosphonate as sole nitrogen source by a strain of Kluyveromyces fragilis. AB - A strain of the yeast Kluyveromyces fragilis was screened for its ability to utilize a range of synthetic and natural organophosphonate compounds as the sole source of phosphorus, nitrogen or carbon. Only 4-aminobutylphosphonate was utilized as sole nitrogen source with protein yields increasing proportionally with substrate concentrations up to 10 mM. No 4-aminobutylphosphonate metabolizing enzyme activity was detectable in cell-free extracts prepared from K. fragilis pregrown on 2.5 mM 4-aminobutylphosphonate. None of the organophosphonates tested served as a source of carbon or phosphorus for K. fragilis. PMID- 10713428 TI - Microwave oven and boiling waterbath extraction of hepatotoxins from cyanobacterial cells. AB - Low-cost, straightforward methods for the extraction of microcystins and nodularins from cyanobacterial cells were developed using a microwave oven and boiling waterbath. The use of organic solvents, such as methanol, which can interfere with sensitive analytical procedures, e.g. immunoassays, can thus be avoided. Analysis by protein phosphatase inhibition assay and high performance liquid chromatography indicated that purified microcystin-LR was unaffected by the microwave oven and boiling waterbath treatments. Four microcystins of differing hydrophobicities were successfully extracted from Microcystis PCC 7813 by both treatments at yields equivalent to those obtained by longer protocols using methanol. Assessment of the microwave oven and boiling waterbath extraction methods with laboratory strains and environmental samples of cyanobacteria showed good correlation with results from lyophilisation and methanol extraction, when extracts were analysed by high performance liquid chromatography with diode array detection (R(2)>/=0.92). The microwave and boiling waterbath extraction methods also sterilised the environmental bloom samples, as evidenced by the abolition of heterotrophic bacterial growth. PMID- 10713429 TI - Binding and utilization of myoglobin by Porphyromonas gingivalis. AB - Myoglobin was found to bind reversibly to the envelope of Porphyromonas gingivalis in a pH-dependent manner; the binding took place below neutral pHs of the incubation mixtures and myoglobin bound released from the envelope at high pHs. The amounts of myoglobin bound to 1 mg of the envelope at pH 5.0 per min under the presence of sufficient myoglobin were 1.4 microg. K(d) for the reaction at pH 5.0 was 2.2 x 10(-10) M. From the dot blot assay, myoglobin obviously bound to hemoglobin-binding protein (HbBP) of P. gingivalis, however, the amounts of myoglobin that bound to HbBP were half those of hemoglobin. One of the fractions, separated by gel filtration, of the digested materials of myoglobin by the detergent-solubilized envelope containing proteinases was found to support the growth of P. gingivalis in the iron source-depleted medium. PMID- 10713430 TI - The Pseudomonas aeruginosa phaG gene product is involved in the synthesis of polyhydroxyalkanoic acid consisting of medium-chain-length constituents from non related carbon sources. AB - We recently identified the phaG(Pp) gene encoding (R)-3-hydroxydecanoyl-ACP:CoA transacylase in Pseudomonas putida, which directly links the fatty acid de novo biosynthesis and polyhydroxyalkanoate (PHA) biosynthesis. An open reading frame (ORF) of which the deduced amino acid sequence shared about 57% identity with PhaG from P. putida was identified in the P. aeruginosa genome sequence. Its coding region (herein called phaG(Pa)) was amplified by PCR and cloned into the vector pBBR1MCS-2 under lac promoter control. The resulting plasmid pBHR88 mediated PHA synthesis contributing to about 13% of cellular dry weight from non related carbon sources in the phaG(Pp)-negative mutant P. putida PhaG(N)-21. The PHA was composed of 5 mol% 3-hydroxydodecanoate, 61 mol% 3-hydroxydecanoate, 29 mol% 3-hydroxyoctanoate and 5 mol% 3-hydroxyhexanoate. Furthermore, an isogenic phaG(Pa) knock-out mutant of P. aeruginosa was constructed by gene replacement. The phaG(Pa) mutant did not show any difference in growth rate, but PHA accumulation from gluconate was decreased to about 40% of wild-type level, whereas from fatty acids wild-type level PHA accumulation was obtained. These data suggested that PhaG from P. aeruginosa exhibits 3-hydroxyacyl-ACP:CoA transacylase activity and strongly enhances the metabolic flux from fatty acid de novo synthesis towards PHA(MCL) synthesis. Therefore, a function could be assigned to the ORF present in the P. aeruginosa genome, and a second PhaG is now known. PMID- 10713431 TI - A simple method to evaluate the concentration of pentachlorophenol degraders in contaminated soils. AB - A new most probable number (MPN) method for the determination of pentachlorophenol (PCP) degraders in soil using the change in pH due to PCP degradation is compared with a well documented MPN method using radiolabeled PCP. The results of all MPN counts were similar within a 95% confidence limit. The results obtained in MPN per gram of dry soil using pH measurements were 1.8 (+3.1, -1.03) x10 (4) compared to 0.64 (+1.34, -0.42) x 10(4) when using production of [(14)C]CO(2). PMID- 10713432 TI - Design and uses of Bdellovibrio 16S rRNA-targeted oligonucleotides. AB - An 18-mer oligonucleotide almost exclusively targeting Bdellovibrio spp. was designed based on available 16S rRNA sequence data. The specificity of this oligonucleotide used as a PCR primer in combination with a Bacteria domain targeted primer as well as used as a probe in rRNA dot blot hybridizations was experimentally confirmed using a variety of alpha-, beta-, gamma-, delta Proteobacteria and Gram-positive bacteria. Similarly, combinations of the Bacteria primer with oligonucleotides targeting the 16S rRNA gene of Bdellovibrio bacteriovorus and Bdellovibrio stolpii were shown to be species specific by PCR. Positive amplification products were obtained from an irrigation water sample in which a low level of bdellovibrios was detected by plating as well as from soil detached from potato tubers, using the Bdellovibrio spp.-Bacteria and the B. bacteriovorus-Bacteria primer pairs. PMID- 10713433 TI - Production of N-acyl-L-homoserine lactones by P. aeruginosa isolates from chronic lung infections associated with cystic fibrosis. AB - The N-acyl-L-homoserine lactones (AHLs) produced by sequential Pseudomonas aeruginosa isolates from chronically infected patients with cystic fibrosis were analyzed by thin-layer chromatography. It is demonstrated that both the amounts and the types of molecules synthesized by isolates from patients who were monitored over periods of up to 11 years do not change significantly during chronic colonization. However, in the case of a patient who became co-infected with an AHL-producing Burkholderia cepacia strain a dramatic reduction in the amounts of AHLs produced by the co-residing P. aeruginosa isolates was observed. PMID- 10713434 TI - Genomic diversity among Vibrio cholerae O139 strains isolated in Bangladesh and India between 1992 and 1998. AB - In order to assess the extent of genomic diversity among Vibrio cholerae O139 strains, restriction fragment length polymorphisms in two genetic loci, rrn and ctx, were studied. Analysis of 144 strains isolated from different regions of Bangladesh and India between 1992 and 1998 revealed the presence of at least six distinct ribotypes (B-I through B-VI) of which three were new ribotypes, and one of these was represented by a nontoxigenic O139 strain. Strains of ribotypes B-I through B-V shared 11 different CTX genotypes (A through K). Antimicrobial resistance patterns of the strains varied independently of their ribotypes and CTX genotypes. Results of this study suggest that V. cholerae O139 is undergoing rapid genetic changes leading to the origination of new variants, and temporal changes in antimicrobial resistance patterns may be contributing to the selection of different variants. PMID- 10713435 TI - Genetic analysis of SecA-SecY interaction required for spore development in Bacillus subtilis. AB - All spontaneous suppressor mutations obtained from a secA12 sporulation-defective mutant in Bacillus subtilis were localized in highly conserved membrane-spanning regions of SecY. The expression of early sporulation genes, kinA and spo0A encoding a histidine kinase and a transcription regulator for several sporulation genes, respectively, was restored in these suppressor mutants. These results indicate that the secretion function of translocase combined with Sec proteins is required for sporulation in B. subtilis. PMID- 10713436 TI - Flow cytometric detection of specific gene expression in prokaryotic cells using in situ RT-PCR. AB - Prokaryotic in situ RT-PCR was coupled with flow cytometry to detect mRNA transcripts of the toluene dioxygenase (todC1) gene in intact cells of the bacterium Pseudomonas putida F1. Recovery efficiency of fixed cells over the course of the entire in situ detection procedure was approximately 81% for both P. putida F1 and AC10R cells. It appeared that lysozyme treatment and PCR thermal cycling were the steps responsible for most of observed cell loss. Bacterial cells expressing the todC1 gene could be discriminated from negative control cells of the same size based on flow cytometrically-measured fluorescence and forward angle light scatter. According to flow cytometric analysis, the fluorescence intensity of positive cells was 4-5 times brighter than that of negative cells. The combination of flow cytometry and a prokaryotic in situ reverse transcription-PCR (RT-PCR) approach make possible the rapid detection and enumeration of functional (based on mRNA) populations of microbial cells. PMID- 10713437 TI - Plasmid curing effect of trovafloxacin. AB - The effect of sub-inhibitory concentrations of trovafloxacin, a recently developed fluoroquinolone molecule, on the capability of Escherichia coli cells to maintain three different types of plasmids has been investigated by a number of approaches, including the quantification of the loss of plasmid-borne functions and of plasmid DNA by quantitative PCR. The results obtained demonstrate that at concentrations ranging from the MIC to 1/8 of the MIC, trovafloxacin induces a clear, albeit incomplete, 'episome-curing' effect which was observed with plasmids differing in copy number, size and nature of the replication origin of the episome. This effect was most likely not due to an alteration of DNA supercoiling. PMID- 10713438 TI - Expression of beta-lactamase in Coxiella burnetii transformants. AB - Coxiella burnetii can be transformed to ampicillin resistance by electroporation with plasmids encoding beta-lactamase. However, non-plasmid emergence of resistance to ampicillin also develops. To validate the usefulness of the bla gene marker for selection and detection, transformed C. burnetii were examined for beta-lactamase expression by use of immunoblotting after SDS-PAGE. The 29-kDa mature form of the beta-lactamase protein was detected in C. burnetii lysates. Quantitation of these immunoblot signals showed that C. burnetii surprisingly expressed low levels of beta-lactamase. The results validate the use of plasmid encoded ampicillin resistance as a means for selecting C. burnetii transformants; they also suggest that levels of ampicillin used for selection pressure should be empirically determined and that detection of beta-lactamase by antibody blotting done to confirm transformants. PMID- 10713439 TI - The SRC family of nuclear receptor coactivators. AB - Nuclear hormone receptors are ligand-dependent transcription factors that regulate genes critical to such biological processes as development, reproduction, and homeostasis. Interestingly, these receptors can function as molecular switches, alternating between states of transcriptional repression and activation, depending on the absence or presence of cognate hormone, respectively. In the absence of hormone, several nuclear receptors actively repress transcription of target genes via interactions with the nuclear receptor corepressors SMRT and NCoR. Upon binding of hormone, these corepressors dissociate away from the DNA-bound receptor, which subsequently recruits a nuclear receptor coactivator (NCoA) complex. Prominent among these coactivators is the SRC (steroid receptor coactivator) family, which consists of SRC-1, TIF2/GRIP1, and RAC3/ACTR/pCIP/AIB-1. These cofactors interact with nuclear receptors in a ligand-dependent manner and enhance transcriptional activation by the receptor via histone acetylation/methylation and recruitment of additional cofactors such as CBP/p300. This review focuses on the mechanism of action of SRC coactivators in terms of interactions with receptors and activation of transcription. Specifically, the roles of the highly conserved LXXLL motifs in mediating SRC function will be detailed. Additionally, potential diversity among SRC family members, as well as several recently cloned SRC-associated cofactors, will be discussed. PMID- 10713440 TI - Initiation of genome replication: assembly and disassembly of replication competent chromatin. AB - Considerable progress has been made in research on the initiation of eukaryotic genome replication. This has generated a number of recent review articles. Here, we briefly summarize the major conclusions described in these articles and also include the results of more recent primary articles. The consensus view that has emerged is that a pre-replication complex assembles during the G1 phase of the cell cycle, making chromatin competent for replication. The complex consists of Orc proteins, Cdc6p, and the family of Mcm proteins. Chromatin, thus 'licenced' for replication, is guided into the S phase by the activation of cell-cycle regulated protein kinases. Upon entry into S phase, the pre-replication complex is partially dissolved, first by the dissociation of Cdc6p and then by the gradual release of Mcm proteins. This appears to be accompanied by a recruitment of chain elongation factors and the establishment of replication forks. PMID- 10713441 TI - Rice gibberellin-insensitive gene homolog, OsGAI, encodes a nuclear-localized protein capable of gene activation at transcriptional level. AB - This paper reports isolation and properties of a rice gene, OsGAI, a putative homolog of the GAI of Arabidopsis thaliana. OsGAI encodes a polypeptide of 625 amino acids, which shows 53-55% identity to GAI and RGA from A. thaliana, and 85% identity to wheat rht-D1a and maize d8. Genomic DNA blot analysis indicated the OsGAI to be a single-copy gene in the rice genome. RNA blot hybridization showed that OsGAI transcripts increased within 6h upon GA(3) but not ABA application. This GA-induced increment in OsGAI transcripts did not require de novo protein synthesis. High levels of OsGAI transcripts were detected in nodes, internodes, leaf sheaths and ears of adult plants and leaf sheaths of young seedlings, where GA enhances cell elongation and division. Transiently expressed OsGAI-GFP fusion protein located to the nucleus in onion epidermal cells. Transactivation assays clearly indicated that OsGAI protein is a transcriptional activator or a coactivator. PMID- 10713443 TI - Determination of heterogeneous transcription start points of two c-myc genes from the common carp (Cyprinus carpio). AB - We determined the heterogeneous transcription start points (tsp) of two c-myc genes from the common carp (Cyprinus carpio), tetraploid teleost, by the oligo capping method and showed the existence of the first exon. This is the first report on the existence of the first exons of the fish c-myc gene. Transcription of the two carp c-myc genes started from at least four sites in CAM1, locating from -752 to -381bp upstream of the translation start site, and from 12 sites in CAM2, locating from -586 to -413bp upstream respectively. The first introns of CAM1 and CAM2 were deduced to be 335 and 356bp, respectively. They shared 86.9% nt identity, lower than those of the second exons (94.1%), and third exons (92.3%), which suggest that the first exons evolved faster. No nt identities were found between the c-myc first exons of carp and other vertebrates. The putative promoter regions in CAM1 and CAM2 contained no obvious TATA or CCAAT boxes in the expected positions. PMID- 10713442 TI - A tyramine receptor gene mutation causes a defective olfactory behavior in Drosophila melanogaster. AB - We characterized molecular profiles of a new olfactory mutant line, honoka (hono), which was found among 500 viable P-element insertion lines screened first by 5-bromo-4-chloro-3-indrolyl-beta-D-galactopyranoside (X-gal) staining on the third segment of the antenna, and then by behavioral assays to several pure chemicals. The behavioral responses of hono mutants to repellents such as ethyl acetate (EA), benzaldehyde (BZ) and 4-methylcycrohexanol (MCH), were reduced compared with those of a control strain. The location of the P-element insertion was determined to be about 100bp) upstream of the first exon of the tyramine receptor gene. The level of 3.6kb tyramine receptor mRNA expression was reduced in hono compared with that of wild-type flies. The tyramine receptor cDNA hybridized to the chromosomal division 79C-D, the same locus as the P-element insertion point in hono, and not to 99A-B, previously reported by Arakawa et al. (1990. Neuron 2, 343-354). Electrophysiological responses to octopamine and tyramine were examined by measuring the excitatory junctional potential (EJP) amplitude from larval body-wall muscles of the hono mutant. The hono was impaired with responding to tyramine, while displaying normal response to octopamine. These results indicate that tyramine has a functional role in the Drosophila olfactory system as a neurotransmitter or a neuromodulator, and hono is the first tyramine receptor mutant. This study provides the first step toward understanding of the molecular genetics of tyramine-mediated neural functions in Drosophila. PMID- 10713444 TI - Cell-cycle regulation of the DNA topoisomerase IIalpha promoter is mediated by proximal CCAAT boxes: possible involvement of acetylation. AB - Expression of DNA topoisomerase (topo) IIalpha is cell-cycle-regulated, with its peak in G(2)/M and its lowest level in G(0)/G(1). In agreement with this expression pattern, we have shown that the topo IIalpha gene promoter shows cell cycle-dependent activity, which is repressed in G(0)/G(1) and activated exclusively in G(2)/M. However, the promoter sequence reveals no canonical CDE/CHR motifs, repressor elements commonly found in promoters of late S/G(2) activated genes. Here, we show that at least two of the three proximal inverted CCAAT boxes (ICBs) are responsible for the G(2)/M-specific activation of the topo IIalpha promoter. Using antibody supershift experiments, we identify NF-Y as the ICB-binding transcription factor. However, the expression profile and binding capacity of NF-Y were constant during the cell cycle, suggesting a more global mechanism in topo IIalpha promoter regulation. Interestingly, we find that trichostatin A (TSA), a specific histone deacetylase inhibitor, greatly enhances topo IIalpha promoter activity in an ICB-dependent manner. In addition, the effect of TSA is predominant in G(0)/G(1) and less obvious in G(2)/M. Our data, along with the recent findings that NF-Y associates in vivo with histone acetyltransferases (HATs), strongly suggest a mechanism, in which histone deacetylation plays a crucial role in the G(0)/G(1)-specific repression of the topo IIalpha promoter, and NF-Y recruits HATs to the promoter region, thereby stimulating histone acetylation and activating transcription in G(2)/M. PMID- 10713445 TI - A T-extended vector using a green fluorescent protein as an indicator. AB - T-extended vector (T-vector) is a useful tool for cloning PCR products directly. We exploited a novel T-vector using a green fluorescent protein (GFP) as an indicator based on insertional inactivation. The brightest GFP mutant was used for easy detection even under daylight. The 100bp and 0.9kb of PCR products were cloned, and the transformant colonies with inserts were adjudged by the fluorescent green-white screening. The GFP system was more sensitive to insertional inactivation than the beta-galactosidase system at the conventional insertion sites. PMID- 10713446 TI - Complete nucleotide sequence and evolutionary significance of a chromosomally encoded naphthalene-degradation lower pathway from Pseudomonas stutzeri AN10. AB - Pseudomonas stutzeri strain AN10 is a naphthalene-degrading strain whose dissimilatory genes are chromosomally encoded. We sequenced the entire naphthalene-degradation lower pathway of P. stutzeri AN10, this being, together with the upper-pathway reported previously (Bosch R. et al., 1999a. Gene 236, 149 157) the first complete DNA sequence for an entire naphthalene-catabolic pathway. Eleven open reading frames were identified. The nahGTHINLOMKJ genes encode enzymes for the metabolism of salicylate to pyruvate and acetyl-CoA, and nahR encodes the NahR regulatory protein. Our findings suggest that catabolic modules were recruited through transposition events and recombination among tnpA-like genes, and subsequent rearrangements and deletions of non-essential DNA fragments allowed the formation of the actual catabolic pathway. Our results also suggest that the genes encoding the xylene/toluene-degradation enzymes of P. putida mt-2 (pWW0) have coexisted with the nah genes of the P. stutzeri AN10 ancestral genome. This could allow the selection, via recombination events among homologous genes, for a combination of genes enabling the metabolism of a given aromatic compound in the ancestral host strain. Such events accelerate the evolution of modern catabolic pathways and provide new genetic material to the environment, ultimately resulting in improved, natural, bioremediation potential. PMID- 10713447 TI - Cloning and characterization of cDNA encoding zebrafish Danio rerio NM23-B gene. AB - A full-length zebrafish NM23-B cDNA was cloned and sequenced. The zebrafish NM23 B cDNA consists of 624bp with an open reading frame of 153 amino acids. NM23-B mRNA of approximately 0.7kb is present in adult zebrafish tissues. Zebrafish NM23 B his-tagged protein (17kDa) was produced in E. coli and characterized by binding and UV-cross-linking to a single-stranded telomeric repeat (TTAGGG)(6). This is the first report to show that fish have a NM23-H2 homologue that is similar to that in humans. PMID- 10713448 TI - Cloning and primary structure of putative cytosolic and mitochondrial malate dehydrogenase from the mollusc Nucella lapillus (L.). AB - The evolutionary history of the malate dehydrogenase (MDH) gene family [NAD dependent MDH; EC 1.1.1.37 and NAD(P)-dependent MDH; EC 1.1.1.82] has received much attention. MDHs have also featured extensively as electrophoretic markers in population genetics and evolutionary ecology, and in many cases, intraspecific variation in MDH has been correlated with environmental variables. However, while the amino acid residues essential for MDH function are known, no studies have examined intraspecific nucleotide variation despite evidence indicating that natural selection may be operating on this locus. This study presents two sets of degenerate oligonucleotide PCR primers to facilitate the cloning of cytosolic MDH (cMDH) and mitochondrial MDH (mMDH) from a broad range of animals (cMDH) and animals and plants (mMDH). These primers were used to obtain putative cMDH and mMDH cDNAs from the mollusc Nucella lapillus. The N. lapillus cMDH cDNA was found to encode a putative cMDH protein of 334aa and 36kDa, while the mMDH cDNA encoded a putative mature mMDH protein of 315aa and 33kDa. The putative amino acid sequences of the two compartmentalised N. lapillus MDHs are presented and compared to other known MDH sequences. PMID- 10713449 TI - Characterization of two members of the maize gene family, Incw3 and Incw4, encoding cell-wall invertases. AB - Two maize putative cell-wall invertase genes (Incw3 and Incw4) have been isolated by screening a genomic DNA library (Zea mays L. W22) using the cDNA probes encoding the two maize cell-wall invertases Incw1 and Incw2. The Incw3 and Incw4 genes contain six exons/five introns and five exons/four introns, respectively. The protein sequences deduced from both genes revealed a beta-fructosidase motif and a cysteine catalytic site known to be conserved in invertase genes. A detailed analysis of the protein and nucleotide sequences provides evidence that the Incw3 and the Incw4 genes encode putative cell-wall invertases. Furthermore, the isoelectric point deduced from the INCW4 protein sequence suggested that the Incw4 gene may encode a unique type of cell-wall invertase unbound in the apoplast. Gene expression studies using RT-PCR and in-situ RT-PCR hybridization showed that the Incw3 expression is organ/tissue-specific and developmentally regulated. In contrast, the Incw4 gene is constitutively expressed in all vegetative and reproductive tissues tested. PMID- 10713450 TI - Molecular cloning, chromosome mapping and characterization of a testis-specific cystatin-like cDNA, cystatin T. AB - The cystatin superfamily of cysteine proteinase inhibitors consists of three major families. In the present study, we report the cloning of the cDNA for mouse cystatin T, which is related to family 2 cystatins. The deduced amino acid sequence of cystatin T contains regions of significant sequence homology including the four highly conserved cysteine residues in exact alignment with all cystatin family 2 members. However, cystatin T lacks some of the conserved motifs believed to be important for inhibition of cysteine proteinase activity. These characteristics are seen in two other recently cloned genes, CRES and Testatin. Thus, cystatin T appears to be the third member of the CRES/Testatin subgroup of family 2 cystatins. The mouse cystatin T gene was mapped on a region of chromosome 2 that contains a cluster of cystatin genes, including cystatin C and CRES. Northern blot analysis demonstrated that expression of mouse cystatin T is highly restricted to the mouse testis. Thus, a shared characteristic of the cystatin family 2 subgroup members is an expression pattern limited primarily to the male reproductive tract. PMID- 10713451 TI - Rad25p, a DNA helicase subunit of yeast transcription factor TFIIH, is required for promoter escape in vivo. AB - The general transcription factor TFIIH is required for initial DNA unwinding and promoter escape by RNA polymerase II in vitro. We examined whether Rad25p, a DNA helicase subunit of TFIIH, mediates promoter opening and promoter escape in the yeast Saccharomyces cerevisiae. DNA unwinding was probed with an in vivo permanganate reactivity assay, in a temperature-sensitive mutant of RAD25. The consequences of Rad25p inactivation were promoter-specific. Whereas in the TDH2 promoter permanganate reactivity was entirely abolished, the reactivity at the GAL1 and GAL10 promoter regions was only moderately affected. In the GAL genes permanganate reactivity uniformly decreased downstream of the transcription start site, indicating that progression of RNA polymerase II to this region was impaired. Our results suggest that in yeast cells, promoter opening is not sufficient for productive initiation and that Rad25p-mediated promoter escape may be a limiting step in the transcription of some promoters. PMID- 10713452 TI - Isolation and analysis of two cellulase cDNAs from Orpinomyces joyonii. AB - Two cellulase cDNAs, celB29 and celB2, were isolated from a cDNA library derived from mRNA extracted from the anaerobic fungus, Orpinomyces joyonii strain SG4. The nucleotide sequences of celB2 and celB29 and the primary structures of the proteins encoded by these cDNAs were determined. The larger celB29 cDNA was 1966bp long and encoded a 477 amino acid polypeptide with a molecular weight of 54kDa. Analysis of the 1451bp celB2 cDNA revealed an 1164bp open reading frame coding for a 44kDa protein consisting of 388 amino acids. Both deduced proteins had a high sequence similarity in central regions containing putative catalytic domains. Primary structure analysis revealed that CelB29 contained a Thr/Pro-rich sequence that separated the N-terminal catalytic domain from a C-terminal reiterated region of unknown function. Homology analysis showed that both enzymes belong to glycosyl hydrolase family 5 and were most closely related to endoglucanases from the anaerobic fungi Neocallimastic patriciarum, Neocallimastix frontalis and Orpinomyces sp. The classification of CelB29 and CelB2 as endoglucanases was supported by enzyme assays. The cloned enzymes had high activities towards barley beta-glucan, lichenan and carboxymethylcellulose (CMC), but not Avicel, laminarin, pachyman, xylan and pullulan. In addition, CelB29 and CelB2 showed activity against p-nitrophenyl-beta-D-cellobioside (pNP G(2)) to p-nitrophenyl-beta-D-cellopentaoside (pNP-G(5)) but not p-nitrophenyl beta-D-glucopyranoside (pNP-G(1)) with preferential activity against p nitrophenyl-beta-D-cellotrioside (pNP-G(3)). Based on these results, we proposed that CelB29 and CelB2 are endoglucanases with broad substrate specificities for short- and long-chain beta-1,4-glucans. PMID- 10713453 TI - The protein Hrb57A of Drosophila melanogaster closely related to hnRNP K from vertebrates is present at sites active in transcription and coprecipitates with four RNA-binding proteins. AB - The hnRNP K protein is among the major hnRNA-binding proteins with a strong preference for cytidine-rich sequences. We have cloned a Drosophila hnRNP protein closely related to this vertebrate protein. The protein first identified by the monoclonal antibody Q18 is encoded by a gene located in 57A on polytene chromosomes and has been consequently named Hrb57A. The amino acid sequence of the Hrb57A KH domains and their overall organisation in the protein are remarkably similar to the vertebrate proteins. As the hnRNP K in vertebrates the M(r) 55 000 Drosophila Hrb57A/Q18 protein strongly binds to poly(C) in vitro and is ubiquitously present in nuclei active in transcription. On polytene chromosomes it is found in many puffs and minipuffs. Hrb57A/Q18 specifically coprecipitates four other proteins: Hrb87F/P11 a Drosophila hnRNP A1 homologue, the hnRNA-binding protein S5, the RNA recognition motif-containing protein NonA and the RNA-binding zinc finger-containing protein on ecdysone puffs PEP/X4. PMID- 10713454 TI - Major transcript variants of VAV3, a new member of the VAV family of guanine nucleotide exchange factors. AB - VAV3 is a new member of the VAV oncogene family with a strong homology to VAV and VAV2. A conceptual translation of the cDNA indicates that VAV3 is between 40 and 77% identical to VAV and VAV2 at the amino acid level in all identified functional motifs. This homology suggests that VAV3 occupies a similar position in signal transduction as the other family members. A major variant transcript, VAV3.1, found in both humans and mice, appears to encode only the 3' SH3-SH2-SH3 region, which suggests that it may substitute for the full-length isoform in functions mediated by this domain, or compete with the full-length isoform in functions mediated by more N-terminal motifs. VAV3.1 either is a partly unspliced mRNA or originates from a different promoter. VAV3 transcripts are found in cells of hematopoietic origin, where VAV is primarily expressed. However, unlike, VAV, the VAV3 and VAV3.1 transcripts are also found at varying levels in a wide variety of other tissues and cell lines. TGF-beta and EGF reversibly down regulate the abundance of the VAV3. 1 transcript in HaCaT keratinocytes, representing the first observation of transcript regulation of a member of the VAV family by a growth factor. PMID- 10713455 TI - Studies of the murine DDB1 and DDB2 genes. AB - Human DDB (Damaged DNA Binding protein) is a heterodimer of 48 and 127kDa subunits whose activity is absent from cell strains derived from a subset of Xeroderma Pigmentosum (XP) complementation group E individuals (Ddb(-)) [Keeney, S., Wein, H., and Linn, S., (1992). Mut. Res. 273, 49-56]. Whereas in vivo DNA repair appears to be compromised in both Ddb(-) and Ddb(+) XPE cells, DDB activity is not necessary for nucleotide excision repair (NER) in vitro. In this study, the presence of a specific UV-damaged DNA binding activity in mouse cell free extracts that is comparable to the activity observed in HeLa cells was demonstrated. The mouse DDB2 cDNA, coding for DDB p48 subunit, was cloned and the partial genomic structure of DDB2 was obtained. A search of current databases revealed amino acid sequences of mouse and Drosophila predicted p127 homologues, but not of a Drosophila p48 homologue. The alignment of these higher eukaryotic p127 sequences uncovered the presence of three highly conserved domains in the p127 polypeptides which we hypothesize could function in DNA binding, transcription-transactivation, and protein-protein interaction, respectively. PMID- 10713457 TI - Rapid, high-level expression of glycosylated rice alpha-amylase in transfected plants by an RNA viral vector. AB - Tobamoviral vectors have been developed for the heterologous expression of glycoproteins in plants. The rice alpha-amylase gene (OS103) was placed under the transcriptional control of a tobamovirus subgenomic promoter in a RNA viral vector. One to two weeks after inoculation, transfected Nicotiana benthamiana plants accumulated glycosylated alpha-amylase to levels of at least 5% total soluble protein. The 46kDa recombinant enzyme was purified, and its structural and biological properties were analyzed. Post-translational modifications of the secreted protein were compared to rice alpha-amylase expressed in amylolytic strains of Pichia pastoris and Saccharomyces cerevisiae. Endo-H analysis revealed that the alpha-amylase was moderately glycosylated in transfected plants and hyperglycosylated in yeast. PMID- 10713456 TI - TfdR, the LysR-type transcriptional activator, is responsible for the activation of the tfdCB operon of Pseudomonas putida 2, 4-dichlorophenoxyacetic acid degradative plasmid pEST4011. AB - In Pseudomonas putida EST4021, the tfdCB operon of plasmid pEST4011 encodes enzymes involved in 2,4-dichlorophenoxyacetic acid degradation. We have identified a gene, tfdR, important for the regulation of the tfdCB operon. Sequence analysis of the tfdR gene revealed an open reading frame with amino acid sequence similar to the LysR family of transcriptional activators. The tfdR gene is located upstream and transcribed divergently from the tfdCB operon. Utilizing primer extension analysis, the transcription initiation sites of the gene tfdR and the tfdCB operon were localized 85 (84)bp and 292bp upstream from the coding sequences of these genes, respectively. Multiple sequence analysis revealed that the genes tfdR, tfdC and tfdB of plasmid pEST4011 are most similar to the regulatory gene tfdR and the module 2 genes tfdC(II) and tfdB(II) of pJP4, respectively. The promoter-operator sequences of tfdR and its target tfdCB operon of pEST4011 have regions with highly conserved nucleotides characteristic for the catechol-subgroup LysR-type transcriptional activators. We showed that the pEST4011 tfdR gene product activates the expression of the tfdCB operon and the effector molecule for TfdR is 2,4-dichloro-cis,cis-muconate. Our data indicate that the structure and the mode of regulation of tfd genes are similar, despite the bacteria being isolated from different geographical regions. PMID- 10713458 TI - Mutational analysis of the RNA component of Saccharomyces cerevisiae RNase MRP reveals distinct nuclear phenotypes. AB - The 340-nucleotide RNA component of Saccharomyces cerevisiae RNase MRP is encoded by the single-copy essential gene, NME1. To gain additional insight into the proposed structure and functions of this endoribonuclease, we have extensively mutagenized the NME1 gene and characterized yeast strains expressing mutated forms of the RNA using a gene shuffle technique. Strains expressing each of 26 independent mutations in the RNase MRP RNA gene were characterized for their ability to grow at various temperatures and on various carbon sources, stability of the RNase MRP RNA and processing of the 5.8S rRNA (a nuclear function of RNase MRP). 11 of the mutations resulted in a lethal phenotype, six displayed temperature-conditional lethality, and several preferred a non-fermentable carbon source for growth. In those mutants that exhibited altered growth phenotypes, the severity of the growth defect was directly proportional to the severity of the 5.8S rRNA processing defect in the nucleus. Together this analysis has defined essential regions of the RNase MRP RNA and provides evidence that is consistent with the proposed function of the RNase MRP enzyme. PMID- 10713459 TI - Characterization and chromosomal mapping of a human Necdin pseudogene. AB - The necdin gene is expressed predominantly in postmitotic neurons and encodes a growth suppressor that interacts with the transcription factors E2F1 and p53. Human necdin gene (NDN) is maternally imprinted and located in Prader-Willi syndrome deletion region 15q11.2-q12. We isolated an NDN homologous sequence from a human genomic DNA library. The homologous sequence is overall 83% identical with necdin cDNA sequence, and possesses a short poly(A) stretch at the 3' end and direct repeats at both ends. Expression of the homologous sequence, which lacks a 5' promoter sequence, was undetected in cultured human cell lines. We mapped this sequence to chromosome 12q14-q21.1 by fluorescence in situ hybridization. These characteristics of the NDN-homologous sequence are consistent with those of processed pseudogenes. The information about the necdin pseudogene in the human genome will be useful for genetic studies on NDN associated neurogenic disorders. PMID- 10713460 TI - Nucleotide sequence and differential expression of the human 3-phosphoglycerate dehydrogenase gene. AB - The nucleotide sequence of Hs 3-PGDH gene, encoding human 3-phosphoglycerate dehydrogenase that catalyzes the initiating step in the phosphorylated pathway of serine biosynthesis, has been determined. The 3-PGDH gene has a predicted 533 amino acid open reading frame, encoding a 56.8kDa protein that shares 94.0% similarity with rat-liver 3-PGDH. Two different transcripts corresponding to 3 PGDH mRNA were detected in human normal tissues. A dominant 2.1kb transcript was expressed at high levels in prostate, testis, ovary, brain, liver, kidney, and pancreas, and weakly expressed in thymus, colon, and heart. A 710bp transcript also appeared as a weaker band where the 2.1kb mRNA was expressed, and it was more significant than the 2.1kb mRNA in heart and skeletal muscle. The TPA induced monocytic differentiation of U937, which also resulted in growth arrest, abruptly downregulated the expression of 3-PGDH. Removal of TPA restored cell growth through the retrodifferentiation process and subsequent expression of 3 PGDH. The 3-PGDH mRNA was markedly expressed in human leukemias, lymphoma Sup-T1, colon adenocarcinoma COLO 320DM, epitheloid carcinoma HeLa S3, and murine lymphoma BW5147.G.1.4, but not in human leukemia K562. This report demonstrates that the human 3-PGDH gene is regulated at the transcriptional level depending on tissue specificty and cellular proliferative status, and its transcriptional regulation mechanism may be a useful target for diagnosis and therapy of cancer. PMID- 10713461 TI - Genetic architecture of the polyketide synthases for methymycin and pikromycin series macrolides. AB - The methymycin and pikromycin series of antibiotics are structurally related macrolides produced by several Streptomyces species, including Streptomyces venezuelae ATCC 15439, which produces both 12-membered ring macrolides methymycin, neomethymycin, and 14-membered ring macrolides pikromycin and narbomycin. Cloning and sequencing of the biosynthetic gene clusters for these macrolides from three selected Streptomyces strains revealed a common genetic architecture of their polyketide synthases (PKSs). Unlike PKS clusters of other 14-membered ring macrolides such as erythromycin and oleandomycin, each of the pikromycin series producers harbors a six module PKS cluster, in which modules 5 and 6 are encoded on two separate proteins instead of one bimodular protein, as well as a thioesterase II gene immediately downstream of the main PKS gene. The results shed new light on the evolution of modular PKSs and provide further evidence on the regulation of methymycin and pikromycin production in S. venezuelae ATCC 15439. PMID- 10713462 TI - A novel type of RNase III family proteins in eukaryotes. AB - The RNase III family of double-stranded RNA-specific endonucleases is characterized by the presence of a highly conserved 9 amino acid stretch in their catalytic center known as the RNase III signature motif. We isolated the drosha gene, a new member of this family in Drosophila melanogaster. Characterization of this gene revealed the presence of two RNase III signature motifs in its sequence that may indicate that it is capable of forming an active catalytic center as a monomer. The drosha protein also contains an 825 amino acid N-terminus with an unknown function. A search for the known homologues of the drosha protein revealed that it has a similarity to two adjacent annotated genes identified during C. elegans genome sequencing. Analysis of the genomic region of these genes by the Fgenesh program and sequencing of the EST cDNA clone derived from it revealed that this region encodes only one gene. This newly identified gene in nematode genome shares a high similarity to Drosophila drosha throughout its entire protein sequence. A potential drosha homologue is also found among the deposited human cDNA sequences. A comparison of these drosha proteins to other members of the RNase III family indicates that they form a new group of proteins within this family. PMID- 10713463 TI - Production of reactive oxygen species by phagocytic cells after exposure to glass wool and stone wool fibres - effect of fibre preincubation in aqueous solution. AB - The potential of four man-made vitreous fibres (MMVFs) (glass wool Code A, stone wool Code G, HT-N and MMVF 21) and of two natural mineral fibres (crocidolite, erionite) to induce production of reactive oxygen species (ROS) by differentiated HL-60 cells (HL-60-M cells) was investigated by determination of luminol-enhanced chemiluminescence (CL). Quartz served as positive control. The same system was used to uncover possible influences of fibre preincubation in aqueous solutions on the ROS-generating potential. Following preincubation in unbuffered saline over about 4 weeks, Code A and G fibres showed decreased ROS-generating potential as compared to freshly suspended fibres. On the other hand, MMVF 21 and HT-N fibres as well as crocidolite and erionite showed no decreased CL after incubation in aqueous solutions. The observed decrease of the ROS-generating potential of Code A and G fibres after preincubation may be an expression of fibre surface alterations (leaching, initiation of dissolution) that influences the response of exposed phagocytic cells. After incubation of both fibres in buffered solutions at different pH values (5.0, 7.4) a reduced ROS-generating potential was still discernible as compared to freshly suspended fibres. PMID- 10713464 TI - Chronic toxic effects of quinalphos on some biochemical parameters in Labeo rohita (Ham.). AB - The effect of exposure to sublethal concentrations of the organophosphate pesticide, quinalphos (1.12, 0.22 mg/l) on biochemical parameters of muscle and enzyme activities in brain, liver and kidney of the Indian major carp, Labeo rohita was studied after 15, 30 and 45 days. The muscle protein and RNA levels decreased whereas DNA levels and acid phosphatase were elevated. Similarly, alkaline phosphatase was depleted. The brain acetyl cholinesterase activity was decreased most (-75.43%) in 1.12 mg/l concentration over a period of 45 days. Lactic dehydrogenase levels in brain and liver were elevated whereas in the kidney they were inhibited. Succinic dehydrogenase and adenosine triphosphatase activities were depleted in brain, liver and kidney. The effects have been discussed for different organ tissues in relation to the pesticide. PMID- 10713465 TI - Diminished decondensation and DNA synthesis in activated sperm from rats treated with cyclophosphamide. AB - Standard andrology tests do not predict fertility or assess the genetic quality of spermatozoa. To address these problems, we have analyzed sperm nuclear activation in vitro using cytoplasmic extracts of Xenopus laevis frog eggs. The objective of this study was to determine if rat sperm chemically damaged in vivo by cyclophosphamide treatment would respond abnormally in an in vitro rat sperm activation assay (RSAA). Male Sprague-Dawley rats were treated for 6 weeks with cyclophosphamide (CP) to induce DNA damage in post-meiotic germ cells. After the treatment period, cauda epididymal sperm were isolated, and incubated in cytoplasmic extracts of X. laevis frog eggs to induce chromatin decondensation and DNA synthesis in vitro. Sperm from treated rats displayed significant decreases in both decondensation and DNA synthesis when compared to sperm from control rats, consistent with the presence of CP-induced DNA crosslinks. No differences in body, testes, or epididymal weights were observed between control and treated rats, nor was sperm count diminished in the treatment group. These results demonstrate that the RSAA can be used to detect damaged sperm chromatin in the absence of detrimental effects on sperm count, and testis and epididymal weights. PMID- 10713466 TI - Effects of neutral ionophores on membrane electrical characteristics of NG108-15 cells. AB - The effects of several K(+)-selective neutral ionophores on membrane electrical characteristics of differentiated NG108-15 (neuroblastoma X glioma hybrid) cells were examined. Specifically, alterations in membrane resting potential (V(m)), input resistance (R(in)) and electrically-induced action potential generation were determined upon bath application of enniatin (0.1-10 microg/ml), nonactin (0. 1-10 microM) and valinomycin (0.1-10 microM). Although some cells exhibited a slight hyperpolarization and/or reduced R(in), i.e. membrane electrical correlates of enhanced K(+) loss, neither V(m) nor R(in) were significantly altered by any of the ionophores. However, valinomycin and especially nonactin affected action potentials induced by electrical stimulation. This was apparent in the ablation of action potentials in some cells and in the occurrence of degenerative changes in action potential shape in others. The simultaneous administration of the neutral ionophores and the protonophore CCCP or the superfusion of enniatin, nonactin or valinomycin in high (50 mM) glucose containing physiological solution did not yield more extensive alterations in V(m) or R(in). These data suggest that the neutral ionophores are unable to materially enhance K(+) flux above the relatively high resting level in NG108-15 cells. Thus, alterations in action potentials appear to be unrelated to K(+) transport activity. PMID- 10713467 TI - Effect of vitamin E on lipid peroxidation and liver monooxigenase activity in experimental influenza virus infection. AB - Influenza virus infection was associated with development of oxidative stress in liver of mice, viz. increase in amount of lipid peroxidation products, decrease in cytochrome P-450 and NADP. H-cytochrome c-reductase activity, and inhibition of liver monooxygenases (aniline hydroxylase, ethylmorphine-N-demethylase, amidopyrine-N-demethylase and analgin-N-demethylase). These effects were most pronounced on the 7th day after virus inoculation as compared to the 5th one. Supplementation of mice with vitamin E before virus inoculation leads to liver protection against oxidative stress and toxicosis. A marked decrease of lipid peroxidation products and an increase of cytochrome P-450 and activities of monooxygenases was established. The stabilizing effect of vitamin E was dose dependent and was most pronounced on the 5th day after virus inoculation as compared to the 7th one. PMID- 10713468 TI - Clinical and pharmacological profile in a clenbuterol epidemic poisoning of contaminated beef meat in Italy. AB - Long-acting beta adrenergic agonists, such as clenbuterol accumulate in the liver, but not meat of treated farm animals, and result in epidemic poisonings in consumers. We describe an outbreak of poisoning in 15 people, following the consumption of meat. Clinical symptoms (distal tremors, palpitations, headache, tachipnoea-dyspnoea, and also moderate hyperglycaemia, hypokalemia and leucocytosis) were seen in nine hospitalised patients, starting about 0.5-3 h after poisoning, and disappearing within 3-5 days later. Clenbuterol was found in the urine of all the symptomatic patients, at higher levels than pharmacokinetic computing (mean level 28 ng/ml, 36 h after ingestion), based on the levels found in the meat (1140-1480 ng/g edible tissue). Thus, epidemic poisoning can be produced following the consumption of contaminated meat. The need for a better definition of pharmaco- and toxico-kinetics, not only for drugs ingested as parent drug, but also when ingested as residues with animal tissues, is recommended. PMID- 10713469 TI - Use of a combined human liver microsome-estrogen receptor binding assay to assess potential estrogen modulating activity of PCB metabolites. AB - Polychlorinated biphenyls (PCBs) are metabolized by hydroxylation; some of these hydroxylated metabolites exhibit estrogen-like activity in animal models. Because PCBs may have effects on human health, it is of interest to determine if human tissues also metabolize PCBs to potentially estrogenic metabolites. In this study metabolites of seven PCBs with different degrees and positions of chlorination, generated by human liver microsomal reaction mixtures (MRM) have been examined, and their affinity for human recombinant estrogen receptor-alpha (ER) has been tested before and after HPLC fractionation. Two of the three MRMs with di-chloro biphenyls (BPs, 2,5BP and 3,4BP), one of the three MRMs with tetra-BPs (2,6,2',6'BP), and one hexa-BP (2,4,6,2',4',6'BP) generated metabolites that competed for ER. HPLC of the ER-binding MRMs generated fractions that also exhibited ER-binding. This study shows the usefulness of combining in vitro metabolism and an ER-binding assay in initial identification of PCBs with estrogen-modulating potential. PMID- 10713470 TI - Potential mechanisms of tumorigenic action of diethanolamine in mice. AB - Diethanolamine (DEA), a secondary amine found in a number of consumer products, reportedly induces liver tumors in mice. In an attempt to define the tumorigenic mechanism of DEA, N-nitrosodiethanolamine (NDELA) formation in vivo and development of choline deficiency were examined in mice. DEA was administered with or without supplemental sodium nitrite to B6C3F1 mice via dermal application (with or without access to the application site) or via oral gavage for 2 weeks. Blood levels of DEA reflected the dosing method used; oral greater than dermal with access greater than dermal without access. No NDELA was observed in the urine, blood or gastric contents of any group of treated mice. Choline, phosphocholine and glycerophosphocholine were decreased 20 kHz. Middle-ear motion (i.e. umbo velocity) was similar in the two strains; although frequencies >20 kHz could not be evaluated. Permanent threshold shift (PTS) and hair cell losses, measured 1 week after high-intensity exposure to an 8-16 kHz noise band, were smaller in129/SvEv at all exposure levels and durations from 97 dB SPLx2 h to 106 dB SPLx8 h. Furthermore, PTS growth with increasing exposure energy was slower in 129/SvEv (<2 dB/dB) than CBA/CaJ (9 dB/dB). These data suggest that the vulnerability differences lie in the inner ear, not the middle ear. Several 129/Sv substrains show age-related hearing loss (AHL): 129/SvEv has not yet been evaluated (Zheng, Q.Y., Johnson, K. R., Erway, L.C., 1999. Assessment of hearing in 80 inbred strains of mice by ABR threshold analyses. Hear. Res. 130, 94-107). Thus, although other strains with AHL, e.g. C57Bl/6J, show increased vulnerability to noise-induced hearing loss (NIHL), pairing of AHL and NIHL vulnerabilities may not be obligatory. PMID- 10713499 TI - Regional variations of noise-induced changes in operating range in cat AI. AB - Regional differences in spectral integration of neurons in cat primary auditory cortex (AI) suggest that regions differ in effects of background noise on operating characteristics of neurons. Therefore, tone-response threshold, best level (peak-rate intensity), dynamic range, and sharpness of tuning in quiet and in continuous broadband noise were mapped for single neurons along the isofrequency domain of AI. Neurons did not show an excitatory response to the noise. Noise invariably increased the tone-response threshold and best levels. Consequently, the dynamic ranges and receptive fields shifted to higher intensity levels without changes of average sharpness of tuning. These shifts were linearly related to noise level and showed little inter-neuronal variability for neurons in the central, mostly sharply tuned part of AI. In more dorsal and ventral parts of AI, neurons were more variable in tone-response threshold, dynamic range and best level, and no systematic relationship between increase in noise level, threshold increase and best-level increase was observed. We conclude that linear shifts in the operating range of neurons in central AI in the presence of continuous noise backgrounds do not affect other response properties and may relate to the unaltered analysis and representation of spectral components of sounds. In contrast, neurons in dorsal and ventral AI change response properties in a non-predictable way in the presence of noise in accordance with the more complex receptive field properties in those areas. PMID- 10713501 TI - Structural/audiometric correlations in a human inner ear with noise-induced hearing loss. AB - A morphological analysis was performed on a human cochlea removed during skull base surgery. The patient experienced a noise-induced hearing loss following 30 years of mechanical exposure. The tissue was processed according to the block surface technique and the organ of Corti, osseous spiral lamina and spiral ganglion were analyzed at different levels. There was a circumscribed lesion approx. 10 mm from the round window extending to about 13 mm. At this site, the dominant pathological feature was the loss of outer hair cells that was comprehensive in the centermost area and partial in the peripheral region of the damage. The degradation of inner hair cells was less severe with signs of cell atrophy yet with limited loss. Outer pillar cells were often collapsed leading to deformation of the acoustic ridge. The Deiters cells were often present and physically interactive with remaining nerve fibers. In the reticular lamina, surgical manipulation and dissection resulted in tears which may be attributed to a reduction of intercellular strength between cells. In the damaged area, there was a 45% loss of myelinated nerve fibers measured at the osseous spiral lamina. Pathological changes could not be observed in the spiral ganglion with certainty although the type II cells innervating the outer hair cells were often difficult to discern. PMID- 10713500 TI - High calcium permeability and calcium block of the alpha9 nicotinic acetylcholine receptor. AB - At the synapse between olivocochlear efferent fibers and outer hair cells (OHCs) of the cochlea, a non-classical ionotropic cholinergic receptor allows Ca(2+) entry into the hair cell, thus activating a Ca(2+)-sensitive K(+) current which hyperpolarizes the cell's membrane. In the mammalian ear, this leads to a reduction in basilar membrane motion, altering auditory nerve fiber activity and reducing the dynamic range of hearing. The alpha9 nicotinic acetylcholine receptor (nAChR) subunit mediates synaptic transmission between cholinergic olivocochlear fibers and OHCs. Given that Ca(2+) is a key player at this inhibitory synapse, we evaluated the permeability to Ca(2+) of the recombinant alpha9 receptor expressed in Xenopus laevis oocytes and the modulation of its activity by extracellular Ca(2+). Our results show that the alpha9 receptor is highly permeable to Ca(2+) and that this cation potently blocks monovalent currents through this channel (IC(50)=100 microM, at -70 mV) in a voltage dependent manner. At a Ca(2+) concentration similar to that found in the perilymph bathing the base of the OHCs, approximately 90% of the Na(+) current through the alpha9 receptor is blocked, suggesting that one of the main functions of this channel could be to provide a pathway for Ca(2+) influx. PMID- 10713502 TI - Detection of small across-channel timing differences by cochlear implantees. AB - Five post-lingually deafened users of the LAURA cochlear implant were presented with two trains of biphasic pulses applied concurrently to two widely separated channels. They could all discriminate between stimuli where pulses on the two channels were nearly synchronous (inter-channel delay=0.1 ms) and those where there was a longer delay applied to one channel. All showed an asymmetry, being more sensitive when the longer delay was on either the more basal or, depending on the listener, the more apical channel. For four out of the five listeners this asymmetry could be at least partly attributed to one stimulus, with a 0.1-ms delay in either the apical (three listeners) or basal (one listener) channel, sounding markedly different from all other stimuli used in the experiment. Both the overall sensitivity of listeners and the general pattern of results survived the presentation of maskers on intermediate channels, and did not vary markedly with changes in the polarity of the pulses applied to one channel. Although the results varied substantially across listeners, it is concluded that they demonstrate a genuine sensitivity to the relative timing of stimulation applied to discrete populations of auditory nerve fibers. PMID- 10713503 TI - Effect of violation of the labyrinth on the sensory epithelium in the chick cochlea. AB - Models in which a single large systemic dose of gentamicin is used to cause near synchronous hair cell (HC) loss in the basal end of the chick cochlea have proven increasingly useful in the study of HC regeneration. We quantified the amount of HC death, as a percentage of the length of the basilar papilla, following single doses of 200 mg/kg and 300 mg/kg of gentamicin in 23-day-old chicks. Following 200 mg/kg of gentamicin, there was total HC loss in the basal 18.0% of the sensory epithelium and partial HC loss in the basal 26.3%. Following 300 mg/kg of gentamicin, there was total HC loss in the basal 30.5% of the epithelium and partial HC loss in the basal 40.9%. The second goal of this study was to determine whether cannula implantation in the inner ear, and infusion of bromodeoxyuridine causes HC damage. We found that creation of a fistula in the labyrinth is not associated with HC damage, but that cannula implantation can cause HC death, and can also cause potentiation of gentamicin-induced HC death. Revision of the cannula and surgical technique to ensure minimal penetration into the labyrinth almost entirely eliminated these effects. We conclude that surgical technique is critical in experimental models in which the labyrinth is violated. PMID- 10713504 TI - Endotracheal isoflurane anesthesia for chick auditory surgery. AB - Survival surgeries upon chicks are commonly used in auditory research. Appropriate anesthesia is usually obtained with intramuscular or intraperitoneal injections of systemic agents. These techniques have several drawbacks, including delayed onset of anesthesia, difficulty in adjusting the dosage to accomodate individual animals' different responses, prolonged recovery times, and in some cases substantial mortality. We present a technique of administering inhaled isoflurane via an endotracheal tube which we have used for over a year with excellent results. With this agent, onset of deep anesthesia is very rapid, dosage can be titrated readily, overdosage is survivable, complete recovery occurs within a few minutes and mortality is rare. This technique may be valuable for other auditory scientists performing survival surgery in avian species. PMID- 10713505 TI - Stapedius muscle fibre composition in the rat. AB - The stapedius muscle (SM) is supposed to prevent cochlear damage by noise. Consequently functional demands are the ability of fast contraction with long endurance. This implies the presence of a large fraction of myosin type II fibres with an appreciable oxidative capacity. We determined the myosin composition of SM fibres using consecutive complete SM cross-sections (6 week old rats) which were processed by enzyme histochemistry (EHC) to determine acid/alkali lability of myofibrillar adenosine triphosphatase (mATPase) or by immunohistochemistry (IHC) using myosin heavy chain (MyHC) antibodies. Method accuracy was determined in co-processed extensor digitorum longus (EDL). Four hundred SM and 200 EDL fibres were assigned to mATPase type I, IIA, IIB, IIX or 'miscellaneous' ('Misc') categories. Per mATPase category the fibres were attributed to groups with specific MyHC composition. In the EDL, mATPase type I and IIB fibres expressed only MyHC I and IIB respectively, whereas about 10% of the type IIA and 40% of the type IIX fibres expressed more than one MyHC. Thus IHC detects amounts of myosin isoforms which are not detected by EHC. The mATPase IIX category criterion leaves the possibility that this category contains fibres with myosin type IIA and/or IIB in larger amounts. The criteria of the mATPase categories type I, IIA or IIB preclude assignment to these categories of fibres which also contain other myosin isoforms in larger amounts. Such fibres were classified in one of the mATPase 'Misc' categories. Thus in the EDL the capability of the EHC criteria to select 'pure' fibres in terms of myosin differs per mATPase category. None of the SM fibres were assigned to the mATPase type I or IIB categories, about 25% to the type IIA, 60% to type IIX and 15% (including most fibres which expressed MyHC I) to a 'Misc' category. All SM fibres expressed two or more MyHC isoforms, MyHC IIB occurring in all fibres and substantial amounts of MyHC IIA and/or IIX in most. These findings confirm the hypothesis that such fibres have the capacity to contract fast and have the better fatigue resistance. PMID- 10713506 TI - A saccular origin of frequency tuning in myogenic vestibular evoked potentials?: implications for human responses to loud sounds. AB - Previous research has indicated that an early component of click-evoked myogenic potentials in the sternocleidomastoid muscle is vestibularly mediated, since it can be obtained in subjects with loss of cochlear function, but is absent in subjects with loss of vestibular function (Colebatch et al., 1994). We report here the results of an experiment to investigate whether this response shows any tuning properties. In a sample of 11 subjects, we obtained acoustically evoked EMG from the sternocleidomastoid muscle in response to 110 dB SPL 10 ms tone pips with frequencies of 100 Hz, 200 Hz, 400 Hz, 800 Hz, 1600 Hz and 3200 Hz. The results of this experiment indicate that this response does indeed have a well defined frequency tuning which may be modelled as a resonance with a maximum response at frequencies between 300-350 Hz. The possible saccular origin of the tuning response and the consequences that this may have in human responses to loud sounds is discussed. Also discussed are the consequences of particular electrode arrangements in relation to the innervation and anatomy of sternocleidomastoid. PMID- 10713507 TI - Comparison of the vascular innervation of the rat cochlea and vestibular system. AB - In order to gain a better understanding of the neuronal and local control of inner ear blood flow, the vascular innervation to the rat cochlea and vestibular system was examined. Specimens were removed in toto beginning at the basilar artery extending to the anterior inferior cerebellar artery, labyrinthine artery, common cochlear artery, modiolar artery and anterior vestibular artery. When possible the vessels were dissected in continuity through the cribrose area. The vestibular endorgans were also removed. Specimens were examined using immunohistochemical techniques for the presence of vasoactive intestinal peptide, neuronal nitric oxide synthase, neuropeptide-Y, substance P and calcitonin gene related peptide. Results show that the vasculature to the cochlea and vestibular portion of the inner ear receive similar types of nonadrenergic innervation, that within the vestibular endorgans, only CGRP and SP were found in the neuroepithelium or in association with vessels, and that within the vestibular system, the majority of the vascular innervation appears to stop at or near the cribrose area. In the cochlea however, it extends to include the radiating arterioles. These findings suggest that cochlear blood flow is under finer control and that neuronally induced changes in blood flow may have a more global effect in the vestibular periphery. PMID- 10713508 TI - The effect of blood flow promoting drugs on cochlear blood flow, perilymphatic pO(2) and auditory function in the normal and noise-damaged hypoxic and ischemic guinea pig inner ear. AB - The effect of blood flow promoting drugs, such as hydroxyethyl starch (HES) either of low or high molecular weight (HES 70, HES 200), pentoxifylline, ginkgo biloba, naftidrofuryl and betahistine, and various combinations of the drugs was studied in unexposed and noise-exposed (broad-band noise, bandwidth 1-12 kHz, 106 dB SPL, 30 min) guinea pigs. The results were compared without therapy and placebo (isotonic saline, NaCl). The cochlear blood flow (CoBF) and the partial pressure of oxygen in the perilymph (PL-pO(2)) were continuously and simultaneously recorded over a period of 210 min. In addition, cochlear microphonics (CMs), compound action potentials of the auditory nerve (CAPs) and auditory brain stem responses (ABRs) were registered. Noise-induced hearing loss (NIHL) paralleled a decrease of PL-pO(2). Both were found to occur before evidence of reduced CoBF. PL-pO(2) and CoBF declined progressively post-exposure, while CMs, CAPs and ABRs showed no further deterioration or signs of recovery up to 180 min after cessation of noise. Treatment started 60 min post-exposure, respectively after 90 min, without manipulation in unexposed animals, and was then studied for a further 120 min. In unexposed animals, CoBF increased significantly during infusion of HES 70, HES 200, pentoxifylline and betahistine. NaCl, ginkgo biloba and naftidrofuryl did not alter CoBF. PL-pO(2) decreased significantly during infusion of all administered drugs and combinations, except for NaCl. CMs, CAPs and ABRs remained constant, with the exception of increased ABRs after infusion of HES 70 and HES 200. In noise-exposed animals, a sustained therapeutic effect on cochlear ischemia was achieved only by HES 200 and pentoxifylline. HES 70, betahistine and ginkgo biloba compensated cochlear ischemia only during infusion; however, 30-60 min after termination of therapy, no significant difference of values for CoBF was observed compared to the untreated noise-exposed groups. NaCl and naftidrofuryl showed no effect on CoBF. None of the applied drugs had a sustained compensatory effect on cochlear hypoxia. CMs, CAPs and ABRs improved significantly after HES 70, HES 200 and betahistine, resulting in partial recovery of CMs, and partial (betahistine) or even full (HES 70 and HES 200) recovery of CAPs and ABRs. In contrast, NaCl, pentoxifylline, ginkgo biloba and naftidrofuryl had no therapeutic effect on NIHL. PMID- 10713509 TI - Longitudinal threshold changes in older men with audiometric notches. AB - Age-related hearing loss (presbycusis) is a multifactorial process that results chiefly from the accumulating effects of noise damage and aging on the cochlea. Noise damage is typically evidenced clinically by a discrete elevation (notch) of the auditory thresholds in the 3-6 kHz region of the audiogram whereas aging affects the highest frequencies first. To determine whether the presence of such high-frequency notches influences auditory aging, we examined the 15 year change in audiometric thresholds in 203 men from the Framingham Heart Study cohort. The mean age at the first hearing test was 64 years (range 58-80). Occupational and recreational noise exposure over the 15 years was assumed to be minimal due to the age of the subjects. The presence or absence of a notch was determined using a piecewise linear/parabolic curve fitting strategy. A discrete elevation of the pure-tone thresholds of 15-34 dB in the 3-6 kHz region was deemed a small notch (N1), and elevations of 35 dB or greater were deemed large notches (N2). Absence of a notch (N0) was encoded those ears with <15 dB elevation in the 3-6 kHz region. The presence and absence of notches correlated with the subjects' history of noise exposure. The 15 year pattern of change in age-adjusted pure-tone thresholds varied significantly by notch category. There was less change over time in the notch frequencies (3-6 kHz) and significantly greater change in the adjacent frequency of 2 kHz in the N2 group as compared to the N0 and N1 groups. The adjacent frequency of 8 kHz showed a significant, but smaller, change in the N1 group as compared to the N0 and N2 groups. The change at 2 kHz was independent of the starting hearing level at E15, whereas the changes at 4-8 kHz were influenced by the hearing level at E15. These data suggest that the noise-damaged ear does not 'age' at the same rate as the non-noise damaged ear. The finding of increased loss at 2 kHz suggests that the effects of noise damage may continue long after the noise exposure has stopped. The mechanism for this finding is unknown but presumably results from prior noise-induced damage to the cochlea. PMID- 10713510 TI - Dendritic arbors on the saccule and lagena in the ear of the goldfish, Carassius auratus. AB - The ear of the goldfish (Carassius auratus) contains three otolithic endorgans: the saccule, lagena, and utricle. The saccule has an auditory function in most teleost fishes for whom data are available, and there is evidence that the lagena is also an auditory endorgan in the goldfish. This study was conducted to compare the innervation of the saccule and the lagena to one another and to previously published data from goldfish and other species. We placed cobaltous-lysine in saccular and lagenar nerves in vivo and permitted uptake over 18-24 h. A total of 59 saccular and 59 lagenar dendritic arbors were labeled in 10 fishes. Our data indicate that arbors on the saccule and lagena have similar morphologies, but differ in relative size. Saccular arbors tend to be smaller than lagenar arbors, with median arbor widths of 50 micrometer on the saccule and 74 micrometer on the lagena. Fiber diameters on the two endorgans are similar. A regional analysis of the saccule indicated that a wide range of arbor sizes are found along the rostral-caudal axis, with larger arbors more common caudally. Our data do not support the presence of two distinct categories of saccular afferents with non overlapping distributions. Moderate arbor widths (50-99 micrometer) were most common in all regions of the lagena. Maximum arbor width and hair cell density do not appear to be correlated with one another on either the saccule or the lagena. Comparisons with published data from goldfish and oscar revealed similarities and differences that may be attributable to variations in label uptake or transport as well as potential species differences. PMID- 10713511 TI - Validation of protein crystal structures. AB - Since the process of building and refining a model of a biomacromolecule based on crystallographic data is subjective, quality-control techniques are required to assess the validity of such models. During the 1990s, much experience was gained; the methods used and some of the lessons learned are reviewed here. In addition, an extensive compendium of quality criteria and quality-control methods that are or have been used to validate models of biomacromolecules has been compiled. The emphasis in this compendium is on the validation of protein crystal structures. PMID- 10713512 TI - Structure of FKBP12.6 in complex with rapamycin. AB - FKBP12.6 is a novel isoform of FKBP12, which selectively binds to the cardiac ryanodine receptor (RyR2). The crystal structure of FKBP12.6 in complex with rapamycin has now been determined at 2.0 A resolution. The structures of FKBP12.6 and FKBP12 are nearly identical, except for a displacement observed in the helical region of FKBP12.6 toward the hydrophobic pocket. This displacement was not predicted by homology modeling studies. Analyses of the residues that are likely to confer the RyR2-binding specificity are presented. PMID- 10713513 TI - Structural study of the complex between human pepsin and a phosphorus-containing peptidic -transition-state analog. AB - The refined crystal structure of the complex between human pepsin and a synthetic phosphonate inhibitor, Iva-Val-Val-Leu(P)-(O)Phe-Ala-Ala-OMe [Iva = isovaleryl, Leu(P) = the phosphinic acid analog of L-leucine, (O)Phe = L-3-phenyllactic acid, OMe = methyl ester], is presented. The structure was refined using diffraction data between 30 and 1.96 A resolution to a final R factor ( summation operator| |F(o)| - |F(c)| | / summation operator|F(o)|, where |F(o)| and |F(c)| are the observed and calculated structure-factor amplitudes, respectively) of 20.0%. The interactions of the inhibitor with the enzyme show the locations of the binding sites on the enzyme from S4 to S3'. Modeling of the inhibitor binding to porcine pepsin shows very similar binding sites, except at S4. Comparison of the complex structure with the structures of related inhibitors bound to penicillopepsin helps to rationalize the observed differences in the binding constants. The convergence of reaction mechanisms and geometries in different families of proteinases is also discussed. PMID- 10713514 TI - Crystal structure of delta-chymotrypsin bound to a peptidyl chloromethyl ketone inhibitor. AB - Chymotrypsin is a member of the trypsin family of serine proteases and is one of the first proteins successfully studied by X-ray crystallography. It is secreted into the intestine as the inactive precursor chymotrypsinogen; four sequential cleavages of the peptide bonds following residues 13, 15, 146 and 148 occur to generate the active pi, delta, kappa and alpha forms of chymotrypsin. (13)C NMR has shown [O'Connell & Malthouse (1995). Biochem. J. 307, 353-359] that when the delta form of chymotrypsin is inhibited by 2-(13)C-enriched benzyloxycarbonylglycylglycylphenylalanyl chloromethane, a tetrahedral adduct is formed which is thought to be analogous to the tetrahedral intermediate formed during catalysis. This inhibitor complex has been crystallized as a dimer in space group P4(1)2(1)2. The structure has been refined at 2.14 A resolution to an R value of 21.2% (free R = 25.2%). Conformational differences between delta chymotrypsin and chymotrypsinogen in the region of the flexible autolysis loop (residues 145-150) were observed. This is the first crystal structure of delta chymotrypsin and includes two residues which are disordered in previous crystal structures of active chymotrypsin. A difference of 11.3 A(2) between the average B values of the monomers within the asymmetric unit is caused by lattice disordering effects approximating to rotation of the molecules about a crystallographic screw axis. The substrate-binding mode of the inhibitor was similar to other chymotrypsin peptidyl inhibitor complexes, but this is the first published chymotrypsin structure in which the tetrahedral chloromethyl ketone transition-state analogue is observed. This structure is compared with that of a similar tetrahedral transition-state analogue which does not alkylate the active site histidine residue. PMID- 10713515 TI - Structure of the bifunctional inhibitor of trypsin and alpha-amylase from ragi seeds at 2.2 A resolution. AB - The crystal structure of a bifunctional inhibitor of alpha-amylase and trypsin (RATI) from ragi seeds (Indian finger millet, Eleusine coracana Gaertneri) has been determined by X-ray diffraction at 2.2 A resolution. The inhibitor consists of 122 amino acids, with five disulfide bridges, and belongs to the plant alpha amylase/trypsin inhibitor family. The crystals were grown by the microdialysis method using ammonium sulfate as a precipitating agent. The structure was determined by the molecular-replacement method using as models the structures of Corn Hageman factor inhibitor (CHFI) and of RATI at 2.9 A resolution determined previously. It has been refined to an R factor of 21.9%. The structure shows an r.m.s. deviation for C(alpha) atoms of 2.0 A compared with its own NMR structure, whereas the corresponding value compared with CHFI is found to be 1.4 A. The r.m.s. difference for C(alpha) atoms when compared with the same protein in the structure of the complex with alpha-amylase is 0.7 A. The conformations of trypsin-binding loop and the alpha-amylase-binding N-terminal region were also found to be similar in the crystal structures of native RATI and its complex with alpha-amylase. These regions differed considerably in the NMR structure. PMID- 10713516 TI - Structure of thrombin complexed with selective non-electrophilic inhibitors having cyclohexyl moieties at P1. AB - The crystal structures of five new non-electrophilic beta-strand-templated thrombin active-site inhibitors have been determined bound to the enzyme. Four co crystallize with hirugen and inhibitor isomorphously to produce thrombin-hirugen crystals (monoclinic, space group C2), while one co-crystallizes in the hexagonal system, space group P6(5). A 1,4-substituted cyclohexyl moiety is conserved at the P1 position of all the inhibitors, along with a fused hetero-bicyclic five- and six-membered ring that occupies the P2 site. Amino, amidino and aminoimidazole groups are attached to the cyclohexyl ring for recognition at the S1 specificity site, while benzylsulfonyl and diphenyl groups enhance the binding at the S3 subsite. The cyclohexyl groups at the P1 positions of three of the inhibitors appear to be in the energetically favored chair conformation, while the imidazole-substituted cyclohexyl rings are in a boat conformation. Somewhat unexpectedly, the two cyclohexyl-aminoimidazole groups bind differently in the specificity site; the unique binding of one is heretofore unreported. The other inhibitors generally mimic arginyl binding at S1. This group of inhibitors combines the non-electrophilicity and selectivity of DAPA-like compounds and the more optimal binding features of the S1-S3 sites of thrombin for peptidic molecules, which results in highly potent (binding constants 12 nM-16 pM, one being 1.1 microM) and selective (ranging from 140 to 20 000 times more selective compared with trypsin) inhibitors of thrombin. The binding modes of these novel inhibitors are correlated with their binding constants, as is their selectivity, in order to provide further insight for the design of therapeutic antithrombotic agents that inhibit thrombin directly at the active site. PMID- 10713517 TI - Structure determination of porcine haemoglobin. AB - To investigate a potential candidate material for making artificial red blood cells to supplement blood transfusion, the X-ray structure of porcine haemoglobin at 1.8 A resolution was determined as part of research towards synthesizing human blood. Porcine haemoglobin was crystallized by the vapor-diffusion method, producing crystals of dimensions 0.3-0.5 mm after successive seeding. The crystals belong to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 68.10, b = 72.27, c = 114.85 A. The initial phase was determined by the molecular-replacement method, using human oxyhaemoglobin as a model. The final R factor was 21.1% for 36 820 reflections after validation of 574 water molecules. The r.m.s. deviations of bond lengths, angles, torsion angles and improper angles from their ideal values are 0.017 A, 3.0, 20.6 and 1.8 degrees, respectively. The average B factor is 33.63 A(2) for the haemoglobin molecule and 50.53 A(2) for the water molecules. The structure could be superimposed on a 2.8 A resolution structure with an r.m.s. difference of 0.59 A in main-chain atomic positions and 1. 27 A in side-chain atomic positions. Porcine and human haemoglobins are compared. A tentative model for artificial blood is proposed based on the complementarity relationship of the surface charges between haemoglobin and the surrounding cell membrane. PMID- 10713518 TI - Pseudo-symmetry characterization and refinement of a trigonal crystal form of naphthalene 1,2-dioxygenase. AB - Two trigonal crystal structures of naphthalene 1,2-dioxygenase from Pseudomonas sp. NCIB 9816-4 have been refined at 2.6 A resolution. The space group is R3, with four heterodimers in the asymmetric unit. The crystallographic threefold axis coincides with the symmetry axis of the active molecule, a mushroom-shaped alpha(3)beta(3) hexamer. The crystal is formed by symmetrical contacts between the hexamers on three different interaction surfaces, one on the beta-subunit and the other two on the alpha--subunits. Nickel ions mediate one of the alpha subunit interactions. The two other types of packing contacts sustain two interlaced and almost independent crystal patterns with significantly different temperature factors. The space group of the individual crystal patterns is R32, with the corresponding twofold axes parallel to each other. The interactions between the crystal patterns separate the two parallel twofolds, eliminating the twofold symmetry for the whole crystal. The differences in temperature factors among the molecules in the asymmetric unit have been refined and are different for the two refined structures. An analysis of the structure factors of the pseudo-equivalent reflections showed that their differences lie in their phases and not in their amplitudes, suggesting that R(merge) is not an appropriate indicator for revealing the correct point group. PMID- 10713519 TI - Properties of an electron-density map derived from a limited number of experimentally determined triplet phases. AB - In a previous communication [Weckert et al. (1999). Acta Cryst. D55, 1320-1328], the feasibility of the measurement of a large set of triplet phases by three-beam interference was demonstrated. This paper reports the methodology for the calculation of an electron-density map from this limited amount of experimental phase information and the map's properties with respect to model building and refinement. The tetragonal form of hen egg-white lysozyme (HEWL) was chosen as a test structure for the development of this method. The quality of the electron density map obtained from all measured triplet phases allows a straightforward and nearly complete interpretation. The starting model was refined to a final R value of 17.4%. In a second step, the minimum number of phased reflections needed for the interpretation of an electron-density map was investigated, applying criteria based on |F| and resolution. PMID- 10713520 TI - Structural changes in a cryo-cooled protein crystal owing to radiation damage. AB - The high intensity of third-generation X-ray sources, along with the development of cryo-cooling of protein crystals at temperatures around 100 K, have made it possible to extend the diffraction limit of crystals and to reduce their size. However, even with cryo-cooled crystals, radiation damage becomes a limiting factor. So far, the radiation damage has manifested itself in the form of a loss of overall diffracted intensity and an increase in the temperature factor. The structure of a protein (myrosinase) after exposure to different doses of X-rays in the region of 20 x 10(15) photons mm(-2) has been studied. The changes in the structure owing to radiation damage were analysed using Fourier difference maps and occupancy refinement for the first time. Damage was obvious in the form of breakage of disulfide bonds, decarboxylation of aspartate and glutamate residues, a loss of hydroxyl groups from tyrosine and of the methylthio group of methionine. The susceptibility to radiation damage of individual groups of the same kind varies within the protein. The quality of the model resulting from structure determination might be compromised owing to the presence of radiolysis in the crystal after an excessive radiation dose. Radiation-induced structural changes may interfere with the interpretation of ligand-binding studies or MAD data. The experiments reported here suggest that there is an intrinsic limit to the amount of data which can be extracted from a sample of a given size. PMID- 10713521 TI - Crystallization and preliminary X-ray study of beta--mannosidase from Trichoderma reesei. AB - beta-Mannosidase from Trichoderma reesei, a 105 kDa glycoprotein, has been crystallized. The crystals belong to the space group P4(1)2(1)2 or P4(3)2(1)2, with unit-cell dimensions a = b = 165.86, c = 122.46 A, and diffract beyond 2.75 A resolution. X-ray diffraction data were collected from a frozen crystal on a synchrotron X-ray source. PMID- 10713522 TI - Crystallization and preliminary X-ray diffraction studies of d(ACGTAGCTACGT)2:[actinomycin D, (echinomycin)2] and d(ACGTAGCTACGT)2:[actinomycin D, (triostin A)2] complexes. AB - A DNA-multiple drug complex, d(ACGTAGCTACGT)(2):[actinomycin D, (echinomycin)(2)] has been crystallized. The crystals belong to the monoclinic space group C2, with unit-cell parameters a = 85.6, b = 72.8, c = 56.6 A, beta = 101.5 degrees at 93 K and Z = 8. The crystal diffracted to 3.0 A resolution along the DNA fiber axis and to 3.5 A resolution in other directions. The Patterson maps indicate that all complexes in the crystal are oriented along their helical axes in the [101;] direction. PMID- 10713523 TI - Structural studies of MIP synthase. AB - The conversion of glucose 6-phosphate to 1-L-myo-inositol 1--phosphate (MIP) by 1 L-myo-inositol 1-phosphate synthase (MIP synthase) is the first committed and rate-limiting step in the de novo biosynthesis of inositol in all eukaryotes. The importance of inositol-containing molecules both as membrane components and as critical second messenger signal-transduction species make the function and regulation of this enzyme important for a host of biologically important cellular functions including proliferation, neurostimulation, secretion and contraction. MIP synthase has been overexpressed in Esherichia coli and purified to homogeneity by chromatographic methods. Two crystal forms of MIP synthase were obtained by the hanging-drop vapor-diffusion method. Native data sets for both crystal forms were collected in-house on a Rigaku R-AXIS IIC imaging-plate detector. Crystal form I belongs to space group C2, with unit-cell parameters a = 153.0, b = 96.6, c = 122.6 A, beta = 126.4 degrees, and diffracts to 2.5 A resolution. Crystal form II belongs to space group P2(1), with unit-cell parameters a = 94.5, b = 186.2, c = 86.5 A, beta = 110.5 degrees, and diffracts to 2.9 A resolution. PMID- 10713524 TI - Purification, crystallization and x-ray analysis of crystals of pectate lyase A from Exwinia chrysanthemi. AB - Pectate lyase A is secreted by Erwinia chrysanthemi and is a virulence factor for soft rot diseases in plants. Crystals of pectate lyase A were obtained by vapor diffusion techniques in the presence of polyethylene glycol. The crystals belong to the monoclinic space group P2(1), with unit-cell parameters a = 48.96, b = 148.86, c = 78.61 A, beta = 97.32 degrees. The crystals contain two protein molecules of 38 kDa per asymmetric unit and diffract to 2.4 A using Cu Kalpha radiation. PMID- 10713525 TI - Crystallization and preliminary X-ray analysis of insect antifreeze protein from the beetle Tenebrio molitor. AB - Hyperactive antifreeze protein from the beetle Tenebrio molitor (TmAFP) was produced in Escherichia coli and purified by gel-permeation chromatography and HPLC. An iodinated derivative was prepared by incubating the 8.5 kDa TmAFP with N iodosuccinimide. Native and iodinated TmAFP produced two different crystal forms when crystallized using the hanging-drop vapor-diffusion technique. Native crystals were rectangular plates that diffracted to approximately 2.5 A resolution. They were monoclinic and belonged to the space group P2(1), with unit cell dimensions a = 38.4, b = 73.4, c = 59.3 A, beta = 97.0 degrees. Crystals of iodinated TmAFP formed elongated hexagons that allowed data to be collected to approximately 1.4 A. These crystals belonged to the space group P6(1) (or P6(5)), with unit-cell dimensions a = 73.85, b = 73.85, c = 53.15 A. There were two molecules per asymmetric unit, which corresponds to V(m) = 2.46 A(3) Da(-1) and 51% solvent content. A twofold non-crystallographic symmetry was evident from self-rotation calculations. PMID- 10713526 TI - Crystallization and preliminary x-ray crystallographic analysis of NAD+-dependent DNA ligase from Thermus filiformis. AB - A highly thermostable DNA ligase from Thermus filiformis has been crystallized at room temperature using methoxypolyethylene glycol 5000 as a precipitant. The crystal belongs to the monoclinic space group P2(1), with unit-cell parameters a = 90.63, b = 117.80, c = 98. 65 A, beta = 115.56 degrees. Two molecules of DNA ligase are present in the asymmetric unit, giving a crystal volume per protein mass (V(m)) of 3.1 A(3) Da(-1) and a solvent content of 61%. A native data set extending to 3.0 A resolution has been collected at 100 K using synchrotron X rays. PMID- 10713527 TI - Crystallization and preliminary X-ray analysis of the catalytic domain of the adenylate cyclase GRESAG4.1 from Trypanosoma brucei. AB - Adenylate cyclases (ACs) are involved in signal transduction by generating the second messenger, cAMP. In Trypanosoma brucei, 3', 5'-cyclic adenosine monophosphate (cAMP) controls the life cycle of this unicellular parasite. cAMP is generated by a class of adenylate cyclases which are either constitutively (GRESAG4.1-4.3) or transiently expressed (ESAG4) during the life cycle. Unlike mammalian ACs, the trypanosomal ACs have a simple topology comprising of a large extracellular region, a transmembrane helix and a cytosolic catalytic region. Two orthorhombic crystal forms of the catalytic AC domain of GRESAG4.1 (residues Ala884-Thr1132) were generated by the hanging-drop vapour-diffusion method. X-ray diffraction data from GRESAG4.1 crystals were collected at 1.9 A resolution using synchrotron radiation. Furthermore, two heavy-metal derivative data sets were collected from crystal form A; heavy-atom sites were subsequently located in difference Patterson maps. PMID- 10713528 TI - Crystallization and preliminary x-ray diffraction analysis of beta-cinnamomin, an elicitin secreted by the phytopathogenic fungus Phytophthora cinnamomin. AB - Cinnamomin (CIN) belongs to a family of 10 kDa proteins designated as elicitins. Some of these proteins induce a hypersensitive response in diverse plant species, leading to resistance against fungal and bacterial plant pathogens. CIN was crystallized by the vapour-diffusion method using either ammonium sulfate or polyethyleneglycol (PEG) as precipitants in solutions buffered at around pH 7. These crystals are isomorphous and belong to the triclinic space group, with unit cell parameters a = 31.69, b = 36. 99, c = 44.09 A, alpha = 76.86, beta = 84.41, gamma = 80.26 degrees. A frozen crystal diffracted X-rays beyond 1.45 A resolution on a synchrotron-radiation source. PMID- 10713529 TI - Purification, crystallization and preliminary x-ray diffraction analysis of carboxyhaemoglobin-II from the fish Piaractus mesopotamicus (pacu). AB - Carboxyhaemoglobin-II isolated from the pacu (Piaractus mesopotamicus) has been crystallized and X-ray diffraction data were collected to 2.0 A resolution using synchrotron radiation. Crystals were characterized as belonging to the space group I23; preliminary structural analysis reveals the presence of one dimer in the asymmetric unit. PMID- 10713530 TI - How do the x-ray structure and the NMR structure of FMN-binding protein differ? AB - The crystal structure of FMN-binding protein (FMN-bp) from Desulfovibrio vulgaris Miyazaki F was solved by the multiple isomorphous replacement method and refined to an R factor of 15.1% at 1.3 A resolution. FMN-bp exists in a dimeric form in the crystal, in contrast to the monomeric structure determined by NMR. R.m.s. deviations between the crystal structure and the solution structure are more than 2 A, which implies significant differences. There are some hydrophobic residues in the interface between the two monomers. In particular, Leu122 in the C terminus has a close contact with the o-xylene moiety of FMN, while solvent molecules may cover the o-xylene moiety in the solution structure. PMID- 10713531 TI - Arabidopsis thaliana peroxidase N: structure of a novel neutral peroxidase. AB - The structure of the neutral peroxidase from Arabidopsis thaliana (ATP N) has been determined to a resolution of 1.9 A and a free R value of 20.5%. ATP N has the expected characteristic fold of the class III peroxidases, with a C(alpha) r.m.s.d. of 0.82 A when compared with horseradish peroxidase C (HRP C). HRP C is 54% identical to ATP N in sequence. When the structures of four class III plant peroxidases are superimposed, the regions with structural differences are non randomly distributed; all are located in one half of the molecule. The architecture of the haem pocket of ATP N is very similar to that of HRP C, in agreement with the low small-molecule substrate specificity of all class III peroxidases. The structure of ATP N suggests that the pH dependence of the substrate turnover will differ from that of HRP C owing to differences in polarity of the residues in the substrate-access channel. Since there are fewer hydrogen bonds to haem C17 propionate O atoms in ATP N than in HRP C, it is suggested that ATP N will lose haem more easily than HRP C. Unlike almost all other class III plant peroxidases, ATP N has a free cysteine residue at a similar position to the suggested secondary substrate-binding site in lignin peroxidase. PMID- 10713532 TI - 1.7 A x-ray structure of space-grown collagenase crystals. AB - Collagenases, divided into metallocollagenases and serine collagenases, are the only proteases that cleave the triple helix of collagen under physiological conditions. In the present work, the serine protease collagenase purified from Hypoderma lineatum larvae is studied. From crystals grown in the International Microgravity Laboratory (IML2), a data set was collected at 1.7 A using synchrotron radiation. Although the resolution is not very different, the signal to-noise ratio and the quality of the electron density are much improved. Alternate conformations were revealed for several residues, in particular Tyr99, suggesting a gate mechanism of recognition. PMID- 10713533 TI - Rational molecular design in drug research PMID- 10713534 TI - The many faces of RNA PMID- 10713535 TI - Electron microscopy methods and protocols PMID- 10713536 TI - Structure-based drug design PMID- 10713538 TI - New cryostream cooler PMID- 10713537 TI - Patterson peaks PMID- 10713539 TI - Methods of isolation of reaction center preparations from photosynthetic purple bacteria. AB - Various modern methods of isolation of functionally active membrane complexes of the reaction centers (RC) from photosynthetic purple bacteria are reviewed. Special attention is given to the methods of RC isolation from bacteria which are widely used in experimental practice. The analysis includes the main steps of RC isolation, evaluation of purity of the resultant preparation, and characterization of its functional activity. Besides description of conventional methods of RC isolation based on ion-exchange chromatography and hydroxyapatite chromatography, some other methods such as affinity chromatography and high performance chromatography at high and fine pressure are also considered. PMID- 10713540 TI - Study of interaction of human replication factor A with DNA using new photoreactive analogs of dTTP. AB - Replication factor A (RPA) is a protein that binds single-stranded DNA in eukaryotic cells; it participates in replication, repair, and recombination of DNA. RPA is composed of three subunits with molecular masses 70 (p70), 32 (p32), and 14 kD (p14). The photoaffinity labeling method was used to study the interaction of RPA with the 3;-end of duplex DNA containing extended 5;-end of a single strand. We have synthesized dTTP analogs containing photoreactive 2,3,5,6 tetrafluoro-4-azidobenzoyl group attached to the 5th position of the uracil residue with linkers of variable length (9, 11, and 13 atom chains). Using these analogs and dTTP analog containing the same photoreactive residue attached to the 5th position of the uracil residue with a 4-atom linker, a number of oligonucleotide primers carrying a single photoreactive group on the 3;-end were enzymatically synthesized. Using the complex of the photoreactive primers with DNA template containing extended 19-base 5;-end, human RPA was photoaffinity modified. The primers were covalently bound to the p70 and p32 subunits of RPA and the p14 subunit was not labeled by the primers. The data are discussed considering the previously suggested model of interaction of RPA with DNA during replication. PMID- 10713541 TI - Aminopeptidase PC from the hepatopancreas of the Kamchatka crab Paralithodes camtshatica. AB - Homogeneous aminopeptidase PC was isolated with yield 67% and purification degree 237 from the hepatopancreas of the Kamchatka crab Paralithodes camtshatica by ion exchange chromatography on DEAE-Sepharose, hydrophobic chromatography on Phenyl Sepharose, and gel-filtration on Sephadex G-150. The enzyme is a homodimer with a molecular mass 220 kD (110 x 2). Aminopeptidase PC has pI = 4.1. It hydrolyzes Leu-pNA optimally at pH 6.0 and at the optimum temperature 36-40 degrees C; in the presence of Ca2+ the enzyme is stable at pH 5.5-8.0. Aminopeptidase PC is activated by Ca2+, Mg2+, and Fe2+; it is completely inhibited by EDTA, o phenanthroline, and bestatin. The enzyme contains four Zn atoms per molecule and is therefore a metalloaminopeptidase. The aminopeptidase PC can effectively cleave N-terminal Arg and Lys residues as well as Leu, Phe, and Met residues. Km and kcat values for hydrolysis of Leu-pNA were 0.075 mM and 0.19 sec-1 and for hydrolysis of Arg-pNA 0.078 mM and 0.48 sec-1, respectively. D-Amino acid residues cannot be cleaved. Thus, aminopeptidase PC of the Kamchatka crab has a mixed substrate specificity which is characteristic of some microbe aminopeptidases. Its N-terminal sequence ESVEIELPEGLSPLV is 46% coincident with that of yeast vacuolar aminopeptidase YSCA. PMID- 10713542 TI - Parathyroid hormone as a modulator of the functional activity of neuron. AB - The effect of 10-10 M parathyroid hormone (PTH) on Ca2+ levels and cAMP and cGMP contents in peripharyngeal ganglions of the snail Helix pomatia was studied by radioisotope and radioimmune methods; also, chemoreception of GABA was determined in the neuronal membrane. The levels of Ca2+ and cyclic nucleotides were increased in the ganglions. Co-addition of PTH with compounds increasing the levels of Ca2+ and cyclic nucleotides lowered these elevations of the levels of Ca2+ and cyclic nucleotides. Depending on the initial levels of Ca2+ and cyclic nucleotides in the neuron, PTH can direct the fluxes of ions either into or out of the cell and can activate the cyclase or phosphodiesterase systems. PTH decreased the binding of GABA by the ganglions at high GABA concentrations and under conditions which promote increased binding of GABA. These data suggest that PTH has neuroprotective effects and protects the neuronal tissue from inhibition. Thus, PTH can modulate the function of neurons. PMID- 10713543 TI - Structure of the O-specific polysaccharide of the bacterium Proteus mirabilis O29. AB - An O-specific polysaccharide was obtained by mild acid degradation of P. mirabilis O29 lipopolysaccharide (LPS) and found to contain 2-acetamido-2-deoxy-D galactose and D-glucuronic acid (D-GlcA) in the ratio 3:1. Studies of the polysaccharide by 1H- and 13C-NMR spectroscopy including two-dimensional correlation spectroscopy (COSY), total correlation spectroscopy (TOCSY), nuclear Overhauser effect spectroscopy (NOESY), and H-detected 1H,13C-heteronuclear multiple-quantum coherence (HMQC) experiments demonstrated the following structure of the branched tetrasaccharide repeating unit: PMID- 10713544 TI - Effect of ethanol on the hemolytic stability of erythrocytes. AB - The stability of rabbit erythrocytes to hemolysis induced by different compounds in the presence or absence of ethanol or acetaldehyde has been analyzed. Ethanol slightly reduced erythrocyte stability against acidic hemolysis only after long term preincubation, but the effect of ethanol on stability to oxidative hemolysis manifested itself immediately after its addition to the cells. Ethanol decreased both stability of cells to oxidative damage and dispersion of the hemolytic curve. Comparison of the effects of ethanol and acetaldehyde showed that the destabilizing effect of ethanol might be caused by either its direct action or the effect of its metabolites formed during preincubation of ethanol with erythrocytes. Possible mechanisms of ethanol and acetaldehyde effects on erythrocyte stability are discussed. PMID- 10713545 TI - Cytochrome P450 1A2: oligomers in proteoliposomes. AB - The presence of oligomers of cytochrome P450 1A2 in membranes of proteoliposomes produced by the cholate-dialysis technique was demonstrated by cross-linking of protein molecules with bifunctional reagents followed by electrophoretic analysis of the modified proteins. A hexameric organization of cytochrome P450 1A2 in the membrane of proteoliposomes is suggested with high probability based on the comparison of the purified hemoprotein oligomeric structure in an aqueous medium and that in the proteoliposomes. The comparison was carried out using the same method. PMID- 10713546 TI - Changes in the hormonal status of the Taraxacum officinale Web. ovary at early stages of embryogenesis. AB - The hormonal status of the Taraxacum officinale Web. ovary was quantitatively assayed for the first time during early stages of embryogenesis. Apparent concentrations of endogenous cytokinins were measured using two systems of enzyme linked immunosorbent assay (ELISA). The ELISA systems differed from one another by the specificity for the main endogenous forms of zeatin. The specificity of two heterological ELISA systems based on zeatin- and kinetin-specific antisera was studied. A new immunochemical approach to the problem of differential quantitative determination of natural zeatin forms is suggested. This approach does not require preliminary separation of experimental samples into individual fractions. True concentrations of zeatin and zeatin riboside in the T. officinale ovary were calculated based on the average values of apparent concentrations of endogenous cytokinins. When the embryo sac maturation had been completed, there was a threefold increase in the zeatin riboside concentration within the following 12 h. By the time of the first division of an unfertilized ovicell (i.e., within the next 12 h), there had been a twofold decrease in the zeatin riboside concentration. Therefore, at early stages of division of the unfertilized ovicell the zeatin riboside concentration virtually returned to the initial level. In contrast to zeatin riboside, there was a steady trend toward an increase in the zeatin concentration in the T. officinale ovary. Within the first 12 h and the next 12 h after completion of the embryo sac maturation, the zeatin concentration was increased 1.5-fold and 2-fold, respectively. The results of this work provide a pioneering insight into the dynamics of various natural forms of zeatin during the reproductive process. The immunochemical approach to quantitative monitoring of various natural forms of zeatin and their dynamics during embryogenesis suggested in this work can be extended to similar biological, medical, and agricultural problems of differential determination of low-molecular-weight agents of similar structure but different biological activity. PMID- 10713547 TI - Isolation and characterization of soybean Bowman-Birk inhibitor from different sources. AB - A chromatographic procedure for isolation of different isoforms of Bowman--Birk soybean trypsin inhibitors was developed. The number of isoforms was shown to depend on soybean cultivar. The amount of the classical Bowman--Birk inhibitor (BBI) in different soybean cultivars, commercial flour, and processing products was analyzed. BBI reaches its highest concentration in freshly milled seeds. Storage conditions optimum for preservation of maximum inhibitory activity in soybean raw material were developed. The use of indirect enzyme immunoassay for BBI detection during its isolation from different sources was demonstrated. PMID- 10713548 TI - Stimulation of nitrate reductase activity of the salt-tolerant yeast Rhodotorula glutinis by tungsten in the presence of molybdenum. AB - Tungsten in the presence of molybdenum stimulates nitrate reductase activity and growth of the salt-tolerant yeast Rhodotorula glutinis on medium with nitrates. Tungsten is not incorporated in proteins possessing nitrate reductase activity. A significant increase in molybdenum cofactor in cells grown on medium with equimolar amounts of molybdenum and tungsten may relate to the stimulatory action of tungsten. PMID- 10713549 TI - Inhibition of heat-induced aggregation of beta- and gamma-crystallin by alpha crystallin evaluated by gel permeation HPLC. AB - The capability of alpha-crystallin (alpha-C), a known molecular chaperon, of protecting beta-C and gamma-C against heat-induced aggregation was studied by gel permeation high performance liquid chromatography. The activity was calculated using a formula based on the changes in the areas under the chromatographic peaks of these proteins, which appeared well separated. When heat-induced aggregation was studied in the range 22-90 degrees C, beta-C appeared more stable than gamma C. The activity of alpha-C in stabilizing gamma-C but not beta-C was already relevant at 60 degrees C, but the maximum activity was higher (about 35%) for beta-C than for gamma-C. This method could be useful for studying the effect of drugs with potential anti-cataract activity on heat-induced aggregation of individual lens proteins. PMID- 10713550 TI - Slowing of proton transport processes in the structure of bacterial reaction centers and bacteriorhodopsin in the presence of dipyridamole. AB - Dipyridamole, 2,6-bis(diethanolamino)-4,8-dipiperidinopyrimido(5, 4-d)pyrimidine, is employed in clinical practice as a vasodilator. It can also inhibit a specific membrane protein (glycoprotein P) which pumps anticancer drugs out of tumor cells. Dipyridamole (10-4 M) markedly slows down the kinetics of the electrogenic phase of the photoelectric response in Rhodobacter sphaeroides chromatophores. This phase is due to proton transfer from the external medium to the secondary quinone acceptor in the reaction center. In purple membranes of bacterium Halobacterium salinarium containing bacteriorhodopsin dipyridamole (in its charged state) significantly slowed the kinetics of proton transfer from the primary donor, Asp-96 (in membranes from bacteria of wild type), or from the external medium (in D96N mutant) to the Schiff base. It is suggested that dipyridamole can influence the structural-dynamic state of membrane proteins including modification of the structure of their hydrogen bonds involved in proton-transport processes. PMID- 10713551 TI - Inhibition of 2,4-dinitrophenol-induced potassium efflux by adenine nucleotides in mitochondria. AB - The influence of nucleotides on 2,4-dinitrophenol (DNP)-induced K+ efflux from intact rat liver mitochondria has been studied. ATP and ADP at micromolar concentrations were found to inhibit mitochondrial potassium transport, whereas GTP, GDP, CTP, and UTP did not show tha same effect. The values of half-maximal inhibition (IC50) were approximately 20 microM for ATP and approximately 60 microM for ADP. It is suggested that adenine nucleotides exert their inhibitory action at the matrix side of the inner mitochondrial membrane since the inhibitor of adenine nucleotide translocase atractyloside at concentration of 1 microM completely removed the inhibitory effect of ATP and ADP. The mitochondrial ATPase inhibitor oligomycin (2 microg/ml) was found to reduce slightly the rate of DNP induced K+ efflux and had no effect on inhibition by adenine nucleotides; the latter was insensitive to Mg2+ and the changes in pH. It seems likely that the regulation of potassium transport is not due to phosphorylation of the channel forming protein but to binding of the nucleotides in specific regulatory sites. The possibility of potassium efflux from mitochondria in the presence of uncoupler via the ATP-dependent potassium channel is discussed. PMID- 10713552 TI - Bioenergetic response of isolated nerve terminals of rat brain to osmotic swelling. AB - The swelling of nerve terminals of rat brain in a hypotonic medium (230 mOsm) induced the potential-independent entrance of 45Ca2+ into synaptosomes and intrasynaptosomal mitochondria that changed the energy status of synaptosomes, the rate of O2 consumption and the content of ATP being decreased. The ratio ATP/ADP decreased from 6.5 +/- 0.26 (310 mOsm medium) to 3.1 +/- 0.18 (the medium 230 mOsm). Studies on the equilibrium distribution of K+ (86Rb+) and [3H]TPP+ showed that contents of these cations in the nerve terminals were virtually the same on incubation in both iso- and hypotonic media. This indicated that the swelling did not damage intrasynaptosomal mitochondria and plasma membranes of the synaptosomes. The inhibition of oxidative phosphorylation increased twofold the rate of glycolysis. The incubation of synaptosomes in calcium-free medium (230 mOsm) in the presence of EGTA (1 mM) prevented the inhibition of oxidative phosphorylation and synthesis of ATP by the osmotic swelling. Ruthenium Red (10 microM) in the medium 230 mOsm inhibited the entrance of 45Ca2+ into the intrasynaptosomal mitochondria and normalized the oxidative phosphorylation to the control level (310 mOsm medium). The decrease in the energy potential of synaptosomes induced by the hypoosmotic shock is suggested to be associated with the increase in Ca2+ content in the cytoplasm, its transport into the mitochondria, and the inhibitory effect on oxidative phosphorylation. PMID- 10713553 TI - N-Ethylmaleimide induces disaggregation of platelets preaggregated by ADP and thrombin and decreases cytoplasmic calcium level. AB - The effect of N-ethylmaleimide (NEM, 1-200 microM) on ADP- and thrombin-induced platelet aggregation and thrombin-induced increase in intracellular Ca2+ concentration was studied. Addition of NEM to platelets preaggregated with ADP or thrombin induces platelet disaggregation. The anti-aggregant activity of NEM was different for ADP- and thrombin-induced aggregations. At 200 microM concentration, NEM completely disaggregated ADP-induced aggregates and only partially disaggregated thrombin-aggregated platelets. NEM did not influence the thrombin-induced increase in cytoplasmic Ca2+ and had no effect on the basal level of Ca2+ in the cytosol of non-activated platelets. However, NEM decreased the level of thrombin-mobilized Ca2+ in the cytosol of activated platelets. Thus, NEM can induce disaggregation of ADP- and thrombin-preaggregated platelets by activating a system which removes Ca2+ from the platelet cytosol. PMID- 10713554 TI - Influence of nucleic acids and polysaccharides on phosphotransferase activity of preparations of secretory immunoglobulin A from human milk. AB - The influence of nucleic acids (DNA, tRNA), synthetic oligonucleotides, and polysaccharides (lipopolysaccharides from Escherichia coli, heparin) on protein kinase and lipid kinase activities of preparations of human secretory immunoglobulin A (sIgA) has been studied. The preparations of sIgA were isolated from human milk by chromatography on the column with Protein A-Sepharose and DEAE sorbent (sIgA1), by affinity chromatography of sIgA1 on DNA-cellulose (sIgA2), and by gel-filtration of sIgA1 in buffer containing 5% dioxane (sIgA3). Two 32P labeled products with high and low electrophoretic mobility in polyacrylamide gel containing SDS were found after incubation of sIgA1 and sIgA2 with [gamma 32P]ATP. The product with low electrophoretic mobility was degraded in 10% trichloroacetic acid giving a radioactive background in lanes of the polyacrylamide gel. 32P-Labeled phospholipids were found among the phosphorylation products. Soluble and immobilized DNA increase lipid kinase activity of preparations of sIgA. In this case the secretory component and H chains of sIgA were degraded. Fractions possessing lipid kinase activity were precipitated in the presence of heparin (1 mg/ml), and lipid kinase activity was separated from sIgA by gel-filtration in buffer containing 5% dioxane. 32P Labeled products were formed in the presence of [gamma-32P]ATP as well as [32P]ortho-phosphoric acid. The influence of heparin and synthetic deoxy- and ribooligonucleotides on casein kinase activity of sIgA3 was studied. It was observed that deoxyribooligonucleotides in micromolar concentrations increased the rate of casein phosphorylation in the presence of sIgA3 and [gamma-32P]ATP. It has been proposed that catalytically active sIgA have an affinity to DNA (anti DNA sIgA) and can be present in human milk as a part of lipoprotein complexes. PMID- 10713555 TI - Highly selective affinity labeling of DNA-polymerase from Thermus thermophilus B35 by a binary system of photoreactive agents. AB - The thermostable DNA-polymerase from Thermus thermophilus B35 (Tte-polymerase) was affinity labeled by a binary system of photoreagents comprising base substituted TTP analogs. The 5;-[32P]-labeled primer was elongated by Tte polymerase in the presence of a TTP analog containing the photoreactive 2,3,5, 6 tetrafluoro-4-azidobenzoyl group (FAB-4-dUTP). Then the reaction mixture was UV irradiated (365-450 nm) in the presence or the absence of a photosensitizer (TTP analog containing a pyrene moiety, Pyr-dUTP). The initial rate of the Pyr-dUTP sensitized photomodification was almost 10-fold higher than the rate of direct photomodification (in the absence of Pyr-dUTP); in the case of the sensitized modification, the product of covalent cross-linking of the photoreactive primer with Tte-polymerase was apparently homogenous according to the data of electrophoresis. The enzyme was protected from the photosensitized modification by dNTP. To confirm the selectivity of the photosensitized modification of Tte polymerase, another DNA-binding protein (human replication factor A, RPA) was added to the reaction mixture. In the presence of the photosensitizer (Pyr-dUTP), RPA was not labeled and only Tte-polymerase was modified, whereas in the case of direct modification, Tte-polymerase and the p32 and p70 subunits of RPA were labeled. The suggested method enables highly selective affinity modification of DNA-polymerases. PMID- 10713556 TI - Effect of gallic acid polydisulfide on activity and stability of catalase in various media. AB - The effect of the active bioantioxidant polydisulfide of gallic acid (PDSG) on the catalytic activity and operational and thermal stability of catalase was studied in three media: distilled water (pH approximately 5.6), phosphate buffer, pH 7.4, and reversed micelles of Aerosol OT (AOT) in heptane of varied hydration degree w0. PDSG inhibited the catalase-induced decomposition of H2O2 by the mixed or noncompetitive mechanism: in various media the inactivation constant Ki varied in the range of (0.63-2.32).10-5 M. PDSG nearly twofold decreased the rate constant of interaction of the complex I of catalase with H2O2 (k2, M-1.sec-1) in water and reversed micelles of AOT and 3-5 times increased the effective rate constant of catalase thermal inactivation, k*in, sec-1, depending on the reaction medium. PDSG significantly decreased the rate constant of catalase inactivation during the enzymatic reaction, kin, sec-1, and thus increased the enzyme operational stability in water and reversed AOT micelles in heptane. The interaction of PDSG with catalase in water and in phosphate buffer was accompanied by significant changes in CD spectra in the far UV-region that indicated disturbances in the secondary structure of catalase subunits induced by the bioantioxidant; the latter was suggested to initiate the reaction of thiol- disulfide exchange with the enzyme. The problem of the compatibility of catalase with disulfide bioantioxidants is discussed. PMID- 10713557 TI - Localization, purification, and characterization of the rabbit sarcoplasmic reticulum associated calmodulin-dependent protein kinase. AB - The Ca2+/calmodulin dependent protein kinase associated with the sarcoplasmic reticulum membranes (SR CaM kinase) plays a specific and important role in the modulation of both Ca2+ uptake and release functions of the sarcoplasmic reticulum itself. In this work we have localized a 60 kD SR CaM kinase in slow and fast twitch rabbit skeletal muscle fractions; the kinase was present in both the longitudinal and the junctional sarcoplasmic reticulum. We then developed a procedure for the purification of the active kinase from the longitudinal sarcoplasmic reticulum and performed biochemical and functional characterization of the enzyme. Differently from what was previously suggested, our analysis shows that the biochemical properties of the purified SR CaM kinase (Ca2+ sensitivity, K0.5 for calmodulin, Km for ATP, IC50 for the specific inhibitory peptide (290 309), autophosphorylation properties) are not significantly different from those of the soluble multifunctional CaM kinase II. Moreover, we show that the purified SR CaM kinase retains the ability to autophosphorylate in a Ca2+/calmodulin dependent manner, becoming a Ca2+-independent enzyme. In the light of the knowledge of the rabbit SR CaM kinase biochemical properties, we propose and discuss the possibility that, under physiological conditions, the activity of the autophosphorylated kinase persists when the Ca2+ transient is over. PMID- 10713558 TI - Safety and cleaning of medical materials and devices. AB - A study was undertaken to evaluate different procedures to safely remove microorganisms, protein, and mammalian cells from materials and provide a suitable method for cleaning and assessing effectiveness of cleaning medical devices for reuse or for analysis of failure. Safety considerations for the personnel performing the cleaning or handling the device after cleaning are important issues. Polystyrene plates (96 well) were used to simulate device surfaces not amenable to manual scrubbing. Staphylococcus epidermidis, Candida albicans, Escherichia coli, Pseudomonas aeruginosa, and oral flora were grown in the plates. The plates were stained with crystal violet and the optical densities recorded. The results indicated that E. coli did not adhere well and Pseudomonas formed clumps that were easily detached from the surface of the plates. However, S. epi, C. albicans, and the oral organisms formed adherent biofilms that were difficult to remove from the plates. Detergents with enzymes and sodium hypochlorite (NaOCl) bleach were both effective in removing the biofilm. Other detergents and surfactants were not effective. The aldehyde agents did not remove the organisms and made further cleaning difficult. Allowing the biofilm to dry first made cleaning very difficult. Only the NaOCl bleach could subsequently remove the dried or aldehyde fixed organisms from the wells. The same 96-well polystyrene plate format was used to measure the amount of protein and cell adherence as well as the effectiveness of subsequent cleaning. Bradford reagent was used to detect protein as a measure of the cleaning efficacy. As with the bacteria, NaOCl bleach was effective at removing the protein and cells that had been dried or fixed by formalin or alcohol, whereas detergent with enzymes was not very effective. This study confirmed that used medical devices, contaminated with microorganisms, protein, and/or mammalian cells, should not be allowed to dry before cleaning and that a thorough cleaning procedure should precede sterilization or disinfection (with the exception of NaOCl bleach which also cleans). PMID- 10713559 TI - Elemental and morphological identification of third-body particulate and calcium stearate inclusions in polyethylene components. AB - Third-body particulate such as human bone chips, hydroxyapatite, and bone cement are considered contributing factors in accelerated wear in total joint replacement. Particulate wear debris is now considered the major contributing factor in aseptic loosening of total joint replacements. The ability to distinguish between different third-body particulate is necessary to better understand wear mechanisms when conducting implant retrieval analysis. The objective of this investigation is to demonstrate that backscattered electron imaging with correlated energy dispersive X-ray analysis can accurately identify third-body particulate in retrieved polyethylene components. It is important that this technique can also distinguish between third-body particulate and normal inclusions in the polyethylene such as calcium stearate, based on the distinct morphology and elemental composition of each material. Therefore, the ability to distinguish third-body particulate from calcium stearate inclusions is essential in gaining a better understanding of the contributing factors associated with coating separation and accelerated wear observed in clinically retrieved polyethylene components. PMID- 10713560 TI - Effect of fabrication method and resin type on performance of tibial bearings. AB - Polyethylene has been used successfully for more than 30 years as an orthopedic bearing material. During this time, several polyethylene resins and fabrication methods have been used to produce bearings. Some bearings fail prematurely due to fatigue, which has been linked to oxidation and degradation of mechanical properties resulting from gamma sterilization in air. Fabrication method and/or resin have been hypothesized to govern whether oxidative degradation occurs in gamma-sterilized bearings. This study evaluates the effect of fabrication (machining/direct compression molding) and resin type on oxidation and the resulting mechanical properties for a large series of never-implanted bearings. While many molded bearings studied exhibit lower oxidation than machined bearings, fabrication method is not a significant predictor of oxidation. Resin type and shelf-age are found to be significant predictors of oxidation. Bearings fabricated from Himont 1900 exhibit lower oxidation than those from GUR 415/412 at comparable times after gamma in air. However, Himont 1900 bearings lose strength and elongation at lower oxidation levels than GUR 415/412 bearings. But since Himont 1900 oxidizes more slowly, Himont 1900 bearings retain mechanical properties for longer shelf times than comparable GUR 415/412 bearings. These effects are seen in retrievals as well. PMID- 10713561 TI - Submicron-size particles of ultrahigh molecular weight polyethylene produced via nonsolvent and temperature-induced crystallization. AB - Submicrometer size particles of ultrahigh molecular weight polyethylene (UHMWPE) were produced by crystallization from dilute (0.1-1.0 wt % of UHMWPE) solvent/nonsolvent emulsions. The procedure consisted of mixing a hot solution of UHMWPE in decalin or decane with a nonsolvent (tetraglyme) at approximately 160 degrees C, followed by rapid cooling of the mixture to zero or subzero temperatures. The rapid cooling causes microphase separation between the two liquids, resulting in the formation of an emulsion, which consists of microdroplets of the supercooled UHMWPE solution dispersed in tetraglyme. The consequent crystallization of the polymer in the microdroplets produces a suspension of fine crystals of UHMWPE, which can easily be isolated. The particles were characterized using scanning electron microscopy, differential scanning calorimetry, and Raman spectroscopy. Their degree of crystallinity is between that of GUR 1050 original (powder) and processed (molded) polymer. By changing the polymer concentration, solvent to nonsolvent ratio, and temperature, the size (from 0.1-1.0 microm) and shape (spheroids or rods) of the particles can be controlled. These particles may be used for immunochemical investigations and the study of the influence of UHMWPE wear debris on cell response. PMID- 10713562 TI - In vitro model for frontal sinus obliteration with bioactive glass S53P4. AB - An in vitro model was used to investigate the behavior of a massive frontal sinus obliteration with bioactive glass S53P4 (BG) for clinical purposes. Two sizes of granules (0.63-0.8 mm or 0.8-1.0 mm) in 16 separate BG amounts, weight 25 g, were tested both in simulated body fluid (SBF) and a buffer containing trishydroxymethyl aminomethane citric acid (TRIS-c.a) in standard conditions. The dissolution of silicon (Si) and phosphate (P) was detected with direct current plasma atom emission spectroscopy (DCP-AES) monthly up to 6 months. The BG masses were scanned by computer tomography (CT) and the scans analyzed by Region of Interest (ROI) technique. Calcium phosphate (CaP)- and silica (Si)-gel-layers were studied by scanning electron microscopy (SEM) at 1, 3, and 6 months. Cumulative loss of Si and P was stronger in TRIS -c.a than in SBF (p < 0.0001), and it was higher with smaller than with larger granules in both solutions (p < 0.0001). This was shown correspondingly by the decrease in Hounsfield units (HU) by ROI analysis (p < 0.0001). In SBF-soaked BG masses, the CaP-layer occurred on the uppermost granules, and in TRIS-c.a at 3-6 months, on the granules in the center and lower parts. The decrease of HU seems to reveal indirectly the resorption of BG. PMID- 10713563 TI - Missing osteoconductive effect of a resorbable PEO/PBT copolymer in human bone defects: a clinically relevant pilot study with contrary results to previous animal studies. AB - PEO/PBT 70/30 (POLYACTIVE(R) 70/30), a degradable porous copolymer with elastic properties, was found to be osteoconductive in many animal studies. The aim of this study was to determine the osteoconductive effect in a human paired control iliac defect model. In seven patients undergoing anterior spinal interbody fusion surgery, two bicortical iliac defects for autograft harvesting were created. The defect size was identical for both defects measuring about 40 x 15 mm (group I). One defect was filled with the degradable implants, whereas the remaining one was left untreated as a control. The defect site for treatment was chosen randomly. In three further patients, only one defect measuring about 40 x 35 mm was created (group II). All patients were examined clinically and radiologically by spiral-CT after 1, 6, 12, 24, and 52 weeks. Three-dimensional reconstructions as well as CT volumetric measurements using 1 mm sections were used as evaluation methods. In group I, a two-tailed paired t-test showed that the treated defects had significantly less formation of new bone than the untreated ones (p < 0.05 after 12 weeks, p < 0.01 after 52 weeks). Also, in group II, not much bone ingrowth could be observed. The histological evaluation of one patient in group I revealed no bone within the pores, and a fibrous layer between bone and implant was always present. Therefore, PEO/PBT 70/30 cannot be recommended as a bone substitute for clinical use. Differences in bone regeneration between humans and certain animal species as well as inapplicable defect models in previous animal studies are discussed as possible reasons for the failure. PMID- 10713564 TI - Hydroxyapatite granules interposed at bone-cement interface in total hip replacements: histological study of retrieved specimens. AB - The effect of hydroxyapatite (HA) granules interposed between bone and polymethylmethacrylate (PMMA) bone cement in total hip replacement was histologically evaluated. The technique consisted of smearing 2-5 g of HA granules (straight phi = 100-300 microm) onto the bone surface just before cementing. Four specimens containing well-fixed bone-cement interface were retrieved at 1, 2, 6, and 10 years postoperatively and examined with back scattered electron microscopy and light microscopy. The majority of HA granules were incorporated into remodeled trabeculae, and highly convoluted bone-cement interface was maintained up to 10 years. The presence of active remodeling in the adjacent bone was observed. There were no significant inflammatory or foreign body reactions against interposed HA granules. In one specimen retrieved from a patient with rheumatoid arthritis, bone formation around HA granules was limited after 1 year. These results have provided histological evidence for the significantly reduced incidence of radiolucent lines in total hip replacement with this cementing technique, reported elsewhere. PMID- 10713565 TI - The piezoelectric cochlear implant: concept, feasibility, challenges, and issues. AB - A better understanding of the fundamental phenomena occurring in both the healthy and the artificially stimulated cochlea will greatly aid in the engineering of more effective cochlear implant devices and will, in general, enhance mankind's knowledge of inner ear function. This study was initiated to probe the feasibility of use of artificial piezoelectric transducer devices, both for the understanding of cochlear phenomena and as a possible cochlear implant. Aspects of feasibility of such an implant, the issues involved, the materials science challenges that need to be overcome to fabricate such a device, and results from initial in vivo experiments are discussed. PMID- 10713566 TI - A simple, nondestructive assay for bound hyaluronan. AB - A simple, convenient, nondestructive method is described for the quantitative determination of bound hyaluronan. The method is based on the binding of the cationic dye Toluidine Blue O to the D-glucuronate component of the hyaluronan repeat disaccharide. Quantification is accomplished without interference by the dye's metachromatic properties. The method is easily adapted to hyaluronan coated medical devices and should be useful to developers and manufacturers of such devices and coatings. PMID- 10713567 TI - Role of NMDA receptors in adult primate cortical somatosensory plasticity. AB - We have previously shown that most of the reorganization that typically follows median nerve transection in adult squirrel monkeys is dependent on normally functioning N-methyl-D-aspartate (NMDA) receptors. Here, we have evaluated two additional hypotheses: (1) is the immediate "unmasking" found after median nerve transection NMDA receptor-dependent? and (2) are NMDA receptors necessary for both the initiation and maintenance of the second phase of reorganizational changes, or only the former? To address these issues, we implanted osmotic minipumps subcutaneously to deliver an NMDA receptor antagonist (3-((+/-)-2- carboxypiperazin-4-yl)propyl-1-phosphonic acid, CPP) systemically either before examining the immediate effects of median nerve transection, or after reorganization had presumably occurred. For the first set of experiments, NMDA receptor blockade was initiated either 1 or 4 weeks prior to multi-unit mapping in area 3b followed by transection of the median nerve and remapping of the cortex. In the second set of experiments, median nerve transection was followed 4 weeks later by either 1 or 4 weeks of NMDA receptor blockade prior to terminal mapping. We report that the immediate unmasking of new receptive fields after acute nerve injury is not prevented by NMDA receptor blockade; nor are completely reorganized cortical maps dependent upon NMDA receptors for their maintenance. We conclude that the immediate changes in cortical topography are not due to an NMDA receptor-dependent mechanism, but more likely due to release from tonic inhibition. Furthermore, the later phase of reorganization, as for some forms of hippocampal long-term potentiation (LTP), is dependent on normally functioning NMDA receptors for its initiation, but not for its maintenance. PMID- 10713568 TI - Comparative study of the apical ganglion in planktotrophic caenogastropod larvae: ultrastructure and immunoreactivity to serotonin. AB - Previous research suggests that a major role of the apical ganglion (also called the apical or cephalic sensory organ) in gastropod larvae is detection and integration of sensory information and relay of motor signals to effectors in the velum. However, the relative impact of ancestry versus velum size and life history on characteristics of the apical ganglion is unresolved. We address this issue by contributing data on the apical ganglion and overlying epidermis in planktotrophic larvae of four caenogastropod species (Euspira [Polinices] lewisii, Lacuna vincta, Trichotropis cancellata, and Amphissa versicolor) derived from light microscopy, scanning and transmission electron microscopy, and immunohistochemical localization of serotonin-like antigenicity. Ultrastructure of the apical ganglion is similar in these caenogastropods, and the basic plan corresponds to previous descriptions of the apical ganglion in planktotrophic opisthobranch larvae (subgroup of Heterobranchia). The only identified structural feature that is unique to all these caenogastropods, relative to opisthobranchs, is modified ciliary axonemes for the ampullary cells, a distinctive type of sensory neuron. Like opisthobranch larvae, caenogastropod larvae have serotonin immunoreactive neurons within the apical ganglion; the number ranges from three to six, but a lateral pair of serotonergic, nonsensory neurons is common to all species. The pattern of serotonergic neurons in E. lewisii, which develops large, subdivided velar lobes, is the same as that of opisthobranch larvae, which have a relatively small, unelaborated velum. These and other data suggest that common ancestry is a major determinant of overall structural design for the apical ganglion in caenogastropods and heterobranchs, which are sister groups within the Gastropoda. Velum size and life history strategy may account for some, but not all, cases of interspecific differences in the serotonergic component. PMID- 10713569 TI - Differentiation of mitral cell dendrites in the developing main olfactory bulbs of normal and naris-occluded rats. AB - The morphological differentiation of mitral cell dendrites during embryonic and early postnatal development was examined in the main olfactory bulb of rats to determine a possible role of afferent activity in the development of the dendrites. Mitral cells and olfactory nerve fibers were labeled with 1,1' dioctadecyl-3,3,3', 3'-tetramethylindocarbocyanine perchlorate (DiI) and fluorescein-conjugated lectin (Ulex europeus agglutinin-I), respectively. Morphogenesis of mitral cell dendrites proceeded as previously described (Malun and Brunjes [1996] J. Comp. Neurol. 368:1-16); that is, undifferentiated dendrites with radial orientation were transformed into a single primary dendrite having a glomerular tuft and secondary dendrites extending tangentially into the external plexiform layer. Quantitative examinations in both pre- and postnatal rats revealed that the differentiation of primary dendrites, including tuft formation, increases in diameter and decreases in branching, started before birth, whereas differentiated secondary dendrites were only observed in postnatal animals. Mitral cells with more than two primary dendrites were found after embryonic day 21. The proportion of the mitral cells with differentiated dendrites increased postnatally. At postnatal day 10, almost all mitral cells had fully differentiated dendrites, and mitral cells with multiple primary dendrites were no longer seen. No significant change was found during development in the number of stem dendrites that arose directly from the cell body. Unilateral naris occlusion started on postnatal day 1 retarded differentiation of primary and secondary dendrites, and increased the proportion of mitral cells with multiple primary dendrites. These finding revealed that differentiation of mitral cell primary dendrites precedes that of secondary dendrites, and suggested that the differentiation of secondary dendrites proceeds in an activity-dependent manner. PMID- 10713570 TI - Altered mRNA expression for brain-derived neurotrophic factor and type II calcium/calmodulin-dependent protein kinase in the hippocampus of patients with intractable temporal lobe epilepsy. AB - The expression of brain-derived neurotrophic factor and the alpha subunit of calcium/calmodulin-dependent protein kinase II mRNA in hippocampi obtained during surgical resections for intractable temporal lobe epilepsy were examined. Both calcium/calmodulin-dependent protein kinase II and brain-derived neurotrophic factor are localized heavily within the hippocampus and have been implicated in regulating hippocampal activity (Kang and Schuman [1995] Science 267:1658-1662; Suzuki [1994] Intl J Biochem 26:735-744). Also, the autocrine and paracrine actions of brain-derived neurotrophic factor within the central nervous system make it a likely candidate for mediating morphologic changes typically seen in the epileptic hippocampus. Quantitative assessments of mRNA levels in epileptic hippocampi relative to autopsy controls were made by using normalized densitometric analysis of in situ hybridization. In addition, correlations between clinical data and mRNA levels were studied. Relative to autopsy control tissue, decreased hybridization to mRNA of the alpha subunit of calcium/calmodulin-dependent protein kinase II and increased hybridization to brain-derived neurotrophic factor mRNA were found throughout the granule cells of the epileptic hippocampus. There also was a significant negative correlation between the duration of epilepsy and the expression of mRNA for brain-derived neurotrophic factor. These results are similar qualitatively to those found in animal models of epilepsy and suggest that chronic seizure activity in humans leads to persistent alterations in gene expression. Furthermore, these alterations in gene expression may play a role in the etiology of the epileptic condition. PMID- 10713571 TI - Changes in efferent and afferent connectivity in rats with induced cerebrocortical microgyria. AB - Freezing injury to the cortical plate at postnatal day (P) 1 initiates a cascade of events that ultimately result in a focal neocortical malformation resembling human 4-layered microgyria. This malformation has been associated with widespread changes in neocortical and thalamic architecture and physiology. It was hypothesized that at least some of these alterations could result from connectional reorganization following early injury. The current experiment was designed to delineate the efferent and afferent connections between the cerebral hemispheres and between the cortex and thalamus of rats with induced cerebrocortical microgyria. Microgyria were induced in the parietal cortex of rats by freezing injury on postnatal day 1. In adulthood, injections of biotinylated dextran amine were made either in the microgyric cortex, in homologous regions of the opposite hemisphere, or in ipsilateral ventrobasal complex of the thalamus. Appropriately directed connections to homotopic areas were seen in some but not all microgyric rats. In addition, heterotopic projections to frontal and secondary sensorimotor cortices were noted. Projections from homotopic regions in the hemisphere opposite to the malformation terminated most often in the medial portions of the microgyrus or avoided it entirely. There were almost no thalamocortical or corticothalamic projections between the ventrobasal complex and the microgyrus itself, although a dense plexus of thalamocortical fibers was often noted at the border between the malformed and normal cortex. These connectional changes may help explain disturbances in architecture, physiology, and behavior associated with these focal malformations. PMID- 10713572 TI - Development of functional connections between thalamic fibres and the visual cortex of the wallaby revealed by current source density analysis in vivo. AB - Functional development of thalamic input to the cortex in anaesthetised wallaby pouch young between postnatal day 25 (P25) and P153 has been studied by electrical stimulation of the optic nerve, current source density (CSD) analysis, and histologic identification of recording sites. Conduction in the optic nerve was recorded prior to P39, by which time responses from the superior colliculus appeared. No evoked potential of cortical origin was recorded until P46, even though thalamic fibres grew into the cortical plate from P15. The first cortical synaptic responses were recorded at the margin of the subplate and the developing cortical plate, where cells that later comprise the adult layer 6 settle. At about P66, an additional short-latency, superficial response appeared, coinciding with the formation of layer 4. The deep response was retained in layer 6. Evoked activity in the presumed layer 4 was found progressively deeper in the cortex over the next few weeks, which would be expected from the addition of layer 3 above it. By P113, a new sink was added superficial in the cortex. Thalamocortical connections follow the same deep-to-superficial order in development as the cellular layers of the cortex. PMID- 10713573 TI - Morphological and electrophysiological characteristics of layer V neurons of the rat medial entorhinal cortex. AB - This study aimed to characterize the morphological and electrophysiological properties of neurons in layer V of the entorhinal cortex in the rat brain. Using the in vitro slice preparation and sharp electrode techniques, we recorded from layer V neurons located in the medial entorhinal cortex. Recorded cells were also labeled with biocytin. Based on morphological criteria, layer V of the entorhinal cortex is comprised of three categories of neurons: pyramidal cells, horizontal cells, and polymorphic cells. Horizontal cells could be easily distinguished from the pyramidal cells because the bulk of their dendritic plexus extended horizontally within layer V. Polymorphic cells vary in size and shape. Interestingly, they typically do not have apical dendrites, and some of them have dendrites that extend into the subiculum. Based on electrophysiological criteria alone, it was not possible to unequivocally distinguish the morphological cell types because they were somewhat heterogeneous with respect to several parameters including inward rectification, spike-frequency adaptation, and intrinsic oscillations. Nevertheless, although most horizontal cells displayed time dependent inward rectification, most pyramidal cells displayed fast inward rectification exclusively. None of the entorhinal cortex layer V cells displayed oscillatory activity like that of neocortical layer V "bursting" cells, although neurons from all groups displayed rhythmic subthreshold membrane potential oscillations. In summary, we have found that layer V of the rat medial entorhinal cortex consists of three morphologically distinct neuronal subtypes that cannot be clearly distinguished from each other by traditional electrophysiological measures. PMID- 10713574 TI - Localization of transitin mRNA, a nestin-like intermediate filament family member, in chicken radial glia processes. AB - We have examined the gene expression of two radial glia intermediate filament proteins, transitin and vimentin, in the developing chick CNS. Despite global similarities in their mRNA distributions, marked regional differences are observed. Most notably, we show that transitin mRNA is localized along radial glial processes and is localized to radial glia endfeet, whereas vimentin mRNA is not localized in radial glia. Localization of transitin mRNA is best shown in the diencephalic radial glia, as well as cerebellar Bergmann glia. In addition, in the early embryonic optic tectum, telencephalon, and retina, transitin mRNA is highly localized to radial glia endfeet, which is suggestive of its transport in these cells. These in vivo demonstrations of transitin mRNA localization are confirmed by in situ hybridization analysis of cultured chick brain radial glia, which demonstrates the presence of granular staining for transitin mRNA in glial processes. Transitin mRNA distribution in developing muscle also shows a highly regulated expression pattern, especially along the Z-lines of myofibrils. As further support for the transport and localization of transitin mRNA in radial glia and muscle, we have identified a consensus RNA transport signal in transitin mRNA that is absent from vimentin. These data suggest that the local regulation of transitin protein synthesis may contribute to its function as an intermediate filament protein in radial glia. PMID- 10713576 TI - The elderly criminal. PMID- 10713575 TI - Pseudorabies virus tracing of neural pathways between the uterine cervix and CNS: effects of survival time, estrogen treatment, rhizotomy, and pelvic nerve transection. AB - The transneuronal tracer, pseudorabies virus (PRV), was used to identify pathways from the uterine cervix which may be involved in induction of analgesia and abbreviation of estrus by vaginocervical stimulation. In Experiment I, PRV immunoreactivity (PRV-IR) in brain and spinal cord was examined 3-5 days after injection into the cervix of ovariectomized (OVX) female rats given estrogen (E) or control treatments. No differences in viral labeling were observed between OVX and OVX+E females at any time. PRV-infected cells were observed to increase as a function of time and at progressively higher CNS levels. PRV-IR neurons were first observed on day 3 post-infection at L6 in the SPN. Increased labeling was observed at day 4 in the SPN and the DGC at L6 and S1 spinal segments. Dorsal horn neurons showed PRV-IR by 4.5 days. Five days post-infection, labeling was seen in the IML and lamina X in T12-L1 segments, and in medullary raphe, A5, nPGi, nGi, DMV, lateral reticular, Barrington's nuclei, and in the midbrain PAG. In Experiment II, the effects of bilateral L6 dorsal root rhizotomy (RH) combined with unilateral (UPx) or bilateral (BPx) pelvic nerve transection on PRV infectivity were examined 5 days after infection. Despite reductions in substance P labeling in the dorsal horn following RH, PRV-IR neurons persisted in this area. In RH+UPx females, labeling persisted bilaterally in the SPN and DGC at L6. RH+BPx almost completely eliminated the PRV labeling in L6 and S1. Horizontal sections showed distinct patterns of infectivity within the IML of thoracolumbar and SPN of lumbosacral segments consistent with infection in the hypogastric and pelvic nerves, respectively. Our data indicate that retrograde transport of PRV occurs via the hypogastric and pelvic nerves after injection of the virus into the uterine cervix. Furthermore, significant intraspinal processing is likely to occur between thoracolumbar and lumbosacral levels in the modulation of reproductive tract function. PMID- 10713577 TI - How useful are cholinesterase inhibitors in the treatment of Alzheimer's disease? A number needed to treat analysis. AB - OBJECTIVES: To report on a Numbers Needed to Treat (NNT) analysis of the literature identified through a systematic review of trials of cholinesterase inhibitors in Alzheimer's Disease. DESIGN: Search of Medline (1966-1998), EMBASE (1994-1999) and Psychlit (1974-1998) using the keywords cholinesterase and placebo dementia. SETTINGS: Double-blind, randomised, placebo-controlled trials. SUBJECTS: People with Alzheimer's Disease. INTERVENTIONS: Drug trials of acetylcholinesterase inhibitors (ChIs). Main outcome measuresAlzheimer's Disease Assessment Scale (Cognitive subscale), Clinician's Interview Based Impression of Change Plus, Mini Mental State Examination, Progressive Deterioration Scale. RESULTS: Small numbers of patients (in most cases between 3 and 7) need to be treated with appropriate dosages of ChIs to ameliorate the clinical symptoms, or postpone deterioration in one of them. CONCLUSIONS: These small NNTs suggest that, despite their expense, the cholinesterase inhibitors have a valuable place in the current clinical management of AD. PMID- 10713578 TI - The economic and social cost of dementia in Ireland. AB - The economic and social burden of dementia on society is the value of all the resources used to prevent, diagnose, treat, and generally cope with the illness. There is increasing pressure to define the cost components of dementia with a view to improving resource allocation and accountability in this area in the future. We have assessed the overall resource implications of dementia in Ireland. Six main areas are covered in the cost analysis as follows: mortality and life years lost, in-patient acute care, in-patient psychiatric care, residential long-stay care, family care, and primary and social care in the community. While the results indicate that the baseline cost of illness associated with dementia is substantial at just under IR pound250 million, the most important aspect of the work is the distribution of the burden. The critical role of carers in maintaining people with dementia in their own home is reflected in the results showing that family care accounts for almost 50% of the overall resource burden, based on an opportunity cost valuation of carer time. PMID- 10713579 TI - Prevalence of dementia in centenarians. AB - Above age 65, the prevalence of dementia rises exponentially from 1 to 15% at age 85. Despite many studies concerning dementia, little is known about the prevalence of dementia in the 'oldest old'. Whether the prevalence levels off around age 95 is yet unanswered. This question is important because it addresses whether dementia is an inevitable consequence of ageing or a disorder occurring within a specific age range. All 17 persons aged 100 or more in three Dutch towns with 250 000 inhabitants were examined by means of cognitive tests, informant questionnaires, clinical interviews and anamneses. Fifteen out of 17 Dutch centenarians in a complete population sample of 250 000 were found to be demented. Two could not be examined. PMID- 10713580 TI - Vitamin B12 deficiency in dementia and cognitive impairment: the effects of treatment on neuropsychological function. AB - BACKGROUND: Vitamin B12 assay is part of the routine investigation of dementia, although few studies have investigated the effects of treatment on cognition. We examined the effects of B12 treatment on neuropsychological function and disease progression in patients presenting with dementia or cognitive impairment. METHODS: From 1432 patients who were assessed at the Bristol Memory Disorders Clinic, 125 patients with low serum B12 were identified. Sixty-six patients presenting with dementia, and 22 with cognitive impairment were seen for a second assessment after treatment. Changes in neuropsychological test scores were compared with those of patients with normal serum B12, matched by age and diagnosis. RESULTS: The majority of patients with low serum B12 had normal Hb and MCV values. We found no cases of reversible B12 deficiency dementia. The B12 treatment patients who presented with dementia showed no significant improvement, and no less deterioration, in their neuropsychological function than their matched group. However, a treatment effect was demonstrated among the patients presenting with cognitive impairment. These improved significantly compared to matched patients on the verbal fluency test (p<0.01). CONCLUSION: All patients with cognitive impairment should be investigated for B12 deficiency. Vitamin B12 treatment may improve frontal lobe and language function in patients with cognitive impairment, but rarely reverses dementia. PMID- 10713581 TI - Diagnostic performance of two mental status tests in the older chinese: influence of education and age on cut-off values. AB - AIMS: To (1) establish the clinical usefulness of the 10-item Abbreviated Mental Test (AMT) and the 18-item Chinese Mini-Mental Status Examination (CMMSE) for detecting cognitive impairment associated with dementia in the elderly Chinese; (2) determine how the tests' optimal cut-off scores varied with the patients' educational level and age; and (3) evaluate which was the more accurate test. METHODS: 151 cognitively-healthy, community dwelling elderly Chinese subjects and 95 elderly Chinese outpatients with dementia were administered the AMT and CMMSE. Receiver-Operating Characteristic (ROC) analysis was used to determine the tests' optimal cut-off scores for each of the education-by-age subgroups and their areas under-the-curve were compared non-parametrically to evaluate which test was more accurate. RESULTS: Both the AMT and CMMSE could identify cognitive impairment accurately, but higher cut-off values were necessary for the younger and more educated cohort, while lower values were adequate for the older and less educated subgroup. The AMT appeared to reach a ceiling effect in the more educated categories. The diagnostic accuracies of the two instruments were statistically equivalent; there was a trend, however, for the CMMSE to be performing better in the more educated subgroups. CONCLUSIONS: To maximise the diagnostic efficiency of these two clinically useful mental status tests, it is important to adjust their cut-off scores for the patients' education and age. Though no clear superiority of either instrument was established in this study, we recommend the AMT for patients with 0-6 years of education, whereas for those with greater levels of literacy, we think it better to administer the CMMSE. PMID- 10713582 TI - Lack of adverse pharmacodynamic drug interactions with rivastigmine and twenty two classes of medications. AB - Alzheimer's disease (AD) is often associated with multiple comorbidities and subsequent polypharmacy. Treatment of AD with acetylcholinesterase (AChE) inhibitors can carry a risk of drug interaction with multiple medications often prescribed for other co-existing illnesses. Rivastigmine is an AChE inhibitor that is enzymatically cleaved by AChE, minimally metabolized by cytochrome P450 enzymes, has low protein binding, has a short plasma half-life, and a relatively short duration of action. Such properties make it ideal for use in this patient population. A pharmacodynamic analysis of rivastigmine administered concomitantly with other medications (22 different therapeutic classes) did not reveal any significant pattern of increase in adverse events that would indicate a drug interaction. In summary, rivastigmine was well tolerated and safely administered to a population receiving multiple medications for 'real-world' comorbidities. PMID- 10713583 TI - Physical morbidity in elderly psychiatric inpatients: prevalence and possible relations between the major mental disorders and physical illness. AB - BACKGROUND: This study examines the prevalence of physical morbidity in elderly psychiatric inpatients and the possible relationships between major psychiatric disorders (organic mental disorders, schizophrenic and mood disorders) and physical illnesses. The clinical implications of such relationships are discussed. METHOD: Data were obtained from two old age psychiatry wards over a six month period. Seventy-nine subjects were studied and information was obtained from their medical files. Demographic characteristics, psychiatric diagnosis, number of physical illnesses and number of body systems affected were collected. Analysis of variance (ANOVA) was used to compare the psychiatric groups on continuous outcome data and chi(2) test to compare psychiatric groups on categorical data. RESULTS: Seventy-five per cent of subjects had at least one physical illness. The number of medical illnesses was independent from the psychiatric disorder. Subjects with mood disorders, and especially depression, were more likely to suffer from hypertension, diabetes and cardiovascular illnesses than subjects with schizophrenic or organic disorders. Subjects with organic disorders had the lowest prevalence of endocrine disease and diabetes. CONCLUSIONS: It was concluded the link between mood disorders (depression), cardiovascular diseases and hypertension could be of a 'cause/effect' type or are the results of a survivor effect. The high prevalence of physical morbidity has implications for training and continuing professional development of those in Old Age Psychiatry Services. It should also be taken into consideration when the location of services is being decided. PMID- 10713584 TI - The elderly, dementia, aggression and risk assessment. AB - There are no clear comprehensive guidelines on risk assessment with the elderly. This paper reviews the literature relevant to the subject in conjunction with the case history of a patient with dementia who committed homicide whilst in residential care. Suggestions concerning factors to be taken into consideration during risk assessment are made. PMID- 10713585 TI - Association of serum AACT levels and AACT signal polymorphism with late-onset Alzheimer's disease in Northern Ireland. AB - alpha1-antichymotrypsin (AACT) is a serine protease inhibitor that has been associated with amyloid plaques in the brains of patients with Alzheimer's disease (AD). It has been reported that AACT serum levels are higher in AD patients than in age and sex matched controls. In addition, polymorphisms in the signal peptide and 5' of the AACT gene have been reported to increase the risk of developing AD. Serum AACT has also been suggested to be associated with cognitive decline in elderly subjects. Our objective was to investigate whether a relationship existed between serum AACT levels, AACT genotypes and risk for AD in a case control association study using 108 clinically well defined late onset AD cases and 108 age and sex matched controls from Northern Ireland. We also wished to determine whether higher serum AACT affected levels of cognition as had been previously reported. Serum AACT levels were found to be significantly raised in cases compared to controls (t=3.8, df=209, p<0.001). However, we detected no relationship between serum AACT levels and cognitive decline. We report allelic association of the AACT signal polymorphism with AD (chi(2)=3.70, df=1, p=0.04) but we failed to show any correlation between AACT serum levels and genotype. PMID- 10713586 TI - Three years survival in patients with a clinical diagnosis of dementia with Lewy bodies. AB - The majority of information available on the prognosis of dementia with Lewy bodies (DLB) is based on retrospective data from autopsy series, which are subject to selection bias due to the specific reasons patients are referred for post-mortem studies. The earlier studies comparing DLB patients with patients with Alzheimer's disease (AD) suggest that the mean duration of illness is shorter in DLB patients than in patients with AD. However, more recent studies have not observed significant differences between DLB and AD in age of onset, age at death or duration of illness. We report a 3 year follow-up of a cohort of 114 consecutive patients with dementia, referred to an old age psychiatric service and diagnosed using ICD 10 criteria and the McKeith and Byrne DLB criteria. The case notes of all patients were reviewed to determine the date of onset of symptoms and the date of first presentation to the psychiatric services. Information about outcome was gathered from case notes, hospital files and general practitioner (GP) records. Of the original sample of 114 patients, 106 could be traced. Sixty-four had died and 42 were still alive at the time of the follow-up. Thirty-two patients had originally been assigned the diagnosis of DLB, 43 the diagnosis of AD, 31 vascular dementia and other diagnoses. There were no differences between the AD and DLB group in age at onset, age at death or survival. We have not found any evidence that the prognosis of clinically diagnosed DLB patients is worse than that of patients with a clinical diagnosis of AD. PMID- 10713587 TI - Recent trends in elderly suicide rates in England and Wales. AB - The proportion of elderly in the population is increasing due to increased life expectancy and falling birth rate, and suicide rates increase with age. This study examined the following in England and Wales: (i) recent trends in the elderly suicide rate; (ii) recent trends in method-specific elderly suicide rate; (iii) the relationship between elderly population size and elderly suicide rate in recent years; and (iv) the sex difference in overall and method-specific elderly suicide rate. Data on the various suicide variables were ascertained from the annually published mortality data for years 1985 to 1996. The main findings of this study were: (i) there is a trend towards decline in the overall pure and combined suicide rates for elderly men and women over the 12 year study period; (ii) the main contributors to this decline are suicides due to poisoning by solid and liquid substances (E950), hanging, strangulation and suffocation (E953), drowning (E954), firearms and explosives (E955), and jumping from high places (E957); (iii) the overall pure and combined suicide rates and that for most categories of suicide was higher in men compared to women; and (iv) suicide rates decreased with an increase in the elderly population size. Suicide rates can decline due to a number of reasons. The challenge now is to ensure further decline in suicide rates to meet the Our Healthier Nations target. PMID- 10713588 TI - Donepezil for the treatment of psychosis in dementia with Lewy bodies. PMID- 10713589 TI - SIADH induced by two atypical antipsychotics. PMID- 10713590 TI - Prevalence of abnormal eating amongst elderly people in residential care. PMID- 10713591 TI - Does oestrogen protect the male brain too? A case of oestrogen withdrawal in a man reveals underlying dementia. PMID- 10713592 TI - Current awareness AB - In order to keep subscribers up-to-date with the latest developments in their field, John Wiley & Sons are providing a current awareness service in each issue of the journal. The bibliography contains newly published material in the field of geriatric psychiatry. Each bibliography is divided into 9 sections: 1 Books, Reviews & Symposia; 2 General; 3 Assessment; 4 Epidemiology; 5 Therapy; 6 Care; 7 Dementia; 8 Depression; 9 Psychology. Within each section, articles are listed in alphabetical order with respect to author. If, in the preceding period, no publications are located relevant to any one of these headings, that section will be omitted PMID- 10713593 TI - When is a sexually transmitted viral disease not an 'STD'? PMID- 10713594 TI - Surgeons who test positive for hepatitis C should be transferred to low risk duties. AB - HCV-infected surgeons may transmit HCV to patients during exposure-prone procedures. Current UK policy allows HCV-infected surgeons to practise unrestricted unless they have been associated with transmission, and, at present, surgeons are not routinely tested for HCV infection. The overall outcome for patients exposed to an HCV-infected surgeon may be worse than that for patients exposed to a surgeon who is an HBeAg negative carrier of HBV. However, because most acute HCV infections are anicteric, surgeon associated HCV transmission is less likely to be detected by surveillance. Surgeons have been observed to sustain intraoperative injuries in around 5% of procedures. If surgeons were required to report every intraoperative injury and to be tested to determine whether the patient could have been exposed to HCV, compliant surgeons would be tested for HCV at least annually. Investigations of HBV transmission, however, have suggested that patients may be exposed to a surgeon's blood in as many as 1 in 5 procedures, and that much surgeon to patient transmission is the result of inapparent intraoperative exposure, which the surgeon does not recognise. Thus, requiring surgeons to report intraoperative injuries would not identify all those patients who might have been exposed to HCV, and, since no vaccine or prophylaxis is available, could not prevent infection. A more satisfactory alternative is regular testing of surgeons for HCV, coupled with restriction of practice of those found to be infected. PMID- 10713595 TI - Surgeons who test positive for hepatitis C should not be transferred to low risk duties. AB - Current UK guidelines allow surgeons who are antibody-positive for hepatitis C virus (HCV) to continue performing exposure-prone procedures (EPPs) unless they have been shown to transmit HCV to a patient. Given the low rate of recognised transmission from surgeon to patient, this recommendation is probably reasonable and is consistent with the management of eAg negative carriers of hepatitis B who are also allowed to continue operating. It seems likely that, in the future, pressure will increase to remove surgeons who are HCV-positive (or positive for HBsAg without HBeAg or HIV-positive) from the list of those able to perform EPPs. If implemented, this would require surgeons to be tested at regular intervals for HCV status. There are no data to demonstrate that such an approach would benefit patients overall and the ethical costs would be high because many surgeons will have acquired HCV occupationally. The financial costs would also be high and, in my opinion, would be better deployed by ensuring that existing simple preventative measures are routinely applied to prevent patient-surgeon-patient transmission of all blood-borne viruses. PMID- 10713596 TI - EBNA-1: a protein pivotal to latent infection by Epstein-Barr virus. AB - Epstein-Barr nuclear antigen 1, or EBNA-1, is required for the replication of the EBV genome as an extra-chromosomal element and is a key transcriptional regulator of this virus's latent gene expression. In this review we will describe the salient features of EBNA-1 and oriP, the latent origin of EBV to which EBNA-1 binds site-specifically. EBNA-1's association with host cellular factors, its association with metaphase chromosomes, and its ability to link DNAs to which it binds will be discussed in relation to its roles in replication and transcriptional activation. Although the mechanisms by which EBNA-1 facilitates replication and transcription largely remain enigmatic, EBV's viral replicon has been exploited successfully for applications in gene therapy and in the design of eukaryotic vectors for use in cell culture. PMID- 10713597 TI - The role of the hepatitis C virus glycoproteins in infection. AB - HCV encodes two glycoproteins, E1 and E2, that are believed to be exposed on the surface of virions. These molecules are likely to be involved in viral interactions with the host immune response and responsible for mediating viral entry into target cells. They are obvious major components for prototype vaccine studies. Recently, E2 has been reported to bind to the tetraspan molecule CD81, which represents a putative receptor for HCV. Here, we discuss the role the HCV gps may play during infection, the contribution of E2 gp variation to HCV evasion from the immune response and possible implications of the E2-CD81 interaction for HCV pathogenesis. PMID- 10713598 TI - Influenza A pandemics of the 20th century with special reference to 1918: virology, pathology and epidemiology. AB - Influenza A virus initiated worldwide epidemics (pandemics) in 1918, 1957, 1968 and 1977. A revised calculation of the 1918-1919 pandemic estimates that 40 million persons died and 500 million were infected. The mortalities in 1957 and 1968 were nearly 6 million. Biological and genetic characteristics of the causative agents of the more recent pandemics, have been well studied but little is known about the causative agent of the Great Pandemic in 1918. Genetic characterisation of the 1918 virus has been achieved by sourcing virus RNA from formalin fixed lung samples or by exhuming frozen victims of the outbreak from Arctic regions. Initial analysis of the HA gene from two USA sources indicates a virus related to swine and human influenza with no base insertion at the HA1-HA2 cleavage junction which, at least in avian influenza A, characterises high virulence. Important unanswered questions are whether the 1918 virus spread pantropically perhaps to include the brain and hence cause encephalitis including the later lethargic forms, or whether infection was confined to the respiratory tract. Re-examination of reports of respiratory disease in England and France in 1916-1917 may indicate a non-Spanish origin of the pandemic and a period of 2 years for the virus to be seeded worldwide. In contrast the other two pandemic viruses in 1957 and 1968 appeared to originate in Asia. New anti-neuraminidase drugs in conjunction with amantadine and novel developments with influenza vaccines would be expected to ameliorate the disease in a future pandemic. PMID- 10713599 TI - Antiviral developments. PMID- 10713619 TI - Myeloablative chemotherapy with stem cell rescue for the treatment of primary systemic amyloidosis: a status report. AB - Stem cell transplantation has been incorporated in the treatment of primary systemic amyloidosis for 5 years. Results reported to date suggest that the response rates are substantially better than those for patients treated with low dose traditional melphalan and prednisone chemotherapy. Unexpectedly high mortality rates have, however, been reported with stem cell transplantation, reaching 40% in some series. This unexpectedly high mortality appears to be related to multiorgan failure of tissues infiltrated with amyloid deposits. Deaths have been reported from gastrointestinal tract hemorrhage, gastrointestinal tract perforation, sudden cardiac death, and renal failure. The best patient for transplantation appears to have single organ involvement, an age <55 years, the absence of renal insufficiency, and no symptomatic cardiac dysfunction. Patients eligible to receive stem cell transplant represent a highly selected population, and before conclusions about the efficacy of transplantation are drawn, comparison with a matched control group is necessary. Amyloidosis should be considered an indication for stem cell transplantation in the context of a clinical trial so that results can be compiled and reported for an accurate assessment of response rate, survival, relapse rates and treatment-related toxicities. Bone Marrow Transplantation (2000) 25, 465-470. PMID- 10713620 TI - Randomised studies in acute myeloid leukaemia: the double truth. PMID- 10713621 TI - Dr Frassoni's paper was shown to Dr sue richards, who has commented as follows PMID- 10713622 TI - HLA-A, -B and -DR antigen frequencies of the London Cord Blood Bank units differ from those found in established bone marrow donor registries. AB - Patients requiring allogeneic stem cell transplantation who do not have an HLA matched related donor can sometimes obtain an unrelated donor by searching volunteer registries. The majority of donors in the registries are Caucasoid, which results in a lower probability of a non-Caucasoid patient finding a suitable donor. Cord blood is increasingly used as a source of haematopoietic stem cells for allogeneic bone marrow reconstitution and so far the London Cord Blood Bank has banked almost 3000 cord blood units. An analysis of the first 1500 units banked showed that more than 30% of the London Cord Blood Bank units are derived from UK ethnic minorities compared with only 2% of individuals recruited locally for the British Bone Marrow Registry (BBMR). The HLA types found in these cord blood units reflect their ethnic diversity and include: HLA-A34, A36, A80, B75, B61, B53, B78, B81 and B82. The units stored by the London Cord Blood Bank show an HLA profile which differs considerably from that of locally typed adult volunteers for the BBMR panel and this should help to increase the chances of obtaining acceptably HLA-matched donors for patients from ethnic minorities. Bone Marrow Transplantation (2000) 25, 475-481. PMID- 10713623 TI - Melphalan plus total body irradiation (MEL-TBI) or cyclophosphamide (MEL-CY) as a conditioning regimen with second autotransplant in responding patients with myeloma is inferior compared to historical controls receiving tandem transplants with melphalan alone. AB - The role of more intense conditioning for second transplant was evaluated in myeloma patients achieving at least partial remission (PR) after first transplant with melphalan at 200 mg/m2. Forty-three patients received more intensive conditioning for the second transplant. Nineteen patients received cyclophosphamide 120 mg/kg along with melphalan 200 g/m2 (MEL-CY; group 1) while 24 patients received total body irradiation (1125 cGy) in conjunction with melphalan 140 mg/m2 (MEL-TBI; group 2). Forty-three matched control patients were identified from 450 patients receiving melphalan alone for second transplant (MEL200; group 3). Engraftment and toxicities were comparable among the groups with the exception of increased treatment-related mortality of 8% in group 2 compared to none in groups 1 and 3 (P = 0.07). Despite identical CR rates of 74, 71 and 70%, respectively, in groups 1, 2 and 3 (P = 1.0), event-free survival (median: 27, 15 and 61; P < 0.0001) and overall survival (median: 39, 25 and 76 months; P = 0.003) were significantly decreased in patients receiving more intensive conditioning (groups 1 and 2). Lymphocyte recovery, evaluated as a surrogate for immune recovery, was inferior in more intensively treated patients (groups 1 and 2 compared to group 3). Our findings suggest that more intense conditioning appears to have no benefit in patients responding to their first cycle of high-dose therapy and may even be detrimental in this setting. Bone Marrow Transplantation (2000) 25, 483-487. PMID- 10713625 TI - Scoring system for the prediction of successful peripheral blood stem cell (PBSC) collection in non-Hodgkin's lymphoma (NHL): application in clinical practice. AB - Fifty-six patients with chemosensitive NHL were studied to assess factors affecting mobilization and peripheral blood stem cell (PBSC) collection: all were mobilized with high-dose cyclophosphamide and etoposide and G-CSF 5 microg/kg/day. None of them had bone marrow involvement at the time of mobilization or a history of extended field irradiation. Previous chemotherapy regimens were divided into two groups: moderately myelotoxic chemotherapy (MMC) and highly myelotoxic chemotherapy (HMC). The adequacy of the PBSC harvest was not associated with age, gender, a past history of bone marrow involvement or disease status. In contrast, the number of MMC cycles (n(MMC)) and the number of HMC cycles (n(HMC)) were both significant (P = 0.009 and P = 0.0004, respectively) and were used to compute a score predictive of a successful PBSC harvest: SCORE = n(MMC) + 4 n(HMC). The estimated successful PBSC collection rate was greater than 80% in patients with a score ranging from 0 to 15 and dropped rapidly to below 20% in patients with a score exceeding 25. This scoring system may help to determine the timing of PBSC mobilization in patients with a score below 15 and suggests that new PBSC mobilization procedures should be investigated in other patients. Bone Marrow Transplantation (2000) 25, 495-499. PMID- 10713624 TI - Allogeneic bone marrow transplantation for children with acute leukemia: cytoreduction with fractionated total body irradiation, high-dose etoposide and cyclophosphamide. AB - Marrow-ablative chemo-radiotherapy followed by hematopoietic stem cell rescue from an allogeneic source improves outcomes for children with high-risk acute leukemia. The first effective pre-transplant preparative regimens consisted of high-dose cyclophosphamide (CY) and total body irradiation (TBI). Subsequent attempts have been made to improve leukemia-free survival, by adding other chemotherapy agents to these agents. In previous clinical studies of total body irradiation, etoposide, cyclophosphamide (TBI-VP-16-Cy) in adult allogeneic bone marrow transplantation, there has been a high incidence of severe regimen-related toxicity. In this study, we investigated the safety and efficacy of this combination in 41 children who received TBI (12-14 Gy), VP-16 (30 mg/kg), and CY (60 mg/kg x 2) and then either matched sibling or alternative donor transplants for acute leukemia. There was only one case of fatal regimen-related toxicity. The estimated 3-year event-free survival for patients with early or intermediate stage disease was 68% (53-88%). The estimated event-free survival of patients with advanced disease was 17% (5-59%). TBI-VP16-CY is safe in pediatric transplantation, and it has good efficacy for transplant recipients with less advanced disease. Bone Marrow Transplantation (2000) 25, 489-494. PMID- 10713626 TI - Comparison of marrow and blood cell yields from the same donors in a double blind, randomized study of allogeneic marrow vs blood stem cell transplantation. AB - Forty healthy adult donors underwent marrow (BM) as well as peripheral blood (PBSC) stem cell collections for their HLA-identical adult siblings with hematologic malignancies. BM was harvested on day 1 (target 3 x 108 nucleated cells/kg, 10 microg/kg lenograstim (glycosylated G-CSF) administered on days 2-6, and a single leukapheresis performed on day 6. The blood volume processed was the higher of 200% donor blood volume or 10 liters. The total nucleated cell (TNC) yields from PBSC were 1.1- to 4.3-fold higher than BM (median 7.0 vs 3.1 x 10(8)/kg, P < 0.0001). Although BM contained a higher proportion of CD34+cells (1.3% vs 0.7%, P < 0. 0001) and a comparable proportion of CD3+ cells (median 29% vs 26%, P = 0.4), the absolute numbers of CD34+ and CD3+ cells and their subsets were several times higher in PBSC. There was a poor correlation between BM and PBSC CD34 and TNC numbers, but a significant correlation between BM and PBSC CD3 numbers. Only five of 40 BM harvests contained >/=2 x 10(6) CD34+ cells/kg compared with 35 of 40 PBSC harvests (P < 0.0001). We conclude that the numbers of progenitor and immunocompetent cells in PBSC are several times higher than in BM. It is possible to collect adequate numbers of progenitor cells from blood after lenograstim stimulation more frequently than from marrow, and donors yielding low quantities of progenitor cells from BM usually deliver better quantities from PBSC. Bone Marrow Transplantation (2000) 25, 501-505. PMID- 10713627 TI - Allogeneic peripheral blood hematopoietic stem cell transplantation: guidelines for red blood cell immuno-hematological assessment and transfusion practice.Societe Francaise de Greffe de Moelle. AB - Allogeneic peripheral blood hematopoietic stem cell transplantation (PBSCT) is presently being evaluated in a French randomized study comparing peripheral blood vs bone marrow. Cases of potentially lethal acute hemolysis have recently been reported after allogeneic PBSCT in the presence of a 'minor' ABO incompatibility. Patients were frequently transfused with recipient-compatible and donor incompatible RBC and usually did not receive methotrexate in addition to cyclosporin A for graft-versus-host disease (GVHD) prophylaxis. In order to homogenize immuno-hematological (IH) assessment and transfusion practices within our protocol, we made proposals to 25 allo-transplant French centers on the following aspects: pre-inclusion IH assessment, IH exclusion criteria, transfusion rules, post-transplant IH surveillance and treatment of hemolysis. Analysis of responses to our proposals led to the elaboration of guidelines which were approved and implemented by the French Bone Marrow Transplantation Society (SFGM). Pre-inclusion IH testing includes mandatory detection and titration of anti-RBC allo-Ab, as well as titration of anti-A and anti-B Ab. The presence in the donor of an anti-A (group A or AB recipients), anti-B (group B or AB recipients) Ab with a titer >1/32 or the presence of allo-Ab against Rh, Kell, Fya, Fyb, Jka, Jkb, Ss Ag present on recipient RBC is an exclusion criterion for the protocol. ABO and RhD compatibility of RBC blood products with both HSC donor and recipient is mandatory. A similar compatibility is also required for Rh (other than D) and Kell Ag. If not possible, compatibility of RBC blood products with the HSC donor is mandatory. Lastly, guidelines regarding post transplantation IH follow-up as well as acute hemolysis treatment have been elaborated. The implementation of these guidelines should contribute to enhancing the quality of transfusion practice after PBSCT. Such an approach will be applied to other aspects of transfusion medicine in the setting of HSC transplantation. Bone Marrow Transplantation(2000) 25, 507-512. PMID- 10713628 TI - Unrelated peripheral blood stem cell transplantation with 'megadoses' of purified CD34+ cells in three children with refractory severe aplastic anemia. AB - Three children with refractory severe aplastic anemia were transfused with high numbers of unrelated matched (n = 2) or C-locus haploidentical mismatched (n = 1) CD34-selected peripheral blood stem cells in the absence of an HLA-identical family donor. Two leukaphereses of the donors yielded a median number of 10.1 x 10(10) nucleated cells (range 9.7-15.4) with a median number of 9.89 x 10(8) CD34+ cells (range 7.46-26.1) and a median percentage of CD34+cells of 0.98% (range 0.77-1.7). After positive selection by magnetic cell sorting the patients received a median of 14.3 x 10(6) CD34+ cells/kg (range 11.7-24.3) and of 1.3 x 10(4) CD3+ cells/kg (range 0.57-5.8). Median time to ANC >/=0.5 x 10(9)/l was 7 days (range 7-12) and to platelets >/=20 x 10(9)/l 13 days (range 13-27). Chimerism analysis of peripheral blood after transplantation revealed permanent 100% donor hematopoiesis in all patients. The patient with the C-locus haploidentical mismatch presented with acute GVHD (grade III-IV) of the skin, liver and lower gastrointestinal tract (onset day +40) and died despite intensive immunosuppressive treatment on day +238. The two survivors developed lymphopoietic recovery of B and T lymphocytes within 3 months after transplantation. To our knowledge this experience represents the first report of transplantation with unrelated CD34+ enriched peripheral blood stem cell in children with refractory severe aplastic anemia. Bone Marrow Transplantation (2000) 25, 513-517. PMID- 10713629 TI - A phase II study of two cycles of high-dose chemotherapy with autologous stem cell support in patients with metastatic breast cancer who meet eligibility criteria for a single cycle. AB - Multi-cycle high-dose chemotherapy with autologous stem cell support (HDC-ASCS) may improve the results obtained with single-cycle HDC-ASCS in metastatic breast cancer (MBC). However, the tolerability and efficacy of additional cycles of HDC ASCS in patients selected using standard eligibility criteria for single cycle HDC-ASCS is uncertain. Twenty-nine patients with MBC and a CR or PR to induction chemotherapy were selected by standard institutional eligibility criteria for single-cycle HDC-ASCS. Cycle 1 HDC-ASCS (cyclophosphamide 6 g/m2; mitoxantrone 70 mg/m2; carboplatin 800 mg/m2) was followed by a planned second cycle (etoposide 1.6 g/m2; thiotepa 800 mg/m2; carboplatin 800 mg/m2 modulated by tamoxifen 120 mg/m2/day x 5 days) with a median interval of 3.2 months. CR rate was 20% after induction chemotherapy and 33% and 54% after HDC cycles I and II, respectively. Sixteen patients (55%) failed to complete HDC cycle II within 200 days because of disease progression, toxicity, inadequate stem cell collection, insurance denials or patient choice. Median progression-free survival (PFS) for all 29 patients entered is 301 days from date of HDC cycle I and actuarial PFS at 2 years is 35%. For the 13 patients who received the two cycles of HDC-ASCS, actuarial PFS at 2 years was 54% (P = NS compared to those receiving only one cycle). These data show that a second cycle of full-dose intensity HDC-ASCS may increase the proportion of patients with MBC that achieve CR and may increase PFS. However, a large proportion of patients that complete HDC-ASCS cycle I may fail to proceed to cycle II in a timely fashion. Bone Marrow Transplantation (2000) 25, 519-524. PMID- 10713630 TI - High-dose immunosuppressive therapy with PBPC support in the treatment of poor risk multiple sclerosis. AB - High-dose immunoablative chemotherapy with autologous haematopoietic cell support might be beneficial in the treatment of intractable forms of MS. We mobilised PBPC in 11 patients with secondary progressive MS and finally eight patients were grafted after high-dose BEAM chemotherapy with either in vitro or in vivo T cell depletion. Median EDSS and SNRS scores at the time of inclusion were 6.5 (6.5 7.5) and 56 (44-65), respectively. PBPC mobilisation was safe with no serious adverse effects, and without significant aggravation of disability. One patient improved significantly (by 1.0 point on EDSS) after the mobilisation. Two mobilisation failures were observed. No life-threatening events occurred during the transplantation. All grafted patients, except one, at least stabilised their disability status. One patient improved significantly (by 1.5 points on EDSS), two patients improved slightly (by 0.5 points on EDSS), one patient worsened by 1.0 point on the EDSS in 10 months. Improvement occurred with a delay of 2-4 months. Median EDSS and SNRS of grafted patients at the last follow up were 6.5 (5.5-8.5) and 64 (39-73), respectively with median follow-up of 8.5 months. Further follow-up is needed to determine the disease course after complete immune reconstitution. Bone Marrow Transplantation (2000) 25, 525-531. PMID- 10713631 TI - The role of autologous transplantation in patients with multiple myeloma aged 65 years and over. AB - Autologous stem cell transplantation after high-dose melphalan for the treatment with multiple myeloma has resulted in prolonged progression-free survival and overall survival in patients under 65 years. We have examined the role of autologous transplantation in 17 patients with multiple myeloma over 65 years at our centre using a matched pair analysis with younger patients. The median age of this cohort of patients over 65 years was 67 years (65-74) and their outcome and transplant-related morbidity was compared with 17 younger pair mates with a median age of 55 years (31-64). Sixteen patients received high-dose melphalan, and one received busulphan with autologous stem cell rescue. The high-dose therapy was well tolerated in both elderly patients and the matched pairs, with comparable time to recover neutrophils and platelets. Treatment-related mortality also did not differ significantly in both the groups. Median overall survival of the elderly patients was 3.59 years similar to 3.01 years of the pair mates (P = 0.92). Autologous stem cell transplantation after high-dose melphalan conditioning was equally well tolerated in groups of patients above and below 65 years. There was no difference in relapse rate, OS and myelotoxicity in both the groups. These findings suggest that advanced age should not be an exclusion criterion from autologous transplant programmes. Bone Marrow Transplantation (2000) 25, 533-539. PMID- 10713632 TI - Measurements from normal umbilical cord blood of four lysosomal enzymatic activities: alpha-L-iduronidase (Hurler), galactocerebrosidase (globoid cell leukodystrophy), arylsulfatase A (metachromatic leukodystrophy), arylsulfatase B (Maroteaux-Lamy). AB - Umbilical cord blood (UCB) has received increasing attention as a source of unrelated hematopoietic stem cells for transplantation. Lysosomal diseases have been effectively treated and normal enzymatic activity has occurred subsequent to engraftment using UCB. The use of donor cells with normal amounts of enzyme, rather than those from carriers whose level may be 50% or less, is an obvious goal. The frequency of such heterozygotes varies from 1:10 to 1:140 or lower depending upon the disease at issue. We assayed the levels of lysosomal enzymes in normal UCB in random samples as well as those used for transplantation. We measured the following enzymatic activities: alpha-l-iduronidase (Hurler), galactocerebrosidase (globoid cell leuko- dystrophy) and arylsulfatase A (metachromatic leukodystrophy). For the latter, levels of activity in UCB are comparable to those found in adult blood. In the case of arylsulfatase B (Maroteaux-Lamy) a level lower than adult level was found. An informed choice by the transplanting physician based on the activity of the relevant enzyme in the UCB donor will provide a better opportunity for an improved prognosis for more complete correction of the recipient's primary disease. Bone Marrow Transplantation (2000) 25, 541-544. PMID- 10713633 TI - Successful ribavirin therapy for severe adenovirus hemorrhagic cystitis after allogeneic marrow transplant from close HLA donors rather than distant donors. AB - Intravenous ribavirin was given to nine patients who had developed severe adenovirus-induced hemorrhagic cystitis (AD-HC) which was resistant to conventional therapy or where there was involvement of other organs after allogeneic BMT. Three patients recovered completely from AD-HC, two of whom had been resistant to vidarabine. All three had received sibling BMTs (2 HLA matched, 1 HLA mismatched). Five patients who received BMTs from related (2 HLA mismatched) or unrelated (1 HLA matched, 2 HLA mismatched) showed an improvement in symptoms but had recurrent AD-HC after discontinuation of ribavirin. Improvement in clinical symptoms and termination of virus excretion were well correlated. The last patient who received a mismatched unrelated BMT died during ribavirin therapy. Ribavirin was notably more effective among patients receiving BMTs from siblings in contrast to patients receiving BMTs from alternative donors (<0.05). One patient experienced severe pancytopenia during the second treatment with ribavirin after HC recurrence and recovered after ceasing ribavirin. Thus, ribavirin seems to be very effective for severe AD-HC for some recipients who receive transplants from a genetically close donor. Bone Marrow Transplantation (2000) 25, 545-548. PMID- 10713634 TI - Diagnosis and management of subdural haematoma complicating bone marrow transplantation. AB - Subdural haematoma (SDH) is a known complication of bone marrow transplantation (BMT). A retrospective review of 657 consecutive patients undergoing allogeneic or autologous bone marrow/stem cell transplantation at the Royal Brisbane Hospital between January 1991 and December 1998 is reported. Seventeen cases of subdural haematoma/hygroma were identified (2.6%). Eleven of these (65%) were bilateral. Four required surgical drainage, with two developing re-accumulation of SDH. All cases presented with a headache and eight of these had associated neurological complications. Diagnosis was made predominately by CT scan: however in 25% of cases definitive diagnosis could only be made in MRI studies. An association with intrathecal methorexate-containing conditioning therapy, post lumbar puncture headache, prolonged thrombocytopenia and coagulopathy was noted. In our experience, conservative management with platelet support and correction of coagulopathy achieved resolution of subdural haematoma in most cases, with surgical intervention being reserved for neurological deterioration. Bone Marrow Transplantation (2000) 25, 549-552. PMID- 10713635 TI - Balancing efficacy with cost: antiemetic control in the pediatric stem cell transplant (SCT) population. AB - We studied the practice patterns regarding intravenous (i.v.) ondansetron in children receiving stem cell transplants (SCT) at The Children's Hospital, Boston to identify cost efficiencies. The pharmacy provided information on material and preparation costs on 36 patients who received i.v. ondansetron during 41 SCT in 1995. We examined the effects of frequency, duration, and route of administration on costs. There were 498 days of ondansetron administration costing $49,083 (95$). Tremendous variation existed in frequency and duration with one third receiving i.v. ondansetron once daily, despite published evidence of equivalence of once a day and divided dosing. A switch to once daily i.v. dosing for all patients would have resulted in >/=28% savings. The median duration of use was 11 days (range 1-48); placing a cap for 7-10 days based on the length of SCT conditioning regimens, would produce savings of 48-60% over current use. By shifting administration route from i.v. to oral, a savings of 67% over current use, without a cap on duration, would be realized. Identifying areas for cost savings can be achieved after thorough analysis of all the component costs. We demonstrated that significant cost reductions could be realized by simple changes in prescribing practices without jeopardizing efficacy. These savings are achieved by standardizing dosing interval, route of administration and duration of treatment without altering daily dosage or access to an effective antiemetic. Bone Marrow Transplantation (2000) 25, 553-557. PMID- 10713637 TI - Central nervous system (CNS) tuberculosis following allogeneic stem cell transplantation. AB - Tuberculosis is an uncommon infectious complication after stem cell transplantation. We report a patient who presented with a brain mass, 3 months after pulmonary tuberculosis had been diagnosed and while he was receiving triple antituberculous therapy. He had extensive chronic GVHD. The diagnosis was made after biopsy of the lesion. The cerebral mass was excised, antituberculous treatment was maintained and the patient made a complete neurologic recovery. Six months later, he died of gram-negative septic shock. Mycobacterial infections should be considered in allograft recipients with chronic GVHD and solid lesions in the brain. Bone Marrow Transplantation (2000) 25, 567-569. PMID- 10713636 TI - CD34+ selection of hematopoietic blood cell collections and autotransplantation in lymphoma: overnight storage of cells at 4 degrees C does not affect outcome. AB - The purpose of this study was to investigate whether storing mobilized peripheral blood progenitor cell (PBPC) collections overnight before CD34+ selection may delay platelet count recovery after high-dose chemotherapy and CD34+-enriched PBPC re-infusion. Lymphoma patients underwent PBPC mobilization with cyclophosphamide 4 g/m2 i.v. and G-CSF 10 microg/kg/day subcutaneously. Patients were prospectively randomized to have each PBPC collection enriched for CD34+ cells with the CellPro CEPRATE SC System either immediately or after overnight storage at 4 degrees C. Thirty-four patients were randomized to overnight storage and 34 to immediate processing of PBPC; 15 were excluded from analysis due to tumor progression or inadequate CD34+ cell mobilization. PBPC from 23 patients were stored overnight, while 30 subjects underwent immediate CD34+ selection and cryopreservation. Median yield of CD34+ enrichment was 43.6% in the immediate processing group compared to 39.1% in the overnight storage group (P = 0.339). Neutrophil recovery >500 x 10(9)/l occurred a median of 11 days (range 9-16 days) in the overnight storage group compared to 10.5 days (range 9-21 days) in the immediate processing group (P = 0.421). Median day to platelet transfusion independence was 13 (range 7-43) days in the overnight storage group vs 13.5 (range 8-35) days in those assigned to immediate processing (P = 0.933). We conclude that storage of PBPC overnight at 4 degrees C allows pooling of consecutive-day collections resulting in decreased costs and processing time without compromising neutrophil and platelet engraftment after infusion of CD34+ selected progenitor cells. Bone Marrow Transplantation(2000) 25, 559-566. PMID- 10713638 TI - Constrictive pericarditis post allogeneic bone marrow transplant for Philadelphia positive acute lymphoblastic leukaemia. AB - We describe two cases of severe constrictive pericarditis arising after allogeneic BMT conditioning involving total body irradiation and melphalan to treat Philadelphia-chromosome positive ALL. Both patients required pericardectomy, resulting in marked improvement in ventricular filling. However, a degree of right-sided cardiac failure persisted in both patients secondary to restrictive cardiomyopathy. Constrictive pericarditis has not been previously described after BMT, but has been observed following thoracic radiotherapy for malignancy, usually involving a substantially higher radiation dose. Pericardial constriction and restrictive cardiomyopathy should be considered as causes of breathlessness and/or oedema occurring late after BMT. Bone Marrow Transplantation (2000) 25, 571-573. PMID- 10713639 TI - Criteria for assessing chronic GVHD. PMID- 10713640 TI - Buccal swabs but not mouthwash samples can be used to obtain pretransplant DNA fingerprints from recipients of allogeneic bone marrow transplants. PMID- 10713641 TI - Photosensitivity in lupus. AB - A wide variety of skin conditions may present in patients with lupus erythematosus (LE). These can be broadly divided into three main groups: cutaneous forms of LE ('LE-specific skin disease'), non-specific cutaneous manifestations of SLE ('LE non-specific skin disease') and cutaneous complications of drug treatments for LE. This review examines clinical photosensitivity in LE, a trait most commonly associated with cutaneous forms of LE but which may also manifest in SLE. All humans are photosensitive, developing reddening of the skin if exposed to sufficient ultraviolet radiation (UVR). Therefore we define photosensitivity in clinical practice as an abnormal cutaneous response to UVR. Abnormal photosensitivity in LE may manifest in a number of different forms. The lesions of LE-specific skin disease may be induced or exacerbated by UVR. Patients with LE who are prescribed photosensitizing medications such as thiazide diuretics, neuroleptics and tetracyclines may also develop phototoxic reactions which usually present as easy sunburn. Photosensitivity may also, rarely, manifest as fragile skin and blistering in patients with both LE and porphyria cutanea tarda. Several other photosensitive disorders have been reported in association with LE, including solar urticaria and erythropoetic protoporphyria (EPP), but these appear to be chance associations. Assessment of patients with LE and photosensitivity requires a careful history and examination. Phototesting and photoprovocation tests may be used to demonstrate photosensitivity in some cases, but these are rarely required for diagnosis. Photosensitive patients should be advised about sun avoidance, photoprotection and sunscreen use as a first line treatment. PMID- 10713642 TI - Intestinal pseudo-obstruction in systemic lupus erythematosus: an uncommon but important clinical manifestation. AB - OBJECTIVES: To document intestinal pseudo-obstruction (IpsO) as a recognised clinical manifestation of systemic lupus erythematosus (SLE) and a possible new clinical entity with its apparent association with ureterohydronephrosis. METHODOLOGY: We report six lupus patients who presented with IpsO and review 12 other cases from an English literature search. IpsO is defined as the presence of clinical features suggestive of intestinal obstruction but without organic obstruction, namely absence of bowel sounds, presence of multiple fluid levels on plain abdominal X-rays and exclusion of organic obstruction by imaging or surgical procedure. Other clinical characteristics related to the underlying lupus, serological and histological findings, treatment modalities and outcomes of these patients were reviewed. RESULTS: All 18 patients fulfilled the ACR revised classification criteria for SLE. None showed any clinical features of scleroderma or overlap syndrome. The mean age of onset of IpsO was 29.0 (15-47) y. The female to male ratio was 16:2. Nine patients had IpsO as the initial presentation of their underlying lupus. Coexisting lupus involvement of other organ systems included glomerulonephritis (n=7), thrombocytopenia (n=5) and cerebral lupus (n=3). The serology data and autoantibody profile of some of the previously reported patients were incomplete. In our series, anti-Ro antibody was positive in 5/6 while anti-RNP was found in 1/6 patients only. All our patients had active lupus serology at presentation. 17/18 patients required the use of high dose systemic corticosteroid therapy while one patient responded to topical adrenocorticotrophin hormone treatment. Response was good and was observed early after commencement. Azathioprine was used as maintenance therapy in 6/18 patients with good effects. An apparent association with the presence of bilateral ureterohydronephrosis was found in 12/18 patients. These patients presented with dysuria without positive bacterial culture though features of chronic interstitial cystitis were not invariably found in these patients. CONCLUSION: IpsO is an uncommon but important manifestation of SLE. The underlying pathology is not fully understood but it may be related to immune complex deposition. The finding of coexisting ureterohydronephrosis suggests that there may also be a central smooth muscle motility problem of neuropathic or myogenic pathophysiology which may or may not be secondary to vasculitis. Early recognition and treatment of IpsO in SLE is important. PMID- 10713643 TI - Usefulness of detection of complement activation products in evaluating SLE activity. AB - Complement activation products, such as C1rs-C1inh, specific for the activation of the classical pathway, C3b(Bb)P, specific for the activation of the alternative pathway and SC5b-9, specific for common terminal pathway of the complement cascade, were measured in healthy donors and in patients with clinically active and inactive systemic lupus erythematosus (SLE). Plasma levels of C3b(Bb)P and SC5b-9 were moderately, those of C1rs-C1inhibitor (C1rs-C1inh) were markedly elevated in patients with clinically inactive SLE, compared with healthy controls. The difference between active and inactive stages of the disease was best reflected by C3b(Bb)P plasma concentration (P<0.001), which also showed the highest correlation with the SLEDAI (Rs=0.41 P<0.001) and which was the most useful in distinguishing active and inactive sample pairs as well. The difference between SC5b-9 levels in the active and inactive stages was also significant (P=0.007), while that of C1rs-C1inh did not differ significantly (P=0.136). The correlation of the SLEDAI with SC5b-9 was 0.3 (P=0.015), while with C1rs-C1inh it was 0.21 (P=0. 089). These findings suggest that the measurement of complement activation products, especially that of the alternative pathway, are sensitive markers of the activity of SLE and can be used for clinical purposes. PMID- 10713644 TI - Effect of intravenous cyclophosphamide in systemic lupus erythematosus: relation to lymphocyte subsets and activation markers. AB - Since the mechanism mediating the beneficial effect of intravenous cyclophosphamide (IVCY) in systemic lupus erythematosus (SLE) is unknown, we investigated lymphocyte subsets and markers of activated lymphocytes in patients received IVCY, and compared the results with the effect of steroid pulse. In 55 patients with SLE, 34 patients receiving IVCY [21 cases (61.8%) were responsive] and 25 patients received steroid pulse [21 cases (84.0%) were responsive] (four patients who were resistant to steroid pulse therapy were transferred to IVCY). When the lymphocyte subsets and markers of activated lymphocytes were compared in the responsive and unresponsive group of IVCY, soluble CD4 levels and the ratio of HLA-DP-positive T cells were significantly higher in the unresponsive group. Further, the changes of these markers and costimulatory molecules [LFA-1 (CD11a), ICAM-1 (CD54), CD40 and CD40-ligand (CD154)] were also examined in the responsive patients. The ratio of HLA-DP-positive T cells did not change in the IVCY responsive group, while it decreased in the steroid pulse therapy-responsive group. The ratio of CD11a on T cells increased and CD54 on B cells decreased in the IVCY-responsive group. The ratio of CD154 on T cells increased in the steroid pulse-responsive group, while it decreased in the IVCY-responsive group. These results suggest that the effect of IVCY is different to that of steroid pulse therapy and mainly related to B cell activation, and that these markers may contribute to predict the responsiveness of IVCY. PMID- 10713645 TI - Testing for the antiphospholipid syndrome: importance of IgA anti-beta 2 glycoprotein I. AB - BACKGROUND: Testing for the antiphospholipid syndrome (APS) using anticardiolipin antibodies (aCL) has been problematic. Titers may fluctuate or even become negative. Anti-beta 2-glycoprotein I assays (abeta2-GPI) may be more reliable for diagnosis. METHODS: In a prospective, blinded study over a nine-month period we retested all patients seen for routine follow-up visits in our clinic who had previously been evaluated for aCL-associated illnesses. Patients were stratified into two groups: group A-patients previously positive for aCL; group B-patients previously negative for aCL. Both groups were further classified according to disease severity. Patients were retested for both aCL and abeta2-GPI (isotypes G, M, A for each) using uniform testing standards. RESULTS: 118 patients with previously positive aCL (group A) were retested. Repeat aCL were positive in 52/118 (44%), abeta2-GPI positive in 69/118 (58%) and 82/118 (69.5%) were positive for one or both assays. In patients with serious organ damage (92% with documented APS), 48.6% were aCL positive, 64% positive for abeta2-GPI, and 75.7% were positive for one or both assays. When only one assay was positive, abeta2 GPI was most frequent (P=0.0096). Overall, IgA abeta2-GPI was the most frequent isotype found (60.9%). On retesting of 73 aCL-negative patients (group B), 9/73 (12%) were aCL positive, 27/73 (36%) were abeta2-GPI positive, with 24/73 (32.9%) having isolated abeta2-GPI. Of those positive for abeta2-GPI, IgA abeta2-GPI was present in 74. 1%. Many of these patients had documented APS. CONCLUSION: Based on our data, abeta2-GPI assays are superior to aCL assays for diagnosis of APS. The combined use of both assays enhance positive testing results in up to 75% of patients with APS at any stage of illness. ACL negative patients suspected of having APS should be retested for both abeta2-GPI and aCL. IgA abeta2-GPI appears to be the most important isotype detected. PMID- 10713646 TI - Dermatomyositis and polymyositis associated with the antiphospholipid syndrome-a novel overlap syndrome. AB - OBJECTIVE: As APS (antiphospholipid syndrome) can be either primary or secondary to a wide range of other conditions (such as autoimmune diseases, malignancies, infectious diseases, and drug-induced conditions) the aim of this study was to describe a novel overlap syndrome of APS. METHODS: All patients diagnosed with either PM (polymytosis) or DM (dermatomytosis) who were treated in the Rheumatology Unit, Sheba Medical Center, were followed-up in the past 8 years for the appearance of a clinical manifestation of the APS, and conversely, patients with APS treated by us were clinically assessed for the presence of signs and symptoms of PM and DM. Both conditions were diagnosed according to accepted diagnostic criteria. RESULTS: Three patients were found to have both APS and PM/DM. A patient with PM had transverse myelopathy, a patient with DM had pulmonary embolism, and a patient with PM had recurrent abortions, stroke, livedo reticularis and mitral regurgitation. Both patients with PM had also SLE (systemic lupus erythematosis). CONCLUSIONS: APS can be associated with a wide range of diseases. Future data would reveal which therapy is the best for the association of PM/DM with APS, and determine the pathogenesis and prognosis in patients with this association. PMID- 10713647 TI - Class specific rheumatoid factors and antiphospholipid syndrome in systemic lupus erythematosus. AB - The relationship of rheumatoid factors (RF) with antiphospholipid syndrome (aPLS) and anticardiolipin antibodies (aCL) has rarely been investigated in systemic lupus erythematosus (SLE). We found IgM-RF, IgG-RF, IgA-RF, IgM-aCL, IgG-aCL, IgA aCL, respectively, in 35.4%, 35.4%, 33.8%, 23.1%, 23.1%, 20.0% of 65 SLE patients. Class specific RFs were negatively associated (P<0.05) with IgG-aCL. The frequency of definite or probable aPLS according to Alarcon-Segovia classification criteria was significantly (P<0.05) different (8.7% vs 30.9%) in patients with or without IgG-RF. Among the other clinical features of SLE, we found that patients with IgG-RF, compared to patients lacking this autoantibody, showed a lower frequency (P<0.05) of serositis (21.7% vs 52.4%) and hematologic (52. 2% vs 80.9%) disorders. The levels of IgG-RF and IgM-RF negatively correlated with the number of ARA criteria (P<0.05) but not with the indices of diseases activity or damage. Our study shows that in SLE the presence of RFs are not markers of severity of the disease, but the negative association between IgG RF and IgG-aCL suggests a distinct role of these autoantibodies in the pathology of SLE, whereas the presence of IgG isotype may identify a subset of SLE patients having a lower risk to develop some clinical manifestations such as aPLS. PMID- 10713648 TI - Primary Sjogren's syndrome in men: clinical and immunological characteristics. AB - OBJECTIVE: To determine the clinical and immunological characteristics of primary Sjogren's syndrome (SS) in men from a large series of unselected patients with this condition. METHODS: We studied 223 consecutive patients (204 women and 19 men; mean age at onset 53 y, range 15-87 y, mean disease duration 77 months) with primary SS visited in our units. All these patients fulfilled 4 or more of the diagnostic criteria for SS proposed by the European Community Study Group in 1993. RESULTS: Nineteen (9%) patients were men and they represent the male group described in this paper. Extraglandular manifestations during the course of their disease were present in 10 (53%) of our male patients with primary SS: articular involvement in 4 (21%) patients, interstitial pneumopathy in 3 (16%) and peripheral neuropathy in 2 (11%). ANA were positive in 13 (68%) patients, RF in 5 (31%), anti-Ro/SS-A in 3 (16%) and cryoglobulins in 1/14 (7%). When compared with women, men with primary SS presented a lower prevalence of articular involvement (21 percent; vs 46%, P=0.03, OR 0.32, CI 0.07-0.97). CONCLUSION: Although primary SS is typically a disease of middle-aged women, clinicians should note that it may be diagnosed in male patients. Except for a lower prevalence of articular involvement, we could no find any notable differences in clinical and immunological characteristics between male and female patients with primary SS. PMID- 10713649 TI - A reversible bilateral renal artery stenosis in association with antiphospholipid syndrome. AB - We describe a 26-year-old white female with a history of Raynaud phenomenon, erythema nodosum, polyarthralgias, migraine, vertigo, seizures, transient ischemic attacks, one fetal loss, and false positive VDRL, who developed milk hypertension without overt lupus nephritis. She had positive antinuclear antibodies (ANA) and double-stranded deoxyribonucleic acid (dsDNA) antibodies. The lupus anticoagulant test (LAC) and cardiolipins antibodies (aCL) were positive. She was diagnosed as having a Systemic Lupus Erythematosus-like illness (SLE-like) with 'secondary' antiphospholipid syndrome (APS). Renal spiral computed tomography (CT) with intravenous (IV) contrast showed bilateral renal artery stenosis. Anticoagulation with acenocumarol was started. She became normotensive without antihypertensive drugs five months later. A follow-up renal spiral CT showed complete recanalization of both renal arteries, making thrombosis the more likely culprit pathology in the stenosis. After two years follow up the patient is normotensive. She remains on acenocumarol. PMID- 10713650 TI - Acquired C1q deficiency caused by monoclonal paraproteinaemia. AB - Acquired C1q deficiency secondary to anti C1q auto-antibodies may result in the hypocomplementaemic urticarial vasculitis syndrome and may also be seen in active systemic lupus erythematosus. Some patients with acquired C1 inhibitor deficiency are found to have an underlying malignancy, most commonly lymphoma. We report a case of a 40-year-old man presenting with a lupus-like illness with acquired C1q deficiency secondary to a monoclonal paraprotein in the presence of splenic lymphoma with villous lymphocytes. His clinical symptoms correlated with the presence of the paraprotein. Relapse coincided with a rise in the paraprotein and fall in C1q, C3 and C4. There was no improvement in his clinical condition following combination chemotherapy. He remains on oral prednisolone. PMID- 10713651 TI - Lupus cystitis: a possible additive risk factor for emphysematous cystitis in diabetes mellitus: discussion about one case. AB - Emphysematous cystitis (EC) is a rare condition in which gas-forming organisms are active in the bladder wall and lumen. Most of the cases have been described in patients suffering from diabetes mellitus due to glucosuria and subsequent anaerobic fermentation of glucose. To our knowledge this condition has never been described in association with systemic lupus erythematosus (SLE). We report here the first case of EC during the course of a chronic lupus cystitis (LC) in a woman suffering from SLE and type-I diabetes mellitus. PMID- 10713652 TI - Liver dysfunction due to apoptosis in a patient with systemic lupus erythematosus. AB - We report on a 23-year-old Japanese female with a 13-year history of systemic lupus erythematosus (SLE), and two episodes of deterioration followed by treatment with high dose prednisolone. Although she had been recently treated with prednisolone (12.5 mg daily), her liver function became worse in July 1998. Results of a liver biopsy revealed multi-focal hepatic cell death in a severe fatty liver, without any inflammatory cell invasion. The biopsy also showed a positive TUNEL (Tdt-catalysed DNA nick end labelling) reaction indicating apoptosis. Her liver function recovered rapidly following steroid pulse therapy. Serum soluble Fas ligand (sFasL) was found to be elevated to a concentration of 0.395 ng/ml at the time of liver damage, but was less than 0.03 ng/ml before liver damage and after prednisolone treatment. The liver damage in this case appeared to be involved with apoptosis induced by sFasL. Although hepatitis associated with SLE is rare, apoptosis directly related to elevated sFasL levels might cause this complication. PMID- 10713653 TI - Overwhelming septic cavernous sinus thrombosis in a woman after combination of high-dose steroid and intravenous cyclophosphamide therapy for lupus nephritis. AB - There are many treatment methods for lupus nephritis, including high-dose steroids, pulse methylprednisolone, and cyclophosphamide therapy. In cyclophosphamide therapy, there can be some side effects such as nausea, vomiting, and infection. We report on a case receiving a combination of high dose steroid and intravenous cyclophosphamide. Following this, she developed a fever and a protruding right eye, and septic cavernous sinus thrombosis was diagnosed. This complication had never been reported in a patient with systemic lupus erythematosus, and related literature is reviewed. PMID- 10713654 TI - Dendritic cell biology and the application of dendritic cells to immunotherapy of multiple myeloma. AB - Dendritic cells (DCs) are extremely efficient antigen-presenting cells that are potent stimulators of both B and T cell immune responses. Although DCs are normally present in extremely small numbers in the circulation, recent advances in DC biology have made it possible to generate DCs in culture. DCs can be generated in vitro from various cellular sources including bone marrow, cord blood and peripheral blood. Although culture conditions are extremely diverse, the majority of protocols grow DCs in GM-CSF and either TNF-alpha and/or IL-4. The addition of other growth factors such as SCF and Flt-3 ligand can dramatically enhance DC recovery. It is important to appreciate that DC subsets have been identified. Thus, DC at different stages of maturation, based on phenotype and capacity to capture antigen, can be obtained depending on culture conditions. For clinical applications, DCs can be generated in serum-free media and cryopreserved for future clinical applications. The ability to obtain DCs in numbers suitable for manipulating immune responses has pushed DC-based immunotherapies into the spotlight for treatment of various malignancies, including multiple myeloma, a B cell malignancy that is presently incurable. Although high-dose chemotherapy and transplantation have improved complete remission rates and overall survival in myeloma, immunotherapeutic strategies are needed for the additional cytoreduction needed to achieve a cure. Because DCs specialize in antigen capture and are extremely potent at stimulating T cell responses, they are ideally suited for generating anti-myeloma T cell responses in vivo. Several studies have demonstrated that myeloma protein, also called idiotype (Id), is sufficiently immunogenic and can be used to generate in vivo T cell responses in myeloma patients. Clinical trials using Id-pulsed DCs as a vaccine to treat minimal residual disease or relapsed myeloma are currently underway. Feasibility studies indicate that antigen-pulsed autologous DCs can be used to elicit in vivo Id-specific T cell responses. Additional studies are needed to optimize current DC vaccination protocols and determine clinical benefits associated with this approach. It is hoped that, following conventional therapies, a combination of adoptive immunotherapeutic modalities such as DCs together with myeloma-specific T cells may lead to improved clinical responses in multiple myeloma, and ultimately lead to complete remission and cure. PMID- 10713655 TI - Myelodysplasia and apoptosis: new insights into ineffective erythropoiesis. PMID- 10713656 TI - c-erb-B2 expression and response to treatment in metastatic breast cancer. AB - The frequency of c-erb-B2 expression, clinical correlates and treatment outcome was investigated in 92 patients with metastatic breast cancer. Positive c-erb-B2 immunostaining was found in 24/92 (26%) of tumours. There was a statistically significant inverse correlation between c-erb-B2 expression and ER status. There was also a significant inverse correlation between c-erb-B2 expression and tumour free interval. c-erb-B2 expression had no influence on response to treatment with tamoxifen among patients co-expressing both c-erb-B2 and estrogen receptor (ER). Following chemotherapy (CAF) treatment there was a trend to a lower response rate among c-erb-B2+ as compared to c-erb-B2- patients. However, more c-erb-B2+ patients had received prior adjuvant chemotherapy, and when this factor was included in a multivariate analysis only prior adjuvant chemotherapy treatment predicted for response to CAF chemotherapy for metastatic disease. Time to treatment failure (TTF) was significantly shorter among cerb B2+ as compared to c erb-B2- patients. The current study suggests that the c-erb-B2 expression is found at similar frequency in metastatic as in primary breast cancer. Although c erb-B2 expression did not predict for response to either hormonal therapy or to chemotherapy for metastatic disease, patients with c-erb-B2+ metastatic lesions appear to have a more aggressive clinical course. PMID- 10713657 TI - Serum erythropoietin level in anemic cancer patients. AB - Anemia is a frequent complication of cancer and its treatment. A defect in erythropoietin production has been advocated as being the main cause of anemia in cancer patients. We studied serum erythropoietin levels in 74 patients with solid tumors and in a control group consisting of 20 otherwise healthy individuals without any malignancy, who have only iron deficiency anemia. Serum erythropoietin levels were measured by enzyme immunoassay in cancer patients without anemia (n=34), and in anemic cancer patients (n=40); either receiving chemotherapy (n=21) or not (n=19). Anemic cancer patients were found to have decreased response of erythropoietin for a given hemoglobin level (mean, 40.1+/ 34.7 u/ml), compared with the patients having only iron deficiency anemia (mean, 69.7+/-68.6 u/ml) (P<0.05). In patients with iron deficiency anemia having no malignancy, erythropoietin response was remarkably high and inversely correlated with the level of hemoglobin (r=-0.69; P=0. 05). Although there was no correlation between hemoglobin and erythropoietin response in cancer anemia (r= 0.07), serum levels of erythropoietin were found to be higher in anemic cancer patients (mean, 40.1+/-34.7 u/ml), compared with cancer patients with normal hemoglobin values (mean, 19.96+/-18.4 u/ml). There was not any statistically significant difference between erythropoietin levels in anemic cancer patients with or without chemotherapy (mean, 43. 7+/-37.7 u/ml and 41.9+/-30.08 u/ml respectively; P>0.05). No difference in serum erythropoietin levels were noted in patients treated with cisplatin or non-cisplatin containing regimens (mean, 48.36+/-33.12 u/ml and 38.55+/-43.52 u/ml, respectively; P>0.05). In this study, we demonstrated that anemia in cancer patients was caused by blunted erythropoietin response, rather than its quantitative deficiency. Serial measurements, however, should be considered in patients receiving chemotherapy. PMID- 10713658 TI - Seropositivity for MIA and S100 in patients with gastrointestinal carcinomas. AB - Serum levels of melanoma inhibiting activity (MIA) and S100, both markers in malignant melanoma, are increased only in few patients with non-melanocytic tumors. We examined a series of serum samples from patients with colorectal (CRC) (N=56), gastric (GC) (N=43), pancreatic (PC) (N=29), hepatocellular (HCC) (N=30), cholangiocellular and gallbladder carcinoma (CCC) (N=18). MIA and S100 were measured by commercially available assays. Positive serum levels for MIA and S100 were found in 16.1% and 5.4% of the patients with CRC, 11.6% and 9.3% with GC, 34.5% and 13.8% with PC, 0% and 30% with HCC and 16.7% with CCC, respectively. All patients with sera positive for either MIA or S100 suffered from advanced tumors and received palliative treatment. Elevated serum levels of MIA and S100 are frequent in patients with gastrointestinal cancer. Further investigation is warranted to define the role of MIA or S100 seropositivity in gastrointestinal cancer with regard to follow-up. PMID- 10713660 TI - Phase II study of etoposide (VP-16) in patients with thyroid cancer with no prior chemotherapy: an Eastern Cooperative Oncology Group Study (E1385). AB - This study of etoposide in thyroid cancer was designed to determine the activity and toxicity of etoposide in a variety of inoperable, thyroid hormone insensitive, and radio-iodine resistant primary cancers of the thyroid. The patients were required to have an ECOG performance status of at least 3 and no previous exposure to chemotherapy. The etoposide was given at a dose of 140 mg/m2 daily for 3 days and every 3 weeks until progression. The study was closed after 18 months because of poor accrual. There were no responses seen among the 10 patients accrued. The toxicity was primarily hematologic. There was no evidence of activity of etoposide in thyroid carcinoma, although this study lacked significant power because of the poor accrual. PMID- 10713661 TI - Induction of matrix metalloprotease-7 is common in mucinous ovarian tumors including early stage disease. AB - Matrix metalloproteases are known to play an important role in tumor invasion by mediating degradation of the extracellular matrix. In this study, we have investigated the immunohistochemical expression of matrix metalloprotease -7 (MMP 7) in 44 mucinous ovarian tumors (9 adenomas, 13 low malignant potential tumors, 22 adenocarcinomas) and 6 normal ovaries. Positive staining of MMP-7 is observed in all mucinous ovarian tumors, whereas little or no staining was observed in surface epithelium as well as the epithelial cells of germinal inclusion cyst of the normal ovary. Positive immunostaining of MMP-7 is also observed in the secreted mucin in the tumor glands, which suggests the secretion of the MMP-7 protein from tumor cells. mRNA expression of MMP-7 was confirmed using RT-PCR. The MMP-7 gene was amplified in parallel with an internal control gene beta tubulin using a thermal cycler. mRNA expression levels of MMP-7 were significantly elevated in mucinous tumor samples compared with that in normal ovaries. Our results suggest that MMP-7 is frequently overexpressed in mucinous ovarian tumors and secreted with the mucin which is produced from the tumor cells. MMP-7 may therefore contribute to mucinous ovarian tumor development or enhanced growth capacity of mucinous ovarian tumors. MMP-7 may also serve as a target for therapeutic intervention in the down regulation of tumor progression. PMID- 10713659 TI - Adding high-dose tamoxifen to CHOP does not influence response or survival in aggressive non-Hodgkin's lymphoma: an interim analysis of a randomized phase III trial. AB - PURPOSE: CHOP is the standard regimen currently used in the management of the majority of patients with aggressive non-Hodgkin's lymphoma (NHL). However, CHOP only produces 30-35% long-term survival. We hypothesized that adding high-dose tamoxifen, which is known to have multiple drug resistance-modulatory effects, to the CHOP regimen could increase the response rate, and consequently enhance the survival of patients with NHL. PATIENTS AND METHODS: In a prospective, controlled, and randomized study, eligible adult patients with aggressive NHL were randomized between CHOP only (Group I), or CHOP plus high-dose tamoxifen (Group II). The primary aim was to assess the effect of tamoxifen on complete response (CR) rate, with the secondary evaluation of tamoxifen potential impact on survival. The interim analysis of this study is presented. RESULTS: Fifty-one and forty-seven evaluable patients were randomized to Group I and Group II, respectively. The median age of all patients was 53 y (range 18-78 y). The two groups had comparable distributions of the pretreatment prognostic variables. The CR for patients in Group I was 80% (41 patients) as compared with 74% (35 patients) in Group II (P=0.48). Likewise, there was no apparent difference in the partial remission rates between the two groups (6% vs 15%, respectively). Of patients who initially attained CR, 15 (37%) and 10 (29%) subsequently relapsed in Groups II and I respectively (P = 0.45). The NHL International Prognostic Index (IPI) was the only factor that predicted attaining CR. At the time of this interim analysis, the actuarial-estimated overall survival (OS) probability (+/ S.E.) for the entire population at 5 y was 58% (+/-6) with no survival difference between the two groups (P=0.51). Only attaining CR and the IPI predicted OS probability. The probability of remaining event-free at 5 y (+/-SE) for those achieving CR was 72% (+/-9), and there was no significant difference between the two treatment groups (P=0.68). Toxicity profile was similar in the two groups. CONCLUSION: Based on this interim analysis, combining high-dose tamoxifen, as used in this study, with the CHOP regimen has failed to have any favorable effect on the outcome of patients with aggressive NHL, and therefore cannot be recommended for future trials. PMID- 10713662 TI - Elevated pretreatment serum levels of Il-10 are associated with a poor prognosis in Hodgkin's disease, the milan cancer institute experience. AB - Alterated cytokine secretion may play a role in determining Hodgkin's disease related immunosuppression. The aim of this study was to analyze the clinical significance of interleukin-10 (IL-10) serum levels in 73 chemotherapy-naive patients with Hodgkin's disease. We evaluated the relationship between pretreatment circulating values of IL-10 and both the clinical characteristics of the disease as well as the prognosis in terms of freedom from progression and overall survival. Abnormally high pre-treatment serum levels (mean+/-standard error: 26.79+/-13.24 pg/ml) were detected in 33/73 (45%) patients. The percentage of patients with enhanced IL-10 secretion was significantly higher in the presence of advanced disease (56% vs 32%, P<0.03), systemic symptoms (57% vs 34%, P<0.04) and more than 3 involved sites (61% vs 36%, P<0.03). The high basal levels of IL-10 negatively influenced long-term results: at 8-years freedom from progression (FFP) and overall survival (OS) for patients with IL-10>6 pg/ml vsF mutation of Y1252-1253, neither ligand binding nor Tpr-mediated dimerization can release this block. Here we show that the M1268T mutation partially rescues the kinase activity (and the transforming ability) of the Y1252-1253F Tpr-Met mutant, but is completely dependent on dimerization for its effect. In contrast, the two D1246H/N mutants strictly depend on Y1252-1253 for activity. Surprisingly, however, they constitutively activate the isolated cytoplasmic TK domain of Met (Cyto-Met). These data indicate that the two mutations operate via distinct mechanisms. PMID- 10713678 TI - Regulation of mda-7 gene expression during human melanoma differentiation. AB - Induction of irreversible growth arrest and terminal differentiation in human melanoma cells following treatment with recombinant human fibroblast interferon (IFN-beta) and mezerein (MEZ) results in elevated expression of a specific melanoma differentiation associated gene, mda-7. Experiments were conducted to define the mechanism involved in the regulation of mda-7 expression in differentiating human melanoma cells. The mda-7 gene is actively transcribed in uninduced HO-1 human melanoma cells and the rate of transcription of mda-7 is not significantly enhanced by treatment with IFN-beta, MEZ or IFN-beta+MEZ. The high basal activity of the mda-7 promoter in uninduced melanoma cells and the absence of enhancing effect upon treatment with differentiation inducers is corroborated by transfection studies using the promoter region of mda-7 linked to a luciferase reporter gene containing the SV40 polyadenylation signal sequence. RT - PCR analysis detects the presence of low levels of mda-7 transcripts in uninduced and concomitant increases in differentiation inducer treated HO-1 cells. However, steady-state mda-7 mRNA is detected only in IFN-beta+MEZ and to a lesser degree in MEZ treated cells. We show that induction of terminal differentiation of HO-1 cells with IFN-beta+MEZ dramatically increases the half-life of mda-7 mRNA while treatment with cycloheximide results in detectable mda-7 mRNA in control and inducer treated cells. These observations confirm constitutive activity of the mda-7 promoter in HO-1 cells irrespective of differentiation status suggesting posttranscriptional processes as important determinants of mda-7 expression during terminal differentiation. The 3' UTR region of mda-7 contains AU-rich elements (ARE) that contribute to rapid mda-7 mRNA turnover during proliferation and reversible differentiation, a process controlled by a labile protein factor(s). Substitution of the SV40 polyadenylation signal sequence in the luciferase reporter plasmid with the mda-7-ARE-3'-UTR renders the Luciferase message unstable when expressed in proliferating and reversibly differentiated melanoma cells. In contrast, the luciferase message is stabilized when the mda-7 ARE-3'-UTR construct is expressed in terminally differentiated HO-1 cells. These results provide compelling evidence that mda-7 expression during terminal differentiation in human melanoma cells is regulated predominantly at a posttranscriptional level. PMID- 10713679 TI - Mouse models for breast cancer. PMID- 10713680 TI - The mammary pathology of genetically engineered mice: the consensus report and recommendations from the Annapolis meeting. AB - NIH sponsored a meeting of medical and veterinary pathologists with mammary gland expertise in Annapolis in March 1999. Rapid development of mouse mammary models has accentuated the need for definitions of the mammary lesions in genetically engineered mice (GEM) and to assess their usefulness as models of human breast disease. The panel of nine pathologists independently reviewed material representing over 90% of the published systems. The GEM tumors were found to have: (1) phenotypes similar to those of non-GEM; (2) signature phenotypes specific to the transgene; and (3) some morphological similarities to the human disease. The current mouse mammary and human breast tumor classifications describe the majority of GEM lesions but unique morphologic lesions are found in many GEM. Since little information is available on the natural history of GEM lesions, a simple morphologic nomenclature is proposed that allows direct comparisons between models. Future progress requires rigorous application of guidelines covering pathologic examination of the mammary gland and the whole animal. Since the phenotype of the lesions is an essential component of their molecular pathology, funding agencies should adopt policies ensuring careful morphological evaluation of any funded research involving animal models. A pathologist should be part of each research team. PMID- 10713681 TI - The interactive Web-based histology atlas system. AB - Molecular pathways and mechanisms underlying human cancer are frequently investigated in animal models. Studies to identify and dissect molecular pathways include the discovery of genes and their subsequent analysis in transgenic and 'knock-out' mice. A critical aspect in such investigations is the evaluation of organ integrity and histology upon the alteration or inactivation of specific genes. Results from such studies are usually published in scientific journals. However, due to print space and costs the display of large high quality images is limited. Furthermore, the printed media does not permit an easy comparison of histological images published in different journals and different years. The Internet provides a tool for the timely and inexpensive dissemination of scientific data to the research community. However, its potential for the analysis of histological images has not been explored. Here we present a web based interactive histology atlas (http://histology.nih.gov) that permits the retrieval of annotated, high-resolution histology images via the Internet. This histology atlas also takes advantage of the interactive nature of the Internet to support the communication between different research groups. As an outline forum this atlas provides the framework to evaluate and understand cancer pathology, and to develop a consensus between veterinary and human pathologists. PMID- 10713683 TI - Use of MMTV-Wnt-1 transgenic mice for studying the genetic basis of breast cancer. AB - Wnt-1 was first identified as a protooncogene activated by viral insertion in mouse mammary tumors. Transgenic expression of this gene using a mouse mammary tumor virus LTR enhancer causes extensive ductal hyperplasia early in life and mammary adenocarcinomas in approximately 50% of the female transgenic (TG) mice by 6 months of age. Metastasis to the lung and proximal lymph nodes is rare at the time tumors are detected but frequent after the removal of the primary neoplasm. The potent mitogenic effect mediated by Wnt-1 expression does not require estrogen stimulation; tumors form after an increased latency in estrogen receptor alpha-null mice. Several genetic lesions, including inactivation of p53 and over-expression of Fgf-3, collaborate with Wnt-1 in leading to mammary tumors, but loss of Sky and inactivation of one allele of Rb do not affect the rate of tumor formation in Wnt-1 TG mice. PMID- 10713682 TI - MMTV-induced mammary tumorigenesis: gene discovery, progression to malignancy and cellular pathways. AB - The study of the mouse mammary tumor virus (MMTV) has provided important insights into the mechanisms of gene transcription regulation by steroid hormones, the mode of action of heritable super antigens and the progressive nature of neoplastic transformation in the mammary gland. Here we describe the current situation with respect to the latter aspect of MMTV biology and the prospects for further advance in our understanding of breast cancer in humans that may be expected from a continued study of MMTV-induced mammary neoplasia. MMTV is a heritable somatic mutagen whose target range is limited. Commonly, the tumorigenic capacity of MMTV is restricted to mammary gland, whereas infection is found in a variety of cell types. In order to replicate, proviral DNA must be inserted into the cell DNA and cell division is required to fix the mutation. Yet only in the mammary epithelium does this lead to neoplastic transformation. This suggests a unique relationship between MMTV and mammary epithelium. In evaluating this relationship, we and others have discovered genes and potential gene pathways that are pertinent in mammary differentiation and neoplasia. In addition, the clonal nature of these progressive events from normal to malignant phenotype has become increasingly clear. The weight of these observations compel us to conclude that mammary neoplasms arise from multipotent mammary epithelial cells through a process of acquired mutations that are reflected in the increasingly malignant nature of the population of progeny produced by these damaged stem cells. PMID- 10713684 TI - WAP-TAg transgenic mice and the study of dysregulated cell survival, proliferation, and mutation during breast carcinogenesis. AB - Understanding the process of carcinogenesis is key to developing therapies which might interrupt or reverse tumor onset and progression. Cell growth and death signals are dependent not only upon molecular mechanisms within a cell but also upon external stimuli such as hormones, cell - cell signaling, and extracellular matrix. Mouse models can be used to dissect these complex processes, to identify key signaling pathways operating at different stages of tumorigenesis, and to test the strength of specific interventions. In the WAP-TAg mouse model, carcinogenesis is initiated by expression of the Simian Virus 40 T antigen (TAg). TAg expression is triggered by hormonal stimulation, either during estrus or pregnancy. Breast adenocarcinomas (ranging from well to poorly differentiated) develop in 100% of the female mice by approximately 8 - 9 months of age. Three distinct stages of tumorigenesis are easily identified: an initial proliferation, hyperplasia, and adenocarcinoma. The mean time to first palpable tumor in mice which undergo at least one pregnancy is 6 months. The tumorigenic process is marked by a competition between proliferation and apoptosis and is characterized by cellular acquisition of genetic mutations and increased stromal fibrosis. Protein levels of cell cycle control genes cyclin D1, cdk2, and E2F-1 are increased in these adenocarcinomas. c-Fos protein levels are slightly increased in these cancers, while c-Jun levels do not change. Hormonal exposure alters progression. Estrogen plays a role during the early stages of oncogenesis although the growth of the resulting adenocarcinomas is estrogen-independent. Transient hormonal stimulation by glucocorticoids that temporarily increases the rate of cell proliferation results in tetraploidy, premature appearance of irreversible hyperplasia, and early tumor development. Tumor appearance also can be accelerated through over expression of the cell survival protein, Bcl-2. Bcl-2 over expression not only reduces apoptosis during the initial proliferative process but also decreases the total rate of cell proliferation. This block in cell proliferation is lost selectively as the cells transition to adenocarcinoma. The WAP-TAg model can be utilized to investigate how the basic processes of cell proliferation, apoptosis, DNA mutation, and DNA repair are modified by external and internal signals during mammary oncogenesis. PMID- 10713686 TI - A transgenic mouse model for the ductal carcinoma in situ (DCIS) of the mammary gland. AB - The ductal carcinoma in situ (DCIS) of the mammary gland represents an early, pre invasive stage in the development of invasive breast carcinoma and is increasingly diagnosed since the introduction of high-quality mammography screening. Uncertainties in the prognosis for patients with DCIS have caused a controversial discussion about adequate treatment, and it is suspected that most patients undergoing mastectomy may be overtreated. In order to improve treatment and treatment decision, it therefore is highly desirable to identify prognostic markers and therapeutic targets for DCIS. We here introduce a set of transgenic mice (WAP-T and WAP-T-NP lines) presenting with various morphological forms of DCIS-like lesions. In these mice the SV40 large tumor antigen is specifically induced in epithelial cells of the terminal duct lobular units (TDLU). As a consequence of continuous expression of the oncogene, the animals develop multifocal DCIS and consequently invasive carcinoma within strain specific periods of latency. DCIS lesions in transgenic mice exhibit distinct architectural and cytological features which closely resemble those commonly present in humans. We therefore propose these transgenic lines as an experimental model to study the underlying molecular events leading to DCIS and its progression to invasive disease. PMID- 10713685 TI - The C3(1)/SV40 T-antigen transgenic mouse model of mammary cancer: ductal epithelial cell targeting with multistage progression to carcinoma. AB - The 5' flanking region of the C3(1) component of the rat prostate steroid binding protein (PSBP) has been used to successfully target the expression of the SV40 large T-antigen (Tag) to the epithelium of both the mammary and prostate glands resulting in models of mammary and prostate cancers which histologically resemble the human diseases. Atypia of the mammary ductal epithelium develops at about 8 weeks of age, progressing to mammary intraepithelial neoplasia (resembling human ductal carcinoma in situ [DCIS]) at about 12 weeks of age with the development of invasive carcinomas at about 16 weeks of age in 100% of female mice. The carcinomas share features to what has been classified in human breast cancer as infiltrating ductal carcinomas. All FVB/N female mice carrying the transgene develop mammary cancer with about a 15% incidence of lung metastases. Approximately 10% of older male mice develop anaplastic mammary carcinomas. Unlike many other transgenic models in which hormones and pregnancy are used to induce a mammary phenotype, C3(1)/Tag mice develop mammary tumors in the mammary epithelium of virgin animals without hormone supplementation or pregnancy. Although mammary tumor development appears hormone-responsive at early stages, invasive carcinomas are hormone-independent, which corresponds to the loss of estrogen receptor-alpha expression during tumor progression. Molecular and biologic factors related to mammary tumor progression can be studied in this model since lesions evolve over a predictable time course. Genomic alterations have been identified during tumor progression, including an amplification of the distal portion of chromosome 6 containing ki-ras and loss of heterozygosity (LOH) in other chromosomal regions. We have demonstrated that stage specific alterations in the expression of genes which are critical regulators of the cell cycle and apoptosis are functionally important in vivo. C3(1)/Tag mice appear useful for testing particular therapies since growth of the mammary tumors can be reduced using chemopreventive agents, cytokines, and an anti-angiogenesis agent. PMID- 10713687 TI - Signal transduction in mammary tumorigenesis: a transgenic perspective. AB - A number of genes have been implicated in breast cancer development, yet few have been demonstrated to play causative roles in mammary tumor formation. The advent of transgenic mouse and embryonic stem cell technologies now permits manipulation of the mouse genome in such a way as to temporally and spatially control a gene product's expression. Thus, the basic researcher now can directly assess the involvement of particular genes in tumorigenesis and disease progression and, in the process, to develop mouse models of human genetic disease. The utility of such technologies is emphasized in transgenic mice expressing genes thought to play important roles in the initiation and progression of mammary carcinomas. As these transgenic strains have been the subject of several reviews, here we focus on two mouse mammary tumor models, Polyomavirus middle T antigen and the Neu/ErbB 2 receptor tyrosine kinase, which are most amenable to study specific signaling pathways in process of mammary tumorigenesis. PMID- 10713688 TI - Mutant p53 and genomic instability in a transgenic mouse model of breast cancer. PMID- 10713689 TI - A mammary-specific model demonstrates the role of the p53 tumor suppressor gene in tumor development. AB - Although alterations in the p53 tumor suppressor gene are detected frequently in human breast cancers, mammary tumors are observed infrequently in p53(null) mice. This has led to the suggestion that absence of p53 alone is not sufficient for induction of mammary tumors. However, early death of p53(null) mice from thymic lymphomas may obscure tumor phenotypes that would develop later. Therefore, p53(null) mammary epithelium was transplanted into cleared mammary fat pads of wild type p53 BALB/c hosts to allow long-term analysis of mammary tumor phenotypes. Five treatments were compared for their effects on tumor incidence in hosts bearing transplants of p53(null) and p53wt mammary epithelium. The treatment groups were: (1) untreated; (2) continuous hormone stimulation with pituitary isografts; (3) multiple pregnancies; (4) DMBA alone; and (5) DMBA+pituitary isografts. The tumor incidences in p53(null) vs p53wt mammary transplants for each treatment group were 62% vs 0%, 100% vs 0%, 68% vs 0%, 60% vs 4% and 91% vs 14%, respectively. The mammary tumors that developed in the p53(null) mammary epithelium were all adenocarcinomas and were frequently aneuploid. These data demonstrate that the absence of p53 is sufficient to cause development of mammary tumors and that hormonal stimulation enhances the tumorigenicity of p53(null) mammary epithelium to a greater extent than DMBA exposure alone. This model provides an in situ approach to examine the molecular basis for the role of p53 in the regulation of mammary tumorigenesis. PMID- 10713690 TI - Role of the tumor suppressor gene Brca1 in genetic stability and mammary gland tumor formation. AB - Germline mutations in the tumor suppressor BRCA1 predispose women to breast and ovarian cancers. Current evidence demonstrates that mutations in BRCA1 do not directly result in tumor formation, but instead cause genetic instability, subjecting cells to high risks of malignant transformation. In an animal model in which Brca1 is mutated specifically in mammary epithelium, tumorigenesis occurs in mutant glands at low frequency after a long latency. Notably, introduction of a p53-null allele significantly enhanced mammary gland tumor formation in Brca1 conditional mutant mice. These results are consistent with a model that Brca1 is a caretaker gene, whose absence causes genetic instability and triggers further alterations, including inactivation of tumor suppressor genes and/or activation of oncogenes, leading to tumor formation. PMID- 10713691 TI - Dual regulation of proliferation and apoptosis: c-myc in bitransgenic murine mammary tumor models. AB - Recent progress in the study of c-Myc has convincingly demonstrated that it possesses a dual role in regulating both proliferation and apoptosis; however, the manner in which c-Myc influences these cellular response pathways remains incompletely characterized. Deregulation of c-Myc expression, via many mechanisms, is a common feature of multiple cancers and is an especially prominent feature of many breast cancers. Of significant interest to those who study mammary gland development and neoplasia is the unresolved nature and contribution of apoptosis to breast tumorigenesis. Recently, the use of transgenic mice and gene-knockout mice has allowed investigators to evaluate the pathological mechanisms by which different genes influence tumor development and progression. In this review, we address two distinct c-myc-containing bitransgenic murine mammary tumor models and discuss the contribution and possible future directions for resolution of cancer-relevant molecular pathways influenced by c-Myc. PMID- 10713692 TI - The role of prolactin and growth hormone in breast cancer. AB - This review will focus on the role for prolactin (PRL) and growth hormone (GH) in mammary tumor formation. Much attention has previously been focused on circulating levels of GH/PRL in relation to mammary tumor formation. We will review data demonstrating that these ligands also could be produced locally in different organs, including the mammary gland and mammary tumors, and suggest that this local production may be of importance for pathological conditions. We will also discuss mechanisms for crosstalk between steroids and GH/PRL. A crosstalk between GH- and PRL response is possible at multiple levels. In the human, GH can activate both the prolactin receptor (PRLR) and the growth hormone receptor (GHR). We have demonstrated that activation of the PRLR, but not the GHR, is inducing mammary tumors in transgenic mice. Furthermore, the elevated levels of insulin-like growth factor 1 (IGF-I) seen in the GHR activating transgenic mice is not sufficient for tumor induction. The induced tumors express functionally active prolactin that could be of importance for the tumor formation. Paracrine/aurocrine stimulation by PRL may be more important than PRL transported via the circulation. In women, the role for stimulation of the PRLR and/or the GHR in mammary tumor formation has not been proven, although experiments from primates suggest that the PRLR could be of importance. PMID- 10713693 TI - Prolactin gene-disruption arrests mammary gland development and retards T-antigen induced tumor growth. AB - Prolactin (PRL), interacting with other hormones from the pituitary, gonad, and placenta, activates specific signals that drive the appropriately timed morphological and functional development of the mammary gland. A mouse model of isolated PRL deficiency (PRL-/-) was created by gene disruption in an effort to further understand the molecular basis of mammary gland development and breast cancer. Whereas primary ductal growth was normal in PRL-/- mice, ductal arborization was minimal (branches/mm2=1.5+/-0.5), and lobular budding was absent. Replacement therapy with PRL injections stimulated a modest degree of lobular budding and ductal arborization (3.75+/-0.9). Pituitary transplants to the kidney capsule of PRL-/- mice restored lobular budding and ductal arborization, to the full extent of that seen in control animals (20. 3+/-5.5). Pregnancy, established by mating progesterone-treated PRL-/- females with PRL-/- males, led to complete morphological development of the mammary gland, appropriate to the gestational stage. PRL treatment stimulated tyrosine phosphorylation and DNA binding activity of Stat5a, but not Stat1 in PRL-/- or PRL+/- females, and Stat5a, but not Stat1, was elevated by estradiol within 24 h. PRL-deficient mice were crossed with mice expressing a dominant oncogene (polyoma middle-T antigen driven by the MMTV promoter, PyVT mice). Palpable (1 mm3) tumors were detected an average of 9 days earlier in hormonally normal females (PRL+/ :PyVT) compared with littermates that were PRL-deficient (PRL-/-:PyVT). The growth rate of PyVT-induced tumors was 30% faster in PRL+/-, than in PRL-/- females. PMID- 10713694 TI - Transforming growth factor alpha and mouse models of human breast cancer. AB - Transforming growth factor alpha (TGFalpha) is a principal molecule in the normal and neoplastic development of the mammary gland. Binding of TGFalpha to the epidermal growth factor receptor (EGFR), activates the EGFRs' endogenous tyrosine kinase activity and stimulates growth of the epithelium in the virgin and pregnant mouse mammary gland. TGFalpha expression can be detected in breast cancer cells in vivo and in vitro and overexpression can elicit partial transformation or immortalized human and rodent mammary epithelial cells. Despite evidence implicating TGFalpha in the development of mammary neoplasia, the actual mechanism of TGFalpha-induced transformation is unclear. Transgenic mouse models targeting heterologus TGFalpha to the mammary gland have established TGFalpha overexpression can induce hyperproliferation, hyperplasia and occasional carcinoma. These transgenic studies demonstrated a facilitating, proliferative role for TGFalpha in the development of neoplasia and implicated several oncogenes that can cooperate with TGFalpha to transform the mammary epithelium. From studies of EGFR signaling pathways, inhibitory and modulating agents such as anti-EGFR antibodies and specific kinases inhibitors have been used to block the action of this pathway and prevent the development of TGFalpha-induced neoplasia and tumor formation. Studies in Stat5a knockout mice have established that the JAK2/Stat5a pathway can facilitate the survival of the mammary epithelium and can impact the progression of TGFalpha-mandated mammary tumorigenesis. Together these experiments indicate that TGFalpha and the EGFR signaling pathway are potentially amenable to therapies for treatment of human breast disease. PMID- 10713695 TI - Transforming growth factor alpha- and c-myc-induced mammary carcinogenesis in transgenic mice. AB - The growth factor transforming growth factor alpha (TGFalpha) and the nuclear transcription factor c-myc often are overexpressed by human breast cancer cells. To produce models of breast disease with these etiologies, mice were generated that carried TGF-alpha- or c-myc-encoding transgenes. Transgene targeting employed the whey acidic protein (WAP) gene promoter, which is expressed in pregnant and lactating mammary epithelial cells. Non-virgin WAP-TGFalpha transgenic mice displayed accelerated mammary development during pregnancy, delayed post-parturient mammary involution, a progressive increase in the number of hyperplastic alveolar nodules (HANs), and development of mammary carcinoma with a mean latency of 9 months. Non-virgin WAP-c-myc transgenic mice displayed accelerated mammary gland development during pregnancy and development of mammary carcinomas with a latency of 8 months. Bitransgenic mice carrying both WAP TGFalpha and WAP-c-myc displayed a dramatic acceleration of tumor development. These models identify the overexpression of TGFalpha or c-myc as etiological factors in the development of mammary neoplasia and demonstrate the increased severity of disease when both molecular alterations are present in the same cell. PMID- 10713696 TI - Mammary gland oncogenes as indicators of pathways important in mammary gland development. AB - The identification of dominant acting proto-oncogenes in mammary tumors from mice and humans has highlighted a number of signal transduction pathways that have subsequently been shown to have a role in normal mammary growth and differentiation. Here we describe the use of two different transgenic mouse strategies to investigate the function of two of these signalling pathways in the normal growth and differentiation of the mouse mammary gland during pregnancy. PMID- 10713698 TI - Genetic determinants of response to chemotherapy in transgenic mouse mammary and salivary tumors. AB - Several transgenic mouse tumor models were utilized to explore how specific genetic alterations affect the tumor cell response to chemotherapeutic agents in vivo. Specifically, MMTV-ras transgenic mice were interbred to p53 knock-out mice to create a model for assessing the role of p53 in chemotherapeutic responses. In addition, MMTV-ras tumors were compared to MMTV-myc and MMTV-ras/myc tumors. Mice of each genotype reproducibly develop mammary and/or salivary tumors, but tumor growth dynamics vary considerably between genotypes. MMTV-ras/p53-/- tumors exhibit higher S phase fractions than MMTV-ras/p53+/+ tumors, although both tumor types display very low apoptosis levels. In contrast, MMTV-myc tumors exhibit both high S phase fractions and spontaneous apoptosis levels. Tumor-bearing mice of each genotype were treated with either doxorubicin or paclitaxel, and effects on overall tumor growth, cell cycle distribution and apoptosis were evaluated. Surprisingly, neither agent efficiently induced apoptosis in any of the tumor models, including those with wildtype p53. Rather, tumor responses were mediated primarily by changes in cell cycle distribution. However, the spontaneous apoptosis levels did serve as a predictor of tumor growth response, in that only those tumors with high pretreatment apoptosis levels underwent significant regression following treatment with either agent. PMID- 10713697 TI - The matrix metalloproteinase stromelysin-1 acts as a natural mammary tumor promoter. AB - Extracellular matrix-degrading matrix metalloproteinases (MMPs) are invariably upregulated in epithelial cancers and are key agonists in angiogenesis, invasion and metastasis. Yet most MMPs are secreted not by the cancer cells themselves, but by stromal cells within and around the tumor mass. Because the stromal environment can influence tumor formation, and because MMPs can alter this environment, MMPs may also contribute to the initial stages of cancer development. Several recent studies in MMP-overexpressing and MMP-deficient mice support this possibility, but have required carcinogens or pre-existing oncogenic mutations to initiate tumorigenesis. Here we review the spontaneous development of premalignant and malignant lesions in the mammary glands of transgenic mice that express an autoactivating form of MMP-3/stromelysin-1 under the control of the whey acidic protein gene promoter. These changes were absent in nontransgenic littermates and were quenched by co-expression of a human tissue inhibitor of metalloproteinases-1 (TIMP-1) transgene. Thus by altering the cellular microenvironment, stromelysin-1 can act as a natural tumor promoter and enhance cancer susceptibility. PMID- 10713699 TI - Selective activation of NF-kappa B subunits in human breast cancer: potential roles for NF-kappa B2/p52 and for Bcl-3. AB - Members of the NF-kappa B/Rel transcription factor family have been shown recently to be required for cellular transformation by oncogenic Ras and by other oncoproteins and to suppress transformation-associated apoptosis. Furthermore, NF kappa B has been shown to be activated by several oncoproteins including HER2/Neu, a receptor tyrosine kinase often expressed in human breast cancer. Human breast cancer cell lines, human breast tumors and normal adjacent tissue were analysed by gel mobility shift assay, immunoblotting of nuclear extracts and immunohistochemistry for activation of NF-kappa B. Furthermore, RNA levels for NF kappa B-activated genes were analysed in order to determine if NF-kappa B is functionally active in human breast cancer. Our data indicate that the p65/RelA subunit of NF-kappa B is activated (i.e., nuclear) in breast cancer cell lines. However, breast tumors exhibit an absence or low level of nuclear p65/RelA but show activated c-Rel, p50 and p52 as compared to nontumorigenic adjacent tissue. Additionally, the I kappa B homolog Bcl-3, which functions to stimulate transcription with p50 or p52, was also activated in breast tumors. There was no apparent correlation between estrogen receptor status and levels of nuclear NF kappa B complexes. Transcripts of NF-kappa B-regulated genes were found elevated in breast tumors, as compared to adjacent normal tissue, indicating functional NF kappa B activity. These data suggest a potential role for a subset of NF-kappa B and I kappa B family proteins, particularly NF-kappa B/p52 and Bcl-3, in human breast cancer. Additionally, the activation of functional NF-kappa B in these tumors likely involves a signal transduction pathway distinct from that utilized by cytokines. PMID- 10713700 TI - Transcriptional activation of the hepatocyte growth factor receptor (c-met) gene by its ligand (hepatocyte growth factor) is mediated through AP-1. AB - Hepatocyte Growth Factor (HGF) exerts its biological effects via binding and activating a transmembrane protein tyrosine kinase receptor known as c-Met. Previous studies from our laboratory demonstrated that c-met gene expression is inducible by its own ligand (HGF). However, the molecular mechanism(s) involved in this process are unknown. The present study was carried out to address this question. Transfection of various c-met-CAT promoter constructs into the mouse hepatocellular carcinoma cell line Hepa 1-6 in combination with electrophoretic mobility shift assays (EMSA) identified the responsive element as an activated protein-1 (AP-1) binding site (TGAGTCA) within the c-met core promoter region at position -158 to -152. The c-met AP-1 element binds specifically to AP-1 protein as verified by supershift assays. EMSA studies and mutational analyses of the promoter region also revealed that the members of the Sp family of transcription factors (Sp-1 and Sp-3) bind to the c-met Sp-1 element (located at position -124) which is adjacent to the AP-1 site. We show that Sp binding dampens binding of AP 1 to its cognate site in the c-met promoter region. Stimulation of Hepa 1-6 cells with HGF resulted in a rapid and dramatic enhancement of the AP-1 binding activity as well as an overall increase in the level of AP-1 protein. Cotransfection of AP-1 expression vectors (c-Fos plus c-Jun) with c-met promoter constructs resulted in stimulation of c-met promoter activity. We found that transactivation of the c-met promoter by AP-1 can be blocked by Curcumin, an inhibitor of AP-1. Moreover, we found that the induction of the endogenous c-met gene by HGF is inhibited by the addition of Curcumin. The results demonstrate that the HGF-induced transcription of the c-met gene by HGF is, at least in part, due to activation of the AP-1 pathway. PMID- 10713701 TI - Positive regulation of skeletal myogenesis by R-Ras. AB - Most of the proteins in the Ras-family proteins, including Ras, Rap and TC21, have been reported to be strong inhibitors of skeletal myogenesis. Here we show that R-Ras, another member of this family, promotes terminal differentiation of C2C12 skeletal myoblasts. In contrast to Ras, which induced a markedly transformed phenotype of C2C12 cells, an activated mutant of R-Ras (R-RasQ87L) did not exhibit any inhibitory effect on the differentiation of C2C12 cells, but enhanced the formation of multinucleated myotubes. Although R-RasQ87L showed little effect on induction of two muscle-specific proteins, creatine kinase and myogenin, it prevented cell death during myoblast differentiation, probably through Akt activation and Bcl-xL induction. Motility of C2C12 cells, which may be involved in fusion of myoblasts, was also stimulated by R-RasQ87L. Furthermore, we observed a transient activation of endogenous R-Ras during differentiation of C2C12 cells. The ectopic expression of R-Ras GAP inhibited the differentiation. These results suggest that R-Ras has a positive effect on the terminal differentiation of myoblasts and may be involved in the program of skeletal myogenesis. PMID- 10713702 TI - Dimerization of the amino terminal domain of p57Kip2 inhibits cyclin D1-cdk4 kinase activity. AB - Previous studies have led to the proposal that a single molecule of Cki can associate with the cyclin/Cdk complex to repress its activity. On the other hand, multiple inhibitor molecules are required to inhibit Cdks. In the present work, by using differently tagged p57Kip2 proteins we demonstrate that p57Kip2 can bind to itself in vitro and in vivo. Mutational deletion analysis showed that the NH2 terminal domain of p57Kip2 is necessary and sufficient to dimerization. Using an in vitro competition/association assay, we demonstrate that cyclin D1 alone, Cdk4 alone and/or cyclin D1/Cdk4 complexes do not compete for the p57Kip2 homodimers formation. However, a mutation in the alpha-helix domain of p57Kip2 (R33L) strongly reduced homodimer formation but did not modify interaction with cyclin D1-Cdk4 complexes. Also, increasing amounts of p57Kip2 lead in vivo to a significant augmentation in the level of p57Kip2 homodimerization associated with cyclin D1-Cdk4 complexes and to a marked inhibition of the cyclin D1-Cdk4 kinase activity. Altogether, these data suggest a model whereby p57Kip2 associates with itself by using the NH2 domain to form a homodimeric species which interacts with and inhibits the cyclin D1-Cdk4 complexes. PMID- 10713703 TI - Caspase-dependent cleavage and inactivation of the Vav1 proto-oncogene product during apoptosis prevents IL-2 transcription. AB - Here we identify the hematopoietic proto-oncogene Vav1 as a caspase substrate during apoptosis in lymphoid cells. Cleavage of Vav1 is prevented by the caspase inhibitors zDEVD and zVAD as well as by expression of CrmA. Vav1 is cleaved in vivo at the evolutionary conserved caspase consensus cleavage site DLYD161C, generating the carboxy-terminal cleavage product Vav1p76 of intermediate stability. In vitro caspase assays reveal cleavage of Vav1 at position 161 either by apoptotic cell lysates or by recombinant caspase-3. Mutation of Asp 161 to Ala leads to the usage of the adjacent alternative cleavage sequence DQID150D. Mutation of both cleavage sites at position 150 and 161 protects Vav1 from caspase-mediated proteolysis in vitro and in vivo. The cleavage product Vav1p76 is capable of activating JNK in T-cells, but fails to induce the phosphorylation of p38/HOG1. Vav1p76 displays a diminished capacity to activate the transcription factors NF-AT, AP-1 and NF-kappaB, and thus completely fails to activate IL-2 transcription. Since Vav1 is essential for IL-2 production and plays a central role for cytoskeletal reorganization, its proteolytic inactivation during apoptosis affects multiple downstream targets. PMID- 10713704 TI - Cooperation between STAT5 and phosphatidylinositol 3-kinase in the IL-3-dependent survival of a bone marrow derived cell line. AB - Cytokine-dependent activation of distinct signaling pathways is a common scheme thought to be required for the subsequent programmation into cell proliferation and survival. The PI 3-kinase/Akt, Ras/MAP kinase, Ras/NFIL3 and JAK/STAT pathways have been shown to participate in cytokine mediated suppression of apoptosis in various cell types. However the relative importance of these signaling pathways seems to depend on the cellular context. In several cases, individual inhibition of each pathway is not sufficient to completely abrogate cytokine mediated cell survival suggesting that cooperation between these pathways is required. Here we showed that individual inhibition of STAT5, PI 3 kinase or MEK activities did not or weakly affected the IL-3 dependent survival of the bone marrow derived Ba/F3 cell line. However, the simultaneous inhibition of STAT5 and PI 3-kinase activities but not that of STAT5 and MEK reduced the IL 3 dependent survival of Ba/F3. Analysis of the expression of the Bcl-2 members indicated that phosphorylation of Bad and Bcl-x expression which are respectively regulated by the PI 3-kinase/Akt pathway and STAT5 probably explain this cooperation. Furthermore, we showed by co-immunoprecipitation studies and pull down experiments with fusion proteins encoding the GST-SH2 domains of p85 that STAT5 in its phosphorylated form interacts with the p85 subunit of the PI 3 kinase. These results indicate that the activations of STAT5 and the PI 3-kinase by IL-3 in Ba/F3 cells are tightly connected and cooperate to mediate IL-3 dependent suppression of apoptosis by modulating Bad phosphorylation and Bcl-x expression. PMID- 10713705 TI - Induction of apoptosis by the transactivating domains of the hepatitis B virus X gene leads to suppression of oncogenic transformation of primary rat embryo fibroblasts. AB - Epidemiology shows a clear correlation between chronic infection with the hepatitis B virus (HBV) and development of hepatocellular carcinoma (HCC). The potential role of the transactivating hepatitis B virus X protein (HBx) in transformation by HBV is controversial. Here we report that HBx suppresses transformation of primary rat embryo fibroblasts (REFs). Cooperating oncogenes like c-Ha-ras and c-myc transform REF very efficiently but cotransfection with HBx suppressed transformation of REFs down to 5%. Similarly, transfection of HBx together with the cooperating oncogenes Ha-ras and SV40 LTAg or c-Ha-ras and mutant p53 reduced the number of foci to 13%. Comparable results were obtained with HBx in the context of the whole HBV. Suppression of focus formation in REF could be partly relieved by cotransfection of apoptosis inhibitors Bcl-2 or E1B. However, cotransfection of apoptosis inhibitors crmA and p35 did not influence the proapoptotic functions of HBx. Thus, HBx may specifically activate the Bcl-2 sensitive pathway leading to apoptosis. Experiments with 13 HBx linker scanning mutants revealed that the domains necessary for HBx dependent transactivation overlap with the domains needed for the apoptotic/growth arrest functions of HBx. PMID- 10713706 TI - Differential role of caspase-8 and BID activation during radiation- and CD95 induced apoptosis. AB - Activation of the CD95 death receptor as well as ionizing radiation induces apoptotic cell death in human lymphoma cells. The activation of caspases is a hallmark of apoptosis induction irrespective of the apoptotic trigger. In contrast to death receptor signaling, the exact mechanisms of radiation-induced caspase activation are not well understood. We provide evidence that both, radiation and CD95 stimulation, induce the rapid activation of caspase-8 and BID followed by apoptosis in Jurkat T-cells. To analyse the relative position of caspase-8 within the apoptotic cascade we studied caspase activation and apoptosis in Jurkat cells overexpressing Bcl-2 or Bcl-xL. Caspase-8 activation, pro-apoptotic BID cleavage and apoptosis in response to radiation were abrogated in these cells, while the responses to CD95 stimulation were only partially attenuated by overexpression of Bcl-2 family members. In parallel, the breakdown of the mitochondrial transmembrane potential (DeltaPsim) in response to radiation was inhibited by overexpression of Bcl-2/Bcl-xL Jurkat cells genetically deficient for caspase-8 were found to be completely resistant towards CD95. However, radiation-induced apoptotic responses in caspase-8-negative cells displayed only a modest reduction. We conclude that ionizing radiation activates caspase-8 and BID downstream of mitochondrial damage suggesting that, in contrast to CD95, both events function as executioners rather than initiators of the apoptotic process. PMID- 10713707 TI - Low frequency of alterations of the alpha (PPP2R1A) and beta (PPP2R1B) isoforms of the subunit A of the serine-threonine phosphatase 2A in human neoplasms. AB - The phosphatase 2A (PP2A) is one of the major cellular serine-threonine phosphatases. It was recently shown that the gene encoding for the beta isoform of its subunit A, PPP2R1B, is altered in human lung and colorectal carcinomas, suggesting a role in human tumorigenesis. Here, we report the detection of mutations in breast, lung carcinomas and melanomas in the genes of both alpha (PPP2R1A) and beta isoforms. Mutations affecting PPP2R1B were found in four breast carcinomas, while mutations in PPP2R1A were found in carcinomas of the breast and of the lung and in one melanoma. Most of the mutations affecting PPP2R1B were exons deletions, suggesting abnormal splicing. These splicing abnormalities were detected in tumor samples in the absence of the normal splicing product, and were not found in several normal controls. In one case, a homozygous deletion present in tumor DNA, and not in the matched normal control was demonstrated. Mutations affecting the PPP2R1A gene were nucleotide substitutions changing highly conserved amino acids and one frame-shift. Although the frequency of alterations is low, the inclusion of both isoforms of subunit A in the genes mutated in human cancer and the addition of breast cancer to the list of neoplasms in which PPP2R1B is altered, strengthen the potential role of PP2A in human tumorogenesis. PMID- 10713708 TI - Myb is required for self-renewal in a model system of early hematopoiesis. AB - In hematopoiesis, self-renewal, proliferation, differentiation and apoptosis represent opposing decisions made by stem cells and progenitor cells, which when dysregulated can result in leukaemia. Here, we have investigated the function of Myb proteins in regulating these key cellular decisions, using the cell line FDCP mix A4 as a model of early hematopoiesis. High concentrations of IL-3 in these cells favour self-renewal over differentiation and apoptosis. However when endogenous Myb activity was inhibited with an inducible dominant interfering protein, self-renewal was replaced by apoptosis and differentiation. Differentiation was to granulocytes and monocyte/macrophages and was closely associated with a G1-S phase block in the cell cycle. As for normal hematopoiesis, cytokine-induced terminal differentiation of FDCP-mix cells is associated with concomitant proliferation prior to its completion. However, when Myb activity was inhibited during this process, proliferation and survival were both reduced, resulting in a much lower yield of mature cells. These results indicate multiple cellular roles of Myb proteins during normal hematopoiesis. PMID- 10713709 TI - The cytoprotective aminothiol WR1065 activates p21waf-1 and down regulates cell cycle progression through a p53-dependent pathway. AB - The phosphoaminothiol WR1065, the active metabolite of the pro-drug amifostine (WR2721), protects cultured cells and tissues against cytotoxic exposure to radiation or chemotherapeutic agents. We show here that WR1065 and the pro-drug WR2721 activate the p53 tumor suppressor protein and induce the expression of the cyclin-dependent kinase inhibitor p21waf-1 in the breast cancer cell line MCF-7, and in the mouse fibroblast cell line balb/c 3T3. Using two MCF-7 derived cell lines, MN1 and MDD2, we show that induction of p21waf-1 is detectable in MN1 (expressing a functional p53) but not in MDD2 (p53 disabled). These effects are observed at concentrations of WR1065 (0.5 to 1 mM) identical to those required to protect against cytotoxicity by hydrogen peroxide. Induction of p53 is not prevented by addition of aminoguanidine, an inhibitor of Cu-dependent amine oxidases which blocks the extra-cellular degradation of WR1065 into toxic metabolites. Moreover, spermidine, a natural polyamine structurally related to amifostine, does not activate p53. Induction of p53 by WR1065 results in a delay in the G1/S transition in MCF-7 and MN-1 cells, but not in the p53 disabled cells MDD2. These data indicate that WR1065, a polyamine analog with thiol anti-oxidant properties, activates a cell cycle check-point involving p53. PMID- 10713710 TI - Developmental expression of pim kinases suggests functions also outside of the hematopoietic system. AB - We have cloned a novel quail cDNA with strong homology to the pim family of proto oncogenes. The deduced amino acid (aa) sequence of the cDNA, named qpim, is more closely related to Xenopus Pim and to the recently identified rat Pim-3 than to human or rodent Pim-1 or Pim-2. The protein encoded by the qpim cDNA can autophosphorylate itself and share substrates with murine Pim-1, suggesting functional redundancy to other Pim family serine/threonine kinases. We have compared the expression of qpim in avian embryos to mouse pim-1, -2 and -3 by in situ hybridization. qpim shows a highly dynamic expression pattern, particularly at early developmental stages. Surprisingly, its expression pattern is not identical to any of the murine pim genes, which show complementary and/or partially overlapping expression sites both in- and outside of the hematopoietic system. Altogether, our results suggest novel functions for Pim family kinases during embryonic development, in particular in epithelia and in the central nervous system. PMID- 10713711 TI - Lipopolysaccharide increases cyclo-oxygenase-2 expression in a colon carcinoma cell line through nuclear factor-kappa B activation. AB - Both nonneoplastic colon epithelium and colon carcinoma cells in situ are continuously exposed to lipopolysaccharide (LPS). Few reports have addressed possible direct effects of LPS in promotion of colon carcinoma and underlying mechanisms. We found evidence that LPS directly stimulated growth of the human colon carcinoma cell line CE-1 through an increase in the production of prostaglandin E2 (PGE2) as a result of cyclo-oxygenase-2 (COX-2) expression. LPS induced significant increases in PGE2 production in CE-1 cells, which were found to express a high-affinity LPS receptor, CD14. Positive correlations were found between PGE2 production and activation of nuclear factor (NF)-kappa B as well as expression of both COX-2 mRNA and protein in LPS-stimulated CE-1 cells. When CE-1 cells were exposed to exogenous PGE2, DNA synthesis increased. These results indicate that LPS may stimulate DNA synthesis in certain colon carcinoma cells as a result of PGE2 production involving increased COX-2 expression that might result in turn from activation of NF-kappa B by LPS. Further investigation of the pathways mediating LPS-induced stimulation of colon carcinoma cells may provide insights into LPS effects in in vivo tumor biology. PMID- 10713712 TI - Cytoplasmic domain mutants of beta1 integrin, expressed in beta 1-knockout lymphoma cells, have distinct effects on adhesion, invasion and metastasis. AB - Structural requirements for beta 1 integrin cytoplasmic domain functions in adhesion, migration and signaling have been studied mainly for fibroblasts in vitro. The relevance for beta 1-dependent in vivo migration of lymphoid cells has not been assessed. To study this, we transfected beta 1 mutants into beta 1 deficient double knockout (DKO) ESb lymphoma cells, and tested the capacity of the cells to metastasize to liver and spleen. This was compared to alpha 4 beta 1 dependent invasion into cell monolayers in vitro and Mn2+-induced adhesion to fibronectin. Deletion of the five C-terminal residues or mutation of both threonines T788 and T789 to alanines blocked invasion and metastasis and greatly reduced adhesion, in line with known in vitro effects. However, mutations of the NPXY motif tyrosines had unexpected consequences. A Y783F mutation had no effect at all, but a Y783,795F double mutation strongly reduced Mn2+-induced adhesion, whereas it had limited effects on invasion and metastasis. Furthermore, cells expressing a beta 1 beta 2 chimeric subunit, which contains phenylalanines in the NPXY/F motifs, adhered poorly but invasion and metastasis was fully restored to the same levels as for cells expressing wild-type beta 1. We conclude that part of the functions of the beta 1 cytoplasmic domain that are required for adhesion are not essential for beta 1-dependent invasion and metastasis. PMID- 10713713 TI - N-terminal determinants of I kappa B alpha necessary for the cytoplasmic regulation of c-Rel. AB - I kappa B alpha is a dual regulator of Rel/NF-kappa B transcription factors. I kappa B alpha retains inactive NF-kappa B dimers in the cytoplasm, and inhibits their DNA-binding and transcriptional activities in the nucleus. Our previous studies identified discrete functional domains in I kappa B alpha responsible for the cytoplasmic and nuclear regulation of c-Rel. Determinants necessary for regulating c-Rel in the nucleus mapped to the central ankyrin domain of I kappa B alpha and a few negatively-charged amino acids that follow in the C-terminal PEST region. In contrast, sequences involved in the cytoplasmic regulation of c-Rel reside in the N-terminal and central ankyrin domains of I kappa B alpha. Here, we present a refined mapping of the N-terminal determinants of I kappa B alpha necessary for the cytoplasmic regulation of c-Rel homodimers. We demonstrate that amino acids 48 - 58 in p40/I kappa B alpha are essential to block the nuclear localization of c-Rel dimers. These data define a region of I kappa B alpha that may be required for optimal masking of the c-Rel NLS, or for the nuclear export of c-Rel/I kappa B alpha complexes. These findings highlight a novel function for the N-terminus of I kappa B alpha in the control of the subcellular localization of Rel/NF-kappa B dimers. Given the implication of deregulated NF-kappa B activity in hematopoietic and solid tumors, our findings predict that certain alterations in this domain of I kappa B alpha may have severe biological repercussions. PMID- 10713714 TI - Cell death in the third millennium. PMID- 10713715 TI - Specificity questions concerning the clone 33 anti-fas ligand antibody. PMID- 10713716 TI - c-Abl: activation and nuclear targets. AB - The c-Abl tyrosine kinase and its transforming variants have been implicated in tumorigenesis and in many important cellular processes. c-Abl is localized in the nucleus and the cytoplasm, where it plays distinct roles. The effects of c-Abl are mediated by multiple protein-protein and protein-DNA interactions and its tyrosine kinase domain. At the biochemical level, the mechanism of c-Abl kinase activation and the identification of its target proteins and cellular machineries have in part been solved. However, the phenotypic outcomes of these molecular events remained in large elusive. c-Abl has been shown to regulate the cell cycle and to induce under certain conditions cell growth arrest and apoptosis. In this respect the interaction of c-Abl with p53 and p73 has attracted particular attention. Recent findings have implicated c-Abl in an ionizing irradiation signaling pathway that elicits apoptosis. In this pathway p73 is an important immediate downstream effector. Here I review the current knowledge about these nuclear processes in which c-Abl is engaged and discuss some of their possible implications on cell physiology. Cell Death and Differentiation (2000) 7, 10 - 16. PMID- 10713717 TI - Nuclear translocation of granzyme B in target cell apoptosis. AB - Granzyme B is the prototypic member of a family of serine proteases localized to the cytolytic granules of cytotoxic lymphocytes. Together with another granule protein, perforin, granzyme B is capable of inducing all aspects of apoptotic death in target cells. A number of granzyme B substrates have been identified and it has been demonstrated that granzyme B is responsible, directly or indirectly, for the morphological nuclear changes observed in target cell apoptosis, including DNA fragmentation. In an earlier study, we showed that granzyme B binds to a nuclear protein in a manner dependent on its enzymatic activity. Here, we demonstrate that granzyme B is translocated rapidly to the nucleus in cells that have been induced to undergo apoptosis by a granzyme-dependent process, and that translocation is dependent on caspase activity. Appearance of granzyme B in the nucleus of target cells precedes the detection of DNA fragmentation. Although not directly responsible for DNA fragmentation, these data suggest a nuclear role for granzyme B in target cell apoptosis. c-Abl nuclear functions. PMID- 10713718 TI - The HIV-1 vpr protein induces anoikis-resistance by modulating cell adhesion process and microfilament system assembly. AB - We have previously shown that CD4+ T Jurkat cells constitutively expressing low levels of the human immunodeficiency virus 1 (HIV-1) vpr protein were less susceptible to undergo apoptosis than control cells.1 In this study we have investigated the role of vpr in affecting mechanisms of importance in the control of apoptosis. Vpr-expressing clones consistently aggregated in clusters with time in culture, whereas mock-transfected cells grew as dispersed cultures. The analysis of adhesion molecules involved in cell-to-cell as well as in cell substrate interactions showed a higher expression of cadherin and integrins alpha5 and alpha6 in vpr-transfected clones with respect to mock-transfected cells. This up-modulation was specifically blocked by cell exposure to antisense oligonucleotides targeted at the vpr. In addition, F-actin microfilament cytoskeletal organization, known to be involved in cell-cell interaction pathways and in the modulation of cell surface molecule expression, was significantly improved in vpr-expressing clones, in which filament polymerization was increased. We thus envisage that vpr viral protein can maintain cell survival via a specific activity on cytoskeleton-dependent cell adhesion pathways, i.e. by inducing anoikis-resistance. These particular effects of vpr might enhance the homing, spreading and survival of the infected lymphocytes, thus contributing to virus persistence in the course of acute HIV-1 infection. PMID- 10713719 TI - Primary HIV-1 infection of human CD4+ T cells passaged into SCID mice leads to selection of chronically infected cells through a massive fas-mediated autocrine suicide of uninfected cells. AB - We have recently shown that a human CD4+ T cell line (CEM-SS) acquires the permissiveness to M-tropic strains and primary isolates of HIV-1 after transplantation into SCID mice. This permissiveness was associated with the acquisition of a memory (CD45RO+) phenotype as well as of a functional CCR5 coreceptor. In this study, we have used this model for invest-igating in vivo the relationships between HIV-1 infection, apoptosis and T cell differentiation. When an in vivo HIV-1 infection was performed, the CEM cell tumors grew to a lower extent than the uninfected controls. CEM cells explanted from uninfected SCID mice (ex vivo CEM) underwent a significant level of spontaneous apoptosis and proved to be CD45RO+, Fas+ and Fas-L+, while Bcl-2 expression was significantly reduced as compared to the parental cells. Acute HIV-1 infection markedly increased apoptosis of uninfected ex vivo CEM cells, through a Fas/Fas-L-mediated autocrine suicide/fratricide, while parental cells did not undergo apoptosis following viral infection. The susceptibility to apoptosis of ex vivo CEM cells infected with the NSI strain of HIV-1, was progressively lost during culture, in parallel with the loss of Fas-L and marked changes in the Bcl-2 cellular distribution. On the whole, these results are strongly reminiscent of a series of events possibly occurring during HIV-1 infection. After an initial depletion of bystander CD4+ memory T cells during acute infection, latently or chronically infected CD4+ T lymphocytes are progressively selected and are protected against spontaneous apoptosis through the development of an efficient survival program. Studies with human cells passaged into SCID mice may offer new opportunities for an in vivo investigation of the mechanisms involved in HIV-1 infection and CD4+ T cell depletion. PMID- 10713720 TI - Complement mediated cell death is associated with DNA fragmentation. AB - In this study, we demonstrate for the first time that complement attack of target cells, in the presence of suitably high levels of serum, can induce the oligonucleosomal DNA fragmentation characteristic of apoptosis. This phenomenon requires membrane permeabilisation induced by formation of the complete membrane attack complex and relies on physiologically relevant levels of serum. TUNEL analysis detected complement mediated DNA fragmentation as early as 30 min after the addition of serum and electron microscopy confirmed that chromatin became condensed after complement attack. Various experiments implicate serum DNase I as the mediator of this DNA fragmentation. Intriguingly, membrane permeability induced by melittin gave rise to similar serum dependent DNA fragmentation. The implications of these results for the study of apoptosis in vitro and in vivo are discussed. PMID- 10713721 TI - Micro-environmental factors in the survival of human B-lymphoma cells. AB - Burkitt lymphoma (BL) cells retain a high inherent propensity to undergo apoptosis indicating that net growth of the tumour population in vivo is likely to be influenced profoundly by its micro-environment. Here we investigate micro environmental factors that affect BL-cell survival in vitro. We show that survival, and consequently net production, of tumour cells is enhanced by autocrine factors and, to a greater extent, by paracrine factors provided by relevant stromal elements of the tumour (fibroblasts and follicular dendritic cells) and by macrophages. Promotion of BL-cell survival by paracrine elements was mediated by cell/cell contact and by short-range soluble factor(s). IL-4, IL 10 and TNF-alpha promoted, whereas TGF-beta1 inhibited, tumour-cell production. Macrophages engaged in phagocytosis of apoptotic BL cells were less effective than untreated macrophages in supporting net expansion of BL populations. These results suggest that the net production of tumour cells in BL is supported by multiple micro-environmental factors that modulate apoptosis. PMID- 10713722 TI - The fate of U1 snRNP during anti-Fas induced apoptosis: specific cleavage of the U1 snRNA molecule. AB - During apoptosis, the U1-70K protein, a component of the spliceosomal U1 snRNP complex, is specifically cleaved by the enzyme caspase-3, converting it into a C terminally truncated 40-kDa fragment. In this study, we show that the 40-kDa U1 70K fragment is still associated with the complete U1 snRNP complex, and that no obvious modifications occur with the U1 snRNP associated proteins U1A, U1C and Sm B/B'. Furthermore, it is described for the first time that the U1 snRNA molecule, which is the backbone of the U1 snRNP complex, is modified during apoptosis by the specific removal of the first 5 - 6 nucleotides including the 2,2, 7 trimethylguanosine (TMG) cap. The observations that U1 snRNA cleavage is very specific (no such modifications were detected for the other U snRNAs tested) and that U1 snRNA cleavage is markedly inhibited in the presence of caspase inhibitors, indicate that an apoptotically activated ribonuclease is responsible for the specific modification of U1 snRNA during apoptosis. PMID- 10713723 TI - Fas-independent apoptosis in T-cell tumours induced by the CD2-myc transgene. AB - Depending on the cellular context, the Myc oncoprotein is capable of promoting cell proliferation or death by apoptosis. These observations suggest that apoptosis in response to deregulated gene expression may represent a natural brake to tumour development. The pathways by which Myc induces apoptosis are as yet poorly characterised although recent observations on rat fibroblasts over expressing Myc have demonstrated a requirement for the Fas pathway. To investigate the role of Fas in Myc-induced lymphomagenesis we backcrossed CD2-myc mice onto an lpr background. Rates of tumour development and phenotypic properties, including levels of apoptosis were indistinguishable from CD2-myc controls. Further, tumour cell lines derived from mice expressing a regulatable form of Myc showed inducible apoptosis at similar rates regardless of their lpr genotype. These results show that activation of c-myc and loss of Fas do not collaborate in T lymphoma development and that Myc-induced apoptosis in T-cells occurs by Fas-independent pathways. PMID- 10713724 TI - The timing of drosophila salivary gland apoptosis displays an l(2)gl-dose response. AB - During Drosophila metamorphosis, larval tissues, such as the salivary glands, are histolysed whereas imaginal tissues differentiate into adult structures forming at eclosion a fly-shaped adult. Inactivation of the lethal(2)giant larvae (l(2)gl) gene encoding the cytoskeletal associated p127 protein, causes malignant transformation of brain neuroblasts and imaginal disc cells with developmental arrest at the larval-pupal transition phase. At this stage, p127 is expressed in wild-type salivary glands which become fully histolysed 12 - 13 h after pupariation. By contrast to wild-type, administration of 20-hydroxyecdsone to l(2)gl-deficient salivary glands is unable to induce histolysis, although it releases stored glue granules and gives rise to a nearly normal pupariation chromosome puffing, indicating that p127 is required for salivary gland apoptosis. To unravel the l(2)gl function in this tissue we used transgenic lines expressing reduced ( approximately 0.1) or increased levels of p127 (3.0). Here we show that the timing of salivary gland histolysis displays an l(2)gl-dose response. Reduced p127 expression delays histolysis whereas overexpression accelerates this process without affecting the duration of third larval instar, prepupal and pupal development. Similar l(2)gl-dependence is noticed in the timing of expression of the cell death genes reaper, head involution defective and grim, supporting the idea that p127 plays a critical role in the implementation of ecdysone-triggered apoptosis. These experiments show also that the timing of salivary gland apoptosis can be manipulated without affecting normal development and provide ways to investigate the nature of the components specifically involved in the apoptotic pathway of the salivary glands. PMID- 10713725 TI - Bcl-2 inhibits Bax translocation from cytosol to mitochondria during drug-induced apoptosis of human tumor cells. AB - The pro-apoptotic protein, Bax, has been reported to translocate from cytosol to mitochondria following exposure of cells to apoptotic stresses including cytokine withdrawal and treatment with glucocorticoids and cytotoxic drugs. These observations, coupled with reports showing that Bax causes the release of mitochondrial cytochrome c, implicate Bax as a central mediator of the apoptotic process. In this report we demonstrate by subcellular fractionation a significant shift in Bax localization from cytosol to cellular membranes in two human tumor cell lines exposed to staurosporine or etoposide. Immunofluorescence studies confirmed that Bax specifically relocalized to the mitochondria. This redistribution of Bax occurred in concert with, or just prior to, proteolytic processing of procaspase-3, activation of DEVD-specific cleavage activity and degradation of poly(ADP-ribose) polymerase. However, Bax membrane translocation was independent of caspase activity as determined using the broad-range caspase inhibitor z-VAD-fmk. High level overexpression of the anti-apoptotic protein Bcl 2 prevented Bax redistribution to the mitochondria, caspase activation and apoptosis following exposure to staurosporine or etoposide. These data confirm the role of Bax in mitochondrial cytochrome c release, and indicate that prevention of Bax translocation to the mitochondrial membrane represents a novel mechanism by which Bcl-2 inhibits drug-induced apoptosis. PMID- 10713726 TI - Inhibition of phosphoinositide 3-kinase impairs pre-commitment cell cycle traverse and prevents differentiation in erythroleukaemia cells. AB - During the early hours after exposure to differentiation inducing agents, Friend erythroleukaemia cells undergo alterations which commit them to cessation of growth and development of the characteristics of differentiation. Our current experiments have compared the expression and activity of phosphoinositide 3 kinase (PI 3-kinase) in control cells with cells undergoing differentiation which has been induced by dimethyl sulfoxide (DMSO). When the cultures were initiated with stationary phase cells and DMSO was added at the time of seeding, PI 3 kinase activity was stimulated in both treated and control cells during the first 3 h from seeding. This event appears to be a rate limiting step in commitment since pretreatment of cells with 10 microM LY294002 or down-regulation of p85 expression prior to adding DMSO completely prevents commitment to erythropoiesis. Accordingly, PI 3-kinase inhibition during the commitment period prevents DNA binding of the transcription factor GATA-1, essential for erythroid differentiation. However, once cells are committed to differentiate, PI 3-kinase activity and expression dramatically decreases along with the differentiation programme, to become barely detectable after 96 h. Remarkably, LY294002 treatment leads to accumulation of cell in G1 phase and prevents DMSO-dependent cyclin D3 induction. Based on these data, we suggest that PI 3-kinase is rate limiting for the completion of the first round cycle of cell division required for initiation of erythrocytic differentiation. On the other hand, the late decrease of PI 3 kinase associated with the differentiation process seems to be part of the programmed shut off of genes not needed in mature erythrocytes. PMID- 10713727 TI - Up-regulation of Bcl-2 by redox signals in glomerular mesangial cells. AB - The mediators nitric oxide (NO) and superoxide (O2-) are known to regulate cell death and survival. In mesangial cells (MC), NO induced apoptosis and in higher concentrations necrosis. Intriguingly, cogeneration of NO and O2- in a balanced ratio promoted cell protection. Under these conditions, we noticed the accumulation of the anti-apoptotic protein Bcl-2. Its up-regulation is based on an increase in mRNA and protein level. To investigate whether oxidative stress elicits Bcl-2 expression in general, we further used the chemically unrelated oxidative agents diamide and butyl hydroperoxide. Both stimulated mRNA and protein up-regulation of Bcl-2. But in contrast to diamide, butyl hydroperoxide evoked apoptosis and necrosis despite Bcl-2 accumulation. As diamide was non toxic, we used diamide as a Bcl-2 activator to protect MC against a subsequent toxic dose of NO. We conclude that redox changes promote Bcl-2 up-regulation that may confer cellular protection towards apoptosis. PMID- 10713728 TI - Mass spectrometric identification of proteins released from mitochondria undergoing permeability transition. AB - Mitochondrial membrane permeabilization is a rate-limiting step of cell death. This process is, at least in part, mediated by opening of the permeability transition pore complex (PTPC) Several soluble proteins from the mitochondrial intermembrane space and matrix are involved in the activation of catabolic hydrolases including caspases and nucleases. We therefore investigated the composition of a mixture of proteins released from purified mitochondria upon PTPC opening. This mixture was subjected to a novel proteomics/mass spectrometric approach designed to identify a maximum of peptides. Peptides from a total of 79 known proteins or genes were identified. In addition, 21 matches with expressed sequence tags (EST) were obtained. Among the known proteins, several may have indirect or direct pro-apoptotic properties. Thus endozepine, a ligand of the peripheral benzodiazepin receptor (whose occupation may facilitate mitochondrial membrane permeabilization), was found among the released proteins. Several proteins involved in protein import were also released, namely the so-called X linked deafness dystonia protein (DDP) and the glucose regulated protein 75 (grb75), meaning that protein import may become irreversibly disrupted in mitochondria of apoptotic cells. In addition, a number of catabolic enzymes are detected: arginase 1 (which degrades arginine), sulfite oxidase (which degrades sulfur amino acids), and epoxide hydrolase. Although the functional impact of each of these proteins on apoptosis remains elusive, the present data bank of mitochondrial proteins released upon PTPC opening should help further elucidation of the death process. PMID- 10713729 TI - Long-term fate of terminally differentiated skeletal muscle cells following E1A initiated cell cycle reactivation. AB - We have previously shown that E1A reactivates the cell cycle in 'irreversibly' growth arrested, terminally differentiated (TD) cells. The molecular events following E1A-mediated reactivation of TD skeletal muscle cells have been extensively investigated. However, the long-term fate of the reactivated cells has not been directly determined. In this paper, E1A is used to reactivate TD myotubes derived from established cell lines or primary myoblasts. We show that the reactivated muscle cells continue proliferating beyond the end of the first cell cycle and progress through at least a second one. Experiments performed with an inducible E1A/estrogen receptor chimera indicate that the reactivated cell cycle is self-sustained, since E1A is exclusively necessary to reactivate TD cells, but is dispensable for both the continuation of the first cycle and the progression into the following one. Finally, we report that E1A-mediated reactivation of muscle cells results in apoptotic cell death that can be delayed by the antiapoptotic, adenoviral E1B 55 kDa oncogene. PMID- 10713730 TI - Subcellular localization and CARD-dependent oligomerization of the death adaptor RAIDD. AB - RAIDD, a caspase recruitment domain (CARD) containing molecule, interacts with procaspase-2 in a CARD-dependent manner. This interaction has been suggested to mediate the recruitment of caspase-2 to the tumour necrosis factor receptor 1 (TNFR1). In this paper we have studied the subcellular localization of RAIDD and its interaction with caspase-2. We demonstrate that endogenous RAIDD is mostly localized in the cytoplasm and to some extent in the nucleus. RAIDD localization is not affected by TNF-treatment of HeLa cells, but in cells ectopically expressing caspase-2, a fraction of RAIDD is recruited to the nucleus. In transfected cells, coexpression of RAIDD and caspase-2 leads to CARD-dependent colocalization of the two proteins to discrete subcellular structures. We further show that overexpression of the RAIDD-CARD results in the formation of filamentous structures due to CARD-mediated oligomerization. These structures were similar to death effector filaments (DEFs) formed by FADD and FLICE death effector domains (DEDs), and partially colocalized with DEFs. Our results suggest that similar to the DED, the RAIDD-CARD has the ability to form higher order complexes, believed to be important in apoptotic execution. We also present evidence that RAIDD-CARD oligomerization may be regulated by intramolecular folding of the RAIDD molecule. PMID- 10713731 TI - Myofibroblast phenotype and apoptosis in keloid and palmar fibroblasts in vitro. AB - Keloid formation is a wound healing response, which fails to resolve and leads to formation of a raised collagen mass extending beyond the original wound margins. Keloids are typically excluded from palms and soles. Therefore we compared keloid and palmar fibroblasts in vitro using fibroblasts from nonaffected individuals as controls. Collagen I, alpha-smooth muscle actin and thrombospondin-1 were found at higher levels in keloid than in palmar fibroblasts. These differences were ameliorated by addition of TGFbeta1. The potential for resolution of the wound healing response was estimated analyzing apoptosis during serum starvation. Annexin V and TUNEL assays showed that palmar fibroblasts underwent faster apoptosis, than did the keloid fibroblasts, and started detaching. Addition of TGFbeta1 counteracted this effect. The weak expression of the myofibroblast phenotype and the advanced apoptosis of palmar fibroblasts suggest mechanisms for the exclusion of keloids from palmar sites. PMID- 10713732 TI - E1A is sufficient by itself to induce apoptosis independent of p53 and other adenoviral gene products. AB - Induction of apoptosis seems to be a key function in maintaining normal cell growth by exerting negative controls on cell proliferation and suppressing tumorigenesis. The adenovirus E1A oncogene shows both cell cycle progression and apoptotic functions. To understand the mechanism of E1A-induced apoptosis, the apoptotic function of E1A 13S was investigated in p53-null cells. We show here that E1A is sufficient by itself to induce substantial apoptosis independent of p53 and other adenoviral genes. The apoptotic function of E1A is accompanied by processing of caspase-3 and cleavage of poly(ADP-ribose)-polymerase. Cell death is significantly blocked by the caspase inhibitor zVAD-fmk and when coexpressed with E1B19K, Bcl-2 or the retinoblastoma protein (RB). Analyses of E1A mutants indicated that the apoptotic activity of E1A correlates closely with the ability to bind the key regulators of E2F1-induced apoptosis, p300 and RB. Finally, in vivo relevance of down-modulation of p53-independent apoptosis for efficient transformation is demonstrated. PMID- 10713733 TI - Cripto-1 induces apoptosis in HC-11 mouse mammary epithelial cells. AB - Cripto-1 (CR-1) is an epidermal growth factor (EGF)-related protein. CR-1 can inhibit beta-casein and whey acidic protein expression in mouse mammary epithelial cells. The present study demonstrates that CR-1 can induce apoptosis in HC-11 mouse mammary epithelial cells, as measured by bis-benzimide stained nuclei, TUNEL assay and cell death ELISA. Apoptosis could be observed after 2 days of exposure of confluent HC-11 cells to CR-1 in the absence of the survival factors EGF and insulin, with maximum apoptosis occurring at 3 days. A reduction in poly(ADP-ribose) polymerase (PARP) expression and an increase in beta-catenin cleavage was found after 18 h of exposure to CR-1 suggesting that apoptosis was preceded by the induction of a caspase activity since the caspase inhibitor ZFAD.FMK could block the CR-1-induced reduction in PARP expression and CR-1 induced apoptosis. CR-1 was found to increase the expression of caspase-3-like protease. Although, the levels of p27kip1 and p21Bax did not change after exposure to CR-1 for 18 h, the levels of Bcl-xL became undetectable. These studies suggest that CR-1 promotes apoptosis by mediating the induction of caspase-3-like protease and downregulating the expression of Bcl-xL. PMID- 10713734 TI - The topoisomerase inhibitors camptothecin and etoposide induce a CD95-independent apoptosis of activated peripheral lymphocytes. AB - The effect of etoposide and camptothecin, two topoisomerase inhibitors directed against topoisomerases II and I, respectively, was evaluated on human peripheral blood lymphocytes. Etoposide and camptothecin induced apoptosis of mitogen activated but not resting CD4+ and CD8+ T lymphocytes. Cell sensitivity to these agents required G1 to S-phase transition of the cell cycle. Conversely, daunorubicin, an intercalating agent and topoisomerase II inhibitor, induced apoptosis of both resting and activated lymphocytes. Although etoposide and camptothecin induced CD95-ligand mRNA expression, drug-induced apoptosis of activated human lymphocytes was not inhibited by CD95 antagonists. Drug-induced cell death was also not inhibited by p55 TNFR-Ig fusion protein. Activation of the caspases cascade was suggested by the partial inhibitory effect of the tripeptide zVAD-fmk and documented by activation of caspase 3. Finally etoposide and camptothecin induced a rapid production of ceramide in activated but not resting peripheral blood lymphocytes, suggesting that ceramide might initiate the signaling apoptotic cascade in sensitive cells. PMID- 10713735 TI - Ceramide selectively inhibits apoptosis-associated events in NGF-deprived sympathetic neurons. AB - Ceramide manifests both neurotoxic and neuroprotective properties depending on the experimental system. Ito and Horigome previously reported that ceramide delays apoptosis in a classic model of developmental programmed cell death, i.e. sympathetic neurons undergoing NGF deprivation.1 Here, we investigated the actions of ceramide upon the biochemical and genetic changes that occur in NGF deprived neurons. We correlate ceramide's neuroprotective actions with the ability of ceramide to antagonize NGF deprivation-induced oxidative stress and c jun induction, both of which contribute to apoptosis in this model. However, ceramide did not block NGF deprivation-induced declines in RNA and protein synthesis, suggesting that ceramide does not slow all apoptosis-related events. Overall, these results are significant in that they show that ceramide acts early in the death cascade to antagonize two events necessary for NGF-deprivation induced neuronal apoptosis. Moreover, these results dissociate declines in neuronal function, i.e. macromolecular synthesis, from the neuronal death cascade. PMID- 10713736 TI - Apoptotic response to growth factor deprivation involves cooperative interactions between c-Fos and p300. AB - Two preneoplastic cell lines have been utilized to study changes in the regulation of apoptosis during neoplastic progression [sup+I (stage I) and sup-II (stage II)]. Sup+I cells are prone to undergo apoptosis, while sup-II cells are relatively resistant. We report that induction of apoptosis in sup+I cells is tightly correlated with the formation of c-Fos/p300 complexes, which were not present in the non-apoptotic sup-II cells under the same conditions. When apoptosis was induced in the sup-II cells by over-expression of c-Fos, concomitant c-Fos:p300 complexes were detected. Over-expression of p300 resulted in apoptosis in sup-II cells and also in p53wt human tumor cells, but not in p53mutant human tumor cells. Over-expression of the C-terminal fragment of p300, which contains the c-Fos binding site, enhanced apoptosis, suggesting that the c Fos:p300 complex is actively involved in apoptosis. We propose that p300 could function as a general mediator of transcription factor-induced apoptosis. PMID- 10713737 TI - Distinct stages of cytochrome c release from mitochondria: evidence for a feedback amplification loop linking caspase activation to mitochondrial dysfunction in genotoxic stress induced apoptosis. AB - Cytochrome c (cyto c) release from mitochondria is a critical event in apoptosis. By investigating the ordering of molecular events during genotoxic stress-induced apoptosis, we found that ionizing radiation (IR) and etoposide induced the release of cyto c from mitochondria in two distinct stages. The early release of low levels of cyto c into the cytosol preceded the activation of caspase 9 and 3, but had no effect on ATP levels or mitochrondrial transmembrane potential (Deltapsim). In contrast, the late stage cyto c release resulted in a drastic loss of mitochondrial cyto c and was associated with reduction of ATP levels and Deltapsim. Moreover, caspases contributed to the late cyto c release since the caspase inhibitor zVAD prevented only the late but not the early-stage cyto c release. Recombinant caspase 3 induced cyto c release from isolated mitochondria in the absence of cytosolic factors. Bcl-2 but not Bid was cleaved during apoptosis after caspase activation. This suggests that Bcl-2 cleavage might contribute to the late cyto c release, which results in mitochondrial dysfunction manifested by the decrease of ATP and Deltapsim. zVAD prevented the reduction of ATP, Deltapsim, and nuclear condensation when added up to 8 h after IR, at the time the caspases were highly activated but when the majority of cyto c was still maintained in the mitochondria. These findings link the feedback loop control of caspase-induced cyto c release with mitochondrial dysfunction manifested by ATP and Deltapsim decline. PMID- 10713738 TI - Comparison of diets of diabetic and non-diabetic elderly men in Finland, The Netherlands and Italy. AB - OBJECTIVE: To evaluate whether dietary recommendations for subjects with diabetes are met among Finnish, Dutch and Italian elderly men with diabetes, and whether the diets of diabetic and non-diabetic men differ in these three countries. DESIGN: A dietary survey using cross-check dietary history method. A cross sectional comparison. SETTING: Thirty-year follow-up of survivors from the Finnish, Dutch and Italian cohorts of the Seven Countries Study. SUBJECTS: 227 elderly men from Finland, 537 from The Netherlands, and 417 from Italy, of whom 8 9% had diabetes. MAIN RESULTS: The diets of non-diabetic men from the three countries differed markedly from each other. In all three countries diabetic men consumed less added sugar than non-diabetic men. In Italy, in addition, diabetic men consumed more fruits and berries and vegetables. The Dutch diabetic men ate relatively more cereal products, fruits and berries, milk and milk products, cheese, and meat and meat products and drank less alcoholic beverages than non diabetic men. The diet of both diabetic and non-diabetic Finnish and Dutch men was characterized by high fat content (41% and 40% of energy, respectively). The fat content of the diet was even higher for diabetic than non-diabetic men in Finland and The Netherlands, but not in Italy. The fibre content of the diet was the highest among Dutch men and diabetic men received more dietary fibre than non diabetic men in The Netherlands and Italy, but not in Finland. The diet of diabetic and non-diabetic Finnish men differed little from each other and was characterized by high nutrient density of several vitamins and minerals. The proportion of protein of energy intake was higher among diabetic than non diabetic Dutch and Italian men. CONCLUSIONS: The diet of the diabetic men from Finland, the Netherlands, and Italy resembled more the diet of non-diabetic men from the respective countries than the diet of diabetic men from the other countries. In the diet of Italian diabetic men, the proportions of fat, saturated fatty acids and carbohydrates were nearest the recommended levels. SPONSORSHIP: The National Institute on Aging, Bethesda, USA, the Dutch Prevention Foundation, the Hague, The Netherlands, the Academy of Finland, and the Sandoz Gerontological Foundation. PMID- 10713739 TI - Nutrient intake of working women in Bangkok, Thailand, as studied by total food duplicate method. AB - OBJECTIVES: To establish a general view of food habits in Thailand, and to make a quantitative assessment of rice dependency of Thai people. DESIGN: Cross sectional study. SETTING: Community. SUBJECTS: 52 non-smoking and non-habitually drinking adult women in Bangkok participated in the study. METHODS: The participants offered 24 h food duplicates and peripheral blood samples, and underwent clinical examination including anthropometry. The duplicates were subjected to nutritional evaluation taking advantage of the Thai food composition tables (FCTs), and analyzed for eight nutrient elements by inductively coupled plasma mass spectrometry (ICP-MS). RESULTS: The participants took 1630 kcal from 55 g protein (63% from animal sources), 57 g lipid (mostly from vegetable oil), and 224 g carbohydrate (60% from rice) daily. Nutrient intake at lunch was as large as that at dinner. About a half of the women had insufficient energy intake (ie <80% RDA) whereas 4% had an excess (>120%). Protein intake was sufficient in most cases, whereas lipid intake was in excess in more than a half of the women. Ca, Fe, Mg, Zn and possibly P intakes were below the RDA values in many participants. FCT-based estimates agreed well with the ICP-MS measures in cases of Fe and Ca but tended to be greater than the measures by 50% with regard to P. CONCLUSIONS: Lunch as substantial as dinner for Thai urbanites. There was a marked dependency on rice as an energy source. Whereas protein intake is generally sufficient, the intake of Ca (and to a lesser extent Fe) was insufficient in a majority of the study participants. SPONSORSHIP: Dai-ichi Mutual Life Insurance, Japan; the Ministry of Health and Welfare, the government of Japan. PMID- 10713740 TI - Total daily energy expenditure and pattern of physical activity measured by minute-by-minute heart rate monitoring in 14-15 year old Swedish adolescents. AB - OBJECTIVE: To assess total daily energy expenditure (TDEE) and patterns of physical activity among Swedish male and female adolescents and to relate the amount and intensity of physical activity to existing recommendations (energy expenditure equal to or above 12.4 kJ/kg/day or accumulation of 30 min/day in moderate physical activity equal to 4.5 times sedentary energy expenditure or more). DESIGN: TDEE, physical activity level (PAL=TDEE/BMR), energy expenditure (EE) and time spent in different intensities of physical activity were assessed by using minute-by-minute heart rate monitoring in combination with laboratory measured sedentary energy expenditure (SEE) and peak oxygen uptake. SETTING: Department of Physical Education and Health, Orebro University, and Department of Clinical Physiology, Orebro Medical Centre Hospital, Sweden. SUBJECTS: Eighty-two 14-15 y old adolescents (42 boys, 40 girls) from the city of Orebro, randomly selected through a two-stage sampling procedure. RESULTS: TDEE was 12.8 MJ/day and 10.0 MJ/day for boys and girls respectively (P<0.001) and PAL was 1.74 and 1.67 (NS). Forty-four percent and 47%, respectively, of TDEE referred to EE in physical activity, of which 70% for both genders referred to light physical activity (corresponding to <4.5 times SEE). Eleven boys and 14 girls had an EE lower than 12.4 kJ/kg/day and/or did not accumulate 30 min/day in physical activity >/=4.5 SEE. Those (n=20) with the highest PAL values (>2.01 and 1.81, respectively) spent 149 min/day at a >/=4.5 SEE intensity level compared to 40 min/day for those (n=30) with the lowest PAL values (<1.55 and 1.45, respectively). CONCLUSIONS: Swedish adolescent boys and girls are similarly physically active. The major amount of time devoted to physical activity refers to light physical activity. At least thirty percent of adolescents seem not to achieve appropriate levels of physical activity considered to be beneficial for health. SPONSORSHIP: Orebro County Council, The Public Health Committee of Stockholm County Council, Sweducation Foundation. PMID- 10713741 TI - Puberty begins with a characteristic subcutaneous body fat mass in each sex. AB - OBJECTIVE: To observe whether there exists a characteristic body fat mass at pubertal onset. DESIGN: Longitudinal clinical follow-up (between ages of 10 and 15 y) with an annual visit in a sample of 469 children. They were grouped according to age of purbertal onset: boys with pubertal onset at the ages of 11 (n=59), 12 (n=88), 13 n=89) and 14 y (n=46), and girls with pubertal onset at the age of 10 (n=68), 11 (n=66), 12 (n=37) and 13 (n=16). METHODS: Height, weight, upper arm circumference and four skinfold thicknesses were recorded annually. In boys testicular volume index was measured, and genital development was assessed on the Tanner scale; in girls mammary development was measured also using the Tanner scale. The sum of four skinfolds, body mass index, upper arm fat estimate and percentage body fat were calculated. RESULTS: Boys presented a positive relation between the age of pubertal onset and body mass index (P<0.001), which was not observed in girls. Body mass index thus varied according to the onset of puberty in boys (P<0.001), but not in girls. The sum of four skinfolds, the upper arm fat estimate index and the percentage of body fat mass did not differ according to age of pubertal onset either in girls (P=NS) or in boys (P=NS). The characteristic adiposity of the puberty onset is progressively acquired during the previous years in all the groups. CONCLUSIONS: Puberty seems to begin with a characteristic subcutaneous body fat mass that is independent of the age of onset. This study supports the hypothesis of a close link between maturation and the development of an energy store in the form of adipose tissue in both sexes. PMID- 10713742 TI - Effect of oral iodized oil on thyroid size and thyroid hormone metabolism in children with concurrent selenium and iodine deficiency. AB - OBJECTIVES: To determine the efficacy of oral iodized oil in goitrous children who are both selenium (Se) and iodine deficient; to investigate if Se status modifies the response of iodine deficient, goitrous children to oral supplementation with iodized oil. DESIGN: A longitudinal intervention trial. SETTING: Two rural villages in the western Cote d'Ivoire. SUBJECTS: 51 goitrous non-anemic schoolchildren with both iodine and Se deficiency. INTERVENTION: Each child received an oral dose of 0.4 ml iodized poppyseed oil containing 200 mg of iodine. They were followed for 1 y with measurements of urinary iodine (UI), thyrotropin (TSH), thyroxine (T4), and thyroid volume by ultrasound. RESULTS: At baseline all children were goitrous and Se deficient; median UI was 29 microg/l and mean serum Se (s.d.) was 14.8 (10.7) microg/l. After receiving iodized oil, thyroid volume decreased significantly vs baseline at 10, 15, 30 and 50 weeks (P<0.001). At 50 weeks mean percentage change in thyroid volume from baseline was 46.6% and only five children remained goitrous. Median TSH values at 5, 10, 15, 30 and 50 weeks were reduced significantly (P<0.001) compared to baseline. Among individual children the severity of Se deficiency predicted the degree of response to iodized oil. Baseline serum Se and percentage change in thyroid volume from baseline at 50 weeks were strongly correlated (r2=0.554). Baseline Se and percentage decrease in TSH from baseline at 30 weeks were also well correlated (r2=0.467). CONCLUSION: Although more severe Se deficiency partially blunts the thyroid response to iodine supplementation, oral iodized oil is an effective method for iodine repletion in goitrous children who are Se deficient. SPONSORSHIP: The Swiss Federal Institute of Technology, Zurich, the Foundation for Micronutrients in Medicine, Rapperswil, Switzerland, and the Thrasher Research Fund, Salt Lake City, USA. PMID- 10713743 TI - Body mass index in children and adolescents according to age and pubertal stage. AB - OBJECTIVE: To evaluate the dependence of body mass index (BMI) values on pubertal stage in subjects similar in age. DESIGN, SUBJECTS AND MEASUREMENTS: Height and weight were recorded cross-sectionally in school subjects from three provinces in central Italy. The subjects were subdivided into three groups: (1) 4271 school subjects (2125 males and 2146 females; 8.5-15.5 y old), in whom the pubertal development was also recorded, were selected to subdivide BMI values according to pubertal stage and age; (2) 6345 females (10.5-14.5 y old), who were asked whether or not they had had their first menstrual period, were selected to subdivide BMI values according to age in pre-menarche and post-menarche girls, separately; and (3) 1919 females (10.5-14.5 y old), who had presented their menarche within the previous 6 months, were selected to subdivide short-term post menarche BMI values according to age. RESULTS: The medians and interquartile ranges of BMI varied according to age and pubertal stage. Kruskall-Wallis test performed in subjects similar in age demonstrated that significant differences existed among the medians of BMI values of subjects at different pubertal stages in 12-14-y-old males (P<0.05), and in 11-14-y- old females (P<0.001). The difference also proved to be significant between stage I and stage II (P<0.05) in 10-y-old females, but not in 10-11-y-old males. The Kruskal-Wallis test performed in subjects similar in pubertal stage demonstrated that significant differences among the medians of BMI at different ages existed only in females at stages II and III. A significant positive trend was observed in both genders according to pubertal stage for BMI values of subjects similar in age (z-test for trend, P<0.01). On the contrary, a negative age trend proved to be significant in females at stages I (P<0.01), II (P<0.01) and III (P<0.001), but not in males when the subdivision of BMI was made according to age in subjects similar in pubertal stage. BMI values were significantly higher in post-menarche girls as compared to pre-menarche girls similar in age (P<0.001). However, at partial regression analysis BMI values were influenced by pubertal stage and, to a lesser extent, by age, but not by menarcheal status. An inverse association between short-term post-menarche BMI and age was observed, with the highest values in girls presenting menarche at 11 y of age (P<0.05). The negative trend was demonstrated at the z-test for trend (P<0.001). CONCLUSIONS: BMI values depend on pubertal degree of maturation, especially in girls. This influence should be taken into account when BMI is evaluated in adolescents. SPONSOR: University of Perugia, Region of Umbria. PMID- 10713744 TI - Skeletal muscle protein mass correlates with the lipid status in children with solid tumors and before bone marrow transplantation. AB - OBJECTIVE: To evaluate the changes in lipid status in children during anticancer therapy, with special reference to the effect of protein-energy malnutrition on plasma lipids. DESIGN: Prospective follow-up study. SETTING: The study was carried out in the Hospital for Children and Adolescents, University of Helsinki, Helsinki, Finland SUBJECTS: The study group consisted of 33 children going through bone marrow transplantation (BMT) and 10 children with malignant solid tumors. The BMT patients were evaluated before transplantation and 1 and 3 months after BMT, and the tumor patients were studied at diagnosis and in remission. The reference group consisted of 23 healthy children. INTERVENTIONS: As indicators of lipid status, lipoproteins and the concentration of cholesterol and triacylglycerol were measured. Protein reserves were expressed as muscle index (MI), derived from ultrasonographic measurement of the femoral quadriceps muscle. Body weight, triceps skinfold thickness and the serum concentration of albumin, prealbumin and transferrin were measured. RESULTS: In both groups, plasma concentration of total triacylglycerol was increased and high-density lipoprotein (HDL) cholesterol decreased as compared to the reference subjects. Plasma triacylglycerol concentration had a negative correlation with skeletal muscle protein mass (MI; r=0.34, P=0.02). The concentration of serum prealbumin correlated positively with plasma total cholesterol concentration (r=0.47, P=0.002). CONCLUSIONS: In children with cancer, abnormalities of lipid status are associated with changes in skeletal muscle protein reserves. SPONSORSHIP: This study was supported by the Foundation of Pediatric Research, Helsinki, Finland and the Nona and Kullervo Vare Foundation, Helsinki, Finland. PMID- 10713745 TI - Appetite dysfunction in obese males: evidence for role of hyperinsulinaemia in passive overconsumption with a high fat diet. AB - OBJECTIVE: To test the hypothesis that caloric and fat intake in a pre-load meal have no subsequent effects upon blood glucose and insulin concentrations, perceived hunger, subsequent food intake and appetite control in lean and obese men. DESIGN: Lean and obese men reported to the laboratory in the morning in a fasted state where they were subject to an eating test based on the pre-load-test meal paradigm, using a double-blind protocol. The breakfast pre-load was either a reduced caloric low-fat (LF) meal or an overfeeding high-fat (HF) meal. LF was 20% of each individual's average daily energy requirement (ADER) and comprised 60% carbohydrate, 27% protein, and 13% fat, whilst HF was adjusted to yield 55% of the ADER, and comprised 45% carbohydrate, 22% protein and 43% fat. The pre loads on both trials were administered as one single mean, and were given in a random order. After 5(1/2) h, an ad libitum test-lunch was given to determine how much energy was consumed. Between the two meals, blood samples were collected and subjective hunger ratings were assessed hourly. These variables were measured at 30-min intervals for 75 min after the ad libitum meal. STUDY PARTICIPANTS: Twelve healthy men, six of whom were lean (BMI 22. 50+/-1.08 kg.m2) and six of whom were obese (BMI 39.05+/-11.63 kg. m2) were recruited. RESULTS: When given 55% of their ADER in a HF pre-load meal, the obese group consumed more energy (5426+/-1126 kJ; F1,20=11.45, P<0.01), than the lean group did (3473+/-1114 kJ), accounting for 45% of the ADER in that meal setting. However, no differences between lean and obese intake were noted at the test meal following a LF pre-load. The lean group exhibited a significant inverse correlation (r=0.628, P<0.05) between serum insulin concentration before eating the test meal and the amount of energy consumed at the test meal, while such a relationship was absent in the obese group. CONCLUSION: The obese males were unable to compensate for the caloric overloading when fed a HF (55% ADER) pre-load at a subsequent test meal, whereas a calorically reduced pre-load (20% ADER) produced similar intakes to the lean control group. The inverse relationship noted in the lean group between insulin levels before the test meal and the energy intake at that test meal suggests that insulin may play a role in the regulation of appetite - satiety mechanism in lean males. The absence of such a relationship in the obese may suggest the site for possible appetite dysfunction contributing to obesity. These results further suggest that when obese individuals consume a high-fat meal they are prone to passive overconsumption, whereas lean study participants appear to be more resistant to such a phenomenon. PMID- 10713746 TI - Essential fatty acids in breast milk of atopic mothers: comparison with non atopic mothers, and effect of borage oil supplementation. AB - OBJECTIVE: To evaluate whether levels of n-6 long chain polyunsaturated fatty acids (LCPs) in human breast milk are related to the mother's atopic constitution, and whether a decreased level can be restored by gamma-linolenic acid supplementation. DESIGN: Cross-sectional study and dietary supplementation trial. SUBJECTS: 20 atopic mothers and 20 non-atopic mothers (controls), all lactating. SETTING: General population. INTERVENTIONS: The atopic mothers were randomly assigned to low (n=10) or high (n=10) dosage oral supplementation with oral borage oil for one week (230 or 460 mg gamma-linolenic acid (18:3n-6) per day). MAIN OUTCOME MEASURES: Essential fatty acid composition of the breast milk total fat fraction, determined by gas liquid chromatography. RESULTS: Arachidonic acid (20:4n-6) was lower in breast milk of atopic mothers compared with non atopic mothers (0.39 wt% vs 0.46 wt%, difference -0.07% wt% (95% confidence limits -0.13, -0.01 wt%; P<0. 05). The ratio between linoleic acid and the sum of n-6 derivatives did not differ between these groups, indicating no difference in delta-6-desaturase (D6D) activity. Supplementation of the atopic mothers significantly increased the levels of gamma-linolenic acid and dihomo-gamma linolenic acid in breast milk in a dose-related way, but the level of arachidonic acid was not increased. CONCLUSION: We found a decreased level of arachidonic acid in breast milk in atopic compared to non-atopic mothers, but no indication that the rate-limiting enzymatic step (D6D) is involved. Supplementation increased the precursor pool but did not restore the level of arachidonic acid. We conclude that atopy is related to a metabolic disturbance beyond the D6D enzymatic step. A low level of arachidonic acid in breast milk may be a risk factor for the development of atopy in the infant, especially when the possible underlying metabolic disturbance of EFA metabolism is inherited by the child. SPONSORSHIP: F Hoffman-La Roche (Basel, Switzerland) and Friesland Dairy Foods (Leeuwarden, The Netherlands). PMID- 10713747 TI - Short-term regulation of food intake in children, young adults and the elderly. AB - OBJECTIVE: The present study investigated the effects of consuming preloads with different macronutrient and energy contents on subsequent intake and subjective feelings of hunger and satiety in children, young adults and the elderly. SUBJECTS: 30 Children (4-6 y), 33 young adults (18-26 y) and 24 elderly (61-86 y). DESIGN: A 'preload test meal' design was applied. Subjects were given four different strawberry yoghurt preloads that varied in energy and macronutrient content, or no yoghurt. Children, young adults and elderly consumed 200, 340 and 300 g of the preload, respectively. One yoghurt was low-fat, low-carbohydrate and low in energy (the control; 0.7 MJ/500 g serving), one yoghurt was high-fat and medium in energy (71% of energy (en%) of fat; 2 MJ/500 g serving), one yoghurt was high-carbohydrate and medium in energy (87 en% of carbohydrate; 2 MJ/500 g serving) and the fourth yoghurt was high-fat and high-carbohydrate and high in energy (42 en% of fat and 53 en% of carbohydrate; 3 MJ/500 g serving). Ninety minutes after preload consumption, subjects had an attractive ad libitum lunch buffet. Energy intake at lunch and subjective feelings of hunger and satiety were analysed. RESULTS: The ability to compensate at lunch did not differ among the three age groups. Compared to the no-preload condition, all children, young adults and elderly ate significantly less after the high-fat and high carbohydrate yoghurt. The energy compensation observed in the children ranged between -21% and 34%, in the young adults between 15% and 44% and in the elderly between 17% and 23%. Hunger responses were clearly different between young adults and the elderly. Compared to the no-preload condition, the young adults showed larger differences in their appetite ratings than the elderly, indicating that the elderly were less sensitive to the energy content of the preload than the young adults. CONCLUSION: We conclude that the ability to regulate the food intake within a preload 90 min test meal paradigm did not differ among children, young adults and the elderly. SPONSORSHIP: This study was funded by the European Commission as part of project FAIR-CT95-0574. PMID- 10713748 TI - Obesity in women from developing countries. AB - OBJECTIVES: The key objective was to estimate obesity (>/=30 kg/m2) in women 15 49 y from developing countries. A second objective was to study how obesity varies by educational level and by residence in urban and rural areas. A third objective was to investigate how national incomes shape the relationship between obesity and eduction or residence. DESIGN: The analyses use cross-sectional data from nationally representative surveys from developing countries carried out in the last decade. Most of the surveys were Demographic Health Surveys (DHS). Data from a survey from the USA are used for comparison. SETTING: The 39 surveys used come from 38 developing countries and the USA. SUBJECTS: A total of 147,938 non pregnant women 15-49 y were included in the analyses. RESULTS: The percentage of obese women was 0.1% in South Asia, 2.5% in Sub-Saharan Africa, 9. 6% in Latin America and the Caribbean, 15.4% in Central Eastern Europe/Commonwealth of Independent States (CEE/CIS), 17.2% in the Middle East and North Africa, and 20.7% in the USA. Levels of obesity in countries increased sharply until a gross national product of US$1500 per capita (1992 values) was reached and changed little thereafter. In very poor countries, such as in Sub-Saharan Africa, obesity levels were greatly concentrated among urban and higher educated women. In more developed countries, such as those in Latin America and the CEE/CIS regions, obesity levels were more equally distributed in the general population. CONCLUSIONS: Based on the analyses presented and on a review of the literature, it is concluded that obesity among women is a serious problem in Latin America and the Caribbean, the Middle East and North Africa, and the CEE/CIS region. Obesity is less of a concern in Sub-Saharan Africa, China and South Asia. Obesity levels increased over time in most of the limited number of countries with data, but at varying rates. Rising national incomes in developing countries and increased 'Westernization' will most likely lead to increased levels of obesity in the future. SPONSORSHIP: Financial support was provided by the Food and Nutrition Program of the Pan American Health Organization and by the World Bank. PMID- 10713749 TI - Dietary patterns in six european populations: results from EURALIM, a collaborative European data harmonization and information campaign. AB - OBJECTIVE: To determine and describe the extent to which European dietary data collected in disparate surveys can be meaningfully compared. DESIGN: Seven independent population-based surveys from six European countries were initially included. Differences in study designs and methodological approaches were examined. Risk factor data for 31,289 adults aged 40-59 y were harmonized and pooled in a common, centralized database. RESULTS: Direct comparisons of dietary measures across studies were not deemed appropriate due to methodological heterogeneity. Nonetheless, comparisons of intra-population contrasts by gender across sites were considered valid. Women consumed fruit and vegetables more often than men. Age-standardized gender differences in the prevalence of low fruit and vegetable consumption ranged from 7 to 18% and 5 to 15%, respectively. Data on energy intake showed good agreement across study populations. The proportion of total energy from macronutrients was similar for women and men. Gender differences for relative intakes of saturated fatty acids (percentage energy) were small and only in France were they significant. Dietary fibre density was significantly higher in women than in men. Overall, the participating Southern European populations from Italy and Spain exhibited more healthful food composition patterns. CONCLUSIONS: Contrasts in dietary patterns by gender across populations may provide the basis for health promotion campaigns. The most favourable patterns observed may serve as attainable goals for other populations. An international risk factor surveillance programme based upon locally run, good quality studies has the potential to provide the needed data. SPONSORSHIP: European Community (DG V), project 96CVVF3-446-0; Swiss Federal Office for Education and Science, OFES 96.0089. PMID- 10713750 TI - Enhancement of natural immune function by dietary consumption of Bifidobacterium lactis (HN019). AB - OBJECTIVE: To determine the effects of dietary consumption of Bifidobacterium lactis (strain HN019, DR10TM) on natural immunity. DESIGN: A randomized, double blind, placebo-controlled clinical trial. SETTING: Janeway Medical Centre, Memorial University, St Johns, Newfoundland. SUBJECTS: Twenty-five healthy elderly volunteers (median age 69 y; range 60-83 y). INTERVENTIONS: Twelve control subjects consumed 180 ml low-fat/low-lactose milk twice daily for a period of 6 weeks; 13 test subjects consumed milk supplemented with 1.5x1011 colony-forming units of B. lactis twice daily. Indices of natural immunity, including interferon production, phagocytic capacity and phagocyte-mediated bactericidal activity, were determined via peripheral blood at 0, 3, 6 and 12 weeks post-trial commencement. RESULTS: Subjects who consumed milk containing B. lactis for 6 weeks produced significantly enhanced levels of interferon-alpha, upon stimulation of their peripheral blood mononuclear cells in culture, in comparison to the placebo control group who received milk alone. There were also significant increases in polymorphonuclear cell phagocytic capacity among test group subjects, following consumption of milk supplemented with B. lactis, while individuals who consumed B. lactis-supplemented milk or milk alone showed enhanced phagocyte-mediated bactericidal activity. CONCLUSIONS: The results demonstrate that dietary consumption of B. lactis HN019 can enhance natural immunity in healthy elderly subjects, and that a relatively short-term dietary regime (6 weeks) is sufficient to impart measurable improvements in immunity that may offer significant health benefits to consumers. SPONSORS: Financial support for this project was provided by the New Zealand Dairy Board. PMID- 10713751 TI - Carotenoids and retinol-equivalents in food composition tables from European countries (EPIC study) PMID- 10713752 TI - Response to the letter submitted by B olmedilla and F granado entitled 'Carotenoids and retinol-equivalents in food composition tables from european countries (EPIC study)' PMID- 10713753 TI - Insulin resistance: the metabolic syndrome X PMID- 10713754 TI - Lipoproteins in health and disease PMID- 10713797 TI - A European perspective on social anxiety disorder. AB - Epidemiologic surveys conducted across Europe indicate that the lifetime prevalence of social anxiety disorder in the general population is close to 7%. The disorder in adulthood rarely presents in its 'pure' form and 70-80% of patients have at least one other psychiatric disorder, most commonly depression. Social anxiety disorder is a risk factor for the development of depression and alcohol/substance use or dependence, especially in cases with an early onset (< 15 years). Individuals with social anxiety disorder have significant functional impairment, notably in the areas of initiation and maintenance of social/romantic relationships and educational and work achievement. The economic consequences of social anxiety disorder are considerable, with a high level of diminished work productivity, unemployment and an increased utilisation of medical services amongst sufferers. Effective treatment of social anxiety disorder would improve its course and its health and economic consequences. PMID- 10713799 TI - Social phobia in the community: relationship between diagnostic threshold and prevalence. AB - This paper investigates the prevalence of symptoms and various diagnostic criteria of DSM-IV social phobia in a French national representative population of 12,873 subjects, aged 15 or more. Respondents filled out a mailed questionnaire based on the social phobia section of the Munich-Composite International Diagnostic Interview (M-CIDI) in the year 1996. Response rate was 80.5%. Sixty-seven point one percent of the sample acknowledge having at least once in their lifetime a strong fear of one or more of the six prototypical social fear situations that are used as the CIDI social phobia stem items. However, only a few fulfilled all DSM-IV diagnostic criteria for social phobia. Depending on the type of diagnostic algorithms used and the stringency in which these criteria are applied, the resulting prevalence varied between 1.9 and 7.3%. These findings provide some further evidence about the considerable effects of varying diagnostic criteria and thresholds on prevalence rates for social phobia, explaining why most recent surveys have reported considerably higher rates of social phobia than those in the early 1980s. PMID- 10713798 TI - Epidemiology of social phobia: a clinical approach. AB - The recent epidemiologic studies report extremely varied rates for social phobia (SP). One of the reasons for this may be the difficulty in diagnosing SP, the boundaries of which are uncertain. A community survey was carried out using doctors with experience in clinical psychiatry as interviewers, and a clinical diagnostic instrument. Two thousand three hundred and fifty-five people (out of the 2,500 randomly selected from the population) living in Sesto Fiorentino, a suburb of Florence, Italy, were interviewed by their own general practitioner, using the MINI plus six additional questions. Six hundred and ten of the 623 subjects that were found positive for any form of psychopathology at the screening interview, and 57 negative subjects, were re-interviewed by residents in psychiatry using the Florence Psychiatric Interview (FPI). The FPI is a validated composite instrument that has the format of a structured clinical research record. It was found that 6.58% of subjects showed social anxiety not attributable to other psychiatric or medical conditions during their life. Social or occupational impairments meeting DSM-IV diagnostic requirements for SP was detected in 76 subjects (lifetime prevalence = 3.27%). Correction for age raises the lifetime expected prevalence to 4%. Sex ratio was approximately (F:M) 2:1. The most common fear was speaking in public (89.4%), followed by entering a room occupied by others (63.1%) and meeting with strangers (47.3%). Eighty-six point nine percent of subjects with SP complained of more than one fear. The mean age of onset (when the subjects first fully met DSM-IV criteria for SP) was 28.8 years, but the first symptoms of SP usually occurred much earlier, with a mean age of onset at 15.5 years. Ninety-two percent of cases with SP also showed at least one other co-morbid psychiatric disorder during their life. Lifetime prevalence of avoidant personality disorder (APD) was 3.6%. Forty-two point nine percent of cases with SP also had APD, whereas 37.9% of cases with APD developed SP. PMID- 10713801 TI - Anxiety disorders in anorexia nervosa and bulimia nervosa: co-morbidity and chronology of appearance. AB - The objectives of the study were to assess lifetime prevalence of specific anxiety disorders, and their age of onset relative to that of eating disorders (ED), in a French sample of patients with anorexia nervosa (AN) or bulimia nervosa (BN). We assessed frequencies of seven anxiety disorders and childhood histories of separation anxiety disorder among 63 subjects with a current DSM-IV diagnosis of an ED, using the Composite International Diagnostic Interview (CIDI). Eighty-three percent of subjects with AN and 71% of those with BN had at least one lifetime diagnosis of an anxiety disorder. By far, the most frequent was social phobia (55% of the anorexics and 59% of the bulimics). When present, the co-morbid anxiety disorder had predated the onset of the ED in 75% of subjects with AN, and 88% of subjects with BN. Our results are consistent with those of studies conducted in other countries, and show that an anxiety disorder frequently exists before an ED. This has to be taken in consideration for successful treatment of patients with AN or BN. PMID- 10713800 TI - Trends in the prevalence of social phobia in the United States: a synthetic cohort analysis of changes over four decades. AB - Previous analysis of data from the U.S. National Comorbidity Survey (NCS) [24] suggested that the lifetime prevalence of social phobia in the community has increased significantly in recent cohorts. Furthermore, a latent class analysis of NCS data [21] revealed two primary classes of persons with social phobia: those with exclusive speaking fears and those with one or more other social evaluative fears. Social phobia in the other social fear group is more persistent, more impairing, and more highly co-morbid with other DSM-III-R disorders. The current report presents data on whether the cohort effect is a general aspect of social phobia or is specific to one of the NCS social phobia subtypes, and whether the cohort effect varies as a function of socio-demographic characteristics. Data were drawn from the NCS. Social phobia was assessed with a revised version of the Composite International Diagnostic Interview. Retrospective age of onset reports were used to estimate Kaplan-Meier survival curves for first onset of social phobia in each cohort represented in the survey. Comparison of these curves allowed us to make synthetic estimates based on retrospective reports of intercohort trends in lifetime prevalence. The lifetime prevalence of social phobia appears to have increased in recent cohorts. However, this increase does not exist among social phobics with exclusive fears of speaking. The increase is most pronounced among white, educated, and married persons, and it is not explained by increased co-morbidity with other mental disorders. The fact that the cohort effect is more pronounced for social phobia with one or more non-speaking fears is important in that this is generally a more severe form of the disorder with an earlier age of onset than social phobia with pure speaking fears. The fact that the cohort effect is most pronounced among people with social and economic advantage (i.e., white, married, well-educated) is intriguing and raises questions about the etiologic process that warrant further study in future research. PMID- 10713802 TI - Disability and quality of life in pure and comorbid social phobia. Findings from a controlled study. AB - Social phobia is increasingly recognized as a prevalent and socially impairing mental disorder. However, little data is available regarding the general and disease-specific impairments and disabilities associated with social phobia. Furthermore, most studies have not controlled for the confounding effects of comorbid conditions. This study investigates: (a) the generic quality of life; (b) work productivity; and, (c) various other disorder-specific social impairments in current cases with pure (n = 65), comorbid (n = 51) and subthreshold (n = 34) DSM-IV social phobia as compared to controls with no social phobia (subjects with a history of herpes infections). Social phobia cases reported a mean illness duration of 22.9 years with onset in childhood or adolescence. Current quality of life, as assessed by the SF-36, was significantly reduced in all social phobia groups, particularly in the scales measuring vitality, general health, mental health, role limitations due to emotional health, and social functioning. Comorbid cases revealed more severe reductions than pure and subthreshold social phobics. Findings from the Liebowitz self-rated disability scale indicated that: (a) social phobia affects most areas of life, but in particular education, career, and romantic relationship; (b) the presence of past and current comorbid conditions increases the frequency and severity of disease-specific impairments; and, (c) subthreshold social phobia revealed slightly lower overall impairments than comorbid social phobics. Past-week work productivity of social phobics was significantly diminished as indicated by: (a) a three-fold higher rate of unemployed cases; (b) elevated rates of work hours missed due to social phobia problems; and (c) a reduced work performance. Overall, these findings underline that social phobia in our sample of adults, whether comorbid, subthreshold, or pure was a persisting and impairing condition, resulting in considerable subjective suffering and negative impact on work performance and social relationships. The current disabilities and impairments were usually less pronounced than in the past, presumably due to adaptive behaviors in life style of the respondents. Data also confirmed that social phobia is poorly recognized and rarely treated by the mental health system. PMID- 10713803 TI - Cognitive aspects of social phobia: a review of theories and experimental research. AB - Cognitive theories of social phobia have largely been inspired by the information processing models of anxiety. They propose that cognitive biases can, at least partially, explain the etiology and maintenance of this disorder. A specific bias, conceived as a tendency to preferentially process socially-threatening information, has been proposed. This bias is thought to intervene in cognitive processes such as attention, memory and interpretation. Research paradigms adopted from experimental cognitive psychology and social psychology have been used to investigate these hypotheses. The existence of a bias in the allocation of attentional resources and the interpretation of information seems to be confirmed. A memory bias in terms of better retrieval for threat-relevant information appears to depend on specific encoding activities. PMID- 10713804 TI - Are social fears and DSM-IV social anxiety disorder associated with smoking and nicotine dependence in adolescents and young adults? AB - To investigate associations between social anxiety and smoking behaviour in order to explore whether social anxiety predicts the first onset of cigarette smoking, regular smoking and the development of nicotine dependence. Baseline and four year follow-up data from the Early Developmental Stages of Psychopathology Study (EDSP), a prospective-longitudinal community study of 3,021 adolescents and young adults, are used. Smoking behaviour and psychopathology were assessed with the M CIDI and its DSM-IV algorithms. At baseline, 35.7% of the sample were regular smokers, and 18.7% fulfilled criteria for DSM-IV nicotine dependence. Twenty seven point two percent reported at least one social fear, and 7.2% met criteria for DSM-IV social phobia, most of whom reported first onset of social fear problems clearly prior to smoking initiation. Cross-sectional retrospective baseline analyses based on retrospective reports revealed that social fears and DSM-IV social phobia were both significantly associated with higher rates of nicotine dependence. Prospective-longitudinal analyses that were conducted in an attempt to confirm cross-sectional retrospective results showed that baseline non users with social fears (OR = 3.85) and baseline non-dependent users with social fears (OR = 1.5) had an increased risk of onset of nicotine dependence during the follow-up period of four years. These findings remained significant even when controlling for co-morbid depressive disorders. Social anxiety was found to be significantly associated with nicotine dependence in both cross-sectional retrospective and prospective-longitudinal analyses. It is suggested that social fears could lead to heavy tobacco use as smoking is a socially acceptable behaviour that relieves anxiety in social situations. Possible differential effects of social anxiety on the early stages of smoking behaviour compared to effects on nicotine dependence are discussed. These findings should stimulate a continued search into potentially causal links between social fear symptoms and the development of tobacco consumption and nicotine dependence in adolescence. PMID- 10713805 TI - The challenge of multidrug resistant typhoid in childhood: current status and prospects for the future. PMID- 10713806 TI - Randomized controlled trial of once vs. twice daily gentamicin therapy in newborn. AB - OBJECTIVE: To compare the efficacy of once daily gentamicin administration to the conventional twice daily dosage schedule by estimation of serum gentamicin concentrations (SGC) in neonates. DESIGN: Randomized controlled trial. SETTING: Medical college hospital. SUBJECTS: Seventy three neonates of gestational age>32 weeks at risk or with clinical features of sepsis. METHODS: The subjects were divided into preterm and term groups. Babies in each of these groups were randomized to receive a single daily dose (4 mg/kg) or a twice daily dose (2.5 mg/kg) of injection gentamicin intravenously. Trough and peak SGC were estimated half an hour prior and one hour after the second dose. Statistical analysis was done using the equivalence method. RESULTS: In preterm as well as term babies, the mean peak and trough gentamicin levels were comparable in the two regimens. There is statistically significant evidence to show that the effect of once daily and twice daily dosage is similar. CONCLUSION: Once daily gentamicin administration is as effective as twice daily therapy and would be more cost effective. PMID- 10713807 TI - Follow-up study of survivors of severe protein energy manlnutrition. AB - OBJECTIVE: To evaluate the nutritional profile of survivors of severe protein energy malnutrition on follow-up at 1-3 years and 5-7 years. DESIGN: Prospective and Cross-sectional point prevalence. METHODS: Group I comprised 50 severely malnourished children dischared 5-7 years from nutritional rehabilitation center and Group II comprised 50 children discharged 1-3 years ago. The nutritional status of these 100 children was compared to the nutritional status of 35 siblings who had not suffered from severe malnutrition in their earlier life (Group III). RESULTS: All the malnourished children showed significant improvement in weight for age. More children in Group I had better height for age compared to Group II(p>0.05). Analysis of weight for height showed that study children has better nutritional status than their siblings who had not suffered from significant malnutrition. CONCLUSION: Significant improvement in nutritional status occurs with nutritional rehabilitation. However, improvement in height for age is more difficult to obtain. Interestingly, rehabilitated malnourished children on follow up had better nutritional status as compared to their siblings. PMID- 10713808 TI - Undernutrition and adolescent growth among rural Indian boys. AB - OBJECTIVE: To evaluate impact of undernutrition on various adolescent growth parameters among rural Indian boys. DESIGN: Adolescent boys covering 8-18 yr age group were observed longitudinally for a period of 3 years. SETTING: Adolescent boys (n=673) from seven different villages within 30 to 40 km from Pune were studied. METHOD: Six monthly measurements on weight (upto 50 g) and height (upto 0.1 cm) were recorded and age assessment was done from school records with reasonable accuracy. RESULTS: Stunted and underweight boys were lighter (by 4 kg) and shorter (by 8 cm) at 10 yr age compared to their normal counterparts but this difference increased to 12 kg and 10 cm respectively by adulthood. Undernourished boys however, revealed significant height gains at later ages especially beyond 14+ yr, compared to normals suggesting slow, gradual but continual growth. Undernutrition delayed age at take-off and age at PHV by about 2 yr, and lowered attained height at PHV (by 5 cm) and adult height (by 7 cm). CONCLUSION: Normal and maluourished children from the same rural community show wide differences in their adolescent growth performance. Nutritional deprivation thus seems to affect almost all growth parameters and final adult size too. PMID- 10713809 TI - Consensus guidelines on management of childhood asthma in India. PMID- 10713810 TI - Neonatal morbidity and mortality: report of the national neonatal and perinatal database. PMID- 10713811 TI - Predicting neuro-developmental outcome at 3 months of age in babies with hypoxic ischemic encephalopathy by Vojta's neurokinesiological examination. PMID- 10713812 TI - Clinical and computerised tomography evaluation of term neonates with perinatal asphyxia. PMID- 10713813 TI - Assessment of iodine deficiency in Ernakulam district, Kerala state. PMID- 10713814 TI - Prevalence of Cryptosporidium associated diarrhea in a community. PMID- 10713815 TI - Esophageal stricture. PMID- 10713816 TI - Primary thyrotoxicosis (Graves disease). PMID- 10713817 TI - Hereditary angioneurotic edema. PMID- 10713818 TI - Amebic abscess of both liver lobes: simultaneous rupture into pleura and stomach. PMID- 10713819 TI - Idiopathic spontaneous rupture of bile duct. PMID- 10713820 TI - Synovial sarcoma of the hand. PMID- 10713821 TI - Role of DPT or TT vaccines after injury or as prophylaxis. PMID- 10713822 TI - Role of DPT or TT vaccines after injury or as prophylaxis - reply PMID- 10713823 TI - Need of rabies vaccine after a course of rabies vaccination. PMID- 10713824 TI - Need of rabies vaccine after a course of rabies vaccination - reply PMID- 10713825 TI - Safety of Ghasa. PMID- 10713826 TI - The updates are for the audience. PMID- 10713828 TI - Current status of iodine deficiency disorders control program -reply PMID- 10713827 TI - Current status of iodine deficiency disorders control program. PMID- 10713829 TI - Current status of iodine deficiency disorders control program. PMID- 10713830 TI - Current status of iodine deficiency disorders control program - reply PMID- 10713832 TI - Hypocalcemic effect of phototherapy - reply PMID- 10713831 TI - Hypocalcemic effect of phototherapy. PMID- 10713833 TI - Foreign bodies in respiratory passages. PMID- 10713834 TI - Foreign bodies in respiratory passages - reply PMID- 10713836 TI - Inaugural function of V international congress of tropical pediatrics february 9, 1999, jaipur PMID- 10713835 TI - Diagnosis and management of Kala azar. PMID- 10713837 TI - Presidental address XXXVI National Conference of IAP, Jaipur, February 13, 1999. PMID- 10713838 TI - Widespread outbreaks of measles in rural Uttar Pradesh, India, 1996: high risk areas and groups. AB - OBJECTIVE: To describe outbreaks of measles which affected many districts in Uttar Pradesh (UP) during 1996. DESIGN: Outbreak investigations. SETTING: The state of Uttar Pradesh, India. METHODS: The reported data on measles morbidity, mortality and vaccine coverage from 1991 through 1996 were reviewed. Reported vaccine coverage levels were compared with the results of coverage surveys carried out in UP from 1992 through 1996. Line lists on measles cases were analyzed to ascertain the age, immunization status, geographical distribution, and age and sex-specific fatality ratios during the outbreaks. A community survey was organized in 7 affected villages to estimate vaccine effectiveness. RESULTS: Fifty one of 68 districts in UP reported 6922 measles cases and 281 deaths in 1996. The majority of cases and deaths occurred in June and July which are usually low transmission months. Overall cases fatality ratio (CFR) was 4.1%. CFRs were significantly higher in females and young children. The median age of cases was found to be below 5 years. There was heavy clustering of cases and deaths in rural areas. About 85% of the cases and virtually all the measles associated deaths occurred in unvaccinated children. Published documents on statewide coverage surveys revealed that the measles vaccine coverage levels ranged between 26% and 36% during 1992-96. Large gaps were found between reported coverage and survey results. Nevertheless, epidemiological studies indicated a vaccine effectiveness of more than 90%. CONCLUSIONS: The outbreaks occurred due to poor vaccine coverage levels and an inefficient surveillance system which failed to generate early warning signals. The study highlights the urgent need to raise the vaccine coverage levels rapidly in all districts to achieve measles control and prevent future outbreaks in UP. PMID- 10713839 TI - Longitudinal growth of arm circumference in Punjabi infants. AB - OBJECTIVE: To study distance and velocity growth pattern of mid-upper-arm circumference in Punjabi infants. DESIGN: Longitudinal, monthly follow up. SETTING: Growth clinic and homes of subjects. SAMPLE: One hundred and fifty four (Male 86, Female 68) Punjabi infants weighing more than 2.5 kg at birth with gestation over 37 weeks. METHODS: Every subject was measured for mid-upper-arm circumference between 1 to 12 months of age at one monthly intervals by the same investigator with a time tolerance of 3 days on the day of measurement. RESULTS: Mid upper arm circumference showed rapid increase between 1 to 6 months whereafter, gain became slower during second half of infancy. It had grown by 34.9% in male and 40.2% in female infants between 1 to 12 months. Sex differences favoring male infants were statistically significant between 1 to 4 months. Monthly growth rates for arm-circumference depicted vascillatory pattern. CONCLUSION: The values presented for arm circumference may be used as reference base to monitor growth of children during infancy. PMID- 10713840 TI - Management of shock. PMID- 10713841 TI - Takayasu's arteritis in young children: a potentially treatable condition. PMID- 10713842 TI - Growth and morbidity patterns of exclusively breast-fed preterm babies. PMID- 10713844 TI - Blood lead levels in urban and rural Indian children. PMID- 10713843 TI - Outbreak of Salmonella worthington meningitis in neonatal intensive care unit. PMID- 10713845 TI - Adenosine infusion in the management of a micropremi neonate with pulmonary hypertension. PMID- 10713846 TI - Pancreatopleural fistula presenting as hemorrhagic pleural effusion. PMID- 10713847 TI - Osteoporosis pseudoglioma syndrome. PMID- 10713848 TI - Measles vaccination and risk of SSPE. PMID- 10713849 TI - Measles vaccination and risk of SSPE - reply PMID- 10713851 TI - Contraindications of OPV - reply PMID- 10713850 TI - Contraindications of OPV. PMID- 10713852 TI - Single cell translocations: is it coincidental or causal? PMID- 10713853 TI - Prevalence of iron deficiency anemia amongst pregnant women in urban slum communities of Delhi. PMID- 10713854 TI - Acute stroke due to left atrial myxoma. PMID- 10713856 TI - Nimesulide toxicity - reply PMID- 10713855 TI - Nimesulide toxicity. PMID- 10713857 TI - Management of breath holding spells. PMID- 10713858 TI - Management of breath holding spells - reply PMID- 10713859 TI - Epigastric heteropagus. PMID- 10713860 TI - Neu-Laxova syndrome. PMID- 10713861 TI - Central pattern generators PMID- 10713862 TI - Multicenter randomized trial comparing meropenem (1.5 g daily) and imipenem/cilastatin (2 g daily) in the hospital treatment of community-acquired pneumonia. AB - An open, multicenter study with 144 patients, aged between 18 and 94 years, was performed to compare the efficacy and safety of meropenem with imipenem/cilastatin in the hospital treatment of community-acquired pneumonia. Patients were randomized to receive either intravenous meropenem (500 mg every 8 h) or intravenous imipenem/cilastatin (1,000 mg every 12 h). The primary end point was considered to be clinical efficacy and the secondary end points were bacteriological response and safety assessment. At the end of therapy, cure or improvement in signs and symptoms as a satisfactory clinical response was observed in 57 of 64 (89.1%) meropenem-treated patients and in 60 of 66 (90.9%) imipenem/cilastatin patients. The mean duration of treatment was 10 days for meropenem and 9.7 days for imipenem/cilastatin. In patients who were followed up for weeks 2-4, the response was satisfactory (100%) for both treatments. A satisfactory bacteriological response, defined as either presumed or confirmed eradication of all pathogens, was found in eight patients who had received meropenem and in 14 patients who had received imipenem/cilastatin. Response was considered satisfactory in 100% of the meropenem group and in 92.9% of the imipenem/cilastatin group and at follow-up, it was 100% for both treatments. Drug related adverse events were reported in three (4.2%) meropenem-treated patients and in eight (11.0%) imipenem/cilastatin-treated patients. None of these events was classified as serious. The results of this study show that the clinical and bacteriological efficacy and tolerability of meropenem (500 mg every 8 h) are similar to that of imipenem/cilastatin (1,000 mg every 12 h) in the hospital treatment of community-acquired pneumonia. PMID- 10713863 TI - Mucosal wound healing after nasal surgery. A controlled clinical trial on the efficacy of hyaluronic acid containing cream. AB - In the present clinical trial the efficacy of a new nasal cream containing hyaluronic acid (Rhinogen) on mucosal wound healing has been evaluated in comparison to an ointment (H.E.C.), which is commonly prescribed for this disorder in Switzerland. A total of 56 patients recovering from surgical operation of the nasal cavities participated in this study. In both treatment groups (Rhinogen n = 27 patients, H.E.C. n = 29 patients) respiration and the condition of the nasal mucosa clearly improved. The statistical comparison between the two treatments showed a significant difference in favor of Rhinogen. With regard to the improvement in respiration, the Rhinogen-treated group showed a faster and greater progress than did the H.E.C.-treated group. Furthermore, hyaluronic acid prevented extensive crust formation during the first week of wound healing. The analysis of the efficacy of the treatments, judged by both the patients and the investigator, showed the overall superiority of Rhinogen (patients: p = 0.0041, investigator: p = 0.0023) after 6 weeks of treatment. Furthermore, Rhinogen scored significantly better than H.E.C. with respect to the organoleptic parameters of smell and sensation of cooling. Both treatments were well tolerated. No adverse reactions were reported or observed for Rhinogen, whereas three patients in the H.E.C.-treated group complained of sore throat and burning sensation when the ointment flowed down into the pharynx. In conclusion, this study confirms the therapeutic benefit of hyaluronic acid in mucosal wound healing. PMID- 10713864 TI - Antiinflammatory effects of seaprose-S on various inflammation models. AB - The antiinflammatory activity of seaprose-S in different experimental models involving different biochemical mediators of inflammation was investigated. In vivo experiments were performed using male Sprague-Dawley rats and in vitro experiments were performed using articular cartilage explants of pig joints. In acute experimental models of inflammation, 0.5, 1 or 2 mg/kg of seaprose-S was injected intravenously (i.v.) before challenge with inflammatory agents. In adjuvant-induced arthritis, seaprose-S was given as a 2 mg/kg i.v. dose once a day for 4 consecutive days from day 8 after injection of the adjuvant. In cartilage-synovium cocultures, seaprose-S was incubated at a concentration of 0.001 microM and 0.05 microM. Paw volume was measured with a plethysmograph and proteoglycan synthesis was determined in articular cartilage-synovium coculture by incorporation of 35S-sulfate. Seaprose-S inhibited inflammation dose dependently in carrageenan, concanavalin-A, FeCl2, nystatin-induced paw edema and in carrageenan-induced pleurisy and acetic acid-induced peritonitis. In Freund's adjuvant-induced arthritis, seaprose-S significantly reduced the primary and secondary lesions. In vitro on articular cartilage, seaprose-S increased proteoglycan synthesis in the cartilage alone and reduced the inhibition of proteoglycan synthesis in the cartilage cocultured with minced synovium. PMID- 10713866 TI - Oxidative stress and antioxidants at skin biosurface: a novel antioxidant from lemon oil capable of inhibiting oxidative damage to the skin. AB - Atmospheric pollutants are an important source of oxidative and nitrosative stress both to terrestrial plants and to animals. Skin, which has a highly differentiated and certainly complex organizational structure, is particularly vulnerable to free radical damage because of its contact with oxygen and with other environmental stimuli. Fruit and vegetables contain several classes of compounds that when ingested can potentially contribute to antioxidant defenses. In the present study we employed a novel gas chromatographic method to assess the antioxidant properties of a natural compound isolated from lemon oil, which we have called Lem1. We provide experimental evidence that Lem1 is endowed with a strong antioxidant activity and that it is capable of inhibiting free radical mediated reactions, as evaluated in vitro and in vivo. The present study extends our previous findings and demonstrates that topical application of Lem1 in healthy volunteers significantly increases the antioxidative potential of skin biosurface, thus highlighting the effectiveness of a natural antioxidant biotechnology in the antiaging management of skin. PMID- 10713865 TI - Vasodilatory effect of diuretics is dependent on inhibition of vascular smooth muscle carbonic anhydrase by a direct mechanism of action. AB - Five years ago, our in vitro and in vivo studies demonstrated for the first time that diuretic agents such as furosemide, hydrochlorothiazide, amiloride, triamterene and spironolactone inhibit carbonic anhydrase (CA) I, II and renal CA IV by a direct mechanism of action. In this paper we investigate the relationship between diuretics and CA I in the vasodilatory mechanism. Both in vitro (on purified CA I, erythrocyte CA I and smooth muscle CA I) and in vivo (in human and rabbits) we studied the effect of acetazolamide, hydrochlorothiazide, indapamide, furosemide, amiloride and triamterene on purified CA I, on human erythrocyte CA I, as well as on CA I isolated from vascular smooth muscle. Our results demonstrate that in vitro all diuretics inhibit CA I by a direct mechanism of action. Inhibition reached 100% with acetazolamide, 45% with hydrochlorothiazide, 82% with indapamide, 85% with furosemide, 68% with amiloride and 58% with triamterene. In vivo, similar inhibition of erythrocyte and smooth muscle CA I was obtained, being parallel with a reduction in arterial blood pressure values. Our data show that in addition to their already known mechanisms, diuretics also inhibit CA in vascular smooth muscle. Our results suggest that this mechanism is achieved by means of pH changes induced by CA I inhibition. PMID- 10713868 TI - Usefulness of routine pre-operative chest radiography for anaesthetic management: a prospective multicentre pilot study. AB - A prospective multicentre pilot study was undertaken in 20 Italian hospitals to assess the influence of a routine pre-operative chest radiograph on anaesthetic management and to characterise which patients might benefit from it. A total of 6111 patients undergoing elective surgery and submitted for routine pre-operative chest radiograph were enrolled. Abnormal preoperative chest radiographs were reported in 1116 patients (18.3%). Pre-operative chest radiograph altered the anaesthetic management (i.e. useful pre-operative chest radiograph) in 313 patients (5.1%). Male sex, age > 60 years, ASA classes > or = 3, respiratory diseases, and the presence of two or more co-existing diseases were significantly related to the probability of a useful pre-operative chest radiograph using multivariate analysis (P < 0.01). The classification of the surgical intervention and, of the co-existing diseases, the presence of cardiac disease had a very low influence when determining the probability that a pre-operative chest radiograph would be useful. A simple equation includes the effects of all the variables studied and allows calculation of the probability of a useful pre-operative chest radiograph. This study indicates that in healthy, female, < or = 60-year-old patients, submitted for standard surgery, the probability of a useful pre operative chest radiograph ranges from 0.2% to 3.5% according to the hospital. The probability increases in male or elderly subjects, or in the presence of co existing respiratory diseases, or in ASA classes > or = 3, but there is a wide variation between hospitals. PMID- 10713869 TI - Comparison of anti-emetic effects of ondansetron, metoclopromide or a combination of both in children undergoing surgery for strabismus. AB - This prospective, randomized and double-blinded study was designed to evaluate the anti-emetic efficacy of a combination of ondansetron and metoclopramide in 100 ASA physical status I and II children of either sex and 1-15 years of age undergoing elective surgery for strabismus. A standardized anaesthetic technique and post-operative analgesia were used for all the children. Children were divided into four groups. They received saline, metoclopromide 250 micrograms kg 1, ondansetron 150 micrograms kg-1 or a combination of metoclopramide 150 micrograms kg-1 and ondansetron 100 micrograms kg-1 intravenously immediately after the insertion of an intravenous cannulae. There were no differences between the groups in their age, gender, weight, duration of surgery, number of muscles subjected to surgery or intravenous fluids received. In the first 24 post operative hours, 18 (72%) patients in the placebo group, 15 (60%) patients in the metoclopramide group, 10 (40%) patients in the ondansetron group and 11 (44%) patients in the combination group had nausea or vomiting. The overall incidence of post-operative nausea and vomiting was significantly (P < 0.05) lower in the combination group and in the ondansetron group compared with the placebo group. Nine (36%) patients in both the placebo and the metoclopramide groups and one (4%) patient in the ondansetron group required rescue anti-emetic treatment. None of the patients in the combination group required rescue anti-emetic and this was significantly less (P < 0.01) when compared with the placebo and the metoclopramide groups. Recovery and sedation scores were comparable in all the four groups. A combination of metoclopramide 150 micrograms kg-1 and ondansetron 100 micrograms kg-1 administered prior to surgery was not found to be more effective than ondansetron 150 micrograms kg-1 alone for the prophylaxis of nausea and vomiting following surgical repair of strabismus in paediatric patients. PMID- 10713867 TI - Remifentanil, propofol or both for conscious sedation during eye surgery under regional anaesthesia. AB - We performed a prospective, randomized study comparing the efficacy and safety of remifentanil, propofol or both for conscious sedation during eye surgery under retrobulbar blockade. Forty-five unpremedicated patients were assigned to receive remifentanil (group R) (n = 15, mean dosage: 0.05 +/- 0.03 microgram kg-1 min-1), propofol (group P) (n = 15, 1.5 +/- 0.5 mg kg-1 h-1) or a combination (group RP) (n = 15, R: 0.03 +/- 0.01 microgram kg-1 min-1; P: 0.7 +/- 0.2 mg kg-1 h-1). Haemodynamic responses were comparable among all groups. Minimum values for respiratory rate were lower in R patients (R: 7 vs. P and RP: 10 breaths min-1). Perioperative blood gas analysis showed differences in maximum carbon dioxide tensions (R: 51.5 vs. P: 48.3 vs. RP: 45.5 mmHg) and decrease in minimum pH values (R: -0.06 vs. P: -0.0 vs. RP: -0.01). All group P patients reported mild to intense pain during retrobulbar block, while 53% of the group R patients were free from pain. In group RP, 60% of patients experienced no pain and the remaining 40% reported mild pain only. Remifentanil, applied as the sole agent, provided superior pain relief and patient comfort when compared with propofol, but produced greater respiratory depression and postoperative nausea. The combination of remifentanil and propofol provided haemodynamic stability, adequate spontaneous respiration and pain relief, with a low risk of untoward side effects. PMID- 10713870 TI - Assessment of ondansetron and droperidol for the prevention of post-operative nausea and vomiting after cholecystectomy and minor gynaecological surgery performed by laparoscopy. AB - The anti-emetic effects of ondansetron and droperidol were evaluated in 134 ASA Grade I and II female patients, scheduled for laparoscopic cholecystectomy and minor gynaecological laparoscopic surgery, who were randomly assigned to receive ondansetron 4 mg or droperidol 75 micrograms kg-1 intravenously immediately after induction of anaesthesia. The patients were assessed 1, 6, 12 and 24 h after surgery for intensity of nausea and number of vomiting episodes. In the case of the patients undergoing laparoscopy, vomiting episodes occurred in a similar proportion in patients treated with ondansetron or droperidol, with the probability of the Type I error of 0.05 and the Type error II of 0.1. Although there was no difference between the two groups in emetic episodes following all laparoscopic procedures and gynaecological laparoscopic surgery, there was a significant difference between these parameters after laparoscopic cholecystectomy. The patients treated with ondansetron experienced a lower intensity of nausea (P = 0.04) after laparoscopic cholecystectomy, less frequent severe nausea (P = 0.02) and episodes of vomiting (P = 0.04) when compared with those in the droperidol group. We conclude, that despite the result the droperidol prophylaxis appears to be an effective alternative to ondansetron in all patients undergoing laparoscopy, the ondansetron prophylaxis is superior to droperidol in patients undergoing laparoscopic cholecystectomy. PMID- 10713871 TI - Anaesthesia for ultrasound guided oocyte retrieval: midazolam/remifentanil versus propofol/fentanyl regimens. AB - To evaluate the quality of intra-operative anaesthesia and recovery characteristics of two different anaesthesia regimens, 60 healthy women undergoing ultrasound guided oocyte retrieval for in vitro fertilization procedures were randomly allocated to receive either a propofol/fentanyl or a midazolam/remifentanil based anaesthesia. The surgical procedure was successful in all patients and no severe side effects were reported by any patient. Four patients in the midazolam/remifentanil group (13%) would not accept the same anaesthetic procedure for further in vitro fertilization treatment due to intra operative awareness, while all propofol/fentanyl patients were prepared to accept the same procedure again (P < 0.05). Patients in the propofol/fentanyl group required manual ventilation more frequently through a facemask than those patients treated with the midazolam/remifentanil combination (50% and 30%, respectively; P < 0.05). The time to achieve an Aldrete's score of 10 was shorter in the midazolam/remifentanil patients (2 +/- 2 min) than in those who received propofol/fentanyl (4 +/- 2 min) (P < 0.001), but no differences were observed in the time required to be 'fit to discharge' from the post-anaesthesia care unit. We conclude that the use of a midazolam/remifentanil regimen is as effective and safe as a fentanyl/propofol regimen in patients undergoing transvaginal oocyte retrieval for in vitro fertilization procedures. PMID- 10713872 TI - The effects of nitrous oxide and ketamine on the bispectral index and 95% spectral edge frequency during propofol-fentanyl anaesthesia. AB - In this study, we have sought to establish whether N2O and ketamine alter the bispectral index during propofol-fentanyl anaesthesia. Fourteen surgical patients were randomly assigned to one of two groups: the N2O group (n = 7) and the ketamine group (n = 7). In both groups, anaesthesia was induced with propofol 1.5 2 mg kg-1 and fentanyl 2 micrograms kg-1 and maintained with propofol 5-7 mg kg-1 hr-1 to target the bispectral index between 40 and 50. After the bispectral index value had stabilized the propofol infusion rate was fixed. In the N2O group, the following concentrations of N2O were subsequently inhaled at 20-min intervals; 20, 40, 60 and 70%, and then N2O was terminated. In the ketamine group, ketamine (0.4 mg kg-1 + 1.0 mg kg-1h-1) was given. The bispectral index and 95% spectral edge frequency were recorded 20 min after each change in concentration of N2O or ketamine infusion. The bispectral index and 95% spectral edge frequency did not change significantly in the N2O group, but increased significantly from 44.1 +/- 0.7 and 16.0 +/- 0.5 to 58.6 +/- 1.4 and 19.5 +/- 0.3 (P < 0.01), respectively, in the ketamine group. Additional N2O or ketamine did not decrease the bispectral index and 95% spectral edge frequency values. The depth of sedation should be assessed carefully using a bispectral index monitor when these anaesthetic agents are used together. PMID- 10713873 TI - A clinical comparison of ropivacaine 0.75%, ropivacaine 1% or bupivacaine 0.5% for interscalene brachial plexus anaesthesia. AB - In order to compare interscalene brachial plexus block performed with ropivacaine or bupivacaine, 45 healthy, unpremedicated patients, undergoing elective shoulder surgery, were randomly allocated to receive interscalene brachial plexus anaesthesia with 20 mL of either ropivacaine 0.75% (n = 15), ropivacaine 1% (n = 15), or bupivacaine 0.5% (n = 15). Readiness for surgery (loss of pinprick sensation from C4 to C7 and inability to elevate the limb from the bed) was achieved later with bupivacaine 0.5% (28 +/- 15 min) than with ropivacaine 1% (10 +/- 5 min) (P = 0.005) and ropivacaine 0.75% (15 +/- 8 min) (P = 0.0005). No differences in success rate were observed between the three groups; however, seven patients receiving bupivacaine 0.5% required intra-operative analgesic supplementation (fentanyl 0.1 mg intravenous) compared with one patient receiving ropivacaine 0.75%, and two patients treated with ropivacaine 1% (P = 0.02). The time from the block placement to first request for pain medication was similar in the three groups (10.7 +/- 2 h, 11 +/- 2.4 h, and 10.9 +/- 3.9 h after 0.75% and 1% ropivacaine or 0.5% bupivacaine, respectively). We conclude that interscalene brachial plexus block performed with 20 mL of either 0.75% or 1% ropivacaine allows for a prolonged post-operative pain relief, similar to that provided by bupivacaine 0.5%, with short onset time of surgical anaesthesia. PMID- 10713874 TI - Droperidol and dimenhydrinate alone or in combination for the prevention of post operative nausea and vomiting after nasal surgery in male patients. AB - Droperidol and dimenhydrinate are inexpensive antiemetic drugs. Droperidol, especially, has been studied extensively, but there are no studies on the combination of both drugs for prevention of post-operative nausea and vomiting. One hundred and forty male hospitalized patients undergoing nasal surgery were randomized to receive one of four anti-emetic regimes: placebo, dimenhydrinate (1 mg kg-1), droperidol (15 micrograms kg-1), or the combination of both drugs (droperidol 15 micrograms kg-1 + dimenhydrinate 1 mg kg-1) administered after induction of anaesthesia. Patients in the dimenhydrinate-group and the combination-group received a second dose of dimenhydrinate 6 h after the first administration to mitigate the short half-life of the drug. For general anaesthesia a standardized technique, including benzodiazepine premedication, propofol, desflurane in N2O/O2, vecuronium, and a continuous infusion of remifentanil, was used. Post-operative analgesia and anti-emetic rescue medication were standardized. Episodes of vomiting, retching, nausea, and the need for additional anti-emetics were recorded for 24 h. The main endpoint of this study was the number of patients who were completely free of post-operative nausea and vomiting (Fisher's Exact Test). Furthermore, the severity of post operative nausea and vomiting was analysed using a standardized scoring algorithm. The incidence of patients completely free of post-operative nausea and vomiting was 62.9% in the placebo-group, 77.1% in the dimenhydrinate-group (P = 0.21), and 82.9% in the droperidol-group (P = 0.07). This increased to 94.3% in the combination-group (P = 0.0015). In all three treatment groups the severity of post-operative nausea and vomiting was reduced significantly compared with placebo treatment (P = 0.0003). The incidence of side effects was similar in the four groups. Dimenhydrinate was ineffective in reducing the incidence of post operative nausea and vomiting and droperidol only reduced the severity of post operative nausea and vomiting. However, the combination of both drugs significantly reduces the incidence of post-operative nausea and vomiting when compared with placebo treatment. PMID- 10713875 TI - Profound bradycardia and hypotension following spinal anaesthesia in a patient receiving an ACE inhibitor: an important 'drug' interaction? AB - An 86-year-old man on whom a transurethral resection of prostate was performed under spinal anaesthesia developed profound bradycardia and hypotension with disturbance of consciousness during transfer to the recovery room. Initial treatment with atropine produced rapid improvement in cardiovascular and cerebral function. A further hypotensive episode (without bradycardia) occurred approximately 1 h later but responded rapidly to methoxamine. The patient made a full recovery during an overnight stay on the High Dependency Unit. Possible mechanisms for this event are discussed, with the proposal that the concomitant administration of captopril and the relative unavailability of Angiotensin II may have significantly contributed to the problem. PMID- 10713876 TI - Pressure limited ventilation with permissive hypoxia and nitric oxide in the treatment of adult respiratory distress syndrome. AB - In the management of adult respiratory distress syndrome pressure limited mechanical ventilation may protect the lungs from overdistention injury. Unacceptable hypoxia may be avoided by adding nitric oxide to the inspiratory gas, and thus make pressure limited ventilation easier to perform. There exists no consensus about an acceptable lower limit of SaO2, and in the present case we gave preference to pressure limitation at the cost of oxygenation. A young woman with severe adult respiratory distress syndrome was set on pressure limited mechanical ventilation with peak pressures of 35-38 cm H2O, PEEP of 10-12 cm H2O, and FiO2 of 0.95 with 20 ppm nitric oxide. SaO2 varied between 75 and 85%, and cardiac output ranged between 5.2 and 7.5 L min-1. Oxygen consumption was in the upper normal range, and she did not became acidotic. After 3 days, she started to improve. In conclusion, it seems that hypoxia might be well tolerated as long as the circulation is not compromised. It might prove beneficial to accept some hypoxia to avoid ventilator induced lung damage. PMID- 10713877 TI - Undiagnosed central anticholinergic syndrome may lead to dangerous complications. AB - This report describes two cases of central anticholinergic syndrome, the first after general anaesthesia and the other during a prolonged stay in the intensive care unit. The symptoms in both patients resolved soon after physostigmine administration. There was a delay in the diagnosis of central anticholinergic syndrome, which resulted in acute lung injury and unanticipated intensive care unit admission. It is suggested that in cases of abnormal mental recovery after anaesthesia or sedation, the diagnosis of central anticholinergic syndrome should be considered. PMID- 10713878 TI - Continuous spinal anaesthesia/analgesia for abdominal aortic aneurysm repair and post-operative pain management. AB - The intra-operative management of two patients with chronic obstructive pulmonary disease and cardiovascular pathology, who underwent peripheral reconstructive vascular surgery under continuous spinal anaesthesia, is described. Furthermore, continuous intrathecal analgesia was also continued in the post-operative period and provided effective pain relief that was reflected by the favourable surgical outcome. PMID- 10713879 TI - Editorial: epidural blood patch. Eur J Anaesthesiol 1999; 16: 211-213. PMID- 10713880 TI - 2000: promises and plans. PMID- 10713881 TI - Formal recognition of the speciality of medical genetics in Portugal. PMID- 10713882 TI - Clinical and molecular advances in autosomal dominant cerebellar ataxias: from genotype to phenotype and physiopathology. AB - Major advances have been made in the understanding of autosomal dominant cerebellar ataxias since genetic markers came into use in the 1980s. The subsequent mapping of nine genes, six of which have been identified, involved in this clinically diverse group of disorders highlighted their great genetic heterogeneity. Evidence is now accumulating that, except for SCA8, the same molecular and physiopathological processes underlie these diseases and other neurodegenerative disorders sharing the same mutational basis, the expansion of a (CAG)n-polyglutamine coding sequence. The clinical overlap among the different genetic entities makes prediction of the molecular origin impossible in a single patient so that molecular characterisation is necessary. However, extended clinical and neuropathological comparisons have shown that each genetic entity has a characteristic constellation of signs and symptoms that are related to CAG repeat size and disease duration. The combined genetic and clinical information form the basis of a new classification that will aid better understanding of disease evolution, assure follow up and permit genetic counselling by the clinician. PMID- 10713883 TI - High carrier frequency of the 35delG deafness mutation in European populations. Genetic Analysis Consortium of GJB2 35delG. AB - Congenital deafness accounts for about 1 in 1000 infants and approximately 80% of cases are inherited as an autosomal recessive trait. Recently, it has been demonstrated that connexin 26 (GJB2) gene is a major gene for congenital sensorineural deafness. A single mutation (named 35delG) was found in most recessive families and sporadic cases of congenital deafness, among Caucasoids, with relative frequencies ranging from 28% to 63%. We present here the analysis of the 35delG mutation in 3270 random controls from 17 European countries. We have detected a carrier frequency for 35delG of 1 in 35 in southern Europe and 1 in 79 in central and northern Europe. In addition, 35delG was detected in five out of 376 Jewish subjects of different origin, but was absent in other non European populations. The study suggests either a single origin for 35delG somewhere in Europe or in the Middle East, and the possible presence of a carrier advantage together with a founder effect. The 35delG carrier frequency of 1 in 51 in the overall European population clearly indicates that this genetic alteration is a major mutation for autosomal recessive deafness in Caucasoids. This finding should facilitate diagnosis of congenital deafness and allow early treatment of the affected subjects. PMID- 10713884 TI - Comparison of fluorescent single-strand conformation polymorphism analysis and denaturing high-performance liquid chromatography for detection of EXT1 and EXT2 mutations in hereditary multiple exostoses. AB - EXT1 and EXT2 are two genes responsible for the majority of cases of hereditary multiple exostoses (HME), a dominantly inherited bone disorder. In order to develop an efficient screening strategy for mutations in these genes, we performed two independent blind screens of EXT1 and EXT2 in 34 unrelated patients with HME, using denaturing high-performance liquid chromatography (DHPLC) and fluorescent single-strand conformation polymorphism analysis (F-SSCP). The mutation likely to cause HME was found in 29 (85%) of the 34 probands: in 22 of these (76%), the mutation was in EXT1; seven patients (24%) had EXT2 mutations. Nineteen of these disease mutations have not been previously reported. Of the 42 different amplicon variants identified in total in the cohort, 40 were detected by DHPLC and 39 by F-SSCP. This corresponds to mutation detection efficiencies of 95% and 93% respectively. We have also found that we can confidently distinguish between different sequence variants in the same fragment using F-SSCP but not DHPLC. In light of this, and the similarly high sensitivities of the two techniques, we propose to continue screening with F-SSCP. PMID- 10713885 TI - A linkage disequilibrium map of the MHC region based on the analysis of 14 loci haplotypes in 50 French families. AB - A sample of 100 individuals from 50 French families of known pedigrees were typed for 14 loci of the HLA region (DPB1, DQB1, DQA1, DRB1, DRB3, 4, 5, C4B, C4A, Bf, C2, TNFa, TNFb, B, Cw, A). Linkage disequilibrium in each pair of loci was investigated by an exact test using a Markov chain algorithm. The results indicate no disequilibrium between DPB1 and the other loci, whereas the other class II genes are all significantly linked to each other. Linkage disequilibrium is also detected between some pairs of class I and class II-class I loci despite the long physical distance separating the loci (e.g. A-B, Cw-DRB1). On the other hand, some contiguous loci of the class III region are found to be in equilibrium with each other. Several hypotheses including selection, but also unequal allelic diversity at different MHC loci are discussed to explain this complex pattern of linkage disequilibrium. PMID- 10713886 TI - APC mutation and phenotypic spectrum of Singapore familial adenomatous polyposis patients. AB - Familial adenomatous polyposis (FAP) is a familial form of colon cancer caused by mutation of the adenomatous polyposis coli (APC) gene. Although the APC gene has been extensively studied in the Caucasian population, it has not been previously described in the Chinese population. In the present study, we investigated APC mutation and phenotypic spectrum in the Singapore FAP families who are predominantly Chinese. The protein truncation test (PTT) was used to screen the entire APC gene for germline mutations in 28 unrelated families. Fifteen different mutations were identified in 22 families. Eight mutations were 1-11 basepair deletions or insertions; three involved deletions of whole exons and four were nonsense mutations. Nine of the mutations, including two complex rearrangements, are novel. Eight families including three de novo cases have the same (AAAGA) deletion at codon 1309, indicating that like the Western families, codon 1309 is also the mutation 'hot spot' for Singapore FAP families. In contrast, we did not find any mutation in codon 1061, the second hot spot for the Western population. Congenital hypertrophy of the retinal pigment epithelium (CHRPE) is consistently associated with the prescribed domain (codons 463 to 1387) and is the only phenotype with no intra-family variation. Other than CHRPE, differences in the type and frequency of extracolonic manifestations within the FAP families suggest the influence of modifying genes and environmental factors. PMID- 10713887 TI - Detection of mutations in mismatch repair genes in Portuguese families with hereditary non-polyposis colorectal cancer (HNPCC) by a multi-method approach. AB - Mutation searching was performed in the hMSH2 and hMLH1 genes in 20 Portuguese families representing 124 registered affected individuals. Of the 20, 16 fulfilled the classic 'Amsterdam' criteria for HNPCC, whereas the remaining four families satisfied a modified set of criteria. These criteria required a CRC diagnosed before age 50 years and cancers diagnosed in two other relatives within the HNPCC spectrum. A multi-method approach was performed using the protein truncation test (PTT), single strand conformation polymorphism (SSCP) with two different sets of conditions, heteroduplex analysis (HA) and denaturing gradient gel electrophoresis (DGGE). Putative phenotype-genotype correlations were also explored. Ten different germline mutations were identified. Six of these were found in hMLH1 in seven families and four in hMSH2 in four families. SSCP and DGGE had the highest diagnostic yields with the percentage of variants detected above 67% and together HA and PTT had the lowest. No single technique detected all variants. Trends for the absence of extracolonic manifestations were observed in families carrying hMLH1 germline mutations (four of seven in hMLH1 vs one of four in hMSH2). Most of the families with rectal cancer were associated with hMLH1 (six of seven in hMLH1 vs two of four in hMSH2). A multi-technique approach is necessary to identify a high percentage of germline mutations. Seven novel mutations were found in this Portuguese population. PMID- 10713888 TI - Phenotypic variation and genetic heterogeneity in Leri-Weill syndrome. AB - Leri-Weill syndrome (LWS) or dyschondrosteosis represents a short stature syndrome characterised by the mesomelic shortening of the forearms and lower legs and by bilateral Madelung deformity of the wrists. Recently, mutations in the pseudoautosomal homeobox gene SHOX have been shown to be causative for this disorder. This gene has previously been described as the short stature gene implicated in Turner syndrome (TS). We studied 32 Leri-Weill patients from 18 different German and Dutch families and present clinical, radiological and molecular data. Phenotypic inter- and intrafamilial heterogeneity is a frequent finding in LWS, and phenotypic manifestations are generally more severe in females. In males, muscular hypertrophy is a frequent finding. To test for SHOX mutations we used FISH, Southern blot and SSCP analysis as well as long-range PCR and sequencing. We identified (sub)microscopic deletions encompassing the SHOX gene region in 10 out of 18 families investigated. Deletion sizes varied between 100 kb and 9 Mb and did not correlate with the severity of the phenotype. We did not detect SHOX mutations in almost half (41%) the LWS families studied, which suggests different genetic etiologies. PMID- 10713889 TI - Opposite deletions/duplications of the X chromosome: two novel reciprocal rearrangements. AB - Paralogous sequences on the same chromosome allow refolding of the chromosome into itself and homologous recombination. Recombinant chromosomes have microscopic or submicroscopic rearrangements according to the distance between repeats. Examples are the submicroscopic inversions of factor VIII, of the IDS gene and of the FLN1/emerin region, all resulting from misalignment of inverted repeats, and double recombination. Most of these inversions are of paternal origin possibly because the X chromosome at male meiosis is free to refold into itself for most of its length. We report on two de novo rearrangements of the X chromosome found in four hypogonadic females. Two of them had an X chromosome deleted for most of Xp and duplicated for a portion of Xq and two had the opposite rearrangement (class I and class II rearrangements, respectively). The breakpoints were defined at the level of contiguous YACs. The same Xp 11.23 breakpoint was found in the four cases. That of the long arm coincided in three cases (Xq21.3) and was more proximal in case 4 (Xq21.1). Thus class I rearrangements (cases 1 and 2) are reciprocal to that of case 3, whilst that of case 4 shares only the Xp breakpoint. The abnormal X was paternal in the three cases investigated. Repeated inverted sequences located at the breakpoints of rearrangements are likely to favour the refolding of the paternal X chromosome and the recombination of the repeats. The repeat at the Xp11 may synapse with either that at Xq21.3 or that at Xq21.1. These rearrangements seem to originate as the Xq28 submicroscopic inversions but they are identifiable at the microscopic level and result from a single recombination event. PMID- 10713890 TI - Axenfeld-Rieger syndrome resulting from mutation of the FKHL7 gene on chromosome 6p25. AB - Mutations in the forkhead-like 7 (FKHL7) gene have been recently shown to cause juvenile glaucoma and anterior segment anomalies. We report on a three-generation family with Axenfeld-Rieger syndrome (ARS), harboring an alteration in the FKHL7 gene. Genetic linkage analyses excluded the ARS phenotype from chromosomes 4q25 and 13q14, the locations of the PITX2 and RIEG2 loci, respectively. Evidence of linkage was observed with markers at 6p25, near the FKHL7 gene. Direct sequencing of FKHL7 detected a C67T mutation that segregated with the ARS phenotype in this family, but was not detected in over 80 control chromosomes. This mutation is predicted to cause a nonsense mutation of the FKHL7 protein (Gln23Stop) upstream of the forkhead DNA-binding domain, and thus to generate a truncated FKHL7 protein product. This discovery broadly implicates FKHL7 in ocular, craniofacial, dental, and umbilical development. PMID- 10713891 TI - Refined mapping of the human serotonin transporter (SLC6A4) gene within 17q11 adjacent to the CPD and NF1 genes. AB - The SLC6A4 gene encodes the serotonin transporter, the target of an important class of antidepressant drugs (serotonin selective reuptake inhibitors). Polymorphisms in the SLC6A4 gene have been reported to be associated with susceptibility to depression and other psychiatric disorders. We have constructed a 1 Mb YAC and PAC contig which harbours both the SLC6A4 and the carboxypeptidase D (CPD) genes. The order of loci within the contig was cen-D17S975-D17S1549-24R D17S1294-SLC6A4-28L+ ++-(CPD, D17S2009, D17S2004)-D17S2120-ter. Both genes were deleted in one of 17 neurofibromatosis type 1 (NF1) patients carrying submicroscopic NF1 contiguous gene deletions. PMID- 10713892 TI - Clinical and neurophysiological correlations of spasticity. AB - The aim of this investigation was to explore the correlations between some neurophysiological methods and spasticity. An examination of 120 patients with spastic hemiparesis was performed. The muscle tone, force and tendon reflexes were assessed using well-known five-point scales. The F wave, T, H and flexor reflex parameters and the Hvibrated/Hmaximal ratio were obtained. Our results revealed moderate correlations (0.3 < r < 0.5) between the amplitudes of F wave, T and H reflexes and muscle tone. The correlations between the amplitude ratios (Fmaximal/M, Fmean/M, T/M, H/M) and muscle tone were poor, as were the correlations between H reflex thresholds and muscle tone. Moderate correlations existed between Hvibration/Hmaximal ratio and muscle tone. Only the second flexor reflex response showed moderate correlations with muscle tone. In conclusion correlations between the neurophysiological methods employed and muscle tone are moderate. Evaluation of all the F wave, T, H reflex and flexor reflex parameters is not necessary, as only some of these parameters show good correlations with spasticity. PMID- 10713893 TI - The tropicamide test in patients with dementia of Alzheimer type and frontotemporal dementia. AB - The tropicamide test was applied to 30 patients with probable Alzheimer's disease (pAD), 12 with frontotemporal dementia (FTD) and 46 healthy subjects. One drop of 0.01% tropicamide was instilled in one eye and one drop of saline solution in the other. The pupil diameter was measured with a Goldmann pupillometer in its basal condition and 10, 15, 20, 25, 30, 35, 45 and 55 minutes afterwards. The results do not show differences between pupil dilation observed in pAD and in FTD; in both groups, from the beginning of the test, the pupil dilated more than in healthy people. A high interindividual variability was observed. The best cutoff point is 38% of interpupillary difference at minute 25 (sensitivity = 63%, specificity = 80%). If we consider the prevalence of AD in a population over 40 years old to be 1.4%, the adjusted positive predictive value of the test would be 4.2%. According to these results, the test is not a true diagnostic marker of AD. PMID- 10713894 TI - Coexistence of cervicogenic headache and migraine without aura (?). AB - It is well known that migraine with aura may coexist with various unilateral headaches, like cluster headache and chronic paroxysmal hemicrania. It may also coexist with cervicogenic headache. The diagnosis of migraine without aura ("common migraine") poses greater problems than the diagnosis of migraine with aura. Cervicogenic headache diagnosis also poses problems when these two headaches coexist, since they have symptoms in common. Therefore, the scientific demonstration of coexistence of migraine without aura and cervicogenic headache is bound to be a difficult task. In the present study, migraine without aura and cervicogenic headache seemed to coexist in 4 patients (3 F and 1 M, mean age 50). Attacks with migraine characteristics fulfilled the IHS and IASP migraine criteria. Out of a maximum of 13 migraine characteristics based on the IHS/IASP migraine criteria, such as unilaterality, aggravation on minor physical activity, etc., none of the patients presented less than 11, as opposed to a mean of < or = 4 of these criteria in the cervicogenic type attacks. A similar system, based on criteria such as: reduction of range of motion in the neck, mechanical precipitation of attacks, etc., was also developed for cervicogenic headache. The mean number of cervicogenic headache criteria was 4.3 (out of a total of 5) in the "cervicogenic part of the picture", as opposed to 1.5 (1.8 if laterality is considered, see text) in the "migraine part of the picture". Drug regimens and anaesthetic blocks also showed different results in the two different headaches in the same patient. All in all, this study seems to support a coexistence of the two headache types. PMID- 10713895 TI - Late whiplash syndrome: a clinical and magnetic resonance imaging study. AB - Cervical hyperextension injuries are common and are associated with significant morbidity. Clinically two syndromes are described: "acute" whiplash syndrome and "late" whiplash syndrome (in which the patients are still symptomatic after six months despite normal physical and radiological examination). In order to clarify the pathology of the persistent pain in late whiplash syndrome we performed a cervical spine magnetic resonance imaging (MRI) in 33 consecutive patients suffering from this condition. Twenty-six patients (78.8%) showed MRI abnormalities, the most common MRI finding (57.6%) was pre-existent spondylosis. Indeed, the group of patients with spondylosis and other MRI changes had higher clinical scores than those without MRI abnormalities as measured by a three-point grading system based upon the symptoms and signs shown. Several MRI changes, most of them already demonstrable by standard X-ray were seen among 33 patients suffering from late whiplash syndrome. Although no one of these findings appears to be specific and certainly related to the previous neck injury, they could represent a risk factor for a longer pain duration. PMID- 10713896 TI - Ability and fitness to drive of Parkinson's disease patients. AB - The authors review reports in the literature on the fitness and ability to drive of neurosurgical patients and subjects afflicted by neurological disorders, before focusing on their own series of 204 idiopathic Parkinson's disease (PD) patients. The study sample comprised 173 men and 31 women (average age 70.6 and 74.2 years, respectively) of whom 51, for various reasons, still drove (albeit only short distances). Different variables were examined (Hoehn & Yahr scale values (of the total group and of the subgroup of active drivers), scores for various clinical diseases, and so on) looking for an association between these variables and the number of accidents incurred by PD patients as compared with the healthy population and with a control group of healthy age-matched subjects (ISTAT data). The need for adequate legislation on driving in PD emerges clearly and recommendations are given on which such legislation might be based. PMID- 10713897 TI - Echocontrast agents in neurosonology. AB - Temporal hyperostosis is the major limitation of transtemporal insonation of the basal cerebral arteries in transcranial Doppler sonography. New contrast agents capable of traversing the pulmonary bed offer new prospects for overcoming this limitation. Echocontrast agents improve the diagnostic potentiality of ultrasound techniques, increasing the diagnostic accuracy of these methods in cerebrovascular diseases, vascular malformations, venous pathologies and tumors, and may contribute to reducing the need for more invasive and expensive examinations. PMID- 10713899 TI - Family stressors as predictors of codependency. AB - Codependency has been defined as an extreme focus on relationships, caused by a stressful family background (J. L. Fischer, L. Spann, & D. W. Crawford, 1991). In this study the authors assessed the relationship of the Spann-Fischer Codependency Scale (J. L. Fischer et al., 1991) and the Potter-Efron Codependency Assessment (L. A. Potter-Efron & P. S. Potter-Efron, 1989) with self-reported chronic family stress and family background. Students (N = 257) completed 2 existing self-report codependency measures and provided family background information. Results indicated that women had higher codependency scores than men on the Spann-Fischer scale. Students with a history of chronic family stress (with an alcoholic, mentally ill, or physically ill parent) had significantly higher codependency scores on both scales. The findings suggest that other types of family stressors, not solely alcoholism, may be predictors of codependency. PMID- 10713898 TI - Prions and prion diseases. PMID- 10713900 TI - Effects of self-schema elaboration on affective and cognitive reactions to self relevant information. AB - The basic assumption of the integrative self-schema model (ISSM; L.-E. Petersen, 1994; L.-E. Petersen, D. Stahlberg, & D. Dauenheimer, 1996; D. Stahlberg, L.-E. Petersen, & D. Dauenheimer, 1994, 1999) is that self-schema elaboration (schematic vs. aschematic) affects reactions to self-relevant information. This assumption is based on the idea that schematic dimensions occupy a more central position in the cognitive system than aschematic dimensions. In the first study, this basic prediction could be clearly confirmed: The results showed that schematic dimensions possessed stronger cognitive associations with other self relevant cognitions as well as a higher resistance to change than aschematic dimensions did. In the second study, the main assumptions of the ISSM concerning the affective and cognitive reactions to self-relevant feedback were tested: The ISSM proposes that, on schematic dimensions, reactions to self-relevant feedback will most likely follow principles of self-consistency theory, whereas on aschematic dimensions positive feedback should elicit the most positive reactions that self-enhancement theory would predict. The experimental results clearly confirmed the hypotheses derived from the ISSM for affective reactions. Cognitive reactions, however, were in line with self-consistency principles and were not modified by the elaboration of the self-schema dimension involved. PMID- 10713901 TI - The use of the thematic apperception test in the study of Native American psychological characteristics: a review and archival study of Navaho men. AB - The present study was designed as a "snapshot in time"--an archival analysis of the psychological characteristics of four Navaho men taken from Navaho Veterans- A Study of Changing Values (E. Z. Vogt, 1951). From a sample of 15 men, Vogt judged 2 as "most acculturated" and 2 as "most unacculturated." The present study was an attempt to examine the Thematic Apperception Test (TAT) protocols of these men on the basis of any recurring and psychologically significant thematic patterns, shared and unshared by the "acculturated" and "unacculturated" subjects. Vogt administered cards from the original TAT set created by H. A. Murray (1943). In this article, a section concerning the definitional use of the term acculturation is provided, followed by a discussion of the limitations inherent in the analysis of TAT protocols. A general survey of psychological and cultural studies of Native Americans using the TAT or TAT modifications is also provided. The examination of the protocol sets resulted in the identification of four themes or approaches to the TAT cards that were thought to be most pervasive and significant across both levels of acculturation: economic deprivation and physical suffering, loneliness/isolation, interpersonal conflict/violence, and individualistic vs. familial orientations. These themes and approaches are illustrated with quotations from the original protocols and are later summarized as the first four categories of a table comparing the psychological characteristics of the most acculturated men with the most unacculturated men. Results of the thematic analyses are discussed within the framework of economic and social pressures traditionally experienced by Native Americans. PMID- 10713902 TI - Gender stereotyping in television advertisements: a study of French and Danish television. AB - Two similar, but not identical, content analyses of the portrayals of men and women in French and Danish television advertisements are reported. By partially replicating and extending past investigations conducted in America, Australia, Britain, Hong Kong, Indonesia, Italy, Kenya, and New Zealand, it was predicted that there would be more gender stereotyping in French television advertisements and less gender stereotyping in Danish television advertisements. In the first study, 165 French television advertisements were analyzed by following established coding categories (A. Furnham & E. Skae, 1997; L. Z. McArthur & B. G. Resko, 1975). Contrary to prediction, the results showed that traditional gender role portrayal on French television was no different from that found in other countries. Separate statistical analyses were carried out for visually versus aurally classified central figures, yet this yielded relatively few significant differences. In the second study, a sample of 151 Danish advertisements was analyzed; results showed that Danish television was generally less gender stereotypic than French television in its portrayal of women. Exactly half (5) of the coding categories showed significant differences. Finally, an international statistical comparison between these two studies and similar research in Australia, Britain, and Italy was carried out. The methodological implications of these results are discussed as well as the theoretical issues arising from other studies of this sort. PMID- 10713903 TI - Antecedents and moderators of behavioral intention: differences between U.S. and Taiwanese students. AB - The purpose of this study was to investigate the relative influence of attitude toward the act, subjective norm, and perceived behavioral control on consumers' purchase intention when consumers possess different levels of product knowledge (subjective and objective). The magnitude of the influence is compared across two different societies (U.S. and Taiwanese). U.S. (N = 295) and Taiwanese (N = 297) college students participated. The results showed that the relative importance of attitude toward the act, subjective norm, and perceived behavioral control in predicting purchase intention varied across consumers with different levels of product knowledge (subjective or objective) for the U.S. participants. However, the moderating effect of product knowledge was less profound for the Taiwanese participants. PMID- 10713904 TI - [ACE gene polymorphism and cardiovascular diseases]. AB - The angiotensin converting enzyme (ACE) is an integral part of enzymatic cascades leading to generation of angiotensin II as well as degradation of bradykinin. For this reason, it represents an important part for the metabolism of 2 vasoactive peptides. Early in this decade, convincing experimental evidence demonstrated the induction of this enzyme in several pathophysiological conditions including myocardial infarction and left ventricular hypertrophy. In parallel, a deletion/insertion (D/I) polymorphism of the human ACE gene was discovered that was related to 14 to 50% of the interindividual variance of serum ACE activity. More recently, this polymorphism was implicated in the pathogenesis of a variety of cardiovascular disorders including myocardial infarction, left ventricular hypertrophy, hypertension as well as nephropathy. PMID- 10713905 TI - [Genetic risk factors for myocardial infarct]. AB - Interactions of genetic and environmental risk factors influence the susceptibility to coronary artery disease (CAD) and myocardial infarction. In myocardial infarction occurring at young age, genetics of this multifactorial disease may be the leading factor. A number of candidate genes have been implicated in the pathogenesis of CAD and myocardial infarction. Mutations in the DNA sequence (gene polymorphisms) have been identified that appear to play a crucial role in blood pressure regulation, lipid metabolism, endothelial function, in the pathophysiology of coagulation or thrombosis, or in interventional cardiology by interfering with restenosis development. Genetic polymorphisms seem to be clinically important because they not only potentiate the individual risk under certain circumstances, but they also determine safety and effectiveness of commonly prescribed drugs. Understanding the complexity and functional relevance of genetic risk factors will be useful in early detection and treatment of individuals that are exposed to higher risk for myocardial infarction. Thus it is important to include genetic risk factors in the concept of the classical risk factor theory. Potentially in future a genetic risk profile including relevant polymorphisms may be an essential part of the clinicians' knowledge in primary and secondary prevention of coronary artery disease. PMID- 10713906 TI - [Role of the angiotensinogen gene for essential hypertension]. AB - Essential hypertension is a complex disease influenced by different genetic and environmental factors. The renin-angiotensin system (RAS) is implicated in blood pressure regulation. Angiotensinogen (AGT) is the precursor of the biologically active angiotensin II (Ang II). Initial studies on hypertensive siblings and case control studies indicated the important role of the angiotensinogen gene (AGT) for the predisposition to essential hypertension, preeclampsia and obesity related hypertension. Recently, different AGT polymorphisms had been identified and analyzed in case-control studies. The aim of present studies is the analysis of potentially functional AGT variants (C-532T, G-6A), which might be responsible for the regulation of gene expression and therefore AGT generation. The A-6 allele is in complete linkage disequilibrium with the T235 allele and is associated with higher AGT expression in vitro. Segregation linkage analysis demonstrated that the C-532T polymorphism influences plasma AGT variability more significantly than the G-6A variant. Since the C-532T polymorphism is located within a AP-2 consensus element, functional promoter analyses are required. The understanding of the molecular basis of RAS in essential hypertension may provide us with new and more specific pharmacological agents and perhaps the ability to individualize antihypertensive treatment. PMID- 10713907 TI - [Genetic polymorphism of the G-protein beta3 subunit, obesity and essential hypertension]. AB - Following a classical candidate gene approach we have detected a C825T polymorphism in the gene GNB3 which encodes the G beta 3 subunit of heterotrimeric G proteins. The 825T allele causes alternative splicing of the gene and the generation of a truncated but functionally active splice variant of G beta 3 which is referred to as G beta 3s. Thus, genotyping for the C825T polymorphism is predictive for the activation of certain G proteins in humans. The 825T allele is significantly associated with an increased risk for hypertension in Caucasians, most likely "low renin hypertension" and it accumulates significantly in individuals with a strong family history of hypertension. Highest frequencies of the 825T allele (up to 80%) are found in old ethnicities, e.g. black Africans, African Americans, bushmen, and Australian aborigines. This suggests that enhanced G protein activation represents a thrifty genotype which might have facilitated survival in our ancestors. Frequencies of the 825T allele are significant lower in Asians (approximately 40 to 50%) and Caucasians (30%). More recent studies show that young 825T allele carriers are predisposed for obesity and this association could be confirmed across different ethnicities including young Germans, as well as Chinese and black African individuals. Thus, genotyping at the GNB3 locus represents an ideal tool for preventive medicine in that individuals at risk for obesity and hypertension can be identified early and counteract their genetic predisposition through changes in lifestyle. In individuals with borderline hypertension genotyping can facilitate the decision for medical treatment as a positive test result confirms an inherited form of hypertension. PMID- 10713908 TI - Genetic risk factors and restenosis after percutaneous coronary interventions. AB - Restenosis is the major limitation of percutaneous coronary interventions. Depending on the form of intervention and patients' characteristics, 20 to 50% of the treated patients incur significant restenosis. Restenosis is caused by a complex and only partially understood cascade of events. Thrombus formation at the injury site, formation of the neointima as a result of the migration and proliferation of smooth muscle cells (SMC) and extracellular matrix production, as well as constrictive remodeling of the vessel wall contribute by a variable degree to restenosis. Restenosis is not a random event but it affects selectively a certain subset of patients. These patients have some peculiar characteristics that help to identify the presence of a higher risk for restenosis. Conventional patient-related factors account only for a relatively small portion of the predictive power, much more contribution comes from lesion and procedural characteristics. There is increasing evidence that inherited factors may explain at least part of the excessive risk for restenosis observed in certain patients. Evidence exists that gene polymorphisms may lead to quantitative or functional alterations of the respective gene products. Recent studies have also found significant associations between several polymorphic alleles encoding for proteins with a relevant role in the process of lumen renarrowing and restenosis after percutaneous coronary interventions. The best studied polymorphisms in this regard are those of the genes encoding for angiotensin-converting enzyme and platelet glycoprotein-IIIa. Completed or ongoing studies have focused on polymorphisms of genes encoding for proteins interfering with lipid metabolism, hemostasis, nitric oxide production, inflammatory mechanisms, SMC proliferation and matrix production. The results of this research will have considerable pathophysiological and therapeutical implications for the battle against restenosis. PMID- 10713910 TI - Hypotony and retinal complications after aqueous humor shunt implantation: the 1999 Dohlman Lecture. PMID- 10713911 TI - Laser applications in oculoplastic surgery and their postoperative complications. AB - Laser surgery for oculoplastic or dermatological indications--whether incisional work, removal of pigmented or vascular lesions, removal of hair, or resurfacing- necessitates that the practitioner have appropriate training in and understanding of not only the techniques but also of their advantages and disadvantages. Understanding laser safety and how to handle complications is critical to appropriate management of laser-assisted surgery. Long-term results are limited at this time, but current information regarding the use of lasers in aesthetic oculoplastic surgery appears promising. New approaches to such operations include combining more than one type of laser or combining traditional cutting blades and lasers in an effort to reduce side effects and improve outcome. PMID- 10713909 TI - Current concepts in secondary prevention after acute myocardial infarction. AB - Acute myocardial infarction (MI) is the leading cause of death around the globe. Advances in the field of cardiology have identified several effective treatments that have lead to decrease in mortality from this cause over the past 3 decades. The purpose of this article is to review the existing literature in regards to secondary prevention after acute MI. A search of MEDLINE through August of 1999 was carried out to identify any available publications on secondary prevention after MI. Evidence on the use of both pharmacological and nonpharmacological interventions that was shown to be effective in improving morbidity and mortality was sought. Recommendations for the treatment of patients with acute MI are made based on existing evidence. Betablockers, aspirin and lipid-lowering agents for patients with low density lipoprotein-cholesterol > 130 mg% should be used for all patients following a MI. Angiotensin converting enzyme inhibitors are indicated for patients with congestive heart failure and/or reduced left ventricular ejection fraction and are likely protective in most patients. Calcium channel blockers (Verapamil and Diltiazem) are indicated as second-line therapy for patients who have contraindications or are intolerant to betablockers. The routine prophylactic use of antiarrhythmic drugs to suppress ventricular ectopic beats should be avoided. Recommendations regarding diet, smoking cessation and achievement of ideal body weight should be an integral part of patient management. Referral for outpatient rehabilitation should also be strongly encouraged. Finally, adequate control of blood pressure and diabetes cannot be overemphasized. Adherence to these goals in patients with acute MI will lead to better long-term outcomes and reduction in cardiac death, recurrent MI, stroke, and need for coronary revascularization. PMID- 10713912 TI - Complications after penetrating keratoplasty. PMID- 10713913 TI - Optical disturbances and their management after myopic laser in situ keratomileusis. PMID- 10713914 TI - Chronic postoperative endophthalmitis. PMID- 10713915 TI - Postoperative sympathetic ophthalmia. AB - Although uncommon, SO is a fearful postoperative complication because of its potential to blind both eyes. It can result not only from penetrating ocular surgery but also from nonpenetrating ocular procedures. Thus, it is important to consider in any patient who has undergone ocular surgery and develops bilateral uveitis, particularly because prompt, sufficient treatment is required to maximize visual outcome. It is also important to note that the disease may present with a spectrum of clinical findings, none of which is pathognomonic. Thus, suspicion is important for making the diagnosis. Treatment should address the T-cell-mediated nature of the disease. With appropriate treatment, visual acuity of no less than 20/60 is likely. However, before the start of treatment, which consists of immunosuppressants, infection must be ruled out and potential side effects of treatments must be considered. Furthermore, any patient with a history of SO needs ample immunosuppressant coverage for ocular procedures. Better understanding of the pathogenesis of the disease may lead to safer treatments that result in improved visual outcome and a cure. Meanwhile, because of its relapsing nature, SO requires continual, close surveillance, even after many years of quiescence. PMID- 10713916 TI - Postoperative complications of periocular anesthesia. PMID- 10713917 TI - Neuroophthalmological complications of ocular surgery. AB - Visual dysfunction, including visual loss and diplopia, may occur in association with ocular surgery. Strabismus may be the most common abnormal eye movement seen as a complication of local anesthesia. Local anesthesia also may cause direct and indirect traumatic optic neuropathy. Vitrectomy is associated with visual-field loss from direct manipulation of the nerve fiber layer during suctioning of the vitreous or by direct compression of intraocular gas. Trabeculectomy may be complicated by visual-field loss. Patients may be at higher risk if their eye is hypotonous after surgery, but the duration or severity of hypotony that places the eye at risk is unknown. The only neuroophthalmic complication directly related to cataract surgery itself is AION. Complications of ONSD include motility disorders, pupillary dysfunction, and vascular compromise. Neuroophthalmic complications are uncommon after blepharoplasty. They include ocular motility disorders, transient pupil dilation, and vision loss. PMID- 10713918 TI - Intraocular lens explantation in uveitis. AB - Visual rehabilitation after cataract surgery has been improved with the development of IOLs. These lenses are well tolerated in many uveitis patients when complete control of preoperative inflammation is achieved. However, in some patients, IOL placement after cataract extraction results in chronic inflammation, deposition of inflammatory cells and debris on the IOL surface, and inflammatory membrane formation despite antiinflammatory coverage. Patients with systemic diseases characterized by chronic inflammation, such as sarcoidosis and JRA, and those with chronic ocular inflammatory conditions or inflammation involving the intermediate segment of the eye may be at high risk for these complications. In patients in whom antiinflammatory therapy fails, adequate control of inflammation may be achieved after lens explantation. PMID- 10713919 TI - Malignant glaucoma. PMID- 10713920 TI - Hypotony after glaucoma filtration surgery. PMID- 10713921 TI - Bleb complications after filtration surgery. PMID- 10713922 TI - Complications of glaucoma drainage implant surgery. AB - Glaucoma drainage implants play an important role in the management of refractory glaucoma. It may be considered as a primary procedure in eyes with severe scarring of the conjunctiva and the limbus, neovascularization, multiple previous, failed filtering procedures, and severely glaucomatous eyes for which corneal graft is planned. Recommendations have been made regarding improving the biocompatibility of the materials used in the manufacture of these implants. The current state of development of glaucoma drainage implants is likened to that of intraocular lenses in the 1970s. Further development of devices and techniques will only serve to improve our armamentarium and success against glaucoma, specifically refractory glaucoma. Glaucoma posterior tube shunts have proved to be safe, with the majority of complications either nonsight-threatening, spontaneously resolving, or reversible. Considered primarily in the most severe cases of refractory glaucoma, it has otherwise changed the outlook for such patients. PMID- 10713923 TI - Postoperative complications of pneumatic retinopexy. PMID- 10713924 TI - Postoperative complications of scleral buckling surgery. AB - The postoperative course of scleral buckling surgery can witness a host of untoward events, which may undo the anatomical success of retinal reattachment. Diligent preoperative identification of all retinal breaks and meticulous intraoperative technique will enhance the likelihood of a tranquil postoperative course. PMID- 10713925 TI - Complications of intraocular tamponade: silicone oil versus intraocular gas. PMID- 10713926 TI - Complications of surgery for subfoveal choroidal neovascularization. AB - Advancing surgical techniques have made the surgical excision of subfoveal CNV possible in all cases. However, serious surgical complications lead to a limited visual outcome in many cases. The major complications that cause poor visual outcome are related to poor case selection and include injury to the RPE, with secondary atrophy of the choriocapillaris and damage to the neurosensory retina, and a high rate of persistent or recurrent CNV. Patients with POHS have localized disease of the RPE-Bruch's membrane complex and typically have CNV growing between the RPE and neurosensory retina (type 2 CNV). These patients have the best visual prognosis postoperatively because of the potential for maintaining native RPE beneath the fovea. Patients with AMD typically have CNV growing beneath the RPE (type I CNV). These patients are poor surgical candidates because the surgical excision of type I CNV is almost always associated with debridement of native subfoveal RPE and a poor visual outcome. Presently, all surgical studies have been retrospective and are characterized by limited follow-up. Therefore, whether surgical excision of subfoveal CNV is beneficial as compared to mere observation is uncertain. A large, randomized, prospective study currently is being performed. These Submacular Surgery Trials will attempt to determine whether these surgical procedures are more efficacious than observation and whether the benefits outweight the risks of surgery in these patients. PMID- 10713927 TI - Postoperative complications of epiretinal membrane surgery. PMID- 10713928 TI - Complications of macular hole surgery. AB - Macular hole is a serious vision-threatening disease for which, until the early 1990s, no effective treatment was available. However, with advanced techniques in microsurgical vitrectomy surgery, effective and successful closure of macular holes now can be obtained. Many complications can occur after any ocular surgery. Some of the specific complications associated with macular hole surgery are RPE alterations, retinal detachments, CME, subretinal neovascular membrane, endophthalmitis, hypopyon, late re-opening of macular holes, increased intraocular pressure, visual field defects, and cataract formation. Because macular hole surgery is a relatively recent operation devised in the last 8 years, data on complications still are being compiled and studied to understand and minimize the rates of complications after such surgery. We hope that, in the future, the rate of complications after macular hole surgery will be no higher than that for any other ophthalmic procedure. PMID- 10713929 TI - [Evaluation of restenosis by serum levels of soluble Fas, Fas ligand and nuclear matrix protein before and after coronary intervention]. AB - Serum levels of soluble Fas, soluble Fas ligand, and nuclear matrix protein (NMP) were measured in 38 patients with ischemic heart disease before and after coronary angiography or coronary intervention. Serum levels of soluble Fas, soluble Fas ligand and NMP were determined by enzyme-linked immunosorbent assay. Patients one week after undergoing stent implantation had much higher levels of soluble Fas ligand and significantly lower levels of soluble Fas than patients without coronary intervention. Serum levels of soluble Fas ligand and NMP in patients with coronary restenosis were significantly higher than those in patients without restenosis. Serum levels of soluble Fas and soluble Fas ligand in patients one week after undergoing stent implantation who had coronary restenosis were markedly lower and higher than in patients without restenosis, respectively. Serum levels of soluble Fas ligand were positively correlated with NMP in those patients. These results indicate that coronary restenosis might be affected by the Fas/Fas ligand system. We conclude that measurement of soluble Fas, soluble Fas ligand and NMP is useful for determination of coronary restenosis in patients with ischemic heart disease. PMID- 10713930 TI - Efficacy of cholesterol-lowering treatment in Japanese elderly patients with coronary artery disease and normal cholesterol level using 3-hydroxy-3 methylglutaryl coenzyme A reductase inhibitor. AB - The clinical benefit of cholesterol-lowering treatment is unknown in the Japanese elderly in whom the prevalence of morbidity and mortality related to coronary artery disease are known to be low. To evaluate the efficacy of cholesterol lowering treatment with 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor in Japanese elderly patients with documented coronary artery disease, 121 patients with serum cholesterol > or = 150 mg/dl prospectively received HMG CoA reductase inhibitor, and 271 patients undergoing cholesterol-lowering treatment based on dietary therapy alone served as historical controls. The 143 elderly patients age > or = 65 years in the 2 groups had similar baseline serum total cholesterol level (201 +/- 30 vs 202 +/- 31 mg/dl), age (71 +/- 4 vs 70 +/- 4 years), proportion of men (37/53 vs 64/90), number of diseased vessels (1.7 +/- 0.9 vs 1.5 +/- 1.0), and incidences of other classical coronary risk factors, including hypertension, diabetes mellitus, smoking, obesity and family history of coronary artery disease. In all 392 patients, similar trends were observed, including serum total cholesterol level (208 +/- 33 vs 201 +/- 34 mg/dl). With HMG-CoA reductase inhibitors, serum total cholesterol level was reduced by 14% in the elderly subjects and by 13% in all patients. During the follow-up of approximately 3 years, cardiac events occurred in 5 patients (one elderly) in the treatment group and 38 patients (12 elderly) in the control group. Kaplan-Meier survival estimates revealed a higher event-free survival rate with HMG-CoA reductase inhibitors in the elderly subjects (98% vs 85%, p < 0.05) and in all patients (94% vs 86%, p < 0.05). Cox proportional hazard modeling also demonstrated a significant reduction in risk for cardiac events with drug therapy (relative risk 0.32, p < 0.05), in addition to the number of diseased vessels (relative risk 1.8, p < 0.01). In contrast, no additional risk was observed with advancing age. Cholesterol-lowering treatment with HMG-CoA reductase inhibitors is effective to improve the prognosis of Japanese elderly patients, including those with normal serum cholesterol level. PMID- 10713931 TI - Angiographically demonstrated coronary collaterals predict residual viable myocardium in patients with chronic myocardial infarction: a regional metabolic study. AB - Angiographical demonstration of coronary collateral circulation may suggest the presence of residual viable myocardium. The development of coronary collaterals was judged according to Rentrop's classification in 37 patients with old anteroseptal myocardial infarction and 13 control patients with chest pain syndrome. The subjects with myocardial infarction were divided into 2 groups: 17 patients with the main branch of the left coronary artery clearly identified by collateral blood flow from the contralateral coronary artery [Coll(+)group, male/female 10/7, mean age 56.6 years]and 20 patients with obscure coronary trunk [Coll(-)group, male/female 16/4, mean age 54.9 years]. Thallium-201 myocardial scintigraphy and examination of local myocardial metabolism were carried out by measuring the flux of lactic acid under dipyridamole infusion load. Coronary stenosis of 99% or total occlusion was found in only 5 of 20 patients (25%)in the Coll(-)group but in 16 of 17 patients(94%)in the Coll(+)group(p < 0.001). Redistribution of myocardial scintigraphy was found in 11 of 15 patients(73%)in the Coll(+)group, but only 3 of 18 patients (17%)in the Coll(-)group(p < 0.01). The myocardial lactic acid extraction rate was--13.2 +/- 17.0% in the Coll(+)group, but 9.1 +/- 13.2% in the Coll(-)group(p < 0.001). These results suggest that coronary collateral may contribute to minimizing the infarct area and to prediction of the presence of viable myocardium. PMID- 10713932 TI - [Does reperfusion therapy reduce complications in acute inferior myocardial infarction?]. AB - Both right ventricular infarction and complete atrioventricular block were frequently seen in patients with acute inferior myocardial infarction before the introduction of reperfusion therapy (RT). However, the effect of reperfusion therapy on these 2 complications is not well known. To evaluate the effect of reperfusion therapy in them, we retrospectively studied the in-hospital outcome of 103 consecutive patients with acute inferior myocardial infarction within 72 hr after the onset, 23 with right ventricular infarction and 36 with complete atrioventricular block. Patients were divided into 2 groups: RT group (n = 63) in which Thrombolysis in Myocardial Infarction (TIMI) III flow was obtained by reperfusion therapy within 24 hr after the onset, and the non-RT group (n = 40) in which TIMI III flow was not obtained or did not receive reperfusion therapy. Patients with right ventricular infarction in the RT group had a larger proportion of proximal occlusion of the right coronary artery and the absence of preinfarction angina. There were no effects of perfusion on complete atrioventricular block. In 23 patients with right ventricular infarction and 36 patients with complete atrioventricular block, in-hospital stay, duration of using temporary pacing and Swan-Ganz catheter were shorter in the RT group than the non-RT group. Reperfusion therapy does not decrease the incidence of both complications. However, successful reperfusion therapy results in a rapid improvement in hemodynamic instability and atrioventricular conduction injury, and early hospital discharge. Preinfarction angina may be associated with a protective effect against the development of these 2 complications. PMID- 10713933 TI - [Diagnostic usefulness of KL-6 measurements in patients with pulmonary complications after administration of amiodarone]. AB - Amiodarone-induced pulmonary toxicity is one of the major complications in patients receiving administration of amiodarone. KL-6 is a useful indicator to evaluate the activity of interstitial pneumonitis. We studied the clinical utility of KL-6 as a marker for amiodarone-induced pulmonary toxicity. We investigated 6 patients in whom chest radiography revealed abnormal consolidations after administration of amiodarone from 1997 to 1999. All patients were male aged 56 to 76 years (mean 66 +/- 7 years). The indications for amiodarone included sustained ventricular tachycardia in 5 patients and atrial fibrillation in one patient with refractory heart failure. The mean left ventricular ejection fraction was 31 +/- 12% (22-52%). KL-6 levels were measured by a sandwich type enzyme immunoassay using a murine monoclonal antibody (KL-6 antibody), and the cutoff level was determined at 520 U/ml. Complications occurred from 17 days to 45 months after treatment with amiodarone. The KL-6 levels were abnormally high (2,100 and 3,000 U/ml) in 2 patients with amiodarone induced pneumonitis but under the cutoff level in the non-pneumonitis patients. In one patient with amiodarone-induced pneumonitis, the KL-6 level increased from 695 to 2,100 U/ml concurrently with worsening interstitial changes shown by high resolution computed tomography. We conclude that KL-6 has practical uses as a marker for the detection and evaluation of amiodarone-induced pulmonary toxicity. PMID- 10713934 TI - [Left atrial pressure gradient and right heart failure secondary to compression of the left atrium by a huge ascending aortic aneurysm: a case report]. AB - A 71-year-old man was admitted to our department with congestive heart failure on June 28, 1998. He previously had an aortic valve replacement because of aortic regurgitation probably due to annuloaortic ectasia in 1984. Thoracic aortic aneurysm was identified during the postoperative course. Magnetic resonance imaging showed a huge saccular ascending aortic aneurysm of 12 x 11.5 x 9.5 cm size, which had severely compressed the left atrium. Doppler echocardiography documented an accelerated flow (2.2 m/sec) in the left atrium in early diastole. The calculated pressure gradient was 19 mmHg. All pressures in the right heart system were elevated. This is the first case of pulmonary hypertension and right heart failure secondary to compression of the left atrium in a patient with thoracic aortic aneurysm. PMID- 10713935 TI - [A 46-year-old woman presenting with abnormal chest X-ray shadow]. PMID- 10713936 TI - [A patient with severe left ventricular hypertrophy suffering sudden death]. PMID- 10713937 TI - Optimization of fast cardiac imaging using an echo-train readout. AB - Fast gradient-echo sequences that use an echo-train readout are becoming more widely used, particularly for imaging the heart. An important issue for these sequences involves determining the optimal duration for the echo-train readout. In normal volunteer scans and theoretically the echo-train readout duration was varied from 2.4 to 32.8 msec. Myocardial signal-to-noise ratio (SNR), myocardium tag signal difference-to-noise ratio (SDNR), flow artifact-to-noise ratio (FNR), and geometric distortion were measured and/or calculated. Our results showed that to obtain high SNR, SDNR, and data acquisition efficiency while minimizing FNR and geometric distortion, the readout duration should be 10-15 msec at 1.5 T. PMID- 10713938 TI - Three-dimensional cardiac cine magnetic resonance imaging with an ultrasmall superparamagnetic iron oxide blood pool agent (NC100150). AB - The purpose of this study was to assess image quality of three-dimensional (3D) cardiac cine magnetic resonance (MR) imaging before and after administration of a T1-shortening ultrasmall superparamagnetic iron oxide blood pool agent (NC100150). 3D cardiac cine MR imaging was performed in 13 volunteers using a radiofrequency-spoiled cardiac-gated 3D cine gradient-echo sequence with short repetition and echo times. Compared with precontrast images, postcontrast images showed no enhancement in fat and skeletal muscle, moderate enhancement in myocardium, and significant enhancement in ventricular cavity. After contrast injection, the signal ratio of the ventricular chamber to the myocardium significantly increased, and dramatic improvements were seen in the quality of the cineangiographic images and the depiction of cardiac valves. This quantitative study has shown that 3D cardiac cine MR imaging using a blood pool agent provided MR ventriculography and cineangiography with excellent image quality. PMID- 10713939 TI - Characterization of breast lesion morphology with delayed 3DSSMT: an adjunct to dynamic breast MRI. AB - The purpose of the study was to determine the sensitivity and specificity of various morphologic criteria in distinguishing malignant from benign breast lesions using a new sequence (3DSSMT) performed immediately after dynamic breast MRI. 3DSSMT combines a water-selective spectral-spatial excitation and an on resonance magnetization transfer pulse with three-dimensional spoiled gradient echo imaging. Morphologic features of 87 pathologically confirmed lesions were analyzed. The presence of either skin thickening, or a combination of a spiculated or microlobulated border, with a rim, ductal, linear, or clumped enhancement pattern was 94% specific and 54% sensitive for malignancy. Conversely, the presence of either a perfectly smooth border, a well-defined margin, non-enhancing internal septations, or a macrolobulated border was 97% specific and 35% sensitive for a benign diagnosis. In conclusion, delayed 3DSSMT discriminates a significant number of benign and malignant breast lesions; it has the potential to improve the diagnostic accuracy of dynamic breast MRI. PMID- 10713940 TI - Detection of regional pulmonary perfusion deficit of the occluded lung using arterial spin labeling in magnetic resonance imaging. AB - Detection of regional perfusion deficit in the lung has been demonstrated using an arterial spin labeling technique called flow-sensitive alternating inversion recovery with an extra radiofrequency pulse (FAIRER). A pulmonary artery was occluded using a nondetachable balloon catheter to simulate an acute pulmonary embolism in 3 of 10 rabbits. Inflating the balloon occludes the artery, and deflating the balloon allows for reperfusion. Perfusion imaging was performed pre occlusion, during occlusion, and after reperfusion. Signal enhancement due to perfusion of the pulmonary parenchyma was observed in the perfusion images with negligible artifacts. The perfusion deficit of the pulmonary parenchyma was detected distal to the site of occlusion in all three rabbits. Return of the pulmonary parenchymal perfusion was observed after reperfusion. Magnetic resonance imaging using FAIRER can detect signal loss due to absence of perfusion caused by pulmonary embolism. PMID- 10713941 TI - Abnormalities of the contrast re-circulation phase in cerebral tumors demonstrated using dynamic susceptibility contrast-enhanced imaging: a possible marker of vascular tortuosity. AB - Dynamic susceptibility contrast-enhanced magnetic resonance (MR) imaging in tumors is restricted by relaxivity effects, which may obscure any abnormality of first-pass kinetics in the re-circulation phase. The purposes of this study were a) to document the magnitude of relaxivity effects with a variety of commonly used MR susceptibility imaging techniques; and b) to determine whether the re circulation phase of the first-pass curve in tumors differs from that in normal tissue. We have confirmed that residual relaxivity effects can be eliminated from dynamic susceptibility contrast-enhanced data by several techniques. Application of these methods to enhancing vascular tumors allows detection of abnormalities in the re-circulation phase, which would otherwise be obscured. These abnormalities are independent of relative cerebral blood volume (rCBV) and presumably represent deviations from the predicted gamma variat flow pattern seen in normal tissues. We believe that the parameter rR described here provides an indicator of the chaotic nature of neovascular angiogenesis, which may be of benefit in diagnosis and management. PMID- 10713942 TI - Dynamic susceptibility contrast MR imaging: correlation of signal intensity changes with cerebral blood volume measurements. AB - Cerebral blood volume (CBV) maps derived from dynamic susceptibility contrast (DSC) magnetic resonance (MR) imaging provide valuable information regarding intracranial micro-hemodynamics and have been helpful in characterizing primary brain tumors and guiding stereotactic biopsy. Another parameter, the maximum signal drop (MSD) during the first pass of intravascular contrast bolus due to T2* effect, can also be measured directly without extensive post-processing and data manipulation. The purpose of our study is to determine whether MSD maps provide information similar to CBV maps in patients presenting with intracranial mass lesions. Twenty-nine patients with various intracranial mass lesions were studied with DSC MR imaging prior to stereotactic biopsy or volumetric resection. Maps of both CBV and MSD are calculated on a pixel-by-pixel basis and displayed as color overlays over the raw images. Relative CBV (rCBV) and MSD (rMSD) values were measured in regions of interest (ROIs) within areas of abnormality and compared. In addition, computer-generated noise was added to the data to estimate the sensitivity of each measurement to noise. The rMSD values were strongly correlated with rCBV values (r = 0.87, P = 0.0001). CBV values were much more sensitive to added noise than MSD values (P < 0.01). MSD maps derived from DSC MR imaging provide information similar to CBV maps in patients with intracranial mass lesions. MSD maps are a simple and reliable indicator of vascularity that can easily be incorporated into routine MR imaging. PMID- 10713943 TI - Evaluation of syringomyelia with three-dimensional constructive interference in a steady state (CISS) sequence. AB - The purpose of this study was to evaluate a three-dimensional (3D) constructive interference in steady state (CISS) sequence in the assessment of syringomyelia. Eleven patients with syringomyelia were prospectively studied with magnetic resonance imaging. All patients underwent sagittal imaging with T1- and T2 weighted spin-echo (SE), and 3D-CISS sequences. The SE and 3D-CISS images, as well as multiplanar reconstruction (MPR) images of the 3D-CISS sequence, were analyzed with regard to image quality, degree of artifacts, visualization of the extent and internal structure of the syringomyelia, and contrast-to-noise ratio (CNR) of the fluid within the syringomyelia. Contrast between the spinal cord and cerebrospinal fluid (CSF), as well as delineation was significantly poorer for the T1-weighted SE sequence than for the 3D-CISS sequence (P < 0.01), while there was no significant difference between the T2-weighted SE sequence and the 3D-CISS sequence. Artifacts induced by CSF flow were significantly more for the T2 weighted SE sequence than for the 3D-CISS sequence (P < 0.01). Although the extent of syringomyelia was delineated equally among the three sequences in 9 of 11 patients, it was better for the 3D-CISS sequence than for the SE sequences in the remaining two. Septation and communication between the cavities were best detected by the 3D-CISS MPR images. The CNR of the 3D-CISS sequence was significantly higher than that of the SE sequence (P < 0.01). The 3D-CISS sequence demonstrates the extent and internal structures of syringomyelia better than conventional SE sequences and should be added to SE sequences in the evaluation of syringomyelia. PMID- 10713944 TI - Crohn's disease evaluation: comparison of contrast-enhanced MR imaging and single phase helical CT scanning. AB - The purpose of this study was to evaluate the use of gadolinium and barium enhanced magnetic resonance (MR) imaging in detecting intestinal and extraintestinal Crohn's disease and compare MRI with contrast-enhanced helical computed tomography (CT). Twenty-six patients with Crohn's disease underwent imaging examinations, including gadolinium-enhanced, fat suppressed fast multiplanar spoiled gradient-recalled (FMPSPGR) MR imaging with oral 2% barium sulfate and rectal water and with helical CT using i.v. and positive (13) or negative (13) intestinal contrast material. MR images and CT scans were reviewed separately by two radiologists for bowel wall thickness and enhancement, presence of abscess, phlegmon, and fistula. MR images and CT scans were then compared side by side. Surgical, endoscopic, and histopathologic findings and results of barium studies were reviewed to determine the location and severity of involvement of intestinal Crohn's disease. Depiction of mural thickening and/or enhancement was superior on the MR images, which showed 55 (85%) and 52 (80%) of 65 abnormal bowel segments for the two observers, compared with helical CT, which showed 39 (60%) and 42 (65%; P < 0.001, P < 0.05) of bowel segments affected by Crohn's disease. Segments of bowel with moderate or marked mural thickening were depicted equally on MR imaging and helical CT. In mildly diseased segments of bowel, with only slight thickening and enhancement, MR imaging depicted 22 (79%) and 19 (68%) of 28 segments, compared with helical CT, which depicted 9 (32%; P < 0.01), and 13 (46%; P > 0.05) of 28 segments. In the side-by side comparison, MR imaging was preferred over helical CT for depicting normal bowel wall (MR 71%, CT 4%, equal 25%; P < 0.001), mural thickening (MR 41%, CT 11% equal 48%; P < 0.01), mural enhancement (MR 89%, equal 11%; P < 0.001), and overall GI tract evaluation (MR 52%, CT 10%, equal 38%; P < 0.001). Gadolinium-enhanced MR imaging with oral dilute barium sulfate and rectal water depicts intestinal and extraintestinal changes of Crohn's disease and shows promise as a clinically useful tool. PMID- 10713945 TI - Enlargement of hilar periportal space: a sign of early cirrhosis at MR imaging. AB - The purpose of this study was to determine whether the finding of an enlarged hilar periportal space is a sign for early cirrhosis at magnetic resonance (MR) imaging. Forty-one pathologically proved cirrhotic patients in the early stage of disease who did not show conventional imaging findings of cirrhosis (early cirrhosis group) and 47 patients without history of chronic liver diseases (control group) were included in this study. MR images were qualitatively and quantitatively evaluated for the presence of enlargement of the periportal space. Enlargement of the periportal space was seen in 98% of patients in the early cirrhosis group, while this finding was seen in 11% of patients in the control group (P < 0.0001). The mean value of the hilar periportal fat thickness was significantly greater (P < 0.0001) in the early cirrhosis group (15.5 +/- 6.2 mm) than in the control group (5.3 +/- 3.1 mm). The sensitivity, specificity, accuracy, and positive predictive value of this finding for the MR diagnosis of cirrhosis with a cutoff value of 10 mm were 93%, 92%, 92%, and 91%, respectively. Enlargement of the hilar periportal space is a helpful sign at MR imaging in the discrimination between normal and early cirrhotic livers. PMID- 10713946 TI - Neuroendocrine tumors of the pancreas: spectrum of appearances on MRI. AB - We reviewed our 8.5 year experience with magnetic resonance imaging (MRI) in the demonstration of neuroendocrine tumors of the pancreas using precontrast fat suppressed T1-weighted, fat-suppressed T2-weighted, and serial post-gadolinium T1 weighted images, to describe the spectrum of appearances of these tumors. All MR examinations of patients with histologically proven neuroendocrine tumors were retrospectively reviewed. Histological type, tumor location, tumor diameter, signal intensity on precontrast images, enhancement patterns, and presence and appearance of metastases were determined. Twenty-two patients had histologically proved neuroendocrine tumors detected by MRI over the 8.5 year period. Histological types were gastrinoma (n = 8), insulinoma (n = 3), glucagonoma (n = 2), somatostatinoma (n = 1), VIPoma (n = 1), ACTHoma (n = 1), carcinoid (n = 1), and five untyped tumors. Primary tumors ranged in diameter from 1 to 6.2 cm. There was one histopathology-proven false-positive neuroendocrine tumor. The positive predictive value for MRI in the detection of these tumors was 96%. The most common appearance on precontrast images was low signal intensity on T1 weighted images and high signal intensity on T2-weighted images, which was observed in tumors in 18 of 22 patients. Moderate or intense early enhancement of all or portions of the primary tumors was observed in tumors in 19 of 22 patients either as uniform homogeneous, ring, or diffuse heterogeneous enhancement. Enhancement was minimal on these images in the other three patients. Gastrinomas enhanced in a ring pattern in 7 of 8 patients whereas the majority (9 of 11 patients) of noninsulinoma-nongastrinoma and untyped tumors enhanced in a diffuse heterogeneous fashion. Liver metastases were present in 13/22 patients including 3/8 with gastrinoma and 9/11 with noninsulinoma-nongastrinoma tumors. Most neuroendocrine tumors of the pancreas are low signal intensity on fat-suppressed T1-weighted images and moderately high in signal intensity on fat-suppressed T2 weighted images, although variations do exist. Tumors most often enhance in an early moderately intense fashion. Gastrinomas are often different in appearance than other neuroendocrine tumors in that they usually enhance in a ring fashion whereas nongastrinoma-noninsulinoma tumors usually enhance in a heterogeneous fashion. PMID- 10713947 TI - Reduction of noise in medullary renograms from dynamic MR images. AB - Dynamic magnetic resonance images of the kidney can be used to acquire separate renograms of the cortex and medulla. A high-quality cortical renogram can be determined directly from a region of interest (ROI) placed in the cortex. Due to partial volume effects, part of the signal from a ROI placed in the medulla is caused by cortical tissue. By subtracting a fraction of the cortical signal from the cortico-medullary signal, a purer medullary renogram can be obtained. A side effect of this subtraction is an increase in noise level. The noise level increases with larger partial volume fractions. Using a matched image filter, it is possible to exclude those areas from the ROI that have a high partial volume content, thus reducing the amount of cortical signal that has to be separated from the medullary signal. Noise reductions of up to 50% have been achieved in the medullary renogram, with an average reduction of 23%. PMID- 10713948 TI - Anisotropic diffusion in kidney: apparent diffusion coefficient measurements for clinical use. AB - The purpose of this study was to evaluate anisotropy of the kidney by measurements of the apparent diffusion coefficient (ADC) using commercially available magnetic resonance (MR) imaging. Fifty-one consecutive patients underwent diffusion-weighted echoplanar MR imaging of the upper abdomen with five different strengths of motion probing gradients (b = 1.51, 55.3, 36.6, 317, and 932 sec/mm2) applied along the z-axis. Four ADC values for the upper pole and central portion of the kidney were calculated from four different b-value ranges and compared. The ADCs for the kidney calculated in the lower b-value ranges were significantly higher than those in the higher ranges. The ADCs for the upper pole portion were significantly higher than those for the central portion except for one in the highest b-value range. Diffusion in the kidney is anisotropic, probably due to the kidney's radially oriented structures such as renal vessels and tubules. PMID- 10713949 TI - Effect of gradient and section orientation on quantitative analysis of knee joint cartilage. AB - The object of this study was to determine the influence of the gradient and section orientation on cartilage thickness and volume measurements in the knee joint. Eight specimens were imaged with a fat-suppressed gradient-echo sequence, applying sagittal, transverse, and coronal section orientations. Images were additionally acquired with exchanged gradient directions, and with computed tomography (CT) arthrography. After segmentation and three-dimensional (3D) reconstruction, the volume, the mean, and the maximal 3D cartilage thickness were computed. No effect of changes in the gradient orientation was found, suggesting that susceptibility-induced geometric distortion is not a relevant problem in quantitative cartilage imaging. Sagittal images produced similar data to that obtained with transverse (patella) or coronal (tibia) sections, demonstrating that all knee joint cartilages can be accurately quantified from a single sagittal data set. Whereas no significant systematic deviation between magnetic resonance imaging (MRI) and CT arthrography was recorded in the patella, there was a 10%-15% underestimation of tibial cartilage thickness in MRI. PMID- 10713950 TI - Microwave coagulation therapy: ex vivo comparison of MR imaging and histopathology. AB - We compared the findings of magnetic resonance (MR) images and pathological examination to determine whether or not MR images reflect pathological changes following microwave coagulation therapy (MCT) on liver tissue. We used microwave (generating frequency 2450 Mhz, wave length 12 cm, output 50 W, 60 second duration) to irradiate six canine livers under general anesthesia. After the animals were sacrificed, the livers were resected. The irradiated regions were cut with margins and divided into two pieces, one for MR study, and the other for pathological examination. The findings were compared. From the center to the marginal layer, the irradiated region presented 4/3 laminal patterns on T1/T2 weighted images: low/high, high/low, very high/high, and iso-low/high intensity. On gradient-echo imaging, the irradiated regions presented no decreasing signals using several echo time lengths. With hematoxylin and eosin stain, MR laminar patterns reflected the histopathological changes, as follows: a tissue loss area surrounding the inserted needle, low/high; decreased sinusoidal width with/without necrotic tissue, high/low; sinusoidal width dilation at the periphery, very high/high; and fatty degenerated tissue surrounding the irradiated area at the boundary of the normal hepatocytes, iso-low/high. The MR signal intensity, which reflected the histopathological changes, presented tissue characterization after MCT, and the macromolecular hydration effect influenced the high intensity on T1-weighted images. PMID- 10713951 TI - Image metric-based correction (autocorrection) of motion effects: analysis of image metrics. AB - Magnetic resonance (MR) imaging of the shoulder necessitates high spatial and contrast resolution resulting in long acquisition times, predisposing these images to degradation due to motion. Autocorrection is a new motion correction algorithm that attempts to deduce motion during imaging by calculating a metric that reflects image quality and searching for motion values that optimize this metric. The purpose of this work is to report on the evaluation of 24 metrics for use in autocorrection of MR images of the rotator cuff. Raw data from 164 clinical coronal rotator cuff exams acquired with interleaved navigator echoes were used. Four observers then scored the original and corrected images based on the presence of any motion-induced artifacts. Changes in metric values before and after navigator-based adaptive motion correction were correlated with changes in observer score using a least-squares linear regression model. Based on this analysis, the metric that exhibited the strongest relationship with observer ratings of MR shoulder images was the entropy of the one-dimensional gradient along the phase-encoding direction. We speculate (and show preliminary evidence) that this metric will be useful not only for autocorrection of shoulder MR images but also for autocorrection of other MR exams. PMID- 10713953 TI - Subacute clot mimicking flow in a thrombosed arterial bypass graft on two dimensional time-of-flight and three-dimensional contrast-enhanced MRA. AB - Subacute intravascular thrombus can contain methemoglobin, which results in very short spin-lattice (T1) relaxation times. We describe a case of a 78-year-old man with increasing right lower extremity claudication. The patient had a thrombosed arterial bypass graft showing high signal intensity that mimicked flow on both two-dimensional time-of-flight and three-dimensional contrast-enhanced MR angiography. Misinterpretation of the high signal thrombus as flowing blood can be avoided by obtaining a precontrast T1-weighted sequence. PMID- 10713952 TI - Physical, chemical, and biological evaluations of P760: a new gadolinium complex characterized by a low rate of interstitial diffusion. AB - An original gadolinium chelate, termed P760, which diffuses through the vascular endothelium but at a much lower rate than nonspecific agents (NSA), is described. P760 is a gadolinium macrocyclic compound based on a DOTA structure that is substituted by hydrophilic bulky groups branched on the amino-carboxylic residues. The molecular weight is 5293, and the molecular volume, measured by light scattering, is 30 times higher (11.5 nm3) than that of gadolinium (Gd)-DOTA (0.38 nm3). The increase in molecular volume and weight has two consequences: a) higher relaxivity (r1; 24.7 mM-1.s-1 compared with 3.4 mM-1.s-1 for Gd-DOTA at 20 Mhz, 37 degrees C); and b) a lengthening of its transport rate through the endothelium. P760 presents a peculiar pharmacokinetic profile: at early times post injection, the blood concentrations are higher than those of Gd-DOTA, but after 20 minutes, the blood concentrations are equal for the two compounds. The body clearances of the products are identical (i.e., glomerular filtration rate). P760 molecules are large enough to have a restricted diffusion through the endothelium but, conversely, small enough to pass freely through the glomerular membrane. This limited extravasation has been observed in rabbits by magnetic resonance angiography or in investigations of tumor permeability. Further experimental imaging studies are needed to define the clinical interest of such properties. PMID- 10713954 TI - Breath-hold 3D gradient-echo MR imaging of the lung parenchyma: evaluation of reproducibility of image quality in normals and preliminary observations in patients with disease. AB - This study evaluates the reproducibility and image quality of a three-dimensional (3D) gradient-echo sequence for imaging the lung parenchyma, with and without gadolinium administration, using a 2D spoiled gradient-echo sequence for comparison. Twenty patients without lung disease (normals) and five patients with lung disease (lung disease) underwent paired 2D and 3D gradient-echo sequences, without contrast (24 patients) and with contrast (18 patients). Images were retrospectively reviewed independently in a blinded fashion by two investigators. Artifacts and demonstration of central lung, peripheral lung, heart, pulmonary arteries, and esophagus were evaluated. Image quality of the central lung was rated as fair or good in 5 and 4 (reader one and two) patients with non-contrast 2D gradient-echo, 24 and 25 patients with non-contrast 3D gradient-echo, 3 and 1 patient(s) with contrast-enhanced 2D gradient-echo, and 19 and 19 patients with contrast-enhanced 3D gradient-echo imaging. Differences in image quality between 2D and 3D sequences were significant (P < 0.001). Heart-related phase artifacts were negligible in 2 and 0 patients with non-contrast 2D gradient-echo, 23 and 25 patients with non-contrast 3D gradient-echo, 0 and 0 patients with contrast enhanced 2D gradient-echo, and 17 and 19 patients with contrast-enhanced 3D gradient-echo imaging. Differences in heart-related phase artifact in the central lung between 2D and 3D sequences were significant (P = 0.001). Infiltrates, lung cancer, and pulmonary metastasis were better shown on the gadolinium-enhanced 3D gradient-echo sequences than on the other sequences. Breath-hold 3D gradient-echo imaging results in good image quality and negligible image artifacts and is superior to 2D spoiled gradient-echo imaging. Preliminary results in patients with disease appear promising. PMID- 10713955 TI - Dynamic three-dimensional magnetic resonance abdominal angiography and perfusion: implementation and preliminary experience. AB - Implementation of and preliminary experience with an ultra-fast partial-Fourier radiofrequency (RF) spoiled gradient-echo sequence for gadolinium-enhanced imaging are presented. Three-dimensional angiograms can be acquired in less than 6 seconds. Repetition of the acquisition allows the three-dimensional visualization of several distinct vascular phases. Feasibility is demonstrated in three healthy volunteers. The trade-offs among spatial resolution, temporal resolution, and spatial coverage as well as the technical aspects of gadolinium enhanced pulse sequences are discussed. PMID- 10713956 TI - Retrospective adaptive motion correction for navigator-gated 3D coronary MR angiography. AB - Retrospective adaptive motion correction (AMC) was developed for reducing effects of residual respiration in real-time navigator-gated three-dimensional (3D) coronary magnetic resonance (MR) angiography. In both motion phantom and in vivo experiments, AMC improved image sharpness of coronary arteries. This navigator based technique combining adaptive correction and real-time gating is potentially an efficient and effective motion reduction method for 3D coronary MR angiography. PMID- 10713958 TI - A versatile, low-cost method for presenting visual stimuli during MRI. AB - A visual presentation system is described that is inexpensive, easily implemented, and adaptable to most laboratory and clinical settings. Performance evaluations demonstrated a field of view of 26 x 56 degrees, no radiofrequency interference, and high image quality. Although the system was designed for brain activation studies with functional magnetic resonance imaging (MRI), it can also be used for positron emission tomography activation studies and for presenting relaxation videos during clinical MRI. PMID- 10713957 TI - In vivo Hahn spin-echo decay (Hahn-T2) observation of regional changes in the time course of oleic acid lung injury. AB - We studied the time course of changes in the Hahn spin-echo decay (Hahn-T2) in lungs of spontaneously breathing living rats at 1 hour, 3 hours, and 7 days following oleic acid injection. Motion artifacts were minimized by using the motion-insensitive interleaved rapid line scan (ILS) imaging technique. Prior to injury, the lungs exhibited two resolvable exponential Hahn-T2 components. One and 3 hours after injury the decay showed a regionally nonuniform behavior, which was fit with one, two, or three exponential components. The short and medium components increased at 1 and 3 hours after injection. The third, much longer, component is probably due to intraalveolar pulmonary edema. After 7 days the Hahn decay was similar to that observed before injury, probably reflecting resolution of the edema. Our data suggest that Hahn-T2 measurements can be used to characterize the time course and regional distribution of lung injury in living animals. PMID- 10713959 TI - Resampling as a cluster validation technique in fMRI. AB - Exploratory, data-driven analysis approaches such as cluster analysis, principal component analysis, independent component analysis, or neural network-based techniques are complementary to hypothesis-led methods. They may be considered as hypothesis generating methods. The representative time courses they produce may be viewed as alternative hypotheses to the null hypothesis, i.e., "no activation." We present here a resampling technique to validate the results of exploratory fuzzy clustering analysis. In this case an alternative hypothesis is represented by a cluster centroid. For both simulated and in vivo functional magnetic resonance imaging data, we show that by permutation-based resampling, statistical significance may be computed for each voxel belonging to a cluster of interest without parametric distributional assumptions. PMID- 10713960 TI - The role of static and dynamic fusimotor activity during locomotion. PMID- 10713961 TI - Inhibition of KCNQ1-4 potassium channels expressed in mammalian cells via M1 muscarinic acetylcholine receptors. AB - 1. KCNQ1-4 potassium channels were expressed in mammalian Chinese hamster ovary (CHO) cells stably transfected with M1 muscarinic acetylcholine receptors and currents were recorded using the whole-cell perforated patch technique and cell attached patch recording. 2. Stimulation of M1 receptors by 10 microM oxotremorine-M (Oxo-M) strongly reduced (to 0-10%) currents produced by KCNQ1-4 subunits expressed individually and also those produced by KCNQ2 + KCNQ3 and KCNQ1 + KCNE1 heteromers, which are thought to generate neuronal M-currents (IK,M) and cardiac slow delayed rectifier currents (IK,s), respectively. 3. The activity of KCNQ2 + KCNQ3, KCNQ2 and KCNQ3 channels recorded with cell-attached pipettes was strongly and reversibly reduced by Oxo-M applied to the extra-patch membrane. 4. It is concluded that M1 receptors couple to all known KCNQ subunits and that inhibition of KCNQ2 + KCNQ3 channels, like that of native M-channels, requires a diffusible second messenger. PMID- 10713962 TI - Solvent effects on squid sodium channels are attributable to movements of a flexible protein structure in gating currents and to hydration in a pore. AB - 1. Solvent effects on the time course of gating and sodium currents were analysed in squid sodium channels using four non-electrolytes of different size, glycerol, erythritol, glucose and sucrose, to separate effects of viscosity from those of osmolarity and to obtain viscosity and osmolarity parameters that were independent of molecular size. 2. The gating and sodium currents were reversibly slowed in a voltage-independent manner as the non-electrolyte concentration increased. 3. Solvent effects were analysed using a model in which the percentage change in time constant was expressed by an equation involving the viscosity parameter alpha and the osmolarity parameter delta: t/t0 = alpha (eta/eta 0) - 1 + 100 alpha-1)exp(delta delta pi), where eta/eta 0 is solution viscosity and delta pi is increase in osmolarity. Since the solution viscosity was found experimentally to be a function of the solution osmolarity, solvent effects are described by an equation with one independent variable eta/eta 0 or delta pi. 4. Voltage sensor movement, reflected in gating currents, was primarily sensitive to viscosity, as its decay time constant was a function of eta/eta 0, with only a minor sensitivity to osmolarity (delta was 2-3 water molecules). 5. For sodium currents, alpha was equal to that of gating currents but delta was 2-3 times greater, suggesting that the final channel opening was primarily sensitive to osmolarity (delta delta was 5 water molecules). The relative ineffectiveness of the largest non-electrolyte, sucrose, suggested that this osmolarity-sensitive step in channel opening occurred in the narrow pore region. 6. Sodium channel inactivation was primarily sensitive to osmolarity (delta delta was 8-12 water molecules). 7. The observed viscosity dependence of the sodium current activation and inactivation processes was attributable to the viscosity-dependent process accompanying the gating current. 8. This model explains why non-electrolytes slow sodium currents while electrolytes do not. 9. Viscosity effects on gating currents can be explained by a process in which non-electrolytes interact with the flexible hydrophilic parts of sodium channel proteins, but osmolarity effects on the final step need to be explained by a local interaction of several water molecules with fluctuating protein segments in the pore. PMID- 10713963 TI - Mitochondrial Ca2+ homeostasis during Ca2+ influx and Ca2+ release in gastric myocytes from Bufo marinus. AB - 1. The Ca(2+)-sensitive fluorescent indicator rhod-2 was used to monitor mitochondrial Ca2+ concentration ([Ca2+]m) in gastric smooth muscle cells from Bufo marinus. In some studies, fura-2 was used in combination with rhod-2, allowing simultaneous measurement of cytoplasmic Ca2+ concentration ([Ca2+]i) and [Ca2+]m, respectively. 2. During a short train of depolarizations, which causes Ca2+ influx from the extracellular medium, there was an increase in both [Ca2+]i and [Ca2+]m. The half-time (t1/2) to peak for the increase in [Ca2+]m was considerably longer than the t1/2 to peak for the increase in [Ca2+]i. [Ca2+]m remained elevated for tens of seconds after [Ca2+]i had returned to its resting value. 3. Stimulation with caffeine, which causes release of Ca2+ from the sarcoplasmic reticulum (SR), also produced increases in both [Ca2+]i and [Ca2+]m. The values of t1/2 to peak for the increase in [Ca2+] in both cytoplasm and mitochondria were similar; however, [Ca2+]i returned to baseline values much faster than [Ca2+]m. 4. Using a wide-field digital imaging microscope, changes in [Ca2+]m were monitored within individual mitochondria in situ, during stimulation of Ca2+ influx or Ca2+ release from the SR. 5. Mitochondrial Ca2+ uptake during depolarizing stimulation caused depolarization of the mitochondrial membrane potential. The mitochondrial membrane potential recovered considerably faster than the recovery of [Ca2+]m. 6. This study shows that Ca2+ influx from the extracellular medium and Ca2+ release from the SR are capable of increasing [Ca2+]m in smooth muscle cells. The efflux of Ca2+ from the mitochondria is a slow process and appears to be dependent upon the amount of Ca2+ in the SR. PMID- 10713964 TI - Protein kinase C enhances the rapidly activating delayed rectifier potassium current, IKr, through a reduction in C-type inactivation in guinea-pig ventricular myocytes. AB - 1. The rapidly activating delayed rectifier potassium current, IKr, was studied in guinea-pig ventricular myocytes in the presence of thiopentone, which blocks the more slowly activating component of the delayed rectifier potassium current, IKs, and using whole cell perforated patch clamp or switched voltage clamp with sharp electrodes to minimise intracellular dialysis. 2. Activation of protein kinase A (PKA) by isoprenaline or forskolin caused an increase in IKr tail currents. Following a 300 ms depolarising step to +20 mV, mean tail current amplitude was increased 47 +/- 12% by isoprenaline, and 73 +/- 13% by forskolin. No increase in IKr was observed when IKr was studied using whole cell ruptured patch clamp and there was no change in the reversal potential of IKr in the presence of isoprenaline. 3. The rectification of the current sensitive to E4031, a selective IKr blocker, was markedly reduced in the presence of isoprenaline and the region of negative slope was absent. This is consistent with a reduction in the inactivation of IKr and was supported by the finding that IKr, in the presence of isoprenaline, was somewhat less sensitive to block. E4031 (5 microM) blocked only 81 +/- 5% of IKr in the presence of isoprenaline compared to 100 +/- 0% in control. 4. The forskolin- and isoprenaline-induced increases in IKr were inhibited by staurosporine and by the selective protein kinase C (PKC) inhibitor bisindolymaleimide I. Direct activation of PKC by phorbol dibutyrate increased IKr tail currents by 24 +/- 5%. Both the isoprenaline- and forskolin-induced increases in IKr were inhibited when calcium entry was reduced by block of ICa with nifedipine or when myocytes were pre-incubated in BAPTA-AM. 5. The selective PKA inhibitor KT5720 prevented the isoprenaline-induced increase in IKr only when the increase in ICa was also suppressed. 6. These data show a novel mechanism of regulation of IKr by PKC and this kinase was activated by beta-adrenoceptor stimulation. IKr seems to be enhanced through a reduction in the C-type inactivation which underlies the rectification of the channel and such a mechanism may occur in other channels with this type of inactivation. PMID- 10713965 TI - Carbachol-induced [Ca2+]i increase, but not activation of protein kinase C, stimulates exocytosis in rat parotid acini. AB - 1. A column perfusion system was applied to rat parotid acinar cells to clarify the roles of Ca2+ and protein kinase C (PKC) in the mechanisms of carbachol (CCh) induced amylase secretion. 2. CCh evoked a biphasic response of amylase secretion with an initial rapid and large peak that reached maximum at about 10 s followed by a sustained plateau. The time profile and the dose-response relationship paralleled with those of cytosolic free Ca2+ concentration ([Ca2+]i). 3. The CCh induced sustained response of amylase secretion maintained by Ca2+ influx into cells was ATP dependent, while the initial peak response regulated by Ca2+ released from InsP3-sensitive stores was relatively ATP independent. 4. Restoration of extracellular Ca2+ during continuous stimulation with CCh in Ca(2+)-free medium evoked a second rapid and large peak of amylase secretion. 5. Phorbol 12,13-dibutyrate (PDBu) potentiated the CCh-induced amylase secretion in both the initial peak and the sustained plateau without enhancing CCh-induced [Ca2+]i changes. 6. PKC inhibitors such as Ro 31-8220 inhibited the potentiating effect of PDBu but only slightly reduced amylase secretion induced by CCh alone. 7. These results suggest that a CCh-induced rise in [Ca2+]i triggers the final fusion and/or exocytosis of amylase secretion. CCh also has some ability to promote ATP-dependent priming of secretory granules that, together with Ca2+ influxed into cells, contributes to the CCh-induced sustained plateau of amylase secretion. PDBu-induced activation of PKC promotes the priming of secretory granules, thereby enhancing the efficacy for Ca2+ to trigger fusion/exocytosis. PMID- 10713966 TI - Diverse roles of intracellular cAMP in early synaptic modifications in the rat visual cortex. AB - 1. The effects of increasing intracellular cAMP concentration were studied using photolysis of caged-cAMP in layer II/III neurons recorded intracellularly in visual cortex slices. The recorded neurons exhibited either after hyperpolarization (AHP) or after-depolarization (ADP) in response to depolarizing current injection. Depending on which afterpotential appeared, the effects of photolysis differed. 2. In ADP-generating neurons, photolysis of caged-cAMP induced long-lasting depression of postsynaptic potentials (PSPs) evoked by grey matter (GM) stimulation, without altering the size of the ADP. In AHP-generating neurons, photolysis induced long-lasting potentiation of GM-evoked PSPs, with the size of the AHP reduced in the same time course. White matter (WM)-evoked PSPs showed no change. 3. Extracellular application of bromo-cAMP depressed both GM- and WM-evoked PSPs in ADP- and AHP-generating neurons. This depression may be due to presynaptic effects of cAMP, since photolysis-evoked postsynaptic increase in cAMP concentration never induced depression of PSPs in AHP-generating neurons. This depression was reversible but continued until bromo-cAMP was washed out, while ADP and AHP in the postsynaptic neurons were depressed only temporarily and returned to the pre-application level even in the continued presence of bromo cAMP. 4. Bromo-cAMP was applied following photolysis of caged-cAMP. In the neurons in which the photolysis potentiated GM-evoked PSPs this potentiation was cancelled out by bromo-cAMP (depotentiation). In the other neurons, PSPs were depressed only reversibly. 5. Thus, a postsynaptic increase in cAMP concentration exerts more diverse effects on synaptic plasticity than thus far reported, depending on the difference in neuronal intrinsic excitability and probably on how much, or the way in which, cAMP concentration is increased. PMID- 10713967 TI - Activity-dependent extracellular K+ accumulation in rat optic nerve: the role of glial and axonal Na+ pumps. AB - 1. We measured activity-dependent changes in [K+]o with K(+)-selective microelectrodes in adult rat optic nerve, a CNS white matter tract, to investigate the factors responsible for post-stimulus recovery of [K+]o. 2. Post stimulus recovery of [K+]o followed a double-exponential time course with an initial, fast time constant, tau fast, of 0.9 +/- 0.2 s (mean +/- S.D.) and a later, slow time constant, tau slow, of 4.2 +/- 1 s following a 1 s, 100 Hz stimulus. tau fast, but not tau slow, decreased with increasing activity dependent rises in [K+]o. tau slow, but not tau fast, increased with increasing stimulus duration. 3. Post-stimulus recovery of [K+]o was temperature sensitive. The apparent temperature coefficients (Q10, 27-37 degrees C) for the fast and slow components following a 1 s, 100 Hz stimulus were 1.7 and 2.6, respectively. 4. Post-stimulus recovery of [K+]o was sensitive to Na+ pump inhibition with 50 microM strophanthidin. Following a 1 s, 100 Hz stimulus, 50 microM strophanthidin increased tau fast and tau slow by 81 and 464%, respectively. Strophanthidin reduced the temperature sensitivity of post-stimulus recovery of [K+]o. 5. Post stimulus recovery of [K+]o was minimally affected by the K+ channel blocker Ba2+ (0.2 mM). Following a 10 s, 100 Hz stimulus, 0.2 mM Ba2+ increased tau fast and tau slow by 24 and 18%, respectively. 6. Stimulated increases in [K+]o were followed by undershoots of [K+]o. Post-stimulus undershoot amplitude increased with stimulus duration but was independent of the peak preceding [K+]o increase. 7. These observations imply that two distinct processes contribute to post stimulus recovery of [K+]o in central white matter. The results are compatible with a model of K+ removal that attributes the fast, initial phase of K+ removal to K+ uptake by glial Na+ pumps and the slower, sustained decline to K+ uptake via axonal Na+ pumps. PMID- 10713968 TI - Spontaneous photo-relaxation of urethral smooth muscle from sheep, pig and rat and its relationship with nitrergic neurotransmission. AB - 1. In the present work we have characterized the relaxant response induced by light stimulation (LS) in the lower urinary tract from sheep, pig and rat, establishing its relationship with nitrergic neurotransmission. 2. Urethral, but not detrusor, preparations showed pronounced photo-relaxation (PR) which declined progressively following repetitive LS. Sheep urethral PR was again restored either spontaneously or (to a greater extent) by exogenous nitric oxide (NO) addition and by electrical field stimulation (EFS) of intrinsic nitrergic nerves. 3. Greater NO generation was detected from sheep urethral than from detrusor homogenates following illumination. 4. Sheep urethral PR was inhibited by oxyhaemoglobin, but not by methaemoglobin, carboxy-PTIO, extracellular superoxide anion generators or superoxide dismutase. Guanylyl cyclase but not adenylyl cyclase activation mediates urethral relaxation to LS. 5. Urethral PR was more resistant to inhibition by L-thiocitrulline than EFS-induced responses, although this agent prevented PR restoration by high-frequency EFS. 6. Urethral PR was TTX insensitive and partially modified in high-K+ solutions. Cold storage for 24 h greatly impaired urethral PR, although it was restored by high-frequency EFS. 7. Repetitive exposure to LS, EFS or exogenous NO induced changes in the shape of the EFS-induced nitrergic relaxation, possibly by pre-synaptic mechanisms. 8. In conclusion, we suggest the presence of an endogenous, photo-labile, nitro compound store in the urethra, which seems to be replenished by neural nitric oxide synthase activity, indicating a close functional relationship with the nitrergic neurotransmitter. PMID- 10713969 TI - The capacity of mdx mouse diaphragm muscle to do oscillatory work. AB - 1. Mdx mice were used as a model for Duchenne muscular dystrophy; both lack dystrophin. It was hypothesized that the mdx condition would have a marked effect on the ability of diaphragm muscle from mdx mice to do active net work and generate power. This hypothesis was tested using the work-loop technique. 2. Specific twitch force, specific tetanic force and maximum power were all significantly less in diaphragm strips from mdx mice than those from control mice. 3. In all preparations muscle length at which maximum power was achieved (Lw) was about 8% less than that at which maximum tetanic force was achieved (L0), both in mdx and control muscle. 4. The isometric force-length curve for mdx muscle was steeper on both sides of the plateau. Similarly, the curve relating net work per cycle to muscle length was steeper for mdx muscle on both sides of the plateau. 5. Maximum power of mdx muscle was achieved at a lower strain than for control muscle; maximum power occurred at a strain of 10.2% for mdx and 14.7% for control. Further increases in strain caused a marked decrease of power production in mdx muscle, whereas they caused a smaller decrease in control muscle. 6. In summary, at muscle lengths longer than Lw and at high strains, performance of mdx muscle was compromised relative to that of control muscle. Work and power were compromised more than isometric force. PMID- 10713970 TI - Evidence for control of adenosine metabolism in rat oxidative skeletal muscle by changes in pH. AB - 1. We investigated the effects of pH elevation or depression on adenosine output from buffer-perfused rat gracilis muscle, and kinetic properties of adenosine forming enzymes, 5'-nucleotidase (5'N) and non-specific phosphatase (PT), and adenosine-removing enzymes, adenosine kinase (AK) and adenosine deaminase (AD), in homogenates of muscle. 2. Depression of the perfusion buffer pH from 7.4 to 6.8, by addition of sodium acetate, reduced arterial perfusion pressure from 8.44 +/- 1.44 to 7.33 +/- 0.58 kPa, and increased adenosine output from 35 +/- 5 to 56 +/- 6 pmol min-1 (g wet wt muscle)-1 and AMP output from 1.8 +/- 0.3 to 9.1 +/- 3.9 pmol min-1 (g wet wt muscle)-1. 3. Elevation of the buffer pH to 7.8, by addition of ammonium chloride, reduced arterial perfusion pressure from 8.74 +/- 0.57 to 6.96 +/- 1.37 kPa, and increased adenosine output from 25 +/- 5 to 47 +/- 8 pmol min-1 (g wet wt muscle)-1 and AMP output from 3.7 +/- 1.1 to 24.6 +/- 6.8 pmol min-1 (g wet wt muscle)-1. 4. Activity of membrane-bound 5'N was an order of magnitude higher than that of either cytosolic 5'N or PT: pH depression reduced the K(m) of 5'N, which increased its capacity to form adenosine by 10-20% for every 0.5 unit decrease inpH within the physiological range. PT was only found in the membrane fraction: its contribution to extracellular adenosine formation increased from about 5% at pH 7.0 to about 15% at pH 8.0. 5. Cytosolic 5'N had a low activity, which was unaffected by pH; the rate of intracellular adenosine formation was an order of magnitude lower than the rate of adenosine removal by adenosine kinase or adenosine deaminase, which were both exclusively intracellular enzymes. 6. We conclude that (i) adenosine is formed in the extracellular compartment of rat skeletal muscle, principally by membrane-bound 5'N, where it is protected from enzymatic breakdown; (ii) adenosine is formed intracellularly at a very low rate, and is unlikely to leave the cell; (iii) enhanced adenosine formation at low pH is driven by an increased extracellular AMP concentration and an increased affinity of membrane-bound 5'N for AMP; (iv) enhanced adenosine formation at high pH is driven solely by the elevated extracellular AMP concentration, since the catalytic capacity of membrane 5'N is reduced at high pH. PMID- 10713971 TI - The effect of serum iron concentration on iron secretion into mouse milk. AB - 1. The concentration of iron in mouse milk is approximately 3 times that of the serum. Although there is clear evidence for the presence of the transferrin receptor in the rodent mammary gland, the precise mechanisms of iron transfer into milk are not known. 2. Milk iron was linearly related to the serum iron:transferrin ratio in lactating mice whose serum iron ranged from 8 to 66 microM. 3. Increasing the iron binding capacity of the milk by 340 microM by targeting the lactoferrin transgene to the mammary gland did not alter the relation between milk iron and the serum iron:transferrin ratio. 4. The steady state distribution ratio of 125I-transferrin between plasma and milk was about 0.2, indicating that transcytosed transferrin contributed a maximum of 6% of the milk iron. 5. Fluorescently labelled transferrin incubated with the in situ gland localized mainly near the basal surface of the mammary alveolar cells. 6. These experiments provide evidence that the initial and rate-limiting step in the transfer of iron into milk is binding to a basal transferrin receptor. 7. A theoretical model of the relation between milk and serum iron suggests that the affinity of apotransferrin for the basal recycling system may be higher than observed in many other cell types. PMID- 10713972 TI - Ventilatory and central neurochemical reorganisation of O2 chemoreflex after carotid sinus nerve transection in rat. AB - 1. The first step of this study was to determine the early time course and pattern of hypoxic ventilatory response (HVR) recovery following irreversible bilateral carotid sinus nerve transection (CSNT). The second step was to find out if HVR recovery was associated with changes in the neurochemical activity of the medullary catecholaminergic cell groups involved in the O2 chemoreflex pathway. 2. The breathing response to acute hypoxia (10% O2) was measured in awake rats 2, 6, 10, 45 and 90 days after CSNT. In a control group of sham-operated rats, the ventilatory response to hypoxia was principally due to increased respiratory frequency. There was a large reduction in HVR in the CSNT compared to the sham operated rats (-65%, 2 days after surgery). Within the weeks following denervation, the CSNT rats progressively recovered a HVR level similar to the sham-operated rats (-37% at 6 days, -27% at 10 days, and no difference at 45 or 90 days). After recovery, the CSNT rats exhibited a higher tidal volume (+38%) than the sham-operated rats in response to hypoxia, but not a complete recovery of respiratory frequency. 3. Fifteen days after CSNT, in vivo tyrosine hydroxylase (TH) activity had decreased in caudal A2C2 (-35%) and A6 cells ( 35%). After 90 days, the CSNT rats displayed higher TH activity than the sham operated animals in caudal A1C1 (+51%), caudal A2C2 (+129%), A5 (+216%) and A6 cells (+79%). 4. It is concluded that HVR following CSNT is associated with a profound functional reorganisation of the central O2 chemoreflex pathway, including changes in ventilatory pattern and medullary catecholaminergic activity. PMID- 10713973 TI - Tension changes in the cat soleus muscle following slow stretch or shortening of the contracting muscle. AB - 1. The permanent extra tension after a stretch and the deficit of tension after a shortening in the soleus muscle of the anaesthetised cat were measured using distributed nerve stimulation across five channels. At low rates of stimulation the optimum length for a contraction was several millimetres longer than that when higher rates of stimulation were used, so that movements applied over the same length range could be on the descending limb of the full activation curve but on the ascending limb of the submaximal activation curve. 2. The extra tension after stretch and the depression after shortening were present only near the peak and on the descending limb of the length-tension curve. Effects on final tension of changing the speed and amplitude of stretches or shortenings were found to be small. 3. Statistical analysis showed that variations in the tension excess or deficit due to changing stimulus rate could be entirely attributed to the effect of stimulus rate on the length-tension relation, as when length was expressed relative to optimum for each rate, stimulus rate was no longer a significant determinant of the tension excess or deficit. 4. The extra tension after stretch and the depression after shortening disappeared if stimulation was interrupted and tension briefly fell to zero. 5. These effects were explained in terms of a non-uniform distribution of sarcomere length changes at long muscle lengths. During stretch some sarcomeres are stretched to beyond overlap while others lengthen hardly at all. During shortening some sarcomeres shorten much further than others. 6. These mechanisms have important implications for exercise physiology and sports medicine. PMID- 10713974 TI - Patterns of fusimotor activity during locomotion in the decerebrate cat deduced from recordings from hindlimb muscle spindles. AB - 1. Recordings have been made from multiple single muscle spindle afferents from medial gastrocnemius (MG) and tibialis anterior (TA) muscles of one hindlimb in decerebrate cats, together with ankle rotation and EMG signals, during treadmill locomotion. Whilst the other three limbs walked freely, the experimental limb was denervated except for the nerves to MG and TA and secured so that it could rotate only at the ankle joint, without any external load. Each afferent was characterised by succinylcholine testing with regard to its intrafusal fibre contacts. Active movements were recorded and then replayed through a servo mechanism to reproduce the muscle length changes passively after using a barbiturate to suppress gamma-motor firing. 2. The difference in secondary afferent firing obtained by subtracting the discharge during passive movements from that during active movements was taken to represent the profile of static fusimotor activity. This indicated an increase before the onset of movement followed by a strongly modulated discharge in parallel with muscle shortening during locomotion. The pattern of static firing matched the pattern of unloaded muscle shortening very closely in the case of TA and with some phase advance in the case of MG. The same effects were observed in primary afferents. 3. Primary afferents with bag1 (b1) contacts in addition showed higher firing frequencies during muscle lengthening in active than in passive movements. This indicated increased dynamic fusimotor firing during active locomotion. There was no evidence as to whether this fluctuated during the movement cycles. 4. When the mean active minus passive difference profile of firing in bag2-chain (b2c) type primary afferents was subtracted from that for b1b2c afferents, the difference was dominated by a peak centred on the moment of maximum lengthening velocity (v). 5. The component of the active minus passive difference firing due to b1 fibre contacts could be modelled by f(t) = av (where a is a constant) during lengthening and by f(t) = 0.2 av during shortening. The remainder of the difference signal matched the predictions of the static fusimotor signal derived from secondary afferents. 6. The findings are discussed in relation to the concept that the modulated static fusimotor pattern may represent a 'temporal template' of the expected movement, though the relationship of the results to locomotion in the intact animal will require further investigation. The analysis of the data indicates that the combined action of muscle length changes and static and dynamic fusimotor activity to determine primary afferent firing can be understood in terms of the interaction between the b1 and b2c impulse initiation sites. PMID- 10713975 TI - Effect of triple therapy on eradication of canine gastric helicobacters and gastric disease. AB - Nine helicobacter-positive pet dogs with upper gastrointestinal signs were studied to evaluate the effect of a triple therapy, normally applied to humans for the eradication of gastric helicobacters, on clinical signs and gastric histology, as well as the recurrence of helicobacters after eradication in an extended follow-up in four dogs. Endoscopy was performed at entry to the study and repeated after eradication therapies and additional treatments. If the triple therapy (amoxycillin, metronidazole and bismuth subcitrate) failed, tetracycline and omeprazole were prescribed. Additional therapies were instituted if clinical signs persisted after eradication therapies. Helicobacter status was verified from gastric biopsy specimens by the urease test and histological examination, and in a few dogs also by brush cytology. Triple therapy eradicated gastric helicobacters in 7/9 dogs; gastric helicobacters were also eradicated in one dog treated with tetracycline and omeprazole. Eradication of helicobacters resulted in significant improvement, but not total resolution, of clinical signs. Subsequent additional therapies resulted in further alleviation of clinical signs. Neither triple therapy nor additional therapies had a significant effect on gastric histological changes. Gastric helicobacters recurred in 4/4 dogs within three years of the eradication treatment. Because canine gastric helicobacters alone were not definitively shown to induce clinical signs, routine eradication therapy seems not to be warranted at present. PMID- 10713976 TI - Deionised water as an adjunct to surgery for the treatment of canine cutaneous mast cell tumours. AB - Medical records of 55 dogs with a diagnosis of cutaneous mast cell tumour were reviewed. Twenty-seven of the dogs were treated with surgery plus deionized water and the remaining 28 with surgery alone. A survival analysis was performed to determine whether deionized water, as an adjunct to surgery for cutaneous mast cell tumour, affected survival time or time to tumour recurrence. Dogs in which mast cell tumour recurred had a significantly shorter survival time compared with dogs with no recurrence (P = 0.05), regardless of the method of treatment. A significant negative association between tumour recurrence and method of treatment (P = 0.0097) and clinical stage (P = 0.0223) was observed. Dogs treated with surgery and deionized water had a significantly shorter time to recurrence of their mast cell tumour (P = 0.0113). Based on these results, deionized water does not appear to be beneficial in prolonging survival time or time to tumour recurrence for dogs with cutaneous mast cell tumours. PMID- 10713977 TI - Influence of feeding regimen on body condition in the cat. AB - The influence of feeding regimen on body condition in the cat was studied in a sample of the UK domestic cat population (n = 136). Data were collected by interviewing cat owners and assessing body condition of cats in the owners' homes. Four main factors were identified which were related to body condition. These were, in descending order of significance: neuter status, age, frequency of treat feeding, and ad libitum feeding. Feeding regimen as a risk factor in feline obesity is discussed. PMID- 10713978 TI - Factors affecting hip dysplasia in German shepherd dogs in Finland: efficacy of the current improvement programme. AB - Hip dysplasia records from 10,335 German shepherd dogs were used to estimate environmental effects and predict breeding values and genetic change achieved with the Finnish Kennel Club's breeding programme. The best linear unbiased prediction (BLUP) procedure was used for the analysis. No clear genetic improvement could be found during the study period (1985 to 1997). This might be due to ineffective selection for good hips. Significant environmental effects included year and month of birth, panellist, screening age and the effect of the genetic group of offspring from imported versus non-imported sires. In order to make the breeding programme more effective, BLUP breeding values should be used instead of phenotypic selection. PMID- 10713979 TI - Dermoid sinus at the lumbosacral junction in an English springer spaniel. AB - A dermoid sinus was identified in a springer spaniel that presented with hindlimb neurological deficits. The sinus was continuous with the dura mater at the level of the lumbosacral junction. The presence of hair and debris adjacent to neural tissue had elicited a myelitis. A dorsal laminectomy was required to allow complete surgical resection of the sinus. The dog's neurological status improved after treatment and this improvement was maintained over a five-month follow-up period. PMID- 10713981 TI - Six pearls to a successful defense. PMID- 10713980 TI - Extrahepatic biliary atresia in a border collie. AB - Progressive lameness and leg pain were the predominant clinical signs in a 17 week-old male border collie presented for examination. On clinical investigation, extrahepatic cholestasis in association with rickets due to inadequate vitamin D resorption was diagnosed. The dog was treated parenterally with vitamin D and a cholecystoduodenostomy was performed. At 25 days postsurgery the lameness had resolved and bone structure was radiographically normal. However, at six weeks postsurgery, the dog's condition deteriorated rapidly and euthanasia was finally performed at eight weeks postsurgery. At postmortem examination, Toxocara canis nematodes were found to have invaded the biliary system via the anastomosis between the gallbladder and duodenum, causing biliary and hepatic toxocariasis. The cause of the primary extrahepatic cholestasis was atresia of the common bile duct at the hepatic end. The liver tissue showed microscopic lesions of chronic extrahepatic cholestasis as well as acute inflammation associated with the nematode invasion. There was no postmortem evidence of bone lesions. Extrahepatic biliary atresia is extremely rare in animals and has not been described before in dogs. In contrast, it represents the most common cause of congenital cholestasis in children, occurring in approximately one per 10,000 to 15,000 live births. PMID- 10713982 TI - Intrapartum fetal pulse oximetry. Part I: Principles and technical issues. AB - Fetal pulse oximetry has been advocated as a means of improving the specificity of cardioto-cography in intrapartum fetal surveillance. The objective of this article, the first of two reviewing the current literature on fetal pulse oximetry, is to discuss the principles of this new and evolving technology, its development, and the various factors that can affect its readings during fetal monitoring. It serves as a prelude to the second article, which profiles the clinical application of fetal pulse oximetry. Literature pertaining to this topic was selected from a MEDLINE search from 1965 through September 1999, with additional sources obtained through cross-referencing. TARGET AUDIENCE: Obstetricians & Gynecologists, Family Physicians LEARNING OBJECTIVES: After completion of this article, the reader will be able to explain the principles of fetal pulse oximetry, and describe the factors that affect the calibration of fetal pulse oximeters and the factors that affect the fetal pulse oximeter readings. PMID- 10713983 TI - Intrapartum fetal pulse oximetry. Part 2: Clinical application. AB - Part II continues by discussing the clinical application of fetal pulse oximetry in intrapartum fetal surveillance as reviewed in the literature. It profiles the predictive ability of fetal pulse oximetry and its correlation with conventional methods of fetal monitoring. In addition, an account is made of its limitations, safety, and acceptability in clinical practice. TARGET AUDIENCE: Obstetricians & Gynecologists, Family Physicians LEARNING OBJECTIVES: After completion of this article, the reader will be able to describe the predictive abilities of fetal pulse oximetry and its correlation with conventional methods of fetal monitoring, to define the critical threshold of fetal oxygen saturation, and to summarize the safety of fetal pulse oximetry. PMID- 10713984 TI - Bell palsy complicating pregnancy: a review. AB - The aim of the present work was to review the published evidence on the association of Bell palsy (BP), an acute idiopathic peripheral facial paralysis of unknown etiology, with pregnancy. Reports have shown that women of reproductive age are affected two to four times more often than men of the same age, and pregnant women 3.3 times more often than nonpregnant women. The apparent predisposition of pregnant women to Bell palsy has been attributed to the high extracellular fluid content, viral inflammation, and immunosuppression characteristic of pregnancy, but findings are controversial. Most cases of Bell palsy occur in the third trimester or the puerperium. Onset is acute and painful. Some authors suggest that Bell palsy increases the risk of hypertension and toxemia of pregnancy, whereas the pregnant state, in turn, may affect the course and severity of disease. Recovery is usually good; poor prognostic markers are recurrence in subsequent pregnancy and bilateral disease, both of which are rare. Neonatal outcome is apparently unaffected, although this has been studied rarely. The preferred mode of management remains undecided; it is usually confined to supportive care. Corticosteroids in pregnancy are controversial. We think clinicians should be aware of these findings to avoid unnecessary testing and treatment and to help the patient cope with this acute, painful disease. TARGET AUDIENCE: Obstetricians & Gynecologists, Family Physicians LEARNING OBJECTIVES: After completion of this article, the reader will be able to identify the potential etiologies of Bell palsy associated with pregnancy and to describe the clinical presentation of this condition in pregnancy and its likelihood for recovery. PMID- 10713986 TI - An alternative view of protein fold space. AB - Comparing and subsequently classifying protein structures information has received significant attention concurrent with the increase in the number of experimentally derived 3-dimensional structures. Classification schemes have focused on biological function found within protein domains and on structure classification based on topology. Here an alternative view is presented that groups substructures. Substructures are long (50-150 residue) highly repetitive near-contiguous pieces of polypeptide chain that occur frequently in a set of proteins from the PDB defined as structurally non-redundant over the complete polypeptide chain. The substructure classification is based on a previously reported Combinatorial Extension (CE) algorithm that provides a significantly different set of structure alignments than those previously described, having, for example, only a 40% overlap with FSSP. Qualitatively the algorithm provides longer contiguous aligned segments at the price of a slightly higher root-mean square deviation (rmsd). Clustering these alignments gives a discreet and highly repetitive set of substructures not detectable by sequence similarity alone. In some cases different substructures represent all or different parts of well known folds indicative of the Russian doll effect--the continuity of protein fold space. In other cases they fall into different structure and functional classifications. It is too early to determine whether these newly classified substructures represent new insights into the evolution of a structural framework important to many proteins. What is apparent from on-going work is that these substructures have the potential to be useful probes in finding remote sequence homology and in structure prediction studies. The characteristics of the complete all-by-all comparison of the polypeptide chains present in the PDB and details of the filtering procedure by pair-wise structure alignment that led to the emergent substructure gallery are discussed. Substructure classification, alignments, and tools to analyze them are available at http://cl.sdsc.edu/ce.html. PMID- 10713985 TI - A proposed model for the PEX5-peroxisomal targeting signal-1 recognition complex. AB - The three-dimensional structure of a protein can greatly illuminate the relationship between its sequence and its function. However, in the absence of a set of experimentally derived coordinates, one often seeks a model of the protein of interest to guide future study. We describe the combined utilization of orthologous sequence information along with knowledge of the related structural fold to model the interaction between PEX5 and its ligand, the peroxisomal targeting signal-1 (PTS1). With this model, we are able to identify residues within PEX5 that appear to be important for peptide recognition, as well as explain some of the sequence requirements of the PTS1. Specifically, our model highlights four asparagine residues as important for ligand backbone atom recognition, which, along with previously observed examples, suggests this as a general mechanism for the binding of extended polypeptides. PMID- 10713987 TI - Water penetration and escape in proteins. AB - The kinetics of water penetration and escape in cytochrome c (cyt c) is studied by molecular dynamics (MD) simulations at various temperatures. Water molecules that penetrate the protein interior during the course of an MD simulation are identified by monitoring the number of water molecules in the first coordination shell (within 3.5 A) of each water molecule in the system. Water molecules in the interior of cyt c have 0-3 water molecules in their first hydration shell and this coordination number persists for extended periods of time. At T = 300 K we identify over 200 events in which water molecules penetrate the protein and reside inside for at least 5 picoseconds (ps) within a 1.5 nanoseconds (ns) time period. Twenty-seven (27) water molecules reside for at least 300 ps, 17 water molecules reside in the protein interior for times longer than 500 ps, and two interior water molecules do not escape; at T = 360 K one water molecule does not escape; at 430 K all water molecules exchange. Some of the internal water molecules show mean square displacements (MSD) of 1 A2 characteristic of structural waters. Others show MSD as large as 12 A2, suggesting that some of these water molecules occupy transient cavities and diffuse extensively within the protein. Motions of protein-bound water molecules are rotationally hindred, but show large librations. Analysis of the kinetics of water escape in terms of a survival time correlation function shows a power law behavior in time that can be interpreted in terms of a broad distribution of energy barriers, relative to kappa BT, for water exchange. At T = 300 K estimates of the roughness of the activation energy distribution is 4-10 kJ/mol (2-4 kappa BT). Activation enthalpies for water escape are 6-23 kJ/mol. The difference in activation entropies between fast exchanging (0.01 ns) and slow exchanging (0.1-1 ns) water molecules is -27 J/K/mol. Dunitz (Science 1997;264:670.) has estimated the maximum entropy loss of a water molecule due to binding to be 28 J/K/mol. Therefore, our results suggest that the entropy of interior water molecules is similar to entropy of bulk water. PMID- 10713988 TI - A novel energy-based stochastic method for positioning polar protons in protein structures from X-rays. AB - A novel automated method for the optimal placement of polar hydrogens in a protein structure is presented. The algorithm adds initially, to a protein data bank file of the protein, nonrotatable hydrogens such as peptide backbone hydrogens according to geometric considerations. Then, water protons and polar side chain protons of lysine, serine, threonine, tyrosine, aspartic acid, glutamic acid, and the C and N termini of a protein are added according to energy considerations. A unique stochastic approach has been developed to overcome a combinatorial explosion in the search for the lowest energy structure. First, the system is divided into ensembles. Each ensemble is treated separately: N conformations are sampled at random, their energies computed, whereas common components of high-energy combinations are gathered on one hand, and low-energy combinations on the other. Components that yield only high-energy conformations and do not contribute to any low energies are excluded. This is reiterated while the total amount of combinations is decreased along the iterative process. When the total number of combinations is lower than a user defined threshold, all remaining combinations are evaluated by exhaustive search. Energy evaluations use nonbonding energy expressions alone. The program was tested on five high resolution crystal structures: bovine pancreatic trypsin inhibitor (Brookhaven Protein Data Bank file 5PTI), RNase-A (5RSA), trypsin (1NTP), and carbon monoxymyoglobin (2MB5), for which neutron diffraction structures are available, as well as phosphate binding protein (1IXH) for which very high resolution X-ray crystallography was used. The low RMS values prove the efficiency of this algorithm as a tool for positioning protons in proteins. It may be used for other biological structures. PMID- 10713989 TI - Environment of tryptophan side chains in proteins. AB - Although relatively rare, the tryptophan residue (Trp), with its large hydrophobic surface, has a unique role in the folded structure and the binding site of many proteins, and its fluorescence properties make it very useful in studying the structures and dynamics of protein molecules in solution. An analysis has been made of its environment and the geometry of its interaction with neighbors using 719 Trp residues in 180 different protein structures. The distribution of the number of partners interacting with the Trp aromatic ring shows a peak at 6 (considering protein residues only) and 8 (including water and substrate molecules also). The means of the solvent-accessible surface areas of the ring show an exponential decrease with the increase in the number of partners; this relationship can be used to assess the efficiency of packing of residues around Trp. Various residues exhibit different propensities of binding the Trp side chain. The aromatic residues, Met and Pro have high values, whereas the smaller and polar-chain residues have weaker propensities. Most of the interactions are with residues far away in sequence, indicating the importance of Trp in stabilizing the tertiary structure. Of all the ring atoms NE1 shows the highest number of interactions, both along the edge (hydrogen bonding) as well as along the face. Various weak but specific interactions, engendering stability to the protein structure, have been identified. PMID- 10713990 TI - Structural alignment of ferredoxin and flavodoxin based on electrostatic potentials: implications for their interactions with photosystem I and ferredoxin NADP reductase. AB - The two proteins ferredoxin and flavodoxin can replace each other in the photosynthetic electron transfer chain of cyanobacteria and algae. However, structure, size, and composition of ferredoxin and flavodoxin are completely different. Ferredoxin is a small iron-sulfur protein (approximately 100 amino acids), whereas flavodoxin is a flavin-containing protein (approximately 170 amino acids). The crystal structure of both proteins from the cyanobacteria Anabeana PCC 7120 is known. We used these two protein structures to investigate the structural basis of their functional equivalence. We apply the Hodgkin index to quantify the similarity of their electrostatic potentials. The technique has been applied successfully in indirect drug design for the alignment of small molecule and bioisosterism elucidation. It requires no predefined atom-atom correspondences. As is known from experiments, electrostatic interactions are most important for the association of ferredoxin and flavodoxin with their reaction partners photosystem I and ferredoxin-NADP reductase. Therefore, use of electrostatic potentials for the structural alignment is well justified. Our extensive search of the alignment space reveals two alignments with a high degree of similarity in the electrostatic potential. In both alignments, ferredoxin overlaps completely with flavodoxin. The active sites of ferredoxin and flavodoxin rather than their centers of mass coincide in both alignments. This is in agreement with electron microscopy investigations on photosystem I cross linked to ferredoxin or flavodoxin. We identify residues that may have the same function in both proteins and relate our results to previous experimental data. PMID- 10713991 TI - When fold is not important: a common structural framework for adenine and AMP binding in 12 unrelated protein families. AB - ATP is a ligand common to many proteins, yet it is unclear whether common recognition patterns do exist among the many different folds that bind ATP. Previously, it was shown that cAMP-dependent protein kinase, D-Ala:D-Ala ligase and the alpha-subunit of the alpha 2 beta 2 ribonucleotide reductase do share extensive common structural elements for ATP recognition although their folds are different. Here, we have made a survey of structures that bind ATP and compared them with the key features seen in these three proteins. Our survey shows that 12 different fold types share a specific recognition pattern for the adenine moiety, and 8 of these folds have a common structural framework for recognition of the AMP moiety of the ligand. The common framework consists of a tripeptide segment plus three additional residues, which provides similar polar and hydrophobic interactions between the protein and mononucleotide. Consensus interactions are represented by four key hydrogen bonds present in each fold type. Two of these four hydrogen bonds, together with three aliphatic residues, form a specific recognition pattern for the adenine moiety in all 12 folds. These similarities point to a structural-functional requirement shared by these different mononucleotide-binding proteins that represent at this time 28% of the adenine mononucleotide complexes found in the Brookhaven Protein Data Bank. PMID- 10713992 TI - Molecular dynamics of mouse and Syrian hamster PrP: implications for activity. AB - Molecular dynamics computer simulations have been performed on Mouse (Mo) and Syrian Hamster (SHa) prion proteins. These proteins differ, primarily, in that the SHa form incorporates additional residues at the C-terminus and also includes a segment of the unstructured N-terminal region that is required for infectivity. The 1-ns simulations have been analyzed by using a combination of dynamical cross correlation maps, residue-residue contact plots, digital filtering, and residue based root-mean-square deviations. The results show that the extra residues present in the SHa form at the C- and N-termini produce changes in the stability of key regions of the protein. The loop region between strand S2 and helix B that contains part of the proposed discontinuous binding site for the chaperone, protein X, is found to be more stable in SHa than in the Mo protein; these results are consistent with the NMR data of James et al. (James et al. Proc Natl Acad Sci USA 1997;94:10086-10091). In addition, a degree of flexibility within the region between and including strand S1 and helix A is also shown in SHa, which is not present in the Mo form; the cross-correlation maps suggest that this is a consequence of the additional unstructured N-terminal region. Furthermore, the extra residues in the N-terminal region of SHa are found to form a beta bridge with the beta-sheet, within which critical point mutations associated with prion diseases lie. The implications of these results for the conformational interconversion pathway of the prion protein are discussed. PMID- 10713993 TI - Structural characterization of a methionine-rich, emulsifying protein from sunflower seed. AB - The 2 S seed storage protein, sunflower albumin 8, contains an unusually high proportion of hydrophobic residues including 16 methionines in a mature protein of 103 amino acids. A structural model, based on the known structure of a related protein, has been constructed as a four-helix bundle cross-linked by four disulphide bonds. This model structure is consistent with data from circular dichroism and nuclear magnetic resonance experiments. Analysis of the model's surface shows the presence of a large hydrophobic face that may be responsible for the highly stable emulsions this protein is known to form with oil/water mixtures. PMID- 10713994 TI - Disclosure, health, and negative emotions. PMID- 10713995 TI - Hypnosis and mind-body research. PMID- 10713997 TI - The placebo effect: if it's all in your head, does that mean you only think you feel better? PMID- 10713996 TI - Diet, exercise and natural killer cells. PMID- 10713998 TI - "Of course mental events affect physical events". PMID- 10713999 TI - "The vast majority of drug studies offer little if any evidence to support the physiologic effects of the placebo". PMID- 10714000 TI - What is so special about the placebo effect? PMID- 10714001 TI - "Is there really a placebo effect, professor"? PMID- 10714002 TI - Do placebo effects in analgesic drug studies demonstrate powerful mind-body interactions? PMID- 10714003 TI - Can a placebo thought make you blush? An evidence-based poem. PMID- 10714004 TI - A contribution to the debate. PMID- 10714005 TI - Tabulating the results. PMID- 10714006 TI - Toward a research agenda on placebo. AB - The placebo effect is about healing. The human healing process can be substantially influenced in actual medical practice by appropriate kinds of caring, communication, and patient empowerment. The creation of "meaning" and of "representations" by physicians and their patients can have dramatic effects on patients for good or ill. These effects are potentially as important in procedures such as surgery or chiropractic as they are in medicine generally. Placebo processes can substantially affect the overall response to health care and its cost, yet very little research specifically explores these phenomena. An explicit research agenda should be developed to investigate the influences of belief, context, and meaning in medicine. Such a research agenda would have significant beneficial scientific and policy consequences. In addition, it would enhance our understanding of those healing processes that underlie most if not all of medicine. PMID- 10714007 TI - Observations on the disintegration of the self: an experiential case for holism. PMID- 10714008 TI - Reflections for Anna. PMID- 10714009 TI - Holistic dying? PMID- 10714010 TI - Senses and the self. PMID- 10714011 TI - [Molecular targeting as a new strategy for cancer chemotherapy]. PMID- 10714012 TI - Inhibin-like immunoreactivity produced by the adrenal gland is circulating in vivo. AB - To clarify the contribution of the inhibin-like immunoreactivity (inhibin-LI) produced by adrenal glands to the total circulating levels of inhibin-LI, we measured serum inhibin-LI in normal and hypogonadal subjects under ACTH-loading or dexamethasone-loading condition. The mean basal concentration of inhibin-LI in the peripheral serum of the hypogonadal cases was 3.6 +/- 1.3 IU/ml (mean +/- SD, n = 5), which corresponded to 19.5 +/- 5.8% of that of normal controls matched for age and sex. The low levels of inhibin-LI in hypogonadal subjects (n = 7) rose significantly (3.6 +/- 1.1 vs 8.1 +/- 1.7 IU/ml, p < 0.001) after the administration of synthetic 1-24ACTH (40 units/day intramuscular injection) for 2 days, while the levels of serum inhibin-LI were not increased in two cases of Addison's disease with hypogonadism after the administration of ACTH. After the oral administration of a low dose of dexamethasone (1 mg) the serum inhibin-LI level in normal subjects (eight males and eight females) decreased significantly (male, 16.2 +/- 3.3 vs 14.5 +/- 4.1 IU/ml; female, 12.9 +/- 6.3 vs 10.8 +/- 5.6 IU/ml; p < 0.01 each) without significant change in the levels of serum gonadotropin (LH and FSH) and those of gonadal steroid (testosterone or estradiol). These results indicate that a small; but significant amount of inhibin-LI is secreted from the adrenal gland and circulating in vivo, and that the proportion of adrenal-derived inhibin-LI is much higher in patients with hypogonadism. PMID- 10714013 TI - Comparative study of the toxic effects of gallium arsenide, indium arsenide and arsenic trioxide following intratracheal instillations to the lung of Syrian golden hamsters. AB - Toxic effects of gallium arsenide (GaAs), indium arsenide (InAs) and arsenic trioxide (As2O3) were studied in male Syrian golden hamsters. GaAs (7.7 mg/kg) and As2O3 (1.3 mg/kg) particles were instilled intratracheally twice a week a total of 16 times, while InAs (7.7 mg/kg) was instilled a total of 14 times. As a control, hamsters were treated with the vehicle, phosphate buffer solution. During the instillation period, the cumulative body weight gain of the InAs-, but not the GaAs- or As2O3-treated hamsters was suppressed significantly, when compared with the control group. Slight to severe inflammatory responses were observed in the lung for all treatment groups. The most severe inflammatory change, characterized by an accumulation of neutrophils and macrophages, exudation, thickness of the pleura and fibrotic proliferation was found in the InAs-treated hamsters. Extensive alveolar or bronchiolar cell hyperplasia with or without keratinizing squamous cell metaplasia was observed in almost all the InAs treated hamsters. Furthermore, squamous cell metaplasia or squamous cell hyperplasia developed in some of the InAs-treated hamsters, but not in the GaAs- or As2O3-treated hamsters. Slight to mild lesions were found in the convoluted tubules of the kidney in both the GaAs and InAs groups. From the present study, the toxic potency of these particles was provisionally estimated to be in the following order: InAs > GaAs > As2O3, at the dosage level used in this study. Furthermore, there was evidence that InAs particles could induce pulmonary, renal or systemic toxicity, and as such, InAs particles may produce pulmonary precancerous change when instilled intratracheally into hamsters. PMID- 10714014 TI - Terminal warm blood cardioplegia improves the recovery of myocardial electrical activity. A retrospective and comparative study. AB - OBJECTIVE: The effect of terminal warm blood cardioplegia was analyzed in 191 patients undergoing either coronary artery bypass grafting (CABG) or prosthetic heart valve replacement between Jan. 1990 and Dec. 1995. METHODS: Patients were subdivided into 3 historical cohorts based on the method of myocardial protection: Group A (n = 106), multidose cold crystalloid glucose-potassium cardioplegia, alone; Group B (n = 37), cold crystalloid glucose-potassium cardioplegia plus terminal warm blood cardioplegia, Group C (n = 48), cardioplegia induction with cold crystalloid glucose-potassium cardioplegia, maintenance with multidose cold blood cardioplegia, and terminal warm blood cardioplegia. RESULTS: Of patients undergoing CABG, 5.6% of group A, 70.4% of group B, and 86.7% of group C spontaneously resumed sinus rhythm after aortic declamping, as did 9.1% of group A, 60.0% of group B, and 55.6% of group C of patients undergoing prosthetic heart valve replacement. The incidence of spontaneous recovery was significantly better in groups B and C than in group A (p < 0.05). Over 90% of patients without terminal warm blood cardioplegia developed ventricular fibrillation or tachycardia requiring electrical cardioversion (p < 0.05). Postoperatively, patients without terminal warm blood cardioplegia required temporary epicardial pacing more frequently than those with terminal warm blood cardioplegia (p < 0.05). In patients undergoing prosthetic heart valve replacement, groups B and C, the incidence of postoperative atrial fibrillation was significantly lower than in group A. CONCLUSION: Terminal warm blood cardioplegia thus promoted better postoperative electrophysiological cardiac recovery. PMID- 10714015 TI - Combined thoracic aortic or upper digestive tract resection for lung cancer and malignant mediastinal tumor. AB - OBJECTIVE: We studied possible indications and combined resection in patients with lung cancer and mediastinal tumors requiring combined thoracic aortic or upper digestive tract resection. METHODS: Ten patients with lung cancer and malignant mediastinal tumors (9 men and 1 woman aged 39 to 72 years; mean: 60.5) underwent combined aortic or upper digestive tract resection. RESULTS: Five--3 [corrected] with primary lung cancer, 1 with thymic cancer, and 1 with liposarcoma--, underwent combined aortic resection. In 2 each, lung cancer and malignant mediastinal tumor had infiltrated the thoracic aorta. The remaining case of lung cancer was complicated by aortic aneurysm in the distal arch. Cardiopulmonary bypass was conducted in 4, and selective cerebral perfusion in 2. Three patients are alive after 11, 22, and 61 months without disease recurrence. Those undergoing combined upper digestive tract resection all had lung cancer, with 4 having tumors infiltrating the esophagus or corpus ventriculi. The remaining patient had both lung and esophageal cancer. The patient treated with combined corpus ventriculi resection has survived 24 months and the patient treated with combined esophageal resection has survived 12 months without disease recurrence. The 1-year survival rate was 60%, 2-year 23%, and 3-year 23%. Prognosis was generally poor with the longest survival 13 months with N2 lung cancer. CONCLUSIONS: In combined resection due to malignant mediastinal tumor, T4N0-1 lung cancer, or diseases such as aortic aneurysm, prognosis can be expected to improve. Despite the often poor prognosis in T4N2 lung cancer, surgical intervention may be indicated to avoid complications due to tumor invasion and to lengthen survival and improve quality of life. PMID- 10714016 TI - Right ventricular volume unloading evaluated by tangential magnetocardiography. AB - OBJECTIVE: The evaluation of right ventricular volume overload in the presence of a right bundle branch block based solely on the results of an electrocardiogram is difficult. The purpose of this study was to purify acute right ventricular volume unloading from the tangential magnetocardiography. METHODS: We measured the tangential (x-y plane) magnetocardiogram and electrocardiogram simultaneously in nine patients with a secundum atrial septal defect. The magnetocardiograms were obtained before surgical closure and during the immediate postoperative period using the 32-channel superconducting quantum interference device system in a magnetically shielded room. RESULTS: The QRS duration on the surface electrocardiogram decreased significantly (p < 0.05) in the immediate postoperative period. The right ventricular depolarization time as measured by the magnetocardiogram was shortened from 40.3 +/- 6.1 to 25.3 +/- 7.4 milliseconds (p < 0.0005). The maximum peak amplitude during right ventricular depolarization decreased from 17.9 +/- 4.8 to 11.1 +/- 4.9 pT (p < 0.01). CONCLUSIONS: We conclude that acute volume unloading of the right ventricle was indicated quantitatively by shortening of the right ventricular depolarization time and a reduction in the amplitude of current vectors originating from the right ventricular depolarization on the tangential magnetocardiography. PMID- 10714017 TI - Nicorandil pretreatment and improved myocardial protection during cold blood cardioplegia. AB - OBJECTIVE: The present study was designed to assess whether pretreatment with nicorandil enhanced myocardial protection provided by cold (15 degrees C) high potassium (25 mmol/l) blood cardioplegia during open heart surgery. METHODS: Subjects were 40 patients with a variety of acquired heart diseases undergoing cardiac surgery involved cardiopulmonary bypass. They were randomly divided into two groups, 25 pretreated nicorandil (0.3 mg/kg) 30 minutes before aortic cross clamping, 15 not pretreated. After aortic cross clamping, the initial dose of cardioplegic solution (10 ml/kg) was administered through the ascending aorta and supplemental doses of cardioplegia (5 ml/kg) given each 30 minutes thereafter. Preoperative and postoperative cardiac troponin-T, myosin light chain 1 and cardiac enzymes were measured and hemodynamic data recorded. RESULTS: Postoperative serum creatine kinase and myosin light chain 1 were significantly lower in the nicorandil pretreatment group than in controls. Serum glutamic oxalacetic transaminase and troponin-T were lower and cardiac output was higher after surgery in the nicorandil group, although not statistically significant. CONCLUSION: This data suggests that pretreatment with nicorandil enhances the myocardial protection achieved by cold blood cardioplegia. PMID- 10714018 TI - Evaluation of postoperative cardiac function and long-term results in patients after aortic valve replacement for aortic valve disease with increased left ventricular mass. AB - OBJECTIVE: This clinical study was designed to evaluate the postoperative cardiac function in patients after aortic valve replacement for aortic valve disease with increased left ventricular mass. METHODS: Aortic valve replacement was performed in 117 patients using the St. Jude Medical valve. Their valve lesion was aortic regurgitation in 71, and aortic stenosis in 46. The mean value of the left ventricular mass index was 272 g/m2. The 117 patients were subdivided into 4 groups according to their preoperative left ventricular mass index-Group I (n = 35) with aortic regurgitation and a large left ventricular mass index (> or = 273 g/m2), Group II (n = 36) with aortic regurgitation and a small left ventricular mass index Group III (n = 19) with aortic stenosis and a large left ventricular mass index, and Group IV (n = 27) with aortic stenosis and a small left ventricular mass index. The cardiac function was evaluated by radionuclide ventriculography. RESULTS: In a comparative study of postoperative parameters among the 4 groups, the postoperative systolic and diastolic parameters of Group I patients were more significantly impaired compared with these parameters of the other 3 groups. The postoperative values the left ventricular mass index were significantly higher in Group I than in the other 3 groups. The 10-year survival rate was significantly lower in Group I than in the other 3 groups (30 +/- 22% in Group I). CONCLUSION: Aortic valve replacement is recommended for patients with eccentric hypertrophy in the adequate clinical phase of patients whose left ventricular mass index is less than 272 g/m2. PMID- 10714019 TI - Clinical experience of Argatroban for anticoagulation in cardiovascular surgery. AB - PURPOSE: We have reviewed our experience with Argatroban-a direct thrombin inhibitor for anticoagulation--in a variety of cardiovascular operations, and in extracorporeal circulation, as a substitute for heparin. SUBJECTS AND METHODS: 60 patients receiving anticoagulation with Argatroban were classified into the following four groups. Group 1; 20 patients with anticoagulation therapy after cardiac surgery. Group 2; 8 patients with extracorporeal circulation for continuous hemofiltration for either pre- or post-operative control of acute renal failure. Argatroban was used alone or in combination with nafamostat mesilate. Group 3, one patient with replacement of the descending aorta with left heart assist and 15 patients with percutaneous cardiopulmonary support. And Group 4, 16 patients undergoing vascular surgery including the abdominal aorta. The target activated clotting time was individually set for each group. In Group 1, the coagulofibrinolytic activity and platelet function were measured precisely. Bleeding and complications were examined in all groups. RESULTS: Group 1; the targeted activated clotting time of 150-180 seconds was achieved by a dosage of 0.4-0.8 microgram/kg/min Argatroban. Group 2; the activated clotting time of 150 180 seconds was achieved by 0.05-1.6 micrograms/kg/min (concomitance), or by 0.02 2.5 micrograms/kg/min (alone). Group 3; the activated clotting time of 180-200 seconds by 0.05-3.86 micrograms/kg/min. And Group 4; the activated clotting time of around 150 seconds by 2.0 micrograms/kg/min with initial bolus infusion of 0.1 mg/kg. Argatroban did not promote post-surgery bleeding and had no unfavorable effect on coagulo-fibrinolysis or on platelet activity. CONCLUSION: Argatroban may be useful as an anticoagulant in the field of cardiovascular surgery as a substitute for heparin, without causing any post-surgery bleeding complication, or influencing the fibrinolytic activities or platelet functions. PMID- 10714020 TI - Risk factors for posttransfusion graft versus host disease, mediastinitis, and late cardiac tamponade in heart surgery. Survey of 119 Japanese institutions. AB - OBJECTIVE: Correlations and risk factors remain to be unclarified for post-heart surgery posttransfusion graft-versus-host disease, mediastinitis, and late cardiac tamponade caused by deteriorated host-defense mechanisms due to cardiopulmonary bypass both with and without steroid usage. METHODS: We sent questionnaires to 298 Japanese cardiovascular institutions asking for institution profiles, including infection control, steroid use in cardiopulmonary bypass, and prevalence of mediastinitis, late cardiac tamponade, and posttransfusion graft versus-host disease during 1994. The overall prevalence of posttransfusion graft versus-host disease since the start of service (from establishment of institution to date) was also requested. RESULTS: The number of pump cases at the 119 institutions responding (40%) were 91.6 +/- 67.9 cases/institution (total = 10,904). The prevalence of mediastinitis was 1.2 +/- 1.8 and that of late cardiac tamponade 1.0 +/- 1.8%. Posttransfusion graft-versus-host disease occurred in 1 of 10,904 patients (0.01%) during 1994 at an institution where steroids and nonirradiated blood were used in surgery. The simple institutional mean prevalence of posttransfusion graft-versus-host disease since establishing institutions was 0.08 +/- 0.13%. Of the 119 institutions surveyed, 86 used steroids in all pump cases (72%); 11 institutions used steroids in a limited number of cases (9%). The institutional mean of methylprednisolone-converted steroid dose was 21.5 +/- 16.4 mg/kg (n = 119). In multivariate regression analysis, operation time (p = 0.005) for mediastinitis, steroid usage (all, limited, or no cases) (p = 0.01) and % aneurysm (p = 0.05) for late cardiac tamponade, and steroid dosage (p = 0.002) for posttransfusion graft-versus-host disease were identified as significant risk factors. CONCLUSION: Our results suggest that massive steroid administration for cardiopulmonary bypass may increase the risk of posttransfusion graft-versus-host disease and late cardiac tamponade, but not mediastinitis. PMID- 10714021 TI - Surgical management of pulmonary aspergilloma. Role of single-stage cavernostomy with muscle transposition. AB - OBJECTIVE: We reviewed the outcome of the patients with aspergilloma who were treated surgically. METHODS: Between July 1991 and October 1996, 11 patients with pulmonary aspergilloma underwent surgery. One underwent sequential bilateral tboracotomy and two underwent re-operation. The total number of operations was 14. Surgical procedures consisted of 5 cavernostomies with muscle transposition, 3 cavernostomies with muscle transposition and thoracoplasty, 1 lobectomy 1 pneumonectomy, 1 segmentectomy and 3 partial resections. RESULTS: Morbidity and mortality rates were 28.6% and 7.1%, respectively Two patients who underwent cavernostomy and muscle transposition experienced a relapse of aspergilloma 19 and 29 months after the operation, respectively, but both successfully underwent re-operation, including cavernostomy. Both are free of symptoms 28 and 30 months after re-operation, respectively. All survivors except for one who died of multiple organ failure remain free of symptoms 14 to 60 months after the most recent operation. CONCLUSION: Our experience was not a controlled trial and two relapsed cases had undergone cavernostomy, our series may suggest that single stage cavernostomy with muscle transposition is a viable surgical option for patients with pulmonary aspergilloma. PMID- 10714022 TI - Evaluation of motor- and sensory-evoked potentials for spinal cord monitoring during thoracoabdominal aortic aneurysm surgery. AB - OBJECTIVE: To assess the utility of spinal cord monitorings for prediction of spinal cord ischemia, we investigated the role of both motor evoked potentials and sensory evoked potentials during thoracoabdominal aortic aneurysm surgeries. METHODS: We monitored two kinds of sensory evoked potentials; descending evoked spinal cord potentials from the lumbar enlargement after cervical spinal cord stimulation and segmental evoked spinal cord potentials at the lumbar enlargement elicited by peroneal nerve stimulation, and motor evoked potentials from the lumbar enlargement elicited by direct subcranial stimulation in 9 thoracoabdomonal aortic aneurysm surgeries. RESULTS: Postoperative paraplegia occurred in one case in which the patients died during the perioperative period. One case showed transient paraparesis, but recovered following rehabilitatation. These cases showed a decrease in the amplitude of descending evoked spinal cord potentials and motor evoked potentials. CONCLUSION: The recovery of the amplitude of the motor evoked potentials and the descending evoked spinal cord potentials after declamping correlated with the neurologic outcome. PMID- 10714023 TI - Mediastinal growing teratoma syndrome. AB - A 27-year-old man had undergone orchiectomy and chemotherapy for testicular cancer. Despite normalization of raised tumor marker levels after postoperative chemotherapy, computed tomographic scanning demonstrated multiple swellings of the para-aortic lymph nodes with extension from beneath the aortic arch to the bifurcation of the descending aorta. Open biopsies of the para-aortic lymph nodes disclosed mature teraroma without malignant cells. The patient presented the typical features of mediastinal and retroperitoneal growing teratoma syndrome. A two stage resection of the tumors was performed via laparotomy and left thoracotomy. Histological examination of the resected specimens revealed a mature teratoma component without malignant cells. Upon follow-up sixteen months later, the patient was well and without recurrence. PMID- 10714024 TI - Stanford type A acute dissection developing acute myocardial infarction. AB - A 75-year-old female, exhibiting epigastric pain and vomiting, underwent treatment for acute gastritis. She also experienced incontinence of urine and chest pain. A diagnosis of acute myocardial infarction was made upon examination of electrocardiographic findings and the patient was transferred to our hospital. Diffuse infarction of the left ventricle and acute aortic dissection (Stanford type A) were diagnosed by electrocardiographic and echo-cardiography. An emergency operation was performed. After induction of anesthesia, elevation of pulmonary artery pressure and fall of pulse pressure were observed, indicating acute cardiac tamponade. Transesophageal ultrasonography disclosed the entry of dissection in the descending aorta. Dissection of the aorta extended proximally up to the annulus of the aortic valve and the right and left coronary arteries were compressed by its aneurysm. As aortic insufficiency was mild, only reconstruction of the ascending aorta was carried out. The patient was discharged in fair condition one month after operation under use of postoperative long-term administration of catecholamines. PMID- 10714025 TI - Aorto-bronchial fistula after implantation of a self-expanding bronchial stent in a patient with aortic dissection. AB - We report a case of aorto-bronchial fistula after implantation of a self expanding stent into the left main bronchus compressed by a dissected descending aorta. A 66-year-old female, who underwent Stanford type-B aortic dissection two years previously, was admitted to our hospital for the treatment of a newly developed false lumen that originated from the ascending aorta and extended to the aortic bifurcation. She was unable to be weaned from the respirator after the graft replacement of the ascending aorta. Fiberoptic bronchoscopic examination revealed complete obstruction of the left main bronchus by extrinsic compression. A self-expanding nitinol stent was implanted in the left main bronchus five days after the operation. Her respiratory condition improved remarkably, allowing her to be successfully weaned from the respirator. Her clinical course was uneventful until she suddenly died from massive hemoptysis 20 days after stent implantation. A communication of 5 mm in diameter between the dissected descending aorta and the left main bronchus was seen at autopsy. Permanent application of a self expanding nitinol stent to relieve extrinsic compression of a left main bronchus by a dissected descending aorta is not recommended because pressure necrosis might lead to fatal aorto-bronchial fistula. PMID- 10714026 TI - Off-pump coronary artery bypass grafting via partial sternotomy. An alternative to left anterior small thoracotomy. AB - Four patients, who were considered to be inappropriate candidates for left anterior small thoracotomy, underwent off-pump coronary artery bypass grafting via partial sternotomy. Under a median skin incision over the lower half of the sternum, the sternum below the second rib was cut in an "inverted L" (or "C") shape. Without cardiopulmonary bypass, the left internal thoracic artery was anastomosed to the left anterior descending artery in all patients, and a saphenous vein graft was anastomosed to the right coronary artery in one of them. Partial sternotomy has some advantages as an alternative to left anterior small thoracotomy, in that it enables multiple-bypass grafting without cardiopulmonary bypass and conversion to cardiopulmonary bypass, should it be come necessary, would be relatively uncomplicated. PMID- 10714027 TI - Acute heart failure due to local dehiscence of aortic wall at aortic valvular commissure. AB - Spontaneous dehiscence of the aortic wall at the aortic commissure is not recognized as one of the usual pathological causes of aortic regurgitation. We describe the case of a 56-year-old man with hypertension, who experienced acutely progressive congestive heart failure due to massive aortic regurgitation. Local layer dehiscence around the commissure was noted with partial detachment of the commissure resulting in the loss of commissural support with secondary rupture of a non-coronary cusp, which led to massive aortic regurgitation. PMID- 10714028 TI - Translocation of aortic valve for calcific aortic stenosis. AB - A 56-year-old man underwent surgery for treatment of severe calcific aortic stenosis. Because it was found after excision of the aortic valve that calcification of the annulus was too extensive for the placement of sutures, translocation of the aortic valve was performed. The results were satisfactory and indicate that translocation is a useful alternative in cases of severe calcification of the aortic valve which cannot be treated by ordinary valve replacement. PMID- 10714029 TI - Innominate artery injury by blunt trauma. PMID- 10714030 TI - Pulmonary trunk aneurysm. PMID- 10714031 TI - Children and youth in out-of-home placements: nursing care opportunities for pediatric nurses. PMID- 10714032 TI - Joining the efforts to prevent and curb youth violence. PMID- 10714033 TI - Intervention studies involving parents of hospitalized young children: an analysis of the past and future recommendations. AB - Hospitalization imposes multiple stressors on young children and their parents that place them at risk for both short-term and long-term negative outcomes. As a result, numerous researchers have developed and tested interventions to enhance coping outcomes in this population. This article describes major sources of stress for hospitalized young children and their parents, outcomes of hospitalization, and major factors influencing adjustment. In addition, a critique of prior intervention studies and recommendations for future research are highlighted. PMID- 10714034 TI - School-based research: problems of access and consent. AB - Public schools are enticing yet intimidating places to conduct research on children and adolescents. A public school provides a large potential subject pool; however, obtaining access to that subject pool can seem impossible with all the layers of permissions that must be obtained. If the study is federally funded, additional regulations apply. This article presents practical and proven approaches to obtaining access to and approval from schools or school systems, as well as parental consent, to conduct school-based research. PMID- 10714035 TI - Children's voices: can we hear them? AB - This article addresses an important but often neglected notion in the care of children--the notion of voice. Recognizing that a crucial role for pediatric nurses is that of advocate for the child, this article poses the questions of how children's voices can be heard and how nurses know whose voice they represent when they act in an advocacy capacity. Drawing on contributions from psychology, sociology, and feminist studies, the analysis narrows our focus to the special challenge created for pediatric nurses when they recognize the importance of voice in caring for children, and examines the complexities inherent in attending to voice in pediatric nursing practice. PMID- 10714036 TI - Sociocultural influences and self-care practices of middle adolescents. AB - The purpose of this study was to describe general self-care practices of middle adolescents. In addition, the relation between general self-care practices and specific sociocultural characteristics including socioeconomics and church attendance were explored. Orem's self-care theory and developmental theory provided the framework for the investigation. Findings from the sample of 15- and 16-year-old adolescents (n = 425) showed that they are engaging in self-care practices. The influence of sociocultural characteristics on self-care practices was supported. Implications from the study include the need to continue research endeavors that describe, explain, and predict health behavior in child and adolescent populations. Practicing nurses in diverse health care settings should consider the results of this study when working with adolescents and their families from diverse sociocultural backgrounds. Results from this investigation should be incorporated into the planning of health education programs for the adolescent population. PMID- 10714037 TI - Ethical voices of pediatric mental health nurses. AB - Ethical issues in pediatric mental health care have undergone little theoretical consideration and empirical study. In this exploratory ethnographic study, 20 Pediatric Mental Health Registered Nurses (PMHRNs) describe the ethical issues they believe arise from the care they deliver to children in school-age and adolescent age groups. Three major themes emerge from the interviews. These themes, the PMHRNs' relational roles, their role as advocate facilitator, and their view of the milieu as an extension of the family, are analyzed for ethical content using several ethical theories. These ethical theories are evaluated for adequacy, and an argument for the use of relational ethical theories in examining pediatric mental health ethical issues, as well as general pediatric nursing practice, is presented. PMID- 10714038 TI - Healthy People 2000/2010: health appraisal of the nation and future objectives. PMID- 10714039 TI - Ethics in pediatric nursing: an international perspective. AB - The goal of this survey was to compare similarities and differences between two groups of pediatric nurses, a group from developing countries and a group from Israel, in confronting ethical issues and in the use of resources for support. Findings indicate similarities between the two groups in a majority of ethical situations that concern them in their work despite differences in cultural backgrounds. PMID- 10714040 TI - Are stimulants overprescribed? PMID- 10714041 TI - Are stimulants overprescribed? PMID- 10714042 TI - Behavioral inhibition and developmental risk: response to commentary. PMID- 10714043 TI - Smoking among Spanish adolescents. PMID- 10714044 TI - "Faced with guilt": a suicide risk education tool. PMID- 10714045 TI - Autism and adenylosuccinase deficiency. PMID- 10714046 TI - Imipramine plus cognitive-behavioral therapy in the treatment of school refusal. AB - OBJECTIVE: To investigate the efficacy of 8 weeks of imipramine versus placebo in combination with cognitive-behavioral therapy (CBT) for the treatment of school refusing adolescents with comorbid anxiety and major depressive disorders. METHOD: This was a randomized, double-blind trial with 63 subjects entering the study and 47 completing. Outcome measures were weekly school attendance rates based on percentage of hours attended and anxiety and depression rating scales. RESULTS: Over the course of treatment, school attendance improved significantly for the imipramine group (z = 4.36, p < .001) but not for the placebo group (z = 1.26, not significant). School attendance of the imipramine group improved at a significantly faster rate than did that of the placebo group (z = 2.39, p = .017). Over the 8 weeks of treatment, there was a significant difference between groups on attendance after controlling for baseline attendance; mean attendance rate in the final week was 70.1% +/- 30.6% for the imipramine group and 27.6% +/- 36.1% for the placebo group (p < .001). Defining remission as 75% school attendance, 54.2% of the imipramine group met this criterion after treatment compared with only 16.7% from the placebo group (p = .007). Anxiety and depression rating scales decreased significantly across treatment for both groups, with depression on the Children's Depression Rating Scale-Revised decreasing at a significantly faster rate in the imipramine group compared with the placebo group (z = 2.08, p = .037). CONCLUSIONS: Imipramine plus CBT is significantly more efficacious than placebo plus CBT in improving school attendance and decreasing symptoms of depression in school-refusing adolescents with comorbid anxiety and depression. PMID- 10714047 TI - Imipramine compliance in adolescents. AB - OBJECTIVE: To investigate side effects, medication compliance, and assumption of medication assignment in adolescents taking imipramine versus placebo in a clinical trial. METHOD: Sixty-three anxious-depressed adolescents in an 8-week double-blind study of imipramine versus placebo, each in combination with cognitive-behavioral therapy for school refusal, were evaluated. Measures of side effects, global improvement, family functioning, medication compliance based on pill counts, and guesses of drug assignment (imipramine versus placebo) were analyzed. RESULTS: Mean side effects ratings were significantly higher for the imipramine group compared with the placebo group (p = .001). Side effects were not associated with noncompliance or with dropping out. Oppositional defiant disorder (ODD) in the adolescents was significantly associated with medication noncompliance (p = .036). On the Family Adaptability and Cohesion Evaluation Scale II (FACES II), low family adaptability (i.e., rigidity), low family cohesion (i.e., disengagement), and extreme family type were significantly associated with greater noncompliance with medications. Accuracy rates for guessing medication assignment (imipramine versus placebo) were 66% for subjects, 62.5% for mothers, and 79.5% for the psychiatrist. Logistic regression demonstrated that side effects (p = .005) and global improvement scores (p = .06) predicted the psychiatrist's guesses of drug assignment. CONCLUSIONS: Side effects were not associated with noncompliance. Nonadherence with taking medications was associated with ODD in the adolescents and problematic family functioning on FACES II. The psychiatrist, who was blind to treatment condition, guessed the subjects' medication assignments with high accuracy. Thus, because of expectancy bias, the data support the use of blind independent evaluators for rating changes in medication trials. PMID- 10714048 TI - Ziprasidone treatment of children and adolescents with Tourette's syndrome: a pilot study. AB - OBJECTIVE: To evaluate the efficacy and tolerability of ziprasidone in children and adolescents with Tourette's syndrome and chronic tic disorders. METHOD: Twenty-eight patients aged 7 to 17 years were randomly assigned to ziprasidone or placebo for 56 days. Ziprasidone was initiated at a dose of 5 mg/day and flexibly titrated to a maximum of 40 mg/day. RESULTS: Ziprasidone was significantly more effective than placebo in reducing the Global Severity (p = .016) and Total Tic (p = .008) scores on the Yale Global Tic Severity Scale. Compared with placebo, ziprasidone significantly reduced tic frequencies as determined by blind videotape tic counts (p = .039). The mean (+/- SD) daily dose of ziprasidone during the last 4 weeks of the trial was 28.2 +/- 9.6 mg. Mild transient somnolence was the most common adverse event. No clinically significant effects were observed on specific ratings of extrapyramidal symptoms, akathisia, or tardive dyskinesia. CONCLUSIONS: In this limited sample, ziprasidone (5-40 mg/day) appears to be effective and well tolerated in the treatment of Tourette's syndrome. Ziprasidone may be associated with a lower risk of extrapyramidal side effects in children. However, additional studies are necessary to evaluate more fully its safety and efficacy in children with tic disorders. PMID- 10714049 TI - Validity of DSM-IV subtypes of attention-deficit/hyperactivity disorder: a family study perspective. AB - OBJECTIVE: To test the hypothesis that the clinical severity of subtypes paralleled a gradient of familial severity. METHOD: One hundred forty children with attention-deficit/hyperactivity disorder (ADHD) and 120 normal control children and their biological relatives were studied: Because these data had been collected prior to the publication of DSM-IV, DSM-III-R symptoms were used to approximate DSM-IV subtypes using a method the authors had validated in prior work. RESULTS: The first prediction from the hypothesis was true: rates of ADHD among relatives of each subtype group were greater than rates among relatives of controls. But the second prediction did not hold: rates of ADHD were not significantly higher among relatives of combined-typed probands compared with relatives of other probands. The "gradient model" also predicted that subtypes would not "breed true" (i.e., the subtype of the relative would not be the same as that of the proband). The prediction of nonspecificity was refuted for the inattentive and combined subtypes, but hyperactive-impulsive ADHD was found almost exclusively among relatives of hyperactive-impulsive probands. CONCLUSIONS: Although the results are limited by some small subsamples along with the use of a DSM-III-R-ascertained sample, they provide little evidence for the idea that DSM-IV subtypes of ADHD correspond to familially distinct conditions. They also do not confirm the idea that the subtypes fall along a gradient of familial severity. Instead, they suggest that symptom differences among subtypes are due to nonfamilial, environmental causes. PMID- 10714050 TI - Parent-teacher concordance for DSM-IV attention-deficit/hyperactivity disorder in a clinic-referred sample. AB - OBJECTIVE: To examine concordance between parent and teacher reports of DSM-IV attention-deficit/hyperactivity disorder (ADHD) and its symptoms. METHOD: Parents and teachers of 74 clinically referred children were interviewed using the ADHD module of the Diagnostic Interview Schedule for Children. Parent-teacher agreement for the diagnosis of ADHD and its subtypes, as defined in DSM-IV, as well as parent-teacher concordance of in-school ADHD symptoms, was examined. RESULTS: Agreement between parents and teachers was found to be relatively poor, with virtually no agreement for individual ADHD subtypes. Diagnoses based on either parent or teacher report frequently yielded a diagnosis of either inattentive or hyperactive-impulsive subtype of ADHD. However, when cross informant data were used to form diagnoses, these subtypes became relatively rare, with most cases meeting criteria for ADHD combined type. In addition, parent reports of in-school behavior were more highly correlated with their own reports of their child's behavior at home than with teacher reports of their child's behavior in school. CONCLUSIONS: These data suggest that the diagnosis of ADHD inattentive or hyperactive-impulsive subtype based on data from a single informant may be of questionable validity, and they point to the importance of using multiple informants when diagnosing this disorder in clinically referred samples. PMID- 10714051 TI - Impact of maternal depression on ratings of comorbid depression in adolescents with attention-deficit/hyperactivity disorder. AB - OBJECTIVE: To assess the degree to which indirect maternal reports of comorbid major depression (MD) in adolescents with and without attention deficit/hyperactivity disorder (ADHD) were influenced by the mother's personal history of MD. METHOD: Bivariate regression was used to model the impact of maternal depression on the direct and indirect report of MD in ADHD (n = 150) and non-ADHD (n = 123) subjects. The dependent variable (i.e., risk for MD) was modeled as a function of the main effect of ADHD, the main effect of reporter, their interaction, and higher-order interactions with maternal depression. RESULTS: There was a significant interaction between maternal depression and the effect of reporter exclusively in non-ADHD control subjects. ADHD continued to be a significant risk factor for MD independent of maternal reporting or maternal depression. CONCLUSIONS: The potential distortion of indirect interviews by depressed mothers may be stronger in community than in clinical settings and does not account for the increased risk for MD in referred adolescents with ADHD. PMID- 10714052 TI - Outcome and follow-up study of an adolescent psychiatric day treatment school program. AB - OBJECTIVES: To evaluate outcome and follow-up of 55 subjects who attended a classroom-based, adolescent psychiatric day treatment unit school program (ADTU) situated in a community high school by examining pre-post changes in emotional, behavioral, family, and academic functioning and to identify preadmission and family variables associated with outcome. METHOD: Student-patients were assessed on clinical/academic variables at admission, discharge, and follow-up, using standardized assessment measures. Patient satisfaction was also evaluated. RESULTS: Significant improvements in emotional-behavioral and academic functioning were found at discharge and follow-up. Hierarchical regression analysis revealed that parent and clinician ratings at admission, total number of separations from family greater than 3 months, parental history of mental illness, and patient history of treatment for emotional problems were significantly associated with outcome. Satisfaction questionnaires revealed a high degree of satisfaction with the program. CONCLUSIONS: The ADTU is a successful treatment program for adolescents with chronic, severe emotional and behavioral disorders, despite limitations of the study. PMID- 10714053 TI - Factors associated with early dropout from adolescent psychiatric outpatient treatment. AB - OBJECTIVE: To examine background factors, psychopathology, and psychosocial impairment among adolescents complying with or dropping out early from outpatient psychiatric treatment. METHOD: Family background, psychiatric history, and other data were collected prospectively on 143 male and 154 female outpatients aged 12 to 22 years. DSM-II-R psychiatric diagnoses were assessed at the end of treatment. RESULTS: Fifty-three adolescents (17.8%) attended 1 or 2 treatment appointments, and 33 of them (11.1% of 297) then dropped out; 50.5% of the total attended 3 to 13, and 31.6% attended 14 or more appointments. Low parental socioeconomic status was more common among the early dropouts than the other patient groups (88%, 69%, 63%, respectively). The early dropouts had had more problems with the law than the adolescents attending 14 or more appointments (18%, 6%), but less suicidal behavior (24%, 56%, respectively). Among the early dropouts, mood disorders were less common (21%, 49%), especially major depression (0%, 20%), and substance abuse was more common (9%, 0%) than among patients attending 14 or more appointments. CONCLUSIONS: Low parental socioeconomic status, not having mood disorder, not having psychotropic medication, and having substance abuse were associated with early dropout of adolescents from outpatient psychiatric treatment. PMID- 10714054 TI - Psychosis in a pediatric mood and anxiety disorders clinic: phenomenology and correlates. AB - OBJECTIVES: To examine the demographics and phenomenology of psychosis in a sample of children and adolescents referred to a mood and anxiety disorders clinic. METHOD: Patients (N = 2,031) were assessed with the Schedule for Affective Disorders and Schizophrenia for School-Age Children-Present Episode version and classified as definite, probable, or nonpsychotic. Clinical and demographic characteristics of the groups were compared,and symptoms of psychosis were analyzed using factor analysis. RESULTS: Definite psychotic symptoms were seen in approximately 90 (4.5%) patients: 80% of these reported hallucinations (mainly auditory), 22% delusions, and 3.3% thought disorder. Of the patients with definite psychotic symptoms, 24% had bipolar disorder, 41% had major depression, 21% had subsyndromal depression, and 14% had schizophrenia spectrum disorders (schizophrenia and schizoaffective disorders). Factor analysis of the definite psychotic symptoms yielded 4 factors: hallucinations, thought disorder, delusions, and manic thought disorder. Psychotic patients had a higher frequency of comorbid disorders and suicidal ideation than nonpsychotic patients. CONCLUSIONS: Outpatient youngsters with mood disorders frequently present with psychotic symptoms, in particular auditory hallucinations. These patients commonly have comorbid psychiatric disorders and suicidal ideation. PMID- 10714055 TI - Subgroups of children with autism by cluster analysis: a longitudinal examination. AB - OBJECTIVES: A hierarchical cluster analysis was conducted using a sample of 138 school-age children with autism. The objective was to examine (1) the characteristics of resulting subgroups, (2) the relationship of these subgroups to subgroups of the same children determined at preschool age, and (3) preschool variables that best predicted school-age functioning. METHOD: Ninety-five cases were analyzed. RESULTS: Findings support the presence of 2 subgroups marked by different levels of social, language, and nonverbal ability, with the higher group showing essentially normal cognitive and behavioral scores. The relationship of high- and low-functioning subgroup membership to levels of functioning at preschool age was highly significant. CONCLUSIONS: School-age functioning was strongly predicted by preschool cognitive functioning but was not strongly predicted by preschool social abnormality or severity of autistic symptoms. The differential outcome of the 2 groups shows that high IQ is necessary but not sufficient for optimal outcome in the presence of severe language impairment. PMID- 10714056 TI - Trauma and dissociation in delinquent adolescents. AB - OBJECTIVES: To assess history of trauma and dissociation in a group of juvenile delinquents and to assess how adolescents would respond to a structured interview for dissociative symptoms. METHOD: Sixty-four adolescents in juvenile probation hall participated in 2 investigational sessions in 1996-1997. For session 1 they answered the Childhood Trauma Questionnaire (CTQ), the Response Evaluation Measure for Youth-71 (REMY-71), and the Weinberger Adjustment Inventory. For session 2 they were given the Childhood Trauma Interview (CTI) and the Structured Clinical Interview for DSM-IV Dissociative Disorders (SCID-D). RESULTS: In this sample 28.3% met criteria for a dissociative disorder and 96.8% endorsed a history of traumatic events. There were significant positive correlations between CTI and CTQ trauma scores and SCID-D and REMY-71 dissociative symptoms. All dissociative symptoms were endorsed, but depersonalization was the most common experience. There was a lack of congruence between the different methods of assessing dissociation. CONCLUSIONS: This study provides support for an early link between history of trauma and dissociation. Adolescents were able to answer questions from a structured interview assessing dissociation. PMID- 10714057 TI - Cultural identification and attempted suicide in Native Hawaiian adolescents. AB - OBJECTIVES: To determine rates of lifetime suicide attempts in a community sample of Native Hawaiian adolescents and determine the contribution of Hawaiian cultural affiliation, socioeconomic status, and psychiatric symptoms as risk factors for suicide. METHOD: High school students were surveyed in the state of Hawaii for lifetime suicide attempts, Hawaiian cultural affiliation, socioeconomic status, and symptoms of depression, substance abuse, aggression, and anxiety. Multiple logistic regressions were used on 3,094 subjects to develop prediction models for lifetime suicide attempts. RESULTS: Native Hawaiian adolescents had significantly higher rates of suicide attempts (12.9%) than other adolescents in Hawaii (9.6%). Hawaiian cultural affiliation rather than ethnicity was uniquely predictive of suicide attempts. Logistic regression indicated that depression, substance abuse, grade level, Hawaiian cultural affiliation, and main wage earner's education best predicted suicide attempts in Native Hawaiian adolescents, while depression, substance abuse, and aggression predicted suicide attempts in non-Hawaiians. CONCLUSIONS: Native Hawaiian adolescents have higher rates of attempted suicide than non-Hawaiian adolescents. Strong Hawaiian cultural affiliation rather than ethnicity is a risk factor for attempted suicide. PMID- 10714058 TI - Psychopathology, adversity, and service utilization of young refugees. AB - OBJECTIVE: To investigate the psychopathology, social impairment, adversities, and service utilization of refugee families and their children seeking help at a child and adolescent psychiatry clinic in London. METHOD: A retrospective case control study of 30 refugee children and families individually matched with nonrefugee immigrant families and white British families. Case note review was carried out to obtain data on diagnosis, social adjustment, past adversity, exposure to violence, current socioeconomic circumstances, and use of the child and adolescent psychiatric service. RESULTS: Refugee children tended to have disorders with a psychosocial etiology rather than neurobiological disorders. Refugees had similar levels of social impairment compared with the other groups. Refugees were much more isolated and disadvantaged and had different referral pathways but were not more likely to drop out of treatment prematurely. CONCLUSIONS: Refugee children and families had been exposed to high levels of adversity. The ability of community agencies to refer families who could use treatment has significant resource implications. PMID- 10714059 TI - Detection and importance of laxative use in adolescents with anorexia nervosa. AB - OBJECTIVE: The frequency of laxative use in adolescents with anorexia nervosa is poorly described. This study of adolescents with anorexia nervosa examined self report and biochemical screening methods for the detection of laxative use, the pattern of laxative use in this population over time, and the associated medical complications and psychopathology. METHOD: Forty-three consecutive patients with anorexia nervosa were studied. Initial assessment encompassed psychiatric history, medical examination, and administration of the Eating Disorders Examination, Child Behavior Checklist, and Youth Self-Report. Biochemical investigations, including random urinary laxative screening, were performed at assessment and follow-up. RESULTS: The frequency of laxative use from self-report alone was 12%; combined with urine screening it was 19%. The frequency of laxative use increased to 32% with prospective follow-up. Medical complications were associated with laxative use at follow-up. Laxative use was associated with longer duration of disease and with higher scores on the Eating Disorders Examination subscale Eating Concern. CONCLUSIONS: Laxative use is common among adolescents with anorexia nervosa, and the risk of associated medical complications increases over time. Biochemical screening will improve detection of laxative use. Longer duration of illness and greater Eating Concern scores are associated with increased risk of laxative use, and monitoring patients at increased risk is important. PMID- 10714060 TI - Genetics of childhood disorders: XII. Genomic imprinting: breaking the rules. PMID- 10714061 TI - [Bone scintigraphy in metastatic bone disease]. AB - Bone scintigraphy has been widely used in the evaluation of metastatic bone disease. It can provide easy performance of whole body evaluation and high sensitivity for the detection of lesions. However, the indication for bone scintigraphy is revised from views of the less specificity and relatively higher cost than newly introduced diagnostic tools such as CT and MRI in Japan. In order to maintain the present state as a screening test for the evaluation of bone metastasis, more appropriate image acquisition and newly developed radiopharmaceuticals which show the specific accumulation to the metastatic bony lesions will be considered in future. PMID- 10714062 TI - [Measurement of myocardial blood flow increase rate at exercise with 99mTc tetrofosmin radionuclide angiography]. AB - We developed new method to calculate myocardial blood flow increase rate at exercise (MBF-IR) with 99mTc-tetrofosmin (TF) radionuclide (RN) angiography and myocardial perfusion SPECT and assessed its feasibility using clinical data. METHOD: Fifteen patients who were suspected to have coronary artery disease underwent TF RN angiography and SPECT at exercise and at rest. Seven patients had coronary stenosis and eight patients had no significant coronary stenosis in coronary angiography. MBF-IRs were calculated by the following equation: [formula: see text] where Cr = regional myocardial count at rest, Ce = regional myocardial count at exercise [formula: see text] = the area under ventricular time activity curve at rest and [formula: see text] = the area under ventricular time activity curve at exercise. RESULT: Rate pressure product (RPP) was similar in patients with and without coronary stenosis (24,509 +/- 6701.9 vs. 27,196 +/- 4862.4, p = 0.39). MBF-IR was 1.88 +/- 0.73 in the area covered by stenosed coronary artery, 2.53 +/- 0.75 in unstenosed coronary artery in patients who have significant coronary stenosis and 2.97 +/- 0.77 in normal coronary patients. MBF IRs in the area covered by stenosed coronary arteries were significantly smaller than that of normal coronary artery patient (p = 0.037). Interobserver and intraobserber reproducibility were good (r = 0.96, 0.95 respectively). There was strong positive correlation between MBF-IR and RPP in normal patients (r = 0.69, p = 0.0018), suggesting MBF increase depends on the cardiac workload. CONCLUSION: MBF-IR can be estimated by the combination of TF RN angiography and SPECT at exercise and at rest. PMID- 10714063 TI - [Assessment of noninvasive therapy in lung cancer using lung perfusion images]. AB - The purpose of this prospective study was to follow the changes in functional parameters of radionuclide lung perfusion scans and their role in prognostication of lung cancer cases after noninvasive therapy. We studied 91 patients of lung cancer treated with chemotherapy and/or radiotherapy during 1993 to 1997 in our hospital. Lung perfusion scans were acquired pre and post-therapy. An index of lung perfusion, called Improvement Ratio (IR) was defined as a change in the perfusion of diseased lung as a result of treatment. IR was calculated by the following equation under the assumption that perfusion of contralateral lung remained unaffected. [formula: see text] where Q and Q' are pulmonary arterial blood flow pre and post therapy respectively, p is perfusion ratio of diseased lung before therapy and q is that after therapy. We further studied the relationship between IR and change in tumor size. The influence of tumor location, histopathological diagnosis and prognosis of lung cancer were correlated with this newly defined index. IR in the group of patients with complete response or partial response was significantly higher than in those with poor response (2.72 +/- 0.78 versus 0.99 +/- 0.09, p < 0.05). There was no statistical difference between the group with and without radiotherapy. The score was significantly higher for patients with hilar disease compared to those with peripheral lesions (2.80 +/- 0.83 versus 1.02 +/- 0.03, p < 0.05). Similarly, patients with small cell lung cancer depicted higher values of IR than non-small cell lung cancer (3.36 +/- 1.10 versus 1.06 +/- 0.07, p < 0.05). All those subjects who showed IR > 1 had longer survival time than those with IR < 1 (p < 0.05). It is suggested that improvement in the perfusion of diseased lung predicted better prognosis. We conclude that the evaluation of physiological parameters during therapy using lung perfusion scanning, in addition to lesion size assessment will contribute to the comprehensive follow-up of lung cancer. PMID- 10714064 TI - [Two cases of microvascular vasospastic angina usefulness of 99mTc-tetrofosmin myocardial SPECT in clinical diagnosis]. AB - Case 1 involved a 52-year-old man with angina chest pain at rest and case 2 involved a 63-year-old woman with chest oppression at rest. An electrocardiogram (ECG) showed negative T wave in III and aVF leads in case 1, and complete atrioventricular block and ST segment depression in II, III, aVF, and V5-6 leads in case 2. In both cases, 99mTc-tetrofosmin myocardial SPECT showed reduced uptake in the inferior and posterior wall. Although bath patients' left coronary arteriographies were normal, right coronary arteriographies revealed severely delayed filling of contrast medium without significant narrowing of epicardial coronary arteries, suggesting microembolism or microvascular vasospasm. An intracoronary infusion of isosorbide dinitrate did not improve the delayed filling of contrast medium or ST segment depression on ECG. Soon after intracoronary infusion of diltiazem in case 1 and nicorandil in case 2, coronary arterial flows were normalized, chest symptoms disappeared, and ECG findings were normalized. The next day, both patients' 99mTc-tetrofosmin myocardial SPECT showed normal uptake. These findings suggest that myocardial ischemia in these cases might be explained as having been caused by microvascular spasm. PMID- 10714065 TI - [Prediction of the clinical efficacy of hepatic arterial chemotherapy for metastatic hepatic cancer by intraarterial infusion of 99mTc-MIBI]. AB - SPECT was performed in 11 patients with metastatic hepatic cancer by intraarterial infusion of 99mTc-MIBI before hepatic arterial chemotherapy was started, and the degree of accumulation and clinical efficacy were compared. Early and delayed SPECT images were obtained and various parameters were calculated, including early ratio (ER), delayed ratio (DR), washout rate (WR), and retention index (RI). Judgement of clinical efficacy was made by CT before and after hepatic arterial chemotherapy and was classified as effective, unchanged, and progressive groups. The mean values of ER and DR in the effective group were higher than those in the progressive group. No relationships were noted among the WR and RI values of the groups. The assessment of ER and DR using 99mTc-MIBI intraarterial SPECT is considered to be useful for prediction of the clinical efficacy of hepatic arterial chemotherapy for metastatic hepatic cancer. PMID- 10714066 TI - [Investigation of the resolution and count recovery coefficients of a whole-body PET scanner (Shimadzu SET-2400W) in 2D and 3D mode image]. AB - We measured the resolution and count recovery coefficients (RC) of the SET-2400W whole-body PET scanner (Shimadzu Co., Japan) in the 2D and 3D clinical modes. METHOD: The 3D images were reconstructed by using the full 3D image reconstruction method (3-D reprojection algorithm: 3DRP) and the Fourier rebinning method (FORE). The 2D images were reconstructed with conventional filtered back-projection method (FBP). The measurements of resolution and recovery coefficient were according to JRIA (Japan Radioisotope Association) protocols. RESULTS: The transaxial resolutions of all methods were better than 7 mm FWHM at a radius of 10 cm with 1.25 cm-1 cutoff frequency. The average slice width of 2D FBP, 3DRP and FORE are 5.8 mm, 8.0 mm and 6.8 mm respectively at the center of transaxial field of view. The RC values were measured in a range from 10 mm to 27 mm at 6 cm from the center with the cylindrical and spherical hot area phantoms. In all methods, RC values at 27 mm diameter were nearly 1.0 in both type of hot area. RC values at 10 mm diameter in 2D FBP, 3DRP and FORE of cylindrical hot area were 0.69, 0.72, 0.73 and those of spherical hot area were 0.52, 0.51, 0.53 respectively. CONCLUSION: At the SET-2400W, resolution and recovery coefficient of 3D mode image under the clinical mode showed the value which did not differ from the 2D mode image. PMID- 10714067 TI - [Evaluation of clinical utility of 123I-MIBG scintigraphy in localization of tumors of sympathetic and adrenomedullary origin--a report of multicenter phase III clinical trials]. AB - Phase III clinical study was performed to evaluate clinical utility of 123I-MIBG in the localization of tumors in 48 patients with tumors of sympathetic and adrenomedullary origin, diagnosed or strongly suspected. Sixteen patients had pheochromocytoma, 23 had neuroblastoma, 7 had medullary carcinoma of the thyroid, and 2 had Sipple syndrome. In 3 out of 48 patients, 123I-MIBG scintigraphy was performed twice. The clinical utility of 123I-MIBG was evaluated in 50 cases. Out of 140 lesions, 123I-MIBG scintigraphy demonstrated 51 true positive, 79 true negative, 1 false positive, and 2 false negative. Seven lesions were not evaluable. Sensitivity was 96.2%, Specificity was 98.8%, and Accuracy was 97.7%. An acquisition between 4 hrs and a day after injection was adequate for tumor detection. Neither adverse reactions nor abnormal laboratory findings were noted in relation to 123I-MIBG injections. Our study indicates that 123I-MIBG is a safe and useful radiotracer for visualization and localization of tumors of sympathetic and adrenomedullary origin. PMID- 10714068 TI - [The evaluation of the usefulness of phacotrabeculectomy as the only surgery for glaucoma and cataract patients]. AB - PURPOSE: To evaluate IOP changes after phacotrabeculectomy. MATERIAL AND METHODS: 24 glaucoma patients (27 eyes) with coexistence of cataract. Age: 56-79 years. Mean IOP level before surgery: 23.2 +/- 2.4 mm Hg. In all cases phacotrabeculectomy was performed. Follow-up ranged from 3 to 12 months. RESULTS: IOP decrease after surgery was achieved: mean decrease 7.1 +/- 2.3 mm Hg after 3 months and 5.5 +/- 2.4 mm Hg, and 4.1 +/- 2.1 mm Hg after 6 and 12 months, respectively. CONCLUSION: Phacotrabeculectomy combines all advantages of phacoemulsification and creates perspective for IOP normalization. PMID- 10714069 TI - [The influence of 0.85% RS-timolol and 0.5% S-timolol on intraocular pressure and systemic arterial blood pressure, heart rate, ECG, expiratory capacity in patients with ocular hypertension and primary open-angle glaucoma]. AB - PURPOSE: Analysis of local and systemic effects of RS-timolol enantiomer and S timolol in patients with ocular hypertension and glaucoma: evaluation of Risk/Benefit Ratio. MATERIAL AND METHODS: 19 patients (38 eyes) receiving 0.85% RS-timolol and 10 patients (20 eyes) receiving 0.5% S-timolol were evaluated using a double-blank test. Intraocular pressure, heart rate, systemic arterial blood pressure, ECG and spirometry were recorded before and 2 hours after drugs administration and after 7 days of treatment. RESULTS: There were no statistically significant differences between intraocular pressure-lowering effects of RS-timolol and S-timolol. In patients receiving both medicines bradycardia was detected 2 hours after drugs administration. Patients receiving RS-timolol showed increased expiratory capacity in comparison to those receiving S-timolol. There was no detectable influence of both medicines on ECG and systemic arterial blood pressure. CONCLUSIONS: 0.85% RS-timolol and 0.5% S timolol produced comparable intraocular pressure-lowering effects. 0.85% RS timolol exerted less severe influence on respiratory system. PMID- 10714070 TI - [Post-steroid elevations of intraocular pressure in patient after radial keratotomy]. AB - PURPOSE: The evaluation of frequency and amplitude of IOP elevations in patients after RK who were administrated topically dexamethasone. MATERIAL AND METHODS: RK was carried out in 90 eyes (55 patients). All these patients had been treated topically with 0.1% dexamethasone (Maxitrol, Alcon) since the first day after surgery till the period of 3 months in lowering doses. IOP was measured using air puff tonometer (Reichert, USA) before surgery and on 1, 2, 3, 14, 30, 50, 90 day after RK. In case of IOP elevations the measurements were made more often. RESULTS: The mean IOP before surgery was 14.88 +/- 2.86 mm Hg for women and 16.14 +/- 2.83 mm Hg for men. In the period of 3 months after RK maximum IOP increased significantly both for women (mean: 21.46 +/- 7.51 mm Hg) and men (mean: 26.14 +/ 8.87 mm Hg). IOP higher than 25 mm Hg was observed in 35 eyes (37.7%). These IOP elevations were observed more often in men than women but this difference was not statistically significant. There was no correlation between frequency of IOP elevations and preoperative refractive error or the age of patients. CONCLUSIONS: The usage of steroids after RK requires careful monitoring of IOP. This subject needs further studies to answer if these IOP elevations can damage eye functions. PMID- 10714071 TI - [Intraocular lens implantation in traumatic cataract]. AB - PURPOSE: The evaluation of postoperative results and complications after traumatic cataract extraction with intraocular lens implantation. MATERIAL AND METHODS: Forty two eyes of 41 patients (31 male and 10 female) aged from 10 to 66 years (mean 37.8) with traumatic cataract were the subject of our study. They were operated on between 1996 and 1997. A penetrating injury in 33 eyes (78.6%) and blunt trauma in 9 eyes (21.4%) caused the cataract. The interval between trauma and cataract surgery ranged from 1 day to 40 years. The mean follow up was 13.5 months. Despite of cataract corneal scars, pupil deformations, posterior and anterior synechiae were observed in most of the eyes. Extracapsular cataract extraction was performed in all cases. Posterior chamber IOLs were implanted in 33 eyes. Anterior vitrectomy and anterior chamber IOL implantation was performed in 9 eyes due to the lack of capsular support. RESULTS: Very good or good visual acuity (5/5-5/16) was achieved in 71.4% of cases. Low visual acuity (below 5/50) was observed in 6 eyes (14.3%) because of severe damage to the retina, optic nerve atrophy or amblyopia. Fibrin reaction in anterior chamber (30.9%), hyphaema (19%) and haemophtalmus (4.8%) were the most severe postoperative complications. CONCLUSIONS: An intraocular lens implantation in traumatic cataract, despite many postoperative complications, enables most of the patients to achieve satisfactory and useful vision. Patients with severe posterior segment damage do not benefit functionally from cataract surgery. PMID- 10714072 TI - [Evaluation of endothelial cells after the usage of gentamycin in irrigation aspiration fluid during extracapsular cataract extraction]. AB - PURPOSE: Assessment of the density of endothelial cells after usage of gentamicin in irrigation-aspiration fluid. MATERIAL AND METHODS: 60 patients subjected to extracapsular cataract extraction with intraocular lens implantation. In 30 eyes irrigation was performed with Ringer fluid (group I) and in 30 eyes Ringer fluid with gentamicin (group II) 4 mg gentamicin were added to 500 ml of Ringer fluid obtaining the concentration 8 micrograms/ml. Evaluation of endothelial cells density was carried before operation, 1 month and 6 months after. RESULTS: Percentage endothelial cells loss in group I was 7.8% 1 month postoperatively and 14.5% 6 months after operation and in group II 8.5% and 14.87%, respectively. CONCLUSION: Gentamicin added to irrigation-aspiration fluid doesn't cause enhanced endothelial cells loss during extracapsular cataract extraction. PMID- 10714073 TI - [Comparative evaluation of corneal endothelial cells after extracapsular cataract extraction with PMMA and heparin-surface modified lens implantation]. AB - PURPOSE: Comparison of endothelial cells density in eyes after PMMA and heparin surface-modified lenses (HSfM1) implantation. MATERIAL AND METHODS: 58 patients (26 male and 32 female) aged from 36 to 89 years. In 30 patients PMMA lenses produced by Pharmacia, Storz and Alcon and in 28 HSM (Pharmacia) lenses were implanted, after ECCE. RESULTS: Percentage loss of endothelial cells in eyes with PMMA lenses was 8.61% after month and 13.56% 6 months post-operatively. In eyes with HSM lenses endothelial cell loss was 8.44% and 13.55% respectively. CONCLUSION: Loss of corneal endothelial cells in eyes with PMMA lenses is almost the same in eyes with HSM lenses. PMID- 10714074 TI - [The use of indocyanine green angiography in diagnosis of occult choroidal neovascularization in age-related macular degeneration]. AB - PURPOSE: To determine whether scanning laser ophthalmoscopy (SLO) indocyanine green (ICG) angiography may be useful in detecting occult or ill-defined choroidal neovascularization (CNV). MATERIAL AND METHODS: ICG angiography using SLO was performed in 28 patients (29 eyes). Indication for this examination was occult or ill-defined CNV on fluorescein angiography. RESULTS: Well-defined hyperfluorescent lesions on ICG angiography were found in 16 eyes (55.2%). Three morphological types of CNV were noted: plaque (82.8%), focal spots (10.4%), combination lesions (3.4%). In one case no manifestations of CNV were observed in ICG angiography (3.4%). In 4 eyes (13.8%) we performed successful laseroterapy of CNV. CONCLUSIONS: ICG angiography performed using SLO is a valuable tool in determining borders of occult and ill-defined CNV. PMID- 10714075 TI - [Visual evoked potentials in diagnosis and monitoring of optic neuropathy in the course of thyroid ophthalmopathy]. AB - PURPOSE: To evaluate the usefulness of pattern visual evoked potentials (PVEP) in early diagnosis and monitoring of dysthyroid optic neuropathy (DON). MATERIAL AND METHODS: We recorded PVEP (UTAS E--1000) in 74 patients with thyroid ophthalmopathy (TO) including 12 patients with clinically evident DON--group I, 13 patients with subclinical DON (prolongation of latency of P100 wave in PVEP)- group II, and 49 patients without clinical or electrophysiological signs of DON- group III. Thirty six healthy subjects served as controls. The latencies of P100 and N75 waves were examined. In 50 TO patients we recorded changes in PVEP before and after treatment. RESULTS: The mean LP100 and LN75 were significantly longer in TO patients compared to control group. In groups I and II they were also significantly longer in comparison to group III. We observed prolongation of LP100 above the upper limit of normal values in all patients with clinically evident DON and in 13/62 (21%) cases without clinical signs of optic nerve dysfunction. The prolongation of LP100 was present in 16/21 (76.2%) eyes in group I and in 15/26 (57.7%) eyes in group II. Prolongation of LN75 was less frequent: group I--5/21 (23.8%), group II--2/26 (7.7%). After treatment in patients with clinically evident DON the latencies of both waves were significantly shorter in comparison to values obtained before therapy. In group II statistically significant differences were observed only in LP100. The percentage of eyes with prolonged latencies was significantly smaller after treatment (group I: LP100- 3/23 (13.0%), LN75--1/23 (4.3%); group II: LP100--2/18 (11.1%), LN75--no prolongation was observed). CONCLUSIONS: PVEP is an useful method in diagnosis and monitoring of dysthyroid optic neuropathy. It can reveal even asymptomatic optic nerve dysfunctions. Detection of subclinical neuropathy using PVEP enables early--thus more effective--treatment. The most important factor in PVEP evaluation is the latency of P100 wave, the latency of N75 wave is less useful. PMID- 10714076 TI - [Local parabulbar anesthesia with Greenbaum cannula for most common ophthalmic operations]. AB - PURPOSE: To compare two methods of local anesthesia in ophthalmic surgery: classic retrobulbar and new--parabulbar made with Greenbaum cannula and to try to assess new method. MATERIAL AND METHODS: 300 patients underwent most common ophthalmic operations: ECCE + PCLI and glaucoma surgery. 150 of them were anesthetised by retrobulbar and 150 by parabulbar (flush) with Greenbaum cannula methods. We compared the presumed influence of these two types of anesthesia on the operations assessing the following: the occurrence "vis a tergo" and posterior capsule rupture. We compared also efficacy of both methods (analgesia and akinesia) and complications (retrobulbar hematoma, globe perforation). We minimalized the volume of anesthetic mixture used in retrobulbar method to 1.5 ml (0.5 ml 0.5% bupivacaine + 1 ml 2% xylocaine). RESULTS: The number of complications was lower in the group anesthetised by parabulbar method. The complications of local anesthesia were bigger in retrobulbar method (retrobulbar hematoma, globe perforation). Anesthesia in parabulbar method in spite of little volume of anesthetic mixture was very good but akinesia--slightly weaker comparing to retrobulbar injection. CONCLUSIONS: Parabulbar anesthesia made with Greenbaum cannula is a very good, safe method giving very good anesthesia no possibility of globe perforation or retrobulbar hematoma, but the method is for skilled surgeons because of weaker akinesia. PMID- 10714077 TI - [Keratectomy for a malignant melanoma of the cornea]. AB - PURPOSE: To present the possibility of malignant melanoma treatment of the cornea using keratectomy. MATERIAL AND METHODS: A 59-year old woman with a primary malignant melanoma of the cornea. Visual acuity before surgery 0.2. Malignant melanoma was removed using keratectomy. Follow-up 18 months. RESULTS: Satisfactory cosmetic result and visual acuity 1.0 was obtained. In 18 months follow-up no local or general symptoms of tumor recurrence are observed. CONCLUSION: Keratectomy should be considered as an alternative treatment for primary malignant melanoma of the cornea. PMID- 10714078 TI - [Capsule contraction syndrome]. AB - AIM: To analyse the occurrence of the anterior capsule contraction following cataract surgery. Capsule contraction syndrome (CCS) is defined as an extreme reduction in diameter of anterior capsulectomy, capsular bag diameter and, occasionally, displacement of the IOL after extracapsular cataract extraction. It is relatively frequent in pseudoexfoliation, advanced age, in association with uveitis, pars planitis and myotonic muscular dystrophy. MATERIALS AND METHODS: 5965 eyes of patients were operated on cataract between 1.01.1994 and 31.12.1997 in Tadeusz Krwawicz Chair of Ophthalmology and 1st Eye Hospital, Medical School in Lublin. Two types of surgical procedures were performed: "divide and conquer" phacoemulsification with 4.5-8 mm continuous curvilinear capsulorhexis (3385 eyes) and extracapsular cataract extraction with "can opener" capsulotomy (2580 eyes). RESULTS: 20 cases of clinically apparent CCS were referred to the Department: in the course of intensive postoperative inflammation--5, in patients over 80--4, in pseudoexfoliation syndrome--2, myotonic dystrophy--1, ectopia lentis--2, other causes--6. In order to improve visual acuity in 4 cases surgical removal of the distorted and opaque anterior capsule was performed, in 3 cases relaxing radial tears were done, in 3 cases secondary anterior capsulotomy was performed using Q-switched Nd:YAG laser. CONCLUSIONS: In cases where the occurrence of CCS is especially high large diameter capsulorhexis should be performed and IOL designed to provide maximal peripheral capsular bag expansion should be implanted. PMID- 10714079 TI - [Ocular complications of cerebrospinal fluid leakage after skull trauma]. AB - PURPOSE: Analysis of diagnostic and therapeutic ophthalmological problems after severe cerebrocranial lesions with liquorrhea. MATERIAL: 3 children between 5 months and 5 years in whom the ophthalmological symptoms were the first and important signs of the cerebrospinal fluid leakage after skull lesion. These patients were observed and diagnosed at the department of ophthalmology and then operated at the department of neurosurgery. RESULTS: In all children skull trauma has led to dura mater tear and leakage of cerebrospinal fluid with its complications. As a result of the ophthalmological complications, two patients developed worse visual acuity evoked potentials. Blind eyeball of the third patient was enucleated. CONCLUSIONS: The ocular findings are not typical for liquorrhea after cerebrocranial trauma and they must be diagnosed and treated by a team of ophthalmologists and neurosurgeons. PMID- 10714080 TI - [Phantom vision phenomenon]. AB - The subject of the paper is the phenomenon of phantom vision. It occurs among the blind (or almost blind) and is characterised by perceiving various visual sensations, for example: light, geometrical shapes, buildings, people, flowers etc. The paper presents views explaining the generation of phantom vision. According to Melzack's theory, the occurrence of these sensations is explained in relation to the phenomenon of transforming the neuro-activity of the brain in a specific--for each organ--neuronal network called neuromatrix. Even if some organs do exist but have ceased to function completely (or to a significant degree) and consequently the brain does not receive any sensory stimuli from them, the neuromatrix in brain can generate visual sensations constituting the phantom vision phenomenon. The paper also presents three cases of phantom vision. PMID- 10714081 TI - [The use of molecular biology achievements in glaucoma diagnosis]. AB - Glaucoma is the third leading cause of blindness in the world. The molecular basis of this disease is unknown, although it is likely to be genetically heterogeneous disorder that results from the interaction of multiple genes and environmental influences. Current achievements and new trends of molecular biology used in glaucoma diagnosis are presented. PMID- 10714082 TI - [Pharmacotherapy of congenital glaucoma in young children]. AB - The aim of congenital glaucoma treatment is to preserve visual function of children's eyes. There are three forms of congenital glaucoma: primary congenital glaucoma of infants, children and the youth, glaucoma in dysgenesis of anterior segment of the eyeballs and glaucoma in congenital systemic diseases. Pharmacotherapy is indicated during preparation for surgical treatment, as an adjunct following surgical procedures and as a protection of retinal and optic nerve function. Carbonic anhydrase inhibitors, hyperosmotic agents, sympathomimetic drugs, parasympathomimetic drugs, prostaglandins may be used. Dopamine antagonists, calcium channel blockers, serotonin antagonists are indicated as a protection of retinal cells and optic nerve function. The most widely used are carbonic anhydrase inhibitors and beta-blocker: betaxolol. Antiglaucoma drugs have proven to be efficacious in treating children with congenital glaucoma but they have several serious side effects. Therefore, pharmacotherapy may be used only periodically. The main treatment of congenital glaucoma is surgical treatment. PMID- 10714083 TI - [Possibilities of diagnosis and treatment of eye disease by general practitioners]. AB - PURPOSE: To analyse possibilities of diagnosis and treatment of eye diseases by practitioner. MATERIAL AND METHODS: The number and diagnosis profile of patients treated in average municipal out-patient ward was calculated and possibilities of their treatment by general practitioner were analysed. RESULTS: In 87.4% of eye patients general practitioner cannot diagnose and treat the disease and he has to send them to an eye-doctor. CONCLUSION: The system that eye-patient cannot check in directly to an eye-doctor is more costly because general practitioners can diagnose and treat only small number of these patients. Therefore nearly every patient has to be consulted be eye-specialist, which increases the costs of treatment because patient has to be seen by two doctors. PMID- 10714084 TI - Eosinophil activation markers in induced sputum in asthmatics. AB - OBJECTIVES: Eosinophils play an important role in asthmatic airway inflammation collaborately with other inflammatory cells. The present study was aimed to determine whether the eosinophil activation markers in induced sputum reflect the clinical status in asthmatics. METHODS: The clinical severity and FEV1 were measured. Hypertonic saline induced sputum was obtained from 25 asthmatics and ten control subjects. We processed freshly expectorated sputum separated from saliva by treatment with an equal volume of dithiothreitol 0.1%, cytospins for cell count and special stain, and a collection of the supernatant for biochemical assay. We used a fluoroimmunoassay to detect eosinophil cationic protein (ECP), and a sandwich ELISA to detect interleukin (IL)-5. RESULTS: Asthmatics, compared with control subjects, had a significantly higher proportion of eosinophils (25.6 +/- 4.6% vs 1.7 +/- 0.2%, p < 0.01) and higher levels of ECP (1117.8 +/- 213.9 micrograms/L vs 154.6 +/- 47.4 micrograms/L, p < 0.01) in their sputum. IL-5 was detected more frequently in asthmatics than in control subjects [11/25 (44%) vs 1/10 (10%), p < 0.05]. Moderate to severe asthmatics had a significantly higher proportion of eosinophils, higher levels of ECP and IL-5 compared to mild asthmatics. FEV1, FEV1/FVC were significantly correlated with the proportion of eosinophils and the levels of ECP and IL-5. Significant positive correlations were noted between the proportion of eosinophils and the level of ECP and IL-5. Sputum ECP level showed a significant positive correlation with IL-5 level. CONCLUSION: These findings demonstrate that eosinophils and the eosinophil activation markers, such as ECP and IL-5 in induced sputum, are closely related to the clinical status in asthmatics. Induced sputum study may thus be useful in clinically measuring indices of airway inflammation in asthma. PMID- 10714085 TI - Initial and late results of Freedom coronary stent. AB - OBJECTIVES: Initial and late results after implantation of Freedom stents, a balloon expandable stainless steel coil stents were evaluated. METHODS: From Jun. 1996 to Nov. 1997, we implanted 123 Freedom stents in 122 lesions in 117 patients and performed follow-up coronary angiograms at 7.0 +/- 3.6 months after stents placement. Clinical courses after stenting and follow-up coronary angiographic findings were evaluated. Comparison of clinical, angiographic, and procedural factors according to the presence or absence of restenosis was performed. RESULTS: In 117 patients who underwent stents implantation, major complications were not observed. Follow-up coronary angiograms were performed in 47 stents in 41 patients (35%). Among 47 stents, angiographic significant restenosis (percent diameter stenosis > 50%) was observed in 13 (28%). Mean age in 41 patients was 59 +/- 9 years, with 27 male patients (66%). Indications for stents implantation were de novo lesions in 18 (38%), suboptimal results after PTCA in 18 (38%), bail out lesions in 4 (9%) and restenotic lesions in 7 (15%). Lesion types by AHA/ACC classification were A in 1 (1%), B1 in 10 (21%), B2 in 17 (36%), and C in 19 (40%). Average lesion length was 13.7 +/- 9.0 mm, stent diameter 3.0 +/- 0.3 mm, and stent length 24.6 +/- 9.0 mm. There were no significant differences of the clinical, angiographic, and procedural characteristics according to the presence or absence of restenosis. CONCLUSION: Freedom coronary stents implantation is safely performed in various morphology of coronary lesions and no significant predictive factors on restenosis in follow-up coronary angiogram were observed. PMID- 10714086 TI - ACE gene polymorphism and renal responsiveness to ACE inhibitors in IgA nephropathy patients. AB - We examined the renal responsiveness to ACE inhibitor in IgA nephropathy (IgAN) patients according to the grouping of ACE gene polymorphism. Sixty one patients diagnosed as IgAN by renal biopsy and prescribed with ACE inhibitors were enrolled. Genomic DNA was extracted from whole blood and PCR was performed. The I/D polymorphism was determined by the presence of the 287 bp fragment in intron 16 of chromosome 17. During the follow-up period (mean; 44.6 months, median: 44.5 months, 5 to 113 months), the blood pressure of 61 patients was controlled below 130/80 mmHg. The renal responsiveness was determined by the degree of changes of proteinuria at 12 months after initiation of ACE inhibitors and by the slope of reciprocal variation of the serum creatinine against follow-up duration ?(1/Cr2 1/Cr1)/durations?. The distribution of the II, ID and DD genotype among 61 patients was 21, 16 and 24 patients, respectively. There were no differences among three genotypes in age, sex, the number of patients with initial blood pressure over 140/90 mmHg, initial serum creatinine level, the number of patients with initial azotemia (> 1.4 mg/dL) and with initial 24-hr proteinuria amount over 2.0 g. Significant anti-proteinuric effect of ACE inhibitor was found in IgAN (p = 0.001), but no significant difference was found among genotypes. Significant difference (p = 0.011) was noticed between II type and DD type in the slope of reciprocal variation of the serum creatinine against follow-up duration. In conclusion, efficacy of ACE inhibitors on renal function preservation in IgAN was more pronounced in DD genotype than II genotype. PMID- 10714087 TI - Changes in gallbladder motility in gastrectomized patients. AB - OBJECTIVES: Gastric resection may predispose gallstone formation. However, the mechanism has not been clearly understood. To evaluate the relationship between gastric resection and gallstone formation, we compared gallbladder(GB) motility in gastrectomized patients and control subjects. METHODS: We compared the GB volume and ejection fraction of the 46 gastrectomized patients with 37 healthy controls using real time ultrasonography. RESULTS: GB volume increased significantly in the gastrectomized group in fasting (30.2 +/- 13.9 ml). The GB volume after a fatty meal was greater in the gastrectomized group (12.6 +/- 6.4 ml) than in the control group (4.3 +/- 3.3 ml) (p < 0.01). A significant reduction of ejection fraction was found in gastrectomized patients (56.9 +/- 13.0%) in comparison with the control group (75.5 +/- 16.1%) (p < 0.01). The GB ejection fraction had a poor correlation to the postoperative period (r = 0.232). CONCLUSION: A gastrectomy appears to be a risk factor of GB dysmotility, which may play a major role in gallstone formation in gastrectomized patients. PMID- 10714088 TI - The influence of gastrectomy on the change of bone metabolism and bone density. AB - OBJECTIVES: Abnormalities of bone metabolism could be followed in gastrectomized patients as a late complication. Nowadays, many biochemical and radiologic measurements are applied to detect these abnormalities. The aim of our study is to determine the valuable parameter as an appropriate screening test during long term follow-up periods and define the usefulness of new biochemical markers for bone metabolism by comparing with traditional markers. METHODS: Fifteen patients who had undergone partial gastrectomy were chosen randomly and fifteen healthy controls were compared. Then, several biochemical and radiologic tests were measured. We excluded subjects who proved to have other causes of bone metabolism abnormalities. Ten patients and 10 controls were finally selected. RESULTS: Comparing the data with those of a corresponding control group, the lumbar bone density measured by quantitative computed tomography (QCT) was statistically significantly lower in the patient group (p < 0.01). The urinary deoxypyridinoline, a biochemical marker for bone resorption, was statistically higher in the patient group (p < 0.025). Osteocalcin, Procollagen I C-terminal peptide (PICP) and Type I collagen C-terminal telopeptide (ICTP) were slightly but not significantly higher in the patient group. The serum parathyroid hormone (PTH) and 25-hydroxy vitamin D levels were similar in both groups. CONCLUSION: We could suggest that urinary deoxypyridinoline and QCT are appropriate parameters as screening tests for the detection of bone metabolism abnormalities in gastrectomized patients during long-term follow-up. Urinary deoxypyridinoline may be a simple and rapid test which could replace cumbersome 24-hour urinary hydroxyproline. PMID- 10714089 TI - Difference in the distribution pattern of Helicobacter pylori and grade of gastritis in the antrum and in the body between duodenal ulcer and benign gastric ulcer patients. AB - OBJECTIVES: To investigate the relationship between the Helicobacter pylori (H. pylori) colonization and the grade of gastritis in the antrum and in the body of patients with duodenal ulcer (DU) or benign gastric ulcer (BGU). METHODS: This study was performed in H. pylori-positive 220 DU patients and 180 BGU patients. H. pylori density was evaluated by modified Giemsa staining and CLO test, and gastritis grade was graded by H&E staining in the antrum and in the body. RESULTS: H. pylori grade by Giemsa staining was 1.24 in the antrum and 0.82 in the body for DU group (p < 0.01), and those of BGU group were slightly reversed, 0.83 and 0.87, respectively, but without statistical significance. Similarly H. pylori grade by CLO test was 3.1 in the antrum and 2.8 in the body for DU group (p < 0.01), and those of BGU group 2.3 and 2.6 (p < 0.05), respectively. In contrast, gastritis grade was 1.7 in the antrum and 1.2 in the body for DU group (p < 0.01), and those of BGU group 1.6 and 1.3 (p < 0.01), respectively, similar to those of DU. However, there was a correlation between H. pylori grade and gastritis grade in the antrum and in the body, not only in DU but also in BGU group (p < 0.01). CONCLUSION: In spite of different distribution patterns of H. pylori between DU group and BGU group, gastritis grade of the antrum was significantly higher than that of the body in both DU and BGU. However, gastritis is correlated with H. pylori density not only in DU but also in BGU patients. It looks like the inflammatory reaction to H. pylori is stronger in the antrum than in the body. PMID- 10714090 TI - Comparison of glucose tolerance categories in the Korean population according to World Health Organization and American Diabetes Association diagnostic criteria. AB - OBJECTIVES: To compare the prevalence and metabolic profiles of glucose tolerance categories according to World Health Organization(WHO) and 1997 American Diabetes Association(ADA) fasting criteria for the diagnosis of diabetes mellitus and impaired glucose metabolism in the Korean population. METHODS: 2251 subjects without previous history of diabetes, who participated in the Yonchon diabetes epidemiology survey in 1993, were classified according to both criteria. The prevalence of glucose tolerance categories and the agreement across all categories of glucose tolerance were calculated. Metabolic characteristics of different glucose tolerance categories were compared. RESULTS: The prevalence of diabetes and impaired fasting glucose(IFG) according to ADA fasting criteria was similar to those of diabetes and impaired glucose tolerance(IGT) according to WHO criteria, respectively. However, 35.5% of the subjects who were diagnosed as diabetes by WHO criteria were reclassified as either IFG or normal fasting glucose (NFG), and 38.5% of diabetic patients according to ADA fasting criteria were IGT or normal glucose tolerance (NGT) by WHO criteria. Only 31.3% of IGT subjects remained as IFG and 62.1% were reclassified as NFG. Similarly, 69.4% of IFG subjects were NGT by WHO criteria. The agreement between the two criteria was poor (K = 0.31). Discordant diabetes groups had higher WHR, systolic and diastolic blood pressure, cholesterol and triglyceride levels than concordant non diabetes group. Non-diabetes(WHO)/diabetes(ADA) group had higher WHR than diabetes (WHO)/non-diabetes(ADA) group. There were no differences in other metabolic characteristics between the two discordant diabetes groups. IGT/NFG and NGT/IFG group showed higher BMI, WHR, systolic and diastolic blood pressure, cholesterol and triglyceride levels than NGT/NFG group. Metabolic characteristics of IGT/NFG group were not different from those of NGT/IFG group except IGT/NFG subjects were older than NGT/IFG subjects. CONCLUSION: The agreement between WHO and ADA fasting criteria was poor. ADA fasting criteria can detect new diabetic patients and subjects with impaired glucose metabolism who are not classified as diabetes or IGT by WHO criteria. However, a substantial number of subjects, who may have increased cardiovascular risk and/or increased risk for the development of diabetes and its complication, will be missed when using ADA fasting criteria. PMID- 10714091 TI - Urinary bladder involvement in patients with systemic lupus erythematosus: with review of the literature. AB - OBJECTIVES: To investigate the etiologies of urinary bladder involvement in patients with systemic lupus erythematosus (SLE), the clinicoradiologic features of gastrointestinal tract manifestations and clinical outcomes in patients with lupus cystitis accompanied by gastrointestinal manifestations. METHODS: We conducted a retrospective chart review on 413 patients with SLE. Patients were selected for review on the basis of lower urinary tract symptoms including urinary frequency, urgency and urinary incontinence. Radiologic studies were analyzed in patients with lupus cystitis. RESULTS: Ten consecutive patients, complicated with lower urinary tract symptoms, were identified. Underlying etiologies were as follows: lupus cystitis in five, neurogenic dysfunction secondary to transverse myelitis in three, cyclophosphamide-induced cystitis in one and tuberculous cystitis in one patient. All patients with lupus cystitis showed gastrointestinal manifestations, such as abdominal pain, nausea, vomiting and/or diarrhea during the periods of cystitis symptoms. In all patients with lupus cystitis, paralytic ileus was demonstrated on plain abdominal X-ray and ascites, bilateral hydroureteronephrosis and thickened bladder wall were identified on abdominal ultrasound or CT. Abdominal CT revealed bowel wall thickening in four of the five patients. The main sites of thickened bowel on abdominal CT were territory supplied by superior mesenteric artery. Two of five patients with lupus cystitis expired during the follow-up period. CONCLUSION: Diverse etiologies may cause lower urinary tract symptoms in patients with SLE. Lupus cystitis is strongly associated with gastrointestinal involvement and abdominal CT can be a useful radiologic tool to investigate the gastrointestinal tract involvement in patients with lupus cystitis. PMID- 10714092 TI - Comparison of acquired cystic kidney disease between hemodialysis and continuous ambulatory peritoneal dialysis. AB - OBJECTIVES: ACKD has been described mainly in patients treated with hemodialysis(HD), and there are only a few reports about the prevalence of ACKD in continuous ambulatory peritoneal dialysis (CAPD) patients. Therefore, we compared the prevalence of ACKD in patients receiving HD and CAPD, and evaluated the possible factors which may affect the development of ACKD. METHODS: Forty nine HD and 49 CAPD patients who had received dialysis therapy for at least 12 months were enrolled in this cross-sectional study. Patients who had a past history of polycystic kidney disease and had acquired cystic kidney disease on predialysis sonographic exam were excluded. Detection of ACKD was made by ultrasonography and ACKD was defined as 3 or more cysts in each kidney. RESULTS: The prevalence of ACKD was about 31% (30/98) and there was no significant difference between HD and CAPD patients(27% vs. 34%, p > 0.05). The prevalence of ACKD was not associated with age, sex, primary renal disease, the levels of hemoglobin, BUN, and serum creatinine. However, the duration of dialysis was significantly related to the development of ACKD (presence of ACKD, 74.4 +/- 42.4 months vs. absence of ACKD, 37.8 +/- 24.1 months, p < 0.05). CONCLUSION: The prevalence of ACKD is not different according to the mode of dialysis, and the major determinant of acquired cyst formation is duration of dialysis. PMID- 10714093 TI - Apoptosis in dilated cardiomyopathy. AB - OBJECTIVE: Cardiomyopathy, a popular diagnosis that always obscures more than it reveals, nevertheless has several characteristic histological features. These prominently include widespread focal myocardial fibrosis and associated hypertrophy of surviving cardiac myocyte. In fact, focal noninflammatory degeneration (not necrosis) has been demonstrated as a feature of many forms of cardiac hypertrophy. We hypothesized that this loss of myocardial cells in dilated cardiomyopathy (DCMP) may result from cell death by apoptosis. METHODS: Endomyocardial biopsy specimens from the right ventricles of six patients who suffered from DCMP were studied, and myocardial specimens from two persons who died in motor vehicle accidents were used as negative controls. For identification of apoptosis, immunohistochemistry with terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick end-labeling was performed. In addition, apoptosis was confirmed morphologically by confocal laser scanning microscopy with propidium iodide. RESULTS: Apoptosis, that was represented by an apoptotic index ranging from 19.8 to 25.4%, could be extensively seen in myocytes and also rarely in non-myocytes of interstitium and vascular endothelium. Morphologically, there were a lot of nuclei with clumps of condensed chromatin, suggestive of apoptosis. CONCLUSION: The present study demonstrated that myocyte loss in DCMP might be mainly due to the apoptosis of myocytes and interstitial cells, rather than inflammation or cell necrosis. PMID- 10714094 TI - Aging affects the association between endothelial nitric oxide synthase gene polymorphism and acute myocardial infarction in the Korean male population. AB - OBJECTIVES: The aging process affects responsiveness and other functions of endothelium and vascular smooth muscle cells, predisposing the old vessels to the development of atherosclerotic lesions. Endothelial nitric oxide synthase (ecNOS) gene polymorphisms were shown to affect the occurrence of acute myocardial infarction (AMI). We hypothesized that aging may affect the association between the ecNOS gene polymorphism and AMI. METHODS: We investigated the age-related distribution of the ecNOS gene a/b polymorphism in 121 male AMI patients and 206 age-matched healthy male controls. RESULTS: The aa, ab and bb genotypes were found in 1, 49 and 156 cases among the control subjects and 5, 23 and 93 cases among the AMI patients, respectively. There was a significant correlation between the ecNOS polymorphism and AMI (p = 0.045). When the correlation was analyzed by age, the significance remained only in the group below the age of 51 (p = 0.009). The proportion of smokers was increased in the young patients when compared to the old patients (p = 0.033), indicating that smoking also has greater effect on the younger population. The incidences of hypertension and diabetes mellitus, however, were similar in both populations. CONCLUSION: Our work provides the first evidence that links ecNOS polymorphism to the risk of AMI in relation to age. Young persons who smoke or have ecNOSaa genotype may have an increased risk of developing AMI. The functional as well as structural changes associated with aging in the vascular endothelium may mask the effect of the ecNOS polymorphism in the development of AMI in old persons. PMID- 10714095 TI - The expression of the high mobility group I(Y) mRNA in thyroid cancers: useful tool of differential diagnosis of thyroid nodules. AB - OBJECTIVE: Thyroid nodule is frequent and occurs in about 5% of the general population. In contrast, thyroid cancer is much less frequent and occurs in about 5-10% of thyroid nodules. Distinguishing between benign and malignant lesions is an important task that is best accomplished by fine needle aspiration. Recently, Chiappetta et al. reported that the expression of the high mobility group (HMG) I(Y) proteins correlates with the malignant phenotype of human thyroid neoplasia, and suggested that the detection of the HMG I(Y) proteins might be a valid tool for an easy and sensitive discrimination assay between benign and malignant neoplastic thyroid disease. METHODS: We evaluated the expression of the HMG I(Y) mRNA in 39 frozen thyroid tissues from patients with thyroid nodule by semiquantitative RT-PCR. RESULTS: The expression of the HMG I(Y) mRNA was low in all of 10 normal thyroid tissues. In all of 3 adenomatous goiters, 6 follicular adenomas and 2 Hurthle cell adenomas, the HMG I(Y) mRNA expression level was low. In 11 of 13 papillary carcinomas and all of 5 follicular carcinomas, the HMG I(Y) mRNA expression level was high. CONCLUSION: These results indicate that there is a correlation between the expression of HMG I(Y) and the malignant phenotype of thyroid cancer, suggesting that these proteins may be useful as a marker in thyroid cancer. PMID- 10714096 TI - Non-Hodgkin's lymphoma of the thyroid and adrenal glands. AB - We report a case of non-Hodgkin's lymphoma(NHL) with simultaneous involvement of both thyroid and bilateral adrenal glands. Literature review on a computerized search showed that this is an extremely rare condition. The final diagnosis of diffuse large B cell lymphoma was confirmed by biopsies of thyroid gland, enlarged cervical lymph node, and adrenal gland. The significant endocrine dysfunction of the thyroid, adrenal or other endocrine glands was absent in our case. The patient responded dramatically to three cycles of chemotherapy with no complication or endocrine dysfunction and continues to be followed. PMID- 10714097 TI - Nonocclusive mesenteric ischemia in a patient on maintenance hemodialysis. AB - Nonocclusive mesenteric ischemia (NOMI) is known to occupy about 25% to 60% of intestinal infarction. NOMI has been reported to be responsible for 9% of the deaths in the dialysis population and the postulated causes of NOMI include intradialytic hypotension, atherosclerosis and medications, such as diuretics, digitalis and vasopressors. Clinical manifestations, such as fever, diarrhea and leukocytosis, are nonspecific, which makes early diagnosis of NOMI very difficult. CASE: A 66-year-old woman on maintenance hemodialysis for 5 years was admitted with syncope, abdominal pain and chilly sensation. Since 7 days prior to admission, blood pressure on the supine position during hemodialysis had frequently fallen to 80/50 mmHg. Four days later, she complained of progressive abdominal pain. Rebound tenderness and leukocytosis (WBC 13900/mm3) with left shift were noted. Stool examination was positive for occult blood. Abdominal CT scan showed a distended gall bladder with sludge. Under the impression of acalculous cholecystitis, she was operated on. Surgical and pathologic findings of colon colon were compatible with NOMI. Because of recurrent intradialytic hypotension, we started midodrine 2.5 mg just before hemodialysis and increased the dose up to 7.5 mg. After midodrine therapy, blood pressure during dialysis became stable and the symptoms associated with hypotension did not recur. CONCLUSION: As NOMI may occur within several hours or days after an intradialytic hypotensive episode, abdominal pain should be carefully observed and NOMI should be considered as a differential diagnosis. In addition, we suggest that midodrine be considered to prevent intradialytic hypotensive episodes. PMID- 10714098 TI - Dermatomyositis without elevation of creatine kinase presented as bronchiolitis obliterans organizing pneumonia. AB - A case of dermatomyositis presented as bronchiolitis obliterans organizing pneumonia has been rarely reported. We describe a 46-year-old female patient with dermatomyositis without elevation of creatine kinase presented as bronchiolitis obliterans organizing pneumonia. She was treated with prednisolone and azathioprine. Over a 2-year follow-up she has had no elevation of creatine kinase. The patient remains asymptomatic and has no medication for dermatomyositis and bronchiolitis obliterans organizing pneumonia two years after initial treatment. It has been suggested that the prognosis of dermatomyositis without creatine kinase elevation may be poor. Because the prognosis of bronchiolitis obliterans organizing pneumonia is generally believed to be good, we tentatively suggest that the normal value of creatine kinase in dermatomyositis does not always seem to herald a poor prognosis, an associated malignancy or severe interstitial lung disease. PMID- 10714099 TI - Systemic mononuclear inflammatory vasculopathy associated with Sjogren's syndrome in a patient with primary biliary cirrhosis. AB - We report a 46-year-old woman with primary biliary cirrhosis (PBC) presenting with Sjogren's syndrome and systemic mononuclear inflammatory vasculopathy. Biopsy specimens of sural nerve showed findings consistent with vasculitic neuropathy. Perivascular inflammatory mononuclear cell infiltration was observed on muscle biopsy specimen. The findings of abdominal computed tomography and brain magnetic resonance imaging were suggestive of vasculitis. Clinical manifestations and radiologic findings were improved after high dose prednisolone therapy. PMID- 10714100 TI - A case of coexisting Behcet's disease and ankylosing spondylitis. AB - Behcet's disease (BD) is a chronic inflammatory condition involving several organs, such as skin, mucous membrane, eye, joint, intestine, lung and central nervous system. Ankylosing spondylitis (AS) is a prototype of seronegative spondyloarthropathy, and a chronic systemic inflammatory disorder of the axial skeleton, mainly affecting the sacroiliac joint and spine. In the latter, systemic complications may develop in addition to joint involvement. The coexistence of BD and AS has been rarely reported in the literature. The inclusion of BD among seronegative spondyloarthritides and whether sacroiliitis (SI) develops in BD are still being debated. We describe a 28-year-old man who has fulfilled the diagnostic criteria for BD and AS as well. PMID- 10714101 TI - Posterior tibial neuropathy by a Baker's cyst: case report. AB - Baker's cysts are rare cause of peripheral nerve entrapment and only a few cases of tibial nerve entrapment resulting from the popliteal cyst in the calf muscle have been reported in the literature. We present a case of rheumatoid arthritis complicated by a Baker's cyst with a tibial nerve entrapment. It is important to diagnose a Baker's cyst early and to differentiate it from thrombophlebitis, a popliteal aneurysm, tumor or muscle tear to effect optimal therapy and to obviate a potential neuropathy. Prompt recognition of these cases may save the patients unnecessary procedures and delay in treatment. PMID- 10714102 TI - Ileocecal ulcer with a cecocecal fistula in Behcet's disease. AB - We describe a case of Behcet's disease (BD) which showed the ileocecal ulcer and cecocecal fistula. This 38-year-old man had appendectomy six years ago because of colicky pain in the right lower abdomen (RLA). There are some reports on fistula formation in BD. In those, some are related to surgery and others are not. BD with cecocecal fistula, possibly associated with a past operation, has not been reported in the literature. PMID- 10714103 TI - [Pulmonary outflow tract reconstruction with autologous tissue during the Ross procedure: right ventricular characteristics in mid-term follow-up]. AB - BACKGROUND: The Ross procedure requires the interposition of prosthetic or homograft extracardiac conduits to establish ventricle-pulmonary artery connection (RV-PA). These materials usually require multiple reoperations because of conduit failure. To avoid the re-replacement of currently available conduits, usage of autologous tissue may be preferable to reconstruct RV-PA connection during the Ross procedure, especially in the pediatric age group. METHOD: Ten patients (mean age 8.7 years, range 2-23) with congenital aortic valve disease underwent the Ross procedure between June, 1996 and July, 1998. To establish RV PA continuity, autologous aortic wall including aortic valve with a gusset of pericardial tissue was used in six patients, rolled pericardial conduit with fresh pericardial bicuspid valve in three and one direct anastomosis of pulmonary posterior wall onto the right ventricle with a fresh pericardial monocusp valved patch. All patient's postoperative courses were uneventful. All patients were followed up (mean follow-up period: 21.6 +/- 6.6 months) and postoperative right ventricular characteristics, cardio-thoracic ratio (CTR) on chest X-ray and pulmonary valve function were evaluated. RESULTS: Postoperative right ventricular end-diastolic volume, right ventricular ejection fraction and right ventricular end-diastolic pressure did not change significantly (RVEDV: 128 to 113% of normal, RVEF: 56.4 to 51.5%, RVEDP: 5.9 to 10.1 mmHg). Pulmonary regurgitation during follow-up was mild in six patients and moderate in four. However, CTR decreased significantly over time (preop.: 56.5% postop.: 58.5%, late period: 53.4%). CONCLUSION: Our results support the concept of the reconstruction of pulmonary outflow tract without foreign materials during the Ross procedure. Longer follow-up are necessary to define the possible limitation of this technique. PMID- 10714104 TI - [Local recurrence in complete resection for non-small cell lung cancer]. AB - 307 patients with complete resection for non-small cell lung cancer between 1989 and 1996 were examined to evaluate the clinical features of local recurrence. Postoperative recurrence was observed in 104 of the 307 patients. At the first recurrence, local recurrences occurred in 21 (20%) of the 104 patients, distant metastases in 72 (69%), and combined recurrences in 11 (11%). 32 patients had local recurrences in 47 localizations, bronchial stump recurrences occurred in 11 (34%) of 47 localizations, hilarmediastinal lymph node metastases in 18 (56%), supraclavicular lymph node metastases in 10 (31%), contralateral mediastinal lymph node metastases in 1 (0%), and malignant effusion in 7 (22%). Lymph node metastases in the hilusmediastinum were the most common mode of the local recurrences (22%), and each mode of solely subclavian lymph node metastases and malignant effusion was 16%. The incidence of local recurrences increased as invasion into the lymphatic and/or blood vessels was demonstrated (p < 0.05). The results of our study implies that despite of lymph node dissection, latent disease persisted in the small lymphatic and/or blood vessels from hilus to subclavian sites. Postoperative adjuvant therapy including radiation will be beneficial to improvement of local control for patients invaded to lymph and/or blood vessels. PMID- 10714105 TI - [A case of tricuspid infective endocarditis in a drug addict]. AB - This report describes a successful operative case of tricuspid infective endocarditis in a drug addict. A 24-year-old man with a history of drug addiction (6 months) complained of general fatigue and high fever. Echocardiography showed a large vegetation attached to the tricuspid valve and severe tricuspid regurgitation. Blood cultures revealed septicemia due to methicillin sensitive Staphylococcus aureus. He was treated for about 1 week with intravenous antibiotics. However, subsequent severe heart failure necessitated emergency operation. The tricuspid valve was replaced with Carpentier-Edwards bioprosthesis because of severe destruction of the tricuspid valve. The postoperative course was uneventful and he has remained free from endocarditis for 15 months after surgery. PMID- 10714106 TI - [Atherosclerosis-related aortic dissection]. AB - Penetrating atherosclerotic aortic ulcers (PAU) can cause aortic dissection. Of 38 autopsy cases with aortic dissection, 6 (15.8%) had severe atherosclerotic changes, resembling those of PAU, at the site of entry (SE). Clinicopathological data on these patients were compared with those on 32 cases with nonatheromatous dissection (5 with Marfan syndrome or its forme fruste and 27 without Marfan syndrome) and 13 with atherosclerotic saccular aneurysms. For control study, the aorta of a 44-year-old woman who died of pulmonary cancer was used. Compare to nonatheromatous dissection, atherosclerosis-related aortic dissections were found in older women. Four cases were complicated by saccular aneurysms of the aorta. The SE was located in the ascending aorta in 1 and the descending aorta in 5. These sites usually were ulcerated atheromatous plaques or longitudinal fissures rather than transverse tears. Immunohistochemical examination of the SE revealed that MMP-1, 2, 9 and TIMP-2 were expressed in macrophages and/or interstitium, similar to the findings in atheromatous plaque or PAU. We propose that atherosclerosis-related aortic dissection differs from the usual classical aortic dissection. Patients with this lesion have a high risk of re-dissection from the new SE in the same lesion. PMID- 10714107 TI - [The management of infectious mediastinitis after the open heart surgery]. AB - Between October of 1992 and September of 1998, we performed 604 open heart operations. Among them, 12 cases (1.9%) were complicated with postoperative infectious mediastinitis. Five patients (Group A) were treated by conservative therapy which consists of open drainage and intermittent closed irrigation with dilute povidone iodine solution. Seven patients (Group B) were treated surgically in addition to the above-mentioned conservative treatment. Among those patients, one patient developed fatal complication. We have realized that mental care of the patients was also very important when long term hospitalization was necessitated. The hyperbaric oxygen therapy seemed to be also effective for postoperative mediastinitis caused by MRSA. PMID- 10714108 TI - [A new cardiovascular surgical instrument: cannula stabilizer]. AB - A new instrument has been developed to safely keep the venous cannula out of the operative field during open heart surgery. This instrument is designed for holding the venous cannula in a curved state at a desired angle. It consists of four semicircular claws mounted on a bar. The size of the claws is tailored so that they comfortably accommodate the standard-size two-stage venous cannula. The bar is flexible enough to be bent manually to form the required curvature, and at the same time it is strong enough to resist the self-uncoiling force of the cannula. When in use, the bar is bent to form the desired curvature and then it is placed on the required portion of the venous cannula. The angle of its curvature and its location on the cannula can be adjusted easily as required during the course of the operative procedure. With this instrument the venous cannula can be easily held in a curved state without disturbing venous return so that the cannula does not come into the way of the surgeon's hands. PMID- 10714109 TI - [Indocyanine green (TCG) clearance as a monitor to evaluate right heart function]. AB - Systemic venous return to the heart is disturbed as a result of right heart failure. ICG clearance is known to be influenced by hepatic venous return to the right atrium. Under a hypothesis that right heart function could be evaluated by ICG clearance test, patients with mitral valve disease (Group M, n = 29), aortic valve disease (Group A, n = 16), ischemic heart disease (Group CABG, n = 19) were studied. Preoperative K-ICG (normal range > 0.17) in the Group M was significantly lower than those in the Group A and Group CABG (0.097 +/- 0.037 vs 0.166 +/- 0.032 and 0.171 +/- 0.027, p < 0.05). In the Group M, patients who underwent tricuspid annuloplasty (TAP) had significantly lower K-ICG than the others (0.077 +/- 0.026 vs 0.113 +/- 0.038, p < 0.05). Postoperatively, K-ICG of TAP patients significantly increased (0.092 +/- 0.031, p < 0.05) when compared to their preoperative value. ICG clearance test was useful to quantify the right heart function, especially in the postoperative evaluation. PMID- 10714111 TI - [Surgical treatment of intracardiac tumors in 25 patients]. AB - For most intracardiac tumors, operation is the only means of therapy. In our institute, we have aggressively performed operation for intracardiac tumors regardless of histological type because resection for tumor had a beneficial effect on the hemodynamics with congestive heart failure. Twenty-five cases of cardiac tumors were operated upon from 1980 through 1998. The follow-up period ranged from 2 months to 19 years. The histological diagnoses of the tumors were as follows: benign tumors 24 (myxoma 21, papillary fibroelastoma 1, fibroma 1, angiomyolipoma 1) and malignant tumor (angiosarcoma 1). There was one hospital death in this series. In the New York Heart Association classification, the cardiac performances of intracardiac benign tumors after operation were Class I or II. The results of surgical treatment of intracardiac benign tumors were satisfactory both in short-term and in long-term. On the other hand the long term result of malignant tumor was extremely poor. A patient with angiosarcoma died 8 months later due to bone metastasis. PMID- 10714110 TI - [Could apoptosis be contributed to the occurrence of aortic dissection?]. AB - This clinical study was conducted to determine whether apoptosis is contributed to the occurrence of aortic dissection. The subjects comprised 11 patients who underwent Stanford type A aortic dissection and 4 autopsy cases, being the control group. The occurrence of apoptosis was determined by the TUNEL assay using an aortic wall specimen, and the distribution of macrophages and h-MMP-9 was examined by immunohistological staining. Apoptotic cells were observed in the aortic specimens for all of the 11 patients who underwent Standford type A aortic dissection, but not in any of the 4 autopsy cases. Moreover, apoptotic cells were present in large numbers on the surface of the false lumen strongly in 8 patients who underwent surgery within 1 month after aortic dissection, but not in 3 who underwent treatment after than 1 month. Those phenomena were also observed in the immunohistological staining of CD 68 and h-MMP-9. These findings indicate that apoptosis could be contributed to the occurrence of aortic dissection. PMID- 10714112 TI - [Coronary revascularization in a patient with bilateral internal carotid artery stenosis and aneurysm of brachiocephalic artery: a case report]. AB - A 67-year-old man with a twenty-year history of effort angina was referred to our hospital. He underwent successful PTCA for the right coronary artery and diagonal branch. However, his angina recurred three months after PTCA due to restenosis and he was recommended to undergo CABG. Because he had concomitant bilateral internal carotid artery stenosis and aneurysm of the brachiocephalic artery, we chose two-staged operation strategy to avoid cerebrovascular complication during CABG. First, he underwent bilateral carotid endarterectomy (CEA), and then he underwent concomitant CABG and grafting of brachiocephalic artery 12 days after CEA. After these operations he recovered uneventfully without neurological complication. PMID- 10714113 TI - [A case of heart operation in infective endocarditis after brain surgery for mycotic cerebral aneurysm]. AB - Complications of infective aneurysm are not rare in patients with infective endocarditis. An optimal timing of heart operation after brain surgery for hemorrhage is controversial. We reported a 19-year-old woman with ventricular septal defect (type II), mitral regurgitation and ruptured cerebral aneurysm with infective endocarditis. Cerebral aneurysm had been ruptured during infective endocarditis treatment. Resection of the aneurysm was performed next day. Vessel spasm occurred, resulting in cerebral infarction 7 days after the operation. Conservative therapy was continued for infective endocarditis until heart failure appeared. Heart operation was successfully performed 41 days after brain surgery without cerebral complication. This report indicates that heart operation might be avoided at the early postoperative stage of brain surgery for cerebral aneurysm with hemorrhage. PMID- 10714114 TI - [Malignant cardiac lymphoma]. AB - A 65-year-old male was admitted to the hospital in shock. The transesophageal echocardiography showed cardiac tumor in the right atrium. The tumor was resected under cardiopulmonary bypass. It was diagnosed as malignant lymphoma of B-cell type by histological examination. After operation, his general condition became satisfactory. Then he received chemotherapy. Twenty days after operation, however, a mass appeared in the right side of chin. It was diagnosed as malignant lymphoma of the same cell type as in the heart. We searched his body with CT and Ga-scanning. No other lesions were found. After the chemotherapy (CHOP, 6 cycles), the mass disappeared. No recurrence was been observed for six months. PMID- 10714115 TI - [A case report of a mediastinal bronchogenic cyst with back pain]. AB - We performed thoracoscopic surgery for a mediastinal bronchogenic cyst with complaining of back pain. The patient, a 38-year-old male, was admitted with an abnormal shadow on chest X-ray. Chest CT showed a localized tumor on the mediastine. MRI showed a cyst. Under thoracoscopy the tumor was based on parietal pleura and movable in chest cavity. We concluded that back pain was caused by a stimulus of a nerve in parietal pleura. Pathological diagnosis was a bronchogenic cyst. PMID- 10714116 TI - [Experience with induction chemoradiotherapy for cT4N2M0 lung cancer]. AB - The 42-year-old man with cT4N2M0 adenocarcinoma is reported. The primary tumor showed no change in size on chest X-ray film after chemoradiotherapy, but chest computed tomography revealed necrosis of the lesion and was supposed to be down staging to cT3N0M0. Right upper lobectomy and lymph node dissection (R 2) was performed. Pathological examination revealed histological CR, and he is still alive 34 months after the start of induction therapy. PMID- 10714118 TI - [Two cases of traumatic diaphragmatic hernia with atypical medical history]. AB - We have recently experienced two cases of traumatic diaphragmatic hernia which has been repaired by surgery. The first case was a 58-year-old man who had suffered left upper abdominal injury with a branch in his childhood. Although he had never symptoms, chest X-ray showed abnormal shadow in the left lower lung field. Radiologic studies indicated that the great omentum was escaped into the thoracic cavity. On patient request, we performed primary repair of the diaphragmatic hernia on thoracotomy. The second case was a 56-year-old woman who had undergone a left nephrectomy for the left renal abscess. Seven months after the operation, she began to feel nausea and vomiting, and the symptom aggravated with time. Chest X-ray showed air bubbles in the left lower lung field. It proved to be a projection of the stomach into the thoracic cavity through the iatrogenic diaphragmatic injury. We successfully performed a repairment of the diaphragm with a mesh. PMID- 10714117 TI - [A thoracoscopic resection of pulmonary metastasis from breast cancer: a case report with a 27-year disease-free interval]. AB - A 70-year-old woman, who had undergone a right radical mastectomy for breast cancer 27 years previously, was found to have a tumor measuring 15 mm in diameter between the S5 area and the S8 area of the left lung. We suspected it to be either metastatic or primary lung cancer based on preoperatively any conclusive diagnosis. We thus performed a thoracoscopic partial lung resection and pericardial resection. The pathological diagnosis of the resected specimen was pulmonary metastasis from the previous breast cancer, since the pathological findings of the lung lesion were closely similar to those of the previous breast lesion. Using immunohistochemical methods, the tumor cells show positive staining for anti-estrogen receptor antibody. In lung tumor cases in which the patient has undergone a breast cancer resection, even more than 20 years previously, surgeons must not rule out the possibility of recurrence. Thoracoscopic surgery is considered to be most effective method for both making a definitive diagnosis and for performing curative treatment. PMID- 10714119 TI - [Videoscopic surgical treatment for the patient of pleuroperitoneal communication complicating CAPD]. AB - A 60-year-old female presented in end-stage renal failure. She was introduced continuous ambulatory peritoneal dialysis in August 1997. After about six months, acute hydrothorax developed in her right side of the pleural cavity. Then she was obligated to give up CAPD because of repeated recurrence of hydrothorax. She wanted to return to CAPD for her life style. In November 1998, she came to our hospital to have a surgical treatment for her pleuroperitoneal communication. We performed video assisted thoracoscopic surgery. Using methylene blue containing dialysis solution through CAPD catheter, we found a bleb on the diaphragm that expanded gradually. We diagnosed that potion was pleuroperitoneal communication. The defect of diaphragm was directly closed with surgical stapler. Post-operative course was very favorable. She could restarted CAPD at 3 postoperative day, and discharged from our hospital after 5 days of operation. No recurrence of hydrothorax has been detected for a years after the surgical treatment. PMID- 10714120 TI - [Surgical treatment for spontaneous hemopneumothorax complicated by delayed re bleeding: a case report]. AB - A 34-year-old man was admitted to the hospital due to spontaneous hemopneumothorax. A chest tube drainage was performed, and hemorrhagic plueral effusion of 1,600 ml was drained. Because of this, the patient was transferred to the emergency center of our hospital. Following a blood transfusion, we continued to treat conservatively for nine days, because no more bleeding was recognized. On day ten, the patient suddenly started bleeding again, thus, an emergency operation was performed. At the operation under a thoracoscope, a bleeding point was ligated with surgical clip, however, it was difficult to remove blood clots that were attached with the lung surface, it was impossible to continue the thoracoscopic surgery. If re-bleeding occurs after the acute phase, problems may arise from conservative treatment. So, early surgical treatment should be considered. PMID- 10714121 TI - [Effects of restrictions on use of vancomycin in a German university hospital]. AB - BACKGROUND: Recently, increasing antibiotic resistance has been observed among gram-positive bacteria. However, only few isolates were found to be resistant against glycopeptides. Therefore, internationally accepted guidelines recommend a restricted use of vancomycin and other glycopeptide antibiotics in order to prevent the development of resistance against these clinically important antibiotics. In many countries, the hospital pharmacies play a key role in control and reinforcement of antibiotic formulary restrictions. In Germany, however, the hospital pharmacies usually do not take over such control functions, and most wards keep a stock of regularly used drugs including antibiotics, which makes reinforcement of restrictions difficult. METHODS: In an attempt to achieve a restriction of vancomycin use, the pharmacy of our university hospital was advised to deliver vancomycin to the wards only on request with a special order form signed by an attending, individually for every patient who should receive vancomycin. The efficacy of this restriction measure was evaluated in 3-month periods before and after the restriction became effective. RESULTS: Hospitalwide, this led to a 20.1% reduction of i.v. vancomycin and an 85.7% reduction of oral vancomycin use per 1000 patient days. If the hematology/oncology units were not considered, the reduction of i.v. vancomycin use was 41.8%, and the total use after the restriction 24.2 g per 1000 patient days. Microbiology results which justified the use of vancomycin decreased by 8.3% (10.9% hematology/oncology units not considered) between the 2 observation periods. Assuming a 7-day mean course of i.v. vancomycin therapy, the empirical use of i.v. vancomycin decreased from 39.9% to 8% after the restriction had been instituted. CONCLUSION: Allowing only experienced physicians (attendings) to decide on the use of vancomycin therapy, proved in our experience to be an effective measure to reduce unnecessary vancomycin use. PMID- 10714122 TI - [Health promotion and cardiovascular risk factors. The level of knowledge among 510 inpatients of an acute coronary care unit]. AB - PATIENTS AND METHODS: A total of 510 patients hospitalized on a cardiologic ward were questioned on cardiovascular disease risk factors using a questionnaire. The knowledge on these risk factors was assessed with a score system. RESULTS: Knowledge of patients on cardiovascular disease risk factors was generally low: One out of 5 did not know about the consequences of obesity, high blood cholesterol or smoking on the coronary vessels. Over 30% did not name hypertension. Only 1 out of 3 patients mentioned diabetes mellitus as a risk factor. There was no change in the knowledge during the hospital stay despite a standardized and intensive information program. The results of the second survey on the day of discharge were equal to the results of the admission day. Hospital stays in the past had no influence on the knowledge. Patients with a diagnosed coronary heart disease had the same results in the survey as patients with other diseases. The presence of risk factors had hardly any influence on the knowledge of these patients. CONCLUSION: The result of this study emphasizes the need for better health information for patients. The repetitive information on health related issues during inpatient treatment does not seem to have a positive effect on patients' knowledge. Therefore other ways of health education have to be introduced and evaluated in acute care. PMID- 10714123 TI - [Drug therapy of endocrine neoplasms. Part II: Malignant gastrinomas, insulinomas, glucagonomas, carcinoids and other tumors]. AB - BACKGROUND: The thyroid gland and the adrenal glands are the most common sites of endocrine carcinomas (see Part I of this review, Med Klin 2000;95: 20-5, Nr. 1). Less frequent are endocrine malignancies of the gastrointestinal tract (gastrinomas, insulinomas, glucagonomas, carcinoids and others). TREATMENT: Because of the rarity and missing prospective studies as well as radiotherapy and chemotherapy resistance of these tumors, generally accepted conventional therapy guidelines for these endocrine carcinomas do not exist. Surgery and radionucleotide treatment should be considered as first line therapy. Somatostatin analogs (octreotide) are frequently used as well. Chemotherapy is usually not effective. Common substances are streptozotocin, 5-fluorouracil, doxorubicin, dacarbazine and cyclophosphamide. PMID- 10714124 TI - [Current concepts in diagnosing brain death in Germany]. AB - Diagnosis of brain death requires definite evidence of an acute CNS catastrophe and exclusion of complicating medical conditions that may confound clinical assessment. Acute CNS catastrophe may be due to direct ("primary") brain damage (e.g., intracerebral hemorrhage, severe concussion, brain tumors), or indirect ("secondary") brain damage (e.g., cerebral hypoxia following cardio-pulmonary resuscitation). The cardinal findings in brain death are coma, absence of brainstem reflexes, and apnea. Persistence of these clinical signs determines brain death. In Germany, the intervals of a repeat clinical evaluation are at least 12 hours in patients with primary, and at least 72 hours in those with secondary brain damage. Electroencephalographically documented absence of electrical activity for at least 30 minutes or by means of transcranial Doppler ultrasonography or isotope angiography documented intracranial circulatory arrest also confirm brain death. Under such conditions, a repeat clinical evaluation is unnecessary in patients with clinical brain death signs. First of all, brain death is a clinical diagnosis. Confirmatory tests are not mandatory in most situations. In Germany, confirmatory tests are required in newborns, infants below the age of 2 years, and patients with infratentorial brain damage. PMID- 10714125 TI - [Delayed aortic rupture following a fall from an examination couch. Clinical and forensic aspects]. AB - BACKGROUND: Physicians frequently have to face an investigation of their work from a judicial point of view. Due to these proceedings the number of cases when they are reproached with malpractice is increasing. CASE REPORT: We report the rare case of a 76-year-old female patient who fell from a 68 cm high couch after a sedative was applied during gastroscopical examination. She died 24 hours after the incident. Forensic necropsy revealed a delayed aortic rupture as the cause of death. The preliminary proceedings against the physician involved for culpable homicide by criminal negligence were suspended after he paid a fine. The possible legal consequences under German law for physicians involved in such cases are discussed as well as strategies to avoid them. CONCLUSION: Physicians who are under investigation for malpractice should act carefully. They are advised to seek professional help at an early stage of preliminary proceedings. A public trial should be avoided if possible. PMID- 10714126 TI - [Mycobacterium avium infection presenting as a mass in the right lung]. AB - BACKGROUND: The diagnostic procedure of pulmonary masses in patients with AIDS is presented. CASE REPORT: A 39-year-old patient with AIDS was admitted to hospital because of a non-productive cough and radiologic evidence of mediastinal and right hilar masses suggestive of lymphoma associated with pneumonia of the right lower lobe. Bronchoscopy revealed a stenosis of the right lower lobe bronchus with small endobronchial lesions. Biopsies showed granulomatous inflammation, but no microorganisms were detected. Chest pain with dyspnea developed and was relieved by evacuation of pus during mediastinoscopy. The diagnosis of Mycobacterium avium infection was established via culture of sputum and bronchoalveolar lavage fluid and via mediastinoscopy. The patient was commenced on a 3-drug regimen with rifabutin, ethambutol and clarithromycin and has remained asymptomatic now for over 9 months. CONCLUSION: Mycobacterium avium infection needs to be included in the differential diagnosis of patients with AIDS presenting with mediastinal and hilar masses. When procedures such as bronchoscopy and chest CT-scans are non-diagnostic, mediastinoscopy may become necessary in order to establish the diagnosis. PMID- 10714127 TI - [Thrombotic thrombocytopenic purpura (Moschkowitz-syndrome) caused by ticlopidine]. AB - BACKGROUND: Only in a few case reports the thrombotic thrombocytopenic purpura was related to ticlopidine with a controversial discussion about this association. CASE REPORT: In a 57-year-old female patient, who was admitted with fluctuating central neurological abnormalities and generalized purpura, was made the diagnosis of a thrombotic thrombocytopenic purpura (TTP, Moschcowitz' syndrome). On admission there were a distinct anemia and thrombocytopenia. Corresponding to the hemolysis the laboratory findings showed raised reticulocytes and elevated LDH with > 900 U/l. The peripheral blood smear showed an enrichment of fragmented red cells (15%) and the bone marrow indicated a hyperplastic erythrocytopoesis and a left shift in megakaryocytopoesis. An increase of eosinophilic granulocytes and the tissue basophilic cells directed to a possible allergic phenomenon of the underlying disease. Until 3 weeks before admission she had been on ticlopidine after a left heart catheter with stenting the left coronary artery 6 weeks earlier. Beside taking of acetylsalicylacid and thyroid hormone there was no other regular medication. An early treatment with fresh frozen plasma and plasmapheresis with plasma exchange with fresh frozen plasma led directly to an elevation of thrombocytes and a normalization of hemolytic parameters. CONCLUSION: This case demonstrates the possible relationship between thrombotic thrombocytopenic purpura and the administration of ticlopidine. PMID- 10714128 TI - [Diagnosis of community-acquired pneumonia]. PMID- 10714129 TI - [Fever, myalgia, and eyelid edema]. PMID- 10714130 TI - [Physician's mission is communication]. PMID- 10714131 TI - [Meta-analysis as a tool for evaluation of evidence]. AB - In these days, more than one clinical trial is mostly performed to evaluate a new treatment or therapeutic intervention. This necessitates a combined evaluation of their results. An integration of evidence from several trials is also helpful to determine the actual knowledge. These are the main goals of meta-analyses. Since the end of the 80s meta-analyses are widely used in clinical research. At the beginning of a meta-analysis, a protocol has to be developed. Similar to a protocol of a clinical trial, the inclusion and exclusion criteria for trials, the hypotheses and the planned analyses have to be fixed. After a careful localization of trials, a combined statistical analysis is performed. An investigation of heterogeneity, i.e., differences between study results, is indispensable. During the last years, the tool meta-analysis has been criticized. The criticism mainly results from poorly conducted meta-analyses which generated results without prespecifying hypotheses or which merely combined study results. Well-planned meta-analyses, on the contrary, have an increasing influence in clinical research. PMID- 10714132 TI - When it hurts to be misled: a Stroop-like effect in a simple addition production task. AB - In four experiments, subjects saw simple addition equations (e.g., 3 + 4 = 9) and produced the sums while ignoring the presented answer. If the presented answer was false, subjects took longer to produce the sum, as compared with when the presented answer was true (Experiment 1), when there was no answer presented (blanks; Experiment 2), when a letter was presented (Experiment 3), and when a symbol was presented (Experiment 4). The results suggest that subjects were unable to ignore the presented answers, which raises problems for theories of arithmetic verification (i.e., deciding whether 3 + 4 = 9 is true or false) that claim that subjects verify equations by first producing the sum and then comparing the produced sum with the presented answer. Our results are more compatible with theories that claim that in verification and production, an arithmetic knowledge base is used in different ways. PMID- 10714133 TI - Working memory, inhibitory control, and reading disability. AB - The relationships among working memory, inhibitory control, and reading skills were studied in 966 individuals, 6-49 years old. In addition to a standardized measure of word recognition, they received a working memory (listening span) task in the standard, blocked format (three sets containing two-, three-, or four-item trials) or in a mixed format (three sets each containing two-, three-, and four item trials) to determine whether scores derived from the standard format are influenced by proactive interference. Intrusion errors were investigated in order to determine whether deficits in working memory were associated with the access, deletion, or restraint functions of inhibitory control. The results indicated that deficits in working memory were characteristic of individuals with reading disabilities at all ages. These deficits may be associated with the access and restraint functions of inhibition. Working memory skills increased until the age of 19. The blocked format showed a gradual decline in adulthood whereas the mixed format did not. The different patterns suggest that the decline in working memory skills associated with aging may result from growing inefficiencies in inhibitory control, and not diminished capacity. PMID- 10714134 TI - The phenomenology of real and illusory tip-of-the-tongue states. AB - The tip-of-the-tongue state (TOT) is the phenomenological experience that a word is on the verge of being recalled. Most research has been directed at TOT etiology and at retrieval processes occurring during a TOT. In this study, TOT phenomenology was examined. In Experiment 1, strong TOTs were more likely than weak TOTs to be followed by correct recognition, and resolution (later recall) of TOTs was higher for strong than for weak TOTs, but only for commission errors. In Experiment 2, emotional TOTs were more likely to be resolved and recognized than nonemotional TOTs. In Experiment 3, imminence was defined as the feeling that retrieval is about to occur. Imminent TOTs were more likely to be followed by resolution and recognition than were nonimminent TOTs. Illusory TOTs (TOTs for unanswerable questions) tended to be weaker, less emotional, and less imminent than TOTs for answerable questions. PMID- 10714135 TI - Interactive property attribution in concept combination. AB - We address the question of how people understand attributive noun-noun compounds. Alignment-and-comparison models suggest that the similarity of the constituent concepts guides interpretation. We propose, as an alternative, an interactive property attribution model wherein the modifier and head concepts have different functions: The head provides relevant dimensions, whereas the modifier provides candidate features for attribution. According to our model, the interaction of dimensions and features, rather than constituent similarity, guides interpretation. In this study, we empirically contrasted the two models by holding constituent similarity of compounds constant while varying the interaction of modifier feature salience and head dimension relevance. Compounds consisting of a head concept with a relevant dimension for attribution and a modifier with a salient property on that dimension were interpreted by means of property attribution. Other compounds with equivalent constituent similarity, but lacking the high salience-relevance interaction, were not interpreted by means of attribution. The interactive property attribution model more accurately predicted interpretation of noun-noun compounds. PMID- 10714136 TI - Similarity, alignment, and conceptual combination: comment on Estes and Glucksberg. PMID- 10714137 TI - Similarity and attribution in concept combination: reply to Wisniewski PMID- 10714138 TI - Tall is typical: central tendency, ideal dimensions, and graded category structure among tree experts and novices. AB - Many accounts of categorization equate goodness-of-example with central tendency for common taxonomic categories; the best examples of a category are average members--those that are most similar to most other category members. In the present study, we asked 24 tree experts and 20 novices to rate goodness-of example for a sample of 48 trees and found (1) that the internal structure of the category tree differed between novices and experts and (2) that central tendency did not determine goodness-of-example ratings for either group. For novices, familiarity determined goodness-of-example ratings. For experts, the "ideal" dimensions of height and weediness, rather than average similarity to other trees, were the primary predictors of goodness-of-example ratings for experts. The best examples of tree were not species of average height, but of extreme height. The worst examples were the weediest trees. We also found systematic differences in predictors of goodness-of-example as a function of type of expertise. We argue that the internal structure of taxonomic categories can be shaped by goal-related experience and is not necessarily a reflection of the attributional structure of the environment. Implications for models of category structure and category learning are discussed. PMID- 10714139 TI - The effects of category use on learned categories. AB - Frequently, people learn to classify instances of a concept and later learn additional information about the concept. What is the effect of this later learning on the original classification? In five experiments, this issue was investigated with a common classification paradigm in which symptom sets were classified into disease categories. After learning to classify these sets, the subjects learned to use the category to decide what treatment should be given for a symptom set. The symptoms that were important for the treatments were later classified by disease more accurately and were generated earlier from the disease name. However, this effect occurred only if the category representation was activated during the learning of the treatments. Thus, later learning about a particular use of the concept can sometimes affect the original classification. PMID- 10714140 TI - Learning categories composed of varying instances: the effect of classification, inference, and structural alignment. AB - The members of a natural category are not usually identical in their appearance, although at some level they can be described as having features in common. For example, birds have wings, but the actual appearance of their wings varies from one bird to another. To examine the effect of this feature variation on category acquisition, subjects in three experiments were asked to learn categories in which individual features were depicted with several different instances. The results of the experiments indicated that subjects had significant difficulty learning these categories when they were given a standard classification learning task. In contrast, subjects were able to acquire the same categories when they were given an inference learning task, in which they learned the categories by predicting a missing feature of a stimulus given the category label and information about the other features. Finally, subjects who were allowed to compare stimuli during learning were able to learn the categories. These results suggest that a common description of different instances emerges in the process of aligning stimuli. PMID- 10714141 TI - Not only base rates are neglected in the Engineer-Lawyer problem: an investigation of reasoners' underutilization of complementarity. AB - The standard Engineer-Lawyer problem (Kahneman & Tversky, 1973) points to the failure of reasoners to integrate mentioned base-rate information in arriving at likelihood estimates. Research in this area nevertheless has presupposed that participants respect complementarity (i.e., participants ensure that competing estimates add up to 100%). A survey of the literature leads us to doubt this pre supposition. We propose that the participants' non-normative performance on the standard Engineer-Lawyer problem reflects a reluctance to view the task probabilistically and that normative responses become more prominent as probabilistic aspects of the task do. In the present experiments, we manipulated two kinds of probabilistic cues and determined the extent to which (1) base rates were integrated and (2) the complementarity constraint was respected. In Experiment 1, six versions of an Engineer-Lawyer-type problem (that varied three levels of cue to complementarity and two base rates) were presented. The results showed that base-rate integration increased as cues to complementary did. Experiment 2 confirmed that Gigerenzer, Hell, and Blank's (1988) random-draw paradigm facilitates base-rate integration; a second measure revealed that it also prompts respect for complementarity. In Experiment 3, we replicated two of our main findings in one procedure while controlling for the potential influence of extraneous task features. We discuss approaches that describe how probabilistic cues might prompt normative responding. PMID- 10714142 TI - Individual differences in metacognition: evidence against a general metacognitive ability. AB - Individual differences in metacognitive accuracy are generally thought to reflect differences in metacognitive ability. If so, memory monitoring performance should be consistent across different meta-cognitive tasks and show high test-retest reliability. Two experiments examined these possibilities, using four common metacognitive tasks: ease of learning judgments, feeling of knowing judgments, judgments of learning, and text comprehension monitoring. Alternate-forms correlations were computed for metacognitive accuracy (with a 1-week interval between tests). Although individual differences in memory and confidence were stable across both sessions and tasks, differences in metacognitive accuracy were not. These results pose considerable practical and theoretical challenges for metacognitive researchers. PMID- 10714143 TI - How analogies are generated: the roles of structural and superficial similarity. AB - Laboratory studies of analogical reasoning have shown that subjects are mostly influenced by superficial similarity in the retrieval of source analogs. However, real-world investigations have demonstrated that people generate analogies using deep structural features. We conducted three experiments to determine why laboratory and real-world studies have yielded different results. In the first two experiments, we used a "production paradigm" in which subjects were asked to generate sources for a given target. Results show that the majority of the analogies that were generated displayed low levels of superficial similarity with the target problem. Moreover, most of the analogies were based on complex underlying structures. The third experiment used a "reception paradigm" methodology. The subjects had to retrieve predetermined sources instead of generate their own. In this case, retrieval was largely constrained by surface similarity. We conclude that people can use structural relations when given an appropriate task and that this ability has been underestimated in previous research on analogy. PMID- 10714144 TI - Spatial frequencies in short-term memory for faces: a test of three frequency dependent hypotheses. AB - Spatial frequency manipulations have been shown to have significant utility in ascertaining the various types of information that might be important in object identification and recognition tasks. This utility suggests that, given some mapping between ranges of spatial frequencies and different types of psychological information, it should be possible to examine the roles of these different types of psychological information by way of spatial frequency manipulations. One potential problem, however, is that there is no well specified, unambiguous mapping between the distinctions in the frequency domain and the distinctions in the informational domain. Three experiments provide tests of three general hypotheses regarding the ways in which different spatial frequencies might map to different information in facial perception and memory tasks: (1) the low-frequency dominance hypothesis, which proposes that low frequency information should be superior (to high-frequency information) as a cue to perception and memory; (2) the distinct informational roles hypothesis, which holds that high spatial frequencies should carry featural information while low spatial frequencies should carry information about the configuration of those features; and (3) the task-dependent information hypothesis, which suggests that high-frequency information should be best suited to discrimination tasks while low-frequency information should be best suited for recognition tasks. Results generally contradict the first two of these hypotheses, while providing support for the third. Implications with regard to the various issues related to the mapping between spatial frequencies and the informational content of faces, as well as the need to consider important interactions among perceptual and memory processes, are discussed. PMID- 10714145 TI - Exposure to print and word recognition processes. AB - The effect of exposure to print on the efficiency of phonological and orthographic word recognition processes was examined by comparing two groups of university students having similar reading comprehension scores but different levels of exposure to print. Participants with a high level of exposure to print were faster and more accurate in naming pseudowords, in choosing the correct member of a homophone pair, and in making lexical decisions when nonwords were pseudohomophones. In the lexical decision task, low-print-exposure participants were more sensitive to the frequency of the orthographic patterns in the stimuli. The results of a form priming task demonstrated that high-print-exposure participants more quickly and strongly activated the orthographic representations of common words and subsequently more strongly activated the corresponding phonological representations. Even among successful students, differences in exposure to print produce large differences in the efficiency of both orthographic and phonological word recognition processes. PMID- 10714146 TI - [Lifestyle and health]. AB - The total environments to which individuals have been exposed throughout the lifestages from birth to the present time have been composing the individual and community lifestyles. Such lifestyles are known to determine the risks for developments of cancers, circulatory diseases, and other chronic diseases. To establish new theory and practice programs for disease prevention and health promotion in the environmental and preventive medicine, we have quantitatively investigated correlations of lifestyles, or ways of daily living, to comprehensive health potentials in the cohort of industrial workers. The total lifestyles were evaluated by the originally-designed 8 health-practices such as smoking, alcohol-drinking, physical exercise, and working and sleeping patterns. The data indicate that individuals having good lifestyles showed much younger health ages calculated based on the health-check-up data, and lower risks for developing lifestyle-related diseases than those with poor lifestyles. The physical health potentials were assessed by the biomarker-measurements such as lymphocyte chromosome-DNA alterations, natural-killer activities and serum IgE levels. The psycho-mental health potentials were evaluated by both the quality-of life-related questionnaires and the stress-related hormonal and cytokine levels such as cortisol and interleukines. The comprehensive health potentials have been shown to be significantly lower in poor-lifestyle people than in good-lifestyle ones. The changes in poor to good lifestyles through health education and learning were also shown to result in promotion of such health potentials. PMID- 10714147 TI - [Effects of Ramadan fasting on the health of Muslims]. AB - The fasting month of Ramadan is the ninth lunar month of the Islamic calendar. It is the most important month for Muslims because in which the Qur'an was revealed, and they abstain from food and drink from dawn to sunset to express their gratitude to God. Eating and drinking is permitted only at night, and Muslims typically eat two meals each day, after sunset and just before dawn. People tend to stay up late watching TV with the family, praying or reading the Qur'an. Ramadan teaches Muslims self-restraint and reminds them of the feelings of the impoverished. On the other hand, the biological effects of changes in lifestyle during Ramadan may also be expected. Some studies have reported substantial weight loss, signs of dehydration, raised serum concentrations of uric acid and cholesterol, etc. during Ramadan. However, these changes are unlikely to have much effect on healthy individuals, because generations of Muslims have undertaken fasting year after year. In conclusion, the observance of the Ramadan fast may produce some ill-effects in patients with some disease, e.g. hypertension, hypercholesterolaemia, hyperuricaemia, hyperglycaemia, and heart, liver and kidney disease. PMID- 10714148 TI - [A Japanese language version of the health-promoting lifestyle profile]. AB - The development and initial psychometric evaluation of a Japanese version of the Health-Promoting Lifestyle Profile II (HPLP II) is described. The 52-item instrument was translated into Japanese and was found to be culturally relevant and reliable in a pilot study. The Japanese version was then administered to adiverse but predominantly Japanese group of 337 subjects residing in northern Japan. The Japanese version of the HPLP II was evaluated using factor analysis and reliability measurement. Six factors similar to those isolated previously during psychometric assessment of the English language version were extracted. Those six dimensions comprise the HPLP II subscales of: 1. Health responsibility, 2. Spiritual growth, 3. Physical activity, 4. Interpersonal relations, 5. Nutrition, and 6. Stress management. The alpha reliability coefficient for the total scale was 0.94 and the 2-week retest reliability was 0.91; the alpha coefficients for the subscales ranged from 0.70 to 0.87. The Japanese language version of the HPLP II appears to have sufficient validity and reliability for use by researchers who wish to describe the health-promoting components of lifestyle among the Japanese population and to explore differences and similarities in the health-promoting lifestyle of Japanese and American subjects or those of other ethnic groups. Further evaluations of measurement with different populations appears warranted. This instrument will enable researchers to investigate patterns and determinants of health-promoting lifestyle, as well as the effects of interventions to alter the lifestyle. PMID- 10714149 TI - [Effects of protein intake or exercise on 24 h urinary solute excretion]. AB - The effects of protein intake or exercise on 24 h urinary solute excretion, were evaluated in 10 female 18-19 yr of age. This study was performed during four periods: a low-protein diet (30 g x 5 days), a normal-protein diet (control, 60 g x 5 days), a high-protein diet (90 g x 5 days), and exercise loading with a normal-protein diet. (The amount of plant protein was kept constant to be 24 g/day) The following results were obtained: 1. In the case of exercise loading, urinary potassium (K) and nitrogen (N) excretions decreased significantly, while urinary sodium (Na), chlorine (Cl), calcium (Ca), and phosphate (P) excretions showed no significant differences compared with control values. 2. With the low protein diet, urinary Ca excretion decreased significantly compared with those in normal or high-protein diet. 3. The apparent fractional absorption of Na, Cl, and Ca in the female on the high-protein diet was significantly higher than that in those on the low-protein diet. These results suggest the following: 1. The amount of urinary K excretion is not only directly influenced by K intake, but also by K metabolism, such as K+ transport between extra- and intracellular spaces. 2. Although urinary Ca excretion was not increased by the increment in protein in the diet from 60 g/day to 90 g/day, it is necessary to evaluate both quantity and quality of a protein diet. 3. Protein intake of more than 60 g/day is necessary for an effective increase in Ca and NaCl absorption. PMID- 10714151 TI - [Trends in variation within the day and between days of subjective symptoms of fatigue in adolescent males]. AB - The purpose of this study was to clarify the trends in the variation within the day, and between days, of subjective symptoms of fatigue (SSF) in the daily life of male students. A SSF questionnaire (54 items) with guaranteed validity and reliability was administered to 104, 15-16 year-old healthy students, 3 times a day, in the morning (about 08:50), mid-day (about 12:10) and afternoon (about 16:10) for 5 days from Monday to Friday. As the main statistical analysis, two way (day and week) ANOVA and post-hoc t tests were used. The SSF questionnaire was considered to have very high reliability because Cronbach's alpha coefficients in each survey point of time were .967-.977. SSF complaints between days were low on the whole, but complaints of drowsiness were relatively high. The trends in variation of SSF between days were greater than those within the day. The trends in variation within the day were noticeable in complaints regarding drowsiness and loss of vigor. Complaints regarding languor became high from the middle of the week to the weekend. On the other hand, SSF complaints except for languor, were high at the beginning of the week, especially on Monday and became lower after Tuesday. There is a trend in variation within the day for symptoms regarding drowsiness and loss of vigor. The trend in variation between days was confirmed for many SSFs, and was noticeable as compared to those within the day. Complaints regarding languor were high on the weekend, and SSF complaints except for languor were high at the beginning of the week, especially on Monday. PMID- 10714150 TI - [The public health significance of the measurement of cytokines in serum]. AB - As variable functions of cytokines have been proved in recent years, cytokine levels in biological fluids such as serum, plasma, and synovial fluid of patients with every kind of disease have been enthusiastically measured. As a result, many studies have shown an increase or decrease in the production of cytokines or abnormal cytokine levels in biological fluids. However, the relationship between the abnormal levels of cytokines and the intensity of the clinical symptoms or the prognosis remains unclear. The significance for the measurement of cytokines depends on whether it should be valid for detecting a preclinical status such as AST or ALT used for health checks or for disease screening such as some tumor markers. The purpose of this study is to know whether or not some cytokine levels in serum could be biomarkers for preventive purposes. Serum cytokine levels (IL 4, 6, 8, 12, and IFN-gamma) were measured in three different types of cohorts (nursery school infants, manufacturing workers and middle and old aged women) with chemiluminescence ELISA. The results showed no differences with atopic status in infants, pulmonary fibrosis in workers or with the decrease in bone stiffness, these results are mainly due to a great inter-individual variability of serum cytokine levels. This study concludes that serum cytokine levels are inappropriate as biomarkers for preventive purposes. However, a further detailed evaluation in healthy people with high serum cytokine levels may be necessary. PMID- 10714152 TI - [A 5 year follow-up study on the relationship between changes in serum lipids and blood pressure, and changes in exercise habits in subjects 30 years of age]. AB - A 5 year follow-up study was conducted to investigate the relationships between %changes in serum lipids and blood pressure and corresponding changes in exercise habits in the middle-aged. The subjects were 152 males and 169 females who received a health check-up at 30-years-old during the period between 1990-1992 and then at 35-years-old during the period between 1995-1997. The results are as follows: 1. In multiple regression analysis, % delta toriglyceroid (TG) and % delta AI(atherogenic index) ratio were associated significantly with changes in exercise habits in males (P < 0.05) and % delta high density lipoprotein cholesterol (HDLC) was associated with changes in exercise habits in females (P < 0.05). % delta TG and % delta HDLC were affected by changes in exercise habits adjusted for BMI, smoking, and drinking. But changes in TC and blood pressure were not affected by changes in exercise habits. 2. In males, % delta TG was significantly lower in those subjects who started their exercise habit than in the no exercise group and than in those within the ceased exercise group who ceased their exercise during the study period (P < 0.05). % delta AI was significantly lower in the started exercise group than in the no exercise group. Also, the continued exercise group had a significantly (P < 0.05) lower % delta AI as compared to the ceased exercise group. In females, % delta TG was significantly lower in the started exercise group than in the no exercise group (P < 0.05). % delta HDLC was higher significantly in the continued exercise group than in the ceased exercise group (P < 0.05). From the results obtained, it is recommended that the no exercise subjects should have exercise more than once time per week and those with more than 1 time per week maintain their exercise habits. PMID- 10714153 TI - [Sjogren's syndrome--recent advances in research and treatment]. PMID- 10714154 TI - [Antiviral effect of (S)-1-(3-hydroxy-2-phosphonylmethoxypropyl) cytosine on adenovirus]. AB - PURPOSE: Adenovirus is the most frequent causative virus of conjunctivitis in Japan. Recently (S)-1-(3-hydroxy-2-phosphonylmethoxypropyl) cytosine (HPMPC) has been promoted as a new drug against adenoviral conjunctivitis. So we examined the antiviral activity of HPMPC against adenoviruses in vitro. METHOD: The antiviral activity of HPMPC against adenovirus (Ad) type 3, type 4, type 19, and type 37 isolated from conjunctivial scrapings in Japan and the prototype of adenovirus type 5 was examined by plaque reduction assay using A 549 cells in vitro. RESULTS: The 50% inhibitory dose (ID50) of HP-MPC was 3.50 (1.44-4.79) micrograms/ml for Ad type 3, 4.50 (4.17-4.92) micrograms/ml for Ad type 4, 2.11 (1.03-3.13) micrograms/ml for Ad type 5, 1.64 (1.40-2.02) micrograms/ml for Ad type 19, and 2.02 (1.17-2.73) micrograms/ml for type 37. The 50% cytotoxic dose of HPMPC for A 549 cells was 205 micrograms/ml by the deoxythimidine uptake inhibition test, and 537 micrograms/ml by the trypan blue exclusion inhibition test. CONCLUSIONS: HPMPC proved to be highly effective in inhibiting replication of adenoviruses at lower concentrations than the cytotoxic level in vitro. PMID- 10714155 TI - [Mitomycin C dissolved in a reversible thermo-setting gel: effect on aqueous flare in the rabbit eye]. AB - PURPOSE: To determine the effects of mitomycin C (MMC) dissolved in a gel, which is as effective as a higher dose of aqueous MMC solution, on aqueous flare in rabbits. METHOD: We injected subconjunctivally a 0.1 ml solution of MMC containing 30, 10, 3.0, or 0.3 micrograms of MMC dissolved in a reversible thermo setting gel in pigmented rabbits. As a control, a 0.1 ml aqueous solution of 30 micrograms MMC was injected subconjunctivally. Aqueous flare was measured for 28 days after the injection. RESULTS: The aqueous flare in the 30 micrograms gel group was also level similar to that in the 30 micrograms aqueous solution group, but the aqueous flare in the 10, 3.0, and 0.3 micrograms gel groups was significantly lower. The increase in the aqueous flare following the administration of MMC gel was dose-dependent. CONCLUSION: The administration of MMC dissolved in a gel can diminish the ocular inflammation by reducing the dose of MMC needed for the antiproliferative effect. PMID- 10714157 TI - [Pediatric cataract surgery with posterior capsulorrhexis and optic capture of the intraocular lens]. AB - We evaluated the result of the pediatric cataract surgery with optic capture. The intraocular lens (IOL) optic is subluxated to the vitreous body side through the posterior continuous curvilinear capsulorrhexis (PCCC). Five eyes with congenital or developmental cataracts were treated with phacoemulsification and IOL implantation, and then PCCC and optic capture was done. Three of the cases were bilateral and one case was unilateral. All cases have remained in good condition without secondary fibrous membrane formation, which influences visual acuity, and transparency of the posterior capsule was maintained in the visual axis area. We found one case of anterior capsule contraction. Long-term follow up should be continued. PMID- 10714156 TI - [Effects of corticosteroid on porcine retinal pigment epithelial cells in culture -2. Effects on phagocytosis and lysosomal activity]. AB - PURPOSE: The effects of a corticosteoid on the phagocytosis and lysosomal activity of cultured porcine retinal pigment epithelium (RPE) cells were investigated. METHODS: After exposing cultured RPE cells to various concentrations (10, 50, 100, 500, 1,000 nM) of betamethasone sodium phosphate (betamethasone), the cells were incubated with latex microspheres for 6 hours. RESULTS: The number of latex microspheres phagocytized by the cultured RPE cells was inhibited by 50 nM betamethasone within 24 hours. Ten-nM betamethasone did not inhibit the proliferation of cultured RPE cells, but ingestion of latex microspheres by the cells was inhibited after 3 days. Lysosomal activity (acid phosphatase, beta-glucuronidase) of RPE cells was inhibited by a high concentration (500 nM) of betametasone. CONCLUSION: These results suggest that corticosteroid inhibits the phagocytosis and lysosomal activity of cultured RPE cells. PMID- 10714158 TI - [Uveitis associated with zoster sine herpete. Diagnosis and clinical findings]. AB - PURPOSE: Zoster sine herpete (ZSH) causes solely neurologic symptoms without the eruption that is evident in herpes zoster ophthalmicus. It is occasionally complicated by acute granulomatous uveitis. We examined patients suspected of ZSH for detection of the varicella-zoster virus (VZV) genome, and discussed its clinical appearance. MATERIALS AND METHODS: Nine patients were presumed to have ZSH. All manifested acute granulomatous iridocyclitis and high intraocular pressure without eruption. A polymerase chain reaction (PCR) analysis of the aqueous humor was performed. RESULTS: Five patients were positive for VZV DNA. They showed mutton-fat keratic precipitates and high intraocular pressure in the early stage. Pigmentation in the anterior chamber angle, pigmented keratic precipitates, and finally sectoral iris atrophy were observed in the recovery stage. These clinical findings were common to ZSH. CONCLUSIONS: The ocular lesions in ZSH were shown to have distinctive characteristics, and PCR is useful to determine etiological agents in the early stage of disease. PMID- 10714159 TI - [Studies on vitrectomy cases associated with complicated branch retinal vein occlusion]. AB - PURPOSE: To examine the factors affecting visual outcome in vitrectomized cases associated with complicated branch retinal vein occlusion (BRVO). MATERIALS AND METHODS: Of 114 eyes of 113 patients with BRVO, ages, general complications, distribution of occluding vessels, location of retinal breaks, classification of vitreoretinal pathology and number of cases, period from onset of BRVO to vitreous hemorrhage and from vitreous hemorrhage to vitrectomy, number of operations, relationship between posterior vitreous detachment (PVD) and number of operations, post and preoperative pholocoagulation status, pre and postoperative visual acuity, and cases with poor visual outcome were analyzed. RESULTS: The visual prognosis was much better in the cases with vitreous hemorrhages only than those with proliferative membrane and retinal detachment (p = 0.0029). Repeated operations were needed in the cases of incomplete PVD (p = 0.0023). CONCLUSIONS: Early vitrectomy, especially in the cases of incomplete PVD, seems to be essential for management and treatment for better visual acuity. PMID- 10714160 TI - [Comparison of primary and secondary Sjogren's syndrome]. AB - PURPOSE: We studied the difference in severity between primary and secondary Sjogren's syndrome (SS). SUBJECTS AND METHODS: Two groups of patients (all females, mean age: 58 years), 31 with primary SS and 18 with secondary SS were studied. We performed the following dry eye tests: fluorescein score and Rose Bengal staining, grading of tear lipid layer interference patterns, measurement of fluorescein break up time, cotton thread test, and Schirmer-I test. Auto antibodies were also investigated. RESULTS: There was no significant difference between primary and secondary SS with respect to any dry eye tests or auto antibodies. In primary SS, however, the presence of anti SS-A antibody was significantly correlated with Rose Bengal scores (p = 0.044). CONCLUSION: The severity of SS is independent of the primary or secondary type. In primary SS, the presence of anti SS-A antibody may be correlated with the severity. PMID- 10714161 TI - [A case of acute diffuse atrophy of retinal pigment epithelium]. AB - PURPOSE: To report a case of acute bilateral visual disturbance where the ocular fundus changed like retinitis pigmentosa in a short time. PATIENT: 26-year-old man. FINDINGS: In initial examination, the patient's visual acuity was 0.01 OD and 0.02 OS with myopic correction, but his fundus did not look abnormal. Fluorescein angiogram showed marked background hyperfluorescence and dye leaking to the vitreous. After 2-3 weeks, the fundus appearance changed like retinitis pigmentosa. Best corrected visual acuity became 1.0 OD and 0.9 OS after steroid pulse therapy. We were unable to find the cause of this disease in spite of blood tests and other examinations. CONCLUSION: This case with acute diffuse atrophy of retinal pigment epithelium and damage of the function of blood-retinal barrier was considered remarkably rare. PMID- 10714162 TI - [A case of ring 14 chromosome with ocular manifestations]. AB - BACKGROUND: Ring 14 chromosome has been reported to be associated with mental retardation, craniofacial dysmorphology, and epilepsy. Flecked and/or pigmented retina are also ocular manifestations of this disease. CASE: A 29-year-old female suffered from seizures and developmental and growth delay. Narrow palpebral fissura, broad flat nose, large auricula, high arched palate, and short neck were present. Chromosomal analysis disclosed her ring 14 chromosome (p 11.2 q 32.3). Ophthalmologically, cortical cataract, refractive error (right--3.00 D, left- 1.50 D), and yellow-white flecks in the macula and yellow-white spots in the mid peripheral retina in both eyes were present. CONCLUSIONS: To date, ophthalmic changes concomitant to a breakpoint at 14 q 32.2 have been reported. We report a case of ring 14 chromosome with breakpoint at 14 q 32.3 which showed yellow flecks in the macula and mid-peripheral retina. PMID- 10714163 TI - [Politics in science]. PMID- 10714164 TI - [Signal transduction in Fas-mediated apoptosis]. PMID- 10714165 TI - [Serralysin Zn-metalloproteinases--structure, function, secretion pathway, and pathogenicity]. PMID- 10714166 TI - [DNA-dependent protein kinase]. PMID- 10714167 TI - [Functional implication of BH3-only proteins (BOP) in apoptosis]. PMID- 10714168 TI - [Intrinsic nucleoside diphosphate kinase-type activity is a novel function of the molecular chaperone and protease]. PMID- 10714169 TI - [Physiological role of mucin-type sugar chain on hinge portion of human serum IgA1]. PMID- 10714170 TI - [Structure-function relationship of gastric proton pump]. PMID- 10714171 TI - [Progress of adenovirus vector development]. PMID- 10714172 TI - [Glucose signaling in Escherichia coli: role of PTS phospho-relay system]. PMID- 10714173 TI - [Development and evolutionary origin of vertebrate limb]. PMID- 10714174 TI - [Biosynthesis of heparan sulfate and the tumor suppressor EXT gene family]. PMID- 10714175 TI - [Excited states and electron transfer in the photosynthetic reaction center of Rhodopseudomonas viridis: SAC-CI study]. PMID- 10714176 TI - [Crystal structure of Z-DNA binding domain bound to left-handed Z-DNA: outstanding achievement of Alexander Rich]. PMID- 10714177 TI - [Application of specific and strong biotin-avidin binding for cell technology]. PMID- 10714178 TI - [Transgenic plants as a medicine production system]. PMID- 10714179 TI - [Neuronics: application of micro-fabrication technique to neuroscience]. PMID- 10714181 TI - [In Process Citation] PMID- 10714180 TI - [Attempts to prepare aseptic and non-salt miso by the use of bacteriocins produced by lactic acid bacteria]. PMID- 10714182 TI - Tennessee's mental health services in crisis. Strained system undergoes comprehensive review. PMID- 10714183 TI - Loss prevention case of the month. Physician + nurse = expensive clash. PMID- 10714184 TI - Health policy report. What is health services research--and why? PMID- 10714185 TI - Autoimmune hepatitis--a diagnostic challenge. AB - Autoimmune hepatitis is a type of chronic hepatitis characterized by hypergammaglobulinemia, hypertransaminasemia, presence of autoantibodies, and active necroinflammatory process in the liver revealed by histology. Its onset is usually acute and has a bad prognosis. Tissue antibodies are found in large proportions of patients. Women outnumber men in a 2-3:1 ratio, and it is mostly young women who are affected. Ten percent of all affected have severe disease characterized by elevations of serum aminotransferase levels greater than five fold, along with a two-fold elevation of gamma globulin or a 10-fold elevation of serum aminotransferase levels alone. We present an unusual case of this disorder in a 60-year-old male patient manifesting its severe form. PMID- 10714186 TI - Cicatricial pemphigoid with an upper airway lesion. AB - Cicatricial pemphigoid is an unusual mucocutaneous disease that is characterized by subepidermal blister formation involving the oral and conjunctival membranes. The oral lesions are expressed as erythema and induration and have rarely been associated with upper airway obstruction. We report the case of a patient with dyspnea and an abnormal flow-volume loop who was found to have subglottic compromise due to cicatricial pemphigoid. Immunosuppressive therapy improved his symptoms and air flow. PMID- 10714187 TI - Vanderbilt Morning Report. A young woman with gastrointestinal bleeding. PMID- 10714188 TI - Department of Health Report. Tobacco control and prevention in Tennessee. PMID- 10714189 TI - [Pathology in Jena]. PMID- 10714190 TI - [Rudolf-Virchow Prize 1999 of the German Pathology Society]. PMID- 10714191 TI - [Biopsy differential diagnosis of gastroesophageal reflux (GERD)]. AB - In biopsy interpretation of GERD demonstration or exclusion of Barrett's esophagus and its complications (dysplasia and carcinoma) is the main task, especially in its differentiation to cardiac gastritis with intestinal metaplasia. Despite of all molecular-biological examinations till today no biomarker exists for the malignant potential of Barrett's esophagus, therefore regular surveillance is necessary. Differentiation of Barrett's esophagus from cardiac mucosa with intestinal metaplasia is not yet possible by using immunochemistry, also there is no marker for the differential diagnosis of low grade dysplasia versus reactive atypia and high grade dysplasia versus early cancer of mucosa type. PMID- 10714192 TI - Pathology of the gastric cardia. AB - Adenocarcinomas of the cardia and distal esophagus have increased in incidence more rapidly than any other type of human cancer in recent years. Whereas the sequence of events leading to esophageal adenocarcinoma (esophagitis, Barrett's esophagus, dysplasia, and carcinoma) has been well described, that of cardiac adenocarcinoma has yet to be defined. We have examined 100 consecutive biopsies of the gastroesophageal (GE) junction in patients with symptoms of GERD, in order to answer the following questions: (1) What is the spectrum of cardiac pathology? (2) Can "carditis" due to GERD be histologically distinguished from carditis due to Helicobacter pylori (HP) infection? (3) Is intestinal metaplasia of the cardia more frequently related to GERD, or to HP infection? (4) What types of esophageal and gastric pathology coexist with carditis? Out of the 100 GE junction biopsies only 63 contained cardiac mucosa and all showed carditis. Carditis was related to HP in 8 cases. Distinguishing histologic features of non-HP carditis included the predominance of reactive epithelial changes over inflammatory changes, a villiform surface and a tendency of the inflammatory infiltrate to decrease with increasing distance from the squamo-columnar junction. Pancreatic metaplasia was seen in 9 cases (14%), none associated with HP. Intestinal metaplasia (IM) was seen in 15 cases (24%) and associated with HP in only 2. Non-HP carditis, with or without IM, was associated with normal or reactive esophageal squamous and distal gastric mucosa in the majority of cases. We conclude that, in our patient population, carditis and cardiac IM is primarily due to GERD and propose that the sequence of events leading to cardiac carcinoma is similar to that of esophageal adenocarcinoma. PMID- 10714193 TI - [Histological and molecular prognostic factors in esophageal cancer]. AB - The prognostic impact of new histological and molecular parameters was tested retrospectively in a series of 149 patients with esophageal squamous cell carcinoma (SCC) who underwent potentially curative resection therapy (no distant metastases, no residual tumor, no radio- or chemotherapy). This analysis was performed in order to identify patients with increased risk for tumor-related death in spite of being treated by standard therapy and thus being candidates that most likely profit from postoperative adjuvant therapy. Additionally, the prognostic value of various molecular markers was investigated in a group of 38 patients with locally advanced esophageal SCC that have been treated with combined therapy modalities (radiochemotherapy and optionally surgery). Among surgically treated carcinomas, the following morphological parameters proved to be prognostically significant in univariate survival analysis and multivariate survival analysis: pattern of invasion, inflammatory response and lymphatic vessel invasion. In contrast, the tumor grading according to the criteria of WHO and tumor cell proliferation did not show significant prognostic impact. Concerning the prognostic influence of molecular parameters strong expression of the proliferation-regulating molecule p21WAF1 and weak expression of the apoptosis-regulating molecule Bcl-XL were predictors of poor survival in univariate and multivariate survival analysis. No prognostic impact could be shown in relation to the expression of the proliferation-regulating molecule p53 and the apoptosis regulating-molecules Bcl-2 and Bax. Among multimodally treated esophageal cancer patients, again strong expression of p21WAF1 as well as accumulation of p53 were predictors of poor survival, whereas expression of Bcl 2, Bax and Bcl-XL did not show any prognostic influence. In conclusion, morphological and molecular parameters may provide important prognostic information for esophageal cancer patients. PMID- 10714194 TI - Classification of gastritis--yesterday, today and tomorrow. AB - The major landmark in the recent history of gastritis was the discovery of Helicobacter pylori as the cause for approximately 90% of cases of chronic gastritis. This was followed by an attempt to rationalise classification from the many conflicting nomenclatures in existence to one, the Sydney System, that might gain more universal acceptance and allow studies from around the world to be compared. In the decade since its inception, including an appropriate update, the system has partially achieved this aim. It is based on documenting the topography of the gastritis, this reflecting the range of possible clinical outcomes and point of progress along the gastritis-metaplasia-dysplasia-carcinoma cascade. The rapidly expanding molecular knowledge about host mucosal immunology, determinants of bacterial virulence and dietary constituents makes it likely "tomorrow's" classification will be an algorithm to include several such factors. From this an exact "peptic" patient profile could be constructed. However one must also speculate that interest in gastritis and its classification could whither just as quickly as it blossomed with the advent of a successful vaccine. PMID- 10714195 TI - [Helicobacter pylori infections and autoimmunity: the interplay in the pathogenesis of gastritis]. AB - Recent studies suggest a significant association of Helicobacter pylori (H.p.) gastritis and antigastric autoimmunity. Sera of H.p. infected patients exhibit antigastric autoantibodies with high prevalence (over 50 percent of the cases) and with antiluminal and/or anticanalicular binding patterns on gastric mucosa. Models for the pathogenesis of this H.p. associated autoimmunity are antigenic mimicry or Th 1-induced expression of MHC class II and costimulatory molecules on gastric epithelial cells. The anticanalicular autoantibodies binding to gastric parietal cells show fine specificities for the alpha- and for the beta-subunit of the gastric H, K-ATPase, which correspond to the findings in classical autoimmune gastritis. They are significantly correlated with higher grades of corpus gastritis as well as morphologic and functional glandular atrophy of the corpus. The development of this antigastric autoimmunity apparently is a relevant pathogenic factor of the host reaction and might be crucial for the different types and outcomes of H.p. gastritis. PMID- 10714196 TI - Well-differentiated adenocarcinoma or dysplasia of the gastric epithelium: rationale for a new classification system. AB - BACKGROUND: Gastric neoplastic lesions labelled as high-grade adenoma/dysplasia by Western pathologists are often diagnosed as mucosal carcinoma by Japanese pathologists. We examined whether stratifying histological diagnoses by invasion status could increase the extent of agreement between Western and Japanese pathologists and could reduce the frequency of discrepant diagnoses between biopsy samples and corresponding resected specimens. METHODS: Thirty-five histological slides of gastric lesions that had previously been individually reviewed by eight expert gastrointestinal pathologists from Japan, North America and Europe were reassessed by the same pathologists with particular attention to the aspect of invasion. Kappa statistics were used to determine the extent of agreement between Western and Japanese pathologists before and after reclassifying the diagnoses according to invasion status. Moreover, we examined the number of discrepant assessments regarding 14 lesions of which there were both biopsy specimens and resected specimens. RESULTS: There was agreement between the Western and Japanese pathologists in only 11 (31%) of the 35 slides (kappa coefficient 0.15) when traditional diagnostic categories were used. However, after high-grade adenoma/dysplasia, noninvasive carcinoma and suspected carcinoma were grouped together, there was better agreement, namely in 22 (63%) of the slides (kappa coefficient 0.41). Moreover, such reclassification significantly reduced the number of discrepant diagnoses between biopsy and endoscopic mucosal resection specimens by three Western pathologists, from 19 (45%) of 42 assessments when high-grade adenoma/dysplasia was grouped together with low-grade adenoma/dysplasia, to 6 (14%) of 42 assessments when grouped with suspected and definite carcinoma. CONCLUSIONS: To improve the international comparability of gastric histological diagnoses we recommend that high-grade adenoma/dysplasia, noninvasive carcinoma and suspicion of invasive carcinoma be grouped together as a single category of noninvasive high-grade epithelial neoplasia. PMID- 10714197 TI - [Correlation between histological and molecular mechanisms of carcinogenesis in stomach cancer]. AB - Since gastric cancer is an exceptional heterogeneous tumor conflicting results have been obtained about the relationship between genotype and phenotype. From the molecular point of view gastric carcinoma diffuse type forms a distinct entity which is microsatellite stable, has almost no p53 mutations and exhibits in at least half of the cases mutations in the E-cadherin gene. In contrast, all other gastric carcinomas comprise a heterogeneous group of which about one third exhibits microsatellite instability (MSI) but no p53 protein stabilization or gene mutations. These tumors are either of pure intestinal (glandular) type or show large solid (medullary) tumor cell clusters. Thereby, in sporadic gastric cancer MSI is caused by loss of hMLH1 expression due to hypermethylation of the promotor region rather than by mutation of the gene itself. Tumors that are microsatellite stable (MSS) and show p53 alterations are either intestinal (about 70%) or a mixed-type encompassing at least 5% glandular and poorly differentiated diffuse components (about 30%). Whereas pure diffuse type gastric cancer is unlikely to develop from intestinal type carcinoma, this may, however, be the case in some advanced mixed-type gastric cancers. PMID- 10714198 TI - [MALT-type non-Hodgkin's lymphoma: advances in biopsy diagnosis and pathogenesis]. AB - Malignant Non-Hodgkin's lymphomas of the mucosa-associated lymphoid tissue display unique morphological and biological features. Unlike the situation in nodal lymphomas, they almost invariably arise from defined and recognisable precursor lesions, such as chronic infections or autoimmune diseases. From the diagnostic point of view, efforts have been undertaken to define (1) the minimal requirements for the unequivocal diagnosis of MALT-type lymphoma, (2) the regression stages after Helicobacter pylori eradication, and (3) to establish a grading system for the respective lesions. Of importance, it must be emphasized that in most cases the biopsy received in first-line is not enough in establishing a correct diagnosis with regard to biological and therapeutic implications. In particular, the accurate estimation of the biological aggressiveness of the tumour will in most cases require extensive rebiopsies. As in nodal tumours, the genetic constitution of low grade MALT-type lymphomas is more and more revealed. While data on the importance of numerical aberrations are conflicting, certain structural aberrations, such as the t(1;14) and the t(11;18), have been unequivocally linked to MALT-type lymphomas. Preliminary data suggest that these aberrations may target the apoptotic pathway. Their recognition may herald the evolution of new tumour features, such as independence from outer regulatory influences, obviously in early stages determined by the microenvironment and might be closely related to the site of tumour development and hence be a feature of the primary inflammatory disorder. PMID- 10714199 TI - [Problems in biopsy differential diagnosis in lymphomas of the small and large intestines]. AB - Besides the problems inherent in endoscopically obtained tissue and the low incidence of intestinal lymphomas, the major difficulties reside in the distinction to reactive processes and in the differential diagnosis among several lymphoma entities. Knowledge of the microanatomical and biological properties of the intestinal MALT, supplemented by sufficient clinical information, are important prerequisites for the diagnostic work-up which has to include immunohistochemical studies. Whereas the diagnosis of aggressive B-cell lymphomas (diffuse large B-cell lymphoma, Burkitt's lymphoma) is usually straightforward, lymphomatous polyposis (LP, the intestinal equivalent of mantle cell lymphoma) and low-grade B-cell lymphoma of MALT-type may be difficult to diagnose and to separate from reactive lymphoproliferations. The characteristic immunohistochemical profile of LP (cyclin D1 + CD5 + CD43 + CD23 - CD10 - IgM kappa or lambda, is very helpful in this regard and similarly useful to exclude intestinal involvement by B-CLL or follicular center lymphoma. In addition, the endoscopic appearance characterized by seeds of small polyps along the colorectum favors LP although MALT-type lymphoma may occasionally produce polypoid lesions. Focal lymphoid hyperplasia occurs in the terminal ileum and may present with a mass in the right iliac fossa. The diagnosis of intestinal T-cell lymphoma (ITL) represents the most challenging task for both clinicians and pathologists. This disease is often associated with and may closely mimick celiac disease of adult onset type, or can be misdiagnosed as inflammatory bowel disease. The presence of an abnormal activated T-cell phenotype, i.e. different from that of normal intraepithelial lymphocytes, strongly suggests ITL and is of particular importance in cases that lack overt cytological atypia. PMID- 10714200 TI - [Biopsy differential diagnosis of non-cancerous small intestinal diseases]. AB - Both pathologists and gastroenterologists are one team working up patients with suspected small bowel diseases. Both have to try to recognize the various entities of disorders found in the small bowel. Some diseases, such as Whipple's disease, are histologically easily recognized by their specific changes. Others, however, show unspecific histological changes and may only be recognized by a certain pattern of inflammatory infiltration of the mucosa and/or by architectural changes. In the present paper giardiasis with its diagnostic trophozoites, Crohn's disease and celiac disease with their typical inflammation and mucosal architecture and their differential diagnoses are discussed. A modified Marsh classification is given, which allows to report the various types of celiac lesions more precisely. Furthermore, the variations found in the normal mucosa are presented. In particular, the presence of lymphocytes and plasma cells in the lamina propria is shown to be a part of the mucosa associated lymphatic tissue (MALT). The importance of giving a clear statement on the etiology of the disease whenever possible is stressed. PMID- 10714201 TI - [Differential diagnosis of colitis]. AB - The aims of the bioptical diagnosis of colitis are delineated in introductory remarks. Subsequently, the range of normality is described for mucosal structures, and the term "nonspecific chronic colitis" is critically discussed. Furthermore, some characteristic (diagnostic) forms of colitis are presented together with their differential diagnoses. Also the "less" diagnostic features of infectious colitis and chronic inflammatory bowel disease are described. An algorithmus of the differential diagnosis of colitis is proposed. Excluding 83 cases of normal findings, the practical impact of such an algorithm was critically analysed in 381 consecutive biopsies of histologically proven colitis. -Several "characteristical" types of colitis could be distinguished in about 95% of the biopsies; in a small percentage of cases only a differential diagnosis could be offered, partly because of inconclusive morphological findings, partly because clinical data were completely lacking and partly because there were no separate rectal biopsies in these cases. We wish to stress, however, that the anamnestical data relating to the length of complaints and the therapeutic interventions are very relevant to the interpretation of the morphological differential diagnosis of chronic colitis, especially in chronic colitis with low activity or inactive disease. PMID- 10714202 TI - [Dysplasia in ulcerative colitis]. AB - Due to their increased risk of developing a carcinoma, patients with long standing ulcerative colitis are usually enrolled in surveillance programs in order to regularly evaluate their risk of developing cancer using endoscopic biopsies. In this evaluation, the histologic identification of dysplasia in connection with the endoscopic findings is of central importance. According to current recommendations, dysplasia which occurs in endoscopically identifiable lesions, i.e. in the form of a DALM ("dysplasia associated lesion or mass"), result in a colectomy independent of the degree of dysplasia, as does high grade dysplasia in a flat mucosal area. For low grade flat dysplasia, the diagnosis should be confirmed. A special diagnostic challenge is the distinction of adenomas in ulcerative colitis, since for these lesions a simple polypectomy is an adequate treatment. For the assessment of the individual cancer risk various molecular biologic techniques have been discussed as additional procedures (p53, Sialyl-Tn, ploidy status by flow cytometry), but H&E histopathology is still the gold standard for the grading of dysplasia and its distinction from regenerative epithelial changes. Due to the increasing refinement of endoscopic techniques and considering the potential new, less invasive methods for treating dysplasia, changes in the management approach for patients with dysplasia in ulcerative colitis are to be expected. PMID- 10714203 TI - [Molecular carcinogenesis in ulcerative colitis-associated and sporadic colorectal carcinoma--differences and similarities]. AB - Like sporadic colorectal cancers, ulcerative colitis (UC)-related cancers are thought to evolve through a multistep progression pathway. The genomic alterations important in sporadic colorectal carcinogenesis are well characterized, with loss of APC function being a frequent and early event. However, the genomic alterations in UC-related carcinogenesis are yet unclear and the role of APC is controversial. In this study genomic alterations in UC-related cancers, dysplasias and nondysplasias were assessed by comparative genomic hybridization (CGH). Alterations of the APC/beta-catenin pathway were evaluated by immunohistochemistry. 32 cases of UC-related cancers (14 with synchronous dysplasias and nondysplasias) and 42 sporadic cancers were matched by UICC stage. CGH was performed using DOP-PCR amplification after microdissection. Expression of beta-catenin, E-cadherin and APC were detected by immunohistochemistry in paraffin sections. Chromosomal alterations were present in 90% of the sporadic and 94% of the UC-related cancers. 86% of the UC-related dysplasias and 36% of the nondysplasias showed changes by CGH. Chromosome 5q was lost in 56% of UC related cancers and 36% of the dysplasias but in only 26% of the sporadic cancers. Other frequent alterations in both cancer groups were loss of 18q, 8p, 17p, and gain of 8q and 20q. Immunohistochemistry showed a decrease of membranous and an increase of cytoplasmic expression of E-cadherin and beta-catenin in UC related and sporadic cancers. APC expression was significantly decreased in both tumor types. Clonal chromosomal alterations occur early in UC-related tumor progression. UC-related and sporadic cancers share a set of common clonal abnormalities. The frequent loss of 5q and the altered expression of APC, beta catenin, and E-cadherin proteins in UC-related cancers indicate a critical role of the APC/beta-catenin pathway in UC-related carcinogenesis. PMID- 10714204 TI - [Are there differences between ex adenoma and de novo colorectal carcinomas?]. AB - Colorectal carcinoma is a major cause of death throughout the Western world. It is increasingly recognized that any reduction in mortality must be achieved through the detection and removal of early and precancerous lesions. The primary attention for such a preventive strategy has been the polypoid adenoma and surveillance studies have shown a significant reduction in the incidence of carcinoma through systematic polypectomy of suspicious lesions. A potential problem with such a program, however, is raised by reports from Japan that some carcinomas seem to arise without a precursor polypoid adenoma, that is de novo. Although the histopathologic findings in such reports seem to clearly support this idea, this concept is not widely accepted in the Western world. We undertook a series of immunohistochemical (p53, bcl-2, Mib-1, E-cadherin, CD44, Stromelysin 3), and microsatellite analysis studies (on 17p (p53), 18q (DCC), 5q (APC), 8p, 2p and 1p), on groups of de novo and ex adenoma carcinomas in order to see if differences between the two groups of lesions exist. The results of these studies demonstrate that de novo carcinomas share several phenotypic and genotypic features with ex adenoma carcinoma (similar CD44 in the carcinomas, similar rates of LOH at APC and DCC loci), but have significantly higher rates of LOH at 17p, p53 over-expression and ST-3 expression indicating that tumor progression in de novo carcinoma is accelerated. These findings should help clarify the concept of de novo carcinoma and contribute to wider recognition of this important clinicopathologic entity. PMID- 10714205 TI - [Diffuse stomach carcinoma: from H&E diagnosis and molecular pathology to specific therapy]. AB - E-cadherin is a homophilic cell adhesion molecule that is mutated in half of diffuse-type gastric cancer patients. Since these mutations generally affect the extracellular portion of the transmembrane molecule and do not interrupt the reading frame, altered E-cadherin protein may be an excellent tumor marker. We established a rapid PCR-based E-cadherin mutation detection technique allowing positive results within a day. Furthermore, we succeeded in the generation of monoclonal antibodies that specifically react with mutant E-cadherin but not with the wild-type protein. In gastric carcinoma specimens known to express mutant E cadherin messenger RNA these monoclonal antibodies target exclusively tumor cells in routine formalin fixed and paraffin embedded material from biopsies, primary tumors and lymph node metastases. Non-tumorous cells, including normal gastric epithelium expressing wild-type E-cadherin, are not stained. The unique type of E cadherin mutations in diffuse-type gastric cancer (missense mutations and complete or partial in-frame deletions of exons) might improve the information of conventional diagnostic techniques. In addition, they may open novel and more selective clinical avenues to treat small tumor deposits for adjuvant-, neoadjuvant- and additive-therapy. PMID- 10714206 TI - [Worldwide consensus: the way from the KIEL classification to the REAL classification to the WHO classification]. AB - The history of lymphoma classification has been long and controversial. However, within the last thirty years much has been learned about the biology of lymphoma. The concept of classifying malignant lymphoma according to the (proposed) normal counterpart was developed in the KIEL classification. In 1994, the so-called revised European/American lymphoma classification was published as a consensus of pathologists from both sides of the Atlantic. Its clinical significance was proven in the International Lymphoma Classification Project. The upcoming WHO Classification is based on the principle to define disease entities that can be recognized by the pathologists and are of clinical relevance. Within these entities, histopathological variants and clinical subtypes are described, which may or may not bear prognostic relevance. Prognostic factors can be defined within an entity, on a clinical, morphological, immunohistochemical or genetic basis. PMID- 10714207 TI - [Suspicious acinar proliferations of the prostate]. AB - A variety of small acinar lesions of the prostate may mimick prostate cancer. In the central and transition zone of the prostate atypical adenomatous hyperplasia (AAH) has to be differentiated from low grade carcinoma (Gleason score 2-6). In the dorso-peripheral zone high grade PIN (prostatic intraepithelial neoplasie) and ASAP (atypical small acinar proliferations) represent the most important mimicers of carcinoma. High grade PIN has to be differentiated from intraductal carcinoma, ASAP on the other hand may mimic low grade carcinoma. The significance of basal cell type cytokeratin immunhistochemistry (IHC) in the differentiation between ASAP and low grade carcinoma of the prostate is assessed by additional MIB-1 IHC. The status of the basal cell layer in ASAP was shown to be variable (complete, fragmented and partial loss). Independently from the status of the basal cell layer the mean MIB-1 proliferation index of ASAP was significantly higher than of clearly benign lesions and did not differ from that of low grade carcinoma. Taking into account the high detection rate of carcinoma in repeat biopsies, close clinical follow up of patients with ASAP should be recommended. PMID- 10714208 TI - [New aspects of lung tumor pathology]. AB - The "new" 3rd WHO classification of lung tumors 1999 is described in nine groups of malignant broncho-pulmonary neoplasias. In recent years, our knowledge of the quite variable biology of tumors has been significantly increased by the use of immunohistochemical methods and molecular biology. These methods facilitated an improved qualitative and quantitative characterization of heterogeneously differentiated long tumors (e.g., neuroendokrine/blastomatoid portions etc.). The detection of genetic alterations of tumor suppressors (Rb/p53/FIHT/TGF beta R-2 etc.) and oncogenes (e.g., myc/fos/blc-2/ras/erbB etc.) is, at the moment, only of scientific interest. The genetic heterogeneity of tumors is emphasized by results obtained with the comparative genomic hybridization (CGH). In small-cell carcinomas of the lung more than seven variable chromosomal alterations- especially loss of genetic material in the regions 3p, 10q, and 4q--can be detected in a tumor. Adenocarcinomas show losses of genetic material on chromosomes 9 and 19, and/or gains on chromosome 1. Squamous cell carcinomas frequently exhibit losses on chromosome 2 and gains on chromosome 3. A connection between the detected genetic anomalies and histomorphological growth patterns can not be seen. At the present time, the validity of individual findings for a correlation between operability, tumor progress, chemotherapy and prognosis is not sufficiently elucidated by investigations nor secured. The histopathological, immunohistochemical, and genetic characterization of specimens of, mostly advanced, lung tumors that show variable phenotypes in biopsies of just 1-2 mm does not allow conclusions regarding causal factors (e.g., smoking, radon, asbestos etc.) or further progress of the disease. Therapeutical approach and the still unfavourable prognosis remain essentially, as in the last thirty years, to be characterized by TNM and performance status of the individual patient and, to a lesser extent, by the main histological type of tumor. PMID- 10714209 TI - [New findings in orthopedic pathology]. AB - The term orthopedic pathology refers to bone- and joint-affecting diseases which are important for the orthopedic surgeon. In the report presented here, emphasis is placed on the membrane-associated proteolysis, which is essential for the degradation of the extracellular matrix. Matrix-degrading processes play a role not only in arthrosis but also in rheumatoid arthritis. Moreover, they are strongly associated with the problem of loosening of protheses, which is of utmost importance for the orthopedic surgeon. In these processes, major roles are played by the plasminogen activator system, plasmin, different matrix metalloproteinases, including the membrane type matrix metalloproteases and different cathepsins. A deeper insight into the function of these proteins and their influence on the matrix degradation in joint diseases will open the way for new diagnostic and therapeutic strategies. Investigations into a large number of chondrosarcomas have shown that for this type of bone lesions, urokinase plasminogen activator and cathepsin B are prognostic parameters that are independent of the differentiation grade. Also, in this context, investigations into the membrane-bound proteases will be of great practical and diagnostic value. PMID- 10714210 TI - [Rudolf-Virchow Prize 1999. Genetics of respiratory tract carcinomas: correlation of genotype and phenotype]. AB - In contrast to carcinomas of the upper respiratory tract lung cancer shows a considerable variety of histological differentiations and is particularly known for its morphological heterogeneity. Of clinical relevance, however, is only the distinction between small cell carcinomas (SCLC) and non-small cell lung cancer (NSCLC). We meanwhile investigated a tumor collective of several hundred respiratory tract carcinomas by Comparative Genomic Hybridization (CGH) and developed computer software for the statistical comparison of tumor groups. The analysis revealed recurrent patterns of chromosomal imbalances which are associated with morphological histotypes and biological phenotypes, e.g. there are chromosomal imbalances predominantly found in SCLC compared to NSCLC but also a considerable overlap between both entities. Specifically, the pattern of metastasizing lung squamous cell carcinomas (SCC) approaches that of SCLC. In addition, the analysis of a metastasizing combined SCLC after microdissection showed a clonal relationship between the SCC component of the primary tumor and the SCLC metastasis. These findings have direct consequences for the pathogenesis and classification of lung cancer. First, SCLC should be differentiated into primary and secondary carcinomas. Whereas primary SCLC as the predominant tumor type evolve directly from a precursor cell of probably epithelial origin, secondary SCLC develop via a NSCLC intermediate. Second, neuroendocrine differentiation in lung carcinomas should be used as a marker for dedifferentiation, worse prognosis and rapid tumor progression rather than an indicator of a putative tumor stem cell. The primary data is available at our online tumor database at http://amba.charite.de/cgh/. PMID- 10714211 TI - [Reduction of acute kidney transplantation rejection by the chemokine receptor antagonist Met-RANTES]. AB - Chemokines contribute to the mononuclear cell infiltrate in vessels and interstitium which is characteristic of renal transplant rejection. By employing the chemokine receptor blocker Met-RANTES it was shown that recruitment of inflammatory cells into renal allografts could be significantly suppressed. In a renal transplant model (Fisher RT1(1v1) rat kidney into Lewis RT1(1) rat) Met RANTES-treated animals showed a significant reduction in vascular injury score (16.10 +/- 5.20 vs. 62.67 +/- 18.64) and tubular damage score (15.70 +/- 5.22 vs. 33.00 +/- 6.44) relative to untreated animals. In a severe rejection model (Brown Norway RT1n rat kidney into Lewis RT1(1) rat), Met-RANTES significantly augmented low-dose cyclosporin A treatment to reduce all aspects of renal injury including interstitial inflammation (score 71.00 +/- 6.10 vs. 157.30 +/- 21.30). In a monocyte attachment assay on microvascular endothelium under physiological flow conditions exposure of microvascular endothelium to RANTES resulted in RANTES immobilization and RANTES-induced firm adhesion of monocytes only after prestimulation of the endothelium with IL-1 beta. Met-RANTES completely inhibited this RANTES-mediated arrest. Thus, Met-RANTES can reduce acute rejection by impeding leukocyte arrest to inflamed endothelium. PMID- 10714212 TI - [Inhibition of angiogenesis on the chicken chorioallantoic membrane by Ets 1 antisense oligodeoxyribonucleotides]. AB - The Ets 1 transcription factor, the founder of the ets gene family of transcriptional regulators, has strongly been supposed to play a role in angiogenesis under both physiological and pathological conditions including tumor vascularization. An in vivo role has nevertheless not yet been proven. We therefore investigated whether an Ets 1 antisense oligodesoxynucleotide (ODN) blockade effectively inhibits in vivo angiogenesis in the chicken chorioallantois membrane-assay. We used a 20-mer phosphorothioate directed against the AUG initiation codon and a short 3' sequence of the c-ets 1 mRNA (5' AGATCGACGGCCGCCTTCAT-3') in order to block Ets 1 expression. Three quantities of either antisense or negative control sense ODNs were directly applied on the chorioallantois membrane on day five of development and results evaluated on day seven. No effect on angiogenesis was seen in the antisense group treated with 2.5 micrograms ODN/egg compared to the sense control group. In contrast chorioallantoic blood vessel numbers and diameters were considerably reduced after application of 5 micrograms antisense ODN. 10 micrograms of either antisense or sense ODNs turned out to be toxic: all 6 embryos had died on day seven. Our results are the first proof that the Ets 1 transcription factor is actually necessary for the regulation of in vivo angiogenesis. Its role is probably not restricted to development but also concerns new blood vessel formation during chronic inflammation and tumor vascularization. PMID- 10714213 TI - [Loss of junB expression causes a myeloproliferative syndrome resembling chronic myeloid leukemia]. AB - JunB, a member of the AP-1 family of transcription factors, has been implicated in the control of proliferation and differentiation of various cell types including myeloid cells. We found that absence of junB expression in the myeloid lineage of mice lead to a myeloproliferative syndrome resembling human chronic myeloid leukaemia. The disease was due to a cell autonomous increase in the proliferation of immature myeloid cells. These results suggest that junB is a negative regulator of myelopoiesis. PMID- 10714214 TI - [Analysis of kidney tumors in trichloroethylene exposed workers by comparative genomic hybridization and DNA sequence analysis]. AB - Environmental and occupational mutagens such as chemical agents are suspected to be involved in cancer development. However, a direct link between carcinogen exposure and genetic alterations has been established only in a few examples. Carcinogen involvement in renal cell carcinoma (RCC) was recently supported by reports of an increased frequency of RCC among workers exposed to Trichloroethylene (TRI) in Germany. The aim of this study was to evaluate the phenotype and the genotype of renal tumors in occupationally TRI-exposed patients. Sporadic RCCs from TRI nonexposed patients served as control. Normal and renal tumor tissues from 12 patients with occupational histories of solvents and TRI exposure were histologically analyzed. Comparative Genomic Hybridization (CGH) was used to screen for genomic alterations along all chromosomal arms. Tumor DNA was analyzed for mutations of the von Hippel-Lindau gene (VHL) on 3p25.5 by PCR and sequencing analyses. The histological analysis revealed 9 clear cell (75%), 2 papillary RCC (18%) and 1 oncocytoma (8%). In clear cell RCC, DNA losses were frequently observed on 3p (66%), 6q (56%), 9p and 4q (44% each) and 2q (33%). DNA gains were detected on 9q, 17 (33% each) and 5q (22%). DNA gains were identified on chromosome 7 and 17 in papillary RCC. Four VHL gene mutations were found in three clear cell RCC (33%) with 3p loss. Comparisons of these results with data obtained from sporadic RCC revealed no unique phenotype, genotype or mutation pattern in the VHL gene of renal tumors after TRI exposure. Therefore, further studies on the relevance of dose and exposure time for the nephrocarcinogenic effect of TRI are indicated. PMID- 10714215 TI - [Identification of prognostic parameters for renal cell carcinoma by cDNA arrays and cell chips]. AB - We report here an example of how the combined application of cDNA and tumor array can lead to the identification of a novel prognostic marker within a very short time. A cDNA array analysis was performed on 5184 cDNA clones on a filter to screen for genes with differential expression between the renal cancer cell line CRL-1933 and normal kidney tissue to identify genes with relevance in RCC. There were 89 differentially expressed genes in the cancer cell line, one of them coding for vimentin, a cytoplasmic intermediate filament. To determine the in vivo prevalence and prognostic significance of vimentin expression, a renal cancer tissue micro array containing 532 RCC specimen was constructed and vimentin expression was determined by immunohistochemistry. Vimentin expression was frequently seen in clear-cell (51%) and papillary RCC (61%), but rarely in chromophobe RCC (4%) and oncocytomas (12%). These results obtained from minute arrayed tumor samples match with previous findings on vimentin expression in renal tumors. Furthermore, vimentin expression was significantly associated with poor patient prognosis (p < 0.007) independently of grade and stage. These results suggest that tissue microarray analysis provides a rapid and powerful method to determine the prevalence and prognostic significance of novel candidate genes discovered to be involved in cancer development with large-scale cDNA expression arrays. The tissue array can be adapted as a routine tool in research laboratories for analyses of large tumor series at the DNA, RNA or protein level. With such a tool, cancer researchers can study vast numbers of tumor samples in a short time and can generate a wealth of data on the application of tumor markers. PMID- 10714216 TI - [Novel mutation-specific monoclonal E-cadherin antibodies make possible allele differentiation at the protein level in tumors]. AB - Somatic deletion mutations in the cell adhesion molecule E-cadherin are present in almost 50% of diffuse type gastric cancer. We recently generated monoclonal antibodies against an in-frame deletion of exon 9. The aim of this study was to generate and characterize monoclonal antibodies against the second mutational hot spot, in-frame deletions of exon 8. Lou/C rats were immunized using a KLH-coupled peptide that represents a unique sequence generated by fusion of exon 7 and exon 9 from an E-cadherin deletion mutation lacking exon 8. Hybridoma supernatants were tested in a solid-phase immunoassay using BSA-coupled peptide. Positive reacting hybridomas were confirmed by Western Blots, FACS analysis, and immunohistochemistry of E-cadherin negative carcinoma cells that had been transfected with mutant and wild-type E-cadherin cDNA, respectively. In addition, routine formalin fixed and paraffin embedded tissues from gastric cancer patients were analyzed using both mutation-specific and commercial monoclonal antibodies against E-cadherin, including HECD-1 and AEC. Two hybridoma supernatants, termed E-cad delta 8-1, were selected that reacted with the mutant peptide used for immunization and gave strong signals in Western Blot and FACS analysis with cells expressing mutant E-cadherin lacking exon 8. Wild-type protein expressing cells only reacted with the commercial antibodies but not with the two selected hybridoma supernatants. In contrast to AEC, monoclonal antibody HECD-1 did not react with exon 8 deleted E-cadherin, suggesting that the previously unknown epitope for this often used monoclonal antibody is located at least in part within exon 8. Four gastric cancer specimens known to express mutated E-cadherin mRNA strongly reacted with both mutation-specific supernatants and with AEC monoclonal antibody but not with HECD-1. Taken together, we succeeded in generating monoclonal antibodies reacting with mutant E-cadherin protein lacking exon 8. Furthermore, using both HECD-1 and the new mutation-specific antibodies E cadherin immunoreactivity can for the first time be evaluated in an allele specific manner in archival tissues. PMID- 10714217 TI - [Aspirin suppresses microsatellite instability]. AB - Nonsteroidal anti-inflammatory drugs (NSAIDs) exhibit cancer preventive effects and have been shown to induce regression of adenomas in FAP patients. In order to elucidate the probable underlying mechanism, the effect of NSAIDs on mismatch repair related microsatellite instability was investigated. Six colorectal cancer cell lines all but one deficient for human mismatch repair (MMR) genes were examined for microsatellite instability (MSI) prior and after treatment with Aspirin or Sulindac. For rapid in vitro analysis of MSI a microcloning assay was developed by combining Laser microdissection and random (PEP-) PCR prior to specific MSI-PCR. Effects of NSAIDs on cell cycle and apoptosis were systematically investigated by using flow cytometry and cell-sorting. MSI frequency in cells deficient of MMR genes (hMSH2, hMLH1, hMSH6) was markedly reduced after long-term (> 10 weeks) NSAID treatment. This effect was reversible, time- and concentration dependent. However, in the hPMS2 deficient endometrial cancer cell line (HEC-1-A) the MSI phenotype kept unchanged. According to cell sorting, non-apoptotic cells were stable and apoptotic cells were unstable. These results suggest that aspirin/sulindac induces a genetic selection for microsatellite stability in a subset of MMR-deficient cells and may thus provide an effective prophylactic therapy for HNPCC related colorectal carcinomas. PMID- 10714218 TI - [MALT-type B-cell lymphomas escape fas-mediated apoptosis]. AB - Important prognostic factors in primary MALT-lymphomas are stage and grading since the localized low-grade lymphoma may be cured with antibiotic treatment that may result in lymphoma regression as exemplified in Helicobacter eradication (H.p.) in gastric low-grade MALT-lymphomas. For obscure reasons, around 20% of low-grade MALT-lymphomas do not respond to eradication therapy or contain in 25 33% high-grade components as a possible consequence of tumor cell transformation. Given that MALT-lymphoma B-cells are autoimmune in nature and proliferate in response to antigen and T-cell-mediated signals, it is suggestive that autoreactive B-cell lymphoma precursors generated during chronic inflammation escaped the tolerance checkpoint. This mechanism normally operates in healthy individuals through CD40-L/FAS-L expressing activated T-cells and is fundamental in the deletion of harmful B-cells. Analyzing the role of FAS/FAS-L mediated apoptosis in lymphomagenesis, we purified B- and T-cells from low- and high-grade MALT B-cell tumors and tonsillar memory B-cells as control. T-cells were stimulated in vitro to express CD40-L and FAS-L using anti-CD3 antibodies and were subsequently cocultivated with B-cells. Apoptosis was 3 times increased in normal memory B-cells and, to a lesser extent, in a subgroup of low-grade MALT lymphomas as compared with the spontaneous apoptosis without T-cell coculture. In striking contrast, all primary high-grade MALT-type lymphomas showed increased survival but were resistant to FAS-mediated apoptosis. Furthermore, 40% of low grade MALT-type lymphomas were resistant to apoptosis and showed mutations in the FAS-gene. The conclusion that self-tolerant high-grade but also a subgroup of low grade MALT-lymphoma B-cells escaped censoring by the FAS pathway may therefore identify a novel regulatory step in early MALT-lymphomagenesis. PMID- 10714219 TI - Identification of p62, a phosphotyrosine independent ligand of p56lck kinase, as a major component of intracytoplasmic hyaline bodies in hepatocellular carcinoma. AB - Intracytoplasmic hyaline bodies (IHBs) resemble a peculiar type of cytoplasmic inclusions in cells of hepatocellular carcinoma (HCC) which so far have escaped further characterization. In order to determine protein composition of IHBs we investigated tissue of a HCC containing numerous IHBs by immunohistochemistry and Western blot analysis using a large panel of different antibodies. Our studies revealed immunoreactivity of IHBs with the monoclonal antibodies SMI 31 and MPM-2 which recognize hyperphosphorylated epitopes present on paired helical filaments in Alzheimer's disease brains (SMI 31) and on proteins hyperphosphorylated by mitotic kinases (MPM-2), respectively. In two-dimensional Western blots of HCC extracts SMI 31 and MPM-2 antibodies detected a 62 to 65 kD protein with an isoelectric point around 4.5. Microsequencing identified this protein as p62, a recently identified phosphotyrosine-independent ligand of the SH2 domain of tyrosine kinase p56lck. Immunoreactivity of p62 protein spots with antibodies to phosphorylated epitopes (i.e. SMI 31 and MPM-2) suggest that p62 is highly phosphorylated in IHBs. This is the first report on accumulation of p62 as cellular inclusions and its association with human disease. PMID- 10714220 TI - [Mutational pattern of Ig-VH genes of synovial B-lymphocytes from different anatomical locations in a rheumatoid arthritis patient]. AB - Synovial B-lymphocytes which are a constant feature of RA synovialitis are expanded in an antigen dependent manner. To understand the nature of the antigen driving the B-cell expansion it would be interesting to know whether there exists a restricted number of antigens. Therefore the usage of germline genes and the mutational pattern of Ig-VH-genes from synovial B-cells from 3 different anatomical regions with different onsets of local disease were analysed. Together with an immunohistological study characterising the inflammatory infiltrate (CD3, CD22, CD23 and Ki-M4), RNA was prepared from tissue sections taken from the right and left peroneal tendons and right elbow of a patient with seropositive RA. The corresponding cDNA was amplified using specific VH primers and the obtained variable region sequences were compared to their closest germline counterparts on the EMBL/Genbank data base. The molecular analysis demonstrated somatically mutated VH genes (total R/S ratios in CDR 1 + 2: right peroneal tendon 7.5; left peroneal tendon 3.5 and right elbow 3.0) in all three different regions, with two clones presenting aminoacid deletions. The values of total R/S ratios correlated directly to the duration of local disease. The aminoacid sequences from the different locations exhibited strong homology among each other. This homology was of 77% for the 19 clones belonging to the VH1 family. Immunohistochemistry demonstrated in two locations a dense follicle-like infiltrate with FDC-network, the third location presented a non-follicular distribution of lymphozytes without FDC. Hence the R/S values were correlated directly with the duration of local disease, it appears that here is an ongoing antigen-dependent B-cell activation in joints probably occurring in the FDC-networks. Moreover this study suggests- due to a high homology among the VH aminoacid sequences from B-cells of different anatomical regions--that a restricted number of antigens is involved in the B cell expansion of RA patients. PMID- 10714222 TI - [List of members]. PMID- 10714221 TI - [Podoplanin--a specific marker for lymphatic endothelium expressed in angiosarcoma]. AB - AIMS: Angiosarcomas apparently derive from endothelia of the blood vasculature, however occasionally their histologic features suggest mixed origin from blood and lymphatic endothelia. In the absence of specific positive markers for lymphatic endothelia the precise distinction between these components was not possible so far. Here we provide evidence that podoplanin, a approximately 38 kD membrane glycoprotein of podocytes is a specific marker of lymphatic endothelium that was used to identify the relative fraction of tumor cells with lymphatic or blood vascular endothelial phenotype in vascular tumors. METHODS: Podoplanin was localized in normal human skin and kidney cortex by immunohistochemistry on paraffin sections, double immunofluorescence on frozen sections with PAL-E, immunoelectron microscopy and by immunoblotting. 45 vascular tumors (29 benign lesions, 11 angiosarcomas and 5 gastrointestinal Kaposi's sarcomas) were evaluated for podoplanin expression. Complementary staining was obtained with established endothelial markers (CD 31, CD 34, Factor VIII related antigen, UEA I) and with podocalyxin, another podocytic protein mainly present in endothelia of blood vessels. RESULTS: In human tissues podoplanin is specifically expressed in the endothelium of lymphatics, but not in blood vasculature or in hemangiomas. This expression is preserved in endothelia of all benign lymphatic tumorous lesions and all Kaposi's sarcomas examined. By contrast 10 out of 11 G3 angiosarcomas contained only variable fractions of podoplanin-expressing tumor cells. Most tumor cells coexpressed podoplanin and markers of blood vessel phenotype. CONCLUSIONS: (1) Podoplanin is a selective marker of lymphatic endothelium; (2) G3 angiosarcomas display a quantitative spectrum of podoplanin expressing tumor cells; (3) In the majority of angiosarcomas tumor cells coexpress podoplanin and endothelial markers of blood vessels; (4) All endothelial cells of Kaposi's sarcomas expressed the lymphatic marker podoplanin. PMID- 10714223 TI - [Current complex noninvasive diagnosis of parathyroid tumors]. AB - The diagnosis of bulky formations of the parathyroid glands (PTG) has become possible since current high-resolution techniques of visualization, such as ultrasound study (USS), computed tomography (CT), magnetic resonance imaging (MRI), were introduced into practice. The presence of clinical and/or laboratory signs of hyperparathyroidism (HPT) is a signal to initiate a goal-oriented search for abnormal PTG formations. The complex diagnosis of HPT involves the methods of detecting osteoporosis ranging from routine X-ray study of the hand and foot to more in-depth techniques: dichromatic X-ray absorptiometry (DXA) and quantitative CT (QCT). USS is an excellent method for screening if abnormal PTC changes are suspected; however, negative USS results in the presence of clinical and/or laboratory signs of HPT should not stop a diagnostic search. CT with intravenous contrast bolus specifies the site and structure of an formation, has some advantage in detecting retrosternal tumors. Due to its high tissue contrast, three-dimensional images, none ionizing radiation and osseous structural artefacts, MRI becomes a preferable tool for studying PTG when they are typically or atypically located. Needle biopsy is required when noninvasive methods cannot characterize the pattern of an abnormal PTG formation properly or their results are contradictory. PMID- 10714224 TI - [Magnetic resonance tomography in the diagnosis of lung metastases in the mediastinum and thorax]. AB - The results of MRI in 81 patients with morphologically verified lung cancer, mainly Stages IIIA and IIIB, were analyzed. They were compared with CT data in 37 cases and surgical findings in 28. MRI was performed by using Magnaview 0.04 T and Vectra 0.5 T apparatus in the T1- and T2-weighted SE and PC sequences as well in the fat-suppression mode. Thoracic metastases were evaluated from the direct signs tumor spread into the adjacent tissue and vessels. The criteria for the involvement of lymph nodes were their over 1-cm enlargement and characteristic changes in the intensity of signals from them. CT was found to yield less information on pleural, pericardial, and vascular invasion (66-75% sensitivity). MRI detected this type of cancer spread (88-94% sensitivity). Both techniques have nearly equal sensitivities in revealing intrathoracic lymphadenopathy. The interpretation of MRI data did not depend on the voltage of a magnetic field. It is recommended that MRI should be made after CT when there is a need for assessing large vessels or for making clear the data that remain open to question following CT. PMID- 10714225 TI - [Radiologic diagnosis of different forms of acute pancreatitis]. AB - By using their data on 123 patients with different forms of pancreatitis, the authors describe the ultrasound and computed tomographic semiotics of this condition. The authors consider ultrasonography and computed tomography to be highly informative in diagnosing different forms of the disease and its complications. Both methods not only assess the pancreatic parenchyma, the extent of the process to the adjacent anatomic structures, but permit a follow-up and diagnostic and treatment measures under their visual guidance. PMID- 10714227 TI - [Diagnosis of remote metastases of rectal cancer with the aid of minimally invasive diagnostic procedures under computerized tomography control]. AB - At 124 patients who have rectum cancer made computer tomography of a abdominal cavity, retroperitoneal space and small pelvis with the purpose of definition of prevalence of a tumour. Was used computer tomograph "Somatom CRX", "Siemens". Remote metastasis are revealed at 26 patients, their most often localization- liver (45.8%). Are described CT-attributes of organs remote metastasis: hypodense spherical formation from 1.0 up to 4.0 CM in a diameter. Pathology changed lymph nodes were characterized by increase of a diameter more 1.5 CM, illegibility of contours, merge in a conglomerate. This changes have not specific character and also can be in case of hyperplasia of lymph nodes. At 14 patients is made puncture biopsy of focal formations under the CT-control. Diagnostic efficiency CT with puncture biopsy have made: accuracy--0.92, sensitivity--0.77, specificity -0.96. Is judged high efficiency of a method. PMID- 10714226 TI - [Long-term results of endovascular treatment of metastatic renal cell cancer]. AB - The authors analyse the long-term results of treatment of 106 patients with advanced renal cell carcinoma. The patients were divided in two groups. Group 1 (42 patients) included cases with single metastatic lesion. Patients of group 2 (n = 64) had more than one site of metastases. All patients were treated by nephrectomy, embolization with chemotherapeutic agent or arterial occlusion without cytostatic drug except nine patient of group 2, who has undergone chemo and hormonal therapy. The survival according to method at treatment was studies. After occlusion satisfactory results were obtained in cases with single metastasis lesion and also in cases with many sites of metastases. Oily chemoembolization in renal cell carcinoma with distant metastases has given better results as compared with the embolization without chemodrugs. We conclude that endovascular method of treatment is an alternative to the traditional nephrectomy in palliation of renal cell carcinoma. PMID- 10714228 TI - [Invasive interventions guided by computerized tomography in the diagnosis and treatment of diseases of the internal organs]. AB - Diagnostic punctures under CT guidance were made in 544 patients with diseases of the chest (n = 303), abdomen (n = 149), and retroperitoneal space (n = 92). In 87 patients, diagnostic punctures were combined with therapeutical manipulations and included aspiration and drainage of visceral organ cysts and abscesses. The proposed procedure of diagnostic biopsies under CT guidance could ascertain the morphological nature of a lesion in 90.1% of cases prior to treatment. Therapeutical aspirations and drainage of visceral cysts and abscesses under CT guidance resulted in their complete recovery in 97.8 and 85.7% of cases, respectively. Diagnostic and therapeutical interventions under CT guidance caused complications in 6.9% of cases. At diagnostic biopsy, pneumothorax is the most frequent complication *5.3%), and lung tissue hemorrhage along the puncture needle passage is the less frequent (1.3%). The use of therapeutical interventions developed complications in 1.1% of cases. PMID- 10714229 TI - [Age-related x-ray features of the spine in patients with achondroplasia]. AB - The paper studies the age-related X-ray features of the spine in patients with achondroplasia. It gives the time course of changes in the shape of vertebrae, the specific features of apophyseal ossification, provides a quantitative account of the shorter caudal lumbar vertebral arch root distance symptom. The time course of changes in the size of the lumbosacral angle was examined. The findings suggest that there are not only considerable static changes in the spine of patients with achondroplasia, but also significant age-related features of vertebral tissue growth and differentiation. PMID- 10714230 TI - [MRI diagnosis of soft tissue tumors]. AB - MRI findings of 13 patients with soft tissue tumors (STT) are presented. There are were abnormalities, such as primary STTs of the hip in 5 patients, the back in 1, and the neck in 1, STT relapses of the hip in 2 and those of the back in 1. Two patients had chronic hip STT hematomas and 1 had hip STT metastatic melanoma. The diagnosis was verified in 11 cases (in 10 cases at surgery and 1 at needle biopsy). MRI makes it possible to define the accurate sizes of the tumor, its structure and relationship to its adjacent tissues, which is important in choosing at treatment policy, the type and scope of a surgical interventions. PMID- 10714231 TI - [Possibilities of the liver MRI in complex diagnosis of primary hemochromatosis and in evaluation of the treatment effectiveness]. PMID- 10714232 TI - [Prospects of radiotherapy in nontumor diseases]. PMID- 10714234 TI - [Lesson 9. Source of radiation]. PMID- 10714233 TI - [Diagnostic effectiveness and safety of currently available X-ray contrast media]. PMID- 10714235 TI - GnRH receptor and apoptotic signaling. AB - In addition to its hypophysiotropic action, gonadotropin-releasing hormone (GnRH) can modify activity in extrapituitary organs and peripheral tumors. GnRH analogs are the preferred treatment for advanced and even metastatic or recurring carcinomas in vivo and in vitro. Hormone-responsive tumors undergo apoptosis with the appropriate stimulus; GnRH-induced tumor growth arrest may result from stimulated apoptotic cell death. The sensitivity of tumors and normal tissue to GnRH is strongly associated with the possession of receptors for GnRH as well as other hormonal control. Despite the lack of a precise apoptotic signaling cascade through GnRH receptors, biochemical events observed within a plasma membrane appear to constitute the most convincing evidence that the membrane event is primarily stimulated during cell activation by GnRH. GnRH receptors in tumors differ from those in pituitary gonadotrophs in some aspects, in particular with regard to the transmembrane signaling cascade. The intramembranous phenomena that occur independently of the contribution of other organelles upon tumoral GnRH receptor engagement include (i) activation of phosphotyrosine phosphatase and loss of phosphotyrosine from the endogenous membrane protein and (ii) phosphoinositide and perhaps sphingomyelin cleavage producing lipid-originated second messengers. GnRH has also been demonstrated to increase Fas ligand expression within plasma membrane, which is known to promote apoptotic cell death through attack on Fas-positive cells within tumors. The Fas-Fas ligand complex might, at least in part, account for the antiproliferative action of the hormone. An understanding of the relationship between the extracellular (hormonal) stimuli that leads to cell death and the intracellular events regulating growth arrest on GnRH action may fundamentally help clarify the therapeutic approach to all hormone-dependent carcinomas that respond to stimuli that lead to apoptosis. In this chapter, we review the recent literature and the results of our studies on GnRH-induced membrane events and summarize what is currently known about this promising antiproliferative function. PMID- 10714236 TI - Structure and function of the growth-hormone-releasing hormone receptor. AB - Growth-hormone-releasing hormone (GHRH) stimulates growth hormone (GH) secretion and GH synthesis and is also thought to cause somatotroph proliferation. Specific high-affinity binding sites for GHRH have been demonstrated on pituitary membranes using iodinated GHRH analogs. The complementary DNA encoding for the human GHRH receptor (GHRH-R) was recently cloned. The open reading frame was shown to extend 1269 bp and thus to encode a protein of 423 amino acids with a predicted molecular weight of 47 kDa. Expression is restricted to specific tissues. Analysis of the genomic structure revealed that the human GHRH-R gene spans 15 kb and consists of 13 exons. The 5'-flanking region of the human GHRH-R gene was recently characterized. Transcriptional regulation of the GHRH-R is discussed in this review. Mechanisms of signal transduction for control of GH transcription and secretion are presented. Furthermore, the role of the GHRH-R in proliferation and differentiation of the somatotrophic pituitary cell as well as in disease is examined. PMID- 10714238 TI - The EGF domain: requirements for binding to receptors of the ErbB family. AB - Epidermal growth factor (EGF) has been the prototype growth-stimulating peptide for many years. It has a characteristic structure with three disulfide bridges, which is essential for its activity. However, many other proteins, including both growth factors and proteins with unrelated functions, have similar EGF-like domains. This indicates that besides a characteristic conformation provided by the EGF-like domain, specific amino acids are required to provide specificity in protein functioning. Currently, more than 10 different growth factors with an EGF like domain have been characterized which all exert their action by binding to the four members of the erbB family of receptors. In this review, studies are described on the structure-function relationship of these EGF-like growth factor molecules in an attempt to analyze the individual amino acids that determine their binding specificity to the individual members of the erbB family. PMID- 10714237 TI - Regulators of growth hormone signaling. AB - Growth hormone acts through binding to membrane receptors that belong to the cytokine receptor superfamily. Ligand binding induces receptor dimerization and activation of the receptor-associated kinase: JAK2; this results in phosphorylation of the kinase itself, of the receptor, and of many cellular proteins. Among these are the Stat proteins as well as adaptors leading to the activation of the Ras/MAP kinase pathway and of the PI-3 kinase pathway. Activation by growth hormone is very transient and several mechanisms are involved in this downregulation: internalization and degradation of the receptor and recruitment of phosphatases or of specific inhibitors of the JAK/Stat pathway, the SOCS proteins. PMID- 10714239 TI - Enkephalin-cell interactions. AB - Enkephalin analogs for multivalent ligand systems, bivalent enkephalins, multivalent enkephalins on polymers, multivalent ligands on vesicles, simultaneous activation of two different receptor systems, and cell interactions of enkephalin/polypeptide conjugates are described. Multivalent ligand systems can trigger receptor-receptor interactions and are considered to possess interesting possibilities in terms of enhanced potency, reduction of side effects, and a new biological activity. PMID- 10714240 TI - Studies on structure-activity relationships of retinoic acid receptor ligands by means of molecular modeling. AB - Vitamin A and its biologically active derivatives, retinal and retinoic acid, play an important role in vision, are required for reproduction, act as morphogenic agents during embryonic development, and regulate the growth and differentiation of a wide variety of cell types throughout the life of an organism. The biological action of retinoic acid and synthetic analogs, referred to as retinoids, is mediated by RAR alpha, RAR beta, or RAR gamma and/or by RXR alpha, RXR beta, or RXR gamma, all being nuclear receptors. Since retinoids exert profound effects on cell proliferation, differentiation, and apoptosis, these compounds seem to be promising agents for the treatment of cancer. Consequently, a large number of retinoids have been synthesized and examined to determine if they exert their biological activity according to retinoic acid receptor interaction. These screening methods are often expensive, time-consuming, and labor-intensive procedures. Since one can construct the pharmacophores of congeneric groups of drug molecules, molecular modeling techniques offer a new way to determine the binding abilities of different agents. We examined the structural properties of retinoids, which allow them to specifically bind to the different receptor subtypes. The thus-generated 3D pharmacophore models were used to predict the binding affinities of several retinoids to the retinoic receptor subtypes. Finally, the 3D models served as criteria for searching the Derwent World Drug Index for compounds that possess the features necessary for favorable ligand receptor interaction. The search resulted in a "hit list" containing 323 compounds, some of which are worth further investigation to determine if they act via retinoic acid receptor binding or not. PMID- 10714241 TI - Glucocorticoid-regulated gene expression during cutaneous wound repair. AB - Glucocorticoids exert a deleterious effect on the wound healing process, which has been suggested to result from the anti-inflammatory action of these steroids. In addition, recent studies have demonstrated that glucocorticoids regulate the expression of various genes at the wound site which are likely to encode key players in the wound repair process. Using a murine full-thickness excisional wound healing model, we analyzed the effect of dexamethasone on the expression of various cytokines, growth factors, enzymes, and extracellular matrix molecules in normal and wounded skin. We demonstrate that the proinflammatory cytokines interleukin-1 alpha and -beta, tumor necrosis factor alpha, keratinocyte growth factor, transforming growth factors beta 1, beta 2, and beta 3 and their receptors, platelet-derived growth factors and their receptors, tenascin-C, stromelysin-2, macrophage metalloelastase, and enzymes involved in the generation of nitric oxide are targets of glucocorticoid action in wounded skin. These results indicate that anti-inflammatory steroids inhibit wound repair at least in part by influencing the expression of these key regulatory molecules. PMID- 10714242 TI - The aromatic hydrocarbon receptor, transcription, and endocrine aspects of dioxin action. AB - The widespread and persistent environmental contaminant 2,3,7,8 tetrachlorodibenzo-p-dioxin elicits adaptive and adverse biological responses by inducing changes in gene transcription. Some of dioxin's effects reflect disruption of endocrine homeostasis. The aromatic hydrocarbon receptor protein, together with its heterodimerization partner, the aromatic hydrocarbon receptor nuclear translocator protein, mediates dioxin action. There are notable similarities between the mechanism of dioxin action and the mechanisms of steroid/retinoid/thyroid hormone action. Studies of dioxin action may provide insights into the regulation of hormone-responsive genes and endocrine physiology. PMID- 10714243 TI - Control of food intake via leptin receptors in the hypothalamus. AB - Food intake is regulated via neural circuits located in the hypothalamus. During the past decade our knowledge on the specific mediators and neuronal networks that regulate food intake and body weight has increased dramatically. An important contribution to the understanding of hypothalamic control of food intake has been the characterization of the ob gene product (leptin) via positional cloning. Absence of circulating, functionally active, leptin hormone results in massive obesity as seen in ob/ob mice. Leptin inhibits food intake and increases energy expenditure via an interaction with specific leptin receptors located in the hypothalamus. Leptin receptors, of which there are several splice variants (Ob-Ra through Ob-Re), belong to the superfamily of cytokine receptors, which use the JAK-STAT pathway of signal transduction. Obese db/db mice, which have a mutation in the db locus, are unable to perform JAK-STAT signal transduction due to absence of functionally active (long form; Ob-Rb) leptin receptors. Ob-Rb is primarily expressed in the hypothalamus, with particularly high levels in the arcuate, paraventricular, and dorsomedial nuclei and in the lateral hypothalamic area. The abundance of leptin receptors in the ventromedial and lateral hypothalamus supports early observations that these two regions are intimately associated with the regulation of food intake. Leptin receptors have been identified in neuropeptide Y (NPY)/lagouti-related peptide (AgRP)- and proopiomelanocortin (POMC)/cocaine- and amphetamine-regulated transcript (CART) containing neurons of the ventromedial and ventrolateral arcuate nucleus, respectively, and in melanin-concentrating hormone (MCH)- and hypocretin/orexin containing neurons of the lateral hypothalamus, suggesting that the above mentioned messengers are mediators of leptin's action in the hypothalamus. Indeed, functional studies show that NPY, AgRP, POMC-derived peptides, CART, MCH, and hypocretins/orexins all are important regulators of food intake. Leptin is essential for normal body weight balance, but the exact mechanisms by which leptin activates hypothalamic neuronal circuitries is known to a limited extent. In order to find pharmaceutical approaches to treat obesity, further studies will be needed to reveal the exact mechanisms by which leptin lowers body weight and which role leptin and leptin receptors have in the pathogenesis of human obesity. PMID- 10714244 TI - Vitamin E and immunity. AB - Vitamin E is a potent antioxidant and has an ability to modulate host immune functions. This chapter consists of five parts: (1) vitamin E deficiency and immunity, (2) vitamin E supplementation and immunity, (3) vitamin E and the decreased cellular immunity with aging, (4) vitamin E and T-cell differentiation in the thymus, and (5) vitamin E and acquired immune deficiency syndrome (AIDS). In vitamin E deficiency most of the immune parameters show a downward trend, which is associated with increased infectious diseases and the incidence of tumors. In contrast, vitamin E supplementation has various beneficial effects on the host immune system. The decreased cellular immunity with aging or during the development of AIDS is markedly improved by the intake of a high vitamin E diet. In addition, vitamin E plays an important role in the differentiation of immature T cells in thymus. Vitamin E deficiency induces the decreased differentiation of immature T cells, which results in the early decrease of cellular immunity with aging in spontaneously hypertensive rats. Conversely, vitamin E supplementation induces a higher differentiation of immature T cells via increased positive selection by thymic epithelial cells, which results in the improvement of decreased cellular immunity in the aged. Furthermore, vitamin E supplementation induces the early recovery of thymic atrophy following X-ray irradiation. Taken together, these results suggest that vitamin E is an important nutrient for maintaining the immune system, especially in the aged. PMID- 10714245 TI - Transcobalamin II and its cell surface receptor. AB - Transcobalamin II (TC II), a nonglycoprotein secretory protein of molecular mass 43 kDa, and its plasma membrane receptor (TC II-R), a heavily glycosylated protein with a monomeric molecular mass of 62 kDa, are essential components of plasma cobalamin (Cbl; vitamin B12) transport to all cells. Evidence from studies over the past 10 years has provided some important information on their structure, regulation of expression, and function. Some of the specific findings include (a) identification of the structural relationship of the ligand TC II with other members of the Cbl-binding family of proteins, intrinsic factor (IF) and haptocorrin (HC), (b) regulation of TC II gene expression, (c) molecular basis for human TC II deficiency in patients with a lack of plasma TC II, (d) membrane expression, interactions, and dimerization of TC II-R, and (e) targeting and function of TC II-R in polarized epithelial cells. It is hoped that some of the recent findings presented in this review will provide new insights into the structure and function of these two fascinating proteins and stimulate future research in this area. PMID- 10714246 TI - Laparoscopic antireflux surgery: what is enough? PMID- 10714247 TI - The management of trauma in Canada. PMID- 10714248 TI - Soft-tissue images. Abdominal actinomycosis. PMID- 10714249 TI - Musculoskeletal images. Osteogenic sarcoma in the soft tissues of the hip. PMID- 10714250 TI - Soft-tissue case 31. Emphysematous cystitis. PMID- 10714251 TI - Musculoskeletal case 8. Mazabraud's syndrome--intramuscular myxoma associated with fibrous dysplasia. PMID- 10714252 TI - Use of abdominal computed tomography in blunt trauma: do we scan too much? AB - OBJECTIVES: To determine what proportion of abdominal computed tomography (CT) scans ordered after blunt trauma are positive and the applicability and accuracy of existing clinical prediction rules for obtaining a CT scan of the abdomen in this setting. SETTING: A leading trauma hospital, affiliated with the University of Ottawa. DESIGN: A retrospective cohort study. PATIENTS AND METHODS: All patients with blunt trauma admitted to hospital over a 1-year period having an Injury Severity Score (ISS) greater than 12 who underwent CT of the abdomen during the initial assessment. Recorded data included age, sex, Glasgow Coma Scale (GCS) score, ISS, type of injuries, number of abdominal CT scans ordered, and scan results. Two clinical prediction rules were found in the literature that identify patients likely to have intra-abdominal injuries. These rules were applied retrospectively to the cohort. The predicted proportion of positive CT scans was compared with the observed proportion, and the sensitivity, specificity, and accuracy were estimated. RESULTS: Of the 297 patients entered in the study, 109 underwent abdominal CT. The median age was 32 years, 71% were male and the median ISS was 24. In only 36.7% (40 of 109) of scans were findings suggestive of intra-abdominal injuries. Application of one of the clinical prediction rules gave a sensitivity of 93.8% and specificity of 25.5% but excluded 23% of patients because of a GCS score less than 11. The second prediction rule tested could be applied to all patients and was highly sensitive (92.5%) and specific (100.0%). CONCLUSIONS: The assessment of the abdomen in blunt trauma remains a challenge. Accuracy in predicting positive scans in equivocal cases is poor. Retrospective application of an existing clinical prediction rule was found to be highly accurate in identifying patients with positive CT findings. Prospective use of such a rule could reduce the number of CT scans ordered without missing significant injuries. PMID- 10714253 TI - A review of trauma systems using the Calgary model. AB - Surgeons caring for severely injured patients have witnessed tremendous change over the past 2 decades with the rapid evolution of trauma systems. This paper describes the evolution of trauma systems in Canada, using the one in Calgary as a model. Canadian system guidelines were produced by the Trauma Association of Canada in 1993. Participation in Canadian accreditation is accelerating as increasingly more centres across the country undergo external review each year. Reporting of trauma outcomes, including standardized mortality and a variety of performance measures, is becoming the norm. Injury is being treated as a disease with comprehensive control strategies aimed at reducing death and disability rates through prevention, treatment and rehabilitation. PMID- 10714254 TI - Value of DNA ploidy and S-phase fraction as prognostic factors in stage III cutaneous melanoma. AB - OBJECTIVE: To determine the prognostic value of flow cytometric analysis (S-phase fraction and DNA index) performed on lymph-node metastases of patients with stage III melanoma. DESIGN: A retrospective chart review with flow cytometric analysis of paraffin-embedded tissues. SETTING: A university teaching hospital. PATIENTS: Among 332 patients with cutaneous melanoma, 33 with stage III were identified. Distant metastases developed in 16 patients; 17 had no further recurrence. Charts were reviewed to obtain clinicopathologic parameters such as sex, age, location of the primary tumour, histologic features, presence or absence of ulceration, and Clark's and Breslow's levels. INTERVENTION: DNA ploidy and S-phase fraction were determined on the paraffin-embedded nodes. MAIN OUTCOME MEASURES: The groups with or without recurrence were compared in terms of disease-free survival (DFS) and overall survival (OS). These survival parameters were correlated with DNA ploidy and S-phase fraction. RESULTS: By univariate analysis, clinicopathologic factors did not predict OS. A higher Clark's level of invasion and more than 3 positive lymph nodes were associated with shorter DFS (p < 0.05). Tumour thickness and S-phase fraction did not correlate with either DFS or OS. Patients with diploid lymph-node metastases had an 87% 12-month survival compared with 41% for those with aneuploid tumours. CONCLUSIONS: DNA ploidy may be used as a prognostic index in patients with lymph-node metastases. This could be particularly useful in the context of sentinel lymph-node mapping by which more patients are being identified with single microscopic lymph-node involvement. PMID- 10714255 TI - Comparison of patellar resurfacing versus nonresurfacing in total knee arthroplasty. AB - OBJECTIVES: To determine whether resurfacing the patellar component during total knee replacement (TKR) influences the clinical outcome. DESIGN: A retrospective study of data gathered prospectively during the recovery course of patients who underwent TKR with or without patellar resurfacing. SETTING: Victoria General Hospital, Halifax, NS. PATIENTS: One hundred and eighty-five patients operated on between 1992 and 1995. The inclusion criteria were (a) osteoarthritis, (b) replacement carried out by 2 independent surgeons, (c) no comorbid illness such as rheumatoid arthritis, cancer or infection, (d) pre- and postoperative attendance at the assessment clinics. INTERVENTION: TKR with (45) or without (140) patellar replacement. MAIN OUTCOME MEASURES: Range of motion (ROM), pain assessment, Hospital Severity Score (HSS) and complications. RESULTS: There was no statistical difference between the 2 groups with respect to ROM, pain, HSS and complications postoperatively. CONCLUSIONS: Resurfacing the patella during TKR does not seem to influence the clinical outcome with respect to ROM, pain and overall complications. The decision should be based on individual criteria, depending on the preoperative and intraoperative findings. Randomized clinical trials assessing ROM, pain, complications and cost-effectiveness with long-term follow-up are necessary to further investigate this controversial issue. PMID- 10714256 TI - Ministernotomy for aortic valve replacement: a study of the preliminary experience. AB - OBJECTIVE: The aim of the study was to evaluate the technical feasibility and the postoperative course of aortic valve replacement through a ministernotomy. SETTING: The Montreal Heart Institute and the Hopital Lariboisiere, Paris, France. DESIGN: A case series from 2 institutions. PATIENTS: Fifty-one patients who underwent aortic valve replacement through a ministernotomy. The sternal incision was started at the level of the sternal notch extending down to the third or fourth intercostal space with a transverse section of the sternum at this level on both sides or limited to the right side (inverted T or L incision). Thirty-nine patients had aortic stenoses, 6 patients were operated for aortic insufficiency and 6 had mixed disease. The mean (and standard deviation) preoperative left ventricular ejection fraction was 0.56 (0.17). MAIN OUTCOME MEASURES: Cardiac bypass time, complications and outcome. RESULTS: The patients received Carbomedics and St. Jude mechanical valves, Hancock and Carpentier Edwards bioprostheses. Thirty-eight patients were administered antegrade and retrograde cardioplegia, 10 patients antegrade and 3 retrograde blood cardioplegia only. The mean (and standard error) cardiopulmonary bypass time and aortic cross-clamp time were 104 (38) minutes and 72 (16) minutes respectively. Two patients (4%) died and 2 patients (4%) showed evidence of a stroke after the procedure. Hospital stay averaged 8 (5) days. CONCLUSION: We conclude that aortic valve replacement can be done through a ministernotomy approach with perioperative results similar to those obtained through a conventional sternotomy. PMID- 10714257 TI - The effect of octreotide on postoperative adhesion formation. AB - OBJECTIVE: To investigate the effect of octreotide, a long-acting analogue of somatostatin, on postoperative adhesion formation, because somatostatin inhibits secretion of some growth factors that have modulatory effects on collagen synthesis. DESIGN: An experimental study. SETTING: Surgical Research and Biochemistry laboratories at Hacettepe University, Ankara, Turkey. SUBJECTS: Male Swiss albino mice. INTERVENTIONS: Both sides of a 5-cm ileal segment from Swiss albino mice were scraped 10 times, and transient ischemia was induced by clamping the segmental artery. Animals were injected subcutaneously with 1 mL/d of saline for 3 days (group 1), a single 5-mL intraperitoneal dose of saline (group 2), subcutaneously with 10 micrograms/kg daily of octreotide for 3 days (group 3) or a single 10 micrograms/kg intraperitoneal dose of octreotide (group 4). In half of the animals repeat laparotomy was performed on postoperative day 5. After adhesions were graded, the scraped ileal segments were excised for determination of hydroxyproline quantity. The same procedure was repeated on postoperative day 14 for the remaining animals. OUTCOME MEASURES: Adhesion grading, hydroxyproline levels. RESULTS: On postoperative day 5, the intraperitoneal octreotide group (group 4) had a significantly lower median adhesion score than groups 1 and 2. On postoperative day 14, both octreotide groups (3 and 4) had a significantly lower median adhesion grading than both saline groups (1 and 2). Hydroxyproline levels of the groups were not significantly different on either day 5 or day 14. CONCLUSION: Octreotide has a beneficial effect in decreasing adhesion formation in the early postoperative period. PMID- 10714258 TI - National trends in gastroesophageal reflux surgery. AB - OBJECTIVES: To assess the surgical technique and the frequency of different types of antireflux surgery used in Canada after the introduction of laparoscopic antireflux surgery. DESIGN: Gastroesophageal reflux (GER) surgery and population data in fiscal years 1992 through 1996. were accessed through the Canadian Institute of Health Information, provincial health ministries, MED ECHO and Statistics Canada databases. Data were also analysed by province and nationally for type of surgery (e.g., open abdominal, thoracic, thoracoscopic and laparoscopic). RESULTS: National data showed a slight increase in GER surgery in the last 5 years. Laparoscopic surgery increased 2.8 fold in 1993 and 1.6 fold in 1995 over the previous years. Open abdominal cases decreased 1.1 fold from 1992 to 1996. Thoracic cases remained essentially unchanged. Provincial and regional disparities in procedures per 100,000 population exist (Ontario 7.1 versus Nova Scotia 20.7). Areas in which little or no laparoscopic surgery was done had an average increase of 3%, whereas areas in which laparoscopic surgery was done had an average increase of 16% in GER surgery during the course of the study. In provinces west of Quebec (with the exception of Manitoba) more than 50% of GER surgery is laparoscopic; in areas east of Ontario less than 25% of GER surgery is performed laparoscopically. Five provinces (Manitoba, Quebec, Nova Scotia, Prince Edward Island and Newfoundland) performed significantly fewer laparoscopic procedures than the national average. CONCLUSIONS: The frequency of GER surgery is increasing modestly in Canada and is performed most often by the open abdominal route. Regional disparities in open and laparoscopic techniques are apparent. Laparoscopic surgery for GER is increasing rapidly and accounts for the decrease in open GER surgery. PMID- 10714260 TI - Isolated metachronous jejunal metastases after resection of bronchogenic carcinoma. PMID- 10714259 TI - Gastrocolic fistulization in Crohn's disease: a case report and a review of the literature. PMID- 10714261 TI - Epithelioid leiomyosarcoma of the stomach: report of a case. PMID- 10714262 TI - Pretreatment with fragments of substance-P or with cholecystokinin differentially affects recovery from sub-total nigrostriatal 6-hydroxydopamine lesion. AB - The neuropeptide substance P is known to have mnemogenic and reinforcing actions and can exert neurotrophic and regenerative effects in vitro as well as in vivo. Furthermore, our previous work in the rat showed that either pre- or post-lesion treatment with substance P can promote functional recovery in cases of partial nigrostriatal dopamine lesions. Other work has provided evidence that the effects of substance P might be differentially encoded by its C- and N-terminal fragments. The C-terminal fragment was found to be reinforcing, whereas the mnemogenic as well as neurotrophic properties have been ascribed to the N terminal sequences. Given these relations, we asked here whether pre-lesion treatment with either a C- or an N-terminal fragment of substance P might differentially affect the behavioral and neurochemical outcome of nigrostriatal dopamine lesions. Therefore, either substance P1-7 or substance P5-11 (37 nmol/kg each) was administered intraperitoneally daily for eight consecutive days before unilateral 6-hydroxy-dopamine lesions of the substantia nigra. Control rats received pre-lesion treatment with vehicle. Furthermore, we investigated the effects of pre-treatment with Boc-cholecystokinin-4 (0.91 nmol/kg), as we had found an increase in dopamine metabolism in animals that were pre-treated with cholecystokinin-8 in a former study. In accordance with our previous work, drug treatment effects were observed when excluding animals with most severe dopamine lesions: In animals with partial lesions (residual neostriatal dopamine levels of more than 10%), lesion-dependent asymmetries in turning behavior were observed in animals that were pre-treated with vehicle-, substance P1-7, or Boc-cholecysto kinin-4, whereas turning after pre-treatment with substance P5-11 was not significantly asymmetrical. Furthermore, the ipsi- and contra-lateral neostriatal dopamine levels did not differ significantly in this group. Moreover, pre treatment with substance P5-11 affected dopamine metabolism in the neostriatum and in the ventral striatum, as indicated by increased ratios of dihydroxyphenyllic acid to dopamine. The data provide the first evidence that the promotive effects of substance-P treatment in the unilateral dopamine lesion model might be mediated by its C-terminal and might depend on actions on residual dopamine mechanisms. PMID- 10714263 TI - Ultrastructure of Mauthner cells in fish adapted to long-duration vestibular stimulation and the effect of ethanol. AB - Adaptation or resistance of fish Mauthner cells (M-cells) to long duration (2 h) vestibular stimulation (LDS) was produced by daily brief and gradually increasing vestibular stimulation (training). The LDS resistance was accompanied by an increase in the number of desmosome-like junctions in the afferent axosomatic synapses. F-actin, the main component of desmosome-like contacts, has been suggested to be responsible for the increased resistance of M-cells to LDS. The purpose of the present study was to investigate the capacity of M-cells to adapt to LDS under the influence of ethanol, which alters the content of F-actin in cells. The experiments were carried out in goldfish fry. Vestibular stimulation (training and LDS) was performed in special drums that were rotated in two planes. The training time was increased from 1 min on day 1 to 30 min on day 30. For ethanol exposure, fish were immersed daily in a 2% ethanol solution for 20 min. To assess the level of resistance to LDS, motor activity indicating the functional state of M-cells was evaluated before and after LDS. The results show that exposure to ethanol reduces the resistance to LDS in both untrained and trained fish. Electron microscopic data demonstrated some structural changes in the synaptic endings located on M-cell soma in ethanol-exposed fish. Wrapping of boutons by cytoplasmic outgrowths and myelin-like structures was observed. Morphometric analysis revealed that exposure to ethanol without training decreases the number of desmosome-like contacts, probably due to ethanol-induced depolymerization of cytoskeletal actin. Ethanol exposure also partly suppressed the increase in the number of desmosome-like contacts that occurs as a result of training. In ethanol-treated trained fish, however, a concomitant increase in the length of desmosome-like contacts was observed. As training alone leads to the formation of additional desmosome-like contacts of standard length, it is possible that although a sufficient amount of such structures cannot be formed in the M-cells of ethanol-exposed trained fish, the existing contacts can be elongated. Thus, possibly changes of the actin state are involved in the adaptation of M-cells to LDS. PMID- 10714264 TI - Fetal spinal cord tissue in mini-guidance channels promotes longitudinal axonal growth after grafting into hemisected adult rat spinal cords. AB - Solid fetal spinal cord (FSC) tissue, seeded into semipermeable mini-guidance channels, was tested for the ability to promote axonal growth across the gap created by a midthoracic (T8) hemisection in adult rats. Fetal thoracic spinal cords, at embryonic days 13 to 15, were harvested and gently aspirated into mini guidance channels (1.25 mm in diameter and 3.0 mm in length). Care was taken to maintain the rostro-caudal orientation of the FSC. In control rats, the FSC channel construct was exposed to 5 freeze/thaw cycles to produce non-viable grafts before implantation into the hemisected cord. All cases revealed intact tissue cables of various diameters spanning the rostro-caudal extent of the lesion cavity, with integration of host-graft tissues at both interfaces. Immunofluorescence results indicated that numerous neurofilament-positive axons were present within the FSC tissue cable. Double-labeling of a subpopulation of these axons with calcitonin gene-related peptide indicated their peripheral nervous system (PNS) origin. Descending serotonergic and noradrenergic axons were found in the proximity of the rostral host-graft interface, but were not observed to grow into the FSC-graft. Anterograde tracing of propriospinal axons with Phaseolus vulgaris-leucoagglutinin demonstrated that axons had regenerated into the FSC-graft and had traveled longitudinally to the distal end of the channel. Few axons were observed to cross the distal host-graft interface to enter the host spinal cord. Cross-sectional analysis at the midpoint of the tissue cable stained with toluidine blue demonstrated a significant increase (P < 0.01) in myelinated axons in viable FSC grafts (1455 +/- 663, mean +/- S.E.M.; n = 6) versus freeze-thaw control grafts (155 +/- 50; n = 5). In addition to the myelinated axons, many unmyelinated axons were observed in the tissue cable at the electron microscopic level. Areas resembling the PNS with typical Schwann cells, as well as those resembling the central nervous system with neurons and central neuropil, were also seen. In freeze-thaw control grafts, neither viable neurons nor central neuropil were observed. Retrograde tracing with Fast Blue and Diamidino Yellow demonstrated that neurons within the FSC graft extended axons into the host spinal cord at least for 2 mm from both the rostral and caudal host graft interfaces. We conclude that viable FSC grafts within semipermeable guidance channels may serve both as a permissive bridge for longitudinally directed axonal growth and a potential relay for conveying information across a lesion site in the adult rat spinal cord. PMID- 10714265 TI - Comparison of the effects of electrolytic and chemical destruction of the red nucleus on the compensatory capacity of rats with rubrospinal tract lesions. AB - Transection of the rubrospinal tract in rats, performed before lesion of the red nucleus, resulted in the facilitated recovery of motor activity and operantly conditioned reflexes. Such facilitation was absent when the red nucleus is lesioned alone. This phenomenon is explained by the switching of descending influences on the corticospinal tract through the participation of the following system: red nucleus--inferior olive--cerebellum--ventrolateral thalamic nucleus- cerebral cortex. The above mentioned facilitating influence on the recovery process was particularly prominent in rats with quinolinic acid-induced lesion of the red nucleus. Under these conditions, the cerebellar ascending fibers to the ventrolateral thalamic nucleus were preserved. Decreased facilitated recovery following electrolytic lesion of the red nucleus suggests the existence of additional cerebello-cortical pathways for the realization of the switching phenomenon. PMID- 10714266 TI - Response features determining spike times. AB - Interpreting messages encoded in single neuronal responses requires knowing which features of the responses carry information. That the number of spikes is an important part of the code has long been obvious. In recent years, it has been shown that modulation of the firing rate with about 25 ms precision carries information that is not available from the total number of spikes across the whole response. It has been proposed that patterns of exactly timed (1 ms precision) spikes, such as repeating triplets or quadruplets, might carry information that is not available from knowing about spike count and rate modulation. A model using the spike count distribution, the low-pass filtered PSTH (bandwidth below 30 Hz), and, to a small degree, the interspike interval distribution predicts the numbers and types of exactly-timed triplets and quadruplets that are indistinguishable from those found in the data. From this it can be concluded that the coarse (< 30 Hz) sequential correlation structure over time gives rise to the exactly timed patterns present in the recorded spike trains. Because the coarse temporal structure predicts the fine temporal structure, the information carried by the fine temporal structure must be completely redundant with that carried by the coarse structure. Thus, the existence of precisely timed spike patterns carrying stimulus-related information does not imply control of spike timing at precise time scales. PMID- 10714267 TI - Temporal coding of periodicity pitch in the auditory system: an overview. AB - This paper outlines a taxonomy of neural pulse codes and reviews neurophysiological evidence for interspike interval-based representations for pitch and timbre in the auditory nerve and cochlear nucleus. Neural pulse codes can be divided into channel-based codes, temporal-pattern codes, and time-of arrival codes. Timings of discharges in auditory nerve fibers reflect the time structure of acoustic waveforms, such that the interspike intervals that are produced precisely convey information concerning stimulus periodicities. Population-wide inter-spike interval distributions are constructed by summing together intervals from the observed responses of many single Type I auditory nerve fibers. Features in such distributions correspond closely with pitches that are heard by human listeners. The most common all-order interval present in the auditory nerve array almost invariably corresponds to the pitch frequency, whereas the relative fraction of pitch-related intervals amongst all others qualitatively corresponds to the strength of the pitch. Consequently, many diverse aspects of pitch perception are explained in terms of such temporal representations. Similar stimulus-driven temporal discharge patterns are observed in major neuronal populations of the cochlear nucleus. Population-interval distributions constitute an alternative time-domain strategy for representing sensory information that complements spatially organized sensory maps. Similar autocorrelation-like representations are possible in other sensory systems, in which neural discharges are time-locked to stimulus waveforms. PMID- 10714268 TI - Intrinsic and extrinsic neuronal mechanisms in temporal coding: a further look at neuronal oscillations. AB - Many studies in recent years have been devoted to the detection of fast oscillations in the Central Nervous System (CNS), interpreting them as synchronizing devices. We should, however, refrain from associating too closely the two concepts of synchronization and oscillation. Whereas synchronization is a relatively well-defined concept, by contrast oscillation of a population of neurones in the CNS looks loosely defined, in the sense that both its frequency sharpness and the duration of the oscillatory episodes vary widely from case to case. Also, the functions of oscillations in the brain are multiple are not confined to synchronization. The paradigmatic instantiation of oscillation in physics is given by the harmonic oscillator, a device particularly suited to tell the time, as in clocks. We will thus examine first the case of oscillations or cycling discharges of neurones, which provide a clock or impose a "tempo" for various kinds of information processing. Neuronal oscillators are rarely just clocks clicking at a fixed frequency. Instead, their frequency is often adjustable and controllable, as in the example of the "chattering cells" discovered in the superficial layers of the visual cortex. Moreover, adjustable frequency oscillators are suitable for use in "phase locked loops" (PLL) networks, a device that can convert time coding to frequency coding; such PLL units have been found in the somatosensory cortex of guinea pigs. Finally, are oscillations stricto sensu necessary to induce synchronization in the discharges of downstream neurones? We know that this is not the case, at least not for local populations of neurones. As a contribution to this question, we propose that repeating patterns in neuronal discharges production may be looked at as one such alternative solution in relation to the processing of information. We review here the case of precisely repeating triplets, detected in the discharges of olfactory mitral cells of a freely breathing rat under odor stimulation. PMID- 10714269 TI - Oxidative stress, growth factor starvation and Fas activation may all cause apoptosis through lysosomal leak. AB - Oxidative stress, growth factor starvation, and activation of the Fas/APO-1/CD95 receptor all induce apoptosis in a variety of cell-types, including the established human Jurkat T-cell line. Oxidative stress, in the form of exposure of the cells to a bolus dose of hydrogen peroxide, results in intralysosomal, iron-catalyzed oxidative reactions. This is accompanied by a time- and dose dependent lysosomal destabilization--as evaluated by a decreased lysosomal uptake of the metachromatic fluorochrome, and weak base, acridine orange--in combination with leakage to the cytosol of lysosomal contents, including hydrolytic enzymes. Moderate lysosomal rupture is followed by apoptosis within initially intact plasma membranes, while necrosis and cell lysis are associated with a more complete lysosomal breach. Prior endocytosis of the potent iron-chelator desferrioxamine, resulting in binding of intralysosomal low molecular weight iron in a non-redox active form, largely prevents not only oxidative stress-induced lysosomal labilization, but apoptosis as well. When apoptosis is induced by the use of a monoclonal IgM anti-human Fas/APO-1/CD95 receptor antibody, the apoptotic process is again found to be accompanied by lysosomal leak. It is, however, not prevented by a preceding endocytosis of desferrioxamine and, consequently, could not be a function of intralysosomal iron-catalyzed oxidative reactions, but must be due to other mechanisms. Growth factor starvation of Jurkat cultures for a few days results in a high proportion of apoptotic cells, which contain lysosomes many of which have lost their proton gradient and appear to have released their contents. Overall, our results indicate that lysosomal leakage/rupture precedes apoptosis in Jurkat cells regardless of the initiating agent, but that such rupture may occur through multiple mechanisms. Lysosomal enzymes, leaking out of their normal vacuolar compartment, may then induce apoptosis, perhaps by proteolytic activation of the caspase-family of enzymes. Regardless of the precise mechanism, these observations suggest that partial rupture of the acidic vacuolar compartment may be one of the final pathways in apoptosis. PMID- 10714270 TI - Antioxidant properties of flavonol glycosides from tea. AB - We have determined the antioxidant activity of the major flavonols found in tea: a monoglycoside, a diglycoside and two triglycosides of kaempferol and three monoglycosides, a diglycoside and two triglycosides of quercetin. The Trolox equivalent antioxidant capacity (TEAC) and inhibition of iron/ascorbate-induced lipid peroxidation of phosphatidyl choline vesicles were measured. In the aqueous phase TEAC assay, the quercetin monoglycosides and diglycoside were approximately half as effective as quercetin aglycone. The quercetin triglycosides were much less effective than the monoglycosides and the diglycoside. The kaempferol glycosides were 32-39% less effective in the aqueous phase antioxidant assay compared to the kaempferol aglycone. Quercetin monoglycosides and diglycoside were potent inhibitors of lipid peroxidation, in contrast to the triglycoside which was much less effective. All the kaempferol glycosides were significantly less potent inhibitors of lipid peroxidation compared to the kaempferol aglycone. The compounds described herein demonstrate the antioxidant activity of the major flavonols in tea and indicate the effect of substituting a range of sugar moieties in the phenolic C ring. PMID- 10714271 TI - Effect of dialysis on oxidative stress in uraemia. AB - It has been postulated that dialysis of patients with chronic renal failure (CRF) is associated with increased lipid peroxidation which may contribute to vascular and other complications of the syndrome. In the present study, a specific and precise technique [ferrous oxidation in xylenol orange (FOX) assay] was used to measure plasma lipid hydroperoxides (ROOHs) in three groups of uraemic patients. Patients were either studied before starting dialysis (n = 12) or on continuous ambulatory peritoneal dialysis (CAPD, n = 12) or haemodialysis (HD, n = 36) and compared to healthy controls (n = 20). Plasma ROOHs were markedly elevated in HD patients compared with the controls (7.01 +/- 2.9 microM versus 4.25 +/- 2.05 microM; P < 0.005, Mann-Whitney test). Plasma ROOH concentrations in the CAPD patients were increased but not significantly higher than controls (5.36 +/- 3.56 microM versus 4.25 +/- 2.05 microM). By contrast, no differences in ROOH levels were found between controls and predialysis patients. There was no difference in plasma thiobarbituric acid reactive substances (TBARS) between control and the three CRF groups. Absolute and cholesterol standardised plasma alpha-tocopherol levels were lower in the patients (whether they were on dialysis or not) than in the controls (18.62 +/- 6.88 microM versus 22.73 +/- 5.33 microM; P < 0.01 and 1.99 +/- 1.88 microM/mM versus 5.25 +/- 1.0 microM/mM; P < 0.0005, respectively). This study provides direct evidence that enhanced oxidative stress in CRF patients is related to the dialysis treatment rather than the disease itself. Further studies will be necessary to establish the relationships between plasma measures of oxidative stress and cardiovascular complications in CRF patients under dialysis and whether treatment with antioxidants may reduce oxidative stress or reverse adverse effects associated with dialysis. PMID- 10714273 TI - Catalase activity and hydrogen peroxide levels are inversely correlated in maize scutella during seed germination. AB - Temporal patterns of hydrogen peroxide (H2O2) levels and total catalase activity are presented for post-imbibition scutella from six maize inbred lines expressing variable catalase activity. In all lines examined, H2O2 levels were highest during the initial days post-imbibition (1-2 dpi) and decreased thereafter, while total catalase activity was lowest during early dpi (1-2 dpi) and reached maximal activity at 4-6 dpi. In three of the six lines tested, a simple inverse correlation between catalase activity and H2O2 level was significant by Spearman's rank (P < 0.01). In addition to the general decline in H2O2 level throughout the dpi period, a reproducible increase in H2O2 level was observed at 4-5 dpi in five of six lines examined. Mutant lines lacking CAT-3 activity demonstrated a temporal shift in the occurrence of this increase. The role of total catalase (and individual isozymes) in controlling H2O2 levels during germination and the role of H2O2 as a potential regulator of catalase expression during germination are discussed. PMID- 10714272 TI - Quinone reductase (NQO1), a sensitive redox indicator, is increased in Alzheimer's disease. AB - NAD(P)H:quinone oxidoreductase 1 (NQO1), a redox-regulated flavoenzyme, plays a central role in monitoring cellular redox state. NQO1 acts to protect against oxidative stress induced by a variety of metabolic situations, including metabolism of quinones and other xenobiotics, by: (i) functioning as a two electron donor to provide a shunt that competes with the formation of reactive oxygen species; (ii) maintaining reduced coenzyme Q; and (iii) regulating the stress activated kinase pathway. In Alzheimer's disease, while there is abundant evidence for the involvement of oxidative stress, the cause or the consequences are largely unresolved. We suspected that increased NQO1 could signal a major shift in redox balance in Alzheimer's disease and, in this study, found that NQO1 is localized not only to neurofibrillary tangles but also the cytoplasm of hippocampal neurons. By marked contrast, there is very little NQO1 in the same neuronal populations in young and age-matched controls. This novel association of NQO1 further buttresses the nexus of oxidative stress, via free radicals, with selective neuronal vulnerability and also supports a fundamental abnormality in redox balance in Alzheimer's disease. PMID- 10714275 TI - 2,4-dinitrophenylhydrazine carbonyl assay in metal-catalysed protein glycoxidation. AB - Using an experimental in vitro glycation model, long-term incubations of bovine serum albumin with glucose (fructose) resulted in a significant increase in protein content of 2,4-dinitrophenylhydrazine (DNPH)-reactive carbonyl groups, which could be strongly inhibited by anaerobiosis and metal chelation. The pattern of yields of the protein-bound DNPH was not in accordance with that of the sugar-derived carbonyls determined as the ketoamine Amadori product. In spite of the fact that the contribution of the final advanced glycation end-products to the total DNPH-reactivity of glycation-altered protein remains unclear, the present results stress the need of oxidative steps in formation of most of the DNPH-reactive carbonyl compounds generated by glycation. The results provide evidence that, in protein glycoxidation, the DNPH assay is selective enough to discriminate between protein-bound carbonyls produced by metal-catalysed oxidations and those formed in the early glycation steps. PMID- 10714274 TI - Bioflavonoid effects on the mitochondrial respiratory electron transport chain and cytochrome c redox state. AB - The polyphenolic structure common to flavonoids enables them to donate electrons and exert antioxidant activity. Since the mitochondrial electron transport chain consists of a series of redox intermediates, the effect of flavonoids in a complex mixture of polyphenols, as well as related pure flavonoids, was evaluated on the rat liver mitochondrial electron transport chain. A French maritime pine bark extract (PBE), a complex mixture of polyphenols and related pure flavonoids, was able to reduce cytochrome c reversibly, possibly by donation of electrons to the iron of the heme group; the donated electrons can be utilized by cytochrome c oxidase. Among single flavonoids tested, (-)-epicatechin gallate had the greatest ability to reduce cytochrome c. In addition, PBE competitively inhibited electron chain activity in both whole mitochondria and submitochondrial particles. A 3.5 fold increase in the apparent Km value for succinate was calculated from reciprocal plots. Among the flavonoids tested, taxifolin and (-)-epicatechin gallate showed minor inhibitory effects, while (+/-)-catechin and (+)-epicatechin were ineffective. Activities of NADH-ubiquinone, succinate-ubiquinone, and ubiquinol-cytochrome c reductases were inhibited by low concentrations of PBE to a similar extent. However, inhibition of cytochrome c oxidase activity required 4 fold higher PBE concentrations. These results suggest that flavonoids reduce cytochrome c and that PBE inhibits electron transport chain activity mainly through NADH-ubiquinone, succinate-ubiquinone, and ubiquinol-cytochrome c reductases. PMID- 10714276 TI - Hydrogen peroxide and catalase are inversely related in adult patients undergoing cardiopulmonary bypass: implications for antioxidant protection. AB - Adult patients undergoing cardiopulmonary bypass (CPB) surgery are subjected to increased oxidative stress and show a spectrum of lung injury. Increased levels of hydrogen peroxide (H2O2) are often seen during episodes of oxidative stress, such as the use of high FiO2s, and this molecule plays a key role in the formation of highly damaging oxidants such as the hydroxyl radical. Oxidative damage to plasma proteins was assessed by measuring free thiol groups, and antioxidant protection against H2O2 by measuring catalase activity. CPB patients (n = 39) receiving either 100% or 50% oxygen at the end of bypass were studied by measuring levels of H2O2 in breath condensate and levels of catalase in their plasma, and comparing these to pre-bypass levels. Post-bypass, all CPB patients exhaled significantly lower levels of H2O2 (P < 0.0001) at a time when they had significantly increased activity (0.809 +/- 0.11 versus 1.688 +/- 0.18 U/mg protein) of catalase in their plasma. There were no significant differences in these parameters between the 100% and 50% oxygen groups. At a time when oxidative stress is greatest, there appears to be a corresponding plasma increase in the antioxidant catalase. Whether this change is fortuitous or a response to oxidative stress is at present under consideration. PMID- 10714277 TI - Blood glutathione homeostasis as a determinant of resting and exercise-induced oxidative stress in young men. AB - Although the importance of glutathione in protection against oxidative stress is well recognized, the role of physiological levels of glutathione and other endogenous antioxidants in protecting against exercise-induced oxidative stress is less clear. We evaluated the role of glutathione and selected antioxidant enzymes as determinants of lipid peroxidation at rest and in response to exercise in men (n = 13-14) aged 20-30 years, who cycled for 40 min at 60% of their maximal oxygen consumption (VO2max). Levels of plasma thiobarbituric acid reactive substances (plasma TBARS) and blood oxidised glutathione (GSSG) increased by about 50% in response to exercise. Mean blood reduced glutathione (GSH) decreased by 13% with exercise. Of the measured red blood cell (RBC) antioxidant enzyme activities, only selenium-dependent glutathione peroxidase (Se GPX) activity rose following exercise. In univariate regression analysis, plasma TBARS levels at rest predicted postexercise plasma TBARS and the exercise-induced change in total glutathione (TGSH). Blood GSSG levels at rest were strongly determinant of postexercise levels. Multiple regression analysis showed blood GSH to be a determinant of plasma TBARS at rest. The relative changes in TGSH were determinant of postexercise plasma TBARS. In summary, higher blood GSH and lower plasma TBARS at rest were associated with lower resting, and exercise-induced, lipid peroxidation. Subjects with a favourable blood glutathione redox status at rest maintained a more favourable redox status in response to exercise-induced oxidative stress. Changes in blood GSH and TGSH in response to exercise were closely associated with both resting and exercise-induced plasma lipid peroxidation. These results underscore the critical role of glutathione homeostasis in modulating exercise-induced oxidative stress and, conversely, the effect of oxidative stress at rest on exercise-induced changes in glutathione redox status. PMID- 10714278 TI - Antioxidant activity of palm oil carotenes in peroxyl radical-mediated peroxidation of phosphatidyl choline liposomes. AB - The antioxidant efficacy of alpha-carotene and comparison with beta-carotene in multilamellar liposomes prepared from egg yolk phosphatidyl choline (EYPC) exposed to the lipid soluble 2,2'-azobis (2,4-dimethyl valeronitrile) (AMVN) was investigated. Lipid peroxidation was measured as thiobarbituric acid reacting substances (TBARS) at 532 nm or as hydroperoxide formation at 234 nm after separation of phosphatidyl choline hydroperoxide (PCOOH) by high-pressure liquid chromatography (HPLC). Lutein and zeaxanthin, the hydroxyl derivatives of alpha- and beta-carotenes, and the chain breaking antioxidant alpha-tocopherol were also included in the study. AMVN being a lipid soluble, non polar azo initiator penetrates into the hydrophobic interior of the phospholipid bilayer, forming peroxyl radicals which peroxidate the phospholipid leading to PCOOH accumulation. All the carotenoids tested at 1 mol% relative to EYPC significantly suppressed the formation of PCOOH compared to control samples. In this system, alpha carotene retarded PCOOH formation better than beta-carotene. Similarly, lutein was a better antioxidant than is zeaxanthin. But lutein and zeaxanthin were more effective antioxidants than alpha- and beta-carotenes, respectively. After 1 h of incubation of the carotenoid with AMVN, alpha-, beta-carotene, lutein and zeaxanthin limited PCOOH formation by 77%, 68%, 85% and 82%, respectively, while alpha-tocopherol elicited 90% reduction. AMVN incubated with EYPC for 2 h induced the formation of TBARS compared to control (P < 0.001). alpha-Carotene significantly suppressed the TBARS formation by 78% whilst beta-carotene, lutein, zeaxanthin and alpha-tocopherol elicited 60%, 91% and 80% reductions, respectively. Increasing the concentration of the carotenoid > 1 mol% to EYPC did not significantly increase protection of the membrane against free radical attack. Our findings suggest that alpha-carotene is a better antioxidant than is beta-carotene in phosphatidyl choline vesicles. It may, therefore, be useful in limiting free radical mediated peroxidative damage against membrane phospholipids in vivo. PMID- 10714279 TI - Measurement of relative antioxidant activity of compounds: a methodological note. AB - The relative activities of some hydrogen-donating antioxidants were assessed by comparing their activities with that of Trolox (Trolox equivalent antioxidant capacity, TEAC) for scavenging the ABTS radical cation (ABTS.+) generated in the aqueous phase. We have verified, however, that TEAC values may change with the concentration of compounds and with the measuring times used. Not withstanding, TEAC values do not differ significantly if the compounds have kinetic curves of ABTS.+ formation similar to that of Trolox. This is the case with ascorbic acid, whose TEAC values, determined by using five concentrations at three different measuring times, are very close. For the flavonoids studied (catechin, rutin, naringenin and silibinin) which have kinetic curves of ABTS.+ formation different from that of Trolox, the TEAC values decrease with increasing concentrations of the compounds for each measuring time, and increase with increasing measuring times for each concentration. In the present study, we conclude that, in order to evaluate relative antioxidant activities of compounds by the ABTS assay, it is essential to perform kinetic studies to assess scavenging of ABTS.+ by these compounds. Therefore, when the TEAC values of compounds are determined for more than one measuring time, we may be sure that all the antioxidant potential of compounds is being considered and whether or not it is possible to establish a hierarchy for their antioxidant activities. PMID- 10714280 TI - [Frontal lobe dysfunction in patients with epilepsy and chronic psychosis]. AB - PATIENTS AND METHODS: Fifty patients diagnosed as having epileptic crises were given neuropsychological tests including the Wechsler Adult Intelligence Scale or WAIS, Luria's neuropsychological test and a quantitative EEG examination. Multivariate analysis was done of the following variables: presence or absence of inter-octal psychosis, onset of crises before the age of 10 years, frequency of crises or status epilepticus greater than 100, known cause or otherwise of epilepsy, and the presence of more than one type of crisis in a particular patient. The working hypothesis was to show that the association of epilepsy and psychosis causes alterations in superior psychic functions (SPF) particularly of the frontal lobes. The WAIS test, intelligence, verbal and executive quotients and the 11 subtests were evaluated using multivariate analysis (ANOVA) conditioned by the different variables studied. The broad band spectral measurements of the quantitative EEG (BBSM) were studied using a statistical programme (COMPARA) by which the groups of individuals were compared with a standard group, using the Student t and Fisher tests. The different BBSM variables studied were: absolute power, relative power and total dominant frequency. RESULTS: Amongst the most important results were: reduction in the performance scale of epileptics with chronic psychosis, alterations on the verbal scale in epileptics with more than one type of crisis, presence of frontal and fronto-temporal dysfunction in epileptics with chronic psychosis and negative signs of schizophrenia. On the quantitative EEG in epileptics with psychosis there was abnormally slow activity predominantly in the frontal lobes. CONCLUSIONS: From the overall results we may conclude that in patients with epilepsy and chronic psychosis there is cortical dysfunction of the frontal lobe. PMID- 10714281 TI - [The use of zygomatic electrodes in the assessment of epileptic patients. Presentation of methodology for recording and assessing a digital EEG]. AB - INTRODUCTION AND OBJECTIVES: Electrographic anomalies of the temporal lobe may be seen in 70% of epileptic patients. Focalization if inter-ictal epileptiform discharges makes a major contribution to clinical diagnosis. Also, it is accepted that the detection and interpretation of such anomalies is influenced by the method of recording. Thus the use of additional extracranial electrodes has been shown to significantly increase the sensitivity of inter-ictal or ictal electroencephalograms (EEG) PATIENTS AND METHODS: We present the results of two years work in the Electroencephalogram Laboratory of the International Centre for Neurological Recovery, La Habana, Cuba (CIREN), using zygomatic electrodes for the standard assessment of epileptic patients. Recordings were made using the FCz position as reference electrode instead of the reference electrode (short circuited ears) used as pre-programmed in the recording module of the software Track Walker 2 for Medicid 3E (used in the Clinical Neurophysiology Laboratories of the National Network). Epileptiform activity was seen on 196 recordings; in 100 (51%) of these, this activity involved the temporal lobe, with strictly temporal localization shown in 25 patients (25%) and in 7 (28%) we recorded focalization on the medial aspect. We present segments of EEGs with epileptiform discharges focalized in the temporal lobe (medial and lateral aspects) in bipolar systems (zygotemporal-parasagittal) and in reference electrodes. CONCLUSIONS: Our results support the usefulness of this methodology for the detection and localization of epileptiform activity in the temporal lobe and suggest an alternative which would increase the diagnostic sensitivity of EEG in epilepsies. PMID- 10714282 TI - [Reference values for the central motor conduction time and silent period obtained by the transcranial magnetic stimulation]. AB - INTRODUCTION: Normal values of reference to transcranial magnetic stimulation for the motor central conduction time (CCT) and silent period (SP) is recorded in 30 healthy control subjects over abductor pollicis brevis. MATERIAL AND METHODS: We get for the CCT four measurements: two with low intensity of stimulus, 5% plus the motor threshold, with and without facilitation (CCT1 and CCT1 fac.); and two with high intensities of stimulus, elevating the magnetic stimulation intensity to 1.5 times the threshold (CCT2 and CCT2 fac.). RESULTS AND CONCLUSIONS: The mean and standard deviation of each measurement are: CCT1: 9.34 +/- 1.19, CCT1 fac.: 7.12 +/- 1.1. CCT2: 8.84 +/- 1.05 and CCT2 fac.: 6.57 +/- 1.05. Given that the CCT and SP doesn't follow a normal distribution, the medium and the 5-95% percentiles for the normal values of reference are calculated; there are: CCT1: 7.15-11.32, CCT1 fac: 5.27-9.42. CCT2: 7.05-10.73 and CCT2 fac: 4.91-9.14. For the silent period gets only one measurement employing high intensities. These last measurement were recorded in two localizations: on vertex and on motor area, selecting the greater duration. Given the great individual variability of this period in normal population absolute and ratio for the difference duration of SP between both sides are calculated. The latency of the SP is 50.2 +/- 5.99, 95 percentiles 39.1-64.63, the duration 151 +/- 32.51, 95 percentiles 102.63-239.55. The total SP measured from the discharge of the stimulus to the end of the silent period is 201.71 +/- 33.27; 95 percentiles: 151.39-296.4. The comparison of both hemispheres would give us pathological security for the 99.99% of the population for more than 14.94 ms of absolute difference, and for less of the 79.81% of ratio difference. A summary of the discoveries of the silent period in different pathologies is contributed in the discussion. PMID- 10714283 TI - [Neuropathy of leprosy: characteristics of cases in 1962-1995]. AB - INTRODUCTION: Neuropathy due to leprosy is the most frequent cause of peripheral nervous system disorders due to an infective agent, one of the commonest aetiologies of peripheral neuropathy, and also one of the few peripheral neuropathies which is curable. Hansen's disease, initially and predominantly involves the skin, so it is not usually seen by a neurologist; a neurologist sees cases which are difficult to diagnose and often after the condition has been ruled out by other doctors from other medical specialties. PATIENTS AND METHODS: In the National Institute of Neurology and Neurosurgery of La Habana, Cuba, 18 nerve biopsies were studied (during its history), that had been diagnosed as leprous neuropathy. We reviewed the clinical histories of all the patients admitted to this centre. The sample was characterized by: age, sex, neurological clinical picture, nerves most affected, skin lesions, electrophysiological studies and a description of the anatomopathological findings. RESULTS AND DISCUSSION: Most patients were male and all (100%) were adults of 'working age' (19-65 years). The most frequent neuropathic pattern was multiple mono-neuropathy (78%) and the quality, purely sensitive or mixed was predominantly sensitive (100%). The nerves most affected were the ulnar (10 patients) and median (6 patients). The most frequent skin lesions were anesthetic maculae. Anatomo pathological study showed inflammatory infiltration in all patients and the bacillus was absent in only one case. PMID- 10714284 TI - [The Guayaquil Declaration]. PMID- 10714285 TI - [Pain]. AB - This text reports the 'introduction remarks' of Professor Ochoa at the inauguration of a Symposium on Pain at the Spanish Neurology Society. Profoundly, but as clearly as possible, the author expounded on the nociceptors to the cerebral cortex, the structures involved in senso-perception--and emotion--which are to be taken into consideration. Some basic facts are: 1. The functional characterization of different nociceptors. 2. The existence of usually 'silent' nociceptors which are only activated under certain conditions. 3. The enumeration of the zones of the posterior horn which are 'specific' for nociception: lamina I and the external zone of lamina II. 4. The great feature of lamina V, with a population of convergent neurones, which receive both tactile and nociceptive stimuli. 5. The contrast between the postero-ventral thalamic nucleus (touch, pain, etc.) and the internal or 'medial' thalamic sectors, which are involved in the most crude and aggravating pain and in the course of impulses which predispose to pathological emotional experiences. This thalamic sector does not generate pain when stimulated under normal conditions, but may do so when the patient has pain of central origin, in which case the condition--sometimes complex--of desafferentization may be assessed. We point out that both the classically named causalgia and the central algias (thalamic, due to pathology of the brainstem and cortico-subcortical lesions) have similar aspects: burning pain with paroxystic exacerbations, sensory defects, hyperpathia and adodynia. We also mention the complex inhibitory effects of impulses conducted by large diameter myelinated fibres or nociceptive effects at different levels. Reference was not made to centrifuge systems (endogenous opiates etc.) which inhibit pain. References 1, 3, 6 and 9 include a considerable quantity of bibliography. PMID- 10714286 TI - [Typical absence epilepsy in a patient with serious cranio-encephalic trauma]. AB - INTRODUCTION: Head injury is the commonest cause of symptomatic or secondary epilepsy and one of its most serious sequelas. Typical absence seizures are well defined clinically and electroencephalographically and are seen in age-related idiopathic epilepsies. There are very few descriptions of seizures of typical absences that were symptomatic of tumors or other structural lesions. CLINICAL CASE: We describe the case of a nine year old boy who had had a severe head injury at the age of four years. When he was seven years old he started to have seizures with all the clinical and electroencephalographic features of typical absences. CONCLUSIONS: In this case, taking the age of the patient into account, the APF, APP, electroclinical characteristics of the seizures, neurological and clinical condition, the problem was to decide whether the seizures were idiopathic or symptomatic of a cerebral lesion. This was important for treatment and prognosis. The answer could only be obtained by follow-up and assessment of the response to specific treatment for petit mal. PMID- 10714287 TI - [Alexia without agraphia due to the lesion in the right occipital lobe in a right handed man. Detection of hemispheric lateralization of handedness and language in a right-handed patient]. AB - INTRODUCTION AND CLINICAL CASE: A 65 year-old man, right-handed, without any family history of left handiness, suddenly developed a left homonymous hemianopia and incapacity for reading. Neurological and neuropsychological examinations showed the presence of a profound alexia with preservation of writing to dictation and spontaneously. He was unable to read what he had written. He could spell the words letter by letter but he was unable to read the complete word. MRI showed an extensive infarct in the territory of the right posterior cerebral artery. The infarct extended anteriously to the right thalamus and to the medial temporal fifth or fusiform gyrus. The splenius was spared. Brain SPECT disclosed the area of the infarct and an extensive area of decreased cerebral perfusion over the right parietal and temporal areas. CONCLUSION: Alexia without agraphia has been reported in right-handed patients with left occipital lesions and in right occipital regions in left-handed patients but rarely if ever in right occipital lesions in right-handed patients. PMID- 10714288 TI - [Neurofibroma with mucus-producing glands. Report of a case and literature review]. AB - INTRODUCTION: Most tumors of peripheral nerve sheaths containing glands are malignant tumors associated with Von Recklinghausen's disease. CLINICAL CASE: A 39 year old man consulted with a tumour on a finger of the right hand, which was not painful, and was slow growing. There was no past history of neurofibromatosis. Histological study showed a tumour of the peripheral nerve sheath, a benign type of neurofibroma containing glands. Immunohistochemical techniques confirmed that it had the stroma of a Schwannoma with well-defined glandular epithelial elements. CONCLUSIONS: A neurofibroma with glands is considered to be a rare type of divergent differentiation, and of considerable interest to pathologists, since it must be differentiated from other tumoral lesions. Immunohistochemical study is very useful for this. PMID- 10714289 TI - [Paroxysmic anarthria in multiple sclerosis]. AB - INTRODUCTION: The paroxystic clinical features of multiple sclerosis (MS) include trigeminal neuralgia, itch, transient diplopia, Lhermitte's sign, akinesia, dystonia, Uhthoff's phenomenon and others which are very characteristic, such as paroxystic ataxia and dysarthria. CLINICAL CASE: We present the case of a 30 year old man who consulted for multiple episodes lasting only a few seconds, of complete inability to speak. This symptom recurred several times a day and in many different situations. It was often triggered off by external stimuli such as having to speak in front of several people. The disorder disappeared without treatment seven days after onset. Magnetic resonance using fast spin echo image sequences showed multiple hyperintense lesions in mid right cerebellar peduncle, right pons, left temporal lobe, white substance of both internal capsules, periventricular and semioval centres. Biochemical study of the cerebrospinal fluid showed that there were 9 cells/microliter (mainly lymphocytes), proteins 45 mg/dl and a normal glucose level. The Tibling-Link level was 0.73. Cortical somestesic evoked potentials showed slowed conduction after stimulation of the right median nerve and both peroneal nerves. Acoustic evoked potentials of the brain stem were conducted more slowly by the right acoustic pathway at intraxial level. The patient was diagnosed as having clinically defined MS. CONCLUSIONS: We consider that our patient's symptom was a kind of paroxystic dysarthria which we call paroxystic anarthria. Differential diagnosis of this symptom should be basically with phonatory or dysphasic simple partial seizures. PMID- 10714290 TI - [Chlamydia pneumoniae and arteriosclerosis]. AB - INTRODUCTION: The relationship between Chlamydia pneumoniae and atherosclerosis has been assessed in several studies. Development and conclusions. Their results suggest a possible role of Chlamydia pneumoniae in the origin and development of the disease, while the actual link between them are still unknown. However, the presence of Chlamydia pneumoniae in patients with atherosclerosis could be just a coincidence. PMID- 10714291 TI - [Current management of carotid artery stenosis. What does the evidence show us?]. AB - OBJECTIVE: Surgery of carotid artery stenosis as treatment for acute cerebral vascular accident was most used in the 1980s. This surgical procedure is one of the few to have been 'examined' to demonstrate its usefulness by means of a series of prospective multicentric trials, carried out between 1980 and 1990. This paper aims to show the results of these trials and give useful advice for current management of these patients. DEVELOPMENT: We describe the most important studies carried out to date, both in the group of patients with symptomatic and those with asymptomatic carotid artery stenosis. We emphasize the importance of achieving low morbi-mortality, both in the surgical procedure and in diagnosis. We describe the advantages and disadvantages of non-invasive methods of diagnosis, alternatives to cerebral angiography, and discuss current indications for carotid artery surgery in symptomatic and asymptomatic groups. CONCLUSIONS: The most important cooperative studies (ECST and NASCET) have shown the validity of surgical in combination with medical treatment, as opposed to medical treatment alone, in order to avoid further neurological incidents in patients with symptomatic carotid stenosis. The only cooperative study which has shown the superiority of surgical treatment, as compared with medical treatment, in a group of asymptomatic patients was ACAS (particularly in male patients). PMID- 10714292 TI - [The report of the 36th Annual Reunion of the Spanish League against Epilepsy]. PMID- 10714293 TI - [International environment of the Spanish League against Epilepsy]. PMID- 10714294 TI - [The latest nosological advances and treatment of progressive myoclonus epilepsies]. PMID- 10714295 TI - [Severity and epidemiology of myoclonic epilepsy]. AB - The author first reviews the definition of myoclonia as an epileptic crisis differentiated from tonic crises and infantile spasms. He reviews the prevalence and incidence found in bibliographic data, under the following headings 1. Early myoclonic encephalopathy or neonatal myoclonic encephalopathy 2. Early epileptic syndrome with bursts of suppression or Otahara's syndrome. 3. West's syndrome. 4. The benign myoclonic epilepsy syndrome of children. 6. Syndrome of myoclonic epilepsy in non-progressive encephalopathy. 7. Early myoclonic epilepsy of children or Dose's syndrome. 8. Lennox-Gastaut syndrome. 9. Syndrome of epilepsy with absences in children. 10. Myoclonic absence epilepsy syndrome. 11. Landau Kleffner syndrome and the syndrome of continuous slow spike-and-wave epilepsy during slow sleep. 12. Photosensitive epilepsy. 13. Absence epilepsy in young patients. 14. Juvenile myoclonic epilepsy. 15. Syndrome of gran mal epilepsy on waking. 16. Progressive myoclonic epilepsies. The author reviews 6,450 cases, 408 patients who had myoclonic crises, that is 6.3%. The differences seen in this total group of patients were: the myoclonic crises which presented alone, myoclonic crises accompanied by simple typical absences, those initially accompanied by generalized tonic-clonic crises and those presenting typical absences, tonic-clonic generalized crises and myoclonus simultaneously. The course of the different groups is analyzed. PMID- 10714296 TI - [Neurophysiological classification of myoclonus]. AB - Myoclonus is defined as a brief, jerky movements of one or several muscular groups, generated in the central nervous system. Myoclonus can be the manifestation of an extensive variety of pathological conditions with different anatomical substrates and physiopathological mechanisms. According to the physiopathology, four myoclonus types are distinguished: cortical, subcortical, corticosubcortical and spinal myoclonus. Its generators, and the neurophysiological approaches are discussed. PMID- 10714297 TI - [Myoclonus and myoclonic epilepsies in childhood]. AB - Myoclonic jerks occur in a number of different syndromes. There is many classifications of myoclonus. It is preferred the Fejerman classification, slightly modified that present the following five groups: 1. Myoclonus without encephalopathy and without epilepsy, which includes physiological myoclonus; 2. Encephalopathies with non epileptic myoclonus, which includes Kinsbourne syndrome and certain types of hyperekplexia which pose differential diagnosis problems with reflex myoclonic epilepsy; 3. Progressive encephalopathies with myoclonic seizures which includes typical and atypical progressive myoclonus epilepsies; 4. Epilepsies and epileptic encephalopathies with myoclonic seizures, which includes severe epilepsies which leads to mental retardation, as Otahara syndrome, West syndrome and Lennox-Gastaut syndrome, and other epilepsies which present sometimes myoclonic seizures, as Landau-Kleffner syndrome, 5. Comprises true myoclonic epilepsies, differentiating syndromes recognized as idiopathic, -benign myoclonic epilepsy of infancy, reflex form of benign myoclonic epilepsy in infancy, eyelid myoclonic with absences, perioral myoclonic with absences and juvenile myoclonic epilepsy-, cryptogenic-severe myoclonic epilepsy of infancy, myoclonic-astatic epilepsy and epilepsy with myoclonic absences-, and symptomatic as the generalized myoclonus in children with static encephalopathies. The epileptic syndromes of the last group are described. Despite this classification, apparently clear, there is still a great deal of confusion and in clinical practice, many cases are difficult to classify. PMID- 10714298 TI - [Myoclonic epilepsies in adolescent and adult patients]. AB - We review the electroclinical characteristics of the main myoclonic epileptic syndromes occurring in adolescents and adults, excepting the progressive myoclonic epilepsies. In the discussion we include some epilepsies which are not currently classified as generalized, but which develop myoclonic crises such as the myoclonic variant of epilepsy when reading. We emphasize strict criteria for diagnosis of juvenile myoclonic epilepsy and discuss the syndromes related to it. Other myoclonic epilepsies, such as those which may occur in adults with Down's syndrome and in Alzheimer's disease, are considered. PMID- 10714299 TI - [Vascular epilepsy in childhood]. AB - OBJECTIVE: To make a summary of the connection between seizures and vascular diseases in childhood. MATERIAL: It is mainly used the experience along a lot of years in our Pediatric Neurology Service. RESULTS: They are directly shown the results observed in every vascular disease using the types of study and therapy existing in every period. CONCLUSIONS: It is possible to appreciate the different capacity of every vascular disease to cause seizures of different type and its response to the therapy depending of the nature of the vascular pathology. PMID- 10714300 TI - [Epidemiology of various types of vascular epilepsy in adults]. AB - From studies of the incidence of epilepsy, figures of between 40 and 70 per 100,000 inhabitants are obtained whilst in studies of prevalence, 5 to 7 cases per 1,000 are found. Cerebrovascular disease is the commonest cause of epileptic crises in the developed world, being more frequent in patients aged over 60, who make up 50% of all cases. Depending on the moment of presentation of crises in relation to the cerebrovascular disease, these may be classified as herald crises or precursors of vascular epilepsy, early crises if they occur during the first week and late crises if they occur after. Whilst early crises are usually due to metabolic or cytotoxic factors, the late crises occur in true vascular epilepsy. When these patients are assessed it is also important to consider increased hospital mortality in those with early crises and possible deterioration of neurological deficit as sequelae following late epileptic crises. PMID- 10714301 TI - [Cerebral ischemia and epilepsy]. AB - INTRODUCTION: We review the characteristics and evolution of epileptic crises (EC) related to non-hemorrhagic ictus. Patients and methods. Since June 1994 we have studied patients with EC both at the time of the ictus (acute symptomatic crises, ASC) and later (remote symptomatic crises RSC). One hundred and fifteen fulfilled the criteria and were followed-up until recurrence of EC, death or the end of the study (30.06.98). There were 66 men and 49 women (average age at the time of ictus = 67.4 +/- 12 years). RESULTS: Ninety one patients had RSC; reversible ischemic neurological deficit (DNIR) (50%), atherothrombotic pathology (58.5%) and anterior territory (70%) predominated. There was a similar proportion of partial and generalized crises (51.5% compared with 48.5%). Fifteen patients had presented with ASC. Thirty-nine patients presented with ASC, with predominance of established ictus (48.5%), atherothrombotic pathology (56.5%), anterior territory (82%) and generalized crises (59%). There was recurrence in 50.5% of those with RSC (follow-up 18.5 +/- 24 months). STATISTICAL ANALYSIS: there was a predominance of ASC in patients with established ictus and RSC in the case of DNIR. In cases of abnormal EEG there was a greater proportion of patients with a history of ASC. In patients over 60 years old, CSR was commoner. In those with atherothrombosis there was a predominance of one crisis and in patients with embolisms two or more crises. There were more recurrences in patients with no previous history of ASC (p = 0.001), those with all the anterior territory affected (p = 0.002), those < 59 years old (p = 0.01), those previously untreated (p = 0.04) and those with abnormal EEG (p = 0.03). There was an increased RR in the abnormal EEG, involvement of the entire anterior territory and age < 59 years. Multivariate analysis showed that the probability of recurrence increased 1.23 times when there was a previous history of ASC; 14.73 times if the EEG was abnormal, and 18.12 times when both these factors were present. PMID- 10714302 TI - [Early presentation of crises and the development of epilepsy in cerebral intra parenchymatous hemorrhage]. AB - PATIENTS AND METHODS: Of a total of 283 patients with spontaneous or hypertensive cerebral intraparenchymatous hemorrhage, 18 (6.3%), with no previous epilepsy, had crises whilst being followed-up for a period of between 2 and 7 years. In 14 cases the hematoma was lobar and 4 involved the basal ganglia or thalamus. In 8 cases (2.8% of all hemorrhage), these crises occurred during the first 24 hours, or as a first symptom of intraparenchymatous hemorrhage. One patient presented with status epilepticus with generalized crises and two had subentrant secondarily generalized partial crises at the time of the ictus. Treatment with anti-epileptic drugs was started in 13 patients. Twelve patients (4.2% of the hemorrhages) developed symptomatic epilepsy with partial crises with or without secondary generalization. RESULTS AND CONCLUSIONS: The maximum rate of recurrence was four crises per year. However, in one patient, reduction of the dose of medication led to the appearance of status epilepticus. Patients with crises of late onset developed epilepsy more often than those who had early crises. In those with crises there was a predominance of bilobular involvement with participation of the parietal lobe and extension of the hematoma or oedema to the cerebral cortex. PMID- 10714303 TI - [Overall treatment of vascular epilepsy]. AB - Cerebrovascular disease has different acute, ischemic and hemorrhagic presentations and may be associated with epileptic crises during the acute phase, or a later epileptic syndrome may develop. Status epilepticus is an infrequent complication which may appear at any time during the course of the illness, sometimes as the first and only sign of epilepsy. The risk of acute crises or of an epileptic syndrome varies depending on the nature of the vascular accident: its occurrence is more likely in hemorrhagic lesions and in those involving the cerebral cortex. The acute crises may be treated with benzodiazepines or with fast acting antiepileptic drugs; parenteral administration may sometimes be necessary. The need for prolonged prophylactic antiepileptic treatment is still under discussion, since there is no evidence that this prevents later development of an epileptic syndrome. The management of status epilepticus is the same whatever the etiology, although one has to take account of the risk of side effects related to the age and general health of the patient. When deciding on treatment for vascular epilepsy consideration should be given not only to which drugs are to be used, but also their pharmacokinetic characteristics and interactions with any treatment required by the patient for coexisting conditions such as arterial hypertension, heart failure, anticoagulation, diabetes, etc. PMID- 10714304 TI - [Epilepsy and arteriovenous malformations]. PMID- 10714306 TI - [Kennedy disease. A report of two cases]. PMID- 10714305 TI - [Clinical implications of pharmacology and pharmacokinetics of tiagabine]. AB - Tiagabine is a new antiepileptic drug which acts by blocking neuronal and glial GABA uptake and it is indicated in the treatment of partial epilepsies. Its pharmacokinetics is lineal, being extensively metabolized in the liver by means of CYP3A4 isoenzyme. Plasma elimination half life ranges between 5-8 hours in healthy volunteers, being markedly reduced when the drug is administered concomitantly with enzyme-inducing antinconvulsants. Tiagabine does not induce nor inhibit hepatic enzymes and, consequently, it does not modify the kinetics of simultaneously prescribed antiepileptic drugs. No relevant kinetic differences have been observed between adults and elderly subjects. Renal impairment does not alter the pharmacokinetic profile of tiagabine; hepatic disease, however, significantly reduces tiagabine elimination and lower daily doses of the drug are necessary in these patients. Although tiagabine elimination half life is short, it has been ascertained that therapeutic efficacy is similar when administered in 2 or 4 divided doses. Tiagabine is usually well tolerated; its most frequent side effects include dizziness, asthenia, nervousness, tremor, diarrhea and depressed mood. PMID- 10714307 TI - [Diagnostic difficulties in Devic's neuromyelitis]. PMID- 10714308 TI - [A case of optic neuritis due to Treponema pallidum and treatment with megadoses of corticosteroids]. PMID- 10714309 TI - [Memory disorders in the epidemic neuropathy]. AB - INTRODUCTION: The epidemic neuropathy that occurred in Cuba between 1992-1993 had three clinical forms: optic, peripheral and mixed. Epidemic neuropathy patients often complained of disorders in the recent memory. PATIENTS AND METHODS: We studied 120 patients: 68 with peripheral form, 26 with the optic form and 26 of the mixed form and 70 healthy subjects as control group. The short term memory was evaluated with the paradigm of Brown and Petersen, task of free recall for verbal nonsense material, during several intervals of retention (0, 15, 30, 45 seconds). Also, we applied tests for general capacity, depression and a subjective questionnaire about neuropathic symptoms intensity. RESULTS: Significant differences were observed in the severity of the forgetfulness and depression level between the peripheral form patients and the others groups. The correlation between severity of visual symptoms and the forgetfulness were not significant (S = -0.05; t (176) = -0.7; NS), whereas there was a significant correlation between severity of neuropathic peripheral symptoms and the forgetfulness (S = 0.2; t (184) = 2.7; p < 0.008). This effect on the mediate memory was not observed in the group of visual neuropathy and only occurred, of attenuated form, in the mixed group. CONCLUSION: An explanation for this dissociation is suggested based on the physiopathogenic mechanisms invoked in the illness. PMID- 10714310 TI - [The Cambridge Cognitive Examination as a tool for detection of dementia]. AB - INTRODUCTION: The efficiency of Cambridge Cognitive Examination' (CAMCOG) is analyzed as a tool to detect dementias in epidemiological studies. PATIENTS AND METHODS: The data were obtained from subjects who enrolled the second phase on a door-to-door field epidemiological study. The tool used was the 'Cambridge Mental Disorders of the Elderly Examination' (CAMDEX). RESULTS: From the total sample of 602 subjects, 189 (31%) were men and the remaining 413 (69%) were women. The age average was 77.21 +/- 6.18 for the men group and 78.76 +/- 5.98 for the women group. The cut-off point that obtained best efficacy was 59/60. A multiple regression analysis was made using CAMCOG total scoring as the dependent variable and clinical diagnosis, age, sex and scholarship as independent variables. All the variables intermediate on CAMCOG total scoring with a multiple correlation coefficient (R) of 0.7061 (p < 0.0000). Five of the twelve CAMCOG subareas were efficient with a significance level of p > 0.0000, classifying correctly the 82.56% of the subjects. CONCLUSION: The CAMCOG is a moderate efficient total to discriminate dementia from no dementia on epidemiological studies. PMID- 10714311 TI - [Electrophysiological characteristics of inflammatory demyelinating chronic polyneuropathy]. AB - INTRODUCTION: The electrophysiological studies, specially nerve conduction studies (NCS) constitute one of the basic supports to obtain an early and accurate diagnosis to perform a successful treatment in the chronic inflammatory demyelinating polyneuropathy (CIDP). There is nevertheless, no definite consensus about which would be the most specific and sensitive electric variables in the illness. OBJECTIVE: To describe the NCS findings in a group of patients with this diagnosis, in order to contribute to its electrophysiological characterization. PATIENTS AND METHODS: We an analyzed the NCS on 37 patients diagnosed with CIDP. The NCS were done using standard techniques. These studies were assessed according to the reference values of a normative study on 90 healthy persons. We performed somatosensory evoked potentials (SSEP), visual (VEP) and brainstem auditory evoked potentials (BAEP) in 8 patients, and motor potentials (MP) by transcranial magnetic stimulation in 3. RESULTS: The registered values are presented in media comparison tables (cases/controls). The frequency of abnormalities in latencies (L), amplitudes, conduction velocity (CV) and the presence of partial blocks is analyzed. The affectation of the evoked potentials in some patients, demonstrated subclinical concomitant demyelinization of the central nervous system. CONCLUSIONS: The most sensitive electrophysiological parameters are the motor CV, the total duration and distal latencies, that gives these variables a confident value in the initial stages of the illness. The relative normality of conduction through the sural nerve, even in the presence of severe abnormalities of the median nerve constitutes a repetitive and specific pattern to this kind of illness. PMID- 10714312 TI - [Sensitivity of motor evoked potentials in detecting cortico-spinal lesions in patients with multiple sclerosis]. AB - INTRODUCTION: Motor evoked potentials (MEP) are very useful for the evaluation of multiple sclerosis patients. OBJECTIVE: To know the sensibility of MEP in a group of patients with diagnosis of multiple sclerosis. PATIENTS AND METHODS: A transversal and descriptive study was done in 56 patients with definite multiple sclerosis (MS). MEP using magnetic stimulation were carried out in all of them, with recordings over abductor pollicis brevis and tibialis anterioris. We analyzed the sensibility of MEP and the clinical correlation of this study. In 22 patients the sensibility of MEP and somatosensory, auditory and visual evoked potentials was compared. RESULTS: Abnormalities were detected in the 87% of records, and in 18% with absences of clinical signs of corticospinal lesion. We found statistically significant differences between relapsing-remitting form and primary chronic and progressive form (Wilk's lambda = 0.606, p = 0.00), with a high lineal relation to Kurtzke Scale (p < 0.05). In relation to the other modalities of evoked potentials, MEP were the most sensible study (68.1%), followed by somatosensory (59%), visual (45.4%) and auditory (22.5%) evoked potentials. CONCLUSIONS: MEP are a very sensible modality of evoked potentials for the detection of corticospinal lesions in MS patients, with a high degree of clinical correlation. PMID- 10714313 TI - [Cortical spectral distribution of infantile electroencephalography in attention disorders]. AB - INTRODUCTION: Sensory and attention disorders cause selective effects on the topographical distribution of electroencephalograph frequencies. OBJECTIVES: In this paper we show changes in spectral topography produced by visual attention tests in a group of healthy children, thus demonstrating the cerebral areas involved in this mental process and the anatomical and bioelectrical organization of the subsystems involved. METHOD: To distinguish the effect due to cognoscitive activity from what is merely a visual sensorial process, firstly we compared resting quantified electroencephalograms with the eyes shut, with those done with the eyes open. Immediately afterwards we evaluated the spectral transformation caused by the test to find the contribution made to the electroencephalogram pattern by intellectual activity, using spectral amplitude analysis as an objective method for the measurement of recordings in a group of 10 children with an average age of 8.85 years. RESULTS AND CONCLUSIONS: The contribution of the cognitive test is seen in an increased beta frequency in the temporal and occipital cortical areas, whilst the maps of cortical distribution show the cerebral areas activated. PMID- 10714314 TI - [The use of brief questionnaire in the diagnosis of attention deficit. Study group of the Manizales University Foundation]. AB - INTRODUCTION: American Psychiatric Association has defined the DSM-IV ADD diagnostic criteria and symptoms, however, there is not a quantitative instrument to evaluate them in Spanish speaker population. OBJECTIVE: To evaluate the utility of a ADD checklist in a Colombian schooling population. PATIENTS AND METHODS: A randomized and stratified by sex, age and socioeconomic level, 4 to 17 year old, sample of 540 schooling subjects was selected from Manizales City, Colombia. An ADD checklist was applied to the parents of these subjects. RESULTS: The reliability of the different dimensions of the questionnaire (18 total items, 9 items for inattention, 9 for hyperactivity-impulsivity, and 6 for hyperactivity) were strong in both sex and in all age groups (Cronbach's alpha coefficient 0.71-0.92). Only the impulsivity dimension formed by three variables showed fairly weak reliability (0.42-0.79 Cronbach's alpha). Some factorial analysis found two dimensions. In the male sample first dimension (inattention) explain around the 45% of the variance, and the second dimension (hyperactivity impulsivity) explain around the 12 to 15% of the variance in the different age groups. In the female sample the first dimension was hyperactivity-impulsivity and the second dimension was inattention. A categorical (yes or not) scored questionnaire found a ADD estimated prevalence of 16.1, distributed in type I (combined) 3.3%, in type II (inattentive) 4.3%, and type III (hyperactive impulsive) 8.5%. Male prevalence was 19.8% and female 12.4%. CONCLUSIONS: ADD checklist Spanish version showed a strong reliability. A bidimensional stable structured was found. A clinical related ADD prevalence was presented, it was much higher than the prevalence of the developed countries. PMID- 10714315 TI - [A descriptive study of migraine in a rural population of Area del Comtat]. AB - INTRODUCTION: Headache is one of the commonest causes of consultation in neurology. There are many studies of the prevalence of migraine showing considerable variation in the results obtained. OBJECTIVES: To find the prevalence of migraine, with and without an aura, by means of a randomized transverse study carried out in a previously selected rural population, using the validated questionnaire 'Alcoi-92'. To find the overall prevalence of migraine, specifically regarding age and sex and adjusted for the European population. PATIENTS AND METHODS: A door-to-door study was made by random selection of 790 persons aged over 18, in three towns in the comarca del Comtat, Alacant, Spain. A self-questionnaire was sent to all persons studies and subsequently they were seen by a medical interviewer. RESULTS: We interviewed 548 persons (overall response rate 78%). The average age was 52.5 +/- 19.3 years. Sex distribution of the population interviewed showed a predominance of females (52.9%) as compared to males (47.1%). According to the type of headache: other types of headache 62.32%, persons with no previous history of headache 20.07%, amplified migraine 16.6%, typical migraine 12.6%, cluster type migraine 0.18%. CONCLUSIONS: The overall prevalence of amplified migraine was 16.6%. According to diagnostic groups the frequency of migraine with aura was 2.9%, migraine without aura 9.7% and typical migraine 12.6%. A predominance of women was seen in all types of migraine. The frequency of migraine, adjusted for the European population was 19.6%. Prevalence during the past year, as a measure of activity of the disorder was 15.7%. PMID- 10714316 TI - [Some hemorheological factors in patients with cerebrovascular ischemia]. AB - INTRODUCTION AND OBJECTIVE: To study hemorheological factors in patients with ischemic cerebrovascular disease (not secondary to embolic cardiopathy). PATIENTS AND METHODS: We assessed 40 patients with an average age of 64.5 years; 26 had cerebral infarcts due to alterations in major blood vessels and 14 had lacunar infarcts. Forty persons with no cerebrovascular disease acted as controls. The hematological studies were done between three weeks and six months after the initial ictus. RESULTS: Blood viscosity and plasma fibrinogen concentrations were significantly higher in the patients than in the controls. However, hematocrit values were similar in all three groups studied. With regard to the hematocrit, fibrinogen levels and blood viscosity, no differences were seen between the group with damage to the great vessels and those with lacunar infarcts. PMID- 10714318 TI - [Recurrent spontaneous carotid dissection]. AB - INTRODUCTION: Arterial dissection is the cause of 20% of the stroke occurring in adults under the age of 45. The existence of recurrence has been discussed in recent studies, and the overall frequency estimated as 4% to 8%, with a risk of 1% per year. The course of the condition is usually oligosymptomatic, so that a high index of suspicion is necessary for diagnosis to be made. We consider that different connective tissue disorders and anomalies of the vascular wall predispose to dissection. It would seen reasonable to think that these same anomalies may lead to recurrence. However, this cannot always be demonstrated. A family history of dissection is also an important factor in recurrence. CLINICAL CASES: We present two cases of recurrent spontaneous dissection of the carotid artery from a series of 22 patients with dissection, during the period 1990-1997. In the first case, the second dissection occurred 15 days after the first and in the second case, seven months later. In both cases the recurrence was in the contra-lateral carotid artery. In the second case the vascular tree was noted to have been formed of ecstatic, tortuous vessels. CONCLUSIONS: Our series shows results similar to others published. In one of these, an underlying arteriopathy which predisposed to the condition was shown. Both followed satisfactory courses. In case 2 a high index of clinical suspicion was necessary, since the recurrence presented as headache alone. PMID- 10714317 TI - [Clinical response of gabapentin for glossopharyngeal neuralgia]. AB - INTRODUCTION: The analgesic effect of the antiepileptic drug gabapentin makes it useful as an alternative for neuropathic pain. Its structural resemblance to other GABAergic antiepileptic drugs does not explain its mechanism of action, which seems not to depend on the activation of GABA receptor. Glossopharyngeal neuralgia is a rare entity which presents paroxystic crisis of pain, often with unknown etiology and poor response to treatment. CLINICAL CASES: Nine patients bearing of IX root neuralgia resistant to other therapies and ages ranged from 43 to 71 years old are being treated with gabapentin at doses between 800 and 3,600 mg daily in a period of time between two and sixteen months. Four cases in which magnetic resonance detected a compression of the nerve by posterior inferior cerebellar artery were submitted to a decompression surgery, but it was not effective. Gabapentin, alone or in association to carbamazepine, reduced the frequency and severity of crisis in seven patients, but in those with vascular compression the response was poorer. CONCLUSION: Gabapentin can be considered as an useful option for the management of glossopharyngeal neuralgic crisis, associated or not to other agents, for a short and long time, because of its good tolerance and lack of interactions. PMID- 10714319 TI - [Survival with no sequelae after near-drowning with very poor signs for prognosis including persistent bilateral non-reactive mydriasis]. AB - INTRODUCTION: The outcome of cases of near-drowning and initially poor prognostic signs are usually discouraging because of the severity of the consequent encephalopathy in most survivors. However, good recovery has been described, in spite of bad prognostic factors initially. It is difficult to establish the predictors of poor outcome which would enable one to decide when to establish and maintain advanced cardio-pulmonary resuscitation measures (CPR), since each case of near-drowning is different. CLINICAL CASE: A four year old boy survived near drowning in cold water without sequelas but with initial signs of very poor prognosis, including prolonged immersion time, coma, severe metabolic acidosis, hyperglycemia and persistent bilateral arreactive mydriasis. DISCUSSION: The beneficial effect of hypothermia is well known, and explains (at least partially) survival in cases of apparently irreversible near-drowning. Potential benefits are reduced metabolic demand which prevents the adverse effects of hypoxia and the 'diving reflex' which short-circuits the blood supply to vital organs such as the brain and heart. We consider that the persistently arreactive pupils were not due to hypoxia, but rather to bilateral uncal compression of the third cranial nerves due to cerebral edema secondary to initial hypoxia and water intoxication. CONCLUSION: This observation is yet another argument for the establishment and maintenance of aggressive manoeuvers of CPR and treatment in all children who have nearly-drowned, independently of the apparent seriousness or irreversibility. PMID- 10714320 TI - [Disseminated encephalic cryptococcosis as a form of presentation of idiopathic T CD4 lymphocytopenia]. AB - INTRODUCTION: The term idiopathic T-CD4 lymphocytopenia is used to describe a new syndrome, defined as reduced T-CD4 lymphocytes in persons with no evidence of HIV infection or other causes which would explain the immunosuppression (secondary to neoplasties, immunosuppressive treatment, hereditary immunodeficiencies, infections, etc.). The reduced number of T-CD4 lymphocytes leads to deterioration in cellular immunity and therefore this leads to a predisposition to develop tumors and opportunist infections in patients with such defects. CLINICAL CASE: We describe a case of depletion of T-CD4 lymphocytes, associated with disseminated encephalic cryptococcosis (multiple cortical, capsulo-ganglionar, thalamic and cerebellar cryptococcomas) in a patient with no evidence of HIV infection. The case we present fulfilled diagnostic criteria for idiopathic T-CD4 lymphocytopenia, a clinical condition seldom described in this country. We discuss the pathogenic mechanisms of cryptococcosis, the different varieties of Cryptococcus neoformans and their different roles as the cause of opportunist infections in humans. CONCLUSIONS: In view of the neurotrophism of this fungus, the neurological signs and symptoms should make one suspect the presence of Cryptococcus neoformans infection in non-HIV carriers with cellular immunity defects such as those present in idiopathic T-CD4 lymphocytopenia. PMID- 10714321 TI - [Chronic inflammatory demyelinating polyneuropathy associated with prostatic adenocarcinoma]. AB - INTRODUCTION AND CLINICAL CASE: Chronic inflammatory demyelinating polyneuropathy (CIDP) is an acquired autoimmune disorder of unknown cause. CIDP most often occurs alone and not as a complication of other disorders but may accompany plasma cell dyscrasias, human immunodeficiency virus infection, systemic lupus erythematosus and others diseases. The association of CIDP and carcinoma has rarely been reported and its relevance is debated. CONCLUSION: We report one patient with CIDP and prostatic adenocarcinoma. Steroids therapy were effective. PMID- 10714322 TI - [Preoperative percutaneous++ vertebroplasty in hemangioma compression]. AB - INTRODUCTION: Vertebroplasty is a new procedure used in interventional neuroradiology, involving percutaneous introduction of acrylic cement. CLINICAL CASE: We present the case of a D10 vertebral hemangioma, which had progressed so as to cause compression of the spinal cord. We used combined treatment. Vertebroplasty was done with acrylic cement, using transpedicular percutaneous puncture, with subsequent bilateral laminectomy to decompress the spinal cord. One year later the clinical condition is completely satisfactory. The signs of paraparesia and dorsalgia have disappeared. Posterior fixation was not necessary. CONCLUSIONS: Vertebroplasty is effective since the vertebral body is consolidated and pain avoided. We give details of the methodology, indications and possible complications of the technique of percutaneous vertebroplasty. PMID- 10714323 TI - [Cerebellar syndrome and myoclonus in a patient with adenocarcinoma of the colon]. AB - INTRODUCTION: Neurological paraneoplastic syndromes (NPS) are usually found in association with bronchogenic and gynecological tumors. Any part of the central or peripheral nervous system may be involved, and the clinical presentation may therefore take any of a large number of forms. Intestinal tumors rarely lead to NPS. We present the case of a patient with adenocarcinoma of the colon, in whom the first clinical sign was NPS. CLINICAL CASE: A 72 year old man presented with subacute onset of generalized myoclonus, predominantly of action, ataxia on walking and changes in speech associated with a constitutional syndrome. There were no ocular changes. Laboratory investigations including immunology, serology and tumour markers were normal. Anti-Hu, Yo and Ri antibodies were negative. Study of the CSF showed the blood-brain barrier to be damaged. Cranial MR, EEG, thoraco-abdominal CT and osseous gammagraphy showed no significant changes. On colonoscopy there was a tumour in the medial zone of the transverse colon (an adenocarcinoma). Treatment was started with high dose steroids and the condition progressively improved. After right hemicolectomy steroid treatment was stopped, and there was complete recovery of the condition. CONCLUSIONS: The presence of cerebellar syndromes and myoclonus of unknown aetiology should lead one to the diagnosis of a paraneoplastic syndrome. Adenocarcinoma of the colon should be included in the differential diagnosis. PMID- 10714324 TI - [Triple association of mesencephalic syndromes]. AB - INTRODUCTION: The mesencephalic alternating syndromes or syndromes of Weber, Benedikt and Claude are uncommon in clinical practice. They are caused by a lesion in the mesencephalus which affects the third cranial nerve bundle, together with the corticospinal pathway, subthalamic nucleus and the dentato rubric path. CLINICAL CASE: We present the case of a normotensive patient who, as a consequence of a hematoma in the mesencephalic tegmentum, had the association of these three syndromes consecutively. The clinical course was favorable, so that after three months of follow-up only paralysis of the third cranial nerve bundle persisted. DISCUSSION: In the syndromes of Weber, Benedikt and Claude there is the association of ophthalmoplegia with hemiplegia, or a cerebellar hemisyndrome or contralateral abnormal movements compatible with hemiballismus, respectively. Amongst the commonest causes are expansive processes, tumors and arteriovenous malformations. More rarely they are due to cerebrovascular accidents, which are usually ischemic and occasionally hemorrhagic in aetiology. CONCLUSIONS: Spontaneous mesencephalic hematomas are infrequent. They make up approximately 1% of all intracranial hematomas. The commonest site is the tegmentum followed by the peduncle and tectum. They have better prognosis than other hematomas of the brainstem. PMID- 10714325 TI - [Multiple cranial neuropathy: an atypical variant of the Guillain-Barre syndrome?]. AB - INTRODUCTION: Multiple cranial neuropathy or polineuritis cranealis is rarely seen in everyday clinical practice. It is considered to be a topographically circumscribed form of the Guillain-Barre syndrome. The cases described have a wide range of clinical and biological characteristics. Some of these may be due to infectious agents. CLINICAL CASE: We present the case of a 50 year old man with acute onset of diplopia, dysphagia, anarthria, cervical and right arm flexor extensor muscle weakness due to involvement of many motor cranial nerves and superior cervical nerve roots. On neurological examination there was mixed involvement, mainly of the axons of the nerve trunks involved. Studies to determine aetiology did not show any demonstrable agent. DISCUSSION AND CONCLUSIONS: Different topographical varieties of the Guillain-Barre syndrome have been described, including: Fisher's syndrome, pharyngo-cervico-brachial paralysis, arreflexive paraparesia, bilateral facial paralysis with paraesthesias, hyporeflexia and bilateral lumbar polyradiculopathy. We compare the clinical characteristics of our patient with those described in the literature. We found a degree of heterogenicity in the clinical and biological characteristics of the cases described, which may mean that they had different aetiologies. Therefore, we consider that before labelling these conditions as atypical variants of the Guillain-Barre syndrome, a thorough search should be made for a precise aetiology. PMID- 10714326 TI - [Myoclonic epilepsies in pediatrics]. AB - INTRODUCTION: In our environment the frequency of epilepsy is 1.9% and in the central anti-epilepsy league (LICCE) of Santafe de Bogota, Colombia, some 1,500 patients are attended each month (approximately 75 daily), of which an average of 7.2 consulted for the first time. DEVELOPMENT: This article about myoclonic epilepsy in paediatrics allows the clinician to have a general view of the patient who consults for the first time, facilitating his subsequent treatment. We emphasize the etiological classification of the first crises, seeking specifically to discover whether it is a disorder, a sequela, a syndrome or an isolated crisis. We present the basic definitions: classification according to physiology, anatomy, symptomatology (epileptic or nonepileptic), cryptogenetics (intermediate or polymorphic) and benign, severe or progressive. CONCLUSION: Finally we draw attention to the description of fifteen syndromes in which the clinical picture of progressive, myoclonic epilepsy is variable and the aetiology complicated. PMID- 10714327 TI - [Cerebral mapping during sleep: a critical review of literature]. AB - INTRODUCTION: Cerebral mapping is a useful tool which permits graphic representation of certain EEG characteristics which are imperceptible on simple visual inspection. This technique offers quite a good solution to analysis of cerebral rhythmicity, by including spectral, spatial and statistical parameters. Sleep is a physiological process during which dynamic interchange of cerebral rhythms has an important part in delimiting the different phases, not to mention the decisive part played by phasic events at each stage of it. DEVELOPMENT: In this paper we review the studies which have used cerebral mapping techniques during the different stages of sleep. In order to give structure to the data obtained from these papers, we have classified the information into aspects related to the methodology used in these studies and the results obtained. Decisions regarding methodology are discussed, and further information given regarding technical aspects which might have had decisive influence on the results obtained using this technique. CONCLUSIONS: We give suggestions as to the possibilities of the use of cerebral mapping in studies on sleep, both clinically and experimentally. PMID- 10714328 TI - [Neuropsychological sequelae of head injury]. AB - INTRODUCTION AND DEVELOPMENT: Traumatic brain injury (TBI) neuropsychological sequelae are consequence of the combination of focal and diffuse cerebral lesions. Foci of concussion usually involves frontal lobes. Fronto-basal lesions produces important changes in mood, personality and behavior, and dorsolateral lesions impairment of executive functions (lack of planning, flexibility and use of strategies). Left temporal concussion can produce aphasia. Hippocampal and parahippocampal atrophy are essentially responsible of memory dysfunctions. Diffuse axonal damage is related to impairment of attention, speed of mental processing and frontal lobe functions. The development of neuroimaging techniques, especially tridimensional magnetic resonance acquisition and analysis, allows accurate anatomo-functional correlates. Genetic variables can explain in part individual differences in the degree of memory impairment and the relationship of TBI with Alzheimer's disease. CONCLUSIONS: The TBI percussion model performed in rat and mouse allows the study of the interrelationship among structural damage, memory changes, genetic factors, and the effects of pharmacological treatment. PMID- 10714329 TI - [Neurology in the medical papyruses of the pharaohs]. AB - The civilization of Ancient Egypt included a long period of almost three millenniums, and is the most interesting example of the so-called pretechnical archaic cultures. Papyrus scrolls are the main source of information about medical activities. There are fourteen medical papyrus scrolls, in varying states of conservation, mostly corresponding to the Middle Empire, but containing references to the Ancient Empire (the period of the pyramids). These are practical treaties with little explanation of the underlying pathology (a primitive theory of the 'flow' of humors, involving the flowing of different malignant entities) within a system of magic and religion. The empirical observations referring to diseases or dysfunctions of the nervous system, although few, seem to be worth reviewing. Remedies for migraine ('the disorder affecting half the head') take up a long chapter of the only complete and best preserved Ebers papyrus. Dementia (deterioration with age), convulsions and tetany are briefly mentioned in several papyrus scrolls. With the detailed description of the clinical findings of cranial and vertebral trauma, and the orderly assessment of severity presented in Edwin Smith's papyrus the neurology of pharaonic Egypt attained its greatest importance. PMID- 10714330 TI - [The parkinsonian view of the Rey-Osterrieth complex figure]. AB - Visuospatial impairment has been frequently reported in Parkinson's disease (PD). We present the progressive distortion performance of the Rey-Osterrieth complex figure in parkinsonian patients at different stages of the disease (PD de novo, PD on long-term treatment, PD with phychosis and PD with dementia). PMID- 10714332 TI - [The ear of the owl: a system of increased temporal dysphase in between depolarization fronts]. PMID- 10714331 TI - [Cervical myelopathy due to a tumor of the foramen magnum]. PMID- 10714333 TI - [Chronic resistant sciatalgia]. PMID- 10714334 TI - [Pseudo-migraine with temporary neurological symptoms and cerebrospinal fluid pleocytosis]. PMID- 10714335 TI - [Optic neuritis as a clinical manifestation of neurobrucellosis]. PMID- 10714336 TI - [Neurophysiological study of thin myelinated and unmyelinated fibers]. AB - INTRODUCTION: Standard neurophysiological techniques evaluate thick myelinated fibers. Yet, peripheral nerves are equally composed of thin myelinated and unmyelinated fibers. The latter are responsible for autonomic function as well as temperature and pain perception. DEVELOPMENT: Microneurographic studies are restricted to investigation laboratories. Since the techniques are complex and invasive, their performance is still poor for clinical purposes and some of the components to be analyzed, such as cardiovagal, cannot be directly recorded. The clinical need to evaluate the functions regulated by the autonomic nervous system (ANS) had led to devising a series of tests which, in most cases, rely on reflex responses evoked by already known standardize stimuli. The battery chosen has to be non invasive, reproducible, specific, providing relevant data to the investigated function, with a readily available technology, which has to be managed being aware of the physiological and pathological factors that might bear an influence on the results. The recent development of heart rate and blood pressure power spectral analysis, provides a new interesting insight for quantification of ANS abnormalities. The study of thermography and thermometry of body surface brings forward evidence on the activity of other thin and unmyelinated fibers components of the peripheral nerve spectrum. CONCLUSION: The adequate management of the above mentioned tests gives rise to a more extensive and appropriate knowledge of the whole peripheral nerve fiber spectrum. PMID- 10714337 TI - [Small fiber dysfunction in peripheral neuropathies]. AB - INTRODUCTION: Disfunction of thin myelinated and unmyelinated fibers may appear isolated or in association with large-myelinated fibers lesion. Small-fiber neuropathy includes autonomic and sensory symptoms, most prominent of them thermo algesic deficits. DEVELOPMENT AND CONCLUSION: In some acute neuropathies, small fiber lesion is relatively pure, as in pandysautonomia, but it also appears in disorders with prominent somatic involvement, such as the Guillain-Barre syndrome, in which case autonomic symptoms worsens the prognosis. Small-fiber dysfunction is important in certain diseases that involve different components of the nervous system, like paraneoplastic syndromes and porphyria. Some drugs and toxic substances may damage thin myelinated and unmyelinated fibers. Nowadays, chronic idiopathic small-fiber neuropathy is diagnosed more frequently, because of the recent development of techniques that selectively evaluate this peripheral nerve component. Hereditary sensory and autonomic neuropathies can also be studied. Small-fiber dysfunction is very prominent in some diseases, e.g. diabetes mellitus and amyloidosis. In the pure autonomic failure, only the peripheral component of the autonomic nervous system is affected, and this feature is the key to make diagnosis versus multisystem atrophy. There are situations in which there is no clear deviation from normality, namely old age autonomic failure and orthostatic intolerance syndrome in which autonomic evaluation is mandatory. PMID- 10714338 TI - [Arousal and its influence on vigilance]. AB - INTRODUCTION: The term arousal usually refers to transient, brief arousal during nocturnal sleep or to a state of vigilance which is maintained during the day. DEVELOPMENT AND CONCLUSION: In this review we consider the various accepted meanings and clinical disorders of 'arousal'. Insomnia (hyperarousal), parasomnias (partial arousal) and the sudden death syndrome of infancy are all attributed to disorders of arousal. It is concluded that diurnal hyperarousal leads to difficulty in getting off to sleep and staying asleep, whilst nocturnal arousal leads to excessive somnolence during the day, due to broken sleep during the night. PMID- 10714339 TI - [Arousal and motor activity during sleep]. AB - OBJECTIVE: To describe different types of motor activity which occur during sleep in relation to episodes of arousal and sleep disorder. DEVELOPMENT: During sleep, normal motor activity should be distinguished from paroxystic episodes: parasomnias; abnormal movements such as nocturnal paroxystic dystonia, which is very similar to epilepsy of frontal origin; nocturnal epileptic crises and especially periodic movements of the limbs and the restless legs syndrome, which is related to it. Physiological cyclical fluctuations of sleep are common to all these conditions and due to cortico-subcortical changes in excitability. CONCLUSION: We review diagnostic, clinical and neurophysiological criteria and aspects of physiopathology and treatment. PMID- 10714340 TI - [Arousal of respiratory origin and upper airway resistance syndrome: pathophysiological and diagnostic aspects]. AB - INTRODUCTION: The description of Upper Airway Resistance Syndrome (UARS) let us to recognize the importance of the pair 'respiratory effort-arousal' on sleep disordered breathing pathophysiology. DEVELOPMENT: First part of this paper reviews knowledge about respiratory arousal pathophysiology. Arousal response is normally needed to end obstructive respiratory episodes, but it is also the cause of sleep fragmentation. Among respiratory stimuli able to provoke arousal (respiratory effort, hypoxemia and hypercapnia), respiratory effort is the most constant. Neurophysiological mechanisms involved in arousal, sleep and vegetative consequences, and the possible role of non visible arousals, are also discussed. In UARS, because of the absence of apnea/hypopnea and significative O2 desaturations, arousals are induced by the increased respiratory effort. Diagnosis needs the simultaneous recording of polysomnography and esophageal pressure. Some symptoms and signs of UARS are similar to those of Obstructive Sleep Apnea Syndrome. However, UARS shows any differences: a lower Body Mass Index, less constant snoring, males and females are similarly affected or higher frequency of craniofacial abnormalities. Diagnostic difficulties may be due to confusion between hypopneas and episodes of increased resistance of upper airway, or to the lack of definitive diagnostic criteria. Finally, differential diagnosis needs a broad knowledge of disorders of excessive daytime sleepiness. PMID- 10714341 TI - [Intraoperative electroneurophysiological monitoring of basal cranial nerve surgery]. AB - INTRODUCTION: Since 1980 when intraoperative monitoring was introduced or acoustic nerve surgery, the technique rapidly was outspreaded to the other basal cranial nerves, and today is the most important and indispensable piece of work first of all because of its effectiveness in the prevention of the neurologic deficits. OBJECTIVE: The aim of this work is to describe the basic requisites needed (personnel, equipment, stimulation and recording electrodes, patient handling), as well as to review the different techniques used (electromyography and electroneurography) in the motor cranial nerve monitorization (facial, motor trigeminal, extraocular muscles, glossopharyngeal, vagus, spinal accessory and hypoglossal) and the sensitive cranial nerves (optic, sensitive trigeminal, and acoustic) especially with evoked potentials. Finally, we pointed out the non neurologic structures monitoring during the surgery of the posterior fossa. PMID- 10714342 TI - [Protocol of intraoperative neurophysiological spinal cord monitoring]. AB - INTRODUCTION: The consequences derived from medullar harm caused during several spinal cord surgical interventions can often be catastrophic for the patient, even more taking into consideration that many of them are young. DEVELOPMENT: This problem is observed specially at interventions of spinal malformations (kyphoscoliosis), but also during other surgical techniques like fractures, degenerations, spinal tumors and aortic lesions. The use of somatosensory evoked potentials (SEP) began at late 70's, as a method to monitorize spinal cord function during surgery; years later, motor evoked potentials (MEP) joined this option, giving us direct information about the functioning of posterior spinothalamic tract (posterior column) and lateral corticospinal tract (pyramidal tract), respectively. This has motivated that, although the degree of surgical difficulty and complexity of spinal instrumentation have raised, the actual percentage of neurological complications derived from them has decreased. CONCLUSIONS: This article describes the intraoperative spinal cord monitoring protocol followed at Hospital Clinico San Carlos of Madrid, Spain, which includes the making of SEP and MEP, the latest according to the translaminar stimulation technique. PMID- 10714344 TI - [Intraoperative electrical cortical stimulation in cerebral lesions]. AB - INTRODUCTION: Intraoperative brain mapping is used during neurosurgery in functional cortex to facilitate the extent of cortical resection and to identify nonfunctional tissue to remove subcortical lesions. OBJECTIVE: To describe the intraoperative brain mapping protocol that has been utilized in the Hospital Clinico San Carlos of Madrid, Spain. Detailed somatotopic motor, sensitive and language localization is possible using direct cortical electric stimulation in the awake patient. CONCLUSIONS: Intraoperative brain mapping is a surgical adjunct used during lesionectomy in functional cortex. In addition to preserving functions, the other essential goal is to achieve a radical removal while attempting to minimize the associated morbidity. PMID- 10714343 TI - [The use of somatosensory evoked potentials for cortical localization during surgical operations]. AB - INTRODUCTION: Identification of the cortical areas of Roland, by means of anatomical references seen on craniotomy, with a view to their conservation during surgical operations, is found to be associated with a high rate of error which may reach 50% with experienced surgeons. MATERIAL AND METHODS: Somatosensory evoked potentials (PESS) following stimulation of the median nerve may easily be recorded using electrodes situated directly over the cerebral cortex of patients under general anaesthesia. To obtain this recording, a bipolar set-up of a series of between 6 and 10 electrodes in line one centimetre apart allows identification, by means of phase opposition, of the cortical zones, usually the parietal zone adjacent to the sulcus of Roland, where these potentials are generated. Identification of motor sites (area 4) can be obtained by electrical stimulation of the cerebral cortex, although we have rejected this type of exploration because of its technical difficulty and possible undesirable effects. CONCLUSION: We consider that, when at surgical operation the cortical area corresponding to the median nerve is identified, and care taken not to damage the cortex 2 cm rostrally, excellent results may be obtained, reducing the sequelae which may occur with this type of surgery. PMID- 10714345 TI - [Protocol of pre-surgical neurophysiological evaluation in epilepsy resistant to treatment]. AB - INTRODUCTION: Nowadays approximately twenty per cent of epileptic patients who are on a pharmacological treatment carry on their fits. Their instability can be eliminated with an operation in any center that is specialized in epileptic surgery. The center must have a cross disciplinary team. DEVELOPMENT: The ideal operation is one that only eliminates the tissue needed to end up with the fits The main goal of the neurophysiological evaluation is to delimit the cerebral area that generates an epileptogenic activity so as to be able to eliminate it without causing further damage. We present a medical record of pre-surgical neurophysiological evaluation that we would like to put into practice in our department in coordination with the services of neurology, radiodiagnosis, and neurosurgery. This medical record contains the following sections: 1. Selection of patients. 2. Non invasive pre-surgical evaluation. 3. Invasive pre-surgical evaluation. 4. Postsurgical evaluation. The invasive pre-surgical evaluation constitutes the most interesting part of the whole process, it allows thanks to intracranial registers to locate accurately epileptogenic focuses; what is more, it allows to carry out a functional mapping of areas that cannot be explored with surface techniques; and finally this evaluation allows to open new fields of investigation about the way in which the encephalon works. CONCLUSION: Epilepsy surgery is a subject to develop in Spain in which the neurophysiologic exploration is something essential. PMID- 10714346 TI - [Stereotactic target identification for neurosurgery of Parkinson disease]. AB - INTRODUCTION: The use of applied neurophysiological methods to improve the stereotactic localization of devices in the deep human brain is a high and systematic technology in Parkinson's neurosurgery today. The available standard equipment for clinical neurophysiology practice may constitute the basic set for high tech functional neurosurgery. Free run and event related multiunit recording, naturalistic and electrical evoked potentials, and deep brain microstimulation responses are the basic methodological set to neurophysiological target localization. DEVELOPMENT AND CONCLUSIONS: This article is concerned with the topic: set out a high technology using low cost equipment. So our 41 cases experienced in pallidal and thalamic nucleolisis and thalamus and subthalamus DBS results suggest that the proposed equipment and methods are the required to assure accuracy and safety for target location. PMID- 10714347 TI - [P300 and neuropsychological tests in schizophrenia and bipolar patients]. AB - INTRODUCTION: A smaller P300 amplitude has been found in schizophrenic patients in the majority of the studies done. And this occur in spite of treatment, sex, clinical stage and evolution time of the illness. It has been already described an association between this amplitude reduction and a poor result on the neuropsychological tests. But these changes in the P300 amplitude are not specific of the schizophrenic patients and they can be found in affective patients. OBJECTIVES: This study try to value the influence of the clinical stage and the cognitive results on the P300 values in the bipolar patients and to compare these results with the schizophrenic patients. PATIENTS AND METHODS: It has been evaluated the evoked potentials and the results of a neuropsychological battery of tests in three patient groups: schizophrenics in patients with acute symptomatology (n = 18), schizophrenic out patients without acute symptomatology (n = 15) and bipolar in patients during a maniac period (n = 16). RESULTS: It has not been found any differences between schizophrenic and maniac patients nor in the amplitude or in the latency of the P300. The results of the neuropsychological tests have not any relation with P300 amplitude and latency except for the verbal fluidity (VF) which has a good correlation with a longer P300 latency. At the same time this was the only test in which schizophrenic patients have smaller results than affective patients with a good correlation. CONCLUSIONS: These results suggest a similar pattern of neuropsychological and neurophysiological damage in schizophrenic and affective patients in acute period of mania. PMID- 10714348 TI - [The neurophysiological diagnosis of myopathies]. AB - INTRODUCTION: Electromyography is a technique widely used to assist diagnosis of neuromuscular disorders. The neuromuscular system as a whole (neuron, nerve, neuromuscular junction and muscle) have several electrochemical properties which may be detected by suitable apparatus and are the basis of electromyography. DEVELOPMENT AND CONCLUSIONS: Healthy muscle has particular characteristics which are affected by structural/functional changes, whether they are of primary origin (myopathies) or secondary to alterations at other levels (secondary muscle involvement). In this paper we wish to summarize the electromyographic findings typical of myopathies, those which differentiate these from other neuromuscular disorders and to establish when possible the characteristic signs of each form of presentation. Finally we give a brief bibliography which we consider to be basic to understanding this diagnostic technique in this type of illness. PMID- 10714349 TI - [Automatic methods of measurement of motor potential units]. AB - INTRODUCTION: The study of motor unit potentials (MUP) may be carried out using qualitative or quantitative methods. The considerable usefulness of quantitative methods in the diagnosis of neuromuscular disorders has led to development of new automatic techniques which permit greater speed and accuracy of clinical investigation. In this review we consider traditional electromyographic techniques and describe some automatic methods. DEVELOPMENT: Qualitative techniques are simpler but give limited information since they only permit recording of very marked alterations. The introduction of trigger and averaging permit quantitative analysis and thus the evaluation of less obvious changes. Although they do not identify all the potentials of an electromyographic signal, the new automatic techniques using partial decomposition avoid the loss of time occurring with earlier techniques, give reliable objective measurements of the MUP and also the possibility of measuring new parameters, such as the area or frequency of discharge. CONCLUSIONS: New methods of evaluation of the MUP have significant advantages over traditional methods, namely increased speed and objectivity. However, this makes it necessary to redefine normal values since different methods are used for obtaining and processing the signals, otherwise the results may not be strictly comparable. PMID- 10714350 TI - [Automatic analysis of interference pattern: principles and applications]. AB - INTRODUCTION: The signals obtained during conventional needle examination are usually assessed subjectively by the neurophysiologist. Buchthal's approach allows quantification of several motor unit parameters during mild effort. DEVELOPMENT: The automatic methods of analysis of the EMG interference pattern (IP), in particular Turns/Amplitude, give us information concerning the number, firing rate, and morphology of the motor units, as well as the recruitment pattern at different degrees of effort. In pathological conditions, the changes in the IP allow distinction between myopathies and neuropathies. CONCLUSION: These methods, which complement other methods of motor unit analysis, have also proved to be excellent for follow up studies. PMID- 10714351 TI - [Macro-electromyography findings in neuropathies and myopathies]. AB - The possibility of overall recording of the electrical activity generated by muscle fibres making up the motor unit in its entire territory, is the characteristic and main objective of macro-EMG studies. This is possible because of the technical characteristics of the electrode and the recording conditions whose methodology is described. We describe how parameters of macro-EMG are obtained in normal healthy people, and define changes in parameters in pathological neurogenic and myogenic disorders and their value in the follow-up of reinnervation processes. PMID- 10714352 TI - [Spinal subdural hematomas: clinical and magnetic resonance findings]. PMID- 10714353 TI - [Epilepsy of late onset]. PMID- 10714354 TI - [Type II schizencephaly: magnetic resonance imaging]. PMID- 10714355 TI - Structure-activity relationships of G protein-coupled receptors. AB - The primary function of cell-surface receptors is to discriminate the specific signaling molecule or ligand from a large array of chemically diverse extracellular substances and to activate an effector signaling cascade that triggers an intracellular response and eventually a biological effect. G protein coupled cell-surface receptors (GPCRs) mediate their intracellular actions through the activation of guanine nucleotide-binding signal-transducing proteins (G proteins), which form a diverse family of regulatory GTPases that, in the GTP bound state, bind and activate downstream membrane-localized effectors. Hundreds of GPCRs signal through one or more of these G proteins in response to a large variety of stimuli including photons, neurotransmitters, and hormones of variable molecular structure. The mechanisms by which these ligands provoke activation of the receptor/G-protein system are highly complex and multifactorial. Knowledge and mapping of the structural determinants and requirements for optimal GPCR function are of paramount importance, not only for a better and more detailed understanding of the molecular basis of ligand action and receptor function in normal and abnormal conditions, but also for a rational design of early diagnostic and therapeutic tools that may allow exogenous regulation of receptor and G protein function in disease processes. PMID- 10714356 TI - The many roles of the calcium-sensing receptor in health and disease. AB - The physiological relevance of calcium in many vital processes requires that its concentration in extracellular fluids be kept within a narrow range. The near constancy of this parameter emphasizes the remarkable sensitivity of cells sensing changes in extracellular calcium concentration to minimal fluctuations (< 2%) and the level of sophistication of the homeostatic system (1). The identification of a cell surface, Ca2+ (polyvalent cation)-sensing receptor (CaR), has shed considerable light on the molecular aspects of hypercalcemia on cell function (2). Activation of the receptor by calcium triggers an intracellular cascade of second messengers producing a variety of biological effects, many of which have yet to be understood. This suggests, for the first time, that Ca2+ can exert its effects in a hormone-like fashion without crossing the plasma membrane. The demonstration that inherited genetic disorders of Ca2+ homeostasis are associated with mutations that reduce or enhance responsiveness of the receptor to extracellular Ca2+ concentration clearly proposes CaR as the main regulator of divalent mineral ion excretion (3). This hypothesis is confirmed by the assessment of the presence of the receptor in all regions involved in Ca2+ homeostasis (e.g., parathyroid glands, kidney, calcitonin secreting C cells, bone-derived cell lines, and intestine) (1,4-8). Recently, the receptor has also been found in regions not normally involved in mineral ion metabolism, such as the brain, eye, stomach, and pancreas (9-13). This clearly indicates a much broader relevance of CaR in the maintenance of local ionic homeostasis and, possibly, in the involvement in vital processes such as the regulation of cell fate. PMID- 10714357 TI - Alpha 1-adrenoceptors: subtypes, signaling, and roles in health and disease. AB - Alpha 1-adrenoceptors mediate some of the main actions of the natural catecholamines, adrenaline, and noradrenaline. They participate in many essential physiological processes, such as sympathetic neurotransmission, modulation of hepatic metabolism, control of vascular tone, cardiac contraction, and the regulation of smooth muscle activity in the genitourinary system. It is now clear that alpha 1-adrenoceptors mediate, in addition to immediate effects, longer term actions of catecholamines such as cell growth and proliferation. In fact, adrenoceptor genes can be considered as protooncogenes. Over the past years, considerable progress has been achieved in the molecular characterization of different alpha 1-adrenoceptor subtypes. Three main subtypes have been characterized pharmacologically and in molecular terms. Splice variants, truncated isoforms, and polymorphisms have also been detected. Similarly, it is now clear that these receptors are coupled to several classes of G proteins that, therefore, are capable of modulating different signaling pathways. In the present article, some of these aspects are reviewed, together with the distribution of the subtypes in different tissues and some of the known roles of these receptors in health and disease. PMID- 10714358 TI - Insulin resistance and type 2 diabetes mellitus: its relationship with the beta 3 adrenergic receptor. AB - The beta 3 subtype of adrenaline and noradrenaline receptors has been extensively characterized at structural and functional levels. Ligand binding and adenyl cyclase activation studies have helped to define their unique beta-adrenergic profile. Humans, other larger mammals, and rodents share most of the characteristic beta 3-adrenergic receptor properties, although obvious species specific differences have been identified. Most studies in animal models have shown a distinct beta 3-adrenergic receptor activity that results in an increase in energy expenditure, decrease of fat mass (especially of intra-abdominal fat), and increased glucose disposal efficiency. It is of interest that mild weight increase was shown to develop in female but not male mice, in whom the beta 3 adrenergic receptor gene was disrupted. Recently, the incidence of a naturally occurring variant of the human beta 3-adrenergic receptor was shown to correlate with hereditary obesity in Pima Indians and Japanese individuals. In Western obese patients, this phenotype increased the capacity to gain weight and develop type 2 diabetes mellitus. Studies of humans with the Trp64Arg variant have shown controversial results. Many studies have failed to show any effect in heterozygous male subjects, and only modest effects in homozygous male subjects. In women, several studies have shown modest-to-significant effects regarding weight gain, intra-abdominal fat, and decreased insulin sensitivity in heterozygous and homozygous women. Other studies have failed to show any effect in heterozygous females. Disruptions in the activity of the beta 3-adrenergic receptor in the homozygous male and the heterozygous or homozygous female appear to have a profound effect in animal models, but a limited consequence in human physiology. Association with obesity or diabetes in humans is still controversial. This difference between animal and human models may be explained by the different quantity and distribution of metabolically active brown adipose tissue in the two. PMID- 10714359 TI - Vasopressin receptor mutations and nephrogenic diabetes insipidus. AB - X-linked recessive nephrogenic diabetes insipidus is caused by mutations in the gene encoding the V2 vasopressin receptor (V2R), the mediator of the antidiuretic effect of arginine vasopressin (AVP) in mammalian kidneys. Upon binding to AVP, the receptor activates the G protein Gs, stimulating a phosphorylation cascade that promotes translocation of presynthesized water channels to the apical surface of the principal cells lining the last segments of the nephron. The presence of these channels allows the flow of water from the hypotonic lumen of the nephron into the hypertonic interstitium. More than 100 different mutations have been identified since the receptor gene was characterized--in most cases one per family, although some families bear two and three mutations in the same gene. The frequency of the de novo mutations identified suggests that the DNA at the end of the long arm of the X chromosome is very susceptible to alteration. The mutations are scattered within the coding region, not confined to a particular segment of the receptor protein, and in most cases confined to a single amino acid change that significantly reduces the number of receptors present on the plasma membrane. Some mutations do not affect protein synthesis but significantly reduce the coupling efficiency between the receptor and G protein. Analysis of the biochemical impact of the mutations has provided valuable information about the synthesis and regulation of the receptor. PMID- 10714360 TI - Isolated glucocorticoid deficiency and ACTH receptor mutations. AB - Familial isolated glucocorticoid deficiency is a form of potentially lethal hereditary unresponsiveness to ACTH that manifests as primary adrenal insufficiency, usually without mineralocorticoid deficiency. Affected children commonly present with hyperpigmentation, recurrent hypoglycemia, chronic asthenia and failure to thrive within the first 2 years of life. Typically, they have deficient production of cortisol and adrenal androgens in the presence of markedly elevated ACTH levels, while renin and aldosterone levels are usually normal and responsive to activation of the renin-angiotensin axis. Clinical awareness of these syndromes is of considerable prognostic and therapeutic importance. The etiological involvement of the ACTH receptor gene in isolated glucocorticoid deficiency has been recently established in many, but not all, affected families. Several naturally occurring mutations of the ACTH receptor gene have been identified to date and have helped illuminate the mechanisms of ligand binding and signal transduction by this receptor. Discovery of the molecular defect(s) responsible for isolated glucocorticoid deficiency in cases with a normal ACTH receptor gene coding region and for the triple A syndrome (adrenal insufficiency, alacrima, achalasia) will hopefully provide further insight into the mechanisms of adrenocortical function and will increase the prospect of new therapeutic approaches. PMID- 10714361 TI - Mutations of the GnRH receptor gene: a new cause of autosomal-recessive hypogonadotropic hypogonadism. AB - Mutations in a few genes have been identified in hypogonadotropic hypogonadism (HH): the gene KAL-1 is involved in X-linked Kallmann syndrome associated with anosmia and mutations in transcription factors, namely, DAX-1 and Prop-1 were also evidenced when associated with other pituitary or endocrine defects. Recently, compound heterozygote mutations in the GnRH receptor gene were described both in males and females and hormonal resistance was confirmed in vitro. There is a wide spectrum of phenotype, ranging from complete HH with lack of pubertal development and cryptorchidism to partial hypogonadism with an arrest of pubertal development. In complete GnRH resistance, endogenous LH secretory patterns were abnormal, either apulsatile or characterized by a low-normal pulse frequency with small pulses or erratic pulses of low amplitude. In patients with partial resistance, basal LH plasma concentration was low, but FSH level was in the normal range. LH pulse analysis revealed normal frequency with decreased amplitude. Mutations are distributed along the coding sequence, as reported for other GPCRs. However, two hot-spots, Q106R and the R262Q, were observed, regardless of the geographic origin of the patients. In most cases, patients responded to GnRH administration, making the GnRH test inappropriate for screening GnRH resistance in IHH. PMID- 10714362 TI - Follicle-stimulating hormone ligand and receptor mutations, and gonadal dysfunction. AB - In contrast to the general contention, infertility can be an inherited condition. Some of the genetic causes of male and female infertility have turned out to be due to inactivating mutations in the gonadotropin and gonadotropin receptor genes. The topic of the present text is to review current knowledge on mutations affecting the function of follicle-stimulating hormone (FSH). This gonadotropin, by binding to its specific G protein-coupled cell membrane receptor (FSHR), is important for normal gonadal function. Mutations affecting gonadotropin genes are extremely rare, but recent genetic studies have revealed that the pathogenesis of subfertility or infertility can be due to mutations in the FSH receptor (FSHR) gene. While mutations affecting FSHR are sporadic, polymorphism of the FSHR gene seems to be a common phenomenon. To date, six inactivating and only one activating mutation have been detected in the FSHR gene. In contrast to LHR gene, the majority of these mutations affect the extracellular domain of the receptor. Together with animal models using the transgenic and knock-out approaches, systematic analysis of alterations in the FSHR gene increases our knowledge on the structure and function of the FSHR and demonstrates that the integrity of each FSHR segment is required for proper expression of the fully active protein and for normal gonadal function. Mutations in the FSHR gene have different consequences in the reproductive function depending on the sex of the patient: while normal ovarian function is critically dependent on FSH, male fertility is possible with minimal or absent FSH action. PMID- 10714363 TI - Male pseudohermaphroditism due to inactivating luteinizing hormone receptor mutations. AB - Human male sexual development is regulated by chorionic gonadotropin (CG) and luteinizing hormone (LH), the action of both mediated by the LH receptor (LHR). Mutations that inactivate the LHR cause Leydig cell hypoplasia (LCH), an autosomal recessive disorder. In its mild form, LCH patients present with male hypogonadism. In its severe form, patients present with male pseudohermaphroditism, with female external genitalia, and cryptorchid testis. Mullerian derivatives are absent. Histological examination of the testis shows absence of mature Leydig cells. LCH patients have elevated plasma levels of LH, normal-to-elevated levels of follicle stimulating hormone (FSH), and low levels of testosterone that do not respond to CG stimulation. Missense mutations, nonsense mutations, deletion mutations, and in-frame insertion mutation of the LHR have been identified in patients with LCH. These mutations are not localized in any particular region of the gene and cause variable degrees of receptor activity loss. The clinical manifestation of patients with LCH with homozygous or compound heterozygous mutations can be correlated with the residual activity of their respective mutated LHRs. Homozygous inactivating mutations of the LHR in the female cause hypergonadotrophic hypogonadism with primary amenorrhea or oligoamenorrhea, cystic ovaries, and infertility. PMID- 10714364 TI - Constitutively active mutations of G protein-coupled receptors: the case of the human luteinizing hormone and follicle-stimulating hormone receptors. AB - Activating mutations of the luteinizing hormone receptor (LHR) and the follicle stimulating hormone receptor (FSHR) have been known for several years. These activating mutations permanently stimulate, in the absence of their cognate ligand, the receptor signaling pathways. In the case of the LHR, the induced chronic stimulation causes sporadic and familial pseudoprecocious puberty, a phenotype observed only in males. The absence of a female phenotype is probably due to the requirement for FSH in the induction of LHR expression. For the FSHR, one activating mutation was found in a patient with normal spermatogenesis without detectable gonadotropins. Whether activating mutations of the gonadotropin receptors are involved in tumor development is not yet clear. Activating mutations of the FSHR were supposedly involved but not found in ovarian tumors. For the LHR, only one patient with a seminoma and an activating mutation was described. The different occurrence of activating mutations of the LHR compared to the FSHR is surprising, since the two genes are adjacently located on chromosome 2 and should therefore be affected by a similar mutation rate. It might well be that mutations occur with the same frequency, but that activating mutations of the FSHR do not result in any particular phenotype. PMID- 10714365 TI - Hyperfunctioning thyroid adenoma and activating mutations in the TSH receptor gene. AB - Thyrotropin (TSH) positively controls the function, differentiation, and growth of thyrocytes. TSH interacts with thyrocytes through the TSH receptor and its action is mediated by cyclic AMP-dependent mechanisms. From data gathered on adrenergic receptors, it was hypothesized that TSH receptor mutations that lead to constitutive activation of the TSH receptor would also result in autonomous thyroid growth and function. Indeed, such mutations were shown to be the main molecular mechanisms leading to toxic thyroid adenomas. The same mechanism was shown to be operating in "hot" thyroid nodules from multinodular goiter. A low iodine supply seems to increase the clinical expression of such somatic mutations responsible for thyroid autonomy. Moreover, the presence of such mutations has helped to define a working model for TSH receptor physiology. The unliganded TSH receptor maintains a negative constraint on the signal transduced, whereas the presence of specific mutations activates the receptor. PMID- 10714367 TI - Clinical implications of genetic defects in G proteins: oncogenic mutations in G alpha s as the molecular basis for the McCune-Albright syndrome. AB - Signal-transducing guanine nucleotide-binding proteins (G proteins) couple extracellular receptor proteins to intracellular effector enzymes and ion channels, and therefore are critical mediators of cellular responses to external stimuli. G proteins are comprised of three subunits (alpha, beta, gamma), each encoded by many different genes. The multiplicity of G protein subunits facilitates great combinatorial variability, which, in part, accounts for the ability of G proteins to interact with many different receptor and effector proteins. Hundreds of G protein-coupled receptors have been identified, and their unique patterns of expression among a restricted number of cell types contributes greatly to the apparent specificity of hormone action. Mutations that either activate or inactivate some of these receptors account for a number of highly specific syndromes, which affect a limited number of target tissues. By contrast, most G proteins are widely expressed in many tissues. Accordingly, mutations in these signaling molecules would be expected to produce a more generalized pattern of hormone dysfunction. Activating mutations in the gene (GNAS1) that encode the alpha subunit of the G protein that stimulates adenylyl cyclase (AC) have been identified in many endocrine neoplasms and diverse tissues of patients with McCune-Albright syndrome. The McCune-Albright syndrome is characterized by autonomous endocrine function, hyperpigmented skin lesions, and fibrous dysplasia of bone--effects which reflect the ability of CAMP to stimulate cell function and proliferation in a wide variety of tissues. The unusual features of the McCune Albright syndrome are explained by the mosaic distribution of cells bearing the mutant allele, an observation that is most consistent with postzygotic mutation of GNAS1. Experimental analysis of this syndrome has extended our understanding of the clinical and biochemical consequences of dysfunctional G protein action and has provided a bench-to-bedside demonstration of the critical role that G proteins play in transmembrane signal transduction in humans. PMID- 10714366 TI - Gs alpha mutations in hyperfunctioning thyroid adenomas. AB - Hyperfunctioning thyroid adenomas are benign tumors characterized by their autonomous growth and functional activity, which frequently cause clinical hyperthyroidism and show a predominant radioactive iodine uptake in the nodule. Activating mutations in the gene encoding the alpha subunit of the stimulatory G protein (Gs alpha), as well as activating mutations in the gene encoding thyrotropin receptor in hyperfunctioning thyroid adenomas, have been reported. The mutations in Gs alpha involved the replacement of either arginine 201 with cysteine or histidine, or glutamine 227 with arginine or leucine. These residues are involved in GDP/GTP binding of Gs alpha and these mutations inhibit intrinsic GTPase activity that results in constitutive activation of adenylyl cyclase. The pathophysiological roles of these mutations in the formation of hyperfunctioning thyroid adenoma have been suggested. PMID- 10714368 TI - G protein-coupled receptors as targets for prolactin actions. AB - Prolactin (PRL) is known to be involved in a wide range of biological functions including osmoregulation, lactation, reproduction, and immunomodulation. The first step in PRL action involves its interaction with a specific membrane receptor that belongs to the cytokine receptor superfamily. In spite of the lack of a kinase domain, receptors of the cytokine superfamily induce tyrosine phosphorylation of cellular substrates including the receptors. The role of PRL in female reproductive functions is well known and a direct effect on ovarian and testicular steroidogenesis has been established. In the ovary, PRL binds to a specific membrane receptor and exerts an inhibitory effect on follicular steroidogenesis. This effect is the result of an impairment involving FSH stimulation of G protein-coupled receptors (GPCR) and cyclic AMP-mediated activation of aromatase cytochrome P450 gene expression. This observation may indicate a direct connection between tyrosine phosphorylation and follicle stimulating hormone (FSH) receptor (FSHR) transduction pathways, as is the case for growth factor receptors with intrinsic tyrosine kinase activity, which share several downstream signaling elements with GPCRs. Some studies leading to our understanding of these pathways are reviewed. PMID- 10714369 TI - Where does cervical screening stand after the Kent and Canterbury Appeal judgement? PMID- 10714370 TI - Screening for the 21st century: learning from the past. PMID- 10714371 TI - Reactivity of six antibodies in effusions of mesothelioma, adenocarcinoma and mesotheliosis: stepwise logistic regression analysis. AB - Anti-CEA, anti-vimentin, CAM5.2, BerEp4, Leu-M1 and anti-EMA were applied to effusions from 36 mesotheliomas, 53 adenocarcinomas and 24 reactive mesothelial proliferations. Stepwise logistic regression analysis selected three criteria of major importance for distinguishing between adenocarcinoma and mesothelioma: BerEp4, CEA and EMA accentuated at the cell membrane (mEMA), these three being of similar diagnostic value. The pattern BerEp4-, CEA- and mEMA+ was fully predictive for mesothelioma (sensitivity 47%), whereas the opposite pattern was fully predictive for adenocarcinoma (sensitivity 80%). Only EMA seemed to distinguish between mesotheliosis and mesothelioma. Comparison of reactivity in cytological and histological material from the same mesotheliomas showed similar staining frequencies for CEA and CAM5.2, with some random variation for Leu-M1 and EMA, whereas vimentin and BerEp4 reactivity was more frequent in cytological specimens. PMID- 10714372 TI - Diagnostic value of lymph node fine needle aspiration cytology: an institutional experience of 387 cases observed over a 5-year period. AB - Lymph node fine needle aspiration (LNFNA) cytology is valuable in solving the diagnostic problems of clinical adenopathy. The usefulness of the procedure in the staging and diagnosis of various malignant and lymphoproliferative tumours, as well as its role in distinguishing reactive hyperplastic lymph nodes from lymphoma, has been documented in the literature generally on an individual basis. We report our cumulative 5 year experience of LNFNA representing 387 cases. Approximately half (n = 182) were diagnosed as either metastatic carcinoma or melanoma; in 54 cases (30%) excisional biopsy or tissue study was performed to confirm the diagnosis; there was only one false-positive diagnosis of a metastatic squamous carcinoma rendered on a submandibular lymph node. Sixty-one lymphoma cases were successfully diagnosed via LNFNA with no false positives; concurrent flow cytometry was utilized in 51% (n = 31) of the 61 cases and supported the cytologic diagnosis of lymphoma in 27 of the 31 cases (87%). A benign or reactive lymph node process was also diagnosed via LNFNA alone or in combination with flow cytometry in 48 cases with only five false negatives, which included four cases of mantle cell lymphoma and one case of melanoma. Unsatisfactory cases accounted for 12%, and represented specimens obtained by 'Wang needle' or other emerging techniques. Our study demonstrates that LNFNA can be an accurate, economical and rapid diagnostic procedure. PMID- 10714373 TI - Improved fine needle aspiration (FNA) cytology results with a near patient diagnosis service for breast lesions. AB - This study is a review of the quality of FNA cytology results for breast lesions approximately 18 months before and 10 months after a change from a rapid diagnosis FNA service with consultant pathologist aspirators to a conventional FNA service with clinician aspirators of varied experience. The setting was symptomatic breast clinic in a large hospital in rural New Zealand acting as a tertiary referral centre for a population of 550,000. The results were collected retrospectively and prospectively. The quality of results for pathologist aspirators (total 810) and clinician aspirators (total 403) was compared using the definitions of the NHS Breast Screening Program Guidelines for Cytology Procedures and Reporting in Breast Cancer Screening. There were statistically significant differences in specificity (biopsy cases only) with 73% for pathologists and 49% for clinicians, specificity (full) with 74% and 56%, inadequate rate with 23% and 37%, and complete sensitivity with 76% and 67%. The use of pathologist aspirators allowed the specimens to be reported in a few minutes. Specimens taken by clinicians took at least 30 min to report. The financial aspects of the two approaches are discussed. When compared with clinician aspirators, pathologist aspirators obtained better quality results and these were reported more quickly. PMID- 10714374 TI - Outcome of women referred to colposcopy for persistently inadequate smears. AB - The outcome of referral to colposcopy of 240 women who had persistently inadequate smears was investigated. Of 232 women who attended colposcopy, 214 (92.2%) had a normal outcome, 12 (5.2%) had low grade abnormalities, and six (2.6%) had high grade abnormalities. This group of women therefore has a negligibly increased risk of harbouring cervical neoplasia. Although not directly comparable, women with a history of previous abnormal cytology did not have a higher risk than those without such a history. Unnecessary colposcopy could have been avoided in the majority of cases if a good quality repeat smear had been taken. Improved smear taker training could decrease the number of referrals. A hospital cytology clinic is proposed as a cost-effective alternative to colposcopy at the first attendance. PMID- 10714375 TI - Cervical disease in women referred to colposcopy following inadequate smears. AB - The aim of this audit was to determine if inadequate cervical smears are associated with significant cervical pathology. Case records for 52 women with three consecutive inadequate smears referred for colposcopy to the Leicester Royal Infirmary (LRI) were retrieved. Sixteen women underwent large loop excision of the transformation zone (LLETZ) and cervical intraepithelial neoplasia (CIN) was identified in six cases. There were no cases of inadequate smears initiating the diagnosis in 100 consecutive women with invasive cervical cancer. Inadequate smears are associated with high rates of treatment for a low yield of CIN. To reduce morbidity associated with colposcopy it may be acceptable to repeat an inadequate smear after 6 months rather than arranging immediate recall. PMID- 10714376 TI - Cervical cytology through the looking glass. PMID- 10714377 TI - Inflammatory pseudotumour of the subcutis: a report on the fine needle aspiration findings in a case misdiagnosed cytologically as malignant. PMID- 10714378 TI - Fine needle aspiration cytology of malignant melanoma of soft parts: a case report and literature review. PMID- 10714379 TI - Fine needle aspiration cytodiagnosis of endometriosis in an abdominal scar after caesarean section. PMID- 10714380 TI - Who should be reporting gynaecological cytopathology? PMID- 10714381 TI - Ingestive behavior microstructure, basic mechanisms and clinical applications. PMID- 10714382 TI - Measuring hedonic impact in animals and infants: microstructure of affective taste reactivity patterns. AB - The hedonic impact of taste is reflected in affective facial reactions made by human infants, other primates, and even rats. Originally studied in human infants, affective reactions to taste have also been used by affective neuroscience to identify hedonic brain systems in studies of animals (via application of neural stimulation, pharmacological activation, and neural lesion manipulations). The chief limitation of measuring affective reactions is that it is difficult for experimenters to know how to interpret them, and therefore how to interpret changes produced by brain manipulations. This paper notes guidelines to interpretation. It examines the phylogenetic continuity between humans, other primates, and rats in terms of the microstructure of taste-elicited affective reactions. It reviews evidence that affective taste reactivity patterns truly reflect a 'core hedonic process' of palatability or affect, rather than being an ingestion measure, consummatory behavior measure, or a sensory reflex measure. It reviews affective neuroscience studies of taste reactivity that have identified true hedonic brain substrates, and discriminated them from false hedonic brain substrates. It considers the neural bases of incentive 'wanting' versus 'liking'. Finally, it notes the difference between human subjective affective ratings of pleasure and 'core hedonic processes' reflected by behavioral affective reactions. PMID- 10714383 TI - Microstructure of the rat's intake of food, sucrose and saccharin in 24-hour tests. AB - A detailed description of a method for collecting meal and drinking bout patterns for the rat over 24-h preference tests is presented. Both the hardware and software are described for data collection with a 6-s resolution. As an example of the value of knowing the details of ingestive patterns, data are presented describing the meal and drinking bout patterns during daylong preference tests between sucrose or saccharin vs. water. During these two-day tests at each of a wide range of concentrations, the intake of powdered Purina Chow, sucrose and saccharin were observed and quantified into meals or bouts. The analysis allowed for a comparison of sucrose and saccharin ingestion in terms of number of bouts, duration of bouts, rate of licking during bouts, the juxtaposition of eating and drinking and the day/night patterns of intake. Comparisons of drinking during bouts were made with electrophysiological and short-term taste test data. The usefulness of microstructure analysis is discussed. PMID- 10714384 TI - Meal patterning in rodents: psychopharmacological and neuroanatomical studies. AB - Studies of meal patterning have made an important contribution to our understanding of ingestive behaviour. This paper initially reviews studies of normal meal patterning in rodents, with an emphasis on the determination of suitable meal criteria. Studies of serotonergic mechanisms in the control of meal size and feeding rate suggest important roles for the 5HT1B and 5-HT2C receptor. Analysis of dopaminergic mechanisms show that dopamine D2 receptor blockade is associated with enhancement of meal size and decrease of meal frequency; this probably represents a failure to switch from feeding to other behaviour when a meal is expected to terminate. Finally studies are described demonstrating that lesions of several forebrain structures, including hippocampus and nucleus accumbens, lead to a similar syndrome of short, frequent meals with little evidence of a deficit in body weight regulation. These structures may play a role in the organisation of meal patterning. PMID- 10714385 TI - Chewing and swallowing as indices of the stimulation to eat during meals in humans: effects revealed by the edogram method and video recordings. AB - Patterns of chewing and swallowing were recorded during standardized meals in humans. Cocktail size (3 cm2) open sandwiches were served in one of five different flavors. An oscillographic recording of chewing and swallowing showed that chewing activity varied with the palatability and variety of foods. Chewing time was shorter and fewer chews were observed as palatability increased. Swallowing did not change as a function of stimulus flavor. Pause duration between two successive food pieces became shorter as palatability increased. The effects of sensory factors were most evident at the beginning of meals and decreased until the end of meals. A later study which compared eating parameters in sandwich, semi-solid, and traditional French meals (different courses ingested in succession: appetizer, main course, cheese and dessert), as assessed from video recordings, found that different microstructure parameters responded to palatability manipulation in different meal types. Strength of mastication and prandial drinking might be other important parameters to look at in order to understand the motivation to eat and its fluctuations during the meal. PMID- 10714386 TI - Rate of intake, bites, and chews-the interpretation of lean-obese differences. AB - The microstructure of eating behavior reflects physical properties of food. Responses of lean and obese subjects to these physical properties are similar. For example, eating smaller bite-sized food units reduces initial ingestion rate and mean and local ingestion rate for the entire meal, but does not affect total intake in either lean or obese women. On the other hand, analysis of the microstructure of eating behavior also suggests that obese subjects are less hungry and are more motivated by food preferences than lean subjects. For example, in meals of bite-sized food units, initial ingestion rate is less affected by deprivation and more affected by food preference in obese than lean women. In buffet meals with a variety of foods, obese men eat dessert earlier in the meal, and eat more dessert and other energy dense foods than lean men. The research reviewed here suggests that treatments for obesity should not focus on modifying bite size and ingestion rate and other microstructural variables, which are largely determined by the physical properties of food. Instead, treatment should focus on food selection and the stimulatory effects of palatability on intake. PMID- 10714387 TI - Eating behavior in humans, characterized by cumulative food intake curves--a review. AB - Cumulative food intake curves have been obtained by monitoring eating from a plate, placed on a scale built into a table, and connected to a digital computer. They describe and integrate parameters of consumption of an ad lib single course meal, i.e. meal size, meal duration, eating rate, change in eating rate, bite size and bite frequency. It is concluded that they are an adequate tool for analyzing dietary and clinical interventions on meal size, because the cumulative food intake curve parameters: are stable and consistent within subjects; show a clear relationship with the subject characteristics dietary restraint and obesity; show a clear relationship with the physiological parameters satiation, diet-induced thermogenesis and body-temperature near the liver, and with the cognitive parameter: estimating forthcoming ingestion; are sensitive to instructions, clinical and dietary interventions (preloads, palatability, energy density, macronutrient composition), and to a state of negative energy balance. Because of possible compensatory post-prandial effects, it is suggested that assessment of meal size should be part of a 24 h appetite profile and food intake observation. PMID- 10714388 TI - Rating changes over the course of meals: what do they tell us about motivation to eat? AB - Detailed analysis of the pattern of change in rated appetite within a meal have proved a useful technique through which to explore appetite control. Variability in individual ratings, and technical difficulties in achieving ratings at equivalent stages of a meal, have lead to the use of curve-fitting techniques to model changes in rated appetite across a meal. These changes could best be described by a quadratic function, in which the three parameters (intercept, linear and quadratic coefficients) represented distinct influences on meal size. In normal subjects, manipulations of palatability and opioid receptor blockade and preloads of alcohol all modified the linear component of this function only, while preloading with maltodextrin reduced appetite at the start of eating (the intercept) but not the pattern of change in ratings within that meal. Thus the linear coefficient appears to measure the degree of stimulation of appetite by the sensory characteristics of the food, while the intercept reflects baseline appetite at the start of a meal. These results suggest that microstructural analyses of rating changes allow some dissociation of the factors underlying motivation to eat, and provide a novel methodology for future experimentation. PMID- 10714389 TI - Microstructural analyses of human ingestive patterns: from description to mechanistic hypotheses. AB - Continuous automated weighing of food while subjects ate was used to test the hypothesis that failure to slow eating rate during a meal indicated a deficient response to satiety signals in obese patients. Cumulative intake curves were fitted to a quadratic equation. The physical form of the food and its palatability were a greater influence on the equation's parameters than the subjects' body weights, and the hypothesis was abandoned for several years (1984 1993). The hypothesis was revived with modifications when we discovered disturbances in eating behavior in patients with bulimia nervosa. The new hypothesis was that overeating was attributable to subjects' inability to detect or respond to satiety-related signals after eating large amounts of food. Patients with eating disorders showed lower ratings of satiety after eating the same amounts of food as controls, but only after eating more than normal. In conclusion, microstructural examination of eating behavior may be more useful for tests of specific hypotheses about the control of eating than as a description of clinical disturbance. PMID- 10714390 TI - Role of the alpha1 and alpha2 integrin cytoplasmic domains in cell morphology, motility and responsiveness to stimulation by the protein kinase C pathway. AB - The alpha1beta1 and alpha2beta1 integrins, extracellular matrix receptors for collagens and/or laminins, have similarities in structure and ligand binding. Recent studies suggest that the two receptors mediate distinct post-ligand binding events and are not simply redundant receptors. To discern the mechanisms by which the two receptors differ, we focused on the roles of the cytoplasmic domains of the alpha subunits. We expressed either full-length alpha1 integrin subunit cDNA (X1C1), full-length alpha2 integrin subunit cDNA (X2C2), chimeric cDNA composed of the extracellular and transmembrane domains of alpha2 subunit and the cytoplasmic domain of alpha1 (X2C1), chimeric cDNA composed of the extracellular and transmembrane domains of alpha1 subunit and the cytoplasmic domain of alpha2 (X1C2), alpha1 cDNA truncated after the GFFKR sequence (X1C0) or alpha2 cDNA truncated after the GFFKR sequence (X2C0) in K562 cells. Although the cytoplasmic domains of the alpha1 and alpha2 subunits were not required for adhesion, the extent of adhesion at low substrate density was enhanced by the presence of either the alpha1 or alpha2 cytoplasmic tail. Spreading was also influenced by the presence of an alpha subunit cytoplasmic tail. Activation of the protein kinase C pathway with phorbol dibutyrate-stimulated motility that was dependent upon the presence of the alpha2 cytoplasmic tail. Both the phosphatidylinosotide-3-OH kinase and the mitogen-activated protein kinase pathways were required for phorbol-activated, alpha2-cytoplasmic tail-dependent migration. PMID- 10714391 TI - Internalization of the E-cadherin/catenin complex and scattering of human mammary carcinoma cells MCF-7/AZ after treatment with conditioned medium from human skin squamous carcinoma cells COLO 16. AB - Cytokines and other paracrine or autocrine factors functionally modulate the invasion-suppressor and signal-transducing E-cadherin/catenin complex. We have used conditioned medium from human squamous carcinoma COLO 16 cells (CM COLO 16) as a source of such factors to modulate the E-cadherin/catenin complex in human breast carcinoma MCF-7 cells. CM COLO 16 induces scattering of MCF-7/AZ, but not of MCF-7/6 cells on tissue culture plastic substratum, and reduces aggregation of MCF-7/AZ cells in suspension. Insulin-like growth factor I counteracts this reduction of aggregation. Confocal laser scanning microscopy of immunocytochemical stainings shows loss of the honeycomb pattern of E-cadherin, alpha-catenin and beta-catenin, and internalization of those elements. Cell surface biotinylation shows a decrease in membrane-bound E-cadherin. Immunoprecipitation and cell fractionation show that the composition of the complex is maintained. Interleukin-1, interleukin-6, granulocyte-monocyte colony stimulating factor, stem cell factor, scatter factor/hepatocyte growth factor and transforming growth factor-beta, added separately to MCF-7/AZ cells, could not mimic the effects of CM COLO 16. Neither could we find evidence that the 80 kDa extracellular fragment of E-cadherin is implicated in scattering of MCF-7/AZ cells. This fragment is present in CM COLO 16, but it is also produced by the MCF 7/AZ cells themselves, even at higher levels. Our data point toward cytoplasmic internalization induced by paracrine factors as one of the downregulating mechanisms for the E-cadherin/catenin complex. PMID- 10714392 TI - Increased resistance to apoptosis in cells overexpressing thymosin beta four: A role for focal adhesion kinase pp125FAK. AB - Loss of adherence to substrate can, by itself, induce apoptosis (anoikis) in epithelial cells, but does not do so in fibroblasts. To test the idea that adherence transmits signals that inhibit apoptosis even in fibroblasts, we took advantage of the greatly increased adherence to the substratum observed in NIH3T3 cell lines that overexpress thymosin beta four. We treated overexpressing (OE) and vector control lines with either ultraviolet light (UV) or tumor necrosis factor alpha (TNF alpha). When the cells were on a substratum, the more adherent OE cells were 2-fold more resistant to apoptosis induced by either treatment than vector controls. In contrast, when the cells were treated with either agent while in suspension, the difference in resistance between OE cells and vector controls was lost. Thus the increased resistance to apoptosis was dependent on adherence. There was no difference in the content of bcl-2 in the OE cells vs the controls. A connection between pp125FAK and resistance to apoptosis has been previously shown in primary cultures of fibroblasts. The Tbeta4 overexpressing cells have approximately 1.4x more pp125FAK than the controls, and the kinase is approximately 2-fold more phosphorylated in adherent OE cells than in the vector controls. The phosphorylation of pp125FAK decreased strikingly when the cells were put into suspension. In addition, twice as much paxillin associated with pp125FAK in OE adherent cells as in vector controls, but this difference was also lost in suspended cells. Our results support the concept of an adherence dependent pp125FAK-paxillin signalling pathway in fibroblasts that inhibits damage-induced apoptosis. PMID- 10714393 TI - Combined interferon-gamma and tumor necrosis factor-alpha treatment differentially affects adhesion and migration of keratinocyte-derived cells to laminin-1. AB - Interactions with the extracellular matrix constitute basic steps in cervix carcinoma cell invasion. In this study, we examined the adhesion and migration profiles of two human papillomavirus (HPV) DNA-transfected keratinocyte-derived cell lines, EIL8 and 18-11S3, and of the cervix adenocarcinoma SiHa cell line, towards laminin-1, and the selective effect of a 24-72 h treatment of 1000 U/ml interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha (TNF-alpha), a treatment that significantly decreases cervix carcinoma cell proliferation and progression in nude mice, on these parameters. Compared to normal cervix keratinocytes (CK) and two HPV DNA-transfected keratinocyte cell lines, in basal conditions, the SiHa cell line was characterized by increased attachment (SiHa, 48.74 +/- 4.02 vs. normal keratinocytes, 4.32 +/- 0.40, EIL8, 17.80 +/- 3.03 and 18-11S3, 17.82 +/- 1.48% of attached cells after 30 min) and marked directed chemotactic migration towards laminin-1. Interestingly, treatment of the cells with the cytokines (1000 U/ml IFN-gamma and TNF-alpha) did not modulate the adhesion properties of the cells, but chemotactic migration of SiHa cells to laminin-1 was significantly decreased, while migration towards type I collagen was increased. Similar results were obtained with the Ca Ski cervix carcinoma cell line. Our results emphasize the altered pattern of interactions of cervix carcinoma cells with extracellular matrix components such as laminin-1, compared to normal and pre-neoplastic cells, and contributes to the understanding of the effects of cytokine treatment on cervix carcinoma cells. PMID- 10714394 TI - The CD44 receptor of lymphoma cells: structure-function relationships and mechanism of activation. AB - Migration of some tumor cells, and their lodgment in target organs, is dependent on the activation of cell surface CD44 receptor, usually detected by its ability to bind hyaluronic acid (HA) or other ligands. In an attempt to reveal the mechanism of tumor cell CD44 activation, we compared the physical and chemical properties of CD44 in nonactivated LB cell lymphoma with those in phorbol 12 myristate 13-acetate (PMA)-activated LB cells and of an LB cell subline (designated HA9) expressing constitutively-active CD44. In contrast to nonactivated LB cells, PMA-activated LB cells and HA9 cells displayed a CD44 dependent ability to bind HA. The ability of activated cell CD44 to bind HA was not dependent on microfilament or microtubule integrity or on changes in CD44 mobility on the membrane plane, indicating that the CD44 activation status is not associated with cytoskeleton function. Aside from the increased expression of CD44 on the surface of PMA-activated LB cells and HA9 cells, qualitative differences between the CD44 of nonactivated and activated LB cells were also detected: the CD44 of the activated lymphoma was (i) larger in molecular size, (ii) displayed a broader CD44 isoform repertoire, including a CD44 variant that binds HA, and (iii) its glycoprotein contained less sialic acid. Indeed, after removal of sialic acid from their cell surface by neuraminidase, LB cells acquired the ability to bind HA. However, a reduced dose of neuraminidase did not confer HA binding on LB cells, unless they were also activated by a low concentration of PMA, which by itself was ineffective. Similarly, under suboptimal conditions, a synergistic effect was obtained with tunicamycin and PMA: each one alone was ineffective but in combination they induced the acquisition of HA binding by the lymphoma cells, while their CD44 expression was not enhanced. Unveiling of the activation mechanism of CD44, by exposing the cells to PMA stimulation or to deglycosylation, is not only academically important, but it also has practical implications, as activated CD44 may be involved in the support of tumor progression. PMID- 10714395 TI - Adherence of platelets under flow conditions results in specific phosphorylation of proteins at tyrosine residues. AB - Collagen is a powerful platelet activating agent that promotes adhesion and aggregation of platelets. To differentiate the signals generated in these processes we have analyzed the tyrosine phosphorylation occurring in platelets after activation with collagen in suspension or under flow conditions. For the suspension studies, washed platelets were activated with different concentrations of purified type I collagen (ColI). Studies under flow conditions were performed using two different adhesive substrata: ColI and endothelial cells extracellular matrix (ECM). Coverslips coated with ColI or ECM were perfused through a parallel plate perfusion chamber at 800 s(-1) for 5 min. After activation of platelets either in suspension or by adhesion, samples were solubilized and proteins were resolved by electrophoresis. Tyrosine-phosphorylated proteins were detected in immunoblots by specific antibodies. Activation of platelet suspensions with collagen induced tyrosine phosphorylation before aggregation could be detected. Profiles showing tyrosine-phosphorylated proteins from platelets adhered on ColI or on ECM were almost identical and lacked proteins p95, p80, p66, and p64, which were present in profiles from platelets activated in suspension. The intensity of phosphorylation was quantitatively weaker in those profiles from platelets adhered on ECM. Results from the present work indicate that activation of platelets in suspension or by adhesion induces differential tyrosine phosphorylation patterns. Phosphorylation of proteins p90 and p76 may be related to early activation events occurring during initial contact and spreading of platelets. Considering that adhesion is the first step of platelet activation, studies on signal transduction mechanisms under flow conditions may provide new insights to understand the signaling processes taking place at earliest stages of platelet activation. PMID- 10714396 TI - The anticonvulsant SGB-017 (ADCI) blocks voltage-gated sodium channels in rat and human neurons: comparison with carbamazepine. AB - PURPOSE: SGB-017 (ADCI) is a novel anticonvulsant that blocks both voltage activated sodium channels and N-methyl-D-aspartate (NMDA)-receptor-gated channels. Results by Rogawski et al. suggested that SGB-017 produces its anticonvulsant action primarily by inhibition of NMDA-receptor channels. However, SGB-017 is effective in several animal models of epilepsy that are unresponsive to NMDA antagonists. These results indicate that block of NMDA-receptor channels is not the only mechanism contributing to its anticonvulsant activity. Thus the effects of SGB-017 on neuronal sodium channels were investigated. METHODS: Whole cell voltage-clamp techniques were used to record sodium currents in freshly dissociated rat superior cervical ganglion (SCG) and hippocampal neurons and cultured human NT2 neurons. The effects of SGB-017 on the amplitude of sodium currents, elicited by a depolarizing pulse to 0 mV from different holding potentials, were measured and compared with those of carbamazepine (CBZ). RESULTS: SGB-017 inhibited sodium currents in rat SCG and hippocampal neurons with a similar potency to CBZ. Like CBZ, the inhibition of sodium channels by SGB 017 was voltage dependent. Its median inhibitory concentration (IC50) for inhibition of sodium channels at depolarized holding potentials is similar to that for its inhibition of NMDA receptor channels. In human hNT2 neurons, SGB-017 was more potent than CBZ at inhibiting sodium currents. CONCLUSIONS: SGB-017 produces its anticonvulsant activity by blocking both sodium- and NMDA-receptor channels in a voltage- and use-dependent manner. The combination of these two mechanisms of action makes SGB-017 an effective AED in several different animal models of epilepsy. PMID- 10714397 TI - Cyclosporine induces epileptiform activity in an in vitro seizure model. AB - PURPOSE: Cyclosporine (CSA) toxicity represents a common cause of seizures in transplant patients, but the specific mechanisms by which CSA induces seizures are unknown. Although CSA may promote seizure activity by various metabolic, toxic, vascular, or structural mechanisms, CSA also has been hypothesized to modulate neuronal excitability directly. The objective of this study was to determine if CSA exerts direct epileptogenic actions on neurons in an in vitro seizure model. METHODS: Combined hippocampal-entorhinal cortex slices from juvenile rats were exposed directly to artificial cerebrospinal fluid (ACSF) containing either (a) 1.0 mM magnesium sulfate (control), (b) 1.0 mM sodium sulfate (low-magnesium), or (c) 1.0 mM magnesium sulfate + CSA (1,000-10,000 ng/ml). Spontaneous and evoked extracellular field potentials were recorded simultaneously from the dentate gyrus (DG) and CA3 hippocampal regions. Evoked synaptic responses were elicited by stimulation of the entorhinal cortex/perforant pathway. RESULTS: CSA elicited spontaneous or stimulation induced epileptiform activity in the DG or CA3 region of approximately 40% of slices, consisting of brief repetitive "interictal" discharges or prolonged stereotypical "ictal" discharges. Mean latency to epileptiform activity was approximately 100 min after onset of CSA application. The interictal discharges were inhibited by the non-NMDA antagonist, NBQX. Similar epileptiform activity was observed in low-magnesium ACSF without CSA. In control ACSF alone, epileptiform activity was not seen, except for rare spontaneous potentials in the DG. CONCLUSIONS: Direct effects of CSA on neuronal excitability and synaptic transmission may contribute to seizures seen in clinical CSA neurotoxicity. PMID- 10714398 TI - Seizures in rural Zambia. AB - PURPOSE: To describe the period prevalence of epilepsy and febrile seizures in a bush hospital and discuss the medical sequelae and social impact of seizures in this population. METHODS: For 13 weeks, an evaluation of inpatients was made at Chikankata Hospital in rural Zambia. Inpatients identified as having seizures, "fits," "spells," or "fainting," were evaluated by a medical records review, basic demographic data, a neurological history and physical examination, and a treatment history. A semistructured questionnaire was administered to evaluate the social impact of seizures and assess factors associated with delayed care seeking. RESULTS: Seizures composed 44% of all inpatient neurologic disease and resulted in 84 admissions. Epilepsy patients received treatment primarily from traditional healers; only 31% reported ever receiving antiepileptic drugs (AEDs). Among those who had received treatment, AEDs were frequently underdosed. Patients with epilepsy had significantly less education than their sex-matched siblings. Patients with untreated epilepsy for >2 years were more likely to have experienced serious burns or falls requiring hospitalization. Children with febrile seizures whose parents held supernatural beliefs regarding seizures were more likely to be treated with traditional medicines, had higher malarial parasite counts, and required longer hospitalizations than children with febrile seizures whose parents recognized the association between seizures and hyperthermia. CONCLUSIONS: Epilepsy and febrile seizures are responsible for a significant burden of disease in rural Zambia. Serious medical complications often result from seizures, especially if untreated for >2 years. Social stigma decreases educational opportunities and misperceptions regarding seizures may result in delayed care for children with febrile seizures. Some evidence suggests that epilepsy is underreported, underrecognized, and undertreated in this population. PMID- 10714399 TI - Chemical shift imaging spectroscopy and memory function in temporal lobe epilepsy. AB - PURPOSE: Hippocampal neuron loss and associated memory deficits are characteristic of intractable temporal lobe epilepsy (TLE). Proton chemical shift imaging (CSI) spectroscopy is a sensitive tool for detecting neuronal loss. The aim of this study was to investigate the correlation between memory functions and results provided by CSI spectroscopy of the hippocampal structures. METHODS: Ten patients with cryptogenic TLE participated. The study protocol involved both the acquisition of high-spatial-resolution CSI spectroscopy and neuropsychological evaluation, including memory testing and intracarotid sodium amytal test (IAT). The analysis of the CSI data was based on normative data obtained in 30 healthy volunteers. Memory functions were represented by verbal, visual, and general memory indices. RESULTS: A significant correlation was found between CSI spectroscopy of the hippocampal formation and the verbal memory indices for the dominant hemisphere. In addition, there was a significant correspondence of the qualitative judgment "hippocampal pathology indicated by CSI spectroscopy" and both "material specific memory deficit" and "memory deficit in the IAT." CONCLUSIONS: Our results demonstrate that CSI spectroscopy of the hippocampal structures is strongly related to lateralized memory deficits in patients with TLE. This suggests that CSI spectroscopy may be useful in the prediction of postoperative outcome in respect of seizure control and memory. PMID- 10714400 TI - MR spectroscopy shows reduced frontal lobe concentrations of N-acetyl aspartate in patients with juvenile myoclonic epilepsy. AB - PURPOSE: Neuropsychological studies suggest frontal lobe dysfunctions in patients with juvenile myoclonic epilepsy (JME). In this study we investigated whether an underlying mechanism could be a regional neuronal damage not visible with structural magnetic resonance (MR), but detectable with magnetic resonance spectroscopy (MRS). METHODS: The study included 15 patients with JME and 10 matched healthy controls. Quantitative single voxel MRS was conducted at 1.5 Tesla by using a STEAM sequence (TR/TE/TM = 6,000/30/13.7 ms). The voxels were placed over the right cerebellum, right thalamus, and the prefrontal and occipital cortex. The quantitation included fitting of transmitter gain, and correction for partial volume of cerebrovascular fluid. LC-Model was used for estimation of the absolute concentrations of total N-acetyl aspartate (NAA), cholines, total creatine, and myoinositol. RESULTS: Patients with JME had significantly reduced prefrontal concentrations of NAA in relation to controls (9.1 +/- 1.0 vs. 10.2 +/- 0.8 mM; p = 0.031 after Bonferroni correction). The other regions showed normal NAA values, as did the other metabolites. CONCLUSIONS: The observed reduction in NAA levels suggests a prefrontal neuronal lesion in patients with JME. PMID- 10714401 TI - Intracranial EEG seizure-onset patterns in neocortical epilepsy. AB - PURPOSE: We investigated neocortical seizure-onset patterns recorded by intracranial EEG with regard to anatomic location, pathologic substrate, and prognostic value for surgical outcome. METHODS: Seizure onset was analyzed in 53 neocortical resective epilepsy surgery patients. Anatomic location was divided into temporal and extratemporal. Pathologic substrate was classified as developmental, mature, and negative or non-specific gliosis. Onset frequency was categorized by visual analysis into tradition EEG frequency bands. Spatial extent was divided into focal (fewer than four contacts) and regional (more than five contacts). Waveform at seizure onset was divided into several types based on their morphology. Onset features were examined with respect to anatomic location, pathologic substrate, and surgical outcome. RESULTS: Seizure-onset frequency was significantly related to spatial distribution and to anatomic location. Extratemporal and regional onset were more commonly in the gamma range, and temporal and focal onset in the beta frequency range or slower. Waveform could be categorized into five different patterns, of which low voltage fast activity (LVFA) was the most common form (57%). LVFA and rhythmic alpha-theta spike activity were more common in developmental than in mature pathology, whereas rhythmic sinusoidal waves at onset were found in only mature substrates. Waveform pattern showed a possible correlation with surgical outcome (p = 0.097): LVFA and rhythmic sinusoidal waves onset patterns were associated with favorable outcome more often (40.4%) than the other three patterns (6.3%). Slow onset suggested poor outcome in the subgroup of developmental pathology (p = 0.062). CONCLUSIONS: Certain electrographic seizure-onset features are associated with specific substrates and outcomes, whereas others reflect the anatomic location and its connections independent of the pathology. PMID- 10714402 TI - Reduction of epileptiform activity in response to low-dose clonazepam in children with epilepsy: a randomized double-blind study. AB - PURPOSE: To evaluate the effect of low-dose clonazepam (CZP) on the amount of epileptiform activity in children with focal and generalized epilepsy. METHODS: In a single-blind pilot study, followed by a double-blind, placebo-controlled, randomized, crossover study, 15 children with epilepsy were evaluated by using 24 h long-term EEG recordings during baseline days and days after injections of placebo and CZP. The drug was given as a single i.m. injection of 0.02 mg/kg BW. Blood samples were obtained regularly for analysis of plasma concentrations of CZP. The number of epileptiform discharges was determined during corresponding periods with the individual child in the same state of alertness, the same real time of day, and with concomitant antiepileptic drugs (AEDs) unchanged. RESULTS: In the double-blind study, low-dose CZP produced a highly significant (p = 0.0015) decrease in the amount of epileptiform activity (mean, -69% vs. placebo, 2%) obtained during periods when median plasma concentrations ranged from 18 to <14 nM. The maximal plasma level (median, 24 nM) was reached before the start of the analysis periods. The pilot study showed reductions of epileptiform discharges within the same range as the double-blind study. In the children with daily seizures, a parallel decrease in seizures and the number of epileptiform discharges was seen after the administration of CZP. CONCLUSIONS: Our data demonstrate a significant reduction of epileptiform discharges on long-term EEGs after a single low dose of CZP with concomitant low plasma levels, which were considerably lower than the doses and plasma levels usually recommended. A concomitant reduction of seizures also was seen. PMID- 10714403 TI - Ictal spiking patterns recorded from temporal depth electrodes predict good outcome after anterior temporal lobectomy. AB - PURPOSE: Investigators have shown that the presence of ictal spiking (IS) recorded from temporal depth electrodes is associated with mesial temporal sclerosis (MTS). We investigated the relation of IS to seizure control and pathology after anterior temporal lobectomy (ATL). METHODS: All patients undergoing intracranial ictal monitoring from a single institution since 1989 were identified. Those who did not undergo ATL or had postoperative follow-up of <1 year were excluded. All received at a minimum bilateral temporal depth electrodes. Ictal recordings were reviewed for the presence of IS, and the proportion of seizures with IS was determined for each patient. Outcome was determined by using Engel's classification. Surgical specimens were reviewed for pathology. Statistics used were chi2, Fisher exact test, and Wilcoxon rank sum. RESULTS: Forty patients with 571 seizures were reviewed. In 292 seizures from 32 patients, IS was seen. Outcomes were 24 class I (22 with IS), five class II (four with IS), three class III (one with IS), seven class IV (four with IS), and one lost to follow-up (with IS). Pathologic review revealed 25 with MTS, 22 of whom had IS. The presence of IS was associated with class I outcomes (p = 0.04), but not MTS (p = 0.06). Patients with class I outcomes had a significantly greater proportion of seizures with IS (mean, 0.58 +/- 0.3) compared with other outcomes (mean, 0.30 +/- 0.3, p = 0.02). CONCLUSIONS: The presence of IS and higher proportion of seizures with IS correlated with good seizure outcome after ATL. This information may be used in preoperative counseling. PMID- 10714404 TI - The role of the intracarotid amobarbital procedure in evaluation of patients for epilepsy surgery. AB - PURPOSE: To examine the role of the intracarotid amobarbital procedure (IAP) in the presurgical evaluation of patients with medically refractory localization related epilepsy. METHODS: We retrospectively studied 111 patients who underwent cortical resective surgery at our center between 1991 and 1996. In patients with mesial temporal lobe epilepsy (mTLE), a presurgical determination of the epileptogenic zone was compared with localization based on IAP memory asymmetry scores, and with ultimate localization after resective surgery. In patients with neocortical or mesial frontal epilepsy, the IAP was evaluated for evidence of unilateral or bilateral poor memory performance. RESULTS: Of 68 patients with mTLE localized by noninvasive tests, 60 had concordant lateralized memory deficits on IAP. Eight patients had lateralized memory deficits on IAP that were discordant with noninvasive tests and with localization as determined by surgical outcome. All 11 mTLE patients requiring invasive EEG monitoring were correctly lateralized by IAP, including one patient in whom the noninvasive evaluation otherwise provided false lateralization. Of 32 patients with neocortical or mesial frontal lobe epilepsy, 21 displayed memory deficits on IAP. Of 10 patients with bilateral deficits, five had mesial frontal lobe epilepsy. In 13 patients with lateralized memory deficits, seven underwent electrode implantation in the mesial temporal lobe, and four ultimately underwent resection of an epileptogenic mesial temporal lobe in addition to a neocortical resection. CONCLUSIONS: In patients with mTLE, lateralized memory deficits on IAP usually confirm localization provided by noninvasive tests. However, in mTLE not well lateralized by the noninvasive evaluation, and in neocortical or mesial frontal epilepsy, the IAP may provide information regarding localization that ultimately alters surgical management. PMID- 10714405 TI - Negative emotions in children with newly diagnosed epilepsy. AB - PURPOSE: To understand the emotional predicament in children with recently diagnosed idiopathic or cryptogenic epilepsy. METHODS: We used the well-tried method of structured projection for the first time in children with epilepsy. Thirty-six children with epilepsy, aged 7-15 years (mean age, 9.5 years) and in 35 control children aged 7-15 years (mean age, 9.4 years), attributed shame and guilt in relation to three types of situation (non-illness related, illness related, and epilepsy related). Children were evaluated twice: shortly after diagnosis, before antiepileptic drug (AED) use and after an interval of 3 months. RESULTS: Children with epilepsy and healthy controls were similar in their way of attributing shame and guilt. However, the type of situation was of influence: Both children with epilepsy and healthy children attributed more shame to incompetence due to epilepsy than to incompetence due to other illnesses. CONCLUSIONS: Increased affective problems in childhood epilepsy cannot be explained by excessive attribution of shame and guilt, affects known to be important precursors of psychopathology, yet both healthy children and children with epilepsy attribute more shame to epilepsy than to other illnesses. Epilepsy is not like any other disease. PMID- 10714406 TI - Improved prediction of nonepileptic seizures with combined MMPI and EEG measures. AB - PURPOSE: Nonepileptic seizures (NESs) are frequently mistaken for epileptic seizures (ESs). Improved detection of patients with NESs could lead to more appropriate treatment and medical cost savings. Previous studies have shown the MMPI/MMPI-2 to be a useful predictor of NES. We hypothesized that combining the MMPI-2 with a physiologic predictor of epilepsy (routine EEG; rEEG) would result in enhanced prediction of NES. METHODS: Consecutive patients undergoing CCTV-EEG monitoring underwent rEEG evaluation and completed an MMPI-2. Patients were subsequently classified as having epilepsy (n = 91) or NESs (n = 76) by using standardized criteria. Logistic regression was used to predict seizure type classification. RESULTS: Overall classification accuracy was 74% for rEEG, 71% for MMPI-2 Hs scale, and 77% for MMPI-2 Hy scale. The model that best predicted diagnosis included rEEG, MMPI-2, and number of years since the first spell, resulting in an overall classification accuracy of 86%. CONCLUSIONS: The high accuracy achieved by the model suggests that it may be useful for screening candidates for diagnostic telemetry. PMID- 10714407 TI - Prognostic factors affecting long-term retention of topiramate in patients with chronic epilepsy. AB - PURPOSE: To determine the long-term retention rate of topiramate (TPM) therapy in patients with chronic epilepsy and to identify the relevant prognostic factors that influence retention. METHODS: All patients with chronic epilepsy (n = 393) prescribed TPM between October 1, 1995, and December 31, 1998, at a tertiary referral centre for epilepsy were analysed. The retention rate for TPM was calculated by using Kaplan-Meier survival analysis, and the prognostic factors influencing retention were analysed by using Cox regression. RESULTS: Of patients prescribed TPM, 30% continued taking the drug beyond 3 years. Discontinuation was mainly due to adverse events and lack of efficacy. Use of more than one new concurrent antiepileptic drug (AED) and lower maximal daily doses were more likely to result in treatment discontinuation due to adverse events. Older age at onset of epilepsy, a history of having previously taken more than one new AED [lamotrigine (LTG), gabapentin (GBP), or vigabatrin (VGB)], and lower maximal daily doses were more likely to lead to discontinuation due to lack of efficacy. CONCLUSIONS: A third of patients with chronic epilepsy started on TPM therapy will continue on treatment for >3 years. Absence of learning disabilities, late age at onset of seizures, previous use of more than one new AED, two or more concurrent AED use, and low maximal daily doses of TPM are more likely to result in discontinuation of medication. These factors should be taken into account when considering the use of TPM for the treatment of chronic epilepsy. PMID- 10714408 TI - The cost of epilepsy in the United States: an estimate from population-based clinical and survey data. AB - PURPOSE: To provide 1995 estimates of the lifetime and annual cost of epilepsy in the United States using data from patients with epilepsy, and adjusting for the effects of comorbidities and socioeconomic conditions. METHODS: Direct treatment related costs of epilepsy from onset through 6 years were derived from billing and medical chart data for 608 population-based incident cases at two sites in different regions of the country. Indirect productivity-related costs were derived from a survey of 1,168 adult patients visiting regional treatment centers. Direct costs separate the effects of epilepsy and comorbidity conditions. Indirect costs account for the effects of other disabilities and socioeconomic conditions on foregone earnings and household activity. The estimates were applied to 1995 population figures to derive national projections of the lifetime and annual costs of the disorder. RESULTS: The lifetime cost of epilepsy for an estimated 181,000 people with onset in 1995 is projected at $11.1 billion, and the annual cost for the estimated 2.3 million prevalent cases is estimated at $12.5 billion. Indirect costs account for 85% of the total and, with direct costs, are concentrated in people with intractable epilepsy. CONCLUSIONS: Direct costs attributable to epilepsy are below previous estimates. Indirect costs adjusted for the socioeconomic conditions of patients are above previous estimates. Findings indicate that epilepsy is unique in the large proportion of costs that are productivity-related, justifying further investment in the development of effective interventions. PMID- 10714409 TI - Bilateral periventricular and subcortical heterotopia in a man with refractory epilepsy. AB - PURPOSE: To report a novel malformation in a male subject with refractory partial seizures. METHODS: Magnetic resonance imaging (MRI) and data reformatting in a subject referred for management of partial seizures. RESULTS: The patient had four distinct partial seizure types, without learning disability. MRI demonstrated the novel association of bilateral laminar subcortical heterotopia, bilateral temporal periventricular heterotopia, and hippocampal malformation. CONCLUSIONS: This previously unreported complex bilateral neocortical and archicortical malformation in a male patient cannot be explained by known genetic causes of heterotopia, raising the possibility of a novel gene involved in brain formation. PMID- 10714410 TI - Practice advisory. PMID- 10714411 TI - Poor surgical outcome in patients with neocortical epilepsy is correlated with interictal epileptiform abnormalities outside the area of surgical resection. PMID- 10714412 TI - Efficacy of lamotrigine (LTG) monotherapy. PMID- 10714413 TI - Efficacy and safety of ten day moxifloxacin 400 mg once daily in the treatment of patients with community-acquired pneumonia. Community Acquired Pneumonia Study Group. AB - Community-acquired pneumonia (CAP) remains a common and serious illness with approximately 2-4 million cases reported annually. Management of CAP is therapeutically challenging due to the increasing prevalence of penicillin- and macrolide-resistant pneumococci and beta-lactamase producing Haemophilus influenzae, as well as the increased recognition of 'atypical' pathogens, such as Chlamydia pneumoniae and Mycoplasma pneumoniae, and the frequent need for empiric therapy. We aimed to evaluate the safety and efficacy of moxifloxacin in the treatment of patients with CAP. To do this we carried out a prospective, uncontrolled, non-blind, Phase III clinical trial, in 27 U.S. centers. Patients included in the study were over 18 years of age with signs and symptoms of CAP confirmed by evidence of a new or progressive infiltrate on chest radiograph. The intervention used was moxifloxacin 400 mg PO once daily for 10 days. Sputum samples were collected pretherapy for Gram stain and culture for typical organisms. Culture and serological testing for Chlamydia pneumoniae and Mycoplasma pneumoniae was also performed. Susceptibility to moxifloxacin was determined by disk diffusion and MIC. Clinical and bacteriological responses were determined at the end of therapy (0-6 days post-therapy), follow-up (14-35 days post-therapy) and overall (end of therapy plus follow-up). Analyses were performed on both valid for efficacy and intent-to-treat populations. The primary efficacy variable was overall clinical resolution. Of 254 patients enrolled in the Study, 196 patients were included in the efficacy analyses. The majority of patients were male (58%) and Caucasian (85%) with a mean age of 49 years (range: 18 to 85 years). Only 3% of patients were hospitalized pretherapy. The most common pretherapy organisms identified, by culture or serology, in the valid for efficacy population (i.e. 147 organisms among 116 patients), were: Chlamydia pneumoniae (n=63; 54%), Mycoplasma pneumoniae (n=29; 25%), Streptococcus pneumoniae (n=14; 12%) and Haemophilus influenzae (n=13; 10%). End of therapy, follow-up and overall clinical resolution rates for the valid for efficacy population were 94%, 93% and 93%, respectively. The 95% CI for the overall clinical resolution rate was 88.1%, 95.9%. The overall bacteriological response for patients diagnosed by culture or serological criteria, was 91% (95% CI=84%, 96%). For patients who only met serological criteria for infection, the overall bacteriological response was 94% (60/64). Bacterial response rates for the four most commonly isolated pathogens were: 89% (56/63) for C. pneumoniae, 93% (27/29) for M. pneumoniae, 93% (13/14) for S. pneumoniae and 85% (11/13) for H. influenzae. Drug-related adverse events were reported in 33% (85/254) of moxifloxacin-treated patients. Nausea (9%), diarrhea (6%) and dizziness (4%) were the most commonly reported adverse events. Atypical organisms were isolated in high frequency among patients with CAP. Moxifloxacin 400 mg once daily for 10 days was effective and well-tolerated in the treatment of these adult patients with CAP. Moxifloxacin offers an effective treatment alternative for CAP due to both typical and atypical bacterial pathogens. PMID- 10714414 TI - Production of the potent neutrophil chemokine, growth-related protein alpha (GROalpha), is not elevated in cystic fibrosis children. AB - Progressive neutrophil-mediated lung damage causes much of the morbidity and mortality in cystic fibrosis (CF). Neutrophil chemoattractants implicated in CF include interleukin (IL-)8, tumour necrosis factor (TNFalpha) and leukotriene (LT)B4, but growth-related protein alpha (GROalpha), a highly potent neutrophil chemokine, has not been investigated. Atopic status has been considered to contribute to the marked heterogeneity of pulmonary disease in CF. We hypothesized that GROalpha may be produced in biologically-significant amounts in the CF lung, and that enhanced production of GROalpha, IL-8 or LTB4 may contribute to the poorer lung function seen in atopic CF patients compared to non atopic CF patients. GROalpha, IL-8 and LTB4 levels in the sputum of atopic and non-atopic CF patients were assessed by immunoassays, and GROalpha and IL-8 levels were also assessed in the plasma of CF patients and normal controls. As expected, there were high levels of IL-8 and LTB4 in most CF sputum samples, and IL-8 levels were higher in CF plasma than in control plasma (P=0.02). In contrast, GROalpha was undetectable (< 5 pg ml(-1)) in the sputum of 21 out of 25 CF patients, with low levels (range 144-825 pg ml(-1)) in the remainder, and median levels of GROalpha in CF plasma (33 pg ml(-1), n=24) were not significantly different from controls (34 pg ml(-1), n=25). Lung function [forced expiratory volume in 1 sec (FEV1) and forced vital capacity (FVC)] was significantly poorer in atopic CF compared to non-atopic CF patients (P<0.02), but sputum levels of GROalpha, IL-8 and LTB4 were not different between the subgroups. Our results suggest that unlike LTB4 and IL-8, GROalpha does not contribute to neutrophilic inflammation in the CF lung, and other factors must determine the impaired lung function observed in atopic CF patients. These results may have important implications in the development of chemokine receptor antagonists as novel anti-inflammatory agents in CF. PMID- 10714415 TI - Effects of 2 weeks of treatment with fluticasone propionate 100 mcg b.d. by comparison with zafirlukast 20 mg b.d. on bronchial hyper-responsiveness in patients with mild to moderate asthma. AB - This study was designed to compare the effects of low-dose inhaled fluticasone propionate (100 mcg twice daily) with those of the leukotriene antagonist, zafirlukast (20 mg twice daily), on bronchial hyper-responsiveness. The study recruited 30 patients (nine men, 21 women; mean age 45 years) with forced expiratory volume in 1 sec (FEV1) > 50% and airway reversibility to salbutamol > or =15%. This was a single centre, double-blind, double-dummy cross-over study, composed of two successive 2-week treatment periods, each preceded by a 2-4 week single-blind placebo period. Following 2 weeks of treatment with fluticasone propionate and zafirlukast, the mean provocational concentration causing a 20% fall in FEV1 (PC20) histamine was 1.61 mg ml(-1) (SD 2.34) and 0.99 mg ml(-1) (SD 1.74) respectively. Taking baseline differences into account, the difference between treatments was equivalent to 0.77 doubling doses of histamine (95% CI, 0.05-1.50; P=0.037). Morning peak flow values were significantly higher (17 l min(-1); P=0.049) after treatment with fluticasone propionate during the second week of treatment. Both treatments were well tolerated. The results of this short term study show that compared with zafirlukast, a low dose of fluticasone propionate offers greater clinical benefit and is more cost effective. PMID- 10714416 TI - Bronchial reactivity to cigarette smoke; relation to lung function, respiratory symptoms, serum-immunoglobulin E and blood eosinophil and leukocyte counts. AB - STUDY OBJECTIVES: The aim of the study was to investigate the relationship between the immediate bronchial response to inhaled cigarette smoke [cigarette smoke bronchial reactivity (CBR)] and lung function, respiratory symptoms and markers of allergy and inflammation. DESIGN, PARTICIPANTS AND MEASUREMENTS: This cross-sectional study included 98 smokers. Their lung function and reversibility to inhaled terbutaline was measured. Their clinical history was obtained, an allergological examination was done, and bronchial reactivity to methacholine and inhaled cigarette smoke was measured. Questionnaires about respiratory symptoms, smoking history and drug usage were completed and a blood sample was obtained. Participants were divided into three groups: with asthma, chronic bronchitis and persons without asthma or chronic bronchitis (the respiratory healthy). RESULTS: Forced expiratory volume in 1sec (FEV1) residuals were independently related to the % fall in FEV1 after 12 cigarette smoke inhalations (DFEV%) in all participants (P<001), in asthmatic smokers (P<0.01) and in smokers with chronic bronchitis (P<0.05). In smokers with asthma and chronic bronchitis FEV1 residuals explained 51% and 13% of the variation in DFEV%, respectively, but only 8% (P<0.05) and 1% (N.S.) of the variation in the methacholine bronchial reactivity. In the total population the presence of wheeze (P<0.01), attacks of breathlessness (P<0.05) and daily expectoration (P<0.001) were related to higher DFEV% readings. Serum immunonoglobulin (ES-IgE) was independently related to DFEV% in all participants (P<0.01), in smokers with chronic bronchitis (P<0.01) and in the respiratory healthy (0.05 15% and 15 patients did not. There was a statistically significant difference in perception of dyspnoea (P<0.01), between the group of patients with a improved FEV1 and the group of patients that were under IGC treatment without improvement in their FEV1. There was also a difference in the mean beta2-agonists consumption between the two groups (P<0.01). Asthmatic patients have a significantly lower perception of dyspnoea compared to normal subjects. IGC treatment was associated with increased perception of dyspnoea. However, this improvement was noted only in patients with improved FEV1, while the patients without improvement remained with an equal degree of dyspnoea perception and beta2-agonists consumption. PMID- 10714424 TI - Pulmonary function and airway responsiveness in mild to moderate asthmatics given repeated inhaled doses of zanamivir. AB - Zanamivir is a potent and specific inhibitor of influenza A and B virus neuraminidase, that is now approved for the treatment, and is currently under development for the prophylaxis of influenza. To assess the safety of this drug in asthmatics, 13 subjects with mild/moderate asthma [forced expiratory volume in 1 sec (FEV1)> or =70% predicted, reversibility of FEV1 to salbutamol > or =15%, concentration of methacholine causing a drop of 20% in the FEV1 (PC20FEV1)< or =8 mg ml(-1)], were recruited to a double-blind, randomized, placebo controlled, two way cross-over study. Subjects received 10 mg zanamivir as a dry powder (2 x 5 mg blisters via a Diskhaler Sovnn Plastics Ltd., Berkshire, U.K.), or a matching placebo, twice daily on day 1 and then four times daily from day 2 to day 14, in two separate periods separated by a washout period of 7 days. PC20FEV1 to methacholine was determined pre-study, on day 1 after the evening dose and on day 14 after the last dose of the study drug. FEV1 was measured pre-study and at regular intervals on days 1 and 14. Laboratory safety tests were performed on days 1, 7 and 15. Morning and evening peak expiratory flow rate (PEFR) and any adverse events were recorded in a diary card. Eleven subjects completed the study. One was withdrawn due to non-compliance, and one due to an adverse event that occurred during the placebo period. On day 1 the geometric mean PC20 for zanamivir was 36% lower than for placebo [ratio to placebo 0.64, (90% CI 0.44, 0.93)] and on day 14 this was 33% lower with zanamivir [ratio to placebo 0.67 (90% CI 0.38, 1.15)]. Both these confidence intervals were within the pre-defined interval of 'no clinically significant effect' of 0.25-4 (i.e. a change of two doubling doses of methacholine PC20FEV1 which was considered clinically significant). The time weighted mean FEV1 was 0.15 l (5.4%) lower for zanamivir on day 1 compared to placebo (90% CI 0.03, 0.28; P=0.050) and 0.01 l higher compared to placebo on day 14 (90%CI -0.12, 0.10; P=0.912). The day 1 changes were not associated with any significant symptoms or requirement for rescue bronchodilator therapy. Furthermore there was no apparent treatment difference over the 14 day dosing period in FEV1 data (90% CI: -0.11, 0.05, P=057). The mean morning PEFR was 4 l min(-1) less for zanamivir than for placebo (90% CI: -11, 3) and mean evening PEFR was 9 l min(-1) less (90% CI: -24, 5). The study treatments were well tolerated by the subjects with no clinically significant adverse events attributable to zanamivir treatment. Zanamivir inhaled as a dry powder does not significantly affect the pulmonary function and airway responsiveness of subjects with mild/moderate asthma and therefore its use in such patients subjects is not precluded. PMID- 10714425 TI - Inhaled heparin is effective in exacerbations of asthma. PMID- 10714426 TI - A hazard of Christmas: Bird Fancier's Lung and the Christmas tree. AB - A lady with alveolitis due to her budgerigar developed recurrent symptoms when exposed to allergen left on her artificial Christmas tree. PMID- 10714427 TI - Equivalence of hydrofluoroalkane (HFA) and chlorofluorocarbons (CFC) formulations of inhaled beclomethasone. PMID- 10714428 TI - Is inhaled beclomethasone (BDP) with a non-CFC propellant equivalent to the CFC propellant formulations? PMID- 10714429 TI - Re: inhaled beclomethassone (BDP) with non-CFC propellent (HFA 134a) is equivalent to BDP-CFC for the treatment of asthma (Respir Med 19999; 93:245-251) PMID- 10714430 TI - Inhaled beclomethasone (BDP) with non-CFC propellant (HFA-134a) is equivalent to BDP-CFC for the treatment of asthma: Milanowski et al. (Respir Med 1999; 93: 245 251) PMID- 10714431 TI - Inhaled beclomethasone (BDP) with non-CFC propellant (HFA 134a) is equivalent to BDP-CFC for the treatment of asthma (Respir Med 1999; 93: 245-251) PMID- 10714432 TI - The pattern of a T(H)1 cytokine in autoimmune thyroiditis. AB - This study performed on 51 patients with autoimmune thyroiditis and 15 healthy subjects was aimed at correlating the activation of T cells and the secretion of inflammatory cytokine with echography and thyroid functional assays. A significant increase of activated T cells was observed in Hashimoto patients illustrated by an increased percentage of CD3+ CD25+ T cells (P < 0.01). Similarly, increased amounts of IL-2, TNF-alpha and IFN-gamma were in the serum of patients compared with the control group in which these cytokines are barely detectable. An in vitro study shows a significant increase of IL-2 and TNF-alpha upon the exposure to Concanavalin A. These results suggest that T(H)1 secreting inflammatory cytokines may contribute to pathogenesis of autoimmune thyroiditis. PMID- 10714433 TI - N-Acetylcysteine and glutathione modulate the behaviour of Trypanosoma cruzi experimental infection. PMID- 10714434 TI - The analysis of in vitro transforming growth factor-beta1 (TGF-beta1) production by peripheral blood in overt and pre-clinical type 1 diabetes mellitus. AB - The alterations of TGF-beta1 production are believed to contribute to the development of insulin-dependent diabetes mellitus (IDDM) in animal models as well as in humans. There is also increasing evidence about the role of this cytokine in the pathogenesis of diabetic vascular complications. The aim of our study was to evaluate in vitro TGF-beta1 production by peripheral blood of newly diagnosed type 1 diabetes patients and subjects in the pre-clinical stage of the disease in comparison to healthy controls and relatives of IDDM patients with low genetic risk for diabetes development. The study was carried out in three groups of subjects: 22 patients with a recently diagnosed type 1 diabetes, their 24 first degree relatives with a different genetic risk of IDDM development and 18 healthy volunteers (control group). In all studied groups whole blood was taken for morphology parameters. HbA1C and for 72 h cultures with PHA stimulation for the estimation of TGF-beta1 in vitro production. TGF-beta1 concentration in supernatants were quantified by ELISA. In the first degree relatives HLA typing (for DR3, DR4 and DQB1*0602 alleles), measurements of anti-pancreatic antibodies (ICA, GADA, IA-2A, IAA) and intravenous glucose tolerance tests were performed. The levels of TGF-beta1 in the supernatants were significantly higher in diabetic patients (P < 0.0002) and in their first degree relatives (P < 0.05) in comparison to the control group. In the group of first degree relatives TGF-beta1 levels were highest in subjects with the presence of two or more pancreatic autoantibodies and/or with impaired insulin release in IVGTT, but lowest in relatives with protective DQB1*0602 alleles (P < 0.01). There was also a significant positive correlation between the TGF-beta1 levels and HbA1C in the IDDM subjects and first degree relatives (P < 0.03). Our study suggests that the alterations of TGF-beta1 levels could be associated with the activity of autoimmune process leading to pancreatic B cells destruction and may have a role in the pathogenesis of diabetic complications, but further studies in humans are needed. PMID- 10714435 TI - Preservative-free influenza vaccine. AB - OBJECTIVE: To investigate the immunogenicity of and tolerance towards the preservative-free inactivated influenza vaccine Begrivac. METHODS: In this prospective, single-centre, non-controlled study, efficacy was evaluated by the change in influenza antibody titre from baseline to 21 days following vaccination. The safety variables included post-injection reactions and adverse events. Blood samples were taken on day 21 and the antibody titre assayed by haemagglutination inhibition test. RESULTS: All three of the European efficacy requirements for influenza vaccines are satisfied by the new preservative-free vaccine described in this report. The mean geometric increase in titre and the proportion of vaccination responders were greater in patients of the adult group than in the elderly. Thus for strain A/Beijing/262/95 66% of subjects seroconverted and 28% showed a significant increase in antibody titre (total 94%), compared to a total of 45 patients (76%) in the elderly group. For strain A/Sydney/5/97 the corresponding figures were total 55 (90%) adult and 47 (80%) elderly, and for B/Beijing/184/93 46 (75%) adult and 31 (53%) elderly. Sixty four subjects (53%) reported adverse events, mainly local reactions at the injection site such as pain, erythema and induration, and systemic reactions such as headache and fatigue. CONCLUSIONS: The absence of preservative in this novel vaccine preparation does not have any detectable impact on its efficacy or safety and tolerability profile. PMID- 10714436 TI - Neutrophils augment the release of TNFalpha from LPS-stimulated macrophages via hydrogen peroxide. AB - We examined the effect of polymorphonuclear cells on the release of tumor necrosis factor (TNFalpha) in endotoxin-treated macrophages. Human peripheral blood neutrophils were co-cultured with mouse peritoneal macrophages stimulated with lipopolysaccharide (LPS). In a dose-dependent manner, FMLP (n-formyl methionyl-leucyl-phenylalanine) augmented the release of TNFalpha by LPS stimulated macrophages in the presence, but not in the absence, of neutrophils. The stimulating effect of neutrophils on macrophages was reversed by catalase, suggesting that the release of hydrogen peroxide from neutrophils was responsible for augmenting macrophage TNFalpha. Moreover, the direct addition of hydrogen peroxide to macrophages resulted in an increased secretion of TNFalpha. In addition, insertion of a porous membrane between the neutrophils and macrophages cancelled the effect, indicating that adherence of neutrophils may be necessary for augmentation of TNFalpha release. In summary, the data suggest that hydrogen peroxide released from stimulated neutrophils may act as an activator of macrophage function by increasing their release of TNFalpha. PMID- 10714437 TI - IL-12 and IFN-gamma production, and NK cell activity, in acute and chronic experimental Trypanosoma cruzi infections. AB - Resistance to acute Trypanosoma cruzi infection is mainly associated with a Th1 immune response, characterized by gamma-interferon (IFN-gamma) production and activation of macrophages. The outcome of the Th1 response in the spleen and serum of BALB/c and C3H mice infected with T. cruzi, Tulahuen strain was studied. The levels of interleukin-12 p40 (IL-12 p40) and IFN-gamma, as well as natural killer (NK) cell cytotoxicity were determined at different time-points during the acute phase, and the production of cytokines was also studied in the chronic infection. At 2 days post-infection (pi), spleen cells from C3H mice increased their NK cell activity and the ex vivo spontaneous release of both IL-12 p40 and IFN-gamma. On the other hand, BALB/c mice reached low levels of NK cell cytotoxicity and no IFN-gamma production was detected at this time pi, but the cytokine was released at high amounts in the second week of the infection. Seric IL-12 p40 concentrations showed a 3-fold increase in both mouse strains on the second day pi and remained high throughout the acute phase. However, seric IFN gamma levels increased during the late acute infection and were higher in BALB/c than in C3H mice. In chronically infected mice IL-12 p40 was as high as in the acute phase in the serum of both strains, but only BALB/c mice still produced IFN gamma. To the authors' knowledge this is the first report showing the protein levels of IL-12 p40 determined in vivo in acute and chronic T. cruzi infections. The results reveal differences between both mouse strains in the mechanisms controlling the onset and fate of the Th1 response triggered by the parasite and a long lasting pro-inflammatory stimuli. PMID- 10714438 TI - Antigen binding characteristics of antibodies against hydroxyl radical modified thymidine monophosphate. AB - Reactive oxygen species (ROS) are formed in living organisms during normal metabolic reactions as well as under different environmental stresses. In this study, thymidine monophosphate (TMP) was exposed to hydroxyl radical (OH) and challenged in rabbits. TMP and OH-modified TMP were found to be nonimmunogenic. The TMP was linked to bovine serum albumin (BSA) by carbodiimide reaction, and then modified with the OH. The neoantigens, TMP-BSA, and ROS-TMP-BSA conjugates induced highly specific antibodies against immunogens. Induced antibodies exhibited appreciable cross-reactivity with various polynucleotides and nucleic acids. In this respect, the induced antibodies resembled the diverse antigen binding characteristics of naturally occurring systemic lupus erythematosus (SLE) anti-DNA autoantibodies. PMID- 10714440 TI - Increased expression of neutral endopeptidase (NEP) and aminopeptidase N (APN) on peripheral blood mononuclear cells in patients with multiple sclerosis. AB - We studied CD10 Ag of neutral endopeptidase (NEP) and CD13 Ag of aminopeptidase N (APN) expression on peripheral blood mononuclear cells (PBMC) as cells activation markers and for their transendothelial migration properties in three groups of MS patients (total 58); with acute exacerbation of MS (n = 18), primary progressive MS (n = 17) and with MS remission (n = 23). The control group (OND) consisted of 24 patients, suffering from other noninflammatory neurological diseases. CD10 Ag and CD13 Ag expression on PBMC was higher in clinically active MS (acute exacerbation and progressive MS) compared to MS remission and OND groups. Our study suggests that CD10 Ag and CD13 Ag can be useful mononuclear cell activation markers in the course of MS. CD13 Ag expression on PBMC may be also the sensitive marker of these cells transendothelial migration properties. PMID- 10714439 TI - Antibodies to variable Plasmodium falciparum-infected erythrocyte surface antigens are associated with protection from novel malaria infections. AB - In areas of unstable transmission malaria affects all age groups, but the malaria incidence is lower in adults compared to children and teenagers. Under such conditions subclinical Plasmodium falciparum infections are common and some infections are controlled, because blood parasitaemia is maintained at low densities. Here, we test the hypothesis that the presence or absence of antibodies against variant antigens on the surface of P. falciparum-infected erythrocytes protect individuals against some infectious challenges and render them susceptible to others. Plasma collected in Daraweesh, eastern Sudan, before and after the malaria season from individuals who had (susceptible) or did not have malaria (protected) during the season, were tested for reactivity against variant antigens on the surface of nine parasite isolates by flow cytometry. Both protected and susceptible individuals acquired antibodies to variant antigens during the malaria season. The presence of antibody to a Ghanaian isolate before the season was statistically significantly associated with protection against malaria. When considering all nine isolates, the patterns of antibody acquisition differed between susceptible and protected individuals. Together, the results indicate that pre-existing anti-PfEMP1 antibodies can reduce the risk of contracting clinical malaria when challenged by novel parasite clones expressing homologous, but not heterologous variable surface antigens. The results also confirm that antibodies to variant antigens are induced by both clinical and subclinical infections, and that antibodies against several var sero-types are induced during an infection. PMID- 10714441 TI - Immune and pathophysiological processes in baboons experimentally infected with Ebola virus adapted to guinea pigs. AB - The dynamics of pathophysiological and immunological parameters monitored in monkeys Papio hamadryas infected with the guinea pig-adapted Ebola virus strain demonstrated that this viral strain preserved its virulence for monkeys and caused the disease with characteristic features similar to those caused by non adapted Ebola virus. However, certain previously unknown patterns have been observed: (1) prolongation of the febrile period by two days; (2) extended period was characterized by stability of serum biochemical parameters; (3) marked vacuolization of the neutrophil cytoplasm; (4) appearance of juvenile lymphocytes on day 3 and by the end of the disease; and (5) a considerable increase in the spontaneous mononuclear proliferation (along with a decrease in the mitogen induced proliferation) during the terminal stage of infection. The severity of pathological coagulation was found to correlate with the activity of serum cytokines IFN-alpha and TNF-alpha: their activities increased about 250- and 100 fold, respectively. There was significant alteration in the activity of natural killer cells, that dropped by the time of animal death. PMID- 10714442 TI - Computer simulation of haemodynamic parameters changes with left ventricle assist device and mechanical ventilation. AB - Left Ventricular Assist Device is used for recovery in patients with heart failure and is supposed to increase total cardiac output, systemic arterial pressure and to decrease left atrial pressure. Aim of our computer simulation was to assess the influence of Left Ventricular Assist Device (LVAD) on chosen haemodynamic parameters in the presence of ventilatory support. The software package used for this simulation reproduces, in stationary conditions, the heart and the circulatory system in terms of pressure and volume relationships. Different circulatory sections (left and right heart, systemic and pulmonary arterial circulation, systemic and pulmonary venous circulation) are described by lumped parameter models. Mechanical properties of each section are modelled by RLC elements. The model chosen for the representation of the Starling's law of the heart for each ventricle is based on the variable elastance model. The LVAD model is inserted between the left atrium and the aorta. The contractility of the heart and systemic arterial resistance were adjusted to model pathological states. Our simulation showed that positive thoracic pressure generated by mechanical ventilation of the lungs dramatically changes left atrial and pulmonary arterial pressures and should be considered when assessing LVAD effectiveness. Pathological changes of systemic arterial resistance may have a considerable effect on these parameters, especially when LVAD is applied simultaneously with mechanical ventilation. Cardiac output, systemic arterial and right atrial pressures are less affected by changes of thoracic pressure in cases of heart pathology. PMID- 10714443 TI - Self-organizing arterial pressure pulse classification using neural networks: theoretical considerations and clinical applicability. AB - A self-organizing classification system for the arterial pressure pulse based on the ART2 (adaptive resonance theory) network was developed. The system consists of a preprocessor and an ART2 recognition network. The preprocessor removes the arterial pressure pulse servo component signals from Finapres, detects the systolic pressure points and divides the acquired signals into minimal cardiac cycles. The ART2 network input is the minimal cardiac cycle detected by the preprocessor. The classification results can be used to assist physicians in evaluating the signs of abnormal and normal autonomic control and has shown its clinical applicability for the examination of the autonomic nervous system. PMID- 10714444 TI - An improved measure for comparing diagnostic tests. AB - We present a loss based method for comparing the predictive performance of diagnostic tests. Unlike standard assessment mechanisms, like the area under the receiver-operating characteristic curve and the misclassification rate, our method takes specific advantage of any information that can be obtained about misclassification costs. We argue that not taking costs into account can lead to incorrect conclusions, and illustrate with two examples. PMID- 10714445 TI - A new way for surgical education--development and evaluation of a computer-based training module. AB - Computer-based training (CBT) programs teach the material of a specific field and at the same time offer various ways of objectively assessing the knowledge gained. The interactive use of multi-media components such as text, graphics, animation, sound, digital slide shows, and videos as well as quizzes can theoretically facilitate the learning process. The aim of this study was the development and evaluation of a CBT-program by surgeons for student training. Using SuperCard, a teaching module for Distal Radius Fracture (DRF) was developed, which contains detailed clinical information. Video clips and vivid animations combine theoretical knowledge with practical experience. Fourth-year medical students (n = 103) were tested after using the module for 90 min. Other students (n = 47) served as the control group. In a 90 min lecture, DRF was discussed. CBT gained in all evaluated criteria (distinctiveness, detailed description, presentation of materials, structure, motivation for learning, time saved learning and memory retention) 15-20% better scores than the lecture. Although 82% of the students stated that their experience with computers was limited or insufficient, 100% found the use of CBT systems useful in student teaching. Most of them suggested the use of such programs as a method of exam preparation/self study (90%) or as a supplement to a lecture (40%). Based on these evaluations, CBT modules are an appropriate future teaching and learning system that is well accepted. In conclusion, the results of this study show that CBT-programs could be a valuable supplement to medical education. In addition, further development of CBT-programs and their use as information systems for surgical residency programs at universities can be suggested. PMID- 10714446 TI - Alcohol and crime in China. AB - This study examines the pattern of alcohol involvement across violent and property crimes in China. We describe and discuss the cultural and biological differences between Chinese and Westerners concerning alcohol and the features of Chinese culture concerning violence. Drawing upon the disinhibition perspective in alcohol and crime, a specific hypothesis derived from the sociocultural context of Chinese society is that alcohol is more likely to be involved with violent crime than with property crime. Using data from a survey of inmates in China, we assess this hypothesis and the possible variables that may moderate this hypothesis. The data support the hypothesis that disinhibition is applicable to the alcohol-violence relationship in Chinese culture. However, the predicted pattern of alcohol use in violent and property crimes does not vary across different offender groups, which is inconsistent with some United States research. A tentative explanation is provided for this inconsistency. PMID- 10714447 TI - Optimal treatment for Maori with alcohol and drug-use-related problems: an investigation of cultural factors in treatment. AB - There is an increasing emphasis on taking account of the diversity of social, psychological, and cultural factors in the assessment and treatment of alcohol and drug-use-related problems. In New Zealand the increasing use of customary Maori values, beliefs, and practices in the treatment of Maori with alcohol and drug-use-related problems has also been accompanied by the adaptation and integration of Western approaches to fit contemporary Maori sociocultural needs. This paper reports on an investigation of cultural factors and cultural identity in the alcohol and drug-user treatment of a clinical sample of Maori. The essential finding was a very high endorsement of the importance of cultural factors in treatment--irrespective of age, gender, mood, level of dependence, previous admissions, cultural connectedness, or whether they were treated in a Maori dedicated program or not. A significant number believed that a sense of belonging to an Iwi (tribe), identifying as a Maori and having pride in being Maori were also important in the recovery/healing process. The findings of this study support the need to investigate the relationship between specific "cultural factors" and other clinical components of effective treatment for Maori. PMID- 10714448 TI - Cannabis users in the general Canadian population. AB - In a national survey conducted in 1994, 29.3% of all respondents reported that they had used cannabis at least once, 7.3% reported using at least once during the year of the survey, and 2.0% reported using cannabis at least once a week during the year of survey. Nonusers and those with different patterns of cannabis use could be distinguished by age; gender; the use of alcohol, tobacco, and other drugs; and involvement with drug users. Frequent cannabis use at the time of the survey was associated with "heavy" drinking and drinking problems, drinking and driving, driving within 2 hours of using cannabis, and the use of other drugs, especially among young males. The association between regular cannabis use, "heavy" drinking, and other risk behaviors poses challenges to prevention and harm-reduction initiatives. PMID- 10714449 TI - "Drug dependence" and death: survival analysis of the Baltimore ECA sample from 1981 to 1995. AB - AIMS: Illicit drug use and dependence often are associated with premature death, but available evidence comes mainly from clinical samples. The present paper examines drug-related mortality experience over 14 years in a United States community sample. PARTICIPANTS: Following probability sampling, 3,481 adult community household residents were recruited for the 1981 NIMH Baltimore Epidemiologic Catchment Area survey. Follow-up occurred in 1993-1996. METHODS: Survival analyses were used to estimate median age at death and relative risk of dying in relation to drug use and dependence as assessed in 1981 using the Diagnostic Interview Schedule (DIS). FINDINGS: Cases with DIS "drug dependence" were more likely to have died and to have a younger median age at death (p < .05), with and without statistical adjustment for confounding variables. Higher levels of drug involvement also were associated with increased age-adjusted mortality. CONCLUSIONS: The evidence favors the hypothesis that DIS-elicited "drug dependence," as well as subthreshold drug use, help to account for premature death in this community sample. PMID- 10714450 TI - Economics as a factor in models of behavioral motivation and change. AB - This note first presents a summary of four main behavioral models that are used to explain behavioral motivation and change. Three models are based on psychosocial theory. They are: 1) the Theory of Reasoned Action, 2) the Theory of Planned Behavior, and 3) the Theory of Stages-of-Change. The fourth model is based on economic theory and is known as the Rational Addiction Model. Each model is analyzed for its strengths and weaknesses. The note concludes by arguing for the usefulness of integrating the economic and the psychosocial models to study drug use. Specific examples and suggestions are presented. PMID- 10714451 TI - Nurses' attitudes toward substance misusers. I. Surveys. AB - Studies of nurses' attitude toward substance misusers, spanning three decades, are reviewed. Survey findings reflect greater acceptance of alcohol and drug misusers in recent years. However, significant minorities of nurses continue to regard substance users as immoral, characterologically defective, and unlikely to recover. Implications of these attitudes for treatment of substance misusers are discussed. PMID- 10714452 TI - Exploring the additive effects of drug misuse treatment and Twelve-Step involvement: does Twelve-Step ideology matter? AB - Previous research revealed an additive effect of recovery activities in that those who attended Twelve-Step meetings on a weekly basis during and after outpatient drug-user treatment had higher rates of abstinence compared to those who participated in either treatment or Twelve-Step programs alone. The current investigation extends the previous research by examining the possible effects of Twelve-Step ideology on participation in Twelve-Step programs and abstinence from drug use. The findings from this treatment outcomes study indicate that the acceptance of Twelve-Step ideology, particularly strong agreement with the need for frequent, lifelong attendance at Twelve-Step meetings, and the need to surrender to a "higher power" are significant predictors of weekly or more frequent attendance at Twelve-Step meetings independent from other potentially mediating variables. Twelve-Step ideology, specifically the notion that controlled or nonproblematic drug use is not possible, predicted abstinence independent from Twelve-Step participation and other potentially mediating variables. These findings often a number of implications concerning group process and recovery from drug misuse which are addressed in the Discussion section under the following topics: 1) spirituality and group cohesion, 2) spiritual transcendence, social transcendence, and recovery; 3) spirituality and the obstruction of recovery; 4) Twelve-Step ideology and learning; 5) perceived control of drug use, self-efficacy theory, and recovery; and 7) perceived control of drug use and optimistic illusions. Directions for future research are discussed. PMID- 10714453 TI - Methods for estimating the number of "hard-core" drug users. AB - This article reviews methods used to estimate the number of "hard-core" drug users. Synthetic estimation methods include the population projection method, which extrapolates from areas where prevalence is known to other areas, and the principal component method, which uses relationships observed among multiple indicators to obtain a single indicator of drug use. Small area estimations project risk estimates developed from small area surveys to larger areas, and capture-recapture methods provide reliable estimates only if the underlying assumptions are met. In making estimates, researchers should understand the limitations of the data and changes in drug use patterns. The challenge for future research is to refine each method and then to combine the strengths of each to produce the best estimates possible. PMID- 10714454 TI - Effects of a brief alcohol preventive intervention for youth attending school sports physical examinations. AB - This pilot study examined the feasibility and efficacy of a brief alcohol misuse preventive intervention for 178 7th-9th grade junior high school students attending sports physical examinations at three schools during the Summer of 1997. At 6-month posttest, fewer suburban intervention youth intended to use alcohol during the next 6 months (chi2 = 7.01, 1 df, p = .01), and fewer rural intervention youth used alcohol during the past 30 days (chi2 = 4.65, 1 df, p = .04), compared to control youth. When suburban and rural school samples were collapsed, intervention youth had significantly lower alcohol use on three of four measures than control subjects (p's < .05). PMID- 10714455 TI - The distribution and epidemiology of bovine trypanosomosis in Malawi. AB - A survey to update the distribution and clarify the epidemiology of bovine trypanosomosis in Malawi was conducted between 1995-97. Use was made of parasitological and serological (anti-trypanosomal antibody-detection Enzyme Linked Immunosorbent Assay (ELISA) diagnostic methods. Trypanosomal infections were detected in cattle sampled adjacent to known tsetse foci. The distribution of cattle with anti-trypanosomal antibodies indicated that the distribution of bovine trypanosomosis was more widespread than the parasitological prevalence data suggested. This is attributed to the seasonal movement of tsetse (mainly Glossina morsitans morsitans and G. pallidipes) from its prime habitat and the presence of localized foci of G. brevipalpis. The odour-baited, insecticide treated, target barriers along the edge of Kasungu National Park and the Nkhotakota Game Reserve appeared to be effective in preventing tsetse from moving outside the game areas and contacting cattle. The anti-trypanosomal antibody detection ELISA proved to be a useful tool in establishing the distribution of bovine trypanosomosis. Moreover, the distribution and prevalence of cattle with anti-trypanosomal antibodies was instrumental in clarifying the epidemiology of bovine trypanosomosis in Malawi. The anti-trypanosomal antibody-detection ELISA had high sensitivity in detecting Trypanosoma congolense infections. In parasitologically negative animals, the average packed cell volume was higher in those that had no anti-trypanosomal antibodies. The packed cell volume decreased with increasing antibody titre. PMID- 10714456 TI - Effect of experimental fasciolosis on antipyrine metabolism and clearance in water buffaloes. AB - The effect of chronic Fasciola hepatica infection on the metabolism of antipyrine, a marker of microsomal oxidative metabolism, was investigated in male water buffaloes dosed daily with 60 F. hepatica metacercariae over 20 days. The plasma elimination half-life of antipyrine was significantly elevated by 23% at 11 weeks postinfection (p.i.) but did not significantly differ from the control period at 20 weeks p.i. The systemic clearance of antipyrine decreased by 48% at 11 weeks p.i. and then returned to normal. The renal clearance for each of the main antipyrine metabolites decreased at 11 weeks p.i. (hydroxymethylantipyrine (HMA), -42%; norantipyrine (NORA), -58%; and 4-hydroxyantipyrine (OHA), -70%) and did not significantly differ from the control period at 20 weeks p.i. These findings indicate that experimental subclinical fasciolosis leads to altered antipyrine kinetics and to an inhibition of the different antipyrine metabolic pathways in water buffaloes. PMID- 10714457 TI - A strategic dosing scheme for the control of fasciolosis in cattle and sheep in Ireland. AB - This study examined the effectiveness of a strategic dosing scheme in lowering the incidence of fasciolosis on a mixed dry-stock farm and in maintaining the reduced incidence following a reduction in dosing intensity. Two neighbouring farms with a history of chronic fluke disease were selected, the strategic dosing scheme being implemented on one (the trial farm) while the other (the control farm) continued to treat according to its normal practice. The strategic dosing scheme was designed to suppress the faecal egg output of Fasciola hepatica at critical times of the year in order to limit infection of the intermediate host snail population and thus reduce the subsequent contamination of the pasture with metacercariae. On the trial farm cattle and sheep were treated three times per year for the first 2 years at approximately 8 week intervals, starting in March of each year. A fourth treatment was given when the cattle were housed and out wintered sheep received an additional treatment in January. In Years 3 and 4 the dosing intensity was reduced. By the end of Year 2, data from faecal egg counts, tracer-sheep fluke burdens and snail infection levels indicated that the treatment strategy had succeeded in suppressing the fluke population and eliminating the occurrence of clinical fasciolosis. The decrease in dosing intensity in Years 3 and 4 maintained both stock and snail infections at low levels and there was no re-emergence of the disease. PMID- 10714458 TI - A model for study of lungworm (Dictyocaulus sp.) and gastrointestinal nematode infection in young red deer (Cervus elaphus). AB - A model of sub-clinical parasitism in young red deer, using concurrent trickle infections of lungworm (Dictyocaulus sp.) and mixed gastro-intestinal (GI) nematodes of deer-origin was evaluated. 20 parasite-free deer calves were artificially reared indoors from 4 days of age. A further five calves were naturally reared on pasture with their dams, treated with anthelmintic and brought indoors at 3-4 months. At 4-4.5 months of age they were individually housed and allocated to five groups (n=5). Groups were dosed 3 x per week, for 9 weeks with 0, 100 and 500, 200 and 1000 (2 groups), 400 and 2000 infective larvae of lungworm and mixed GI nematodes, respectively, cultured from deer faeces. Liveweight and voluntary feed intake measurements and faecal and blood samples were taken weekly. In the fourth week following cessation of trickle infection, deer were euthanased and lung and GI nematodes recovered. Both lungworm and GI nematode infections became patent at Week 4 of infection. Maximum group arithmetic mean faecal egg counts were 100-190 epg. Maximum group arithmetic mean faecal lungworm larval counts were 58-123 lpg. Group arithmetic mean nematode counts at slaughter ranged from 439-806 for GI nematodes and 31-73 for lungworm, respectively. Despite low nematode counts, reduced liveweight gain, voluntary feed intake and serum albumin concentration, elevated serum pepsinogen, gastrin and globulin concentrations and elevated peripheral eosinophil counts and slight haemoconcentration, but no clinical signs, were observed. The reduction in liveweight gain was related to the reduction in voluntary feed intake (r2=0.83; p<0.088). Naturally-reared deer had similar liveweight gains, voluntary feed intake and nematode counts to artificially-reared deer. Thus, methods of infection to produce concurrent sub-clinical lungworm and GI nematode burdens for study of sub-clinical parasitism in young deer have been defined. PMID- 10714459 TI - Induction of protective immunity to Dictyocaulus viviparus in calves while under treatment with endectocides. AB - Three groups of five parasite-naive calves were used. The treatments were: (a) Group 1 calves were weighed on Day 0 and injected with doramectin at 200 microg/kg. From Day 1 to 19 they were dosed orally with 2000 infective larvae of Dictyocaulus viviparus. On Day 28 they were again injected with doramectin, and infected with D. viviparus larvae from Days 33 to 41. They were then left untreated until Day 81 when they were infected with 20 infective larvae of D. viviparus per kg body weight. They were killed on Day 110 and lungworms were counted; (b) Group 2 calves were immunised with oral lungworm vaccine on Days 0 and 28, and infected and slaughtered as Group 1 on Days 81 and 110, respectively; (c) Group 3 calves acted as infection controls. Blood samples were taken at Days 0, 21, 49, 77 and 110 for antibody tests to D. viviparus. At autopsy there were no significant differences between the number of lungworms from Groups 1 and 2 (Means 17.4 and 31.3, respectively); Group 1 had significantly less value than Group 3 (Mean 228) (p < 0.05). Increased antibody titres to the larval sheath of the infective larvae were observed from Groups 1 and 2, showing that the larvae in Group 1 had penetrated the intestine before being killed by the circulating anthelmintic. This experiment shows that if calves are exposed to infective larvae while under systemic endectocide cover, an immune reaction is stimulated. PMID- 10714460 TI - Screening for infection of Trichinella in red fox (Vulpes vulpes) in Denmark. AB - A total of 6141 foxes (Vulpes vulpes) were examined for infection with Trichinella. The foxes were killed in Denmark during the hunting season 1995-1996 and 1997-1998; 3133 and 3008, respectively. Foxes included in the investigation came from throughout the country with the exception of the island of Bornholm. The right foreleg from each fox was submitted for investigation. The legs were stored at -20 degrees C for 3-10 months prior to examination. Following thawing, muscle tissue (10 g) from each leg was examined by trichinoscopy and by a pepsin HCl digestion technique. In 1995-1996, three foxes were found positive corresponding to a prevalence of 0.001. Each of the infected foxes harboured an extremely low infection, i.e. about one larva per 10 g muscle tissue. It was not possible to obtain sufficient larval material for species identification. All three foxes were shot in the vicinity of a small village in the north-western part of Denmark. In 1997-1998 no Trichinella cases were found. The results, compared with previous studies, indicate that the prevalence of infection of Trichinella sp. among wild living foxes in Denmark is very low. This is further supported by the fact, that no infection of Trichinella sp. has been found in slaughtered pigs in Denmark for more than 65 years, which suggests that the infection pressure is very low. Considering the facts above we conclude that the risk of Trichinella infections is negligible in intensive indoor pig production units in Denmark whereas high local prevalence of Trichinella infections in the wildlife might constitute a serious risk for the expanding outdoor pig production. PMID- 10714461 TI - Parenterally administered gastrointestinal nematode infective larvae viable after more than 15 years in liquid nitrogen. AB - After cryopreservation for 13.3-15.8 years, the viability of the infective larvae (L3) of Trichostrongylus axei, T. colubriformis, Oesophagostomum columbianum, Haemonchus contortus, Ostertagia circumcincta, T. falculatus, Nematodirus spathiger, Chabertia ovina and Dictyocaulus filaria was assessed in sheep, by being deposited at their predilection sites. D. filaria was, however, an exception, in that the L3 were injected into the jugular vein. The mean development of all the species was 22.8%, but if three species (O. columbianum, C. ovina and D. filaria), that developed poorly are disregarded, then the mean development was 33.4%, similar to previous tests after shorter periods of cryopreservation. The L3 of some of the species appeared sluggish when examined 10-15 min after being thawed, and in the case of H. contortus practically all the larvae of the original batch tested in the previous trials of the series appeared dead when thawed for use in the present trial, and were replaced by another batch of L3 of the same species. When re-examined after about 8 h, however, a high percentage of the L3 of the original batch appeared to have become revitalised, and their viability was tested in a trial reported elsewhere. The intestinal cells of the majority of the L3 of N. spathiger, O. circumcincta and C. ovina were vesiculated when they were thawed. Nevertheless, the degree of development of the former two species was of the highest in the trial, and it can be concluded that this phenomenon does not necessarily impede the viability of larvae. PMID- 10714462 TI - Vaccination of calves with Haemonchus placei intestinal homogenate. AB - The ability of adult Haemonchus placei intestinal homogenate to confer protection against homologous challenge infection was evaluated. Calves were immunized twice with 100 microg H. placei intestinal protein in 5% dextran-sulfate/PBS (vaccinates) or PBS alone (controls) and were challenged with approximately 3300 infective H. placei larvae. There was no significant difference between groups in the total number of nematodes recovered but significantly fewer (p < 0.001) adult females were recovered from vaccinates. The proportion of fourth-stage larvae in vaccinates was significantly greater (p < or = 0.05) than in controls. Lengths of adult male and female nematodes were significantly shorter (p < 0.001) in vaccinated calves, and the numbers of eggs present in the uteri of female nematodes from vaccinates were significantly decreased (p < 0.001). Counts of nematode eggs per gram of feces (EPG) of vaccinates were significantly less than that for controls on Days 29-49 post-challenge (p < or = 0.05). Vaccinates had significant increases in serum IgG1 and IgG2 log(10) titers (p < or = 0.05) but not in serum IgM. EPG, numbers of females, and size of males and females were negatively correlated with increased mean post-challenge IgG1 and IgG2 titers. Reduction in binding of periodate-treated gut homogenate by immune serum indicated a carbohydrate specific component in the immune response. PMID- 10714463 TI - In vitro insecticidal effects of fipronil and beta-cyfluthrin on larvae of the blowfly Lucilia sericata. AB - The insecticidal effects of the phenylpyrazole, fipronil, and a pyrethroid, beta cyfluthrin, on larvae of the blowfly Lucilia sericata were determined in laboratory assays. When first stage larvae of L. sericata were reared on homogenized pig liver which had been treated with known amounts of test compounds, both fipronil and beta-cyfluthrin induced significant levels of mortality compared to acetone and water controls. However, fipronil was approximately 10 times more toxic than beta-cyfluthrin to L. sericata larvae following ingestion. Beta-cyfluthrin had little effect on mortality until concentrations of approximately 0.5 ppm were reached. In contrast, fipronil effected L. sericata mortality at a concentration of 0.05 ppm and 100% mortality was reached by 0.5 ppm. The lethal concentration (LC50) value for beta-cyfluthrin was 1.56 ppm as compared to 0.14 ppm for fipronil. Following contact of first and third stage larvae with cloth impregnated with known amounts of test compound, the mortality profiles of fipronil and beta-cyfluthrin were similar. At short contact times, the LC50 values for fipronil were lower than those for beta cyfluthrin. However, at the highest contact time evaluated for the first stage larvae, 300 s, there was a reversal in this trend. The results suggest that the phenylpyrazole fipronil may represent a new potential insecticide for development against blowfly strike of sheep. PMID- 10714464 TI - Prevalence and larval burden of oestrus ovis (Linne 1761) in sheep and goats in northern mediterranean region of France. AB - A slaughterhouse survey to determine prevalence and larval burden of Oestrus ovis larvae in sheep and goats was performed monthly during one year in Pezenas, South of France, northern mediterranean region. A total of 1303 sheep and goat heads were selected at random. O. ovis larvae were found in 274 sheep out of 631 (43.4%), and the prevalence rate varied from 14.3% in February to 65% in October. The mean number of larvae in infected sheep heads was 10.86 with 9.24 L1, 0.91 L2 and 0.71 L3. One hundred and ninety-one goats out of 672 were infected (28.4%), and the prevalence rate varied from 6.25% in September to 47.1% in April. In infected goat heads, the mean parasitic burden was 5.35 with 4.04 L1, 0.73 L2 and 0.58 L3. These results confirm worldwide observations indicating that the prevalence and the parasitic burdens are less in goats than in sheep. PMID- 10714465 TI - Evaluation of tickGARD(PLUS), a novel vaccine against Boophilus microplus, in lactating Holstein-Friesian cows. AB - The effects of vaccination with the Bm 86 vaccine TickGARD(PLUS) against infestation with cattle tick (Boophilus microplus) and of holding cattle on a feedpad until 09:00 hours after the morning milking was tested on 40 mid lactation Holstein cattle using a factorial design. Vaccination resulted in a 56% reduction in tick numbers in the field over one generation, and a 72% reduction in laboratory measures of the reproductive efficiency of ticks. The liveweight gain of vaccinated cattle over 27 weeks was 18.6 kg higher than that of controls, and vaccinated cattle tended to have lower somatic cell count in milk (SCC). There were no other significant differences in measures of production. Cattle kept on the feedpad after the morning milking carried 26% more ticks than those returned immediately to their paddocks. PMID- 10714466 TI - Efficacy of moxidectin equine oral gel against endoscopically-confirmed Gasterophilus nasalis and Gasterophilus intestinalis (Diptera: Oestridae) infections in horses. AB - A 3 m, video gastroscope was used to screen 47 horses suspected of being naturally infected with equine bot larvae. 17 of 47 (36.2%) candidate horses harbored Gasterophilus nasalis larvae in the proximal duodenum and 46 of 47 (97.9%) had G. intestinalis larvae in the stomach. All horses infected with G. nasalis had concurrent infections with G. intestinalis. 14 horses with dual infections were allocated randomly to two treatment groups. Seven horses in Group 1 received 2% moxidectin oral gel once at a dosage of 0.4 mg/kg bodyweight (BW), and seven horses in Group 2 were untreated controls. 14 days after treatment, all horses were necropsied and the stomach and proximal duodenum harvested from each. Bot larvae were recovered, identified to species and instar, and counted. At the label dosage, moxidectin oral gel was 100 and 97.6% effective (P < 0.05) against third-instar G. nasalis and G. intestinalis, respectively. In addition to demonstrating the boticidal efficacy of moxidectin, this trial illustrated that gastroscopy/duodenoscopy is a feasible method for confirming infections with different species of bot larvae in the horse. PMID- 10714467 TI - Efficacy of an ivermectin controlled-release capsule against some rarer nematode parasites of sheep. AB - A controlled trial was conducted to evaluate the efficacy of the intraruminal ivermectin controlled-release capsule (CRC) (IVOMEC Maximizer CR Capsule for Sheep, Merial Ltd.) against induced incoming third-stage larvae and established adult infections with some rarer gastrointestinal nematode parasites of sheep. Twenty-one worm-free lambs were allocated by restricted randomisation based on body weight within sex to one of the following treatments: unmedicated control, ivermectin CRC given on Day 0 prior to induced infection, and ivermectin CRC given on Day 70 after establishment of induced infection. The ivermectin CRC delivers ivermectin at a minimum dose rate of 20 microg/kg/day for 100 days. Infections were induced by daily administration of third-stage larvae for five consecutive days. Nematodes were counted on Day 84, 14 days after treatment of established infection. The treatment with the ivermectin CRC prevented the establishment of Ostertagia leptospicularis, O. ostertagi, Bunostomum trigonocephalum, Cooperia oncophora, C. punctata, C. surnabada, Nematodirus helvetianus, N. roscidus and Strongyloides papillosus by >99% as compared with the untreated controls (p < 0.01). The administration of the ivermectin CRC reduced established adult infections of O. ostertagi, B. trigonocephalum, C. oncophora, C. punctata, C. surnabada, N. roscidus and S. papillosus by >99% (p < 0.01), and reduced established adult infections of O. leptospicularis and N. helvetianus by 96.5 and 98.4% (p < 0.01), respectively. PMID- 10714468 TI - The effect of doramectin, moxidectin and netobimin against natural infections of Syphacia muris in rats. AB - In this study, the effect of doramectin, moxidectin and netobimin was investigated in naturally infected Syphacia muris in rats. The natural infection was determined by the use of cellophane tape method on the perianal region and by the technique of centrifugal flotation of feces. The infected rats were divided into three treated and one control group (N = 10). Doramectin and moxidectin at the dose of 0.2 mg/kg per day and netobimin at the dose of 7.5 mg/kg per day were given in the diet for 4 days. Cellophane tape preparations were performed in all groups on 4th and 7th day after the last treatment. The rats of treated groups were necropsied on 7th day after the last treatment together with that of control group. While doramectin and netobimin were highly effective against S. muris, moxidectin was not found to be effective for eradication of S. muris. PMID- 10714469 TI - Evidence of Onchocerca fasciata (Filarioidea: Onchocercidae) in camels (Camelus dromedarius): I-prevalence, nodular lesions appearance and parasite morphology. AB - Examination of 3376 imported and 200 local Egyptian camels was carried out during the period extending between September 1997 and August, 1998. These animals were carefully examined for the presence of Onchocerca fasciata nodules and subsequently for O. fasciata adult parasites. Results of this study revealed that imported camels had the higher infection rate (2.75%), while those of local origin showed no palpable or detected Onchocerca nodules. Distribution of these nodules was mainly on the two sides of abdomen, hind limbs (concentrated in thigh region) and fore limbs particularly on the shoulders and nuchal ligament. This distribution varied according to the degree of infection. Searches for microfilariae were also performed using either blood samples, or in the subcutis and fascial sheath near or around the detected nodules. PMID- 10714470 TI - Effects of sarcoptic mange on serum proteins and immunoglobulin G levels in chamois (Rupicapra pyrenaica) and Spanish ibex (Capra pyrenaica). AB - Three groups of chamois (Rupicapra pyrenaica) and three groups of Spanish ibex (Capra pyrenaica) were established to study the effects of sarcoptic mange on serum proteins and immunoglobulin G (IgG) levels. The first group of chamois consisted of 22 healthy Pyrenean chamois (R. pyrenaica pyrenaica) from a non infested area, the second group consisted of 20 healthy Cantabrian chamois (R. p. parva) from an area where sarcoptic mange has been reported since 1994 and the third group consisted of 16 Cantabrian chamois from the same area but naturally infested by Sarcoptes scabiei. The first group of Spanish ibex was 39 healthy animals from a sarcoptic mange non-infested area, the second group was 23 healthy animals from a sarcoptic mange infested area and the third group consisted of 20 animals from the same area but naturally infested with the parasite. Blood samples were taken after killing the animals as part of hunting programmes. Values for total proteins, gamma-globulin and IgG were higher in infested and healthy chamois from the infested area compared to healthy chamois from the non infested area, and IgG levels were higher in infested chamois compared to healthy exposed chamois. Values for alpha2-globulin were higher in healthy Cantabrian chamois. In Spanish ibex, albumin, alpha2-globulin and IgG levels were lower in the healthy Spanish ibex from the non-infested area than in healthy animals from an infested area. The differences found in the chamois were indicative of the establishment of a humoral antibody response in the animals in contact with the disease. As the IgG levels were not significantly different between healthy and infested Spanish ibex from the same area, a different pattern of chronic infection with humoral response to the disease was suggested. PMID- 10714471 TI - Further evidence on the internal life cycle of Przhevalskiana silenus (Diptera, Oestridae). AB - Knowledge of the internal life cycle of goat warble fly infestation is scarce despite ample data available on the aetiology, epidemiology, immunodiagnosis and treatment of such infestations. This study was carried out at the slaughterhouse of Rossano Calabro (Cosenza, southern Italy) on 154 animals from 10 months to 6 years of age from May 1997 to June 1998. 1206 Przhevalskiana silenus larvae were collected during the trial period from the subcutaneous tissue of the slaughtered animals. The larval stage average size ranged from 4.7 mm, for first instar larvae (May), to 16.6 mm, for third instar larvae (February), in the first cycle of infestation. No larvae were found in March-April, coinciding with the pupation period. Small first instar larvae were found at the beginning of the second cycle of infestation (May-June). Necroscopic examinations were also carried out on internal organs and no larvae were found. The results pointed out that the internal life cycle of P. silenus is exclusively subcutaneous and there is no internal migration of the larvae. PMID- 10714472 TI - Susceptibility to two pyrethroids in Boophilus microplus (Acari: Ixodidae) populations of northwest Argentina. Preliminary results. AB - Cattle are treated 6-12 times yearly to control Boophilus microplus ticks in the east zone of the Argentinean infested region, while 1-4 treatments are applied for tick control in the west zone. In the 1970s resistance to organo-phosphate acaricides was found in the east zone, but not in the west zone. However, a shift to synthetic pyrethroids (SP) was made through all infested regions. Currently, indications of resistance to SP in the east zone, but not in the west zone, are provoking to a switch to formamidine acaricides. During 1998 a total of 147 B. microplus engorged females were collected from 20 beef cattle ranches from the west zone of the Argentinean infested region. Individual progenies of these ticks were tested ('larval packet test') with cypermethrin and deltamethrin, and their LC 50 and LC 90 were compared to those estimated for the Milagro susceptible strain. No evidence of resistance to these SP was found. Due to sampling restraints the results are presented as preliminary. Nevertheless, a shift away from use of SP for control of B. microplus in the west zone appears to be unjustified and should be independent of the resistance circumstances observed in the east zone of the Argentinean tick infested region. PMID- 10714473 TI - Improving patient compliance with asthma therapy. AB - Patients fail to comply with asthma medication for a variety of reasons. These range from physical inability to use an inhaler, through simple forgetfulness, to a conscious decision not to use medication as prescribed due to internal or cultural health beliefs or socioeconomic factors. In some patients, poor self care because of deep-rooted psychological factors (i.e. factors of which patients have only limited awareness) can affect compliance. Poor doctor-patient communication can be the cause in many other individuals. Thus, there is no single solution that will improve compliance in all patients. Simplifying the regimen or providing memory aids will be sufficient for some patients, while education or psychological counselling will be more appropriate for others. Doctors can also use a range of communication skills to improve the way in which they present information, motivate patients and reinforce progress. These approaches, plus respect for patients' health beliefs and involving them in treatment decisions, can help foster an atmosphere of mutual responsibility and concordance over medicine taking. PMID- 10714474 TI - Regulated exocytosis in immune function: are SNARE-proteins involved? AB - Inflammation is an important feature in the pathogenesis of most chronic lung diseases. It is characterized by tissue infiltration with various inflammatory cells, including eosinophils, mast cells, basophils, macrophages, neutrophils, T- and B-lymphocytes and dendritic cells (1). In the tissue granulocytes release their toxic granule proteins after being stimulated by soluble mediators released by other inflammatory cells (2). Therefore, it is important to characterize the intracellular mechanisms regulating the transport of the granule contents in inflammatory cells. Intracellular vesicle-traffic in mammalian cells is mediated by transport vesicles that emerge from donor compartments and are specifically targeted to acceptor compartments where they deliver their contents after membrane fusion (3). This traffic leads to three types of fusion: vesicle intracellular membranes, vesicle-vesicle or vesicle-plasma membrane. The process leading to fusion of vesicle-plasma membrane is called exocytosis, and it delivers proteins to the cell surface (receptors e.g. CD11b, CD18) and exports soluble molecules (mediators e.g. ECP) from the cell. A number of key proteins involved in regulated exocytosis have been identified from inflammatory cells. This review is a brief summary of these proteins and it includes recent results from studies on regulated exocytosis in inflammatory cells. PMID- 10714475 TI - Short-course moxifloxacin therapy for treatment of acute bacterial exacerbations of chronic bronchitis. The Bronchitis Study Group. AB - Chronic bronchitis is common among adults and infectious exacerbations contribute considerably to morbidity and mortality. We aimed to compare the safety and efficacy of moxifloxacin to clarithromycin for the treatment of patients with acute bacterial exacerbations of chronic bronchitis (ABECB) using a prospective, randomized, double-blind, parallel group trial. Between November 21, 1996 and April 7, 1998, 936 patients with acute exacerbations of chronic bronchitis (AECB) were enrolled at 56 centers across the United States of which 491 (52%) had ABECB (i.e. pretherapy pathogen). Patients were randomized to either oral moxifloxacin 400 mg administer once daily, for either 5 or 10 days, or clarithromycin 500 mg bid for 10 days. For the purpose of study blinding, the patients taking moxifloxacin received placebo to maintain uniform dosing. The main outcome measures were bacteriological response at the end of therapy (post-therapy days 0 6) and follow-up (7-17 days post-therapy) visits, as well as overall clinical response, clinical response at the end of therapy and clinical response at follow up. Two patient populations were analyzed: efficacy-valid (i.e., those with a pretherapy pathogen) and intent-to-treat (i.e., all subjects that took drug). In 420 efficacy valid patients with a pretherapy organism, overall clinical resolution was 89% for 5 days moxifloxacin vs. 91% for 10 day moxifloxacin vs. 91% for 10 day clarithromycin. Bacteriological eradication rates at the end of therapy were 94% and 95% for 5-day moxifloxacin and 10-day moxifloxacin, respectively, and 91% for the clarithromycin group. Eradication rates at follow up were 89% and 91% for 5-day moxifloxacin and 10-day moxifloxacin respectively, and 85% for the clarithromycin group. Among 926 intent-to-treat patients (312 5 day moxifloxacin, 302 10-day moxifloxacin and 312 clarithromycin), drug-related events were reported for 26%, 30% and 35%, respectively. Moxifloxacin 400 mg once daily, as a 5 or 10 day regimen, was found to be clinically and bacteriologically equivalent to 10 day clarithromycin for the treatment of ABECB. Given its favorable safety and tolerability profile, moxifloxacin administered once daily for 5 days may be as effective and a more convenient treatment than a standard course of clarithromycin for patients with ABECB. PMID- 10714476 TI - Importance of adjusting carbon monoxide diffusing capacity (DLCO) and carbon monoxide transfer coefficient (KCO) for alveolar volume. AB - The volume dependence of single breath carbon monoxide diffusing capacity (DLCO) and carbon monoxide transfer coefficient (KCO) was determined in 24 healthy subjects. The change in DLCO [fraction of DLCO measured at total lung capacity (TLC)] to change in alveolar volume [fraction of alveolar volume (VA) at TLC] closely fitted a simple linear regression and matched a theoretical model. As VA decreased, DLCO fell linearly and KCO increased as expected from the relation of DLCO to VA. The equations for adjustment of predicted DLCO and KCO for alveolar volume are: DLCO/DL COtlc = 0.58 + 0.42 VA/VAtlc, KCO/KCOtlc = 0.42 + 0.58/(VA/VAtlc). DLCO and KCO were evaluated in 2313 patients. Subgroups of patients with asthma, emphysema, extrapulmonary lung disease, interstitial lung disease and lung resection were identified. Unadjusted DLCO and KCO percent predicted values showed large differences and much variability, so can be misleading. As expected, KCO and DLCO percent predicted values adjusted for alveolar volume were nearly identical. Subgroups have characteristic patterns of VA and unadjusted and adjusted DLCO and KCO. Changes in DLCO and KCO with alveolar volume are relevant for accurate interpretation of diffusion in patients with low lung volumes. Adjusting predicted DLCO and KCO for alveolar volume provides a better assessment of lung function. PMID- 10714477 TI - Emphysematous lesions and lung function in healthy smokers 60 years of age. AB - We aimed to study the occurrence of emphysematous lesions in symptom free smoking men of about 60 years of age and in a matching group of never-smoking men and the relationship between pulmonary changes at high resolution computed tomography (HRCT) and lung function tests. Our investigation included 57 smoking and 32 never-smoking healthy men from a randomized epidemiological study. HRCT was performed at full inspiration with a 1.5 mm slice thickness and a 3 cm inter slice distance. Evaluation was made by two radiologists unaware of smoking history. Emphysematous lesions were scored visually. Pulmonary function tests were performed including spirometry and diffusion capacity test (DLCO). Emphysematous changes were demonstrated in 25 of 57 smokers but in only one never smoker. DLCO/VA was the most sensitive test for early emphysematous lesions. It also correlated with radiographical scoring. Emphysematous lesions were evident in 44% of the healthy symptom free smokers. HRCT may reveal early emphysematous lesions in smokers before clinical symptoms have developed. PMID- 10714478 TI - Antigen-induced airway inflammation and hyper-responsiveness does not enhance airway responses to a subsequent antigen challenge in rats. AB - Brown-Norway (BN) rats develop airway hyper-responsiveness and lung eosinophilia 18-24 h after ovalbumin (OA) challenge. We hypothesized therefore that allergen induced airway inflammation would further enhance airway responses to a subsequent antigen challenge. Animals were sensitized to both OA and bovine serum albumin (BSA) and, 14 days later, challenged by aerosols with both antigens 24 h apart. Measurements of pulmonary resistance (RL) were made for 8 h after the second antigen challenge and bronchoalveolar lavage (BAL) was performed. Animals were divided into three groups and received two challenges as follows: saline-BSA (n=9), OA-saline (n=8) and OA-BSA (n=10). Sensitization was confirmed by measurements of specific OA-IgE and BSA-IgE. Early responses [determined as the highest value of RL within the first 30 min after the challenge] were absent in all study groups. The late responses [determined from the area under the RL versus time curve from 120 to 480 min after the challenge] were significantly greater in animals challenged with BSA (15.16+/-3.86) compared to saline (3.76+/ 4.09; P<0.05). However previous exposure to OA did not further increase the late response in animals subsequently challenged with BSA (20.11+/-3.67) despite enhanced airway responsiveness to LTD4 at this time point. BAL eosinophils and lymphocytes were significantly increased following BSA challenge in previously OA challenged animals, compared to numbers retrieved from animals previously exposed to saline (P<0.05). These data indicate that previous exposure to OA did not further increase the LR to a second antigen challenge despite substantial increases in airway inflammatory cells and airway hyper-responsiveness to LTD4. PMID- 10714479 TI - The relative bioavailability of salbutamol to the lung using urinary excretion following inhalation from a novel dry powder inhaler: the effect of inhalation rate and formulation. AB - Each dry powder inhaler has a different resistance so that a respirable dose can be generated from the formulation by the patient's inspiratory effort. It is important to recognize that this effect is achievable. The inspiratory flow characteristics of asthmatics inhaling through a Clickhaler were determined. In a separate study 10 volunteers inhaled separate 2 x 100 microg salbutamol doses from a Clickhaler (ML Laboratories plc U.K.). Two different formulations (one with a high and one with a low respirable fraction) were each inhaled using an inspiratory flow of 30 and 60 l min(-1). A urine sample was collected 30 min post inhalation and then pooled for the next 24 h. The mean (SD) inhalation rate of 24 asthmatics when they inhaled from a Clickhaler was 37.3 (14.6) l min(-1). The mean (SD) urinary salbutamol excretion during the first 30 min, by the volunteers, using the high respirable dose formulation at 30 and 60 l min(-1) was 5.59 (1.87) and 4.62 (1.49) microg respectively (P<0.01). Similar values using the low respirable dose formulation were 4.84 (1.58) and 3.86 (2.14) microg. There was no significant difference between the amounts excreted in the 24 h post dose. The different 30-min urinary excretions following inhalation of the two formulations suggest a link between the relative bioavailability of salbutamol to the lung and the respirable dose, and that a slow inhalation rate should be used when using a Clickhaler. The patient study shows that this rate is achieved by most asthmatics without further training. PMID- 10714481 TI - Preferential recruitment of phagocytes into the lung of patients with advanced acquired immunodeficiency syndrome and tuberculosis. AB - Limited data are available on the cellular and immunocytological characteristics of bronchoalveolar lavage (BAL) fluid in individuals infected with the human immunodeficiency virus (HIV) and pulmonary tuberculosis (TB). The immune host response against tuberculosis in early HIV-infection may differ from that in later stages of HIV disease, as is strongly suggested by different clinical and radiographic patterns. We studied the cellular elements in the lungs of 15 HIV infected patients with advanced immunosuppression and pulmonary tuberculosis (TB/AIDS). The findings were compared with data from four other groups: 1) 15 HIV seronegative patients with pulmonary TB; 2) 12 HIV-seropositive TB patients without previous AIDS-defining illnesses and with CD4+ >200 cells mm(-3); 3) five AIDS patients without pulmonary lesions; and 4) five healthy controls. BAL fluid and differential cell counts, as well as lymphocyte subsets, were determined. Despite a low CD4/CD8 ratio, the TB/AIDS group had a higher absolute number of CD8+ lymphocytes in the BAL fluid than the other groups. Alveolar macrophages and neutrophils were significantly increased in TB/AIDS patients compared to control groups. The number of eosinophils was increased in TB/HIV--patients but not in TB/AIDS patients. We conclude that tuberculosis in late stage HIV-infected patients has a distinct inflammatory cell profile, suggesting an enhanced compensatory mechanism that amplifies the unspecific inflammatory reaction. PMID- 10714480 TI - Therapeutic equivalence of inhaled beclomethasone dipropionate with CFC and non CFC (HFA 134a) propellants both delivered via the Easibreathe inhaler for the treatment of paediatric asthma. AB - Chlorofluorocarbon (CFC)-containing inhalers for use in the treatment of asthma are to be phased out under the terms of the Montreal Protocol (1). In this multi centre, randomized, double-blind study, the therapeutic equivalence of two formulations of beclomethasone dipropionate (BDP) containing CFC or non-CFC (HFA134a) propellant, both delivered via the Easibreathe (Norton Healthcare Ltd, London, U.K.) inhaler, was determined in 229 asthmatic children. Each child received 100 microg doses of BDP (containing either CFC or HFA propellant) twice daily for 12 weeks. Both CFC and HFA formulations produced statistically and clinically significant improvements in patient's lung function and symptom scores when administered via the Easibreathe inhaler. The improvements in mean morning peak expiratory flow (PEF) were 41 l min(-1) and 34 l min(-1) for the BDP-HFA and BDP-CFC products respectively (P<0.001) and for mean evening PEF the improvements were 38 l min(-1) and 38 l min(-1), respectively (P<0.001). Similar findings were demonstrated for the other efficacy parameters. The two formulations were statistically equivalent with respect to efficacy. For mean morning PEF the estimated treatment difference (BDP-CFC/BDP-HFA ratio) was 102.6% (95% CI 99.1, 106.2). Similar equivalence was shown for the other efficacy parameters. Both products were well tolerated, with no difference in the adverse event profiles, effects on 24 h urinary cortisol or Candida colonisation. This study demonstrates that the new formulation of BDP with HFA-134a propellant is equivalent to and directly substitutable for BDP with the older CFC propellant in a dose for dose manner. This should enable a seamless transition from one product to the other when CFC containing products are eventually phased out. In addition this study has also shown that the Easibreathe inhaler is an effective delivery system for use with inhaled products for the treatment of asthma in children. PMID- 10714482 TI - Clinical and in vitro evaluation of membrane humidifier that does not require addition of water. AB - It is well known that conventional bubbling humidifiers are capable of producing micro-aerosols contaminated with bacteria. We developed a unique humidifier, named a membrane humidifier, that does not require an external water supply. This new system obtains moisture from room air. We investigated the clinical and in vitro evaluation of the membrane humidifier. Ten patients with chronic pulmonary disease participated in the study. We evaluated the partial pressure of oxygen in arterial blood (PaO2) of 10 patients who used the new device. We conducted an in vitro study to determine whether the device could prevent the bacterial contamination of humidified-oxygen. We passed compressed air contaminated with Pseudomonas aeruginosa outside the hollow fibres of the membrane humidifier, and the humidified-oxygen passed inside the hollow fibres was sampled into nutrient broth periodically for 10 days. We also compared the relative humidity of oxygen humidified by a membrane humidifier with that of oxygen humidified by a bubbling humidifier. There was no significant difference between measured PaO2 while breathing oxygen humidified using a membrane humidifier and that while breathing oxygen humidified using a bubbling humidifier. Cultures of the humidified-oxygen passed through the hollow fibres were negative for bacteria. The membrane humidifier could produce good humidification. The new device appeared to prevent bacterial contamination, and may help to reduce the risk of infection in patients at hospital and home. PMID- 10714483 TI - Compliance with nasal continuous positive airway pressure assessed with a pressure monitor: pattern of use and influence of sleep habits. AB - The aim of the study was to assess compliance with nasal continuous positive airway pressure (N-CPAP) at home in patients with obstructive sleep apnoea syndrome (OSAS) and to search for predictors of compliance. We studied a cohort of 106 consecutive patients (91 men, 15 women) with a median apnoea hypopnoea index of 62.4 (range 21-132) h(-1), equipped at home with a Rem+ Soft device (Sefam, France), including a pressure monitor and a real-time clock. During the third and fourth months of treatment, the patients used their machine a median of 88% of days (16-100%), with a mean effective use of 5.6 (1.3-11.2) h per effective day. Residual apnoea index on N-CPAP, as recorded by the monitor, was 1.5 (0.3-27.6) h(-1). Mean clock-time for starting with N-CPAP was 23 h 54 min (21 h 34-01 h 42). The mean effective use per effective day correlated negatively with the minimal (and the mean) level of oxyhaemoglobin saturation (r(s) = -0.24, P < 0.05) while the percentage of days the machine was used correlated negatively with the percentage of slow wave sleep (r(s) = -0.22, P < 0.05) at baseline polysomnography. In a subset of 30 subjects, earlier start on N-CPAP correlated with longer use of the device in 22 patients (median r--0.48). We conclude that a pressure monitor allows reporting on compliance in terms of regularity (% of days the machine is used) and length of sleep on N-CPAP (effective use per effective day). These compliance variables show modest correlations with baseline polysomnographic features. Late bedtime should be discouraged as it might decrease compliance. PMID- 10714484 TI - Effect of hospital asthma nurse appointment on inpatient asthma care. AB - While asthma nurses are funded by many health authorities within the U.K. National Health Service, for the improvement of clinical management in both inpatient and outpatient settings in secondary care, the effect of asthma nurse appointment on acute asthma care in hospitalized children has been inadequately studied. Here, we test the hypothesis that the employment of a full-time hospital asthma nurse improves quality of care for children admitted to hospital with acute asthma. Prospective in design, the study compares analyses of indicators of good clinical practice for hospitalized asthmatic children (2-16 yrs) before and after the appointment of a hospital asthma nurse. Both management [oxygen saturation check (35/106 vs. 111/126, P<0.05)] and discharge planning [self management plan/asthma education (17/106 vs. 49/126, P<0.05), follow-up arrangements with general practice (8/106 vs. 25/126, P<0.05)] improved. There was, however, no significant change in oral steroid administration, peak flow check, inhaler technique assessment, inhaled drug prophylaxis or arrangements for hospital follow-up at discharge. Employment of a hospital-based children's asthma nurse leads to significant improvement in aspects of routine in-patient asthma management. However, other important areas of in-patient asthma care did not improve following nurse-led interventions. A clearer evidence base may improve compliance with asthma management guidelines, and could make the role of hospital asthma nurse more effective. PMID- 10714485 TI - Long-term effects nasal continuous positive airway pressure on daytime sleepiness, mood and traffic accidents in patients with obstructive sleep apnoea. AB - To describe the long-term effects of nasal continuous positive airway pressure (CPAP) on the rate of traffic car accidents, excessive daytime sleepiness (EDS) and mood in patients with obstructive sleep apnoea syndrome (OSAS), we investigated the changes of these parameters before and after nasal CPAP treatment using a questionnaire. Seventy-five male patients who were diagnosed with severe OSAS by polysomnography were evaluated for driving competence, by looking at their driving history for 2 yr, for EDS by the Epwarth Sleepiness Scale (ESS) and for mood by the Self-related Depression Scale (SDS), and then underwent nasal CPAP treatment. After 2 yr of treatment, questionnaires inquiring about the patients' use of CPAP, their ESS, SDS and driving history during treatment were sent to the patients. A total of 47 patients (63%) responded to these questionnaires. Forty-six of the 47 responders had continued to use the nasal CPAP and completed the questionnaire. No traffic car accidents were observed among the 39 routine car users during treatment, while 13 of 39 patients (33%) had had car accidents before treatment. Although near-miss accidents had been reported by 32 of 39 patients (82%) before treatment, only four patients reported near-miss accidents during nasal CPAP treatment. The mean score of ESS was significantly (P<0.01) reduced in 46 patients after nasal CPAP. The mean score of SDS was also decreased (P<0.01) after nasal CPAP in 46 patients. Although 26 of 41 patients had been depressive on SDS before treatment, the mood was improved in 13 patients after nasal CPAP. These results suggest that long term nasal CPAP treatment reduces the rate of traffic car accidents and improves the EDS and the mood in patients with OSAS. PMID- 10714486 TI - Pet ownership and asthma morbidity. PMID- 10714487 TI - Eosinophilic pleural effusion due to gliclazide. PMID- 10714488 TI - Secondary amines as new pharmacophores for macrofilaricidal drug design. AB - Several secondary amines exhibit promising macrofilaricidal response in vivo through oral route of administration against Acanthocheilonema viteae in which N hexylcyclohexylamine (1) shows 100% macrofilaricidal activity while a tertiary amine such as 9 elicits predominantly microfilaricidal (93%) response. PMID- 10714489 TI - 4-Aza-2,3-dehydro-4-deoxypodophyllotoxins: simple aza-podophyllotoxin analogues possessing potent cytotoxicity. AB - 4-Aza-2,3-dehydro-4-deoxypodophyllotoxin analogues 3a-n were synthesized through quinolines 2a-n. Comparison of their cytotoxicity against P-388 leukemia cells revealed that the steric effects of the ring B substituents on the activity are greater than the electronic effects, while the presence of a methoxy group on the ring E is not essential to exhibit potent cytotoxicity. Analogues 3a and 3b proved to be more than twice as cytotoxic as natural podophyllotoxin (1). PMID- 10714490 TI - Antihypertensive activity of substituted 2,3,8,8a-tetrahydro-7H-oxazolo[3,2 a]pyridinedicarboxylate enantiomers. AB - The synthesis and antihypertensive activity of racemates and enantiomers of substituted 2,3,8,8a-tetrahydro-7H-oxazolo[3,2-a]pyridinedicarboxylates have been reported. PMID- 10714491 TI - Effects of stilbene constituents from rhubarb on nitric oxide production in lipopolysaccharide-activated macrophages. AB - Two new anthraquinone glucosides [chrysophanol 8-O-beta-D-(6'-galloyl) glucopyranoside, aloe-emodin 1-O-beta-D-glucopyranoside] together with various known stilbenes and their glucosides, anthraquinone glucosides, and a naphthalene glucoside were isolated from the rhizome of Rheum undulatum L. Three stilbenes (rhapontigenin, piceatannol, resveratrol), a naphthalene glucoside (torachrysone 8-O-beta-D-glucopyranoside), and two stilbene glucoside gallates (rhaponticin 2'' O-gallate, rhaponticin 6''-O-gallate) showed inhibitory activity of NO production in lipopolysaccharide-activated macrophages, (IC50 = 11-69 microM). The oxygen functions (-OH,-OCH3) at the benzene ring were found to be essential to show the activity. Whereas, the glucoside moiety reduced the activity, while the alpha,beta-double bond did not affect the activity. Furthermore, the active stilbenes (rhapontigenin, piceatannol, resveratrol) inhibited iNOS induction. PMID- 10714492 TI - 2-Methyladenosine-Substituted 2',5'-oligoadenylates: conformations, 2-5A binding and catalytic activities with human ribonuclease L. AB - 2-Methyladenosine-substituted analogues of 2-5A, p5'A2'p5'A2'p5'(me2A), p5'(me2A)2'p5'A2'p5'A, and p5'(me2A) 2'p5'(me2A)2'pS'(me2A), were prepared via a modification of a lead ion-catalyzed ligation reaction. These 5'-monophosphates were subsequently converted into the corresponding 5'-triphosphates. Both binding and activation of human recombinant RNase L by various 2-methyladenosine substituted 2-5A analogues were examined. Among the 2-5A analogues, p5'A2'p5'A2'p5'(me2A) showed the strongest binding affinity and was as effective as 2-5A itself as an activator of RNase L. The CD spectra of both p5'(me2A)2'p5'A2'p5'A and p5'A2'p5'A2'p5'(me2A) were superimposable on that of p5'A2'p5'A2'p5'A, indicative of an anti orientation about the base-glycoside bonds as in naturally occurring 2-5A. PMID- 10714493 TI - Synthesis and antibacterial activity of 2alpha-functionalized 1beta methylcarbapenems related to KR-21012. AB - The 2alpha-functionalized 1beta-methylcarbapenems 3 were synthesized from the 2 formyl 1beta-methylcarbapenem intermediate 5. The best compound in the series of 2alpha-(hydroxy)alkylcarbapenems, KR-21012, displayed well balanced in vitro and in vivo activity as a parent compound of oral carbapenem. PMID- 10714494 TI - Antibody against a novel, myriocin (ISP-I)-based immunosuppressant, FTY720. AB - An antibody was prepared by immunizing rabbits with an ovalbumin conjugate of 2 amino-2-(2-(4-(4-mercaptobutyl)phenyl)ethyl)propane-1,3-diol HCl (AMPD-4), which contains the essential structure of the novel immunosuppressant FTY720. As the antibody reacted to not only AMPD-4, but also FTY720, it should be useful for immunoassay of FTY720 in body fluids, tissues and cells. PMID- 10714495 TI - Synthesis of a polymeric 4-N-linked sialoside which inhibits influenza virus hemagglutinin. AB - A multiple sialic acid-bearing polymer 7 has been made in which a novel 4-N substituted sialoside 5 has been coupled to polyacrylamide. The conjugate 7 has been found to inhibit the agglutination of influenza virus to red blood cells with HAI inhibition constants of around 10(-6) M, based on the sialic acid concentration. PMID- 10714496 TI - Design and biological evaluation of non-peptide analogues of omega-conotoxin MVIIA. AB - Omega-conotoxin MVIIA, a highly potent antagonist of the N-type voltage sensitive calcium channel, has shown utility in several models of pain and ischemia. We report a series of three alkylphenyl ether based analogues which mimic three key amino acids of the toxin. Two of the compounds have been found to exhibit IC50 values of 2.7 and 3.3 microM at the human N-type voltage sensitive calcium channel. PMID- 10714497 TI - Synthesis of a dihydroxythiophene analogue of catechosporines. AB - A vinylogous cephalosporine bearing a dihydroxythiophene moiety as a potential catechol surrogate has been synthesised (I-e-beta). Even if the anti staphylococcus spectrum displayed by this compound is of interest, its activity against Pseudomonas species, expected for such a structure, remains disappointing. PMID- 10714498 TI - Synthesis of 8-chloro-benzo[c]quinolizin-3-ones as potent and selective inhibitors of human steroid 5alpha-reductase 1. AB - The synthesis of a series of differently substituted 8-chloro-benzo[c]quinolizin 3-ones, as potent and selective human steroid 5alpha-reductase type 1 inhibitors, has been accomplished by a four-step procedure based on the TiCl4-promoted tandem Mannich-Michael cyclization of 2-silyloxy-1,3-butadienes with N-t-Boc iminium ions from quinolin-2-ones. The presence on the benzo[c]quinolizinone nucleus of a methyl group and a double bond at positions 6 and 4-4a, respectively, as in compound 1d, gave rise to one of the most potent non-steroidal 5alphaR-1 inhibitors reported so far (IC50 = 14 nM). PMID- 10714499 TI - Synthesis of the fatty sterol bound protein for a new sterol antibody. AB - For the purpose of applying the particular antibodies as a new diagnostic procedure for atherosclerosis and related diseases, we successfully achieved the synthesis of the fatty sterol with a linker, then linked the target protein to this sterol. Synthesis was started from pregnenolone and achieved by the Grignard reaction with pentenyl magnesium bromide, regioselective photoaddition of thiolacetic acid toward the 25-double bond, esterification of 3-OH with linoleic anhydride, in situ conjunction of the cross-linker (MBS) to the thiol group after selective deprotection from its acetyl ester, and finally by the reaction with protein such as KLH or albumin through this linker. PMID- 10714500 TI - Development of an affinity-driven cross-linker: isolation of a vitamin D receptor associated factor. AB - A vitamin D analogue containing an affinity and a photoaffinity probe (affinity driven cross-linker, Double Label) was synthesized. An unknown factor, associated with vitamin D receptor (VDR), was isolated from rat liver nuclear extract using a GST-VDR-ligand-binding domain fusion protein (GST-VDR-LBD), affinity labeled with Double Label, and protein-protein cross-linking by photolysis. PMID- 10714501 TI - A novel thermophilic glycosynthase that effects branching glycosylation. AB - A novel thermophilic glycosynthase that effects branching glycosylation has been obtained by mutation of the nucleophile in the active site of the glycosidase from Sulfolobus solfataricus. Two methods for the use of this mutant are reported. PMID- 10714502 TI - Novel C-ring analogues of 20(S)-camptothecin. Part 3: synthesis and their in vitro cytotoxicity of A-, B- and C-ring analogues. AB - Several 5-substituted alkoxy 20(S)-camptothecin analogues having A- and B-ring substituents were prepared via semi-synthesis. Most of these compounds were found to exhibit potent anti-cancer activity based on their in vitro cytotoxicity data obtained against human tumor cell lines. PMID- 10714503 TI - Novel oximes having 5-benzyl-2,4-thiazolidinedione as antihyperglycemic agents: synthesis and structure-activity relationship. AB - Oximes having 5-benzyl-2,4-thiazolidinedione were prepared, and their PPAR gamma agonistic activities and blood glucose lowering activities were evaluated. Biaromatic and methyl groups, attached to the oxime moiety, and the ethylene bridge between oxime and phenoxy groups are favorable to biological activities. PMID- 10714504 TI - Construction of HIV Rev peptides containing peptide nucleic acid that bind HIV RRE IIB RNA. AB - Peptides containing peptide nucleic acid (PNA) have been designed and synthesized to construct molecules recognizing a bulge or a loop structure of RNA. Such peptides were here designed from the HIV Rev protein that can bind the stem-loop IIB of the Rev responsive element (RRE) RNA. Variations of PNA modulated the binding affinities of the peptides to RRE IIB RNA. PMID- 10714505 TI - Enhancing the aqueous solubility of d4T-based phosphoramidate prodrugs. AB - A range of polyether para-substituted phosphoramidates were synthesised and found to have substantially elevated aqueous solubilities compared to the underivatised parent prodrug. A 30-fold increase in aqueous solubility could be achieved without a substantial decrease of in vitro activity against HIV-1. Replacement of the aryl (i.e. phenolic) moiety by tyrosine led to a substantial enhancement in aqueous solubility but also to a decrease in antiviral potency. A previously unobserved trend was identified, relating increased aryl substituent steric bulk to decreased antiviral activity. PMID- 10714506 TI - Fused bicyclic Gly-Asp beta-turn mimics with potent affinity for GPIIb-IIIa. Exploration of the arginine isostere. AB - 6-[4-Amidinobenzoyl]amino]-tetralone-2-acetic acid is a potent antagonist of GPIIb-IIIa. Substitution in the meta position of the benzamidine, or replacement with a heteroaryl amidine was tolerated in this series. Use of an acyl-linked 4 alkyl piperidine as an arginine isostere also provided active compounds. Compounds from this series provided substantial systemic exposure in the rat following oral administration. PMID- 10714507 TI - Mechanism-based inactivation of thymidylate synthase by 5-(3-fluoropropyn-1-yl) 2'-deoxyuridine 5'-phosphate. AB - 5-Fluoropropynyl-2'-deoxyuridine 5'-phosphate (3) was designed as a mechanism based inactivator of thymidylate synthase (TS). The inhibitor was synthesized from 5-iodo-2'-deoxyuridine and propargyl alcohol by palladium-catalyzed coupling, followed by fluorination and selective phosphorylation. Incubation of TS with 3, in the presence or absence of the CH2H4folate cofactor, caused rapid, irreversible inactivation of the enzyme. PMID- 10714508 TI - 2-(Alkylthio)pyrimidin-4-ones as novel, reversible inhibitors of lipoprotein associated phospholipase A2. AB - Starting from two weakly active hits from high throughput screening, a novel series of 2-(alkylthio)-pyrimidin-4-ones with high potency and selectivity for lipoprotein-associated phospholipase A2 has been designed. In contrast to previously known inhibitors, these have been shown to act by a non-covalent and substrate competitive mechanism. PMID- 10714509 TI - Synthesis and pharmacology of a novel pyrrolo[2,1,5-cd] indolizine (NNC 45-0095), a high affinity non-steroidal agonist for the estrogen receptor. AB - 1-Ethyl-2-(4-hydroxyphenyl)pyrrolo[2,1,5-cd]indolizine (NNC 45-0095) is a novel compound which represents the parent pharmacophore structure of a series of pyrrolo[2,1,5-cd]indolizine derivatives with mixed estrogen agonist/antagonist properties. NNC 45-0095 binds with high affinity to the estrogen receptor (IC50=9.5 nM) and exhibits full protection of bone loss in the ovariectomized mouse model for post-menopausal osteoporosis. PMID- 10714510 TI - The discovery and synthesis of highly potent, A2a receptor agonists. AB - A series of N6,2-disubstituted adenosine analogues have been synthesized and their functional activity measured against A2a and A1 receptors. Examples of compounds with both a lipophilic N6-substituent and amino-functionalized 2 position were highly active at the A2a receptor on the human neutrophil. PMID- 10714511 TI - Limb loading activity of adult horses confined to box stalls in an equine hospital barn. AB - OBJECTIVE: To determine a range of limb loading activity for healthy adult horses confined to box stalls in an equine veterinary teaching hospital and determine the effects of hospital environmental factors on load rates and daily limb loading patterns. ANIMALS: 6 mature healthy horses of various ages, breeds, and sexes, and 1 horse with a repaired metatarsal fracture. PROCEDURE: Step monitors were placed on 2 limbs of adult horses confined to box stalls. Relocation steps and weight shifts were recorded, as loading events, for 24 hours. Influence of forelimb versus hind limb and environmental factors on load rate (loading events per hour) were assessed with repeated-measures ANOVA. RESULTS: Loading activity was greater for the forelimb than the hind limb and was greater during the day than the night. Loading activity differences were not associated with daytime environmental factors. CONCLUSIONS AND CLINICAL RELEVANCE: Horses with normal locomotor activity appear to have higher load rates for forelimbs compared with hind limbs and higher load rates during the day compared with night. Knowledge of influence of environmental factors and mechanical restraint on limb loading activity may be useful in management of horses with musculoskeletal disorders. This information may also be used for in vitro simulation of in vivo loading of limbs during cyclic biomechanical investigations. PMID- 10714512 TI - Detection of attenuated wavy fibers in the myocardium of Newfoundlands without clinical or echocardiographic evidence of heart disease. AB - OBJECTIVES: To determine whether attenuated wavy fibers may be found in the myocardium of Newfoundlands without clinical or echocardiographic evidence of heart disease. ANIMALS: 15 Newfoundlands from a kennel with a known predisposition to dilated cardiomyopathy (DCM) and 32 dogs of other breeds that died suddenly or were euthanatized for reasons unrelated to heart disease and did not have gross postmortem evidence of heart disease. PROCEDURE: Echocardiography was performed on all Newfoundlands on a yearly basis. Necropsy specimens from all dogs were evaluated for attenuated wavy fibers (i.e., myocardial cells <6 microm in diameter with a wavy appearance). RESULTS: None of the Newfoundlands had clinical signs of heart disease, and results of echocardiographic examinations were within reference ranges. Seven Newfoundlands had histologic evidence of attenuated wavy fibers, whereas attenuated wavy fibers were not found in the remaining 8 Newfoundlands or in any of the 32 dogs of other breeds. CONCLUSIONS AND CLINICAL RELEVANCE: Findings suggest that attenuated wavy fibers in dogs with a known predisposition for DCM may indicate an early stage of the disease. However, further studies on a larger number of dogs are needed to confirm this hypothesis. PMID- 10714513 TI - Calcium regulation by skeletal muscle membranes of horses with recurrent exertional rhabdomyolysis. AB - OBJECTIVE: To determine whether an alteration in calcium regulation by skeletal muscle sarcoplasmic reticulum, similar to known defects that cause malignant hyperthermia (MH), could be identified in membrane vesicles isolated from the muscles of Thoroughbreds with recurrent exertional rhabdomyolysis (RER). SAMPLE POPULATION: Muscle biopsy specimens from 6 Thoroughbreds with RER and 6 healthy (control) horses. PROCEDURES: RER was diagnosed on the basis of a history of > 3 episodes of exertional rhabdomyolysis confirmed by increases in serum creatine kinase (CK) activity. Skeletal muscle membrane vesicles, prepared by differential centrifugation of muscle tissue homogenates obtained from the horses, were characterized for sarcoplasmic reticulum (SR) activities, including the Ca2+ release rate for the ryanodine receptor-Ca2+ release channel, [3H]ryanodine binding activities, and rate of SR Ca2+-ATPase activity and its activation by Ca2+. RESULTS: Time course of SR Ca2+-induced Ca2+ release and [3H]ryanodine binding to the ryanodine receptor after incubation with varying concentrations of ryanodine, caffeine, and ionized calcium did not differ between muscle membranes obtained from control and RER horses. Furthermore, the maximal rate of SR Ca2+ ATPase activity and its affinity for Ca2+ did not differ between muscle membranes from control horses and horses with RER. CONCLUSIONS AND CLINICAL RELEVANCE: Despite clinical and physiologic similarities between RER and MH, we concluded that RER in Thoroughbreds does not resemble the SR ryanodine receptor defect responsible for MH and may represent a novel defect in muscle excitation contraction coupling, calcium regulation, or contractility. PMID- 10714514 TI - Role of platelet-activating factor in alveolar septal injury associated with experimentally induced pneumonic pasteurellosis in calves. AB - OBJECTIVE: To determine whether platelet-activating factor (PAF) is involved in acute lung microvascular injury associated with pneumonic pasteurellosis in calves. ANIMALS: 15 healthy 2- to 4-week-old male Holstein calves. PROCEDURE: Calves were anesthetized and inoculated intrabronchially with saline (0.9% NaCl) solution (n = 5) or 1x10(9) Pasteurella haemolytica organisms (n = 10). Of the 10 calves inoculated with P haemolytica, 5 also were treated with WEB 2086, a potent inhibitor of PAF, and 5 were treated with vehicle. Blood and bronchoalveolar lavage samples were collected before and 1, 2, 4, and 6 hours after inoculation of P. haemolytica. Blood samples were analyzed to evaluate total number and differential counts of leukocytes, dilute whole-blood leukocyte deformability, size of neutrophils, and neutrophil CD11b expression. Bronchoalveolar lavage samples were analyzed for total number and differential counts of nucleated cells, total protein concentration, and hemoglobin concentration. Size and gross and histologic appearance of lung lesions also was determined. RESULTS: Treatment of calves with WEB 2086 reduced size of lung lesions, attenuated the increase in microvascular permeability, and reduced neutrophil infiltration in the first 4 hours after inoculation. Treatment with WEB 2086 also attenuated a decrease in leukocyte deformability, increase in size of neutrophils, and CD11b expression by circulating neutrophils. CONCLUSIONS AND CLINICAL RELEVANCE: It appears that PAF is a major mediator for altered lung microvascular permeability and activation of circulating neutrophils in the first 4 hours after onset of pneumonic pasteurellosis in calves. PMID- 10714515 TI - Digital angiography of the feet of horses. AB - OBJECTIVE: To describe the vascular anatomy of the palmar digital artery and its major branches in the equine foot and to quantify the diameter of these vessels by use of digital angiograms. Sample Population-6 thoracic limbs obtained from 6 horses. PROCEDURE: Distal portions of each limb were perfused with aerated Krebs Henseleit solution. Digital angiograms were acquired in standing and lateral recumbent positions, following an intra-arterial injection of iopamidol. Select vessels were measured on radiographic views, and values were corrected for magnification. RESULTS: The palmar digital artery tapered from 2.28 mm at the coronary region to 1.61 mm at the entrance to the solar canal, and the major arterial branches ranged in diameter from 0.71 to 1.42 mm in the standing position. CONCLUSIONS AND CLINICAL RELEVANCE: Digital angiography is useful for imaging small vessels, but penumbra limits the image resolution of the macrovasculature of the foot. The palmarodorsal projection is more useful for evaluation of the terminal arch and solar branches, but 2 projections are necessary for a thorough examination of the foot. Image magnification, position of horse, and vascular response to contrast medium must be considered in the quantitative assessment of vessel diameter. Digital angiography may be performed in clinical cases and research models for examination of vascular perfusion of the distal portion of the limb. PMID- 10714516 TI - Respiratory reflexes in response to nasal administration of halothane to anesthetized, spontaneously breathing dogs. AB - OBJECTIVE: To characterize and determine the sensory innervation of respiratory reflexes elicited by nasal administration of halothane to dogs. ANIMALS: 10 healthy Beagles. PROCEDURE: Dogs underwent permanent tracheostomy and, 2 to 3 weeks later, were anesthetized with thiopental and alpha-chloralose administered IV. The nasal passages were functionally isolated so that halothane could be administered to the nasal passages while dogs were breathing 100% O2 via the tracheostomy. Respiratory reflexes in response to administration of halothane at concentrations of 1.25, 1.75, and 2.5 times the minimum alveolar concentration (MAC), and 5% (administered in 100% O2 at a flow rate of 5 L/min) were recorded. Reflexes in response to administration of 5% halothane were also recorded following transection of the infraorbital nerve, transection of the caudal nasal nerve, and nasal administration of lidocaine. RESULTS: Nasal administration of halothane induced an inhibition of breathing characterized by a dose-dependent increase in expiratory time and a resultant decrease in expired volume per unit time. Effects were noticeable immediately after the onset of halothane administration and lasted until its cessation. Reflex responses to halothane administration were attenuated by transection of the caudal nasal nerve and by nasal administration of lidocaine, but transection of the infraorbital nerve had no effect. CONCLUSIONS AND CLINICAL RELEVANCE: Nasal administration of halothane at concentrations generally used for mask induction of anesthesia induces reflex inhibition of breathing. Afferent fibers in the caudal nasal nerve appear to play an important role in the reflex inhibition of breathing induced by halothane administration. PMID- 10714517 TI - Histomorphometric analysis of the proximal portion of the femur in healthy dogs. AB - OBJECTIVE: To describe the cancellous bone architecture of the head and neck of the femur in healthy dogs by use of automated histomorphometry techniques in conjunction with histologic grading of articular cartilage. ANIMALS: 30 mature male dogs with healthy coxo-femoral joints PROCEDURE: Dogs were 1.5 to 4 years old and weighed 27 to 37 kg. Computer images of fine-detail radiographs of 100 microm-thick coronal and transverse plane sections of the head and neck of the femur (14 dogs) were analyzed by use of histomorphometry software. Statistical comparisons among histomorphometric indices of 4 regions were performed. Histologic preparations of coronal and transverse plane sections of femoral head articular cartilage (16 dogs) were graded. Median grades for lateral, medial, cranial, and caudal halves of the femoral head articular cartilage were determined. RESULTS: Bone volume/total volume, trabecular thickness and number, and bone surface/total volume were significantly higher in the femoral head than in the femoral neck. Anisotropy (trabecular alignment) and trabecular separation were significantly higher in the femoral neck than in the femoral head. Anisotropy was significantly higher in the caudal half of the femoral neck than in the cranial half. Cartilage had histologic grades indicating health without significant differences among lateral, medial, cranial, and caudal halves of femoral head cartilage. CONCLUSIONS AND CLINICAL RELEVANCE: A predictable cancellous architecture in the head and neck of the femur is associated with healthy cartilage. PMID- 10714518 TI - Effects of lactoferrin and milk on adherence of Streptococcus uberis to bovine mammary epithelial cells. AB - OBJECTIVE: To determine whether lactoferrin (LF) or milk influenced adherence of Streptococcus uberis to bovine mammary epithelial cells. SAMPLE POPULATION: Three strains of S uberis from cows with mastitis, pooled milk samples from 3 clinically healthy Jersey cows early in the lactation period, and bovine mammary epithelial cells from a clonal cell line. PROCEDURES: Adherence of S uberis to bovine mammary epithelial cells in the presence of various concentrations of LF or milk and after pretreatment of bacteria with LF or milk was tested. Bacteria were cultured with mammary epithelial cell monolayers for 1 hour. The culture supernatant was removed, and the epithelial cells were lysed. Adherence index was calculated as number of colony-forming units (CFU) in the cell lysate divided by number of CFU in the supernatant times 10,000. RESULTS: All 3 strains of S uberis were found to bind to purified LF and LF in milk. Addition of LF to the culture medium enhanced adherence of all 3 strains to mammary epithelial cells, whereas addition of milk enhanced adherence of 2 strains and decreased adherence of the third. Pretreatment of bacteria with LF or milk increased adherence of 1 of the strains but decreased adherence of the other 2. Increased adherence was antagonized by rabbit antibovine LF antibody. CONCLUSIONS: Results suggest that LF may function as a bridging molecule between S uberis and bovine mammary epithelial cells, facilitating adherence of the bacteria to the cells. PMID- 10714519 TI - Effect of sample number and time on determination of plasma clearance of technetium Tc 99m pentetate and orthoiodohippurate sodium I 131 in dogs and cats. AB - OBJECTIVE: To determine the effect of number of blood samples and sampling times on plasma clearance of technetium Tc 99m pentetate (Tc99mP) and orthoiodohippurate sodium I 131(OIH). ANIMALS: 20 dogs and 14 cats. PROCEDURE: Plasma clearances of OIH and Tc99mP were calculated by use of a 2-compartment model, on the basis of a 12-point curve as a reference method. Plasma clearance was calculated by use of all possible combinations of 4 to 11 samples. Time schedule yielding the smallest difference from the reference method was considered to be optimal. Regression analysis was performed between the 12-point model and models using a reduced number of samples. RESULTS: SD of the difference between the 12-point clearance and the models with reduced numbers of samples increased when the number of samples decreased. The SD of the difference between 12-point clearance and 4-point clearance was 4.17 ml/min for OIH and 0.94 ml/min for Tc99mP in dogs and 0.45 ml/min for OIH and 0.11 ml/min for Tc99mP in cats. Optimal schedules were distributed logarithmically and included an early sample at 5 or 10 minutes, a late sample at 2.5, 3, 4, or 5 hours for OIH, and a late sample at 4 or 5 hours for Tc99mP. CONCLUSIONS AND CLINICAL RELEVANCE: Plasma clearances of OIH and Tc99mP can be accurately calculated in dogs and cats by use of a single-injection 2-compartment pharmacologic model with a reduced number of blood samples, resulting in an acceptable margin of error. PMID- 10714520 TI - Doppler ultrasonography and single-fiber laser Doppler flowmetry for measurement of hind limb blood flow in anesthetized horses. AB - OBJECTIVE: To use Doppler ultrasonography and single-fiber laser Doppler flowmetry (LDF) to evaluate blood flow in the dependent and nondependent hind limbs of anesthetized horses and to evaluate changes in femoral arterial blood flow and microvascular skeletal muscle perfusion in response to administration of phenylephrine hydrochloride or dobutamine hydrochloride. ANIMALS: 6 healthy adult horses. PROCEDURE: Horses were anesthetized and positioned in left lateral recumbency. Doppler ultrasonography was used to measure velocity and volumetric flow in the femoral vessels. Single-fiber LDF was used to measure relative microvascular perfusion at a single site in the semimembranosus muscles. Phenylephrine or dobutamine was then administered to decrease or increase femoral arterial blood flow, and changes in blood flow and microvascular perfusion were recorded. RESULTS: Administration of phenylephrine resulted in significant decreases in femoral arterial and venous blood flows and cardiac output and significant increases in mean aortic blood pressure, systemic vascular resistance, and PCV. Administration of dobutamine resulted in significant increases in femoral arterial blood flow, mean aortic blood pressure, and PCV. Significant changes in microvascular perfusion were not detected. CONCLUSION AND CLINICAL RELEVANCE: Results suggest that Doppler ultrasonography and single-fiber LDF can be used to study blood flows in the hind limbs of anesthetized horses. However, further studies are required to determine why changes in femoral arterial blood flows were not associated with changes in microvascular perfusion. PMID- 10714521 TI - Bovine respiratory syncytial virus-specific IgE is associated with interleukin-2 and -4, and interferon-gamma expression in pulmonary lymph of experimentally infected calves. AB - OBJECTIVE: To study the local immune response of calves to bovine respiratory syncytial virus (BRSV) infection with emphasis on IgE production and cytokine gene expression in pulmonary lymph. ANIMALS: Twelve 6- to 8-week-old Holstein bull calves. Six similar control calves were mock infected to obtain control data. PROCEDURE: Lymphatic cannulation surgery was performed on 12 calves to create a long-term thoracic lymph fistula draining to the exterior. Cannulated calves were exposed to virulent BRSV by aerosol. Lymph fluid collected daily was assayed for BRSV and isotype-specific IgE antibody, total IgG, IgA, IgM, and protein concentrations. Interleukin-4 (IL-4), interleukin-2 (IL-2), and interferon-gamma were semi-quantitated by reverse transcription-polymerase chain reaction (RT-PCR). Cell counts and fluorescence-activated cell scanner (FACSCAN) analysis of T-cell subsets were performed on lymph cells. RESULTS: Calves had clinical signs of respiratory tract disease during days 5 to 10 after infection and shed virus. Bovine respiratory syncytial virus-specific IgE in infected calves was significantly increased over baseline on day 9 after infection. Mean virus-specific IgE concentrations strongly correlated with increases in severity of clinical disease (r = 0.903). Expression of IL-2, IL-4, and interferon-gamma was variably present in infected and control calves, with IL-4 expression most consistent during early infection. CONCLUSIONS AND CLINICAL RELEVANCE: Infection with BRSV was associated with production of BRSV-specific IgE, and IL-4 message was commonly found in lymph cells of infected calves. This finding supports the concept that BRSV-induced pathophysiology involves a T helper cell type-2 response. Effective therapeutic and prophylactic strategies could, therefore, be developed using immunomodulation to shift the immune response more toward a T helper cell type-1 response. PMID- 10714522 TI - Blood oxygen binding in double-muscled calves and dairy calves with conventional muscle conformation. AB - OBJECTIVE: To assess in vivo blood oxygen binding in double-muscled calves and dairy calves with conventional muscle conformation. ANIMALS: 58 dairy and 48 double-muscled calves. PROCEDURE: Calves were classified as neonatal (24 hours old) or older calves (2 to 26 days old). Venous and arterial blood samples were collected, and hemoglobin concentration, pH, PCO2, and PO2 were determined. Blood oxygen equilibrium curves (OEC) under standard conditions were constructed, and the oxygen exchange fraction (OEF) and the amount of oxygen released at the tissue level by 100 ml of blood (OEF Vol%) were calculated. RESULTS: In each breed, partial pressure of oxygen at 50% saturation of hemoglobin (P50) under standard conditions was significantly higher in older than in neonatal calves, indicating a right shift in OEC with age. Venous P50 was significantly lower in neonatal double-muscled calves than in neonatal dairy calves, but arterial and venous P50 were significantly higher in older double-muscled calves than in older dairy calves. In double-muscled, but not in dairy, calves, OEF was significantly higher in older than in neonatal calves. In neonatal calves, OEF Vol% was not significantly different between breeds, but OEF Vol% was significantly higher in older double-muscled calves than in older dairy calves. CONCLUSIONS AND CLINICAL RELEVANCE: The lower OEF in neonatal double-muscled calves, compared with dairy calves, could contribute to the higher sensitivity of double-muscled calves to hypoxia. Blood oxygen affinity decreased with age, but OEF and OEF Vol% were unchanged with age in dairy calves, whereas they increased with age in double muscled calves. PMID- 10714523 TI - Single primer polymerase chain reaction fingerprinting for Pasteurella multocida isolates from laboratory rabbits. AB - OBJECTIVE: To evaluate a rapid polymerase chain reaction (PCR) fingerprinting technique for discriminating among Pasteurella multocida isolates from laboratory rabbits. SAMPLE POPULATION: 33 P multocida isolates from rabbits with clinical pasteurellosis. PROCEDURE: PCR assays were conducted with 2 minisatellites (core sequence and modified core sequence of phage M13) and 2 microsatellites ([GTG]5 and [GACA]4). Each bacterium was assigned to a PCR type for each of the primers used. Boiled bacterial extracts and purified genomic DNA were compared by use of PCR assays for phage M13 and (GACA)4. Plasmids were isolated from each bacterium, and their influence on PCR fingerprint was determined, using boiled extracts as a DNA source. RESULTS: M13 core sequence and M13 modified core sequence yielded 5 and 8 PCR types, respectively. The microsatellites (GTG)5 and (GACA)4 yielded 4 and 9 PCR fingerprint types, respectively. Fingerprint patterns obtained by use of isolated DNA differed from those obtained by use of boiled extracts, although discrimination among P multocida isolates was similar. The presence or absence of plasmids did not affect PCR fingerprints. CONCLUSION: Single primer PCR fingerprinting with minisatellite and microsatellite primers is an efficient and reproducible method for the discrimination of P multocida isolates from rabbits and can be performed directly, using boiled bacterial extracts as a source of template, although more bands were obtained from pure genomic DNA. PMID- 10714524 TI - Comparison of nuclear scintigraphy and acetaminophen absorption as a means of studying gastric emptying in horses. AB - OBJECTIVE: To evaluate the correlation between halftime of liquid-phase gastric emptying (T50), determined with nuclear scintigraphy using technetium Tc 99m pentetate, and absorption variables of orally administered acetaminophen. ANIMALS: 6 mature horses. PROCEDURE: Technetium Tc 99m pentetate (10 mCi) and acetaminophen (20 mg/kg of body weight) were administered simultaneously in 200 ml of water. Serial left and right lateral images of the stomach region were obtained with a gamma camera, and T50 determined separately for counts obtained from the left side, the right side and the geometric mean. Power exponential curves were used for estimation of T50 and modified R2 values for estimation of goodness of fit of the data. Serial serum samples were taken, and acetaminophen concentration was determined, using fluorescence polarization immunoassay. Maximum serum concentration (Cmax), time to reach maximum serum concentration (Tmax), area under the curve for 240 minutes and the absorption constant (Ka) were determined, using a parameter estimation program. Correlations were calculated, using the Spearman rank correlation coefficient. RESULTS: Correlations between T50 and Tmax and between T50 and Ka were significant. CONCLUSIONS AND CLINICAL RELEVANCE: Tmax and Ka are valuable variables in the assessment of liquid-phase gastric emptying using acetaminophen absorption. Acetaminophen absorption may be a valuable alternative to nuclear scintigraphy in the determination of gastric emptying rates in equine patients with normally functioning small intestine. PMID- 10714525 TI - Comparison of polymerase chain reaction assays with bacteriologic culture, immunofluorescence, and nucleic acid hybridization for detection of Leptospira borgpetersenii serovar hardjo in urine of cattle. AB - OBJECTIVE: To compare sensitivity and specificity of various polymerase chain reaction (PCR) assays for detection of Leptospira borgpetersenii serovar hardjo in bovine urine and to compare results of the optimal PCR assay with results of immunofluorescence, nucleic acid hybridization, and bacteriologic culture. ANIMALS: 6 heifers. PROCEDURE: Heifers were exposed to serovar hardjo type hardjo bovis by conjunctival instillation of 10(6) leptospires on 3 successive days. Urine samples were collected before and after infection. Sensitivity and specificity of 5 PCR assays were compared, to determine the optimal assay for use with bovine urine samples. The optimal PCR assay was then compared with results of bacteriologic culture, nucleic acid hybridization, and immunofluorescence. RESULTS: A PCR assay with the best combination of specificity (100%) and sensitivity (91%) was selected for comparison with the other diagnostic tests. Sensitivity for nucleic acid hybridization was 55%, whereas sensitivity for bacteriologic culture and immunofluorescence was 89 to 93%. CONCLUSIONS AND CLINICAL RELEVANCE: Bacteriologic culture, PCR, and immunofluorescence were sensitive for detection of L borgpetersenii serovar hardjo type hardjo-bovis in urine specimens of cattle, but a single technique was not the most sensitive for each animal tested. Therefore, the use of 2 techniques in combination is warranted for maximal sensitivity for diagnosis. PMID- 10714526 TI - Comparison of results of scanning electron microscopy and magnetic resonance imaging before and after administration of a radiographic contrast agent in the tendon of the deep digital flexor muscle obtained from horse cadavers. AB - OBJECTIVE: To analyze the tendon of the deep digital flexor (TDDF) muscle of the forelimb in horses by use of a contrast radiographic agent (gadopentate dimeglumine [Gd-DTPA/Dimeg]) and magnetic resonance imaging (MRI) and to determine the concentration of water protons in the tendons by use of MRI. SAMPLE POPULATION: 8 TDDF harvested from the forelimbs of 6 horse cadavers. PROCEDURE: Examinations were performed on the same portion of each tendon. Tendons were examined by use of two techniques: MRI before and after treatment with Gd DTPA/Dimeg as well as scanning electron microscopy. RESULTS: Tendons did not have detectable signal intensity on MRI before treatment with Gd-DTPA/Dimeg; however, intravascular injection of Gd-DTPA/Dimeg allowed evaluation of the internal structure of the tendons Scanning electron microscopy images correlated well with images obtained by use of MRI before and after administration of Gd-DTPA/Dimeg. Localized spectra revealed the concentration of water protons in the TDDF. CONCLUSIONS AND CLINICAL RELEVANCE: The techniques used in this study provided information about internal organization of the TDDF in horses. Analysis of results revealed that the best technique involved vascular injection of contrast medium. Results of MRI correlated well with results for scanning electron microscopy. After administration of Gd-DTPA/Dimeg, MRI provided additional information about tendon morphologic characteristics. This technique may be of value for examination of tendons in lame horses. PMID- 10714527 TI - Comparison of equine amnion and a nonadherent wound dressing material for bandaging pinch-grafted wounds in ponies. AB - OBJECTIVE: To evaluate healing of pinch-grafted wounds on the distal aspect of the limbs of ponies bandaged with equine amnion or a standard nonadherent wound dressing material. ANIMALS: 6 ponies. PROCEDURE: A 2.5x2.5-cm full-thickness section of skin was removed from the dorsal aspect of each limb at the midpoint of the metacarpus or metatarsus. Six days later, wounds were grafted with partial thickness pinch grafts. Half the wounds were bandaged with amnion, and the other half were bandaged with a nonadherent dressing. Bandages were changed every 3 days until wound healing was complete. At each bandage change, numbers of grafts lost were recorded, and wounds were measured. RESULTS: Percentage of grafts lost from wounds bandaged with amnion was not significantly different from percentage lost from wounds bandaged with the nonadherent dressing. Median healing time for wounds bandaged with amnion (30 days) was significantly less than median healing time for wounds bandaged with the nonadherent dressing (39 days). All wounds were healed by day 45. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that amnion can be used for bandaging pinch-grafted wounds on the distal aspect of the limbs of ponies. PMID- 10714528 TI - Evaluation of the safety of fenbendazole in cats. AB - OBJECTIVE: To evaluate the safety of fenbendazole in domestic cats. ANIMALS: 28 six- to seven-month old domestic short-hair cats. PROCEDURE: Cats were randomly assigned to 1 of 3 treatment groups or a control group (n = 7/group). Cats in the treatment groups were given fenbendazole at a dosage of 50, 150, or 250 mg/kg, PO, every 24 hours for 9 days; control cats were given a placebo. A fecal examination, coagulation tests, serum biochemical analyses, CBC, and urinalyses were performed before and 5, 9, and 21 days after initiation of treatment; cats were closely monitored for adverse reactions. After the last dose of fenbendazole was given, 4 control cats and 4 cats given fenbendazole at the highest dosage were euthanatized, and necropsies were performed. RESULTS: None of the cats developed any adverse reactions. For cats in the control and all treated groups, laboratory test results were within reference limits, and there were no significant differences in results of laboratory tests among groups. No gross or histologic lesions were identified in the control or treated cats that were euthanatized. CONCLUSIONS AND CLINICAL RELEVANCE: Fenbendazole administered to healthy cats at a dosage 5 times the dosage and 3 times the duration approved for use in dogs and wild felids did not cause any acute or subacute adverse reactions or pathologic changes. Results suggest that cats may be safely treated with fenbendazole. PMID- 10714529 TI - Short-term effects of ecadotril in dogs with induced congestive heart failure. AB - OBJECTIVE: To evaluate short-term hemodynamic effects of ecadotril in a model of congestive heart failure in dogs. ANIMALS: 6 conscious adult male dogs. PROCEDURES: Instruments were placed in dogs to measure left ventricular, aortic, and atrial blood pressures. Heart failure was induced by repeated coronary embolization with latex microspheres. Four times, and in random order, dogs were given vehicle or active drug (3, 10, or 30 mg/kg of body weight) orally. Hemodynamic variables, urine flow, and urinary electrolyte excretion were measured before and 30, 90, and 150 minutes, and 10 and 21 hours after drug administration. RESULTS: Changes in urine flow, heart rate, mean arterial pressure, or peak positive and negative rate of change in ventricular pressure were not apparent. Urinary sodium excretion significantly increased in response to the low and high doses of ecadotril but not in response to the 10 mg/kg dose. Left ventricular end diastolic pressure (LVEDP) consistently decreased in dose- and time-dependent manner. Maximal group-averaged reductions in LVEDP were 5.2, 8.1, and 10 mm Hg for the low, middle, and high doses, respectively. The magnitude of the decrease in LVEDP was not related to cumulative change in urine flow. CONCLUSIONS AND CLINICAL RELEVANCE: Orally administered ecadotril reduced left ventricular filling pressures in these dogs by a mechanism that does not require a substantial diuretic effect. Ecadotril may be effective for alleviating clinical signs in dogs with left-sided heart failure and may be particularly beneficial for use in dogs that are refractory to traditional diuretic therapy. PMID- 10714530 TI - Immune function of bovine leukocytes after in vitro exposure to selected heavy metals. AB - OBJECTIVE: To study effects of in vitro exposure of bovine leukocytes to mercury, cadmium, and lead on phagocytosis, natural killer cell activity, and lymphocyte proliferation. SAMPLE POPULATION: Leukocytes from 6 nonpregnant Holstein heifers. PROCEDURE: Leukocytes were exposed in vitro to the aforementioned metals, and leukocyte functions were assessed. RESULTS: Phagocytosis was suppressed by 10(-5) to 10(-7) M CdCl2 and by 10(-5) and 10(-6) M HgCl2, but not 10(-7) M HgCl2 nor 10(-4) to 10(-6) M PbCl2. Spontaneous and concanavalin A- or phytohemagglutinin stimulated proliferation of metal-treated bovine blood mononuclear cells was not significantly different from that of nontreated control cells, except for enhanced spontaneous proliferation in response to 10(-5) M HgCl2. When proliferation was expressed as a stimulation index, a dose-dependent increase of spontaneous proliferation was observed in response to exposure to HgCl2 and PbCl2. Compared with response to 10(-6) or 10(-7) M CdCl2, reduction of mitogen induced and spontaneous proliferation was observed on exposure to 10(-5) M CdCl2. Natural killer cell activity against YAC-1 target cells, evaluated by flow cytometry, was decreased only in cells exposed to 10 M HgCl2. CONCLUSION AND CLINICAL RELEVANCE: Bovine leukocytes are susceptible to the immunomodulatory effects of in vitro exposure to heavy metals at concentrations equal to or higher than those at which similar effects are seen for leukocytes from most other animal species for which data are available for comparison. Exception is phagocytosis, which is severely affected by low concentrations of CdCl2 and HgCl2 in cattle. Reduction of defense mechanisms on exposure to metals could lead to increased susceptibility to potential pathogens. PMID- 10714531 TI - Conservative lower back treatment reduces inhibition in knee-extensor muscles: a randomized controlled trial. AB - BACKGROUND: Knee-joint pathologies, such as anterior knee pain (AKP), are associated with strength deficits and reduced activation of the knee extensors, which is referred to as muscle inhibition (MI). MI is thought to prevent full functional recovery, and treatment modalities that help to reduce or eliminate MI appear necessary for successful rehabilitation. Clinical observations suggest that AKP is typically associated with sacroiliac (SI) joint dysfunction. It is unknown whether SI-joint dysfunction contributes to knee-extensor deficits and whether correction of SI-joint dysfunction alleviates MI. OBJECTIVE: The objective of this study was to assess whether conservative low back treatment reduces lower limb MI. STUDY DESIGN: In a randomized, controlled, double-blind study the effects of conservative lower back treatment on knee-extensor strength and MI were evaluated in patients with AKP. METHODS: Twenty-eight patients with AKP were randomly assigned to either a treatment or a control group. After a lower back functional assessment, the treatment group received a conservative treatment in the form of a chiropractic spinal manipulation aimed at correcting SI-joint dysfunction. The control group underwent a lower back functional assessment but received no joint manipulation. Before and after the manipulation or the lower back functional assessment, knee-extensor moments, MI, and muscle activation during full effort, isometric knee extensions were measured. RESULTS: Patients showed substantial MI in both legs. Functional assessment revealed SI joint dysfunction in all subjects (23 symptomatic and 5 asymptomatic). After the SI-joint manipulation, a significant decrease in MI of 7.5% was observed in the involved legs of the treatment group. MI did not change in the contralateral legs of the treatment group or the involved and contralateral legs of the control group. There were no statistically significant changes in knee-extensor moments and muscle activation in either group. CONCLUSIONS: The results of this study suggest that SI-joint manipulation reduces knee-extensor MI. Spinal manipulation may possibly be an effective treatment of MI in the lower limb musculature. PMID- 10714532 TI - Effects of diphtheria-tetanus-pertussis or tetanus vaccination on allergies and allergy-related respiratory symptoms among children and adolescents in the United States. AB - BACKGROUND: Findings from animal and human studies confirm that diphtheria and tetanus toxoids and pertussis (DTP) and tetanus vaccinations induce allergic responses; associations between childhood vaccinations and subsequent allergies have been reported recently. OBJECTIVE: The association of DTP or tetanus vaccination with allergies and allergy-related respiratory symptoms among children and adolescents in the United States was assessed. METHODS: Data were used from the Third National Health and Nutrition Examination Survey on infants aged 2 months through adolescents aged 16 years. DTP or tetanus vaccination, lifetime allergy history, and allergy symptoms in the past 12 months were based on parental or guardian recall. Logistic regression modeling was performed to estimate the effects of DTP or tetanus vaccination on each allergy. RESULTS: The odds of having a history of asthma was twice as great among vaccinated subjects than among unvaccinated subjects (adjusted odds ratio, 2.00; 95% confidence interval, 0.59 to 6.74). The odds of having had any allergy-related respiratory symptom in the past 12 months was 63% greater among vaccinated subjects than unvaccinated subjects (adjusted odds ratio, 1.63; 95% confidence interval, 1.05 to 2.54). The associations between vaccination and subsequent allergies and symptoms were greatest among children aged 5 through 10 years. CONCLUSIONS: DTP or tetanus vaccination appears to increase the risk of allergies and related respiratory symptoms in children and adolescents. Although it is unlikely that these results are entirely because of any sources of bias, the small number of unvaccinated subjects and the study design limit our ability to make firm causal inferences about the true magnitude of effect. PMID- 10714533 TI - A randomized controlled trial of chiropractic spinal manipulative therapy for migraine. AB - OBJECTIVE: To assess the efficacy of chiropractic spinal manipulative therapy (SMT) in the treatment of migraine. DESIGN: A randomized controlled trial of 6 months' duration. The trial consisted of 3 stages: 2 months of data collection (before treatment), 2 months of treatment, and a further 2 months of data collection (after treatment). Comparison of outcomes to the initial baseline factors was made at the end of the 6 months for both an SMT group and a control group. SETTING: Chiropractic Research Center of Macquarie University. PARTICIPANTS: One hundred twenty-seven volunteers between the ages of 10 and 70 years were recruited through media advertising. The diagnosis of migraine was made on the basis of the International Headache Society standard, with a minimum of at least one migraine per month. INTERVENTIONS: Two months of chiropractic SMT (diversified technique) at vertebral fixations determined by the practitioner (maximum of 16 treatments). MAIN OUTCOME MEASURES: Participants completed standard headache diaries during the entire trial noting the frequency, intensity (visual analogue score), duration, disability, associated symptoms, and use of medication for each migraine episode. RESULTS: The average response of the treatment group (n = 83) showed statistically significant improvement in migraine frequency (P < .005), duration (P < .01), disability (P < .05), and medication use (P< .001) when compared with the control group (n = 40). Four persons failed to complete the trial because of a variety of causes, including change in residence, a motor vehicle accident, and increased migraine frequency. Expressed in other terms, 22% of participants reported more than a 90% reduction of migraines as a consequence of the 2 months of SMT. Approximately 50% more participants reported significant improvement in the morbidity of each episode. CONCLUSION: The results of this study support previous results showing that some people report significant improvement in migraines after chiropractic SMT. A high percentage (>80%) of participants reported stress as a major factor for their migraines. It appears probable that chiropractic care has an effect on the physical conditions related to stress and that in these people the effects of the migraine are reduced. PMID- 10714534 TI - A combined approach for the treatment of cervical vertigo. AB - BACKGROUND: Cervical vertigo is a diagnosis commonly made at both otorhinolaryngologist and chiropractic offices. Hypothesized non-vascular mechanisms are reviewed. Therapeutic approaches have been suggested in the literature, ranging from cervical immobilization to vertebral manipulation. OBJECTIVE: To characterize the patient population with cervical vertigo and observe therapeutic results of a treatment protocol by using distinct conservative modalities. METHODS: Fifteen subjects with cervical vertigo were selected from patients presenting with dizziness at an otorhinolaryngology medical office. Diagnosis was based on specific criteria and results of an otoneurologic examination. All patients were submitted to a treatment protocol, including spinal manipulation, manual therapy on affected muscle groups, analgesic electrotherapy, labyrinth sedation, surface electromyography biofeedback, and an exercise program. Evolution of dizziness complaints and related musculoskeletal dysfunction was observed. RESULTS: Musculoskeletal complaints were present in 93% of the patients, mainly cervical pain, shoulder girdle pain, and tension-type headache. Median duration of musculoskeletal symptoms was 7.5 years, whereas the median duration of dizziness before the beginning of treatment was 52 days. Treatment duration averaged 5 sessions and 41 days. At the end of treatment, 60% of patients reported remission, 20% reported consistent improvement of vertigo. Remission of musculoskeletal symptoms was observed in 26.7% of patients, and improvement was observed in 60% of patients. CONCLUSION: Chronic, nontraumatic, cervical and shoulder-girdle dysfunction was an important causal and perpetuating factor of cervical vertigo in the population studied, and a consistent improvement was observed with the use of a conservative treatment protocol involving multiple modalities for patients with cervical vertigo. Further controlled studies are needed to access its validity. PMID- 10714535 TI - Reflex effects of vertebral subluxations: the peripheral nervous system. An update. AB - BACKGROUND: The traditional chiropractic vertebral subluxation hypothesis proposes that vertebral misalignment cause illness, disease, or both. This hypothesis remains controversial. OBJECTIVE: To briefly review and update experimental evidence concerning reflex effects of vertebral subluxations, particularly concerning peripheral nervous system responses to vertebral subluxations. DATA SOURCE: Information was obtained from chiropractic or scientific peer-reviewed literature concerning human or animal studies of neural responses to vertebral subluxation, vertebral displacement or movement, or both. CONCLUSION: Animal models suggest that vertebral displacements and putative vertebral subluxations may modulate activity in group I to IV afferent nerves. However, it is not clear whether these afferent nerves are modulated during normal day-to-day activities of living and, if so, what segmental or whole-body reflex effects they may have. PMID- 10714536 TI - Reflex effects of subluxation: the autonomic nervous system. AB - BACKGROUND: The collective experience of the chiropractic profession is that aberrant stimulation at a particular level of the spine may elicit a segmentally organized response, which may manifest itself in dysfunction within organs receiving autonomic innervation at that level. This experience is at odds with classic views of neuroscientists about the potential for somatic stimulation of spinal structures to affect visceral function. OBJECTIVE: To review recent findings from basic physiologic research about the effects of somatic stimulation of spinal structures on autonomic nervous system activity and the function of dependent organs. DATA SOURCE: Findings were drawn from a major recent review of the literature on the influences of somatic stimulation on autonomic function and from recent original physiologic studies concerning somatoautonomic and spinovisceral reflexes. CONCLUSIONS: Recent neuroscience research supports a neurophysiologic rationale for the concept that aberrant stimulation of spinal or paraspinal structures may lead to segmentally organized reflex responses of the autonomic nervous system, which in turn may alter visceral function. PMID- 10714537 TI - Mechanisms of neurovascular compression within the spinal and intervertebral canals. AB - OBJECTIVE: To describe some possible causes of encroachment on human spinal and intervertebral canal (foramen) neurovascular II structures. DATA SELECTION AND SYNTHESIS: A review of some imaging films of patients aged 38 to 52 years and some human autopsy histopathologic sections from 40- to 60-year-old cadavers to determine what structures may be responsible for neurovascular compression in individuals in this relatively young-to-middle-age group and to illustrate some examples. RESULTS: Stenosis of the spinal and intervertebral canal neurovascular structures can be caused by various bony and soft-tissue structures. Stenosis can be related to osteophytosis of the vertebral body, uncoverte-intervertebral disc protrusion, ossification of the posterior longitudinal ligament, and ligamentum flavum hypertrophy or buckling. DISCUSSION: Various forms of spinal and intervertebral canal stenosis can cause compression of neurovascular structures that may, in turn, be responsible for symptomatology. Of course, autopsy findings cannot be equated with painful syndromes in patients. PMID- 10714538 TI - Neurological effects of the adjustment. AB - This paper discusses the several theories pertaining to the chiropractic adjustment, including the nerve compression theory, reflex theories, and pain relief theories. There is now sufficient scientific research to consider these theories reasonable working models to explain the effects of the adjustment but insufficient to consider them valid. PMID- 10714539 TI - Motor control problems in patients with spinal pain: a new direction for therapeutic exercise. AB - Recent research into muscle dysfunction in patients with low back pain has led to discoveries of impairments in deep muscles of the trunk and back. These muscles have a functional role in enhancing spinal segmental support and control. The muscle impairments are not those of strength but rather problems in motor control. These findings call for a different approach in therapeutic exercise, namely a motor learning exercise protocol. The specific exercise approach has an initial focus on retraining the cocontraction of the deep muscles (ie, the transversus abdominis and lumbar multi-fidus Initial clinical trials point to the effectiveness of the approach in patients with both acute and chronic low back pain in terms of reducing the neuromuscular impairment and in control of pain. PMID- 10714540 TI - Economic case for the integration of chiropractic services into the health care system. AB - The role and position of chiropractic care in the health care system must be transformed from being alternative and separate to alternative and mainstream. This transformation requires that chiropractic services become integrated in the many health care delivery organizations that collectively constitute the health care system. There is solid and impressive economic and related justification for the desired integration. Chiropractic care is a cost-effective alternative to the management of neuromusculoskeletal conditions by other professions. It is also safer and increasingly accepted by the public, as reflected in the growing use and high patient retention rates. There is much and repeated evidence that patients prefer chiropractic care over other forms of care for the more common musculoskeletal conditions. The public interest will be well served by this transformation. Musculoskeletal disorders and injuries are the second and third most costly categories of health problems in economic burden-of-illness studies. They rank first as a cause in the prevalence of chronic health problems and long term disability and rank at the top for activity limitations and short-term disability. They rank first as a reason for consultation with a health professional and second as a reason for the use of prescription and nonprescription drugs. These conditions are more prevalent among the poor, lower middle income groups, and the elderly, yet those are precisely the groups that make the least use of chiropractic care for reasons of inadequate insurance coverage. The integration of chiropractic care into the health care system should serve to reduce health care costs, improve accessibility to needed care, and improve health outcomes. PMID- 10714541 TI - Public demand and the integration of complementary and alternative medicine in the US health care system. AB - Public use of complementary and alternative medicine (CAM) grew 25% between 1990 and 1997 and had a number of implications for chiropractic and the US health care system. Recent surveys describe the issues surrounding definitions of CAM; patterns of CAM use and its costs; attitudes of the public, health care providers and business entities; increasing scientific research; and changes in the health care system. Almost one third (192 million) of the 629 million visits to CAM providers in 1997 were to chiropractors. The new US National Center for Complementary and Alternative Medicine have funded chiropractic and other CAM research as a regular part of its scientific portfolio. Health maintenance organizations and other health care business entities have created new markets for CAM services, including chiropractic. profession, chiropractic appears to be positioned somewhere between mainstream practice and CAM, with conflicting opinions held by the public, the health care industry, and chiropractors themselves. The benefits and risks of chiropractic being identified with the CAM movement must be weighed carefully. PMID- 10714542 TI - Manipulation under joint anesthesia/analgesia: a proposed interdisciplinary treatment approach for recalcitrant spinal axis pain of synovial joint origin. AB - BACKGROUND: Manipulation under joint anesthesia/analgesia (MUJA) is an approach to treatment for patients with chronic, recalcitrant spinal axis pain of synovial joint origin. MUJA is the synthesis of fluoroscopically and corticosteroid agents with targeted, manual mobilizations and/or manipulations of the injected joint(s). DISCUSSION: MUJA should be viewed with guarded optimism because its success is based solely on anecdotal experience. Many physicians (specializing in targeted intraarticular "blocks" of spinal synovial joints) and chiropractors (specializing in manual mobilization and manipulation of spinal synovial joints) in the Tyler, Texas, area have treated more than 1000 patients over a 7-year period with the MUJA protocol. This protocol includes treatment of the atlanto occipital and lateral atantoaxial joints of the upper cervical spine, the zygapophysial joints of the cervical spine from C2-3 to C6-7, the thoracic spine and the lumbar spine, and the pelvic sacroiliac joints. CONCLUSION: The following patient types are suitable candidates for MUJA: patients with dominant spinal axis pain who have been unable to progress despite the passage of sufficient time (>2 months) and the delivery of prior treatments, including spinal manipulative therapy; patients with pain so severe that standard manipulative therapy cannot be delivered with technical success; and patients with complex problems in whom the diagnosis of synovial joint-mediated spinal pain must be established before the safe delivery of manipulative therapy. PMID- 10714543 TI - Integration of chiropractic education into a hospital setting: A South African experiences. AB - This article examines differences between chiropractic and medical internship experiences, both internationally and in South Africa. The South African hospital experience is described, and the future is discussed. PMID- 10714544 TI - Qualitative review of studies of manipulation-induced hypoalgesia. AB - BACKGROUND: The number of studies that have investigated the direct analgesic effect of a spinal manipulation on spinal or referred pain is small, making knowledge of this crucial aspect of manipulation sparse. This paper reviews a set of studies that measure the immediate effect of manipulation on pain or pain related phenomena in the spinal and peripheral soft tissues. METHODS: The literature was accessed through MEDLINE. Key words used were "manipulation," "pain," and "chiropractic." This search was complemented by citation reviews of important research and chapters on the topic. Only studies that directly measured the effect of at least a single spinal manipulation on pain (eg, tenderness, biochemical assay, referred pain) were selected. The selected studies were reviewed descriptively; no systematic assessment of their quality was conducted. RESULTS: The electronic search yielded 738 citations. Six hundred and forty-two were relevant to chiropractic. Of these, most were clinically descriptive articles about diagnostic and therapeutic procedures or case management. Most of the remaining articles were clinical trial reports or letters to the editor. Only 5 studies were selected according to the established criteria. Thus less than 1% of the indexed literature on chiropractic, manipulation, and pain involved studies that explored the mechanism of the putative effect of spinal manipulation on pain mechanisms. Six other studies were retrieved from citation reviews. These 11 studies were reviewed in order publication. CONCLUSION: Few studies have investigated the effects of spinal manipulation on pain directly. If the theory of manipulation exerting its therapeutic effects posits that the sensory input created by the intervention results in some form of inhibition of pain, then the results of these studies are largely consistent with one another and with this theory. This review has highlighted the deficiencies in the extant studies and many remaining questions. Only more high-quality research will permit a full elucidation of the hypoalgesic effects of spinal manipulation. PMID- 10714545 TI - Integration of traditional and complementary medicine into national health care systems. AB - The growth of complementary and alternative medicine in the Far East is discussed. The role of traditional medicine in the development of health care in Asia is presented, and integration of traditional and modern methods of health care are described. PMID- 10714546 TI - Effects of interleukin-15 on in vitro human T cell proliferation and activation. AB - Interleukin-15 (IL-15) has been reported to have many activities on T cell populations, including a potential role in improving antigen-specific proliferation in HIV-1 disease. We tested this response in healthy adults by studying the response of T cell populations after stimulation with medium, tetanus, cytomegalovirus (CMV) antigens in cultures from 21 volunteers. IL-15 caused a dose-dependent increase in medium and antigen-induced proliferation. The expansion was due to CD8>natural killer (NK)>CD4 lymphocytes and memory > naive cells. The IL-15-stimulated CD8 cells had increased levels of the activation markers CD69 and DR. The published CMV-induced expression of CD57 on CD8+ cells was increased in CMV seronegative and seropositive subjects by IL-15. IL-15 appears to be a stimulator of T cell populations in healthy adults and may be useful in settings to enhance nonspecific NK activity or antigen-specific CD8 activity. PMID- 10714547 TI - Interleukin-4 and IL-10 bind covalently to activated human alpha2-macroglobulin by a mechanism that requires Cys949. AB - Alpha2-macroglobulin (alpha2M) functions as an extracellular carrier of diverse cytokines, including transforming growth factor-beta1 (TGF-beta1), that expresses anti-inflammatory activities. The results presented here demonstrate that interleukin-10 (IL-10) and IL-4, which also regulate the inflammatory response, bind to alpha2M. Unlike TGF-beta, IL-4 and IL-10 bind almost exclusively to the receptor-recognized, or activated, form of alpha2M. Purified IL-4-alpha2M complexes were predominantly covalent due to thiol disulfide exchange involving Cys949 in the alpha2M subunit. Blocking Cys949 with iodoacetamide significantly inhibited IL-4- and IL-10 binding. Bovine serum albumin (BSA), which possesses a free Cys residue and undergoes thiol disulfide exchange reactions, did not compete with alpha2M for the binding of IL-4 or IL-10. These results suggest a model in which IL-4 and IL-10 associate with activated alpha2M to form complexes that are initially noncovalent but unstable. In these complexes, Cys949 is properly aligned to undergo thiol disulfide exchange and generate stable, covalent IL-4-alpha2M and IL-10-alpha2M complexes. PMID- 10714548 TI - Analysis of transcription factors regulating induction of indoleamine 2,3 dioxygenase by IFN-gamma. AB - IFN-gamma treatment of the human carcinoma cell line ME180 causes cell death due to induction of indoleamine 2,3-dioxygenase (IDO) and resulting starvation for tryptophan. A mutant cell line 3B6A derived from ME180 was resistant to IFN-gamma because of loss of IDO activity. Cotransfecting an IDO promoter-chloramphenicol acetyl transferase (CAT) construct with IFN regulatory factor-1 (IRF-1) resulted in induction of CAT activity in both ME180 and 3B6A cells even in the absence of IFN-gamma. This induction was reduced by cotransfection with IRF-2. However, IRF 1 was not able to restore IDO activity, suggesting a possible repressor site outside the IDO promoter region. Stat1alpha (p91) restored both CAT and IDO activities in 3B6A cells following IFN-gamma treatment. 3B6A cells doubly treated with IFN-gamma and IFN-alpha or IFN-beta restored IDO activity, although neither cytokine on its own could induce IDO. Western blot analysis showed that both constitutive expression and induction of Stat1alpha by IFN-gamma were reduced in 3B6A cells, and double treatment of IFN-gamma with IFN-alpha or IFN-beta restored the expression level of Statla. Electrophoretic mobility shift assays indicated that Stat1 binds to the IFN-gamma-activated sequence (GAS) region in the IDO promoter in ME180 cells following IFN-gamma treatment. Our results indicated that the defect in 3B6A cells was reduced expression of Stat1alpha and that IRF-1, NF kappaB, and PKR were all involved to some extent in the induction of IDO following IFN-gamma treatment. PMID- 10714549 TI - Transforming growth factor-beta mRNA and protein in hypertrophic scar tissues and fibroblasts: antagonism by IFN-alpha and IFN-gamma in vitro and in vivo. AB - Hypertrophic scarring (HSc) following burn injury is a common, disfiguring, and functionally limiting form of dermal fibrosis, compromising recovery. Previously, elevated levels of transforming growth factor-beta1 (TGF-beta1), a fibrogenic cytokine, were found in wounds and serum of severely injured patients, antagonized in part by treatment with systemic interferon-alpha2b (IFN-alpha2b) both in vitro and in vivo. It is hypothesized that in wound healing after injury, platelets are an initial source of TGF-beta, but wound fibroblasts may be capable, after activation, of autoamplification of the initial response to injury by increasing TGF-beta mRNA and protein that may subsequently be responsive to IFN therapy with IFN-alpha or IFN-gamma or both. Using three pairs of site matched HSc and normal fibroblasts from the same individuals, nonconfluent and near confluent fibroblasts were treated with TGF-beta, and cell proliferation and collagen production were assayed using cell counting and 18O2 isotopic uptake into hydroxyproline before analysis by gas chromatography-mass spectrometry (GC MS). HSc and normal fibroblasts were assayed for the production of TGF-beta protein secretion using ELISA for TGF-beta1, TGF-beta2, and TGF-beta3 after acidification of medium samples from 96-h cultures. HSc and normal fibroblasts were treated with IFN-alpha2b or IFN-gamma or both for 96 h. Quantitative RT-PCR and Northern analysis were performed using newly synthesized internal standards for human TGF-beta1. TGF-beta stimulates both HSc and normal fibroblast proliferation. Collagen synthesis is greater in HSc than in normal fibroblasts and is maximally stimulated at 75 pM TGF-beta. TGF-beta stimulated collagen metabolism is antagonized by IFN-alpha or IFN-gamma or both in an additive fashion. HSc and normal fibroblasts not only possess the mRNA for TGF-beta1 but also secrete mature TGF-beta protein. Treatment of HSc and normal fibroblasts with IFN-alpha2b or IFN-gamma antagonizes TGF-beta protein production, and additive effects occur. RT-PCR demonstrates that after IFN treatment, downregulation of TGF-beta1 mRNA accounts in part for the reduction in protein secretion in HSc fibroblasts. Elevations of systemic TGF-beta may be due to wound fibroblasts. TGF-beta synthesis and antagonism of fibroblast TGF-beta protein secretion occurs with either IFN-alpha or IFN-gamma, in part by downregulation of TGF-beta1 mRNA levels. PMID- 10714550 TI - Pharmacologic effect of recombinant human IFN-alpha, continuously released from a matrix prepared from a polyglycerol ester of fatty acids, on 2',5'-oligoadenylate synthetase activity in murine liver. AB - The objective of this study was to assess the pharmacologic effect of continuously released recombinant human interferon-alpha (rHuIFN-alpha) in the liver, the target organ of chronic hepatitis B and C, using 2',5'-oligoadenylate synthetase (2',5'-OAS) activity as an indicator of an antiviral state. A cylindrical matrix prepared from tetraglycerol dipalmitate (TGDP), a polyglycerol ester of fatty acids (PGEF), released rHuIFN-alpha in a pseudo-zero-order manner for about 1 week after implantation into mice, without any major loss of rHuIFN alpha biologic activity during the release period. To evaluate the pharmacologic effect of the rHuIFN-alpha continuously released from this type of matrix, we established a murine test system. Bolus injections of rHuIFN-alpha solution at three doses increased 2',5'-OAS activities in murine liver extract and serum in a dose-dependent manner, indicating that this system is suitable for evaluating rHuIFN-alpha activity. After subcutaneous insertion of TGDP-matrix implants containing 5.5x10(7) IU rHuIFN-alpha per animal, 2',5'-OAS activities in both liver extracts and serum increased rapidly and remained high for over 1 week. Subcutaneous injections of an equivalent total dose (5.0x10(7) IU/animal per week) of rHuIFN-alpha solution in three or seven fractions prolonged 2',5'-OAS activities compared with a single bolus injection. Comparing 2',5'-OAS activity on day 7 and the portion of the area under the 2',5'-OAS activity-time curve above the normal level (deltaAUC) between the TGDP-matrix implant and multiple injections of the solution revealed that continuously released rHuIFN-alpha has an effect almost equivalent to that of three or seven injections of the solution per week. PMID- 10714551 TI - Turkey and chicken interleukin-2 cross-react in in vitro proliferation assays despite limited amino acid sequence identity. AB - We cloned the cDNA of turkey interleukin-2 (IL-2), initially using oligonucleotide primers based on the sequence of the chicken IL-2 gene. Compared with the only other cytokines available for comparison, the interferons (IFN), the coding regions of the turkey and chicken IL-2 genes are much less conserved (86.24% nucleotide identical and 69.93% amino acid identical). The lack of nucleotide conservation was spread across the entire length of the coding region. In comparison, the promoters of the two avian IL-2 genes shared a high degree of identity (95.71% identical over 380 nucleotides). Phylogenetic analysis shows that turkey and chicken IL-2 have diverged to a greater extent than IL-2 from closely related mammalian species. Surprisingly, considering the low level of amino acid identity, including residues known to be important in binding the IL-2 receptor in mammalian species, both turkey and chicken IL-2 cross-react in functional assays. PMID- 10714552 TI - Randomized trial of sequential administration of G-CSF and GM-CSF vs. G-CSF alone following peripheral blood progenitor cell autograft in solid tumors. AB - A trial was conducted to investigate whether the sequential administration of recombinant human granulocyte colony-stimulating factor (G-CSF) and recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF) could accelerate reconstitution of hematopoiesis, compared with G-CSF alone following high-dose chemotherapy (HDCT). A group of 34 consecutive patients with solid tumors undergoing HDCT and autologous peripheral blood progenitor cell (PBPC) transplantation was studied. Conditioning regimen included carboplatin, etoposide, mitoxantrone, and melphalan for breast cancer and cyclophosphamide or ifosfamide, carboplatin, and etoposide for the other tumors. HDCT was delivered from day -3 to day -1. PBPC were infused on day 0, and on the same day growth factors were administered subcutaneously (s.c.) 5 microg/kg each. Seventeen patients were randomized to receive G-CSF from day 0 to day 13 after HDCT (arm A), and 17 patients received G-CSF from day 0 to day 6 and GM-CSF from day 7 to day 13 (arm B). Patients were stratified, and their characteristics were homogeneous in both arms for age, performance status, and number of previous chemotherapy courses and CD34+ infused. The median time to absolute neutrophil count (ANC) >500/microl was 10 days in arm A and 9 days in arm B (p = 0.96). Days to platelet (PLT) count >20,000 were not different in the two treatment arms (p = 0.1), but patients randomized to arm A had a lower platelet count compared with patients in arm B. One month after PBPC transplantation, a statistically significant difference in PLT count was observed (arm A median 150x10(3)/microl (90-310), arm B median 254x10(3)/microl (117-387),p = 0.0013). The days patients had fever >38 degrees C were 39 in arm A and 26 in arm B (p = 0.18). The difference in the length of hospital stay was not statistically significant between the groups (Mann-Whitney sum rank test). After a median follow-up of 30 months, 21 patients were alive and 20 were disease free. These data show that the two growth factors are associated with different patterns of hematopoietic recovery, and larger randomized trials in groups of more homogeneous patients will be needed to define the effects and benefits of combination growth factor therapies. PMID- 10714553 TI - A comparative analysis of the antigenic, structural, and functional properties of three different preparations of recombinant human interleukin-18. AB - We compared the structural and functional properties of three recombinant human interleukin-18 (rIL-18) preparations, commercially available (Pep rIL-18) and prepared in our laboratory (active and inactive, according to their ability to potentiate IL-12-mediated interferon-gamma [IFN-gamma] induction in lymphocytes). All three preparations showed multimer formation on SDS-PAGE/immunoblotting using monoclonal antibodies (mAb) against the inactive form of rIL-18. In contrast, only the 18-kDa bands were recognized in each sample by mAb against the active form of rIL-18. The amounts of multimers and the 18-kDa moiety of Pep rIL-18 resembled those of the inactive rather than the active form. Likewise, the reaction profile of Pep rIL-18 toward mAb was very similar to that of inactive but not active rIL-18 on sandwich ELISA. Pep rIL-18 potentiated IFN-gamma inducing activity together with IL-12, but its potency was 100-fold less than that of the active rIL-18, and excess doses were required for its activity. The inactive rIL-18 showed virtually no IFN-gamma-inducing ability, but when reduced and reconstituted, it inhibited the IFN-gamma-inducing activity of active rIL-18. These results suggest that there are two categories of recombinant IL-18 that are structurally, functionally, and antigenically different, and the mAb 125-2H and 21 can discriminate these two IL-18 populations by recognizing the epitopes specifically expressed on active and inactive IL-18, respectively. PMID- 10714554 TI - A CA repeat polymorphism of the IFN-gamma gene is associated with susceptibility to type 1 diabetes. AB - Recent studies have shown that loci outside the HLA region are involved in determining susceptibility to type 1 diabetes. Polymorphisms in the coding and noncoding regions of the genes encoding cytokines may be involved in modulating the immune response to self and nonself antigens. There is increasing evidence that an imbalance and disruption of the Thl and Th2 T cell subsets play a key role in the development of experimental and clinical type 1 diabetes. The aim of this study was to investigate the frequency of a CA dinucleotide repeat polymorphism in the interferon-gamma (IFN-gamma) gene (IFNG) and a C(-590)T polymorphism of the interleukin-4 (IL-4) gene in 236 Caucasoid patients with type 1 diabetes. There was a highly significant increase in the 3/3 IFNG genotype in the patients compared with normal healthy controls (34.3% vs. 13.5%, p<0.0001) as well as a significant increase in allele 3 of the IFNG locus in the patients compared with controls (51.9% vs. 31.7%, p<0.00001). In contrast, no significant differences were found in the frequency of the C(-590)T IL-4 polymorphism between patients and controls. These results suggest that polymorphisms of the IFNG gene may modify the function of this proinflammatory mediator and the response to pancreatic islet beta cells. PMID- 10714555 TI - Transtracheal administration of interleukin-12 induces neutrophil responses in the murine lung. AB - Although the roles of interleukin-12 (IL-12) in the immunomodulation of antigen specific responses are well characterized, the effects of IL-12 on the respiratory tract following mucosal administration are not well defined. Therefore, we investigated changes in the murine lung shortly after intranasal (i.n.) administration of murine IL-12. We showed that IL-12 induced neutrophil influx to the murine lung in both C57BL/6 and BALB/c mice. Histologic examination revealed that intranasal administration of IL-12 with liposomes induced focal neutrophil infiltration into the alveoli and a significant increase in neutrophils in bronchoalveolar lavage fluids when compared with administration of liposomes alone. In vitro chemotaxis assays indicated that the observed pulmonary neutrophil response induced by IL-12 could have been due in part to the direct chemotactic activity of IL-12 for murine neutrophils. PMID- 10714556 TI - Partial IFN-alpha/beta and IFN-gamma receptor knockout trisomy 16 mouse fetuses show improved growth and cultured neuron viability. AB - The trisomy 16 mouse fetus is a well-studied model for Down syndrome (trisomy 21), the leading genetic cause of mental retardation in the newborn population. Human chromosome 21 and mouse chromosome 16 each carry a large cluster of genes that code for components of the interferon (IFN)-alpha/beta and IFN-gamma receptors, and Down syndrome cells display significantly increased sensitivity to IFN action. We have previously reported that in utero anti-IFN IgG treatment of mice pregnant with trisomy 16 fetuses results in a significant improvement in trisomy 16 fetus growth and morphology and that anti-IFN-gamma IgG treatment can prevent the premature death of trisomy 16 fetal mouse cortical neurons in culture. We have now used IFN receptor subunit knockout mice to produce mouse fetuses that carry three No. 16 chromosomes and one copy each of disabled IFN gamma receptor (IFNGR) and IFN-alpha/beta receptor (IFNAR-2) component genes. We report here that this partial IFN receptor knockout trisomy (PIRKOT) mouse fetus has significantly improved growth and yields cortical neurons whose viability is the equivalent of that seen in their euploid counterparts. PMID- 10714557 TI - Early expression of IFN-alpha/beta and iNOS in the brains of Venezuelan equine encephalitis virus-infected mice. AB - To investigate the roles of type I interferon (IFN-alpha/beta) and other mediators of innate immune responses (e.g., inducible nitric oxide synthase [iNOS]) in early dissemination of Venezuelan equine encephalitis virus (VEE) infection, we used mice with targeted deletions in either their IFN-alpha/beta receptor (IFNAR-1-/-) or interferon regulatory factor 2 (IRF-2-/-) genes. Following footpad infection, both IFNAR-1-/- and IRF-2-/- mice were more susceptible than control mice to VEE. The IFNAR-1-/- mice also exhibit accelerated VEE dissemination to serum, spleen, and brain, and compared with control mice, they evidenced faster kinetics in the upregulation of proinflammatory genes. In contrast, in IRF-2-/- mice, iNOS gene induction was completely absent following peripheral virulent VEE infection. In evaluating the role of cells involved in iNOS production, primary microglial cell cultures were found to be highly permissive to VEE infection. Moreover, VEE infection increased levels of nitric oxide (NO) in resting microglial cultures but decreased NO production in IFN-gamma-stimulated microglia. Thus, these findings suggest that reactive nitrogen species play an important contributory role in VEE dissemination and survival of the host. Our results further suggest the necessity for a carefully balanced host response that follows a middle course between immunopathology and insufficient inflammatory response to VEE infection. PMID- 10714558 TI - Interleukin-18 in combination with IL-2 enhances natural killer cell activity without inducing large amounts of IFN-gamma in vivo. AB - Interleukin-18 (IL-18) is known to synergistically enhance murine natural killer (NK) cell activity in vitro when combined with either IL-12 or IL-2. However, it has also been demonstrated that simultaneous administration of IL-18 and IL-12 to mice induces extraordinarily large amounts of interferon-gamma (IFN-gamma) in the serum. In this study, we examined the effects of a combination of IL-18 and IL-2 on in vivo NK cell activation in parallel with the induction of toxicity. In contrast to the IL-18 and IL-12 combination, the combined administration of IL-18 and IL-2 to BALB/c mice for 3 days induced neither high levels of IFN-gamma production nor other visible side effects. When compared with treatment with IL 18 or IL-2 alone, the combined treatment resulted in a significant increase in the number of DX-5 (pan-NK cells marker)-positive cells in spleens and a marked enhancement of splenic NK activity, as determined by standard cytotoxicity assays. Enhanced splenic cytotoxicity generated in the mice treated with both IL 18 and IL-2 was also observed against syngeneic Colon 26 adenocarcinoma cells. Consistent with this in vitro observation, combined treatment produced a significantly stronger inhibitory effect on the pulmonary metastases following i.v. injection of Colon 26 tumor cells than treatment with either cytokine alone. These results suggest that IL-18 combined with IL-2 potentiates in vivo NK cell activity and contributes to inhibition of tumor metastasis without inducing significant toxicity. PMID- 10714559 TI - Acute effects of interferon on estrogen receptor function do not involve the extracellular signal-regulated kinases p42mapk and p44mapk. AB - Exposure to type I interferons (IFN) increased estrogen receptor (ER) ligand binding and induced protein kinase C (PKC) translocation within 30 min but had no effect on net incorporation of [32P] into ER in Madin Darby bovine kidney (MDBK) cells. Ligand binding was also increased within 30 min by phorbol ester and the protein phosphatase inhibitor okadaic acid. Mitogen-activated protein (MAP) kinase phosphorylation was initially inhibited between 2 and 30 min and subsequently activated between 30 and 60 min after treatment with IFN. The activatory response was blocked by the PKC inhibitor Ro 31-8220. Following transient transfection with an ERE-CAT reporter construct, IFN increased CAT expression after 6 h but decreased ER ligand binding, transcriptional activity and phosphorylation after 48 h, probably as a result of decreased ER concentrations. The results rule out rapid activation of ER ligand binding through phosphorylation at Ser118 by MAP kinase because (1) the increase in ligand binding preceded activation of MAP kinase, and (2) IFN had no short-term effect on [32P]incorporation or ER transcriptional activity. The rapid effect of IFN on ER ligand binding is postulated to reflect phosphorylation of the receptor at Tyr537 by p56lck, a member of the Src family of PKC-activated tyrosine kinases. PMID- 10714560 TI - Corrective effects of interleukin-12 on age-related deficiencies in IFN-gamma production and IL-12Rbeta2 expression in virus-specific CD8+ T cells. AB - Interleukin-12 receptor beta2 (IL-12Rbeta2) has been shown to be selectively expressed on Th1 T cell subsets, and we have previously shown that influenza specific CD8+ cytotoxic T lymphocyte (CTL) deficiency in old mice was associated with deficient Th1 (interferon-gamma [IFN-gamma]) cytokine production. This study tested whether IL-12Rbeta2 expression was also deficient in CD8+ CTL from old mice and the effect of IL-12 treatment on these responses. Splenic lymphocytes from influenza-primed old and young BALB/c mice were stimulated with influenza virus in vitro with and without IL-12 and then enriched for CD8+ T cells. IFN gamma was significantly reduced, whereas IL-4 and IL-12p40 (an antagonist of IL 12 function) were evaluated in old when compared with young mice. This was true for secreted protein measured by ELISA and for mRNA levels quantitated by RT-PCR. IL-12Rbeta2 mRNA expression in CD8+ CTL was also significantly reduced in old mice. IL-12 treatment in vitro caused significant upregulation of IFN-gamma and IL-12Rbeta2 and downregulation of IL-4 in CD8+ T cells from old mice and young mice. The present demonstration of an age-related downregulation in IL-12Rbeta2 expression and our previous data showing reduced IFN-gamma and elevated IL-4 production provide strong evidence that CD8+ CTL deficiency in aging results from a Th1/Th2 cytokine production switch. Agents that increase IL-12Rbeta2 expression and redirect Th2 to Thl immune responses are likely to enhance CD8+ CTL-mediated control of viral infections in aging. PMID- 10714561 TI - Effects of 2-deoxy-D-glucose administration on cytokine production in BDF1 mice. AB - Physical exercise and diet changes have been shown to affect immune parameters, and similar effects are also induced by the administration of a nonmetabolizable glucose analog, 2-deoxy-D-glucose (2-DG). The present study was designed to characterize the effects of glucoprivation induced by 2-DG administration on concentrations of tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL 1beta), and IL-6 in the blood and interferon-gamma (IFN-gamma), IL-2, and IL-4 in vitro production by partially purified T splenocytes in BDF1 mice. Mice (n = 8 per group) were injected intraperitoneally one or three times with 0, 500, 750, or 1000 mg/kg of 2-DG, and blood and spleens were collected 2 h after the last injection. Partially purified T splenocytes were cultured 24 h in the presence of concanavalin A (ConA). A significant increase in the corticosterone levels with the amount of 2-DG injected was observed after one or three injections (p<0.05). The amount of 2-DG injected was associated with an increase in TNF-alpha, IL 1beta, and IL-6 concentrations in the blood of mice after one or three injections of 2-DG (p<0.05). A significant decrease in in vitro proliferation of partially purified splenocytes in the presence of ConA was associated with a decrease in IFN-gamma production in the culture supernatants and an increase in IL-1 receptor expression on the cell surface (p<0.05). PMID- 10714562 TI - Immunogenicity of a novel DNA vaccine cassette expressing multiple human immunodeficiency virus (HIV-1) accessory genes. AB - OBJECTIVE: To develop an HIV-1 accessory gene immunogen using a DNA vaccine approach. METHODS: HIV-1 accessory genes vif, vpu and nef were modified to express under the control of a single promoter with cellular proteolytic cleavage sites between the coding sequences (VVN-P). Immune responses induced by these constructs were evaluated in mice. RESULTS: DNA vaccine construct (pVVN-P) expressing Vif, Vpu and Nef was processed and the fusion protein was cleaved appropriately. Vif, Vpu and Nef as a fusion protein with proteolytic cleavage sites (VVN-P) is able to induce a significant level of cellular immune responses. We also observed that accessory genes Vif, Vpu and Nef (VVN-P) induced an effective T helper 1 proliferative response measured by cytokine production. Furthermore, expression cassette pVVN-P was able to induce cytotoxic T lymphocyte (CTL) responses against diverse HIV-1 viruses in infected target cells. CONCLUSION: We conclude that cell-mediated immune responses induced by accessory gene constructs from clade B may have a broader recognition of divergent HIV-1 viruses and should be further examined for both prophylactic and therapeutic vaccination schemes against HIV-1. PMID- 10714563 TI - Response to immunization with recall and neoantigens after prolonged administration of an HIV-1 protease inhibitor-containing regimen. ACTG 375 team. AIDS Clinical Trials Group. AB - OBJECTIVES: To ascertain if immunization results in the restoration of responses to recall antigens, in the development of responses to presumed neoantigens, and to identify the virologic and immunologic correlates of these responses in persons with HIV-1 infection. DESIGN AND SETTING: Open-label study carried out at three university-affiliated AIDS Clinical Trials Units in the United States. SUBJECTS AND METHODS: Thirty-one subjects participating in AIDS Clinical Trials Group Protocol 375 who had received zidovudine, lamivudine, and ritonavir for at least 48 weeks. Subjects were immunized with tetanus toxoid (TT) at entry and with inactivated hepatitis A vaccine (hep A) and keyhole limpet hemocyanin (KLH) at entry and 6 weeks. The development of antibody, lymphocyte proliferative assay (LPA), and delayed-type hypersensitivity (DTH) responses after immunization were monitored. RESULTS: The LPA and DTH responses to TT improved in 57 and 68% of participants, respectively; 73 and 65% developed enhanced LPA and DTH responses to KLH. Forty-eight percent of patients developed a four-fold increase in antibody concentration to tetanus. Seventy-three percent of patients without detectable hepatitis A antibodies at baseline developed antibodies after immunization. Eighty-three percent of patients experienced at least a four-fold rise in KLH antibody concentration. Immune activation and viral load predicted poor recall responses and the number of memory CD4+ T-cells predicted good responses to recall antigens. Naive CD4+ T-cell numbers, decrease in viral load, increases in CD4+ and CD28+ cells, and decreases in immune activation were associated with responses to presumed neoantigens. CONCLUSIONS: Most HIV-infected patients treated with potent combination antiretrovirals develop responses to recall and presumed neoantigens after immunization. Functional immune restoration in response to immunization is related to control of viral replication, decreased immune activation as well as to both quantitative and qualitative restoration of circulating T- lymphocyte subpopulations. PMID- 10714564 TI - Cellular proviral HIV-DNA decline and viral isolation in naive subjects with <5000 copies/ml of HIV-RNA and >500 x 10(6)/l CD4 cells treated with highly active antiretroviral therapy. AB - OBJECTIVE: To evaluate the decay rate of cellular proviral HIV-DNA and viral replication in patients receiving highly active antiretroviral therapy (HAART) in the very early phase of infection. METHODS: Thirty-four patients treated with HAART and retrospectively selected for progressive decline of plasma viraemia up to undetectable levels (< 20 copies/ml), were stratified according to CD4+ cell count and plasma viraemia at base line: > 500 x 10(6) cells/l with < 5000 copies/ml (group 1) or with > 5000 copies/ml (group 2), > 5000 copies/ml with 300 500 x 10(6) cells/l (group 3) or with < 300 x 10(6) cells/l (group 4). Plasma HIV RNA and proviral HIV-DNA were analysed at baseline and after 1, 2, 3, 6, 9 and 12 months of treatment. RESULTS: After 1 year of treatment, a significant decrease of proviral DNA titre was observed in all patients and a decrease > 1 log was achieved in 24 of 29 subjects of the first three groups. The more pronounced decay of HIV-DNA (half-life 28 weeks) up to < 50 HIV-DNA copies/10(6) CD4+ cells was detected in patients of group 1. At the year's endpoint, five patients (four in group 1 and one in group 2) had < 20 HIV-DNA copies. However, HIV strains sensitive to antiretroviral drugs were isolated from peripheral lymphocytes of 16 out of 34 patients. CONCLUSION: In patients with undetectable plasma viraemia after 1 year of HAART, the highest reduction of proviral DNA up to < 50 copies/10(6) CD4+ cells was obtained only in subjects in the early asymptomatic phase of infection. Nevertheless, a replication-competent virus can be detected in all phases of antiretroviral therapy. PMID- 10714565 TI - Multiple sites in HIV-1 reverse transcriptase associated with virological response to combination therapy. AB - OBJECTIVE: To determine whether analysis of sequence variation in reverse transcriptase at baseline can explain differences in response to combination antiretroviral therapy. METHODS: Amino acid sequences of reverse transcriptase obtained from baseline isolates from 55 patients included in a trial of zidovudine and didanosine versus zidovudine/didanosine/nevirapine (ACTG241) were analysed. Simple and multiple linear regression were used to determine the relationship between numbers and identity of mutations at baseline and virological response after 8 and 48 weeks. RESULTS: Numbers of baseline zidovudine resistance mutations were predictive of short-term response (week 8). Amino acid identity at position 215 explained > 20% of the variation in response at week 8, but less at week 48. Multiple regression identified the combinations: 215 + 44 and 41 + 202, each of which explained about 30% of the variation in week 8 response. A model incorporating amino acids 214 + 215 + 60 + 202 + baseline viral load explained > 40% of the variation in response at week 48. Unexpectedly, the mutant combination 601 + 215Y/F responded threefold better than 60V + 215Y/F over 48 weeks. CONCLUSIONS: Use of clinical data to analyse virological response to combination therapy has revealed effects of baseline amino acid mutations at sites not previously identified as being important in antiretroviral resistance. Predictors of long-term responses were different from those involved in the short term and may require more complex analysis. PMID- 10714566 TI - Fat distribution evaluated by computed tomography and metabolic abnormalities in patients undergoing antiretroviral therapy: preliminary results of the LIPOCO study. AB - BACKGROUND: Fat distribution abnormalities have been reported in patients treated with various antiretroviral drug regimens. The LIPOCO study is an ongoing observational study of unselected HIV-infected patients which aims to better characterize such disorders and their metabolic correlations. METHODS: Cross sectional analysis of data collected at baseline in the first 154 male patients included. Investigators divided patients into four predetermined clinical categories of fat distribution: lipoatrophy, obesity, mixed condition and normal. Body composition (tetrapolar bioelectrical impedance analysis and skinfold thickness), fat distribution [computed tomography (CT) scan], plasma glucose and insulin concentrations both fasting and during an oral glucose tolerance test and endocrine and lipid profile were measured and compared between the four groups. RESULTS: Patients in the lipoatrophy group had significantly decreased abdominal and mid-thigh subcutaneous fat area values and elevated levels of plasma triglycerides. Patients in the obese and mixed groups had significantly increased intra-abdominal fat area values and elevated levels of plasma insulin and C peptide. The CT scans identified some patients with isolated subcutaneous fat accumulation but no other alterations in fat distribution and no insulin resistance. Visceral adipose tissue measured by CT scan was positively correlated with fasting insulin and the sum of insulin levels (P < 0.0001). Fasting insulin as well as the sum of insulin levels were negatively correlated with the delta HIV-RNA (log(10)). In a multivariate logistic regression model, the use of stavudine significantly correlated with fat wasting in both nucleoside reverse transcriptase inhibitor and protease inhibitor groups: odds ratio (OR), 413 [95% confidence interval (CI), 5.2-999; P = 0.0068] and OR, 2.08 (95% CI, 0.92-7.0; P = 0.058) respectively, when compared with the use of zidovudine. Neither lamivudine or didanosine use, nor the use of protease inhibitors were significantly associated with fat distribution abnormalities or fat wasting. CONCLUSIONS: These preliminary results suggest that three major types of fat distribution abnormalities may occur in isolation or in association in HIV infected patients undergoing active antiretroviral therapy: a fat depletion or 'lipoatrophy' syndrome which might be related to the use of stavudine; a mixed or fat redistribution syndrome related to an unusual side-product of effective virus control; and a subcutaneous adiposity syndrome reflecting increase in caloric intake. PMID- 10714567 TI - Effect of ritonavir on lipids and post-heparin lipase activities in normal subjects. AB - BACKGROUND: Intensive therapy of HIV infection with highly active antiretroviral therapy (HAART) dramatically reduces viral loads and improves immune status. Abnormalities of lipid levels, body fat distribution, and insulin resistance have been commonly reported after starting HAART. Whether the lipid abnormalities result from changes in metabolism after an improvement in HIV status or are partly attributable to the effects of protease inhibitor use is unknown. METHODS: Twenty-one healthy volunteers participated in a 2 week double-blind, placebo controlled study on the effect of the protease inhibitor ritonavir on total lipids, apolipoproteins, and post-heparin plasma lipase activities. RESULTS: Those taking ritonavir (n = 11) had significantly higher levels of plasma triglyceride, VLDL cholesterol, IDL cholesterol, apolipoprotein B, and lipoprotein (a) compared with placebo (n = 8). HDL cholesterol was lower with therapy as a result of a reduction in HDL3 cholesterol. Post-heparin lipoprotein lipase (LpL) activity did not change but hepatic lipase activity decreased 20% (P < 0.01) in those taking ritonavir-compared with placebo. Although all lipoprotein subfractions became triglyceride enriched, most of the increase in triglyceride was in VLDL and not in IDL particles. CONCLUSION: Treatment with ritonavir in the absence of HIV infection or changes in body composition results in hypertriglyceridemia that is apparently not mediated by impaired LpL activity or the defective removal of remnant lipoproteins, but could be caused by enhanced formation of VLDL. Long-term studies of patients with HIV infection receiving HAART will be necessary to determine the impact of these drugs and associated dyslipidemia on the risk of coronary artery disease. PMID- 10714568 TI - Toxicity and drug exposure in a quadruple drug regimen in HIV-1 infected patients participating in the ADAM study. AB - OBJECTIVE: To study the relationship between toxicity and the exposure to nelfinavir and saquinavir as part of a quadruple drug regimen. DESIGN: The ADAM study is a randomized study to investigate the feasibility of induction maintenance therapy in HIV-1 infection. METHODS: HIV-1-infected patients with no prior use of antiretroviral treatment started induction therapy consisting of stavudine + lamivudine + nelfinavir + saquinavir for a period of 26 weeks. Data regarding toxicity of the quadruple regimen and exposure to the protease inhibitors were collected. RESULTS: Seven of the 65 patients enrolled had to switch therapy for reasons of toxicity within the first 26 weeks. Diarrhoea was frequently reported (49 of 65, one discontinuation), but could be relieved by using antidiarrhoeal agents. Laboratory monitoring revealed elevated liver enzymes (leading to four discontinuations) and mild to moderate elevations of triglycerides and cholesterol (nine and 23 of 65, respectively). The exposure to saquinavir and nelfinavir was lower than expected. Abdominal pain was associated with a higher exposure to nelfinavir or saquinavir. The association of nausea and abdominal distension with drug exposure appeared to vary over time. CONCLUSIONS: The quadruple drug regimen was quite well tolerated. Diarrhoea was frequently reported but could be relieved by the use of antidiarrhoeal agents. In comparison with other protease inhibitor combinations, lipid abnormalities in plasma were infrequent and mild. With the exception of diarrhoea, all gastrointestinal complaints observed were found to be associated with the level of exposure to nelfinavir or saquinavir. The exposure to the protease inhibitors was relatively low, although the virologic efficacy of the regimen used was satisfactory. PMID- 10714569 TI - Changes in pathological findings at autopsy in AIDS cases for the last 15 years. AB - OBJECTIVE: To analyze changes in frequency of systemic AIDS pathology over time and its relationship to central nervous system pathology. DESIGN AND METHODS: A total of 390 AIDS autopsy cases obtained at University of California at San Diego Medical Center from 1982 to 1998 were reviewed retrospectively and linear regression analysis was used to evaluate significance of changes over time. RESULTS: Overall, the frequency of cytomegalovirus, Pneumocystis carinii pneumonia and Mycobacterium avium complex decreased, whereas bacterial infections increased and the frequency of fungal infection remained unchanged over time. The frequency of non-Hodgkin's lymphoma showed an upward trend over time, while the frequency of Kaposi's sarcoma remained unchanged. Following involvement of the lung (84%), the brain continued to be the second most frequently affected organ (63%). Whereas alterations of the brain by opportunistic infections or non Hodgkin's lymphoma showed a downward trend, HIV encephalitis continued to be detected in at least 25% of the cases. Cases with advanced HIV-related neuropathology and cases with no HIV involvement of the brain showed significant systemic pathology with opportunistic infections and neoplasms. In contrast, cases with early brain pathology (e.g., lymphocytic meningitis) showed minimal systemic pathology. Overall these trends remained unchanged throughout the total period covered by this study. CONCLUSIONS: This study suggests that despite the beneficial effects of antiretroviral and anti-opportunistic infection therapy, involvement of the brain by HIV continues to be a frequent autopsy finding. PMID- 10714570 TI - Overland heroin trafficking routes and HIV-1 spread in south and south-east Asia. AB - OBJECTIVES: Burma produces approximately 60% of the world's heroin, Laos is the third leading producer. Recent outbreaks of injecting drug use and HIV-1 in Burma, India, China, and Vietnam have been associated with Burmese and Laotian overland heroin trafficking routes. We analyzed findings from narcotics investigations, molecular epidemiology studies of HIV-1, and epidemiologic and behavioral studies of injecting drug use, to evaluate the roles that the heroin export routes play in the spread of drug use and HIV-1 in south and south-east Asia. METHODS: We reviewed the medical and narcotics literature, the molecular epidemiology of HIV, and did key informant interviews in India, China, and Burma with injecting drug users, drug traffickers, public health staff, and narcotics control personnel. RESULTS: Four recent outbreaks of HIV-1 among injecting drug users appear linked to trafficking routes. Route 1: From Burma's eastern border to China's Yunnan Province, with initial spread of HIV-1 subtype B, and later C. Route 2: Eastern Burma to Yunnan, going north and west, to Xinjiang Province, with B, C, and a B/C recombinant subtype. Route 3: Burma and Laos, through northern Vietnam, to China's Guangxi Province, subtype E. Route 4: Western Burma, across the Burma-India border to Manipur, predominant subtype C, and B and E. CONCLUSIONS: Overland heroin export routes have been associated with dual epidemics of injecting drug use and HIV infection in three Asian countries and along four routes. Molecular epidemiology is useful for mapping heroin routes. Single country narcotics and HIV programs are unlikely to succeed unless the regional narcotic-based economy is addressed. PMID- 10714571 TI - Safety of multiple daily applications of COL-1492, a nonoxynol-9 vaginal gel, among female sex workers. COL-1492 Phase II Study Group. AB - RATIONALE: COL-1492 is a nonoxynol-9 (N-9)-containing vaginal gel and may be a potential microbicide. As part of an effectiveness trial, an initial toxicity study was conducted. OBJECTIVES: The main objective of the reported study was the assessment of the toxicity of a 52.5 mg N-9 gel, COL-1492, when used a number of times each day by female sex workers. METHODS: This was a randomized, placebo controlled triple-blinded trial among female sex workers. The participants were asked to use the product for each vaginal sexual act. At each monthly visit a gynaecological examination with sexually transmitted disease sampling and colposcopy was performed. Venous blood was drawn for syphilis and HIV serology. All women received intensive counselling on condom use. Male condoms and sexually transmitted disease treatment were given free of charge. RESULTS: Only blinded results on the colposcopic examinations are reported. The incidence of lesions with or without an epithelial disruption was low: 0.06 and 0.29, respectively, per 100 woman-days in group A; 0.09 and 0.26 respectively per 100 woman-days in group B. There was no significant difference between the two arms. CONCLUSION: The multiple daily use of COL-1492 by female sex workers did not show an increase of local toxicity over that of a placebo. Colposcopy was discontinued in the autumn of 1997 in accordance with a Data Safety Monitoring Board decision. In the currently ongoing effectiveness trial the assessment of the product's toxicity continues to be monitored by simple visual examination. PMID- 10714572 TI - Itraconazole as an alternative for ritonavir liquid formulation when combined with saquinavir. PMID- 10714573 TI - Coreceptor usage of HIV-1 after deletion of Gly317-Pro318-Gly319 motif in the gp120 V3 loop. PMID- 10714574 TI - The cytokine milieu of HIV-associated non-Hodgkin's lymphoma favours aggressive tumours. PMID- 10714575 TI - Is there really a correlation between AIDS dementia and Kaposi's sarcoma? PMID- 10714576 TI - Are nelfinavir-containing regimens effective as second-line triple therapy? PMID- 10714577 TI - Effect of interferon and ribavirin on HIV viral load. PMID- 10714578 TI - Induction of human herpesvirus-8 gene expression by recombinant interferon gamma. PMID- 10714579 TI - Pharmacokinetics of nelfinavir during haemodialysis in a patient with HIV infection. PMID- 10714580 TI - In-vitro tipranavir susceptibility of HIV-1 isolates with reduced susceptibility to other protease inhibitors. PMID- 10714581 TI - Fibrillation potential amplitude to quantitatively assess denervation muscle atrophy. AB - Denervated muscle fibers exhibit spontaneous, repetitive single muscle fiber discharges and display fibrillation potentials detectable by electromyography. To explore the changing pattern of fibrillation potential amplitude after peripheral nerve injury and its relationship to the degree of muscle atrophy, fibrillation potential amplitudes were recorded on completely denervated biceps brachii of 173 patients with brachial plexus injury. Biceps brachii biopsies were taken at the same sites as the electromyogram recordings in 63 patients. The biopsies were analyzed by ATPase staining and the cross-sectional areas of fast and slow-twitch fibers were calculated. We found that the fibrillation potential amplitude and the cross-sectional areas of denervated muscle decay over time (P < 0.05), and both correlate negatively with denervation time (P < 0.01-0.05) within the first 15 months. The fibrillation potential amplitude correlates positively with both type I and II fiber cross-sectional areas (P < 0.0005-0.01). Our results show that fibrillation potential amplitude is closely correlated with muscle fiber size during the first 15 months after nerve injury, and it may therefore serve as a convenient index to evaluate quantitatively the degree of atrophy of denervated muscles. Electromyographic studies thus may help in designing treatment strategies. PMID- 10714582 TI - Clinical and neuropathological parameters affecting the diagnostic yield of nerve biopsy. AB - The value of nerve biopsy in the investigation of peripheral neuropathies is an important and controversial issue, partially obscured by the large variations in the diagnostic yield routinely reported for this procedure. The aim of this study was to evaluate the clinical and neuropathological parameters affecting the yield of nerve biopsy. We compared the experience of two independent neuropathology laboratories with different patient recruitment and neuropathological methods over 11 years (01/1987-12/1997). Clinicopathological correlations were studied retrospectively in 355 patients. Using the same criteria of evaluation, contributive biopsies accounted for 35.5% in one laboratory, and 47.3% in the other. Clinical parameters affecting the yield of nerve biopsy were: (a) the presumptive diagnosis at time of referral for biopsy; (b) the distribution of symptoms; and (c) the interval between disease onset and biopsy. Greater yield was associated with clinically suspected vasculitis, inflammatory demyelinating neuropathy or hereditary sensorimotor neuropathies. Contributive findings were more often reported with multifocal or asymmetrical presentations, and onset-to biopsy interval of less than 6 months. The contribution of nerve biopsy varied according to neuropathological techniques: (a) serial sections on frozen. paraffin-embedded and resin-embedded material improved sensitivity for interstitial pathology: (b) combined muscle biopsy increased sensitivity in the detection of vasculitis; and (c) teasing of nerve fibers added critical information to other classical techniques in only 4/102 cases. PMID- 10714583 TI - Absence of SMN gene deletion in post-polio syndrome. PMID- 10714584 TI - Severe gamma-sarcoglycanopathy caused by a novel missense mutation and a large deletion. AB - We report two siblings with a relatively severe limb-girdle muscular dystrophy. The elder sister presented at 8 years of age with inability to climb and abnormal gait. At 12 years she was barely ambulant. Her sister followed a similar course. Serum creatine kinase was 8500-10000 IU (N 25-200) in the elder sister and 17000 19000 IU in the younger sister. Muscle biopsy of the elder sister at 8 years showed chronic myopathic changes with loss of muscle fibres, active necrosis and regeneration. Immunocytochemistry demonstrated normal spectrin and dystrophin, reduced alpha-sarcoglycan and absent gamma-sarcoglycan--indicating a gamma sarcoglycanopathy. Haplotype analysis for the markers D13S115, D13S232, D13S292, D13S787, D13S1243 and D13S283 internal to and flanking the gamma-sarcoglycan gene showed the affected sisters shared haplotypes, indicating it was possible they were suffering from a gamma-sarcoglycanopathy. Non-inheritance of paternal alleles for D13S232, D13S292 and D13S1243 suggested the inheritance of a deletion, which was confirmed by FISH, using a genomic probe from the gamma sarcoglycan gene. The gamma-sarcoglycan cDNA was amplified by reverse transcriptase PCR from the muscle biopsy of the elder sister and sequenced. A missense mutation changing codon 69 from GGC glycine to CGC arginine was identified. HhaI digestion of exon 3 genomic PCR products showed the two affected sisters were hemizygous for the mutation, while the mother and grandmother were heterozygotes. The mutation, identified by SSCP analysis, was not observed in 116 unrelated, unaffected individuals. Previously, only two other missense mutations, the Cys283Tyr missense mutation in Gypsies and the Leu193Ser mutation in a Dutch family, have been described in the gamma-sarcoglycan gene. The fact that the affected individuals in the current and Gypsy families are gamma-sarcoglycan negative may indicate that codons 69 and 283 are important in gamma-sarcoglycan function. PMID- 10714585 TI - Genetic heterogeneity in three Chinese children with Fukuyama congenital muscular dystrophy. AB - Three Chinese patients, two boys and one girl, were afflicted with the typical clinical, myopathological and neuroradiological findings of Fukuyama congenital muscular dystrophy (FCMD). Polymorphism analysis of our patients did not reveal the founder haplotype (138-192-147-183 in D9S2105-D9S2170-D9S2171-D9S2107) of Japanese FCMD, even though one patient was descended from Japanese ancestry. Full mutational analysis of the fukutin gene revealed that there is neither 3 kb insertion nor point mutation. These findings suggest genetic heterogeneity between Chinese and Japanese FCMD patients. PMID- 10714586 TI - Shorter telomeres in dystrophic muscle consistent with extensive regeneration in young children. AB - Muscular dystrophies are characterised by continuous cycles of degeneration and regeneration resulting in an eventual diminution of the muscle mass and extensive fibrosis. In somatic cells chromosomal telomeres shorten with each round of cell division and telomere length is considered to be a biomarker of the replicative history of the cell. We have previously shown that human myoblasts have a limited proliferative capacity, and that normal skeletal muscle has a very low level of nuclear turnover. However, in patients suffering from muscular dystrophy the satellite cells will be forced to make repeated rounds of cell division, driving the cells towards senescence. In this study we have used the telomere length to quantify the intensity of the muscle cell turnover in biopsies from dystrophic patients of different ages. Our results show that as soon as the first clinical symptoms become apparent the muscle has already undergone extensive regeneration and the rate of telomere loss is 14 times greater than that observed in controls. This confirms that the decline in regenerative capacity is due to the premature senescence of the satellite cells induced by their excessive proliferation during muscle repair. PMID- 10714587 TI - Integrin and dystrophin associated adhesion protein complexes during regeneration of shearing-type muscle injury. AB - In shearing injury both the myofibres and connective tissue framework are breached and the muscle tendon continuity is disrupted. During regeneration the firm myofibre to extracellular matrix (ECM) adhesion must be re-established. We have analysed the expression of selected molecules implementing this adhesion in regenerating myofibres 2-56 days after transection of rat soleus muscle using quantitative immunohistochemistry and Northern blotting. Beta1 integrin mRNA level and alpha7 integrin and vinculin immunoreactivities were transiently increased in both the intact and regenerating parts of the transected myofibres by day 5-7 with normalization by day 10-14. After day 14, alpha7 integrin and vinculin accumulated at the tips of the regenerating myofibres, indicating formation of new mini-myotendinous junctions (mMTJ). Immunoreactivities for dystrophin and associated proteins as well as merosin appeared in regenerating myotubes by day 3-4 reaching control levels by day 56. Our results suggest that integrin and dystrophin associated molecules are complementary in myofibre-ECM adhesion. During regeneration, ruptured myofibres temporarily reinforce their integrin mediated lateral adhesion until mMTJs are formed. Thereby the load on the newly formed scar and the risk of rerupture are reduced. Dystrophin associated molecules appear later and replace integrin on the lateral aspects, while both complexes are abundant at the mMTJs. These molecular events correspond to our previous results on tensile strength. PMID- 10714588 TI - A clinical and genetic study of a manifesting heterozygote with X-linked myotubular myopathy. AB - X-linked myotubular myopathy (XLMTM) characteristically causes severe or fatal muscle weakness in male infants. Mutations in the gene MTM1, encoding the protein myotubularin, can be identified in most families. Prior to this report, XLMTM was thought not to cause symptomatic manifestations in female carriers. We describe an adult female from a large family with typical XLMTM. The patient had progressive disabling muscle weakness of later onset and lesser severity than that observed in affected males. The distribution of weakness resembled typical XLMTM with facial weakness, marked limb-girdle weakness, respiratory muscle involvement and dysphagia. Analysis of the MTM1 gene identified a heterozygous missense mutation (G378R) within the highly conserved tyrosine phosphatase site of myotubularin. We did not identify significantly skewed X-inactivation. We conclude that XLMTM is capable of causing significant disability in heterozygotes. PMID- 10714589 TI - A missense mutation T487N in the myophosphorylase gene in a Spanish patient with McArdle's disease. AB - A heterozygous C-to-A substitution at codon 487, changing a highly conserved threonine to an asparagine (T487N) was identified in two siblings with McArdle's disease who were also heterozygous for the nonsense mutation at codon 49 (R49X). Our data further expand the genetic heterogeneity in patients with McArdle's disease. PMID- 10714590 TI - A family with PROMM not linked to the recently mapped PROMM locus DM2. AB - Proximal myotonic myopathy is an autosomal dominantly inherited multisystem disorder, clinically similar to but genetically distinct from myotonic dystrophy (DM). A recently mapped second locus for myotonic dystrophy was thought to be an attractive candidate locus for PROMM, and this hypothesis was supported by reports of linkage to this locus in some PROMM families. We present a large German pedigree with PROMM in which linkage to this locus could be excluded, showing that PROMM is genetically heterogeneous. PMID- 10714592 TI - Generalized calcification in a case of dermatomyositis. PMID- 10714591 TI - Immunolabelling of mitochondrial superoxide dismutase and of Hsp60 in muscles harbouring a respiratory chain deficiency. AB - In mitochondrial encephalomyopathies, impairment of the electron transfer chain may lead to overproduction of reduced oxygen species because oxygen consumption is decreased. Whether heat shock proteins (Hsp) are induced or not in mitochondria against oxidative stress is questionable. Muscle ragged-red fibres are the histological hallmark of most respiratory chain deficiencies in humans. They exhibit abnormal mitochondria which accumulate mainly under their sarcolemma. Within these fibres, immunolabelling demonstrated strong expression of mitochondrial manganese-dependent superoxide dismutase and a lack of expression of mitochondrial Hsp60 within the subsarcolemmal spaces. In contrast, Hsp60 was overexpressed within the intermyofibrillar mitochondria. These findings suggest enhanced generation and dismutation of superoxide anions and that processing and integration of imported precursor proteins is impaired within the subsarcolemmal mitochondrial aggregates of ragged-red fibres, whereas protein import and assembly may still be efficient in the intermyofibrillar mitochondria of these fibres. PMID- 10714593 TI - Neuromuscular disorders: gene location.Vol. 10 No. 2, February 2000. PMID- 10714594 TI - Mitochondrial encephalomyopathies: gene mutation. Vol. 10 No. 2, February 2000. PMID- 10714595 TI - Emergency physicians and their attrition rate. PMID- 10714596 TI - Communicating rupture of pulmonary hydatid cysts with resulting acute respiratory distress syndrome. PMID- 10714597 TI - Allergic reaction associated with intravenous marijuana use. PMID- 10714598 TI - The resuscitation alphabet--when does "U" come before "E"? PMID- 10714599 TI - Lecture on insect stings at Queen of the Valley Hospital in Napa, California. PMID- 10714600 TI - An interview with a distinguished pharmaceutical scientist: Douwe D. Breimer- elected Foreign Associate Member of the Institute of Medicine of the National Academy of Sciences of the U.S.A. (1999). PMID- 10714601 TI - Systematic investigations of the influence of molecular structure on the transport of peptides across cultured alveolar cell monolayers. AB - PURPOSE: To determine how the structures of peptides influence their alveolar permeability. METHODS: The studies were performed using 14 synthetic 'model' peptides, labelled with a novel, non-intrusive amino acid fluorophore, and their transport studied using rat alveolar cell monolayers cultured on permeable supports. RESULTS: The passage of the peptides across the epithelial cell monolayers is shown to be primarily paracellular, with an inverse dependence on molecular size, and an enhanced flux observed for cationic peptides. The apparent permeability coefficients (Papp) for the peptides (together with those for other organic solutes, taken from the literature) are shown to be well-modelled assuming two populations of 'pores' in the monolayers, modelled as cylindrical channels of radii 15 A and 22 nm. The former pores are shown to be numerically equitable with the monolayer tri-junctional complexes, and the latter are taken as monolayer defects. CONCLUSIONS: The various monolayer Papp values correlate well with the results from in vivo transport experiments, and the conclusion is drawn that the pulmonary delivery of peptide drugs is perfectly exploitable. PMID- 10714602 TI - N-terminal halves of rat H+/peptide transporters are responsible for their substrate recognition. AB - PURPOSE: Peptide transporters PEPT1 and PEPT2 differ substantially in their substrate affinity and recognition. The aim of this study is to define the structural domains which influence the functional characteristics of both transporters METHODS: Two kinds of chimeric peptide transporters (PEPT-N1C2 and PEPT-N2C1) were constructed, and their functional characteristics were compared with those of wild-type transporters in stable transfectants. RESULTS: PEPT-N1C2, the N-terminal half of rat PEPT1 and the C-terminal half of rat PEPT2, and the reciprocal chimera PEPT-N2C1 were functionally expressed in LLC-PK1 cells. The pH profiles of [14C] glycylsarcosine uptake by PEPT-N1C2 and PEPT-N2C1 were close to those of PEPT1 and PEPT2, respectively. Substrate recognition for PEPT-N1C2 and PEPT-N2C1 was also similar to that of PEPT1 and PEPT2, respectively. However, substrate affinities for PEPT-N1C2 were higher than those for PEPT1, although those for PEPT-N2C1 and PEPT2 were comparable. CONCLUSIONS: These results indicate that functional regions which are associated with the extracellular pH changes and are responsible for substrate recognition of PEPT1 and PEPT2 may be located in the N-terminal halves of the proteins. In addition, it is suggested that the domain to affect the substrate affinity exists in the C-terminal as well as in the N-terminal half of rat PEPT2. PMID- 10714603 TI - Induction of UDP-glucuronosyltransferase by the flavonoids chrysin and quercetin in Caco-2 cells. AB - PURPOSE: Dietary flavonoids have been reported to be potent inhibitors of drug metabolizing enzymes. In the present study we examined the inducing effect of three of these compounds, chrysin, quercetin and genistein, on UDP glucuronosyltransferase (UGT) in the human intestinal cell line Caco-2. METHODS: The induction of UGT by flavonoid pretreatment was studied both in the intact cells and cell homogenates, measured as the glucuronidation of chrysin, and by immunoblot analysis of the UGT 1A protein. RESULTS: Exposure of Caco-2 cells to 50 microM chrysin resulted in a 3.8-fold increase in chrysin glucuronidation in intact cells (p < 0.0001) with a 38% decrease in sulfation (p < 0.01). In the cell homogenate the induction was much larger, 14-fold. The induction was slow to develop with maximum induction after 3-4 days. Interestingly, the isoflavonoid genistein was without effect. Immunoblot analysis of Caco-2 cell microsomes with a UGT1A subfamily-selective antibody showed a markedly increased band at about 59 kDa, consistent with induction of one or more UGT1A isoforms. A 5-week exposure of Caco-2 cells to low concentrations (10 microM) of chrysin or quercetin also showed markedly increased glucuronidation activity. CONCLUSIONS: Diet-mediated induction of intestinal UGT may be important for the bioavailability of carcinogens and other toxic chemicals as well as therapeutic drugs. PMID- 10714604 TI - N-trimethylated chitosan chloride (TMC) improves the intestinal permeation of the peptide drug buserelin in vitro (Caco-2 cells) and in vivo (rats). AB - PURPOSE: To evaluate N-trimethyl chitosan chloride (TMC) of high degrees of substitution as intestinal permeation enhancers for the peptide drug buserelin in vitro using Caco-2 cell monolayers, and to investigate TMCs as enhancers of the intestinal absorption of buserelin in vivo, in rats. METHODS: TMCs were tested on Caco-2 cells for their efficiency to increase the paracellular permeability of the peptide buserelin. For the in vivo studies male Wistar rats were used and buserelin was administered with or without the polymers intraduodenally. Both types of experiments were performed at pH 7.2. RESULTS: Transport studies with Caco-2 cell monolayers confirmed that the increase in buserelin permeation is dependent on the degree of trimethylation of TMC. In agreement with the in vitro results, in vivo data revealed highly increased bioavailability of buserelin following intraduodenal co-administration with 1.0% (w/v) TMCs. Intraduodenally applied buserelin resulted in 0.8% absolute bioavailability, whereas co administrations with TMCs resulted in mean bioavailability values between 6 and 13 %. Chitosan HCl (1.0%; pH = 7.2) did not significantly increase the intestinal absorption of buserelin. CONCLUSIONS: Both the in vitro and in vivo results indicate that TMCs are potent mucosal permeation enhancers of the peptide drug buserelin at neutral pH values. PMID- 10714605 TI - Transdermal delivery of macromolecules using skin electroporation. AB - PURPOSES: (1) To evaluate the feasibility of transdermal delivery of macromolecules by skin electroporation. (2) To assess the influence of the molecular weight of the permeant on transport and examine whether there exists a "cut-off" value of molecular weight. (3) To localize the transport pathways of the macromolecules in the skin. METHODS: FITC-dextran (FD) of increasing molecular weight (4.4, 12 and 38 kDa) were used as model macromolecules to study the extent of transport across hairless rats skin in vitro and to localize their distribution in the skin by confocal scanning laser microscopy. RESULTS: Electroporation enhanced the transport of the macromolecules as compared to passive diffusion. The transdermal delivery by skin electroporation of FITC and FD 4.4 was equivalent whereas transport of higher molecular weight FD was lower but significant. FITC and FD 38 were observed in the epidermis both around and in the keratinocytes. CONCLUSIONS: Transdermal and topical delivery of macromolecules of at least 40 kDa can be achieved by skin electroporation. PMID- 10714606 TI - Surface active agents as enhancers of alveolar absorption. AB - PURPOSE: Small solutes which are deposited in the alveoli by aerosol inhalation will be absorbed across the alveolo-capillary barrier. Inhalation of dioctyl sodium sulfosuccinate (DOSS) enhances absorption while having little or no effect on lung function, suggesting that surface active agents may be used as enhancers of alveolar absorption of inhaled pharmaceuticals. The purpose of this study was to examine the effects of a selection of different surface active agents on alveolar absorption. METHODS: The absorption of 99mTc-diethylene triamine pentaacetate (99mTc-DTPA) from the lungs was studied in rabbits. We studied five different surface active agents: DOSS, sodium glycodioxycholate (GDCA), sodium lauryl sulphate (NaLS), lysophosphatidyl choline (LPC) and polyoxyethylene-23 laurylether (P23LE). RESULTS: DOSS and GDCA both dramatically enhanced the absorption of 99mTc-DTPA. There was a moderate effect of NaLS, no significant effect of LPC and P23LE reduced the rate of absorption. None of the compounds affected gas exchange or lung compliance. CONCLUSIONS: There is a wide spectrum of effects of inhaled surface active agents on the alveolar absorption of 99mTc DTPA. Ionic compounds such as DOSS and GDCA have the greatest effect, and further studies of these classes of surface active agents for use as enhancers of alveolar absorption of pharmaceuticals seem warranted. PMID- 10714608 TI - Detection of the membrane protein recognized by the kidney-specific alkylglucoside vector. AB - PURPOSE: Previously, we suggested that alkylglucoside can be an effective vector for renal-specific drug delivery (Suzuki et al., J. Pharmacol. Exp. Ther, 288:57 61, 1999). The purpose of the present study is to characterize the membrane protein which is recognized by this alkylglucoside. METHODS: The binding of [125I] tyrosine conjugated with a octylthioglucoside (Glc-S-C8-[125I]Tyr) Glc-S C8-[125I]Tyr to crude membrane fractions of kidney was determined. In addition, the membrane was cross-linked with this alkylglucoside and examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. RESULTS: Glc-S-C8-[125I]Tyr was shown to have a specific binding site on the kidney membrane (Kd = 931 nM and Bmax = 987 pmol/mg protein). Cross-linking of the membrane with Glc-S-C8 [125I]Tyr resulted in the detection of a protein (Mr = 62,000), which was unaffected by reducing agents. The results of this cross-linking study were consistent with previous information on its localization and binding characteristics. CONCLUSIONS: The kidney membrane protein, to which alkylglucoside binds in a specific manner, has a molecular weight of 62,000. Crosslinking is a useful tool for detecting this novel membrane protein in kidney. PMID- 10714607 TI - Liposomes containing interferon-gamma as adjuvant in tumor cell vaccines. AB - PURPOSE: Liposomal systems may be useful as a cytokine supplement in tumor cell vaccines by providing a cytokine reservoir at the antigen presentation site. Here, we examined the effect of liposome incorporation of mIFNgamma on its potency as adjuvant in an established tumor cell vaccination protocol in the murine B16 melanoma model. Adjuvanticity of the mIFNgamma-liposomes was compared to that achieved by mIFNgamma-gene transfection of the B16 tumor cells. Furthermore, we studied whether liposomal incorporation of mIFNgamma indeed increases the residence time of the cytokine at the vaccination site. METHODS: C57B1/6 mice were immunized with i) irradiated IFNgamma-gene transfected B16 melanoma cells or ii) irradiated wild type B16 cells supplemented with (liposomal) mIFNgamma, followed by a challenge with viable B16 cells. The residence time of the (liposomal) cytokine at the subcutaneous (s.c.) vaccination site was monitored using radiolabeled mIFNgamma and liposomes. RESULTS: Immunization with irradiated tumor cells admixed with liposomal mIFNgamma generated comparable protection against B16 challenge as immunization with mIFNgamma-gene modified tumor cells. Irradiated tumor cells admixed with soluble mIFNgamma did not generate any protective responses. Radiolabeling studies indicated that free mIFNgamma rapidly cleared from the s.c. injection site. Association of [125I]-mIFNgamma with liposomes increased the local residence time substantially: liposomal association of mIFNgamma resulted in a prolonged local residence time of the cytokine as reflected by a 4-fold increase of the area under the curve. The amount of released cytokine in the optimal dose range corresponds to the amount released by the gene-transfected cells. Moderate but significant CTL-activity against B16 cells was found for mice immunized with irradiated cells supplemented with mIFNgamma-liposomes compared to untreated control animals. CONCLUSIONS: Prolonged presence of mIFNgamma at the site of antigen presentation is crucial for the generation of systemic immune responses in the B16 melanoma model. These studies show that liposomal encapsulation of cytokines is an attractive strategy for paracrine cytokine delivery in tumor vaccine development. PMID- 10714609 TI - Functional clarification of MCT1-mediated transport of monocarboxylic acids at the blood-brain barrier using in vitro cultured cells and in vivo BUI studies. AB - PURPOSE: To prove the functional significance of monocarboxylic acid transporter, MCT1 at the blood-brain barrier (BBB) for the passage of both endogenous and exogenous monocarboxylic acids into the central nervous system. METHODS: Monocarboxylic acid transport at the BBB was studied in rats by using a newly established immortalized brain capillary endothelial cell (BCEC) line, RBEC1, and the results were compared with those obtained by using primary cultured BCECs, cells stably expressed with rat MCT1, and the in vivo brain uptake index (BUI) method. RESULTS: The cell line, RBEC1 meets various morphological and enzymatic criteria of BCECs and appears to be suitable for the study of BBB transport of monocarboxylic acids. The presence of MCT1-transcript in RBEC1 was confirmnned by the RT-PCR method, as previously observed in isolated brain capillaries. A typical substrate of MCT1, lactic acid, was taken up by RBEC1 in a stereospecific and saturable manner. The value of the kinetic parameter Km showed good agreement with values previously obtained in studies using an in vivo BUI and in vitro MCT1 transfected cells. An organic weak acid, benzoic acid, which has been considered to cross biological membranes by passive diffusion, exhibited carrier-mediated transport properties, such as saturation, pH dependence, and stereospecific inhibition in RBEC1, similar to those we observed in primary cultured rat BCECs. The Km values in RBEC1, in primary cultured BCECs and in the in vivo BUI method were comparable and well agreed with that obtained in MCT1-transfected cells, suggesting that the transport features of benzoic acid observed by in vitro methods well reflect the in vivo transport activity. Furthermore, hybrid depletion of MCT1 in RBEC1 using an antisense oligonucleotide against rat MCT1 abolished the saturable transport of benzoic acid. CONCLUSIONS: These observations show that MCT1 plays a significant role in the transport of monocarboxylic acids, including the exogenous organic weak acid benzoic acid, as well as native lactic acid. PMID- 10714610 TI - Blood-brain barrier transport of 125I-labeled basic fibroblast growth factor. AB - PURPOSE: This study was carried out to examine the blood-brain barrier (BBB) transport of human basic fibroblast growth factor (bFGF) and investigate its mechanism. METHODS: The BBB transport of 125I-bFGF was measured by several in vivo methods including intravenous administration, in situ internal carotid artery perfusion, and intracerebral microinjection. The in vitro binding of 125I bFGF was characterized using freshly prepared bovine brain capillaries. RESULTS: The distribution volume of 125I-bFGF in the postvascular supernatant increased with the perfusion time, and exceeded the space occupied by the brain microvasculature and its trichloroacetic acid (TCA) precipitability was more than 90%. 125I-bFGF avidly bound to isolated bovine brain capillaries with a Bmax of 206 +/- 48 pmol/mg protein, and a Kd of 36.5 +/- 15.7 nM. This binding was significantly inhibited by unlabeled bFGF and heparin in a concentration dependent manner. The cationic peptides, protamine and poly-L-lysine (each 300 microM), produced over 85% inhibition of 125I-bFGF binding to brain capillaries. Furthermore, glycosaminoglycans with a sulfate residue, chondroitin sulfate B and C (each 10 microg/mL) also inhibited the binding of 125I-bFGF The in vivo transcytosis of 125I-bFGF from the luminal side to the brain was also inhibited by the presence of heparin (10 microg/mL) and poly-L-lysine (300 microM), whereas neither hyaruronic acid (10 microg/mL) nor insulin (10 microM) had any effect. In addition to these results, the brain efflux index method was used to confirm that the transcytosis of 125I-bFGF from brain to blood across the BBB was negligible. CONCLUSIONS: These results suggest that 125I-bFGF is transported across the BBB, possibly by an adsorptive-mediated transcytosis mechanism that is triggered by binding to negatively charged species on the luminal membrane surface of the brain microvasculature, such as heparan sulfate proteoglycans. PMID- 10714611 TI - Evaluation of blood-brain barrier passage of a muscarine M1 agonist and a series of analogous tetrahydropyridines measured by in vivo microdialysis. AB - PURPOSE: To investigate the blood-brain barrier (BBB) passage of the M1 muscarine agonist Lu 25-109 (5-(2-Ethyl-2H-tetrazol-5-yl)-1,2,3,6-tetrahydro methylpyridine) and potential metabolites using in vivo microdialysis. METHODS: Anesthetized rats were administered an intravenous infusion of one of seven analogs with a Log D7.4 ranging from 0.35 to -2.4. Microdialysis probes were implanted in the brain and the jugular vein. The integrity of the BBB was evaluated using 2-amino-3-(3-hydroxy-5-phenylisoxazol-4-yl)propionic acid (APPA), a compound not expected to penetrate the BBB. The data was corrected for in vitro recovery. RESULTS: Lu 25-109, Lu 24-165 (demethylated Lu 25-109) and Lu 25-077 (N demethylated Lu 25-109) entered the brain in a 1:1 ratio with the blood. Although Lu 29-081 (hydroxylated Lu 25-109) presented a similar Log D7.4 to Lu 25-109 and Lu 24-164, it entered the brain with a lower brain:blood ratio of 0.5. Lu 32-181 (Lu 25-109 N-oxide), Lu 35-026 (deethylated and oxidized Lu 25-109) and Lu 31-126 (deethylated Lu 25-109) were not detected in the brain samples, indicating no penetration. Infusion of Lu 25-109 resulted in a time perspective of the formation and distribution of the two metabolites Lu 25-077 and Lu 32-181. Although the hydroxylated compound (Lu 29-081) had a Log D74 of -0.6, within the range 0.35 to -0.83 of the compounds penetrating the BBB, it showed a brain: blood ratio of 0.5. Lu 35-026 showed an unusual infusion profile with a tmax of 100-150 min and a subsequent decrease in blood concentration. CONCLUSIONS: Compounds with Log D7.4 above -0.83 penetrated the BBB, whereas compounds below 1.5 did not. Knowledge of Log D7.4 values is not sufficient to evaluate BBB passage because the value does not predict the influence of active transport processes. PMID- 10714612 TI - Population pharmacokinetics of fast release oral diclofenac in healthy volunteers: relation to pharmacodynamics in an experimental pain model. AB - PURPOSE: Population pharmacokinetics of a fast release diclofenac were assessed with special focus on pharmacodynamic implications. METHODS: In a double blind four-way crossover study, 20 healthy volunteers received orally 50 and 100 mg diclofenac-Na effervescent ("fast-release NSAID"), 50 mg diclofenac tablets ("control"), or placebo. Population pharmacokinetics of the fast release diclofenac were assessed using a nonlinear mixed effects modeling approach (NON MEM). Analgesic effects were investigated by means of an experimental pain model based on both pain-ratings and cortical evoked potentials after specific stimulation of nasal nociceptors with short pulses of gaseous CO2. RESULTS: Pharmacokinetics of fast release diclofenac were best described by a two compartment population model, with an estimated terminal half-life of 1.2 hours. Pharmacokinetics of diclofenac tablets were highly variable and a population pharmacokinetic model could not be obtained. As an indication of an early onset of analgesic effects, 100 mg fast release diclofenac but not the tablets significantly reduced the amplitudes of pain-related evoked potentials at 30 min after administration. CONCLUSIONS: Earlier drug absorption and lower pharmacokinetic variability of the fast-release formulation are likely to be preserved in a population. PMID- 10714613 TI - pH-metric solubility. 2: correlation between the acid-base titration and the saturation shake-flask solubility-pH methods. AB - PURPOSE: The objective of this study was to compare the results of a normal saturation shake-flask method to a new potentiometric acid-base titration method for determining the intrinsic solubility and the solubility-pH profiles of ionizable molecules, and to report the solubility constants determined by the latter technique. METHODS: The solubility-pH profiles of twelve generic drugs (atenolol, diclofenac.Na, famotidine, flurbiprofen, furosemide, hydrochlorothiazide, ibuprofen, ketoprofen, labetolol.HCl, naproxen, phenytoin, and propranolol.HCl), with solubilities spanning over six orders of magnitude, were determined both by the new pH-metric method and by a traditional approach (24 hr shaking of saturated solutions, followed by filtration, then HPLC assaying with UV detection). RESULTS: The 212 separate saturation shake-flask solubility measurements and those derived from 65 potentiometric titrations agreed well. The analysis produced the correlation equation: log(1/S)titration = -0.063(+/- 0.032) + 1.025(+/- 0.011) log(1/S)shake-flask, s = 0.20, r2 = 0.978. The potentiometrically-derived intrinsic solubilities of the drugs were: atenolol 13.5 mg/mL, diclofenac.Na 0.82 microg/mL, famotidine 1.1 mg/ mL, flurbiprofen 10.6 microg/mL, furosemide 5.9 microg/mL, hydrochlorothiazide 0.70 mg/mL, ibuprofen 49 microg/mL, ketoprofen 118 microg/mL, labetolol.HCl 128 microg/mL, naproxen 14 microg/mL, phenytoin 19 microg/mL, and propranolol.HCl 70 microg/mL. CONCLUSIONS: The new potentiometric method was shown to be reliable for determining the solubility-pH profiles of uncharged ionizable drug substances. Its speed compared to conventional equilibrium measurements, its sound theoretical basis, its ability to generate the full solubility-pH profile from a single titration, and its dynamic range (currently estimated to be seven orders of magnitude) make the new pH-metric method an attractive addition to traditional approaches used by preformulation and development scientists. It may be useful even to discovery scientists in critical decision situations (such as calibrating computational prediction methods). PMID- 10714614 TI - Quantitative relationship between solubility, initial dissolution rate and heat of solution of chiral drugs. AB - PURPOSE: The aim of this study was to clarify the quantitative relationship between solubility, initial dissolution rate and heat of solution of racemic compound and its enantiomers. METHODS: Propranolol, propranolol HCl, tyrosine, and tryptophan were used as typical chiral drugs. The heat of solution of chiral drug was measured by an isothermal microcalorimeter and the heat of fusion was measured by a DSC. The free energy difference for the dissolution of drug was calculated from the solubility and initial dissolution rate data. RESULTS: The free energy difference and enthalpy difference of the dissolution between the racemic compound and enantiomer of propranolol, propranolol hydrochloride, tyrosine, and tryptophan were obtained by the solubility, initial dissolution rate and heat of solution data. A good linearity was observed in the free energy difference and the enthalpy difference for the dissolution of them, except for propranolol HCl data. By considering the dissociation in solution, the data of propranolol HCl followed the regression line. CONCLUSIONS: The free energy difference of the dissolution was linearly dependent on the enthalpy difference for the racemic compound and its enantiomers. The results fit the theoretical equation. It could be possible to estimate the solubility of chiral insoluble drug from the thermal data. PMID- 10714615 TI - Spray-dried lactose composite particles containing an ion complex of alginate chitosan for designing a dry-coated tablet having a time-controlled releasing function. AB - PURPOSE: The properties of novel spray-dried lactose composite particles suitable for the coating filler of a dry-coated tablet having a long induction period in drug release were investigated. METHODS: To prepare spray-dried composite particles containing alginate-chitosan complex (SD(L/AL-CS)), an aqueous solution of lactose and sodium alginate and the acetic acid solution of chitosan were concomitantly fed into the rotary atomizer of a spray-dryer. The formation of the alginate-chitosan complex was confirmed by measuring the weight of insoluble portion in the mixture of sodium alginate and chitosan solutions. The dissolution properties of the dry-coated tablet were measured with the JP specified paddle method. RESULTS: The micromeritic properties of SD(L/AL-CS) were compared to those of the SD composite particles of lactose-sodium alginate, having a good compacting property. The drug release profiles of dry-coated tablet with SD(L/AL CS) contained a long induction period followed by a rapid drug release phase in the artificial intestinal fluid. The induction period for drug release to occur was increased with an increase in the degree of deacetylation of chitosan and in the amount of chitosan in the formulation. The prolongation of induction period was attributed to the formation of an insoluble ion complex between sodium alginate and chitosan in the composite particles, which could form a rigid gel structure on the tablet surface. CONCLUSIONS: A time-controlled release tablet was designed with the composite particles of lactose containing the alginate chitosan ion complex. The induction period of the dry-coated tablet could be prolonged in order to deliver the drug to the colon by controlling the type and amount of chitosan formulated in the composite particles. PMID- 10714616 TI - Visual evidence of acidic environment within degrading poly(lactic-co-glycolic acid) (PLGA) microspheres. AB - PURPOSE: In the past decade, biodegradable polymers have become the materials of choice for a variety of biomaterials applications. In particular, poly(lactic-co glycolic acid) (PLGA) microspheres have been extensively studied for controlled release drug delivery. However, degradation of the polymer generates acidic monomers, and acidification of the inner polymer environment is a central issue in the development of these devices for drug delivery. METHODS: To quantitatively determine the intrapolymer acidity, we entrapped pH-sensitive fluorescent dyes (conjugated to 10,000 Da dextrans) within the microspheres and imaged them with confocal fluorescence microscopy. The technique allows visualization of the spatial and temporal distribution of pH within the degrading microspheres (1). RESULTS: Our experiments show the formation of a very acidic environment within the particles with the minimum pH as low as 1.5. CONCLUSIONS: The images show a pH gradient, with the most acidic environment at the center of the spheres and higher pH near the edges, which is characteristic of diffusion-controlled release of the acidic degradation products. PMID- 10714617 TI - Approaches to the enhancement of patient adherence to antidepressant medication treatment. AB - The number of safe and effective medication treatments for depression has increased significantly over the past 10 years. Relative to the older tricyclic antidepressants and monoamine oxidase inhibitors, the newer medications offer comparable efficacy with fewer side effects and a markedly reduced risk for serious adverse effects. In spite of these benefits, and in spite of the extensive and successful efforts that have been made to inform the general population about the diagnosis and treatment of depression, many patients do not comply with treatment recommendations. Although specific factors such as side effects lead to high rates of noncompliance with medication treatment, noncompliance is a multifactorial phenomenon. The reasons for noncompliance can include rational and intentional decisions based on beliefs about the illness, concerns over side effects, ineffectiveness of treatment. costs of the medication, decisions influenced by the symptoms of the disorder, and many other cultural and attitudinal factors. Some of the important concepts that should be addressed with depressed patients are reviewed. Strategies aimed at informing patients about depression and its treatment and providing a collaborative treatment environment have the potential to significantly improve treatment outcome and treatment adherence. PMID- 10714618 TI - Management of nonresponse and intolerance: switching strategies. AB - Approximately 29% to 46% of depressed patients show only partial or no response to treatment with antidepressants, with intolerance a frequent cause of treatment failure or discontinuation. Clinicians frequently switch to other antidepressants patients who have failed to tolerate or to respond to antidepressant treatment. The switching strategy involves substitution of another agent for the agent that has either caused intolerable side effects or has failed to induce a response. Fredman and colleagues have recently surveyed 402 psychiatrists from various parts of the country and asked them what steps they would take for patients who fail to respond to 8 weeks or more of an adequate dose of a selective serotonin reuptake inhibitor (SSRI). Interestingly, switching to a non-SSRI agent was the most popular choice indicated by psychiatrists (44% of respondents), with dual acting agents and bupropion being the next most commonly chosen agents. Even though there are no controlled trials of switching strategies in the literature to date, clinicians often choose this course of action. This article will review some of the currently available studies on switching strategies. PMID- 10714619 TI - Augmentation strategies in depression 2000. AB - Augmentation strategies and combination treatments have become a popular method of treating refractory depression, enhancing therapeutic response in partial responders and increasing the likelihood of more rapid response. The evidence supporting these strategies will be reviewed, their methods of administration discussed, and the relative advantages and disadvantages considered. PMID- 10714620 TI - Clinical issues in long-term treatment with antidepressants. AB - Historically, the emphasis in treating depression has been focused on the acute phase of treatment, with few published data on the continuation and maintenance phases of treatment. Yet the risk of depression increases with each episode, with a 50% to 90% chance of developing another episode after 1 or 2 prior episodes of depression. Moreover, subsequent episodes of depression are often of longer duration, more severe, and less responsive to treatment. Most patients with major depression require some form of long-term antidepressant treatment, and many need lifelong treatment. Optimizing efficacy and minimizing side effects are essential during both the acute and long-term phases of antidepressant treatment. Antidepressant side effects, including insomnia or somnolence, weight gain, asthenia, and sexual dysfunction, can significantly decrease patient compliance with long-term treatment for depression. Identification and management of side effects, combined with early and ongoing educational messages to the patient about treatment issues and the importance of sustaining illness remission, help improve compliance and reduce the potential for premature discontinuation of an otherwise optimal antidepressant. PMID- 10714622 TI - Whipple or pylorus preservation? A critical reappraisal and some new insights into pancreaticoduodenectomy. PMID- 10714621 TI - Outcome of pancreaticoduodenectomy with pylorus preservation or with antrectomy in the treatment of chronic pancreatitis. AB - OBJECTIVE: To compare the short- and long-term results of pancreaticoduodenectomy with pylorus preservation (PPPD) or with antrectomy (Whipple procedure) in the treatment of selected patients with chronic pancreatitis. BACKGROUND: PPPD may be preferred over Whipple because of its purported nutritional advantages and the reduced likelihood of postgastrectomy syndromes. METHODS: A retrospective review was performed of 72 consecutive patients undergoing pancreaticoduodenectomy for chronic pancreatitis between 1991 and 1997. RESULTS: PPPD was performed in 39 patients and Whipple in 33. The two patient populations had similar characteristics. Short-term complications included (PPPD vs. Whipple): pancreatic or biliary fistulas (5.1% vs. 15%), delayed gastric emptying (33% vs. 12%), cholangitis (2.6% vs. 6.1%), and death (0 vs. 3%). Delayed gastric emptying was not associated with other complications and resulted in longer hospital stays for PPPD than for Whipple patients (15 vs. 12 days). The duration of follow-up averaged 41 +/- 24 months. Long-term weight status was similar, with body-mass indices of 22.1 and 22.9 after PPPD and Whipple, respectively. Postoperative enzyme supplementation (63% vs. 77%) and new-onset diabetes (10% vs. 12%) did not differ significantly between the PPPD and Whipple groups. Dumping, bile gastritis, or peptic ulcer disease occurred in three patients after PPPD and in three after Whipple. Complete or partial pain relief was attained in 60% and 70% of patients after PPPD and Whipple, respectively. Multivariate analysis of preoperative variables revealed that site-specific pathology in the head of the pancreas was the only independent factor associated with successful pain relief after pancreatic resection. CONCLUSION: PPPD results in higher frequencies of postoperative delayed gastric emptying compared with the Whipple procedure. Both operations achieve comparable long-term nutritional results, cause new insulin dependence in surprisingly few patients, and provide equivalent pain relief to 65% of selected patients. Patients with disproportionate pathology in the head of the pancreas have a higher likelihood of successful pain relief. PMID- 10714623 TI - Columnar mucosa and intestinal metaplasia of the esophagus: fifty years of controversy. AB - OBJECTIVE: To outline current concepts regarding etiology, diagnosis, and treatment of intestinal metaplasia of the esophagus and cardia. SUMMARY BACKGROUND DATA: Previously, endoscopic visualization of columnar mucosa extending a minimum of 3 cm into the esophagus was sufficient for the diagnosis of Barrett's esophagus, but subsequently the importance of intestinal metaplasia and the premalignant nature of Barrett's have been recognized. It is now apparent that shorter lengths of intestinal metaplasia are common, and share many features of traditional 3-cm Barrett's esophagus. METHODS: Themes and concepts pertaining to intestinal metaplasia of the esophagus and cardia are developed based on a review of the literature published between 1950 and 1999. RESULTS: Cardiac mucosa is the precursor of intestinal metaplasia of the esophagus. Both develop as a consequence of gastroesophageal reflux. Intestinal metaplasia, even a short length, is premalignant, and the presence of dysplasia indicates progression on the pathway to adenocarcinoma. Antireflux surgery, as opposed to medical therapy, may induce regression or halt progression of intestinal metaplasia. The presence of high-grade dysplasia is frequently associated with an unrecognized focus of adenocarcinoma. Vagal-sparing esophagectomy removes the diseased esophagus and is curative in patients with high-grade dysplasia. Invasion beyond the mucosa is associated with a high likelihood of lymph node metastases and requires lymphadenectomy. CONCLUSIONS: Despite improved understanding of this disease, controversy about the definition and best treatment of Barrett's esophagus continues, but new molecular insights, coupled with careful patient follow-up, should further enhance knowledge of this disease. PMID- 10714624 TI - Ultrasonically activated shears in thyroidectomies: a randomized trial. AB - OBJECTIVE: To test whether the advantages of the ultrasonically activated shears (UAS) observed in thyroidectomies in a previous matched-pair study could be repeated in a randomized trial. SUMMARY BACKGROUND DATA: The UAS has been documented, mainly in nonrandomized studies, to be a safe and fast device in video-assisted and conventional surgery. METHODS: Thyroidectomies and lobectomies performed for benign or malignant thyroid disease between August 1997 and January 1999 were included in this series. Separate randomization, resulting in four sets of envelopes, was done for one consultant endocrine surgeon and for senior residents for both lobectomies and for total thyroidectomies. The operations performed with the UAS were compared with operations performed with the conventional method, using ligatures as the main hemostatic method. Main outcome measures were operating time, postoperative serum calcium level, palsy of the recurrent laryngeal nerve, and amount of intraoperative and postoperative bleeding. Possible bias that could have been caused by imbalance between treatment groups for surgeon experience was tested by two-way analysis of covariance. RESULTS: Thirty-six patients were randomized, 19 to the UAS and 17 to the conventional group. Mean operating time was 99.1 minutes in the UAS group and 134.9 minutes in the conventional group. The average savings in operating time with the UAS was thus 35.8 minutes. There was no difference in complications between the groups. The estimated savings in operating time would have been 1.66 times that observed in this study if the groups had been unbalanced with reference to surgeon experience. CONCLUSION: The UAS is a usable device in total thyroidectomies and lobectomies. PMID- 10714625 TI - Insular and anaplastic carcinoma of the thyroid: a 45-year comparative study at a single institution and a review of the significance of p53 and p21. AB - OBJECTIVE: To analyze the clinicopathologic features of a large cohort of patients with insular or anaplastic carcinomas treated at a single institution. SUMMARY BACKGROUND DATA: Insular and anaplastic carcinomas of the thyroid, although uncommon, have more aggressive clinical behavior than well differentiated carcinomas of the thyroid. In the literature, the incidence and features of these carcinomas have not been fully characterized. METHODS: The authors reclassified 740 primary thyroid carcinomas diagnosed and treated between January 1, 1954, and December 30, 1998, to select those with features that met the histologic criteria of insular or anaplastic carcinoma. The clinicopathologic features of these carcinomas were studied and compared. The expression of p53 and p21 in these tumors was analyzed by immunohistochemistry. RESULTS: Twenty-two patients (5 men, 17 women) with insular carcinoma and 38 patients (7 men, 31 women) with anaplastic carcinoma were found. Patients with insular carcinomas were younger (mean age 45 vs. 70 years) and had smaller tumors than those with anaplastic carcinomas (mean diameter 5 vs. 8 cm). Insular carcinomas were commonly mislabeled as other histologic subtypes, whereas anaplastic carcinomas might be overdiagnosed on pathologic examination. A history of longstanding goiter (>10 years) was noted in 27% of patients with insular carcinoma and 24% of patients with anaplastic carcinomas. Concomitant well-differentiated carcinomas of the thyroid were noted in 59% of patients with insular carcinoma and 39% of patients with anaplastic carcinoma. In anaplastic carcinomas, 13% of patients had concomitant insular carcinoma. Calcification or bone was noted in the stroma of 23% of patients with insular carcinomas and 47% of those with anaplastic carcinomas. The 10-year survival rates for patients with insular carcinoma and anaplastic carcinoma were 42% and 3%, respectively. Distant metastases were seen in 32% of patients with insular carcinoma and in 47% of patients with anaplastic carcinomas. In both types of carcinomas, metastatic tumors were often seen in bone and lung. Distant metastases were noted in a variety of organs in anaplastic carcinomas. In insular carcinoma, neither p53 nor p21 expression was present. In anaplastic carcinoma, p53 and p21 expression was identified in 69% and 3%, respectively. Concomitant expression of p53 and p21 was noted in one tumor. CONCLUSIONS: Insular carcinoma and anaplastic carcinoma had distinctive clinicopathologic features, and recognition of these histologic variants is important for better management of these tumors in the future. p53 overexpression might have a role in dedifferentiation from insular carcinoma to anaplastic carcinoma. PMID- 10714626 TI - Standardization of surgeon-controlled variables: impact on outcome in patients with acute cholecystitis. AB - OBJECTIVE: To examine the effect of standardization of surgeon-controlled variables on patient outcome after cholecystectomy for two cohorts of patients with acute cholecystitis (AC). SUMMARY BACKGROUND DATA: Laparoscopic cholecystectomy (LC), when performed efficiently and safely, offers patients with AC a more rapid recovery and decreases the length of stay, thus reducing the health care utilization. Numerous studies have focused on the characteristics of patients with AC that may predict the conversion of LC to open cholecystectomy. However, analysis of these factors offers little insight for improving the outcome of patients with AC, because patient-controlled variables are difficult to influence. In the present study, treatment variables that were under the surgeon's control were standardized and the effects of these changes on the outcome of patients with AC were quantified. METHODS: Beginning in August 1997, a standardized treatment protocol was initiated for patients with suspected AC. LC was initiated as early as practical from the time of admission. All operations were performed in a specially equipped and staffed laparoscopic surgery suite, and all patients were supervised by one of two attending surgeons with a special interest in laparoscopic interventions. Two cohorts of patients with AC were retrospectively analyzed: 39 patients from the 12 months before initiation of this protocol (period 1) and 49 patients from the 12 months after its inception (period 2). Medical records were reviewed for demographic, perioperative, and outcome data. Surgical reports were reviewed to ascertain the reason for conversion and whether laparoscopic technical modifications were used. RESULTS: No significant difference was noted between the groups with regard to patient demographics, clinical presentation, or radiologic or laboratory parameters. After protocol initiation, patients received definitive treatment closer to the time of admission and had a greater percentage of laparoscopically completed cholecystectomies. Furthermore, the patients in period 2 had a significantly decreased postoperative length of stay and hospital charges than the earlier ones. Complications were infrequent and not significantly different between the groups. Two or more laparoscopic technical modifications were used in 95% of the successful LCs during period 2 versus 33.3% during period 1. CONCLUSIONS: By controlling when, where, and by whom LC for AC was performed, the authors have significantly improved the percentage of cholecystectomies that were completed laparoscopically. This has led to improved outcomes and lower hospital charges for patients with AC at this municipal hospital. PMID- 10714627 TI - Local excision of rectal cancer without adjuvant therapy: a word of caution. AB - OBJECTIVE: To evaluate the results of local excision alone for the treatment of rectal cancer, applying strict selection criteria. BACKGROUND DATA: Several retrospective studies have demonstrated that tumor control in properly selected patients with rectal cancer treated locally is comparable to that observed after radical surgery. Although there is a consensus regarding the need for patient selection for local excision, the specific criteria vary among centers. METHODS: The authors reviewed 82 patients with T1 (n = 55) and T2 (n = 27) rectal cancer treated with transanal excision only during a 10-year period. At pathologic examination, all tumors were localized to the rectal wall, had negative excision margins, were well or moderately differentiated, and had no blood or lymphatic vessel invasion, nor a mucinous component. End points were local and distant tumor recurrence and patient survival. RESULTS: Ten of the 55 patients with T1 tumors (18%) and 10 of the 27 patients with T2 tumors (37%) had recurrence at 54 months of follow-up. Average time to recurrence was 18 months in both groups. Seventeen of the 20 patients with local recurrence underwent salvage surgery. The survival rate was 98% for patients with T1 tumors and 89% for patients with T2 tumors. Preoperative staging by endorectal ultrasound did not influence local recurrence or tumor-specific survival. CONCLUSION: Local excision of early rectal cancer, even in the ideal candidate, is followed by a much higher recurrence rate than previously reported. Although most patients in whom local recurrence develops can be salvaged by radical resection, the long-term outcome remains unknown. PMID- 10714628 TI - Pathophysiologic role of oxygen free radicals in acute pancreatitis: initiating event or mediator of tissue damage? AB - BACKGROUND AND OBJECTIVE: Oxidative stress is an important factor in the pathogenesis of acute pancreatitis, as shown in vivo by the beneficial effects of scavenger treatment and in vitro by the potential of free radicals to induce acinar cell damage. However, it is still unclear whether oxygen free radicals (OFR) act only as mediators of tissue damage or represent the initiating event in acute pancreatitis in vivo as well. In the present study the authors aimed to address this issue in an experimental set-up. MATERIALS AND METHODS: Two hundred male Wistar rats were randomly assigned to one of the following experimental groups. In two groups, acute necrotizing pancreatitis was induced by retrograde intraductal infusion of 3% sodium taurocholate. Through the abdominal aorta, a catheter was advanced to the origin of the celiac artery for continuous regional arterial (CRA) pretreatment with isotonic saline (NP-S group) or superoxide dismutase/catalase (NP-SOD/CAT group). In another group, oxidative stress was generated by CRA administration of xanthine oxidase and intravenous administration of hypoxanthine (HX/XOD group). Sham-operated rats received isotonic saline both arterially and intraductally. After observation periods of 5 and 30 minutes and 3 and 6 hours, the pancreas was removed for light microscopy and determination of reduced glutathione (GSH), oxidized glutathione (GSSG), conjugated dienes (CD), and malondialdehyde as a marker for OFR-induced lipid peroxidation as well as myeloperoxidase as a parameter for polymorphonuclear leukocyte accumulation. RESULTS: A significant decrease of GSH was paralleled by an increased ratio of GSSG per total glutathione and elevated CD levels after 5 minutes in the NP-S group versus the sham-operated group. Thereafter, the percentage of GSSG and GSH returned to normal levels until the 6-hour time point. After a temporary decrease after 30 minutes, CD levels increased again at 3 hours and were significantly higher at 6 hours in contrast to sham-operated rats. Myeloperoxidase levels were significantly elevated at 3 and 6 hours after pancreatitis induction. In contrast to NP-S rats, treatment with SOD/CAT significantly attenuated the changes in glutathione metabolism within the first 30 minutes and the increase of CDs after 6 hours. HX/XOD administration lead to changes in levels of GSH, GSSG, and CDs at 5 minutes as well as to increased myeloperoxidase levels at 3 hours; these changes were similar to those observed in NP-S rats. Acinar cell damage including necrosis was present after 5 minutes in both NP groups, but did not develop in HX/XOD rats. In addition, serum amylase and lipase levels did not increase in the latter group. SOD/CAT treatment significantly attenuated acinar cell damage and inflammatory infiltrate compared with NP-S animals during the later time intervals. CONCLUSION: OFRs are important mediators of tissue damage. However, extracellular OFR generation alone does not induce the typical enzymatic and morphologic changes of acute pancreatitis. Factors other than OFRs must be involved for triggering acute pancreatitis in vivo. PMID- 10714629 TI - Does an infected peripancreatic fluid collection or abscess mandate operation? AB - OBJECTIVE: To assess the treatment of peripancreatic fluid collections or abscess with percutaneous catheter drainage (PCD). SUMMARY BACKGROUND DATA: Surgical intervention has been the mainstay of treatment for infected peripancreatic fluid collections and abscesses. Increasingly, PCD has been used, with mixed results reported in the literature. METHODS: A retrospective chart review of 1993 to 1997 was performed on 82 patients at a tertiary care public teaching hospital who had computed tomography-guided aspiration for suspected infected pancreatic fluid collection or abscess. Culture results, need for subsequent surgical intervention, length of stay, and death rate were assessed. RESULTS: One hundred thirty-five aspirations were performed in 82 patients (57 male patients, 25 female patients) with a mean age of 40 years (range 17-68). The etiologies were alcohol (41), gallstones (32), and other (9). The mean number of Ranson's criteria was four (range 0-9). All patients received antibiotics. Forty-eight patients had evidence of pancreatic necrosis on computed tomography scan. Cultures were negative in 40 patients and positive in 42. Twenty-five of the 42 culture-positive patients had PCD as primary therapy, and 6 required subsequent surgery. Eleven patients had primary surgical therapy, and five required subsequent surgery. Six patients were treated with only antibiotics. The death rates were 12% for culture-positive patients and 8% for the entire 82 patients. CONCLUSIONS: Historically, patients with positive peripancreatic aspirate culture have required operation. This series reports an evolving strategy of reliance on catheter drainage. PCD should be considered as the initial therapy for culture positive patients, with surgical intervention reserved for patients in whom treatment fails. PMID- 10714630 TI - Fas and Fas-ligand expression in human pancreatic cancer. AB - OBJECTIVE: To investigate Fas and FasL expression in pancreatic tissues and cultured pancreatic cancer cell lines, and to assess the ability of anti-Fas antibodies to induce apoptosis. SUMMARY BACKGROUND DATA: Activation of the Fas receptor by Fas-ligand (FasL) results in apoptosis, and dysregulation of this pathway may contribute to abnormal cell proliferation. METHODS: Northern blotting and immunohistochemistry were used to compare Fas and FasL expression in normal and cancerous tissues. DNA 3'-OH end labeling was used to detect apoptotic cells. The effects of Fas activation on cell growth and signaling pathways were investigated in culture. RESULTS: Pancreatic cancers exhibited increased Fas RNA levels, whereas FasL mRNA levels were similar in both groups. Despite the colocalization of Fas and FasL in the cancer cells, an apoptotic signal was present in approximately 10% of these cells in only 2 of 16 cancer samples. Fas and FasL were coexpressed in all four cell lines, whereas Fas-associated phosphatase 1 was below the level of detection in all cell lines. Only COLO-357 cells underwent apoptosis after Fas activation. Apoptosis was associated with enhanced activation of jun kinase (JNK) and p38 mitogen-activated protein kinase (MAPK). In the presence of actinomycin D, Fas antibody also induced apoptosis in the other three cell lines. CONCLUSIONS: These results suggest that pancreatic cancer cells are resistant to Fas-mediated apoptosis by mechanisms excluding receptor downregulation or Fas-associated phosphatase upregulation and raise the possibility that Fas-mediated apoptosis may be dependent on the activation of the JNK/p38 MAPK pathway in these cells. PMID- 10714631 TI - Clinical value of [18-F]] fluorodeoxyglucose positron emission tomography imaging in soft tissue sarcomas. AB - OBJECTIVE: To evaluate positron emission tomography (PET) using 2-fluoro-2-deoxy D-glucose (FDG) for clinical application in soft tissue sarcomas. SUMMARY AND BACKGROUND DATA: FDG PET is a promising noninvasive method for the preoperative assessment of soft tissue sarcomas and may complement radiologic tomography. METHODS: Data from 50 consecutive patients with 59 masses, either suspicious for primary or locally recurrent soft tissue sarcoma, were prospectively gathered. The semiquantitative FDG uptake (standardized uptake values [SUVs]) was calculated in tumor and normal tissue (muscle). Histopathology of surgical specimens and follow-up data were used as control criteria. RESULTS: In primary soft tissue sarcomas, PET displayed a sensitivity of 91% and a specificity of 88%. Local recurrence was detected with a sensitivity of 88% and a specificity of 92%. All intermediate-grade and high-grade soft tissue sarcomas (primary and locally recurrent) were visualized with a precise differentiation from muscle. Fifty percent of the low-grade sarcomas showed an FDG uptake equivalent to muscle (false-negative results in one primary and three recurrent soft tissue sarcomas). Benign soft tissue tumors (e.g., lipoma, leiomyoma, ganglion) did not accumulate FDG. Inflammation resulted in an increased FDG uptake. The semiquantitative FDG uptake (SUVs) correlated with tumor grade but not with size and histologic type. CONCLUSION: High-grade and intermediate-grade soft tissue sarcomas are amenable to PET imaging, whereas low-grade lesions may not be depicted. SUVs for FDG correlate with tumor grade in soft tissue sarcomas. Benign soft tissue tumors are differentiated from higher-grade soft tissue sarcomas. These data show that FDG PET can complement preoperative radiologic assessment for soft tissue sarcomas and that FDG-PET is a powerful diagnostic tool for detecting high-grade and intermediate-grade local recurrence. PMID- 10714632 TI - Interaction of platelet activating factor, reactive oxygen species generated by xanthine oxidase, and leukocytes in the generation of hepatic injury after shock/resuscitation. AB - OBJECTIVE: To evaluate the putative relation of platelet activating factor (PAF), xanthine oxidase, reactive oxidants, and leukocytes in the pathogenesis of hepatic injury after shock/resuscitation (S/R) in vivo. BACKGROUND: Reactive oxygen metabolites generated by xanthine oxidase at reperfusion have been found to trigger postischemic injury in many organs, including the liver. However, the precise linear sequence of the mechanism of consequent hepatic injury after S/R remains to be characterized. METHODS: Unheparinized male rats were bled to a mean blood pressure of 45 +/- 3 mmHg. After 2 hours of shock, they were resuscitated by reinfusion of shed blood (anticoagulated with citrate-phosphate-dextrose) and crystalloid and observed for the next 6 or 24 hours. RESULTS: S/R caused the oxidation of hepatic glutathione and generated centrolobular leukocyte accumulation at 6 hours, followed by predominantly centrolobular hepatocellular injury at 24 hours. Each of these components was attenuated by PAF inhibition with WEB 2170, xanthine oxidase inhibition with allopurinol, antioxidant treatment with N-acetylcysteine, or severe leukopenia induced by vinblastine. In each case, the degree of leukocyte accumulation at 6 hours correlated with the hepatocellular injury seen at 24 hours. However, xanthine oxidase inhibition with allopurinol failed to attenuate further the small level of residual hepatocellular injury seen in leukopenic rats. CONCLUSION: These findings suggest that reactive oxidants generated by xanthine oxidase at reperfusion, stimulated by PAF, mediate hepatocellular injury by triggering leukocyte accumulation, primarily within the centrolobular sinusoids. PMID- 10714633 TI - Inhibition of tyrosine kinase signaling after trauma-hemorrhage: a novel approach for improving organ function and decreasing susceptibility to subsequent sepsis. AB - OBJECTIVE: To determine whether administration of a tyrosine kinase inhibitor after trauma-hemorrhage has any beneficial effects on cardiovascular parameters and hepatocellular function and on survival rate after subsequent sepsis. BACKGROUND: Increased inflammatory cytokine release and concomitant activation of intracellular signaling pathways contributes to multiple organ dysfunction and increased susceptibility to subsequent sepsis after severe hemorrhagic shock. METHODS: Male Sprague-Dawley rats underwent a midline laparotomy (i.e., soft tissue trauma induced) and were then bled to and maintained at a mean arterial pressure of 40 mmHg until 40% of the maximal shed blood volume was returned in the form of Ringer's lactate. The rats were then resuscitated with four times the shed blood volume in the form of Ringer's lactate during a 60-minute period. A tyrosine kinase inhibitor, AG 556 (7.5 mg/kg), or vehicle was administered intraperitoneally at the middle of resuscitation. At 24 hours after resuscitation, various in vivo parameters such as heart performance, cardiac index, and hepatocellular function (i.e., the maximum velocity and the overall efficiency of indocyanine green clearance) were determined. Phosphorylation state of the mitogen-activated protein kinases p44/42 and p38 in the liver was assessed by Western blot analysis. In additional groups of rats, sepsis was induced by cecal ligation and puncture at 20 hours after hemorrhage. The necrotic cecum was excised 10 hours thereafter, and the survival rate was monitored for a period of 10 days. RESULTS: AG 556 treatment restored the depressed cardiovascular and hepatocellular functions after trauma-hemorrhage and resuscitation, which was associated with reduced phosphorylation of mitogen-activated protein kinases p44/42 and p38. Moreover, treatment with AG 556 significantly increased the survival rate of rats after trauma-hemorrhage and induction of subsequent sepsis compared with vehicle-treated rats. CONCLUSION: Inhibition of tyrosine kinase signaling after trauma-hemorrhage may represent a novel therapeutic approach for improving organ functions and decreasing the death rate from subsequent sepsis under such conditions. PMID- 10714634 TI - Insulin-like growth factor I in combination with insulin-like growth factor binding protein 3 affects the hepatic acute phase response and hepatic morphology in thermally injured rats. AB - OBJECTIVE: To modulate the hepatic acute phase response after a thermal injury by the administration of insulin-like growth factor I (IGF-I) in combination with its principal binding protein 3 (IGFBP-3). SUMMARY BACKGROUND DATA: The hepatic acute phase response is a cascade of events initiated to restore homeostasis after trauma; however, a prolonged response contributes to multiorgan failure, hypermetabolism, complications, and death. Although IGF-1 has been shown to improve cell recovery and play a major role in liver regeneration, its effect on the hepatic acute phase response is not known. METHODS: Sprague-Dawley rats (56 males) received a 60% total body surface area third-degree scald burn and were randomly divided to receive either rhIGF-I/BP-3 (10 mg/kg/day given subcutaneously) or saline (control). Rats were killed on postburn days 1, 2, 5, and 7 and serum glucose, electrolytes, acute phase reactant proteins, tumor necrosis factor alpha, interleukin 1 beta, interleukin 6, and rat and human serum IGF-I and IGFBP-3 were measured. Hepatic protein concentrations, hepatocyte proliferation, and hepatocyte apoptosis were determined. RESULTS: No hypoglycemia or electrolyte imbalance could be shown in rats receiving the growth factor complex compared with saline. rhIGF-I/BP-3 increased serum protein on postburn days 2 and 7, albumin on days 5 and 7, and transferrin on days 1, 5, and 7, and decreased haptoglobin and alpha1-acid glycoprotein on postburn days 5 and 7 compared with controls. IGF-I/ BP-3 had no effect on type II acute phase proteins. Rats receiving IGF-I/BP-3 had lower serum levels of interleukin 1 beta and tumor necrosis factor alpha on the first day after burn compared with controls, whereas serum levels of interleukin 6 did not change. rhIGF-I/BP-3 significantly increased total liver protein content on postburn days 1, 2, 5, and 7 compared with controls. IGF-I/BP-3 increased hepatocyte proliferation and decreased hepatocyte apoptosis versus controls. CONCLUSION: In combination with its principal binding protein, rhIGF-I decreases the proinflammatory cytokines interleukin 1 beta and tumor necrosis factor alpha, followed by a decrease in type I acute phase proteins. IGF-I/BP-3 had no effect on interleukin 6 and type II acute phase proteins. Decreases in acute phase protein and proinflammatory cytokine synthesis were associated with increases in constitutive hepatic proteins, total liver protein content, and hepatocyte proliferation. IGF-I/BP-3 attenuates the hypermetabolic response after thermal injury and may improve the clinical outcome. PMID- 10714635 TI - Simultaneous pancreas-kidney transplantation and living related donor renal transplantation in patients with diabetes: is there a difference in survival? AB - OBJECTIVE: To compare the outcome of simultaneous pancreas-kidney transplantation (SPK) and living related donor renal transplantation (LRD) in patients with diabetes. SUMMARY BACKGROUND DATA: It remains unanswered whether diabetic patients with end-stage renal failure are better served by LRD or SPK. METHODS: Using a longitudinal database, data from all diabetic patients receiving LRD or cadaveric renal transplants or SPKs from January 1986 through January 1996 were analyzed. Patient and graft survival, early graft function, and the cause of patient and graft loss were compared for 43 HLA-identical LRDs, 87 haplotype identical LRDs, 379 SPKs, and 296 cadaveric renal transplants. RESULTS: The demographic composition of the SPK and LRD groups were similar, but because of less strict selection criteria in the cadaveric transplant group, patients were 10 years older, more patients received dialysis, and patients had been receiving dialysis longer before transplantation. Patient survival was similar for the SPK and LRD groups but was significantly lower for the cadaveric renal transplant group. Similarly, there was no difference in graft survival between SPK and LRD recipients, but it was significantly lower for recipients in the cadaveric renal transplant group. Delayed graft function was significantly more common in the cadaveric renal transplant group. Discharge creatinine, the strongest predictor of patient and graft survival, was highest in the SPK group and lowest in the HLA identical LRD group. The rate of rejection within the first year was greatest in SPK patients (77%), intermediate in the haplotype-identical LRD and cadaveric transplant groups (57% and 48%, respectively), and lowest (16%) in the HLA identical LRD group. Cardiovascular disease was the primary cause of death for all groups. Acute rejection, chronic rejection, and death with a functioning graft were the predominant causes of graft loss. CONCLUSIONS: This study demonstrates that there was no difference in patient or graft survival in diabetic patients receiving LRD or SPK transplants. However, graft and patient survival rates in diabetic recipients of cadaveric renal transplants were significantly lower than in the other groups. PMID- 10714637 TI - Physician impact on the total cost of care. AB - BACKGROUND AND OBJECTIVES: Physicians' efforts at cost containment focus on decreased resource utilization and reduced length of stay. Although these efforts appear to be appropriate, little data exist to gauge their success. As such, the goal of this study is to determine trauma service cost allocations and how this information can help physicians to contain costs. MATERIALS AND METHODS: The authors analyzed the costs for 696 trauma admissions at a level I trauma center for fiscal year 1997. Data were obtained from the hospital costing system. Costs analyzed were variable direct, fixed direct, and Indirect costs. Together, the fixed and indirect costs are referred to as "hospital overhead." Total Cost equals variable direct plus fixed direct plus indirect costs. RESULTS: The mean variable, fixed, and indirect costs per patient were $7,998, $3,534, and $11,086, respectively. Mean total cost per patient was $22,618. CONCLUSION: The 35% variable direct cost represents the percentage of total cost that is typically under the immediate influence of physicians, in contrast to the 65% of total cost over which physicians have little control. Physicians must gain a better understanding of cost drivers and must participate in the operations and allocations of institutional fixed direct and indirect costs if the overall cost of care is to be reduced. PMID- 10714636 TI - Cell death in human lung transplantation: apoptosis induction in human lungs during ischemia and after transplantation. AB - OBJECTIVE: To examine the presence and extent of apoptosis as well as the affected cell types in human lung tissue before, during, and after transplantation. SUMMARY BACKGROUND DATA: Apoptosis has been described in various human and animal models of ischemia-reperfusion injury, including heart, liver, and kidney, but not in lungs. Therefore, the presence of apoptosis and its role in human lungs after transplantation is not clear. METHODS: Lung tissue biopsies were obtained from 20 consecutive human lungs for transplantation after cold ischemic preservation (1-5 hours), after warm ischemia time (during implantation), and 30, 60, and 120 minutes after graft reperfusion. To detect and quantify apoptosis, fluorescent in situ end labeling of DNA fragments (TUNEL assay) was used. Electron microscopy was performed to verify the morphologic changes consistent with apoptosis and to identify the cell types, which were lost by apoptosis. RESULTS: Almost no evidence of apoptosis was found in specimens after immediate cold and warm ischemic periods. Significant increases in the numbers of cells undergoing apoptosis were observed after graft reperfusion in a time-dependent manner. The mean fraction of apoptotic cells at 30, 60, and 120 minutes after graft reperfusion were 16.6%, 22.1%, and 34.9% of total cells, respectively. Most of the apoptotic cells appeared to be alveolar type II pneumocytes, as confirmed by electron microscopy. CONCLUSIONS: Programmed cell death (apoptosis) appears to be a significant type of cell loss in human lungs after transplantation, and this may contribute to ischemia-reperfusion injury during the early phase of graft reperfusion. This cell loss might be responsible for severe organ dysfunction, which is seen in 20% of patients after lung transplantation. Therefore, this work is of importance to surgeons for the future development of interventions to prevent cell death in transplantation. PMID- 10714638 TI - The search for an ideal method of abdominal fascial closure: a meta-analysis. AB - BACKGROUND AND OBJECTIVE: The ideal suture for abdominal fascial closure has yet to be determined. Surgical practice continues to rely largely on tradition rather than high-quality level I evidence. The authors conducted a systematic review and meta-analysis of randomized controlled trials to determine which suture material and technique reduces the odds of incisional hernia. METHODS: MEDLINE and Cochrane Library databases were searched for articles in English published from 1966 to 1998 using the keywords "suture", "abdomen/surgery", and "randomized controlled trials". Randomized controlled trials, trials of adult patients, and trials with a Jadad Quality Score of more than 3, comparing suture materials, technique, or both, were included. Two independent reviewers critically appraised study quality and extracted data. The reviewers were masked to the study site, authors, journal, and date to minimize bias. The primary outcome was postoperative incisional hernia. Secondary outcomes included wound dehiscence, infection, wound pain, and suture sinus formation. RESULTS: The occurrence of incisional hernia was significantly lower when nonabsorbable sutures were used. Suture technique favored nonabsorbable continuous closure. Suture sinuses and wound pain were significantly lower when absorbable sutures were used. There were no differences in the incidence of wound dehiscence or wound infection with respect to suture material or method of closure. Subgroup analyses of individual sutures showed no significant difference in incisional hernia rates between polydioxanone and polypropylene. Polyglactin showed an increased wound failure rate. CONCLUSIONS: Abdominal fascial closure with a continuous nonabsorbable suture had a significantly lower rate of incisional hernia. The ideal suture is nonabsorbable, and the ideal technique is continuous. PMID- 10714639 TI - Repair of pectus excavatum deformities: 30 years of experience with 375 patients. AB - OBJECTIVE: To review the surgical experience with pectus excavatum chest deformities at UCLA Medical Center during a 30-year period. BACKGROUND: Pectus excavatum is a relatively common malformation that is often symptomatic; however, children's physicians often do not refer patients for surgical correction. METHODS: Hospital records from 375 patients who underwent repair of pectus excavatum deformities between 1969 and 1999 were reviewed. Decrease in stamina and endurance during exercise was reported by 67%; 32% had frequent respiratory infections, 8% had chest pain, and 7% had asthma. The mean pectus severity score (width of chest divided by distance between posterior surface of sternum and anterior surface of spine) was 4.65 (normal chest = 2.56). All patients had marked cardiac deviation into the left chest. Repair was performed with subperiosteal resection of the abnormal cartilages, transverse wedge osteotomy of the anterior sternum, and internal support with a steel strut for 6 months. Repair was performed on 177 children before age 11 years; 38 adults with severe symptoms underwent repair. RESULTS: The mean hospital stay was 3.1 days. With a mean follow-up of 12.6 years, all patients with preoperative respiratory symptoms, exercise limitation, and chest pain experienced improvement. Vital capacity increased 11% (mean) within 9 months in 35 patients evaluated. There were no deaths. Complications included hypertrophic scar formation (35), atelectasis (12), pleural effusion (13), recurrent sternal depression (5), and pericarditis (3). More than 97% had a very good or excellent result. CONCLUSION: Pectus excavatum deformities can be repaired with a low rate of complications, a short hospital stay, and excellent long-term physiologic and cosmetic results. PMID- 10714640 TI - Mammalian microsporidiosis. AB - The phylum Microspora contains a diverse group of single-celled, obligate intracellular protozoa sharing a unique organelle, the polar filament, and parasitizing a wide variety of invertebrate and vertebrate animals, including insects, fish, birds, and mammals. Encephalitozoon cuniculi is the classic microsporidial parasite of mammals, and encephalitozoonosis in rabbits and rodents has been and continues to be recognized as a confounding variable in animal-based biomedical research. Although contemporary research colonies are screened for infection with this parasite, E. cuniculi remains a cause of morbidity and mortality in pet and conventionally raised rabbits. In addition, E. cuniculi is a potential pathogen of immature domestic dogs and farm-raised foxes. The recent discovery and identification of Encephalitozoon intestinalis, Encephalitozoon hellem, and Enterocytozoon bieneusi, in addition to E. cuniculi, as opportunistic pathogens of humans have renewed interest in the Microspora. Veterinary pathologists, trained in the comparative anatomy of multiple animal species and infectious disease processes, are in a unique position to contribute to the diagnosis and knowledge of the pathogenesis of these parasitic diseases. This review article covers the life cycle, ultrastructure, and biology of mammalian microsporaidia and the clinical disease and lesions seen in laboratory and domestic animals, particularly as they relate to Encephalitozoon species. Human microsporidial disease and animal models of human infection are also addressed. Often thought of as rabbit pathogens of historical importance, E. cuniculi and the related mammalian microsporidia are emerging as significant opportunistic pathogens of immunocompromised individuals. PMID- 10714641 TI - Expression of viral proteins in feline leukemia virus-associated enteritis. AB - Fourteen cases of feline leukemia virus (FeLV)-associated enteritis were immunohistologically examined for the expression of FeLV proteins gp70, p27, and p15E in the jejunum, mesenteric lymph nodes, spleen, and bone marrow. Results were compared with those of FeLV-infected cats without intestinal alterations. Other viral infections and specific bacterial, fungal, and parasitic infections were excluded by standard microbiologic methods, histopathology, immunohistology, and in situ hybridization. In FeLV-associated enteritis, FeLV gp70 and p15E were strongly expressed in intestinal crypt epithelial cells. In contrast, FeLV positive cats without intestinal alterations showed only faint staining for gp70 and p15E and comparatively strong p27 expression in these cells. Findings suggest a direct relation between FeLV infection and alterations in intestinal crypt epithelial cells that may be attributed to the envelope proteins gp70 and p15E and/or their precursor protein. Distinct similarities to the intestinal changes in the experimentally induced FeLV-feline AIDS syndrome are obvious, suggesting that naturally occurring feline AIDS variants may be responsible for FeLV associated enteritis. PMID- 10714642 TI - Subcutaneous atypical mycobacteriosis in captive tiger quolls (Dasyurus maculatus). AB - From July 1989 to October 1998, 9/37 (24%) adult captive tiger quolls (Dasyurus maculatus) were diagnosed with atypical mycobacterial infection involving the subcutis and skin. Females were more often affected than males (seven females, two males). Grossly, lesions presented as focal thickenings, plaques, and abscesses within the subcutis, often with fistulous tracts. The subcutis and skin overlying cervical and thoracic regions were the primary sites of infection. Cytology of subcutaneous impression smears from all nine affected tiger quolls revealed pyogranulomatous inflammation admixed with several acid-fast bacilli. Histologically, all tiger quolls had nodular to diffuse pyogranulomatous panniculitis and cellulitis. Small numbers of acid-fast bacilli were noted histologically in 7/9 (78%) animals. Skin cultures from seven tiger quolls were positive for one or more different Runyon group IV mycobacteria. The disease described in these tiger quolls is similar to subcutaneous atypical mycobacteriosis of humans and domestic animals. PMID- 10714643 TI - Pathogenesis of porcine reproductive and respiratory syndrome virus-induced increase in susceptibility to Streptococcus suis infection. AB - Eighty 3-week-old crossbred pigs were randomly assigned to six groups (13-14 pigs/group). Group 1 pigs served as uninoculated controls, group 2 pigs were inoculated intranasally (i.n.) with Streptococcus suis serotype 2, group 3 pigs were inoculated i.n. with a modified live porcine reproductive and respiratory syndrome virus (PRRSV) vaccine, group 4 pigs were inoculated i.n. with the same vaccine and with S. suis, group 5 pigs were inoculated i.n. with VR-2385 (a high virulence strain of PRRSV), and group 6 pigs were inoculated i.n. with VR-2385 and S. suis. Pigs exposed to both PRRSV and S. suis were inoculated with PRRSV 7 days prior to S. suis inoculation. The pigs were 26 days old when inoculated with S. suis. Respiratory disease was significantly more severe in groups 5 and 6. Mortality rate was the highest in group 6 (87.5%). This rate was significantly higher than that observed in all other groups except group 4 (37.5%). The mortality rate in group 2, inoculated with S. suis alone, was 14.3%. No pigs from groups 1, 3, or 5 died prior to the scheduled necropsies at 10 and 28 days postinoculation with PRRSV (DPI). To study the effect of PRRSV and/or S. suis on pulmonary clearance by pulmonary intravascular macrophages, six pigs from each group were intravenously infused with 3% copper phthalocyanine tetrasulfonic acid in saline prior to necropsy at 10 DPI. Mean copper levels in the lungs of pigs in groups 2, 5, and 6 were significantly lower than those in control pigs. The mean percentage of lung tissue grossly affected by pneumonia at 10 DPI was 0%, 1%, 0%, 3%, 64%, and 62% for groups 1-6, respectively. Both gross and microscopic interstitial pneumonia lesions were significantly more severe in the VR2385 inoculated groups (5 and 6). PRRSV was isolated from bronchoalveolar lavage fluid collected at necropsy from 100% of the pigs in groups 5 and 6, 71.4% of pigs in group 4, 38.5% of pigs in group 3, and none of the pigs in groups 1 or 2. Streptococcus suis serotype 2 was cultured from the internal tissues of 7.7%, 28.6%, and 78.6% of the pigs in groups 2, 4, and 6, respectively. Streptococcus suis serotype 2 was isolated from whole blood at necropsy from 7.7%, 35.7%, and 78.6% of pigs in groups 2, 4, and 6, respectively. Significantly more pigs in group 6 had S. suis isolated from whole blood and internal tissues. In summary, both high-virulence PRRSV and S. suis decreased copper clearance, and the incidence of isolation of S. suis and PRRSV was higher in dually inoculated pigs. PRRSV-induced suppression of pulmonary intravascular macrophage function may in part explain PRRSV-associated increased susceptibility to S. suis infection. PMID- 10714644 TI - Lysosomal storage disease caused by Sida carpinifolia poisoning in goats. AB - A neurologic disease characterized by ataxia, hypermetria, hyperesthesia, and muscle tremors of the head and neck was observed for 2 years in a flock of 28 Anglo-Nubian and Saanen goats on a farm with 5 ha of pasture. Six newborns died during the first week of life, and five abortions were recorded. The predominant plant in the pasture was Sida carpinifolia. The disease was reproduced experimentally in two goats by administration of this plant. Three goats with spontaneous disease and the two experimental animals were euthanatized and necropsied. No significant gross lesions were observed. Fragments of several organs, including the central nervous system, were processed for histopathology. Small fragments of the cerebellar cortex, liver, and pancreas of two spontaneously poisoned goats and two experimentally poisoned goats were processed for electron microscopy. Multiple cytoplasm vacuoles in hepatocytes, acinar pancreatic cells, and neurons, especially Purkinje cells, were the most striking microscopic lesions in the five animals. Ultrastructural changes included membrane-bound vacuoles in hepatocytes, Kupffer cells, acinar pancreatic cells, Purkinje cells, and the small neurons of the granular cell layer of the cerebellum. Paraffin-embedded sections of the cerebellum and pancreas were submitted for lectin histochemical analysis. The vacuoles in different cerebellar and acinar pancreatic cells reacted strongly to the following lectins: Concanavalia ensiformis, Triticum vulgaris, and succinylated Triticum vulgaris. The pattern of staining, analyzed in Purkinje cells and acinar pancreatic cells coincides with results reported for both swainsonine toxicosis and inherited mannosidosis. PMID- 10714645 TI - Clinical and pathologic features of oligodendrogliomas in two cats. AB - Two oligodendrogliomas in two domestic cats involved mainly the rostral brain stem, midbrain, fourth ventricle, and cerebellum. Both cats were aged neutered males presenting with clinical neurologic deficits suggestive of a brain stem lesion. Magnetic resonance imaging of both tumors demonstrated lesions with a pattern of heterogeneous contrast enhancement and multifocal lesions in one cat. Routine cerebrospinal fluid analysis was normal in one cat and suggestive of an inflammatory disease in the other. Oligodendroglioma cells were seen in cytospin preparations of cerebrospinal fluid from both cats. In each cat, the tumors occurred intraventricularly in the midbrain and fourth ventricle with aggressive intraparenchymal infiltration. There was extensive growth into the basilar subarachnoid space of the midbrain and brain stem in one cat. One tumor was well differentiated, and the other was an anaplastic subtype. Immunostaining for several myelin- and oligodendroglia-specific antigens was negative with formalin fixed tumors and with unfixed frozen samples from one cat. In both tumors, component cells of the intratumoral vascular proliferations were positive for human von Willebrand factor VIII antigen or smooth muscle actin. Immunocytochemical reactivity for glial fibrillary acidic protein identified both reactive astrocytes and a subpopulation of minigemistocytes in both tumors. Ultrastructurally, the tumor cells were unremarkable except for their prominent desmosomal junctions and paucity of microtubules. PMID- 10714646 TI - Distribution of cells immunopositive for AM-3K, a novel monoclonal antibody recognizing human macrophages, in normal and diseased tissues of dogs, cats, horses, cattle, pigs, and rabbits. AB - The monoclonal antibody AM-3K, which was developed using human pulmonary macrophages as the immunogen, immunocytochemically labels most human macrophages except for blood monocytes and dendritic cell populations. AM-3K also shows cross reactivity in some animal species. To evaluate the usefulness of AM-3K, the present study investigated the detailed distribution of AM-3K-immunopositive macrophages in normal and diseased tissues of dogs, cats, horses, cattle, pigs, and rabbits. Zamboni's solution-fixed, paraffin-embedded sections were the most available for the immunocytochemistry with AM-3K. In all animal species examined, AM-3K labeled most macrophages in splenic red pulp, lymph node sinuses and thymus, and tissue macrophages in the interstitium of various organs and sites such as the kidneys, lungs, heart, pancreas, intestines, and skin. Alveolar macrophages and perivascular microglial cells were also immunoreactive for AM-3K. Interestingly, Kupffer cells of dogs, cats, and horses were labeled for AM-3K, but those of cattle, pigs, and rabbits were not. Furthermore, in tumor tissues and inflammatory lesions such as liver fibrosis and encephalomalacia that were obtained from dogs, infiltrating macrophages were stained with AM-3K, but not all infiltrating macrophages reacted to AM-3K. In addition, only 30-50% of pulmonary and peritoneal macrophages obtained from cats and dogs were reactive for AM-3K. AM-3K did not react with blood monocytes, dendritic cell populations, and osteoclasts. These observations indicate that AM-3K specifically labels most exudate and tissue macrophages in the animal species examined. However, the expression of antigens recognized by AM-3K on macrophages may be dependent on differential maturation stages or different functions evoked by some conditions. AM-3K immunoreaction products were seen on the cytoplasmic membrane of macrophages by immunoelectron microscopy. AM-3K would be useful for detection of macrophage populations in the animal species examined here. PMID- 10714647 TI - Cellular proliferative and telomerase activity in canine mammary gland tumors. AB - In canine mammary tumors, we examined the telomerase activity, proliferative activity by proliferative cell nuclear antigen (PCNA) immunohistochemistry, and percentage of apoptotic cells by the deoxynucleotidyl transferase-mediated dUTP biotin nick end-labeling (TUNEL) method. The relationship between these measures and histopathologic malignancy was also investigated. PCNA index was highest in malignant tumors (adenocarcinoma: 27.0%; malignant mixed tumor: 15.7%), followed by benign tumors (adenoma: 4.4%; benign mixed tumor: 5.3%), hyperplasia (2.1%), and normal mammary gland (0.9%). In adenoma and adenocarcinoma, papillary and solid types showing higher cellularity tended to have higher PCNA indices than did cystic and tubular types. Although the TUNEL index was <1% in all cases, the relationship between this measure and histopathologic diagnosis showed the same tendency as observed in PCNA immunostaining. Telomerase activity was detectable in all adenomas, benign mixed tumors, and adenocarcinomas examined. In contrast, all normal mammary glands, hyperplasias, and malignant mixed tumors were negative for telomerase. Relative telomerase activity (RTA) of adenocarcinoma (56.5) was significantly higher than that of adenoma (27.8) and benign mixed tumor (33.9), and a significant positive correlation (P < 0.001) was noted between RTA and PCNA index. No significant correlations were noted between either PCNA or TUNEL index and clinical features such as metastasis and tumor diameter. PCNA index and telomerase activity may be useful markers for judging malignancy of canine mammary tumors. PMID- 10714648 TI - Primary epitheliotropic T-cell lymphoma of the urinary bladder in a dog. AB - A 7-year-old, intact female mixed-breed dog was presented for evaluation of hematuria. Physical examination revealed a suprapubic mass. Ultrasonographic examination showed a large lobular mass occupying the urinary bladder. At the owners' request, the dog was euthanatized and a postmortem examination was performed. Necropsy confirmed the presence of a lobular mass of about 5- to 6-cm diameter protruding into the lumen of the bladder. Histologically, the mass was composed of a large number of atypical lymphoid cells in the lamina propria and mucosal epithelium. Immunohistochemically, the neoplastic cells expressed CD3 but not CD79alpha or keratin and vimentin, supporting a diagnosis of T-cell lymphoma. PMID- 10714649 TI - Choriocarcinoma and teratoma in the ovary of a cynomolgus monkey. AB - An ovarian choriocarcinoma was found in a 13-year-old cynomolgus monkey (Macaca fascicularis). The tumor was accompanied by a mature teratoma in the contralateral ovary. Histologically, the choriocarcinoma was characterized by nests of cells where cytotrophoblasts occupied the periphery with syncytiotrophoblasts at the center. Immunohistochemical staining for anti-human chorionic gonadotropin was positive in the syncytiotrophoblasts. The teratoma consisted of well-differentiated epidermal cells, sebaceous glands, hair follicles, cartilage, bone, and teeth. Choriocarcinoma metastases were in multiple organs. The concomitant development of choriocarcinoma and teratoma in the ovary is a consistent finding with the human counterparts of these lesions. PMID- 10714650 TI - Bilateral renal oncocytoma in a Greyhound dog. AB - A bilateral, locally invasive renal oncocytoma was diagnosed in a 10-year-old spayed female Greyhound dog. The diagnosis was based on positive staining of the tumor with the periodic acid-Schiff reaction prior to diastase treatment, on the immunohistochemical expression of cytoplasmic cytokeratin, and on the prominence of mitochondria in the tumor cells. PMID- 10714651 TI - Polysaccharide storage myopathy in Morgan, Arabian, and Standardbred related horses and Welsh-cross ponies. AB - Polysaccharide storage myopathy is an equine neuromuscular disorder characterized by accumulation of glycogen-related polysaccharide inclusions within skeletal muscle fibers. The pathologic criteria for diagnosis of this disorder are somewhat controversial; however, periodic acid-Schiff-positive, amylase-resistant inclusions are considered pathognomonic. Although these inclusions are most often found in affected horses related to the Quarter Horse, draft horse, and Warmblood breeds, this report describes these characteristic inclusions in muscle of five horses from nonrelated breeds (two Morgans, one Arabian, one Arabian x Thoroughbred, and one Standardbred) and two Welsh cross ponies. Affected horses had histories of recurrent exertional rhabdomyolysis, and one developed progressive weakness leading to increased recumbency. The affected ponies were part of an unrelated research project and had no apparent clinical signs. PMID- 10714652 TI - Aspirin benefit remains elusive in primary stroke prevention. PMID- 10714653 TI - Sporadic cases of possible genetic diseases: to test or not to test? PMID- 10714654 TI - Methods for discerning disease-modifying effects in Alzheimer disease treatment trials. PMID- 10714655 TI - Phantom limbs and neural plasticity. AB - The study of phantom limbs has received tremendous impetus from recent studies linking changes in cortical topography with perceptual experience. Systematic psychophysical testing and functional imaging studies on patients with phantom limbs provide 2 unique opportunities. First, they allow us to demonstrate neural plasticity in the adult human brain. Second, by tracking perceptual changes (such as referred sensations) and changes in cortical topography in individual patients, we can begin to explore how the activity of sensory maps gives rise to conscious experience. Finally, phantom limbs also allow us to explore intersensory effects and the manner in which the brain constructs and updates a "body image" throughout life. PMID- 10714656 TI - Human immunodeficiency virus-associated dementia. AB - It is clear that optimal control of HIV infection using cocktails of antiretrovirals has an important beneficial effect on the neurologic manifestations of HIV. Research is required to define the pathophysiology of HIV associated disease in the central nervous system, and to enhance delivery of therapy to this important compartment. Concurrently, trials of potentially neuroprotective agents are needed to optimize central nervous system therapy. No neuroprotective treatment to date has been successfully proven to be beneficial. However, progress has been very rewarding, with rapidly declining incidence of neurologic disease associated with dramatic improvement in HIV therapy. The prolonged life span of patients with HIV leaves the possibility that prevalence of HIV-associated neurologic disease might even increase in coming years. Therapy for HIV in the central nervous system compartment remains potentially the most challenging therapeutic frontier for HIV control. PMID- 10714657 TI - Aspirin for the primary prevention of stroke and other major vascular events: meta-analysis and hypotheses. AB - BACKGROUND: Aspirin therapy reduces stroke by about 25% for persons with atherosclerotic vascular disease, but the effect in those without clinically apparent vascular disease is distinctly different. OBJECTIVE: To define the effect of aspirin use on stroke and other major vascular events when given for primary prevention to persons without clinically recognized vascular disease. DATA SOURCES AND EXTRACTION: Systematic review of randomized clinical trials and large prospective observational cohort studies examining the relation between aspirin use and stroke in persons at low intrinsic risk. Studies were identified by a computerized search of the English-language literature. DATA SYNTHESIS: Five randomized trials of primary prevention included 52 251 participants randomized to aspirin doses ranging from 75 to 650 mg/d; the mean overall stroke rate was 0.3% per year during an average follow-up of 4.6 years. Meta-analysis revealed no significant effect on stroke (relative risk = 1.08; 95% confidence interval, 0.95 1.24) contrasting with a decrease in myocardial infarction (relative risk = 0.74; 95% confidence interval, 0.68-0.82). The lack of reduction of stroke by aspirin for primary prevention was incompatible with its protective effect against stroke in patients with manifest vascular disease (P = .001). Intracranial hemorrhage was increased by the regular use of aspirin (relative risk = 1.35; P = .03), similarly for both primary and secondary prevention. In 4 large observational studies, self-selected use of aspirin was consistently associated with higher rates of stroke. CONCLUSIONS: The effect of aspirin therapy on stroke differs between individuals based on the presence or absence of overt vascular disease, in contrast with the consistent reduction in myocardial infarction by aspirin therapy observed in all populations. We hypothesize that the effect of aspirin therapy on stroke for persons with major risk factors for vascular disease may be intermediate between a substantial decrease for those with manifest vascular disease and a possible small increase for healthy persons due to accentuated intracranial hemorrhage. When aspirin is given for primary prevention of vascular events, available data support using 75 to 81 mg/d. PMID- 10714658 TI - Frequency of the DYT1 mutation in primary torsion dystonia without family history. AB - BACKGROUND: Idiopathic torsion dystonia is a clinically and genetically heterogeneous movement disorder. A GAG deletion at position 946 of the DYT1 gene was the first mutation found, in early-onset dystonia, with an autosomal dominant transmission and reduced penetrance. OBJECTIVE: To evaluate the frequency of the DYT1 mutation in patients with idiopathic torsion dystonia but without a family history. DESIGN: Prospective cohort study. SETTING: Four botulinum toxin clinics in the Paris, France, area. PATIENTS: A French population of 100 patients with dystonia. MAIN OUTCOME: Frequency of the DYT1 mutation tested by polymerase chain reaction and enzyme restriction analysis for the 946 GAG deletion, and genotype to-phenotype correlation. RESULTS: Only 5 mutation carriers were identified, 4 of whom were part of a group of 10 patients with generalized dystonia. Onset was between ages 5 and 12 years as in typical early-onset dystonia. All 4 patients had cranial muscle involvement, which is atypical for DYT1 mutation carriers. One had segmental dystonia. Molecular analysis of relatives in 2 families demonstrated that the lack of family history was due to reduced penetrance. CONCLUSIONS: For accurate diagnosis and genetic counseling, screening for the DYT1 deletion is of great interest in cases with generalized dystonia without a family history. In other cases, positive results are rare. PMID- 10714659 TI - Using serial registered brain magnetic resonance imaging to measure disease progression in Alzheimer disease: power calculations and estimates of sample size to detect treatment effects. AB - OBJECTIVE: To evaluate the rate of brain atrophy calculated from serial magnetic resonance imaging (MRI) registration as a surrogate marker of disease progression for use in clinical trials in Alzheimer disease (AD). METHODS: Eighteen patients with mild to moderate AD and 18 age-matched normal controls underwent 2 MRI brain scans separated by a 12-month interval. Each individual's later scan was registered to their first scan, and the volume of cerebral tissue loss calculated directly from the registered and subtracted MRI scan pairs. The mean and SD of the rate of brain volume changes were used to estimate the sample sizes that would be needed in a clinical trial with a drug anticipated to modify disease progression by varying degrees. Comparable sample size estimates were performed with data for other methods of monitoring rates of brain atrophy, extracted from published papers. RESULTS: The mean (SD) rate of brain atrophy for the patients with AD was 2.37% (1.11%) per year, while in the control group it was 0.41% (0.47%) per year. Based on these figures, to have 90% power to detect a drug effect equivalent to a 20% reduction in the rate of atrophy, 207 patients would be needed in each treatment arm. This assumes a 1-year placebo-controlled trial with a 10% patient dropout rate, and that 10% of scan pairs are unusable. CONCLUSION: Registration of serial MRI volume images provides a powerful method of quantification of brain atrophy that can be used to monitor progression of AD in clinical trials. PMID- 10714660 TI - Diagnostic accuracy of dementia with Lewy bodies. AB - BACKGROUND: Diagnostic criteria for dementia with Lewy bodies (DLB) are still evolving. No data exist on prospective differentiation of DLB and Alzheimer disease (AD). OBJECTIVE: To examine the clinician's diagnostic accuracy for DLB and analyze factors contributing to false-positive DLB diagnoses. METHODS: A prospective series of 10 patients with clinically diagnosed DLB who came to autopsy was compared with 32 autopsy-confirmed cases of DLB (27 Lewy body variant, 5 diffuse Lewy body disease) and 20 autopsy-confirmed cases of AD (matched on age, sex, education, and initial Mini-Mental State Examination score) with regard to distinguishing and/or confounding clinical features. RESULTS: The clinical diagnostic accuracy for DLB was 50%, with 5 of the 10 patients clinically presumed to have DLB confirmed at autopsy. Of the 5 misdiagnosed cases, 4 had AD and 1 had progressive supranuclear palsy. The misdiagnosed DLB cases who had pure AD had fewer hallucinations (25%) than those with Lewy body variant (63%) or diffuse Lewy body disease (100%) (P = .048); however, an equal amount of spontaneous (in the absence of neuroleptics) extrapyramidal signs was found. There were no differences among groups with regard to daily fluctuations in cognition or falls. Compared with the AD control group, the misdiagnosed DLB cases with pure AD showed significantly more spontaneous extrapyramidal signs (P< or =.02). CONCLUSIONS: The clinician's diagnostic accuracy for DLB was poor. Early spontaneous extrapyramidal signs in AD were associated with false-positive clinical diagnoses of DLB. The distinction between DLB and AD may be improved by greater emphasis on hallucinations. PMID- 10714661 TI - Progressive necrotic myelopathy: clinical course in 9 patients. AB - OBJECTIVE: To review the clinical, laboratory, and radiological findings of 9 patients who had progressive idiopathic myelopathy with evidence of spinal cord necrosis. DESIGN AND METHODS: We reviewed personally examined cases of myelopathy that fulfilled the following criteria: (1) regional loss of reflexes, flaccidity, and muscle atrophy; (2) magnetic resonance imaging showing a shrunken or cavitated cord without evidence of arteriovenous malformation; (3) electromyogram showing denervation over several contiguous spinal cord sgements with preservation of sensory potentials in some cases; and (4) the absence of evidence of systemic disease or neoplasm. RESULTS: The illness began in these patients after the age of 40 years, with prominent burning or tingling limb pain, occasionally with radicular features or with less well-defined back, neck, or abdominal pain. Leg or infrequently arm weakness appeared concurrently or soon after the onset of pain. The most distinctive feature was a saltatory progression of symptoms, punctuated by both acute and subacute worsenings approximately every 3 to 9 months, culminating in paraplegia or tetraplegia. The distinguishing clinical findings, together indicative of destruction of gray matter elements of the cord, were limb atrophy, persistent areflexia, and flaccidity. The concentration of cerebrospinal fluid protein was typically elevated between 500 g/L and 1000 g/L, without oligoclonal bands, accompanied infrequently by pleocytosis. Magnetic resonance imaging showed features suggesting cord necrosis, specifically swelling, T2-weighted hyperintensity, and gadolinium enhancement over several spinal cord segments, succeeded months later by atrophy in the same regions. Necrosis of the cord was found in biopsy material from one patient and postmortem pathology in another case, but inflammation and blood vessel abnormalities were absent. Only 2 patients had prolonged visual evoked responses. The disease progressed despite immune-modulating treatments although several patients had brief epochs of limited improvement. CONCLUSIONS: The saltatory course, prolonged visual evoked responses in 2 patients, and a cranial abnormality on magnetic resonance imaging in another, raised the possibility of a link to multiple sclerosis. However, the normal cranial magnetic resonance imaging scans in 6 other patients, uniformly absent oligoclonal bands, and poor response to treatment were atypical for multiple sclerosis. On the basis of shared clinical and laboratory features, idiopathic progressive necrotic myelopathy is indistinguishable from a limited form of Devic disease. PMID- 10714662 TI - Reduction of rapid eye movement sleep by diurnal and nocturnal seizures in temporal lobe epilepsy. AB - BACKGROUND: Patients with brief, complex partial seizures frequently suffer from tiredness and decreased productivity that continue well beyond the postictal period. A possible explanation is that seizures, even when occurring during the day, disrupt sleep the following night. OBJECTIVE: To determine the effect of temporal lobe complex partial seizures on sleep structure and daytime drowsiness. METHODS: Patients with temporal lobe epilepsy were admitted for video electroencephalography monitoring. All-night polysomnography was recorded under the following 3 conditions: seizure free, seizure during the day before the recording, and seizure during the recording. Percentage of time in each sleep stage, sleep efficiency, and time to first and second rapid eye movement (REM) period were compared for seizure vs control conditions. Daytime drowsiness was also measured, using a modified maintenance of wakefulness test and 2 subjective drowsiness tests. RESULTS: Daytime seizures reduced REM from 18%+/-1% to 12%+/-2% (P = .003). Night seizures reduced REM from 16%+/-1% to 6.8%+/-2% (P<.001). Night seizures also significantly reduced stages 2 and 4 while increasing stage 1 sleep. Night seizures, but not day seizures, significantly reduced sleep efficiency, increased time to first REM period, and increased drowsiness as measured by the maintenance of wakefulness test. CONCLUSIONS: Temporal lobe complex partial seizures decrease REM sleep, particularly when occurring during sleep but also when occurring on the previous day. This may, in part, be responsible for the prolonged impairment of functioning that some patients report following seizures. PMID- 10714663 TI - The evolution of diagnosis in early Parkinson disease. Parkinson Study Group. AB - CONTEXT: Since there is no diagnostic biological marker for Parkinson disease (PD), the diagnosis is based on the results of clinical assessment. The accuracy of diagnosis improves with time and repeated assessments. Studies that require only inclusion of early cases of PD present a diagnostic challenge. Previous studies concluded that initial diagnoses of PD made by general neurologists were incorrect in 24% to 35% of the cases when patients were examined at autopsy. Experts in movement disorders are expected to have greater accuracy of initial diagnosis of PD. OBJECTIVE: To determine the evolution of clinical diagnosis in patients with early PD made initially by experts in PD. DESIGN: Eight hundred patients with mild parkinsonian symptoms (Hoehn and Yahr stage 1 or 2) who received a diagnosis of PD less than 5 years before the beginning of the study were included in the original Deprenyl and Tocopherol Antioxidative Therapy for Parkinson's Disease study. These patients were followed up prospectively with repeated clinical assessments. The following clinical criteria were used to reassess the initial diagnosis: investigator's confidence in the diagnosis of PD, presence of atypical clinical features, findings of imaging studies, response to levodopa, and results of autopsy examinations. RESULTS: The mean +/- SD duration of illness in the 800 cases at enrollment was 2.2+/-1.3 years, and the mean +/- SD Hoehn and Yahr stage was 1.6+/-0.5. The mean +/- SD follow-up was 6.0+/-1.4 years (range, 0.2-7.6 years). In 5 cases, PD was not confirmed at autopsy, and in 15 patients, the results of imaging studies indicated the presence of other pathological conditions. Of the 550 cases treated with levodopa, 49 (8.9%) had little or no improvement; 6 of these cases overlap with either autopsy or imaging study exclusion criteria. Two other cases had at least 4 of the 6 atypical clinical features arguing against the diagnosis of PD. Thus, of the 800 patients, 65 (8.1%) did not have PD according to the study criteria. Compared with those patients with the final diagnosis of PD, in the diagnoses of 60 patients without autopsy, the duration of symptoms (mean +/- SD, 7.2+/-2.0 years vs. 8.3+/-1.9 years; P<.001) and the duration of follow-up (5.3+/-1.6 years vs. 6.1+/-1.3 years; P<.001) were shorter. CONCLUSIONS: We found that 65 (8.1%) of patients initially diagnosed as having PD were later found to have an alternate diagnosis based on multifactorial clinical diagnostic criteria. This alternate diagnosis indicated that experts in PD changed their diagnoses infrequently during the 7.6 year follow-up. PMID- 10714664 TI - Diagnosis and treatment of intravascular lymphomatosis. AB - OBJECTIVE: To describe a patient with unusually good outcome of a rare, high grade lymphoma that often involves the nervous system. DESIGN: Case report. SETTING: University hospital. CASE: A 70-year-old pharmacist first presented with meningoencephalitislike symptoms and 6 months later with acute confusional state followed by complex partial status epilepticus. Diagnosis of intravascular lymphomatosis was made using detection and biopsy of a bilateral adrenal tumor. MAIN OUTCOME AND RESULTS: Polychemotherapy consisting of CHOP (cyclophosphamide, doxorubicin hydrochloride, vincristine sulfate, and prednisone) led to complete remission. The patient's survival time currently exceeds 21/2 years. CONCLUSIONS: The possibility of intravascular lymphomatosis should be considered in adult patients with unclear meningoencephalitic syndrome, acute confusional state, dementia, or other unexplained neurologic conditions with signs of a systemic disease. In intravascular lymphomatosis, as in other high-grade non-Hodgkin lymphomas, CHOP polychemotherapy should be the standard treatment. PMID- 10714665 TI - Nitrous oxide anesthesia-associated myelopathy. AB - BACKGROUND: The role of nitrous oxide exposure in neurologic complications of subclinical cobalamin deficiency has been reported, but few cases are well documented. OBSERVATION: Two weeks after surgery for prosthetic adenoma, a 69 year-old man developed ascending paresthesia of the limbs, severe ataxia of gait, tactile sensory loss on the 4 limbs and trunk, and absent tendon reflexes. After a second surgical intervention, the patient became confused. Four months after onset, the patient had paraplegia, severe weakness of the upper limbs, cutaneous anesthesia sparing the head, and confusion. Moderate macrocytosis, low serum B12 levels, and a positive Schilling test result led to the diagnosis of pernicious anemia. Results of electrophysiologic examinations showed a diffuse demyelinating neuropathy. Magnetic resonance imaging of the spinal cord disclosed hyperintensities of the dorsal columns on T2-weighted images. CONCLUSIONS: Pernicious anemia can result in severe neurologic symptoms with only mild hematologic changes. The role of nitrous oxide anesthesia in revealing subclinical B12 deficiency must be emphazised. Magnetic resonance imaging of the spinal cord might be helpful in making the diagnosis. PMID- 10714666 TI - Effect of liver transplantation on neurological manifestations in Wilson disease. AB - BACKGROUND: Liver transplantation (LT) is the sole resolutive therapy for Wilson disease (WD) and is the treatment of choice for patients with WD who have fulminant hepatic failure or end-stage cirrhosis. Although its role in managing the neurological manifestations of WD is not yet conclusive, LT has recently been advocated as a therapy for neurologically affected patients with WD with stable liver function. OBJECTIVE: To evaluate the effect of LT on the neurological manifestations of WD. OBSERVATION: A 44-year-old man with WD with cirrhosis and neurological symptoms (motor dysfunction and cognitive impairment) experienced a dramatic improvement in motor function early after LT, as well as normalization of copper balance and the disappearance of Kayser-Fleischer rings. Abnormalities seen on magnetic resonance imaging scans were reversed 18 months after LT. Cognitive testing 2 years after LT showed a moderate global improvement. CONCLUSIONS: In this case, LT healed the neurological manifestations of WD. To date, this favorable result has been seen in almost 80% of cases. However, the decision to perform LT in patients with WD solely on the basis of neurological impairment must be considered experimental. PMID- 10714667 TI - Myoclonus from selective dentate nucleus degeneration in type 3 Gaucher disease. AB - OBJECTIVE: To describe a case with a new genetic variant of type 3 Gaucher disease presenting with stimulus-sensitive and action myoclonus in the presence of selective dentate abnormalities. DESIGN: Clinical, pathologic, and molecular genetic studies. SETTING: Medical school departments. PATIENT: A 6-year-old girl with type 3 Gaucher disease experienced progressively crippling generalized stimulus-sensitive and action myoclonus. Repeated electroencephalographic examination did not show cortical activity associated with the myoclonus, suggesting its subcortical origin. Neuropathological examination revealed selective degeneration of the cerebellar dentate nucleus and dentatorubrothalamic pathway in the face of essentially complete lack of storage in the brain. Mutation analysis identified the following 2 mutant alleles: one with a V394L mutation and the other with the lesion RecTL (D409H + L444P + A456P + V460V), which resulted from a recombination event, with the pseudogene located 16 kilobases downstream from the structural gene. CONCLUSION: Given the restricted abnormalities, this genetically unique case provides insight into the pathogenesis of myoclonus and suggests a prominent role for the cerebellar dentate nucleus in its genesis. PMID- 10714668 TI - Cognitive and behavioral abnormalities in a case of central nervous system Whipple disease. AB - BACKGROUND: Whipple disease is a rare condition characterized by migratory polyarthralgias, fever, and chronic diarrhea. A subset of patients with the disease may either initially have or eventually develop symptoms of central nervous system involvement. DESIGN AND METHODS: The cognitive and behavioral functioning of a patient with central nervous system involvement from Whipple disease was studied during a 7-month period. Serial neuropsychological evaluations were used to quantify the nature of his cognitive and behavioral profile. SETTING: Neurology department of a university medical center. RESULTS: A variety of cognitive impairments were noted, most prominently in the domains of sustained attention, memory, executive function, and constructional praxis. There were striking behavioral manifestations as well, including disinhibition and confabulation. CONCLUSIONS: The case demonstrates a degree of higher-order central nervous system dysfunction rarely observed and quantified in connection with Whipple disease, and with important implications for differential diagnosis of certain neurologic conditions. We also call attention to some of the neuroanatomical correlates of this encephalopathic condition. PMID- 10714669 TI - Wernicke encephalopathy. PMID- 10714670 TI - Levodopa toxicity in Parkinson disease: reality or myth? Reality--practice patterns should change. PMID- 10714671 TI - Is levodopa toxic? PMID- 10714672 TI - Neurological manifestations in Sjogren syndrome. PMID- 10714673 TI - Barriers to care for patients with neurologic disease in rural Zambia. AB - The awesome burden of treatable yet untreated neurologic disease in the developing world presents a humanitarian crisis to those of us with neurologic expertise from more privileged situations. Although increased economic resources are critically needed, a shortage of personnel to care for these patients is as great a problem. It is neither feasible nor desirable to propose training neurologists to work in these regions. However, COs could be selected to receive additional training and return to their home regions to serve as resources for referrals and as community educators. Such a training program would not require massive financial commitments. A handful of dedicated neurologists could conceivably accomplish this in 6- to 8-week training sessions. Ideally, educational materials, such as posters and pamphlets in both English and the native language of the various regions, would be provided at no cost. Existing textbooks in neurology are written for physicians and often focus on diagnostic evaluations and therapies far beyond the services available in developing countries. A text for practical use by COs and community health workers that discusses the application of available medicines and therapies for common neurologic problems would be invaluable. Similar books exist that address general medical and obstetrical problems (for example, Where There Is No Doctor: A Village Health Care Handbook). Where There Is No Neurologist could be developed as a primary teaching tool and a valuable reference for COs with neurologic expertise. Neuroscience researchers, clinical neurologists, and neurology residents from industrialized countries have much to offer and to gain by working in the Third World. Research to monitor the incidence and resource utilization of emerging problems such as stroke is needed to influence public policy. The economic burden and lost productivity caused by neurologic disease in this part of the world has not been appreciated or explored. Disease beyond the scope of Western experience manifests daily in places like Chikankata. Entities such as tabes neurosyphilis, which previous generations of neurologists used as the basis for their training, still abound in Zambia. Much personal satisfaction can be gained in providing care to this vulnerable and underserved population. PMID- 10714674 TI - The global burden of disease study: implications for neurology. AB - Because of the epidemiological transition, the global burden of illness has changed. Several factors have contributed to this change, including improvements in maternal and child health, increasing age of populations, and newly recognized disorders of the nervous system. It is now evident that neurologic disorders have emerged as priority health problems worldwide. This is reflected in the Global Burden of Disease Study, jointly published by the World Health Organization and other groups. The proportionate share of the total global burden of disease resulting from neuropsychiatric disorders is projected to rise to 14.7% by 2020. Although neurologic and psychiatric disorders comprise only 1.4% of all deaths, they account for a remarkable 28% of all years of life lived with a disability. This study provides compelling evidence that one cannot assess the neurologic health status of a population by examining mortality statistics alone. Health ministries worldwide must prioritize neurologic disorders, and neurologists must be prepared to provide care for increased numbers of people individually and in population groups. PMID- 10714675 TI - How does one define progression of disease in patients with relapsing-remitting multiple sclerosis? PMID- 10714676 TI - Interferons in the treatment of multiple sclerosis: errors and misrepresentations. PMID- 10714677 TI - Association of varicella-zoster virus with cervical artery dissection in 2 cases. PMID- 10714678 TI - Chemokines: a new classification system and their role in immunity. PMID- 10714679 TI - Posttranslational regulation of IRF-4 activity by the immunophilin FKBP52. AB - Interferon regulatory factor-4 (IRF-4) plays an important role in immunoregulatory gene expression in B and T lymphocytes and is also highly expressed in human T cell leukemia virus type 1 infected cells. In this study, we characterize a novel interaction between IRF-4 and the FK506-binding protein 52 (FKBP52), a 59 kDa member of the immunophilin family with peptidyl-prolyl isomerase activity (PPIase). IRF-4-FKBP52 association inhibited IRF4-PU.1 binding to the immunoglobulin light chain enhancer E(lambda2-4) as well as IRF-4-PU.1 transactivation, effects that were dependent on functional PPIase activity. FKBP52 association also resulted in a structural modification of IRF-4, detectable by immunoblot analysis and by IRF-4 partial proteolysis. These results demonstrate a novel posttranslational mechanism of transcriptional control, mediated through the interaction of an immunophilin with a transcriptional regulator. PMID- 10714680 TI - WECHE: a novel hematopoietic regulatory factor. AB - Previously, we described AGM-derived endothelial cell lines that either inhibited or permitted the development of erythroid or B cells. We utilized a differential gene expression method to isolate a chemokine, termed WECHE, from one of these cell lines. WECHE inhibited the formation of erythroid cells but had no effect on either myeloid or B cell formation. WECHE repressed BFU-E development from either mouse fetal liver or bone marrow progenitor cells but had no effect on colony formation induced by IL-3 or IL-7. WECHE reduced HPP-CFC production from fetal liver-derived stem cells. WECHE hindered the growth of yolk sac-derived endothelial cells. WECHE was also chemotactic for bone marrow cells. Thus, WECHE is a novel chemokine that regulates hematopoietic differentiation. PMID- 10714681 TI - A novel functional interaction between Vav and PKCtheta is required for TCR induced T cell activation. AB - Vav and PKCtheta play an early and important role in the TCR/CD28-induced stimulation of MAP kinases and activation of the IL-2 gene. Vav is also essential for actin cytoskeleton reorganization and TCR capping. Here, we report that PKCtheta function was selectively required in a Vav signaling pathway that mediates the TCR/CD28-induced activation of JNK and the IL-2 gene and the upregulation of CD69 expression. Vav also promoted PKCtheta translocation from the cytosol to the membrane and cytoskeleton and induced its enzymatic activation in a CD3/CD28-initiated pathway that was dependent on Rac and on actin cytoskeleton reorganization. These findings reveal that the Vav/Rac pathway promotes the recruitment of PKCtheta to the T cell synapse and its activation, essential processes for T cell activation and IL-2 production. PMID- 10714682 TI - Triggering the TCR complex causes the downregulation of nonengaged receptors by a signal transduction-dependent mechanism. AB - Downregulation of the TCR complex is believed to be intimately tied to T cell activation, allowing serial triggering of receptors and desensitization of stimulated cells. We studied transfected and transgenic T cells expressing CD3zeta chimeras to demonstrate that ligand engagement of the TCR or chimeras causes comodulation of nonengaged receptors. Comodulation required protein tyrosine kinase activity but not trans-phosphorylation of nonengaged receptors. The TCR appears to be downregulated by at least two mechanisms. One mechanism requires direct engagement, independent of signaling. The second requires signaling and downregulates nontriggered receptors. These results shed new light on the process of TCR downregulation and indicate that the number of downregulated TCRs cannot be assumed to equal the number of engaged receptors. PMID- 10714683 TI - Abrogation of TGFbeta signaling in T cells leads to spontaneous T cell differentiation and autoimmune disease. AB - Targeted mutation of TGFbeta1 in mice demonstrated that TGFbeta1 is one of the key negative regulators of immune homeostasis, as its absence leads to activation of a self-targeted immune response. Nevertheless, because of the highly pleiotropic properties of TGFbeta and the presence of TGFbeta receptors on most cell types, its biologic role in the regulation of immune homeostasis is not yet understood. To limit the consequences of TGFbeta effects to a single cell type, we developed a transgenic approach to abrogate the TGFbeta response in key immune cells. Specifically, we expressed a dominant-negative TGFbeta receptor type II under a T cell-specific promoter and created a mouse model where signaling by TGFbeta is blocked specifically in T cells. Using this transgenic model, we show that T cell homeostasis requires TGFbeta signaling in T cells. PMID- 10714684 TI - Thymocyte resistance to glucocorticoids leads to antigen-specific unresponsiveness due to "holes" in the T cell repertoire. AB - We have proposed that glucocorticoids antagonize TCR-mediated activation and influence which TCR avidities result in positive or negative selection. We now analyze the immune response of mice whose thymocytes express antisense transcripts to the glucocorticoid receptor (TKO mice). TKO mice responded normally to the complex antigen PPD but were proliferative nonresponders to pigeon cytochrome c 81-104 (PCC), having a large decrease in the frequency of PCC responsive CD4+ T cells. Unlike wild-type T cells, few TKO T cells in PCC specific cell lines expressed V alpha11+Vbeta3+. Furthermore, for naive CD4+ T cells from unimmunized TKO mice, the frequencies of many of the molecular features common to the CDR3 regions of PCC-responsive V alpha11+Vbeta3+ cells were substantially decreased. Thus, thymocyte glucocorticoid hyporesponsiveness resulted in loss of cells capable of responding to PCC, corresponding to an antigen-specific "hole" in the T cell repertoire. PMID- 10714685 TI - Expression of CD27 on murine hematopoietic stem and progenitor cells. AB - Hematopoietic stem cells (HSC) are defined by self-renewal and multilineage differentiation potentials. In order to uncover the genetic program of HSC, we utilized high-density arrays to compare gene expression in highly purified mouse HSC and their mature progeny. One molecule specifically expressed in immature cells is CD27, a member of the TNF receptor family previously shown to play roles in lymphoid proliferation, differentiation, and apoptosis. We show here that the CD27 protein is expressed by about 90% of cells in a purified HSC population. Interestingly, the CD27pos cells are enriched for cells with short-term hematopoietic activities (colony forming potential in vivo and in vitro), while the minority CD27neg population is more effective in clonal long-term transplantation. PMID- 10714686 TI - Impaired immunity and enhanced resistance to endotoxin in the absence of neutrophil elastase and cathepsin G. AB - While the critical role of reactive oxygen intermediates (ROI) in the microbicidal activity of polymorphonuclear granulocytes is well established, the function of the nonoxidative effector mechanisms in vivo remains unclear. Here we show that mice deficient in the neutrophil granule serine proteases elastase and/or cathepsin G are susceptible to fungal infections, despite normal neutrophil development and recruitment. The protease deficiencies but not the absence of ROI leads to enhanced resistance to the lethal effects of endotoxin LPS, although normal levels of TNFalpha are produced. The data demonstrate a critical role of the nonoxidative effector mechanisms of neutrophils in host immunity and immunopathology and identify elastase and cathepsin G as effectors in the endotoxic shock cascade downstream of TNFalpha. PMID- 10714687 TI - Murine CD1d-restricted T cell recognition of cellular lipids. AB - NKT cells are associated with immunological control of autoimmune disease and cancer and can recognize cell surface mCD1d without addition of exogenous antigens. Cellular antigens presented by mCD1d have not been identified, although NKT cells can recognize a synthetic glycolipid, alpha-GalCer. Here we show that after addition of a lipid extract from a tumor cell line, plate-bound mCD1d molecules stimulated an NKT cell hybridoma. This hybridoma also responded strongly to three purified phospholipids, but failed to recognize alpha-GalCer. Seven of sixteen other mCD1d restricted hybridomas also showed a response to certain purified phospholipids. These findings suggest NKT cells can recognize cellular antigens distinct from alpha-GalCer and identify phospholipids as potential self-antigens presented by mCD1d. PMID- 10714688 TI - Ly6d-L, a cell surface ligand for mouse Ly6d. AB - The mouse Ly6 gene family encodes proteins found in lymphocytes and other cells. Some are involved in cell activation; no ligands have been found. A ligand for Ly6d (ThB) was identified on lymphocytes using microspheres loaded with Ly6d and the cDNA isolated from a spleen/thymus library by panning on Ly6d. The Ly6d ligand (Ly6d-L) is a nonglycosylated protein of 9 kDa of broad distribution, rich in cysteine, with no discernable transmembrane sequence. Its N and C termini are on the cell surface, where it associates with a 30 kDa protein. Ly6d-L is homologous with an EGF repeat of Notch. PMID- 10714689 TI - Childhood myasthenia. PMID- 10714690 TI - Combination cyclopentolate and phenylephrine for mydriasis in premature infants with heavily pigmented irides. AB - PURPOSE: This study examined whether safe and effective mydriasis can be achieved in premature infants with heavily pigmented irides using combination cyclopentolate 0.2% and phenylephrine 1% eyedrops. METHODS: A prospective, randomized double-blind study was performed to compare combination cyclopentolate 0.2% and phenylephrine 1% eye-drops with triple instillation of tropicamide 0.5% and phenylephrine 2.5%. Twenty-eight consecutive babies with dark irides and birthweight <1600 g referred for screening for retinopathy of prematurity comprised the study population. Infants' eyes were randomly dilated twice with both regimens within a 2-week period. Blood pressure, heart rate, and pupil size were measured. RESULTS: Good mydriasis was achieved in both groups with no significant differences in pupil size or blood pressure (systolic, diastolic, or mean arterial pressures) over starting baseline values. Pulse rates decelerated below the baseline values in both groups, but these differences were not large. CONCLUSION: The single combination eyedrop of cyclopentolate 0.2% and phenylephrine 1% is as effective and safe a mydriatic for infants with dark irides as both tropicamide 0.5% and phenylephrine 2.5%. PMID- 10714691 TI - Diagnosis of amblyopia using the 10-diopter fixation test: a proposed modification for patients with unilateral ptosis. AB - PURPOSE: The 10-diopter (D) fixation test is a useful test for detecting amblyopia in children without a manifest deviation and in whom a reliable visual acuity assessment is difficult. The 10 prism diopter is conventionally placed base down. METHODS: Seventeen children were studied over a 12-month period to determine the effect of prism orientation on the accuracy of the 10-D fixation test in children with unilateral ptosis. RESULTS: Anomalous results were obtained in 4 of 17 patients with the prism held base up and base down. CONCLUSIONS: To prevent an erroneous diagnosis of amblyopia in children with unilateral ptosis, the prism should be held base up when performing the 10-D fixation test. PMID- 10714692 TI - Binocular fixation pattern and visual acuity in children with strabismic amblyopia. AB - BACKGROUND: A prospective study was undertaken to compare the binocular fixation pattern and presence of amblyopia in strabismic children. METHODS: Fifty-three children with manifest strabismus and the ability to cooperate with an optotype acuity test were examined. The binocular fixation pattern and logMAR visual acuity were recorded by separate, masked observers under standardized conditions. The binocular fixation pattern was divided into four grades from alternation to uniocular fixation. RESULTS: Patients who freely alternated did not have amblyopia, while those who maintained or preferred fixation with a given eye tended to have amblyopia in the nonpreferred eye. CONCLUSION: The binocular fixation pattern can be rapidly assessed with minimal equipment and training. These findings confirm the usefulness of a graded assessment of the binocular fixation pattern in the detection of amblyopia. PMID- 10714694 TI - Conservative management of orbital lymphangioma. PMID- 10714693 TI - Treatment of glaucoma in children with Sturge-Weber syndrome. AB - BACKGROUND: Sturge-Weber syndrome is a rare congenital neuro-oculocutaneous disorder. Ocular involvement can include glaucoma and vascular malformations of the conjunctiva, episclera, choroid, and retina. METHODS: The records of 19 Sturge-Weber syndrome patients (mean age 8.2 years) treated at our institution were reviewed to determine the incidence of ophthalmologic manifestations in Sturge-Weber syndrome. RESULTS: Glaucoma occurred in 42% of all patients and was more frequent in patients with a port-wine stain involving both upper and lower eyelids. Other ocular manifestations included conjunctival/episcleral hemangioma, choroidal hemangioma, iris heterochromia, retinal detachment, strabismus, and homonymous hemianopia. In 7 of 8 patients with glaucoma, topical pharmacotherapy (beta-blockers and carbonic anhydrase inhibitors) alone failed to normalize intraocular pressures. In those patients, cryocoagulation of the ciliary body was performed. Mean postoperative intraocular pressure after a mean follow-up of 4-5 years was <22 mm Hg in 6 patients. CONCLUSIONS: Cryocoagulation of the ciliary body combined with topical medication is an effective and safe treatment option in the management of glaucoma in children with Sturge-Weber syndrome. PMID- 10714695 TI - Bilateral congenital glaucoma in a child with hydrops fetalis, congenital pulmonary lymphangiectasia, and lymphoedema. PMID- 10714696 TI - Infantile lipemia retinalis and conjunctivalis. PMID- 10714697 TI - Anterior transposition of the inferior oblique for the treatment of a lost inferior rectus muscle. PMID- 10714698 TI - Hemophilus aegyptius endophthalmitis following strabismus surgery. PMID- 10714699 TI - Hydrogen atom abstraction by Cu(II)- and Zn(II)-phenoxyl radical complexes, models for the active form of galactose oxidase. AB - The Cu(II) and Zn(II) complexes of phenoxyl radical species [M(II)(L1*)(NO3)]+ (M=Cu or Zn, L1H: 2-methylthio-4-tert-butyl-6-[[bis[2-(2 pyridyl)ethyl]amino]methyl]phenol ) and [M(II)(L2*)(NO3)]+ (M=Cu or Zn, L2H: 2,4 di-tert-butyl-6-[[bis[2-(2-pyridyl)ethyl]amino]methyl]phenol) are prepared as model complexes of the active form of galactose oxidase (GAO). Hydrogen atom abstraction of 1,4-cyclohexadiene and tert-butyl substituted phenols by the GAO model complexes proceeds very efficiently to give benzene and the corresponding phenoxyl radical or its C-C coupling dimer as the oxidation products, respectively. Kinetic analyses on the oxidation reactions have shown that the hydrogen atom abstraction of the phenol substrates is significantly enhanced by the coordinative interaction of the OH group to the metal ion center of the complex, providing valuable insight into the enzymatic mechanism of the alcohol oxidation. Details of the substrate-activation process have been discussed based on the activation parameters (deltaH* and deltaS*) of the reactions. PMID- 10714700 TI - Desaturation of trans-octadecenoyl-acyl carrier protein by stearoyl-acyl carrier protein delta9 desaturase. AB - Positional isomers of mono-unsaturated 18:1-ACP have been used as substrates for stearoyl-acyl carrier protein delta9 desaturase to test whether a C-H bond abstraction from either the C-9 or C-10 position could lead to rearranged products diagnostic for the production of an allylic radical intermediate. The reconstituted enzyme complex was able to desaturate trans-delta11-18:1-ACP and trans-delta7-18:1-ACP, but not trans-delta9-18:1-ACP, or any of the corresponding cis-isomers. Enzymatic desaturation of trans-delta11-18:1-ACP gave a single product, cis-delta9,trans-delta11-18:2-ACP, as characterized by gas chromatography-electron ionization mass spectrometry of the molecular ions, the fragmentation products of pyrrolidide and 4,4-dimethyloxazoline derivatives, and by comparison of chromatographic retention times with authentic standards. Reaction of trans-delta7-18:1-ACP gave two enzymic products, trans-delta7,cis delta9-18:2 (approximately 80%) and trans-delta7,cis-delta11-18:2 (approximately 20%). The major product was likely formed in a reaction identical to that of 18:0 ACP desaturation, while the minor product was likely formed by alternative placement of the C-10 and C-11 positions of the substrate analog in a cis configuration relative to the diiron oxidant. Since none of the products observed are indicative of rearrangements originating with an allylic radical, a discussion of the origins and possible implications of these results is presented. PMID- 10714701 TI - Towards an artificial model for Photosystem II: a manganese(II,II) dimer covalently linked to ruthenium(II) tris-bipyridine via a tyrosine derivative. AB - In order to model the individual electron transfer steps from the manganese cluster to the photooxidized sensitizer P680+ in Photosystem II (PS II) in green plants, the supramolecular complex 4 has been synthesized. In this complex, a ruthenium(II) tris-bipyridine type photosensitizer has been linked to a manganese(II) dimer via a substituted L-tyrosine, which bridges the manganese ions. The trinuclear complex 4 was characterized by electron paramagnetic resonance (EPR) and electrospray ionization mass spectrometry (ESI-MS). The excited state lifetime of the ruthenium tris-bipyridine moiety in 4 was found to be about 110 ns in acetonitrile. Using flash photolysis in the presence of an electron acceptor (methylviologen), it was demonstrated that in the supramolecular complex 4 an electron was transferred from the excited state of the ruthenium tris-bipyridine moiety to methylviologen, forming a methylviologen radical and a ruthenium(III) tris-bipyridine moiety. Next, the Ru(III) species retrieved the electron from the manganese(II/II) dimer in an intramolecular electron transfer reaction with a rate constant kET > 1.0 x 10(7) s(-1), generating a manganese(II/III) oxidation state and regenerating the ruthenium(II) photosensitizer. This is the first example of intramolecular electron transfer in a supramolecular complex, in which a manganese dimer is covalently linked to a photosensitizer via a tyrosine unit, in a process which mimics the electron transfer on the donor side of PS II. PMID- 10714702 TI - Two-step concerted mechanism for methane hydroxylation on the diiron active site of soluble methane monooxygenase. AB - A new concerted mechanism is proposed for the conversion of methane to methanol on intermediate Q of soluble methane monooxygenase (sMMO), the active site of which is considered to involve an Fe2(mu-O)2 diamond core. A hybrid density functional theory (DFT) method is used for our mechanistic study on the important reactivity of the bare FeO+ complex and a diiron model of intermediate Q. The reaction pathway for the methane hydroxylation on the diiron complex is essentially identical to that for the gas-phase reaction by the bare FeO+ complex. Methane is highly activated on the dinuclear iron model through the formation of a methane complex, in which a coordinatively unsaturated iron plays a central role in the bonding interaction between the diiron model and substrate methane. A H atom abstraction via a four-centered transition state and a recombination of the OH and CH3 groups via a three-centered transition state successively occur on the dinuclear iron-oxo species, leading to the formation of a methanol complex that corresponds to intermediate T. These electronic processes take place in a concerted manner. Our mechanism for methane hydroxylation by sMMO is different from the radical mechanism that has been widely accepted for enzymatic hydrocarbon hydroxylation, especially by cytochrome P450. PMID- 10714703 TI - Stability and folding of the ferredoxin from the hyperthermophilic archaeon Acidianus ambivalens. AB - The ferredoxin from the thermophilic archaeon Acidianus ambivalens is a small monomeric protein containing two iron-sulfur centres, one [3Fe-4S](1+/0) and one [4Fe-4S](2+/1+). It is an intrinsically hyperstable protein, being expressed at the organism's extreme optimal growth temperature: 80 degrees C. Using spectroscopic methods we have investigated the unfolding reaction of the Acidianus ambivalens ferredoxin. No unfolding of the oxidised ferredoxin was observed at pH 7.0, even in the presence of 8 M GuHCl. Upon increasing the pH to 10.0, the unfolding transition showed a midpoint at 6.3 M GuHCl and an unfolding free energy of 70 kJ mol(-1) in buffer (pH 10) was estimated. Kinetic-unfolding experiments showed that the polypeptide unfolding correlated with rearrangement of the iron-sulfur centres to new ones which had strong absorption maxima at 520 and 610 nm. These new, possibly linear three-iron, clusters were coordinated to the unfolded protein but degraded slowly. From thermal experiments in the presence of GuHCl we estimated the melting temperature for the Acidianus ambivalens ferredoxin in buffer (at pH 7) to be 122 degrees C. Possible structural properties that contribute to the large thermal stability of the Acidianus ambivalens ferredoxin are discussed using a three-dimensional protein model. PMID- 10714704 TI - Inhibition of the aminopeptidase from Aeromonas proteolytica by L-leucinethiol: kinetic and spectroscopic characterization of a slow, tight-binding inhibitor enzyme complex. AB - The peptide inhibitor L-leucinethiol (LeuSH) was found to be a potent, slow binding inhibitor of the aminopeptidase from Aeromonas proteolytica (AAP). The overall potency (K(I)*) of LeuSH was 7 nM while the corresponding alcohol L leucinol (LeuOH) was a simple competitive inhibitor of much lower potency (K(I) = 17 microM). These data suggest that the free thiol is likely involved in the formation of the E x I and E x I* complexes, presumably providing a metal ligand. In order to probe the nature of the interaction of LeuSH and LeuOH with the dinuclear active site of AAP, we have recorded both the electronic absorption and EPR spectra of [CoCo(AAP)], [CoZn(AAP)], and [ZnCo(AAP)] in the presence of both inhibitors. In the presence of LeuSH, all three Co(II)-substituted AAP enzymes exhibited an absorption band centered at 295 nm, characteristic of a S --> Co(II) ligand-metal charge-transfer band. In addition, absorption spectra recorded in the 450 to 700 nm region all showed changes characteristic of LeuSH and LeuOH interacting with both metal ions. EPR spectra recorded at high temperature (19 K) and low power (2.5 mW) indicated that, in a given enzyme molecule, LeuSH interacts weakly with one of the metal ions in the dinuclear site and that the crystallographically identified mu-OH(H) bridge, which has been shown to mediate electronic interaction of the Co(II) ions, is likely broken upon binding LeuSH. EPR spectra of [CoCo(AAP)]-LeuSH, [ZnCo(AAP)]-LeuSH, and [Co_(AAP)]-LeuSH were also recorded at lower temperature (3.5-4.0 K) and high microwave power (50-553 mW). These signals were unusual and appeared to contain, in addition to the incompletely saturated contributions from the signals characterized at 19 K, a very sharp feature at g(eff) approximately 6.5 that is characteristic of thiolate Co(II) interactions. Combination of the electronic absorption and EPR data indicates that LeuSH perturbs the electronic structure of both metal ions in the dinuclear active site of AAP. Since the spin-spin interaction seen in resting [CoCo(AAP)] is abolished upon the addition of LeuSH, it is unlikely that a mu S(R) bridge is established. PMID- 10714705 TI - The preparation, characterisation, and DNA adduct profile of 2-amino-2-methyl-3 butanoneoximedichloroplatinum(II), a platinum(II) complex designed to bind to GpA sequences of DNA. AB - The complex, 2-amino-2-methyl-3-butanoneoximedichloroplatinum(II), [Pt(ambo)Cl2], was chosen because of its potential to bind to GpA sequences of duplex DNA. Crystals of [Pt(ambo)Cl2] are monoclinic, space group, P2(1)/n, a = 6.799(4), b = 17.642(5), c = 8.193(2) A, beta = 102.10(3) degrees, Z = 4, R = 0.033 (1864 F). The binding of [Pt(ambo)Cl2] to salmon-sperm DNA was studied using enzymatic digestion and HPLC analysis. [Pt(ambo)Cl2] was found to form fewer GpG and ApG intrastrand adducts and more monofunctional adducts than [Pt(en)Cl2]. Binding to GpA sequences could not be established, but [Pt(ambo)Cl2] forms substantially more adducts with adenine than does [Pt(en)Cl2]. PMID- 10714706 TI - Control of CooA activity by the mutation at the C-terminal end of the heme binding domain. AB - A constitutively active mutant of a CooA, in which Met131 was replaced by Leu, was isolated by random mutagenesis. Site-directed mutagenesis at position 131 revealed that M131R-CooA was also constitutively active even in the absence of CO and that M131P-, M131D-, and M131E-CooA were constitutively inactive regardless of the presence or absence of CO. While M131L- and M131E-CooA showed almost the same electronic absorption spectra as those of the wild type in the ferric, ferrous, and CO-bound forms, M131D-CooA showed the typical spectrum of a five coordinate heme protein in the ferric form. The conformational change around the heme induced by CO binding, which triggers the activation of CooA, is thought to be linked to the rearrangement of the conformation around the hinge region between the heme-binding and DNA-binding domains and/or of the relative orientation of the two domains to activate CooA. PMID- 10714707 TI - Effects of cetyltrimethylammonium bromide on reactions catalyzed by maxizymes, a novel class of metalloenzymes. AB - We demonstrated previously that some shortened forms of hammerhead ribozymes had high cleavage activity that was similar to that of the wild-type parental hammerhead ribozyme. Moreover, the active species appeared to form dimeric structures with a common stem II (in order to distinguish monomeric forms of conventional minizymes that have low activity from our novel dimers with high level activity, the latter very active short ribozymes were designated 'maxizymes'). The dimers can be homodimeric (with two identical binding sequences) or heterodimeric (with two different binding sequences). In the case of heterodimers, they are in equilibrium with inactive homodimers. In this study, we investigated the effects of cationic detergent, cetyltrimethylammonium bromide (CTAB), on reactions catalyzed by a variety of maxizymes. The slope of close to unity in profiles of pH versus rate demonstrated that the deprotonation was important in catalysis and that the rate-limiting chemical step was followed in these reactions. Addition of appropriate amounts of CTAB enhanced the activity of a variety of maxizymes. The activity of our least stable, least active maxizyme was enhanced 100-fold by CTAB. Thus, CTAB effectively enhanced the conversion of kinetically trapped inactive conformations to active forms. Moreover, we suggest that the activity and specificity of catalytic RNAs in vivo might be better estimated if their reactions are monitored in vitro in the presence of appropriate amounts of CTAB. PMID- 10714708 TI - Valence tautomerism and macrocycle ruffling in nickel(III) porphyrins. AB - Nonlocal density functional calculations with full geometry optimization have been carried out on the low-lying electronic states of oxidized nickel porphyrins. For [NiIII(P)(Py)2]+, the ground state corresponds to a t2g6(z2)1 configuration and the t2g6(x2-y2)1 configuration is 0.43 eV higher in energy. In contrast, the ground state of [NiIII(P)(CN)2]- corresponds to a t2g6(x2-y2)1 configuration, the t2g6(z2)1 configuration being 0.96 eV higher in energy. The results are consistent with EPR spectroscopic results on the TPP analogs of these complexes. For [Ni(P)(Py)2]+, the a2u- and a1u-type Ni(II) porphyrin cation radical states are higher in energy by 0.63 and 1.23 eV, respectively, relative to the t2g6(z2)1 Ni(III) ground state. The Ni-N(Porphyrin) distance is significantly shorter in [NiIII(P)(Py)2]+ (196 pm) than in [NiIII(P)(CN)2]- (206 pm), which is consistent with the ruffled and planar macrocycle conformations, respectively, in the two complexes. PMID- 10714709 TI - The reduction of the Rieske iron-sulfur cluster in naphthalene dioxygenase by X rays. AB - Naphthalene 1,2 dioxygenase (NDO) displays characteristic UV-Vis spectra depending on the oxidation state of the Rieske center. Investigations on crystals of NDO grown for X-ray diffraction experiments showed spectra characteristic of the oxidized form. Crystals reduced in an anaerobic glovebox using sodium dithionite showed a characteristic reduced spectrum. Spectra of crystals (cooled to 100 K) after being exposed to X-rays for data collection showed spectra corresponding to a reduced Rieske iron center, demonstrating the ability of X rays to change the oxidation state of the Rieske iron-sulfur cluster in NDO. PMID- 10714710 TI - Ligand replacement study at the His120 site of purple CuA azurin. AB - The CuA center is a dinuclear Cu2S2(Cys) electron transfer center found in cytochrome c oxidase and nitrous oxide reductase. In a previous investigation of the equatorial histidine ligands' effect on the reduction potential, electron transfer and spectroscopic properties of the CuA center, His120 in the engineered CuA azurin was mutated to Asn, Asp, and Ala. The identical absorption and EPR spectra of these mutants indicate that a common ligand is bound to the copper center. To identify this replacement ligand, the His120Gly CuA azurin mutant was constructed and purified. Absorption and X-band EPR spectra show that His120Gly is similar to the other His120X (X = Asn, Asp, Ala) mutant proteins. Titrations with chloride, imidazole, and azide suggest that the replacement ligand is not exchangeable with exogenous ligands. The possibility of an internal amino acid acting as the replacement ligand for His120 in the His120X mutant proteins was investigated by analyzing the CuA azurin crystal structure and then converting the likely internal ligand, Asn 119, to Asp, Ser, or Ala in the His120Gly mutant. The double mutants H120G/Asn 119X (X = Asp, Ser, or Ala) displayed UV-Vis absorption and EPR spectra that are identical to His120Gly and the other His120X mutants, indicating that Asn119 is not the internal ligand replacing His120 in the His120X mutant proteins. These results demonstrate the remarkable stability of the dinuclear His120 mutants of CuA azurin. PMID- 10714711 TI - Salvage surgery for patients with recurrent squamous cell carcinoma of the upper aerodigestive tract: when do the ends justify the means? AB - OBJECTIVES/HYPOTHESES: Salvage surgery is widely viewed as a "double-edged sword." It is the best option for many patients with recurrent cancer of the upper aerodigestive tract, especially when original therapy included irradiation, yet it may provide only modest benefit at high personal cost to the patient. The stakes are high because alternatives are of limited value. The primary objective of this study was to fully assess the value of salvage surgical procedures in the treatment of local and regional recurrence. The following hypotheses were developed to focus the study design and data analysis. 1) The efficacy of salvage surgery correlates recurrent stage, recurrent site, and time to presalvage recurrence. 2) The economic and noneconomic costs of salvage surgery increase with higher recurrent stage. 3) Information relating the value of salvage surgery to recurrent stage and recurrent site will be useful to these patients and the physicians who treat them. STUDY DESIGN: Two complimentary methods of investigation were used: a meta-analysis of the published literature and a prospective observational study of patients undergoing salvage surgery for recurrent cancer of the upper aerodigestive tract. METHODS: The meta-analysis combined 32 published reports to obtain an estimate of average treatment effect for salvage surgery with regard to survival, disease-free survival, surgical complications, and operative mortality. The prospective observational study included detailed data in 109 patients who underwent salvage surgery. In addition to parameters studied in the meta-analysis, we obtained baseline and interval quality of life data (Functional Living Index for Cancer [FLIC] scores), baseline and interval performance status evaluations (Performance Status Scale for Head and Neck Cancer Patients [PSS head and neck scores]), length of hospital stay, and hospital and physician charges, and related this data primarily to recurrent stage, recurrent site, and time to presalvage recurrence. RESULTS: The weighted average of 5-year survival in the meta-analysis was 39% in 1,080 patients from 28 different institutions. In the prospective study, median disease-free survival was 17.9 months in 109 patients, and this correlated strongly with recurrent stage, weakly with recurrent site, and not at all with time to presalvage recurrence. Noneconomic costs for patients and economic costs correlated with recurrent stage, but not with site. Baseline FLIC and PSS head and neck scores correlated with recurrent stage, but not with site. After salvage surgery the percentage of patients reaching or exceeding baseline was 51% for FLIC scores, and this differed significantly with recurrent stage. Postoperative interval "success" in PSS head and neck subscale scores for diet and eating in public also correlated with recurrent stage. CONCLUSIONS: Overall, the expected efficacy for salvage surgery in patients with recurrent head and neck cancer was surprisingly good, but success was limited and costs were great in stage III and, especially, in stage IV recurrences. A strong correlation of efficacy and noneconomic costs with recurrent stage allowed the creation of expectation profiles that may be useful to patients. Additional systematic clinical research is needed to improve results. In the end, the decision to undergo salvage surgery should be a personal choice made by the patient after honest and compassionate discussion with his or her surgeon. PMID- 10714712 TI - Clinical aspects of genetic variability in Helicobacter pylori. PMID- 10714713 TI - Psychiatrists help survivors in the Balkans. PMID- 10714714 TI - New twist in knee repair. PMID- 10714715 TI - HIV risk from oral sex higher than many realize. PMID- 10714716 TI - HIV's origins traced to 1930s. PMID- 10714717 TI - Clearing the way for new combination vaccine use. PMID- 10714718 TI - From the Centers for Disease Control and Prevention. Achievements in public health, 1900-1999: fluoridation of drinking water to prevent dental caries. PMID- 10714719 TI - Preventing medication errors in the intensive care unit. PMID- 10714720 TI - Preventing medication errors in the intensive care unit. PMID- 10714721 TI - Preventing medication errors in the intensive care unit. PMID- 10714722 TI - Fairness in fraud and abuse enforcement. PMID- 10714723 TI - Fairness in fraud and abuse enforcement. PMID- 10714724 TI - Follow-up of unsatisfactory Papanicolaou test results. PMID- 10714725 TI - Susceptibility to gonococcal infection during the menstrual cycle. PMID- 10714726 TI - A program to provide antiretroviral prophylaxis to health care personnel working overseas. PMID- 10714727 TI - Pott puffy tumor associated with intranasal methamphetamine. PMID- 10714728 TI - Effects of controlled-release metoprolol on total mortality, hospitalizations, and well-being in patients with heart failure: the Metoprolol CR/XL Randomized Intervention Trial in congestive heart failure (MERIT-HF). MERIT-HF Study Group. AB - CONTEXT: Results from recent studies on the effects of beta1-blockade in patients with heart failure demonstrated a 34% reduction in total mortality. However, the effect of beta1-blockade on the frequency of hospitalizations, symptoms, and quality of life in patients with heart failure has not been fully explored. OBJECTIVE: To examine the effects of the beta1-blocker controlled release/extended-release metoprolol succinate (metoprolol CR/XL) on mortality, hospitalization, symptoms, and quality of life in patients with heart failure. DESIGN: Randomized, double-blind controlled trial, preceded by a 2-week single blind placebo run-in period, conducted from February 14, 1997, to October 31, 1998, with a mean follow-up of 1 year. SETTING: Three hundred thirteen sites in 14 countries. PARTICIPANTS: Patients (n = 3991) with chronic heart failure, New York Heart Association (NYHA) functional class II to IV, and ejection fraction of 0.40 or less who were stabilized with optimum standard therapy. INTERVENTIONS: Patients were randomized to metoprolol CR/XL, 25 mg once per day (NYHA class II), or 12.5 mg once per day (NYHA class III or IV), titrated for 6 to 8 weeks up to a target dosage of 200 mg once per day (n = 1990); or matching placebo (n = 2001). MAIN OUTCOME MEASURES: Total mortality or any hospitalization (time to first event), number of hospitalizations for worsening heart failure, and change in NYHA class, by intervention group; quality of life was assessed in a substudy of 741 patients. RESULTS: The incidence of all predefined end points was lower in the metoprolol CR/XL group than in the placebo group, including total mortality or all-cause hospitalizations (the prespecified second primary end point; 641 vs 767 events; risk reduction, 19%; 95% confidence interval [CI], 10%-27%; P<.001); total mortality or hospitalizations due to worsening heart failure (311 vs 439 events; risk reduction, 31%; 95% CI, 20%-40%; P<.001), number of hospitalizations due to worsening heart failure (317 vs 451; P<.001); and number of days in hospital due to worsening heart failure (3401 vs 5303 days; P<.001). NYHA functional class, assessed by physicians, and McMaster Overall Treatment Evaluation score, assessed by patients, both improved in the metoprolol CR/XL group compared with the placebo group (P = .003 and P = .009, respectively). CONCLUSIONS: In this study of patients with symptomatic heartfailure, metoprolol CR/XL improved survival, reduced the need for hospitalizations due to worsening heart failure, improved NYHA functional class, and had beneficial effects on patient well-being. PMID- 10714729 TI - Methadone maintenance vs 180-day psychosocially enriched detoxification for treatment of opioid dependence: a randomized controlled trial. AB - CONTEXT: Despite evidence that methadone maintenance treatment (MMT) is effective for opioid dependence, it remains a controversial therapy because of its indefinite provision of a dependence-producing medication. OBJECTIVE: To compare outcomes of patients with opioid dependence treated with MMT vs an alternative treatment, psychosocially enriched 180-day methadone-assisted detoxification. DESIGN: Randomized controlled trial conducted from May 1995 to April 1999. SETTING: Research clinic in an established drug treatment service. PATIENTS: Of 858 volunteers screened, 179 adults with diagnosed opioid dependence were randomized into the study; 154 completed 12 weeks of follow-up. INTERVENTIONS: Patients were randomized to MMT (n = 91), which required 2 hours of psychosocial therapy per week during the first 6 months; or detoxification (n = 88), which required 3 hours of psychosocial therapy per week, 14 education sessions, and 1 hour of cocaine group therapy, if appropriate, for 6 months, and 6 months of (nonmethadone) aftercare services. MAIN OUTCOME MEASURES: Treatment retention, heroin and cocaine abstinence (by self-report and monthly urinalysis), human immunodeficiency virus (HIV) risk behaviors (Risk of AIDS Behavior scale score), and function in 5 problem areas: employment, family, psychiatric, legal, and alcohol use (Addiction Severity Index), compared by intervention group. RESULTS: Methadone maintenance therapy resulted in greater treatment retention (median, 438.5 vs 174.0 days) and lower heroin use rates than did detoxification. Cocaine use was more closely related to study dropout in detoxification than in MMT. Methadone maintenance therapy resulted in a lower rate of drug-related (mean [SD] at 12 months, 2.17 [3.88] vs 3.73 [6.86]) but not sex-related HIV risk behaviors and in a lower severity score for legal status (mean [SD] at 12 months, 0.05 [0.13] vs 0.13 [0.19]). There were no differences between groups in employment or family functioning or alcohol use. In both groups, monthly heroin use rates were 50% or greater, but days of use per month dropped markedly from baseline. CONCLUSIONS: Our results confirm the usefulness of MMT in reducing heroin use and HIV risk behaviors. Illicit opioid use continued in both groups, but frequency was reduced. Results do not provide support for diverting resources from MMT into long-term detoxification. PMID- 10714730 TI - Extraimmunization among US children. AB - CONTEXT: Little is known about the extent of extraimmunization, ie, vaccine doses given in excess of the recommended schedule, and whether it should be a public health concern. OBJECTIVES: To determine the extent and cost of extraimmunization in children and to identify its associated factors. DESIGN, SETTING, AND PARTICIPANTS: United States 1997 National Immunization Survey, in which telephone interviews were conducted with parents of 32742 19- to 35-month-old children and vaccination histories were collected from health care providers for 22806 of these children (overall response rate, 68.5%). Estimates were weighted to represent the full sample. MAIN OUTCOME MEASURES: Frequency of extraimmunization compared by vaccine type as well as with adequate immunization; factors associated with extraimmunization; and vaccine and visit costs associated with extraimmunization. RESULTS: Frequency of extraimmunization was less than 5% for each vaccine considered except poliovirus (14.1%). Overall, 21% of children were extraimmunized for at least 1 vaccine vs 31% underimmunized for at least 1 vaccine. In a multivariate model, the strongest contributors to extraimmunization were having more than 1 immunization provider (odds ratio [OR], 2.8; 95% confidence interval [CI], 2.4-3.2) and having multiple types of providers (eg, private and public health department; OR, 2.0; 95% CI, 1.6-2.4). Children seen only in public health department clinics were significantly less likely to be extraimmunized (OR, 0.3; 95% CI, 0.2-0.3). Annual costs associated with extraimmunization for this cohort of children were estimated conservatively at $26.5 million. CONCLUSIONS: These data indicate that extraimmunization can be costly. The challenge is to reduce extraimmunization without interfering with more important efforts to combat underimmunization. Improvements in immunization record keeping and sharing practices may help reduce extraimmunization. PMID- 10714731 TI - Monitoring osteoporosis therapy with bone densitometry: misleading changes and regression to the mean. Fracture Intervention Trial Research Group. AB - CONTEXT: The principle of "regression to the mean" predicts that patients with unusual responses to treatment might represent outliers who are likely to have more typical responses if treatment is continued without change. OBJECTIVE: To test whether women who lose bone mineral density (BMD) during the first year of treatment for osteoporosis continue to lose BMD if the same treatment is continued beyond 1 year. DESIGN AND SETTING: Two randomized, double-blind, placebo-controlled trials in 11 US clinical research centers for the Fracture Intervention Trial and 180 centers in the United States and other countries for the Multiple Outcomes of Raloxifene Evaluation Trial. PARTICIPANTS AND INTERVENTIONS: Postmenopausal women with low BMD assigned to treatment with 5 mg/d of alendronate sodium in the Fracture intervention Trial who completed 2 years of BMD monitoring and adhered to study medication (n = 2634), and postmenopausal women with osteoporosis assigned to treatment with 60 or 120 mg/d of raloxifene hydrochloride in the Multiple Outcomes of Raloxifene Evaluation trial who similarly completed 2 years of monitoring while adhering to study medication (n = 3954). MAIN OUTCOME MEASURES: Baseline, 12-, and 24-month hip and spine BMD. RESULTS: Women with the greatest loss of BMD during the first year of treatment were the most likely to gain BMD during continued treatment. Specifically, among women taking alendronate whose hip BMD decreased by more than 4% during the first year, 83% (95% confidence interval [CI], 82%-84%)had increases in hip BMD during the second year, with an overall mean increase of 4.7%. In contrast, those who seemed to gain at least 8% during the first year lost an average of 1% (95% CI, 0.1%-1.9%) during the next year. Similar results were observed among women taking raloxifene for 2 years. CONCLUSIONS: Our data suggest that most women who lose BMD during the first year of treatment with alendronate or raloxifene will gain BMD if the same treatment is continued for a second year. These results illustrate the principle of regression to the mean and suggest that effective treatments for osteoporosis should not be changed because of loss of BMD during the first year of use. PMID- 10714732 TI - Bipolar permanent magnets for the treatment of chronic low back pain: a pilot study. AB - CONTEXT: Chronic low back pain is one of the most prevalent and costly medical conditions in the United States. Permanent magnets have become a popular treatment for various musculoskeletal conditions, including low back pain, despite little scientific support for therapeutic benefit. OBJECTIVE: To compare the effectiveness of 1 type of therapeutic magnet, a bipolar permanent magnet, with a matching placebo device for patients with chronic low back pain. DESIGN: Randomized, double-blind, placebo-controlled, crossover pilot study conducted from February 1998 to May 1999. SETTING: An ambulatory care physical medicine and rehabilitation clinic at a Veterans Affairs hospital. PATIENTS: Nineteen men and 1 woman with stable low back pain of a mean of 19 years' duration, with no past use of magnet therapy for low back pain. Twenty patients were determined to provide 80% power in the study at P<.05 to detect a difference of 2 points (the difference believed to be clinically significant) on a visual analog scale (VAS). INTERVENTIONS: For each patient, real and sham bipolar permanent magnets were applied, on alternate weeks, for 6 hours per day, 3 days per week for 1 week, with a 1-week washout period between the 2 treatment weeks. MAIN OUTCOME MEASURES: Pretreatment and posttreatment pain intensity on a VAS; sensory and affective components of pain on the Pain Rating Index (PRI) of the McGill Pain Questionnaire; and range of motion (ROM) measurements of the lumbosacral spine, compared by real vs sham treatment. RESULTS: Mean VAS scores declined by 0.49 (SD, 0.96) points for real magnet treatment and by 0.44 (SD, 1.4) points for sham treatment (P = .90). No statistically significant differences were noted in the effect between real and sham magnets with any of the other outcome measures (ROM, P = .66; PRI, P = .55). CONCLUSIONS: Application of 1 variety of permanent magnet had no effect on our small group of subjects with chronic low back pain. PMID- 10714733 TI - Jockey injuries in the United States. AB - CONTEXT: In the sport of horse racing, the position of the jockey and speed of the horse predispose the jockey to risk of injury. OBJECTIVE: To estimate rates of medically treated injuries among professional jockeys and identify patterns of injury events. DESIGN: Cross-sectional survey from data compiled by an insurance broker. Information on the cause of injury, location on the track, and body part injured was evaluated. SETTING: Official races at US professional racing facilities (n = 114) from January 1, 1993, through December 31, 1996. PARTICIPANTS: A licensed jockey population of approximately 2700 persons. MAIN OUTCOME MEASURES: Annual injury incidence rates per 1000 jockey-years, as well as injury type, cause, and location on the track. RESULTS: A total of 6545 injury events occurred during official races between 1993 and 1996 (606 per 1000 jockey years). Nearly 1 in 5 injuries (18.8%) was to the jockey's head or neck. Other frequent sites included the leg (15.5%), foot/ankle (10.7%), back (10.7%), arm/hand (11.0%), and shoulder (9.6%). The most frequent location where injuries occurred was entering, within, or leaving the starting gate (35.1%), including 29.5% of head injuries, 39.8% of arm/hand injuries, and 52.0% of injuries to the leg/foot. Most head injuries resulted from being thrown from the horse (41.8%) or struck by the horse's head (23.2%). Being thrown from the horse was the cause of 55.1% of back and 49.6% of chest injuries. CONCLUSIONS: Our data suggest that jockeys have a high injury rate. Efforts are needed to reduce the number of potential injury events on the track and to improve protective equipment so events do not lead to injury. PMID- 10714734 TI - Strategies for long-term success in the treatment of HIV infection. AB - Highly active antiretroviral therapy has revolutionized the treatment of human immunodeficiency virus (HIV) infection, which can now be viewed as a chronic and manageable disease. However, HIV infection differs from other chronic diseases in that early treatment decisions can irrevocably alter the patient's response to future therapy. Despite the large number of approved antiretroviral agents, the number of sequential treatment regimens that will be effective for an individual patient is sharply limited by cross-resistance within the 3 drug classes. Because of the complexity of antiretroviral therapy, clinicians prescribing it require considerable expertise. Treatment should be deferred until the patient has been educated about the importance of strict adherence and has demonstrated willingness and motivation to begin therapy. Drug regimens should be chosen that the patient can tolerate and adhere to, and the consequences of resistance should be considered before therapy is begun. When treatment fails, the timing and choice of subsequent therapy can be critical in determining the magnitude and durability of response. Resistance testing can help guide the clinician in the choice of therapy. In patients who have been treated with numerous antiretroviral agents, it may be impossible to achieve significant viral suppression. Therapy may still be beneficial for such patients, but it should be tolerable and should not increase resistance to drugs that may become available in the near future. Drug resistance and treatment failure are not random events, but are the result of factors over which clinicians and their patients have some control. The treatment of drug-resistant patients is challenging; the best way to deal with resistance is to prevent it. PMID- 10714735 TI - Beta-blocker therapy for heart failure: the evidence is in, now the work begins. PMID- 10714736 TI - Treatment for opioid dependence: quality and access. PMID- 10714737 TI - Vaccine extraimmunization--too much of a good thing? PMID- 10714738 TI - Provision of methadone treatment in primary care medical practices: review of the Scottish experience and implications for US policy. AB - CONTEXT: Under new proposed regulations, US physicians outside of traditional methadone clinics could prescribe methadone to patients with opioid dependence. No large-scale evaluations of US programs in which methadone maintenance is provided by primary care physicians are available, but primary care physicians in Scotland have participated in such programs on a large scale. OBJECTIVE: To review the history, operation, and outcome data on the efficacy and safety of 2 Scottish primary care-based opioid agonist treatment programs to derive lessons for the US context. DESIGN AND SETTING: Naturalistic study of programs in Edinburgh and Glasgow, Scotland, with data obtained through site visits and interviews conducted in 1996 and 1998, as well as from published reports and retrospective analysis of electronic databases. MAIN OUTCOME MEASURES: Proportions of injection drug users who were enrolled in the methadone maintenance programs, average methadone doses in the programs, and methadone related deaths. RESULTS: A total of 60% to 80% of injection drug users in Edinburgh and 41% to 73% of those in Glasgow were enrolled in methadone maintenance in 1998-1999. Dose levels are consistent with US recommendations (for Edinburgh in 1998, 61 mg; for Glasgow in 1994-1996, 54 mg). The Glasgow program required supervised consumption of methadone in community pharmacies for the first year and experienced significantly fewer methadone-related deaths than Edinburgh in 1997 (17 vs 30 deaths; P<.0001). Programs in both Edinburgh and Glasgow provided support to primary care physicians and achieved levels of general practitioner participation of 59% (1998) and 30% (1999), respectively. CONCLUSIONS: The Scottish experience indicates that prescription of methadone in office-based settings can expand access to an important treatment modality. Primary care physicians safely prescribed methadone for maintenance treatment when provided with adequate support. Diversion of methadone was minimized by requiring supervised consumption in community pharmacies. PMID- 10714739 TI - JAMA Patient Page: drug abuse. PMID- 10714740 TI - The role of qualitative research in a national project on decision making about hysterectomy and the use of hormone replacement therapy. PMID- 10714741 TI - Overview of women's decision making regarding elective hysterectomy, oophorectomy, and hormone replacement therapy. AB - Over 600,000 hysterectomies are performed each year in the United States, the majority of which are to improve quality of life for perimenopausal women. Hysterectomy rates for common conditions differ between African American and white women, and African American women undergo surgery at a younger age for most diagnoses. Many hysterectomies are accompanied by elective oophorectomy, and hormone replacement therapy (HRT) is commonly used, especially among women experiencing surgical menopause, despite questions about its long-term benefits and risks. Despite the high rates of hysterectomy in the United States, little is known about how women make decisions regarding this surgery and, in particular, how ethnic and cultural factors may influence these decisions. This article provides a review of what is currently known about the epidemiology of hysterectomy, oophorectomy, and HRT use and identifies gaps in knowledge about women's decision making, with a special focus on ethnic variations and cultural influences, issues addressed by the Ethnicity, Needs, and Decisions of Women (ENDOW) project. PMID- 10714742 TI - A qualitative study of women's hysterectomy experience. AB - The purpose of this qualitative study was to elicit women's perceptions of their experiences with hysterectomy, oophorectomy, and surgical menopause. Focus group and individual interviews were used to obtain data from a sample of southern urban women who had had hysterectomies for benign reasons. Of the 38 women who participated, 22 were African American and 16 were Caucasian, the mean age was 48 years, and most were low to middle income. Findings revealed that biophysical, psychosocial, and spiritual domains were important in the decision to have a hysterectomy. For many, the choice to have a hysterectomy was a last resort and was viewed as a technique that could relieve a myriad of symptoms. Although most participants described the hysterectomy experience as positive, they expressed a variety of concerns from diagnosis through recovery. Participants expressed a need for information about women's gynecological health for themselves and their male partners. African American women expressed a need for change in attitudes and beliefs in the black community about women undergoing hysterectomy. Many spouses, brothers, uncles, and other African American male friends were nonsupportive, and a few women revealed that they had not told a new partner about the surgery. The findings have implications for women's healthcare providers. Provider training and education are needed that integrate biophysical care of women with the psychological, sociological, and spiritual domains. Efforts must be directed to the community to enlighten men and families about hysterectomy by dispelling myths and providing current health information related to women's gynecological health and alternatives to, indications for, and types of hysterectomy. PMID- 10714743 TI - Women's stories: ethnic variations in women's attitudes and experiences of menopause, hysterectomy, and hormone replacement therapy. AB - To increase understanding of women's midlife changes, 23 focus groups were held to investigate the possible ethnic variations in attitude toward and experience with menopause, hysterectomy, and hormone replacement therapy (HRT) in non Hispanic white, Hispanic, and Navajo women in New Mexico. The medical definition of menopause, no menstrual bleeding for 12 months, did not coincide with the women's definition of menopause as the hormonal fluctuations they experienced before, during, and after any change in their menstrual cycle. More women reported having to fight to have a hysterectomy than having one unnecessarily. Women complained about the lack of information and preparation prior to having a hysterectomy and expressed dissatisfaction with doctor-patient communication, but they were satisfied with their hysterectomy because they felt better after the surgery. Although women in the study reported that menopause and hysterectomy are seldom discussed openly, they all participated freely in the storytelling focus groups. The most traditional women, primarily rural Navajo and newly immigrated Latina, related few or no menopausal symptoms with natural menopause or after hysterectomy. Many of these women had not even heard of HRT. Many women who had been prescribed HRT expressed dissatisfaction with the side effects and dosage. Unsupervised tinkering with the dosage was the rule rather than the exception. The study revealed that women are much more alike than they are different. Traditional women in all ethnic groups had more in common with each other than they did with the least traditional women in their own ethnic group. PMID- 10714744 TI - Decision making, beliefs, and attitudes toward hysterectomy: a focus group study with medically underserved women in Texas. AB - Variations in hysterectomy rates have been associated with assorted physician and patient characteristics, and the disproportionate rate of hysterectomies in African American women has been attributed to a higher prevalence of leiomyomas. The role of women's beliefs and attitudes toward hysterectomy and participation in decision making for medical treatment has not been explored as a source of variance. The purposes of this qualitative study were to explore these constructs in a triethnic sample of women to understand beliefs, attitudes, and decision making preferences among underserved women; to facilitate development of a quantitative survey; and to inform development of interventions to assist women with such medical decisions. Twenty-three focus groups were conducted with 148 women from community sites and public health clinics. Thirteen self-identified lesbians participated in three groups. Analysis of audiotaped transcripts yielded four main themes: perceived outcomes of hysterectomy, perceived views of men/partners, opinions about healthcare providers, decision-making process. Across groups, the women expressed similar expectations from hysterectomy, differing only in the degree to which dimensions were emphasized. The women thought men perceived women with hysterectomy as less desirable for reasons unrelated to childbearing. Attitudes toward physicians were negative except among Hispanic women. All women expressed a strong desire to be involved in elective treatment decisions and would discuss their choice with important others. Implications for intervention development include enhancing women's skills and confidence to evaluate treatment options and to interact with physicians around treatment choices and creation of portable educational components for important others. PMID- 10714745 TI - The role of male partners in women's decision making regarding hysterectomy. AB - Although hysterectomy is a frequently performed surgical procedure, little is known about how women make decisions regarding hysterectomy. This report details the women's perceptions of male partners' knowledge and attitudes about hysterectomy and the role women expect or allow men to play in their decision making process. Seventeen focus groups were conducted with a total of 82 African American and Caucasian women aged 30-65 years in two coastal counties of South Carolina. Transcripts were coded and analyzed using the nonnumerical unstructured data indexing searching and theory building (QSR NUD*IST) software program. Results indicate that women perceive men to be not well informed or knowledgeable about hysterectomy, to be concerned about the quality of sexual relations after hysterectomy, and, in some cases, to be neutral about hysterectomy. African American women reported that men hold more negative perceptions about hysterectomized women. Caucasian women stressed men's inability to understand what a woman is going through and men's concern with the hysterectomy's effect on their own egos. Nonhysterectomized women felt that men would be more bothered by a surgical procedure that left more visible effects (such as mastectomy). These women defined a limited role for men in their decision making regarding hysterectomy, consisting of discussion and offering of support/sympathy, but they reserved the actual decision for themselves. In a few instances, women accorded men a role in the hysterectomy decision based on a religious interpretation of marriage. Intervention programs are recommended that target women and their partners together, using hysterectomized women and their partners as peer educators. PMID- 10714746 TI - Talking about hysterectomy: the experiences of women from four cultural groups. AB - As part of the Ethnicity, Needs, and Decisions of Women (ENDOW) project, in-depth qualitative interviews and focus groups were conducted at four sites, Alabama, New Mexico, South Carolina, and Texas. In South Carolina and Alabama, African American and white women were interviewed. In Texas, African American, Caucasian, and Hispanic women were interviewed, and in New Mexico, focus groups with Caucasian, Hispanic, and Navajo women were conducted. The Texas site also conducted focus groups with lesbian women. Data were collected on women's experiences with and attitude toward menopause, hysterectomy, and hormone replacement therapy (HRT). Information also was gathered on women's concerns and what experiences they have had or expect to have with healthcare providers and what they perceive their friends', families', and sexual partners' attitudes are toward hysterectomy. Numerous commonalties of experience existed across racial and ethnic groups. Overall, the women who participated believed that doctors do not take the time to explain issues related to menopause, hysterectomy, and HRT. Most of the women who have had a hysterectomy were satisfied with the outcome of surgery, as painful symptoms were relieved. There are also several interesting differences among the groups. Decision-making patterns differed among the ethnic groups, as did experience with healthcare providers. Many women in the focus groups expressed mistrust of or negative opinions of healthcare providers. African Americans expressed mistrust of their motives for recommending surgery, as did several of the Caucasian, non-Hispanic women. Most of the Hispanic participants respected and trusted their providers. All groups said they would seek additional medical opinions if they could afford to do so. PMID- 10714747 TI - New aspects of proinsulin physiology and pathophysiology. PMID- 10714748 TI - Minimally invasive surgery in pediatric endocrinology. AB - The use of minimally invasive surgery (MIS) in children and adolescents is steadily increasing. The aim of the present review was to summarize the status of MIS in pediatric endocrinology. We found that laparoscopic procedures have been proven useful for the diagnosis or treatment of endometriosis and its associated manifestations, undescended testicles, ambiguous genitalia, adnexal torsion and ovarian cyst. Considering the safety and efficacy of these applications, the more rapid recovery of the patients, and the considerably less pain induced, we believe MIS will gradually take precedence over standard procedures in many areas of endocrinology in the young population. PMID- 10714749 TI - Thyroid gland volume and urinary iodine excretion in children 6-11 years old in an endemic area. AB - Goiter prevalence and urinary iodine excretion levels were assessed in 605 schoolchildren (301 males and 304 females), aged 6-11 years, living in the Antalya region, a well known endemic goiter area in Turkey. Goiter prevalence was evaluated by clinical examination and ultrasound of the thyroid gland. Urinary iodine levels were expressed as microg/g creatinine. Goiter by inspection and palpation was found in 35% (n = 212) of all subjects, in 37.5% (n = 114) of girls and 32.5% (n = 98) of boys. Iodine deficiency of moderate degree was detected from the point of goiter prevalence. With regard to the upper limits of reference thyroid volumes reported by WHO and ICCIDD, goiter by ultrasonography was found in 34% (n = 206) of all subjects, in 36.8% (n = 112) of girls and 31% (n = 94) of boys. Median iodine/creatinine ratios of all subjects, and goitrous and non goitrous subjects, were 64.1+/-20.1, 62.8+/-21.8 and 64.9+/-19.1 microg/g, respectively. Urinary iodine excretion levels revealed mild iodine deficiency in the region. No significant correlation was observed between urinary iodine excretion levels and thyroid volumes (r = 0.12, p>0.05). Iodine deficiency of mild to moderate degree in schoolchildren aged 6-11 years was detected in Antalya. It was concluded that urgent measures must be undertaken to eradicate iodine deficiency in the region. PMID- 10714750 TI - Increased 5alpha-reductase and normal 11beta-hydroxysteroid dehydrogenase metabolism of C19 and C21 steroids in a young population with polycystic ovarian syndrome. AB - OBJECTIVE: To test the hypothesis that 5alpha-reductase (5alphaR) and 11beta hydroxysteroid dehydrogenase (11beta-HSD) activity are increased in adolescent and young-adult women with PCOS and that an altered regulation of the hypothalamic-pituitary-adrenal (HPA) axis occurred in these subjects. DESIGN: Prospective non-randomized study in an academic research environment. PATIENTS: Eleven women, aged 14 to 25 years, were studied who were at least one year post menarche and who had a diagnosis of PCOS based on a history of oligomenorrhea and elevated total and or free serum testosterone. INTERVENTION: 24-Hour urinary metabolites were assessed in nine subjects and five underwent stimulation with ovine corticotropin releasing factor (oCRF). OUTCOME MEASURES: C19 and C21 steroid urinary metabolite 5-alpha/5-beta pairs, 11-oxo/11-hydroxy products and the ratio of the total 5-alpha/5-beta reduced and 11-oxo/11-hydroxy products were compared to values in control women. Urinary cortisol (F) (sum of conjugated and free, and free F) and total F metabolites (the sum of THE, THF, 5alpha-THF, cortolones, and cortols) were determined. A 1 microg/kg oCRF stimulation test was performed with timed samples determined for plasma ACTH and serum F levels. RESULT: The 24-hour total and free urinary F were not different from control. However, the total F metabolites were markedly elevated (7922+/-2666 vs 5418+/ 1549 microg/24 h, p<0.01). A marked increase in the total 5-alpha reduced C19 and C21 metabolites was observed in the PCOS population vs control (5084+/-1977 vs 2681+/-1188 microg/24 h, p<0.01). The total urinary 11-oxo/11-hydroxy metabolite ratio was not different, p=0.23. The basal values and response of both ACTH and F to oCRF stimulation were not different from those of controls. CONCLUSION: There is a marked increase in 5alphaR metabolism of both C19 and C21 steroids in younger women with PCOS. PMID- 10714752 TI - Fetal size to final height in Hong Kong Chinese children. AB - OBJECTIVES: It has been known that size at birth is important for postnatal growth and final height. However, there are few data in the literature on the difference in height growth patterns from fetal size to final height between less privileged and more privileged populations. The aim of this study was to describe the important features in height growth from birth to maturity in an underprivileged Hong Kong Chinese cohort in comparison to a more privileged Swedish cohort. METHODS: The longitudinal height growth data from birth to maturity in full-term healthy Hong Kong Chinese children (n=132) who were born in 1967 were analyzed, and compared with those for Swedish children who were born in 1973-75 (n=3650). RESULTS: Children with longer birth length achieved taller adult stature with respect to their target height. The mean final height retained the same order as that of the mean length at birth for various birth length groups. All children in the Hong Kong Chinese series showed catch-down height growth during the first 2 years of life, in contrast to the catch-up in smaller babies and catch-down in larger babies for the Swedish series. The growth deficit for the Hong Kong Chinese was -0.9 SDS at birth, -1.8 SDS at 2.0 years of age, 2.1 SDS at 8 years of age, and -1.7 SDS at final height. CONCLUSIONS: Fetal size is important for postnatal growth and attained final height with respect to a child's familial genetic potential in stature, not only for privileged populations, but also for underprivileged populations. However, children in underprivileged populations experience a persistent increasing growth deficit during infancy and childhood. Special attention should be given to monitor their growth status in early years and to institute appropriate intervention programs. PMID- 10714751 TI - Peripubertal prolactinomas: clinical presentation and long-term outcome with different therapeutic approaches. AB - The evolution of prolactinomas in children and adolescents continues to be controversial. We report on the long-term evolution (2-20 yr) of prolactinomas in 40 patients (29 F, 11 M). In females, the age for the onset of symptoms ranged between 8 and 16 yr and the age at which diagnosis was made ranged from 15 to 19 yr; in males, ages ranged from 8 to 17 yr and from 13.8 to 19 yr, respectively. In females, there was predominance of microprolactinomas (22/ 29) and the symptomatology resulted from functional disorders, whereas in males there was predominance of macroprolactinomas (8/11) and symptoms were caused by tumor mass disorders. Surgery was used as primary therapy in nine patients and as supplemental therapy in six patients. Twenty-four patients were treated primarily with bromocriptine and seven with cabergoline. Of the nine patients treated primarily with surgery, only one achieved gonadotropic axis restoration; in 25/31 patients receiving drug therapy gonadotropic function was restored to normal. Fifteen patients showed complete resolution or substantial shrinkage of tumor. CONCLUSION: In pediatric and adolescent age, there seem to be age- and sex dependent differences in the clinical presentation of prolactinomas that cannot be accounted for only in terms of time of evolution. Drug therapy can control the disease, normalize prolactin levels and achieve gonadotropic axis restoration in most patients. PMID- 10714753 TI - Suppression and recovery of GH secretion after GH injection in non-GH deficient short children. AB - OBJECTIVE: To identify the effect of exogenous GH on endogenous GH secretion in 48 non-GH deficient short children participating in a placebo-controlled trial of GH therapy on final adult height. DESIGN: Night GH secretion (mean of levels every 20 min from 20.00 to 08.00 h) was evaluated at baseline, 6 months before starting placebo or GH (somatotropin, 0.222 mg/kg/ week, divided into 3 doses each week). At 6 months after starting injections, blood samples for GH were obtained hourly for 24 h after an injection, and every 20 min on each of the next two nights (with no additional placebo or GH injection). RESULTS: IGF-I levels in the treatment group were elevated at 12 and 24 h but not at 36 h compared to the placebo group. Mean GH levels in the placebo group did not vary significantly among the four sampling periods. In the treatment group, the mean serum GH rose to a supraphysiological peak at an average time of 4 h after injection. Subsequently, mean GH level was significantly suppressed compared to placebo on the second night following GH injection, but returned to normal by the third night. CONCLUSION: After 6 months of a thrice weekly GH treatment regimen in non GH deficient short children, endogenous GH secretion was reduced from 24 to 36 h after injection compared to placebo and returned to control levels by 48 to 60 h after injection. PMID- 10714754 TI - The relationship of the levels of leptin, insulin-like growth factor-I and insulin in cord blood with birth size, ponderal index, and gender difference. AB - In humans, serum levels of leptin correlate with total body fat in both adults and children. After collecting cord blood from 156 term neonates (82 males, 74 females; 132 AGA and 22 LGA), we measured the cord levels of leptin, insulin and IGF-I to determine the relationships between these three hormones and relationships of these hormones with birth size (birth weight and ponderal index for adiposity in newborn) and gender. The leptin and IGF-I levels were significantly higher in the LGA group (9.2+/-4.0 ng/ml and 96.1+/-34.1 ng/ml, respectively) than in the AGA group (4.8+/-3.8 ng/ml and 56.4+/-37.6 ng/ml, respectively). A significant positive correlation was observed between leptin levels and birth weight, and a weaker correlation between leptin levels and birth height. IGF-I level significantly correlated with birth weight and birth height, but there was no correlation between the levels of insulin and birth weight. There was no relationship between the levels of IGF-I, insulin and leptin. Ponderal index was higher in LGA than in AGA. A significant correlation was also observed between the levels of leptin and ponderal index, but not between the levels of insulin or IGF-I and ponderal index. The levels of leptin and ponderal index were higher in females than males despite no gender differences in gestational age and birth weight. In conclusion, our results suggest that the level of IGF-I is a useful index for fetal growth during late gestation, and the development of adipose tissue is the major determinant of fetal serum leptin levels, the production of which is not regulated by insulin or IGF-I. In addition, a gender difference with a higher level of leptin in female neonates suggests that sexual dimorphism in adipose tissue already exists in utero. PMID- 10714755 TI - Epidemiology of diabetes mellitus in children in Hong Kong: the Hong Kong childhood diabetes register. AB - OBJECTIVES: To establish a registry for Chinese children with onset of type 1 (insulin dependent) diabetes mellitus before 15 years of age and to determine the incidence of childhood onset type 1 diabetes mellitus in Chinese children in Hong Kong. RESEARCH DESIGN AND METHODS: A registry was established in 1997 to collect childhood diabetes cases retrospectively from all districts in Hong Kong. The study included all newly diagnosed cases of diabetes with onset < 15 yr of age from 1st January 1984 to 31 December 1996. Primary ascertainment was based on review of medical records at all regional public hospitals in Hong Kong and survey of all the registered practitioners in Hong Kong. The secondary source of validation was made impractical, if not impossible, because of the recent implementation of the Personal Data Privacy Ordinance in Hong Kong. RESULTS: A total of 255 diabetic cases were identified, 227 type 1 diabetes mellitus (218 were Chinese), 18 type 2 diabetes mellitus and 11 secondary diabetes. 246 patients were Chinese and 9 non-Chinese. The age-standardized incidence of type 1 and type 2 diabetes mellitus in southern Chinese children in Hong Kong was 1.4/100,000/yr and 0.1/100,000/yr respectively for children < 15 yr of age during the study period. The incidence rates for type 1 diabetes were 0.9, 1.5 and 1.7 per 100,000/yr for 0-4 years, 5 to 9 years and 10 to 14 years age-groups respectively. The incidence for males was 1.2/100,000/yr and for females 1.7/100,000/yr. A significant increase in the incidence was demonstrated during the study period by simple linear regression (slope 0.14/100,000/year, r2 = 0.73, p = 0.0002) CONCLUSIONS: A diabetic registry is established in Hong Kong. This study documents a very low incidence rate of childhood type 1 diabetes mellitus in southern Chinese children in Hong Kong and we have seen an increasing incidence of the disease in the past 13 years. PMID- 10714756 TI - Ultrasonography of the pancreas, as a function index, in children with beta thalassemia. AB - Increased echogenicity of the pancreas, due to hemosiderosis, is a frequent laboratory finding in children and adolescents with beta-thalassemia. The aim of this study was to investigate whether increased echogenicity of the pancreas is associated with dysfunction. The ultrasonic image of the pancreas was examined in 34 children aged 12+/-3.8 years old and was compared to the endocrine and exocrine functioning of the gland. Oral glucose tolerance test (OGTT) was performed with simultaneous measurement of insulin and serum trypsin. Twenty-six of the 34 patients (76.5%) presented increased echogenicity, while 8 (23.5%) had a normal ultrasonic pancreatic image. 77% of the patients with increased echogenicity had abnormal OGTT, 46%, with subnormal or increased insulin values, and 32.5% manifested low levels of trypsin. Among the patients with normal ultrasound, 25% had abnormal OGTT and 37.5% abnormal insulin values. Statistical analysis with Student's t-test revealed that patients with increased echogenicity had significantly higher glucose values on OGTT at 60: 7.6 +/- 1.8 mmol/l (137.3 +/- 33.7 mg/dl) as compared to the patients with normal ultrasound: 6.1 +/- 1.2 mmol/l (110.75 +/- 21.72 mg/dl) (p<0.05). Insulin values were significantly affected at 30, 60, and 90 min: 570+/-301, 332+/-156, 294+/-158 pmol/l (79.54 +/- 42, 46.4 +/- 21.8, 41.04 +/- 22 mU/l) respectively in patients with increased echogenicity in comparison to those with normal ultrasonographic image of the gland: 301 +/- 170, 192 +/- 52, 135 +/- 63 pmol/l (42 +/- 23.7, 26.85 +/- 7.36, 18.9 +/- 8.8 mU/l) (p<0.05). No statistical significance was observed between the two groups regarding trypsin levels, even though abnormal values were observed in more children with increased echogenicity than in patients with a normal ultrasound. The above findings confirm that increased echogenicity of the pancreas is associated with disturbance of its function. This simple imaging method could be used as a rough early index of detection of an increased risk for developing diabetes mellitus in patients with beta-thalassemia. PMID- 10714757 TI - Bone mineral density in childhood obesity. AB - There are several metabolic and hormonal disturbances in childhood obesity. The purpose of this study was to determine the relationship between childhood obesity and bone mineral density (BMD). We studied BMD in 37 obese children and in 37 non obese children. BMD was measured at L2-L4 level by using dual energy X-ray absorptiometry. BMD was significantly related to age, height and weight. The mean BMD in the obese children and control group was 0.655 +/- 0.175 and 0.626 +/- 0.159 g/cm2, respectively, without any statistically significant difference (p>0.05). There was no correlation between BMD values and osteocalcin or calcitonin levels. According to Tanner's pubertal staging, the mean BMD of pubertal obese children was higher than that of prepubertal obese children. BMD of the pubertal obese children was significantly higher than that of the pubertal control group (p<0.05). Girls had higher mean BMD values than boys. In conclusion, our results show that BMD is not influenced by obesity in children but higher values in puberty were observed in obese children which may due to hormonal changes. PMID- 10714758 TI - A comparison of the use of premixed insulins in pen-injectors with conventional patient-mixed insulin treatment in children and adolescents with IDDM. Is there a decreased risk of night hypoglycemia? AB - Insulin injection is a problem in pediatric and adolescent age, and premixed insulin therapy given in pen-injectors (Novopen II) is expected to increase compliance. Compliance with treatment and safety of this kind of insulin substitution was investigated in 20 IDDM patients (8.2-19.6 years old). The study was of randomized cross-over design and its duration was 6 (2x3) months. Metabolic parameters were compared between premixed insulin therapy via pen injector and patient-mixed insulin therapy via conventional syringe, and no differences were observed except for the postponing of night hypoglycemic attacks to 07.00 a.m. during premixed insulin therapy. No technical or medical problems occurred. Patients were more satisfied with the new therapy regimen as determined by questionnaire. We concluded that this kind of insulin substitution is safe in pediatric and adolescent IDDM patients. PMID- 10714759 TI - Islet autoantibodies and insulin dependent diabetes mellitus in cystic fibrosis. AB - Cystic fibrosis-related diabetes mellitus (CF-DM) is thought to be secondary to beta-cell destruction by fibrous tissue replacing the exocrine pancreas. The aim of this study was to investigate the hypothesis that other factors may also be responsible. Glutamic acid decarboxylase (GAD) and islet cell (IA-2) antibodies were measured by quantitative ELISA in a group of patients with CF (n=30) in comparison to a group of newly diagnosed DM type 1 (IDDM) patients (n=30) and normal subjects (n=30). GAD antibodies were positive (>32 ng/ml) in 50% of the CF, 93% of the IDDM and 0% of the control group. IA-2 antibodies were detected (>0.9 U/ml) in 40% of the CF, 93% of the IDDM and 0% of the control group. Among the fifteen CF patients with positive GAD and IA-2 antibodies, four already had IDDM and another five abnormally low (<45 mU/l) first phase insulin response (FPIR) indicating a prediabetic state. We conclude that factors other than mechanical may be involved in the development of CFDM. The presence of autoantibodies predicting IDDM supports the hypothesis that CF-DM may have a multifactorial pathogenesis. PMID- 10714760 TI - Multiple endocrine abnormalities in a child with Blackfan-Diamond anemia and hemochromatosis. Significant improvement of growth velocity and predicted adult height following growth hormone treatment despite liver damage. AB - We evaluated a short, prepubertal 13.9 year-old boy with Blackfan-Diamond anemia and significant liver iron stores due to multiple blood transfusions and found him to have several endocrine abnormalities, including hypothyroidism, hypoparathyroidism, primary and secondary hypogonadism and IGF-I insufficiency. Growth velocity was poor despite treatment with levothyroxine, calcitriol, calcium and aggressive therapy with chelating agents. After 25 months of treatment with rhGH his growth velocity, height for age and PAH increased significantly, suggesting a degree of sensitivity to GH despite his liver damage. PMID- 10714761 TI - Triple A syndrome mimicking cystic fibrosis. AB - We report a 2-8/12 year-old male who presented with symptoms resembling cystic fibrosis (failure to thrive, developmental delay and recurrent diarrhea) and had elevated sweat chloride concentration. Mucosal hyperpigmentation led to the diagnosis of adrenal insufficiency which was ultimately shown to be a component of triple A syndrome (achalasia, alacrima, adrenal insufficiency). Elevated sweat chloride concentration normalized after initiation of adrenal replacement therapy. We suggest that non-CF conditions causing elevated sweat chloride concentration should be considered in patients with atypical findings or who do not have objective evidence of pulmonary or exocrine pancreatic disease. PMID- 10714762 TI - VII International "Onnela" Workshop: gut immune system and type 1 diabetes mellitus held in Janakkala, Finland, June 25-26, 1998. PMID- 10714763 TI - Transcriptional regulation of the cellular retinoic acid binding protein I gene in F9 teratocarcinoma cells. AB - Retinoic acid (RA) induces the differentiation of many murine teratocarcinoma cell lines such as F9 and P19. In F9 cells, the level of the cellular retinoic acid binding protein I (CRABP I) mRNA is greatly reduced after exposure of the cultured cells to exogenous RA. In P19 cells, the level of CRABP I mRNA is greatly increased after RA exposure. We have identified a 176-bp region in the murine CRABP I promoter, between -2.9 and -2.7 kb 5' of the start site of transcription, which acts as an enhancer in undifferentiated F9 stem cells and through which RA effects inhibition of CRABP I transcription. Within this region are two footprinted sites at -2763 and -2834. This 176-bp regulatory region does not function to enhance CRABP I transcription in P19 stem cells. Several DNA sequences within these two footprinted regions bind proteins from F9 nuclear extracts but not from P19 nuclear extracts (e.g., FP1B, FP1A, and FP2B), as assessed by gel shift assays. This 176-bp CRABP I genomic region has not been sequenced previously and functionally analyzed in cultured cells because it was not present in the murine CRABP I clones used for the promoter analyses reported earlier by another laboratory. The function of this enhancer may be to reduce the expression of the CRABP I gene in specific embryonic cell types in order to regulate the amount of RA to which the cells are exposed. PMID- 10714764 TI - Adherence of human erythroleukemia cells inhibits proliferation without inducing differentiation. AB - To investigate the effect of extracellular matrix molecules in the megakaryocytic lineage, we studied the role of integrin engagement in the proliferation and differentiation of human erythroleukemia (HEL) cells. HEL cells grew in suspension, but their adherence depended upon the presence of matrix proteins or protein kinase C signaling. Adherence by itself did not trigger commitment of these cells but accelerated phorbol 12-myristate 13-acetate-induced differentiation. HEL cells adhered to fibronectin mainly through alpha5beta1, and this receptor acted synergetically with alpha4beta1. Integrin engagement induced cell growth arrest through mitogen-activated protein kinase inactivation. Such down-regulation of the mitogen-activated protein kinase pathway by integrin engagement was suggested as a megakaryocytic-platelet lineage specificity. This signaling was not restricted to a peculiar integrin but was proposed as a general mechanism in these cells. PMID- 10714765 TI - A role for E2F1 in the induction of apoptosis during thymic negative selection. AB - Thymic negative selection is the process in which maturing thymocytes that express T-cell receptors recognizing self are eliminated by apoptotic cell death. The molecular mechanism by which this occurs is poorly understood. Notably, genes involved in cell death, even thymocyte death, such as Fas, Fas-ligand, p53, caspase-1, caspase-3, and caspase-9, and Bcl-2 have been found to not be required for normal thymic negative selection. We have demonstrated previously that E2F1 deficient mice have a defect in thymocyte apoptosis. Here we show that E2F1 is required for normal thymic negative selection. Furthermore, we observed an E2F1 dependent increase of p53 protein levels during the process of thymic clonal deletion, which suggests that E2F1 regulates activation-induced apoptosis of self reactive thymocytes by a p53-dependent mechanism. In contrast, other apoptotic pathways operating on developing thymocytes, such as glucocorticoid-induced cell death, are not mediated by E2F1. The T lymphocytes that escape thymic negative selection migrate to the peripheral immune system but do not appear to be autoreactive, indicating that there may exist E2F1-independent mechanisms of peripheral tolerance, which protect mice from developing an autoimmune response. We expect that E2F1-deficient mice will provide a useful tool for understanding the molecular mechanism of and the immunological importance of thymic negative selection. PMID- 10714766 TI - Inhibition of mitogen-activated protein kinase and phosphatidylinositol 3-kinase activity in MCF-7 cells prevents estrogen-induced mitogenesis. AB - Estrogen acts to promote DNA synthesis in the MCF-7 human breast cancer cell line via its interaction with high levels of estrogen receptor. The primary mode of estrogen action has been considered to be through transcriptional activation of genes containing estrogen response elements, including the immediate early genes c-myc and fos. Recent reports have indicated that estrogen, acting through the estrogen receptor, is capable of inducing the mitogen-activated protein kinase (MAPK) cytoplasmic signaling cascade. In this study, specific small molecule inhibitors of MAPK and phosphatidylinositol 3-kinase activity were used to determine the influence of these cascades on estrogen-mediated mitogenesis. Phosphatidylinositol 3-kinase inhibitors, LY294002 and wortmannin, as well as inhibitors of MAPK kinase-1, PD098059 and U0126, decreased the fraction of cells entering DNA synthesis after treatment with 17beta-estradiol. These compounds did not inhibit expression of myc or fos. However, the drugs did prevent the accumulation of cyclin D1 and hyperphosphorylated retinoblastoma protein, indicating that the block occurred at, or prior to, this point in the cell cycle. Although these compounds were effective in preventing estrogen-mediated mitogenesis, the downstream kinases extracellular signal-regulated kinase 1, extracellular signal-regulated kinase 2, and protein kinase B were not activated over basal levels by estrogen treatment. These studies suggest that estrogen initiates mitogenesis by inducing the transcription of immediate early genes, but cytoplasmic signaling pathways play an important role in the control of subsequent events in the cell cycle. PMID- 10714767 TI - Epidermal overexpression of granulocyte-macrophage colony-stimulating factor induces both keratinocyte proliferation and apoptosis. AB - Granulocyte-macrophage colony-stimulating factor (GM-CSF) is released by keratinocytes in sizeable amounts only under pathological conditions, e.g., after topical application of a tumor promoter, in atopic dermatitis (AD), and after wounding. To study the biological function of this cytokine release, we generated transgenic mice that constitutively overexpress GM-CSF in the epidermis. An increase in the numbers of mast cells and Langerhans cells (LCs) in transgenics versus nontransgenic controls was observed but no severe inflammation. This is consistent with a central role of this cytokine in the development and maturation of LCs. Mitotic activity in the epidemnis of transgenic mice was elevated, but epidermal thickness and differentiation were normal. Homeostasis is maintained by an increase of apoptosis in the epidermis. We describe the differential expression of regulators of apoptosis and discuss a potential mechanism for this novel proapoptotic activity of GM-CSF on keratinocytes. Both stimulation of proliferation and promotion of apoptosis are of great relevance to tumorigenesis. The latter may be a means of removing damaged cells after genotoxic stress or injury. PMID- 10714768 TI - Cooperative effect of hepatocyte growth factor and fibronectin in anchorage independent survival of mammary carcinoma cells: requirement for phosphatidylinositol 3-kinase activity. AB - Anchorage-independent survival and growth are critical characteristics of malignant cells. We showed previously that the addition of exogenous hepatocyte growth factor (HGF) and the presence of fibronectin fibrils stimulate anchorage independent colony growth of a murine mammary carcinoma, SP1, which expresses both HGF and HGF receptor (Met; R. Saulnier et al., Exp. Cell Res., 222: 360-369, 1996). We now show that tyrosine phosphorylation of Met in carcinoma cells is augmented by cell adhesion and spreading on fibronectin substratum. In contrast, detached serum-starved cells exhibit reduced tyrosine phosphorylation of Met and undergo apoptotic cell death within 18-24 h. Under these conditions, the addition of HGF stimulates tyrosine phosphorylation of Met and restores survival of carcinoma cells. Soluble fibronectin also stimulates cell survival and shows a cooperative survival response with HGF but does not affect tyrosine phosphorylation of Met; these results indicate that fibronectin acts via a pathway independent of Met in detached cells. We demonstrated previously that inhibition of phosphatidylinositol (PI) 3-kinase activity blocks HGF-induced DNA synthesis of carcinoma cells (N. Rahimi et al., J. Biol. Chem., 271: 24850-24855, 1996). We now show in detached cells a cooperative effect of HGF and FN in the activation of PI 3-kinase and on the phosphorylation of PKB/Akt at serine 473. PI 3-kinase activity is also required for the HGF- and fibronectin-induced survival responses, as well as anchorage-independent colony growth. However, c-Src kinase or MEK1/2 activities are not required for the cell survival effect. Together, these results demonstrate that the PI 3-kinase/Akt pathway is a key effector of the HGF- and fibronectin-induced survival response of breast carcinoma cells under detached conditions and corroborate an interaction between integrin and HGF/ Met signalling pathways in the development of invasive breast cancer. PMID- 10714769 TI - Meeting report: Second European Phycological Congress, Montecatini Terme, Italy, September 20-26, 1999. PMID- 10714770 TI - Protist diversity: estimates of the near-imponderable. PMID- 10714771 TI - Uniform distribution of transcription complexes over the entire Leishmania donovani clpB (hsp 100) gene locus. AB - We have analyzed the RNA polymerase density on the Leishmania donovani clpB gene locus. Our results show an even distribution of RNA polymerase over the clpB locus indicating an undiscriminative transcription. We conclude that, unlike the hsp70 genes, the clpB gene is not transcribed individually, but rather as part of a larger, polycistronic transcription unit. PMID- 10714772 TI - The effect of electrostatic charge of food particles on capture efficiency by Oxyrrhis marina Dujardin (dinoflagellate). AB - Laboratory experiments were carried out to investigate the effect of food quality, measured as surface charge of the particles, on capture efficiency and ingestion rate by the heterotrophic dinoflagellate Oxyrrhis marina. Fluorescent particles in two size classes of around 1 and 4 microm and of 7 different qualities were offered to the flagellate: carbohydrate and albumin particles, the algae Synechocystis spec. and Chlorella spec., carboxylated microspheres, silicate particles and bacteria. Rates of particle uptake showed significant differences depending on particle size and quality, and ranged from 0 to 4 particles cell(-1) h(-1). Ingestion rates were up to 4 times higher for 4 pm particles than for 1 microm particles, which indicates strong size-selective feeding. Our main result is that the surface charge or zeta potential, of artificial particles, i.e. carboxylated microspheres (> or = -107 mV) and silicate particles, strongly differ from more natural and natural food (< or = 17 mV). For both size classes Oxyrrhis had ingestion rates up to 4 times higher for particles with less negative charge, such as albumin particles or algae. Thus, the zeta potential of the model food should be considered in experimental design. Particles with a zeta potential similar to that of natural food, e.g. albumin, seem to be the preferred model food. PMID- 10714774 TI - Mesostigmatophyceae, a new class of streptophyte green algae revealed by SSU rRNA sequence comparisons. AB - Complete nuclear-encoded SSU rRNA sequences have been obtained from three taxa of streptophyte green algae (Klebsormidium nitens, Nitella capillaris, Chaetosphaeridium globosum) and two strains of the scaly green flagellate Mesostigma viride. Phylogenetic analyses of 70 taxa of Viridiplantae (Chlorophyta and Streptophyta) and 57 taxa of streptophyte green algae and embryophyte plants using distance, parsimony and likelihood methods revealed a novel monophyletic lineage among the Streptophyta comprising the genera Mesostigma and Chaetosphaeridium. This lineage is described here as the Mesostigmatophyceae classis nova. Our analyses demonstrate that (1) scaly green flagellates (prasinophytes) are polyphyletic, (2) a scaly green flagellate is a member of the Streptophyta and forms a clade with the oogamous, filamentous Chaetosphaeridium to the exclusion of all other known streptophyte green algae, (3) a previously published SSU rRNA sequence of Chaetosphaeridium (AF113506) is chimeric and contains part of a fungal SSU rRNA, and (4) the phylogenetic relationships between the Mesostigmatophyceae and other streptophyte green algae remain unresolved by SSU rRNA sequence comparisons. PMID- 10714773 TI - Symbiomonas scintillans gen. et sp. nov. and Picophagus flagellatus gen. et sp. nov. (Heterokonta): two new heterotrophic flagellates of picoplanktonic size. AB - Two new oceanic free-living heterotrophic Heterokonta species with picoplanktonic size (< 2 microm) are described. Symbiomonas scintillans Guillou et Chretiennot Dinet gen. et sp. nov. was isolated from samples collected both in the equatorial Pacific Ocean and the Mediterranean Sea. This new species possesses ultrastructural features of the bicosoecids, such as the absence of a helix in the flagellar transitional region (found in Cafeteria roenbergensis and in a few bicosoecids), and a flagellar root system very similar to that of C. roenbergensis, Acronema sippewissettensis, and Bicosoeca maris. This new species is characterized by a single flagellum with mastigonemes, the presence of endosymbiotic bacteria located close to the nucleus, the absence of a lorica and a R3 root composed of a 6+3+x microtubular structure. Phylogenetical analyses of nuclear-encoded SSU rDNA gene sequences indicate that this species is close to the bicosoecids C. roenbergensis and Siluania monomastiga. Picophagus flagellatus Guillou et Chretiennot-Dinet gen. et sp. nov. was collected in the equatorial Pacific Ocean. Cells are naked and possess two flagella. This species is characterized by the lack of a transitional helix and lateral filaments on the flagellar tubular hairs, the absence of siliceous scales, two unequal flagella, R1 + R3 roots, and the absence of a rhizoplast. SSU rDNA analyses place this strain at the base of the Chrysophyceae/Synurophyceae lineages. PMID- 10714775 TI - Apparent global ubiquity of species in the protist genus Paraphysomonas. AB - Evidence is presented for the ubiquity of protist species. Using the example of protists that leave traces (siliceous scales) of their recent population growth, we show that most - perhaps all species in the genus Paraphysomonas, are ubiquitous. Of the species recorded in surveys carried out worldwide, we have identified 78% of their number in 0.1 cm2 of sediment collected from a freshwater pond (total area 10(8) cm2) in England. Moreover, the pond appears to act like a microcosm of aquatic environments in general, for species that are globally rare or abundant, are likewise rare or abundant in the pond. We assume that the rate of neutral migration to the pond is greatest for the globally abundant species. As these species are probably capable of growth in a broad range of conditions, they will more frequently encounter the environment they require for population growth. Thus globally abundant species are also locally abundant in the pond - a pattern that will be amplified by periodic cyst production. Ubiquitous dispersal is probably driven by very high absolute abundance of individuals, and the water column of the pond was estimated to support >10(14) Paraphysomonas individuals. Ubiquity will dampen rates of speciation, and the evidence presented here indicates that global species richness of Paraphysomonas is indeed modest - perhaps close to what is already known. PMID- 10714776 TI - Per aspera ad protozoa et astra, Yuri I. Polyansky and Russian protistology. PMID- 10714777 TI - High and low pressure pulsatile lavage of contaminated tibial fractures: an in vitro study of bacterial adherence and bone damage. AB - OBJECTIVE: This study was designed to examine the effect of pulsatile irrigation on microscopic bone architecture and its time-dependent efficacy in removing adherent slime-producing bacteria from cortical bone. DESIGN: Using an in vitro model, ten-millimeter transverse cut sections from five human tibiae were contaminated with Staphylococcus aureus and subjected to either high pressure pulsatile lavage (HPPL; seventy pounds per square inch, normal saline) or low pressure pulsatile lavage (LPPL; fourteen pounds per square inch, normal saline) or served as controls. Alteration of bony architecture was quantified by using a previously described ordinal scale and histomorphometric analysis of each transverse cut section of tibia. To assess the time-dependent effectiveness of pulsatile lavage in removing adherent bacteria from bone, ten-millimeter transverse cut sections from ten canine tibiae were contaminated with S. aureus and subjected to high or low pressure pulsatile lavage immediately or after one, three, or six hours. Scanning electron microscopy and bacterial cultures were used to assess the removal of adherent bacteria. RESULTS: HPPL resulted in significantly greater macroscopic damage than was seen with LPPL or in controls (ANOVA, p < 0.001). Histomorphometry revealed that HPPL was associated with significantly larger and more numerous fissures or defects in the cortical bone when compared with low pressure irrigation (p < 0.001). However, high and low pressure lavage were associated with similar degrees of periosteal separation from the cortical bone surface (p = 0.87). Both high and low pressure lavage were effective in removing adherent bacteria from bone at three hours irrigation delay, but only high pressure lavage removed adherent bacteria from bone at six hours delay. CONCLUSION: In this in vitro study, compared with HPPL, LPPL led to less structural damage and was equally effective in removing bacteria within three hours debridement delay; however, the efficacy of LPPL at six hours debridement delay is questionable. This finding may have clinical significance in the development of infection following open tibial fractures. PMID- 10714778 TI - Mechanical comparison of plates used in the treatment of unstable subtrochanteric femur fractures. AB - OBJECTIVES: To determine the stiffness and strength characteristics of certain plate-composite femur models designed to simulate unstable subtrochanteric femur fractures (OTA 31-A2.3). DESIGN: Fifteen identical composite femora were osteotomized to produce like models of an unstable subtrochanteric femur fracture. The femora were fixed with either the Synthes 95 degree angled condylar blade plate, a 95 degree dynamic condylar screw plate (DCS), or a 135 degree dynamic compression hip screw (DHS). MAIN OUTCOME MEASUREMENTS: A materials testing machine was used to apply compression to the femoral head through an adapter plate. Stiffness values were calculated from the load-deformation curves obtained. RESULTS: The DHS-femur model was the stiffest (586 newtons/ millimeter), followed by the 95 degree DCS (404 newtons/millimeter) and the 95 degree condylar blade plate (260 newtons/ millimeter). The DHS also had the highest ultimate load-to-failure (4,877 newtons), followed by the 95 degree DCS (3,107 newtons) and the 95 degree condylar blade plate (2,272 newtons). All of these differences were statistically significant (p < 0.00001 ). CONCLUSIONS: Our findings suggest that the Synthes 95 degree DCS has greater stiffness and strength than the Synthes 95 degree condylar blade plate when tested in this model of an unstable subtrochanteric femur fracture. This model may not be completely appropriate for testing the 135 degree DHS because the hard plastic "cortex" of the model prevented cut-out of the screw. PMID- 10714779 TI - Biomechanical analysis of supracondylar femoral fractures fixed with modern retrograde intramedullary nails. AB - OBJECTIVES: Several new retrograde supracondylar intramedullary nails have been developed to specifically address fractures of the distal femur. The nails appear clinically effective, but there are few biomechanical data documenting the stability of the fixation or the mechanical stiffness of the different designs. The goal of this study was to assess the torsional and bending stiffness of four designs of intramedullary nails developed for this application. METHODS: Four nail designs were tested in torsion and bending to determine system stiffness: Ace supracondylar, Richards "five hole" and "multi-hole" supracondylar, and Biomet retrograde. The nails were inserted into cadaveric femurs in which a one centimeter distraction osteotomy had been created seven centimeters proximal to the condyles. The constructs were then tested on an Instron biaxial testing system. RESULTS: There were no statistically significant differences in bending stiffness among the groups of nails (range 0.79 to 1.18 newtons/meter; p > 0.1). However, the Ace nails (1.10 newtonmeters/degree) did exhibit a statistically lower torsional stiffness compared with the other nails (2.20 to 2.21 newton meters/ degree; p < 0. 1). No differences were noted as a function of the number of locking holes. CONCLUSIONS: The bending stiffness of four currently available designs of retrograde intramedullary nails does not appear to be dependent on design variations. The torsional stiffness did vary among the four designs, but this was not determined by the number of fixation holes provided. It appears that a well-placed retrograde supracondylar nail of modern design should have sufficient stiffness to support the femur and provide stability during fracture healing. PMID- 10714780 TI - A biomechanical analysis of internal fixation of complex tibial plateau fractures. AB - OBJECTIVE: To compare the mechanical stability of fixation of an unstable bicondylar tibial plateau fracture with several different fixation techniques in a cadaveric model. DESIGN: Randomized laboratory investigation using a simulated bicondylar tibial plateau fracture with metaphyseal-diaphyseal dissociation. SETTING: Complex tibial plateau fractures were instrumented and tested under ramp and cyclic loading conditions on a servohydraulic materials testing machine. INTERVENTION: Each tibia was instrumented sequentially with a lateral buttress plate, a lateral and a medial buttress plate, and a lateral buttress and an anteromedial antiglide plate for ramp load testing. For cyclic testing, one of the three constructs was used on each specimen. MAIN OUTCOME MEASUREMENTS: Vertical subsidence of the medial tibial plateau was measured in both ramp and cyclic loading in order to evaluate the three internal fixation techniques. RESULTS: No significant difference was measurable between the dual buttress construct and the lateral buttress/anteromedial antiglide construct. However, the lateral buttress plate alone provided significantly less stability. CONCLUSIONS: A lateral buttress plate with an anteromedial antiglide plate may provide equally effective fixation as compared with the dual buttress plating technique in complex tibial plateau fractures. This less invasive technique may also be associated with fewer complications due to the lack of soft tissue stripping that is required for its application. PMID- 10714781 TI - The mechanical effect of blocking screws ("Poller screws") in stabilizing tibia fractures with short proximal or distal fragments after insertion of small diameter intramedullary nails. AB - OBJECTIVES/HYPOTHESIS: To evaluate the mechanical effects of medial and lateral blocking screws in supplementing intramedullary nail fixation of high proximal and low distal tibial fractures treated with small-diameter intramedullary nails. STUDY DESIGN: Intact fresh human cadaveric tibiae were sectioned to provide ten distal segments measuring seventy millimeters and ten proximal segments measuring ninety millimeters. In the distal segments, stainless steel solid eight millimeter tibial nails were advanced to eight millimeters from the ankle joint. Two transverse and one anterior-posterior (AP) locking screw were inserted using a custom-made jig. The same jig was used for the placement of a medial and a lateral blocking screw (BS) in the AP direction, nine millimeters above the superior most interlocking screw and eight millimeters distal to the lower end of the segment. In the proximal segments, two interlocking screws (both static and dynamic screws) were placed in a medial-lateral direction with the use of the insertion handle. A jig was used for placement of a medial and a lateral BS in the AP direction, nine millimeters below the lower transverse interlocking screw and sixteen millimeters proximal to the lower end of the segment. The bone implant construct (BIC) was embedded and fixed in a materials testing machine. The BICs were loaded in the medial-lateral direction at a distance of 185 millimeters from the nail ends with loads from -150 newtons to + 150 newtons. Force-displacement curves were recorded before and after insertion of the BSs. RESULTS: In proximal BICs, the addition of BSs decreased the deformation of the BICs 25 percent, from 8.9 +/- 1.9 degrees [mean +/- standard deviation (SD)] in the control group to 6.8 +/-1.1 degrees in the BS group (mean +/- SD) (p < 0.0001). In distal BICs, the addition of BSs decreased the deformation of the BICs 57 percent, from 9.5 +/- 1.4 degrees (mean +/- SD) in the control group to 4.0 +/- 1.0 degrees in the BS group (mean +/- SD) (p < 0.0001). CONCLUSIONS: The study suggests that medial and lateral blocking screws can increase the primary stability of distal and proximal metaphyseal fractures after nailing and can be an effective tool for selected cases that exhibit malalignment and/or instability. PMID- 10714783 TI - Plate fixation of fractures of the distal aspect of the radius: relative indications. PMID- 10714782 TI - Bohler's angle: correlation with outcome in displaced intra-articular calcaneal fractures. AB - OBJECTIVES: Bohler's tuber joint angle is commonly assessed when evaluating calcaneal fractures. A severe heel fracture will result in a significant decrease or loss of this angle. The purpose of this study was to evaluate the correlation between Bohler's angle and functional outcome in displaced intra-articular calcaneal fractures. DESIGN: Prospective cohort study. SETTING: Level I trauma center. METHODS: Ninety-five fractures in eighty-eight patients were analyzed for clinical outcome as measured by previously validated visual analogue scale (VAS) and SF-36 Health Survey scores. Radiographic results were measurements of Bohler's angle on plain x-rays. Angles were measured twice by two observers working independently. RESULTS: This prospective randomized cohort study indicates that patients initially presenting with a severely depressed Bohler's angle have a poor two-year outcome regardless of treatment. As well, fractures of lesser initial displacement, as measured by Bohler's angle, had higher functional scores on both VAS and SF-36 scoring scales. CONCLUSION: In this study, Bohler's angle had significant prognostic value in terms of predicting morbidity. Fractures with a markedly diminished Bohler's angle demonstrated a much poorer two-year outcome regardless of treatment. It would seem that the initial Bohler's angle was highly prognostic, regardless of treatment modality. PMID- 10714784 TI - External fixation, not ORIF, as the treatment of choice for fractures of the distal radius. PMID- 10714785 TI - Open reduction and internal fixation of pilon fractures. PMID- 10714786 TI - External fixation is the treatment of choice for fractures of the tibial plafond. PMID- 10714787 TI - Posterolateral approach for tibial pilon fractures: a report of two cases. AB - Open reduction and internal fixation (ORIF) of displaced tibial pilon fractures can lead to a high percentage of good and excellent functional results, but has also been associated with a meaningful incidence of wound breakdown and infection. The use of the posterolateral approach to the distal tibia for ORIF of tibial pilon fractures is presented. This may be used instead of the standard anteromedial incision in certain fracture configurations. The flexor hallucis longus muscle coverage overlying the plate fixation of the tibia and ability to fix both the tibia and fibula through the same incision may decrease the risk of deep infection and wound complications in these injuries frequently associated with marked soft tissue trauma. PMID- 10714788 TI - Immediate tibiocalcaneal arthrodesis with interposition fibular autograft for salvage after talus fracture: a case report. AB - Treatment goals in the operative management of talus fractures include prompt, anatomic, open reduction with rigid internal fixation; functional outcome is measured by degree of arthrosis, pain, range of motion, limb length, cosmesis, and return to premorbid activities. If restoration of the articular surfaces is precluded secondary to comminution, immediate and/or staged reconstructive salvage procedures must be considered. This report describes an immediate reconstructive procedure for salvage after a comminuted talus fracture with an ipsilateral tibia fracture. A standard antegrade tibial nail extending into the calcaneus was selected to stabilize both fracture sites. The technique of tibiocalcaneal arthrodesis using interposition fibular autograft and intramedullary fixation is presented as a unique treatment option. PMID- 10714789 TI - Evidence-based medicine: empiric antibiotic therapy in community-acquired pneumonia. AB - A number of national society guidelines exist for empiric management of community acquired pneumonia but these are, to a large extent, not evidence-based, but based on clinical experience, in vitro data, pragmatism and common sense. Many randomized controlled trials of antibiotic therapy in community-acquired pneumonia have been conducted, but most of these have been powered to demonstrate equivalent efficacy of new treatments in comparison with conventional antimicrobial therapy. Development of new antibiotics has been driven by the emergence of penicillin-resistant Streptococcus pneumoniae, but so far there is no hard evidence that beta-lactam therapy fails in community-acquired pneumonia, at least with the higher doses of penicillins that are commonly used in hospital practice. Nonetheless, newer antibiotics have been deployed including macrolides and quinolones, and have demonstrated equivalent (and in some cases, marginally improved) efficacy to older antibiotic treatments in randomized control trials. A number of studies have shown that it is possible to stratify patients according to severity of illness, to in-patient or out-patient management protocols. These have been validated and refined. PMID- 10714790 TI - Immunotherapy with Mycobacterium vaccae in the treatment of mycobacterial disease. PMID- 10714791 TI - What is the future of exchange transfusion for falciparum malaria? PMID- 10714792 TI - What is the future of exchange transfusion in severe malaria? PMID- 10714793 TI - Cytomegalovirus viraemia has poor predictive value for the development of cytomegalovirus disease in patients with advanced HIV-infection. AB - OBJECTIVE: Cytomegalovirus (CMV) continues to be one of the most important opportunistic infections associated with human immunodeficiency virus (HIV) infection. This study investigated the value of CMV-viraemia in predicting the development of clinical CMV disease in patients with advanced HIV infection. METHODS: This was a prospective observational study performed over a 2-year period between 1994-96 in the Department of Infection and Tropical Medicine at Leicester Royal Infirmary. Adult HIV-positive patients attending a hospital clinic were included if they were CMV-seropositive with CD4 counts < or =50 cells/mm3. Subjects were seen at approximately 6-weekly intervals in the clinic and were reviewed by an experienced ophthalmologist. Serum for CMV PCR was taken and stored at regular intervals and qualitative and quantitative PCR was performed at the end of the study period. The value of PCR in predicting the development of CMV disease was then assessed. RESULTS: Twenty-six patients were followed up during the study period and 77 evaluable specimens were analysed for CMV PCR. Twenty-three (30%) samples were positive and 54 negative. Seven (27%) patients developed CMV disease (five retinitis alone, and two with retinitis and oesophagitis) during the study period. Viraemia was often intermittent and there was no significant difference in the proportions of patients with positive or negative tests who subsequently developed CMV disease. The sensitivity, specificity, positive and negative predictive values of the qualitative PCR were 71%, 47%, 33% and 82% respectively and 57%, 74%, 44% and 82% respectively for the quantitative PCR (>10(3) copies/ml). CONCLUSIONS: The results from this study, which was performed before the introduction of protease inhibitors, found that cytomegalovirus PCR was of limited clinical value in predicting the patients at greatest risk of developing CMV-disease and provided little useful prognostic information. PMID- 10714794 TI - Recovery of long-term natural protection against reactivation of CMV retinitis in AIDS patients responding to highly active antiretroviral therapy. AB - OBJECTIVES: To see whether in severely immunosuppressed AIDS patients (with prior Cytomegalovirus retinal disease) who have significant increases in CD4+ lymphocytes following the initiation of highly active antiretroviral therapy (HAART) anti-Cytomegalovirus (CMV) maintenance therapy can be withdrawn with no subsequent progression of CMV retinitis. METHODS: Eight patients with AIDS and one or more previous episodes of CMV retinitis interrupted anti-CMV maintenance therapy following the successful beginning of HAART. CD4 cell counts and HIV-RNA were monitored monthly while measurement of CMV antigenemia and ophthalmoscopy were carried every 2 weeks thereafter. RESULTS: The HAART recipients in whom anti CMV maintenance therapy had been interrupted had measureable increases of CD4+ T lymphocytes, substantial control of both HIV-RNA and CMV viraemia and did not show recurrence of retinitis during a mean follow-up of 98.4 weeks (range 78-120, SD 15.2). CONCLUSIONS: Anti-CMV maintenance therapy can be interrupted with no subsequent progression of retinal damage over a long time in patients with AIDS who successfully respond to HAART with a significant increase in CD4 cell count. PMID- 10714795 TI - Clinical impact of rapid oxacillin susceptibility testing using a PCR assay in Staphylococcus aureus bactaeremia. AB - OBJECTIVES: The aim of this work was to establish the clinical impact of rapid oxacillin susceptibility testing in nosocomial Staphylococcus aureus bacteraemia. METHODS: This study was performed in 145 critically ill patients infected by S. aureus. Patients were randomly assigned to one of two groups: patients for whom susceptibility testing was performed using a rapid same day multiplex PCR assay for detection of the staphylococcal mecA (mean delay of response: 6 h) and those for whom testing was accomplished using traditional overnight techniques (21 h). RESULTS: The results of this study showed no significant difference between the two groups in terms of age, Simplified Acute Physiologic Score, severity of infection, severity of underlying disease and clinical outcome (control vs. PCR): unfavourable outcome of infection, 12.32 vs. 12.5%; 95% CI for the difference = 11.49 to 11.09 (P = 0.975); unfavourable general outcome, 16.43 vs. 20.83%; 95% CI for the difference = -17.35 to 8.50 (P = 0.497). For the oxacillin-susceptible S. aureus bactaeraemia, results were: unfavourable outcome of infection = 13.04 vs. 11.11%; 95% CI for the difference = -11.38 to 16.18 (P = 0.767); unfavourable general outcome = 13.04 vs. 20.37%; 95% CI for the difference = -22.12 to 8.07 (P = 0.331). CONCLUSION: This study seemed to demonstrate that rapid oxacillin susceptibility testing using a PCR assay did not have a major impact on the care and outcome of patients with S. aureus bactaeremia. PMID- 10714796 TI - Length of time to laboratory diagnosis of Mycobacterium tuberculosis infection: comparison of in-house methods with reference laboratory results. AB - OBJECTIVES: To audit the time taken to obtain laboratory confirmation of infection with Mycobacterium tuberculosis using in-house methods of polymerase chain reaction (PCR) and culture and referral to a reference laboratory. METHODS: Retrospective collection of data from laboratory records covering a period of 1 year. RESULTS: Median time to microbiological diagnosis of a new infection using the in-house services in addition to the reference laboratory was 22.0 days. Using reference laboratory results alone, median time to diagnosis would have been 61.5 days. CONCLUSIONS: Development of on-site laboratory facilities to identify Mycobacterium tuberculosis can reduce the time to its identification by almost two-thirds. PMID- 10714797 TI - Is VZV reactivation a common cause of unexplained unilateral pain? Results of a prospective study of 57 patients. AB - OBJECTIVE: Pain is a common reason for patients to present to a doctor. Many patients with zoster have seen their doctor with pain during the days before the rash and zoster sine herpete is well described. If early varicella zoster virus (VZV) reactivation could be identified confidently, it could provide an opportunity for early antiviral intervention. This prospective study was performed to assess how often patients presenting to their general practitioner with unilateral pain of no obvious clinical cause proved to have evidence of VZV reactivation. METHODS: Fifty-seven patients were recruited and followed for 28 days; laboratory testing included VZV polymerase chain reaction (PCR) from peripheral blood mononuclear cells, VZV IgG, IgA and IgM. The control group consisted of 81 blood donors. RESULTS: Only two study patients developed the rash of zoster. There was no significant difference in PCR or serological responses between the study group and control group. Clinical characteristics did not enable identification of patients presenting to their doctor with unilateral pain who had prodromal zoster. CONCLUSION: There was no evidence on clinical or laboratory tests used in this study to support the view that reactivation of VZV is a common cause of unexplained unilateral pain. PMID- 10714798 TI - Maternal humoral factors associated with perinatal human immunodeficiency virus type-1 transmission in a cohort from Kigali, Rwanda, 1988-1994. AB - OBJECTIVES: to study different parameters of humoral immunity responses in the serum of 39 human immunodeficiency virus type-1 infected pregnant women from Kigali, (Rwanda) in correlation with perinatal transmission. METHODS: this study was done between 1988 and 1994. Thirty nine HIV-1 infected women, 18 transmitting (T) and 21 non-transmitting (NT) mothers, have been chosen based on the quantity of sera available for analysis. Maternal data were collected at the time of delivery or during the preceding month. Quantification of viral load was performed by the signal amplification bDNA assay. Specific reactivity of antibody was tested against recombinant p24 protein and five different synthetic peptides from gp120 and gp41 based on HIV LAI-strain sequences. Neutralization assays were performed against laboratory (RII strain of the HIV-1 C subtype) and primary strains (two NSI and one SI of the HIV-1 A subtype). Antibody Dependent Cellular Cytotoxicity assay was performed with CEM.NK(R) cells against a laboratory HIV-1 strain. RESULTS: absence of correlation regarding maternal viral load, or viral subtype and vertical transmission was observed. By contrast, the CD4/CD8 ratio was significantly higher in non-transmitting mothers compared to transmitting mothers. Moreover, high anti-p24 antibody avidity was correlated with a lower risk of perinatal transmission. Furthermore, transmission risk appeared significantly higher with reactivity of serum samples to linear epitopes of gp41 (amino acids 566-582, 578-594), whereas risk appeared lower with reactivity to the immunodominant domain of gp41 (amino acids 597-609). No significant difference was observed in titres of antibody neutralizing primary isolates (two NSI (non syncitium inducer) and one SI (syncitium inducer) of the HIV-1 A subtype) and laboratory strain (RII strain, of the HIV-1 C subtype) between transmitting and non-transmitting mother's sera. In addition, titres of Antibody Dependent Cellular Cytotoxicity were similar in transmitting versus non transmitting mothers. However, high Antibody Dependent Cellular Cytotoxicity titres were correlated with a good clinical status of children. CONCLUSIONS: three parameters such as high CD4/CD8 ratio, high anti-p24 antibody avidity and high reactivity against the immunodominant epitope of gp41 have been shown to be correlated with no perinatal transmission. High Antibody Dependent Cellular Cytotoxicity titres appeared to be linked to a good clinical status of children after birth. One parameter, reactivity against two linear epitopes of gp41, appeared to be correlated with vertical transmission. PMID- 10714799 TI - Influenza diagnosis: from dark isolation into the molecular light. West of Scotland Respiratory Virus Study Group. AB - OBJECTIVES: To compare the conventional virus isolation method for diagnosis of influenza infection with reverse-transcription polymerase chain reaction (RT-PCR) in prospectively collected nose and throat swabs from elderly patients during the winter influenza season. The use of a denaturing buffer as an alternative to viral transport medium (VTM) for submission of combined nose and throat swabs to the laboratory for PCR was then investigated in a second study. METHODS: Virus was cultured in microtitre plates using two different cell lines and detected using monoclonal antibody staining. A multiplex, matrix gene PCR assay was optimized to increase the sensitivity and specificity of detection of influenza A (H3 and H1) and B nucleic acid. RESULTS: The multiplex assay detected all viruses with equal sensitivity to individual assays. In a large, multicentre field study PCR detected twice as many influenza infections compared with virus isolation. No positive culture was missed. PCR has a rapid turn around time (< 36 h) vs. a minimum of 7 days for virus isolation. Greater sensitivity and specificity in the PCR were achieved using a 'hot-start' method. Although the numbers were small, the detection rate using PCR was greater for swabs submitted in denaturing buffer than in VTM. CONCLUSIONS: PCR significantly increased the sensitivity and clinical utility of influenza A (H3 and H1) and B diagnosis. There were a number of advantages in using denaturing buffer for submission of samples, including high sensitivity, rapidity, ease of use and no requirement for the virus to be viable on arrival at the laboratory. Therefore, PCR is a rapid, sensitive and user-friendly alternative for influenza diagnosis. Virus isolation technology should be limited to referral centres for further epidemiological characterization. PMID- 10714800 TI - Concomitant poliovirus infection during an outbreak of hepatitis A. AB - AIM OF THE STUDY: The present study was designed to evaluate the possible co infection, with other enteric viruses, during an outbreak of hepatitis A (HA). MATERIAL AND METHODS: Forty-two stool samples and sera were collected during an outbreak of hepatitis A. Sera were analysed by the Abbott test for IgG-IgM anti HAV antibodies. Stool samples were used to identify the presence of enteric viruses. HAV genome was identified by a RT-PCR test, other enteric viruses were identified, after cell passage and seroneutralization test on BGM cells, by RT PCR and RFLP assay. RESULTS: The samples were obtained from 27 employees of an industrial plant, nine household contacts and six non-employee controls. The attack rate was 12.5%, whereas the overall prevalence was 63%. In the employee group, 12 out of 27 stool samples were positive for the presence of HAV by reverse transcriptase polymerase chair reaction (RT-PCR). All the other samples (30) were negative. Five samples from employees, three from household contacts and one from non-employees were also found positive for enteroviruses. These viruses were classified by seroneutralization as poliovirus and RFLP assay as Sabin poliovirus type 1. Four samples were positive both for HAV and poliovirus. CONCLUSIONS: This study confirms co-infection with different enteric viruses may occur and also emphasizes the wide circulation of HAV and the existence of silent infection with poliovirus. PMID- 10714801 TI - Automated exchange transfusion for life-threatening plasmodium falciparum malaria -lessons relating to prophylaxis and treatment. AB - We report a case of traveller to Kenya who contracted severe plasmodium falciparum malaria complicated by disseminated intravascular coagulation and acute renal failure. She had taken no antimalarial prophylaxis in view of concerns in the media regarding the adverse effects of mefloquine. There was a protracted delay before the diagnosis of malaria was made. Clinical recovery occurred following treatment with intravenous quinine, haemofiltration and manual/automated red-cell exchange transfusions. Automated red-cell exchange transfusion resulted in a marked decrease in the parasitaemia, before a response to quinine therapy would have been anticipated, leading to a successful outcome thereafter. In conjunction with other groups we therefore feel that exchange transfusions should be considered in seriously ill patients with falciparum malaria, multiorgan complications and parasitaemias greater than 10%. PMID- 10714802 TI - Automated erythrocytapheresis in the treatment of severe falciparum malaria. AB - Removal of parasitized erythrocytes is generally considered to be of value as adjunctive therapy in severe falciparum malaria with high parasitaemia. This is commonly achieved by exchange transfusion. We describe three cases of severe falciparum malaria treated by automated erythrocytapheresis (red cell exchange) in addition to quinine and conventional supportive therapy. Erythrocytapheresis consists of removal of the red-cell fraction by apheresis. Plasma, leukocyte and platelet fractions are returned to the patient. In all cases, dramatic reduction in parasitaemia was achieved within 2 h with subsequent complete clinical recovery. Erythrocytapheresis has significant advantages over exchange transfusion in terms of speed, efficiency, haemodynamic stability and retention of plasma components such as clotting factors and may thus represent an improvement in adjunctive therapy for severe malaria. PMID- 10714803 TI - Use of corticosteroids to suppress drug toxicity in complicated tuberculosis. AB - Four cases of tuberculosis complicated by allergic or toxic reactions to antibiotic treatment are presented. In each, it was considered essential to suppress the reactions in order to give effective chemotherapy. This was achieved by using prednisolone generally in a dose of 40 mg/day or less during the continuation phase of therapy. Reactions to essential treatment are an important indication for corticosteroid treatment in tuberculosis. PMID- 10714804 TI - Tuberculous tenosynovitis and carpal tunnel syndrome as a presentation of HIV disease. AB - We describe a patient who presented with carpal tunnel syndrome secondary to tuberculous tenosynovitis and who was subsequently shown to have HIV infection. Recognition of this atypical presentation of tuberculosis is important for early, effective treatment. PMID- 10714805 TI - Successful treatment by meropenem of Campylobacter jejuni meningitis in a chronic alcoholic following neurosurgery. AB - Meningitis caused by Campylobacter jejuni is rare, we describe a case following neurosurgery for intra-cranial haematoma in a chronic alcoholic patient. Conventional culture of CSF and blood was supplemented by polymerase chain reaction (PCR) detection of Campylobacter jejuni. PMID- 10714806 TI - Acute adrenal insufficiency precipitated by isolated involvement of the adrenal gland by tuberculosis. PMID- 10714807 TI - Empyema of the thorax due to Gemella haemolysans. PMID- 10714808 TI - Probiotics and life-threatening infection. PMID- 10714809 TI - Relative bradycardia in infectious diseases. PMID- 10714810 TI - Effects of growth hormone on male reproductive functions. PMID- 10714811 TI - A role for mitochondrial DNA and sperm survival. PMID- 10714812 TI - Cloning: a basic bioethical analysis. PMID- 10714813 TI - Commentary: forging a partnership between total quality management and the andrology laboratory. PMID- 10714814 TI - Measurement of protein C inhibitor in seminal plasma is useful for detecting agenesis of seminal vesicles or the vas deferens. AB - Protein C inhibitor (PCI), a plasma serine protease inhibitor of activated protein C, is present at high concentrations in the seminal plasma of normal subjects and is decreased in some infertile patients. We measured the concentrations of PCI, prostate-specific antigen, and fructose in the seminal plasma of infertile patients (n = 125) and of normal subjects (n = 13). We also measured time-dependent changes in the concentrations of PCI and fructose in seminal plasma after ejaculation. A weak correlation was found between the levels of PCI and fructose (r = 0.268, P = 0.016). The PCI level in seminal plasma of patients with seminal vesicle and/or vasal agenesis was significantly lower (P < .01) than in normal subjects. The level of fructose in seminal plasma decreased in vitro in a time-dependent manner after ejaculation, whereas the concentration of PCI was stable at 48 hours after ejaculation. These data suggest that PCI in seminal plasma, as well as fructose, may become one of the markers for agenesis of seminal vesicles and/or the vas deferens. PMID- 10714815 TI - Effects of accessory sex gland fluid on viability, capacitation, and the acrosome reaction of cauda epididymal bull spermatozoa. AB - The effect of accessory sex gland fluid (AGF) on viability and acrosomal integrity of spermatozoa was examined with cauda epididymal spermatozoa and AGF from the same Holstein bull (n = 6). Surgical cannulation of the vasa deferentia enabled the separate collection of cauda epididymal effluent and AGF from each bull. Cauda epididymal effluent was incubated with either AGF collected from the same bull or medium alone. Following coincubation, spermatozoa (5 x 10(7) sperm/mL) were incubated in medium alone or under capacitating conditions (10 microg/mL heparin) for 16 hours. Every 2 hours, an aliquot of spermatozoa was exposed to lysophosphatidylcholine (100 microg/mL) to induce the acrosome reaction in capacitated spermatozoa. Sperm motility decreased over time regardless of treatment. Overall, spermatozoa incubated in AGF had fewer acrosome intact live spermatozoa than did those not incubated in AGF. Viability was significantly (P < .05) compromised over time when spermatozoa were exposed to AGF, compared with those not preincubated in AGF. Significantly more (P < .05) acrosome-reacted live spermatozoa were seen following exposure to heparin and lysophosphatidylcholine when spermatozoa were not preincubated in AGF. We conclude that exposure of spermatozoa to AGF accelerates cell death and that rapid removal of spermatozoa from seminal plasma is critical for maximal viability. PMID- 10714816 TI - Dipeptidylpeptidase IV activities are elevated in prostate cancers and adjacent benign hyperplastic glands. AB - Dipeptidylpeptidase IV (DPP IV) is a serine exopeptidase that has been implicated in cell-extracellular matrix interactions and bioactive peptide/cytokine/growth factor metabolism. The objective of this study was to determine if DPP IV activities were changed with development of cancer in the prostate. DPP IV activity was measured in human prostate cancer and benign prostatic hyperplasia (BPH) tissues by biochemical assays with glycylprolyl-p-nitroanalide as substrate in tissue extracts (BPH, n = 8: cancer, n = 7; 2 with Gleason score 5 and 5 with Gleason score 7) and quantitative morphometry of histochemical activities with glycylproline-4-methoxy-beta-naphthylamide as substrate (BPH, n = 9: cancer, n = 13, 1 with Gleason score 4, 10 with Gleason score 6, 2 with Gleason score 8) in frozen-tissue sections. Data were analyzed by analysis of variance. The peptidase activity was detected in epithelial but not stromal cells of BPH and cancer tissues, and it was present as a single band of activity of approximately 160 kDa in electrophoretically separated activity blots of the extracts. DPP IV activity was increased approximately twofold in cancer versus BPH tissues as determined by biochemical and quantitative histochemical methods. In addition, DPP IV activity was increased to a similar extent in BPH glands associated with the cancers. These data indicate that DPP IV activity is increased not only in primary prostatic cancers but also in associated BPH glands, suggesting that there may be some local factors produced by cancer cells that influence adjacent BPH epithelial cells to positively affect the immediate growth environment of the cancer. PMID- 10714817 TI - Changes in the expression of junctional and nonjunctional complex component genes when inter-sertoli tight junctions are formed in vitro. AB - Throughout spermatogenesis, germ cells move progressively from the basal to the adluminal compartment, which is accompanied by continual disassembly and reassembly of intercellular junctions suggesting germ cell movement is composed of intermittent phases of junction disassembly and reassembly. A study was performed to correlate the expression of junctional-complex components (such as zonula occludens-1 [ZO-1], a tight-junction component protein) and nonjunctional complex components (such as urokinase-type plasminogen activator [uPA], a serine protease; cathepsin L, a cysteine protease; alpha2-macroglobulin, a nonspecific protease inhibitor; and cystatin C, a cysteine protease inhibitor) at the time when inter-Sertoli tight junctions were established in vitro. This is an attempt to investigate whether the expression of nonjunctional component genes also correlates with the formation of inter-Sertoli tight junctions in vitro. This is part of an effort to understand the physiologic elements of germ cell movement in the epithelium. Sertoli cells cultured in vitro are known to undergo programmed cell death. To ensure that the changes in target gene expression were not the result of apoptosis, Sertoli cells were cultured in vitro at densities of 0.25, 0.75, and 3 x 10(6) cells/cm2 for up to 7 days on bicameral culture units coated with Matrigel (Collaborative Research) and were assessed by morphologic analysis and agarose gel electrophoresis. It was noted that many of the Sertoli cells cultured at 3 x 10(6) cells/cm2 underwent apoptosis by day 7, in contrast to cultures at 0.25 and 0.75 x 10(6) cells/cm2 illustrating the Sertoli cell number per unit of area may be an important parameter to be considered when studying Sertoli cell function in vitro. Also, it was shown that the expression of ZO-1 increased significantly between days 2 and 3 prior to the establishment of inter Sertoli tight junctions assessed by transepithelial resistance measurement (TER), which illustrates that ZO-1 can be used as a marker to monitor this cellular event. More interestingly, there was also a transient increase in the expression of uPA and cathepsin L between days 2 and 3 at the time preceding the formation of tight junctions. In Sertoli cells cultured at low density (2 x 10(4) cells/cm2), when a confluent monolayer of cells could not form, there were no changes in the expression of either ZO-1, uPA, or cathepsin L throughout the 7 day culture period. These results show that the establishment of specialized junctions, such as tight junctions between Sertoli cells in vitro, may require the participation of both junctional and nonjunctional complex components. PMID- 10714819 TI - An immunochemical assay to detect DNA damage in bovine sperm. AB - An immunochemical assay has been developed to detect oxidative damage in bovine sperm DNA. Sperm DNA contains a large amount of oxidative damage as a result of exposure to exogenous agents, but damage also can caused by normal metabolic processes and the absence of DNA repair in the later stages of spermatogenesis. A freeze-thaw procedure performed on extended bovine sperm in straws did not induce additional DNA damage immediately after thawing compared with nonfrozen extended sperm. The data suggest that the amount of oxidative damage correlated to the percentage of artificially inseminated cows returning to service within 56 days postinsemination, because a number of sires with high sperm concentrations had a large variation in fertility after artificial insemination. These observations have led to the conclusion that by measuring DNA damage in thawed sperm, one might predict the fertility of bulls with high semen concentration. PMID- 10714818 TI - Measurement of intracellular calcium concentration and plasma membrane potential in human spermatozoa using flow cytometry. AB - We report 2 novel approaches using flow cytometry to measure intracellular calcium concentration and plasma membrane potential in human spermatozoa. Both approaches have the potential to measure different responses in subpopulations of cells, which is particularly useful when studying heterogeneous populations such as human spermatozoa. Intracellular calcium concentration ([Ca2+]i) was measured using the probe indo-1/AM. This allowed measurements to be made that were independent of variation in cell size and dye loading. It also enabled dead cells to be directly identified and excluded from the analyses without the need for counterstaining. Mean basal [Ca2+]i was determined as 50 nM (25-75 nM range) and, in response to the agonist progesterone (20 microM), this increased transiently to 195 nM (125-285 nM range) before declining to approximately half the maximal level within 2 minutes (values in parentheses correspond to the range of values typically found within a sperm population from 1 sample). These results are comparable with previously published data on whole sperm populations. Sperm membrane potential (VM) was assayed using the probe DiOC6(3). In carefully controlled experiments, a marked depolarization of the plasma membrane potential of capacitated spermatozoa was observed in response to progesterone (20 microM). Following in vitro capacitation, the sperm plasma membrane potential became hyperpolarized compared with the noncapacitated state. Therefore, this technique may be used to assay for sperm capacitation in vitro. PMID- 10714820 TI - Influence of sex hormones on prostate volume in men on hemodialysis. AB - The relationship between sex hormones and prostate volume in 90 men on hemodialysis and 91 healthy men was investigated. In men on hemodialysis, serum levels of total and free testosterone were significantly reduced compared with the controls (P < .001), whereas the serum estradiol level was significantly elevated in men on hemodialysis (P < .001). However, prostate volumes were not different between the 2 groups. Serum estradiol level correlated with the prostate volume in controls (P < .001), whereas total and free testosterone levels did not correlate with the prostate volume. In men on hemodialysis, serum levels of total and free testosterone and estradiol did not correlate with the prostate volume. The present study suggests that sex hormones do not play important roles in the prostate growth in men on hemodialysis. PMID- 10714821 TI - Factors involved in the biochemical etiology of human seminal plasma hyperviscosity. AB - Semen rheology was studied to elucidate the biochemical basis of seminal plasma hyperviscosity. Semen proved to fit in with a power law model, by presenting a pseudoplastic behavior. Apparent viscosity at 230 s(-1) and 25 degrees C (eta(a)) was 4.3 /- 0.2 cp and 5.4 +/- 0.4 cp in normal and high-consistency semen, respectively. The effect of enzymes and mucolytic agents on human seminal plasma viscosity were evaluated by incubating normal and hyperviscous semen pool aliquots with trypsin, dithiothreitol, EDTA, alpha-amylase and deoxyribonuclease I. After incubation, trypsin treatment reduced eta(a) by 36% in normal semen and by 44% in hyperviscous semen. There was a decrease in eta(a) following incubation of hyperviscous samples with dithiothreitol (33%) and alpha-amylase (44%) that was not observed in the normal consistency samples. No decrease was observed in eta(a) after EDTA or DNAse treatment of both groups. Comparison of normal and hyperviscous seminal plasmas revealed no difference in the concentration of total proteins, DNA, or in the percentage of water content. These findings indicate that the primary substances responsible for basic normal semen rheologic behavior are proteins. A comparison of rheological properties between normal and hyperviscous semen samples indicates the existence of a highly organized network in the latter group, in which disulfide bonds and oligosaccharide chains complexed to the peptide core may play a key role. PMID- 10714822 TI - The antigonadotropic action of testosterone but not 7alpha-methyl-19 nortestosterone is attenuated through the 5alpha-reductase pathway in the castrated male rat pituitary gland. AB - The enzyme 5alpha-reductase plays a significant role in the prostate to amplify the action of testosterone (T) by converting it to a more potent androgen, dihydrotestosterone (DHT). The role of 5alpha-reductase in the testosterone feedback inhibition of gonadotropin secretion from the pituitary has not been elucidated. Therefore, we investigated the role of 5alpha-reductase on T action in in vitro and in vivo models. Castration has been reported to increase the 5alpha-reductase activity in pituitary glands. Hence, the effect of castration duration on the conversion of T to DHT by pituitary homogenates and the responsiveness of pituitary monolayer cell cultures to gonadotropin-releasing hormone (GnRH) challenge exposure were investigated. Incubation of [3H]-T with pituitary homogenates showed that the conversion of T to 5alpha-reduced metabolites was two- to threefold greater in pituitaries from rats who had been castrated for 14 days compared with those castrated for 1 day. In addition, the GnRH-stimulated release of LH from monolayer cell cultures of pituitaries from rats castrated for 1 day was twofold greater, whereas that from rats castrated for 2 weeks was six- to sevenfold greater compared with basal luteinizing hormone (LH) release. Hence we used rats castrated for 2 weeks to elucidate the role of 5alpha-reductase in T feedback inhibition. The inhibitory effects of the androgens T, 19-nortestosterone (19-NT), and 7alpha-methyl-19-nortestosterone (MENT) at 3 different concentrations (10(-9), 10(-7), and 10(-5) mol/L) on GnRH stimulated LH release from monolayer cell cultures of pituitaries from rats castrated for 2 weeks were examined. All 3 androgens showed dose-dependent inhibition of LH release. MENT showed the greatest inhibition, followed by 19-NT and T. In the presence of finasteride (a 5alpha-reductase inhibitor), the inhibition of LH released by T and 19-NT were significantly greater. The inhibitory effect of MENT, which does not undergo 5alpha-reduction, was not altered by finasteride. In an in vivo study, rats castrated for 2 weeks received T with or without finasteride. There was a significantly greater suppression of serum LH in rats receiving T plus finasteride compared with those receiving T alone. These results suggested that 5alpha-reductase in the pituitary is not obligatory for the inhibitory action of T on gonadotropin secretion in the castrated rat. The action of MENT, a nonreducible androgen, on the pituitary is not affected by 5alpha-reductase. PMID- 10714823 TI - Postvasectomy alterations in protein synthesis and secretion in the rat caput epididymidis are not repaired after vasovasostomy. AB - Many men who have undergone vasectomy later request vasovasostomy. Unfortunately, significant numbers of these men remain infertile despite the reestablishment of patent ducts. This report examines the possibility that epididymal function remains compromised after vasovasostomy in the rat by examination of quantifiable, in vivo protein synthesis and secretion in the caput epididymidis. Rats were studied 30 days after vasectomy, 30 days after a vasovasostomy (which was performed 30 days after vasectomy), or after sham operations. Epididymal lumen fluids (LF) were collected by micropuncture after 3 hours' in vivo microperifusion of tubules with 35S-amino acids. Proteins were separated by 2 dimensional electrophoresis and were detected by Coomassie blue staining. Synthesized proteins in tubule extract and synthesized and secreted proteins in LF were detected by autoradiography and image analysis. Specific proteins that appeared to be affected by vasectomy-vasovasostomy were identified by internal sequence analysis. LF contained an average of 87 detectable proteins synthesized and secreted in the control caput. Nineteen of the most prominent LF proteins were selected for more focused study. The most prominent proteins were clusterin, cysteine-rich secretory protein (CRISP)-1, and epididymal retinoic acid-binding protein. Among these, CRISP-1 remained reduced in LF after vasovasostomy. Two more minor proteins that remained reduced after vasovasostomy were identified as prostaglandin D2 synthase and phosphatidylethanolamine-binding protein. All 3 of these proteins occur in the epididymides of multiple species and have been associated with sperm fertilizing capacity. PMID- 10714825 TI - Initial predominance of the oxidative activity of type I 11beta-hydroxysteroid dehydrogenase in primary rat Leydig cells and transfected cell lines. AB - Glucocorticoids suppress testosterone production in Leydig cells. The level of glucocorticoid action is set within the Leydig cell by the number of glucocorticoid receptors and by the activity of 11beta-hydroxysteroid dehydrogenase (11betaHSD). This enzyme acts either as an oxidase inactivating glucocorticoid or as a reductase amplifying its action. It is currently unknown whether extracellular conditions might cause 11betaHSD oxidative and reductive activities in Leydig cells to change inversely or independently. The aim of the present study was to determine whether extracellular conditions set in vitro by various culture time and media components, such as glucose and pyruvate, affect the relative rates of 11betaHSD oxidation and reduction. Primary rat Leydig cells and cell lines (COS1 and CHOP cells) transfected with 11betaHSD-I complementary DNA (cDNA), were incubated with 25 nmol/L (physiologic range) or 500 nmol/L (stress range) concentrations of radiolabeled substrates, corticosterone or 11 hydrocorticosterone, for 0 to 24 hours. Oxidative activity predominated over reductive activity under initial conditions when product formation increased linearly with time. For example, in Dulbecco's modified Eagle medium/F12 medium (containing 5.5 mmol/L glucose), the peak ratio of oxidation to reduction (with 1 denoting equivalence of oxidative and reductive activities) was 5.5 in rat Leydig cells, 19.7 in COS1 cells, and 20.8 in CHOP cells. Glucose stimulated reductive activity but did not change the predominant direction of 11betaHSD catalysis at earlier times. In COS1 cells transfected with 11betaHSD-I cDNA, oxidative activity rapidly increased during the first 2 hours of the incubation, then gradually decreased while reductive activity increased steadily. The relative ratio of oxidation to reduction rapidly declined to less than 0.5 at 6 hours, and thus the favored direction of catalysis changed from oxidation to reduction. However, in transfected CHOP cells, 11betaHSD oxidative activity rapidly increased during the first 2 hours and continued to increase for 24 hours. The ratio of oxidative to reductive activity rapidly declined but kept above 1 in CHOP cells for 24 hours, and the favored direction of catalysis remained predominantly oxidative. These results revealed that 11betaHSD-I is a predominant oxidase initially in Leydig cells and 2 cell lines, and that the oxidative activity is gradually lost over time. The data suggest that type I 11betaHSD is a predominant oxidase in Leydig cells in vivo. PMID- 10714826 TI - Intracavernosal pressure monitoring in mice: responses to electrical stimulation of the cavernous nerve and to intracavernosal drug administration. AB - With the development of transgenic mice to evaluate mechanisms of erectile function, it appears particularly advantageous to develop a standardized mouse model of penile erection. The purpose of the study reported here was to evaluate the novel application of intracavernosal pressure (ICP) monitoring in the mouse during electrophysiologic and pharmacologic induction of penile erection. In anesthetized adult male mice, the cavernous nerves (CN) were isolated unilaterally, and the corpora cavernosa were exposed. A 24-gauge angiocath (intravenous catheter) was inserted into the right corpus cavernosum to monitor the ICP, and a 30.5-gauge needle was inserted into the left corpus cavernosum for intracavernosal drug administration. ICP was recorded during CN-stimulated or pharmacostimulated erections. Electrical stimulation of the CN significantly increased the ICP (from 10.09 +/- 2.01 to 34.62 +/- 2.71 mm Hg, P < .05), which then returned to baseline pressure after termination of the electrical stimulation. Pretreatment with intracavernosal administration of the nitric oxide synthase inhibitor, nitro-L-arginine methyl ester (0.1 mg), inhibited the electrical stimulation-induced changes in ICP (7.17 +/- 1.46 vs 10.38 +/- 2.17 mm Hg, not significant [NS]). Also, intracavernosal administration of papaverine (0.4 mg) produced a significant increase in ICP (from 8.51 +/- 0.69 to 26.37 +/- 5.7 mm Hg, P < .05). We concluded that this technique might be applied to perform quantitative erection physiologic experiments with the mouse as an economical and experimentally advantageous animal model, particularly with the development of transgenic mice to evaluate mechanisms of erectile function. PMID- 10714824 TI - Identification and regulation of receptor tyrosine kinases Rse and Mer and their ligand Gas6 in testicular somatic cells. AB - Receptor tyrosine kinases act to convey extracellular signals to intracellular signaling pathways and ultimately control cell proliferation and differentiation. Rse, Axl, and Mer belong to a newly identified family of cell adhesion molecule related receptor tyrosine kinase. They bind the vitamin K-dependent protein growth arrest-specific gene 6 (Gas6), which is also structurally related to the anticoagulation factor Protein S. The aim of this study is to investigate the possible role of Rse/Axl/Mer tyrosine kinase receptors and their ligand in regulating testicular functions. Gene expression of Rse, Axl, Mer, and Gas6 in the testis was studied by reverse transcriptase-polymerase chain reaction (RT PCR) and Northern blot analysis. The results indicated that receptors Rse and Mer and the ligand Gas6 were expressed in the rat endothelial cell line (TR1), mouse Leydig cell line (TM3), rat peritubular myoid cell line (TRM), mouse Sertoli cell line (TM4), and primary rat Sertoli cells. Axl was not expressed in the testicular somatic cells by RT-PCR or Northern blot analysis. The highest level of expression of Gas6 messenger RNA (mRNA) was observed in the Sertoli cells, and its expression was responsive to the addition of forskolin in vitro. The effects of serum, insulin, and transferrin on Gas6 expression by TM4 cells were examined. It was shown that they all exhibited an up-regulating effect on Gas6 expression. The forskolin-stimulated Gas6 expression was accompanied by an increase in tyrosine phosphorylation of the Rse receptor in vitro, suggesting that Gas6 may exhibit an autocrine effect in the Sertoli cells through multiple tyrosine kinase receptors. Our studies so far have demonstrated that tyrosine kinase receptors Rse and Mer and their ligand Gas6 are widely expressed in the testicular somatic cell lines and may play a marked role in promoting testicular cell survival. PMID- 10714827 TI - Alterations in sperm characteristics of follicle-stimulating hormone (FSH) immunized men are similar to those of FSH-deprived infertile male bonnet monkeys. AB - The quality of sperm ejaculated by bonnet monkeys and normal, healthy proven fertile volunteer men, both actively immunized with ovine follicle-stimulating hormone (oFSH), was examined at different times of study for chromatin packaging and acrosomal glycoprotein concentration by flow cytometry. Susceptibility of sperm nuclear DNA to dithiothreitol (DTT)-induced decondensation, as measured by ethidium bromide binding, was markedly high compared with values at day 0 in men and monkeys during periods when FSH antibody titer was high. Sperm chromatin structure assay yields alphat values, which is another index of chromatin packaging. Higher alphat values, signifying poor packaging, occurred in both species following immunization with heterologous pituitary FSH. The binding of fluorosceinated pisum sativum agglutinin (PSA-FITC) to acrosome of sperm of monkeys and men was significantly low, compared with values at day 0 (control) during periods when cross-reactive FSH antibody titer was high and endogenous FSH was not detectable. Blockade of FSH function in monkeys by active immunization with a recombinant oFSH receptor protein corresponding to a naturally occurring messenger RNA (mRNA) also resulted in production of sperm with similar defects in chromatin packaging and reduced acrosomal glycoprotein concentration. Thus, it appears that in monkeys and men, lack of FSH signaling results in production of sperm that exhibit defective chromatin packaging and reduction in acrosomal glycoprotein content. These characteristics are similar to that exhibited by sperm of some class of infertile men. Interestingly, these alterations in sperm quality occur well ahead of decreased sperm counts in the ejaculate. PMID- 10714829 TI - Future of andrology. PMID- 10714828 TI - Human glyceraldehyde 3-phosphate dehydrogenase-2 gene is expressed specifically in spermatogenic cells. AB - Although the process of glycolysis is highly conserved in eukaryotes, several glycolytic enzymes have unique structural or functional features in spermatogenic cells. We previously identified and characterized the mouse complementary DNA (cDNA) and a gene for 1 of these enzymes, glyceraldehyde 3-phosphate dehydrogenase-s (Gapds). This gene is expressed only in spermatids. The enzyme appears to have an essential role in energy production required for fertilization, and it is reported to be susceptible to inhibition by certain environmental chemicals. We have now cloned and sequenced the cDNA for the human homologue of glyceraldehyde 3-phosphate dehydrogenase (GAPD2) and determined the structure of the gene. The messenger RNA (mRNA) was detected in testis, but not in 15 other human tissues analyzed by Northern blot technique. The deduced GAPD2 protein contains 408 amino acids and is 68% identical with somatic cell GAPD. GAPD2 has a 72-amino acid segment at the amino terminal end that is not present in somatic cell GAPD. This segment is proline-rich but contains smaller stretches of polyproline and is 30 amino acids shorter than the comparable segment of mouse GAPDS. The structure of the human GAPD2 gene was determined by polymerase chain reaction (PCR) to identify exon-intron junctions in a genomic clone and in total genomic DNA. The locations of these junctions in the GAPD2 gene corresponded precisely to those of the 11 exon-intron junctions in the mouse Gapds gene. Immunohistochemical studies found that GAPD2 is located in the principal piece of the flagellum of human spermatozoa, as are GAPDS in mouse and rat spermatozoa. GAPD2 extracted from human spermatozoa and analyzed by Western blot technique migrated with an apparent molecular weight of approximately 56,000, although the calculated molecular weight is 44 501. The conserved nature of the mouse, rat, and human enzymes suggests that they serve similar roles in these and other mammalian species. PMID- 10714830 TI - Future of andrology. PMID- 10714831 TI - A step in the right direction. PMID- 10714832 TI - Breathing pattern associated with respiratory comfort during automatic tube compensation and pressure support ventilation in normal subjects. AB - BACKGROUND: Automatic tube compensation (ATC) is a new option to support spontaneously breathing tracheally intubated patients. We have previously demonstrated an increased respiratory comfort compared to pressure support ventilation (PSV) in volunteers. Here we characterized the breathing pattern during ATC associated with respiratory comfort in comparison to PSV. Furthermore, we studied whether ATC can be substituted by a simple modification of PSV. METHODS: We exposed 10 volunteers breathing through a 7.5 mm endotracheal tube via mouthpiece to PSV with 1) immediate and 2) delayed pressure rise and to 3) ATC. Immediate changes of the respiratory pattern after mode shifts were analyzed in detail. Furthermore, the volunteers were instructed to indicate changes in comfort after transitions between these modes as increased, unchanged, or decreased. RESULTS: Decreased comfort was associated with a substantial increase of tidal volume, minute ventilation, gas flow, and pressure. No differences in respiratory comfort were perceived between immediate and delayed pressure rise during PSV. CONCLUSION: PSV resulted in excessive tidal volumes and airflow, which was perceived as discomfort. This cannot be avoided by a delayed pressure rise but can be by the more comfortable ATC. ATC seems to adapt better to the ventilatory demand than PSV. PMID- 10714833 TI - Estimation of cardiac preload changes by systolic pressure variation in pigs undergoing pneumoperitoneum. AB - BACKGROUND: Variations in systolic pressure arterial waveform (SPV) and its component have been shown to be a reasonable indicator of left ventricular preload. Creation of a pneumoperitoneum (PMOP) by insufflation of CO2 increases intrathoracic pressure, leading to overestimation of preload as assessed by pressure methods. The purpose of this study was to compare SPV with other standard methods in anaesthetized pigs. METHODS: We measured SPV and its DeltaDown component (deltaDown), pulmonary artery occlusion pressure (PAOP) and left ventricular short-axis cross-sectional area using transthoracic echocardiography (TTE) in 7 pigs, at baseline, after 12 mmHg PMOP and after an intravascular load with 10 ml/kg hydroxylethylstarch (HES). RESULTS: PMOP increased SPV from 12.9+/-4.9 to 16.9+/-5.5 mmHg (P<0.05) and decreased pulmonary compliance, with no change in PAOP or end-diastolic area assesssed by TTE. Intravascular volume loading significantly decreased SPV from 16.9+/-5.5 to 11.2+/-4.9 mmHg and deltaDown from 9.9+/-7.1 to 5.2+/-4.5 (P<0.05), and increased PAOP and end-diastolic area. Significant correlation between changes in deltaDown and EDA was noted following HES (r=0.78, P<0.05). CONCLUSION: In anaesthetized pigs, the creation of a PMOP alters SPV, likely by decreasing lung compliance. Once PMOP is established, changes in cardiac preload could be estimated by SPV analysis. PMID- 10714834 TI - Impact of acute renal failure on antioxidant status in multiple organ failure. AB - BACKGROUND: Acute renal failure (ARF) can be triggered or aggravated by reactive oxygen species (ROS) but established ARF per se might also affect the antioxidant defence mechanisms of the organism. We evaluated a broad pattern of antioxidants in critically ill patients with multiple organ failure with (MOF-ARF) and without acute renal failure (MOF) to identify any potential involvement of renal dysfunction in the depletion of the antioxidant system. METHODS: Observational study; 13 patients with MOF were investigated (9 with and 4 without ARF), and 17 healthy subjects served as controls. Blood samples were drawn after establishment of MOF. Plasma levels of ascorbate, alpha-tocopherol, retinol, beta-carotene, selenium and lipid peroxidation products (MDA) were determined and the activities of the antioxidant enzymes catalase (CAT), superoxide dismutase (SOD) and glutathione-peroxidase (GSH-PX) in erythrocytes were measured. In addition, ROS production (superoxide anion and hydrogen peroxide) in activated neutrophils was assessed. RESULTS: Plasma levels of ascorbate, beta-carotene and selenium were reduced in both patient groups, when compared to controls. Decrease in selenium was more pronounced in the MOF-ARF group. MDA levels were increased in both groups, again being more pronounced in MOF-ARF patients compared to MOF patients. Hydrogen peroxide production by neutrophils from both patient groups was lower than from controls. CONCLUSION: A depression of the antioxidative system is present in critically ill patients with MOF. In patients with associated ARF this was even more pronounced and plasma MDA levels were higher, suggesting an additional effect on the antioxidative potential in the presence of renal dysfunction and/or renal replacement therapy. PMID- 10714835 TI - Supraceliac aortic cross-clamping and declamping. Effects of dopexamine and dopamine on systemic and mesenteric hemodynamics, metabolism and intestinal tonometry in a rat model. AB - BACKGROUND: The effects of dopexamine and dopamine on mesenteric ischemia during reperfusion following aortic cross-clamping are not known. We determined intramucosal tonometric PCO2 and PCO2 gap using a rat model of supraceliac aortic cross-clamping and declamping. METHODS: Under pentobarbital and fentanyl anesthesia, 24 rats were surgically instrumented with arterial, right atrial, and portal venous catheters, ultrasonic flowprobes for measurements of abdominal aortic, superior mesenteric and carotid artery blood flow, and a pediatric tonometer for intestinal mucosal PCO2 measurements. Rats were randomized to receive a continuous infusion of dopexamine (10 x microg(-1) x kg(-1) x min(-1), n=8), dopamine (10 microg x kg(-1) x min(-1), n=8 ), or physiologic saline (control, n= 8), infused at a rate of 4 ml x kg(-1) x h(-1), administered throughout the experimental protocol. After 30 min of drug infusion, the aorta was cross-clamped at the supraceliac level for 30 min. Reperfusion following declamping was observed for 180 min. RESULTS: Intestinal tonometric PCO2 remained unchanged during drug treatment before aortic cross-clamping, increased similarly in all groups following declamping during early reperfusion, and recovered to baseline within 30 min of reperfusion. Dopexamine treatment was associated with higher lactate levels and increased heart rate (P<0.05) during aortic cross clamping. CONCLUSIONS: 1) Mesenteric ischemia, determined by intestinal tonometric PCO2 and PCO2 gap, recovers within 30 min of reperfusion following 30 min of aortic cross-clamping irrespective of drug treatment and, 2) dopexamine induced higher lactate levels and increased heart rate during aortic cross clamping and should be carefully analyzed for potentially adverse effects on cardiac function. PMID- 10714836 TI - Core and thenar skin temperature variation during prolonged abdominal surgery: comparison of two sites of active forced air warming. AB - BACKGROUND: This study was designed to compare the efficacy of two different sites of active forced air warming, upper body or lower body, to maintain normothermia; and their respective effect on thenar skin temperature in relation to the accelerographic monitoring of neuromuscular blockade during long-lasting abdominal surgery. METHODS: Twenty-six patients were randomised into two groups: upper body, (n=13) and lower body, (n=13), for intraoperative forced air warming. General anaesthesia was induced with thiopentone, sufentanil, and maintained with a mixture of N2O/O2/isoflurane. Pancuronium, 0.1 mg x kg(-1) was used to facilitate tracheal intubation. Reinjection doses of 0.01 mg x kg(-1) were administered once 25% recovery of first twitch height of train-of-four stimulation had occurred, or if surgical relaxation was estimated as inadequate by the surgeon. Thenar skin temperature and core temperature were monitored continuously. RESULTS: A similar trend for core temperature profile was observed in both groups. After an initial mild hypothermia, normothermia was reached progressively. Normothermia was obtained faster with lower body forced air warming than with upper body (2 h versus 3 h), P<0.05. Thenar skin temperature significantly increased during the first 90 min of surgery. This rise was significantly higher in the upper body group at 40 min and 60 min, P=0.03 and P=0.01, respectively. Stabilisation of thenar skin temperature occurred after 2 h without any further significant difference between groups. Muscle relaxant requirements were not significantly different between the groups. CONCLUSION: This study suggests that during long-lasting abdominal surgery, normothermia is maintained after 2-3 h by either upper or lower body active forced air warming. After an initial post-induction mild hypothermia, normothermia was achieved faster with lower body surface warming. Thenar skin temperature trend showed that it remained above 32 degrees C during most of the procedure in both groups. PMID- 10714837 TI - Lung lymph response to overinfusion with hydroxyethyl starch in sheep. Comparative studies of high and low molecular weight compounds. AB - BACKGROUND: Several hydroxyethyl starch (HES) solutions are available clinically. We performed comparative studies of low and high molecular weight HES to evaluate the effects on lung lymph flow in sheep, to see the difference in the types of HES. METHODS: We prepared awake sheep with vascular monitorings and lung lymph fistulas. We measured systemic artery pressure (Psa), pulmonary artery pressure (Ppa), and left atrial pressure (Pla) continuously. Cardiac output (CO) was measured every 30 min. Lung lymph flow (Qlym) was collected every 15 min. After baseline measurements, two HES solutions were infused over 2 h, respectively. Experiment 1 (n=6): low molecular weight HES (MW 70 000, substitution ratio 0.5 0.55), Experiment 2 (n=5): high molecular weight HES (MW 450 000, substitution ratio 0.7). RESULTS: Both low and high molecular HES behaved similarly as a volume expander, increasing Psa, CO, Pla and Ppa, and decreasing hematocrit. In addition, the actual oncotic pressure gradient (plasma - lymph) was widened after the start of either low or high molecular HES, but the value for high molecular HES was significantly higher than that for low molecular HES. Qlym of low molecular HES rose significantly from the baseline and the percent increase in Qlym for low molecular HES was significantly higher than that for high molecular HES. CONCLUSION: These data suggest that low molecular HES is as useful a plasma substitute as high molecular HES, but may increase lung fluid filtration in the overinfused state. PMID- 10714838 TI - The pharmacokinetics of anesthetic drugs and adjuvants during cardiopulmonary bypass. AB - The institution of cardiopulmonary bypass during cardiac surgery has profound effects on the plasma concentration of drugs and thus their therapeutic effectiveness. These changes occur through acute hemodilution, altered plasma protein binding, hypotension, as well as the use of hypothermia and heparin administration. Isolation of the lungs from the circulation and the possible sequestration of drugs in the bypass circuit also affect drug plasma concentrations on bypass. The individual characteristics of the drug in question are also important in determining the final plasma concentration: Lipid soluble drugs with a high volume of distribution may be more readily taken up by bypass equipment, but the initial fall in concentration at the start of cardiopulmonary bypass may be more readily counteracted by back diffusion into plasma, if large tissue stores have accumulated. The extent of the drug's plasma protein binding is of importance as the effective free fraction in plasma for highly bound drugs will be sensitive to changes in plasma protein binding brought on by factors such as hemodilution, heparin administration as well as alpha, acid-glycoprotein binding. Clearly the fate of drugs administered before or on bypass is complex and can only be accurately determined by specific studies evaluating drug plasma concentrations. This review updates the available data on anesthetics and drugs used during cardiac surgery in order that anesthetists may predict better the likely effect of drugs administered before or during cardiopulmonary bypass. PMID- 10714839 TI - A comparison of coracoid and axillary approaches to the brachial plexus. AB - BACKGROUND: Brachial plexus block by the coracoid approach does not require arm abduction and may be more effective than the axillary approach because of a more proximal injection of local anaesthetic. However, the clinical usefulness of the coracoid approach has not been tested in prospective controlled trials. The present randomized, observer-blinded study compared success rates, time to obtain a complete block, frequency of adverse effects and block discomfort in two groups of 30 patients, anaesthetized for hand surgery using either the coracoid or the axillary approach to the brachial plexus. METHODS: After subcutaneous infiltration with 5 ml of 1% mepivacaine/adrenaline the brachial plexus was located using a nerve stimulator and an insulated pencil-point needle. Ropivacaine 0.75%, 20-40 ml, depending on body weight, was used for the initial block. In the coracoid (C) group two plexus cords, and in the axillary (A) group four terminal nerves were electrolocated and the volume of ropivacaine was divided equally between them. Spread of analgesia to the arm was assessed every 5 min, by an anaesthetist unaware of the block technique. The block was defined as effective (complete) when analgesia was present in all five sensory nerve areas distal to the elbow. Incomplete blocks were supplemented 30 min after the initial block. RESULTS: In the C group a median 11 min was required for block performance as compared to 12 min in the A group (NS). Onset of block was shorter and the frequency of incomplete blocks lower in the A group (median 17 min and 17%) than in the C group (30 min and 47%, respectively). Lack of analgesia of the ulnar nerve was the main cause of incomplete initial blocks in the C group. All incomplete blocks were successfully supplemented. However, total time to obtain complete block was shorter in the A group than in the C group (29 min vs. 41 min, P<0.05). Accidental arterial puncture occurred in seven patients (five in C and two in A group), which resulted in two haematomas, both in the C group (NS). No permanent sequelae were observed. CONCLUSION: The axillary approach to the brachial plexus using four injections of ropivacaine results in a faster onset of block and a better spread of analgesia than the coracoid approach using two injections. PMID- 10714840 TI - Intraperitoneal lidocaine for postoperative pain after laparoscopy. AB - BACKGROUND: A controversy exists over the effectiveness and clinical value of intraperitoneal local anaesthetics for treating pain after laparoscopic cholecystectomy. The use of intraperitoneal lidocaine was evaluated in this study. METHODS: At the end of surgery, 200 ml saline containing 200 mg lidocaine, or the same volume of saline, were randomly splashed under the right diaphragmatic surface in 50 patients in a double-blind manner. Postoperative shoulder and abdominal pain intensity were recorded on a numeric grading scale and a visual analogue scale, respectively. Analgesic consumption was also recorded. Respiratory function tests were compared before and after surgery. Side effects and recovery variables were assessed by the nurses at 2-h intervals. RESULTS: The incidence, severity and duration of shoulder pain were reduced from 40% of patients scoring 3.9+/-0.2 for duration of 17.9+/-0.2 h in the control group to 12% scoring 2.5+/-0.5 for duration of 1.6+/-0.01 h in the lidocaine group. Lidocaine treated patients had significantly less abdominal postoperative pain immediately on return to the ward and during the first postoperative day (P<0.05). "No pain on deep inspiration" was reported by 72% of patients in the lidocaine group immediately on return to the ward compared to 8% of those in the control group. Analgesic consumption for 24 h after surgery was significantly less in the lidocaine group (P<0.05). There were no significant differences in respiratory function tests, recovery variables or incidence of side effects between the two groups. CONCLUSION: Intraperitoneal lidocaine is simple to use and results in a long-lasting reduction of pain after a single administration. PMID- 10714841 TI - The incidence of transient neurologic symptoms (TNS) after spinal anaesthesia in patients undergoing surgery in the supine position. Hyperbaric lidocaine 5% versus hyperbaric bupivacaine 0.5%. AB - BACKGROUND: The incidence of TNS after spinal anaesthesia is a problem. Especially the use of hyperbaric lidocaine in patients placed in the lithotomy position during surgery has been associated with a high incidence of TNS. The present study was performed to investigate whether TNS is present more frequently in patients undergoing surgery in the supine position with use of hyperbaric lidocaine compared with hyperbaric bupivacaine. METHOD: Seventy patients were included and randomised to receive either hyperbaric lidocaine or hyperbaric bupivacaine. All patients were contacted on the first and third postoperative days by an anaesthesiologist blinded to the local anaesthetic used. The patients were asked about symptoms of TNS, pain not associated with the operation area, and asked to grade the complaints after a verbal analogue score from 0 to 10. RESULTS: We found a total of ten patients who showed signs of TNS. There were nine patients in the lidocaine group (26%) who showed signs of TNS compared to only one patient in the bupivacaine group (3%) (P<0.01). The average score of TNS complaints was 3.5. A total of 13 patients (19%) complained of back pain. There were no significant differences with regard to which local anaesthetic was used. The average score of back pain was 3.3. CONCLUSION: TNS is a significant problem in patients having spinal anaesthesia with hyperbaric lidocaine compared to hyperbaric bupivacaine, both in the supine position. For day-case surgery, TNS would start after dismissal from hospital. The use of hyperbaric lidocaine is therefore questionable, even though these problems are of an order that the majority of patients would still choose spinal anaesthesia for future operations. PMID- 10714842 TI - Diclofenac or acetaminophen for analgesia in paediatric tonsillectomy outpatients. AB - BACKGROUND: In order to establish an effective drug regimen, we compared the analgesic efficacy of oral diclofenac and high-dose acetaminophen on pain after tonsillectomy. METHODS: In this randomised, double-blind study 48 children, 5 to 15 years of age, following tonsillectomy were assigned to receive either diclofenac 2-3 mg kg(-1) 24 h(-1) (n=24) or acetaminophen 90 mg kg(-1) 24 h(-1) (n=24) for the first three days after surgery. Postoperative pain was assessed by self-report each day before scheduled medication at 7 h, 12 h, 18 h and 23 h. RESULTS: The number of children rating severe pain was high in both the diclofenac group, 5-50%, and in the acetaminophen group, 12-58% during the three day study period. Pain scores in the diclofenac group were only significantly lower at 12 h on day 1-3 compared to pain scores in the acetaminophen group (P<0.05). None of the children in the diclofenac group experienced any episodes of nausea/vomiting compared to 9 children in the acetaminophen group on day 1. The incidences of nausea/vomiting increased with pain (P<0.05). None of the 48 children experienced any episodes of bleeding. CONCLUSIONS: This study indicates that diclofenac was no more effective than high-dose acetaminophen (90 mg vs. 60 mg kg(-1) 24 h(-1)) for analgesia, but resulted in a lower incidence of nausea and vomiting in patients following tonsillectomy. PMID- 10714843 TI - Postoperative pain control by epidural analgesia after transabdominal surgery. Efficacy and problems encountered in daily routine. AB - BACKGROUND: The efficacy of postoperative epidural pain treatment has been well documented in controlled studies. However, the literature concerning results of daily routine use of this method often only emphasises certain aspects of it. METHODS: A prospective study of 168 patients scheduled for major surgery with transabdominal access was performed in order to evaluate efficacy, side effects, complications and rate of acceptance of postoperative epidural pain treatment. The epidural catheter was placed before surgery and the patients received epidural analgesia by a bupivacaine/morphine mixture for 3 days postoperatively, continued by another 3 days with bolus injections of morphine only. RESULTS: Only few complications followed the insertion of the epidural catheter, but in about 16% of the patients the epidural catheter or the drugs administered by it made reinsertion or change in the type of analgesia necessary during the first 3 post operative days. Despite the possibility for individualising the treatment and p.r.n. analgesics, pain relief when coughing and moving during day 1-3 was insufficient in 30-50% of the patients. Serious side effects were rare, but pruritus was frequent, as were the symptoms of nausea and vomiting. The patients were generally satisfied with the treatment; however, a small group had unacceptable pain when the epidural catheter was inserted. CONCLUSION: Analgesia was insufficient when coughing and moving in an unacceptably large number of the patients. Also the number of epidural catheter related problems was high. In order to make early intervention possible, the patients and epidural catheters should be observed daily and systematically by members of the staff competent to detect possible problems. PMID- 10714844 TI - High-dose rectal and oral acetaminophen in postoperative patients--serum and saliva concentrations. AB - BACKGROUND: The primary purpose of the study was to examine the absorption of acetaminophen by measuring serum and saliva concentrations produced by a standard postoperative acetaminophen dosing regimen and secondary to examine the correlation between saliva and serum concentrations of acetaminophen after rectal and oral dosing. METHODS: Twenty-four women, aged 18-60 years, scheduled for minor gynaecological laparoscopic surgery were studied. Patients received acetaminophen 2000 mg suppositories after surgery and oral doses of 1000 mg at 4 and 8 h postoperatively. Alfentanil was available via patient-controlled analgesia. Saliva and blood samples were collected postoperatively. RESULTS: At 1, 2, 3, and 4 h after rectal dosing the saliva concentrations (mean+/-SD) were 15.2+/-5.9 micromol/l, 33.7+/-12.5 micromol/l, 45.5+/-19.1 micromol/l, and 55.4+/ 23.1 micromol/l, respectively. The serum concentrations at 2 and 4 h were 31.0+/ 11.2 micromol/l and 54.8+/-23.8 micromol/l, respectively. Additional oral dosing resulted in saliva concentrations at 5, 8, and 9 h of 99.7+/-49.5 micromol/l, 106.9+/-31.7 micromol/l, and 139.3+/-55.4 micromol/l, respectively, with coincident serum concentrations of 100.1 +/- 50.2 micromol/l, 105.6+/-29.0 micromol/l, and 141.2+/-52.1 micromol/l. After rectal dosing the linear regression resulted in r2=0.96, P<0.001 and saliva/ serum-ratio=0.99. After additional oral dosing the outcome of linear regression was: r2=0.90, P<0.001 and saliva/serum-ratio= 1.00. CONCLUSION: The slow and ongoing absorption process resulting in no maximum concentration within 4 h after administration of 2000 mg acetaminophen suppositories makes this rectal regimen therapeutically irrational for treatment of postoperative pain. The significant ratio and linear correlation between saliva and serum concentrations of acetaminophen suggests that saliva could be used instead of blood to monitor acetaminophen administration in patients. PMID- 10714845 TI - Quality of recovery in children: sevoflurane versus propofol. AB - BACKGROUND: Sevoflurane, with its low pungency and low blood and tissue solubility, is an attractive anaesthetic in paediatric outpatient surgery. Propofol-anaesthesia is recognised for its rapid and clear-headed emergence. This study was designed to compare emergence and recovery characteristics of sevoflurane and propofol anaesthesia for tonsillectomy in children. METHODS: Children aged 3-10 years, undergoing elective tonsillectomy, were randomly assigned to receive propofol (n=25, induction with 3 mg x kg(-1), maintenance with 100-250 microg x kg(-1) min(-1)) or sevoflurane anaesthesia (n=25, induction 7 vol.%, maintenance 2-3 vol.%). Tracheal intubation was performed with alfentanil 20 microg x kg(-1) and atracurium 0.5 mg x kg(-1). Ventilation was controlled to maintain normocapnia and all patients received N2O/O2 (60:40 vol.%) for induction and maintenance of anaesthesia. At the end of surgery infiltration of the operative sites with bupivacaine 2 mg x kg(-1) was provided for postoperative analgesia. Emergence, recovery, discharge times, and incidence of side effects were compared between the two groups. RESULTS: Time to extubation (14 vs 15 min), time to response to simple verbal command (21 vs 21 min) and time to discharge from the recovery room (45 vs 50 min) were similar in the sevoflurane and propofol groups, respectively. There was a significantly greater incidence of postoperative agitation in the sevoflurane group (46%) compared with the propofol group (9%) (P=0.008). This did not, however, delay discharge from the recovery room. The incidence of nausea and vomiting was not significantly different (8% vs 0%; P=0.49). CONCLUSION: In children, recovery from anaesthesia with sevoflurane results in a higher incidence of agitation compared with propofol. PMID- 10714846 TI - Comparison between pentazocine, pethidine and placebo in the treatment of post anesthetic shivering. AB - BACKGROUND: We have compared the effects of pethidine, pentazocine and placebo in the treatment of post-anesthetic shivering. METHODS: Forty-five patients who shivered after routine surgery were allocated randomly to receive normal saline (n=15), pentazocine 7.5 mg (n=15) or pethidine 17.5 mg (n=15). RESULTS: After 10 min, 13 patients had stopped shivering in the pethidine group, which was significantly more than the incidence in the two other groups (placebo=2; pentazocine=4) (P<0.01). Pentazocine was not significantly different from normal saline in affecting shivering. CONCLUSION: We conclude that pentazocine 7.5 mg was not effective in the treatment of post-anesthetic shivering. PMID- 10714847 TI - Midlatency median nerve evoked responses during recovery from propofol/sufentanil total intravenous anaesthesia. AB - BACKGROUND: Median nerve somatosensory evoked responses (MnSSER) are frequently used to monitor the integrity of the somatosensory pathway during surgery. We investigated MnSSER components during the wakeup phase from anaesthesia with propofol/sufentanil, because detailed information is lacking about the reversibility of anaesthetic induced changes of MnSSER. The aim of the study was to document precisely the MnSSER waves in relation to the clinical awakening. The hypothesis was that anaesthetic induced MnSSER changes are reversed when the patient becomes responsive after anaesthesia. METHODS: In 20 gynaecological patients anaesthesia was maintained with propofol 8 mg kg(-1) h(-1) supplemented by bolus injections of sufentanil. MnSSER were recorded at C4' (N20, P25, N35, P45, N50) following electrical median nerve stimulation on the day before surgery, after the end of surgery during anaesthesia and every 5 min during recovery, till the patients were responsive again and able to identify a shown object. RESULTS: While the primary cortical MnSSER complex N20P25 regained baseline values, the cortical latencies > or =35 ms remained prolonged (P<0.001) and the amplitudes P45N50 were suppressed (P< or =0.013), when the patients were responsive after 26+/-7 min following anaesthesia. However, the amplitudes P25N35 exceeded their corresponding baseline value (P<0.01) CONCLUSION: Persistent changes of MnSSER waves > or =35 ms reflect impaired signal processing along the somatosensory pathway following propofol/sufentanil anaesthesia when the patients are responsive again. Further studies combining MnSSER recording with distinct neuro-psychological tests are needed to define the clinical relevance of these findings. PMID- 10714848 TI - Halogenated volatile anesthetics inhibit carbon monoxide-stimulated soluble guanylyl cyclase activity in rat brain. AB - BACKGROUND: Because of halogen contents, halogenated volatile anesthetics (HVA) have a similarity to nitric oxide (NO) in terms of great affinity for the ferrous ion. Interactions between HVA and NO at the ferrous ion of soluble guanylyl cyclase (sGC) have been reported in different tissues. Carbon monoxide (CO), a more stable gas than NO, activates sGC by the same mechanism as NO. This study was undertaken to examine the effect of HVA on CO-stimulated sGC activity in rat brain. METHODS: Sprague-Dawley rat brain was homogenized and ultracentrifuged. The resulting supernatant was used as sGC fraction. The fraction was incubated with CO and HVA, and the activity of sGC was determined by measuring cyclic guanosine monophosphate (cGMP) production using an enzyme immunoassay in aliquots of the supernatant. RESULTS: CO clearly increased cGMP production in a dose dependent manner. Sevoflurane and isoflurane produced significant and dose dependent inhibition of CO-stimulated sGC activity. There was no difference in the inhibitory effect between the two anesthetics. GTP dose-dependently increased CO-stimulated cGMP production. Both anesthetics decreased GTP production, but the inhibition by the anesthetics was not significant at higher GTP concentrations. CONCLUSIONS: These results suggest that HVA can compete with CO at the ferrous ion of sGC and inhibit the activity of this enzyme. PMID- 10714849 TI - Safety, efficacy, and long-term results of a modified version of rapid opiate detoxification under general anaesthesia: a prospective study in methadone, heroin, codeine and morphine addicts. AB - BACKGROUND: In the present study a method of rapid opiate detoxification under general anaesthesia has been evaluated regarding the safety, the efficacy in preventing withdrawal symptoms, and the long-term results. In addition, it was investigated whether the profile and severity of withdrawal symptoms depend on the type of opiate abused (methadone, heroin, codeine, morphine). METHODS: Seventy-two opiate addicts were detoxified in an intensive care unit (ICU). Anaesthesia was induced and maintained using propofol infusion. Patients were endotracheally intubated. The opiate receptor antagonist naltrexon was administered into the stomach via a nasogastric tube. Withdrawal symptoms before and after the detoxification treatment were assessed using an objective and a subjective opiate withdrawal scale (OOWS, SOWS). After detoxification patients entered a long-term naltrexone maintenance programme as well as a supportive psychotherapy programme. Vital organ function was monitored using haemodynamic and respiratory parameters as well as body temperature. RESULTS: Organ function parameters were stable during the whole treatment in all patients and no anaesthetic complications were registered. Minor side effects such as bradycardia or hypotension were observed in 20 patients. Compared to patients with pre existing heroin, codeine, or morphine abuse respectively, patients from the methadone maintenance programme had significantly higher (P<0.01) OOWS as well as SOWS values after the treatment. Twelve months after the detoxification 49 patients (68%) were abstinent from opiates whereas 17 patients had relapsed during the period of follow-up. Six patients were lost during follow-up. CONCLUSIONS: Rapid opiate detoxification under general anaesthesia is a safe and efficient method to suppress withdrawal symptoms. This treatment may be of benefit in patients who particularly suffer from severe withdrawal symptoms during detoxification and who have failed repeatedly to complete conventional withdrawal. Methadone patients have more withdrawal symptoms than other opiate addicts. PMID- 10714850 TI - Contractures in skeletal muscle of malignant hyperthermia susceptible patients after in vitro exposure to sevoflurane. AB - BACKGROUND: Sevoflurane, a potent inhalational anaesthetic agent that is structurally similar to halothane, has some favourable characteristics, but may also be able to trigger malignant hyperthermia (MH) in susceptible patients. The diagnosis of malignant hyperthermia susceptibility relies on the in vitro contracture test on skeletal muscle. The present study was undertaken to investigate whether exposure to sevoflurane of muscles of malignant hyperthermia susceptible (MHS) patients would also cause an abnormal contracture. METHODS: Muscle fascicles obtained from three MHS patients, one malignant hyperthermia non susceptible (MHN) patient, two control patients and one malignant hyperthermia equivocal (MHE) patient were exposed to sevoflurane instead of halothane in the in vitro contracture test, carried out according to the protocol of the European Malignant Hyperthermia Group. The muscle fascicles were surplus to diagnostic requirements. Sevoflurane concentrations in the testbath were measured using a headspace gas chromatographic technique. RESULTS: The kinetics of sevoflurane concentration in the testbath were similar to those of halothane. An in vitro contracture response of 2 mN or more was seen in all four MHS/MHE patients with sevoflurane but not in the three control/MHN patients. The magnitude of muscle contracture in the sevoflurane test was less than in the conventional halothane test at comparable testbath concentrations. CONCLUSIONS: Sevoflurane can trigger an abnormal contracture in human muscle in vitro. This is indicative of malignant hyperthermia susceptibility. Exposure to sevoflurane should be avoided in patients thought to be susceptible to malignant hyperthermia. PMID- 10714851 TI - The impact of 4-chloro-m-cresol in heparin formulas on malignant hyperthermia: in vitro and in vivo. AB - BACKGROUND: The preservative 4-chloro-m-cresol (4CmC) is a specific activator of sarcoplasmic Ca2- release and induces contractures in skeletal muscles of malignant hyperthermia susceptible (MHS) patients in vitro. Clinical formulas of heparin contain 4CmC. We studied whether (a) these heparin formulas induce contractures in isolated MHS and normal (MHN) human skeletal muscles and whether (b) significant serum levels of 4CmC are reached after heparinization in cardiopulmonary bypass patients. METHODS: (a) In vitro, muscle bundles of 16 MHS and 22 MHN patients were exposed to the heparin formula Liquemin (containing 4CmC 0.08 mg/500 IU), to chlorocresol and to preservative-free heparin in the in vitro contracture test. (b) In vivo, serum 4CmC levels of 12 patients receiving Liquemin 500 IU/ kg before cardiopulmonary bypass were determined at 1, 5 and 60 min by high-pressure liquid chromatography. RESULTS: (a) For Liquemin and 4CmC, significant contractures were measured with MHS muscles compared to MHN muscles at 61.4 microM 4CmC. (b) In control sera, the detection threshold for 4CmC was 4 microM. Concentrations of 4CmC in all patients' serum samples were below this threshold. CONCLUSION: Heparin formulas, containing 4CmC, induce dose-dependent contractures in vitro in MHS human skeletal muscle at about 60 microM 4CmC. However, in vivo, 4CmC serum concentrations with therapeutic heparinization are less than 1/15 of the in vitro concentration. The lipophilicity of 4CmC with a high volume of distribution may account for these findings. MHS patients seem not to be at risk from clinical heparin formulas containing chlorocresol. PMID- 10714852 TI - Less adrenergic activation during cataract surgery with total intravenous than with local anesthesia. AB - BACKGROUND: Previous studies reported that, contrary to local anesthesia, cataract surgery under inhalational anesthesia is associated with substantial adrenergic activation. We tested the hypothesis that total intravenous anesthesia (TIVA) with propofol and alfentanil produces less or comparable adrenergic activation during cataract surgery than local anesthesia. METHODS: Patients were randomly assigned to peribulbar local block (n=10) or TIVA (n=10). The heart rate, blood pressure, plasma concentrations of catecholamines, cortisol, and glucose were assessed at seven pre-, intra-, and post-operative time points. RESULTS: In the patients given local anesthesia, plasma concentrations of epinephrine, norepinephrine and cortisol did not change significantly. In contrast, plasma epinephrine decreased by roughly 66% during TIVA: from 45+/-16 to 15+/-8 pg/ml. Plasma norepinephrine concentration decreased by roughly 50%, from 462+/-265 to a minimum value of 219+/-6 pg/ml and plasma cortisol concentrations decreased by roughly 61%, from 16.4 ng/ml to 6.4 ng/ml. Blood pressure and heart rates remained near baseline values during local anesthesia. In contrast, systolic blood pressure decreased by 30% and heart rate by 12 beats/min during TIVA. CONCLUSION: The presented study and available data clearly suggest that local anesthesia produces the best adrenergic and hemodynamic stability during cataract surgery. Contrary to previously reported results on inhalational anesthesia (thiopentone/enflurane), the TIVA regimen used effectively prevents the adrenergic and metabolic response during cataract surgery. PMID- 10714853 TI - Treatment of severe low back pain with opioids during pregnancy in a patient with incomplete tetraplegia. AB - We report a case of severe low back pain during pregnancy in a woman with incomplete tetraplegia due to viral myelitis. The pain was interpreted as a radiculopathy in the presence of multiple herniated discs. Surgical intervention was not indicated and physiotherapy failed; therefore, a symptomatic drug treatment with oral analgesics was initiated. To minimise the total daily opioid dosage and the potential risk of a neonatal withdrawal syndrome due to opioids, the route of administration was changed from oral to epidural. Adequate pain relief was maintained with this regimen until caesarean section was necessary. The neonatal withdrawal symptoms after delivery were mild. Residual pain slowly diminished after delivery and the patient was able to discontinue opioid therapy. The aetiology of low back pain remains unclear and may be multifactorial. PMID- 10714854 TI - Sevoflurane as a sole anaesthetic for thymectomy in myasthenia gravis. AB - Myasthenia gravis is a challenging situation for anaesthesiologists due to its neuromuscular involvement. The main concerns are respiratory muscle weakness and side effects due to a heavy dose of anticholinesterases. This limits the use of sedatives, hypnotics and muscle relaxants. Inhalational anaesthetics are best suited. We describe our experience with sevoflurane as a sole anaesthetic in a child having juvenile-type myasthenia gravis with thymoma, who underwent thymectomy by midsternal incision. Very smooth and short duration of induction (35 s) and easy intubation within 60 s without use of muscle relaxant were the remarkable features. Sevoflurane in oxygen and nitrous oxide (MAC=0.5-0.7) was used for maintenance of anaesthesia. Recovery was smooth and fast with no residual respiratory insufficiency. Hence we found sevoflurane to be a highly suitable agent for thymectomy in mysthenia gravis. PMID- 10714855 TI - Perfusion method to measure oesophageal pressure. PMID- 10714856 TI - A review and proposed nomenclature for major proteins of the milk-fat globule membrane. AB - The characteristics and possible functions of the most abundant proteins associated with the bovine milk-fat globule membrane are reviewed. Under the auspices of the Milk Protein Nomenclature Committee of the ADSA, a revised nomenclature for the major membrane proteins is proposed and discussed in relation to earlier schemes. We recommend that proteins be assigned specific names as they are identified by molecular cloning and sequencing techniques. The practice of identifying proteins according to their Mr, electrophoretic mobility, or staining characteristics should be discontinued, except for uncharacterized proteins. The properties and amino acid sequences of the following proteins are discussed in detail: MUC1, xanthine dehydrogenase/oxidase, CD36, butyrophilin, adipophilin, periodic acid Schiff 6/7 (PAS 6/7), and fatty acid binding protein. In addition, a compilation of less abundant proteins associated with the bovine milk-fat globule membrane is presented. PMID- 10714857 TI - Cheese maturity assessment using ultrasonics. AB - The relationship between Mahon cheese maturity and ultrasonic velocity was examined. Moisture and textural properties were used as maturity indicators. The ultrasonic velocity of the cheese varied between 1630 and 1740 m/s, increasing with the curing time mainly because of loss of water, which also produced an increase of the textural properties. Because of the nature of low-intensity ultrasonics, velocity was better related to those textural parameters that involved small displacements. Ultrasonic velocity decreased with increasing temperature because of the negative temperature coefficient of the ultrasonic velocity of fat and the melting of fat. These results highlight the potential use of ultrasonic velocity measurements to rapidly and nondestructively assess cheese maturity. PMID- 10714858 TI - Effect of administration of fermented milk containing whey protein concentrate to rats and healthy men on serum lipids and blood pressure. AB - The effect of fermented milk supplemented with whey protein concentrate on the serum lipid level of rats was investigated. The serum total cholesterol level for the group fed fermented milk with both Lactobacillus casei TMC0409 and Streptococcus thermophilus TMC 1543 was significantly lower than that of the control group (P<0.05) in rats. Furthermore, the effect of the longterm intake of this fermented milk on the serum lipid level of twenty healthy adult men was investigated. During the 8-wk study, the volunteers consumed 200 ml of fermented milk or placebo in the morning and evening. Blood samples were drawn for analysis three times, just before taking the experimental diet, and after 4 wk and 8 wk of consumption. After 8 wk, the high density lipoprotein cholesterol level for the fermented milk group showed a significant rise after 4 wk (P<0.05), whereas that of the placebo group showed no change even after 4 wk (P>0.05). The triglyceride level for the fermented milk group lowered significantly after 4 wk (<0.05), whereas that of the placebo group showed no change even after 4 wk (P>0.05). The atherogenic index [(total cholesterol - high density lipoprotein cholesterol)/high-density lipoprotein cholesterol] for the fermented milk group decreased significantly from 4.24 to 3.52 (P<0.05). The systolic blood pressure lowered significantly by the intake of fermented milk (P<0.05) On the other hand, such effect was not observed in the placebo group (P>0.05). These results indicate potential of the development of fermented milk with multiple therapeutic effects. PMID- 10714859 TI - Effects of somatic cell count on quality and shelf-life of pasteurized fluid milk. AB - Milk was collected from eight Holstein cows four times before and four times after intramammary infection with Streptococcus agalactiae. Postinfection milk had significantly higher somatic cell count (SCC) (849,000 cells/ml) than preinfection milk (45,000 cells/ml). High SCC raw milk had more lipolysis and proteolysis than low SCC raw milk. Pasteurized, homogenized, 2% fat milks from pre- and postinfection periods were stored at 5 degrees C and analyzed for lipolysis, proteolysis, microbial quality, and sensory attributes at 1, 7, 14, and 21 d post processing. During refrigerated storage, the average rates of free fatty acid increase (i.e., lipolysis) and casein hydrolysis in high SCC milk were, respectively, three and two times faster than those in low SCC milk. In general, standard plate counts, coliform counts, and psychrotrophic bacterial counts of both the high and low SCC milks remained low (<100,000 cfu/ ml) during 5 degrees C storage. Low SCC milk maintained high organoleptic quality for the entire 21-d shelf-life period. However, for high SCC milk, between 14 and 21 d, sensory defects were detected, which resulted in low overall quality ratings. The sensory defects mainly included rancidity and bitterness and were consistent with higher levels of lipolysis and proteolysis. Hence, mastitis adversely affected the quality of pasteurized fluid milk. It is recommended that the fluid milk industry consider implementation of premium quality payment programs for low SCC milks. PMID- 10714860 TI - Organic acids and volatile flavor components evolved during refrigerated storage of kefir. AB - Kefir samples were prepared and transferred to sterile jars for storage at 4 degrees C. After 0, 7, 14, and 21 d of storage, the pH, organic acid, and volatile flavor component content were determined to monitor possible flavor changes during storage. Stored samples were analyzed for organic acid (orotic, citric, pyruvic, lactic, uric, acetic, propionic, butyric, and hippuric) content by HPLC with UV detection at 275 nm. Acetoin, ethanol, acetaldehyde, and diacetyl were monitored using gas chromatography equipped with a headspace autosampler. There was no significant decrease in average pH of samples between d 0 and 21 of storage (P>0.05). Lactic acid concentration increased during storage, reaching a maximum of 7739 ppm by d 21. Orotic and citric acids increased slightly during storage. Although pyruvic and hippuric acids are produced during fermentation, neither was detected during storage. Acetic, propionic, and butyric acids were not detected during kefir storage. Ethanol concentrations increased during storage and reached 0.08% by d 21. The amounts of acetaldehyde and acetoin, common flavor substances in many cultured dairy products, increased during fermentation. Acetaldehyde content in kefir samples doubled from d 0 to 21, reaching a final concentration of 11 microg/g. During storage, the concentration of acetoin decreased from 25 ppm on d 0 to 16 ppm on d 21. However, diacetyl, another common flavor component in cultured dairy products, was not detected during fermentation or storage. PMID- 10714861 TI - Factors associated with cure after therapy of clinical mastitis caused by Staphylococcus aureus. AB - One hundred and fifty-nine cases of clinical Staphylococcus aureus mastitis were analyzed to detect factors associated with bacteriological cure after therapy. On 100 Dutch dairy farms, data were collected from four clinical trials with five intramammary treatment regimes designed to treat beta-lactamase-positive pathogens. Infected quarters were treated three times, with a 12-h interval between treatments. Treatment was extended for 2 d if results of the trial treatment were, according to the owner, not satisfactory. The overall bacteriological cure rate was 52%. The bacteriological cure rate of clinical beta lactamase-negative S. aureus mastitis was significantly higher than that of clinical beta-lactamase-positive S. aureus mastitis. Bacteriological cure was also significantly higher if somatic cell count of the cow was low at the milk recording prior to the onset of the clinical mastitis. The bacteriological cure rate of clinical beta-lactamase-negative S. aureus mastitis was also significantly higher after an extended treatment compared with no extended treatment. The seriousness of the various clinical symptoms and the bacteriological cure rate of clinical S. aureus mastitis were not associated. PMID- 10714862 TI - Autocrine insulin-like growth factor II inhibits beta-casein mRNA expression in a mammary cell line. AB - A hallmark of mammary cell differentiation is the induction of beta-casein mRNA expression. A mouse mammary epithelial cell line (COMMA-1D) was treated with insulin, hydrocortisone (HC), and prolactin (Prl) at concentrations (50, 500, and 20 ng/ml, respectively) that resulted in less than half-maximal beta-casein mRNA expression. The cells secreted insulin-like growth factor (IGF)-II (106 pg/ml per 24 h) in the condition media under these conditions. Replacement of insulin with rhIGF-II (150 ng/ml) resulted in significantly less beta-casein mRNA expression. Long-Arg IGF-I (50 ng/ml) was similar to insulin in terms of its ability to induce differentiation, but its activity differed from that of insulin in that it also induced cell proliferation. When the two receptor-specific IGF-II analogs, Arg54,55IGF-II and Leu27IGF-II, were used in studies, only at high concentrations (150 ng/ml) was either analog capable of stimulating any beta-casein mRNA expression. When autocrine IGF-II was immuno-neutralized or bound by the addition of rhIGF binding protein-3 (IGFBP-3)beta-casein mRNA expression was enhanced seven-fold and three-fold, respectively. Exogenous application of IGF-II to counteract the IGF-II mAb stimulation resulted in increased cellular growth and reduced differentiation. We conclude that autocrine IGF-II inhibits mammary cell differentiation and that the blockage of autocrine IGF-II benefits mammary cell differentiation. PMID- 10714864 TI - Correlation between bovine milk somatic cell count and polymorphonuclear leukocyte level for samples of bulk milk and milk from individual cows. AB - The influence of various factors on the concentrations of polymorphonuclear neutrophilic leukocytes (PMN) in milk samples from bulk tanks and individual cows was investigated. While somatic cell counts (SCC) and PMN level were in both cases significantly correlated, lower correlation coefficients were found between SCC and PMN for samples of bulk tank milks than for milk samples from individual cows. Furthermore, plots of PMN concentrations versus SCC showed great variability in PMN in milk samples of similar total SCC. One factor that may lead to variability in bulk tank PMN levels was shown to be increased proportions of high SCC milk in the bulk tank mixture, which result in relatively high PMN levels without excessive elevation of total SCC. In milk samples from individual cows, it was found that there was also a significant seasonal influence on milk PMN content, with milk from cows calving in the spring having, at SCC > 160,000 cells/ ml, higher proportions of PMN in the total milk SCC than milk from autumn calving cows. The results of this study suggest that the concentration of PMN may be a useful indicator of herd status in bulk tank monitoring schemes. PMID- 10714863 TI - Evaluation of eight cow-side ketone tests in milk for detection of subclinical ketosis in dairy cows. AB - The objective of this study was to evaluate eight cowside ketone tests when used with milk for detection of subclinical ketosis. A total of 469 dairy cows in the first week of lactation were studied. Twelve percent of these cows had subclinical ketosis, defined as >1400 micromol of beta-hydroxybutyrate/L of blood serum. The Pink test liquid and the Ketolac test strip were highly sensitive for subclinical ketosis when used with milk. The Uriscan and Rapignost test strips were poorly sensitive; the Ketostix, Ketur-Test, and Medi-Test-Keton test strips and the Acetonreagenz test tablet were insensitive for subclinical ketosis when used with milk. Pink and Ketolac milk ketone tests are potentially useful tools for use in a routine monitoring program to detect subclinical ketosis in early postpartal dairy cows. PMID- 10714865 TI - Short communication: seasonal effects on development of bovine embryos produced by in vitro fertilization in a hot environment. AB - The objective of this study was to determine if season affected the production of in vitro-derived bovine embryos from oocytes of cattle in a subtropical environment. Ovaries (approximately 75% beef cattle, including many with Bos indicus breeding) were collected from an abattoir. Oocytes were obtained and subjected to in vitro maturation and fertilization. Embryos were then cultured in CR1aa medium. Cleavage rate averaged 72.2+/-9.7% and was not different between months of collection. In addition, no differences were observed in the percent of oocytes or embryos that became blastocysts on d 8 or 9 after insemination. Least squares means averaged across months for percent oocytes and cleaved embryos to blastocyst on d 8 were 22.8+/-7.5% and 31.2+/-9.4%, respectively. When d 8 blastocysts were classified according to stage of development (nonexpanded, expanded, and hatched), an effect of month was observed that reflected month-to month variation and not a consistent change associated with season. Taken together, results failed to indicate an effect of season on in vitro production of embryos in a subtropical environment. PMID- 10714866 TI - In situ disappearance of malate from alfalfa and bermudagrass hay. AB - The objectives of this study were to determine the rate of malate and dry matter disappearance from different forages in the rumen. Four nonlactating, ruminally cannulated Holstein cows were fed a hay-based diet. Samples of early and late harvested alfalfa, Coastal bermudagrass, and Tifton 85 bermudagrass hays were ground, placed in nylon in situ bags, and ruminally incubated for 0, 0.5, 1, 2, 4, 8, 12, 24, and 48 h. After incubation, samples were rinsed, freeze-dried, extracted, and analyzed for malate content by HPLC with an organic acid column. When forages were incubated in the rumen, malate concentrations were less than 0.55 mg/g of dry matter at 0.5 h and remained low for the 48-h incubation period. These results suggest that malate was solublized and utilized within 30 min after reaching the rumen. Dry matter digestibility of both forages increased with time and was different across forages. Both alfalfa samples were digested to a greater extent between 0.5 and 24 h than either type of bermudagrass, but after 48 h the early maturity Tifton 85 digestibility was similar to alfalfa. Even though it is more common to feed unground forages to ruminants, these in situ results suggest that once malate is available in the rumen it will disappear quickly. PMID- 10714867 TI - Effects of starch source and level of forage neutral detergent fiber on performance by dairy cows. AB - The effectiveness of neutral detergent fiber (NDF) from soyhulls and whole cottonseed for replacing NDF from forage was evaluated in a lactation trial during wk 10 to 25 of lactation. Forty-eight cows were blocked and randomly assigned within a block to one of four diets: 1) 21% forage NDF with corn 2) 16% forage NDF with corn, 3) 16% forage NDF with corn and wheat (1:1) and, 4) 11% forage NDF with cottonseed and corn. Soybean hulls were added at approximately 23.0% of dry matter (DM) for the 16 and 11% forage NDF diets to replace forage and formulate diets with 35% nonfiber carbohydrates. Actual forage NDF concentration were 17.8, 14.0, 13.9, and 9.4%, respectively. Dry matter intake and milk yield were highest for cows fed 11% forage NDF with cottonseed. Milk fat percentage was higher for cows consuming 21% forage NDF and 16% forage NDF with corn than for cows fed the two other diets. Cows fed 16% forage NDF with corn and wheat experienced milk fat-protein inversion, but ruminal acetate:propionate was lower for cows fed 11% forage NDF than cows fed 16% forage NDF. Body weight (BW) and BW change were not different among treatments. Time spent chewing was similar among all diets. For cows in midlactation, forage NDF may be reduced to 9 to 11% when cottonseed is at 11% of DM and dietary nonstructural carbohydrates are at 30% of DM. Forage NDF may be reduced to 14 to 16% without cottonseed when nonstructural carbohydrates are at 30% of DM. PMID- 10714868 TI - Interactions between forage and wet corn gluten feed as sources of fiber in diets for lactating dairy cows. AB - Twelve early lactation Holstein cows (4 fistulated) were used in replicated 4x4 Latin squares with 4-wk periods to determine the effective neutral detergent fiber (NDF) content of wet corn gluten feed and to measure the effect of forage particle size on ruminal mat consistency and passage rate of wet corn gluten feed. Diets were 1) 23.3% NDF (17.4 percentage units of NDF from alfalfa silage), 2) diet 1 plus 11.1 additional percentage units of NDF from alfalfa silage, 3) diet 1 plus 10.7 percentage units of NDF from wet corn gluten feed, and 4) 8.6 percentage units of NDF from alfalfa silage plus 8.9 percentage units of NDF from coarsely chopped alfalfa hay and 10.7 percentage units of NDF from wet corn gluten feed. The calculated effective NDF factor for wet corn gluten feed, using change in milk fat concentration per unit change in NDF, was 0.74 compared with an assumed 1.0 for alfalfa silage. Rumination activity was measured to calculate a physically effective NDF factor for wet corn gluten feed, which was only 0.11 compared with 1.0 for alfalfa silage. Physically effective NDF also was determined for wet corn gluten feed by wet sieving; 22% of the particles were retained on the 3.35-mm screen or greater. Ruminal mat consistency increased and passage rate of wet corn gluten feed decreased with added hay. The inclusion of chopped alfalfa hay to a diet containing wet corn gluten feed increased ruminal mat consistency, rumination activity, and slowed passage rate, resulting in greater ruminal digestion of NDF from wet corn gluten feed. Depending on the response variable, the effectiveness of NDF from wet corn gluten feed varied from 0.11 to 0.74. PMID- 10714870 TI - The influence of long-term supplementation with biotin on the prevention of lameness in pasture fed dairy cows. AB - In a double-blind study, the influence of biotin supplementation on lameness in dairy cows was investigated over a 13-mo period. The experimental site was a tropical upland environment and involved over 2705 Holstein and Friesian cows on 20 participating farms. Cows on 10 farms received biotin at a rate of 20 mg/head per day in the concentrate, and cows on 10 other farms received feed without the biotin supplement. Premixes with or without biotin were incorporated into a grain concentrate that was fed at a constant rate to cows at milking. Farmers maintained accurate records of the nature of hoof problems and any treatment applied. Each herd was evaluated for locomotion scores at 8-wk intervals. Locomotion scores were significantly correlated with the number of days with measurable rainfall per month (r = 0.88). The biotin-supplemented herds exhibited better locomotion scores than the unsupplemented herds. In the wet summer period the number of lame cows, as observed by the farmer, were significantly fewer during the rainy period for the biotin-supplemented herds and required fewer antibiotic treatments than unsupplemented herds. Most hoof lesions were most commonly observed in the outer claws of the hind limb. Daily milk production (17.3 vs. 18.5 L) was not affected by biotin supplementation. Reduced milk fat percentage and somatic cell counts of bulk milk were recorded in the biotin supplemented herds during the wet, summer period. PMID- 10714869 TI - Feeding oleamide to lactating Jersey cows 1. Effects on lactation performance and milk fatty acid composition. AB - Oleamide was previously reported to resist ruminal biohydrogenation and elevate milk oleic acid concentration when fed to lactating Holstein cows. To determine if Jersey cows responded similarly to oleamide, four lactating Jersey cows (mean 417 kg of body weight and 64 days in milk) were fed four diets in a 4x4 Latin square with 2-wk periods. Diets were total mixed ration containing 47% corn silage and 53% concentrate (dry matter basis) and were supplemented with no added fat (control), or with 3.5% added fat from either higholeic canola oil, a commercial source of oleamide, or oleamide synthesized from oleic acid and urea. The canola oil supplement had no effect on milk yield or composition. Compared to canola oil, the oleamide supplements reduced milk yield, dry matter intake, and milk fat and protein contents. Milk oleic acid concentration increased from 17.4% of total fatty acids for the control diet to 22.1% for the canola oil diet. Both oleamides further increased milk oleic acid to 30.0 and 27.1% of total fatty acids for the commercial and synthesized oleamides, respectively. Milk palmitic acid was reduced and stearic acid was increased by all fat supplements but more so by the oleamides than by the canola oil. Consistent with previous reports that fatty acyl amides resist ruminal biohydrogenation, feeding oleamide to Jersey cows in this study increased milk oleic acid concentration but had negative effects on feed intake and milk yield. PMID- 10714871 TI - Effect of alpha-tocopherol deprivation on the involution of mammary gland in sheep. AB - The objective of this study was to investigate the effect of alpha-tocopherol deprivation on mammary gland involution and apoptosis in sheep. Two groups of four single lamb ewes were used. The control group received 100 mg/d of RRR-alpha tocopherol supplementation and the experimental group received no vitamin E supplementation. After 3 mo of suckling, ewes were dried off, and blood samples from the jugular vein and tissue biopsies from the mammary gland were collected at d 1, 3, 5, and 8 after dry-off. The experimental group had lower plasma concentrations of alpha-tocopherol (1.8 vs. 4.2 micromol/L), lower glutathione peroxidase activity in erythrocytes, and higher concentration of malondialdehyde in plasma than the control group. Immunohistochemical analysis of tissue samples resulted in marked differences of bcl-2 and bax protein expressions during involution and between groups. The bax expression was decreased by alpha tocopherol deprivation at 1, 3, and 5 d, but not at 8 d, while the bcl-2 score was higher only at 8 d (1.5 vs. 0.0 for experimental and control groups, respectively). As a result, the bcl-2 to bax ratios were increased for the experimental group at 1 and 8 d. During involution, apoptotic counts increased (from 0.12 to 4.06%), but no effects were detected in relation to bcl-2 to bax ratio and alpha-tocopherol. These results indicate that alpha-tocopherol can control bcl-2 expression, but not apoptosis in cells of the mammary gland during involution. PMID- 10714872 TI - Effect of oil content and kernel processing of corn silage on digestibility and milk production by dairy cows. AB - Corn silages were produced from a high oil corn hybrid and from its conventional hybrid counterpart and were harvested with a standard silage chopper or a chopper equipped with a kernel processing unit. High oil silages had higher concentrations of fatty acids (5.5 vs. 3.4% of dry matter) and crude protein (8.4 vs. 7.5% of dry matter) than the conventional hybrid. Processed silage had larger particle size than unprocessed silage, but more starch was found in small particles for processed silage. Dry matter intake was not influenced by treatment (18.4 kg/d), but yield of fat-corrected milk (23.9 vs. 22.6 kg/d) was increased by feeding high oil silage. Overall, processing corn silage did not affect milk production, but cows fed processed conventional silage tended to produce more milk than did cows fed unprocessed conventional silage. Milk protein percent, but not yield, was reduced with high oil silage. Milk fat percent, but not yield, was higher with processed silage. Overall, processed silage had higher starch digestibility, but the response was much greater for the conventional silage hybrid. The concentration of total digestible nutrients (TDN) tended to be higher for diets with high oil silage (71.6 vs. 69.9%) and tended to be higher for processed silage than unprocessed silage (71.7 vs. 69.8%), but an interaction between variety and processing was observed. Processing conventional corn silage increased TDN to values similar to high oil corn silage but processing high oil corn silage did not influence TDN. PMID- 10714873 TI - Chemical and physical properties of processed newspaper compared to wheat straw and wood shavings as animal bedding. AB - Because of continuing concerns about the safety and the suitability of recycled newspaper as an animal bedding material, municipal curbside-collected newspaper was processed into chopped and pelleted forms for comparison studies with wheat straw and kiln-dried pinewood shavings. Measurements included nutrient, heavy metal, dioxin and furan content, particle size distribution, density, combustion potential, and water-holding capacity. Recycled newspaper, straw, and wood shavings tested below or equivalent to National Research Council dietary tolerance levels and US Environmental Protection Agency toxic equivalent levels. Small particle size distribution was shavings > straw > all forms of newspaper. The density of pelleted newspaper was 50-fold greater than that of chopped newspaper and straw and 15-fold greater than shavings. In simulated flash burns, chopped newspaper, straw, and shavings ignited, and flames spread rapidly in newspaper and shavings and lasted the longest in shavings. Pelleted newspaper did not ignite. Chopped and pelleted forms of newspaper and wood shavings had higher water holding capacities (>400%) than did straw (200%). Animal industries can, in confidence, utilize recycled newspaper as an animal bedding material, providing that sources of low toxicity are identified, and suitable processed forms are produced. PMID- 10714874 TI - Feed-borne transmission and case clustering of BSE. AB - An unresolved issue in the epidemiology of bovine spongiform encephalopathy (BSE) in the UK is what precisely determines the degree to which cases of disease in cattle are clustered within herds throughout the course of the epidemic. This paper presents an analysis of feed-borne transmission at the herd level and tests various models of case-clustering mechanisms, associated with heterogeneity in exposure to infectious feed, against observed epidemic pattern. We use an age structured metapopulation framework in which the recycling of animal tissue between herds via feed producers is explicitly described. We explore two alternative assumptions for the scaling with herd size of the within-herd risk of exposure of an animal to infectious material. We find that whereas exposure heterogeneity caused by variation in feed and offal processing methods and by variation in per-animal feed uptake can explain the pattern of case clustering seen in the BSE epidemic, exposure heterogeneity due to the aggregation of infectivity within feed cannot. PMID- 10714875 TI - Constraints on polyploid evolution: a test of the minority cytotype exclusion principle. AB - Polyploid evolution is often considered a mechanism of instant speciation; yet the establishment of rare tetraploids within diploid populations may be constrained by a frequency-dependent mating disadvantage (minority cytotype exclusion principle). I tested this hypothesis using experimental populations of Chamerion angustifolium (Onagraceae) that contained different proportions of tetraploids and diploids. Fitness, measured as total seed production over the entire flowering season, was calculated from a census of flower number and estimates of ovule number per flower and proportion of seed set per fruit. The fitness of tetraploids relative to diploids was frequency dependent, increasing from 0.4, when tetraploids were rare, to 0.7 when at 50% and 1.15 when they were in the majority (67%). This pattern exists because of a negative relationship between tetraploid frequency and seed set per fruit in diploids. Seed set in tetraploids was independent of cytotype frequency. The frequency-independent effect in tetraploids reflects higher assortative mating, partly because of non random patterns of bee visitation. Bees visited a disproportionately high number of diploid inflorescences; however, the proportion of successive flights between tetraploids increased above random expectations as the frequency of tetraploids decreased. These results provide the first experimental test of frequency dependent fitness in diploid-polyploid mixtures and suggest an important role for more gradual, population processes governing the evolution of polyploidy in natural populations. PMID- 10714876 TI - Monophyly of brachiopods and phoronids: reconciliation of molecular evidence with Linnaean classification (the subphylum Phoroniformea nov.). AB - Molecular phylogenetic analyses of aligned 18S rDNA gene sequences from articulate and inarticulate brachiopods representing all major extant lineages, an enhanced set of phoronids and several unrelated protostome taxa, confirm previous indications that in such data, brachiopod and phoronids form a well supported clade that (on previous evidence) is unambiguously affiliated with protostomes rather than deuterostomes. Within the brachiopod-phoronid clade, an association between phoronids and inarticulate brachiopods is moderately well supported, whilst a close relationship between phoronids and craniid inarticulates is weakly indicated. Brachiopod-phoronid monophyly is reconciled with the most recent Linnaean classification of brachiopods by abolition of the phylum Phoronida and rediagnosis of the phylum Brachiopoda to include tubiculous, shell-less forms. Recognition that brachiopods and phoronids are close genealogical allies of protostome phyla such as molluscs and annelids, but are much more distantly related to deuterostome phyla such as echinoderms and chordates, implies either (or both) that the morphology and ontogeny of blastopore, mesoderm and coelom formation have been widely misreported or misinterpreted, or that these characters have been subject to extensive homoplasy. This inference, if true, undermines virtually all morphology-based reconstructions of phylogeny made during the past century or more. PMID- 10714877 TI - What is not a bird of paradise? Molecular and morphological evidence places Macgregoria in the Meliphagidae and the Cnemophilinae near the base of the corvoid tree. AB - The cnemophiline 'birds of paradise' (Cnemophilinae) and Macgregor's 'bird of paradise' (Macgregoria) have traditionally been included in the Paradisaeidae although their relationships within the group have been enigmatic and subject to repeated discussion in the literature. Here we use sequences from two mitochondrial genes, cytochrome b and cytochrome oxidase I, along with a suite of morphological characters, to investigate their relationships to paradisaeids and other members of the passerine Parvorder Corvida. The combined data strongly support the removal of both groups from the birds of paradise: the cnemophilines are basal members of the Corvoidea and Macgregoria is a member of the Meliphagoidea and embedded in the honeyeaters (Meliphagidae) close to the genus Melipotes. The amount of sequence divergence among basal passeriforms and members of the Corvida, as well as available fossil evidence for Australian corvidans, suggest that cnemophilines represent an ancient lineage within the corvoid radiation. Because cnemophilines and Macgregoria have been placed at the base of the paradisaeid tree, hypotheses of morphological, behavioural and ecological character-state transformations within the family will require reanalysis. PMID- 10714878 TI - Coalitions of relatives and reproductive skew in cooperatively breeding white winged choughs. AB - We used DNA fingerprinting to examine reproductive skew in cooperatively breeding white-winged choughs, Corcorax melanorhamphos, which live in groups of up to 20 individuals. Before a severe drought, groups that had been stable for multiple years were characterized by long-term monogamy involving a single breeding pair (high skew). After the drought, new groups formed from the amalgamation of multiple individuals and coalitions of relatives. At most one member of each faction succeeded in breeding, such that skew was dependent on the number of unrelated factions, and not group size. In the new groups, dominant males and females with supporting relatives were always successful. Whereas most females without support also gained breeding positions, many males without family support failed to breed. Thus subordinates gain indirect fitness by first helping related males to secure a breeding position, and then helping to raise their young. Our study demonstrates the advantage of operating in coalitions, and suggests that the acquisition of future allies may be a major benefit of helping behaviour in this species. PMID- 10714879 TI - Dynamic mate-searching tactic allows female satin bowerbirds Ptilonorhynchus violaceus to reduce searching. AB - Females can maximize the benefits of mate choice by finding high-quality mates while using search tactics that limit the costs of searching for mates. Mate searching models indicate that specific search tactics would best optimize this trade-off under different conditions. These models do not, however, consider that females may use information from previous years to improve mate searching and reduce search costs in subsequent years. We followed female satin bowerbirds Ptilonorhynchus violaceus during mate searching and reconstructed their search patterns. We found that females who chose very attractive males typically mated with the same male in the following year, resulting in these females sampling fewer males than those who switched mates. In contrast, females who mated with less attractive males typically rejected their previous mates and searched longer for more attractive mates in the following mating season. A potential cost to mate searching is suggested by the observed increase in the likelihood of force copulation attempts from marauding males with increased searching. Our results suggest that by using past experiences to adjust their search tactics, females may obtain high-quality mates while limiting search costs. These results emphasize the need to consider historical effects in studies of sexual selection, especially for long-lived species with stable display sites. PMID- 10714881 TI - Origins and ecological consequences of pollen specialization among desert bees. AB - An understanding of the evolutionary origins of insect foraging specialization is often hindered by a poor biogeographical and palaeoecological record. The historical biogeography (20,000 years before present to the present) of the desert-limited plant, creosote bush (Larrea tridentata), is remarkably complete. This history coupled with the distribution pattern of its bee fauna suggests pollen specialization for creosote bush pollen has evolved repeatedly among bees in the Lower Sonoran and Mojave deserts. In these highly xeric, floristically depauperate environments, species of specialist bees surpass generalist bees in diversity, biomass and abundance. The ability of specialist bees to facultatively remain in diapause through resource-poor years and to emerge synchronously with host plant bloom in resource-rich years probably explains their ecological dominance and persistence in these areas. Repeated origins of pollen specialization to one host plant where bloom occurs least predictably is a counter-example to prevailing theories that postulate such traits originate where the plant grows best and blooms most reliably Host-plant synchronization, a paucity of alternative floral hosts, or flowering attributes of creosote bush alone or in concert may account for the diversity of bee specialists that depend on this plant instead of nutritional factors or chemical coevolution between floral rewards and the pollinators they have evolved to attract. PMID- 10714880 TI - Sex allocation and population structure in apicomplexan (protozoa) parasites. AB - Establishing the selfing, rate of parasites is important for studies in clinical and epidemiological medicine as well as evolutionary biology Sex allocation theory offers a relatively cheap and easy way to estimate selfing rates in natural parasite populations. Local mate competition (LMC) theory predicts that the optimal sex ratio (r*; defined as proportion males) is related to the selfing rate (s) by the equation r* = (1-s)/2. In this paper, we generalize the application of sex allocation theory across parasitic protozoa in the phylum Apicomplexa. This cosmopolitan phylum consists entirely of parasites, and includes a number of species of medical and veterinary importance. We suggest that LMC theory should apply to eimeriorin intestinal parasites. As predicted, data from 13 eimeriorin species showed a female-biased sex ratio, with the sex ratios suggesting high levels of selfing (0.8-1.0). Importantly, our estimate of the selfing rate in one of these species, Toxoplasma gondii, is in agreement with previous genetic analyses. In contrast, we predict that LMC theory will not apply to the groups in which syzygy occurs (adeleorins, gregarines and piroplasms). Syzygy occurs when a single male gametocyte and a single female gametocyte pair together physically or in close proximity, just prior to fertilization. As predicted, data from four adeleorin species showed sex ratios not significantly different from 0.5. PMID- 10714882 TI - Imperfect assessment and limited information preclude optimal strategies in male male fights in the orb-weaving spider Metellina mengei. AB - Agonistic behaviour between male orb-web spiders Metellina mengei competing for access to female webs was examined in field experiments to test the major predictions of game theory. Winners of fights were significantly larger than losers, particularly with respect to the length of the first pair of legs, which are sexually dimorphic in this species and used extensively in agonistic encounters. The size of the winning male had no influence on contest intensity or duration, and neither did relative size. However, fight intensity and duration were both positively correlated with the size of the losing male. Resident males won significantly more contests than intruders. Winning intruders were significantly larger than winning residents and it was these winning intruders that tended to produce the longer fights. Female weight and hence reproductive value had a marked influence on fight intensity and duration of fights won by the intruder but not those won by the resident. This indicates that only the resident obtains information about the female. These data are discussed with reference to the discrepancy with theory and a failure of some contestants to obtain information on resource value and relative contestant size necessary to optimize fight strategy. PMID- 10714883 TI - Phylogeography and population history of Atlantic mackerel (Scomber scombrus L.): a genealogical approach reveals genetic structuring among the eastern Atlantic stocks. AB - Despite the resolving power of DNA markers, pelagic and migratory marine fish species generally show very little geographical population structuring. In mackerel (Scomber scombrus L.) population differentiation has been detected only at a transatlantic scale. By applying two regions in mitochondrial DNA (mtDNA) (D loop and cytochrome b (cytb)) in combination with genealogical and frequency based statistical approaches, our data suggest population differentiation among eastern Atlantic spawning stocks. In contrast, and indicative of homing behaviour, no genetic structuring was observed among shoals of individuals outside the spawning season. Among spawning stocks, mtDNA D-loop sequences detected differentiation within the eastern Atlantic, while the cytb gene detected transatlantic differentiation. The impact of recurrent events (e.g. gene flow restricted by isolation by distance) and historic events (e.g. population range expansions) among spawning stocks was investigated applying a nested cladistic analysis of geographical distribution of cytb haplotype lineages. In the eastern Atlantic, historical population range expansion, presumably in connection with recolonization of northern areas after the last glaciation, is suggested to be the main factor determining mtDNA lineage distribution. This was supported by estimates of mtDNA nucleotide diversity, where the highest diversity was observed for the stock spawning in the Bay of Biscay, for which the size estimate is only 15% of the largest stock (Celtic Sea). In addition to revealing population differentiation, our data demonstrate the importance of sampling strategy and the power of applying statistical methods addressing both ongoing and historical population processes. PMID- 10714884 TI - Dissecting latitudinal diversity gradients: functional groups and clades of marine bivalves. AB - The latitudinal diversity gradient, with maximum taxonomic richness in the tropics, is widely accepted as being pervasive on land, but the existence of this pattern in the sea has been surprisingly controversial. This is partly due to Thorson's influential claim that the normal latitudinal diversity gradient occurs in marine epifauna (taxa living on the surface of the substratum) but not in infauna (burrowing or boring into the substratum), a contrast he attributed to the greater spatial and temporal environmental homogeneity of infaunal habitats. In an analysis of 930 species of north-eastern Pacific marine shelf bivalves, we found that bivalves as a whole, and both infauna and epifauna separately, show a strong latitudinal diversity gradient (measured as number of species per degree latitude) that is closely related to mean sea surface temperature (SST), even in analyses of residuals and first differences. This agrees with results for marine gastropods, but contradicts Thorson's environmental homogeneity hypothesis. The relationship between SST and diversity is consistent with a species-energy hypothesis, but the linkages from SST to diversity remain unclear. Most bivalve clades within broad functional groups conform to the general latitudinal trend, except for the deposit-feeding protobranchs. This group's non-directional pattern may be related to its mode of development, because a similar effect is seen in several other groups locked into this low-fecundity, non-feeding larval mode. PMID- 10714885 TI - Individual contributions to babysitting in a cooperative mongoose, Suricata suricatta. AB - Evolutionary explanations of cooperative breeding based on kin selection have predicted that the individual contributions made by different helpers to rearing young should be correlated with their degree of kinship to the litter or brood they are raising. In the cooperative mongoose or meerkat, Suricata suricatta, helpers babysit pups at the natal burrow for the first month of pup life and frequent babysitters suffer substantial weight losses over the period of babysitting. Large differences in contributions exist between helpers, which are correlated with their age, sex and weight but not with their kinship to the young they are raising. Provision of food to some group members raises the contributions of individuals to babysitting. We discuss the implications of these results for evolutionary explanations of cooperative behaviour. PMID- 10714886 TI - The evolution of sperm length in moths. AB - Sperm form and function remain poorly understood despite being of fundamental biological importance. An instructive approach has been to examine evolutionary associations across comparable taxa between sperm characters and other, potentially selective reproductive traits. We adopt this approach here in a comparative study examining how sperm lengths are associated with male and female reproductive characters across moths. Primary data have revealed Lepidoptera to be an ideal order for examination: there is profound variation in the dimensions (but not organization) of the reproductive traits between closely related species which all share a monophyletic ancestry, for example, eupyrene sperm length varies from 110 to 12,675 microm. Eupyrene (normal fertilizing) and apyrene (anucleate and non-fertile) sperm lengths are positively correlated across taxa and both sperm types show positive associations with mating pattern (as measured by the residual testis size). At fertilization, eupyrene sperm must migrate down the often elongated female spermathecal duct from storage to unite with the ovum. Across taxa, the elongation of this duct is associated with increased eupyrene sperm length, suggesting a positive female influence on sperm size since longer, more powerful sperm may be selected to migrate and/or compete successfully down greater ductal lengths. Apyrene sperm length is not associated with female reproductive tract dimensions. However, we found a positive relationship between the residual testis volume and spermathecal volume, suggesting coevolution between male investment in spermatogenesis and the extent of the female sperm storage capacity. Within males, there is a positive association between the two organs which form the ejaculate-containing spermatophore: the testes and the accessory gland. The 'trade-up' in investment to these components is discussed in relation to paternal investment and mating patterns. PMID- 10714887 TI - Adrenomedullin: potential in physiology and pathophysiology. AB - Adrenomedullin (ADM), a 52-amino acid ringed-structure peptide with C-terminal amidation, was originally isolated from human pheochromocytoma. ADM mediates vasodilatory and natriuretic properties through the second messenger cyclic adenosine 3',5'-monophosphate (cAMP), nitric oxide and the renal prostaglandin system. ADM immunoreactivity and its gene are widely distributed in cardiovascular, pulmonary, renal, gastrointestinal, cerebral and endocrine tissues. ADM is also synthesized and secreted from vascular endothelial and smooth muscle cells. When injected intravenously, ADM increases flow rates predominantly in organs in which the ADM gene is highly expressed, suggesting that ADM acts as a local autocrine and/or paracrine vasoactive hormone. In addition, ADM is a circulating hormone and its plasma concentration is increased in various cardiorenal diseases such as hypertension, chronic renal failure and congestive heart failure. Current evidence suggests that ADM plays an important role in fluid and electrolyte homeostasis and cardiorenal regulation, however further investigations are required to address the importance of ADM under various physiological and pathophysiological conditions. PMID- 10714888 TI - Adenosine receptors are involved in the control of acute naloxone-precipitated withdrawal: in vitro evidence. AB - The effects exerted by adenosine A1 and A2 receptor agonists and antagonists on the acute opiate withdrawal induced by morphine were investigated in vitro. Following a 4 min in vitro exposure to morphine, the guinea-pig isolated ileum exhibited a strong contracture after the addition of naloxone. The P1 adenosine receptor agonist, adenosine, was able to reduce dose-dependently naloxone precipitaded withdrawal. The same effect was induced by the adenosine A1 receptor agonist, N6-Cyclopentyladenosine (CPA) whereas the selective adenosine A2A receptor agonist CGS 21680 increased the naloxone-precipitated withdrawal phenomenon. Dipyridamole, a blocker of adenosine reuptake, induced a significant reduction of morphine dependence. Caffeine, an adenosine receptor antagonist, significantly increased the naloxone-precipitated withdrawal effect in a concentration dependent manner. The same effect was observed with 8 phenyltheophylline (8PT), an A1 adenosine receptor antagonist, whereas 3,7 dimethyl-1-propargylxanthine (DMPX), an A2 adenosine receptor antagonist, reduced the naloxone-precipitated withdrawal phenomenon. The results of our experiments indicate that both A1 and A2 adenosine receptor agonists and antagonists are able to influence opiate withdrawal in vitro, suggesting an important functional interaction between the adenosine receptors and opioid withdrawal. PMID- 10714889 TI - Evaluation of Z-(R,R)-IQNP for the potential imaging of m2 mAChR rich regions of the brain and heart. AB - Alterations in the function or density of the m2 muscarinic (mAChR) subtype have been postulated to play an important role in various dementias such as Alzheimer's disease. The ability to image and quantify the m2 mAChR subtype is of importance for a better understanding of the m2 subtype function in various dementias. Z-(R)-1-Azabicyclo[2.2.2]oct-3-y (R)-alpha-hydroxy-alpha-(1-iodo-1 propen-3-yl)-alpha-phenylacetate (Z-(R,R)-IQNP) has demonstrated significant uptake in cerebral regions that contain a high concentration of m2 mAChR subtype in addition to heart tissue. The present study was undertaken to determine if the uptake of Z-(R,R)-IQNP in these regions is a receptor mediated process and to identify the radiospecies responsible for binding at the receptor site. A blocking study demonstrated cerebral and cardiac levels of activity were significantly reduced by pretreatment (2-3 mg/kg) of (R)-3-quinuclidinyl benzilate, dexetimide and scopolamine, established muscarinic antagonists. A direct comparison of the cerebral and cardiac uptake of [I-125]-Z-(R,R)-IQNP and [I-131]-E-(R,R)-IQNP (high uptake in ml, m4 rich mAChR cerebral regions) demonstrated Z-(R,R)-IQNP localized to a higher degree in cerebral and cardiac regions containing a high concentration of the m2 mAChR subtype as directly compared to E-(R,R)-IQNP. In addition, a study utilizing [I-123]-Z-(R,R)-IQNP, [I 131]-iododexetimide and [I-125]-R-3-quinuclidinyl S-4-iodobenzilate, Z-(R,R)-IQNP demonstrated significantly higher uptake and longer residence time in those regions which contain a high concentration of the m2 receptor subtype. Folch extraction of global brain and heart tissue at various times post injection of [I 125]-Z-(R,R)-IQNP demonstrated that approximately 80% of the activity was extracted in the lipid soluble fraction and identified as the parent ligand by TLC and HPLC analysis. These results demonstrate Z-(R,R)-IQNP has significant uptake, long residence time and high stability in cerebral and cardiac tissues containing high levels of the m2 mAChR subtype. These combined results strongly suggest that Z-(R,R)-IQNP is an attractive ligand for the in vivo imaging and evaluation of m2 rich cerebral and cardiac regions by SPECT. PMID- 10714890 TI - Radioimmunoassay for orexin A. AB - A radioimmunoassay for orexin A has been developed. Anti-orexin A antiserum was raised in New Zealand white rabbits immunized with a conjugate of synthetic orexin A with bovine serum albumin. This antibody did not crossreact with orexin B, hypothalamic hormones, pituitary hormones, neuropeptides or gut hormones. Radioiodination of orexin A was performed with the chloramin T method, followed by purification of radioiodinated material on Sephadex G-25 column. Orexin A was extracted from tissues using acid-acetone. The assay was performed with a double antibody system. The dilution curve of acid-acetone-extracts of rat hypothalamus in the radioimmunoassay system was parallel to the standard curve. The recovery of tissue orexin A was about 80%,and the intra-assay and inter-assay variations were 5.2% and 7.8%, respectively. Orexin A was found in the hypothalamus, cerebrum and testis. These data suggest that this assay system is suitable for the measurement of tissue orexin A and that orexin A is found in the central nervous system and testis. PMID- 10714891 TI - Vessel dilator, long acting natriuretic peptide, and kaliuretic peptide increase circulating prostaglandin E2. AB - Prostaglandin E2 (PGE2) increases in the circulation of persons with congestive heart failure (CHF), but the cause of this increase is unknown. Prostaglandins are not stored, therefore, they cannot be released in response to congestive heart failure itself but rather need to have their synthesis stimulated by a hormone or some other substance. Prostaglandin E2's biologic properties are nearly identical to four peptide hormones originating from amino acids 1-30 [long acting natriuretic peptide], 31-67 [vessel dilator], 79-98 [kaliuretic peptide] and 99-126 [atrial natriuretic peptide, ANP] of the 126 amino acid ANP prohormone. ANP previously has been found to have no effect on circulating PGE2 concentrations in persons with CHF. The present investigation was designed to determine if one or more of the other three atrial natriuretic peptides might increase PGE2 when infused at their respective 100 ng/kg body weight/minute concentrations for 60 minutes in persons with congestive heart failure. Vessel dilator increased PGE2 8-fold (P<0.001) in the first 20 minutes of its infusion with PGE2 remaining 2-3 fold increased (P<0.05) for 60 minutes after stopping its infusion. Long acting natriuretic peptide did not increase PGE2 until 40 minutes of its infusion but it caused the maximal increase (27-fold; P<0.001) of PGE2 of the three peptide hormones tested. Kaliuretic peptide's stimulated increase of PGE2 also began in a delayed fashion but its effects lasted the longest, with PGE2 being increased (P<0.05) for two hours after the cessation of kaliuretic peptide's infusion. This investigation demonstrates that 1) three endogenous peptide hormones increase PGE2 in the circulation and 2) suggests that the known increase in PGE2 in CHF may be in part secondary to these peptides. PMID- 10714892 TI - Distribution of alpha1-adrenoceptor subtype mRNA and identification of subtype responsible for renovascular contraction in human renal artery. AB - This study was intended to quantify the amounts of the alpha1-adrenoceptor subtype mRNAs in human renal artery and to demonstrate the distribution of receptor subtypes responsible for the contraction of the renal artery. RNase protection assay showed that the mean amount of alpha1a mRNA was much greater than that of alpha1b or alpha1d mRNAs in both the main and branch renal arteries. However, the abundance of alpha1a mRNA in human renal artery was much less than in our previous data in the prostate. In situ hybridization showed that all alpha1 subtype mRNAs were localized in the smooth muscle cells of the tunica media of the artery, and the distribution pattern of these three mRNAs in the main artery was the same as in the branch artery. However, the intensity of signals for alpha1d and alpha1b antisense RNAs probes was lower than that for the alpha1a antisense RNA probe. In the functional study, concentration-response curves to noradrenaline pretreated with KMD-3213, an alpha1A/L-adrenoceptor selective antagonist, seemed to be biphasic in nature. Chloroethyclonidine (CEC) failed to inactivate the noradrenaline-induced contraction, and prazosin showed relatively low affinity with a pA2 value of 8.8. These data suggest that the alpha1A/L-adrenoceptor mediates primarily those responses to noradrenaline in this artery. The other alpha1-adrenoceptor subtypes could also mediate the secondary contractile response to noradrenaline in this artery. PMID- 10714893 TI - Neurotensin increases endogenous glutamate release in rat cortical slices. AB - In the present study, the effects of the tridecapeptide neurotensin [NT(1-13)] and its fragments, NT(1-7) and NT(8-13), on endogenous glutamate release from rat cortical slices, were evaluated. NT(1-13) (100-1000 nM) slightly increased spontaneous glutamate release, while it was ineffective at 1 and 10 nM concentrations. Neither the biologically active NT fragment NT(8-13) nor the inactive one NT(1-7) affected basal glutamate release. NT(1-13) (1-1000 nM) enhanced potassium (35 mM)-evoked glutamate release displaying a bell-shaped concentration response curve. In addition NT(8-13) (10 nM) increased K+-evoked glutamate release similarly to the parent peptide (10 nM), while the biologically inactive fragment NT(1-7) (10-100 nM) was ineffective. The effects of NT(1-13) and NT(8-13) were fully counteracted by the selective neurotensin receptor antagonist SR48692 (100 nM). These findings suggest that NT plays a role in regulating cortical glutamate transmission. PMID- 10714894 TI - Direct determination of endothelin receptor antagonist levels in plasma using a scintillation proximity assay. AB - An assay using scintillation proximity bead technology has been developed suitable for the quantitation of endothelin (ET) receptor antagonists in preclinical and clinical samples of plasma. The assay measures the competitive inhibition of radiolabelled ET-1 binding to ET(A) receptor membranes bound to wheat germ agglutinin (WGA)-coated scintillation proximity assay (SPA) beads in the presence of plasma containing A-127722, a potent orally active, ET(A) selective ET antagonist. The assay requires as little as 50 microl plasma and no extraction procedure is needed. The SPA methodology eliminates the need for the separation of bound from free ligand. Using this method, A-127722 could be directly quantified in rat plasma with a detection limit of 1 ng/ml. PMID- 10714895 TI - Long term neurological and behavioral effects of graded perinatal asphyxia in the rat. AB - Perinatal hypoxic-ischemic states can cause irreversible damage to the brain, ranging from minimal brain dysfunction to death. Only few studies have been reported describing neurological, cognitive and behavioral deficits following perinatal asphyxia. We therefore decided to study long term effects of perinatal asphyxia in a well-documented animal model resembling the clinical situation. Caeserean section in rats was performed and the pups, still in the uterus horns, were placed into a water bath at 37 degrees C for periods of 5-20 min; pups were then given to surrogate mothers and examined at three month of age. Examinations consisted of a battery of motor and reflex tests, Morris water maze, multiple T maze, elevated plus maze and open field studies. No abnormalities were found in rats even with long periods of perinatal asphyxia by neurological examination, in the open field and in mazes. Interestingly, in the elevated plus maze rats with long lasting exposure to hypoxia (15 and 20 min of asphyxia) showed reduced anxiety-related behavior. This finding may be relevant for the explanation of anxiety related disorders in adulthood with a tentative history in the perinatal period. PMID- 10714896 TI - Paracrine effects of angiotensin-converting-enzyme- and angiotensin-II-receptor- inhibition on transcapillary glucose transport in humans. AB - The paracrine renin-angiotensin-system (RAS) is increasingly recognized to play an important role in the regulation of both, regional vascular tone and regional glucose metabolism. To date, however, a selective investigation of paracrine RAS effects in an in vivo clinical setting was beyond technical reach. We here set out to selectively study the metabolic effects of paracrine RAS inhibition at different levels in healthy volunteers (n = 8). For this purpose bradykinin, enalaprilate and losartan were administered locally to the interstitial space fluid in skeletal muscle by means of reverse microdialysis and transcapillary glucose transport was measured simultaneously. During reverse microdialysis with bradykinin and enalaprilate a significant decrease in arterial-interstitial gradient for glucose (AIG(glu)) was observed (from 1.49 +/- 0.08 mM to 0.12 +/- 0.63 mM (p = 0.018) for bradykinin and from 1.5 +/- 0.07 mM to 0.24 +/- 0.67 mM (p = 0.043) for enalaprilate). In contrast, losartan had no effect on AIG(glu). The changes in transcapillary glucose transport during bradykinin and enalaprilate administration were accompanied by significant increases in interstitial lactate levels which was most pronounced for bradykinin (from 0.14 +/- 0.01 mM to 0.40 +/- 0.07 mM, p = 0.018). We conclude that paracrine angiotensin-converting-enzyme (ACE) inhibition but not angiotensin II (AT-II) receptor blockade decreases AIG(glu) and facilitates transcapillary glucose transport due to an increase in interstitial bradykinin concentration. These results support the concept that blood pressure control with ACE-inhibitors but not with AT-II-receptor-antagonists has beneficial long term metabolic consequences in hypertensive, hyperinsulinemic subjects. PMID- 10714897 TI - Highlights of marine natural products chemistry (1972-1999). AB - Marine Natural Products Chemistry is essentially a child of the 1970's that developed rapidly during the 1980's and matured in the last decade. With a few notable exceptions, it is difficult to select individual papers that significantly impacted the field. However, marine natural products chemistry has often influenced other fields and that aspect is the focus of this review. It is clear that the early directions taken by marine natural products chemists drew as much from the examples provided by insect chemical ecology as from the longer history of phytochemistry. By 1975 there were already three parallel tracks in marine natural products chemistry: marine toxins, marine biomedicinals and marine chemical ecology. It is the integration of the three fields of study that has given marine natural products chemistry its unique character and vigour. PMID- 10714898 TI - Marine natural products. PMID- 10714899 TI - Amaryllidaceae, muscarine, imidazole, oxazole, thiazole and peptide alkaloids, and other miscellaneous alkaloids. PMID- 10714900 TI - Lysine biosynthesis and metabolism in fungi. PMID- 10714901 TI - Iron chelators from mycobacteria (1954-1999) and potential therapeutic applications. PMID- 10714902 TI - Recent progress in the chemistry of the Stemona alkaloids. PMID- 10714903 TI - Estrogen with interrupted progestin HRT: a review of experimental and clinical studies. AB - This review outlines the basic principles of a novel interrupted progestin HRT regimen in which estrogen is administered continuously, and progestin is given in a 3-days on, 3-days off pulsed fashion. The rationale for this regimen is to prevent receptor down-regulation and allow increased estrogen and progestin sensitivity during the progestin-free periods. Background information is provided including the reasons for poor patient acceptance of HRT, and the concerns of the potential association of HRT with breast and endometrial cancer. Experimental studies in the rat are described which provide evidence in support of the rationale for the interrupted progestin regimen. Clinically, two pilot studies examining symptom control, bleeding rates and safety of the interrupted progestin regimen, as well as preliminary results of a third study examining the usefulness of this regimen for addback therapy in GnRH agonist treated patients, are outlined. The preliminary results of phase III trials are presented. These clinical studies all demonstrated good symptom control, low bleeding rates, endometrial protection, and excellent patient acceptance. The combination of continuous estrogen with interrupted progestin appears to result in increased sensitivity to estrogen and progestin in estrogen responsive tissues. As a result, lower doses of estrogen and progestin may be used for HRT with good biological effects. Further clinical studies, preferably in prospective randomized trials, are required to demonstrate an advantage of this new regimen compared to continuous combined HRT. PMID- 10714904 TI - Commentary on the Women's Health Initiative. AB - The Women's Health Initiative (WHI), established by the National Institutes of Health in 1991, is a long-term national health study that focuses on strategies for preventing heart disease, breast and colorectal cancer and osteoporosis in postmenopausal women. These chronic diseases are the major causes of death, disability and frailty in older women of all races and socioeconomic backgrounds. The WHI a 15-year multi-million dollar endeavor, and one of the largest U.S. prevention studies of its kind. The study involves over 161,000 women aged 50-79, and is one of the most definitive, far reaching clinical trials of women's health ever undertaken in the U.S. The WHI Clinical Trial and Observational Study will attempt to address many of the inequities in women's health research and provide practical information to women and their physicians about hormone replacement therapy, dietary patterns and calcium/vitamin D supplements, and their effects on the prevention of heart disease, cancer and osteoporosis. Emerging information from the NIH Women's Health Initiative and other studies of women's health begun in the 1990's should be changing the landscape of options for older women in the years to come. PMID- 10714905 TI - Hormonal substitution during menopause: what are we treating? AB - OBJECTIVES: It is suggested that during menopausal transition, women with vasomotor symptoms benefit from HRT, (hormone replacement therapy) whereas, the use of HRT for other cognitive-vegetative symptoms is questionable. METHODS: The occurrence of menopausal complaints and depressive symptoms was assessed cross sectionally in 5896 Dutch Caucasian women (47-54 years) of a large community sample in the city of Eindhoven, The Netherlands. Menopausal complaints were assessed using a 22 items self-rating scale (consisting of a vasomotor, uro genital and a cognitive-vegetative subscale). Depressive symptoms were assessed using the Edinburgh depression scale (EDS). Differences in mean scores were analysed between groups using ANOVA. The independent relationship of depressive symptoms to the intensity of menopausal complaints was assessed, by multiple linear regression analysis. RESULTS: Women using HRT showed the highest scores on all subscales. Oral contraceptive users had significantly lower scores on the vasomotor subscale compared to HRT users and to non users. Depressive symptoms contributed the most, to the explained variance on scores on the menopausal subscales. CONCLUSIONS: Women during menopause presenting several complaints, other than vasomotor origin might be suffering from underlying depression which makes it questionable to prescribe HRT for the latter symptoms. PMID- 10714906 TI - Determinants of age at menopause in Italy: results from a large cross-sectional study. ICARUS Study Group. Italian Climacteric Research Group Study. AB - OBJECTIVE: To identify the determinants of age at menopause in an Italian population, using data from the Italian Climacteric Research Group Study (ICARUS). METHODS: ICARUS is a prospective study of the effect of menopause on women's health that has been running in menopause clinics throughout Italy since 1995. A total of 4300 women with spontaneous menopause, aged 55 years or more and observed for the first time at the participating centres are included in the present analysis. RESULTS: The mean age at menopause in the total population was 50.9 years. After taking into account potential covariates, the women reported smoking, had a slightly lower mean age at menopause than non smokers 50.4 versus 50.9 years; P = 0.01. The mean age at menopause in nulliparae was 50.0 years, and, respectively 50.4, 50.6, 50.9, 51.2 and 50.9 years in those reporting 1, 2, 3, 4 and 5 or more births (P < 0.01). A low body mass index and an early age at menarche were associated with early menopause in the crude analysis, but these associations disappeared after taking into account the confounding factors. CONCLUSIONS: This study offers an estimate of the mean age at menopause of women attending menopause clinics in Italy, on the basis of the data obtained from a large sample. It also indicates that smoking and nulliparity are associated with early menopause. PMID- 10714907 TI - A longitudinal cohort study of elderly women with urinary tract infections. AB - AIMS: the prevalence of urinary tract infections (UTI), urinary incontinence (UI), estrogen-use and overall mortality in a cohort of elderly women who had been treated for UTI in 1985-86 was re-assessed 10 years later. MATERIAL AND METHODS: a random sample of 6000 women from the birth cohorts 1900, 1905, 1910, 1915 and 1920 were invited in 1986 to complete a questionnaire about UTI, UI and estrogen use (response rate 70%; n = 4206). Treatment with antibiotics for UTI during 1985-86 was reported by 688 (17%) women. In 1995 a similar questionnaire was sent to the women from this group who were still alive (n = 434). Mortality in the women with a history of UTI was compared with an aged-matched control group of women who did not have UTI during 1985-86. RESULTS: the questionnaire was completed and returned by 361 (83%) women. Treatment for at least one UTI during the last 9 years was reported by 219 (61%) women. The number of episodes varied: 35% had one to two UTI, 28% had three to four UTI, 27% five to ten UTI and 10% had had more than 10 UTI. In 1986, the prevalence of UI was higher in women with a history of UTI than in the total population sample (30 vs. 17%; P < 0.001). The prevalence of UI had increased from 30% in 1986 to 33% in 1995 (P < 0.05). Mortality in the women with a history of UTI was higher than in the aged matched control group (37 vs. 28%; P < 0.001). A total of 162 (45%) women had received estrogen therapy at some time after the age of 60 years and 140 (39%) reported that they were currently taking low potency estrogens. CONCLUSION: elderly women with a history of UTI had a continued high occurrence of UTI and UI, and overall mortality was higher in these women than in an age-matched control group of women from the total population. PMID- 10714908 TI - A double-blind, randomized, comparative study evaluating clinical effects of two sequential estradiol-progestogen combinations containing either desogestrel or medroxyprogesterone acetate in climacteric women. AB - OBJECTIVES: The aim of this study was to compare a new sequential estradiol desogestrel (E2-DSG) hormone replacement regimen (Liseta) with one of the standard treatments i.e. estradiol valerate-medroxyprogesterone acetate (E2V-MPA) combination (Klimalet) regarding the alleviation of climacteric symptoms, vaginal bleeding pattern and the occurrence of adverse experiences. METHODS: In a multicenter study performed in Denmark, a total of 376 perimenopausal women with climacteric symptoms were randomly allocated to oral sequential treatment with either E2-DSG (1.5 mg E2 for 24 days with 0.15 mg DSG for the last 12 days followed by a placebo tablet for 4 days) (n = 186) or with E2V-MPA (2 mg E2V for 21 days with 10 mg MPA for the last 10 days) (n = 190). Treatments were administered, using a double-blind, double-dummy technique for 6 cycles of 28 days. RESULTS: Three hundred and seventeen women, 158 in the E2-DSG and 159 in the E2V-MPA group, completed six treatment cycles. Both treatments reduced menopausal symptoms rapidly and to a similar extent. Hot flushes were present in 88% of the women in both groups. After six treatment cycles, hot flushes were no longer present in 71 and 62% of the women in the E2-DSG and E2V-MPA group, respectively. Perspiration decreased from 80 to 65% in the E2-DSG group and from 82 to 63% in the E2V-MPA group. Mood disturbances were present in 82% of the women in the E2-DSG at baseline, and in 52% after six cycles. In the E2V-MPA group the corresponding figures were 68 and 42%, respectively. The bleeding pattern was comparable in both treatment groups. Regular withdrawal (expected) bleeding appeared in 90-92% and in 85-90% of the women in cycles 1-5 with E2-DSG and E2V-MPA, respectively. Irregular bleeding (including spotting) occurred in 15.2% of the women receiving E2-DSG and in 20.1% of the women treated with E2V MPA in cycle 6. In both treatment groups there was a tendency of a slight decrease in blood pressure. Adverse events were in less than 10% in each group the reason to discontinue treatment. CONCLUSIONS: Both treatments effectively alleviated menopausal complaints and presented good cycle control. Bleeding pattern and mood disturbances appeared to be more favorable influenced by E2-DSG. PMID- 10714909 TI - Efficacy and tolerability of a new 7-day transdermal estradiol patch versus placebo in hysterectomized women with postmenopausal complaints. AB - OBJECTIVES: To investigate the efficacy and tolerability of a continuously applied 7-day-Estradiol patch (Fem7, Merck KGaA, Germany) delivering 50 microg estradiol per day in the treatment of hysterectomized women with postmenopausal complaints compared with placebo. DESIGN: A multicentre, randomized, double-blind study with an initial screening phase (phase I), a 3-month double-blind placebo controlled phase (phase II) and a 3-month open follow-up phase (phase III). METHODS: 186 patients were randomized for a 3-cycle placebo-controlled study followed by a 3-cycle open follow-up (total duration; 6 months). The changes in Kupperman Index (primary efficacy variable), hot flushes and urogenital symptom score were studied from baseline to the end of the study. In addition, skin tolerability was assessed and patients were also asked to grade the subjective acceptance of therapy. RESULTS: A reduction in Kupperman Index was observed in both groups, and at each cycle of the placebo-controlled treatment phase the 7 day-Estradiol patch was superior compared with placebo (last value vs. baseline P = 0.0006). From the second treatment week onwards a distinct difference was noted in the reduction of hot flushes from baseline between the 7-day-Estradiol patch group and the placebo group. The difference between the groups was statistically significant for each cycle and at the end of the controlled treatment phase (mean weekly hot flush reduction at the end of the placebo-controlled treatment phase: 32.5 for the 7-day-Estradiol patch vs. -22.0 for placebo, P = 0.0025). The efficacy of the 7-day-Estradiol patch within the application period did not show any difference between days 1-3 and 4-7. Subjective acceptance of the 7-day Estradiol patch was good and 72.4% of patients who took active medication throughout the study were willing to consider continuing its use. CONCLUSIONS: The 7-day-Estradiol patch is well tolerated and provides effective relief of moderate to severe vasomotor symptoms in hysterectomized women, with a rapid onset of action and 7-day duration of therapeutic effect. Although a placebo effect was observed, the 7-day-Estradiol patch significantly reduced hot flushes and other menopausal symptoms throughout the application period. PMID- 10714910 TI - Can endometrial protection be inferred from the bleeding pattern on combined cyclical hormone replacement therapy. AB - OBJECTIVE: To investigate the relationship between the timing of withdrawal bleeding on hormone replacement therapy and the state of the endometrium. DESIGN: Double-blind, prospectively randomised dose-ranging study. SETTING: Menopause clinics in the UK and the Netherlands. SUBJECTS: Two hundred and seventy one postmenopausal women aged 40-60. INTERVENTIONS: Administration of six 28-day treatment cycles of a continuous daily dose of 2 mg of micronised 17beta oestradiol with a randomly allocated dose of 5-20 mg of dydrogesterone added for the last 14 days of each. METHODS: Comparison of the timing of the withdrawal bleed recorded in subject-held diaries with an endometrial biopsy obtained toward the end of the last cycle. RESULTS: There was a trend towards later withdrawal bleeding with secretory endometrium and earlier bleeding with inactive or atrophic endometrium, but with too much overlap for this to be of clinical relevance. There were two cases of proliferative and one of hyperplastic endometrium, with no characteristic bleeding pattern. CONCLUSION: Combined sequential HRT with progestogen given for 12-14 days very rarely fails to protect the endometrium. Such failures can not be detected by noting the bleeding pattern. The only suspicious pattern is non-cyclic bleeding, but this will not detect every case of hyperplasia or persistent proliferative endometrium. PMID- 10714911 TI - Comparative effects of estrogens plus androgens and tibolone on bone, lipid pattern and sexuality in postmenopausal women. AB - BACKGROUND: The main goals of estrogen replacement therapy (ERT) are the prevention of osteoporosis and cardioprotection and the improvement of quality of life (QL). Androgens and tibolone therapy may increase bone mineral density (BMD) to a greater extent than ERT and offer an increase in QL. Lipid and cardiovascular effects, however, are still a major concern. AIM: To evaluate whether the addition of a weak androgen to ERT may improve postmenopausal bone loss and sexual activity without adverse effects on lipid pattern and to compare these effects with those observed after tibolone therapy. SUBJECTS AND METHODS: This prospective study enrolled 120 surgical postmenopausal women; of these, 96 completed the 1-year follow-up. Patients were allocated to one of four groups. The first group (A; n = 23) received 4 mg of estradiol valerate plus 200 mg of enanthate of dihydroandrosterone im monthly. The second group (E; n = 26) received 50 microg/day of transdermal 17-b-estradiol continuously; the third (T; n = 23) received 2.5 mg of tibolone every day; and finally, the fourth group (C; n = 24) constituted a treatment-free control group. Bone mass (dual X-ray absorptiometry), serum total cholesterol, HDL, LDL, triglycerides, apolipoproteins A1 and B and sexual activity were evaluated before starting therapy and at the end of follow-up. RESULTS: All active treatment groups showed an increase in BMD. This increase was higher in the A treatment group (4.08% P < 0.01). Sexuality improved significantly with therapy; however, tibolone and androgens increased scores to a greater extent than ERT. Androgen therapy was associated with significant increases in total cholesterol, LDL and triglycerides. Cholesterol and LDL fall into groups E and T, HDL into groups A and T and triglycerides in group T only. CONCLUSION: The combined regimen of androgens and ERT increased vertebral bone mass and enhance sexual activity in postmenopausal women equal to that of tibolone and to a greater extent than ERT alone; its effects on lipids, however, are clearly adverse. PMID- 10714912 TI - Efficacy and safety of oral estriol for managing postmenopausal symptoms. AB - OBJECTIVE: to assess the therapeutic efficacy and safety of oral estriol for the treatment of climacteric symptoms in postmenopausal women. METHODS: 68 postmenopausal women with climacteric symptoms received oral estriol, 2 mg/day, daily for 12 months. We evaluated the degree of climacteric complaints with estriol therapy; serum levels of gonadotropins, estradiol (E2) and lipids; biochemical markers of bone metabolism; blood pressure; and side effects both at baseline and during treatment. Climacteric symptoms were assessed according to the menopausal index (MI), a version of the Kupperman index that had been modified for Japanese women. RESULTS: oral estriol therapy significantly reduced total MI scores. The greatest relief was noted for hot flushes, night sweats, and insomnia. Estriol treatment significantly lowered serum follicle stimulating hormone (FSH) and luteinizing hormone (LH) concentrations but did not affect any of the other parameters (lipids, bone, liver and blood pressure) during the study period. Slightly vaginal bleeding occurred in 14.3% of those who underwent natural menopausal women. Histologic evaluation of the endometrium and ultrasound assessment of the breasts following 12 months of estriol treatment found normal results in all women. CONCLUSION: Estriol is a safe and effective alternative for relieving climacteric symptoms in postmenopausal Japanese women. PMID- 10714913 TI - Prevention of postmenopausal bone loss by low and conventional doses of calcitriol or conjugated equine estrogen. AB - OBJECTIVES: Estrogen deficiency is the most common cause of postmenopausal osteoporosis and estrogen replacement is well known to retard postmenopausal bone loss. Calcium supplement alone is generally considered to be insufficient for the prevention of bone loss associated with estrogen deficiency while the role of calcitriol is unclear. In the present study we examined the efficacy different doses of estrogen or calcitriol in the prevention of postmenopausal bone loss in Thais. METHODS: The subjects consisted of 146 Thai women no more than 6 years postmenopausal. The subjects were randomly allocated to receive 750 mg supplemental calcium alone, calcium and conjugated equine estrogen (CEE) at 0.3 or 0.625 mg, calcium and calcitriol at 0.25 or 0.5 microg daily. Those receiving CEE also took 5 mg medrogestone for 12 days each month. BMD at L2-4 and femoral neck were measured at baseline 1 year and 2 years after treatments. Data were expressed as mean +/- S.E. RESULTS: Subjects on supplemental calcium alone had approximately 2.5% decreases in L2-4 (P < 0.05) and femoral BMD (P < 0.01) at 2 years. CEE (0.3 mg) resulted in 3.20 +/- 1.2% increase in vertebral BMD (P < 0.05) while no significant change in BMD was demonstrated at the femoral neck. Likewise, 0.625 mg of CEE induced 5.4 +/- 1.4% increase in vertebral BMD at 2 years (P < 0.001) without change in the femoral BMD. In regard to calcitriol, no significant change in vertebral or femoral BMD was demonstrated with either 0.25 or 0.5 microg calcitriol. CONCLUSION: We concluded that calcitriol is effective in the prevention of early postmenopausal bone loss in Thais. It represents an option for the prevention of osteoporosis in postmenopausal women who are contraindicated for estrogen replacement. PMID- 10714914 TI - Evaluation of hormone replacement therapy use by the sales figures. AB - Despite the efficiency of hormone replacement therapy (HRT) to prevent climacteric manifestations and possibly the long-term deleterious influences of menopause, the prevalence of HRT is relatively low, and quite variable, depending on the population studied. Presently, there is no information regarding HRT in Switzerland and in the region of Geneva, which have particularly aged populations, with a life expectancy among the longest in the Western world. In this study, the number of women treated per year in 1993 and 1996, as well as the prevalence of HRT were estimated, based on the total amount of hormone preparations sold for HRT. In Switzerland, for a female population older than 45 years of about 1.45 million, the number of women on HRT was approximately 166,000 in 1993 and 202,000 in 1996. For Geneva, the female population was more than 86,000, and the number of treated women was about 14,000 and 21,000 in 1993 and 1996, respectively. Depending on the age class considered as susceptible of receiving HRT, the prevalence of this therapy may vary between 15 and 20% for Switzerland, and between 21 and 27% for Geneva in 1993. It was estimated between 17 and 24%, and 31 and 41% in 1996. These values are quite comparable to those reported for other countries with a similar socioeconomic level and obtained using different methods of evaluation. PMID- 10714915 TI - Geographic research at the end of the century: papers from the Eighth International Symposium on Medical Geography. PMID- 10714916 TI - Finke's 1792 map of human diseases: the first world disease map? AB - Documentary evidence reveals that a German physician L.L. Finke produced a world map of diseases in 1792. This is much earlier than any world disease map previously known. Contrary to the contemporary literature in medical cartography this data proves that: (1) It was neither yellow fever nor cholera epidemics but indigenous diseases that were the catalyst for this earlier world disease map. (2) It predates Humboldt's influence on thematic mapping. PMID- 10714917 TI - Our sense of Snow: the myth of John Snow in medical geography. AB - In 1854, Dr. John Snow identified the Broad Street pump as the source of an intense cholera outbreak by plotting the location of cholera deaths on a dot-map. He had the pump handle removed and the outbreak ended...or so one version of the story goes. In medical geography, the story of Snow and the Broad Street cholera outbreak is a common example of the discipline in action. While authors in other health-related disciplines focus on Snow's "shoe-leather epidemiology", his development of a water-borne theory of cholera transmission, and/or his pioneering role in anaesthesia, it is the dot-map that makes him a hero in medical geography. The story forms part of our disciplinary identity. Geographers have helped to shape the Snow narrative: the map has become part of the myth. Many of the published accounts of Snow are accompanied by versions of the map, but which map did Snow use? What happens to the meaning of our story when the determinative use of the map is challenged? In his book On the Mode of Communication of Cholera (2nd ed., John Churchill, London, 1855), Snow did not write that he used a map to identify the source of the outbreak. The map that accompanies his text shows cholera deaths in Golden Square (the subdistrict of London's Soho district where the outbreak occurred) from August 19 to September 30, a period much longer than the intense outbreak. What happens to the meaning of the myth when the causal connection between the pump's disengagement and the end of the outbreak is examined? Snow's data and text do not support this link but show that the number of cholera deaths was abating before the handle was removed. With the drama of the pump handle being questioned and the map, our artifact, occupying a more illustrative than central role, what is our sense of Snow? PMID- 10714918 TI - Geography, ecology and emerging infectious diseases. AB - Emerging infectious diseases are the focus of increased attention and even alarm in the scholarly and popular literature. The emergence of new diseases and the resurgence of older and previously recognized infectious diseases both in developing and developed country poses challenges for understanding the ecological web of causation, including social, economic, environmental and biological components. This paper is a synthesis of the major characteristics of emerging diseases, in an interdisciplinary context. Political ecology is one framework for analysis that is promising in developing a modified ecology of disease. PMID- 10714919 TI - Ecological and cultural barriers to treatment of childhood diarrhea in riverine areas of Ondo State, Nigeria. AB - In Nigeria diarrhea still poses the greatest health problem to the survival of the under-fives in spite of the fact that the majority of mothers are reportedly to have been reached by health education on oral rehydration therapy (ORT) regardless of their ecological and socioeconomic situations. This study assesses the effect of different ecological and sociocultural conditions on use of ORT in riverine areas of Ondo State for the identification of the most effective means of dissemination of information on ORT in similar geographically disadvantaged localities in Nigeria and elsewhere. It is a formative study, but its results are expected to lead to identification of potentially effective intervention modalities to improve diarrhea treatment in remote areas. Of great concern in this study are communities whose awareness and acceptance of ORT may be more dictated by environmental conditions. This study combines two different research methodologies; namely, semistructured questionnaires and in-depth interviews to gain 'focused' insight into the communities. The study was carried out in Ilaje Ese-Odo local government area (LGA) in southwestern Nigeria. It covered 308 mothers from 2 subethnic groups (Ilaje and Apoi) from a set of randomly chosen villages situated in 3 ecological strata and the in-depth interviews with 42 key informants. The majority of the mothers described some dangerous signs of last diarrhea suffered by their children under the age of five. Preliminary results indicate that awareness of actual causation was lowest in the remotest saltwater areas compared with other mothers in the study communities. Sixty-eight percent of the mothers in Ilaje mainland, 57% in fresh water Apoi and 44% in saltwater Ilaje stated that they have ever heard of salt, sugar solution (SSS). Furthermore, only 43% of them said they could prepare SSS while 42% ever made it. None of those mothers in saltwater Ilaje who confirmed awareness of and how to administer and prepare SSS could in reality do it correctly. When shown a typical sachet of ORS, only 8% of mothers living in salt water swamps said they have seen one before. Thus, promotive health services to reduce high mortality rates of children under five have passed the study mothers whose area reported the highest incidence of diarrhea in the state. PMID- 10714920 TI - Health and disease in southern Africa: a comparative and vulnerability perspective. AB - Using a vulnerability and comparative perspective, this paper examines the status of health in southern Africa highlighting the disease complex and some of the factors for the deteriorating health conditions. It is argued that aggregate social and health care indicators for the region such as life expectancy and infant mortality rates often mask regional variations and intra-country inequalities. Furthermore, the optimistic projections of a decade ago about dramatic increases in life expectancy and declines in infant mortality rates seem to have been completely out of line given the current and anticipated devastating effects of the HIV/AIDS pandemic in southern Africa. The central argument is that countries experiencing political and/or economic instability have been more vulnerable to the spread of diseases such HIV/AIDS and the collapse of their health care systems. Similarly, vulnerable social groups such as commercial sex workers and women have been hit hardest by the deteriorating health care conditions and the spread of HIV/AIDS. The paper offers a detailed discussion of several interrelated themes which, through the lense of vulnerability theory, examine the deteriorating health care conditions, disease and mortality, the AIDS/HIV situation and the role of structural adjustment in the provision of health care. The paper concludes by noting that the key to a more equitable and healthy future seems to lie squarely with increased levels of gender empowerment. PMID- 10714921 TI - Using survival analysis to study spatial point patterns in geographical epidemiology. AB - The spatial K-function has become a well accepted method of investigating whether significant clustering can be detected in spatial point patterns. Unlike nearest neighbor-based methods, the K-function approach has the advantage of exploring spatial pattern across a range of spatial scales. However, K-functions still have a number of drawbacks. For instance, although K-functions are based on inter event distances, they only use a count of the number of point events within successive distance bands. This represents data aggregation and information loss. Secondly, and perhaps more significantly, K-functions are based on a cumulative count of point events with distance. This feature raises the possibility that the investigation of pattern at different scales is compromised by the dependency of any one count to previous counts. This paper proposes a new approach to the analysis of spatial point patterns based upon survival analysis. Although typically used in the temporal domain, there is no reason why survival analysis cannot be applied to any positively-valued, continuous variable as well as time. In this paper, survival analysis is applied to the inter-event distance measures of bivariate spatial point patterns to investigate the 'random labeling' hypothesis. It is shown, through both a controlled data situation and empirical epidemiological applications, that such an approach may be a very useful complement to K-function analysis. PMID- 10714922 TI - Approaches to sampling and case selection in qualitative research: examples in the geography of health. AB - This paper focuses on the question of sampling (or selection of cases) in qualitative research. Although the literature includes some very useful discussions of qualitative sampling strategies, the question of sampling often seems to receive less attention in methodological discussion than questions of how data is collected or is analysed. Decisions about sampling are likely to be important in many qualitative studies (although it may not be an issue in some research). There are varying accounts of the principles applicable to sampling or case selection. Those who espouse 'theoretical sampling', based on a 'grounded theory' approach, are in some ways opposed to those who promote forms of 'purposive sampling' suitable for research informed by an existing body of social theory. Diversity also results from the many different methods for drawing purposive samples which are applicable to qualitative research. We explore the value of a framework suggested by Miles and Huberman [Miles, M., Huberman,, A., 1994. Qualitative Data Analysis, Sage, London.], to evaluate the sampling strategies employed in three examples of research by the authors. Our examples comprise three studies which respectively involve selection of: 'healing places'; rural places which incorporated national anti-malarial policies; young male interviewees, identified as either chronically ill or disabled. The examples are used to show how in these three studies the (sometimes conflicting) requirements of the different criteria were resolved, as well as the potential and constraints placed on the research by the selection decisions which were made. We also consider how far the criteria Miles and Huberman suggest seem helpful for planning 'sample' selection in qualitative research. PMID- 10714924 TI - Ageing in Portugal: regional iniquities in health and health care. AB - The health of the Portuguese has improved considerably in the last twenty years. Economic and social transformations that have contributed to the progressive amelioration of problems of feeding, sanitation, hygiene, housing and social conditions in general, as well as health services, have had decisive effect on this phenomenon. The spectacular regression of the indicators related to transmitted diseases, infant, perinatal (more than 50% between 1985 and 1994) and maternal mortality, and the mortality of children 1 to 4 yr old, also reflects this impact. The positive changes that took place in health indicators were reflected in the growth of life expectancy at birth (2.2 yr more for male and 2.3 more for women between 1985 and 1994) in spite of the fact that the difference in life expectancy in relation to EU countries has grown. Improvement in life expectancy, especially in the older age groups, is not normally associated with significant reductions in morbidity. In fact, increased longevity has become more generally associated with chronic illness or other disabilities requiring more medical services and other forms of personal care. This paper reviews some of the evidence for regional differences in the health status of elderly people in Portugal and considers how health services have reacted to these differences. A preliminary study of health status and patterns of utilisation of elderly people was undertaken. After 30 yr of a National Health Service (NHS) in Portugal we may ask why do inequities in health and access to health care of the elderly population persist? Proactive policies to prevent illness and promote health are still relatively underdeveloped in the Portuguese NHS, and the factors that influence health, such as housing, diet and occupational health hazards, remain largely absent from health and welfare policies. Poor accessibility to health services is the most serious barrier consumers have to face in order to get a medical appointment, and this is more relevant to the oldest part of the population. Geographical location of health care facilities unequally affects the ease of access of different groups of consumers and influences utilisation patterns. Examining the distribution of health services resources is an important way to understand the inequities of access to health and to health care. PMID- 10714923 TI - Regional assessment of elderly disability in the U.S. AB - This study examines regional variation of elderly disability in the United States. Elderly disability measurements are derived from two newly available questions on mobility and self-care limitations in the 1990 census. Substantial regional differences in elderly disability rates exist, with a higher prevalence of disability in the Southeast. These differences persist after controlling for age and socioeconomic status (SES). The study findings suggest that some public health policy should be regionally formulated and some government actions should be devoted to reduce the excessive elderly disability in the South. PMID- 10714925 TI - Long-term care restructuring in rural Ontario: retrieving community service user and provider narratives. AB - This paper examines the extensive restructuring of community-based long-term care that was initiated in Ontario, Canada in 1996, and does so with particular reference to longstanding problems of provision in rural communities. Specifically, it draws on a case study focussed on two small rural towns to develop a 'situated understanding' of service-user and service-provider perspectives on service coordination issues and on service cuts, particularly as they affect the ability of elderly people reliant on publicly-funded community services to stay in their homes, to continue to 'age in place'. The general and specific antecedents of long-term care reform are considered prior to the presentation of the case study. General antecedents include the rapid aging of Canada's population and aggressive strategies to reduce government deficits, while specific antecedents flow from a decade of failed attempts to address longstanding issues of service coordination and from the ideologically-driven, free market stance of the provincial government elected in 1995. The analysis of interviews conducted with 14 community-service users and 17 providers suggests that the managed competition system introduced as the centerpiece of long-term care reform has resulted in increasing diversity and uncertainty on both sides of the service provision equation. Despite continued attempts by rural elderly people and their families to 'cut and paste' support packages, it seems that the restructuring of publicly-funded community services, combined with a substantial re-investment in long-term care facilities, will make some elderly people more vulnerable to institutionalization. PMID- 10714926 TI - Widening inequality in mortality between 160 regions of 15 European countries in the early 1990s. AB - This paper presents maps of geographical patterns in mortality for the 160 mainland regions of the 15 countries of the European Union. Standardised mortality ratios (SMRs) for all ages are presented for all causes of death and for lung cancer, ischaemic heart disease, road traffic accidents and suicide. All cause standardised mortality ratios (for deaths under the age of 65) for the years 1990 and 1994 are presented. These data show that while most regions of Europe had decreasing SMRs over this time period, SMRs increased for the 10% of the population with the highest SMRs and the gap between the most and least healthy regions grew. Possible reasons for the observed patterns, the limitations of currently available data and the limitations of studying nation states, are suggested. PMID- 10714927 TI - Death and deprivation: an exploratory analysis of deaths in the health and lifestyle survey. AB - An analysis is undertaken of deaths of respondents in the UK- representative Health and Lifestyle Survey. The sample was originally interviewed in 1984/5 and followed initially until May 1997. Using multilevel logistic and Cox-proportional hazards models, the relationships between death and a wide range of social circumstances and behaviours is explored. It is found that place deprivation interacts with individual social class in accounting for variations in mortality. This is the case even when account is taken of personal health-related behaviour. There appears to be some evidence of a threshold relationship such that the differential effects of social class are only found at high-levels of deprivation. No statistically significant interactions are found for social and behavioural variables, for behavioural and place deprivation variables, and for social and place deprivation variables with the exception of social class. The study is deliberately exploratory and a wide range of models have been fitted which will be subject to more rigorous evaluation as the HALS death study proceeds. PMID- 10714929 TI - Etiology of limited transmission diseases among drug users: does recent migration magnify the risk of sharing injection equipment? AB - Epidemiological studies have attributed area-specific changes in infectious disease prevalence to human migration. There is a paucity of research investigating the postmigration adjustment period as an effect on risk behaviors that are required in limited transmission diseases. A two-group typology, derived by cluster analysis, allowed for an analytical differentiation in the postmigration period. The cluster variable and other possible cofactors were included in linear and logistic regression modeling of sharing drug injection equipment among drug users in Alaska. The results indicate that among participants who have injected drugs, those in the postmigration adjustment period are nearly six times more likely to share injection equipment than those drug users who are not in a postmigration period. Further research is suggested and limitations are discussed. PMID- 10714928 TI - Problems of providing health care in British island communities. AB - Issues and problems relating to the funding and provision of health services to islands are examined with particular reference to the island communities of Britain. The results of a telephone survey of key representatives of relevant health authorities in the British Isles indicate the general problems faced by health care providers in island localities. The Isle of Wight is used as a case study and exemplar of the specific difficulties faced in island settings. The paper concludes by examining the relevance of current resource allocation policy to island contexts. PMID- 10714930 TI - The geography of survival after surgery for colo-rectal cancer in southern England. AB - This study investigates variations in survival following surgery for colo-rectal cancer in the Wessex region (part of southern England), using 5147 cases diagnosed between 1 September 1991 and 31 August 1995. Survival curve estimation by life tables and Cox's proportional hazards model were used to examine geographical variation in cancer survival, with a specific focus on distance between place of residence and treatment centre, and district of treatment. We also consider whether area deprivation has an impact on survival. In seeking to answer these questions we control for possible confounders, including: age, gender, site of tumour, stage of disease at operation, hospital size and surgery type (whether elective or non-elective). District of treatment, distance and deprivation all show a relationship to outcome using survival curves, but when adjusting for other covariates using the Cox model, and considering deaths from all causes, only district of treatment was a very significant covariate (p < 0.0001). Distance, deprivation, and gender were only weakly significant (p < 0.10). Considering only deaths related to operation (within 30 days) district of treatment remained significant, but while distance had some effect on outcome, deprivation and gender ceased to be significant covariates. There is some evidence that those who live furthest from centres of treatment have the worst outcomes but the 'geography of survival' manifests itself more through where patients are treated than through area (deprivation) effects or relative location. The results have important policy implications, as they show variations among treatment centres having controlled for potentially confounding factors. PMID- 10714931 TI - Predicting small-area health-related behaviour: a comparison of smoking and drinking indicators. AB - Health-related behaviours are of central importance to health promotion and to the promotion of enhanced population health. In the UK, localised knowledge of the quantitative dimensions of health-related behaviours is traditionally attained by conducting a costly sample survey. Such surveys seldom generate reliable data at scales more local than that of the health authority, they also need to be repeated regularly. This paper outlines an alternative framework for generating statistics on small-area health related behaviours using routinely available data from the annual Health Survey for England (N = 17,000) and the decennial Population Census. Using a multilevel modelling approach nesting individuals within postcode sectors within health authorities, and focusing on the prevalence of smoking and 'problem' drinking, the paper comprises four sections: a consideration of the modelling strategy, a comparison of the smoking and drinking models, an outline of the estimation strategy, and the presentation and discussion of ward-level estimates of smoking and drinking behaviour for England. The paper concludes that the method is better at estimating smoking than drinking but that it offers a feasible, cheap and more informative alternative to the survey approach to the generation of information on smoking and drinking behaviour. PMID- 10714932 TI - Modelling exposure opportunities: estimating relative risk for motor neurone disease in Finland. AB - This paper addresses the issues surrounding an individual's exposure to potential environmental risk factors, which can be implicated in the aetiology of a disease. We hope to further elucidate the 'lag' or latency period between the initial exposure to potential pathogens and the physical emergence of the disease, with specific reference to the rare neurological condition, motor neurone disease (MND), using a dataset obtained from the Finnish Death Certificate registry, for MND deaths between the period 1985-1995. A space-time approach is adopted, whereby patterns in both time and space are considered. No prior assumptions about the aetiology of MND are adopted. By using methods for the analysis of point processes, which preserve the continuous nature of the data, we resolve some of the problems of analysis that are often based on arbitrary areal units, such as postcode boundaries, or political boundaries. We use kernel estimation to model space-time patterns. Raised relative risk is assessed by adopting appropriate adjustments for the underlying population at risk, with the use of controls. Significance of the results is assessed using Monte Carlo simulation, and comparisons are made with results obtained from Openshaw's geographical analysis machine (GAM). Our results demonstrate the utility of kernel estimation as a visualisation tool. Small areas of elevated risk are identified, which need to be more closely examined before any firm conclusions can be drawn. We highlight a number of issues concerning the inadequacies of the data, and possibly of the techniques themselves. PMID- 10714933 TI - Environmental risk perception and well-being: effects of the landfill siting process in two southern Ontario communities. AB - In the context of siting (environmentally) noxious land uses, recent research suggests that the well-being of individuals and communities is impacted as much by the decision-making process as the outcome itself. The study results presented in this paper stem from an ongoing, two-stage quantitative/qualitative investigation of impacts on individual and community well-being associated with the environmental assessment process in Ontario, Canada. This research uses a parallel case-study design to investigate two proposed landfill sites in southern Ontario. Qualitative in-depth interviews (n = 36), conducted across a variety of stakeholder groups, were used to address the following objectives: to explore the nature of concerns experienced by individuals faced with a local landfill site proposal; to explore the effects of the siting process on individuals and communities; and to examine the coping strategies employed by individuals in response to impacts experienced. The work attempts to apply theories of risk society (as conceptualised by Beck and Giddens) at a community scale. In so doing, we build on the work of health geographers attempting to link the social and contextual with the medical. Overall, substantial impacts on individual and community well-being were reported across all stakeholder groups interviewed: these included stress, disempowerment, hostility and divisions within the community. The experience of psychosocial impacts and effectiveness of coping strategies is shaped by certain factors associated with the site and the siting process (including uncertainty and the perceived lack of meaningful participation). The links between risk, process and impacts are theorized using a conceptual framework which incorporates site and process factors, effects on daily life (e.g. feelings of losing control, mistrust), and Gidden's conception of 'ontological security'. These findings have implications for environmental decision-making, as they suggest a need to locate the delicate balance point between community involvement and an expedient decision-making process within variable community contexts. PMID- 10714934 TI - A longitudinal study of the health impacts of a petroleum refinery. AB - Emissions from a petroleum refinery in Oakville, Ont., have been the source of longstanding health concerns among residents in the surrounding community. Between 1992 and 1997, the refinery implemented extensive odour reduction measures through improvements in waste water treatment, in sulphur recovery and combustion. In this paper, we present the main findings of a recent longitudinal analysis using data from community health surveys conducted in 1992 and 1997, before and after implementation of the odour reduction plan. The results show a decline in the frequency of odour perception and annoyance by residents whereas the reporting of cardinal and general symptoms among adults and children was virtually unchanged. Odour perception and annoyance were strongly related to symptom reporting in both years supporting the hypothesis that the effect of refinery emissions on residents' health is odour mediated. The findings extend our understanding of the psychosocial basis of symptom reporting in the vicinity of refineries. PMID- 10714935 TI - In vivo regulation of cerebral monoamine oxidase activity in senescent controls and chronically stressed mice by long-term treatment with Ginkgo biloba extract (EGb 761). AB - It is well recognized that Ginkgo biloba extract (EGb 761) exert beneficial effects against various age-related changes and is able to reduce the negative influence of stress. In view of the age-dependent increase in the activity of the B form of monoamine oxidase (MAO-B) and in view of the anti-stress action of EGb 761 hypothetically attributed to an inhibition of monoamine oxidase by this substance, we investigated the effects of long-term treatment with EGb 761 upon in vivo cerebral MAO-A and -B activities of stressed and unstressed 17- and 18 month-old mice. The stress was a 'chronic mild stress' regimen whose behavioral impact is known to be reduced by EGb 761. The results showed that: (1) EGb761 induced reductions in MAO activity in 18-month-old, but not in 17-month-old mice; the older animals having higher basal MAO activity; (2) in unstressed mice, EGb 761 appeared to reduce the age-induced increase in cerebral MAO activity; (3) MAO A and -B activities of stressed and treated 18-month-old mice did not differ significantly from the levels observed in unstressed and untreated 17-month-old mice. These results may shed light on the anti-stress effects of Ginkgo biloba extract. PMID- 10714936 TI - Ceramide induces expression of the senescence histochemical marker, beta galactosidase, in human fibroblasts. AB - We recently showed that ceramide is elevated in senescence and that when administered to low-passage cells induces biochemical changes characteristic of senescence. The in situ histochemical marker beta-galactosidase (beta-Gal) has provided an important tool in the study of cellular senescence. We investigated the ability of ceramide to induce the expression of beta-Gal and correlated this with cell proliferation. We find that D-e-C6-ceramide, induces the expression of acidic beta-Gal in fetal lung-derived Wi-38 human diploid fibroblasts. Our results show that this induction is: (1) time and concentration dependent; and (2) reversible upon ceramide removal. We also find that concomitant with the onset of beta-Gal staining, DNA synthesis is blocked. These conditions are reversible. The induction of beta-Gal expression is specific to C6-ceramide. We discuss a potential role of beta-Gal in the regulation of senescence. Although signal transduction of senescence is still not fully understood, this new evidence strengthens the hypothesis that ceramide plays a key role in signaling down stream biochemical changes in cellular senescence. PMID- 10714937 TI - Nitric oxide myotoxicity is age related. AB - Nitric oxide (NO) is generated under normal conditions in skeletal muscle and acts as a messenger that influences contractility, blood flow, and glucose metabolism. Excess NO generation may occur in pathological states, in particular inflammatory conditions. We demonstrate that incubation of rat extensor digitorum longus muscle with the NO donor, S-nitrosocysteine, leads to release of creatine kinase, a marker of muscle injury after a delay of 90 min. Muscle of old animals was more sensitive to the NO donor. Light microscopic analysis does not show abnormalities, with the exception of an increase in interfiber distance. Histological staining identified no pathological elevations of calcium. The study demonstrates the direct toxicity of NO to skeletal muscle, and that muscle of older animals is differentially susceptible to NO toxicity. PMID- 10714938 TI - Changes in glycosylation of rat liver arylsulfatase B in relation to age. AB - The aim of this study was to determine how glycosylation of the rat liver arylsulfatase B was influenced by the age of the animal. The enzyme was purified from a liver lysosomal fraction obtained from male Wistar rats aged 18 days of gestation, 1 week, and 1, 1.5, 3 and 18 months by an affinity chromatography. Examination of the carbohydrate structures was performed after electrophoresis and blotting, followed by a very sensitive detection system with a set of six highly specific digoxygenin-labelled lectins. After densitometric measurement of the intensity of a digoxigenin-labelled lectin binding to arylsulfatase B, it could be stated that, at least, changes in sialylation are related to the growth and development of rats. Sialylation increases while fucosylation slightly decreases with age of the animal. PMID- 10714939 TI - Growth factor receptor gene and protein expressions in the human lens. AB - In this study the mRNAs encoding epidermal growth factor receptor (EGFR), basic fibroblast growth factor receptor (FGFR-2) and insulin-like growth factor receptor (IGFR-1) genes of the human normal lenses at ages varying from 0.5 to 72 years, were identified by semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR). Regulation of EGFR gene expression in the lens did not change with aging, and of FGFR-2 and IGFR-1 genes also remained unaltered up to age 53 years. However, expressions of FGFR-2 and IGFR-1 genes were decreased at ages above 60 years. EGFR, FGFR-2 and IGFR-1 proteins were detected by immunoblot analysis in the epithelial cell membranes of lens at age varying from 40 to 72 years. There was no detectable amount of EGFR protein in fiber cell membranes of the lens, and the levels of FGFR-2 and IGFR-1 proteins were much lower than those in the epithelial cell membranes. The low levels of these receptor proteins in the fiber cell membranes of lens, suggest their possible role in keeping the differentiated function of these unique transparent cells. The findings of the increased protein levels with age of EGFR with the appearance of some degradation products at age 48 years and higher, and the increased FGFR-2 protein at age 60 years and higher in the epithelial cell membranes of lens, were of interest. It appears that this could be a compensatory protective response of the lens to aging process for lifelong continuation of normal growth by proliferation and differentiation of its epithelial cells into new fiber cells in the germinative zone at the equatorial region. Thus, these results could provide a basis for further studies on growth factor receptor gene and protein regulations in the mechanism of lens aging and progression of age-related human cataract. PMID- 10714940 TI - Lymphatic lipid transport is not impaired in ageing rat intestine. AB - Lymphatic lipid transport in the intestine of adult and ageing rats was compared. Adult (8-10 months old) and old (24-26 months old) male Wistar rats were cannulated into the mesenteric lymph under ethrane anesthesia. On the following day, lipid emulsion containing 35.4 mg/h of olive oil was infused intraduodenally for 7 h and lymph collected hourly was assayed for triglyceride and apolipoprotein A-IV (apo A-IV). The results showed there was no difference in lymphatic lipid and apo A-IV transport between adult and old rats. Since apo A-IV synthesis in the enterocytes is linked to the intracellular assembly of lipoprotein, it is likely that in addition to lymphatic transport, production of chylomicrons is not impaired in ageing rats. PMID- 10714941 TI - Insulin receptors in the brain cortex of aging mice. AB - The aim of the present study was to analyze whether aging also affects central insulin receptors in brain cortex as it does in whole brain of BALB/c-nu mice. Results showed statistically significant decrease of number and increase of affinity of insulin high affinity binding sites in old animals. As a consequence, central insulin actions, among which neuromodulation of monoaminergic system, can result altered during aging. PMID- 10714942 TI - Asthma mediators: current views. PMID- 10714943 TI - Brittle-ductile transitions in sucrose and the influence of lateral stresses during compaction. AB - Sucrose, in a range of particle sizes, has been compacted to investigate both the effect of brittle-ductile transition and the effect of lateral stresses on the deformation stress as measured using Heckel plots. All particles with a diameter greater than 30 microm exhibited cracking in line with both theoretical predictions and literature data from hammer and ball milling. In addition, crack lengths in compressed particles examined microscopically were very similar to those predicted from the deformation stress, confirming the applicability of the model. PMID- 10714944 TI - Controlled transdermal delivery of propranolol using HPMC matrices: design and in vitro and in-vivo evaluation. AB - To improve bioavailability and achieve a smoother plasma-concentration profile as compared with oral administration, a matrix-dispersion-type transdermal delivery system was designed and developed for propranolol using different ratios of hydroxypropylmethylcellulose (HPMC) K4M, K15M and K100M. Formulations were evaluated for in-vitro dissolution characteristics using a Cygnus' sandwich-patch holder. Drug release followed Higuchi rather than zero-order or first-order kinetics. In-vivo evaluation was carried out on healthy volunteers (21+/-1.41 years; 60.89+/-5.35 kg) following the balanced incomplete block design. The dissolution rate constant (k) and data generated from plasma and urine (Cmax, maximum plasma concentration; t(max), time to reach peak plasma concentration; AUC, area under the curve; k(e), elimination rate constant; t1/2e, elimination half-life; k(a), absorption rate constant; t1/2a, absorption half-life) were evaluated statistically by two-way analysis of variance. Statistically excellent correlation was found between the percentage of drug absorbed and Cmax, AUC0-24 and AUC0-infinity. A highly significant difference (P < 0.001) was observed when Cmax and AUC0-infinity, generated from plasma and urine were compared, but k(e), t1/2e, k(a) and t1/2a did not differ significantly (P > 0.1). We conclude that urinary excretion data may be used as a simpler alternative to blood level data in studying the kinetics of absorption and deriving the absorption parameters. PMID- 10714945 TI - Effects of some non-ionic surfactants on transepithelial permeability in Caco-2 cells. AB - The effects of the non-ionic surfactants polysorbate 20, polysorbate 60, polysorbate 85, cholesteryl poly (24) oxyethylene ether (Solulan C24) and the lanolin-based poly (16) oxyethylene ether (Solulan 16) on the epithelial integrity of monolayers of human intestinal epithelial (Caco-2) cells has been studied using metformin as a model drug. The aim was to identify the surfactants and their optimal concentrations capable of enhancing drug transport while causing no, or only minor, cellular damage. Effects on cell permeability were assessed by measurements of the transport of metformin, a hydrophilic drug, by monitoring transepithelial electrical resistance. Cell viability was determined by the diphenyltetrazolium bromide test (the MTT test). All the surfactants studied demonstrated concentration-dependent effects on cell permeability and cell viability. The effects on transepithelial electrical resistance correlated with cell viability, i.e. increased transepithelial electrical resistance and increased cell-monolayer permeability for metformin corresponded to decreased cell viability. The results indicate that the Solulan and polysorbate surfactants were active as absorption enhancers, Solulan C24 and 16 being more effective than polysorbates 20, 60 or 85, causing an increase in metformin transport at lower concentrations than the polysorbates. Polysorbate 20 exerted its greatest effect at a concentration of 5%-increasing the flux of metformin after 3 h by a factor of around 20 over the control. Large increases in the transport of metformin, especially at surfactant levels of 0.05%, 0.1% and 0.5%, were related to the effect of Solulan C24 and Solulan 16 on the cell permeability. The Caco-2 cell monolayer experiments confirmed the ability, especially of polysorbate 20, Solulan C24 and Solulan 16, to increase the absorption of metformin. The polysorbates increased permeability as a result of solubilisation of membrane components, while Solulans did so by penetrating and solubilising the membrane. Correlation between increase in membrane permeability and the toxicity of the surfactants towards the cell membrane has been established. PMID- 10714946 TI - Synthesis and anti-inflammatory effect of chalcones. AB - The process of degranulation of mast cells and neutrophils contributes to inflammatory disorders. Activation of microglial cells and macrophages is believed to be involved in inflammatory, infectious and degenerative diseases of the CNS. Combining the potent inhibition of chemical mediators released by the degranulation of mast cells or neutrophils and from the activated microglial cells or macrophages, would lead to a promising anti-inflammatory agent for the treatment of peripheral and central inflammation. A series of chalcone derivatives have been reported to have potent anti-inflammatory activity. In an effort to continually develop potent anti-inflammatory agents, novel series of chalcones, 2'-hydroxy- and 2',5'-dihydroxychalcones were synthesized and their inhibitory effects on the activation of mast cells, neutrophils, microglial cells and macrophages were evaluated in-vitro. The chalcones were prepared by Claisen Schmidt condensation of appropriate acetophenones with an appropriate aromatic aldehyde. The alkoxychalcones were prepared with appropriate hydroxychalcones and alkyl iodide and the dihydroxychalcones were prepared by hydrogenation of an appropriate chalcone with Pd/C. Almost all of the hydroxychalcones exhibited potent inhibitory effects on the release of beta-glucuronidase and lysozyme from rat neutrophils stimulated with formyl-Met-Leu-Phe/cytochalasin B (fMLP/CB). Of the hydroxychalcones, compound 1 was the most potent inhibitor of the release of beta-glucuronidase (IC50=1.6+/-0.2 microM) and lysozyme (IC50=1.4+/-0.2 microM) from rat neutrophils stimulated with fMLP/CB. Almost all of the 2',5' dialkoxychalcones exhibited potent inhibitory effects on nitric oxide (NO) formation from murine microglial cell lines N9 stimulated with lipopolysaccharide (LPS). Of these, compound 11 showed the greatest effect (IC50=0.7+/-0.06 microM). The present results demonstrated that most of the chalcone derivatives have an anti-inflammatory effect. The inhibitory effects of dialkoxychalcones, 10-12 on inflammation are probably not due to the inhibition of mast cells and neutrophil degranulation, but are mediated through the suppression of NO formation from N9 cells. PMID- 10714947 TI - Evaluation of the pharmacological actions and pharmacokinetics of BOF-4272, a xanthine oxidase inhibitor, in mouse liver. AB - BOF-4272 (+/-)-8-(3-methoxy-4-phenylsulphinylphenyl) pyrazolo[1,5-a]-1,3,5 triazine-4-(1H)-one, a new synthetic anti-hyperuricaemic drug, which has a chiral centre and exists as racemates, is a potent inhibitor of xanthine oxidase/dehydrogenase in the purine catabolism pathways. The present studies using mice demonstrated that BOF-4272 was specifically distributed in the liver, which is the main organ of uric acid production. Therefore, a decrease in uric acid concentration in the liver, rather than the plasma, was identified as a pharmacological action of BOF-4272. The ratio of liver to plasma concentrations of BOF-4272 increased from 2.5 to 6.3 over time, up to 8 h after oral administration. The elimination half-life of BOF-4272 in the liver was 5-1-fold longer than that in the plasma. High concentrations of BOF-4272 were observed in the liver up to 8 h after oral administration. Furthermore, the influx of BOF 4272 into hepatocytes occurred in a temperature-dependent manner. The liver concentrations of uric acid from 1 h to 8 h after the oral administration of BOF 4272 (0.34-0.75 microg (g tissue)(-1)) were significantly lower than those in control animals (5.03-10.96 microg (g tissue)(-1)). BOF-4269 (the sulphide metabolite of BOF-4272) was the only metabolite detected in plasma or faeces after intravenous or oral administration. BOF-4269, which has no inhibitory action on the uric acid biosynthesis system, is generated by the metabolism of BOF-4272 in the intestinal tract. In conclusion, this work using the liver as the target organ has allowed us to identify the pharmacological actions of BOF-4272 in mice. The long-lasting effect of BOF-4272 in reducing levels of hepatic uric acid was consistent with the prolonged high BOF-4272 concentrations in the liver. These results also demonstrate that the mouse is a suitable animal species for evaluating the clinical pharmacology and pharmacokinetics of BOF-4272. PMID- 10714948 TI - Development and validation of a recirculatory physiological model of the myocardial concentrations of lignocaine after intravenous administration in sheep. AB - A recirculatory physiological model of the determinants of the myocardial concentrations of lignocaine after intravenous administration was developed in sheep and validated with the intention of analysing and predicting the outcome of altered dose regimens and various pathophysiological states on the initial myocardial concentrations of lignocaine. The structure and parameters of the model were determined by hybrid modelling of the time-courses of the pulmonary artery, arterial and coronary sinus concentrations of lignocaine after the intravenous administration of 100 mg of lignocaine over 5 min to 5 chronically instrumented sheep. The model accounted for the determinants of the myocardial concentrations via compartments for venous mixing, the lung (a single-compartment model with a first-order loss) and the heart (a single flow-limited compartment). Recirculation and the remainder of the body were represented as a single tissue pool with a clearance term. The distribution volume of the heart was 0.42+/-0.009 L, which gave a half-time of myocardium:blood equilibration of 2.37 min. The distribution volume of the lungs was 5.40+/-0.23 L, with an apparent first-order loss of 1.02 L min(-1) representing deep distribution or metabolism. The validity of the model was tested by comparing the predictions of the model with the equivalent data collected in 6 sheep when lignocaine (89 mg) was administered via a complex dose regimen with a faster initial rate of infusion (39.1 mg min(-1)), declining exponentially to basal infusion rate (7.02 mg min(-1)) over 8 min. The predictions of the model were in general agreement with these data. It is concluded that the model was sufficient to account for the effect of altered dose regimens of lignocaine on the time-course of its myocardial concentrations. PMID- 10714950 TI - Rapid detection of CYP2C18 genotypes by real-time fluorescence polymerase chain reaction. AB - In man, CYP2C19, a liver enzyme, plays an important role in the metabolism of several drugs. Mutation of the CYP2C19 gene results in a poor metaboliser phenotype. S-Mephenytoin hydroxylation genetic polymorphism is due to two mutations of the CYP2C19 gene, namely CYP2C19*2, located in exon 5, and CYP2C19*3, located in exon 4. CYP2C18 is also polymorphically expressed. The mutant alleles of this enzyme are CYP2C18m1, located in exon 2 and CYP2C18m2, located in the 5'-flanking region. We have developed an allele-specific TaqMan polymerase chain reaction (PCR) assay with which to detect CYP2C18 mutant alleles. This assay combines hybridization of the TaqMan probe and allele specific amplification primers to the target DNA. The TaqMan probe is labelled with 6-carboxyfluorescein at the 5' end and 6-carboxytetramethylrhodamine together with a phosphate at the 3' end. Genotypes are separated according to the different threshold cycles of the wild type and mutant primers. We applied this procedure to DNA extracted from the blood or saliva of 144 healthy Japanese volunteers. The wt/wt, wt/m1, wt/m2, m1/m1, m1/m2 and m2/m2 genotypes of the CYP2C18 alleles detected by the assay were consistent with the results obtained from restriction enzyme cleavage. In accordance with a previous report, the genotypes of CYP2C18m1 and CYP2C18m2 coincided with those of CYP2C19*3 and CYP2C19*2, respectively. Therefore, detection of CYP2C18 mutant alleles also allows that of CYP2C19 mutant alleles. Among 19 poor metabolisers, eight showed the homozygous CYP2C19*2/CYP2C19*2, two the homozygous CYP2C19*3/CYP2C19*3 and nine the compound heterozygous CYP2C19*2/CYP2C19*3 genotype. We found the allele specific TaqMan PCR assay rapid, simple and cost-effective, as well as suitable for high-throughput applications in a routine laboratory. This assay allows the fast and reliable detection of inherited disorders that might influence diagnosis and treatment. PMID- 10714949 TI - Stereoselective metabolism of the monoterpene carvone by rat and human liver microsomes. AB - The large amounts of carvone enantiomers consumed as food additives and in dental formulations justifies the evaluation of their biotransformation pathway. The in vitro metabolism of R-(-)- and S-(+)-carvone was studied in rat and human liver microsomes using chiral gas chromatography. Stereoselective biotransformation was observed when each enantiomer was incubated separately with liver microsomes. 4R, 6S-(-)-Carveol was NADPH-dependently formed from R-(-)-carvone, whereas 4S, 6S (+)-carveol was produced from S-(+)-carvone. Metabolite formation followed Michaelis-Menten kinetics exhibiting a significant lower apparent Km (Michaelis Menten Constant) for 4R, 6S-(-)-carveol compared with 4S, 6S-(+)-carveol in rat and human liver microsomes (28.4+/-10.6 microM and 69.4+/-10.3 microM vs 33.6+/-8 55 microM and 98.3+/-22.4 microM). The maximal formation rate (Vmax) determined in the same microsomal preparations yielded 30.2+/-5.0 and 32.3+/-3.9 pmol (mg protein)(-1) min(-1) in rat liver and 55.3+/-5.7 and 65.2+/-4.3 pmol (mg protein)(-1) min(-1) in human liver microsomes. Phase II conjugation of the carveol isomers by rat and human liver microsomes in the presence of UDPGA (uridine S'-diphosphogluaronic acid) only revealed glucuronidation of 4R, 6S-(-) carveol. Vmax for glucuronide formation was more than 4-fold higher in the rat liver compared with human liver preparations (185.9+/-34.5 and 42.6+/-7.1 pmol (mg protein)(-1) min(-1), respectively). Km values, however, showed no species related difference (13.9+/-4.1 microM and 10.2+/-2.2 microM). This study demonstrated stereoselectivity in phase-I and phase-II metabolism for R-(-)- and S-(+)-carvone and might be predictive for carvone biotransformation in man. PMID- 10714951 TI - Specific binding of nicergoline on an alpha1-like adrenoreceptor in the rat retina. AB - Systemic treatment with nicergoline, an ergoline derivative showing alpha1 antagonist properties, causes vasodilatation in the eye without apparent untoward cardiovascular effects. In the present work we investigated the ability of nicergoline to inhibit the binding of radiolabelled prazosin in the rat retina and cortex. We found that nicergoline inhibited [3H]prazosin binding in both tissues, being more potent than unlabelled prazosin in the retinal tissue. The competition curves of the ergoline derivative were well fitted by a one-site model in the cortical tissue, with an IC50 (concentration of the drugs needed to inhibit the binding of labelled prazosin by 50%) of 2.54 x 10(-8) M, and by a two site model in the retinal tissue, with IC50 values of 7.08 x 10(-12) M and 1.82 x 10(-5) M. 2-(2,6 dimetoxyphenoxyethyl) aminomethyl-1,4-benzodioxane hydrochloride (WB4101) and phentolamine, selective ligands for the high-affinity binding site for prazosin, in particular the alpha1A-site, fully inhibited prazosin binding in the cortex but only partially inhibited prazosin binding in the retina, being less potent in this tissue than either nicergoline or prazosin. Our results suggest that a binding component of alpha1-adrenoreceptors is expressed to a lesser extent in the retina than the cortex, leading to a reduced response of the retinal tissue to prazosin, and more particularly to WB4101 and phentolamine. The selective binding of the nicergoline on this retinal adrenoreceptor may explain the peculiar efficacy of the drug in ocular pathophysiology. PMID- 10714952 TI - Comparison of the vasorelaxing effect of cromakalim and the new inodilator, levosimendan, in human isolated portal vein. AB - In the present study the vasorelaxing capacity of cromakalim, an ATP-sensitive potassium-channel (KATP channel) activator, and that of levosimendan, a new positive inotropic and vasodilating drug with calcium sensitizing and potassium channel-activating properties, were compared in human isolated portal vein. Based on the 50% effective concentrations (EC50), levosimendan was found to be about 16 fold more potent (EC50 = 0.281+/-0.03 microM) as a relaxing agent than cromakalim (EC50 = 4.53+/-0.12 microM) in noradrenaline-precontracted portal venous preparations. Glibenclamide, the known inhibitor of KATP channels, was able to prevent the cromakalim-induced venodilation completely. Glibenclamide (15 microM) decreased the quasi-maximal effect of levosimendan (at 1.27 microm by about 60%) and also the effects of those submicromolar concentrations of the inodilator (at 0.1 microM by 23%, at 0.3 microM by 27% and at 0.7 microM by 19%, on average) which were therapeutically effective in preliminary human studies. These findings indicate that, in the human portal vein, both cromakalim and levosimendan are powerful vasorelaxants and that a considerable part of the relaxing effect induced by levosimendan is of cromakalim type. PMID- 10714953 TI - Protection of mitochondrial functions against oxidative stresses by isoflavans from Glycyrrhiza glabra. AB - Isoflavan derivatives, glabridin (1), hispaglabridin A (2), hispaglabridin B (3), 4'-Omethylglabridin (4) and 3'-hydroxy-4'-O-methylglabridin (5), isolated from Glycyrrhiza glabra, were investigated for their ability to protect liver mitochondria against oxidative stresses. Mitochondrial lipid peroxidation linked to respiratory electron transport and that induced non-enzymatically were inhibited by these isoflavans. Hispaglabridin A (2) strongly inhibited both peroxidations and 3'-hydroxy-4'-O-methylglabridin (5) was the most effective at preventing NADH-dependent peroxidation. 3'-Hydroxy-4'-O-methylglabridin (5) protected mitochondrial respiratory enzyme activities against NADPH-dependent peroxidation injury. Dihydroxyfumarate-induced mitochondrial peroxidation was also prevented by this isoflavan. Isoflavans from G. glabra were shown to be effective in protecting mitochondrial function against oxidative stresses. PMID- 10714954 TI - Endothelin-1 mediates hypocapnic constriction of the rabbit basilar artery in vitro. PMID- 10714955 TI - Do biocides select for antibiotic resistance? AB - Some similarities exist between bacterial resistance to antibiotics and to biocides, and gram-negative bacteria that have developed resistance to cationic biocides may also be insusceptible to some antibiotics. Outer membrane changes are believed to be responsible for this non-specific increase in resistance. Efflux, another important resistance mechanism, is associated with the qacA/B gene system in staphylococci that confers low-level resistance to cationic agents including chlorhexidine salts and quaternary ammonium compounds. It has been proposed that the introduction into clinical practice of chlorhexidine and quaternary ammonium compounds has resulted in the selection of staphylococci containing qacA genes on multiresistance plasmids. A linkage between low-level resistance to triclosan and to antibiotics has recently been claimed to occur in Escherichia coli, with the bisphenol selecting for chromosomally-mediated antibiotic resistance. A key issue in many studies has been the use of biocides at concentrations significantly below those used clinically. It remains to be determined how an increase to low-level resistance to cationic biocides can be held responsible for the selection of antibiotic-resistant bacteria. PMID- 10714956 TI - Extract of Juglandaceae regia inhibits growth, in-vitro adherence, acid production and aggregation of Streptococcus mutans. AB - Aqueous and alcoholic extracts from Juglandaceae regia, used as chewing sticks to maintain oral hygiene, were tested for their ability to inhibit the growth and some physiological functions of Streptococcus mutans. Both the aqueous and the alcoholic extract strongly inhibited the growth, in-vitro adherence, acid production and glucan-induced aggregation of S. mutans. At a concentration of 8% w/v, the aqueous extract produced a 95% inhibition (P < 0.05) of adherence of S. mutans to glass and a 40% inhibition (P < 0.05) of adherence to tooth surface. The alcoholic extract at a concentration of 10% w/v produced a 95% inhibition (P < 0.05) of adherence of S. mutans to glass and a 56% inhibition (P < 0.05) of adherence to tooth surface. At concentrations of 2% w/v the aqueous and alcoholic extracts significantly inhibited (P < 0.05) glucan-induced aggregation of S. mutans and the in-vitro salivary glycolytic reaction for up to 5 h. Bactericidal effects on S. mutans were also evident. At a concentration of 10% w/v, the zone of inhibition observed with the aqueous extract was 12+/-0.01 mm and that observed with the alcoholic extract was 12.6+/-0.02 mm. As the in-vitro studies had shown that both the aqueous and the alcoholic extract of J. regia, at concentrations of 10% w/v, could inhibit the growth as well as the acid-producing ability of S. mutans, they were tested at the same concentration for their activity in-vivo. Three subjects were employed. Parameters monitored were salivary bacterial count and salivary glycolysis. Mouth-rinsing with the aqueous but not the alcoholic extract significantly reduced total streptococcal counts in the salivary samples obtained up to, and including, 3 h after rinsing, compared with the counts obtained pre-rinsing or after placebo rinsing. Mouth-rinsing with the aqueous extract produced a 65%, 27% and 78% reduction (P < 0.05) in the streptococcal count in the salivary samples obtained 10 min, 1 h and 3 h after rinsing, respectively. Both the aqueous and the alcoholic extract also inhibited the glycolytic reaction by the salivary bacteria for up to 90 min post-rinsing. This study provides evidence to justify the use of J. regia sticks as an aid to maintain oral hygiene. PMID- 10714957 TI - Stimulatory effect of procaine on the growth of several microalgae and cyanobacteria. AB - Procaine has been used to stimulate plant growth and it has been noted that it also promotes growth of microorganisms. The effect of procaine hydrochloride concentration on the growth rates of several species of microalgae and cyanobacteria was studied under both photoautotropic and heterotrophic growth conditions. Procaine hydrochloride was added to cultures at concentrations over the range 0.01-1000 mg L(-1). A stimulating effect of procaine hydrochloride on photoautotrophic growth was observed for the cyanobacteria Anabaena cylindrica and Anabaena variabilis, and for the salt-tolerant green algae Dunaliella primolecta and Dunaliella parva. During active growth in batch culture an increase in growth rate (compared with control culture without procaine hydrochloride) of about 25% was observed at 0.1 mgL(-1) of procaine hydrochloride for A. cylindrica. However, procaine hydrochloride was toxic at concentrations of > 10 mgL(-1). Simultaneous administration of hydrolysis products of procaine, p aminobenzoic acid and diethyl aminoethanol, in lieu of procaine hydrochloride, was as effective as procaine in stimulating growth of A. cylindrica. Heterotrophic growth of Chlorella ellipsoidea and Prototheca zopfii was not stimulated by procaine hydrochloride over the concentration range studied (0.1-10 mg L(-1)). The combined effects of procaine hydrochloride concentration and four other environmental factors (temperature, light intensity, CO2 concentration in the flushing gas and NaCl concentration) on growth rate of D. primolecta was modelled using both a neural network approach and a response surface method. These results indicate that procaine hydrochloride exerts different effects on the growth of microalgal and cyanobacterial cells as functions of dosage, species and culture conditions. PMID- 10714958 TI - The role of p53-target genes in human cancer. AB - The tumor suppressor gene p53 is mutated in a large proportion of human cancers. In some cellular conditions like DNA damage, the p53 gene is induced and its gene product is posttranscriptionally activated. p53 works as a transcriptional activator and induces the expression of its downstream target genes. This review will explain why expression of the normal p53 gene leads to tumor growth suppression. The p53 has several biological effects involving cell-cycle arrest, DNA replication and repair, proliferation, apoptosis, angiogenesis inhibition, and cellular stress response. These effects of the p53 result mainly from the activation of expression of a large number of p53-target genes. Here we have focused on the biological functions of the transcriptional targets of p53. PMID- 10714959 TI - Inhibition of RAS-targeted prenylation: protein farnesyl transferase inhibitors revisited. AB - The ras oncogene and its 21 kD protein product, Ras, has emerged during the last decade as a potentially exploitable target for anticancer drug development. The knowledge that Ras was readily prenylated by protein farnesyl transferase (PFTase) and that inhibition of this prenylation had functional consequences for the transformed phenotype that expressed oncogenic Ras provided the rational for the development of PFTase inhibitors. The initial enthusiasm for this approach seemed justified by the early identification of PFTase inhibitors that were able potently and specifically to block Ras processing, signalling and transformation in transformed and tumour cell lines in vitro and in certain selected animal models. More recently the recognition that geranylgeranyl transferase (GGTase) I might also be a therapeutic target is being actively researched. The last couple of years though have proved remarkable with the disclosure of a series of structurally-diverse molecules, whose major in vivo preclinical activites have been well documented against experimental animal tumours, and culminating this year in preliminary reporting of their Phase I clinical evaluations. Nevertheless, during the research and development phases of PFTase inhibitors as pharmaceutical agents for clinical use, there have been several unexpected findings which have raised intriguing and potentially crucial questions about their activities. This review aims to highlight and offer new insights into many of these issues and to bring into perspective concerns arising from basic research, as well as from clinical studies. There seems little doubt that these inhibitors of RAS-targeted prenylation represent a new generation of anticancer drugs for the preclinical researcher, whether they can be successfully exploited in clinical practice should be resolved early in the next millenium. PMID- 10714960 TI - Glycosyl phosphatidylinositol (GPI)-anchored molecules and the pathogenesis of paroxysmal nocturnal hemoglobinuria. AB - Paroxysmal nocturnal hemoglobinuria (PNH) is characterized by the expansion of one or more clones of stem cells producing progeny of mature blood cells deficient in the plasma membrane expression of all glycosyl phosphatidylinositol (GPI)-anchored proteins (AP). This is due to somatic mutations in the X-linked gene PIGA, encoding one of the several enzymes required for GPI anchor biosynthesis. More than 20 GPI-APs are variously expressed on hematological cells. GPI-APs may function as enzymes, receptors, complement regulatory proteins or adhesion molecules; they are often involved in signal transduction. The absence of GPI-APs may well explain the main clinical findings of PNH, i.e., hemolysis and thrombosis in the venous system. Other aspects of PNH pathophysiology such as various degrees of bone marrow failure and the dominance of the PNH clone may also be linked to the biology and function of GPI-APs. Results of in vitro and in vivo experiments on embryoid bodies and mice chimeric for nonfunctional Piga have recently demonstrated that Piga inactivation confers no intrinsic advantage to the affected hematopoietic clone under physiological conditions; thus additional factors are required to allow for the expansion of the mutated cells. A close association between PNH and aplastic anemia suggests that immune system mediated bone marrow failure creates and maintains the conditions for the expansion of GPI-AP deficient cells. In this scenario, a PIGA mutation would render GPI-AP deficient cells resistant to the cytotoxic autoimmune attack, enabling them to emerge. Even though the 'survival advantage' hypothesis may explain all the various aspects of this intriguing disease, a formal proof of this theory is still lacking. PMID- 10714961 TI - The comprehensive geriatric assessment: when, where, how. AB - Cancer is increasingly a disease of the aged, a segment of the population that is the fastest growing. Often, cancer adds on to the progressive deterioration of normal aging and to the impairment associated with the presence of multiple concomitant medical problems. Thus, the likelihood that cancer leads to disability is much greater among older patients than younger ones. In consideration of the dimension of the problem, and of the peculiarities of the elderly patient, it has recently been proposed that a new approach termed 'comprehensive geriatric assessment' (CGA) might allow a better management and more efficient care of elderly patients with cancer. The systematic introduction of CGA in clinical research and in daily practice can contribute to: identify cancer patients for whom we could expect the greatest benefit from treatment; assess their physiologic, functional and health-related quality of life; formulate appropriate treatment and management strategies; monitor clinical and functional outcomes; provide a more accurate evaluation of prognostic indicators. PMID- 10714962 TI - Integration of geriatrics in oncology training--the relationship between the academic center and the community. AB - The aging of the population and the ensuing large increase in the number of older cancer patients require that the subspeciality of Medical Oncology respond quickly and effectively to this need. Sixty percent of all cancer occurs in persons aged 65 years or older and there is an unprecedented growth of this segment of the population. Their treatment is further complicated by the preexisting medical conditions and their unique social and economic needs. The response of both the academic and private practice communities will greatly effect the future of cancer care for the elderly. There is an immediate need for physicians and other health care providers to better understand the influence of advancing age on diagnosis and treatment of cancer. Integration of geriatric and medical oncology training would be an important step in this process. This paper discusses various aspects of a combined educational proposal. PMID- 10714964 TI - FDA gives ground to supplement makers. PMID- 10714963 TI - Idarubicin and cyclophosphamide--an active oral chemotherapy regimen for advanced breast cancer. AB - Between October 1993 and September 1994, 33 women with metastatic breast cancer aged between 29 and 74 years with a median age of 58 were entered into a study of oral chemotherapy from three UK centres. Patients by definition had metastatic disease and were fit and well with performance status 0 or 1 in 23 cases, 2 in seven cases and 3 in two cases (one missing). Five patients had received prior adjuvant CMF chemotherapy, nine first line non-anthracycline containing chemotherapy for relapse, eight patients second line non-anthracycline containing chemotherapy and all patients had had hormone therapy either as adjuvant or for relapsed disease. Adjuvant radiotherapy had been given to 17 and palliative radiotherapy to 12 patients. In nine patients there was one site of disease at start of therapy, in 10 two sites, in 11 three sites and in three patients four or more sites. The regimen comprised oral idarubicin 15 mg/m2 on day 1, 10 mg/m2 on days 2 and 3 and oral cyclophosphamide 250 mg/m2 (maximum 400 mg) on days 1, 2 and 3. Treatment was continued until disease progression or toxicity. RESULTS: Overall 25% of 32 evaluable patients responded objectively including one complete response; 50% of patients had stable disease and 25% of patients progression. Among patients who had had no prior chemotherapy the objective response rate was 37.5%; 45% of patients had symptomatic improvement. The most common severe toxicity was granulocytopenia WHO grade 3 or more in 69.7% of patients. Thrombocytopenia grade 3 or 4 was seen in four patients. Six patients had documented infections and all but four patients had alopecia. All patients complained of mild or moderate fatigue. Nausea and vomiting occurred in 75% of patients but only four individuals had grade 3 toxicity. Two patients stopped therapy after myocardial infarction and one after impaired cardiac function was noted. The median time to progression was 2.7 months (1-11.5 months) and median survival time 8.8 months (1-13+ months). CONCLUSION: The combination chemotherapy is active in heavily treated patients with manageable toxicity but there are problems in heavily pre-treated patients. There was good compliance in taking medication and at the doses chosen the drugs appear to be suitable for younger fitter patients. PMID- 10714966 TI - Disclosure of errors may have financial benefit. PMID- 10714965 TI - VA releases adverse event data. Clinicians must report to nationwide system. PMID- 10714967 TI - Greater patient involvement needed in ambulatory care decisions. Study finds fully informed decision-making rare. PMID- 10714968 TI - Illinois revises requirements for error records. PMID- 10714969 TI - Valerian: Valeriana officinalis. PMID- 10714970 TI - Too many medication errors, not enough pharmacists. PMID- 10714971 TI - Cost-effectiveness analysis of six strategies for cardiovascular surgery prophylaxis in patients labeled penicillin allergic. AB - The cost-effectiveness of different approaches to antimicrobial prophylaxis for cardiovascular surgery patients labeled penicillin allergic was studied. A decision-analytic model was used to examine the cost-effectiveness of six strategies for antimicrobial prophylaxis in cardiovascular surgery patients at a tertiary care hospital. The strategies consisted of (1) giving vancomycin to all patients labeled penicillin allergic, (2) giving cefazolin to all patients labeled penicillin allergic, (3) giving vancomycin to all patients with a history suggesting an immunoglobulin E (IgE)-mediated reaction to penicillin and cefazolin to patients without such a history, (4) administering a penicillin skin test to patients with a history suggesting an IgE-mediated reaction to penicillin and giving vancomycin to patients with positive results and cefazolin to all others, (5) skin testing all patients labeled penicillin allergic and giving vancomycin to those with positive results and cefazolin to those with negative results, regardless of history, and (6) skin testing all patients and giving vancomycin to those with positive results or a history suggesting an IgE-mediated reaction to penicillin and cefazolin to all others. Giving cefazolin to all patients labeled penicillin allergic was the least expensive strategy but was associated with the highest rate of both anaphylactic and non-life-threatening serious reactions. Selective use of vancomycin in patients with a history suggesting an IgE-mediated reaction to penicillin was associated with an added cost and a slightly lower rate of anaphylaxis. Although skin-testing strategies may decrease both non-life-threatening and anaphylactic reactions, the incremental cost was high. When vancomycin was given to all patients labeled penicillin allergic, the incremental cost was very high. A decision-analytic model indicated that selective use of vancomycin is more cost-effective than indiscriminate use of vancomycin for surgical prophylaxis in cardiovascular surgery patients labeled penicillin allergic. PMID- 10714972 TI - Understanding the critical components of a successful cleanroom and barrier isolator project. AB - The critical components of clean-room and barrier isolator systems are described. Cleanroom and barrier isolator systems have four basic parts: the physical structure, the internal environment, the interaction technology, and the monitoring system. To create an aseptic environment, pharmacists must understand each of these components and be able to provide vendors with clear specifications. Among the decisions pharmacists must make are what materials should be used in construction, how many filters to use and where these should be placed, how to best reduce the contamination challenge to the area, and the best means of monitoring the room's air cleanliness. Evaluation of each component should be made on the basis of durability, functionality, and cost. Pharmacists must also have a set of criteria to help them choose a vendor. They must know their state's requirements and ensure that whatever system is built for their organization will meet all regulations. Cleanroom and barrier isolator systems must adhere to Federal Standard 209E, which defines standard classes of air cleanliness that are based on specific concentrations and sizes of airborne particles. Having an understanding of the basic components of cleanroom and barrier isolator systems will help pharmacists define their needs and describe them to vendors. PMID- 10714973 TI - Modern inventory analysis techniques. AB - Modern techniques for managing pharmacy inventories are described. Pharmacists should rely on modern techniques, such as sort-based and activity-based analyses, for managing pharmacy inventories, containing drug costs, performing replacement and-elimination analysis, and monitoring the health system's operations. Unit price and quantity are the two basic inventory-control approaches; however, modern techniques recognize quantity as the more useful of the two. The primary areas of the pharmacy's activities must be taken into consideration. Pharmacists must learn to divide inventory analysis problems into sets of smaller issues. Modern inventory analyses that take into account annual quantity, unit price, total annual cost, and the health system's unique activities provide the pharmacist with a practical basis for inventory management. PMID- 10714974 TI - Comparative effectiveness of low-molecular-weight heparins after therapeutic interchange. AB - Management Case Studies describe approaches to real-life management problems in health systems. Each installment is a brief description of a problem and how it was dealt with. The cases are intended to help readers deal with similar experiences in their own work sites. Problem solving, not hypothesis testing, is emphasized. Successful resolution of the management issue is not a criterion for publication-important lessons can be learned from failures, too. PMID- 10714975 TI - Pharmacy practice in Russia amid economic turmoil. AB - The Pharmacy Abroad section of AJHP features brief, informal, and topical communications related to pharmacy in other countries. Contributions are welcomed from pharmacists abroad or from pharmacists who have traveled abroad. AJHP also encourages pharmacists from outside of the United States to submit traditional manuscripts (e.g., scientific studies, descriptions of practice innovations), which are evaluated for publication in the primary sections of the journal. PMID- 10714976 TI - Depression in patients with HIV infection. AB - The epidemiology, clinical features, and drug treatment of depression in HIV infected patients are discussed. The lifetime prevalence of depression in patients infected with HIV has been estimated at 22-45%. The signs and symptoms of depression are similar in HIV-infected and noninfected patients, but patients with HIV infection may more frequently have sleep and appetite disturbances. Diagnosis should focus on affective or cognitive depression symptoms that reflect mood state alone. Patients with a history of depression, homosexual men, women, and i.v. drug abusers are among HIV-infected individuals who may be at increased risk for depression. Depression may alter the course of HIV infection by impairing immune function or influencing behavior. Depression my contribute to nonadherence to therapy. Antidepressant therapy is effective in most HIV-positive patients with major depression. Tricyclic antidepressants (TCAs) have produced response rates as high as 89%, but their usefulness has been limited by adverse effects. Selective serotonin-reuptake inhibitors and other non-TCAs have also demonstrated efficacy and are generally better tolerated. Psychostimulants have improved mood, cognition, and energy level, and androgens have been used for their anabolic effects. The systemic concentrations of antidepressants may be altered by coadministered drugs that affect their cytochrome P-450 isoenzyme mediated metabolism; in turn, the metabolism and toxicity of certain antiretrovirals may be affected by antidepressants. Guidelines on the treatment of depression in the general population may be applied to patients with HIV infection. Depressive disorders are prevalent among patients with HIV infection but often respond to a variety of treatments. PMID- 10714977 TI - Premixed i.v. admixtures for reducing costs and labor time in a home infusion company. PMID- 10714978 TI - Consider potential for drug interactions during formulary review. PMID- 10714979 TI - A novel diagnostic method for pulmonary aspiration in a murine model. Immunocytochemical staining of milk proteins in alveolar macrophages. AB - Aspiration of foreign material into the lungs has been implicated in the etiology of a variety of pulmonary disorders. Although aspiration is a common clinical problem, its diagnosis represents a major challenge due to the lack of sensitive and/or specific tests. In this study, we evaluated the sensitivity and specificity of a novel diagnostic method in a murine model of milk aspiration. Under light anesthesia, BALB/c mice received either single or repeated intranasal instillation of milk. Control animals received sterile physiologic saline or were infected with respiratory pathogens in a similar manner. After isolation and cannulation of the trachea, mouse lungs were lavaged with PBS at various time points after the last aspiration event. Cells were recovered for Oil Red O (ORO) staining as well as immunocytochemistry for milk proteins: alpha-lactalbumin and beta-lactoglobulin. After single aspiration of milk, a large number of alveolar macrophages displayed a strong immunoreactivity for alpha-lactalbumin for 2-96 h. After single and repeated aspiration, the percentage of positive cells for alpha lactalbumin was significantly higher when compared with ORO staining at 24, 48, and 72 h (p < 0.05). No immunoreactivity for milk proteins was found in alveolar macrophages obtained from our control groups. These findings demonstrate that immunocytochemical staining of milk proteins within alveolar macrophages represents a novel, sensitive, and specific test for the diagnosis of aspiration in a murine model. PMID- 10714980 TI - An exclusive contract: specificity in the Vibrio fischeri-Euprymna scolopes partnership. PMID- 10714981 TI - Flavodoxin mutants of Escherichia coli K-12. AB - The flavodoxins are flavin mononucleotide-containing electron transferases. Flavodoxin I has been presumed to be the only flavodoxin of Escherichia coli, and its gene, fldA, is known to belong to the soxRS (superoxide response) oxidative stress regulon. An insertion mutation of fldA was constructed and was lethal under both aerobic and anaerobic conditions; only cells that also had an intact (fldA(+)) allele could carry it. A second flavodoxin, flavodoxin II, was postulated, based on the sequence of its gene, fldB. Unlike the fldA mutant, an fldB insertion mutant is a viable prototroph in the presence or absence of oxygen. A high-copy-number fldB(+) plasmid did not complement the fldA mutation. Therefore, there must be a vital function for which FldB cannot substitute for flavodoxin I. An fldB-lacZ fusion was not induced by H(2)O(2) and is therefore not a member of the oxyR regulon. However, it displayed a soxS-dependent induction by paraquat (methyl viologen), and the fldB gene is preceded by two overlapping regions that resemble known soxS binding sites. The fldB insertion mutant did not have an increased sensitivity to the effects of paraquat on either cellular viability or the expression of a soxS-lacZ fusion. Therefore, fldB is a new member of the soxRS (superoxide response) regulon, a group of genes that is induced primarily by univalent oxidants and redox cycling compounds. However, the reactions in which flavodoxin II participates and its role during oxidative stress are unknown. PMID- 10714982 TI - The Staphylococcus aureus lrgAB operon modulates murein hydrolase activity and penicillin tolerance. AB - Previous studies in our laboratory have shown that the Staphylococcus aureus LytSR two-component regulatory system affects murein hydrolase activity and autolysis. A LytSR-regulated dicistronic operon has also been identified and shown to encode two potential membrane-associated proteins, designated LrgA and LrgB, hypothesized to be involved in the control of murein hydrolase activity. In the present study, a lrgAB mutant strain was generated and analyzed to test this hypothesis. Zymographic and quantitative analysis of murein hydrolase activity revealed that the lrgAB mutant produced increased extracellular murein hydrolase activity compared to that of the wild-type strain. Complementation of the lrgAB defect by providing the lrgAB genes in trans restored the wild-type phenotype, indicating that these genes confer negative control on extracellular murein hydrolase activity. In addition to these effects, the influence of the lrgAB mutation on penicillin-induced lysis and killing was examined. These studies demonstrated that the lrgAB mutation enhanced penicillin-induced killing of cells approaching the stationary phase of growth, the time at which the lrgAB operon was shown to be maximally expressed. This effect of the lrgAB mutation on penicillin-induced killing was shown to be independent of cell lysis. In contrast, the lrgAB mutation did not affect penicillin-induced killing of cells growing in early-exponential phase, a time in which lrgAB expression was shown to be minimal. However, expression of the lrgAB operon in early-exponential-phase cells inhibited penicillin-induced killing, again independent of cell lysis. The data generated by this study suggest that penicillin-induced killing of S. aureus involves a novel regulator of murein hydrolase activity. PMID- 10714983 TI - Characterization of the Mycobacterium tuberculosis iniBAC promoter, a promoter that responds to cell wall biosynthesis inhibition. AB - The cell wall provides an attractive target for antibiotics against Mycobacterium tuberculosis. Agents such as isoniazid and ethambutol that work by inhibiting cell wall biosynthesis are among the most highly effective antibiotics against this pathogen. Although considerable progress has been made identifying the targets for cell wall active antibiotics, little is known about the intracellular mechanisms that are activated as a consequence of cell wall injury. These mechanisms are likely to have an important role in growth regulation and in the induction of cell death by antibiotics. We previously discovered three isoniazid induced genes (iniB, iniA, and iniC) organized in tandem on the M. tuberculosis genome. Here, we investigate the unique features of the putative iniBAC promoter. This promoter was specifically induced by a broad range of inhibitors of cell wall biosynthesis but was not inducible by other conditions that are toxic to mycobacteria via other mechanisms. Induction required inhibitory concentrations of antibiotics and could be detected only in actively growing cells. Analysis of the iniBAC promoter sequence revealed both a regulatory element upstream and a potential repressor binding region downstream of the transcriptional start site. The induction phenotype and structure of the iniBAC promoter suggest that a complex intracellular response occurs when cell wall biosynthesis is inhibited in M. tuberculosis and other mycobacteria. PMID- 10714984 TI - Reinvestigation of the proteolytic activity of neocarzinostatin. AB - Neocarzinostatin (NCS) is the most studied member of a family of chromoproteins secreted by a range of actinomycetes species. It has been proposed that in addition to their antitumoral activity related to the bound chromophores, this group of related proteins could be a secreted proteases superfamily. With the aim of dissecting the molecular basis of the proteolytic activity of NCS, an expression system allowing efficient expression of apo-NCS in Escherichia coli was constructed. The recombinant protein was properly folded and functional. Its histone-specific proteolytic activity was similar to the activity described for the natural protein. Further analyses unambiguously demonstrated that the proteolytic activity could be physically separated from NCS. This activity is therefore due not to NCS itself but to minor contaminating proteases, the nature of which differed in the recombinant and natural NCS preparations. The histone degradation test commonly used to monitor proteolytic activity is extremely sensitive and may easily generate false-positive results. These results strongly suggest that the possible proteolytic activity of the proteins of this family should be critically reconsidered. PMID- 10714985 TI - trp RNA-binding attenuation protein-5' stem-loop RNA interaction is required for proper transcription attenuation control of the Bacillus subtilis trpEDCFBA operon. AB - The trp RNA-binding attenuation protein (TRAP) regulates expression of the Bacillus subtilis trpEDCFBA operon by a novel transcription attenuation mechanism. Tryptophan-activated TRAP binds to the nascent trp leader transcript by interacting with 11 (G/U)AG repeats, 6 of which are present in an antiterminator structure. TRAP binding to these repeats prevents formation of the antiterminator, thereby promoting formation of an overlapping intrinsic terminator. A third stem-loop structure that forms at the extreme 5' end of the trp leader transcript also plays a role in the transcription attenuation mechanism. The 5' stem-loop increases the affinity of TRAP for trp leader RNA. Results from RNA structure mapping experiments demonstrate that the 5' stem-loop consists of a 3-bp lower stem, a 5-by-2 asymmetric internal loop, a 6-bp upper stem, and a hexaloop at the apex of the structure. Footprinting results indicate that TRAP interacts with the 5' stem-loop and that this interaction differs depending on the number of downstream (G/U)AG repeats present in the transcript. Expression studies with trpE'-'lacZ translational fusions demonstrate that TRAP 5' stem-loop interaction is required for proper regulation of the trp operon. 3' RNA boundary experiments indicate that the 5' structure reduces the number of (G/U)AG repeats required for stable TRAP-trp leader RNA association. Thus, TRAP 5' stem-loop interaction may increase the likelihood that TRAP will bind to the (G/U)AG repeats in time to block antiterminator formation. PMID- 10714986 TI - Morphogenetic proteins SpoVID and SafA form a complex during assembly of the Bacillus subtilis spore coat. AB - During endospore formation in Bacillus subtilis, over two dozen polypeptides are assembled into a multilayered structure known as the spore coat, which protects the cortex peptidoglycan (PG) and permits efficient germination. In the initial stages of coat assembly a protein known as CotE forms a ring around the forespore. A second morphogenetic protein, SpoVID, is required for maintenance of the CotE ring during the later stages, when most of proteins are assembled into the coat. Here, we report on a protein that appears to associate with SpoVID during the early stage of coat assembly. This protein, which we call SafA for SpoVID-associated factor A, is encoded by a locus previously known as yrbA. We confirmed the results of a previous study that showed safA mutant spores have defective coats which are missing several proteins. We have extended these studies with the finding that SafA and SpoVID were coimmunoprecipitated by anti SafA or anti-SpoVID antiserum from whole-cell extracts 3 and 4 h after the onset of sporulation. Therefore, SafA may associate with SpoVID during the early stage of coat assembly. We used immunogold electron microscopy to localize SafA and found it in the cortex, near the interface with the coat in mature spores. SafA appears to have a modular design. The C-terminal region of SafA is similar to those of several inner spore coat proteins. The N-terminal region contains a sequence that is conserved among proteins that associate with the cell wall. This motif in the N-terminal region may target SafA to the PG-containing regions of the developing spore. PMID- 10714987 TI - The Yersinia pestis YscY protein directly binds YscX, a secreted component of the type III secretion machinery. AB - Human pathogenic yersiniae organisms export and translocate the Yop virulence proteins and V antigen upon contact with a eukaryotic cell. Yersinia pestis mutants defective for production of YscX or YscY were unable to export the Yops and V antigen. YscX and YscY were both present in the Y. pestis cell pellet fraction; however, YscX was also found in the culture supernatant. YscY showed structural and amino acid sequence similarities to the Syc family of proteins. YscY specifically recognized and bound to a region of YscX that included a predicted coiled-coil region. These data suggest that YscY may function as a chaperone for YscX in Y. pestis. PMID- 10714988 TI - Identification of two novel hrp-associated genes in the hrp gene cluster of Xanthomonas oryzae pv. oryzae. AB - We have cloned a hrp gene cluster from Xanthomonas oryzae pv. oryzae. Bacteria with mutations in the hrp region have reduced growth in rice leaves and lose the ability to elicit a hypersensitive response (HR) on the appropriate resistant cultivars of rice and the nonhost plant tomato. A 12,165-bp portion of nucleotide sequence from the presumed left end and extending through the hrpB operon was determined. The region was most similar to hrp genes from Xanthomonas campestris pv. vesicatoria and Ralstonia solanacearum. Two new hrp-associated loci, named hpa1 and hpa2, were located beyond the hrpA operon. The hpa1 gene encoded a 13 kDa glycine-rich protein with a composition similar to those of harpins and PopA. The product of hpa2 was similar to lysozyme-like proteins. Perfect PIP boxes were present in the hrpB and hpa1 operons, while a variant PIP box was located upstream of hpa2. A strain with a deletion encompassing hpa1 and hpa2 had reduced pathogenicity and elicited a weak HR on nonhost and resistant host plants. Experiments using single mutations in hpa1 and hpa2 indicated that the loss of hpa1 was the principal cause of the reduced pathogenicity of the deletion strain. A 1,519-bp insertion element was located immediately downstream of hpa2. Hybridization with hpa2 indicated that the gene was present in all of the strains of Xanthomonas examined. Hybridization experiments with hpa1 and IS1114 indicated that these sequences were detectable in all strains of X. oryzae pv. oryzae and some other Xanthomonas species. PMID- 10714989 TI - Capsule biosynthesis and basic metabolism in Streptococcus pneumoniae are linked through the cellular phosphoglucomutase. AB - Synthesis of the type 3 capsular polysaccharide of Streptococcus pneumoniae requires UDP-glucose (UDP-Glc) and UDP-glucuronic acid (UDP-GlcUA) for production of the [3)-beta-D-GlcUA-(1-->4)-beta-D-Glc-(1-->](n) polymer. The generation of UDP-Glc proceeds by conversion of Glc-6-P to Glc-1-P to UDP-Glc and is mediated by a phosphoglucomutase (PGM) and a Glc-1-P uridylyltransferase, respectively. Genes encoding both a Glc-1-P uridylyltransferase (cps3U) and a PGM homologue (cps3M) are present in the type 3 capsule locus, but these genes are not essential for capsule production. In this study, we characterized a mutant that produces fourfold less capsule than the type 3 parent. The spontaneous mutation resulting in this phenotype was not contained in the type 3 capsule locus but was instead located in a distant gene (pgm) encoding a second PGM homologue. The function of this gene product as a PGM was demonstrated through enzymatic and complementation studies. Insertional inactivation of pgm reduced capsule production to less than 10% of the parental level. The loss of PGM activity in the insertion mutants also caused growth defects and a strong selection for isolates containing second-site suppressor mutations. These results demonstrate that most of the PGM activity required for type 3 capsule biosynthesis is derived from the cellular PGM. PMID- 10714990 TI - Characterization of hydrogenase II from the hyperthermophilic archaeon Pyrococcus furiosus and assessment of its role in sulfur reduction. AB - The fermentative hyperthermophile Pyrococcus furiosus contains an NADPH utilizing, heterotetrameric (alphabetagammadelta), cytoplasmic hydrogenase (hydrogenase I) that catalyzes both H(2) production and the reduction of elemental sulfur to H(2)S. Herein is described the purification of a second enzyme of this type, hydrogenase II, from the same organism. Hydrogenase II has an M(r) of 320,000 +/- 20,000 and contains four different subunits with M(r)s of 52,000 (alpha), 39,000 (beta), 30,000 (gamma), and 24,000 (delta). The heterotetramer contained Ni (0.9 +/- 0.1 atom/mol), Fe (21 +/- 1.6 atoms/mol), and flavin adenine dinucleotide (FAD) (0.83 +/- 0.1 mol/mol). NADPH and NADH were equally efficient as electron donors for H(2) production with K(m) values near 70 microM and k(cat)/K(m) values near 350 min(-1) mM(-1). In contrast to hydrogenase I, hydrogenase II catalyzed the H(2)-dependent reduction of NAD (K(m), 128 microM; k(cat)/K(m), 770 min(-1) mM(-1)). Ferredoxin from P. furiosus was not an efficient electron carrier for either enzyme. Both H(2) and NADPH served as electron donors for the reduction of elemental sulfur (S(0)) and polysulfide by hydrogenase I and hydrogenase II, and both enzymes preferentially reduce polysulfide to sulfide rather than protons to H(2) using NADPH as the electron donor. At least two [4Fe-4S] and one [2Fe-2S] cluster were detected in hydrogenase II by electron paramagnetic resonance spectroscopy, but amino acid sequence analyses indicated a total of five [4Fe-4S] clusters (two in the beta subunit and three in the delta subunit) and one [2Fe-2S] cluster (in the gamma subunit), as well as two putative nucleotide-binding sites in the gamma subunit which are thought to bind FAD and NAD(P)(H). The amino acid sequences of the four subunits of hydrogenase II showed between 55 and 63% similarity to those of hydrogenase I. The two enzymes are present in the cytoplasm at approximately the same concentration. Hydrogenase II may become physiologically relevant at low S(0) concentrations since it has a higher affinity than hydrogenase I for both S(0) and polysulfide. PMID- 10714991 TI - Multiple factors independently regulate hilA and invasion gene expression in Salmonella enterica serovar typhimurium. AB - HilA activates the expression of Salmonella enterica serovar Typhimurium invasion genes. To learn more about regulation of hilA, we isolated Tn5 mutants exhibiting reduced hilA and/or invasion gene expression. In addition to expected mutations, we identified Tn5 insertions in pstS, fadD, flhD, flhC, and fliA. Analysis of the pstS mutant indicates that hilA and invasion genes are repressed by the response regulator PhoB in the absence of the Pst high-affinity inorganic phosphate uptake system. This system is required for negative control of the PhoR-PhoB two component regulatory system, suggesting that hilA expression may be repressed by PhoR-PhoB under low extracellular inorganic phosphate conditions. FadD is required for uptake and degradation of long-chain fatty acids, and our analysis of the fadD mutant indicates that hilA is regulated by a FadD-dependent, FadR independent mechanism. Thus, fatty acid derivatives may act as intracellular signals to regulate hilA expression. flhDC and fliA encode transcription factors required for flagellum production, motility, and chemotaxis. Complementation studies with flhC and fliA mutants indicate that FliZ, which is encoded in an operon with fliA, activates expression of hilA, linking regulation of hilA with motility. Finally, epistasis tests showed that PhoB, FadD, FliZ, SirA, and EnvZ act independently to regulate hilA expression and invasion. In summary, our screen has identified several distinct pathways that can modulate S. enterica serovar Typhimurium's ability to express hilA and invade host cells. Integration of signals from these different pathways may help restrict invasion gene expression during infection. PMID- 10714992 TI - The Bacillus subtilis yabG gene is transcribed by SigK RNA polymerase during sporulation, and yabG mutant spores have altered coat protein composition. AB - The expression of six novel genes located in the region from abrB to spoVC of the Bacillus subtilis chromosome was analyzed, and one of the genes, yabG, had a predicted promoter sequence conserved among SigK-dependent genes. Northern blot analysis revealed that yabG mRNA was first detected from 4 h after the cessation of logarithmic growth (T(4)) in wild-type cells and in a gerE36 (GerE(-)) mutant but not in spoIIAC (SigF(-)), spoIIGAB (SigE(-)), spoIIIG (SigG(-)), and spoIVCB (SigK(-)) mutants. The transcription start point was determined by primer extension analysis; the -10 and -35 regions are very similar to the consensus sequences recognized by SigK-containing RNA polymerase. Inactivation of the yabG gene by insertion of an erythromycin resistance gene did not affect vegetative growth or spore resistance to heat, chloroform, and lysozyme. The germination of yabG spores in L-alanine and in a mixture of L-asparagine, D-glucose, D-fructose, and potassium chloride was also the same as that of wild-type spores. On the other hand, the protein preparation from yabG spores included 15-, 18-, 21-, 23-, 31-, 45-, and 55-kDa polypeptides which were low in or not extracted from wild type spores under the same conditions. We determined their N-terminal amino acid sequence and found that these polypeptides were CotT, YeeK, YxeE, CotF, YrbA (31 and 45 kDa), and SpoIVA, respectively. The fluorescence of YabG-green fluorescent protein fusion produced in sporulating cells was detectable in the forespores but not in the mother cell compartment under fluorescence microscopy. These results indicate that yabG encodes a sporulation-specific protein which is involved in coat protein composition in B. subtilis. PMID- 10714993 TI - pTAR-encoded proteins in plasmid partitioning. AB - Partition cassettes, essential for the segregational stability of low-copy-number bacterial plasmids, typically encode two autoregulated proteins and an adjacent cis-acting centromere analog to which one or perhaps both proteins bind. The diminutive partition region of pTAR of Agrobacterium spp. was reported to be exceptional, encoding only a single protein, ParA (D. R. Gallie and C. I. Kado, J. Mol. Biol. 193:465-478, 1987). However, resequencing of the region revealed two small downstream genes, parB and orf-84, of which only parB was found to be essential for partitioning in A. tumefaciens. Purified ParA exhibited a weak ATPase activity that was modestly increased by nonspecific DNA. ParB bound in vitro to repeated sequences present in a region, parS, that possesses centromere and operator functions and within which we identified the primary transcription start site by primer extension. In certain respects the Par proteins behave normally in the foreign host Escherichia coli. In E. coli, as in A. tumefaciens, ParB repressed the partition operon; ParA, inactive alone, augmented this repression. Functional similarities between the partition system of pTAR and those of other plasmids and bacteria are prominent, despite differences in size, organization, and amino acid sequence. PMID- 10714995 TI - Purification and characterization of a catalase from the facultatively psychrophilic bacterium Vibrio rumoiensis S-1(T) exhibiting high catalase activity. AB - Catalase from the facultatively psychrophilic bacterium Vibrio rumoiensis S-1(T), which was isolated from an environment exposed to H(2)O(2) and exhibited high catalase activity, was purified and characterized, and its localization in the cell was determined. Its molecular mass was 230 kDa, and the molecule consisted of four identical subunits. The enzyme, which was not apparently reduced by dithionite, showed a Soret peak at 406 nm in a resting state. The catalytic activity was 527,500 U. mg of protein(-1) under standard reaction conditions at 40 degrees C, 1.5 and 4.3 times faster, respectively, than those of the Micrococcus luteus and bovine catalases examined under the same reaction conditions, and showed a broad optimum pH range (pH 6 to 10). The catalase from strain S-1(T) is located not only in the cytoplasmic space but also in the periplasmic space. There is little difference in the activation energy for the activity between strain S-1(T) catalase and M. luteus and bovine liver catalases. The thermoinstability of the activity of the former catalase were significantly higher than those of the latter catalases. The thermoinstability suggests that the catalase from strain S-1(T) should be categorized as a psychrophilic enzyme. Although the catalase from strain S-1(T) is classified as a mammal type catalase, it exhibits the unique enzymatic properties of high intensity of enzymatic activity and thermoinstability. The results obtained suggest that these unique properties of the enzyme are in accordance with the environmental conditions under which the microorganism lives. PMID- 10714994 TI - Characterization of an HPr kinase mutant of Staphylococcus xylosus. AB - The Staphylococcus xylosus gene hprK, encoding HPr kinase (HPrK), has been isolated from a genomic library. The HPrK enzyme, purified as a His(6) fusion protein, phosphorylated HPr, the phosphocarrier protein of the bacterial phosphotransferase system, at a serine residue in an ATP-dependent manner, and it also catalyzed the reverse reaction. Therefore, the enzyme constitutes a bifunctional HPr kinase/phosphatase. Insertional inactivation of the gene in the genome of S. xylosus resulted in the concomitant loss of both HPr kinase and His serine-phosphorylated-HPr phosphatase activities in cell extracts, strongly indicating that the HPrK enzyme is also responsible for both reactions in vivo. HPrK deficiency had a profound pleiotropic effect on the physiology of S. xylosus. The hprK mutant strain showed a severe growth defect in complex medium upon addition of glucose. Glucose uptake in glucose-grown cells was strongly enhanced compared with the wild type. Carbon catabolite repression of three tested enzyme activities by glucose, sucrose, and fructose was abolished. These results clearly demonstrate the prominent role of HPr kinase in global control to adjust catabolic capacities of S. xylosus according to the availability of preferred carbon sources. PMID- 10714996 TI - CelE, a multidomain cellulase from Clostridium cellulolyticum: a key enzyme in the cellulosome? AB - CelE, one of the three major proteins of the cellulosome of Clostridium cellulolyticum, was characterized. The amino acid sequence of the protein deduced from celE DNA sequence led us to the supposition that CelE is a three-domain protein. Recombinant CelE and a truncated form deleted of the putative cellulose binding domain (CBD) were obtained. Deletion of the CBD induces a total loss of activity. Exhibiting rather low levels of activity on soluble, amorphous, and crystalline celluloses, CelE is more active on p-nitrophenyl-cellobiose than the other cellulases from this organism characterized to date. The main product of its action on Avicel is cellobiose (more than 90% of the soluble sugars released), and its attack on carboxymethyl cellulose is accompanied by a relatively small decrease in viscosity. All of these features suggest that CelE is a cellobiohydrolase which has retained a certain capacity for random attack mode. We measured saccharification of Avicel and bacterial microcrystalline cellulose by associations of CelE with four other cellulases from C. cellulolyticum and found that CelE acts synergistically with all tested enzymes. The positive influence of CelE activity on the activities of other cellulosomal enzymes may explain its relative abundance in the cellulosome. PMID- 10714997 TI - Purification and characterization of the DeoR repressor of Bacillus subtilis. AB - Transcription of the Bacillus subtilis dra-nupC-pdp operon is repressed by the DeoR repressor protein. The DeoR repressor with an N-terminal His tag was overproduced with a plasmid under control of a phage T5 promoter in Escherichia coli and was purified to near homogeneity by one affinity chromatography step. Gel filtration experimental results showed that native DeoR has a mass of 280 kDa and appears to exist as an octamer. Binding of DeoR to the operator DNA of the dra-nupC-pdp operon was characterized by using an electrophoretic gel mobility shift assay. An apparent dissociation constant of 22 nM was determined for binding of DeoR to operator DNA, and the binding curve indicated that the binding of DeoR to the operator DNA was cooperative. In the presence of low-molecular weight effector deoxyribose-5-phosphate, the dissociation constant was higher than 1,280 nM. The dissociation constant remained unchanged in the presence of deoxyribose-1-phosphate. DNase I footprinting exhibited a protected region that extends over more than 43 bp, covering a palindrome together with a direct repeat to one half of the palindrome and the nucleotides between them. PMID- 10714998 TI - Distribution of intervening sequences in the genes for 23S rRNA and rRNA fragmentation among strains of the Salmonella reference collection B (SARB) and SARC sets. AB - Intervening sequences (IVSs) occur sporadically in several bacterial genera in the genes for 23S rRNA at relatively conserved locations. They are cleaved after transcription and lead to the presence of fragmented rRNA, which is incorporated into the ribosomes without religation but is nevertheless functional. The fragmentation of rRNA and the number of IVSs in all 72 strains of the Salmonella Reference Collection B set and 16 strains of the Salmonella Reference Collection C set, which have been established on the basis of multilocus enzyme electrophoresis (MLEE), were analyzed in the present study. Fragmentation of 23S rRNA was restricted to conserved cleavage sites located at bp 550 (helix 25) and bp 1170 (helix 45), locations where IVSs have been reported. Random cleavage at sites where IVSs could not be detected was not seen. Uncleaved IVSs were not detected in any case; thus, the IVSs invariably led to rRNA fragmentation, indicating a strong selection for maintenance of RNase III cleavage sites. The distribution of the number of IVSs carried by the different strains in the seven rrl genes is diverse, and the pattern of IVS possession could not be related to the MLEE pattern among the various Salmonella strains tested; this indicates that the IVSs are frequently exchanged between strains by lateral transfer. All eight subspecies of the genus Salmonella, including subspecies V represented by Salmonella bongori, have IVSs in both helix 25 and helix 45; this indicates that IVSs entered the genus after its divergence from Escherichia coli (more than 100 million years ago) but before separation of the genus Salmonella into many forms or that they were in the ancestor but have been lost from Escherichia. PMID- 10714999 TI - Spirochaeta aurantia has diacetyl chloramphenicol esterase activity. AB - The free-living spirochete Spirochaeta aurantia was nearly as susceptible to diacetyl chloramphenicol, the product of chloramphenicol acetyltransferase, as it was to chloramphenicol itself. This unexpected susceptibility to diacetyl chloramphenicol was wholly or partly the consequence of intrinsic carboxylesterase activity, as indicated by high-performance liquid chromatography, thin-layer chromatography, and microbiological assays. The esterase converted the diacetate to chloramphenicol, thus inhibiting spirochete growth. The esterase activity was cell associated, reduced by proteinase K, eliminated by boiling, and independent of the presence of either chloramphenicol or diacetyl chloramphenicol. S. aurantia extracts also hydrolyzed other esterase substrates, and two of these, alpha-napthyl acetate and 4-methylumbelliferyl acetate, identified an esterase of approximately 75 kDa in a nondenaturing gel. Carboxylesterases occur in Streptomyces species, but in this study their activity was weaker than that of S. aurantia. The S. aurantia esterase could reduce the effectiveness of cat as either a selectable marker or a reporter gene in this species. PMID- 10715000 TI - Sinorhizobium meliloti putA gene regulation: a new model within the family Rhizobiaceae. AB - Proline dehydrogenase (PutA) is a bifunctional enzyme that catalyzes the oxidation of proline to glutamate. In Sinorhizobium meliloti, as in other microorganisms, the putA gene is transcriptionally activated in response to proline. In Rhodobacter capsulatus, Agrobacterium, and most probably in Bradyrhizobium, this activation is dependent on an Lrp-like protein encoded by the putR gene, located immediately upstream of putA. Interestingly, sequence and genetic analysis of the region upstream of the S. meliloti putA gene did not reveal such a putR locus or any other encoded transcriptional activator of putA. Furthermore, results obtained with an S. meliloti putA null mutation indicate the absence of any proline-responsive transcriptional activator and that PutA serves as an autogenous repressor. Therefore, the model of S. meliloti putA regulation completely diverges from that of its Rhizobiaceae relatives and resembles more that of enteric bacteria. However, some differences have been found with the latter model: (i) S. meliloti putA gene is not catabolite repressed, and (ii) the gene encoding for the major proline permease (putP) does not form part of an operon with the putA gene. PMID- 10715001 TI - The Bacillus subtilis HBsu protein modifies the effects of alpha/beta-type, small acid-soluble spore proteins on DNA. AB - HBsu, the Bacillus subtilis homolog of the Escherichia coli HU proteins and the major chromosomal protein in vegetative cells of B. subtilis, is present at similar levels in vegetative cells and spores ( approximately 5 x 10(4) monomers/genome). The level of HBsu in spores was unaffected by the presence or absence of the alpha/beta-type, small acid-soluble proteins (SASP), which are the major chromosomal proteins in spores. In developing forespores, HBsu colocalized with alpha/beta-type SASP on the nucleoid, suggesting that HBsu could modulate alpha/beta-type SASP-mediated properties of spore DNA. Indeed, in vitro studies showed that HBsu altered alpha/beta-type SASP protection of pUC19 from DNase digestion, induced negative DNA supercoiling opposing alpha/beta-type SASP mediated positive supercoiling, and greatly ameliorated the alpha/beta-type SASP mediated increase in DNA persistence length. However, HBsu did not significantly interfere with the alpha/beta-type SASP-mediated changes in the UV photochemistry of DNA that explain the heightened resistance of spores to UV radiation. These data strongly support a role for HBsu in modulating the effects of alpha/beta type SASP on the properties of DNA in the developing and dormant spore. PMID- 10715002 TI - A 90-kilobase conjugative chromosomal element coding for biphenyl and salicylate catabolism in Pseudomonas putida KF715. AB - The biphenyl and salicylate metabolic pathways in Pseudomonas putida KF715 are chromosomally encoded. The bph gene cluster coding for the conversion of biphenyl to benzoic acid and the sal gene cluster coding for the salicylate meta-pathway were obtained from the KF715 genomic cosmid libraries. These two gene clusters were separated by 10-kb DNA and were highly prone to deletion when KF715 was grown in nutrient medium. Two types of deletions took place at the region including only the bph genes (ca. 40 kb) or at the region including both the bph and sal genes (ca. 70 kb). A 90-kb DNA region, including both the bph and sal genes (termed the bph-sal element), was transferred by conjugation from KF715 to P. putida AC30. Such transconjugants gained the ability to grow on biphenyl and salicylate as the sole sources of carbon. The bph and sal element was located on the chromosome of the recipient. The bph-sal element in strain AC30 was also highly prone to deletion; however, it could be mobilized to the chromosome of P. putida KT2440 and the two deletion mutants of KF715. PMID- 10715003 TI - Characterization of the gene cluster involved in isoprene metabolism in Rhodococcus sp. strain AD45. AB - The genes involved in isoprene (2-methyl-1,3-butadiene) utilization in Rhodococcus sp. strain AD45 were cloned and characterized. Sequence analysis of an 8.5-kb DNA fragment showed the presence of 10 genes of which 2 encoded enzymes which were previously found to be involved in isoprene degradation: a glutathione S-transferase with activity towards 1,2-epoxy-2-methyl-3-butene (isoI) and a 1 hydroxy-2-glutathionyl-2-methyl-3-butene dehydrogenase (isoH). Furthermore, a gene encoding a second glutathione S-transferase was identified (isoJ). The isoJ gene was overexpressed in Escherichia coli and was found to have activity with 1 chloro-2,4-dinitrobenzene and 3,4-dichloro-1-nitrobenzene but not with 1, 2-epoxy 2-methyl-3-butene. Downstream of isoJ, six genes (isoABCDEF) were found; these genes encoded a putative alkene monooxygenase that showed high similarity to components of the alkene monooxygenase from Xanthobacter sp. strain Py2 and other multicomponent monooxygenases. The deduced amino acid sequence encoded by an additional gene (isoG) showed significant similarity with that of alpha methylacyl-coenzyme A racemase. The results are in agreement with a catabolic route for isoprene involving epoxidation by a monooxygenase, conjugation to glutathione, and oxidation of the hydroxyl group to a carboxylate. Metabolism may proceed by fatty acid oxidation after removal of glutathione by a still-unknown mechanism. PMID- 10715004 TI - Repair of DNA lesions induced by hydrogen peroxide in the presence of iron chelators in Escherichia coli: participation of endonuclease IV and Fpg. AB - In Escherichia coli, the repair of lethal DNA damage induced by H(2)O(2) requires exonuclease III, the xthA gene product. Here, we report that both endonuclease IV (the nfo gene product) and exonuclease III can mediate the repair of lesions induced by H(2)O(2) under low-iron conditions. Neither the xthA nor the nfo mutants was sensitive to H(2)O(2) in the presence of iron chelators, while the xthA nfo double mutant was significantly sensitive to this treatment, suggesting that both exonuclease III and endonuclease IV can mediate the repair of DNA lesions formed under such conditions. Sedimentation studies in alkaline sucrose gradients also demonstrated that both xthA and nfo mutants, but not the xthA nfo double mutant, can carry out complete repair of DNA strand breaks and alkali labile bonds generated by H(2)O(2) under low-iron conditions. We also found indications that the formation of substrates for exonuclease III and endonuclease IV is mediated by the Fpg DNA glycosylase, as suggested by experiments in which the fpg mutation increased the level of cell survival, as well as repair of DNA strand breaks, in an AP endonuclease-null background. PMID- 10715005 TI - Regulation of rRNA transcription is remarkably robust: FIS compensates for altered nucleoside triphosphate sensing by mutant RNA polymerases at Escherichia coli rrn P1 promoters. AB - We recently identified Escherichia coli RNA polymerase (RNAP) mutants (RNAP beta' Delta215-220 and beta RH454) that form extremely unstable complexes with rRNA P1 (rrn P1) core promoters. The mutant RNAPs reduce transcription and alter growth rate-dependent regulation of rrn P1 core promoters, because the mutant RNAPs require higher concentrations of the initiating nucleoside triphosphate (NTP) for efficient transcription from these promoters than are present in vivo. Nevertheless, the mutants grow almost as well as wild-type cells, suggesting that rRNA synthesis is not greatly perturbed. We report here that the rrn transcription factor FIS activates the mutant RNAPs more strongly than wild-type RNAP, thereby compensating for the altered properties of the mutant RNAPs. FIS activates the mutant RNAPs, at least in part, by reducing the apparent K(ATP) for the initiating NTP. This and other results suggest that FIS affects a step in transcription initiation after closed-complex formation in addition to its stimulatory effect on initial RNAP binding. FIS and NTP levels increase with growth rate, suggesting that changing FIS concentrations, in conjunction with changing NTP concentrations, are responsible for growth rate-dependent regulation of rrn P1 transcription in the mutant strains. These results provide a dramatic demonstration of the interplay between regulatory mechanisms in rRNA transcription. PMID- 10715006 TI - Characterization of the uup locus and its role in transposon excisions and tandem repeat deletions in Escherichia coli. AB - Null mutations in the Escherichia coli uup locus (at 21.8 min) serve to increase the frequency of RecA-independent precise excision of transposable elements such as Tn10 and to reduce the plaque size of bacteriophage Mu (Uup(-) phenotype). By the combined approaches of physical mapping of the mutations, complementation analyses, and protein overexpression from cloned gene fragments, we have demonstrated in this study that the Uup(-) phenotype is the consequence of the absence of expression of the downstream gene (uup) of a two-gene operon, caused either directly by insertions in uup or indirectly by the polar effect of insertions in the upstream gene (ycbY). The promoter for uup was mapped upstream of ycbY by primer extension analysis on cellular RNA, and assays of reporter gene expression indicated that it is a moderately active, constitutive promoter. The uup mutations were also shown to increase, in a RecA-independent manner, the frequencies of nearly precise excision of Tn10 derivatives and of the deletion of one copy of a chromosomal tandem repeat, suggesting the existence of a shared step or intermediate in the pathways of these latter events and that of precise excision. Finally, we found that mutations that increase the frequency of precise excision of Tn10 are divisible into two categories, depending upon whether they did (uup, ssb, polA, and topA) or did not (mutHLS, dam, and uvrD) also increase precise excision frequency of the mini-Tn10 derivatives. It is suggested that the differential response of mini-Tn10 and Tn10 to the second category of mutations is related to the presence, respectively, of perfect and of imperfect terminal inverted repeats in them. PMID- 10715007 TI - Mutations in the gerP locus of Bacillus subtilis and Bacillus cereus affect access of germinants to their targets in spores. AB - The gerP1 transposon insertion mutation of Bacillus cereus is responsible for a defect in the germination response of spores to both L-alanine and inosine. The mutant is blocked at an early stage, before loss of heat resistance or release of dipicolinate, and the efficiency of colony formation on nutrient agar from spores is reduced fivefold. The protein profiles of alkaline-extracted spore coats and the spore cortex composition are unchanged in the mutant. Permeabilization of gerP mutant spores by coat extraction procedures removes the block in early stages of germination, although a consequence of the permeabilization procedure in both wild type and mutant is that late germination events are not complete. The complete hexacistronic operon that includes the site of insertion has been cloned and sequenced. Four small proteins encoded by the operon (GerPA, GerPD, GerPB, and GerPF) are related in sequence. A homologous operon (yisH-yisC) can be found in the Bacillus subtilis genome sequence; null mutations in yisD and yisF, constructed by integrational inactivation, result in a mutant phenotype similar to that seen in B. cereus, though somewhat less extreme and equally repairable by spore permeabilization. Normal rates of germination, as estimated by loss of heat resistance, are also restored to a gerP mutant by the introduction of a cotE mutation, which renders the spore coats permeable to lysozyme. The B. subtilis operon is expressed solely during sporulation, and is sigma K-inducible. We hypothesize that the GerP proteins are important as morphogenetic or structural components of the Bacillus spore, with a role in the establishment of normal spore coat structure and/or permeability, and that failure to synthesize these proteins during spore formation limits the opportunity for small hydrophilic organic molecules, like alanine or inosine, to gain access to their normal target, the germination receptor, in the spore. PMID- 10715008 TI - Cooperative action of the catabolite activator protein and AraC in vitro at the araFGH promoter. AB - Full activation of transcription of the araFGH promoter, p(FGH), requires both the catabolite activator protein (CAP) and AraC protein. At p(FGH), the binding site for CAP is centered at position -41.5, an essential binding site for AraC is centered at position -79.5, and a second, nonessential binding site is centered at position -154.5. In this work, we used the minimal promoter region required for in vivo activation of p(FGH) to examine the roles of CAP and AraC in stimulating formation of open complexes at p(FGH). Migration retardation assays of open complexes showed that RNA polymerase binds exceptionally tightly to the AraC-CAP-p(FGH) complex and that the order of addition of proteins to the initiating complex is important. Similar assays with RNA polymerase containing truncated alpha subunits suggest that AraC interacts with the C-terminal domain of the alpha subunit. Finally, AraC protein also acts to prevent the improper binding of RNA polymerase at a pseudo promoter near the true p(FGH) promoter. PMID- 10715009 TI - Pyruvate kinase of the hyperthermophilic crenarchaeote Thermoproteus tenax: physiological role and phylogenetic aspects. AB - Pyruvate kinase (PK; EC 2.7.1.40) of Thermoproteus tenax was purified to homogeneity, and its coding gene was cloned and expressed in Escherichia coli. It represents a homomeric tetramer with a molecular mass of 49 kDa per subunit. PK exhibits positive binding cooperativity with respect to phosphoenolpyruvate and metal ions such as Mg(2+) and Mn(2+). Heterotropic effects, as commonly found for PKs from bacterial and eucaryal sources, could not be detected. The enzyme does not depend on K(+) ions. Heterotrophically grown cells exhibit specific activity of PK four times higher than autotrophically grown cells. Since the mRNA level of the PK coding gene is also accordingly higher in heterotrophic cells, we conclude that the PK activity is adjusted to growth conditions mainly on the transcript level. The enzymic properties of the PK and the regulation of its expression are discussed with respect to the physiological framework given by the T. tenax specific variant of the Embden-Meyerhof-Parnas pathway. T. tenax PK shows moderate overall sequence similarity (25 to 40% identity) to its bacterial and eucaryal pendants. Phylogenetic analyses of the known PK sequences result in a dichotomic tree topology that divides the enzymes into two major PK clusters, probably diverged by an early gene duplication event. The phylogenetic divergence is paralleled by a striking phenotypic differentiation of PKs: PKs of cluster I, which occur in eucaryal cytoplasm, some gamma proteobacteria, and low-GC gram positive bacteria, are only active in the presence of fructose-1,6-bisphosphate or other phosphorylated sugars, whereas PKs of cluster II, found in various bacterial phyla, plastids, and in Archaea, show activity without effectors but are commonly regulated by the energy charge of the cell. PMID- 10715010 TI - Kinetic analysis of Clostridium cellulolyticum carbohydrate metabolism: importance of glucose 1-phosphate and glucose 6-phosphate branch points for distribution of carbon fluxes inside and outside cells as revealed by steady state continuous culture. AB - During the growth of Clostridium cellulolyticum in chemostat cultures with ammonia as the growth-limiting nutrient, as much as 30% of the original cellobiose consumed by C. cellulolyticum was converted to cellotriose, glycogen, and polysaccharides regardless of the specific growth rates. Whereas the specific consumption rate of cellobiose and of the carbon flux through glycolysis increased, the carbon flux through the phosphoglucomutase slowed. The limitation of the path through the phosphoglucomutase had a great effect on the accumulation of glucose 1-phosphate (G1P), the precursor of cellotriose, exopolysaccharides, and glycogen. The specific rates of biosynthesis of these compounds are important since as much as 16.7, 16.0, and 21.4% of the specific rate of cellobiose consumed by the cells could be converted to cellotriose, exopolysaccharides, and glycogen, respectively. With the increase of the carbon flux through glycolysis, the glucose 6-phosphate (G6P) pool decreased, whereas the G1P pool increased. Continuous culture experiments showed that glycogen biosynthesis was associated with rapid growth. The same result was obtained in batch culture, where glycogen biosynthesis reached a maximum during the exponential growth phase. Glycogen synthesis in C. cellulolyticum was also not subject to stimulation by nutrient limitation. Flux analyses demonstrate that G1P and G6P, connected by the phosphoglucomutase reaction, constitute important branch points for the distribution of carbon fluxes inside and outside cells. From this study it appears that the properties of the G1P-G6P branch points have been selected to control excretion of carbon surplus and to dissipate excess energy, whereas the pyruvate-acetyl coenzyme A branch points chiefly regulate the redox balance of the carbon catabolism as was shown previously (E. Guedon et al., J. Bacteriol. 181:3262-3269, 1999). PMID- 10715011 TI - sal genes determining the catabolism of salicylate esters are part of a supraoperonic cluster of catabolic genes in Acinetobacter sp. strain ADP1. AB - A 5-kbp region upstream of the are-ben-cat genes was cloned from Acinetobacter sp. strain ADP1, extending the supraoperonic cluster of catabolic genes to 30 kbp. Four open reading frames, salA, salR, salE, and salD, were identified from the nucleotide sequence. Reverse transcription-PCR studies suggested that these open reading frames are organized into two convergent transcription units, salAR and salDE. The salE gene, encoding a protein of 239 residues, was ligated into expression vector pET5a. Its product, SalE, was shown to have esterase activity against short-chain alkyl esters of 4-nitrophenol but was also able to hydrolyze ethyl salicylate to ethanol and salicylic acid. A mutant of ADP1 with a Km(r) cassette introduced into salE had lost the ability to utilize only ethyl and methyl salicylates of the esters tested as sole carbon sources, and no esterase activity against ethyl salicylate could be detected in cell extracts. SalE was induced during growth on ethyl salicylate but not during growth on salicylate itself. salD encoded a protein of undetermined function with homologies to the Escherichia coli FadL membrane protein, which is involved in facilitating fatty acid transport, and a number of other proteins detected during aromatic catabolism, which may also function in hydrocarbon transport or uptake processes. A Km(r) cassette insertion in salD deleteriously affected cell growth and viability. The salA and salR gene products closely resemble two Pseudomonas proteins, NahG and NahR, respectively encoding salicylate hydroxylase and the LysR family regulator of both salicylate and naphthalene catabolism. salA was cloned into pUC18 together with salR and salE, and its gene product showed salicylate-inducible hydroxylase activity against a range of substituted salicylates, with the same relative specific activities as found in wild-type ADP1 grown on salicylate. Mutations involving insertion of Km(r) cassettes into salA and salR eliminated expression of salicylate hydroxylase activity and the ability to grow on either salicylate or ethyl salicylate. Studies of mutants with disruptions of genes of the beta-ketoadipate pathway with or without an additional salE mutation confirmed that ethyl salicylate and salicylate were channeled into the beta-ketoadipate pathway at the level of catechol and thence dissimilated by the cat gene products. SalR appeared to regulate expression of salA but not salE. PMID- 10715012 TI - Isolation and characterization of vicH, encoding a new pleiotropic regulator in Vibrio cholerae. AB - During the last decade, the hns gene and its product, the H-NS protein, have been extensively studied in Escherichia coli. H-NS-like proteins seem to be widespread in gram-negative bacteria. However, unlike in E. coli and in Salmonella enterica serovar Typhimurium, little is known about their role in the physiology of those organisms. In this report, we describe the isolation of vicH, an hns-like gene in Vibrio cholerae, the etiological agent of cholera. This gene was isolated from a V. cholerae genomic library by complementation of different phenotypes associated with an hns mutation in E. coli. It encodes a 135-amino-acid protein showing approximately 50% identity with both H-NS and StpA in E. coli. Despite a low amino acid conservation in the N-terminal part, VicH is able to cross-react with anti-H-NS antibodies and to form oligomers in vitro. The vicH gene is expressed as a single gene from two promoters in tandem and is induced by cold shock. A V. cholerae wild-type strain expressing a vicHDelta92 gene lacking its 3' end shows pleiotropic alterations with regard to mucoidy and salicin metabolism. Moreover, this strain is unable to swarm on semisolid medium. Similarly, overexpression of the vicH wild-type gene results in an alteration of swarming behavior. This suggests that VicH could be involved in the virulence process in V. cholerae, in particular by affecting flagellum biosynthesis. PMID- 10715013 TI - Dephosphorylation of the Escherichia coli transcriptional antiterminator BglG by the sugar sensor BglF is the reversal of its phosphorylation. AB - The Escherichia coli BglF protein catalyzes transport and phosphorylation of beta glucosides. In addition, BglF is a membrane sensor which reversibly phosphorylates the transcriptional regulator BglG, depending on beta-glucoside availability. Therefore, BglF has three enzymatic activities: beta-glucoside phosphotransferase, BglG phosphorylase, and phospho-BglG (BglG-P) dephosphorylase. Cys-24 of BglF is the active site which delivers the phosphoryl group either to the sugar or to BglG. To characterize the dephosphorylase activity, we asked whether BglG-P can give the phosphoryl group back to Cys-24 of BglF. Here we provide evidence which is consistent with the interpretation that Cys-24-P is an intermediate in the BglG-P dephosphorylation reaction. Hence, the dephosphorylation reaction catalyzed by BglF proceeds via reversal of the phosphorylation reaction. PMID- 10715014 TI - Analysis of the bmp gene family in Borrelia burgdorferi sensu lato. AB - BmpA, BmpB, BmpC, and BmpD are homologous Borrelia burgdorferi lipoproteins of unknown functions, encoded by the bmp genes of paralogous chromosomal gene family 36. At least some of the Bmp proteins are immunogens in infected vertebrate hosts. The genetic organization of the bmp region has been characterized for a variety of B. burgdorferi sensu lato strains by Southern hybridization, PCR amplification, and DNA sequencing. All four bmp genes were present in the same relative order in all B. burgdorferi sensu lato low- and high-passage-number isolates. While there were no differences in the relative orders of the bmp genes in these species, variations in DNA sequence in the bmpD-bmpC and bmpC-bmpA intergenic regions were significantly more common than in the corresponding 3' bmpD and bmpC coding regions. The genetic structure of the chromosomal region containing the bmp genes thus appears to be well conserved across different species of B. burgdorferi, but variations in DNA fine structure that prevent PCR primer annealing may occur in this region and make Southern hybridization much more reliable than PCR for detection of the presence of these genes. Our results also suggest that bmp gene products may be used as reagents in the preparation of vaccines and diagnostic assays to protect against and diagnose Lyme disease produced by B. burgdorferi sensu lato. PMID- 10715016 TI - Disruption of the Borrelia burgdorferi gac gene, encoding the naturally synthesized GyrA C-terminal domain. AB - The C-terminal domain of the A subunit of DNA gyrase, which we term Gac, is naturally synthesized in Borrelia burgdorferi as an abundant DNA-binding protein. Full-length GyrA, which includes the C-terminal domain, is also synthesized by the spirochete and functions as a subunit of DNA gyrase. We have disrupted synthesis of Gac as an independent protein and demonstrated that it is not essential for growth in a coumarin-resistant background. We detected no alterations in DNA maintenance, condensation, or topology in B. burgdorferi lacking this small DNA-binding protein. PMID- 10715015 TI - Mobilization of plasmids and chromosomal DNA mediated by the SXT element, a constin found in Vibrio cholerae O139. AB - The Vibrio cholerae SXT element encodes resistance to multiple antibiotics and is a conjugative, self-transmissible, and chromosomally integrating element (a constin). Excision and self-transfer of the SXT element require an element encoded integrase. We now report that the SXT element can also mobilize the plasmids RSF1010 and CloDF13 in trans as well as chromosomal DNA in an Hfr-like manner. SXT element-mediated mobilization of plasmids and chromosomal DNA, unlike its self-transfer, is not dependent upon excision of the element from the chromosome. These results raise the possibility that the SXT element and other constins play a general role in horizontal gene transfer among gram-negative bacteria. PMID- 10715017 TI - Purification and properties of ornithine racemase from Clostridium sticklandii. AB - Ornithine racemase has been purified to homogeneity from Clostridium sticklandii, as shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This is the first racemase known to be highly specific to ornithine. This PLP-dependent enzyme has an M(r) of 92, 000, with a K(m) for L-ornithine of 0.77 +/- 0.05 mM and a k(cat) of 980 +/- 20 s(-1). PMID- 10715018 TI - The sulfur-regulated arylsulfatase gene cluster of Pseudomonas aeruginosa, a new member of the cys regulon. AB - A gene cluster upstream of the arylsulfatase gene (atsA) in Pseudomonas aeruginosa was characterized and found to encode a putative ABC-type transporter, AtsRBC. Mutants with insertions in the atsR or atsB gene were unable to grow with hexyl-, octyl-, or nitrocatecholsulfate, although they grew normally with other sulfur sources, such as sulfate, methionine, and aliphatic sulfonates. AtsRBC therefore constitutes a general sulfate ester transport system, and desulfurization of aromatic and medium-chain-length aliphatic sulfate esters occurs in the cytoplasm. Expression of the atsR and atsBCA genes was repressed during growth with sulfate, cysteine, or thiocyanate. No expression of these genes was observed in the cysB mutant PAO-CB, and the ats genes therefore constitute an extension of the cys regulon in this species. PMID- 10715019 TI - Colorectal cancer imaging: a challenge for radiologists. PMID- 10715020 TI - Reflections on the early years of nuclear medicine. AB - In the early years of nuclear medicine, physicians explored applied nuclear physics, and physicists pursued uncharted areas in medicine. Reflections from Jim Adelstein, MD, PhD, John McAfee, MD, Henry Wagner, MD, Fred Bonte, MD, Dave Kuhl, MD, and Alex Gottschalk, MD, add to the appreciation of the diversity in those early years. These reflections may serve many purposes. For some, they may provoke nostalgia for the better life gone by. For others, reflections may create an awareness of the people and the process of what it took to be where we are today. For still others, this may provide some impetus to better understand the origins of modern imaging technologies and their diffusion. Which techniques in use today will be in use 30 years from now? Why will some survive and others go by the wayside? From research into the process of technology transfer and diffusion, can we learn to put our efforts today where they will have the greatest benefits to human beings some 30 years from now? How can we maximize the present value of our efforts to improve diagnostic imaging? Reflections from the past may help. PMID- 10715021 TI - The Fleischner Society: a 30th anniversary retrospective. PMID- 10715022 TI - Age-related accuracy of screening mammography: how should it be measured? PMID- 10715023 TI - The future of graduate medical education: summary of the proceedings of the American Board of Radiology seminar, March 5 and 6, 1998. PMID- 10715024 TI - MR coronary angiography: are we there yet? PMID- 10715025 TI - MR imaging evaluation of seizures. AB - It is imperative for a radiologist to determine the type of seizure a patient has prior to magnetic resonance (MR) imaging to optimally provide the clinician with the information he or she requires. Specifically, complex partial seizures require evaluation of the frontal lobes and the hippocampus (for mesial temporal sclerosis). These are best evaluated with fluid-attenuated inversion recovery (FLAIR) imaging; the use of intravenously administered contrast material is not required. Other types of chronic seizures are best evaluated with nonenhanced FLAIR or T2-weighted imaging for low-grade tumors, vascular malformations, gliosis after infarction, inflammation, or trauma. The presence of new-onset seizures in an adult or the worsening of chronic seizures warrants T2-weighted or FLAIR imaging and gadolinium-enhanced T1-weighted imaging (to look for primary or metastatic tumors, infections, or inflammatory lesions). If available, echo planar diffusion imaging should be used also (to look for acute infarcts). PMID- 10715026 TI - Cerebrospinal fluid cleft with cortical dimple: MR imaging marker for focal cortical dysgenesis. AB - PURPOSE: To determine whether the magnetic resonance (MR) imaging feature of a cerebrospinal fluid (CSF) cleft and cortical dimple can be used as a marker for cortical dysgenesis. MATERIALS AND METHODS: MR images in 875 patients with epilepsy were evaluated for the following features of focal cortical dysgenesis: cortical thickening, indistinct junction between gray and white matter, macrogyria, and abnormal sulcal pattern. Images with these features were reevaluated to determine the relationship between the CSF cleft-cortical dimple complex and focal cortical dysgenesis and its contribution to diagnosis. The cleft-dimple complex was defined as a prominent CSF space associated with a region of cortical volume loss adjacent to the dysgenesis. RESULTS: Seventy-one patients had cortical dysgenesis, including 27 with cellular proliferation abnormalities, 18 with migration abnormalities, 25 with cortical organization abnormalities, and one with miscellaneous anomalies. Histologic correlation was available in 20 patients. There was an associated cortical dimple in 29 (41%) patients. This association was strongest in patients with cortical organization abnormalities: It was present in 22 of the 25 (88%) patients. In 12 (48%) patients with abnormalities of cortical organization, the CSF cleft was easier to detect than other features of cortical dysgenesis or contributed directly to the MR imaging diagnosis. CONCLUSION: The cleft-dimple complex is a marker for subtle cases of focal cortical dysgenesis and may be due to maldevelopment. In patients with seizures, the presence of a cleft-dimple complex should alert the physician to scrutinize adjacent regions for developmental anomalies. PMID- 10715027 TI - Brain atrophy in relapsing-remitting multiple sclerosis and secondary progressive multiple sclerosis: longitudinal quantitative analysis. AB - PURPOSE: To determine annual rates of volumetric changes in the whole-brain parenchyma of patients with relapsing-remitting and secondary progressive multiple sclerosis (MS) and test the hypothesis that these changes correlate with clinical disability. MATERIALS AND METHODS: A computer-assisted segmentation technique with thin-section magnetic resonance (MR) imaging was used in 36 patients with MS (27 relapsing-remitting, nine secondary progressive) and in 20 control subjects to quantify brain and cerebrospinal fluid volumes. To determine the degree of brain atrophy, the percentage brain parenchyma volume (PBV) relative to that of intracranial contents was calculated. RESULTS: At the beginning of the study, the PBV was smaller in the MS group than in the control group (P = .007); brain parenchyma volumes were similar. The median rate of brain volume loss was 17.3 mL per year in patients with relapsing-remitting MS and 23.6 mL per year in those with secondary progressive MS. There was a negative correlation between brain atrophy and Expanded Disability Status Scale (EDSS) score in patients with secondary progressive MS (r = -0.69, P = .004) and no correlation in patients with relapsing-remitting MS. T2 lesion volume did not correlate with brain atrophy in either group. CONCLUSION: The correlation between brain atrophy and EDSS score was better in patients with secondary progressive MS than in those with relapsing-remitting MS. PMID- 10715028 TI - MR imaging and histologic features of subinsular bright spots on T2-weighted MR images: Virchow-Robin spaces of the extreme capsule and insular cortex. AB - PURPOSE: To determine the cause and frequency of high-signal-intensity foci detected in the insular cortex and extreme capsule on thin-section, high-spatial resolution, coronal, T2-weighted magnetic resonance (MR) images. MATERIALS AND METHODS: The authors assessed high-signal-intensity areas in the insular cortex and extreme capsule on coronal MR images obtained in 56 patients with seizure and five control subjects. Images were obtained with thin-section, high-spatial resolution, T2-weighted, fast spin-echo; three-dimensional, spoiled gradient recalled-echo; and fluid-attenuated inversion-recovery sequences. In two formalin fixed brain specimens, MR imaging findings were correlated with gross anatomic and histologic findings. RESULTS: Subinsular bright spots were found in 53 of the 56 (95%) patients (96 of 112 [86%] hemispheres) and all five control subjects. The spots were elliptical in 30 patients, round in 14 patients, linear in 22 patients, and dotlike in seven patients and often had a featherlike configuration. The spots were isointense to cerebrospinal fluid on T2-weighted, fast SE images and were located in the anterior extreme capsule white matter and insular cortex. MR imaging of brain specimens revealed bilateral elliptical areas of high signal intensity that corresponded to small multiple cavities at gross anatomic inspection. At microscopic examination, these cavities were perivascular spaces of mostly arteriolar origin. CONCLUSION: High-signal-intensity subinsular foci at MR imaging are due to enlarged perivascular spaces. In most cases, these foci can be visualized on thin-section, high-spatial-resolution, coronal T2 weighted images; they should not be mistaken for pathologic conditions when they occur unilaterally. PMID- 10715029 TI - MR angiography of the intracranial venous system. AB - PURPOSE: To compare the effectiveness of different imaging planes at time-of flight (TOF) magnetic resonance (MR) angiography and phase-contrast MR angiography in the visualization of the normal intracranial venous system. MATERIALS AND METHODS: In 12 healthy volunteers, two-dimensional (2D) TOF MR angiography and three-dimensional (3D) phase-contrast MR angiography were performed in transverse, sagittal, and coronal planes. All data were displayed as maximum intensity projection (MIP) images. Four neuroradiologists assessed the visibility of 28 intracranial venous structures on the MIP images. Statistical analysis was performed by using the Friedman two-way analysis of variance and the Cochran Q test. RESULTS: Visualization of the normal intracranial venous system was better with 3D phase-contrast and coronal 2D TOF MR angiography than with transverse or sagittal 2D TOF MR angiography (P < .05, Friedman test) for each observer and the group of observers. Differences were found between each of the 2D TOF and 3D phase-contrast MR angiographic sequences in the visualization of individual venous structures (Cochran Q test). The kappa values ranged from 0.36 to 0.71, which indicated a moderate to good agreement between observers. CONCLUSION: The normal intracranial venous system is adequately visualized with 3D phase-contrast and coronal 2D TOF MR angiography. PMID- 10715030 TI - Laryngeal or hypopharyngeal squamous cell carcinoma: can follow-up CT after definitive radiation therapy be used to detect local failure earlier than clinical examination alone? AB - PURPOSE: To determine if follow-up computed tomography (CT) after definitive radiation therapy for laryngeal or hypopharyngeal (laryngopharyngeal) carcinoma allows the detection of local failure earlier than clinical examination alone. MATERIALS AND METHODS: Pre- and post-radiation therapy follow-up CT scans in 66 patients were reviewed retrospectively. All patients underwent definitive hyperfractionated radiation therapy and were followed up clinically for at least 2 years after its completion. Post-radiation therapy CT scans (N = 153) were evaluated for posttreatment changes with a three-point score: A score of 1 represented expected posttreatment changes; 2, focal mass with a maximal diameter of less than 1 cm and/or asymmetric obliteration of laryngeal tissue planes; or 3, focal mass with a maximal diameter equal to or greater than 1 cm or estimated tumor volume reduction of less than 50%. All patients underwent the first posttreatment CT study 1-6 months after therapy. New or progressive laryngeal cartilage changes were noted. The clinical impression of the larynx at the time of each follow-up CT scan was also recorded. RESULTS: In 12 of 29 (41%) patients with treatment failure at the primary site, follow-up CT scans were definite for local failure (score, 3) a mean of 5.5 months (median, 3.5 months; range, 1-17 months) before clinical examination results. CONCLUSION: In many patients, follow up CT shows local failure earlier than does clinical examination alone. PMID- 10715031 TI - Unresectable primary and recurrent head and neck tumors: effect of hyperthermia and carboplatin--preliminary experience. AB - PURPOSE: To perform a single-arm study to determine the effectiveness of and potential toxic reactions to local hyperthermia and systemic carboplatin (cis diammine-1,1-cyclobutane dicarboxylate platinum II) for the treatment of advanced or recurrent squamous cell carcinomas of the head and neck. MATERIALS AND METHODS: Eight patients with squamous cell carcinoma of the head and neck and stage IV disease (N2 or N3 neck adenopathy) or recurrent local-regional disease and who were previously and definitively treated were included in the study. Thermochemotherapy was administered every 4 weeks. Recorded end points were tumor response, duration of response, incidence of distant metastases, survival, cause of death, and toxic reactions. RESULTS: One patient had a complete response to therapy, and two had a partial response. Five patients had no response or developed progressive disease during therapy. Six patients died after 4-13 months of progressive disease. Two long-term survivors received radiation therapy; one also underwent surgical resection for residual neck disease. Each thermochemotherapeutic session was well tolerated, with minimal discomfort. Toxic reactions included hypotension, vomiting, hyponatremia, anemia, thrombocytopenia, and infection at the site of administration. There were no life-threatening toxic reactions. CONCLUSION: The combined use of hyperthermia and carboplatin shows potential in the management of unresectable head and neck tumors and is safe and well tolerated. Further studies on thermochemotherapy are warranted to assess its potential. PMID- 10715033 TI - Tear of the peroneus longus tendon: MR imaging features in nine patients. AB - PURPOSE: To determine the magnetic resonance (MR) imaging features that characterize tear of the peroneus longus tendon at the midfoot. MATERIALS AND METHODS: Medical records and MR images in nine patients with a tear of the middle segment of the peroneus longus tendon were retrospectively reviewed. All nine patients had undergone routine ankle MR imaging; three had undergone additional oblique coronal MR imaging. Surgical proof of a tear was available for three patients. RESULTS: Partial tear was present in four patients, and complete tear was present in five. Partial tears were characterized by heterogeneous signal intensity and thickening of the tendon. Complete tears were characterized by discontinuity of the tendon. Additional findings included fluid in the tendon sheath (n = 6), marrow edema of the lateral calcaneal wall (n = 3), enlarged peroneal tubercle (n = 3), and tear of the peroneus brevis tendon (n = 2). The extent of the tear was better assessed with oblique coronal MR images. CONCLUSION: The characteristic MR imaging appearance of complete or partial tear of the middle portion of the peroneus longus tendon includes foci of increased signal intensity in the distal tendon, morphologic alterations, and/or discontinuity of tendon. Bone marrow edema along the lateral calcaneal wall may be suggestive of the diagnosis. Additional oblique coronal midfoot MR images may help in assessment of the extent of the tear. PMID- 10715032 TI - Can imaging findings help differentiate spinal neuropathic arthropathy from disk space infection? Initial experience. AB - PURPOSE: To determine if radiographic, computed tomographic (CT), and magnetic resonance (MR) imaging findings can help differentiate spinal neuropathic arthropathy from disk space infection. MATERIALS AND METHODS: Imaging studies in 33 patients were evaluated, including 14 patients with spinal neuropathic arthropathy (12 radiographic, seven CT, and six MR studies) and 19 with disk space infection (13 radiographic, nine CT, and 12 MR studies). Potential imaging discriminators, including endplate sclerosis or erosions, osteophytes, spondylolisthesis, facet involvement (narrowing or erosions), vacuum disk, paraspinal soft-tissue mass, joint disorganization, and osseous joint debris, were recorded, as were MR imaging signal intensity and gadolinium-enhancement characteristics. RESULTS: The most helpful findings for diagnosis of spinal neuropathic arthropathy were vacuum disk on radiographs and CT images, debris on radiographs and CT and MR images, disorganization on radiographs and CT and MR images, facet involvement on radiographs and CT and MR images, spondylolisthesis on CT and MR images, diffuse signal intensity patterns in vertebral bodies on MR images, and rim enhancement of disks on gadolinium-enhanced MR images. Findings that were not helpful included endplate sclerosis and erosions, osteophytes, paraspinal soft-tissue mass, and decreased disk height. CONCLUSION: Vacuum disk, facet involvement, vertebral body spondylolisthesis, joint disorganization and debris, and gadolinium-enhancement patterns of vertebral bodies and disks may help differentiate spinal neuropathic arthropathy from infection. PMID- 10715034 TI - Normal maturation of the distal femoral epiphyseal cartilage: age-related changes at MR imaging. AB - PURPOSE: To determine how signal intensity in the cartilaginous distal part of the femoral epiphysis varies with (a) age, (b) sex, and (c) distribution to the medial or lateral condyle on magnetic resonance (MR) images. MATERIALS AND METHODS: Sixty-six sagittal T2-weighted or inversion-recovery MR images of the distal femoral epiphysis in children aged 2 months to 5 years 5 months were evaluated. Epiphyses were categorized into five types on the basis of progressive signal intensity changes within the epiphyseal cartilage along the weight-bearing region and posterior condyles. Epiphyseal type was compared with age, sex, and distribution of signal intensity changes within the condyle. RESULTS: In early infancy, epiphyseal cartilage was homogeneous. During the 2nd year, signal intensity along the weight-bearing region decreased. With further advancing age, signal intensity in the posterior femoral condyles increased and became progressively more focal. The increase in epiphyseal grade correlated with age for both the medial and the lateral femoral condyles (r = 0.71 and r = 0.77, respectively; P < .001). There was no significant difference in epiphyseal changes between boys and girls or between medial and lateral condyles. CONCLUSION: There is normal age-related variation in MR imaging signal intensity within the cartilaginous epiphysis of the distal femur. This may be related to weight bearing and epiphyseal maturation and should not be confused with disease. PMID- 10715035 TI - Hypertensive encephalopathy: complication in children treated for myeloproliferative disorders--report of three cases. AB - We routinely perform echo-planar diffusion-weighted sequences in all brain magnetic resonance (MR) imaging studies. When three children undergoing chemotherapy for acute leukemia presented with seizures, conventional MR images demonstrated what appeared to be acute, posterior, parasagittal infarcts. However, diffusion-weighted images were normal. These MR imaging findings were consistent with those of hypertensive encephalopathy. Early recognition and treatment of minimal hypertension in these patients allows reversal of encephalopathy. PMID- 10715036 TI - Fetal skeletal dysplasia: three-dimensional US--initial experience. AB - PURPOSE: To compare the prenatal ultrasonographic (US) features of skeletal dysplasia by using two-dimensional (2D) and three-dimensional (3D) US to determine whether 3D US can reveal additional diagnostic information. MATERIALS AND METHODS: Seven pregnant women suspected of having skeletal dysplasia were examined by using 2D US and 3D US. Data regarding the thorax, spine, face, limbs, hands, and feet were compared. Multiplanar and volume-rendered US images were evaluated. RESULTS: The skeletal dysplasias studied included camptomelic dysplasia (n = 2), thanatophoric dysplasia (n = 1), osteogenesis imperfecta (n = 1), arthrogryposis (n = 2), and short-limbed dysplasia (n = 1). Three-dimensional US, by allowing review in a standard anatomic orientation, was better than 2D US in depicting abnormal spatial relationships such as short ribs, splayed digits, and absent bones. Three-dimensional US enabled the acquisition of additional information in two fetuses with facial abnormalities and in two fetuses with scapular aplasia or hypoplasia (one fetus had both facial and scapular anomalies); it enabled a specific diagnosis in one fetus. The archiving capabilities of 3D US allow the review and manipulation of data after the patient has left the clinic. CONCLUSION: In three of seven patients, 3D US provided additional information in the evaluation of skeletal dysplasias, as compared with 2D US. PMID- 10715037 TI - Twin fetuses: intravascular microbubble US contrast agent administration--early experience. AB - PURPOSE: To explore the feasibility of administering SH U 508A by using a single needle procedure at ultrasonography (US) in twin pregnancies to confirm interfetal transfusion in monochorionic twins and delineate placental angioarchitecture in pregnancies with twin-twin transfusion syndrome. MATERIALS AND METHODS: Fourteen twin pregnancies were studied over 12 months: seven with monochorionic twins, including six with twin-twin transfusion syndrome; two of unknown chorionicity; and five with known dichorionic twins discordant for fetal karyotype or anomaly and undergoing selective feticide in the third trimester. Bolus injection of 100 microL/kg of estimated fetoplacental weight of 400 mg/mL of SH U 508A was performed in the intrahepatic vein of one twin, and evidence of interfetal transfusion was sought by means of digital analysis of power Doppler signals in the contralateral twin. RESULTS: Contralateral twin echo enhancement was seen in four of the nine ultimately histopathologically proved monochorionic twins. As expected, no evidence of echo enhancement in the contralateral twin was seen in any of the five dichorionic twin pregnancies. There was no evidence of fetal compromise associated with the procedure. CONCLUSION: These pilot results suggest that microbubbles can be used to demonstrate interfetal transfusion but not to delineate placental vascular anatomy. PMID- 10715038 TI - Uterine fibroleiomyoma: MR imaging appearances before and after embolization of uterine arteries. AB - PURPOSE: To evaluate the magnetic resonance (MR) imaging appearances of uterine fibroleiomyoma before and after embolization and to determine whether there are preembolization MR imaging characteristics that are predictive of a successful outcome. MATERIALS AND METHODS: MR imaging was performed in 18 patients (32 fibroleiomyomas) before and at 2 and 6 months after embolization of the uterine arteries. On each occasion, fibroleiomyoma signal intensity and gadolinium enhancement characteristics were assessed in comparison with those of myometrium on T1-weighted and gadolinium-enhanced images or with those of skeletal muscle on T2-weighted images. Fibroleiomyoma volume was measured by using the ellipsoid formula. RESULTS: The mean fibroleiomyoma volume before embolization was 340 cm3 (range, 15-1,383 cm3). The mean reduction in fibroleiomyoma volume was 43% at 2 months and 59% at 6 months. Before embolization, high signal intensity on T1 weighted images was predictive of a poor response (P = .008), and high signal intensity on T2-weighted images was predictive of a good response (P = .007). The degree of gadolinium enhancement was not correlated with fibroleiomyoma volume reduction (P = .46). CONCLUSION: MR imaging was useful for evaluation of changes in fibroleiomyoma volume after uterine arterial embolization. MR imaging characteristics of fibroleiomyomas before embolization can help predict subsequent response to treatment. PMID- 10715039 TI - Fetus in fetu: CT appearance--report of two cases. AB - Fetus in fetu is a rare abnormality secondary to the abnormal embryogenesis in a diamniotic, monochorionic pregnancy. It is an unusual condition in which a vertebrate fetus is enclosed within the abdomen of a normally developing fetus. Presented are the computed tomographic findings in two cases in which imaging findings were diagnostic of this entity. PMID- 10715040 TI - Renal arteries in patients at risk of renal arterial stenosis: multicenter evaluation of the echo-enhancer SH U 508A at color and spectral Doppler US. Levovist Renal Artery Stenosis Study Group. AB - PURPOSE: To assess SH U 508A in the diagnosis of suspected renal arterial stenosis by means of ultrasonography (US) and to confirm the safety of SH U 508A in a clinical setting. MATERIALS AND METHODS: A randomized crossover study was performed in 198 patients from 14 European centers who were referred for renal arterial angiography because they were suspected of having renal arterial stenosis. All patients underwent nonenhanced and SH U 508A-enhanced Doppler US of the renal arteries. Doppler criteria included measurement of renal arterial peak systolic velocity (threshold, 1.4-2.0 m/sec) in all centers and renoaortic ratio (threshold, 3.0-3.5) in nine. RESULTS: The number of examinations with successful results increased following enhanced Doppler US examination--160 (83.8%) compared with 122 (63.9%) with nonenhanced Doppler US (P = .001), including patients with obesity or renal dysfunction. Renal arterial stenosis (> or =50%) was detected at angiography in 72 patients. Results at enhanced Doppler US were in agreement with results at angiography more often than with results at nonenhanced Doppler US in the diagnosis or exclusion of renal arterial stenosis (P = .001). For patients examined with nonenhanced and enhanced Doppler US, sensitivity (80.0% and 83.7%, respectively) and specificity (80.8% and 83.6%, respectively) remained unchanged. There were no clinically important adverse events following use of SH U 508A. CONCLUSION: In patients suspected of having renal arterial stenosis, SH U 508A increased the number of diagnostic renal arterial Doppler studies. PMID- 10715041 TI - Soft-tissue vascular anomalies: utility of US for diagnosis. AB - PURPOSE: To determine the ultrasonographic (US) features that distinguish soft tissue hemangioma from vascular malformation and one type of malformation from another. MATERIALS AND METHODS: Eighty-seven vascular anomalies were evaluated by means of US. Lesions were assessed for the presence of solid tissue and abnormal arteries, veins, or cysts. Vessel density, peak flow velocities, and resistive indexes were compared. RESULTS: There were 49 hemangiomas and 38 vascular malformations. A significantly greater proportion of hemangiomas (48 of 49) compared with vascular malformations (zero of 38) consisted of a solid-tissue mass (P < .001). Vessel density was comparable for hemangioma and arteriovenous malformation (AVM) but significantly greater compared with the other vascular malformations (P < .001 in each case). No differences in mean arterial peak velocity were detected between hemangiomas and malformations. Mean venous peak velocity was significantly higher for AVM than for other vascular malformations and hemangioma. Mean resistive index was greater for lymphatic malformation than for hemangioma or AVM. Abnormal veins, arteries and veins, or cysts were univariate predictors for distinguishing between venous, arteriovenous, and lymphatic malformations (P < .001 in all cases). Solid-tissue mass was the only multivariate predictor for differentiating hemangioma from vascular malformation (likelihood ratio test = 109.8, P < .001). CONCLUSION: US can be used to distinguish hemangioma from vascular malformation and detect arterial flow. These distinctions are critical for subsequent management and assessing prognosis. PMID- 10715042 TI - Arteriographic detection of renovascular disease in potential renal donors: incidence and effect on donor surgery. AB - PURPOSE: To determine the arteriographic incidence and severity of renal arterial disease in potential renal donors and to evaluate the effect of identifying vascular abnormalities on subsequent donor surgery. MATERIALS AND METHODS: The records of 716 potential living renal donors who underwent conventional arteriography were reviewed. Abnormal arteriograms were reexamined to characterize vascular disease, and the effect of identifying renovascular disease on subsequent donor surgery was ascertained with chart review. RESULTS: Renovascular abnormalities were noted in the dictated reports in 78 patients (10.9%). The most common causes were fibromuscular dysplasia and atherosclerosis. The arteriograms of 64 patients were available for retrospective review. Abnormalities were characterized as minimal stenosis (<30% narrowing) in 42 patients and mild stenosis (30%-50% narrowing) in 19 of 61 patients with arteriographic abnormalities at retrospective review. In three patients, no significant abnormality was seen at retrospective review. The effect of detecting renovascular disease on donor selection was determined in 74 of the 78 patients. In 73 of these 74 patients (99%), detection of an abnormality directly affected donor surgery. CONCLUSION: In this population of potential renal donors, the arteriographic incidence of renovascular disease (10.9%) was higher than previously reported. Although renovascular abnormalities were mild, their detection influenced the plan for donor surgery in almost all patients. PMID- 10715043 TI - Hepatocellular carcinoma: radio-frequency ablation of medium and large lesions. AB - PURPOSE: To study local therapeutic efficacy, side effects, and complications of radio-frequency (RF) ablation in the treatment of medium and large hepatocellular carcinoma (HCC) lesions in patients with cirrhosis or chronic hepatitis. MATERIALS AND METHODS: One-hundred fourteen patients who were under conscious sedation or general anesthesia had 126 HCCs greater than 3.0 cm in diameter treated with RF by using an internally cooled electrode. Eighty tumors were medium (3.1-5.0 cm), and 46 were large (5.1-9.5 cm). The mean diameter for all tumors was 5.4 cm. At imaging, 75 tumors were considered noninfiltrating, and 51 were considered infiltrating. RESULTS: Complete necrosis was attained in 60 lesions (47.6%), nearly complete (90%-99%) necrosis in 40 lesions (31.7%), and partial (50%-89%) necrosis in the remaining 26 lesions (20.6%). Medium and/or noninfiltrating tumors were treated successfully significantly more often than large and/or infiltrating tumors. Two major complications (death, hemorrhage requiring laparotomy) and five minor complications (self-limited hemorrhage, persistent pain) were observed. The single death was due to a break in sterile technique rather than to the RF procedure itself. CONCLUSION: RF ablation appears to be an effective, safe, and relatively simple procedure for the treatment of medium and large HCCs. PMID- 10715044 TI - Small hepatocellular carcinoma: safety and efficacy of single high-dose percutaneous acetic acid injection for treatment. AB - PURPOSE: To evaluate the safety and efficacy of single high-dose percutaneous acetic acid injection (PAI) for treatment of small (<3-cm-diameter) hepatocellular carcinoma (HCC). MATERIALS AND METHODS: Eighteen patients with HCC (22 nodules; diameter range, 1.5-3.0 cm) underwent single PAI. With ultrasonographic or computed tomographic (CT) guidance, 4-11 mL of 50% acetic acid was slowly injected into the center of the nodule through a skinny multiple side-hole needle. Follow-up was performed with helical contrast material-enhanced CT. Complications of high-dose PAI were recorded. RESULTS: Seventeen nodules showed no local recurrence (follow-up, 6-29 months) after single PAI. At a mean follow-up of 15.6 months, mean tumor diameter was 2.1 cm and mean injected volume was 6.4 mL. Four nodules showed residual tumor (mean tumor diameter, 2.6 cm; mean injected volume, 5.8 mL). The mean ratio of injected to estimated volume of acetic acid was 1.21 in cases of successful single PAI and 0.72 in cases of local recurrence (P < .001). One patient with preexistent right portal venous thrombosis died of hepatic failure 37 days after PAI. Other complications included severe pain (11%), high fever (4%), and segmental wedge infarction (4%). CONCLUSION: Single high-dose PAI is safe and effective for treatment of small HCC. PMID- 10715045 TI - Hepatic arterial complications in liver transplant recipients treated with pretransplantation chemoembolization for hepatocellular carcinoma. AB - PURPOSE: To compare the prevalence of hepatic arterial complications in patients who underwent hepatic arterial chemoembolization for hepatocellular carcinomas before orthotopic liver transplantation with the prevalence of hepatic arterial complications in the total population of liver transplant recipients. MATERIALS AND METHODS: Forty-seven patients underwent selective hepatic arterial chemoinfusion with mitomycin C, doxorubicin hydrochloride, and cisplatin combined with embolization. The prevalence rates for hepatic arterial complications, including pseudoaneurysm, stenosis, anastomotic disruption, and thrombosis, were tabulated and compared with results in 1,154 patients who underwent orthotopic liver transplantation but not chemoembolization. RESULTS: Of the 47 patients who had undergone preoperative hepatic arterial chemotherapy, 13% developed hepatic arterial complications within a mean of 7 days after transplantation; an 8% prevalence of hepatic arterial thrombosis was observed. Of the 1,154 patients who underwent orthotopic liver transplantation but not chemotherapy, 6% developed hepatic arterial complications; a 5% prevalence of hepatic arterial thrombosis was observed. There was no statistically significant difference in the prevalence rates for thrombosis and complications between the patients who underwent chemoembolization before orthotopic liver transplantation and those who did not. The mean interval between chemotherapy and orthotopic liver transplantation was 111 days (range, 3-428 days). CONCLUSION: Patients who undergo hepatic arterial chemotherapy are not at an increased risk of developing hepatic arterial thrombosis or other hepatic arterial complications after orthotopic liver transplantation. PMID- 10715046 TI - Balloon catheter dilation in common canalicular obstruction of the lacrimal system: safety and long-term effectiveness. AB - PURPOSE: To evaluate the safety and long-term effectiveness of balloon catheter dilation in the treatment of common canalicular obstruction of the lacrimal system. MATERIALS AND METHODS: Fluoroscopically guided dilation with a 3-mm diameter balloon catheter was attempted in 195 eyes of 148 patients (26 men, 122 women; mean age, 57 years; age range, 33-78 years) with epiphora due to common canalicular obstruction. Eighty-four of 195 eyes had complete obstruction, and 111 had partial obstruction. RESULTS: Complications were self-limited nasal bleeding (n = 8), false passage (n = 7), and extravasation of contrast material (n = 6). Initial technical success was achieved in 76 (90%) of 84 eyes with complete obstruction and in 104 (94%) of 111 eyes with partial obstruction. In the 180 eyes with technical success, immediate clinical improvement was achieved in 76 eyes with complete obstruction and in 100 eyes with partial obstruction. The mean follow-up period was 36 weeks (range, 4-168 weeks). The cumulative patency rates were 51% at 6-month, 43% at 12-month, and 40% at 24-month follow up. CONCLUSION: Although the long-term recurrence rate is relatively high, balloon catheter dilation is a safe and effective therapeutic technique to be used initially in common canalicular obstruction. PMID- 10715047 TI - Comparison of two blood pool contrast agents for 0.5-T MR angiography: experimental study in rabbits. AB - PURPOSE: To evaluate two experimental blood pool agents for potential use in equilibrium phase abdominal magnetic resonance (MR) angiography. MATERIALS AND METHODS: MR imaging at 0.5 T was performed in 37 rabbits before and after intravenous injection of a gadolinium-based blood pool contrast agent (SH L 643 A), superparamagnetic iron oxide blood pool agent (SH U 555 C), or gadopentetate dimeglumine. T1-weighted fast spoiled gradient-echo images from the renal arteries to below the iliac bifurcation were obtained. The aorta-to-tissue signal difference-to-noise ratio (SDNR) was measured over time. RESULTS: Both blood pool agents yielded excellent demonstration of the rabbit abdominal aorta. At a dose of 0.1 mmol/kg, both provided a statistically significant increase in aorta-to tissue SDNR in comparison with that achieved with gadopentetate dimeglumine (200% increase for SH L 643 A, 95% increase for SH U 555 C; P < .05). A 0.1 mmol/kg dose of SH L 643 A provided a 24% increase in SDNR relative to the increase with a 0.37 mmol/kg dose of gadopentetate dimeglumine. Time-dependent enhancement properties of the blood pool agents differed due to differences in elimination method. CONCLUSION: Both blood pool agents were found to be promising contrast agents for 0.5-T MR angiography; however, their clinical applicability warrants further investigation. The gadolinium-based agent had several advantages over the iron oxide compound, including less T2* dephasing, lack of susceptibility artifacts, and fast renal elimination. PMID- 10715048 TI - Radiolabeled annexin V imaging: diagnosis of allograft rejection in an experimental rodent model of liver transplantation. AB - PURPOSE: To assess the value of imaging rejection-induced apoptosis with technetium 99m and annexin V, a human protein-based radiopharmaceutical used in the diagnosis of acute rejection of a liver transplant, in a well-characterized rodent model of orthotopic liver transplantation. MATERIALS AND METHODS: 99mTc radiolabeled annexin V was intravenously administered to six allografted (immunologically mismatched) and five isografted (immunologically matched) recipient rats on days 2, 4, and 7 after orthotopic liver transplantation. Animals were imaged 1 hour after injection of 0.2-2.0 mCi (8.0-74.0 MBq) of radiolabeled annexin V by use of clinical nuclear scintigraphic equipment. RESULTS: All animals in the allografted group demonstrated marked increases of 55% and 97% above the activity in the isografted group in hepatic uptake of annexin V on days 4 and 7, respectively. Severe acute rejection was histologically detected in all allografted livers on day 7. There was no histologic evidence of acute rejection in isografted animals. Dynamic hepatobiliary imaging with 99mTc and mebrofenin, an iminodiacetic acid derivative, demonstrated no correlation with the presence or absence of acute rejection or with annexin V uptake. CONCLUSION: Noninvasive imaging with radiolabeled annexin V is more sensitive and specific than imaging with 99mTc mebrofenin in the diagnosis of acute rejection of a liver transplant. PMID- 10715049 TI - Coronary angiogenesis: detection in vivo with MR imaging sensitive to collateral neocirculation--preliminary study in pigs. AB - PURPOSE: To assess the ability to track neovascularization over time with a magnetic resonance (MR) imaging technique sensitized to new intramyocardial collateral development as a means of evaluating therapeutic angiogenesis. MATERIALS AND METHODS: Magnetization preparation plus spatial frequency reordering was applied to distinguish new intramyocardial collateral vessels from normal circulation on the basis of geometric differences. A vascular occluder was inserted in 34 pigs, and they were assigned randomly to treatment groups with either placebo or angiogenic growth factor. Collateral extent determined with collateral-sensitive MR imaging was correlated with direct measurements by means of three-dimensional (3D) computed tomography (CT), coronary blood flow distribution determined with microspheres, and findings at histologic examination. Changes in the signal at collateral-sensitive MR imaging before and after treatment were assessed by two observers blinded to treatment. RESULTS: The collateral extent determined with collateral-sensitive MR imaging correlated well with findings at 3D CT (r = 0.95) and with microspheres (r = 0.86). Furthermore, the collateral extent determined with collateral-sensitive MR imaging increased significantly (P < .001) in response to the administration of an angiogenic growth factor but not to placebo. The correspondence of findings at collateral sensitive MR imaging to collateral neovascularization was confirmed at histologic examination. CONCLUSION: The presence of intramyocardial collateral microvessels was accurately determined with collateral-sensitive MR imaging. The technique may be useful in clinical studies of therapeutic angiogenesis. PMID- 10715050 TI - Three-dimensional, navigator-echo MR coronary angiography in detecting stenoses of the major epicardial vessels, with conventional coronary angiography as the standard of reference. AB - PURPOSE: To test three-dimensional (3D), navigator-echo magnetic resonance (MR) coronary angiography in detecting stenoses of the coronary arteries. MATERIALS AND METHODS: Forty-two patients (age range, 50-79 years) underwent MR coronary angiography (1.5 T). A navigator-echo sequence was used. Two or three 15% overlapped transverse slabs were acquired. Data were analyzed by readers blinded to conventional coronary angiographic results. On conventional coronary angiograms, coronary arterial stenoses of 50% or greater narrowing were considered significant. On MR coronary angiograms, the major coronary vessels were subdivided into proximal (within 5 cm) and distal (beyond 5 cm) segments, except for the left main vessel. Stenoses of 50% or greater were identified on reformatted multiplanar MR coronary angiograms. RESULTS: Three MR coronary angiographic examinations were aborted because of patient claustrophobia; 39 of 39 left main, 117 of 117 proximal, and 78 of 117 distal segments were visualized. MR coronary angiography showed a sensitivity of 82% (95% CI: 73%, 91%) and a specificity of 89% (95% CI: 85%, 94%) in overall stenoses identification, of 90% (95% CI: 81%, 99%) and 90% (95% CI: 83%, 96%) for proximal segments, and of 68% (95% CI: 50%, 86%) and 81% (95% CI: 71%, 92%) for distal segments, respectively. CONCLUSION: Navigator-echo, 3D MR coronary angiography is a promising sequence for assessing coronary arterial stenoses, but further improvements are required for distal segments. PMID- 10715051 TI - Radiology groups' workload in relative value units and factors affecting it. AB - PURPOSE: To measure diagnostic radiology groups' workload in physician work relative value units (RVUs) and identify factors affecting it. MATERIALS AND METHODS: In 1996 and 1997, the authors surveyed diagnostic radiology and radiation oncology groups regarding finances, workload, and basic characteristics. The study was based on approximately 100 diagnostic radiology groups. The authors analyzed the distribution of workload in different categories of groups, conducting multiple statistical analyses. RESULTS: The annual numbers of procedures were approximately 10%-15% lower than those in a comparison survey with a good response rate. The annual number of RVUs per full-time equivalent (FTE) diagnostic radiologist was highly variable in every group category, as was the number of RVUs per clinical work hour. Multivariate regression analysis indicated that variation in the annual number of hours worked did not explain variation in annual workload. RVUs per FTE radiologist were higher the greater the percentage of a group's workload from interventional, computed tomographic, and magnetic resonance imaging procedures. CONCLUSION: Given the likely response bias, the annual workload per FTE radiologist probably averaged approximately 4,000 RVUs in academic groups and approximately 6,000 in nonacademic groups, but the large, unexplained variance means the average values should not be taken as norms. PMID- 10715052 TI - Computerized analysis of the likelihood of malignancy in solitary pulmonary nodules with use of artificial neural networks. AB - PURPOSE: To develop a computer-aided diagnostic scheme by using an artificial neural network (ANN) to assist radiologists in the distinction of benign and malignant pulmonary nodules. MATERIALS AND METHODS: Fifty-six chest radiographs of 34 primary lung cancers and 22 benign nodules were digitized with a 0.175-mm pixel size and a 10-bit gray scale. Eight subjective image features were evaluated and recorded by radiologists in each case. A computerized method was developed to extract objective features that could be correlated with the subjective features. An ANN was used to distinguish benign from malignant nodules on the basis of subjective or objective features. The performance of the ANN was compared with that of the radiologists by means of receiver operating characteristic (ROC) analysis. RESULTS: Performance of the ANN was considerably greater with objective features (area under the ROC curve, Az = 0.854) than with subjective features (Az = 0.761). Performance of the ANN was also greater than that of the radiologists (Az = 0.752). CONCLUSION: The computerized scheme has the potential to improve the diagnostic accuracy of radiologists in the distinction of benign and malignant solitary pulmonary nodules. PMID- 10715053 TI - Correlation of aging and smoking with air trapping at thin-section CT of the lung in asymptomatic subjects. AB - PURPOSE: To assess the frequency and degree of air trapping at thin-section computed tomography (CT) of the lung in relation to age and smoking history in asymptomatic subjects. MATERIALS AND METHODS: Thin-section CT of the lung was performed prospectively at end inspiration and end expiration in 82 subjects (27 smokers, 55 nonsmokers) without any history of pulmonary diseases and without present pulmonary symptoms. The frequency and degree of air trapping were evaluated according to age and smoking status. RESULTS: The overall frequency of air trapping was 52% (43 of 82 subjects, kappa = 0.72). Air trapping was found in three of 13 (23%), seven of 17 (41%), nine of 18 (50%), 11 of 17 (65%), and 13 of 17 (76%) subjects aged 21-30, 31-40, 41-50, 51-60, and greater than or equal to 61 years, respectively. The frequency of air trapping increased with age (P < .05). The degree of air trapping had a significant correlation with age (r = 0.523, P < .001) and was higher in smokers with a smoking history of more than 10 pack-years (P < .05). CONCLUSION: Air trapping was found in approximately 50% of asymptomatic subjects. The frequency of air trapping increased with age, and its severity increased with age and smoking. PMID- 10715054 TI - Symptomatic brachial plexopathy following treatment for breast cancer: utility of MR imaging with surface-coil techniques. AB - PURPOSE: To investigate the clinical utility and diagnostic accuracy of magnetic resonance (MR) imaging in patients with symptomatic brachial plexopathy following treatment for breast cancer. MATERIALS AND METHODS: Fifty patients with symptoms of brachial plexopathy (principally pain, weakness, and paresthesia) who had received treatment for breast cancer, which included surgery, radiation therapy, and cytotoxic chemotherapy, underwent MR imaging at 1.5 T. MR imaging was performed by using a body coil, which was supplemented with surface-coil imaging of the cervical spine and shoulder-coil imaging of the brachial plexus. At review, two observers attempted to discriminate between tumor recurrence and nonmalignant causes of symptoms. The diagnosis was verified with histologic analysis or a follow-up of at least 12 months. RESULTS: Of 27 patients demonstrated to have tumor recurrence, 26 were correctly identified by using MR imaging; the recurrence was directly related to the brachial plexus in 17. During the follow-up, 21 patients remained free of recurrence, 20 of whom were determined to have a nonmalignant cause of symptoms. Two of the 50 patients were excluded from the analysis. The MR criteria used for detection of tumor yielded a sensitivity of 96%, specificity of 95%, positive predictive value of 96%, and negative predictive value of 95%. CONCLUSION: MR imaging is reliable and accurate in the diagnosis of symptomatic brachial plexopathy following breast cancer therapy. PMID- 10715055 TI - Case 24. PMID- 10715056 TI - Case 20: Biliary ascariasis. PMID- 10715057 TI - Severe chronic pancreatitis versus suspected pancreatic disease: dynamic MR cholangiopancreatography after secretin stimulation. AB - PURPOSE: To assess whether secretin stimulation improves visualization of the pancreatic ducts at magnetic resonance (MR) cholangiopancreatography (MRCP) in patients with severe chronic pancreatitis or suspected pancreatic disease. MATERIALS AND METHODS: Thirty-one patients (group 1) with chronic pancreatitis and 84 patients (group 2) with clinical and/or laboratory findings suggestive of pancreatic disease who did not have ductal alterations at ultrasonography (US) and/or computed tomography (CT) underwent MRCP before and up to 10 minutes after secretin stimulation. Size of the main pancreatic duct (head, body, tail) and duodenal filling before and after secretin stimulation were measured quantitatively. Image quality, number of main pancreatic ductal segments visualized, visualization of side branches, ductal narrowing, endoluminal filling defects, and presence of pancreas divisum were analyzed qualitatively. RESULTS: In both groups, the size of the main pancreatic duct increased significantly 3 minutes after secretin stimulation. Reduced duodenal filling was detected in patients with severe chronic pancreatitis (P < .001). The number of segments of the main pancreatic duct visualized improved from 85 (91%) to 93 (100%) of 93 in group 1 and from 164 (65%) to 245 (97%) of 252 (P < .001) in group 2. Visualization of side branches improved from 22 (71%) to 31 (100%) of 31 in group 1 and from three (4%) to 53 (63%) of 84 (P < .001) in group 2. Pancreas divisum was visualized in one additional patient in group 1 and in six additional patients in group 2. CONCLUSION: The administration of secretin improves visualization of the pancreatic ducts and helps in the evaluation of exocrine reserve. PMID- 10715058 TI - Pseudo-obstruction of the extrahepatic bile duct due to artifact from arterial pulsatile compression: a diagnostic pitfall of MR cholangiopancreatography. AB - PURPOSE: To evaluate the frequency of artifact from arterial pulsatile compression as the cause of pseudo-obstruction of the extrahepatic bile duct at magnetic resonance (MR) cholangiopancreatography (MRCP) and specify the causative vessels. MATERIALS AND METHODS: In 234 patients (102 men, 132 women; age range, 25-80 years), MRCP images obtained by using a single-shot turbo spin-echo sequence were reviewed to assess pseudo-obstruction of the extrahepatic bile duct caused by vascular compression. Dual-phase spiral computed tomography, contrast material-enhanced three-dimensional MR angiography, and/or digital subtraction angiography also were performed to determine the vessel that caused the pseudo obstruction. RESULTS: Thirty-six pseudo-obstructions due to vascular compression were found in 33 (14%) patients. The common hepatic duct (27 [75%] sites) was the most common pseudo-obstruction site, followed by the left hepatic duct (four [11%] sites), proximal common bile duct (three [8%] sites), and right hepatic duct (two [6%] sites). The causative vessels were identified as the right hepatic artery at 24 (67%) sites; gastroduodenal artery, two (6%) sites; cystic artery, two (6%) sites; proper hepatic artery, one (3%) site; and an unspecified branch of the common hepatic artery, seven (19%) sites. CONCLUSION: At MRCP, pseudo obstruction of the extrahepatic bile duct can be caused by pulsatile vascular compression of the hepatic and gastroduodenal arteries, and it should not be misdiagnosed as a bile duct tumor or biliary stone. PMID- 10715059 TI - Hepatocellular adenoma: multiphasic CT and histopathologic findings in 25 patients. AB - PURPOSE: To evaluate multiphasic computed tomographic (CT) findings of hepatic adenomas and to correlate these findings with those of histopathologic analysis. MATERIALS AND METHODS: Multiphasic helical CT was performed in 25 patients with 44 hepatic adenomas. Nonenhanced scans were obtained in all cases, along with hepatic arterial-dominant phase (HAP) and portal venous-dominant phase (PVP) images at 25-28 and 60-70 seconds after intravenous contrast material injection at 3-5 mL/sec. Twelve patients with 24 adenomas also underwent delayed-phase (5 10-minute) CT. Two independent readers retrospectively reviewed each case for the number of detectable lesions in each CT phase, morphologic features of tumors, and degrees of enhancement. RESULTS: Thirteen patients had solitary adenomas; 12 patients had two or three adenomas. Both observers agreed on the numbers of lesions detected in all cases and in all phases of enhancement. The detection rate for all 44 adenomas per type of examination was as follows: nonenhanced, 86% (38 of 44); HAP, 100% (44 of 44); PVP, 82% (36 of 44), and delayed, 88% (21 of 24). Tumor margins were well defined in 38 adenomas (86%), and the surface was smooth in 42 adenomas (95%). The right hepatic lobe was the only site of adenoma or was a site along with the left lobe in 29 cases (66%). Tumor fat and calcifications were uncommon (three cases [7%] and two cases [5%], respectively). Other than areas of fat, hemorrhage, or necrosis, the adenomas enhanced nearly homogeneously, especially on PVP and delayed-phase scans. Five patients had coexistent hepatic masses, which were focal nodular hyperplasia (n = 3) or hepatocellular carcinoma (n = 2). CONCLUSION: Hepatic adenomas often have characteristic features at multiphasic CT that may allow their distinction from other hepatic masses. PMID- 10715060 TI - Dysplastic nodules in liver cirrhosis: evaluation of hemodynamics with CT during arterial portography and CT hepatic arteriography. AB - PURPOSE: To evaluate the portal and arterial blood supplies to dysplastic nodules in the cirrhotic liver with computed tomography (CT) during arterial portography (CTAP) and CT hepatic arteriography (CTHA). MATERIALS AND METHODS: Nineteen histopathologically proved low-grade dysplastic nodules and 13 high-grade dysplastic nodules in 17 patients with liver cirrhosis were evaluated with CTAP and CTHA for the presence of portal and arterial blood supplies to the nodules. The nodules ranged from 0.4 to 4.5 cm in diameter (mean, 1.6 cm). RESULTS: The portal supply was present in 14 of the 19 (74%) low-grade dysplastic nodules and in seven of the 13 (54%) high-grade dysplastic nodules. The hepatic arterial supply was increased in four of the 19 (21%) low-grade dysplastic nodules, present in nine (47%), and absent in six (32%). The arterial supply was increased in four of the 13 (31%) high-grade dysplastic nodules, present in four (31%), and absent in five (38%). CONCLUSION: The portal and arterial supplies to the low- and high-grade dysplastic nodules were variable and inconsistent. Therefore, it is difficult to detect and characterize the dysplastic nodules on the radiologic images on the basis of the blood supply. PMID- 10715062 TI - The WES sign. PMID- 10715061 TI - Hepatic parenchymal enhancement during triple-phase helical CT: can it be used to predict which patients with breast cancer will develop hepatic metastases? AB - PURPOSE: To evaluate the efficacy of hepatic enhancement characteristics for identification of patients with breast cancer who are at risk for future hepatic metastases. MATERIALS AND METHODS: Triple-phase helical computed tomography (CT) was performed in 60 patients with known breast cancer without visible hepatic metastases. Peak hepatic attenuation and enhancement, and attenuation and enhancement at 25 and 30 seconds were obtained. Ratios of hepatic attenuation or enhancement at 25 and 30 seconds to peak hepatic attenuation or enhancement were calculated. A Wilcoxon rank sum test was used to compare patients with and those without subsequent hepatic metastases. RESULTS: During a mean 18-month follow-up, 18 patients (30%) developed hepatic metastases. Decreases in peak hepatic attenuation and enhancement and increases in hepatic attenuation and enhancement ratios at 25 and 30 seconds were seen in patients who developed metastases compared with those who did not (P < .05). When corrected for chemotherapy interval, these differences were not statistically significant. Using a threshold value of 0.40 or more for the enhancement ratio at 30 seconds resulted in sensitivity of 28%, specificity of 92%, and accuracy of 55%. CONCLUSION: Patients with breast cancer who develop subsequent hepatic metastases have higher relative hepatic arterial perfusion during triple-phase CT; however, after correction for chemotherapy interval, this difference was not statistically significant. Threshold values cannot be used reliably to identify patients who will develop metastases. PMID- 10715063 TI - Apparent ipsilateral decrease in breast size at mammography: a sign of infiltrating lobular carcinoma. AB - PURPOSE: To assess if infiltrating lobular carcinoma (ILC) is associated with an ipsilateral mammographic decrease in breast size. MATERIALS AND METHODS: Mammographic change in size was evaluated by measuring the distance from the nipple to the pectoralis major muscle on the mediolateral oblique view of the diagnostic mammogram and on a preceding mammogram in 30 patients with ILC. Clinical, mammographic, and histopathologic findings were retrospectively reviewed. RESULTS: Five patients (17%) had an ipsilateral decrease in mammographic size. No patients noticed a physical decrease in breast size. Patients with an ipsilateral decrease in mammographic size most commonly had breast thickening at examination (four of five patients [80%], P < .001) and either a focal asymmetry density (three of five patients [60%]) or architectural distortion (one of five patients [20%]) at mammography; those patients with no change in size most commonly had a palpable mass (six of 25 patients [24%]) or normal findings (19 of 25 patients [76%]) and a mass (13 of 25 patients [52%]) at mammography. The mean tumor size was 66 mm for those with an ipsilateral size decrease and 16 mm for those with no size decrease (P < .001). At histologic analysis, tumors associated with an ipsilateral decrease in mammographic size had more diffuse involvement of the breast, and discrete masses were not seen. CONCLUSION: An apparent decrease in mammographic size may help identify cases of ILC, especially when associated with thickening at clinical examination and focal asymmetric density at mammography. PMID- 10715064 TI - Metallic punctate densities in the breast after Chinese herbal treatment: mammographic findings. AB - PURPOSE: To describe the mammographic features of metallic punctate densities seen in women who were treated with the herb go-yak for breast abscess and to explain the cause of these findings. MATERIALS AND METHODS: Mammograms showing metallic punctate densities that appeared to be microcalcifications in 34 women were analyzed retrospectively with attention to the location, shape, distribution, and depth of the lesions. In all patients, go-yak was applied into the open wound after abscess drainage 6-42 years before mammography. In six patients, histopathologic specimens were obtained after needle localization. RESULTS: Metallic densities were in the subareolar or central breast in 24 (71%) of 34 patients. The shape was predominantly round or punctate in all patients, but rod-shaped or linear lesions were found in seven patients. The distribution and depth of lesions were variable, but they extended to the subcutaneous fat in 29 patients (85%). A high concentration of lead was found in the histopathologic specimens and herb samples. CONCLUSION: Lead deposits associated with go-yak treatment should be included in the differential diagnosis when the suspected microcalcifications are of unusually high density, are central in location, and extend into the subcutaneous fat in Asian women with a history of breast abscess. PMID- 10715066 TI - Ligating clips for three-dimensional MR angiography at 1.5 T: in vitro evaluation. AB - Artifact size on three-dimensional (3D) magnetic resonance (MR) angiograms and safety of various vascular clips (15 titanium and three absorbable polydioxanone clips) were assessed. All evaluated clips were completely safe. Biodegradable clips rendered no artifacts; titanium clips were associated with susceptibility effects. Artifact size was dependent on clip size, clip orientation, echo time, and degree of k-space coverage. In the presence of titanium vascular clips, fast 3D MR angiography should be performed with the shortest echo time and full k space coverage. PMID- 10715065 TI - Human breast cancer specimens: diffraction-enhanced imaging with histologic correlation--improved conspicuity of lesion detail compared with digital radiography. AB - Seven breast cancer specimens were examined with diffraction-enhanced imaging at 18 keV with a silicon crystal with use of the silicon 333 reflection in Bragg mode. Images were compared with digital radiographs of the specimen, and regions of increased detail were identified. Six of the seven cases (86%) showed enhanced visibility of surface spiculation that correlated with histopathologic information, including extension of tumor into surrounding tissue. PMID- 10715067 TI - Balloon-occluded endoscopic retrograde ileography. AB - For diagnostic ileography, the authors developed balloon-occluded endoscopic retrograde ileography and performed 77 studies in 36 consecutive patients with Crohn disease. Balloon-occluded endoscopic retrograde ileography proved to be useful in visualization of minute mucosal lesions such as aphthous ulcers and lymphoid hyperplasia in the distal ileum, and satisfactory ileographic images of Crohn disease were obtained in 54 (70%) studies. PMID- 10715069 TI - Carbon dioxide as a low-attenuation oral contrast agent. PMID- 10715068 TI - Vascular permeability: quantitative measurement with double-echo dynamic MR imaging--theory and clinical application. AB - Double-echo dynamic magnetic resonance (MR) imaging was used to evaluate both vascularity and permeability of tissues simultaneously. Vascularity was evaluated on the basis of the T2*-shortening effect due to the intravascular fraction of the contrast agent and permeability on the basis of the T1-shortening effect due to the extravascular fraction. Meningioma was characterized on the basis of higher vascularity and neurinoma on the basis of higher permeability. The proposed method enables better tissue characterization. PMID- 10715070 TI - In Memoriam. PMID- 10715071 TI - In Memoriam. PMID- 10715072 TI - Abstracts of Current Literature. PMID- 10715074 TI - What goes around comes around PMID- 10715073 TI - Tityustoxin effect on nerve compound action potentials requires extracellular sodium. AB - Previous studies have demonstrated that Li(+) ions can substitute for Na(+) in a variety of functional systems. Using the single sucrose-gap recording technique, we measured the nerve compound action potential to study the effects of tityustoxin (an alpha-scorpion toxin that selectively inhibits fast Na(+) channel inactivation) upon removal of extracellular Na(+). Our results suggest that tityustoxin requires the presence of extracellular Na(+) to produce its typical pharmacological effect on Na(+) channel inactivation kinetics, but not to bind to its site. PMID- 10715075 TI - A vision for nursing: the future revisited PMID- 10715076 TI - Future think PMID- 10715077 TI - Brandon/Hill selected list of print nursing books and journals. PMID- 10715078 TI - Preparing the nursing profession for participation in a genetic paradigm in health care. AB - It is critical that nurses be recognized for their ability to deliver genetic services in collaboration with medical geneticists, genetic counselors, physicians, and providers from other disciplines. The purpose of this special communication is to describe progress made by the International Society of Nurses in Genetics toward incorporating genetics into nursing education and practice. PMID- 10715079 TI - Prescription of physical activity by adult nurse practitioners: a national survey. PMID- 10715081 TI - President's message: stepping out: issuing position statements to inform health policy. PMID- 10715080 TI - Assessing the involvement of retired health professionals in meeting the needs of underserved populations. PMID- 10715082 TI - New role for AAN publication advisory committee. PMID- 10715083 TI - Position statement on adolescent health endorsed by American Academy of Nursing. PMID- 10715084 TI - Academy recognizes nursing legends. PMID- 10715085 TI - Building bridges over changing times. PMID- 10715086 TI - Improved clinical use of Twin-block and Herbst as a result of radiating viscoelastic tissue forces on the condyle and fossa in treatment and long-term retention: growth relativity. AB - Understanding mechanisms of action for orthopedic appliances is critical for orthodontists who hope to treat and retain the achieved corrections in patients with initial Class II mandibular retrognathism. That knowledge can help orthodontists produce clinically significant bone formation and avoid compression at the condyle-glenoid fossa region. It also assists us to understand the differences between short-term and long-term treatment results. It was previously thought that increased activity in the postural masticatory muscles was the key to promoting condyle-glenoid fossa growth. By analyzing results from several studies, we postulate that growth modification is associated with decreased activity, which leads to our nonmuscular hypothesis. This premise has its foundation on 3 key specific findings: significant glenoid fossa bone formation occurs during treatment that includes mandibular displacement; glenoid fossa modification is a result of the stretch forces of the retrodiskal tissues, capsule, and altered flow of viscous synovium; observations that glenoid fossa bone formation takes place a distance from the soft tissue attachment. The latter observation is explained by transduction or referral of forces. Evidence is presented, therefore, that the 3 trigger switches for glenoid fossa growth can similarly initiate short-term condylar growth modifications because the 2 structures are contiguous. These are displacement, several direct viscoelastic connections, and transduction of forces. Histologic evidence further shows that stretched retrodiskal tissues also insert directly into the condylar head's fibrocartilaginous layer. The impact of the viscoelastic tissues may be highly significant and should be considered along with the standard skeletal, dental, neuromuscular, and age factors that influence condyle-glenoid fossa growth with orthopedic advancement. These biodynamic factors are also capable of reversing effects of treatment on mandibular growth direction, size, and morphology. Relapse occurs as a result of release of the condyle and ensuing compression against the newly proliferated retrodiskal tissues together with the reactivation of muscle activity. To describe condyle-glenoid fossa growth modification, an analogy is made to a light bulb on a dimmer switch. The condyle illuminates in treatment, dims down in the retention period, to near base levels over the long term. PMID- 10715087 TI - Nonsurgical and nonextraction treatment of skeletal Class III open bite: its long term stability. AB - Two female patients, aged 14 years 5 months and 17 years 3 months with skeletal Class III open bite and temporomandibular dysfunction are presented. They had previously been classified as orthognathic surgical cases, involving first premolar removal. The primary treatment objective was to eliminate those skeletal and neuromuscular factors that were dominant in establishing their malocclusions. These included abnormal behavior of the tongue with short labial and lingual frenula, bilateral imbalance of chewing muscles, a partially blocked nasopharyngeal airway causing extrusion of the molars, with rotation of the mandible and narrowing of the maxillary arch. Resultant occlusal interference caused the mandible to shift to one side, which in turn produced the abnormal occlusal plane and curve of Spee. As a result, the form and function of the joints were adversely affected by the structural and functional asymmetry. These cases were treated by expanding the maxillary arch, which brought the maxilla downward and forward. The mandible moved downward and backward, with a slight increase in anterior facial height. Intruding and uprighting the posterior teeth, combined with a maxillary protraction, reconstructed the occlusal plane. A favorable perioral environment was created with widened tongue space in order to produce an adequate airway. Myofunctional therapy after lingual and labial frenectomy was assisted by vigorous gum chewing during and after treatment, together with a tooth positioner. Normal nasal breathing was achieved. PMID- 10715088 TI - Healing of autogenous intramembranous bone in the presence and absence of homologous demineralized intramembranous bone. AB - This study was designed to examine the osteogenecity of demineralized bone matrix (DBM) prepared from intramembranous (IM) bone and to quantitatively assess the amount of new bone formed by IM autogenous bone grafts with or without DBM(IM). Forty-two defects were created in 42 New Zealand White rabbits. Twenty-one defects were grafted with IM bone alone, and the other 21 defects were grafted with composite IM-DBM(IM). Eleven rabbits, 22 defects were used as controls, where 11 defects were left empty (passive control) and the other 11 defects were filled with rabbit skin collagen (active control). Tissues were retrieved on days 1, 2, 3, 4, 5, 6, 7, and 14 for qualitative and quantitative analysis. Cells involved in the healing of composite IM and IM-DBM(IM) bone grafts were identified. No cartilage cells were detected during the healing of either grafts. Appearance of small blood vessels into the newly formed matrix was seen on day 5 in IM bone grafts and on day 4 in composite IM-DBM(IM) bone graft. Quantitative analysis was performed by means of image analysis on 100 sections of tissues retrieved after 14 days. Approximately 204% more new bone was formed in defects grafted with composite IM-DBM(IM) than in those grafted with IM bone alone (P <.0001). No bone was formed across the defects in either active or passive controls. In conclusion, DBM(IM) significantly increases the osteogenicity of IM bone grafts. PMID- 10715089 TI - Otic axis locator: closing the accuracy gap in cephalometrics and cast mounting. AB - An overview is presented of a new technique to improve the accuracy and reproducibility of earpost-dependent orientation in procedures such as cephalometric radiography and cast mounting, using a custom-molded ear canal insert to provide a stable fitted socket for more exact earpost fit. This is especially valuable in follow-up treatment and serial studies because the insert can be filed for future use on the same patient to provide identical positioning in successive registrations. Cephalometric radiography is further enhanced by the addition of a radiopaque x-ray marker deep in the canal, otherwise inaccessible, close to the bony ear canal (anatomical porion) and the mandibular condyle. This marker is also visible in all 3 cephalometric views-lateral, frontal (posteroanterior) and coronal (submental-vertex)-providing an exact common horizontal axis (Otic axis) for a three-dimensional Cartesian coordinate system of measurement. PMID- 10715090 TI - An inference modeling of human visual judgment of sagittal jaw-base relationships based on cephalometry: part II. AB - In a clinical situation, visual judgment of sagittal jaw-base relationships is described and handled as fuzzy terms, such as "slight" skeletal 3 tendency or "relatively severe" skeletal 2. The purpose of this study was to develop an inference system in order to be able to describe the degree of certainty for sagittal skeletal discrepancy automatically and in a mathematical way. Orthodontic records of 137 adult female patients were judged by 3 orthodontists. Based on these data and the multiple regression model described in Part I of this study, we developed an inference system that consisted of 2 membership functions and 6 incorporating rules. The performance reliability of the system was tested for 175 female adult cases by 7 orthodontic experts. In 97% (170 of 175) of the cases, the orthodontic experts agreed with advice proposed by the inference system. We conclude that the current model is an effective means of deriving the opinion of an experienced clinician from a cephalometric tracing and provides an interesting insight into how orthodontists are influenced by the face when attempting to judge the sagittal relationship clinically. In addition, the current system would be an initial stage in the development of the decision making system for the orthodontic diagnosis and treatment planning. PMID- 10715091 TI - Extraction of maxillary first bicuspids and mandibular lateral incisors, combined with orthognathic surgery to correct a severe class II skeletal malocclusion. AB - This is a case report of a 21-year-old female with a Class II Division 1 malocclusion. The maxillary arch was constricted with an associated anterior open bite. The lower facial height was excessive, and the mandibular plane angle was high. The treatment options were limited due to a previously extracted mandibular right lateral incisor. The patient was successfully treated by a surgical rapid palatal expansion procedure, extraction of the mandibular left lateral incisor, extraction of the maxillary first premolars at the time of a 3-piece Lefort 1 maxillary osteotomy procedure, and a bilateral sagittal split osteotomy advancement procedure. PMID- 10715092 TI - Dental and facial skeletal characteristics and growth of males and females with class II, division 1 malocclusion between the ages of 10 and 14 (revisited)-part I: characteristics of size, form, and position. AB - The purpose of this study was to describe and analyze the craniofacial and dentofacial skeletal characteristics associated with Angle's Class II, Division 1 malocclusion. The material examined included 613 lateral head radiographs comprising 2 series: (1) 278 films of children with "normal" occlusion and (2) 335 films of children with Class II, Division 1 malocclusion. Each series was subdivided into 6 samples (3 female and 3 male; skeletal ages 10, 12, 14, [+/-6 months]), representing children with chronological ages ranging from 8.5 to 15.5 years. The radiographs were converted to computer-readable X and Y coordinate data and 52 linear, angular, and coordinate axis measurements were taken. Findings were visually verified by superimposing the computer-drawn composite plots of the Class II, Division 1 series over those of the normal series. In all 6 intergroup comparisons, it was found that: (1) the mandible and its dentition is similar to the controls in size, form, and position except for the position of the lower incisors in males; (2) the forehead (Gl), anteriorcranial base (Nas), maxilla (A) and dentition (molars and incisors) are protrusive (mesial positioned), with an increased frontal bone thickness at the level of the sinus, and a larger A-P maxilla, the palate of which is inclined superiorly at its anterior half; (3) no vertical dysplasia was evident; (4) the cranial base angle is larger, as are the anterior and posterior sections that compose it, but it is not related to mandibular position; (5) angular indexes of maxillary and mandibular position that included point Nasion are highly misleading indicators of maxillary and mandibular size and position. Visualized diagnosis via a composite norm based on age and sex might offer a more reliable alternative or supplement to the numeric reference standards now in use. Enlarged sinuses may contribute to the cause of Class II, Division 1 malocclusion. PMID- 10715093 TI - Dentoalveolar and skeletal changes associated with the pendulum appliance. AB - The purpose of the study was to examine the dentoalveolar and skeletal effects of the pendulum appliance in Class II patients at varying stages of dental development and with varying facial patterns (high, neutral, and low mandibular plane angles). Specifically, the amount and nature of the "distalization" of the maxillary first molars and the reciprocal effects on the anchoring maxillary first premolars and incisors were studied, as were skeletal changes in the sagittal and vertical dimensions of the face. Pretreatment and posttreatment cephalometric radiographs obtained from 13 practitioners were used to document the treatment of 101 patients (45 boys and 56 girls). The average maxillary first molar distalization was 5.7 mm, with a distal tipping of 10.6 degrees. The anchoring anterior teeth moved mesially, as indicated by the 1.8-mm anterior movement of the upper first premolars, with a mesial tipping of 1.5 degrees. The maxillary first molars intruded 0.7 mm, and the first premolars extruded 1.0 mm. Lower anterior facial height increased 2.2 mm; there was no significant difference in lower anterior facial height increase between patients of high, neutral, or low mandibular plane angles. In patients with erupted maxillary second molars, there was a slightly greater increase in lower anterior face height and in the mandibular plane angle and a slightly greater decrease in overbite in comparison to patients with unerupted second molars. Similar findings were observed in patients with second premolar anchorage versus those with second deciduous molar anchorage. The results of this study suggest that the pendulum appliance is effective in moving maxillary molars posteriorly during orthodontic treatment. For maximum maxillary first molar distalization with minimal increase in lower anterior facial height, this appliance is used most effectively in patients with deciduous maxillary second molars for anchorage and unerupted permanent maxillary second molars, although significant bite opening was not a concern in any patient in this study. PMID- 10715094 TI - A clinical evaluation of a locking orthodontic facebow. AB - Standard orthodontic facebows may accidentally detach from the appliance buccal tubes at night; this could reduce the effectiveness of extra oral traction and occasionally cause an injury. To try and prevent facebow detachment at night a facebow with a locking mechanism was introduced. This study assessed the ability of 706 consecutively treated patients to learn to wear and use this facebow. The facebows were fitted in 9 different practices supervised by 12 orthodontists. Data from the patients and orthodontists were collected over a 2-year period and covered approximately 166,550 nights. All the orthodontists were able to fit and adjust the facebow; a total of 697 patients successfully used the facebow. Accidental detachment of the facebow at night was reported to be less than 1%. This indicates a significant improvement in the safety of the facebow and should help to improve compliance by increasing the number of hours of wear achieved by the patients. PMID- 10715095 TI - An occlusal and cephalometric analysis of lower first and second premolar extraction effects. AB - This study was designed to examine lateral cephalometric and arch dimensional changes that occur in the mandibular arch during orthodontic treatment involving the extractions of various premolars. Pretreatment and posttreatment records of 73 patients were chosen at random from completed cases in the practice of one experienced orthodontist. Eighteen involved the extraction of lower first premolars, and 55 involved the extraction of lower second premolars. Of these 55, 29 involved the extraction of upper first premolars and 26 involved the extraction of upper second premolars. In the lower first premolar group, however, all 18 involved the extraction of upper first premolars. Males and females were evenly represented in the 3 subgroups. Pretreatment factors that suggested a basis for the extraction choice in this group of patients were found to include incisal overjet, molar relationship, and underlying vertical facial pattern. A wide variety of arch dimensional changes was found with different lower premolar extraction patterns. There was evidence, however, of more intermolar arch width reduction after the extraction of lower second premolars than lower first premolars. Orthodontic treatment with the extraction of premolars did not consistently cause a retrusive effect on the incisors. In fact, instances of proclination of the incisors occurred within all of the extraction groups. A large amount of individual variation in incisor and molar changes accompanied treatment involving all lower premolar extraction patterns. PMID- 10715096 TI - Brainerd F. Swain, 1911-1999. PMID- 10715097 TI - Rodrigues Ottolengui, MDS, DDS, LLD, FACD. PMID- 10715098 TI - Data loss prevention and useful utilities. PMID- 10715099 TI - Litigation, legislation, and ethics. Thwarting vicarious liability. PMID- 10715100 TI - HLA class II alleles associations of anticardiolipin and anti-beta2GPI antibodies in a large series of European patients with systemic lupus erythematosus. AB - The objective of this study was to determine the HLA class II associations of the anticardiolipin (aCL) and anti-beta2GPI (abeta2GPI) antibodies in a large series of European patients with systemic lupus erythematosus (SLE). A cohort of 577 European SLE patients was enrolled. aCL and abeta2GPI were measured by ELISA methods. Molecular typing of HLA-DRB1, DRB3, DRB4, DRB5, DQA1 and DQB1 loci was performed by the polymerase chain reaction-sequence specific oligonucleotide probes (PCR-SSOP) method. aCL of IgG, IgM and IgA isotypes were detected in 22.8%, 14% and 13.9% of patients, respectively. IgG and IgM abeta2GPI were detected in 20% of patients. aCL showed positive association with HLA DRB1*04, DRB1*0402, DRB1*0403, DRB1*07, DRB3*0301, DQA1*0201, DQA1*0301, DQB1*0302, and negative association with DQA1*0501, DRB3*0202. abeta2GPI showed positive association with DRB1*0402, DRB1*0403, DQB1*0302. DRB1*0402 carried the highest relative risk for the presence of both aCL (RR=8. 1) and abeta2GPI (RR=4.6). Our results confirm the already described associations of aCL with HLA DR4 and DR7, but also demonstrate that, among the alleles at the DRB1*04 locus, the *0402 was most represented both in aCL and in abeta2GPI positive patients. In addition, HLA class II associations of abeta2GPI are for the first time extensively examined in a large cohort of European SLE patients. PMID- 10715101 TI - FTIR analysis of the SII540 intermediate of sensory rhodopsin II: Asp73 is the Schiff base proton acceptor. AB - Sensory rhodopsin II (SRII), a repellent phototaxis receptor found in Halobacterium salinarum, has several homologous residues which have been found to be important for the proper functioning of bacteriorhodopsin (BR), a light-driven proton pump. These include Asp73, which in the case of bacteriorhodopsin (Asp85) functions as the Schiff base counterion and proton acceptor. We analyzed the photocycles of both wild-type SRII and the mutant D73E, both reconstituted in Halobacterium salinarum lipids, using FTIR difference spectroscopy under conditions that favor accumulation of the O-like, photocycle intermediate, SII540. At both room temperature and -20 degrees C, the difference spectrum of SRII is similar to the BR-->O640 difference spectrum of BR, especially in the configurationally sensitive retinal fingerprint region. This indicates that SII540 has an all-trans chromophore similar to the O640 intermediate in BR. A positive band at 1761 cm-1 downshifts 40 cm-1 in the mutant D73E, confirming that Asp73 undergoes a protonation reaction and functions in analogy to Asp85 in BR as a Schiff base proton acceptor. Several other bands in the C=O stretching regions are identified which reflect protonation or hydrogen bonding changes of additional Asp and/or Glu residues. Intense bands in the amide I region indicate that a protein conformational change occurs in the late SRII photocycle which may be similar to the conformational changes that occur in the late BR photocycle. However, unlike BR, this conformational change does not reverse during formation of the O-like intermediate, and the peptide groups giving rise to these bands are partially accessible for hydrogen/deuterium exchange. Implications of these findings for the mechanism of SRII signal transduction are discussed. PMID- 10715102 TI - Mechanism of nonphotochemical quenching in green plants: energies of the lowest excited singlet states of violaxanthin and zeaxanthin. AB - The xanthophyll cycle is an enzymatic, reversible process through which the carotenoids violaxanthin, antheraxanthin, and zeaxanthin are interconverted in response to the need to balance light absorption with the capacity to use the energy to drive the reactions of photosynthesis. The cycle is thought to be one of the main avenues for safely dissipating excitation energy absorbed by plants in excess of that needed for photosynthesis. One of the key factors needed to elucidate the molecular mechanism by which the potentially damaging excess energy is dissipated is the energy of the lowest excited singlet (S(1)) state of the xanthophyll pigments. Absorption from the ground state (S(0)) to S(1) is forbidden by symmetry, making a determination of the S(1) state energies of these molecules by absorption spectroscopy very difficult. Fluorescence spectroscopy is potentially the most direct method for obtaining the S(1) state energies. However, because of problems with sample purity, low emission quantum yields, and detection sensitivity, fluorescence spectra from these molecules, until now, have never been reported. In this work these technical obstacles have been overcome, and S(1) --> S(0) fluorescence spectra of violaxanthin and zeaxanthin are presented. The energies of the S(1) states deduced from the fluorescence spectra are 14 880 +/- 90 cm(-)(1) for violaxanthin and 14 550 +/- 90 cm(-)(1) for zeaxanthin. The results provide important insights into the mechanism of nonphotochemical dissipation of excess energy in plants. PMID- 10715103 TI - Aminoglycoside-arginine conjugates that bind TAR RNA: synthesis, characterization, and antiviral activity. AB - Regulation of HIV gene expression is crucially dependent on binding of the trans activator protein, Tat, to the trans-activation response RNA element, TAR, found at the 5' end of all HIV-1 transcripts. Tat-TAR interaction is mediated by a short arginine-rich domain of the protein. Disruption of this interaction could, in theory, create a state of complete viral latency. A new class of small molecule peptidomimetic TAR RNA binders, conjugates of aminoglycosides and arginine, was recently designed [Litovchick, A., Evdokimov, A. G., and Lapidot, A. (1999) FEBS Lett. 445, 73-79]. Two of these compounds, the tri-arginine derivative of gentamicin C (R3G) and the tetra-arginine derivative of kanamycin A (R4K), bind efficiently and specifically to TAR RNA. These compounds display negligible toxicity while being transported and accumulated in cell nuclei. Here we present a detailed synthesis and chemical characterization of the aminoglycoside-arginine conjugates R3G and R4K as well as GB4K, the tetra-gamma guanidinobutyric derivative of kanamycin A. Their binding sites on TAR RNA were assigned by RNase A, uranyl nitrate, and lead acetate footprinting. The conjugates interact with TAR RNA in the widened major groove, formed by the UCU bulge and the neighboring base pairs of the upper stem portion of TAR, the binding site of Tat protein, and Tat-derived peptides (e.g., R52). Our results suggest an additional binding site of R4K and R3G compounds, in the lower stem bulge region of TAR. The antiviral activity of the conjugates in cultured equine dermal fibroblasts infected with equine infectious anemia virus, used as a model system of HIV-infected cells, is also presented. PMID- 10715104 TI - Tracking structural features leading to resistance of activated protein C to alpha 1-antitrypsin. AB - Activated protein C (APC) is a multi-modular anticoagulant serine protease, which degrades factor V/Va and factor VIIIa. Human APC (hAPC) is inhibited by human alpha 1-antitrypsin (AAT), while the bovine enzyme (bAPC) is fully resistant to this serpin. Structural features in the catalytic domains between the two species cause this difference, but detailed knowledge about the causal molecular difference is missing. To gain insight into the APC-AAT interaction and to create a human protein C resistant to AAT inhibition, we have used molecular modeling and site-directed mutagenesis. First, a structural model for bAPC based on the Gla-domainless X-ray structure of hAPC was built. Screening the molecular surface of the human and bovine APC enzymes suggested that a hAPC molecule resistant to AAT inhibition could be constructed by substituting only a few amino acids. We thus produced recombinant hAPC molecules with a single mutation (S173E, the numbering follows the chymotrypsinogen nomenclature), two mutations (E60aS/S61R) or a combination of all these substitutions (E60aS/S61R/S173E). Amidolytic and anticoagulant activities of the three mutant APC molecules were similar to those of wild-type hAPC. Inhibition of wild-type hAPC by AAT was characterized by a second-order rate constant (k2) of 2.71 M-1 s-1. The amino acid substitution at position 173 (S173E mutant) led to partial resistance to AAT (k2 = 0.84 M-1 s-1). The E60aS/S61R mutant displayed mild resistance to AAT inhibition (k2 = 1.70 M-1 s-1), whereas the E60aS/S61R/S173E mutant was inefficiently inactivated by AAT (k2 = 0.40 M-1 s-1). Inhibition of recombinant APC molecules by the serpin protein C inhibitor (PCI) in the presence and absence of heparin was also investigated. PMID- 10715106 TI - Inhibitory activity and structural characterization of a C-terminal peptide fragment derived from the prosegment of the proprotein convertase PC7. AB - Mammalian proprotein convertases (PCs) belong to the family of recently discovered serine proteases responsible for the processing of a large number of precursor proteins into their active forms. The enzymatic activities of the convertases have been implicated in a variety of disease states, such as cancer and infectious and inflammatory diseases. Like many other proteases, PCs are also synthesized as inactive proenzymes with N-terminal extensions as their prosegments. Here, we present the inhibitory activities of a number of "putative" interfacial peptide fragments derived from the proregion of PC7. We found that a peptide fragment corresponding to the C-terminal region (residues 81p-104p, or C24: E(1)-A-V-L-A-K-H-E-A-V-R-W-H-S-E-Q-R-L-L-K-R-A-K-R(24)) of the PC7 prosegment displays a strong inhibition (K(i) = 7 nM) of the PC7 enzyme comparable to that of the full-length (104 residue) prosegment. The same 24 residue peptide shows significantly populated helical conformations in an aqueous solution close to the physiological condition. Structure calculations driven by NOE distance restraints revealed a slightly kinked helical conformation for the entire peptide, characterized by many side-chain/side-chain interactions including those involving charged residues E8-R11-E15 and hydrophobic residues W12 and L19. These results suggest that the C-terminal region of the prosegment of PC7 may play a dominant role in conferring the inhibitory potency to the cognate enzyme and this strong inhibitory activity may be a direct consequence of the folded conformation of the peptide fragment in solution. We surmise that such a structure-function correlation for an inhibitory peptide could lead to the design and discovery of molecules mimicking the specific interactions of the PC prosegments for their cognate proteases. PMID- 10715105 TI - Substitution of asparagine for arginine 347 of recombinant factor Xa markedly reduces factor Va binding. AB - Herein we describe a recombinant factor X (fX) with a single substitution at position 347 (fXR347N). Activated fXR347N had a reduced affinity for factor Va (fVa), although the catalytic impact of fVa binding remained intact. The mutation was selective as demonstrated by normal activation and inhibition, except in the presence of subsaturating heparin where the rate of inhibition by antithrombin III (ATIII) was 15% of normal. The reactivity of fXaR347N toward prothrombin was equivalent to wild-type fXa (fXaWT) in the absence of fVa and phospholipid. Addition (without phospholipid) of fVa dramatically increased the catalytic efficiency of fXaWT toward prothrombin but had a negligible effect on fXaR347N. On addition of phosphatidylcholine:phosphatidylserine (PC:PS, 3:1) vesicles, fXaR347Ndisplayed an increased catalytic activity in response to fVa, but the apparent affinity for fVa on the phospholipid surface was 5-20-fold lower than that of fXaWT. On an activated platelet surface, however, fXaWT and fXaR347N activated prothrombin similarly. In a competitive binding assay that measures the displacement of radiolabeled fXa from fVa on a phospholipid surface, fXaR347N was approximately 10-fold less effective than fXaWT. Substitution of fXa at position 347 selectively attenuates the interaction between fXa and fVa without affecting its catalytic activity. PMID- 10715107 TI - Design and solution structure of a well-folded stack of two beta-hairpins based on the amino-terminal fragment of human granulin A. AB - Four amino acid substitutions were introduced into a peptide corresponding to the amino-terminal subdomain (30-31 residues) of human granulin A (HGA) in order to assess the contributions of a hydrophobic framework and other interactions to structure stabilization of the stack of two beta-hairpins. The resulting hybrid peptide, HGA 1-31 (D1V, K3H, S9I, Q20P) with four free cysteines, spontaneously formed a uniquely disulfide-bonded isomer with an expected [1-3, 2-4] disulfide pairing pattern. This peptide was characterized in detail by use of NMR and shown to assume a highly stable structure in solution, in contrast to the amino terminal 1-30 fragment of human granulin A. The prototype peptide, or HGA 1-30 (C17S, C27S), had lower resistance to chemical reduction and proteolysis, broad NH and H(alpha) proton resonances, lower proton resonance dispersion, and no slowly exchanging amide protons. Two other peptides, HGA 1-30 (C17S, Q20P, C27S) and HGA 1-31 (D1V, K3H, S9I, C17S, C27S), with either Pro20 stabilizing a potential reverse turn or with a hydrophobic cluster consisting of Val1, His3, and Ile9, had sharper and slightly better dispersed NH and H(alpha) proton resonances, but still no slowly exchanging amide protons. The solution structure of HGA 1-31 (D1V, K3H, S9I, Q20P) indicates that it adopts a well-folded conformation of a stack of two beta-hairpins, as found for the amino-terminal subdomain of the prototypic carp granulin-1 with representative beta-hairpin stacks. These results highlight the importance of both hydrophobic and turn stabilizing interactions for the structural integrity of the hairpin stack scaffold. The conformational stability appears to be maintained by a combination of the well-formed second beta-hairpin and two hydrophobic clusters, one located at the interface between the two beta-hairpins and the other on "top" of the first beta-hairpin. The implications of these findings for the design of conformationally stable hairpin stacks are discussed. PMID- 10715108 TI - Identification of residues involved in the interaction of Staphylococcus aureus fibronectin-binding protein with the (4)F1(5)F1 module pair of human fibronectin using heteronuclear NMR spectroscopy. AB - Many pathogenic Gram-positive bacteria express cell surface proteins that bind to components of the extracellular matrix. This paper describes studies of the interaction between ligand binding repeats (D3 and D1-D4) of a fibronectin binding protein from Staphylococcus aureus with a module pair ((4)F1(5)F1) from the N-terminal region of fibronectin. When D3 was added to isotope-labeled (4)F1(5)F1, (1)H, (15)N, and (13)C NMR chemical shift changes indicate that binding is primarily via residues in (4)F1, although a few residues in (5)F1 are also affected. Both hydrophobic and electrostatic interactions appear to be involved. The NMR data indicate that part of the D3 repeat converts from a disordered to a more ordered, extended conformation on binding to (4)F1(5)F1. In further NMR experiments, selective reduction of the intensity of D1-D4 resonances was observed on binding to (4)F1(5)F1, consistent with previous suggestions that in each of D1, D2, and D3 repeats, the main fibronectin binding site is in the C terminal region of the repeat. In D1-D4, these regions also appear to go from a disordered to a more ordered conformation of fibronectin binding. Although the regions of the two proteins which interact had been previously identified, the findings presented here identify, for the first time, the specific residues in both proteins that are likely to be involved in the interaction. PMID- 10715109 TI - Native and non-native secondary structure and dynamics in the pH 4 intermediate of apomyoglobin. AB - The partly folded state of apomyoglobin at pH 4 represents an excellent model for an obligatory kinetic folding intermediate. The structure and dynamics of this intermediate state have been extensively examined using NMR spectroscopy. Secondary chemical shifts, (1)H-(1)H NOEs, and amide proton temperature coefficients have been used to probe residual structure in the intermediate state, and NMR relaxation parameters T(1) and T(2) and ?(1)H?-(15)N NOE have been analyzed using spectral densities to correlate motion of the polypeptide chain with these structural observations. A significant amount of helical structure remains in the pH 4 state, indicated by the secondary chemical shifts of the (13)C(alpha), (13)CO, (1)H(alpha), and (13)C(beta) nuclei, and the boundaries of this helical structure are confirmed by the locations of (1)H-(1)H NOEs. Hydrogen bonding in the structured regions is predominantly native-like according to the amide proton chemical shifts and their temperature dependence. The locations of the A, G, and H helix segments and the C-terminal part of the B helix are similar to those in native apomyoglobin, consistent with the early, complete protection of the amides of residues in these helices in quench-flow experiments. These results confirm the similarity of the equilibrium form of apoMb at pH 4 and the kinetic intermediate observed at short times in the quench-flow experiment. Flexibility in this structured core is severely curtailed compared with the remainder of the protein, as indicated by the analysis of the NMR relaxation parameters. Regions with relatively high values of J(0) and low values of J(750) correspond well with the A, B, G, and H helices, an indication that nanosecond time scale backbone fluctuations in these regions of the sequence are restricted. Other parts of the protein show much greater flexibility and much reduced secondary chemical shifts. Nevertheless, several regions show evidence of the beginnings of helical structure, including stretches encompassing the C helix-CD loop, the boundary of the D and E helices, and the C-terminal half of the E helix. These regions are clearly not well-structured in the pH 4 state, unlike the A, B, G, and H helices, which form a native-like structured core. However, the proximity of this structured core most likely influences the region between the B and F helices, inducing at least transient helical structure. PMID- 10715110 TI - Role of the structural domain of troponin C in muscle regulation: NMR studies of Ca2+ binding and subsequent interactions with regions 1-40 and 96-115 of troponin I. AB - The interaction between the calcium binding and inhibitory components of troponin is central to the regulation of muscle contraction. In this work, two-dimensional heteronuclear single-quantum coherence nuclear magnetic resonance (2D-?1H,15N? HSQC NMR) spectroscopy was used to determine the stoichiometry, affinity, and mechanisms for binding of Ca2+ and two synthetic TnI peptides [TnI1-40 (or Rp40) and TnI96-115] to the isolated C-domain of skeletal troponin C (CTnC). The Ca2+ titration revealed that 2 equiv of Ca2+ binds to sites III and IV of CTnC with strong positive cooperativity and high affinity [dissociation constant (KD) Met; Q151M) of HIV-1 RT has been shown to confer resistance to the virus against dideoxy nucleoside analogues [Shirasaka, T., et al. (1995) Proc. Natl. Acad. Sci. U.S.A. 92, 2398], suggesting that Gln 151 may be involved in conferring sensitivity to nucleoside analogues. To understand its functional implication, we generated two mutant derivatives of this residue (Q151M and Q151N) and examined their sensitivities to ddNTPs and their ability to discriminate against rNTPs versus dNTP substrates on natural U5 PBS HIV-1 RNA template. We found that Q151M was highly discriminatory against all four ddNTPs but was able to incorporate rNTPs as efficiently as the wild type enzyme. In contrast, the Q151N mutant was only moderately resistant to ddNTPs but exhibited a higher level of discrimination against rNTPs. The fidelity of misinsertion was found to be highest for the Q151N mutant followed by Q151M and the wild type enzyme. These results point toward the importance of the amino acid side chain at position 151 in influencing the ability of the enzyme in recognition and discrimination against the sugar moieties of nucleotide substrates. PMID- 10715112 TI - Role of an electrostatic network of residues in the enzymatic action of the Rhizomucor miehei lipase family. AB - We have used continuum electrostatic methods to investigate the role of electrostatic interactions in the structure, function, and pH-dependent stability of the fungal Rhizomucor miehei lipase (RmL) family. We identify a functionally important electrostatic network which includes residues S144, D203, H257, Y260, H143, Y28, R80, and D91 (residue numbering is from RmL). This network consists of residues belonging to the catalytic triad (S144, D203, H257), residues located in proximity to the active site (Y260), residues stabilizing the geometry of the active site (Y28, H143), and residues located in the lid (D91) or close to the first hinge (R80). The lid and the first hinge are associated with the interfacial activation of lipases, where an alpha-helical lid opens up by rotating around two hinge regions. All network residues are well conserved in a set of 12 lipase homologues, and 6 of the network residues are located in sequence motifs. We observe that the effects of modeled mutations R86L, D91N, and H257F on the pH-dependent electrostatic free energies differ significantly in the closed and open conformations of RmL. Mutation R86L is especially interesting since it stabilizes the closed conformation but destabilizes the open one. Site site electrostatic interaction energies reveal that interactions between R86 and D61, D113, and E117 stabilize the open conformation. PMID- 10715113 TI - Topography of the surface of the Escherichia coli phosphotransferase system protein enzyme IIAglc that interacts with lactose permease. AB - The unphosphorylated form of enzyme IIAglc of the Escherichia coli phosphoenolpyruvate:sugar phosphotransferase system inhibits transport catalyzed by lactose permease. We (Seok et al. (1997) Proc. Natl. Acad. Sci. U.S.A. 94, 13515-13519) previously characterized the area on the cytoplasmic face of lactose permease that interacts with enzyme IIAglc, using radioactive enzyme IIAglc. Subsequent studies (Sondej et al. (1999) Proc. Natl. Acad. Sci. U.S.A. 96, 3525 3530) suggested consensus binding sequences on proteins that interact with enzyme IIAglc. The present study characterizes a region on the surface of enzyme IIAglc that interfaces with lactose permease. Acetylation of lysine residues by sulfosuccinimidyl acetate treatment of enzyme IIAglc, but not lactose permease, reduced the degree of interaction between the two proteins. To localize the lysine residue(s) on enzyme IIAglc that is(are) involved in the regulatory interaction, selected lysine residues were mutagenized. Conversion of nine separate lysines to glutamic acid resulted in proteins that were still capable of phosphoryl acceptance from HPr. Except for Lys69, all the modified proteins were as effective as the wild-type enzyme IIAglc in a test for binding to lactose permease. The Lys69 mutant was also defective in phosphoryl transfer to glucose permease. To derive further information concerning the contact surface, additional selected residues in the vicinity of Lys69 were mutagenized and tested for binding to lactose permease. On the basis of these studies, a model for the region of the surface of enzyme IIAglc that interacts with lactose permease is proposed. PMID- 10715114 TI - Specificity of Ca2+-dependent protein interactions mediated by the C2A domains of synaptotagmins. AB - Synaptotagmins represent a family of neuronal proteins thought to function in membrane traffic. The best characterized synaptotagmin, synaptotagmin I, is essential for fast Ca2+-dependent synaptic vesicle exocytosis, indicating a role in the Ca2+ triggering of membrane fusion. Synaptotagmins contain two C2 domains, the C2A and C2B domains, which bind Ca2+ and may mediate their functions by binding to specific targets. For synaptotagmin I, several putative targets have been identified, including the SNARE proteins syntaxin and SNAP-25. However, it is unclear which of the many binding proteins are physiologically relevant. Furthermore, more than 10 highly homologous synaptotagmins are expressed in brain, but it is unknown if they execute similar binding reactions. To address these questions, we have performed a systematic, unbiased study of proteins which bind to the C2A domains of synaptotagmins I-VII. Although the various C2A domains exhibit similar binding activities for phospholipids and syntaxin, we found that they differ greatly in their protein binding patterns. Surprisingly, none of the previously characterized binding proteins for synaptotagmin I are among the major interacting proteins identified. Instead, several proteins that were not known to interact with synaptotagmin I were bound tightly and stoichiometrically, most prominently the NSF homologue VCP, which is thought to be involved in membrane fusion, and an unknown protein of 40 kDa. Point mutations in the Ca2+ binding loops of the C2A domain revealed that the interactions of these proteins with synaptotagmin I were highly specific. Furthermore, a synaptotagmin I/VCP complex could be immunoprecipitated from brain homogenates in a Ca2+-dependent manner, and GST-VCP fusion proteins efficiently captured synaptotagmin I from brain. However, when we investigated the tissue distribution of VCP, we found that, different from synaptic proteins, VCP was not enriched in brain and exhibited no developmental increase paralleling synaptogenesis. Moreover, binding of VCP, which is an ATPase, to synaptotagmin I was inhibited by both ATP and ADP, indicating that the native, nucleotide-occupied state of VCP does not bind to synaptotagmin. Together our findings suggest that the C2A-domains of different synaptotagmins, despite their homology, exhibit a high degree of specificity in their protein interactions. This is direct evidence for diverse roles of the various synaptotagmins in brain, consistent with their differential subcellular localizations. Furthermore, our results indicate that traditional approaches, such as affinity chromatography and immunoprecipitations, are useful tools to evaluate the overall spectrum of binding activity for a protein but are not sufficient to estimate physiological relevance. PMID- 10715115 TI - Mirroring perfection: the structure of methylglyoxal synthase complexed with the competitive inhibitor 2-phosphoglycolate. AB - The crystal structure of the transition-state analogue 2-phosphoglycolate (2PG) bound to methylglyoxal synthase (MGS) is presented at a resolution of 2.0 A. This structure is very similar to the previously determined structure of MGS complexed to formate and phosphate. Since 2PG is a competitive inhibitor of both MGS and triosephosphate isomerase (TIM), the carboxylate groups of each bound 2PG from this structure and the structure of 2PG bound to TIM were used to align and compare the active sites despite differences in their protein folds. The distances between the functional groups of Asp 71, His 98, His 19, and the carboxylate oxygens of the 2PG molecule in MGS are similar to the corresponding distances between the functional groups of Glu 165, His 95, Lys 13, and the carboxylate oxygens of the 2PG molecule in TIM. However, these spatial relationships are enantiomorphic to each other. Consistent with the known stereochemical data, the catalytic base Asp 71 is positioned on the opposite face of the 2PG-carboxylate plane as Glu 165 of TIM. Both His 98 of MGS and His 95 of TIM are in the plane of the carboxylate of 2PG, suggesting that these two residues are homologous in function. While His 19 of MGS and Lys 13 of TIM appear on the opposite face of the 2PG carboxylate plane, their relative location to the 2PG molecule is quite different, suggesting that they probably have different functions. Most remarkably, unlike the coplanar structure found in the 2PG molecule bound to TIM, the torsion angle around the C1-C2 bond of 2PG bound to MGS brings the phosphoryl moiety out of the molecule's carboxylate plane, facilitating elimination. Further, the superimposition of this structure with the structure of MGS bound to formate and phosphate suggests a model for the enzyme bound to the first transition state. PMID- 10715116 TI - EPR investigation of Cu2+-substituted photosynthetic bacterial reaction centers: evidence for histidine ligation at the surface metal site. AB - The coordination environments of two distinct metal sites on the bacterial photosynthetic reaction center (RC) protein were probed with pulsed electron paramagnetic resonance (EPR) spectroscopy. For these studies, Cu2+ was bound specifically to a surface site on native Fe2+-containing RCs from Rhodobacter sphaeroides R-26 and to the native non-heme Fe site in biochemically Fe-removed RCs. The cw and pulsed EPR results clearly indicate two spectroscopically different Cu2+ environments. In the dark, the RCs with Cu2+ bound to the surface site exhibit an axially symmetric EPR spectrum with g(parallel) = 2.24, A(parallel) = 160 G, g(perpendicular) = 2.06, whereas the values g(parallel) = 2.31, A(parallel) = 143 G, and g(perpendicular) = 2.07 were observed when Cu(2+) was substituted in the Fe site. Examination of the light-induced spectral changes indicate that the surface Cu2+ is at least 23 A removed from the primary donor (P+) and reduced quinone acceptor (QA-). Electron spin-echo envelope modulation (ESEEM) spectra of these Cu-RC proteins have been obtained and provide the first direct solution structural information about the ligands in the surface metal site. From these pulsed EPR experiments, modulations were observed that are consistent with multiple weakly hyperfine coupled 14N nuclei in close proximity to Cu2+, indicating that two or more histidines ligate the Cu2+ at the surface site. Thus, metal and EPR analyses confirm that we have developed reliable methods for stoichiometrically and specifically binding Cu2+ to a surface site that is distinct from the well characterized Fe site and support the view that Cu2+ is bound at or near the Zn site that modulates electron transfer between the quinones QA and QB (QA-QB --> QAQB-) (Utschig, L. M., Ohigashi, Y., Thurnauer, M. C., and Tiede, D. M (1998) Biochemistry 37, 8278-8281) and proton uptake by QB- (Paddock, M. L., Graige, M. S., Feher, G., and Okamura, M. Y. (1999) Proc. Natl. Acad. Sci. U.S.A. 96, 6183-6188). Detailed EPR spectroscopic characterization of these Cu2+-RCs will provide a means to investigate the role of local protein environments in modulating electron and proton transfer. PMID- 10715117 TI - Azotobacter vinelandii nitrogenases containing altered MoFe proteins with substitutions in the FeMo-cofactor environment: effects on the catalyzed reduction of acetylene and ethylene. AB - Altered MoFe proteins of Azotobacter vinelandii Mo-nitrogenase, with amino acid substitutions in the FeMo-cofactor environment, were used to probe interactions among C(2)H(2), C(2)H(4), CO, and H(2). The altered MoFe proteins used were the alpha-195(Asn) or alpha-195(Gln) MoFe proteins, which have either asparagine or glutamine substituting for alpha-histidine-195, and the alpha-191(Lys) MoFe protein, which has lysine substituting for alpha-glutamine-191. On the basis of K(m) determinations, C(2)H(2) was a particularly poor substrate for the nitrogenase containing the alpha-191(Lys) MoFe protein. Using C(2)D(2), a correlation was shown between the stereospecificity of proton addition to give the products, cis- and trans-C(2)D(2)H(2), and the propensity of nitrogenase to produce ethane. The most extensive loss of stereospecificity occurred with nitrogenases containing either the alpha-195(Asn) or the alpha-191(Lys) MoFe proteins, which also exhibited the highest rate of ethane production from C(2)H(2). These data are consistent with the presence of a common ethylenic intermediate on the enzyme, which is responsible for both ethane production and loss of proton-addition stereochemistry. C(2)H(4) was not a substrate of the nitrogenase with the alpha-191(Lys) MoFe protein and was a poor substrate of the nitrogenases incorporating either the wild-type or the alpha-195(Gln) MoFe protein, both of which had a low V(max) and high K(m) (120 kPa). Ethylene was a somewhat better substrate for the nitrogenase with the alpha-195(Asn) MoFe protein, which exhibited a K(m) of 48 kPa and a specific activity for C(2)H(6) formation from C(2)H(4) 10-fold higher than the others. Neither the wild-type nitrogenase nor the nitrogenase containing the alpha-195(Asn) MoFe protein produced cis-C(2)D(2)H(2) when turned over under trans-C(2)D(2)H(2). These results suggest that the C(2)H(4)-reduction site is affected by substitution at residue alpha-195, although whether the effect is related to the substrate reduction site directly or is mediated through disturbance of the delivery of electrons/protons is unclear. Ethylene inhibited total electron flux, without uncoupling MgATP hydrolysis from electron transfer, to a similar extent for all four A. vinelandii nitrogenases. This observation indicates that this C(2)H(4) flux-inhibition site is remote from the C(2)H(4)-reduction site. Added CO eliminated C(2)H(4) reduction but did not fully relieve its electron-flux inhibition with all four A. vinelandii nitrogenases, supporting the suggestion that electron-flux inhibition by C(2)H(4) is not directly connected to C(2)H(4) reduction. Thus, C(2)H(4) has two binding sites, and the presence of CO affects only the site at which it binds as a substrate. When C(2)H(2) was added, it also eliminated C(2)H(6) production from C(2)H(4) and also did not relieve electron flux inhibition fully. Thus, C(2)H(2) and C(2)H(4) are likely reduced at the same site on the MoFe protein. Two schemes are presented to integrate the results of the interactions of C(2)H(2) and C(2)H(4) with the MoFe proteins. PMID- 10715118 TI - FTIR study of the monosialoganglioside GM1 in perdeuterated dimyristoylglycerophosphocholine (DMPCd54) multilamellar bilayers: spectroscopic evidence of a significant interaction between Ca2+ ions and the sialic acid moiety of GM1. AB - Fourier transform infrared (FTIR) spectroscopy was employed to study bovine brain GM1 and perdeuterated dimyristoylglycerophosphocholine (DMPCd54) multilamellar dispersions (mole fractions of GM1 in DMPCd54: 0.12, 0.15, 0.19, 0.26, 0.34, 0.41, and 0.58), in the absence and presence of 10 mM CaCl2. GM1 micelles did not display a thermal phase transition in the temperature range 5-60 degrees C. Moreover, the ceramide moiety of GM1 inserted into the hydrophobic core of DMPCd54 bilayers and was capable of undergoing a single, cooperative phase transition (Tm = 22-28 degrees C, depending on GM1 content) in a bilayer system. This suggested that the mixed bilayers consisted of a homogeneous mixture and that GM1 was uniformly dispersed in the bilayer plane rather than segregated into regions of relative enrichment. The coexistence of GM1 and DMPCd54 in a bilayer environment induced a rearrangement of the interfacial hydrogen bonding network of the amide I and ester C=O groups, relative to GM1 micelles and DMPCd54 bilayers, respectively. The modifications induced by GM1 might ultimately modulate surface events such as lipid-lipid and/or lipid-protein interactions. The spectroscopic results also suggested that the glycolipid's headgroup surface location and conformation in bilayers allow GM1 to act as a receptor for Ca2+ via its sialic acid moiety. PMID- 10715119 TI - Mutations in the putative H-channel in the cytochrome c oxidase from Rhodobacter sphaeroides show that this channel is not important for proton conduction but reveal modulation of the properties of heme a. AB - As the final electron acceptor in the respiratory chain of eukaryotic and many prokaryotic organisms, cytochrome c oxidase catalyzes the reduction of oxygen to water, concomitantly generating a proton gradient. X-ray structures of two cytochrome c oxidases have been reported, and in each structure three possible pathways for proton translocation are indicated: the D-, K-, and H-channels. The putative H-channel is most clearly delineated in the bovine heart oxidase and has been proposed to be functionally important for the translocation of pumped protons in the mammalian oxidase [Yoshikawa et al. (1998) Science 280, 1723 1729]. In the present work, the functional importance of residues lining the putative H-channel in the oxidase from Rhodobacter sphaeroides are examined by site-directed mutagenesis. Mutants were generated in eight different sites and the enzymes have been purified and characterized. The results suggest that the H channel is not functionally important in the prokaryotic oxidase, in agreement with the conclusion from previous work with the oxidase from Paracoccus denitrificans [Pfitzner et al. (1998) J. Biomembr. Bioenerg. 30, 89-93]. Each of the mutants in R. sphaeroides, with an exception at only one position, is enzymatically active and pumps protons in reconstituted proteoliposomes. This includes H456A, where in the P. denitrificans oxidase a leucine residue substituted for the corresponding residue resulted in inactive enzyme. The only mutations that result in completely inactive enzyme in the set examined in the R. sphaeroides oxidase are in R52, a residue that, along with Q471, appears to be hydrogen-bonded to the formyl group of heme a in the X-ray structures. To characterize the interactions between this residue and the heme group, resonance Raman spectra of the R52 mutants were obtained. The frequency of the heme a formyl stretching mode in the R52A mutant is characteristic of that seen in non hydrogen-bonded model heme a complexes. Thus the data confirm the presence of hydrogen bonding between the heme a formyl group and the R52 side chain, as suggested from crystallographic data. In the R52K mutant, this hydrogen bonding is maintained by the lysine residue, and this mutant enzyme retains near wild type activity. The heme a formyl frequency is also affected by mutation of Q471, confirming the X-ray models that show this residue also has hydrogen-bonding interactions with the formyl group. Unlike R52, however, Q471 does not appear to be critical for the enzyme function. PMID- 10715120 TI - Effects of mutations in M4 of the gastric H+,K+-ATPase on inhibition kinetics of SCH28080. AB - The effects of site-directed mutagenesis were used to explore the role of residues in M4 on the apparent Ki of a selective, K+-competitive inhibitor of the gastric H+,K+ ATPase, SCH28080. A double transfection expression system is described, utilizing HEK293 cells and separate plasmids encoding the alpha and beta subunits of the H+,K+-ATPase. The wild-type enzyme gave specific activity (micromoles of Pi per hour per milligram of expressed H+,K+-ATPase protein), apparent Km for ammonium (a K+ surrogate), and apparent Ki for SCH28080 equal to the H+, K+-ATPase purified from hog gastric mucosa. Amino acids in the M4 transmembrane segment of the alpha subunit were selected from, and substituted with, the nonconserved residues in M4 of the Na+, K+-ATPase, which is insensitive to SCH28080. Most of the mutations produced competent enzyme with similar Km,app values for NH4+ and Ki,app for SCH28080. SCH28080 affinity was decreased 2-fold in M330V and 9-fold in both M334I and V337I without significant effect on Km,app. Hence methionine 334 and valine 337 participate in binding but are not part of the NH4+ site. Methionine 330 may be at the periphery of the inhibitor site, which must have minimum dimensions of approximately 16 x 8 x 5 A and be accessible from the lumen in the E2-P conformation. Multiple sequence alignments place the membrane surface near arginine 328, suggesting that the side chains of methionine 334 and valine 337, on one side of the M4 helix, project into a binding cavity within the membrane domain. PMID- 10715121 TI - ATP-dependent substrate occlusion by the human erythrocyte sugar transporter. AB - Human erythrocyte sugar transport presents a functional complexity that is not explained by existing models for carrier-mediated transport. It has been suggested that net sugar uptake is the sum of three serial processes: sugar translocation, sugar interaction with an intracellular binding complex, and the release from this complex into bulk cytosol. The present study was carried out to identify the erythrocyte sugar binding complex, to determine whether sugar binding occurs inside or outside the cell, and to determine whether this binding complex is affected by cytosolic ATP or transporter quaternary structure. Sugar binding assays using cells and membrane protein fractions indicate that sugar binding to erythrocytes is quantitatively accounted for by sugar binding to the hexose transport protein, GluT1. Kinetic analysis of net sugar fluxes indicates that GluT1 sugar binding sites are cytoplasmic. Intracellular ATP increases GluT1 sugar binding capacity from 1 to 2 mol of 3-O-methylglucose/mol GluT1 and inhibits the release of bound sugar into cytosol. Reductant-mediated, tetrameric GluT1 dissociation into dimeric GluT1 is associated with the loss of ATP and 3-O methylglucose binding. We propose that sugar uptake involves GluT1-mediated, extracellular sugar translocation into an ATP-dependent cage formed by GluT1 cytoplasmic domains. Caged or occluded sugar has three possible fates: (1) transport out of the cell (substrate cycling); (2) interaction with sugar binding sites within the cage, or (3) release into bulk cytosol. We show how this hypothesis can account for the complexity of erythrocyte sugar transport and its regulation by cytoplasmic ATP. PMID- 10715122 TI - Annexins V and XII insert into bilayers at mildly acidic pH and form ion channels. AB - The functional hallmark of annexins is the ability to bind to the surface of phospholipid membranes in a reversible, Ca(2+)-dependent manner. We now report that human annexin V and hydra annexin XII reversibly bound to phospholipid vesicles in the absence of Ca(2+) at low pH; half-maximal vesicle association occurred at pH 5.3 and 5. 8, respectively. The following biochemical data support the hypothesis that these annexins insert into bilayers at mildly acidic pH. First, a photoactivatable reagent (3-trifluoromethyl)-3-(m [(125)I]iodophenyl)diazirine) which selectively labels proteins exposed to the hydrophobic domain of bilayers reacted with these annexins at pH 5.0 and below but not at neutral pH. Second, in a Triton X-114 partitioning assay, annexins V and XII act as integral membrane proteins at low pH and as hydrophilic proteins at neutral pH; in the presence of phospholipids half-maximal partitioning into detergent occurred at pH approximately 5.0. Finally, annexin V or XII formed single channels in phospholipid bilayers at low pH but not at neutral pH. A model is discussed in which the concentrations of H(+) and Ca(2+) regulate the reversible conversion of three forms of annexins-soluble, peripheral membrane, and transmembrane. PMID- 10715123 TI - Detailed analysis of the phosphorylation of the human La (SS-B) autoantigen. (De)phosphorylation does not affect its subcellular distribution. AB - The La (SS-B) autoantigen is an evolutionarily conserved phosphoprotein which plays an important role, most likely as an RNA chaperone, in various processes, such as the biosynthesis and maturation of RNA polymerase III transcripts in the cell nucleus and (internal) initiation of translation in the cytoplasm. In this study, the phosphorylation state of this protein from human HeLa and HEp-2 cells was characterized by high-resolution two-dimensional IEF/SDS-PAGE analysis, and phosphorylation sites were mapped by nanoelectrospray mass spectrometry. Furthermore, the effect of phosphorylation at the sites identified on the subcellular distribution of the protein was studied by site-directed mutagenesis. At least 14 isoelectric isoforms were discerned on 2-D gels with La protein from both types of cells. Metabolic labeling in combination with alkaline phosphatase treatment revealed that only a limited number of these isoforms could be attributed to phosphorylation. Four phosphorylation sites, Thr-302, Ser-325, Thr 362, and Ser-366, were mapped by mass spectrometric analysis of the isolated La protein from HeLa cells or the carboxy-terminal half of this protein. The analysis of mutants of La, in which the respective phosphorylated residues were replaced by either a neutral (alanine) or an acidic (aspartate) residue, by microinjection into Xenopus laevis oocytes on the one hand and transfection of HEp-2 cells on the other hand revealed that the subcellular distribution of this protein was not affected by these amino acid substitutions. These results strongly suggest that the signals that determine the subcellular distribution of this protein are not regulated by (de)phosphorylation of the target residues examined. PMID- 10715124 TI - Photoaffinity labeling of Torpedo nicotinic receptor with the agonist [3H]DCTA: identification of amino acid residues which contribute to the binding of the ester moiety of acetylcholine. AB - Torpedo marmorata acetylcholine binding sites were photolabeled using 360 nm light, at equilibrium in the desensitized state, with the agonist [3H]DCTA utilizing the CeIV/glutathione procedure described previously (Grutter, et al. (1999) Biochemistry 38, 7476-7484). Photoincorporation of [3H]DCTA was concentration-dependent with a maximum of 7.5% specific labeling on the alpha subunit and 1.2% on the gamma-subunit. The apparent dissociation constants for labeling of the alpha- and gamma-subunits were 2.2 +/- 1.1 and 3.6 +/- 2.8 microM, respectively. The alpha-chains isolated from receptor-rich membranes photolabeled in the absence or in the presence of carbamylcholine were cleaved with CNBr using an efficient "in gel" procedure. The resulting peptide fragments were purified by HPLC and further submitted to trypsinolysis. The digest was analyzed by HPLC leading to a single radioactive peak which, by microsequencing, revealed two sequences extending from alpha Lys-179 and from alpha His-186, respectively. Radioactive signals could be unambiguously attributed to positions corresponding to residues alpha Tyr-190, alpha Cys-192, alpha Cys-193, and alpha Tyr-198. These four identified [3H]DCTA-labeled residues, which have been also labeled with other affinity and photoaffinity probes including the agonist [3H]nicotine, belong to loop C of the ACh binding site. The chemical structure of [3H]DCTA, together with its well-defined and powerful photochemical reactivity, provides convincing evidence that loop C-labeled residues are primarily involved in the interaction with the ester moiety of acetylcholine. PMID- 10715125 TI - The glycan domain of thrombopoietin enhances its secretion. AB - Thrombopoietin (TPO) is the major regulator of megakaryocyte development and platelet production. The hormone is structurally characterized by an amino terminal receptor binding domain (amino acid residues 1-152) predicted to encode a left-handed four-helix bundle structure, and a carboxyl-terminal domain (residues 153-335) that is remarkable for its abundant carbohydrate modification and a lack of homology to other proteins. To investigate the functional role of the carboxyl-terminal glycan domain, we generated truncated forms of murine TPO (TPO1-238, TPO1-174, and TPO1-152) and glycomuteins in which the predicted asparagine (N)-linked sites of glycosylation were sequentially mutated to glutamine (Q), and assayed their secretion and function by comparing them to the native sequence (TPO1-335). Following transient transfection of the corresponding cDNA expression vectors into mammalian cell lines, the secretory efficiencies of the proteins were compared with those of the native hormone. Transfection efficiencies were monitored by cotransfection and reporter gene assay, and TPO secretion was assessed by functional and immunologic assays. We found that full length TPO was 5-29-fold more efficiently secreted than any of the truncated forms of the hormone in fibroblast and hepatocyte cell lines. Elimination of carboxyl-terminal sites of N-linked glycosylation had a minor impact on secretion of the protein. We conclude that the carboxyl-terminal domain of TPO serves the important role of enhancing secretion of the protein, and in this manner functions as a prosequence. PMID- 10715126 TI - Determining the DNA bending angle induced by non-specific high mobility group-1 (HMG-1) proteins: a novel method. AB - To study the DNA bending induced by non-sequence-specific HMG-1 domain proteins, we have engineered a fusion protein linking the yeast NHP6A with a sequence specific DNA binding domain, the DNA binding domain of the Hin recombinase, Hin DBD. A series of biochemical experiments were carried out to characterize the DNA binding property of this fusion protein. Our data showed that the fusion protein not only specifically recognizes a DNA fragment containing the Hin-DBD binding site, but also binds DNA with a higher affinity in comparison with either domain alone. Both domains of the fusion protein are bound to the DNA in juxtaposition. Permutation assays showed that the fusion protein induced a DNA bending at the site of NHP6A binding by an estimated value of 63 degrees. We believe that this experimental design provides an effective vehicle to determine the DNA bending induced by nonspecific HMG-1 proteins. PMID- 10715127 TI - Histidine 450 plays a critical role in catalysis and, with Ca2+, contributes to the substrate specificity of aminopeptidase A. AB - Aminopeptidase A (EC 3.4.11.7, APA) is a 130 kDa membrane-bound protease that contains the HEXXH consensus sequence found in the zinc metalloprotease family, the zincins. In addition to the catalytic zinc atom, APA contains a Ca2+ ion that increases its enzymatic activity. Aligning the sequences of the mouse APA, APN, and other monozinc aminopeptidases led to the identification of a conserved histidine (His 450 in mouse APA). Replacing this residue with a phenylalanine (Phe 450) by site-directed mutagenesis resulted in markedly lower levels of APA activity and in a change in the sensitivity of APA to Ca2+ (the EC50 for Ca2+ was 25 microM in the wild type and only 279 microM in the mutant). Kinetic studies, with a supramaximal Ca2+ concentration (4 mM), showed that the Km of the mutant enzyme for the substrate alpha-L-glutamyl-beta-naphthylamide was 25 times higher than that of the wild type, whereas the kcat value was much lower (factor of 22). Thus, overall, the wild-type enzyme had a cleavage efficiency that was 571 times higher than that of the mutant. The inhibitory potencies of two different classes of inhibitors, a glutamate thiol and a glutamate phosphonate compound, were significantly lower (factors of 19 and 22, respectively) for the mutated enzyme than for the wild-type enzyme. In contrast, inhibition by lysine thiol was unaffected. These data strongly suggest that His 450 is critical for catalytic activity and is involved in substrate binding via interaction with the P1 carboxylate side chain of the substrate. Furthermore, His 450, together with Ca2+, may contribute to the substrate specificity of APA for N-terminal acidic amino acid residues. PMID- 10715128 TI - Binding of nicotinamide adenine dinucleotide phosphate to the tetratricopeptide repeat domains at the N-terminus of p67PHOX, a subunit of the leukocyte nicotinamide adenine dinucleotide phosphate oxidase. AB - The nicotinamide adenine dinucleotide phosphate (NADPH) binding site of the NADPH oxidase complex is believed to be located on the beta, subunit of cytochrome b558. However, our previous studies showed that p67PHOX also contains an NADPH binding site that is essential for normal oxidase activity and that p67PHOX is able to mediate a slow electron transfer from a reduced pyridine nucleotide to an artificial electron acceptor. Using both affinity labeling and fluorescence quenching, we have obtained further evidence that p67PHOX is able to bind NADPH. We have used a number of truncated forms of p67PHOX, including p67PHOX(1-243), p67PHOX(1-210), p67PHOX(1-199), and p67PHOX(244-526) (where the numbers represent the initial and final amino acids in the truncated p67PHOX) in order to localize the binding site. We found that NADPH could bind to p67PHOX(1-243), p67PHOX(1 210), and p67PHOX(1-199) but not to p67PHOX(244-526). The p67PHOX(1-199) fragment consists largely of four tetratricopeptide (TPR) domains. We showed further that Rac2-GTP gamma S and to a lesser extent Rac2-GDP beta S could modulate the binding of NADPH to p67PHOX. PMID- 10715129 TI - Tracking sliding clamp opening and closing during bacteriophage T4 DNA polymerase holoenzyme assembly. AB - The bacteriophage T4 DNA polymerase holoenzyme, consisting of the DNA polymerase (gp43), the sliding clamp (gp45), and the clamp loader (gp44/62), is loaded onto DNA in an ATP-dependent, multistep reaction. The trimeric, ring-shaped gp45 is loaded onto DNA such that the DNA passes through the center of the ring. gp43 binds to this complex, thereby forming a topological link with the DNA and increasing its processivity. Using stopped-flow fluorescence-resonance energy transfer, we have investigated opening and closing of the gp45 ring during the holoenzyme assembly process. Two amino acids that lie on opposite sides of the gp45 subunit interface, W91 and V162C labeled with coumarin, were used as the fluorescence donor and acceptor, respectively. Free in solution, gp45 has two closed subunit interfaces with W91 to V162-coumarin distances of 19 A and one open subunit interface with a W91 to V162C-coumarin distance of 40 A. Making the assumption that the distance across the two closed subunit interfaces is unchanged during the holoenzyme assembly process, we have found that the distance across the open subunit interface is first increased to greater than 45 A and is then decreased to 30 A during a 10-step assembly mechanism. The gp45 ring is not completely closed in the holoenzyme complex, consistent with previous evidence suggesting that the C-terminus of gp43 is inserted into the gp45 subunit interface. Unexpectedly, ATP-hydrolysis events are coupled to only a fraction of the total distance change, with conformational changes linked to binding DNA and gp43 coupled to the majority of the total distance change. Using the nonhydrolyzable ATP analogue ATP-gamma-S results in formation of a nonproductive gp45 x gp44/62 complex; however, adding an excess of ATP to this nonproductive complex results in rapid ATP/ATP-gamma-S exchange to yield a productive gp45 x gp44/62 complex within seconds. PMID- 10715130 TI - Reactions of glutamate 1-semialdehyde aminomutase with R- and S-enantiomers of a novel, mechanism-based inhibitor, 2,3-diaminopropyl sulfate. AB - Glutamate semialdehyde aminomutase is a recognized target for selective herbicides and antibacterial agents because it provides the aminolevulinate from which tetrapyrroles are synthesized in plants and bacteria but not in animals. The reactions of the enzyme with R- and S-enantiomers of a novel compound, diaminopropyl sulfate, designed as a mechanism-based inhibitor of the enzyme are described. The S-enantiomer undergoes transamination without significantly inactivating the enzyme. The R-enantiomer inactivates the enzyme rapidly. Inactivation is accompanied by the formation of a 520 nm-absorbing chromophore and by the elimination of sulfate. The inactivation is attenuated by simultaneous transamination of the enzyme to its pyridoxamine phosphate form but inclusion of succinic semialdehyde to reverse the transamination leads to complete inactivation. The inactivation is attributed to further reactions arising from generation of an external aldimine between the pyridoxal phosphate cofactor and the 2,3-diaminopropene that results from enzyme-catalyzed beta-elimination of sulfate. PMID- 10715131 TI - Identification of the metal-binding sites of restriction endonucleases by Fe2+ mediated oxidative cleavage. AB - Fenton chemistry [Fenton (1894) J. Chem. Soc. 65, 899-910] techniques were employed to identify the residues involved in metal binding located at the active sites of restriction endonucleases. This process uses transition metals to catalytically oxidize the peptide linkage that is in close proximity to the amino acid residues involved in metal ligation. Fe2+ was used as the redox-active transition metal. It was expected that Fe2+ would bind to the endonucleases at the Mg2+-binding site [Liaw et al. (1993) Biochemistry 32, 7999-4003; Ermacora et al. (1992) Proc. Natl. Acad. Sci. U.S.A. 89, 6383-6387; Soundar and Colman (1993) J. Biol. Chem. 268, 5264-5271; Wei et al. (1994) Biochemistry 33, 7931-7936; Ettner et al. (1995) Biochemistry 34, 22-31; Hlavaty and Nowak (1997) Biochemistry 36, 15515-15525). Fe2+-mediated oxidation was successfully performed on TaqI endonulease, suggesting that this approach could be applied to a wide array of endonucleases [Cao and Barany (1998) J. Biol. Chem. 273, 33002-33010]. The restriction endonucleases BamHI, FokI, BglI, BglII, PvuII, SfiI, BssSI, BsoBI, EcoRI, EcoRV, MspI, and HinP1I were subjected to oxidizing conditions in the presence of Fe2+ and ascorbate. All proteins were inactivated upon treatment with Fe2+ and ascorbate. BamHI, FokI, BglI, BglII, PvuII, SfiI, BssSI, and BsoBI were specifically cleaved upon treatment with Fe2+/ascorbate. The site of Fe2+/ascorbate-induced protein cleavage for each enzyme was determined. The Fe2+ mediated oxidative cleavage of BamHI occurs between residues Glu77 and Lys78. Glu77 has been shown by structural and mutational studies to be involved in both metal ligation and catalysis [Newman et al. (1995) Science 269, 656-663; Viadiu and Aggarwal (1998) Nat. Struct. Biol. 5, 910-916; Xu and Schildkraut (1991) J. Biol. Chem. 266, 4425-4429]. The sites of Fe2+/ascorbate-induced cleavage for PvuII, FokI, BglI, and BsoBI agree with the metal-binding sites identified in their corresponding three-dimensional structures or from mutational studies [Cheng et al. (1994) EMBO J. 13, 3297-3935; Wah et al. (1997) Nature 388, 97-100; Newman et al. (1998) EMBO J. 17, 5466-5476; Ruan et al. (1997) Gene 188, 35-39]. The metal-binding residues of BglII, SfiI, and BssSI are proposed based on amino acid sequencing of their Fe2+/ascorbate-generated cleavage fragments. These results suggest that Fenton chemistry may be a useful methodology in identifying amino acids involved in metal binding in endonucleases. PMID- 10715132 TI - Capping DNA with DNA. AB - Twelve classes of deoxyribozymes that promote an ATP-dependent "self-capping" reaction were isolated by in vitro selection from a random-sequence pool of DNA. Each deoxyribozyme catalyzes the transfer of the AMP moiety of ATP to its 5' terminal phosphate group, thereby forming a 5',5'-pyrophosphate linkage. An identical DNA adenylate structure is generated by the T4 DNA ligase during enzymatic DNA ligation. A 41-nucleotide class 1 deoxyribozyme requires Cu(2+) as a cofactor and adopts a structure that recognizes both the adenine and triphosphate moieties of ATP or dATP. The catalytic efficiency for this DNA, measured at 10(4) M(-1) x min(-1) using either ATP or dATP as substrate, is similar to other catalytic nucleic acids that use small substrates. Chemical probing and site-directed mutagenesis implicate the formation of guanine quartets as critical components of the active structure. The observation of ATP-dependent "self-charging" by DNA suggests that DNA could be made to perform the reactions typically associated with DNA cloning, but without the assistance of protein enzymes. PMID- 10715133 TI - Kinetic framework for ligation by an efficient RNA ligase ribozyme. AB - The class I RNA ligase ribozyme, isolated previously from random sequences, performs an efficient RNA ligation reaction. It ligates two substrate RNAs, promoting the attack of the 3'-hydroxyl of one substrate upon the 5'-triphosphate of the other substrate with release of pyrophosphate. This ligation reaction has similarities to the reaction catalyzed by RNA polymerases. Using data from steady state kinetic measurements and pulse-chase/pH-jump experiments, we have constructed minimal kinetic frameworks for two versions of the class I ligase, named 207t and 210t. For both ligases, as well as for the self-ligating parent ribozyme, the rate constant for the chemical step (k(c)) is log-linear with pH in the range 5.7-8.0. At physiological pH, the k(c) is 100 min(-1), a value similar to those reported for the fastest naturally occurring ribozymes. At higher pH, product release is limiting for both 207t and 210t. The 210t ribozyme, with its faster product release, attains multiple-turnover rates (k(cat) = 360 min(-1), pH 9.0) exceeding those of 207t and other reported ribozyme reactions. The kinetic framework for the 210t ribozyme describes the limits of this catalysis and suggests how key steps can be targeted for improvement using design or combinatorial approaches. PMID- 10715134 TI - Tryptophan-anchored transmembrane peptides promote formation of nonlamellar phases in phosphatidylethanolamine model membranes in a mismatch-dependent manner. AB - To better understand the mutual interactions between lipids and membrane-spanning peptides, we investigated the effects of tryptophan-anchored hydrophobic peptides of various lengths on the phase behavior of 1,2 dielaidoylphosphatidylethanolamine (DEPE) dispersions, using (31)P nuclear magnetic resonance and small-angle X-ray diffraction. Designed alpha-helical transmembrane peptides (WALPn peptides, with n being the total number of amino acids) with a hydrophobic sequence of leucine and alanine of varying length, bordered at both ends by two tryptophan membrane anchors, were used as model peptides and were effective at low concentrations in DEPE. Incorporation of 2 mol % of relatively short peptides (WALP14-17) lowered the inverted hexagonal phase transition temperature (T(H)) of DEPE, with an efficiency that seemed to be independent of the extent of hydrophobic mismatch. However, the tube diameter of the H(II) phase induced by the peptides was clearly dependent on mismatch and decreased with shorter peptide length. Longer peptides (WALP19-27) induced a cubic phase, both below and above T(H). Incorporation of WALP27, which is significantly longer than the DEPE bilayer thickness, did not stabilize the bilayer. The longest peptide used, WALP31, hardly affected the lipid's phase behavior, and appeared not to incorporate into the bilayer. The consequences of hydrophobic mismatch between peptides and lipids are therefore more dramatic with shorter peptides. The data allow us to suggest a detailed molecular model of the mechanism by which these transmembrane peptides can affect lipid phase behavior. PMID- 10715135 TI - Thiol cross-linking of cytoplasmic loops in the lactose permease of Escherichia coli. AB - The N- and C-terminal halves of lactose permease, each with a single-Cys residue in a cytoplasmic loop, were coexpressed, and cross-linking was studied in the absence or presence of ligand. Out of the 68 paired-Cys mutants in cytoplasmic loops IV/V and VIII/IX or X/XI, three pairs in loop IV/V and X/XI, (i) Arg135 --> Cys/Thr338 --> Cys, (ii) Arg134 --> Cys/Val343 --> Cys, and (iii) Arg134 --> Cys/Phe345 --> Cys, form a spontaneous disulfide bond, indicating that loops IV/V and X/XI are in close proximity. In addition, specific paired-Cys residues in loop IV/V (132-138) and loop VIII/IX (282-290) or loop X/XI (335-345) cross-link with iodine and/or the homobifunctional cross-linking agents N, N'-o phenylenedimaleimide, N,N'-p-phenylenedimaleimide, and 1, 6-bis(maleimido)hexane. The results demonstrate that loop IV/V is close to both loop VIII/IX and loop X/XI. On the other hand, similar though less extensive cross-linking studies indicate that neither the N terminus nor loop II/III appear to be close to loops VIII/IX or X/XI. The findings suggest that the longer cytoplasmic loops are highly flexible and interact in a largely random fashion. However, although a Cys residue at position 134 in loop IV/V, for example, is able to cross-link with a Cys residue at each position in loop VIII/IX or loop X/XI, Cys residues at other positions in loop IV/V exhibit markedly different cross-linking patterns. Therefore, although the domains appear to be very flexible, the interactions are not completely random, suggesting that there are probably at least some structural constraints that limit the degree of flexibility. In addition, evidence is presented suggesting that ligand binding induces conformational alterations between loop IV/V and loop VIII/IX or X/XI. PMID- 10715136 TI - Activation of JNK3 alpha 1 requires both MKK4 and MKK7: kinetic characterization of in vitro phosphorylated JNK3 alpha 1. AB - JNK3 alpha 1 is predominantly a neuronal specific MAP kinase that is believed to require, like all MAP kinases, both threonine and tyrosine phosphorylation for maximal enzyme activity. In this study we investigated the in vitro activation of JNK3 alpha 1 by MAP kinase kinase 4 (MKK4), MAP kinase kinase 7 (MKK7), and the combination of MKK4 + MKK7. Mass spectral analysis showed that MKK7 was capable of monophosphorylating JNK3 alpha 1 in vitro, whereas both MKK4 and MKK7 were required for bisphosphorylation and maximal enzyme activity. Measuring catalysis under Vmax conditions showed MKK4 + MKK7-activated JNK3 alpha 1 had Vmax 715-fold greater than nonactivated JNK3 alpha 1 and MKK7-activated JNK3 alpha 1 had Vmax 250-fold greater than nonactivated JNK3 alpha 1. In contrast, MKK4-activated JNK3 alpha 1 had no increase in Vmax compared to nonactivated levels and had no phosphorylation on the basis of mass spectrometry. These data suggest that MKK7 was largely responsible for JNK3 alpha 1 activation and that a single threonine phosphorylation may be all that is needed for JNK3 alpha 1 to be active. The steady-state rate constants kcat, Km(GST-ATF2++), and Km(ATP) for both monophosphorylated and bisphosphorylated JNK3 alpha 1 were within 2-fold between the two enzyme forms, suggesting the addition of tyrosine phosphorylation does not affect the binding of ATF2, ATP, or maximal turnover. Finally, the MAP kinase inhibitor, SB203580, had an IC50 value approximately 4-fold more potent on the monophosphorylated JNK3 alpha 1 compared to the bisphosphorylated JNK3 alpha 1, suggesting only a modest effect of tyrosine phosphorylation on inhibitor binding. PMID- 10715137 TI - Substrate recognition through a PDZ domain in tail-specific protease. AB - Tail-specific protease (Tsp) is a periplasmic enzyme that selectively degrades proteins bearing a nonpolar C-terminus. Its substrate specificity suggests that Tsp may contain a substrate recognition domain, which selectively binds to the nonpolar C-termini of substrate proteins, separate from its catalytic site. In this work, we show that substrate recognition of Tsp is mediated by a PDZ domain, a small protein module that promotes protein-protein interactions by binding to internal or C-terminal sequences of their partner proteins. Partial proteolysis by V8 protease at a single peptide bond immediately N-terminal to the PDZ domain resulted in two distinct and relatively stable fragments and complete loss of catalytic activity. Photoaffinity labeling with a fluorescent nonpolar peptide caused the covalent attachment of the peptide to a single site on the Tsp protein. Systematic deletion mutagenesis of Tsp localized the binding site to amino acids 206-307, a region that completely encompasses the putative PDZ domain (217-301). The isolated PDZ domain (amino acids 206-334) is capable of folding into a well-behaved structure and binds to a nonpolar peptide with a dissociation constant (K(D)) of 1.9 microM, similar to that of the intact Tsp protein. Site directed mutagenesis of a surface residue at the peptide binding site of the PDZ domain, valine 229, to Glu or Gln resulted in an increase in the K(M) value but had no effect on the k(cat) value. The use of a separate substrate recognition domain such as a PDZ domain may be a general mechanism for achieving selective protein degradation. PMID- 10715138 TI - Crystal structure of Escherichia coli malate synthase G complexed with magnesium and glyoxylate at 2.0 A resolution: mechanistic implications. AB - The crystal structure of selenomethionine-substituted malate synthase G, an 81 kDa monomeric enzyme from Escherichia coli has been determined by MAD phasing, model building, and crystallographic refinement to a resolution of 2.0 A. The crystallographic R factor is 0.177 for 49 242 reflections observed at the incident wavelength of 1.008 A, and the model stereochemistry is satisfactory. The basic fold of the enzyme is that of a beta8/alpha8 (TIM) barrel. The barrel is centrally located, with an N-terminal alpha-helical domain flanking one side. An inserted beta-sheet domain folds against the opposite side of the barrel, and an alpha-helical C-terminal domain forms a plug which caps the active site. Malate synthase catalyzes the condensation of glyoxylate and acetyl-coenzyme A and hydrolysis of the intermediate to yield malate and coenzyme A, requiring Mg(2+). The structure reveals an enzyme-substrate complex with glyoxylate and Mg(2+) which coordinates the aldehyde and carboxylate functions of the substrate. Two strictly conserved residues, Asp631 and Arg338, are proposed to provide concerted acid-base chemistry for the generation of the enol(ate) intermediate of acetyl-coenzyme A, while main-chain hydrogen bonds and bound Mg(2+) polarize glyoxylate in preparation for nucleophilic attack. The catalytic strategy of malate synthase appears to be essentially the same as that of citrate synthase, with the electrophile activated for nucleophilic attack by nearby positive charges and hydrogen bonds, while concerted acid-base catalysis accomplishes the abstraction of a proton from the methyl group of acetyl-coenzyme A. An active site aspartate is, however, the only common feature of these two enzymes, and the active sites of these enzymes are produced by quite different protein folds. Interesting similarities in the overall folds and modes of substrate recognition are discussed in comparisons of malate synthase with pyruvate kinase and pyruvate phosphate dikinase. PMID- 10715139 TI - Q-band ENDOR (electron nuclear double resonance) of the high-affinity ubisemiquinone center in cytochrome bo3 from Escherichia coli. AB - Electron nuclear double resonance (ENDOR) was performed on the protein-bound, stabilized, high-affinity ubisemiquinone radical, QH*-, of bo3 quinol oxidase to determine its electronic spin distribution and to probe its interaction with its surroundings. Until this present work, such ENDOR studies of protein-stabilized ubisemiquinone centers have only been done on photosynthetic reaction centers whose function is to reduce a ubiquinol pool. In contrast, QH*- serves to oxidize a ubiquinol pool in the course of electron transfer from the ubiquinol pool to the oxygen-consuming center of terminal bo3 oxidase. As documented by large hyperfine couplings (>10 MHz) to nonexchangeable protons on the QH*- ubisemiquinone ring, we provide evidence for an electronic distribution on QH*- that is different from that of the semiquinones of reaction centers. Since the ubisemiquinone itself is physically nearly identical in both QH*- and the bacterial photosynthetic reaction centers, this electronic difference is evidently a function of the local protein environment. Interaction of QH*- with this local protein environment was explicitly shown by exchangeable deuteron ENDOR that implied hydrogen bonding to the quinone and by weak proton hyperfine couplings to the local protein matrix. PMID- 10715141 TI - Ultraviolet-resonance raman spectroscopy of the filamentous virus pf3: interactions of trp 38 specific to the assembled virion subunit PMID- 10715140 TI - Modulation of MutS ATP hydrolysis by DNA cofactors. AB - Escherichia coli MutS protein, which is required for mismatch repair, has a slow ATPase activity that obeys Michalelis-Menten kinetics. At 37 degrees C, the steady-state turnover rate for ATP hydrolysis is 1.0 +/- 0.3 min(-1) per monomer equivalent with a K(m) of 33 +/- 6 microM. Hydrolysis is competitively inhibited by the ATP analogues AMPPNP and ATPgammaS, with K(i) values of 4 microM in both cases, and by ADP with a K(i) of 40 microM. The rate of ATP hydrolysis is stimulated 2-5-fold by short hetero- and homoduplex DNAs. The concentration of DNA cofactor that yields half-maximal stimulation is lowest for oligodeoxynucleotide duplexes that contain a mismatched base pair. Pre-steady state chemical quench analysis has demonstrated a substoichiometric initial burst of ADP formation by free MutS that is governed by a rate constant of 78 min(-1), indicating that the rate-limiting step for the steady-state reaction occurs after hydrolysis. Prebinding of MutS to homoduplex DNA does not alter the burst kinetics or amplitude but only increases the steady-state rate. In contrast, binding of the protein to heteroduplex DNA abolishes the burst of ADP formation, indicating that the rate-limiting step now occurs before hydrolysis. Gel filtration analysis indicates that the MutS dimer assembles into higher order oligomers in a concentration-dependent manner, and that ATP binding shifts this equilibrium to favor assembly. These results, together with kinetic findings, indicate nonequivalence of subunits within a MutS oligomer with respect to ATP hydrolysis and DNA binding. PMID- 10715142 TI - alpha(2) Adrenoceptor agonists as potential analgesic agents. 2. Discovery of 4 (4-Imidazo)-1,3-dimethyl-6,7-dihydrothianaphthene [corrected] as a high-affinity ligand for the alpha(2D) adrenergic receptor. PMID- 10715143 TI - 1-(2-Nitrophenyl)thiosemicarbazides: a novel class of potent, orally active non peptide antagonist for the bradykinin B(2) receptor. PMID- 10715144 TI - Dual function glutamate-related ligands: discovery of a novel, potent inhibitor of glutamate carboxypeptidase II possessing mGluR3 agonist activity. PMID- 10715145 TI - 4-[5-Methyl-3-phenylisoxazol-4-yl]- benzenesulfonamide, valdecoxib: a potent and selective inhibitor of COX-2. PMID- 10715146 TI - Potent anti-HIV (type 1 and type 2) activity of polyoxometalates: structure activity relationship and mechanism of action. AB - A series of polyoxometalates have been synthesized and evaluated for their inhibitory effects on HIV-1(III(B)) and HIV-1(ROD) replication in MT-4 cells. All compounds showed activity against HIV-1 and HIV-2, but the antiviral potency of the heteropolytungstates varied considerably depending on their chemical structure. The antiviral activity of single, double, and triple Keggin-type of compounds against HIV-1(III(B)) replication was comparable (IC(50): 0.4-0.5 microgram/mL), whereas HIV-2(ROD) appeared to become less sensitive with the increasing number of Keggin structures per compound. The same trend was observed for single and double Dawson structures. Some of these compounds were examined for their inhibitory effect on the replication of HIV-1(RF) and SIV(MAC(251)) in MT-4 cells. Their anti-HIV-1(RF) and anti-SIV(MAC(251)) potencies were comparable to those for the HIV-1(III(B)) or HIV-2(ROD) strain, respectively. The polyoxometalates represent a class of polyanionic compounds, which block the binding of the envelope glycoprotein gp120 of HIV to CD4(+) cells. The compounds interfered with the binding of anti-CD4 mAb to the OKT4A/Leu3a epitope of the CD4 receptor, compound 24 being the most active in this regard, and inhibited the binding of anti-gp120 mAb to infected MT-4 cells. None of the polyoxometalates inhibited the binding of a specific CXCR4 mAb to SUP-T1 cells, suggesting that they do not interact with CXCR4, the main co-receptor for T-tropic HIV strains, and thus act as virus binding, and not as fusion, inhibitors. PMID- 10715147 TI - Design of potent dicyclic (4-10/5-8) gonadotropin releasing hormone (GnRH) antagonists. AB - With the ultimate goal of identifying a consensus bioactive conformation of GnRH antagonists, the compatibility of a number of side chain to side chain bridges in bioactive analogues was systematically explored. In an earlier publication, cyclo[Asp(4)-Dpr(10)]GnRH antagonists with high potencies in vitro and in vivo had been identified.(1) Independently from Dutta et al. (2) and based on structural considerations, the cyclic [Glu(5)-Lys(8)] constraint was also found to be tolerated in GnRH antagonists. We describe here a large number of cyclic (4 10) and (5-8) and dicyclic (4-10/5-8) GnRH antagonists optimized for affinity to the rat GnRH receptor and in vivo antiovulatory potency. The most potent monocyclic analogues were cyclo(4-10)[Ac-DNal(1), DFpa(2),DTrp(3),Asp(4),DArg(6),Xaa(10)]GnRH with Xaa = D/LAgl (1, K(i) = 1.3 nM) or Dpr (2, K(i) = 0.36 nM), which completely blocked ovulation in cycling rats after sc administration of 2.5 microgram at noon of proestrus. Much less potent were the closely related analogues with Xaa = Dbu (3, K(i) = 10 nM) or cyclo(4 10)[Ac-DNal(1), DFpa(2),DTrp(3),Glu(4),DArg(6),D/LAgl(10)]GnRH (4, K(i) = 1.3 nM). Cyclo(5-8)[Ac-DNal(1),DCpa(2),DTrp(3),Glu(5),DArg++ +(6),Lys(8), DAla(10)]GnRH (13), although at least 20 times less potent in the AOA than 1 or 2 with similar GnRHR affinity (K(i) = 0.84 nM), was found to be one of the most potent in a series of closely related cyclo(5-8) analogues with different bridge lengths and bridgehead chirality. The very high affinity of cyclo(5,5'-8)[Ac DNal(1), DCpa(2),DPal(3),Glu(5)(betaAla),DArg(6),(D or L)Agl,(8)DAla(10)]GnRH 14 (K(i) = 0.15 nM) correlates well with its high potency in vivo (full inhibition of ovulation at 25 microgram/rat). Dicyclo(4-10/5-8)[Ac DNal(1),DCpa(2),DTrp(3),Asp (4),Glu(5),DArg(6), Lys(8),Dpr(10)]GnRH (24, K(i) = 0.32 nM) is one-fourth as potent as 1 or 2, in the AOA; this suggests that the introduction of the (4-10) bridge in 13, while having little effect on affinity, restores functional/conformational features favorable for stability and distribution. To further increase potency of dicyclic antagonists, the size and composition of the (5-8) bridge was varied. For example, the substitution of Xbb(5') by Gly (30, K(i) = 0.16 nM), Sar (31, K(i) = 0.20 nM), Phe (32, K(i) = 0.23 nM), DPhe (33, K(i) = 120 nM), Arg (36, K(i) = 0.20 nM), Nal (37, K(i) = 4.2 nM), His (38, K(i) = 0.10 nM), and Cpa (39, K(i) = 0.23 nM) in cyclo(4-10/5,5' 8)[Ac-DNal(1),DCpa(2),DPal(3),Asp(4),G lu(5)(Xbb(5')), DArg(6),Dbu,(8)Dpr(10)]GnRH yielded several very high affinity analogues that were 10, ca. 10, 4, >200, 1, ca. 4, >2, and 2 times less potent than 1 or 2, respectively. Other scaffolds constrained by disulfide (7, K(i) = 2.4 nM; and 8, K(i) = 450 nM), cyclo[Glu(5)-Aph(8)] (16, K(i) = 20 nM; and 17, K(i) = 0.28 nM), or cyclo[Asp(5)-/Glu(5)-/Asp(5)(Gly(5'))-Amp(8)] (19, K(i) = 1.3 nM; 22, K(i) = 3.3 nM; and 23, K(i) = 3.6 nM) bridges yielded analogues that were less potent in vivo and had a wide range of affinities. The effects on biological activity of substituting DCpa or DFpa at position 2, DPal or DTrp at position 3, and DArg, DNal, or DCit at position 6 are also discussed. Interestingly, monocyclo(5 8)[Glu(5), DNal(6),Lys(8)]GnRH (18, K(i) = 1. (ABSTRACT TRUNCATED) PMID- 10715148 TI - Design of monocyclic (1-3) and dicyclic (1-3/4-10) gonadotropin releasing hormone (GnRH) antagonists. AB - Careful analysis of the NMR structures of cyclo(4-10)[Ac Delta(3)Pro(1),DFpa(2),DTrp(3),Asp(4),DNal (6), Dpr(10)]GnRH, dicyclo(4-10/5 8)[Ac-DNal(1),DCpa(2),DTrp(3), Asp(4), Glu(5),DArg(6),Lys(8),Dpr(10)]GnRH, and dicyclo(4-10/5, 5'-8)[Ac-DNal(1),DCpa(2),DPal(3),Asp(4), Glu(5)(Gly),DArg(6),Dbu(8), Dpr(10)]GnRH showed that, in the N-terminal tripeptide, a type II beta-turn around residues 1 and 2 was probable along with a gamma-turn around DTrp(3)/DPal(3). This suggested the possibility of constraining the N-terminus by the introduction of a cyclo(1-3) scaffold. Optimization of ring size and composition led to the discovery of cyclo(1-3)[Ac DAsp(1),DCpa(2),DLys(3),DNal(6), DAla(10)]GnRH (5, K(i) = 0.82 nM), cyclo(1,1' 3)[Ac-DAsp(1)(Gly), DCpa(2),DOrn(3),DNal(6),DAla(10)]GnRH (13, K(i) = 0.34 nM), cyclo(1, 1'-3)[Ac-DAsp(1)(Gly),DCpa(2),DLys(3),DNal(6),DA la(10)]GnRH (20, K(i) = 0.14 nM), and cyclo(1,1'-3)[Ac-DAsp(1)(betaAla), DCpa(2), DOrn(3),DNal(6),DAla(10)]GnRH (21, K(i) = 0.17 nM), which inhibited ovulation significantly at doses equal to or lower than 25 microgram/rat. These results were particularly unexpected in view of the critical role(s) originally ascribed to the side chains of residues 1 and 3.(1) Other closely related analogues, such as those where the [DAsp(1)(betaAla), DOrn(3)] cycle of 21 was changed to [DOrn(1)(betaAla), DAsp(3)] of cyclo(1,1'-3)[Ac-DOrn(1)(betaAla), DCpa(2),DAsp(3),DNal(6),DAla(10)]GnRH (22, K(i) = 2.2 nM) or where the size of the cycle was conserved and [DAsp(1)(betaAla), DOrn(3)] was replaced by [DGlu(1)(Gly), DOrn(3)] as in cyclo(1, 1'-3)[Ac DGlu(1)(Gly),DCpa(2),DOrn(3),DNal(6),DA la(10)]GnRH (23, K(i) = 4.2 nM), were approximately 100 and 25 times less potent in vivo, respectively. Analogues with ring sizes of 18 ?cyclo(1, 1'-3)[Ac-DGlu(1)(Gly),DCpa(2),DLys(3),DNal(6),DA la(10)]GnRH (24)? and 19 ?cyclo(1,1'-3)[Ac-DGlu(1)(betaAla),DCpa(2),DLys( 3),DNal(6), DAla(10)]GnRH (25)? atoms were also less potent than 21 with slightly higher K(i) values (1.5 and 2.2 nM, respectively). These results suggested that the N-terminal tripeptide was likely to assume a folded conformation favoring the close proximity of the side chains of residues 1 and 3. The dicyclic analogue dicyclo(1-3/4-10)[Ac-DAsp(1),DCpa(2),DLys(3),Asp (4),DNal(6), Dpr(10)]GnRH (26) was fully active at 500 microgram, with a K(i) value of 1 nM. The in vivo potency of 26 was at least 10-fold less than that of monocyclic cyclo(1-3)[Ac DAsp(1),DCpa(2),DLys(3),DNal(6), DAla(10)]GnRH (5); this suggested the existence of unfavorable interactions between the now optimized and constrained (1-3) and (4-10) cyclic moieties that must interact as originally hypothesized. Tricyclo(1 3/4-10/5-8)[Ac-DGlu(1),DCpa(2), DLys(3),Asp(4),Glu(5), DNal(6),Lys(8),Dpr(10)] GnRH (27) was inactive at 500 microgram/rat with a corresponding low affinity (K(i) = 4.6 nM) when compared to those of the most potent analogues (K(i) < 0.5 nM). PMID- 10715149 TI - Design of potent dicyclic (1-5/4-10) gonadotropin releasing hormone (GnRH) antagonists. AB - In three earlier papers, the structures and biological potencies of numerous mono and dicyclic antagonists of GnRH were reported. Among these, two families, each containing two to four members were identified that had very high antagonist potencies in an antiovulatory assay (within a factor of 2 of those of the most potent linear analogues) and high affinities (K(i) < 0.5 nM) for the rat GnRH receptor (rGnRHR). The most favored cycles bridged the side chains of residues (4 10),(1,2) (5-8),(2) (4-10/5-8),(2) (1-3),(3) and (1-3/4-10).(3) Our goal was to identify a consensus model of bioactive conformations of GnRH antagonists, yet these biocompatible constraints did not sufficiently restrain the spatial location of the N-terminal tripeptide with respect to the C-terminal heptapeptide, due largely to the rotational freedom about the bonds connecting these regions. Examination of models derived from NMR studies of cyclo(4-10) analogues suggested a large number of possible cyclic constraints such as cyclo (0-8), (1-8), or (2-8). All analogues tested with these substitutions were inactive as antiovulatory agents at 1 mg/rat (5-9) and had low affinity for rGnRHR. On the other hand, bridging positions 3 and 8 with a [DAsp(3)] to [Dbu(8)] (12, K(i) = 13 nM) or [Orn(8)] (13, K(i) = 14 nM) in the parent compound cyclo(3-8)[Ac-DNal(1),DCpa(2),DXaa(3), Arg(5),DNal(6),Xbb(8),DAla(10)]GnRH yielded analogues that blocked ovulation at 250 microgram/rat. Analogue 14 (K(i) = 2.3 nM), with a [DAsp(3), Lys(8)] bridge, was fully active at 50 microgram/rat. Loss of potency (>20-fold) was observed with the substitution of [DAsp(3)] in 14 by [DGlu(3)] in 15 (K(i) = 23 nM). Dicyclic analogues possessing the (4-10) cycle and selected (1-6), (2-6), and (2-8) cycles led to analogues that were inactive at doses of 500 microgram/rat or larger. Two analogues with (1-8/4-10) cycles (16, K(i) = 1.1 nM) or (3-8/4-10) cycles (22, K(i) = 17 nM) showed full antiovulatory potency at 250 microgram/rat. None of these substitutions yielded analogues potent enough (>80% inhibition of ovulation at 5 microgram/rat or less and K(i) < 0.5 nM) to be candidates for structural analysis by NMR. On the other hand, four dicyclic (1, 1'-5/4-10) analogues met this criterion: dicyclo(1, 1' 5/4-10)[Ac-Asp(1)(Gly),DCpa(2),DTrp(3),Asp(4),Dbu(5 ), DNal(6), Dpr(10)]GnRH (32, K(i) = 0.22 nM), dicyclo(1, 1'-5/4-10)[Ac Asp(1)(Gly),DCpa(2),DNal(3),Asp(4),Dbu(5 ), DNal(6), Dpr(10)]GnRH (34, K(i) = 0.38 nM), dicyclo(1, 1'-5/4-10)[Ac-Asp(1)(betaAla),DCpa(2), DTrp(3),Asp(4),Dbu(5),DNal(6), Dpr(10)]GnRH (40, K(i) = 0.15 nM), and dicyclo(1, 1'-5/4-10)[Ac-Glu(1)(Gly), DCpa(2),DTrp(3),Asp(4),Dbu(5),DNal(6), Dpr(10)]GnRH (41, K(i) = 0.24 nM). Since they differed slightly in terms of the (1,1'-5) bridge length (21 and 22 atoms) and bridgehead configuration, we may hypothesize that they assume similar bioactive conformations that satisfy a very discriminating receptor, since many other closely related analogues were significantly less potent. PMID- 10715150 TI - Consensus bioactive conformation of cyclic GnRH antagonists defined by NMR and molecular modeling. AB - Little is known of the conformation of peptide hormones as they interact with their receptors for a number of reasons: peptide hormones are notoriously flexible in solution, their receptors are particularly complex, and there is strong evidence that receptor-ligand interaction leading to activation is a dynamic process. Insights into the active conformation of the decapeptide gonadotropin releasing hormone (GnRH) have been obtained previously from the solution structures of four constrained GnRH antagonists ?cyclo(1-10)[Ac-Delta(3) Pro(1),DCpa(2),DTrp(3,6),NMeLeu+ ++(7), betaAla(10)]GnRH (1), cyclo(4-10)[Ac Delta(3)Pro(1),DFpa(2),DTrp(3), Asp(4),DNal(6),Dpr(10)]GnRH (2), dicyclo(4-10/5 8)[Ac-DNal(1), DCpa(2),DTrp(3),Asp(4),Glu(5),DArg(6),Lys(8),Dpr (10)]GnRH (3), and dicyclo(4-10/5-5'-8)[Ac-DNal(1),DCpa(2),DPal(3), Asp(4),Glu(5)(Gly), DArg(6),Dbu(8),Dpr(10)]GnRH (4)?. However, the precise location of the N-terminal tripeptide in the highly potent (K(i) < 0.4 nM) 2-4 remained unclear due to the lack of constraints in this region. The NMR structure of the newly discovered and potent (K(i) = 0.24 nM) dicyclo(1-1'-5/4-10)[Ac-Glu(1)(Gly),DCpa(2),DTrp(3),As p(4),Dbu(5), DNal(6),Dpr(10)]GnRH (5) now allows the definition of the conformation of this region. A combined computational analysis (consensus forcing) of compounds 2-5, designed to explore the common conformations available to them that are simultaneously consistent with the NMR data corresponding to each compound, leads to a consensus structural model for the GnRH pharmacophore. This model shares some common features with the structure of the nonpeptidic GnRH mimetic T-98475. In the course of that comparative study, two additional contact points to those proposed by the authors are identified, suggesting that this model has predictive value. PMID- 10715151 TI - Synthesis, biological activity, and molecular modeling of ribose-modified deoxyadenosine bisphosphate analogues as P2Y(1) receptor ligands. AB - The structure-activity relationships of adenosine-3', 5'-bisphosphates as P2Y(1) receptor antagonists have been explored, revealing the potency-enhancing effects of the N(6)-methyl group and the ability to substitute the ribose moiety (Nandanan et al. J. Med. Chem. 1999, 42, 1625-1638). We have introduced constrained carbocyclic rings (to explore the role of sugar puckering), non glycosyl bonds to the adenine moiety, and a phosphate group shift. The biological activity of each analogue at P2Y(1) receptors was characterized by measuring its capacity to stimulate phospholipase C in turkey erythrocyte membranes (agonist effect) and to inhibit its stimulation elicited by 30 nM 2-methylthioadenosine-5' diphosphate (antagonist effect). Addition of the N(6)-methyl group in several cases converted pure agonists to antagonists. A carbocyclic N(6)-methyl-2' deoxyadenosine bisphosphate analogue was a pure P2Y(1) receptor antagonist and equipotent to the ribose analogue (MRS 2179). In the series of ring-constrained methanocarba derivatives where a fused cyclopropane moiety constrained the pseudosugar ring of the nucleoside to either a Northern (N) or Southern (S) conformation, as defined in the pseudorotational cycle, the 6-NH(2) (N)-analogue was a pure agonist of EC(50) 155 nM and 86-fold more potent than the corresponding (S)-isomer. The 2-chloro-N(6)-methyl-(N)-methanocarba analogue was an antagonist of IC(50) 51.6 nM. Thus, the ribose ring (N)-conformation appeared to be favored in recognition at P2Y(1) receptors. A cyclobutyl analogue was an antagonist with IC(50) of 805 nM, while morpholine ring-containing analogues were nearly inactive. Anhydrohexitol ring-modified bisphosphate derivatives displayed micromolar potency as agonists (6-NH(2)) or antagonists (N(6)-methyl). A molecular model of the energy-minimized structures of the potent antagonists suggested that the two phosphate groups may occupy common regions. The (N)- and (S)-methanocarba agonist analogues were docked into the putative binding site of the previously reported P2Y(1) receptor model. PMID- 10715152 TI - 5,6-Dihydropyran-2-ones possessing various sulfonyl functionalities: potent nonpeptidic inhibitors of HIV protease. AB - On the basis of previous SAR findings and molecular modeling studies, a series of compounds were synthesized which possessed various sulfonyl moieties substituted at the 4-position of the C-3 phenyl ring substituent of the dihydropyran-2-one ring system. The sulfonyl substituents were added in an attempt to fill the additional S(3)' pocket and thereby produce increasingly potent inhibitors of the target enzyme. Racemic and enantiomerically resolved varieties of selected compounds were synthesized. All analogues in the study displayed decent binding affinity to HIV protease, and several compounds were shown to possess very good antiviral efficacy and safety margins. X-ray crystallographic structures confirmed that the sulfonamide and sulfonate moieties were filling the S(3)' pocket of the enzyme. However, the additional substituent did not provide improved enzymatic inhibitory or antiviral activity as compared to the resolved unsubstituted aniline. The addition of the sulfonyl moiety substitution does not appear to provide favorable pharamacokinectic parameters. Selected inhibitors were tested for antiviral activity in clinical isolates and exhibited similar antiviral activity against all of the HIV-1 strains tested as they did against the wild-type HIV-1. In addition, the inhibitors exhibited good antiviral efficacies against HIV-1 strains that displayed resistance to the currently marketed protease inhibitors. PMID- 10715153 TI - 1,2-Dibenzamidobenzene inhibitors of human factor Xa. AB - High-throughput screening of a combinatorial library of diamidophenols yielded lead compounds with the ability to inhibit human factor Xa (fXa) at micromolar concentrations (e.g. compound 4, fXa apparent K(ass) = 0.64 x 10(6) L/mol). SAR studies in this novel structural series of fXa inhibitors showed that the phenolic hydroxyl group was not essential for activity. The best activity was found in substituted 1,2-dibenzamidobenzenes in which the phenyl group of one benzoyl group (A-ring) was substituted in the 4-position with relatively small lipophilic or polarizable groups such as methoxy, vinyl, or chloro and the phenyl group of the other benzoyl group (B-ring) was substituted in the 4-position with larger lipophilic groups such as tert-butyl or dimethylamino. The central phenyl ring (C-ring) tolerated a wide variety of substituents, but methoxy, methanesulfonamido, hydroxyl, and carboxyl substitution produced slightly higher levels of activity than other substituents when present in combination with favorable B-ring substitution. Methylation of the amide nitrogen atoms was found to greatly decrease activity. Compound 12 is the highest affinity fXa inhibitor in this group of compounds, having fXa apparent K(ass) = 25.5 x 10(6) L/mol, about 40x more active than the original lead. This lead series does not show potent inhibition of human thrombin. A model for the binding of these ligands to the fXa active site is proposed. The model is consistent with the observed SAR and can serve to guide future SAR studies. PMID- 10715154 TI - N(2)-Aroylanthranilamide inhibitors of human factor Xa. AB - Reversal of the A-ring amide link in 1,2-dibenzamidobenzene 1 (fXa K(ass) = 0.81 x 10(6) L/mol) led to a series of human factor Xa (hfXa) inhibitors based on N(2) aroylanthranilamide 4. Expansion of the SAR around 4 showed that only small planar substituents could be accommodated in the A-ring for binding to the S1 site of hfXa. Bulky groups such as 4-isopropyl, 4-tert-butyl, and 4-dimethylamino were favored in the B-ring to interact with the S4 site of hfXa. The central (C) ring containing a 5-methanesulfonamido group yielded greater activity than carbamoyl groups. Combining the beneficial features from the B- and C-ring SAR, compound 55 represents the most potent hfXa inhibitor in the N(2) aroylanthranilamide 4 series with hfXa K(ass) = 58 x 10(6) L/mol (K(i) = 11.5 nM). PMID- 10715156 TI - Selective ET(A) antagonists. 5. Discovery and structure-activity relationships of phenoxyphenylacetic acid derivatives. AB - The fifth paper in this series describes the culmination of our investigations into the development of a potent and selective ET(A) receptor antagonist for the treatment of diseases mediated by ET-1. Receptor site mapping of several ET(A) antagonists prepared previously identified a common cationic binding site which prompted synthesis of phenoxyphenylacetic acid derivative 13a, which showed good in vitro activity (IC(50) 59 nM, rat aortic ET(A)). Optimization of 13a led to the identification of 27b, which exhibited an IC(50) of 4 nM. Although this did not translate into the expected in vivo potency, a compound of comparable in vitro activity, 27a (RPR118031A), showed a far better pharmacokinetic profile and in vivo potency (75 micromol/kg) and was duly proposed and accepted as a development candidate. PMID- 10715155 TI - Structure-based design of potent, amidine-derived inhibitors of factor Xa: evaluation of selectivity, anticoagulant activity, and antithrombotic activity. AB - To enhance the potency of 1,2-dibenzamidobenzene-derived inhibitors of factor Xa (fXa), an amidine substituent was incorporated on one of the benzoyl side chains to interact with Asp189 in the S1 specificity pocket. Lead molecule 1 was docked into the active site of fXa to facilitate inhibitor design. Subsequently, iterative SAR studies and molecular modeling led to a 1000-fold increase in fXa affinity and a refined model of the new inhibitors in the fXa active site. Strong support for the computational model was achieved through the acquisition of an X ray crystal structure using thrombin as a surrogate protein. The amidines in this series show high levels of selectivity for the inhibition of fXa relative to other trypsin-like serine proteases. Furthermore, the fXa affinity of compounds in this series (K(ass) = 50-500 x 10(6) L/mol) translates effectively into both anticoagulant activity in vitro and antithrombotic activity in vivo. PMID- 10715157 TI - Inhibition of Grb2 SH2 domain binding by non-phosphate-containing ligands. 2. 4 (2-Malonyl)phenylalanine as a potent phosphotyrosyl mimetic. AB - Nonhydrolyzable phosphotyrosyl (pTyr) mimetics serve as important components of many competitive Grb2 SH2 domain inhibitors. To date, the most potent of these inhibitors have relied on phosphonate-based structures to replace the 4 phosphoryl group of the parent pTyr residue. Reported herein is the design and evaluation of a new pTyr mimetic, p-malonylphenylalanine (Pmf), which does not contain phosphorus yet, in Grb2 SH2 domain binding systems, approaches the potency of phosphonate-based pTyr mimetics. When incorporated into high affinity Grb2 SH2 domain-directed platforms, Pmf is 15-20 times more potent than the closely related previously reported pTyr mimetic, O-malonyltyrosine (OMT). Pmf containing inhibitors show inhibition constants as low as 8 nM in extracellular Grb2 binding assays and in whole cell systems, effective blockade of both endogenous Grb2 binding to cognate erbB-2, and downstream MAP kinase activation. Evidence is provided that use of an N(alpha)()-oxalyl auxiliary enhances effectiveness of Pmf and other inhibitors in both extracellular and intracellular contexts. As one of the most potent Grb2 SH2 domain-directed pTyr mimetics yet disclosed, Pmf may potentially have utility in the design of new chemotherapeutics for the treatment of various proliferative diseases, including breast cancer. PMID- 10715158 TI - Conformationally constrained analogues of diacylglycerol (DAG). 16. How much structural complexity is necessary for recognition and high binding affinity to protein kinase C? AB - The design of potent protein kinase C (PK-C) ligands with low nanomolar binding affinities was accomplished by the combined use of pharmacophore- and receptor guided approaches based on the structure of the physiological enzyme activator, diacylglycerol (DAG). Earlier use of the former approach, which was based on the structural equivalence of DAG and phorbol ester pharmacophores, identified a fixed template for the construction of a semirigid "recognition domain" that contained the three principal pharmacophores of DAG constrained into a lactone ring (DAG-lactones). In the present work, the pharmacophore-guided approach was refined to a higher level based on the X-ray structure of the C1b domain of PK Cdelta complexed with phorbol-13-O-acetate. A systematic search that involved modifying the DAG-lactone template with a combination of linear or branched acyl and alpha-alkylidene chains, which functioned as variable hydrophobic "affinity domains", helped identify compounds that optimized hydrophobic contacts with a group of conserved hydrophobic amino acids located on the top half of the C1 domain where the phorbol binds. The hydrophilic/hydrophobic balance of the molecules was estimated by the octanol/water partition coefficients (log P) calculated according to a fragment-based approach. The presence of branched alpha alkylidene or acyl chains was of critical importance to reach low nanomolar binding affinities for PK-C. These branched chains appear to facilitate important van der Waals contacts with hydrophobic segments of the protein and help promote the activation of PK-C through critical membrane interactions. Molecular modeling of these DAG-lactones into an empty C1b domain using the program AutoDock 2.4 suggests the existence of competing binding modes (sn-1 and sn-2) depending on which carbonyl is directly involved in binding to the protein. Inhibition of epidermal growth factor (EGF) binding, an indirect PK-C mediated response, was realized with some DAG-lactones at a dose 10-fold higher than with the standard phorbol-12, 13-dibutyrate (PDBU). Through the National Cancer Institute (NCI) 60 cell line in vitro screen, DAG-lactone 31 was identified as a very selective and potent antitumor agent. The NCI's computerized, pattern-recognition program COMPARE, which analyzes the degree of similarity of mean-graph profiles produced by the screen, corroborated our principles of drug design by matching the profile of compound 31 with that of the non-tumor-promoting antitumor phorbol ester, prostratin. The structural simplicity and the degree of potency achieved with some of the DAG-lactones described here should dispel the myth that chemical complexity and pharmacological activity go hand in hand. Even as a racemate, DAG lactone 31 showed low namomolar binding affinity for PK-C and displayed selective antitumor activity at equivalent nanomolar levels. Our present approach should facilitate the generation of multiple libraries of structurally similar DAG lactones to help exploit molecular diversity for PK-C and other high-affinity receptors for DAG and the phorbol esters. The success of this work suggests that substantially simpler, high-affinity structures could be identified to function as surrogates of other complex natural products. PMID- 10715159 TI - Design and synthesis of 13,14-dihydro prostaglandin F(1alpha) analogues as potent and selective ligands for the human FP receptor. AB - The in vitro evaluation of a new class of potential bone anabolic agents for the treatment of osteoporosis is described. These compounds are potent and selective ligands for the human prostaglandin F receptor (hFP receptor). The compounds lack the olefin unsaturation required for potency in the natural ligand PGF(2)(alpha) yet retain binding affinity for the hFP receptor in the nanomolar to micromolar range. Removal of the alkenes also results in a better selectivity ratio for the hFP receptor over the other prostaglandin receptors tested. A rationale for the selectivity differences of various analogues, based on ligand docking experiments to a putative hFP receptor model, is also described. PMID- 10715160 TI - Substituent effects on the antibacterial activity of nitrogen-carbon-linked (azolylphenyl)oxazolidinones with expanded activity against the fastidious gram negative organisms Haemophilus influenzae and Moraxella catarrhalis. AB - A series of new nitrogen-carbon-linked (azolylphenyl)oxazolidinone antibacterial agents has been prepared in an effort to expand the spectrum of activity of this class of antibiotics to include Gram-negative organisms. Pyrrole, pyrazole, imidazole, triazole, and tetrazole moieties have been used to replace the morpholine ring of linezolid (2). These changes resulted in the preparation of compounds with good activity against the fastidious Gram-negative organisms Haemophilus influenzae and Moraxella catarrhalis. The unsubstituted pyrrolyl analogue 3 and the 1H-1,2,3-triazolyl analogue 6 have MICs against H. influenzae = 4 microgram/mL and M. catarrhalis = 2 microgram/mL. Various substituents were also placed on the azole moieties in order to study their effects on antibacterial activity in vitro and in vivo. Interesting differences in activity were observed for many analogues that cannot be rationalized solely on the basis of sterics and position/number of nitrogen atoms in the azole ring. Differences in activity rely strongly on subtle changes in the electronic character of the overall azole systems. Aldehyde, aldoxime, and cyano azoles generally led to dramatic improvements in activity against both Gram-positive and Gram-negative bacteria relative to unsubstituted counterparts. However, amide, ester, amino, hydroxy, alkoxy, and alkyl substituents resulted in no improvement or a loss in antibacterial activity. The placement of a cyano moiety on the azole often generates analogues with interesting antibacterial activity in vitro and in vivo. In particular, the 3-cyanopyrrole, 4-cyanopyrazole, and 4-cyano-1H-1,2,3-triazole congeners 28, 50, and 90 had S. aureus MICs 1000 times more potent at the NR1A/2B subtype than at either the NR1A/2A or NR1A/2C subtypes. The binding affinities of 21 at alpha(1) adrenergic receptors ([(3)H]prazosin, IC(50) = 0.54 microM) and dopamine D2 receptors ([(3)H]raclopride, IC(50) = 1.2 microM) are reduced by incorporating a hydroxy group onto the 4-position of the piperidine ring and the beta-carbon of the N-alkyl spacer to give (+/-)-27: IC(50) NR1A/2B, 0.026; alpha(1), 14; D2, 105 microM. The high-potency phenolic antagonist 21 and its low-potency O-methylated analogue 18 are both potent anticonvulsants in a mouse maximal electroshock induced seizure (MES) study (ED(50) (iv) = 0.23 and 0.56 mg/kg, respectively). These data indicate that such compounds penetrate the blood-brain barrier but their MES activity may not be related to NMDA receptor antagonism. PMID- 10715163 TI - New azolidinediones as inhibitors of protein tyrosine phosphatase 1B with antihyperglycemic properties. AB - Insulin resistance in the liver and peripheral tissues together with a pancreatic cell defect are the common causes of type 2 diabetes. It is now appreciated that insulin resistance can result from a defect in the insulin receptor signaling system, at a site post binding of insulin to its receptor. Protein tyrosine phosphatases (PTPases) have been shown to be negative regulators of the insulin receptor. Inhibiton of PTPases may be an effective method in the treatment of type 2 diabetes. A series of azolidinediones has been prepared as protein tyrosine phosphatase 1B (PTP1B) inhibitors. Several compounds were potent inhibitors against the recombinant rat and human PTP1B enzymes with submicromolar IC(50) values. Elongated spacers between the azolidinedione moiety and the central aromatic portion of the molecule as well as hydrophobic groups at the vicinity of this aromatic region were very important to the inhibitory activity. Oxadiazolidinediones 87 and 88 and the corresponding acetic acid analogues 119 and 120 were the best h-PTP1B inhibitors with IC(50) values in the range of 0.12 0.3 microM. Several compounds normalized plasma glucose and insulin levels in the ob/ob and db/db diabetic mouse models. PMID- 10715164 TI - 2-Substituted tryptamines: agents with selectivity for 5-HT(6) serotonin receptors. AB - Several 2-alkyl-5-methoxytryptamine analogues were designed and prepared as potential 5-HT(6) serotonin agonists. It was found that 5-HT(6) receptors accommodate small alkyl substituents at the indole 2-position and that the resulting compounds can bind with affinities comparable to that of serotonin. In particular, 2-ethyl-5-methoxy-N, N-dimethyltryptamine (8) binds with high affinity at human 5-HT(6) receptors (K(i) = 16 nM) relative to 5-HT (K(i) = 75 nM) and was a full agonist, at least as potent (8: K(act) = 3.6 nM) as serotonin (K(act) = 5.0 nM), in activating adenylate cyclase. Compound 8 displays modest affinity for several other populations of 5-HT receptors, notably h5-HT(1A) (K(i) = 170 nM), h5-HT(1D) (K(i) = 290 nM), and h5-HT(7) (K(i) = 300 nM) receptors, but is otherwise quite selective. Compound 8 represents the first and most selective 5-HT(6) agonist reported to date. Replacing the 2-ethyl substituent with a phenyl group results in a compound that retains 5-HT(6) receptor affinity (i.e., 10: K(i) = 20 nM) but lacks agonist character. 2-Substituted tryptamines, then, might allow entry to a novel class of 5-HT(6) agonists and antagonists. PMID- 10715165 TI - Monocyclic L-nucleosides with type 1 cytokine-inducing activity. AB - A series of 1,2,4-triazole L-nucleosides were synthesized and evaluated for their ability to stimulate type 1 cytokine production by activated human T cells in direct comparison to the known active agent ribavirin. Among the compounds prepared, 1-beta-L-ribofuranosyl-1,2,4-triazole-3-carboxamide (5, ICN 17261) was found to be the most uniformly potent compound. Conversion of the 3-carboxamide group of 5 to a carboxamidine functionality resulted in 1-beta-L-ribofuranosyl 1,2,4-triazole-3-carboxamidine hydrochloride (10), which induced cytokine levels comparable to 5 for two of the three type 1 cytokines examined. Modification of the carbohydrate moiety of 5 provided compounds of reduced activity. Significantly, ICN 17261 offers interesting immunomodulatory potential for the treatment of diseases where type 1 cytokines play an important role. PMID- 10715166 TI - N-Terminal dipeptides of D(-)-penicillamine as sequestration agents for acetaldehyde. AB - Since acetaldehyde (AcH), a toxic oxidation product of ethanol, may play an etiologic role in the initiation of alcoholic liver disease, we had earlier pioneered the development of beta, beta-disubstituted-beta-mercapto-alpha-amino acids as AcH-sequestering agents. We now report the synthesis of a series of N terminal dipeptides of D(-)-penicillamine, prepared from the synthon 3-formyl 2,2,5,5-tetramethylthiazolidine-4S-carboxylic acid (3), a cyclized N-protected derivative of D(-)-penicillamine. These dipeptides were equally or more effective than penicillamine in trapping AcH in a cell-free system. In experiments using a hepatocyte culture system, two of the dipeptides, D-penicillamylglycine (6a) and D-penicillamyl-beta-alanine (6d), at 1/20 the molar concentration of ethanol, lowered the concentration of ethanol-derived AcH by 79% and 84%, respectively, at 2 h. The presence of cyanamide (an inhibitor of aldehyde dehydrogenase) in the incubation medium resulted in a 45-fold increase in ethanol-derived AcH; nevertheless, dipeptides 6a and 6c (D-penicillamyl-alpha-aminoisobutyric acid) were able to reduce this AcH level by approximately one-third. PMID- 10715167 TI - Novel 1,5-diphenylpyrazole nonnucleoside HIV-1 reverse transcriptase inhibitors with enhanced activity versus the delavirdine-resistant P236L mutant: lead identification and SAR of 3- and 4-substituted derivatives. AB - Through computationally directed broad screening, a novel 1, 5-diphenylpyrazole (DPP) class of HIV-1 nonnucleoside reverse transcriptase inhibitors (NNRTIs) has been discovered. Compound 2 (PNU-32945) was found to have good activity versus wild-type (IC(50) = 2.3 microM) and delavirdine-resistant P236L (IC(50) = 1.1 microM) reverse transcriptase (RT). Also, PNU-32945 has an ED(50) for inhibition of viral replication in cell cultures of 0.1 microM and was shown to be noncytotoxic with a CC(50) > 10 microM. Structure-activity relationship studies on the 3- and 4-positions of PNU-32945 led to interesting selectivity and activity within the class. In particular, the 3-hydroxyethyl-4-ethyl congener 29 is a potent inhibitor of the P236L mutant (IC(50) = 0.65 microM), whereas it is essentially inactive versus the wild-type enzyme (IC(50) > 50 microM). Furthermore, this compound was significantly more active versus the P236L mutant than delavirdine. The synthesis and RT inhibitory activity of various 3- and 4 substituted analogues are discussed. PMID- 10715168 TI - Alternative mechanisms of nonindependent mate choice. AB - Aspects of the environment, including the social environment, can contribute to intrapopulation variation in mating preferences. One example of the effect of social environment on mate preferences is mate choice copying; however, other types of socially influenced (nonindependent) choice might exist. We develop a list of such alternatives based on possible physiological or psychological mechanisms, evaluate the evidence distinguishing one from another and clarify some controversial aspects of mate choice copying. This framework reveals many ways in which one female's mate choice can influence that of another, and suggests a broader array of hypotheses about the selective forces acting on such mechanisms. Because nonindependent choice can occur in a variety of ways, it could be more important for understanding patterns of mate choice than current theory suggests. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715169 TI - An experimental analysis of territory size in juvenile steelhead trout. AB - I experimentally manipulated levels of food abundance and density of competitors to determine how these factors influence the territory size of juvenile steelhead trout, Oncorhynchus mykiss. Steelhead trout were held in artificial stream channels and I followed cohorts that were fed at one of three levels of food abundance and stocked at one of three levels of fish density. By measuring territory size over a 2-month period, while the fish were growing, I was also able to assess the effects of body size in determining the size of a territory. Defended and foraging areas were similar in absolute size, but the frequency of space use was different for defence than for foraging. As predicted, territory size decreased with increasing levels of food abundance and increased with decreasing levels of fish density. In addition, territory size increased with increasing body size even after controlling the effects of food abundance and competitor density. In comparison to previous studies, territory size of steelhead trout changed more dramatically in response to changing levels of food and competitors. For territorial animals with indeterminate growth, territory size is not only adjusted as a trade-off between the costs and benefits of defence, but also with respect to body size due to increasing metabolic demands as individuals grow. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715170 TI - Retention of social recognition after hibernation in Belding's ground squirrels. AB - The retention of social memory during long periods of separation, such as hibernation or migration, has not been well documented, despite evidence for long term social relationships in migrating species or in long-lived sedentary species. We investigated the ability of captive Belding's ground squirrels, Spermophilus beldingi, to remember previously familiar individuals as well as littermates after 9 months of isolation. Before hibernation, young ground squirrels discriminated between odours of familiar and unfamiliar individuals, as shown by greater investigation of a novel individual's odour. The following spring, these yearlings did not respond differentially to odours of previously familiar and unfamiliar individuals, suggesting that memory for familiar conspecifics was lost during hibernation. In contrast, both female and male yearlings continued to discriminate between odours of littermates and previously familiar nonlittermates. Thus, recognition of close kin was maintained during prolonged social isolation, but recognition of familiar, unrelated individuals was not. If re-establishment of familiarity is not costly or if adults rarely interact with the same individuals in successive years, then selection may not favour retention of individual memories of particular conspecifics over the winter. Even though males rarely encounter kin after dispersal, yearling males did recognize their siblings, suggesting that the relative costs of maintaining kin-recognition abilities year-round may be low. Possible mechanisms underlying the formation and maintenance of individual and kin recognition are discussed. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715171 TI - Agonistic screams differ among four species of macaques: the significance of motivation-structural rules. AB - We compared screams of four species of macaques (rhesus monkey, Macaca mulatta; pigtailed monkey, M. nemestrina; Sulawesi crested black macaque, M. nigra; stumptailed macaque, M. arctoides) with respect to predictions of Morton's motivation-structural rules (Morton 1977, American Naturalist, 111, 855-869). We examined screams produced by victims of attack that involved contact aggression (pulling, pushing, slapping, grappling and biting) from a higher-ranking opponent. For each macaque species, we digitized 100 screams from females 3 years of age or older and measured acoustic features of each call. We used discriminant function analysis to determine whether the 400 vocalizations could be assigned to the correct caller species on the basis of their acoustic structure. Calls were assigned to the correct species at a significantly higher rate (93.5%) than expected by chance (25%). Each of the four macaque species used acoustically distinct screams in a shared context. While the differences in the macaque species' vocalizations suggest no simple correlation between immediate context and the acoustic forms of screams, there was general correspondence between the acoustic structure predicted by motivation-structural rules and inferences about the internal state of the vocalizer derived from the typical intensity of aggressive patterns that characterize each of the four species. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715172 TI - Effects of residence time on displays during territory establishment in a lizard. AB - We know little about how signals are used during territory establishment, particularly when potential competitors are separated by distances that are typical of those between neighbours. I studied the effects of residence time on the display behaviour of male Anolis sagrei lizards in long- and short-distance contexts. In the long-distance context, the habitat patches of two male lizards were 5 m apart, separating the males by a distance typical of that in territorial neighbourhoods. For the short-distance context, I placed two males in one habitat patch. In both contexts, either 1-day residents were paired with new arrivals, or both individuals were new arrivals. In the long-distance contexts only, I also created situations in which both individuals were 1-day residents. Residence time affected the relative frequencies of headbob displays ('bobbing displays' and 'nodding displays'). However, the direction of the effect depended on opponent proximity. In long-distance contexts, 1-day residents performed fewer bobbing displays relative to nodding displays than did new arrivals; in short-distance contexts 1-day residents performed more bobbing displays relative to nodding displays than did new arrivals. The results suggest that signalling during territory establishment is governed by a qualitatively different set of rules when potential competitors are at short versus long distances. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715173 TI - Sex ratio determination by queens and workers in the ant Pheidole desertorum. AB - Because workers in colonies of eusocial Hymenoptera are more closely related to sisters than to brothers, theory predicts workers should bias investment in reproductive broods to favour reproductive females over males. However, conflict between queens and workers is predicted. Queens are equally related to daughters and sons, and should act to prevent workers from biasing investment. Previous study of the ant Pheidole desertorum showed that workers are nearly three times more closely related to reproductive females than males; however, the investment sex ratio is very near equal, consistent with substantial queen control of workers. Near-equal investment is produced by an equal frequency of colonies whose reproductive broods consist of only females (female specialists) and colonies whose reproductive broods consist of only males or whose sex ratios are extremely male biased (male specialists). Because natural selection should act on P. desertorum workers to bias investment in favour of reproductive females, why do workers in male-specialist colonies rear only (or mostly) males? We tested the hypothesis that queens prevent workers from rearing reproductive females by experimentally providing workers with immature reproductive broods of both sexes. Workers reared available reproductive females, while failing to rear available males. Worker preference for rearing reproductive females is consistent with queens preventing their occurrence in colonies of male specialists. These results provide evidence that queens and workers will act in opposition to determine the sex ratio, a fundamental prediction of queen-worker conflict theory. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715174 TI - Behavioural and hormonal responses to capture stress in the male red-sided garter snake, Thamnophis sirtalis parietalis. AB - We measured the behavioural and hormonal responses to capture stress in male red sided garter snakes. Four hours of capture stress resulted in no suppression of mating behaviour relative to control individuals. In contrast, the same stress resulted in a significant increase in plasma levels of corticosterone and a significant decrease in plasma levels of testosterone. There was a significant negative correlation between plasma levels of corticosterone and testosterone in both control and capture-stress groups, suggesting that the increase in corticosterone directly drives the decrease in testosterone. While there was no relation between body size and initial plasma levels of the two steroids, longer individuals had a significantly greater increase in corticosterone following capture stress than did shorter individuals. Snakes display indeterminate growth, suggesting that older individuals have decreased sensitivity to negative feedback in the hypothalamic-pituitary-adrenal axis and thus hypersecrete glucocorticoids. These results suggest that male red-sided garter snakes have uncoupled their behavioural stress response from their hormonal stress response to maximize reproductive opportunities. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715175 TI - Antipredator vigilance of juvenile and adult thirteen-lined ground squirrels and the role of nutritional need. AB - Juvenile thirteen-lined ground squirrels, Spermophilus tridecemlineatus, are less vigilant (i.e. they spend less time visually scanning the environment) than adults. To determine whether nutritional need was a potential cause of this difference, we supplemented two groups of free-ranging juveniles during the predispersal stage, while juveniles were still near and around the natal burrows. The high-energy food group (HEF: 11 squirrels) received peanut butter and oats while the low-energy food group (LEF: seven squirrels) received lettuce. Adults (14 squirrels) were also supplemented, but due to their greater home range sizes, it was not feasible to classify them as either HEF or LEF. To evaluate the effect of supplementation on antipredator vigilance, the behavioural act of visually scanning for predators, we videotaped individuals while they were foraging above ground during 5-min observation periods. Each squirrel was observed and weighed during three time periods over 23 days. From the videotape, we extracted measures of time spent vigilant, locomoting and foraging. All three categories of squirrels gained mass over the study period, but the HEF juveniles rapidly exceeded that of the LEF juveniles. Early in the study, LEF and HEF juveniles did not significantly differ in either body mass or time budgets, and, initially, both juvenile groups were similar to adults in the amount of time devoted to vigilance. Later in the study, the behaviour of HEF juveniles closely resembled that of adults (increased time devoted to vigilance and decreased time devoted to foraging), while LEF juveniles decreased vigilance and increased their foraging time. This study indicates that for thirteen-lined ground squirrels the lower vigilance of juveniles is due, at least in part, to the greater nutritional needs of young animals with consequent increases in foraging, which is largely incompatible with vigilance. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715176 TI - Parasites reduce attractiveness and reproductive success in male grain beetles. AB - Sexual characters may reveal the quality of a potential mate, including the mate's level of infection with parasites. Females that prefer males with low levels of infection or no infection may benefit in several ways. Direct benefits may include avoidance of infection, acquistition of larger nuptial gifts or enhancement in fecundity due to differences in male fertility. Females may also benefit indirectly by producing offspring that are more resistant to infections. We measured female preference for odours produced by male grain beetles, Tenebrio molitor, that were either infected by a tapeworm, Hymenolepis diminuta, or uninfected. This parasite is not transmitted directly between conspecifics. Females were attracted to odours of all males, but they were less attracted to those from parasitized males. To the contrary, females were preferentially attracted to infected females. Males did not show any biased attraction to odours from infected and uninfected male beetles. Females that mated with highly infected males produced fewer offspring than females mated to uninfected males, indicating parasitic infection inflicts multiple costs to males. These results are consistent with models of parasite-mediated sexual selection. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715177 TI - Female song sparrow, Melospiza melodia, response to simulated conspecific and heterospecific intrusion across three seasons. AB - To investigate female responses to territorial intrusion I presented female song sparrows with either a simulated female song sparrow intrusion or a simulated spotted towhee, Pipilo maculatus, intrusion as a control during either the prebreeding, breeding or postmoult seasons. Aggressive and nonaggressive behaviours and vocalizations were compared between intrusion types and across seasons. Principle components analysis suggested that female responses fell into three categories: (1) responses directed towards the intruder, mostly aggressive; (2) responses directed towards the mate; and (3) lack of response to the intruder. In every season, females responded more aggressively to simulated female song sparrow intrusion than simulated towhee intrusion. Responses directed towards female song sparrow intruders dropped across the three seasons and were significantly higher in the prebreeding season than in the breeding or postmoult seasons. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715178 TI - Individual and geographical variation in display behaviour of male harbour seals in Scotland. AB - Studying variations in behaviour at the individual or population level enables insight into the reproductive strategies within a species. We examined individual and geographical variation in the vocal and dive behaviour of male harbour seals, Phoca vitulina, which is associated with aquatic mating. This display behaviour was recorded in the Moray Firth, Scotland, from July 1994 to 1997, and in Orkney, Scotland, during July 1998. One vocalization type was apparent in the Moray Firth and two in Orkney. Time parameters (total and pulse duration) varied between males in the population in the Moray Firth. We used both frequency and time parameters in a discriminant analysis, which showed that 73.2% of individual male vocalizations could be correctly classified; 94.6% of male vocalizations from the Moray Firth and Orkney could be correctly classified according to their geographical areas. Therefore, vocal variation was greater between geographical areas than between individuals. No individual variation was apparent between dive and surface interval durations. However, individuals varied significantly in the percentage of short surface intervals. Male harbour seals showed substantial variability in the parameters affecting their vocal and dive behaviour during the mating season. We suggest that these variations may be indicative of adaptations to varying environmental challenges influencing the reproductive strategies of discrete populations. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715179 TI - Social tactics of pigs in a competitive foraging task: the 'informed forager' paradigm. AB - Studies of the social dynamics in foraging groups have focused primarily on birds, rodents and nonhuman primates. We extended the study of animal social tactics to the domestic pig, Sus scrofa, by using an experimental analogue of natural foraging skills, the 'informed forager' paradigm. We investigated the behaviour of 16 pigs foraging in pairs in an arena in which food had been hidden in one of eight monopolizable buckets. Before each pair trial, one of the pigs, the 'informed' pig, was given privileged knowledge about the location of the food during a solitary search trial. The 'noninformed' pig was naive about the location of the food during pair trials, but heavier than its informed partner and thus able to displace the latter from the baited bucket. By first focusing on the informed pigs' behaviour, we show that pigs are able to remember and relocate the food site. They found the food in relocation trials, using fewer bucket investigations than expected of a random searcher. Second, by focusing on the noninformed pigs, we show that pigs are able to exploit the knowledge of others by following them to a food source. They investigated more buckets immediately after their informed partners significantly more often than expected by chance and required fewer bucket investigations to find the food in pair trials than expected from a random searcher, but not in solitary search trials. We discuss these latter findings with reference to social foraging tactics. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715180 TI - The diving behaviour of green turtles at Ascension Island. AB - For six green turtles, Chelonia mydas, that had nested on Ascension Island in the South Atlantic, we used time-depth recorders to examine their diving behaviour during the subsequent internesting interval (10-12 days). All the turtles performed dives where they remained at a fixed depth for a long period, surfaced briefly and then dived to the same depth again. It is generally believed these dive profiles are caused by the turtles resting on the sea bed. The maximum depth that turtles routinely reached on these resting dives was between 18 and 20 m, with resting dives deeper than 20 m being extremely rare. Resting dive duration increased significantly with deeper dives. From this relationship, and assuming that turtles with fully inflated lungs at the surface need to dive to 19 m to achieve negative buoyancy, we estimated for two turtles that the oxygen consumption during resting dives was 0.016 and 0.020 litres O(2)/kg per h, respectively. This is similar to the value predicted from the allometric scaling relationship for the minimal oxygen consumption of turtles. We calculated that the energy conserved by resting during the internesting period may appreciably increase the reproductive output of females. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715181 TI - Leaf caching in Atta leafcutting ants: discrete cache formation through positive feedback. AB - We examined the occurrence, mechanism and costs and benefits of leaf caching in laboratory colonies of two species of leafcutting ants, Atta cephalotes and A. colombica. If foragers returning to the nest are unable to enter because of a temporary bottleneck caused by leaves building up they may deposit their leaf pieces outside the nest entrance, forming a leaf cache. Similar leaf caches occur in the field at foraging trail junctions, obstacles on the trail and within nest entrance tunnels. Foraging ants carrying leaves were presented with different sized leaf caches and the number dropping their leaves on the cache was recorded. The probability of a forager dropping her leaf was positively correlated with the size of the cache that she encountered. Therefore, positive feedback played a role in the formation of nest entrance caches. Cached pieces were more likely to be retrieved than noncached pieces but the time taken to retrieve leaf pieces from a cache was greater than from scattered groups of leaves. We suggest that the strategy of flexible nest entrance caching is an adaptive response to fluctuating food availability and collection. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715182 TI - Escape flights of yellowhammers and greenfinches: more than just physics. AB - Wintering birds increase their fat reserves throughout the day, and impaired escape performance is often considered to be an important cost of fat reserves. Since lifting a larger mass requires more energy, if birds escape at maximum power output, an increase in mass will impair the escape flight. In this study we did not find support for mass-dependent escape performance for yellowhammers, Emberiza citrinella, and greenfinches, Carduelis chloris, with natural daily mass increases of 7-8%. This suggests either that the birds were not performing at maximum output at dawn, when light, or that maximum power output was higher at dusk, when heavy. Either way, the birds seemed to be able to put more effort into their escape flight when heavier. In both species, when alarmed, birds took off significantly faster and at a steeper angle than when not alarmed. Yellowhammers escaped at a higher speed and angle than greenfinches, and reacted faster to the predator model. This suggests that predator escape is more than just Newtonian physics, and may be influenced by behavioural, as well as morphological, adjustments. Different species may have evolved different responses to predation risk. Our results seem to be in disagreement with recent ideas about mass dependent predation risk. However, to build up reserves, birds have to increase exposure time, which increases predation risk. This cost may be more important than impaired escape performance when relatively small, daily, changes in body mass are considered. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715183 TI - Web-building behaviour in the orb-weaving spider Zygiella x-notata: influence of experience. AB - Zygiella x-notata is an orb-weaving spider that often renews its trap daily. Web building has associated costs and benefits, and building successive webs may have consequences for lifetime reproductive success. In the laboratory, we tested the ability of Z. x-notata to modify its building behaviour in response to various stages in predation (prey detection, capture and ingestion) experienced with a previous web. We determined which stages provided information for the spiders. Spiders that detected, captured and ingested prey and then rebuilt their web used less silk and made a smaller capture area than in the previous web. There was no effect of prey detection alone on the next web. Capture without feeding gave the same results as capture followed by feeding. The spiders that ate prey without detection and capture (feeding by hand) had the same energetic gains as spiders that caught prey but delayed building a new web. The spiders thus showed plasticity in web-building behaviour and in the amount of silk used (energetic investment) in the short term (from one web to the next). Changes in body condition may therefore influence web construction. Moreover, information gained during prey capture appeared to influence the size and structure of the next web. This ability should enable spiders to adapt their web building to maximize their fitness. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715184 TI - Two intertidal fish species use visual association learning to track the status of food patches in a radial maze. AB - We tested fifteen-spined sticklebacks, Spinachia spinachia, and corkwing wrasse, Crenilabrus melops, for their ability to associate visual spatial cues with food sources in a radial maze and so to track renewal frequencies and productivity. When all locations contained food or were empty, subjects displayed win-shift or lose-shift behaviour by avoiding recently visited locations; this behaviour therefore appeared to be a basic trait. Both species readily learned food-cue associations, although with less efficiency as the diversity of cues increased. They used this information to distinguish food sources renewed within and between daily foraging bouts from those that remained empty. Moreover, both species distinguished between renewable food sources differing in productivity, preferentially visiting those containing more food. Reversal of cue roles caused an immediate decline in foraging efficiency, followed by rapid recovery as new food-cue associations were learned. Evidently, therefore, subjects tracked the status of potential food sources by continued sampling and could switch from win shift to win-stay behaviour, when preferred locations were persistently revisited, as appropriate. The formation of food-cue associations, together with sampling and flexible use of win-shift and win-stay behaviour, would enable these rocky intertidal fish species to exploit patchily distributed food sources whose status changes during the tidal cycle. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715185 TI - Sex-dependent risk taking in the collared flycatcher, Ficedula albicollis, when exposed to a predator at the nestling stage. AB - An increased mortality rate is a cost of parental care, and can be high during the provisioning phase of altricial nestlings. When a parent stops feeding the nestlings temporarily after seeing a predator, it can reduce its own predation risk, but the suspension of parental care may also reduce its offspring's chances of surviving. We modelled this situation by exposing a stuffed sparrowhawk near collared flycatcher nests and removing it when both parents had seen it. We measured the time (return time) between the removal and when each parent entered the nestbox. The parents' risk taking and the return time are assumed to be inversely related. We studied which brood variables the parents take into account when deciding how much risk they are willing to take during the provisioning period. Males took more risk for older and better-quality nestlings and earlier broods. The females' behaviour was opposite to that of the males: they took significantly less risk for older and better-quality offspring and visited the nestbox later for earlier broods. The males' behaviour supported the reproductive value hypothesis, that risk taking is related to brood value and survival chances, whereas the females' behaviour supported the harm to offspring hypothesis, that risk taking is related to the broods' vulnerability. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715186 TI - Kin-biased dispersal behaviour in the mango shield scale, Milviscutulus mangiferae. AB - When fitness decreases with increasing density in a habitat, dispersal behaviour is expected to evolve. To avoid competition between kin, dispersal behaviour based on kin recognition should be more likely to occur when the individuals in a habitat are closely related. I tested this prediction with first-instar larvae (crawlers) of the mango shield scale, Milviscutulus mangiferae. The body size of adult females, a measure of fecundity, was larger when only one female was present on a leaf than when two were present. When I placed two crawlers on a leaf, they emigrated more frequently when they were siblings than when they were not related. I discuss the implication of the results for kin recognition in thelytokous parthenogenetic animals. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715187 TI - Territorial intrusions and copulation patterns in red kites, Milvus milvus, in relation to breeding density. AB - Two main paternity assurance strategies are generally found in birds: mate guarding and frequent copulations. The latter is expected particularly in species such as raptors that cannot guard their mates efficiently because of ecological constraints, such as frequent courtship feeding. I investigated the prelaying behaviour of red kites, in which the males courtship feed. I compared pair behaviour in situations of varying breeding density and simulated male territorial intrusions by presenting decoys. Males' certainty of paternity was likely to decrease with increasing breeding density, because of the proximity of other males and more frequent male territorial intrusions during the presumed fertile period. The percentage of time spent by males within their breeding territory during the prelaying period was positively related to the number of close breeding neighbours, suggesting territory surveillance and mate guarding. The kites copulated frequently and over a long period. Copulation frequency prior to and during laying increased with breeding density, and in isolated pairs in response to simulated male territorial intrusions. The results support the idea of paternity assurance through frequent copulations during the presumed fertile period of the female, and suggest that early copulation activity is related to functions other than fertilization, such as pair bonding or mate assessment. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715188 TI - The role of food distribution and nutritional quality in behavioural phase change in the desert locust. AB - The behaviour of herbivorous insects is influenced by their nutritional state. Nutrition-induced behavioural changes are often interpreted as adaptive mechanisms for controlling nutrient intake; however, their influence on other life history traits has received far less attention. We investigated the effect of food quality and distribution on the behaviour and phase state of desert locusts, Schistocerca gregaria Forskal (Orthoptera, Acrididae), which change from the 'solitarious' to the 'gregarious' phase in response to population density. Phase change involves many morphological, physiological and behavioural changes. Solitarious insects are cryptic whereas gregarious locusts aggregate. Individual phase change is stimulated by mechanical contact with other locusts. A clumped resource distribution promotes change to the gregarious phase by increasing crowding and contact between individuals. In this study, we found that the effect of food distribution on locust phase depended on the nutritional quality of the food. We used three synthetic food treatments: near optimal, dilute and a choice of two unbalanced but complementary foods. Clumped resource distribution led to increased gregarization in the dilute and the complementary diet treatments. This effect was particularly pronounced on the complementary foods, owing to the interaction of crowding and locomotion. Gregarization was most pronounced in the dilute diet treatment, owing to increased activity. These diet-induced effects are explained in terms of behavioural changes in locomotion, quiescence and feeding that are consistent with what is known from earlier work on locust feeding behaviour and behavioural phase change. Copyright 2000 The Association for the Study of Animal Behaviour. PMID- 10715189 TI - Randomization tests and the equality of variance assumption when comparing group means. PMID- 10715190 TI - Body size enhances mating success in male garter snakes. PMID- 10715191 TI - Reply to Shine et al. (2000). PMID- 10715192 TI - Estimation of stature from the skeletal reconstruction of an immature Neandertal from Dederiyeh cave, Syria. AB - Skeletal reconstruction of a child Neandertal unearthed at Dederiyeh Cave, Syria in 1993, is undertaken and the acquired stature discussed. Although the skeletal remains were well preserved, the reconstruction required several assumptions to be made because of the immature status of the specimen. The assumptions were mainly concerned with distances between bones in the inter-vertebral spaces and in the joints of the hip, knee, and ankle. These were estimated from X-ray films of modern children and data from previous studies. Stature was directly measured on the reconstruction, and found to be 79.2 cm. After corrections for soft tissue thickness and shrinkage of the casts, the stature became 81.7 cm. This estimate is consistent with estimates based on regression equations of long bone lengths, especially from those of the lower extremity. In comparison with longitudinal data for white American boys, the assessment of stature for Dederiyeh varied according to the estimated age. For a younger estimated age, the stature falls in the lower half of the white American range of variation, but with an older estimated age, it falls below the lower limit of the range of variation. Other immature Neandertals including two European specimens, Roc de Marsal and La Ferrassie 6, fall below the lower limit of the 5th to 95th percentile range based on the estimated statures from their long bone lengths. More comprehensive age assessment covering both fossil and modern humans is required before accurate conclusions in relation to Neandertal growth can be drawn. PMID- 10715193 TI - The place of Neandertals in the evolution of hominid patterns of growth and development. AB - This study uses the two developmental fields of dental maturation and femoral growth to determine if the pattern of growth and development in Neandertals (archaic Homo sapiens) was intermediate between that of Homo erectus and recent modern humans. Specimens used in the analysis included Neandertals and Upper Palaeolithic early modern Homo sapiens from Europe and individuals from two recent modern human populations. Ontogenetic data for the H. erectus adolescent KNM-WT 15000 and for Gorilla gorilla were included for comparison. Previous reports have indicated that H. erectus demonstrates a pattern of ontogeny characterized by earlier and more rapid linear growth than in modern humans. Results reported here demonstrate that Upper Paleolithic early modern Homo sapiens display a growth trajectory indistinguishable from that of recent modern humans. The pattern of Neandertal ontogeny is not intermediate between the pattern displayed in H. erectus and the derived pattern seen in the modern reference samples and the early modern H. sapiens sample. The Neandertal growth trajectory is consistent with either slow linear growth or advanced dental development. PMID- 10715194 TI - Adults only. Reindeer hunting at the middle palaeolithic site salzgitter lebenstedt, northern Germany. AB - The Middle Palaeolithic site Salzgitter Lebenstedt (northern Germany), excavated in 1952, is well known because of its well-preserved faunal remains, dominated by adult reindeer (Rangifer tarandus). The archaeological assemblage accumulated in an arctic setting in an earlier part of the last (Weichsel) glacial (OIS5-3). The site is remarkable because of the presence of unique Middle Palaeolithic bone tools and the occurrence of the northernmost Neanderthal remains, but this paper focuses on an analysis of its reindeer assemblage. The results indicate autumn hunting of reindeer by Middle Palaeolithic hominids. After the hunt, carcasses were butchered and in subsequent marrow processing of the bones a selection against young and sub-adult animals occurred. Adults were clearly preferred, and from their bones, again, poorer marrow bones were neglected. This focus on primeness of resources has been documented in other domains of Neanderthal behaviour, but Salzgitter Lebenstedt is the best example yet known in terms of systematic and routinized processing of game. The Salzgitter Lebenstedt assemblage displays some remarkable similarities to the Late Glacial reindeer assemblages from the Ahrensburg tunnel valley sites. The subsequent review of the evidence on subsistence strategies from earlier periods of the European Palaeolithic shows that hunting of large mammals may have been a part of the behavioural repertoire of the Middle Pleistocene occupants of Europe from the earliest occupation onwards. At the same time, it is suggested that these early hunting strategies were incorporated in ways of moving through landscapes ("settlement systems") which were different from what we know from the middle parts of the Upper Palaeolithic onwards. PMID- 10715195 TI - The depositional context of the early upper paleolithic human fossils from the Koneprusy (Zlaty kun) and Mladec caves, Czech republic. AB - The caves of Mladec I and II (Moravia) and Koneprusy (Bohemia) are principal hominid Early Upper Paleolithic sites in Central Europe that require a complex reconsideration from several viewpoints. The focus of this paper is on the depositional context of human fossils, which is clearer from the documentation of Koneprusy, excavated during the 1950s, than from the early reports about Mladec. Both caves are multi-floor underground karstic systems penetrated by vertical fissures and chimneys, where the fossils were found in restricted areas, related to debris cones accumulated under the chimneys. These associations are confirmed using Surfer reconstruction of the original fillings. It appears certain for Koneprusy and highly probable for Mladec that the fossils fell in through the chimneys. This does not mean that living animals and humans never entered the interior of the caves (traces of gnawing by hyenas are visible), but it makes it unlikely that the human paleontological accumulations were the result of human activity within the cave chambers. PMID- 10715196 TI - The phylogenetic affinities of Otavipithecus namibiensis. AB - The middle Miocene hominoid Otavipithecus namibiensis is the first and most complete fossil ape from sub-equatorial Africa and represents a significant addition to the taxonomically sparse African middle Miocene hominoid fossil record. The Otavipithecus hypodigm comprises the holotype mandible, which presents a unique mosaic of dental and gnathic characters, and several attributed cranial and postcranial elements which resemble the stem hominoid Proconsul. Contrary to initial hopes that this discovery would provide new insights into hominoid morphological diversity and phylogenetic relationships, a variety of conflicting phylogenetic hypotheses have been advanced suggesting ties to virtually every major large-bodied hominoid group (Conroy et al., 1992; Andrews, 1992 a; Conroy, 1994; Pickford et al., 1994; Begun, 1994 a). Cladistic analysis of a matrix of 22 qualitative and ten quantitative characters of the mandible and mandibular dentition found no support for a close phylogenetic relationship between Otavipithecus and either the African ape or great ape clades, or with any of the Eurasian fossil hominoids with which it has previously been compared. A close relationship between Otavipithecus and Kenyapithecus cannot be ruled out, but is deemed unlikely on the basis both of morphological comparisons and the absence of support within a cladistic framework. The present analysis indicates that Otavipithecus is most closely related to Afropithecus, as previously suggested by Andrews (1992 a) among others. Due to lack of statistical support for this result, a conservative interpretation, that these taxa represented related but divergent lineages of a late early Miocene hominoid radiation, is currently favored. Findings are consistent with the allocation of Otavipithecus to Andrews' (1992 a) tribe Afropithecini which represents the sister group to Kenyapithecus and the extant ape clade. PMID- 10715198 TI - The smallest primates. PMID- 10715197 TI - Middle paleolithic human deciduous incisor from Khudji, Tajikistan. AB - In 1997 a human mandibular second deciduous incisor was discovered during excavations at the central Asian Middle Paleolithic site of Khudji, Tajikistan. The specimen was associated with a late Middle Paleolithic assemblage in a minimally disturbed cultural layer. The specimen is average in size for Late Pleistocene archaic human di(2)s and differs from many late archaic human di(2)s in having minimal marginal ridges and tapering markedly distally. In these features it resembles a minority of specimens from the later Pleistocene. PMID- 10715199 TI - Extraterrestrial evidence on the age of the hominids from Java. PMID- 10715200 TI - Structure and function of the hairpin ribozyme. AB - The hairpin ribozyme belongs to the family of small catalytic RNAs that cleave RNA substrates in a reversible reaction that generates 2',3'-cyclic phosphate and 5'-hydroxyl termini. The hairpin catalytic motif was discovered in the negative strand of the tobacco ringspot virus satellite RNA, where hairpin ribozyme mediated self-cleavage and ligation reactions participate in processing RNA replication intermediates. The self-cleaving hairpin, hammerhead, hepatitis delta and Neurospora VS RNAs each adopt unique structures and exploit distinct kinetic and catalytic mechanisms despite catalyzing the same chemical reactions. Mechanistic studies of hairpin ribozyme reactions provided early evidence that, like protein enzymes, RNA enzymes are able to exploit a variety of catalytic strategies. In contrast to the hammerhead and Tetrahymena ribozyme reactions, hairpin-mediated cleavage and ligation proceed through a catalytic mechanism that does not require direct coordination of metal cations to phosphate or water oxygens. The hairpin ribozyme is a better ligase than it is a nuclease while the hammerhead reaction favors cleavage over ligation of bound products by nearly 200 fold. Recent structure-function studies have begun to yield insights into the molecular bases of these unique features of the hairpin ribozyme. PMID- 10715201 TI - Structure and function of the mouse DNA methyltransferase gene: Dnmt1 shows a tripartite structure. AB - Dnmt1 is the predominant DNA methyltransferase (MTase) in mammals. The C-terminal domain of Dnmt1 clearly shares sequence similarity with many prokaryotic 5mC methyltransferases, and had been proposed to be sufficient for catalytic activity. We show here by deletion analysis that the C-terminal domain alone is not sufficient for methylating activity, but that a large part of the N-terminal domain is required in addition. Since this complex structure of Dnmt1 raises issues about its evolutionary origin, we have compared several eukaryotic MTases and have determined the genomic organization of the mouse Dnmt1 gene. The 5' most part of the N-terminal domain is dispensible for enzyme activity, includes the major nuclear import signal and comprises tissue-specific exons. Interestingly, the functional subdivision of Dnmt1 correlates well with the structure of the Dnmt1 gene in terms of intron/exon size distribution as well as sequence conservation. Our results, based on functional, structural and sequence comparison data, suggest that the gene has evolved from the fusion of at least three genes. PMID- 10715202 TI - Transposition and exon shuffling by group II intron RNA molecules in pieces. AB - In the realms of RNA, transposable elements created by self-inserting introns recombine novel combinations of exon sequences in the background of replicating molecules. Although intermolecular RNA recombination is a wide-spread phenomenon reported for a variety of RNA-containing viruses, direct evidence to support the theory that modern splicing systems, together with the exon-intron structure, have evolved from the ability of RNA to recombine, is lacking. Here, we used an in vitro deletion-complementation assay to demonstrate trans-activation of forward and reverse self-splicing of a fragmented derivative of the group II intron bI1 from yeast mitochondria. We provide direct evidence for the functional interchangeability of analogous but non-identical domain 1 RNA molecules of group II introns that result in trans-activation of intron transposition and RNA-based exon shuffling. The data extend theories on intron evolution and raise the intriguing possibility that naturally fragmented group III and spliceosomal introns themselves can create transposons, permitting rapid evolution of protein coding sequences by splicing reactions. PMID- 10715203 TI - Constructing high complexity synthetic libraries of long ORFs using in vitro selection. AB - We present a method that can significantly increase the complexity of protein libraries used for in vitro or in vivo protein selection experiments. Protein libraries are often encoded by chemically synthesized DNA, in which part of the open reading frame is randomized. There are, however, major obstacles associated with the chemical synthesis of long open reading frames, especially those containing random segments. Insertions and deletions that occur during chemical synthesis cause frameshifts, and stop codons in the random region will cause premature termination. These problems can together greatly reduce the number of full-length synthetic genes in the library. We describe a strategy in which smaller segments of the synthetic open reading frame are selected in vitro using mRNA display for the absence of frameshifts and stop codons. These smaller segments are then ligated together to form combinatorial libraries of long uninterrupted open reading frames. This process can increase the number of full length open reading frames in libraries by up to two orders of magnitude, resulting in protein libraries with complexities of greater than 10(13). We have used this methodology to generate three types of displayed protein library: a completely random sequence library, a library of concatemerized oligopeptide cassettes with a propensity for forming amphipathic alpha-helical or beta-strand structures, and a library based on one of the most common enzymatic scaffolds, the alpha/beta (TIM) barrel. PMID- 10715204 TI - DNA constraints on transcription activation in vitro. AB - Activators of eukaryotic transcription often function over a range of distances. It is commonly hypothesized that the intervening DNA between the transcription start site and the activator binding sites forms a loop in order to allow the activators to interact with the basal transcription apparatus, either directly or through mediators. If this hypothesis is correct, activation should be sensitive to the presence of intrinsic bends in the intervening DNA. Similarly, the precise helical phasing of such DNA bends and of the activator binding sites relative to the basal promoter should affect the degree of transcription activation. To explore these considerations, we designed transcription templates based on the adenovirus E4 promoter supplemented with upstream Gal4 activator binding sites. Surprisingly, we found that neither insertion of intrinsically curved DNA sequences between the activator binding sites and the basal promoter, nor alteration of the relative helical alignment of the activator binding sites and the basal promoter significantly affected in vitro transcription activation in HeLa cell nuclear extract. In all cases, the degree of transcription activation was a simple inverse function of the length of intervening DNA. Possible implications of these unexpected results are discussed. PMID- 10715205 TI - Inferring regulatory elements from a whole genome. An analysis of Helicobacter pylori sigma(80) family of promoter signals. AB - Helicobacter pylori is adapted to life in a unique niche, the gastric epithelium of primates. Its promoters may therefore be different from those of other bacteria. Here, we determine motifs possibly involved in the recognition of such promoter sequences by the RNA polymerase using a new motif identification method. An important feature of this method is that the motifs are sought with the least possible assumptions about what they may look like. The method starts by considering the whole genome of H. pylori and attempts to infer directly from it a description for a family of promoters. Thus, this approach differs from searching for such promoters with a previously established description. The two algorithms are based on the idea of inferring motifs by flexibly comparing words in the sequences with an external object, instead of between themselves. The first algorithm infers single motifs, the second a combination of two motifs separated from one another by strictly defined, sterically constrained distances. Besides independently finding motifs known to be present in other bacteria, such as the Shine-Dalgarno sequence and the TATA-box, this approach suggests the existence in H. pylori of a new, combined motif, TTAAGC, followed optimally 21 bp downstream by TATAAT. Between these two motifs, there is in some cases another, TTTTAA or, less frequently, a repetition of TTAAGC separated optimally from the TATA-box by 12 bp. The combined motif TTAAGCx(21+/-2)TATAAT is present with no errors immediately upstream from the only two copies of the ribosomal 23 S-5 S RNA genes in H. pylori, and with one error upstream from the only two copies of the ribosomal 16 S RNA genes. The operons of both ribosomal RNA molecules are strongly expressed, representing an encouraging sign of the pertinence of the motifs found by the algorithms. In 25 cases out of a possible 30, the combined motif is found with no more than three substitutions immediately upstream from ribosomal proteins, or operons containing a ribosomal protein. This is roughly the same frequency of occurrence as for TTGACAx(15-19)TATAAT (with the same maximum number of substitutions allowed) described as being the sigma(70 )promoter sequence consensus in Bacillus subtilis and Escherichia coli. The frequency of occurrence of the new motif obtained, TTAAGCx(19-23)TATAAT, remains high when all protein genes in H. pylori are considered, as is the case for the TTGACAx(15-19)TATAAT motif in B. subtilis but not in E. coli. PMID- 10715206 TI - Co-evolution of the tuf genes links gene conversion with the generation of chromosomal inversions. AB - The tufA and tufB genes in Salmonella typhimurium co-evolve by recombination and exchange of genetic material. A model is presented which predicts that co evolution is achieved by gene conversions and chromosomal inversions. Analysis of recombinants reveals that conversion and inversion each occur with similar rates and each depends on RecBCD activity. The model predicts sequence structures for different classes of post-recombination tuf genes. Sequence analysis reveals the presence of each of these structures and classes, with a predicted bias in the absence of mismatch repair. An implication of these data is that co-evolution of gene families can be linked with the generation of chromosomal rearrangements. PMID- 10715207 TI - Complete sequence of the mitochondrial genome of Tetrahymena pyriformis and comparison with Paramecium aurelia mitochondrial DNA. AB - We report the complete nucleotide sequence of the Tetrahymena pyriformis mitochondrial genome and a comparison of its gene content and organization with that of Paramecium aurelia mtDNA. T. pyriformis mtDNA is a linear molecule of 47,172 bp (78.7 % A+T) excluding telomeric sequences (identical tandem repeats of 31 bp at each end of the genome). In addition to genes encoding the previously described bipartite small and large subunit rRNAs, the T. pyriformis mitochondrial genome contains 21 protein-coding genes that are clearly homologous to genes of defined function in other mtDNAs, including one (yejR) that specifies a component of a cytochrome c biogenesis pathway. As well, T. pyriformis mtDNA contains 22 open reading frames of unknown function larger than 60 codons, potentially specifying proteins ranging in size from 74 to 1386 amino acid residues. A total of 13 of these open reading frames ("ciliate-specific") are found in P. aurelia mtDNA, whereas the remaining nine appear to be unique to T. pyriformis; however, of the latter, five are positionally equivalent and of similar size in the two ciliate mitochondrial genomes, suggesting they may also be homologous, even though this is not evident from sequence comparisons. Only eight tRNA genes encoding seven distinct tRNAs are found in T. pyriformis mtDNA, formally confirming a long-standing proposal that most T. pyriformis mitochondrial tRNAs are nucleus-encoded species imported from the cytosol. Atypical features of mitochondrial gene organization and expression in T. pyriformis mtDNA include split and rearranged large subunit rRNA genes, as well as a split nad1 gene (encoding subunit 1 of NADH dehydrogenase of respiratory complex I) whose two segments are located on and transcribed from opposite strands, as is also the case in P. aurelia. Gene content and arrangement are very similar in T. pyriformis and P. aurelia mtDNAs, the two differing by a limited number of duplication, inversion and rearrangement events. Phylogenetic analyses using concatenated sequences of several mtDNA-encoded proteins provide high bootstrap support for the monophyly of alveolates (ciliates, dinoflagellates and apicomplexans) and slime molds. PMID- 10715208 TI - Expression of mitochondrial protein-coding genes in Tetrahymena pyriformis. AB - In the ciliate protozoon, Tetrahymena pyriformis, mitochondrial protein-coding genes are highly divergent in sequence, and in a number of cases they lack AUG initiation codons. We asked whether RNA editing might be acting to generate protein sequences that are more conventional than those inferred from the corresponding gene sequences, and/or to create standard AUG initiation codons where these are absent. However, comparison of genomic and cDNA sequences (the latter generated by reverse transcriptase sequencing of T. pyriformis mitochondrial mRNAs) yielded no evidence of mitochondrial RNA editing in this organism. To delineate the 5' ends of mitochondrial protein-coding transcripts, primer extension experiments were conducted. In all cases, 5' termini were found to map within a few nucleotides of potential initiation codons, indicating that T. pyriformis mitochondrial mRNAs have little or no 5' untranslated leader sequence. The pattern of strong primer extension stops suggested that both standard (AUG) and non-standard (AUU, AUA, GUG, UUG) initiation codons are utilized by the Tetrahymena mitochondrial translation system. We also investigated expression of the nad1 gene, which in both T. pyriformis and Paramecium aurelia is split into two portions that are encoded by and transcribed from different DNA strands. Northern hybridization analysis showed that the corresponding transcripts are not trans-spliced, implying that separate N terminal and C-terminal portions of Nad1 are made in this system. Finally, in a search for primary transcripts, we isolated from a T. pyriformis mitochondrial fraction several small RNAs that were reproducibly labeled by incubation in the presence of [alpha-(32)P]GTP and guanylyltransferase. Partial sequence information revealed that none of these cappable RNAs is encoded in the T. pyriformis mitochondrial genome. PMID- 10715209 TI - Thioesterase domain of delta-(l-alpha-Aminoadipyl)-l-cysteinyl-d-valine synthetase: alteration of stereospecificity by site-directed mutagenesis. AB - The carboxy-terminal thioesterase domain of delta-(l-alpha-aminoadipyl)-l cysteinyl-d-valine synthetase catalyzes the hydrolytic release of the tripeptide product (LLD-ACV). By site-directed mutagenesis an S3599A change was introduced into the highly conserved GXSXG motif, resulting in a more than 95 % decrease of penicillin production. Purification of the modified multienzyme showed surprisingly only a 50 % reduction of the peptide formation rate, with the stereoisomer delta-(l-alpha-aminoadipyl)-l-cysteinyl-l-valine (LLL-ACV) as the dominating product. Thioesterases of ACV synthetases differ from other thioesterases integrated in non-ribosomal peptide synthetases in their direct association with an epimerase domain, and their respective GXSXG-seryl residue is apparently not essential in acyl transfer leading to peptide release. Instead, this motif may be involved in the control of tripeptide epimerization by selection of the isomer to be released, and the construct supports the presence of LLL-ACV as an intermediate in penicillin biosynthesis. PMID- 10715210 TI - The major surface antigens of Entamoeba histolytica trophozoites are GPI-anchored proteophosphoglycans. AB - Trophozoites of the parasitic protozoa, Entamoeba histolytica, synthesize a cell surface lipoglycoconjugate, termed lipophosphoglycan, which is thought to be an important virulence factor and potential vaccine candidate against invasive amebiasis. Here, we show that the E. histolytica lipophosphoglycans are in fact glycosylphosphatidylinositol (GPI)-anchored proteophosphoglycans (PPGs). These PPGs contain a highly acidic polypeptide component which is rich in Asp, Glu and phosphoserine residues. This polypeptide component is extensively modified with linear glycan chains having the general structure, [Glcalpha1-6](n)Glcbeta1-6Gal (where n=2-23). These glycan chains can be released after mild-acid hydrolysis with trifluoroacetic or hydrofluoric acid and are probably attached to phosphoserine residues in the polypeptide backbone. The PPGs are further modified with a GPI anchor which differs from all other eukaryotic GPI anchors so far characterized in containing a glycan core with the structure, Gal(1)Man(2)GlcN myo-inositol, and in being heterogeneously modified with chains of alpha galactose. Trophozoites of the pathogenic HM-1:IMSS strain synthesize two distinct classes of PPG which have polydisperse molecular masses of 50-180 kDa (PPG-1) and 35-60 kDa (PPG-2) and are modified with glucan side-chains of different average lengths. In contrast, the non-pathogenic Rahman strain synthesizes one class of PPG which is only elaborated with short disaccharide side-chains (i.e. Glcbeta1-6Gal). However, the PPGs are abundant in all strains (8x10(7) copies per cell) and are likely to form a protective surface coat. PMID- 10715211 TI - High resolution crystal structure of bovine mitochondrial EF-Tu in complex with GDP. AB - The crystal structure of bovine mitochondrial elongation factor Tu (EF-Tu) in complex with GDP has been determined at a resolution of 1. 94 A. The structure is similar to that of EF-Tu:GDP from Escherichia coli and Thermus aquaticus, but the orientation of the GDP-binding domain 1 is changed relative to domains 2 and 3. Sixteen conserved water molecules common to EF-Tu and other G-proteins in the GDP binding site are described. These water molecules create a network linking separated parts of the binding pocket. Mitochondrial EF-Tu binds nucleotides less tightly than prokaryotic EF-Tu possibly due to an increased mobility in regions close to the GDP-binding site. The C-terminal extension of mitochondrial EF-Tu has structural similarities with DNA recognising zinc fingers suggesting that the extension may be involved in recognition of RNA. PMID- 10715212 TI - Crystal structure of MEF2A core bound to DNA at 1.5 A resolution. AB - Members of the myocyte enhancer factor-2 (MEF2) family of transcription factors bind to and activate transcription through A+T-rich DNA sequences found primarily, but not exclusively, in the promoters of muscle-specific genes. Their importance has been established for myogenic development and in activation of the immediate-early gene, c-jun, and recently further functional roles in the immune system have emerged. The MEF2 factors belong to the MADS-box superfamily, sharing homology in a 58 amino acid domain that mediates DNA binding and dimerization. The structures of two MADS-box proteins, SRF and MCM1, bound to their cognate DNA have been previously reported and shown to share extensive similarity in their mode of DNA binding. We have solved the structure of MEF2A 2-78 bound to its DNA consensus sequence at 1.5 A resolution. It reveals how the absence of amino acids N-terminal to the MADS-box contributes to the DNA binding properties of MEF2 proteins and shows that the MEF domain C-terminal to the MADS-box adopts a conformation considerably different from the same region in SRF and MCM1. PMID- 10715213 TI - Crystal structure of a NifS-like protein from Thermotoga maritima: implications for iron sulphur cluster assembly. AB - NifS-like proteins are ubiquitous, homodimeric, proteins which belong to the alpha-family of pyridoxal-5'-phoshate dependent enzymes. They are proposed to donate elementary sulphur, generated from cysteine, via a cysteinepersulphide intermediate during iron sulphur cluster biosynthesis, an important albeit not well understood process. Here, we report on the crystal structure of a NifS-like protein from the hyperthermophilic bacterium Thermotoga maritima (tmNifS) at 2.0 A resolution. The tmNifS is structured into two domains, the larger bearing the pyridoxal-5'-phosphate-binding active site, the smaller hosting the active site cysteine in the middle of a highly flexible loop, 12 amino acid residues in length. Once charged with sulphur the loop could possibly deliver S(0) directly to regions far remote from the protein. Based on the three-dimensional structures of the native as well as the substrate complexed form and on spectrophotometric results, a mechanism of sulphur activation is proposed. The His99, which stacks on top of the pyridoxal-5'-phosphate co-factor, is assigned a crucial role during the catalytic cycle by acting as an acid-base catalyst and is believed to have a pK(a) value depending on the co-factor redox state. PMID- 10715214 TI - Structure of the utrophin actin-binding domain bound to F-actin reveals binding by an induced fit mechanism. AB - Utrophin is a large ubiquitously expressed cytoskeletal protein, homologous to dystrophin, the protein disrupted in Duchenne muscular dystrophy. The association of both proteins with the actin cytoskeleton is functionally important and is mediated by a domain at their N termini, conserved in members of the spectrin superfamily, including alpha-actinin, beta-spectrin and fimbrin. We present the structure of the actin-binding domain of utrophin in complex with F-actin, determined by cryo-electron microscopy and helical reconstruction, and a pseudo atomic model of the complex, generated by docking the crystal structures of the utrophin domain and F-actin into the reconstruction. In contrast to the model of actin binding proposed for fimbrin, the utrophin actin-binding domain appears to associate with actin in an extended conformation. This conformation places residues that are highly conserved in utrophin and other members of the spectrin superfamily at the utrophin interface with actin, confirming the likelihood of this binding orientation. This model emphasises the importance of protein flexibility in modeling interactions and presents the fascinating possibility of a diversity of actin-binding mechanisms among related proteins. PMID- 10715215 TI - The crystal structure of d-glyceraldehyde-3-phosphate dehydrogenase from the hyperthermophilic archaeon Methanothermus fervidus in the presence of NADP(+) at 2.1 A resolution. AB - The crystal structure of the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from the archaeon Methanothermus fervidus has been solved in the holo form at 2.1 A resolution by molecular replacement. Unlike bacterial and eukaryotic homologous enzymes which are strictly NAD(+)-dependent, GAPDH from this organism exhibits a dual-cofactor specificity, with a marked preference for NADP(+) over NAD(+). The present structure is the first archaeal GAPDH crystallized with NADP(+). GAPDH from M. fervidus adopts a homotetrameric quaternary structure which is topologically similar to that observed for its bacterial and eukaryotic counterparts. Within the cofactor-binding site, the positively charged side-chain of Lys33 decisively contributes to NADP(+) recognition through a tight electrostatic interaction with the adenosine 2'-phosphate group. Like other GAPDHs, GAPDH from archaeal sources binds the nicotinamide moiety of NADP(+) in a syn conformation with respect to the adjacent ribose and so belongs to the B stereospecific class of oxidoreductases. Stabilization of the syn conformation is principally achieved through hydrogen bonding of the carboxamide group with the side-chain of Asp171, a structural feature clearly different from what is observed in all presently known GAPDHs from bacteria and eukaryotes. Within the catalytic site, the reported crystal structure definitively confirms the essential role previously assigned to Cys140 by site-directed mutagenesis studies. In conjunction with new mutation results reported in this paper, inspection of the crystal structure gives reliable evidence for the direct implication of the side-chain of His219 in the catalytic mechanism. M. fervidus grows optimally at 84 degrees C with a maximal growth temperature of 97 degrees C. The paper includes a detailed comparison of the present structure with four other homologous enzymes extracted from mesophilic as well as thermophilic organisms. Among the various phenomena related to protein thermostabilization, reinforcement of electrostatic and hydrophobic interactions as well as a more efficient molecular packing appear to be essentially promoted by the occurrence of two additional alpha-helices in the archaeal GAPDHs. The first one, named alpha4, is located in the catalytic domain and participates in the enzyme architecture at the quaternary structural level. The second one, named alphaJ, occurs at the C terminus and contributes to the molecular packing within each monomer by filling a peripherical pocket in the tetrameric assembly. PMID- 10715216 TI - The spatial orientation of the essential amino acid residues arginine and aspartate within the alpha1beta1 integrin recognition site of collagen IV has been resolved using fluorescence resonance energy transfer. AB - The interaction of collagen IV with cells is mediated mainly by the integrin alpha1beta1. The recognition site has been located to a segment of the triple helical domain 100 nm away from the N terminus of the collagen molecule. The three essential amino acid residues of the alpha1beta1 binding site, arginine alpha2(IV)461 and the two aspartate residues alpha1(IV)461, are all located on different chains. Since the spatial array of the three residues depends on the stagger of the chains within the triple helix, the stagger has been elucidated using fluorescence resonance energy transfer with phenylalanine alpha1(IV)473 and tryptophan alpha2(IV)479 as the fluorescent donor/acceptor pair. The distance R between phenylalanine and tryptophan was determined by analysis of the energy transfer efficiency, E, and the orientation factor, kappa(2). In parallel, distance R and orientation factor, kappa(2 )were also calculated from the coordinates of the triple helix. Comparison of the calculated and empirically determined values unequivocally showed the stagger to be alpha1'alpha1alpha2. This arrangement of the three alpha chains describes the conformation of the alpha1beta1 integrin recognition site, that is the distinct orientation of the side-chains of the essential residues aspartate and arginine in respect to the helix axis. PMID- 10715217 TI - The kinetics of oligonucleotide replacements. AB - The formation of a duplex between two nucleic acid strands is restricted if one of the strands forms an intra- or intermolecular secondary structure. The formation of the new duplex requires the dissociation and replacement of the initial structure. To understand the mechanism of this type of kinetics we studied the replacement of a labeled DNA oligonucleotide probe bound to a complementary DNA target with an unlabeled probe of the same sequence. The replacement kinetics were measured using a gel-shift assay for 12, 14 and 16 nucleotide probes as a function of temperature and concentration of the unlabeled probe. The results demonstrate that the overall replacement rate is a combination of two kinetic pathways: dissociative and sequential displacement. The dissociative pathway occurs by the spontaneous dissociation of the initial duplex followed by association of the target and unlabeled probe. The sequential displacement pathway requires only the partial melting of the initial duplex to allow for the formation of a branched nucleation complex with the unlabeled probe, followed by the complete displacement of the labeled probe by migration of the branch point. The contribution from the dissociative pathway is predominant at temperatures close to the melting point of the labeled probe, whereas the contribution from the displacement pathway prevails at lower temperatures and when the concentration of the replacing unlabeled probe is high. The results show that at physiological conditions, duplex formation between a single-stranded oligonucleotide probe and a structured region of a target molecule occurs mainly by the sequential-displacement mechanism. PMID- 10715218 TI - Mechanism of DNA cleavage by the DNA/RNA-non-specific Anabaena sp. PCC 7120 endonuclease NucA and its inhibition by NuiA. AB - A structural model of the DNA/RNA non-specific endonuclease NucA from Anabaena sp. PCC7120 that has been obtained on the basis of the three-dimensional structure of the related Serratia nuclease, suggests that the overall architecture of the active site including amino acid residues H124, N155 and E163 (corresponding to H89, N119 and E127 in Serratia nuclease) is similar in both nucleases. Substitution of these residues by alanine leads to a large reduction in activity (<0.1 %), similarly as observed for Serratia nuclease demonstrating that both enzymes share a similar mechanism of catalysis with differences only in detail. NucA is inhibited by its specific polypeptide inhibitor with a K(i) value in the subpicomolar range, while the related Serratia nuclease at nanomolar concentrations is only inhibited at an approximately 1000-fold molar excess of NuiA. The artificial chromophoric substrate deoxythymidine 3',5'-bis-(p nitrophenyl phosphate) is cleaved by NucA as well as by Serratia nuclease. Cleavage of this analogue by NucA, however, is not inhibited by NuiA, suggesting that small molecules gain access to the active site of NucA in the enzyme inhibitor complex under conditions where cleavage of DNA substrates is completely inhibited. The active site residue E163 seems to be the main target amino acid for inhibition of NucA by NuiA, but R93, R122 and R167 (corresponding to K55, R87, R131 in Serratia nuclease) are also involved in the NucA/NuiA interaction. NuiA deletion mutants show that the structural integrity of the N and C-terminal region of the inhibitor is important for complex formation with NucA and inhibition of nuclease activity. Based on these results a mechanism of DNA cleavage by NucA and its inhibition by NuiA is proposed. PMID- 10715219 TI - Use of health hazard criteria for estimating the hazard potential of chemicals to water in case of a spill. AB - Accidental spills resulting in severe pollution can occur during transportation or handling of large volumes of chemicals. To address this problem, chemicals are classified according to the level of hazard to man and the environment in order to then define graduated technical standards. Three regulatory examples (enforced or drafted for transport and industrial installations in Europe) covering aspects of limnic as well as sea water are discussed in regard to health aspects of pollution. Whereas for the safety of seagoing tankships an exposure orientated combination of health and environmental aspects is used, for industrial plants in Germany a scoring system based on the European Union's Risk Phrase system is applied. The health-related parameters primarily used for hazard classification are repeated-dose toxicity and acute oral and dermal toxicity. Acute oral toxicity is most widely used because of the ready availability of data. Carcinogenicity is treated as the most important hazard. The report discusses the importance of dermal exposure, aspiration, and endocrine disruption as parameters as well as the importance of health criteria for the protection of aquatic organisms. PMID- 10715221 TI - Use of partition models in setting health guidelines for volatile organic compounds. AB - Partition models based on the octanol-air partition coefficients and associated quantitative structure-activity relationships (QSARs) have been developed to describe the triggering of odor detection, nasal irritation, and narcosis by common volatile organic compounds (VOCs). This study made use of the QSARs developed by Hau and Connell (1998), Indoor Air 8, 23-33) and Hau et al. (1999a, Toxicol. Sci. 47, 93-98; 1999b, Environ. Toxicol. Pharmacol. 7, 159-167) to predict the odor thresholds, nasal pungency thresholds, and anesthetic potency in humans for four groups of VOCs, namely, alkanes, alcohols, ketones, and acetates. The predicted outcomes with their estimated variability were used to evaluate the relevant guidelines on the airborne concentrations of these test groups. Threshold limit values (TLVs) for the test compounds were found to be generally capable of offering adequate protection against nasal pungency and narcosis, except for the higher alcohols (C6-C8) and sec-amyl acetate. The QSARs can also be used to set tentative guidelines for those compounds not having a TLV; values of 5 and 75 ppm were proposed for heptan-1-ol and dibutyl ketone respectively as examples. PMID- 10715220 TI - Dichlorvos and carcinogenicity: a systematic approach to a regulatory decision. AB - On the request of the Belgian Health Council, the authors performed a systematic review of the available evidence in the literature and in expert panel reports, with regard to a possible carcinogenic effect of dichlorvos. Following the evaluation procedure developed by IARC, they first concluded that dichlorvos should be classified as a possible carcinogen for man. This preliminary conclusion, and its possible consequences of withdrawal of several product authorizations, was then communicated by the Health Council to all stakeholders. As a result, the interpretation of the animal experimental data was confronted with the conclusions of a U.S. "Blue Ribbon Panel" of independent experts, who reviewed all the data that were available to them. After an exchange of views, the Health Council downgraded its classification of dichlorvos toward nonclassifiable with regard to cancer in man. This paper describes the review and decision-making processes, focusing on the major arguments underlying the original interpretation of the animal data and its eventual modification. PMID- 10715222 TI - Assessing the risks of exposures to multiple chemicals with a common mechanism of toxicity: how to cumulate? AB - The Food Quality Protection Act (FQPA) of 1996 requires the U.S. EPA to consider the "cumulative effects" of pesticides and other substances that have a "common mechanism of toxicity." Several different methods for combining the exposures to estimate the risk of groups of common mechanism chemicals with different potencies and exposure characteristics are critically evaluated. These are the hazard index (HI), toxicity equivalence factor (TEF), and combined margin of exposure (MOE(T)) procedures as well as the point of departure index (PODI) and cumulative risk index (CRI) methods that are the reciprocals of the HI and MOE(T) approaches, respectively. Each of these methods ideally requires, at a minimum, the availability of in vivo toxicology data for the same toxicological endpoint in the same animal species. Furthermore, all assume that the effects of the individual components in the mixture are independent in nature (i.e., are additive rather than synergistic or antagonistic) and that the dose-response functions for all compounds have a similar slope. The point of departure (POD), preferably the dose corresponding to a given effect level (e.g., the ED(10)), can be used as a measure of the relative potency of the different chemicals in the group. If appropriate exposure and toxicology data are available, and the chemicals in the group have a common uncertainty factor (UF), all the procedures yield a numerically identical result. The fact that different chemicals in the group often have different UFs raises issues for all summation procedures and, in the case of the TEF approach, the UF of the index chemical selected dictates the final result of the assessment. A major distinction between the different methods for addition is the point in the process at which uncertainty is considered. The HI and CRI approaches are problematic because they require application of policy driven UFs (in the form of RfDs) at that stage of the process where exposure should be expressed in terms of potency. In contrast, the PODI and MOE(T) approaches require application of a single group UF(G) at the end of the risk assessment process although they will also accommodate the application of data based adjustments earlier in the analysis. Importantly, both the PODI and the MOE(T) approaches allow policy- and data-driven UFs to be separated and thus make the process more transparent; these should be considered the methods of choice for cumulative risk assessment. Assignment of a single group UF is somewhat different from developing an UF for a single chemical and the total weight of evidence available in the group database can be used to advantage to reduce the UFs that need to be applied to the group. This larger database can also be used to refine the PODs for individual members of the group. It is important to emphasize that there remains a great deal of scientific uncertainty about how to proceed with cumulative risk assessment as described in the FQPA. The serious difficulties associated with defining "common mechanism of toxicity" and "concurrent exposure" combined with the current paucity of data and methodology required to conduct cumulative risk assessment suggest that the procedure is not yet ready for use in pesticide regulation. PMID- 10715223 TI - Reducing uncertainty of risk estimates for mixtures of chemicals within regulatory constraints. AB - Reducing uncertainty in estimated risks is always desirable. While regulatory assumptions and policies regarding risk assessment of chemicals can be debated, these are the rules under which many risk assessments are currently conducted. Methods for reducing the uncertainty in risk estimates generated under those rules are therefore useful, whether or not one agrees with the models and the underlying assumptions that comprise those rules. The guidance for risk assessment of mixtures of chemicals used by EPA was reexamined to determine methods for reducing the uncertainty in the cumulative risk estimates. It was found that the uncertainty could be significantly reduced if the assumptions concerning the assessment of mixtures of chemicals were combined with the assumptions for evaluating the risks of individual chemicals. Methods are proposed for reducing the uncertainty of mixtures of chemicals whose individual constituents are evaluated by cancer potency factors, hazard quotients, or margins of exposure. The methods developed do not require data beyond that which would be required for generating the current risk estimates for the individual chemicals. These analyses also demonstrated that some of the assumptions currently in use for regulatory risk assessment may lead to inconsistencies that should be reevaluated. As the proposed methods do not require any change in the current assumptions to reduce uncertainty in the risk estimates, the proposed methods should prove useful until such time as the assumptions are reevaluated and possibly changed. PMID- 10715224 TI - Conservatism in pesticide exposure assessment. AB - Three important factors are commonly encountered in exposure assessment that when combined could overestimate the exposure to pesticides by as much as two orders of magnitude. The three factors discussed are dermal absorption from laboratory animal studies, daily dose extrapolated from partial day monitoring, and nonbolus dose from dermal or inhalation exposure. Conservatism built into the process by each of these three factors is substantiated with available empirical data. The dose overestimation from these factors varies discriminatively by exposure scenarios and peculiarities of a given chemical. It is for this reason that a generic overestimation factor cannot be ascribed. Following the empirical illustrations, the authors conclude that the most effective approach for dealing with the problem is to generate the most appropriate data possible. This means producing human rather than laboratory animal dermal absorption data, conducting full-day exposure monitoring studies, and whenever feasible generating dermal rather than oral toxicology data (or alternatively data on both oral and dermal pharmacokinetics) in those cases where the dermal route predominates. PMID- 10715225 TI - Effects of olestra and sorbitol consumption on objective measures of diarrhea: impact of stool viscosity on common gastrointestinal symptoms. AB - The aim of this study was to determine the effects of olestra and sorbitol consumption on three accepted objective measures of diarrhea (stool output >250 g/day, liquid/watery stools, bowel movement frequency >3/day), and how stool composition influences reports of common gastrointestinal symptoms. A double blind, placebo-controlled study compared the effects of sorbitol (40 g/day in candy), a poorly absorbed sugar-alcohol with known osmotic effects, with those of olestra (20 or 40 g/day in potato chips), a nonabsorbed fat, on objective measures of stool composition and GI symptoms. Sixty-six subjects resided on a metabolic ward for 12 days: 2 days lead-in, 4 days baseline, 6 days treatment. Sorbitol 40 g/day resulted in loose/liquid stools within 1-3 h of consumption. In contrast, olestra resulted in a dose-responsive stool softening effect after 2-4 days of consumption. Subjects reported "diarrhea" when mean stool apparent viscosity (peak force (PF), g) decreased from a perceived "normal" (mean +/- SE, 1355 +/- 224 g PF; firm stool) to loose (260 +/- 68 g PF) stool. Mean apparent viscosity of stool during treatment: placebo, 1363 +/- 280 g (firm); olestra 20 g/day 743 +/- 65 g (soft); olestra 40 g/day, 563 +/- 105 g (soft); and sorbitol 40 g/day, 249 +/- 53 g (loose). Of the 1098 stool samples collected, 38% (419/1098) were rated by subjects as "diarrhea," yet only 2% of treatment days (all in the sorbitol treatment group) met commonly accepted criteria for a clinical diarrhea. Sorbitol, but not olestra, increased the severity of abdominal cramping, urgency and nausea compared to placebo. Olestra consumption, at levels far in excess of normal snacking conditions, resulted in a gradual stool softening effect after several days of consumption, did not meet any of the three objective measures of diarrhea, and did not increase GI symptoms. Sorbitol consumption, at only 80% of the dose requiring a "laxative effect" information label, resulted in rapid onset loose/liquid stools and a significant increase in abdominal cramping, urgency and nausea. Overall, subjects categorized stool as "diarrhea" when stool decreased from their perceived "normal," but the vast majority of these reports were not associated with clinically significant diarrhea. PMID- 10715226 TI - Dietary supplements and lessons to be learned from GRAS. AB - The demand for dietary supplements by the public has transformed this once cottage industry into a 12-billion-dollar-per-year business. Restrictive actions against dietary supplements by the Food and Drug Administration (FDA) prompted Congress to enact new and more permissive amendments governing dietary supplements (Dietary Supplement Health and Education Act, DSHEA) to the Federal Food, Drug and Cosmetic (FFD&C) Act in 1994. A comparison is made between the present status of dietary supplement regulation and the concept of general recognition of safety (GRAS) under conditions of intended use as set forth by the landmark 1958 Food Additive Amendment to the FFD&C Act. An argument is posited for use of applicable principles learned in nearly 40 years of experience with determining the GRAS status for hundreds of substances to problems posed by dietary supplements. PMID- 10715227 TI - An analysis of the possibility for health implications of joint actions and interactions between food additives. AB - The possibility that structurally unrelated food additives could show either joint actions or interactions has been assessed based on their potential to share common sites and mechanisms of action or common pathways of elimination. All food additives approved in the European Union and allocated numerical acceptable daily intake values were studied, initially based on the reports by the FAO-WHO Joint Expert Committee for Food Additives. Target organs were identified based on the effects reported at doses above the no-observed-adverse-effect level (NOAEL) in animal and human studies. The descriptions of the pathological and other changes reported were used to assess whether different additives, sharing the same target organ, would produce a common toxic effect. In all but a very few cases, the possibility of joint actions or interactions could be excluded on scientific grounds. The exceptions were on the liver (curcumin, thiabendazole, propyl gallate, and BHT), the kidney (diphenyl, o-phenylphenol, and ferrocyanide salts), the blood (azorubine and propyl gallate), and the thyroid (erythosine, thiabendazole, and nitrate). Toxicokinetic interactions were considered unlikely because of the low dosages involved, the diverse nature of the routes of metabolism and elimination, and the fact that enzyme induction or inhibition would have influenced selection of the NOAEL. Many of those additives which could not be excluded from showing joint actions or interactions would have low intakes; in some cases they were alternatives for the same application, thereby further lowering the combined intake. In consequence, joint actions or interactions between additives do not represent a significant health concern. PMID- 10715228 TI - Food regulation: use of science-based decisions to determine appropriate levels of protection. PMID- 10715229 TI - The National Academy of Sciences offers a new framework for addressing global warming issues. AB - The recent landmark report by the National Academy of Sciences reviewed the science on which the Kyoto Protocol was based. NAS concluded that the policy choices and the mandatory reductions in greenhouse gases by the developed nations were based on incomplete science with significant uncertainties. In view of these uncertainties the NAS report developed a comprehensive strategic 10-year research program to address the basic issue of whether human activity that results in environmental changes is responsible for climate changes. The report provides a new framework for consideration of global warming issues. The UN International Panel on Climate Change (the UN science advisor) in its 1997 report to the Kyoto parties pointed out the confusing difference between scientific usage of the term "climate change" that distinguishes human from natural causes of change and the official usage that combines natural and human causes of changes in climate. The conclusion of the UN panel on human causes is equivocal. The 1999 report of the U.S. Global Science Research Committee also reached an equivocal conclusion on human causes and announced a 10-year research program to be developed in consultation with NAS. The precautionary measures provided in the 1992 UN Framework Convention differ from the ill-defined "precautionary principle" based on fear of uncertainty, and are consistent with the objectives of the NAS proposed research program. These developments together with the third report of the UN Intergovernmental Science Panel on developments in climate science due in 2001 merit consideration by the convention of the parties under the Kyoto Protocol. PMID- 10715230 TI - First Results from Viper: Detection of Small-scale Anisotropy at 40 GHz. AB - Results of a search for small-scale anisotropy in the cosmic microwave background (CMB) are presented. Observations were made at the South Pole using the Viper telescope, with a 0&fdg;26 (FWHM) beam and a passband centered at 40 GHz. Anisotropy band-power measurements in bands spanning the range of l in which the first acoustic peak is expected (bands centered at l=108, 173, 237, 263, 422, and 589) are reported. Statistically significant CMB anisotropy is detected in all bands. PMID- 10715231 TI - Can a Changing alpha Explain the Supernovae Results? AB - We show that the supernovae results, which imply that there is evidence for an accelerating universe, may be closely related to the recent discovery of redshift dependence in the fine-structure constant alpha. The link is a class of varying speed-of-light (VSL) theories that contain cosmological solutions that are similar to quintessence. During the radiation-dominated epoch, the cosmological constant Lambda is prevented from dominating the universe by the usual VSL mechanism. In the matter-dominated epoch, the varying-c effects switch off, allowing Lambda to eventually surface and lead to an accelerating universe. By the time this happens, the residual variations in c imply a changing alpha at a rate that is in agreement with observations. PMID- 10715233 TI - The Extreme Compact Starburst in Markarian 273. AB - Images of neutral hydrogen 21 cm absorption and radio continuum emission at 1.4 GHz from Mrk 273 were made using the Very Long Baseline Array and Very Large Array. These images reveal a gas disk associated with the northern nuclear region with a diameter 0&farcs;5 (370 pc) at an inclination angle of 53 degrees. The radio continuum emission is composed of a diffuse component plus a number of compact sources. This morphology resembles those of nearby, lower luminosity starburst galaxies. These images provide strong support for the hypothesis that the luminosity of the northern source is dominated by an extreme compact starburst. The H i 21 cm absorption shows an east-west gradient in velocity of 450 km s-1 across 0&farcs;3 (220 pc), which implies an enclosed mass of 2x109 M middle dot in circle, comparable to the molecular gas mass. The brightest of the compact sources may indicate radio emission from an active nucleus, but this source contributes only 3.8% to the total flux density of the northern nuclear regions. The H i 21 cm absorption toward the southeast radio nucleus suggests infall at 200 km s-1 on scales /=0.55 and beta2) solar wind ions and interstellar neutrals. The high charge state solar wind ions resulting from these collisions are left in highly excited states and emit extreme ultraviolet or soft X-ray photons. This solar wind charge exchange mechanism applied to cometary neutrals has been used to explain the soft X-ray emission observed from comets. A simple model demonstrates that heliospheric X-ray emission can account for about 25%-50% of the observed soft X-ray background intensities. The spatial and temporal variations of heliospheric X-ray emission should reflect variations in the solar wind flux and composition as well as variations in the distribution of interstellar neutrals within the heliosphere. The heliospheric X-ray "background" can perhaps be identified with the "long-term enhancements" in the soft X-ray background measured by ROSAT. PMID- 10715248 TI - Erratum. PMID- 10715249 TI - Circulation online only : march 14, 2000 PMID- 10715250 TI - Peak VO(2): a simple yet enduring standard. PMID- 10715251 TI - Low molecular weight heparin after mechanical heart valve replacement. AB - BACKGROUND: Patients with mechanical heart valves require life-long anticoagulation. We report here the first large and comparative series of consecutive patients anticoagulated with low molecular weight heparin (LMWH) after mechanical heart valve replacement. METHODS AND RESULTS: In this comparative, nonrandomized study, 208 consecutive patients who underwent a single or double heart valve replacement with mechanical prostheses were anticoagulated subcutaneously with unfractionated heparin (UH) in the first period (n=106) and LMWH in the second phase (n=102) of the study. Baseline characteristics were similar in the 2 groups. The mean durations of UH and LMWH treatments were 13.6+/ 0.5 and 14.1+/-0.6 days, respectively (not significant). On the second day of treatment, 87% of patients treated with LMWH had an anti-Xa activity within the range of efficacy (0.5 to 1 IU/mL), but only 9% of UH-treated patients had an activated partial-thromboplastin time value within the therapeutic range (1.5 to 2.5 times control, P<0.0001 between the 2 groups). On the last day of prescription, all LMWH-treated patients had anti-Xa activity above 0.5 IU/mL, but 19% were above 1 IU/mL. In the UH group, 27% of patients had an activated partial thromboplastin time above 1.5 times control, but 62% were overanticoagulated. Two major bleedings occurred in each group, and one stroke occurred in the UH group. CONCLUSIONS: In this first comparative study, anticoagulation with LMWHs after mechanical heart valve replacement appears feasible, provides adequate biological anticoagulation, and compares favorably with UH anticoagulation. Randomized studies are now needed to further evaluate this new therapeutic approach. PMID- 10715252 TI - The effect of endovascular irradiation on platelet recruitment at sites of balloon angioplasty in pig coronary arteries. AB - BACKGROUND: Endovascular irradiation (EI) inhibits balloon-induced neointima formation in animals and is now in clinical trials for restenosis prevention. However, little is known of the effect of EI on vessel thrombogenicity due to delayed arterial healing. We investigated EI effects on platelet recruitment in pig coronary arteries. METHODS AND RESULTS: EI was performed using (90)Sr/Y at 0 Gray (Gy), 15Gy, or 30Gy at 2 mm after balloon overstretch injury. At 1 day, 1 week, and 1 month, platelet recruitment and thrombus formation were assessed using autologous (111)In-oxine-platelet labeling and light and scanning electron microscopy. In balloon-injured nonirradiated vessels, there was complete reendothelialization at 1 month, and platelet recruitment was similar to normal uninjured arteries. In irradiated vessels, scanning electron microscopy showed incomplete reendothelialization at 1 month, and these areas demonstrated attachment of activated platelets. Light microscopy of irradiated coronaries showed adherent partially organized thrombi and incomplete resolution of intramural hemorrhages. There was a significant increase in platelet recruitment at 1 month in arteries receiving EI at 15Gy (5.1+/-2. 8x10(6), P=0.02) or 30Gy (12.5+/-9.9x10(6), P=0.005) compared with nonirradiated controls (2.7+/ 1.5x10(6)); 30Gy was also higher than 15Gy (P=0.05). Platelet recruitment was also increased for 30Gy compared with control at 1 day. CONCLUSIONS: Endovascular irradiation at 15Gy or 30Gy after balloon angioplasty results in incomplete endothelial recovery, impaired resolution of intramural hemorrhage, and a dose dependent increase in platelet recruitment at 1 month. PMID- 10715253 TI - Adenovirus-mediated gene transfer of a secreted form of human macrophage scavenger receptor inhibits modified low-density lipoprotein degradation and foam cell formation in macrophages. AB - BACKGROUND: Macrophage scavenger receptors (MSRs) play an important role in the pathogenesis of atherosclerosis. Therefore, local modulation of MSR activity could have a beneficial effect on atherogenesis. METHODS AND RESULTS: We cloned a secreted "decoy" MSR (sMSR) that contains an extracellular portion of the human MSR type AI and constructed an adenoviral vector that directs high-level expression of sMSR in macrophages under the control of the human CD68 promoter. Expression of the sMSR protein inhibited the degradation of (125)I-labeled acetylated LDL and oxidized LDL by murine macrophages up to 90%. sMSRs also reduced acetylated LDL degradation in MSR knockout mouse peritoneal macrophages by 60% to 80%, which suggests that the decoy construct can compete for the uptake mediated via other related scavenger receptors. In addition, sMSRs inhibited foam cell formation in murine macrophages in the presence of cytochalasin D. The mechanism of inhibition is through ligand binding to the sMSRs, which prevents the ligand binding to MSRs on cell membranes. CONCLUSIONS: The demonstration that recombinant adenovirus-mediated gene transfer of decoy sMSRs can block foam-cell formation suggests a possible new strategy for gene therapy of atherosclerosis and for the treatment of lipid accumulation after arterial manipulations. PMID- 10715254 TI - Aspirin use is low among United States outpatients with coronary artery disease. AB - BACKGROUND: The goal of the present study was to assess national trends and patterns of aspirin use among outpatients with coronary artery disease. Although there is strong evidence that the use of aspirin reduces the risk of death and recurrent events in patients with coronary artery disease, current national patterns of aspirin use are unknown. METHODS AND RESULTS: We used data from the 1980 to 1996 National Ambulatory Medical Care Surveys. These surveys provide a nationally representative sample of physician activities during patient visits to physician offices. We evaluated the report of aspirin as a new or continuing medication in 10 942 visits to cardiologists and primary care physicians by patients with coronary artery disease. We evaluated trends in the use of aspirin for 1980 to 1996 and used logistic regression to identify independent predictors of aspirin use for 1993 to 1996. Aspirin use in outpatient visits by persons with coronary artery disease without reported contraindications increased from 5.0% in 1980 to 26.2% in 1996. Large increases occurred in the early 1990s. Independent predictors of aspirin use in 1993 to 1996 were male patient gender (29% versus 21% for females), patient age of <80 years (28% versus 17% for age of >/=80 years), and presence of hyperlipidemia (45% versus 24% for patients without hyperlipidemia; all comparisons P<0. 001). Cardiologists (37%) were more likely to report aspirin use than were internists (20%), family physicians (18%), or general practitioners (11%; P<0.001). These effects persisted after we controlled for potential confounders with the use of logistic regression. CONCLUSIONS: Although aspirin use in patients with coronary artery disease has increased dramatically, it remains suboptimum. Low rates of aspirin use and variations in use suggest a need to better translate clinical recommendations into practice. PMID- 10715256 TI - Myocardial infarction in treated hypertensive patients: the paradox of lower incidence but higher mortality in young blacks compared with whites. AB - BACKGROUND: Despite the impressive decline in coronary heart disease death rates, a mortality differential between blacks and whites persists. Our study objective was to determine whether excess mortality among well-controlled hypertensive black men compared with whites is due to differences in disease incidence or in case fatality. METHODS AND RESULTS: Of 3382 male subjects (1266 blacks and 2116 whites) enrolled between 1973 and 1996 and followed up through 1997 in a work site hypertension control program, 2343 were followed up until 60 years of age, and 1884 were followed up until >60 years of age (either continuing after 60 years [n=845] or beginning treatment at >/=60 years [n=1039]), with a mean follow up of 5.2 and 5.5 years, respectively. During follow-up, 186 myocardial infarction (MI) events (including 31 revascularizations) occurred, with 63 in patients <60 years and 123 in patients >/=60 years of age. Age-adjusted MI incidence was nearly twice as high for whites as blacks in younger (6.3 versus 3.4/1000 person-years) and older (14.1 versus 7.5 person-years) subjects. In contrast, the age-adjusted case fatality rate was 3-fold higher for younger blacks than for whites (37.8% versus 12.2%). In older patients, case fatality did not differ significantly between blacks and whites (37.6% versus 50. 3%). In separate Cox regression analyses, among younger blacks but not younger whites, history of diabetes and smoking were significantly associated with both incidence and fatality. CONCLUSIONS: In these treated male hypertensive patients with good blood pressure control (139.6/85.7 mm Hg), young blacks, despite a lower MI incidence, had higher MI mortality than did their white counterparts. Their higher case fatality rate was associated with fewer coronary artery revascularizations and a higher prevalence of diabetes and smoking. PMID- 10715255 TI - Major racial differences in coronary constrictor response between japanese and caucasians with recent myocardial infarction. AB - BACKGROUND: Enhanced coronary vasomotion may contribute to acute coronary occlusion during the acute phase of myocardial infarction (AMI). Japanese have a higher incidence of variant angina than Caucasian patients, but racial differences in vasomotor reactivity early after AMI are controversial. METHODS AND RESULTS: The same team studied 15 Japanese and 19 Caucasian patients within 14 days of AMI by acetylcholine injection into non-infarct-related (NIRA) and infarct-related (IRA) coronary arteries followed by nitroglycerin. Incidence of vasodilation, vasoconstriction, spasm, and basal tone were assessed in proximal, middle, and distal segments after each drug bolus by quantitative angiography. Japanese patients had much lower cholesterol levels than Caucasians (183+/-59 versus 247+/-53 mg/dL, P<0.006) but showed a lower incidence of vasodilation (2% versus 9% of coronary segments) and a greater incidence of spasm after acetylcholine (47% versus 15% of arteries, P<0.00001). Incidence of spasm was higher in IRAs than in NIRAs in both populations (67% versus 39% and 23% versus 11%, respectively). Multivessel spasm was more common (64% versus 17%, P<0.02) and vasoconstriction of nonspastic segments was greater in Japanese patients ( 23.4+/-14.9% versus -20.1+/-15.7%, P<0.02) in the presence of similar average basal coronary tone with respect to post-nitroglycerin dilation and of nonsignificant differences of coronary atherosclerotic score. CONCLUSIONS: Soon after AMI, Japanese patients exhibited a 3-fold-greater incidence of spasm and greater vasoconstriction of nonspastic segments after acetylcholine than Caucasians. The causes of such differences warrant further investigation because they may have relevant pathophysiological and therapeutic implications. PMID- 10715257 TI - New body surface isopotential map evaluation method to detect minor potential losses in non-Q-wave myocardial infarction. AB - BACKGROUND: Potential losses caused by stable non-Q-wave myocardial infarction (MI) are too small to diagnose with the use of standard ECG. The aim of the present study was to obtain accurate diagnostic criteria for this prognostically important disease with the help of body surface mapping. METHODS AND RESULTS: Body surface potentials were recorded with the use of 63 unipolar leads in 45 patients with a non-Q-wave MI (41 to 75 years old); 24 healthy adults, 42 patients with unstable angina, and 70 patients with Q-wave MI served as reference groups. Qualitative pathological features of the isopotential maps, such as onset time and site and magnitude of the first right-anterior/anterior minimum, as well as pathological negativities at that time, were defined in non-Q-wave MI cases. These features, which account for the activation sequence and the body surface projections of specific cardiac regions (Selvester classification), showed a 91% sensitivity and an 88% specificity for the detection of non-Q-wave MI. In comparison, the different departure maps (first third QRS, QRS, and QRST isoarea) resulted in less favorable specificities (50% to 58%). Concordance between the isopotential maps and the acute-phase ECG (90%), hypokinesis (64%), fixed perfusion defects (59%), and significant stenosis of the infarct-related coronary artery (87%) supported the concept that these isopotential map changes correspond to the supposed sites of MI. There were pathological features in 69% of patients with unstable angina, with similar concordances as in non-Q-wave MI. CONCLUSIONS: Isopotential maps revealed characteristic features that were suitable for the detection and localization of non-Q-wave MI in the clinical setting of unstable coronary artery disease. PMID- 10715258 TI - Effects of abciximab, ticlopidine, and combined Abciximab/Ticlopidine therapy on platelet and leukocyte function in patients undergoing coronary angioplasty. AB - BACKGROUND: Abciximab and ticlopidine reduce adverse cardiovascular events after percutaneous transluminal coronary angioplasty (PTCA). The goal of the current study was to determine if combined abciximab/ticlopidine therapy inhibits arterial thrombosis more effectively than either treatment alone. The effect of each therapy on platelet-leukocyte interactions was also investigated. METHODS AND RESULTS: Whole blood samples from 14 patients undergoing PTCA who received abciximab therapy, ticlopidine therapy, or both treatments were evaluated using dynamic experimental systems. Mural thrombus formation under arterial shear conditions (1500 s(-1)) was determined in a parallel plate flow chamber. Shear induced platelet aggregation was evaluated using a cone-and-plate viscometer at a shear rate of 3000 s(-1). Of the 3 treatments, combined abciximab/ticlopidine therapy produced the most consistent reduction in shear-induced platelet aggregation and the most prolonged inhibition of mural thrombosis. Three days after PTCA, abciximab/ticlopidine treatment decreased mural thrombus formation to approximately 50% of baseline values. Abciximab treatment alone inhibited mural thrombosis for only 1 day after PTCA, whereas ticlopidine treatment alone had no significant effect. Two hours after PTCA, abciximab therapy significantly decreased the number of circulating platelet-neutrophil aggregates but significantly enhanced P-selectin-mediated leukocyte adhesion on the collagen/von Willebrand factor-platelet surface. CONCLUSIONS: Combined therapy with abciximab and ticlopidine has a prolonged inhibitory effect on mural thrombosis formation relative to either treatment alone. Further, we demonstrated an unexpected effect of abciximab in enhancing P-selectin-mediated leukocyte adhesion. PMID- 10715259 TI - Angiotensin II has multiple profibrotic effects in human cardiac fibroblasts. AB - BACKGROUND: Angiotensin II (Ang II) is implicated in cardiac remodeling and is increasingly recognized for its profibrotic activity. METHODS AND RESULTS: Because little is known about the direct cellular effects of Ang II on human cardiac fibroblasts, we isolated fibroblasts from ventricles of explanted human hearts and added Ang II (100 nmol/L) to determine its role in growth, extracellular matrix accumulation, and adhesion. To assess which Ang II receptor is involved, Ang II was added in the presence of irbesartan (10 micromol/L), a specific AT(1) receptor antagonist; PD 123319 (10 micromol/L), a specific AT(2) receptor antagonist, or divalinil (100 nmol/L), an AT(4) receptor inhibitor. In human ventricles (n=13), message levels of atrial natriuretic peptide and AT(1) receptor were inversely correlated, which suggests a decrease in AT(1) receptor expression with progressive heart failure. Northern analysis and fluorescence activated cell sorting demonstrated AT(1) receptor in cultured human cardiac fibroblasts. Ang II increased mitogen-activated protein kinase activity and in DNA synthesis (5-fold, P<0.01) stimulated a 2-fold increase in transforming growth factor-beta(1) (P<0.05) mRNA levels at 2 hours and a 2-fold increase in laminin (P<0.05) and fibronectin (P<0.05) mRNA levels at 24 hours. Ang II also enhanced plasminogen activator inhibitor-1 expression, which inhibits metalloproteinases that degrade the extracellular matrix. All of these effects were inhibited by irbesartan but not PD 123319 or divalinil. In addition, Ang II increased cardiac fibroblast attachment to collagens I and III, which was associated with an increase in focal adhesion kinase activity. CONCLUSIONS: Activation of the AT(1) receptor in human heart promotes fibrosis. Ang II plays a novel role in stimulation of plasminogen activator inhibitor-1 expression and adhesion of cardiac fibroblasts to collagen. PMID- 10715260 TI - Clinical outcomes after ablation and pacing therapy for atrial fibrillation : a meta-analysis. AB - BACKGROUND: Radiofrequency ablation of the atrioventricular node and permanent pacing are used for symptomatic relief in patients with medically refractory atrial fibrillation. In this study, meta-analysis was used to clarify clinical outcomes and survival after ablation and pacing therapy using data from the published literature. METHODS AND RESULTS: We used 21 studies with a total of 1181 patients in the meta-analysis. All patients had medically refractory atrial tachyarrhythmias, primarily atrial fibrillation (97%). Nineteen measures of clinical outcome, encompassing quality of life, ventricular function, exercise duration, and healthcare use, were derived from the studies. The meta-analysis demonstrated significant improvement after ablation and pacing therapy in all outcome measures except fractional shortening, which demonstrated a trend toward improvement (P=0.08). Ejection fraction did show significant improvement (P<0.001). The calculated 1-year total and sudden death mortality rates after ablation and pacing therapy were 6.3% and 2.0%, respectively. CONCLUSIONS: Ablation and pacing therapy improves a broad range of clinical outcomes for patients with medically refractory atrial fibrillation. The calculated 1-year mortality rates after this therapy are low and comparable with medical therapy. PMID- 10715261 TI - Reversal of atrial electrical remodeling after cardioversion of persistent atrial fibrillation in humans. AB - BACKGROUND: Although atrial electrical remodeling has been studied extensively in animal models, the reversibility of this phenomenon after termination of clinical atrial fibrillation (AF) has not been demonstrated. We aimed to examine this important question of reversibility by using AF cycle length (AFCL) and coupling intervals of atrial premature beats after cardioversion as measures of atrial refractoriness. METHODS AND RESULTS: We measured AFCL at the right atrial appendage and distal coronary sinus before attempting internal cardioversion in 39 patients with persistent AF. Patients were monitored by daily transtelephonic recordings after discharge and admitted rapidly for repeat internal cardioversion if there was spontaneous AF recurrence. Measurements of AFCL were repeated immediately before repeat cardioversions in the 17 patients who had recurrence of AF. There was an increase in AFCL from the initial cardioversion to that measured at the time of first AF recurrence at both the right atrial appendage (161+/-22 vs 167+/-26 ms, P=0.05) and distal coronary sinus (162+/-20 vs 168+/-22 ms, P=0.01) sites. The magnitude of increase in AFCL was positively correlated with duration of sinus rhythm before AF recurrence (r=0.524, P=0.001). Other measures of refractoriness (shortest coupling interval of atrial premature beats and directly measured refractory periods after cardioversion) also increased from initial to subsequent cardioversions. CONCLUSIONS: These findings demonstrate that changes in atrial electrophysiology associated with chronic AF in humans are reversible after cardioversion and that the extent of this reversal is dependent on the duration of sinus rhythm after cardioversion. PMID- 10715262 TI - Peak oxygen uptake better predicts outcome than submaximal respiratory data in heart transplant candidates. AB - BACKGROUND: Many studies have focused on the prognostic power of peak oxygen uptake VO(2) in patients with chronic heart failure, but maximal exercise testing is not without risk. The purpose of the present study was, therefore, to assess the prognostic significance of the steepness of changes in ventilation and carbon dioxide output VO(2) during submaximal exercise in comparison with VO(2). METHODS AND RESULTS: The study population consisted of 284 adult heart transplant candidates who performed a graded maximal bicycle ergometer test with respiratory gas analysis. Using the respiratory data up to a gas exchange ratio of 1.0, 3 submaximal slopes were calculated in each patient. During follow-up (median, 1.33 years), 57 patients died and 149 had >/=1 cardiovascular event. When using Cox proportional hazards analysis, both peak VO(2) and submaximal respiratory slopes predicted outcome before and after accounting for age, sex, and body mass index. However, whereas the prognostic power of peak VO(2) was independent of submaximal respiratory data, the prognostic significance of the slopes was lost after controlling for peak VO(2). Stepwise regression analysis even selected peak VO(2) as an independent prognostic index among the following factors: cause of heart failure, ejection fraction, pulmonary vascular resistance, natremia, and the forced expiratory volume in 1 s. CONCLUSIONS: Respiratory data during submaximal exercise are significant predictors of outcome in patients with chronic heart failure, but their prognostic power is inferior to that of peak VO(2). However, these data may be useful when maximal exercise is contraindicated or not achievable. PMID- 10715263 TI - Construction and functional evaluation of a single-chain antibody fusion protein with fibrin targeting and thrombin inhibition after activation by factor Xa. AB - BACKGROUND: Recombinant technology was used to produce a new anticoagulant that is preferentially localized and active at the site of the clot. METHODS AND RESULTS: The variable regions of the heavy and light chains of a fibrin-specific antibody were amplified by polymerase chain reaction (PCR) with hybridoma cDNA. To obtain a functional single-chain antibody (scFv), a linker region consisting of (Gly(4)Ser)(3) was introduced by overlap PCR. After the scFv clones were ligated with DNA encoding the pIII protein of the M13 phage, high-affinity clones were selected by 10 rounds of panning on the Bbeta15-22 peptide of fibrin (beta peptide). Hirudin was genetically fused to the C-terminus of the variable region of the light chain. To release the functionally essential N-terminus of hirudin at the site of a blood clot, a factor Xa recognition site was introduced between scFv(59D8) and hirudin. The fusion protein was characterized by its size on SDS PAGE (36 kDa), by Western blotting, by its cleavage into a 29-kDa (single chain alone) and 7-kDa (hirudin) fragment, by its binding to beta-peptide, and by thrombin inhibition after Xa cleavage. Finally, the fusion protein inhibited appositional growth of whole blood clots in vitro more efficiently than native hirudin. CONCLUSIONS: A fusion protein was constructed that binds to a fibrin specific epitope and inhibits thrombin after its activation by factor Xa. This recombinant anticoagulant effectively inhibits appositional clot growth in vitro. Its efficient and fast production at low cost should facilitate a large-scale evaluation to determine whether an effective localized antithrombin activity can be achieved without systemic bleeding complications. PMID- 10715265 TI - The cardiac Fas (APO-1/CD95) Receptor/Fas ligand system : relation to diastolic wall stress in volume-overload hypertrophy in vivo and activation of the transcription factor AP-1 in cardiac myocytes. AB - BACKGROUND: Fas (APO-1/CD95) is a transmembrane receptor belonging to the tumor necrosis factor receptor superfamily. Cross-linking of Fas by Fas ligand (FasL), a tumor necrosis factor-alpha-related cytokine, promotes apoptosis and/or transcription factor activation in a highly cell-type-specific manner. The biological consequences of Fas activation in cardiomyocytes and the regulation of Fas and FasL abundance in the myocardium in vivo remain largely unknown. METHODS AND RESULTS: As shown by immunohistochemistry, Fas was expressed on the sarcolemma of cardiomyocytes in left ventricular tissue sections. Moreover, FasL was constitutively expressed in the myocardium and in isolated cardiomyocytes, as revealed by reverse transcription polymerase chain reaction and Western blotting. Left ventricular abundance of Fas but not FasL was upregulated in a rat model of compensated volume-overload hypertrophy and was closely related to diastolic but not systolic wall stress as determined by MRI. Cardiomyocyte apoptosis was not enhanced in volume-overload hypertrophy despite the increased expression of Fas and the presence of FasL in the myocardium. Moreover, injection of mice with an agonistic anti-Fas antibody promoted hepatocyte but not cardiomyocyte apoptosis in vivo. Stimulation of isolated cardiomyocytes with recombinant FasL promoted an activation of the transcription factor AP-1 as shown by electrophoretic mobility shift assays but did not induce cell death. CONCLUSIONS: Fas and FasL are constitutively expressed in the myocardium and in cardiomyocytes. Myocardial expression of Fas is closely related to diastolic loading conditions in vivo. Signaling pathways emanating from Fas are coupled to an activation of the transcription factor AP-1 in cardiomyocytes. PMID- 10715264 TI - Troglitazone improves recovery of left ventricular function after regional ischemia in pigs. AB - BACKGROUND: This study determined whether treatment of normal (nondiabetic) pigs with the insulin-sensitizing agent troglitazone improves recovery of left ventricular (LV) function after acute ischemia and whether such effects are associated with altered myocardial substrate metabolism. METHODS AND RESULTS: Juvenile pigs (n=6) were treated with troglitazone (75 mg. kg(-1). d(-1) PO) for 8 weeks. Untreated pigs (n=8) served as controls. Under anesthetized, open-chest conditions, pigs underwent 90 minutes of moderate regional LV ischemia and 90 minutes of reperfusion. Regional LV function was assessed with subendocardial sonomicrometry crystals. Fasting plasma insulin and free fatty acid levels were lower in troglitazone-treated pigs than in untreated pigs, whereas blood glucose did not differ between groups. These findings suggest that treatment enhanced systemic insulin sensitivity. Baseline hemodynamics and regional LV function did not differ between groups. After ischemia and reperfusion, systolic function (external work) of the ischemic region recovered to 44+/-6% of baseline in troglitazone-treated pigs versus 18+/-6% of baseline in untreated pigs (P<0.05). Regional diastolic function (maximum rate of wall expansion) recovered to 78+/-7% of baseline in treated pigs versus 52+/-7% of baseline in untreated pigs (P<0.05). Recovery of global LV systolic and diastolic function was also significantly greater in treated pigs. Myocardial glucose uptake did not differ between groups under any condition; however, net myocardial lactate uptake after reperfusion was 7 times greater in troglitazone-treated pigs than in untreated pigs, suggesting that treatment enhanced myocardial carbohydrate oxidation after reperfusion. CONCLUSIONS: In nondiabetic pigs, chronic troglitazone treatment improves recovery of LV systolic and diastolic function after acute ischemia. PMID- 10715266 TI - Dose-dependence of 4-aminopyridine plasma concentrations and electrophysiological effects in dogs : potential relevance to ionic mechanisms in vivo. AB - BACKGROUND: Previous investigators have administered 4-aminopyridine (4AP) to dogs to evaluate the role of transient outward current (I(to)) in vivo; however, plasma concentrations of 4AP were not measured, and it is therefore uncertain which cardiac ion channels were blocked at the concentrations achieved. METHODS AND RESULTS: We applied high-performance liquid chromatography to measure 4AP concentrations produced by intravenous 4AP administration to dogs. A previously described dose regimen produced plasma concentrations that increased during the maintenance infusion but never exceeded 250 micromol/L and caused significant mortality. Whole-cell patch-clamp experiments on isolated canine myocytes showed that even the maximum 4AP concentrations achieved in vivo failed to alter ventricular I(to) and had very small effects on atrial I(to); however, concentrations achieved in vivo had a strong inhibitory effect on the dog ultrarapid delayed rectifier (I(Kur.d)), present only in atrial cells. We designed a loading and maintenance infusion regimen to produce stable 4AP plasma concentrations. At concentrations in the range of 25 and 50 micromol/L, 4AP had no effect on ventricular refractory period but increased atrial refractoriness significantly, consistent with the results of voltage clamp studies. CONCLUSIONS: The interpretation of previous studies using intravenous 4AP administration to inhibit I(to) in dogs in vivo needs to be reevaluated in light of the fact that the infusion regimens used produce plasma concentrations that are inadequate to affect ventricular I(to). Our findings also support the concept that selective inhibition of ultrarapid delayed rectifier current can prolong atrial refractory periods without affecting ventricular refractoriness. PMID- 10715267 TI - Atrial fibrillation produced by prolonged rapid atrial pacing is associated with heterogeneous changes in atrial sympathetic innervation. AB - BACKGROUND: Structural and electrophysiological changes of the atria occur with prolonged rapid rates; however, the effects of sustained atrial fibrillation (AF) on autonomic innervation of the atria are unknown. We hypothesized that electrophysiological remodeling from rapid atrial rates is accompanied by altered atrial autonomic innervation. METHODS AND RESULTS: Six dogs (paced group) underwent atrial pacing at 600 bpm; 9 dogs (control animals) were not paced. All paced dogs developed sustained AF by week 4 of pacing. All 15 animals underwent positron emission tomography imaging of the atria with [C-11] hydroxyephedrine (HED) to label sympathetic nerve terminals. HED retention in the atria was significantly greater in paced dogs compared with control animals (P=0.03). Tissue samples from the atrial appendages had a greater concentration of norepinephrine in paced animals than in control animals (P=0.01). The coefficient of variation of HED retention was also greater in paced animals (P=0.05) and was greater in the right atrium than in the left atrium (P=0.004). Epicardial activation maps of AF were obtained in the paced animals at baseline and with autonomic manipulation. Mean AF cycle length was longer in the right atrium (109.2+/-5 ms) than in the left atrium (85.8+/-5.5 ms) at baseline (P=0.005). AF cycle length did not vary significantly from baseline (97.6+/-13.4 ms) with stellate stimulation (100.5+/-6 ms) but lengthened with propranolol (107.5+/-6.1 ms, P=0.03). CONCLUSIONS: Rapid rates of AF produce a heterogeneous increase in atrial sympathetic innervation. These changes parallel disparate effects of rapid pacing-induced AF on atrial electrophysiology. PMID- 10715268 TI - Cellular arrhythmogenic effects of congenital and acquired long-QT syndrome in the heterogeneous myocardium. AB - BACKGROUND: Certain alterations by mutations or drugs of the potassium currents I(Ks) and I(Kr) and the sodium current I(Na) give rise to several types of the long-QT syndrome. I(Ks) is heterogeneously distributed across the ventricular wall. METHODS AND RESULTS: We investigated the effects of reducing I(Ks) or I(Kr) or enhancing late I(Na) (to simulate the 3 forms of long-QT syndrome) on action potential duration (APD) in the context of I(Ks) heterogeneity. We introduced I(Ks) heterogeneity in the Luo-Rudy dynamic cell model to simulate epicardial, endocardial, and midmyocardial (M) cells. Results demonstrated higher susceptibility of M cells to the development of arrhythmogenic early afterdepolarizations (EADs) in isolated cells and poorly coupled tissue. An important observation is that I(Kr) block or late I(Na) acts to increase APD differences between the cell types, whereas I(Ks) block minimizes such differences. Also, for normal transverse coupling, EADs develop in the endocardial region rather than in the M region as the result of strong electrotonic interaction. CONCLUSIONS: I(Ks) heterogeneity and intercellular coupling strongly influence EAD development during interventions or disorders that prolong APD. M cells in isolation or in poorly coupled tissue are more susceptible to EAD development than epicardial or endocardial cells. In well coupled myocardium, EAD formation in the subendocardium can be the source of focal arrhythmias or provide the trigger for reentrant excitation. The efficacy of I(Ks) block in minimizing APD dispersion could have important implications for antiarrhythmic therapy. PMID- 10715269 TI - Losartan and its metabolite E3174 modify cardiac delayed rectifier K(+) currents. AB - BACKGROUND: The effects of type 1 angiotensin II receptor antagonist losartan and its metabolite E3174 on transmembrane action potentials, hKv1.5, HERG, and I(Ks) currents were analyzed. METHODS AND RESULTS: Guinea pig ventricular action potentials were recorded with microelectrode techniques and hKv1.5 and HERG currents with the whole-cell patch-clamp technique. I(Ks) was recorded in guinea pig ventricular myocytes with the perforated-nystatin-patch configuration. Losartan and E3174 transiently increased the hKv1.5 current by 8.0+/-1.4% and 7.4+/-1.6%, respectively. Thereafter, they produced a voltage-dependent block, E3174 being more potent than losartan (P<0.05) for this effect. Losartan decreased HERG currents elicited at 0 mV (23.3+/-4.8%), whereas E3174 increased the current (30.5+/-6.2%). Both drugs shifted the midpoint of the activation curve of HERG channels to more negative potentials. In ventricular myocytes, losartan and E3174 inhibited the I(Ks) (18.4+/-3.2% and 6. 5+/-0.7%, respectively). Losartan-induced block was voltage-independent, whereas E3174 shifted the midpoint of the activation curve to more negative potentials. Losartan lengthened the duration of the action potentials at both 50% and 90% of repolarization, whereas E3174 slowed only the final phase of the repolarization process. CONCLUSIONS: These results demonstrated that at therapeutic concentrations, both losartan and E3174 modified the cardiac delayed rectifier hKv1.5, HERG, and Ks currents. PMID- 10715270 TI - Aspirin. PMID- 10715272 TI - New national institutes of health website helps locate clinical trials PMID- 10715271 TI - Images in cardiovascular medicine. Myocardial infarction in children with hypoplastic coronary arteries. PMID- 10715273 TI - Physiology of ventricular septal defect shunt flow in the fetus examined by color Doppler M-mode. PMID- 10715274 TI - Chlamydia pneumoniae in coronary artery disease. PMID- 10715276 TI - Qualification of the concepts of unqualified success and unmitigated failure. PMID- 10715277 TI - Qualification of the concepts of unqualified success and unmitigated failure. PMID- 10715279 TI - A troubled field scores a Hit PMID- 10715278 TI - Assessment of coronary flow reserve by contrast-enhanced second harmonic echo Doppler. PMID- 10715280 TI - Antibodies that protect mice against ebola virus hold promise of vaccine and therapy for disease PMID- 10715281 TI - New National Institutes of Health website helps locate clinical trials. PMID- 10715282 TI - Depression after coronary artery bypass surgery increases risk of more heart problems. PMID- 10715283 TI - Prostate biopsy: a wealth of information when done and interpreted correctly. PMID- 10715284 TI - Clinical utility of the percentage of positive prostate biopsies in defining biochemical outcome after radical prostatectomy for patients with clinically localized prostate cancer. AB - PURPOSE: To determine the clinical utility of the percentage of positive prostate biopsies in predicting prostate-specific antigen (PSA) outcome after radical prostatectomy (RP) for men with PSA-detected or clinically palpable prostate cancer. METHODS: A Cox regression multivariable analysis was used to determine whether the percentage of positive prostate biopsies provided clinically relevant information about PSA outcome after RP in 960 men while accounting for the previously established risk groups that are defined according to pretreatment PSA level, biopsy Gleason score, and the 1992 American Joint Committee on Cancer (AJCC) clinical T stage. The findings were then tested using an independent surgical database that included data for 823 men. RESULTS: Controlling for the known prognostic factors, the percentage of positive prostate biopsies added clinically significant information (P <.0001) regarding time to PSA failure after RP. Specifically, 80% of the patients in the intermediate-risk group (1992 AJCC T2b, or biopsy Gleason 7 or PSA > 10 ng/mL and /= 60 g underwent treatment with NAAD before TIPPB to reduce the prostate volume (n = 163). The median duration of NAAD was 3.4 months before TIPPB (range, 1 to 8 months). To assess the benefit of NAAD, a matched-pair analysis was performed. The American Society of Therapeutic Radiology and Oncology Consensus Group definition of prostate-specific antigen (PSA) relapse-free survival (RFS) was used with the added caveat of an absolute increase of >/= 1.0 ng/mL. Differences in pretreatment PSA, Gleason scores, and stage were analyzed by Kaplan-Meier curves and the log-rank test. RESULTS: Two hundred sixty-three patients were matched, with a median follow-up duration of 46 months (range, 24 to 76 months). The actuarial 5-year PSA-RFS rate for all 263 patients is 86.5%. The 5-year PSA RFS rate for patients treated with NAAD and TIPPB was 87.1% compared with 86.9% for those treated with TIPPB only (P =.935). Subgroup analysis by Gleason score groupings, pretreatment PSA, or stage of disease failed to identify any factors for which androgen ablation was beneficial. CONCLUSION: We were unable to identify any improvement with the addition of NAAD to TIPPB in patients with localized prostate cancer in this retrospective matched-pair analysis. Furthermore, there was no subset for which the addition of NAAD was found to be beneficial. Clarification of the role and duration of NAAD in patients with early stage prostate cancer will require prospective data. PMID- 10715288 TI - Oxaliplatin or paclitaxel in patients with platinum-pretreated advanced ovarian cancer: A randomized phase II study of the European Organization for Research and Treatment of Cancer Gynecology Group. AB - PURPOSE: This was a multicentric, open, randomized, phase II study of single agent paclitaxel and oxaliplatin to evaluate the efficacy of oxaliplatin in a relapsing progressive ovarian cancer patient population and to analyze the safety profile and impact of both agents on quality of life, time to progression, and survival. PATIENTS AND METHODS: Eighty-six patients with platinum-pretreated advanced ovarian cancer were randomly assigned to two arms: 41 received paclitaxel at 175 mg/m(2) over 3 hours every 3 weeks, and 45 received oxaliplatin at 130 mg/m(2) over 2 hours every 3 weeks. For inclusion, patients had to have a performance status of 0 to 2 and to have received at least one and no more than two prior cisplatin- and/or carboplatin-containing chemotherapy regimens within the last 12 months. RESULTS: Seven confirmed responses were observed in each arm, for an overall response rate in the total treated population of 17% (95% confidence interval [CI], 7% to 32%) in the paclitaxel arm and 16% (95% CI, 7% to 29%) in the oxaliplatin arm. Median time to progression was 14 weeks and 12 weeks, and overall survival was 37 weeks and 42 weeks in the paclitaxel and oxaliplatin arms, respectively. Among 63 patients with a 0- to 6-month progression-free, platinum-free interval, there were five objective responses with paclitaxel in 31 patients and two objective responses with oxaliplatin in 32 patients. Nine patients (22%) in the paclitaxel arm had grade 3 or 4 neutropenia (National Cancer Institute of Canada [NCIC] Common Toxicity Criteria). Two patients (4%) experienced grade 3 thrombocytopenia in the oxaliplatin arm. Maximum grade (grade 3) NCIC neurosensory toxicity was experienced by three patients (7%) in the paclitaxel arm and by four patients (9%) in the oxaliplatin arm. CONCLUSION: Single-agent oxaliplatin at 130 mg/m(2) every 3 weeks is active with moderate toxicity in patients with cisplatin-/carboplatin-pretreated advanced ovarian cancer. PMID- 10715289 TI - Factors that predict the referral of breast cancer patients onto clinical trials by their surgeons and medical oncologists. AB - PURPOSE: To improve understanding of physicians' reluctance to refer patients to clinical trials. METHODS: This study was conducted in a large metropolitan region from 1993 to 1995 using a two-staged population-based sampling strategy. A total of 147 physicians discussed 245 patient cases and their own knowledge, attitudes, and practices toward clinical trials. RESULTS: Ninety-three patients (38. 0%) were offered a trial, and 49 (52.7%) of them agreed to participate. Forty-five patients (18.4%) actually received their adjuvant therapy on trial. Older patients and those with a poorer prognosis were less likely to be referred. Patients who delayed their decision were more than three times as likely to participate in a trial and more than eight times as likely to participate when they were reported to be actively involved in making the decision. Generally, physicians in university settings and who had formal support from a cooperative group were more likely to refer patients to trials. More specifically, surgeons referred more patients to trials when they felt comfortable explaining trials or believed that treatment should not stray from protocol. Oncologists were less likely to make referrals if they perceived the paperwork to be onerous or entry requirements to be too stringent. Surgeons' participation in recommending adjuvant therapy to patients resulted in more trial referrals unless they treated their patients with tamoxifen. CONCLUSION: (1) Physicians still need to overcome attitudinal and practical barriers to trial participation, (2) more support for physicians is needed, (3) surgeons may play a pivotal role in the recruitment of patients to adjuvant therapy trials, and (4) garnering patient enthusiasm for trial participation and involving them in the choice of adjuvant therapy may be key components to increasing trial enrollment. PMID- 10715290 TI - Docetaxel administered on a weekly basis for metastatic breast cancer. AB - PURPOSE: To evaluate the safety and efficacy of weekly docetaxel in women with metastatic breast cancer. PATIENTS AND METHODS: Twenty-nine women were enrolled onto a study of weekly docetaxel given at 40 mg/m(2)/wk. Each cycle consisted of 6 weeks of therapy followed by a 2-week treatment break, repeated until disease progression or removal from study for toxicity or patient preference. Fifty-two percent of patients had been previously treated with adjuvant chemotherapy; 21% had received prior chemotherapy for metastatic breast cancer, and 31% had previously received anthracyclines. All patients were assessable for toxicity; two patients were not assessable for response but are included in an intent-to treat analysis. RESULTS: Patients received a median of 18 infusions, with a median cumulative docetaxel dose of 720 mg/m(2). There were no complete responses. Twelve patients had partial responses (overall response rate, 41%; 95% confidence interval, 24% to 61%), all occurring within the first two cycles. Similar response rates were observed among subgroups of patients previously treated either with any prior chemotherapy or with anthracyclines. An additional 17% of patients had stable disease for at least 6 months. The regimen was generally well tolerated. There was no grade 4 toxicity. Only 28% of patients had any grade 3 toxicity, most commonly neutropenia and fatigue. Acute toxicity, including myelosuppression, was mild. Fatigue, fluid retention, and eye tearing/conjunctivitis became more common with repetitive dosing, although these side effects rarely exceeded grade 2. Dose reductions were made for eight of 29 patients, most often because of fatigue (n = 5). CONCLUSION: Weekly docetaxel is active in treating patients with metastatic breast cancer, with a side effect profile that differs from every-3-weeks therapy. PMID- 10715291 TI - Does locoregional radiation therapy improve survival in breast cancer? A meta analysis. AB - PURPOSE: Recent randomized trials in women with node-positive breast cancer who received systemic treatment report that locoregional radiation therapy improves survival. Previous trials failed to detect a difference in survival that results from its use. A systematic review of randomized trials that examine the effectiveness of locoregional radiation therapy in patients treated by definitive surgery and adjuvant systemic therapy was conducted. METHODS: Randomized trials published between 1967 and 1999 were identified through MEDLINE database, CancerLit database, and reference lists of relevant articles. Relevant data was abstracted. The results of randomized trials were pooled using meta-analyses to estimate the effect of treatment on any recurrence, locoregional recurrence, and mortality. RESULTS: Eighteen trials that involved a total of 6,367 patients were identified. Most trials included both pre- and postmenopausal women with node positive breast cancer treated with modified radical mastectomy. The type of systemic therapy received, sites irradiated, techniques used, and doses of radiation delivered varied between trials. Data on toxicity were infrequently reported. Radiation was shown to reduce the risk of any recurrence (odds ratio, 0.69; 95% confidence interval [CI], 0.58 to 0.83), local recurrence (odds ratio, 0.25; 95% CI, 0.19 to 0.34), and mortality (odds ratio, 0.83; 95% CI, 0.74 to 0.94). CONCLUSION: Locoregional radiation after surgery in patients treated with systemic therapy reduced mortality. Several questions remain on how these results should be translated into current-day clinical practice. PMID- 10715292 TI - Building bridges between physicians and patients: results of a pilot study examining new tools for collaborative decision making in breast cancer. AB - PURPOSE: To present the results of a pilot study testing an intervention designed to improve the quality of medical consultations between breast cancer patients and physicians and, in particular, to report the effects of the intervention on the quality of treatment decisions, the quality of communication, and the satisfaction of patients and physicians. PATIENTS AND METHODS: We enrolled 24 predominantly white, well-educated, early-stage breast cancer patients who were facing local or systemic treatment decisions in a sequential, controlled trial. All patients received a visit preparation session before the consultation in which a trained researcher helped patients organize their questions and concerns. In the control, a researcher observed the consultation. In the intervention, a researcher helped create an agenda, facilitated the discussion, and created a record of the consultation in real time. Valid and reliable surveys measured the quality of treatment decisions and satisfaction with the consultation. RESULTS: Patients in the intervention achieved significantly higher final decision quality scores compared with control patients (median score, 14 v 10, respectively; P =.008) and a significantly higher level of intersubjective agreement with their physicians about decision quality (Cohen's kappa, 0.49 v 0.285, respectively; P <.0001). Consultation recording methods did not affect the length of time required for the consultation. CONCLUSION: Consultation recording methods provide a promising innovation for medical consultations. Further studies are warranted to broaden the findings, assess impacts on the quality of decisions, cost, and health, and develop practical ways to integrate consultation recording methods into clinics. PMID- 10715293 TI - Sucralfate in the prevention of treatment-induced diarrhea in patients receiving pelvic radiation therapy: A North Central Cancer Treatment Group phase III double blind placebo-controlled trial. AB - PURPOSE: Randomized studies have suggested that sucralfate is effective in mitigating diarrhea during pelvic radiation therapy (RT). This North Central Cancer Treatment Group study was undertaken to confirm the antidiarrheal effect of sucralfate. Several other measures of bowel function were also assessed. PATIENTS AND METHODS: Patients receiving pelvic RT to a minimum of 45 Gy at 1.7 to 2.1 Gy/d were eligible for the study. Patients were assigned randomly, in double-blind fashion, to receive sucralfate (1.5 g orally every 6 hours) or an identical looking placebo during pelvic RT. RESULTS: One hundred twenty-three patients were randomly assigned and found assessable. Overall, there was no significant difference in patient characteristics between those receiving sucralfate and those receiving placebo. Moderate or worse diarrhea was observed in 53% of patients receiving sucralfate versus 41% of those receiving placebo. Compared with patients receiving placebo, more sucralfate-treated patients reported fecal incontinence (16% v 34%, respectively; P =. 04) and need for protective clothing (8% v 23%, respectively; P =. 04). The incidence and severity of nausea were worse among those taking sucralfate (P =.03). Analysis of patient reported symptoms 10 to 12 months after RT showed a nonsignificant trend toward more problems in patients taking sucralfate than in those taking placebo (average, 2.3 v 1.9 problems, respectively; P =.34). CONCLUSION: Sucralfate did not decrease pelvic RT-related bowel toxicity by any of the end points measured and seems to have aggravated some gastrointestinal symptoms. PMID- 10715294 TI - Radiation therapy and high-dose tamoxifen in the treatment of patients with diffuse brainstem gliomas: results of a Brazilian cooperative study. Brainstem Glioma Cooperative Group. AB - PURPOSE: The efficacy of radiation therapy (RT) combined with tamoxifen (TX) was tested in patients diagnosed with diffuse brainstem gliomas in a multicenter trial. PATIENTS AND METHODS: TX was administered orally (maintenance dose: 200 mg/m(2) per day) along with conventional local RT and then continued for 52 additional weeks. Survival, tumoral radiologic response, and toxicity were evaluated. Compliance was assessed using pharmacokinetic measurements. RESULTS: Of 29 patients, 27 completed RT (median dose, 54 Gy). Of 22 assessable patients, 11 (50%) had an objective radiologic response. The mean TX steady-state serum level was 2.44 micromol/L +/- 1.02 micromol/L. Only three patients completed the entire course of treatment without tumoral progression or significant toxicity. Common side effects included nausea and vomiting. Hepatotoxicity (five patients), neurotoxicity (two patients), venous thrombosis (one patient), bilateral ovarian cysts (two patients), and transient neutropenia (one patient) were also observed. Median survival was 10.3 months. Only four patients remain alive without tumoral progression. The 1-year survival rate (mean +/- SD) was 37.0% +/- 9.5%. CONCLUSION: This treatment combination produced no significant change in the overall poor prognosis of these patients. Most tumors responded initially to treatment but recurred as the study progressed. A minority of patients seemed to benefit from the extended use of TX. Generally, treatment was well tolerated, with good patient compliance, but we recommend continuous close monitoring for side effects. Based on our poor results, we recommend that alternative treatments be tested in patients with this type of tumor. PMID- 10715296 TI - Biologic factors determine prognosis in infants with stage IV neuroblastoma: A prospective Children's Cancer Group study. AB - PURPOSE: A prospective Children's Cancer Group study, CCG-3881, has been completed to determine if a more accurate prediction of prognosis by biologic features can identify subgroups of infants with stage IV neuroblastoma (NBL) who require differing intensities of treatment. PATIENTS AND METHODS: One hundred thirty-four infants were registered from June 1989 to August 1995, with a median follow-up of 47.1 months (range, 0 to 88 months). The biologic factors examined were tumor MYCN copy number, Shimada histopathologic classification, serum ferritin, and bone marrow immunocytology (sensitivity, one tumor cell per 10(5) bone marrow cells). Patients treated on CCG-3881 (n = 116) received four-drug chemotherapy for 9 months (cisplatin, cyclophosphamide, doxorubicin, and etoposide), with surgery and local radiation to residual disease. After January 1991, all subsequent infants with tumor MYCN amplification (n = 18) were transferred after one cycle of therapy to the high-risk CCG-3891 protocol (open January 1991 to April 1996) for more intensive treatment. RESULTS: The 3-year event-free survival (EFS) and overall survival (mean +/- SD) for the 134 infants were 63% +/- 5% and 71% +/- 5%, respectively. Patients whose tumors were without MYCN amplification had a 93% +/- 4% 3-year EFS, whereas those with amplified MYCN had a 10% +/- 7% 3-year EFS (P <. 0001). Each of the other biologic features studied had prognostic significance in univariate analysis but not after stratifying by MYCN copy number. CONCLUSION: Infants less than 1 year of age at diagnosis with stage IV NBL have a much improved outcome compared with children >/= 1 year of age. Nonamplified MYCN tumors identify a group of infants with a 93% +/- 4% EFS, whereas amplified MYCN copy number clearly identifies patients who are unlikely to survive despite intensive chemotherapy. PMID- 10715295 TI - Single-arm, open-label phase II study of intravenously administered tirapazamine and radiation therapy for glioblastoma multiforme. AB - PURPOSE: This phase II study tested the efficacy and safety of tirapazamine (Sanofi Synthelabo Research, Malvern, PA), a bioreductive agent, in glioblastoma multiforme (GBM) patients. The patients were staged according to a model constructed by a recursive partitioning analysis (RPA) of glioma patients in prior Radiation Therapy Oncology Group (RTOG) trials and compared with a matched standard population, as predicted by the model. PATIENTS AND METHODS: A total of 124 patients diagnosed with a GBM were treated with radiation therapy and intravenous tirapazamine between January 27,1995, and April 25,1997. All patients received 60 Gy in 2-Gy fractions. Tirapazamine was delivered three times a week for 12 treatments during radiotherapy. Fifty-five patients received tirapazamine at 159 mg/m(2). A second dose level, 260 mg/m(2), was opened, and 69 patients were entered. RESULTS: There was no significant survival advantage to the drug in any RPA class at either dose level. The median survival time was 10.8 months for the patient population treated with the 159-mg/m(2) dose of tirapazamine and 9.5 months for the group treated with the 260-mg/m (2) dose of tirapazamine. Survival times by RPA class for patients receiving tirapazamine at 159 mg/m(2) were 27.4 months (class III), 10.8 months (class IV), 7.9 months (class V), and 3.8 months (class VI). Survival times by RPA class for patients receiving tirapazamine at 260 mg/m(2) were 16.2 months (class III), 10.3 months (class IV), 5. 1 months (class V), and 1.3 months (class VI). Patients in RPA class III treated in the 159 mg/m(2) dose arm had a notably longer survival than patients in the RTOG database RPA class III, but the difference did not reach statistical significance. There were no fatal toxicities. Grade 3/4 toxicities were more frequent at the higher dose level. CONCLUSION: Survival in the population treated with radiation and tirapazamine was equivalent to the control population. Patients in RPA class III treated with radiation and tirapazamine at the 159 mg/m(2) dose had a longer survival when compared with the historical controls. The improvement in survival did not reach statistical significance. Toxicity was acceptable in both treatment arms, but grade 3/4 toxicities were more frequent in the higher dose regimen. PMID- 10715297 TI - Prevention of central venous catheter-related infections and thrombotic events in immunocompromised children by the use of vancomycin/ciprofloxacin/heparin flush solution: A randomized, multicenter, double-blind trial. AB - PURPOSE: To determine whether an antibiotic flush solution containing vancomycin, heparin, and ciprofloxacin (VHC) can prevent the majority of line infections. PATIENTS AND METHODS: A prospective double-blind study was performed comparing VHC to vancomycin and heparin (VH) to heparin alone in 126 pediatric oncology patients. RESULTS: The 153 assessable lines resulted in 36,944 line days studied. There were 58 blood stream infections (43 gram-positive, 14 gram-negative, and one fungal). Forty were defined as line infections (31 heparin, three VH, six VHC). The time to develop a line infection was significantly increased using either antibiotic flush (VH, P =.011; VHC, P =.036). The rate of total line infections (VH, P =.004; VHC, P =.005), gram-positive line infections (VH, P =. 028; VHC, P =.022), and gram-negative line infections (VH, P =.006; VHC, P =.003) was significantly reduced by either VH or VHC. Sixty-two (41%) of the lines developed 119 occlusion episodes (heparin, 3.99 per 1,000 line days; VHC, 1.75 per 1,000 line days; P =.0005). Neither antibiotic could be detected after flushing, and no adverse events were detected, including increased incidence of vancomycin-resistant Enterococcus colonization or disease. CONCLUSION: The use of either VH or VHC flush solution significantly decreased the complications associated with the use of tunneled central venous lines in immunocompromised children and would save significant health care resources. PMID- 10715298 TI - Feasibility and toxicity of chemoembolization for children with liver tumors. AB - PURPOSE: To determine the feasibility, toxicity, and efficacy of hepatic arterial chemoembolization (HACE) in pediatric patients with refractory primary malignancies of the liver. PATIENTS AND METHODS: Six patients with hepatoblastoma (HB), three with hepatocellular carcinoma (HCC), and two with undifferentiated sarcoma of the liver were treated with HACE every 2 to 4 weeks until their tumors became surgically resectable or they showed signs of disease progression. All but one newly diagnosed patient with HCC had previously received systemic chemotherapy. RESULTS: All patients with HB and HCC responded to HACE, as measured by imaging studies and alpha-fetoprotein levels. Surgical resection (complete or microscopic residual disease) was feasible in five of 11 patients, and three patients remain alive with no evidence of disease. Elevated liver transaminase and bilirubin levels were seen after each one of the 46 courses of HACE. Other toxicities included fever, pain, nausea, vomiting, and transient coagulopathy. CONCLUSION: HACE is feasible, well tolerated, and effective in inducing surgical resectability of primary hepatic tumors in children. PMID- 10715299 TI - Intensification with intermediate-dose intravenous methotrexate is effective therapy for children with lower-risk B-precursor acute lymphoblastic leukemia: A Pediatric Oncology Group study. AB - PURPOSE: To determine whether early intensification with 12 courses of intravenous (IV) methotrexate (MTX) and IV mercaptopurine (MP) is superior to 12 courses of IV MTX alone for prevention of relapse in children with lower-risk B lineage acute lymphoblastic leukemia (ALL). PATIENTS AND METHODS: Six hundred fifty-one eligible patients were entered onto the study. Vincristine, prednisone, and asparaginase were used for remission induction therapy. Patients were randomized to receive intensification with IV MTX 1,000 mg/m(2) plus IV MP 1,000 mg/m(2) (regimen A) or IV MTX 1,000 mg/m(2) alone (regimen C). Twelve courses were administered at 2-week intervals. Triple intrathecal therapy was used for CNS prophylaxis. Continuation therapy included standard oral MP, weekly MTX, and triple intrathecal therapy every 12 weeks for 2 years. RESULTS: Six hundred forty five patients (99.1%) achieved remission. Three hundred twenty-five were assigned to regimen A and 320 to regimen C. The estimated 4-year overall continuous complete remission for patients treated with regimen A is 82.1% (SE = 2.4%) and for regimen C is 82.2% (SE = 2.6%; P =.5). No significant difference in overall outcome was shown by sex or race. Serious grade 3/4 neurotoxicity, principally characterized by seizures, was observed in 7.6% of patients treated with either regimen. CONCLUSION: Intensification with 12 courses of IV MTX is an effective therapy for prevention of relapse in children with B-precursor ALL who are at lower risk for relapse but may be associated with an increased risk for neurotoxicity. Prolonged infusions of MP combined with IV MTX did not provide apparent advantage. PMID- 10715300 TI - CD56 expression is an indicator of poor clinical outcome in patients with acute promyelocytic leukemia treated with simultaneous all-trans-retinoic acid and chemotherapy. AB - PURPOSE: Preliminary reports suggest that leukemic cell expression of CD56, a neural cell adhesion molecule, is associated with adverse clinical outcome in either acute myeloid leukemia with t(8;21) or acute promyelocytic leukemia (APL). We investigated the prognostic relevance of CD56 in a series of patients with APL who were treated homogeneously with all-trans-retinoic acid (ATRA) and chemotherapy. PATIENTS AND METHODS: Clinicobiologic presenting features and therapeutic results were analyzed in a series of 100 patients with genetically proven APL who were treated, according to the example of the Gruppo Italiano Malattie Ematologiche Maligne dell'Adulto multicenter trial, with ATRA plus idarubicin (AIDA) and for whom data on CD56 expression were available at diagnosis. RESULTS: Fifteen patients (15%) showed expression of CD56 in greater than or equal to 20% blasts at diagnosis and were considered as CD56(+). No differences were found regarding age, sex, WBC and platelet counts, incidence of coagulopathy, hemoglobin and fibrinogen levels, promyelocytic leukemia/retinoic acid receptor (PML/RAR) alpha fusion type, or complete remission (CR) rate in the comparison of the CD56(+) and CD56(-) populations. Conversely, compared with patients who were CD56(-), patients with CD56(+) APL had shorter CR duration (P =.04) and overall survival (P =.002). In the multivariate analysis, CD56 positivity and initial WBC count greater than 10 x 10(9) cells/L retained statistical significance in overall survival (P =.04 and P =.02, respectively). CONCLUSION: The expression of CD56 is significantly associated with inferior CR duration and survival in patients with APL who were treated with modern frontline treatment that included ATRA and simultaneous chemotherapy. Combined with other well-established prognostic factors such as WBC count, CD56 expression at diagnosis might be used to build prognostic scores for risk-adapted therapy in APL. PMID- 10715301 TI - Risk factors for severe neuropsychiatric toxicity in patients receiving interferon alfa-2b and low-dose cytarabine for chronic myelogenous leukemia: analysis of Cancer and Leukemia Group B 9013. AB - PURPOSE: Recombinant interferon alfa-2b (rIFNalpha2b) is a standard therapy for chronic myelogenous leukemia (CML). Severe neuropsychiatric toxicity has been described in patients receiving rIFNalpha2b, although the frequency of and the risk factors for developing this toxicity are not well described. The purpose of this study was to identify predictors for the development of severe neuropsychiatric toxicity in CML patients receiving rIFNalpha2b-based therapy. PATIENTS AND METHODS: From a prospective cohort of 91 Philadelphia chromosome positive, previously untreated, chronic-phase CML patients treated on Cancer and Leukemia Group B (CALGB) 9013, a phase II trial of rIFNalpha2b plus cytarabine, the following were recorded at baseline: age, sex, race, pretreatment history of neurologic or psychiatric diagnosis, spleen size, blood counts, and peripheral blast count. Best response to treatment, rIFNalpha2b cumulative dose, dose duration, and dose-intensity were recorded during follow-up. Severe neuropsychiatric toxicity was defined as grade 3 or 4 events, according to CALGB expanded common toxicity criteria. Univariate and multivariate logistic regression analyses were used to identify variables that were associated with the development of severe neuropsychiatric toxicity. RESULTS: Severe neuropsychiatric toxicity developed in 22 patients (24.0%; 95% confidence interval [CI], 15.2% to 32.8%). Toxicity resolved after withdrawal of treatment in all patients. Five of six patients developed recurrence of symptoms with rechallenge. Twelve (63%) of 19 patients with a pretreatment neurologic or psychiatric diagnosis developed severe neuropsychiatric toxicity, as compared with 10 (14%) of 72 patients without a pretreatment neurologic or psychiatric diagnosis (P =.001), resulting in a relative risk of 4. 55 (95% CI, 2.33 to 8.88) for developing severe neuropsychiatric toxicity. No other variables were independently associated with the development of neuropsychiatric toxicity. CONCLUSION: CML patients with a pretreatment history of a neurologic or psychiatric diagnosis are at significantly increased risk of developing severe neuropsychiatric toxicity during therapy with rIFNalpha2b plus cytarabine. Monitoring for neuropsychiatric symptoms and avoiding rechallenge are recommended measures for such patients receiving rIFNalpha2b-based therapy. PMID- 10715302 TI - Randomized comparison of ACVBP and m-BACOD in the treatment of patients with low risk aggressive lymphoma: the LNH87-1 study. Groupe d'Etudes des Lymphomes de l'Adulte. AB - PURPOSE: To compare a short intensified regimen followed by sequential consolidation therapy (doxorubicin, cyclophosphamide, vindesine, bleomycin, and prednisone [ACVBP]) to the standard regimen of methotrexate, bleomycin, cyclophosphamide, and etoposide (m-BACOD) in patients with low-risk aggressive lymphoma. PATIENTS AND METHODS: A total of 752 patients with intermediate- or high-grade lymphoma and no adverse prognostic factors (Eastern Cooperative Oncology Group performance status of 2 to 4, >/= two extranodal sites of disease, tumor burden >/= 10 cm in largest dimension, bone marrow or CNS involvement, Burkitt's or lymphoblastic subtypes) were registered. Of 673 eligible patients, 332 received ACVBP and 341 received m-BACOD. RESULTS: The complete remission rate was identical (86%) in the two groups. With a median follow-up duration of 7 years, the 5-year failure-free survival (FFS) rate was 65% in the ACVBP group and 61% in the m-BACOD group (P =.16). The 5-year overall survival rate was 75% in the ACVBP group and 73% in the m-BACOD group (P =.47). ACVBP was responsible for more severe and life-threatening infections (P <.01), but m-BACOD caused more pulmonary toxicity (P <.001). The number of treatment-related deaths did not differ between the two regimens. A multivariate analysis indicated that ACVBP was associated with a longer FFS in patients with two or three risk factors of the International Prognostic Index. CONCLUSION: In this population of patients with low-risk aggressive lymphoma, toxicities of the regimens are different, but the rates of response and survival are identical. The survival advantage of ACVBP over standard regimen in patients with advanced disease is suggested by this analysis but remains to be assessed in prospective studies specifically designed for this purpose. PMID- 10715303 TI - Multicenter phase II study of iodine-131 tositumomab for chemotherapy relapsed/refractory low-grade and transformed low-grade B-cell non-Hodgkin's lymphomas. AB - PURPOSE: This multicenter phase II study evaluated the efficacy, dosimetry methodology, and safety of iodine-131 tositumomab in patients with chemotherapy relapsed/refractory low-grade or transformed low-grade non-Hodgkin's lymphoma (NHL). PATIENTS AND METHODS: Patients received a dosimetric dose that consisted of 450 mg of anti-B1 antibody followed by 35 mg (5 mCi) of iodine-131 tositumomab. Serial total-body gamma counts were then obtained to calculate the patient-specific millicurie activity required to deliver the therapeutic dose. A therapeutic dose of 75 cGy total-body dose (attenuated to 65 cGy in patients with platelet counts of 101,000 to 149,000 cells/mm(3)) was given 7 to 14 days after the dosimetric dose. RESULTS: Forty-five of 47 patients were treated with a single dosimetric and therapeutic dose. Twenty-seven patients (57%) had a response. The response rate was similar in patients with low-grade (57%) or transformed low-grade (60%) NHL. The median duration of response was 9.9 months. Fifteen patients (32%) achieved a complete response (CR; 10 CRs and five clinical CRs), including five patients (50%) with transformed low-grade NHL. The median duration of CR was 19.9 months, and six patients have an ongoing CR. Treatment was well tolerated, with the principal toxicity being hematologic. The most common nonhematologic toxicities that were considered to be possibly related to the treatment included mild to moderate fatigue (32%), nausea (30%), fever (26%), vomiting (15%), infection (13%), pruritus (13%), and rash (13%). Additionally, one patient developed human-antimouse antibodies. CONCLUSION: Iodine-131 tositumomab produced a high overall response rate, and approximately one third of patients had a CR despite having chemotherapy-relapsed or refractory low-grade or transformed low-grade NHL. PMID- 10715304 TI - Long-term disease-free survivors in metastatic undifferentiated carcinoma of nasopharyngeal type. AB - PURPOSE: To review incidence and analyze profile of long-term complete responders among patients with undifferentiated carcinoma of nasopharyngeal type (UCNT) treated at a single institution. PATIENTS AND METHODS: We present a cohort of 20 long-term unmaintained complete responders to chemotherapy for metastatic UCNT treated at the Institut Gustave Roussy between April 1978 and November 1996. A patient was considered a long-term survivor if he or she was disease-free for more than 36 months without treatment after obtaining a complete response by chemotherapy. Patient characteristics were as follows: sex, 17 men and three women; median age, 28 years (range, 9 to 62 years); median World Health Organization performance status, 1; and initial tumor-node-metastasis stage (International Union Against Cancer-American Joint Committee on Cancer, 1987) of T3 to T4, 60%, and of N2b to N3, 75%. Epstein-Barr virus serology was characteristic in 19 patients. Of 16 pretreated patients, 11 were pretreated by radiotherapy alone and five by chemotherapy and radiotherapy. Thirteen patients had metastatic relapses of locally controlled UCNT. Tumor sites were bone in 15 patients, lung in four, and liver (biopsy-proven) in two. Chemotherapy included the following: cisplatin, bleomycin, and fluorouracil in five patients; bleomycin, epirubicin, and cisplatin in seven patients; fluorouracil, mitomycin, epirubicin, and cisplatin in four patients; and fluorouracil, bleomycin, epirubicin, and cisplatin in one patient. Three patients were treated with platinum-based regimens before 1985. Patients received a median of six cycles (range, three to 13). Thirteen patients with bone metastases received consolidating radiotherapy. RESULTS: As of June 1999, 14 of 20 patients were still alive with no evidence of disease after treatment (disease-free survival time, 82+ to 190+ months), three patients died of other causes while in complete response at 61, 109, and 208 months after treatment, and three patients died of disease at 42, 89, and 115 months after treatment. Long-term complete responses were obtained in both bone and visceral disease. CONCLUSION: Our data support a curative role for chemotherapy in metastatic UCNT and are a major incentive to continue research for better combinations to increase the percentage of patients with metastatic UCNT who attain complete responses and long-term survival. PMID- 10715305 TI - Value of peptide receptor scintigraphy using (123)I-vasoactive intestinal peptide and (111)In-DTPA-D-Phe1-octreotide in 194 carcinoid patients: Vienna University Experience, 1993 to 1998. AB - PURPOSE: To report our experience with both (123)I-vasoactive intestinal peptide (VIP) and (111)In-DTPA-D-Phe(1)-octreotide for imaging to identify primary and metastatic tumor sites in carcinoid patients. PATIENTS AND METHODS: One hundred ninety-four patients with a verified or clinically suspected diagnosis of a carcinoid tumor were injected with (111)In-DTPA-D-Phe(1)-OCT for imaging purposes, while 133 patients underwent scanning with both (123)I-VIP and (111)In DTPA-D-Phe(1)-OCT in random order. Imaging results were compared with computed tomography scans, results of conventional ultrasound, endosonography, and endoscopy, and results of surgical exploration in case of inconclusive conventional imaging. RESULTS: Primary or recurrent carcinoid tumors could be visualized with (111)In-DTPA-D-Phe(1)-OCT in 95 (91%) of 104 patients; metastatic sites were identified in 110 (95%) of 116 patients. In 11 (51%) of 21 patients with suggestive symptoms but without identified lesions by conventional imaging, focal tracer uptake located the carcinoid tumor. In addition, metastatic disease was demonstrated in three patients after resection. In a direct comparison in the 133 patients who underwent both imaging modalities, (111)In-DTPA-D-Phe(1)-OCT was found to be superior to (123)I-VIP, with 35 (93%) of 38 versus 32 (82%) of 38 scans being positive in primary or recurrent tumors, 58 (90%) of 65 versus 53 (82%) of 65 being positive in patients with metastatic sites, and seven (44%) of 16 versus four (25%) of 16 being positive in patients with symptoms but otherwise negative work-ups. Overall, additional lesions not seen on conventional imaging were imaged in 43 (41%) of 158 versus 25 (25%) of 103 scans with (111)In-DTPA-D Phe(1)-OCT and (123)I-VIP, respectively. CONCLUSION: Both peptide tracers have a high sensitivity for localizing tumor sites in patients with ascertained or suspected carcinoid tumors, with (111)In-DTPA-D-Phe(1)-OCT scintigraphy being more sensitive than (123)I-VIP receptor scanning. Both, however, had a higher diagnostic yield than conventional imaging, as verified by surgical intervention or long-term follow-up. The combination of both peptide receptor scans does not seem to further enhance diagnostic information. PMID- 10715306 TI - Capecitabine, an oral fluoropyrimidine carbamate with substantial activity in advanced colorectal cancer: results of a randomized phase II study. AB - PURPOSE: To evaluate in patients with advanced colorectal cancer (CRC) three treatment regimens of oral capecitabine in order to select the most appropriate regimen for testing in phase III. PATIENTS AND METHODS: Three capecitabine schedules were evaluated in a randomized phase II design: arm A, 1,331 mg/m(2)/d bid continuously; arm B, 2,510 mg/m(2)/d bid intermittently (2 weeks on/1 week off); and arm C, 1,657 mg/m(2)/d plus oral leucovorin 60 mg/d bid intermittently (2 weeks on/1 week off). RESULTS: One hundred nine patients were randomized; 39 patients were assessable for efficacy in arm A, 34 in arm B, and 35 in arm C. Patient characteristics were balanced in the arms. Confirmed tumor responses (partial response [PR] + complete response [CR]) were reported for eight patients with two CRs (21%; 95% confidence interval [CI], 9% to 36%) in arm A, eight patients with one CR (24%; 95% CI, 11% to 41%) in arm B, and eight patients with two CRs (23%; 95% CI, 10% to 40%) in arm C. Median times to progression (TTP) in arms A, B, and C were 127, 230, and 165 days, respectively. Overall, more toxicity was seen with capecitabine plus leucovorin, particularly diarrhea and hand-foot syndrome. There was no grade 3 or 4 marrow toxicity. CONCLUSION: Capecitabine offers a new, effective treatment option as an oral single agent in advanced CRC. Promising overall response rates were reported for all three regimens. The addition of leucovorin to the intermittent regimen had no marked effect on tumor response or median TTP. The intermittent single-agent capecitabine schedule is proposed for phase III evaluation, based on considerations of toxicity, dose-intensity, response rate, and TTP. PMID- 10715307 TI - Phase II trial of docetaxel and vinorelbine in patients with advanced non-small cell lung cancer. AB - PURPOSE: Docetaxel and vinorelbine are active agents in advanced non-small-cell lung cancer (NSCLC) and demonstrate preclinical synergism perhaps, in part, through their inactivation of the proto-oncogene bcl-2. We show that docetaxel (60 mg/m(2)) and vinorelbine (45 mg/m(2)) can be safely combined when given on an every 2-week schedule with filgrastim, with encouraging antitumor activity observed. PATIENTS AND METHODS: Thirty-five chemotherapy naive patients with advanced NSCLC received vinorelbine as an intravenous push immediately followed by docetaxel as a 1-hour intravenous infusion once every 2 weeks. Prophylactic corticosteroids, ciprofloxacin, and filgrastim were used. RESULTS: We delivered median doses of 450 mg/m(2) of vinorelbine and 600 mg/m(2) of docetaxel. The major objective response rate was 51% (95% confidence interval [CI], 34% to 68%). With a median follow-up of 14 months, the predicted median survival time was 14 months, and the 1-year survival rate was 60% (95% CI, 44% to 80%). Febrile neutropenia occurred in five patients and five (1.3%) of 384 treatments. No dose limiting neurotoxicity occurred. Symptomatic onycholysis and excessive lacrimation were observed after several months or more of therapy. CONCLUSION: Docetaxel 60 mg/m(2) and vinorelbine 45 mg/m(2), both given every 2 weeks, is a highly active combination for the treatment of advanced NSCLC. Filgrastim largely obviates neutropenic fever and allows for the single-agent dose-intensity of both drugs to be delivered. The occurrence of certain late toxicities can limit use in some cases and suggests that the combination could also be beneficial in settings requiring briefer, fixed periods of treatment, such as in induction or postoperative therapy. PMID- 10715309 TI - Therapeutic relevance of CD34 cell dose in blood cell transplantation for cancer therapy. AB - PURPOSE: To review recent advances in peripheral-blood progenitor-cell (PBPC) transplantation in order to define the optimal cell dose required for autologous and allogeneic transplantation. MATERIALS AND METHODS: A search of MEDLINE was conducted to identify relevant publications. Their bibliographies were also used to identify further articles and abstracts for critical review. RESULTS: The CD34(+) cell content of a graft is regarded as an accurate predictor of engraftment success. Postchemotherapy autologous PBPC transplantation with >/= 5 x 10(6) CD34(+) cells/kg body weight leads to more rapid engraftment than does transplantation of lower cell doses. Further increases in transplant cell dose further accelerate platelet but not neutrophil engraftment. Evidence that long term hematopoietic recovery may be more accurately predicted by the subpopulation of primitive progenitors transplanted suggests that the content of CD34(+)CD33(-) and long-term culture-initiating cells in cell collection samples may be important for predicting successful engraftment, particularly in patients with poor mobilization. Allogeneic transplantation has been limited by concerns regarding graft-versus-host disease and the use of hematopoietic growth factors in donors. The risk of graft rejection and engraftment failure after HLA mismatched allogeneic transplantation may be overcome by intensive chemoradiotherapy and the infusion of large numbers of T cell-depleted hematopoietic stem cells. CONCLUSION: An optimal cell dose of >/= 8 x 10(6) CD34(+) cells/kg seems to be recommended for autologous PBPC transplantation. This dose facilitates the administration of scheduled chemotherapy on time and reduces the demand for other supportive therapies. A combination of growth factors may enable patients with poor mobilization to achieve a collection sufficient to allow transplantation. The optimum PBPC dose for allogeneic transplantation remains to be defined. PMID- 10715308 TI - Tirapazamine plus cisplatin versus cisplatin in advanced non-small-cell lung cancer: A report of the international CATAPULT I study group. Cisplatin and Tirapazamine in Subjects with Advanced Previously Untreated Non-Small-Cell Lung Tumors. AB - PURPOSE: A phase III trial, Cisplatin and Tirapazamine in Subjects with Advanced Previously Untreated Non-Small-Cell Lung Tumors (CATAPULT I), was designed to determine the efficacy and safety of tirapazamine plus cisplatin for the treatment of non-small-cell lung cancer (NSCLC). PATIENTS AND METHODS: Patients with previously untreated NSCLC were randomized to receive either tirapazamine (390 mg/m(2) infused over 2 hours) followed 1 hour later by cisplatin (75 mg/m(2) over 1 hour) or 75 mg/m(2) of cisplatin alone, every 3 weeks for a maximum of eight cycles. RESULTS: A total of 446 patients with NSCLC (17% with stage IIIB disease and pleural effusions; 83% with stage IV disease) were entered onto the study. Karnofsky performance status (KPS) was >/= 60 for all patients (for 10%, KPS = 60; for 90%, KPS = 70 to 100). Sixty patients (14%) had clinically stable brain metastases. The median survival was significantly longer (34.6 v 27. 7 weeks; P =.0078) and the response rate was significantly greater (27.5% v 13.7%; P <.001) for patients who received tirapazamine plus cisplatin (n = 218) than for those who received cisplatin alone (n = 219). The tirapazamine-plus-cisplatin regimen was associated with mild to moderate adverse events, including acute, reversible hearing loss, reversible, intermittent muscle cramping, diarrhea, skin rash, nausea, and vomiting. There were no incremental increases in myelosuppression, peripheral neuropathy, or renal, hepatic, or cardiac toxicity and no deaths related to tirapazamine. CONCLUSION: The CATAPULT I study shows that tirapazamine enhances the activity of cisplatin in patients with advanced NSCLC and confirms that hypoxia is an exploitable therapeutic target in human malignancies. PMID- 10715310 TI - American Society of Clinical Oncology guideline on the role of bisphosphonates in breast cancer. American Society of Clinical Oncology Bisphosphonates Expert Panel. AB - PURPOSE: To determine clinical practice guidelines for the use of bisphosphonates in the prevention and treatment of bone metastases in breast cancer and their role relative to other therapies for this condition. METHODS: An expert multidisciplinary panel reviewed pertinent information from the published literature and meeting abstracts through May 1999. Additional data collected as part of randomized trials and submitted to the United States Food and Drug Administration were also reviewed, and investigators were contacted for more recent information. Values for levels of evidence and grade of recommendation were assigned by expert reviewers and approved by the panel. Expert consensus was used if there were insufficient published data. The panel addressed which patients to treat and when in their course of disease, specific drug delivery issues, duration of therapy, management of bony metastases with other therapies, and the public policy implications. The guideline underwent external review by selected physicians, members of the American Society of Clinical Oncology (ASCO) Health Services Research Committee, and the ASCO Board of Directors. RESULTS: Bisphosphonates have not had an impact on the most reliable cancer end point: overall survival. The benefits have been reductions in skeletal complications, ie, pathologic fractures, surgery for fracture or impending fracture, radiation, spinal cord compression, and hypercalcemia. Intravenous (IV) pamidronate 90 mg delivered over 1 to 2 hours every 3 to 4 weeks is recommended in patients with metastatic breast cancer who have imaging evidence of lytic destruction of bone and who are concurrently receiving systemic therapy with hormonal therapy or chemotherapy. For women with only an abnormal bone scan but without bony destruction by imaging studies or localized pain, there is insufficient evidence to suggest starting bisphosphonates. Starting bisphosphonates in patients without evidence of bony metastasis, even in the presence of other extraskeletal metastases, is not recommended. Studies of bisphosphonates in the adjuvant setting have yielded inconsistent results. Starting bisphosphonates in patients at any stage of their nonosseous disease, outside of clinical trials, despite a high risk for future bone metastasis, is currently not recommended. Oral bisphosphonates are one of several options which can be used for preservation of bone density in premenopausal patients with treatment-induced menopause. The panel suggests that, once initiated, IV bisphosphonates be continued until evidence of substantial decline in a patient's general performance status. The panel stresses that clinical judgment must guide what is a substantial decline. There is no evidence addressing the consequences of stopping bisphosphonates after one or more adverse skeletal events. Symptoms in the spine, pelvis, or femur require careful evaluation for spinal cord compression and pathologic fracture before bisphosphonate use and if symptoms recur, persist, or worsen during therapy. The panel recommends that current standards of care for cancer pain, analgesics and local radiation therapy, not be displaced by bisphosphonates. IV pamidronate is recommended in women with pain caused by osteolytic metastasis to relieve pain when used concurrently with systemic chemotherapy and/or hormonal therapy, since it was associated with a modest pain control benefit in controlled trials. CONCLUSION: Bisphosphonates provide a meaningful supportive but not life-prolonging benefit to many patients with bone metastases from cancer. Further research is warranted to identify clinical predictors of when to start and stop therapy, to integrate their use with other treatments for bone metastases, to identify their role in the adjuvant setting in preventing bone metastases, and to better determine their cost-benefit consequences. PMID- 10715311 TI - Chemotherapy with and without anthracycline. PMID- 10715312 TI - Docetaxel, cisplatin, fluorouracil, and leucovorin in locally advanced head and neck cancer. PMID- 10715313 TI - Expression of Ikaros isoforms in acute lymphoblastic leukemia cell lines. PMID- 10715314 TI - Proteomics: broad strokes of expressionism? PMID- 10715315 TI - Self-incompatibility in Brassica: the elusive pollen S gene is identified! PMID- 10715316 TI - How can two-gene models of self-incompatibility generate new specificities? PMID- 10715317 TI - Evolutionary dynamics of dual-specificity self-incompatibility alleles. PMID- 10715318 TI - Reply. Establishing A paradigm for the generation of new s alleles PMID- 10715319 TI - INTERFASCICULAR FIBERLESS1 is the same gene as REVOLUTA. PMID- 10715320 TI - Proteomics of the chloroplast: systematic identification and targeting analysis of lumenal and peripheral thylakoid proteins. AB - The soluble and peripheral proteins in the thylakoids of pea were systematically analyzed by using two-dimensional electrophoresis, mass spectrometry, and N terminal Edman sequencing, followed by database searching. After correcting to eliminate possible isoforms and post-translational modifications, we estimated that there are at least 200 to 230 different lumenal and peripheral proteins. Sixty-one proteins were identified; for 33 of these proteins, a clear function or functional domain could be identified, whereas for 10 proteins, no function could be assigned. For 18 proteins, no expressed sequence tag or full-length gene could be identified in the databases, despite experimental determination of a significant amount of amino acid sequence. Nine previously unidentified proteins with lumenal transit peptides are presented along with their full-length genes; seven of these proteins possess the twin arginine motif that is characteristic for substrates of the TAT pathway. Logoplots were used to provide a detailed analysis of the lumenal targeting signals, and all nuclear-encoded proteins identified on the two-dimensional gels were used to test predictions for chloroplast localization and transit peptides made by the software programs ChloroP, PSORT, and SignalP. A combination of these three programs was found to provide a useful tool for evaluating chloroplast localization and transit peptides and also could reveal possible alternative processing sites and dual targeting. The potential of proteomics for plant biology and homology-based searching with mass spectrometry data is discussed. PMID- 10715321 TI - A conserved domain of the arabidopsis GNOM protein mediates subunit interaction and cyclophilin 5 binding. AB - The Arabidopsis GNOM protein, a guanine nucleotide exchange factor (GEF) that acts on ADP ribosylation factor (ARF)-type G proteins, is required for coordination of cell polarity along the apical-basal embryo axis. Interallelic complementation of gnom mutants suggested that dimerization is involved in GNOM function. Here, direct interaction between GNOM molecules is demonstrated in vitro and by using a yeast two-hybrid system. Interaction was confined to an N terminal domain conserved within a subgroup of large ARF GEFs. The same domain mediated in vitro binding to cyclophilin 5 (Cyp5), which was identified as a GNOM interactor in two-hybrid screening. Cyp5 displayed peptidylprolyl cis/trans isomerase and protein refolding activities that were sensitive to cyclosporin A. Cyp5 protein accumulated in several plant organs and, like GNOM, was partitioned between cytosolic and membrane fractions. Cyp5 protein was also expressed in the developing embryo. Our results suggest that Cyp5 may regulate the ARF GEF function of the GNOM protein during embryogenesis. PMID- 10715322 TI - Silencing of retrotransposons in arabidopsis and reactivation by the ddm1 mutation. AB - Gene silencing associated with repeated DNA sequences has been reported for many eukaryotes, including plants. However, its biological significance remains to be determined. One important function that has been proposed is the suppression of transposons. Here, we address transposon suppression by examining the behavior of the tobacco retrotransposon Tto1 and endogenous retrotransposons in Arabidopsis. After an initial increase in copy number because of active transposition in the Arabidopsis genome, Tto1 became silent. The amount of transcript was reduced, and the inactivated Tto1 became methylated. This silencing correlated with an increase in copy number. These phenomena mimic repeat-induced gene silencing. The homozygous ddm1 (for decrease in DNA methylation) mutation of Arabidopsis results in genomic DNA hypomethylation and the release of silencing in repeated genes. To investigate the role of DNA methylation and the gene-silencing machinery in the suppression of Tto1, we introduced the ddm1 mutation into an Arabidopsis line carrying inactivated Tto1 copies. In the homozygous ddm1 background, Tto1 became hypomethylated and transcriptionally and transpositionally active. In addition, one of the newly isolated endogenous Arabidopsis retrotransposon families, named Tar17, also became hypomethylated and transcriptionally active in the ddm1 mutant background. Our results suggest that the inactivation of retrotransposons and the silencing of repeated genes have mechanisms in common. PMID- 10715323 TI - Potato virus X amplicons in arabidopsis mediate genetic and epigenetic gene silencing. AB - Amplicon transgenes from potato virus X (PVX) are based on a modified version of the viral genome and are efficient activators of post-transcriptional gene silencing (PTGS). To determine whether PVX amplicons activate PTGS in Arabidopsis, we used constructs based on the genome of PVX carrying a green fluorescent protein (GFP) reporter gene. Our analysis of the transgene phenotype exploited previous observations indicating that PTGS is associated with short 25 nucleotide RNA species, transgene methylation, and homology-dependent virus resistance. We also used the ability of turnip mosaic virus to suppress gene silencing as a means of dissecting stages of the mechanism. The results showed that a PVX:GFP amplicon induces weak PTGS and that this PTGS was enhanced in the presence of a GFP reporter gene. Our interpretation of these data is that the PTGS induced by the amplicon was genetically determined and equivalent to the initiation stage of the PTGS mechanism. The PTGS induced by the combined amplicon and reporter gene was equivalent to the maintenance stage and was associated with an epigenetic conversion of the transgene. The distinction between genetic and epigenetic PTGS explains the well-characterized effects of transgene dosage on PTGS that have been previously interpreted in terms of RNA expression thresholds. PMID- 10715324 TI - The complete sequence of 340 kb of DNA around the rice Adh1-adh2 region reveals interrupted colinearity with maize chromosome 4. AB - A 2.3-centimorgan (cM) segment of rice chromosome 11 consisting of 340 kb of DNA sequence around the alcohol dehydrogenase Adh1 and Adh2 loci was completely sequenced, revealing the presence of 33 putative genes, including several apparently involved in disease resistance. Fourteen of the genes were confirmed by identifying the corresponding transcripts. Five genes, spanning 1.9 cM of the region, cross-hybridized with maize genomic DNA and were genetically mapped in maize, revealing a stretch of colinearity with maize chromosome 4. The Adh1 gene marked one significant interruption. This gene mapped to maize chromosome 1, indicating a possible translocation of Adh1 after the evolutionary divergence leading to maize and sorghum. Several other genes, most notably genes similar to known disease resistance genes, showed no cross-hybridization with maize genomic DNA, suggesting sequence divergence or absence of these sequences in maize, which is in contrast to several other well-conserved genes, including Adh1 and Adh2. These findings indicate that the use of rice as the model system for other cereals may sometimes be complicated by the presence of rapidly evolving gene families and microtranslocations. Seven retrotransposons and eight transposons were identified in this rice segment, including a Tc1/Mariner-like element, which is new to rice. In contrast to maize, retroelements are less frequent in rice. Only 14.4% of this genome segment consist of retroelements. Miniature inverted repeat transposable elements were found to be the most frequently occurring class of repetitive elements, accounting for 18.8% of the total repetitive DNA. PMID- 10715325 TI - Arabidopsis ethylene-responsive element binding factors act as transcriptional activators or repressors of GCC box-mediated gene expression. AB - Ethylene-responsive element binding factors (ERFs) are members of a novel family of transcription factors that are specific to plants. A highly conserved DNA binding domain known as the ERF domain is the unique feature of this protein family. To characterize in detail this family of transcription factors, we isolated Arabidopsis cDNAs encoding five different ERF proteins (AtERF1 to AtERF5) and analyzed their structure, DNA binding preference, transactivation ability, and mRNA expression profiles. The isolated AtERFs were placed into three classes based on amino acid identity within the ERF domain, although all five displayed GCC box-specific binding activity. AtERF1, AtERF2, and AtERF5 functioned as activators of GCC box-dependent transcription in Arabidopsis leaves. By contrast, AtERF3 and AtERF4 acted as repressors that downregulated not only basal transcription levels of a reporter gene but also the transactivation activity of other transcription factors. The AtERF genes were differentially regulated by ethylene and by abiotic stress conditions, such as wounding, cold, high salinity, or drought, via ETHYLENE-INSENSITIVE2 (EIN2)-dependent or independent pathways. Cycloheximide, a protein synthesis inhibitor, also induced marked accumulation of AtERF mRNAs. Thus, we conclude that AtERFs are factors that respond to extracellular signals to modulate GCC box-mediated gene expression positively or negatively. PMID- 10715326 TI - Deficiency in fatty acid synthase leads to premature cell death and dramatic alterations in plant morphology. AB - An Arabidopsis mosaic death1 (mod1) mutant, which has premature cell death in multiple organs, was isolated. mod1 plants display multiple morphological phenotypes, including chlorotic and curly leaves, distorted siliques, premature senescence of primary inflorescences, reduced fertility, and semidwarfism. The phenotype of the mod1 mutant results from a single nuclear recessive mutation, and the MOD1 gene was isolated by using a map-based cloning approach. The MOD1 gene encodes an enoyl-acyl carrier protein (ACP) reductase, which is a subunit of the fatty acid synthase complex that catalyzes de novo synthesis of fatty acids. An amino acid substitution in the enoyl-ACP reductase of the mod1 mutant causes a marked decrease in its enzymatic activity, impairing fatty acid biosynthesis and decreasing the amount of total lipids in mod1 plants. These results demonstrate that a deficiency in fatty acid biosynthesis has pleiotropic effects on plant growth and development and causes premature cell death. PMID- 10715327 TI - The thylakoid FtsH protease plays a role in the light-induced turnover of the photosystem II D1 protein. AB - The photosystem II reaction center D1 protein is known to turn over frequently. This protein is prone to irreversible damage caused by reactive oxygen species that are formed in the light; the damaged, nonfunctional D1 protein is degraded and replaced by a new copy. However, the proteases responsible for D1 protein degradation remain unknown. In this study, we investigate the possible role of the FtsH protease, an ATP-dependent zinc metalloprotease, during this process. The primary light-induced cleavage product of the D1 protein, a 23-kD fragment, was found to be degraded in isolated thylakoids in the dark during a process dependent on ATP hydrolysis and divalent metal ions, suggesting the involvement of FtsH. Purified FtsH degraded the 23-kD D1 fragment present in isolated photosystem II core complexes, as well as that in thylakoid membranes depleted of endogenous FtsH. In this study, we definitively identify the chloroplast protease acting on the D1 protein during its light-induced turnover. Unlike previously identified membrane-bound substrates for FtsH in bacteria and mitochondria, the 23-kD D1 fragment represents a novel class of FtsH substrate-functionally assembled proteins that have undergone irreversible photooxidative damage and cleavage. PMID- 10715328 TI - Nuclear gamma-tubulin during acentriolar plant mitosis. AB - Neither the molecular mechanism by which plant microtubules nucleate in the cytoplasm nor the organization of plant mitotic spindles, which lack centrosomes, is well understood. Here, using immunolocalization and cell fractionation techniques, we provide evidence that gamma-tubulin, a universal component of microtubule organizing centers, is present in both the cytoplasm and the nucleus of plant cells. The amount of gamma-tubulin in nuclei increased during the G(2) phase, when cells are synchronized or sorted for particular phases of the cell cycle. gamma-Tubulin appeared on prekinetochores before preprophase arrest caused by inhibition of the cyclin-dependent kinase and before prekinetochore labeling of the mitosis-specific phosphoepitope MPM2. The association of nuclear gamma tubulin with chromatin displayed moderately strong affinity, as shown by its release after DNase treatment and by using extraction experiments. Subcellular compartmentalization of gamma-tubulin might be an important factor in the organization of plant-specific microtubule arrays and acentriolar mitotic spindles. PMID- 10715330 TI - Special communication (1): the RSNA scientific program: 2000 PMID- 10715331 TI - Practice corner-(1): optimal imaging coverage: local or from afar? PMID- 10715329 TI - A strong loss-of-function mutation in RAN1 results in constitutive activation of the ethylene response pathway as well as a rosette-lethal phenotype. AB - A recessive mutation was identified that constitutively activated the ethylene response pathway in Arabidopsis and resulted in a rosette-lethal phenotype. Positional cloning of the gene corresponding to this mutation revealed that it was allelic to responsive to antagonist1 (ran1), a mutation that causes seedlings to respond in a positive manner to what is normally a competitive inhibitor of ethylene binding. In contrast to the previously identified ran1-1 and ran1-2 alleles that are morphologically indistinguishable from wild-type plants, this ran1-3 allele results in a rosette-lethal phenotype. The predicted protein encoded by the RAN1 gene is similar to the Wilson and Menkes disease proteins and yeast Ccc2 protein, which are integral membrane cation-transporting P-type ATPases involved in copper trafficking. Genetic epistasis analysis indicated that RAN1 acts upstream of mutations in the ethylene receptor gene family. However, the rosette-lethal phenotype of ran1-3 was not suppressed by ethylene-insensitive mutants, suggesting that this mutation also affects a non-ethylene-dependent pathway regulating cell expansion. The phenotype of ran1-3 mutants is similar to loss-of-function ethylene receptor mutants, suggesting that RAN1 may be required to form functional ethylene receptors. Furthermore, these results suggest that copper is required not only for ethylene binding but also for the signaling function of the ethylene receptors. PMID- 10715332 TI - Resolving breast microcalcifications. PMID- 10715333 TI - Pitfalls in the diagnosis of thoracic aortic dissection at CT angiography. AB - Two hundred seventy-five computed tomographic (CT) angiograms of the thoracic aorta were obtained over a period of approximately 4 years in patients with suspected or known aortic dissection. In all cases, unenhanced images were initially obtained, followed by contrast material-enhanced images. A variety of pitfalls were encountered that mimicked aortic dissection. These pitfalls were attributable to technical factors (eg, improper timing of contrast material administration relative to image acquisition); streak artifacts generated by high attenuation material, high-contrast interfaces, or cardiac motion; periaortic structures (eg, aortic arch branches, mediastinal veins, pericardial recess, thymus, atelectasis, pleural thickening or effusion adjacent to the aorta); aortic wall motion and normal aortic sinuses; aortic variations such as congenital ductus diverticulum and acquired aortic aneurysm with thrombus; and penetrating atherosclerotic ulcer. Although several of these pitfalls are easy to recognize and therefore unlikely to present a diagnostic problem, others are potentially confusing. Familiarity with these common pitfalls, coupled with a knowledge of normal intrathoracic anatomy, will facilitate recognition of true aortic dissection and help avoid misdiagnosis at thoracic aortic CT angiography. PMID- 10715334 TI - Congenital tarsal coalition: multimodality evaluation with emphasis on CT and MR imaging. AB - Congenital tarsal coalition is a diagnosis that is often overlooked in young patients who first present with foot and ankle pain. Calcaneonavicular and talocalcaneal coalitions are encountered most frequently; fusion at other sites is much less common. Tarsal coalitions may be osseous, cartilaginous, or fibrous. Calcaneonavicular coalitions are readily detected on oblique radiographs. Radiographic confirmation of talocalcaneal coalition is more difficult than for fusion at other locations, although several secondary radiographic signs may indirectly suggest the diagnosis. Computed tomography (CT) and magnetic resonance (MR) imaging are invaluable for assessment of tarsal coalitions because they allow differentiation of osseous from nonosseous coalitions and because they depict the extent of joint involvement as well as secondary degenerative changes, features of vital importance in surgical planning. Short-inversion-time inversion recovery MR images may reveal bone marrow edema along the margins of the abnormal articulation, an important clue to the diagnosis. Moreover, CT or MR imaging may be required to confirm the diagnosis of talocalcaneal coalition when radiographic findings are equivocal. Because the diagnosis of tarsal coalition is often not entertained by the clinician ordering a CT or MR imaging examination, multiplanar imaging of the ankle and hindfoot is required. PMID- 10715335 TI - Painful heel: MR imaging findings. AB - Heel pain is a common and frequently disabling clinical complaint that may be caused by a broad spectrum of osseous or soft-tissue disorders. These disorders are classified on the basis of anatomic origin and predominant location of heel pain to foster a better understanding of this complaint. The disorders include plantar fascial lesions (fasciitis, rupture, fibromatosis, xanthoma), tendinous lesions (tendinitis, tenosynovitis), osseous lesions (fractures, bone bruises, osteomyelitis, tumors), bursal lesions (retrocalcaneal bursitis, retroachilleal bursitis), tarsal tunnel syndrome, and heel plantar fat pad abnormalities. With its superior soft-tissue contrast resolution and multiplanar capability, magnetic resonance (MR) imaging can help determine the cause of heel pain and help assess the extent and severity of the disease in ambiguous or clinically equivocal cases. Careful analysis of MR imaging findings and correlation of these findings with patient history and findings at physical examination can suggest a specific diagnosis in most cases. The majority of patients with heel pain can be successfully treated conservatively, but in cases requiring surgery (eg, plantar fascia rupture in competitive athletes, deeply infiltrating plantar fibromatosis, masses causing tarsal tunnel syndrome), MR imaging is especially useful in planning surgical treatment by showing the exact location and extent of the lesion. PMID- 10715336 TI - Biliary obstruction: findings at MR cholangiography and cross-sectional MR imaging. AB - Twenty-two patients with malignant biliary obstruction and 21 patients with suspected obstruction of biliary-enteric anastomoses were evaluated over a 12 month period with magnetic resonance (MR) cholangiography and cross-sectional MR imaging. In patients with malignant obstruction, MR cholangiography helped accurately determine the status of the biliary ductal system by identifying the exact location and extent of the obstruction and the severity of duct dilatation. In so doing, MR cholangiography helped determine whether percutaneous transhepatic cholangiography with antegrade stent placement or retrograde cholangiography with stent placement constituted the more suitable treatment. Cross-sectional MR imaging was necessary to identify the organ of tumor origin, define the tumor margins, and determine the stage of disease. This information helped evaluate the appropriateness of curative surgical therapy versus palliative drainage procedures. In patients with biliary-enteric anastomoses, MR cholangiography clearly depicted the site of the anastomosis and demonstrated the status of the intrahepatic ducts, thereby helping determine which patients would benefit from undergoing antegrade duct cannulation with a drainage procedure or perhaps balloon dilation. In some of these patients, MR cholangiography was sufficient to help plan therapeutic intervention. MR cholangiography also demonstrates the presence and size of biliary stones and associated findings such as intraductal tumor growth. In addition, MR cholangiography may obviate retrograde cholangiography, which can be technically difficult to perform. PMID- 10715337 TI - Imaging of nontraumatic hemorrhagic hepatic lesions. AB - Spontaneous hepatic bleeding is a rare condition. In the absence of trauma or anticoagulant therapy, hepatic hemorrhage may be due to underlying liver disease. The most common causes of nontraumatic hepatic hemorrhage are hepatocellular carcinoma and hepatic adenoma. Such hemorrhage can also occur in patients with other liver tumors, such as focal nodular hyperplasia, hemangiomas, and metastases. Other conditions associated with this entity include HELLP syndrome, amyloidosis, and miscellaneous causes. Imaging plays a significant role in the diagnosis and management of this potentially lethal entity. In the appropriate clinical setting, the diagnosis of a hemorrhagic liver lesion is suggested when a hyperechoic mass or a mass with hyperechoic areas is seen at ultrasonography, a hyperattenuating mass is seen at computed tomography (CT), or a mass with high signal-intensity areas is seen at T1-weighted magnetic resonance (MR) imaging. The signal intensity of blood can be increased or decreased on MR images depending on when the hemorrhage is imaged. The presence and extent of commonly associated subcapsular hematomas and hemoperitoneum can be easily ascertained with CT. During the first 24-72 hours, acute hematomas are hyperattenuating on nonenhanced CT scans; later, they decrease in attenuation and sometimes develop a pseudocapsule. PMID- 10715338 TI - Imaging of atypical hemangiomas of the liver with pathologic correlation. AB - Compared with the imaging features of typical hepatic hemangiomas, the imaging features of atypical hepatic hemangiomas have not been well studied or well described. Knowledge of the entire spectrum of atypical hepatic hemangiomas is important and can help one avoid most diagnostic errors. A frequent type of atypical hepatic hemangioma is a lesion with an echoic border at ultrasonography. Less frequent types are large, heterogeneous hemangiomas; rapidly filling hemangiomas; calcified hemangiomas; hyalinized hemangiomas; cystic or multilocular hemangiomas; hemangiomas with fluid-fluid levels; and pedunculated hemangiomas. Adjacent abnormalities consist of arterial-portal venous shunt, capsular retraction, and surrounding nodular hyperplasia; hemangiomas can also develop in cases of fatty liver infiltration. Associated lesions include multiple hemangiomas, hemangiomatosis, focal nodular hyperplasia, and angiosarcoma. Types of atypical evolution are hemangiomas enlarging over time and hemangiomas appearing during pregnancy. Complications consist of inflammation, Kasabach Merritt syndrome, intratumoral hemorrhage, hemoperitoneum, volvulus, and compression of adjacent structures. In some cases, such as large heterogeneous hemangiomas, calcified hemangiomas, pedunculated hemangiomas, or hemangiomas developing in diffuse fatty liver, a specific diagnosis can be established with imaging, especially magnetic resonance imaging. However, in other atypical cases, the diagnosis will remain uncertain at imaging, and these cases will require histopathologic examination. PMID- 10715339 TI - CT evaluation of the colon: inflammatory disease. AB - Computed tomography (CT) is valuable for detection and characterization of many inflammatory conditions of the colon. At CT, a dilated, thickened appendix is suggestive of appendicitis. A 1-4-cm, oval, fatty pericolic lesion with surrounding mesenteric inflammation is diagnostic of epiploic appendagitis. The key to distinguishing diverticulitis from other inflammatory conditions of the colon is the presence of diverticula in the involved segment. In typhlitis, CT demonstrates cecal distention and circumferential thickening of the cecal wall, which may have low attenuation secondary to edema. In radiation colitis, the clinical history is the key to suggesting the diagnosis because the CT findings can be nonspecific. The location of the involved segment and the extent and appearance of wall thickening may help distinguish Crohn disease and ulcerative colitis. In ischemic colitis, CT typically demonstrates circumferential, symmetric wall thickening with fold enlargement. CT findings of graft-versus-host disease include small bowel and colonic wall thickening, which may result in luminal narrowing and separation of bowel loops. In infectious colitis, the site and thickness of colon affected may suggest a specific organism. The amount of wall thickening in pseudomembranous colitis is typically greater than in any other inflammatory disease of the colon except Crohn disease. PMID- 10715340 TI - Spiral CT of colon cancer: imaging features and role in management. AB - Colorectal cancer is a common malignancy that results in significant morbidity and mortality. Abdominal computed tomography (CT) is valuable in planning surgery for colon cancer because it can demonstrate regional extension of tumor as well as adenopathy and distant metastases. At CT, colorectal cancer typically appears as a discrete soft-tissue mass that narrows the colonic lumen. Colorectal cancer can also manifest as focal colonic wall thickening and luminal narrowing. Complications of primary colonic malignancies such as obstruction, perforation, and fistula can be readily visualized with CT. At CT, local extension of tumor appears as an extracolic mass or simply as thickening and infiltration of pericolic fat. Extracolic spread is also suggested by loss of fat planes between the colon and adjacent organs. The liver is the predominant organ to be involved with metastases from colorectal cancer. At CT, hepatic metastases usually appear as hypoattenuating masses, which are best visualized during the portal venous phase of liver enhancement. Other common sites of metastases from colon cancer include the lungs, adrenal glands, and bones. Use of CT is critical for identifying recurrences, evaluating anatomic relationships, documenting "normal" postoperative anatomy, and confirming the absence of new lesions during and after therapy. PMID- 10715341 TI - Preoperative use of 3D volume rendering to demonstrate renal tumors and renal anatomy. AB - With increased use of computed tomography (CT) and abdominal ultrasonography, the indications for nephron-sparing surgery are also increasing. Triphasic helical CT and three-dimensional (3D) volume rendering can be combined into a single noninvasive test to delineate renal tumors and normal and complex renal anatomy prior to nephron-sparing surgery. This combination technique has proved accurate and very useful for both preoperative and intraoperative planning by demonstrating renal position, tumor location and depth of tumor extension into the kidney, relationship of the tumor to the collecting system, and renal vascular anatomy. Knowledge of the position of the kidney relative to the lower rib cage, iliac crest, and spine helps in planning the initial surgical incision. By depicting tumor location and depth of extension, helical CT with 3D volume rendering helps ensure complete tumor excision and conservation of adjacent normal renal parenchyma. Depiction of the relationship of the tumor to the collecting system helps anticipate further tumor extension and minimize postoperative complications. Identification of normal renal vasculature and anatomic variants can help minimize ischemic injury and intraoperative bleeding. Radiologists should be familiar with current indications for nephron-sparing surgery and understand what information is required prior to surgery. PMID- 10715342 TI - Pearls and pitfalls in the diagnosis of ureterolithiasis with unenhanced helical CT. AB - Several signs to assist interpretation of unenhanced helical computed tomographic (CT) scans obtained for suspected ureterolithiasis have been described. Because signs such as perinephric stranding are not always readily apparent, a methodical approach to interpretation of CT studies is important in determining the presence or absence of ureterolithiasis. Evaluation of the poles of the kidneys is helpful in detecting subtle stranding of the perinephric fat. Inspection of the intrarenal collecting system within the poles of the kidneys is helpful in identifying subtle collecting system dilatation and can help prevent mistaking an extrarenal pelvis for hydronephrosis. Careful inspection of the ureter throughout its course is the most reliable method of distinguishing between ureteral stones and phleboliths. However, when the ureter cannot be followed antegrade, the pelvic portion can often be identified in a retrograde fashion. When secondary signs of obstruction are present but no stone is present, differential diagnostic considerations include a recently passed stone, pyelonephritis, urinary tract obstruction unrelated to stone disease, and protease inhibitor deposition disease. PMID- 10715345 TI - Educational materials review : contrast ultrasound for the radiologist PMID- 10715344 TI - Imaging of extrapulmonary tuberculosis. AB - Diagnosis of extrapulmonary tuberculosis is often difficult. Although positive chest radiographic findings or a positive tuberculin skin test supports the diagnosis, negative results do not exclude extrapulmonary tuberculosis. However, recognition and understanding of the radiologic findings of extrapulmonary tuberculosis can help in diagnosis. The spine is the most common site of skeletal involvement. The femur, tibia, and small bones of the hands and feet are most commonly involved by tuberculous osteomyelitis. Tuberculosis of the joints is characteristically monoarticular; the knee and hip are most frequently affected. Central nervous system tuberculosis takes various forms, including meningitis, tuberculoma, abscess, cerebritis, and miliary tuberculosis. Ileocecal involvement is seen in 80%-90% of patients with abdominal tuberculosis. The most common manifestation of abdominal tuberculosis is lymphadenopathy. Genitourinary tuberculosis is the most common manifestation of extrapulmonary tuberculosis. Lymphatic tuberculosis is more common among children, with cervical or supraclavicular nodes most frequently involved. Tuberculosis of the breast is extremely rare and occurs most often in young, multiparous, lactating women. The radiologic features of extrapulmonary tuberculosis mimic those of many diseases. A high level of suspicion is required, especially in high-risk populations. A positive culture or histologic analysis of biopsy specimens is still required in many patients for definitive diagnosis. PMID- 10715343 TI - Tuberculosis from head to toe. AB - Tuberculosis can affect virtually any organ system in the body and can be devastating if left untreated. The increasing prevalence of tuberculosis in both immunocompetent and immunocompromised individuals in recent years makes this disease a topic of universal concern. Because tuberculosis demonstrates a variety of clinical and radiologic findings and has a known propensity for dissemination from its primary site, it can mimic numerous other disease entities. Primary pulmonary tuberculosis typically manifests radiologically as parenchymal disease, lymphadenopathy, pleural effusion, miliary disease, or lobar or segmental atelectasis. In postprimary tuberculosis, the earliest radiologic finding is the development of patchy, ill-defined segmental consolidation. Both computed tomography (CT) and magnetic resonance (MR) imaging are helpful in diagnosing tuberculous spondylitis and tuberculous arthritis. CT is especially useful in depicting gastrointestinal and genitourinary tuberculosis. In tuberculosis involving the central nervous system, CT and MR imaging findings vary depending on the stage of disease and the character of the lesion. A high degree of clinical suspicion and familiarity with the various radiologic manifestations of tuberculosis allow early diagnosis and timely initiation of appropriate therapy, thereby reducing patient morbidity. PMID- 10715346 TI - Illuminations PMID- 10715347 TI - From the archives of the AFIP: pulmonary vasculature: hypertension and infarction. AB - Pulmonary hypertension is the hemodynamic consequence of vascular changes within the precapillary (arterial) or postcapillary (venous) pulmonary circulation. These changes may be idiopathic, as in primary pulmonary hypertension or pulmonary veno-occlusive disease, but more commonly they represent a secondary response to alterations in pulmonary blood flow. The pulmonary and systemic bronchial circulations form broad anastomoses that largely prevent infarction except in settings of markedly elevated pulmonary venous pressure, underlying malignancy, or excessive embolic burden. Causes of precapillary pulmonary hypertension include long-standing cardiac left-to-right shunt, chronic thromboembolic disease, and widespread pulmonary embolism arising from intravascular malignant cells, parasites, or foreign materials. The classic radiologic features of precapillary pulmonary hypertension are central arterial enlargement, sharply pruned peripheral vascularity, and right-sided heart hypertrophy and chamber dilatation. Postcapillary pulmonary hypertension may develop secondary to focal venous constriction or to compromised pulmonary venous drainage due to left atrial neoplasia, mitral stenosis, or left ventricular failure. Radiologic manifestations of postcapillary pulmonary hypertension include prominent septal lines, small pleural effusions, and occasionally air space opacities. In addition, radiologic evaluation of postcapillary pulmonary hypertension may demonstrate evidence of pulmonary arterial hypertension, secondary to the retrograde transmission of elevated pulmonary venous pressure across the capillary bed. PMID- 10715348 TI - The AAPM/RSNA physics tutorial for residents: internal radiation dosimetry: principles and applications. AB - Internal dose calculations in nuclear medicine normally use the techniques, equations, and resources provided by the Medical Internal Radiation Dose (MIRD) Committee of the Society of Nuclear Medicine. The MIRD schema uses a unique set of symbols and quantities to calculate the absorbed dose of radiation in any target organ per radioactive decay in any source organ. The calculations involve the energy emitted per radioactive decay, the fraction of the emitted energy that is absorbed in various target organs, the masses of these organs, and both the physical decay and biologic clearance of the injected radioactive material. Standardized mathematical models (phantoms) of the human body and standardized biokinetic models are also used. A computer program, MIRDose, calculates dose tables per unit administered activity of various radiopharmaceuticals. Special care must be taken when nuclear medicine procedures involve pregnant or lactating patients. New methodologies are becoming available to calculate doses to individual patients. PMID- 10715349 TI - Virtual labyrinthoscopy: visualization of the inner ear with interactive direct volume rendering. AB - Computed tomography (CT) is the modality of choice for detailed imaging of the bony labyrinth. Usually, information about the complex three-dimensional anatomic structures of the inner ear is presented as two-dimensional section images. Interactive direct volume rendering is a powerful method for visualization of the labyrinth. Unlike other visualization methods, direct volume rendering enables direct visualization of the bony labyrinth without explicit segmentation prior to the visualization process. Direct volume rendering was applied to visualization of the structures of the temporal bone in five patients without pathologic conditions and four patients with pathologic conditions. In all cases, clear representations of the bony labyrinth and the facial canal were provided. Because standard CT examinations combined with interactive visualization based on direct volume rendering are used, the method is fast and flexible. Therefore, this approach is applicable in routine clinical work. Problems occur in patients with effusion in the temporal bone because adjustment of imaging parameters for proper delineation of the target structures is difficult in this situation. However, direct volume rendering can produce meaningful images of high quality even in these problematic cases. The term virtual labyrinthoscopy is suggested for visualization of the labyrinth by using direct volume rendering. PMID- 10715350 TI - Clinical utility of three-dimensional US. AB - Three-dimensional (3D) ultrasonography (US) is rapidly gaining popularity as it moves out of the research environment and into the clinical setting. This modality offers several distinct advantages over conventional US, including 3D image reconstruction with a single pass of the US beam, virtually unlimited viewing perspectives; accurate assessment of long-term effects of treatment; and more accurate, repeatable evaluation of anatomic structures and disease entities. In obstetric imaging, 3D US provides a novel perspective on the fetal anatomy, makes anomalies easier to recognize, facilitates maternal-fetal bonding, and helps families better understand fetal abnormalities. Three-dimensional pelvic US allows volume data sets to be acquired with both transvaginal and transabdominal probes. Viewing multiple 3D power Doppler US images in a fast cine loop has proved useful in angiographic applications. Three-dimensional prostate US can help make accurate volume assessments for dosimetry planning or for estimating prostate-specific antigen levels. In breast imaging, 3D US has the capacity to demonstrate lesion margins and topography, thereby helping differentiate benign from malignant masses. Three-dimensional US can also help determine the need for biopsy and help facilitate needle localization and guidance during biopsy. With recent advances in computer technology and display techniques, 3D US will likely play an increasingly important role in medicine. PMID- 10715351 TI - Development of an electronic radiologist's office in a private institute. AB - A computer system that improves the quality, user-friendliness, accessibility, and management of radiology data (images, reports, databases, knowledge) was implemented at a private institute. A picture archiving and communication system (PACS) was integrated with the radiology information system (RIS). Two servers and 12 personal computers form the integrated system. The first server is dedicated to management and archiving of Digital Imaging and Communications in Medicine (DICOM) images. The second server is dedicated to management of the RIS and archiving of patient data (Structured Query Language database), reports (hypertext markup language [HTML]), and images in the Joint Photographic Experts Group (JPEG) format (mini-PACS). There are three main client-server networks: a common network of imaging modalities (magnetic resonance imaging, computed tomography, ultrasonography, digital radiography) and two fast Ethernet networks (the PACS network and the RIS network). The RIS-PACS is linked remotely with other workstations and servers via Integrated Services Digital Network (ISDN). Images and reports can be distributed to referring physicians in the form of multimedia HTML and JPEG documents, which can also be used for quick and easy archiving, distribution, and reviewing within the institute. However, referring physicians have been reluctant to use electronic reports and images. PMID- 10715352 TI - Distributing medical images with internet technologies: a DICOM web server and a DICOM java viewer. AB - With the advent of filmless radiology, it becomes important to be able to distribute radiologic images digitally throughout an entire hospital. A new approach based on World Wide Web technologies was developed to accomplish this objective. This approach involves a Web server that allows the query and retrieval of images stored in a Digital Imaging and Communications in Medicine (DICOM) archive. The images can be viewed inside a Web browser with use of a small Java program known as the DICOM Java Viewer, which is executed inside the browser. The system offers several advantages over more traditional picture archiving and communication systems (PACS): It is easy to install and maintain, is platform independent, allows images to be manipulated and displayed efficiently, and is easy to integrate with existing systems that are already making use of Web technologies. The system is user-friendly and can easily be used from outside the hospital if a security policy is in place. The simplicity and flexibility of Internet technologies makes them highly preferable to the more complex PACS workstations. The system works well, especially with magnetic resonance and computed tomographic images, and can help improve and simplify interdepartmental relationships in a filmless hospital environment. PMID- 10715353 TI - Decreased dorsolateral prefrontal N-acetyl aspartate in bipolar disorder. AB - BACKGROUND: N-acetyl aspartate (NAA) is an amino acid present in high concentrations in neurons and is thus a putative neuronal marker. In vivo proton magnetic resonance spectroscopy ((1)H MRS) studies have shown lower NAA concentrations in patients with various neurodegenerative disorders, suggesting decreased neuronal number, size, or function. Dorsolateral prefrontal (DLPF) NAA has not been extensively assessed in bipolar disorder patients, but it could be decreased in view of consistent reports of decreased DLPF cerebral blood flow and metabolism in mood disorders. We measured DLPF NAA in patients with bipolar disorder and healthy control subjects using in vivo (1)H MRS. METHODS: We obtained ratios of NAA, choline, and myoinositol (mI) to creatine-phosphocreatine (Cr-PCr) in bilateral DLPF 8-mL voxels of 20 bipolar patients (10 Bipolar I, 10 Bipolar II) and 20 age- and gender-matched healthy control subjects using (1)H MRS. RESULTS: DLPF NAA/Cr-PCr ratios were lower on the right hemisphere (p<.03) and the left hemisphere (p<.003) in bipolar disorder patients compared with healthy control subjects. CONCLUSIONS: These preliminary data suggest that bipolar disorder patients have decreased DLPF NAA/Cr-PCr. This finding could represent decreased neuronal density or neuronal dysfunction in the DLPF region. PMID- 10715354 TI - [123I]-beta-CIT SPECT imaging shows reduced brain serotonin transporter availability in drug-free depressed patients with seasonal affective disorder. AB - BACKGROUND: Numerous findings indicate alterations in brain serotonin systems in seasonal affective disorder (SAD). [(123)I]-2-beta-carbomethoxy-3-beta-(4 iodophenyl)-tropane ([(123)I]-beta-CIT) labels serotonin transporters (5-HTTs) in the midbrain. We performed a [(123)I]-beta-CIT single photon emission computer tomography (SPECT) study under the hypothesis of lower [(123)I]-beta-CIT binding reflecting reduced central 5-HTT availability in depressed SAD patients. METHODS: Depressed SAD patients and healthy control subjects were investigated using [(123)I]-beta-CIT SPECT 4 hours and again 24 hours after tracer injection. Subjects had either never used psychotropic medication or had been drug-free for at least 6 months prior to the investigation. Specific-to-nondisplaceable partition coefficient (V(3)") was calculated for the thalamus-hypothalamus and the midbrain-pons; the cerebellum served as a reference region. RESULTS: Patients showed a reduction in V(3)" in thalamus-hypothalamus (2.41+/-0.3 vs. 2.84+/-0.4; p = .026) 24 hours post tracer injection (p.i.). No difference between patients and control subjects was found in midbrain-pons (1.31+/-0.2 vs. 1.42+/-0.2; p = .39). No differences were detected in the SPECT acquisitions 4 hours p.i. CONCLUSIONS: Depressed SAD patients showed lower specific-to-nondisplaceable [(123)I]-beta-CIT binding in the region of interest (ROI) thalamus-hypothalamus. The small size of the midbrain-pons ROI may have contributed to the failure to show a difference in this ROI as well. Similar to reduced midbrain 5-HTT availability in nonseasonal depression, depression in SAD seems to be associated with reduced 5-HTT availability to the thalamus-hypothalamus. PMID- 10715355 TI - Increased risk of learning disabilities in low birth weight boys at age 11 years. AB - BACKGROUND: Few studies have examined learning disabilities among low birth weight (< or =2500 g) children, and those that have, have focused on very low birth weight children (<1500 g). We tested the hypothesis that low birth weight increases the risk of reading and math disabilities, examined possible sex differences in the effect of low birth weight, and assessed risk across the entire range of low birth weight. METHODS: Low birth weight and normal birth weight children were randomly selected from the 1983-1985 newborn lists of an urban and a suburban hospital in southeast Michigan. Children with neurological impairments were excluded. Children were evaluated at age 6 years and at age 11 years. Of the 823 children in the initial assessment, 717 (87.1%) participated in the second assessment. The Wechsler Intelligence Scale for Children--Revised and the Woodcock-Johnson Psycho-Educational Battery--Revised were used to identify children with learning disabilities. Learning disabilities were estimated in 574 children with IQs of > or =85. RESULTS: Low birth weight was associated with increased risk for reading and math disability in male children (odds ratio = 3.3 and odds ratio = 6.5, respectively) but not in female children. The increased risk of learning disabilities among male children applied to the entire range of low birth weight and was observed in both the urban and suburban communities. CONCLUSIONS: The effect of low birth weight on learning disabilities appears to be specific to male children. Although this sex-specific effect is consistent with previous findings of a greater vulnerability of male children to pregnancy and birth complications, it remains to be replicated and clarified. PMID- 10715356 TI - Developmental changes in postural sway in children at high and low risk for developing alcohol-related disorders. AB - BACKGROUND: To utilize the power of latent growth analysis to evaluate changes in postural sway during development in children who are either at high or low risk for developing alcoholism. METHODS: A total of 629 assessments of postural sway have been performed in children and adolescents (n = 126) who were evaluated annually over a 7-year period. RESULTS: Latent curve models indicated that these children/adolescents show a linear decrease in sway with age. Moreover, significantly different rates of change in the amount of sway between high- and low-risk offspring were seen. With the exception of one of the four stances tested, high-risk boys consistently showed a slower rate of improvement with respect to the amount of sway exhibited compared to low-risk boys. In girls, similar rates of improvement with age were seen in high- and low-risk individuals, though in one stance the high-risk girls showed a deterioration (greater sway with increasing age). CONCLUSIONS: Previous reports of increased postural sway in high-risk offspring most likely reflect a developmental delay (high-risk children have greater sway than is appropriate for their age based on normative values by age). PMID- 10715357 TI - Psychophysiologic assessment of aversive conditioning in posttraumatic stress disorder. AB - BACKGROUND: The objective of this study was to evaluate the acquisition, generalization, and extinction of conditioned physiologic responses to aversive stimuli in posttraumatic stress disorder (PTSD). METHODS: Thirty-six PTSD patients, 20 individuals with past trauma and no current PTSD, and 30 mentally healthy individuals without exposure to major trauma underwent a differential aversive conditioning experiment. Bursts of 105 dB white noise were used as unconditioned stimuli (UCSs), and 35x24 mm slides of different colors served as either CS+ (paired) or CS- (unpaired) stimuli. Heart rate (HR) and nondominant palm skin conductance (SC) were measured at rest and between 1 and 4 sec following each CS presentation. RESULTS: The PTSD group showed higher levels of resting SC and resting HR, larger SC responses to the initial presentation of unpaired CSs, larger HR responses following paired CS+ stimuli, larger SC responses to unpaired CS- during acquisition and extinction, and larger SC and HR responses to CS+ during extinction. The group differences in responses to CS+ during extinction remained statistically significant after controlling for age, resting physiologic levels, and initial responsivity. CONCLUSIONS: PTSD is associated with elevated autonomic responses to both innocuous and aversive stimuli, with larger responses to unpaired cues and with reduced extinction of conditioned responses. PMID- 10715358 TI - Sleep in a community sample of elderly war veterans with and without posttraumatic stress disorder. AB - BACKGROUND: Although sleep disturbances are commonly reported by individuals with posttraumatic stress disorder (PTSD), objective findings have been inconsistent, due in part to small sample sizes, comorbid psychiatric disorders, variations in the recentness of trauma exposure, and the use of PTSD subjects involved in psychiatric treatment. METHODS: A community sample of elderly males (n = 59) exposed to war trauma 28-50 years ago and free from sleep-affecting medications and disorders other than PTSD completed 3 nights of polysomnography. Of these participants, 30 met criteria for current PTSD; three were receiving supportive outpatient psychotherapy. RESULTS: Two statistically significant differences were observed: Those with PTSD had a higher percentage of rapid eye movement (REM) sleep and fewer arousals from non-REM sleep. The perceptions of sleep quality among the participants with PTSD were lower than the perceptions of non-PTSD participants. Although participants with untreated obstructive sleep apnea and sleep movement disorders were not included in the sample, many cases were detected on initial screening. Treatment resulted in improved sleep and increased feelings of well being. CONCLUSIONS: Alterations in REM and arousals characterized PTSD in this sample. When comorbid sleep disorders were ruled out, sleep was clinically similar across the groups. Trauma-related sleep disturbances that subjects reported as arising early in the course of the disorder appear to have declined over time. PMID- 10715359 TI - Low baseline and yohimbine-stimulated plasma neuropeptide Y (NPY) levels in combat-related PTSD. AB - BACKGROUND: Consistent with many studies demonstrating enhanced reactivity of the sympathetic nervous system in posttraumatic stress disorder (PTSD), the administration of yohimbine, a noradrenergic alpha(2)-antagonist, has been shown to increase core symptoms of PTSD and to induce greater increases in plasma 3 methyl-4-hydroxy-phenyl-glycol (MHPG) in subjects with PTSD compared with healthy control subjects. In turn, neuropeptide Y (NPY) has been shown to inhibit the release of norepinephrine from sympathetic noradrenergic neurons. METHODS: In the following study, plasma NPY responses to yohimbine and placebo were measured in a subgroup of 18 subjects with PTSD and 8 healthy control subjects who participated in the previous study of the effect of yohimbine on plasma MHPG. RESULTS: The PTSD subjects had lower baseline plasma NPY and blunted yohimbine-stimulated increases in plasma NPY compared with the healthy control subjects. Within the PTSD group, baseline plasma NPY levels correlated negatively with combat exposure scale scores, baseline PTSD and panic symptoms, and yohimbine-stimulated increases in MHPG and systolic blood pressure. CONCLUSIONS: This study suggests that combat stress-induced decreases in plasma NPY may mediate, in part, the noradrenergic system hyperreactivity observed in combat-related PTSD. The persistence of this decrease in plasma NPY may contribute to symptoms of hyperarousal and the expression of exaggerated alarm reactions, anxiety reactions, or both in combat veterans with PTSD long after war. PMID- 10715360 TI - A fenfluramine-activated FDG-PET study of borderline personality disorder. AB - BACKGROUND: Impulsive aggression in patients with personality disorders is associated with diminished levels of cerebrospinal fluid (CSF) 5-HIAA, blunted neuroendocrine responses to serotonergic agonists, and decreased glucose utilization in the prefrontal cortex. We tested the hypothesis that impulsive aggression in borderline personality disorder (BPD) may be associated with diminished serotonergic regulation in the prefrontal cortex, using positron emission tomography (PET) neuroimaging during pharmacologic challenge with d,l fenfluramine (FEN). METHODS: A 2-day, single-blind, placebo-controlled FEN challenge study was conducted in five patients with BPD (and no Axis I MDD) and eight healthy control participants. On Day 1, 4 mCi [(18)F]-fluorodeoxyglucose (FDG) was injected 3 hours after ingestion of placebo; on Day 2, FDG was injected 3 hours after ingestion of.8 mg/kg to 60 mg of d,l fenfluramine. After 30 min, a 45-min emission scan was acquired on the Siemans/CTI 951r/31 scanner. PET data were aligned to MR images and analyzed by Statistical Parametric Mapping (SPM96). RESULTS: In response to placebo, uptake of FDG was greater in control participants than patients in large areas of the prefrontal cortex including medial and orbital regions bilaterally (BA 10-11), left superior temporal gyrus, and right insular cortex. There were no areas in which patients had greater relative regional uptake than control participants. In response to FEN, relative regional uptake of FDG (relative to placebo) was greater in control participants compared to patients in medial and orbital regions of right prefrontal cortex (BA 10), left middle and superior temporal gyri (BA 22-23), left parietal lobe (BA 40), and left caudate body. CONCLUSIONS: Patients with BPD have diminished response to serotonergic stimulation in areas of prefrontal cortex associated with regulation of impulsive behavior. PMID- 10715361 TI - Serotonergic blunting to meta-chlorophenylpiperazine (m-CPP) highly correlates with sustained childhood abuse in impulsive and autoaggressive female borderline patients. AB - BACKGROUND: Disturbances of affect, impulse regulation, and autoaggressive behavior, which are all said to be related to an altered function of the central serotonergic (5-HT) system, are prominent features of borderline personality disorder (BPD). A high coincidence of childhood physical and sexual abuse is reported in these patients. Animal studies indicate that early, sustained stress correlates with a dysfunctional central 5-HT system. Therefore, we hypothesize that sustained traumatic stress in childhood affects the responsivity of the postsynaptic serotonergic system of traumatized BPD patients. METHODS: Following Axis I, Axis II, and trauma assessment, a neuroendocrine challenge test was performed with the postsynaptic serotonergic agonist meta-chlorophenylpiperazine (m-CPP) in 12 impulsive and autoaggressive female patients with BPD and 9 matched healthy volunteers. RESULTS: The cortisol and prolactin responses to the m-CPP challenge in BPD patients were significantly lower compared to those in controls. Within the group of patients with BPD, the net prolactin response showed a high inverse correlation with the frequency of the physical (r = -.77) and sexual abuse (r = -.60). CONCLUSIONS: Our data suggest that severe and sustained traumatic stress in childhood affects the 5-HT system and especially 5-HT(1A) receptors. This finding confirms the data from animal research. The blunted prolactin response to m-CPP appears to be the result of severe traumatization and independent of the BPD diagnosis. PMID- 10715362 TI - Hippocampus in Alzheimer's disease: a 3-year follow-up MRI study. AB - BACKGROUND: Due to the progressive nature of Alzheimer's disease (AD), it has been proposed that serial imaging studies tracking the course of progression might improve the diagnostic accuracy of AD. METHODS: Longitudinal changes in hippocampal volumes were evaluated using magnetic resonance imaging (MRI) over a period of 3 years in 27 AD patients and 8 control subjects. RESULTS: A statistically nonsignificant trend towards accelerated volume loss in the AD group compared to control subjects was observed. During the study period, the average shrinkage of the hippocampal volume ranged from -2.2% to -5.8% in control subjects, and from -2.3% to -15.6% in AD patients. CONCLUSIONS: The observed changes at an individual level were small, and within the accuracy range of the measurements. Therefore, serial MRI of the hippocampus did not offer any advantage over a single MRI to support the diagnosis of AD in this study sample. PMID- 10715363 TI - Steroid regulation of tryptophan hydroxylase protein in the dorsal raphe of macaques. AB - BACKGROUND: Tryptophan hydroxylase (TPH) is the rate-limiting enzyme for the synthesis of serotonin, and serotonin is a pivotal neurotransmitter in the regulation of mood, affective behavior, pituitary hormone secretion, and numerous autonomic functions. We previously demonstrated that estradiol (E) and progesterone (P) increase TPH mRNA levels in the dorsal raphe of macaques. METHODS: This study employed western blotting and densitometric quantitation to determine whether the changes observed at the level of gene expression were manifested by changes in TPH protein expression and whether modified estrogens or progestins had actions similar to the native ligands. In addition, the effect of the antiestrogen tamoxifen was examined. Ovariectomized (ovx) rhesus and cynomolgus macaques were untreated or treated with E, P, E+P, equine estrogens (EE), medroxyprogesterone (MPA), EE+MPA, or tamoxifen. The dorsal raphe region was subjected to Western analysis. RESULTS: E treatment for 28 days increased TPH protein mass four to six fold over ovariectomized controls. Addition of P to the E regimen or treatment with P for 28 days after E priming did not alter TPH from E treatment alone. Treatment of ovx macaques with a low dose of P caused a two fold increase in TPH protein. Treatment of ovariectomized macaques for 30 months with EE alone or MPA alone significantly increased TPH protein; however, unlike P, the addition of MPA to the EE regimen blocked the stimulatory effect of EE. Tamoxifen treatment significantly reduced TPH protein compared to EE and ovariectomized control animals. CONCLUSION: The stimulatory effect of E and P on TPH protein in the dorsal raphe of macaques correlates with the previously observed effect at the level of mRNA expression. P had no effect on the stimulatory action of E, whereas MPA blocked the stimulatory effect of EE. Tamoxifen acted as a potent antiestrogen on TPH protein expression. If TPH protein mass influences serotonin synthesis, then these steroids will impact many autonomic systems that are regulated by serotonin. PMID- 10715364 TI - Nomenclature of the gonane progestins. PMID- 10715365 TI - Failure rates among perfect users and during perfect use: a distinction that matters. AB - To make an informed decision when choosing a contraceptive, women and couples need to know how effective different methods are when used perfectly, where perfect use is defined as following the directions for use. In this article, we show that unbiased estimates of pregnancy rates during perfect use can be guaranteed only if information on consistency and correctness of use is available for each menstrual cycle. The estimated probability of pregnancy during a year of perfect use among the subset of women who always used a method perfectly will be biased upward. PMID- 10715366 TI - Efficacy, cycle control, and side effects of low- and lower-dose oral contraceptives: a randomized trial of 20 micrograms and 35 micrograms estrogen preparations. AB - Estrogen content represents a tradeoff between cycle control and side effects, but few direct comparisons of 20 and 30/35 micrograms preparations are available. To address this issue, we conducted a randomized, open-label multicenter clinical trial comparing Alesse (20 micrograms ethinyl estradiol [EE]), Mircette (20 micrograms EE), and Ortho Tri-Cyclen (35 micrograms EE) among 463 OC starters or switchers. Bloating, breast tenderness, and nausea were approximately 50% more common in women using 35 micrograms EE as compared to 20 micrograms EE preparations. Cycle control was similar in all products, although during the first two cycles among starters; users of Mircette and Ortho Tri-Cyclen (Tri Cyclen) exhibited better cycle control than Alesse users. Discontinuation and pregnancy rates were not significantly higher in 35 micrograms EE users. PMID- 10715367 TI - Oral contraception and other factors in relation to back disorders in women: findings in a large cohort study. AB - The Oxford-Family Planning Association contraceptive study includes 17,032 women, initially aged 25-39 years, recruited at 17 British family planning centers during the interval 1968-1974 and subsequently followed-up for periods up to 26 years. This article examines the pattern of referral to hospital for back disorders among these women. Certain back disorders have been reported to occur more frequently in oral contraceptive users than in other women, and back pain has also been reported in some women consequent to using an intrauterine device. The disorders considered were spinal osteoarthritis, displaced cervical disc, displaced lumbar disc, other and unspecified displaced disc, cervicalgia, unspecified back pain, and sprains and strains of the back. Spinal osteoarthritis and unspecified backache were the only two conditions significantly related (both positively) to age. Displaced lumbar disc and other and unspecified displaced disc were strongly positively related to height and weight. Unspecified backache showed similar, but less striking (in terms of the magnitude of the relative risks), associations with height and weight. Little evidence was found of any association between oral contraceptive use and any of the back disorders, and the same was true for intrauterine device use. PMID- 10715368 TI - Metabolic and fibrinolytic response to changed insulin sensitivity in users of oral contraceptives. AB - The fundamental role of insulin resistance for metabolic changes linked to cardiovascular disease and type 2 diabetes is increasingly recognized. Oral contraceptives (OC) may affect insulin sensitivity, and a detailed characterization hereof, as well as the secondary effects on related metabolic systems, are relevant in the evaluation of the risk of developing vascular disorders or diabetes in OC users. We studied insulin sensitivity index (S(I)), glucose effectiveness (S(g)), and insulin response in young, healthy women by frequently sampled intravenous glucose tolerance tests before and after randomization to 6 months of treatment with ethinyl estradiol in triphasic combination with norgestimate (n = 17) or gestodene (n = 20). Measurements of fasting triglycerides and antigen concentrations of tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor type 1 (PAI-1) were also included. Both compounds increased fasting plasma insulin and reduced S(i) but did not affect S(g). The relationships between S(i) and insulin response were unchanged. No consistent correlation between insulin sensitivity and triglycerides, t-PA, or PAI-1 were demonstrated before or during treatment. We conclude that the treatments were followed by a compensated decrease in insulin sensitivity that was unrelated to changes in triglycerides, t-PA, or PAI-1 antigen. PMID- 10715369 TI - Factors affecting continuation rates of DMPA. AB - A prospective study was conducted with 430 new depot medroxyprogesterone acetate (DMPA) acceptors to estimate continuation rates and investigate factors associated with length of use. Data were collected on services received and sociodemographic characteristics of participants. Women were enrolled over the course of 1 year and were followed for up to 13 months. Failure to return to the same clinic within 104 days of the last injection was the outcome of interest. The 3-, 6-, 9-, and 12-month continuation rates were 68%, 67%, 55%, and 51%, respectively. In the bivariate analysis, women who were told to return to the clinic for side effects were more likely to continue using DMPA than those who were not given such advice (p <0.05). Likewise, women who received information on DMPA efficacy, side effects, and amenorrhea were more likely to continue using DMPA compared to those who did not receive such information (p <0.05). A proportional hazards regression model was constructed to estimate the simultaneous effect of various factors on length of use. In results consistent with the bivariate analysis, women who were told to return to the clinic were 2.7 times more likely to continue using DMPA compared to women who did not receive that advice. Likewise, women who were told about the possibility of amenorrhea were 2.5 times more likely to continue using DMPA compared to those who did not receive that information. The regression model also identified new factors such as number of children, attitude toward menstruation, lactating at admission, and spousal input on method choice. The findings suggest that providers play an important role in ensuring the highest possible continuation rates for DMPA. PMID- 10715371 TI - A fixed formula to define the fertile window of the menstrual cycle as the basis of a simple method of natural family planning. AB - A significant number of women worldwide use periodic abstinence as their method of family planning. Many of them use some type of calendar-based approach to determine when they should abstain from unprotected intercourse to avoid pregnancy; yet they often lack correct knowledge of when during their menstrual cycle they are most likely to become pregnant. A simple method of natural family planning (NFP) based on a fixed formula to define the fertile window could be useful to these women. This article reports the results of an analysis of the application of a fixed formula to define the fertile window. A large existing data set from a World Health Organization study of the Ovulation Method was used to estimate the theoretical probability of pregnancy using this formula. Information about the variable probability of pregnancy on different cycle days relative to ovulation also was considered in the analysis. Results suggest that a fixed formula in which days 8-19 of the menstrual cycle are considered to be the fertile window would provide the appropriate basis of a simple, effective, family planning method. PMID- 10715370 TI - A comparison of tamoxifen and misoprostol to misoprostol alone for early pregnancy termination. AB - A study was undertaken to determine whether the combination of oral tamoxifen and moistened misoprostol administered vaginally was superior to that of placebo and moistened misoprostol administered vaginally for elective termination of early pregnancies.A clinical trial was conducted with a study group of 150 healthy women with pregnancies of or =18 years, mean age: 30.1 years+/-10.7) were randomised into 3 groups to receive a single dose of one of the study vaccines. In all groups, clinically significant reactions (severe) were infrequent (0-6%) and no serious adverse events were reported during the study. The incidence of local and systemic reactions following the administration of dTpa was comparable to the Td vaccine group. Of the total study group, prior to vaccination 52. 3 and 93.2% of the subjects had anti-diphtheria and anti-tetanus antibody levels > or = 0.1 IU/ml, respectively; and 73.1, 98.2 and 74.5% of the subjects were seropositive for pertussis toxin (PT), filamentous hemagglutinin (FHA) and pertactin (PRN) antibodies, respectively. One month after vaccination, a similar percentage of subjects in the dTpa and Td groups had anti diphtheria (88.4% vs 90. 1%) and anti-tetanus (100% vs 98.9%) antibody levels > or =0.1 IU/ml. Similar anti-FHA (100%) and anti-PRN (98.9%) vaccine response rates were seen in the dTpa and pa groups, while the anti-PT vaccine response rates were 96.8 and 100.0%, respectively. The dTpa vaccine is as well tolerated and immunogenic as the licensed Td vaccine, and additionally, can also boost antibodies against pertussis. PMID- 10715522 TI - Antibody responses, cytokine levels and protection of mice immunised with HSV-2 antigens formulated into NISV or ISCOM delivery systems. AB - The immunogenicity of a type 2 herpes simplex virus (HSV-2) antigen preparation following its formulation into immunostimulating complexes (ISCOMs) or non-ionic surfactant vesicles (NISV) was investigated in a murine model. The immune responses induced by each formulation were characterised by antigen specific total and subclass serum responses, and by lymphocyte proliferation and cytokine (interleukin-2 (IL-2), interleukin-4 (IL-4) and interferon-gamma (IFN-gamma)) production by in vitro restimulated spleen cells. The degree of protection afforded to mice by these various HSV-2 vaccine preparations against homologous (HSV-2) and heterologous (HSV-1) challenge infection was also determined. The findings suggest that formulation of the HSV-2 glycoprotein antigens with ISCOM or NISV delivery vehicles, and the methods used to prepare these formulations, influenced the immunogenicity of the final preparation. Higher IgG2a and neutralising antibody levels, IL-2 and IFN-gamma levels and lymphoproliferative responses were noted in mice immunised with the HSV-2 ISCOM formulated vaccine preparation. Furthermore, although HSV-2 antigens formulated in dehydration rehydration NISV, or entrapped in NISV by freeze-thawing at 30 degrees C (HSV-2 NISV 30), also elicited relatively high antibody, IL-2 and IFN-gamma levels and relatively high lymphoproliferative responses, formulation of HSV-2 antigens by freeze-thawing with NISV at 60 degrees C (HSV-2 NISV 60) did not. There were no differences between any of the HSV-2 vaccine formulations in terms of IL-4 induction in in vitro stimulated spleen cell cultures. Almost complete protection against HSV-2 challenge was afforded by the HSV-2 ISCOM preparation, while partial protection against challenge infection was afforded by the HSV-2 NISV 30 vaccine formulation. The findings are discussed in relation to the nature of the immune mechanisms, particularly Th1- or Th2-like responses, that may be elicited by HSV-2 antigen preparations formulated into various delivery systems and the relevance of these immune responses to protection against HSV infection in the murine model. PMID- 10715523 TI - A hepatitis B vaccine formulated with a novel adjuvant system. AB - Although more than 95% of the vaccinated population responds to the currently licensed vaccines against hepatitis B, some groups were found to be low responders. Lipid A as adjuvant, through its ability to activate macrophages, might improve humoral as well as cellular immune response. Therefore we evaluated the profile of a hepatitis B vaccine with the new adjuvant system SBAS4. 150 young adults were enrolled and randomized into three groups: one received the SBAS4 hepatitis B vaccine, the second Engerix-B(TM) and the third a hepatitis B vaccine with an alternative formulation on alum. Vaccinations were at 0 and 6 months. The vaccine was well tolerated. At month 7 all vaccinees were protected but with significant differences in GMTs between groups: 13,271 mIU/ml for the SBAS4 group versus 1203 and 1823 mIU/ml. Hence the hepatitis B vaccine with the new adjuvant system is more immunogenic compared to the other vaccines containing the same antigen and could be suitable for a two dose schedule. PMID- 10715524 TI - Immunogenicity of plasmid DNA encoding the 62 kDa fragment of Schistosoma japonicum myosin. AB - The recombinant Schistosoma mansoni 62 kDa myosin fragment, rIrV-5, is highly protective in experimental animals, however, vaccination of mice and rats with the recombinant Schistosoma japonicum homologue, rSj62, did not induce significant resistance against S. japonicum infection. To explore alternative ways of presenting this antigen, we further constructed a plasmid (VRSj62) which encodes Sj62 using the VR1020 vector and tested it in vaccination experiments. Four immunisations with 10 microg VRSj62 DNA alone were sufficient to induce high and progressively increasing levels of IgG antibodies against rSj62 with increasing numbers of injections in CBA/Ca mice (IgG titre > or =1:25000), and three injections with 50 microg VRSj62 DNA alone induced significant IgG responses in C57Bl/6 mice (IgG titre, 1:1600). However, vaccination with plasmid DNA entrapped in cationic liposomes or together with pUC19 DNA as a source of CpG motifs, both of which have been reported to enhance immune responses, did not enhance specific antibody production. In spite of the stimulation of specific antibodies against rSj62 with the naked DNA construct no resistance to challenge was demonstrated. PMID- 10715525 TI - Effect of pertussis toxin on the induction of nitric oxide synthesis in murine macrophages and on protection in vivo. AB - Macrophages from mice immunised with whole cell pertussis vaccine (WCV) responded in vitro to selected antigens by nitric oxide (NO) synthesis. This process was closely associated with macrophage activation. Because of the postulated role of traces of pertussis toxin (PT) in the protective effects of WCV, native PT and a genetically detoxified PT (g-PT) in combination with either a heat-treated whole cell pertussis vaccine (dWCV) or a three component acellular vaccine (ACV), were examined for their effects on NO induction in murine macrophages. The protective effects of these two forms of PT were examined in parallel using the intracerebral (ic) and aerosol challenge routes. Cultures of macrophages from mice immunised with dWCV and ACV, PT or g-PT produced less NO than comparable cultures from mice vaccinated with WCV. However, vaccination with either dWCV or ACV in combination with PT but not with g-PT, induced a significant increase (126 157%) in NO production by cultured cells and was associated with increased protection against challenge by both the ic and aerosol routes. These data indicate that a low concentration of PT acting as a co-factor in combination with other Bordetella pertussis antigens, can potentiate the activation of macrophages and that this process plays a key role in protection against infection. PMID- 10715526 TI - A gE-negative BHV1 vaccine virus strain cannot perpetuate in cattle populations. AB - Three identical transmission experiments were successively performed to quantitatively evaluate the possible transmission of a gE-negative bovine herpesvirus 1 (BHV1) vaccine strain among cattle. After intranasal inoculation, the vaccine virus was excreted in high titers in nasal fluids. However, the vaccine virus was transmitted to only one sentinel in one experiment, and not to any of the 10 sentinel cattle in the other two experiments. Based on these observations, it can be concluded that the expected number of cases per vaccine inoculated animal, i.e. the transmission ratio R(0) of the vaccine strain, is significantly below 1. The R(0) was estimated to be 0.14. After intramuscular inoculation, shedding of vaccine virus was not detected. Therefore, we concluded that it is highly unlikely that this live gE-negative BHV1 vaccine strain will perpetuate in the cattle population. PMID- 10715527 TI - Immunostimulatory effects of polar glycopeptidolipids of Mycobacterium chelonae for inactivated rabies vaccine. AB - Humoral and cellular immune responses were analyzed with Fuenzalida-Palacios rabies vaccine associated with pGPL-Mc, polar glycopeptidolipids extracted from Mycobacterium chelonae, aiming at its use as adjuvant. These results were compared to those obtained with BCG, a well-known immunostimulator, under the same conditions. Rabies vaccine plus pGPL-Mc (2.5 mg/kg) induced a significant increase in serum neutralizing activity, in vitro lymphocyte proliferation (spontaneous, specific and mitogen stimulation) and delayed type hypersensibility. In addition, pGPL-Mc, as well as BCG, enhanced the vaccine potency. Our results support further studies to encourage the use of pGPL-Mc as an immunostimulator of veterinary vaccines, before consideration for human vaccines. PMID- 10715528 TI - Quantitative and qualitative analyses of the immune responses induced by a multivalent minigene DNA vaccine. AB - Vaccines containing minigenes - isolated antigenic epitopes encoded by short open reading frames - can, under certain circumstances, confer protective immunity upon the vaccinee. Here we evaluate the efficacy of the minigene vaccine approach using DNA immunization and find that, to be immunogenic, a minigene-encoded epitope requires a perfect "Kozak" translational initiation region. In addition, using intracellular cytokine staining, we show that immunization with a plasmid encoding a full-length protein induces epitope-specific CD8(+) T cells which are detectable directly ex vivo, and constitute approximately 2% of the vaccinee's splenic CD8(+) T cells. In contrast, such cells are undetectable directly ex vivo in recipients of a minigene vaccine. Nevertheless, the minigene plasmid does induce a low number of epitope-specific CD8(+) T cells, which can be amplified to detectable levels by in vivo stimulation. Indeed, 4 days after in vivo stimulation (by virus infection), all vaccinated mice - regardless of whether they had been vaccinated with the minigene or with the full-length gene - had similar numbers of epitope-specific CD8(+) T cells. However, despite these strong responses at 4 days post-infection, recipients of the minigene vaccine showed no enhanced ability to limit virus replication and dissemination. We therefore observe a dichotomy; minigene vaccinees are not protected, despite the presence of strong virus-specific immune responses at 4 days post-challenge. We suggest that the protective benefits of vaccination exert themselves very soon - perhaps within minutes or hours - after virus challenge. If the vaccine-induced immune response is too low to achieve this early protective effect, virus-specific T cells will expand rapidly, but ineffectually, leading to the strong but non protective response measured at 4 days post-infection. Thus, vaccine-induced immunity should be monitored very early in infection, since the extent to which these responses may later be amplified is largely irrelevant to the protection observed. PMID- 10715529 TI - Vaccination of domestic pig with recombinant paramyosin. against Schistosoma japonicum in China. AB - Paramyosin (PM), a myosin-like protein is a major antigen on Schistosoma japonicum (Sj). We reported that passive transfer of a monoclonal IgE SjE18varepsilon.1 which recognizes PM of Sj (SJPM), partially protected mice from challenge infection. In the present study, we developed an experimental model system of schistosomiasis japonica with domestic pigs in China and used it for the evaluation of vaccination with recombinant SJPM (rSJPM). Sixteen-week-old pigs were successfully infected by dermal penetration of 120 cercariae of a domestic strain of Sj (50-60% worm recovery 11 weeks after challenge). The pigs vaccinated with 400 UV attenuated cercariae showed a reduction of worm recovery (53%, p<0.001). The experimental groups were immunized intradermally with rSJPM and alum or TiterMax and were partially protected against the challenge infection (32-35% reduction). PMID- 10715530 TI - Immunoadjuvant activities of E. coli- and plasmid-expressed recombinant chicken IFN-alpha/beta, IFN-gamma and IL-1beta in 1-day- and 3-week-old chickens. AB - In the present study we assessed the capacity of recombinant E. coli- or plasmid expressed chicken interferons (IFN) and chicken IL-1beta, to exert immunostimulatory activities for humoral immune responses, in day-old and adult chickens. We observed that both recombinant E. coli-expressed chicken IFN alpha/beta and IFN-gamma facilitated the induction of a primary and also a secondary antibody response, using tetanus toxoid (TT) as a bacterial model antigen, in immunologically mature 3-week-old chickens. In contrast, no improvement of antibody either type of chicken IFN was co-injected with inactivated Infectious Bursal Disease Virus (IBDV) antigen. TT-specific antibody formation was marginally increased by co-injection of recombinant E. coli expressed chicken IL-1beta. Combined administration of IFN-alpha/beta plus IFN gamma or IL-beta increased responses to TT in an additive, but not synergistic fashion. Remarkably, no augmentation of antibody responses specific for TT, nor IBDV, was noted in day-old birds, receiving IFN-alpha/beta or IFN-gamma as adjuvant. Also, intramuscular immunization of 3-week-old birds, using plasmids encoding IFN-alpha/beta together with TT protein antigen, significantly increased the speed and magnitude of TT-specific antibody responses. Plasmids encoding chicken IL-beta or IFN-gamma had a minimal or inhibitory effect, respectively. These data indicate a potential for chicken cytokines as immunoadjuvant for particular types of chicken vaccine antigens. PMID- 10715531 TI - The immunogenicity of single and combination DNA vaccines against tuberculosis. AB - DNA immunization is a promising new approach for the development of novel tuberculosis vaccines. In this study, the immune responses following the administration of single and combination tuberculosis DNA vaccines were evaluated. Single DNA vaccines encoding the MPT-63 and MPT-83 tuberculosis antigens evoked partial protection against an aerogenic challenge with M. tuberculosis Erdman in the mouse model of pulmonary tuberculosis. Immunization with a multivalent combination DNA vaccine (containing the ESAT-6, MPT-64, MPT 63, and KatG constructs) generated immune responses that indicated an absence of antigenic competition since antigen-specific cell-mediated and humoral responses were detected to each component of the mixture. More importantly, the combination vaccine elicited a strong protective response relative to the protection evoked by live BCG vaccine. PMID- 10715532 TI - Safety evaluation of recombinant cholera toxin B subunit produced by Bacillus brevis as a mucosal adjuvant. AB - Mucosal immune responses are known to play important roles in the establishment of protective immunity to microbial infections through mucosa. We examined the toxic effects of recombinant cholera toxin B subunit (rCTB) secreted by Gram positive bacterium Bacillus brevis as a mucosal adjuvant. Incubation of guinea pig peritoneal macrophages with cholera toxin (CT) or aluminium hydroxide gel (Al gel) released a significantly higher activity of lactate dehydrogenase than did commercial natural CTB (CTB) or rCTB. Intraintestinal or intramuscular administration of CT, CTB or Al-gel caused severe histopathological reactions. CT also caused infiltration of neutrophils and irregular arrangement or partial loss of the respiratory epithelium. In addition, CT and CTB elicited vascular permeability-increasing effects. rCTB elicited no toxic effects to macrophages and no vascular permeability-increasing effects. Moreover, it is noticeable that no distinct local histopathological reactions were observed in the nasal cavity, the small-intestinal loop or the muscle given rCTB. These results suggest that, from a safety standpoint, rCTB is a useful candidate as mucosal vaccine adjuvant. PMID- 10715533 TI - Signaling mechanisms of cytokine receptors and their perturbances in disease. AB - Cytokines regulate the proliferation and differentiation of cells through their interaction with specific receptors on the surface of target cells which are coupled to intracellular signal transduction pathways. The cytokine receptor class I superfamily, characterized by structural homology in the extracellular domain, includes receptors for many interleukins and hematopoietic growth factors, but also those of growth hormone, leptin, ciliary neurotrophic factor (CNTF), oncostatin M (OSM), leukemia inhibitory factor (LIF) and cardiotrophin-1 (CT-1). The receptors for interferons are structurally distinct and have therefore been categorized separately (class II cytokine receptors). The discovery of the JAK/STAT pathway in the early 1990s has been an important step forward in deciphering cytokine mediated signaling. This pathway connects activation of the receptor complexes directly to transcription of genes. Studies of humans and mice, deficient for one of the JAKs or STATs, have revealed crucial roles of these molecules in embryonic development, blood cell formation and immune responses. In addition, recent studies have revealed some of the mechanisms that control the activation of the JAKs and STATs, which contribute to signal intensity and specificity. In this review we will summarize these recent insights and discuss their implications for a variety of pathological conditions. PMID- 10715534 TI - The role of the testicular factor INSL3 in establishing the gonadal position. AB - INSL3, also designated Leydig insulin-like (Ley I-L) or relaxin-like factor (RLF), belongs to the insulin-like hormone superfamily. It is expressed in pre- and postnatal Leydig cells of the testis and in postnatal theca cells of the ovary. This sexual dimorphic pattern of INSL3 expression during development led us to suggest that the INSL3 factor could play an essential role in sexual differentiation, gonadal function and germ cell development. Key insights into the role of INSL3 came from analyses of INSL3 knockout mice. These mice showed impaired development of the gubernaculum ligament, a structure that is believed to mediate transabdominal descent of the testis during male embryogenesis. In double mutant XY-mice lacking INSL3 and a functional androgen receptor, it was demonstrated that both are essential for establishment of the sexual dimorphic position of the gonads through regulation of gubernaculum development and regression of the cranial suspensory ligament (CSL) during fetal life. Defects in this developmental process can cause cryptorchidism in the male, which is a most common disorder of sexual differentiation in human. PMID- 10715535 TI - Effect of estrogen on intracellular signaling pathways linked to activation of M(2)- and M(3)-muscarinic acetylcholine receptors in the rat myometrium. AB - The estrogen treatment of adult female rats induces an increase in myometrium sensitivity to cholinergic agonists and in this tissue the presence of M(2)- and M(3)-muscarinic acetylcholine (mACh) receptor was shown. We now report the effect of estrogen on intracellular signaling pathways linked to activation of M(2)- and M(3)-mACh receptor subtypes. The intracellular cyclic AMP accumulation and [3H] inositol phosphates content were measured in myometrium strips from rats in estrus (control) and estradiol-treated rats (12.5 microg/100 g body weight, sc, 24 h before experiments) (the plasma estradiol level was 30.9+/-3.5 pg/ml and 119.3+/-14.1 pg/ml from control and estrogen-treated rats, respectively). Estrogen treatment increased 2.5-fold the intracellular cyclic AMP accumulation induced by 10 microM forskolin. The effects of muscarinic agonist and antagonists on cyclic AMP accumulation were tested. Carbachol reduced the forskolin-induced intracellular cyclic AMP content, 3.0 and 10.5-fold, in myometrium from control and estradiol-treated rats, respectively. This inhibitory effect failed to occur when carbachol was incubated in the presence of methoctramine. Carbachol also induced increase on total [3H]-inositol phosphates accumulation in myometrium from estradiol-treated rats when compared with control rats. This effect was reversed by pfHHSiD. These studies suggest the modulation by estrogen of intracellular signaling pathways linked to activation of M(2)- and M(3)-mACh receptors in the rat myometrium. PMID- 10715536 TI - Expression of the early-late gene encoding the nuclear receptor HR3 suggests its involvement in regulating the vitellogenic response to ecdysone in the adult mosquito. AB - The insect steroid hormone, 20-hydroxyecdysone (20E), is a key factor controlling critical developmental events of embryogenesis, larval molting, metamorphosis, and, in some insects, reproduction. We are interested in understanding the molecular basis of the steroid hormone ecdysone action in insect egg development. The yellow fever mosquito, Aedes aegypti, in addition to being an important vector of human diseases, represents an outstanding model for studying molecular mechanisms underlying egg maturation due to stringently controlled, blood meal activated reproductive events in this insect. To elucidate the genetic regulatory hierarchy controlling the reproductive ecdysone response, we have investigated ecdysone-regulated gene expression in vitellogenic mosquito ovaries and fat bodies. We have previously demonstrated the conservation of a primary ecdysone triggered regulatory hierarchy, implicated in development of immature stages of Drosophila, represented by the ecdysone receptor/Ultraspiracle complex and an early gene E75 during the reproductive ecdysone response (Wang, S.-F., Miura, K., Miksicek, R.J., Segraves, W.A., Raikhel, A.S., 1998. DNA binding and transactivation characteristics of the mosquito ecdysone receptor - Ultraspiracle complex. J. Biol. Chem. 273, 27531-27540; Pierceall, W. E., Li, C., Biran, A., Miura, K., Raikhel, A.S., Segraves, W.A., 1999. E75 expression in mosquito ovary and fat body suggests reiterative use of ecdysone-regulated hierarchies in development and reproduction. Mol. Cell. Endocrinol. 150, 73-89). The present paper demonstrates that conservation of the factors involved in the ecdysone responsive genetic hierarchy regulating female reproduction extends beyond the early genes. Here, we identify AHR3, a highly conserved homologue of the Drosophila HR3 early-late ecdysone-inducible gene in the mosquito. We show that AHR3 is expressed in both vitellogenic tissues of the female mosquito, the fat body and the ovary. The expression of AHR3 correlates with the ecdysteroid titer, reaching a peak at 24 h after a blood meal. Moreover, in vitro fat body culture experiments demonstrate that the kinetics and dose response of AHR3 to 20 hydroxyecdysone (20E), an active ecdysteroid in the mosquito, is similar to those of the late vitellogenic genes rather than the early E75 gene. However, as shown for other early and early-late genes, the 20E activation of AHR3 is not inhibited by the presence of cycloheximide, a protein synthesis inhibitor. Taken together, these findings strongly suggest AHR3 involvement in regulating the vitellogenic response to ecdysone in the adult mosquito. PMID- 10715538 TI - The human growth hormone receptor gene - characterisation of the liver-specific promoter. AB - Several variants (V1-V8) have been described for the 5' untranslated region of human growth hormone receptor cDNA (Pekhletsky, R.I., Chernov B.K., Rubtsov, P.M., 1992. Variants of the 5'-untranslated sequence of human growth hormone receptor mRNA. Mol. Cell. Endocrinol. 90, 103-109). Transcription of one of these, variant V1, is under the control of a liver-specific promoter (Zou, L., Burmeister L.A., Sperling, M.A., 1997. Isolation of a liver-specific promoter for human growth hormone receptor gene. Endocrinology 138, 1771-1774). Further characterisation of this promoter shows that: (1) a cluster of exon 1 variants, which includes that coding for V1, is located about 13 kb upstream of the first coding exon; (2) the human V1 promoter (and not V7, as previously suggested) is homologous to liver-specific regulatory regions of rat and mouse GH receptor genes; (3) the transcription start site for V1 is 27 bp further upstream than previously reported; (4) a 158-bp sequence is sufficient for basal promoter activity, while larger constructs provide evidence for negative elements further upstream; and (5) nuclear proteins from HepG2 hepatoma cells protect regions of the V1 promoter from DNase digestion, revealing putative regulatory sites. PMID- 10715537 TI - A stable prostatic bioluminescent cell line to investigate androgen and antiandrogen effects. AB - We developed a new stable prostatic cell line expressing the human androgen receptor (AR) and the AR-responsive reporter gene to generate a powerful tool for investigating androgen action and for rapid screening of agonists and antagonists. The AR-deficient PC-3 cells were stably transfected with pSG(5)-puro hAR and pMMTV-neo-Luc. After selection with puromycin and neomycin, one highly inducible clone was isolated and named PALM, for PC-3-Androgen receptor Luciferase-MMTV. The expression of hAR was confirmed by western blot and steroid binding assays on the whole cells. The transcriptional activity of the clone was measured after incubation of cells with increasing concentrations of synthetic R1881 or natural androgens (DHT and testosterone). The three agonists had the same maximal activity at 0.1 microM and the fold induction was equal to 20. The agonist and antagonist activities of the steroidal antiandrogens (cyproterone acetate and RU2956) and the non-steroidal antiandrogens (nilutamide, bicalutamide, inocoterone and hydroxyflutamide) measured with the PALM cells were in good correlation with the results obtained with transiently transfected cells. The selectivity in steroid transactivation was demonstrated with estradiol, progesterone, cortisol, dexamethasone and aldosterone. Spironolactone and RU486 showed partial agonist and antagonist activities, whereas R5020 presented only a partial antagonist activity. We here demonstrate that this stable transfectant provides an accurate tool for studying wild-type human AR activation and its regulation by androgens and antiandrogens in a human prostatic epithelial cell, which is routinely available and remains androgen-responsive in vitro. PMID- 10715539 TI - Inhibition of deoxyglucose uptake in MCF-7 breast cancer cells by 2 methoxyestrone and 2-methoxyestrone-3-O-sulfamate. AB - Most cancer cells are dependent on glucose uptake to fulfil their energy requirements. In the present investigation we have examined the ability of 2 methoxyestrone (2-MeOE1), 2-methoxyestradiol (2-MeOE2), 2-methoxyestrone-3-O sulfamate (2-MeOEMATE), and a number of related compounds, to inhibit 2-deoxy-D [1-(3)H]-glucose uptake in MCF-7 breast cancer cells. Glucose uptake was shown to be linear with respect to cell number and time over a 5-35min period. 2-MeOE2, 2 MeOE1 and 2-MeOEMATE inhibited glucose uptake by 25-49% at 10 microM. 2 Hydroxyestradiol and estrone sulfate had little effect on glucose uptake, whereas estrone glucuronide inhibited uptake by 29%. There is evidence that 2 methoxyestrogens may exert an anti-mitotic effect on cells by stabilizing microtubules in a similar manner to that of paclitaxel. We therefore examined the effect of exposing cells to 2-MeOEMATE or paclitaxel for 24 h on basal or insulin stimulated glucose uptake. Using these conditions, 2-MeOEMATE and paclitaxel inhibited basal glucose uptake by 50 and 22%, respectively, and insulin stimulated uptake by 36 and 51%, respectively. The development of drugs that can inhibit glucose uptake could have therapeutic potential for the treatment of breast cancer. PMID- 10715540 TI - Differential transcriptional activation of peroxisome proliferator-activated receptor gamma by omega-3 and omega-6 fatty acids in MCF-7 cells. AB - While the role of dietary fats in breast cancer remains controversial, the recent cloning of peroxisome proliferator-activated receptor gamma (PPARgamma), a nuclear hormone receptor, from human breast cancer cells lines provides a potential molecular link. Several fatty acids from four classes of dietary fats were tested for their ability to mediate the transcriptional activity of PPARgamma in MCF-7 and MDA-MB-231 cells using growth media with minimal serum. Whereas omega-3 fatty acids inhibit transactivation of PPARgamma to levels below control, omega-6, monounsaturated and saturated fatty acids stimulate the activity of the transcriptional reporter. These studies indicate that individual fatty acids differentially regulate the transcriptional activity of PPARgamma by selectively acting as agonists or antagonists. Furthermore, the transcriptional activation of PPARgamma correlates with cell proliferation in MCF-7 cells. Understanding the effects of individual fats on breast cancer cells and PPARgamma transactivation could provide important new insights into the epidemiology of breast cancer and the role of dietary fat. PMID- 10715541 TI - Direct action of GnRH variants on goldfish oocyte meiosis and follicular steroidogenesis. AB - This study investigated the effects of gonadotropin-releasing hormones (GnRH) present in the goldfish brain (sGnRH and cGnRH-II) as well as a number of other GnRH variants on the reinitiation of meiosis and testosterone production in the follicle-enclosed goldfish oocytes, in vitro. All GnRH peptides tested individually stimulated oocyte meiosis in vitro, as determined by germinal vesicle breakdown (GVBD) as well as histone H1 kinase activity, which is an indicator of maturation promoting factor (MPF) production. The GnRH peptides tested had no detectable effect on basal follicular testosterone production with the exception of lGnRH-III, which had a relatively small stimulatory effect. In the presence of gonadotropin hormone (GTH), however, both sGnRH and lGnRH-III inhibited GTH-induced meiosis and steroidogenesis, whereas other GnRH peptides had no effect on GTH-induced responses. Addition of a GnRH antagonist effectively blocked the stimulatory effect of all GnRH peptides on oocyte meiosis, but was without effect on the inhibitory actions of sGnRH and lGnRH-III on GTH-induced meiosis, suggesting the involvement of different pathways mediating the stimulatory and inhibitory actions of sGnRH and lGnRH-III. These GnRH peptides were found to bind to the GnRH receptors in the goldfish ovary with different affinities (equilibrium association constant, K(a)). The findings provide novel information on the activity of GnRH variants in the goldfish ovary and provide a strong support for the hypothesis that GnRH plays a paracrine/autocrine role in the regulation of ovarian function in goldfish. PMID- 10715542 TI - Cloning and characterization of prostaglandin endoperoxide synthase-1 and -2 from the brook trout ovary. AB - Using a combination of reverse transcription-PCR and library screening, the cDNAs for prostaglandin endoperoxide synthase-1 (PGS-1) and 2 (PGS-2) were isolated from the brook trout ovary. The brook trout PGS-1 cDNA encodes for a 598 amino acid protein that is 69% identical to mammalian PGS-1. PGS-1 transcripts were observed in the ovary, spleen, gills, head kidney, trunk kidney, intestine, stomach, skin and heart. To our knowledge, this is the first characterization of a non-mammalian PGS-1 cDNA. The brook trout PGS-2 encodes for a 607 amino acid protein that is 69% identical to mammalian PGS-2 and was observed in the skin, gills, stomach and heart. PGS-2 transcripts were highly upregulated in the ovaries by the phorbol ester, phorbol-12-myristate-13-acetate, in combination with the calcium ionophore, A23187. However, PGS-2 was not observed in the ovary of brook trout undergoing natural oocyte maturation and ovulation. PMID- 10715543 TI - Nitric oxide production of rat Leydig and Sertoli cells is stimulated by round spermatid factor(s). AB - In this study, we provide evidence of cell-to-cell interaction between rat germ cells and Leydig or Sertoli cells in relation to nitric oxide (NO) production and inducible nitric oxide synthase (iNOS) messenger RNA (mRNA) expression. As a result of being cultured in a round spermatid-conditioned medium (RSd-CM), NO production in both Leydig and Sertoli cells increased in proportion to the length of the culture period. iNOS mRNA expression in both types of cells also increased in a dose-dependent manner as a result of being cultured with RSd-CM. This increase was detected as early as 3 h and was maintained up to 24 h. In contrast, neither NO production nor iNOS mRNA increased in either type of cell following culture in a pachytene spermatocyte-conditioned medium (PS-CM). Our findings suggest that RSd may control NO production of Leydig and Sertoli cells. This cell to-cell interaction may be an important mechanism of regulation of testicular function. PMID- 10715544 TI - Activation of local renin-angiotensin system by chronic hypoxia in rat pancreas. AB - Previous studies have provided evidence that several key elements of renin angiotensin system (RAS) are present in the rat pancreas, notably angiotensinogen, which is mandatory for intracellular generation of physiologically active angiotensin II. The data support the existence of an intrinsic RAS, which may be important for pancreatic blood flow and ductal anion secretion. In the present study, the effect of chronic hypoxia on the expression of RAS components, particularly at the levels of its precursor angiotensinogen and its receptor subtypes AT(1) and AT(2), were investigated in the rat pancreas. Results from western blot and semi-quantitative reverse-transcription polymerase chain reaction (RT-PCR) analyses unequivocally showed that chronic hypoxia caused a marked increase in angiotensinogen both at the protein and gene levels when compared with that in the normoxic pancreas. However, results from RT-PCR showed that there was a differential effect of chronic hypoxia on the expression of AT(1) and AT(2) receptor subtypes, which exhibited subtype-specific changes in gene expression. For AT(1), chronic hypoxia did not cause a significant change in mRNA expression for AT(1a) but a significant increase in mRNA expression for AT(1b). For AT(2), chronic hypoxia caused a marked increase in its mRNA expression. The increased expression of RAS component genes by chronic hypoxia and its significance of changes may be important for physiological and pathophysiological aspects of the pancreas. PMID- 10715545 TI - Growth hormone and insulin-like growth factor I protect intestinal cells from radiation induced apoptosis. AB - We studied whether programmed cell death (or apoptosis) is the predominant mechanism in radiation-induced cell damage to rat intestinal mucosa and investigated the mechanism of the protective effect of GH and IGF-I in the same model. Male albino Wistar rats were divided into four groups: controls, radiation, radiation plus GH and radiation plus IGF-I. Radiation was administered on the first day and on day 4. All animals were sacrificed and segments of the terminal ileum were stained with hematoxylin-eosin. Apoptosis of the epithelial cells was identified at the cellular level by the TUNEL stain and was distinguished from necrosis by the characteristic morphology of the cells (cytoplasmic shrinkage, marginal chromatin condensation and generation of nuclear apoptotic bodies). Apoptotic cells in the control animals were few and detected only at the tips of the villi while in the irradiated animals almost all the epithelial cells were apoptotic, distributed from the crypts to the tips of the villi and the mucosa showed severe epithelial atrophy and ulceration. The histologic picture of the mucosa in the GH and IGF-I treated animals was similar to normal controls and apoptotic cells were restricted only at the tips of the villi. DNA and RNA from the mucosa cells were isolated and analyzed by electrophoresis. DNA fragmentation and RNA 28s band ribonuclease cleavage was observed only in the irradiated animals. We have shown that abdominal radiation causes intestinal epithelial cell damage mainly through the induction of apoptosis and the treatment with GH and IGF-I inhibits apoptosis of the cells and preserves the mucosal integrity. PMID- 10715546 TI - TGF-beta1 stimulates expression of the aromatase (CYP19) gene in human osteoblast like cells and THP-1 cells. AB - Recent evidence has shown that bone is not only a target of estrogen action but also a source of local estrogen production. Bone cells such as osteoblasts express aromatase (P450arom) and the expression of P450arom in osteoblasts is positively regulated in a tissue specific fashion, as in the case of other tissues which express P450arom. To clarify the physiological factors regulating expression of P450arom in bone, we tested TGF-beta1 using osteoblast-like cells obtained from human fetuses as well as THP-1 cells. TGF-beta1 increased IL 1beta+DEX- induced aromatase activity in osteoblast-like cells, while it inhibited activity in skin fibroblasts. Similar enhancement of aromatase activity by TGF-beta1 was found in DEX-stimulated THP-1 cells and this cell line was used for further experiments. In THP-1 cells, TGF-beta1 enhanced DEX-induced aromatase activity almost linearly by 12 h and thereafter. Increased levels of P450arom transcripts were also demonstrated by RT-PCR at 3 h of TGF-beta1 treatment and thereafter. Cyclohexamide abolished enhancement of activity but did not inhibit the accumulation of P450arom transcripts induced by TGF-beta1. Increase in P450arom expression by TGF-beta1 was attributable to expression driven by promoter I.4. TGF-beta1 did not change the half life of P450arom transcripts. To identify the cis-acting elements responsible for TGF-beta1 action on aromatase expression, transient transfection assays were performed using a series of deletion constructs for promoter I.4 (P450-I.4/Luc). Two constructs (-410/+14 and 340/+14) that contain a functional glucocorticoid response element (GRE) and downstream sequence showed significant increase of luciferase activity in response to TGF-beta1. Deletion and mutation of the GRE in P450-I.4/Luc ( 340/+14) abolished the TGF-beta1. The luciferase activity of a (GRE)(1)-SV40/Luc construct was also stimulated by TGF-beta1. These results indicate that TGF-beta1 increases the expression of P450arom at the level of transcription through promoter I.4, at least in part via an enhancement of transactivation activity of the GR in THP-1 cells. TGF-beta1 is suggested to be one of the physiological up regulatory factors of bone aromatase. PMID- 10715547 TI - Active variants of human parathyroid hormone (1-34) with multiple amino acid substitutions. AB - We used site-directed mutagenesis to construct 55 single-site variants of rhPTH, a recombinantly-expressed form of human parathyroid hormone (1-34) containing three amino acid changes compared to the natural sequence (ML8, ML18 and FY34). We identified several mutations, at residues Lys(13), Glu(19), Val(21), Glu(22), Lys(27) and Asp(30), that increase biological activity by up to 2. 5-fold, as measured by stimulation of adenylate cyclase activity in rat UMR-106 cells. We constructed a series of 15 variants in which two to eight substitutions at these positions were combined, and found that the mutations behaved additively, leading to peptides with significantly enhanced potency. The most active combination variant, with six substitutions (KS13, ES19, VQ21, ES22, KQ27 and DN30), is 15 times more active than the parent molecule. However, the extent to which such combinations increase the activity of the peptide depends critically on the identity of the residues at positions 8 and 18. We constructed two of the combination variants in a variety of sequence backgrounds containing different combinations of leucine, methionine and norleucine at positions 8 and 18. Enhancements in potency were significantly reduced when Met or Nle was present at either of these positions, both in UMR-106 cells and human SaOS-2 cells. A corresponding non-additivity was observed in direct measurements of receptor binding affinity on UMR-106 cells. These results suggest that interactions, either direct or indirect, between certain PTH side chains prevent these mutations from behaving in an additive manner. PMID- 10715548 TI - Prostacyclin IP receptor up-regulates the early expression of C/EBPbeta and C/EBPdelta in preadipose cells. AB - Prostacyclin (PGI(2)) and its stable analogue carbacyclin (cPGI(2)) are known to trigger the protein kinase A pathway after binding to the cell surface IP receptor and to promote or enhance terminal differentiation of adipose precursor cells to adipose cells. The early expression of C/EBPbeta and C/EBPdelta is known to be critical for adipocyte differentiation in vitro as well as in vivo. We report herein that in Ob1771 and 3T3-F442A preadipose cells, activation of the IP receptor by specific agonists (PGI(2), cPGI(2) and BMY 45778) is sufficient to up regulate rapidly the expression of C/EBPbeta and C/EBPdelta. Cyclic AMP-elevating agents are able to substitute for IP receptor agonists, in agreement with the coupling of IP receptor to adenylate cyclase. Consistent with the fact that PGI(2) is released from preadipose cells and behaves as a paracrine/autocrine effector of adipose cell differentiation, the present results favor a key role of prostacyclin by means of the IP receptor and its intracellular signaling pathway in eliciting the critical early expression of both transcription factors. PMID- 10715549 TI - Additional N-glycosylation at Asn(13) rescues the human LHbeta-subunit from disulfide-linked aggregation. AB - CG, LH, FSH, and TSH are a family of heterodimeric glycoprotein hormones that contain a common alpha-subunit, but differ in their hormone-specific beta subunits. Despite the considerable homology between LHbeta and CGbeta, we previously demonstrated that, when expressed in GH(3) cells, the secreted form of LHbeta showed mispaired disulfide-linked aggregation in addition to monomer, whereas no aggregation was observed in CGbeta. To determine the domains which are associated with the LHbeta-aggregation and which prevent CGbeta-aggregation, mutant beta-subunits in glycosylation and carboxy-terminus were expressed in GH(3) cells, and the occurrence of aggregation was assessed by continuous labeling with [35S]methionine/cysteine, immunoprecipitation with anti-hCGbeta serum, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis in a non reducing condition. No aggregation was seen when N-linked oligosaccharides were attached to the Asn(13) of LHbeta. Removal of the carbohydrate unit at the Asn(13) of CGbeta caused aggregation, although the amount was less than 10% of monomer. The carboxy-terminal regions of neither LHbeta nor CGbeta were associated with their aggregation. Both CGbeta wild-type (WT) and CGbeta lacking N-glycosylation at Asn(13) (CGbeta-N13) showed aggregates in lysate. However, in contrast to CGbeta-N13, CGbetaWT revealed no aggregation in medium. These results indicate that the backbone structure consisting of 114 amino acids and N-linked glycosylation at Asn(30) is involved in the aggregation of LHbeta. Moreover, N glycosylation at Asn(13) does not prevent such aggregation, but instead plays an important role in correct folding for both LHbeta- and CGbeta-subunits to be secreted as monomer. PMID- 10715550 TI - Cyclic AMP stimulates the gene expression of a non-selective cation channel, mNSC1, in pancreatic beta-cell line, MIN6. AB - Mouse non-selective cation channel 1 (mNSC 1) cDNA from mouse pancreatic beta cell line, MIN6, have recently been cloned. Since the number of non-selective cation channel in pancreatic duct cells has been reported to increase 9-fold in 5 h incubation with cAMP, the effect of cAMP on the gene expression of mNSC1 in MIN6 cells was examined. Dibutyryl cAMP (db-cAMP) was shown to increase the level of the mRNA by reverse transcription-polymerase chain reaction (RT-PCR). The copy number of the mRNA was increased 4-fold in 6 h incubation with db-cAMP by competitive PCR. Western blot analysis also indicated a 4-fold increase in the quantity of the newly synthesized protein in 9 h incubation with db-cAMP. Experiments with 5'-flanking region and with a transcriptional inhibitor suggested that db-cAMP affected transcription, and protected the mRNA from its degradation as well. It is concluded that the expression of mNSC1 is indeed increased by cAMP in the pancreatic beta-cells. PMID- 10715551 TI - Biochemical and molecular biological evidence for the presence of type II iodothyronine deiodinase in mouse mammary gland. AB - In the present study we have obtained several lines of evidence indicating the presence of type II iodothyronine deiodinase (DII) in the mouse mammary gland. 5' deiodinase activity in the mammary gland has an apparent K(m) value of 4.4 nM for T(4) and is inhibited by aurothioglucose but not by propylthiouracil. These characteristics are similar to those of DII in other tissues. We cloned a 1.4-kb cDNA, which contains the entire mouse DII coding region and has high homology with the rat DII cDNA, from the mammary gland and brain. Northern blot analysis showed the presence of 7.9 kb DII mRNA in the mammary gland and brain. The levels of DII activity and mRNA in lactating gland were significantly lower than those in virgin and pregnant glands, suggesting that DII is regulated at the pretranslational level. In addition, we found the low level of DII enzyme activity and transcript in various other mouse tissues. PMID- 10715552 TI - Activator protein-2 regulates human placental lactogen gene expression. AB - DNase I footprint analysis of the human placental lactogen-A (hPL-A) promoter using nuclear extracts from purified human trophoblast cells and BeWo choriocarcinoma cells revealed five protected regions within the proximal 325 bp. Two of the protected regions, FP4 (-289--267) and FP5 (-167--154), are homologous to regions on the human growth hormone (hGH) promoter that bind transcription factors AP-2 and/or NFI. Competitive gel shift assays and supershift assays indicated that FP4 forms complexes with activator protein-2 (AP-2) and nuclear factor I (NFI), while FP5 forms a complex with AP-2 alone. In transient transfection studies in human trophoblast cells, hPL promoter constructs containing point mutations in the AP-2 binding sites of FP4 and/or FP5 were 60 80% less active than plasmids containing the wild-type promoter. A mutation in the NFI binding site of FP4, however, had little effect on promoter activity in these cells. Overexpression of AP-2 in HepG2 cells co-transfected with the wild type hPL promoter resulted in a significant increase in promoter activity. Taken together, these findings suggest a critical role for AP-2 in the regulation of hPL gene expression. PMID- 10715553 TI - Transcriptional regulation of the expression of macrophage colony stimulating factor. AB - The regulatory regions for transcriptional control of the MCSF gene are unknown. We examined regulatory control in a 774-bp murine MCSF promoter transfected into MC3T3-E1 osteoblast-like and COS-7 cells. Deletion of upstream sequence from -635 increased basal activity of the promoter by at least four-fold, an increase that was maintained when PU.1, NFkappaB and Egr1/Sp1 consensus sequences were subsequently removed. Mutagenesis identified a suppressor element between -635 and -642 from the transcriptional start site and an oligonucleotide representing this sequence was retarded by nuclear cell protein. TNFalpha (1 ng/ml), PTH (5x10(-8) M), and IL-1alpha (100 pg/ml), which increased MCSF protein secretion, failed to enhance the transcriptional rate of the full-length promoter. TNFalpha was able to stimulate transcription of a heterologous reporter transfected into COS-7 containing multiple copies of the murine MCSF NFkappaB site inserted before a minimal promoter. In contrast, deletion of the same NFkappaB response element increased basal activity in the native promoter. Thus, the NFkappaB sequence may act as a negative regulator in the context of the endogenous promoter. Our results indicate that constitutive transcriptional activity conferred by the MCSF promoter may be damped by a suppressor protein. Transcriptional regulation, however, does not appear to be a major stimulatory mechanism for MCSF secretion. PMID- 10715554 TI - Human follicle stimulating hormone receptor variants lacking transmembrane domains display altered post-translational conformations. AB - Variant splicing of gonadotropin receptor mRNA commonly occurs, however expression of receptor protein variants and their trafficking has yet to be studied in detail. To determine receptor variant trafficking and intracellular processing in mammalian cells, the intracellular fate of intentionally truncated variants of human follicle stimulating hormone receptor (hFSH-R) expressed in CHO cells was examined. Monoclonal antibodies (mAbs) were made against the hFSH-R's extracellular domain (ECD) expressed in insect cells. Four mAbs 106.156, 106.290, 106.318, and 106.263 were chosen as probes. Epitope mapping using synthetic peptides, and truncated hFSH-R variants revealed that mAb 106.156 bound to ECD residues 183-220, while mAbs 106.318, 106.290, 106.263 bound ECD residues 300 331. Immunofluorescence microscopy showed that mAbs 106.318 and 106.156 stained the surface of fixed, intact CHO cells expressing wild type hFSH-R. However, following cell permeabilization all four antibodies stained hFSH-R in Golgi and endoplasmic reticulum. Permeabilized cells expressing truncated variants ECD213 and ECD254 showed staining accumulated in the endoplasmic reticulum/nuclear envelope continuum. ECD335/His was found to accumulate in extended endoplasmic reticulum (ER). The ER location of ECD335/His was confirmed by double labeling experiments with concanavalin A and ECD mAb. Glycosidase digestion followed by Western blot analysis show ECD213 and ECD335/His to be glycosylated, but not ECD254. Both glycosylated truncated hFSH-R variants were sensitive to peptide-N glycanase F and endoglycosidase H but insensitive to neuraminidase indicating that these variants possess high mannose type oligosaccharides. Thus truncated hFSH-R variants do not reach the medial or trans Golgi where high mannose oligosaccharides are trimmed and sialic acid is added. These data suggest that the conformation the ECD of the wild type receptor is different from the ECD alone expressed in the endoplasmic reticulum. This information suggests that the ECD serves two distinct roles; the first is to bind FSH and the other is likely to contact the endodomain of the receptor, which presumably leads to activation of the endodomain for signal transduction. PMID- 10715555 TI - Cellular mechanisms of CNS pericytes. AB - Three major functional roles have been ascribed to pericytes associated with central nervous system microvasculature-contractility, regulation o f endothelial cell activity, and macrophage activity. A host of different cell factors and signalling agents appear to be involved with these cellular functions, some effecting the pericyte and others produced by this cell. These include neuromodulators, vasoactive peptides, metabolic factors, growth factors and cytokines. The specific compounds and their actions are collectively viewed in an effort to provide an overall picture of the regulation of pericyte functional activity. This small vascular cell is emerging as a significant entity in several physiological processes through the functions of above; these processes include control of blood flow, regulation of vascular development and immune responses. Defining the regulatory agents and their mechanisms is key to understanding the role that pericytes play in these processes. Because these cells have begun to receive increasing attention in neurobiological studies, an overview of signalling properties should be timely and beneficial. PMID- 10715556 TI - Enhancing effect of electric stimulation on cytotoxicity of anticancer agents against rat and human glioma cells. AB - Electropermeabilization, or electroporation, has been used to deliver genes or drugs into the cytoplasm through micropores in the cell membrane caused by electric stimulation. The cytotoxic effect of a combination of anticancer agents with electric stimulation on rat C6 and human T98G glioma cells was examined in vitro. Electric pulses of 100 microsec square waves (eight cycles at 1 Hz) at various electric fields were delivered to C6 or T98G glioma cell suspensions in combination with several anticancer agents. Cell growth was evaluated 48-72 h after treatment. Measurement of cell lysis by electric stimulation was used to assess the optimum field strength for electroporation. Electric stimulation enhanced significantly the cytotoxicity of bleomycin to both C6 and T98G cells by more than 1000-fold using an electric field of 1750 V/cm for C6 cells and 1000 V/cm for T98G cells. The enhancement disappeared when bleomycin concentration was reduced to 100 pg/ml. The cytotoxicity of carboplatin was weakly but significantly enhanced by electric stimulation when a high dose of carboplatin was used. However, there was no enhancement of the cytotoxicity of nimustine hydrochloride (ACNU), etoposide, and vincristine. These results indicate that the combination of bleomycin and electroporation is the most potent candidate for electrochemotherapy in vivo. PMID- 10715557 TI - Distribution of neurons immunoreactive for calcium-binding proteins varies across areas of cat primary somatosensory cortex. AB - The primary somatosensory (SI) cortex in the cat contains four cytoarchitectonic areas that appear to contain separate body representations and have different functions. We tested whether functional differences among these areas are reflected in the densities of neurons containing each of three calcium-binding proteins: parvalbumin (PV), calbindin (CB), and calretinin (CR). Colocalization experiments revealed that CR was localized in a population of neurons distinct from those containing PV or CB. The general laminar distributions of the three calcium-binding proteins were similar to those described in other species and cortical areas, but there were significant density differences in layers II and III across SI. The density of PV-immunoreactive neurons was higher in areas 3b and 1 than in areas 3a and 2. CB-immunoreactive neurons were found in higher densities in anterior SI than in posterior SI, and the pattern of CR immunoreactive neurons was reciprocal to that of CB, with significantly higher densities in posterior regions of SI. Since the firing characteristics of nonpyramidal neurons appear to be related to their calcium-binding protein content, differences in regional distributions of these neurons in layers II and III may contribute to functional differences between the cytoarchitectonic areas of SI cortex. PMID- 10715558 TI - Serotonergic modulation of the P13 midlatency auditory evoked potential in the rat. AB - The vertex-recorded, sleep state-dependent P13 midlatency auditory evoked potential in the rat may be generated, in part, by pedunculopontine nucleus (PPN) projections. Injections into the PPN of the 5-HT(1A) serotonin receptor agonist, 8-hydroxy-2-di-n-propylaminotetralin hydrobromide (DPAT), were found to reduce the amplitude of the P13 potential in a dose- and time-dependent manner. The suppressive effect of DPAT was blocked or reduced by pretreatment with the 5 HT(1A) serotonin receptor antagonist, Pindobind. These results show that the P13 potential can be modulated by known inhibitory serotonergic inputs to the PPN. PMID- 10715559 TI - A reduced compartmental model of the mitral cell for use in network models of the olfactory bulb. AB - We have developed two-, three- and four-compartment models of a mammalian olfactory bulb mitral cell as a reduction of a complex 286-compartment model [1]. A minimum of three compartments, representing soma, secondary (basal) dendrites and the glomerular tuft of the primary dendrite, is required to adequately reproduce the behaviour of the full model over a broad range of firing rates. Adding a fourth compartment to represent the shaft of the primary dendrite gives a substantial improvement. The reduced models exhibit behaviours in common with the full model which were not used in fitting the model parameters. The reduced models run 75 or more times faster than the full model, making their use in large, realistic network models of the olfactory bulb practical. PMID- 10715561 TI - Effects of metabolic fragments of [Arg(8)]-vasopressin on nerve growth in cultured hippocampal neurons. AB - The effects of metabolic fragments of [Arg(8)]-vasopressin (AVP), [pGlu(4), Cyt(6)]AVP (AVP(4-9)), and desglycinamide-[pGlu(4), Cyt(6)]AVP (AVP(4-8)) on the growth of hippocampal neurons in culture were investigated in comparison with those of AVP. AVP(4-9) caused a significant increase in filopodial length following 96 h of exposure at concentrations higher than 300 nM. AVP(4-9) was more potent than AVP. AVP(4-8) also induced an increase in filopodial length, but this effect was less than that of AVP. The selective V(1) agonist [Phe(2), Ile(3), Orn(8)]-vasopressin caused a significant increase in filopodial length, whereas the selective V(2) agonist [deamino-Cys(1), D-Arg(8)]-vasopressin showed no such effect. OPC-21268, a vasopressin V(1) antagonist, blocked AVP and AVP fragment-induced increases in filopodial length. However, the V(2) antagonist OPC 31260 showed no such effect. A23187, a representative Ca ionophore, also increased filopodial length, and the A23187-induced increase in filopodial length was potentiated by AVP and AVP fragments. These results indicated that AVP(4-9) and AVP(4-8) increased filopodial length in cultured hippocampal neurons by activating V(1) receptors. Both phenomena induced by AVP(4-9) and AVP(4-8) were associated with intracellular calcium mobilization. PMID- 10715560 TI - Nitric oxide modulates release of noradrenaline in guinea-pig gastric fundus. AB - The interaction between nitric oxide (NO) and the release of [(3)H]noradrenaline ([(3)H]NA) in conditions of non-activated and activated nicotinic receptors in guinea-pig gastric fundus preincubated with [(3)H]NA was studied. Nicotinic receptor agonist, 1,1-dimethyl-4-phenyl-piperazinium iodide (DMPP) (100 microM) significantly increased the resting release of [(3)H]NA. NO-synthase inhibitor, N(omega)-nitro-L-arginine (L-NNA) (100 microM) significantly decreased DMPP induced release of [(3)H]NA. Field electrical stimulation (FES) (2Hz; 1 ms; 360 st) significantly increased the release of [(3)H]NA above the basal levels. L-NNA significantly decreased the stimulation-evoked release of [(3)H]NA. DMPP increased the stimulation-evoked release of [(3)H]NA, effect which was significantly decreased by L-NNA. The data suggests that endogenous NO increases the release of [(3)H]NA, evoked either by activation of the nicotinic receptors or by electrical stimulation in guinea-pig gastric fundus. PMID- 10715562 TI - The effect on opioid peptides in the rat brain, after chronic treatment with the anabolic androgenic steroid, nandrolone decanoate. AB - In recent years, an increase in abuse of anabolic androgenic steroids (AAS) has been seen among individuals not directly connected to sports. Clinical evidence suggests that abuse of these steroids may result in profound changes in personality, expressed by depressive symptoms, irritability and increased aggression. It is still unknown whether these alterations are related to changes in any particular transmitter system or whether they are persistent or reversible. In this study we focused on AAS effect on the endogenous dynorphin and enkephalin system in the brain. Male rats were given intramuscular injections of the AAS nandrolone decanoate (15 mg/kg), once daily for 2 weeks. The levels of the opioid peptide immunoreactivities (ir) were assessed by radioimmunoassay in two groups immediately after the treatment and in two other groups after additional 3 weeks without any drug treatment (recovery period). The result indicates that chronic AAS treatment increased the activity in the dynorphin B- and Met-enkephalin-Arg(6)Phe(7)-ir in the hypothalamus, striatum and periaqueductal gray (PAG) compared to controls. In addition, the steroid induced an imbalance between the dynorphin and the enkephalin opioid system in the nucleus accumbens, hypothalamus and PAG. This imbalance remained after the recovery period. Since increased peptide activity was found in brain regions regulating emotions, dependence, defensive reactions and aggression, it was suggested that the actual endogenous opioid systems are involved in previously reported AAS-induced changes in these behaviours. PMID- 10715563 TI - Repeated prenatal cocaine increases met-enkephalin immunoreactivity in respiratory-related medulla of developing swine. AB - Repeated prenatal exposure to cocaine is associated with attenuated respiratory and arousal responses in infants and piglets. As the normal development of these functions is influenced by medullary opioid systems, the present study explored the possible contribution of the opioid systems to the attenuation induced by cocaine. Methionine-enkephalin (met-enkephalin), an endogenous opioid peptide, was delineated by immunocytochemistry in respiratory- and arousal-related medullary regions of relatively immature (6-7-day-old) and more mature piglets (20-21-day-old). The animals were either unexposed, or exposed prenatally to 2 mg/kg cocaine four times daily administered to the pregnant sows intravenously throughout the last third of gestation. At control, met-enkephalin was found in the neurons, fibers and terminals of the respiratory- and arousal-related medullary regions throughout the age range studied. Prenatal cocaine exposure increased met-enkephalin immunoreactivity in the respiratory-related hypoglossal and solitary tract nuclei of both age groups. These findings support a modulatory role of met-enkephalin in the normal development of respiratory control, and an involvement of this peptide in the attenuation of respiration by repeated prenatal exposure to cocaine. PMID- 10715564 TI - ACTH- and alpha-MSH-induced grooming, stretching, yawning and penile erection in male rats: site of action in the brain and role of melanocortin receptors. AB - The effect of adrenocorticotropin (ACTH)(1-24) and alpha-melanocyte stimulating hormone (alpha-MSH) on grooming, stretching, yawning and penile erection was studied after injection into different brain areas. Both peptides induce the above responses when injected into the hypothalamic periventricular region of the third ventricle. This region includes the paraventricular nucleus, the dorsomedial nucleus, the ventromedial nucleus and the anterior hypothalamic area. The minimal effective dose of both peptides was 0.5 microg and the maximal effect was seen with 2 microg, the highest dose tested. Irrespective of the injection site, grooming started 5-7 min after injection of either peptide, while stretching, yawning and penile erection started only after 15-35 min and lasted for 90-120 min. In contrast both peptides were ineffective when injected into the preoptic area, the caudate nucleus or the CA1 field of the hippocampus. Grooming, stretching and yawning, but not penile erection, were prevented by cyclic[AcCys(11), D-Nal(14), Cys(18), AspNH(2)(22)]-beta-MSH (11-22) (HS014), a selective melanocortin 4 receptor antagonist, injected into the same periventricular area 10 min before of ACTH(1-24) or alpha-MSH. The results show that ACTH(1-24) and alpha-MSH act in the hypothalamic periventricular region to induce the above responses and that grooming, stretching and yawning, but not penile erection, are mediated by melanocortin 4 receptors. PMID- 10715566 TI - Multiple mechanisms in brain maturation and degeneration PMID- 10715565 TI - Multiple substrates for serotonergic modulation of rat locus coeruleus neurons and relationships with kainate receptors. AB - The ultrastructural substrates of serotonin (5-hydroxytryptamine [5-HT]) immunoreactive terminals with respect to noradrenergic neurons of the locus coeruleus (LC) have only been suggested from immunocytochemical analysis in adjacent tissue sections using antisera directed against tryptophan and tyrosine hydroxylase (TH) enzymes. Here, we conducted dual immunoelectron microscopy in the same section of tissue using antisera directed against 5-HT and TH to determine cellular substrates for proposed interactions between these two transmitter systems. Axon terminals containing peroxidase labeling for 5-HT possessed small clear as well as large dense core vesicles. Of 176 5-HT-labeled axons and terminals, 19% contacted TH-labeled dendrites. When a synaptic specialization was detectable, it was more often of the asymmetric type. Electrophysiological studies have also shown that 5-HT selectively attenuates excitatory amino acid-induced activation of neurons in the LC. Thus, to further examine the cellular relationship between 5-HT-labeled axon terminals and excitatory amino acid receptors, we conducted immunogold-silver labeling of an antibody which recognized the three identified members of the kainate receptor (KAr) subunit class (GluR 5,6,7) and peroxidase localization of 5-HT. Similar proportions of 5-HT-labeled terminals (9%) were either apposed to KAr-labeled dendrites or exhibited KAr immunoreactivity. Twenty-four percent of the 5-HT axon terminals examined were apposed by glial processes that contained KAr. These data indicate that 5-HT axon terminals are in direct contact with LC neurons and also suggest pre- and postsynaptic sites for modulation of 5-HT terminals by excitatory amino acid ligands as well as indirect sites via glial processes. PMID- 10715567 TI - Cell culture as a model to study cell-cell interactions during development aging and neurodegenerative diseases. PMID- 10715568 TI - Induction of tissue plasminogen activator in differentiated NG108-15 cells. AB - Plasminogen activators may facilitate neurite outgrowth and neuronal migration in the developing nervous system. The expression of tissue plasminogen activator by NG108-15 neuroblastoma grown under a variety of conditions has been explored. High tissue plasminogen mRNA expression correlates with growth conditions which induce morphological differentiation and neurite outgrowth; however, NG108-15 cells grown in suspension with dibutyryl-cAMP also show a high level of tissue plasminogen activator expression. PMID- 10715569 TI - Heterogeneity of astroglia cultured from adult human temporal lobe. AB - This study characterized the morphological and electrophysiological diversity of astroglia cultured from adult human cerebral temporal lobe, and explored the influence of the cytokine interleukin-1beta on these cells. The cultures contained astroglia positive for glial fibrillary acidic protein which were flat, bipolar or multipolar in shape and variable in size. A subpopulation of the bipolar and multipolar cells was positive for S100 protein. The most striking feature of these cultures was the presence of glia with long (600 micrometer) processes with few branches or only terminal branches. Patch clamp recordings of the non-stellate process bearing cells revealed prominent inward Na(+) and transient and sustained outward K(+) conductances. Distinct differences in the relative proportion of these conductances were evident among cells but did not appear to be correlated with cell morphology. Treatment of cultures with interleukin-1beta for 96 h did not change total protein content, but increased the content of S100beta protein and decreased the content of glial fibrillary acidic protein. The findings indicate that cultures of adult human cerebrum contain subpopulations of morphologically and electrophysiologically pleomorphic glial fibrillary acidic protein positive astroglia, exhibit increased levels of the neurotrophic factor S100beta when exposed to interleukin-1beta, and may serve as a useful model for investigation of glial involvement in neuropathology. PMID- 10715570 TI - Association between cell swelling and glycogen content in cultured astrocytes. AB - Treatment of cultured rat astrocytes with hypotonic media or with 1 mM glutamate for 90 min caused cell swelling and a significant increase in glycogen content. Conversely, treatment with hypertonic media caused cell shrinkage with a corresponding decrease in astrocyte glycogen, which was proportional to the increasing osmolality of the hypertonic media. The glutamate receptor antagonist, MK-801, lowered both the glutamate-induced swelling and glycogen increase. These findings demonstrate a correlation between changes in cell volume and astrocyte glycogen content. This may explain the increased astrocytic glycogen observed in many neuropathological conditions where astrocyte swelling occurs. Because glycogen represents the largest energy reserve in the central nervous system, a swelling-induced disturbance in glycogen metabolism may lead to abnormal glial neuronal interactions resulting in impaired brain bioenergetics. PMID- 10715571 TI - In utero exposure to serotonergic drugs alters neonatal expression of 5-HT(1A) receptor transcripts: a quantitative RT-PCR study. AB - In embryonic rat brain, serotonin (5-HT) acts as a differentiation signal for 5 HT neurons and their target cells during midgestation. Serotonin receptors expressed during this period include the 5-HT(1A) subtype, which may mediate some of these developmental effects. Using the highly sensitive method of competitive RT-PCR, we quantified the effects of maternal treatment with either p chlorophenylalanine (pCPA; which depletes 5-HT in embryonic rat brain) or 5 methoxytryptamine (5-MT; a general 5-HT(1) /5-HT(2) agonist) from embryonic day E12-17 on expression of 5-HT(1A) receptor mRNA transcripts in brains of offspring at postnatal day 4 (PND 4). In offspring of both pCPA and 5-MT treated mothers, 5 HT(1A) transcripts were significantly reduced compared to vehicle controls, although effects of pCPA were greater than those of 5-MT. These results indicate that either under-stimulation of 5-HT(1A) receptors (due to pCPA-induced 5-HT depletion) or over-stimulation (by the agonist 5-MT) during prenatal development significantly reduced expression of 5-HT(1A) receptor transcripts in neonatal offspring. This may occur by disruption of 5-HT(1A) gene transcription or by post transcriptional mechanisms (such as altered translation or turnover of mRNA). Whatever the mechanism, reductions in 5-HT(1A) receptor transcripts following in utero exposure to serotonergic drugs could significantly impact the number of 5 HT(1A) receptors expressed in neonatal rat brain. Whether such effects will persist into adulthood remains to be determined. PMID- 10715572 TI - Repeated immunolesions display diminished stress response signal. AB - Cholinergic basal forebrain neurons (CBFNs) retrogradely transport neurotrophins released in the hippocampus and cortex as part of a general response to injury in a process that is impaired in the aged rodent and can be spared by the exogenous addition of pharmacological doses of nerve growth factor (NGF). This observation suggests that components of stress response signal transduction pathways in the aged CNS can be exogenously activated. The extent and mechanism of the endogenous stimulation of NGF in response to injury can be mimicked via treatment with 192 IgG-saporin of rat CNS, an immunolesion model. Here we report on the use of a conditioning lesion paradigm to determine if repeated partial immunolesions have a conditioning effect on the immunolesion-induced increases in NGF protein or decreases in choline acetyltransferase (ChAT) and acetylcholinesterase (AChE) activity. We report that chronic repeated immunolesions, as used here, were not as effective as a one time equivalent immunolesion in terms of induced NGF protein increases or decreasing ChAT and AChE activity in the hippocampus and cortex. Thus, chronic lesions resulting in cholinergic impairment typical of the aged CNS may differ from acute toxic models as a result of desensitization due to a conditioning effect of chronic subthreshold lesioning events in the CNS. PMID- 10715574 TI - The neural cell adhesion molecule in synaptic plasticity and ageing. AB - By mediating cell adhesion and signal transduction, the neural cell adhesion molecule (NCAM) regulates neurite outgrowth, fasciculation and target recognition in the developing nervous system. In addition, a number of studies suggest an important role for the NCAM in regeneration and learning in the adult nervous system. NCAM-deficient mice are impaired in spatial learning. Moreover, by interfering with normal NCAM function by intracranial injections of NCAM antibodies, long-term potentiation (LTP) in rat hippocampal slices and learning in rats and chicks have been inhibited. In the vertebrate nervous system, NCAM is the dominant carrier of polysialic acid (PSA), an unusual carbohydrate consisting of long homopolymers of sialic acid. The PSA-NCAM expression decreases markedly during development. However, an upregulation of polysialic acid (PSA) in restricted brain areas including the hippocampus has been observed following learning. Moreover, enzymatic removal of PSA results in impaired LTP and learning. In muscle, the PSA-NCAM expression is upregulated following denervation. This response is weakened in aging rats. The expression of NCAM and PSA have been shown to be regulated by neuronal activity suggesting that the NCAM may promote structural remodelling in an activity dependent manner associated with learning and regeneration. PMID- 10715573 TI - Differential alterations of NF-kappaB to oxidative stress in primary basal forebrain cultures. AB - Oxidative stress has been linked to neuronal cell death resulting from either acute insults due to ischemia, trauma, excitotoxicity, or chronic neurodegenerative diseases. Cholinergic basal forebrain neurons (CBFNs) compete for nerve growth factor (NGF) synthesized in the hippocampus and cortex via retrograde transport. NGF affects CBFN survival and cholinergic function via activation of the NF-kappaB transcription factor and this signaling pathway appears to be impaired in aged rats. Here, we demonstrate that activation of NF kappaB in basal forebrain primary culture via treatment with hydrogen peroxide or TNF-alpha is predominantly restricted to CBFNs, and that NF-kappaB activation appears to mostly affect p65 translocation to the nucleus, but not the p50 subunit. These results are consistent with NF-kappaB activation being a part of recovery processes after acute oxidative stress. Since p50 or p49 (also called p52) binding to promoter sites does not stimulate transcription - both p50 and p49 lack an activating domain - and p65 does contain an activating domain and thus can act as a transcription enhancer, differential translocation of different NF-kappaB dimers can act as repressors of constitutive activity or enhancers. These results are in agreement with the hypothesis that p50/p65 is the active trans-activating species of NF-kappaB, as compared to p50/p50 homodimers which bind to NF-kappaB binding sites but do not trans-activate promoters. Our results also suggest that selective activation of different NF-kappaB dimer species may have regulatory significance in neuronal responses to acute or chronic insults to CNS. Thus, increased p65 translocation could have enhancing effects while increased p50 translocation could have a repressor role. Manipulation of the types of NF-kappaB species being translocated could provide a basis for therapeutic strategies. PMID- 10715575 TI - Embryonic stem-cell derived neurones express a maturation dependent pattern of voltage-gated calcium channels and calcium-binding proteins. AB - There are remarkable changes of calcium binding proteins and voltage dependent Ca(2+) channel subtypes during in vitro differentiation of embryonic stem cell derived neurons. To observe these maturation dependent changes neurones were studied using combined immunohistochemical, patch clamp and videomicroscopic time lapse techniques. Embryonic stem cell derived neuronal maturation proceeds from apolar to bi- and multipolar neurones, expressing all Ca(2+) channel subtypes. There is, however, a clear shift in channel contribution to whole cell current from apolar neurones with mainly N- and L-type channel contribution in favour of P/Q- and R-type participation in bi- and multipolar cells. Expression of the calcium binding protein parvalbumin could be detected in bipolar, while calretinin and calbindin was preferentially found in multipolar neurones. Our data provides new insights into fundamental neurodevelopmental mechanisms related to Ca(2+) homeostasis, and clarifies contradictory reports on the development of Ca(2+) channel expression using primary cultures of neurones already committed to certain brain compartments. PMID- 10715576 TI - PSA-NCAM: an important regulator of hippocampal plasticity. AB - The Neural Cell Adhesion Molecule (NCAM) serves as a temporally and spatially regulated modulator of a variety of cell-cell interactions. This review summarizes recent results of studies aimed at understanding its regulation of expression and biological function, thereby focussing on its polysialylated isoforms (PSA-NCAM). The detailed analysis of the expression of PSA and NCAM in the hippocampal mossy fiber system and the morphological consequences of PSA-NCAM deficiency in mice support the notion that the levels of expression of NCAM are important not only for the regulation and maintenance of structural changes, such as migration, axonal growth and fasciculation, but also for activity-induced plasticity. There is evidence that PSA-NCAM can specifically contribute to a presynaptic form of plasticity, namely long-term potentiation at hippocampal mossy fiber synapses. This is consistent with previous observations that NCAM deficient mice show deficits in spatial learning and exploratory behavior. Furthermore, our data points to an important role of the hypothalamic-pituitary adrenal axis, which is the principle adaptive response of the organism to environmental challenges, in the control of PSA-NCAM expression in the hippocampal formation. In particular, we evidence an inhibitory influence of corticosterone on PSA-NCAM expression. PMID- 10715577 TI - Axonal sprouting regulates myelin basic protein gene expression in denervated mouse hippocampus. AB - The regulation of oligodendrocyte gene expression and myelination in vivo in the normal and injured adult CNS is still poorly understood. We have analyzed the effects of axotomy-induced axonal sprouting and microglial activation, on oligodendrocyte myelin basic protein (MBP) gene expression from 2 to 35 days after transection of the entorhino-hippocampal perforant path axonal projection. In situ hybridization analysis showed that anterograde axonal and terminal degeneration lead to upregulated oligodendrocyte MBP mRNA expression starting between day 2 and day 4, in (1) the deep part of stratum radiatum of CA3 and the dentate hilus, which display axonal sprouting but no degenerative changes or microglial activation, and (2) the outer part of the molecular layer of the fascia dentata, and in stratum moleculare of CA3 and stratum lacunosum-moleculare of CA1, areas that display dense anterograde axonal and terminal degeneration, myelin degenerative changes, microglial activation and axotomi-induced axonal sprouting. Oligodendrocyte MBP mRNA expression reached maximum in both these areas at day 7. MBP gene transcription remained constant in stratum radiatum, stratum pyramidale and stratum oriens of CA1, areas that were unaffected by perforant path transection. These results provide strong evidence that oligodendrocyte MBP gene expression can be regulated by axonal sprouting independently of microglial activation in the injured adult CNS. PMID- 10715578 TI - IL-1beta and ICE mRNA are not altered upon beta-amyloid(25-35) induced neurotoxicity in human neuroblastoma cells. AB - The specific beta-amyloid(25-35) fragment induced cellular degradation of the human neuroblastoma cell line SH-SY5Y, but did not elicit an effect on the levels of interleukin-1beta and interleukin-1beta converting enzyme, as determined by semiquantitative reverse transcription-polymerase chain reaction and immunocytochemical analysis. The assays revealed constitutive expression of these proteins both at mRNA and protein level. It is conceivable that in the absence of glial elements, such as in the present neuroblastoma cell line, beta-amyloid triggers the toxicity through a direct action and/or through the production of other harmful molecules. PMID- 10715579 TI - Mechanisms of apoptosis in embryonic cortical neurons (E6 and E7) in culture involve lipid signalling, protein phosphorylation and caspase activation. AB - Cultured embryonic (E7) chick neurons, derived from cerebral hemispheres, underwent apoptosis in response to inhibitors of protein kinase C (staurosporine) and phosphatidylinositol-3-kinase (wortmannin and LY294002), in a dose- and time dependent manner. This was monitored by loss of cell viability, increased DNA fragmentation, and activation of caspase-3-like activity, all of which were partially reversed by elevating the level of cAMP in the cells with Bt(2)cAMP or (Sp)cAMPS. Further studies revealed that an early step in apoptosis was the formation of ceramide from sphingomyelin, resulting from the activation of a neutral pH sphingomyelinase activity. Thus inhibitors of protein kinase C and phosphatidylinositol-3-kinase increased ceramide levels in the same time-frame as caspase-3 activation and DNA fragmentation. Neurons could also be killed by the addition of either water-soluble C2-ceramide (30 microM) or natural C22/24 ceramide (0.5 microM). In contrast to the apparent protective effect of ser/thr protein phosphorylation, a pro-apoptotic role for tyrosine phosphate phosphorylation was suggested by the ability of protein tyrosine phosphate phosphatase inhibitor, Bis(maltolato)oxovanadium (IV) (BMOV), to induce apoptosis in E7CH neurons. Thus BMOV (25 microM) killed 50% of E7CH neurons and B lymphocytes but not glial cells, or T-lymphocytes, suggesting the existence of a common apoptotic pathway in neurons and B-cells. We conclude that the major pathway for programmed cell death in embryonic chick neurons has many elements in common with that described for other cells but that there may be some unique aspects which can be used to protect embryonic neurons from opioid and other drug enhanced apoptosis. PMID- 10715580 TI - Oxidative stress-induced apoptosis of cochlear sensory cells: otoprotective strategies. AB - Apoptosis is an important process, both for normal development of the inner ear and for removal of oxidative-stress damaged sensory cells from the cochlea. Oxidative-stressors of auditory sensory cells include: loss of trophic factor support, ischemia-reperfusion, and ototoxins. Loss of trophic factor support and cisplatin ototoxicity, both initiate the intracellular production of reactive oxygen species and free radicals. The interaction of reactive oxygen species and free radicals with membrane phospholipids of auditory sensory cells creates aldehydic lipid peroxidation products. One of these aldehydes, 4-hydroxynonenal, functions as a mediator of apoptosis for both auditory neurons and hair cells. We present several approaches for the prevention of auditory sensory loss from reactive oxygen species-induced apoptosis: 1) preventing the formation of reactive oxygen species; (2) neutralizing the toxic products of membrane lipid peroxidation; and 3) blocking the damaged sensory cells' apoptotic pathway. PMID- 10715581 TI - Role of fibroblast growth factor-2 in human brain: a focus on development. AB - Trophic factors have gained a great degree of attention as regulators of neural cells proliferation and differentiation as well as of brain maturation. Very little is known, however, about their effects on human immature nervous system. In this paper, data on expression of fibroblast-growth factor-2 and its receptors are reviewed and discussed in the light of its possible role in human brain development. PMID- 10715582 TI - Excitotoxicity in neurological disorders--the glutamate paradox. AB - Beneficial effects of glutamate-receptor antagonists in models of neurological disorders are often used to support the notion that endogenous excitotoxicity (i.e. resulting from extracellular accumulation of endogenous glutamate) is a major contributor to neuronal death associated with these conditions. However, this interpretation conflicts with a number of robust and important experimental evidence. Here, emphasis is placed on two key elements: (i) very high extracellular levels of glutamate must be reached to initiate neuronal death, far above those measured in models of neurological disorders; and (ii) changes in extracellular glutamate as measured by microdialysis are not related to changes in the synaptic cleft, i.e. the compartment where neurotransmitter glutamate interacts with its receptors. It has become clear that the diversity and complexity of glutamate-mediated processes allow for a wide range of potential abnormalities (e.g. loss of selectivity of glutamate-operated ion channels, abnormal modulation of glutamate receptors). In addition, as neuronal death subsequent to ischemia and other insults is likely to result from multifactorial processes that may be inter-related, inhibition of glutamate-mediated synaptic transmission may be neuroprotective by increasing the resistance of neurons to other deleterious mechanisms (e.g. inadequate energy supply) that are not directly related to glutamatergic transmission. PMID- 10715583 TI - Protective action of 17beta-estradiol and tamoxifen on glutamate toxicity in glial cells. AB - Estrogens influence differentiation, growth and function of neurons, but less is known of their effects on glia. In our experiments reported here, the ovarian steroid, 17beta-estradiol, and the "designer", non-steroidal estrogen, tamoxifen, effectively protected C-6 glioma 2B clone cells from the cytotoxicity of the excitatory neurotransmitter, glutamate. Exposure of these cells to 10-20 mM glutamate induced 61-78% cell death. Pre-treatment of the cells with 0.01 mM estradiol or with 2 microM tamoxifen significantly reduced the glutamate-induced cell death, estradiol being the most effective in this regard. Estradiol- or tamoxifen-treated cells that had survived glutamate damage appeared more mature than controls. Thus, estrogens often used in therapy (estradiol as replacement after menopause and tamoxifen for treatment/prevention of breast cancer) may significantly protect glial cells against glutamate toxicity and stimulate cell differentiation. PMID- 10715584 TI - Excitatory amino acids and neurodegeneration: a hypothetical role of calcium precipitation. AB - Activation of excitatory amino acid (EAA) receptors can induce neurodegeneration by two major mechanisms of excitotoxicity, one related to the influx of Na(+), Cl(-) and water, and the other to the increase in intracellular calcium concentration ([Ca(2+)](i)). Thus, acute microinjection of EAAs in several areas of the central nervous system (CNS) has been used to produce neurodegenerative models. We studied the excitotoxic pattern associated with acute microinjection of AMPA in rat hippocampus, medial septum-diagonal band of Broca (MS-DBB), prefrontal cortex and retina. In all cases progressive neuronal loss, glial reaction and development of intra- and extracellular calcium concretions were observed. However, a CNS-area differential vulnerability was revealed, as shown by the specific atrophy of MS-DBB and its limited calcification. Whether calcium deposits are a defensive mechanism against the massive increment of free cytoplasmatic calcium is discussed on the basis of ultrastructural data and previous results. PMID- 10715585 TI - Depolarization-induced release of [(3)H]D-aspartate from GABAergic neurons caused by reversal of glutamate transporters. AB - Cultured neocortical neurons, which predominantly consist of GABAergic neurons exhibit a pronounced stimulus-coupled GABA release. Since the cultures may contain a small population of glutamatergic neurons and the GABAergic neurons have a high content of glutamate it was of interest to examine if glutamate in addition to gamma-aminobutyric acid (GABA) could be released from these cultures. The neurons were preloaded with [(3)H]D-aspartate and subsequently its release was followed during depolarization induced by a high potassium concentration or the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor agonists, AMPA and kainate. Depolarization of the neurons with 55 mM potassium increased the release of [(3)H]D-aspartate by more than 10-fold. When the non specific calcium-channel blockers cobalt or lanthanum were included in the stimulation buffer with potassium, the release of [(3)H]D-aspartate was decreased by about 40%. These results indicated that some of the released [(3)H]D-aspartate might originate from a vesicular pool. When AMPA was applied to the neurons, the release of [(3)H]D-aspartate was increased 2-fold and could not be prevented or decreased by addition of cobalt. Since AMPA has a rapid desensitizing effect on AMPA receptors, it was examined whether AMPA under non-desensitizing conditions was able to induce an increased release of [(3)H]D-aspartate as compared to the conditions of applying AMPA alone. The desensitization of AMPA receptors was blocked by 6-chloro-3,4-dihydro-3-(2-norbornen-5-yl)-2H-1,2, 4-benzothiadiazine-7 sulphonamide-1,1-dioxide (cyclothiazide). Under the non-desensitizing conditions, the AMPA-induced release of [(3)H]D-aspartate was highly enhanced showing about a 10-fold increase over basal release. Addition of cobalt or lanthanum did not decrease the amount of [(3)H]D-aspartate released, indicating that the release originated from a cytoplasmic pool. Kainate, which induces an almost non desensitizing effect on AMPA receptors, showed similar results as observed for AMPA under non-desensitizing conditions. The NMDA receptor antagonist (5R,10 S) (+)-5-methyl-10, 11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine (MK-801) had only minor effects on the [(3)H]D-aspartate release induced by AMPA and kainate. Thus, the depolarization-induced release of [(3)H]D-aspartate from cultured GABAergic neurons appears to be caused mainly by reversal of the glutamate transporters. PMID- 10715586 TI - Expression of the estrogen-regulated gene Nip2 during rat brain maturation. AB - The present study stems from previous observations demonstrating that in the neuroblastoma cell line SK-ER3 the mRNA content of the pro-apoptotic gene Nip2 is decreased following treatment with estradiol. We investigate the content of Nip2 mRNA during the maturation of rat embryo brain and we show that Nip2 mRNA is very low at embryo day 15 and steadily increases up to day 20. At day 21 Nip2 mRNA is decreased almost to the low levels observed in the mature brain. Studies in neurons from rat embryo at day 18 show that Nip2 mRNA content is significantly decreased by exposure to estradiol at 1 nM concentration demonstrating that the observations previously done in the SK-ER3 neuroblastoma cell line can be reproduced in neurons in culture. The finding that estrogens may modulate the activity of Nip2 gene activity may be of relevance not only with regard to the effects of estradiol on brain maturation, but also for the understanding of the neuroprotective effects recently described for this hormone. PMID- 10715587 TI - Early expression of the BM88 antigen during neuronal differentiation of P19 embryonal carcinoma cells. AB - Previous studies have shown that the BM88 antigen, a neuron-specific molecule, promotes the differentiation of mouse neuroblastoma cells [23] (Mamalaki A., Boutou E., Hurel C., Patsavoudi E., Tzartos S. and Matsas R. (1995) The BM88 antigen, a novel neuron-specific molecule, enhances the differentiation of mouse neuroblastoma cells. J. Biol. Chem. 270, 14201-14208). In particular, stably transfected with the BM88 cDNA, Neuro 2a cells over-expressing the BM88 antigen are morphologically distinct from their non-transfected counterparts; they exhibit enhanced process outgrowth and a slower rate of division. Moreover, they respond differentially to growth factors [10] (Gomez J., Boutou E., Hurel C., Mamalaki A., Kentroti S. , Vernadakis A. and Matsas R. (1998) Overexpression of the neuron-specific molecule BM88 in mouse neuroblastoma cells: Altered responsiveness to growth factors. J. Neurosci. Res. 51, 119-128). In order to further elucidate the role of the BM88 antigen in the differentiation of developing neurons we used the in vitro system of differentiating P19 cells which closely resembles early murine development in vivo. In this study, P19 cells were driven to the neuronal pathway with retinoic acid. We examined by immunofluorescence studies the expression of the BM88 antigen in these cells and we found that it correlates well with the expression of the polysialylated form of the neural cell adhesion molecule (PSA-NCAM) which characterizes early differentiating post-mitotic neurons. In contrast, very few of the BM88 antigen positive/PSA-NCAM-positive cells expressed neurofilament protein, a marker of more mature neurons. Our findings, in accordance with previously reported data, strongly suggest that the BM88 antigen is involved in the early stages of differentiation of neuronal cells. PMID- 10715589 TI - A comparison of iron availability from commercial iron preparations using an in vitro digestion/Caco-2 cell culture model. AB - The objectives of this study were to compare iron availability from commercial preparations of FeSO(4), ferrous gluconate, ferrous fumarate, and a polysaccharide-iron complex using an in vitro digestion/Caco-2 cell culture model. In addition, we sought to determine if calcium carbonate and calcium acetate (common phosphate binding agents) inhibited iron availability from an oral iron supplement when digested simultaneously. Caco-2 cell ferritin formation following exposure to simulated gastric and intestinal digests of the iron supplements was used as a measure of iron uptake and availability. Plates without cell monolayers were included in each replication of the experiment to measure the total amount of soluble iron that resulted from the in vitro digestion. Significantly more iron was taken up from the FeSO(4), ferrous gluconate, and ferrous fumarate than the polysaccharide-iron complex. Similar results comparing FeSO(4) and the polysaccharide-iron complex have been observed in humans. In addition, less iron was taken up from digests with calcium carbonate relative to calcium acetate even though similar amounts of soluble iron were observed in these experiments. The results indicate that when iron supplements and phosphate binders are consumed simultaneously, calcium acetate may be the preferred phosphate binder to maximize iron availability. PMID- 10715588 TI - Expression of the glucocorticoid receptor in early and late passage C-6 glioma cells and in normal astrocytes derived from aged mouse cerebral hemispheres. AB - The presence of the glucocorticoid receptor in early and late passage C-6 glioma cells 2B clone and in astrocytes derived from aged mouse cerebral hemispheres has been documented by immunoblotting and/or immunofluorescence labelling. All cell types studied express the glucocorticoid receptor of molecular weight 97 KDa. In addition, in astrocytes derived from aged mouse cerebral hemispheres a smaller molecular weight polypeptide reacting with anti-glucocorticoid receptor antibody was also demonstrated. No difference in the amount of the 97 KDa glucocorticoid receptor between early and late C-6 2B cells was observed, whereas the astrocytes from aged cerebral hemispheres contained considerably reduced amounts of the glucocorticoid receptor compared to C-6 2B cells. Late passage C-6 2B cells were immunofluorescence labelled with the anti-glucocorticoid antibody, the receptor being almost exclusively present in the cytoplasm, with particular concentration in the perinuclear region. The presence of glucocorticoid receptor of molecular weight 97 KDa in glial cells corroborates and expands the existing data based on radioligand binding and immunocytochemical studies. These cell populations can be exploited as a model system for the study of the effects of glucocorticoids on senescence and brain aging. PMID- 10715590 TI - Hypocholesterolemic effect of anhydrous milk fat ghee is mediated by increasing the secretion of biliary lipids. AB - The anhydrous milk fat ghee is one of the important sources of fat in the Indian diet. Our earlier studies showed that rats fed diets containing greater than 2.5 wt% of ghee had lower levels of serum cholesterol compared with rats fed diets containing groundnut oil. To evaluate the mechanism of the hypocholesterolemic effect of ghee, male Wistar rats were fed a diet containing 2.5 or 5.0 wt% ghee for a period of 8 weeks. The diets were made isocaloric with groundnut oil. Both native and ghee heated at 120 degrees C containing oxidized lipids were included in the diet. The ghee in the diet did not affect the 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase activity in the liver microsomes, but it significantly increased biliary excretion of cholesterol, bile acids, uronic acid, and phospholipids. The rats fed ghee had lower levels of cholesterol esters in the serum as well as in the intestinal mucosa. Both native and oxidized ghee influenced cholesterol metabolism. These results indicate that supplementation of diets with ghee lipids would increase the excretion of bile constituents and lower serum cholesterol levels. PMID- 10715591 TI - Beer increases plasma antioxidant capacity in humans. AB - The positive association of a moderate intake of alcoholic beverages with a low risk for cardiovascular disease, in addition to ethanol itself, may be linked to their polyphenol content. This article describes the effect of acute ingestion of beer, dealcoholized beer, and ethanol (4.5% v/v) on the total plasma antioxidant status of subjects, and the change in the high performance liquid chromatography profile of some selected phenolic acids (caffeic, sinapic, syringic, and vanillic acids) in 14 healthy humans. Plasma was collected at various times: before (T0), 1 hour after (T1), and 2 hours after (T2) drinking. The study is part of a larger research planned to identify both the impact of brewing on minor components potentially present in beer and their metabolic fate in humans. Beer was able to induce a significant (P < 0.05) increase in plasma antioxidant capacity at T1 (mean +/- SD: T0 1,353 +/- 320 microM; T1 1,578 +/- 282 microM), returning close to basal values at T2. All phenolic acids measured in plasma tended to increase after beer intake (20% at T1, 40% at T2). Syringic and sinapic acid reached statistical significance (P < 0.05 by one-way analysis of variance-Fisher's test) at T1 and T2, respectively. Plasma metabolic parameters (glucose, total cholesterol, triglycerides, and uric acid) and plasma antioxidants (alpha tocopherol and glutathione) remained unchanged. Ethanol removal impaired the absorption of phenolic acids, which did not change over the time of the experiment, accounting for the low (and not statistically significant) increase in plasma antioxidant capacity after dealcoholized beer drinking. Ethanol alone did not affect plasma antioxidant capacity or any of the antioxidant and metabolic parameters measured. PMID- 10715592 TI - Participation of blood cells in the changes of blood amino acid concentrations during maximal exercise. AB - We determined the participation of the cellular compartment in the changes of plasma amino acid concentrations during maximal exercise test on a cycle ergometer. Following an overnight fast, male athletes were submitted to a maximal exercise test until fatigue (for 25 min approximately) to determine maximal oxygen uptake. The amino acid concentrations in total blood, plasma, and blood cells were determined before and after the maximal exercise test. Most essential amino acids were decreased significantly in the total blood concentration as a result of the maximal exercise test. However, the concentrations of most nonessential amino acids tended to be significantly increased. Amino acid concentration was increased most in plasma. Concentrations of blood cell alanine and proline were significantly increased by 26% and 15%, respectively, after the maximal exercise test. No significant differences in blood cell concentrations of other amino acids induced by the exercise test were found, although the amount of tryptophan in blood cells was increased after exhaustive exercise. PMID- 10715593 TI - A comparison of extrahepatic lipogenesis from a small glucose meal in obob and gold thioglucose obese mice fed low- or high-fat diets with or without the addition of Delta22-5beta-taurocholenic acid. AB - Extrahepatic fatty acid synthesis from a 250 mg meal of [U-(14)C]-glucose was measured in epidymal fat pads and the remaining carcass of hyperglycemic obese (obob), gold thioglucose obese, and nonobese controls under conditions of maximum and minimum lipogenesis. Also assessed was the effect of Delta(22)-5beta taurocholenic acid, previously shown to inhibit hepatic fatty acid synthesis. Both types of obese and nonobese mice were fed for 6 weeks glucose-based diets containing either 1% corn oil or 40% lard with or without the addition of 0.05% taurocholenic acid. In mice fed 1% corn oil, incorporation of labeled glucose into carcass fatty acids was 25% greater in nonobese than obese mice of either type of obesity. On this diet incorporation of labeled glucose into epididymal fatty acids was reduced 83% in hyperglycemic obese mice compared with nonobese littermates. The corresponding reduction in lipogenesis in gold thioglucose obese mice was only 23% compared with nonobese controls. Feeding 40% lard reduced incorporation of labeled glucose into epididymal and carcass fatty acid 67 to 95% compared with mice fed 1% corn oil in both types of obese and nonobese mice whether or not taurocholenic acid had been fed. Both types of obesity or feeding 40% lard reduced lipogenesis in fat pads to a greater extent than glucose uptake by the pads with the reductions additive. Feeding taurocholenic acid reduced pad weight 30% across strain and obesity status, increased uptake of labeled glucose into epididymal fat pads and increased the percentage of the labeled glucose in the pad recovered as fatty acid in both types of obese and nonobese mice when the diet was 1% corn oil. Similarities and differences between the two obesity models are discussed. PMID- 10715594 TI - Identification of lactoperoxidase in mature human milk. AB - Myeloperoxidase (MPO) derived from milk leukocytes and lactoperoxidase (LPO) secreted from the mammary gland have been identified previously in human colostrum. These peroxidases are known to play host defensive roles through antimicrobial activity. The goals of this study were to measure the peroxidase activity in mature human milk and to characterize the enzyme responsible for the activity. As determined using 3,3',5,5'-tetramethylbenzidine as substrate, whey prepared from human milk samples obtained 1 and 5 months postpartum showed levels of peroxidase activity equivalent to 0.13 +/- 0.18 and 0.24 +/- 0.21 microg/mL bovine LPO (bLPO; n = 13), respectively. Whey from early milk was fractionated into two peaks of peroxidase activity by cation-exchange chromatography; the peroxidase in the first peak was sensitive to dapsone, which is an inhibitor of LPO, whereas the second peroxidase was not. Whey from mature milk showed only the first peak. Purified bLPO and MPO showed chromatographic behaviors that were similar to the first and second peaks, respectively. The dapsone-sensitive peroxidase from mature milk was further purified (952-fold from whey) by hydrophobic interaction chromatography. This preparation showed two bands with molecular masses of 80 and 90 kDa by polyacrylamide gel electrophoresis and immunoblotting using an antibody against bLPO. After deglycosylation, two distinct proteins with lower molecular weights were observed. Amino acid sequencing indicated that both of these proteins are LPO. These results provide evidence that LPO is present in mature human milk and that it is responsible for most of the peroxidase activity in mature milk. PMID- 10715595 TI - Influence of magnesium deficiency on the bioavailability and tissue distribution of iron in the rat. AB - We investigated the effect of dietary magnesium (Mg) deficiency on the nutritive utilization and tissue distribution of iron (Fe). Wistar rats were fed an Mg deficient diet (56 mg/kg) for 70 days. Absorbed Fe, Fe balance, number of the erythrocytes [red blood cells (RBC)] and leukocytes white blood cells (WBC)], hemoglobin (Hb), and Fe content were determined in samples of plasma, whole blood, skeletal muscle, heart, kidney, liver, spleen, femoral bone, and sternum obtained on experimental days 21, 35, and 70. The Mg-deficient diet significantly increased Fe absorption and Fe balance from week 5 until the end of the experimental period. This effect was accompanied by a significant decrease in the concentration of RBC and Hb from day 35, which caused the decrease in whole blood Fe seen on day 70. However, WBC were significantly increased from day 21 until the end of the experimental period. Mg deficiency significantly increased plasma and liver Fe at all three time points investigated. Spleen, heart, and kidney Fe were significantly increased only at the end of the study. However, on day 70, Fe concentration in the sternum had decreased significantly. No changes were found in skeletal muscle or femur Fe content. Mg deficiency led to increased intestinal absorption of Fe and decreased RBC counts, possibly as a result of increased fragility of the erythrocytes. Intestinal interactions between Fe and Mg, together with activation of erythropoiesis as a result of hemolysis, favored intestinal absorption of Fe. This situation gave rise to an increase in plasma Fe levels, which in turn favored Fe uptake and storage by different organs, especially the liver and spleen. However, despite the increased Fe content seen in the tissues of rats fed the Mg-deficient diet, these animals were unable to compensate for the hemolysis caused by this nutritional deficiency. PMID- 10715596 TI - Piperine derived from black pepper increases the plasma levels of coenzyme Q10 following oral supplementation. AB - An extract from the fruits of black pepper consisting of a minimum of 98% pure piperine was evaluated in a clinical study using a double-blind design. The relative bioavailability of 90 mg and 120 mg of coenzyme Q10 administered in a single-dose experiment or in separate experiments for 14 and 21 days with placebo or with 5 mg of piperine was determined by comparing measured changes in plasma concentration. The inter-subject variability was minimized by limiting the selection of individuals to healthy adult male volunteers with (presupplementation) fasting coenzyme Q10 values between 0.30 and 0.60 mg/L. The results of the single-dose study and the 14-day study indicate smaller, but not significant, increases in plasma concentrations of coenzyme Q10 in the control group compared with the group receiving coenzyme Q10 with a supplement of piperine. Supplementation of 120 mg coenzyme Q10 with piperine for 21 days produced a statistically significant (p = 0.0348), approximately 30% greater, area under the plasma curve than was observed during supplementation with coenzyme Q10 plus placebo. It is postulated that the bioenhancing mechanism of piperine to increase plasma levels of supplemental coenzyme Q10 is nonspecific and possibly based on its description in the literature as a thermonutrient. PMID- 10715597 TI - Antioxidant effect of red wine polyphenols on red blood cells. AB - The protective effect of red wine polyphenols against hydrogen peroxide (H(2)O(2))-induced oxidation was investigated in normal human erythrocytes (RBCs). RBCs, preincubated with micromolar amounts of wine extract and challenged with H(2)O(2), were analyzed for reactive oxygen species (ROS), hemolysis, methemoglobin production, and lipid peroxidation. All these oxidative modifications were prevented by incubating the RBCs with oak barrel aged red wine extract (SD95) containing 3.5 mM gallic acid equivalent (GAE) of phenolic compounds. The protective effect was less apparent when RBCs were incubated with wines containing lower levels of polyphenols. Furthermore, resveratrol and quercetin, well known red wine antioxidants, showed lower antioxidant properties compared with SD95, indicating that interaction between constituents may bring about effects that are not necessarily properties of the singular components. Our findings demonstrate that the nonalcoholic components of red wine, mainly polyphenols, have potent antioxidant properties, supporting the hypothesis of a beneficial effect of red wine in oxidative stress in human system. PMID- 10715598 TI - Strategies for the assessment of acute skin irritation potential. PMID- 10715599 TI - The tachycardia-induced dog model of atrial fibrillation. clinical relevance and comparison with other models. AB - In the past, investigators have relied extensively on acute in vivo models of atrial fibrillation (AF), in which AF was induced either pharmacologicly or by vagal stimulation. More recently, there is a need and desire for more clinically relevant models that can only be achieved with the use of chronically instrumented animals. One of these models is the atrial tachycardia-induced AF dog model, which is the main focus of this review. The model produces a persistent AF in 80% of animals paced at 400 beats/min for 6 weeks. Atrial tachycardia also induces various pathophysiologic and ultrastructural changes that often resemble electrical remodeling of atria in patients that have a high susceptibility to AF. This model can also be used to evaluate drug efficacy with respect to attenuation of AF duration or conversion of AF to sinus rhythm. The model may therefore be used to provide further insights into the discovery of new therapeutic approaches to modifying this atrial arrhythmic disorder in man. PMID- 10715600 TI - Pharmacologic evaluation of isometric contraction-relaxation coupling indexes in rabbit ventricular muscle. AB - Investigations of the coupling between contraction and relaxation (contraction relaxation [CRC] process) in isometric conditions are essential in determining whether pharmacologic interventions or cardiac diseases specifically modify isometric relaxation (intrinsic lusitropic effect) or change it in proportion with the accompanying changes in contractility (or inotropy). For this purpose, the CRC process is quantified by various indexes, derived from differentiation and/or curve fitting the whole or relaxation phase of the isometric twitch, one of the most used being tau, the time constant of the final iso(volu)metric phase of relaxation. Nevertheless, the possible redundancy and validity of such indexes have not been thoroughly investigated. Accordingly, we performed a pharmacologic evaluation of such indexes in isolated rabbit ventricular muscles isometrically contracting in vitro, using modifiers of either intracellular Ca(2)+ handling (nifedipine, ryanodine, 2,5-di-tert-butyl-benzohydroquinone, all negative inotropic compounds, and BAY K 8644, a positive inotropic drug), or myofibrillar Ca(2)+ sensitivity (CGP 48506, a Ca(2)+ sensitizer, and butanedione monoxime, a Ca(2)+ desensitizer, respectively positive and negative inotropic compounds). The isometric twitch in control conditions and in the presence of increasing concentration of each compound was analyzed to determine the classically used CRC and/or lusitropic indexes, derived either from single parameters such as the maximal rate or contraction and relaxation (+dT(max) and -dT(max), respectively), or from curve fitting of the whole, or part, of the twitch. As the rate of isometric relaxation is dependent on myofilament properties, we expected that compounds modifying myofibrillar Ca(2)+ sensitivity in an opposite direction (CGP 48506 vs butanedione monoxime) would be the only drugs exerting an intrinsic lusitropic and opposite effect on a validated CRC index. Results showed that (1) none of the tested compounds affected the slope of the linear relationship between peak twitch tension and dT(max), a previously assumed CRC index, sensitive only to myofibrillar Ca(2)+ sensitivity modifiers; (2) the lusitropic parameter B, derived from mathematical curve fitting of the whole isometric twitch, and the ratio +dT(max)/dT(max), exhibited similar drug- and dose dependency, but no opposite sensitivity to CGP 48506 and BDM for either index; and (3) negative inotropic compounds dose-dependently slowed relaxation (and conversely for positive inotropes), whether the latter was quantified by the rate constant beta, derived from double exponential curve fitting of the whole relaxation phase, or by the time constants tau(L) and tau(E), derived from the curve fitting (logistic and monoexponential, respectively) of the final phase of relaxation. Nevertheless, the pharmacologicly induced changes in beta were statistically significant at lower concentrations and exhibited less individual variability, compared with the time constants. We demonstrate that intrinsic lusitropic changes can be quantified by the value of the slope of the relationship relating beta to peak isometric tension: the slope value was unchanged by Ca(2)+ handling modifiers, decreased by CGP 48506, and reversed by BDM (indicating number, negative, and positive intrinsic lusitropic effects respectively). Based on these data, we propose that the linear relationship between beta and peak isometric tension could be used a new method to assess whether pharmacologic interventions or cardiac diseases exert intrinsic effects on isometric relaxation. PMID- 10715601 TI - An investigation of the relationship between the liver and brain using an isolated perfused rat brain preparation. AB - The pathogenesis of portal-systemic encephalopathy (PSE) and hepatic encephalopathy (HE), disorders of the brain attributed to abnormal liver function, are poorly understood. This study was conducted to examine if a fundamental, and possibly exclusive, homeostatic interrelationship exists between the liver and brain that might deteriorate during liver failure to result in the syndrome of PSE and HE. An isolated organ perfusion circuit was devised to accommodate an isolated rat brain and an isolated rat liver preparation perfused concomitantly. The survival time of the brain preparation was measured by the maintenance of the spontaneous electroencephalocorticogram and was extended from a median survival time of 35 min (range 22 to 53 min), when perfused alone, to 210 min (range 172 to 480 min), when perfused simultaneously with a liver. Also, concomitant perfusion with an isolated rat liver reduced perfusate glucose concentrations from 200 mg% to a range between 45 to 60 mg%. These data support our hypothesis that a homeostatic interrelationship exists between the liver and brain that is independent of other metabolic influences; disturbance of this relationship may contribute towards the syndrome of PSE and HE. PMID- 10715602 TI - Dexamethasone enhances the activity of rSP-C surfactant but not of exosurf in a rat model of the acute lung injury. AB - The possible enhancement of surfactant activity by pretreatment with a glucocorticosteroid (dexamethasone) was investigated in a rat lung lavage model of acute lung injury. Animals received a dose of dexamethasone (10 mg/kg i.p.) prior to the protein-free surfactant preparation Exosurf (pure phospholipids containing surfactant, Wellcome GmbH, Burgwedel, Germany) and a rSP-C based surfactant, respectively. Both surfactants were intratracheally instilled at doses of 25 and 100 mg phospholipids per kg body weight and were compared with pretreatment with dexamethasone at each dose level. These groups were also compared with untreated controls and to pretreatment with dexamethasone alone with respect to improvements in oxygenation, to inhibition of infiltration of polymorphonuclear neutrophil leukocytes (PMNL) and to influence formation of hyaline membranes. Dexamethasone alone had no influence on the reduced PaO(2) but reduced the infiltration of PMNL and the formation of hyaline membranes. Dexamethasone improved the oxygenation at both doses of rSP-C surfactant. At the low dose of rSP-C surfactant there were additional effects detectable with regard to histopathologic improvements. In contrast, dexamethasone had no additional effect on oxygenation and formation of hyaline membranes when combined with Exosurf. Only the infiltration of PMNL was decreased by combined treatment with dexamethasone and Exosurf. The effect was comparable to that of pretreatment with dexamethasone alone. In this animal model, pretreatment with dexamethasone showed additional effects on rSP-C surfactant that were superior to each treatment alone. From the comparison of rSP-C surfactant with the synthetic surfactant preparation Exosurf, we conclude that the activity of Exosurf cannot be improved substantially by additional pretreatment with drugs like glucocorticosteroids. PMID- 10715603 TI - An updated two-dimensional gel electrophoresis technique for the detection of drug-induced changes in protein phosphorylation in intact smooth muscle. AB - Two-dimensional gel electrophoresis is widely used in many areas of scientific research. The necessity for greater resolution and more sensitive protein detection with this technique have resulted in a steadily changing methodology. Complete descriptions of some aspects of two-dimensional gel electrophoresis are available in the earlier literature. However, simplified methods incorporating recent advances specifically designed to use two-dimensional gel electrophoresis for the measurement of protein phosphorylation in intact tissue are lacking. This report describes, in detail, each of the steps involved in carrying out such measurements including intact tissue labeling with 32P, homogenization and protein sample preparation, two-dimensional gel electrophoresis using isoelectric focusing followed by vertical second-dimension SDS-PAGE, staining, autoradiography, and quantitative analysis of changes in phosphorylation of specific proteins. This method incorporates a number of modifications taken from other published sources and includes several novel changes developed in our laboratory. To illustrate the utility of this technique we have included a set of results analyzing the phosphorylation patterns induced by the addition of a nitric oxide donor, sodium nitroprusside, to intact strips of rat aorta. We were able to demonstrate SNP-induced phosphorylation of a number of proteins, several of which have not been previously described in earlier reports in which the patterns of PKG-mediated phosphorylation were investigated. PMID- 10715606 TI - Hormesis: interpreting the beta-curve using control theory. AB - Data from experiments exposing colonial hydroids to toxic growth inhibitors have provided evidence of growth control mechanisms that respond adaptively to counter toxic inhibition. Analysis of growth data and the development of simulation models provide an interpretation of both alpha- and beta-curves. The hypothesis also suggests that hormesis is related to adaptation by growth control mechanisms that confer tolerance to subsequent exposure. PMID- 10715607 TI - Hormesis: an adaptive expectation with emphasis on ionizing radiation. AB - Non-linear fitness gradients with maxima between extremes are expected for any environmental variable to which free-living populations are exposed. For exceedingly toxic agents, including ionizing radiation, such deviations from linearity are close to zero exposure and are conventionally called hormesis. Accordingly, hormesis is an extreme version of the non-linear fitness gradients for general environmental stresses such as temperature fluctuations, for which maximum fitness occurs at the moderate temperature fluctuations to which free living populations are most commonly exposed. Some metabolic reserves should occur under moderate temperature stresses because of the need for pre-adaptation enabling survival during exposure to occasional periods of more extreme stress, especially at species borders where selection for stress resistance is likely to be most intense. Because heat shock proteins are induced by all stresses, adaptation to extreme temperatures should translate into adaptation to other stresses. Consequently, metabolic reserves from adaptation to extreme temperatures in the past should translate into protection from correlated abiotic stresses, especially in human populations where modern cultural processes are now ameliorating exposure to extreme stresses. Ambient and man-made radiations of non catastrophic dimensions should therefore lead to stress-derived radiation hormesis. Other stresses can, in principle, be incorporated into this model. Accordingly, evolutionary and ecological considerations suggest two components of hormesis in relation to ionizing radiation: background radiation hormesis based upon the background exposure to which all organisms on earth are subjected; and stress-derived radiation hormesis. Exposure under stress-derived radiation hormesis is considerably larger than under background radiation hormesis, so significant deleterious effects from non-catastrophic radiation normally may be impossible to detect. Suggestions are provided for testing such postulated deviations from the commonly assumed linear no-threshold (LNT) hypothesis for the biological consequences of exposure to radiation. PMID- 10715608 TI - Hazard assessment of chemical carcinogens: the impact of hormesis. AB - The recent report of reductions in the number and area of preneoplastic hepatic lesions in response to low doses of the tumor promoter phenobarbital provides important new support for the existence of hormetic responses to carcinogens. The presence of hormetic responses to carcinogenic agents and the corollary that beneficial doses of these compounds can be determined have several implications for the bioassay and hazard assessment of carcinogens as well as for public policy regulating exposure to these agents. To be adequately sensitive to detect and quantify hormetic or other non-linear dose-response functions, current study designs must be modified to include lower doses and sufficiently large numbers of animals. Short- or medium-term animal studies are a cost-effective means of addressing these needs and have been used recently to describe a classical hormetic response to the non-genotoxic carcinogen phenobarbital. These basic changes should be supported by a continuing emphasis on mechanistic research and the development of biologically based quantitative models of toxicant action. Linking these models with physiologically based pharmacokinetic model descriptions of target dose holds the greatest promise for improving the description of the dose-response curve at low doses. These approaches are generally encouraged by the USEPA in the form of The 1996 Proposed Carcinogen Risk Assessment Guidelines. However, there remain substantial questions regarding integration of the concept of hormesis into hazard testing and public policy that require careful consideration. Herein, we explore the issues that surround testing for hormetic responses and the implications for public policy. PMID- 10715609 TI - Defining the baseline for inhibition concentration calculations for hormetic hazards. AB - The use of endpoint estimates based on modeling inhibition of test organism response relative to a baseline response is an important tool in the testing and evaluation of aquatic hazards. In the presence of a hormetic hazard, the definition of the baseline response is not clear because non-zero levels of the hazard stimulate an enhanced response prior to inhibition. In the present study, the methodology and implications of how one defines a baseline response for inhibition concentration estimation in aquatic toxicity tests was evaluated. Three possible baselines were considered: the control response level; the pooling of responses, including controls and all concentration conditions with responses enhanced relative to controls; and, finally, the maximal response. The statistical methods associated with estimating inhibition relative to the first two baseline definitions were described and a method for estimating inhibition relative to the third baseline definition was derived. These methods were illustrated with data from a standard aquatic zooplankton reproductive toxicity test in which the number of young produced in three broods of a cladoceran exposed to effluent was modeled as a function of effluent concentration. PMID- 10715610 TI - Reference dose (RfD): the possible impact of hormesis. AB - A fictitious US Environmental Protection Agency Administrator in the year 2005 is confronted with making a risk management decision on a chemical that exhibits an unambiguous hormetic effect in an animal system. Dose-response curves for average humans and sensitive humans are derived from the animal data. The question is posed: What should the reference dose (RfD) be in this case? A series of outstanding scientific and policy questions are discussed that have a bearing on the answer. The concept of the 'regulatory dose (RgD)' is revived to address, if not resolve, the issue. PMID- 10715611 TI - Ecological risk assessment: implications of hormesis. AB - Hormesis is a widespread phenomenon across many taxa and chemicals, and, at the single species level, issues regarding the application of hormesis to human health and ecological risk assessment are similar. For example, convincing the public of a 'beneficial' effect of environmental chemicals may be problematic, and the design and analysis of laboratory studies may require modifications to detect hormesis. However, interpreting the significance of hormesis for even a single species in an ecological risk assessment can be complicated by considerations of competition with other species, predation effects, etc. Ecological risk assessments involve more than a single species; they may involve communities of hundreds or thousands of species as well as a range of ecological processes. Applying hormetic adjustments to threshold effect levels for chemicals derived from sensitivity distributions for a large number of species is impractical. For ecological risks, chemical stressors are frequently of lessor concern than physical stressors such as habitat alteration or biological stressors such as introduced species, but the relevance of hormesis to non chemical stressors is unclear. Although ecological theories such as the intermediate disturbance hypothesis offer some intriguing similarities between chemical hormesis and hormetic-like responses resulting from physical disturbances, mechanistic explanations are lacking. Further exploration of the relevance of hormesis to ecological risk assessment is desirable. Aspects deserving additional attention include developing a better understanding of the hormetic effects of chemical mixtures, the relevance of hormesis to physical and biological stressors and the development of criteria for determining when hormesis is likely to be relevant to ecological risk assessments. PMID- 10715612 TI - Hormesis: policy implications. AB - Protecting workers and the public from toxic chemicals, particularly carcinogens, has been a principal focus of public policy. Uncertainty regarding the toxicity of particular chemicals and their dose-response relationship has led to the use of the 'precautionary principle' in which regulators are willing to accept more costly regulation than necessary in order to prevent exposure and disease from these toxic chemicals. The Environmental Protection Agency's (EPA's) current policy of using 'mechanism of action' to set regulations means that hormesis could be used by the EPA without any change in policy if hormesis is accepted as scientifically valid. Hormesis could result in a qualitative change in regulatory policy. Because exposure to toxic chemicals conveys no health benefit in the current dose-response model, public risk aversion leads to a Delaney Clause-like 'no-risk' model for policy: ban toxic chemicals or lower exposure to trivial levels. Hormesis implies that individuals benefit from low exposure to toxicants. Although hormesis may not be relevant for individuals with compromised immune systems, it would be expected to help the vast majority of people. If so, permitting exposure levels that provided the greatest health benefit to most people would be balanced against these same levels hurting the most immune compromised individuals. Public health routinely makes these trade-offs using a 'risk-risk' model. Thus, hormesis could transform the 'no-risk' approach into a 'risk-risk' approach that could tolerate much higher exposures to toxic chemicals than the current policy. PMID- 10715613 TI - Risk assessment and risk management implications of hormesis. AB - International and US radiation protection standards are based upon risk assessment and risk management processes. The assessment of radiation risk is derived from the linear no-threshold (LNT) model. Risk management is based on more subjective value judgements. If the radiation dose-response was found to be hormetic, considerable quantitative data would be needed before current radiation protection standards would change. There would be added complexity, and consideration might have to be given to the additive effects of an individual's exposures to medical radiation and other potential carcinogens. PMID- 10715614 TI - Environmental law applications of hormesis concepts: risk assessment and cost benefit implications. AB - This article focuses on legal structures that influence the degree to which hormesis can be incorporated into environmental law and policy. Three statutes the Occupational Safety and Health Act, the Food Quality Protection Act, and the Clean Air Act-are used to illustrate the varied ways in which Congress, agencies and the courts have approached risk assessment and cost-benefit analyses that are relevant to the hormesis issue. This discussion features several examples of regulations and judicial decisions that have begun to recognize hormetic effects. The article concludes that hormesis concepts could be incorporated effectively into present risk assessment and cost-benefit mechanisms. In the context of agency action, an express policy decision might be made to broaden the typical scope of risk assessment and cost-benefit processes by including hormetic effects. In the judicial context, recognition of hormesis may occur where relevant statutory language is read to contemplate that an agency will consider both the beneficial and the detrimental effects of a particular substance in formulating regulations; in this circumstance, a reviewing court could reverse an agency decision that focuses solely on detrimental effects and ignores hormetic effects. Based on these evolving trends, the time may be ripe to seek further incorporation of hormesis concepts into environmental law and policy decisions. PMID- 10715615 TI - Chemical hormesis in cell growth: a molecular target at the cell surface. AB - A multifunctional ubiquinol (NADH) oxidase with protein disulfide-thiol interchange activity of the cell surface, abbreviated as NOX, is described as a molecular target for chemical hormesis of cell growth. The activity of the NOX correlates with rate of cell enlargement, which helps to determine how rapidly cells will divide. When NOX activity is inhibited, cells fail to enlarge following division and the result is a population of small cells unable to reach the minimum size required for them to divide again. In plants, cells fail to enlarge when NOX activity is inhibited. When NOX activity is stimulated or constitutively activated, as in cancer, cells enlarge more rapidly and the rate of cell division also is enhanced. Both cell growth and NOX activity are sometimes stimulated by low concentrations of normally inhibitory molecules. These properties define chemical hormesis, making the NOX molecule a molecular target to explain hormetic growth responses and to utilize hormetic principles to increase, for example, crop yields with plants. The NOX activity at the cell surface oscillates with a temperature-compensated 24-min ultradian (<24 h) periodicity. The indicated function of the NOX protein as a time-keeping mechanism adds to its potential importance as a molecular target for chemical hormesis. PMID- 10715616 TI - The relevance of the rat lung response to particle overload for human risk assessment: a workshop consensus report. AB - On 23-24 March 1998, the International Life Sciences Institute (ILSI) Risk Science Institute convened a workshop entitled "Relevance of the Rat Lung Response to Particle Overload for Human Risk Assessment." The workshop addressed the numerous study reports of lung tumors in rats resulting from chronic inhalation exposures to poorly soluble, nonfibrous particles of low acute toxicity and not directly genotoxic. These poorly soluble particles, indicated by the acronym PSPs (e.g., carbon black, coal dust, diesel soot, nonasbestiform talc, and titanium dioxide), elicit tumors in rats when deposition overwhelms the clearance mechanisms of the lung resulting in a condition referred to as "overload." These PSPs have been shown not to induce tumors in mice and hamsters, and the available data in humans are consistently negative. The objectives were twofold: (1) to provide guidance for risk assessment on the interpretation of neoplastic and nonneoplastic responses of the rat lung to PSPs; and (2) to identify important data gaps in our understanding of the lung responses of rats and other species to PSPs. Utilizing the five critical reviews of relevant literature that follow herein and the combined expertise and experience of the 30 workshop participants, a number of questions were addressed. The consensus views of the workshop participants are presented in this report. Because it is still not known with certainty whether high lung burdens of PSPs can lead to lung cancer in humans via mechanisms similar to those of the rat, in the absence of mechanistic data to the contrary it must be assumed that the rat model can identify potential carcinogenic hazards to humans. Since the apparent responsiveness of the rat model at overload is dependent on coexistent chronic active inflammation and cell proliferation, at lower lung doses where chronic active inflammation and cell proliferation are not present, no lung cancer hazard is anticipated. PMID- 10715617 TI - Dosimetry of particles in laboratory animals and humans in relationship to issues surrounding lung overload and human health risk assessment: a critical review. PMID- 10715618 TI - Pulmonary responses to inhaled poorly soluble particulate in the human. PMID- 10715619 TI - Rat lung tumors induced by exposure to selected poorly soluble nonfibrous particles. AB - Rodent bioassays have been used to assess the carcinogenicity of several inhaled, poorly soluble, nonfibrous particles that vary in toxicity and carcinogenic potency. There is substantial published information from chronic inhalation bioassays of diesel exhaust, carbon black, titanium dioxide, talc, and coal dust. This review summarizes data from studies with exposures for 2 yr or more using these 5 materials. The review has four objectives: (1) to summarize the current information available from these bioassays concerning exposure-dose-carcinogenic response in rats, (2) to summarize the pathologic and phenotypic features of the neoplastic response in rats, (3) to examine possible strain- and gender-related differences, and (4) to compare the neoplastic responses of rat to those of other species exposed to these materials. PMID- 10715620 TI - Nonneoplastic lung responses induced in experimental animals by exposure to poorly soluble nonfibrous particles. AB - The intent of this article is to review the comparative nonneoplastic pathological responses (inflammation, fibrosis, and epithelial proliferation) of rats, hamsters, mice, and monkeys to low-toxicity dusts (TiO(2), carbon black, talc, coal-mine dust, and diesel particulate). The paucity of information on some of these comparisons and the important information on other dusts (e.g., silica) that illuminate this question have necessitated inclusion of a number of papers that go beyond these nontoxic dusts. PMID- 10715621 TI - Mechanisms of action of poorly soluble particulates in overload-related lung pathology. AB - For reasons that are unclear, poorly soluble particulates are associated with the development of inflammation, fibrogenesis, and carcinogenesis in the rat. The pathogenesis of these changes may be triggered by distinct or species-specific cellular responses to inhaled particulates in a manner similar to known fibrogenic and carcinogenic fibers, such as asbestos. Data reviewed here suggest that generation of oxidants by poorly soluble particulates is a key factor in the initiation of inflammation and generation of chemokines and cytokines in the rat. These substances then cause hyperplasia of epithelial cells and fibroblasts. The diminished or lack of proliferative responses by poorly soluble particulates in mice and primates, in comparison to rats, may be reflected by intrinsic differences in their oxidant-generating capacities or repair after oxidant injury or DNA damage. PMID- 10715622 TI - Time course of response to ozone exposure in healthy adult females. AB - Ozone exposure causes acute decrements in pulmonary function, increases airway responsiveness, and changes the breathing pattern. We examined these responses in 19 ozone-responsive (DeltaFEV(1) > 5%) young females exposed to both air and 0.35 ppm ozone. The randomized 75-min exposures included two 30-min exercise periods at V(E) approximately 40 L/min. Responses were measured before, during, and after exposure and at 18 and 42 h postexposure. FVC, FEV(1), and FIV(0.5) decreased (p <.01) immediately postexposure by 13.2%, 19.9%, and 20.8%, respectively, and the airway responsiveness was significantly increased. Raw increased (p <.05), while TGV remained essentially unchanged. At 18 h postexposure, the airways were still hyperresponsive and FEV(1) and FIV(0.5) were still 5% below the preexposure levels. There were no residual effects in any of the variables at 42 h postexposure. During exercise in ozone the tidal volume was decreased (-14%) and respiratory frequency increased (+15%). The changes in airway responsiveness were not related to changes in spirometric measurements. We found no significant differences between postair and postozone mouth occlusion pressure (Pm(0.1)) and the hypercapnic response to CO(2) rebreathing. We conclude that ozone induced typical acute changes in airway responsiveness and that ventilatory (exercise), spirometric (inspiratory and expiratory), and plethysmographic pulmonary function may show some residual effects for up to 18 h after exposure. The ozone-induced alteration in breathing pattern during exercise does not appear to be related to a change in ventilatory drive. PMID- 10715623 TI - Inhaled particle-bound sulfate: effects on pulmonary inflammatory responses and alveolar macrophage function. AB - Acid sulfate-coated solid particles are a significant environmental hazard produced primarily by the combustion of fossil fuels. We have previously described a system for the nascent generation of carbonaceous particles surface coated with approximately 140 microg/m(3) acid sulfate [cpSO(4)(2-); 10 mg/m(3) carbon black (CB) and 10 ppm sulfur dioxide (SO(2)) at 85% relative humidity (RH)]. The effects of inhaled cpSO(4)(2-) on pulmonary host defenses are assessed in the present work. Mice were acutely exposed (4 h) to either 10 mg/m(3) CB, 10 ppm SO(2), or their combination at 10% or 85% RH in a nose-only inhalation chamber. No evidence of an inflammatory response was found following any of the exposures as assessed by total cell counts and differential cell counts from bronchoalveolar lavage fluid. However, alveolar macrophage Fc receptor-mediated phagocytosis decreased only following exposure to 140 microg cpSO(4)(2-), significant suppression occurred after 24 h, maximal suppression occurred at 3 days postexposure, and recovery to preexposure levels required 7-14 days. Intrapulmonary bactericidal activity (IBA) was also suppressed only after exposure to 140 microg cpSO(4)(2-); suppression was maximal at 1 day postexposure and recovered by day 7. To assess the effects of lower cpSO(4)(2-) concentrations, mice were repeatedly exposed to 1 mg/m(3) CB and 1 ppm SO(2) at 85% RH ( approximately 20 microg/m(3) cpSO(4)(2-) for 4 h/day) for up to 6 days. A significant decrement in IBA was observed following 5 and 6 days of exposure. These studies indicated that acute or repeated exposure to cpSO(4)(2-) could alter pulmonary host defense mechanisms. PMID- 10715624 TI - Antioxidant and inflammatory response after acute nitrogen dioxide and ozone exposures in C57Bl/6 mice. AB - Ozone (O(3)) and nitrogen dioxide (NO(2)) are highly reactive and toxic oxidant pollutants. The objective of this study is to compare chemokine, cytokine, and antioxidant changes elicited by acute exposures of O(3) and NO(2) in a genetically sensitive mouse. Eight-week-old C57Bl/6J mice were exposed to 1 or 2.5 ppm ozone or 15 or 30 ppm NO(2) for 4 or 24 h. Changes in mRNA abundance in lung were assayed by slot blot and ribonuclease protection assay (RPA). Messages encoding metallothionein (Mt), heme oxygenase I (HO-I), and inducible nitric oxide synthase (iNOS) demonstrated increased message abundance after 4 and 24 h of exposure to either O(3) or NO(2). Furthermore, increases in message abundance were of a similar magnitude for O(3) and NO(2). Messages encoding eotaxin, macrophage inflammatory protein (MIP)-1alpha, and MIP-2 were elevated after 4 and 24 h of exposure to 1 ppm ozone. Interleukin-6 was elevated after 4 h of exposure to ozone. After 4 h of 2.5 ppm ozone exposure, increased mRNAs of eotaxin, MIP 1alpha, MIP-2, Mt, HO-I, and iNOS were elevated to a higher magnitude than were detected after 1 ppm ozone. Monocyte chemoattractant protein (MCP-1) was elevated following 15 ppm NO(2) exposure. After 4 h of 30 ppm NO(2) exposure, messages encoding eotaxin, MIP-1alpha, MIP-2, and MCP-1 were elevated to levels similar to those detected after ozone exposure. Our results demonstrate a similar antioxidant and chemokine response during both O(3) and NO(2) exposure. Induction of these messages is associated with the duration and concentration of exposure. These studies suggest that these gases exert toxic action through a similar mechanism. PMID- 10715625 TI - Newborn mice differ from adult mice in chemokine and cytokine expression to ozone, but not to endotoxin. AB - Neonatal animals of some mammalian species are more tolerant to several pulmonary oxidative stress-inducing toxicants than adults. Our initial studies during hyperoxic injury demonstrated a rapid chemokine and cytokine response early in the development of injury in newborn mice, whereas adult mice demonstrated little alteration in cytokine abundance until lethality was imminent. Our hypothesis is that altered response between newborn and adult mice is associated with differential cell injury, rather than alterations in the regulation of the inflammatory response. To test this hypothesis we utilized two distinct models of inducing pulmonary toxicity: ozone (O(3)), which causes epithelial cell injury, and endotoxin, which causes pulmonary inflammation independent of direct epithelial cell injury. C57Bl/6J mice (36 h or 8 wk old) were exposed to O(3) at 1 or 2.5 ppm for 4, 20, or 24 h or to a 10-min inhalation of 10 ng endotoxin per mouse (estimated deposited dose) and were examined 2, 6, or 24 h postexposure. Adult mice displayed increased sensitivity to O(3), as demonstrated by increased abundance of mRNAs encoding eotaxin, macrophage inflammatory protein (MIP) 1alpha, MIP-2, interleukin (IL)-6, and metallothionein (Mt). In newborn mice, only Mt was increased after 4 h of exposure. In contrast, newborn and adult mice responded similarly at 2 h post endotoxin exposure, inducing messages encoding tumor necrosis factor (TNF)-alpha, eotaxin, MIP-1alpha, MIP-1beta, MIP-2, interferon inducible protein (IP)-10, and monocyte chemoattractant protein (MCP) 1. Furthermore, interleukin-6 (IL-6) was increased in adults but not newborns. Similar chemokine and cytokine responses of newborn and adult mice in response to an agent not causing epithelial injury (endotoxin) suggest that altered inflammatory control observed between newborn and adult mice following O(3) exposure is secondary to epithelial cell injury. PMID- 10715626 TI - Induction of adaptation to inhaled lipopolysaccharide in young and old rats and mice. AB - Lipopolysaccharide (LPS) is a component of the gram-negative bacterial cell wall that is known to activate inflammatory cells and enhance the production of inflammatory mediators in the lung. As it is a ubiquitous compound, inhalation exposure is highly likely in the human environment. Adaptation is a phenomenon by which a previous exposure results in improved survival or reduced injury as compared to a single exposure alone. We hypothesized that the basic proinflammatory effects of LPS in the lung could result in the development of adaptation in animals. Based on evidence of age- and species-related differences in lung injury, we used an acute lung injury model with inhaled LPS to compare the development of adaptation in young and old Fisher 344 rats and C57Bl/6J mice. Animals were exposed to low-dose (predicted lung deposition approximately 20 ng in rats and approximately 5 ng in mice) LPS aerosols for 10 min on 3 consecutive days; on day 4, a high dose (rats approximately 200 ng; mice approximately 25 ng) was delivered. Another group of animals received only the high LPS dose on day 4, whereas controls were unexposed. Twenty-four hours after the last exposure, cellular and inflammatory parameters in bronchoalveolar lavage (BAL) were determined. An adaptive response was found in both rats and mice. Adapted animals showed significantly fewer BAL neutrophils compared to nonadapted ones; there was also a significantly lower release of oxidants from phorbol methyl ester stimulated BAL cells from adapted compared to nonadapted animals, which, in turn, showed a greater response than controls. Furthermore, studies in old animals (21 mo of age) showed that adaptation also occurs in this age group. The adaptive response is clear in old mice; in rats, there is greater variability in the response, but an adaptive trend is apparent. Therefore, we have demonstrated that inhaled low-dose LPS can induce adaptation to subsequent higher doses, much as has been shown for other toxicants that induce oxidative lung injury. PMID- 10715627 TI - Ovalbumin aeroallergen exposure-response in Brown Norway rats. AB - A major route of exposure to allergens is through the respiratory tract. Comparatively few animal studies have used aerosolized high-molecular-weight allergens for sensitization, and in these studies, proper characterization of the aeroallergen exposure was usually missing. The purpose of this study was to profile the exposure-response relationship in Brown Norway rats (BNR) to well characterized ovalbumin (OVA) aerosols. Rats were exposed 30 min/wk x 6 wk to respirable OVA aerosols from <1 mg/m(3) to 64 mg/m(3) air. Ovalbumin-specific circulating immunoglobulin (Ig)E, IgG, and IgA were measured throughout the study period. Rats were sacrificed 1 day after the last exposure. Pulmonary tissue was processed for histopathological and histochemical analysis. Tracheas were isolated, perfused, and assessed for in vitro responsiveness to methacholine. Serum concentrations of OVA-specific antibodies increased with both exposure concentration and number of exposures. The number of BNR with measurable titers also increased with both dose and time. Pulmonary inflammatory changes were noted only in BNR exposed to higher OVA concentrations (15 and 64 mg/m(3) air). Increased tracheal reactivity to methacholine was not found in any of the sensitized BNR. In summary, sustained aeroallergen concentration-dependent changes in specific antibody responses and pulmonary inflammation have been demonstrated. PMID- 10715628 TI - Estimating in vitro glass fiber dissolution rate from composition. AB - A method is presented for calculating the dissolution rate constant of a borosilicate glass fiber in the lung, as measured in vitro, from the oxide composition in weight percent. It is based upon expressing the logarithm of the dissolution rate as a linear function of the composition. It was found that the calculated dissolution rate constant agreed with the measured value within the variation of the measured data in a set of compositions in which the dissolution rate constant ranged over a factor of 100. The method was shown to provide a reasonable estimate of dissolution over a considerably wider range of composition than what was used to determine the parameters, such as a set of data in which the dissolution rate constant varied over a factor of 100,000. The dissolution rate constant may be used to estimate whether disease would ensue following animal inhalation or intraperitoneal studies. PMID- 10715629 TI - Air particulate pollution and hospital admissions for chronic obstructive pulmonary disease in Reno, Nevada. AB - This study assessed the association between ambient PM(10) pollution and daily hospital admissions for chronic obstructive pulmonary disease (COPD) in Reno Sparks, Nevada, for the period 1990-1994. All three hospitals in the region were included. There was a total of 3115 admissions for COPD during this period. Daily ambient PM(10) values were available from one of seven air monitoring stations in this region. Weather variables including temperature and wind speed were also collected from this station. The daily average concentration of PM(10) was 36.55 microg/m(3). The generalized additive model (GAM) was used in the whole analysis. After adjusting for the effects of weather variables, day of week, seasons, and time trend, the results show that PM(10) is a statistically significant predictor for daily hospital admissions for COPD. The relative risk (RR) of hospital admissions for COPD for an interquartile increase (26.6 microg/m(3)) of the 24-h average level of PM(10) is 1.049 (95% CI 1.011-1.087). PMID- 10715630 TI - Feasibility study of prolonged ozone inhalation exposure via face mask. AB - Because of the interest attendant to establishing an 8-h ozone (O(3)) federal air quality standard, acute pulmonary function responses to prolonged (6.6 to 8 h) O(3) exposure between 0.08 and 0. 24 ppm have been examined in chamber studies. Given time constraints for O(3) concentration changes in room-sized chambers and the need to simulate rapid fluctuations in O(3) levels, such as occurs when one goes from indoors to outdoors during an air pollution episode, mouthpiece or face mask exposure systems offer potential advantages over chamber exposure systems. In a recent study in this laboratory, subjects indicated that over 2 h of continuous exposure via an obligatory mouthpiece inhalation system (with noseclip) was more than they could tolerate with this type of exposure system. The purpose of this study was to compare O(3)-induced responses observed following 2-h exposures via an obligatory mouthpiece inhalation system and a newly devised face-mask exposure system, and to determine whether the latter could be tolerated during 4-h exposures. Six young adults completed 5 experimental protocols (three 2-h and two 4-h) while exposed to O(3) concentrations ranging from 0.00 ppm (filtered air) to 0.24 ppm. Exercise and resting minute ventilation (V(E)) were measured continuously. Pulmonary function, subjective symptoms (SS) of breathing discomfort, and exercise ventilatory pattern responses were obtained. Exposure to 0.24 ppm O(3) with intermittent exercise (IE) for 2 h via a newly devised inhalation system, consisting of a nylon plastic non-rebreathing respiratory valve and a silicone rubber face mask with Teflon overlay on the inner surface, yielded pulmonary function, SS, and exercise ventilatory pattern responses nearly identical to those obtained in the same O(3) exposure effected via a Teflon-coated Hans-Rudolph respiratory valve obligatory mouthpiece inhalation system. It was concluded that the newly devised face-mask inhalation system was well tolerated by all subjects during the 4-h exposure protocols, with each subject indicating that longer 6.6-h exposures with this system would be tolerable. PMID- 10715631 TI - Inhalation toxicity studies of aqueous dispersion resin. AB - Aqueous dispersion resin (ADR) is a water-based acrylate copolymer designed to allow a range of ethanol concentrations for hair-spray formulations. A series of inhalation (whole-body) aerosol toxicity studies, including acute, 9-day, and 90 day exposures, was conducted to determine any toxicological effects for ADR. An acute study of ADR was conducted for 4 h with a 14-day observation period. The maximum ADR aerosol concentration was 1.07 (+/- 0.04) mg/L with a mass medium aerodynamic diameter (MMAD) value of 1.46 microm and a geometric mean (sigma(g)) of 1.42. No deaths or exposure-related lesions were observed. Thus, the LC50 value was greater than 1.07 mg/L. For the 9-day study, each group contained 10 animals/sex and was exposed for 2 h/day. The mean exposure concentrations (+/- standard deviation) were 244 (+/- 39.6), 543 (+/- 71.9), and 843 (+/- 186.2) mg/m(3) for the 250-, 500-, and 1000-mg/m(3) groups, respectively. The mean MMADs were 2.83, 2.90, and 4.09 microm for the 250-, 500-, and 1000-mg/m(3) groups, respectively, with sigma(g) values ranging from 3.15 to 6.22. Unkempt fur and alopecia in the males and females from the 1000-mg/m(3) group at sacrifice were the only exposure-related clinical signs observed. Increased lung weights in the 500- and 1000-mg/m(3) exposure groups were found. Histopathological effects, such as alveolar histiocytosis and interstitial pneumonitis, were also noted in these two exposure groups. For the 90-day study, 10 animals per sex were included in each group with a 6-wk recovery group of 5 female rats/group. Exposures were conducted for 2 h/day. The mean exposure concentrations were 30.4 (+/- 2.3), 102 (+/- 11.6), and 308 (+/- 19. 3) mg/m(3) with mean MMADs of 3.09, 2.64, and 2.67 microm and sigma(g) values ranging from 3.54 to 3.90 for exposure concentrations of 30, 100, and 300 mg/m(3), respectively. The only findings at the 90-day sacrifice were increased lung weights for the males and females in the 300 mg/m(3) group and males in the 100-mg/m(3) group. For the 6-wk recovery sacrifice of the females, the lung weights for the 300- and 100-mg/m(3) groups were increased. Histopathological effects at the 90-day sacrifice included alveolar histiocytosis and lymphadenitis in the mediastinal lymph nodes in the 100- and 300-mg/m(3) exposure groups for males and females, while alveolar histiocytosis, intraalveolar cellular debris, lymphadenitis in the mediastinal lymph nodes, and/or interstitial pneumonitis were noted in these 2 exposure groups of the females after the 6-wk recovery period. Animals exposed for 90 days to 100 and 300 mg/m(3) for 2 h had increased lung weights. However, no effects were observed in the 30-mg/m(3) exposure group. Thus, under the conditions of the present 90 day study, a no-observable-effect level (NOEL) was found to be 30 mg/m(3). PMID- 10715632 TI - Intratracheal instillation of zinc-cadmium sulfide (ZnCdS) in Fischer 344 rats. AB - The purpose of this study was to assess the bioavailability and pulmonary toxicity of ZnCdS in rats. Groups of 30 male Fischer 344 rats each were anesthetized and dosed via intratracheal instillation with 5 mg of either ZnCdS, quartz (positive control), or titanium dioxide (TiO(2), negative control) suspended in 0.5 ml saline. A vehicle control group received 0.5 ml saline. Ten animals from each test group were sacrificed at 1 day, 1 wk, and 14 wk after dosing for bronchoalveolar lavage fluid (BALF) analysis and histopathology. The BALF was analyzed for alkaline phosphatase, acid phosphatase, lactate dehydrogenase (LDH), beta-glucuronidase (beta-glu), total protein, and cell counts. Two separate groups of 24 rats each were dosed as already described with either ZnCdS or saline. Eight rats from each group were sacrificed at 1 day, 1 wk, and 14 wk after dosing for determination of cadmium (Cd) and zinc (Zn) concentrations in the lung, liver, kidney, and blood. Results indicate that at 1 day after dosing, all enzyme activities (except acid phosphatase) and cell counts in BALF from the quartz and ZnCdS groups were significantly higher than in the TiO(2) and saline groups. At 7 days after dosing, high enzyme activity persisted in the quartz group, while the ZnCdS group showed only LDH and total protein levels significantly higher than the saline group. At 14 wk after dosing, LDH, total protein, beta-glu, and cell counts in the quartz group were significantly higher than all other groups. Histologic examination revealed interstitial inflammation and accumulation of foreign material in the lungs and mediastinal lymph nodes of quartz-, TiO(2)-, and ZnCdS-treated rats. Metal analyses in tissues showed profuse Cd and Zn concentrations in the lung 1 day after dosing, followed by a successive decline at 7 days and 14 wk after dosing. A very small, but statistically significant, amount of Cd and Zn was found in the kidneys at 14 wk after dosing. In conclusion, ZnCdS appears to cause temporary lung inflammation, is cleared slowly, and is poorly bioavailable. PMID- 10715633 TI - Posttreatment with ETYA protects against phosgene-induced lung injury by amplifying the glutathione to lipid peroxidation ratio. AB - Exposure to phosgene has been shown to cause severe and life-threatening pulmonary edema. There is evidence that successful treatment of phosgene-induced acute lung injury may be related to increased antioxidant activity. Acetylenic acids such as 5,8,11, 14-eicosatetraynoic acid (ETYA) have been shown to be effective in preventing pulmonary edema formation (PEF). In phosgene-exposed guinea pigs, we examined the effects of ETYA on PEF. Lipid peroxidation (thiobarbituric acid-reactive substance, TBARS) and total glutathione (GSH) were measured in lung tissue from isolated, buffer-perfused guinea pig lungs at 180 min after start of exposure. Guinea pigs were challenged with 175 mg/m(3) (44 ppm) phosgene for 10 min (1750 mg( small middle dot)min/m(3)). Five minutes after removal from the exposure chamber, guinea pigs were treated, ip, with 200 microl of 100 microM ETYA in ethanol (ETOH). Two hundred microliters of 50 microM ETYA in ETOH was added to the 200 ml perfusate every 40 min beginning at 60 min after start of exposure (t = 0). There were four groups in this study: air-exposed, phosgene-exposed, phosgene + ETYA-posttreated, and air + ETYA-posttreated. Posttreatment with ETYA prevented GSH depletion, 2. 7 +/- 0.5 micromol/mg protein versus 1 +/- 0.2 micromol/mg protein, for the untreated phosgene-exposed lungs (p < or =.05). ETYA posttreatment also significantly decreased PEF (p 0.5 root surface caries per year after the operation but none before (p<.01). Salivary chloride was > 12 meq/l in 3 of the 18 JI bypass patients but in none of 5 controls (p<.05). The salivary pH and bicarbonate were 6.38+/-0.48 vs 6.92+/-0.21 and 2.81+/-2.1 meq/l vs 5.8+/-1.2 meq/l in the JI bypass group and the control group respectively (p<.05). The stimulated saliva was 2.3+/-1.2 vs 4.5+/-1.4 cc/min in the JI bypass group and control group, respectively (p<.02). CONCLUSIONS: Root surface caries are more frequent after JI bypass. This may be due to decreased saliva flow and a reduced salivary buffering capacity. PMID- 10715643 TI - Vertical banded gastroplasty: long-term results comparing three different techniques. AB - BACKGROUND: Vertical banded gastroplasty (VBG) has been our procedure of choice for the treatment of morbid obesity from 1981-1995, at which time it was replaced by laparoscopic gastric banding. Three different techniques have been used for banding: silastic band, marlex mesh, adjustable sphincter. The purpose of this paper is to present the long-term results. METHODS: The charts from all patients operated on during the aforementioned period were reviewed and the data analysed retrospectively. RESULTS: This series comprises 197 patients, 172 females and 25 males, with a mean initial excess weight of 94.8% (6-300%) and a mean initial Body Mass Index (BMI) of 42.9 kg/m2 (23-88 kg/m2). 73 patients had a silastic band, 40 Marlex mesh, and 84 an adjustable sphincter. Overall excess weight loss was 66% after 12-24 months, and remained between 50 and 60% up to 9 years postoperatively. There was no difference between the 3 groups. 82 patients (41%) developed a total of 117 complications during follow-up. Among them were stenosis 20%, staple-line disruption 11%, incisional hernia 13%, severe esophagitis 7% and band migration 1.5%. Stenosis developed more often with a silastic band or an adjustable sphincter, and severe esophagitis was more prevalent after the adjustable sphincter. 58 patients required one or more procedures for correction, including dilatation in 21, band removal in 17, band replacement in 15, restapling in 19 and incisional hernia repair in 11 patients. Overall, 29.4% of patients had to be reoperated. There were more reoperations in the silastic and adjustable sphincter groups compared with the Marlex mesh group. CONCLUSIONS: VBG is associated with a rapid weight loss that is relatively well-maintained over time, although there is a tendency to slight weight regain after 2 years. The price for these results is high if complications and further necessary procedures are considered, especially after banding with a silastic band or an adjustable sphincter. Marlex mesh represents the banding material of choice if VBG is chosen. PMID- 10715642 TI - Dietary changes after vertical banded gastroplasty. AB - BACKGROUND: Gastric restriction surgery relies on obstruction to oral intake by formation of a gastric pouch. Therefore, the therapeutic effect is closely related to intolerance for different types of food, and an ingestion of an unbalanced diet. We investigated dietary changes after VBG and their associations with therapeutic success. METHODS: 70 patients (4 men, 66 women, median age 32) with a median preoperative BMI of 44.6 were examined > or =3 years after VBG. Weight reduction, nutritional changes (type of diet, number of daily meals, amount of food that could be ingested, intolerance for different types of food, frequency of vomiting), satisfaction with results, and willingness to undergo the operation once again were investigated. RESULTS: The average reduction of the BMI was 13, with sufficient weight loss in 80%. 36% were eating a solid, 43% a soft, and 21% a liquid diet. Weight reduction did not depend on the type of diet eaten but on the ingestion of sweets. 93% indicated they could take only small amounts of food. The average number of daily meals was 3.76% reported an intolerance for some type of food (most often meat, fruit, or vegetable). Vomiting was the most common problem and occurred in 71%. 71% indicated a high level of satisfaction with the results of the operation, and 96% said they would undergo the operation again. CONCLUSIONS: The investigation demonstrated successful weight reduction despite dietary changes in 80% of patients after VBG. Weight reduction was not influenced by type of, diet but depended on consumption of sweets. PMID- 10715644 TI - Variation in VBG. PMID- 10715645 TI - Some hormonal changes before and after vertical banded gastroplasty for severe obesity. AB - BACKGROUND: Hormonal disturbances play a role in the development of obesity, but may be a consequence of obesity itself. In this study we assessed the influence of the surgically-induced weight loss on some important hormonal abnormalities in the morbidly obese patients. MATERIAL AND METHODS: Fasting serum prolactin, insulin, cortisol and thyroid hormones: free thyroxin (FT4), free triiodothyronine (FT3) and thyrotropin (TTH), have been studied by radioimmune methods before vertical banded gastroplasty (VBG) and after operation in the early (10-14 days) and late period when excess weight loss (EWL) 51.7-57.1% had been achieved. RESULTS: On the 10-14 day after VBG, prolactin increased significantly in women (p<0.05), but decreased after weight loss (p<0.01). Fasting insulin was lowered significantly (p<0.05) soon after VBG in the hyperinsulinemic (51.7% of the total group) and diabetic (n-9) patients. After weight loss, insulin decreased significantly (p<0.0001) vs. preoperative. Concentration of cortisol was unchanged both in the early and in the late postoperative period. On the days 10-14, significant elevation of TTH and decrease of FT3 (p<0.05) have been noted. After essential weight loss TTH dropped significantly vs. preoperative (p<0.05) with no changes in FT3 and FT4 concentration. CONCLUSIONS: VBG and consequent weight loss favorably influence the hormonal abnormalities in the morbidly obese. Further studies are needed to make clear a relationship between this and other parameters of metabolic syndrome. The hormonal abnormalities may influence the indications for surgery in less than morbidly obese patients with metabolic syndrome. PMID- 10715646 TI - Report on bariatric surgery in the Ukraine. AB - BACKGROUND: Morbid obesity (MO) is a problem internationally, including in the Ukraine. We present the surgical treatment of MO in the Ukraine over the last 15 years, during which intestinal bypasses and various gastric reduction procedures were performed. METHODS: 198 patients with MO underwent: jejunoileal (JI) bypass 64, non-adjustable gastric banding (NGB) 34, Roux-en-Y gastric bypass (RYGBP) 1, horizontal gastroplasty 1, vertical banded gastroplasty (VBG) 2, and abdominal lipectomy 96. The 96 men and 102 women weighed 160-290 kg (mean 210+/-SD18 kg). Mean body mass index was >60 kg/m2. These patients had a high incidence of hypertension, diabetes, sleep apnea, menstrual disorders, impotency in men and infertility in women. RESULTS: At 1 year, after JI bypass 61 patients lost a mean of 62+/-17 kg and after NGB 11 kg. After JI bypass, 1 patient died in the early postoperative period from acute respiratory insufficiency and 2 died in the first year from acute liver insufficiency. The JI bypass was reversed in 2 patients due to uncontrollable malabsorption syndrome; 1 year after reversal, the weight of these patients exceeded their preoperative weight. In the early postoperative period, 1 patient died after NGB and 1 after RYGBP, from acute respiratory insufficiency. Postoperative weight loss was associated with decrease in the co morbidities of MO, but after JIB, there was a high incidence of bypass enteritis, excessive malabsorption, formation of renal stones and gallstones. After NGB, no complications have been identified. Isolated lipectomy was performed in 44 patients, lipectomy combined with a bariatric operation in 31, and lipectomy after loss of the excess body weight in 21. CONCLUSIONS: Bariatric surgery was very effective in weight loss, accompanied by reduction or disappearance of the co-morbidities of MO, with considerable improvement in quality of life. PMID- 10715647 TI - Intragastric balloons for preoperative weight reduction. AB - BACKGROUND: If medical treatment of obesity fails and if surgical gastroplasty is not indicated, insertion of an intragastric balloon may represent an intermediate modality. METHODS: Two patients are reported in whom a balloon was placed for weight reduction before elective surgery: 1) A 53-year-old woman with a BMI of 41.3 kg/m2 lost 18 kg in 6 months and then underwent surgical repair of a huge incisional hernia; 2) A 58-year-old woman with a BMI of 35.8 kg/m2 had total hip arthroplasty after losing 15.5 kg in 5 months. RESULTS: The uneventful postoperative recovery in both patients was thought to be positively influenced by their preoperative weight loss. CONCLUSION: In morbidly obese patients, intragastric balloon placement may contribute to preoperative weight reduction before elective surgery. PMID- 10715648 TI - Vertical banded gastroplasty staple-line dehiscence after blunt trauma to the abdomen. AB - An extensive Medline search revealed no documentation in the literature regarding staple-line disruption of a vertical banded gastroplasty after blunt abdominal trauma. A 41-year-old woman with a past surgical history significant for a vertical banded gastroplasty 13 months ago was admitted to our hospital one week after an episode of blunt trauma to the abdomen. The patient presented with complaints of nausea, vomiting, and anorexia after the incident. The suspected diagnoses were stenosis of the stoma due to hematoma after the blunt trauma as well as staple-line dehiscence. A partial disruption of the vertical staple-line was identified by esophagogastroduodenoscopy and barium swallow. PMID- 10715649 TI - Hypopharyngeal perforation during laparoscopic Roux-en-Y gastric bypass. AB - Laparoscopic Roux-en-Y gastric bypass was recently introduced as an alternative surgical treatment for morbid obesity. The technique involves placement of a 21 mm anvil transorally down to the gastric pouch for creation of the gastroenterostomy anastomosis using an EEA stapler placed transabdominally. Esophageal injury is a theoretical concern with transoral manipulation of the anvil. The authors present a case of hypopharyngeal perforation after an attempted transoral insertion of an EEA anvil. The perforation was treated with neck exploration and drainage. We discuss the mechanism of injury and alternative method for placement of the gastric anvil. PMID- 10715650 TI - Modified VBG vs gastric bypass. PMID- 10715652 TI - Public relations as a means of public education on the role of bariatric surgery. PMID- 10715651 TI - 650 year anniversary of Charles University, Prague. PMID- 10715653 TI - An International Surgical Journal for Research and Treatment of Massive Obesity. PMID- 10715654 TI - A Past Presidential Address to the American Society for Bariatric Surgery. PMID- 10715655 TI - A Tribute to Edward Eaton Mason: the First Annual Edward E. Mason Founders Lecture. PMID- 10715656 TI - The Etiology of Obesity. AB - Obesity likely results from a genetically predetermined body mass set-point that exerts control of body weight through alterations in basal metabolic rate. This set-point may be further influenced by learned eating behaviour, perception of body image, socioeconomic status and the availability of food. This article examines the complex etiology and pathogenesis of obesity from biological, sociocultural and psychological perspectives. Endocrine abnormalities and defective brown fat thermogenesis are also examined and found not to be primary etiologic factors in the development of obesity. PMID- 10715657 TI - Predeposit Autologous Blood Donation: a survey of patient attitudes. AB - Attitudes to autologous blood donation have been surveyed in a group of 38 postoperative bariatric patients. Only two patients (5%) declined to participate. Twenty-eight of 38 ( 70%) successfully predeposited autologous blood. Twenty-five of 28 donors (90%) had done so at the suggestion of their surgeon. Concern about contracting AIDS was the motivating factor in the majority of patients (21 patients = 55%). Lack of infectious complications in general was cited by an additional four (11%). All respondents would donate autologous blood in the future, and would recommend the procedure to others who were about to undergo elective surgery. There was an increase from 29% to 50% who stated that, following their autologous donation experience, they would consider being homologous volunteer blood donors in the future. PMID- 10715658 TI - Effect of Wound Closure Technique on Wound Infection in the Morbidly Obese: results of a randomized trial. AB - The effect of suture obliteration of the subcutaneous dead space in morbidly obese abdominal wounds was studied in a randomized trial, comparing a pre-fascial retention suture technique (utilized for approximated of the thick panniculus) to controls where the skin was simply closed with staples. The wound infection rates were similar (11.8% for the sutured group versus 12.3% for controls, p 0.4), as were the total wound complication rates (26.5% for sutured group versus 21.9% for controls, p 0.4). Ultrasound study of the wounds closed without suturing the panniculus demonstrated no dead spaces. We conclude that no advantage is to be gained by suturing the subcataneous fat, however thick. The finding is of general application in wound closures involving thick layers of fat. PMID- 10715659 TI - Exercise-induced Wall Motion Abnormalities and Resting Left Ventricular Dysfunction in the Morbidly Obese as Assessed by Radionuclide Ventriculography. AB - Rest and exercise first pass radionuclide ventriculograms were obtained in 62 morbidly obese subjects (56 women, six men, mean age 38 years, mean weight 269.2 +/- 46.0 lb, mean height 65.2 +/- 3.1 in., mean Body Mass Index 44.5 +/- 6.2 kg/m(2), mean excess body weight 134.1 +/- 41.1 lb) scheduled for vertical banded gastroplasty. Fifty-six percent demonstrated exercise-induced wall motion abnormalities mimicking coronary disease, compared to 12% of controls (p = 0.03). No subject with exercise-induced abnormalities had coronary disease at cardiac catheterization although only those with an anginal chest pain history underwent angiography. Twenty-six percent demonstrated resting left ventricular systolic dysfunction as manifested by a reduced resting left ventricular ejection fraction ( <0.50). Thirty-one percent of these patients demonstrated exercise-induced abnormalities, versus 65% of morbidly obese subjects with normal resting ejection fractions (p = 0.04). Obesity-induced left ventricular hypertrophy with associated reduced coronary vasodilator reserve could explain the abnormalities. Six month post-gastroplasty follow-up radionuclide ventriculograms show group normalization of the resting left ventricular ejection fraction in those with preoperative dysfunction, possibly due to left ventricular unloading with some regression of hypertrophy. PMID- 10715660 TI - Age, Obesity Surgery, and Weight Loss. AB - Some centers consider an age over 50 to be a contraindication for obesity surgery. This study was conducted to examine the relationship between age and one year postoperative weight of patients receiving gastric restrictive surgery (n = 616) for morbid obesity. Patients were divided into four age groups (18-29, 30 39, 40-49, 50-65 years) matched for preoperative obesity. At one year there were no statistically significant differences in weight loss or postoperative obesity. There were four (0.6%) surgically-related deaths. The mortality of patients aged 50 or older (1.1%) was not significantly higher than that of younger patients (0.6%). It was concluded that older age per se need not be a contraindication for surgery. PMID- 10715661 TI - Gallbladder Disease in the Morbidly Obese Patient. AB - Obesity alone and rapid weight loss induced by bariatric surgical procedures are both recognized risk factors for the development of cholelithiasis. The advisability of performing routine concomitant cholecystectomy with bariatric surgery has been controversial. This prospective clinical study was performed to determine if preoperative ultrasonography, preoperative cholescintigraphy and intraoperative surgical findings could help determine which patients would benefit from concomitant cholecystectomy. Eighteen morbidly obese patients undergoing bariatric surgery were studied preoperatively with gallbladder ultrasonography and cholescintigraphy. All patients had a description of intraoperative findings. Gallbladders were removed routinely and examined pathologically. Sixty percent of patients in this study had abnormal gallbladder pathology. Preoperative ultrasonography and/or cholescintigraphy was inaccurate at predicting reliably those patients who would benefit from cholecystectomy concomitant with gastric-restrictive surgery. lntraoperative findings other than palpable gallstones were also unreliable. PMID- 10715662 TI - An Analysis of the Predictability of Hypoxemia Following Vertical Banded Gastroplasty. AB - Postoperative (post-op) hypoxemia is unpredictable, often undetected by physical examination, sometimes fatal. We studied 45 morbidly obese patients with an average age of 37, including 16 smokers, having vertical banded gastroplasty (VBG) for useful preoperative (preop) predictor(s) of post-op hypoxemia during the first five days following VBG. Patient blood gases (arterial blood oxygen, P&infa;O&inf2; in mmHg), pre-op and five post-op days (POD), after 30 min in room air were: pre-op, 85 +/- 9; POD1, 63 +/- 9*; POD2, 61 +/- 9*; POD3, 63 +/- 10*; POD4, 63 +/- 9* POD5, 64 +/- 1 * (* p < 0.05, Student's t-test compared with pre op). Linear regression showed no practical, predictive value for P.02 for age, Body Mass Index (BMI), pulmonary function tests (PFTs), smokers or preop P&infa;O&inf2;. Post-op atelectasis occurred in 84% of patients, mostly the posterior basilar regions on chest X-ray. No patient developed clinically diagnostic pneumonia. VBG patients experienced profound hypoxemia post-op, the lowest on POD2. There is no reliable method to predict which patient may develop severe hypoxemia. It is, therefore, extremely helpful to uniformly monitor P&infa;O&inf2; post-op in morbidly obese patients. PMID- 10715663 TI - Pathologic Changes in the Stomach at the Site of Silicone Gastric Banding. AB - Silicone gastric banding procedures for severe obesity have been done on 303 patients; 214 as primary operations and 89 as revisions of different types of gastric restriction operations. Twenty-five bands were removed, and in 16 of these cases full thickness gastric wall biopsies were done at the banding site. The pathologic changes at the banding site in these 16 patients consisted of variable local tissue responses consistent with constriction due to the banding process. The most significant finding, seen in only one patient, was full thickness fibrous replacement of the muscular wall. Ten patients had only serosal fibrosis at the banding site and four had both serosal fibrosis and patchy fibrous replacement of the muscularis propria. One biopsy specimen showed no pathologic changes. Notably absent in all cases were evidence of ischemia and evidence of penetration of the gastric wall by the band. PMID- 10715664 TI - Folate Status Following Gastric Bypass Surgery (The Great Folate Mystery). AB - Several previous investigators have reported an incidence of folic acid deficiency following gastric bypass surgery of up to 38%. Failure to encounter any folic acid deficiencies in our postoperative patients led us to discontinue follow-up folate studies for several years. However, due to repeated references to this deficiency in the literature, we re-instituted folate studies as part of the routine follow-up of our patients. Preoperative serum folate levels were obtained in 1,067 patients and preexisting deficiencies found in 63, an incidence of 6%. Of the 588 folate levels determined 1 to 10 years following gastric bypass, only six were less than 3.0 ng/di, an incidence of 1%. All patients were instructed preoperatively and postoperatively to take multivitamin/mineral supplements after gastric restrictive surgery, and were continually educated on their importance. In a bariatric surgery practice in which patients are instructed, reminded, encouraged and even badgered into taking postoperative vitamin/mineral supplements, folate deficiency should be a rarity. In such circumstances, folate deficiency may well act as a sensitive marker of non compliance. PMID- 10715665 TI - Lengthening the Roux-Y Limb Increases Weight Loss after Gastric Bypass: a preliminary report. AB - We compared two variants of gastric bypass which have been used at our hospital since 1984. Initially all patients had a standard 45 cm Roux-Y anastomosed to a 30 cc gastric pouch. Subsequently we increased the length of the Roux-Y from 45 to 90 cm. In all patients the jejunum was divided 15-20 cm from the ligament of Treitz. There were six males, and 49 females with a mean age of 35 years. All were at least twice their ideal weights (range 91.5 to 179, X = 127.6). Percentage follow-up ranged from 100% at three months to 13% at 66 months for both the standard and lengthened Roux-Y groups. There were no major technical or metabolic complications. Doubling the length of the standard Roux-Y limb increased the percentage excess weight lost by approximately 6% without diarrhea or other apparent metabolic, sequelae. PMID- 10715666 TI - Microorganisms in the Stomach During Vertical Banded Gastroplasty. AB - Vertical banded gastroplasty involves the cutting of a stapled window against an Ewald tube Passed perorally, and creation of a vertical staple-line partition. The channel at the window is banded by a polypropylene mesh collar. A study of the gastric rings removed at this operation found that the bacterial colonization rate was 46.8%, so that the potential for infection of the collar exists. The actual significance of this finding is unknown. PMID- 10715667 TI - Long-term Result of Treatment of Prader-Willi Syndrome by Scopinaro's Bilio pancreatic Diversion. Study of Three Cases and the Effect of Dextrofenfluramine on the Postoperative Evolution. AB - The Prader-Willi Syndrome shortens the life of patients due to the morbid obesity which it entails. The compulsive hyperphagia associated with it makes a dietetic treatment or a gastroplasty difficult. This study presents the case histories of three patients suffering from the Prader-Willi syndrome who were operated on by means of a Scopinaro's bilio-pancreatic diversion. Following a marked reduction the first year, the weight loss stabilized and then tended to diminish. The observation of three cases which continued for two and a half to six years did not reveal any considerable metabolic problems. The deficiency of iron, vitamins D and B12 as well as folic acid had to be made up by supplementation. These results are comparable with the most favorable ones in the literature. Even if the effect on the weight loss is not spectacular, the operation manages to hold off the development of the obesity, inexorable for those with the Prader-Willi syndrome, and prevents lethal complications, without having notable side effects. Lifting coercive dietary measures improves the quality of life. PMID- 10715668 TI - The Use of the Biliopancreatic Diversion as a Treatment for Failed Gastric Partitioning in the Morbidly Obese. AB - The dilemma with which every bariatric surgeon is confronted is: What to do with the inevitable failures? In vertical gastric partitioning, revising the gastroplasty results in a high second failure rate. In an effort to improve the Success rate in failed gastroplasty patients who request revisionary surgery, the biliopancreatic bypass (classic Scopinaro procedure) was carried out on 57 patients. They have been followed for up to 10 years. The long-term weight loss has averaged 69.4 lb, which is 87% of the pregastroplasty excess weight. The price paid by these patients, in terms of complications, has been significant. Twenty-two Percent have developed hypoalbuminemia with its accompanying peripheral edema; 24% have required i.v. hyperalimintation because Of malnutrition. Sixteen percent of the patients developed a late post-op bowel obstruction, one resulting in death. Osteomalacia, spontaneous fractures have occurred. The biliopancreatic diversion procedure (BPD) is an effective weight loss operation in the failed gastroplasty patients, but a significant price must be paid in terms of careful follow-up, nutritional deficiencies, and rehospitalizations. PMID- 10715669 TI - Use of a Calibration Balloon Tube to Size the Pouch and Channel in Gastroplasty Procedures. AB - Gastroplasty is currently one of the most common surgical procedures performed on the morbidly obese for weight loss. An adequate result can be assured only if the pouch that is created is less than 30 ml and the channel that connects that pouch to the distal stomach is approximately 1 cm in diameter. The current method to size the pouch is to occlude the esophagus and the outlet of the pouch and to measure with a manometer through a naso-gastric tube. We contend this method is both time consuming and adds to the potential of complications. Through the use of a calibration balloon tube the size of the pouch can be quickly and safely estimated. It can also be used to size the channel between the pouch and the distal stomach and check for leaks. The technique of how this tube has been used over the past 6 years is described. By the use of a calibration balloon tube, three problem areas in gastric stapling surgery for morbid obesity are avoided, namely: inappropriate pouch size, inappropriate channel size and postoperative leaks. PMID- 10715670 TI - Efficient Decompression and Immediate Enteral Hyperalimentation via Gastrostomy as an Adjunct to Gastroplasty. AB - A triple-lumen Moss(c) gastrostomy tube was advanced into the proximal duodenum as an adjunct to the postoperative management of patients who underwent vertical banded gastroplasty (VBG) for the treatment of morbid obesity. The tube efficiently aspirated the proximal duodenum and stomach to prevent postoperative ileus and allow maximum immediate postoperative absorption of an elemental diet fed simultaneously into the distal duodenum. Decompression and feeding were started as soon as the patient arrived in the recovery room. Both were continued for at least the first 48 h after surgery. Patients seemed to improve better clinically on this postoperative regimen than with only the traditional i.v. infusion of fluids, carbohydrates, and electrolytes. The length of stay also was shorter than that allowed in the Diagnosis Related Group (DRG) for the; surgical management of morbid obesity: with the average of 3.0 days (range 2-6 days) versus the general mean length of stay of 7.4 days. Immediate enteral decompression and hyperalimentation through a gastroduodenostomy tube is a useful adjunct to the post-VBG treatment of morbidly obese patients. PMID- 10715671 TI - Adjustable Gastric Banding. AB - Gastric banding is an exceptionally safe operation for the surgical control of morbid obesity. Most failures are due to a stoma that is too large or too small. To obviate the 20% failure rate (removal of band or readjustment with major surgery), a band which is adjustable from the subcutaneous tissues is described. This revisional surgery can be done on an out-patient basis under local anesthesia. Major intraperitoneal surgery is avoided with the great expense, risk, pain, and disability. PMID- 10715672 TI - Genomic organization, transcription, splicing and gene regulation in Leishmania. AB - The parasitic protozoan Leishmania is the aetiological agent of a spectrum of clinical diseases, ranging from disfiguring skin lesions to life-threatening visceral infection, and is a serious health problem in tropical and subtropical areas world-wide. Leishmania parasites undergo a dramatic transformation as they move between the different environments of an extracellular insect stage and an intracellular form in the vertebrate host. In an attempt to develop new strategies for the treatment of leishmaniasis, the techniques of molecular genetics have been utilised to elucidate the mechanisms which direct and control this cyclical differentiation. This review discusses current knowledge concerning the organization and regulation of the Leishmania nuclear genome and includes a discussion of chromosomal organization, genomic arrangement, transcription, transcript processing by trans-splicing and polyadenylation, and post transcriptional regulation. The salient features as well as the supporting evidence for each topic are briefly reviewed. PMID- 10715673 TI - Efficacies of 5- and 14-day primaquine regimens in the prevention of relapses in Plasmodium vivax infections. AB - Vivax malaria accounts for 80% of malaria cases in Mumbai (Bombay) and has high morbidity. In India, the standard treatment to prevent relapses of vivax malaria is a 5-day regimen of primaquine. However, between 1977 and 1997, the efficacy of this treatment declined from approximately 99% to 87%. The efficacy of the 5-day regimen was therefore compared with that of the 14-day regimen currently recommended by the World Health Organization, in Mumbai. The relapse rates observed, over a 6-month period of follow-up, were 0% with the 14-day regimen, 26.7% with the 5-day, and 11.7% when no primaquine treatment was given. The expenditure incurred on the door-to-door dispensing of the 5-day regimen appears to be without benefit. There is an urgent need to review the present strategy for controlling relapses in vivax malaria, at least for the city of Mumbai, and similar studies need to be carried out in other parts of India, to make all anti relapse strategies more appropriate. PMID- 10715674 TI - Poor gametocytocidal activity of 45 mg primaquine in chloroquine-treated patients with acute, uncomplicated, Plasmodium falciparum malaria in Mumbai (Bombay): an issue of public-health importance. AB - In the city of Mumbai (formerly Bombay), chloroquine (CQ) continues to be recommended as the drug of first choice for the treatment of Plasmodium vivax and P. falciparum infections, even though > 50% of local isolates of P. falciparum are resistant to it. Primaquine, an 8-aminoquinoline is also given to patients with falciparum malaria, in a single, 45-mg dose, to kill the gametocytes and so reduce transmission. The gametocytocidal activity of supervised primaquine (45 mg given on day 8) was investigated in 90 patients who had been treated with CQ. Of these, 15 were found to be CQ-sensitive patients, 61 were resistant (49, eight and four considered RI, RII and RIII, respectively) and 14 were lost before completion of the follow-up. The mean (S.D.) baseline gametocytaemias in the CQ sensitive and RI-resistant cases were 665.1 (411.3) and 1537.4 (1045.5)/microliter, respectively. Despite supervised primaquine treatment, four of the 15 CQ-sensitive patients and 32 of the 49 patients found to be RI resistant had gametocytes on day 29. There therefore appears to be a need to review the current, gametocytocidal, primaquine-dosage schedule and to re-treat patients who remain gametocytaemic with higher doses of primaquine, as an important, transmission-blocking strategy. PMID- 10715675 TI - Evidence for the occurrence of Trypanosoma brucei rhodesiense sleeping sickness outside the traditional focus in south-eastern Uganda. AB - The occurrence of Trypanosoma brucei rhodesiense west of the River Nile, in Masindi district in the mid-western part of Uganda, is confirmed. Masindi borders the traditional belt of T. b. gambiense infection in the north-west, Gulu in the north and the Democratic Republic of Congo in the west. Of the 702 persons tested for sleeping sickness in Masindi, 113 (16%) were positive by the card agglutination test for trypanosomiasis (CATT). Trypanosomes were observed in samples of cerebrospinal fluid (CSF) from two (0.3%) of the subjects: a 7-year old girl, who had been ill for 2 weeks and yet was in good general condition, with three white blood cells (WBC)/microliter CSF; and a 47-year-old woman who had been ill for 8 months, looked sickly, had seven WBC/microliter CSF, but was still able to dig in her gardens. Rats and mice inoculated with blood from the two parasitologically confirmed cases became parasitaemic on day 3 post inoculation, indicating that the parasites were T. b. rhodesiense. Isoenzyme analysis revealed that the parasites isolated from one of these confirmed cases belonged to a zymodeme (449) which has not been previously observed among isolates from south-eastern or north-western Uganda. Although the isolate shared PGM2 and ICD3 patterns with T. b. gambiense and T. b. rhodesiense, respectively, it did not have the SOD3:5 pattern characteristic of T. b. gambiense. The spread of T. b. rhodesiense beyond its traditional focus and the development of areas where this subspecies and T. b. gambiense are co-endemic will complicate the control of sleeping sickness in Uganda; although the CATT is very useful for the mass screening of populations for T. b. gambiense area, it is not applicable in the detection of T. b. rhodesiense. PMID- 10715676 TI - Haemoglobin concentrations and infection by Giardia intestinalis in children: effect of treatment with secnidazole. AB - The blood concentrations of haemoglobin were investigated in 82 children aged 2-9 years. Fifty-seven (31 boys and 26 girls) were stool-positive for Giardia intestinalis but the other 25, used as controls, were negative. The mean (S.D.) haemoglobin concentration among the infected children was significantly lower pre treatment than that for the control group [11.6 (1.2) v. 12.6 (1.5) g/dl; P < 0.05]. Treatment of the infected children with a single oral dose of secnidazole (30 mg/kg) led to a significant increase in their mean haemoglobin level 15 days later, from 11.6 (1.2) g/dl pre-treatment to 12.4 (1.2) g/dl post-treatment (P < 0.05). The results indicate that the therapeutic control of giardiasis could be important in programmes to combat anaemia in children living in endemic areas. PMID- 10715677 TI - Symptomatic and asymptomatic amoebiasis in two heterosexual couples. AB - Four cases of amoebiasis are described: two symptomatic with intestinal and hepatic involvement and two asymptomatic, diagnosed in two, heterosexual, Italian couples. Infection was probably acquired first by the men, via an indirect faccal oral route, and then transmitted to their partners in the same way. The two amoebic strains isolated, from the woman of one couple and the man of the other, were characterized by electrophoresis as zymodemes II alpha- and XIX of Entamoeba histolytica. These four cases emphasise once more the role of cyst-passers in the spread of infection and the importance of biochemical identification of the amoebic isolates, enabling more specific treatment. PMID- 10715678 TI - Analysis of climatic data and forecast indices for human fascioliasis at very high altitude. AB - Human infection with Fasciola hepatica has recently been recognized as an important health problem worldwide, and particularly at very high altitudes in South America. The highest prevalences and intensities of human fascioliasis known are those of the northern Bolivian Altiplano, where infected Lymnaea truncatula occur at altitudes of 3800-4100 m. In the present study, the climatic data for this area of the Altiplano, which differ markedly from those of endemic areas in the lowlands, were analysed. There is no marked seasonality in temperature but there are large variations in temperature within a daily, 24-h period. Rainfall is seasonal, with a long dry season, coinciding with the lowest minimum temperatures, and a long wet season. The rate of evapotranspiration is very high, and temporary water bodies dry out very quickly. Solar radiation at ground level is intense, not only because of the altitude but also because of the lack of trees and shrubs. Two climatic indices for forecasting fascioliasis, Mt and Wb-bs, were calculated. Modifications in these forecast indices are proposed, to reflect the environment at high altitude and low latitude. Estimates, based on climadiagrammes, of the durations of the wet and dry seasons were greatly effected by the inclusion of an aridity-index modification. The usefulness of the modified indices was examined using prevalence data for human and cattle fascioliasis collected in the neighbourhoods of the stations providing the meteorological data. Values for both indices indicated that conditions were optimum for transmission between December and March. The results were statistically significant for the modified Wb-bs index when the data for a meteorological station in which no lymnaeids were found were excluded. The modified Mt index did not appear sufficiently accurate to be useful. The values for the modified Wb-bs index permitted the study areas to be designated low-, moderate- or high-risk areas for the transmission of fascioliasis to man and domestic animals. PMID- 10715679 TI - Polyamines: agents with macrofilaricidal activity. AB - There is a need for effective macrofilaricidal drugs. The polyamine metabolism of filarial worms has been recognized as a possible target for effective drug action. In an attempt to identify agents that might provide leads in developing an effective macrofilaricide, 78 polyamine compounds were selected from among > 250,000 structures that have been amassed by the Walter Reed Army Institute of Research, in the U.S.A. These thousands of agents have been chosen principally for drug-development programmes for other parasitic diseases. The 78 prospective drugs selected were evaluated for their macrofilaricidal activity against Brugia pahangi and Acanthocheilonema viteae, in male Mongolian jirds (Meriones unguiculatus). The animal models using these two parasites were designed to mimic, in so far as possible, human lymphatic filariasis and onchocerciasis, respectively. Thirteen of the compounds were found to be active although none of these has been previously reported to be macrofilaricidal. Two were suppressive for B. pahangi and 11 for A. viteae. These active agents may represent a nucleus around which highly effective drugs can be synthesised. PMID- 10715680 TI - The distribution of helminth infections along the coastal plain of Kwazulu-Natal province, South Africa. AB - The results of a previous study indicated that, in the province of KwaZulu-Natal, South Africa, Necator americanus and Strongyloides stercoralis were endemic to the coastal lowlands only. The prevalences of these helminths, as well as those of Trichuris trichiura and Ascaris lumbricoides, have now been investigated along a 1000-km-long transect through the coastal plain, at altitudes of < 300 m, from the Mozambique border (26 degrees 57'S) to the border with Eastern Cape province (30 degrees 53'S). Necator americanus was by far the most dominant hookworm species. Although prevalences of N. americanus and S. stercoralis infection decreased with increasing southerly latitude, those of T. trichiura and A. lumbricoides did not. Determinants of these distribution patterns are examined in terms of a suite of temperature- and rainfall-related variables. PMID- 10715681 TI - An unusual 'infection' of a child in Sri Lanka, with Taenia taeniaeformis of the cat. PMID- 10715682 TI - Malaria in pregnancy. PMID- 10715683 TI - Malaria in pregnant women: research, epidemiology, policy and practice. PMID- 10715684 TI - Epidemiological and control issues related to malaria in pregnancy. AB - Pregnant women infected with malarial parasites have an increased risk of maternal anaemia, abortion, stillbirth, prematurity, intra-uterine growth retardation, and infants of low birthweight. A 'state-of-the-art' symposium on malaria in pregnancy was convened in Kisumu, Kenya, in November 1997, to discuss the biological and clinical impact of malaria in pregnancy, and to identify antimalarial drugs and control strategies to protect pregnant women. The deleterious effects of malarial infection during pregnancy were shown to be associated both with Plasmodium falciparum and P. vivax infections, and to occur under a wide range of malaria transmission pressures. Control interventions, thus, need to be targeted at pregnant women in all endemic areas. Alternative antimalarial drugs to chloroquine have been tested and shown to be effective (and safe) against malaria in pregnancy. Delivery of cost-effective control interventions has been explored; investments are needed to facilitate the scaling up of successful approaches to national-programme level. Several important research questions related to malaria in pregnancy were highlighted at the Kisumu meeting. Increased international and local commitment, to resource effective malaria control in pregnancy adequately, is a public-health priority. PMID- 10715685 TI - The Thai-Burmese border: drug studies of Plasmodium falciparum in pregnancy. AB - Plasmodium falciparum malaria is increasing world-wide, as is resistance to the available antimalarials. On the Thai-Burmese border this problem is most acute in pregnant women, as options for their treatment are even more restricted because of the unknown effects of antimalarials on the foetus. Presented here are the results of descriptive, clinical, drug studies on quinine, mefloquine and artemisinin derivatives for P. falciparum in pregnant women. Mefloquine and quinine have high failure rates for primary and recrudescent infections. Artemisinin-based treatments in pregnant women have proved safe, tolerable and efficacious. However, randomized drug studies with these drugs and other new antimalarials are required to define the true safety and efficacy of these drugs in pregnant women. PMID- 10715686 TI - Malaria in pregnancy and its consequences for the infant in rural Malawi. AB - Maternal malaria and anaemia, pregnancy and infant outcomes are reviewed among a cohort of mothers and their babies living in Chikwawa district, southern Malawi. Overall, 4104 women were screened at first antenatal visit and 1523 at delivery. Factors independently associated with moderately severe anaemia (MSA; < 8 g haemoglobin/dl) in primigravidae were malaria (relative risk = 1.9; 95% confidence interval = 1.6-2.3) and iron deficiency (relative risk = 4.2; 95% confidence interval = 3.5-5.0). Only iron deficiency was associated with MSA in multigravidae. After controlling for antimalarial use, parasitaemia was observed in 56.3% of the HIV-infected primigravidae and 36.5% of the non-infected (P = 0.04). The corresponding figures for multigravidae were 23.8% and 11.0%, respectively (P = 0.002). Over 33% of the infants born alive to primigravidae were of low birthweight (LBW; < 2500 g), and 23.3% of all newborns had foetal anaemia (< 12.5 g haemoglobin/dl cord blood). LBW was significantly associated in primigravidae with pre-term delivery, placental malaria and frequency of treatment with sulfadoxine-pyrimethamine (SP), and in multigravidae with pre-term delivery, adolescence, short stature and MSA. LBW was significantly reduced with a second SP treatment in primigravidae, and with iron-folate supplementation in multigravidae. Mean haemoglobin concentrations were significantly lower in the infant who had been LBW babies than in the others, and significantly associated with parity, peripheral parasitaemia at delivery and placental malaria. At 1 year post-delivery, life status was known for 364 (80.7%) of the 451 infants enrolled in the follow-up study. Independent risk factors for post-neonatal mortality were maternal HIV infection, LBW, and iron deficiency at delivery. This study identifies priorities for improving the health of pregnant women and their babies in this rural area of Malawi. PMID- 10715687 TI - The placenta in malaria: mechanisms of infection, disease and foetal morbidity. AB - There is still much to discover about the reasons for the increased susceptibility of pregnant women to malaria or the pathogenesis of placental malaria. More systematic and detailed examination of the placenta may help. In many ways, the placenta can be seen as the flight recorder of the pregnancy; by examining it carefully it should be possible to tell much about how smooth a 'flight' the mother and baby experienced. It is hoped that, by probing the secrets of this 'squishy black box', the causes of adverse effects in pregnancy are elucidated, and the safe 'travel' of babies and their mothers in the future is ensured. In this review, the features of parasite accumulation in the placenta, parasite adherence, and hormonal and inflammatory responses to placental malaria are discussed, focussing on infection with Plasmodium falciparum. The results of recent research indicating an interaction between HIV and malaria in pregnancy are summarized. Ten questions for basic researchers are posed. The answers may help direct future efforts to control malaria in pregnancy. PMID- 10715688 TI - A birthweight nomogram for Africa, as a malaria-control indicator. AB - Low birthweight (LBW) attributable to malaria in pregnancy is a significant risk for millions in Africa. Infants born to primigravidae are at greatest risk and it is proposed that this excess risk can be used as a simple indicator of malaria transmission and exposure in pregnant women in Africa. Birthweight data from different regions in 11 malarious and three non-malarious African countries were investigated. A regression analysis of the excess risk of low birthweight in first pregnancies, compared with later ones, was completed and interpreted in relation to malaria-transmission intensities. The aim was to develop a simple birthweight chart (nomogram) as a tool for monitoring malaria transmission or malaria control in pregnancy. Low-birthweight risk in first pregnancies was associated with levels of malaria-transmission intensity amongst different African countries. The nomogram distinguished longitudinal changes in malaria exposure, related to season and changes in antimalarial-drug policy. Malaria is one of the most important causes of LBW in first pregnancies in Africa. As birthweight and parity are routinely recorded in many delivery centres across Africa, the nomogram provides a simple, available and inexpensive tool for monitoring malaria transmission and exposure in pregnant women and the effectiveness of malaria-control activities for this high-risk group. PMID- 10715689 TI - Malaria in pregnancy: its relevance to safe-motherhood programmes. AB - Severe anaemia in pregnancy is an important contributor to maternal and perinatal morbidity and mortality. In sub-Saharan Africa severe anaemia in pregnancy is very common, the main causes being iron and folate deficiency, malaria, hookworm infestation and advanced HIV infection. Though most of these causes are preventable, the overall prevalence of anaemia has not changed over many years. This is probably due to a mixture of reasons, including operational problems and inadequate interventions. In addition, a true effect on severe anaemia may have been missed if the only measure taken is of the overall prevalence of anaemia. One cause of anaemia that has been neglected by safe-motherhood programmes has been malaria in pregnancy. In endemic areas, malaria in pregnancy is usually asymptomatic and often associated with a negative peripheral-blood film. Hence the condition needs to be treated and prevented as a matter of routine in all women at risk of infection. A trial conducted in Kenya demonstrated that intermittent treatment with the antimalarial sulfadoxine-pyrimethamine (SP), given a couple of times during pregnancy when women attend for antenatal care, can reduce severe anaemia in primigravidae by 39%. The results of this study demonstrate the important contribution of malaria to severe anaemia in pregnancy in areas of endemic transmission. Intermittent treatment with SP in pregnancy has also been shown to be effective in improving birthweight. Though questions remain about the optimal way to deliver this intervention to different groups of women, we cannot afford to wait for all of the answers. The degree to which malaria contributes to severe anaemia in pregnancy is now clear. In Kenya intermittent SP is now policy for pregnant women from malarious areas. The challenge now is for this regimen to be successfully implemented as part of an integrated programme of anaemia control in pregnancy. PMID- 10715690 TI - Malaria and pregnancy: the implementation of interventions. AB - The choice of interventions for improving malaria control during pregnancy depends on several factors. These include the efficacy of the intervention, consumer acceptability and compliance, provider acceptability, cost, safety, integration with other interventions and the local system of health-care delivery, and the degree of combination of these factors. For successful implementation the following should be recognized: appropriate policy formulation based on a process of advocacy, research and demonstration programmes; regulation and legislation; resource mobilization; consumer awareness; and appropriate delivery, targeting, monitoring and evaluation. Malaria in pregnancy is a priority area for control and a major public-health problem. Improved control of such disease requires better integration into health-care practices. PMID- 10715691 TI - Priority areas for current research on malaria during pregnancy. PMID- 10715692 TI - A rational approach to malaria control in pregnancy in sub-Saharan Africa: the need for a link between scientific research and public-health interventions. PMID- 10715693 TI - The host response in malaria and depression of defence against tuberculosis. AB - Malaria causes significant morbidity and mortality world-wide. Both asymptomatic and symptomatic malarial infections cause immune depression, which predisposes the host to infection with other microorganisms. Specific clinical investigations have shown, for example, that those with malaria-attributable anaemia are particularly likely to have Salmonella septicaemia, and that asymptomatic malarial infection causes diminished response to polysaccharide vaccine. The results of clinical studies and experiments with animal models have revealed that malarial parasites can decrease their vertebrate host's effective humoral and cellular immune responses. In this review, the possible ways in which this malaria-induced immune impairment could affect the host's response to Mycobacterium tuberculosis infection are considered. Could malarial infection be one of the reasons for the persistence of tuberculosis in malaria-endemic regions? PMID- 10715694 TI - Strong association, but incomplete correlation, between chloroquine resistance and allelic variation in the pfmdr-1 gene of Plasmodium falciparum isolates from India. AB - The increasing prevalence of chloroquine-resistant Plasmodium falciparum has complicated the control of falciparum malaria. It has been suggested that point mutations at nucleotide positions 754, 1049, 3598, 3622 and 4234 in the parasite's pfmdr-1 gene are associated with such resistance, although this is a matter of controversy. Eighteen chloroquine-sensitive and 22 resistant isolates of P. falciparum from India were investigated, to examine the role of the pfmdr-1 gene in the resistance, and to determine whether any of the point mutations could be used as a marker for the rapid identification of the chloroquine-resistant strains. As this investigation failed to reveal an explicit association between allelic variation in the pfmdr-1 gene and chloroquine resistance, the use of point mutations to identify the resistant strains does not appear feasible. PMID- 10715695 TI - The diagnosis of Plasmodium falciparum infection in Gambian children, by field staff using the rapid, manual, ParaSight-F test. AB - A rapid immunodiagnostic test for Plasmodium falciparum, the ParaSight-F test, was evaluated in the diagnosis of malaria in 139 children with uncomplicated malaria, who presented at the Medical Research Council's clinic at Basse in Upper River division, The Gambia. The aim was to evaluate the performance and usefulness of the test as a diagnostic method in a malaria-endemic area, when performed by a field worker. Compared with microscopy, the test had a sensitivity of 96.5%, a specificity of 90.5%, a negative predictive value of 94.2% and a positive predictive value of 94.3%. Because of its sensitivity, specificity and simplicity, the ParaSight-F test will be of value in situations where microscopy is not possible. PMID- 10715696 TI - An improved, PCR-based strategy for the detection of Trypanosoma cruzi in human blood samples. AB - Attempts were made to improve the PCR-based detection of Trypanosoma cruzi in blood samples, primarily for screening blood donors. Samples were obtained from candidate donors who were reactive in one or two of three serological tests for Chagas disease (and therefore considered 'indeterminate') or in all three tests (3+). Each sample was then examined using three different, PCR-based techniques: 'PCR-I' (in which the target DNA is a nuclear repetitive sequence); 'PCR-II' [amplifying a conserved region of the T. cruzi kinetoplast DNA (kDNA)]; and 'PCR III' (a new strategy in which the target kDNA is amplified by 'nested' PCR). Among the samples from 3+ individuals, PCR-I, PCR-II and PCR-III amplified two (3.8%) out of 52, four (4.5%) out of 88, and 27 (25.7%) out of 105 samples tested, respectively. Seven, 69 and 70 samples from 'indeterminate' subjects were tested by PCR-I, PCR-II and PCR-III, respectively; there was not a single positive result by PCR-I or PCR-II, but three (4.3%) of the samples tested by PCR III were positive. In a reconstruction experiment, in conditions in which PCR-I and PCR-II could not detect 10,000 parasites/ml, PCR-III was able to detect one parasite/ml. Although all three PCR-based strategies examined had rather poor sensitivities, PCR-III was far more sensitive than PCR-I or PCR-II. PMID- 10715697 TI - Thioridazine treatment modifies the evolution of Trypanosoma cruzi infection in mice. AB - Thioridazine, a tricyclic drug, is known to have a direct effect on Trypanosoma cruzi, disrupting the parasites' mitochondria and kinetoplasts. In the present study, the drug was used orally, at 80 mg/kg.day for 3 days, to treat mice inoculated with low numbers of T. cruzi. The drug caused no apparent toxicity in the host. It cleared trypomastigotes from the bloodstream, prevented the histological and functional alterations of the heart normally observed in the chronic phase of the experimental disease, and greatly reduced the mortality rate compared with that in untreated, infected controls. When checked 135 days post infection, the density of cardiac beta receptors and the cardiac histology of the treated mice were indistinguishable from those of uninfected, untreated controls. The drug is already used to treat humans, as a neuroleptic drug. It appears to be able to prevent acute infection with T. cruzi evolving into chronic disease, at least in mice, and may be a useful base from which to design new agents for the treatment of Chagas disease. PMID- 10715698 TI - Evaluation of a standardized direct agglutination test (DAT) for the diagnosis of visceral leishmaniasis in Kenya. AB - A prototype test kit being developed, by the World Health Organization (WHO), for the diagnosis of visceral leishmaniasis (VL) was evaluated in the Baringo district of Rift Valley province in Kenya. The screening of approximately 10,000 individuals for the signs of VL produced 305 suspected cases. These cases and 304 controls matched for sex and age (+/- 2 years) were then tested with the kit, which is based on a direct agglutination test (DAT). The evaluation was a three stage process. The first stage, the field screening, involved screening filter paper samples of dried blood from the suspects and controls at a DAT titre of 1:500. The second stage, the laboratory titration, involved screening of the same individuals by testing freshly eluted filter-paper samples at 1:500 to 1:2000 dilution. In the third stage, the full-scale titration, all samples that had been positive at 1:2000 were titrated at 1:500-1:512,000. All the suspects giving DAT titres of 1:2000 or higher were considered positive for VL. This diagnosis was checked, whenever possible, by the examination of smears and/or cultures of splenic aspirates for leishmanial parasites. Those found to be parasitologically positive were put on a standard treatment regime of 20 mg sodium stibogluconate (Pentostam)/kg.day. Although 42 (13.8%) of the 305 clinical suspects investigated were DAT-positive (at 1:2000), it was only possible to take splenic aspirates from 32. Four (12.5%) of these 32 were apparently false-positives by DAT, as no parasites could be detected in their splenic aspirates. The others provided positive smears and cultures (27 cases) or a negative smear but a positive culture (one case). It was possible to re-examine two of the four serologically positive but parasitologically negative VL suspects at a 3-month follow-up: neither had a palpable spleen, one had seroconverted and the other had much lower DAT titre (1:32,000) than when investigated previously (1:128,000). All the parasitologically confirmed cases remained DAT-positive (1:2000) at this follow up. The low cut-off titre (1:2000) and the simple procedure should make the kit suitable for use by health workers at all levels of primary-health care, including those with limited training and skills, for screening rural communities at risk of VL. PMID- 10715699 TI - Modification of behaviour and attitude in the control of schistosomiasis. 1. Observations on water-contact patterns and perception of infection. AB - Observations on the water-contact patterns of 2136 residents of Admin community in Nigeria were conducted at four streams between February 1993 and January 1994. Urine samples collected from those observed were used to estimate the prevalence and intensity of Schistosoma haematobium infection. A questionnaire was also completed for each of the subjects, to test their perception of urinary schistosomiasis and its transmission. Infection was detected in 1076 (50.4%) of the subjects, with peak prevalence among those aged 10-14 years. Intensity of infection was more closely correlated with the number of water contacts (r = 0.97) than with the total duration of the exposure (r = 0.77), emphasising the importance of specific/multiple activities, and of the surface area of the body submerged, in transmission. One stream (Culvet) was identified as the main transmission point, with bathing/swimming and fishing as the main activities predisposing people to infection. The awareness of urinary schistosomiasis and its symptom (blood in urine) were high but specific knowledge about the parasite, its vector and the interaction between the parasite and vector in the parasite's life-cycle were extremely low. Activities that require behaviour and attitude modification have been identified and encouraged as components in the control of schistosome-attributable morbidity (in the absence of pipe-borne water). PMID- 10715700 TI - Cross-validation of the rapid epidemiological mapping of onchocerciasis endemicity in Anambra state, Nigeria. AB - In this evaluation study, the results of a previous, rapid, epidemiological mapping of onchocerciasis (REMO) from quota samples of six communities (two hyper , two meso-, one hypo- and one non-endemic) in Anambra state, Nigeria, were cross validated. The findings were based on observations on the prevalence of palpable onchocercal nodules and skin dipigmentation (leopard skin) in a sample of 50 adults resident in each community for at least 5 years. They indicate that the REMO results obtained earlier were reasonably valid. The influence of several factors (mobilization techniques, mobilization message, previous exposure of subjects to previous research studies and treatment intervention, community members' perception of susceptibility and their desire to benefit from treatment programmes) on the representativeness of cases and non-cases in the samples used for REMO is discussed. PMID- 10715701 TI - The community-directed, ivermectin-treatment programme for onchocerciasis control in Uganda--an evaluative study (1993-1997). AB - The first 5 years of a community-directed, ivermectin-treatment programme, to control onchocerciasis in 1805 endemic communities in 10 districts in Uganda, are evaluated. Each year, the desired treatment coverage of the population eligible to take invermectin (90%) was achieved in 42.6%-51% of the 1713 communities for which complete data were available; 67%-74.8% achieved 80% coverage. The annual cost per person treated with ivermectin (ACPTI) was much higher in the districts with small populations to be treated (< 15,000) than in those with large populations (> 40,000) (U.S.$0.40 v. U.S.$0.10 or less). The community members' acceptance of the programme was related to their attendance at health-education sessions (P = 0.009), and their participation in the mobilisation of other community members increased greatly when they were allowed to take part in the selection of the community-based distributors (CBD) and the choice of treatment sites. The overall target ratio of one CBD/71 families was attained by 1997. However, the failure of some trained CBD to participate in the treatment exercise prevented some communities achieving 90% treatment coverage. Providing CBD with cash incentives or externally derived incentives 'in kind' proved counter productive whereas locally generated incentives 'in kind' were simply regarded as the normal obligations of the community. District health staff successfully integrated the programme with their other health commitments, but the involvement of CBD in other programmes proved detrimental to their performance. Other constraints identified were rebel insurgency in some areas, and abnormally heavy rains in hilly areas with poor roads. PMID- 10715702 TI - The predominant genotypes of hepatitis B virus in Thailand. AB - In Thailand, chronic liver disease (CLD) as a consequence of infection with hepatitis B virus (HBV) constitutes a public-health burden. Control and treatment are complicated by the virus exhibiting an unusually high mutation rate, with some genotypes apparently causing more severe disease than others. Restriction fragment-length-polymorphism (RFLP) analysis of the pre-S region of the viral genome, amplified by PCR, was used to determine which genotypes were most prevalent among Thai patients chronically infected with the virus. The patients were chronic HBV carriers (40) or cases of chronic hepatitis (34), cirrhosis (14) or hepatocellular carcinoma (30). As indicated by the results of earlier studies on CLD patients in South-east Asia, genotype C (68.6%) was clearly predominant. RFLP patterns permitted the C1 (12.7%), C7 (45.7%), C8 (10.2%) and B1 (29.7%) subtypes to be identified. Two samples that could not be typed by RFLP were analysed by direct sequencing, categorized as type C, and tentatively designated as subtype C9. As comparison of the present data with those previously obtained by direct sequencing of PCR products indicates that RFLP analysis is as specific and reliable as sequencing and less expensive and time-consuming, RFLP analysis may be particularly useful for epidemiological studies. PMID- 10715703 TI - Acute, hepatitis-A super-infection in HBV carriers, or chronic liver disease related to HBV or HCV. AB - The impact of acute super-infection with hepatitis A virus (HAV) was determined in 20 asymptomatic carriers of the surface antigen (HBsAg) of hepatitis B virus (HBV), eight patients with HBV-related chronic liver disease (CLD), and four patients with CLD related to hepatitis C virus (HCV). For comparison, 100 patients with isolated HAV infection were also studied. The HBsAg carriers and patients with CLD related to HBV or HCV were significantly older than the patients with isolated HAV infection, with mean (S.D.) ages of 43.9 (14.1), 46.4 (16.0), 52.5 (8.6) and 28.4 (10.7) years, respectively (P < or = 0.02). There were no significant between-group differences in the baseline serum concentrations of alanine aminotransferase. All the patients with isolated HAV infection fully recovered. Fulminant or submassive hepatitis occurred in 11 (55%) of the HBsAg carriers and four (33%) of the 12 patients with CLD related to either HBV or HCV. Nine of the 15 patients with severe hepatitis died and the mortality rate among the HBsAg carriers was not significantly different from that among the CLD patients (25% v. 33%; P = 0.15). These fatal cases were all aged > 50 years and were significantly older [59.0 (2.1) years] than the six severe cases who recovered [43.2 (10.7) years] as well as the remaining 17 uncomplicated cases with CLD or HBsAg [40.3 (13.0) years] (P < or = 0.001). The results indicate that acute HAV is rarely fatal in young adults but may be severe and potentially fatal in patients with underlying chronic HBV or HCV infection, especially among the elderly. Vaccination against HAV should be considered for the patients at high risk who are negative for anti-HAV. PMID- 10715704 TI - Morphometric characterization of members of the Simulium damnosum Theobald complex (Diptera: Simuliidae) from East and West Africa. AB - Morphometric multivariate analyses were carried out in order to separate the females of the closely related anthropophilic and non-anthropophilic members of the Simulium damnosum s.l. complex occurring in western Uganda. Simulium kilibanum (= 'Nyamagasani'), 'Nkusi', 'Sebwe' and S. pandanophilum were compared with the West African S. damnosum subcomplex, S. soubrense and S. yahense to correlate the results with those of previous studies. Simulium pandanophilum could be clearly discriminated. However, only 72%-88% of identifications among the closely related S. kilibanum, 'Sebwe' and 'Nkusi', which all belong to the 'Sanje' subcomplex, were correct. The two forest species S. soubrense and S. yahense slightly overlapped but were distinct from all others. PMID- 10715705 TI - Prevalence of syphilis and parasitic infection among blood donors in a tertiary care centre in southern India. PMID- 10715706 TI - A non-lethal, autosomal, recessive, melanotic mutant of Anopheles minimus species A. PMID- 10715707 TI - Clinical research in the tropics: some thoughts for the beginner. PMID- 10715708 TI - Response to antimicrobial therapy in childhood bacterial meningitis in tropical Africa: report of a bi-centre experience in Nigeria, 1993-1998. AB - Recent reports of a high prevalence of in-vitro resistance to chloramphenicol (CHL) and penicillin (PEN)/ampicillin (AMP) cause concern because of cost implications in using the newer cephalosporins (CEPH) to treat meningitis in resource-poor countries. However, the clinical significance of many of the observations is uncertain because of limited back-up by clinical data. We analysed the response in an open study of 161 patients with bacterial meningitis treated with CHL (n = 31), CHL plus PEN or AMP (n = 101), PEN or AMP (n = 14) and CEPH (n = 15). No significant differences were observed in clinical course and outcome in the four treatment groups, other than a higher prevalence of seizures after 72 h of treatment and a higher prevalence of neurological sequelae in survivors in the CEPH and CHL groups. This may reflect the higher number of infants and greater frequencies of uncommon aetiological agents in the CHL and CEPH groups. It is concluded that response to initial chloramphenicol-based treatment regimens remains satisfactory and that there is as yet no compelling reason to switch to the cephalosporins as first-line therapy for bacterial meningitis in developing countries. PMID- 10715709 TI - Pre-school follow-up of a cohort of children with myelomeningocele in Cape Town, South Africa. AB - A cohort of South African children with myelomeningocele was followed for 5 years. Fifty-three were from metropolitan Cape Town and 65 from rural areas including the tribal region of Transkei. The mean general developmental quotient (GQ) at 5 years of age was lower than that previously reported. Black and coloured children had lower GQs than white children, but there were no significant differences between those from the urban and the rural areas. Early closure of the lesion and delivery by caesarean section were associated with higher levels of general developmental functioning. Central nervous system infections and the placement of more than one shunt for hydrocephalus resulted in lower GQs. Lesions above L2 were associated with non-ambulation. Ambulation was more likely in children in Cape Town than in rural children. Urinary incontinence occurred more frequently in rural children and among those in lower socio economic circumstances. PMID- 10715710 TI - Henoch-Schonlein purpura and streptococcal infection: a prospective case-control study. AB - A prospective, matched, case-control study conducted over a period of 3 years was designed to examine the association of group A beta-haemolytic streptococcal infections and Henoch-Schonlein purpura. Demographic and clinical data were collected as well as measurement of antistreptolysin O titres and throat swab culture on all children admitted with Henoch-Schonlein purpura, as well as their matched controls. Antistreptolysin O titre positivity was associated with a 10 fold increase in the risk of Henoch-Schonlein purpura. Renal involvement was common among cases with positive antistreptolysin O titres (27%) compared with cases with a negative titre (8%) but this difference has no statistical significance. PMID- 10715711 TI - Breastfeeding, socio-economic conditions and nutritional status of children younger than 12 months in Brazil. AB - Taking account of socio-economic determinants such as maternal education and family income, nutritional status and its relationship to breastfeeding and socio economic conditions were studied in 419 children aged 3-12 months in the city of Niteroi in south-eastern Brazil. Data were obtained by sampling a population during a high coverage (90%) vaccination campaign in 1992. After stratifying by maternal educational level, the relative risk (prevalence ratio) for height-for age (H/A) Z-score < -1 in relation to no breastfeeding was 2.2 (95% CI 1.1-4.2) for families where mothers had been educated for less than 4 years and 0.7 (95% CI 0.4-1.1) otherwise, indicating a significant interaction (modifying effect) between breastfeeding and the control variable (chi 2 = 7.4; p = 0.006). Similar results were found when family income was used as the stratification variable (RR 2.2, 95% CI 1.1-4.3 in the lower and RR 0.7, 95% CI 0.4-1.2 in the higher income stratum; chi 2 = 6.7; p = 0.009). The population-attributable risk fraction indicated that in the age group analysed a reduction of about 40% in the number of children with HAZ < -1 could be achieved in low-income/low-education families by the widespread adoption of breastfeeding. PMID- 10715712 TI - Pyomyositis in children: analysis of 31 cases. AB - In a 10-year period, 31 children with 35 pyomyositis were managed in Zaria, northern Nigeria. Twenty-two (71%) were less than 10 years of age, with a peak incidence at between 5 and 9 years. The leg muscles, mainly the quadriceps, were most frequently involved (51.4%), followed by the trunk muscles (25.7%), predominantly those of the anterior abdominal wall. Arm and shoulder girdle muscles were less frequently affected (11.4% each). Staphylococcus aureus was the most frequently cultured organism (75%) and was usually sensitive to cloxacillin and, to a lesser degree, to erythromycin and chloramphenicol. Incision and adequate drainage was usually very effective with recurrence at only one site. Antibiotics were used routinely. Involvement of the heart and lungs occurred in two children respectively, the former causing the only death. The average duration of hospital stay was 20 days. PMID- 10715713 TI - Typhoid ileal perforation in children: a scourge in developing countries. AB - Over a 10-year period, 64 children aged < or = 12 years were treated for typhoid perforation, accounting for 56% of all cases of typhoid perforation at our institution. The perforation rates in the age groups < 1, 1-4, 5-9 and 10-12 years were 4%, 1.7%, 12.4% and 29.3%, respectively, with an overall perforation rate of 10.3%. The main features were fever (93.4%) and abdominal pain and tenderness (93.4%). Thirteen children (20.3%) had associated haemorrhage, presenting as haematochezia. The incidence of perforations was 52% during the rainy season and 48% during the dry season, but the disease occurred throughout the year with a peak in October, the beginning of the dry season, which was also the time of peak occurrence of typhoid without perforation. An average of 14 h (range 5-30) was required for resuscitation. Ketamine was used for anaesthesia in most cases. Treatment was by segmental resection (67%), wedge excision (17%) and simple closure (6%). Morbidity was high (53%), and wound infection (53%) and chest infection (30%) were the most common complications. There were 25 deaths (39%), most the result of overwhelming sepsis. Late presentation at > 7 days was associated with high mortality (p < 0.05). Typhoid perforation continues to be a scourge in children in developing countries and, in addition to preventive measures such as improved sanitation and the provision of safe water supplies, public enlightenment is necessary to ensure early presentation and improved survival. PMID- 10715714 TI - Study of the prevalence of and high risk factors for fetal malnutrition in term newborns. AB - This observational study was done to discover the prevalence of fetal malnutrition (FM) in term newborns using clinical assessment of nutritional status (CANS score) and to identify associated risk factors. All term babies born in a referral teaching hospital during the 1-year study period were included in the sample. Gestational age and weight-for-gestational-age were assessed, and babies were classified as appropriate-for-gestational-age (AGA), small-for gestational-age (SGA) or large-for-gestational-age (LGA). Maternal risk factors were recorded in each case. Fetal malnutrition was present in 19.6% of babies, of whom 40.7% had intrauterine growth retardation. Of the babies with FM, 59.9% were AGA and 1.9% were SGA even though they had no signs of FM. FM was evident in 84.2% of SGA babies, and 12.9% of AGA babies showed FM. The weights of babies with FM were significantly lower than of those without FM. Maternal risk factors for FM included adverse age, primiparity, low pre-pregnancy weight and height, a bad obstetric history and pregnancy-induced hypertension. Malnutrition in the newborn might be missed if intrauterine growth curves only are used for assessment. The CANS score is a simple and rapid clinical scoring system for diagnosing fetal malnutrition. Not all SGA babies are malnourished and those without FM have a better outcome and faster catch-up growth. PMID- 10715715 TI - Are orphans at increased risk of malnutrition in Malawi? AB - The objective of this study was to compare the nutritional status and health problems of village orphans, non-orphans and orphanage children, and to identify factors associated with undernutrition. A cross-sectional study was conducted in three orphanages and two villages near Blantyre, Malawi. Seventy-six orphanage children, 137 village orphans and 80 village non-orphans were recruited. Anthropometric measurement was done and guardians were interviewed. In the group of children aged < 5 years, the prevalence of undernutrition in orphanage children was 54.8% compared with 33.3% and 30% of village orphans and non orphans, respectively. Sixty-four per cent of young orphanage children were stunted compared with 50% of village orphans and 46.4% of non-orphans. The mean (SD) Z-score of height/age was significantly lower in the orphanage group, -2.75 (1.29) compared with -2.20 (1.51) and -1.61 (1.57) in the village orphan and non orphan groups (p < 0.05). Conversely, older orphanage children (> or = 5 years) were less stunted and wasted than orphans and non-orphans in villages. Illness of children in the last month was reported to be higher in the non-orphan group, especially diarrhoeal disease, which occurred in 30% compared with 10.8% of village orphans and 6.6% of orphanage children. More than three children in a family being cared for by guardians was significantly associated with undernutrition. Orphanage girls were more likely to be malnourished than orphanage boys. Children who had been admitted to an orphanage for more than a year were less malnourished. In village orphans, there was no association between undernutrition and duration of stay in extended families. Age and education of guardians were not associated with the nutritional status of children. We conclude that young orphanage children are more likely to be undernourished and more stunted than village children. Older orphanage children seem to have better nutrition than village orphans. There was no significant difference in nutritional status between village orphans and non-orphans. PMID- 10715716 TI - Striking differences in the nasopharyngeal flora of healthy Angolan, Brazilian and Dutch children less than 5 years old. AB - Community-acquired pneumonia from enteric gram-negative bacilli is more common in developing than in industrialized countries. We investigated the nasopharyngeal flora in healthy children from Angola, Brazil and The Netherlands to see whether enteric gram-negative bacilli are more often part of the commensal flora in developing countries. Nasopharyngeal specimens were collected from children aged between 4 months and 5 years in day-care centres and immunization clinics. Children who had received antibiotics or were malnourished were excluded. Brazilian and Angolan children had a higher number of household members than Dutch children (5.5 and 7 vs 3.9 mean number of household members, respectively) (p < 0.0001). Enteric and non-fermentative gram-negative bacilli were much more prevalent in Brazilian (50%) and Angolan (57%) children than in Dutch (4%) children (p < 0.0001). By univariate analysis, carriage of enteric gram-negative bacilli was associated with the number of household members (r = 0.26; p < 0.001). The high carriage rate of enteric gram-negative bacilli in children from Angola and Brazil may explain why enteric gram-negative bacilli are a common cause of pneumonia in developing countries. PMID- 10715717 TI - Penetrating abdominal injuries in children in Nigeria. AB - This is a report of a retrospective study of 24 children managed for penetrating abdominal injury over 10 years, and it represents 34% of all abdominal injuries in children in that period. Falls onto sharp objects within and around the home were responsible for ten of the injuries, seven were injured by animal horns and four were sporting injuries. Violence and road traffic accidents were uncommon. Most patients (67%) had evisceration of omentum or intestine, and one of these was found at laparotomy to have a jejuno-jejunal intussusception. Seven children had injury to hollow viscera. There were three deaths, one each from overwhelming sepsis, tetanus and haemorrhage. PMID- 10715718 TI - Nephrotic syndrome associated with Toxocara canis infection. AB - This case report describes nephrotic syndrome in a 7-year-old boy coincident with Toxocara canis infection. This rare association was confirmed by elevated Toxocara-specific IgM titres. Treatment with corticosteroids resulted in remission of renal symptoms as well as abatement of the T. canis infection. The relationship between T. canis infection and glomerular disease is still unclear; nephrotic syndrome may be another manifestation of T. canis infection. PMID- 10715719 TI - Pyknodysostosis: a report of two siblings with unusual manifestations. AB - We report pyknodysostosis presenting as extramedullary haematopoiesis in one of two siblings and as obstructive airway disease in the other. Visceral manifestations are rare and have been reported in only two cases in the Indian literature. They have often been mistaken for osteopetrosis, haemolytic anaemia and other osteochondrodystrophies. The cases we report illustrate that, though the physical characteristics may be similar, it is the radiological features that are typical and help establish the diagnosis. PMID- 10715720 TI - Maternal nutritional status and dietary supplementation and the growth of suckling infants. PMID- 10715721 TI - Chronic subdural haematoma: an everyday problem for the neurosurgeon. PMID- 10715722 TI - Spinal intradural tumours: Part I--Extramedullary. AB - Sixty-six patients had surgery for an intramedullary nerve sheath tumour under the care of one surgical team in a 16-year period. Surgery concentrated on radical intra- and extradural excision combined if necessary with vertebral column reconstruction. Ninety procedures were used in 35 males and 30 females with an age range 12-81 years. Forty-five per cent were located in the cervical, 26% in the thoracic and 29% in the lumbosacral region. Eighteen patients had NF1 and two patients NF2. Sixty-five per cent were schwannomas, 27% were mixed histology and 6% malignant. In terms of functional outcome, 37 patients improved by one or more Frankel grades, three deteriorated by one Frankel grade and no one who presented with symptoms alone deteriorated. There were no operative deaths; no instrumentation failures and five patients developed a CSF leak. PMID- 10715723 TI - Spinal intradural tumours: Part II--Intramedullary. AB - The results of surgical management in 54 patients with intramedullary spinal cord tumours are presented. Cervical tumours were most frequent (25/54) followed by thoracic (16/54) and then lumbar (14/54). Ependymomas and astrocytomas were the most common tumour types. Total tumour removal was possible in just over half of the cases. Surgical complications included: two deaths, six patients with CSF leaks and one with wound infection. Postoperatively three patients had worsening of their motor deficit (unable to walk) and three patients had worsening of urinary sphincter function. Conversely, three patients who were unable to walk preoperatively were able to walk postoperatively, whilst four patients with sphincter disturbance showed improvement. Total tumour removal was not associated with increased risk of postoperative neurological deficit. Long-term follow up (2 18 years) was possible in 40 patients; 90% were still independently mobile. Our results compare favourably with other European studies and data from the North American units which have pioneered this surgery. PMID- 10715725 TI - Intraoperative evoked facial muscle responses and recovery process of the facial nerve in acoustic neuroma surgery. AB - The prognostic value of intraoperative evoked facial muscle responses (EFMR) was studied and correlated with the recovery process of the facial nerve during a follow-up period of 18 months. The patients were classified into four groups according to EFMR amplitudes, group A (150 microV or greater, 190.8, SD28.9 microV, n = 24), group B (100-149 microV, 125.7, SD14.3 microV, n = 14), group C (50-99 microV, 79.0, SD17.0 microV, n = 13) and group D (less than 50 microV, 22.1, SD13.3 microV, n = 15). Significant improved facial function appeared at 3 months after the operation in group A, at 6 months in group B, at 9 months in group C and at 12 months in group D. The early postoperative facial function and facial outcome of groups A and B were significantly better than those of groups C and D. Our data revealed that the intraoperative EFMR amplitudes have more prognostic value in predicting the recovery process of the nerve than functional outcome. PMID- 10715724 TI - Head injuries in four British neurosurgical centres. AB - An issue in the design of trials in traumatic brain injury is whether variation amongst centres in 'conventional' management could mask the impact of a powerful new pharmacological agent. We report the results of an observational study of 988 patients admitted to one of four British neurosurgical units between 1986 and 1988 within 3 days of a severe head injury. The centres fell into two pairs on the basis of the 'intensity' of management. In Edinburgh and Southampton, more frequent use of intracranial pressure monitoring, ventilation and osmotic diuretics was made than in Glasgow and Liverpool. The odds ratio for an independent outcome at 6 months in Edinburgh or Southampton, relative to Glasgow or Liverpool, controlling for case mix, was 1.43 (95% CI, 1.03-1.98, p = 0.033). Thus, there is weak evidence of an association between the approach to management and clinical outcome at 6 months. PMID- 10715726 TI - A prospective study of computer-aided design and manufacture of titanium plate for cranioplasty and its clinical outcome. AB - The use of computerized three dimensional imaging and automated milling of models to produce accurate titanium plates for the reconstruction of craniofacial defects is described. A total of 148 patients have had extensive calvarial defects repaired using this (computer aided design and manufacture) technique developed in our unit. Of these, 141 were repaired secondarily (delayed cranioplasty), whilst seven were repaired immediately following craniectomy (single stage cranioplasty). All cases were assessed for accuracy of fit, restoration of natural skull contour and aesthetics. Seventy-two patients were reviewed after 1 year to determine the effect on adverse preoperative symptoms. Of the plates 97% had an excellent or good intraoperative fit. The modal insertion time was only 15 minutes. Postoperatively 98% resulted in the restoration of natural skull shape and symmetry. After 1 year, 82% of patients had complete resolution or diminution in severity of the adverse symptoms. A staphylococcus infection necessitated the temporary removal of one plate. PMID- 10715728 TI - One-stage removal of a large dumb-bell-shaped cervical neurinoma without laminectomy or interbody fusion in a child. AB - A 12-year-old boy had a large dumb-bell-shaped cervical neurinoma originating at the C5 spinal root that was removed in a one-stage operation through the enlarged C4/5 intervertebral foramen. This technique required no laminectomy, discectomy or interbody fusion, which may frequently produce spinal deformity in children. PMID- 10715727 TI - A comparative study of the Spiegelberg compliance device with a manual volume injection method: a clinical evaluation in patients with hydrocephalus. AB - A new automated method of compliance measurement has been developed which may overcome some of the problems of the manual method. Measurement of craniospinal compliance in brain-injured patients offers the potential for early detection of raised intracranial pressure (ICP) before it rises to levels that may damage brain parenchyma. However, limitations of the existing manual volume pressure techniques have meant few centres routinely perform compliance testing. We report on the results of testing this new method against a manual volume pressure response method (VPR) in 10 patients with hydrocephalus. In this comparison study, 19 pairs of compliance measurements were obtained from 10 patients. The compliance values obtained ranged from 0.141 to 1.407 ml/mmHg. There was a good correlation between the two methods (r2 = 0.8508). The average bias in compliance between the two methods was 0.111 ml/mmHg (95% CL for the bias = 0.0438, 0.1788) with the new method reading higher compliance than the manual method. These results indicate that the new automatic method of compliance measurement correlates well with an independent and classical measurement of compliance, and defines the bias and limits of agreement by which the new method measures craniospinal compliance in patients with hydrocephalus. Further work is needed to validate this device over a wider compliance range, especially at the lower compliance range often found in head injured patients. Studies are also required to determine the normal range of compliance values in the patient populations who undergo ICP monitoring. Research into determining which patient populations may benefit from continuous compliance measurement is warranted. PMID- 10715729 TI - Factitious neurosurgical emergencies: report of five cases. AB - Patients with factitious symptoms occasionally present as neurosurgical emergencies. We have had six admissions over an 8-year period. They all presented at night, half of them during a weekend. Although an organic cause had to be excluded, they were all discharged within 24 h of admission without surgical intervention. PMID- 10715730 TI - Intracranial haemangioendothelioma mimicking a meningioma. AB - A 33-year-old man presented with a history of fits and on initial investigation was suspected of having a left frontal parafalcine meningioma. Initial surgical procedure to excise the lesion had to be abandoned owing to the extreme vascularity of the lesion. Histology revealed it to be a haemangioendothelioma. At a second operation the tumour was completely removed. The histology of this rather uncommon tumour is discussed and the literature is reviewed. PMID- 10715731 TI - Local anaesthesia for laminectomy surgery. AB - Neurosurgical patients presenting for laminectomy surgery may have premorbid pathology that either contraindicates general anaesthesia or at least represents a significant risk to the patient. We present a sample case from a series of ten patients in whom laminectomy surgery was performed under local anaesthesia. The mean duration of surgery was 98 minutes and the average dose of lignocaine used was 1.91 mg/kg and, therefore, within safe limits. One patient was converted to a general anaesthetic. We believe that local anaesthesia can offer a safe and satisfactory alternative, in patients who may otherwise be denied surgery. The additional advantage of awake neuro-monitoring, may also reduce the risk of inadvertant spinal cord injury. PMID- 10715732 TI - Spinal cord compression caused by adjacent adenocystic carcinoma of the skin. AB - Adenocystic carcinomas are malignant tumours that arise from the major accessory salivary glands. Cutaneous involvement can result from direct extension from a salivary gland neoplasm. Cutaneous adenocystic carcinomas remote from adjacent salivary tissue are rare. We present the case of an elderly patient with primary cutaneous adenoid cystic carcinoma causing spinal cord compression at the L1-L2 level. The patient was operated on and the tumour totally removed. No similar cases have been found in our review of the literature. PMID- 10715733 TI - Rapid spontaneous resolution of an acute extradural haematoma: case report. AB - A case of acute extradural haematoma with spontaneous resolution within 6 h of the head injury is presented. The literature is reviewed. PMID- 10715734 TI - Wandering intraspinal bullet. AB - A case of gun shot injury to the spine, with the bullet entering the thecal sac via the right side of the lower chest and wandering freely in the subarachnoid space, is reported. The patient was neurologically intact initially and developed radicular symptoms with foot drop and urinary retention on the third day after injury. The radiological findings and the problems faced at surgery are discussed, and the relevant literature of this uncommon condition is reviewed. PMID- 10715735 TI - Aneurysm of persistent primitive hypoglossal artery. AB - A patient with a ruptured intracranial aneurysm of the right persistent primitive hypoglossal artery is described. Clipping of the aneurysmal neck was achieved by a far lateral approach together with drilling of the jugular tubercle, and it was concluded that this approach should be followed for safe and complete clipping. PMID- 10715736 TI - Occipital condyle fracture with peripheral neurological deficit. AB - A 24-year-old woman sustained a type III Anderson and Montesano fracture in a road traffic accident. Acute respiratory stridor, multiple cranial nerve palsies and right upper limb neurological deficits associated with a C1 to T2 extradural haematoma were unique features of this case. The patient made a full and uncomplicated recovery with conservative management. PMID- 10715737 TI - Neurohypophyseal pilocytic astrocytoma invading the skull base. AB - We describe the clinical presentation, neuroradiological and histological findings of an unusual case of pilocytic astrocytoma of the neurohypophysis, and discuss the related surgical and prognostic issues of this neoplasm which invaded the skull base and the sphenoid sinus. Only four histologically proven cases of such a tumour have been reported in the English literature, and the pathological features and behaviour of this neoplasm still await definition. PMID- 10715738 TI - Promoting mental health: recent progress and problems in Australia. PMID- 10715739 TI - Neonatal and postneonatal mortality in Germany since unification. AB - BACKGROUND: After unification, the gap in infant mortality rates between the two parts of Germany widened until 1996 before converging. The reasons for these changes have not, so far, been apparent. OBJECTIVES: To investigate trends in neonatal and postneonatal mortality in the eastern (the new Lander) and western (the old Lander) part of Germany after unification in 1990 and to identify the scope for further improvement. DESIGN: Examination of trends in birth weights, birth weight specific neonatal mortality and cause specific postneonatal mortality in the two parts of Germany from 1990 to 1996 and 1997 by analysing routinely available vital statistics data. RESULTS: In both parts of Germany, neonatal mortality fell considerably, by 33 per cent in the east and 17 per cent in the west, from 4.5 and 3.5 per thousand live births in 1990 to 3.0 and 2.9 in 1997, respectively. This was attributable to an improvement in survival of infants at all birth weights but especially among those with very low birth weights, accounting for an estimated 83 to 85 per cent of the overall improvement. The birth weight distribution showed a slight worsening in the new and the old Lander with an increase in the proportion of those under 1500 g and, in the east, a 24 per cent increase in the proportion of high birth-weight infants of 4000 and more grams. Trends in postneonatal mortality revealed a worsening of about 32 per cent in the east from 1990 to 1991 followed by a decline of over 50 per cent up to 1997, leading to comparable mortality rates of 1.8 per thousand live births in the east and 2.0 in the west. While both parts experienced a decrease of 40 to 48 per cent in deaths from all diseases, the decline in deaths because of accidents and injuries was markedly higher in the new Lander although they are still exceeding the western rate by 3.7 per 100,000 live births in 1997. CONCLUSIONS: Since unification, the two parts of Germany underwent a complex process that has led finally to convergence of parameters of infant health that are most likely to have been because of improvements in the quality of perinatal care. To improve infant mortality in Germany, policy measures should focus on preventive rather than curative measures as the proportion of very low birthweight babies is increasing in both parts of Germany. PMID- 10715740 TI - Own education, current conditions, parental material circumstances, and risk of myocardial infarction in a former communist country. AB - OBJECTIVE: To study the association between own education, adult and parental circumstances and the risk of myocardial infarction in a former communist country. DESIGN: Population based case-control study. SETTING: General population of five districts of the Czech Republic in the age group 25-64 years. PARTICIPANTS: Random sample of population (938 men and 1048 women, response rate 77%) served as controls to 282 male and 80 female cases of non-fatal first myocardial infarctions. MAIN OUTCOME MEASURES: Myocardial infarction was defined by the WHO MONICA criteria based on ECG, enzymes and symptoms. The following socioeconomic indicators were studied: own education, crowded housing conditions (more than one person per room), car ownership, and education and occupation of mother and father. RESULTS: There was a weak correlation between education and car ownership, and a strong association between own education and parental education and occupation. Crowding was not related to other socioeconomic factors. The risk of myocardial infarction was inversely related to education, and was unrelated to material conditions and parental education and occupation. The age-sex-district adjusted odds ratios for apprenticeship, secondary, and university education, compared with primary education, were 0.87, 0.74 and 0.46, respectively (p for trend 0.009); odds ratios for car ownership and crowding were 1.01 (95% confidence intervals 0.77, 1.34) and 0.92 (0.76, 1.12), respectively. Further adjustment for parental circumstances and adult height did not change these estimates but adjustment for coronary risk factors reduced the gradient. Increased height seemed, anomalously, to confer a small increased risk. CONCLUSIONS: In this population, the social gradient in non-fatal myocardial infarction is only apparent for own education. Materialist explanations for this gradient seem unlikely but behaviours seem responsible for a part of the gradient. PMID- 10715741 TI - Height and risk of death among men and women: aetiological implications of associations with cardiorespiratory disease and cancer mortality. AB - OBJECTIVES: Height is inversely associated with cardiovascular disease mortality risk and has shown variable associations with cancer incidence and mortality. The interpretation of findings from previous studies has been constrained by data limitations. Associations between height and specific causes of death were investigated in a large general population cohort of men and women from the West of Scotland. DESIGN: Prospective observational study. SETTING: Renfrew and Paisley, in the West of Scotland. SUBJECTS: 7052 men and 8354 women aged 45-64 were recruited into a study in Renfrew and Paisley, in the West of Scotland, between 1972 and 1976. Detailed assessments of cardiovascular disease risk factors, morbidity and socioeconomic circumstances were made at baseline. MAIN OUTCOME MEASURES: Deaths during 20 years of follow up classified into specific causes. RESULTS: Over the follow up period 3347 men and 2638 women died. Height is inversely associated with all cause, coronary heart disease, stroke, and respiratory disease mortality among men and women. Adjustment for socioeconomic position and cardiovascular risk factors had little influence on these associations. Height is strongly associated with forced expiratory volume in one second (FEV1) and adjustment for FEV1 considerably attenuated the association between height and cardiorespiratory mortality. Smoking related cancer mortality is not associated with height. The risk of deaths from cancer unrelated to smoking tended to increase with height, particularly for haematopoietic, colorectal and prostate cancers. Stomach cancer mortality was inversely associated with height. Adjustment for socioeconomic position had little influence on these associations. CONCLUSION: Height serves partly as an indicator of socioeconomic circumstances and nutritional status in childhood and this may underlie the inverse associations between height and adulthood cardiorespiratory mortality. Much of the association between height and cardiorespiratory mortality was accounted for by lung function, which is also partly determined by exposures acting in childhood. The inverse association between height and stomach cancer mortality probably reflects Helicobacter pylori infection in childhood resulting in--or being associated with--shorter height. The positive associations between height and several cancers unrelated to smoking could reflect the influence of calorie intake during childhood on the risk of these cancers. PMID- 10715742 TI - Incidence of myocardial infarction in women. A cohort study of risk factors and modifiers of effect. AB - STUDY OBJECTIVE: To assess whether the increased incidence of myocardial infarction and death associated with smoking, hypertension, hyperlipidaemia and diabetes varies significantly between groups defined in terms of occupation, education and marital status. SETTING: Malmo, Sweden. PARTICIPANTS: 9351 women, aged 28-55, with a mean follow up of 10.7 years. MAIN RESULTS: Smoking, hypertension (> or = 160/95 mm Hg or treatment), hyperlipidaemia (cholesterol > or = 6.5 mmol/l or triglycerides > or = 2.3 mmol/l), diabetes, low occupation and education levels were significantly more common among women who experienced a fatal or nonfatal myocardial infarction during the follow up (n = 104) than in other women (n = 9247). Exposure to smoking, hypertension and hyperlipidaemia showed substantial differences between groups defined in terms of education, occupation and marital status. The association between low occupation and myocardial infarction remained statistically significant after adjustments for several potential confounders (RR = 2.6, 95% CI 1.1, 6.0). Single women had similarly higher adjusted mortality rates than married women (RR = 1.4, 95% CI 1.1, 1.8). When other major risk factors were taken into account, the relative risk for mortality and myocardial infarction associated with smoking was 2.6 (95% CI 2.0, 3.4) and 7.8 (95% CI 4.4, 13.9), respectively. CONCLUSION: In this urban female population, short education and low occupation level were both associated with an increased prevalence of smoking, hypertension, hyperlipidaemia and diabetes. Low occupation level increases the rate of cardiac events caused by exposure to these four risk factors. PMID- 10715743 TI - Socioeconomic status and trends in risk factors for cardiovascular diseases in the Danish MONICA population, 1982-1992. AB - STUDY OBJECTIVE: The decline in cardiovascular mortality in Denmark during the 1980s has been greatest in the highest socioeconomic groups of the population. This study examines whether the increased social inequality in cardiovascular mortality has been accompanied by a different trend in cardiovascular risk factors in different educational groups. DESIGN: Data from three cross sectional WHO MONICA surveys conducted in 1982-84, 1987, and 1991-92, were analysed to estimate trends in biological (weight, height, body mass index, blood pressure, and serum lipids) and behavioural (smoking, physical activity during leisure, and eating habits) risk factors in relation to educational status. SETTING: County of Copenhagen, Denmark. PARTICIPANTS: 6695 Danish men and women of ages 30, 40, 50, and 60 years. MAIN RESULTS: The prevalence of smoking and heavy smoking decreased during the study but only in the most educated groups. In fact, the prevalence of heavy smoking increased in the least educated women. There was no significant interaction for the remaining biological and behavioural risk factors between time of examination and educational level, indicating that the trend was the same in the different educational groups. However, a summary index based on seven cardiovascular risk factors improved, and this development was only seen in the most educated men and women. CONCLUSION: The difference between educational groups in prevalence of smoking increased during the 1980s, and this accounted for widening of an existing social difference in the total cardiovascular risk. PMID- 10715744 TI - Psychosocial aetiology of chronic disease: a pragmatic approach to the assessment of lifetime affective morbidity in an EPIC component study. AB - OBJECTIVES: The Health and Life Experiences Questionnaire (HLEQ) was developed for use in a prospective cohort study of 25,000 men and women living in Norfolk and forms a component study of the European Prospective Investigation into Cancer and Nutrition (EPIC). The HLEQ includes an assessment of mood status over the life course allowing a limited capacity for the imposition of diagnostic criteria to enable eventual evaluation of mental health status for chronic disease outcomes. This paper reports estimates of HLEQ Major Depressive Disorder (MDD) prevalence and compares them with those obtained through interviewer-based methods. In addition evidence for the impact of recall, clustering or cohort effects on these estimates are examined. PARTICIPANTS: 3491 eligible respondents to EPIC in Norfolk, aged 45-74 years, recruited from the first five general practices who completed the HLEQ. MAIN RESULTS: MDD prevalence estimates were found to be closely comparable to those obtained recently (by interview) in the UK and to those lifetime MDD rates determined through international studies. Risk of MDD onset was found to vary with age as expected from earlier studies using interviewer-based assessments. Limited evidence was found to show that the distribution of first onset MDD episodes were compressed during the immediate pre assessment period. Results were also consistent with previous evidence demonstrating the raised risk of MDD among women and of the decline in gender differences with advancing age. CONCLUSIONS: These results suggest that estimates of putative MDD diagnostic status, derived through the HLEQ, and of associated demographic risk are similar to those derived by more intensive and costly assessment methods. Implications for the future study of MDD both as an outcome and as a risk factor for chronic disease are discussed. PMID- 10715745 TI - The predictive value of self assessed general, physical, and mental health on functional decline and mortality in older adults. AB - OBJECTIVE: To examine the extent to which older people's self assessments of general health, physical health, and mental health predict functional decline and mortality. DESIGN: The study uses population-based secondary data from the US Longitudinal Study of Aging (LSOA). PARTICIPANTS: A total of 7527 persons aged 70 years or above living in the community. METHODS: Eight different measures on self reported general, physical, and mental health were used. Change in functional status was measured using a composite index of ADLs and IADLs over a period of six years. Duration of survival was calculated over a period of seven years. Adjusting for age and gender, multiple logistic regression was used in analysing functional decline, and Cox proportional hazard model, for mortality. Then all of the self assessed health measures were incorporated into the final model- controlling for baseline sociodemographic characteristics, functional status, disease/conditions, and use of health and social services--to assess the independent contribution of each measure in predicting future health outcomes. MAIN RESULTS: Overall, older people's self assessed general, physical, and mental health were predictive of functional decline and mortality. In multivariate analyses, older people who assessed their global health, self care ability, and physical activity less favourably were more likely to experience poor health outcomes. Gender disparity, however, was observed with poor global health affecting functional decline in men only. Self care ability was predictive of functioning in women only, whereas it was predictive of mortality in men only. CONCLUSIONS: Self assessed global health, as well as, specific dimensions of health act as significant, independent predictors of functioning and mortality in a community dwelling older people. PMID- 10715746 TI - Overall mortality among patients surviving an episode of peptic ulcer bleeding. AB - STUDY OBJECTIVE: The authors investigated whether patients who have survived an acute episode of peptic ulcer bleeding (PUB) have an excess long term all cause mortality compared with the general population free of PUB. DESIGN: Follow up study of previously identified cohort of patients with a PUB episode and a general population cohort. SETTING: The source population included all people aged 30 to 89 years, registered with general practitioners in the United Kingdom. PATIENTS: All patients alive one month after the PUB episode constituted the cohort of PUB patients (n = 978). A control group of 5000 people was randomly sampled from the source population. The same eligibility criteria as for patients with PUB were applied to the control series. Also, controls had to be free of PUB before start date. MAIN RESULTS: Relative risk of mortality among PUB patients was 2.1, 95% CI: 1.7, 2.6) compared with the general population. This increased mortality risk occurred mainly in the patients less than 60 years old. No difference was observed between men and women. The excess mortality was not only circumscribed to deaths attributable to recurrent gastrointestinal bleed, but also cardiovascular, cancer and other causes. CONCLUSIONS: People who have survived an acute episode of PUB have a reduced long term survival compared with the general population. This reduction was stronger among middle age patients than in the elderly. PMID- 10715747 TI - Continuing the debate on the philosophy of modern public health: social quality as a point of reference. PMID- 10715748 TI - Accuracy of the estimated prevalence of obesity from self reported height and weight in an adult Scottish population. AB - STUDY OBJECTIVE: To determine whether self reported heights and weights from Scottish adults can provide an accurate assessment of obesity prevalence in the population. DESIGN: Standardised clinic measurements of weight and height were compared against self reported values on a postal questionnaire in the fourth Scottish MONICA cross sectional study. SETTING: A sex and five year age band stratified random population sample drawn from general practitioner registers in north Glasgow in 1995. Response rate 63% for men and 62% for women. PARTICIPANTS: A total of 865 men and 971 women aged between 25 and 64 years. RESULTS: Men and women under-reported their weight by a mean (SD) of 0.63 (3.45) kg and 0.95 (2.64) kg respectively, and their height by a mean (SD) of 1.3 (2.50) cm and 1.7 (2.37) cm respectively. Estimated body mass index, BMI (kg/m2) varied from true (measured) BMI by +0.19 (1.40) for men and by +0.17 (1.34) for women. The only age/sex group in which BMI was under-estimated from self reports (mean 0.2) was the 55-64 year old women. Prediction equations that explained 90% (men) and 88% (women) of the difference between self reported and measured height included age and self reported weight. The equivalent prediction equations for weight explained 93% of the difference between self reported and measured weight for men and included smoking and diabetic status, while for women 96% of the variance was explained with no further variables being significant. Sensitivity and specificity for determining clinical obesity (BMI > or = 30) were 83% and 96% respectively for men, and 89% and 97% for women. CONCLUSIONS: This Scottish population was unique in the under-reporting of height as well as weight, which resulted in BMI estimates with low error. These data suggest that self reported weights and heights would be satisfactory for the monitoring of obesity prevalence in Scotland. PMID- 10715749 TI - Evaluation of outreach clinics held by specialists in general practice in England. AB - OBJECTIVES: To measure the processes of care, health benefits and costs of outreach clinics held by hospital specialists in primary care settings. DESIGN: The study was designed as a case-referent (comparative) study in which the features of 19 outreach clinics (cases) were compared with matched outpatient clinics (controls). The measuring instruments were self administered questionnaires. Patients were followed up at six months to reassess health status. The specialties included in the study were cardiology, ENT, general medicine, general surgery, gynaecology and rheumatology. SETTING: Specialist outreach clinics in general practice in England, with matched outpatient clinic controls. SUBJECTS: Consecutive patient attenders in the outreach and outpatient clinics, their specialists, the outreach patients' general practitioners, practice managers and trust accountants. Patients' response rate at baseline: 78% (1420). MAIN OUTCOME MEASURES: Patient satisfaction, doctors' attitudes, processes and health outcomes, costs. RESULTS: Outreach patients were more satisfied with the processes of their care than outpatients, their access to specialist care was better than that for outpatients and they were more likely to be discharged. Doctors reported that the main advantages of the outreach clinic were improved patient access to specialists and convenience for patients, in comparison with outpatients, and most GPs and specialists felt the outreach clinic was "worthwhile". At six month follow up, the health status of the outreach sample had significantly improved more than that of the outpatients on all eight sub-scales of the HSQ-12, but this was probably because of their better starting point at baseline. The impact of outreach on health outcomes was small. The NHS costs of outreach were significantly higher than outpatients. An increase in outreach clinic size would reduce cost per patient, but would lead to the loss of most of the clinics' benefits. CONCLUSIONS: While the process of care was of higher quality in outreach than in outpatients, and the efficiency of care was also greater in the latter, the effect on patients' health outcomes was small. Responsiveness to patients' views and preferences is an essential component of good quality service provision. However, the greater cost of outreach raises the issue of whether improvements in the quality and efficiency of health care, without a substantial impact on health outcomes, is money well spent in a publicly funded health service. On the other hand, the real costs of outreach in comparison with outpatients clinics can probably only be truly estimated in a longitudinal study with a resource based costing model derived from documented patient attendances and treatment costs over time in relation to longer term outcome (for example, at a two year end point). PMID- 10715750 TI - Absence of economic barriers does not reduce disparities in the access to renal transplantation: a population based study in a region of central Italy. Dialysis Register of Lazio Region. PMID- 10715751 TI - Eight aphorisms for palliative care. PMID- 10715752 TI - Communication and awareness about dying in the 1990s. AB - Since the 1960s communication and awareness about dying in modern western societies have been topics for debate, with a considerable amount of literature on the need for open communication and the strategies which can be used by health professionals to improve their communication with patients facing a terminal prognosis. Despite the difficulties of comparing studies using different methodologies and the additional problems of ascertaining patients' knowledge and awareness about their impending deaths, the trend is clear. In advanced industrial societies there is increasing evidence that doctors have shifted from a policy of 'withholding' to a policy of 'revealing' to the patient his or her terminal prognosis. This change in medical practice is further supported by other data which show an increase in the percentage of those patients who were aware they were dying from chronic diseases, especially cancer. However, despite the perceived trend towards open disclosure of the patient's terminal illness, recent studies have suggested that in their daily encounters with dying patients health workers employ 'conditional' rather than 'full open disclosure'. Such moderating strategies in discussing the patient's prognosis may be employed despite open awareness of a patient's prognosis. This paper examines this apparent paradox by analysing the complex tensions and conflicts of such communication through a discussion of existing literature on modes of communication and patient awareness. PMID- 10715753 TI - How do terminally ill patients at home take their medication? AB - Compliance with prescribed medication was assessed in 111 terminally ill patients referred to a community palliative care team using semistructured interviews and pill counting. One-hundred-and-six patients were prescribed a total of 597 drugs; of these patients, 64 (60%) were noncompliant. Thirty-five patients (33%) took less medication than prescribed, usually due to experiencing, or anxieties about, adverse events; the commonest drugs involved were analgesics. Seventeen patients (16%) took additional medication, usually purchased over the counter in response to inadequate symptom control or to adverse events from other drugs; the most common preparations were vitamins and analgesics. Twelve patients (11%) both took less medication than prescribed and also purchased medication over the counter. Most patients (90%) had two or more prescribers; patients who saw their general practitioners as their main prescriber were more likely to adhere to their prescribed medication. Patients who omitted and/or reduced their medication were more likely to see the hospital as their main prescriber. Drugs prescribed four times daily were most likely to be omitted and/or reduced. PMID- 10715754 TI - Specialist palliative care and patients with noncancer diagnoses: the experience of a service. AB - This retrospective review was undertaken to identify the pattern of noncancer referrals to a specialist palliative care service, comprising a teaching hospital support team, home care, outpatients and inpatient hospice, over a 1-year period. Of 287 hospital ward referrals, 83 patients had a noncancer diagnosis (29%); they were referred predominantly for symptom control (92%), particularly of pain (84%). Of 130 outpatient referrals, 30 had a noncancer diagnosis (23%) and were also referred mainly for the management of pain (85%). Of 196 home care referrals, 18 had a noncancer diagnosis (9%); they tended to be referred for multiprofessional care of endstage disease. Of 421 hospice inpatient admissions, 17 were for patients with a noncancer diagnosis (4%) and were predominantly for respite care. These admissions accounted for 2% of occupied bed days. It is concluded that specialist palliative care skills are perceived to be transferable to patients with noncancer diagnoses. Resource implications focus on hospital and outpatient services, where shared care with medical teams is usual practice. Defining management goals at the outset is particularly important. PMID- 10715755 TI - An annotated bibliography of the publications of Cicely Saunders--2: 1968-77. PMID- 10715756 TI - Hypokalaemic paralysis. AB - Hypokalaemic paralysis is a relatively uncommon but potentially life-threatening clinical syndrome. If recognised and treated appropriately, patients recover without any clinical sequellae. The syndrome of hypokalaemic paralysis represents a heterogeneous group of disorders characterised clinically by hypokalaemia and acute systemic weakness. Most cases are due to familial or primary hypokalaemic periodic paralysis; sporadic cases are associated with numerous other conditions including barium poisoning, hyperthyroidism, renal disorders, certain endocrinopathies and gastrointestinal potassium losses. The age of onset, race, family history, medications, and underlying disease states can help in identifying the cause of hypokalaemic paralysis. Initial therapy of the patient with hypokalaemic paralysis includes potassium replacement and search for underlying aetiology. Further management depends on the aetiology of hypokalaemia, severity of symptoms, and duration of disease. This review presents the differential diagnosis for hypokalaemic paralysis and discusses management of the syndrome. PMID- 10715757 TI - Transmission of Helicobacter pylori. AB - Helicobacter pylori is found predominantly in human gastric mucosa. Transfer of the bacterium remains an open topic, but it is likely that infection is usually acquired at a young age, particularly where lower socio-economic conditions prevail. Transmission via an external source such as water supply is a possibility but, in general, infection is probably passed from person to person. Arguments for and against faecal-oral, oral-oral and gastric-oral transmission have been presented, but the dominance of one of these routes is still to be determined. PMID- 10715758 TI - Non-organic visual loss. AB - Visual complaints without a physical basis are not uncommon presentations to the general physician, the neurologist, or the ophthalmologist. These alleged visual disturbances may be psychogenic or feigned. The diagnosis is made when all possible contributory pathology of the visual system is excluded, and reassurance remains the cornerstone of management. PMID- 10715759 TI - Targeting pneumococcal vaccination to high-risk groups: a feasibility study in one general practice. AB - The Department of Health recommends pneumococcal vaccination opportunistically or when immunising against influenza. This was a study in one general practice to assess the feasibility of targeting patients for pneumococcal vaccination in primary care. We also examined the rate of uptake of pneumococcal vaccine in identified risk groups after one year of a pneumococcal vaccination programme. A self-administered questionnaire was given to patients attending for influenza vaccine between September and December 1996. A total of 551/747 (73.8%) patients returned completed questionnaires. Few patients receiving influenza vaccination (133/509, 26%) were aware of pneumococcal vaccine. Only 55/108 (51%) of those given influenza vaccination were in a clinical risk group for pneumococcal vaccine. Attitudes towards vaccination were more positive and intention to take up pneumococcal vaccination significantly greater in high-risk patients compared to those who were not in a risk group. A targeted vaccination campaign directed at high-risk patients, both opportunistically and those attending for influenza vaccination over one year, resulted in the following proportions of patients in at-risk groups being vaccinated: coronary disease 144/312 (46%), diabetes 79/132 (60%), splenectomy 2/2 (100%), chronic obstructive airways disease and asthma 135/700 (19%), and chronic renal failure 5/9 (56%). Most doses of pneumococcal vaccine (336/463; 73%) were delivered to patients in high-risk groups. We conclude that a well-organised pneumococcal vaccination campaign can improve coverage of at-risk patients in general practice. Programmes to increase patient awareness of the vaccine, improved availability of vaccine, and practice guidelines, would help to target the vaccine to at-risk patients. Patients with chronic lung disease and asthma were particularly difficult to define and target in this study. A review of the UK guidelines, aligning those for pneumococcal and influenza vaccination and including patients over 65 years, would improve the logistics of vaccine delivery. PMID- 10715760 TI - Orthostatic haemodynamic responses in acute stroke. AB - Little is known about orthostatic blood pressure regulation in acute stroke. We determined postural haemodynamic responses in 40 patients with acute stroke (mild or moderate severity) and 40 non-stroke control in-patients, at two days ('Day 1') and one week ('Week 1') post-admission. Following a 10-minute supine rest and baseline readings, subjects sat up and blood pressure and heart rate were taken for 5 minutes. The procedure was repeated with subjects moving from supine to the standing posture. Haemodynamic changes from supine data were analysed. On standing up, the control group had a transient significant fall in mean arterial blood pressure on Day 1 but not Week 1. No significant changes were seen on either day when sitting up. In contrast to controls, the stroke group showed increases in mean arterial blood pressure on moving from supine to the sitting and standing positions on both days. Persistent postural hypotension defined as > or = 20 mmHg systolic fall occurred in < 10% of either of the study groups on both days. Sitting and standing heart rates in both groups were significantly faster than supine heart rate on both days. The orthostatic blood pressure elevation is consistent with sympathetic nervous system overactivity which has been reported in acute stroke. Upright positioning as part of early rehabilitation and mobilisation following mild-to-moderate stroke would, therefore, not predispose to detrimental postural reductions in blood pressure. PMID- 10715761 TI - ACE inhibitors and heart failure in hospital: any difference between cardiologists and general physicians? AB - Cardiologists and generalists have been reported to diverge in terms of their self-reported use of angiotensin-converting enzyme (ACE) inhibitors, but information on their actual use of ACE inhibitors has been lacking. In order to assess ACE inhibitor use in patients with heart failure in a teaching hospital and any differences between specialties we studied all patients in the Western Infirmary of Glasgow between 1 April and 1 October 1996 with an echocardiogram showing moderate or severe left ventricular systolic dysfunction (n = 236). We found that most patients were on an ACE inhibitor (66%), 12% had been tried but found to be intolerant, 10% had not been tried because of a contraindication, but 12% had not been tried despite no contraindication. Of those on treatment, 58% were on a dose used in a major survival study (38% of all patients). Most patients were treated by a cardiologist (64%). Of these, more were on an ACE inhibitor (77% vs 53%, p < 0.01), fewer had been tried but found intolerant (11% vs 18%), and fewer had never been tried (11% vs 29%, p < 0.01), irrespective of whether they had a contraindication (5% vs 18%, p < 0.01) or not (6% vs 12%). More were on a dose used in a major survival study (48% vs 31%, p < 0.05). We conclude that, despite improvements over time, ACE inhibitors are still under used, sometimes without good reason. There are also differences in the use of ACE inhibitors between cardiologists and generalists which may affect outcome, and could affect resource utilisation. PMID- 10715762 TI - The use of hypnosis in gastroscopy: a comparison with intravenous sedation. AB - A total of 124 subjects who were undergoing routine endoscopy were randomly assigned to one of three groups. All three groups received lignocaine throat spray. The first group additionally received midazolam, the second received hypnosis, whilst the third only received lignocaine throat spray. Although hypnotized patients were deemed by an independent observer to be less agitated than the other two groups (p < 0.03), they reported the gastroscopy to be significantly more uncomfortable (p < 0.042) and scored higher in their memory for the procedure (p < 0.001). They also took slightly longer to induce than the midazolam group. The midazolam group on the other hand rated the procedure as significantly more comfortable although paradoxically were seen by an independent observer as being more agitated. They were also significantly more amnesic. The endoscopist encountered more procedural difficulties with this group but this did not reach levels of significance. Hypnosis was not shown to be an effective alternative to intravenous sedation in gastroscopy. PMID- 10715763 TI - Mature bone metaplasia in abdominal wall scar. AB - A 58-year-old man who had had three laparotomies for gastric surgery, developed a painful mass in the abdominal wall scar. Radiology confirmed bone formation in the scar. The bone was excised and the wound repaired. Histology confirmed metaplastic mature bone formation. This case draws the attention to the clinical condition of bone formation in midline scars. Clinically, it should be differentiated from scar recurrence following surgery for abdominal malignancy. PMID- 10715764 TI - High-altitude illness induced by tooth root infection. AB - High-altitude illness may occur after recent pulmonary infection, but high altitude illness after root canal therapy has not been described previously. A 44 year-old man is presented who skied to a 3333 m high peak in the Eastern Alps one day after he had undergone root canal therapy because of a tooth root infection. After 4 hours above 3000 m severe symptoms of high-altitude illness, including pulmonary oedema, developed. His condition improved after immediate descent. The next day he presented with local and general signs of infection which were successfully treated with gingival incisions and antibiotics. In conclusion, acute tooth root infection and root canal therapy may induce high-altitude illness at an altitude just above 3000 m. PMID- 10715765 TI - Gynaecomastia as a presenting feature of thyrotoxicosis. AB - The association between gynaecomastia and thyrotoxicosis is well recognised. However, the reported frequency of the association is variable, partly depending upon the defining criteria used. Here we report two patients with thyrotoxicosis in whom gynaecomastia was the presenting feature. Both patients had other contributing factors, which are assumed to have predisposed to gynaecomastia. In both patients, the gynaecomastia resolved with successful treatment of the thyrotoxicosis. When gynaecomastia is a presenting, or prominent feature of thyrotoxicosis, the possibility of additional underlying pathology should be considered. PMID- 10715766 TI - Lumbar hernia: a rare cause of large bowel obstruction. AB - We describe a 70-year-old woman presenting with large bowel obstruction secondary to incarceration of the mid descending colon within a lumbar hernia. This was diagnosed on barium enema and successfully treated surgically. PMID- 10715767 TI - Warfarin-induced skin necrosis. AB - Skin necrosis is a rare but serious side-effect of treatment with warfarin. At particular risk are those with various thrombophilic abnormalities, especially when warfarinization is undertaken rapidly with large loading doses of warfarin. With the increasing number of patients anticoagulated as out-patients for thromboprophylaxis, we are concerned that the incidence of skin necrosis may increase. If skin necrosis does occur, prompt remedial action may be of benefit in preventing permanent tissue damage. PMID- 10715768 TI - Calcium-channel blockade can mask the diagnosis of Conn's syndrome. AB - A 30-year-old woman presented with hypertension and hypokalaemia, and was found to have primary aldosteronism due to a Conn's adenoma, whose removal cured the hypertension. Before surgery, the characteristic biochemical changes which enabled the diagnosis were completely masked by administration of a calcium channel blocker, amlodipine. It is likely that widespread use of this class of drugs contributes to under-diagnosis of Conn's syndrome as a curable cause of hypertension. PMID- 10715769 TI - An unusual rash. PMID- 10715770 TI - Large subcutaneous nodules in lower limbs following thyroiditis. PMID- 10715771 TI - Gestational pancreatitis complicating uncontrolled diabetes mellitus. PMID- 10715772 TI - A young woman with fluctuating hypo- and hyperthyroidism. PMID- 10715773 TI - A newborn with watery diarrhoea. PMID- 10715774 TI - An unusual cause of leg ulceration. PMID- 10715775 TI - A new diabetic patient with an abdominal mass. PMID- 10715776 TI - Fever and dyspnoea in a young man with a rash. PMID- 10715777 TI - Ion-selective electrodes. PMID- 10715778 TI - A young man with repeated thromboses. PMID- 10715779 TI - Communication skills. PMID- 10715780 TI - Lord Todd 1907-1997. PMID- 10715781 TI - Melvin Calvin (1911-1997). PMID- 10715782 TI - A fresh understanding of the stereochemical behavior of glycosylases: structural distinction of "inverting" (2-MCO-type) versus "retaining" (1-MCO-type) enzymes. PMID- 10715783 TI - [Expression of TGFb1 and its mRNA in liver tissues of patients with chronic viral hepatitis]. AB - OBJECTIVE: To investigate the relationship between expression of transforming growth factor beta 1(TGF beta 1), TGF beta 1 mRNA in liver tissue of patients with chronic viral hepatitis and degree of liver fibrosis. METHODS: TGF beta 1 protein was detected in 45 patients by immunohistochemistry and TGF beta 1 mRNA was detected in 21 of the 45 cases by in situ hybridization. RESULTS: TGF beta 1 protein was 88.89%(40 of 45) positive in their liver tissues and was mainly distributed in interstitial cells, portal areas and the areas where fibrosis actively occurred, and the expression of TGF beta 1 protein was positively correlated with degree of liver fibrosis(r = 0.73, P < 0.05). TGF beta 1 mRNA was 80.95%(19 of 21)positive in their liver tissues. Positive stains were not only detected in sinusoidal cells and mononuclear cells, but also in some parenchymal cells. CONCLUSIONS: The expression of TGF beta 1 correlated closely with liver fibrosis of viral hepatitis. Not only interstitial cells, but also parenchymal cells in liver might participate in the fibrogenisis by producing TGF beta 1. PMID- 10715784 TI - [The effect of serum TGFb1 of patients with chronic hepatitis B in liver fibrosis formation]. AB - OBJECTIVES: To observe the relation between serum transforming growth factor beta 1(TGF beta 1) and level of PCIII, LN, HA, proliferation of hepatic fibrotic tissue. METHODS: Serum TGF beta 1 was detected by ELISA in 58 patients with chronic hepatitis B(CHB) and 18 patients with liver cirrhosis(LC). 20 healthy persons served as normal control(NC). Liver puncture was performed in 29 patients with CHB. Diagnosis of pathological histology and quantitative analysis of collagenous and reticular fibers were made. RESULTS: (1)Serum TGF beta 1 levels in CHB and LC were notably higher than in NC (P < 0.01) and increased successively in mild, moderate and severe degrees of CHB and LC(P < 0.01/0.05). (2)Serum TGF beta 1 was correlated with serum levels of PCIII, LN, HA(P < 0.01/0.05)positively. The quantity of hepatic collagenous and reticular fibers went up in mild, moderate and severe degrees of CHB(P < 0.01/0.05). Meanwhile serum TGF beta 1 rose at the same degree. CONCLUSION: Serum TGF beta 1 levels increase in CHB and LC. With hepatic pathological change worsening, serum TGF beta 1 and the quantity of hepatic collagenous and reticular fibers increased to the same degree. This suggests that TGF beta 1 is a crucial factor in accelerating liver fibrosis and plays a decisive role in liver fibrosis. PMID- 10715785 TI - [Relationship between serum fibrosis markers and fibrosis quantitative analysis of liver tissue]. AB - OBJECTIVE: To study relation of serum fibrosis markers with quantitative analysis of liver tissue. METHODS: We studied the area of reticular fibrin, collagen fibrin and elasticity fibrin in 77 patients with chronic liver disease using color image analysis system. RESULTS: From S0 to S4, the area of three kinds of liver fibrosis were all escalate, especially collagen fibrosis. Prominent differences existed between S3 and S2, S2 and S1, S1 and S0. There was a good correlation between the serum fibrosis markers and area of collagen fibrosis of liver tissue, r = 0.70775(HA), 0.59402(CIV), 0.52593(LN), 0.52198 (hPCIII). CONCLUSIONS: Serum fibrosis markers have affirmative value to help diagnose liver fibrosis. PMID- 10715786 TI - [Study on esophageal motility and its effect by EVL in patients with esophageal varices]. AB - OBJECTIVE: To investigate the esophageal motility and its effect by EVL in patients with cirrhotic esophageal varices. METHODS: LESP, LESR, PA, PD, PV and 24 h pH-monitoring were measured by PC Polygraf HR system in subjects of control and cases of esophageal varices before and after EVL. RESULTS: In varices group, PA, PD, and PV were significantly abnormal in comparison with those in control group (P < 0.001, 0.01, 0.01). There were pathologic acid-reflux in patients with esophageal varices. After EVL, PA, PV and acid-reflux were significantly recovered. CONCLUSION: Patients with esophageal varices suffer from the significant abnormal esophageal motility and acid-reflux. All of the abnormal changes can be recovered by EVL treatment. PMID- 10715787 TI - [HBV transmission from father to foetus and HBV DNA in tissues outside the liver]. AB - OBJECTIVE: To study the possibility of HBV Transmission from father to foetus and HBV DNA in tissues outside liver. METHODS: Paired sera were from 8 HBV man carriers whose wives were negative for HBVM and 8 foetuses who were infected with HBV in the womb. S gene nt 451-660 nucleotide, C gene nt 2,022-2,321 nucleotide were directly sequenced. RESULTS: The homology of HBV sequence between father and foetus was very high. The mutations of 491, 494, 530, 546 and 581 nucleotide in the S gene caused 113, 114, 126, 131 and 143 amino acid substitution. HBV DNA can be detected in the tissues outside liver of foetus. CONCLUSION: HBV transmission from father to foetus may be present. HBV DNA in tissues outside liver of foetus can be detected. PMID- 10715788 TI - [Long-term evolution of C-terminal of HCV NS3 protein containing the predicted CTL epitopes in two HCV patients]. AB - OBJECTIVE: To investigate the Long-term evolution of NS3 C-terminal protein containing the predicted CTL epitopes. METHODS: The HLA typing was determined serologically in two chronic HCV patients. Based on this, the HLA restricted CTL epitopes were predicted by HLA binding motif. Sera samples were amplified by PCR, HCV was sequenced and translated into proteins. The changes of predicted CTL epitopes were analysed. RESULTS: No obvious change was observed in predicted CTL epitopes in two chronic HCV patients during 5 and 3 years. CONCLUSION: It demonstrated that the C-terminal of HCV NS3 protein containing the predicted CTL epitopes may have no relationship with HCV persistence. PMID- 10715789 TI - [Interleukin 10(IL-10) levels in serums and in culture supernatants of peripheral blood mononuclear cells from patients with chronic hepatitis B]. AB - OBJECTIVE: To understand the impacts of interleukin 10(IL-10) in serums and in culture supernatants of peripheral blood mononuclear cells (PBMCs) from patients with chronic hepatitis B (CHB) on the persistent infection of hepatitis B virus (HBV). METHODS: The levels of IL-10 in serums and in culture supernatants of PBMCs from 15 patients with CHB and 6 normal controls were measured by ELISA method. PBMCs were cultured with or without Staphylococcus aureus enterotoxin B(SEB; 0.2 microgram/ml) or recombinant HBcAg(rHBcAg; 1.0 microgram/ml) for 48 hours in vitro. RESULTS: The levels of IL-10 in serums was greater in patients with CHB (123.11 +/- 13.89 ng/L) than in controls (95.97 +/- 11.68 ng/L, P < 0.01). Spontaneous, SEB-induced and rHBcAg-induced IL-10 production by PBMCs after culturing in vitro for 48 hours was also higher in patients with CHB (280.82 +/- 50.56 ng/L, 321.69 +/- 37.04 ng/L and 369.58 +/- 30.52 ng/L) than in controls (100.2 +/- 8.54 ng/L, 203.41 +/- 12.02 ng/L and 202.38 +/- 15.79 ng/L; P < 0.01). In addition, IL-10 production by rHBcAg-stimulated PBMCs was significantly greater than by SEB-stimulated PBMCs in patients with CHB. Patients with seropositive for HBV DNA had higher levels of IL-10 in serums and in culture supernatants of PBMCs than patients with seronegative for HBV DNA. But the levels of IL-10 were lower in patients with chronic heavy and severe hepatitis B than in patients with chronic mild and moderate hepatitis B. CONCLUSION: IL-10 may have an important role in persistent infection of HBV in patient with CHB. PMID- 10715790 TI - [Expression of glycoprotein hepatitis C virus in mammalian cell and application of purified protein for detection of antibody against E2 in hepatitis C patients]. AB - OBJECTIVE: E2 glycoprotein of hepatitis C virus was expressed in mammalian cell and purified for detection of antibody against E2 in hepatitis C patients. METHODS: E2/NS1 gene derived from HCV was inserted into expression vector containing six His tag. The recombinant plasmid was transfected into mammalian cells to express E2 glycoprotein expression. E2 glycoprotein was purified by affinity chromophotography. The purified protein was used to establish EIA method for detection of antibodies against E2 in hepatitis C patients. RESULT: Expressed E2 glycoprotein was 7.0 x 10(4). Purification of the purified E2 protein was 90.2%. Twenty-nine patients were anti-E2 antibody positive(82.9%). CONCLUSION: It was the first time to establish EIA method for detection of anti-E2 antibody by purified E2 glycoprotein in China. E2 glycoprotein expressed in mammalian cells had good immunogenity and could increase the sensibility of anti-HCV detection. It suggests that E2 glycoprotein may be useful for development of new anti-HCV reagents. PMID- 10715791 TI - [WAF1/CIP1 gene expression in human hepatocellular carcinoma and its relationship with p53 mutation]. AB - OBJECTIVE: To study the WAF1 gene expression in human hepatocellular carcinoma(HCC) and its relationship with p53 mutation. METHODS: Using semiquantitative reverse transcriptase-polymerase chain reaction(RT-PCR) and immunohistochemistry, the expression of WAF1 and p53 in 32 HCC samples and their surrounding liver tissues and 5 normal liver tissues was detected respectively. RESULTS: The expression level of WAF1 mRNA in HCC samples was lower than that in paratumor liver tissues(1.06 U +/- 0.37 U vs 1.30 U +/- 0.37 U, P < 0.01) and its positive rate of p21WAF1 was also lower than that in paratumor liver tissues(53.1% vs 100%, P < 0.01). The expression level of WAF1 mRNA in HCC samples with intra-hepatic metastatic lesions was lower than that without intra hepatic metastatic lesions(0.92 U +/- 0.33 U vs 1.24 U +/- 0.35 U, P < 0.01) and its positive rate of p21WAF1 was also lower than that without intra-hepatic metastatic lesions(33.3% vs 78.6%, P < 0.05). The expression level of WAF1 mRNA in HCC samples with p53 mutation was lower than that without p53 mutation (0.92 U +/- 0.28 U vs 1.16 U +/- 0.40 U, P < 0.05) and its positive rate of p21WAF1 was also lower than that without p53 mutation(15.4% vs 78.9%, P < 0.01). The percentage of cells in S-phase in p21WAF1-positive HCC samples was lower than that in p21WAF1-negative HCC samples(44.57% +/- 7.56% vs 51.83% +/- 8.51%, P < 0.01). However, no significant difference in expression of WAF1 mRNA and p21WAF1 in HCC samples was found in other clinical pathological parameters such as the degree of differentiation, the histological pattern, the size of tumors, the status of capsule in tumors and the formation of tumor thrombus of portal vein. The expression level of WAF1 mRNA in normal liver tissues was strikingly lower(0.17 U +/- 0.06 U), and the immunohistochemical staining of p21WAF1 in all normal liver tissues was positive(100%). CONCLUSION: The expression of WAF1 gene in HCC is predominantly regulated by dependence on p53 and the reduced WAF1 expression may participate in the carcingenesis and progression of HCC and the mechanisms of infiltration and metastasis of HCC. PMID- 10715792 TI - [Quality control of plasma(serum) specimen for the detection of HCV RNA with PCR]. AB - OBJECTIVE: To define the optimal collection and storage parameters of serum (plasma) for HCV RNA detection. METHODS: The serum(plasma) obtained from HCV RNA positive patients by different means and different processing and storage conditions, then HCV RNA was detected with the fluorescent quantitative PCR. RESULTS: The centrifugation was performed within 2 hours after formation of the clot, loss of HCV RNA titers was 10.42% and significant loss of HCV RNA titers 40.49% was observed after 4 h. The HCV RNA titers in plasma samples with anticoagulants were much higher than that in serum samples(citrate sodium anticoagulized 40.97%, EDTA-anticoagulated 53.14%). There was no loss of HCV RNA with up to three freeze-thaw cycles. A significant decrease in HCV RNA(15.6%) was observed at -20 degrees C after one year. CONCLUSIONS: The results suggested that it is important to process and store clinical samples for HCV RNA detection. The EDTA-K2 as anticoagulants the plasma must be collected within 3 h, and the serum must be prepared within 2 hours after hemosposia. It is essential to store the sample under -70 degrees C for the stability of HCV RNA in a longer time. PMID- 10715793 TI - [Expression and its clinical significance of tissue factor pathway inhibitor and antithrombin-III in patients with hepatic inflammatory diseases]. AB - OBJECTIVE: To investigate the change and clinical significance of tissue factor pathway inhibitor (TFPI) and antithrombin-III (AT-III) in the plasma of patients with hepatic inflammatory disease (HID). METHODS: The plasma levels and activation of TFPI and AT-III in patients with HID (Group I, n = 82) were compared to that in healthy people (Group II as control, n = 30). RESULTS: The plasma level of TFPI:Ag in Group I was higher (P < 0.05) and its activation was also increased dramatically (P < 0.01), the level of AT-III: A was lower in patients with chronic hepatitis B (P < 0.05) and no difference was found in other indices between the two groups. CONCLUSIONS: The elevation of TFPI:Ag and TFPI:A in hepatic inflammatory diseases is an indication of inflammation, and hepatic tissue injury in broad, the lower level of AT-III liver secretions. PMID- 10715794 TI - [Construction of mammalian expression plasmid of Flag-tagged rat collagenase and its in vitro transfection study]. AB - OBJECTIVE: A mammalian expression plasmid of rat collagenase was constructed and its expression in NIH3T3 cells in vitro was studied. METHODS: PCR and gene recombinant techniques were used to construct a mammalian expression plasmid of Flag-fusion rat collagenase. This plasmid was then transferred into cultured NIH3T3 cells mediated by lipofectamine. The mRNA and protein expression of the Flag epitope tagged rat collagenase were detected by RT-PCR and western blot techniques. The enzyme activity of expressed collagenase was detected by gelatin zymography. RESULTS: After transfection the mRNA expression of Flag-fusion rat collagenase could be detected in NIH3T3 cells and the secretion of this collagenase could be detected in the culture medium, and this recombinant collagenase kept the gelatin degradation activity. CONCLUSION: This constructed plasmid can express active rat collagenase in vitro and can be used for further study. PMID- 10715795 TI - [Effects of MMP-1 expressing plasmid on rat liver fibrosis]. AB - OBJECTIVE: To observe the effects of MMP-1(Matrix Metalloproteinase-1) expressing plasmid on rat liver fibrosis. METHODS: We constructed the rat's MMP-1 recombinant plasmid which can be expressed in eucaryotic cells. After encapsulating the MMP-1 recombinant plasmid by lipofectin, we transferred it to the rat's fibrotic liver in vivo intraperitoneally, then observed the effects of the plasmid on the fibrotic liver by RT-PCR, immunohistochemistry and the observation of pathology. RESULTS: The recombinant plasmid of MMP-1 could increase the degradation of type I and type III collagen in the rat's fibrotic liver according to immunohistochemistry compared with fibrotic modal group(P < 0.01) or compared with colchicine group(P < 0.05). According to the observation of pathology, recombinant plasmid of MMP-1 has some reverse effects on the rat's fibrotic liver compared with fibrotic model group(P < 0.05), but no obvious difference was found between the plasmid and colchicine(P > 0.05). CONCLUSION: The recombinant plasmid of MMP-1 has some reverse effects on liver fibrosis, but the effects is limited. PMID- 10715796 TI - [Study on endothlin and its gene expression in splanchnic vessels in cirrhotic rats]. AB - OBJECTIVE: To investigate the role of endothelin(ET) in the pathogenesis of portal hypertension, ET-1 level and its gene expression in splanchnic vessels from cirrhotic rats were observed. METHODS: ET levels of both plasma and vascular tissues were determined by radioimmunoassay. ET mRNA in vascular tissues was probed with RT-PCR technique expressed as optical density(OD) value from image analysis. RESULTS: ET peptide and mRNA in PV and SMA vessels were all significantly higher in cirrhotic rats than those in normal rats, while in cirrhotic rats, ET and its gene expression of PV was dramatically higher than that of SMA(P < 0.05). In addition, the positive correlation was observed in difference between PV and SMA in ET concentration with portal pressure(r = 0.737, P < 0.01). CONCLUSION: ET may be involved in the mechanisms of the formation of portal hypertension mainly due to constructing portal vein, increasing portal flow resistance. PMID- 10715797 TI - [DNA vaccination of the induction of immune responses by codelivery of IL-12 expression vector with hepatitis C structural antigens]. AB - OBJECTIVE: In an attempt to demonstrate the utility of DNA vaccines for the tailored methods, the efficacy of immune responses would be enhanced and the types of immune responses shifted. METHODS: Four recombinant plasmids were constructed. These included the HCV coding regions for the core protein(pC) and for the core E1 and E2 together(pCE1E2) IL-12 p35 and p40. These plasmids were transfected into mammalian cells to test their protein expression and were injected into the quadriceps muscles of BALB/C mice to measure specific antibodies and cytotoxic T-lymphocyte responses. RESULTS: All the recombinant plasmids were shown to express specific antigens in cells transiently and stably. Codelivery of pIL-12 expression cassettes with pC and pCE1E2 in mice resulted in splenomegaly and the increasing number of the splenocytes. It also resulted in the enhancement of Ag-dependent CTL responses and the reduction of specific Ab response. The CTL activity were pC = 18.65% +/- 5.71%, pCE1E2 = 20.07% +/- 11.11%, pC + pIL-12 = 60.11% +/- 17.37%, pCE1E2 + pIL-12 = 67.48% +/- 15.57%, respectively. The Anti-HCV activity were pC = 0.415 +/- 0.127, pCE1E2 = 0.358 +/- 0.096, pC + pIL-12 = 0.210 +/- 0.086, pCE1E2 + pIL-12 = 0.258 +/- 0.125, respectively. CONCLUSION: Codelivery of pIL-12 with plasmid DNA can enhance the efficacy of immune responses and shift the type of immune responses. This work demonstrates the power of DNA delivery in vivo for both the production of a new generation of more effective and targeted vaccines or immuno-therapies as well as an analytic tool for the molecular dissection of the mechanisms of immune function. PMID- 10715798 TI - [Inhibited effect of phosphorylated insulin-like growth factor binding protein-1 on hepatoma cells in vitro]. AB - OBJECTIVE: To elucidate the inhibited effect and regulated mechanism of IGFBp-1 on the carcinoma cells. METHODS: The recombinant human IGF II (rhIGF II), phosphorylated and nonphosphorylated binding protein-1(pBp-1 and npBp-1) that were purified from pregnant woman's amnoitic fluid were used to cultivate cancer cells from 15 cases with hepatocellular carcinoma and ascites. The cell counting and MTT methods were used for determining the dynamics change of proliferation on HCC, and the in situ or dot hybridization with Digoxin label used for the expression of IGF II mRNA qualitatively and quantitatively at different times. RESULTS: The cells grew more slowly, in pBp-1 group, than in control group at the same time, which began to decrease at two days(P < 0.05) by MTT. In situ hybridization showed that there are hybridization signal of IGF II mRNA in all groups during the culture, and scanning dot hybridization showed the OPTMD of IGF II mRNA in pBp-1 group was statistically different from control and other groups (P < 0.01), while in npBp-1 group there was no significant effect compared with other groups (P > 0.05). CONCLUSION: The pBp-1 can inhibit the growth of hepatoma cells and the expression of IGF II mRNA, which may be regulated and inhibited by pBp-1. PMID- 10715799 TI - Long-term outcomes of patients with flail mitral valve leaflets. PMID- 10715800 TI - Doppler echocardiographic assessment of mitral regurgitation. PMID- 10715801 TI - Vasoactive medications in the treatment of severe mitral regurgitation. PMID- 10715802 TI - Exercise echocardiographic assessment in severe mitral regurgitation. AB - In chronic severe mitral regurgitation, minimum morbidity and mortality is achieved by applying surgical correction before left ventricular dysfunction becomes irreversible. This requires detection of subtle signs of early ventricular decompensation, for which isotonic stress echocardiography is more accurate than is use of resting indices of contractile function alone. We perform serial 6-monthly stress echocardiography for patients with severe mitral regurgitation, and recommend surgery when the exercise end-systolic volume index or ejection fraction reaches the cutoff values in Table 4 or if there is a clear adverse trend. Exercise echocardiography is more accurate than is exercise electrocardiography for detecting concomitant coronary disease prior to revascularization. Stress testing is also an objective measure of symptoms. Color Doppler stress echocardiography can detect those patients whose mitral regurgitation worsens (or even develops de novo) with exercise, which can explain unexpected symptoms. Stress echocardiography, therefore, provides a comprehensive and cost-effective evaluation of patients with mitral regurgitation that combines functional, diagnostic, and prognostic information. PMID- 10715803 TI - Repair versus replacement of mitral valve for treating severe ischemic mitral regurgitation. PMID- 10715804 TI - Cardiovascular response to combined static-dynamic exercise of patients with myocardial infarction. AB - BACKGROUND: Graded dynamic exercise-stress testing of patients with acute myocardial infarction prior to discharge from hospital has an important diagnostic and prognostic implication. Although many daily tasks involve combinations of static and dynamic exercise, little is known about cardiovascular responses during combined static-dynamic exercise. OBJECTIVE: To determine the difference between cardiovascular responses during two types of combined static dynamic exercise (a 10 kg weight in one hand, and a 10 kg weight bearing on the shoulder). METHODS: We studied 27 male patients who had recently suffered myocardial infarction using ear densitography. The patients were divided into two groups: group 1 was comprised of 14 patients with resting left ventricular end diastolic volumes > or = 140 ml, and group 2 was comprised of 13 patients with left ventricular end-diastolic volumes < 140 ml. RESULTS: For eight patients in group 1 we detected positive electrocardiographic changes during one-hand weight carrying exercise, but for none of these patients was there an electrocardiographic change during weight-bearing exercise. All the patients in group 2 completed both types of exercise without significant ST-segment change. Although there were no significant differences between values of any of the indices measured for the two groups during weight-bearing exercise, patients in group 1 had significantly shorter diastolic times/min (21.8 +/- 2.1 versus 25.1 +/- 2.4 s/min, P < 0.01) during one-hand weight carrying. CONCLUSIONS: In addition to decrease in subendocardial coronary blood flow associated with increase in left ventricular end-diastolic volume, shortening of diastolic perfusion time during one-hand weight-carrying exercise for patients in group 1 can potentially contribute to subendocardial ischemia, which was favorably altered by bearing a weight on the shoulder. PMID- 10715805 TI - Accelerated coronary atherosclerosis and arteriosclerosis in young human immunodeficiency-virus-positive patients. AB - OBJECTIVE: To determine the type of lesions observed in young patients infected with human immunodeficiency virus-1 (HIV-1). DESIGN: Examination of coronary networks in corpses of 13 men and two women who had died aged 23-32 years after having been infected with HIV-1 virus, having been seropositive for 2-5 years. Causes of death were infectious complications (five cases), infection with cytomegalovirus leading to gastro-intestinal haemorrhaging (one case), infection with cytomegalovirus and Kaposi's sarcoma (one case), overdoses of drugs (five cases) and sudden death (three cases). METHODS: The pathological analysis was carried out on the proximal and distal coronary networks. In order to characterize the lesions better, the cells and the cytokines involved were characterized by immunohistochemistry. RESULTS: In all 15 cases we observed thickening of intima in the proximal network at least as great as that of the media, caused by a proliferation of secreting cells, phenotypically identified as smooth muscle cells, with exaggerated production of elastic fibres and in association with an increase in the expression of tumor necrosis factor-alpha and interleukin-1 alpha. In nine cases, atherosclerosis had developed from and on the surface of this proliferation and in four cases arteriosclerosis had an unusual appearance, in the form of mamillated vegetations with endoluminal protrusions. A similar proliferation was found in the distal network in four cases, but with a significantly smaller proportion of elastic fibres. CONCLUSIONS: The lesions we examined in these young HIV-1-infected patients presented particular features and were intermediate between the lesions observed during common coronary atherosclerosis and atherosclerosis associated with chronic rejection of cardiac transplants. PMID- 10715806 TI - Genetic polymorphism of 5,10-methylenetetrahydrofolate increases risk of myocardial infarction and is correlated to elevated levels of homocysteine in the Japanese general population. AB - BACKGROUND: Hyperhomocysteinemia, an independent and graded risk factor for coronary artery disease, can result from both environmental and hereditary factors. C677T mutation of the 5,10-methylenetetrahydrofolate reductase (MTHFR) gene [alanine/valine (A/V) polymorphism], one of the key enzymes involved in catalyzing the remethylation of homocysteine, has recently been reported. OBJECTIVE: To evaluate the incidence of the MTHFR genotypes and their significance in determining the risk for myocardial infarction of Japanese men. METHOD: The subjects consisted of 199 healthy men (mean age, 60 years) and 230 male patients with myocardial infarction (mean age, 59 years). The coronary artery lesions were evaluated by coronary angiography. The MTHFR genotype was analyzed by polymerase chain reaction and then by digestion with Hinfl. Total plasma levels of homocysteine for each MTHFR genotype were compared with those in healthy controls. RESULTS: The prevalences of the A and V alleles among the healthy male subjects were 0.652 and 0.348 in the Hardy-Weinberg equilibrium. The total levels of homocysteine in the plasma of the healthy male subjects were 8.6 +/- 3.3, 8.9 +/- 4.1, and 11.6 +/- 5.6 mumol/l, for AA, AV, and VV genotypes, respectively. Individuals with the VV homozygous mutant genotype thus had the highest plasma levels of homocysteine. Logistic analysis revealed that the levels of high-density lipoprotein cholesterol, hypertension, diabetes mellitus, MTHFR VV genotype, and triglycerides were all independent risk factors for myocardial infarction. The VV genotype was more prevalent among patients with myocardial infarction (mean age, 59 years) than it was among the control subjects (17.0 versus 10.6%, P < 0.05). However, there were no differences in the numbers of stenotic coronary arteries among the MTHFR genotypes. CONCLUSION: The VV genotype of MTHFR increases plasma levels of homocysteine in healthy controls, and this mutation indicates a genetic predisposition toward a greater than normal risk of myocardial infarction for Japanese men. PMID- 10715807 TI - Enhanced accumulation of pericardial fluid ferritin in patients with coronary artery disease. AB - BACKGROUND: Ferritin is a storage protein for iron that can either represent a source of iron or perform a cytoprotective action as an iron sequestrant. OBJECTIVE: To compare the concentrations of ferritin in pericardial fluid of patients with valvular heart disease, serving as controls, and in patients with coronary artery disease. DESIGN: We studied a total of 59 consecutive male patients undergoing elective heart valve replacement (group 1: n = 22, mean +/- SD age 55 +/- 11 years) or elective coronary artery bypass grafting (group 2: n = 37, mean +/- SD age 59 +/- 9 years). METHODS: Iron status indicators, total protein and albumin concentrations, and lactate dehydrogenase activities were determined in pericardial fluid and serum samples obtained from patients during surgery. RESULTS: Pericardial fluid concentrations of ferritin in both patient populations were significantly (P < 0.001) greater than the concentrations in sera: group 1, 375 (107-2030) micrograms/l compared with 146.5 (21-407) micrograms/l; group 2, 1115 (226-2500) micrograms/l compared with 152.0 (16-398) micrograms/l (median (range)), respectively. Moreover, pericardial fluid ferritin concentration was significantly (P < 0.01) greater in patients undergoing coronary artery bypass grafting than in those undergoing heart valve replacement, whereas serum ferritin concentrations did not differ between the two patient populations. CONCLUSIONS: As pericardial fluid reflects the composition of the myocardial interstitium, we suggest that ferritin released can serve as a potential source of iron in the cardiac interstitium that may promote the generation of oxygen free radicals. Conversely, we presume that induction of ferritin synthesis, representing an important mechanism by which tissue adapts to hypoxic damage, can afford myocardial cytoprotection. PMID- 10715808 TI - Greater than normal expression of the collagen-binding stress protein heat-shock protein-47 in the infarct zone in rats after experimentally-induced myocardial infarction. AB - BACKGROUND: The heat-shock protein with relative molecular mass 47,000 (HSP47) can bind to procollagen molecules in the endoplasmic reticulum, and acts as a molecular chaperone during the processing and secretion of procollagen. OBJECTIVE: To test our hypothesis that HSP47 is expressed in the myocardial infarct zone. METHODS: We induced myocardial infarction in male Sprague-Dawley rats by ligation of left coronary artery. The expression of HSP47 was examined by Northern blotting, in-situ hybridization, Western blotting and immunohistochemistry. The time-dependent change in the distribution of HSP47 messenger RNA (mRNA) signal was compared with the changes in expression of alpha 1(I) and alpha 1(III) collagen mRNA by in-situ hybridization. The hypoxic induction of HSP47 in cultured cardiac fibroblasts was examined by Northern-blot analysis. RESULTS: Northern blotting demonstrated that the expression of HSP47 mRNA had increased on day 2, reaching a maximum level around day 14 (induced 3.5 fold compared with the preligation hearts) and was maintained at a high level up to day 28. In-situ hybridization analysis revealed HSP47 mRNA signals in spindle shaped mesenchymal cells located between surviving myocytes in the infarct's peripheral zone 24 h after the ligation, and in the entire infarct zone on day 14. The sequential changes in distribution of HSP47 mRNA signal were identical to those of the alpha 1(I) and alpha 1(III) collagen mRNA. Western blotting demonstrated that expression of HSP47 protein in the infarct zone had increased. Immunofluorescent staining revealed positivity for HSP47 in the infarct's peripheral zone on day 2 and in the entire infarct zone on day 14. Northern blotting revealed that the expression of HSP47 mRNA in cultured cardiac fibroblasts in hypoxic cultures was greater than that in normoxic cultures. CONCLUSION: The present data demonstrated that an increase in expression of HSP47 is produced by spindle-shaped mesenchymal cells in the infarct zone. Expression of HSP47 mRNA was concurrent with the expression of collagen mRNA of types I and III. Hypoxia is one of the factors which induces expression of HSP47. PMID- 10715809 TI - Insulin and insulin-like growth factor-I cause vasorelaxation in human vessels in vitro. AB - BACKGROUND: Insulin and insulin-like growth factor-I (IGF-I) are endogenous peptides with vasoactive activities. OBJECTIVE: To evaluate the vasodilatory effects of insulin and IGF-I on human vessels taken from patients with and without noninsulin-dependent diabetes mellitus (NIDDM) and to elucidate their mechanisms of action. METHODS: Vascular rings of human internal mammary artery (IMA) and saphenous vein harvested from 54 patients with and without NIDDM undergoing coronary bypass surgery were studied in vitro. RESULTS: For samples from patients without NIDDM both insulin and IGF-I (10(-12)-10(-7) mol/l) evoked greater relaxation in IMA rings (30 +/- 4 and 29 +/- 6%, maximal relaxation +/- SEM, respectively) than they did in saphenous-vein rings (43 +/- 4 and 42 +/- 5%, respectively, P < 0.05 both for insulin and for IGF-I). Similar results were obtained with vessels from patients with NIDDM. Relaxation was not affected by the removal of the endothelium and by inhibition of the production of nitric oxide. However, the vascular relaxation caused by insulin and IGF-I was completely abolished by KCI, and was attenuated by the nonspecific potassium channel blocker tetraethylammonium (for IMA rings, to 77 +/- 8 and 66 +/- 4% with insulin and IGF-I, respectively; for saphenous vein rings, 73 +/- 2 and 77 +/- 1% for insulin and IGF-I, respectively, P < 0.001). CONCLUSIONS: Both insulin and IGF-I induced endothelial-independent, nitric oxide-independent vasorelaxation of rings from human IMA and saphenous veins, through a mechanism involving activation of potassium channels. This response remained intact in vessels from patients with NIDDM. This result supports the hypothesis that insulin and IGF-I play roles in the regulation of vascular tone in human vessels. PMID- 10715810 TI - Circulating adhesion molecules and severity of coronary atherosclerosis. AB - BACKGROUND: Circulating leukocytes are recruited at atherosclerotic sites through a family of adhesion molecules. Circulating forms of adhesion molecules in peripheral blood can be quantified now. OBJECTIVE: To evaluate the relationship between circulating adhesion molecules and severity of coronary atherosclerosis. METHODS: Subjects included 81 patients undergoing diagnostic coronary angiography, 12 of whom had normal coronary arteries (control group). The remaining 69 patients with demonstrable coronary atherosclerosis were divided into two groups by use of Gensini scores, namely mild atherosclerosis (n = 36, Gensini score 1-20) and severe atherosclerosis (n = 33, Gensini score > 20). Serum levels of circulating intercellular adhesion molecule-1 (ICAM-1), vascular cellular adhesion molecule-1 (VCAM-1), and E-selectin of groups measured before angiography were compared. RESULTS: Circulating levels of ICAM-1 in members of mild and severe atherosclerosis groups were significantly higher than those in members of the control group, whereas there was no significant difference among circulating levels of VCAM-1 in members of the three groups. Circulating levels of E-selectin in members of the mild atherosclerosis group were significantly higher than those in members of the severe atherosclerosis and control groups. CONCLUSIONS: These findings suggest that E-selectin is related to the early stage, and ICAM-1 is related to the advanced stage, of coronary atherosclerosis. With progression of atherosclerosis, one-step adhesion by ICAM-1 could become more important than multistep adhesion involving E-selectin, ICAM-1, and VCAM-1. These molecules may serve as markers for severity of coronary atherosclerosis. PMID- 10715811 TI - Effects of pimobendan and EGIS 9377, cardiotonic agents, and OG-VI, a nucleoside nucleotide mixture, administered during reperfusion after ischemia on stunned myocardium in dogs. AB - BACKGROUND: Pimobendan is a so-called calcium sensitizer that exerts a positive inotropic action. EGIS 9377 is synthesized as a calcium sensitizer. OG-VI is a nucleoside-nucleotide mixture that ameliorates the myocardial dysfunction (myocardial stunning) after ischemia. OBJECTIVE: To determine whether administration of these agents after the onset of reperfusion after ischemia improves the condition of stunned myocardium. METHODS: Dogs anesthetized with pentobarbital were subjected to 20 min ligation of left anterior descending coronary artery and then 60 min reperfusion. The corresponding vehicle, 0.3 and 1 mg/kg pimobendan, or 1 and 3 mg/kg EGIS 9377 was injected intravenously 30 min after the onset of reperfusion. Saline solution or 1.2 mumol/kg per min OG-VI was infused for 30 min, starting 30 min after the reperfusion. Shortening of myocardial segment was measured by sonomicrometry. The tissue levels of energy and carbohydrate metabolites in the 60 min-reperfused hearts were determined. RESULTS: Shortening of myocardial segments significantly decreased during ischemia, and returned toward preischemic level after reperfusion for all groups, although the contractile dysfunction still remained. Injections and infusion of pimobendan, EGIS 9377, and OG-VI after the onset of reperfusion ameliorated the contractile dysfunction. Systemic vascular resistance was decreased by administrations of pimobendan and OG-VI. The levels of high-energy phosphates in 60 min-reperfused heart were not changed by either treatment. CONCLUSION: Administration of pimobendan, EGIS 9377, and OG-VI ameliorate the myocardial contractile dysfunction after ischemia even when these agents are administered after the onset of reperfusion. The increase in contractile function due to these agents did not worsen the myocardial energy balance. PMID- 10715812 TI - Bibliography. Current world literature. PMID- 10715813 TI - Characterization of the capsid protein gene from a nodavirus strain affecting the Atlantic halibut Hippoglossus hippoglossus and design of an optimal reverse transcriptase polymerase chain reaction (RT-PCR) detection assay. AB - A 1349 nucleotide fragment of the RNA2 from a nodavirus affecting Atlantic halibut Hippoglossus hippoglossus was characterised and the nuclotide sequence (accession no. AJ245641) was employed to develop an optimal reverse-transcriptase polymerase chain reaction (RT-PCR) detection assay. The sequenced part of the RNA2 of Atlantic halibut nodavirus (strain AH95NorA) was highly similar in organisation to that of the RNA2 of striped jack nervous necrosis virus (SJNNV), and comprised features common to all nodaviruses. These characteristics confirmed that the virus that causes viral encephalopathy and retinopathy (VER) in Atlantic halibut is a nodavirus. The nucleotide sequence of the 1349 nucleotide fragment of Atlantic halibut nodavirus RNA2 was 80% identical to the RNA2 of SJNNV. The T2 region (830 nucleotides) of the RNA2 of Atlantic halibut nodavirus shared 98% of the nucleotide sequence when compared with the homologous region of barfin flounder nervous necrosis virus (BFNNV), while the nucleotide sequence identity to SJNNV in this region was 76%. Phylogenetic analysis based on the nucleotide sequences of the T4 region (421 nucleotides) of Atlantic halibut nodavirus and of other fish nodaviruses revealed a close relationship to the nodaviruses of the barfin flounder clad that have been found in other cold-water species (Pacific cod Gadus macrocephalus and barfin founder Verasper moseri). The nucleotide sequence of the RNA2 of Atlantic halibut nodavirus included some features that differ from that of SJNNV. The ORF of the RNA2 of Atlantic halibut nodavirus lacked 6 nucleotides through a single deletion and a 5-nucleotide deletion, separated by 4 nucleotides. The 3'-non-encoding region contained a 21 nucleotide insert and a 3 nucleotide deletion when compared with SJNNV. In comparison with the RNA2 of SJNNV, the 3'-non-encoding region showed a nucleotide sequence identity of 84.5%. A primer set based on the Atlantic halibut nodavirus nucleotide sequence was employed in order to design an optimal RT-PCR. The detection limit of the PCR was 10 to 100 copies of plasmid, while the detection limit of the RT-PCR assay was 100 to 1000 copies of in vitro transcribed viral RNA. PMID- 10715814 TI - Surface disinfection of Atlantic halibut Hippoglossus hippoglossus eggs with ozonated sea-water inactivates nodavirus and increases survival of the larvae. AB - Disinfection by ozonation of sea-water may reduce the risk of transmission of nodavirus, a major fish pathogen, via Atlantic halibut Hippoglossus hippoglossus eggs. In the present study, eggs at 4 d prior to hatching were exposed to nodavirus and then to ozonated sea-water using different concentrations (0.3 to 10 mg l-1) and exposure times (0.5 to 10 min). None of the larvae from virus exposed eggs washed with ozonated sea-water developed viral encephalopathy and retinopathy (VER), which was detected in all dead larvae from eggs exposed to nodavirus but not washed with ozonated sea-water. In the non-treated control group about 20% of the dead larvae developed the disease. This suggests that the halibut eggs taken from a large-scale production facility were already contaminated with nodavirus. The egg groups which had been treated with 4 mg O3 l 1 for 0.5 min or with lower total ozone exposures had a higher survival and no adverse effects on the development of the larvae after hatching were observed. Although a slight delay in hatching was found, after 2 d the cumulative hatching had normalised. In the egg groups with high total exposure (4 mg O3 l-1 for 1 min or higher total ozone exposures) a pronounced negative effect on hatching was observed. Our results indicate that the egg surface may be important in the transfer of nodavirus and that nodavirus associated with the surface of the egg may be inactivated by ozonated sea-water. PMID- 10715815 TI - Platyspondyly and shortness of vertebral column in farmed Atlantic salmon Salmo salar in Norway--description and interpretation of pathologic changes. AB - Body malformation due to shortness of the vertebral column, in most cases of unknown cause, has been observed in fish for more than 100 yr. It periodically occurs with high prevalence in farmed Atlantic salmon Salmo salar in Norway, and this paper describes the results of macroscopic, radiographic and histologic examination of parr and seawater-transferred fish. The vertebral bodies in both age groups did not acquire the length that they normally should due to a growth disturbance leading to the condition of platyspondyly and shortness in the column. The pathologic changes became visible at different ages in both groups and the process apparently starts in intervertebral tissues. There was proliferation of connective tissue and blood vessels, and sometimes infiltration with inflammatory cells, around affected vertebrae, especially in seawater transferred fish. This is the first description of inflammation in abnormally short-spined fish, and it may indicate an infectious etiology, at least in farmed seawater-transferred salmon. PMID- 10715816 TI - Genetic diversity of the fish pathogen Aeromonas salmonicida demonstrated by random amplified polymorphic DNA and pulsed-field gel electrophoresis analyses. AB - The current taxonomy of Aeromonas salmonicida includes 4 subspecies. A. salmonicida subsp. salmonicida is associated with salmonid furunculosis, and A. salmonicida subsp. achromogenes, A. salmonicida subsp. masoucida, and A. salmonicida subsp. smithia are strains that show variation in some biochemical properties. This classification does not readily encompass isolates from a wide range of fish hosts currently described as atypical A. salmonicida. This study examined 17 typical strains, 39 atypical strains and 3 type A. salmonicida subspecies strains for genetic similarity using the random amplified polymophic DNA (RAPD) and pulsed-field gel electrophoresis (PFGE) techniques. On the basis of RAPD- and PFGE-derived profiles, similarity matrices and dendrograms were constructed. The results showed that species A. salmonicida constituted a genetically heterogeneous group of strains, encompassing within an homogeneous or clonal lineage comprised solely of typical strains and the A. salmonicida subsp. salmonicida type strain. PMID- 10715817 TI - Histopathological changes in the swimbladder wall of the European eel Anguilla anguilla due to infections with Anguillicola crassus. AB - The histopathological changes in swimbladders of European eels naturally and experimentally infected with Anguillicola crassus were studied using transmission and scanning electron microscopy. During the course of probably several infections swimbladders undergo characteristic changes. In addition to the thickening of the entire swimbladder wall, and to the folded internal surface of this organ, inflammation, migration of white blood cells, fibrosis and changes in the epithelial cells are frequently seen. Epithelial cells tend to proliferate heavily and form hyperplastic tissues; these processes are accompanied by changes in the internal structure of the cells. The normally cubic cells become spherical or columnar and form folds facing the lumen of the swimbladder. As a consequence, most of these cells lose contact with the blood vessels and show no strict polarity. In heavily affected swimbladders the basal labyrinth of the epithelial cells is reduced, i.e. becomes shorter and less densely packed. The lamina propria shows severe fibrosis with infiltration of white blood cells. Larvae of A. crassus, inhabiting the wall of the swimbladder, were found to be surrounded by cell debris, but this local necrosis does not affect the entire swimbladder in its overall structure. These histological findings can partly explain changes in the gas composition in eels infected with A. crassus. PMID- 10715818 TI - In vitro studies on optimal requirements for the growth of Spironucleus vortens, an intestinal parasite of the freshwater angelfish. AB - Spironucleus vortens were cultivated in either an artificial medium at different temperatures, or in medium at various pH conditions or supplemented with different bile concentrations at 25 degrees C. Temperature, pH and bile requirements for the optimal growth of the parasite were determined. Parasites multiplied quickly at 28 and 31 degrees C and reached maximum numbers on Day 4 of cultivation, whereafter they did not survive. At 25 degrees C, parasites survived longer than those at 28 and 31 degrees C with no difference in multiplication rate during the exponential phase. The longest survival period was seen at 22 degrees C, although the growth rate of the parasite was not as high as those at 25 degrees C. At a higher temperature of 37 degrees C, no parasites were observed alive after the second day of cultivation. Optimal pH range for the parasite's growth was 6.5 to 7.5, with the highest cell number at pH 7.5. Parasites survived longest (15 d) at pH 6.0, although the maximum number of cells was lower than those at the optimal pH. Parasites were dead within 24 h at pH levels above 8.5 or below 5.5. All cultures supplemented with either bovine or fish bile yielded numbers of parasites lower than cultures with no bile. In addition, parasite growth was significantly suppressed in medium supplemented with higher concentrations of bile. These results indicate that the optimal condition for the in vitro cultivation of S. vortens is 25 degrees C and pH 6.5 to 7.5 without supplementation with bile. PMID- 10715819 TI - Preliminary description of lesions in juvenile largemouth bass injected with largemouth bass virus. AB - Juvenile largemouth bass Micropterus salmoides were intraperitoneally injected with largemouth bass virus (LMBV), a member of the genus Ranavirus, family Iridoviridae. Moribund fish which had been injected with 10(6.2) tissue culture infectious doses, 50% endpoint (TCID50), were sampled 4 d after injection; other largemouth bass injected with this dose died between 3 and 5 d after injection. Fish injected with 10(2.8) TCID50 of LMBV were also examined after 4 d and had lesions similar to those of fish injected with the high dose. Clinical signs included darker pigmentation, inflammation and necrosis at the site of injection, distended abdomen, corkscrew swimming, and lateral recumbency. Internally, fish had focally pale livers, bright red spleens and reddened intestinal ceca. Histologically acute fibrinous peritonitis affected the surface of all organs in the peritoneal cavity, but deeper portions of organs appeared normal. There was also necrosis of the gastrointestinal mucosa. Except for the injection site, lesions were confined to the peritoneal cavity. PMID- 10715820 TI - Virus-like particles associated with mortalities of the carpet-shell clam Ruditapes decussatus. AB - During the late summer and early fall of 1997 and 1998 heavy mortalities were detected in the carpet-shell clam Ruditapes decussatus cultured in Galicia (NW Spain). The prevalences and intensities of the parasites found in the histopathological studies were not high enough to explain the high mortality rates. Unenveloped virus-like particles were detected by transmission electron microscopy in the cytoplasm of the connective tissue cells. They had an icosahedrical-spherical shape with a diameter of 27 to 35 nm. These virus-like particles appeared free in the cytoplasm or associated with endoplasmic reticulum membranes. These characteristics suggest that they belong to the Picornaviridae family. PMID- 10715821 TI - Recovery of ranavirus dsDNA from formalin-fixed archival material. AB - The extraction and amplification of nucleic acid from formalin-fixed and paraffin embedded tissues has become an important exercise in the collection of retrospective epidemiological data. A protocol is described that enables the extraction and amplification of dsDNA from fixed tissues within paraffin blocks and from specimens stored in 10% (aq) formalin. The procedure can be used for the examination of ranavirus DNA within archival tissues thereby providing valuable data for identifying the origin and tracing the spread of ranaviruses. PMID- 10715822 TI - Reduced superoxide dismutase activity in Palaemonetes argentinus (Decapoda, Palemonidae) infected by Probopyrus ringueleti (Isopoda, Bopyridae). AB - Cellular oxidative stress may promote damage or death in biological systems and may be caused by production of pro-oxidant molecules known as reactive oxygen species (ROS). The aim of this work was to analyze the activity of antioxidant enzymes (catalase [CAT], superoxide dismutase [SOD] and glutathione peroxidase [GPx]) in the shrimp Palaemonetes argentinus Nobili, 1901 infected by Probopyrus ringueleti (Verdi & Schuldt, 1987), a gill chamber parasite known for its capacity to cause host metabolic changes, including changes in oxygen consumption rates. Infested and non-infested shrimp were collected in the Patos Lagoon estuary (southern Brasil), where the prevalence of the parasite may be as high as 70%. No significant differences were observed for either CAT or GPx activities. However, SOD activity was significantly reduced in infected shrimp, suggesting that bopyrid isopod respiratory impairment resulted in reduced SOD enzyme activity. PMID- 10715823 TI - Meeting the preteen vaccine law: a pilot program in urban middle schools. AB - California, the most populous state in the nation, is one of many states that implemented vaccination requirements for preteens. While kindergarten requirements are well-established and accepted by parents, implementation of preteen vaccination requirements requires inter- and intra-institutional adjustments, educational and public relations efforts, and an augmentation of vaccination delivery systems. This article describes a pilot program in two middle schools in an urban school district and offers planning strategies and practical tools to assist school nurses and health providers to implement preteen requirements. PMID- 10715824 TI - A model for mapping linkages between health and education agencies to improve school health. AB - Efforts to develop effective and sustainable school health programs evolved in sophistication the past 20 years through research and practical experience. This paper reviews these developments, arguing they were significantly driven by public health priorities, and have not adequately accounted for educational perspectives and priorities. To better understand the differences in perspective, a model is presented which illustrates linkages between different school-based inputs and strategies, and long-term health and educational outcomes. The model describes similarities and differences between the two perspectives. A significant coincidence exists in factors that determine educational attainment and improved health outcomes for students. A more holistic and integrated approach to school health is emerging, and at these interfaces our implementation and research efforts for the 21st century should be concentrated. PMID- 10715825 TI - Kindergarten children's knowledge and perceptions of alcohol, tobacco, and other drugs. AB - Kindergarten children's knowledge and perceptions of alcohol, tobacco, and other drugs (ATODs) were assessed and the congruence between parent ATOD use and children's knowledge of ATODs was examined. Data were collected during the pre intervention phase of an ATOD prevention trial with 5- and 6-year-old children and their parents. Three elementary schools were randomly selected from a population of 15 high-risk elementary schools in Lexington, Ky., (n = 126 parent child dyads). Children were interviewed about their knowledge, feelings, and attitudes toward ATODs using the Child Drug Awareness Inventory. Parents self reported ATOD use. Almost all (95%) kindergarten children recognized cigarettes; 56% correctly identified alcoholic beverages; and 17% recognized at least one illicit drug. Minority children were almost four times more likely to recognize illicit drugs than were non-minority children. Children's knowledge of ATODs was not correlated with the parents' reported drug use. ATOD prevention programs for young children merit greater emphasis. PMID- 10715826 TI - Utilizing the SIECUS Guidelines to assess sexuality education in one state: content scope and importance. AB - Comprehensive sexuality education programs, advocated as the most effective means of promoting sexual health for youth, include a broad range of sexuality topics taught throughout the school years as a part of comprehensive health education. Programs must include not only the negative consequences of sexual behavior, but content that cultivates a positive view of sexuality as well. This study assessed the scope of sexuality education topics taught, using the SIECUS Guidelines for Comprehensive Sexuality Education: Kindergarten--12th Grade, and rated importance of these topics, by public high school teachers within a state that mandates sexuality education. The majority of the 261 teachers responding to the survey reported they taught most of the topics recommended by the Guidelines. They also rated most Guidelines' topics as "important" to teach. However, numerous teachers omitted topics crucial to comprehensive sexuality education. The relationship between teacher ratings of importance and what was taught, and implications for future research are discussed. PMID- 10715827 TI - Health, medicine, and food messages in television commercials during 1992 and 1998. AB - The potential effects of television advertisements on knowledge, attitudes, and behavior have generated considerable concern. Part of this concern arises from the overall exposure of children to this medium. By the time they graduate from high school, the time devoted to watching television will exceed the hours spent in school. Hence, health professionals should recognize the disproportionate role of television as an informational and attitudinal source for children. The potential impact of television advertisements, particularly those promoting health-related products such as medications and foods, coupled with the changing nature of television points to this medium as an important candidate for examination. The purpose of this study was to content analyze and compare advertisements broadcast in 1992 and 1998 to create a description of the health information conveyed in top-rated, prime time network television advertisements and to determine the congruence of this information with current health recommendations. PMID- 10715828 TI - Health education and physical education: disciplines preparing students as productive, healthy citizens for the challenges of the 21st century. PMID- 10715829 TI - A cooperative approach to promoting health literacy: the Current Health Issues Project. PMID- 10715831 TI - Contributions of C3H6O+. ions with the oxygen on the middle carbon to gas phase ion chemistry AB - The numerous ways in which studies of C3H6O+. ions with the oxygen on the second carbon have added to our knowledge of gas phase ion chemistry are reviewed. The enol form of this ion (1) first attracted interest during early investigations of the mechanism of the McLafferty Rearrangement and later in characterizing the double McLafferty Rearrangement. Next, it was found that 1 isomerizes to the higher energy acetone ion (2). This discovery sparked studies of the relative stabilities of ionized and neutral enol and ketone species. It also led to the discovery that 1-->2 surmounts a substantial barrier, and that 2 dissociates faster than the excess energy deposited in it by the isomerization can become randomly distributed; i.e., dissociation following 1-->2 is nonergodic. This fact is manifested by a more abundant loss of the methyl formed by isomerization and a greater associated translational energy release. Propene oxide and methyl vinyl ether ions also appear to ionize to 2 and to dissociate nonergodically. Methane is eliminated from 2 through a methyl-acetyl ion complex. Characterization of this reaction by photoionization mass spectrometry, ab initio theory, and RRKM calculations helped to establish that complex-mediated alkane eliminations are generally confined to a region just above threshold. At higher energies, because attractions between ions and nonpolar neutrals are weak, simple dissociation is too fast for complex-mediated H-transfer to compete with it. Studies of the collision-induced dissociations of 2 demonstrate that the first electronically excited A state of 2 is very long-lived and efficiently releases its energy into translational energy when the ion collides with a neutral at low impact energies. Finally, ion-molecule reactions of 2 and acetone-containing ion clusters in the gas phase are described. PMID- 10715830 TI - Membrane introduction mass spectrometry: trends and applications. AB - Recent advances in membrane introduction mass spectrometry (MIMS) are reviewed. On-line monitoring is treated by focusing on critical variables, including the nature and dimensions of the membrane, and the analyte vapor pressure, diffusivity, and solubility in the membrane barrier. Sample introduction by MIMS is applied in (i) on-line monitoring of chemical and biological reactors, (ii) analysis of volatile organic compounds in environmental matrices, including air, water and soil, and (iii) in more fundamental studies, such as measurements of thermochemical properties, reaction mechanisms, and kinetics. New semipermeable membranes are discussed, including those consisting of thin polymers, low vapor pressure liquids, and zeolites. These membranes have been used to monitor polar compounds, selectively differentiate compounds through affinity-binding, and provide isomer differentiation based on molecular size. Measurements at high spatial resolution, for example, using silicone-capped hypodermic needle inlets, are also covered, as is electrically driven sampling through microporous membranes. Other variations on the basic MIMS experiment include analyte preconcentration through cryotrapping (CT-MIMS) or trapping in the membrane (trap and-release), as well as differential thermal release methods and reverse phase (i.e., organic solvent) MIMS. Method limitations center on semivolatile compounds and complex mixture analysis, and novel solutions are discussed. Semivolatile compounds have been monitored with thermally assisted desorption, ultrathin membranes and derivatization techniques. Taking advantage of the differences in time of membrane permeation, mixtures of structurally similar compounds have been differentiated by using sample modulation techniques and by temperature programmed desorption from a membrane interface. Selective ionization techniques that increase instrument sensitivity towards polar compounds are also described, and comparisons are made with other direct sampling (nonchromatographic) methods that are useful in mixture analysis. PMID- 10715832 TI - Nutrition and blood lipids. PMID- 10715833 TI - Nutrition and blood pressure. AB - Nutrition plays a very important role in regulating blood pressure. If we reverted to our evolutionary diet the problem of high blood pressure would disappear. However, this is unlikely and we, therefore, need to identify the most important factors in our diet that predispose to high blood pressure and, therefore, to vascular disease. Studies clearly demonstrate the very important role of our current intake of salt in our diet as being the major factor in regulating blood pressure in populations. Other dietary factors have also been identified as playing an important role, particularly potassium intake and fruit and vegetable consumption. A more healthy diet, that is a diet with much less salt and increased potassium through an increase in fruit and vegetable consumption, a reduction in fat intake with substitution of saturated by monounsaturated fat, a reduction in meat and dairy products with an increase in fish consumption will have large effects on blood pressure but, at the same time, will decrease other cardiovascular risk factors, particularly cholesterol and glucose intolerance. This healthier diet will reduce cardiovascular disease and is similar to the diet now being advocated for the prevention of some forms of cancer. Diet is by far the most important environmental factor determining our longevity and for those who wish to live longer, a change in diet as early in life as possible will have substantial effects. PMID- 10715834 TI - Nutrition and thrombogenic factors. AB - In the 80's, three retrospective studies showed an inverse relation between fish consumption and ischemic heart disease (IHD) mortality. In parallel, fish fats containing the polyunsaturated fatty acid eicosapentaenoic (EPA) were shown to impair platelet aggregation and thromboxane formation. The results of the large prospective Diet and Reinfarction (DART) Study in the secondary prevention of myocardial infarction have further supported the possibility of potential interrelationships between diet and thrombogenic factors with respect to IHD. More recently these concepts have been confirmed and extended in the prospective Lyon Heart Study. However, in the latter in addition to changes in the content of EPA, changes in other well known variables (i.e. leukocytes and vitamin E) often abnormal in subjects prone to arterial thrombosis have been found. From studies on polygenic disorders, we have learned that by lowering the threshold and becoming a susceptibility gene, a polymorphism can lead to an effect assuming that it is present in the appropriate milieu. Nutrients have been shown to affect major determinants of myocardial ischemia such as fibrinogen or factor VII. However, the extent to what these and other hemostatic variables have been affected in studies devoted to dietary prevention of ischemia remains presently elusive. PMID- 10715835 TI - Nutrition and obesity: prevention and treatment. AB - The increased risk of morbidity and mortality from obesity, central body fat, and weight gain, and the beneficial effects of weight reduction argue that the cost associated with obesity could be beneficially affected by prevention of weight gain or induction of weight loss. Genetic, metabolic, and demographic predictors of weight gain have been identified that allow selection of high-risk individuals. Among the metabolic predictors are a low metabolic rate, insulin sensitivity, and a high respiratory quotient. Demographic predictors include current smokers, certain dieting behaviors, lower socio-economic class, a low level of education, use of contraceptives, status post-partum, and rapid weight gain in childhood. Several studies suggest that weight gain can be prevented. Targets for such strategies might be high-risk families, current smokers, those who are planning to stop smoking, and those with a low metabolic rate. For those who fail primary prevention, treatment may be appropriate. The greater the degree of excess weight, the greater the risk and the more appropriate treatment becomes to reduce body weight. PMID- 10715836 TI - Diabetes: nutrition in prevention and management. PMID- 10715837 TI - Nutrition and longevity: multifaceted improvements in population dietary patterns to end the cardiovascular disease epidemic. PMID- 10715838 TI - Cancer and diet. PMID- 10715839 TI - Nutritional education in the community. PMID- 10715840 TI - Nutrition and longevity: relevance to health transition in the developing countries. PMID- 10715841 TI - Summing up. Nutrition and longevity in industrialized countries. PMID- 10715842 TI - Apoptosis induction in HCT116 cells by cytochalasins isolated from the fungus Daldinia vernicosa. AB - We examined the induction of apoptosis by cytochalasin (cc) derivatives (1-14) isolated from the Japanese fungus Daldinia vernicosa to HCT116 human colon cancer cell line based on their cytotoxicity, DNA ladder and DNA fragmentation ratio in agarose gel electrophoresis, and morphological changes. Most cc derivatives tested here induced apoptosis. Particularly cytochalasin 1 (cc1), monoacetate of 1 (cc1Ac), and cc14 were the most potent apoptosis inducers. These apoptotic activities were stronger than that of cytochalasin D as a known apoptosis inducer in HCT116 cell. However, cc4 and cc12 induced necrosis. The structure-activity relationship including their cytotoxicity will be discussed. PMID- 10715843 TI - Anti-herpes simplex virus activities of crude water extracts of Thai medicinal plants. AB - A number of Thai medicinal plants, recommended as remedies for herpesvirus infection and have been used in primary health care were investigated for their intracellular activities against herpes simplex viruses (HSV). Centella asiatica L., Maclura cochinchinensis Cornor, and Mangifera indica L. contained both anti HSV-1 and -2 activities, as determined by plaque inhibition assay. An inhibition of the production of infectious HSV-2 virions from infected Vero cells could also be demonstrated. Combinations of each of these reconstituted extracts with 9-(2 hydroxyethoxymethyl) guanosine (acyclovir; ACV) resulted either in subadditive, additive, or synergistic interaction, against HSV-2, depending on the dose of ACV used; mixture of C. asiatica and M. indica exerted an additive effect in a similar assay. Furthermore, the inhibitory effects of these plant extracts were also substantiated by flow cytometric analysis of virus-specific antigens in the infected cells. The active constituent present in C. asiatica extract was determined to be asiaticoside while in M. indica was mangiferin. Thus, these data suggest therapeutic potential of these plant extracts. PMID- 10715845 TI - Orally administered Kampo (Japanese herbal) medicine, "Juzen-Taiho-To" modulates cytokine secretion in gut associated lymphoreticular tissues in mice. AB - The influence of the oral administration of Juzen-Taiho-To (JTT; Si-Quan-Da-Bu Tang in Chinese), a Kampo (Japanese herbal) medicine, on the cytokine production in mice were investigated. Lymphocytes from spleen (SPN), mesenteric lymph node (MLN) and Peyer's patches (PP) from mice, which received orally administered JTT for 2 weeks, were stimulated with concanavalin A (Con A), and the resulting conditioned medium was tested for cytokine production by ELISA. Administration of JTT caused enhancement of interferon g (IFN-gamma) and interleukin-4 (IL-4) production to some extent but decreased IL-5 production from the SPN. On the other hand, notable changes in cytokine production were observed in lymphocytes from MLN and PP. Administration of JTT increased IFN-gamma production remarkably, as well as IL-5 secretion from both MLN and PP, whereas IL-2 secretion was plainly reduced. The ratio of IFN-gamma and IL-4 was shifted to Th1 dominant in MLN and PP, however changed little in SPN. The flow cytometric analysis revealed that the population of CD3+, CD4+, CD45R/B220+, and CD45RBlowCD4+ cells in each lymphocyte was not changed significantly after oral administration of JTT. These findings demonstrate that the lymphocytes from gut associated lymphoreticular tissues (GALT) are more sensitive to produce cytokines on cytokine production than those from SPN by oral administration of JTT, and that the modulation of cytokine production may be due to functional change of lymphocytes but not change in lymphocytes population. PMID- 10715844 TI - Anti-herpes simplex virus component isolated from Maclura cochinchinensis. AB - The powerful anti-herpes simplex virus (HSV) activity of Maclura cochinchinensis in vitro prompted us to carry out biologically-guided separation of the active component(s). Ethyl acetate and methanol extracts exhibited anti-HSV-2 activity at EC50 values of 38.5 micrograms/ml and 50.8 micrograms/ml, respectively. Although petroleum ether extract was inactive, a chloroform extract was too toxic to the test cell culture to perform the test. Biologically-guided chromatographic separation of the ethyl acetate extract yielded compound A, identified as morin using a spectroscopic method. Morin exhibited anti-HSV-2 activity at an EC50 value of 53.5 micrograms/ml. Morin pentaacetate was synthetized, but was inactive. This result suggested that free hydroxyl groups are required for anti HSV-activity, as demonstrated previously for the antiviral activity of other flavonoids. PMID- 10715846 TI - The molluscicidal activity of plants used in Brazilian folk medicine. AB - In a continuing search for new compounds for the control of the vectors of schistosomiasis, we have tested the activity of some Brazilian medicinal plants as sources of molluscicidal natural compounds, using two molluscicidal bioassays. Twenty-seven crude extracts, from twenty-six species belonging to nineteen families, were tested. Seven extracts showed significant molluscicidal activity against Biomphalaria glabrata adults with DL50 values of less than 50 ppm, and five of them were very active in the test using egg masses. The species most active against B. glabrata adults (LD50 value = 3.65 ppm) and their egg masses (LD50 value = 0.13 ppm) was Derris sp. Annona muricata [LD50 value (adult) = 11.86 ppm and LD50 value (egg) = 49.62 ppm], Jatropha elliptica (from Goias state) [LD50 value (adult) = 24.80 ppm and LD50 value (egg) = 3.03 ppm] and Renealmia exaltata [LD50 value (adult) = 28.03 ppm and LD50 value (egg) = 21.67 ppm], were also considered promising molluscicidal plants. PMID- 10715848 TI - Screening Brazilian plant species for in vitro inhibition of 5-lipoxygenase. AB - Plants from the Brazilian flora were evaluated for the inhibition of 5 lipoxygenase. The species were selected based on their traditional use and on a chemosystematic approach. In total, 19 species belonging to 13 families have been investigated. Hedychium coronarium J. Koenig (Zingiberaceae), Xylopia frutescens Aubl. (Annonaceae) and Hymenaea courbaril L. (Leguminosae) presented a high 5 lipoxygenase inhibitory activity. Some hypothesis about the nature of the active compounds are discussed, based on reports of the chemical constitution of these species or other species from the same botanical family. PMID- 10715847 TI - Protective effects of Glycyrrhizae radix extract and its compounds in a renal hypoxia (ischemia)-reoxygenation (reperfusion) model. AB - Glycyrrhizae radix water extract (GRWE) and its two major constituents glycyrrhizin and 3-glycyrrhetinic monodesmoside, significantly suppressed LDH leakage and MDA release, whereas glycyrrhetinic acid had no effect. On the other hand, in rats subjected to ischemia-reperfusion, the activities of endogenous antioxidant enzymes including catalase and glutathione peroxidase showed recovery, whereas the levels of urea nitrogen and creatinine in serum were reduced by administration of glycyrrhizin orally for 30 days prior to ischemia reperfusion. These results indicate that GRWE and its two constituents may be promising for amelioration of hypoxia (ischemia)-reoxygenation (reperfusion) injury and improvement of renal function by acting directly or indirectly as antioxidant and oxygen radical-scavenging agents. PMID- 10715849 TI - Direct scavenging of nitric oxide by traditional crude drugs. AB - Thirty-one traditional crude drugs and several pure compounds were examined for their possible regulatory effect on nitric oxide (NO) levels using sodium nitroprusside as a NO donor in vitro. Most of the crude drugs tested demonstrated direct scavenging of NO. Eight crude drugs, including Sanguisorbae Radix, Caryophylli Flos, Gambir, Coptidis Rhizoma, Granati Cortex, Gallae Rhois, Rhei Rhizoma and Cinnamomi Cortex exhibited significant activity (IC50 values < 1000 micrograms/ml), and with the exception of Coptidis Rhizoma, all were found to contain tannins as their major constituents. In addition, some crude drugs containing flavonoids or essential oils also appeared to act against NO. Ten major tannins contained in Sanguisorbae Radix and Rhei Rhizoma showed high scavenging activity (IC50 values < 326.3 micrograms/ml), and 6 of 8 alkaloids obtained from Coptidis Rhizoma also effectively scavenged the NO radical (IC50 values < 455.4 micrograms/ml). It was indicated that these compounds may be the active principles of the crude drugs responsible for NO scavenging. The present results suggest that traditional crude drugs might be potent and novel therapeutic agents for scavenging of NO and the regulation of pathological conditions caused by excessive NO and its oxidation product, peroxynitrite. These findings may also help to explain, at least in part, certain pharmacological activities of crude drugs, especially anti-infection and anti-inflammatory activities. PMID- 10715850 TI - Antihyperglycemic activity of Teramnus labialis (Fabaceae). AB - In vivo bioassay-guided fractionation of the aqueous alcohol extract of the aerial parts of Teramnus labialis (Roxb.) Benth. (Fabaceae), using C57BL/Ks-db/db mice as a model for type 2 diabetes, yielded an active fraction containing a mixture of coumarins. The major coumarin present in the active fraction was identified as fraxidin. PMID- 10715851 TI - Physicochemical properties of harpagoside and its in vitro release from Harpagophytum procumbens extract tablets. AB - The objective of this investigation was to characterize the active-component harpagoside of Harpagophytum extract from a physico-chemical perspective and to determine its in-vitro release from tablets according to DAB 1996. It was found that both pure harpagoside and harpagoside in Harpagophytum extract have an octanol-water distribution coefficient of approximately 4 which is neither dependent on temperature nor on pH. The mean harpagoside content in Harpagophytum tablets of Batch 9102 was 16.4 mg (S.D. 0.2; S.E. 0.03). Related to a tablet weight of 365 mg (100%), this corresponds to a haragoside content of 4.5% (S.D. 0.049; S.E. 0.006). On average the tablets disintegrate after 18 +/- 3 minutes (mean +/- SD). The tablets taken from Batch 9102 released the active component harpagoside well, with a t50 of 13.5 min, a t90 of 23 min and a t95 of 25 min in relation to 16.5 mg of harpagoside per dose. Harpagoside content decreased by about 10% in artificial gastric fluid within a period of 3 hours and remained stable in artificial intestinal fluid for a period of 6 hours. PMID- 10715852 TI - Arnebins and antimicrobial activities of Arnebia hispidissima DC. cell cultures. AB - The whole plant of Arnebia hispidissima DC. (Boraginaceae) is used for the treatment of tongue and throat ailments in Indian traditional medicine. The present paper deals with the plants phytochemical constituents, the arnebins, and antimicrobial activities of its root extract. The antimicrobial activities were tested against Gram-positive and Gram-negative bacteria and fungi. The crude hexane extract demonstrated a potent antimicrobial effect against bacteria and a mild effect against fungi. Likewise, the hexane extract of cell cultures of A. hispidissima also showed mild bioefficacy against the select microorganisms. PMID- 10715853 TI - Bringing treatment by halves to an end. PMID- 10715854 TI - A mother with bruising and a black eye. PMID- 10715855 TI - Key developments in psychiatry. PMID- 10715856 TI - Patients who experience traumatic stress. PMID- 10715857 TI - Effective brief interventions in problem drinking. PMID- 10715858 TI - Managing myocarditis. PMID- 10715859 TI - Allergic rhinitis: not just a runny nose. PMID- 10715860 TI - Recurrent vaginal thrush and soreness. PMID- 10715861 TI - Constipation, soiling and encopresis. PMID- 10715862 TI - Hyperactivity: a rational strategy. PMID- 10715863 TI - Early diagnosis of child cancer. PMID- 10715864 TI - Diagnosis and treatment of parvovirus B19. PMID- 10715865 TI - Managing the clumsy and non-reading child. PMID- 10715866 TI - A guide to open-access echocardiography. PMID- 10715867 TI - Evidence-based medicine in practice. PMID- 10715868 TI - A troubled colleague who asks for help. PMID- 10715869 TI - Key developments in unstable angina. PMID- 10715870 TI - Evidence in cardiovascular medicine. PMID- 10715871 TI - Post-MI care in general practice. PMID- 10715872 TI - Valvular disease: the GP's key role. PMID- 10715873 TI - Atrial fibrillation in general practice. PMID- 10715874 TI - Oral cancer: the importance of early referral. PMID- 10715875 TI - The video assessment. PMID- 10715876 TI - The real dangers of too much sun. PMID- 10715877 TI - A depressed man requesting St John's wort. PMID- 10715878 TI - Key developments in dermatology. PMID- 10715879 TI - Managing fungal infections. PMID- 10715880 TI - Allergic drug reactions. PMID- 10715881 TI - Skin presentations: refer or reassure? PMID- 10715882 TI - The science behind head lice treatment. PMID- 10715883 TI - Diagnosing and treating exercise-induced asthma. PMID- 10715884 TI - A rational approach to measuring BP. PMID- 10715885 TI - Tackling the written paper 1. PMID- 10715887 TI - Using of thin-layer chromatography for identification and quantitative determination of benzalkonium chloride in eye drops. AB - Conditions for thin-layer chromatographic separation of components (therapeutic and auxiliary substances) in selected eye drops have been established and next used for the densitometric determination of benzalkonium chloride content in the drugs. PMID- 10715886 TI - Quantitative determination of selected tricyclic biological active compounds by using capillary electrophoresis. AB - Five tricyclic antidepressants: imipramine hydrochloride, noxiptilin hydrochloride, hydroxyzine dihydrochloride, diethiazine hydrochloride and amitryptyline hydrochloride were determined by the capillary electrophoresis method. The determinations were performed by using a fused sillica capillary (60 cm x 50 microns i.d., effective length 53 cm). Acetate buffer (25 mM ammonium acetate and 1 mM acetic acid) in (1:1) methanol:water solution was used as an electrolyte. The determination voltage was 20 kV. The UV-detector at 214 nm was applied. PMID- 10715888 TI - The application of VIS spectrophotometric determination of enalapril maleate in substance, in tablets and estimation of ester group stability. AB - A new spectrophotometric VIS method is proposed for the determination of enalapril maleate in pure substance and in tablets. Attempts have been made to estimate stability of the ester group in the molecule of enalapril maleate in the solid phase at 70 degrees C. PMID- 10715889 TI - The effect of dexamethasone pretreatment on chlordiazepoxide and oxazepam levels in rat plasma. AB - The effect of dexamethasone pretreatment on the level of chlordiazepoxide and oxazepam was examined in the plasma of rats administered with 80 mg/kg of chlorodiazepoxide per os. The concentration of free chlordiazepoxide and unconjugated oxazepam was determined by HPLC method after solid-phase extraction. Pretreatment with dexamethasone significantly increased chlordiazepoxide plasma level 3, 6 and 12 hrs after ingestion, however increase of oxazepam concentration was not statistically significant. AUC 1-24 h after dexamethasone was elevated, for chlordiazepoxide and for oxazepam. PMID- 10715890 TI - The investigations into the interferon-like activity of Polygonum L. genus. AB - The interferon inducing abilities of eleven ethanolic extracts obtained from nine taxons of Polygonum L. genus, were tested in the cell culture of monkey kidney. The extracts from the herb of Polygonum amphibium L. and the rhizome and fruit of Polygonum bistorta L. induced a substance showing an interferon-like activity. Biological activity studies showed that the protective titre (the highest dilution which protected cells by 50% against virus infection) of the interferon like materials was 1:10-1:15. PMID- 10715891 TI - Comparative study of antithrombotic and antiaggregatory activity of acetylsalicylic acid, ticlopidine and a new noncarboxylic acid antiinflammatory pyrazine derivative HF90. AB - The action of acetylsalicylic acid, ticlopidine and a new pyrazine derivative HF90 selected in preliminary screenings (11, 18, 19) was studied by using the mouse antithrombotic assay according to DiMinno and Silver (22) and in vitro blood platelet aggregation method according to Born (23). Acute pulmonary thromboembolism was induced by injection of a mixture of collagen and epinephrine into the mouse tail vein. The effect of HF90, an acidic pyrazine derivative possessing active methylene moiety, administered at doses of 50 and 100 mg/kg, was compared to the action of the well established antithrombotic agents: ticlopidine (100 mg/kg) and acetylsalicylic acid (20 mg/kg). The compounds were administered i.p. in single doses 1 h and 24 h before the thrombotic challenge or once a day per three consecutive days before the thrombotic challenge. Ticlopidine appeared to provide the better protection against microembolism than acetylsalicylic acid although its effect has not manifested itself immediately after administration. The pyrazine derivative examined has a lower but significant antithrombotic activity. The chemical class of pyrazine derivatives with active methylene moiety (the so called pyrazine CH/NH-acids) (16) provides a new original antiinflammatory pharmacophore and HF90 may serve as the "lead compound" in the search for new agents of pharmacological interest. PMID- 10715892 TI - Can magnesium sulfate reduce the risk of cerebral injury after perinatal asphyxia? AB - Hypoxia-ischemia produces brain damage by processes that continue for many hours after reoxygenation/reperfusion. This provides a window of opportunity for therapy aimed at preventing further loss of brain cells. Sulfate magnesium can prevent posthypoxic brain injury by blocking glutamate receptors within the calcium (Ca++) ion channel. We used sulfate magnesium in nine newborn infant after perinatal hypoxia. We investigated the brain damage, by ultrasound examination, on third day, in first, second and third week, and third, sixth month of life. We have estimated the neurological development in the first week of life and third and twelfth month of life. We did not find deviations in ultrasound examination. We did not observe convulsions. We did not observe any side effect of this therapy. The examination at 1 of year of life in all of children was correct. PMID- 10715893 TI - Morphology of lymphatic malformations: a pictorial review. AB - A pictorial review of the many clinical manifestations of lymphatic malformations is presented, changes to the terminology of lymphatic malformations are reviewed and the 'pros and cons' of relevant investigative techniques are discussed. PMID- 10715894 TI - Australian teledermatology: the patient, the doctor and their government. AB - Telemedicine is an emerging technology within Australia. We review the historical development of telemedicine and discuss the clinical and non-clinical issues surrounding its practice in this country. Teledermatology is one application of telemedicine. We discuss the potential impact of teledermatology on patients, doctors and third parties such as government. So far, teledermatology has received little attention from Australian dermatologists. By contrast, the Government and other organizations are showing keen interest in establishing infrastructure within this country. We believe it is time for dermatologists to become more involved in the practice and politics of telemedicine within Australia. PMID- 10715895 TI - New approaches to the laser treatment of vascular lesions. AB - The pulsed dye laser (PDL) was developed based on the concept of selective photothermolysis. Using a wavelength of light well absorbed by the target and a pulse duration short enough to spatially confine thermal injury, specific vascular injury could be produced. While the PDL revolutionized the treatment of port wine stains (PWS) and a variety of other vascular lesions, the mathematical model predicted that the ideal thermal relaxation time for the vessels in PWS is actually 1-10 msec, not 450 microseconds. These original theoretical calculations have been proved correct recently in a study using both an animal vessel model and in human PWS. Longer wavelengths of light within the visible spectrum penetrate deeper into the skin and are more suitable for deeper vessels; while longer pulse duration is required for larger calibre vessels. A variety of lasers have been developed recently for the treatment of vascular lesions that incorporate these concepts into their design, including PDL at 1.5 msec, a filtered flash-lamp pulsed light source with pulse durations of 1-20 msec, several 532 nm pulsed lasers with pulse durations of 1 to as high as 100 msec, long pulsed alexandrite lasers at 755 nm with pulse durations up to 20 msec, pulsed diode lasers in the 800-900 nm range, and long pulsed 1064 Nd:YAG sources. Preliminary results are encouraging. PMID- 10715896 TI - Curettage, electrosurgery and skin cancer. AB - The scientific literature is replete with reports extolling the virtues of curettage and electrosurgery in the treatment of skin disease. Published cure rates for selected skin cancers consistently equal those for other treatment modalities, including scalpel excision. Despite this, curettage is often overlooked as a first line treatment for skin cancer. We review the evidence based literature for patient selection criteria and curettage and electrosurgery techniques. PMID- 10715897 TI - Toxic epidermal necrolysis in Western Australia. AB - Toxic epidermal necrolysis is a severe, life-threatening illness with up to one third mortality. We report a retrospective analysis of all cases treated in Royal Perth Hospital over a 20-year period from July 1978 to June 1998, by analysis of medical records. A total of 12 patients with an age range of 23-73 years was identified. The female to male ratio was 2:1, with age of onset earlier in females. All cases were associated with medications, most commonly antibiotics, anticonvulsants and allopurinol. The mortality rate was one-third (four deaths), mostly resulting from cardiorespiratory failure, renal failure and sepsis. Risk factors for death were advanced age and severe underlying disease, including diabetes, alcoholic liver disease sepsis and malignancy. Among the six patients treated with systemic corticosteroids there was only one death. Treatment with corticosteroids appeared to be beneficial, with such patients having both fewer complications and a lower mortality rate. PMID- 10715898 TI - Topical applications and perioral dermatitis. AB - Perioral dermatitis (POD) is a common dermatosis and is considered by most dermatologists to be increasing in incidence. An Australia-wide, questionnaire based study investigating the aetiology of POD was conducted, with particular emphasis on the significance of cosmetic use. One hundred and thirty-three cases were obtained from dermatologists across Australia and were compared with 99 randomly selected controls who were matched for age and sex. Application of foundation, in addition to moisturizer and night cream resulted in a 13-fold increased risk for POD (odds ratio 13.5; P < 0.001). The combination of moisturizer and foundation was associated with a lesser but significantly increased risk for POD (odds ratio 2.9; P = 0.017). Moisturizer alone was not associated with an increased risk of POD. These findings suggest that cosmetic preparations play a vital role in the aetiology of POD, perhaps by an occlusive mechanism. In 83% of cases, topical steroid application to the face occurred after the development of a facial rash. PMID- 10715899 TI - Nuchal fibroma associated with scleredema, diabetes mellitus and organic solvent exposure. AB - A case of scleredema diabeticorum of Buschke associated with nuchal fibroma and organic solvent exposure is reported. The patient presented with a neck mass causing discomfort and restriction of movement. Histological examination showed this to be a nuchal fibroma. Additionally, there was widespread induration of the skin of his trunk which was asymptomatic. A biopsy showed features of scleredema. This is the first reported association of these two conditions, both of which show increased and thickened collagen bundles without significant fibroblast proliferation. They differ by the occurrence of mucin in scleredema, although this is not always demonstrable, particularly in late lesions. The possibility that nuchal fibroma is an end stage, localized form of scleredema is canvassed. The patient's medical history included insulin-dependent diabetes mellitus with complications of retinal vessel thrombosis and peripheral neuropathy. The patient also had significant past exposure to a wide variety of chemicals, including organic solvents. PMID- 10715900 TI - Acute generalized exanthematous pustulosis associated with oral terbinafine. AB - A case history of acute generalized exanthematous pustulosis (AGEP) following oral terbinafine is reported. A 64-year-old woman presented with a rapidly spreading micropustular eruption 3 days after completing a 28-day course of oral terbinafine. There was a positive family history of psoriasis but no personal history. The clinical presentation and histopathology were consistent with AGEP. There was nearly complete resolution of the pustular eruption within 3.5 weeks following cessation of oral terbinafine and treatment with topical and systemic corticosteroids. The patient has remained free of any recurrence 18 months later. A summary of drugs known to be associated with AGEP is presented. Prompt recognition of AGEP is stressed in order to avoid confusion with acute generalized pustular psoriasis or a systemic infection. The most important aspect of management is the immediate withdrawal of the suspect drug. PMID- 10715901 TI - Tungiasis. AB - A 24-year-old man developed slow-growing lesions on subungual and plantar areas that appeared a few weeks after returning from a trip to South America. The diagnosis of tungiasis was established by microscopic examination of a lesion. Tungiasis is rarely seen in non-endemic areas. PMID- 10715902 TI - Follicular B-cell pseudolymphoma. AB - A 60-year-old woman presented with a 3-week history of a pruritic papulo-nodular eruption on the face and trunk after a bee sting. Histological examination showed a predominantly lymphocytic infiltrate with follicular centres and tingible body macrophages. Immunohistochemically, positive staining for both kappa and lambda light chains was noted. The eruption settled with oral antihistamine and topical corticosteroid. These findings support the diagnosis of follicular B-cell pseudolymphoma. PMID- 10715904 TI - Reconstruction of the nasal tip using a nasalis myocutaneous flap. AB - Reconstruction of nasal tip defects is a technical and aesthetic challenge for dermatological surgeons, due to the limited reservoir of available skin and its thick, inflexible nature. The myocutaneous flap described in this paper, using the vascular pedicle of the transverse nasalis muscle, is an excellent option for repair of defects of the nasal tip. It provides a cosmetically superior result to skin grafting, particularly in noses of 'sebaceous' quality, by providing similar local skin. The flap maintains normal nasal tip contour and, being an axial pattern flap, its blood supply is reliable. PMID- 10715903 TI - Allergy to lichen acids in a fragrance. AB - A 48-year-old clerical officer with a recurrent facial eruption had positive patch test reactions to nickel, fragrance mix and lichen acid mix. On testing to individual ingredients of fragrance mix and lichen acid mix, she had 2+ reactions to oak moss, which is thought to be the main allergen in fragrance mix, and to usnic acid, which is one of a number of lichen acids comprising oak moss. Avoidance of fragrance use resulted in clearing of the eruption but, subsequently, an acute vesicular flare on her face and hands occurred after exposure to lichen on garden shrubs. PMID- 10715905 TI - Reversible painful gynaecomastia induced by low dose finasteride (1 mg/day) PMID- 10715906 TI - The prevalence of hepatitis C in a community-based population, Ontario, 1996. PMID- 10715907 TI - Measles surveillance: guidelines for laboratory support. PMID- 10715908 TI - [The effect of umbilical cord blood cytokines on clonogenicity of hemopoietic stem cells isolated from umbilical cord blood]. AB - Human umbilical cord blood contains haematopoietic stem cells, which are a potential source of cells for hematopoietic transplants. Early cord blood hematopoietic cells are influenced by so called proinflammatory cytokines, which are present in cord blood serum. In this study we tried to correlate the concentration of these cytokines with the number, viability and clonogenicity of cord blood mononuclear cells. Accordingly, cord blood samples were harvested by employing an "open" collection method. Subsequently, we measured in those samples the concentration of selected pro inflammatory cytokines (Il-1 alpha, Il-1 beta, Il-6, Il-8 and TNF alpha), number of mononuclear cells and evaluated in vitro clonogenicity of myeloid progenitors (CFU-GM). We found the negative correlation between number of mononuclear cells and concentration of TNF alpha, and between number of detectable CFU-GM and concentration of IL-1 beta. Other cytokines, which were studied in this report did not correlate with evaluated parameters. PMID- 10715909 TI - [The activities of some glycosaminoglycan degrading enzymes in the wall of the umbilical cord vein and their alteration in EPH-gestosis]. AB - EPH-gestosis evokes great connective tissue rebuilding in the wall of umbilical cord arteries and Wharton's jelly. Not much is known about umbilical cord vein maintenance. For better understanding of processes proceeding in it we decided to determine activities of some glycosaminoglycan-degrading enzymes. It was found that specific activities of some neutral endoglycosidases and exoglycosidases decreased during EPH-gestosis. Acidic endoglycosidases and sulphatases specific activities did not change. Those results with no changes in content and proportional relationship of umbilical cord vein glycosaminoglycans suggest that EPH-gestosis disturbs markedly those connective tissue element metabolism. PMID- 10715910 TI - [Using of Misoprostol for preinduction and induction of labor in term pregnancy]. AB - Labour induction is frequently indicated in women with an unfavourable cervix. Oxytocin and prostaglandins are the most common drugs used for labour induction. Induction of labour with prostaglandins offers the advantage of promoting both cervical ripening and myometrial contractility. The purpose of the study was to evaluate the safety and efficacy intravaginal administration prostaglandin E1 methyl analogue, misoprostol in cervical ripening and induction of labour in term pregnancy and in women with unfavourable cervix (Bishop score < = 4). The approval for this study was given by the Board of Medical Ethics, University Medical School of Lublin in Poland. MATERIALS AND METHODS: 64 women with indication for termination of pregnancy received either misoprostol (Cytotec Searle) vaginally (group M, n = 30) or intravenous drip infusion of oxytocin(group Ox, n = 34). We evaluated profile of the studied women (gravidity, weight, height, maternal age, gestational age), effectiveness and safety of the misoprostol and need for oxytocin administration in group M, start of induction to-active labour interval (contractions), start of induction-to-vaginal delivery interval, hyperstimulation syndrome, delivery within 24 hours of drug application and caesarean section rate. Before starting labour induction a Bishop score was obtained. Statistical analysis was performed. Baseline and outcome variables were tested with student's t-test and c2 analysis. We needed p < 0.05 for statistical significance. RESULTS: There were no differences in the patient profiles (gravidity, weight, height, maternal age, gestational age) between groups except the score of cervical ripening. The Bishop score before induction was lower in group M. The interval between the initiation of induction to active labour was shorter in the misoprostol group (334.23 +/- 126.35 versus 610.00 +/- 352.14 minutes). The mean time between the initiation of induction to delivery was shorter in group M (707.69 +/- 341.15 +/- versus 1025.77 +/- 369.16 minutes). These differences were statistically significant. 28 (93.33%) patients in the misoprostol group delivered within 24 hours compared with 24 (70.59%) women in the oxytocin group. 8 patients in the misoprostol group and 8 patient in the oxytocin group had caesarean section. Labor induction was successful in 30 (100%) women in the misoprostol group compared with 24 (70.59%) patients in the group Ox. CONCLUSIONS: Intravaginal misoprostol is an effective, easy to use and cheap drug for the induction of labour, especially for cervical ripening in women with unfavourable cervix (Bishop score < = 4). PMID- 10715911 TI - [The usefulness of color doppler placenta mapping in the prediction of the condition of neonate in pregnancies complicated by intrauterine growth retardation]. AB - OBJECTIVE: The aim of the study was to assess usefulness of placenta mapping as a screening method for early detection of the state of emergency of fetus and neonate in pregnancies complicated with intrauterine growth retardation (IUGR). MATERIALS AND METHODS: We investigated 48 pregnant women with intrauterine growth retardation detected by ultrasound screening. RESULTS: Color Doppler placenta mapping as a method of early detection of intrauterine growth retardation has sufficient sensitiveness and specificness. CONCLUSIONS: Changes arising in spiral arteries of the placenta cause vanishing of Color Doppler signal in the area of intervillous circulation. Lack of Doppler signal from spiral arteries correlates with bad condition of neonate after delivery. PMID- 10715912 TI - [The use of Gynipral (hexoprenaline) in suppression of uterus contractions]. AB - OBJECTIVE: The aim of of this study was to estimate the effectiveness and tolerance of Gynipral (hexoprenaline)-beta-2 sympathomimetic used as oral and intravenous tocolytic. MATERIAL AND METHODS: We analysed 110 pregnant women admitted to the hospital with premature uterus contractions between 17 and 37 week of pregnancy. Pregnant women were divided into two groups. Group I--86 patients with regular uterus contractions. In this group Gynipral was administered to suppress uterus contractions. Group two consisted of 24 patients without regular uterus contractions. In this group women were treated with Gynipral to keep pregnant uterus in maximum relaxation in such cases as IUGR, oligohydramnion, after Strassman procedure before pregnancy or cerclage procedure in current pregnancy. Group I was treated with intravenous Gynipral infusion and after suppression of uterus contractions the way of administration was changed into oral. Group II from the beginning was treated with oral tocolysis with Gynipral. Pregnant women were continually under CTG control and all ailments such as tachycardia, flapping tremor and anxiety were analysed and noted. RESULTS: Gynipral has effectively suppressed premature uterus contractions in 80 cases in group I (93%) and in 24 cases in group II (100%). In 106 cases (96%) patients have not presented any side effects. CONCLUSIONS: Gynipral is an effective tocolytic successfully used in premature labor treatment. Gynipral has a good tolerance administered both intravenously and orally as well and in most cases there was no need to add antiarrhythmic drugs to reduce side effects. PMID- 10715913 TI - [Giant placental angioma with polyhydramnios, high level of alpha-fetoprotein and neonatal congenital lactic acidosis]. AB - Placental tumors are rare in pregnancy. They cause nonspecific complications such as polyhydramnios, fetal anemia, fetal thrombocytopenia and cardiac decompensation with non-immunological hydrops fetalis. In the presented case a very large placental hemangioma was connected with polyhydramnios, premature delivery, fetal anemia and thrombocytopenia, maternal serum alpha fetoprotein elevation and congenital lactic acidosis. After delivery a severe state of the newborn was caused by oligovolemic shock. In the course of the disease the neonatal state steadily deteriorated mainly because of sepsis, cerebral hemorrhage and metabolic acidosis despite adequate therapy. The organic acids assessment in the blood serum of a newborn child showed a very strong signal of lactic acid and an increase in parahydroksyfenylolactic acid. Postmortal examination confirmed prematurity, respiratory distress syndrome, sepsis and cerebral hemorrhage. These symptoms probably resulted from the presence of a placental tumor of considerable size of 12 cm and congenital lactic acidosis, which to our knowledge, have not been described in the available literature until now. In conclusion it should be underlined, that there exists difficult to explain relationship between the presence of a placental haemangioma and severe metabolic changes resulting in high mortality and morbidity of the newborn. PMID- 10715914 TI - [Pregnancy and delivery after liver transplantation: case report]. AB - We report a case of successful pregnancy in a 22-year-old primigravida who became pregnant 19 months after orthotopic liver transplantation for end-stage liver dysfunction caused by Wilson disease. The mother continued immunosuppressive therapy (tacrolimus) during the pregnancy. No complications were observed. Labour started spontaneously at 40th week's gestation. As a result the healthy male infant weighing 3900 g was born. Patient and the baby were discharged from hospital, both in good condition, on the fifth day after delivery. PMID- 10715915 TI - [Problems of contraception in women with Wilson disease]. AB - Because of the constant progress in medical therapy, the fertility status of patients with Wilson's disease may be preserved. Contraceptive counselling is, therefore, essential to these women. Some contraceptive methods can have a negative influence on hepatic function or hepatic insufficiency may reduce the efficacy of other contraceptives. Different contraceptive options for patients with Wilson's disease are described. Second generation intrauterine devices with copper and estrogen-containing oral contraceptives are contraindicated. Spermatocide and barrier contraceptives and progesterone-only preparations can be safely used. PMID- 10715916 TI - Managing pain. PMID- 10715917 TI - Fraud the old-fashioned way. PMID- 10715918 TI - Chronic TMD. PMID- 10715919 TI - Access and competency. PMID- 10715920 TI - Study shows link between antidepressants, bruxism. PMID- 10715921 TI - Smoking prevention programs for students should stress social skills. PMID- 10715922 TI - The impact of tissue engineering on dentistry. AB - BACKGROUND: Tissue engineering is a novel and highly exciting field of research that aims to repair damaged tissues as well as create replacement (bioartificial) organs. OVERVIEW: The authors provide a general review of the principles underlying key tissue engineering strategies, as well as the typical components used. Several examples of preclinical and clinical progress are presented. These include passive approaches, such as dental implants, and inductive approaches that activate cells with specific molecular signals. PRACTICE IMPLICATIONS: Tissue engineering will have a considerable effect on dental practice during the next 25 years. The greatest effects will likely be related to the repair and replacement of mineralized tissues, the promotion of oral wound healing and the use of gene transfer adjunctively. PMID- 10715923 TI - Postoperative pulpal and repair responses. AB - BACKGROUND: Each year in the United States, the success of 10 million surgically restored carious lesions depends on a favorable tertiary dentin repair response to preparation, restoration and patient factor variables. The authors investigated the relationship between these variables and dentinal response. METHODS: Standardized rectangular Class V restoration preparations were cut into the buccal dentin of intact first or second premolars of 27 patients without exposing the pulp and were restored. The patients were between 9 and 17 years of age. The treated teeth were scheduled for extraction for orthodontic reasons. After tooth extraction, the tertiary dentin was analyzed histomorphometrically. RESULTS: The area of tertiary reactionary dentin was found to be correlated using linear regression analysis of variance with restoration residual dentin thickness (P = .0024), age of the patient (P = .0045), restoration floor surface area (P = .0266) and restoration width (P = .0415). The authors did not find a correlation with the premolar position (P = .0594), sex of the patient (P = .650), pulpal inflammatory reaction (P = .613) or the time elapsed since surgery (P = .531). Restoration with zinc oxide eugenol was found to negatively influence tertiary dentin matrix secretion (post hoc analysis of variance, P = .030). CONCLUSIONS: The age of a patient at treatment, the choice of restorative material and the size of the restoration preparation are all factors that can positively or negatively affect the pulpal repair response. CLINICAL IMPLICATIONS: Age of the patient affects dentin repair capacity and may be a factor in treatment planning decisions. Minimizing the cutting of dentin, especially the width and base of the preparation, reduces the probability of recurrent pulpal complications. PMID- 10715924 TI - Does low-dose aspirin therapy complicate oral surgical procedures? AB - BACKGROUND: The fear of uncontrolled bleeding often prompts medical practitioners to stop aspirin intake for seven to 10 days before any surgical procedure. The authors initiated this study to evaluate the effect of aspirin on bleeding in patients undergoing oral surgery. METHODS: The study group consisted of 39 patients who were scheduled to undergo dental extractions. All patients were receiving 100 milligrams of aspirin daily on a regular basis. The authors randomly divided the patients into two groups: those who stopped the aspirin therapy before the procedure and those who continued the aspirin therapy. One hour before the procedures, all patients underwent a bleeding time test. In addition, the amount of bleeding during the procedure was measured. RESULTS: The mean (+/- standard deviation) bleeding time was 1.8 +/- 0.47 minutes for patients who stopped aspirin therapy one week before the procedure. For patients who continued aspirin therapy, the bleeding time was 3.1 +/- 0.65 minutes. The difference was statistically significant (P = .004). However, both groups were within the normal bleeding time range, and in both groups, a local hemostatic method was sufficient to control bleeding. No episodes of uncontrolled intraoperative or postoperative bleeding were noted. CONCLUSION: Low-dose aspirin therapy should not be stopped before oral surgery. Local hemostasis is sufficient to control bleeding. CLINICAL IMPLICATIONS: Patients receiving aspirin therapy to prevent blood clot formation may be subject to emboli formation if the treatment is stopped. The results of this study show that aspirin therapy should be continued throughout oral surgical procedures. Local measures are sufficient to control any bleeding during surgery. PMID- 10715925 TI - Efficacy of preformed metal crowns vs. amalgam restorations in primary molars: a systematic review. AB - BACKGROUND: The authors evaluated the treatment efficacy of preformed metal crowns, or PMCs, vs. amalgam restorations in primary molars by means of a literature review and meta-analysis. TYPES OF STUDIES REVIEWED: From a literature search, the authors selected clinical studies that evaluated treatment with PMCs vs. amalgam control restorations in primary molars and provided data against which treatment outcomes could be compared. RESULTS: Ten studies with durations ranging from 1.6 to 10 years fulfilled the selection criteria. Their failure rates, based on need for subsequent treatment or retention of the restoration at final evaluation, ranged from 1.9 to 30.3 percent for PMCs and 11.6 to 88.7 percent for amalgam restorations. Overall, PMCs demonstrated greater longevity and reduced retreatment need compared with amalgam control restorations. The odds ratio for all studies fell within the boundary favoring treatment with PMCs. CLINICAL IMPLICATIONS: Analysis of the literature, though mainly retrospective studies, demonstrated evidence of a more favorable outcome for PMCs than for amalgam restorations in primary molars requiring multisurface restorations. PMID- 10715927 TI - Implants and general practitioners. PMID- 10715926 TI - Oral manifestations of primary immunological diseases. AB - BACKGROUND: Primary immunodeficiencies have many oral manifestations. The clinical presentation of these diseases demonstrates the roles of different immune cells for the maintenance of oral health. METHODS: The authors reviewed selected literature describing systemic and oral manifestations of the primary immunodeficiencies published between 1966 and 1999. RESULTS: The authors found that oral candidiasis and herpetic infections are seen frequently in patients with T-cell deficiencies, while patients with B-cell deficiencies are most susceptible to bacterial infections. Periodontitis and oral candidiasis are found in some, but not all, phagocyte deficiencies. CONCLUSIONS: These findings demonstrate that T cells, B cells and phagocytes all have roles in oral immunity. CLINICAL IMPLICATIONS: Acquired conditions that affect the immune system such as diabetes, alcoholism and acquired immunodeficiency syndrome, as well as certain medications, will affect oral defense mechanisms. The effects that acquired immunodeficiencies will have on oral health can be predicted from the oral manifestations of primary immunodeficiencies. PMID- 10715928 TI - The body's skin frontier and the challenges of wound healing: keloids. PMID- 10715929 TI - Antibiotic prophylaxis in dentistry: a review and practice recommendations. AB - BACKGROUND: The American Heart Association, or AHA, and the American Dental Association recently changed their recommended protocols for antibiotic prophylaxis against bacterial endocarditis. A new recommendation also has been issued by the ADA and the American Academy of Orthopaedic Surgeons, or AAOS, against routine antibiotic prophylaxis in patients with prosthetic joint replacements. These changes reflect increasing scientific evidence and professional experience in opposition to widespread use of antibiotic prophylaxis in these specific situations and others faced in dentistry. METHODS: The authors reviewed the medical and dental literature for scientific evidence regarding the use of antibiotics to prevent local and systemic infections associated with dental treatment. Situations commonly considered by dentists for potential use of prophylactic antibiotics were reviewed to determine current evidence with regard to use of antimicrobial agents. This included prevention of distant spread of oral organisms to susceptible sites elsewhere in the body and the reduction of local infections associated with oral procedures. RESULTS: There are relatively few situations in which antibiotic prophylaxis is indicated. Aside from the clearly defined instances of endocarditis and late prosthetic joint infections, there is no consensus among experts on the need for prophylaxis. There is wide variation in recommended protocols, but little scientific basis for the recommendations. The emerging trend seems to be to avoid the prophylactic use of antibiotics in conjunction with dental treatment unless there is a clear indication. CONCLUSIONS: Aside from the specific situations described, there is little or no scientific basis for the use of antibiotic prophylaxis in dentistry. The risk of inappropriate used of antibiotics and widespread antibiotic resistance appear to be far more important than any possible perceived benefit. CLINICAL IMPLICATIONS: Dentists are wise to use antibiotic prophylaxis in only those specific situations in which there is a valid scientific basis for it. Whenever possible, dentists should follow the standard protocols recommended by the ADA, AHA or AAOS. PMID- 10715930 TI - The new posterior resins and a simplified placement technique. AB - BACKGROUND: New heavy-body (packable) composites have been developed for use in posterior direct resin restorations. These materials are promoted as having better handling characteristics and higher physical properties than previous microhybrid composites. METHODS: The authors describe an incremental layering technique that takes advantage of the improved handling characteristics and proposed reduced shrinkage and greater depth of cure. CLINICAL IMPLICATIONS: When this new technique is used with one-bottle adhesives and improved instrumentation, posterior heavy-body composites can be placed faster, easier and possibly more predictably than when medium-body resins and previous techniques are used. PMID- 10715931 TI - A practice-based study of a power toothbrush: assessment of effectiveness and acceptance. AB - BACKGROUND: The greater effectiveness of the power toothbrush compared with a manual toothbrush is well-documented. Despite this, acceptance by dental professionals is still low. METHODS: This general practice study evaluated the effectiveness of a power toothbrush (Braun Oral-B Ultra Plaque Remover, Braun GmbH) in 16,903 patients, based on the clinical opinions of dental professionals in regard to patients' changing oral health status. In addition, a survey assessed the attitudes of dental professionals and patients toward the power toothbrush. RESULTS: The power toothbrush was considered by dental professionals to have had a positive effect on the oral health of 80.5 percent of their patients; the noticeable benefits with respect to a number of clinical criteria included plaque removal and improved gingival condition. Most patients in the study (88.9 percent) reported that they would continue using the power toothbrush once the study was completed. At the end of the study, many more dentists and hygienists considered the power toothbrush to be the most effective way of brushing, and almost 70 percent said that they would now be more likely to recommend a power toothbrush to their patients. CONCLUSIONS: The power toothbrush improved the oral health of patients in this practice-based study, and the number of dental professionals who said they would recommend a power toothbrush increased markedly during the study. CLINICAL IMPLICATIONS: Enabling dental professionals to evaluate the effect of a power toothbrush reinforces the findings from controlled clinical studies and increases their awareness of its potential to improve oral hygiene. PMID- 10715932 TI - Challenging a negative report. PMID- 10715933 TI - [Physicians--failure in marriage?]. PMID- 10715934 TI - [Deep venous thrombosis of the leg: when thrombophilia diagnosis?]. PMID- 10715935 TI - [Drug interactions with Hypericum. Is nature not so mild after all?. Interview by Petra Eiden]. PMID- 10715936 TI - [Back pain: guidelines for drug therapy. Utilize the therapeutic spectrum]. AB - For the treatment of back pain, behavioral modification, pharmacotherapy, non pharmacological conservative treatment and surgical procedures are available. Systemic and metabolic disorders require specific treatment. Medication that may be considered includes non-opioid analgesics with or without an antiphlogistic action, opioid analgesics, muscle relaxants and antidepressants. The choice of substance(s) will depend on the pathophysiology and the duration of the pain condition. In the case of acute pain with an inflammatory component, NSAIDs are the drugs of first choice. However, prolonged administration carries a risk of gastrointestinal and renal complications. Muscle relaxants may be useful adjuvants for a limited time. For myofascial pain, flupirtine is to be recommended because of its analgetic and muscle-tone-normalizing actions. Local anesthetic infiltration or nerve blocks are useful in blocking nociception with its pathophysiological sequelae. For chronic back pain, opioids and some antidepressants have a more favorable benefit-risk profile than NSAIDs. PMID- 10715937 TI - [Prevention of relapse in schizophrenia is discontinued too soon. Many colleagues make this mistake!]. AB - OBJECTIVE: The recommendations given by general practitioners in the treatment of schizophrenia were investigated empirically and compared with those given by psychiatrists as well as with the accepted standards for the prophylactic treatment of schizophrenic psychoses. METHOD: In an interview based on case reports, 64 randomly selected general practitioners in private practice were asked 6 questions on the treatment of schizophrenic psychoses. RESULTS: 75% of the general practitioners recommend a duration of prophylactic measures that is significantly shorter than that recommended by international standards, and also significantly shorter than that recommended by psychiatrists. This undertreatment correlates with an overestimation of the risk of side effects and--at least in first-time patients--an underestimation of the relapse risk. CONCLUSION: Despite a number of significant differences between general practitioners and psychiatrists, both groups fall far short of international treatment standards in terms of recommended duration of prophylaxis, both underestimate the relapse risk and overestimate the risk of side effects. Possible approaches to improving the quality of relapse prevention in schizophrenia are, for example, joint educational activities for family doctors and psychiatrists, and the establishment and implementation of joint treatment guidelines. PMID- 10715938 TI - [When necrosis smells of heating oil... what damage fuels can do]. AB - The subcutaneous injection of heating oil or other crude oil distillates are rare injuries. In the present case, a 26-year-old man injected heating oil subcutaneously into the left cubital region. He then developed massive swelling, pain, local necrosis and abscess, accompanied by fever and leukocytosis. Radical surgical debridement and open wound treatment successfully stopped the necrotic process. Subsequently, a mesh-graft was applied to the wound, which healed with no residual defects. The course of the present case, and the results of a review of the literature on similar occurrences involving mineral oil suggest early extensive debridement of such injuries. PMID- 10715939 TI - [Respiratory tract infections in general practice. Cefpodoxime proxetil in patients with risk factors and concomitant illnesses]. PMID- 10715941 TI - [24-hour blood pressure measurement. Ambulatory blood pressure monitoring. 2: Tips for use in general practice]. PMID- 10715940 TI - [Medical history in panarteritis nodosa. Individual therapy in relation to disease activity]. PMID- 10715942 TI - [Abdominal pain after taking drug tablets. Acute intermittent porphyria]. PMID- 10715943 TI - [Your severely handicapped patient gains his rights. Handicapped status: tax loophole for the common man?]. PMID- 10715944 TI - [Tuberculosis epidemiological situation in the Republic of Belarus]. AB - The paper gives data on the spread of tuberculous infection in Byelarus. More preventive measures are shown to improve major epidemic indices, by stabilizing the situation primarily among children and adolescents. The epidemiological situation remains to be alarming among the contingents of the boarding facilities of the Ministry of Social Welfare and convicts at reformatories. PMID- 10715945 TI - [Clinical and epidemiological characteristics of patients with drug-resistant tuberculosis]. AB - The paper analyzes the drug-resistant causative agent of tuberculosis. The bacterial isolation rates were studied among the cohorts registered at tuberculosis control facilities in the Republic of Byelarus in 1997. The epidemiological significance of patients isolating polyresistant Mycobacterium tuberculosis strains is shown. PMID- 10715946 TI - [Tuberculosis in penitentiaries of the Republic of Belarus]. AB - The paper analyzes the prevalence of tuberculous infection among special cohorts of penitentiaries in the past decade. The incidence of tuberculosis in prisons in 1997 was 28.3 greater than that in the general population, which negatively affects the whole epidemiological situation. The optimal methods of its detection, treatment, and prevention among the convicts in the Republic of Byelarus are substantiated. PMID- 10715947 TI - [Disability in patients with pulmonary tuberculosis: prognosis of its onset]. AB - Disability prediction can timely detect the signs of disability in the patient, greatly enhance the quality of sociomedical expert examination. A method for prediction of the onset of disability in patients with pulmonary tuberculosis has been developed on the basis of the correlation-regression analysis of the factors that influence disability. Verification of the efficiency of prediction of disability in control patients has provided evidence for the feasibility of using this method in the work of phthisiologists and experts of medical rehabilitative expert examination commissions. PMID- 10715948 TI - [Enhancing treatment efficiency of patients with pulmonary tuberculosis by regional lymphotropic therapy]. AB - To reduce the duration of treatment, to reduce the incidence of adverse effects, to enhance the total efficiency of chemotherapy in the complex treatment of 95 patients with different forms of pulmonary tuberculosis, lymphotropic therapy was used with subcutaneous 10% isoniazid in combination with heparin, lazix or lidase. Lymphotropic therapy caused no side effects in response to isoniazid. Lymphotropic therapy was found to substantially accelerate abacillation, the cessation of intoxication symptoms, the closure of decay cavities and to reduce hospital stay by 1.0-1.5 months. PMID- 10715949 TI - [Problem of drug-resistant tuberculosis and the way of its solution]. AB - Drug-resistant tuberculosis in the Ukraine and in other countries as well is a great important problem. The presence of drug-resistant Mycobacteria worsens clinical and bacteriological responses to therapy, prolongs hospital stay, and increases costs. The novel derivative of rifampicin, rifabutin, produced by Pharmacia & Upjohn under the trade name Mycobutin is more effective and safe than rifampicin. Rifabutin is approved for clinical application for tuberculosis and other mycobacterial infections. It is also recommended for rifampicin-resistant tuberculosis. PMID- 10715950 TI - [Efficiency of lomefloxacine in combined treatment of patients with multiresistant pulmonary tuberculosis complicated with nonspecific bronchopulmonary infection]. PMID- 10715951 TI - [Surgical methods on the diagnosis of interstitial lung diseases and intrathoracic adenopathies]. AB - The efficiency of surgical methods for diagnosis of unclear cases of pulmonary interstitial diseases and intrathoracic adenopathies in 136 patients over 5.5 years was analyzed. A high percentage (45.7%) of discrepancies was found in the clinical and conclusive diagnosis in patients undergone test therapy when the diagnosis was unclear over 2 months. The efficiency of surgical diagnostic methods was 90-97.6%. PMID- 10715952 TI - [Tuberculosis and opportunistic infections]. AB - A serological study of blood from 44 patients with acutely progressive forms of pulmonary tuberculosis revealed their rather common association with active forms of the following opportunistic infections, such as M. pneumonia, Pn. carinii, and herpes simplex virus. PMID- 10715953 TI - [Incidence and pattern of bronchial system lesions in patients with respiratory tuberculosis under the present conditions]. AB - The paper presents the detection rates of active and inactive bronchial tuberculosis in patients with different clinical forms of respiratory tuberculosis in 1988-1989 (90 patients) and 1996-1997 (384 patients), comparatively characterizes the clinical forms of bronchial tuberculosis. There was a substantial increase in the detection rates of postprimary active and inactive bronchial tuberculosis. PMID- 10715954 TI - [Impact of combined magnetic and laser radiation of regional pulmonary blood flow in patients with destructive pulmonary tuberculosis]. AB - Rheopulmonography was used to study regional pulmonary blood flow in 30 patients with destructive pulmonary tuberculosis before and after combined magnetic and laser radiation (an experimental group) and in 28 patients receiving the routine chemotherapy (a control group). The use of combined exposure of a constant magnetic field and laser radiation was found to promote pulmonary vascular tone, better microcirculatory blood flow, and increased pulse blood filling in the affected portion of the lung. PMID- 10715955 TI - [Surgical treatment of common forms of urinary system tuberculosis]. AB - The paper deals with the efficiency of various operations on the urinary system. It emphasizes a strict approach to surgical interventions by taking into account the functional status of the kidneys, the magnitude of chronic renal failure, the need for antibiotic therapy in the pre- and postoperative periods. PMID- 10715957 TI - [Helium-neon laser radiation in the therapy of urinary bladder and ureteral tuberculosis]. AB - The paper presents a comparative assessment of conventional therapy methods and the author's proposed treatment by using low-intensity helium-neon laser radiation in combination with endovesical submucous administration of tuberculostatics and glucocorticosteroids into the area of lesion in patients with tuberculosis of the bladder and intramural ureter. PMID- 10715956 TI - [Low-intensity laser radiation in combined treatment of urinary system tuberculosis]. AB - Combined treatment using low-intensity laser radiation (LILR) was performed in 125 patients with urinary tuberculosis: 54 patients had combined intravascular blood laser radiation (IBLR) and external renal laser radiation, 71 patients were exposed to IBLR with an AZOR-2K device. The use of LILR in the combined treatment contributed to the stabilization of a specific process, to significant renal functional improvement, and a reduction of a postoperative period from 26 to 20 days. PMID- 10715958 TI - [Osteoarticular tuberculosis: clinical and social aspects]. AB - A hundred and ninety four history cases of inpatients were analyzed to study such clinical and social aspects of active osteoarticular tuberculosis as age-sex structure; a role of residence, occupation, and bad habits in the onset of the disease; the pattern of clinical forms and complications; bacterial isolation rates; the incidence of mixed forms and complications. The major clinical symptoms and outcomes are described. PMID- 10715960 TI - [Gastroduodenal mucosal morphological changes in tuberculosis-infected children]. AB - Three groups of children aged 4 to 14 years who complained of having dyspepsia were examined. Group 1 included 31 children without tuberculous infection; Group 2 comprised 30 newly tuberculosis-infected children; Group 3 consisted of 35 children who had an over one-year history of the infection. Esophagoduodenoscopy and biopsy indicated that the incidence of Helicobacter infection and gastritis was higher in children with tuberculous infection than in those without it and it was directly related to the duration of infection. The specific features of chronic gastritis in the infected children were the totality of gastric mucous lesion with more profound antral changes, the predominance of progressive gastritis, as appeared as neutrophilic infiltration, gastric mucous contamination with Helicobacter and Candida, and metaplasia. PMID- 10715959 TI - [Efficiency of ramipril treatment in patients with chronic obstructive bronchitis complicated by chronic cor pulmonale]. AB - The supplementation of the angiotensin-converting enzyme inhibitor ramipril to combined therapy in patients with chronic obstructive bronchitis complicated by chronic cor pulmonale resulted in positive central, pulmonary, and peripheral hemodynamic changes, improved peripheral tissue oxygenation, by exerting beneficial effects on the course of the disease. PMID- 10715961 TI - [Contribution of immune complex pathological reactions to immunogenesis of pulmonary tuberculosis]. AB - The association of long-term persistence of circulating immune complexes in blood flow with their deposits in the lung in 54.4% of cases and with morphological reactions of cellular and tissue structures, with the functional status of the immune system and with the progression of a tuberculous process, which was found immunohistochemical, immunological, and light microscopic studies in 68 patients with pulmonary tuberculosis, and the high reproducibility of lung-damaging effects of immune complexes in the experiment suggest that the immune complex pathological mechanism is involved into the pathogenesis of progressive pulmonary tuberculosis. PMID- 10715962 TI - [Clinical and morphological characteristics of deaths in patients with tuberculosis]. AB - The clinical, morphological, and tanatological features of tuberculosis have been studied in patients who died in 1994-1996. Group 1 and Group 2 included 91 and 190 patients, respectively. It has been found that the present-day stage is marked by a trend for tuberculosis reversion with the characteristics typical of the preantibiotic period. Along with worse epidemiological parameters, there is an increase in the incidence of miliary tuberculosis and caseous pneumonia. The common cause of death is tuberculosis progression. The present stage differs from the preantibiotic period by the fact that progression rapidly occurs with specific therapy due to an increase in cases with documented drug resistance. PMID- 10715963 TI - [A rare case of bilateral hemorrhagic pleurisy]. PMID- 10715964 TI - [Possible use of polymerase chain test (DNA diagnosis) in detection of mycobacterium tuberculosis]. AB - The polymerase chain test (PCT) (DNA diagnosis) is comparable with other routine diagnostic methods, but has a variety of advantages over then: it is much more highly sensitive and specific, provides results more promptly than culturing. The tests have indicated that the Polytub diagnostic kit may be recommended for use at medical facilities in Russia to detect tuberculosis pathogen DNA in the sputum, urine, exudate, and spinal fluid using PCT. PMID- 10715965 TI - [Variant of lymphogenic tuberculosis simulating cavity in lung]. PMID- 10715966 TI - [Fournier disease and genital tuberculosis]. PMID- 10715967 TI - Equity and health. PMID- 10715968 TI - [Echinococcosis/hydatidosis in the VII Region of Chile: diagnosis and educational intervention]. AB - This study was designed to embrace three areas: a) serologic and radiologic diagnosis and surgical treatment of hydatidosis in an asymptomatic human population, b) animal diagnosis and the treatment of dogs, and c) evaluation of extent of knowledge and performance of educational interventions among rural families and health, livestock, and education professionals and technicians, in order to help control the disease transmission cycle. Indirect hemagglutination and ELISA tests were performed on 5,556 apparently healthy people. Of these, 42 (0.8%) had positive results on both tests, for a seroprevalence of 754.6 per 100,000. These 42 subjects were scheduled for liver ultrasonography and a chest x ray; of the 26 who complied, 16 showed images compatible with a hydatid cyst. Those 16 cases were sent to the hospital for surgery. In 9 of the cases the diagnosis was confirmed surgically, for a prevalence of 161.7 per 100,000. Arecoline hydrobromide was administered as a laxative to 2,358 dogs to detect the strobilar form of Echinococcus granulosus, and positive results were found in 11% of the dogs. Official data for slaughterhouses indicated the presence of hydatid cysts in 13% of the cattle, 4.4% of the sleep, and 4.2% of the pigs slaughtered in the region. The educational program included an evaluation of the extent of knowledge by surveying heads of household; an educational intervention among families through an informal active participatory process using educational games, in which 1,082 families participated; and an educational intervention with professionals and technicians using distance and in-person approaches. To evaluate the program, the results of knowledge tests before and after educational interventions with 200 families (cases) were compared with those from 95 families who did not participate (controls). Of the 1,423 heads of household initially surveyed about their knowledge of echinococcosis/hydatidosis, 783 of them (55%) said they knew nothing about the infection. It was found that the participatory educational games were well adapted to the lifestyle of people from rural areas and made change possible. Training was provided to 276 health professionals, 201 technical assistants, and 453 rural teachers. The program reached 100% of the staff members of the area's rural primary health care services. PMID- 10715969 TI - [The use of psychotropics by dental patients in Minas Gerais, Brazil]. AB - In Brazil, psychotropics and other drugs are often indiscriminately overused. Nevertheless, there are few studies regarding the use of psychotropics, especially among dental patients. The purpose of this study was to assess the prevalence of psychotropic consumption among patients of the general primary care clinic of the Dentistry School at the Minas Gerais Federal University, in Brazil. To collect data, students working in the clinic interviewed all patients over 12 years of age seen at the clinic during June 1997 and asked them about their use of psychotropics during the preceding 2-week and 12-month periods. The results showed that 4% of the patients had used psychotropic drugs in the 2 weeks before the study and that 10% of them had used psychotropics in the preceding 12 months. The drugs used most frequently in the 12-month period were anxiolytics (around 40% of total use). The median age of the patients was 23 years old. Persons under the median had used psychotropics less in comparison with older persons (P < 0.01). A significant association (P < 0.05) was found between using drugs and being female, and also between the use of drugs and being a housekeeper or a housewife (P < 0.03). Patients with a regular relationship (married or living together) used more psychotropics than patients who were single, widowed, or divorced (P < 0.03). There was no association between the use of drugs and the level of education. Even though information on the use of psychotropics is important for dental diagnosis and planning, only 40% of the students said they noted this information in their patients' charts. That fact suggests that dental student education may be lacking in this regard and that dentistry training should take into consideration the issue of patients' use of drugs. PMID- 10715970 TI - Epidemiology of malaria in the Amazon basin of Ecuador. AB - Malaria is reemerging in most endemic countries of South America. In Ecuador, malaria is endemic on the Pacific coast, in the inter-Andean valleys, and in the Amazon River basin. In the Lower-Napo region of northeastern Ecuador, malaria was considered eliminated in the 1970s, but the disease has reemerged in recent years. Three organizations are involved in malaria-related work in the area, but they are not coordinating their efforts. This study was designed to describe the epidemiology of malaria incidence in the Lower-Napo region for the period of January 1992 through December 1995, and to determine the extent of seasonality in transmission in the area. To determine malaria incidence, data were collected for that 4-year period from the records of the three malaria-related organizations: the office of the National Center for Malaria Eradication (NCME) in the town of Coca, the district hospital in Nuevo Rocafuerte (DHNR), and an association of community health workers called Sandi Yura. Data on climatic conditions for the same period were collected from the Ecuadorian Air Force and civil aviation authorities. During the 1992-1995 period, NCME diagnosed a total of 773 malaria cases, DHNR diagnosed 485, and Sandi Yura clinically diagnosed 859. For the 4 year period, an annual parasite index of 40.4 was found with the DHNR data, 35.8 with the Sandi Yura data, and 6.2 with the NCME data. The predominant parasite in the area was Plasmodium vivax (92% of all the cases). Twenty-eight percent of the infected persons were under 10 years old. No discernible differences between the genders were found. There was also no seasonal variation among the cases. Further research is needed in order to confirm these findings and better understand malaria transmission in the region. The study highlights the need for a closer coordination among the area's malaria-control organizations so as to have an improved understanding of malaria epidemiology and to design and implement effective control strategies. PMID- 10715971 TI - [The importance of genetics in the public health care system: report on the closing down of a genetics division in Sao Paulo, Brazil]. AB - The availability of genetics services in hospitals should be an issue of public concern. Having genetics services helps shorten the time spent in making diagnoses and reduces the average number of inpatient days; it also helps accelerate the choice of adequate treatments, prevents or minimizes sequelae, and, ultimately, reduces costs. The objective of the present study was to describe the closing down in 1996 of the genetics division at the Menino Jesus Children's Hospital, a pediatric institution located in the city of Sao Paulo, SP, Brazil. A retrospective analysis was carried out of the work performed by this division between 1992 and 1996, with an emphasis on the detection of chromosomal abnormalities. Of all cases assessed during the study period, 571 were entered into a database. Some kind of chromosomal abnormality was observed in 20% of the 350 karyotypes performed. The existence of genetics services in hospitals helps minimize the appearance of clinical symptoms in carriers of genetic abnormalities, improves the quality of life of these patients, and enables them to receive information regarding risk of recurrence, while preventing the waste of resources that results from tests that are costly and unnecessary. Such benefits amply justify the investment in setting up genetics services of the type described here. PMID- 10715972 TI - [Report on a leptospirosis outbreak in the city of Santa Fe, Argentina, March April 1998]. AB - In March-April 1998 in a neighborhood in the city of Santa Fe, Argentina, there was an outbreak of an acute disease characterized by fever, headaches, and intense myalgias. This article presents the studies surrounding this outbreak and the attempts to identify the source and the mode of transmission. The epidemiological, serological, and clinical findings indicated that the causative agent was Leptospira interrogans. As a screening test, macroscopic agglutination with heat-resistant antigen was applied, followed by the ELISA test, and, as a confirmatory test, microscopic agglutination for 10 serotypes of L. interrogans. The study covered 32 persons, 8 dogs, and 8 water samples. Among the 32 persons, 12 cases were confirmed, 2 were suspicious, and 18 were negative. Six dogs were found to be infected, and motile spirochetes were found in the water samples. The human sera reacted with the ballum, canicola, icterohaemorrhagiae, and pyrogenes serotypes; the canine sera reacted with the ballum, canicola, and pomona serotypes. The coagglutination found in all the confirmed cases indicates that they were acute cases of leptospirosis, but it was impossible to identify the causal serotype. Except for the index case, the disease was not recognized clinically. Several facts suggest that the outbreak was caused by rain that had flooded the study area. The results of this study emphasize the need for active surveillance of leptospirosis when there are floods and other natural disasters. PMID- 10715973 TI - Pregnancy-related mortality in Guatemala, 1993-1996. AB - To select the proper interventions that could prevent maternal mortality, adequate and appropriate maternal mortality data are needed. Nevertheless, the quality and quantity of information and the scope of maternal health- and death related data are inadequate in many countries, particularly in the developing world. From January 1993 to December 1996 a surveillance program in maternal mortality was developed to conduct surveillance studies in the department of Guatemala, Guatemala. With an active surveillance system, our approach gave a more complete picture of maternal death and produced information on the specific causes of maternal mortality. Using multiple sources of information, we reviewed and analyzed all deaths of women of childbearing age (10 to 49 years). Each death was investigated to determine whether it was pregnancy-related or not. The maternal mortality ratio for the four-year study period was 156.2 deaths per 100,000 live births. Women 35 and older had a higher risk of maternal death than women under that age. Women who were 35-39 years old had a maternal death risk almost three times as high as women aged 20-24. For women who were 40 or older the risk was more than double that of women 20-24 years old. Overall, the two leading causes of maternal mortality were infection and hemorrhage. Vaginal deliveries where there was medical assistance had the highest rate of delivery related maternal death from general infection. In deliveries attended by nonmedical personnel, delivery-related maternal deaths from hemorrhage were most frequently associated with retained placenta. Developing countries are called on to implement systems that can provide continuous and systematic data collection so that policymakers and health managers have adequate information to design proper interventions to save women's lives. PMID- 10715974 TI - [Aging and health: a change in paradigm]. AB - The aging of the population due to increasing life spans requires a reevaluation of the health care models that prevail in the countries of the Region of the Americas, given that the illnesses typical of old age make up an ever-increasing share of the morbidity burden. The countries in the Region are in an intermediate stage of demographic transition. They face the challenge of balancing priority care for the health problems characteristic of developing countries along with care for the chronic and debilitating diseases that afflict the elderly. An even greater challenge is gauging, in a timely manner, the health, social, and economic consequences of the demographic transition over the next millennium, to allow the countries to respond to these consequences through an in-depth restructuring of their health and social services. The extension of human life imposes a new vision of the health of the elderly that is oriented toward self sufficiency, economic independence, and living life to its fullest. It will not be easy to integrate this new paradigm into the countries' health services, which are now primarily oriented toward curative care and are structured to fulfill the requirements of an epidemiological scenario that is undergoing a rapid transformation. The challenge is even greater because of the wide range of needs that the elderly have across countries with varying degrees of development. Nevertheless, sooner or later the governments of all the countries of the Region will be forced to adopt concrete measures to deal with the problems posed by the population's gradual aging. PMID- 10715975 TI - Pulmonary and hemodynamic changes during laparoscopy--are they important? PMID- 10715976 TI - Improvement in the information content of the Glasgow Coma Scale for the prediction of full cognitive recovery after head injury using fuzzy logic. AB - BACKGROUND: The objective of this study was to modify the existing Glasgow Coma Scale (GCS) into a fuzzy GCS by using fuzzy information representation and fuzzy inferencing. The study compared the information content of the existing GCS with the new fuzzy GCS for prediction of full cognitive recovery in patients with head injury. METHODS: A record-based study was conducted at the Government Medical College and Hospital, a tertiary care facility in Nagpur, India. The study, which covered the period from January 1 to December 31, 1997, included 253 patients with head injuries. Opinions of 17 clinical experts who routinely deal with head injury cases were used for the construction of the fuzzy GCS. RESULTS: By using the max operator for summarization, eye, motor, and verbal stimuli were all significantly associated with the possibility of full cognitive recovery with the fuzzy GCS (P < .001). Nonspecificity of the classical GCS, the min-operated fuzzy GCS, and the max-operated fuzzy GCS was comparable. A reduction in Shannon entropy was maximum with the max-operated fuzzy GCS. Min-operated fuzzy GCS better predicted a lack of full cognitive recovery. CONCLUSIONS: Fuzzy GCS substantially improves the information content for prediction of the possibility of full cognitive recovery after head injury. Eye, motor, and verbal stimuli all uniquely and significantly contribute to prediction of this possibility. We recommend the use of fuzzy GCS for prediction of the possibility of full cognitive recovery in patients with head injuries. PMID- 10715977 TI - Invited commentary: fuzzy logic--an introduction. PMID- 10715978 TI - Invited commentary: fuzzy logic, clear reasoning. PMID- 10715979 TI - Measuring improved quality of life after laparoscopic Nissen fundoplication. AB - BACKGROUND: While the correction of pathologic gastroesophageal reflux by means of laparoscopic Nissen fundoplication (LNF) has been well documented, the psychological profiles of patients with this disease and the impact on their quality of life are less well understood. We obtained a baseline psychological profile and measured the impact of LNF on patients' quality of life with 2 standardized instruments: the psychological general well-being index (PGWB) and the gastrointestinal symptoms rating scale (GSRS). The study included 34 consecutive patients with typical symptoms of gastroesophageal reflux who underwent LNF in 1995 at a tertiary care university medical center. METHODS: Patients filled out PGWB and GSRS surveys preoperatively and at 2 weeks, 2 months, and 12 months postoperatively. Data were collected in a blinded fashion by a study nurse and analyzed after completion of the study. Data are expressed as mean +/- standard deviation. RESULTS: The mean preoperative PGWB score (69.6 +/- 17.3) of study patients with gastroesophageal reflux disease was lower than that expected for a healthy population. This was primarily attributable to low scores in the general health domain of the questionnaire, although LNF patients also had low scores in the vitality and positive well-being domains of the PGWB scale. LNF improved the PGWB score to a normal level (78.7 +/- 19.3) (P = .05 vs the preoperative PGWB score) at 12 months post surgery. The GSRS also showed improvement from 34.7 +/- 7.8 to 28.1 +/- 10 (P = .008). The improvement in GSRS was attributed to improvement in the heartburn (7.12 +/- 2.4 to 2.72 +/- 1.2, P < .001) and abdominal pain (6.58 +/- 2.5 to 4.92 +/- 1.6, P = .006) domains of the scale. LNF had no impact on the diarrhea, indigestion, and obstipation domains of the GSRS. CONCLUSIONS: Patients with gastroesophageal reflux disease who are candidates for LNF have low psychological and general well-being scores that are restored to normal levels by successful LNF. When compared with baseline measurements, LNF effectively relieved heartburn and did not cause significant new gastrointestinal complaints. PMID- 10715980 TI - Carotid endarterectomy in women: early and long-term results. AB - BACKGROUND: Although many randomized trials and other multicenter studies have demonstrated the benefits of carotid endarterectomy (CEA) in selected symptomatic and asymptomatic patients, including women, there is a remarkable lack of reports regarding the outcome of CEA with respect to sex. To analyze and compare the outcome of CEA in men and women in a single-group experience, we reviewed a consecutive series of 619 CEAs performed in 539 patients, 371 men (423 CEAs) and 168 women (196 CEAs). METHODS: Data collection was retrospective up to August 1, 1992 and prospective for all 405 patients treated thereafter. RESULTS: Women were significantly less likely than men to have overt evidence of coronary artery disease (P < .001) and had a significantly higher incidence of diabetes (P < .001). No perioperative death occurred in the female group (P = NS), and no statistical difference was found in perioperative stroke risk incidence. Women had a significantly higher incidence of late occlusive events (P = .01), which were all asymptomatic. No late stroke occurred in the female group (P = NS). Life table cumulative survival rates at 1, 3, 5, and 7 years were 99.3%, 90.5%, 85.9%, and 82.3%, respectively, in women, and 98.9%, 91.9%, 85.2%, and 79.6% in men (log rang P = .8). CONCLUSIONS: These findings show that perioperative stroke risk and mortality rates, as well as late stroke-free, mortality, and recurrence rates, in patients undergoing CEA, are comparable in men and women. Further, larger comparative studies are necessary to provide more information on the benefit and durability of CEA in asymptomatic patients, but the results of this study suggest that the early and late outcomes are excellent and comparable in symptomatic and asymptomatic men and women. PMID- 10715981 TI - The influence of female gender on the outcome of carotid endarterectomy: a challenge to the ACAS findings. AB - BACKGROUND: In the Asymptomatic Carotid Endarterectomy Study (ACAS) the perioperative stroke and mortality rate was more than twice as high in women as in men, markedly reducing the long-term benefit of the operation; therefore the role of carotid endarterectomy (CEA) among women with asymptomatic carotid stenoses remains unclear. The current study was undertaken to further examine the influence of gender on the outcome of the operation. METHODS: To control for all variables except gender, the records of all patients in an academic medical center who underwent elective CEA for asymptomatic disease, performed by one surgeon employing a uniform technique, over a 7-year interval were reviewed. RESULTS: From January 1992 through September 1998, 156 CEA procedures for asymptomatic carotid stenoses were performed on 66 (44%) women (n = 68) and 83 (56%) men (n = 88). There were no differences in the prevalence of hypertension (69% vs 69%), diabetes mellitus (24% vs 19%), hyperlipidemia (47% vs 47%), or smoking (46% vs 60%) between women and men, respectively, although a history of angina (28% vs 13%, P < .05) and myocardial infarction (23% vs 6%, P < .01) was more common among men. The mean stenosis was 86% for men and 83% for women. The incidence of perioperative mortality, stroke, and transient ischemic events was 0%, 0.6%, and 0%, with no differences between women and men: 0% vs 0%, 0% vs 1.3%, and 0% vs 0%, respectively. CONCLUSIONS: These findings indicate that female gender does not adversely influence the outcome of CEA when performed for treatment of asymptomatic disease. Gender should not be a consideration in the decision to perform CEA because of asymptomatic disease. PMID- 10715982 TI - Lichtenstein patch or Perfix plug-and-patch in inguinal hernia: a prospective double-blind randomized controlled trial of short-term outcome. AB - BACKGROUND: Open mesh used in anterior inguinal hernia repair can be configured as a flat patch (Lichtenstein operation) or as a cone-shaped preformed plug and supplementary patch (plug-and-patch operation; Perfix Plug; Davol Inc, Cranston, RI). METHODS: One hundred forty-one patients were randomly allocated and blinded to receive either a Lichtenstein patch or a Perfix plug-and-patch. Information before the operation and on postoperative days 3 and 14 was recorded by an independent blinded observer to include operating time, postoperative pain, analgesic medication, return to activity and work, and quality of life assessment. RESULTS: Operating time (32 vs 37.6 minutes) was significantly shorter in the plug-and-patch group (P = .01). During days 1 through 8, patients who had undergone the plug-and-patch operation experienced less pain, and their physical functioning on day 3 was significantly better (P = .013). Days of analgesic medication (4.0 vs 4.6 days), return to normal activity (2.8 vs 3.6 days), return to work (17.0 vs 20.8 days), and total days of work missed (14.3 vs 16.1 days) were similar in both groups (P = NS for all comparisons). CONCLUSIONS: Compared with patients who received the Lichtenstein patch for ambulatory inguinal hernia repair, patients who underwent the Perfix plug-and-patch operation experienced less postoperative pain in the first 8 days after the operation but consumed similar postoperative analgesic medication. The rate of return to normal activity and work is similar in both groups, which indicates no superiority for the plug-and-patch operation in overall rehabilitation and societal costs. Overall hospital costs are greater for the plug-and-patch operation ($120 [US]) compared with the Lichtenstein patch ($20 [US]), with a negligible (5.6 minutes) saving of operating room time for the plug-and-patch operation. PMID- 10715983 TI - Minimally invasive esophagectomy for Barrett's esophagus with high-grade dysplasia. AB - BACKGROUND: Barrett's esophagus with high-grade dysplasia (BE/HGD) is associated with invasive carcinoma in 30% to 70% of cases. Esophagectomy is the treatment of choice for patients with BE/HGD but esophagectomy can be associated with high morbidity and mortality. The aim of our study was to report our preliminary experience in applying minimally invasive surgical techniques to esophagectomy for BE/HGD. METHODS: From August 1996 to February 1999, 12 consecutive patients underwent minimally invasive esophagectomy for biopsy-proven BE/HGD. Our consort consisted of 7 men and 5 women; average age was 64 years (range, 40-78 years). All patients underwent a complete laparoscopic or combined laparoscopic and thoracoscopic resection of the esophagus with cervical anastomosis. RESULTS: Mean operative time was 7.8 +/- 2.1 hours, mean intensive care unit stay was 2.6 +/- 2.2 days, and mean length of hospital stay was 8.3 +/- 4.7 days. Five patients (42%) had carcinoma in situ or carcinoma identified on pathologic specimen. Analyses of all resected lymph nodes in the 12 patients were negative for metastatic disease. There were 6 major complications in 5 patients: 1 patient had a small bowel perforation requiring operative repair, 2 patients needed prolonged ventilatory support for respiratory insufficiency, and 3 patients had delayed gastric emptying requiring revision of the pyloromyotomy. The single minor complication in this series was a jejunostomy tube-site infection. There were no conversions to laparotomy or thoracotomy. All patients were alive and free of metastatic disease at a mean follow-up of 12.6 months. CONCLUSIONS: Minimally invasive esophagectomy is a feasible and safe alternative to conventional open esophagectomy for patients with BE/HGD. PMID- 10715984 TI - Fecal continence following partial resection of the anal canal in distal rectal cancer: long-term results after coloanal anastomoses. AB - BACKGROUND: The aim of the study was to assess the influence of partial excision of the superior portion of the anal canal (AC) when necessary for tumor margin clearance in distal rectal cancer on fecal continence after coloanal anastomoses. METHODS: Between 1977 to 1993, 209 patients with middle and lower third rectal cancers underwent complete rectal excision and coloanal anastomoses. For very low tumors, located at or below 5 cm from the anal verge (AV), varying portions of the superior segment of the AC were excised for tumor margin clearance. The magnitude of resections was inversely proportional to the height of the anastomosis from the AV. The patients were categorized into 3 groups according to their level of anastomoses from AV: group 1, patients with anastomoses from 0.5 to less than 2 cm from AV (1 to 2.5 cm of AC resected, i.e., major resection); group 2, anastomoses at 2 to less than 3 cm from AV (less than 1 cm of AC resected, i.e., minor resection); group 3, with anastomoses at 3 to 3.5 cm from AV (AC completely preserved). A standard questionnaire, physical examination, and anal manometry at intervals of 3, 6, 12, 24, 36, and 48 months were performed prospectively to assess anal continence. RESULTS: The patients in the 3 categories were matched for age, gender, stage, presence or absence of a colonic J-pouch, preoperative neoadjuvant radiotherapy and surgical technique. Fourteen patients with postoperative radiotherapy were excluded from the clinical assessment. Mean follow-up was 33.5 months. There were 43 patients in group 1, 75 in group 2, and 73 in group 3 for clinical assessment. In the first year, there was progressive improvement in anal continence in all 3 groups. At 2 years, 50% in group 1, 73% in group 2, and 62% in group 3 were fully continent. The proportion of patients fully continent in group 1 remained unchanged as compared to continued improvement for groups 2 and 3 following the first year. At 4 years, 50% in group 1, 80% in group 2, and 68% in group 3 were completely continent. The difference among the 3 groups was not statistically significant. CONCLUSIONS: For distal rectal cancer, where tumor margin clearance necessitates partial resection of the superior portion of the AC, when limited to less than 1 cm, the proportion of patients remaining fully continent is similar to those with complete AC preservation. More substantial excisions of the AC can still result in satisfactory anal continence, such that following the fourth year, one half of the patients can expect to be fully continent. PMID- 10715985 TI - Wound recurrence from gallbladder cancer after open cholecystectomy. AB - BACKGROUND: Reports of port site recurrences from gallbladder cancer after laparoscopic cholecystectomy have raised considerable concern as to whether the laparoscopic technique implies an increased risk of metastatic disease. In a previous study of gallbladder cancer and laparoscopic cholecystectomy, we reported a frequency of 16% port site metastases. The purpose of the present study was to determine the frequency of wound metastases from gallbladder cancer after open cholecystectomy. METHODS: The registers from the Swedish Oncological Centers and the National Board of Health and Welfare were checked for reported cases of gallbladder cancer and surgical classification codes for open cholecystectomy from 1991 to 1994. The study included all 8 university and 24 county hospitals in Sweden. The files from all patients with gallbladder cancer who had an open cholecystectomy were retrospectively reviewed. RESULTS: The study included 270 patients who had a cholecystectomy, of which 215 were classified as open and 55 as laparoscopic. Of the 215 patients, 11 patients were excluded because of an incorrect or deficient histopathologic or surgical classification. In 186 patients (91%), sufficient data were obtained for follow-up. Twelve patients (6.5%) had wound metastases from their gallbladder cancer. All patients with wound metastases died with a median survival of 10 months (range, 3 to 65 months). CONCLUSIONS: Wound metastases from gallbladder cancer after open cholecystectomy may be more common than previously assumed. PMID- 10715986 TI - Suppression of humoral immunization against encapsulated xenogeneic hepatocytes and prolongation of their function by 2-week cyclosporine treatment in the rat. AB - BACKGROUND: Xenogeneic liver transplantation may induce immune reactions not only against the grafted liver but also against the proteins that it synthesizes. We investigated whether 2-week cyclosporine treatment could suppress immunization and improve graft function in a xenogeneic hepatocyte transplantation model. METHODS: Free or encapsulated human hepatoma cells (HepG2) were cocultured for 28 days with splenocytes from Lewis rats or implanted for 60 days into the peritoneum of Lewis rats. RESULTS: Anti-HepG2 and antialbumin antibodies were detected in the supernatants of rat splenocytes that were cocultured with HepG2 cells and in the serum of rats that had undergone transplantation with HepG2 cells. Cyclosporine suppressed this antibody production both in vitro and in vivo. Human alpha-GST blood levels, which reflect hepatocyte injury, were low in cyclosporine-treated animals but high when encapsulated HepG2 cells were transplanted without cyclosporine therapy. Western blots revealed human albumin from day 3 to day 60 in the serum of rats treated with cyclosporine, but not after day 30 in untreated rats. CONCLUSIONS: Xenogeneic hepatocytes induce a humoral response that impairs their viability and function. A 2-week course of cyclosporine suppresses this immune response and improves graft function for up to 60 days. PMID- 10715987 TI - Sepsis after major visceral surgery is associated with sustained and interferon gamma-resistant defects of monocyte cytokine production. AB - BACKGROUND: Recent clinical trials failed to demonstrate beneficial effects of anti-inflammatory sepsis therapy. The present study therefore asked the following questions: Is there evidence for immunosuppression during postoperative sepsis? When, during the septic course, may immunosuppression develop? Can defective cellular functions be restored by in vitro treatment with interferon-gamma (IFN gamma)? METHODS: The study included 35 patients with sepsis after major visceral surgery and 85 control patients. Monocyte secretion of interleukin (IL)-1 beta, IL-12 p40 and p70, IL-18, tumor necrosing factor, and IL-10 with or without IFN gamma treatment and its correlation with the course and outcome of postoperative sepsis were determined. RESULTS: Postoperative sepsis was associated with an immediate defect of endotoxin-stimulated monocyte production of IL-12 p40, IL-1 beta, and IL-10 in both surviving and nonsurviving patients. During the final phase of postoperative sepsis, a significant recovery of IL-12 p40 and IL-1 beta secretion, but not of IL-10 production, correlated with survival. Despite the exposure of monocytes in vitro to IFN-gamma for 16 hours, the production of the biologically active IL-12 p70 heterodimer was severely suppressed both in survivors and nonsurvivors, although the secretion of the p40 subunit was restored. In contrast, IFN-gamma treatment resulted in a significant suppression of monocyte IL-1 beta production in all patient subgroups. Alterations of monocyte tumor necrosing factor secretion were not observed. The production of IL 18 was below the limits of detection in all samples. CONCLUSIONS: Postoperative sepsis was associated with immediate monocyte defects that affected both pro- and anti-inflammatory cytokine secretion, which suggests that immunosuppression is a primary rather than a compensatory response to a septic challenge. Sepsis survival correlated with the recovery of the proinflammatory, but not the anti inflammatory, response. The treatment of monocytes with IFN-gamma did not reconstitute defective proinflammatory cytokine production. PMID- 10715988 TI - Nicotine-induced smooth muscle cell proliferation is mediated through bFGF and TGF-beta 1. AB - BACKGROUND: Cigarette smoking influences and enhances the development of atherosclerosis. We investigated if nicotine, an important constituent of cigarette smoking, has a stimulatory effect on bovine smooth muscle cell proliferation in vitro through the mediation of bFGF and TGF-beta 1. METHODS: Bovine aortic smooth muscle cells (SMC) were stimulated with (-)-nicotine at various concentrations ranging from 6 x 10(-4) mol/L to 6 x 10(-8) mol/L. SMC viability and count were assessed. The presence of bFGF and TGF-beta 1 in serum free conditioned media was determined by the inhibition antibody-binding assay, and the mitogenic activity of (-)-nicotine on SMC was analyzed by the 3H thymidine uptake. Polymerase chain reaction was used to study the expression of bFGF and TGF-beta 1. RESULTS: The bFGF release after (-)-nicotine stimulation was greater than in the controls, whereas TGF-beta 1 release was lower. The greatest mitogenic activity was found at a (-)-nicotine concentration of 6 x 10(-6) mol/L. The addition of monoclonal antibody anti-bFGF decreased the 3H-thymidine uptake of SMC exposed to (-)-nicotine, whereas the addition of monoclonal antibody anti TGF-beta 1 increased the 3H-thymidine uptake of stimulated SMC. bFGF mRNA expression was significantly higher in SMC exposed to (-)-nicotine than in the controls, but TGF-beta 1 mRNA expression was significantly lower in SMC exposed to 6 x 10(-6) mol/L (-)-nicotine than in SMC treated with the other concentrations of (-)-nicotine and in controls. CONCLUSIONS: Nicotine is a potent regulator of bFGF and TGF-beta 1 production and release by aortic SMC, and it seems to play an important role in the development and progression of atherosclerosis and neointimal fibrous hyperplasia. PMID- 10715989 TI - Polymorphonuclear neutrophil (PMN) recruitment in a 2-front murine injury model: triage of PMNs to competing stimuli of recruitment. AB - BACKGROUND: Intensive care unit patients as a group have the highest rate of nosocomial infections, such as pneumonia, urinary tract infections, and wound infections. The triage of polymorphonuclear neutrophils (PMNs) during an acute inflammatory response was investigated to determine if the severity of injury or infection contributes to PMN delivery. METHODS: A murine cecal ligation and puncture-induced peritonitis model with polyvinyl sponge discs were used to collect the PMNs in the abdomen (primary site) and in the subcutaneous tissue of the dorsum (remote site). Eighty CD1 male mice--20 in each of 4 groups--were assigned to the following: cecal ligation and puncture (CLP), sham laparotomy with cecal manipulation (CM), polyvinyl sponge placement in the abdomen and back only (SP), and sponge placement in the back alone (CON [control]). After 24 hours, the sponges were harvested, and the PMNs were collected and counted on a hemocytometer. RESULTS: These data, reported as mean PMN cells x 10(5) +/- SEM, demonstrated that back sponges contained significantly fewer PMNs in the CLP group (3.29 +/- 1.1) than in the CM group (7.77 +/- 1.61, P = .04), the SP group (8.69 +/- 1.67, P = .01), and the CON group (11.04 +/- 1.91, P < .001). CONCLUSIONS: These results demonstrate that PMN delivery to sites of secondary injury are inversely correlated to the severity of the primary injury or peritonitis. PMID- 10715990 TI - Suppression of cellular immunity by surgical stress. AB - BACKGROUND: Suppression of cellular immunity is one of the host responses to surgical stress. In cancer patients this immunosuppression may accelerate the growth and metastasis of residual cancer cells, so it is desirable to restrict immunosuppression by surgical stress to a minimum. However, the extent and duration of immunosuppression caused by operations on gastrointestinal cancer, as well as the mechanisms involved, have not been determined. METHODS: To clarify these points, we investigated immunocyte function and measured the blood levels of hormones, cytokines, and acute phase reactants from before to after operation in 20 patients with stage I gastrointestinal cancer. RESULTS: In patients exposed to surgical stress, peripheral blood lymphocyte numbers and function were suppressed until at least 2 weeks postoperatively. This immunosuppression was mainly due to a decrease of helper-inducer T cells, cytotoxic T cells, natural killer cells, and interleukin-2 receptor-positive cells, as well as an increase of suppressor T cells. In addition, hypersecretion of cortisol and overproduction of immunosuppressive acidic protein were observed. CONCLUSIONS: Cellular immunosuppression by surgical stress was mainly due to an increase of lymphocyte subsets that depress cellular immunity coupled with a decrease of the subsets that promote it. Overproduction of cortisol and immunosuppressive acidic protein in response to surgical stress may play an important role in the development of immunosuppression. PMID- 10715991 TI - Somatic mutations of the multiple endocrine neoplasia type 1 (MEN1) gene in patients with sporadic, nonfamilial primary hyperparathyroidism. AB - BACKGROUND: Multiple endocrine neoplasia type 1 (MEN 1) is a syndrome with tumors of many endocrine tissues. Germline MEN1 gene mutations were found in most patients with familial or sporadic MEN 1. Recently, somatic MEN1 gene mutations were also detected in sporadic non-MEN 1 endocrine tumors. METHODS: We used direct sequence analysis to investigate MEN1 gene mutations in 30 parathyroid tumors obtained from 30 patients with sporadic, nonfamilial primary hyperparathyroidism. RESULTS: Four patients had somatic mutations of the MEN1 gene, comprising 1 small insertion (1091insAGC), one missense mutation (G42S), and 2 non-sense mutations (E388X, R460X). Identical missense and non-sense mutations were found in patients with familial and non-familial MEN 1. There were no differences between clinical features of patients with and without MEN1 gene mutations. CONCLUSIONS: The incidence of somatic MEN1 gene mutations (13.3%) in Japanese patients with sporadic, nonfamilial primary hyperparathyroidism is almost equal to those of such patients in the United States and Sweden. Occasionally, the MEN1 gene mutation sites in sporadic parathyroid tumors are identical to those reported in tumors from patients with familial or sporadic MEN 1. PMID- 10715992 TI - B7.1 costimulation increases T-cell proliferation and cytotoxicity via selective expansion of specific variable alpha and beta genes of the T-cell receptor. AB - BACKGROUND: Optimal T-cell activation requires not only ligation of the T-cell receptor (TcR) but also delivery of costimulatory signals by various accessory molecules. The interaction of the costimulatory molecule B7.1 (CD80) with its receptor CD28 provides a strong positive signal to T cells. METHODS: The B7.1 gene was transduced into cultured human ovarian, breast, and pancreatic tumor cells by using a retroviral vector. Autologous as well as allogeneic naive T cells were stimulated with either wild-type or B7.1-transduced tumor cells in a mixed lymphocyte tumor cell culture (MLTC). In addition to cytolytic activity, T cell proliferation, T-cell subset composition, and the frequencies of TcR variable (V) alpha and beta genes were compared in T cells from both types of MLTC. RESULTS: Introduction of the B7.1 gene into tumor cells was successful in all tumors to a varying degree. Those tumors expressing high levels of B7.1 induced significantly higher levels of T-cell proliferation than wild-type tumor cells. T-cell subset composition did not markedly differ between T cells stimulated with wild-type tumor cells or B7.1-expressing tumor cells. However, T cells stimulated with B7.1-expressing tumor cells showed a significantly increased cytolytic potential. The increased cytotoxic T lymphocyte activity was associated with a higher frequency of specific TcR V alpha and V beta genes. In addition, B7.1 costimulation promoted oligoclonality among the responding T cells. CONCLUSIONS: These data suggest that costimulation through B7.1 promotes T cell proliferation and cytotoxic activity through clonal expansions of T cells bearing antigen-specific TcR V alpha and V beta genes and through promotion of oligoclonality. The data also suggest that promoting B7.1-mediated costimulation is an important aspect of immune therapies. PMID- 10715993 TI - Sunday school. PMID- 10715994 TI - Enterocutaneous fistula 14 years after prosthetic mesh repair of a ventral incisional hernia: a life-long risk? PMID- 10715995 TI - Fatal bilhemia. PMID- 10715996 TI - An association of congenital mid-tracheal stenosis with Down syndrome. PMID- 10715997 TI - BDA accreditation causes concern. PMID- 10715998 TI - Use of terminology questioned. PMID- 10715999 TI - Dental fear in children--a proposed model. AB - Over the past eleven years, we have worked together to treat children who are dentally phobic. This has enabled us to develop an understanding of how children come to be dentally fearful. We have constructed a model of child dental fear which helps us in our work. It is important to acknowledge that fear is a normal phenomenon when any of us are exposed to threat. Helping dentally fearful children appraise or evaluate threat, face their fear and build upon their strengths is the task facing dentists and, occasionally, psychologists. The consequences for children of not doing so are extreme difficulty with accepting and ultimately total avoidance of treatment. Both of these can persist into adulthood. First, we propose to discuss the normality of fear in children, placing dental fear within a developmental context. We will then outline a model for assessing and treating dental fear which identifies five discrete but interrelated factors. Each of the factors and its treatment is illustrated with examples. PMID- 10716000 TI - Basic implant surgery. AB - Implant surgery protocols differ slightly with individual systems. However, basic surgical principles are required to ensure successful osseointegration of the implant in the correct location which allows good aesthetics and loading. PMID- 10716001 TI - The design and use of special trays in prosthodontics: guidelines to improve clinical effectiveness. AB - This paper looks at how carefully prescribed special trays can be helpful in everyday dental practice. Guidelines are suggested for the design of custom trays, that will, hopefully, lead to improvements in the quality of working impressions. PMID- 10716002 TI - How long do routine dental restorations last? A systematic review. AB - OBJECTIVE: To conduct a systematic review of the literature on the longevity of routine dental restorations in permanent posterior teeth, and to identify and examine factors influencing its variability. METHOD: Accepted guidelines were followed. An advisory group oversaw the project. Simple Class I and Class II amalgam, composite resin, glass ionomer and cast gold restorations were covered. Comprehensive searching of electronic databases, hand-searching, and location of 'grey' literature, generated 124 research reports. Those considered relevant were assessed for validity and quality according to agreed criteria. The analysis was descriptive. RESULTS: Eight of 58 relevant research reports were categorised, according to agreed criteria, as being of satisfactory validity and quality. They suggested that 50% of all restorations last 10 to 20 years, although both higher and lower median survival times were reported. The findings were supported by the totality of studies reviewed. However, variability was substantial. Restoration type, materials, the patient, the operator, the practice environment and type of care system appeared to influence longevity. CONCLUSIONS: Many studies were imperfect in design. Those considered to be the most appropriate for analysis were too limited to undertake a formal statistical exploration. Therefore there remains a need for definitive randomised controlled trials of restoration longevity, of sound design and adequate power, employing standardised assessments and appropriate methods of analysis. PMID- 10716003 TI - Referrals for dental general anaesthetics--how many really need GA? AB - AIM: To find out how many patients for whom dental general anaesthesia was requested actually needed it in order to complete treatment. DESIGN: Analysis of clinical outcomes supported by telephone canvassing of parents. METHOD: In summer 1998, eighty two child patients were seen in the Community Dental Service in Rochdale with a request for the provision of dental general anaesthesia (DGA) for the extraction of teeth. Their ages ranged from 3 to 14 years and all were required to attend for a pre-anaesthetic visit. Unless objective indicators of a need for DGA applied, the parents and children were actively discouraged from having DGA, and the alternative of local anaesthetic (LA) was offered. Clinical outcomes and parent satisfaction were recorded after treatment was finished. RESULTS: In 75% of cases it proved possible to complete the extractions without need for DGA; in the 10% of cases where DGA was necessary, it was to deal with the sequelae of dental caries. Fifteen percent of subjects failed to complete treatment. Subjects found to have a need for DGA tended to be younger and with treatment required in more than one sextant. Pain as a presenting symptom, young age and multiple treatment needs were found to be poor predictors of need for DGA and did not automatically preclude successful treatment without DGA. The satisfaction ascertained from users of the service was high and explanation of proposed treatments, especially the comparative risks and benefits of DGA versus LA, was well received. CONCLUSION: There is scope for significant reduction in provision of dental general anaesthesia if current professional guidelines are followed. PMID- 10716004 TI - A framework for the evaluation of continuing education short courses in dentistry. AB - The objective of this paper is to propose an evaluation framework for short courses in continuing education for general dental practitioners (GDPs) (so called, Section 63 courses). Existing monitoring and evaluation procedures in the West Midlands deanery were examined and an improved evaluation framework was then devised, piloted and revised. A 5 phase method was used incorporating the examination of existing practice (Phases 1 and 2), development of a new framework (Phase 3), piloting (Phase 4) and revision of the evaluation framework in the light of the pilot. This approach will be implemented in the West Midlands and may be adapted for national use (Phase 5). It was found that existing monitoring and evaluation was inconsistent in prevalence and scope. Those involved in short courses were in favour of a more consistent and visible evaluation, including some assessment of impact-on-practice and cost-effectiveness. In conclusion, meaningful evaluation needs to include four key processes: data gathering; data analysis; dissemination and, action planning (reviewing provision in the light of the data analysis). Thus, this evaluation framework feeds into a quality development cycle designed to ensure high quality and relevant short course provision for general dental practitioners. PMID- 10716005 TI - Understanding HIV risks of chronic drug-using men who have sex with men. AB - Focus groups and individual structured interviews were conducted in six cities with 98 predominantly street-recruited men who had a recent history of smoking crack or injecting drugs and who reported having had sex with other men (MSM) in the past year. Twenty-six focus groups explored the cultural and social context of participant's drug use and sexual activity and addressed outreach and HIV prevention issues pertinent to this population. Narrative summaries developed from verbatim focus group transcripts identified seven themes: (a) sexual orientation and gender identity; (b) interactions within and between MSM networks; (c) drug use, sexual activity and personal relationships; (d) HIV transmission bridges; (e) preferred HIV information sources; (f) HIV knowledge, prevention practices and risk behaviours; and (g) availability of HIV and drug related services. Of the 98 MSM drug users, 42% identified publicly as gay or homosexual; 35% identified publicly, but only 21% privately, as heterosexual. A total of 51% had one or more female sex partners in the past year. There was a high frequency of unprotected sex in conjunction with drug use and a distinct preference for having sex when high. For most participants, drug use rather than sexual orientation formed the core of personal identity. Participants reported associating primarily with other drug users, usually MSM, and had limited contact with people who did not use drugs and the mainstream gay community. Participants' sexual and drug-injecting activities were judged to be a bridge for transmission of HIV to both people who used drugs and those who did not. PMID- 10716006 TI - Differences in sexual behaviour between HIV-infected pregnant women and their husbands in Bangkok, Thailand. AB - In a Bangkok antenatal clinic, we interviewed 102 HIV-infected pregnant women and their husbands, 30% of whom were HIV-negative. We evaluated these data by matched and unmatched analysis, compared men and women in stable couple relationships on a number of sociodemographic and risk factor indicators and investigated further whether there were any differences in sociodemographic or risk factor profiles between HIV-serodiscordant couples and seroconcordant couples. When compared to wives, more of the husbands were working (p = 0.001), earning more money (p = 0.001), had had more than two sex partners (p = 0.001) and had had syphilis (p = 0.001). Serodiscordant couples did not differ greatly from seroconcordant couples except that women married to HIV-negative men were more likely to have been divorced or separated than their husbands which was not the case for women married to HIV-positive men (p = 0.02). There was poor agreement between husband and wife reports of husband risk behaviour and this did not differ between concordant and discordant couples. These findings suggest that assessment of risk and counselling of Thai women is incomplete without information on the HIV status and risk behaviour of her partner. Prevention strategies to decrease heterosexual transmission among couples need to target both the man and the woman. PMID- 10716007 TI - HIV testing practices of Zimbabwean physicians and their perspectives on the future use of rapid on-site tests. AB - To improve HIV testing procedures, rapid on-site HIV tests have been introduced in Zimbabwe. At present, little is known about physicians' perspectives on the potential use of rapid tests in their clinics or about their current laboratory based testing practices. In a sample of 63 general practitioners in Harare, this study found physicians were generally testing individuals, not couples, and an important reason for suggesting a patient be tested was medical symptoms; frequent reasons for patients requesting the test were insurance purposes, being about to get married or having suspicions about a partner. A primary deterrent to physicians testing patients, even when patients requested it, was fear of traumatizing them. Fifty-six per cent of the physicians believed rapid tests would increase the number of HIV tests they performed; significant associations were found between this belief and whether physicians ever chose not to test patients they suspected were HIV-positive (a positive association) and whether they chose not to test specifically out of fear that patients would commit suicide (a negative association). Prior to any expansion of testing with rapid tests, training in counselling and confidentiality measures is essential, given that over half the medical personnel providing counselling to these physicians' patients had received no training in pre- and post-test HIV counselling. PMID- 10716008 TI - Effects of ZDV-based patient education on intentions toward ZDV use, HIV testing and reproduction among a US cohort of women. AB - This study examined the immediate effects of exposure to a patient education brochure concerning the risks and benefits of zidovudine (ZDV) therapy during pregnancy to reduce perinatal HIV transmission (protocol ACTG 076) on related knowledge, behavioural intentions and attitudes of women with and at-risk for HIV infection. Self-reports were collected from 653 women of childbearing age from community family planning clinics and hospital-based HIV centres in 19 sites from nine US cities between May and November 1995. The intervention was a nine-page patient education brochure in Spanish, Creole and English versions, evently presenting the pros and cons of ZDV therapy to reduce perinatal HIV-transmission. Brochure exposure increased knowledge (p < 0.001) for all but one scale concerning ZDV resistance and increased the likelihood of women reporting intentions to take ZDV during pregnancy (p < 0.001) and to believe ZDV reduced transmission (p < 0.001). Brochure exposure had differential effects for some subpopulations. Intentions to have or terminate current or future pregnancies, knowledge about ZDV and attitudes toward ZDV varied mostly by ethnicity/race, language preference and HIV status. Pregnancy status, age, education and having an HIV-positive child had less impact on the brochure's effect, while income had no impact. PMID- 10716009 TI - Women's attitudes to condoms and female-controlled means of protection against HIV and STDs in south-western Uganda. AB - The consistent and correct use of the male condom makes it highly effective in both disease prevention and as a contraceptive method. However, it is also well recognized that its use is under men's control. Because of this vital limitation, there have been frequent calls for female-controlled methods of HIV prevention, particularly from women from sub-Saharan Africa. Here we report on data collected in focus-group discussions (FGDs) with women aged 17-54 in South-Western Uganda. A total of 138 women, from rural villages, urban family planning clinics and a truck-stop town, were recruited to participate in 18 FGDs on the male condom, the female condom and existing formulations of vaginal microbicidal products. Three themes emerged: (i) problems with men's control over the male condom, (ii) the importance of control over and secrecy about protective measures and (iii) sexual pleasure associated with different methods. We found that the female condom, while being perceived as an improvement over the male condom, was recognized as having limited value because of the need to agree its use prior to sex taking place. Other products were considered to be significantly better than the female condom; the sponge, in particular, was perceived as having advantages over every other product. Women like the fact that it could be inserted some time before, and left in place some time after, sexual intercourse, that it was effective for multiple instances of intercourse, and that men would be unaware that it was being employed. Female-controlled methods to prevent sexually transmitted infections, including HIV, and to increase reproductive choice, hold the promise of ceding some control over sexual and reproductive health to women. PMID- 10716010 TI - The impact of British and US guidelines for initiating combination antiretroviral therapy on estimating national treatment requirements for HIV-positive patients. Collaborating Immunologists and the Scottish HIV and AIDS Group. AB - This paper uses comprehensive national data on HIV positive patients in Scotland to carry out a needs assessment exercise for combination antiretroviral therapy. The objective of this study was to estimate the numbers of HIV positive patients in Scotland who would be eligible for combination antiretroviral therapy under current British and US guidelines and to demonstrate the impact of these different guidelines on the resources required. The proportion of the Scottish population that would be eligible for combination therapy ranged from 76% to 91%, under different guidelines for initiating therapy. We thus estimate that for countries such as Scotland, including western Europe and the United States, where a large proportion of the HIV population became infected in the early to mid 1980s, the majority of patients will be eligible for combination therapy, regardless of the guideline. PMID- 10716011 TI - Adherence to antiretroviral therapy in HIV-infected children in Italy. AB - Adherence to antiretroviral therapy is a major problem in children with HIV infection, who depend on parents or foster parents for receiving drugs. During an ongoing investigation on intestinal function in children with symptomatic HIV infection who were treated with zidovudine, blood samples were obtained six hours after the administration of zidovudine as reported by the parents and, again, one and six hours after its administration in the hospital, and drug concentration was measured by radioimmunoassay. Both peak and steady state zidovudine levels were within the expected concentration ranges after administration in the hospital. In contrast, they were below the effective concentration in five of the 10 children that reportedly had received the drug at home by the parents. These data directly show poor compliance with antiretroviral therapy in children. Compliance with antiretroviral therapy should be carefully checked in children and strategies are needed to increase full and permanent adherence with antiretroviral therapy by people in charge to administer drugs to HIV-infected children. PMID- 10716012 TI - Orphans of the AIDS epidemic in the United States: transition-related characteristics and psychosocial adjustment at 6 months after mother's death. AB - This study has two purposes: (1) to describe the characteristics related to the transition to orphanhood for children whose mothers die from AIDS and (2) to examine the psychosocial adjustment of these children at six months following maternal death. Twenty orphans and a control sample of 40 children from the same neighbourhoods, as well as their mothers or care-givers, served as participants. Two assessments occurred: (1) prior to the death of the mother in the orphan group and (2) six months after her death. The results indicated that relatives, particularly maternal grandparents, became the new care-giver of the orphans, no more than one residential move had occurred following the mother's death, and the new care-givers were providing a stable home environment. Child psychosocial adjustment did not change following maternal death. PMID- 10716013 TI - Predictors of dropout and burnout in AIDS volunteers: a longitudinal study. AB - Burnout among HIV/AIDS volunteers contributes to the loss of dedicated personnel resulting in strain on the HIV/AIDS care system. Past research has suggested that there were significant stresses and burnout associated with AIDS caregiving. We investigated the predictors of dropout in AIDS volunteers over time, and specifically, which of the variables of the stressors and rewards of being a volunteer (collected at baseline) predicted who would drop out two years later. The volunteers were the subjects of Nesbitt et al. (1996), who were members of an interfaith religious-based organization in Houston, Texas. The subjects were re contacted by mail after two years, and 76 of the 174 respondents completed a brief questionnaire which gave details of current volunteering activity, reasons for dropout (if they had dropped out) and completed the Texas Revised Inventory of Grief (TRIG). Forty dropped-out from volunteering while 36 continued. Data show the independent variables of total stressor score, the Maslach Burnout Inventory score of Depersonalization intensity and the three subscale scores involving stress: client problems and role ambiguity, emotional overload and organizational factors as being significant in predicting dropout in HIV/AIDS volunteers over time. The best predictors of the dropping-out of HIV/AIDS volunteers can be divided into the stresses (client problems and role ambiguity, emotional overload and organizational factors) and depersonalization intensity. The results showed that volunteers who experienced more client problems and role ambiguity, more emotional overload and more problems with organizational factors are more likely to drop out from the volunteer programme. They also show that the dropout volunteers have a significantly higher level of depersonalization intensity than the continuing volunteers, with the risk of dropout increasing by almost a third in the highest tertile of depersonalization intensity scores compared with those with lower scores. These data indicate that it is the stressors of AIDS volunteering, including the intensity of depersonalization, which lead to dropout, and that rewards do not appear to have a protective effect. PMID- 10716014 TI - HIV prevention with young men who have sex with men: what young men themselves say is needed. Medical College of Wisconsin CITY Project Research Team. AB - Young men who have sex with men (YMSM), and particularly ethnic minority YMSM, experience high incidence HIV infection due to continued patterns of high-risk sexual behaviour. The intent of this research was to systematically solicit input and recommendations from YMSM themselves concerning the kinds of HIV prevention programmes that would best meet their needs and would address risk issues they believed are critical. In-depth qualitative interviews were conducted with a sample of 72 purposively selected YMSM to identify necessary components of HIV prevention targeting YMSM. Respondents noted a need for comprehensive HIV prevention programmes that addressed issues related to dating and intimacy, sexuality and arousal, drugs and alcohol, self-esteem and self-worth, abuse and coercion, and sexual identity. Respondents emphasized the importance of keeping programmes confidential, fun, comfortable, accepting and open to all YMSM regardless of sexual identity. Identified community resource needs included safe havens for youth, more peer educators and older MSM mentors, increased school based sexuality education, and greater support from the society at large as well as from churches, the gay community and communities of Color. Implications of these findings for HIV prevention are discussed. PMID- 10716015 TI - Preventing HIV infection through peer education and condom promotion among truck drivers and their sexual partners in Tanzania, 1990-1993. AB - HIV prevention through peer education and condom promotion among truck drivers and their sexual partners is described. Trends during an initial 18-month intensive phase, followed by a 24-month maintenance phase, were monitored with surveys. Trends for self-reported condom use were: increase among men (56 to 74%) during the first phase with a decrease (72%) during the maintenance phase. Respective figures for women were 51%, 91% and 70%. Multivariate analyses revealed that men most likely to report using condoms were unmarried, had children, were more educated, had previously reported a genital ulcer, and perceived themselves at risk for HIV infection (OR = 1.95-3.47). Women tending to use condoms were unmarried, aware of the limitations of condoms, not in denial as to the existence of HIV, harboured inaccurate information about HIV transmission and were afraid (OR = 1.35-2.52). Both sets of results suggest that the most sexually experienced men and women who did not have a permanent stable relationship and who perceived themselves at risk, were most likely to use a condom. Peer education was an effective tool for increasing knowledge and encouraging appropriate behaviour change. It was most effective as an intensive high-input intervention and sustainable with the relatively stable population of truck drivers. PMID- 10716016 TI - Capturing the paradigm shift in HIV treatment: changing attitudes in the choice of combination antiretroviral drugs by high HIV caseload Australian GPs (1996 1997). AB - The use of combination antiretroviral therapy for HIV infection is a rapidly changing field. To assess the impact of recent studies on prescribing patterns, two surveys of 21 high HIV caseload Australian GPs were undertaken in June 1996 and June 1997 to plot changes in the choice of combination antiviral therapy. Of the 17 GPs who responded to the survey in each year, the number of HIV-infected patients seen at their practices were estimated to be 5,061 in 1996 and 5,912 in 1997. In 1996, 40% of their patients were estimated to be on antiretroviral therapy compared to 60% in 1997 (p < 0.05). In 1996, most GPs preferred using dual combination therapy (59%); whereas in 1997, triple combination therapy was preferred (82%). Between 1996 and 1997, there was a significant change by high caseload Australian GPs in the choice of antiretroviral drugs with many combinations being preferred prior to presentation of efficacy data for those combinations, or recommendation through national guidelines. PMID- 10716017 TI - Social and behavioural factors associated with HIV seroconversion in homosexual men attending a central London STD clinic: a feasibility study. AB - An unmatched retrospective case control study was conducted to test the feasibility of investigating social and behavioural factors which may have contributed to recent HIV seroconversion in a group of homosexual men. Participants, recruited from a London sexually transmitted disease (STD) clinic, were sexually active and had had a negative HIV test with a subsequent test (positive (cases) or negative (controls)) within three to 15 months. Twenty cases and 22 controls were recruited between February and October 1995. There was no difference between cases and controls in: the number of regular or casual sexual partners, the proportion who were unaware of their regular partners' serostatus (cases 60%, controls 59%), or the proportion who had known HIV-positive regular partners (cases 20%, controls 23%). A significant difference in sexual behaviour was found only when the HIV status of partners, if known, was taken into account: cases were more likely than controls to have had unprotected receptive anal intercourse with a partner not known to be HIV-negative (OR = 5.5, CI = 1.15 29.50). Fifty per cent of the cases and 27% of the controls acquired acute STDs between the two HIV tests. All participants achieved high self-efficacy scores, but the controls believed their peers placed a greater value on safer sex. Cases cited emotional issues and the use of drugs and alcohol as contributing to their seroconversion, whereas controls cited a commitment to safer sex and the avoidance of high-risk situations as contributing to their remaining HIV negative. The results illustrate the importance of acknowledging the concept of 'negotiated safety' in studies of sexual behaviour; seroconversion was only associated with unprotected sex with a partner not known to be HIV-negative. Despite high self-efficacy scores, indicating the skills to negotiate safer sex, high levels of unsafe anal intercourse were reported. Differences between cases and controls included the importance of safer sex, periods of emotional vulnerability, influence of peers and the appropriate use of condoms. There is a need for these results to be confirmed in a larger and more powerful study. PMID- 10716018 TI - HIV-infected adolescent and adult perceptions of tuberculosis testing, knowledge and medication adherence in the USA. AB - HIV-infected adolescent and adult perceptions of tuberculosis (TB) infection rates and physician TB behaviour, and patient knowledge of TB transmission and treatment adherence were assessed. HIV-infected youth (N = 199) from adolescent clinical care sites in three cities and HIV-infected adults (N = 133) in New York were interviewed. Adolescent self-report was compared to medical chart review. Adolescents reported they were significantly less likely to be tested, although testing rates were high for both samples. Approximately 9% of both samples reported infection with TB; the majority of whom reported receiving medication (97%), and consistent medication adherence (93%). The overall mean knowledge score was 66%, with significant age differences: adolescents were less knowledgeable than adults, and young males tended to be less knowledgeable than young females. Age, gender and experience with TB (self-perception of TB, testing history and clinic choice) significantly predicted accuracy of knowledge about TB. Results suggest that if HIV-infected individuals--a population at very high risk and often among the least able to afford health care resources--receive the education and support they need from their community health care sources they may substantially reduce their chances of contracting and spreading TB. PMID- 10716019 TI - HIV infection and patterns of risk among women drug injectors and crack users in low and high sero-prevalence sites. AB - As AIDS cases among US women continue to increase, a better understanding of women's behavioural risk patterns is needed to inform intervention efforts. Data were from 2,945 women drug injectors and crack users. Statistical analyses compared sociodemographic variables, lifetime behavioural risk patterns, HIV sero prevalence and history of sexually transmitted diseases, and determined predictors of HIV infection separately in 16 low and four high sero-prevalence sites. Based on risk patterns, four behaviourally-defined sub-groups were constructed, and rates of HIV sero-prevalence were compared. In comparisons between low and high sero-prevalence sites, there were significant differences on most variables examined, and in the relative importance of the sociodemographic characteristics and risk patterns predicting HIV. Drug injection and sex exchange were each independent, significant, behavioural predictors of infection, with no significant difference between the odds ratios attributed to each predictor. HIV sero-prevalence was significantly different among four sub-groups. Interventions must be tailored to address observed differences among women in low and high sero prevalence sites. Injection drug use and exchanging sex each play a major role in the transmission of HIV infection to US women. Prevention efforts targeted at women should address differences in behavioural risk patterns. Aggressive and innovative interventions are needed for women who exchange sex. AIDS research must investigate how socioeconomic factors impact women's risk for HIV infection. PMID- 10716020 TI - Gender differences in sharing injecting equipment by drug users in England. AB - The study investigates whether the higher rates of sharing needles and syringes reported by female injecting drug users (IDUs) also occur in sharing other types of injecting equipment. Structured interviews were carried out with 181 IDUs in two cities (100 in Bournemouth, 81 in Bath), with almost equal numbers of males and females in each sample, recruited through needle exchanges and 'snowballing'. Almost all (92%) had shared some equipment in the previous six months: 40% had shared syringes in the month before interview. Several methods of assessing sharing found that women received previously used injecting equipment significantly more often than men. Significantly more males had passed on equipment other than syringes in the previous six months. Fine-grained analyses of 547 injecting episodes found that women received needles and syringes, and syringes significantly more often than did men. The pattern of gender differences reported suggests that women are at higher risk of blood borne viral infections because they receive more types of used equipment and do so more frequently. These results have implications for practice and research. PMID- 10716021 TI - Doctors' attitudes to the care of children with HIV in South Africa. AB - A descriptive survey was conducted on the attitudes of medical staff to caring for HIV-infected children, in three teaching hospitals in Cape Town, South Africa. The study was designed to determine whether the knowledge of a patient's HIV-positive status affects the doctor's attitude and management, to determine doctors' perceived competence with regard to the management of paediatric AIDS and to identify their major concerns in the management of paediatric patients with HIV infection. The response rate was 86%, including 89% of consultants, 81% of registrars and 87% of medical officers employed at the hospitals during the period July/August 1996. The study has highlighted that doctors working in a situation where the epidemic has only recently emerged perceive themselves as being inadequate with regard to managing HIV infection in children. There are indications that doctors may be influenced by the HIV-positive status of children when making decisions regarding their medical management. One of the major concerns with regard to the management of patients with HIV, expressed by doctors in the study, was the lack of management protocols and policy guidelines. PMID- 10716022 TI - The effect of advance care planning on completion of advance directives and patient satisfaction in people with HIV/AIDS. AB - The effects of advance care planning are poorly understood. The purpose of this study was to evaluate the effect of an advance care planning intervention on the completion of advance directives (ADs) and patient satisfaction. A volunteer sample of persons with HIV/AIDS received advance directive documents, watched an educational video and received individual counselling on completing an advance directive during three face-to-face interviews over approximately six months. The advance care planning intervention was associated with an increase in advance directive completion rates from 16.4% to 40.7% (p = 0.001), but 23.1% of advance directives reported as completed were legally invalid. There was a trend towards decreased overall patient satisfaction with health care (p = 0.07). Advance are planning increases the rate of AD completion but many 'completed' advance directives are legally invalid. Advance care planning did not improve patient satisfaction with health care. PMID- 10716023 TI - Asian paralysis syndrome. AB - We report 20 children admitted to the paediatric ward of a public general hospital for acute flaccid paralysis, which was bilaterally symmetrical in all cases and was associated with bulbar involvement in eight of them. Recovery was partial. Nerve conduction studies showed motor axonal neuropathy. This new disease, variously termed as non-inflammatory neuropathy/Chinese paralysis syndrome must be differentiated from Guillain-Barre syndrome (GBS) and poliomyelitis. Both GBS and Asian paralysis syndrome have bilaterally symmetrical flaccid paralysis but GBS tends to have sensory involvement, full recovery occurs in 90% of cases and nerve conduction shows demyelinating neuropathy. Asian paralysis syndrome and poliomyelitis are pure motor lesions without sensory changes and partial recovery, but poliomyelitis differs in that paralysis is asymmetrical and unequal, muscle spasm is always present in the initial stage and there are prodromal symptoms. Nerve conduction studies show anterior horn cell disease. This new entity, common in Asian populations, assumes public health importance when it mimics poliomyelitis in a country that has tried to eliminate poliomyelitis by universal immunization. To the best of our knowledge, this is the first report of Asian paralysis syndrome in children in our area. PMID- 10716024 TI - Neurological features of cerebral malaria in Nigerian children. AB - Cerebral malaria is one of the commonest causes of an acute neurological syndrome in malaria-endemic areas. However, there are few detailed reports of findings on clinical neurological examination of the condition. The neurological features of cerebral malaria in 103 children aged 5 years or less were studied in Ibadan, Nigeria, an area of high malaria transmission. The correlation of these features with prognosis was also studied. Convulsions occurred in 87% of subjects and were in most cases of a generalized tonic-clinic nature. Abnormalities of posture were observed in 41%, abnormal tone in 70% and abnormal deep tendon reflexes in 74%. Absent corneal reflexes were found in about 14%. The time interval between the last seizure episode and presentation in hospital, abnormal posture (decerebrate or decorticate), absence of corneal reflex and depth and duration of coma were indicators of poor prognosis. In this study, cerebral malaria presented with non specific features of diffuse, symmetrical, upper motor neurone dysfunction, and some specific neurological features were associated with poor prognosis. It is important that cerebral malaria be considered in any child with features of acute encephalopathy in a malaria-endemic area. Careful clinical examination of such children is essential as neurological features of the condition may provide a clue to prognosis. PMID- 10716025 TI - Biochemical and haematological variables in Gambian children with cerebral malaria. AB - Biochemical and haematological measurements were made in Gambian children who satisfied the criteria for the diagnosis of cerebral malaria over a 3-year period. Biochemical and haematological values were available for 388 and 624 children, respectively. Biochemical signs of renal and hepatic dysfunction were found and these may have contributed in a cumulative way to the high mortality seen in the study children. Cerebral involvement in children with cerebral malaria is only one, though the most important, manifestation of a multi-organ disease. PMID- 10716026 TI - Laryngeal swabs for diagnosing tuberculosis. AB - The value of smear and culture of laryngeal swabs as a method of confirming pulmonary tuberculosis was investigated in Indian children. A total of 116 children with 'suspected' tuberculosis had a Mantoux test and chest X-ray. Of these, 51 had a positive Mantoux and/or chest X-ray compatible with tuberculosis, and this group had two laryngeal swabs taken on each of 3 consecutive days. The Mantoux test was positive in 37 (73%) of the 51 'probable' cases. Chest X-ray was abnormal in 36 (71%) cases and compatible with tuberculosis in 20 (39.7%). Mycobacterium tuberculosis was cultured from laryngeal swabs in 14 (28%) children and in another three children smears were positive but culture-negative. The overall confirmation rate for tuberculosis was 33%. Laryngeal swabs are a simple method of confirming tuberculosis and may be undertaken in out-patients. PMID- 10716027 TI - A high prevalence of metabolic bone disease in exclusively breastfed very low birthweight infants in Dar-es-Salaam, Tanzania. AB - Metabolic bone disease (MBD), or rickets, is common in very low birthweight infants. A descriptive, cross-sectional, hospital-based study was carried out at Muhimbili Medical Centre, Dar-es-Salaam from 15 April to 30 June, 1995 to discover the magnitude, contributory factors, morbidity and suitable biochemical diagnostic tests for MBD. One hundred infants with a postnatal age of 6-12 weeks, whose birthweights were 1500 g or less were studied. Thirty-three of 100 (33%) infants, 16 boys and 17 girls, were radiographically diagnosed as having metabolic bone disease. The mean (SD) gestational age of those infants was 30.4 (2.7) weeks, while that of the infants without metabolic bone disease was 32.4 (3) weeks (p = 0.003). There was no significant difference in birthweight, serum calcium and serum phosphate levels between those infants with MBD and those without. The mean (SD) serum alkaline phosphatase in infants with MBD was 1052.9 (493.3) U/l and 766.8 (301.7) in those without MBD (p = 0.006). Thus, metabolic bone disease is common in very low birthweight infants. Wrist radiography and serum alkaline phosphatase levels remain important diagnostic tools. MBD should be considered seriously in very low birthweight infants. PMID- 10716028 TI - Hyperekplexia: a rare differential of neonatal fits described in a developing country. AB - Hyperekplexia is a rare condition in which there is an exaggerated startle response. We report how a case presented in Papua New Guinea (PNG) and was diagnosed with international support. This is the first reported case in PNG. It is an important diagnosis to make to prevent sudden death and inappropriate treatment. The case illustrates the benefit of having a link with an international specialist and we discuss the importance of communication between developing and industrialized countries. PMID- 10716029 TI - Maternal and neonatal anthropometry. AB - Anthropometric measurements were assessed in 434 Brazilian mother-baby pairs (263 appropriate-for-gestational-age [AGA] and 171 intrauterine growth-retarded [IUGR]) to compare their distribution and to evaluate associations in AGA and IUGR pairs. Mothers who delivered IUGR babies were thinner and shorter than mothers of AGA babies; the cut-off points of risk for delivering an IUGR baby were 50 kg for weight (OR = 3.8, p < 0.001) and 150 cm for height (OR = 3.6, p < 0.001). IUGR mothers also tended to gain less weight in pregnancy than AGA mothers, presenting a risk 6.1 times higher for weight gain < or = 7 kg (p < 0.001). There were weak though statistically significant correlations between AGA mother-baby pairs, and a few weak correlations between IUGR mother-baby pairs. The larger number of statistically significant correlations between anthropometric measurements in AGA mother-baby pairs than in IUGR pairs shows that in this region of the country, where maternal malnutrition has a low prevalence, probably other factors are associated with IUGR. It seems that the influence of maternal nutrition on a baby's size at birth is more important in populations with moderate-to-severe malnutrition. In Brazil, as in some other developing countries, overweight is becoming an important issue and the prevalence of malnutrition has decreased. In this study, there were few mothers (n = 17) with a body mass index (BMI) < or = 20. On the other hand, there were many (n = 209) overweight (BMI 25-30) and obese (BMI > 30) mothers. We advise further large epidemiological studies to assess the diet of pregnant women and its relationship to maternal weight, weight gain and low birthweight (particularly IUGR) in countries with a considerable prevalence of maternal undernutrition and maternal overweight/obesity. PMID- 10716030 TI - Neuroblastoma in southern Africa: epidemiological features, prognostic factors and outcome. AB - We retrospectively analysed the epidemiological features and the importance of biochemical, histological and genetic parameters in predicting survival in 14 Namibian and 34 South African children treated for neuroblastoma (NB) from 1983 to 1997. Curative treatment consisted mainly of total (13%) or partial (44%) resection after chemotherapy (cyclophosphamide and doxorubicin x6 courses or carboplatin, etoposide, epirubicin and cyclophosphamide x6 courses). Localized radiotherapy with curative intent was given to 33% of patients. The male:female ratio was 0.9. The median age was 18 months (range 1-116) and was comparable in white, black and mixed ethnic patients. Primary disease was located in the abdomen (75%), thorax (15%), pelvis (5%) or elsewhere (5%). Evans stage distribution was: stage I, 2%; stage II, 19%; stage III, 21%; stage IV, 50%; and stage IVS, 8%. Stage III/IV disease was more common in black than in white children (p = 0.0001). Urinary vanillyl mandelic acid was elevated in 63% of those tested. Survival after 5-163 months' follow-up was 90% for stages I and II combined (median 2983, range 798-4661 days), 51% for stage III (median 367, range 61-5001 days), 6% for stage IV (median 227, range 20-4379 days) and 50% for stage IVS (median 532, range 54-1543 days). All seven children with para-spinal tumours survived. Individual factors associated with significantly poorer survival were elevated serum lactate dehydrogenase (p < 0.001), Joshi histological risk categorization adapted for age (p = 0.039), n-myc amplification (p = 0.006) and diploidy or tetraploidy (p = 0.006). All seven children with serum ferritin exceeding 149 ng/ml at the time of diagnosis died and survival was 33% in children with 1p deletion and 67% in those without, but the numbers were too small to achieve significance. These findings confirm the benefit of simple biochemical tests and histology in identifying those who are likely to respond favourably to conventional chemotherapy and surgery. Supportive genetic tests on formalin-fixed paraffin-embedded tumour tissue contributed to predicting outcome in 21 patients. PMID- 10716031 TI - Management of childhood fractures in Anjouan Island, Comoros. AB - Limb fractures were reviewed in 489 children, 390 boys and 99 girls whose ages ranged between 2 days and 14 years (mean 8 yrs). Only 54 (11%) were caused by road traffic accidents; 435 (89%) were due to other causes. Fractures of the arm were seen in 332 (68%) children and there were leg fractures in 157 (32%), of which 155 (32%) were undisplaced and were treated by first aid. Closed reduction was performed on 138 (28%) children and 196 (40%) were treated by internal fixation. Infection and non-union occurred in 28 (14%) and 13 (7%), respectively. Re-operation was performed because of complications in 21 (11%). PMID- 10716032 TI - The impact of breastfeeding on serum electrolytes in infants hospitalized with severe dehydrating diarrhoea in Yemen. AB - The effect of breastfeeding on serum electrolytes and case fatality was studied in a group of 430 children admitted with severe dehydrating diarrhoea. Hyponatraemia and hypokalaemia were significantly more prevalent in infants who were exclusively bottle-fed (37.3% and 46.3%) compared with exclusively breastfed (12.2% and 16.7%) and among bottle-fed weaning children (46.3% and 62.6%) compared with weaning children who continued to breastfeed (24.7% and 36.7%, respectively). Mortality was lower in exclusively breastfed children (4.4%) than in those receiving formula feeds (16.4%); it was also lower in weaning children who continued to receive breast-milk (6%) than in bottle-fed weaning children (13.8%). Breastfeeding significantly reduces case fatality and the likelihood of electrolyte disturbances among infants hospitalized with severe dehydrating diarrhoea. PMID- 10716033 TI - Purulent pericarditis: clinical profile and outcome following surgical drainage and intensive care in children in Chandigarh. AB - Purulent pericarditis, though rare in developed countries, is not uncommon in developing countries. However, the type of pericardial drainage required and the risk of subsequent constrictive pericarditis has not been clearly defined. Thirty children between the ages of 3 months and 12 years with a diagnosis of purulent pericarditis were studied retrospectively. Pericardial effusion was confirmed in all by echocardiography and the diagnosis of bacterial pericarditis was based on aspiration of purulent fluid with leucocytosis and high proteins. Purulent pericarditis was a part of the disseminated sepsis in 25 (83%) children. Fever was present in all, hepatomegaly in 28 and breathlessness in 25, whereas muffled heart sounds, raised JVP and pericardial rub were found in only 18, 16 and 7, respectively. The ECG was abnormal in only 16 children. Staphylococcus aureus was the causative organism in 24 (96%). Open surgical drainage was done in 26 children, 23 of whom underwent anterior pericardiectomy. Two children died of disseminated sepsis. None of the 21 who returned for follow-up for periods of between 4 and 24 months had any long-term sequelae. PMID- 10716034 TI - Geographical comparison of the prevalence of childhood asthma and allergies in Singapore. AB - A previous study suggested that differences in the prevalence of respiratory illnesses such as asthma in school children in different regions of Singapore were not due to the influence of air pollution or environmental factors but possibly to cultural and socio-economic factors. The effects of socio-economic or demographic variables were, however, not shown in that study. In this study, we set out to discover whether regional differences in the prevalence of atopic diseases such as asthma, rhinitis and eczema in Singapore school children could be explained by different demographic profiles. The prevalence of asthma and allergies were evaluated in 6238 Singapore school children in two age groups (6-7 years [n = 2030] and 12-15 years [n = 4208]). They were from four regions, based on residential post codes. Demographic and socio-economic data were also obtained. The questionnaire of the International Study on Asthma and Allergies in Childhood (ISAAC) was used. The data showed that children residing in the northern regions of Singapore had a significantly lower prevalence of asthma and rhinitis than those in other regions. When controlled for demographic influences (age, sex and race) and socio-economic factors (type of housing), however, the differences between these regions were reduced. No geographical difference in the prevalence of eczema was observed. Thus, geographical differences in the prevalence of asthma and rhinitis in Singapore could in part be explained by demographic and socio-economic differences in the population. PMID- 10716035 TI - Acute pancreatitis with cholestatic hepatitis: an unusual manifestation of hepatitis A. AB - Acute hepatitis A infection is an uncommon cause of pancreatitis in children. To date, only four cases have been reported in the paediatric literature. We report a 7-year-old girl with acute pancreatitis associated with hepatitis A infection who made a satisfactory recovery. The report highlights the CT findings including focal necrosis not previously reported. Because of the extreme rarity of the complication, the four previous reports have also been single case reports. This paper reviews all these cases with a view to elucidating the aetiopathogenesis of the pancreatitis. PMID- 10716036 TI - Recurrent bacterial meningitis: report of two cases from Riyadh, Saudi Arabia. AB - We report two cases of recurrent bacterial meningitis after head injury in two Saudi boys. The brain CT scan showed bony defects in both despite normal otolaryngeal clinical findings. One child remained well after surgical repair but the other was lost to follow-up. PMID- 10716037 TI - A case report of spherocytosis presenting with choledocholithiasis in early childhood and a review of the literature. AB - This is a report of spherocytosis presenting unusually early with choledocholithiasis secondary to low grade haemolysis. PMID- 10716038 TI - Evidence for the biosynthesis of A glucuronide conjugate of (S)-(-)-nicotine, but not (S)-(-)-cotinine or (+/-)-trans-3'-hydroxycotinine by marmoset hepatic microsomes. AB - Recently, the detection of urinary glucuronide conjugates of nicotine and its two major metabolites, trans-3'-hydroxycotinine and cotinine, showed that glucuronidation is an important pathway of nicotine metabolism in humans. (S)-(-) Nicotine-N(+)-1-beta-glucuronide (quaternary N-glucuronide with linkage through the pyridino-nitrogen of nicotine) was shown to be an important nicotine metabolite of humans in vivo. The present study was undertaken to develop an animal model for this process, in order to ascertain the factors influencing quaternary N-glucuronide formation. (S)-(-)-Nicotine-N(+)-1-beta-glucuronide was formed in vitro when [2'-14C]-nicotine was incubated with Triton X-100 activated marmoset hepatic microsomes in the presence of uridine diphosphoglucuronic acid; it was not formed when activated microsomal preparations of rabbit, guinea-pig, or rat were used as enzyme source. The glucuronide was characterised by comparison with authentic synthetic (S)-(-)-nicotine-N(+)-1-beta-glucuronide using HPLC. The rate of formation of the glucuronide was almost linear during up to four hours of incubation, but still only accounted for a maximum of 6.0% of the available substrate at the end of five hours incubation. The synthetic and biosynthetic (S)-(-)-nicotine-N(+)-1-beta-glucuronides were hydrolysed by beta glucuronidase and alkali, but were resistant to acid hydrolysis. The results support the concept that the marmoset may be a good animal species to mimic man in studies of nicotine metabolism during exposure to tobacco smoke. In vitro studies using (+/-)-trans-3'-hydroxycotinine or (S)-(-)-cotinine (as potential substrate) and [14C]-uridine diphospho-glucuronic acid (as cofactor) failed to produce any new radiolabelled glucuronide when the above microsomal preparations were used. PMID- 10716039 TI - Effect of inhibitors on the biotransformation of tamoxifen by female rat and mouse liver slices and homogenates. AB - The metabolism of tamoxifen was studied in female Sprague-Dawley rat and mouse liver slices and homogenates, and the three principal tamoxifen metabolites, 4 hydroxytamoxifen, N-desmethyl-tamoxifen and tamoxifen N-oxide, were identified by HPLC using authentic standards. It was not possible to identify any of the minor metabolites such as the epoxides using this technique. The N-oxide metabolite only appeared when NADPH was added to the system; this is because the production of tamoxifen N-oxide is primarily mediated by microsomal flavin monooxygenase (FMO) which is NADPH dependent. However, this metabolite did appear in incubations with mouse liver slices only, because they are rich in flavin monooxygenases (FMOs). It did not appear in female rat or mouse liver homogenates, because the NADPH present is destroyed during homogenisation, therefore it was necessary to add NADPH to the system to produce the N-oxide metabolite. The purpose of this study was to investigate the effect of inhibitors on the biotransformation of tamoxifen by female rat and mouse liver slices and homogenates. Female rat liver slices and homogenates were incubated with the following inhibitors (1 mM): cimetidine, ascorbate, sodium azide and reduced glutathione. Cimetidine, a general P-450 inhibitor, inhibited the production of the N-desmethyl metabolite by about 80%; this is in agreement with the action of the other inhibitors. Reduced glutathione, ascorbate and sodium azide are mainly peroxidase inhibitors, so therefore from these novel and interesting results it was possible to suggest that peroxidases play a role in the metabolism of tamoxifen. This observation was also strengthened when the production of the N desmethyl metabolite increased when horseradish peroxidase was added to the incubate. The production of 4-hydroxytamoxifen was reduced and the N-oxide metabolite was completely inhibited in the presence of peroxidase inhibitors. When rat liver homogenates was incubated with superoxide dismutase (SOD) and catalase, it was observed that the N-desmethyl metabolite disappeared completely at 60 min and the N-oxide and 4-hydroxy metabolites were completely inhibited. However, this phenomenon was only observed when SOD and catalase were preincubated for 30 min with the rat liver homogenate at 37 degrees C; without preincubation the production of these metabolites was unaffected. Finally, the effect of long incubation periods (300 min) on the production of metabolites was examined. It was found that there was a reduction in the concentration of metabolite produced after 60 min which was due to enzyme and co-factor degradation. PMID- 10716040 TI - Time dependent pharmacokinetic interaction between phenylpropanolamine and chlorpheniramine maleate in human subjects. AB - The influence of time of administration on the serum levels of phenylpropanolamine when administered alone and in combination with chlorpheniramine maleate at two different times of a day was studied in healthy human volunteers in a randomized 4 x 4 Latin square crossover design with a washout period of ten days. Blood samples were collected at predetermined time intervals and serum samples were analysed for unchanged phenylpropanolamine using high performance liquid chromatography. The various pharmacokinetic parameters of phenylpropanolamine were calculated using model independent methods. There was a significant (P < 0.05) decrease in the rate of absorption of phenylpropanolamine following its administration in combination with chlorpheniramine maleate at 2200 hours. However, such a change was not observed for treatment at 1000 hours. The observed change may be due to the time dependent gastrointestinal effect of these drugs. PMID- 10716041 TI - Pharmacokinetic interaction between diltiazem and tolbutamide. AB - The effect of co-administered tolbutamide and diltiazem on each drug's pharmacokinetics was studied in eight healthy male volunteers aged 21-25 years, with a 3 x 3 randomised crossover design. Each subject received orally 60 mg of diltiazem hydrochloride or 500 mg of tolbutamide, or both drugs. The washout period between each treatment was 7 days. Serum levels of diltiazem and tolbutamide were determined by HPLC. Serum profiles were analysed using a non compartmental model. There was no change in the pharmacokinetics of diltiazem in the presence of tolbutamide. There was approximately 10% increase in AUC0-24 and Cmax for tolbutamide in the presence of diltiazem. PMID- 10716042 TI - Metabolism of doxorubicin in long-term bone marrow cultures and SR-4987 stromal established cell line. AB - The metabolism of doxorubicin was studied in murine long-term bone marrow cultures (LTBMC) and in SR-4987 established stromal cells in comparison with primary cultures of murine and rat hepatocytes. The toxicity of metabolites was verified by testing their effects on the clonogenicity of granulo-macrophage progenitors. Metabolic activity was compared in subcellular fractions of SR-4987 cells and murine hepatocytes. Doxorubicin was transformed in long-term bone marrow cultures, SR-4987 cells and murine/rat hepatocytes to less toxic metabolites: 13-OH doxorubicin and a less polar metabolite which were non-toxic on granulo-macrophage progenitors. Among the hemopoietic compartments, stromal cells were responsible for the biotransformation of doxorubicin. The capability of the SR-4987 established stromal cell line to metabolize doxorubicin was higher than that of primary cultures of hepatocytes and bone marrow, and the highest activity was concentrated in the microsomes. These results suggest that in vitro models using primary cell cultures and established cell lines could be a useful tool for investigating the mechanisms underlying detoxification in the bone marrow stromal population. PMID- 10716043 TI - Fixation of patella fractures with braided polyester suture: a biomechanical study. AB - We tested the quality of fixation of displaced transverse patella fractures using braided polyester suture to investigate the suitability of this material as an alternative to stainless steel wire for fixation of these fractures. Osteotomies were created to simulate fractures of the patella in ten cadaveric knee specimens and were sequentially fixed using two techniques: the modified tension-band technique and the longitudinal anterior band (Lotke) technique. Each technique was implemented using either 1.25-mm stainless steel wire or 7-metric braided polyester suture (No. 5 Ethibond). The quality of fixation for each technique was tested by measuring the fracture gap during three simulated extensions of the knee against gravity on a materials testing machine. All techniques behaved comparably under the loading conditions used. In the four groups, there was no fixation failure (fracture gap > 3 mm) nor any significant difference between the mean maximum fracture gaps. The quality of fixation for braided polyester suture was comparable to that of stainless steel wire for such fractures, providing sufficient stability to withstand loads likely to be encountered during postoperative rehabilitation. Our results support the use of braided polyester suture as an alternative to stainless steel wire for fixation of displaced patella fractures. PMID- 10716044 TI - Blood transfusion requirements in femoral neck fracture. AB - The blood transfusion requirements of a consecutive series of 249 unselected patients with femoral neck fracture were studied retrospectively. A total of 339 Units of blood were transfused (a mean of 1.36 Units per patient). Blood transfusion occurred in 132 patients (53.0%), with each receiving a mean of 2.57 Units. Patients aged 80 years and above as a group were transfused significantly more blood than those aged less than 80 years: 1.64 vs 0.94 Units, X2 = 12.09, p < 0.001. Patients with intertrochanteric fractures were transfused significantly more blood than those with intracapsular fractures (1.74 vs 1.00 Units: X2 = 13.4, p < 0.001). PMID- 10716045 TI - Seasonal variation of hip fracture at three latitudes. AB - We studied the seasonal variation of hip fracture admissions at three different latitudes: Scotland (56 degrees North; 54,399 admissions); Shatin, Hong Kong (22 degrees North; 4180 admissions); and Auckland, New Zealand (36 degrees South; 2257 admissions). We calculated the extent of seasonal variation (amplitude) and the time of year of the peak value (acrophase) by fitting a sine curve to monthly data using cosinor analysis. A significant seasonal variation was found in all three countries, at a high level in Scotland (p < 0.01) and Hong Kong (p < 0.001), but just significant in New Zealand (p < 0.05). The extent of the seasonal change was very similar in Scotland and New Zealand, but, as expected, the peak in New Zealand (early September) was approximately six months ahead of Scotland (mid February). In Hong Kong, the amplitude was three times greater than in Scotland and the peak occurred a month earlier. There is neither snow nor ice in Hong Kong, and this provides powerful evidence against a major influence of conditions underfoot causing extra falls in winter. In Scotland there was a significant increase in the proportion of deaths in winter as compared to summer. The Scotland/Hong Kong amplitude difference is striking, but it is unknown whether this has a genetic or environmental explanation. The cause of seasonal death difference to a given injury is also unknown. Possible mechanisms are discussed, but the purpose is to report two new epidemiological features, without wild speculative hypotheses. The findings should be viewed as leads to further epidemiological, clinical and more basic research. PMID- 10716046 TI - A new technique for delayed primary closure of fasciotomy wounds. AB - Fasciotomy for compartment syndrome in the lower limb is a surgical emergency to preserve future limb function. The advised standard procedure involves both medial and lateral dermotomy in addition to the fasciotomy. There is often concern before and after performing fasciotomy about the cosmetic appearance and prolonged hospital stay if split skin grafting is required to cover the resultant skin defect. This is the case in over 50% of lower limb fasciotomies. We have used a technique of subcuticular prolene suture, first described for the delayed primary closure of contaminated abdominal wounds, in six patients who had undergone lower limb fasciotomies. In all of these cases delayed primary closure was easily achieved without the need for skin grafting. Experiments using a synthetic skin model have shown a 60% reduction in suture tension when compared with interrupted vertical mattress suturing. The subcutaneous prolene suture has the advantage of being both the method of approximation and final closure whilst spreading tension evenly across the wound edges without causing skin edge necrosis. It appears to be simpler and more economical than any technique so far described for the successful delayed primary closure of fasciotomy wounds. PMID- 10716047 TI - Femoral nailing for metastatic disease of the femur: a comparison of reamed and unreamed femoral nailing. AB - A total of 73 consecutive intramedullary femoral nails were inserted for metastatic disease of the femur; 43 were reamed and 30 were solid nails. The two groups were similar with regards to age, type of primary tumour, anatomical site, acute or 'impending' fracture and postoperative survival. The 'solid' nail offers a satisfactory alternative form of stabilisation for metastatic disease of the femur with rates of implant failure which are comparable with the reamed nail. In this series bilateral nailing was not associated with any increase in mortality. Contrary to other reports, imposing a delay in patients with pain and a short life expectancy seems unjustified. The use of the 'solid' femoral nail does not prevent sudden death due to massive fat embolism. PMID- 10716048 TI - Results of 1018 digital replantations in 552 patients. AB - To find out the influencing factors of the immediate and late outcome of replantation and revascularization of the digits the study was carried out as a prospective survey research during 1983 to 1995 with at least 2 year follow up. Traumatic total or subtotal amputation with inadequate circulation of the digits distal to the metacarpal head were included in the study. There were 552 patients with 366 males (675 digits) and 186 females (343 digits). Successful operation was found in 508 patients (92%) with 946 digits (92.9%). Type of injury was the most important factor influencing immediate and late outcome. Regular cigarette smoking resulted in poor immediate survival rate. Prolonged ischaemia had a significant influence in final functional outcome. Using composite skin and subcutaneous vein graft gave good survival rate. Injury at the no man's land area resulted in poor range of movement of the digit. Connecting the profundus tendon stump of the proximal part to the superficialis tendon of the amputated part gave a better result than 2 tendon repair and repairing only the profundus tendon. Replantation should be carried out even if only one digit is involved. At the 2 year follow up 195 patients (38%) were classified in grade I of Chen et al., functional outcome, while 153 (31%) were in grade II, 124 (24%) were in grade III and 36 (7%) were in grade IV. PMID- 10716049 TI - The price of peace: the personal and financial cost of paramilitary punishments in Northern Ireland. AB - The aim of this study was to compare the injuries sustained during paramilitary punishments both before and after the onset of "peace" in Northern Ireland. A retrospective chart review was performed looking at age, injuries, treatments instituted, theatre time, length of hospital stay and overall cost of care. In the 10-month period before the ceasefire, 31 patients were treated after sustaining paramilitary punishment shootings. Mean age was 25.2 years. All patients had small entrance and exit wounds with minimal soft tissue disruption. A total of 18 fractures were recorded in 15 patients and 14 arteries required repair. Mean operative time was 2.6 h, mean hospital stay 7.61 days and mean cost per patient 3102 Pounds. In the following 10-month period 28 patients were admitted after punishments, only one of whom had been shot, all others had been beaten with sticks and clubs. Mean age was 27.4 years. In 52 limbs, 64 fractures were recorded and 44% of these were open; 15 of the fractures were Gustilo and Anderson Grade III or greater and 11 of the fractures were intra-articular. There were no arterial injuries. Mean operative time was 2.6 h, mean hospital stay 12.4 h and mean cost per patient 3849 Pounds. PMID- 10716050 TI - Antegrade intramedullary fixation of displaced fifth metacarpal fractures. AB - We report a new and simple modification for antegrade insertion of intramedullary K-wires used in the treatment of displaced fifth metacarpal fractures. This method of fixation was performed on six patients all of whom had excellent results when reviewed clinically and radiologically with a mean follow-up of 9 months. PMID- 10716051 TI - Scintigraphic evaluation of tibial shaft fracture healing. AB - A scintigraphic study of the healing process of type A and B closed tibial shaft fractures was carried out in 40 cases treated non-operatively, comprising 32 men and eight women aged 30.6 yr on average. Scintigraphic scans were obtained with technetium methylenediphosphonate (MDP-Tc99m, 25 mCi) at 6, 12 and 24 weeks after the fracture and an activity index was calculated taking the mean of three consecutive uptake counts for both fractured and normal opposite leg, used for comparison. The results showed that the activity index in general decreased progressively from the first to the third evaluation, with little difference in behaviour between the two types of fractures. However, for B type fractures the activity index remained stable from the first to the second evaluation, followed by a marked decrease at the third evaluation, with a comparable end result for both fracture types. It was concluded that a decrease of the activity index occurs in both types of closed fractures undergoing uneventful healing and that such a decrease can be taken as a parameter for further studies which include delayed union and non-union. PMID- 10716052 TI - Chorioretinitis sclopetaria. PMID- 10716053 TI - Unsuspected agenesis of the odontoid peg in a trauma patient. PMID- 10716054 TI - Avascular necrosis of the talus after a minimally displaced neck of talus fracture in a 6 year old child. PMID- 10716055 TI - Recurrent acute compartment syndrome. PMID- 10716056 TI - Open pelvic fractures with vaginal laceration: an unusual clinical feature. PMID- 10716057 TI - Consequence of equal absorption, distribution and/or elimination rate constants. AB - When fitting experimental data to an open one- or two-compartment model, with first order kinetics, it may happen that no optimized value is obtained for model parameters. Several authors pointed out that this case is especially encountered when absorption and elimination coefficients approach each other in a one compartment model or when absorption and exponential elimination or distribution rate constants are equal in a two-compartment model. We analyze these situations of equal coefficients here. Firstly, dealing with a one-compartment model, we get the concentration in the central compartment after a single oral dose and after successive various doses at various times (first order kinetics). Secondly, dealing with a two-compartment model, also for single or successive various doses, the concentration is expressed when absorption and exponential elimination or distribution rate constants are equal. In all cases, the areas under concentration curves and the mean residence time of the drug are calculated even when cancellation of one exponential term occurs. Furthermore, the concentration at steady-state is taken into account. PMID- 10716058 TI - Prediction of distribution coefficients from structure. Comparison of calculated and experimental data for various drugs. AB - The efficiency of the program PrologD to predict distribution coefficients (D) at any pH and pairing ion concentration has been tested using experimental logD values for various drugs measured under standard conditions of buffers and ionic strength. Clonidine derivatives, fluoroquinolones and beta-blockers were included as particular pharmacological classes within the testing data set. Calculations were performed using the three logP estimation options implemented in the program. PrologD proved to be very efficient and can be of great advantage in drug research. Prediction patterns and correlations between experimental and calculated data indicate acceptable results for more than 80% of the data. In addition, comparable studies using the different options permitted suggestions for the more suitable logP estimation method in respect of the particular classes of compounds. PMID- 10716060 TI - Bioequivalence studies: biometrical concepts of alternative designs and pooled analysis. AB - A bioequivalence study compares the bioavailability between a test and a reference drug product in terms of the rate and extent of drug absorption. Area under the plasma concentration-time curve (AUC) and maximum plasma concentration (Cmax) are the pharmacokinetic parameters that serve as characteristics for the assessment of the extent and rate of absorption, respectively. The experimental design of a bioequivalence study is usually a crossover and rarely a parallel or a paired comparative. The statistical assessment of bioequivalence is based on the 90% confidence interval for the ratio of the test mean to the reference mean for AUC and Cmax The aims of this paper are to: (i) investigate alternative designs to a crossover design for conducting bioequivalence studies; (ii) propose the statistical analysis of different designs for bioequivalence studies on the same products; and (iii) discuss their usefulness for the approval of new generic drug products. For this purpose, three case studies are illustrated and analysed. The first case study concerns the investigation of the merits of a crossover design relative to a parallel group design for highly variable drugs using as an example a bioequivalence study of tamoxifen products. The second case study concerns the pooled statistical analysis of two bioequivalent studies of the same levodopa products. The analyses of the individual studies failed to meet the regulatory criteria for bioequivalence. The one study design was a paired comparative and the other one a crossover. Under some assumptions the crossover design may be considered as a paired comparative and the data from the two studies may be analysed together as a paired comparative design. The third case study concerns the statistical pooled analysis of two bioequivalent studies of the same clodronate products. The one study was a three-period crossover pilot study and it was used to identify the variability of the active substance. Then, this variability was used to determine the number of subjects for the main pivotal study which was a two-period crossover. The pilot study design was converted into a two-period crossover design and the data from the two studies were analysed together as a two-period crossover design. The original data of the studies were modified accordingly. PMID- 10716059 TI - Arteether toxicokinetics and pharmacokinetics in rats after 25 mg/kg/day single and multiple doses. AB - Multiple doses of arteether (ARTE) at 25 mg/kg cause CNS and anorectic toxicities in rats. The same dose of ARTE was used to study the toxicokinetics (TK) after multiple injections and the pharmacokinetics (PK) following single administration. Animals were administered ARTE in sesame oil for 7 days, blood samples were collected using destructive sampling for up to 192 h after dosing and assayed by HPLC-ECD. Two other groups of rats were administered either a single 25 mg/kg i.v. or i.m. dose. In addition, the drug remaining in the i.m. injection site was measured. During the 7 day treatments, anorectic toxicity of ARTE was observed, and that caused significant reductions in food consumption and body weight after day 2. TK data on days 2-7 revealed marked changes compared to the PK parameters estimated on day 1. AUC (4367 ng x h/ml) on day 7 was 5-fold higher than AUC (905 ng x h/ml) on day 1. The volume of distribution at steady state (V(SS)) on day 7 (41.8 l) was 40% of the day 1 value of the V(SS) (104.3 l). Clearance (CL) was increased by 89% of the day 1 value, from 0.98 l/h to 1.85 l/h on day 7. The elimination t(1/2) of ARTE was also prolonged from 13.7 h (day 1) to 31.2 h (day 7). These data suggest that ARTE may have altered its distribution and elimination in rats as a result of the systemic toxicity. Analysis of the injection sites showed that 38% and 91% of the total amount of ARTE single dose remained in the muscles at 24 h (after first injection) and 168 h (at 24 h after 7 daily multiple doses), respectively. Fast and slow absorption phases from muscle were seen with t(1/2) of 0.97 h and 26.3 h, respectively. The apparent elimination t(1/2) of ARTE after i.m. injection (13.7 h) was much longer than that after i.v. dosing (0.67 h) due to the prolonged muscle absorption phase. Acute toxicity data of artemisinin drugs demonstrated that animals receiving a high single ARTE dose in sesame oil died between days 5-11, similar to artemether. When animals received dihydroartemisinin formulated in 50% DMAC/oil, or artesunic acid and artelinic acid in 0.9% saline vehicle, they died between days 1 and 2. This suggests that delayed onset toxicity and death in the ARTE rats may also be due to slow absorption and prolonged drug exposure. Therefore multiple i.m. administrations cause anorexia and drug accumulation, possibly affecting the toxicokinetics and efficacy of the drug. PMID- 10716061 TI - Urinary excretion of phenobarbital and its metabolite p-hydroxyphenobarbital in convulsing and non-convulsing patients. AB - As part of an investigation of phenobarbital (PB) pharmacokinetics in patients with status epilepticus (SE), urinary excretion of PB and its main metabolite, hydroxyphenobarbital (HPB), was studied in patients who had an episode of SE, as well as in non-convulsing ones. Eleven in-patients were studied:(group 1) five patients (4 M + 1 F; 48 +/- 28 years old; 64 +/- 6 kg body weight; mean +/- SD) with convulsive status epilepticus, and (group 2) six patients (5 M + 1 F; 37 +/- 13 years old; 71 +/- 15 kg body weight) with epilepsy, seizure-free at the moment of PB administration and without established anti-epileptic therapy. All subjects received a single intravenous dose of PB (15 mg/kg) at a rate of 100 mg/min. PB and HPB concentrations were measured by high performance liquid chromatography with UV detection at 220 nm in urine samples collected throughout 24 h. The comparison of pharmacokinetic parameters of urinary excretion of PB and HPB showed a statistically significant difference in the values of recovery of HPB and total barbiturate (higher values in the patients with SE) in 24 h urine. Differences in the excretion of PB between the two groups of patients--higher values in the patients who had had an episode of SE, and in urine flow--slightly elevated volumes in the same group, failed to reach statistical significance, probably due to the small number of participants in the study. PMID- 10716062 TI - Overview of stobadine bioanalysis: evaluation and application in pharmacokinetics. AB - Besides its many pharmacodynamic actions, the pyridoindole stobadine was found to exert antioxidant activity and thus possesses the potential to protect various tissues against oxidative stress. This overview is focussed on both the evaluation of the chemical procedures used in the bioassay of stobadine and its metabolites and on the comparison of their quality in the light of applicability for preclinical and clinical pharmacokinetic experiments. All methods and applications were performed at the Institute of Experimental Pharmacology, SASc in Bratislava, Slovakia. In pharmacokinetic and toxicokinetic studies, [3H] labeled stobadine dihydrochloride was administered intravenously or orally to rats in single and repeated doses. Liquid-liquid extraction was used for selective isolation of stobadine and its metabolites from biological matrix, followed by liquid scintillation quantification. A TLRC method was developed both to check the radiochemical purity of [3H]-stobadine and to quantify the labeled drug in rat plasma. A spectrofluorometric approach was used for determination of stobadine in dog serum and urine after its administration in the form of either the dihydrochloride or the dipalmitate. The method allowed us to perform a bioavailability study and a long-term toxicological study. The HPLC method with a limit of detection of 10 ng/ml of plasma proved suitable for calculating the compartmental pharmacokinetic parameters of both salt forms of stobadine administered to dog and man. This method was based on solid-phase extraction procedure by using Separcol SI C18 cartridges. In a GC method, the combination of capillary column separation and nitrogen-specific detection permitted the assay of serum stobadine concentrations as low as 5 ng/ml. The detection limit of the GC/MS method was 1 ng/ml of plasma or of phosphate buffer saline. This method was used for a bioequivalence study of two stobadine dipalmitate dosage forms and for a transdermal penetration study of stobadine acyl derivatives. All the developed assays proved to be appropriate for low-concentration determination of stobadine in a wide range of pharmacokinetic studies. PMID- 10716064 TI - Characterization of the highly variable bioavailability of tiludronate in normal volunteers using population pharmacokinetic methodologies. AB - Currently, the use of classical bioequivalence criteria is being called into question for certain classes of drugs such as bisphosphonates. These compounds typically possess a wide therapeutic index but may be characterized by low and variable absorption. The purpose of this communication was to characterize the highly variable bioavailability of tiludronate using a population pharmacokinetic method (NONMEM program) and compare the results to a standard 2 way cross-over bioequivalence trial in healthy subjects. Over 3500 plasma samples from 153 healthy subjects, representing 12 different clinical trials were pooled for mixed effect modeling purposes (complete data set). These studies, conducted under single and multiple dose conditions, contained all the directly comparable data available in healthy subjects administered a 400 mg dose of tiludronate. A two compartment model with first order absorption was fit to the plasma concentration time data and a term for relative bioavailability (BA) was included. Intersubject and residual variability were modeled using a constant coefficient of variation (CCV) model. A pilot model development data set was obtained from a 24 subject cross-over bioequivalence study. Population estimates of BA and its associated 90% confidence interval of 1.12 and 0.89-1.35 compared favorably to standard bioequivalence methodology (1.15 and 0.93-1.42, respectively). Since a good fit of predicted and observed plasma concentrations as well as estimates of BA were obtained, a two compartment model with a term for BA was then applied to the complete data set. Under these conditions, BA and its 90% confidence interval were found to be 1.17 and 0.98-1.36. Intersubject variability of 31%, compared with 38% in the pilot model development data set and residual variability of 38% were seen. No differences in absorption characteristics as measured by Ka were found. Good agreement between the population pharmacokinetic parameters were observed when the pilot data set was compared with the full data set. The proposed model was confirmed by creating 10 additional smaller data sets that were matched for the number of subjects given both formulations under single and multiple dose conditions. No change in the estimate of BA was observed under these study conditions. This study demonstrated that population pharmacokinetic methodology can be applied successfully to problematical bioequivalence issues that may occur during the development process. Increasing the number of subjects in the overall analysis did not alter the estimate of BA or its 90% confidence interval, when compared to the original cross-over bioequivalence study. Bayesian approaches can be of value in large clinical trials where typically relatively few plasma samples are obtained from individual subjects. PMID- 10716063 TI - Inhibiting effect of ethinylestradiol/levonorgestrel combination on microsomal enzymatic activities in rat liver and kidney. AB - The aim of the study was to evaluate the effects of two therapeutic combinations of ethinylestradiol (EE) and levonorgestrel (LE), which are used in triphasic contraceptives, on the activities of drug-metabolizing enzymes in rat liver and kidney. Sexually mature female Wistar rats were given 0.03 mg EE and 0.05 mg LE, or 0.03 mg EE and 0.125 mg LE for 6 or 18 sexual cycles, i.e. for 30 or 90 days. EE/LE inhibited not only the metabolic capacity of P450, a protein which directly undergoes suicide inhibition, but also the level of rat liver cytochrome b5 (dependent on the heme pool) as well as the activities of NADPH-cytochrome P450 reductase and NADH-cytochrome b5 reductase in the liver and kidney. The majority of these effects were independent of the gestagen dose and of the duration of treatment, suggesting that estrogen is a predominant inhibiting factor in the EE/LE combination. The study has revealed differences in the enzyme activities between the liver and kidney, which may result from the fact that these organs display different sets of P450 isoforms and, therefore, their monooxygenase systems show distinct capacities to metabolize exogenous steroids. PMID- 10716065 TI - Blood and cerebrospinal fluid pharmacokinetics of primidone and its primary pharmacologically active metabolites, phenobarbital and phenylethylmalonamide in the rat. AB - Primidone is a clinically useful antiepileptic drug that is metabolised to two pharmacologically active metabolites phenobarbital and phenylethylmalonamide. As data on the inter-relationship between the systemic and central nervous system pharmacokinetics of primidone and its metabolites are sparse, we have investigated their temporal inter-relationship using a freely behaving rat model which allows repeated sampling of blood (100 microl) and cerebrospinal fluid (CSF; 20 microl). After administration, by intraperitoneal injection (50, 100 or 200 mg/kg), primidone rapidly appeared in both serum (Tmax mean range 1.5-2.5 h) and CSF (Tmax mean range 2.0-3.5 h), suggesting ready penetration of the blood brain-barrier. This was also the case for phenylethylmalonamide and phenobarbital but peak concentration occurred later. Primidone, phenylethylmalonamide and phenobarbital concentrations rose linearly and dose-dependently in both serum and CSF. The mean free fraction (free/total concentration ratio) for primidone, phenylethylmalonamide and phenobarbital was 0.86, 0.97 and 0.88, respectively, and, as their respective mean CSF/serum ratio values were 0.73, 1.06 and 0.65, it would suggest that equilibration between the blood and CSF compartments is rapid. CSF mean t(1/2) values for primidone, phenylethylmalonamide and phenobarbital were similar to those of sera and essentially paralleled the pattern seen in sera. PMID- 10716066 TI - Pulsed estrogen therapy: pharmacokinetics of intranasal 17-beta-estradiol (S21400) in postmenopausal women and comparison with oral and transdermal formulations. AB - Pharmacokinetics of estradiol and estrone were assessed in postmenopausal women receiving S21400, a novel 17beta-estradiol formulation administered by nasal route; the results were compared with those obtained with oral and transdermal routes. Thirty six women received three treatments: a specified dose of 17beta estradiol (100, 300 or 450 microg) given once and as 2 doses, 12 h apart, using three parallel dose groups in a randomised, crossover study. Thereafter, a reservoir patch (50 microg/day of 17beta-estradiol) or a tablet of 2 mg micronised 17beta-estradiol were randomly administered. Plasma concentrations of estradiol and estrone were measured by radioimmunoassays. Following intranasal dosing, estradiol was rapidly absorbed with plasma concentrations reaching maximal values (approximately 1400 pg/ml with a single 300 microg dose) after 10 30 min and returning within 12 h to levels of untreated postmenopausal women. Systemic exposure to estradiol was dose proportional and independent of the treatment regimen. Moreover, the dose of 300 microg gave an estimated 24 h systemic exposure to exogenous estradiol close to that of the 50 microg/day reservoir patch or the 2 mg tablet. The mean estrone to estradiol ratio was similar and 4-fold lower than those with the patch and the tablet, respectively. In conclusion, by this new route for estrogen replacement therapy, the nasal route, the pharmacokinetics of estradiol as S21400 were linear and displayed a 'pulsed' kinetic profile, different from those obtained with the usual routes of administration. PMID- 10716067 TI - A further interaction study of quinine with clinically important drugs by human liver microsomes: determinations of inhibition constant (Ki) and type of inhibition. AB - Our previous study showed that several drugs inhibited quinine 3-hydroxylation, a cytochrome P450 (CYP) 3A4-mediated reaction, in vitro. In this extended study, 13 drugs were selected and tested by human liver microsomes in order to further determine their respective inhibition constant (Ki) and type of inhibition. According to the apparent Ki values, the inhibitory rank order of these tested drugs was as follows: ketoconazole > doxycycline > omeprazole > tetracycline > troleandomycin (with pre-incubation) > primaquine > troleandomycin (without pre incubation) > nifedipine > erythromycin > verapamil > oleandomycin > diltiazem > cimetidine > hydralazine. Among these drugs, doxycycline, tetracycline, ketoconazole, nifedipine and hydralazine were judged as mixed inhibitors; whereas, the remaining other drugs tested were judged as competitive inhibitors. When the plasma/serum concentrations possibly attained after their usual therapeutic doses were taken into account, tetracycline, doxycycline, omeprazole, ketoconazole, nifedipine, troleandomycin and erythromycin are likely to be inhibitors of quinine metabolism in patients when these drugs are co administrated with quinine. PMID- 10716068 TI - Biotransformation of BOF-4272, a sulfoxide-containing drug, in the cynomolgus monkey. AB - BOF-4272, (+/-)-8-(3-methoxy-4-phenylsulfinylphenyl) pyrazolo[1,5-a]-1,3,5 triazine-4(1H)-one), is a new drug intended for the treatment of hyperuricemia. This report describes the pharmacokinetics and detailed metabolic pathways of BOF 4272 in the cynomolgus monkey, which were investigated using the metabolites found in plasma, urine, and faeces after intravenous and oral administration. M-4 was the main metabolite in plasma after intravenous administration. M-3 and M-4 were the main metabolites in plasma after oral administration. The Cmax and AUC(0 t) of M-4 were the highest of all the metabolites after intravenous administration. The Cmax and AUC(0-t) of M-3 were the highest of all the metabolites, and those of M-4 were the second highest, after oral administration. M-4 and M-3 were the main metabolites detected in urine and faeces, respectively, after intravenous administration, with M-4 and M-3 at 47.2% in urine and 19.1% in faeces, respectively, within 120 h after administration. M-4 was the only metabolite detected in urine after oral administration, at about 5% within 120 h after administration. M-3 was detected in faeces at 17.0% within 120 h after oral administration. These results suggest that, in the cynomolgus monkey, BOF-4272 is rapidly biotransformed to a main metabolite (M-4, a sulphoxide-containing metabolite of BOF-4272) and that M-4 is mainly excreted in urine and possibly also in bile, with subsequent conversion to M-3 by the intestinal flora. It is expected that the biotransformation of BOF-4272 would be similar in healthy human volunteers. PMID- 10716069 TI - Hepatitis B, C and human immunodeficiency virus infections in multiply-injected kala-azar patients in Delhi. AB - Sera from 164 patients with parasitologically confirmed kala-azar and 100 patients with non-kala-azar Delhite in 2 Delhi hospitals were tested for anti human immunodeficiency (anti-HIV) and anti-hepatitis C virus (anti-HCV) antibodies and hepatitis B surface antigens to determine which group is more likely to contract these infections. The mean age of the patients was 32.5 y (+/ 6.5 y), (120 M, 44 F). Two patients were from Nepal and the others from the kala azar endemic state of Bihar, India. As geographical controls, 50 serum samples from sex- and age-matched healthy Bihar residents were also tested for the blood borne viral infections. All patients had been treated with injectable medicines by 1 or more local physicians before they were referred to the Delhi hospitals. The prevalence of hepatitis B virus (HBV) and HCV infection was significantly different between the 2 patient groups. While 2 kala-azar patients (1.21%) were found to be HIV-1 positive, 54 (32.9%) patients had anti-HCV antibodies detected by ELISA and 51 (31.1%) by RIBA test. The seroprevalence of HCV was only 2% in hospitalized non-kala-azar cases and 4% in the geographical controls (p < 0.001). The seroprevalence of HBV was 13.2% in hospitalized kala-azar cases, but only 1.75% in disease control cases and 1.6% in geographical control cases. The difference in infection rates between cases and controls was significant (p < 0.001). The results indicate that kala-azar patients treated locally in Bihar have a greater chance of contracting blood-borne infections. Interestingly, we found that HCV was more prevalent than HBV. These infections were most likely acquired through the re-use of needles by local medical and paramedical practitioners for administering anti-leishmanial drugs. This trend, if not checked immediately, may have drastic consequences in the horizontal transmission of HIV in Bihar. PMID- 10716070 TI - Different expression of ICAM-1 and LFA-1 alpha by peripheral leukocytes during respiratory syncytial virus and influenza virus infection in young children. AB - We compared the expression of intercellular adhesion molecule (ICAM)-1 and lymphocyte function-associated antigen (LFA)-1 alpha (alpha) on the surfaces of peripheral immunocompetent cells of young children infected with respiratory syncytial virus (RSV) and influenza virus. Flow cytometric analysis revealed a significantly higher percentage of CD14 + monocytes/macrophages that were strongly ICAM-1-positive in the influenza group than in age-matched controls, whereas the ICAM-1 expression levels of the RSV and control groups did not differ significantly. Analysis of LFA-1alpha expression by CD3 + T lymphocytes showed a significantly higher percentage of strongly positive cells in the influenza group than in the age-matched controls. By contrast, the percentage of cells that were strongly LFA-1alpha-positive was significantly lower in the RSV group than in the age-matched controls. These results suggest that no increase of adhesion molecule expression occurs in vivo in patients with RSV infection, and also that there is little activation of peripheral blood monocytes/macrophages and T lymphocytes in young children with RSV infection. PMID- 10716071 TI - Chlamydia pneumoniae and Mycoplasma pneumoniae in children with acute respiratory infection in general practices in The Netherlands. AB - In this retrospective study Chlamydia pneumoniae and Mycoplasma pneumoniae infections were detected by polymerase chain reaction (PCR) in samples (n = 457) from children presenting with acute respiratory infection to general practitioners during 1992-97. Samples were collected in autumn and winter, and from 1994 onwards in spring and summer also. Overall, C. pneumoniae and M. pneumoniae were detected in throat or nasal samples by PCR in 3.1% and 2.4% of the cases, respectively. The proportion of both C. pneumoniae and M. pneumoniae infections varied between 0% and 6.9% over the years studied, whereas seasonal proportions varied from 1.8 to 9.1% and 1.2 to 4.5%, respectively. For both microorganisms the lowest proportion was detected during winter and the highest in summer. C. pneumoniae could already be detected by PCR in patients under 4 y of age, an observation not made in sero-epidemiological studies. In conclusion, both C. pneumoniae and M. pneumoniae infections play a minor role in children presenting with acute respiratory infection. PMID- 10716072 TI - Sero-prevalence of granulocytic Ehrlichia spp. and Borrelia burgdorferi sensu lato in Swedish dogs 1991-94. AB - In Sweden, 2 tick-borne zoonotic diseases, granulocytic ehrlichiosis and borreliosis, are frequently diagnosed in dogs, using serological assays. The aims of this study were to determine the sero-prevalences of antibodies to Ehrlichia spp. and Borrelia burgdorferi sensu lato during 1991-94 in dogs, not clinically suspected to be infected with either of the 2 agents. Samples (n = 611) were selected from a serum bank using a systematic sampling strategy, stratified across the 4-y period. The stored sera had originally been submitted in order to verify or rule out infection with Sarcoptes scabiei. The overall sero-prevalence for Ehrlichia spp. was 17.7% and for Borrelia burgdorferi sensu lato 3.9% (n = 588). Only a few dogs in the northern part of Sweden were sero-positive for Ehrlichia spp. and none were positive for Borrelia burgdorferi sensu lato. An increased sero-prevalence of Ehrlichia spp. was seen during the years studied. The sero-prevalence of Ehrlichia spp. varied with season. Sero-positivity to both agents increased with age. Both diseases are considered zoonotic, and the increase in sero-prevalence of Ehrlichia spp. over the years may reflect the degree of infection in ticks and may have implications for human health. PMID- 10716073 TI - Clostridial bacteremia in the community hospital. AB - Anaerobic infections are not commonly studied in the community hospital. The aim of this study was to determine demographic factors, the portals of entry and underlying disorders for clostridial bacteremia and to determine whether appropriate (recommended) treatment is effective. Medical records were reviewed for 42 patients with clostridial bacteremia at 1 Florida, USA, hospital and 4 Dayton, Ohio, USA, hospitals. Fourteen (33.3%) of the patients had clostridial micro-organisms that were isolated in cultures with polymicrobial isolates. Only about half of the patients had fever at the onset of their bacteremia and only slightly more than half had elevated leukocyte counts. The most common portals of entry for the micro-organisms were gastrointestinal (42.9%), unknown (35.7%) and skin (16.7%). The most common underlying disorders were advanced malignancy (31.0%), diabetes mellitus (14.3%), none determined (12.0%) and acute cholecystitis (9.5%). The mortality rate was 23.8%. Timely appropriate treatment was started in only about half of the instances. Appropriate (recommended) treatment did not significantly affect survival (p = 0.469). Clostridial infections and bacteremia exist in the community hospital most commonly in severely ill patients. The fact that clostridia are commonly cultured in blood cultures positive for other bacterial pathogens and that appropriate treatment for clostridia did not affect patient survival, call into question the significance and pathogenicity of clostridial organisms. On the other hand, if clostridial bacteremia was not considered in half these patients with bacteremia, it is possible that more indolent clostridial infections are being overlooked. PMID- 10716074 TI - Exposure of hospital personnel to Brucella melitensis and occurrence of laboratory-acquired disease in an endemic area. AB - In 1997, 7 cases of laboratory-acquired Brucella melitensis infections were detected among the hospital personnel of a medical centre serving an endemic area in southern Israel. Although the onset of symptoms in 6 of the 7 patients occurred during a 2-week period, suggesting a point source exposure, biotype analysis showed that the outbreak was caused by 3 different B. melitensis serovars, indicating multiple exposures. Review of the laboratory records showed that during 1997, the microorganism was recovered from 146 blood and synovial fluid cultures, and that during the 2 months in which the laboratory-acquired cases occurred (April and June), 53 of 530 positive aerobic blood culture bottles (10.0%) grew B. melitensis. The epidemiological investigation did not reveal the source of the outbreak, and no noticeable breaches in laboratory safety practices could be demonstrated. It is concluded that in areas endemic for brucellosis, hospital personnel are frequently exposed to Brucella microorganisms. Under these circumstances, significant morbidity may occur despite observance of recommended safety practices. Biotyping of Brucella isolates may contribute to the elucidation of complex epidemiological situations. PMID- 10716075 TI - Tuberculous subcutaneous abscesses developing during miliary tuberculosis therapy. AB - Although rare, paradoxical subcutaneous abscesses may develop during appropriate treatment of miliary tuberculosis. While the pathogenesis of this phenomenon is not clear, some theories have been postulated. A case of a 37-y-old woman diagnosed as having miliary tuberculosis who developed subcutaneous abscesses within the 5 months of antituberculous treatment is described and all 6 similar cases published in English from 1954 to 1999 are discussed. PMID- 10716076 TI - Tuberculous meningitis in children: clinical features and outcome in 40 cases. AB - In order to assess the epidemiology, clinical features and outcome of tuberculous meningitis, a retrospective review of patients was conducted between January 1989 and December 1995. Forty cases (representing 10%, of all paediatric patients with tuberculosis) were included. Mean age was 46 months (range 1-165 months). Eighteen (45%) children were classified as stage I (non-specific febrile illness without neurological signs), 16 (40%) as stage II (neurological signs without marked changes in sensorium) and 6 (15%) as stage III (major neurological signs with sensorial changes and/or coma). Twenty-seven (67%) patients had received BCG vaccination and 14 (35%) displayed an induration zone higher than 10 mm after a 2 TU PPD test. Mycobacterium tuberculosis was recovered from 24 (61%) patients. Hydrocephalus was demonstrated by cranial computed tomography in 31 (78%) children. Overall, 18 (45%) children had a full recovery. Mild, moderate and severe neurological sequelae were shown by 7 (18%), 3 (8%) and 9 (22%) of the patients, respectively. Three fatal cases (7%) were observed. The presence of seizures (RR 15.6, 2.02-119.1) and absence of extrameningeal foci (RR 4.95, 1.10 22.1) were identified as risk factors by multivariate analysis. These findings emphasize the need quickly to diagnose tuberculosis in children in order to give appropriate and early treatment. PMID- 10716077 TI - Identification of non-tuberculous mycobacteria: 16S rRNA gene sequence analysis vs. conventional methods. AB - In a retrospective study, 45 clinical isolates of non-tuberculous mycobacteria were identified to the species level by biochemical profile, gas liquid chromatography and partial sequence analysis of 16S rRNA, and were found to represent 13 different species. The results of sequence analysis showed 100% identity with conventional tests for 34 isolates (76%) and could identify species such as M. bohemicum which are difficult to characterise with conventional methods. Most of the discrepant results for the remaining 11 isolates resulted in species of the same group of mycobacteria. Based on these findings. we concluded that direct sequence analysis of amplified 16S rRNA gene is a promising rapid and accurate method for species determination of non-tuberculous mycobacteria. PMID- 10716078 TI - Genetic analysis and clinical evaluation of vacuolating cytotoxin gene A and cytotoxin-associated gene A in Taiwanese Helicobacter pylori isolates from peptic ulcer patients. AB - The aims of this study were to investigate the cytotoxin-associated gene A (cagA) and vacuolating cytotoxin gene A (vacA) subtype in Taiwanese H. pylori isolates from patients with gastroduodenal diseases and to assess the relationship between genotypes of isolates and clinical features. The vacA s1a allele was found in all isolates and vacA m1 allele was found in 15% of isolates. The cagA gene was found in 82.5% of isolates. The vacA s1a/m2 strains had a significantly higher prevalence rate than vacA s1a/m1 strains in Taiwan (p < 0.05). By aligning and comparing the nucleotide and amino acid sequences of vacA from the Taiwanese isolates, the signal sequence and N-terminal region were found to be highly conserved, but the middle region was found to be highly heterogeneous. Determining the relationship between the genotypes and clinical features, we found that the cagA gene was more closely associated with duodenal ulcer than with gastric ulcer and the vacA s1a/m2 strain was more closely associated with active chronic gastritis and atrophic gastritis than with chronic gastritis. Together, our results indicated that (i) the middle region of vacA gene in Taiwanese isolates was heterogeneous; (ii) s1a/m2 vacA strains had a high prevalence in Taiwanese peptic ulcers; and (iii) the cagA gene was significantly associated with duodenal ulcer. PMID- 10716079 TI - Metronidazole and clarithromycin susceptibility and the subtypes of vacA of Helicobacter pylori isolates in Estonia. AB - The prevalence of metronidazole and clarithromycin resistance of Helicobacter pylori strains under different growth conditions (microaerophilic or anaerobic preincubation) was tested in 56 patients suffering from gastritis and peptic ulcer. vacA subtypes were detected in 46 H. pylori strains and were subsequently compared with the antibiotic resistance pattern. From 56 isolates, 26 proved resistant and 30 sensitive to metronidazole. The patients with peptic ulcer and gastritis were infected with both metronidazole-sensitive and metronidazole resistant strains. In anaerobic preincubation all the strains were sensitive to metronidazole (MIC < 8 mg/l). All the strains were clarithromycin-sensitive (MIC < 2 mg/l). In the patients with gastritis and peptic ulcer s1 was the predominant vacA subtype. Comparison of vacA subtypes with the diagnoses revealed no correlation; different virulence factors such as vacA subtypes and antibiotic resistance to metronidazole in a microaerophilic milieu proved unrelated. PMID- 10716080 TI - Effects of anti-neoplastic agents on the recovery of bacteria and yeasts in an automated blood culture system. AB - The effects of 7 anti-neoplastic agents on the recovery of 5 aerobic gram positive cocci, gram-negative rods and yeasts were studied in 2 different automated blood culture systems using an experimental model. In the absence of anti-neoplastic agents, the growth of gram-positive cocci was detected significantly earlier in standard than in FAN aerobic bottles. In the presence of 100 microM doxorubicin, however, the growth of gram-positive cocci was totally inhibited in standard culture conditions, while in FAN bottles the agent has no inhibitory effect. Etoposide at a concentration of 100 micromol/l also significantly delayed the growth of cocci in standard conditions. Neither the culture bottles nor the anti-neoplastic agents tested had any effect on the growth of gram-negative rods and yeasts. The results suggest that the anti neoplastic agents present in blood might disturb the growth of gram-positive cocci in blood culture. This should be considered when validating blood culture systems and evaluating blood cultures of chemotherapy-receiving febrile patients. PMID- 10716081 TI - Bacteremia among kidney transplant recipients: a case-control study of risk factors and short-term outcomes. AB - Kidney transplant recipients are highly susceptible to life-threatening infections, including bacteremia. To determine the risk factors for bacteremia within the first month after renal transplantation we performed a non-concurrent transplant population-based case-control study involving all 1,000 consecutively operated adult patients at Helsinki University Central Hospital in 1987-93. All patients with at least 1 positive blood culture within 31 d of transplantation were defined as cases. Control patients were drawn systematically from the transplant population with no positive blood cultures within the first 31 d post transplant. The study included 35 cases and 123 controls. The overall rate of bacteremia in the population was 3.5%. The case patients were more likely to have been on haemodialysis prior to transplantation (71%, vs. 43%, p < 0.05) and to have experienced acute rejection (46% vs. 20%, p < 0.05) than the controls. Local infections (46% vs. 12%, p < 0.05) were also more common among case patients. In the crude analysis an additive interaction of acute rejection and haemodialysis was found, with a 10% rate of bacteremia occurring if both conditions were present. The mortality rate within 2 months of follow-up was higher among case patients than among controls (14%, vs. 1%, p < 0.05) and they also returned more often to dialysis (23% vs. 4%, p < 0.05). Bacteremia during the immediate postoperative period might still have severe outcomes measured as allograft and patient survival at 2 months post-transplant. Further evaluation will confirm whether a lower rate of bacteremia among kidney transplantation patients can be achieved if peritoneal dialysis is preferred to haemodialysis whenever possible. PMID- 10716082 TI - TNF-alpha and IL-8 in consecutive sputum samples from cystic fibrosis patients during antibiotic treatment. AB - Proinflammatory cytokines in sputum are useful markers of the activity of lung disease in cystic fibrosis (CF). Tumour necrosis factor alpha (TNF-alpha) and interleukin-8 (IL-8) concentrations in sputum of 10 CF patients were determined during exacerbation and IL-8 in sputum of 48 patients at a yearly follow-up when patients were in optimal clinical condition. In 9 patients of the former group, TNF-alpha levels were increased during exacerbation. In 4 patients, the peak occurred within 2 d (median value > 1500 ng/l), whereas the remaining 5 had peak values on days 3-6 (median value 720 ng/l). IL-8 levels were > 800 microg/l in all 10 patients, and in 9 cases there was a positive correlation between IL-8 and TNF-alpha. Baseline IL-8 levels of 48 patients showed considerable variation (median 207 microg/l, range 1.5-392). There was a significant correlation between IL-8 concentrations and current colonization with either Pseudomonas aeruginosa or Staphylococcus aureus in the lower airways (p = 0.002), immunoglobulin G levels (p = 0.02) and the severity of the pathological findings shown by chest X ray (p = 0.008). High IL-8 and TNF-alpha values correlated with symptoms of deterioration. IL-8 levels seemed to be markers of both current bacterial colonization and the degree of lung damage. PMID- 10716084 TI - Successful non-surgical treatment of Candida tropicalis endocarditis with liposomal amphotericin-B (AmBisome). AB - Fungal endocarditis in children is most commonly a complication of palliative or curative surgery for congenital heart disease, rheumatic valvulitis and prolonged indwelling central venous and umbilical catheters. We describe here the case of a 3-y-old patient with chronic diarrhoea and prolonged total parenteral alimentation who developed severe C. tropicalis endocarditis and was treated successfully using a liposomal preparation of amphotericin-B (AmBisome) without surgical intervention. PMID- 10716083 TI - Comparative effects of moxifloxacin and clarithromycin on the normal intestinal microflora. AB - Twelve healthy male subjects age range 24-40 y participated in the investigation. The trial was divided into 2 35-d periods. The 2 treatment regimens were: (i) 1 x 400 mg moxifloxacin tablet in the morning and 1 placebo tablet in the evening for 7 d; and (ii) 1 x 500 mg clarithromycin tablet in the morning and 1 x 500 mg clarithromycin tablet in the evening for 7 d. Each subject received firstly I treatment regimen and secondly the other treatment regimen. The wash-out period was 6 weeks between the two treatment regimens. Moxifloxacin caused significant decreases of enterococci and enterobacteria during the administration period while the numbers of staphylococci, streptococci, Bacillus and Candida were not affected. No impact on peptostreptococci, lactobacilli, Veillonella, Bacteroides or fusobacteria was observed, while bifidobacteria and clostridia decreased during moxifloxacin administration. The microflora was normalized after 35 d. Clarithromycin caused significant reduction of Escherichia coli while the numbers of enterococci, Enterobacter, Citrobacter, Klebsiella and Pseudomonas increased markedly. No significant changes in the numbers of staphylococci, streptococci, Bacillus and Candida were noticed. In the anaerobic microflora bifidobacteria, lactobacilli and clostridia were suppressed, while no changes in peptostreptococci, Veillonella, Bacteroides and fusobacteria were found. The microflora was normalized in all volunteers after 35 d. PMID- 10716085 TI - Hospital-acquired brevundimonas vesicularis septicaemia following open-heart surgery: case report and literature review. AB - Brevundimonas vesicularis (B. vesicularis) is a pseudomonad rarely encountered in human infection. A case of nosocomial septicaemia with this organism following open-heart surgery is presented, with a review of the literature. The isolate demonstrated resistance to ciprofloxacin and aztreonam, which has not yet been reported. Treatment with piperacillin/tazobactam resulted in full recovery. A review of the literature reveals that B. vesicularis is a virulent organism involved in serious infections such as central nervous system infection or bacteraemia, some of which are nosocomial. Meanwhile, empiric therapy for B. vesicularis infection should include a broad-spectrum antimicrobial agent until susceptibility results are known. PMID- 10716086 TI - Melioidosis presenting as urinary tract infection in a previously healthy tourist. AB - Melioidosis is a tropical disease caused by Burkholderia pseudomallei, which is common in southeast Asia and Australia, but which is rarely diagnosed in Scandinavia. An increasing number of cases are being reported among tourists to infected areas. We report the first Finnish case of melioidosis, which presented as urinary tract infection in a previously healthy male tourist. PMID- 10716088 TI - Paradoxical response to anti-tuberculous drugs: resolution with corticosteroid therapy. AB - During the course of appropriate treatment, patients with tuberculosis occasionally have unusual paradoxical reactions, with transient worsening of lesions or the development of new lesions. A 23-y-old housewife presented with abdominal tuberculosis. She was treated with anti-tuberculous agents to which the micro-organisms were susceptible. During therapy, there was an expansion of her abdominal lesions and her symptoms worsened. However, with the addition of steroids and the continuation of the same anti-tuberculous agents the patient eventually recovered completely. We emphasize that the worsening of tuberculous lesions may occur during chemotherapy and does not necessarily indicate treatment failure. This phenomenon may be immunologically based. PMID- 10716087 TI - Mycobacterium fortuitum osteomyelitis in a peripheral blood stem cell transplant recipient. AB - Mycobacterium fortuitum is an uncommon, but well-recognized, pathogen in immunocompromised hosts, with special predilection for bone and soft tissue infections in solid organ transplant recipients. We describe a case of osteomyelitis due to this pathogen in a peripheral blood stem cell transplant recipient. PMID- 10716090 TI - Pericarditis due to Tsutsugamushi disease. AB - Tsutsugamushi Disease is an acute febrile illness caused by Rickettsia tsutsugamushi, which enters into the human bloodstream through the bite of leptotrombidium. It is characterized by eschar, fever and cutaneous rash. Pericardial effusion in Tsutsugamushi Disease is not a common manifestation, although a high rate of effusion was reported in autopsy in those who had died of the disease. Here, we report a case of Tsutsugamushi pericarditis documented by indirect immunofluorescent test of pericardial fluid, and give a brief review of the literature. PMID- 10716089 TI - Bacillus cereus fatal bacteremia and apparent association with nosocomial transmission in an intensive care unit. AB - Bacillus cereus has sometimes been implicated in food poisoning and in opportunistic infections of seriously ill patients. This report describes an unusual case of persistent bacteremia and multiple organ failure associated with B. cereus in a patient admitted to our institution for lung cancer. The patient was undergoing treatment with an antimicrobial agent (imipenem) that was shown to be effective against the micro-organism in vitro. No portal of entry for the strain was detected. After treatment with vancomycin, also shown to be effective in vitro, no clinical improvement was noted and the patient died. Molecular studies showed that the same strain caused an episode of pseudobacteremia in another patient admitted to the same ICU room. PMID- 10716091 TI - Bipolaris spicifera meningitis complicating a neurosurgerical procedure. AB - Bipolaris spicifera, one of the darkly pigmented (dematiaceous) fungi commonly found in soil, is an uncommon cause of infection in humans and an unusual cause of meningitis and nosocomial infections. An 18-y-old boy who experienced meningitis with this micro-organism after acoustic neuroma resection was successfully treated with amphotericin B. PMID- 10716092 TI - Serum procalcitonin and proinflammatory cytokines in a patient with acute severe leptospirosis. AB - Leptospirosis is a zoonosis, with clinical manifestations ranging from the imperceptible to severe, potentially fatal renal and liver failure accompanied by haemorrhage and jaundice. In this case report of a patient with severe leptospirosis, serum levels of procalcitonin decreased ahead of any obvious clinical improvement, and thus may be useful as a prognostic marker. Levels of soluble IL-2 receptor were very high and correlated well with the clinical course. PMID- 10716093 TI - Intraperitoneal treatment of peritoneo-venous shunt infection in a cancer patient. PMID- 10716094 TI - An agency follow-up outcome study of graduates from four inner-city therapeutic community programs. AB - Using its own resources, a follow-up outcome study of 83 out of 119 (70%) graduates from four therapeutic community programs at two inner-city sites was conducted using the Tennessee Self Concept Scale:2 (TSCS:2) and the Post Treatment Follow-Up Survey (PTFUS). The TSCS:2 and the PTFUS were completed anonymously by the respondent graduates and collected by research staff independently of program administrative staff and clinical staff. On average, the graduates had completed all formal agency required treatment and had been living independently in the community for nearly 12 months at the time of follow-up. Graduates who participated in the study were separated into one of three TSCS:2 profile groupings: Valid TSCS Profile, Invalid Faking Good (FG), and Invalid Inconsistency, to provide a more meaningful analysis of their outcomes data. All three TSCS:2 groups had favorable outcomes; however, to the extent that PTFUS questions were less specific than more specific, the Invalid FG group self reported more positive outcomes. Where the PTFUS questions were very specific and concrete, the Valid TSCS Profile group generally had the most favorable outcomes. Implications of these findings are discussed. PMID- 10716095 TI - Cost-effectiveness of mental health services for persons with a dual diagnosis: a literature review and the CCMHCP. The Cost-Effectiveness of Community Mental Health Care for Single and Dually Diagnosed Project. AB - People suffering from comorbid mental illness and substance abuse disorders (the dually diagnosed) are thought to constitute large portions of clients treated as outpatients by public-sector community-based mental health providers. These providers dispense units of ambulatory mental health services and treatments incrementally to maintain clients in the community and out of psychiatric hospitals. Community maintenance is one step, albeit critical, toward quitting drugs and eventual abstinence. Thus, there is a need for information that compares the effectiveness and cost of such services on dually diagnosed clients to identify appropriate low-cost high-yield treatment and service options and packages. This article provides a review of the literature on the effectiveness of ambulatory mental health services and recent emergent reports of cost effectiveness of programs for the dually diagnosed, paying special attention to the gray areas and gaps. This article also describes a new project; an inexpensive add-on to an existing community mental health center. The project will be examining over 4 years of data to compare influence and cost of different ambulatory mental health services and treatments delivered to a matched pair group of clients with dual disorders and those with only mental illness. The intention of this project is not only to address gray areas and gaps in the literature, but also to inform a more rational deployment of mental health services. PMID- 10716096 TI - The Addiction Severity Index: a field study of internal consistency and validity. AB - This study investigated whether the use of the Addiction Severity Index (ASI) in a network of inner-city alcohol and drug abuse clinics under nonideal conditions would yield internally consistent and valid data. A sample of 8,984 ASI scores was collected over a 34-month period. Construct validity was examined by computing the internal consistency of all subscales. Convergent and divergent validity of composite scores and of severity ratings were evaluated using correlation matrices. Findings demonstrated that ASI scores were internally consistent and valid, even though the recommended administration protocol may not always have been followed as faithfully as might be desirable. This robustness bodes well for the use of the ASI in on-line clinical environments. Results should be viewed with caution until the reliability of ASI administration is tested under similar nonideal conditions and until permissible deviations from standard protocol can be quantified. PMID- 10716097 TI - Case management for dually diagnosed individuals involved in the criminal justice system. AB - A case-management model for individuals with substance abuse and mental health disorders who are involved in the criminal justice system is described, based on the experience of a rural demonstration project. Detailed descriptions of case management activities and the philosophy underlying this model of case management are provided. A major goal of these case-management services was to improve access to appropriate treatment for the target population. Evaluation data describing the population served, case-management implementation, and outcomes are presented along with a case vignette. Six-month follow-up data revealed significantly fewer legal problems and apparent symptom relief for participants in the project. Participants reported improvement in most life areas measured compared to the year before, and were generally satisfied with the case management services. Barriers observed in implementing these types of services and issues for replication are outlined and discussed. PMID- 10716098 TI - Estimating the willingness to pay for drug abuse treatment: a pilot study. AB - Previous economic studies of the benefits of drug treatment have limited their estimation to tangible benefits, and thus have underestimated the benefits of drug treatment. The willingness-to-pay (WTP) approach is a more encompassing benefit valuation method that captures both tangible and intangible benefits and accords with valuation concepts used by economists. In this study, we report the results of a pilot study in which we used the contingent valuation (CV) method to value drug treatment. We conducted mall intercept surveys in two communities: the Triad area in North Carolina and Brooklyn, New York. We estimated WTP models for two different drug treatment programs: a program for all drug users and a program specifically targeted to women drug users. We modeled respondents' WTP for drug treatment as a function of their demographics and to responses from attitudinal/experience questions. The mean WTP for both types of drug treatment programs was estimated to be approximately $37 per respondent. Finally, we demonstrated how the results of the CV method may be used in a benefit-cost analysis of drug treatment. PMID- 10716099 TI - Depressive symptoms, stress, and coping among women recovering from addiction. AB - This article focuses on the variability in well-being of 102 women in continuous recovery from addiction for 1 to 5 years. Univariate and bivariate analyses of cross-sectional data on recent depressive symptomatology, and psychosocial stress and coping strategies before and during recovery yielded the following findings: (a) Nearly a third of the sample reported scores above the 16-point cut-off on the Center for Epidemiologic Studies Depression Scale, indicating risk for depression; (b) over half had a history of diagnosed depression; (c) perceived stress in 16 life domains significantly decreased from prerecovery to recovery; (d) by recovery, participants significantly increase their use of positive strategies, but they continued use some negative ones; and (e) risk for high depressive symptomatology was greatest among those who were married or cohabiting, had a history of clinical of depression, high perceived stress in areas of money and emotional and physical health. Findings are discussed in terms of their implications for treatment and aftercare. PMID- 10716100 TI - Stability in the drinking habits of older problem-drinkers recruited from nontreatment settings. AB - Few prospective studies have examined older problem-drinkers not currently in treatment to determine the stability in alcohol problems over time. Seventy-eight currently drinking, older adults meeting a diagnosis of alcohol abuse or dependence were recruited via advertising to complete a health interview; 48 were reinterviewed approximately 3 years later. Participants were categorized based on alcohol consumption (risk) and alcohol-related diagnostic symptoms (problem) at baseline and follow-up. At follow-up, few older adults (11.4%) were resolved using both risk and problem criteria. Alcohol risk/problem groups were not significantly stable between baseline and follow-up. Health problems was the most common reason for changing drinking habits. Average and maximum consumption at baseline and follow-up were significant markers of follow-up risk group and follow-up alcohol-related consequences, respectively, with maximum consumption being more robust. The course of alcohol problems among older adults fluctuates over time, and heavy drinking appears to be the best indicator of problem continuation. PMID- 10716101 TI - Testosterone abuse and affective disorders. PMID- 10716102 TI - Rapid opiate detoxication in outpatient treatment: relationship with naltrexone compliance. AB - A variety of detoxification methods have been utilized for the treatment of heroin withdrawal before individuals begin long-term opiate-free and naltrexone programs. While methadone in decreasing doses is still widely used for detoxication procedures, rapid and ultrarapid protocols including clonidine and opiate receptors antagonists have been proposed. This study compares the efficacy of different detoxification methods and investigates possible changes in naltrexone compliance. Ninety-eight heroin-addicted individuals were studied to evaluate withdrawal symptoms, craving, mood, urine toxicologic screens, and drop out rate during therapy with: Group A: clonidine only (5 days); Group B: clonidine, oxazepam, baclofen, and ketoprofene with naloxone and naltrexone (2 days); and Group C: methadone in decreasing doses (10 days). Naltrexone compliance and relapse rates were evaluated during a 6-month follow-up period. Rapid detoxification with opiate antagonists (Group B) induced slight and transient withdrawal symptoms, and resulted in a significantly lower percentage of heroin catabolites in urine controls during the detoxification procedure, lower negative and positive craving, less mood problems, and higher compliance in extended naltrexone treatment. In comparison with clonidine only (Group A) and methadone (Group C), the early use of naltrexone during detoxification in combination with benzodiazepines and clonidine facilitated extended naltrexone acceptance and improved the recovery outcome in outpatients. PMID- 10716103 TI - Brief substance use screening instruments for primary care settings: a review. AB - Chemical dependence, including nicotine, alcohol, prescription drugs, and illicit drugs, is one of the leading causes of morbidity and mortality in the United States. Primary care physicians and nurses routinely provide preventive health care and rely on routine screening to detect diseases and promote wellness. These primary care practitioners are in a unique position to assess and detect such dependence at its earliest stages. However, previous research indicates that little such screening is actually conducted. This literature review gathered and examined substance use screening instruments in four categories to assess their feasibility for use in primary care settings. Although substance use screening tools are available, most are not appropriate for screening in a primary care setting. There clearly remains a need for the development of a valid, reliable screening instrument that can be easily incorporated into the practices and procedures found in primary care settings. PMID- 10716104 TI - Ethical dilemmas in managed care. PMID- 10716105 TI - The association of temporomandibular joint pain with abnormal bone marrow in the mandibular condyle. AB - PURPOSE: This study investigated the association between temporomandibular joint pain and bone marrow alterations in the mandibular condyle seen on magnetic resonance (MR) images. PATIENTS AND METHODS: The study was based on 112 temporomandibular joints in 112 patients with disc displacement without reduction. Thirty-four patients with abnormal bone marrow on MR images were compared with a control group of 78 patients with normal bone marrow. The analysis was based on proton density and T2-weighted MR images in the oblique sagittal and coronal planes. The degree of pain was correlated to the status of the bone marrow using statistical methods. RESULTS: The degree of pain in joints with abnormal bone marrow was higher than in joints with normal bone marrow signal on MR images (P = .0045). CONCLUSION: Because the stage of internal derangement was similar in both groups, more intensive pain appears to be associated with bone marrow alterations. PMID- 10716106 TI - Occlusal results after open or closed treatment of fractures of the mandibular condylar process. AB - PURPOSE: This study compared the occlusal relationships after open or closed treatment for fractures of the mandibular condylar process. PATIENTS AND METHODS: A total of 137 patients with unilateral fractures of the mandibular condylar process (neck or subcondylar), 77 treated closed and 65 treated open, were included in this study. Standardized occlusal photographs obtained at several postsurgical time intervals were examined and scored by a surgeon and an orthodontist. Standard statistical methods were used to assess differences between groups. RESULTS: Patients treated by closed techniques had a significantly greater percentage of malocclusion compared with patients treated by open reduction, in spite of the fact that the initial displacement of the fractures was greater in patients treated by open reduction. CONCLUSIONS: Based on this study, more consistent occlusal results can be expected when fractures of the mandibular condylar process are treated by open reduction. PMID- 10716107 TI - Resorbable fixation techniques for genioplasty. AB - PURPOSE: This study evaluated the capability and effectiveness of resorbable bone fixation devices in genioplasty surgery. MATERIALS AND METHODS: Twenty patients underwent different genial movements that were stabilized with either 2.5-mm polylactic-polyglycolic acid lag screws or 2.0-mm polylactic-polyglycolic acid plates and screws. RESULTS: Twenty-one anterior mandibular osteotomies were performed in 20 patients. Sixteen patients had advancement (80%), 2 had horizontal setback (10%), and 2 had vertical reduction (10%). The average advancement was 7.6 mm (range, 4 to 14 mm), the average horizontal setback was 6.0 mm (range, 4 to 8 mm), and the average vertical reduction was 7.0 mm (range, 5 to 9 mm). Fixation was done using the lag screw technique in 13 patients (65%) and plate and screw fixation in 7 patients. (35%) Intraoperative stability was satisfactory in all cases. There were no postoperative infections or segmental instability up to 6 months after surgery. CONCLUSION: Resorbable polylactic polyglycolic acid lag screw and plate and screw fixation is a viable alternative for fixation of anterior horizontal osteotomies of the mandible. PMID- 10716108 TI - Complication rates associated with different treatments for mandibular fractures. AB - PURPOSE: This study compared the complication rate with different types of mandibular fracture treatment (maxillomandibular fixation, 2-mm miniplates, 2.4 mm AO plates, and 2.7-mm AO plates). PATIENTS AND METHODS: A total of 245 patients who presented with 386 fractures were retrospectively analyzed. Patient characteristics, type of fracture, severity of fracture, type of treatment used, and occurrence of complications were recorded. Statistical analysis was used to compare complication rates, fracture severity, and type of treatment. RESULTS: There were no differences in the complication rates for the different types of treatment. There was a significant correlation (P < .05) between fracture severity and the overall complication rate, postoperative infection, and postoperative malocclusion, but there was no significant correlation between these complications and the type of treatment applied. CONCLUSION: The occurrence of postoperative complications in the treatment of mandibular fractures is fundamentally related to the severity of the fracture rather than to the type of treatment used. PMID- 10716109 TI - Assessment of the pharyngeal airway space after mandibular setback surgery. AB - PURPOSE: This retrospective study evaluated the change in pharyngeal airway space associated with surgical mandibular setback. PATIENTS AND METHODS: Lateral cephalograms of 14 adult patients taken preoperatively, immediately postoperatively, and at long-term follow-up were traced, and the width of the pharyngeal airway space and the pharyngeal airway space area were calculated and compared. RESULTS: At long-term follow-up, the mean amount of mandibular setback was 9.7 mm. The mean reduction in the distance from the tongue base to the posterior pharyngeal wall was 4.77 mm (28% decrease). The mean reduction in pharyngeal airway space area was 1.52 cm2, which corresponded to a 12.8% reduction. There was a strong correlation between the amount of mandibular setback and the decrease in pharyngeal airway space area. CONCLUSION: Mandibular setback surgery causes a long-term decrease in pharyngeal airway space area. In patients who have other risk factors, for example, overweight, short necks, or large tongues, a mandibular setback procedure could possibly predispose to the development of sleep apnea syndrome. PMID- 10716110 TI - Long-term evaluation of estimates of need for third molar removal. AB - PURPOSE: The aim of this study was to evaluate the estimates on need for third molar removals made at age 20 after 12 years. PATIENTS AND METHODS: The series consisted of 81 university students followed from age 20 to 32 years. At baseline and at study end, these students were clinically examined, and panoramic radiographs were taken. At baseline in 1982, a qualified oral surgeon had made estimates on need for removal of third molars within 5 years; 75% of students needed removals. Actual treatment performed was evaluated after 12 years. A questionnaire served to determine symptoms related to third molars during the 12 year period. RESULTS: During the follow-up, one or more third molars had been removed from 67% of the former students. A total of 155 third molar removals had been estimated, but by age 32 years the percentage actually removed was only 59%. Of the 79 third molars taken out at the Finnish Student Health Service, 77% were initially estimated to need a surgical procedure, but actually 66% were simply extracted. Most were removed at around age 27 years. According to the questionnaire, 67% of the students were asymptomatic in the third molar region during 12 years. CONCLUSION: Because need for surgical removal decreases during early adulthood, routine prophylactic extraction of asymptomatic third molars in young adults cannot be recommended. Well-defined indications for prophylactic removals are needed. PMID- 10716111 TI - Histologic analysis of prefabricated, vascularized bone grafts: an experimental study in rabbits. AB - PURPOSE: The purpose of this study was to investigate the cellular quality of prefabricated bone grafts. MATERIALS AND METHODS: Small pieces of iliac bone were placed around the neurovascular bundle on the dorsal aspect of the ear of 11 one month-old baby rabbits to create a prefabricated vascularized graft. In five animals, the prefabricated bone grafts were harvested for histologic examination 12 months later. In a second group of 6 rabbits, the prefabricated bone grafts, with the neurovascular bundle as a pedicle, were transferred after 30 days to a defect created by removing 1 cm of the midportion of zygoma on the right side of the face. The transferred bone was removed for histologic examination 11 months later. RESULTS: In both groups, microscopic examination showed the presence of a rich, vascular network and similar histologic characteristics to those of normal iliac bone. CONCLUSION: The findings support the concept that prefabricated bone grafts are a potentially useful source for bony reconstruction. PMID- 10716112 TI - Quantification of growth factor levels using a simplified method of platelet-rich plasma gel preparation. AB - PURPOSE: This study compared two methods of preparing platelet-rich plasma (PRP) gel and the levels of PDGF and TGFbeta in each preparation. MATERIALS AND METHODS: Platelet-rich plasma gel was prepared by centrifugation and clotted using the ITA gelling agent (Natrex Technologies Inc, Greenville, NC) or by the addition of thrombin and calcium chloride. The levels of platelet-derived growth factor (PDGF) and transforming growth factor beta (TGFbeta) generated by clot formation were assayed by enzyme-linked immunoassay (ELISA). RESULTS: Both methods of preparation yielded PRP gel in less than 30 minutes. However, the ITA preparation did not require thrombin to achieve adequate gel formation. The levels of PDGF and TGFbeta were similar regardless of which method was used for initiation of clot formation. CONCLUSION: Use of ITA for gel preparation is equivalent to using calcium chloride and thrombin, without the need for special equipment and the risk of coagulopathy. PMID- 10716114 TI - The sagittal split ramus osteotomy as the preferred treatment for mandibular prognathism. PMID- 10716113 TI - Iron-dependent generation of free radicals: plausible mechanisms in the progressive deterioration of the temporomandibular joint. AB - PURPOSE: The purposes of this study were 1) to determine whether iron concentrations detected in temporomandibular joint (TMJ) lavage fluid samples obtained from symptomatic patients are sufficient to catalyze the degradation of specific extracellular matrix (ECM) molecules in vitro, and 2) to provide evidence of oxidative stress in symptomatic TMJs by the detection of protein carbonyls in lavage fluids. PATIENTS AND METHODS: Iron concentrations in TMJ lavage samples (19 joints in 14 patients) were determined colorimetrically, and the ability of the sample to produce free radicals in the presence of hydrogen peroxide was determined with the chromogen 2,2'-azinobis (3-ethylbenzothizoline-6 sulfonic acid), diammonium salt (ABTS). The presence of oxidized proteins was measured fluorimetrically using Bodipy FL hydrazide (Molecular Probes, Eugene, OR). Degradation of fibronectin was visualized by Western blot. Relative susceptibilities of fibronectin and collagen I to free radical cleavage were measured with the Fenton reaction. RESULTS: Redox-active iron concentration in lavage samples was found to be as high as 3.66 micromol/L. A 70-kd protein band, presumed to be albumin, was found to contain higher levels of carbonyls than peripheral serum albumin, which correlated with a greater degree of oxidative damage. Fibronectin was found to be more susceptible than collagen I to free radical degradation, and fragments of the former were found in the lavage. The TMJ lavage fluid was capable of producing free radicals in the presence of peroxide. CONCLUSION: Circumstantial evidence is provided that the presence of modified and cleaved proteins isolated from lavage of symptomatic TMJs may have been subjected to oxidative stress. PMID- 10716115 TI - Intraoral vertical ramus osteotomy as the preferred treatment for mandibular prognathism. PMID- 10716116 TI - Painless mass in the parotid region. PMID- 10716117 TI - Sigmund Freud: psychoanalysis, cigars, and oral cancer. PMID- 10716118 TI - Rectal adenocarcinoma metastatic to the masseter muscle. PMID- 10716119 TI - A case of metastatic calcinosis of the oral cavity. PMID- 10716120 TI - Post-traumatic hemorrhage in a patient with previously undiagnosed von Willebrand's disease. PMID- 10716121 TI - Aggressive form of cherubism: report of a case. PMID- 10716122 TI - Malignant granular cell tumor of the masseter muscle: case report. PMID- 10716123 TI - Use of a stereolithography model for accurate, preoperative adaptation of a reconstruction plate. PMID- 10716124 TI - A simple template for orbital floor defects. PMID- 10716125 TI - A case for prophylactic removal of impacted third molars in young patients. PMID- 10716126 TI - Inflammation, hormones, the blood and the heart; are cardiologists learning to be internists again? PMID- 10716127 TI - Haematological abnormalities in rheumatic mitral valve disease. PMID- 10716128 TI - Inflammatory mediators in heart failure. AB - Cytokines have been identified as a major player in the pathogenesis and the functional status of patients with heart failure. Herein I review studies which are under way to assess the effects of anti-cytokine therapy, where soon we may see treatments directed against bacteria in the bowel, the translocation process, and endotoxin itself, the binding sites of bacterial endotoxin on immune competent cells, or both. PMID- 10716129 TI - Myocardial revascularisation by laser. AB - Myocardial revascularisation by laser is an emerging treatment for refractory angina in patients with coronary artery disease that is not amenable to conventional revascularisation. With the original technique, laser energy was applied to epicardial surface of the heart through a lateral thoracotomy; so called transmyocardial laser revascularisation (TMR). It is now possible, using catheter-based percutaneous myocardial revascularisation (PMR) to deliver laser energy to the myocardium via the endocardial surface. In this article we discuss the possible mechanism of action of laser revascularisation, and summarise the results of recent randomised controlled trials of TMR, PMR systems are described, and the growing evidence for their efficacy is reviewed. PMID- 10716130 TI - Radiofrequency catheter ablation of tachycardia in children with and without congenital heart disease: indications and limitations. AB - From 1993 to 1998, a total of 100 consecutive pediatric patients with tachycardia (45 male and 55 female, aged 1 year 10 months to 17 years, 11+/-4 year) who underwent electrophysiological study were reviewed. Eleven of them were younger than 5 years. Two had tachycardia-related cerebrovascular accident. Congenital heart disease was found in 12 patients. After propofol anesthesia, the clinical tachycardia could not be induced in three (two atrial tachycardia and one AV nodal re-entrant tachycardia) and became nonsustained in five (atrial tachycardia). Mechanical ablation occurred in three and two had subsequent recurrences. Among the 85 cases who received radiofrequency ablation, the overall final success rate of RF ablation for all diagnoses was 94% with a diagnosis specific success rate ranging from 100 to 57%. Tachycardia cardiomyopathy was noted in four (three atrial tachycardia and one junctional ectopic tachycardia) and all regressed after successful ablation. Success in two patients with left posterioseptal accessory pathway could only be achieved by delivering the energy at the middle cardiac vein. Two patients with right atrial isomerism had an 'AV nodal-to-AV nodal tachycardia' which was eliminated by ablation. Total recurrence rate was 13% but final success was achieved in all during re-study except the three patients who refused re-intervention. The atrial tachycardia developed in postoperative congenital heart disease was associated with the lowest success rate (57%) and highest recurrence rate (25%). Procedure-related complications occurred in four; two with transient brachial palsy, one with first-degree AV block and one with blood loss requiring blood transfusion. In conclusion, the experience of this single center confirmed the efficacy and safety of radiofrequency catheter ablation in treating pediatric arrhythmias, but the limitations in postoperative arrhythmias and the effects of propofol on tachycardia induction (especially the atrial tachycardia) need to be improved. PMID- 10716131 TI - Mild mitral regurgitation was associated with increased prevalence of thromboembolic events in patients with nonrheumatic atrial fibrillation. AB - Although several studies demonstrated that the presence of significant mitral regurgitation was associated with reduced occurrence of thromboembolism, little data is available concerning the effect of mild mitral regurgitation on the occurrence of thromboembolic events. To evaluate the association between mild mitral regurgitation and thromboembolic events, we reviewed 232 patients' records between January 1996 and September 1997 who had nonrheumatic atrial fibrillation. There were 59 patients (25%) with mitral regurgitation > or = grade 2, 69 patients (30%) with grade 1 mitral regurgitation, and 104 patients (45%) with no mitral regurgitation. Patients with grade 1 mitral regurgitation had significantly higher prevalence of thromboembolic events (28%) than those with mitral regurgitation > or = grade 2 (8%, P=0.006) or those with no mitral regurgitation (11%, P=0.007). A history of previous thromboembolic events were compared between 173 patients with grade 1 mitral regurgitation and those with no mitral regurgitation using the logistic regression analysis adjusted for age, sex, administration of warfarin, and presence of hypertension, diabetes mellitus, structural heart disease, enlarged left atrium (> or = 40 mm), chronic atrial fibrillation, and grade 1 mitral regurgitation. Grade 1 mitral regurgitation (odds ratio=2.689, 95% confidence interval=1.039-7.189, P=0.0434) and no warfarin administration (odds ratio=0.045, 95% confidence interval=0.002-0.242, P=0.0036) were significantly associated with the history of thromboembolic events. The presence of mild mitral regurgitation in nonrheumatic atrial fibrillation was associated with higher prevalence of thromboembolic events. PMID- 10716132 TI - Mitral regurgitation and atrial fibrillation: milder the disease, higher the risk? PMID- 10716133 TI - The sensitivity of transesophageal pacing for screening in atrial tachycardias. AB - Transesophageal atrial pacing and recording performed in 128 patients for palpitations or tachycardia was retrospectively evaluated and compared to the same procedure in 77 routinely evaluated patients after a catheter ablation procedure. The sensitivity and specificity of the described protocol was 74 and 90% respectively. The procedure was well tolerated and a majority of patients could be completely evaluated according to the protocol. The outcome of the first time investigation influenced the subsequent choice of therapy in the studied population. The results suggest that transesophageal pacing is a valuable tool for evaluation of atrial tachycardias with specificity, sensitivity and tolerability comparable to other noninvasive methods used in cardiology. PMID- 10716134 TI - Left ventricular function and autonomic nervous system balance during two different stages of the menstrual cycle. AB - We studied the left ventricular function and cardiac autonomic nervous system balance variations during two different stages of the menstrual cycle. These two variables, as well as plasmatic estradiol and progesterone concentrations, were measured in a drug-free state in 20 women (29+/-6 year-old) with regular menstrual periods. A clinical evaluation, an echo-Doppler and a Valsalva manoeuvre were performed in all the patients on the third day of their menstrual cycle (follicular phase) and three days prior to their next menstrual cycle (luteinizing phase). When comparing the results obtained in these two phases, a statistically significant increase was put forward in plasmatic estradiol (50.6+/ 24 vs. 127.3+/-52.8 pg/ml) and progesterone (0.37+/-0.42 vs. 11.92+/-10.8 ng/ml) concentrations, Valsalva index (1.55+/-0.22 vs. 1.67+/-0.33; P=0.044) and E/A mitral wave ratio (1.63+/-0.36 vs. 1.75+/-0.35, P=0.02). The right and left atrial volumes, left ventricular volumes and ejection fraction were similar in the two menstrual phases studied. We conclude that the autonomic nervous system balance and the left ventricular diastolic function suffer significant changes during the luteinizing phase of the menstrual cycle in normal women. PMID- 10716135 TI - Efficacy and safety of valsartan compared with enalapril at different altitudes. AB - OBJECTIVE: To compare the safety, tolerability, and antihypertensive efficacy of valsartan with enalapril at different altitudes. METHOD: A total of 142 adult Colombian outpatients with mild to moderate essential hypertension were recruited in 3 cities at different altitudes (Bogota at 2600 m, Medellin at 1538 m and Barranquilla at 100 m) and randomized in an open label fashion to receive either valsartan 80 mg once daily or enalapril 20 mg once daily for 8 weeks. Those patients not responding at 4 weeks received additional 1.25 mg indapamide daily during the remaining trial period. The primary efficacy variable was the change in mean sitting diastolic blood pressure (SDBP) from baseline to 4 weeks. Secondary efficacy variables included the change in mean sitting systolic blood pressure (SSBP). The primary criterion for tolerability was the incidence of adverse experiences. RESULTS: Both valsartan and enalapril reduced mean SDBP and SSBP with similar efficacy, independent of altitude. Adverse events irrespective of relationship to trial drug were reported by 12 patients (18.8%) on valsartan and by 15 (23.4%) patients on enalapril. Enalapril was associated with a significantly (P<0.05) higher rate of dry cough and more cases of headache than valsartan. CONCLUSIONS: Valsartan 80 mg once daily is as effective as enalapril 20 mg once daily in reducing blood pressure, with tolerability profile at least as good as enalapril's. PMID- 10716136 TI - Cardiopulmonary exercise parameters in relation to all-cause mortality in patients with chronic heart failure. AB - In this study we analysed the all-cause mortality over a period of maximal 6 years in 60 male patients (age: 63.4+/-8.3 years, mean+/-S.D.), suffering from chronic heart failure with resting left ventricular ejection fraction and E/O2 slope as independent factors. We assessed functional NYHA class (II: n=36, III: n=24), radionuclide left ventricular ejection fraction (29.2+/-10.4%) and peak values of heart rate, O2, CO2, E, anaerobic threshold and exercise duration with an incremental work load test on the treadmill. O2 relative to E was based on the individual slopes of the regression of O2 on E during the first 6 min of exercise. These slopes with other exercise-related variables and factors such as etiology, medication, and NYHA class were analysed with a Cox's Regression Method. A survival time analysis (Kaplan-Meier survival curve) was done to establish the influence of E/O2 slope and left ventricular ejection fraction (both split into above and below median values), as well as their interaction, on survival. From all investigated exercise-related variables. E/O2 slope is the most powerful variable regarding prediction of all-cause mortality in our group of chronic heart failure patients. Concerning risk stratification, the subgroup (n=18) with a relatively high left ventricular ejection fraction (>28%) and flat E/O2 slope (<27.6) had most survivors (77.8%) after about 3 years, while the subgroup (n=12) with a relatively high left ventricular ejection fraction (>28%), but a steep E/O2 slope (>27.6) had least survivors (33.3%). This difference in percentage is highly significant (P=0.0025). The fact that E/O2 slope and left ventricular ejection fraction show comparable main and interaction effects between measures of exercise tolerance (e.g., anaerobic threshold, peak O2, exercise duration) on the one hand, and all-cause mortality on the other, suggests the existence of common sources of variance. Based on our analysis, it is unlikely that effects on all-cause mortality are mediated through phenomena related to exercise tolerance. Therefore, we hypothesize that the effects on exercise tolerance and all-cause mortality both depend on common factors, which cause both cardiac and peripheral organ (c.q. muscular) dysfunctions. Moreover, this study clearly shows that E/O2 slope during incremental exercise is an important prognostic marker for risk stratification in chronic heart failure patients, NYHA class II and III. PMID- 10716137 TI - The carvedilol hibernation reversible ischaemia trial, marker of success (CHRISTMAS) study. Methodology of a randomised, placebo controlled, multicentre study of carvedilol in hibernation and heart failure. AB - BACKGROUND: Carvedilol reduces mortality and improves symptoms and ejection fraction in ischemic heart failure, but its mode of action is not well defined and not all patients respond to treatment. The aim of the CHRISTMAS (Carvedilol Hibernation Reversible Ischaemia Trial, Marker of Success) study is to examine whether hibernation may be a significant factor determining this response. This paper describes the methodology and the rationale for the choice of the nuclear cardiology and echocardiography imaging techniques used in the study. METHODS AND RESULTS: The CHRISTMAS study is a double-blind, randomised, parallel group, multinational study of oral carvedilol versus placebo in patients with chronic stable heart failure due to left ventricular systolic dysfunction from coronary artery disease. The study aims to randomise 400 patients who are on optimal treatment. Two parallel groups will be randomised to carvedilol or placebo, namely 200 with hibernating myocardium at baseline and 200 matched patients without. The presence of hibernation is defined from a mismatch between regional contractile function and regional viability, measured by echocardiography (severe segmental asynergy) and nitrate prepared resting Tc99m-MIBI myocardial perfusion imaging (segmental activity >60%). The primary treatment-related end-point of the study is the comparison of the mean change, from baseline to the final visit, in radionuclide-determined left ventricular ejection fraction in patients on placebo with those on carvedilol, between the groups designated as hibernating and non hibernating. Other end-points being examined include the prevalence of hibernation in heart failure, the relationship between the volume of hibernating myocardium and the ejection fraction response, the prevalence of reversible ischemia in heart failure, and the comparison of echo with gated SPECT. To date, 303 patients have been screened and 251 patients randomised in the study. The study aims to report in 2000. CONCLUSIONS: The CHRISTMAS study addresses the issue of whether the presence of hibernation is a predictor of the ejection fraction response to carvedilol in heart failure. It also examines the potential role of medical therapy in hibernation as well as a number of other end-points. The study may potentially lead to an important new role for nuclear cardiology in heart failure, and demonstrates important synergy between cardiac imaging and the pharmaceutical industry. PMID- 10716138 TI - Endothelial dysfunction associated with left ventricular diastolic dysfunction in patients with coronary heart disease. AB - OBJECTIVE: We sought to assess the correlation between endothelial vasodilation and left ventricular diastolic function. BACKGROUND: Previous studies have demonstrated that similar neurohumoral factors are involved in myocardial and vascular endothelial impairment. The degree of endothelial dysfunction is related to the clinical severity of the heart failure. However, it is not clear whether endothelial dysfunction develops with the progression of left ventricular diastolic dysfunction. We hypothesize that the endothelial dysfunction is associated with left ventricular diastolic dysfunction. METHODS: Using high resolution ultrasound, we measured the dilator response of the brachial artery to hyperemia (endothelium-dependent vasodilation) and to 0.5 mg nitroglycerin (endothelium-independent vasodilation), and measured peak velocities of the early wave (Evmax) and the atrial wave (Avmax) in 40 coronary heart disease (CHD) patients and 20 normal subjects. We analyzed the relationship between the Evmax/Avmax ratio and endothelium-dependent vasodilation. RESULTS: The results showed that endothelium-dependent and endothelium-independent vasodilation as well as the Evmax/Avmax ratio were lower in the CHD group than those in the control group (4.29%+/-1.42%, 17.58%+/-2.99%, 0.81+/-0.24 vs. 9.62%+/-2.34%, 24.18%+/-3.15%, 1.07+/-0.29, respectively, P<0.01). The Evmax/Avmax ratio was related to endothelium-dependent vasodilation (r=0.45, P<0.01). CONCLUSIONS: Our results showed that the development of endothelial dysfunction was associated with the progression of myocardial diastolic dysfunction, which suggests that the same mechanisms may be involved in the impairment of endothelium and myocardium. PMID- 10716140 TI - Management of acute myocardial infarction in geriatric population. PMID- 10716139 TI - Are there gender differences in patients presenting with unstable angina? AB - BACKGROUND: There are limited studies on gender differences in patients with unstable angina. We investigated the influence of gender in these patients in a tertiary referral centre. METHODS AND RESULTS: Three hundred and thirteen consecutive patients (210 men and 103 women) with unstable angina were studied over a 42-month period. Patient characteristics, cardiovascular risk factors and subsequent management including coronary artery bypass graft (CABG) operation and percutaneous transluminal coronary angioplasty (PTCA) were investigated. There was no difference in age [61.6 (11.0) (S.D.) years for men vs. 63.5 (10.5) years for women]. Diabetes mellitus and hypertension were more common in women (diabetes, 11% vs. 23%, P = 0.007; hypertension, 32% vs. 52%; P = 0.001). The number of smokers was greater in men (73% vs. 46%, P = 0.00001). There was no difference in the prevalence of hypercholesterolaemia or in the incidence of previous myocardial infarction, previous history of angina and family history of ischaemic heart disease. The duration of unstable angina before presentation to the referring hospital was similar in both sexes. The use of aspirin, intravenous heparin and antianginal drugs was also comparable in the two genders. The number of coronary arteries involved in men and women appeared similar (one vessel, 22% vs. 27%; two vessels, 26% vs. 21%; three vessels, 52% vs. 52% in men and women, respectively). The proportion of men and women who underwent subsequent revascularisation was also similar (CABG, 31% vs. 33%; PTCA, 42% vs. 40%). The overall in-hospital mortality was higher in women (6.8% vs. 2.8%), but was not statistically significant (P = 0.18). CONCLUSIONS: Gender differences in unstable angina manifest in the preponderance of selected risk factors including diabetes mellitus and hypertension in women and smoking in men. There is no difference in age, the degree of coronary artery involvement and the subsequent management in a tertiary referral centre. PMID- 10716141 TI - Therapeutic strategy with total coronary artery occlusions. PMID- 10716142 TI - Secondary prevention in early coronary disease. PMID- 10716143 TI - A rise of troponin and/or von Willebrand factor over the first 48 h is associated with a poorer 1-year outcome in unstable angina patients. PMID- 10716144 TI - Anti-cytokine treatment in patients with chronic heart failure--a clinical approach. PMID- 10716145 TI - Physical activity promotes health also among dialysis patients. PMID- 10716146 TI - Pigmented (melanotic) neurofibroma: a clinicopathologic and immunohistochemical analysis of 19 lesions from 17 patients. AB - Neurofibromas with melanin-laden pigmented cells are rare, accounting for less than 1% of all neurofibromas accessioned to the Soft Tissue Registry of the Armed Forces Institute of Pathology between the years 1970 and 1996. This study analyzes the clinicopathologic features associated with 19 specimens removed from 17 patients. Eleven males and six females, ranging in age from 2 to 61 years (median, 28 years), participated in the study. Nine of 15 patients whose race was provided were black. Eight patients (47%) are known to have neurofibromatosis, and two others (12%) are strongly suspected of having this disorder; two patients have similarly affected family members. Eight patients were noted to have multiple skin tumors, and in each of two cases, two pigmented neurofibromas were available for review. Two patients had hypertrichosis and cutaneous hyperpigmentation resembling a hairy nevus, and one had a cafe au lait spot directly overlying a pigmented neurofibroma. Tumors ranged in size from 1.7 to 50 cm in greatest dimension and involved the buttock or leg (n = 6), head or neck (n = 8), trunk (n = 2), wrist or hand (n = 2), and an unspecified site (n = 1). The neurofibromas exhibited diffuse (n = 15), combined diffuse and plexiform (n = 2), combined diffuse and intraneural epithelioid (n = 1), and nonspecific (n = 1) growth patterns. The process involved the skin (n = 14), subcutis (n = 18), and/or skeletal muscle (n = 3). Wagner-Meissner-like bodies were identified in 11 tumors, and mitoses (average, less than one mitosis per 10 high-power fields) were present in three lesions. All examples contained scattered pigmented cells with dendritic, tadpole-shaped, spindled or epithelioid morphology. These cells were positive with Fontana-Masson (nine of nine) and Warthin-Starry (pH, 3.2; four of four) stains, and were depigmented with a melanin bleach method (two of two). An iron stain was negative. The tumors had immunoreactivity for S-100 protein (11 of 11), HMB-45 ( 10 of 11), Melan-A (four of four), tyrosinase (four of four), and CD34 (four of four). Although recurrences are documented, none of the tumors are known to have undergone malignant transformation. A pigmented neurofibroma can be confused with a pigmented dermatofibrosarcoma protuberans (Bednar tumor) because the melanin-laden cells of both processes are similar. However, the latter entity exhibits a more extensive storiform growth, has greater immunoreactivity for CD34, and lacks a diffuse proliferation of S-100 protein-positive Schwann cells. PMID- 10716147 TI - Definition of histopathologic changes in gastroesophageal reflux disease. AB - A series of 71 patients with multiple measured biopsies of the gastroesophageal junctional region permitting assessment of the presence and length of different glandular epithelial types is presented. All but nine of 53 patients in whom a 24 hour pH study was performed had abnormal reflux, suggesting that endoscopic recognition of an abnormal columnar mucosa at the gastroesophageal junction sufficient to precipitate multiple-level biopsies indicates a high probability of abnormal reflux. All patients had cardiac mucosa (CM) or oxyntocardiac mucosa (OCM). CM was present in 68 of 71 patients. The prevalence of intestinal metaplasia increased with increasing CM+OCM length, and was present in all 22 patients with a CM+OCM length >2 cm and in 20 of 49 patients with a CM+OCM length <2 cm. Patients with a CM+OCM length >2 cm had a markedly higher acid exposure than patients with a CM+OCM length <2 cm. The findings suggest that the presence of CM and OCM in the junctional region are predictive of abnormal acid exposure, and that increasing OCM+CM length correlates strongly with the amount of acid exposure. The histologic finding of CM and OCM represents a sensitive histologic criterion for gastroesophageal reflux rather than normal epithelia. These diagnostic criteria represent the first useful histologic definitions for assessing the presence and severity of reflux. PMID- 10716148 TI - Composite hemangioendothelioma: a complex, low-grade vascular lesion mimicking angiosarcoma. AB - Eight cases of a previously uncharacterized vascular neoplasm, showing varying combinations of benign, low-grade malignant, and malignant vascular components are described. Seven tumors occurred in the dermis and/or subcutis and one occurred in the oral submucosa. The patients were all adults with a median age of 39.5 years (range, 21-71 years). Five patients were men. The tumors arose predominantly in the hands and feet, and the lesions were usually of several years duration. The tumors were composed of a complex admixture of histologic components that varied from tumor to tumor, such that no two tumors looked precisely the same. This was due to variation in the proportions of each component as well as the manner in which each component was distributed throughout each lesion. The predominant histologic components were epithelioid hemangioendothelioma (HE) and retiform HE, which were each present in seven of the tumors. Areas of spindle cell HE were identified in four lesions. Angiosarcoma-like elements were identified in seven tumors. One of the tumors was associated with an arteriovenous malformation and one was associated with an area of lymphangioma circumscriptum. Of six cases with follow up (median duration, 6.5 years), three have recurred locally and, to date, only one has metastasized. We think composite HE is best regarded as a low-grade malignant vascular neoplasm, and the available data suggest that it behaves more favorably than conventional angiosarcoma. The existence of these composite lesions has led to careful reexamination of the concept of HE. The term HE, in that it is currently synonymous with a low-grade malignant vascular tumor, should be reserved for lesions that have true metastatic potential, albeit with low frequency. PMID- 10716149 TI - Pituicytoma: a distinctive low-grade glioma of the neurohypophysis. AB - Pituicytoma is a rare, poorly characterized tumor of the sella and suprasellar region that is distinct morphologically from other local tumors and is thought to be derived from neurohypophyseal pituicytes. Clinical data, neuroimaging studies, and microsections were reviewed from nine such low-grade gliomas. Immunostains for glial, neuronal, and proliferation markers were performed on all nine tumors and six control neurohypophyses. Three tumors were studied ultrastructurally. Six men and three women, age 30 to 83 years (mean, 48 years), presented with visual symptoms, headache, or hypopituitarism. Magnetic resonance images showed solid, discrete, contrast-enhancing masses, four within the sella and five in the suprasellar space. The tumors consisted of sheets and/or fascicles of plump spindle cells with slightly fibrillar cytoplasm and slightly pleomorphic, oval-to elongate nuclei with pinpoint nucleoli. Extracellular mucin was prominent in one tumor. Rosenthal fibers, granular bodies, and Herring bodies (granular axonal dilatations characteristic of the normal neurohypophysis) were lacking. Mitoses were rare or absent. MIB-1 labeling indices were low (0.5-2%). Tumor cells were strongly reactive for vimentin and S-100 protein, variably positive for glial fibrillary acidic protein, and nonreactive for synaptophysin and neurofilament protein. Cytoplasm varied in electron density and contained intermediate filaments. Neither meningothelial nor ependymal features were noted. Two tumors recurred at 20 and 26 months after subtotal resection, but none of the six completely resected tumors have done so. Pituicytomas are discrete, largely noninfiltrative low-grade gliomas of the sellar region that occur in adults. Their histologic appearance is distinct from pilocytic and ordinary, infiltrative astrocytomas. The distinction between pituicytoma and normal neurohypophysis is aided by the latter's content of axons, Herring bodies, and perivascular anucleate zones rich in axonal terminations. Although curable by total excision, subtotal resection can be associated with recurrence. PMID- 10716150 TI - Microcystic endocervical adenocarcinomas: a report of eight cases. AB - Eight endocervical adenocarcinomas with a prominent cystic component that resulted in a resemblance, in part, to certain benign lesions are described. The patients ranged in age from 34 to 78 years (average age, 48.6 years), and three women were postmenopausal. Presentations included abnormal cervical cytology smears (n = 4) and vaginal bleeding (n = 3). One patient was pregnant at the time of diagnosis. Six tumors were typical endocervical-type and two tumors were intestinal-type adenocarcinomas. The cysts occupied 50% to approximately 90% of the tumor and ranged from 1 to 8 mm in diameter. They were lined by flattened to low cuboidal to pseudostratified epithelium with focal goblet cells and Paneth cells in the two intestinal-type tumors, and the epithelial lining of the cysts was denuded occasionally. Seven tumors contained luminal mucin, which was brightly eosinophilic in one patient and resembled the contents of mesonephric tubules, but was mucin positive on special stains. A desmoplastic stroma was identified in two patients; the remainder had no stromal reaction. The features that resulted in mimicry of benign lesions were the cystic glands, their sometimes orderly distribution, and focal, deceptively bland cytology. All tumors contained, at least focally however, architecturally abnormal glands lined by cytologically malignant cells. PMID- 10716151 TI - Epstein-Barr virus-negative post-transplant lymphoproliferative disorders: a distinct entity? AB - Post-transplant lymphoproliferative disorders (PTLDs) are usually but not invariably associated with Epstein-Barr virus (EBV). The reported incidence, however, of EBV-negative PTLDs varies widely, and it is uncertain whether they should be considered analogous to EBV-positive PTLDs and whether they have any distinctive features. Therefore, the EBV status of 133 PTLDs from 80 patients was determined using EBV-encoded small ribonucleic acid (EBER) in situ hybridization stains with or without Southern blot EBV terminal repeat analysis. The morphologic, immunophenotypic, genotypic, and clinical features of the EBV negative PTLDs were reviewed, and selected features were compared with EBV positive cases. Twenty-one percent of patients had at least one EBV-negative PTLD (14% of biopsies). The initial EBV-negative PTLDs occurred a median of 50 months post-transplantation compared with 10 months for EBV-positive cases. Although only 2% of PTLDs from before 1991 were EBV negative, 23% of subsequent PTLDs were EBV negative (p <0.001). Of the EBV-negative PTLDs, 67% were of monomorphic type (M-PTLD) compared with 42% of EBV-positive cases (p <0.05). The other EBV negative PTLDs were of infectious mononucleosis-like, plasma cell-rich (n = 2), small B-cell lymphoid neoplasm, large granular lymphocyte disorder (n = 4) and polymorphic (P) types. B-cell clonality was established in 14 specimens and T cell clonality was established in three (two patients). None of the remaining specimens were studied with Southern blot analysis and some had no ancillary studies. Rearrangement of c-MYC was identified in two M-PTLDs with small noncleaved-like features, and rearrangement of BCL-2 was found in one large noncleaved-like M-PTLD. Ten patients were alive at 3 to 63 months (only three patients received chemotherapy). Seven patients, all with M-PTLDs, are dead at 0.3 to 6 months. Therefore, EBV-negative PTLDs have distinct features, but some do respond to decreased immunosuppression, similar to EBV-positive cases, suggesting that EBV positivity should not be an absolute criterion for the diagnosis of a PTLD. PMID- 10716152 TI - Giant cell tumors of soft tissue: a clinicopathologic study of 18 benign and malignant tumors. AB - Primary giant cell tumors (GCTs) of soft tissue resembling osseous GCTs are uncommon but distinct entities. Malignant GCTs of soft tissue have been designated giant cell malignant fibrous histiocytomas; however, there is scant data regarding benign GCTs of soft tissue. Eleven benign and seven malignant GCTs of soft tissue were identified from the authors' consultation files and the surgical pathology files of the Vancouver General Hospital and Massachusetts General Hospital. The tumors occurred in adults (eight men, 10 women; age range, 25-89 years; mean age, 54 years) in the extremities (n = 14) and in the trunk, abdomen, and pelvis (n = 4). In each patient the skeleton was normal and there was no history of prior osseous GCT. Tumors ranged in size from 0.8 to 9.0 cm. Eleven occurred in the superficial soft tissue and seven occurred in deep soft tissue. Grossly they were circumscribed and frequently hemorrhagic. Cystic change was present in seven tumors. Nine tumors were partially surrounded by a shell of reactive bone. In all tumors, multinucleated osteoclast-like giant cells were distributed uniformly and evenly among mononuclear cells. The histologically benign GCTs of soft tissue were identical to typical osseous GCTs. The mononuclear cells in these tumors lacked nuclear atypia or pleomorphism, and the mitotic rate within this population was low (mean, three mitoses per 10 high power fields [HPF]). In the malignant GCTs of soft tissue, the mononuclear cells exhibited anisocytosis, nuclear atypia, pleomorphism, and readily detectable mitoses including atypical forms (mean, 25 mitoses per 10 HPF). None of the benign or malignant tumors exhibited neoplastic bone production. The benign and malignant GCTs of soft tissue demonstrated a similar immunohistochemical staining profile to GCT of bone ( 12 tumors examined), exhibiting strong positive staining for CD68 within multinucleated osteoclastlike cells, and focal staining of mononuclear cells for CD68, Ham 56, and smooth muscle actin. All tumors were treated by surgical resection. Follow-up information is available for 15 patients (range, 0-108 months). No benign tumor has recurred or metastasized. Of the four patients with malignant tumors for whom follow-up information is available, one died of metastatic disease at 13 months and one developed a local recurrence at 84 months but is alive, apparently free of disease after additional excisional surgery. Primary GCTs of soft tissue are distinctive neoplasms that, like osseous GCTs, exhibit a wide clinicopathologic spectrum. These neoplasms should be distinguished from other giant cell-rich soft-tissue tumors with which they may be confused. PMID- 10716153 TI - Myxoid neoplasms of the adrenal cortex: a rare histologic variant. AB - The myxoid variant of adrenocortical carcinoma is a rare neoplasm described previously in only two case reports. Because of the rarity of these lesions, the presence of myxoid changes in adrenal cortical neoplasms usually raises the possibility of malignancy. We studied the histopathologic features of 14 cases of myxoid adrenocortical neoplasms, including six adenomas and eight carcinomas. All patients with adenomas with sufficient follow-up (n = 5) were alive with no recurrence of their tumors or evidence of metastatic disease. Four patients with carcinomas died of their disease, two were alive with metastatic disease, and one was alive with no evidence of recurrence or metastatic disease. Histologically, the 14 tumors varied in their myxoid composition, ranging from 10% to 95%. The myxoid foci stained positively with Alcian blue and were usually negative with periodic acid-Schiff and mucicarmine stains. As a group, the immunophenotype of the lesions was typical of other adrenal cortical neoplasms, with positive immunostaining for vimentin, synaptophysin, and alpha-inhibin. One tumor was focally positive for keratin. Myxoid adrenal cortical neoplasms should be included in the differential diagnosis of myxoid retroperitoneal neoplasms. Myxoid changes in adrenal cortical neoplasms may be present in both adenomas and carcinomas, and the usual clinical and histopathologic features for adrenocortical neoplasms should be used to diagnose these neoplasms. PMID- 10716154 TI - Histology of the gastroesophageal junction: an autopsy study. AB - Current diagnostic criteria for reflux disease and Barrett's esophagus are based on the belief that the gastroesophageal junction normally contains 2 cm of cardiac mucosa composed of mucous glands devoid of parietal cells. This autopsy study disproves this belief. Even when the entire circumference of the gastroesophageal junction is examined, pure cardiac mucosa was completely absent in 56% of patients. All patients had oxyntocardiac mucosa, in which glands contained a mixture of mucous and parietal cells. Cardiac and oxyntocardiac mucosae were present only in part of the circumference of the junction in 50% of patients. The measured maximum length of cardiac plus oxyntocardiac mucosa was less than 0.5 cm in 76% of patients. There was a tendency for the presence and extent of cardiac mucosa to increase with age. Cardiac mucosa at the junction is therefore frequently absent, has considerable individual variation, is very small in extent when present, is commonly absent from some part of the circumference of the junction, and increases in prevalence and length with age. These characteristics of cardiac mucosa make it highly unlikely that it is a normal structure. We develop the hypothesis that cardiac mucosa represents an early histologic manifestation of gastroesophageal reflux. PMID- 10716155 TI - Cytokeratin immunoreactivity in Ewing's sarcoma: prevalence in 50 cases confirmed by molecular diagnostic studies. AB - Ewing's sarcoma (ES) and primitive neuroectodermal tumor (PNET) are characterized by the presence of the specific t(11;22)(q24;q12) or variants thereof, producing diagnostic EWS fusion transcripts. Cytokeratin has been reported sporadically to be expressed in some cases of ES/PNET. However, its prevalence has not been assessed systematically in a series of cases with confirmatory molecular or cytogenetic evidence of a diagnostic translocation. We present in detail three index patients in whom strong cytokeratin immunoreactivity was a confounding factor in the diagnosis. To establish further the prevalence of cytokeratin immunoreactivity in a series of well-characterized ES/PNET, we then performed immunohistochemical studies with antibodies CAM5.2 and AE1/AE3 on 50 cases of ES/PNET diagnosed at Memorial Sloan-Kettering Cancer Center in which molecular evidence of a specific ES/PNET-associated translocation were available. Immunoreactivity to cytokeratin was present in 10 cases (20%), in five diffusely and five focally. There was no significant association between cytokeratin expression and the following parameters: patient age, sex, skeletal and extraskeletal primary site, and the type of EWS fusion transcript. Cytokeratin expression, a manifestation of epithelial differentiation, is present in as many as 20% of ES/PNET in either a diffuse or focal pattern. PMID- 10716156 TI - Telomerase activity in pulmonary neuroendocrine tumors: correlation with histologic subtype (MS-0060). AB - The authors measured telomerase activity using the telomeric repeat amplification protocol-enzyme-linked immunosorbent assay method in 13 neuroendocrine pulmonary neoplasms and in non-neoplastic frozen lung samples from the same patients. These cases belonged to the complete neuroendocrine neoplastic spectrum: four typical carcinoids, three atypical carcinoids, four large cell neuroendocrine lung carcinomas, and two small cell lung carcinomas. The authors performed the same assay for 52 non-neoplastic lung tissues from the surgical files in their department (negative controls). They verified the presence (or absence) of neoplastic tissue in every case by looking at one frozen section done in the same tissue used for telomerase assay. The telomerase activity level in non-neoplastic tissues (mean, 182 A450nm U) was similar to that obtained in the typical carcinoids (mean, 104.5 A450nm U). All neuroendocrine tumors but the typical carcinoids showed high levels of telomerase activity (mean, 1,750.8 A450nm U). According to the telomerase hypothesis, typical carcinoid cells are mortal pre-M2 stage cells, but atypical carcinoid, large cell neuroendocrine lung carcinoma, and small cell lung carcinoma cells are immortal post-M2 stage cells. This finding may be of important prognostic significance in these kinds of tumors. Measurement of enzyme activity with a good morphologic control could be necessary in telomerase activity assay. PMID- 10716157 TI - Microinvasive carcinoma (T1mic) of the breast: clinicopathologic profile of 21 cases. AB - Clinicopathologic data on microinvasive carcinoma of the breast (MICB) as defined by the 1997 TNM criteria (T1mic < or = 1 mm) is scarce. Histologic slides of 109 cases from 1993 through 1997, in which microinvasion was either suspected or diagnosed initially, were reviewed. A double immunoenzyme-labeling technique using antismooth muscle actin and anticytokeratin antibody on the same section was used to confirm invasion in equivocal cases. All foci of invasion were measured by ocular micrometer. Twenty-one cases were confirmed to be MICB. The mean age of the patients was 60.9 years. Thirteen patients presented with mammographic abnormalities on routine examination (60.9%). MICB was ductal in 18 patients, including one tubular carcinoma, and was lobular in three patients. The mean number of invasive foci was two per patient (range, one to seven foci). The accompanying duct carcinoma in situ had high-grade nuclei and necrosis in 16 of 18 patients (89%), 13 of which (72%) were comedo-type. Two of the 15 patients had one positive axillary lymph node each (13.3%). Eleven patients underwent mastectomy, nine received radiation therapy, one received chemotherapy, and two underwent lumpectomy only. Median follow up was 28 months (range. 18-63 months). One patient had a chest wall recurrence of infiltrating duct carcinoma and another recurred with duct carcinoma in situ. PMID- 10716158 TI - Vulvar intraepithelial neoplasia of the simplex (differentiated) type: a clinicopathologic study including analysis of HPV and p53 expression. AB - The simplex (differentiated) variant of vulvar intraepithelial neoplasia (VIN) has not been well characterized. The authors studied the clinicopathologic features of 12 cases of simplex VIN and obtained follow-up data to assess its relationship to vulvar invasive squamous cell carcinoma (InvSCC). Expression of p53 protein was analyzed immunohistochemically and compared with adjacent non neoplastic epidermal lesions. Assessment of human papilloma virus (HPV) deoxyribonucleic acid was done by polymerase chain reaction amplification and in situ hybridization. All patients were of postmenopausal age (mean age, 66.8 years). Three patients had a history of prior vulvar InvSCC and one had a separate synchronous vulvar InvSCC. Squamous hyperplasia was present in the adjacent epidermis in 10 patients and lichen sclerosus (LS) was present in four patients. Histologically, simplex VIN differed from "classic" VIN by its highly differentiated features. The characteristic features included parakeratosis, thickened epidermis with elongated and anastomosing rete ridges, enlarged abnormal keratinocytes with premature eosinophilic cytoplasmic differentiation extending deeply within the epidermis, whorling of enlarged keratinocytes or keratin pearl formation within rete ridges, prominent intercellular bridges, and dysplastic basilar cells. One patient had minimal microinvasion (0.6 mm). Ten of 12 patients had positive p53 immunostaining staining with suprabasilar extension of p53 positive cells in each patient. The labeling index (LI) of basilar cells ranged from 0% to 99% (median, 94.5%). Non-neoplastic lesions in the adjacent epidermis had p53-positive basal cells in nine of 11 evaluable cases. The LI was significantly lower in these lesions, with a median of 4% in squamous hyperplasia and 7.5% in LS; none had suprabasilar extension of p53-positive cells. HPV (type 31/35/51) was identified in only one simplex VIN--a p53-negative lesion. Staining for p53 often delineated sharply the junction between simplex VIN and squamous hyperplasia. Four patients subsequently developed vulvar InvSCC at 5, 6, 9, and 55 months. All four InvSCCs were of the conventional keratinizing type and were HPV negative, as were the one synchronous and two prior InvSCCs. The authors conclude that (1) simplex VIN has a strong association with vulvar InvSCC and is a probable precursor lesion of HPV-negative vulvar InvSCCs, (2) HPV is very uncommon in simplex VIN and probably does not play an important role in its genesis, (3) alteration of the p53 gene appears to be involved in the development of simplex VIN, and (4) immunostaining for p53 protein may be helpful in the differential diagnosis of simplex VIN. PMID- 10716159 TI - A critical examination of the immunophenotype of pulmonary sclerosing hemangioma. AB - The authors studied a series of 21 cases of pulmonary sclerosing hemangioma (SH) to address conflicting and unconfirmed reports of immunohistologic evidence of differentiation that have been made in the literature. They found the lesional cells of SH to be epithelial membrane antigen (EMA) positive (21 of 21 cases), to be keratin positive only infrequently and focally (six of 21), and to be nonreactive for carcinoembryonic antigen, S-100, smooth muscle actin, and CD34. Faint nuclear staining was seen for estrogen receptors, whereas progesterone receptors were expressed strongly in 17 cases. Neuroendocrine markers (chromogranin A, adrenocorticotrophic hormone, human growth hormone, and calcitonin) were negative uniformly on the lesional cells except for one case in which rare chromogranin-positive cells were present and another case in which rare human growth hormone-positive cells were seen. In contrast to the general EMA-positive, keratin-negative phenotype of the lesional cells, the cells lining the papillae or air spaces within the SH were typically positive for both markers. The following other lesions were identified in the cases studied: carcinoid tumorlets (n = 2), a neuroendocrine body (n = 1), and multiple meningothelial-like nodules (n = 1). All were clearly separable from the SH on morphologic grounds. The authors interpreted these to be chance occurrences of unrelated lesions. Recognition of the phenotype of SH as EMA positive, keratin weak to negative, and negative for S-100, smooth muscle actin, and neuroendocrine markers is notable in its differential diagnosis from other lesions. This phenotype does not suggest a precise lineage or type of differentiation for SH. PMID- 10716160 TI - Renal sinus involvement in renal cell carcinomas. AB - The renal sinus is the fatty compartment located within the confines of the kidney not delineated from the renal cortex by a fibrous capsule. Because it contains numerous veins and lymphatics, invasion into this compartment may permit dissemination of a tumor otherwise regarded as renal-limited. Thirty-one consecutive renal carcinomas were studied: 22 clear cell renal cell carcinomas (3 multilocular cystic renal cell carcinomas), 4 chromophobe renal carcinomas, and 5 papillary renal carcinomas. The entire interface between the neoplasm and the sinus was embedded. Seventeen carcinomas did not invade the renal sinus and 16 were pT1 or pT2 tumors. Fourteen carcinomas, 13 clear cell renal cell carcinoma and one chromophobe renal carcinoma, invaded the renal sinus fat, and 9 of 14 invaded the lumen of renal sinus veins (all clear cell renal carcinomas). Although 14 of 22 clear cell renal carcinomas appeared to be renal limited pT1 and pT2 cancers, 6 of 14 carcinomas invaded sinus fat and 4 invaded into the lumen of renal sinus veins. Compared with the nine sinus-negative clear cell renal cell carcinomas, the 13 sinus-positive cancers were larger, exhibited more frequent renal capsule and renal vein involvement, and had higher nuclear grades. Renal sinus invasion was most common in clear cell renal cell carcinomas but was uncommon (one in 12) in 3 more indolent renal cell carcinomas: multilocular cystic renal cell carcinoma, chromophobe renal carcinoma, and papillary renal carcinoma. The follow-up period was short (1-17 months), but metastases developed in four of 31 cases. In three cases with metastases, carcinoma had involved the lumen of sinus veins but not the main renal vein, although two of three had also invaded through the renal capsule. This study shows that in carcinomas which appear to be renal limited (pT1/pT2), seven of 23 (30.4%) had invaded sinus fat and four of 23 (17.4%) had invaded sinus veins. We conclude that renal sinus invasion, especially sinus vein invasion, could identify a patient at risk for metastases even in a putative renal limited tumor, and suggest that all cases be examined for this feature. Renal sinus invasion merits further investigation to establish its prognostic importance and possible incorporation into future revisions of the TNM staging system for renal cell carcinomas. PMID- 10716161 TI - Hepatosplenic T-cell lymphoma of alphabeta lineage in a 16-year-old boy presenting with hemolytic anemia and thrombocytopenia. AB - The authors report an unusual case of peripheral T-cell lymphoma in a 16-year-old boy who presented initially with jaundice, splenomegaly, anemia, and thrombocytopenia. A lymphoma was found subsequently in the spleen, which was infiltrated extensively in the red pulp by medium-sized, blastic-appearing lymphoma cells. Immunologic characterization of these cells revealed positivity for CD3, CD5, CD45RO, CD56, and T-cell intracellular antigen (TIA), and negativity for CD2, CD3, CD4, CD8, CD57, CD34, and terminal deoxynucleotidyl transferase (TdT). Conventional cytogenetic studies revealed the presence of isochromosome 7q. On follow up, this patient deteriorated rapidly, with evidence of liver and bone marrow involvement. Although the overall clinical and pathologic features of this disease were characteristic of hepatosplenic gammadelta T-cell lymphoma, the T-cell receptor of this tumor showed an immunophenotype of alphabeta not gammadelta lineage. Using the Southern blot technique, the authors demonstrated monoclonal gene rearrangement of the T-cell receptor beta-chain. Thus, they confirmed the existence of hepatosplenic alphabeta T-cell lymphoma. In view of its overall similarity to hepatosplenic gammadelta T-cell lymphoma, this unusual entity probably represents a slight biologic variation of the same disease. PMID- 10716162 TI - Insufficiency subchondral fracture of the femoral head. AB - The authors recently encountered a 65-year-old osteoporotic woman who had had intractable pain in the hip joint that was diagnosed clinically as osteonecrosis. She was treated by total hip replacement. Histopathologically, the most striking finding was the presence of a subchondral fracture with associated callus formation and granulation tissue along both sides of the fracture line. There was no evidence of antecedent osteonecrosis. This case was diagnosed histopathologically as insufficiency subchondral fracture of the femoral head. This is the first case report to substantiate the presence of insufficiency subchondral fracture of the femoral head by both gross and microscopic examination. Because the treatment and management of insufficiency subchondral fracture are entirely different from osteonecrosis, it is important to differentiate between these two conditions. PMID- 10716163 TI - Dedifferentiation in salivary gland carcinomas. PMID- 10716164 TI - Review of extensive workups of 34 patients overexposed to radiofrequency radiation. AB - BACKGROUND: The medical records of 34 patients seen at the Aerospace Medicine Directorate, U.S. Air Force Research Laboratory for confirmed exposure to radiofrequency radiation (RFR) exceeding the permitted exposure limits were reviewed to see if RFR overexposure created any detectable clinical or laboratory alterations that could be correlated with power density or the product of power density and time exposed. The goal of this study was to determine which physiological and laboratory parameters required closest attention on work up of future patients with RFR exposure. METHODS: All 34 patients received an extensive history and physical examination, and a large battery of laboratory studies. Clinical findings were also compared with laboratory results. RESULTS: A sensation of warmth was positively associated with power density. A negative correlation was observed between an abnormal tissue destruction screen and power density. Sophisticated neurological tests in 23 patients and extensive psychometric and psychological exams in 30 patients revealed no neurological or ophthalmologic findings attributable to RFR. A few patients reported burning pain that resolved over several weeks; neurological findings were minimal or absent. CONCLUSIONS: Patients with suspected RFR overexposures need to be seen promptly at the nearest medical facility. Based on this study, an extensive evaluation of persons overexposed to non-ionizing radiation should not be routinely performed. However, a careful history and physical examination with laboratory studies as indicated should be performed and the patient's concerns about RFR effects addressed fully. PMID- 10716165 TI - Cancer incidence and mortality among flight personnel: a meta-analysis. AB - BACKGROUND: Increased cancer risk among flight personnel have previously been noted, including breast cancer among flight attendants and acute myeloid leukemia among pilots. HYPOTHESIS: Exposure to cosmic radiation and other physical or chemical agents may pose health risks for flight personnel. METHODS: We performed an exhaustive search for published and unpublished cohort studies of flight personnel from 1986-98. We combined relative risks (RR) for selected causes from four mortality and/or incidence studies of pilots and two incidence studies of flight attendants, using standard meta-analytic methods. Heterogeneity among the combined studies was explored and adjustments were made for possible confounding by socioeconomic status (SES), where indicated, using correction factors from published studies. RESULTS: SES-adjusted combined RRs were elevated (>1.2) among male pilots for mortality from melanoma 11.97 (95%, CI: 1.02-3.82)] and brain cancer [1.49 (0.89-2.20)], and for cancer incidence of the prostate [1.65 (1.19 2.29)] and the brain [1.74 (0.87-3.30)]. Among female flight attendants, increases were seen for incidence of all cancers [1.29 (0.98-1.70)], melanoma [11.54 (0.83-2.87)], and breast cancer [1.35 (1.00-1.83)]. CONCLUSIONS: Flight personnel appear to be at increased risk for several types of cancer. Both occupational exposures and well-established non-occupational risk factors may contribute to this increased risk. To better control for confounding factors and to identify exposures potentially amenable to preventive measures, future studies should compare risks within cohorts by flight routes, work history, and exposure to cosmic and UV radiation, electromagnetic fields, and chemical substances. PMID- 10716166 TI - Time-course of sleep inertia upon awakening from nighttime sleep with different sleep homeostasis conditions. AB - BACKGROUND: We assessed the time-course of sleep inertia during the first 75 min after morning awakening from regular nocturnal sleep, as well as from nighttime sleep episodes with altered sleep homeostasis conditions. METHODS: Ten normal males slept for 6 nights in the laboratory: 1 adaptation (AD), 2 baseline (BSL, BSL-A), 2 selective Slow-Wave Sleep (SWS) deprivation (DEP-1, DEP-2), and 1 recovery night (REC). On morning awakening, performance was assessed by means of: a) Descending Subtraction Task (DST); b) Auditory Reaction Time task (ART); and c) Finger Tapping Task (FTT). The test battery, lasting about 13 min, was repeated for 5 times. RESULTS: In regard to DST, the Correct Response ratio (CR/NR) showed a great increase of sleep inertia on the first testing session of REC. Regarding sleep inertia time-course, a significant linear decrease across the testing sessions during the BSL-A and the DEP-2 was present, whereas a significant quadratic trend during the AD, the DEP-1 and the REC was found. On the other hand, ART performance showed a significant quadratic trend across testing sessions, while FTT performance did not show any significant variation. CONCLUSIONS: A uniform pattern of variation of time-course of sleep inertia as a function of the different sleep homeostasis conditions was not recognized. Performance accuracy (CR/NR) on the DST showed the hypothesized increasing linear trend across testing sessions only during 2 out of 6 nights, while the unexpected quadratic trend of ART performance is probably due to a fatigue effect. During sleep inertia, cognitive performance reached the baseline level about 30 min after awakening, while motor performance was still below the baseline levels 75 min after awakening. The finding that cognitive performance recovery is greater and more rapid than motor performance recovery could be very important for operational settings and in sustained operations. PMID- 10716167 TI - Dermatologic disease: twenty-two year experience at the USAF Aeromedical Consultation Service and review of other military and civilian experiences. AB - The Aeromedical Consultation Service (ACS) is a U.S. Air Force tertiary referral service which evaluates aviators with complex medical problems and makes recommendations for their aeromedical disposition. This study reports the experience with dermatologic disease in aviators referred to the ACS over a 22-yr period from 1975-1997. A review of other military and civilian experiences with dermatologic disease is also presented. The potential impact of skin disease on aviators and support personnel, in peacetime and wartime, can be surmised by this collective account of dermatologic experiences. PMID- 10716168 TI - The use of melatonin: an information paper. AB - The use of melatonin has been a topic of debate for the past several years. Patients frequently ask their physicians about its use, and many physicians are at a loss about what to tell them. Aviators who have trouble sleeping may choose to buy melatonin and use it since it is a "natural" substance. However, they may lack proper education about its use and the issues of concern. Flight surgeons can help educate their patients in the use of melatonin. This paper will briefly discuss the role of melatonin in humans, its effects on circadian rhythms, its sleep-inducing properties, its effects on mood and performance, and issues pertaining to safety. Flight surgeons and other physicians cannot "prescribe" melatonin, but they at least can offer information about its effects and what is not known about melatonin at this time to the aviators who may ask questions concerning this product. PMID- 10716169 TI - Aeromedical decision-making for aviators with malignant melanoma: an update and review. AB - A review of the recent medical literature on malignant melanoma yielded sufficient information to correlate readily identifiable tumor characteristics to disease-free intervals, and in turn to subsequent risk of cerebral metastasis and neurologic incapacitation. These data were used to construct a new decision table to help guide flight surgeons considering a flying waiver for aviators with a history of melanoma. PMID- 10716170 TI - Early menopause presenting with mood symptoms in a student aviator. AB - The clinical presentation of menopause can resemble the symptoms of a mood disorder. We describe the case of a 31-yr-old student helicopter pilot who presented to the Aviation Psychiatry Department with a several-month history of inconsistent training performance, mood lability, tearfulness, anxiety, insomnia, fatigue, and decreased concentration. Symptoms persisted despite stress management training and resolution of family stressors, and further evaluation revealed other symptoms consistent with early menopause. Symptoms responded to estrogen/progesterone therapy, and patient returned to flight training. The clinical presentation, differential diagnosis, treatment, and aeromedical disposition of perimenopause and menopause are discussed. PMID- 10716171 TI - The effect of baroreflex adaptation on the dynamic cardiovascular response to head-up tilt. AB - BACKGROUND: Baroreflex adaptation to repetitive +Gz has been reported previously. The underlying mechanism may involve different responses of stroke volume (SV) and total peripheral resistance (TPR) to +Gz. HYPOTHESIS: The previously observed enhanced mean arterial pressure (MAP) regulation in fighter pilots (FP) is mediated by increases in SV and/or TPR. METHODS: There were 8 pilots and 12 non pilots who underwent head-up tilt. SV was determined using impedance cardiography. RESULTS: MAP increased significantly in FP, due to heart rate (HR) and TPR increasing more and SV decreasing less. CONCLUSION: Baroreflex adaptation results in better performance of HR, SV and TPR in response to +Gz. PMID- 10716172 TI - Protecting the health of U.S. military forces: a national obligation. PMID- 10716173 TI - A novel method of affinity-purifying proteins using a bis-arsenical fluorescein. AB - Genetically-encoded affinity tags constitute an important strategy for purifying proteins. Here, we have designed a novel affinity matrix based on the his arsenical fluorescein dye FlAsH, which specifically recognizes short alpha helical peptides containing the sequence CCXXCC (Griffin BA, Adams SR, Tsien RY, 1998, Science 281:269-272). We find that kinesin tagged with this cysteine containing helix binds specifically to FlAsH resin and can be eluted in a fully active form. This affinity tag has several advantages over polyhistidine, the only small affinity tag in common use. The protein obtained with this single chromatographic step from crude Escherichia coli lysates is purer than that obtained with nickel affinity chromatography of 6xHis tagged kinesin. Moreover, unlike nickel affinity chromatography, which requires high concentrations of imidazole or pH changes for elution, protein bound to the FlAsH column can be completely eluted by dithiothreitol. Because of these mild elution conditions, FlAsH affinity chromatography is ideal for recovering fully active protein and for the purification of intact protein complexes. PMID- 10716174 TI - 2.9 A crystal structure of ligand-free tryptophanyl-tRNA synthetase: domain movements fragment the adenine nucleotide binding site. AB - The crystal structure of ligand-free tryptophanyl-tRNA synthetase (TrpRS) was solved at 2.9 A using a combination of molecular replacement and maximum-entropy map/phase improvement. The dimeric structure (R = 23.7, Rfree = 26.2) is asymmetric, unlike that of the TrpRS tryptophanyl-5'AMP complex (TAM; Doublie S, Bricogne G, Gilmore CJ, Carter CW Jr, 1995, Structure 3:17-31). In agreement with small-angle solution X-ray scattering experiments, unliganded TrpRS has a conformation in which both monomers open, leaving only the tryptophan-binding regions of their active sites intact. The amino terminal alphaA-helix, TIGN, and KMSKS signature sequences, and the distal helical domain rotate as a single rigid body away from the dinucleotide-binding fold domain, opening the AMP binding site, seen in the TAM complex, into two halves. Comparison of side-chain packing in ligand-free TrpRS and the TAM complex, using identification of nonpolar nuclei (Ilyin VA, 1994, Protein Eng 7:1189-1195), shows that significant repacking occurs between three relatively stable core regions, one of which acts as a bearing between the other two. These domain rearrangements provide a new structural paradigm that is consistent in detail with the "induced-fit" mechanism proposed for TyrRS by Fersht et al. (Fersht AR, Knill-Jones JW, Beduelle H, Winter G, 1988, Biochemistry 27:1581-1587). Coupling of ATP binding determinants associated with the two catalytic signature sequences to the helical domain containing the presumptive anticodon-binding site provides a mechanism to coordinate active-site chemistry with relocation of the major tRNA binding determinants. PMID- 10716175 TI - Comparison of sequence profiles. Strategies for structural predictions using sequence information. AB - Distant homologies between proteins are often discovered only after three dimensional structures of both proteins are solved. The sequence divergence for such proteins can be so large that simple comparison of their sequences fails to identify any similarity. New generation of sensitive alignment tools use averaged sequences of entire homologous families (profiles) to detect such homologies. Several algorithms, including the newest generation of BLAST algorithms and BASIC, an algorithm used in our group to assign fold predictions for proteins from several genomes, are compared to each other on the large set of structurally similar proteins with little sequence similarity. Proteins in the benchmark are classified according to the level of their similarity, which allows us to demonstrate that most of the improvement of the new algorithms is achieved for proteins with strong functional similarities, with almost no progress in recognizing distant fold similarities. It is also shown that details of profile calculation strongly influence its sensitivity in recognizing distant homologies. The most important choice is how to include information from diverging members of the family, avoiding generating false predictions, while accounting for entire sequence divergence within a family. PSI-BLAST takes a conservative approach, deriving a profile from core members of the family, providing a solid improvement without almost any false predictions. BASIC strives for better sensitivity by increasing the weight of divergent family members and paying the price in lower reliability. A new FFAS algorithm introduced here uses a new procedure for profile generation that takes into account all the relations within the family and matches BASIC sensitivity with PSI-BLAST like reliability. PMID- 10716176 TI - Characterization of the functional role of Asp141, Asp194, and Asp464 residues in the Mn2+-L-malate binding of pigeon liver malic enzyme. AB - Pigeon liver malic enzyme was inactivated and cleaved at Asp141, Asp194, and Asp464 by the Cu2+-ascorbate system in acidic environment. Site-specific mutagenesis was performed at these putative metal-binding sites. Three point mutants, D141N, D194N, and D464N; three double mutants, D(141,194)N, D(194,464)N, and D(141,464)N; and a triple mutant, D(141,194,464)N; as well as the wild-type malic enzyme (WT) were successfully cloned and expressed in Escherichia coli cells. All recombinant enzymes, except the triple mutant, were purified to apparent homogeneity by successive Q-Sepharose and adenosine-2',5'-bisphosphate agarose columns. The mutants showed similar apparent Km,NADP values to that of the WT. The Km,Mal value was increased in the D141N and D194N mutants. The Km,Mn value, on the other hand, was increased only in the D141N mutant by 14-fold, corresponding to approximately 1.6 kcal/mol for the Asp141-Mn2+ binding energy. Substrate inhibition by L-malate was only observed in WT, D464N, and D(141,464)N. Initial velocity experiments were performed to derive the various kinetic parameters. The possible interactions between Asp141, Asp194, and Asp464 were analyzed by the double-mutation cycles and triple-mutation box. There are synergistic weakening interactions between Asp141 and Asp194 in the metal binding that impel the D(141,194)N double mutant to an overall specificity constant [k(cat)/(Kd,Mn Km,Mal Km,NADP)] at least four orders of magnitude smaller than the WT value. This difference corresponds to an increase of 6.38 kcal/mol energy barrier for the catalytic efficiency. Mutation at Asp464, on the other hand, has partial additivity on the mutations at Asp141 and Asp194. The overall specificity constants for the double mutants D(194,464)N and D(141,464)N or the triple mutant D(141,194,464)N were decreased by only 10- to 100-fold compared to the WT. These results strongly suggest the involvement of Asp141 in the Mn2+-L-malate binding for the pigeon liver malic enzyme. The Asp194 and Asp464, which may be oxidized by nonspecific binding of Cu2+, are involved in the Mn2+-L-malate binding or catalysis indirectly by modulating the binding affinity of Asp141 with the Mn2+. PMID- 10716178 TI - Structure of tick anticoagulant peptide at 1.6 A resolution complexed with bovine pancreatic trypsin inhibitor. AB - The structure of tick anticoagulant peptide (TAP) has been determined by X-ray crystallography at 1.6 A resolution complexed with bovine pancreatic trypsin inhibitor (BPTI). The TAP-BPTI crystals are tetragonal, a = b = 46.87, c = 50.35 A, space group P41, four complexes per unit cell. The TAP molecules are highly dipolar and form an intermolecular helical array along the c-axis with a diameter of about 45 A. Individual TAP units interact in a head-to-tail fashion, the positive end of one molecule associating with the distal negative end of another, and vice versa. The BPTI molecules have a uniformly distributed positively charged surface that interacts extensively through 14 hydrogen bonds and two hydrogen bonded salt bridges with the helical groove around the helical TAP chains. Comparing the structure of TAP in TAP-BPTI with TAP bound to factor Xa(Xa) suggests a massive reorganization in the N-terminal tetrapeptide and the first disulfide loop of TAP (Cys5T-Cys15T) upon binding to Xa. The Tyr1(T)OH atom of TAP moves 14.2 A to interact with Asp189 of the S1 specificity site, Arg3(T)CZ moves 5.0 A with the guanidinium group forming a cation-pi-electron complex in the S4 subsite of Xa, while Lys7(T)NZ differs in position by 10.6 A in TAP-BPTI and TAP-Xa, all of which indicates a different pre-Xa-bound conformation for the N-terminal of TAP in its native state. In contrast to TAP, the BPTI structure of TAP-BPTI is practically the same as all those of previously determined structures of BPTI, only arginine and lysine side-chain conformations showing significant differences. PMID- 10716177 TI - Structure-based thermodynamic analysis of the dissociation of protein phosphatase 1 catalytic subunit and microcystin-LR docked complexes. AB - The relationship between the structure of a free ligand in solution and the structure of its bound form in a complex is of great importance to the understanding of the energetics and mechanism of molecular recognition and complex formation. In this study, we use a structure-based thermodynamic approach to study the dissociation of the complex between the toxin microcystin-LR (MLR) and the catalytic domain of protein phosphatase-1 (PP-1c) for which the crystal structure of the complex is known. We have calculated the thermodynamic parameters (enthalpy, entropy, heat capacity, and free energy) for the dissociation of the complex from its X-ray structure and found the calculated dissociation constant (4.0 x 10(-11)) to be in excellent agreement with the reported inhibitory constant (3.9 x 10(-11)). We have also calculated the thermodynamic parameters for the dissociation of 47 PP-1c:MLR complexes generated by docking an ensemble of NMR solution structures of MLR onto the crystal structure of PP-1c. In general, we observe that the lower the root-mean-square deviation (RMSD) of the docked complex (compared to the X-ray complex) the closer its free energy of dissociation (deltaGd(o)) is to that calculated from the X-ray complex. On the other hand, we note a significant scatter between the deltaGd(o) and the RMSD of the docked complexes. We have identified a group of seven docked complexes with deltaGd(o) values very close to the one calculated from the X-ray complex but with significantly dissimilar structures. The analysis of the corresponding enthalpy and entropy of dissociation shows a compensation effect suggesting that MLR molecules with significant structural variability can bind PP 1c and that substantial conformational flexibility in the PP-1c:MLR complex may exist in solution. PMID- 10716179 TI - Conservative mutation Met8 --> Leu affects the folding process and structural stability of squash trypsin inhibitor CMTI-I. AB - Protein molecules can accommodate a large number of mutations without noticeable effects on their stability and folding kinetics. On the other hand, some mutations can have quite strong effects on protein conformational properties. Such mutations either destabilize secondary structures, e.g., alpha-helices, are incompatible with close packing of protein hydrophobic cores, or lead to disruption of some specific interactions such as disulfide cross links, salt bridges, hydrogen bonds, or aromatic-aromatic contacts. The Met8 --> Leu mutation in CMTI-I results in significant destabilization of the protein structure. This effect could hardly be expected since the mutation is highly conservative, and the side chain of residue 8 is situated on the protein surface. We show that the protein destabilization is caused by rearrangement of a hydrophobic cluster formed by side chains of residues 8, Ile6, and Leu17 that leads to partial breaking of a hydrogen bond formed by the amide group of Leu17 with water and to a reduction of a hydrophobic surface buried within the cluster. The mutation perturbs also the protein folding. In aerobic conditions the reduced wild-type protein folds effectively into its native structure, whereas more then 75% of the mutant molecules are trapped in various misfolded species. The main conclusion of this work is that conservative mutations of hydrophobic residues can destabilize a protein structure even if these residues are situated on the protein surface and partially accessible to water. Structural rearrangement of small hydrophobic clusters formed by such residues can lead to local changes in protein hydration, and consequently, can affect considerably protein stability and folding process. PMID- 10716180 TI - An interdomain distance in cardiac troponin C determined by fluorescence spectroscopy. AB - The distance between Ca2+-binding site III in the C-terminal domain and Cys35 in the N-terminal domain in cardiac muscle troponin C (cTnC) was determined with a single-tryptophan mutant using bound Tb3+ as the energy donor and iodoacetamidotetramethylrhodamine linked to the cysteine residue as energy acceptor. The luminescence of bound Tb3+ was generated through sensitization by the tryptophan located in the 12-residue binding loop of site III upon irradiation at 295 nm, and this sensitized luminescence was the donor signal transferred to the acceptor. In the absence of bound cation at site II, the mean interdomain distance was found to be 48-49 A regardless of whether the cTnC was unbound or bound to cardiac troponin I, or reconstituted into cardiac troponin. These results suggest that cTnC retains its overall length in the presence of bound target proteins. The distribution of the distances was wide (half-width >9 A) and suggests considerable interdomain flexibility in isolated cTnC, but the distributions became narrower for cTnC in the complexes with the other subunits. In the presence of bound cation at the regulatory site II, the interdomain distance was shortened by 6 A for cTnC, but without an effect on the half-width. The decrease in the mean distance was much smaller or negligible when cTnC was complexed with cTnI or cTnI and cTnT under the same conditions. Although free cTnC has considerable interdomain flexibility, this dynamics is slightly reduced in troponin. These results indicate that the transition from the relaxed state to an activated state in cardiac muscle is not accompanied by a gross alteration of the cTnC conformation in cardiac troponin. PMID- 10716181 TI - Microscopic stability of cold shock protein A examined by NMR native state hydrogen exchange as a function of urea and trimethylamine N-oxide. AB - Native state hydrogen exchange of cold shock protein A (CspA) has been characterized as a function of the denaturant urea and of the stabilizing agent trimethylamine N-oxide (TMAO). The structure of CspA has five strands of beta sheet. Strands beta1-beta4 have strongly protected amide protons that, based on experiments as a function of urea, exchange through a simple all-or-none global unfolding mechanism. By contrast, the protection of amide protons from strand beta5 is too weak to measure in water. Strand beta5 is hydrogen bonded to strands beta3 and beta4, both of which afford strong protection from solvent exchange. Gaussian network model (GNM) simulations, which assume that the degree of protection depends on tertiary contact density in the native structure, accurately predict the strong protection observed in strands beta1-beta4 but fail to account for the weak protection in strand beta5. The most conspicuous feature of strand beta5 is its low sequence hydrophobicity. In the presence of TMAO, there is an increase in the protection of strands beta1-beta4, and protection extends to amide protons in more hydrophilic segments of the protein, including strand beta5 and the loops connecting the beta-strands. TMAO stabilizes proteins by raising the free energy of the denatured state, due to highly unfavorable interactions between TMAO and the exposed peptide backbone. As such, the stabilizing effects of TMAO are expected to be relatively independent of sequence hydrophobicity. The present results suggest that the magnitude of solvent exchange protection depends more on solvent accessibility in the ensemble of exchange susceptible conformations than on the strength of hydrogen-bonding interactions in the native structure. PMID- 10716182 TI - Protein global fold determination using site-directed spin and isotope labeling. AB - We describe a simple experimental approach for the rapid determination of protein global folds. This strategy utilizes site-directed spin labeling (SDSL) in combination with isotope enrichment to determine long-range distance restraints between amide protons and the unpaired electron of a nitroxide spin label using the paramagnetic effect on relaxation rates. The precision and accuracy of calculating a protein global fold from only paramagnetic effects have been demonstrated on barnase, a well-characterized protein. Two monocysteine derivatives of barnase, (H102C) and (H102A/Q15C), were 15N enriched, and the paramagnetic nitroxide spin label, MTSSL, attached to the single Cys residue of each. Measurement of amide 1H longitudinal relaxation times, in both the oxidized and reduced states, allowed the determination of the paramagnetic contribution to the relaxation processes. Correlation times were obtained from the frequency dependence of these relaxation processes at 800, 600, and 500 MHz. Distances in the range of 8 to 35 A were calculated from the magnitude of the paramagnetic contribution to the relaxation processes and individual amide 1H correlation times. Distance restraints from the nitroxide spin to amide protons were used as restraints in structure calculations. Using nitroxide to amide 1H distances as long-range restraints and known secondary structure restraints, barnase global folds were calculated having backbone RMSDs <3 A from the crystal structure. This approach makes it possible to rapidly obtain the overall topology of a protein using a limited number of paramagnetic distance restraints. PMID- 10716183 TI - Characterization of a comparative model of the extracellular domain of the epidermal growth factor receptor. AB - The Epidermal Growth Factor (EGF) receptor is a tyrosine kinase that mediates the biological effects of ligands such as EGF and transforming growth factor alpha. An understanding of the molecular basis of its action has been hindered by a lack of structural and mutational data on the receptor. We have constructed comparative models of the four extracellular domains of the EGF receptor that are based on the structure of the first three domains of the insulin-like growth factor-1 (IGF-1) receptor. The first and third domains of the EGF receptor, L1 and L2, are right-handed beta helices. The second and fourth domains of the EGF receptor, S1 and S2, consist of the modules held together by disulfide bonds, which, except for the first module of the S1 domain, form rod-like structures. The arrangement of the L1 and S1 domains of the model are similar to that of the first two domains of the IGF-1 receptor, whereas that of the L2 and S2 domains appear to be significantly different. Using the EGF receptor model and limited information from the literature, we have proposed a number of regions that may be involved in the functioning of the receptor. In particular, the faces containing the large beta sheets in the L1 and L2 domains have been suggested to be involved with ligand binding of EGF to its receptor. PMID- 10716184 TI - Resolution of ligand positions by site-directed tryptophan fluorescence in tear lipocalin. AB - The lipocalin superfamily of proteins functions in the binding and transport of a variety of important hydrophobic molecules. Tear lipocalin is a promiscuous lipid binding member of the family and serves as a paradigm to study the molecular determinants of ligand binding. Conserved regions in the lipocalins, such as the G strand and the F-G loop, may play an important role in ligand binding and delivery. We studied structural changes in the G strand of holo- and apo-tear lipocalin using spectroscopic methods including circular dichroism analysis and site-directed tryptophan fluorescence. Apo-tear lipocalin shows the same general structural characteristics as holo-tear lipocalin including alternating periodicity of a beta-strand, orientation of amino acid residues 105, 103, 101, and 99 facing the cavity, and progressive depth in the cavity from residues 105 to 99. For amino acid residues facing the internal aspect of cavity, the presence of a ligand is associated with blue shifted spectra. The collisional rate constants indicate that these residues are not less exposed to solvent in holo tear lipocalin than in apo-tear lipocalin. Rather the spectral blue shifts may be accounted for by a ligand induced rigidity in holo-TL. Amino acid residues 94 and 95 are consistent with positions in the F-G loop and show greater exposure to solvent in the holo- than the apo-proteins. These findings are consistent with the general hypothesis that the F-G loop in the holo-proteins of the lipocalin family is available for receptor interactions and delivery of ligands to specific targets. Site-directed tryptophan fluorescence was used in combination with a nitroxide spin labeled fatty acid analog to elucidate dynamic ligand interactions with specific amino acid residues. Collisional quenching constants of the nitroxide spin label provide evidence that at least three amino acids of the G strand residues interact with the ligand. Stern-Volmer plots are inconsistent with a ligand that is held in a static position in the calyx, but rather suggest that the ligand is in motion. The combination of site-directed tryptophan fluorescence with quenching by nitroxide labeled species has broad applicability in probing specific interactions in the solution structure of proteins and provides dynamic information that is not attainable by X-ray crystallography. PMID- 10716185 TI - Copper binding to octarepeat peptides of the prion protein monitored by mass spectrometry. AB - Electrospray ionization mass spectrometry (ESI-MS) was used to measure the binding of Cu2+ ions to synthetic peptides corresponding to sections of the sequence of the mature prion protein (PrP). ESI-MS demonstrates that Cu2+ is unique among divalent metal ions in binding to PrP and defines the location of the major Cu2+ binding site as the octarepeat region in the N-terminal domain, containing multiple copies of the repeat ProHisGlyGlyGlyTrpGlyGln. The stoichiometries of the complexes measured directly by ESI-MS are pH dependent: a peptide containing four octarepeats chelates two Cu2+ ions at pH 6 but four at pH 7.4. At the higher pH, the binding of multiple Cu2+ ions occurs with a high degree of cooperativity for peptides C-terminally extended to incorporate a fifth histidine. Dissociation constants for each Cu2+ ion binding to the octarepeat peptides, reported here for the first time, are mostly in the low micromolar range; for the addition of the third and fourth Cu2+ ions to the extended peptides at pH 7.4, K(D)'s are <100 nM. N-terminal acetylation of the peptides caused some reduction in the stoichiometry of binding at both pH's. Cu2+ also binds to a peptide corresponding to the extreme N-terminus of PrP that precedes the octarepeats, arguing that this region of the sequence may also make a contribution to the Cu2+ complexation. Although the structure of the four octarepeat peptide is not affected by pH changes in the absence of Cu2+, as judged by circular dichroism, Cu2+ binding induces a modest change at pH 6 and a major structural perturbation at pH 7.4. It is possible that PrP functions as a Cu2+ transporter by binding Cu2+ ions from the extracellular medium under physiologic conditions and then releasing some or all of this metal upon exposure to acidic pH in endosomes or secondary lysosomes. PMID- 10716186 TI - Proline inhibits aggregation during protein refolding. AB - The in vitro refolding of hen egg-white lysozyme is studied in the presence of various osmolytes. Proline is found to prevent aggregation during protein refolding. However, other osmolytes used in this study fail to exhibit a similar property. Experimental evidence suggests that proline inhibits protein aggregation by binding to folding intermediate(s) and trapping the folding intermediate(s) into enzymatically inactive, "aggregation-insensitive" state(s). However, elimination of proline from the refolded protein mixture results in significant recovery of the bacteriolytic activity. At higher concentrations (>1.5 M), proline is shown to form loose, higher-order molecular aggregate(s). The supramolecular assembly of proline is found to possess an amphipathic character. Formation of higher-order aggregates is believed to be crucial for proline to function as a protein folding aid. In addition to its role in osmoregulation under water stress conditions, the results of this study hint at the possibility of proline behaving as a protein folding chaperone. PMID- 10716187 TI - Substrate- and pH-dependent contribution of oxyanion binding site to the catalysis of prolyl oligopeptidase, a paradigm of the serine oligopeptidase family. AB - Prolyl oligopeptidase, an enzyme implicated in memory disorders, is a member of a new serine peptidase family. Crystallographic studies (Fulop et al., 1998) revealed a novel oxyanion binding site containing a tyrosine residue, Tyr473. To study the importance of Tyr473 OH, we have produced prolyl oligopeptidase and its Tyr473Phe variant in Escherichia coli. The specificity rate constant, k(cat)/Km, for the modified enzyme decreased by a factor of 8-40 with highly specific substrates, Z-Gly-Pro-Nap, and a fluorogenic octapeptide. With these compounds, the decline in k(cat) was partly compensated for by reduction in Km, a difference from the extensively studied subtilisin. With the less specific suc-Gly-Pro-Nap, the Km value, which approximates Ks, was not significantly changed, resulting in greater diminution (approximately 500-fold) in k(cat)/Km. The second-order rate constant for the reaction with Z-Pro-prolinal, a slow tight-binding transition state analogue inhibitor, and the Ki values for a slow substrate and two product like inhibitors were not significantly affected by the Tyr473 OH group. The mechanism of transition-state stabilization was markedly dependent upon the nature of substrate and varied with pH as the enzyme interconverted between its two catalytically competent forms. PMID- 10716188 TI - Role of the switch II region in the conformational transition of activation of Ha ras-p21. AB - The role of the switch II region in the conformational transition of activation of Ha-ras-p21 has been investigated by mutating residues predicted to act as hinges for the conformational transition of this loop (Ala59, Gly60, and Gly75) (Diaz JF, Wroblowski B, Schlitter J, Engelborghs Y, 1997, Proteins 28:434-451), as well as mutating the catalytic residue Gln61. The proposed mutations of the hinge residues decrease the rate of the conformational transition of activation as measured by the binding of BeF3- to the GDP-p21 complex. Also, the thermodynamic parameters of the binding reaction are altered by a factor between three and five, depending on the temperature. (Due to changes in activation and reaction enthalpies, partially compensated by entropy changes.) The control mutation Q61H in which only the catalytic residue is changed has only a limited effect on the kinetic rate constants of the conformational transition and on the thermodynamic parameters of the reaction. The fact that mutations of the hinge residues of the switch II region affect both the binding of the phosphate analog and the conformational transition of activation indicates that the switch II is implicated both in the early and the late states of the transition. PMID- 10716189 TI - Amyloid protofilament formation of hen egg lysozyme in highly concentrated ethanol solution. AB - Mutant human lysozymes (Ile56Thr & Asp67His) have been reported to form amyloid deposits in the viscera. From the standpoint of understanding the mechanism of amyloid formation, we searched for conditions of amyloid formation in vitro using hen egg lysozyme, which has been extensively studied from a physicochemical standpoint. It was found that the circular dichroism spectra in the far ultraviolet region of the hen egg lysozyme changed to those characteristic of a beta-structure from the native alpha-helix rich spectrum in 90% ethanol solution. When the concentration of protein was increased to 10 mg/mL, the protein solution formed a gel in the presence of 90% ethanol, and precipitated on further addition of 10 mM NaCl. The precipitates were examined by electron microscopy, their ability to bind Congo red, and X-ray diffraction to determine whether amyloid fibrils were formed in the precipitates. Electron micrographs displayed unbranched protofilament with a diameter of approximately 70 A. The peak point of the difference spectrum for the Congo red binding assay was 541 nm, which is characteristic of amyloid fibrils. The X-ray diffraction pattern showed a sharp and intense diffraction ring at 4.7 A, a reflection that arises from the interstrand spacing in beta-sheets. These results indicate that the precipitates of hen egg lysozyme are amyloid protofilament, and that the amyloid protofilament formation of hen egg lysozyme closely follows upon the destruction of the helical and tertiary structures. PMID- 10716190 TI - Human RhoA/RhoGDI complex expressed in yeast: GTP exchange is sufficient for translocation of RhoA to liposomes. AB - The human small GTPase, RhoA, expressed in Saccharomyces cerevisiae is post translationally processed and, when co-expressed with its cytosolic inhibitory protein, RhoGDI, spontaneously forms a heterodimer in vivo. The RhoA/RhoGDI complex, purified to greater than 98% at high yield from the yeast cytosolic fraction, could be stoichiometrically ADP-ribosylated by Clostridium botulinum C3 exoenzyme, contained stoichiometric GDP, and could be nucleotide exchanged fully with [3H]GDP or partially with GTP in the presence of submicromolar Mg2+. The GTP RhoA/RhoGDI complex hydrolyzed GTP with a rate constant of 4.5 X 10(-5) s(-1), considerably slower than free RhoA. Hydrolysis followed pseudo-first-order kinetics indicating that the RhoA hydrolyzing GTP was RhoGDI associated. The constitutively active G14V-RhoA mutant expressed as a complex with RhoGDI and purified without added nucleotide also bound stoichiometric guanine nucleotide: 95% contained GDP and 5% GTP. Microinjection of the GTP-bound G14V-RhoA/RhoGDI complex (but not the GDP form) into serum-starved Swiss 3T3 cells elicited formation of stress fibers and focal adhesions. In vitro, GTP-bound-RhoA spontaneously translocated from its complex with RhoGDI to liposomes, whereas GDP RhoA did not. These results show that GTP-triggered translocation of RhoA from RhoGDI to a membrane, where it carries out its signaling function, is an intrinsic property of the RhoA/RhoGDI complex that does not require other protein factors or membrane receptors. PMID- 10716191 TI - Contribution of proton linkage to the thermodynamic stability of the major cold shock protein of Escherichia coli CspA. AB - The stability of protein is defined not only by the hydrogen bonding, hydrophobic effect, van der Waals interactions, and salt bridges. Additional, much more subtle contributions to protein stability can arise from surface residues that change their properties upon unfolding. The recombinant major cold shock protein of Escherichia coli CspA an all-beta protein unfolds reversible in a two-state manner, and behaves in all other respects as typical globular protein. However, the enthalpy of CspA unfolding strongly depends on the pH and buffer composition. Detailed analysis of the unfolding enthalpies as a function of pH and buffers with different heats of ionization shows that CspA unfolding in the pH range 5.5 9.0 is linked to protonation of an amino group. This amino group appears to be the N-terminal alpha-amino group of the CspA molecule. It undergoes a 1.6 U shift in pKa values between native and unfolded states. Although this shift in pKa is expected to contribute approximately 5 kJ/mol to CspA stabilization energy the experimentally observed stabilization is only approximately 1 kJ/mol. This discrepancy is related to a strong enthalpy-entropy compensation due, most likely, to the differences in hydration of the protonated and deprotonated forms of the alpha-amino group. PMID- 10716192 TI - Electrospray ionization mass spectrometry of zinc, cadmium, and copper metallothioneins: evidence for metal-binding cooperativity. AB - Electrospray ionization (ESI) mass spectra of both well-characterized and novel metallothioneins (MTs) from various species were recorded to explore their metal ion-binding modes and stoichiometries. The ESI mass spectra of the zinc- and cadmium-binding MTs showed a single main peak corresponding to metal-to-protein ratios of 4, 6, or 7. These findings combined with data obtained by other methods suggest that these MTs bind zinc or cadmium in a single predominant form and are consistent with the presence of three- and four-metal clusters. An unstable copper-specific MT isoform from Roman snails (Helix pomatia) could be isolated intact and was shown to preferentially bind 12 copper ions. To obtain additional information on the formation and relative stability of metal-thiolate clusters in MTs, a mass spectrometric titration study was conducted. One to seven molar equivalents of zinc or of cadmium were added to metal-free human MT-2 at neutral pH, and the resulting complexes were measured by ESI mass spectrometry. These experiments revealed that the formation of the four-metal cluster and of the thermodynamically less stable three-metal cluster is sequential and largely cooperative for both zinc and cadmium. Minor intermediate forms between metal free MT, Me4MT, and fully reconstituted Me7MT were also observed. The addition of increasing amounts of cadmium to metal-free blue crab MT-I resulted in prominent peaks whose masses were consistent with apoMT, Cd3MT, and Cd6MT, reflecting the known structure of this MT with two Me3Cys9 centers. In a similar reconstitution experiment performed with Caenorhabditis elegans MT-II, a series of signals corresponding to apoMT and Cd3MT to Cd6MT species were observed. PMID- 10716194 TI - Cleaved antitrypsin polymers at atomic resolution. AB - Alpha1-antitrypsin deficiency, which can lead to both emphysema and liver disease, is a result of the accumulation of alpha1-antitrypsin polymers within the hepatocyte. A wealth of biochemical and biophysical data suggests that alpha1 antitrypsin polymers form via insertion of residues from the reactive center loop of one molecule into the beta-sheet of another. However, this long-standing hypothesis has not been confirmed by direct structural evidence. Here, we describe the first crystallographic evidence of a beta-strand linked polymer form of alpha1-antitrypsin: the crystal structure of a cleaved alpha1-antitrypsin polymer. PMID- 10716193 TI - A new approach to the design of uniquely folded thermally stable proteins. AB - A new computer program (CORE) is described that predicts core hydrophobic sequences of predetermined target protein structures. A novel scoring function is employed, which for the first time incorporates parameters directly correlated to free energies of unfolding (deltaGu), melting temperatures (Tm), and cooperativity. Metropolis-driven simulated annealing and low-temperature Monte Carlo sampling are used to optimize this score, generating sequences predicted to yield uniquely folded, stable proteins with cooperative unfolding transitions. The hydrophobic core residues of four natural proteins were predicted using CORE with the backbone structure and solvent exposed residues as input. In the two smaller proteins tested (Gbeta1, 11 core amino acids; 434 cro, 10 core amino acids), the native sequence was regenerated as well as the sequence of known thermally stable variants that exhibit cooperative denaturation transitions. Previously designed sequences of variants with lower thermal stability and weaker cooperativity were not predicted. In the two larger proteins tested (myoglobin, 32 core amino acids; methionine aminopeptidase, 63 core amino acids), sequences with corresponding side-chain conformations remarkably similar to that of native were predicted. PMID- 10716195 TI - Solution NMR evidence for a cis Tyr-Ala peptide group in the structure of [Pro93Ala] bovine pancreatic ribonuclease A. AB - Proline peptide group isomerization can result in kinetic barriers in protein folding. In particular, the cis proline peptide conformation at Tyr92-Pro93 of bovine pancreatic ribonuclease A (RNase A) has been proposed to be crucial for chain folding initiation. Mutation of this proline-93 to alanine results in an RNase A molecule, P93A, that exhibits unfolding/refolding kinetics consistent with a cis Tyr92-Ala93 peptide group conformation in the folded structure (Dodge RW, Scheraga HA, 1996, Biochemistry 35:1548-1559). Here, we describe the analysis of backbone proton resonance assignments for P93A together with nuclear Overhauser effect data that provide spectroscopic evidence for a type VI beta bend conformation with a cis Tyr92-Ala93 peptide group in the folded structure. This is in contrast to the reported X-ray crystal structure of [Pro93Gly]-RNase A (Schultz LW, Hargraves SR, Klink TA, Raines RT, 1998, Protein Sci 7:1620-1625), in which Tyr92-Gly93 forms a type-II beta-bend with a trans peptide group conformation. While a glycine residue at position 93 accommodates a type-II bend (with a positive value of phi93), RNase A molecules with either proline or alanine residues at this position appear to require a cis peptide group with a type-VI beta-bend for proper folding. These results support the view that a cis Pro93 conformation is crucial for proper folding of wild-type RNase A. PMID- 10716197 TI - Cell immortalisation--a few words of caution. PMID- 10716196 TI - Direct measurement of islet amyloid polypeptide fibrillogenesis by mass spectrometry. AB - A novel method for monitoring fibrillogenesis is developed and applied to the amyloidogenic peptide, islet amyloid polypeptide (IAPP). The approach, based on electrospray ionization mass spectrometry, is complementary to existing assays of fibril formation as it monitors directly the population of precursor rather than product molecules. We are able to monitor fiber formation in two modes: a quenched mode in which fibril formation is halted by dilution into denaturant and a real time mode in which fibril formation is conducted within the capillary of the electrospray source. Central to the method is the observation that fibrillar IAPP does not compromise the ionization of monomeric IAPP. Furthermore, under mild ionization conditions, fibrillar IAPP does not dissociate and contribute to the monomeric signal. Critically, we introduce an internal standard, rat IAPP, for analysis on the mass spectrometer. This standard is sufficiently similar in sequence in that it ionizes identically to human IAPP. Furthermore, the sequence is sufficiently different in that it does not form fibrils and is distinguishable on the basis of mass. Applied to IAPP fibrillogenesis, our technique reveals that precursor consumption in seeded reactions obeys first-order kinetics. Furthermore, a consistent level of monomer persists in both seeded and unseeded experiments after the fibril formation is complete. Given the inherent stability of fibrils, we expect this approach to be applicable to other amyloid systems. PMID- 10716198 TI - The failure of axon regeneration in the CNS is not absolute. PMID- 10716199 TI - A new model of the blood--brain barrier: co-culture of neuronal, endothelial and glial cells under dynamic conditions. AB - Developing in vitro blood-brain barrier (BBB) models that closely mimic the natural state is important for theoretical and practical applications, including drug development. We previously developed an in vitro BBB model based on co culturing endothelial cells with glia in the presence of flow on hollow fiber tube culture substrates. We now report that this dynamic in vitro BBB (DIV-BBB) can be successfully used to co-culture differentiated serotonergic neurons in the presence of a BBB. These neurons demonstrated fluoxetine-sensitive serotonin (5HT) uptake and depolarization-induced release of [3H]5HT. Our results demonstrate that the DIV-BBB is a suitable model for culturing of neurons in a quasi-physiological microenvironment and in the presence of a high-resistance, stereoselective BBB. PMID- 10716200 TI - Human thalamus: neurochemical mapping of inferior pulvinar complex. AB - The primate pulvinar connects with the entire array of known visual areas and is postulated to play a role in selective visual attention. Recently, five separate neurochemical subdivisions of a region termed the inferior pulvinar (PI) complex were identified in monkeys. In the present study, similar histochemical procedures were applied to map the extent of the PI complex in humans. Acetylcholinesterase histochemistry and cytochrome oxidase staining demarcated four histochemical zones in human pulvinar, corresponding to the medial, central, lateral and lateral-shell (PI(M), PI(C), PI(L), and PI(L-S)) divisions of the PI complex in monkeys. PMID- 10716201 TI - Role of ET-1 in hypoxia-induced mitosis of cultured rat carotid body chemoreceptors. AB - The mammalian carotid body (CB) contains O2-chemoreceptors, i.e. glomus cells, which display increased mitoses and endothelin-1 (ET-1) expression during chronic hypoxia. To investigate whether endogenous ET-1 might mediate these mitogenic effects, we quantified bromodeoxyuridine (BrdU) uptake by tyrosine hydroxylase (TH)-positive glomus cells in rat CB cultures using double-label immunofluorescence. In normoxia (20% O2), 2-day exposure to ET-1 (10-1000 nM) caused a dose-dependent increase in BrdU uptake which peaked (approximately 55% of TH+ cells) at around 500 nM ET-1. In chronic hypoxia (5% O2) alone, BrdU uptake was stimulated (approximately 46% of TH+ cells) relative to normoxia (approximately 30%), but the effect was abolished in the presence of specific (BQ 123) or non-specific (PD 142893) ETA receptor antagonists (10(-5) M). Thus paracrine/autocrine release of ET-1 in the hypoxic carotid body may promote glomus cell mitosis via ET(A) receptors. PMID- 10716202 TI - Sleep deprivation-induced reduction in cortical functional response to serial subtraction. AB - Thirteen normal volunteers were studied with fMRI during arithmetic performance after a normal night of sleep and following sleep deprivation (SD). Aims included determining whether the prefrontal cortex (PFC) and the parietal lobe arithmetic areas are vulnerable to the effects of SD. After a normal night of sleep, activation localized to the bilateral PFC, parietal lobes and premotor areas. Following SD, activity in these regions decreased markedly, especially in the PFC. Performance also dropped. Data from the serial subtraction task are consistent with Horne's PFC vulnerability hypothesis but, based on this and other studies, we suggest the localized, functional effects of SD in the brain may vary, in part, with the specific cognitive task. PMID- 10716203 TI - Attention-dependent allocation of auditory processing resources as measured by mismatch negativity. AB - Mismatch negativity (MMN) is a pre-attentive event-related potential measure of echoic memory. However, recent studies suggest attention-related modulation of MMN. This study investigates duration-elicited MMN in healthy subjects (n = 12) who were performing a visual discrimination task and, subsequently, an auditory discrimination task in a series of increasing task difficulty. MMN amplitude was found to be maximal at centro-frontal electrode sites without hemispheric differences. Comparison of both attend conditions (visual vs. auditory), revealed larger MMN amplitudes at Fz in the visual task without differences across task difficulty. However, significantly smaller MMN in the most demanding auditory condition supports the notion of limited processing capacity whose resources are modulated by attention in response to task requirements. PMID- 10716204 TI - HMBA inhibits growth and induces differentiation of astrocytes from neonatal rat brain. AB - HMBA, a potent differentiation inducer belonging to the class of hybrid polar compounds, inhibited the growth of astrocytes from neonatal rat brain. At a concentration of 5.0 mM, which is necessary for growth inhibition and differentiation of a number of leukemic and solid tumour cell lines, astrocyte proliferation was reduced by 74.5+/-5.5%. On HMBA treatment astrocyte morphology changed from flat polygonal with phase translucent cytoplasm to compact retracted appearance with phase dark cytoplasm. HMBA also increased expression of GFAP, a marker for mature astrocytes. HMBA thus induces both morphological and biochemical differentiation of astrocytes. Like dbcAMP and staurosporine, two other known differentiation inducers of astrocytes, HMBA was also found to induce expression of the immediate early gene c-fos. However, unlike dbcAMP and staurosporine, which also induce fra-1, HMBA repressed cycloheximide-induced fra 1 gene expression. PMID- 10716206 TI - P and M channel-specific interference in the what and where pathway. AB - It was the purpose of this experiment to study interference effects of colour or luminance peripheral flicker (in order to saturate either the parvocellular or the magnocellular stream) on object identity and spatial location memory. Colour flicker interfered with object identity recognition, whereas luminance flicker affected memory for spatial location. Moreover, overall performance was worse if coloured rather than grey-scaled objects were used in the stimulus display. There was no selective effect of coloured flicker affecting coloured objects and chromatic flicker affecting chromatic objects. These results provide strong evidence for the theoretical position that the what pathway relies heavily on information derived from the P stream and the where pathway on the M stream. PMID- 10716205 TI - Non-conscious recognition of affect in the absence of striate cortex. AB - Functional neuroimaging experiments have shown that recognition of emotional expressions does not depend on awareness of visual stimuli and that unseen fear stimuli can activate the amygdala via a colliculopulvinar pathway. Perception of emotional expressions in the absence of awareness in normal subjects has some similarities with the unconscious recognition of visual stimuli which is well documented in patients with striate cortex lesions (blindsight). Presumably in these patients residual vision engages alternative extra-striate routes such as the superior colliculus and pulvinar. Against this background, we conjectured that a blindsight subject (GY) might recognize facial expressions presented in his blind field. The present study now provides direct evidence for this claim. PMID- 10716207 TI - Delta-9-tetrahydrocannabinol interferes with the establishment and the expression of conditioned rejection reactions produced by cyclophosphamide: a rat model of nausea. AB - Reliable animal models of nausea are necessary to better understand the neurobiology of nausea and to assess treatment effectiveness. We present such a model based on conditioned rejection reactions in rats. Our results demonstrate that delta-9-tetrahydrocannabinol (THC), a treatment reported to reduce chemotherapy-induced nausea in humans, also reduces conditioned rejection reactions in rats. Rats were administered THC or vehicle prior to a pairing of saccharin solution with cyclophosphamide or saline during conditioning and/or prior to test. THC interfered with the establishment of cyclophosphamide-induced conditioned rejection during conditioning and with the expression of conditioned rejection during testing. Our results confirm that the conditioned rejection reaction in the rat is a useful animal model of nausea. PMID- 10716208 TI - Tryptophan hydroxylase gene polymorphism (A218C) and suicidal behaviors. AB - Serotonergic dysfunction is implicated in mood disorders and suicidal behaviors. Genetic variants of tryptophan hydroxylase (TPH), a rate-limiting enzyme in the biosynthesis of serotonin, were associated with suicidal behaviors in three reports, but were not found in other studies. We investigated the TPH A218C polymorphism in 151 subjects with mood disorders and 200 control subjects. The results demonstrated a significant difference in genotypic distribution between controls and depressed patients, but not bipolar patients. A positive association between TPH polymorphism and suicidal behaviors was found in depressed patients and not in bipolar patients. We suggest that the association of TPH variants and suicide might depend on the diagnosis, and TPH mutation plays no major role in the pathogenesis of bipolar disorders. PMID- 10716209 TI - Administration of either non-NMDA receptor agonists or NMDA receptor antagonists into the substantia nigra or the globus pallidus reduces the psychostimulant effect of high helium pressure on locomotor activity in rats. AB - Helium pressure of >2 MPa is a well known factor underlying pressure-dependent central neuroexcitatory disorders that include locomotor and motor activity (LMA) and myoclonia. We investigated the effects of bilateral injection in either the substantia nigra (SN) or the globus pallidus (GP) of the AMPA receptor agonist (+/-)AMPA, the kainate receptor agonist kainic acid, the NMDA receptor agonist (+/-)-cis-piperidine-2,3-dicarboxylic acid (PDA), and the NMDA receptor antagonist (+/-)-2-amino-7-phosphono-heptanoic acid (AP-7) in the occurrence of helium pressure-induced LMA and myoclonia. Administration of AMPA, kainate, or AP 7 in either the SN or the GP significantly reduced high helium pressure-induced LMA, whereas the NMDA receptor agonist showed no significant effect. Injection in the SN of the non-NMDA receptor agonist AMPA and the NMDA receptor agonist PDA increased the development of high helium pressure-induced myoclonia, whereas injection of the NMDA receptor antagonist AP-7 into the SN or the GP decreased it. This confirms that NMDA transmission in the SN and the GP would play a major role in the development of helium pressure-induced LMA; manipulation of AMPA and kainate systems may have therapeutic potential. The opposite effects of AMPA on LMA and myoclonia also confirm the neural substrates involved in the motor disorder produced by helium pressure differ substantially between LMA and myoclonia. PMID- 10716210 TI - Regional distribution of tau, beta-amyloid and beta-amyloid precursor protein in the Alzheimer's brain: a quantitative immunolabelling study. AB - Regional variation in the distribution of SP and NFT within the brain is well documented. Consideration of such variation is potentially of help in formulating models of disease progression. Several models propose that pathological changes in Alzheimer's disease (AD) progress in a step-wise fashion along neuronally connected regions. In this study, we measured tau, Abeta and betaAPP load in different brain regions and examined our results against models of AD progression. Blocks of brain tissue from 45 AD and 15 control cases were immunolabelled for tau, Abeta and betaAPP. Immunolabelled areas were measured as a proportion of the area of the field. Tau load was almost twice as great in the entorhinal cortex than elsewhere in the brain and was least in the cingulate gyrus. In contrast, Abeta was greatest in the cingulate gyrus and least in the entorhinal cortex. BetaAPP rankings were similar to those of tau. Thus the site with the greatest Abeta load (cingulate cortex) had the least tau and the site with the greatest tau load (entorhinal cortex) had the least Abeta. The entorhinal and cingulate cortex are neuronally interconnected. Our results might be explained on the basis that a neurone with its cell body in the entorhinal cortex and axonal terminals in the cingulate cortex shows predominately tau pathology in relation to the cell body and predominately Abeta pathology in relation to its axonal terminals. We conclude that our observations are consistent with previously described models of AD progression. It is possible that tau-rich neurones are associated through their projections to Abeta rich sites. Further work of this kind analysing differential pathological profiles in interconnected brain regions may contribute to refining this model. PMID- 10716211 TI - Enlarged gamma band response of neuromagnetic auditory evoked fields in a visually impaired subject. AB - Under acoustic stimulation a phase-locked response in the gamma band (near 40 Hz) in the latency range between 20 and 130 ms is evoked. We report on a considerably visually impaired woman with Gronblad-Strandberg syndrome which involves degeneration at the level of retina, but has no overt central nervous component to the degeneration. The subject exhibited an extraordinarily high power in the phase-locked gamma band response (GBR) which was found to be more than three, and sometimes more than four, standard deviations above the average of a group of 25 subjects with normal vision. Furthermore, the dipoles of her mismatch reaction and M200 were found to be located posteriorly to the dipoles of the M100. Overall, both enlarged GBR and changed cortical representation could be results of cortical plasticity related to visual impairment. PMID- 10716212 TI - Oxidative stress increases ubiquitin--protein conjugates in synaptosomes. AB - Synaptosomes were incubated in the presence of FeSO4 to test the hypothesis that iron-catalyzed oxidative damage causes an increase in the ubiquitination of synaptosomal proteins. Incubation with 10 or 50 microM FeSO4 caused concentration dependent increases in carbonyl groups (an indication of protein oxidation) and ubiquitinated proteins (determined by probing Western blots with a monoclonal antibody to ubiquitin). Differences in protein ubiquitination occurred within 5 min of incubation, indicating a rapid response to oxidative stress. Results of experiments with MG-132, an inhibitor of the degradation of ubiquitinated proteins, suggested that oxidative damage stimulated ubiquitination rather than inhibited degradation of ubiquitinated proteins. The data are consistent with the hypothesis that synaptic terminals utilize the ubiquitin/proteasome proteolytic pathway to degrade oxidatively damaged proteins. PMID- 10716213 TI - Stages in motion processing revealed by the ocular following response. AB - Motion perception and associated involuntary eye movements depend on factors such as the physical attributes of the stimulus and visual attention. Cues from spatial changes in luminance (first-order motion in the Fourier domain) or more complicated transitions involving two-dimensional patterns (second-order, non Fourier) require rather different kinds of analyses to detect their net motion. During a fixation task we monitored eye movements induced by the onset of motion to examine the functional properties of the monkey cortical motion processing system. Eye movement velocity was indistinguishable to first- and second-order motion; concomitant response latency confirmed an additional calculation is required to detect the direction and velocity of second-order motion. PMID- 10716215 TI - Complex cells as linear mechanisms receiving sequential afferents. AB - Complex cells in mammalian visual cortex appear to be non-linear mechanisms lacking a structured receptive field, and different complex cells display mutually inconsistent behaviors. Current models postulate nonlinear interactions among multiple simultaneous afferents, but none explains the diversity of complex cell behaviors. We propose that complex cell inputs are sequential and cyclic. Cells receiving such input behave as if their spatial receptive field changes shape over time. Different putative time-varying receptive fields arise when the number of afferents, their characteristics and/or the sequence of their inputs vary, and simulations show that they exhibit all reported varieties of complex cell behavior. Our results suggest a common functional description for simple and complex cells. Additional non-linearities are unnecessary to explain complex cell behavior. PMID- 10716214 TI - Functional localization of the sensory hand area with respect to the motor central gyrus knob. AB - The aim of this work was to investigate the topography of the primary sensory hand cortex with magnetoencephalography in order to define the functional anatomy of this area in healthy humans. Previous studies denoted an inverted Ohm or an horizontal epsilon shaped knob on the pre-central gyrus as a landmark for the motor hand area; therefore a systematic difference between the orientation of the source for thumb with respect to little finger should be observed. We found this systematic difference, but the direction of the sources activated during thumb and little finger stimulation did not converge, as would be expected if only the Ohm convexity is activated: in fact our results suggest that thumb sensory area also extends to the area lateral to this convexity. PMID- 10716216 TI - Bradykinin-induced neurogenic migration of neutrophils into the rat knee joint. AB - This study examined the dependence of neurogenic and non-neurogenic synovial plasma extravasation on neutrophils. Perfusion of bradykinin into the knee joint produced both a rapid increase in the magnitude of plasma extravasation and a significant increase in number of neutrophils in the synovium. Both bradykinin induced plasma extravasation and neutrophil accumulation were dependent on sympathetic post-ganglionic neuron terminals, since both were blocked in sympathectomized rats. Platelet activating factor, which produces plasma extravasation independent of sympathetic neurons, did not increase the number of neutrophils in the synovium. These findings support the suggestion that bradykinin acts on sympathetic nerve terminals in the knee leading to attraction of neutrophils, which promotes plasma extravasation. PMID- 10716217 TI - A common neural substrate for the analysis of pitch and duration pattern in segmented sound? AB - The analysis of patterns of pitch and duration over time in natural segmented sounds is fundamentally relevant to the analysis of speech, environmental sounds and music. The neural basis for differences between the processing of pitch and duration sequences is not established. We carried out a PET activation study on nine right-handed musically naive subjects, in order to examine the basis for early pitch- and duration-sequence analysis. The input stimuli and output task were closely controlled. We demonstrated a strikingly similar bilateral neural network for both types of analysis. The network is right lateralised and includes the cerebellum, posterior superior temporal cortices, and inferior frontal cortices. These data are consistent with a common initial mechanism for the analysis of pitch and duration patterns within sequences. PMID- 10716218 TI - Platelet-activating factor inhibits ionotropic GABA receptor activity in cultured hippocampal neurons. AB - Platelet-activating factor (PAF), one of the most potent bioactive lipids, has been implicated in modulating long-term potentiation (LTP) and neurotoxicity. In the CNS, glutamate and GABA are the major excitatory and inhibitory neurotransmitters, respectively. Previous work has focused on the effects of PAF on glutamatergic receptor responses. The purpose of the present study was to investigate the possible actions of PAF on ionotropic GABA receptor responses in primary cultured hippocampal neurons using the whole-cell and single channel patch clamp techniques. Extracellular application of PAF induced a reduction of the GABA gated Cl- current in a majority of cells (29 of 44 cells), while it caused an enhancement in 10 of 44 cells. A similar heterogeneous modulation of PAF on the GABA receptor activities was also observed in outside-out patch recordings. Moreover, the cell-attached single channel recordings showed that PAF decreased the GABA channel activity. Therefore, PAF may modulate synaptic activity by inhibiting GABA receptor channels. During seizures and neural injury, when enhanced synthesis of this lipid mediator takes place, the actions of PAF on inhibitory GABA receptors may contribute to synaptic dysfunction. PMID- 10716219 TI - Electrophysiological evidence for sequential activation of multiple brain regions during the auditory selective attention process in humans. AB - In an attempt to examine dynamic involvement of multiple brain regions in the auditory selective attention process, negative difference wave (Nd) generators were assessed using a high-resolution EEG system (128ch) and scalp current density (SCD) analysis. Ten normal volunteers participated in the study. Event related potentials were recorded during a selective attention task. Sequential SCD mappings revealed that current sinks were located in the bilateral temporal regions at 160 ms subsequent to the onset of stimuli, shifting the dipole orientation more tangentially to the scalp at around 220 ms. Moreover, a current sink was demonstrated in the midfrontal region at around 320 ms. These findings confirm that different cortical regions are sequentially involved in the auditory selective attention process. PMID- 10716220 TI - Human NT2/D1 cells differentiate into functional astrocytes. AB - As the most numerous cell type in the brain, astrocytes are coupled via gap junction channels. It is believed that communication among astrocytes is normally regulated by extracellular ions, neurotransmitters and neuromodulators. However, the level of astrocytic coupling is altered in abnormal conditions such as stroke, brain trauma and Alzheimer's disease. A well established human progenitor cell line, NT2/D1, has been previously differentiated into pure neuronal cultures. In the current study, for the first time, we report the differentiation of NT2/D1 cells into astrocytes, which express connexin43 and are coupled via gap junctions. Thus, human NT2/D1 cells are not merely committed neuronal progenitors but, similar to the embryonal stem cells, they can give rise to both lineages. PMID- 10716221 TI - Repeated injections of lipopolysaccharide attenuate the satiety effects of cholecystokinin. AB - The present study examined the effects of repeated injections of lipopolysaccharide (LPS) and cholecystokinin (CCK) on both sucrose palatability and body weight. Rats received repeated injections of LPS (200 microg/kg), CCK (8 microg/kg) and/or NaCl on days 1, 4, 7 and 10. Body weight was monitored on each test day and sucrose palatability was assessed using the taste reactivity test (TRT) on days 7 and 10. Rats treated with LPS developed a rapid tolerance to the reductions in body weight; however, this tolerance did not affect sucrose palatability. Furthermore it was found that repeated exposures to LPS and CCK attenuated the typical satiety related behaviors produced in the TRT by administration of CCK. This desensitization to CCK with repeated exposures to LPS may be one of the mechanisms that could account for the rapid development of tolerance to LPS-induced hypophagia. PMID- 10716222 TI - NTS catecholamine cell recruitment by hemorrhage and hypoxia. AB - Immunolabelling for Fos and tyrosine hydroxylase was used to determine the patterns of activation of nucleus tractus solitarius catecholamine cells in response to graded levels of hemorrhage (0, 4, 8, 12 and 16 ml/kg) and systemic hypoxia (21, 14, 12, 10 and 8% O2) in conscious rats. Both stimuli elicited graded catecholamine cell recruitment with thresholds of 8 ml/kg and 12% O2. The majority of responsive neurons were A2 noradrenergic rather than C2 adrenergic cells. After hemorrhage most Fos-positive catecholamine cells were found below obex whereas most hypoxia-responsive cells were rostral to obex. These distinctive patterns of catecholamine cell recruitment may explain the differences in neuroendocrine responses to these stimuli. PMID- 10716223 TI - Maturation of endomorphin-2 in the dorsal horn of the medulla and spinal cord of the rat. AB - The endomorphins are potent and selective endogenous agonists at the mu opioid receptor. We describe here the postnatal ontogeny of endomorphin-2 like immunoreactivity (EM2-LI) in the dorsal horn of the rat medulla and spinal cord. EM2-LI is dense in the superficial lamina of the dorsal horn of the adolescent and adult rat, with fibers also present in skin and dorsal root ganglia. No staining was noted at 3 days of age or younger. Faint and limited staining was noted by 7 days of age. The density and distribution of the immunoreactivity increased with age, reaching an adult-like distribution by 21 days of age. Stress induced responses mediated by endogenous opioids occur late in development and may be related to the late appearance of endomorphin-2. PMID- 10716224 TI - Expression of metabotropic glutamate receptor subtype mRNA (mGluR1-8) in human cerebellum. AB - The expression of metabotropic glutamate receptor (mGluR) mRNAs in the human cerebellar neocortex was investigated using in situ hybridization. mGluR1, 3 and 4 mRNAs were most abundant and widely distributed, whereas mGluR2, 5 and 7 mRNA expression was circumscript. mRNAs coding for mGluR1, 3, 4 and 7 were expressed in Purkinje cells. mGluR1 and 4 mRNAs detectable in granule cells and mGluR1, 2, 3, 4 and 7 mRNAs in Golgi cells. mGluR5 mRNA was detectable in putative Bergmann glia as well as mGluR3 mRNA, which was widely expressed in glial cells. mGluR8 mRNA expression was very low in a subset of stellate/basket cells of the molecular layer whereas mGluR6 mRNA was not detectable in the cerebellar cortex. PMID- 10716225 TI - NMDA receptor blockade prevents LTD, but not LTP induction by intracellular tetanization. AB - Intracellular tetanization, the activation of a postsynaptic cell without concomitant presynaptic stimulation, was applied to layer II/III pyramidal cells in slices of rat visual cortex. In standard extracellular medium, intracellular tetanization led to LTP (21 of 43 inputs) or LTD (14 of 43 inputs), the direction of the amplitude change depending on initial paired-pulse facilitation (PPF) ratio: inputs with high initial PPF ratio were usually potentiated, and inputs with initially low PPF were most often depressed or did not change. When applied during blockade of NMDA receptors (50 microM APV), intracellular tetanization failed to induce LTD, but was still capable of inducing LTP (14 of 26 inputs). Although LTP could occur in inputs with both, low and high initial PPF ratio, the correlation between the amplitude change and initial PPF ratio remained: potentiation was stronger in inputs with initially higher PPF. These data suggest that intracellular tetanization activated simultaneously NMDA receptor-dependent LTD mechanisms and NMDA receptor-independent LTP mechanisms, the final change of synaptic gain depending on their balance. PMID- 10716226 TI - Processing of the beta-amyloid precursor protein in ex vivo human brain cells. AB - We studied distribution and processing of the Alzheimer's beta-amyloid precursor protein (betaAPP) in immediately ex vivo human brain cells obtained during neurosurgical procedures. Immunoblotting and flow cytometry studies revealed that brain cells supported betaAPP as a transmembrane holoprotein. Brain cells in short-term suspension culture were competent to process betaAPP into Abeta as shown by [35S]methionine pulse-chase studies. Brain cell Abeta was immunoprecipitated as SDS-stable dimers and higher-order multimers. Cleavage of cell surface betaAPP with trypsin prior to metabolic labeling reduced cellular Abeta by approximately 50%. We conclude that plasmalemmal betaAPP in human brain cells is a source of cellular Abeta, presumably via endosomal-lysosomal processing. PMID- 10716227 TI - Violation of homogeneity by the vestibulo-ocular reflex of the goldfish. AB - The vestibulo-ocular reflex (VOR) allows animals to maintain stable gaze during head rotations by generating compensatory eye rotations. The VOR is typically tested using sinusoidal head rotation, and VOR gain is calculated as the ratio of the amplitude of eye to head rotational velocity. Through habituation, prolonged exposure to lower frequency sinusoidal head rotation in the dark decreases VOR gain. The VOR has been treated and modeled as a linear system. If it is linear, then the VOR must obey the principle of homogeneity: VOR gain at a particular frequency should be the same regardless of head velocity. We examined the habituated goldfish VOR for homogeneity. We found that it violated this basic principle of linear systems and is therefore non-linear. PMID- 10716228 TI - 85 kDa cytosolic phospholipase A2 is a target for chronic lithium in rat brain. AB - The mechanism by which chronic lithium exerts its therapeutic effect in brains of bipolar patients is not known. One possibility, suggested by our demonstration in the rat brain, is that chronic lithium inhibits turnover of arachidonic acid (AA) by reducing the activity of an AA-specific phospholipase A2 (PLA2). To test this further, mRNA levels of two AA-specific PLA2s, cytosolic PLA2 (cPLA2) type IV and intracellular PLA2 (iPLA2) type VIII, and protein level of cPLA2 were quantified in the brain of rats given lithium for 6 weeks. Chronic lithium markedly reduced brain mRNA and protein level of cPLA2, but had no effect on mRNA level of iPLA2. These results suggest that the final common path effect of chronic lithium administration is to reduce turnover of AA in brain by down-regulating cPLA2. PMID- 10716229 TI - Intranuclear inclusions in subtypes of striatal neurons in Huntington's disease transgenic mice. AB - R6/2 transgenic mice express exon 1 of an abnormal human Huntington's disease (HD) gene and develop a neurological phenotype similar to HD. These mice develop ubiquitinated neuronal intranuclear inclusions (NII) which might play a central role in the pathophysiology of HD. We studied the distribution of NII in subpopulations of striatal neurons in 12-week-old R6/2 transgenic mice using fluorescent double label immunohistochemistry. We observed that most of the Calbindin-D28K positive projection neurons (89%) and the Parvalbumin positive interneurons (86%) showed ubiquitinated NII. In interneurons, however, which contain either choline acetyltransferase, neuronal nitric oxide synthase, or Calretinin, the frequency of NII was much lower (22%, 8%, 9%, respectively). Our data suggest that subpopulations of striatal neurons differ remarkably in their capability of forming ubiquitinated NII. Interneurons which are known to resist neurodegeneration in HD show less NII. PMID- 10716230 TI - Caffeine-sensitive Ca2+ stores in carp retinal bipolar cells. AB - High K+- or caffeine-induced Ca2+ signal was studied in freshly dissociated carp retinal ON-type bipolar cells using a confocal laser-scanning microscope. In response to 35 mM K+ exposure, a rise in [Ca2+]i appeared in both the terminal and soma, but was absent after removal of external Ca2+ or in the presence of 100 microM nifedipine. It is indicated that, for high K+-induced increase of [Ca2+]i, Ca2+ influx through voltage-gated L-type Ca2+ channels is essential and Ca2+ entry through reversed Na+/Ca2+ exchange may be negligible. Interestingly, caffeine-induced elevation of [Ca2+]i was restricted to the soma, and could be abolished by 50 microM ryanodine, suggesting that caffeine-sensitive Ca2+ stores gated by ryanodine receptors were present in the soma but not in the terminal of bipolar cells. After treatment with 50 microM ryanodine for 20 min, the peak of the Ca2+ transients evoked by 35 mM K+ in the soma decreased to 48.2+/-5.7% of the control. The results suggest that depolarization-evoked Ca2+ influx can cause Ca2+ release from caffeine-sensitive Ca2+ stores, and in turn amplify Ca2+ signal in the soma of retinal bipolar cells. PMID- 10716231 TI - Localization of FGFR-1 in axotomized and peripheral nerve transplanted ferret retina. AB - Retinal ganglion cells (RGCs) survive axotomy and regenerate their axons into the peripheral nerve graft in adult mammals. To understand the potential role of fibroblast growth factors (FGFs) in survival and regeneration of axotomized RGCs, we examined FGF receptor 1 (FGFR-1) localization in normal and PN grafted ferret retinas by immunohistochemistry in combination with retrograde labeling. Prominent expression of FGFR-1 was observed in outer plexiform layer and ganglion cell layer of normal ferret retina. In the ganglion cell layer, FGFR-1 immunoreactivity was detected in about 30% of RGCs, predominantly in large cells. In the PN grafted ferret retina, 90% of RGCs with regenerated axons expressed FGFR-1. Our findings suggest that FGFs may play an important role in the survival and axonal regeneration of RGCs of adult mammals. PMID- 10716232 TI - The nicotinic acetylcholine receptor agonist ABT-594 increases FGF-2 expression in various rat brain regions. AB - The present experiments were designed to extend previous work showing that acute intermittent (-)nicotine treatment upregulates the level of fibroblast growth factor-2 (FGF2) mRNA in several rat brain regions, by the use of the nicotinic acetylcholine receptor (nAChR) agonist ABT-594 with preferential selectivity for the alpha4beta2 nAChR subtype. ABT594 treatment led to a well-defined temporal and regional upregulation of FGF-2 mRNA. A double labelling analysis showed that the up-regulation of FGF-2 mRNA involves both neuronal and non-neuronal cells. The effects of ABT-594 on FGF-2 expression were antagonized by the preferential alpha4beta2 antagonist dihydrobetaerythroidine (DHbetaE), but not by alpha7 antagonist methyllycaconitine (MLA). In conclusion, FGF-2 mRNA levels can be increased in several brain regions upon alpha4beta2 nAChR activation, suggesting a therapeutic significance in neurodegenerative disorders. PMID- 10716233 TI - Linear spatio-temporal convergence in vestibular neurons of the primate nucleus fastigii. AB - Vestibular responses in the primate fastigial nucleus (FN) do often not follow the simple cosine tuning observed in primary vestibular afferents. The present report demonstrates that these more complex patterns can mostly be attributed to simple linear summation of spatially and temporally diverse cosine-tuned input signals (linear spatio-temporal convergence, STC). Analyses following from this elementary finding, however, reveal frequency-dependent properties in many FN neurons, which are difficult to reconcile with existing concepts of possible functions of STC in central vestibular areas. The demonstration that STC linearity holds for FN responses is thus of both theoretical and practical relevance, allowing shortening of future experimental protocols and facilitating comparison of the observed spatio-temporal response dynamics with those at other stages of vestibular signal processing. PMID- 10716234 TI - Interaction of CPCCOEt with a chimeric mGlu1b and calcium sensing receptor. AB - 7-Hydroxyiminocyclopropan[b]chromen-1a-carboxylic acid ethyl ester (CPCCOEt) has previously been shown to be a selective non-competitive antagonist at the metabotropic glutamate (mGlu) receptor subtype 1. In this study we have tested the effect of CPCCOEt on mGlu1b, the calcium sensing receptor (CaR) and a chimeric receptor consisting of the agonist binding amino-terminal domain (ATD) of CaR and the seven transmembrane (7TM) domain of mGlu1b (named Ca/1b). CPCCOEt inhibited responses of (S)-glutamic acid and Ca2+ at mGlu1b and Ca/1b, applied at EC50 values, with IC50 values of 10.2 microM and 13.4 microM, respectively, whereas it was weak as an antagonist of Ca2+ at CaR. These data provides strong evidence that CPCCOEt exerts its antagonistic effect on mGlu1 solely by binding to the 7TM domain. PMID- 10716235 TI - Protection of neonatal rat brain from hypoxic-ischemic injury by LY379268, a Group II metabotropic glutamate receptor agonist. AB - Neuroprotective effects of a Group II metabotropic glutamate receptor agonist, LY379268, were examined in a neonatal rat model of hypoxia-ischemia (unilateral common carotid artery ligation followed by hypoxic exposure for 1.5h in 7-day-old rat pups). LY379268 administered 5 min after hypoxic exposure (2, 5, or 10 mg/kg, i.p.) significantly reduced brain injury as measured by reductions in the ipsilateral brain weight and in CA1 hippocampal neuron density. The significant neuroprotective effects were also observed when this compound (5 mg/kg) was administered 30 min, but not 60 min, after hypoxic exposure. The neonatal hypoxia ischemia (HI) procedure significantly increased caspase-3 activity and induced DNA fragmentation in the ipsilateral cortex compared with that in the contralateral cortex 24 and 72h after the insult, respectively. LY379268 did not prevent this increase in caspase-3 activity and DNA fragmentation in the ipsilateral cortex. These results suggest that activation of Group II metabotropic glutamate receptors may provide neuroprotection against HI brain injury. However, blockade of caspase-3 activation and the apoptotic pathway appears not to be involved in the neuroprotective effects of LY379268 observed in the neonatal rat model of HI. PMID- 10716236 TI - Effect of adenosine A2A receptor stimulation on GABA release from the striatum of young and aged rats in vivo. AB - The effect of the adenosine A2A receptor agonist CGS 21680 on GABA release from the striatum was investigated in vivo in young and old rats by microdialysis experiments. CGS 21680 at 10 microM significantly increased GABA spontaneous outflow in young but not in old rats. Conversely, CGS 21680 significantly increased potassium-evoked GABA release in old but not in young rats. The stimulating effect of CGS 21680 on spontaneous GABA release may be both secondary to an increased outflow of excitatory amino acids and to the removal, brought about by A2A receptor stimulation, of the inhibitory influence exerted by dopamine through D2 receptors on GABA-enkephalinergic neurones. The stimulating effect of CGS 21680 on potassium-evoked GABA release in old rats may be ascribed to an age-dependent imbalance in favour of adenosine vs. dopamine, recently described in the striatum of old rats. PMID- 10716237 TI - Reactive astrocytes express p53 in the spinal cord of transgenic mice expressing a human Cu/Zn SOD mutation. AB - In a previous study, we reported increased NOS expression in the astrocytes in the spinal cord of SOD mutant transgenic mice that are used as ALS animal model. Recently, Messmer and Brune suggested that nitric oxide-induced apoptosis is intimately related with p53-dependent signaling pathway, and de la Monte et al. reported increased p53-immunoreactivity in the spinal cord of ALS patients. In the present study, we performed immunocytochemical studies to investigate the changes of p53-immunoreactivity in the brains of the mutant transgenic mice expressing a human Cu/Zn SOD mutation. Immunocytochemistry showed intensely stained p53-IR glial cells with the appearance of astrocytes in all levels of the spinal cord of the mutant transgenic mice, but no p53-IR glial cells were observed in the spinal cord of the control mice. P53-IR astrocytes were also detected in the brain stem of the mutant transgenic mice. In the medulla, they were observed in the medullary reticular formation, hypoglossal nucleus, vestibular nucleus, dorsal motor nucleus of the vagus and nucleus ambiguus. In the pons, their presences were noted in the pontine reticular formation, and trigeminal and facial nuclei. In the midbrain, astrocytes were detected in the mesencephalic reticular formation, red nucleus and periaqueductal gray matter. In the cerebellum, intensely stained p53-IR astrocytes were detected in the intracerebellar nuclei. In contrast to the mutant transgenic mice, no p53-IR astrocytes were detected in the brain stem and spinal cord of the control mice. Further multidisciplinary investigations involving p53-mediated cellular damage and pathogenesis of ALS are needed to clarify the importance of these results. PMID- 10716238 TI - APOE epsilon4 allele and amyloid beta-protein deposition in long term survivors of head injury. AB - Head injury and APOE epsilon4 are risk factors for Alzheimer's disease (AD). We previously found that deposits of amyloid beta-protein (Abeta) occur in fatal head injury, more frequently in patients with APOE epsilon4. We postulated that Abeta deposits triggered by injury could, in survivors, lead to AD-like pathology later in life. Here, we compared Abeta deposits in 21 long term survivors of head injury (up to 20 years) with age and APOE genotype matched controls. In both groups Abeta deposits were more common among patients with APOE epsilon4. However, Abeta deposits were not more common among survivors of head injury than controls. The findings support previous studies associating APOE epsilon4 with deposition of Abeta. However pathogenetic mechanisms other than Abeta deposition may explain the association of head injury with AD. PMID- 10716239 TI - Immunosuppressants promote adult dorsal root regeneration into the spinal cord. AB - Immunosuppressants promote neurite extension in culture and facilitate regeneration of peripheral nerves in vivo. However, their neurotrophic effects in the CNS have not been well studied. We utilized a rat dorsal root transaction model to examine the effects of cyclosporine A (CsA) and FK 506 on regeneration of the dorsal root into the spinal cord. After surgery, the rats received daily subcutaneous injections of CsA or FK 506. One month after surgery, dorsal root axons were immunohistochemically labeled to evaluate the extent of regeneration into the spinal cord. Dorsal root axons of CsA/FK 506-treated rats frequently entered into the spinal cord and arborized extensively. Administration of immunosuppressants markedly promoted regeneration of dorsal root axons into the spinal cord. PMID- 10716240 TI - Spontaneous axonal regeneration after optic nerve injury in adult rat. AB - Optic nerves of adult rats were crushed 2 mm behind the eye to examine the ability of retinal ganglion cells (RGCs) to regenerate their axons. Some animals were treated with the immunophilin ligands FK 506 or GPI 1046 for up to 4 weeks. After 10 days to 16 months, regenerating RGC axons were visualized using anterograde tracing and/or electron microscopy. A small proportion of RGC axons regenerated across the lesion site and grew very slowly along the entire optic nerve. Immunophilin ligands had no obvious effect. The regenerating axons were about 0.2 microm in diameter, and usually in clusters surrounded by astrocyte processes. Thus, some CNS axons can spontaneously regenerate long distances within degenerate white matter and this slow regeneration is not accelerated by immunophilin ligands. PMID- 10716241 TI - Magnetic resonance imaging of transplanted oligodendrocyte precursors in the rat brain. AB - The lack of any markers for oligodendrocyte precursors that can be visualized within the intact CNS is a significant barrier to trials of transplantation of these cells which aim to enhance remyelination in multiple sclerosis. We have therefore asked whether dextran-coated superparamagnetic iron oxide (SPIO) can be used to label cells prior to transplantation and then visualized within the brain using MRI. We have shown that an oligodendrocyte precursor cell line CG-4 will take up dextran-coated SPIO particles in vitro. The label remains within the cells after transplantation into adult rat brain, as assessed by electron microscopy, and is visible by MRI as a reduction in signal intensity at the transplant site at both 1 and 7 days after surgery. We conclude that MRI detection of SPIO-labelled cells represents a promising and novel approach to the analysis of oligodendroglial cell behaviour following transplantation that has very significant advantages over currently available methods. PMID- 10716242 TI - Serotonin and substance P co-exist in dorsal raphe neurons of the human brain. AB - A substance P antagonist has recently been reported to have clinical efficacy in the treatment of depression. We have therefore analysed sections from human pons, including the raphe region, with double in situ hybridization using riboprobes complementary to substance P and the 5-hydroxytryptamine transporter (5-HT-T mRNAs). A distinct overlap of cell bodies expressing these two markers was observed in the dorsal and median raphe nuclei. Analysis of double-labelled sections revealed that almost half of the 5-HT neurons in the dorsal raphe and around 25% in the median raphe nucleus expressed substance P mRNA. The highest percentage was observed in the ventrolateral dorsal raphe nucleus and the lowest in the caudal raphe nucleus. These results demonstrate that the phenotype of the raphe 5-HT neurons varies between species, since so far no 5-HT-substance P co existence has been demonstrated in the dorsal raphe complex of rat. The question is raised whether the present results may be of significance for understanding a possible role of substance P in depression. PMID- 10716243 TI - Differentiation of transplanted neural precursors varies regionally in adults striatum. AB - In the current experiments, we address the emerging hypothesis that transplanted neural precursor cells can respond to local microenvironmental signals in the post-developmental brain and exhibit patterns of differentiation that depend critically on specific location within the brain. HiB5 precursor cells were transplanted into adult mouse cortex, corpus callosum, and multiple positions in striatum, and assessed for differentiation by morphology and immunocytochemistry. Our results indicate that the likelihood of both neuronal and glial differentiation of transplanted precursors depends on proximity to the medial striatum or subventricular zone of the adult host, supporting the concept that microenvironmental signals can critically affect the differentiation fate of neural precursors, and suggesting the potential to manipulate such signals in the adult brain. PMID- 10716244 TI - Localization of cerebral activity during simple singing. AB - Cerebral blood flow (CBF) was measured with PET during rudimentary singing of a single pitch and vowel, contrasted to passive listening to complex tones. CBF increases in cortical areas related to motor control were seen in the supplementary motor area, anterior cingulate cortex, precentral gyri, anterior insula (and the adjacent inner face of the precentral operculum) and cerebellum, replicating most previously seen during speech. Increases in auditory cortex were seen within right Heschl's gyrus, and in the posterior superior temporal plane (and the immediately overlying parietal cortex). Since cortex near right Heschl's has been linked to complex pitch perception, its asymmetric activation here may be related to analyzing the fundamental frequency of one's own voice for feedback guided modulation. PMID- 10716245 TI - Peroxisome 1, 2, 3... PMID- 10716246 TI - Alzheimer's disease risk and the interleukin-1 genes. PMID- 10716247 TI - Pharmacological induction of peroxisomes in peroxisome biogenesis disorders. AB - Inherited aberrant peroxisome assembly results in a group of neurological diseases termed peroxisome biogenesis disorders (PBDs). PBDs include three major clinical phenotypes that represent a continuum of clinical features from the most severe form, Zellweger syndrome (ZS), through neonatal adrenoleukodystrophy (NALD) to the least severe form, infantile Refsum's disease (IRD). Somatic cell complementation studies have identified 13 PBD complementation groups, each representing a defect in a peroxisomal protein (peroxin) involved in peroxisome biogenesis. Most complementation groups include a range of clinical phenotypes. In this study, peroxisome numbers were determined in fibroblasts from 29 PBD (ZS, NALD, and IRD) patients, with various phenotypes from nine complementation groups, using antibodies against either a peroxisomal membrane protein (anti Pex14p) or peroxisomal matrix proteins (anti-SKL). A correlation between the number of peroxisomes, determined with either antibody, and PBD phenotype was found, suggesting that induction of peroxisome number might have a favorable effect on PBD. After treatment of PBD fibroblasts with sodium 4-phenylbutyrate, a human peroxisome proliferator, there was an approximate twofold increase in peroxisome number. After 4-phenylbutyrate treatment, an increase in transcription of the adrenoleukodystrophy-related gene and the peroxin gene, PEX11alpha, was found in PBD fibroblasts. In NALD and IRD, but not ZS, fibroblasts there was an increase in very-long-chain fatty acid beta-oxidation and plasmalogen concentrations, and a decrease in very-long-chain fatty acid concentrations. These data suggest that pharmacological agents that induce peroxisome proliferation, such as 4-phenylbutyrate, may have therapeutic potential in the treatment of PBD patients with milder phenotypes (NALD and IRD). PMID- 10716248 TI - New Purkinje cell antibody (PCA-2): marker of lung cancer-related neurological autoimmunity. AB - Neuron-restricted autoantibodies are important markers of neurological autoimmunity related to cancer. We identified a new paraneoplastic IgG, PCA-2 (Purkinje cell cytoplasmic antibody type 2), in 10 patients. Nine had mixed subacute neurological presentations (5 brainstem or limbic encephalitis, 3 cerebellar ataxia, 2 Lambert-Eaton myasthenic syndrome, 1 autonomic neuropathy, and 1 motor neuropathy). All 9 were smokers, and 8 had definite or probable lung cancer (7 with biopsy-confirmed small cell lung carcinoma [SCLC]; 1 imaged only). One patient had no follow-up information. A 10th patient was among 58 with uncomplicated SCLC. PCA-2 binds to a cytoplasmic antigen in neurons and SCLC cells. Its immunostaining pattern in mouse tissues is distinct from that of the paraneoplastic autoantibodies PCA-1 (anti-Yo, marker of immune response initiated by ovarian or breast carcinoma) and PCA-Tr (anti-Tr, immune response marker of Hodgkin's lymphoma). PCA-2 binds to cerebellar Purkinje somata and dendrites, neurons in internal granular layer and dentate nucleus, and neuronal elements in gut and kidney. Western blots of reduced/denatured cerebellar and SCLC proteins reveal a common antigenic band, of approximately 280 kd. PCA-2 is the seventh IgG neuronal autoantibody marker of paraneoplastic autoimmunity identifiable unambiguously by standardized immunofluorescence criteria. PMID- 10716249 TI - Increased lymphoproliferative response to human herpesvirus type 6A variant in multiple sclerosis patients. AB - Several reports have suggested an association of human herpesvirus 6 (HHV-6) and multiple sclerosis (MS) based on immunohistochemical demonstration of HHV-6 antigens in inflammatory lesions, detection of increased HHV-6 specific serum antibody titers, and amplification of HHV-6 DNA from sera and cerebrospinal fluid of MS patients but not in controls. Characterization of the cellular immune response of MS patients to HHV-6 may further clarify the role of HHV-6 in MS and provide insight into the pathogenesis of this immune-mediated disease. We have compared lymphoproliferative responses to HHV-6A (U1102)-, HHV-6B (Z29)-, and HHV 7 (H7SB)-infected cell lysates in healthy controls and patients with MS. Most healthy controls (71%) proliferated to HHV-6B lysate, and fewer (33%) responded to the HHV-6A lysate. In contrast, 67% of MS patients had a lymphoproliferative response to HHV-6A, which is a significant increase in comparison with healthy controls. A similar frequency of lymphoproliferative response (78%) to HHV-6B was demonstrated in MS patients. Lymphoproliferation to HHV-7 lysate was demonstrated in 23% of healthy controls and 28% of MS patients. These results indicate that the lymphoproliferative response to the HHV-6A variant, which was recently reported to have greater neurotropism, is increased in MS patients. PMID- 10716250 TI - Clinical features and response to treatment in Guillain-Barre syndrome associated with antibodies to GM1b ganglioside. AB - GM1b is a minor ganglioside in human peripheral nerves. Serum anti-GM1b antibodies frequently are present in patients with Guillain-Barre syndrome (GBS). In this collaborative study, we investigated the antecedent infections, clinical features, and response to treatment of GBS patients with anti-GM1b antibodies. Of 132 GBS patients who participated in the Dutch GBS trial that compared the effect of intravenous immunoglobulins and plasma exchange, 25 (19%) patients had anti GM1b antibodies. IgM antibodies were present in 14, IgG antibodies in 15, and both isotypes in 4 patients. The 25 patients with anti-GM1b antibodies had a clinical pattern distinct from that of the other 107 GBS patients. They more often had an episode of gastrointestinal illness and frequently showed serological evidence of recent infection by Campylobacter jejuni. The anti-GM1b positive subgroup was marked by more rapidly progressive, more severe, and predominantly distal weakness. Cranial nerve involvement and sensory deficits were less common in the patients with anti-GM1b antibodies. The presence of anti GM1b antibodies was associated with slower recovery. The clinical manifestations predominantly were associated with anti-GM1b antibodies of the IgG isotype. Fourteen (56%) of the 25 patients with anti-GM1b antibodies also had anti-GM1 antibodies. The group of patients with both antibodies was clinically more homogeneous and had a more rapidly progressive, pure motor neuropathy. The subgroup of anti-GM1b-positive GBS patients responded well to treatment with immunoglobulins but not to plasmapheresis. The distinctive clinical features of the patients with anti-GM1b antibodies show that acute motor neuropathy represents a specific subgroup within GBS and that recognizing these patients may have consequences as to the choice of therapy. PMID- 10716251 TI - Sensory tricks in cervical dystonia: perceptual dysbalance of parietal cortex modulates frontal motor programming. AB - Cervical dystonia is a disabling basal ganglia disorder characterized by an involuntary head deviation to one side. A typical but also mysterious feature is the impressive improvement of muscle spasms and involuntary head posture by application of a sensory facial stimulus (sensory trick). Here, we report the effect of a sensory trick on cortical activation patterns in 7 patients with cervical dystonia by using H2(15)O positron emission tomography. The application of the sensory trick stimulus, resulting in a near-neutral head position, led to an increased activation mainly of the superior and inferior parietal lobule (ipsilateral to the original head turn) and bilateral occipital cortex and to a decreased activity of the supplementary motor area and the primary sensorimotor cortex (contralateral to the head turn). We propose that a perceptual dysbalance induced by a sensory trick maneuver leads to a relative displacement of the egocentric midvertical reference to the opposite side and a decrease in motor cortex activity. This modulation of motor programming gives novel insights into the mechanisms involved in sensorimotor integration in movement disorders. PMID- 10716252 TI - Endothelin inhibition improves cerebral blood flow and is neuroprotective in pneumococcal meningitis. AB - By using an infant rat model of pneumococcal meningitis, we determined whether endothelins contribute to neuronal damage in this disease. Cerebrospinal fluid analysis demonstrated a significant increase of endothelin-1 in infected animals compared with uninfected controls. Histopathological examination 24 hours after infection showed brain damage in animals treated with ceftriaxone alone (median, 9.2% of cortex; range, 0-49.1%). In infected animals treated intraperitoneally with the endothelin antagonist bosentan (30 mg/kg, every 12 hours) also, injury was reduced to 0.5% (range, 0-8.6%) of cortex. Cerebral blood flow was reduced in infected animals (6.5 +/- 4.0 ml/min/100 g of brain vs 14.9 +/- 9.1 ml/min/100 g in controls. Treatment with bosentan restored cerebral blood flow to levels similar to controls (12.8 +/- 5.3 ml/min/100 g). Improved blood flow was not mediated by nitric oxide production, because bosentan had no effect on cerebrospinal fluid or plasma nitrite/nitrate concentrations at 6, 12, or 18 hours. These data indicate that endothelins contribute to neuronal injury in this model of pneumococcal meningitis by causing cerebral ischemia. PMID- 10716253 TI - Prolonged febrile seizures in the immature rat model enhance hippocampal excitability long term. AB - Febrile seizures (FSs) constitute the most prevalent seizure type during childhood. Whether prolonged FSs alter limbic excitability, leading to spontaneous seizures (temporal lobe epilepsy) during adulthood, has been controversial. Recent data indicate that, in the immature rat model, prolonged FSs induce transient structural changes of some hippocampal pyramidal neurons and long-term functional changes of hippocampal circuitry. However, whether these neuroanatomical and electrophysiological changes promote hippocampal excitability and lead to epilepsy has remained unknown. By using in vivo and in vitro approaches, we determined that prolonged hyperthermia-induced seizures in immature rats caused long-term enhanced susceptibility to limbic convulsants that lasted to adulthood. Thus, extensive hippocampal electroencephalographic and behavioral monitoring failed to demonstrate spontaneous seizures in adult rats that had experienced hyperthermic seizures during infancy. However, 100% of animals developed hippocampal seizures after systemic administration of a low dose of kainate, and most progressed to status epilepticus. Conversely, a minority of normothermic and hyperthermic controls had (brief) seizures, none developing status epilepticus. In vitro, spontaneous epileptiform discharges were not observed in hippocampal-entorhinal cortex slices derived from either control or experimental groups. However, Schaeffer collateral stimulation induced prolonged, self-sustaining, status epilepticus-like discharges exclusively in slices from experimental rats. These data indicate that hyperthermic seizures in the immature rat model of FSs do not cause spontaneous limbic seizures during adulthood. However, they reduce thresholds to chemical convulsants in vivo and electrical stimulation in vitro, indicating persistent enhancement of limbic excitability that may facilitate the development of epilepsy. PMID- 10716254 TI - Degeneration of the centre median-parafascicular complex in Parkinson's disease. AB - Two major noncortical inputs to the striatum originate from the substantia nigra and the thalamic centre median-parafascicular complex. Although it is established that in Parkinson's disease there is degeneration of the nigral dopaminergic neurons, there has been little analysis of the glutamatergic centre median parafascicular complex. We therefore evaluated these and neighboring thalamic nuclei (for specificity of any changes) in 9 Parkinson's disease patients and 8 age-matched controls. Degeneration in the substantia nigra and centre median parafascicular complex was estimated by using quantitative neuronal counts. On average, 70% of the pigmented nigral neurons degenerated and there was 30% to 40% neuronal loss in the centre median-parafascicular complex in Parkinson's disease. Thalamic degeneration was marked in neuronal subpopulations (50% loss of parvalbumin-positive neurons in the parafascicular, and 70% loss of non parvalbumin-positive neurons in the centre median nuclei). In contrast, adjacent thalamic nuclei did not degenerate, which supports a selective neurodegeneration of the centre median-parafascicular complex. Our results show that the thalamic centre median-parafascicular complex is an additional nondopaminergic site of neurodegeneration in Parkinson's disease. Because this thalamic region provides important sensorimotor feedback to the striatum, degeneration of this region is likely to exacerbate the clinical signs and symptoms of Parkinson's disease. PMID- 10716255 TI - Specific changes in somatosensory evoked magnetic fields during recovery from sensorimotor stroke. AB - We studied recovery-induced changes in the responsiveness of the primary somatosensory cortex in stroke patients with sensory and/or motor symptoms. Somatosensory evoked magnetic fields, in response to median nerve stimulation, were recorded in 14 patients with their first symptomatic unilateral stroke 1 to 15 days from the first symptoms and again 2 to 3 months later. Neuronal activity at the contralateral primary somatosensory cortex was modeled with equivalent current dipoles at the peak latencies of the first two cortical deflections at about 20 msec (N1m) and at 28 to 91 msec (P1m). Twenty-three age-matched healthy volunteers, 9 of whom were tested also in serial recordings, served as control subjects. At follow-up, 6 patients showed a significant increase of P1m amplitude, whereas N1m increased only in 1. Clinical improvement of two-point discrimination ability, but not of other basic somatosensory skills, was significantly correlated with the increase of P1m. We conclude that the recovery of discriminative touch after stroke is paralleled by the growth of the P1m somatosensory evoked magnetic field deflection, and we propose that this may reflect re-establishment of lateral inhibitory functions at the primary somatosensory cortex. PMID- 10716256 TI - Association of early-onset Alzheimer's disease with an interleukin-1alpha gene polymorphism. AB - Overexpression of the pluripotent cytokine interleukin-1 (IL-1) by microglial cells correlates with formation of neuritic beta-amyloid plaques in Alzheimer's disease (AD). We evaluated polymorphisms in the genes coding for the IL-1alpha, IL-1beta, and IL-1 receptor antagonist cytokines, and tested their association with the occurrence and age at onset of sporadic AD. We found a strong association between the IL-1A T/T genotype and AD onset before 65 years of age (odds ratio, 4.86), with carriers of this genotype showing an onset of disease 9 years earlier than IL-1A C/C carriers. A weaker association with the age at onset was also shown for the IL-1B and IL-1RN genes. These data suggest either a direct effect of the IL-1 gene family, mainly IL-1A, on the clinical onset of AD, or a linkage dysequilibrium with an unknown locus relevant to AD on chromosome 2. PMID- 10716257 TI - Association of interleukin-1 gene polymorphisms with Alzheimer's disease. AB - Interleukin-1 (IL-1) is markedly overexpressed in Alzheimer's disease. We found the IL-1A 2,2 genotype in 12.9% of 232 neuropathologically confirmed Alzheimer's disease patients and 6.6% of 167 controls from four centers in the United Kingdom and United States (odds ratio, 3.0; controlled for age and for ApoE [apolipoprotein E] genotype). Homozygosity for both allele 2 of IL-1A and allele 2 of IL-1B conferred even greater risk (odds ratio, 10.8). IL-1 genotypes may confer risk for Alzheimer's disease through IL-1 overexpression and IL-1-driven neurodegenerative cascades. PMID- 10716259 TI - Corticobasal degeneration shares a common genetic background with progressive supranuclear palsy. AB - Corticobasal degeneration is a sporadic form of tauopathy, involving the cerebral cortex and extrapyramidal motor system. A series of affected subjects was genotyped for a set of genetic markers along the tau protein gene. A specific haplotype is significantly overrepresented in patients versus controls. This haplotype is the same already reported in association with progressive supranuclear palsy. These data show that corticobasal degeneration and progressive supranuclear palsy, in addition to several clinical, pathological, and molecular features, may have the same genetic background. PMID- 10716258 TI - Evaluation of the role of the D2 dopamine receptor in myoclonus dystonia. AB - A novel Val154-->Ile mutation in the D2 dopamine receptor (DRD2) on chromosome 11q23 has recently been shown to be associated with myoclonus dystonia (M-D) in one large family. Sequence analysis of the DRD2 gene in 5 M-D patients from different families did not reveal any mutations, nor was there evidence of linkage to the 11q23 region in the DRD2 gene in four other families. Receptor binding and signal transduction assays of the DRD2 mutant and wild-type receptors revealed identical agonist and antagonist affinities and functional responses. These studies suggest that M-D is genetically heterogeneous. The molecular mechanisms through which the Val-->Ile mutation may contribute to M-D remain to be determined. PMID- 10716260 TI - Sensory discrimination capabilities in patients with focal hand dystonia. AB - To explore the concept that dystonia may result from dysfunction of the sensory system, 14 patients with focal hand dystonia were tested during two somatosensory discrimination tasks. Compared with controls, patients had a higher threshold in a task involving discrimination of two electric stimuli closely related temporally, an abnormality that correlated with the degree of severity of dystonia. There was no significant difference in a single-touch, gross localization task. The possible relevance of these findings to the pathogenesis of dystonia is discussed. PMID- 10716261 TI - Very low levels of the mtDNA A3243G mutation associated with mitochondrial dysfunction in vivo. AB - We studied mitochondrial function in vivo in 2 brothers harboring the mitochondrial DNA A3243G mutation by using magnetic resonance spectroscopy. One brother presented with recurrent strokes and had a mitochondrial respiratory chain complex I defect, with 85% A3243G mutation in his quadriceps. The maximum rate of mitochondrial ATP production in his calf, measured in vivo, was reduced to 21% of the normal mean value. The second brother had mild exercise intolerance, normal muscle histochemistry, and normal respiratory chain activity in vitro. Despite a level of the A3243G mutation of only 5.95% (SD, 4.45; range, 0.7-16.1%) within single muscle fibers from the gastrocnemius muscle, the maximum rate of mitochondrial ATP production in his calf, measured in vivo, was reduced to 35% of the normal mean value. These findings suggest that there may not be a clear genetic threshold level for the expression of the A3243G mutation in skeletal muscle in vivo. PMID- 10716262 TI - Increased bone turnover in epileptic patients treated with carbamazepine. AB - Bone turnover has been investigated in 12 epileptic patients before and after treatment with carbamazepine and in 15 sex- and age-matched control subjects. We found higher values of markers of bone formation (serum bone alkaline phosphatase, osteocalcin, and propeptides of types I and III procollagen) and of bone resorption (serum telopeptide of type I collagen and urine N-telopeptides of type I collagen) in patients than in controls. Our study demonstrates that carbamazepine induces an increase of bone turnover. PMID- 10716263 TI - De novo mutation in the Notch3 gene causing CADASIL. AB - CADASIL, an autosomal dominant arteriopathy responsible for stroke and dementia, is caused by strongly stereotyped mutations in the Notch3 gene. We report a patient with a condition strongly suggestive of CADASIL (migraine, stroke, and white matter abnormalities), except that this patient did not have any first degree relatives with similar symptoms. This patient carried a heterozygous Arg182Cys mutation in the Notch3 gene; this mutation was absent in his two biological parents. These data demonstrate the occurrence of a de novo noninherited mutation in the Notch3 gene, which indicates that CADASIL should not be rejected in the absence of a family history. Therefore, our finding suggests that CADASIL may be more frequent than anticipated. PMID- 10716264 TI - Quantitative pathological evidence for axonal loss in normal appearing white matter in multiple sclerosis. AB - We assessed axonal loss in the normal appearing white matter of the corpus callosum in postmortem brains of patients with multiple sclerosis, using quantitative measures of both axonal density and white matter atrophy. The calculated total number of axons was reduced significantly (mean +/- SD, 5.4 x 10(7) +/- 3.1 x 10(7)) compared with normal controls (11.6 x 10(7) +/- 2.2 x 10(7), p = 0.001) with a reduction both in axonal density (median, 34%; range, 16 56%; p = 0.004) and area (mean +/- SD: multiple sclerosis, 584 +/- 170 mm2; controls, 871 +/- 163 mm2; p = 0.004). These results confirm substantial axonal loss in the normal appearing white matter and demonstrate that measures of both axonal density and white matter volume are necessary to appreciate the full extent of axonal loss. PMID- 10716265 TI - Congenital encephalomyopathy with epilepsy, chorioretinitis, basal ganglia involvement, and muscle minicores. AB - A woman had severe psychomotor retardation, epilepsy, rigidity, and chorioretinitis. Magnetic resonance imaging showed cerebellar and cerebral atrophy and hypointensities in T2-weighted images of the thalami and basal ganglia. Muscle biopsy documented size variations in rounded muscle fibers, fibrosis, and minicores on electron microscopy. Merosin staining was normal. These hitherto unreported features do not permit classification of our patient within the current types of encephalomyopathy and congenital muscular dystrophies. PMID- 10716266 TI - A genetic variation of cathepsin D is a major risk factor for Alzheimer's disease. AB - Cathepsin D (catD) is an intracellular acid protease possibly involved in Alzheimer's disease (AD)-related neurodegeneration through cleavage of amyloid precursor protein into amyloidogenic components. We studied whether an exonic polymorphism of the catD gene (C --> T [Ala --> Val] transition at position 224), which possibly influences pro-catD secretion and intracellular maturation of the enzyme, was associated with the risk for the development of AD in 127 demented patients and 184 controls. The catD*T allele was significantly overrepresented in demented patients (11.8%) compared with nondemented controls (4.9%). Carriers of the catD*T allele had a 3.1-fold increased risk for developing AD than noncarriers. Carriers of the apolipoprotein E (ApoE) epsilon4 allele (ApoE*4) had a 3.9-fold increased risk than non-carriers. The adjusted odds ratio for subjects with the ApoE*4 and the catD*T allele was 19.0 compared with subjects with neither of these two alleles. Our data confirm the results of a recently performed pilot study in an independent sample and suggest that the catD genotype is strongly associated with the risk for AD. PMID- 10716267 TI - William Osler: on Chorea: on Charcot. AB - As the first Professor of Diseases of the Nervous System at the Faculte de Paris, Jean-Martin Charcot was an immensely powerful figure at the end of the 19th century who engendered both wide admiration and resentment. William Osler offers a particularly valuable resource to view Charcot's place in neurology in a relatively unbiased and balanced perspective. Although Osler made numerous seminal neurological contributions, he never considered himself a neurologist, had no formal training with Charcot, and, as a North American, was not tied to the European academic hierarchy of university medicine. One year after Charcot's death, Osler published On Chorea and Choreiform Affectations (1894), and in this pithy monograph, Osler offered a particularly useful evaluation of Charcot's neurological contributions. Whereas in most instances, Osler and Charcot agreed, Osler used data from the new fields of genetics and bacteriology to draw a dear distinction between two entities that Charcot had failed to separate, Sydenham's chorea and Huntington's disease. Osler's On Chorea uniquely captures the transition period between the 19th and 20th centuries. With clarity and insight, Osler documents Charcot's important contributions on disease description, differential diagnosis, and treatment. But with equal sobriety, he delineates Charcot's and his generation's limitation, as the 20th century opens toward the search for neurological causes and embraces new laboratory and experimental methodologies. PMID- 10716268 TI - Multiple sclerosis and Chlamydia pneumoniae. PMID- 10716269 TI - Chlamydia, Rickettsia, and antibiotic treatment of multiple sclerosis. PMID- 10716270 TI - The contribution of HLA to multiple sclerosis susceptibility in Sardinian affected sibling pairs. PMID- 10716271 TI - In vivo visualization of human neural pathways by magnetic resonance imaging. PMID- 10716272 TI - Evaluation of the feasibility of and results of measuring health-status changes in patients undergoing surgical treatment for skeletal metastases. AB - The goal of treating patients with skeletal metastases is to decrease pain and improve or maintain physical function. Assessment of the effectiveness of treatment should therefore include evaluation of patient-rated measures of quality of life. The primary objective of the study was to determine the feasibility of studying the effect of surgical treatment of skeletal metastases on quality of life. The secondary objective was to provide data that begin to characterize this effect. The characteristics of patients with skeletal metastases are heterogeneous, patient enrollment in the study may be low, high attrition occurs secondary to death, and well accepted health-status measures (such as the Short Form-36) may be ineffective at detecting changes in health status; therefore, it is difficult to study these patients. High attrition and adjuvant treatment with radiation or chemotherapy made it impractical to draw firm conclusions about the effect of surgical treatment, but a trend toward improvement in selected health-status measures for both physical and mental health was noted. Analysis of patient-rated health-status scores as predictors of survival indicates that improvement in these scores 6 weeks after surgery is associated with an increase in the length of survival following surgery. PMID- 10716273 TI - Drug-induced apoptosis in osteosarcoma cell lines is mediated by caspase activation independent of CD95-receptor/ligand interaction. AB - Osteosarcoma is one of the most common primary malignant tumors of bone. Treatment of this tumor with systemic chemotherapy dramatically improves the prognosis, although the molecular mechanisms involved in the drug action are poorly understood. In chemosensitive leukaemic T cells and certain solid tumors, cytotoxic drugs mediate the induction of apoptosis by activation of the CD95/APO 1/Fas system. Triggering of the corresponding signaling pathway may involve CD95 receptor/ligand interaction, activation of caspases, or alterations in mitochondrial function. The purpose of our study was to determine if similar mechanisms are involved in the chemosensitivity of osteosarcomas. We found that cytotoxic drugs induce characteristic biochemical and morphological alterations related to apoptosis in osteosarcoma cell lines, including activation of caspases and disturbance of mitochondrial function. However, drug treatment did not result in activation of CD95-receptor or CD95-ligand mRNA. In addition, drug-induced apoptosis was blocked by caspase inhibitors but not by inhibition of CD95-ligand action, indicating a CD95-receptor/ligand-independent mechanism in osteosarcoma cell lines. PMID- 10716274 TI - Effect of age and sampling site on the chondro-osteogenic potential of rabbit marrow-derived mesenchymal progenitor cells. AB - Four sequential bone-marrow aspirations from the ipsilateral tibia and iliac crest of New Zealand White rabbits, aged 4 months or 1, 2, or 3 years, were assayed in vitro and in vivo for their chondro-osteogenic potential. Nonhematopoietic cells from the samples of bone marrow were isolated and expanded in culture; their colony-forming efficiency was determined, and second-passage marrow-derived cells, referred to as mesenchymal progenitor cells, were loaded into porous calcium-phosphate ceramic cubes as carrier vehicles for an in vivo cartilage and bone-formation assay. The cubes were placed subcutaneously in nude BALB/C mice for 3 and 6 weeks. On histological examination, the cubes were scored for the presence of bone and cartilage in their pores, and average values for age groups and location were compared. At aspiration, the samples from the iliac crest exhibited an overall reduction in cell concentration with increasing age, and at the first harvest time, they showed a decrease in colony-forming efficiency and cube score with increasing age. This study demonstrated that repeated bone-marrow aspirations may be performed and may have an enhancing effect on the osteochondral progenitor cells of older animals. PMID- 10716276 TI - Relationship between body mass index and activity in hip or knee arthroplasty patients. AB - The weight of patients has not been demonstrated to have a consistent effect on the rate of polyethylene wear in clinical studies of total joint replacement. For this reason, we analyzed the relationship between quantitative activity, measured with a pedometer, and body mass index, a measure of obesity. Data were acquired for 209 individuals, 22-82 years of age; all were independent community walkers. One hundred and fifty-one had a well functioning total hip or knee replacement. Analysis of variance was used to evaluate the relationship between activity and body mass index, with adjustments for confounding variables. The 58 individuals with no total joint prosthesis averaged 7,781 steps per day, which was higher (p < 0.01) than those with a total hip (5,869 steps per day) or knee (4,597 steps per day) replacement. The subjects with no total joint prosthesis were, however, younger than the patients with a prosthesis (p < 0.01), and the body mass index of the patients with a total knee replacement was higher than that of the patients with a hip replacement and that of the subjects with no prosthesis (p < 0.01). After adjustment for differences in age, gender, and Charnley class, a higher body mass index (greater obesity) was associated with lower activity (p = 0.05). With regard to the rate of polyethylene wear, decreased ambulatory activity may counterbalance increased weight, which could, at least in part, explain why weight has not had a consistent effect on polyethylene wear in clinical studies. Wear is a function of use, not time. The effect of obesity on activity should be considered in radiographic studies of wear and other outcome assessments of total joint replacements. PMID- 10716275 TI - Cellular responses of embryonic hyaline cartilage to experimental wounding in vitro. AB - It is well established that the reparative potential of many tissues is greatest during embryonic development. Despite the extensive literature documenting repair in nonembryonic cartilage models, there is no comparable wealth of experience relating to embryonic cartilage repair. With the embryonic chick sternum as a model of hyaline cartilage, this paper accounts cellular responses and alterations in extracellular matrix composition in response to experimental wounding in vitro. Creation of an experimental lesion induced a rapid (<20 minutes) apoptotic response in chondrocytes adjacent to the lesion edge; the presence of perichondrium delayed this response. Alterations in the extracellular matrix included immediate mechanical damage to type-II collagen fibrils and an increase in the expression of chondroitin-4 sulphate next to the lesion. Creation of the lesion induced an increased proliferative response in chondrocytes behind the zone of apoptosis and the expression of alpha5 and alpha6 integrin subunits. PMID- 10716277 TI - Changes in bone mineral density at the proximal tibia after total knee arthroplasty: a 2-year follow-up of 28 knees using dual energy X-ray absorptiometry. AB - The change in bone mineral density at the proximal tibia during 2 years after total knee arthroplasty was studied in 28 knees (28 patients: 10 men and 18 women; median age: 71 years) with dual energy x-ray absorptiometry. Bone mineral density was measured at the proximal tibia at nine regions of interest below the tibial component within 1 week after the operation (baseline); measurements were repeated at 3, 6, 12, and 24 months. All but one knee was malaligned before the operation, and all but three were corrected to within the normal range of alignment after it. The mean bone mineral density of all nine regions of interest at the proximal tibia temporarily decreased by 13% (p = 0.001) during the initial 3 months, probably due to a general metabolic reaction of the skeleton to the operative trauma combined with the effect of the postoperative immobilization, and then the initial level was regained for as long as 2 years. The overall changes in mean bone mineral density to 2 years were insignificant (p > 0.05); however, a great variation (43.9% decrease to 98.0% increase) was observed on an individual basis. This change over time was significantly associated (R2 = 0.36, p = 0.002) with the level of the baseline bone mineral density, which in turn was partly related (R2 = 0.24, p = 0.009) to the amount of malalignment of the knee before the operation. Knees with high baseline levels (n = 14: 11 with varus and three with valgus alignment) displayed a decrease of 10.0 +/- 14.0% (mean +/- SD, p > 0.05) for as long as 2 years, whereas those with low baseline levels (n = 14: seven with varus and six with valgus alignment and one neutrally aligned) had an increase of 19.1 +/- 38.2% (p = 0.038). In both groups, the mean bone mineral density converged to a level of 0.75-0.95 g/cm2 at 2 years. PMID- 10716278 TI - Controlling the motion of total knee replacements using intercondylar guide surfaces. AB - Total knee replacements using intercondylar cams, such as posterior stabilized types, have been in use for many years. In a previous study, software was written to analyze an alternative shape of the intercondylar cams. The goal of the current study was to investigate in a more general way the potential of intercondylar cams, or guide surfaces, for reproducing the anterior-posterior motion of the natural knee throughout the flexion range. Typical sagittal outlines for the femoral and tibial bearing surfaces were defined, and a parametrized shape for the femoral guide surface was defined to produce a wide range of shapes. Software was written in which the femoral component was flexed in increments, with the posterior translation defined as a function of the flexion angle. The shape of the tibial guide surface was derived from the locus of the femoral guide surface at its multiple flexion positions. By iterating methodically through possible shapes of femoral guide surfaces, several types of total knee replacement components in common use today were identified, as well as other configurations of potential interest. For quantification of a given design, the software calculated the anterior and posterior laxity at each flexion angle. Laxity was defined as the motion before the femoral guide surface impacted the tibial guide surface or until the contact point of the bearing surfaces reached a specified slope. Convex femoral and concave saddle-shaped tibial guide surfaces produced small laxities in both directions over most of the flexion range. A saddle design with small laxities in the first half of flexion, combined with a posterior stabilized feature, was an interesting combination. Potential improvements to the currently used designs were shown in this study, and new shapes of intercondylar guide surfaces were derived that could be considered for application. PMID- 10716279 TI - Comparison of three methods of gluteal muscle attachment to an allograft/endoprosthetic composite in a canine model. AB - This study used radiography, gait analysis, gluteal muscle mass, mechanical testing, and qualitative histology to compare three methods of gluteal muscle attachment to an allograft/endoprosthetic composite of the proximal 25% of the femur in an in vivo canine model. The three methods of gluteal muscle attachment were identical to those used clinically in human patients for hip revision and proximal femoral limb salvage: the host gluteal tendon sutured to the allograft tendon (tendon group), the host greater trochanter with intact gluteal tendons secured to the allograft with a cable-grip system (grip group), and periosteally vascularized proximal femoral bone onlay with intact tendons wrapped around the allograft (wrap group). On the basis of radiographs taken every 2 months, the tendon group had more graft fractures than did the grip or wrap group. Radiographic union of the graft-host bone junction occurred more rapidly and there was less graft resorption in the wrap group than in the other two groups. In all dogs, peak vertical ground-reaction forces in the treated limb decreased immediately after surgery and then slowly increased over the length of the study. The dogs in the wrap group regained normal weight-bearing on the treated limb more quickly than did those in the other groups. The constructs in the tendon group were weaker and less stiff immediately after surgery than were those in the other groups or in intact controls. Histologic analysis confirmed that the wrap technique resulted in complete union of the host bone-allograft junction more often than did the other techniques. The wrap method had the best functional outcome after 9 months when an allograft/endoprosthetic composite was used during total hip arthroplasty in this canine model. PMID- 10716280 TI - Collagen crosslinked N-telopeptides as markers for evaluating particulate osteolysis: a preliminary study. AB - The purpose of this study was to determine whether a marker of bone resorption could be used noninvasively to diagnose and assess treatment of periprosthetic osteolysis. The crosslinked N-telopeptide marker of osteoclast-mediated bone resorption potentially has the sensitivity to detect periprosthetic osteolysis. Second-morning urine specimens were obtained from (a) seven age-matched controls, (b) eight patients who had a hip arthroplasty, hybrid implants at least 1 year after surgery, and no osteolysis, (c) 11 patients who had a hip arthroplasty and osteolysis, and (d) 10 patients who had a hip arthroplasty and with osteolysis before and after 6 weeks of oral Fosamax (alendronate) treatment. The Fosamax treatment consisted of one 10-mg dose per day for 6 weeks. Men and young women (less than 40 years old) were chosen for this study to avoid bone resorption enhanced after menopause as a possible confounder. An enzyme-linked immunosorbent assay technique for quantifying crosslinked N-telopeptides of type-I collagen was performed on all specimens. The patients with osteolysis had significantly elevated levels of N-telopeptide compared with the implant control group. In addition, levels of N-telopeptide were significantly lowered after Fosamax treatment. These findings indicate that a systemic bone-resorption marker (N telopeptide) can be used to evaluate local particulate-induced osteolysis and its treatment. The study also provides clinical evidence that osteolysis is associated with increased osteoclast-mediated bone resorption and that this locally induced bone resorption can be suppressed by certain bisphosphonates (Fosamax). These insights have potential value in the understanding and clinical management of aseptic loosening. PMID- 10716281 TI - Responsiveness of bovine chondrocytes to growth factors in medium with different serum concentrations. AB - Autologous transplantation of chondrocytes is currently under investigation as a potential therapy to stimulate intrinsic repair in articular cartilage defects. The quality of the repair tissue may benefit from the preservation of the characteristic chondrocytic phenotype of the transplanted cells together with the production of a new extracellular matrix composed of collagen type II and larger proteoglycans. A number of growth factors are believed to play an important role in the process of generating new cartilage repair tissue. In this study, the dose dependent response of bovine chondrocytes to recombinant human insulin-like growth factor-1, recombinant human transforming growth factor-beta2, and recombinant human bone morphogenetic protein-2 was studied in an alginate culture system under different culture conditions. The chondrocytes were cultured in medium with increasing concentrations of fetal calf serum. The cultures were assessed by the total amount of DNA, quantitative and qualitative synthesis of proteoglycan, production of nitric oxide, and histology. Cells cultured in the presence of each growth factor had an equal, nonsignificant stimulation of DNA synthesis compared with those cultured in basal medium alone. Recombinant human insulin-like growth factor-1 and recombinant human transforming growth factor beta2 stimulated proteoglycan synthesis in a dose-dependent and reversed dose dependent fashion, respectively. Recombinant human bone morphogenetic protein-2 stimulated proteoglycan synthesis significantly only in the absence of fetal calf serum or in the presence of small amounts of the serum. Overall, proteoglycan synthesis dramatically decreased with the addition of each growth factor as the concentration of fetal calf serum in the medium decreased, and the dose-dependent stimulation pattern, as observed for recombinant human insulin-like growth factor 1 and recombinant human transforming growth factor-beta2, disappeared. Apart from a moderate increase in mRNA for aggrecan and decorin, the growth factors did not greatly affect the type of proteoglycans synthesized. Histological examination confirmed the presence of a dense pericellular matrix deposition, especially when the chondrocytes were cultured in the presence of recombinant human insulin-like growth factor-1 or recombinant human transforming growth factor-beta2. The results indicate that these growth factors can stimulate qualitatively superior matrix production and that the responsiveness of the chondrocytes to the growth factors changes with the culture conditions. Further knowledge about the interaction between chondrocytes, growth factors, and the external environment is important to stimulate chondrocytes to produce adequate repair tissue in cartilage defects in vivo. Insulin-like growth factor-1 especially seems capable of stimulating, in the most consistent and predictable fashion, qualitatively superior proteoglycan synthesis by differentiated chondrocytes. Additional in vivo studies are needed to evaluate the potential of these growth factors as stimulators in cartilage repair. PMID- 10716282 TI - Effect of compressive loading on chondrocyte differentiation in agarose cultures of chick limb-bud cells. AB - It is well established that mechanical loading is important to homeostasis of cartilage tissue, and growing evidence suggests that it influences cartilage differentiation as well. Whereas the effect of mechanical forces on chondrocyte biosynthesis and gene expression has been vigorously investigated, the effect of the mechanical environment on chondrocyte differentiation has received little attention. The long-term objective of this research is to investigate the regulatory role of mechanical loading in cell differentiation. The goal of this study was to determine if mechanical compression could modulate chondrocyte differentiation in vitro. Stage 23/24 chick limb-bud cells, embedded in agarose gel, were subjected to either static (constant 4.5-kPa stress) or cyclic (9.0-kPa peak stress at 0.33 Hz) loading in unconfined compression during the initial phase of commitment to a phenotypic lineage. Compared with nonloaded controls, cyclic compressive loading roughly doubled the number of cartilage nodules and the amount of sulfate incorporation on day 8, whereas static compression had little effect on these two measures. Neither compression protocol significantly affected overall cell viability or the proliferation of cells within nodules. Since limb-bud mesenchymal cells were seeded directly into agarose, an assessment of cartilage nodules in the agarose reflects the proportion of the original cells that had given rise to chondrocytes. Thus, the results indicate that about twice as many mesenchymal cells were induced to enter the chondrogenic pathway by cyclic mechanical compression. The coincidence of the increase in sulfate incorporation and nodule density indicates that the primary effect of mechanical compression on mesenchymal cells was on cellular differentiation and not on their subsequent metabolism. Further studies are needed to identify the primary chondrogenic signal associated with cyclic compressive loading and to determine the mechanism by which it influences commitment to or progression through the chondrogenic lineage, or both. PMID- 10716283 TI - Microstructural properties of the distal growth plate of the rabbit radius and ulna: biomechanical, biochemical, and morphological studies. AB - The purposes of this study were to define the tensile properties of each zone of the rabbit growth plate and to correlate them with the microarchitecture and biochemical composition of the zones. The epiphysis-growth plate-metaphysis complex was obtained from the radius and ulna of 20 8-week-old rabbits. Four dye markers were placed on the growth plate. The complex was loaded to failure with a tensile testing machine, and the strain behavior was recorded simultaneously with a microscope, a charge-coupled device camera, and a video dimension-analyzer system. The collagenous fiber architecture of each zone was examined with a microscope, and the collagen content of each zone was also determined. The tangent modulus of the resting zone was 75% stiffer than that of the other two zones. The highest values for strain at failure and energy absorbed to failure were observed in the hypertrophic zone, and the total collagen content was highest in the proliferating zone. The collagen fibers were more randomly aligned in the resting zone than in the other two zones. The diversity observed in the microarchitecture of the rabbit growth plate correlates with the zone-dependent differences in its mechanical properties. PMID- 10716284 TI - Stiffness, viscosity, and upper-limb inertia about the glenohumeral abduction axis. AB - To evaluate the dynamic properties of the shoulder and understand how they are controlled by the central nervous system, glenohumeral-joint stiffness and viscosity and upper-limb inertia were quantified under various levels of muscle contraction in seven healthy human subjects. Through a cast attachment, the upper limb was perturbed in a precise pattern by a computer-controlled servomotor to manifest the dynamic properties of the joint. The recorded joint position and torque were used to estimate joint stiffness and viscosity and upper-limb inertia. With moderate muscle contraction, the stiffness and viscosity increased several fold. A stiffer shoulder joint associated with stronger muscle contraction made the shoulder more stable and protected it from potential injuries during strenuous tasks. Joint viscosity, especially the stronger viscous damping associated with more strenuous contraction, smoothed shoulder movement and stabilized the joint. From the control viewpoint, the glenohumeral joint responded to the central nervous system more quickly with increasing muscle contraction, which was useful during strenuous tasks. On the other hand, the central nervous system controlled stiffness and viscosity synchronously so that it dealt with only a nearly constant damping ratio of the joint over various levels of contraction, which simplified its task substantially. This approach quantified the dynamic and static properties of the shoulder under various levels of contraction more accurately and completely than a manual test, and it can potentially be used to evaluate changes in these properties caused by musculoskeletal injuries and their surgical treatments. PMID- 10716285 TI - Hamstrings and iliotibial band forces affect knee kinematics and contact pattern. AB - Many clinical studies have emphasized the role of the hamstrings and the iliotibial band on knee mechanics, although few biomechanical studies have investigated it. This study therefore examined two hypotheses: (a) with loading of the hamstrings, the tibia translates posteriorly and rotates externally and the tibial contact pattern shifts anteriorly; furthermore, the changes in tibial kinematics alter patellar kinematics and contact; and (b) loading the iliotibial band alters the kinematics and contact pattern of the tibiofemoral joint similarly to loading the hamstrings, and loading the iliotibial band laterally translates the patella and its contact location. Five cadaveric knee specimens were tested with a specially designed knee-joint testing machine in an open-chain configuration. At various flexion angles, the knees were tested always with a quadriceps force but with and without a hamstrings force and with and without an iliotibial band force. The results support the first hypothesis. Hence, the hamstrings may be important anterior and rotational stabilizers of the tibia, a role similar to that of the anterior cruciate ligament. The results also support the second hypothesis, although the iliotibial band force had a smaller effect on the tibia than did the hamstrings force. Both forces also changed patellar kinematics and contact, demonstrating that these structures should also be considered during the clinical management of patellar disorders. PMID- 10716286 TI - Importance of the medial meniscus in the anterior cruciate ligament-deficient knee. AB - The incidence of meniscal tears in the chronically anterior cruciate ligament deficient knee is increased, particularly in the medial meniscus because it performs an important function in limiting knee motion. We evaluated the role of the medial meniscus in stabilizing the anterior cruciate ligament-deficient knee and hypothesized that the resultant force in the meniscus is significantly elevated in the anterior cruciate ligament-deficient knee. To test this hypothesis, we employed a robotic/universal force-moment sensor testing system to determine the increase in the resultant force in the human medial meniscus in response to an anterior tibial load following transection of the anterior cruciate ligament. We also measured changes in the kinematics of the knee in multiple degrees of freedom following medial meniscectomy in the anterior cruciate ligament-deficient knee. In response to a 134-N anterior tibial load, the resultant force in the medial meniscus of the anterior cruciate ligament deficient knee increased significantly compared with that in the meniscus of the intact knee; it increased by a minimum of 10.1 N (52%) at full knee extension to a maximum of 50.2 N (197%) at 60 degrees of flexion. Medial meniscectomy in the anterior cruciate ligament-deficient knee also caused a significant increase in anterior tibial translation in response to the anterior tibial load, ranging from an increase of 2.2 mm at full knee extension to 5.8 mm at 60 degrees of flexion. Conversely, coupled internal tibial rotation in response to the load decreased significantly, ranging from a decrease of 2.5 degrees at 15 degrees of knee flexion to 4.7 degrees at 60 degrees of flexion. Our data confirm the hypothesis that the resultant force in the medial meniscus is significantly greater in the anterior cruciate ligament-deficient knee than in the intact knee when the knee is subjected to anterior tibial loads. This indicates that the demand on the medial meniscus in resisting anterior tibial loads is increased in the anterior cruciate ligament-deficient knee compared with in the intact knee, suggesting a mechanism for the increased incidence of medial meniscal tears observed in chronically anterior cruciate ligament-deficient patients. The large changes in kinematics due to medial meniscectomy in the anterior cruciate ligament-deficient knee confirm the important role of the medial meniscus in controlling knee stability. These findings suggest that the reduction of resultant force in the meniscus may be a further motive for reconstructing the anterior cruciate ligament, with the goal of preserving meniscal integrity. PMID- 10716287 TI - An in vivo model for load-modulated remodeling in the rabbit flexor tendon. AB - This study tested the hypothesis that eliminating in vivo compression to the wrap around, fibrocartilage-rich zone of the flexor digitorum profundus tendon results in rapid depletion of fibrocartilage and changes in its mechanical properties, microstructure, extracellular matrix composition, and cellularity. The right flexor digitorum profundus tendons of 2.5-3-year-old rabbits were translocated anteriorly to eliminate in vivo compression and shear to the fibrocartilage zone and, at 4 weeks after surgery, were compared with tendons that had sham surgery and with untreated tendons. The translocated tissue showed a significant increase in equilibrium strain under a compressive creep load (p < 0.05). The thickness and area of the fibrocartilage zone also decreased significantly (p < 0.05). The nuclear density decreased by 40% in the fibrocartilage zone (p < 0.005); however, nuclear shape and orientation were not significantly altered. Glycosaminoglycan content in the fibrocartilage zone was also depleted by 40% (p < 0.02). The tightly woven basket weave-like mesh of collagen fibers in the zone appeared more loosely organized, suggesting matrix reorganization due to translocation. Moreover, immunoreactive type-II collagen and link protein in the fibrocartilage zone also decreased. With use of this unique in vivo model, this research clearly elucidates how changing tissue function (by removing compressive forces) rapidly alters tissue form. PMID- 10716288 TI - Biomechanical evaluation of early fracture healing in normal and diabetic rats. AB - Diabetes mellitus has been shown to alter the properties of bone and impair fracture healing in both humans and animals. The objective of this study was to document changes in the structural and material properties of intact bone and bone with healed fractures in diabetic rats compared with nondiabetic controls after 3 and 4 weeks of healing. Rods were inserted in the right femurs of control rats and rats with streptozotocin-induced diabetes, and the femurs were fractured in a standardized procedure and then allowed to heal for 3 and 4 weeks. After death, all femurs were mechanically tested to failure in torsion. The degree of healing was quantified for each animal by normalizing mechanical parameters for the femur with a healed fracture with those for the intact contralateral femur. At both time points of healing, diabetic rats exhibited inferior healing compared with that of control animals in terms of failure torque, failure stress, structural stiffness, and material stiffness of the femur with the healed fracture relative to the intact contralateral femur (p < 0.05). Our results demonstrate that the recovery of structural and material strength in femurs with healed fractures in diabetic rats is delayed by at least 1 week compared with that in controls. PMID- 10716289 TI - Osteogenic growth peptide normally stimulated by blood loss and marrow ablation has local and systemic effects on fracture healing in rats. AB - Osteogenic growth peptide, a histone H4-related, 14-amino-acid peptide, is an active mediator of local, as well as systemic, osteogenic activity in response to marrow ablation, trauma, and blood loss. In this study, the effect of exogenous osteogenic growth peptide on the healing of femoral fractures in rats was investigated. A fracture at the midshaft of the femur was created in 50 rats. Half of the rats were injected subcutaneously with 25 ng of osteogenic growth peptide per rat per day for the first 7 days after fracture. Radiographs were taken each week, and the diameter of the callus was measured. The femurs of four animals from each group were harvested 1, 2, 3, and 4 weeks after fracture. Two femurs from each group were sectioned for histologic examination, and two were sectioned for measurement of density and mineral content. Marrow was aspirated from the contralateral femurs to establish adhering cell cultures, which were examined for osteogenicity. At 2 weeks, a large increase in mitogenicity and osteogenicity was seen in the marrow-derived cultures from the rats treated with osteogenic growth peptide; this increase was sustained through 4 weeks. Extraction of RNA from the contralateral marrow (systemic expression) and callus (local expression) for amplification with reverse transcription-polymerase chain reaction revealed greater systemic expression of transforming growth factors beta1, beta2, and beta3, fibroblast growth factor-2, insulin-like growth factor 1, and aggrecan throughout the 4 weeks after fracture, whereas types IIA and IIB collagen, link protein, and fibroblast growth factor receptor-3 had a greater local expression. The specimens treated with osteogenic growth peptide had a stronger expression of transforming growth factor-beta1, both locally and systemically. The average diameter of the callus was greater for the treated rats at all time intervals, and peak diameters were 7.58 mm at 3 weeks for the treated rats and 6.64 mm at 2 weeks and 6.63 mm at 3 weeks for the controls. Histological study revealed an earlier organization and faster healing of the treated fractures, as evidenced by the larger, earlier appearance of cartilaginous soft callus and the more rapid organization of bridging trabecular bone. No statistical significance was obtained when these comparisons were made between the groups. These results suggest that osteogenic growth peptide can be used to promote earlier proliferation and differentiation of osteogenic cells in marrow and bone-repair callus, possibly through its effect on the transforming growth factor-beta family. PMID- 10716290 TI - Assessment of resorbable bioactive material for grafting of critical-size cancellous defects. AB - Bioactive glasses form a surface apatite layer in vivo that enhances the formation and attachment of bone. Sol-gel Bioglass graft material provides greater nanoscale porosity than bioactive glass (on the order of 50-200 A), greater particle surface area, and improved resorbability, while maintaining bioactivity. This study histologically and biomechanically evaluated, in a rabbit model, bone formed within critical-sized distal femoral cancellous bone defects filled with 45S5 Bioglass particulates, 77S sol-gel Bioglass, or 58S sol-gel Bioglass and compared the bone in these defects with normal, intact, untreated cancellous bone and with unfilled defects at 4, 8, and 12 weeks. All grafted defects had more bone within the area than did unfilled controls (p < 0.05). The percentage of bone within the defect was significantly greater for the 45S5 material than for the 58S or 77S material at 4 and 8 weeks (p < 0.05), yet by 12 weeks equivalent amounts of bone were observed for all materials. By 12 weeks, all grafted defects were equivalent to the normal untreated bone. The resorption of 77S and 58S particles was significantly greater than that of 45S5 particles (p < 0.05). Mechanically, the grafted defects had compressive stiffness equivalent to that of normal bone at 4 and 8 weeks. At 12 weeks, 45S5-grafted defects had significantly greater stiffness (p < 0.05). At 8 and 12 weeks, all grafted defects had significantly greater stiffness than unfilled control defects (p < 0.05). In general, the 45S5-filled defects exhibited greater early bone ingrowth than did those filled with 58S or 77S. However, by 12 weeks, the bone ingrowth in each defect was equivalent to each other and to normal bone. The 58S and 77S materials resorbed faster than the 45S5 materials. Mechanically, the compressive characteristics of all grafted defects were equivalent or greater than those of normal bone at all time points. PMID- 10716291 TI - Changes in conduction, blood flow, histology, and neurological status following acute nerve-stretch injury induced by femoral lengthening. AB - The effects of an acute stretch on evoked potential, blood flow, histological change, and clinical neurological state were studied in a rat model of acute nerve stretch induced by femoral lengthening. The purposes of this study were to assess, in a model of acute limb lengthening, the safe limits of nerve stretch for nerve function, the pathogenesis of nerve dysfunction, the sensitivity of spinal somatosensory evoked potential, and one of the proposed criteria for irreversible compromise of the sciatic nerve. Thirty-two rats were assigned to one of four groups defined by the degree of acute femoral lengthening (8, 16, 24, and 32%). Spinal somatosensory evoked potential at L5/6 following stimulation of the sciatic nerve was recorded before, immediately after, and 30 minutes after lengthening. Sciatic nerve blood flow was measured by laser Doppler flowmetry at the stretched site before and after lengthening. One week after the operation and without further lengthening, clinical neurological status was evaluated by the functional index of the sciatic nerve and histological examination was performed. At the measurement immediately after the procedure, amplitude changed significantly in all groups except for the group with 8% lengthening. In all groups, sciatic nerve blood flow also dropped significantly compared with values for the control side. Moreover, a greater percentage increase in acute lengthening corresponded with more marked changes in spinal somatosensory evoked potential and sciatic nerve blood flow. The groups that underwent acute lengthening of 24 and 32% had significant neurological deficits and histological changes and demonstrated a significant and profound (50%) drop in amplitude and blood flow. We concluded that spinal somatosensory evoked potential is very sensitive and may serve as an effective tool for the early detection of impending acute nerve-stretch injury and that a 50% reduction in amplitude indicates irreversible damage. PMID- 10716293 TI - Re: Quantitative determination of joint incongruity and pressure distribution during simulated gait and cartilage thickness in the human hip joint. PMID- 10716292 TI - Role of alpha-1 adrenoceptor subtypes mediating constriction of the rabbit ear thermoregulatory microvasculature. AB - An acute in vivo preparation of the microvasculature of the rabbit ear was used to evaluate the functional role of alpha1 (alpha1)-adrenoceptor subtypes in thermoregulatory microcirculation. The effect of alpha1-adrenoceptor subtype blockade on phenylephrine-induced vasoconstriction was assessed with the alpha1A, alpha1B, and alpha1D-adrenoceptor-selective antagonists 5-methyl-urapidil (10(-8) M), chloroethylclonidine (10(-5) M), and 8-[2-[4(2-methoxyphenyl)-1 piperazinyl]ethyl]-8-azaspirol[4.5]deca ne-7,9-dione dihydrochloride (BMY7378) (10(-6) M), respectively. The results demonstrated that pretreatment of the ear microvasculature with 5-methyl-urapidil or BMY7378 shifted the phenylephrine concentration-response curve rightward and significantly changed the log of the phenylephrine concentration, causing half-maximum stimulation (EC50) in arterioles (p < 0.05). BMY7378 shifted the phenylephrine concentration-response curve of the arteriovenous anastomoses about 100-fold rightward (p < 0.05). All three alpha1-adrenoceptor antagonists eliminated the vasoconstrictive effects of phenylephrine on venules. The results indicate that the ear microvasculature has a heterogenous distribution of alpha1-adrenoceptor subtypes. The alpha1A and alpha1D-adrenoceptor subtypes appear to have a greater influence on constrictive function in arterioles, whereas the alpha1D-adrenoceptor is the dominant constrictor of arteriovenous anastomoses. In general, the alpha1-adrenoceptor does not play a major vasoconstrictor role in venules. Chloroethylclonidine, an irreversible alpha1B-adrenoceptor antagonist, induced contractile responses in the ear microvasculature, probably due to its alpha2-adrenoceptor agonist effects. This study extended our understanding of the adrenergic receptor control mechanisms of a cutaneous thermoregulatory end organ characterized by two parallel perfusion circuits providing nutritional and thermoregulatory functions. PMID- 10716294 TI - Umbilical cord knots. AB - BACKGROUND: Umbilical cord knots may represent a hazard to the fetus, particularly as regards intrauterine death and fetal distress or asphyxia in labor. The object of this study was to analyze the impact of associated umbilical cord encirclements and cord length on fetal outcome and fetal weight deviation. METHODS: Among 22,012 births occurring in Akershus Central Hospital, there were 216 instances of umbilical cord knots. Fetal outcome, fetal weight deviation, associated umbilical cord encirclements and cord length were assessed. RESULTS: Neonates with a knotted cord are more often large-for-gestational age compared to other babies, and have longer umbilical cords. There is a 10 times higher chance of intrauterine fetal death with a knotted cord, but if this does not occur then there is no increased risk of obstetrical intervention and Apgar scores are the same as in other babies and other fetuses. CONCLUSION: There is an association between umbilical cord knots and umbilical cord encirclements. Knotting of the cord is not by itself lethal. Pregnancies with knotted cords have characteristics different from those with ordinary umbilical cord encirclements. PMID- 10716295 TI - Increased risk of preterm delivery with elevated maternal alpha-fetoprotein and plasma zinc levels in African-American women. AB - BACKGROUND: This study evaluated the relationship of maternal serum alpha fetoprotein (MSAFP) and plasma zinc levels (PZn) to pregnancy outcome. METHODS: The subjects for this investigation consisted of 917 African-American women, who on registration for prenatal care between 7-22 weeks gestational age (GA), had PZn levels determined and also had MSAFP recorded in their charts. RESULTS: MSAFP levels greater than the 90th percentile significantly increased the risk of PTD (adjusted odds ratio or AOR=2.5, 95% C.I.=1.5-4.2) but not of IUGR. There was no significant relationship between maternal PZn level and PTD or IUGR. When subjects were stratified by MSAFP levels, in women with MSAFP greater than the 90th percentile, the AOR for PTD was 4.0 (95% C.I.=1.2-13.5) for women with PZn levels greater than the median vs. those with PZn equal to or less than the median. In women with MSAFP equal to or less than the 90th percentile, there was no such difference. Multiple regression analyses, using GA at birth as the dependent variable, indicated an interaction between MSAFP and PZn levels. CONCLUSION: In this population, the adverse pregnancy outcome associated with elevated MSAFP was seen only in women with PZn levels greater than the median. The reason for this association is not currently apparent. PMID- 10716296 TI - Longitudinal distance standards of fetal growth. Intrauterine and Infant Longitudinal Growth Study: IILGS. AB - BACKGROUND: Most ultrasonographic fetal growth norms are derived from cross sectional data or from longitudinal data treated as coming from cross-sectional studies, although only longitudinal models may detect particular aspects of fetal growth shape, such as peak of growth velocity. MATERIALS AND METHODS: The sample included 238 singleton normal pregnancies. All the fetal traits under study (biparietal diameter, occipito-frontal diameter, head circumference, femur diaphysis length and abdomen circumference) were measured according to the classical ultrasound techniques by highly trained operators. Individual growth profiles (made up of 5 to 9 measures) were taken at regular intervals between the 12th and the 40th week. Growth norms were traced by means of a two-stage linear model: (I) a 3-constant fetal growth function was fitted to each individual growth profile, (II) growth centiles were based upon the weighted mean and covariance matrix of the individual growth constants. RESULTS: Fetal growth curves show a sigmoid shape with a maximum slope (i.e. a peak growth velocity) which occurs earlier for head diameters (about 18 weeks), later for femur diaphysis length (20 weeks) and abdomen circumference (22 weeks). During intrauterine growth, all traits show a progressive increase in interindividual variability, which is more prominent for abdomen circumference. CONCLUSION: The mathematical model applied to a large sample of growth profiles provided a satisfactory description of the individual fetal development and its biological variability, and allowed the construction of longitudinal distance standards useful for clinical purposes. PMID- 10716297 TI - Oxytocin-induced oscillations of cytoplasmic Ca2+ in human myometrial cells. AB - BACKGROUND: To investigate the mechanisms of oxytocin (OT) induced oscillations of the cytoplasmic Ca2+ concentration ([Ca2+]i) in cultured human myometrial cells. METHODS: [Ca2+]i was measured in individual myometrial cells by dual wavelength spectrophotofluorometry using the fluorescent indicator fura-2. Myometrium was obtained at abdominal hysterectomy (n=8) and during cesarean section (n=7). RESULTS: OT (10-300 nM) typically induced [Ca2+]i oscillations with frequencies in the 0.6-0.8/min range. There were no obvious differences in the responses of cells taken from non-pregnant and term pregnant women. The frequency and amplitude of the oscillations were not significantly affected by OT concentrations up to 300 nM. The amplitude of the oscillations decreased in the presence of the voltage-dependent Ca2+ channel antagonist verapamil and gradually disappeared in Ca2+-free medium. The oscillations were further blocked by the inorganic Ca2+ antagonist La3+ and by the intracellular Ca2+-ATPase inhibitor 2.5 di-tert-butylhydroquinone (DTBHQ). Caffeine inhibited the OT-induced oscillations in a concentration-dependent manner. DTBHQ and high concentrations of OT made [Ca2+]i remarkably sensitive to changes in the external Ca2+ concentration. CONCLUSIONS: The results indicate that OT-induced [Ca2+]i oscillations in human myometrial cells are due to inositol 1,4,5-trisphosphate-mediated release of intracellular Ca2+ combined with capacitative as well as voltage-dependent influx of the ion. PMID- 10716298 TI - Effects of electro-acupuncture on anovulation in women with polycystic ovary syndrome. AB - BACKGROUND: The present study was designed to evaluate if electro-acupuncture (EA) could affect oligo-/anovulation and related endocrine and neuroendocrine parameters in women with polycystic ovary syndrome (PCOS). METHODS: Twenty-four women (between the ages of 24 and 40 years) with PCOS and oligo-/amenorrhea were included in this non-randomized, longitudinal, prospective study. The study period was defined as the period extending from 3 months before the first EA treatment, to 3 months after the last EA treatment (10-14 treatments), in total 8 9 months. The menstrual and ovulation patterns were confirmed by recording of vaginal bleedings and by daily registrations of the basal body temperature (BBT). Blood samples were collected within a week before the first EA, within a week after the last EA and 3 months after EA. RESULTS: Nine women (38%) experienced a good effect. They displayed a mean of 0.66 ovulations/woman and month in the period during and after the EA period compared to a mean of 0.15 before the EA period (p=0.004). Before EA, women with a good effect had a significantly lower body-mass index (BMI) (p<0.001), waist-to-hip circumference ratio (WHR) (p=0.0058), serum testosterone concentration (p=0.0098), serum testosterone/sex hormone binding globulin (SHBG) ratio (p=0.011) and serum basal insulin concentration (p=0.0054), and a significantly higher concentration of serum SHBG (p=0.040) than did those women with no effect. CONCLUSION: Repeated EA treatments induce regular ovulations in more than one third of the women with PCOS. The group of women with good effect had a less androgenic hormonal profile before treatment and a less pronounced metabolic disturbance compared with the group with no effect. For this selected group EA offers an alternative to pharmacological ovulation induction. PMID- 10716299 TI - DNA strand breaks in human spermatozoa: correlation with fertilization in vitro in oligozoospermic men and in men with unexplained infertility. AB - BACKGROUND: The purpose of this study was to examine the correlation between DNA strand breaks in human spermatozoa and semen quality, fertilization rate and IVF outcome. METHODS: A total of 50 men suffering from unexplained infertility and 50 men with oligozoospermia undergoing IVF treatment entered a prospective study. Sperm samples were assessed according to the WHO manual and for the presence of DNA strand breaks in spermatozoa. The study was blinded for the technician involved in assessment of DNA strand breaks. IVF was carried out according to a long down regulation protocol using GnRH, FSH and hCG. The ova were inseminated with 200,000 spermatozoa/ml. Embryos were transferred on day 2 after fertilization with a maximum of three embryos. RESULTS: This study demonstrates a negative correlation between the proportion of spermatozoa having DNA strand breaks and the proportion of oocytes fertilized after IVF. CONCLUSION: The number of human spermatozoa with DNA strand breaks is a good predictor for fertilization rate in couples suffering from unexplained infertility and undergoing IVF treatment. PMID- 10716300 TI - Use of hormone replacement therapy among Danish nurses in 1993. AB - BACKGROUND: To describe the prevalence of women using systemic hormone replacement therapy in various age groups. To identify their reasons for choosing or not choosing the therapy, reasons for discontinuing the treatment, the prevalence of side effects among current users, and to estimate the duration of treatment. METHODS: The study is based on postal questionnaires sent to 23,000 female Danish nurses above the age of 44 years. Out of these 19,953 (86%) responded. The questionnaire gave information on age, use of hormone replacement therapy, use of oral contraceptives, family predisposition and diseases. Duration of hormone replacement therapy was calculated by Cox regression analysis. Chi square tests were used to evaluate differences and 5% was used as the level of significance. RESULTS: Overall, 6673 (33%) had ever used hormone replacement therapy. The prevalence was highest in the age group 55-59, where 29.3% were currently using hormones. The most cited reasons for choosing hormone replacement therapy were vasomotor symptoms (62%) and prevention of osteoporosis (44%). Among never users 43% had not experienced climacteric symptoms, 24% found the therapy unnatural, and 22% were afraid of side effects. It was estimated that 70% still were using hormones five years after the start of therapy, 57% after ten years, and 48% after fifteen years. Women with a family history of osteoporosis used hormones longer than women without this predisposition. CONCLUSIONS: One third of all the women had ever used hormone replacement therapy and more than half of ever users used the therapy for more than ten years. PMID- 10716301 TI - Transvaginal ultrasonographic findings in the uterus and the endometrium: low prevalence of leiomyoma in a random sample of women age 25-40 years. AB - BACKGROUND: In articles and textbooks the prevalence of uterine leiomyomas is said to be 20-25% in women over the age of 30. The aim of this study was to investigate the rate of uterine leiomyoma, the thickness and the texture of the endometrium, and the size of the uterus in a random sample of asymptomatic women 25-40 years old. METHODS: A random sample of women 25-40 years old was offered a transvaginal ultrasonographic examination and 335 (72%) accepted the invitation. RESULTS: In 18 women uterine leiomyomas were detected, i.e. 5.4% (95% CI 3.0 7.8%). The prevalence of leiomyomas increased with age, being 3.3% (95% CI 0.7 6.0%) in the 25-32 years age group and 7.8% (95% CI 3.6-12.0%) in the 33-40 age group. The size of the uterus correlated to parity, age and height. In women on combined oral contraceptives the size of the uterus was smaller than in women with natural cycles. The size of the uterus did not correlate to body mass index, cycle day or smoking habits. The endometrium increased in thickness and had in most cases a triple line appearance during the proliferative phase until day 15, whereafter it was unchanged in thickness throughout the secretory phase and hyperechogenic in appearance. CONCLUSIONS: This study confirms earlier studies on the endometrium based on selected populations. The size of the uterus increased with parity, age and height, and was smaller in combined oral contraceptive users. The prevalence figures for uterine leiomyomas in textbooks are not confirmed. PMID- 10716302 TI - Determinants of urinary incontinence in a population of young and middle-aged women. AB - BACKGROUND: Urinary incontinence and genital prolapse are prevalent conditions in the female population. The aim of this study was to study possible determinants of female urinary incontinence in a population-based sample of young and middle aged women. METHODS: Of 641 eligible women aged 20-59 years in a primary health care district, 487 (76%) responded to a questionnaire and accepted an invitation to a gynecological examination. The examination included digital assessment of the pelvic floor muscle strength (PFMS). Genital prolapse presence (cystocele, rectocele, uterine prolapse or absence of the urethrovesical crease) was graded in relation to the vaginal introitus. RESULTS: The prevalence of urinary incontinence was 28%, 3.5% having daily leakage. Stress urinary incontinence was the dominant type. The odds ratio (OR) of having incontinence increased from 1 to 3.5 with increasing age and from 1 to 2.7 with increasing parity. The OR also increased with decreasing PFMS; from 1 in the group with the best PFMS to 3.4 in the group unable to contract their pelvic musculature. In addition, women with cystocele and/or absence of the urethrovesical crease had a 2.5-fold increased OR of incontinence (95% CI 1.5-4.2), smoking increased the OR 1.9 times (95% CI 1.1 3.2) and estrogen replacement therapy (ERT) increased the OR 2.9 times (95% CI 1.4-5.9). There were no significant correlations with the presence of chronic disease, episiotomy or the birth weights of children but small non-significant correlations with performed hysterectomy and the woman's weight. CONCLUSIONS: Urinary incontinence is a frequent symptom in the female general population and related to age, pelvic floor muscle strength, genital prolapse, smoking, parity and estrogen replacement therapy. PMID- 10716303 TI - Aggressive angiomyxoma in females: is radical resection the only option? AB - BACKGROUND: Aggressive angiomyxoma is a rare mesenchymal tumor, characterized by frequent local recurrences. Our aim is to assess the role of radical resection. METHODS: Retrospective case review. The records of five patients with aggressive angiomyxoma during the period from 1984 to 1998 were reviewed and analyzed. A MEDLINE search from 1983 to May 1999 was performed. The clinical presentation, surgical treatment, resection margin involvement and clinical outcomes were analyzed. RESULTS: Together with our five cases, 106 cases have been reported in the world literature. The female-to-male ratio was 6.6:1. The age distribution was wide, with the peak incidence at 31 to 35. The local recurrence rate was high. Seventy-one percent of recurrence occurred within the first 3 years. Patients with clear resection margins have similar chances of remaining disease free compared with those having tumor-involved resection margins. There was no correlation between the size of the tumors and the chance of recurrence. CONCLUSIONS: Though we aim for complete resection, incomplete or partial resection is acceptable, especially when high operative morbidity is anticipated and preservation of fertility is an issue. Long-term follow-up and careful monitoring with imaging techniques are essential for timely identification of recurrence and prompt resection. PMID- 10716304 TI - The pap-smear history of women with invasive cervical squamous carcinoma. A case control study from Sweden. AB - BACKGROUND: Screening by cytology is a potentially highly effective procedure for preventing carcinoma of the uterine cervix. To elucidate any weaknesses in the screening procedure in a Swedish county where screening started many years ago, the detection of invasive cervical squamous cell carcinoma was compared to the prior cytological screening. METHODS: On the basis of the complete Pathology data files, including cytology and histology, all 112 women with invasive cervical squamous carcinoma were compared to 112 matched controls from the Swedish Population Register, regarding attendance rate and results of Pap-smears prior to the date of discovery of an invasive carcinoma in the case. RESULTS: Almost as many cases as controls had a history of pap-smear testing, but the cases had significantly more prior atypias registered. Only 16% of women with cervical carcinoma and younger than 60 years were lacking Pap-smear tests prior to the carcinoma diagnosis, but 46% had former atypias registered. More than half of them presented, however, a negative Pap-smear test less than three years before the diagnosis. Among the controls, 10% were lacking prior Pap-smears and only 9% had former atypias registered. CONCLUSION: The policy for follow-up and treatment of cervical dysplasias has to be improved in order to achieve a further reduction of the incidence of invasive carcinoma. PMID- 10716305 TI - Stratified rates of cesarean sections and spontaneous vaginal deliveries. Data from five labor wards in Denmark--1996. AB - BACKGROUND: A fundamental point when auditing labor management is to ensure present and stratified process data. METHOD: Stratification of deliveries into ten mutually exclusive groups enabled comparisons of rates of cesarean sections and rates of spontaneous vaginal deliveries between labor wards. RESULTS: Data from five labor wards in Denmark in 1996 were included in the study comprising a total of 11,287 women. The overall cesarean section rates were between 13.2 and 15.2% which was not a significant difference, whereas cesarean section rates in several of the ten groups and the rates of spontaneous vaginal delivery in group 1 and 3 were significantly different between the labor wards. DISCUSSION: The method presented here is simple and can be used as an integrated part of the daily work and quality assurance. We advocate that stratification of the delivering women into ten groups should take place in every labor ward with focus on both the cesarean section rate and the rate of spontaneous vaginal delivery. Stratification provides data for periodical evaluation of the outcome within a department and for comparison between departments with different populations and policy. PMID- 10716306 TI - Aggressive angiomyxoma of the vagina. Report of a distinctive type gynecologic soft tissue neoplasm. PMID- 10716307 TI - A case-control teratological study of spiramycin, roxithromycin, oleandomycin and josamycin. PMID- 10716308 TI - Relationship between resting 201Tl reverse redistribution, microvascular perfusion, and functional recovery in acute myocardial infarction. AB - 201TI reverse redistribution is a common finding early after reperfusion therapy for myocardial infarction. Its mechanism and clinical implications remain unclear. The aim of this study was to clarify the relationships between reverse redistribution, microvascular perfusion, and myocardial viability. METHODS: Resting, 10-min-postinjection, and redistribution 201TI data obtained for 33 patients 8 and 42 d after the onset of acute myocardial infarction were compared with echocardiographic wall motion measured acutely and on day 42. Microvascular perfusion was assessed by myocardial contrast echocardiography performed 10 min after restoration of complete patency of the infarct artery. RESULTS: Marked significant reverse redistribution was found on day 8 (absolute change, 7.5%+/ 7.9% of the 10-min-postinjection defect size; P<5x0.000001) and significantly decreased on day 42 (2.7%+/-6.8%; P = 0.004 between days 8 and 42). The 10-min postinjection defect size best predicted the final infarct size on day 42 and was closely related to microvascular perfusion. Patients with adequate reperfusion had a smaller postinjection defect on day 8 (21.1%+/-14.6%) and a larger reverse redistribution (10.2%+/-6.1%) than did patients with no reflow (35.3%+/-13% and 3.2%+/-9.2%, respectively; P<0.04 for both). CONCLUSION: Reverse redistribution was marked early after myocardial infarction in patients with complete patency of the infarct artery and decreased in subsequent weeks. Reverse redistribution was associated with restoration of adequate microvascular reperfusion and with myocardial salvage and viability. The early postinjection scans on day 8 were the relevant images for assessing myocardial salvage and predicting wall motion recovery. PMID- 10716309 TI - Impairment of cardiac neuronal function in childhood dilated cardiomyopathy: an 123I-MIBG scintigraphic study. AB - Abnormalities of norepinephrine uptake have been found to reflect impairment of cardiac adrenergic neuronal function in adults with heart failure. To our knowledge, no data on childhood dilated cardiomyopathy (DCM) are available. The aim of this study was to assess the cardiac neuronal function using 123I metaiodobenzylguanidine (MIBG) scintigraphy in children with idiopathic DCM. METHODS: We studied 26 patients (mean age, 44+/-50 mo) with DCM and left ventricular dysfunction and 12 control subjects (mean age, 49+/-65 mo) with normal left ventricular function. All subjects underwent planar cardiac imaging after intravenous injection of 20-75 MBq 123I-MIBG. A static anterior view was acquired 4 h after injection. The heart-to-mediastinum count ratio was measured as described previously. RESULTS: On the basis of a reduction of the heart-to mediastinum count ratio, cardiac neuronal uptake of 123I-MIBG was significantly decreased in patients with DCM compared with cardiac uptake in control subjects (172%+/-34% versus 277%+/-14%; P<0.0001). A significant correlation was found between left ventricular ejection fraction and 123I-MIBG cardiac uptake in patients with DCM (y = 2.5x + 113.3; r = 0.80; P<0.0001). CONCLUSION: Cardiac adrenergic neuronal function is impaired in children with idiopathic DCM. 1231 MIBG cardiac scintigraphy is a useful tool to assess cardiac neuronal function in childhood DCM. PMID- 10716310 TI - Relationship between cerebral perfusion in frontal-limbic-basal ganglia circuits and neuropsychologic impairment in patients with subclinical hepatic encephalopathy. AB - Early detection of neuropsychologic impairment in cirrhotic patients with subclinical hepatic encephalopathy (SHE) is important for their prognosis and quality of life. Abnormal MRI and MR spectroscopy (MRS) findings have been proposed as early markers of brain damage in these patients, but the role of functional neuroimaging in this field still has to be defined. In this study, the SPECT perfusion pattern in patients with SHE was investigated, and the relationship between regional cerebral blood flow (rCBF) and the MRI, MRS, neuropsychologic evaluation and biochemical data of these patients was assessed. METHODS: Data were obtained from 13 cirrhotic patients with SHE and 13 age matched healthy volunteers. Fasting venous blood ammonia and manganese sampling and a battery of standardized neuropsychologic tests related to basal ganglia function and sensitive to the effects of liver disease were all performed on the same day. MRI and 99mTc-hexamethyl propyleneamine oxime SPECT were performed within 2 wk. RESULTS: A pattern of decreased prefrontal rCBF was found in patients with SHE compared with healthy volunteers. Basal ganglia and mesial temporal rCBF correlated inversely with performance on motor tasks involving speed (Purdue pegboard test) and frontal premotor function (Luria graphic alternances and Stroop tests). Thalamic rCBF correlated positively with T1 weighted MRI signal hyperintensity in the globus pallidus and with abnormal MRS findings. Neither the MRI signal intensity of the globus pallidus nor MRS correlated with neuropsychologic test results. CONCLUSION: Cirrhotic patients with SHE show a SPECT pattern of impaired prefrontal perfusion that does not seem to account for their neuropsychologic deficits. On the other hand, perfusion in some parts of the limbic system and limbic-connected brain regions, such as the striatum and the mesial temporal regions, increased with neuropsychologic impairment. These findings suggest that brain SPECT may be more sensitive than MRI in delineating cirrhotic patients requiring in-depth clinical testing to reveal basal ganglia-related neuropsychologic alterations. PMID- 10716311 TI - Visualization of the motor activation area using SPECT in neurosurgical patients with lesions near the central sulcus. AB - The purpose of this study was to visualize the motor area related to finger movement and a fist-making task using SPECT in patients with lesions near the central sulcus. METHODS: Eleven patients (9 with a brain tumor, 1 with cerebral infarction, and 1 with an arteriovenous malformation) were investigated. The first intravenous injection of 99mTc-ethyl cysteinate dimer (ECD) for the motor activation SPECT images was administered 2 min after completion of the fist making task with the hand contralateral to the brain lesion. The movement was stopped 2 min after injection, and activation SPECT was performed. After the scan, the second dose of 99mTc-ECD was injected into resting patients, and a second set of SPECT images was acquired. The first set of images was subtracted from the second set to obtain control images. Regions of interest were set bilaterally on the sensorimotor hand area; the supplementary motor area; the frontal, temporal, and occipital lobes; and the cerebellar hemispheres. The results of activation SPECT were expressed as positive or negative for a high count area, and the regional percentage change for activation images relative to resting images was calculated. RESULTS: Visual assessment of activation images was positive in 9 patients for the sensorimotor hand area and 7 patients for the supplementary motor area. The regional percentage change between activation and resting images for the high-count areas was 19.7% for the sensorimotor hand area and 18.2% for the supplementary motor area. Both values were significantly higher than those for other areas (P<0.05). CONCLUSION: Motor activation SPECT using a 99mTc-ECD split-dose method is easy to perform and may be helpful for presurgical visualization and identification of the sensorimotor hand area or the supplementary motor area. PMID- 10716312 TI - Cerebellar vasoreactivity in stroke patients with crossed cerebellar diaschisis assessed by acetazolamide and 99mTc-HMPAO SPECT. AB - Crossed cerebellar diaschisis (CCD) tends to persist or even worsen after supratentorial infarction. Several studies have shown impairment of cerebral vasomotor responsiveness in the hemispheric area of diaschisis in patients with hemispheric infarction. This finding has led to the concern that the lack of CCD reversibility might be associated with chronic circulatory abnormalities. We therefore assessed the vasoreactivity in the cerebellar hemisphere in which diaschisis is manifested using acetazolamide (ACZ) and SPECT. METHODS: Eight stroke patients with CCD (5 with unilateral hemispheric infarcts and 3 with unilateral intracerebral hemorrhage) had 99mTc-HMPAO SPECT scanning at rest and 20 min after intravenous injection of 1.0 g ACZ. The time interval after stroke ranged from 25 to 904 d. From the total counts obtained from each cerebellar hemisphere, the asymmetry index (AI) was calculated as (unaffected - affected cerebellar hemisphere)/unaffected cerebellar hemisphere x100. RESULTS: After ACZ, the mean AI (8.7+/-6.6) was significantly decreased (P<0.05) compared with that at rest (17.7+/-5.8). Seven of the 8 patients showed decrease in the AI after ACZ. In 1 patient, the direction of the asymmetry was reversed after ACZ so that the AI was negative. The ACZ-induced change in the AI did not show a significant correlation with the time interval after stroke, whether calculated in absolute terms or as a percentage change. CONCLUSION: This study shows that normal vascular supply is maintained in the CCD-affected cerebellar hemisphere over long periods of time after a stroke. Thus, the lack of CCD reversibility may not be attributed to a chronic circulatory insufficiency. The results lend support to the concept of functional deactivation and subsequent transneuronal degeneration as a likely explanation for CCD. It is unclear whether decreased AI after ACZ indicates a higher vascular response of the affected cerebellar hemisphere than that of the normal side. PMID- 10716313 TI - Response to percutaneous transhepatic portal embolization: new proposed parameters by 99mTc-GSA SPECT and their usefulness in prognostic estimation after hepatectomy. AB - Accumulation of 99mTc-galactosyl human serum albumin (GSA) in the liver correlates well with the parameters of hepatic function tests. We performed 99mTC GSA SPECT before and after percutaneous transhepatic portal embolization (PTPE) to induce compensatory hypertrophy of the remnant lobe before extensive hepatic resection and analyzed the responses of new proposed parameters in the future remnant lobe that showed hypertrophy. The aim of this study was to evaluate the usefulness of these parameters in prognostic estimation after hepatectomy. METHODS: We studied 10 patients with cholangiocarcinoma and 1 patient with metastatic liver tumor from sigmoid colon cancer. 99mTc-GSA SPECT was performed immediately before and 2 wk after PTPE. We analyzed the responses of the liver uptake ratio (LUR), functional volume (FV), and liver uptake density (LUD) in the future remnant lobe and evaluated their relationship with the prognosis after subsequent hepatic resection. RESULTS: LUR and FV increased slightly but were not associated with the prognosis after hepatic resection. LUD increased significantly after PTPE in the group showing a good outcome after hepatic resection but decreased after PTPE in the group showing a poor outcome (post-PTPE LUD, 0.064+/-0.017%/cm3 versus 0.035+/-0.006%/ cm3, P<0.05; response rate, 22.2%+/-11.9% versus -8.9%+/-17.6%, P<0.01). CONCLUSION: Responses of LUD to PTPE before hepatic resection in the future remnant lobe represent changes in asialoglycoprotein receptor activity per hepatocyte and predict responses to subsequent hepatic resection. LUD may be an important parameter for determining the outcome after hepatic resection. PMID- 10716314 TI - Use of 99mTc-furifosmin scintigraphy--planar and SPECT--to evaluate suggestive palpable and nonpalpable breast lesions. AB - Our objective was to evaluate the role of 99mTc-furifosmin scintigraphy--planar and SPECT--in discriminating benign from malignant breast disease. METHODS: The trial was prospective, open, and diagnostic. We recruited 30 consecutive patients with 14 palpable and 16 nonpalpable breast lesions. After receiving informed consent, we injected 555-640 MBq 99mTc-furifosmin intravenously in the arm contralateral to the breast lesion. Planar imaging and SPECT were performed. All patients underwent excision of the tumor within 2 wk. Using histology as the gold standard, we calculated sensitivity, specificity, and positive and negative predictive values for 99mTc-furifosmin in planar and SPECT technique. RESULTS: For 18 malignant and 12 benign breast lesions, a sensitivity of 50% for planar imaging and 72% for SPECT was seen. Specificity and positive and negative predictive values were 83%, 82%, and 53%, respectively, for planar imaging and 50%, 68%, and 55%, respectively, for SPECT. For the 14 palpable tumors (10 malignant, 4 benign), which averaged 17+/-10 mm in size (size range, 4-45 mm), a sensitivity of 60% for planar imaging and 80% for SPECT was achieved. Sixteen lesions were not palpable (median size, 9+/-3 mm [size range, 4-13 mm]). In this subgroup, 99mTc-furifosmin scintigraphy yielded a sensitivity of 37% for planar and 62% for SPECT technique (P>0.05). CONCLUSION: 99mTc-furifosmin scintigraphy is not a potent competitor to established scintigraphic procedures. In comparing this tracer with 99mTc-sestamibi and 99mTc-tetrofosmin, we cannot recommend 99mTc furifosmin for the diagnosis of breast cancer. PMID- 10716315 TI - 111In-labeled EGF is selectively radiotoxic to human breast cancer cells overexpressing EGFR. AB - Our objective was to determine whether the internalization and nuclear translocation of human epidermal growth factor (hEGF) after binding to its cell surface receptor (EGFR) could be exploited to deliver the Auger electron emitter 111In into EGFR-positive breast cancer cells for targeted radiotherapy. METHODS: hEGF was derivatized with diethylenetriamine pentaacetic acid (DTPA) and radiolabeled with 111In-acetate. The internalization of 111In-DTPA-hEGF by MDA-MB 468 breast cancer cells (1.3x10(6) EGFRs/cell) was determined by displacement of surface-bound radioactivity by an acid wash. The radioactivity in the cell nucleus and chromatin, isolated by differential centrifugation, was measured. The effect on the growth rate of MDA-MB-468 or MCF-7 (1.5x10(4) EGFRs/cell) cells was determined after treatment in vitro with 111In-DTPA-hEGF, unlabeled DTPA-hEGF, or 111In-DTPA. The surviving fraction of MDA-MB-468 or MCF-7 cells treated in vitro with 111In-DTPA-hEGF was determined in a clonogenic assay. The radiotoxicity in vivo against normal hepatocytes or renal tubular cells was evaluated by measuring alanine aminotransferase (ALT) or creatinine levels in mice administered high amounts of 111In-DTPA-hEGF (equivalent to human doses up to 14,208 MBq) and by light and electron microscopy of the tissues. RESULTS: Approximately 70% of 111In DTPA-hEGF was internalized by MDA-MB-468 cells within 15 min at 37 degrees C and up to 15% was translocated to the nucleus within 24 h. Chromatin contained 10% of internalized radioactivity. The growth rate of MDA-MB-468 cells was decreased 3 fold by treatment with 111In-DTPA-hEGF (45-60 mBq/cell). Treatment with unlabeled DTPA-hEGF caused a 1.5-fold decrease in growth rate, whereas treatment with 111In DTPA had no effect. Targeting of MDA-MB-468 cells with up to 130 mBq/cell of 111In-DTPA-hEGF resulted in a 2-logarithm decrease in their surviving fraction. No decrease in the growth rate or surviving fraction of MCF-7 cells was evident. There was no evidence of hepatotoxicity or renal toxicity in mice administered high amounts of 111In-DTPA-hEGF. Radiation dosimetry estimates suggest that the radiation dose to an MDA-MB-468 cell targeted with 111In-DTPA-hEGF could be as high as 25 Gy with up to 19 Gy delivered to the cell nucleus. CONCLUSION: 111In DTPA-hEGF is a promising novel radiopharmaceutical for targeted Auger electron radiotherapy of advanced, hormone-resistant breast cancer. PMID- 10716316 TI - Three-dimensional clinical PET in lung cancer: validation and practical strategies. AB - The feasibility of 3-dimensional acquisition mode for semiquantitative analysis in thoracic PET studies was compared to the conventional 2-dimensional mode. Several practical considerations were analyzed to propose an optimized scanning protocol for clinical use. METHODS: Twenty-one patients with focal thoracic abnormalities were evaluated with FDG PET. The acquisition consisted of 3 consecutive static scans for a single bed position: 3-dimensional (10 min), 2 dimensional (15 min), and 3-dimensional (5 min). On the basis of the average and maximum activity values per region of interest, standardized uptake value (SUV) normalized for total body weight (TBW), lean body mass (LBM), body surface area (BSA), and blood glucose level (PGL) were evaluated. The effect of the delay between tracer injection and PET scanning on the SUV, as well as on the relative error of the activity distribution, was studied from 40-134 min after tracer injection. RESULTS: A strong positive correlation was observed among SUVs from 2 dimensional and both 3-dimensional acquisitions. The mean SUV percentage differences between both acquisition modes were about 17%, differences that were not statistically significant when time postinjection was addressed in the analysis of covariance. SUVs provided the greatest variability and differences among studies on experimental periods up to 70 min postinjection. Indeed, the variability of 20% observed on the SUVs from 2 PET scans 13 min apart was reduced to 9% when the acquisitions started at least 70 min after tracer injection. In addition, a two-fold reduction in the relative error of the activity distribution was observed over this period of time. The reproducibility coefficient was increased from 0.87 to 0.95 before and after 70 min postinjection, respectively. No correlation was found between different normalization procedures of SUV and LBM, BSA, TBW, or height, whereas a weak correlation was found between SUV and PGL. CONCLUSION: 18F-FDG 3-dimensional PET is a realistic alternative to the gold standard 2-dimensional for clinical nonkinetic studies. A short, 5-min 3 dimensional acquisition at 70 min postinjection is proposed as the best protocol for the clinical evaluation of thoracic pathologies. PMID- 10716317 TI - Neutrophil-specific 99mTc-labeled anti-CD15 monoclonal antibody imaging for diagnosis of equivocal appendicitis. AB - We evaluated 99mTc-labeled anti-CD15 immunoglobulin M monoclonal antibody (LeuTech) for diagnosing acute appendicitis in patients with an equivocal clinical presentation. LeuTech avidly binds to circulating and sequestered human polymorphonuclear neutrophils in vivo, eliminating in vitro cell labeling and blood handling. METHODS: We studied 49 patients to evaluate the safety and efficacy of LeuTech imaging. 99mTc-labeled LeuTech was prepared on site using a lyophilized kit, 99mTc-labeled pertechnetate, and 2 different incubation techniques, 1 at room temperature and the other at 37 degrees C. The abdomen was serially imaged for up to 3 h after the intravenous administration of 370-740 MBq 99mTc-labeled LeuTech. Scans were read as positive or negative for acute appendicitis or other intraabdominal infection. The institutional diagnosis was established by surgery, other diagnostic studies, or 1-mo clinical follow-up. RESULTS: Scans were positive for appendicitis in all 26 patients with appendicitis, for a sensitivity of 100%, and negative for appendicitis in 19 of 23 patients without appendicitis, for a specificity of 83%. Accuracy, positive predictive value, and negative predictive value were 92%, 87%, and 100%, respectively. Results were not different between the LeuTech preparations. The rate of laparotomies with negative findings in patients who underwent surgery was 10%. The average time from injection to LeuTech visualization in the appendix for cases positive for appendicitis was 9 min. No serious adverse reactions occurred. CONCLUSION: LeuTech imaging is safe, rapid, and sensitive for diagnosis of appendicitis in equivocal cases. The potential advantages of LeuTech over currently available radiopharmaceuticals for infection imaging are ease of preparation, absence of blood handling, excellent image quality, no requirement for SPECT, and rapid diagnostic uptake. PMID- 10716318 TI - Imaging acute appendicitis: an opportunity for nuclear medicine in the surgical emergency room. PMID- 10716319 TI - 111In-pentetreotide scintigraphy in the detection of insulinomas: importance of SPECT imaging. AB - The aim of this study was to determine whether the systematic use of SPECT can increase the reported low sensitivity of somatostatin receptor scintigraphy (SRS) in detecting insulinomas. METHODS: Fourteen patients were evaluated. After 111In pentetreotide injection (approximately 250 MBq intravenously), abdominal SPECT images were obtained at 4 h and multiple planar images were obtained at 4 and 24 h. MRI and CT were performed within 1 mo of SRS. Sixteen tumors were histologically verified after surgery in 14 patients. RESULTS: SPECT revealed 14 lesions in 12 patients (sensitivity, 87.5%), both planar SRS and MRI revealed 7 tumors in 7 patients (sensitivity, 43.8%), and CT revealed only 5 lesions in 4 patients (sensitivity, 31.3%). Moreover, in 4 patients SPECT was the only examination with positive findings. CONCLUSION: SPECT at 4 h is mandatory for preoperative detection of insulinomas using SRS because the images are more sensitive than planar images and are superior to images from other conventional methods. PMID- 10716320 TI - Radiolabeled interleukin-8: specific scintigraphic detection of infection within a few hours. AB - Several small receptor-binding agents have been tested for imaging of infection and inflammation. The potential of chemotactic peptides and of interleukins is promising and superior to that of conventional agents. In this study, we investigated the potential of interleukin-8 (IL-8) to image infection in rabbits. METHODS: IL-8 was labeled with 123I using the Bolton-Hunter method. Twenty-fours hours after induction of Escherichia coli abscesses in the left thigh muscle, rabbits were injected intravenously with 18.5 MBq 123I-IL-8. Gamma camera images were obtained at 5 min and at 1, 4, and 8 h after injection. Biodistribution was determined 8 h after injection. RESULTS: 123I-IL-8 rapidly cleared from the blood. Accumulation of 123I-IL-8 in the abscess was visible as early as 1 h after injection. The highest abscess uptake was obtained 4 h after injection (2.6+/-0.2 percentage injected dose [%ID]), whereas 123I-IL-8 rapidly cleared from all other tissues. This resulted in increases in abscess-to-background ratios to 13.0+/-0.7 (8 h after injection), as determined by quantification of the images. In tissue biodistribution (8 h after injection), the abscess uptake was 0.057+/-0.011 %ID/g with abscess-to-contralateral muscle ratios of 114.7+/-23.0. The radioiodination method clearly affected the in vivo biodistribution of IL-8 because IL-8 iodinated using the lodo-Gen method cleared significantly slower from the blood and most other organs, resulting in poor visualization of the abscess. CONCLUSION: The superior characteristics of IL-8 radioiodinated using the Bolton Hunter method--i.e., high abscess uptake and rapid background clearance within a few hours--make IL-8 a promising agent to image infection and inflammation. PMID- 10716321 TI - When is too much too much and yet not enough? Alas, a plethora of opportunities but where's the beef? PMID- 10716322 TI - Radionuclide analysis of drug-induced blood-pool changes in liver and other organs. AB - Although it is possible to repopulate the animal liver with transplanted hepatocytes, the success of such transplants depends, in part, on the number of transplanted cells that enter the hepatic sinusoids. Pharmacologic alteration of hepatic vascular tone, and hence blood volume, can increase the number of cells that are successfully transplanted. Although analysis of changes in vascular beds is helpful for developing strategies for cell transplantation, convenient methods to analyze such changes are lacking. The objective of this study was to determine whether 99mTc-labeled red blood cells could be used to reveal pharmacologically induced blood pool changes in various organs. METHODS: F344 rats were injected with syngeneic labeled red blood cells and subjected to blood pool analysis with gamma camera imaging. Animals were treated with phenylephrine, phentolamine, labetalol, and nitroglycerine. To correlate hepatic blood pool changes with structural alterations at the vascular level, microspheres were injected into the portal circulation of these animals. RESULTS: Phenylephrine significantly increased cardiac and pulmonary blood pools, findings in agreement with its alpha adrenergic effects. Phentolamine increased the hepatic, splenic, and pulmonary blood pools, whereas labetalol increased only the pulmonary blood pool. Nitroglycerine increased both hepatic and splenic blood pools. Prior administration of phentolamine, labetalol, and nitroglycerine prevented the phenylephrine-induced changes. When microspheres were injected into the portal circulation after nitroglycerine administration, they penetrated more distal locations in the liver lobule. CONCLUSION: These data indicate that it is possible, using radionuclide methods, to noninvasively show pharmacologically induced hemodynamic changes. This finding is potentially useful for studying hepatic physiology and may also have applications for cell therapy. PMID- 10716323 TI - Pretargeted radioimmunotherapy of human colorectal xenografts with bispecific antibody and 131I-labeled bivalent hapten. AB - We have developed a pretargeting strategy, called the affinity enhancement system (AES), which uses bispecific antibodies to target radiolabeled bivalent haptens to tumor cells. The aim of this study was to evaluate the potential of the AES for the radioimmunotherapy (RIT) of LS174T colorectal xenografts in comparison with RIT with directly labeled F(ab')2 fragment. METHODS: A total of 6 groups of tumor-bearing mice were treated using anticarcinoembryonic antigen (CEA) x anti diethylenetriamine pentaacetic acid (DTPA)-In bispecific antibody (BsF(ab')2) and 131I-labeled di-DTPA-In bivalent hapten. Three groups of mice were injected with various activities of 131I-labeled bivalent hapten (75, 96, and 112 MBq) 20 h after administration of BsF(ab)'2. Three other groups were injected with an almost constant activity of labeled hapten (102 MBq) at 3 time periods (15, 30, and 48 h) after BsF(ab')2 administration. For conventional RIT, mice were treated with 96 MBq 131-labeled anti-CEA F(ab')2. Control groups were left untreated. Toxicity and tumor growth were monitored at weekly intervals. RESULTS: Doses used for conventional RIT induced severe toxicity and resulted in death of several treated animals. Nevertheless, all surviving animals treated with 131I-labeled anti-CEA F(ab')2 relapsed shortly after treatment (tumor growth delay = 48+/-13 d). For animals treated with the AES reagents, toxicity varied with the pretargeting time interval and the administered activity. For 20-h pretargeting time, the maximum tolerated dose was 96 MBq. For all AES RIT except 1 (with 48-h pretargeting time interval and growth delay of 82+/-26 d), no tumor growth was observed over a period of 8 mo. Furthermore, based on clinical and histologic criteria, 33% of the treated mice were considered cured. CONCLUSION: High cure rates of LS174T colon carcinoma were achieved with the AES, and the flexibility of the pretargeting approach allowed the control of hematologic toxicity, which is the main limitation to dose escalation with conventional RIT. PMID- 10716324 TI - Kinetic differences and similarities among 3 tracers of myocardial glucose uptake. AB - Two glucose tracer analogs, uniformly labeled [14C]2-deoxyglucose ([U-14C]2DG) and FDG, are widely used to assess myocardial glucose uptake. Despite the similar electron configuration of the fluorine and hydrogen atoms, uptake of the 2 tracer analogs may not be the same because of their different electronegativity. METHODS: To test this hypothesis, we determined glucose uptake in isolated rat hearts simultaneously from the accumulation of [U-14C]2DG radioactivity in the tissue, by continuous monitoring of FDG accumulation with a pair of coincidence detectors and by cumulative release of 3HOH from [2-3H]glucose. A first group of hearts was perfused at physiologic workload with Krebs-Henseleit buffer containing 10 mmol/L glucose; a second group, with the buffer containing 5 mmol/L glucose plus 0.4 mmol/L oleate and 1 mU/mL insulin. Third and fourth groups were subjected to ischemia (i.e., a 75% reduction in coronary flow) and reperfused. For the third group, the buffer contained 5 mmol/L glucose; for the fourth, 5 mmol/L glucose plus 0.4 mmol/L oleate. RESULTS: No difference in the total amount of tracer accumulation in any group was seen between the 2 tracer analogs. The ratio [+/-SD] of [U-14C]2DG to FDG ranged from 0.93+/-0.09 to 1.31+/-0.11. However, both tracer analogs paralleled glucose uptake in the absence of insulin but underestimated glucose uptake significantly in the presence of insulin. Changes in 2DG uptake with ischemia and reperfusion could be detected only with FDG. CONCLUSION: Although uptake of [U-14C]2DG equals uptake of FDG quantitatively, acute changes in 2DG uptake (and, thus, in the tracer-tracee relationship) are detectable only with the fluorine-labeled tracer. PMID- 10716325 TI - Evaluation of 11C-colchicine for PET imaging of multiple drug resistance. AB - Overexpression of P-glycoprotein (P-gp) can confer multiple drug resistance (MDR) phenotype on cancer cells and tumors by reducing intracellular accumulation of various cytotoxic agents. Early diagnosis of MDR in the clinic will serve to improve the efficacy of chemotherapeutic intervention and the quality of life of patients. In this article we describe use of a positron-emitting MDR tracer, 11C colchicine (CHC), to evaluate MDR by PET imaging. Unlike existing MDR tracers such as 99mTc-sestamibi, this compound is electroneutral, with biodistribution not affected by perturbations of membrane potential. METHODS: In vitro studies showed that resistance to CHC is correlated to resistance to Taxol (paclitaxel). The results of biodistribution experiments were found to be consistent with previously reported experiments with CHC labeled with other isotopes. On the basis of in vitro experiments with a series of drug-resistant variants of the human neuroblastoma BE (2)-C cell line, a mathematic model of 11C-CHC distribution in tumors was formulated. Dynamic PET 11C-CHC imaging experiments were performed with nude rats xenografted with the BE (2)-C-sensitive and resistant strains. Each scan was accompanied by a transmissions scan and a static FDG scan. These scans allowed improved image localization. RESULTS: We observed an approximately 2-fold difference between 11C-CHC accumulation in sensitive and resistant tumors. Imaging data were analyzed using the mathematic model, and various parameters characterizing resistance could be identified and estimated. In particular, the parameter r, proportional to the level of resistance of the tumors, was obtained. We showed that the ratio of these r parameters determined from the sensitive and resistant tumors was identical to the ratio of CHC accumulation in the corresponding sensitive and resistant cell lines used for xenografting. CONCLUSION: These in vivo experiments provided additional evidence for the indirect effect of P-gp action on CHC-to-tubulin binding, which in turn determines CHC uptake in tumors. The significance of these findings and future plans is discussed. PMID- 10716326 TI - Receiver operating characteristic evaluation of iterative reconstruction with attenuation correction in 99mTc-sestamibi myocardial SPECT images. AB - The purpose of this study was to evaluate differences in myocardial defect detection between 99mTc-sestamibi myocardial SPECT images reconstructed using conventional filtered backprojection (FBP) without attenuation correction (AC) and those reconstructed using maximum-likelihood expectation maximization with nonuniform attenuation correction (MLAC). METHODS: An observer study and receiver operating characteristic (ROC) curve analysis were performed using simulated 99mTc-sestamibi SPECT data from a population of 24 mathematic anthropomorphic torso phantoms, which realistically modeled a wide range of anatomic variations. The phantoms modeled male patients with a flat diaphragm, male patients with a diaphragm raised to the level of the heart, and female patients with large breasts. Transmural, cold defects with a contrast of 0.25 were simulated in the left ventricular wall for 6 locations. Noisy projection data were generated from the phantoms and included the effects of nonuniform attenuation, collimator detector response, and scatter. The data were then reconstructed using FBP and MLAC. Images were displayed in the short- and long-axis formats, as in clinical practice. Eight observers viewed blocks of FBP and MLAC images and, for each image, indicated on a continuous rating scale the probability that a defect was present. From the rating data, FBP and MLAC ROC curves were generated, and their areas (Az) were estimated and compared. RESULTS: In general, the FBP and MLAC ROC curves did not cross and the MLAC curve showed a higher Az than did the corresponding FBP curve. For male phantoms with a flat diaphragm, the average difference in Az was 0.04 and was not statistically significant (at the P = 0.05 level) for 6 of 8 observers. For male phantoms with a raised diaphragm, the average difference in Az was 0.22 and was statistically significant for 6 of 8 observers. For female phantoms with large breasts, the average difference in Az was 0.19 and was statistically significant for all 8 observers. CONCLUSION: This study showed an improvement in defect detection in myocardial SPECT images using MLAC in comparison with images using FBP without AC, particularly for patients with large breasts or with a diaphragm raised to the level of the heart. PMID- 10716327 TI - Channelized hotelling and human observer correlation for lesion detection in hepatic SPECT imaging. AB - Mathematic "model" observers that predict human performance are of interest in medical imaging as substitutes in psychophysical studies. We have examined the correlations between human observers and several forms of the channelized Hotelling observer (CHO) for a tumor detection task with simulated SPECT liver images that were used to study the effects of scatter and scatter correction on detection. METHODS: A receiver operating characteristic (ROC) study was devised to investigate the relative value of a scatter-subtraction strategy in SPECT imaging. The study used simulated images of the biodistribution of 99mTc-labeled FO23C5 anticarcinoembryonic antigen antibodies within the liver. Projection data for 3 separate tumor locations and 5 strategies for handling scatter were obtained using Monte Carlo software applied to an anthropomorphic phantom. The strategies were (a) perfect scatter rejection, (b) no scatter correction, (c) no scatter correction under an assumption of an elevated amount of scatter, (d) an energy-spectrum-based scatter compensation of the normal-scatter case (b), and (e) similar scatter compensation for the elevated-scatter case (c). Image reconstruction approximated current clinical procedures at the University of Massachusetts Medical School. Human performance for each combination of location and strategy was based on averaging the areas under the ROC curve for 7 individuals. A set of 15 signal-to-noise ratios (SNRs) was derived from these averages for comparison with SNRs for CHO models featuring constant-Q and difference-of-gaussian (DOG) filters. RESULTS: The Spearman rank correlation coefficient was 0.92 (P = 0.000001) when comparing task performances for the average human and a constant-Q CHO using 4 square-profile channels. For the DOG version of the CHO, comparison with the average human found a coefficient of 0.84 (P = 0.00005). CONCLUSION: The significant positive correlations found between the rankings of the average human observer and the CHOs for our detection task indicate that a channelized model observer could eventually serve as a replacement for human observers. The specific CHO models we have used are best suited to screen for significant differences between strategies before a human psychophysical study. PMID- 10716328 TI - Measurement of dopamine release with continuous infusion of [11C]raclopride: optimization and signal-to-noise considerations. AB - PET studies with [11C]raclopride provide an indirect measure of changes in synaptic dopamine. Previously, we used the bolus-plus-infusion (B/I) method to assess dopamine response from the percentage change in binding potential (deltaBP) before and after administration of amphetamine. The goal of this work is to optimize the measurement of changes in neurotransmitter with the B/I method by choosing the optimal timing for pre- and poststimulus scanning. METHODS: Two sources of variability in deltaBP were considered: within-subject and between subject noise. A noise model based on a phantom study and human data was used to evaluate the within-subject noise. For between-subject noise, simulated time- activity curves were generated from measured [11C]raclopride input functions. Optimal timing to measure deltaBP was determined and applied to human data. RESULTS: According to the simulation study, the optimal scan times for pre-and poststimulus scans were 39-50 and 58-100 min, respectively. The optimal timing resulted in a 28% noise reduction compared with the original timing. By applying the optimal timing to human studies, the statistical significance of the difference in deltaBP between patients with schizophrenia and healthy volunteers increased from P = 0.038 to 0.012. CONCLUSION: Careful assessment of the sources of noise in receptor imaging studies can increase the sensitivity of the B/I method for the detection of biologic signals. PMID- 10716329 TI - Radioimmunotherapy for the intensification of conditioning before stem cell transplantation: differences in dosimetry and biokinetics of 188Re- and 99mTc labeled anti-NCA-95 MAbs. AB - A new concept is the intensification of preparative regimens for patients with advanced leukemia using monoclonal antibodies (MAbs) with an affinity for beta emitter-labeled bone marrow. 188Re is a high-energy beta emitter that has therapeutic promise. Our first aim was to clarify whether the therapeutic application of 188Re-MAb against nonspecific cross-reacting antigen 95 (NCA-95) can be predicted from biokinetic data derived from 99mTc-labeled NCA-95. Our second aim was to show that a radiation absorbed dose of > or =12 Gy in the bone marrow can be achieved using 188Re-MAb. METHODS: Dosimetric data were obtained for both radiotracers from multiple planar whole-body scans (double-head gamma camera), blood samples, and urine measurements from 12 patients with advanced leukemia. Radiation absorbed doses were calculated using MIRDOSE 3 software. RESULTS: Radiation absorbed doses to bone marrow, liver, spleen, lung, and kidney were 2.24, 0.50, 1.93, 0.05, and 0.90 mGy/MBq, respectively, using 99mTc-MAb and 1.45, 0.43, 1.32, 0.07, and 0.71 mGy/MBq, respectively, using 188Re-MAb. These differences were statistically significant for bone marrow, spleen, and kidney. The main differences were less accumulation of 188Re-MAb in bone marrow (31%+/ 13% compared with 52%+/-13%) and faster elimination through urine (25%+/-3% compared with 15%+/-5% after 24 h). On the basis of these data, a mean marrow dose of 14+/-7 Gy was achieved in 12 patients suffering from leukemia after application of approximately 10+/-2 GBq 188Re-MAb. CONCLUSION: Myeloablative radiation absorbed doses can easily be achieved using 188Re-MAb. 99mTc- and 188Re MAb showed similar whole-body distributions. However, direct prediction of radiation absorbed doses from the 99mTc-MAb, assuming identical biokinetic behavior, is not valid for the 188Re-MAb in a single patient. Therefore, individual dosimetry using 188Re-MAb is needed to calculate therapeutic activity. PMID- 10716330 TI - Single-dose versus fractionated radioimmunotherapy: model comparisons for uniform tumor dosimetry. AB - Targeting molecules with reduced immunogenicity will enable repetitive administrations of radioimmunotherapy. In this work a mathematical model was used to compare 2 different treatment strategies: large single administrations (LSAs) and rapid fractionation (RF) of small individual administrations separated by short time intervals. METHODS: An integrated compartmental model of treatment pharmacokinetics and tumor response was used to compare alternative treatments that delivered identical absorbed doses to red marrow. RESULTS: Based on the key assumption of uniform dose distributions, the LSA approach consistently produced smaller nadir values of tumor cell survival and tumor size. The predicted duration of remission was similar for both treatment structures. These findings held for both macroscopic and microscopic tumors and were independent of tumor cell radiosensitivity, proliferation rate, rate of tumor shrinkage, and uptake characteristics of radiolabeled material in tumor. CONCLUSION: Clinical situations for which each treatment is most appropriate may be tentatively identified. An LSA using a short-range-emitting radionuclide would be most appropriate for therapy of microscopic disease, if uptake is relatively homogeneous. RF using a longer range emitter would be most appropriate for macroscopic disease, if uptake is heterogeneous and varies from one administration to another. There is a rationale for combining LSA and RF treatments in clinical situations in which slowly growing macroscopic disease and rapidly growing microscopic disease exist simultaneously. PMID- 10716331 TI - Angulation errors in parallel-hole and fanbeam collimators: computer controlled quality control and acceptance testing procedure. AB - Angulation errors in collimators of 1 degrees or even less can seriously diminish the resolution of SPECT images. We have developed a computer-controlled quality control procedure that can be used for acceptance testing and regular routine checks. METHODS: Using a marker point source and a computer-controlled x-y positioning table, we investigated 7 parallel-hole and 3 fanbeam collimators. The results are presented as collimator surface maps, which are easy to interpret visually. RESULTS: The measurement accuracy for absolute angulation errors was better than 0.32 degrees. Regional variations in channel tilt could be detected with an accuracy better than 0.16 degrees. Six parallel-hole collimators were found acceptable for high-resolution SPECT imaging. For a parallel-hole collimator that had to be replaced because of nonoptimal image quality, our measurements clearly identified regions of directionally uniform angulation errors. Two fanbeam collimators showed slight concavities. CONCLUSION: Automation of the measurement and evaluation process make this procedure suitable for both acceptance tests and routine quality control checks. It can be applied to parallel-hole, fanbeam, converging, and diverging collimators, regardless of their individual geometry. No technical collimator specifications are needed. Our results reveal subtle mechanical deformations of collimators. They also show that for a detailed investigation, angulation error surface maps should be used to discover regional preferences in channel orientation. PMID- 10716332 TI - A cell-culture reactor for the on-line evaluation of radiopharmaceuticals: evaluation of the lumped constant of FDG in human glioma cells. AB - A fluidized-bed cell-culture reactor with on-line radioactivity detection was developed for the in vitro evaluation of radiopharmaceuticals. The technique was applied to measure the dependency of the lumped constant (LC) of FDG on the glucose concentration in the culture medium in a human glioma cell line. METHODS: Human glioblastoma cells (86HG39) immobilized in open porous microcarriers were cultivated in a continuously operating fluidized-bed bioreactor. At different glucose concentrations in the culture medium, step inputs (0.1 MBq/mL) of FDG were performed and the cellular uptake of FDG was measured on-line and compared with analyzed samples. From these results, the LC of FDG and its dependency on the glucose concentration were calculated. RESULTS: This fluidized-bed technique enabled precise and reproducible adjustment of all relevant experimental parameters, including radiotracer time-concentration course, medium composition, pH, dissolved oxygen and temperature under steady-state conditions, and an on line determination of the intracellular radiotracer uptake. The immobilized glioma cells formed stable, 3-dimensional, tumor-like spheroids and were continuously proliferating, as proven by an S-phase portion of 25%-40%. For further examination of the cells, an enzymatic method for detachment from the carriers without cellular destruction was introduced. In the FDG experiments, a significant dependency of the LC on the glucose level was found. For normoglycemic glucose concentrations, the LC was determined to be in the range of 0.7+/-0.1, whereas in hypoglycemia LC increased progressively up to a value of 1.22+/-0.01 at a glucose concentration of 3 mmol/L. CONCLUSION: The bioreactor represents an improved in vitro model for the on-line evaluation of radiotracers and combines a wide range of experimental setups and 3-dimensional, tissue-like cell cultivation with a technique for on-line radioactivity detection. PMID- 10716333 TI - Induction of recombinant gene expression in stably transfected cell lines using attenuated vaccinia virus MVA expressing T7 RNA polymerase with a nuclear localisation signal. AB - There are major drawbacks using vaccinia virus (VV) expressing T7 polymerase for eukaryotic expression. VV is infectious for humans and due to cytosolic replication of Poxviridae, transient transfection of T7 promoter containing plasmids is necessary, which varies in efficiency. Several improvements have been introduced to this system to enhance expression of herpes viral glycoproteins. Stably transfected cell lines were generated with an EBV-based episomal plasmid vector which can be pushed to increasing copy numbers under selective pressure. The avirulent vaccine MVA strain was adopted to generate a safe laboratory vector for inserting the bacteriophage T7 RNA polymerase gene with (+) or without (-) a nuclear localisation signal. Constructs were designed for recombination into the VV haemagglutinin gene as recombinants could not be isolated successfully when inserting into the MVA thymidine kinase locus. Both T7 MVA recombinants induced foreign protein expression in transiently transfected cells but only the T7-/+ MVA induced target protein expression in stably transfected cells. The level of protein expression by this induction mechanism was comparable to, or superior to levels obtained with VV recombinants expressing the gene under control of the VV 11 k IE promoter. The results suggests that the T7+ MVA virus can be used to induce gene expression in stable recombinant cell lines and offers an attractive and safe alternative to other inducible eucaryotic expression systems. PMID- 10716334 TI - Routine detection and quantification of hepatitis B virus DNA in clinical laboratories: performance of three commercial assays. AB - The detection and quantification of hepatitis B virus (HBV) genomes in molecular biology-based assays appear to be the most reliable methods for monitoring HBV infection and assessing responses to antiviral treatment. The aim of this study was to evaluate the performance of three HBV-DNA detection and quantification assays currently used for the management of HBV-infected patients: a solution hybridization assay based on hybrid-capture (Digene Hybrid-Capture, Murex Diagnostics, Dartford, UK); a signal-amplification assay based on 'branched-DNA' (bDNA) technology (Quantiplex HBV DNA, Bayer Diagnostics, Emeryville, CA); and a target-amplification assay based on competitive polymerase chain reaction (Amplicor HBV Monitor, Roche Molecular Systems, Pleasanton, CA). The Monitor assay was significantly more sensitive than both the hybrid-capture and bDNA methods. This better sensitivity appeared to be clinically relevant. The linear ranges of quantification in the hybrid-capture, bDNA and Monitor methods were 6.5 9 log10 genome copies/ml, 6.5-9.5 log10 genome equivalents/ml, and 3-5.5 log10 genome copies/ml, respectively. However, the HBV-DNA units used in the three assays were not comparable. The specificity of the hybrid-capture, bDNA and Monitor assays was 99.2% (95% confidence interval: 97.7-100.0%), 99.2% (97.7 100.0%), and 97.8% (95.3-100%), respectively. Their within-run coefficients of variation and log10 SDs were 5.5% (+/- 0.025 log10 copies/ml), 6.7% (+/- 0.029 log10 Eq/ml) and 21.0% (+/- 0.093 log10 copies/ml), respectively. Between-run coefficients of variation ranged from 4.4-39.1%, 5-39.5%, and 17.8-96.1%, respectively. The competitive PCR-based Monitor assay appears to be significantly more sensitive but slightly less specific and reproducible than the hybrid capture and bDNA methods. Given their respective performance, these three assays should be used in complementary fashion in the management of HBV-infected patients. PMID- 10716335 TI - A method for the preparation of highly purified adeno-associated virus using affinity column chromatography, protease digestion and solvent extraction. AB - Recombinant adeno-associated virus (AAV) is becoming the vector of choice for many gene therapy protocols. There has been much recent progress made toward increasing AAV titres but a continuing problem in using AAV has been that it is relatively difficult to concentrate and purify. Traditional methods, such as caesium chloride (CsCl) gradients, have drawbacks, notably extended purification times and the ability to process only limited volumes. Where the target cells of interest require a high multiplicity of infection (MOI), or to complete in vivo experiments, there is a requirement for both the production of high titre and a large volume of virus. This is laborious to obtain using traditional methods. A simple technique is described here for purifying AAV, involving affinity chromatography, protease digestion and solvent extraction that retains both the high yields and titres obtained using CsCl gradients. In addition, this technique displays a fast throughput and may be used to purify AAV from larger volumes than CsCl gradients. The high yield and purity of these virus preparations has allowed us to achieve good levels of expression in the target cell types tested. The purification technique described here will be applicable to any protocol that requires high titre, high purity recombinant AAV (rAAV). PMID- 10716336 TI - A novel EBNA-1 tag system for high level expression and efficient detection of fusion proteins in vitro and in vivo. AB - A highly specific monoclonal antibody, which recognises an epitope between amino acids (aa) 408 and 446 of EBNA-1, was found to immunoprecipitate this protein with great efficiency. By adding a 39-aa tag, the BGLF4 protein product of EBV was shown to express in the cytoplasm of transfected cells. This EBNA-1 tag could be used for the detection of specific protein expression by immunoblotting, immunofluorescence and, especially, immunoprecipitation assays. A plasmid vector encoding this EBNA-1 tag sequence was therefore designed for efficient expression both in vitro and in vivo. The efficient translational start signal of black beetle virus (BBV) was placed under the control of the SV40 or T7 promoters, and the beta-globin splicing signal used to enhance the transportation of mRNA from the nucleus to the cytoplasm. PMID- 10716337 TI - Optimal conditions for the growth, purification and storage of the avian reovirus S1133. AB - In spite of their importance as avian pathogens causing important losses in poultry farming, the biochemistry of avian reoviruses has been little investigated. In order to facilitate the handling of these agents in the laboratory, a study was carried out to establish the best conditions both for growing the avian reovirus S1133 in primary cultures of chicken embryo fibroblasts and for purification and storage of viral suspensions. The results indicate that the conditions used currently for the manipulation of mammalian reoviruses are not always the best for handling their avian counterparts. In particular, avian reoviruses are much less stable than mammalian reoviruses, and specific conditions for the purification and storage of avian reoviruses therefore should be used. Furthermore, the instability of avian reovirions may have important implications for the life cycle and pathogenesis of the virus within the animal host. PMID- 10716338 TI - Measuring infectious bursal disease virus RNA in blood by multiplex real-time quantitative RT-PCR. AB - A quantitative reverse transcription polymerase chain reaction (RT-PCR) protocol for assessing infectious bursal disease virus (IBDV) RNA levels in blood was developed using the ABI PRISM 7700 Sequence Detection System coupled with TaqMan chemistry. To control for variations in sampling and processing between samples 28S rRNA was co-amplified in a multiplex reaction and used to quantify total RNA. Relative quantification and standardisation was achieved using a log10 dilution series of RNA extracted from IBDV stock. A linear relationship was observed between input RNA and cycle threshold values (C(T)) over 5 log10 dilutions for the IBDV-specific product and 6 log10 dilutions for the 28S rRNA-specific product. As a test of the assay it was used to determine whether differences in susceptibility to IBDV observed between inbred lines of chickens could be detected at the level of viral load in the blood. Viral RNA levels peaked 2 days post-infection when there was significantly less viral RNA in the blood of resistant line 6(1) chickens compared with the more susceptible Brown Leghorns (P = 0.01). These results demonstrate that the course of IBDV infection can be monitored by quantifying IBDV RNA extracted from blood of infected chickens using TaqMan technology. PMID- 10716339 TI - Comparison of three polymerase chain reaction methods for routine detection of bovine herpesvirus 1 DNA in fresh bull semen. AB - Five bulls were inoculated intrapreputially with Bovineherpes virus 1 (BHV 1), in order to compare the relative sensitivity of three polymerase chain reaction (PCR) assays for routine diagnosis of fresh bovine semen for the presence of BHV 1 Semen was collected twice a week up to 107 days post-infection (dpi). To reactivate latent virus, the bulls were treated with dexamethasone from 44 until 48 dpi. All samples were examined before and after cryopreservation treatment using a standard virus isolation (VI) method and three PCR assays: PCR A, PCR B and PCR C. PCR A and PCR C used an internal control plasmid DNA template and PCR B used the split sample method in order to control for false negative results. Of the 149 fresh semen samples that were tested, PCR A detected 45 positive, PCR B detected 39 positive and PCR C detected 66 positive, while virus was isolated from 22 samples. Of the 149 samples treated by cryopreservation, the virus was isolated from 13 samples and PCR C was positive in 21 samples. The results demonstrate that all three PCR assays are more sensitive than virus isolation, particularly during the later phases of infection. PMID- 10716340 TI - Sensitive and accurate quantitation of hepatitis B virus DNA using a kinetic fluorescence detection system (TaqMan PCR). AB - The laboratory diagnosis of hepatitis B virus (HBV) infection is based mainly on serological assays. Yet the detection and quantitation of viral DNA is necessary when addressing directly the question of infectivity or when monitoring the viral load during therapy. Standard hybridization assays allow for exact quantitation, but their sensitivity is limited to 10(5)-10(6) viral genomes per ml of serum. The most sensitive tests for HBV DNA are nested PCR systems, which recognize virtually one molecule of the target DNA per reaction. However, these assays only provide very coarse quantitative statements. To take advantage of both methods, a new assay for HBV DNA is described based on the commercial TaqMan system. This assay is capable of quantifying HBV DNA from the theoretical lower limit up to 10(10) genome equivalents per ml of serum and, thus, covers the complete range of naturally occurring states of infections. The method was calibrated on the basis of serial plasmid dilutions and compared with a well-established nested PCR system. More than 100 HBV positive sera and serial dilutions of the Eurohep standard for both ad and ay subtypes were analyzed. The assay reliably detected all HBV positive samples. It shows minimal run-to-run deviations, allows for quantitation that covers eight orders of magnitude, and finally, completely avoids the risk of cross-contamination by PCR products. Thus, this technique combines the sensitivity of PCR amplification and the quantitation potential of hybridization tests and it is time efficient and safer. PMID- 10716341 TI - A new procedure to differentiate citrus tristeza virus isolates by hybridisation with digoxigenin-labelled cDNA probes. AB - A non-isotopic hybridisation procedure was developed to differentiate isolates of citrus tristeza virus (CTV) using digoxigenin (DIG)-labelled cDNA probes and different kinds of target RNA. Hybridisation of DIG-probes with purified double stranded RNA (dsRNA) or concentrated total RNA extracts spotted on nylon membranes allowed detection of CTV nucleic acid equivalent to as little as 0.1-1 mg infected tissue, when the reaction was developed with a chemiluminiscent substrate. This sensitivity was similar to or slightly better than that obtained by hybridisation with a 32P-labelled probe. CTV was also detectable by hybridisation of DIG-probes with tissue prints from freshly cut young citrus shoots. Hybridisation of tissue prints with DIG-probes under stringent conditions (60 degrees C and 50% formamide) could differentiate CTV isolates in citrus, whether grown in the greenhouse or in the field. This rapid and sensitive procedure can easily be applied to many samples, even under field conditions, and opens the way to monitoring spatio-temporal movement of specific CTV strains (or groups of strains) in epidemiological studies. PMID- 10716342 TI - Rapid simultaneous detection of two orchid viruses using LC- and/or MALDI-mass spectrometry. AB - Liquid chromatography/mass spectrometry (LC/MS) and matrix-assisted laser desorption-ionization (MALDI) mass spectrometry are capable of providing molecular mass information on biological samples with high speed, accuracy and sensitivity. With mass spectrometry, identifying a virus based on the molecular weight of its coat protein is relatively simple and accurate. The technique can be applied to all viruses with known coat protein molecular weights. Using the LC/MS and/or MALDI, this paper describes rapid simultaneous detection of the two most prevalent orchid viruses, namely cymbidium mosaic potexvirus (CymMV) and odontoglossum ringspot tobamovirus (ORSV). The coat protein molecular weights of CymMV and ORSV were detected accurately using an extract from 1 g of virus infected Oncidium orchid flower. Because LC/MS and MALDI allow automated analyses of multiple samples with simple preparation steps, both techniques are ideal for rapid identification of viruses from a large number of samples. This is the first report on the application of LC/MS and/or MALDI for simultaneous detection of two plant viruses from an infected plant extract. PMID- 10716343 TI - Use of denaturing gradient gel electrophoresis for human polyomavirus JC sequence analysis. AB - Five genotypes of human polyomavirus JC (JCV) have been detected so far by nucleotide sequencing and by restriction fragment length polymorphism analysis. These genotypes are the result of geographically based virus evolution. However, interesting aspects of genotyping could involve both pathogenetic and diagnostic aspects. The product from amplification of JCV sequences, found in urine from 12 healthy individuals from different countries, have been analysed by denaturing gradient gel electrophoresis (DGGE) and by nucleotide sequencing. The aim of this study was to assess if the DGGE analysis could be used to study the variability of the JCV genome. The target sequence of this study was 233 bp long, within the gene coding for the VP1, known to contain several type-determining sites. Four DGGE patterns have been observed among our strains. Five strains, from African individuals, were of type 3 and exhibited the same electrophoretic pattern, clearly distinguishable from that of the type 1 strains detected in the urine from 6 European individuals. Five type 1 strains shared a similar DGGE pattern, slightly different from that of the sixth. A different pattern characterised as a type 2 strain was detected in the urine from a Peruvian individual. These results suggest that DGGE analysis could be used as a screening assay for choosing strains for nucleotide sequencing. The analysis of a fragment larger than the one used in this study could allow the identification of more types and subtypes. PMID- 10716344 TI - A viral transmembrane recombinant protein-based enzyme-linked immunosorbent assay for the detection of bovine immunodeficiency virus infection. AB - The expression of bovine immunodeficiency virus (BIV) truncated transmembrane envelope protein (designated hereafter tTM) in insect cells has been described previously (Abed, Y., St-Laurent, G., Zhang, H., Jacobs, R.M., Archambault, D., 1999. Development of a Western blot assay for detection of bovine immunodeficiency-like virus using capsid and transmembrane proteins expressed from recombinant baculovirus. Clin. Diagn. Lab. Immunol. 6, 168-172). In this study, a tTM-based enzyme-linked immunosorbent assay (ELISA) was developed for the serodetection of BIV infection. A total of 109 bovine sera including 86 BIV negative and 23 BIV-positive serum samples were tested. The ELISA results were compared with those of three Western blot assays using, as test antigens, cell culture-derived whole virus proteins (WB1), and the tTM (WB2) and p26 (WB3) fusion proteins expressed from recombinant baculovirus in insect cells, respectively. The concordances of the ELISA results with those of the WB1, WB2, and WB3 were 97.2, 100 and 97.2%, respectively. The tTM protein-based ELISA and Western blot permitted the detection of BIV infection in cattle whose sera failed to react with the p26 fusion protein and the whole virus protein preparation. The tTM recombinant protein was also used to study the kinetics of appearance of antibodies against BIV transmembrane envelope protein in rabbits infected experimentally with BIV. Antibodies to tTM were detected at 28 days post infection and persisted through the entire 36-39.5 months experimental time period. The results of this study showed that the tTM-ELISA might be useful for the serodetection of BIV-infected animals, and for basic studies on BIV replication life cycle. PMID- 10716345 TI - Lessons from a multicentre study of the detectability of viral genomes based on a two-round quality control of GB virus C (GBV-C)/hepatitis G virus (HGV) polymerase chain reaction assay. AB - The aim of this study was to determine whether multicentre quality controls for the detectability of viral genomes could contribute to the improvement of diagnostic performance in the participating laboratories. The study was carried out during two successive rounds, during which 18 laboratories specialized in nucleic acid testing analyzed, through a polymerase chain reaction (PCR) assay, a common panel of GB virus C (GBV-C)/hepatitis G virus (HGV) RNA-positive and negative samples. During the first round, the laboratories used either an 'in house' PCR procedure or a partly standardized commercial test. After decoding the results of the first round, the procedures of the participating laboratories were compared in order to establish a consensus procedure deduced from those of the laboratories which provided the best results. During the second round, each participating laboratory could use the resulting consensus procedure, or its own procedure, or both. The results of this quality control study indicated that, whatever method used, even specialized and trained laboratories may give false negative or false-positive results. The commercial assay did not guarantee a systematic high quality level of results. The striking heterogeneity of results observed among laboratories using the same commercial assay confirm that molecular biology methods need skilled technicians. The results of this quality control study suggest that full standardization of viral genome detection, including all steps of the procedure, is necessary and that the laboratories performing PCR should participate in repeated quality control studies, whatever technique is being used. PMID- 10716346 TI - Use of recombinatory PCR to insert subtle genetic markers into Moloney murine leukemia virus-based retroviral vectors. AB - As tools to examine template switches and recombination events during the process of reverse transcription, two nearly identical Moloney murine leukemia virus based (MoMLV) retroviral vectors were constructed using the technique of recombinatory polymerase chain reaction (PCR). The experimental vectors designed for this study were based on the well-characterized LN series vectors. The protein coding regions normally present in the retroviral genome have been replaced by the coding regions for two drug resistance markers, neomycin phosphotransferase (Neo) and hygromycin phosphotransferase (Hyg). With only one functional drug resistance gene in each vector, the individual vectors as well as recombination events between them can be followed by phenotypic selection. Utilization of recombinatory PCR allowed the insertion of very subtle nucleotide changes resulting in a series of restriction site polymorphisms in the two retroviral vectors. The ability to create these subtle mutations in specific locations of these retroviral vectors allowed the utilization of naturally occurring areas of variability in the vectors and avoid regions important for replication. PMID- 10716347 TI - Laboratory and field studies of an antigen capture ELISA for bluetongue virus. AB - An improved bluetongue antigen capture ELISA (BTACE) technique was evaluated for its ability to detect the full range of 24 bluetongue (BLU) serotypes. The BTACE detected all 24 serotypes in cell culture fluids, including eight serotypes where the representative strains originated from both Australia and also from the South African reference collection. The amount of infectious virus required to obtain a positive BTACE result varied between 100-1000 TCID50. This was approximately 10 fold more sensitive than the antigen capture test described previously (Hosseini, M., Hawkes, R.A., Kirkland, P.D., Dixon, R., 1998. J. Virol. Methods 75, 39-46.). The BTACE method was compared with conventional passage in cell culture to detect the presence of virus in the tissues of embryonated chicken eggs (ECEs) which had been inoculated intravenously with the blood of sheep and cattle infected experimentally with the eight Australian serotypes of BLU (1, 3, 9, 15, 16, 20, 21, and 23). The BTACE method was at least as sensitive as the conventional cell culture detecting virus in ECEs, obviating the need for prolonged cell culture passage to detect the virus. A comparison of the amount of antigen detected in different embryo tissues indicated that liver homogenates gave the highest positive to negative ratios in the BTACE and were selected as the specimen of choice. In studies of sheep infected with all 24 South African reference BLU serotypes this new BTACE was able to detect viraemia with all serotypes. Finally, the BTACE was validated in surveillance programs for BLU in both New South Wales, Australia and in Yunnan Province, People's Republic of China. Blood samples from sentinel cattle were inoculated into ECEs. Homogenised ECE livers were tested by BTACE and those positive were passaged subsequently in cell culture for virus isolation and identification. This protocol led to the efficient isolation of field isolates of many serotypes. The high sensitivity and broad reactivity of the method indicates that it should be valuable for BLU diagnosis and surveillance programs. PMID- 10716348 TI - Rapid phenotypic drug susceptibility assay for HIV-1 with a CCR5 expressing indicator cell line. AB - Phenotypic drug susceptibility assays of human immunodeficiency virus type 1 (HIV 1) isolates generally use time-consuming, expensive assays with peripheral blood mononuclear cells. A new HIV-1 indicator cell line, MAGI-CCR5, has been developed and applied for this purpose. This cell line expresses human CD4, the two major HIV-1 coreceptors, CCR5 and CXCR4, the reporter gene beta-galactosidase driven by the HIV-1 LTR, and quantitates infection within 48 h. A panel of reference strains and primary HIV-1 isolates were all found to infect this cell line. Susceptibility assays with a nucleoside (zidovudine, ZDV) and a non-nucleoside reverse transcriptase inhibitor (nevirapine, NVP) were performed with reference and primary isolates. The assay was modified into two steps for protease inhibitor (indivinavir, IDV and ritonavir, RTV) susceptibility assays. Primary isolates derived from drug naive patients displayed mean baseline 50% effective concentrations (EC50) of 0.14 microM for ZDV, 0.33 microM for NVP, and 0.02 microM for IDV. Isolates derived from patients under treatment displayed increased EC50 concentrations. The MAGI-CCR5 cell line offers a rapid, efficient, and reproducible method of testing a wide range of HIV-1 isolates for drug susceptibility. PMID- 10716349 TI - Disinfection potential of electrolyzed solutions containing sodium chloride at low concentrations. AB - Electrolyzed products of sodium chloride solution were examined for their disinfection potential against hepatitis B virus (HBV) and human immunodeficiency virus (HIV) in vitro. Electrolysis of 0.05% NaCl in tap water was carried out for 45 min at room temperature using a 3 A electric current in separate wells installed with positive and negative electrodes. The electrolyzed products were obtained from the positive well. The oxidation reduction potential (ORP), pH and free chlorine content of the product were 1053 mV, pH 2.34 and 4.20 ppm, respectively. The products modified the antigenicity of the surface protein of HBV as well as the infectivity of HIV in time- and concentration-dependent manner. Although the inactivating potential was decreased by the addition of contaminating protein, recycling of the product or continuous addition of fresh product may restore the complete disinfection against bloodborne pathogens. PMID- 10716350 TI - Flow cytometric detection of viruses. AB - Representatives from several different virus families (Baculoviridae, Herpesviridae, Myoviridae, Phycodnaviridae, Picornaviridae, Podoviridae, Retroviridae, and Siphoviridae) were stained using a variety of highly fluorescent nucleic acid specific dyes (SYBR Green I, SYBR Green II, OliGreen, PicoGreen) and examined using a standard flow cytometer equipped with a standard 15 mW argon-ion laser. The highest green fluorescence intensities were obtained using SYBR Green I. DNA viruses with genome sizes between 48.5 and 300 kb could easily be detected. The fluorescence signals of the small genome-sized RNA viruses (7.4-14.5 kb) were found at the limit of detection. No significant linear relationship could be found between genome size and the green fluorescence intensity of the SYBR Green I stained virus preparations. To our knowledge, this is the first report of detecting and discriminating between a wide range of different viruses directly using flow cytometry. This rapid and precise assay represents a new and promising tool in the field of virology. PMID- 10716351 TI - Immunological analysis of the tegument phosphoprotein ppUL83 of human cytomegalovirus. AB - Immunological properties of the tegument phosphoprotein, ppUL83, of human cytomegalovirus (HCMV), expressed using a replication deficient recombinant adenovirus vector (RAd83) are described. The initial characterisation of this protein was carried out by immunofluorescence (IF), immunoprecipitation (RIP) and immunoblotting using nine mouse monoclonal antibodies (Mabs) directed against five linear and four conformational epitopes of ppUL83. The reactivity of the recombinant protein with the Mabs was similar to that observed with native ppUL83, although, the kinetics of its expression was in agreement with expression derived from the HCMV major immediate early promoter (MIEP). The recombinant antigen was used successfully in an Enzyme Immunoassay (EIA) for the detection of IgG class antibodies in 171 sequential sera taken from 21 heart transplant recipients. Comparison of HCMV-infected and RAd83-infected cell extracts in this experiment showed that recombinant antigen could substitute whole virus extracts as a single well-characterised protein in EIA. Serum IgG avidity measurements, using the recombinant ppUL83, differentiated between primary and past HCMV infections in the population studied. PMID- 10716352 TI - Validation of the specific isotype assay to detect antibodies against foot-and mouth disease virus in bovine milk. AB - The specific isotype assay (SIA) detects IgG1 against foot-and-mouth disease (FMD) virus in bovine milk. A strong correlation was demonstrated between milk antibody titres, and those in serum as measured by the liquid phase blocking ELISA. Thus the SIA would be useful on a herd basis to monitor the milk of vaccinated cattle to determine when re-immunisation is advisable. The SIA titration ELISA was then simplified to a single dilution test and optimised to differentiate the reactions in the milk of FMD-naive cows from those in animals which had been infected with FMD or vaccinated against the disease. For milk from immunised cattle, the pH of the sample was important and borderline positive specimens with a pH of 6.0 or below gave negative results. For milk from naive animals, the optical density (OD) registered in the SIA varied according to the time of year that samples were collected which, in turn, influenced the OD above which milks might be considered positive. Studies showed that the pH of milk could be maintained within the range suitable for the SIA by either storing for up to 1 week at 4 degrees C or by freezing at -20 degrees C for an indefinite period. PMID- 10716353 TI - Parameters that affect in vitro splicing of bovine papillomavirus type 1 late pre mRNAs. AB - During the course of study on regulated viral pre-mRNA splicing, an in vitro RNA splicing assay was developed to analyze how an exonic splicing enhancer stimulates splicing of bovine papillomavirus type 1 (BPV-1) late pre-mRNAs. The optimal concentration of HeLa nuclear extract (HNE) in a standard RNA splicing reaction depends on the individual substrate pre-mRNA. Splicing of a BPV-1 late pre-mRNA required 40% HNE and 2 h incubation at 30 degrees C. Higher HNE was detrimental to splicing and longer incubation times lead to RNA degradation. In the reaction containing 40% EDTA-treated HNE, 1.5-3.0 mM Mg2+ or 3 mM Mn2+, but not Co2+, were required to catalyze an efficient splicing reaction. Surprisingly, EDTA-untreated HNE in the absence of exogenous Mg2+ catalyzed very efficiently splicing of the RNA. Addition of Mg2+ from 0.1 to 0.5 mM, only enhanced slightly the splicing in EDTA-untreated HNE and excessive Mg2+ concentration (above 1.5 mM) in the reaction resulted in production of aberrant splicing products or intermediates. In contrast, addition of Mn2+ to EDTA-untreated HNE severely suppressed splicing. In addition, it was observed that the RNA transcribed from vector sequences downstream of the polylinker region of pSP72 vector, when connected to the 3' terminus of chimeric Drosophila doublesex-BPV-1 SE1 pre mRNAs, suppressed dramatically splicing. RNA transcribed from the pSP72 polylinker region, when supplied in trans, also suppressed splicing. These results suggest that a DNA template used to make RNA transcripts should avoid these sequences as much as possible. PMID- 10716354 TI - Historical review of the use of silver in the treatment of burns. I. Early uses. PMID- 10716355 TI - A historical review of the use of silver in the treatment of burns. II. Renewed interest for silver. AB - In 1965, Moyer revived interest in silver nitrate solution. He concluded on the basis on in vitro and in vivo studies that a 0.5% solution represented the lowest concentration at which antibacterial action (against Staphylococcus aureus, haemolytic streptococci and generally against Pseudomonas aeruginosa and E. coli) was obtained. Mafenide acetate was introduced a short time after the reintroduction of silver nitrate, followed a few years later by silver sulphadiazine. Thus, in a short period of time three medicaments appeared on the market which represented a radical change in the topical treatment of burns. The action of silver sulphadiazine has been intensively studied. Since silver sulphadiazine does not offer sufficient protection to prevent or retard the growth of gram-negative bacteria in patients with burns covering more than 50% of body surface, Monafo introduced the combined preparation silver sulphadiazine and cerium nitrate. Although various attempts have been made to develop more effective silver compounds, so far silver sulphadiazine still remains the most widely used substance of this type. PMID- 10716356 TI - Bound and soluble adhesion molecule and cytokine levels in patients with severe burns. AB - We measured endotoxins, inflammatory cytokines and soluble adhesion molecules in the blood of 17 severe burn patients to determine the involvement of these factors in the pathophysiology of severe burns. All seventeen patients had burns with a total burn surface area of 20% or more and a burn index of 15% or more. Endotoxin was measured by an endotoxin-specific assay and tumor necrosis factor alpha, interleukin 6 and interleukin 8 and soluble adhesion molecules were measured by enzyme-linked immunosorbent assay. CD11a, CD11b and CD18, measured by flow cytometry, were elevated in the non-surviving group, the septic shock group and the multiple organ dysfunction syndrome group, suggesting a close connection between these adhesion molecules and burns complicated by infection. Soluble adhesion molecules were found to indirectly reflect the level of endothelial cell adhesion molecules, suggesting that inflammatory cytokines may also be involved in their production. PMID- 10716357 TI - Role of mast cells in the pathogenesis of postburn inflammatory response: reactive oxygen species as mast cell stimulators. AB - Thermal trauma has a direct effect on mast cells, triggering the secretion of histamine. This secretion leads to an enhanced xanthine oxidase activity and an increased production of reactive oxygen species (ROS), the latter being produced after burns through differing mechanisms. As ROS have been shown to have deleterious effects on cellular membranes, a lesion of the mast cell membrane could close the circle of autoinjury due to the vasoactive actions of mast cell mediators. Our studies were designed to assess the potentiality of ROS as stimulators of mast cell degranulation after burns by comparing two groups of rats treated, respectively, with SOD and saline solution after a scald injury. Plasma levels of tryptase and histamine were analyzed as markers of mast cell activity. A comparison of the mean increases of tryptase between baseline and 3-h postburn levels in the two groups shows significant differences (p < 0.001) (control: 0.13+/-0.04, SOD: 0.03+/-0.01). When comparing the mean increases between the baseline and 3 h postburn levels of histamine in the two groups, significant differences were also found (p < 0.001) (control group: 2.70+/-0.57. SOD group: 1.22+/-0.32). The lower levels of histamine and tryptase induced by SOD provides indirect evidence that ROS are involved in the process, causing the release of such mediators by mast cells, which may in turn suggest that ROS can act as stimulators of mast cell degranulation in burns. PMID- 10716358 TI - An experimental study on systemic inflammatory response syndrome induced by subeschar tissue fluid. AB - OBJECTIVE: To investigate the biological activity of subeschar tissue fluid (STF) and its probable mechanism in the genesis of systemic inflammatory response syndrome(SIRS). METHODS: The changes of heart rate (HR), respiratory rate (RR), white blood cell count (WBCC) as well as the major organ function were observed in the animals injected with STF. The inflammatory mediators TNF-alpha and IL-1 in the supernatants of macrophages cultured with STF were assayed. RESULTS: The HR, RR and WBCC were elevated in animals after injection with STF. STF showed a deleterious effect on function and structure of the major visceral organs. Macrophages were activated to produce excessive TNF-alpha and IL-1. CONCLUSION: The findings suggest that STF may be one of the inducing factors involved in the genesis of SIRS and the development of MODS in the early postburn stage. PMID- 10716359 TI - The inter-rater reliability of estimating the size of burns from various burn area chart drawings. AB - The accuracy and variability of burn size calculations using four Lund and Browder charts currently in clinical use and two Rule of Nine's diagrams were evaluated. The study showed that variability in estimation increased with burn size initially, plateaued in large burns and then decreased slightly in extensive burns. The Rule of Nine's technique often overestimates the burn size and is more variable, but can be performed somewhat faster than the Lund and Browder method. More burn experience leads to less variability in burn area chart drawing estimates. Irregularly shaped burns and burns on the trunk and thighs had greater variability than less irregularly shaped burns or burns on more defined anatomical parts of the body. PMID- 10716360 TI - A burn prevention program as a long-term investment: trends in burn injuries among Jews and Bedouin children in Israel. AB - In order to broaden our long-term intervention efforts in elementary schools in Israel (underway since 1988) and to set priorities for further population specific actions, we compared the pattern of burn injuries among two age groups (0-4; 5-14) of two ethnic groups of Jews and Bedouins admitted to a regional hospital between 1986 and 1995 (n = 1050). The findings indicated a significant downward trend, though somewhat nonlinear, in burn admissions among the older age groups. A relatively less favorable trend was observed for the younger age groups. Consistently across years, burn rates in the younger group of Bedouin children were the highest. For the 10-year period, a significant season by ethnic group variation in burn admissions was observed, with a peak in the spring and in the wintertime for the Jews and Bedouins, respectively. A significant trend of decrease, mostly among older children, in average lengths of hospital stay, was also evident. Yet, regardless of age group and across years, Bedouin children stayed longer in the hospital than Jewish children. The overall leading causes of injury (for 1992-1995) were hot liquids (69%), fire (17%), chemicals (9.5%) and contact (2%). In our view, there is a need to address at-risk populations through environmental, community and family-oriented interventions and to venture beyond the pathogenic factors to the investigation of the salutary factors of health under diverse life conditions. PMID- 10716361 TI - Bacteriology of burn wounds in Enugu, Nigeria. AB - A retrospective study of bacterial infection in 71 burned patients over a 5-year period (1993-1997) was carried out. The commonest colonizing organism was Klebsiella species (26.7%) followed by Staph aureus (25.6%). There was a very high degree of resistance by these organisms to commonly available antibiotics in Nigeria, with the result that more expensive antibiotics such as the cephalosporins were required. The poor socioeconomic condition of most of the patients was a very important pre-disposing factor to burn wound infection, as only 25% of patients were able to afford the cost of wound microscopy and culture, thus leading to limited numbers of cultures being performed, the result being their prescription of antibiotics was made generally on an empirical basis. Restriction in the misuse of antibiotics and establishment of an infection control until will help to lower the incidence of infection. PMID- 10716362 TI - The role of a 'satellite-service' in the national organisation of burn care in the Sultanate of Oman. AB - Khoula hospital, one of the major tertiary care hospitals, situated in the capital city, is the national trauma centre of the Sultanate of Oman. The burns unit at Khoula hospital has developed an unique collaborative service of active participation in the initial management of burns patients admitted to the peripheral hospitals of the country. The description 'Satellite-service' is used in this paper to describe this 'remote-control service' offered by the main burns center, for the management of burned patients admitted the various peripheral hospitals (the 'satellites') throughout the country, during the shock-period of the first 36 hour post-burn. Thereafter the patients are transferred to the main center for further management. This system was instigated with a common protocol for the initial management of burns all over the country in 1981. Central registration of burns patients from peripheral hospitals was started in 1994. Retrospective analysis of the statistics and records of 151 burns patients transferred from 19 peripheral hospitals to Khoula burns center during a 3 year period from September 1994 to August 1997, showed that the satellite service is very effective in the initial management of patients during the shock period, as there was no mortality due to hypovolemia. The advantages of this service include uniform and appropriate 'initial' management of all the burns patients admitted to any of the hospitals of the country. The major disadvantages of the system were the increased work-load of the main center and poor patient-compliance with post-discharge instructions. This paper also describes how the Satellite service works in practice. PMID- 10716363 TI - Innovations in flap design: modified groin flap for closure of multiple finger defects. AB - The groin flap is frequently used for covering soft-tissue defects of the hand. It is normally utilised as a single unit to cover the defect. When used for coverage of multiple digital defects, it requires syndactylisation of the digits with a further procedure to divide the syndactylised digits some time after division of the main pedicle. We report a new technique of fashioning 'daughter flaps' from the groin flap at the time of elevation, and their use to cover full thickness burns to the dorsum of PIP joints in both hands thus avoiding the need for syndactylisation of the digits. PMID- 10716364 TI - Effects of tracheostomies on infection and airway complications in pediatric burn patients. AB - Considerable controversy exists as to whether tracheostomy is ever indicated in burn patients. New advents in the treatment of inhalation injury have improved survival, making the use of tracheostomy more usual. The purpose of this study was to analyze the outcome of tracheostomies, and the effect of time on complications. Patients requiring ventilatory support and tracheostomies were studied. Demographic data, hospital course, ventilatory parameters and complications were analyzed. Two hundred ninety patients required ventilation and 36 tracheostomy. Mean percentage of TBSA burned was 59%+/-4. Ninety percent of these patients presented with inhalation injury. Mortality in tracheostomy patients was 25 and 16% in all ventilated patients. Thirty-five percent of the patients developed late complications. Patients who had their airway converted to tracheostomy before day 10 postinjury had a significantly lower incidence of subglottic stenosis. and patients who required airway pressures over 50 cm H2O for more than 10 days had a significantly higher incidence of tracheomalacia. Pneumonia occurred at similar incidence in ventilated and tracheostomy patients. The mortality and late complications of pediatric burn patients with tracheostomy has decreased over the last decade. They do not present with higher incidence of pneumonia. Maintenance of airway pressures below 50 cm H2O and conversion of the artificial airway to tracheostomy before day 10 postinjury may be advisable in patients requiring long term ventilation to prevent late complications. PMID- 10716365 TI - Herpes simplex virus infection in a paediatric burn patient: case report and review. AB - Herpes simplex virus (HSV) infection in the burn patient is thought to occur relatively frequently. Most commonly, children with significant burns, particularly involving the head and neck, are affected. Burn related immunosuppression is thought to allow reactivation of latent HSV in most cases, although primary HSV infection has been recognized. Clinical manifestations vary from asymptomatic viral shedding, to prolonged fever with eruption of vesicles, to rare cases of systemic visceral dissemination. Healing partial thickness wounds and donor sites are most prone to infection. Laboratory confirmation of HSV infection relies on direct demonstration of the virus and/or observation of a rise in antibody titer. Treatment of an established HSV infection includes use of IV Acyclovir, meticulous wound care, and efforts to prevent nosocomial spread. The vast majority of cases resolve without sequelae unless complicated by systemic, multiorgan HSV infection. PMID- 10716366 TI - Cannula related suppurative thrombophlebitis in the burned patient. AB - Suppurative thrombophlebitis is a well recognised and potentially fatal complication of intravenous cannulation in burns patients. We report a case of an Afro-Caribbean patient with noninsulin-dependent diabetes who developed signs of systemic sepsis two weeks after a 14% total body surface area flame burn. Despite an initial paucity of clinical signs at the cannulation site, exploratory venotomy revealed frank suppuration within the long saphenous vein from the ankle to the groin. This was treated successfully by total excision of the vein and its tributaries and delayed wound closure. Following this, a retrospective analysis of the measured clinical parameters and blood tests revealed no obvious, missed pointers to the impending sepsis other than a dramatic increase in the overall daily insulin requirement. This had doubled over a 48-h period, preceding the clinical diagnosis by three days. The relevant literature and guidelines for management are reviewed. PMID- 10716367 TI - Characteristics of bath-related burns in Japan. PMID- 10716368 TI - Alloimmunization as a strategy for vaccine design against HIV/AIDS. PMID- 10716369 TI - Predominance of subtype A and G HIV type 1 in Nigeria, with geographical differences in their distribution. AB - The purpose of this study was to generate data on the relative prevalences of the HIV-1 subtypes circulating in Nigeria. A total of 252 HIV-1-positive samples collected during an epidemiologic survey conducted in April 1996 were genetically characterized by HMA (heteroduplex mobility assay) and/or sequencing. Samples were collected in Lagos, Calabar, Kano, and Maiduguri. Overall, the predominant env subtypes were A (61.3%) and G (37.5%). Subtype A is more prevalent in the south (p < 0.001), about 70% in Lagos and Calabar, whereas a quarter of the samples was classified as subtype G in these states. In contrast, subtype G is predominant in the north ( < 0.001), representing 58% of the samples in Kano. In the northeastern region, Maiduguri, almost similar proportions of subtype A and G were seen, 49 and 47.4%, respectively. A total of 37 samples was also sequenced in the p24 region from the gag gene; 13 (35%) had discordant subtype designations between env and gag. The majority of the gag (12 of 17) and env (14 of 22) subtype A sequences clustered with the A/G-IBNG strain. Within subtype G, three different subclusters were seen among the envelope sequences. These different subclusters are observed among samples obtained from asymptomatic individuals and AIDS patients from the four Nigerian states studied. In conclusion, we observed a limited number of HIV-1 subtypes circulating in Nigeria, with subtypes A and G being the major env subtypes responsible for the HIV-1 epidemic. Nevertheless, the high rate of recombinant viruses (A/G) and the different A/G recombinant structures indicate a complex pattern of HIV-1 viruses circulating in this country. PMID- 10716370 TI - Genetic evidence of multiple transmissions of HIV type 1 subtype F within Romania from adult blood donors to children. AB - We studied the phylogeny of HIV-1 subtype F viruses from children and adults in Romania in order to (1) clarify whether the Romanian subtype F epidemic was caused by one or several virus introductions and (2) gain insight into the route of spread of the HIV-1 subtype F virus among children and adults in Romania. env (V3), gag (p17/half p24), and pol (prot/half RT) sequences were obtained from three districts in Romania: Tirgu Mures (n = 9, children), Craiova (n = 15, children), and Bucharest (n = 13, adults). Of 37 HIV V3 sequences from Romania, 35 belonged to the genetic subtype F in the neighbor-joining tree, whereas 2 sequences from adults clustered with subtypes A and C. Within the subtype F cluster, no bootstrap-supported subclusters were observed according to geographic area in Romania. Two of the adult V3 sequences that clustered with the children were obtained from individuals who tested HIV seropositive in 1989 and 1990, showing that the subtype F virus was present among adults when the HIV epidemic began among children in Romania. The HIV-1 subtype F viruses obtained from children showed a mean pairwise V3 nucleotide distance of 7.9% and maximum distances of between 18 and 19%; both are higher than previously described. The mean V3 distances (overall, synonymous, and nonsynonymous) were significantly higher for adults than for children. One V3 sequence from the Democratic Republic of Congo clustered within the Romanian sequences, suggesting that the subtype F virus in Romania may originate from this area. Our data also suggest that HIV-1 subtype F was present among Romanian adults before it appeared in 1989 among institutionalized children. The juvenile population was most likely infected with the HIV-1 subtype F virus on more than one occasion, presumably through HIV contaminated blood (products) obtained from adults. PMID- 10716371 TI - Evaluation of a lipopeptide immunogen as a therapeutic in HIV type 1-seropositive individuals. AB - A 32-amino acid HIV-1 Gag immunogen was assessed for its ability to augment existing virus-specific CTL responses in chronically HIV-1-infected individuals. The immunogen was an HIV-1 synthetic lipopeptide conjugate composed of an N palmitoyl-S-[2,3-bis(palmitoyloxy)-(2R)-propyl-N-(R)-cysteinyl] group covalently coupled to a synthetic 32-amino acid Gag peptide containing at least 5 CTL epitopes known to be restricted by HLA-A33, -B8, -B27, -B35, and -Bw62. This potential immunotherapeutic was first determined to be safe in six HIV-1 seropositive subjects, with no adverse clinical effects noted during a 182-day period after administration of a dose of 350 microg. The immunogenicity of this lipopeptide conjugate was then assessed in a pilot study in nine HIV-1 seropositive volunteers with peripheral blood CD4+ lymphocyte counts of >500/microl. Three groups of individuals were studied: HLA-selected subjects who received 350 microg of the immunogen on days 0, 28, and 56 (four subjects); HLA selected subjects who received a placebo according to a similar inoculation schedule (three subjects); and HLA-mismatched subjects who received the experimental immunogen (two subjects). All subjects were monitored for 26 weeks. After treatment, PBLs from two of the four HLA-selected subjects who received the experimental immunogen showed a transient increase in Gag peptide-specific bulk CTL activity. None of the placebo-vaccinated or vaccinated HLA-mismatched subjects showed any change in bulk Gag peptide-specific CTL activity. However, no consistent decrease in plasma HIV-1 RNA levels was noted in any of the subjects. The present study illustrates that this peptide formulation may not be a sufficiently potent immunogen to significantly augment HIV-1-specific CTLs and to decrease virus load in HIV-1-seropositive individuals. PMID- 10716372 TI - Interleukin 18 and interleukin 1beta production is decreased in HIV type 1 seropositive hemophiliacs but not in HIV type 1-seropositive nonhemophiliacs. AB - In Japan, the proportion of hemophiliacs infected with human immunodeficiency virus type 1 (HIV-1) is 40%, whereas more than 90% are infected with hepatitis C virus (HCV). To evaluate the immunological status of hemophiliacs infected with HIV-1, we investigated the pattern of cytokine production in peripheral blood mononuclear cells (PBMCs) of HIV-1-seropositive and -seronegative hemophiliacs, HIV-1-seropositive non-hemophiliacs, and healthy individuals. The production of IL-18 and IL-1beta from PBMCs stimulated with Staphylococcus aureus Cowan strain 1 (SAC) in the HIV-1-seropositive hemophiliacs was significantly decreased in comparison with the other groups. On the other hand, IL-12 production in both HIV 1-seropositive groups was significantly lower than in HIV-1-seronegative groups. TNF-alpha and IL-6 production was similar among the four groups. In contrast, plasma levels of TGF-beta1 were increased in HIV-1-seropositive hemophiliacs, HIV 1-seropositive nonhemophiliacs, and HIV-1-seronegative hemophiliacs, with the highest levels being in HIV-1-seropositive hemophiliacs, suggesting that coinfection with HIV-1 and HCV increases the level of plasma TGF-beta in HIV-1 seropositive hemophiliacs. Treatment of PBMCs from healthy individuals with TGF beta1 inhibited IL-18 and IL-1beta production without affecting IL-6, IL-10, or TNF-alpha production. Suppression of the expression of caspase 1 mRNA, which is known to be an IL-1beta-converting enzyme and which also cleaves the precursor of IL-18, was observed in the SAC-stimulated PBMCs from healthy individuals after treatment with TGF-beta1 and in the SAC-stimulated PBMCs from HIV-1-seropositive hemophiliacs, suggesting that the decreased production of IL-18 and IL-1beta in HIV-1-seropositive hemophiliacs may be related to the downregulation of caspase 1 mRNA induced by high levels of TGF-beta1 in plasma. PMID- 10716373 TI - Increased infectivity of HIV type 1 particles bound to cell surface and solid phase ICAM-1 and VCAM-1 through acquired adhesion molecules LFA-1 and VLA-4. AB - HIV-1 incorporates a variety of host membrane proteins during budding. We have previously shown that adhesion molecules are acquired by the virus in their activated or functional states. Our studies and those of others indicate that adhesion molecules can have profound effects on virus infectivity and its resistance to neutralization by antiviral antibodies. In this study we have examined the effect on infectivity of immobilization or margination of HIV-1 through acquired integrins LFA-1 and VLA-4 onto nonsusceptible cells and solid phase adhesion ligands (ICAM-1 and VCAM-1, respectively). LFA-1- and VLA-4 mediated HIV-1 binding was supported by ICAM-1 and VCAM-1 immunoglobulin Fc chimeras, respectively. Integrin-mediated HIV-1 binding was also supported by 293 cells transfected with ICAM-1. In both cases the specificity of binding was confirmed with the appropriate blocking monoclonal antibodies or soluble adhesion ligands. We used a sensitive single-cycle infection assay based on a cell line expressing an LTR-luciferase cDNA construct to compare the infectivity of bound virus with that of free virus. Our results show that the binding of HIV-1 to nonsusceptible cells or immobilized adhesion ligands through acquired integrins can increase its infectivity by as much as two orders of magnitude. These results have implications for in vivo dissemination and transmission of HIV-1 and may also explain the high level of virus replication seen in solid lymphoid organs. PMID- 10716374 TI - HIV virions and HIV infection in vitro are unaffected by human granzymes A and B. AB - Granzymes are a family of serine proteinases commonly found in the granules of CD8+ T cells. In HIV infection, CD8+ cells show cytotoxic and noncytotoxic antiviral activities. The latter is mediated, at least in part, by a secreted CD8+ cell antiviral factor, CAF. Because of the antiviral nature of CD8+ cells, we examined the potential anti-HIV activity of free granzymes that can be found in CD8+ cell culture fluids. Pretreatment of CD4+ T cells with granzyme A or granzyme B had no effect on their susceptibility to infection with HIV, nor did incubation of the granzymes with HIV virions alter their infectivity. Continuous culture of acutely infected CD4+ T cells with granzyme A or B showed no effect on cell viability or the replication of HIV. The findings of this study suggest that free granzymes do not control HIV infection and spread in CD4+ T cells. PMID- 10716375 TI - Pertussis toxin enhances human immunodeficiency virus type 1 replication. AB - Pertussis toxin (PTX) has been used as a reagent to identify involvement of the G protein-mediated signal transduction pathway. In this study, we found that PTX enhanced HIV-1 replication in acute infection systems at a high dose (1-10 microg/ml) in vitro. PTX treatment enhanced the infectivity of HIV-1-based pseudovirus enveloped with HIV-1 or amphotropic murine leukemia virus (A-MuLV), but not with vesicular stomatitis virus (VSV). This high dose of PTX treatment did not affect HIV-1 gene expression. These data suggested that the effect was virus envelope dependent and that PTX acted on an early stage of viral infection. Treatment with B-oligomer, a nonenzymatic subunit of PTX, mimicked this enhancing effect of PTX. However, desialylation of viral and cellular surface glycoproteins, which are receptors for B-oligomer, did not affect the augmentation induced by PTX. These results indicate that the enhancement of HIV-1 replication is mediated through an unknown biological function of B-oligomer. PMID- 10716376 TI - Generation of CD8+ T cell-generated suppressor factor and beta-chemokines by targeted iliac lymph node immunization in rhesus monkeys challenged with SHIV 89.6P by the rectal route. AB - The targeted lymph node (TLN) immunization strategy was investigated in macaques, in order to determine the efficacy in generating secretory, systemic, and cellular immune responses, CD8+ T cell-generated suppressor factors, and beta chemokines. TLN immunization of the rectal and genital mucosa-associated iliac lymph nodes (TILNs) was compared with axillary TLN immunization (TAxLN) using HIV 1 MN/LAI gp140env and SIV p27gag in alum. Significantly higher immune responses, as well as CD8+ T cell-generated anti-SIV factors and the beta-chemokines RANTES, MIP-1alpha, and MIP-1beta, were elicited by iliac as compared with axillary TLN immunization. The immune responses induced by TLN immunization were examined for their capacity to prevent rectal mucosal infection by the pathogenic dual-tropic SHIV-89.6P. Despite significant secretory, serum, cellular, and beta-chemokine responses, the macaques were infected by SHIV-89.6P. Whether the lack of protection was associated with the antigenic unrelatedness of SHIV-89.6P to the immunizing HIV-1 MN/LAI gp140 or to the virus utilizing CXCR4 to a much greater extent than CCR5, remains to be determined. PMID- 10716377 TI - Ilizarov bone transport treatment for tibial defects. AB - OBJECTIVES: To evaluate the results and complications of Ilizarov bone transport in the treatment of tibial bone defects. DESIGN: Retrospectively reviewed consecutive series. METHODS: Nineteen patients with tibial bone defects were treated by the Ilizarov bone transport method. The mean bone defect was ten centimeters, and there were eight soft-tissue defects. The mean external fixation time was sixteen months. Ten patients required debridement of the bone ends and/or bone grafting of the docking site at the end of transport. RESULTS: Union was achieved in all cases. One refracture of the docking site required retreatment with the Ilizarov apparatus to achieve union. There was one residual leg length discrepancy greater than 2.5 centimeters and two angular deformities greater than 5 degrees. There were no recurrent or residual infections. Seven of the eight soft-tissue defects were closed by soft-tissue transport; the eighth required a free-vascularized flap. The bone results were graded as fifteen excellent, three good, and one fair. The functional results were graded as twelve excellent, six good, and one poor. There were twenty-two minor complications, sixteen major complications without residual sequelae, and three major complications with residual sequelae. To treat the bone defect and the complications, a mean of 2.9 operations per patient was required. CONCLUSIONS: Our results compare favorably with those for other methods of bone grafting as well as with those from other published accounts of the Ilizarov method, especially considering the large defect size in this series. The main disadvantage of the Ilizarov method is the lengthy external fixation time. PMID- 10716378 TI - Mechanical strength of fracture callus in osteopenic bone at different phases of healing. AB - OBJECTIVES: To quantify and compare peak bending force and stiffness of fractured femurs during healing of ovariectomized (OVX) and sham-operated (SHAM) rats. DESIGN: Temporal biomechanical animal study. SETTING: Rat femurs were fractured and surgically fixed by a qualified surgeon. The inherent instability of the fixation system employed produced delayed union of the fracture. All biomechanical assessments were performed with servohydraulic test machines (Instron Inc., Canton, MA, U.S.A.; and MTS Corp., Eden Prairie, MN, U.S.A.). INTERVENTION: OVX was performed sixteen weeks before femur fracture, and the effect of OVX on healing fractures was determined. MAIN OUTCOMES: Peak bending force and stiffness of the healing femurs at four, six, and eight weeks after fracture. RESULTS: Peak bending loads of the healing fractured femurs in the OVX and SHAM animals were not significantly different. Peak bending loads for the OVX animals at four and six weeks were significantly lower than the peak load at eight weeks (p < 0.05), whereas no difference was found in the peak load with respect to time for the SHAM animals. Both SHAM and OVX animals had greater bending stiffness of the healing fractured femur after eight weeks of healing than at four weeks (p < 0.05). CONCLUSIONS: OVX is known to reduce cancellous bone mass and strength, but the effect of OVX on healing of fractures in cortical bone is controversial. This study, using a delayed-union model, found no significant differences between OVX and SHAM animals in the breaking strength of healing fractures. PMID- 10716379 TI - Effects of pulsed electromagnetic fields on bone healing in a rabbit tibial osteotomy model. AB - OBJECTIVE: The purpose of this study was to determine the effect of pulsed electromagnetic field (PEMF) exposure on healing tibial osteotomies in New Zealand White rabbits. DESIGN: One-millimeter Gigli saw osteotomies were stabilized by external fixation. One day after surgery, rabbits were randomly assigned to receive either no exposure (sham control) or thirty minutes or sixty minutes per day of low-frequency, low-amplitude PEMF. Radiographs were obtained weekly throughout the study. Rabbits were euthanized at fourteen, twenty-one, or twenty-eight days, and tibiae underwent either destructive torsional testing or histologic analysis. To determine the baseline torsional strength and stiffness of rabbit tibiae, eleven normal intact tibiae were tested to failure. RESULTS: Sixty-minute PEMF-treated osteotomies had significantly higher torsional strength than did sham controls at fourteen and twenty-one days postoperatively. Thirty minute PEMF-treated osteotomies were significantly stronger than sham controls only after twenty-one days. Normal intact torsional strength was achieved by fourteen days in the sixty-minute PEMF group, by twenty-one days in the thirty minute PEMF group, and by twenty-eight days in the sham controls. Maximum fracture callus area correlated with the time to reach normal torsional strength. CONCLUSION: In this animal model, low-frequency, low-amplitude PEMF significantly accelerated callus formation and osteotomy healing in a dose-dependent manner. PMID- 10716380 TI - Retrograde nailing of humeral shaft fractures: a biomechanical study of its effects on the strength of the distal humerus. AB - OBJECTIVES: The purpose of this study was to evaluate the loss of strength in the distal humerus that resulted from the creation of two different entry portals used for retrograde humeral nailing. DESIGN: Nine pairs, treated as blocks of size two, of fresh frozen humeri from individuals free of musculoskeletal disease were randomly divided into three groups in a balanced incomplete block experimental design. INTERVENTION: The specimens were tested intact (control) or with an entry portal drilled in either the distal metaphyseal triangle or the proximal slope of the olecranon fossa. Two of these three conditions were applied to each pair of the bones. Therefore, three pairs of bones accommodated testing of all possible combinations of the two treatments. All specimens were tested in torsion at a rate of 30 degrees per second until failure or fracture. RESULTS: The creation of an entry portal reduced the ultimate torque to 63 percent of that of the intact specimens (p = 0.044) and the energy absorbed to failure to 27 percent of that of the intact specimens (p = 0.039). The metaphyseal entry portal reduced the torque to failure to 71 percent of that of the intact specimens (p = 0.143) and the energy absorbed to failure to 37 percent of that of the intact specimens (p = 0.073), The olecranon fossa entry portal reduced the torque to failure to 55 percent of that of the intact specimens (p = 0.035) and the energy absorbed to failure to 18 percent (p = 0.058) of that of the intact specimens. CONCLUSIONS: Surgeons should be aware of the loss of strength in the distal humerus after retrograde humeral nailing. This is especially important when prescribing postoperative mobilization in which the upper extremities will be used for weight-bearing in either transfers or ambulation. PMID- 10716381 TI - Dislodgement of the locking cap from the AO unreamed intramedullary nail: a case report. AB - We report on the dislodgement of the proximal locking cap from an AO solid femoral intramedullary nail with spiral blade. The pathologic femoral fracture subsequently went on to heal, and the spiral blade did not back out of the femoral neck. Meticulous intraoperative attention to detail is required to select the correct offset for the proximal cap, because the turning moments of the hip abductor muscles are sufficient to unscrew a prominent cap. PMID- 10716382 TI - Posterior locked lateral compression injury of the pelvis: report of three cases. AB - Lateral compression injuries to the pelvis typically result in a rotationally unstable and vertically stable condition including an impaction and compression fracture of the posterior pelvic ring. The operative and postoperative management, as well as the morbidity and mortality, of these fractures differs significantly from vertical shear injuries to the pelvis, which are characterized by vertical and rotational instability. We report on three unusual lateral compression injuries to the pelvis, resulting in a complete disruption of the pelvic ring with vertical and rotational instability, by definition. Nevertheless, in these patients, locking of the posterior pelvic ring with medial translation of the iliac wing anterior to the sacrum resulted in a pseudostable condition. Their high rate of fracture-related associated injuries and possible complications, as well as the malalignment of the pelvis, required surgical restoration of the pelvic ring. Fracture reduction was successfully performed through an anterior approach in one patient and a posterior approach in two patients; the posterior approach was preferred. Open reduction and internal fixation of these pelvic ring fractures can result in a satisfactory outcome if the associated injuries are successfully dealt with. PMID- 10716383 TI - Ankle fracture with syndesmotic injury. PMID- 10716384 TI - Thrombolytic therapy with use of alteplase (rt-PA) in peripheral arterial occlusive disease: review of the clinical literature. The Advisory Panel. AB - PURPOSE: The clinical literature describing the use of alteplase in the treatment of peripheral arterial occlusive (PAO) disease is reviewed. MATERIALS AND METHODS: The literature database was acquired by a MEDLINE search using the Boolean keyword string: tissue plasminogen activator and/or rt-PA and peripheral not animal. A review was performed to identify the dose range of alteplase, technique of infusion, use of anticoagulation, clinical success rates, and risk of complications. RESULTS: Forty-six clinical studies were identified. There are few prospective, randomized clinical trials and a lack of standardized protocols and endpoints. Use of catheter-directed infusions of recombinant tissue plasminogen activator (rt-PA) may be beneficial versus surgery in the initial management of acute limb ischemia (< 14 days) and in reducing the magnitude of subsequent surgical or percutaneous revascularization. For patients with chronic limb ischemia (> 14 days), irreversible acute limb ischemia, or advanced diabetic arteriopathy, catheter-directed infusion of rt-PA or other plasminogen activators may be unsuitable. The risk of adverse bleeding appears related to the overall dose and duration of infusion. These risks appear similar to those of urokinase. The role of heparin in increasing adverse bleeding during rt-PA therapy is unclear. CONCLUSIONS: There is no generally accepted dose or technique for administering catheter-directed thrombolysis using alteplase; however, several studies have demonstrated its clinical safety and efficacy. Formal studies will be required to determine the optimal dose, technique of infusion, the role of anticoagulation, and complication rates when alteplase is used for PAO disease. PMID- 10716385 TI - Assessment of CO2 arteriography in arterial occlusive disease of the lower extremities. AB - PURPOSE: To compare the diagnostic ability and usefulness of carbon dioxide arteriography with that of angiography using iodinated contrast medium in ischemia of the lower extremities. METHODS: Between April 1997 and February 1998 arteriography was performed systematically in 50 consecutive patients (42 men, eight women; average age, 65 years), who presented with peripheral vascular disease of the arteries of the lower extremities, with use of both CO2 and iodinated contrast medium. Untoward events that occurred during the examinations and the resulting clinical problems were recorded. Subsequently, two radiologists carried out a double-blind evaluation of the images obtained for each segment (aorta, pelvis, thighs, knees, legs, and feet) using the two different contrast agents to diagnose the arterial condition (normal, aneurysm, stenosis, and occlusion). Afterward, the two types of study performed for each patient were compared to assess the overall quality of CO2 arteriography as opposed to arteriography performed with use of iodinated contrast material. RESULTS: Forty eight percent of the patients reported discomfort during the CO2 examinations and 18% of the studies had to be discontinued as a result. When problems relating to poor image quality were included, only 36% of the arteriograms obtained with use of CO2 were complete. Evaluation was possible in only 25% of CO2 studies of the feet. On average, the overall quality of the arteriograms obtained with use of CO2 was insufficient for diagnosis. CONCLUSION: In the authors' experience, CO2 arteriography cannot replace procedures performed with use of iodinated contrast medium for routine examination of ischemia of the lower limbs. In most cases, because of lower tolerance to the procedure and poorer image quality, CO2 imaging was not of sufficient quality to permit diagnosis, particularly at the infrapopliteal level. PMID- 10716386 TI - New hemodynamic test for assessment of failing hemodialysis grafts: the saline infusion test. AB - PURPOSE: To validate the saline infusion test, a new hemodynamic test for assessment of failing hemodialysis access grafts. MATERIALS AND METHODS: Over a 12-month period, 31 procedures were performed in 25 patients with synthetic forearm loop grafts for hemodialysis. Pre- and postangioplasty measurements of static graft pressures and infusion pressures were obtained. For the saline infusion test, graft pressure was measured while saline was infused at a rate of 600 mL/min for 10 seconds with arterial inflow occluded. Comparison was made to percent outflow stenosis as determined with pre- and postangioplasty angiograms. RESULTS: There was no correlation between either the static intragraft pressure (r = .085, P = .654) or the normalized pressure ratio (r = .136, P = .4676) and venous outflow stenosis in the preangioplasty group. When pressure was measured during infusion, a significant Pearson correlation was observed between infusion pressure and percent of angiographic stenosis (r = .60, P = .0002). All three pressure tests were significantly correlated to the percent stenosis identified after angioplasty. CONCLUSIONS: Pressure measured in the graft during the saline infusion test at a standard rate that simulates optimal graft flow correlates with the angiographic degree of stenosis and warrants further investigation as a useful adjunct to the assessment of revascularization results. PMID- 10716387 TI - Primary Gianturco stent placement for inferior vena cava abnormalities following liver transplantation. AB - PURPOSE: To determine the efficacy of primary Gianturco stent placement for patients with inferior vena caval (IVC) abnormalities following liver transplantation. MATERIALS AND METHODS: From August 1996 through March 1999, nine adult patients developed significant IVC abnormalities following liver transplantation. Patients were referred for vena cavography on the basis of abnormal clinical findings, laboratory values, liver biopsy results, Doppler findings, or a combination. Those patients demonstrating a significant caval or hepatic venous gradient were treated with primary Gianturco stent placement. Patients were followed clinically (nine patients), with duplex ultrasound (nine patients), vena cavography (four patients), and biopsy (seven patients). RESULTS: Original pressure gradients ranged from 3 to 14 mm Hg, with a mean of 9 mm Hg. Gradients were reduced to 3 mm Hg or less in all nine patients; presenting signs and symptoms either resolved or improved in eight of nine patients. The ninth patient required repeated transplantation 2 days later. A second patient died 433 days after stent placement of recurrent hepatitis C. Another initially improved following caval stent placement, but underwent repeated transplantation 7 days later due to hepatic necrosis from hepatic arterial thrombosis. Follow-up for the remaining six patients has averaged 491 days, with no clinical, venographic, or ultrasound evidence for recurrent caval stenosis. CONCLUSIONS: Intermediate term results suggest that primary Gianturco stent placement for IVC stenosis, compression, or torsion resulting after liver transplantation is safe and effective. PMID- 10716388 TI - Primary placement of Palmaz long medium stents in transjugular intrahepatic portosystemic shunts. AB - PURPOSE: To describe our results with primary placement of the long-medium Palmaz stent for transjugular intrahepatic portosystemic shunts (TIPS). MATERIALS AND METHODS: Between December 1997 and December 1998 primary placement of long-medium Palmaz stents was performed for TIPS procedures in 17 patients. Patency was determined with ultrasound, angiography, or pathologic examination in the event of transplant. RESULTS: Primary patency was achieved in 13 of 17 patients (76.5%) (follow up, 1-399 days; mean, 99 days). Secondary patency was achieved in 17 of 17 patients (100%) (follow-up, 1-399 days; mean, 110 days). Among the four patients who required revision, the mean time to revision from initial shunt creation was 81 days (range, 13-125 days). Two of these four patients had symptoms of worsening ascites as well as abnormal ultrasound findings prior to their revision; the other two patients were asymptomatic and had abnormal ultrasound findings only. Revisions were performed for intimal hyperplasia within the stent in three of the patients and acute thrombus within the stent in the remaining patient. Kaplan-Meier survival analysis for primary patency yielded mean survival time of 265 days (standard error, 52 days). CONCLUSION: The long medium Palmaz stent is a viable stent for creation of TIPS shunts. PMID- 10716389 TI - Transhepatic mechanical thrombectomy followed by infusion of TPA into the superior mesenteric artery to treat acute mesenteric vein thrombosis. PMID- 10716390 TI - Mechanical versus chemical thrombolysis: an in vitro differentiation of thrombolytic mechanisms. AB - PURPOSE: To assess differing mechanisms of thrombolysis determining time to reperfusion, completeness of thrombus dissolution, and embolic potential. MATERIALS AND METHODS: An in vitro perfusion model designed to mimic arterial flow conditions was created. Bifurcated limbs allowed continuous flow through one channel and the placement of radiolabeled (iodine-125) thrombus housed in a 5-cm segment of polytetrafluoroethylene graft in the other. The three experimental groups consisted of a standard continuous urokinase infusion, a pulsed pressurized injection of saline, and a similar injection with urokinase. A continuous infusion of 5% dextrose served as a control group. Time to reflow (as assessed with ultrasonic flow monitoring), completeness of thrombus dissolution (I-125 liberated into solution), and the number and size of embolic particles produced (detected by a series of graduated filter sizes) were analyzed. RESULTS: Time to reflow was significantly faster for both groups when pressurized injections were used (P < .001). There was no reflow in the control arm at 90 minutes. Completeness of thrombus dissolution was higher when a continuous infusion of urokinase was used in comparison to either of the power injection groups or the control (P < .05). The amount of embolic debris produced was significantly lower with a continuous infusion of urokinase compared with either of the power lysis groups (P < .05), but significantly greater than the control arm (P < .001). The size of the embolic particles in the power pulsed lysis groups was significantly decreased by the addition of urokinase (P < .05). CONCLUSIONS: Reflow is more rapidly established by the use of mechanical means. However, a less complete dissolution of thrombus in conjunction with a greater amount of embolic debris is achieved with this approach. The size of the embolic particles produced is reduced by the addition of a thrombolytic agent. PMID- 10716391 TI - Immunolocalization of proliferating cells in the rabbit iliac artery after balloon angioplasty. AB - PURPOSE: Experiments were performed to characterize the location of proliferating cells in the balloon-dilated rabbit iliac artery. MATERIALS AND METHODS: Balloon angioplasty was performed on the external iliac arteries in each of four rabbits. The arteries were removed 3 days later, frozen, cryosectioned, and immunostained with Ki-67, an antibody that identifies proliferating cells. The sections were then examined to determine the patterns of cell proliferation within the arterial media and the ratio of proliferating to nonproliferating cells. RESULTS: Of the 31 arterial cross-sections examined, cell proliferation was circumferential in five (16%), and focal in 26 (84%). Of the 86 foci of proliferation examined within the 31 cross-sections, proliferation was localized to the inner media in 30 (35%), to the outer media in four (5%), and was transmural in 52 (60%). The internal elastica lamina (IEL) appeared normal at 22 foci (26%), but appeared stretched or torn at 64 (74%). Proliferation was usually confined to the inner media at foci having no IEL injury (18 of 22; 82%), but was most often transmural where the IEL was stretched or torn (49 of 64; 77%). The ratio of proliferating to nonproliferating cells, which averaged 0.31 +/- .20, was greater (P < .01) in areas with IEL injury than in areas without IEL injury. CONCLUSION: These results suggest that angioplasty-induced cell proliferation is typically focal rather than circumferential and is associated with stretching or tearing of the IEL. PMID- 10716392 TI - A practical approach for repositioning flipped venous access ports. PMID- 10716393 TI - Placement of SVC stents over pacemaker wires for the treatment of SVC syndrome. PMID- 10716394 TI - Intravascular occluding device using a modified Gianturco stent as a coil cage. PMID- 10716395 TI - Comparison of technical success and outcome of tunneled catheters inserted via the jugular and subclavian approaches. AB - PURPOSE: To compare the technical success and immediate and long-term outcomes of tunneled central venous catheters placed in comparative cohorts via the subclavian vein (SCV) and the internal jugular vein (IJV) routes. MATERIALS AND METHODS: This was a prospective observational single-center study of consecutive procedures. Between November 1993 and June 1995, 99 catheters were placed via the SCV and between December 1997 and July 1998, 109 catheters were placed via the IJV. Procedural data were recorded in both cohorts by completion of a proforma by the primary operator. RESULTS: Follow-up data were available in 96% of the SCV and 87% of the IJV cohorts. The average procedure time was significantly shorter in the IJV group and technical success was 100% versus 97% in the SCV group, but this did not reach statistical significance. The procedure-related pneumothorax rate and the rate of symptomatic venous thrombosis were significantly lower in the IJV cohort (P = .023, P = .015). Fewer catheters were removed prematurely due to sepsis in the IJV group (P = .043). CONCLUSIONS: The IJV route is associated with comparable technical success, and lower major procedural complication and venous thrombosis rates, with fewer catheters removed prematurely. The right IJV approach with ultrasound guidance is recommended as the route of choice for the placement of tunneled central venous catheters. PMID- 10716396 TI - Importance of US findings in access planning during jugular vein hemodialysis catheter placements. AB - PURPOSE: To evaluate the significance of internal jugular vein ultrasound (US) findings in long-term hemodialysis patients and to assess how frequently these findings lead to a change in access approach. MATERIALS AND METHODS: One hundred consecutive hemodialysis catheter placements in 79 patients were retrospectively analyzed. Prior to catheter insertion, each patient underwent an US examination of the proposed access site by an interventional radiologist or interventional radiology fellow. The examinations were recorded on VHS tapes. The procedure notes, dictated radiology reports, and VHS tapes were reviewed for evidence of total occlusion, non-occlusive thrombus, presence of venous collaterals, stenosis, or variation in normal anatomy. The number of months that the patient required hemodialysis prior to catheter placement was also noted. RESULTS: Significant US findings were present in 28 patients (35%). Findings included total occlusion (n = 18), non-occlusive thrombus (n = 11), stenosis (n = 5), and anatomic variation (n = 1). These required a change in access approach in 21 patients. Unexpectedly, 54% of the patients with US findings had been undergoing dialysis for 12 months or less. CONCLUSION: These results underscore the importance of sonography in planning and performing vascular access procedures. A thorough US examination of the internal jugular veins is warranted prior to hemodialysis catheter placement, especially in patients with previous temporary or tunneled catheters. Three-quarters of patients with sonographic abnormalities required a change in access approach. PMID- 10716397 TI - Fluoroscopically guided percutaneous placement of large-bore gastrostomy and gastrojejunostomy tubes: review of 109 cases. AB - PURPOSE: To evaluate our experience with percutaneous placement, management, and complications of large-bore (20-24 F) gastrostomy and gastrojejunostomy feeding tubes. MATERIALS AND METHODS: A retrospective review was performed on 109 consecutive patients who underwent placement of percutaneous large-bore feeding tubes between January 1994 and May 1998. Data were collected with respect to underlying illness, technical success, number of replaced tubes, and immediate and late complications. No patient had a small-bore tube placed during this series. RESULTS: A total of 109 cases were reviewed. Immediate follow-up within the first 2 weeks was available for all 109. Follow-up after 2 weeks was available for 61 (56%) patients. Tubes were placed in patients aged 15 to 94 years. Neurologic dysfunction from a variety of causes was the most common underlying illness and occurred in 52% of patients. There were nine (8.3%) immediate, treatable complications: three major and six minor. There was one procedure-related death (0.9%). Persistent fistula tracts following tube removal occurred in three patients (4.9%). Balloon rupture was the most common reason for tube exchange (40.7%). CONCLUSION: Percutaneous large-bore gastrostomy and gastrojejunostomy tubes are safe to place and have technical success, morbidity, and mortality rates comparable to those of tubes placed surgically or endoscopically as well as small-bore tubes placed with fluoroscopic guidance. PMID- 10716399 TI - SCVIR Annual Meeting Film Panel Session PMID- 10716398 TI - Alcohol ablation of a mesenteric lymphangioma. PMID- 10716400 TI - Nutritive characterization of purslane accessions as influenced by planting date. AB - The uniqueness of purslane (Portulaca spp.) as the richest vegetable source of omega-3 (omega-3) fatty acids is well documented. However, purslane has not been domesticated or fully evaluated for its nutritive value. The objective of this study was to determine the influence of planting date on chemical composition of purslane accessions. Eight accessions from different geographical locations were planted 12 days apart, and whole plants harvested at full bloom. Chemical analysis (DM basis) of leaves showed significant differences among varieties for all the characteristics measured. Accession by planting date interaction influenced (p <0.05) levels of crude protein, total lipids, and carbohydrate contents. Wild Greek accession had the highest, while a Beltsville (Maryland) wild type had the lowest crude protein content (27.1 vs 20.5%) at the second planting date. Crude protein, lipid and ash levels were most influenced (p <0.05) by planting date. Total lipids varied from 4.0-5.8% and 3.7-5.1% for the first and second planting dates, respectively. Selected fatty acid content indicated significantly (p <0.05) higher levels of 18: 2omega6, and 18: 3omega3 in the Dutch Garden accession compared with other varieties. The Egyptian wild accession had the lowest level of 18: 3omega3. The ratio of omega3 to omega6 acids, which ranged from 5.5 to 22.3 indicated a high nutritive value of purslane compared to other oil crops such as soybeans and perrilla. The high levels of protein in purslane compete with those of other commercially important vegetable crops. The study shows that, in spite of its genetic diversity, purslane remains one of the most abundant terrestrial vegetable sources of Omega-3 fatty acids and other essential nutrients potentially beneficial for humans as well as animals. PMID- 10716401 TI - Production of African breadfruit (Treculia africana) and soybean (Glycine max) seed based food formulations, 1: Effects of germination and fermentation on nutritional and organoleptic quality. AB - Germination and fermentation were investigated as methods of improving the nutritional and organoleptic properties of soybean and African breadfruit seed based food formulations. Four products consisting of germinated-fermented soy breadfruit seeds (GFSB), nongerminated-fermented soy-breadfruit seeds (NGFSB), germinated-nonfermented soy-breadfruit seeds (GNFSB) and nongerminated nonfermented soy-breadfruit seeds (NGNFSB) were prepared. Phytic acid contents, in vitro protein digestibilities, protein efficiency ratios (PER), net protein ratios (NPR), flavor, appearance and overall acceptability were evaluated. Germination followed by natural lactic fermentation significantly (p < 0.05) reduced the phytic acid by a factor of 11.6 in NGNFSB compared to reduction factors of 2.1 and 1.5 in GNFSB and NGFSB, respectively. The in vitro protein digestibility (%), PER and NPR values of 73.4, 2.46 and 3.62 for GFSB; 71.1, 2.35 and 3.46 for NGFSB; 68.7, 2.16 and 3.41 for GNFSB were significantly (p < 0.05) higher than the 64.7, 1.82 and 2.11 for NGNFSB. The mean sensory scores were 5.26 5.67 for GNFSB, 4.66-4.94 for NGNFSB, 4.33-4.80 for GFSB and 4.27-4.34 for NGFSB on a 7-point rating scale. PMID- 10716402 TI - Production of African breadfruit (Treculia africana) and soybean (Glycine max) seed based food formulations, 2: Effects of germination and fermentation on microbiological and physical properties. AB - The effects of germination (G) and naturally fermented (F) on the bacterial flora, viscosities and moisture sorption isotherms of soybean (S) and African breadfruit (B) seed based food products were investigated. Bacillus, Enterobacter, Enterobacteriaceae, Proteus, Serratia and Staphylococcus species dominated in the nonfermented products. Lactobacillus, Leuconostoc, Pediococcus and yeast species dominated in the fermented products whose gruels also inhibited growth of coagulase positive Staphylococcus aureus in challenge tests. Germination and fermentation resulted in significant (p < 0.05) decreases in cooked paste viscosities which is advantageous in increasing nutrient density. The monolayer moisture contents (g H2O/g solid) and surface areas for monolayer adsorption (m2/g solid) derived from BET model were 0.0422 and 148.1 (GFSB); 0.0428 and 150.4 (NGFSB); 0.0436 and 153.3 (NGNFSB); 0.0531 and 186.6 (GNFSB), respectively. PMID- 10716403 TI - Studies on the baking properties of wheat: pigeonpea flour blends. AB - Pigeonpea flour was substituted at levels of 0, 5, 10, 15, 20, 25% to wheat flour and whole wheat meal for bread and Chapatti making, respectively. Blends were prepared up to 50% for cookie making. Increasing levels of pigeonpeas in the blends significantly increased the protein and mineral content of the baked products. The bread from 10% pigeonpea flour blend with 2-3% vital gluten and 0.5% SSL had high loaf volume and loaf quality. Blends containing 15% pigeonpea flour were acceptable for Chapatti and 30% pigeonpea flour with 0.25% SSL were acceptable for cookie making. PMID- 10716404 TI - Proximate composition and selected functional properties of fermented and unfermented African oil bean (Pentaclethra macrophylla) seed flour. AB - Flour samples were prepared from fermented and unfermented African oil bean (Pentaclethra macrophylla) seeds (AOBS). The flour samples were evaluated for proximate composition and certain functional properties. The influence of defatting on these properties was also determined. Fermentation significantly increased (p < 0.05) the protein and decreased the crude fiber, ash, fat and carbohydrate contents of the AOBS flours. The nitrogen solubility of both fermented and unfermented flours was pH dependent with minimum and maximum solubility at pH 4.0 and pH 8.0, respectively, and with increased nitrogen solubility in the fermented sample. The fermented and unfermented flour samples had least gelation concentrations of 14 and 16% (w/v), respectively. The water absorption capacity and foam capacities of the fermented flour were 36 and 34%, respectively, over the unfermented seed flour. On the other hand, fermentation decreased the fat absorption capacity, emulsion activity and emulsion and foam stabilities. Fermentation decreased (p < 0.05) the bulk density of AOBS flour by 15%. Defatting improved all the functional properties evaluated except emulsion activity. These results indicate potential food uses of fermented and unfermented AOBS flour samples as protein supplements in diets and as functional ingredients in formulated foods. PMID- 10716405 TI - Effect of process improvement on the physico-chemical properties of infant weaning food from fermented composite blends of cereal and soybeans. AB - Saccharomyces cerevisiae and Lactobacillus plantarum ATCC 10776 were used as starters to ferment various composite blends of cereals and legumes produced through malting and toasting of two varieties of maize--Zea mays (DMR-LSR white & DMR-ESR yellow), sorghum--Sorghum bicolor (Dawa white & Dawa red) and one variety of soybeans (Glycine ax). Compared to the untoasted and unmalted fermented blends, a relatively lower pH (3.6), highly sour product was obtained with 12 h of fermentation. Results also showed that cereal and soybean toasting brought about a better reconstitution indices (B25, 84 ml; B45, 87 ml), water holding capacities (B25, 0.68 ml/g; B45, 0.62 ml/g), bulk densities (C15, 11.6; C35, 10.8) and gross energy (B15, 501.5 k cal/100g; B45, 508.5 kcal/100g) at the end of fermentation. Furthermore, reductions in total polyphenol and tannin contents were observed with fermentation of toasted and malted cereal blends supplemented with toasted and malted soybeans while porridges from the same blends displayed desirable starch stability and consistent gelling tendency, although B15 (a ferment of malted, toasted white maize supplemented with toasted and malted soybean) fell within acceptable limits. In all, the physical characteristics were affected by varieties of cereal and soybeans. PMID- 10716406 TI - Protein extraction from Atriplex lampa leaves: potential use as forage for animals used for human diets. AB - The purpose of this study was to evaluate the effect of pH on the extraction of protein nitrogen from Atriplex lampa leaves (Moquin) Dietrich. The chemical characterization of the dry matter indicated the following (g/100 g): protein, 26.93; ash, 21.80; ether extract, 4.65; dry matter, 37.30; sodium, 6.05; and calcium, 0.41. Non-critical values were obtained for saponins and nitrates. The high concentration of oxalic acid (8.52 g/100 g), together with elevated salt content account for the low palatability of the studied species. In order to determine the parameters needed to improve the extraction in protein nitrogen from leaves, fresh material was macerated with 2% sodium metasulfite, followed by pulping with a hand-driven grinder. Extractions were performed at different pH values (2-12) adjusting the value with 5N HCL or NaOH, with agitation followed by centrifugation and pressing. Supernatants were collected and kept. The last extraction was performed with Tween 20 in order to obtain maximum nitrogen recovery from the residue cake. Highest extraction (41.23%) was obtained at pH 10 with a 1:5 ratio (leaf: deionized water, w/v). It is proposed that this regional natural resource may be used to elaborate a protein concentrate, which can be made more palatable by decreasing potassium and sodium salt content with the use of membrane technology. PMID- 10716407 TI - Effect of blanching and ripening on functional properties of plantain (Musa aab) flour. AB - Flours were prepared from raw and blanched samples of unripe and ripe mature plantain Musa aab and examined for their proximate composition, physical characteristics and functional properties. The plantain flours contained 3.5 g crude protein, 2.5-3.5 g crude fat, 5.7-7.1 g moisture, 1.33-2.0 g crude fiber, 1.66-2.0 g ash, and 82.25-86.07 g carbohydrate per 100 g sample. The flours had bulk densities between 0.42-0.72 g/ml, emulsion capacities of 4.7-14.7%, water absorption capacities of 250-338%, oil absorption capacities of 214-371%, foaming capacities of 1.90-5.79%, least gelation concentrations of 6-8%, and viscosities of 23.7-46.7 CP at 2% slurry concentration. Foaming capacity increased with increasing flour concentration. Blanching considerably reduced the emulsion capacity and viscosity, while bulk density, water and oil absorption capacities were increased by blanching. Ripening was found to have a negative effect on all the functional properties examined except the bulk density, and gelation property. Unripe plantain could be used as an emulsifier and thickener in a food system. PMID- 10716409 TI - Albright's last scientific stand PMID- 10716410 TI - A critical nuclear appointment PMID- 10716408 TI - Nutritive value and effect of blanching on the trypsin and chymotrypsin inhibitor activities of selected leafy vegetables. AB - Proximate composition, energy, mineral and vitamin contents and the effect of blanching methods and times on the trypsin and chymotrypsin inhibitor activities were studied using cabbage, collard, turnip, peanut, and sweet potato leaves. Results of this study indicated that, crude protein, crude fat, carbohydrate and ash contents were in the range of 15.5-25.6%, 1.4-6.5%, 60.4-73.1% and 6.8-7.5%, respectively. Total dietary fiber was lowest in cabbage (28.2 g/100 g) and highest in the collard leaves (43.1%) while energy content per 100 g of vegetables was highest in sweet potato leaves (402 kcal) and lowest in cabbage (379 kcal). The mineral content per 100 g of vegetables were in the range of 33.4 249.8 mg, 241.2-471.2 mg, 12.1-75.1 mg, 14.9-98.9 mg, 0.5-3.5 mg and 0.9-3.1 mg for Ca, K, Na, Mg, Fe and Zn, respectively. For ascorbic acid, riboflavin, thiamin and total carotenoids, concentrations in 100 g of vegetables were in the range of 45.1-112.7 mg, 0.2-0.3 mg, 0.3-0.8 mg and 2.0-7.3 mg, respectively. The trypsin inhibitory activity per gram of the vegetables was highest in collard (60.1 TIU/g) and lowest in peanut leaves (41.0 TIU/g). Chymotrypsin inhibitor activity was highest in the peanut (69.6 CIU/g) but lowest in the collard leaves (48.0 CIU/g). Both trypsin and chymotrypsin inhibitor activities were significantly (p < 0.05) reduced by most of the treatments in either the conventional or microwave blanching methods. In the conventional blanching method, trypsin inhibitor activity was reduced by 0.5, 6.8, 11.9, 9.0 and 19.3 percent in cabbage, collard, turnip, sweet potato and peanut leaves, respectively, when the vegetables were blanched for 2.5 minutes but after blanching for 10 minutes, the trypsin inhibitor activity was reduced by 29.7, 34.9, 54.3, 52.3 and 65.6 percent in cabbage, collard, turnip, sweet potato and peanut greens, respectively. For the microwave oven blanching, trypsin inhibitor activity was reduced by 3.8, 3.3, 32.7, 5.0 and 9.5 percent in cabbage, collard, turnip, sweet potato and peanut leaves, respectively when the vegetables were blanched for 30 seconds. When blanched for 60 seconds, trypsin inhibitor activity was reduced by 16.2, 45.8, 46.2, 51.0 and 42.4 percent in cabbage, collard, turnip, sweet potato and peanut greens, respectively. Similar trends in the reduction of chymotrypsin inhibitor activity were observed when the vegetables were conventionally blanched for 2.5, 5 and 10 minutes and when blanched by microwave oven for 30, 45 and 60 seconds. Based on the results of this study, the vegetables were good dietary sources of minerals, vitamins, carbohydrate and proteins. Also, blanching was an effective method for reducing the trypsin and chymotripsin inhibitor activities in the leafy vegetables, however, further investigation on the heating times for both conventional and microwave blanching methods is suggested. PMID- 10716411 TI - Germany challenges human stem cell patent awarded 'by mistake'. PMID- 10716412 TI - NIH tightens up monitoring of gene-therapy mishaps. PMID- 10716413 TI - Culling plans put future of red deer study in jeopardy. PMID- 10716414 TI - Young, worldly and unhelpful all miss out on data sharing. PMID- 10716415 TI - Promega and Roche take up battle over PCR patents. PMID- 10716416 TI - Funding woes spell doom for US radio dish PMID- 10716417 TI - Post-cold war needs 'new forms of scientific linkage' PMID- 10716418 TI - Analysis of polio vaccine could end dispute over how AIDS originated. PMID- 10716419 TI - Japanese officials found guilty in HIV blood case. PMID- 10716420 TI - Crowded universities cramp more than just students' style PMID- 10716421 TI - Patent confusion in law on new plant varieties. PMID- 10716422 TI - Reversing Rorschach. PMID- 10716423 TI - Danger--hard hack area PMID- 10716424 TI - Of ice and elephants PMID- 10716425 TI - p53 sends nucleotides to repair DNA. PMID- 10716426 TI - Circadian clocks. Heartfelt enlightenment. PMID- 10716427 TI - Physics. Brane new worlds PMID- 10716428 TI - Imaging in the fourth dimension. PMID- 10716429 TI - Brimstone on mars. PMID- 10716430 TI - Errors in judging 'offside' in football. PMID- 10716431 TI - Finding genes in Plasmodium falciparum. PMID- 10716432 TI - Fish do not avoid survey vessels. PMID- 10716433 TI - Migration and speciation. PMID- 10716434 TI - The importance of repairing stalled replication forks. AB - The bacterial SOS response to unusual levels of DNA damage has been recognized and studied for several decades. Pathways for re-establishing inactivated replication forks under normal growth conditions have received far less attention. In bacteria growing aerobically in the absence of SOS-inducing conditions, many replication forks encounter DNA damage, leading to inactivation. The pathways for fork reactivation involve the homologous recombination systems, are nonmutagenic, and integrate almost every aspect of DNA metabolism. On a frequency-of-use basis, these pathways represent the main function of bacterial DNA recombination systems, as well as the main function of a number of other enzymatic systems that are associated with replication and site-specific recombination. PMID- 10716435 TI - A ribonucleotide reductase gene involved in a p53-dependent cell-cycle checkpoint for DNA damage. AB - The p53 gene is frequently inactivated in human cancers. Here we have isolated a p53-inducible gene, p53R2, by using differential display to examine messenger RNAs in a cancer-derived human cell line carrying a highly regulated wild-type p53 expression system. p53R2 contains a p53-binding sequence in intron 1 and encodes a 351-amino-acid peptide with striking similarity to the ribonucleotide reductase small subunit (R2), which is important in DNA synthesis during cell division. Expression of p53R2, but not R2, was induced by ultraviolet and gamma irradiation and adriamycin treatment in a wild-type p53-dependent manner. Induction of p53R2 in p53-deficient cells caused G2/M arrest and prevented cells from death in response to adriamycin. Inhibition of endogenous p53R2 expression in cells that have an intact p53-dependent DNA damage checkpoint reduced ribonucleotide reductase activity, DNA repair and cell survival after exposure to various genotoxins. Our results indicate that p53R2 encodes a ribonucleotide reductase that is directly involved in the p53 checkpoint for repair of damaged DNA. The discovery of p53R2 clarifies a relationship between a ribonucleotide reductase activity involved in repair of damaged DNA and tumour suppression by p53. PMID- 10716437 TI - Fabrication of photonic crystals for the visible spectrum by holographic lithography AB - The term 'photonics' describes a technology whereby data transmission and processing occurs largely or entirely by means of photons. Photonic crystals are microstructured materials in which the dielectric constant is periodically modulated on a length scale comparable to the desired wavelength of operation. Multiple interference between waves scattered from each unit cell of the structure may open a 'photonic bandgap'--a range of frequencies, analogous to the electronic bandgap of a semiconductor, within which no propagating electromagnetic modes exist. Numerous device principles that exploit this property have been identified. Considerable progress has now been made in constructing two-dimensional structures using conventional lithography, but the fabrication of three-dimensional photonic crystal structures for the visible spectrum remains a considerable challenge. Here we describe a technique--three dimensional holographic lithography--that is well suited to the production of three-dimensional structures with sub-micrometre periodicity. With this technique we have made microperiodic polymeric structures, and we have used these as templates to create complementary structures with higher refractive-index contrast. PMID- 10716436 TI - Evidence of atmospheric sulphur in the martian regolith from sulphur isotopes in meteorites. AB - Sulphur is abundant at the martian surface, yet its origin and evolution over time remain poorly constrained. This sulphur is likely to have originated in atmospheric chemical reactions, and so should provide records of the evolution of the martian atmosphere, the cycling of sulphur between the atmosphere and crust, and the mobility of sulphur in the martian regolith. Moreover, the atmospheric deposition of oxidized sulphur species could establish chemical potential gradients in the martian near-surface environment, and so provide a potential energy source for chemolithoautotrophic organisms. Here we present measurements of sulphur isotopes in oxidized and reduced phases from the SNC meteorites--the group of related achondrite meteorites believed to have originated on Mars- together with the results of laboratory photolysis studies of two important martian atmospheric sulphur species (SO2 and H2S). The photolysis experiments can account for the observed sulphur-isotope compositions in the SNC meteorites, and so identify a mechanism for producing large abiogenic 34S fractionations in the surface sulphur reservoirs. We conclude that the sulphur data from the SNC meteorites reflects deposition of oxidized sulphur species produced by atmospheric chemical reactions, followed by incorporation, reaction and mobilization of the sulphur within the regolith. PMID- 10716438 TI - Electrophoretic assembly of colloidal crystals with optically tunable micropatterns AB - The production of materials with micrometre- and submicrometre-scale patterns is of importance in a range of applications, such as photonic materials, high density magnetic data storage devices, microchip reactors and biosensors. One method of preparing such structures is through the assembly of colloidal particles. Micropatterned colloidal assemblies have been produced with lithographically patterned electrodes or micromoulds. Here we describe a different method that combines the well-known photochemical sensitivity of semiconductors with electric-field-induced assembly to create ordered arrays of micrometre-sized colloidal particles with tunable patterns. We show that light affects the assembly processes, and demonstrate how to produce patterns using electrophoretic deposition in the presence of an ultraviolet (UV) illumination motif. The distribution of current across an indium tin oxide (ITO) electrode can be altered by varying the illumination intensity: during the deposition process, this causes colloidal particles to be swept from darkened areas into lighted regions. Illumination also assists in immobilizing the particles on the electrode surface. Although the details of these processes are not well understood, the patterning effects of the UV light are discussed in terms of alterations in the current density that affects particle assembly on an ITO electrode. PMID- 10716439 TI - Shape control of CdSe nanocrystals AB - Nanometre-size inorganic dots, tubes and wires exhibit a wide range of electrical and optical properties that depend sensitively on both size and shape, and are of both fundamental and technological interest. In contrast to the syntheses of zero dimensional systems, existing preparations of one-dimensional systems often yield networks of tubes or rods which are difficult to separate. And, in the case of optically active II-VI and III-V semiconductors, the resulting rod diameters are too large to exhibit quantum confinement effects. Thus, except for some metal nanocrystals, there are no methods of preparation that yield soluble and monodisperse particles that are quantum-confined in two of their dimensions. For semiconductors, a benchmark preparation is the growth of nearly spherical II-VI and III-V nanocrystals by injection of precursor molecules into a hot surfactant. Here we demonstrate that control of the growth kinetics of the II-VI semiconductor cadmium selenide can be used to vary the shapes of the resulting particles from a nearly spherical morphology to a rod-like one, with aspect ratios as large as ten to one. This method should be useful, not only for testing theories of quantum confinement, but also for obtaining particles with spectroscopic properties that could prove advantageous in biological labelling experiments and as chromophores in light-emitting diodes. PMID- 10716440 TI - Evidence from U-Th dating against Northern Hemisphere forcing of the penultimate deglaciation AB - Milankovitch proposed that summer insolation at mid-latitudes in the Northern Hemisphere directly causes the ice-age climate cycles. This would imply that times of ice-sheet collapse should correspond to peaks in Northern Hemisphere June insolation. But the penultimate deglaciation has proved controversial because June insolation peaks 127 kyr ago whereas several records of past climate suggest that change may have occurred up to 15 kyr earlier. There is a clear signature of the penultimate deglaciation in marine oxygen-isotope records. But dating this event, which is significantly before the 14C age range, has not been possible. Here we date the penultimate deglaciation in a record from the Bahamas using a new U-Th isochron technique. After the necessary corrections for alpha recoil mobility of 234U and 230Th and a small age correction for sediment mixing, the midpoint age for the penultimate deglaciation is determined to be 135 +/- 2.5 kyr ago. This age is consistent with some coral-based sea-level estimates, but it is difficult to reconcile with June Northern Hemisphere insolation as the trigger for the ice-age cycles. Potential alternative driving mechanisms for the ice-age cycles that are consistent with such an early date for the penultimate deglaciation are either the variability of the tropical ocean-atmosphere system or changes in atmospheric CO2 concentration controlled by a process in the Southern Hemisphere. PMID- 10716441 TI - Stable sulphate clusters as a source of new atmospheric particles AB - The formation of new atmospheric particles with diameters of 3-10 nm has been observed at a variety of altitudes and locations. Such aerosol particles have the potential to grow into cloud condensation nuclei, thus affecting cloud formation as well as the global radiation budget. In some cases, the observed formation rates of new particles have been adequately explained by binary nucleation, involving water and sulphuric acid, but in certain locations--particularly those within the marine boundary layer and at continental sites--observed ambient nucleation rates exceed those predicted by the binary scheme. In these locations, ambient sulphuric acid (H2SO4) levels are typically lower than required for binary nucleation, but are sufficient for ternary nucleation (sulphuric acid ammonia-water). Here we present results from an aerosol dynamics model with a ternary nucleation scheme which indicate that nucleation in the troposphere should be ubiquitous, and yield a reservoir of thermodynamically stable clusters 1-3 nm in size. We suggest that the growth of these clusters to a detectable size (> 3 nm particle diameter) is restricted by the availability of condensable vapour. Observations of atmospheric particle formation and growth from a continental and a coastal site support this hypothesis, indicating that a growth process including ternary nucleation is likely to be responsible for the formation of cloud condensation nuclei. PMID- 10716442 TI - Geodetic evidence for a low slip rate in the Altyn Tagh fault system AB - The collision between India and Asia has been simulated with a variety of computational models that describe or predict the motions of the main faults of east Asia. Geological slip-rate estimates of 20-30 mm yr(-1) suggest that the largest of these faults, the 2,000-km-long Altyn Tagh fault system on the northern edge of the Tibetan plateau, absorbs as much of the Indo-Asian convergence signal as do the Himalayas--partly by oblique slip and partly by contraction and mountain growth. However, the predictions of dynamic models for Asian deformation and the lower bounds of some geological slip-rates estimates (3 9 mm yr(-1); refs 7, 8) suggest that the Altyn Tagh system is less active. Here, we report geodetic data from 89-91 degrees E that indicate left-lateral shear of 9 +/- 5 mm yr(-1) and contraction of 3 +/- 1 mm yr(-1) across the Altyn Tagh system. This result--combined with our finding that, at 90 degrees E, Tibet contracts north-south at 9 +/- 1 mm yr(-1)--supports the predictions of dynamic models of Asian deformation. PMID- 10716443 TI - Tree species impoverishment and the future flora of the Atlantic forest of northeast Brazil. AB - Estimates of species extinction due to human impact on tropical forests have previously been based on the relationship between species number and area. Here we use a different approach to estimate loss of tree species in the Atlantic forest of northeast Brazil. We evaluate the characteristics of plant species, their avian dispersers and the distribution of the forest remnants on the landscape to estimate that about 33.9% of tree species in this region will become extinct on a regional scale. Because northeast Brazil is the most threatened sector of South American Atlantic forest, our results highlight the need to change the current conservation paradigm for this region. Rather than focus on the creation of isolated reserves in any medium-to-large forest remnant, a bioregional planning approach is urgently required to rescue this unique biota from extinction. PMID- 10716444 TI - Egg investment is influenced by male attractiveness in the mallard. AB - Why females prefer to copulate with particular males is a contentious issue. Attention is currently focused on whether females choose males on the basis of their genetic quality, in order to produce more viable offspring. Support for this hypothesis in birds has come from studies showing that preferred males tend to father offspring of better condition or with increased survivorship. Before attributing greater offspring viability to a male's heritable genetic quality, however, it is important to discount effects arising from confounding sources, including maternal effects. This has generally been addressed by comparing offspring viability from two different breeding attempts by the same female: one when offspring are sired by a preferred male, and one when offspring are sired by a less preferred male. However, here we show that individual female mallard (Anas platyrhynchos) lay larger eggs after copulating with preferred males and smaller eggs after copulating with less preferred males. As a result, females produced offspring of better body condition when paired with preferred males. After controlling for these differences in maternal investment, we found no effect of paternity on offspring condition. This shows that differences between half-sibs cannot always be attributed to paternal or maternal genetic effects. PMID- 10716445 TI - Macaque monkeys categorize images by their ordinal number. AB - The recall of a list of items in a serial order is a basic cognitive skill. However, it is unknown whether a list of arbitrary items is remembered by associations between sequential items or by associations between each item and its ordinal position. Here, to study the nonverbal strategies used for such memory tasks, we trained three macaque monkeys on a delayed sequence recall task. Thirty abstract images, divided into ten triplets, were presented repeatedly in fixed temporal order. On each trial the monkeys viewed three sequentially presented sample stimuli, followed by a test stimulus consisting of the same three images and a distractor image (chosen randomly from the remaining 27). The task was to touch the three images in their original order without touching the distractor. The most common error was touching the distractor when it had the same ordinal number (in its own triplet) as the correct image. Thus, the monkeys' natural tendency was to categorize images by their ordinal number. Additional, secondary strategies were used eventually to avoid the distractor images. These included memory of the sample images (working memory) and associations between sequence triplet members. Thus, monkeys use multiple mnemonic strategies according to their innate tendencies and the requirements of the task. PMID- 10716446 TI - Temporal patterns of human cortical activity reflect tone sequence structure. AB - Despite growing interest in temporal aspects of auditory neural processing, little is known about large-scale timing patterns of brain activity during the perception of auditory sequences. This is partly because it has not been possible to distinguish stimulus-related activity from other, endogenous brain signals recorded by electrical or magnetic sensors. Here we use amplitude modulation of unfamiliar, approximately 1-minute-long tone sequences to label stimulus-related magnetoencephalographic neural activity in human subjects. We show that temporal patterns of activity recorded over particular brain regions track the pitch contour of tone sequences, with the accuracy of tracking increasing as tone sequences become more predictable in structure. In contrast, temporal synchronization between recording locations, particularly between sites over the left posterior hemisphere and the rest of the brain, is greatest when sequences have melody-like statistical properties, which may reflect the perceptual integration of local and global pitch patterns in melody-like sequences. This method is particularly well suited to studying temporal neural correlates of complex auditory sequences (such as speech or music) which engage multiple brain areas as perception unfolds in time. PMID- 10716447 TI - Cannabinoids control spasticity and tremor in a multiple sclerosis model. AB - Chronic relapsing experimental allergic encephalomyelitis (CREAE) is an autoimmune model of multiple sclerosis. Although both these diseases are typified by relapsing-remitting paralytic episodes, after CREAE induction by sensitization to myelin antigens Biozzi ABH mice also develop spasticity and tremor. These symptoms also occur during multiple sclerosis and are difficult to control. This has prompted some patients to find alternative medicines, and to perceive benefit from cannabis use. Although this benefit has been backed up by small clinical studies, mainly with non-quantifiable outcomes, the value of cannabis use in multiple sclerosis remains anecdotal. Here we show that cannabinoid (CB) receptor agonism using R(+)-WIN 55,212, delta9-tetrahydrocannabinol, methanandamide and JWH-133 (ref. 8) quantitatively ameliorated both tremor and spasticity in diseased mice. The exacerbation of these signs after antagonism of the CB1 and CB2 receptors, notably the CB1 receptor, using SR141716A and SR144528 (ref. 8) indicate that the endogenous cannabinoid system may be tonically active in the control of tremor and spasticity. This provides a rationale for patients' indications of the therapeutic potential of cannabis in the control of the symptoms of multiple sclerosis, and provides a means of evaluating more selective cannabinoids in the future. PMID- 10716448 TI - Light acts directly on organs and cells in culture to set the vertebrate circadian clock. AB - The expression of clock genes in vertebrates is widespread and not restricted to classical clock structures. The expression of the Clock gene in zebrafish shows a strong circadian oscillation in many tissues in vivo and in culture, showing that endogenous oscillators exist in peripheral organs. A defining feature of circadian clocks is that they can be set or entrained to local time, usually by the environmental light-dark cycle. An important question is whether peripheral oscillators are entrained to local time by signals from central pacemakers such as the eyes or are themselves directly light-responsive. Here we show that the peripheral organ clocks of zebrafish are set by light-dark cycles in culture. We also show that a zebrafish-derived cell line contains a circadian oscillator, which is also directly light entrained. PMID- 10716449 TI - Delayed activation of the paternal genome during seed development. AB - Little is known about the timing of the maternal-to-zygotic transition during seed development in flowering plants. Because plant embryos can develop from somatic cells or microspores, maternal contributions are not considered to be crucial in early embryogensis. Early-acting embryo-lethal mutants in Arabidopsis, including emb30/gnom which affects the first zygotic division, have fuelled the perception that both maternal and paternal genomes are active immediately after fertilization. Here we show that none of the paternally inherited alleles of 20 loci that we tested is expressed during early seed development in Arabidopsis. For genes that are expressed at later stages, the paternally inherited allele becomes active three to four days after fertilization. The genes that we tested are involved in various processes and distributed throughout the genome, indicating that most, if not all, of the paternal genome may be initially silenced. Our findings are corroborated by genetic studies showing that emb30/gnom has a maternal-effect phenotype that is paternally rescuable in addition to its zygotic lethality. Thus, contrary to previous interpretations, early embryo and endosperm development are mainly under maternal control. PMID- 10716450 TI - Eomesodermin is required for mouse trophoblast development and mesoderm formation. AB - The earliest cell fate decision in the mammalian embryo separates the extra embryonic trophoblast lineage, which forms the fetal portion of the placenta, from the embryonic cell lineages. The body plan of the embryo proper is established only later at gastrulation, when the pluripotent epiblast gives rise to the germ layers ectoderm, mesoderm and endoderm. Here we show that the T-box gene Eomesodermin performs essential functions in both trophoblast development and gastrulation. Mouse embryos lacking Eomesodermin arrest at the blastocyst stage. Mutant trophoectoderm does not differentiate into trophoblast, indicating that Eomesodermin may be required for the development of trophoblast stem cells. In the embryo proper, Eomesodermin is essential for mesoderm formation. Although the specification of the anterior-posterior axis and the initial response to mesoderm-inducing signals is intact in mutant epiblasts, the prospective mesodermal cells are not recruited into the primitive streak. Our results indicate that Eomesodermin defines a conserved molecular pathway controlling the morphogenetic movements of germ layer formation and has acquired a new function in mammals in the differentiation of trophoblast. PMID- 10716451 TI - p73-deficient mice have neurological, pheromonal and inflammatory defects but lack spontaneous tumours. AB - p73 (ref. 1) has high homology with the tumour suppressor p53 (refs 2-4), as well as with p63, a gene implicated in the maintenance of epithelial stem cells. Despite the localization of the p73 gene to chromosome 1p36.3, a region of frequent aberration in a wide range of human cancers, and the ability of p73 to transactivate p53 target genes, it is unclear whether p73 functions as a tumour suppressor. Here we show that mice functionally deficient for all p73 isoforms exhibit profound defects, including hippocampal dysgenesis, hydrocephalus, chronic infections and inflammation, as well as abnormalities in pheromone sensory pathways. In contrast to p53-deficient mice, however, those lacking p73 show no increased susceptibility to spontaneous tumorigenesis. We report the mechanistic basis of the hippocampal dysgenesis and the loss of pheromone responses, and show that new, potentially dominant-negative, p73 variants are the predominant expression products of this gene in developing and adult tissues. Our data suggest that there is a marked divergence in the physiological functions of the p53 family members, and reveal unique roles for p73 in neurogenesis, sensory pathways and homeostatic control. PMID- 10716452 TI - Single-molecule studies of the effect of template tension on T7 DNA polymerase activity. AB - T7 DNA polymerase catalyses DNA replication in vitro at rates of more than 100 bases per second and has a 3'-->5' exonuclease (nucleotide removing) activity at a separate active site. This enzyme possesses a 'right hand' shape which is common to most polymerases with fingers, palm and thumb domains. The rate limiting step for replication is thought to involve a conformational change between an 'open fingers' state in which the active site samples nucleotides, and a 'closed' state in which nucleotide incorporation occurs. DNA polymerase must function as a molecular motor converting chemical energy into mechanical force as it moves over the template. Here we show, using a single-molecule assay based on the differential elasticity of single-stranded and double-stranded DNA, that mechanical force is generated during the rate-limiting step and that the motor can work against a maximum template tension of approximately 34 pN. Estimates of the mechanical and entropic work done by the enzyme show that T7 DNA polymerase organizes two template bases in the polymerization site during each catalytic cycle. We also find a force-induced 100-fold increase in exonucleolysis above 40 pN. PMID- 10716453 TI - The role of tumor necrosis factor in the pathophysiology of heart failure. AB - Recent studies have focused their attention on the role of the proinflammatory cytokine tumor necrosis factor (TNF) in the development of heart failure. First recognized as an endotoxin-induced serum factor that caused necrosis of tumors and cachexia, it is now recognized that TNF participates in the pathophysiology of a group of inflammatory diseases including rheumatoid arthritis and Crohn's disease. The normal heart does not express TNF; however, the failing heart produces robust quantities. Furthermore, there is a direct relationship between the level of TNF expression and the severity of disease. In addition, both in vivo and in vitro studies demonstrate that TNF effects cellular and biochemical changes that mirror those seen in patients with congestive heart failure. Furthermore, in animal models, the development of the heart failure phenotype can be abrogated at least in part by anticytokine therapy. Based on information from experimental studies, investigators are now evaluating the clinical efficacy of novel anticytokine and anti-TNF strategies in patients with heart failure; one such strategy is the use of a recombinantly produced chimeric TNF alpha soluble receptor. Thus, in view of the emerging importance of proinflammatory cytokines in the pathogenesis of heart disease, we review the biology of TNF, its role in inflammatory diseases, the effects of TNF on the physiology of the heart and the development of clinical strategies that target the cytokine pathways. PMID- 10716454 TI - Atheromas of the thoracic aorta: clinical and therapeutic update. AB - Atherosclerotic lesions of the thoracic aorta have recently been recognized as an important cause of stroke and peripheral embolization, which may result in severe neurologic damage as well as multiorgan failure and death. Their prevalence is approximately 27% in patients with previous embolic events. Transesophageal echocardiography is the modality of choice for the diagnosis of these atheromas, although computed tomography, magnetic resonance imaging and intraoperative epiaortic ultrasound are complementary. Two clinical syndromes account for the embolic phenomena, atheroemboli and, more commonly, thromboemboli. In addition to such superimposed thrombi, plaque thickness (especially > or =4 mm) also correlates with embolic risk. This risk is high, with 12% of patients having a recurrent stroke within approximately one year, and up to 33% of patients having a stroke or peripheral embolus. In addition, aortic atheromas (as seen with intraoperative transesophageal echocardiography and intraoperative epiaortic ultrasound) are an important cause of stroke during heart surgery requiring cardiopulmonary bypass. Such strokes occur during approximately 12% of cardiac operations employing cardiopulmonary bypass when aortic arch atheromas are seen with transesophageal echocardiography (six times the general intraoperative stroke rate). Although anticoagulant strategies have been reported with encouraging results in nonrandomized studies, prospective, randomized data must be developed before an effective and safe treatment strategy can be determined. This review details the current state of knowledge in this area, including the clinical and pathologic evidence that thoracic aortic atherosclerosis is an important embolic source, data which guide current therapy and future directions for clinical investigation. PMID- 10716455 TI - Platelets and restenosis. AB - Restenosis is currently the major limitation of percutaneous transluminal coronary angioplasty (PTCA). Factors such as elastic recoil, migration of vascular smooth muscle cells from media to intima, neointimal proliferation and vascular remodeling underly the restenotic process. Presently there is no effective therapy available for restenosis. The role of platelets in the development of thrombosis and abrupt closure after PTCA is well recognized. However, the effects of platelets in PTCA extend well beyond the early phase. Although antiplatelet agents such as glycoprotein IIb/IIIa antagonists have been reported to reduce target vessel revascularization, major unresolved controversies still exist. This report reviews the potential role of platelets in restenosis. Various drugs, successfully tested in experimental studies and in a small number of human studies, that inhibit the effect of platelets on the restenotic process are also reviewed. PMID- 10716456 TI - Emerging concepts in the management of acute myocardial infarction in patients with diabetes mellitus. AB - Although fibrinolysis has improved survival of patients after myocardial infarction (MI), such therapy is less likely to be administered to patients with diabetes. Furthermore, these patients present later (15 min) than nondiabetics. Moreover, even with the use of early potent fibrinolytic agents, patients with diabetes continued to suffer excessive morbidity and mortality. This finding is not related to the ability of fibrinolytic agents to restore complete reperfusion or increased risk of reocclusion of the infarct-related artery. Instead, the impaired ventricular performance at the noninfarct areas and metabolic derangements during the acute phase of MI may account for the adverse outcome. The efficacy of percutaneous coronary revascularization procedures for treatment of acute MI requires further evaluation. Therapeutic approaches should consider correcting these abnormalities to afford greater survival benefit for this subset of high-risk patients. PMID- 10716457 TI - Cardiac remodeling--concepts and clinical implications: a consensus paper from an international forum on cardiac remodeling. Behalf of an International Forum on Cardiac Remodeling. AB - Cardiac remodeling is generally accepted as a determinant of the clinical course of heart failure (HF). Defined as genome expression resulting in molecular, cellular and interstitial changes and manifested clinically as changes in size, shape and function of the heart resulting from cardiac load or injury, cardiac remodeling is influenced by hemodynamic load, neurohormonal activation and other factors still under investigation. Although patients with major remodeling demonstrate progressive worsening of cardiac function, slowing or reversing remodeling has only recently become a goal of HF therapy. Mechanisms other than remodeling can also influence the course of heart disease, and disease progression may occur in other ways in the absence of cardiac remodeling. Left ventricular end-diastolic and end-systolic volume and ejection fraction data provide support for the beneficial effects of therapeutic agents such as angiotensin-converting enzyme (ACE) inhibitors and beta-adrenergic blocking agents on the remodeling process. These agents also provide benefits in terms of morbidity and mortality. Although measurement of ejection fraction can reliably guide initiation of treatment in HF, opinions differ regarding the value of ejection fraction data in guiding ongoing therapy. The role of echocardiography or radionuclide imaging in the management and monitoring of HF is as yet unclear. To fully appreciate the potential benefits of HF therapies, clinicians should understand the relationship between remodeling and HF progression. Their patients may then, in turn, acquire an improved understanding of their disease and the treatments they are given. PMID- 10716458 TI - Vascular remodeling and the local delivery of cytochalasin B after coronary angioplasty in humans. AB - OBJECTIVES: This study sought to determine the safety, feasibility and outcome of local delivery of cytochalasin B at the site of coronary angioplasty. BACKGROUND: Previous failures in the pharmacologic prevention of restenosis may have been related to inadequate dosing at the angioplasty site as a result of systemic drug administration. Alternatively, although previous experimental protocols have typically targeted control of excess tissue growth (intimal hyperplasia), it now appears that overall arterial constriction (vascular remodeling) is the major contributor to late lumen loss. Cytochalasin B inhibits the polymerization of actin and has proved to be a potent inhibitor of vascular remodeling in animal models. METHODS: In this phase I, multicenter, randomized, controlled trial, cytochalasin B (or matching placebo) was administered to the site of a successful balloon angioplasty using a microporous local delivery infusion balloon. RESULTS: The rate of drug delivery at a constant infusion pressure varied significantly from patient to patient (range 1.7 to 20.2 ml/min), perhaps related to a variable constricting effect of the atherosclerotic plaque on the infusion balloon. The minimal stenosis diameter after the procedure was slightly better in the active drug group (1.86 +/- 0.44 vs. 1.49 +/- 0.63 mm, p < 0.03), but this difference was not seen at four to six weeks. Although the study was not powered for clinical outcomes (n = 43), the combined end point (death, nonfatal infarction or repeat revascularization) was encountered in 20% of the patients receiving cytochalasin B and in 38% of the patients receiving placebo. Clinical restenosis occurred in 18% of the treatment group and 22% of the placebo group. There were no significant differences between groups in biochemical or electrocardiographic variables. CONCLUSIONS: Cytochalasin B can be safely administered by local delivery after successful coronary angioplasty and warrants further study of its efficacy in reducing restenosis. PMID- 10716459 TI - Restenosis and clinical outcome in patients treated with amlodipine after angioplasty: results from the Coronary AngioPlasty Amlodipine REStenosis Study (CAPARES). AB - OBJECTIVES: Our intent was to investigate the effect of the dihydropyridine calcium channel blocker amlodipine on restenosis and clinical outcome in patients undergoing percutaneous transluminal coronary angioplasty (PTCA). BACKGROUND: Amlodipine has sustained vasodilatory effects and relieves coronary spasm, which may reduce luminal loss and clinical complications after PTCA. METHODS: In a prospective, double-blind design, 635 patients were randomized to 10 mg of amlodipine or placebo. Pretreatment with the study drug started two weeks before PTCA and continued until four months after PTCA. The primary angiographic end point was loss in minimal lumen diameter (MLD) from post-PTCA to follow-up, as assessed by quantitative coronary angiography (QCA). Clinical end points were death, myocardial infarction, coronary artery bypass graft surgery and repeat PTCA (major adverse clinical events). RESULTS: Angioplasty was performed in 585 patients (92.1%); 91 patients (15.6%) had coronary stents implanted. Follow-up angiography suitable for QCA analysis was done in 236 patients in the amlodipine group and 215 patients in the placebo group (per-protocol group). The mean loss in MLD was 0.30 +/- 0.45 mm in the amlodipine group versus 0.29 +/- 0.49 mm in the placebo group (p = 0.84). The need for repeat PTCA was significantly lower in the amlodipine versus the placebo group (10 [3.1%] vs. 23 patients [7.3%], p = 0.02, relative risk ratio [RR]: 0.45, 95% confidence interval [CI]: 0.22 to 0.91), and the composite incidence of clinical events (30 [9.4%] vs. 46 patients (14.5%), p = 0.049, RR: 0.65, CI: 0.43 to 0.99) within the four months follow-up period (intention-to-treat analysis). CONCLUSIONS: Amlodipine therapy starting two weeks before PTCA did not reduce luminal loss, but the incidence of repeat PTCA and the composite major adverse clinical events were significantly reduced during the four-month follow-up period after PTCA with amlodipine as compared with placebo. PMID- 10716460 TI - High dose heparin as pretreatment for primary angioplasty in acute myocardial infarction: the Heparin in Early Patency (HEAP) randomized trial. AB - OBJECTIVES: In the Heparin in Early Patency (HEAP) pilot study a beneficial effect of high-dose heparin on early patency in acute myocardial infarction (MI) was observed in a matched-control study. BACKGROUND: High dose bolus intravenous injection of heparin may achieve lysis of coronary thrombi and could enhance early patency of the infarct related vessel in patients with MI scheduled for primary angioplasty. METHODS: Before primary angioplasty, 584 patients with MI entered an open randomized trial of high dose (300 IU/kg) or low dose (0 or 5,000 IU) heparin. Of the 584 patients, 299 were randomized to high dose and 285 patients to low dose heparin. RESULTS: Thrombolysis In Myocardial Infarction (TIMI) flow grade 2 or 3 was observed before primary angioplasty in 65 patients (22%) in the high dose group and 60 patients (21%) in the low dose heparin group (p > 0.1), whereas TIMI flow grade 3 was observed in 38 (13%) and 24 patients (9%), respectively (p = 0.11). There were no differences in the clinical end points between the two groups. There were no hemorrhagic strokes, while 10% of the patients in the high dose group required blood transfusion versus 6% in the low dose/no heparin group (p = 0.07). No subsets of patients showed beneficial effects of high dose heparin, such as patients with longer delay between heparin administration and diagnostic angiogram or patients with short delay between symptom onset and admission. CONCLUSIONS: There is no benefit of high dose bolus heparin on early patency compared with no or low dose heparin. PMID- 10716461 TI - Clinical and angiographic outcomes in patients with previous coronary artery bypass graft surgery treated with primary balloon angioplasty for acute myocardial infarction. Second Primary Angioplasty in Myocardial Infarction Trial (PAMI-2) Investigators. AB - OBJECTIVES: We sought to characterize the presenting characteristics of patients with previous coronary artery bypass graft surgery (CABG) and acute myocardial infarction (AMI) and to determine the angiographic success rate and clinical outcomes of a primary percutaneous transluminal coronary angioplasty (PTCA) strategy. BACKGROUND: Patients who have had previous CABG and AMI comprise a high risk group with decreased reperfusion success and increased mortality after thrombolytic therapy. Little is known about the efficacy of primary PTCA in AMI. METHODS: Early cardiac catheterization was performed in 1,100 patients within 12 h of onset of AMI at 34 centers in the prospective, controlled Second Primary Angioplasty in Myocardial Infarction trial (PAMI-2), followed by primary PTCA when appropriate. Data were collected by independent study monitors, end points were adjudicated and films were read at an independent core laboratory. RESULTS: Of 1,100 patients with AMI, 58 (5.3%) had undergone previous CABG. The infarct related vessel in these patients was a bypass graft in 32 patients (55%) and a native coronary artery in 26 patients. Compared with patients without previous CABG, patients with previous CABG were older and more frequently had a previous myocardial infarction and triple-vessel disease. Coronary angioplasty was less likely to be performed when the infarct-related vessel was a bypass graft rather than a native coronary artery (71.9% vs. 89.8%, p = 0.001); Thrombolysis in Myocardial Infarction trial (TIMI) flow grade 3 was less frequently achieved (70.2% vs. 94.3%, p < 0.0001); and in-hospital mortality was increased (9.4% vs. 2.6%, p = 0.02). As a result, mortality at six months was 14.3% versus 4.1% in patients with versus without previous CABG (p = 0.001). By multivariate analysis, independent determinants of late mortality in the entire study group were advanced age, triple-vessel disease, Killip class and post-PTCA TIMI flow grade <3. CONCLUSIONS: Reperfusion success of a primary PTCA strategy in patients with previous CABG, although favorable with respect to historic control studies, is reduced as compared with that in patients without previous CABG. New approaches are required to treat patients with previous CABG and AMI, especially when the infarct-related vessel is a diseased saphenous vein graft. PMID- 10716462 TI - Procedural results and late clinical outcomes after percutaneous interventions using long (> or = 25 mm) versus short (< 20 mm) stents. AB - OBJECTIVES: To evaluate clinical outcomes after the use of long coronary stents. BACKGROUND: The use of long slotted-tube stents has been recently approved in the U.S. to treat long lesions or dissections. Procedural success and long-term outcomes of long versus short stents have not been established. METHODS: We evaluated procedural success, major in-hospital complications, target lesion revascularization and long-term (one year) clinical outcomes in 1,226 consecutive patients (1,259 native coronary lesions) who underwent a single vessel intervention using a single long (> or =25 mm, 116 patients) or short (<20 mm, 1,110 patients) tubular-slotted stent. RESULTS: Patients treated with long stents had more diffuse (>10 mm length) lesions (63% vs. 28%, p = 0.001). The mean stent length was 28 +/- 5 mm versus 15 +/- 2 mm for long versus short stent groups (p = 0.001). Overall procedural success was similar in the long versus short stent groups (96% vs. 98%, p = 0.08). However, major in-hospital complications tended to occur more frequently in patients treated with longer stents (3.4% vs. 1.0%, p = 0.04). The rate of periprocedural non-Q-wave myocardial infarction (MI) (creatine kinase-MB > or =5 times normal) was notably higher after long stent implantation (23% vs. 11%, p = 0.001). Target lesion revascularization at one year was 14.5% vs. 13.8% (p = 0.69), and target vessel revascularization rate was 19.6% vs. 17.3% (p = 0.41) in the long versus short stent group, respectively. There was no difference in one year mortality (2.5% vs. 3.5%, p = 0.49) or Q-wave MI (2.7% vs. 1.2%, p = 0.48), and the overall cardiac event-free survival was similar for the two groups (81%). CONCLUSIONS: The use of single coronary long (> or =25 mm) versus short (<20 mm) stents is associated with: 1) somewhat increased major procedural complications, 2) significantly higher frequency of periprocedural non-Q-wave MIs, and 3) equivalent repeat revascularization risk and cardiac event-free survival out-of-hospital up to one year. PMID- 10716463 TI - Application of a continuous regression model of restenosis to saphenous vein grafts after successful percutaneous transluminal coronary angioplasty or directional coronary atherectomy. AB - OBJECTIVES: To evaluate a quantitative model of restenosis in patients with vein graft disease undergoing percutaneous transluminal coronary angioplasty (PTCA) or directional coronary atherectomy (DCA). BACKGROUND: A quantitative relationship between acute gain and late loss has been developed to describe the late changes in lumen dimension after native vessel coronary intervention. This same relationship may also be seen after treatment of saphenous vein graft disease. METHODS: Patients with native coronary artery stenoses (CAVEAT-I) or saphenous vein graft lesions (CAVEAT-II) were randomized to either DCA or PTCA, and data from these trials were analyzed retrospectively. Angiographic results of the target lesions were reviewed, and each lesion was assessed for vessel caliber and reference diameter, absolute minimal lumen diameter, percent diameter stenosis, percent stenosis of the cross-sectional area, acute gain and late loss. Linear regression models were used to determine late loss and to detect differences in angiographic outcomes. RESULTS: Vein grafts had significantly larger reference vessel diameters than native coronary arteries; they also had significantly more acute gain and more late loss. Directional coronary atherectomy was associated with a larger acute gain in both studies. Patients undergoing DCA also experienced greater late loss although the effect was statistically significant only in the CAVEAT-I study. After adjusting for the acute gain, the treatment effect on late loss became nonsignificant in both studies. CONCLUSIONS: In patients undergoing DCA or PTCA of saphenous vein graft narrowings, the relationship between late loss and acute gain is also demonstrated, similar to the device-independent relationships seen in native coronary lesions. In CAVEAT II, larger degrees of acute gain were also associated with higher degrees of late lumen loss. PMID- 10716464 TI - Functional significance of recruitable collaterals during temporary coronary occlusion evaluated by 99mTc-sestamibi single-photon emission computerized tomography. AB - OBJECTIVES: The present study evaluated the impact of recruitable collaterals on regional myocardial perfusion measured by 99mtechnetium (Tc)-sestamibi single photon emission computerized tomography (SPECT) during temporary coronary occlusion and related these estimates to the coronary wedge pressure and electrocardiographic (ECG) ST-segment changes. BACKGROUND: Clinical variables (angina and ECG changes) and intracoronary flow and pressure recordings have indicated a protective role of recruitable collaterals on myocardial perfusion during percutaneous transluminal coronary angioplasty (PTCA). METHODS: Thirty patients (mean age 55 years, SD 9; 20 men) with stable angina pectoris and proximal nonocluding single-vessel left anterior descending coronary artery (LAD) stenosis scheduled for PTCA were included. Visualization of recruitable collaterals by ipsilateral and contralateral contrast injection, registration of coronary wedge pressure and injection of 99mTc-sestamibi during 90-s LAD occlusions were undertaken. A rest perfusion study was performed within four days before PTCA. As an estimate of the severity of regional hypoperfusion during occlusion, an occlusion/rest count ratio was calculated (mean defect pixel count during occlusion divided by mean pixel count in identical regions at rest). RESULTS: The scintigraphic occlusion/rest count ratio was higher in patients with recruitable collaterals (n = 16), 67 +/- 11%, compared to patients without collaterals (n = 14), 60 +/- 6% (p < 0.05). The occlusion/rest count ratio correlated with the coronary wedge pressure (R2 = 0.34; p < 0.001). The occlusion/rest count ratio was lower, 61 +/- 6%, in patients with ST-segment elevation (n = 23) versus 74 +/- 9% in patients without ST-segment elevation (n = 7) (p < 0.0001). CONCLUSIONS: Using 99mTc-sestamibi SPECT imaging during brief episodes of coronary occlusion, the severity of regional myocardial hypoperfusion was reduced by the presence of recruitable collaterals in a selected patient population with proximal LAD stenoses. Our results demonstrate a protective effect of recruitable collaterals on myocardial perfusion during temporary coronary occlusion. PMID- 10716465 TI - Large, sustained cardiac lipid peroxidation and reduced antioxidant capacity in the coronary circulation after brief episodes of myocardial ischemia. AB - OBJECTIVES: We sought to investigate whether a brief episode of myocardial ischemia produces a detectable cardiac oxidative stress in patients undergoing elective coronary angioplasty (PTCA). BACKGROUND: Although cardiac oxidative stress has been clearly demonstrated in experimental models of ischemia reperfusion, its presence in patients after transient myocardial ischemia is still unclear. METHODS: In order to evaluate oxidative stress in ischemic cardiac regions, plasma conjugated dienes (CD), lipid hydroperoxides (ROOHs) and total antioxidant capacity (TRAP), independent indexes of oxidative stress, were measured in the aorta and great cardiac vein (GCV) before (t0), 1, (t1), 5 (t5) and 15 min (t15) after first balloon inflation in 15 patients undergoing PTCA on left anterior descending coronary artery (Group 1); six patients with right coronary artery stenosis (Group 2), which is not drained by the GCV, were studied as controls. RESULTS: In Group 1 at baseline, CD and ROOHs levels were higher in GCV than in aorta (p < 0.01 for both), and TRAP levels were lower (p < 0.01). Aortic levels of CD, ROOHs and TRAP did not change at any time after to; venous levels of CD and ROOHs levels markedly increased at t1, at t5 and remained elevated at t15 (p < 0.01 for all comparisons vs. to); venous levels of TRAP decreased at t1 and t5 (p < 0.01 vs. t0) and returned to normal at t15. In Group 2, CD, ROOHs and TRAP levels were similar in the aorta and GCV and did not change throughout the study. CONCLUSIONS: Short episodes of myocardial ischemia during PTCA induce a sustained oxidative stress, which is detectable in the venous effluent of reperfused myocardium. PMID- 10716466 TI - Gemfibrozil, nicotinic acid and combination therapy in patients with isolated hypoalphalipoproteinemia: a randomized, open-label, crossover study. AB - OBJECTIVES: To assess the effects of nicotinic acid (NA), gemfibrozil and combination therapy on the lipid profile of patients with clinical atherosclerotic disease and isolated hypoalphalipoproteinemia. BACKGROUND: Isolated hypoalphalipoproteinemia (low high density lipoprotein cholesterol [HDL C] alone) accounts for a significant percentage of patients with premature atherosclerosis. However, it remains unclear whether currently available pharmacotherapy has the ability to favorably affect the lipid profile and therefore potentially reduce clinical events. METHODS: Twenty-three patients with clinically well-defined atherosclerosis and isolated hypoalphalipoproteinemia were prospectively randomized to receive gemfibrozil, NA or combination therapy in an open-label, crossover design trial to assess the effects on serum lipids. Lipid profiles and other relevant laboratory variables were monitored while the patients were on and off pharmacologic lipid-modulating therapy. RESULTS: In those 14 patients able to tolerate all forms of pharmacotherapy, HDL-C of 0.89 +/ 0.17 mmol/liter (34.5 +/- 6.5 mg/dl) increased by 15%, to 1.02 +/- 0.18 mmol/liter (39.7 +/- 7.1 mg/dl), while taking gemfibrozil (1,200 mg/day); by 35%, to 1.20 +/- 0.21 mmol/liter (46.5 +/- 8.1 mg/dl), while taking NA (mean dose 2,250 mg/day); and by 45%, to 1.29 +/- 0.19 mmol/liter (50.0 +/- 7.5 mg/dl), while taking combination therapy of gemfibrozil plus NA (p < 0.001 for all interventions as compared with baseline/washout; p < 0.005 NA vs. gemfibrozil; p < 0.001 combination therapy vs. gemfibrozil alone; p = 0.088 combination therapy vs. NA alone). Statistically significant favorable alterations were also observed with low density lipoprotein cholesterol (LDL-C), LDL-C/HDL-C, non-HDL-C/HDL-C, apolipoprotein (Apo) B and Apo B/Apo A1. CONCLUSIONS: In the majority of patients with clinical atherosclerotic disease and isolated hypoalphalipoproteinemia, pharmacologic therapy to raise HDL-C is not only feasible but is also effective with currently available agents, particularly when used in combination. PMID- 10716467 TI - Serum insulin-like growth factor-I level is independently associated with coronary artery disease progression in young male survivors of myocardial infarction: beneficial effects of bezafibrate treatment. AB - OBJECTIVES: We investigated whether the effect of bezafibrate on progression of coronary atherosclerosis in the BEzafibrate Coronary Atherosclerosis Intervention Trial (BECAIT) was related to insulin-like growth factor (IGF)-I and glucose insulin homeostasis. BACKGROUND: BECAIT, the first double-blind, placebo controlled, randomized, serial angiographic trial of a fibrate compound, demonstrated that progression of focal coronary atherosclerosis in young patients after infarction could be retarded by bezafibrate treatment. METHODS: The treatment effects on serum concentrations of IGF-I and insulin-like growth factor binding protein (IGFBP)-1, as well as on basal and postload glucose and insulin levels, were examined, and on-trial determinations were related to the angiographic outcome measurements. RESULTS: Bezafibrate treatment resulted in a significant reduction of serum IGF-I levels, both at two and five years, and on trial serum IGF-I levels were directly related to changes in both minimal lumen diameter (r = 0.25, p < 0.05) and mean segment diameter (r = 0.29, p < 0.05). In contrast, none of the available indexes of insulin resistance (homeostasis model assessment estimate, basal and postload plasma insulin concentrations and serum IGFBP-1 levels) were related to the angiographic changes, nor were they significantly affected by bezafibrate treatment. Multiple stepwise regression analysis showed that the relation between on-trial serum IGF-I level and coronary artery disease (CAD) progression was independent of baseline angiographic score, age, body mass index, serum lipoprotein and plasma fibrinogen concentrations and measures of glucose-insulin homeostasis. CONCLUSIONS: IGF-I could be implicated in the progression of premature CAD, and a reduction of serum IGF-I concentration could account partly for the effect of bezafibrate on progression of focal coronary atherosclerosis. PMID- 10716468 TI - Clinical significance of increased plasma concentration of macrophage colony stimulating factor in patients with angina pectoris. AB - OBJECTIVES: To determine the effect of macrophage colony-stimulating factor (MCSF) on atherogenesis in patients with coronary artery disease (CAD), we assessed the relation between the plasma concentration of MCSF and the incidence of acute coronary events in patients with CAD. BACKGROUND: Cytokines such as MCSF play a central role in inflammatory and proliferative responses in patients with acute coronary syndromes. However, the effect of MCSF on the clinical course in patients with CAD is still not known. METHODS: We measured the plasma MCSF concentration in 142 patients with documented CAD (62 +/- 9 years) and followed up for a mean period of 14 +/- 6 months. The study included 97 patients with stable angina (SA), 45 patients with unstable angina (UA) and 22 age-matched control subjects. The predictors of coronary events were analyzed by using a Cox proportional hazards model. RESULTS: The mean plasma MCSF concentration in patients with UA was significantly higher than that in patients with SA and in control subjects (981 +/- 277 vs. 693 +/- 223 vs. 680 +/- 158 pg/ml, p < 0.001). The mean plasma MCSF concentration in the 20 patients with coronary events was significantly higher than that in patients without coronary events (1,192 +/- 232 vs. 690 +/- 213 pg/ml, p < 0.001). The predictors of unfavorable outcome were an increased MCSF concentration, the presence of CAD and a low ejection fraction. CONCLUSIONS: These findings suggest that an increased circulating MCSF concentration reflects atherosclerotic progression in patients with CAD and predicts future cardiac events. PMID- 10716469 TI - Prognostic implications of TIMI flow grade in the infarct related artery compared with continuous 12-lead ST-segment resolution analysis. Reexamining the "gold standard" for myocardial reperfusion assessment. AB - OBJECTIVE: To compare the prognostic significance of reperfusion assessment by Thrombolysis in Myocardial Infarction (TIMI) flow grade in the infarct related artery and ST-segment resolution analysis, by correlating with clinical outcomes in patients with acute myocardial infarction (AMI). BACKGROUND: Angiographic assessment, based on epicardial coronary anatomy, has been considered the "gold standard" for reperfusion. The electrocardiogram (ECG) monitoring provides a noninvasive, real-time physiologic marker of cellular reperfusion and may better predict clinical outcomes. METHODS: Two hundred fifty-eight AMI patients from the Thrombolytics and Myocardia Infarction phase 7 and Global Utilization of Streptokinase tPA for Occluded coronary arteries phase 1 trials were stratified based on blinded, simultaneous reperfusion assessment on the acute angiogram (divided into TIMI grades 0 & 1, TIMI grade 2 and TIMI grade 3) and ST-segment resolution analysis (divided into: <50% ST-segment elevation resolution or reelevation and > or =50% ST-segment elevation resolution). In-hospital mortality, congestive heart failure (CHF) and combined mortality or CHF were compared to determine the prognostic significance of reperfusion assessment by each modality using chi-square and Fisher's Exact tests for univariable correlation and logistic regression analysis for univariable and multivariable prediction models. RESULTS: By logistic regression analysis, ST-segment resolution patterns were an independent predictor of the combined outcome of mortality or CHF (p = 0.024), whereas TIMI flow grade was not (p = 0.693). Among the patients determined to have failed reperfusion by TIMI flow grade assessment (TIMI flow grade 0 & 1), the ST-segment resolution of > or =50% identified a subgroup with relatively benign outcomes with the incidence of the combined end point of mortality or CHF 17.2% versus 37.2% in those without ST-segment resolution (p = 0.06). CONCLUSION: Continuous 12-lead ECG monitoring can be an inexpensive and reliable modality for monitoring nutritive reperfusion status and to obtain prognostic information in patients with AMI. PMID- 10716470 TI - A decrease in diastolic blood pressure combined with an increase in systolic blood pressure is associated with a higher cardiovascular mortality in men. AB - OBJECTIVES: The study evaluated the risk of cardiovascular mortality according to combined spontaneous (non-treatment-related) changes in both systolic and diastolic blood pressure (BP). BACKGROUND: Long-term longitudinal changes in blood pressure may be a more accurate determinant of cardiovascular risk since changes in systolic or diastolic blood pressure over a period of time reflect the evolution of arterial and arteriolar alterations. METHODS: Two independent French male cohorts were studied: the IPC cohort (Investigations Preventives et Cliniques) composed of 15,561 men aged 20 to 82 years who had had two visits spaced four to 10 years apart, and the Paris Prospective Study composed of 6,246 men aged 42 to 53 years, examined annually for a period of four years. None of the subjects were taking antihypertensive medication. Annual changes in BP were estimated, and subjects were divided into groups according to the increase, lack of change, or decrease of systolic or diastolic BP. Nine groups were formed by combining the changes of systolic and diastolic BP. Cardiovascular mortality was assessed for a mean period of 13.5 years for the IPC Study and 17 years for the Paris Prospective Study. RESULTS: In both cohorts, after adjustment for age and major risk factors, the group with an increase in systolic and a decrease in diastolic BP presented the highest relative risk of cardiovascular mortality compared to the group with no changes in either systolic or diastolic BP (relative risk: 2.07 [1.05 to 4.06] in the IPC Study and 2.16 [1.16 to 4.01] in the Paris Prospective Study). CONCLUSIONS: Assessment of spontaneous changes of BP over a long period of time can contribute to the evaluation of cardiovascular risk. Subjects whose systolic BP increased while their diastolic BP decreased had the highest cardiovascular risk independently of absolute values of BP or other risk factors. PMID- 10716471 TI - The prognostic implications of renal insufficiency in asymptomatic and symptomatic patients with left ventricular systolic dysfunction. AB - OBJECTIVES: The present analysis examines the prognostic implications of moderate renal insufficiency in patients with asymptomatic and symptomatic left ventricular systolic dysfunction. BACKGROUND: Chronic elevations in intracardiac filling pressures may lead to progressive ventricular dilation and heart failure progression. The ability to maintain fluid balance and prevent increased intracardiac filling pressures is critically dependent on the adequacy of renal function. METHODS: This is a retrospective analysis of the Studies of Left Ventricular Dysfunction (SOLVD) Trials, in which moderate renal insufficiency is defined as a baseline creatinine clearance <60 ml/min, as estimated from the Cockroft-Gault equation. RESULTS: In the SOLVD Prevention Trial, multivariate analyses demonstrated moderate renal insufficiency to be associated with an increased risk for all-cause mortality (Relative Risk [RR] 1.41; p = 0.001), largely explained by an increased risk for pump-failure death (RR 1.68; p = 0.007) and the combined end point death or hospitalization for heart failure (RR 1.33; p = 0.001). Likewise, in the Treatment Trial, multivariate analyses demonstrated moderate renal insufficiency to be associated with an increased risk for all-cause mortality (RR 1.41; p = 0.001), also largely explained by an increased risk for pump-failure death (RR 1.49; p = 0.007) and the combined end point death or hospitalization for heart failure (RR 1.45; p = 0.001). CONCLUSIONS: Even moderate degrees of renal insufficiency are independently associated with an increased risk for all-cause mortality in patients with heart failure, largely explained by an increased risk of heart failure progression. These data suggest that, rather than simply being a marker of the severity of underlying disease, the adequacy of renal function may be a primary determinant of compensation in patients with heart failure, and therapy capable of improving renal function may delay disease progression. PMID- 10716472 TI - Expiratory flow limitation as a determinant of orthopnea in acute left heart failure. AB - OBJECTIVES: To assess the contribution of expiratory flow limitation (FL) in orthopnea during acute left heart failure (LHF). BACKGROUND: Orthopnea is typical of acute LHF, but its mechanisms are not completely understood. In other settings, such as chronic obstructive pulmonary disease, dyspnea correlates best with expiratory FL and can, therefore, be interpreted as, in part, the result of a hyperinflation-related increased load to the inspiratory muscles. As airway obstruction is common in acute LHF, postural FL could contribute to orthopnea. METHODS: Flow limitation was assessed during quiet breathing by applying a negative pressure at the mouth throughout tidal expiration (negative expiratory pressure [NEP]). Flow limitation was assumed when expiratory flow did not increase during NEP. Twelve patients with acute LHF aged 40-98 years were studied seated and supine and compared with 10 age-matched healthy subjects. RESULTS: Compared with controls, patients had rapid shallow breathing with slightly increased minute ventilation and mean inspiratory flow. Breathing pattern was not influenced by posture. Flow limitation was observed in four patients when seated and in nine patients when supine. In seven cases, FL was induced or aggravated by the supine position. This coincided with orthopnea in six cases. Only one out of the five patients without orthopnea had posture dependent FL. Control subjects did not exhibit FL in either position. CONCLUSIONS: Expiratory FL appears to be common in patients with acute LHF, particularly so when orthopnea is present. Its postural aggravation could contribute to LHF-related orthopnea. PMID- 10716473 TI - Peripartum cardiomyopathy: analysis of clinical outcome, left ventricular function, plasma levels of cytokines and Fas/APO-1. AB - OBJECTIVES: 1) To evaluate the outcome of patients with peripartum cardiomyopathy (PPC) on current treatment for heart failure, 2) to assess the circulating plasma levels of cytokines and Fas receptors and 3) to identify predictors of prognosis. BACKGROUND: Previous studies in patients with PPC were done when angiotensin converting enzyme (ACE) inhibitors and beta-adrenergic blocking agents were not routinely used in heart failure. Inflammatory cytokines play an important role in the pathogenesis and progression of heart failure of other etiologies. However, there is a paucity of data regarding cytokine expression in patients with PPC. Plasma concentrations of Fas receptors (an apoptosis-signalling receptor) have not been reported in this population. METHODS: We followed prospectively 29 consecutive black women with PPC. All patients were treated with diuretics, digoxin, enalapril and carvedilol. Echocardiograms were performed at baseline and after six months of treatment. Cytokine and soluble Fas/APO-1 plasma levels were measured at baseline. RESULTS: Tumor necrosis factor-alpha, interleukin-6 and Fas/APO-1 levels were significantly elevated in the study patients compared with 20 healthy volunteers. Eight patients died. sFas/APO-1 levels were significantly higher in patients who died compared with survivors (8.98 +/- 4.5 vs. 5.33 +/- 3 U/ml, respectively, p = 0.02). At six months, ejection fraction improved from 26.7 +/- 10 to 42.7 +/- 16%, p = 0.00003, with an increment of more than 10 U in 10 patients (28.1 +/- 4 to 51.9 +/- 8%, p = 0.000008). CONCLUSIONS: Cytokine and sFas levels are elevated in patients with PPC. Despite treatment with ACE inhibitors and beta-blockers, mortality remains high. However, in 34% of the patients, left ventricular function almost completely normalized. PMID- 10716474 TI - Correction of endothelial dysfunction in chronic heart failure: additional effects of exercise training and oral L-arginine supplementation. AB - OBJECTIVES: The aim of this study was to analyze whether L-arginine (L-arg.) has comparable or additive effects to physical exercise regarding endothelium dependent vasodilation in patients with chronic heart failure (CHF). BACKGROUND: Endothelial dysfunction in patients with CHF can be corrected by both dietary supplementation with L-arg. and regular physical exercise. METHODS: Forty patients with severe CHF (left ventricular ejection fraction 19 +/- 9%) were randomized to an L-arg. group (8 g/day), a training group (T) with daily handgrip training, L-arg. and T (L-arg. + T) or an inactive control group (C). The mean internal radial artery diameter was determined at the beginning and after four weeks in response to brachial arterial administration of acetylcholine (ACh) (7.5, 15, 30 microg/min) and nitroglycerin (0.2 mg/min) with a transcutaneous high-resolution 10 MHz A-mode echo tracking system coupled with a Doppler device. The power of the study to detect clinically significant differences in endothelium-dependent vasodilation was 96.6%. RESULTS: At the beginning, the mean endothelium-dependent vasodilation in response to ACh, 30 microg/min was 2.54 +/- 0.09% (p = NS between groups). After four weeks, internal radial artery diameter increased by 8.8 +/- 0.9% after ACh 30 microg/min in L-arg. (p < 0.001 vs. C), by 8.6 +/- 0.9% in T (p < 0.001 vs. C) and by 12.0 +/- 0.3% in L-arg. +/- T (p < 0.005 vs. C, L-arg. and T). Endothelium-independent vasodilation as assessed by infusion of nitroglycerin was similar in all groups at the beginning and at the end of the study. CONCLUSIONS: Dietary supplementation of L-arg. as well as regular physical exercise improved agonist-mediated, endothelium-dependent vasodilation to a similar extent. Both interventions together seem to produce additive effects with respect to endothelium-dependent vasodilation. PMID- 10716475 TI - Angiotensin II type 1 receptor antagonist decreases plasma levels of tumor necrosis factor alpha, interleukin-6 and soluble adhesion molecules in patients with chronic heart failure. AB - OBJECTIVES: To evaluate the effects of an angiotensin (Ang II) type 1 receptor antagonist on immune markers in patients with congestive heart failure (CHF). BACKGROUND: Ang II stimulates production of immune factors via the Ang II type 1 receptor in vitro, and the long-term effects of Ang II type 1 receptor antagonists on plasma markers of immune activation are unknown in patients with CHF. METHODS: Twenty-three patients with mild to moderate CHF with left ventricular dysfunction were randomly divided into two groups: treatment with Ang II type 1 receptor (candesartan cilexetil) (n = 14) or placebo (n = 9). We measured plasma levels of immune factors such as tumor necrosis factor alpha (TNFalpha), interleukin-6 (IL-6), soluble intercellular adhesion molecule-1 (sICAM-1) and soluble vascular cell adhesion molecule-1 (sVCAM-1). We also measured plasma levels of the neurohumoral factors such as atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) and cyclic guanosine monophosphate (cGMP), a biological marker of ANP and BNP. RESULTS: Plasma levels of TNFalpha, IL-6, sICAM-1 and sVCAM-1 were increased in the 23 CHF patients compared with normal subjects and significantly decreased after 14 weeks of candesartan cilexetil treatment, but did not change in the placebo group. Plasma levels of BNP, which is a marker of ventricular injury, significantly decreased, and the molar ratio of plasma cGMP to cardiac natriuretic peptides (ANP + BNP) was significantly increased after candesartan cilexetil treatment, but did not change in the placebo group. CONCLUSIONS: These findings suggest that 14 weeks of treatment with an Ang II type 1 receptor antagonist (candesartan cilexetil) decreased plasma levels of the immune markers such as TNFalpha, IL-6, sICAM-1 and sVCAM-1 and that it improved the biological compensatory action of endogenous cardiac natriuretic peptides in patients with mild to moderate CHF. PMID- 10716476 TI - Combined assessment of T-wave alternans and late potentials used to predict arrhythmic events after myocardial infarction. A prospective study. AB - OBJECTIVES: The aim of the present study was to determine whether the combination of two markers that reflect depolarization and repolarization abnormalities can predict future arrhythmic events after acute myocardial infarction (MI). BACKGROUND: Although various noninvasive markers have been used to predict arrhythmic events after MI, the positive predictive value of the markers remains low. METHODS: We prospectively assessed T-wave alternans (TWA) and late potentials (LP) by signal-averaged electrocardiogram (ECG) and ejection fraction (EF) in 102 patients with successful determination results after acute MI. The TWA was analyzed using the power-spectral method during supine bicycle exercise testing. No antiarrhythmic drugs were used during the follow-up period. The study end point was the documentation of ventricular arrhythmias. RESULTS: The TWA was present in 50 patients (49%), LP present in 21 patients (21%), and an EF <40% in 28 patients (27%). During a follow-up period of 13 +/- 6 months, symptomatic, sustained ventricular tachycardia or ventricular fibrillation occurred in 15 patients (15%). The event rates were significantly higher in patients with TWA, LP, or an abnormal EF. The sensitivity and the negative predictive value of TWA in predicting arrhythmic events were very high (93% and 98%, respectively), whereas its positive predictive value (28%) was lower than those for LP and EF. The highest positive predictive value (50%) was obtained when TWA and LP were combined. CONCLUSIONS: The combined assessment of TWA and LP was associated with a high positive predictive value for an arrhythmic event after acute MI. Therefore, it could be a useful index to identify patients at high risk of arrhythmic events. PMID- 10716477 TI - Outcomes of cardiac surgery in patients > or = 80 years: results from the National Cardiovascular Network. AB - OBJECTIVES: The purpose of this study was to evaluate characteristics and outcomes of patients age > or =80 undergoing cardiac surgery. BACKGROUND: Prior single-institution series have found high mortality rates in octogenarians after cardiac surgery. However, the major preoperative risk factors in this age group have not been identified. In addition, the additive risks in the elderly of valve replacement surgery at the time of bypass are unknown. METHODS: We report in hospital morbidity and mortality in 67,764 patients (4,743 octogenarians) undergoing cardiac surgery at 22 centers in the National Cardiovascular Network. We examine the predictors of in-hospital mortality in octogenarians compared with those predictors in younger patients. RESULTS: Octogenarians undergoing cardiac surgery had fewer comorbid illnesses but higher disease severity and surgical urgency than younger patients. Octogenarians had significantly higher in-hospital mortality after cardiac surgery than younger patients: coronary artery bypass grafting (CABG) only (8.1% vs. 3.0%), CABG/aortic valve (10.1% vs. 7.9%), CABG/mitral valve (19.6% vs. 12.2%). In addition, they had twice the incidence of postoperative stroke and renal failure. The preoperative clinical factors predicting CABG mortality in the very elderly were quite similar to those for younger patients with age, emergency surgery and prior CABG being the powerful predictors of outcome in both age categories. Of note, elderly patients without significant comorbidity had in-hospital mortality rates of 4.2% after CABG, 7% after CABG with aortic valve replacement (CABG/AVR), and 18.2% after CABG with mitral valve replacement (CABG/MVR). CONCLUSIONS: Risks for octogenarians undergoing cardiac surgery are less than previously reported, especially for CABG only or CABG/AVR. In selected octogenarians without significant comorbidity, mortality approaches that seen in younger patients. PMID- 10716478 TI - Early and long-term (one-year) effects of the association of aspirin and oral anticoagulant on thrombi and morbidity after replacement of the mitral valve with the St. Jude medical prosthesis: a clinical and transesophageal echocardiographic study. AB - OBJECTIVES: The aim of the study was to test the value of low dose aspirin associated with standard oral anticoagulants (OAC) after mechanical mitral valve replacement (MMRV) to reduce strands, thrombi and thromboembolic events. BACKGROUND: Strands and thrombi are thought to increase the risk of embolic events after MMVR, particularly in the immediate postoperative period. METHODS: Two hundred twenty-nine patients were prospectively recruited: 109 patients (group A+) were randomly assigned to aspirin (200 mg per day) with OAC and 120 patients (group A-) to OAC alone (international normalized ratio 2.5 to 3.5). All patients were subjected to multiplane transesophageal echocardiography at nine days and five months and were followed up for one year. RESULTS: At nine days and five months, there was a high and comparable incidence of strands in the two groups (group A+: 44%, 58%; group A-: 49%, 63%). However, the incidence of nonobstructive periprosthetic valve thrombi was significantly lower in group A+ at 9 days: 5% versus 13%, p = 0.03. Total thromboembolic events were reduced in group A+ (9% vs. 25%, p = 0.004) although there was an increased incidence of gastrointestinal hemorrhage (7% vs. 0%). Overall mortality was 9% in group A+ and 4% in group A-. Valve-related events were similar in both groups. Early thrombi, but not strands, were associated with higher morbidity, especially thromboembolic events (30% vs. 13%, p = 0.003). CONCLUSIONS: One year after MMVR, the association of aspirin with OAC reduced thrombi and thromboembolic events, but not morbidity, due to an increase in hemorrhagic complications. PMID- 10716479 TI - Observed and relative survival after aortic valve replacement. AB - OBJECTIVES: We sought to evaluate the effects of a number of factors that can potentially determine the optimal time for aortic valve replacement (AVR) and the observed and relative survival after the operation. BACKGROUND: Aortic valve replacement is performed in patients within a wide age span, but the proportion of elderly patients is increasing. In survival analyses, adjustment for the effects of age is therefore essential. Analysis of relative survival provides additional information on excess or disease-specific mortality and its risk factors. METHODS: Survival was analyzed in 2,359 patients (1,442 without and 917 with concomitant coronary artery bypass graft surgery) undergoing their first AVR. By relating observed survival to that expected among the general Swedish population stratified by age, gender and five-year calendar period, the relative survival and disease-specific survival were estimated. RESULTS: Early mortality after AVR (death within 30 days) was 5.6%. Relative survival rates (excluding early deaths) after 5, 10 and 15 years were 94.6%, 84.7% and 74.9%, respectively. There was an excess risk of dying during the entire follow-up period. Advanced New York Heart Association functional class, preoperative atrial fibrillation and pure aortic regurgitation were independent risk factors for observed and relative survival. Patients in the oldest age group showed decreased observed survival but excellent relative survival. CONCLUSIONS: Old age was not a risk factor for excess mortality after AVR, whereas atrial fibrillation decreased relative survival substantially. PMID- 10716480 TI - Exercise-induced U-wave alterations as a marker of well-developed and well functioning collateral vessels in patients with effort angina. AB - OBJECTIVES: We sought to determine whether exercise-induced U-wave alterations are observed in association with well-developed and well-functioning collateral vessels. BACKGROUND: Although exercise-induced electrocardiographic (ECG) U-wave alterations including negative and prominent U waves have been established as a marker of significant or critical narrowing of a major coronary artery, the relation between this finding and the degree of collateral development has not yet been determined. METHODS: Patients with stable effort angina were divided into two groups according to the presence (group A, n = 46) or absence (group B, n = 79) of exercise-induced either negative or prominent U waves in the precordial leads; the clinical profiles, coronary angiographic findings and also ischemic status during 60 s of coronary balloon occlusion were compared between the two groups. RESULTS: The incidence of severe angina (CCS [Canadian Cardiovascular Society] class III or IV) was higher (p < 0.05) in group A (52%) than in group B (32%) patients. Good collateral vessels (Rentrop grade 2 or 3) into the perfusion territory of the culprit vessel were observed more frequently (p < 0.05) in group A (70%) than in group B (43%) patients. Coronary balloon angioplasty was carried out in 23 patients of group A and 40 patients of group B. Both ischemic ST changes (52% vs. 85%) and angina (57% vs. 80%) during balloon inflation were less (p < 0.05) frequently observed in group A than in group B. The incidence of no apparent myocardial ischemia with ST deviation or angina during the balloon inflation was higher (p < 0.05) in group A (39%) than in group B (10%) patients. In the prediction of the absence of myocardial ischemia during balloon inflation by the presence of exercise-induced U-wave alterations, the sensitivity was 69% (9/13) and the specificity was 72% (36/50) in the study patients. CONCLUSIONS: Exercise-induced U-wave alterations are a marker for well developed collateral circulation in patients with stable but severe effort angina. This finding is also highly predictive of the absence of myocardial ischemia during transient coronary balloon occlusion and possibly of low-risk for development of acute myocardial infarction or hemodynamic instability upon abrupt closure of the culprit coronary artery. PMID- 10716481 TI - Clinical features and management of isolated cleft mitral valve in childhood. AB - OBJECTIVES: We reviewed an institutional experience of isolated cleft mitral valve (ICMV), its clinical features, and management in a pediatric population. BACKGROUND: As ICMV is relatively uncommon, earlier reports highlighted its anatomical and echocardiographic features. Few studies have collated their clinical features with their outcome. METHODS: All patients with ICMV were retrospectively reviewed. Patients who were considered to have an atrioventricular septal defect or variant were excluded. RESULTS: Twenty patients (9 male, 11 female) were diagnosed with ICMV. Seven patients had associated cardiac lesions. The median age of diagnosis was 5.2 years (range 0.4 to 13.6 years). Echocardiography aided by color Doppler demonstrated the ICMV in all patients. However, an incomplete diagnosis was made in 4 of 20 patients before surgery. The severity of the mitral regurgitation (MR) at presentation was mild in 11, moderate in 8, and severe in 1 patient. In the 13 patients without associated cardiac lesions, 5 underwent mitral valve (MV) repair at median age of 5.2 years (range 1.2 to 7.7 years) for moderate to severe MR, 4 being symptomatic. The severity of the MR in seven of the eight unoperated patients has remained unchanged over the follow-up period (median 8.3 years, range 0.7 to 14.4 years). In total, 10 patients underwent MV repair (median 6.4, range 0.4 to 13.8 years). No patient required MV replacement. None of the 10 patients had more than mild MR over the follow-up period (median 0.6, range 0.2 to 11.0 years). CONCLUSIONS: Now readily diagnosable by echocardiography, ICMV is a correctable cause of MR with a good outcome. Surgery is indicated in those patients with moderate to severe MR and probably should be done early following diagnosis. PMID- 10716482 TI - Atrial flutter in the perinatal age group: diagnosis, management and outcome. AB - OBJECTIVES: The aim of this retrospective study was to evaluate perinatal atrial flutter (AF) and the efficacy of maternally administered antiarrhythmic agents, postpartum management and outcome. BACKGROUND: Perinatal AF is a potentially lethal arrhythmia, and management of this disorder is difficult and controversial. METHODS: Forty-five patients with documented AF were studied retrospectively. RESULTS: Atrial flutter was diagnosed prenatally in 44 fetuses and immediately postnatally in 1 neonate. Fetal hydrops was seen in 20 patients; 17 received maternal therapy, 2 were delivered and 1 was not treated because it had a severe nontreatable cardiac malformation. In the nonhydropic group of 24 patients, 18 were treated and the remaining 6 were delivered immediately. In the hydropic group, 10 received single-drug therapy (digoxin or sotalol) and 7 received multidrug therapy. In the nonhydropic group, 13 received a single drug (digoxin or sotalol) and 5 received multiple drugs. One patient with rapid 1:1 atrioventricular conduction (heart rate 480 beats/min) died in utero and another died due to a combination of severe hydrops because of the AF, sotalol medication, stenosis of the venous duct and hypoplastic placenta. Of the 43 live born infants, 12 were in AF at birth. Electrical cardioversion was successful in eight of nine patients. No recurrences in AF have occurred beyond the neonatal period. Four patients with fetal flutter and hydrops showed significant neurological pathology immediately after birth. CONCLUSIONS: Fetal AF is a serious and threatening rhythm disorder, particularly when it causes hydrops, it may be associated with fetal death or neurological damage. Treatment is required and primarily aimed at reaching an adequate ventricular rate and preferably conversion to sinus rhythm. Digoxin failed in prevention of recurrence at time of delivery in a quarter of our patients, whereas with sotalol no recurrence of AF has been reported, suggesting that class III agents may be the future therapy. Once fetuses with AF survive without neurological pathology, their future is good and prophylaxis beyond the neonatal period is unnecessary. PMID- 10716483 TI - Differential effects of beta-adrenergic agonists and antagonists in LQT1, LQT2 and LQT3 models of the long QT syndrome. AB - OBJECTIVES: To define the cellular mechanisms responsible for the development of life-threatening arrhythmias in response to sympathetic activity in the congenital and acquired long QT syndromes (LCQTS). METHODS: Transmembrane action potentials (AP) from epicardial (EPI), M and endocardial (ENDO) cells and a transmural electrocardiogram were simultaneously recorded from an arterially perfused wedge of canine left ventricle. We examined the effect of beta adrenergic agonists and antagonists on action potential duration (APD90), transmural dispersion of repolarization (TDR) and the development of Torsade de Pointes (TdP) in models of LQT1, LQT2 and LQT3 forms of LQTS. RESULTS: I(Ks) block with chromanol 293B (LQT1) homogeneously prolonged APD90 of the three cell types without increasing TDR. Addition of isoproterenol prolonged QT and APD90 of M but abbreviated that of EPI and ENDO, causing a persistent increase in TDR; Torsade de Pointes developed or could be induced only in the presence of isoproterenol. I(Kr) block with d-sotalol (LQT2) and augmentation of late I(Na) with ATX-II (LQT3) prolonged APD90 of M more than EPI and ENDO, causing increases in QT and TDR. TdP developed in the absence of isoproterenol. In LQT2 isoproterenol initially prolonged, then abbreviated, the APD90 of M but always abbreviated EPI, thus transiently increasing TDR and the incidence of TdP. In LQT3, isoproterenol always abbreviated APD90 of the three cell types, causing a persistent decrease in TDR and suppression of TdP. The arrhythmogenic as well as protective actions of isoproterenol were reversed by propranolol. CONCLUSIONS: Our data suggest that beta-adrenergic stimulation induces TdP by increasing transmural dispersion of repolarization in LQT1 and LQT2 but suppresses TdP by decreasing dispersion in LQT3. The data indicate that beta-blockers are protective in LQT1 and LQT2 but may facilitate TdP in LQT3. PMID- 10716484 TI - Comparative effects of pretreatment with captopril and losartan on cardiovascular protection in a rat model of ischemia-reperfusion. AB - OBJECTIVES: We sought to assess the comparative effects of pretreatment with captopril and losartan on myocardial infarct size and arrhythmias in a rat model of ischemia-reperfusion. BACKGROUND: Angiotensin-converting enzyme (ACE) inhibitors and angiotensin II receptor blockers (ARBs) inhibit the renin angiotensin system in different ways. However, the comparative effects of pretreatment with ACE inhibitors or ARBs on acute myocardial infarct size and arrhythmias are unknown. METHODS: We randomly assigned 117 female Sprague-Dawley rats into three groups: group N was the normal control; group C was given 40 mg/kg body weight per day of captopril in drinking water; and group L was given 40 mg/kg per day of losartan in drinking water. After 10 weeks of pretreatment, 25 rats in each group were subjected to 17 min of left anterior descending coronary artery occlusion and 2 h of reperfusion with hemodynamic and electrocardiographic monitoring. Fourteen rats in each group had blood samples drawn and aortic rings removed to study vascular reactivity. RESULTS: Mortality during ischemia and reperfusion was lower in combined groups L and C than in group N (4.2% vs. 19.2%, p = 0.042). Rats treated with losartan had significantly higher levels of angiotensin II in their plasma. Hemodynamic variables were not significantly different among the three groups. The thresholds of ventricular fibrillation (VF) before occlusion and after reperfusion were significantly higher in groups L and C than in group N (1.99 +/- 0.24 and 1.93 +/- 0.27 vs. 1.23 + 0.17 mA, p = 0.04; 2.13 +/- 0.25 and 1.78 +/- 0.22 vs. 0.95 +/- 0.11 mA, p = 0.001). The average episodes of ventricular tachycardia (VT) and VF per rat were significantly less in groups L and C than in group N (0.96 +/- 0.2 and 1.2 +/- 0.3 vs. 2.8 + 0.4 mA, p < 0.001). Myocardial infarct size was significantly smaller in groups L and C than in group N (34 +/- 3% and 35 +/- 3% vs. 44 +/- 3%, p = 0.031, 0.043). Endothelium-dependent vasorelaxation induced by a calcium ionophore (A23187) was increased in both groups but was only statistically significant in group C (p = 0.020). CONCLUSIONS: Losartan and captopril have similar cardiovascular protective effects in a rat model of ischemia-reperfusion. They increased the threshold of VF, decreased mortality and decreased episodes of VT and VF, as well as decreased myocardial infarct size. PMID- 10716486 TI - Pharmacy madness. PMID- 10716485 TI - Vascular actions of brain natriuretic peptide: modulation by atherosclerosis and neutral endopeptidase inhibition. AB - OBJECTIVES: We sought to define the vascular actions of the cardiac hormone brain natriuretic peptide (BNP) on cellular proliferation and cyclic guanosine monophosphate (cGMP) in human aortic vascular smooth muscle cells (HAVSMCs). Secondly, we investigated BNP and acetylcholine (ACh) vasorelaxations in aortic rings from normal and atherosclerotic rabbits in the presence and absence of long term oral inhibition of neutral endopeptidase (NEP). BACKGROUND: The vascular actions of BNP are not well defined, despite the presence of its receptor in vascular smooth muscle and the upregulation of NEP, the ectoenzyme that degrades BNP, in the vascular wall in atherosclerosis. METHODS: HAVSMCs stimulated with fetal calf serum (FCS) were pulsed with bromodeoxyuridine (BrdU) with and without BNP. The HAVSMCs were incubated in the presence and absence of BNP to assess cGMP. Vasorelaxations to BNP and ACh were assessed in rings in normal and atherosclerotic rabbits in the presence and absence of long-term oral inhibition of NEP, together with assessment of atheroma formation. RESULTS: FCS-stimulated BrdU uptake in HAVSMCs was suppressed with BNP. BNP potentiated cGMP in HAVSMCs. BNP resulted in potent vasorelaxation in normal isolated aortic rings, which were impaired in atherosclerotic versus normal rabbits and preserved with NEP inhibition, which also decreased atheroma formation. Relaxations to ACh, which were also impaired in atherosclerosis, were preserved with inhibition of NEP. CONCLUSIONS: We conclude that BNP potently inhibits vascular smooth muscle cell proliferation and potentiates the generation of cGMP. BNP potently relaxes the normal rabbit aorta, and this response is impaired in atherosclerosis but preserved with inhibition of NEP, together with a reduction in atheroma formation and preservation of relaxations to ACh. PMID- 10716487 TI - The ACC training outcomes survey of recently trained cardiology fellows. PMID- 10716488 TI - Mechanism of protection against vascular smoking-induced changes by hormone replacement therapy. PMID- 10716489 TI - Lack of effect of coenzyme Q on left ventricular function in patients with congestive heart failure. PMID- 10716490 TI - Is the lower mortality in patients treated with aspirin and angiotensin converting enzyme inhibitors due to decreased norepinephrine release? PMID- 10716491 TI - Mercury and idiopathic dilated cardiomyopathy. PMID- 10716492 TI - Risk of sulfonylurea drugs is underappreciated. PMID- 10716493 TI - Describing patients with discordant ventriculoarterial connections. PMID- 10716494 TI - Coronary endothelial dysfunction after Kawasaki disease. PMID- 10716495 TI - A syndrome of lipoatrophy, lactic acidaemia and liver dysfunction associated with HIV nucleoside analogue therapy: contribution to protease inhibitor-related lipodystrophy syndrome. AB - BACKGROUND: Lipodystrophy (LD; peripheral lipoatrophy, central adiposity) hyperlipidaemia and insulin resistance often complicate protease inhibitor containing antiretroviral therapy. Lipoatrophy and abdominal distension were observed in protease inhibitor-naive nucleoside analogue reverse transcriptase inhibitor (NRTI) recipients with lactic acidaemia and hepatic impairment, which are known NRTI-induced mitochondrial toxicities. DESIGN AND SETTING: Case-control study in a university-based outpatient clinic. PATIENTS AND METHODS: The patients studied included 14 NRTI recipients with lipoatrophy, 32 antiretroviral-naive patients without LD, 28 NRTI recipients without LD, 44 combined NRTI-protease inhibitor recipients without LD, and 102 NRTI-protease inhibitor recipients with LD. Data was obtained on body composition (questionnaire, physical examination, dual-energy x-ray absorptiometry and abdominal computerized tomography), with biochemical, lipid and glycaemic parameters. RESULTS: The NRTI-LD syndrome was characterized by recent onset fatigue and nausea, peripheral lipoatrophy (6 kg loss over 4 months), abdominal distension (ascites +/- hepatomegaly) and elevated lactate (4.6, 1.1, 1.2, 1.4 and 1.7 mmol/l, respectively; P< 0.0001) and liver enzymes. Cases without hepatic involvement also had lower body fat and greater lactate than unaffected controls. Metabolic disturbances and weight improved after cessation. The NRTI-LD syndrome differed from protease inhibitor-related LD syndrome by the presence of recent onset symptoms and weight loss, higher lactate and alanine aminotransferase, and lower albumin, cholesterol, triglycerides, glucose and insulin. In treated controls, current stavudine therapy, protease inhibitor duration, and lactic acidaemia were independently associated with both lipoatrophy and abdominal obesity; total NRTI duration was also associated with lipoatrophy, and lamivudine and protease inhibitor duration with buffalo hump. CONCLUSIONS: A syndrome of lipoatrophy, constitutional illness, lactic acidaemia and hepatic dysfunction can complicate NRTI therapy. Both protease inhibitor and NRTI therapies, particularly if associated with lactic acidaemia, contribute to LD syndrome, but have some distinguishable clinical and metabolic effects. PMID- 10716496 TI - HIV-1 strains from a cohort of American subjects reveal the presence of a V2 region extension unique to slow progressors and non-progressors. AB - OBJECTIVES: To determine the molecular nature of HIV-1 quasispecies and their evolution, in vivo over time, in an American cohort of 22 homosexual men [four rapid progressors (RP), 15 slow progressors (SP) and three long-term non progressors (LTNP)], infected with HIV-1 between 1982 and 1983, and to assess the possible role of the HIV-1 V2 region extension in HIV disease progression. DESIGN: Genetic and phylogenetic analyses of the V3 region and the nef gene clones over time from uncultured peripheral blood mononuclear cells (PBMC) of American patients with varying HIV disease progression rates. METHODS: Proviral DNA from longitudinally collected uncultured PBMC were subjected to PCR amplification in the nef gene and env V2 and V3 regions, followed by cloning, sequencing and phylogenetic analysis to establish evolutionary relationships between HIV-1 strains over time. RESULTS: Analysis of multiple viral clones showed nef gene deletions/insertions in 10 out of 15 SP, along with the coexistence of intact and defective nef gene lineages in the same individual over time, whereas these nefgene abnormalities were absent from HIV-1 strains from LTNP. Increasing quasispecies diversity in HIV-1 strains, over time, abrogation of a V3 region N-linked glycosylation site in > 60% of the clones, and, importantly, an extended V2 region were unique features of HIV-1 strains from SP and LTNP. CONCLUSIONS: The V2 region extension was unique to only SP and LTNP, and so may have a role in slow progression or non-progression of HIV disease. Increasing genetic diversity in HIV-1 strains in SP and LTNP correlated with the immunocompetent status of the host. PMID- 10716497 TI - Direct visualization of HIV-1-specific cytotoxic T lymphocytes during primary infection. AB - OBJECTIVE: HIV-specific cytotoxic T lymphocytes (CTL) are believed to play an important role in containing viral replication throughout HIV-1 infection. Previous studies have attempted to quantify the HIV-1-specific CTL precursor frequency during primary HIV infection by using limiting dilution analysis, which almost certainly underestimates the true CTL frequency. Here we use a relatively new technique to quantify HIV-specific CD8 T cells in primary HIV infection. METHODS: We have used soluble tetrameric complexes of HLA class I molecules complexed with HIV epitope peptides to study the dynamics and frequency of HIV specific CD8 T cells in relation to plasma viral load in early HIV infection, in three patients with a highly focused HIV-specific CTL response. RESULTS: We show that the frequencies of HIV-1-specific CD8 T cells in acute infection are significantly higher than previously documented and can be demonstrated well before full seroconversion. These studies also confirm the immunodominance of the B27-restricted response in HIV infection and demonstrate a close temporal relationship between the numbers of circulating HIV-specific CD8 T cells and viral load. CONCLUSIONS: These findings strongly suggest that HIV-1-specific CD8 T cells are responding directly to the level of viral replication in early HIV infection and are a major factor in its control. In addition, the data indicate that immunodominance for CD8 T-cell responses is established in the acute phase of HIV infection. PMID- 10716498 TI - Significance of P-glycoprotein for the pharmacology and clinical use of HIV protease inhibitors. PMID- 10716499 TI - Significance of P-glycoprotein for the pharmacology and clinical use of HIV protease inhibitors. PMID- 10716500 TI - Time of initiation of antiretroviral therapy: impact on HIV-1 viraemia. The Swiss HIV Cohort Study. AB - OBJECTIVE: The current recommendation that patients infected with HIV-1 be treated early is based on little evidence. We examined whether the early initiation of antiretroviral treatment affects residual HIV-1 viraemia. METHODS: Viraemia was measured using an assay with a detection limit of 3 HIV-1 RNA copies/ml in drug-naive patients who started antiretroviral therapy at the time of primary HIV-1 infection (PHI) (n = 10), during chronic infection without immune suppression (CD4 cell counts > or = 500/mm3; median 577) (n = 10), or after immune suppression developed (CD4 cell counts < 500/mm3; median 113) (n = 21). RESULTS: In 249 samples collected 24 to 120 weeks after treatment initiation, the mean proportion of samples with HIV-1 RNA levels of less than 3 copies/ml was 75% for PHI patients compared with 32 and 8% for immunocompetent and immunosuppressed chronically infected patients, respectively. Fifty per cent of PHI patients, but none of the chronically infected patients, had persistently fewer than 3 HIV-1 RNA copies/mL. PHI patients had lower residual HIV-1 RNA levels than chronically infected patients, and immunocompetent patients had lower residual HIV-1 RNA levels than immunosuppressed patients (all pairwise, P< 0.001). The mean residual HIV-1 RNA level was independently associated with the initiation of therapy during PHI and baseline CD4 cell counts (P < 0.001 for both associations). CONCLUSION: Viraemia levels are associated with clinical progression and predict virological treatment failure. The initiation of antiretroviral therapy at the time of PHI and while CD4 cell counts are high results in lower residual viraemia. These results support early antiretroviral therapy in HIV-1-infected patients. PMID- 10716501 TI - Early immune reconstitution after potent antiretroviral therapy in HIV-infected children correlates with the increase in thymus volume. AB - DESIGN: Despite significant rises in total CD4 T cells, the process of immune reconstitution in adults with HIV infection treated with potent antiretroviral treatment results in a rather slow increase in phenotypically naive lymphocytes. In children more than in adults, thymic function may be at least partly restored when disease-induced immunosuppression is attenuated by pharmacological means. METHODS: Twenty-five vertically infected and antiretroviral-experienced [zidovudine (ZDV)/ZDV plus didanosine (ddl)] children were prospectively followed during 12 months of treatment with lamivudine (3TC), stavudine (d4T) and indinavir (IDV). The plasma HIV viral load and phenotypic and functional cellular immunity-defining parameters were examined. The relationship between the degree of immune reconstitution and thymus volume assessed by nuclear magnetic resonance was also examined. RESULTS: An early and steep increase in CD45RA+62L+ T cells was observed in parallel with a sustained decrease in plasma HIV RNA levels and a significant rise in total CD4 T cells. This increase was significantly greater than that observed in CD4+CD45RO+ T cells. Analysis of the CD4 T cell receptor (TCR) beta repertoire and T helper function showed the ability to reconstitute families almost completely absent at baseline, and a substantial improvement of antigen-specific responses by peripheral blood lymphocytes. The rise in CD4 cells and in CD4+CD45RA+62L+ T cells was statistically associated with changes in thymus size observed over time. CONCLUSION: These data suggest a relevant contribution of the thymus to reconstitution of the peripheral pool of T cells in vertically HIV-infected children treated with potent antiretroviral regimens. PMID- 10716502 TI - HIV-1 genotypic zidovudine drug resistance and the risk of maternal--infant transmission in the women and infants transmission study. The Women and Infants Transmission Study Group. AB - OBJECTIVES: Although the treatment of pregnant women and their infants with zidovudine (ZDV) has been remarkably effective in preventing the perinatal transmission of human HIV-1, many potentially preventable infections still occur. To examine whether the risk of perinatal infection is increased among women who carry ZDV-resistant HIV-1, the role of genotypic ZDV resistance in perinatal transmission was evaluated. METHODS: The reverse transcriptase (RT) region of clinical isolates from culture supernatants of 142 HIV-1-infected women enrolled in the Women and Infants Transmission Study (WITS), who had been treated with ZDV during pregnancy was sequenced. Results from genotypic sequencing were linked to demographic, laboratory, and obstetrical databases, and the magnitude of association of having consensus drug-resistant HIV-1 RT mutations with transmission was estimated. RESULTS: Twenty-five per cent (34/142) of maternal isolates had at least one ZDV-associated resistance mutation. A lower CD4 cell percentage and count (P= 0.0001) and higher plasma HIV-1 RNA (P=0.006) were associated with having any ZDV resistance mutation at delivery. Having any RT resistance mutation [odds ratio (OR): 5.16; 95% confidence interval (CI): 1.40, 18.97; P=0 0.01], duration of ruptured membranes [OR: 1.13 (1.02, 1.26) per 4 h duration; P= 0.02], and total lymphocyte count [OR: 1.06 (1.01, 1.10) per 50 cells higher level; P=0.009] were independently associated with transmission in multivariate analysis. CONCLUSION: Maternal ZDV resistant virus was predictive of transmission, independent of viral load, in these mothers with moderately advanced HIV-1 disease, many of whom had been treated with ZDV before pregnancy. PMID- 10716503 TI - Incidence of neuropathy in HIV-infected patients on monotherapy versus those on combination therapy with didanosine, stavudine and hydroxyurea. AB - BACKGROUND: Sensory neuropathy is a common adverse effect of the nucleoside analogue anti-retroviral drugs didanosine (ddl) and stauvudine (d4T). These drugs are increasingly being used in combination, and it is not currently known whether the incidence of neuropathy is higher with combination compared to individual drug use. It is also not known if hydroxyurea, used to potentiate the antiviral efficacy of these drugs, may also increase the risk of neuropathy. The purpose of this analysis is to investigate if the combination of ddl and d4T, with or without hydroxyurea, has a higher incidence of neuropathy than a single drug regimen. METHODS: Data were obtained from patients followed longitudinally by the Johns Hopkins AIDS Services. Incidence rates of development of neuropathy were calculated for each of five regimens: ddl (+/- hydroxyurea), ddl + d4T (+/- hydroxyurea), and d4T. Cox proportional hazard regression was used to compare the relative risk of neuropathy for each regimen adjusting for CD4 cell count, other drugs received, and time on therapy. RESULTS: A total of 1116 patients received at least one of the five regimens. There were 117 cases of neuropathy. The crude incidence rate of neuropathy ranged from 6.8 cases per 100 person-years for ddl to 28.6 cases per 100 person-years for ddl + d4T + hydroxyurea. Compared with ddl alone, and adjusting for CD4 cell counts and other variables, the relative risk of neuropathy was 1.39 [95% confidence interval (CI): 0.84-2.32] for d4T alone, 2.35 (95% CI: 0.69-8.07) for ddl + hydroxyurea, 3.50 (95% CI: 1.81-6.77) for ddl + d4T, and 7.80 (95% CI: 3.92-15.5) for ddl + d4T + hydroxyurea. CONCLUSIONS: Based on the data, the risk of neuropathy is additive or even synergistic for ddl + d4T + hydroxyurea compared with ddl or d4T alone. The combination of ddl + d4T also increases the risk of neuropathy but less than when hydroxyurea is included. PMID- 10716504 TI - Geographic variation of HIV infection in childbearing women with syphilis in the United States. AB - OBJECTIVES: Substantial biologic and epidemiologic data indicate the importance of syphilis as a potential cofactor for sexual transmission of HIV infection, but few detailed data exist on the geographic covariation of these two important sexually transmitted infections. DESIGN: HIV prevalence in childbearing women and primary and secondary (P&S) syphilis data from 29 states were examined to explore the importance of the epidemiology of syphilis as a factor in facilitating HIV transmission. METHOD: The spatial relationship between P&S syphilis and HIV infection in the health districts of 29 states was analyzed and adjusted for demographic and socioeconomic factors such as racial composition, income, housing, education levels, and access to medical services using the 1990 US census, and geographic location. RESULTS: In 29 states and the District of Colombia, 448 health districts, representing more than 75% of the US population, reported HIV prevalence rates for mothers' district of residence. The HIV seroprevalence ranged from 0 to 1258/10 000 in these health districts. The incidence of P&S syphilis from 1984-1994 in these districts ranged from 0 to 87/100 000. The P&S syphilis incidence was positively associated with the prevalence of HIV infection among childbearing women (P < 0.0001). CONCLUSIONS: Syphilis that persists in communities in the United States appears to represent a 'sentinel public health event' reflecting risk for sexual HIV transmission. These findings, along with other biologic and epidemiologic information, reinforce the importance of syphilis as an indicator for targeting HIV prevention efforts generally, as well as syphilis control as a specific HIV-prevention strategy. PMID- 10716505 TI - HIV-1 diversity in France, 1996-1998. The AC 11 laboratory network. AB - OBJECTIVE: To study the distribution of HIV-1 subtypes in France and to describe the characteristics of patients infected with non-B subtypes. METHODS: All adults who tested HIV-1 positive on Western blot for the first time in one of the participating laboratories between September 1996 and March 1998 were eligible, whether or not they had been diagnosed previously elsewhere. Data on age, sex, country of birth, HIV-transmission group, dates of the last negative and first positive HIV test and clinical stage were collected. Serotyping was performed with a peptide subtype-specific enzyme immunoassay on each plasma sample and genotyping with heteroduplex mobility assay on each non-B serotype-infected patient. Patients characteristics were compared in B and non-B subtypes. RESULTS: Of the 2168 HIV-positive patients included by 32 laboratories, subtype,results were available for 2042. Among those, 73.4% were men, 12.2% born in sub-Saharan Africa, 41.5% infected through heterosexual contact and 67.6% in CDC stage A. Among the 2042 patients, 1 725 (84.5%) were infected with B subtype. Among the 317 non-B subtypes, subtype A was predominant (66.9%); all other subtypes (C, D, E, F, G, H, O) were present. Factors independently associated with a non-B subtype were to be included in the Paris area [adjusted odds ratio (aOR), 1.6; 95% confidence interval (CI), 1.1-2.3], to be born in sub-Saharan Africa (aOR, 26.0; 95% CI, 17.5-37.8) and to be infected through heterosexual contact (aOR, 4.2; 95% CI, 2.8-6.4). CONCLUSIONS: In France, although B subtype is still predominant, all non-B subtypes are now present. The diversity of HIV strains may affect diagnostic tests and clinical practice, especially viral load measurements. Moreover, the decreased susceptibility of non-B subtypes to antiretroviral drugs emphasizes the importance of surveillance of HIV diversity. PMID- 10716506 TI - Behavioral and biologic evidence of persistent high-risk behavior in an HIV primary care population. AB - OBJECTIVE: To define the prevalence of gonorrhea, chlamydial infection, and high risk sexual behavior in an HIV primary care clinic. DESIGN: Subjects enrolling in this cross-sectional study answered a brief interviewer-administered questionnaire and provided a urine sample for gonorrhea and chlamydia testing. SETTING: A large urban HIV primary care clinic. PARTICIPANTS: HIV-infected patients presenting for a scheduled medical visit from June 1997 to April 1998. MAIN OUTCOME MEASURES: Prevalence of self-reported high-risk sexual behavior and gonorrhea and chlamydial infection. RESULTS: Of 691 patients consenting to the study over a 10-month period, 58% reported sexual activity in the past 90 days, 7.4% reported multiple sexual partners in the past month, and 34.6% did not use a condom at last sexual encounter. Overall, 4.6% reported a history of either gonorrhea or a chlamydial infection in the past year. Of 637 giving a urine sample for testing, the prevalence of chlamydial infection was 2.4%; the prevalence of gonorrhea was 1.6%. Overall, 7.5% of those screened had either current or recent (within 1 year) gonorrhea or chlamydial infection. Current or recent gonorrhea or chlamydial infection was not associated with age, gender, HIV transmission risk, CD4 cell count, HIV viral load, symptoms, or self-reported risk behavior. CONCLUSION: High-risk sexual behavior and unrecognized sexually transmitted diseases (STD) are common among HIV-infected persons followed in primary medical care. Enhanced detection of treatable STD among this population coupled with improved risk-reduction counselling may be important clinical practice measures that can curb the spread of HIV. PMID- 10716507 TI - Comparison of sexual behaviors, unprotected sex, and substance use between two independent cohorts of gay and bisexual men. AB - OBJECTIVE: To compare demographic characteristics, sexual practices, unprotected receptive and insertive anal intercourse, substance use and rates of HIV-1 seroconversion between two prospective cohorts of HIV-negative men who have sex with men. DESIGN: Comparative analysis of two independent cohorts. METHODS: Between May 1995 and April 1996, 235 HIV-negative Vanguard Project (VP) participants were enrolled and between January and December 1985, 263 HIV negative participants in the Vancouver Lymphadenopathy AIDS Study (VLAS) completed a follow-up visit. The VP participants were compared with VLAS participants with respect to self-reported demographic variables, sexual behaviors, unprotected sex, substance use and rates of HIV-1 seroconversion during follow-up. RESULTS: In comparison with the VLAS participants the VP participants were younger (median age, 26 versus 34 years; P< 0.001), more likely to be non-Caucasian (75 versus 97%; P< 0.001), and were less likely to have attended university/college (35 versus 46%; P = 0.014). The VP participants reported a higher mean number of male sex partners in the previous year (15 versus 12; P= 0.026) and a higher mean number of regular partners (1.7 versus 0.6; P < 0.001). The VP participants were more likely to report engaging in receptive (92 versus 60%; P< 0.001) and insertive (90 versus 69%; P < 0.001) anal intercourse with regular partners and receptive anal intercourse with casual partners (62 versus 38%; P< 0.001). The VLAS participants were more likely to report never using condoms during insertive and receptive anal intercourse with both regular and casual partners. The VP participants were less likely to report using nitrite inhalants (34 versus 43%; P= 0.033), but more likely to report the use of cocaine (30 versus 8%; P< 0.001), LSD (21 versus 3%; P < 0.001), amphetamine (11 versus 1%; P< 0.001), heroin (3 versus 0%; P= 0.010) and methyldiamphetamine (17 versus 10%; P= 0.034). The VLAS participants were nine times more likely to report high-risk sexual behavior, after controlling for differences in age, ethnicity, substance use, and method of recruitment between cohort members. After adjustment for differences in demographics, sexual behaviors, and level of substance use, the risk ratio for seroconversion among VLAS participants remained significantly elevated compared with VP participants. CONCLUSION: These data provide evidence that men who have sex with men who were enrolled in the VP were more sexually active than their VLAS counterparts were 10 years ago as measured by self-reported numbers of regular and casual partners and frequency of anal intercourse with these partners. However, condom use appears to be significantly higher among VP participants, which has contributed to a lower rate of HIV-1 infection. PMID- 10716509 TI - Epidemiology of thrombosis in HIV-infected individuals. The Adult/Adolescent Spectrum of HIV Disease Project. AB - OBJECTIVE: To describe the incidence of clinically recognized thrombosis and associated factors among individuals infected with HIV. DESIGN: A longitudinal medical record review. SETTING: Over 100 medical clinics in nine US cities. PATIENTS: A total of 42 935 individuals aged 13 years or older with HIV infection, observed for an average of 2.4 years. MAIN OUTCOME MEASURES: The incidence of thrombosis among HIV-infected individuals; adjusted odds ratios for factors associated with thrombosis. RESULTS: The incidence of thrombosis among HIV-infected individuals was 2.6/1000 person-years (PY). Factors significantly associated with thrombosis included: age of 45 or more years (adjusted odds ratio [AOR], 1.9; 95% confidence interval [CI], 1.4-2.7); a diagnosis of cytomegalovirus disease or retinitis (AOR, 1.9; CI, 1.2-2.9), or other AIDS defining opportunistic illness (AOR, 1.5; CI, 1.1-2.2); hospitalization (AOR, 3.3; CI, 2.5-4.4); and the prescription of megestrol acetate (AOR, 2.0; CI 1.3 2.9) or indinavir (AOR, 2.4; CI 1.4-4.3). The prescription of other protease inhibitors, sex, race, and mode of HIV exposure were not associated with thrombosis. CONCLUSION: Among HIV-infected individuals, clinically detected thrombosis is more common in those who have opportunistic illnesses, for whom megestrol acetate or indinavir have been prescribed, who have been hospitalized, and who are aged 45 years or older. Physicians should be aware of the risks of thrombosis in order to promote the early identification and appropriate treatment or prophylaxis. Further study is needed to characterize the association between indinavir and thrombosis. PMID- 10716508 TI - The prevalence and incidence of HIV-1 infection and syphilis in a cohort of police officers in Dar es Salaam, Tanzania: a potential population for HIV vaccine trials. AB - OBJECTIVES: To assess the suitability of a cohort of police officers in Dar es Salaam for HIV vaccine trials by determining the prevalence and incidence of HIV 1 infection, active syphilis and their associated factors. DESIGN AND SETTING: An open cohort study of police officers in Dar es Salaam, Tanzania. METHODS: Recruitment of police officers began in 1994. A standardized questionnaire was completed at enrolment and subsequent visits. HIV antibodies were determined using two consecutive enzyme-linked immunosorbent assays. Samples repeatedly discordant on the two tests were tested by a Western blot assay. Treponema pallidum antibodies were first determined by Venereal Disease Research Laboratory (VDRL) test and reactive sera were confirmed by Treponema pallidum hemagglutination test. RESULTS: At the end of 1996 a total of 2850 police officers had been recruited of whom 2733 (96%) consented to be tested for HIV. The overall HIV-1 seroprevalence at recruitment was 13.8% (378 of 2733). Females had a significantly higher HIV-1 seroprevalence, 18.0% (55 of 306), as compared to males, 13.3% (323 of 2427), P< 0.05. From a total of 2215 married police officers, 585 (26.4%) responded to a question on extramarital sex within the previous 3 months of whom 36.2% (212 of 585) admitted to have had at least one extramarital sexual intercourse. Condoms were not used during these encounters by 178 of 212 (84.0%). As of 31st December 1998, among the 1524 males observed for 2553 person-years (PYAR), 50 had seroconverted and among 200 females observed for 357 PYAR, eight had seroconverted. The overall crude HIV-1 incidence was thus 19.9/1000 PYAR; 19.6 and 22.4/1000 PYAR for males and females, respectively. The overall prevalence and incidence of active syphilis were 3.1% (88 of 2850) and 8.6/1000 PYAR (26 of 3149), respectively. Males had a higher prevalence of active syphilis, 84 of 2525 (3.3%) than females, five of 325 (1.5%), P = 0.09. CONCLUSIONS: There was high risk sexual practice including low condom use in this cohort of police officers. The incidence and prevalence of HIV infection were high. Police officers in Dar es Salaam are therefore a potential population group for HIV vaccine evaluation. PMID- 10716510 TI - Is post-exposure prophylaxis affordable? PMID- 10716511 TI - A reality check: the cost of making post-exposure prophylaxis available to gay and bisexual men at high sexual risk. PMID- 10716512 TI - Diversity of HIV-1 genetic subtypes in France, in the context of mother-to-child transmission. The French Pediatric Cohort Study Group. PMID- 10716513 TI - HIV virions and HIV replication are unaffected by granulysin. PMID- 10716514 TI - Normalization of the CD4 T cell receptor repertoire after evolution of syncytium inducing HIV-1 variants. PMID- 10716515 TI - Efavirenz associated with corticosteroids in patients with previous severe hypersensitivity reaction due to nevirapine. PMID- 10716516 TI - Excess peripheral neuropathy in patients treated with hydroxyurea plus didanosine and stavudine for HIV infection. PMID- 10716517 TI - Relationship between Kaposi's sarcoma, Kaposi's sarcoma-associated herpesvirus and AIDS dementia complex. PMID- 10716518 TI - Rash as side-effect of nelfinavir in children. PMID- 10716519 TI - Carpal tunnel syndrome in HIV-1 patients: a metabolic consequence of protease inhibitor use? PMID- 10716520 TI - Effect of beta-blocker therapy on severe ventricular arrhythmias in patients with idiopathic dilated cardiomyopathy. AB - Beta-blocker therapy has been shown to improve cardiac function and prognosis in patients with idiopathic dilated cardiomyopathy (DCM). However, whether beta blockers reduce severe ventricular arrhythmias and sudden cardiac death has not been clarified. The present study was designed to investigate the effects of beta blockers on non-sustained ventricular tachycardia (VT) and sudden cardiac death in patients with DCM. Sixty-five patients with DCM treated with diuretics, digitalis and angiotensin-converting enzyme inhibitors were assigned to receive beta-blockers (n = 33) or not (n = 32). Mean follow-up was 53+/-30 months. The echocardiographic indices of cardiac function, the incidence of non-sustained VT on Holter monitoring electrocardiograms, and sudden cardiac death rate were compared between the 2 groups. Comparable improvement in cardiac function on echocardiograms was found in the 2 treatment groups. The patient group treated with beta-blockers showed a significant reduction in the prevalence of VT (from 43 to 15%, p<0.05) and the development of new episodes of VT (5 vs. 16%) compared to the group without beta-blockers. The sudden cardiac death rate did not differ between the 2 groups. The results of the present study suggest that beta-blockers are effective in reducing severe ventricular arrhythmias in patients with DCM. PMID- 10716521 TI - Lipoprotein(a), left atrial appendage function and thromboembolic risk in patients with chronic nonvalvular atrial fibrillation. AB - Lipoprotein(a) (Lp(a)) has a prothrombotic effect by modulating the fibrinolytic system. The purpose of the present study was to determine whether serum Lp(a) levels are associated with an increased risk of thromboembolism in chronic nonvalvular atrial fibrillation (NVAF). Clinical, laboratory and transesophageal echocardiographic data were collected in 172 consecutive, non-anticoagulated patients with chronic NVAF. Thirty-four patients (thromboembolic group) had a recent (<1 month) embolic event and/or a left atrial thrombus on transesophageal echocardiography. The thromboembolic group had a higher frequency of spontaneous echo contrast (94 vs. 58%, p<0.0001), increased concentrations of Lp(a) (median: 31.5 vs. 15.5 mg/dl, p<0.0001) and fibrinogen (median: 352 vs. 314 mg/dl, p = 0.0015), larger left atrial dimensions (median: 5.1 vs. 4.8cm, p = 0.0078), and reduced left atrial appendage (LAA) flow velocities (median: 9.5 vs. 21.2 cm/s, p<0.0001) than the nonthromboembolic group. Multivariate analysis identified 3 independent predictors of thromboembolism: Lp(a) level > or =30 mg/dl (odds ratio (OR) 9.5, 95% confidence interval (CI) 4.4-20.4, p<0.0001), LAA flow velocity of <20 cm/s (OR 8.7, 95% CI 3.3-23.0, p = 0.0003) and a fibrinogen concentration of <377mg/dl (OR 3.2, 95% CI 1.5-6.9, p = 0.0201). The Lp(a) elevations and reduced LAA flow velocities are independently associated with thromboembolism in chronic NVAF. PMID- 10716522 TI - Covered stent implantation by the puncture method for the treatment of a small aneurysm of the common iliac artery. AB - A method to repair endovascular aneurysms with covered stents has recently been developed. In the present paper, the implantation of a covered stent through a 12Fr sheath by the puncture method for the treatment of an isolated aneurysm of the right common iliac artery is reported. The aneurysm was less than 3 cm in diameter, and computed tomography showed no signs of aneurysm rupture, but the patient nonetheless complained of right lower abdominal pain and constipation. It was decided to implant a covered stent in lieu of surgical repair because it was difficult to prove a causal relationship between the aneurysm and the patient's complaints. Fortunately, after implantation, the symptoms were resolved. In conclusion, it is possible to choose this less invasive type of therapy for the treatment of an isolated iliac artery aneurysm if the patient complains only of general malaise and there are no certain signs of an impending rupture, although surgery should be indicated regardless of aneurysm size. PMID- 10716523 TI - Risk factors for non-fatal acute myocardial infarction in middle-aged and older Japanese. Fukuoka Heart Study Group. AB - It remains uncertain whether established risk factors for coronary heart disease in middle-aged persons can be generalized to elderly persons. Based on a case control study, risk factors for nonfatal acute myocardial infarction (AMI) were assessed separately in middle-aged (40-64 years) and older (65-79 years) Japanese. Eligible cases were patients who were admitted to 22 collaborating hospitals for the first AMI between September 1996 and January 1998. Community controls were recruited by using the resident registers of the municipalities with individual matching by gender, year of birth (within 2 years), and proximity in residence. The present study used 384 sets of 384 cases and 656 controls. Smoking, hypertension, and angina pectoris were associated with an increased risk of AMI, and alcohol use and leisure-time exercise were related to a decreased risk of AMI in the elderly as well as in middle-aged persons. There was no apparent relation between body mass index and AMI in either middle-aged or older adults. Diabetes mellitus was significantly associated with an increased risk of AMI in older persons, but not in middle-aged persons. Hypercholesterolemia was related to an increased risk of AMI in middle-aged individuals alone. The findings suggest that risk factors for AMI in the elderly are generally similar to those of middle-aged persons, but provide no evidence that hypercholesterolemia in the elderly is an important risk factor. PMID- 10716524 TI - Second derivative of photoplethysmogram in children and young people. AB - The characteristics of the second derivative of the photoplethysmogram (SDPTG) were clarified in children and young people, and the factors affecting the SDPTG wave pattern were examined. The study group comprised 775 healthy subjects aged 3 20 years (mean, 10+/-5). The blood pressure of the left brachial artery was determined in the resting sitting position and then the fingertip PTG and the SDPTG were automatically measured using a digital photoplethysmograph, with the sensor located at the cuticle of the second digit of the right hand. The values used were the b/a, c/a, d/a, and e/a ratios, and the SDPTG aging index (SDPTG AI). With increasing age, the systolic blood pressure and height increased (r = 0.52, 0.92). Aging decreased the b/a ratio and SDPTG-AI (r = -0.58, -0.67) and increased the c/a and e/a ratios (r = 0.42 and 0.42). There was no significant correlation between blood pressure and indices of SDPTG. As height increased, the b/a ratio and SDPTG-AI decreased (r = -0.57, -0.71), whereas the c/a and e/a ratios increased (r = 0.42 and 0.46). In males the SDPTG-AI decreased with age from 3 to 18 years and then increased, and in females it decreased with age from 3 to 15 years and then increased. Overall, the SDPTG-AI decreased with age between 3 and 18 years and then increased, forming a J curve. In the children's and young people's SDPTG, the b/a and SDPTG-AI decreased and the c/a and e/a ratios increased with age. The length of the vascular system and the inner diameter and wall thickness of vessels may modify the SDPTG wave pattern in the growth period. Thereafter, as the effects of these factors decrease, the increase in intravascular pressure and decreasing wall elasticity due to aging may affect the wave pattern. PMID- 10716525 TI - Improvement in fatty acid utilization in relation to a change in left ventricular hypertrophy in spontaneously hypertensive rats. AB - Although fatty acid metabolism is reportedly impaired in myocardial hypertrophy, it is unclear whether the antihypertensive drugs are associated with improved fatty acid metabolism. In order to evaluate the effects of antihypertensive drugs on fatty acid metabolism and myocardial perfusion, the simultaneous uptake of iodine-125(125I)-15-(p-iodophenyl)-3-(R,S)-methylpentadecanoic acid (BMIPP) and thallium-201 (Tl) were measured in 3 groups of rats: (1) spontaneously hypertensive rats (SHR) without treatment (SHR-N), (2) SHR chronically treated with captopril (SHR-C), and (3) SHR chronically treated with hydralazine (SHR-H). Captopril and hydralazine were administered to their respective groups for 3 weeks from 12 weeks of age. The hearts were removed 10 min after simultaneous intravenous injections of BMIPP and Tl and the 125I and 201Tl counts were measured to calculate the uptake ratio. The systolic blood pressure (SBP) in SHR N was 222+/-10 mm Hg, whereas the SHR-C and SHR-H groups showed significant SBP reduction (156+/-11, and 158+/-10 mm Hg, respectively) (p<0.01 each). The heart/bodyweight ratio was significantly lower in SHR-C (2.48+/-0.09) than in SHR N (2.74+/-0.11) (p<0.05). However, there was no significant difference in the heart/bodyweight ratio between SHR-N and SHR-H (2.65+/-0.09). The ratio of BMIPP uptake to Tl uptake (BMIPP/Tl) was significantly higher in SHR-C (0.71+/-0.13) than in SHR-N (0.50+/-0.09) (p<0.05). However, BMIPP/Tl in SHR-H (0.53+/-0.09) was similar to that in SHR-N. These results suggest that captopril improves fatty acid metabolism in the hypertrophied ventricle in SHR. The metabolic alterations may improve with left ventricular hypertrophy regression but are not effected by the reduction of blood pressure only. PMID- 10716526 TI - Spatial heterogeneity in refractoriness as a proarrhythmic substrate: theoretical evaluation by numerical simulation. AB - Spatial heterogeneity in the refractoriness of the ventricular myocardium due to a regionally prolonged refractory period has often been observed in patients with cardiovascular disease as the substrate for functional reentrant tachyarrhythmias. The present study sought to determine how functional reentrant activity could occur due to the spatial heterogeneity, using numerical simulation. Spatial heterogeneity in the refractoriness was introduced into a two dimensional array by the regionally prolonged refractory period expressed as a square cluster. Double stimulation, conducted from a single source, was introduced into 4 types of matrices, which differed in their level of spatial heterogeneity. A pseudoelectrocardiogram was calculated from these matrices. Spiral waves were initiated in all the matrices except for the lowest heterogeneous matrix. A vulnerable window of the coupling interval, which induced spiral waves, was observed and was wider in proportion to the level of the heterogeneity. A higher level of heterogeneity and more limited range of coupling intervals were required to sustain the spiral waves. Furthermore, in the pseudoelectrocardiogram, sustained spiral waves exhibited a waveform like that in torsades de pointes (TdP) and their transformation into ventricular fibrillation (VF). Spatial heterogeneity in refractoriness due to a regionally prolonged refractory period could be a substrate for functional reentrant tachyarrhythmias, possibly including TdP and VF. PMID- 10716527 TI - A case of aortic dissection with transient ST-segment elevation due to functional left main coronary artery obstruction. AB - A 48-year-old man with a history of hypertension and diabetes mellitus was hospitalized with sudden onset of severe chest pain. He was in cardiogenic shock with a systolic pressure of 60 mm Hg. His electrocardiogram (ECG) showed ST segment elevation in the precordial leads suggestive of acute anteroseptal myocardial infarction. The ST-segment returned to baseline after the systolic blood pressure rose to 100 mm Hg with the administration of sympathomimetic agents. Aortography and transesophageal echocardiography demonstrated type A aortic dissection and aortic regurgitation. Aortography and short-axis transesophageal echocardiography showed during diastole almost complete collapse of the true lumen of the ascending aorta caused by the intimal flap. The patient underwent surgical repair of the aortic dissection and implantation of Palmaz stents in the carotid arteries. Decreased blood pressure and the presence of aortic regurgitation accelerated the collapse of the true lumen during diastole in the ascending aorta, resulting in functional obstruction of the left main coronary artery, which may have been related to ST-segment changes in this case. PMID- 10716528 TI - A case of primary cardiac B cell lymphoma associated with ventricular tachycardia, successfully treated with systemic chemotherapy and radiotherapy: a long-term survival case. AB - We experienced a long-term survival case of primary cardiac lymphoma (PCL) demonstrating ventricular tachycardia (VT) as an initial sign, which was related to localized myocardial damage by lymphoma cells. A 70-year-old woman with sustained VT was admitted to the Kofu Municipal Hospital. VT ceased with the administration of disopyramide intravenously. The origin of the VT was the free wall of the right ventricular outflow tract (RVOT) as observed by electrocardiography on admission. A solitary mass in the free wall of the RVOT was found by echocardiography, chest computed tomographic scanning and magnetic resonance imaging. There was no evidence of extracardiac involvement. The patient was histologically diagnosed as PCL by endomyocardial biopsy. Chemotherapy started immediately after the diagnosis and the mass showed a marked reduction in size. After 8 cycles of chemotherapy, radiotherapy was performed. Pericardial thickness in the free wall of the RVOT developed without severe side effects. Complete remission has been maintained for 30 months after the initial diagnosis, and no recurrence and arrhythmias have been detected during the follow-up period. It was demonstrated that rapid diagnosis and chemotherapy followed by radiotherapy for PCL achieved better survival. PMID- 10716529 TI - Resuscitation from fulminant myocarditis associated with refractory ventricular fibrillation. AB - Resuscitation was possible in a case of fulminant myocarditis with refractory ventricular fibrillation (Vf) using a percutaneous cardiopulmonary support system (PCPS). A 46-year old Japanese man suddenly experienced cardiopulmonary dysfunction shortly after the onset of flu symptoms, was promptly diagnosed as having fulminant myocarditis and PCPS was immediately initiated. On the second day in the hospital, refractory Vf occurred, which lasted for approximately 2h despite repeated efforts to terminate it. Finally, a large dose of steroids was administered. From the third day of hospitalization and onwards, the Vf disappeared totally. The patient completely recovered from such a serious state in 6 months. During the following 3 years, he has had no clinical symptoms of worsening. As in this case demonstrates, most myocarditis is curable and invasive measures are very helpful in rescuing patients from the fulminant type with refractory Vf. PMID- 10716530 TI - Echocardiographic observation of acute myocarditis with systemic lupus erythematosus. AB - Although myocarditis from a series of autopsies of patients with systemic lupus erythematosus was frequently observed, the incidence of clinically apparent myocardial dysfunction was low. A 30-year-old woman with systemic lupus erythematosus was examined by echocardiography. An acoustic densitometry was followed at the left ventricular posterior wall throughout the clinical course. A decrease in the magnitude of cyclic variation of integrated backscatter (IB) was observed before treatment. Following the combined treatment, steroid and cyclophosphamide, a repeated ultrasonic tissue characterization showed an increase in the magnitude of cyclic variation of IB. It is thought that ultrasonic tissue characterization may be a useful method to evaluate the impairment of contraction, and to follow up the clinical course of myocardial involvement in systemic lupus erythematosus. PMID- 10716531 TI - Demonstration of phase-3 and phase-4 retrograde block in a second concealed accessory pathway after an initial successful radiofrequency ablation of a 'normal' concealed accessory pathway. AB - We report a patient with concealed Wolff-Parkinson-White syndrome who, following catheter ablation, demonstrated phase-3 and phase-4 retrograde block in a concealed accessory pathway. After an initial 'apparently successful' ablation, retrograde conduction was through the atrioventricular node during constant ventricular pacing. Ventricular extrastimulus testing was performed at a basic drive cycle length of 600 ms. Unexpectedly, ventricular extrastimuli at coupling intervals of 440-380 ms were conducted retrogradely over an accessory pathway, consistent with a phase-3 and phase-4 retrograde block in the accessory pathway. Residual accessory pathway conduction was eliminated in a single ablation session. PMID- 10716532 TI - Shortening of conduction time over arborized atrioventricular accessory pathway with Mahaim fibers physiology just before interruption during radiofrequency ablation. AB - A 21-year-old woman had paroxysmal wide QRS tachycardia with a left bundle branch block configuration and a retrograde conducted P wave just behind the QRS complex. An electrophysiological study revealed antidromic atrioventricular tachycardia involving an atrioventricular connection with decremental conduction as the anterograde limb and normal atrioventricular node as the retrograde limb. During constant pacing from the high right atrium (HRA) at the cycle length (CL) of 600 ms, the QRS configurations were not identical to those during the wide QRS tachycardia or constant pacing at the CL of less than 500 ms. The process by which this arborized atrioventricular accessory pathway with the Mahaim fibers physiology was interrupted by radiofrequency catheter ablation is described. Radiofrequency energy was delivered to the site recording a Mahaim potential at the tricuspid annulus during constant pacing from the HRA at the CL of 429 ms. The stimulus-QRS interval gradually shortened as it reached the power plateau without changing the preexcited QRS configuration. Shortening of the conduction time over the Mahiam pathway might have resulted in changing of the propagation from a slow to fast conduction zone or acceleration in response to thermal effect in a node-like structure on the atrial insertion site. PMID- 10716533 TI - Ampulla cardiomyopathy ('Takotusbo' cardiomyopathy)--reversible left ventricular dysfunction: with ST segment elevation. AB - The clinicopathologic findings of reversible ampulla-like ventriculogram of the left ventricle were studied in 8 elderly women and one middle-aged man. Their coronary arteriograms were normal, even in the 7 patients who had ST elevation on electrocardiogram. Coronary spasm was positive in only 2 of the 7 patients who received provocation tests. Biopsy specimens revealed focal myocyte injury. Normal coronary arteriograms during ST elevation and the presence of pathologic myocardial lesions were not consistent with a concept of stunned myocardium. The presence of myocardial lesions suggested that focal and disseminated myocardial damage had occurred. PMID- 10716534 TI - Effects of hypothermia and gender on survival and behavior after perinatal asphyxia in rats. AB - Previous studies in rats have demonstrated that perinatal asphyxia (PA) produces long-term morphological alterations, particularly affecting hippocampus. neostriatum, and cerebral cortex. These changes were prevented by applying hypothermia during the asphyctic insult. Because these cerebral areas are involved in cognitive and motor functions, the aim of the present study was to determine whether periods of PA during normothermia or hypothermia produces long term behavioral impairments in rats of both sexes. The cognitive and motor functions were studied using the spatial Morris water maze (MWM) task at 1.5 months, and the open field at 5 months, respectively. The present study revealed that female rats had a higher survival rate than males after PA in normothermic conditions (p < 0.014). and that hypothermia drastically prolonged the time of survival in both sexes (p < 0.001). There were no differences in learning and memory functions between groups or male and female rats when tested with MWM. Rats subjected to hypothermia treatment did not show differences in the MWM compared to controls. A lower locomotor activity in the open field test was only observed in male rats that suffered 15 and 20 min of PA in normothermia (p < 0.05). Hypothermia treatment prevented this hypoactivity. PA in females, even if severe, did not affect the motor activity. The data of both behavioral tests showed differences between sexes, i.e., the female rats learned the MWM task slower, and were more active in the open field. This work lends further support for the hypothesis that hypothermia can prevent mortality as well as long-term sequelae induced by PA. PMID- 10716535 TI - Palatability and intake relationships in free-living humans: measurement and characterization in the French. AB - To investigate palatability influences on the ad lib eating behavior of free living humans, 54 French participants were paid to maintain food intake diaries for four 7-day periods. They recorded their intake along with palatability ratings, on a seven-point scale, of each individual item eaten and also a global rating of the palatability of the entire meal. Higher levels of palatability were found to be related to larger meal sizes, durations, and deprivation ratios, smaller satiety ratios, greater hunger, and lower depression and anxiety. The global palatability rating was found to be superior to individual item palatability ratings as a measure of the palatability of the meal. Although palatability was found to have fairly large effects on intake, it accounted for less than 2% of the variance. It was concluded that, in the natural environment, there are a large number of other powerful variables present that add variance. In addition, people tend to self-select only a restricted range of highly palatable foods. As a result, in the natural environment, the influence of palatability on intake is limited. PMID- 10716536 TI - Lateral septal neuronal firing rate increases during proestrus-estrus in the rat. AB - Neuronal activity of the lateral septal nucleus (LSN) is related to motivational and hedonic behavior. Even though some changes in mood and anxiety during proestrus and pregnancy have been reported, the possible changes in the neuronal activity of the LSN through the phases of the estrous cycle are unknown. In the present study we explored the neuronal activity from the LSN using glass micropipettes (NaCl 1 M, and Evans blue 2.5%; 3-8 Mohms in 30 urethane (1 g/kg) anesthetized Wistar rats. Analysis of data included a total of 88 single-unit extracellular recordings taken from the LSN during proestrus (n = 22), estrus (n = 23), diestrus (n = 22), and metestrus (n = 21). The highest values of firing rate were found in proestrus, and the lowest in metestrus, F(3,84) = 3.78, p < 0.01. During estrous cycles, in the phase characterized by high plasma levels of estradiol and progesterone, i.e., proestrus-estrus, the neurons from the dorsal aspect of the LSN fired at significantly (p < 0.05) higher frequencies, shorter first-order intervals and a lower coefficient of variation than those in the phase characterized by lower levels of estradiol and progesterone (metestrus diestrus). In another group of rats (n = 12), immobility in the forced-swim test was assessed. Consistently, a longer latency (p < 0.05) for the first period of immobility and a nonsignificant trend to a lowered total time in immobility were found in proestrus and estrus. It is concluded that the higher firing rate in neurons from the dorsal aspect of the LSN during proestrus-estrus, may be associated with an increased motivation to escape from a stressful situation. PMID- 10716537 TI - Novel male mice disrupt pregnancy despite removal of vesicular-coagulating and preputial glands. AB - Exposure to novel males can disrupt intrauterine implantation of fertilized ova in inseminated female mice, an effect established to involve androgen-dependent male excretions. These experiments were designed to examine potential roles of vesicular and coagulating glands, independently or in conjunction with preputial glands. Inseminated females were randomly assigned to conditions of housing below (1) no males; (2) males with vesicular-coagulating gland removal; (3) males with preputial, vesicular, and coagulating gland removal; or (4) males subjected to sham surgery. Males with accessory glands removed disrupted pregnancy to the same extent as did sham-operated males. Long-term testosterone replacement permitted pregnancy disruption in castrated males with vesicular-coagulating and preputial glands removed. Fertility was not disrupted by preputialectomy, but half of the males without vesicular-coagulating glands could not inseminate females. We suggest that males' capacity to disrupt pregnancy could derive from androgen metabolism that is independent of actions on major accessory glands. PMID- 10716538 TI - Involvement of central beta-adrenoceptors in the tachycardia induced by water immersion stress in rats. AB - The purpose of the present study was to investigate whether central beta adrenoceptors are involved in stress-induced cardiovascular responses in rats. Using a biotelemetry system, blood pressure and heart rate were measured at rest and during stress induced by immersion in 1 cm-deep water. Intracerebroventricular (i.c.v.) injections of a nonselective beta-adrenoceptor antagonist, DL-propranolol (5 or 50 microg), significantly and dose dependently attenuated the tachycardia induced by water immersion stress (drug-induced reduction of tachycardia at 5 min after the start of stress: 61.4 +/- 13.2% for 5 microg, 72.5 +/- 8.2% for 50 microg). The same doses of DL-propranolol had no effect on the resting heart rate. Injection (i.c.v.) of a lower dose (5 microg) of D-propranolol--which has a lower potency as a beta-adrenoceptor antagonist than DL-propranolol, but a similar local anesthetic, membrane-stabilizing activity--did not attenuate the stress-induced tachycardia, although a higher dose (50 microg) did. Intravenous administration of DL-propranolol (5 or 50 microg) significantly attenuated the stress-induced tachycardia (drug-induced reduction of tachycardia at 5 min after the start of stress: 20.0 +/- 7.5% for 5 microg, 42.4 +/- 3.4% for 50 microg). However, the attenuation was much smaller than in the i.c.v. DL-propranolol-injected group. The i.c.v. injection of the 50 microg dose of DL-propranolol significantly augmented both the resting blood pressure and the pressor response to water immersion stress, whereas the lower dose (5 microg) had no effect. The i.c.v. injection of 50 microg D-propranolol also augmented, although not significantly, the resting blood pressure and the pressor response to stress. These results suggest that central beta-adrenoceptors are involved in the tachycardia induced by water immersion stress in rats. PMID- 10716539 TI - Modalities of the food intake-reducing effect of sibutramine in humans. AB - Sibutramine is a serotonin and noradrenaline reuptake inhibitor exerting a weight reducing effect partly via its anorectic properties. We investigated the effects of 15 mg sibutramine on objective (intake) and subjective (sensations) parameters of eating behavior in 24 young male subjects. At 0830 h subjects took either placebo or sibutramine in a counterbalanced order, followed by a fixed amount of breakfast. Intake was covertly recorded in the laboratory until the dinner meal, and then until the next morning using diary reports. Sibutramine induced a highly significant reduction in energy (1304 kJ, p < 0.001), protein (294 kJ, p < 0.001), fat (414 kJ, p < 0.01), and carbohydrate (CHO, 594 kJ, p < 0.001) intakes compared to placebo. This reduction was further enhanced when 24-h intake was analyzed (1601 kJ, p < 0.001). The effect of sibutramine occurred mainly at lunch (637 kJ, p = 0.005). Throughout the test day the number of items consumed and the weight of food were reduced by sibutramine (1.6, p < 0.01 and 222 g, p < 0.001, respectively), whereas energy density was not changed. Meals minus dessert items were the most altered by sibutramine. A specific CHO reduction was found in the dinner meal, although the proportions of macronutrients in total daily energy intake were not changed by sibutramine. Hunger ratings began to be lower than placebo 240 min after sibutramine. These results show that a single dose of sibutramine in lean humans induces a potent reduction in intake, and that its action is modulated according to the time of occurrence and the structure of the meal. PMID- 10716540 TI - Social stability attenuates the stress in the modified multiple platform method for paradoxical sleep deprivation in the rat. AB - The instrumental methods to induce paradoxical sleep (PS) deprivation are stressful. The modified multiple platform method (MMPM), in which animals are placed with new cohorts inside the water tanks, results in augmented ACTH and corticosterone (CORT) responses. We hypothesised that this increased response could be attributed to social instability. In addition, we tested a new environmental control, a grid (GR) placed on the tank floor. Animals were submitted to the MMPM for 4 days as socially unstable (UG--coming from several cages) or stable groups (SG--coming from one cage), placed either on narrow platforms or on the grid. All UG animals presented higher ACTH levels than their SG counterparts, including home-cage controls. CORT levels of manipulated animals were higher than controls only in the stable group. UG animals presented heavier adrenals than their SG counterparts. Only adrenals from SG animals placed on the grid were similar to cage controls. SG rats lost less weight than UG animals. While UG animals ate the same amount of chow as home-cage controls, SG animals ate more than control and UG animals. These results suggest that the stress of the MMPM can be attenuated in stable groups. The introduction of a grid on the tank floor may serve an adequate environmental control as far as stress-related variables are considered. PMID- 10716541 TI - Cell-body lesions of the posterodorsal preoptic nucleus or posterodorsal medial amygdala, but not the parvicellular subparafascicular thalamus, disrupt mating in male gerbils. AB - In gerbils, the posterodorsal preoptic nucleus (PdPN) and the lateral part of the posterodorsal medial amygdala (MeApd) express Fos with ejaculation. In contrast, the medial/central part of the MeApd expresses Fos when a sexually experienced male reenters the environment associated with mating. The parvicellular part of the subparafascicular thalamic nucleus (SPFp) of gerbils expresses Fos under both conditions. To study the role of the PdPN and MeApd in male sex behavior, male gerbils were tested for mating before and after these areas were bilaterally lesioned by infusions of N-methyl-D-aspartate (NMDA). Controls received the vehicle or inactive isomer, NMLA. Lesions in either area reduced mounting, but MeApd lesions, which were more complete than PdPN lesions, also delayed ejaculation when males intromitted. To determine if the MeApd and PdPN affect mating via a common pathway, they were bilaterally disconnected by lesioning them unilaterally, contralateral to each other. Other groups received ipsilateral lesions, NMLA, or bilateral lesions of the PdPN or MeApd. In addition, the SPFp was studied using bilateral lesions. MeApd and PdPN lesions again decreased mounting, and this time both lesions, which were quite complete, delayed ejaculation when males intromitted. Contralateral lesions that bilaterally disconnected these cell groups from each other mimicked both effects. Thus, the MeApd and PdPN affect mounting and ejaculation, at least in part, via their connections with each other. In contrast, SPFp lesions did not affect mating. Thus, SPFp cells activated at ejaculation may react to ejaculation rather than trigger it, possibly initiating preparations for paternity. PMID- 10716542 TI - Maturation of taste buds on the soft palate of the postnatal rat. AB - Taste bud distribution on the soft palate and within three types of tongue papillae (fungiform, foliate, and circumvallate) were examined histologically in the rat at different postnatal ages. After paraffin embedding, serial sections (10 microm) were made and stained by HE, and digitized images of each section were examined. The existence of a taste pore was used to identify mature taste buds. At birth, 53% (68 of 127 observed) of the taste buds on the soft palate, but only 14% (14 of 110 observed) within fungiform papillae, contained a taste pore. One week after birth, the number of mature taste buds increased rapidly, resulting in 90% of soft palate taste buds and 80% of fungiform taste buds containing taste pores. In contrast, no taste buds with pores were observed at birth within foliate and circumvallate papillae; however, at two weeks after birth 52% (71 of 132 observed) of the foliate and 68% (180 of 267 observed) of the circumvallate taste buds examined contained taste pores. These results suggest that taste buds within the soft palate play an important role in the detection of nutrients in the neonatal rat. PMID- 10716543 TI - Dehydroepiandrosterone-mediated decrease in caloric intake by obese Zucker rats is not due to changes in serum entrostatin-like immunoreactivity. AB - To understand the mechanism(s) of appetite modulation by DHEA, we have undertaken a series of studies to examine the effects of DHEA on neurotransmitters and neuropeptides known to affect appetitive behavior. Here, we report the effect of DHEA on serum enterostatin-VPDPR or E, a pentapeptide known to cause selective diminution in fat intake. Four-week-old lean (fa/+) and obese (fa/fa) Zucker rats were divided into control and treatment groups. DHEA-treated groups received powdered chow containing 0.6% DHEA ad lib for 16 weeks. Another group of obese rats was pair fed to match the intake of the obese DHEA-treated rats. At the end of this period, trunk blood was collected from fasted rats for assay of E-like immunoreactivity (E-LI) by ELISA. DHEA treatment caused a significant diminution in circulating E-LI in both lean (control: 2030 +/- 226; treated: 752 +/- 145 ng/mL; n = 10, p < 0.0001) and obese (control: 2489 +/- 391, n = 6; treated: 1123 +/- 185 ng/mL, n = 7; p = 0.0003) rats. Because DHEA treatment decreases caloric intake and body weight, we examined the effect of caloric intake and body weight on E-LI levels. Serum ELI levels were lower in the obese DHEA-treated group compared to that of obese pair fed (pair fed: 1589 +/- 313, n = 6; DHEA: 1123 +/- 185 ng/mL, n = 7), but the differences were statistically insignificant (p = 0.185). Also, both weight-matched lean and obese control rats had significantly (p < 0.008) higher E-LI than their DHEA-treated counterparts. To examine whether the decrease in serum E-LI following DHEA treatment could be due to increased peptide metabolism, the rate of disappearance of endogenous E-LI from serum (obese control and DHEA-treated) at 37 degrees C was evaluated. The results show an attenuation of peptide metabolism in serum from DHEA-treated rats, a finding contrary to our expectations. In summary, DHEA treatment lowers serum E-LI levels both in lean and obese Zucker rats. This decrement in peptide level is not secondary to changes in body weight or caloric intake due to DHEA, or due to altered serum peptide metabolism. Although DHEA appears to be a potent modulator of E-LI levels, the relationship between DHEA and E-LI in relation to appetitive behavior remains to be clarified. PMID- 10716544 TI - Circadian sleep, illumination, and activity patterns in women: influences of aging and time reference. AB - Patterns of sleep, illumination, and activity of women of different ages were continuously monitored in their natural environments with a wrist activity monitor. Partial correlation analyses were performed to determine relationships between age and sleep and several circadian rhythm measures including the amplitudes, mesors, and timings of sleep, of illumination, and of activity. We found no age-related decline in actigraphic sleep duration. Age was not a significant correlate of circadian rhythm parameters of sleep. Moreover, no age effects were found on daily illumination exposure or on the circadian timing of illumination and activity patterns. However, the level and amplitude of the circadian activity rhythm showed a gradual decline with aging, independent of the time reference (i.e., Daylight Saving Time versus Standard Time) when recordings were obtained. As expected, significant associations were observed between local time reference and the level and timing of peak of illumination patterns. However, changes in local time reference were not significantly and consistently associated with actigraphic sleep or activity measures. PMID- 10716545 TI - Dietary monotony and food cravings in young and elderly adults. AB - Many hypothesized mechanisms for food cravings focus on nutritional deprivation. However, outside of the laboratory, nutritional inadequacy is often confounded with dietary monotony. Therefore, one aim was to examine the effects of a nutritionally adequate, liquid, sweet, monotonous diet on food cravings in young and elderly adults. In addition, previous retrospective questionnaire work has indicated that elderly individuals report fewer food cravings than do young adults. Because there are possible age differences in cognitive capacity, another goal of this work was to develop a prospective method (i.e.. a method that depends less on memory) of studying cravings that permits comparison of adults of different ages. Young adults reported significantly more cravings per day during the monotony manipulation than during the baseline period. Therefore, nutritional deprivation is not a necessary condition for food cravings. There were no gender differences. The increase was due primarily to a greater number of cravings for entrees (i.e., foods that differed in sensory quality from the monotonous diet). Cravings for sweets did not change. In contrast to predictions that liking for the monotonous diet would decline as a result of "repetition revulsion," there was no significant change in liking for the diet over the study period. In contrast to the young adults, elderly subjects were not responsive to the monotony manipulation. Elderly men reported almost no cravings at any time during the study. Elderly women reported as many cravings as did young adults during the baseline period, but did not show an increase in cravings during the monotony period. This lack of response to dietary monotony by the elderly could result in nutritionally inadequate diets. PMID- 10716546 TI - The effects of changes in housing on feeding and wheel running. AB - The experiments explored the effects on feeding when rats were moved between individual and paired housing. In Experiment 1, rats moved to paired housing showed a 3-day suppression in feeding (initially 23%) compared to chronically individual- or pair-housed rats. In Experiment 2, half of the rats from the two control groups of Experiment 1 were moved between individual and paired housing on alternate days. Only the rats moved to paired housing showed a feeding suppression (initially 40%), but the nature of the suppression differed from Experiment 1: it appeared that only one rat of each pair showed a feeding suppression. Experiment 3 examined simultaneous introduction of running wheels and moves to paired housing. The feeding suppression induced by the move to paired housing was more immediate and shorter lived than the wheel-induced suppression. Unlike wheel access, paired housing produced only a temporary suppression of body weight. These experiments suggest that the relatively simple manipulation of moving rats from individual to paired housing results in a temporary stress-induced decrease in feeding. PMID- 10716547 TI - Sodium detection during the water absorption response in Rana pipiens. AB - Although taste in vertebrates is typically associated with specialized receptors in the lingual epithelium, Hoff and Hillyard reported that the toad, Bufo punctatus, is able to "taste" sodium with the abdominal skin. This was reflected in a differential behavioral response to hypertonic NaCl. The present study tests for the presence of such abdominal chemoreceptors in the frog Rana pipiens. The experiment was a five-condition design in which frogs were placed on filter paper saturated with: deionized water, 250 mM NaCl, 350 mM NaCl, 12.9 microM amiloride, or 350 mM NaCl + 12.9 microM amiloride. The time that the frogs remained on the test substrate before moving to a surface of deionized water was recorded. It was necessary to dehydrate the frogs to 80% of their body weight to elicit a behavioral response to the NaCl whereas dehydration to 90% of their body weight has been reported effective in Bufo punctatus. The frogs displayed significantly shorter mean times to move on both concentrations of NaCl compared to deionized water, with the shortest times occurring when 350 mM NaCl was used. Amiloride alone did not have an effect upon times to move to deionized water, but did significantly reduce the response to 350 mM NaCl. Movement to amiloride + 350 mM NaCl did not differ significantly from that to deionized water. The results indicate that Rana pipiens, like Bufo punctatus, have epithelial chemoreceptors for the detection of NaCl on hydrated surfaces and that these receptors, like those of mammals, are amiloride sensitive. PMID- 10716548 TI - The nature of the metabolic signal that triggers onset of puberty in female rats. AB - These experiments examined the effects of different refeeding stimuli on reproductive activity as measured by the onset of first estrus in prepubertal, food-restricted female rats. Such rats were placed on a restricted food intake until day 50 of age to maintain a weight of 80-90 g, and to suppress onset of puberty (normal time of puberty: 37 +/- 1.4 days of age). In Experiment 1, rats were refed at different daily caloric intakes from day 50 through day 62. First estrus was observed in all rats, with highest caloric intake after 5.7 +/- 0.6 days of refeeding. Progressively fewer rats achieved first estrus, and the time to first estrus increased as the caloric intake per day decreased. These results suggest that the highest caloric intake most closely resembles an ad lib diet in such realimented rats. The second experiment determined the duration of an ad lib food stimulus needed to initiate first estrus. Similarly growth-restricted rats were refed (on Day 50 of age) ad lib meals of 67.2 +/- 0.1 kcal, presented for periods of 12, 24, 48, or 72 h. The majority of rats (6 of 7) achieved first estrus when the ad lib meals were presented for 72 h, but progressively fewer rats achieved first estrus when such meals were presented for less time. These data indicate that an extended ad lib food stimulus (72 h) is necessary to cause onset of cycling in the majority of food-restricted, prepubertal female rats. PMID- 10716549 TI - Prolactin levels in juvenile and adult rats following acute restraint and the open field. AB - Acute restraint and exposure to a novel environment alter behavior and increase prolactin levels in rats quickly and reliably. However, little research is available that examines behavior and levels of prolactin as a result of acute exposure to one stressor immediately followed by a second stressor. Similarly, a relationship between prolactin and behavior has not been established. In the present study, juvenile (35-day-old) and adult (5-month-old) rats were either placed in a novel open field for 10 min or restrained for 10 min prior to exposure to the open field. Restrained juveniles groomed more than control juveniles and restrained adults. Conversely, restraint + open field reduced ambulation and rearing among juvenile males and females, and adult females ambulated less than control females and restrained males across both behaviors. In addition, results from the present study demonstrated the first reported relationship between prolactin and open-field behaviors. Prolactin was positively correlated with rearing and number of fecal boli, and levels were negatively correlated with freezing. Among adult females, prolactin levels were lower following restraint + open field than after exposure only to the open field. This may be explained by the psychological response to the open field when it directly followed the physical stressor of acute restraint. PMID- 10716550 TI - The genetic liability to stress and postweaning isolation have a competitive influence on behavioral organization in rats. AB - Rats housed in social isolation postweaning (isolates) show profound behavioral and neurobiological differences when compared to socially housed rats (socials). Fischer rats (F344) relative to Lewis rats are hyperresponsive and significantly more susceptible to stressful stimuli. This investigation tested the hypothesis that the behavioral effects of postweaning isolation are more pronounced in a strain of rats with high susceptibility to stress compared to a strain with low susceptibility to stress. Seventy male Sprague-Dawley, Lewis, and F344 rats were housed individually or in groups at weaning on Day 21 and tested on Day 85 in the Behavioral Pattern Monitor. There was no interaction between strain and postweaning isolation for measures of locomotor activity and exploratory behavior (holepoking). However, the postweaning isolation-induced increase in the frequency of repetitive straight movements, a measure of behavioral organization, was more pronounced in Lewis isolates compared to Sprague-Dawley and F344 isolates. These results do not support the hypothesis that rats with a higher susceptibility to stress show more pronounced changes in behavior following postweaning isolation; instead, increased susceptibility to stress may counteract the repetitive movement patterns induced by social isolation. PMID- 10716551 TI - Analysis of the feeding behavior of pigs using different models. AB - Short-term feeding behavior is conventionally analysed using random process models. The assumption underlying these models have recently been questioned and this article describes the application of both random, and more biologically based, models to the feeding behavior of pigs. Feeder visits of 16 growing pigs, housed individually from 17 to 52 kg live weight, were recorded electronically over a continuous period of 35 days. Daily food intake increased linearly with time, but there was considerable individuality in the degree of order. Pigs made between 18.8 and 80.3 (mean 47.9) daily visits to the feeder. Intervals between visits could be described by two log-normal distributions. Two Gaussian density functions were fitted to the distribution of the log-transformed intervals. For the combined data from all animals the within- and between-meal intervals were 11.2 s and 100.1 min, respectively. A model with three Gaussian functions gave an improved fit to the interval distribution. The within and between meal intervals were then estimated to be 4.2 s and 93.9 min, respectively. The middle distribution of intervals ranged from 0.5 to 38.1 min. The intervals were also described by random process models; again, a three-process model gave an improved fit compared to a two-process model. The mean estimated number of meals per day from the three Gaussian model was 14.3, and from the three process random model, 16.3. A biological interpretation of the three types of interval suggests that: (1) pigs eat in meals separated by long intervals; (2) meals consist of clusters of eating bouts separated by shorter intervals, sometimes associated with drinking; (3) within each eating bout short intervals occur as pigs constantly move in and out of the feeder. It remains unclear what underlies the observed patterns of eating. PMID- 10716552 TI - Taste effects of lingual application of cardiovascular medications. AB - Medications used to treat cardiovascular diseases such as congestive heart failure, high blood pressure, and arrhythmia, are prescribed extensively in Western countries. However, taste complaints are common side effects of many of these cardiovascular medications. Although clinical observations are helpful in determining potential taste problems from a medication, experimental studies are necessary to obtain quantitative data on taste. In the studies performed here, nine cardiovascular medications (labetalol HCl, captopril, diltiazem HCl, enalapril maleate, hydrochlorothiazide, propranolol HCl, mexiletine HCl, procainamide HCl, and propafenone HCl) were applied to the tongue in human volunteers to measure the direct effect of these drugs on taste receptors. The medications were applied topically to the tongue surface of both young and elderly subjects to mimic the situation in which the drug is secreted into the saliva. Detection thresholds ranged from 0.048 mM (propafenone) to 0.438 mM (procainamide). The detection thresholds of healthy elderly subjects did not significantly differ from young controls. The compounds tested had a predominantly bitter taste with other qualities as well. In addition, topical application of the medications to the tongue affected the taste of one or more taste stimuli, with medications differing in the pattern of taste effects exhibited. The mechanism of taste effects is not fully known, but the results of this study suggest one route may be due to medications' effect on peripheral taste receptors. PMID- 10716553 TI - Effects of acute behavioral stress and LPS-induced cytokine release on growth and energetics in mice. AB - We examined the biological cost of a single episode of acute restraint stress on the growth and energetics of mice, and compared these effects with the effects of stress associated with LPS injection and the total cost of experiencing both stressors simultaneously. We monitored growth and energetics over a 24-h period in 31-day-old mice exposed to either 4 h of restraint stress (R), i.p. injection of 0.45 mg/g of lipopolysaccharide (L), or restraint plus L injection (RL). Compared to controls, either restraint or L significantly (p < 0.05) impaired growth, feed intake, and lean and fat energy deposition. The R and L treatments depressed (>100% lower than controls) body weight change. However, when applied in concert, the effect of restraint and LPS on growth, energy deposition, and feed intake did not summate. All stressors significantly increased circulating corticosterone (p < 0.05) at 1 and 4 h following the initiation of treatment, and there was no difference between the R, L, or RL treatments. Only L and RL increased (p < 0.05) circulating IL-1b at 1 and 4 h. LPS injection elevated (p < 0.05) IL-1b. Although a single episode of behavioral stress altered growth and energy partitioning, LPS-induced cytokine release inhibited energy deposition and growth to a greater extent than behavioral stress (p < 0.05), and no further suppression of energy deposition or growth occurred when the two stressors were combined over 24 h. PMID- 10716554 TI - Conditioned taste aversion in rats for a threonine-deficient diet: demonstration by the taste reactivity test. AB - Rats avoid a diet that is deficient in one or more essential amino acids (EAAs). This phenomenon is thought to involve the development of a "learned aversion" for the sensory properties or spatial placement associated with the deficient diet. The dietary self-selection technique has been widely used to show this avoidance of the deficient diet. Because avoidance does not necessarily imply taste aversion, we used the Taste Reactivity Test initially created by Grill and Norgren (1978) to analyze the affective reactivity pattern of rats that ingested a threonine-deficient diet. The results showed that there was an increase in the aversive responses when ingesting the threonine-deficient (Thr-Dev) diet, compared to a control diet, without changes in the hedonic responses. The aversive reactions were mainly gaping, and to a lesser extent chin rubbing and head shaking. This asymmetrical shift in the Thr-Dev diet palatability is consistent with a two-dimensional hypothesis of palatability, indicating that the aversive palatability of the deficient diet was increased while the positive palatability did not change. Further evidence indicates that rats do not develop a normal behavioral satiety sequence after ingesting the threonine-deficient diet. These results indicate that a true aversion is formed to the taste of a diet that is deficient in an essential amino acid. PMID- 10716555 TI - Detection of thermophilic Campylobacter spp. in blood-free enriched samples of inoculated foods by the polymerase chain reaction. AB - The detection of thermophilic Campylobacter spp., as represented by Campylobacter jejuni, by the polymerase chain reaction (PCR) was investigated and compared with the selective agar isolation (SAI) method. Stationary-phase cultures of C. jejuni were inoculated into either blood-free enrichment broth (BFEB) or BFEB that contained 10% broccoli, crabmeat, mushroom, raw milk, and raw oyster rinses. Following a 48-h enrichment period, aliquots of food test portions were removed for simultaneous analysis by PCR and SAI. It was determined that the presence of charcoal and iron in the enrichment broth interfered with the PCR assay. Therefore, three DNA extraction techniques were developed and evaluated using a 16S rRNA primer pair in the PCR assay. The 50% end point (DL50) values (determined upon six initial C. jejuni spiking levels) were used to assess the frequency of isolation utilizing PCR versus SAI for the detection of this organism in the enrichment matrices. There were virtually no differences in detection of C. jejuni among enriched samples analyzed by PCR and SAI. Mean DL50 values (n = 3) for plain BFEB, broccoli, crabmeat, mushroom, raw milk, and raw oyster were, respectively, 0.02 (PCR) versus 0.01 (SAI), 0.01 versus 0.06, 0.07 versus 0.04, 0.03 versus 0.08, 0.01 versus 0.01, and 0.01 versus 0.01 CFU/5 g food. Significant variability in the detection limit of C. jejuni by PCR in the food enrichments was observed among DNA extraction techniques. Using 48-h enrichment cultures followed by PCR analysis could save 1 day of the time required for the presumptive identification of C. jejuni in suspected foods. PMID- 10716556 TI - Isolation and identification of enteropathogenic Campylobacter spp. from chicken samples in Taipei. AB - A total of 95 chicken samples that consisted of 34 whole chickens, 32 organs (gizzards and livers), and 29 chicken parts (drumsticks, wings, and breasts), collected from traditional retail markets (no chilling facilities) and supermarkets in Taipei, were examined for the occurrence of enteropathogenic campylobacters. Three selective media, Peterz's charcoal cefoperazone deoxycholate agar, Campy-Cefex agar, and charcoal-based selective medium, were evaluated for their efficacy to isolate Campylobacter spp. from chicken samples. The results showed that there were no differences among the three media to isolate Campylobacter spp. from all chicken samples (P > 0.05). However, there were markedly different isolation rates of campylobacters between supermarket and retail market (P < 0.05). Enteropathogenic campylobacters (C. jejuni and C. coli) were found on 68% of whole chickens, 100% of chicken parts, and 100% of organs from retail markets. In supermarkets, the isolation rates of these campylobacters from whole chickens, chicken parts, and organs were 42%, 53%, and 60%, respectively. The low isolation rates of the two campylobacters isolated from chicken samples in supermarkets differed statistically from those obtained from traditional retail markets (P < 0.10). The API CAMPY test kit also was evaluated for the identification of Campylobacter spp. as compared with the conventional identification method. The results showed that the API CAMPY test kit (Biomerieux, Marcy-l'Etoile, France) could efficiently detect 87 Campylobacter spp. isolates from chicken samples examined, with 100% agreement at the genus level to 94% at the species level as compared with conventional methods. PMID- 10716557 TI - Continuous source outbreak of campylobacteriosis traced to chicken. AB - Poultry is a source of human campylobacteriosis, but a large continuous source outbreak, heretofore, has not been attributed to both a single source of poultry and single serotype of Campylobacter. Here we report an outbreak of C. jejuni affecting 6 catering college trainees and 13 patrons of a restaurant in southern England. An epidemiological investigation successfully tracked the outbreak source to the farm of origin. Frequency of occurrence of campylobacters and outbreak serotype distribution were determined in index cases, the local population, and local chicken suppliers. The source farm was investigated and the effect of interventions assessed. A single outbreak serotype of C. jejuni was isolated from trainee chefs, patrons, and chicken supplied to the college by Wholesaler A. The Campylobacter isolation rate for Wholesaler A was 89% (98% outbreak serotype), compared to 40% for non-Wholesaler A (10% outbreak serotype). The isolation rate for 14 months averaged 85% (99% outbreak serotype) in chickens grown on two farms (X and Y) supplying Wholesaler A, contributing approximately 40% to all local cases. In the research reported here, a specific strain and hygiene practice were found to be important for understanding transmission of Campylobacter from poultry to humans in this outbreak. PMID- 10716558 TI - Psychrobacters and related bacteria in freshwater fish. AB - Three phenotypic identification systems were employed to identify 106 strains of gram-negative, nonmotile, aerobic bacteria obtained during iced storage of wild (Salmo trutta and Esox lucius) and farmed (Oncorhynchus mykiss) freshwater fish. Using diagnostic tables and computer-assisted identification, the isolates were Psychrobacter (64 strains), Acinetobacter (24 strains), Moraxella (6 strains), Chryseobacterium (5 strains), Myroides odoratus (2 strains), Flavobacterium (1 strain), Empedobacter (1 strain), and unidentified (3 strains). Overall similarities of all strains were determined for 108 characters by numerical analysis (simple matching coefficient of similarity [S] and clustering by unweighted pair group average linkage [UPGMA]). At the 77% similarity level, 92 strains formed nine major clusters (3 or more strains) and four small clusters (2 strains). Cluster 1 (25 isolates divided into two main subclusters) could be assigned to Psychrobacter phenylpyruvicus, clusters 2 and 3 (26 isolates) were designated as Psychrobacter immobilis, and clusters 4 (3 isolates) and 7 (4 isolates) were identified as Psychrobacter urativorans and Psychrobacter spp., respectively. Clusters 5 (five isolates), 6 (three isolates), and 9 (five isolates) were labeled as Acinetobacter spp., Acinetobacter johnsonii, and Acinetobacter lwoffii, respectively. Cluster 8 (12 isolates), with a high resemblance to Thornley's phenon 4 (a heterogeneous group of bacteria isolated from poultry and related to Acinetobacter), remained unnamed. The restriction pattern was identical for strains grouped into clusters 2 and 3 (P. immobilis) but was different for the remaining Psychrobacter isolates. A large proportion of isolates belonging to the family Moraxellaceae were closely related. Psychrobacters and A. johnsonii were present in freshly caught fish and river water. In the latter stages of storage, P. phenylpyruvicus and acinetobacters tended to decrease, whereas P. immobilis increased. PMID- 10716559 TI - An enzymatic-colorimetric assay for the quantification of Bifidobacterium. AB - An enzymatic-colorimetric assay for the quantification of Bifidobacterium was developed. The method, based upon the standard detection of fructose-6-phosphate phosphoketolase activity, was optimized with respect to bacterial cell pretreatment, time of incubation, and substrate concentration. The relationship between bacterial biomass and phosphoketolase activity was linear in a wide spectrum of bacterial densities. Higher sensitivity over the standard method was achieved by using 0.25% Triton X-100 in the reaction mixture to pretreat the bacterial cells. Because autoaggregation is a frequent feature among Bifidobacterium strains, this simple and reproducible method offers good advantage over viable plate count and turbidimetric techniques. The methodology can also be applied to the assessment of adherent Bifidobacterium strains to human epithelial cells. PMID- 10716560 TI - Viability of bifidobacteria in commercial dairy products during refrigerated storage. AB - Commercial milk and two brands of yogurt containing bifidobacteria were obtained from retail outlets. All products were evaluated for viability of bifidobacteria and lactic acid bacteria during refrigerated storage at 4 degrees C. Milk was evaluated at 9, 6, and 3 days prior and past its expiration date. The yogurts were evaluated at 3, 2, and 1 week prior and past their expiration. Viability of bifidobacteria and lactic acid bacteria in milk and yogurt remained above 10(6) CFU/ml or g until the expiration date of the respective products. This microbial concentration is the recommended minimum dose to receive the health benefits of these organisms. PMID- 10716561 TI - Single-strand conformation polymorphisms in the hly gene and polymerase chain reaction analysis of a repeat region in the iap gene to identify and type Listeria monocytogenes. AB - Two novel methods that allow the powerful identification of Listeria monocytogenes by polymerase chain reaction (PCR) and simultaneous differentiation by special electrophoresis formats are described. The first method involves a PCR driven single-strand conformation polymorphism (SSCP-PCR) assay using a portion of the noncoding region of the hlv gene. The assay was evaluated with 120 genetically distinct L. monocytogenes strains of either foodborne or clinical origin. Distribution of listerial strains to at least 14 SSCP types was observed. In respect to the panel of strains, 39.7% were assigned to SSCP type 3, and 19% showed SSCP type 5. Further, SSCP type 1 was found in 7.5% of all strains, SSCP type 10 in 6.7%, and 5.8% each for SSCP types 6 and 7. The SSCP types 4, 9, and 11 were infrequently described in 2.55%, 3.3%, and 4.2%, respectively, of all isolates. At least 0.85% represented each of the SSCP types 2, 13, and 14, and 1.7% displayed SSCP types 8 and 12. In the second method, the internal threonine asparagine repeat portion of the L. monocytogenes p60 protein was used for setting up a PCR-based identification and parallel differentiation assay. Ten different repeat types (RTs), according to different sizes of PCR products, were observed. Of 163 strains tested, 35.58% of samples were assigned to RT 1, 39.26% to RT 2, 3.68% to RT 3, 6.13% to RT 4, 4.29% to RT 5, 2.45% to RT 6, 5.52% to RT 7, 0.61% to RT 8, 0.61% to RT 9, and 1.83% to RT 10. The data suggest that both methods allow the simple identification and differentiation of L. monocytogenes isolates. Therefore, both the SSCP-PCR and the PCR-based identification and parallel differentiation assay could represent single-strand pretyping assays before laborious reference typing methods are applied. PMID- 10716562 TI - Rapid polymerase chain reaction/DNA probe membrane-based assay for the detection of Listeria and Listeria monocytogenes in food. AB - We describe the development of polymerase chain reaction (PCR)/DNA probe membrane based colorimetric assays for the detection and identification of Listeria and L. monocytogenes. PCR primers designed from the 16S to 23S rRNA intergenic spacer region amplified products that were reverse hybridized to membrane-bound oligonucleotide probes specific for Listeria and L. monocytogenes with a detection limit of 1 to 10 CFU/25 ml in inoculated raw and pasteurized milk samples. These qualitative assays have the potential to be integrated into testing laboratories for monitoring the microbiological quality of foods. PMID- 10716563 TI - Application of the BAX for screening/genus Listeria polymerase chain reaction system for monitoring Listeria species in cold-smoked fish and in the smoked fish processing environment. AB - The cold-smoked fish industry was used as a model for the development of a system for monitoring Listeria spp. in foods and in the food processing environment. A total of 214 samples including raw fish, fish during the cold-smoking process, finished product, and environmental samples were collected from three processing facilities over two visits to each facility. Samples were screened for Listeria spp. using the BAX for Screening/genus Listeria polymerase chain reaction system (PCR) and by culture. Listeria spp., confirmed by the API Listeria test strip or by a PCR assay targeting the L. monocytogenes hlyA gene, were isolated from a total of 89 (41.6%) samples. Of these, 80 samples also tested positive for Listeria spp. using the BAX system. Specifically, 42 (55.3%) environmental samples (n = 76), 11 (25.6%) raw materials samples (n = 43), 20 (35.1%) samples from fish in various stages of processing (n = 57), and 7 (18.4%) finished product samples (n = 38) tested positive for Listeria spp. using the BAX system. Five (4.0%) of the 125 culture-negative samples yielded BAX system-positive results. Listeria isolates from each of nine culture-positive/BAX system-negative samples yielded a positive reaction when tested in pure culture by the BAX system, suggesting that our false-negative results were likely due to the presence of low Listeria numbers in the initial enrichment as opposed to nonreacting isolates. The employment of alternative enrichment protocols, such as the two-step enrichment recommended by the manufacturer, may increase the sensitivity of the assay. PMID- 10716564 TI - Development and evaluation of a 24-hour method for the detection and quantification of Listeria monocytogenes in meat products. AB - A 24-h filter monitor-based test, Listeria-SELeCT, has been developed to quantify Listeria monocytogenes organisms in meat samples with a sensitivity of < or = 1.0 CFU/g. The technique comprises a filter monitor-based system and a colony lift immunoassay to identify and enumerate the target organism. Meat homogenates were centrifuged and the eluate was filtered to trap and immobilize the microorganisms on the filter. Fraser broth was then added to the filter apparatus to allow the organisms to become established overnight and to inhibit contaminants, after which the filters were transferred onto Modified Oxford medium agar, a selective medium for L. monocytogenes. After 10 to 12 h, a colony lift immunoassay was used to confirm and enumerate suspect colonies on the filter. A correlation study between the Listeria-SELeCT method and the most probable number technique showed the Listeria-SELeCT to be considerably more accurate than the most probable number for quantitatively determining the number of viable organisms in meat samples. Because of ease and speed of testing, the Listeria-SELeCT system also provided major advantages over the most probable number technology. PMID- 10716565 TI - Twenty-four-hour direct presumptive enumeration of Listeria monocytogenes in food and environmental samples using the ISO-GRID method with LM-137 agar. AB - A new culture medium, LM-137 agar, was developed for use with the ISO-GRID hydrophobic grid membrane filter system for direct presumptive enumeration of Listeria monocytogenes in 24 h. The method was validated against three-replicate, three-dilution most probable number procedures based on enrichment methods specified by the U.S. Department of Agriculture, the Association of Official Analytical Chemists International and the U.S. Food and Drug Administration. The study encompassed meats, dairy products, egg, produce, seafood, and environmental samples. The ISO-GRID filter method produced significantly higher recovery of L. monocytogenes from fermented sausage, hot dogs, pasteurized and raw milk, raw shrimp, and environmental swab samples (P < 0.05). The reference methods yielded significantly higher counts from frozen raw pork and cole slaw (P < 0.05). Confirmation rates of presumptive positive isolates from the filter method ranged from a low of 92% (frozen raw pork) to 100% (most other products). Neither the recovery efficiency nor the confirmation rate were affected by the presence of competing aerobic flora. PMID- 10716566 TI - Inhibitory power of kefir: the role of organic acids. AB - Milk and MRS broth fermented with kefir grains from different households were examined for inhibitory activity toward gram-negative and gram-positive strains. Fermented milk obtained with 10 g per 100 ml of inoculum (final pH 3.32 to 4.25) and MRS broth fermented with 1 and 10 g per 100 ml of inocula (final pH 4.18 to 5.25) had inhibitory power demonstrated by spot test and agar well diffusion assay. This inhibitory effect could be assigned to the undissociated form of lactic and acetic acid produced during the fermentation process. Kefir supernatants inhibited the growth of Escherichia coli 3 in nutrient broth at 37 degrees C for 24 h. However, supernatants of yogurt or milk artificially acidified with lactic and acetic acids allowed the growth of E. coli 3 in the same conditions. A bacteriostatic effect of milk fermented with kefir grains over E. coli 3 was also demonstrated. PMID- 10716567 TI - An effective lacticin biopreservative in fresh pork sausage. AB - Lacticin 3147 is a novel heat-stable bacteriocin, produced by Lactococcus lactis DPC 3147, that exhibits a broad-range inhibition spectrum similar to nisin. In this study, the effect of lacticin 3147 and nisin on the shelf life of fresh pork sausage and their ability to control pathogens (Clostridium perfringens DSM 756, Salmonella Kentucky AT1) and nonpathogenic Listeria innocua DPC 1770 was investigated. The following preservative regimens were evaluated, both in broth and sausage systems: (i) 450 ppm of sodium metabisulphite; (ii) 500 IU g(-1) or ml(-1) of nisin, (iii) 2500 arbitary units (AU) g(-1) or ml(-1) of lacticin 3147; (iv) 2% sodium lactate and 500 IU of nisin; (v) 2% sodium citrate and 500 IU g( 1) or ml(-1) of nisin; (vi) 2% sodium lactate and 2500 AU g(-1) or ml(-1) of lacticin 3147, (vii) 2% sodium citrate and 2500 AU g(-1) or ml(-1) of lacticin 3147, (viii) 2% sodium lactate, and (ix) 2% sodium citrate. There was no significant difference in the activity of nisin and lacticin 3147 against any of the target strains used, both bacteriocins performing significantly better than sodium metabisulfite against gram-positive strains in broth systems. Trends indicate that the combination of organic acids with either bacteriocin enhanced its activity against Salmonella Kentucky and L. innocua and was particularly effective in the inhibition of C. perfringens in fresh pork sausage. In addition, lacticin 3147 combined with either sodium citrate or sodium lactate maintained significantly lower (P < 0.05) total aerobic plate counts for the duration of the trials and may function as an alternative to sodium metabisulfite in the preservation of fresh pork sausage. PMID- 10716568 TI - Antimicrobial properties of pepsin-digested lactoferrin added to carrot juice and filtrates of carrot juice. AB - The objective of this study was to determine the antimicrobial activity of pepsin digested lactoferrin added to carrot juice and filtrates prepared from carrot juice. Lactoferrin isolated from raw skim milk was digested by pepsin for 4 h at pH 3. The digest of lactoferrin was lyophilized, and the antimicrobial activity of the digests was determined in peptone-yeast-glucose broth, carrot juice, permeate from carrot juice, and the dialysate of carrot juice permeate using Escherichia coli (American Type Culture Collection strain 35343) as the test organism. Growth of E. coli and the inhibitory effect of the peptide were greater in peptone-yeast-glucose broth at pH 7 than at pH 4. The peptic digest of lactoferrin did not have antimicrobial properties in carrot juice at concentrations of less than 10 mg/ml of juice. Carrot juice was filtered through a membrane with a molecular weight rejection of 10,000 or 500 Da, and the permeate was dialyzed against distilled water. Growth of E. coli was delayed in the filtrate by 5 mg but not by 1 mg of the peptic digest of lactoferrin per ml of filtrate. Bacterial counts of the control and experimental samples were not significantly different after 24 h of incubation. The peptic digest of lactoferrin at a concentration of 5 mg of digest per ml of dialysate was bacteriostatic toward E. coli after 24 h of incubation at 23 degrees C. Dialysis of permeate caused a percentage reduction in cation concentration in the permeate ranging from 69.23% (Co) to 99.32% (Na). The antimicrobial activity of lactoferrin added to carrot juice was probably inhibited by cations. PMID- 10716569 TI - Food handlers and foodborne diseases: knowledge, attitudes, and reported behavior in Italy. AB - The purpose of this study was to evaluate knowledge, attitudes, and behavior concerning foodborne diseases and food safety issues among food handlers in Italy. Face-to-face interviews were conducted within a random sample using a structured questionnaire. Of the 411 food handlers responding, 48.7% knew the main foodborne pathogens (Salmonella spp., Staphylococcus aureus, Vibrio cholerae or other Vibrio spp., Clostridium botulinum, hepatitis A virus), and this knowledge was significantly greater among those with a higher education level, in practice from a longer period of time, and who had attended education courses (P < 0.05). A vast majority (90.4%) correctly indicated those foods classified as common vehicles for foodborne diseases, and only 7.1% of food handlers were able to name five different food vehicles, each of which transmit one of the five pathogens. The proportion of those who were able to specify a food vehicle that transmitted hepatitis A virus was significantly higher for those with a higher educational level and with a longer food-handling activity. A positive attitude toward foodborne diseases control and preventive measures was reported by the great majority of food handlers, and it was more likely achieved by those who had attended education courses. This attitude was not supported by some of the self reported safe practices observed for hygienic principles, because only 20.8% used gloves when touching unwrapped raw food, and predictors of their use were educational level and attending education courses. Results strongly emphasize the need for educational programs for improving knowledge and control foodborne diseases. PMID- 10716570 TI - Isolation and characterization of antagonists for the biocontrol of the postharvest wound pathogen Botrytis cinerea on strawberry fruits. AB - Antagonistic bacteria and yeasts were isolated from the epiphytic flora of stored strawberry fruits and evaluated for their ability to protect strawberry fruit wounds after harvest against Botrytis cinerea. Among selected potential antagonists, three strains of Candida reukaufii (5L3, 10CL4, 10L2) and one strain of Candida pulcherima (10L8) still protected fruit wounds when applied at 10(3) CFU/wound, reducing lesion or conidiophore development. In the same conditions, two Enterobacteriaceae (10B1, 5B4) highly reduced pathogen development. Strain 5B4 was still highly inhibitory when inoculated at 10(2) CFU/wound. The six strains applied on fruits did not produce any significant change in color, brightness, and firmness of fruits. The two yeasts, 5L3 and 10L8, and particularly the two bacteria, 5B4 and 10B1, were selected for further studies. The four antagonists effectively colonized fruit wounds and strongly inhibited spore germination of B. cinerea in vitro. The bacterial cells surrounded the germinating spores of B. cinerea and attachment of 5L3 cells on germinating spores were additionally observed. Bacterial antagonists, particularly the strain 5B4, multiplied and rapidly used carbohydrates in strawberry fruit juice despite the low pH (pH 3.5). The efficiency of the bacterial antagonists on fruit wounds was related to their growth and nutritional properties. PMID- 10716571 TI - In vitro susceptibility of Yersinia enterocolitica isolated from the oral cavity of swine. AB - Antimicrobial susceptibility of 181 (107 ail-harboring isolates and 74 non-ail harboring) Yersinia enterocolitica isolates obtained from the oral cavity of swine was determined against 24 antimicrobial agents. All Y. enterocolitica isolates were susceptible to sulfamethoxazole/trimethoprim, enrofloxacin, aminoglycosides, and nitrofurantoin. Susceptibility to tetracycline appeared to vary by lot of origin. Isolates were resistant to sulfonamides (other than sulfamethoxazole/trimethoprim), penicillin, ampicillin, ticarcillin, cephalothin, macrolides, and tiamulin. PMID- 10716572 TI - The activity of myrosinase from broccoli (Brassica oleracea L. cv. Italica): influence of intrinsic and extrinsic factors. AB - The potential of some intrinsic (MgCl2, ascorbic acid, pH) and extrinsic (temperature, pressure) factors for controlling/altering activity of myrosinase from broccoli was investigated in this paper. A combination of MgCl2 and ascorbic acid was found to enhance enzyme activity. Concentrations resulting in optimal activity were determined as 0.1 g/liter and 2 g/liter, respectively. Both in the absence and presence of this enzyme activator, the optimal pH was situated between 6.5 and 7, corresponding to the natural pH of fresh broccoli juice. At atmospheric pressure, the enzyme was optimally active at a temperature about 30 degrees C. Application of low pressure (50 to 100 MPa) slightly enhanced the activity while at higher pressure (300 MPa), the activity was largely reduced. Future work should focus on the extension of this work to real food products in order to take cellular disruption into account. In intact vegetable tissues, the enzyme myrosinase is present in compartments separated from its substrate, the glucosinolates. Hence, enzymatic hydrolysis can merely occur after cellular disruption. In this respect, processes such as cutting, cooking, freezing, or pressurizing of the vegetables will have a large effect on the glucosinolate hydrolysis by myrosinase. This work could then be the basis for controlling glucosinolate hydrolysis in food preparation and processing. PMID- 10716573 TI - Antioxidative/Antimicrobial effects of galangal and alpha-tocopherol in minced beef. AB - The antioxidant and microbial stabilities of galangal (Alpinia galanga) extract in raw minced beef were examined at 4 +/- 1 degree C. Raw minced beef containing galangal extracts (0 to 0.10%, wt/wt) were prepared. Lipid oxidation during refrigerated storage was assessed by monitoring malonaldehyde formation, using the thiobarbituric acid reactive substances method. In minced beef, added galangal extract improved oxidative stability. Galangal extract at higher concentrations of 0.05% and 0.10% (wt/wt) were also found to extend the shelf life of minced beef. Addition of alpha-tocopherol (0.02%, wt/wt) to galangal extract (0.05%, wt/wt) were observed to increase the oxidative but not the microbial stability of minced beef during the storage of 7 days. Galangal extract may prove useful in inhibiting lipid oxidation and increasing microbial stability of minced meat. PMID- 10716574 TI - Species origin of milk in Italian mozzarella and Greek feta cheese. AB - Several animal species such as cattle, goats, sheep, and water buffalo provide milk for dairy products. We describe a simple procedure for detecting the species origin of milk used for cheese production. DNA was isolated from Italian mozzarella or Greek feta by sequential organic extractions and resin purification. This DNA was analyzed by polymerase chain reaction-restriction fragment length polymorphism as described previously for meat samples. This procedure differentiated mozzarella made from water buffalo milk and from less expensive bovine milk and also feta cheeses made from bovine, ovine, and caprine milk. PMID- 10716575 TI - Binding of aflatoxin B1 alters the adhesion properties of Lactobacillus rhamnosus strain GG in a Caco-2 model. AB - Lactic acid bacteria have been previously reported to possess antimycotoxigenic activities both in vitro and in vivo. The objective of this study was to investigate the effect of aflatoxin B1 on adhesion capability of Lactobacillus rhamnosus strain GG using a Caco-2 adhesion model. Removal of aflatoxin B1 by L. rhamnosus strain GG reduced the adhesion capability of this strain from 30% to 5%. It is therefore concluded that aflatoxins may influence the adhesion properties of probiotics able to sequester them, and subsequently these bacteria may reduce the accumulation of aflatoxins in the intestine via increased excretion of an aflatoxin-bacteria complex. PMID- 10716576 TI - Preliminary evidence that degradation of aflatoxin B1 by Flavobacterium aurantiacum is enzymatic. AB - The ability of crude protein extracts from Flavobacterium aurantiacum to degrade aflatoxin B1 (AB1) in aqueous solution was evaluated. Crude protein extracts (800 microg of total protein per ml) degraded 74.5% of AB1 in solution. An average of 94.5% of AB1 was recovered after incubation with heat-treated crude protein extracts (800 microg of total protein per ml). DNase I-treated crude protein extracts degraded 80.5% of AB1 in solution, suggesting that removal of aflatoxin by F. aurantiacum is not due to nonspecific binding with the bacterium's genomic DNA. Proteinase K-treated crude protein extracts degraded 34.5% of AB1, providing evidence that degradation of aflatoxin is linked to a protein that is possibly an enzyme. Solution pH affected the amount of AB1 degraded by crude protein extracts after 24 h. Maximum degradation was observed at pH 7 (pH levels tested: 5, 6, 7, and 8), with some AB1 degradation occurring at pH levels as low as 5 and as high as 8. Acidic pH levels were more detrimental to the ability of crude protein extracts to degrade AB1 than was basic pH. The results of this work indicate that the degradation of AB1 by F. aurantiacum may be enzymatic. PMID- 10716577 TI - A call to arms: economic barriers to optimal dialysis care. AB - Epidemic growth rates and the enormous cost of dialysis pressure end-stage renal disease (ESRD) delivery systems around the world. Payers of dialysis services can constrain costs through (1) limiting access to dialysis, (2) reducing the quality of dialysis, and (3) placing constraints on modality distribution. In order to secure the necessary resources for ESRD care, we propose that the nephrology community consider the following suggestions: First, future leaders in dialysis should acquire additional advanced training in innovative pathways such as health care economics, business and health care administration, and health care policy. Second, the international nephrology community must strongly engage in ongoing advocacy for accessible, high quality, cost-effective care.Third, efforts should be made to better define and then implement optimal dialysis modality distributions that maximize patient outcomes but limit unnecessary costs. Fourth, industry should be encouraged to lower the unit cost of dialysis, allowing for improved access to dialysis, especially in developing countries. Fifth, research should be encouraged that seeks to identify measures that will reduce dialysis costs but will not impair quality of care. Finally, early referral of patients with progressive renal disease to nephrology clinics, empowerment of informed patient choice of dialysis modality, and proper and timely access creation should be encouraged and can be expected to help limit overall expenditures. Ongoing efforts in these areas by the nephrology community will be essential if we are to overcome the challenges of ESRD growth in this new decade. PMID- 10716579 TI - Influence of initial nutritional status on continuous ambulatory peritoneal dialysis patient survival. AB - OBJECTIVES: To evaluate the influence of initial nutritional status on continuous ambulatory peritoneal dialysis (CAPD) patient survival and to identify factors determining initial nutritional status and patient mortality. DESIGN: Prospective single-center study. SETTING: Kidney Center, Soon Chun Hyang University Hospital. PATIENTS: A total of 91 consecutive CAPD patients, who underwent initial nutritional assessment at Soon Chun Hyang University Hospital, Seoul, Korea, between September 1994 and January 1999, was included in this study. All patients were assessed at a mean of 7 days after beginning CAPD (range 3 - 24 days). Forty eight patients were male, 50 were diabetics, and their mean age was 53.9 years (range 22 - 76 years). METHODS: Nutritional status was assessed by subjective global assessment (SGA), biochemical and anthropometric measurements, fat-free edema-free (FFEF) body mass by creatinine kinetics, urea kinetic studies, and calculation of the normalized protein equivalent of total nitrogen appearance (nPNA). RESULTS: By SGA, 55% were classified as having normal nutrition while 45% had signs of malnutrition; 61% of female and 31% of male patients, and 54% of diabetics and 34% of nondiabetics were classified as malnourished. Initial FFEF body mass, blood urea nitrogen (BUN), serum albumin (sAIb), residual renal function (RRF), and weekly total creatinine clearance were significantly lower in the malnourished patients than in the patients with normal nutrition. On multiple regression analysis, only FFEF body mass was an independent determinant of SGA score. On 31 January 1999, 41 patients were still on CAPD, 15 patients had died, and 27 patients had been transferred to hemodialysis. Those who died during observation were older and had lower initial FFEF body mass, % lean body mass, BUN, sAIb, RRF, and SGA score. The 2-year patient survival rate was significantly lower in the malnourished than in normal patients (67.1 % vs 91.7%, p = 0.02). On Cox proportional hazards analysis, initial age, malnutrition assessed by SGA, and FFEF body mass were identified as factors determining death. CONCLUSION: Malnutrition was present in 45% of patients commencing CAPD when assessed by SGA. Initial FFEF body mass was a determinant of SGA score and predicted death. Malnutrition as assessed by SGA was also an independent predictor of death. Initial nutritional status, therefore, appears to exert a powerful influence on CAPD patient survival. PMID- 10716578 TI - Peritoneal membrane transport and hypoalbuminemia: cause or effect? AB - OBJECTIVE: Peritoneal membrane transport has been associated with serum albumin and clinical outcome. We examined the relationship between serum albumin and peritoneal membrane transport status before and after the initiation of peritoneal dialysis. SETTING: Patients were followed at a tertiary-care regional nephrology program at St. Joseph's Hospital, McMaster University, Hamilton, Ontario, Canada. METHODS: Incident peritoneal dialysis patients between 1 January 1995 and 31 May 1998 were eligible if there was a peritoneal equilibration test within 180 days of starting dialysis, and a serum albumin value measured within 90 days prior to, and 20 to 70 days after initiating dialysis. MAIN OUTCOME MEASURES: Serum albumin, before and after the initiation of dialysis, and the presence of proteinuric renal disease were compared with the peritoneal equilibration test results. RESULTS: Among 67 identified patients, there were 7 high, 27 high-average, 26 low-average, and 7 low transporters and the mean serum albumin values before dialysis were 35.1, 37.4, 37.8, and 40.4 g/L, respectively (p < 0.001). Serum albumin values prior to the initiation of dialysis correlated significantly with the 4-hour D/P creatinine ratio (r = -0.251, p = 0.040). After initiation of dialysis, the correlation was stronger (r= -0.447, p< 0.001). Serum albumin prior to initiation of dialysis was lower for those with proteinuric than nonproteinuric renal disease (36.4 g/L vs 38.8 g/L, p = 0.04). The trend to lower serum albumin in high transporters was seen in patients with both proteinuric and nonproteinuric renal disease. CONCLUSION: The association between higher peritoneal membrane transport and lower serum albumin is present before initiation of dialysis in both proteinuric and nonproteinuric renal disease. The poor outcomes associated with low serum albumin and higher peritoneal membrane transport might be explained by other underlying factors. The contribution of inflammation, malnutrition, and fluid overload requires further study. PMID- 10716580 TI - Intraperitoneal insulin reduces plasma leptin concentration in diabetic patients on CAPD. AB - OBJECTIVE: To determine the effects of subcutaneous (SC) and intraperitoneal (IP) insulin on serum leptin concentration in type I diabetic patients with end-stage renal failure treated with continuous ambulatory peritoneal dialysis (CAPD). DESIGN: Prospective, open, before-after study. SETTING: Tertiary-care university hospital. PARTICIPANTS: Twelve type I diabetic patients with stabilized CAPD, age 43.9 +/- 2.8 years, and duration of diabetes 30.4 +/- 3.5 years. INTERVENTION: After stabilized CAPD therapy, all patients were treated first with SC insulin for a median of 3 months, and thereafter with IP insulin for another 3 months. MAIN OUTCOME MEASURES: Plasma leptin, insulin sensitivity with euglycemic clamp, and glycemic and uremic status after both treatment periods. RESULTS: During SC insulin therapy, plasma leptin concentration was significantly higher than during IP insulin (19.8 +/- 5.9 ng/mL and 12.8 +/- 6.2 ng/mL, respectively; p < 0.001). Leptin concentration was higher in CAPD patients and was related to body mass index in both genders. No correlation was detected between plasma leptin and fasting insulin, glycemic control, glucose disposal rate, or serum lipids. CONCLUSION: Plasma leptin concentration is lower during IP insulin therapy compared to SC insulin. Insulin has probably a direct effect on both peritoneal leptin clearance and adipose tissue leptin production. The significance of leptin in regulating appetite and anorexia in uremia remains unclear. PMID- 10716581 TI - Total lymphocyte count: a promising prognostic index of mortality in patients on CAPD. AB - OBJECTIVE: In view of the limitations of albumin in peritoneal dialysis (PD), we set out to evaluate whether total lymphocyte counts (TLC) could serve as a better prognostic indicator. We were also interested in how these parameters might differ between PD and hemodialysis (HD) patients. DESIGN: In a retrospective study, we reviewed 113 charts from our dialysis unit. All laboratory analyses were performed by the Department of Clinical Pathology of the Nassau County Medical Center, using standard procedures. Intact parathyroid hormone (PTH) was sent out to Nichols Laboratories. SETTING: All patients originated from the renal clinic at Nassau County Medical Center, a 612 bed public hospital. PATIENTS: The 38 PD and 75 HD patients selected had been receiving dialysis for at least 12 months and up to 3 years. The PD patients received either continuous ambulatory and/or cycler PD. For the survivors, the averages of their routine chemical analyses were considered their representative values. For the nonsurvivors, the most recent laboratory values prior to their end point were considered. MAIN OUTCOME MEASURES: Mortality or apparent malnutrition leading to transfer to HD represented the end points for PD patients. Mortality alone was used as the end point for HD patients. RESULTS: Within the PD population, serum albumin was not significantly lower in nonsurvivors compared to survivors, while the TLC was significantly lower in nonsurvivors (1277 +/- 146/mm3 vs 2249 +/- 236/mm3, p = 0.0036). The HD population demonstrated a significant difference in both TLC and serum albumin levels between its two prognostic groups; albumin was the better discriminator. Nonsurvivors had a 20% lower serum albumin than did the survivors (27.0 +/- 1.6 g/L vs 34.0 +/- 0.5 g/L, p = 0.0001). Patients on PD had a higher TLC than those on HD (p = 0.0001). CONCLUSIONS: In the HD population, but not in the PD population, both serum albumin and TLC were significantly higher in the group that survived. Serum albumin is a more powerful discriminator of mortality in the HD population, while TLC is a better discriminator of mortality in the PD population. For uncertain reasons, PD patients have a higher TLC than those on HD. PMID- 10716582 TI - Discriminative impact of ultrafiltration on peritoneal protein transport. AB - OBJECTIVE: The dialysate concentration of large proteins increases, on average, linearly during the whole peritoneal dialysis dwell, and this linear pattern seems to be independent of the rate of ultrafiltration induced by dialysis fluid. However, we observed a high variability of protein kinetics in individual dwell studies. Therefore, we studied the details of the kinetic pattern of peritoneal transport. DESIGN AND METHODS: Kinetics of beta2-microglobulin, albumin, and total protein was examined in 23 clinically stable continuous ambulatory peritoneal dialysis patients using Dianeal 3.86% (15 dwell studies) or Dianeal 1.36% (9 dwell studies) dialysis fluid. Dialysate volume was measured using radioisotopically labeled albumin as a volume marker, with corrections for sample volume and absorption of fluid and marker from the peritoneal cavity. The generalized version of the Babb-Randerson-Farrell model was applied to estimate diffusive mass transport coefficient (K(BD)) and sieving coefficient (S) for proteins and small solutes (urea, creatinine, glucose, sodium, potassium). To quantify deviations from the linear pattern of protein dialysate concentration increase, the ratio (SR) of the slope of the linear regression line for the initial 3-30 minutes, divided by the slope for the next 60 - 360 minutes, was evaluated for albumin. RESULTS: In 5 dwell studies with Dianeal 3.86% fluid, SR was lower than 1 [low albumin transport (LAT) group, median SR = 0.49, range 4.39 - 0.71], while in the other 10 dwell studies with this solution, SR was higher than 1 [high albumin transport (HAT) group, median SR = 2.77, range 1.32 - 7.56]. Clearances of albumin up to 120 minutes were higher in the HAT group than in the LAT group. The transport of fluid, beta2-microglobulin, and small solutes did not differ between the LAT and the HAT groups. K(BD) values for proteins did not differ between the groups, but S values for albumin and total protein were lower for the LAT group than for the HAT group. A similar diversity was found in the dwell studies with Dianeal 1.36%: In three dwell studies, SR for albumin was lower than 1 (median SR = 0.95, range 0.70 - 0.97), and in six dwells it was higher than 1 (median SR = 1.55, range 1.23 - 1.98). In general, the SR values observed with Dianeal 1.36% were closer to 1 than those for Dianeal 3.86%. CONCLUSIONS: Ultrafiltration may affect the initial kinetic patterns of large protein (such as albumin) transport in two opposing ways: (1) by slowing the increase of protein concentration in dialysate (due to a low sieving coefficient, LAT group), and (2) by speeding up the increase of protein concentration in dialysate (due to a high sieving coefficient, HAT group). The average pattern in a non-selected group of studies is, however, close to a steady (linear) increase. PMID- 10716583 TI - Effect of pyrazinamide and probenecid on peritoneal urate transport kinetics during continuous ambulatory peritoneal dialysis. AB - OBJECTIVE: We administered pyrazinamide (PZA) and probenecid (PB) --two well known modulators of urate transport via the proximal tubules - to evaluate their impact on urate transport through the peritoneal membrane and to clarify mechanisms affecting peritoneal transport. SETTING: A continuous ambulatory peritoneal dialysis (CAPD) unit in 2nd Hospital of IKA (Social Services Institute), Greece. PATIENTS: In 20 stable CAPD patients, on the study day, a 4 hour, 2-L, 1.36% glucose exchange was performed (control exchange). Pyrazinamide 3 g was given orally and another identical exchange was performed (study exchange). The same protocol was repeated with 2 g PB. KtN, peritoneal clearances of urea, creatinine, and urate for each exchange, and mass transfer area coefficients (MTAC) for the three solutes and their dialysate-to-plasma concentration (D/P) ratios were used to estimate peritoneal transport. RESULTS: Administration of PZA resulted in decreased clearances and MTAC values for the three solutes. The D/P ratio decreased significantly only for urate, indicating a more intense influence of PZA on urate. After PB administration, clearances of urea, creatinine, and urate were increased. MTAC and DIP ratio increased significantly only for urate (p < 0.05), demonstrating an action similar to that exerted on renal tubules. CONCLUSIONS: These findings provide evidence that unrestricted diffusion is not the only transport mechanism in the case of urate, and demonstrate the existence of an active mechanism in peritoneal urate transport with a reabsorptive and, probably, a secretive component that resembles that of renal tubule urate transport. Attention should be given in the case of CAPD patients undergoing antituberculous (PZA) treatment: it might have a negative impact on urea, creatinine, and urate peritoneal transport rates. PMID- 10716584 TI - Correlation between peritoneal equilibration test and dialysis adequacy and transport test, for peritoneal transport type characterization. Mexican Nephrology Collaborative Study Group. AB - OBJECTIVE: The aim of this study was to analyze the correlation between the peritoneal equilibration test (PET) and the dialysis adequacy and transport test (DATT) for peritoneal transport type characterization, and the degree of patients' acceptance for each test. DESIGN: Cross-sectional, observational multicenter study. SETTING: Five referral (tertiary) dialysis centers of institutional practice. PATIENTS: The study included 107 adult continuous ambulatory peritoneal dialysis (CAPD) patients with a prescription of four exchanges of 2 L per day, irrespective of age, gender, cause of end-stage renal disease, time on dialysis, nutritional status, or residual renal function. Patients on immunosuppressive therapy and those with cancer, hepatitis B, or HIV, and those having a peritonitis episode within the previous 30 days, or three or more episodes during the previous 12 months, were excluded. MAIN MEASURES: Peritoneal transport type as classified by creatinine and urea dialysis-to-plasma (D/P) ratios by PET and DATT. RESULTS: Correlation coefficients between D/P ratios for creatinine and urea, obtained for the PET and the DATT, were 0.73 for D/P creatinine and 0.96 for D/P urea. Patients were classified into high, high average, low-average, and low transport categories according to the mean and standard deviation of D/P creatinine values obtained from the PET at 4 hours. These values showed excellent concordance with those generated from the DATT data (alpha = 0.82, 95% confidence interval 0.67 - 0.93). Nineteen percent of patients showed discordance in their category when classified according to the PET versus the DATT. Patients' acceptance was better for the DATT than for the PET, as evaluated with a questionnaire. CONCLUSION: The DATT is an easy, inexpensive, and reliable test to assess peritoneal transport type, and it also provides information about peritoneal clearance of solutes and ultrafiltration. The DATT has better patient acceptance than the PET. Since the DATT has only been validated for patients on a fixed CAPD daily schedule of 4 x 2 L, the results should be confined only to patients receiving such a prescription. PMID- 10716585 TI - Normalization of clearances in peritoneal dialysis using a formula for body water derived from an end-stage renal disease population. AB - OBJECTIVE: To compare body water (V) estimates from the Chertow formula (Vc), which was derived in an end-stage renal disease population, to V estimates from the Watson formulas (Vw) in continuous ambulatory peritoneal dialysis (CAPD) patients. To identify CAPD patients in whom Vc is preferred to Vw for clearance studies. DESIGN: Retrospective analysis of clearance studies. SETTING: Dialysis units of four academic medical centers. PARTICIPANTS: 302 subjects on CAPD. INTERVENTION: 613 clearance studies by standard methods. MAIN OUTCOME MEASURES: Comparisons between Vc and Vw, and between urea clearance normalized by Vc [(KtVc)ur] and Vw [(Kt/Vw)ur]. RESULTS: Vc exceeded Vw by 3.5 +/- 1.6 L (p < 0.001), or 9.6% on average. This degree of overestimation of Vw is in the range of body water estimates found in CAPD subjects with severe volume overload (> 5% of body weight) in previous studies. Total (Kt/Nw)ur exceeded total (Kt/Vc)ur by 8.6%. By linear regression, Vc = -0.589 + (1.112 x Vw), r = 0.983. Vw exceeded Vc in only 12 studies. Young age, short height, low body weight, and low prevalence of diabetes characterized the studies with Vw > Vc. Total (Kt/Vw)ur was adequate (> or = 2.0 weekly) in 276 studies. Among these, 74 studies had inadequate total (Kt/Vc)ur (< 2.0 weekly). By logistic regression, the predictors of inadequate (Kt/Vc)ur, when (Kt/Vw)ur was adequate, included the presence of diabetes, great height, and long duration of CAPD. CONCLUSIONS: Vc provides estimates of body water exceeding those provided by Vw in a great majority of CAPD patients. Consequently, approximately 25% of the clearance studies that are adequate when Vw is used as the normalizing parameter may be inadequate when Vc is used. Vc may provide a more appropriate estimate of body water than Vw in CAPD patients with volume overload. PMID- 10716586 TI - Prevalence of TT virus in serum and peripheral mononuclear cells from a CAPD population. AB - BACKGROUND: A novel virus named TT virus (TTV) has been isolated recently from patients with posttransfusional hepatitis of unknown etiology. The prevalence of TTV in several groups at risk has been reported, however, there is no information about the prevalence of TTV in patients on continuous ambulatory peritoneal dialysis (CAPD) without blood transfusions or hemodialysis antecedents. OBJECTIVE: To study the incidence of TTV in serum and peripheral blood mononuclear cells (PBMC) of CAPD patients. DESIGN: TTV DNA was detected by polymerase chain reaction, using primers from the open reading frames (ORF) 1 and 2, in serum and PBMC from 22 CAPD patients who had not received blood transfusions or hemodialysis therapy prior to CAPD. As controls, sera from 20 patients with chronic viral hepatitis (10 with HBV and 10 with HCV) and 20 healthy donors were included in the study. RESULTS: TTV DNA was detected in the serum of 5 of 22 (22.7%) CAPD patients with both sets of primers. Four of the 5 (80%) patients with TTV DNA in their serum were TTV positive in their PBMC with primers from ORF1 and ORF2. Five of 20 (25%) patients with chronic viral hepatitis (2 patients with HBV and 3 with HCV) and 4 of 20 (20%) healthy donors were TTV DNA positive in serum. No relation was found between TTV infection and the underlying kidney disease, previous surgery, and abnormal alanine aminotransferase levels. CONCLUSION: We have found a relatively high prevalence of TTV that is similar to that found in healthy donors and in patients with chronic viral hepatitis. PMID- 10716587 TI - Quality of life in predialysis end-stage renal disease patients at the initiation of dialysis therapy. The NECOSAD Study Group. AB - OBJECTIVE: To assess health-related quality of life (QL) in a group of Dutch predialysis end-stage renal disease (ESRD) patients prior to the initiation of dialysis, and to compare QL between patients with different intended initial dialysis treatments. DESIGN: In a prospective cohort study, demographic, clinical, and QL data were obtained from Dutch adult patients who were consecutively enrolled from 27 different centers 0 - 4 weeks prior to the beginning of their chronic dialysis treatment. PATIENTS: Of the 301 patients who completed the QL questionnaires (of a possible 337 enrolled patients), 152 intended to start with hemodialysis (pre-HD) and 149 patients with peritoneal dialysis (pre-PD). MAIN OUTCOME MEASURE: Perceived QL of pre-HD and pre-PD patients. Quality of life was assessed with two generic health assessment instruments: the SF-36 and the EuroQol. RESULTS: After correction for group differences, pre-HD patients scored consistently, but not significantly, lower for all separate dimensions of the SF-36 and the overall health score of the EuroQol compared to pre-PD patients. However, analyzing the dimensions of the SF 36 together, adjusted for case-mix, pre-HD patients scored significantly lower than pre-PD patients. Mean difference was 6.5 points (p = 0.04). CONCLUSION: Multivariate adjustment for known case-mix differences at the start of dialysis therapy was not sufficient to adjust for all patient selection effects on QL. Consequently, published QL comparisons between HD and PD in nonrandomized cohort studies should be interpreted with caution. Assessment of QL just before start of dialysis therapy and subsequent adjustment for baseline values may be the only valid alternative for randomized studies. PMID- 10716588 TI - A retrospective study of seven cases of Candida parapsilosis peritonitis in CAPD patients: the therapeutic implications. AB - BACKGROUND: Candida peritonitis accounts for the majority of fungal peritonitis in continuous ambulatory peritoneal dialysis (CAPD), but the Candida species were not routinely subtyped in previous studies. The clinical course and the outcome of Candida parapsilosis peritonitis remain unclear. OBJECTIVE: To study the clinical course and outcome of C. parapsilosis peritonitis in CAPD patients. SETTING: Peritoneal dialysis unit in a regional hospital. PATIENTS AND DESIGN: A retrospective study on seven cases of C. parapsilosis peritonitis occurring in a single center over 3 years. RESULTS: The 7 patients included 4 males and 3 females. Their mean age was 62 +/- 11.5 years. Two (29%) were diabetic. Three (43%) had a history of preceding peritonitis and 5 (71 %) had received broad spectrum antibiotic within the previous 1 month. All presented with cloudy dialysate, abdominal pain, and fever. The mean dialysate white cell count was 300 +/- 168/mm3 with a predominance of neutrophils (81.4% +/- 13.1%). The mean time from onset of symptoms to diagnosis was 5.7 +/- 3.1 days. All had been treated with immediate catheter removal within 24 hours of diagnosis and antifungal therapy, including oral fluconazole, intravenous (IV) amphotericin, or their sequential combination. Environmental samplings were negative for C. parapsilosis. The overall complication rate was exceptionally high (71%), with three (43%) complicated by abscess formation requiring surgical drainage, one peritoneal adhesion (14%), and one mortality (14%). In the end, only two (29%) could resume CAPD. CONCLUSIONS: The outcome of this study group appeared worse than those previously described in the literature, and the optimal treatment for this group of patients remains unclear. PMID- 10716589 TI - Influence of plasticizer-free CAPD bags and tubings on serum, urine, and dialysate levels of phthalic acid esters in CAPD patients. AB - OBJECTIVES: To evaluate the impact of a plasticizer-free device on exposure to di (2-ethylhexyl) phthalate (DEHP) and its major metabolites in patients on continuous ambulatory peritoneal dialysis (CAPD). DEHP is the most commonly used plasticizer in polyvinyl chloride (PVC) products; it is added to CAPD bags in order to improve the flexibility of the material. Since DEHP leaches out of the plastic matrix, patients on CAPD are exposed to considerable amounts of DEHP and its metabolites. DESIGN: A prospective cross-over study. SETTING: Department of nephrology in a teaching hospital. PARTICIPANTS: Six patients (4 female, 2 male) stable on peritoneal dialysis (PD) for at least 6 months. INTERVENTIONS: Patients were switched from a plasticizer-containing PVC CAPD system (A.N.D.Y. Plus, Fresenius Medical Care, Bad Homburg, Germany) to a polyolefine-made plasticizer free system (stay-safe, Fresenius). MAIN OUTCOME MEASURES: Prior to and 42 days after the switch, 24-hour effluent dialysate and urine collections were performed and 10 mL blood was drawn. Concentrations of DEHP, mono-(2-ethylhexyl) phthalate (MEHP), phthalic acid (PA), and 2-ethylhexanol (2-EH) in urine, dialysate, and serum were determined using gas chromatography/mass spectrometry. RESULTS: Complete data were obtained from 5 patients. Serum levels of PA decreased significantly during the study period (0.137 +/- 0.078 mg/L vs 0.124 +/- 0.049 mg/L, p = 0.04), and the respective levels of DEHP decreased insignificantly (0.097 +/- 0.076 mg/L vs 0.069 +/- 0.046 mg/L, p = 0.07), whereas the concentrations of MEHP and 2-EH remained unchanged. Urine concentrations of PA were high (0.81 +/- 0.69 mg/L) but did not change substantially (0.70 +/- 0.50 mg/L). Effluent dialysate concentrations of MEHP and PA decreased significantly (0.0176 +/- 0.004 mg/L vs 0.0040 +/- 0.0007 mg/L, p = 0.043 and 0.158 +/- 0.056 mg/L vs 0.111 +/- 0.051 mg/L, p = 0.043, respectively). CONCLUSIONS: Although PD patients seem to be exposed to other sources of phthalates in addition to dialysis, use of plasticizer-free devices may help to reduce potentially immunosuppressive exposure to phthalate esters. PMID- 10716590 TI - A simple and inexpensive method of subcutaneous implantation of catheter distal segment using a Tenckhoff curled catheter. PMID- 10716591 TI - Stability of amphotericin B-lipid complex (Abelcet) in peritoneal dialysis solutions. PMID- 10716592 TI - Pichia ohmeri peritonitis in a patient on CAPD: response to treatment with amphotericin. PMID- 10716593 TI - Trichoderma pseudokoningii peritonitis in automated peritoneal dialysis patient successfully treated by early catheter removal. PMID- 10716594 TI - Abdominal pain in a CAPD patient treated with intravenous adriamycin: a chemical peritonitis? PMID- 10716596 TI - Literature. January--February. PMID- 10716595 TI - Nursing application: development of peritoneal dialysis care and nephrology nursing in Brazil. PMID- 10716597 TI - Analysis of the stability and degradation products of triptolide. AB - Triptolide is the major active ingredient of the Chinese herbal remedy Tripterygium wilfordii Hook F. (TwHF). As triptolide content is used to estimate the potency of preparations of TwHF, assessment of its stability is warranted. The accelerated stability of triptolide was investigated in 5% ethanol solution in a light-protected environment at pH 6.9, within a temperature range of 60-90 degrees C. The observed degradation rate followed first-order kinetics. The degradation rate constant (K25 degrees C) obtained by trending line analysis of Arrhenius plots of triptolide was 1.4125 x 10(-4) h(-1). The times to degrade 10% (t1/10) and 50% (t1/2) at 25 degrees C were 31 and 204 days, respectively. Stability tests of triptolide in different solvents and different pH conditions (pH4-10) in a light-protected environment at room temperature demonstrated that basic medium and a hydrophilic solvent were the major factors that accelerated the degradation of triptolide. Triptolide exhibited the fastest degradation rate at pH 10 and the slowest rate at pH 6. In a solvent comparison, triptolide was found to be very stable in chloroform. The stability of triptolide in organic polar solvents tested at both 100% and 90% concentration was greater in ethanol than in methanol than in dimethylsulphoxide. Stability was also greater in a mixture of solvent:pH6 buffer (9:1) than in 100% solvent alone. An exception was ethyl acetate, which is less polar than the other solvents tested, but permitted more rapid degradation of triptolide. Two of the degradation products of triptolide were isolated and identified by HPLC and mass spectroscopy as triptriolide and triptonide. This suggested that the decomposition of triptolide occurred at the C12 and C13 epoxy group and the C14 hydroxyl. The opening of the C12 and C13 epoxy is an irreversible reaction, but the reaction occurring on the C14 hydroxyl is reversible. These results show that the major degradation pathway of triptolide involves decomposition of the C12 and C13 epoxy group. Since this reaction is very slow at 4 degrees C at pH 6, stability is enhanced under these conditions. PMID- 10716598 TI - Triboelectrification of spray-dried lactose prepared from different feedstock concentrations. AB - Powder systems may acquire electrostatic charge during various pharmaceutical processing operations and may give rise to difficulties in handling and powder flow, mainly due to adhesion/cohesion effects. We have investigated the electrostatic charging of spray-dried lactose prepared from different feedstock concentrations using a laboratory spray-dryer. Triboelectrification of the spray dried lactose samples was effected through contact with the stainless steel surface of either a mixing vessel or a cyclone separator. Results from both techniques showed differences in charge accumulation and particle-steel adhesion between the spray-dried lactose samples. As the feedstock concentration used to produce the spray-dried lactose was increased in the range 10-50% w/v, the mean charge on the lactose decreased from -20.8 to -1.3 nC g(-1) and -54.9 to -4.1 nC g(-1) for the mixing vessel and cyclone separator, respectively, with a corresponding decrease in adhesion. In addition, as the feedstock concentration was increased from 10 to 50% w/v, decreases were obtained in surface area values (1.06 to 0.56 m2 g(-1)), pore diameter (198.7 to 83.5 microm) and pore volume (1.09 to 0.75 cm3 g(-1)), and together with differences in crystal form correlated with the charge and adhesion results. The results suggested that the feedstock concentration could have a considerable influence on the charging and adhesional properties of spray-dried lactose. This may have relevance during pharmaceutical processing and manufacturing operations. PMID- 10716599 TI - Development of a lyophilization formulation that preserves the biological activity of the platelet-inducing cytokine interleukin-11 at low concentrations. AB - Recombinant human interleukin-11 (rhIL-11) is a licensed biological therapeutic product in at least one country and is used to combat thrombocytopenia during chemotherapeutic regimens, as well as undergoing clinical trials for a range of other disorders. Following attempts to lyophilize IL-11 at low concentrations, it was clear that a significant loss of recoverable biological activity occurred. Investigation of a variety of factors, including the type of container in which the rhIL-11 was lyophilized, revealed that surface adsorption to glass was a major factor resulting in loss of activity of rhIL-11 in solution (> 40% reduction after 3 h at room temperature), in addition to losses of activity post lyophilization. To overcome this problem, different formulations containing combinations of human serum albumin (HSA), trehalose and Tween-20 have been investigated. Two formulations were successful in entirely preserving the biological activity of rhIL-11 through lyophilization and subsequent reconstitution (potency estimates of formulated relative to original material being > or =0.97). Accelerated degradation studies, performed at intervals over a six-month period, demonstrated the stability of freeze-dried rhIL-11 using these formulations (predicted annual reduction in potency after storage at -20 degrees C < or =1.4%). In conclusion, we have developed a working combination of excipients (0.5% HSA, 0.1% trehalose and 0.02% Tween-20 in potassium phosphate buffer (pH 7.4)) to formulate a stable rhIL-11 freeze-dried product in glass containers, with no loss in potency. These findings should facilitate development of low dose rhIL-11 products and be an indicator of caution to those using this and other material with similar physical properties, without taking appropriate precautions to avoid losses through adsorption. PMID- 10716600 TI - Tissue distribution and pharmacological potential of SM-16896, a novel oestrogen bisphosphonate hybrid compound. AB - Postmenopausal osteoporosis is caused mainly by a deficiency of oestrogen with rapid bone loss. To target oestrogen to the bone effectively, we have synthesized and evaluated the effects of a novel hybrid compound of oestrogen and bisphosphonate, SM-16896. The tissue distribution pattern and pharmacological potential are reported. Although the affinity for calf uterine oestrogen receptor was very low (IC50: 73.3 microM; 1/25000 of that of 17beta-oestradiol (2.84 nM)), SM-16896 showed oestrogenic activity. SM-16896 (1 microM) induced a 4.5-fold transcriptional activity in rat osteosarcoma UMR-106 cells compared with vehicle treated control, when we used the expression vector for human oestrogen receptor and a CAT reporter plasmid containing an oestrogen-responsive element. The distribution of SM-16896 after a subcutaneous administration to 7-week-old female rats was examined by radioluminography using 3H-labelled SM-16896. At 30 min after the administration, significant radioactivity was detected in the bone. At 24 h after administration, a high level of radioactivity was detected in the bone, but in the uterus it was only at a background level. Daily subcutaneous administration of 0.5 mgkg(-1) SM-16896 for 12 weeks (five times per week) to 13 week-old ovariectomized rats suppressed the ovariectomized-induced reduction in bone mineral density. A bone mineral density ratio of 120% was maintained compared with sham-operated rats, whereas a relatively low suppression of uterine weight was observed (about 50% loss compared with sham-operated rats). In the same experiment, the implantation of a 17beta-oestradiol time-release pellet (0.25 mg/pellet/90 days) almost completely suppressed the reduction of both the bone mineral density and uterine tissue weight. It is likely that the effect of SM-16896 on bone was due to its oestrogenic activity, since 1.0 mgkg(-1) SM 18108, the bisphosphonate moiety of this compound, had no effect on bone in 7 week-old ovariectomized rats. The results suggest that SM-16896, a bisphosphonate conjugated oestrogen, showed a preference profile in the uterus and bone due to its characteristic distribution pattern compared with the natural oestrogen analogue 17beta-oestradiol. Thus, bisphosphonate-conjugated oestrogens have the potential to improve patient compliance in oestrogen therapy by minimizing adverse effects and reducing the frequency of medication. PMID- 10716601 TI - The effect of divalent cations on the membrane properties and pharmacokinetics in rat of the lipid A analogue E5531. AB - To obtain information on the effects of Ca2+ on the membrane properties of the lipid A analogue E5531, we have determined the aggregate size, zeta potential, membrane fluidity, micropolarity and permeability of the E5531 membrane as a function of Ca2+ levels. Within the molar ratios of [Ca2+]/[E5531] = 1 and 3, Ca2+ increased the zeta potential of the E5531 membrane but had no effect on aggregate size (approximately 20 nm). Within the above ratios, Ca2+ decreased the membrane fluidity, as measured by micropolarity of E5531 and increased the phase transition temperature. The pharmacokinetics in rats for these samples with different membrane fluidity, prepared by changing the pre-dose formulation concentration of Ca2+, was determined and a correlation between membrane fluidity and pharmacokinetics was clearly observed. It thus appears that Ca2+ effects the membrane fluidity of E5531 as well as its pharmacokinetics in rats. PMID- 10716602 TI - Oral, intraperitoneal and intravenous pharmacokinetics of deramciclane and its N desmethyl metabolite in the rat. AB - The pharmacokinetic properties of deramciclane fumarate (EGIS-3886), a new potential anxiolitic agent, and its N-desmethyl metabolite have been investigated in Wistar rats after 10 mgkg(-1) deramciclane fumarate was administered orally, intraperitoneally or intravenously. A highly sensitive, validated and optimized gas chromatographic method with nitrogen selective detection (GC-NPD) using a solid-phase extraction technique was used to determine plasma levels of the parent compound and its N-desmethyl metabolite. After oral administration the absorption of the parent compound was very fast (t(max) 0.5h). The maximum plasma concentration (C(max)) was detected at 44.9, > or =177.8 and > or =2643.0 ngmL( 1) after oral, intraperitoneal and intravenous administration of deramciclane, respectively. For the metabolite the respective Cmax values were 32.0, > or =25.4 and 51.0 ngmL(-1). The pharmacokinetic curves of both the parent compound and its metabolite showed enterohepatic recirculation for all administration routes. The biological half-life (tbeta 1/2) for deramciclane ranged from 3.42 to 5.44 h and for the N-desmethyl metabolite the range was 2.90-5.44 h, after administration of the drug by the three different routes. After intravenous administration AUC0 infinity, of deramciclane was 29.2- and 5.4-times higher than that observed after oral and intraperitoneal treatment, respectively. These AUC0-infinity ratios were only 2.1- and 1.5-times higher for the metabolite. The absolute bioavailability of deramciclane in rats was 3.42% after oral and 18.49% after intraperitoneal administration. The comparative pharmacokinetic study of deramciclane in rat after the different administration routes showed fast absorption. Furthermore, plasma levels were found to be administration route-dependent, low bioavailability of the parent compound indicated an extremely fast and strong first-pass metabolism. The apparent volume of distribution suggested strong tissue binding after administration of the drug by any of the three routes studied. PMID- 10716603 TI - The effect of fenfluramine on the pulmonary disposition of 5-hydroxytryptamine in the isolated perfused rat lung: a comparison with chlorphentermine. AB - A possible mechanism for fenfluramine-induced pulmonary hypertension has been investigated. Fenfluramine, like chlorphentermine, may inhibit the pulmonary uptake and/or metabolism of 5-hydroxytryptamine (5-HT). This allows more 5-HT to remain in the pulmonary circulation, where it may exert a greater vasoconstrictor action resulting in pulmonary hypertension. Chlorphentermine has been shown to inhibit the uptake and metabolism of 5-HT. The effect of fenfluramine on the pulmonary disposition of [14C]5-HT has been investigated, in comparison with chlorphentermine, using a recirculating isolated perfused rat lung system. The pulmonary disposition of [14C]5-HT was assessed by measuring the change in [14C]5 HT concentration in the perfusion medium during the experiment and at the end, and the concentration in the lung at the end of the experiment. The concentration of 5-hydroxyindoleacetic acid, a metabolite of 5-HT, was measured in perfusate and lung samples. Mean pulmonary clearance of 5-HT for the control lung and lungs challenged with either fenfluramine (2.5 microM) or chlorphentermine (25 microM) was 4.514, 1.316 and 1.007 mL min(-1), respectively (n = 5). The concentration of 5-HT found in the lungs at the end of the experiment for the control and the lungs preloaded with fenfluramine or chlorphentermine was 695.05+/-9.69, 638.65+/ 10.27 and 617.3+/-14.38 ng g(-1), respectively. Fenfluramine, like chlorphentermine, inhibited the pulmonary disposition of 5-HT resulting in an elevated perfusate level of 5-HT. This is a possible contributing mechanism for fenfluramine-induced pulmonary hypertension. The effect of fenfluramine was less pronounced than chlorphentermine. PMID- 10716604 TI - Glutamate and kynurenate in the rat central nervous system following treatments with tail ischaemia or diclofenac. AB - Kynurenate is an endogenous antagonist at the allosteric glycine site on the N methyl-D-aspartate (NMDA) receptor, and may have a role in ameliorating nociceptive processes through modulation of NMDA receptor function. While antinociceptive effects of nonsteroidal anti-inflammatory drugs (NSAIDs) are mediated peripherally and possibly centrally through inhibition of prostaglandin synthesis, there is also evidence for centrally mediated prostaglandin independent antinociceptive effects that may result from increased central nervous system (CNS) concentrations of kynurenate. We have investigated the effects of the NSAID diclofenac, (40 mg kg(-1), s.c.; administered to rats 1 h before killing) or the exposure of rats to noxious stimulation (tail ischaemia for 20 min before killing), on the concentrations of glutamate and kynurenate in discrete CNS regions. Regional CNS tissue concentrations of diclofenac were between 3.0-3.8 nmolg(-1). The corresponding regional glutamate concentrations ranged between 4.8-10.6 micromol g(-1), and were significantly lower in the ischaemia group when compared with both control (15%, P < 0.05) and diclofenac treated (19%, P < 0.002) groups. Kynurenate concentrations in these CNS regions ranged between 3.3-45.8 pmol g(-1). Pairwise comparisons between the control and diclofenac-treated groups found significant increases in kynurenate concentrations in the diencephalon and lumbo-sacral regions of the CNS (P = 0.05). Noxious stimulation from tail ischaemia appeared to be associated with increased release of glutamate. Additionally, NSAIDs appeared to increase kynurenate concentrations in the spinal cord and diencephalon. Antagonism by kynurenate of glutamate effects at NMDA receptors may contribute to the antinociceptive effects of NSAIDs. PMID- 10716605 TI - Facile synthesis of a chitosan hybrid of a laminin-related peptide and its antimetastatic effect in mice. AB - Laminin, a cell adhesion protein, consists of three peptide chains (alpha-1, beta 1 and gamma-1). The beta-1 chain contains a Tyr-Ile-Gly-Ser-Arg (YIGSR) sequence that has been found to inhibit experimental metastasis in mice. We have prepared a hybrid of a water-soluble chitosan and a laminin-related peptide, and have examined its inhibitory effect on experimental metastasis in mice. A laminin related peptide, acetyl-Tyr-Ile-Gly-Ser-Arg-betaAla-OH (Ac-YIGSRbetaA-OH), was prepared by a solid-phase method. Ac-YIGSRbetaA-OH was then reacted with a water soluble chitosan. BetaAla is a spacer and was placed to avoid racemization of the Arg residue when the peptide was coupled with chitosan. Although chitosan has amino groups, they did not react with the peptide. Four methods were tried to achieve a coupling reaction, the diphenylphosphoryl azide method, the diisopropylcarbodiimide/1-hydroxybenzotriazole method, the water-soluble carbodiimide (WSC), and the 2-(1H-benzotriazole-1-yl)-1,1,3,3-tetramethyluronium tetrafluoroborate (TBTU) method, but all four methods were unsuccessful. Therefore, a small spacer, tert-butyloxycarbonyl-Gly, was intercalated in chitosan, by the TBTU method, to facilitate its coupling with the peptide. After removal of the protecting group, the Gly-chitosan was coupled with Ac-YIGSRbetaA OH by the water-soluble carbodiimide method to give Ac-YIGSRbetaAG-chitosan. Conjugation of the peptide with the larger chitosan molecule did not reduce the inhibitory effect of the peptide on experimental metastasis in mice, it actually potentiated the antimetastatic effect, demonstrating that chitosan may be effective as a drug carrier for peptides. PMID- 10716606 TI - Cyclic dipeptides in the induction of maturation for cancer therapy. AB - Studies have suggested a possible form of therapy based on the use of maturation inducing compounds to induce differentiation of neoplastic cells and stimulate faster recovery of the normal cell population. The study of the effects of nine cyclic dipeptides on biochemical markers of differentiation implicated their potential to induce differentiation. Studies were undertaken to determine the specificity of these agents for HT-29 cell cultures as well as the identification of the signal transduction pathways affected by these agents inducing the differential gene expression observed in the cells. The cyclic dipeptides studied showed a high degree of specificity, having no significant effect on Caco-2 cells (P > 0.05), representing the normal gastrointestinal mucosa. All inducers administered were shown to affect the total energy state of HT-29 cells, an effect which increased the probability of HT-29 cell differentiation. Results indicated that those agents which induced differential gene expression acted at different steps in the isolated signal transduction pathway. Cyclo(Trp-Trp) and cyclo(Phe-Pro) induced a high degree of acetylation of histones (P < 0.05), while the remaining cyclic dipeptides induced a high degree of phosphorylation of histones (P < 0.05) (cyclo(Trp-Trp) induced a moderate degree of histone phosphorylation). The results from histone phosphorylation and acetylation and cyclic AMP responsive element binding protein phosphorylation studies suggest that the cyclic dipeptides activate a chromatin switch, which leads to the increase in accessibility of lineage-specific genes for transcription. PMID- 10716607 TI - Investigation of endothelial hyperreactivity in the obese Zucker rat in-situ: reversal by vitamin E. AB - The obese Zucker rat, a popular model of insulin resistance allied with oxidant stress, is associated with either normal or paradoxically enhanced endothelial vasodilator function compared with its lean litter mate. We have investigated hindquarter endothelium-dependent vasodilation in the obese Zucker rat in-situ and have examined its relationship with oxidant stress. In perfused hindquarter preparations equivalently preconstricted with phenylephrine, vasodilator responses to the endothelium-dependent agent acetylcholine (0.03-1000 pmol) were greater in obese (pD2 = 11.03+/-0.19) compared with lean (pD2 = 10.53+/-0.13) animals (P < 0.01, two-way analysis of variance). In contrast, maximal vasodilation to the nitric oxide (NO) donor sodium nitroprusside (100 nmol) was similar in obese (59.6+/-19.8%) and lean (51.9+/-2.6%) preparations (P > 0.05). However, this exaggerated vasodilator reactivity to acetylcholine in obese animals was abolished following four-week dietary supplementation with the lipophilic antioxidant vitamin E (obese pD2 = 10.74+/-0.18; lean pD2 = 10.74+/ 0.08). This antioxidant-mediated effect was associated with a reduction (P < 0.02, two-way analysis of variance) and an enhancement (P < 0.01, two-way analysis of variance) in endothelium-dependent vasodilator responses in obese and lean hindlimb preparations, respectively. Our data therefore now point to a differential modulation of hindquarter endothelium-dependent vasodilation in the obese and lean Zucker rat by the prevailing oxidant tone, resulting in an agonist stimulated endothelial vasodilator hyperreactivity in obese animals. PMID- 10716608 TI - Long-term clomipramine treatment upregulates forebrain acetylcholine muscarinic receptors, and reduces behavioural sensitivity to scopolamine in mice. AB - We have investigated the effects of long-term treatment with clomipramine, a tricyclic antidepressant, on central muscarinic acetylcholine receptors (mAChR) in mice. Repeated clomipramine administration resulted in an increase in the forebrain receptor density value (Bmax) for [3H]quinuclidinyl benzilate, a muscarinic ligand (P < 0.05), that was dependent on dose per administration (saline or 5, 10, or 20 mg kg(-1) once a day for 7 days) and number of days treated (20 mg kg(-1) for 1, 3, 5, or 7 days). No change in apparent affinity (defined as the reciprocal of the dissociation constant) (KD) occurred. Seven daily treatments with clomipramine (saline or 5, 10, or 20 mg kg(-1)) reduced hyperlocomotion induced by scopolamine (0.5 mg kg(-1), s.c.) dose-dependently, and the effect of 20 mg kg(-1) clomipramine was significant (P < 0.05). These results suggest that an upregulation of mAChR is produced by repeated clomipramine administration, and such a change is responsible for the decreased sensitivity to the muscarinic antagonist scopolamine. PMID- 10716609 TI - Time dependent effects of glucocorticoids on adrenocorticotropin secretion of rat pituitaries ex-vivo. AB - Different glucocorticoids have been compared with respect to the inhibition of corticotropin-releasing factor (CRF)-mediated adrenocorticotropin (ACTH) secretion from pituitary fragments of the rat. The influence of time of exposure to glucocorticoids and glucocorticoid concentration has been investigated. CRF stimulated ACTH secretion of perifused rat pituitary fragments was measured by a chemiluminescence immunoassay. ACTH secretion was monitored over three days. Inhibition of CRF-stimulated ACTH secretion by glucocorticoids was quantified by the area under the curve of CRF-stimulated ACTH secretion over baseline. Concentrations needed to inhibit ACTH secretion decreased with the receptor affinities of the glucocorticoids as follows: fluticasone propionate; receptor affinity 1800, concentration 10(-8) M; budesonide, 935 and 3-2.5 x 10(-8) M; flunisolide, 478 and 5 x 10(-7) M; prednisolone, 10 and 10(-6) M. CRF-stimulated secretion was inhibited by glucocorticoids after incubation for 1 min at concentrations between 10(-8) and 10(-6) M. The same absolute quantity of the glucocorticoids produced no inhibition when incubation was prolonged to 50 min or when a lower concentration was used. Immediately after the perifusion stimulation of ACTH secretion was observed. The results suggest the possibility of minimizing the side effects of glucocorticoids by prolonging drug release. PMID- 10716610 TI - Influence of extracellular K+ concentrations on quinidine-induced K+ current inhibition in rat ventricular myocytes. AB - Hypokalaemia is one of the important risk factors for development of torsades de pointes. We recently reported that hypokalaemia increased the electrocardiographic QT interval in rats treated with quinidine, but did not alter the arrhythmogenic potency of quinidine. In this study, we have investigated the influence of extracellular potassium concentration ([K+]o) on the inhibition of several types of cardiac potassium currents by quinidine. Such types of currents include the delayed rectifier potassium current (I(K)), the transient outward current (Ito), and the inward rectifier potassium current (I(K1)), as measured in isolated rat ventricular cells using patch-clamp techniques. Concentration-dependent effects of quinidine on I(K), Ito, and I(K1) were evaluated under both normal ([K+]o = 5.4 mM) and hypokalaemic ([K+]o = 3.5 mM) conditions. In contrast to both I(K) and Ito, which were barely influenced by changes in [K+]o, I(K1) was significantly inhibited by hypokalaemia. Furthermore, while quinidine suppressed both I(K) and Ito in a concentration-dependent manner, the inhibitory potency of quinidine on these currents was not influenced by changes in [K+]o. The respective normal and hypokalaemic IC50 values for quinidine were 11.4 and 10.0 microM (I(K)), and 17.6 and 17.3 microM (Ito). Although higher concentrations of quinidine were required to inhibit I(K1), the inhibitory potency of quinidine was also found to be insensitive to changes in [K+]o. Thus, in rats, the inhibitory potency of quinidine for the K+ current types I(K), Ito and I(K1) is barely influenced by changes in [K+]o. These findings are consistent with our previous report showing that the QT-prolonging potency of quinidine was not altered under hypokalaemic conditions. However, whilst hypokalaemia does not affect I(K) or Ito, it can inhibit I(K1) and can result in QT prolongation in-vivo. PMID- 10716611 TI - Comparison of analgesic effects of khat (Catha edulis Forsk) extract, D amphetamine and ibuprofen in mice. AB - We have compared the analgesic properties of khat (Catha edulis Forsk) extract, amphetamine and ibuprofen in mice. After intragastric administration of the drugs analgesia was measured relative to water-injected controls using the hot-plate, the tail-flick, and abdominal-constriction tests. At the highest doses examined (amphetamine 1.8 mg kg(-1), ibuprofen 90 mg kg(-1), khat extract 1800 mg kg(-1)), all three substances produced analgesia, but the order of efficacy varied with the test. Khat and ibuprofen were significantly different from the control in the hot-plate assay at three or more time points post-injection. In the tail-flick test, khat and amphetamine were efficacious; ibuprofen means were somewhat lower but still significantly different from control. Higher doses of the drugs decreased the number of responses in the acetic acid-induced abdominal constriction assay. We conclude that khat, like amphetamine and ibuprofen, can relieve pain. Differences in assay results may reflect differences in modes and sites of action, as well as in the type of pain generated by the chemical and thermal stimuli for nociception. PMID- 10716612 TI - Effect of Sho-saiko-to extract on HGF and TGF-beta levels of intraorgans in liver injured rats after partial hepatectomy. AB - To examine the effects of Sho-saiko-to extract on liver regeneration, Sho-saiko to extract (0.75%, 1.5% or 3%) was administered to 70% partial hepatectomized rats with dimethylnitrosamine-induced liver-injury. S phase cell number, liver retinoid levels, hepatocyte growth factor (HGF) and transforming growth factor beta (TGF-beta) levels in each intraorgan were measured as indicators of liver regeneration. Three to seven days after hepatectomy, HGF and TGF-beta levels of the liver and spleen of the Sho-saiko-to extract groups were significantly different from the levels of the ordinary food group (P < 0.05-0.1). HGF levels in the Sho-saiko-to extract groups were approximately 1.3-1.8 times higher in the liver and approximately 1.8-2.1 times higher in the spleen compared with the levels found in the ordinary food group. TGF-beta levels in the Sho-saiko-to extract groups were approximately 0.38-0.47 times the level in the liver and 0.58 0.77 times the level in the spleen of the ordinary food group. There was no difference in HGF and TGF-beta levels of the kidney and lung between the Sho saiko-to extract group and the ordinary food group. There was a significant and positive correlation between HGF level and S phase cell number in the liver (r = 0.826, P < 0.01). There was a significant and negative correlation between TGF beta level and the retinoid level in the liver (r = -0.696, P < 0.01). In addition, the levels of the active constituents of Sho-saiko-to extract (glycyrrhetic acid, baicalin and baicalein) showed high values in the liver and spleen of partial hepatectomized rats, and increased from the third day after partial hepatectomy. These results show that Sho-saiko-to extract induces liver regeneration by increasing the production of HGF and suppressing the production of TGF-beta in the liver and spleen of partial hepatectomized rats. It was considered that the increase in the Sho-saiko-to extract active constituent levels in the liver and spleen greatly influences this action. PMID- 10716613 TI - Inhibitory effect of euphol, a triterpene alcohol from the roots of Euphorbia kansui, on tumour promotion by 12-O-tetradecanoylphorbol-13-acetate in two-stage carcinogenesis in mouse skin. AB - The anti-inflammatory activity of euphol, twelve other triterpene alcohols and sitosterol-beta-D-glucopyranoside, isolated from the dichloromethane extract of the roots of Euphorbia kansui, has been evaluated in mice with inflammation induced by 12-O-tetradecanoylphorbol-13-acetate (TPA). TPA (1.7 nmol; 1.0 microg/ear) was dissolved in acetone and 10 microL delivered to the inner and outer surfaces of the right ear of ICR mice. A triterpene alcohol, sterol glucoside or vehicle (20 microL; chloroform-methanol 1:1), was applied topically approximately 30 min before each TPA treatment. The ear thickness was measured before treatment and then oedema was measured 6 h after TPA treatment. For the two-stage carcinogenesis experiment, initiation was accomplished by administration of a single topical application of 7,12-dimethylbenz[a]anthracene (DMBA; 195 nmol; 50 microg/mouse) to the shaved backs of mice. Promotion was with 1.7 nmol (1.0 microg) TPA, applied twice weekly to the same shaved area, begun one week after the initiation. Euphol (2.0 micromol; 853 microg), or its vehicle (acetone-dimethylsulphoxide, 9:1; 100 microL), was applied topically 30 min before each TPA treatment. The number and diameter of skin tumours were measured every other week for 20 weeks. All the compounds were found to possess marked inhibitory activity and their 50% inhibitory dose for TPA-induced inflammation was 0.2-1.0 mg/ear. Topical application of euphol (2.0 micromol; 853 microg/mouse) markedly suppressed the tumour-promoting effect of TPA (1.7 nmol; 1.0 microg/mouse) in mouse skin initiated with DMBA. PMID- 10716614 TI - Bone surface strains and internal bony pressures at the jaw joint of the miniature pig during masticatory muscle contraction. AB - The long-standing debate on whether the jaw joint is loaded is due in part to the complexity of the factors involved, including a number of different muscles, each with a potentially unique role. This study sought to elucidate how two major jaw muscles, the masseter and the lateral pterygoid, influence jaw-joint loading. Twenty-five 10-month-old miniature pigs were divided into two groups, controls and pigs with the lateral capsular ligament of the jaw joint stripped surgically; this was expected to affect loading by destabilizing the joint. Rosette strain gauges were bonded to: (1) the lateral surface of the squamosal bone (equivalent to the squamosal portion of the temporal bone in humans) at the level of the articular eminence; (2) the lateral surface of the condylar neck; and (3) the lateral surface of the mandibular corpus below the molar region. Semiconductor pressure transducers were placed underneath the surfaces of the anterior slope of the condyle and the posterior slope of the articular eminence through drilled holes. Strains and internal bony pressures were recorded during stimulated tetanic contractions of the masseter or lateral pterygoid muscles. Masseter contraction, either alone or with the contralateral muscle, caused net tension in the squamosal bone and net compression in the condylar neck. The orientations were approximately vertical to the occlusal plane. Masseter contraction elevated both the condylar and eminence pressures from their resting values. The strains caused by lateral pterygoid contractions were much smaller than for the masseter with the exception of the condylar location. Ipsilateral lateral pterygoid contraction decreased both the condylar and eminence pressures from their resting values, perhaps because condylar movement altered the contact between the joint surfaces. Surgical disruption enhanced both pressure changes and bone strains under either muscle contraction but their overall patterns were not altered. In conclusion, both strains and pressures in the jaw joint varied according to specific muscle activity. PMID- 10716615 TI - A method for protrusive mandibular force measurement in children. AB - With this new method, protrusive mandibular force was studied in a homogeneous group of 69 children with similar occlusions. Maximum protrusive force ranged from 18.5 to 160 N (mean +/- SD = 81.3+/-31.6 N). Maximum protrusive force was significantly higher in males (90.7+/-30.2 N) than females (66.6+/-28.6 N) while fatigue time was not significantly different between the two groups (70.6+/-38.5 s for males and 65.1+/-33.6 s for females). Although protrusive force was stable in each session, it varied considerably between different experimental days within the same individual. No statistically significant correlation was found between maximum protrusive force and age, skeletal maturity, height, weight, overjet, maxillomandibular relation, facial height, facial widths or facial proportions. PMID- 10716616 TI - A histological study of the effect of growth hormone on odontogenesis in the Lewis dwarf rat. AB - The effect of growth hormone (GH) on the dentition has been described in children with pituitary dwarfism where teeth fail to form; those that do form tend to be reduced in size and the eruption potential is diminished. The aim here was to examine the effect of GH on odontogenesis via molar development in Lewis (control), dwarf (Dw) and Dw GH-treated (Dw+GH) rats aged 3, 6, 9, 12 and 15 days. Dw+GH animals received a twice-daily dose (65 microg/kg) of GH which commenced at 2 days of age. Animals were killed, mandibles removed, processed to embedding in paraffin, sectioned and stained for histological examination of molar morphology during development. Variations in enamel mineralization and root development were observed. In 6-day-old animals, enamel mineralization was delayed in Dw and Dw+GH animals. Root initiation was evident at 6 days of age in controls but was not observed until 9 days of age in Dw and Dw+GH animals. At 12 days of age, maturation of enamel in Dw and Dw+GH animals remained delayed. By 15 days of age no variation in tooth development was evident. These data indicate that enamel mineralization is affected by the level of circulating GH in the rat. A specific deficiency of GH did not appear to delay bone resorption prior to tooth emergence. PMID- 10716617 TI - Alteration in the expression of bone morphogenetic protein-2,3,4,5 mRNA during pathogenesis of cleft palate in BALB/c mice. AB - To identify the function of these bone morphogenetic proteins (BMP) during pathogenesis of cleft palate, an experimental model was established in BALB/c mice. Cleft palate was induced by exposure to retinoic acid on embryonic day (E)12. The expression of BMP-2,3,4,5 mRNA in normal and abnormal embryonic palatal shelves was then examined from E13 to E16 by in situ hybridization. The results showed that BMP-4 mRNA was expressed strongly and uniformly in normal epithelial cells and dispersed mesenchymal cells on E13. BMP-2,5 mRNA expression appeared only in dispersed mesenchymal cells. With the development of shelves, the staining density of BMP-2,4,5 decreased gradually in mesenchymal cells outside of the condensation and increased inside the condensation. After shelves had fused on E16, no positive signals for BMP-2,4,5 were detected in dispersed mesenchymal cells, but their expression persisted in the condensation. Exposure to retinoic acid delayed the formation of the condensation and decreased BMP 2,4,5 mRNA dramatically in mesenchyme from E13 to E15. BMP-3 mRNA expression were almost negative in either control or retinoic acid-treated groups during all stages. It was concluded that spatial and temporal expression of BMP-2,4,5 was required during normal palatogenesis, and that a deficiency of their mRNA expression may contribute to the pathogenesis of cleft palate. PMID- 10716618 TI - In vitro antimicrobial activity of propolis and Arnica montana against oral pathogens. AB - Arnica and propolis have been used for thousands of years in folk medicine for several purposes. They possess several biological activities such as anti inflammatory, antifungal, antiviral and tissue regenerative, among others. Although the antibacterial activity of propolis has already been demonstrated, very few studies have been done on bacteria of clinical relevance in dentistry. Also, the antimicrobial activity of Arnica has not been extensively investigated. Therefore the aim here was to evaluate in vitro the antimicrobial activity, inhibition of adherence of mutans streptococci and inhibition of formation of water-insoluble glucan by Arnica and propolis extracts. Arnica montana (10%, w/v) and propolis (10%, w/v) extracts from Minas Gerais State were compared with controls. Fifteen microorganisms were used as follows: Candida albicans--NTCC 3736, F72; Staphylococcus aureus--ATCC 25923; Enterococcus faecalis--ATCC 29212; Streptococcus sobrinus 6715; Strep. sanguis--ATCC 10556; Strep. cricetus--HS-6; Strep. mutans--Ingbritt 1600; Strep. mutans--OMZ 175; Actinomyces naeslundii- ATCC 12104, W 1053; Act. viscosus OMZ 105; Porphyromonas gingivalis; Porph. endodontalis and Prevotella denticola (the last three were clinical isolates). Antimicrobial activity was determined by the agar diffusion method and the zones of growth inhibition were measured. To assess cell adherence to a glass surface, the organisms were grown for 18 h at 37 degrees C in test-tubes at a 30 degree angle. To assay water-insoluble glucan formation, a mixture of crude glucosyltransferase and 0.125 M sucrose was incubated for 18 h at 37 degrees C in test-tubes at a 30 degree angle. Arnica and propolis extracts (20 microl) were added to these tubes to evaluate the % of inhibition of cell adherence and water insoluble glucan formation. The propolis extract significantly inhibited all the microorganisms tested (p < 0.05), showing the largest inhibitory zone for Actinomyces spp. The Arnica extract did not demonstrate significant antimicrobial activity. Cell adherence and water-insoluble glucan formation were almost completely inhibited by the propolis extract at a final concentration of 400 microg/ml and 500 microg/ml, respectively. The Arnica extract showed slight inhibition of the adherence of the growing cells (19% for Strep. mutans and 15% for Strep. sobrinus) and of water-insoluble glucan formation (29%) at these same concentrations. Thus, the propolis extract showed in vitro antibacterial activity, inhibition of cell adherence and inhibition of water-insoluble glucan formation, while the Arnica extract was only slightly active in those three conditions. PMID- 10716619 TI - Intrinsic regulation of differentiation markers in human epidermis, hard palate and buccal mucosa. AB - Different epithelia show extensive variation in differentiation. Epidermis and epithelium from the hard palate are both typical examples of orthokeratinized epithelia whereas buccal mucosa is an example of a non-keratinized epithelium. Each of these tissues can be distinguished morphologically and also by the expression of a number of structural proteins. Tissue explants derived from epidermis, hard palate or buccal mucosa were cultured at the air-liquid interface on collagen gels containing human dermal fibroblasts. Reconstructed epithelia that retained many of the morphological and immunohistochemical characteristics of the original tissue were formed. Cultures derived from epidermis and the hard palate both had a well-defined stratum basale, stratum spinosum, stratum granulosum and stratum corneum whereas cultures derived from buccal mucosa had no stratum granulosum or corneum and the cells retained their nuclei. Significantly more living cell layers were observed in both types of epithelia obtained from the mouth than in epidermis. The specific localization of proliferation and differentiation markers (Ki67, loricrin, involucrin, SPRR2, SPRR3 and keratin 10) closely resembled that of the tissue from which the cultures were derived. As identical three-dimensional culture models were used here, it is concluded that the differences observed between these epithelia were due to intrinsic properties of the keratinocytes. PMID- 10716620 TI - The effect of bone morphogenetic protein-7 on the expression of type I inositol 1,4,5-trisphosphate receptor in G-292 osteosarcoma cells and primary osteoblast cultures. AB - Bone morphogenetic protein-7 (BMP-7) affects differentiation of preosteoblasts enabling the resultant cells to respond optimally to acutely acting regulators. As the phosphoinositide cascade and, particularly, the calcium-mobilizing inositol 1,4,5-trisphosphate (InsP3) receptor are integral to stimulus-secretion coupling in osteoblasts, the hypothesis that BMP-7 affects InsP3 receptor expression was examined in the G-292 human osteosarcoma cell line and in primary cultures of human osteoblasts. G-292 osteosarcoma cells were found to be a valid experimental model for primary human osteoblasts, expressing osteoblastic mRNAs encoding osteocalcin, bone sialoprotein, alkaline phosphatase, alpha1-collagen, epidermal growth-factor receptor, and BMP type II receptor. When cultured long term in the presence of ascorbic acid and beta-glycerophosphate, G-292 cells underwent further osteoblastic differentiation, forming nodules and exhibiting restricted mineralization. G-292 cells responded to BMP-7 with an increase in InsP3 receptor density. Ligand-binding studies established that BMP-7 (50 ng/ml) treatment of G-292 cells increased InsP3 receptor density 2.4-fold with no apparent change in affinity. Immunoblot analysis with antibodies specific for type I, type II, and type III InsP3 receptors revealed that BMP-7 (50 ng/ml) treatment resulted in a specific increase (206+/-8%) in the type I receptor. Reverse transcription-polymerase chain reaction and Northern blot analyses of G 292 and primary human osteoblasts confirmed an increase in type I InsP3 receptor mRNA upon BMP-7 treatment. These results demonstrate that G-292 cells respond to BMP-7 with an increase InsP3 receptor density, consistent with the enhanced capacity of these cells to respond to Ca2+-mobilizing secretory hormones during osteoblast differentiation. PMID- 10716621 TI - Electrical enhancement of Streptococcus gordonii biofilm killing by gentamicin. AB - This electrical enhancement was demonstrated in an in vitro model. Streptococcus gordonii biofilms were grown for 6 days in continuous-flow reactors on one-tenth strength trypticase peptone broth. The biofilms attained a mean areal cell density of 2.4 x 10(8) c.f.u./cm2 and a thickness of approx. 19 microm. Biofilms exhibited characteristic resistance to killing by an antibiotic. When treated with 2 microg/ml gentamicin for 24 h, they exhibited a 0.84 log reduction in viable cell numbers; a 4.7 log reduction was measured in a planktonic culture. Killing of planktonic bacteria by this treatment was reduced to 1.2 log when an oxygen-scavenging enzyme was added to the medium. When a 2-mA direct current was applied during antibiotic treatment, biofilm killing increased to a 4.3 log reduction. Electrical current alone caused a 1.9 log reduction in biofilm cell counts. It is suggested that gentamicin was less effective against Strep. gordonii under anaerobic conditions than it was under aerobic conditions and that this can explain both the reduced susceptibility of the biofilm (due to oxygen depletion) and electrical enhancement of efficacy (due to oxygen generation by electrolysis). PMID- 10716622 TI - Stimulation of the rat dentine-pulp complex by bone morphogenetic protein-7 in vitro. AB - Human recombinant bone morphogenetic protein-7 (BMP-7), when applied to freshly cut dentine in monkey teeth, stimulated tertiary dentine formation, but it is unclear whether this response involved upregulation of the synthetic and secretory activity of existing odontoblasts or the induction of differentiation of new odontoblast-like cells. Using a recently developed organ-culture system for whole tooth slices, the aim here was to examine the effects of BMP-7 on the stimulation and modulation of existing odontoblasts in the absence of tissue injury. Agarose beads were soaked in a 500 ng/ml or 100 ng/ml solution of BMP-7 in culture medium and placed on the odontoblast area of the dentine pulp complex of rat tooth slices. The slices were embedded in a semisolid agar-based medium and cultured at the liquid gas interface for 7 days. Results showed that beads soaked in 500 ng/ml BMP-7 stimulated a localized increase in extracellular matrix secretion by odontoblasts at the site of application, with greater stimulatory effects than from the lower concentration. These effects may be important in the reparative processes after tissue injury within the dentine-pulp complex and may be useful in the therapeutic induction of tertiary dentinogenesis. PMID- 10716623 TI - Suppression of interleukin-10 release from human periodontal ligament cells by interleukin-1beta in vitro. AB - Periodontitis is characterized by an inflammatory process induced by periodontopathogenic bacteria in the subgingival plaque. Periodontal inflammation can be enhanced by both an increase of inflammatory stimulators, e.g. interleukin (IL)-6, and a decrease of inflammatory inhibitors, e.g. IL-10. The amount of IL 1beta is known to be increased in gingival tissues and in the gingival crevicular fluid from inflamed sites compared to healthy sites. This in vitro study sought to clarity whether IL-1beta (1 ng/ml) has a regulatory effect on the release of these two cytokines from human periodontal ligament (PDL) cells. PDL cells derived from healthy premolars were grown in the presence and absence (control) of IL-1beta. The concentration of IL-6 and IL-10 in the supernatants was assessed by enzyme-linked immunosorbent assay after 48 h of culture. PDL cells incubated with IL-1beta released significantly (p < 0.05) higher amounts of IL-6 and significantly (p < 0.01) smaller amounts of IL-10 compared to control. These results give further support to the observation that IL-1beta can increase the IL 6 secretion from PDL cells. Moreover, they provide original evidence that PDL cells secrete IL-10, which can be suppressed by IL-1beta. It is concluded that PDL cells can function as accessory immunoinflammatory cells amplifying the inflammatory process in periodontitis and, thereby, contributing to periodontal breakdown. PMID- 10716624 TI - Fibroblast growth factor-2. AB - Fibroblast growth factor-2 (FGF-2) is a heparin-binding growth factor which occurs in several isoforms resulting from alternative initiations of translation: an 18 kD cytoplasmic isoform and four larger molecular weight nuclear isoforms (22, 22.5, 24 and 34 kD). FGF-2 has pleiotropic roles in many cell types and tissues; it is a motogenic, angiogenic and survival factor which is involved in cell migration, cell differentiation and in a variety of developmental processes. Although devoid of signal peptide, it could be secreted. It acts mainly through a paracrine/autocrine mechanism involving high affinity transmembrane receptors and heparan sulfate proteoglycan low affinity receptors, but also through still unknown intracrine process(es) on intracellular targets. FGF-2 has many biological functions which are probably isoform-specific. Nevertheless, FGF-2 is not essential for embryonic development as knock-out mice for the growth factor are viable and fertile although they exhibit abnormalities in neuronal differentiation. Use of FGF-2 as therapeutic agent for the treatment of ischemic cardiovascular disease is promising and clinical trials are in progress. PMID- 10716625 TI - Heparan sulfate proteoglycans on the cell surface: versatile coordinators of cellular functions. AB - Heparan sulfate proteoglycans are complex molecules composed of a core protein with covalently attached glycosaminoglycan chains. While the protein part determines localization of the proteoglycan on the cell surfaces or in the extracellular matrix, the glycosaminoglycan component, heparan sulfate, mediates interactions with a variety of extracellular ligands such as growth factors and adhesion molecules. Through these interactions, heparan sulfate proteoglycans participate in many events during cell adhesion, migration, proliferation and differentiation. We are determining the multitude of proteoglycan functions, as their intricate roles in many pathways are revealed. They act as coreceptors for growth factors, participate in signalling during cell adhesion, modulate the activity of a broad range of molecules, and partake in many developmental and pathological processes, including tumorigenesis and wound repair. This review concentrates on biological roles of cell surface heparan sulfate proteoglycans, namely syndecans and glypicans, and outlines the progress achieved during the last decade in unraveling the molecular interactions behind proteoglycan functions. PMID- 10716626 TI - Serine hydroxymethyltransferase and threonine aldolase: are they identical? AB - Serine hydroxymethyltransferase, a pyridoxal phosphate-dependent enzyme, catalyses the interconversion of serine and glycine, both of which are major sources of one-carbon units necessary for the synthesis of purine, thymidylate, methionine, and so on. Threonine aldolase catalyzes the pyridoxal phosphate dependent, reversible reaction between threonine and acetaldehyde plus glycine. No extensive studies have been carried out on threonine aldolase in animal tissues, and it has long been believed that serine hydroxymethyltransferase and threonine aldolase are the same, i.e. one entity. This is based on the finding that rabbit liver serine hydroxymethyltransferase possesses some threonine aldolase activity. Recently, however, many kinds of threonine aldolase and corresponding genes were isolated from micro-organisms, and these enzymes were shown to be distinct from serine hydroxymethyltransferase. The experiments with isolated hepatocytes and cell-free extracts from various animals revealed that threonine is degraded mainly through the pathway initiated by threonine 3 dehydrogenase, and there is little or no contribution by threonine aldolase. Thus, although serine hydroxymethyltransferase from some mammalian livers exhibits a low threonine aldolase activity, the two enzymes are distinct from each other and mammals lack the "genuine" threonine aldolase. PMID- 10716627 TI - How do Rab proteins function in membrane traffic? AB - The Rabs are a group of GTP-binding proteins implicated for some time in the targeting of different transport vesicles within the cell, but it has been unclear how they function, or how they relate to a second group of targeting proteins, the SNAREs. Recent work, discussed in this review, has used biophysical, biochemical and genetic approaches to begin to answer these questions for Rab3, Rab5 and the yeast protein Sec4p. However, the results from these three Rabs lead to a surprising conclusion: the different Rabs seem to function via highly diverse target proteins. PMID- 10716628 TI - Heat-stress induced modulation of protein phosphorylation in virulent promastigotes of Leishmania donovani. AB - In parasites such as Leishmania, the study of molecular events induced in response to heat stress is of immense interest since temperature increase is an integral part of the life cycle. Protein phosphorylation is known to control major steps of proliferation and differentiation in eukaryotic cells. Studies on intracellular signaling systems in protozoa are relatively recent. We have examined the effect of heat shock on the protein phosphorylation status in promastigotes of Leishmania donovani. The patterns of total protein phosphorylation and specific phosphorylation at tyrosine residues were examined using [32P]-orthophosphate labelling of the parasites and immunoblotting with a monoclonal anti-phosphotyrosine antibody. The major proteins of L. donovani that were phosphorylated at 24 degrees C had apparent molecular weights of 110, 105, 66-68, 55, 36-40 and 20 kDa. Heat shock (from 24 to 37 degrees C) led to a significant decrease in phosphorylation of the majority of phosphoproteins in the virulent promastigotes. On the other hand, the avirulent promastigotes did not show any decrease in protein phosphorylation on exposure to heat stress. Predominant phosphorylation at tyrosine residues was detectable in proteins of putative size 105-110 kDa in both virulent and avirulent parasites. Heat shock led to a reduction in the level of phosphotyrosine in both these proteins in the case of virulent parasites, while no such reduction was detectable in avirulent parasites. Significant modifications in the phosphorylation status of proteins in response to heat stress including that of tyrosine containing proteins, observed exclusively in virulent parasites, suggest that modulation of protein phosphorylation/dephosphorylation may play a role in signal transduction pathways in the parasite upon heat shock encountered on entering the mammalian host. PMID- 10716629 TI - Induction of apoptosis by mistletoe lectin I and its subunits. No evidence for cytotoxic effects caused by isolated A- and B-chains. AB - Type II ribosome inactivating proteins (RIP II) are generally known to induce apoptosis in human cells by the inhibition of protein biosynthesis. Recent data from mistletoe RIP II proteins (eg. mistletoe lectin I; ML1) suggest an additional mode of apoptosis induction through the binding of their lectin part to certain cell surface receptors as is known for some human galectins. In order to clarify this possibility, we used highly sensitive flow cytometric apoptosis assays and mistletoe hololectin subunits of proven purity to show that neither human lymphocytes nor Molt-4 cells undergo apoptosis after treatment with isolated lectin-type B-chains. In contrast to earlier investigations, only the hololectin was able to induce apoptosis in these assays. We conclude that direct apoptosis induction by mistletoe lectins occurs only after uptake of the molecules into the cell due to the action of the ribosome inactivating A-chain. PMID- 10716630 TI - Dihydrofolate influences the activity of Escherichia coli dihydrofolate reductase synthesised de novo. AB - The folding of proteins into their native structures is known to depend on molecular chaperones. However, other ligands or cofactors which still require characterisation are also likely to influence protein folding. The intention of this study was to reveal how the folding of an enzyme, Escherichia coli dihydrofolate reductase, was affected by a substrate ligand, i.e. dihydrofolate. The enzyme was synthesised by coupled transcription/translation in a bacterial cell-free system. Correct folding of the protein into its native structure was measured by its enzymatic activity. Synthesis of dihydrofolate reductase was found to be inhibited, at the level of translation, by dihydrofolate. The syntheses of other proteins were also inhibited by this compound and the reasons for this inhibition could not be determined. Most notably, the specific activity of the dihydrofolate reductase formed in the presence of the substrate dihydrofolate was increased and this effect was specific for dihydrofolate reductase since it was not observed with other proteins synthesised in the same system. The increase in dihydrofolate reductase specific activity could not be attributed to mere thermal stabilisation of the fully folded enzyme by dihydrofolate. The effects of dihydrofolate on dihydrofolate reductase synthesis and activity were similar to those of the molecular chaperone DnaJ which is known to promote the folding of newly synthesised proteins. It is suggested that dihydrofolate may interact with the newly synthesised dihydrofolate reductase polypeptide chain and promote its productive folding. PMID- 10716631 TI - Characterisation of the enzymatic complex for the first step in glycosylphosphatidylinositol biosynthesis. AB - The mammalian N-acetylglucosaminyl transferase for the first step in glycosylphosphatidylinositol biosynthesis has been shown to consist of at least four components: PIG-A, PIG-C, PIG-H and GPI1. Here, the enzymatic complex is further characterised. PIG-A protein, which is thought to represent the catalytic subunit of the complex, was expressed in an epitope-tagged form in the PIG-A deficient JY5 lymphoblastoid cell line. Subcellular localisation of this protein was studied using immunofluorescence and immunoelectron microscopy. The protein was localised to both perinuclear and mitochondria-associated lamellae of the endoplasmic reticulum. Using affinity chromatography, epitope-tagged PIG-A protein was partially purified. To identify regions that might be involved in the catalytic process, computer-aided comparison was performed between PIG-A and 26 distantly related glycosyl transferases. A number of residues in the membrane proximal region of the cytoplasmic domain (230-340) were found highly conserved. Finally, a topological model of the four partners participating in the enzymatic complex is introduced to provide a working model for further structural and functional analysis. PMID- 10716632 TI - Tissue specific expression of Yrk kinase: implications for differentiation and inflammation. AB - The Src family of proto-oncogenes is a highly conserved group of non-receptor tyrosine kinases with very similar, but not identical, tissue distributions and functions. Yrk is a recently discovered new member of this family. Here we report the patterns of expression of this kinase in a variety of chicken tissues during development and after hatching, and experiments that correlate some of the observed patterns of expression with potential functions. The results show that the Yrk protein is primarily found in neuronal and epithelial cells and in monocyte/macrophages. In neuronal tissues of hatched chicks, Yrk is expressed in Purkinje cells, in the gigantocellularis of the brain-stem, and in retinal ganglion cells. In addition, staining for this kinase is also seen as thread-like and punctate patterns suggesting staining in neurites and growth cones. Epithelial cells express Yrk in the stomach during late developmental stages and after hatching but, in other epithelia such as in the peridermis, intestine and kidney, expression is high during development but low (skin) or undetectable (intestine and kidney) after hatching. These results suggest that Yrk may have several functional roles, specifically in cell migration and or differentiation during neuronal and epithelial cell development and in maintenance of the differentiated phenotype. In this study we also show that significant levels of Yrk are detected in monocytes of the blood and in tissue macrophages. Analysis of chicken hematopoietic cell lines confirmed the expression of Yrk in cells of monocyte/macrophage lineage and show for the first time in experimentally-induced inflammation that Yrk kinase activity is high during the period of monocyte infiltration, raising the possibility that this kinase plays a role in inflammation and/or response to injury. PMID- 10716633 TI - Effects of lectins with different carbohydrate-binding specificities on hepatoma, choriocarcinoma, melanoma and osteosarcoma cell lines. AB - The effects of lectins with different carbohydrate-binding specificities on human hepatoma (H3B), human choriocarcinoma (JAr), mouse melanoma (B16) and rat osteosarcoma (ROS) cell lines were investigated. Cell viability was estimated by uptake of crystal violet. Wheat germ lectin was the lectin with the most deleterious effect on the viability of H3B, JAr and ROS cell lines. The cytotoxicity of lectins with similar sugar-binding specificity to wheat germ lectin, including Maackia amurensis lectin and Solanum tuberosum lectin, was weaker than that of wheat germ lectin. N-acetylgalactosamine-and galactose binding Tricholoma mongolicum lectin ranked third, after wheat germ lectin and Maackia amurensis lectin, with regard to its effect on H3B, and ranked, together with Maackia amurensis lectin, as the lectins with the second most pronounced effects on ROS. However, the cytotoxic effects of Tricholoma mongolicum lectin on JAr were much weaker than those of Maackia amurensis lectin, Solanum tuberosum lectin and Anguilla anguilla lectin. Artocarpus integrifolia lectin, Lens culinaris lectin and Anguilla anguilla lectin possessed milder cytotoxicity than the remaining lectins. which were approximately equipotent. The mannose-binding Narcissus pseudonarcissus and Lens culinaris lectins were only weakly cytotoxic, the exception being a stronger effect on H3B. The N-acetylgalactosamine-binding Glycine max lectin and methylgalactose-binding Artocarpus integrifolia lectin similarly exhibited low cytotoxicity. It can thus be concluded that in general the ranking was wheat germ lectin > Maackia amurensis lectin approximately Trichloma mongolicum lectins > other aforementioned lectins in cytotoxicity. A particular lectin may manifest more conspicuous toxicity on certain cell lines and less on others. PMID- 10716634 TI - Phylogenetic relationships and theoretical model of human cathepsin W (lymphopain), a cysteine proteinase from cytotoxic T lymphocytes. AB - The recently described cysteine proteinase cathepsin W, also known as lymphopain, which is expressed specifically by CD8+ T lymphocytes, is phylogenetically related to the cruzipain-like group of the C1 family of peptidases. We have constructed sequence alignments and a theoretical three dimensional homology model of cathepsin W. These have allowed the characterization of signature features of cathepsin W in particular and the cruzipain lineage in general. The signature features are (1) an extended loop structure, Gly 170-Trp 200, in the second or beta-sheet domain; (2) an additional disulfide bond, Cys 25/Cys 60; (3) an additional "orphan" cysteine, Cys 5; (4) an additional residue. Ala 11, inserted after the first beta-sheet sheet; and (5) an S2 pocket lined with Phe 68 and Phe 230 which explains the preference for substrates containing Leu at P2. Further, the model suggested that cathepsin W could exist as a dimer with the Cys 5 of each monomer forming a disulfide bond and the Arg 40 Phe 46 loop (RISFWDF) forming part of the dimeric interface. By comparing cathepsin W with other members of the cruzipain group and with other C1 peptidases, six conserved residues were identified which appear in general to be characteristic of the cruzipain group, and which differentiate cruzipain group members from other C1 peptidases including those of the related cathepsin L lineage. The signature residues of the cruzipain lineage are (cruzipain numbering) Asn 33, Trp 38, Ala 124, Leu 127, Leu 164, and Pro 174. PMID- 10716635 TI - Underprediction of visibly complex chromosome aberrations by a recombinational repair ('one-hit') model. AB - PURPOSE: Published low-LET FISH data were used to test two models of chromosome aberration production based on breakage-and-reunion or recombinational repair. MATERIALS AND METHODS: Randomness of DNA double strand break induction and misrejoining is analyzed comprehensively and adopted as a working hypothesis. Proximity effects are approximated by using interaction sites. Model results are calculated using CAS (chromosome aberration simulator) Monte Carlo computer software with two adjustable parameters. CAS can emulate the specifics of any experimental painting protocol, allowing very detailed tests of the models. RESULTS: To reasonable approximation, breakage-and-reunion model predictions are consistent with low-LET FISH results, including two large, elaborate, one-paint data sets. An explicitly specified version of the recombinational-repair model severely underpredicts the frequency of the visibly complex aberration patterns most commonly observed with one-paint FISH, and is inconsistent with some observed multi-paint patterns. When high-dose effects (distortion and saturation) are taken into account quantitatively, a dose-response relation for apparently simple interchanges slightly favours the breakage-and-reunion model over the recombinational-repair model, despite being approximately linear over the dose range 2-6 Gy. CONCLUSIONS: The random breakage-and-reunion model gives comprehensive baseline predictions that are sufficiently accurate for the organization of experimental results. The data speak against complex aberrations being formed by the random recombinational repair pathway discussed here. PMID- 10716636 TI - High-LET-induced chromosome aberrations in V79 cells analysed in first and second post-irradiation metaphases. AB - PURPOSE: As an extension of previous studies, the time-course of high-LET-induced chromosomal damage was investigated in first- and second-cycle V79 Chinese hamster cells. MATERIALS AND METHODS: Cells were exposed in G1 to 10.4 MeV/u Ar ions (LET = 1226 keV/microm) and chromosomal damage was measured at 2h sampling intervals between 10 h and 34 h after irradiation. To distinguish between cells in different post-irradiation cycles, the fluorescence-plus-Giemsa technique was applied. RESULTS: For first- and second-generation cells, the number of aberrant metaphases and aberrations per metaphase were found to increase markedly with sampling time, demonstrating that cell cycle progression was delayed according to the number of lesions carried by the cell. To account for the time-dependent expression of chromosomal damage a mathematical approach was used based on the integrated flux of aberrant cells entering mitosis. Moreover, the analysis of Ar ion-induced chromosome lesions confirmed that high-LET radiation results in specific changes in the spectrum of aberration types. In particular, an increased rate of chromatid-type aberrations as well as a high frequency of chromosomal breaks was found, although the cells were exposed in G1. CONCLUSIONS: Due to the fact that cells collected at one sampling time are not representative of the entire population, the complete time-course of chromosomal damage has to be taken into account for the determination of a meaningful RBE value. Otherwise, the analysis of chromosomal damage can result in a pronounced over- or underestimation of the RBE depending on the subpopulation of cells entering mitosis at that particular sampling time. PMID- 10716637 TI - Analysis of radiation-induced chromosomal aberrations using telomeric and centromeric PNA probes. AB - PURPOSE: To generate dose-response curves for X-ray-induced chromosomal aberrations analysed in human blood lymphocytes using telomeric and centromeric peptide nucleic acid (PNA) probes. MATERIALS AND METHODS: Isolated human lymphocytes were X-irradiated with doses of 0, 1, 2, 3, 4 and 6 Gy. Aberrations were analysed in the first post-irradiation metaphases using telomeric and centromeric PNA probes. RESULTS: Similar to the dose-response curves for the yield of dicentrics and centric rings, the dose-response curves for interstitial fragments and incomplete elements (derived from either terminal deletions or incomplete exchanges) follow a linear-quadratic function. Furthermore, it was estimated that 76% of excess acentric fragments originate from complete exchanges (interstitial deletions) and only 24% from incomplete exchanges or terminal deletions. CONCLUSIONS: Interstitial fragments form a major class of radiation induced chromosomal aberrations. They are induced about half as frequently as dicentrics over the whole dose range investigated. The comparable trend of the dose-response curve for the different aberrations, including incomplete elements, indicates that all detected aberrations are formed by a similar underlying mechanism. It also suggests that the ratio between non- or incomplete repair (leading to open ends of broken chromosomes) and incorrect repair (leading to exchange aberrations) is independent of dose. PMID- 10716638 TI - Dose-response relationship for radiation-induced mutations at micro- and minisatellite loci in human somatic cells in culture. AB - PURPOSE: The study was designed to determine the dose-response relationship for radiation induction of mutations at mini- and microsatellite loci in human somatic cells. Mutations induced by graded doses of gamma-irradiation were quantified by screening clones derived from single irradiated cells for micro- and minisatellite alterations following irradiation with 1, 2 or 3 Gy. MATERIALS AND METHODS: After irradiation, the moderately radioresistant glioma cell line UVW was seeded at low density into Petri dishes to allow formation of discrete colonies, 100 of which were examined at each dose. All the cells within a colony were presumed to have arisen from a single irradiated cell. Radiation-induced microsatellite alterations were determined at 16 different loci, by PCR amplification and visualization on polyacrylamide gels. Minisatellite alterations were identified at four different minisatellite loci by restriction enzyme digestion and Southern blotting. RESULTS: A dose-response curve for mutation frequency was obtained by analysis of 100 clones, yielding a minisatellite mutation rate of 5.5x10(-3) mutations/locus/Gy/cell and a microsatellite mutation rate of 8.75x10(-4) mutations/locus/ Gy/cell. At microsatellite loci, alterations were predominantly simple loss or gain of repeat units and loss of heterozygosity (LOH). The mutations in minisatellite loci resulted predominantly in LOH and variation in repeat number. The background instability at each locus was determined by analysis of non-irradiated clones. Only 2% and 1% of the micro-and minisatellite loci respectively showed altered bands. CONCLUSIONS: This is the first report of a dose-response relationship for radiation-induced micro- and minisatellite mutations in human somatic cells. Described is a sensitive method for analysis of low-dose radiation mutagenesis in somatic cells that may prove to be a useful tool for radiation protection and dosimetry. PMID- 10716639 TI - Ionizing radiation enhances double-strand-break repair in rapamycin-treated ataxia telangiectasia lymphoblasts. AB - PURPOSE: Unlike normal cells, ataxia telangiectasia (A-T) cells do not exhibit enhanced double-strand-break (dsb)-repair fidelity after ionizing radiation (IR) exposure. DNA-repair induction and other responses to IR are mediated by signalling pathways that are defective in A-T cells. Ataxia telangiectasia mutated (ATM) protein, the mutated protein in A-T cells, is homologous to members of the P13-kinase, including the rapamycin target FRAP. Rapamycin kills normal cells more readily in normal than in A-T cells, and inhibits the FRAP target p70 S6 kinase (p70S6K) more readily in normal than in A-T cells. Also, PHAS-1, another FRAP target, may also be a substrate for ATM. Because ATM plays a role in the response of mammalian cells to rapamycin, the effect of rapamycin on IR enhancement of dsb repair was investigated. MATERIALS AND METHODS: Double-strand break repair by normal and A-T lymphoblasts, either untreated or treated with rapamycin and y-irradiated, was analysed using shuttle pZ189 containing a dsb. RESULTS: Double-strand-break-rejoining fidelity in linear plasmids (linDNA) processed in either untreated or rapamycin-treated normal cells increased about twofold if transfection occurred immediately after host irradiation. In contrast, radiation did not increase or decrease the repair fidelity by untreated A-T host cells. But, like normal hosts, dsb-repair fidelity improved twofold in A-T hosts treated with rapamycin. Treatment with the P13-kinase inhibitor LY294002 did not alter dsb-rejoining fidelity in normal or in A-T hosts. CONCLUSIONS: These findings demonstrate that rapamycin does not affect IR enhancement of dsb-repair fidelity in normal cells but restores this phenomenon in A-T lymphoblasts, and suggests the involvement of a rapamycin-sensitive pathway in radiation-enhanced dsb repair. PMID- 10716640 TI - Identification of genes regulated by UV/salicylic acid. AB - PURPOSE: Previous work from the authors' group and others has demonstrated that some of the effects of UV irradiation on gene expression are modulated in response to the addition of salicylic acid to irradiated cells. The presumed effector molecule responsible for this modulation is NF-kappaB. In the experiments described here, differential-display RT-PCR was used to identify those cDNAs that are differentially modulated by UV radiation with and without the addition of salicylic acid. MATERIALS AND METHODS: Differential-display RT PCR was used to identify differentially expressed genes. RESULTS: Eight such cDNAs are presented: lactate dehydrogenase (LDH-beta), nuclear encoded mitochondrial NADH ubiquinone reductase 24 kDa (NDUFV2), elongation initiation factor 4B (eIF4B), nuclear dots protein SP100, nuclear encoded mitochondrial ATPase inhibitor (IF1), a cDNA similar to a subunit of yeast CCAAT transcription factor HAP5, and two expressed sequence tags (AA187906 and AA513156). CONCLUSIONS: Sequences of four of these genes contained NF-kappaB DNA binding sites of the type that may attract transrepressor p55/p55 NF-kappaB homodimers. Down-regulation of these genes upon UV irradiation may contribute to increased cell survival via suppression of p53 independent apoptosis. PMID- 10716641 TI - Multidentate hydroxypyridinonate ligands for Pu(IV) chelation in vivo: comparative efficacy and toxicity in mouse of ligands containing 1,2-HOPO or Me 3,2-HOPO. AB - PURPOSE: To identify the most effective multidentate 1,2-HOPO and Me-3,2-HOPO ligands for chelation of Pu(IV) in vivo. MATERIALS AND METHODS: Two sets of ligands with four identical backbones were prepared containing two, three or four bidentate 1,2-HOPO or Me-3,2-HOPO groups, and 3,4,3-LI(1,2-HOPO) was resynthesized in a higher yielding procedure. They were evaluated in mouse for acute toxicity and reduction of tissue 238Pu, in comparison with CaNa3-DTPA (30 micromol kg(-1)). RESULTS: Nine HOPO ligands, promptly injected or given orally or injected at low dosage, are superior to CaNa3-DTPA for reducing 238Pu retention in mouse. Five, given by delayed injection or promptly injected or orally administered as ferric complexes, are superior to CaNa3-DTPA or FeNa2-DTPA respectively. The Me-3,2-HOPO ligands are more effective than their structural 1,2-HOPO analogues, demonstrating the greater affinity of Me-3,2-HOPO for Pu(IV) in vivo. CONCLUSIONS: The most efficacious ligand, 3,4,3-LI(1,2-HOPO), contains the less stably binding 1,2-HOPO group; therefore, its linear spermine backbone must confer advantages for Pu(IV) binding (greater solubility, more favorable arrangement of ligating groups, more flexible backbone). Effective low toxicity tetradentate 5-LIO(Me-3,2-HOPO) and hexadentate TREN-(Me-3,2-HOPO) and highly effective but moderately toxic 3,4,3-LI(1,2-HOPO) (LD50 approximately 300 micromol kg(-1) in mouse) are recommended for further investigation. PMID- 10716642 TI - Comparative biokinetics of plutonium and americium after inhalation of PuO2 and mixed oxides (U, Pu)O2 in rat. AB - PURPOSE: To compare the biokinetics of Pu and Am in rat after inhalation of PuO2 and two (U, Pu) mixed oxides (MOX), referred to as MIMAS and SOLGEL. MATERIALS AND METHODS: Lung clearance was measured in vivo by X-and y-ray spectrometry. Retention of Pu and Am in femurs, liver and kidneys was measured by alpha spectrometry. RESULTS: Observed lung clearance was in the same range for all three powders. Extra-pulmonary transfers were expressed as the percent of the initial deep lung deposit (IDLD) measured 7 days after inhalation. After PuO2 exposure, bone retention remained nearly constant throughout the 270-day experiment. It was approximately 0.7% of the IDLD for Pu and Am. By contrast, a gradual increase was observed for the two MOX. After 7 days, bone retention of Pu and Am was respectively 0.05 and 0.08% for MIMAS, and 0.2 and 0.6% for SOLGEL. The retention reached maximal values between 180 and 270 days post-exposure, which were 0.2 and 0.3% for MIMAS, and 1.2 and 2.8% for SOLGEL for Pu and Am respectively. CONCLUSIONS: Different transfer rates of Pu and Am from the lung were observed depending on the chemical composition of the oxides and/or the method of their preparation. PMID- 10716643 TI - The Bowman-Birk protease inhibitor enhances clonogenic cell survival of ionizing radiation-treated nucleotide excision repair-competent cells but not of xeroderma pigmentosum cells. AB - PURPOSE: The radioprotective effect of the Bowman-Birk protease inhibitor (BBI) was previously shown to result from a TP53 dependent mechanism. Whether this effect involves specific DNA repair mechanisms is now tested. MATERIAL AND METHODS: Normal human fibroblasts were pre-treated with BBI before exposure to X rays, UVB or to chemical agents (bleomycin, N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), cisplatin). These agents were chosen because of their ability to induce different spectra of DNA damage. The radiometric agent bleomycin primarily induces double-strand breaks (dsb), which are repaired by recombination; MNNG results in alkylated bases which are repaired by base excision repair (BER); cisplatin results in DNA-crosslinks which are repaired mainly by nucleotide excision repair (NER); and finally UVB generates thymine dimers and thymine cytosine-6-4 products which are also repaired by NER. Cell survival was analysed by colony formation assay and DNA dsb by constant field gel electrophoresis. The combined effect of BBI and X-rays was also tested for XP-fibroblasts, which are defective in NER. RESULTS: For normal human fibroblasts the radioprotective effect of BBI was clearly found by using a delayed plating procedure. The radioprotective effect was found to be unrelated to an altered induction or repair of radiation-induced DNA dsb. Pretreatment with BBI did not affect cell killing after exposure to bleomycin or MNNG, but resulted in a significant protection of cells exposed to cisplatin or UVB. These results indicate that pre treatment with BBI did not alter recombination repair or BER, but was able to modify NER. The latter finding was supported by the observation made for XP cells, where pretreatment with BBI failed to result in radioprotection after exposure to ionizing radiation. CONCLUSIONS: On the basis of these data it is proposed that the radioprotective effect of BBI is the result of an improved nucleotide excision repair mechanism. PMID- 10716644 TI - In vitro differentiation characteristics of human skin fibroblasts: correlations with radiotherapy-induced breast fibrosis in patients. AB - PURPOSE: To determine whether there is an association between dermal fibroblast differentiation characteristics in vitro and breast fibrosis developing in patients following radiotherapy for breast cancer. MATERIALS AND METHODS: Three hundred and eighty-five patients had been characterized for the degree of breast fibrosis and the level of clinical risk factors for fibrosis as established by logistic regression. Early-passage fibroblasts from 79 patients with a high (HR) or low (LR) level of risk factors were studied in vitro. The percentage differentiated cells (%DC) 7 days after 0 and 8 Gy was scored, and unirradiated colonies were scored for the ratio of early:late fibroblast differentiation stages (E:L ratio). RESULTS: %DC: For the 0 Gy data there was a significant interpatient variation (CoV = 55%, p = 0.0001). HR patients with breast fibrosis had a higher %DC compared with patients without (p = 0.017). E:L ratio: for HR patients there was a significant interpatient variation (82%, p = 0.0030) and a lower E:L ratio for patients with fibrosis compared with those without (p = 0.086), but for LR patients this relationship was reversed (p = 0.079) CONCLUSIONS: There was a true interpatient variation in the in vitro parameters of fibroblast differentiation but insufficient correlation with observed fibrosis after radiotherapy for use as a predictive test. PMID- 10716645 TI - Radiation effects in the small bowel of the diabetic mouse. AB - PURPOSE: Irradiation of the small intestine in the mouse induces damaging structural alterations to the architecture of the enteric mucosa. There is growing interest in the possible relevance of underlying additional pathology when appreciating the total response of tissues to irradiation. The possibility that small intestinal mucosal abnormalities in the streptozotocin-induced diabetic mouse may exacerbate radiation-induced injury was tested by examining the combined effects of the two treatments. MATERIALS AND METHODS: Streptozotocin diabetic and -non-diabetic mice were exposed to 10 Gy abdominal X-radiation. Profiles of mucosal epithelial cell populations were quantified and comparisons with corresponding groups of unirradiated mice made on the third day post irradiation. RESULTS: The histological appearances of the small intestinal mucosa were similar in both groups of irradiated mice, but the numbers of profiles of crypts and of columnar, goblet, Paneth and entero-endocrine cells were depressed in these groups when compared with values in corresponding groups of unirradiated mice. However, the expression of radiation damage in the diabetic mouse was less severe than in the non-diabetic mouse, particularly in the jejunum where the changes attendant on the onset of diabetes were most marked. CONCLUSION: These findings suggest that the response of mouse to radiation may be moderated by the presence of this type of pathophysiology. However, there is no evidence that the damage produced by streptozotocin-induced diabetes and radiation is additive. PMID- 10716646 TI - Correlation of micronucleus and apoptosis assays with reproductive cell death can be improved by considering other modes of death. AB - PURPOSE: To evaluate, using 10 cell lines, the already shown improved correlation between the sum of micronucleated plus apoptotic cells and reproductive cell death. MATERIALS AND METHODS: The cell lines were X-irradiated with doses selected to obtain comparable survival levels. Micronucleated and apoptotic cells as well as abnormal cells (multinucleated and necrotic cells) were counted over several days. Apoptosis was also confirmed by gel electrophoresis. RESULTS: The data confirmed the already shown improved relationship over the solely performed micronucleus or apoptosis assays with reproductive cell death and radiation dose. However, even this approach revealed a saturation after irradiation in certain cell lines. Including other modes of cell death such as mitotic failure or necrosis could eliminate this effect. CONCLUSION: Multiple parameters should be considered when evaluating the use of a predictive assay for ionizing radiation induced cell killing or biological dosimetry. PMID- 10716647 TI - WR-151327 increases resistance to Klebsiella pneumoniae infection in mixed-field- and gamma-photon-irradiated mice. AB - PURPOSE: To determine the efficacy of WR-151327 (WR) [S-3-(3 methylaminopropylamino) propylphosphorothioic acid; (CH3-HN-(CH2)3-NH-(CH2)3-S PO3H2)] in increasing resistance to bacterial infection after a sublethal dose of gamma-photons or mixed-field neutrons plus gamma-photons. MATERIALS AND METHODS: B6D2F1/J female mice received 200 mg/kg WR i.p. or saline vehicle 20-30 min before or after sham (0 Gy) or 7.0 Gy 60Co gamma-photon irradiation. WR or saline vehicle was given only before 3.5 Gy TRIGA-reactor-produced mixed-field [n/(n+y) = 0.67] irradiation. Four days after drug treatment or drug treatment and irradiation, graded doses of Klebsiella pneumoniae were injected s.c. into mice, and 30-day survival was recorded. To assess haemopoietic changes other unirradiated and irradiated mice not injected with bacteria were given WR or saline. Peripheral blood (PB) and femoral bone marrow (BM) cells were measured 1, 3 or 4, 7, 10 and 14 or 15 days later. RESULTS: WR pretreatment increased resistance to infection in irradiated but not in unirradiated mice. Bacterial CFU LD50/30 values for 0 Gy saline-treated mice were 1.20x10(6); for 0 Gy WR-treated mice 1.16x10(6); for gamma-photon-irradiated saline-treated mice 3.02x10(1); for gamma-photon-irradiated WR-treated mice 1.24x10(4); for mixed-field-irradiated saline-treated mice 1.94x10(2); and for mixed-field-irradiated WR-treated mice 6.13x10(3). WR-induced resistance to infection paralleled increased numbers of PB white cells, neutrophils, platelets, femoral BM cells and granulocyte macrophage colony-forming cells (GM-CFC) in irradiated mice not given bacteria. CONCLUSIONS: These studies quantify the resistance to bacterial infection in mice treated with WR before sublethal irradiation. The findings suggest that WR treatment increases resistance to infection in immunocompromised hosts. PMID- 10716648 TI - Enhancement of radiation-induced mitotic catastrophe by moderate hyperthermia. AB - PURPOSE: To determine if a maximally radiosensitizing but non-toxic moderate hyperthermia treatment enhances radiation-induced mitotic catastrophe. MATERIALS AND METHODS: HeLa S3 cells were given a non-lethal heat treatment (41.5 degrees C, 4 h) and irradiated with 5 Gy X-irradiation. Alterations in cell-cycle distribution, intracellular cyclin B1 levels, and the yield of mitotic catastrophe were then measured and compared to heat-only and radiation-only groups, as a function of time following treatment. RESULTS: A greater accumulation of cells in S and G2 phases of the first cell-cycle post-treatment was observed in the combined heat and radiation groups, when compared with that observed following treatment with heat or radiation alone. Similarly, intracellular levels of cyclin B1 and the incidence of mitotic catastrophe were found to be greater in the combined treatment groups. CONCLUSIONS: This study provides further evidence that delays late in the cell cycle are implicated in increases in intracellular cyclin B1 levels and the subsequent development of mitotic catastrophe. Further, these data suggest a role for mitotic catastrophe occurring as a result of G2/M checkpoint abrogation in the process of thermal radiosensitization. PMID- 10716649 TI - Transgenics and psychopharmacology--introduction. PMID- 10716650 TI - Practical and theoretical issues in gene-targeting studies and their application to behaviour. AB - Some of the major practical and theoretical issues that are associated with gene targeting studies in mice are discussed. The availability of sufficient space to house the extensive breeding colonies associated with studies in gene-manipulated mice is an important logistical consideration that requires consideration at an early stage. A practical example is discussed which illustrates some of these issues. Problems associated with disease control and methods of maintaining the health status of valuable colonies are also outlined. Differences in the behavioural phenotype of inbred mouse strains pose important issues for study design and selection of host mouse lines. The results from studies exploring variations in the behavioural phenotype of six common inbred strains are briefly outlined. The impact of phenotypic variation on behavioural studies is considered and the implications for experimental design are discussed. PMID- 10716651 TI - Targeting genes and proteins in the analysis of learning and memory: caveats and future directions. AB - Gene targeting using homologous recombination in embryonic stem (ES) cells and transgenic approaches in general allow one to precisely manipulate single genes and investigate their in vivo function in the mouse. Geneticists argue that these techniques are superior to pharmacological approaches as they obviate the lack of highly specific pharmacological agents in the study of brain function and behavior. However, by now it has become clear that transgenic approaches also have some limitations. One problem is spatial and temporal specificity of the genetic manipulation. The other is the possibility that the introduced genetic alteration gives rise to complex, secondary phenotypic changes. This may be a disadvantage in the functional analysis of genes associated with learning and memory especially if the gene of interest plays roles in embryonic development of the brain as well as in adult neural function. Examples of such genes include, but are not limited to, those encoding neurotrophins, cell adhesion molecules, and protein kinases. Second generation gene targeting with inducible and cell type restricted knock-out, or transgenic approaches with inducible gene expression systems, will solve some problems. However, at present these strategies also suffer from difficulties inherent to the traditional knock-out. Several strategies alternative to transgenic approaches are also available. Antisense oligonucleotides, antibodies, or pharmacological agents may be used to manipulate molecular events at the transcription, translation, or protein function levels. I review these strategies briefly and suggest yet another approach: protein targeting with the use of recombinant immunoadhesins. I suggest that this latter approach has the specificity of gene targeting but lacks some of its disadvantages. PMID- 10716652 TI - Transgenic animal models for neuropharmacology. AB - The establishment of novel animal models using gene targeting and transgenic technology has opened a new area of neuropharmacological research. For the first time, it became possible to alter the expression of a gene in a specific cell type of an intact animal by either overexpression, inhibition or ablation. This review describes the technology and lists the relevant tools, such as reporter genes, suicide genes, immortalizing genes, and promoters, necessary for the targeted expression of these and other genes in specific cells of the central nervous system. In addition, the problem is discussed that the mouse is the species in which this technology is by far the most developed, while the rat has been used as the model species for neuropharmacology during the last century. PMID- 10716653 TI - From genotype to phenotype--behavior of the transgenic rat TGR(mRen2)27 as an example. AB - Transgenic techniques provide a tool to generate animals that differ from the wild-type by one or more genes, either by introducing foreign genes (transgenic animals) or by specific mutations of genes (knock-out animals). Most transgenic and knock-out animals are mice and not rats. The frequent use of rat models in the behavioral laboratory, however, will require the increasing application of transgenic techniques in this species. This paper reviews behavioral data from our laboratory as an example of characterizing the behavioral phenotype of a particular transgenic rat, the TGR(mRen2)27 rat. By describing the anxiogenic profile of this rat we also consider some problems associated with such an analysis, with the intention to raise issues that may also apply to studies of behavior in transgenic animals in general. PMID- 10716655 TI - Animal models of depression: utility for transgenic research. AB - The utility of available animal models of depression for transgenic research is reviewed. Criteria for usefulness are non-dependence on a mechanism of action, pharmacological validity, existence of genetic determinants, availability of a mouse version, procedural simplicity, and reproducibility. The following models are reviewed: behavioral despair, tail suspension, learned helplessness, chronic mild stress, olfactory bulbectomy, DRL behavior and conditioned place preference. It is concluded that the behavioral despair and tail suspension models satisfy the criteria most closely. On the other hand, despite its procedural complexity and poor reproducibility, the chronic mild stress model shows high promise for the future. PMID- 10716654 TI - Transgenic approaches to model Alzheimer's disease. AB - Two transgenic mouse lines were generated which express human APP751 containing familial Alzheimer's disease (AD) mutations in brain neurons. These mice develop pathological features reminiscent of AD. The degree of pathology depends on both expression levels and specific mutations. In mice with more advanced pathology (APP 23), typical plaques appear at six months which increase with age and are Congo Red positive at first detection. These congophilic plaques are accompanied by neuritic changes and dystrophic cholinergic fibers. Furthermore, inflammatory processes indicated by a massive glial reaction are apparent. Most notably, plaques are immunoreactive for hyperphosphorylated tau, reminiscent of early tau pathology. A quantitative analysis of degenerative changes by state-of-the-art unbiased stereological methods revealed a significant reduction in neuronal cell bodies of the CA1 field of the hippocampus when compared to controls. This reduction is directly related to plaque load. When subjected to analysis in the Morris water maze, 18 month old APP 23 mice show a significant increase in platform finding latency throughout the entire trial when compared to non transgenic littermates. PMID- 10716656 TI - Measurement of anxiety in transgenic mice. AB - A wide range of approaches has been used to study anxiety in mice. All presuppose that aversive stimuli, such as foot shock or novelty, induce a central state of fear, which can be quantified through specific behavioural and physiological measures. This review discusses the validity of the various approaches in terms of their similarity to different human anxiety disorders, their ability to detect compounds which modulate human anxiety, and their relevance to animal defensive processes. The most commonly used models of anxiety suitable for screening transgenic and knockout mice are discussed, with an emphasis placed on controlling for factors which could confound results. As all models used to date have limitations and no single paradigm adequately models all aspects of anxiety, this review recommends the use of a broad range of anxiety models in order to provide a comprehensive characterisation of the behavioural phenotype of transgenic mice. PMID- 10716657 TI - Oxidative phosphorylation disease diagnosis. AB - Although the mitochondrial (mtDNA) encodes only 13 polypeptide subunits of the oxidative phosphorylation (OXPHOS) enzymes, approximately 1,000 proteins are estimated to be necessary for proper OXPHOS function. Over the past ten years, a wide variety of adult and pediatric OXPHOS diseases were found to be caused by or associated with mtDNA mutations and nuclear DNA mutations. These advances enhanced the ability to definitively diagnose patients, develop management plans, and provide genetic counseling. However, in most individuals, diagnosing OXPHOS diseases is difficult and depends on assessing complex data derived from clinical, neuroradiologic, metabolic, biochemical, and pathologic evaluations. As understanding of nuclear OXPHOS genes grows, a more coherent approach to diagnosis, management, and treatment is likely to emerge. This article reviews major classes of OXPHOS diseases, a diagnostic algorithm, and recent advances in this complex field. PMID- 10716658 TI - Inherited peripheral neuropathy. AB - Hereditary disorders of the peripheral nerves constitute a group of frequently encountered neurological diseases. Charcot-Marie-Tooth neuropathy type 1 (CMT1) is genetically heterogeneous and characterized by demyelination with moderately to severely reduced nerve conduction velocities, absent muscle stretch reflexes and onion bulb formation. Genetic loci for CMT1 map to chromosome 17 (CMT1A), chromosome 1 (CMT1B), and another unknown autosome (CMT1C). CMT1A is most often associated with a tandem 1.5-megabase (Mb) duplication in chromosome 17p11.2-12, or in rare patients may result from a point mutation in the peripheral myelin protein-22 (PMP22) gene. CMT1 B result from point mutations in the myelin protein zero (Po or MPZ) gene. The molecular defect in CMT1 C is unknown. Mutations in the early growth response 2 gene (EGR2) are also associated with demyelinating neuropathy. Other rare forms of demyelinating peripheral neuropathies map to chromosome 8q, 10q, and 11q. X-linked Charcot-Marie-Tooth neuropathy (CMTX), which has clinical features similar to CMT1, is associated with mutations in the connexin32 gene. Charcot-Marie-Tooth neuropathy type 2 (CMT2) is characterized by normal or mildly reduced nerve conduction velocity with decreased amplitude and axonal loss without hypertrophic features. One form of CMT2 maps to chromosome 1 p36 (CMT2A), another to chromosome 3p (CMT2B) and another to 7p (CMT2D). Dejerine Sottas disease (DSD), also called hereditary motor and sensory neuropathy type III (HMSNIII), is a severe, infantile-onset demyelinating polyneuropathy that may be associated with point mutations in either the PMP22 gene or the Po gene and shares considerable clinical and pathological features with CMT1. Hereditary neuropathy with liability to pressure palsies (HNPP) is an autosomal dominant disorder that results in a recurrent, episodic demyelinating neuropathy. HNPP is associated with a 1.5-Mb deletion in chromosome 17p11.2-12 and results from reduced expression of the PMP22 gene. CMT1A and HNPP are reciprocal duplication/deletion syndromes originating from unequal crossover during germ cell meiosis. PMID- 10716659 TI - Ion channel diseases: episodic disorders of the nervous system. AB - Electrical excitability of skeletal and cardiac muscle cells and neurons results from a balance of inhibitory and excitatory influences. Ionic concentration gradients established by adenosine 5'-triphosphate (ATP)-dependent pumps can be maintained because the lipid bilayer is an extremely good insulator. Once ionic concentrations are established, movement of one or more ions down their respective concentration gradients can establish voltage differences across a membrane. The Nernst equation allows prediction of membrane potentials based on the particular ion involved and the concentration gradient for that ion in the cell. A large number of voltage-gated ion channels, ligand-gated channels, and transporters are involved in maintaining this balance. The specific channels and transporters involved differ in various cell types. In any case, normal membrane excitability is tightly regulated by the balance of these opposing influences. It is not surprising that the disruption of the balance of excitability of various cells might lead to neurological phenotypes. However, large changes in excitability of muscle or nerve may well be lethal. Therefore, nature may select against such major changes. A growing body of evidence suggests that subtle changes in some ion channels can lead to a slight increase in membrane excitability that results in a neurological phenotype. Interestingly, these phenotypes are frequently episodic. That is, under many circumstances, the nerve or muscle may be functioning properly; however, under certain circumstances, a precipitating event can lead to abnormal excitability resulting in one of any number of phenotypes discussed below. In this chapter, discussion will be focused on a number of monogenic disorders of the nervous system where episodic phenotypes are known to result from specific mutations of ion channels. The similarities between a large group of seemingly disparate disorders will be emphasized. Finally, some energy will be directed at developing the hypothesis that more subtle variations in proteins regula ting membrane excitability-though not causing a Mendelian disorder-may yield a predisposition to certain episodic phenomenon such as seizures and migraine headache. PMID- 10716660 TI - Molecular genetics of Alzheimer disease. AB - Epidemiological and individual case studies indicate that genetic factors play a significant role in the genesis of Alzheimer Disease (AD). To date, molecular genetic studies in families multiply affected with AD have identified three genes (Presenilin 1-PS1, Presenilin 2-PS2, and beta-amyloid precursor protein--betaAPP) associated with highly penetrant early onset AD, and one gene (Apolipoprotein E) associated with late onset AD. A fifth potential AD susceptibility locus has been mapped to a broad region of chromosome 12, but the responsible gene defect has not yet been identified. Case-control studies comparing the frequency of alleles in numerous other candidate genes have identified a number of additional potential AD genes. However, methodological difficulties and conflicting results in follow-up studies, make it unclear whether allelic variations in these genes are truly pathogenic. Nevertheless, analysis of the biochemical effects of mutations in PS1, PS2, betaAPP at least, suggest a common biochemical effect namely disturbances in the processing of betaAPP protein. In addition to utility in defining potential therapeutic targets, in some circumstances these genes can also potentially be used as adjunctives in clinical presymptomatic, symptomatic or pharmacogenomic diagnosis. PMID- 10716661 TI - Recent insights into the molecular pathogenesis of Huntington disease. AB - Huntington disease (HD) is a neurodegenerative disorder caused by a CAG repeat expansion in the HD gene resulting in expression of an uninterrupted polyglutamine stretch within the N-terminus of its protein product huntingtin (htt). In this article we review the clinical, genetic, and neuropathological features of HD and discuss recent insights into the pathogenesis of HD. Examining the role of CAG repeat size on age of onset and penetrance in HD using a refined database of human HD patients has provided further support for the importance of the CAG repeat in the pathogenesis of HD and information leading to a predictive model for the likelihood of being affected by a specific age for a particular CAG expansion. In a YAC transgenic mouse model that replicates key elements of the HD phenotype, the development of selective striatal neurodegeneration is coincident with cleavage of htt and translocation of the N-terminal htt fragment into the nucleus. We also review in vitro evidence that htt is a substrate for cleavage by a group of cysteine proteases involved in apoptotic death-the caspases, and that caspase cleavage of htt results in the generation of a toxic N-terminal fragment. Inhibiting caspase cleavage of huntingtin eliminates the toxicity of the mutant htt protein. These results suggest that cleavage of huntingtin resulting in production of a truncated N-terminal fragment may be a crucial step in the pathogenesis of Huntington disease and that inhibition of this process may be a potential therapeutic strategy for this currently untreatable disorder. PMID- 10716662 TI - Susceptibility genes in human epilepsy. AB - Major advances in the identification of genetic loci and genes that predispose individuals to epilepsy have been made in the last several years. Two main themes for human, idiopathic epilepsies are emerging; genetic, or locus heterogeneity is not uncommon, and the discovery that epilepsy susceptibility genes are voltage gated and ligand-gated ion channels. Knowledge that more than a single genetic locus is responsible for a single seizure type, along with a wide spectrum of disease mutations among families will complicate clinical, diagnostic issues. Disease gene identification, such as the two potassium ion channels (KCNQ2 and KCNQ3) for the two forms of benign familial neonatal seizures (BFNC) and the alpha4 subunit of the nicotinic receptor for autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE), however, should yield significant advances in drug discoveries. Understanding the primary defect in inherited epilepsies provides for specific protein and pathway targets for potential drug intervention. PMID- 10716663 TI - Genetic disorders of motor neurons. AB - Disorders of the motor neuron are etiologically and clinically heterogeneous and cause serious disability and death. Whereas mendelian inheritance can be demonstrated in a subset of these disorders, the genetic contribution to the sporadic forms of motor neuron degeneration are not well understood. In families with spinal muscular atrophy, Kennedy disease and amyotrophic lateral sclerosis, genetic linkage analysis and positional cloning have proven to be extremely productive. The genetics of these neurodegenerative disorders are reviewed. PMID- 10716664 TI - Dominantly inherited ataxias. AB - Molecular genetic studies in the past decade have demonstrated the enormous genetic heterogeneity among the dominantly inherited ataxias. Mutations at several distinct loci give rise to the progressive dominant ataxias and at least 2 different mutations cause episodic ataxias with dominant inheritance. The well established genotypes for progressive dominant ataxias have all involved expansions of repeated CAG sequences. Clinically these patients present with progressive cerebellar deficits as well as signs relating to pathology in other neural systems in a variable fashion. Some of these other signs serve as diagnostic clues to the underlying genotype, but the identification of the genotype from the clinical phenotype alone is usually difficult. The CAG expansions involved usually are unstable with intergenerational expansions as well as contractions of the repeat size. Phenotypic features such as age of onset and to a lesser extent disease progression rate and the presence of specific clinical signs depend on the CAG repeat size. Identification of the mutations has allowed precise genotypic diagnosis in several families allowing more accurate genetic counseling, including predictive testing of at risk individuals when sought. Also, increasing information about the gene products and their abnormal distributions in disease brain is rapidly giving rise to rational ideas about the pathogenesis of neuronal degeneration in these diseases and raising hope for meaningful treatment strategies. PMID- 10716666 TI - The intracellular target of butyrate's actions: HDAC or HDON'T? PMID- 10716665 TI - Is liver fibrosis reversible? PMID- 10716667 TI - Think cytokines before you drink. PMID- 10716668 TI - LKM antibody: getting in some target practice. PMID- 10716669 TI - Combination therapy of hepatitis B. PMID- 10716670 TI - Uteroglobin deficient mice-a novel animal model for IgA nephropathy? PMID- 10716671 TI - The human two domain trefoil protein, TFF2, is glycosylated in vivo in the stomach. AB - BACKGROUND: TFF2, a member of the trefoil factor family (TFF) of peptides, is a secreted protein of 106 amino acids that is expressed in mucous neck cells of the fundus and glands at the base of the antrum in normal human stomach. TFF2 is also detected at high concentrations around sites of ulceration. It is protective against mucosal damaging agents and stimulates cell motility. AIMS: To measure the expression of TFF2 in normal human stomach and its secretion into gastric juice. METHODS: TFF2 cDNA was amplified by reverse transcription polymerase chain reaction from gastric mucosa and sequenced. Gastric juice or cytosol, prepared from gastric mucosa, was obtained from individuals with macroscopically normal stomachs. TFF2 concentrations were measured by quantitative western transfer analysis. RESULTS: Sequencing of TFF2 cDNA revealed a single amino acid change from the published sequence. Significant amounts of 12 kDa TFF2 were detected in human gastric juice. Larger quantities of a protein of higher apparent molecular mass were also detected. This was shown to be N-glycosylated TFF2 using the endoglycosidase, peptide-N-Gycosidase F. The majority of TFF2 in normal gastric mucosa was also glycosylated. CONCLUSIONS: Human TFF2 is glycosylated via an N linkage, presumably on Asn(15) which forms part of the single consensus site for N-glycosylation in human TFF2. The glycosylation may be of functional significance. Future studies of human TFF2 should use antibodies raised against the correct amino acid sequence. Biological studies should be performed with recombinant protein of the correct sequence, and the biological consequences of glycosylation investigated. PMID- 10716673 TI - Detection of upper gastrointestinal cancer in patients taking antisecretory therapy prior to gastroscopy. AB - BACKGROUND: The incidence of early gastric cancer has not increased despite better access to endoscopic facilities for general practitioners. Many patients receive a course of symptomatic treatment while waiting for gastroscopy. AIMS: To ascertain the effect of antisecretory therapy on the diagnostic process and findings for patients with upper gastrointestinal cancer. METHODS: A consecutive case study survey of the primary care records of 133 patients who had died of upper gastrointestinal cancer during 1995-97 in the South Tees health district in the north-east of England (population 300 000). RESULTS: From the 133 patients identified, 116 had died from adenocarcinoma of the oesophagus (31) or stomach (85). Failure to reach the diagnosis of cancer at the index gastroscopy was associated with prior acid suppression therapy. Only one of 54 patients on no treatment or antacids alone was erroneously diagnosed as suffering from benign disease, whereas 22 of 62 patients treated with acid suppression were diagnosed as suffering from benign disease but at varying times later turned out to have adenocarcinoma. Twenty of 45 patients taking a proton pump inhibitor had a delayed diagnosis compared with two of 17 taking an H(2) receptor antagonist. The commonest lesion seen at index gastroscopy in those in whom the diagnosis was initially missed was gastric ulcer. Healing occurred in six patients taking a proton pump inhibitor, despite their later diagnosis of malignancy. CONCLUSIONS: The treatment of dyspeptic symptoms with acid suppression prior to gastroscopy masks and delays the detection of gastric and oesophageal adenocarcinoma on endoscopy in one third of patients. PMID- 10716672 TI - Atrophic gastritis and Helicobacter pylori infection in outpatients referred for gastroscopy. AB - BACKGROUND: Atrophic gastritis has been shown to be one of the long term sequelae of Helicobacter pylori infection. AIMS: To determine the prevalence of atrophic gastritis in outpatients, to study the accuracy of serological methods for revealing atrophy, and to define the association of H pylori infection with atrophic gastritis in these patients. PATIENTS/METHODS: A total of 207 consecutive outpatients referred for gastroscopy were included. Biopsy specimens from the antrum and corpus were assessed histologically according to the Sydney system. Serum samples were studied for H pylori IgG and IgA antibodies by enzyme immunoassay, CagA antibodies by immunoblot, pepsinogen I by an immunoenzymometric assay, gastrin by radioimmunoassay, and parietal cell antibodies by indirect immunofluorescence. RESULTS: Histological examination revealed atrophic gastritis in 52 (25%) of 207 patients. H pylori and CagA antibodies were strongly associated with atrophic antral gastritis but poorly associated with atrophic corpus gastritis. Low serum pepsinogen I was the most sensitive and specific indicator of moderate and severe atrophic corpus gastritis. All six patients with moderate atrophic corpus gastritis had H pylori infection but eight of 10 patients with severe atrophic corpus had increased parietal cell antibodies and nine had no signs of H pylori infection. CONCLUSIONS: Atrophic antral gastritis was strongly associated with CagA positive H pylori infection. Severe atrophic corpus gastritis was not determined by H pylori tests but low serum pepsinogen I, high gastrin, and parietal cell antibodies may be valuable in detecting these changes. PMID- 10716674 TI - Influence of sumatriptan on gastric fundus tone and on the perception of gastric distension in man. AB - BACKGROUND: In animals, activation of 5-HT(1) like receptors causes a relaxation of the gastric fundus through the activation of intrinsic inhibitory neurones. AIMS: To investigate the effect of sumatriptan, an agonist at enteric neuronal 5 HT(1) receptors, on fasting fundus tone and sensitivity to gastric distension in man. METHODS: A gastric barostat was used to study the effect of placebo and sumatriptan, 6 mg subcutaneously, on basal fundic tone in healthy subjects. In addition, stepwise isobaric and isovolumetric gastric distensions were performed and perception was measured before and after the administration of placebo and sumatriptan. RESULTS: Placebo had no significant effects on gastric tone and on perception. Sumatriptan induced an immediate relaxation of the gastric fundus, reflected by an intragastric volume increase of 209 (39) ml (p<0.0005). After sumatriptan, intragastric pressures at the thresholds for perception or discomfort were not significantly altered. However, the intragastric volumes and the corresponding calculated wall tensions at perception and discomfort thresholds were significantly increased. CONCLUSIONS: Administration of the 5 HT(1) receptor agonist sumatriptan induces a relaxation of the gastric fundus in man, allowing larger intragastric volumes before thresholds for perception or discomfort are reached. The effects of sumatriptan on the gastric fundus may have therapeutic potential in the treatment of patients with functional dyspepsia. PMID- 10716675 TI - The 5-HT(3) receptor antagonist alosetron inhibits the colorectal distention induced depressor response and spinal c-fos expression in the anaesthetised rat. AB - BACKGROUND: Noxious intestinal distention elicits a reflex depressor response in the sodium pentobarbitone anaesthetised rat, which can be used as an index of visceral nociception. 5-HT(3) receptor antagonists inhibit this reflex. Repeated colorectal distention (CRD) induces Fos like immunoreactivity (Fos-LI) in the rat spinal cord. AIMS: To examine the effect of the 5-HT(3) receptor antagonist alosetron on the depressor response to CRD, and on Fos expression in the lumbosacral spinal cord. METHODS: Male rats were anaesthetised with sodium pentobarbitone, and mean arterial blood pressure monitored during repeated colorectal balloon inflation before and after treatment with alosetron or saline. Rats anaesthetised with urethane and treated with alosetron or saline underwent a repeated CRD paradigm, after which the lumbosacral spinal cord was removed and processed for visualisation of Fos-LI. RESULTS: CRD elicited reproducible, volume dependent falls in arterial blood pressure, and repeated distention-effect curves were constructed. Alosetron (1-100 microg/kg intravenously) inhibited the depressor response to CRD in a dose related manner, with an ID(50) value of 3.0 microg/kg. Following repeated CRD, numbers of Fos-LI neurones were significantly increased to 1246 (total in 12 sections at 120 microm intervals from L6 to S1) compared with 49 in sham distended animals. Pretreatment with alosetron (100 microg/kg) significantly reduced numbers of Fos-LI neurones to 479.8. CONCLUSION: The 5-HT(3) receptor antagonist alosetron inhibits the depressor response to CRD in a potent and dose dependent manner. It also inhibits CRD induced Fos-LI in the spinal cord. These results suggest that 5-HT(3) receptors are involved in visceral nociceptive transmission, perhaps located on primary afferent or spinal neurones. PMID- 10716676 TI - Clostridium difficile toxin A excites enteric neurones and suppresses sympathetic neurotransmission in the guinea pig. AB - BACKGROUND AND AIMS: Evidence suggests that the intestinal actions of Clostridium difficile toxin A-stimulation of secretion and motility, and an acute inflammatory response-have a neurally mediated component. METHODS: Direct intracellular electrophysiological recording of electrical and synaptic behaviour in enteric neurones was performed in the submucous plexus of guinea pig small intestine during exposure to the toxin. RESULTS: Application of toxin A affected both the electrical behaviour of the neuronal cell bodies and inhibitory noradrenergic neurotransmission to the cell bodies. Altered electrical behaviour included depolarisation and increased excitability. Tetrodotoxin or a histamine H(2) receptor antagonist did not affect the depolarisation evoked by toxin A. Failure of the histamine antagonist to suppress the actions of toxin A is evidence that its actions were not mediated by degranulation of intramural mast cells. The action of toxin A on neurotransmission was suppression of inhibitory postsynaptic potentials evoked in the neuronal cell bodies by stimulation of sympathetic nerve fibres that synapsed with the cell bodies. The inhibitory postsynaptic potentials were mediated by norepinephrine (noradrenaline) acting at postsynaptic alpha adrenoceptors on the cell bodies. Hyperpolarising responses evoked in the cell bodies by micropressure application of norepinephrine were unaffected by toxin A. This fulfils criteria for a presynaptic inhibitory action of toxin A to suppress release of norepinephrine from sympathetic postganglionic axons. CONCLUSIONS: Results suggest that the neural component of the action of toxin A involves both direct excitation of enteric neurones and suppression of norepinephrine release from postganglionic sympathetic nerve fibres in the enteric nervous system. PMID- 10716677 TI - Measurement of in vivo rectal mucosal cytokine and eicosanoid production in ulcerative colitis using filter paper. AB - BACKGROUND: Excessive mucosal generation of cytokines and eicosanoids has been reported in vitro in ulcerative colitis (UC) using traumatising biopsy techniques, and in vivo using time consuming rectal dialysis. AIMS: To validate a simple filter paper technique to profile rectal mucosal production of cytokines and eicosanoids in vivo in patients with UC compared with controls. PATIENTS: Forty one patients with UC (21 with active disease) and 16 controls were studied. METHODS: In vitro, recovery of known concentrations of cytokine or mediator applied to filter papers was measured by ELISA following incubation in buffer. In vivo, patients and controls had filter papers apposed to the rectal mucosa briefly through a rigid sigmoidoscope. Filter papers were then incubated prior to assay by ELISA. RESULTS: In vitro validation studies showed that the filter paper technique could be used to measure mucosal release of interleukin-1beta (IL 1beta), tumour necrosis factor alpha (TNF-alpha), thromboxane B(2) (TXB(2)), and prostaglandin E(2) (PGE(2)), but not interferon gamma (IFN-gamma). Mucosal release of IL-1beta, TNF-alpha, TXB(2) and PGE(2) were significantly increased in active UC (p=0.001) and correlated directly with disease activity (p=0.02). CONCLUSIONS: The filter paper technique confirmed increased rectal mucosal release of cytokines and eicosanoids in UC, in proportion to disease activity. The simplicity, safety and speed of the technique make it a practicable option for use in the outpatient clinic to study the pathogenesis of inflammatory bowel disease, and potentially its response to treatment. PMID- 10716678 TI - Butyrate and glucose metabolism by colonocytes in experimental colitis in mice. AB - BACKGROUND/AIMS: Impaired colonocyte metabolism of butyrate has been implicated in the aetiopathogenesis of ulcerative colitis. Colonocyte butyrate metabolism was investigated in experimental colitis in mice. METHODS: Colitis was induced in Swiss outbred white mice by oral administration of 4% dextran sulphate sodium (DSS). Colonocytes isolated from colitic and normal control mice were incubated with [(14)C]butyrate or glucose, and production of (14)CO(2), as well as of intermediate metabolites (acetoacetate, beta-hydroxybutyrate and lactate), was measured. The effect of different substrate concentrations on oxidation was also examined. RESULTS: Butyrate oxidation (micromol/h per mg protein; mean (SEM)) was significantly reduced in DSS colitis, values on day 7 of DSS administration being 0.177 (0.007) compared with 0.406 (0.035) for control animals (p<0.001). Glucose oxidation (micromol/h per mg protein; mean (SEM)) on day 7 of DSS administration was significantly higher than in controls (0.06 (0.006) v 0.027 (0.004), p<0.001). Production of beta-hydroxybutyrate was decreased and production of lactate increased in DSS colitis compared with controls. Increasing butyrate concentration from 10 to 80 mM enhanced oxidation in DSS colitis (0.036 (0.002) to 0.285 (0.040), p<0.001), although it continued to remain lower than in controls. Surface and crypt epithelial cells showed similar ratios of butyrate to glucose oxidation. When 1 mM DSS was added to normal colonocytes in vitro, it did not alter butyrate oxidation. The initial histological lesion of DSS administration was very patchy and involved crypt cells. Abnormal butyrate oxidation became apparent only after six days of DSS administration, at which time histological abnormalities were more widespread. CONCLUSIONS: Colonocyte metabolism of butyrate, but not of glucose, is impaired in DSS colitis, and may be important in pathophysiology. Histological abnormalities preceded measurable defects in butyrate oxidation. PMID- 10716679 TI - A decision analysis of surveillance for colorectal cancer in ulcerative colitis. AB - BACKGROUND: Patients with long standing, extensive ulcerative colitis have an increased risk of developing colorectal cancer. AIMS: To assess the feasibility of surveillance colonoscopy in preventing death from colorectal cancer. PATIENTS: A hypothetical cohort of patients with chronic ulcerative colitis. METHODS: The benefits of life years saved were weighted against the costs of biannual colonoscopy and proctocolectomy, and the terminal care of patients dying from colorectal cancer. Two separate Markov processes were modelled to compare the cost-benefit relation in patients with or without surveillance. The cumulative probability of developing colorectal cancer served as a threshold to determine which of the two management strategies is associated with a larger net benefit. RESULTS: If the cumulative probability of colorectal cancer exceeds a threshold value of 27%, surveillance becomes more beneficial than no surveillance. The threshold is only slightly smaller than the actual cumulative cancer rate of 30%. Variations of the assumptions built into the model can raise the threshold above or lower it far below the actual rate. If several of the assumptions are varied jointly, even small changes can lead to extreme threshold values. CONCLUSIONS: It is not possible to prove that frequent colonoscopies scheduled at regular intervals are an effective means to manage the increased risk of colorectal cancer associated with ulcerative colitis. PMID- 10716680 TI - Butyrate and trichostatin A effects on the proliferation/differentiation of human intestinal epithelial cells: induction of cyclin D3 and p21 expression. AB - BACKGROUND: Sodium butyrate, a product of colonic bacterial fermentation, is able to inhibit cell proliferation and to stimulate cell differentiation of colonic epithelial cell lines. It has been proposed that these cellular effects could be linked to its ability to cause hyperacetylation of histone through the inhibition of histone deacetylase. AIM: To analyse the molecular mechanisms of butyrate action on cell proliferation/differentiation and to compare them with those of trichostatin A, a well known inhibitor of histone deacetylase. METHODS: HT-29 cells were grown in the absence or presence of butyrate or trichostatin A. Cell proliferation and cell cycle distribution were studied after DNA staining by crystal violet and propidium iodide respectively. Cell cycle regulatory proteins were studied by western blot and reverse transcription-polymerase chain reaction. Cell differentiation was followed by measuring brush border enzyme activities. Histone acetylation was studied by acid/urea/Triton acrylamide gel electrophoresis. RESULTS: Butyrate blocked cells mainly in the G(1) phase of the cell cycle, whereas trichostatin A was inhibitory in both G(1) and G(2) phases. Butyrate inhibited the mRNA expression of cyclin D1 without affecting its protein expression and stimulated the protein expression of cyclin D3 without affecting its mRNA expression. Trichostatin A showed similar effects on cyclin D1 and D3. Butyrate and trichostatin A stimulated p21 expression both at the mRNA and protein levels, whereas their effects on the expression of cyclin dependent kinases were slightly different. Moreover, butyrate strongly stimulated the activity of alkaline phosphatase and dipeptidyl peptidase IV, whereas trichostatin A had no effect. Finally, a six hour exposure to butyrate or trichostatin A induced histone H4 hyperacetylation. At 15 and 24 hours, histone H4 remained hyperacetylated in the presence of butyrate, whereas it returned to control levels in the presence of trichostatin A. CONCLUSIONS: The data may explain how butyrate acts on cell proliferation/differentiation, and they show that trichostatin A does not reproduce every effect of butyrate, mainly because of its shorter half life. PMID- 10716681 TI - Surface hydrophobicity of the rat colonic mucosa is a defensive barrier against macromolecules and toxins. AB - BACKGROUND: Mucosal surface hydrophobicity is a key factor of the gastric acid defence barrier. In the colon, surface hydrophobicity is high but its biological function remains unexplored. AIMS: To investigate the functional changes of the barrier due to removal of the surface active phospholipid layer by a detergent, or to reinforcement of the surface active phospholipid by local application of a suspension of lipids. METHODS: Surface hydrophobicity (contact angle measurement), colonic permeability (lumen to blood clearance of mannitol and dextran), and mucosal resistance against luminal aggression (distal colitis induced by dextran sodium sulphate, DSS) were investigated in three study groups: (a) rats pretreated with a detergent (Brij 35) known to remove surfactant lipids; (b) rats pretreated with a suspension of surface active lipids (tripalmitin and dipalmitoyl-phosphatidylcholine); and (c) control rats pretreated with the corresponding vehicles. RESULTS: In controls, surface hydrophobicity was low on the caecal mucosa and high in colon and rectum. Detergent treatment reduced surface hydrophobicity, and increased colonic permeability to mannitol and dextran. Conversely, treatment with lipids increased surface hydrophobicity, and reduced colonic permeability. Administration of DSS induced a progressive loss of colonic surface hydrophobicity, and an increase in permeability to mannitol and dextran. Detergent treatment increased susceptibility to epithelial damage and mucosal inflammation by DSS. Treatment with lipids reduced susceptibility to DSS colitis. CONCLUSION: Colonic surface hydrophobicity modulates permeability to hydrophilic molecules and protects against toxins. PMID- 10716682 TI - Long term efficacy, safety, and tolerabilitity of low daily doses of isosmotic polyethylene glycol electrolyte balanced solution (PMF-100) in the treatment of functional chronic constipation. AB - AIMS: To assess the long term therapeutic effectiveness, safety, and tolerability of low daily doses of isosmotic PEG electrolyte solutions (PMF-100) administered for a six month period for the treatment of functional constipation, in a double blind, placebo controlled, parallel group study. METHODS: After an initial four week run in period with PMF-100 (250 ml twice daily; PEG 14.6 g twice daily), 70 patients suffering from chronic constipation (58 females, aged 42 (15) years) with normalised bowel frequency (>3 bowel movements (bm)/week) were randomly allocated to receive either PMF-100 or placebo, contained in sachets (one sachet in 250 ml of water twice daily) for 20 weeks. Patients were assessed at four week intervals, and reported frequency and modality of evacuation, laxative use, and relevant symptoms on a diary card. At weeks 1, 12, and 24, a physical examination and laboratory tests were performed. RESULTS: Complete remission of constipation was reported by a significantly (p<0.01) higher number of patients treated with PMF-100 compared with placebo at each four week visit. At the end of the study, 77% of the PMF-100 group and 20% of the placebo group were asymptomatic. Compared with placebo, patients treated with PMF-100 reported hard/pellety stools and straining at defecation less frequently, a significantly higher bowel frequency (week 12: 7. 4 (3.1) v 4.3 (2.5) bm/week, 95% CI 1.64, 4.42; week 24: 7.4 (3.2) v 5.4 (2.1) bm/week, 95% CI 0.13,3.93), reduced consumption of laxative/four weeks (week 12: 0.7 (2.7) v 2.2 (3.3), 95% CI -2.29, 0. 03; week 24: 0.2 (0.8) v 1.4 (2), 95% CI -2.07, -0.023), reduced mean number of sachets used (week 12: 33 (13) v 43 (12), 95% CI -17. 24, 4.56; week 24: 33 (13) v 44 (12), 95% CI -19.68, 2.24), and reduced number of drop outs for therapy failure (16 v 3; p<0.005). Adverse events, physical findings, laboratory values, palatability, and overall tolerance of the solutions did not differ between groups. CONCLUSIONS: Administration of small daily doses of isosmotic PEG electrolyte balanced solutions was effective over a six month period for the treatment of functional constipation. A mean daily dose of approximately 300 ml of PEG solution (PEG 17.52 g) appeared to be safe, well tolerated, and devoid of significant side effects. PMID- 10716683 TI - Sodium handling in patients with well compensated cirrhosis is dependent on the severity of liver disease and portal pressure. AB - BACKGROUND AND AIMS: To test the contribution of portal pressure gradient (PPG) and neurohumoral factors to sodium handling in cirrhotic patients without ascites, by comparing preascitic cirrhotic patients with patients with transjugular intrahepatic portosystemic stent shunt (TIPSS) and previous ascites. PATIENTS: Ten patients with TIPSS and 10 preascitic cirrhotic patients. METHODS: Changes in glomerular filtration, renal plasma flow, urinary sodium excretion (U(Na)V), and neurohumoral factors were measured before and for two hours after infusion of one litre of 0. 9% saline over one hour. RESULTS: Glomerular filtration rate and renal plasma flow were significantly higher in patients with TIPSS compared with preascitic cirrhotic patients. Following saline infusion both parameters increased significantly; this increase was significantly greater in patients with TIPSS. U(Na)V increased significantly in both groups following saline infusion. The increase in U(Na)V was significantly greater in the TIPSS group. Plasma renin activity and angiotensin II decreased significantly in both groups. Basal U(Na)V was independently correlated with angiotensin II concentration and PPG and the change in U(Na)V correlated with the PPG. CONCLUSIONS: Results suggest that patients with advanced liver disease and low portal pressure handle sodium as well as patients with compensated liver disease and high portal pressure. These results are consistent with the notion that in addition to peripheral vasodilatation and severity of liver disease, the severity of portal hypertension contributes to the abnormalities of sodium retention in cirrhosis. PMID- 10716684 TI - Deficiency of natural anticoagulant proteins C, S, and antithrombin in portal vein thrombosis: a secondary phenomenon? AB - BACKGROUND: Hereditary deficiencies of natural anticoagulant proteins are implicated in the pathogenesis of portal vein thrombosis (PVT). Secondary deficiencies of these proteins have also been reported in PVT, making interpretation of concentrations difficult. AIMS: To characterise the coagulation profiles in adult patients with PVT and to investigate the possible mechanisms of natural anticoagulant protein deficiency. PATIENTS: Twenty nine adult patients with portal hypertension caused by PVT, and normal biochemical liver function tests. METHODS: Routine coagulation profiles and concentrations of proteins C, S, and antithrombin were measured; where indicated, corresponding concentrations in parents were also measured. Synchronous peripheral and hepatic or splenic vein concentrations were compared in seven patients undergoing interventional procedures, as were peripheral concentrations before and after shunt surgery in three patients. RESULTS: Deficiencies of one or more of the natural anticoagulant proteins occurred in 18 patients (62%), with six patients having combined deficiency of all three proteins. There were strong correlations between prothrombin and partial thromboplastin time ratios and concentrations of natural anticoagulant proteins. Family studies in nine cases of anticoagulant protein deficiency revealed possible hereditary deficiency in only three cases, and significantly lower concentrations of anticoagulant proteins in all PVT cases compared with parents. Levels of anticoagulant proteins tended to be lower in hepatic veins but higher in splenic veins compared with peripheral vein concentrations. Peripheral concentrations decreased after shunt surgery. CONCLUSIONS: Deficiency of natural anticoagulant proteins is common in PVT and is probably a secondary phenomenon in most cases, occurring as part of a global disturbance of coagulation variables. The mechanism for this remains unclear but may result from a combination of reduced hepatic blood flow and portosystemic shunting itself. PMID- 10716685 TI - Interleukin 10 promoter region polymorphisms and susceptibility to advanced alcoholic liver disease. AB - BACKGROUND: The factors determining why less than 10% of heavy drinkers develop advanced alcoholic liver disease (ALD) remain elusive, although genetic factors may be important. Interleukin 10 (IL-10) is an important cytokine with anti inflammatory, anti-immune, and antifibrotic functions. Several polymorphisms have been identified in the IL-10 promoter and recent evidence suggests that some of these may have functional effects on IL-10 secretion. AIMS: To test the hypothesis that IL-10 promoter region polymorphisms are associated with susceptibility to ALD. METHODS: The allele frequencies for the two single base pair substitutions at positions -627 (C-->A) and -1117 (A-->G) in the IL-10 promoter were determined in 287 heavy drinkers with biopsy proved advanced ALD, 107 heavy drinkers with no evidence of liver disease or steatosis only on biopsy, and 227 local healthy volunteers. RESULTS: At position -627, 50% of patients with advanced ALD had a least one A allele compared with 33% of controls (p<0.0001) and 34% of drinkers with no or mild disease (p=0.017). At position -1117, the slight excess of the A allele in drinkers with advanced disease was because of linkage disequilibrium between the A alleles at the two sites. CONCLUSIONS: Among heavy drinkers, possession of the A allele at position -627 in the IL-10 promoter is associated with an increased risk of advanced liver disease. This is consistent with recent functional data that the -627*A allele is associated with low IL-10 expression which will favour inflammatory, immune mediated, and profibrotic mechanisms of alcohol related liver injury. PMID- 10716686 TI - Increased availability of central benzodiazepine receptors in patients with chronic hepatic encephalopathy and alcohol related cirrhosis. AB - BACKGROUND/AIMS: To measure cerebral benzodiazepine receptor binding using (11)C flumazenil positron emission tomography in patients with stable chronic hepatic encephalopathy, who were also characterised by proton magnetic resonance spectroscopy. METHODS: Six abstinent patients of mean age 61 years with alcohol related cirrhosis and grade I-II hepatic encephalopathy and 11 matched healthy volunteers were studied. Each patient's encephalopathy was defined according to clinical, psychometric, electroencephalographic, and magnetic resonance spectroscopy criteria. Using positron emission tomography, the brain volume of distribution of (11)C-flumazenil was obtained; this reflects benzodiazepine receptor availability. Proton magnetic resonance spectra were acquired at 1.5 T using a multivoxel technique; peak area ratios were calculated for choline, glutamine/glutamate, N-acetylaspartate, and creatine resonances. RESULTS: The mean volume of distribution of (11)C-flumazenil was significantly higher in the cortex, cerebellum, and the basal ganglia in the patients compared with controls (p<0.001). In the patient group, the mean glutamine/glutamate to creatine ratio was significantly increased and the mean choline to creatine ratio was significantly decreased in all brain areas, compared with healthy volunteers. However, the N-acetylaspartate to creatine ratio was unchanged compared with controls. CONCLUSIONS: The spectroscopy results reflect the cerebral metabolic derangement associated with hepatic encephalopathy. Stable grade I-II chronic hepatic encephalopathy in alcohol related cirrhosis may be associated with increased cerebral benzodiazepine receptor availability. However, a direct effect of previous chronic exposure to alcohol cannot be excluded. PMID- 10716687 TI - Liver/kidney microsomal antibody type 1 targets CYP2D6 on hepatocyte plasma membrane. AB - BACKGROUND: Liver/kidney microsomal antibody type 1 (LKM1) is the marker of type 2 autoimmune hepatitis (AIH) and is detected in up to 6% of patients with hepatitis C virus (HCV) infection. It recognises linear and conformational epitopes of cytochrome P450IID6 (CYP2D6) and may have liver damaging activity, provided that CYP2D6 is accessible to effector mechanisms of autoimmune attack. METHODS: The presence of LKM1 in the plasma membrane was investigated by indirect immunofluorescence and confocal laser microscopy of isolated rat hepatocytes probed with 10 LKM1 positive sera (five from patients with AIH and five from patients with chronic HCV infection) and a rabbit polyclonal anti-CYP2D6 serum. RESULTS: Serum from both types of patient stained the plasma membrane of non permeabilised cells, where the fluorescent signal could be visualised as discrete clumps. Conversely, permeabilised hepatocytes showed diffuse submembranous/cytoplasmic staining. Adsorption with recombinant CYP2D6 substantially reduced plasma membrane staining and LKM1 immunoblot reactivity. Plasma membrane staining of LKM1 colocalised with that of anti-CYP2D6. Immunoprecipitation experiments showed that a single 50 kDa protein recognised by anti-CYP2D6 can be isolated from the plasma membrane of intact hepatocytes. CONCLUSIONS: AIH and HCV related LKM1 recognise CYP2D6 exposed on the plasma membrane of isolated hepatocytes. This observation supports the notion that anti CYP2D6 autoreactivity may be involved in the pathogenesis of liver damage. PMID- 10716688 TI - Lamivudine and alpha interferon combination treatment of patients with chronic hepatitis B infection: a randomised trial. AB - BACKGROUND, AIM, AND METHODS: Alpha interferon is the generally approved therapy for HBe antigen positive patients with chronic hepatitis B, but its efficacy is limited. Lamivudine is a new oral nucleoside analogue which potently inhibits hepatitis B virus (HBV) DNA replication. To investigate the possibility of an additive effect of interferon-lamivudine combination therapy compared with interferon or lamivudine monotherapy, we conducted a randomised controlled trial in 230 predominantly Caucasian patients with hepatitis B e antigen (HBeAg) and HBV DNA positive chronic hepatitis B. Previously untreated patients were randomised to receive: combination therapy of lamivudine 100 mg daily with alpha interferon 10 million units three times weekly for 16 weeks after pretreatment with lamivudine for eight weeks (n=75); alpha interferon 10 million units three times weekly for 16 weeks (n=69); or lamivudine 100 mg daily for 52 weeks (n=82). The primary efficacy end point was the HBeAg seroconversion rate at week 52 (loss of HBeAg, development of antibodies to HBeAg and undetectable HBV DNA). RESULTS: The HBeAg seroconversion rate at week 52 was 29% for the combination therapy, 19% for interferon monotherapy, and 18% for lamivudine monotherapy (p=0.12 and p=0.10, respectively, for comparison of the combination therapy with interferon or lamivudine monotherapy). The HBeAg seroconversion rates at week 52 for the combination therapy and lamivudine monotherapy were significantly different in the per protocol analysis (36% (20/56) v 19% (13/70), respectively; p=0.02). The effect of combining lamivudine and interferon appeared to be most useful in patients with moderately elevated alanine aminotransferase levels at baseline. Adverse events with the combination therapy were similar to interferon monotherapy; patients receiving lamivudine monotherapy had significantly fewer adverse events. CONCLUSIONS: HBeAg seroconversion rates at one year were similar for lamivudine monotherapy (52 weeks) and standard alpha interferon therapy (16 weeks). The combination of lamivudine and interferon appeared to increase the HBeAg seroconversion rate, particularly in patients with moderately elevated baseline aminotransferase levels. The potential benefit of combining lamivudine and interferon should be investigated further in studies with different regimens of combination therapy. PMID- 10716689 TI - Gallstone disease in Peruvian coastal natives and highland migrants. AB - BACKGROUND: In a previous study, we found that gallstones were a common occurrence in the high altitude villages of the Peruvian Andes. AIMS: To determine if high altitude (> or = 1500 m) is a contributing risk factor for gallstone disease. METHODS: We conducted a cross sectional study in a periurban community in Lima, Peru, and compared the prevalence of gallstone disease between coastal natives, highland (Sierra) natives and Sierra natives who had migrated to the coast. We also compared the prevalence rates from this study with those from a previous study conducted at high altitude. We examined 1534 subjects >15 years of age for gallstone disease. Subjects were interviewed for the presence or absence of risk factors. RESULTS: Gallstone disease was more common in females (16.1 cases per 100, 95% CI 13.8-18.2) than in males (10.7 per 100, 95% CI 8.0 13.4). Females had a greater risk of gallstone disease, especially if they had used oral contraception and/or had four or more children. The age adjusted prevalence was not significantly different between coastal natives, Sierra migrants, and Andean villagers. The prevalence of gallstone disease was not associated with time since migration or with having native Sierra parents. After adjusting for other risk factors, Sierra natives who migrated to the coast had a lower prevalence of gallstone disease than coastal natives (odds ratio 0.74, 95% CI 0.58-0.94). CONCLUSIONS: This study indicates that high altitude is not a positive risk factor for gallstone disease and confirms that this disease is common in Peruvians, which may be attributable to Peruvian-Indian ethnicity. PMID- 10716690 TI - Intramucosal adenocarcinoma arising under squamous re-epithelialisation of Barrett's oesophagus. AB - BACKGROUND: Eradication of Barrett's mucosa by thermal or photoablation combined with high doses of proton pump inhibitors is a potentially attractive strategy in the management of this preneoplastic condition. However, major concerns of this method are the persistence of residual metaplastic glands beneath the new squamous epithelium and the absence of any knowledge of its impact on long term outcome. CASE REPORT: The case of an intramucosal adenocarcinoma diagnosed 18 months after apparently complete squamous re-epithelialisation achieved using argon plasma coagulation and high dose omeprazole (40 mg/daily) is reported in a 68 year old patient presenting initially with a Barrett's oesophagus without dysplasia. Intramucosal adenocarcinoma was located under the new squamous layer and presented as a bulging area covered by the squamous epithelium. It probably originates from residual metaplastic glands after therapy although a pre-existing tumour cannot be definitely excluded. CONCLUSION: This observation might question future application of this experimental endotherapy in non-dysplastic Barrett's oesophagus. It suggests that the residual glands might still be premalignant and that the early diagnosis of neoplastic changes might be compromised by the squamous re-epithelialisation. PMID- 10716691 TI - TIPSS 10 years on. PMID- 10716693 TI - Tcl1 enhances Akt kinase activity and mediates its nuclear translocation. AB - The TCL1 oncogene at 14q32.1 is involved in the development of human mature T cell leukemia. The mechanism of action of Tcl1 is unknown. Because the virus containing the v-akt oncogene causes T-cell lymphoma in mice and Akt is a key player in transduction of antiapoptotic and proliferative signals in T-cells, we investigated whether Akt and Tcl1 function in the same pathway. Coimmunoprecipitation experiments showed that endogenous Akt1 and Tcl1 physically interact in the T-cell leukemia cell line SupT11; both proteins also interact when cotransfected into 293 cells. Using several AKT1 constructs in cotransfection experiments, we determined that this interaction occurs through the pleckstrin homology domain of the Akt1 protein. We further demonstrated that, in 293 cells transfected with TCL1, the endogenous Akt1 bound to Tcl1 is 5-10 times more active compared with Akt1 not bound to Tcl1. The intracellular localization of Tcl1 and Akt1 in mouse fibroblasts was investigated by immunofluorescence. When transfected alone, Akt1 was found only in cytoplasm whereas Tcl1 was localized in the cytoplasm and in the nucleus. Interestingly, Akt1 was also found in the nucleus when AKT1 was cotransfected with TCL1, suggesting that Tcl1 promotes the transport of Akt1 to the nucleus. These findings were supported by the intracellular localization of Akt1 or Tcl1 when Tcl1 or Akt1, respectively, were confined to the specific cellular compartments. Thus, we demonstrate that Tcl1 is a cofactor of Akt1 that enhances Akt1 kinase activity and promotes its nuclear transport. PMID- 10716694 TI - Design and properties of human D-amino acid oxidase with covalently attached flavin. AB - An "artificial flavinylation" approach was developed to replace a native noncovalent flavin prosthetic group with a covalently attached flavin analogue in recombinant human d-amino acid oxidase. The protein residue Gly-281 was replaced with Cys by site-directed mutagenesis, followed by reaction between mutated apoenzyme and the thiol-reactive flavin analogue, 8-methylsulfonyl FAD. The stoichiometric process of flavin attachment was accompanied by gain in enzymatic activity, reaching up to 26% activity of the recombinant native enzyme. The steady-state kinetic data together with the results of limited proteolysis and benzoate-binding studies suggest that, although mutation perturbs protein structural and catalytic properties, the flavinylation alone does not have any negative impact. We conclude that, despite the implemented restraints on its mobility, the covalently attached flavin is properly positioned within the protein active site and acts efficiently during d-amino acid oxidase catalytic turnover. PMID- 10716695 TI - Heterologous expression in Escherichia coli of the first module of the nonribosomal peptide synthetase for chloroeremomycin, a vancomycin-type glycopeptide antibiotic. AB - The gene cluster from Amycolotopsis orientalis responsible for biosynthesis of the vancomycin-type glycopeptide antibiotic chloroeremomycin was recently sequenced, indicating that this antibiotic derives from a seven-residue peptide synthesized by a three-subunit (CepA, CepB, and CepC) modular nonribosomal peptide synthetase. Expression of all or parts of the peptide synthetase in Escherichia coli would facilitate biochemical characterization of its substrate specificity, an important step toward the development of more potent glycopeptides by combinatorial biosynthesis. To determine whether CepA, a three module 3,158-residue peptide synthetase expected to assemble the first three residues of the heptapeptide precursor, could be heterologously expressed in E. coli and converted to active, holo form by posttranslational priming with a phosphopantetheinyltransferase, we expressed two CepA fragments (CepA1-575 and CepA1-1596) as well as full-length CepA (CepA1-3158). All three constructs were expressed in soluble form. We find that the CepA1-575 fragment, containing adenylation and peptidyl carrier protein domains (A1-PCP1), specifically adenylates l-leucine and d-leucine in a 6:1 ratio, and it can be converted to holo form by the phosphopantetheinyltransferase Sfp; also, we find that holo CepA1-575 can be covalently aminoacylated with l-leucine on the peptidyl carrier protein 1 domain. However, no amino acid-dependent adenylation or aminoacylation activity was detected for the larger CepA constructs with l-leucine or other expected amino acid substrates, suggesting severe folding problems in the multidomain proteins. PMID- 10716696 TI - Millisecond time scale conformational flexibility in a hyperthermophile protein at ambient temperature. AB - Rubredoxin from the hyperthermophile Pyrococcus furiosus is the most thermostable protein characterized to date with an estimated global unfolding rate of 10(-6) s(-1) at 100 degrees C. In marked contrast to these slow global dynamics, hydrogen exchange experiments here demonstrate that conformational opening for solvent access occurs in the approximately millisecond time frame or faster at 28 degrees C for all amide positions. Under these conditions all backbone amides with exchange protection factors between 10(4) and 10(6), for which EX(2) exchange kinetics were directly verified, have exchange activation energy values within 2-3 kcal/mol of that observed for unstructured peptides. The conformational flexibility of this protein is thus sufficient for water and base catalyst access to the exchanging amide with quite limited structural disruption. The common hypothesis that enhanced conformational rigidity in the folded native state underlies the increased thermal stability of hyperthermophile proteins is not supported by these data. PMID- 10716697 TI - Isolation and characterization of pollen coat proteins of Brassica campestris that interact with S locus-related glycoprotein 1 involved in pollen-stigma adhesion. AB - Adhesion of pollen grains to the stigmatic surface is a critical step during sexual reproduction in plants. In Brassica, S locus-related glycoprotein 1 (SLR1), a stigma-specific protein belonging to the S gene family of proteins, has been shown to be involved in this step. However, the identity of the interacting counterpart in pollen and the molecular mechanism of this interaction have not been determined. Using an optical biosensor immobilized with S gene family proteins, we detected strong SLR1-binding activity in pollen coat extracts of Brassica campestris. Two SLR1-binding proteins, named SLR1-BP1 and SLR1-BP2, were identified and purified by the combination of SLR1 affinity column chromatography and reverse-phase HPLC. Sequence analyses revealed that these two proteins (i) differ only in that a proline residue near the N terminus is hydroxylated in SLR1 BP1 but not in SLR1-BP2, and (ii) are members of the class A pollen coat protein (PCP) family, which includes PCP-A1, an SLG (S locus glycoprotein)-binding protein isolated from Brassica oleracea. Kinetic analysis showed that SLR1-BP1 and SLR1-BP2 specifically bound SLR1 with high affinity (K(d) = 5.6 and 4.4 nM, respectively). The SLR1-BP gene was specifically expressed in pollen at late stages of development, and its sequence is highly conserved in Brassica species with the A genome. PMID- 10716698 TI - Increased nutritive value of transgenic potato by expressing a nonallergenic seed albumin gene from Amaranthus hypochondriacus. AB - Improvement of nutritive value of crop plants, in particular the amino acid composition, has been a major long-term goal of plant breeding programs. Toward this end, we reported earlier the cloning of the seed albumin gene AmA1 from Amaranthus hypochondriacus. The AmA1 protein is nonallergenic in nature and is rich in all essential amino acids, and the composition corresponds well with the World Health Organization standards for optimal human nutrition. In an attempt to improve the nutritional value of potato, the AmA1 coding sequence was successfully introduced and expressed in tuber-specific and constitutive manner. There was a striking increase in the growth and production of tubers in transgenic populations and also of the total protein content with an increase in most essential amino acids. The expressed protein was localized in the cytoplasm as well as in the vacuole of transgenic tubers. Thus we have been able to use a seed albumin gene with a well-balanced amino acid composition as a donor protein to develop a transgenic crop plant. The results document, in addition to successful nutritional improvement of potato tubers, the feasibility of genetically modifying other crop plants with novel seed protein composition. PMID- 10716699 TI - An imprinted transcript, antisense to Nesp, adds complexity to the cluster of imprinted genes at the mouse Gnas locus. AB - The Gnas locus in distal mouse chromosome (Chr) 2 is emerging as a complex genomic region. It contains three imprinted genes in the order Nesp-Gnasxl-Gnas. Gnas encodes a G protein alpha-subunit, and Nesp and Gnasxl encode proteins of unknown function expressed in neuroendocrine tissue. Together, these genes form a single transcription unit because transcripts of Nesp and Gnasxl are alternatively spliced onto exon 2 of Gnas. Nesp and Gnasxl are expressed from opposite parental alleles, with Nesp encoding a maternal-specific transcript and Gnasxl encoding a paternal-specific transcript. We now identify a further imprinted transcript in this cluster. Reverse transcription-PCR analysis of Nesp expression in 15. 5-days-postcoitum embryos carrying only maternal or paternal copies of distal Chr 2 revealed an isoform that is exclusively paternally, rather than maternally, expressed. Strand-specific reverse transcription-PCR showed that this form is an antisense transcript. The existence of a paternally expressed antisense transcript was confirmed by Northern blot analysis. The sequence is contiguous with genomic sequence downstream of Nesp and encompasses Nesp exons 1 and 2 and an intervening intron. We propose that Nespas is an additional control element in the imprinting region of mouse distal Chr 2; it adds further complexity to the Gnas-imprinted gene cluster. PMID- 10716700 TI - Insertion site preferences of the P transposable element in Drosophila melanogaster. AB - We determined the genomic sequence at the site of insertion in 2,266 unselected P element insertion events. Estimating physical properties of the genomic DNA at these insertion sites-such as base composition, bendability, A-philicity, protein induced deformability, and B-DNA twist-revealed that they differ significantly from average chromosomal DNA. By examining potential hydrogen bonding sites in the major groove, we identified a 14-bp palindromic pattern centered on the 8-bp target site duplication that is generated by P element insertion. Our results suggest that the P-element transposition mechanism has a two-fold dyad symmetry and recognizes a structural feature at insertion sites, rather than a specific sequence motif. PMID- 10716701 TI - Identification and characterization of an amino acid transporter expressed differentially in liver. AB - Cellular metabolic needs are fulfilled by transport of amino acids across the plasma membrane by means of specialized transporter proteins. Although many of the classical amino acid transporters have been characterized functionally, less than half of these proteins have been cloned. In this report, we identify and characterize a cDNA encoding a plasma membrane amino acid transporter. The deduced amino acid sequence is 505 residues and is highly hydrophobic with the likely predicted structure of 9 transmembrane domains, which putatively place the amino terminus in the cytoplasm and the carboxy terminus on the cell surface. Expression of the cRNA in Xenopus laevis oocytes revealed strong transport activities specific for histidine and glutamine. This protein is a Na(+)- and pH dependent transporter and tolerates substitution of Na(+) by Li(+). Furthermore, this transporter is not an obligatory exchanger because efflux occurs in the absence of influx. This transporter is expressed predominantly in the liver, although it is also present in the kidney, brain, and heart. In the liver, it is located in the plasma membrane of hepatocytes, and the strongest expression was detected in those adjacent to the central vein, gradually decreasing towards the portal tract. Because this protein displays functional similarities to the N system amino acid transport, we have termed it mNAT, for murine N-system amino acid transporter. This is the first transporter gene identified within the N system, one of the major amino acid transport systems in the body. The expression pattern displayed by mNAT suggests a potential role in hepatocyte physiology. PMID- 10716702 TI - Mechanisms of use-dependent plasticity in the human motor cortex. AB - Practicing movements results in improvement in performance and in plasticity of the motor cortex. To identify the underlying mechanisms, we studied use-dependent plasticity in human subjects premedicated with drugs that influence synaptic plasticity. Use-dependent plasticity was reduced substantially by dextromethorphan (an N-methyl-d-aspartate receptor blocker) and by lorazepam [a gamma-aminobutyric acid (GABA) type A receptor-positive allosteric modulator]. These results identify N-methyl-d-aspartate receptor activation and GABAergic inhibition as mechanisms operating in use-dependent plasticity in intact human motor cortex and point to similarities in the mechanisms underlying this form of plasticity and long-term potentiation. PMID- 10716703 TI - Photophysics and optical switching in green fluorescent protein mutants. AB - We demonstrate by using low-temperature high-resolution spectroscopy that red shifted mutants of green fluorescent protein are photo-interconverted among three conformations and are, therefore, not photostable "one-color" systems as previously believed. From our experiments we have further derived the energy level schemes governing the interconversion among the three forms. These results have significant implications for the molecular and cell biological applications of the green fluorescent protein family; for example, in fluorescence resonant energy transfer experiments, a change in "color" on irradiation may not necessarily be due to energy transfer but can also arise from a photo-induced conversion between conformers of the excited species. PMID- 10716704 TI - Unambiguous demonstration of triple-helix-directed gene modification. AB - Triple-helix-forming oligonucleotides (TFOs), which can potentially modify target genes irreversibly, represent promising tools for antiviral therapies. However, their effectiveness on endogenous genes has yet to be unambiguously demonstrated. To monitor endogenous gene modification by TFOs in a yeast model, we inactivated an auxotrophic marker gene by inserting target sequences of interest into its coding region. The genetically engineered yeast cells then were treated with psoralen-linked TFOs followed by UV irradiation, thus generating highly mutagenic covalent crosslinks at the target site whose repair could restore gene function; the number of revertants and spectrum of mutations generated were quantified. Results showed that a phosphoramidate TFO indeed reaches its target sequence, forms crosslinks, and generates mutations at the expected site via a triplex mediated mechanism: (i) under identical conditions, no mutations were generated by the same TFO at two other loci in the target strain, nor in an isogenic control strain carrying a modified target sequence incapable of supporting triple helix formation; (ii) for a given target sequence, whether the triplex was formed in vivo on an endogenous gene or in vitro on an exogenous plasmid, the nature of the mutations generated was identical, and consistent with the repair of a psoralen crosslink at the target site. Although the mutation efficiency was probably too low for therapeutic applications, our results confirm the validity of the triple-helix approach and provide a means of evaluating the effectiveness of new chemically modified TFOs and analogs. PMID- 10716705 TI - Crystal structures of ligand complexes of P450eryF exhibiting homotropic cooperativity. AB - Several mammalian cytochrome P450 (P450) isoforms demonstrate homotropic cooperativity with a number of substrates, including steroids and polycyclic aromatic hydrocarbons. To identify structural factors contributing to steroid and polycyclic aromatic hydrocarbon binding to P450 enzymes and to determine the location of the allosteric site, we investigated interactions of the macrolide hydroxylase P450eryF from Saccharopolyspora erythraea with androstenedione and 9 aminophenanthrene. Spectroscopic binding assays indicate that P450eryF binds androstenedione with an affinity of 365 microM and a Hill coefficient of 1.31 +/- 0.6 and coordinates with 9-aminophenanthrene with an affinity of 91 microM and a Hill coefficient of 1.38 +/- 0.2. Crystals of complexes of androstenedione and 9 aminophenanthrene with P450eryF were grown and diffracted to 2.1 A and 2.35 A, respectively. Electron density maps indicate that for both complexes two ligand molecules are simultaneously present in the active site. The P450eryF/androstenedione model was refined to an r = 18.9%, and the two androstenedione molecules have similar conformations. The proximal androstenedione is positioned such that the alpha-face of carbon-6 is closest to the heme iron, and the second steroid molecule is positioned 5.5 A distal in the active site. The P450eryF/9-aminophenanthrene model was refined to an r = 19.7% with the proximal 9-aminophenanthrene coordinated with the heme iron through the 9-amino group and the second ligand positioned approximately 6 A distal in the active site. These results establish that homotropic cooperativity in ligand binding can result from binding of two substrate molecules within the active site pocket without major conformational changes in the protein. PMID- 10716706 TI - Amplification of the neu/erbB-2 oncogene in a mouse model of mammary tumorigenesis. AB - The neu (c-erbB-2, Her-2) protooncogene is amplified and overexpressed in 20-30% of human breast cancers. Although transgenic mouse models have illustrated the role of Neu in the induction of mammary tumors, Neu expression in these models is driven by a strong viral promoter of questionable relevance to the human disease. To ascertain whether expression of activated Neu under the control of the endogenous promoter in the mammary gland could induce mammary tumors we have generated mice that conditionally express activated Neu under the transcriptional control of the intact endogenous Neu promoter. Expression of oncogenic neu in the mammary gland resulted in accelerated lobulo-alveolar development and formation of focal mammary tumors after a long latency period. However, expression of activated Neu under the normal transcriptional control of the endogenous promoter was not sufficient for the initiation of mammary carcinogenesis. Strikingly, all mammary tumors bear amplified copies (2-22 copies) of the activated neu allele relative to the wild-type allele and express highly elevated levels of neu transcript and protein. Thus, like human erbB-2-positive breast tumors, mammary tumorigenesis in this mouse model requires the amplification and commensurate elevated expression of the neu gene. PMID- 10716707 TI - Progesterone-metabolite prevents protein kinase C-dependent modulation of gamma aminobutyric acid type A receptors in oxytocin neurons. AB - Gonadal steroid feedback to oxytocin neurons during pregnancy is in part mediated via the neurosteroid allopregnanolone (3alpha-OH-DHP), acting as allosteric modulator of postsynaptic gamma-aminobutyric acid type A (GABA(A)) receptors. We describe here a form of nongenomic progesterone signaling by showing that 3alpha OH-DHP not only potentiates GABA(A) receptor-channel activity but also prevents its modulation by protein kinase C (PKC). Application of oxytocin or stimulation of PKC suppressed the postsynaptic GABA responses of oxytocin neurons in the absence, but not in the presence of 3alpha-OH-DHP. This finding was true at the juvenile stage and during late pregnancy, when the GABA(A) receptor is sensitive to 3alpha-OH-DHP. In contrast, after parturition, when the GABA(A) receptors expressed by oxytocin neurons are less sensitive to 3alpha-OH-DHP, this neurosteroid no longer counteracts PKC. The change in GABA(A)-receptor responsiveness to 3alpha-OH-DHP helps to explain the onset of firing activity and thus the induction of oxytocin release at parturition. PMID- 10716708 TI - SAC3 may link nuclear protein export to cell cycle progression. AB - Selective movement of proteins between the nucleus and the cytoplasm is a regulatory mechanism exploited extensively by the eukaryotic cell. We have identified the evolutionarily conserved Sac3 protein, which was implicated previously in the regulation of mitosis [Bauer, A. & Kolling, R. (1996) J. Cell Sci. 109, 1575-1583] as a novel mediator of nuclear protein export. We show that Sac3p is localized to the nuclear pore, where it interacts with nucleoporins. Loss of SAC3 function results in a block in nuclear export of a nuclear export signal-containing reporter protein. Our results also demonstrate that SAC3 interacts genetically with the nuclear protein export factors Crm1p/Xpo1p and Yrb2p. Taken together, these data indicate a link between nuclear protein export and transition through the cell cycle. PMID- 10716709 TI - An approach to gene-specific transcription inhibition using oligonucleotides complementary to the template strand of the open complex. AB - The single-stranded region of DNA within the open complex of transcriptionally active genes provides a unique target for the design of gene-specific transcription inhibitors. Using the Escherichia coli lac UV5 and trp EDCBA promoters as in vitro models of open complex formation, we have identified the sites inside these transcription bubbles that are accessible for hybridization by short, nuclease-resistant, non-extendable oligoribonucleotides (ORNs). Binding of ORNs inside the open complex was determined by linking the chemical nuclease bis(1,10-phenanthroline) cuprous chelate [(OP)(2)Cu(+)] to the ORN and demonstrating template-specific DNA scission. In addition, these experiments were supported by in vitro transcription inhibition. We find that the most effective inhibitors are 5 nt long and have sequences that are complementary to the DNA template strand in the region near the transcription start site. The ORNs bind to the DNA template strand, forming an antiparallel heteroduplex inside the open complex. In this system, RNA polymerase is essential not only to melt the duplex DNA but also to facilitate hybridization of the incoming ORN. This paradigm for gene-specific inactivation relies on the base complementarity of the ORN and the catalytic activity and sequence specificity of RNA polymerase for the site- and sequence-specific recognition and inhibition of transcriptionally active DNA. PMID- 10716711 TI - An archaeal genomic signature. AB - Comparisons of complete genome sequences allow the most objective and comprehensive descriptions possible of a lineage's evolution. This communication uses the completed genomes from four major euryarchaeal taxa to define a genomic signature for the Euryarchaeota and, by extension, the Archaea as a whole. The signature is defined in terms of the set of protein-encoding genes found in at least two diverse members of the euryarchaeal taxa that function uniquely within the Archaea; most signature proteins have no recognizable bacterial or eukaryal homologs. By this definition, 351 clusters of signature proteins have been identified. Functions of most proteins in this signature set are currently unknown. At least 70% of the clusters that contain proteins from all the euryarchaeal genomes also have crenarchaeal homologs. This conservative set, which appears refractory to horizontal gene transfer to the Bacteria or the Eukarya, would seem to reflect the significant innovations that were unique and fundamental to the archaeal "design fabric." Genomic protein signature analysis methods may be extended to characterize the evolution of any phylogenetically defined lineage. The complete set of protein clusters for the archaeal genomic signature is presented as supplementary material (see the PNAS web site, www.pnas.org). PMID- 10716710 TI - Differential requirement for p19ARF in the p53-dependent arrest induced by DNA damage, microtubule disruption, and ribonucleotide depletion. AB - p19ARF has been implicated as a key regulator of p53 stability and activation. While numerous stresses activate the p53 growth arrest pathway, those requiring p19ARF remain to be elucidated. We used p19ARF knockout mouse embryo fibroblasts to show that DNA damage and microtubule disruption require p19ARF to induce p53 responses, whereas ribonucleotide depletion and inhibition of RNA synthesis by low doses of actinomycin D do not. The data provide evidence that the arrest pathway activated by ribonucleotide depletion involves some different signal transducers than those activated by DNA damage or microtubule disruption. We also present biochemical analyses that provide insights into the mechanism by which p53 and p19ARF cooperate in normal cells to induce cell cycle arrest. PMID- 10716712 TI - New studies on the heat resistance of hamster-adapted scrapie agent: threshold survival after ashing at 600 degrees C suggests an inorganic template of replication. AB - One-gram samples from a pool of crude brain tissue from hamsters infected with the 263K strain of hamster-adapted scrapie agent were placed in covered quartz glass crucibles and exposed for either 5 or 15 min to dry heat at temperatures ranging from 150 degrees C to 1,000 degrees C. Residual infectivity in the treated samples was assayed by the intracerebral inoculation of dilution series into healthy weanling hamsters, which were observed for 10 months; disease transmissions were verified by Western blot testing for proteinase-resistant protein in brains from clinically positive hamsters. Unheated control tissue contained 9.9 log(10)LD(50)/g tissue; after exposure to 150 degrees C, titers equaled or exceeded 6 log(10)LD(50)/g, and after exposure to 300 degrees C, titers equaled or exceeded 4 log(10)LD(50)/g. Exposure to 600 degrees C completely ashed the brain samples, which, when reconstituted with saline to their original weights, transmitted disease to 5 of 35 inoculated hamsters. No transmissions occurred after exposure to 1, 000 degrees C. These results suggest that an inorganic molecular template with a decomposition point near 600 degrees C is capable of nucleating the biological replication of the scrapie agent. PMID- 10716713 TI - Understanding hippocampal activity by using purposeful behavior: place navigation induces place cell discharge in both task-relevant and task-irrelevant spatial reference frames. AB - Continuous rotation of an arena in a cue-rich room dissociates the stationary room-bound information from the rotating arena-bound information. This disrupted spatial discharge in the majority of place cells from rats trained to collect randomly scattered food. In contrast, most place cell firing patterns recorded from rats trained to solve a navigation task on the rotating arena were preserved during the rotation. Spatial discharge was preserved in both the task-relevant stationary and the task-irrelevant rotating reference frames, but firing was more organized in the task-relevant frame. It is concluded that, (i) the effects of environmental manipulations can be understood with confidence only when the rat's purposeful behavior is used to formulate interpretations of the data, and (ii) hippocampal place cell activity is organized in multiple overlapping spatial reference frames. PMID- 10716714 TI - Sequence-related protein export NTPases encoded by the conjugative transfer region of RP4 and by the cag pathogenicity island of Helicobacter pylori share similar hexameric ring structures. AB - RP4 TrbB, an essential component of the conjugative transfer apparatus of the broad-host-range plasmid RP4, is a member of the PulE protein superfamily involved in multicomponent machineries transporting macromolecules across the bacterial envelope. PulE-like proteins share several well conserved motifs, most notable a nucleoside triphosphate binding motif (P-loop). Helicobacter pylori HP0525 also belongs to the PulE superfamily and is encoded by the pathogenicity island cag, involved in the inflammatory response of infected gastric epithelial cells in mammals. The native molecular masses of TrbB and HP0525 as determined by gel filtration and glycerol gradient centrifugation suggested a homohexameric structure in the presence of ATP and Mg(2+). In the absence of nucleotides and bivalent cations, TrbB behaved as a tetramer whereas the hexameric state of HP0525 remained unaffected. Electron microscopy and image processing demonstrated that TrbB and HP0525 form ring-shaped complexes (diameter: 12 nm) with a central region (diameter: 3 nm) of low electron density when incubated in the presence of ATP and Mg(2+). However, the TrbB average image appeared to be more elliptical with strong twofold rotational symmetry whereas HP0525 complexes are regular hexagons. Six well defined triangle-shaped areas of high electron density were distinguishable in both cases. Covalent crosslinking of TrbB suggests that the hexameric ring is composed from a trimer of dimers, because only dimeric, tetrameric, and hexameric species were detectable. The toroidal structure of TrbB and HP0525 suggests that both proteins catalyze a repetitive process, most probably translocating a cognate substrate across the inner membrane. PMID- 10716715 TI - Severe B cell hyperplasia and autoimmune disease in TALL-1 transgenic mice. AB - TALL-1/Blys/BAFF is a member of the tumor necrosis factor (TNF) ligand superfamily that is functionally involved in B cell proliferation. Here, we describe B cell hyperplasia and autoimmune lupus-like changes in transgenic mice expressing TALL-1 under the control of a beta-actin promoter. The TALL-1 transgenic mice showed severe enlargement of spleen, lymph nodes, and Peyer's patches because of an increased number of B220+ cells. The transgenic mice also had hypergammaglobulinemia contributed by elevations of serum IgM, IgG, IgA, and IgE. In addition, a phenotype similar to autoimmune lupus-like disease was also seen in TALL-1 transgenic mice, characterized by the presence of autoantibodies to nuclear antigens and immune complex deposits in the kidney. Prolonged survival and hyperactivity of transgenic B cells may contribute to the autoimmune lupus like phenotype in these animals. Our studies further confirm TALL-1 as a stimulator of B cells that affect Ig production. Thus, TALL-1 may be a primary mediator in B cell-associated autoimmune diseases. PMID- 10716716 TI - Human neuronal threonine-for-leucine-248 alpha 7 mutant nicotinic acetylcholine receptors are highly Ca2+ permeable. AB - A cDNA coding for the human neuronal nicotinic alpha7 receptor subunit with Leu 248 mutated to threonine was expressed in Xenopus oocytes. When activated by acetylcholine (AcCho), the receptors expressed generated currents that had low desensitization, linear current-voltage relation, and high apparent affinity for both AcCho and nicotine. These characteristics are similar to those already described for the chick threonine-for-leucine-247 alpha7 nicotinic AcCho receptor (nAcChoR) mutant (L247Talpha7). These properties were all substantially maintained when the human L248Talpha7 mutant was transiently expressed in human Bosc 23 cells. Simultaneous whole-cell clamp and fluorescence measurements with the Ca(2+) indicator dye Fura-2 showed that nicotine induced a Ca(2+) influx in standard 2 mM Ca(2+) solution. The average fractional Ca(2+) current flowing through L248Talpha7 nAcChoRs was 6.7%, which is larger than that flowing through muscle alpha(beta)epsilon(delta) nAcChoRs (4.1%). The relative Ca(2+) permeability, determined in oocytes in the absence of Cl(-), was measured from the shift in reversal potential caused by increasing the external Ca(2+) concentration from 1 to 10 mM. The human wild-type alpha7 nAcChoR was found to be more permeable than the L248Talpha7 mutant to Ca(2+). Our findings indicate that the Ca(2+) permeability of the homomeric alpha7 nAcChoR is larger than that of the heteromeric neuronal nicotinic receptors studied to date and is possibly similar to that of the N-methyl-D-aspartate subtype of brain glutamate receptors. PMID- 10716717 TI - Synthesis and antitumor activity of an inhibitor of fatty acid synthase. AB - Compared to normal human tissues, many common human cancers, including carcinoma of the colon, prostate, ovary, breast, and endometrium, express high levels of fatty acid synthase (FAS, EC ), the primary enzyme responsible for the synthesis of fatty acids. This differential expression of FAS between normal tissues and cancer has led to the notion that FAS is a target for anticancer drug development. Recent studies with C75, an inhibitor of fatty acid synthesis, have shown significant antitumor activity with concomitant inhibition of fatty acid synthesis in tumor tissue and normal liver. Importantly, histopathological analysis of normal tissues after C75 treatment showed no adverse effects on proliferating cellular compartments, such as bone marrow, gastrointestinal tract, skin, or lymphoid tissues. In this study, we describe the de novo synthesis of C75 based on the known mechanism of action of cerulenin and the theoretical reaction intermediates of the beta-ketoacyl synthase moiety of FAS. In addition, we demonstrate that C75 is a synthetic, chemically stable inhibitor of FAS. C75 inhibits purified mammalian FAS with characteristics of a slow-binding inhibitor and also inhibits fatty acid synthesis in human cancer cells. Treatment of human breast cancer cells with [5-(3)H]C75 demonstrates that C75 reacts preferentially with FAS in whole cells. Therefore, we have shown that the primary mechanism of the antitumor activity of C75 is likely mediated through its interaction with, and inhibition of, FAS. This development will enable the in vivo study of FAS inhibition in human cancer and other metabolic diseases. PMID- 10716718 TI - Abnormal development of Purkinje cells and lymphocytes in Atm mutant mice. AB - Motor incoordination, immune deficiencies, and an increased risk of cancer are the characteristic features of the hereditary disease ataxia-telangiectasia (A T), which is caused by mutations in the ATM gene. Through gene targeting, we have generated a line of Atm mutant mice, Atm(y/y) mice. In contrast to other Atm mutant mice, Atm(y/y) mice show a lower incidence of thymic lymphoma and survive beyond a few months of age. Atm(y/y) mice exhibit deficits in motor learning indicative of cerebellar dysfunction. Even though we found no gross cerebellar degeneration in older Atm(y/y) animals, ectopic and abnormally differentiated Purkinje cells were apparent in mutant mice of all ages. These findings establish that some neuropathological abnormalities seen in A-T patients also are present in Atm mutant mice. In addition, we report a previously unrecognized effect of Atm deficiency on development or maintenance of CD4(+)8(+) thymocytes. We discuss these findings in the context of the hypothesis that abnormal development of Purkinje cells and lymphocytes contributes to the pathogenesis of A-T. PMID- 10716719 TI - Functional polymorphisms of the human multidrug-resistance gene: multiple sequence variations and correlation of one allele with P-glycoprotein expression and activity in vivo. AB - To evaluate whether alterations in the multidrug-resistance (MDR)-1 gene correlate with intestinal MDR-1 expression and uptake of orally administered P glycoprotein (PGP) substrates, we analyzed the MDR-1 sequence in 21 volunteers whose PGP expression and function in the duodenum had been determined by Western blots and quantitative immunohistology (n = 21) or by plasma concentrations after orally administered digoxin (n = 8 + 14). We observed a significant correlation of a polymorphism in exon 26 (C3435T) of MDR-1 with expression levels and function of MDR-1. Individuals homozygous for this polymorphism had significantly lower duodenal MDR-1 expression and the highest digoxin plasma levels. Homozygosity for this variant was observed in 24% of our sample population (n = 188). This polymorphism is expected to affect the absorption and tissue concentrations of numerous other substrates of MDR-1. PMID- 10716720 TI - Tumorigenesis in the multiple intestinal neoplasia mouse: redundancy of negative regulators and specificity of modifiers. AB - The interaction between mutations in the tumor-suppressor genes Apc and p53 was studied in congenic mouse strains to minimize the influence of polymorphic modifiers. The multiplicity and invasiveness of intestinal adenomas of Apc(Min/+) (Min) mice was enhanced by deficiency for p53. In addition, the occurrence of desmoid fibromas was strongly enhanced by p53 deficiency. The genetic modifier Mom1 and the pharmacological agents piroxicam and difluoromethylornithine each reduced intestinal adenoma multiplicity in the absence of p53 function. Mom1 showed no influence on the development of desmoid fibromas, whereas the combination of piroxicam and difluoromethylornithine exerted a moderate effect. The ensemble of tumor suppressors and modifiers of a neoplastic process can be usefully analyzed in respect to tissue specificity and synergy. PMID- 10716721 TI - The decomposition of peroxynitrite to nitroxyl anion (NO-) and singlet oxygen in aqueous solution. AB - The mechanism of decomposition of peroxynitrite (OONO(-)) in aqueous sodium phosphate buffer solution at neutral pH was investigated. The OONO(-) was synthesized by directly reacting nitric oxide with superoxide anion at pH 13. The hypothesis was explored that OONO(-), after protonation at pH 7.0 to HOONO, decomposes into (1)O(2) and HNO according to a spin-conserved unimolecular mechanism. Small aliquots of the concentrated alkaline OONO(-) solution were added to a buffer solution (final pH 7.0-7.2), and the formation of (1)O(2) and NO(-) in high yields was observed. The (1)O(2) generated was trapped as the transannular peroxide (DPAO(2)) of 9, 10-diphenylanthracene (DPA) dissolved in carbon tetrachloride. The nitroxyl anion (NO-) formed from HNO (pKa 4.5) was trapped as nitrosylhemoglobin (HbNO) in an aqueous methemoglobin (MetHb) solution. In the presence of 25 mM sodium bicarbonate, which is known to accelerate the rate of decomposition of OONO(-), the amount of singlet oxygen trapped was reduced by a factor of approximately 2 whereas the yield of trapping of NO(-) by methemoglobin remained unaffected. Because NO(3)(-) is known to be the ultimate decomposition product of OONO(-), these results suggest that the nitrate anion is not formed by a direct isomerization of OONO(-), but by an indirect route originating from NO(-). PMID- 10716722 TI - An artificial tetramerization domain restores efficient assembly of functional Shaker channels lacking T1. AB - One feature shared by all Shaker-type voltage-gated K(+) channels is a highly conserved domain (T1) located in the cytoplasmic N terminus. The T1 domain is a key determinant of which subtypes can form heteromultimeric channels, suggesting that T1 functions during channel assembly. To better define the role of T1 during channel assembly and separate this function from potential contributions to channel permeation and gating, we replaced the T1 domain (residues 96-183) of ShakerB with a coiled-coil sequence (GCN4-LI) that forms parallel tetramers. Deleting T1 dramatically, but not completely, abolished channel formation under most expression conditions. Channels lacking T1 are functional and K(+) selective, although they activate at more hyperpolarized membrane potentials and inactivate less completely. Insertion of the artificial tetramerization domain (GCN4-LI) restored efficient channel formation, suggesting that tetramerization of the cytoplasmic T1 domain promotes transmembrane channel assembly by increasing the effective local subunit concentration for T1 compatible subunits. We propose that T1 tetramerization promotes subfamily-specific assembly through kinetic partitioning of the assembly process, but is not required for subsequent steps in channel assembly and folding. PMID- 10716723 TI - The molecular basis of vernalization: the central role of FLOWERING LOCUS C (FLC). AB - In Arabidopsis, the MADS-box protein encoded by FLOWERING LOCUS C (FLC) is a repressor of flowering. Vernalization, which promotes flowering in the late flowering ecotypes and many late-flowering mutants, decreases the level of FLC transcript and protein in the plant. This vernalization-induced reduction in FLC transcript levels is mitotically stable and occurs in all tissues. FLC activity is restored in each generation, as is the requirement of a low-temperature exposure for the promotion of flowering. The level of FLC determines the extent of the vernalization response in the promotion of flowering, and there is a quantitative relationship between the duration of cold treatment and the extent of down-regulation of FLC activity. We conclude that FLC is the central regulator of the induction of flowering by vernalization. Other vernalization-responsive late-flowering mutants, which are disrupted in genes that encode regulators of FLC, are late-flowering as a consequence of their elevated levels of FLC. PMID- 10716724 TI - AiiA, an enzyme that inactivates the acylhomoserine lactone quorum-sensing signal and attenuates the virulence of Erwinia carotovora. AB - N-acylhomoserine lactones, known as autoinducers (AIs), are widely conserved signal molecules present in quorum-sensing systems of many gram-negative bacteria. AIs are involved in the regulation of diverse biological functions, including expression of pathogenic genes in the plant pathogens Pseudomonas solanacearum, several Erwinia species, and the human pathogen Pseudomonas aeruginosa. A bacterial isolate, Bacillus sp. 240B1, is capable of enzymatic inactivation of AIs. The gene (aiiA) for AI inactivation from Bacillus sp. 240B1 has been cloned and shown to encode a protein of 250 amino acids. Sequence alignment indicates that AiiA contains a "HXHXDH" zinc-binding motif that is conserved in several groups of metallohydrolases. Site-directed mutagenesis showed that conserved aspartate and most histidine residues are required for AiiA activity. Expression of aiiA in transformed Erwinia carotovora strain SCG1 significantly reduces the release of AI, decreases extracellular pectolytic enzyme activities, and attenuates pathogenicity on potato, eggplant, Chinese cabbage, carrot, celery, cauliflower, and tobacco. Our results indicate that the AI-inactivation approach represents a promising strategy for prevention of diseases in which virulence is regulated by AIs. PMID- 10716725 TI - G protein-coupled receptors regulate Na+,K+-ATPase activity and endocytosis by modulating the recruitment of adaptor protein 2 and clathrin. AB - Inhibition of Na(+),K(+)-ATPase (NKA) activity in renal epithelial cells by activation of G protein-coupled receptors is mediated by phosphorylation of the catalytic alpha-subunit followed by endocytosis of active molecules. We examined whether agonists that counteract this effect do so by dephosphorylation of the alpha-subunit or by preventing its internalization through a direct interaction with the endocytic network. Oxymetazoline counteracted the action of dopamine on NKA activity, and this effect was achieved not by preventing alpha-subunit phosphorylation, but by impaired endocytosis of alpha-subunits into clathrin vesicles and early and late endosomes. Dopamine-induced inhibition of NKA activity and alpha-subunit endocytosis required the interaction of adaptor protein 2 (AP-2) with the catalytic alpha-subunit. Phosphorylation of the alpha subunit is essential because dopamine failed to promote such interaction in cells lacking the protein kinase C phosphorylation residue (S18A). Confocal microscopy confirmed that oxymetazoline prevents incorporation of NKA molecules into clathrin vesicles by inhibiting the ability of dopamine to recruit clathrin to the plasma membrane. Dopamine decreased the basal levels of inositol hexakisphosphate (InsP(6)), whereas oxymetazoline prevented this effect. Similar increments (above basal) in the concentration of InsP(6) induced by oxymetazoline prevented AP-2 binding to the NKA alpha-subunit in response to dopamine. In conclusion, inhibition of NKA activity can be reversed by preventing its endocytosis without altering the state of alpha-subunit phosphorylation; increased InsP(6) in response to G protein-coupled receptor signals blocks the recruitment of AP-2 and thereby clathrin-dependent endocytosis of NKA. PMID- 10716726 TI - Large exons encoding multiple ectodomains are a characteristic feature of protocadherin genes. AB - Recent studies revealed a striking difference in the genomic organization of classic cadherin genes and one family of "nonclassic cadherin" genes designated protocadherins. Specifically, the DNA sequences encoding the ectodomain repeats of classic cadherins are interrupted by multiple introns. By contrast, all of the encoded ectodomains of each member of the protocadherin gene clusters are present in one large exon. To determine whether large ectodomain exons are a general feature of protocadherin genes we have investigated the genomic organization of several additional human protocadherin genes by using DNA sequence information in GenBank. These genes include protocadherin 12 (Pcdh12), an ortholog of the mouse vascular endothelial cadherin-2 gene; hFmi1 and hFmi2, homologs of the Drosophila planar cell polarity gene, flamingo; hFat2, a homolog of the Drosophila tumor suppressor gene fat; and the Drosophila DN-cadherin and DE-cadherin genes. Each of these genes was found to be a member of the protocadherin subfamily, based on amino acid sequence comparisons of their ectodomains. Remarkably, all of these protocadherin genes share a common feature: most of the genomic DNA sequences encoding their ectodomains are not interrupted by an intron. We conclude that the presence of unusually large exons is a characteristic feature of protocadherin genes. PMID- 10716727 TI - Protonation of the beta-lactam nitrogen is the trigger event in the catalytic action of class A beta-lactamases. AB - The pH dependence of the pK(a) values of all ionizable groups and of the electrostatic potential at grid points corresponding to catalytically important atoms in the active site of TEM-1 beta-lactamase has been calculated by a mean field approach for reaction intermediates modeled on the basis of energy minimized x-ray crystallographic coordinates. By estimating electrostatic contributions to the free energy changes accompanying the conversion of the free enzyme into the acylenzyme reaction intermediate, we found that acid-catalyzed protonation of the beta-lactam nitrogen is energetically favored as the initiating event, followed by base-catalyzed nucleophilic attack on the carbonyl carbon of the beta-lactam group. N-protonation is catalyzed through a hydrogen bonded cluster involving the 2-carboxylate group of the substrate, the side chains of S130 and K234, and a solvent molecule. Nucleophilic attack on the carbonyl carbon is carried out by the side chain of S70 with proton abstraction catalyzed by a water molecule hydrogen-bonded to the side chain of E166. Stabilization of ion pairs in the active site through interactions with distant clusters of charged residues in the enzyme was concluded to be an important driving force of the catalytic mechanism. PMID- 10716728 TI - NF-kappa B activation induced by T cell receptor/CD28 costimulation is mediated by protein kinase C-theta. AB - Protein kinase C-theta (PKCtheta) is a Ca(2+)-independent member of the PKC family that is selectively expressed in skeletal muscle and T lymphocytes and plays an important role in T cell activation. However, the molecular basis for the important functions of PKCtheta in T cells and the manner in which it becomes coupled to the T cell receptor-signaling machinery are unknown. We addressed the functional relationship between PKCtheta and CD28 costimulation, which plays an essential role in T cell receptor-mediated IL-2 production. Here, we provide evidence that PKCtheta is functionally coupled to CD28 costimulation by virtue of its selective ability to activate the CD28RE/activator protein-1 (AP-1) element in the IL-2 gene promoter. First, CD28 costimulation enhanced the membrane translocation and catalytic activation of PKCtheta. Second, among several PKC isoforms, PKCtheta was the only one capable of activating NF-kappaB or CD28RE/AP 1 reporters in T cells (but not in 293T cells). Third, wild-type PKCtheta synergized with CD28/CD3 signals to activate CD28RE/AP-1. In addition, PKCtheta selectively synergized with Tat to activate a CD28RE/AP-1 reporter. Fourth, CD3/CD28-induced CD28RE/AP-1 activation and NF-kappaB nuclear translocation were blocked by a selective PKCtheta inhibitor. Last, PKCtheta-mediated activation of the same reporter was inhibited by the proteasome inhibitor MG132 (which blocks IkappaB degradation) and was found to involve IkappaB-kinase beta. These findings identify a unique PKCtheta-mediated pathway for the costimulatory action of CD28, which involves activation of the IkappaB-kinase beta/IkappaB/NF-kappaB-signaling cascade. PMID- 10716729 TI - Lipid rafts function in biosynthetic delivery of proteins to the cell surface in yeast. AB - Lipid rafts, formed by lateral association of sphingolipids and cholesterol, have been implicated in membrane traffic and cell signaling in mammalian cells. Sphingolipids also have been shown to play a role in protein sorting in yeast. Therefore, we wanted to investigate whether lipid rafts exist in yeast and whether these membrane microdomains have an analogous function to their mammalian counterparts. We first developed a protocol for isolating detergent-insoluble glycolipid-enriched complexes (DIGs) from yeast cells. Sequencing of the major protein components of the isolated DIGs by mass spectrometry allowed us to identify, among others, Gas1p, Pma1p, and Nce2p. Using lipid biosynthetic mutants we could demonstrate that conditions that impair the synthesis of sphingolipids and ergosterol also disrupt raft association of Gas1p and Pma1p but not the secretion of acid phosphatase. That endoplasmic reticulum (ER)-to-Golgi transport of Gas1p is blocked in the sphingolipid mutant lcb1-100 raised the question of whether proteins associate with lipid rafts in the ER or later as shown in mammalian cells. Using the sec18-1 mutant we found that DIGs are present already in the ER. Taken together, our results suggest that lipid rafts are involved in the biosynthetic delivery of proteins to the yeast plasma membrane. PMID- 10716730 TI - Evidence for secretory pathway localization of a voltage-dependent anion channel isoform. AB - Voltage-dependent anion channels (VDACs) are pore-forming proteins (porins) that form the major pathway for movement of adenine nucleotides through the outer mitochondrial membrane. Electrophysiological studies indicate that VDAC-like channel activity is also prevalent in the cell membranes of many mammalian cells. However, the multitopological localization of porins outside the mitochondrion has remained an extremely controversial issue. Herein, we show that usage of two alternative first exons of the murine VDAC-1 gene leads to expression of two porins differing within their N termini. One porin (plasmalemmal VDAC-1) harboring a hydrophobic leader peptide is primarily targeted through the Golgi apparatus to the cell membrane. In contrast, the second isoform lacking the N terminal leader (mitochondrial VDAC-1) is translocated more efficiently into the outer mitochondrial membrane. Thus, our data provide unique genetic evidence in favor of a multitopological localization of a mitochondrial porin. PMID- 10716731 TI - Male-male competition magnifies inbreeding depression in wild house mice. AB - The detrimental effects of inbreeding on vertebrates are well documented for early stages of the life cycle in the laboratory. However, the consequences of inbreeding on long-term survival and reproductive success (Darwinian fitness) are uncertain for vertebrates in the wild. Here, we report direct experimental evidence for vertebrates that competition increases the harmful effects of inbreeding on offspring survival and reproduction. We compared the fitness of inbred (from full-sib matings) and outbred wild house mice (Mus domesticus) in large, seminatural enclosures. Inbred males sired only one-fifth as many surviving offspring as outbred males because of their poor competitive ability and survivorship. In laboratory conditions, inbreeding had relatively minor effects on male reproductive success and no effect on survivorship. Seminatural conditions did not increase inbreeding depression for females, probably because females were not competing for any critical resources. The overall reduction in fitness from inbreeding was 57%, which is 4.5 times as great as previous estimates from the laboratory. These results have important implications for medicine, conservation, evolutionary biology, and functional genomics. PMID- 10716732 TI - Experimental evolution of aging, growth, and reproduction in fruitflies. AB - We report in this paper an evolutionary experiment on Drosophila that tested life history theory and the evolutionary theory of aging. As theory predicts, higher extrinsic mortality rates did lead to the evolution of higher intrinsic mortality rates, to shorter lifespans, and to decreased age and size at eclosion; peak fecundity also shifted earlier in life. These results confirm the key role of extrinsic mortality rates in the evolution of growth, maturation, reproduction, and aging, and they do so with a selection regime that maintained selection on fertility throughout life while holding population densities constant. PMID- 10716733 TI - Oligomerization of serotonin transporter and its functional consequences. AB - Two forms of serotonin transporter (SERT) were prepared with different epitope tags. When co-expressed in HeLa cells, the form containing a FLAG tag (Res-FLAG) was associated with the form containing a c-myc tag (Sens-myc). Antibody against c-myc precipitated Res-FLAG from detergent extracts of cells expressing both forms, but not when Res-FLAG was expressed alone. The specificity of the interaction was demonstrated by the observation that anti-myc antibodies did not precipitate the unrelated vesicular stomatitis virus coat glycoprotein when it was co-expressed with Sens-myc. Sens-myc contained a reactive cysteine at position 172, which reacted with both (2-aminoethyl)methanethiosulfonate and N biotinylaminoethyl methanethiosulfonate on the surface of intact cells. Sens-myc, but not Res-FLAG, was inactivated by these reagents. When co-expressed with Sens myc, functionally active Res-FLAG was precipitated by immobilized streptavidin from digitonin-solubilized cells that had been treated with N-biotinylaminoethyl methanethiosulfonate. In cells co-expressing mixtures of Sens-myc and Res-FLAG, the amount of inactivation by (2-aminoethyl)methanethiosulfonate was less than expected if the two forms were independent. The results are consistent with a dimeric form of SERT with functional interactions between subunits, and with association of dimers into a higher order complex, possibly a tetramer. PMID- 10716734 TI - Reciprocal domain evolution within a transactivator in a restricted sequence space. AB - offhough the concept of domain merging and shuffling as a major force in protein evolution is well established, it has been difficult to demonstrate how domains coadapt. Here we show evidence of coevolution of the Sinorhizobium meliloti NifA (SmNifA) domains. We found that, because of the lack of a conserved glycine in its DNA-binding domain, this transactivator protein interacts weakly with the enhancers. This defect, however, was compensated by evolving a highly efficient activation domain that, contrasting to Bradyrhizobium japonicum NifA (BjNifA), can activate in trans. To explore paths that lead to this enhanced activity, we mutagenized BjNifA. After three cycles of mutagenesis and selection, a highly active derivative was obtained. Strikingly, all mutations changed to amino acids already present in SmNifA. Our artificial process thus recreated the natural evolution followed by this protein and suggests that NifA is trapped in a restricted sequence space with very limited solutions for higher activity by point mutation. PMID- 10716735 TI - A method to identify cDNAs based on localization of green fluorescent protein fusion products. AB - We previously established a high-efficiency, retrovirus-mediated expression cloning method. Using this system, we now have developed an expression cloning method (FL-REX; fluorescence localization-based retrovirus-mediated expression cloning) in which cDNAs can be isolated based on the subcellular localization of their protein products. Complementary DNAs generated from mRNA using random hexamers were fused to the cDNA of green fluorescent protein (GFP) in the pMX retrovirus vector. The resulting cDNA-GFP fusion library was transfected into retrovirus-packaging cells, and the derived retroviruses were used to infect NIH 3T3 cells. Infected cells then were screened to identify cDNAs of interest through the subcellular localization of the GFP-fusion products. Using FL-REX, we have identified 25 cDNAs, most of which showed reasonable subcellular localization as GFP-fusion proteins, indicating that FL-REX is useful for identification of proteins that show specific intracellular localization. PMID- 10716736 TI - The dynamics of repeated elements: applications to the epidemiology of tuberculosis. AB - We propose a stepwise mutation model to describe the dynamics of DNA fingerprint variation in Mycobacterium tuberculosis. The genome of M. tuberculosis carries insertion sequences (IS6110) that are relatively stable over time periods of months but have an observable transposition rate over longer time scales. Variability in copy number and genomic location of (IS6110) can be harnessed to generate a DNA fingerprint for each strain, by digesting the genome with a restriction enzyme and using a portion of the element as a probe for Southern blots. The number of bands found for a given genome approximates the number of copies of IS6110 it carries. A large data set of such fingerprints from tuberculosis (TB) cases in San Francisco provides an observed distribution of IS6110 copy number. Implementation of the model through deterministic and stochastic simulation indicates some general features of IS/TB dynamics. By comparing observations with outcomes of the model, we conclude that the IS/TB system is very heterogeneous and far from equilibrium. We find that the transposition parameters have a much stronger effect than the epidemic parameters on copy number distribution. PMID- 10716737 TI - Inhibition of integrin-linked kinase (ILK) suppresses activation of protein kinase B/Akt and induces cell cycle arrest and apoptosis of PTEN-mutant prostate cancer cells. AB - PTEN is a tumor suppressor gene located on chromosome 10q23 that encodes a protein and phospholipid phosphatase. Somatic mutations of PTEN are found in a number of human malignancies, and loss of expression, or mutational inactivation of PTEN, leads to the constitutive activation of protein kinase B (PKB)/Akt via enhanced phosphorylation of Thr-308 and Ser-473. We recently have demonstrated that the integrin-linked kinase (ILK) can phosphorylate PKB/Akt on Ser-473 in a phosphoinositide phospholipid-dependent manner. We now demonstrate that the activity of ILK is constitutively elevated in a serum- and anchorage-independent manner in PTEN-mutant cells, and transfection of wild-type (WT) PTEN into these cells inhibits ILK activity. Transfection of a kinase-deficient, dominant negative form of ILK or exposure to a small molecule ILK inhibitor suppresses the constitutive phosphorylation of PKB/Akt on Ser-473, but not on Thr-308, in the PTEN-mutant prostate carcinoma cell lines PC-3 and LNCaP. Transfection of dominant-negative ILK and WT PTEN into these cells also results in the inhibition of PKB/Akt kinase activity. Furthermore, dominant-negative ILK or WT PTEN induces G(1) phase cycle arrest and enhanced apoptosis. Together, these data demonstrate a critical role for ILK in PTEN-dependent cell cycle regulation and survival and indicate that inhibition of ILK may be of significant value in PTEN-mutant tumor therapy. PMID- 10716738 TI - Navigation-related structural change in the hippocampi of taxi drivers. AB - Structural MRIs of the brains of humans with extensive navigation experience, licensed London taxi drivers, were analyzed and compared with those of control subjects who did not drive taxis. The posterior hippocampi of taxi drivers were significantly larger relative to those of control subjects. A more anterior hippocampal region was larger in control subjects than in taxi drivers. Hippocampal volume correlated with the amount of time spent as a taxi driver (positively in the posterior and negatively in the anterior hippocampus). These data are in accordance with the idea that the posterior hippocampus stores a spatial representation of the environment and can expand regionally to accommodate elaboration of this representation in people with a high dependence on navigational skills. It seems that there is a capacity for local plastic change in the structure of the healthy adult human brain in response to environmental demands. PMID- 10716739 TI - Recognition of a conserved class of RNA tetraloops by Saccharomyces cerevisiae RNase III. AB - Ribonucleases III are double-stranded RNA (dsRNA) endonucleases required for the processing of a large number of prokaryotic and eukaryotic transcripts. Although the specificity of bacterial RNase III cleavage relies on antideterminants in the dsRNA, the molecular basis of eukaryotic RNase III specificity is unknown. All substrates of yeast RNase III (Rnt1p) are capped by terminal tetraloops showing the consensus AGNN and located within 13-16 bp to Rnt1p cleavage sites. We show that these tetraloops are essential for Rnt1p cleavage and that the distance to the tetraloop is the primary determinant of cleavage site selection. The presence of AGNN tetraloops also enhances Rnt1p binding, as shown by surface plasmon resonance monitoring and modification interference studies. These results define a paradigm of RNA loops and show that yeast RNase III behaves as a helical RNA ruler that recognizes these tetraloops and cleaves the dsRNA at a fixed distance to this RNA structure. These results also indicate that proteins belonging to the same class of RNA endonucleases require different structural elements for RNA cleavage. PMID- 10716740 TI - The RheA repressor is the thermosensor of the HSP18 heat shock response in Streptomyces albus. AB - Microorganisms have mechanisms to sense their environment and rapidly adapt to survive changes in conditions. In Streptomyces albus, various transcriptional repressors mediate the induction of heat shock genes. The RheA repressor regulates the synthesis of HSP18, a small heat shock protein, which plays a role in thermotolerance. The RheA protein was purified to determine how it responds rapidly to temperature. Gel retardation assays and footprinting experiments identified the specific target of RheA as an inverted repeat (TGTCATC 5N GATGACA) located in Phsp18, PrheA which is the common promoter region of the divergon. Gel retardation assays detected RheA-complexes formed with the hsp18-rheA promoters. The complexes did not form at higher temperature. In vitro transcription experiments showed that RheA is an autoregulatory protein and that its activity is inhibited by high temperature. The temperature-induced derepression by RheA is reversible. Dichroism circular spectroscopy revealed a reversible change of RheA conformation in relation with the temperature that could represent a transition between an active and an inactive form. Our experiments demonstrate that RheA acts as a cellular thermometer in hsp18 regulation. PMID- 10716741 TI - Epidemiologic methods in the search for causes of human birth defects. PMID- 10716742 TI - Preimplantation teratology and the placenta. PMID- 10716743 TI - Possible mechanism of congenital malformations induced by exposure of mouse preimplantation embryos to mitomycin C. AB - ICR mice were treated intraperitoneally with mitomycin C at 5 mg/kg on day 3 of gestation. On day 18 of gestation, fetuses of treated dams were inspected for external, skeletal and visceral malformations. At 6 or 12 hr after mitomycin C treatment, the blastocysts were obtained from the uteri of treated dams and the degenerated cells within inner cell mass (ICM) and trophectoderm (TE) tissues were examined microscopically. On day 5, 8, 11, or 18 of gestation, the uteri of treated dams were obtained and those including embryos/fetuses and placentae were examined histologically. Finally, on each of gestational days 5-14, the blood of the treated dams was collected and the hematological parameters determined. Pre- and postimplantation losses in the dams treated with mitomycin C were significantly increased; increased frequency of abdominal wall defects and lumbar ribs in term fetuses, decreased fetal weight, and increased placental weight were noted as well. No significant increase in visceral malformations was found in term fetuses treated with mitomycin C. Frequency of degenerated cells within ICM and TE of blastocysts from dams treated with mitomycin C was significantly increased as compared with the controls. In dams treated with mitomycin C, decidua developed insufficiently and the trophoblast giant cell layer was not separated from the uterine lumen by maternal components; hemorrhage from the denuded trophoblast giant cell layer into the uterine lumen was noted. The number of erythrocytes, as well as hemoglobin concentration, hematocrit, and the percentage of reticulocytes in blood of dams treated with mitomycin C were significantly lower from days 6-12 of gestation, as compared with controls. The results of the present study showed that an increase in number of degenerated cells within blastocysts results in preimplantation loss and both maternal and embryonic hypoxia during major organogenesis results in postimplantation loss and congenital fetal malformations. PMID- 10716744 TI - Hypoplastic basement membrane of the lens anlage in the inheritable lens aplastic mouse (lap mouse). AB - Adult homozygous lap mice show various eye abnormalities such as aphakia, retinal disorganization, and dysplasia of the cornea and anterior chamber. In the fetal eye of a homozygous lap mouse, the lens placode appears to develop normally. However, the lens vesicle develops abnormally to form a mass of cells without a cavity, and the mass vanishes soon afterward. Apoptotic cell death is associated with the disappearance of the lens anlage. We examined the basement membranes of the lens anlage of this mutant by immunohistochemical methods under light microscopy using antibodies against basement membrane components of the lens anlage, type IV collagen, fibronectin, laminin, heparan sulfate proteoglycan, and entactin and by transmission electron microscopy. Immunohistochemistry showed the distribution and intensity of antibody binding to the lens anlage to be almost the same for each these antibodies regardless of the stage of gestation or whether the anlagen were from normal BALB/c or lap mice. Thus, positive continuous reactions were observed around the exterior region of the lens anlage from day 10 of gestation for type IV collagen, fibronectin, laminin, heparan sulfate proteoglycan antibodies, and at least from day 11of gestation for entactin antibody. The basement membrane lamina densa of both normal and lap mice was shown by electron microscopy to be discontinuous at days 10 and 10.5 of gestation. However, by day 11 the lamina densa was continuous in the lens anlagen of normal mice but still discontinuous in the lap mice. By day 12 of gestation, the lamina densa had thickened markedly in normal mice, whereas in lap mice it remained discontinuous and its thinness indicated hypoplasia. These results indicate that, while all basement components examined are produced and deposited in the normal region of the lens anlage in the lap mouse, the basement membrane is, for some reason, imperfectly formed. The time at which hypoplasia of the basement membrane was observed in this mutant coincided with the stage during which apoptosis in the lens anlage occurred. This result may indicate a possibility of the relationship between the basement membrane and apoptosis in this mutant. PMID- 10716745 TI - Large-scale double-staining of rat fetal skeletons using Alizarin Red S and alcian blue. AB - A critical component in the conduct of a prenatal developmental toxicity study is the evaluation of fetal skeletal development. As the developing rodent fetus is typically evaluated at gestation day 20, at a time when ossification of the skeleton is incomplete, a thorough assessment of skeletal development would include both ossified and cartilaginous structures. Current methods to double stain the fetal skeleton using Alizarin Red S and Alcian Blue are typically described for small sample sizes or using time allotments for each processing step that are unsuitable for industry. In an industrial setting, there is a need for an effective means to double-stain fetal skeletons on a large scale (i.e., hundreds of fetuses simultaneously). This article describes a method used in our laboratory to stain both fetal bone and cartilage using solutions and procedures on an industrial scale. PMID- 10716746 TI - Digit effects produced by prenatal exposure to antiepileptic drugs. AB - The hypothesis tested was that digit anomalies among individuals exposed in utero to antiepileptic drugs (AED) are best identified by a systematic search, including radiographs and dermatoglyphics, rather than relying only on visual inspection. A systematic search was made for five types of digit abnormalities in 46 AED-exposed individuals ages 5-29 years in comparison with controls: visible anomalies, size of fingernails, dermal ridge patterns, length of metacarpals and phalanges, and qualitative changes in the distal phalanges. Among the AED exposed, nail size was not decreased. However, there was a 10.8% frequency of digit anomalies, a 12% frequency of three or more arch patterns, and significant shortening and qualitative changes in the distal phalanges, all of which are consistent with the fetal effects of AED. Among the 42 individuals who underwent all evaluations, 14.3% had two or more of these abnormalities, most of which would not be identified by clinical inspection. This frequency is much higher in these AED-exposed individuals than in the general population. Radiographs in 13 individuals over a period of several years showed that the changes in the phalanges and metacarpals persisted. PMID- 10716747 TI - Central nervous system in twin reversed arterial perfusion sequence with special reference to examination of the brain in acardius anceps. AB - The twin-reversed arterial perfusion (TRAP) sequence, or acardia, is the most severe complication in monozygotic twinning. Although more than 400 cases with TRAP sequence were reported since 1533, thorough investigations of the brain in those cases with a rudimentary head remained infrequent. We report a TRAP sequence with microcephaly and a severely rudimentary brain anlage. Neuropathologic examination clearly demonstrated two types of change: (1) developmental arrest of brain at the prosencephalic stage (holoprosencephaly), and (2) hypoxic damage to the holospheric brain mantle with cystic change (hydranencephaly). With reference to previous studies in experimental animals showing that lack of oxygen during early embryogenesis can induce severe disruptions of head-brain and heart formation, it is concluded that oxygen deficiency due to TRAP may be responsible not only for the encephaloclastic changes in the acardius anceps, but for the developmental arrest of the brain cases as well. This would make it unnecessary to postulate additional primary causes such as asymmetric zygote cleavage (Schwalbe, '07) for the maldevelopment. PMID- 10716748 TI - Teratogenic effect of ketamine and cocaine in CF-1 mice. AB - Pregnant CF-1 mice were used to study the teratogenic effect of ketamine and cocaine, alone and in combination. The dose of ketamine was 50 mg/kg and that of cocaine was 20 mg/kg, given intravenously (tail) once daily (these doses of ketamine and cocaine are comparable to doses used by addicted humans). Treatment was started from day 6 to day 15 of gestation, and dams were sacrificed on day 18. There were significant decreases in the fetal weight and length in the combined group. Skeletal defects such as incomplete ossification of skull bones and vertebrae were observed in both the cocaine and combined group, compared with the control. An increased frequency of cerebral and abdominal hemorrhages as well as hydrocephalus and hydronephrosis was observed in the combined group. This study showed that fetal exposure to ketamine and cocaine in combination was more teratogenic than each drug alone in CF-1 mice. PMID- 10716749 TI - Treatment with all-trans-retinoic acid decreases levels of endogenous TGF-beta(1) in the mesenchyme of the developing mouse inner ear. AB - BACKGROUND: Previous studies have shown that in utero exposure of the mouse embryo to high doses of all-trans-retinoic acid (atRA) produces defects of the developing inner ear and its surrounding cartilaginous capsule, while exposure of cultured periotic mesenchyme plus otic epithelium to high doses of exogenous atRA results in an inhibition of otic capsule chondrogenesis. METHODS: In this study, we examine the effects of atRA exposure on the endogenous expression of transforming growth factor-beta(1) (TGF-beta(1)), a signaling molecule that mediates the epithelial-mesenchymal interactions that guide the development of the capsule of the inner ear. RESULTS: Our results demonstrate a marked reduction in immunostaining for TGF-beta(1) in the periotic mesenchyme of atRA-exposed embryos of age E10.5 and E12 days in comparison with control specimens. Consistent with these in vivo findings, high-density cultures of E10.5 periotic mesenchyme plus otic epithelium, treated with doses of atRA that suppress chondrogenesis, showed significantly decreased levels of TGF-beta(1), as compared with TGF-beta(1) levels in untreated control cultures. Furthermore, we demonstrate a rescue of cultured periotic mesenchyme plus otic epithelium from atRA-induced chondrogenic suppression by supplementation of cultures with excess TGF-beta(1). CONCLUSIONS: Our results support the hypothesis that TGF-beta(1) plays a role in mechanisms of atRA teratogenicity during inner ear development. PMID- 10716750 TI - Ribonucleotide reductase subunit R1: a gene conferring sensitivity to valproic acid-induced neural tube defects in mice. AB - Neural tube defects (NTDs), although prevalent and easily diagnosed, are etiologically heterogeneous, rendering mechanistic interpretation problematic. To date, there is evidence that mammalian neural tube closure (NTC) initiates and fuses intermittently at four discrete locations. Disruption of this process at any of these four sites may lead to a region-specific NTDs, possibly arising through closure site-specific genetic mechanisms. Although recent efforts have focused on elucidating the genetic components of NTDs, a void persists regarding gene identification in closure site-specific neural tissue. To this end, experiments were conducted to identify neural tube closure site-specific genes that might confer regional sensitivity to teratogen-induced NTDs. Using an inbred mouse strain (SWV/Fnn) with a high susceptibility to VPA- induced NTDs that specifically targets and disrupts NTC between the prosencephalon and mesencephalon region (future fore/midbrain; neural tube closure site II), we identified a VPA-sensitive closure site II-specific clone. Sequencing of this clone from an SWV neural tube cDNA library confirmed that it encodes the r1 subunit of the cell cycle enzyme ribonucleotide reductase (RNR). The abundance of rnr-r1 mRNA was significantly increased in response to VPA drug treatment. This upregulated expression was accompanied by a significant decrease in cellular proliferation in the closure site II neural tube region of the embryos, as determined by ELISA cellular proliferation assays performed on BrdU-pulsed neuroepithelial cells in vivo. We hypothesize that rnr-r1 plays a critical role in the development of VPA-induced exencephaly. PMID- 10716752 TI - Genetic susceptibility to birth defects in humans: from gene discovery to public health action AB - Khoury MJ. 2000. Genetic susceptibility to birth defects in humans: from gene discovery to public health action. Teratology 61:17-20. PMID- 10716751 TI - Mini-review: history of organized teratology information services in North America. AB - A history of the Organization of Teratology Information Services (OTIS) is presented in context of the history of teratology information services. During the late 1970s, teratology information services grew out of the need for current and accurate information about fetal effects of environmental exposures in pregnancy. Over the next decade, teratology information services networked and collaborated, developing their own professional organization. A description of the activities of OTIS is described. PMID- 10716753 TI - Treatment of spasticity with botulinum toxin. PMID- 10716754 TI - Ulnar neuropathy at the elbow. PMID- 10716755 TI - Acetylcholine receptors and myasthenia. AB - Much progress has been made in the 26 years since initial studies of the first purified acetylcholine receptors (AChRs) led to the discovery that an antibody mediated autoimmune response to AChRs causes the muscular weakness and fatigability characteristic of myasthenia gravis (MG) and its animal model, experimental autoimmune myasthenia gravis (EAMG). Now, the structure of muscle AChRs is much better known. Monoclonal antibodies to muscle AChRs, developed as model autoantibodies for studies of EAMG, were used for initial purifications of neuronal AChRs, and now many homologous subunits of neuronal nicotinic AChRs have been cloned. There is a basic understanding of the pathological mechanisms by which autoantibodies to AChRs impair neuromuscular transmission. Immunodiagnostic assays for MG are used routinely. Nonspecific approaches to immunosuppressive therapy have been refined. However, fundamental mysteries remain regarding what initiates and sustains the autoimmune response to muscle AChRs and how to specifically suppress this autoimmune response using a practical therapy. Many rare congenital myasthenic syndromes have been elegantly shown to result from mutations in muscle AChRs. These studies have provided insights into AChR structure and function as well as into the pathological mechanisms of these diseases. Evidence has been found for autoimmune responses even to some central nervous system neurotransmitter receptors, but only one neuronal AChR has so far been implicated in an autoimmune disease. Thus far, only two neuronal AChR mutations have been found to be associated with a rare form of epilepsy, but many more neuronal AChR mutations will probably be found to be associated with disease in the years ahead. PMID- 10716756 TI - Diagnosis of ulnar neuropathy: a new approach. AB - Conventional electrodiagnosis used to detect an ulnar neuropathy at the elbow depends on accurate determination of ulnar nerve length across this segment. We present a new approach, using the difference in latency of the compound nerve action potentials (CNAPs) of the ulnar and median nerves elicited by stimulation at the wrist and recorded 10 cm above the elbow. Sixty normal controls were examined in order to determine the normal upper limit (1.4 ms) of the difference in CNAP latency of the ulnar and the median nerves (Dlat index). Values obtained in 10 patients with ulnar nerve lesions are discussed. This test was shown to be both sensitive and specific, was independent of ulnar nerve length, and was easy to perform. PMID- 10716757 TI - Spatial distribution of blood flow in electrically stimulated human muscle: a positron emission tomography study. AB - Neuromuscular electrical stimulation (NMES) was studied with positron emission tomography (PET) and H(2)(15)O in the quadriceps muscle of 11 men. The subjects were submitted to simultaneous bilateral isometric contraction (5 s)-rest (5 s) cycles for 12 min, with a workload corresponding to 5% of quadriceps maximal isometric voluntary torque (QMIVT) for one thigh (5%T) and 10% of QMIVT for the other (10%T). Scans were centered at the electrodes and tissue blood flow (TBF) was evaluated in square regions of interest (ROIs) (3.5 cm(2)) in the transverse section (TS) of both thighs. The mean TBF reached 8.9 mL min(-1) 100 g(-1) in the TS of the 5%T and 11.5 mL min(-1) 100 g(-1) in the TS of the 10%T (P > 0.05). A negative linear relationship was found for both thighs between the ROI-electrode distance and the TBF (P 5 mL min(-1) 100 g(-1)) was lower in the 5%T than in the 10%T (50.6% vs. 62.2%; P = 0.017). With NMES, the pattern of spatial recruitment appears linked to electrode proximity and is spatially extended. These results confirm the utility of combining NMES with voluntary exercise in the treatment of atrophied muscle. PMID- 10716758 TI - An overexpression of fibroblast growth factor (FGF) and FGF receptor 4 in a severe clinical phenotype of facioscapulohumeral muscular dystrophy. AB - We evaluated the expression of a select panel of growth factors and their receptors, including fibroblast growth factor 1 (FGF-1), fibroblast growth factor 2 (FGF-2), platelet-derived growth factor (PDGF), FGF receptor 1 (FGF-R1), FGF receptor 3 (FGF-R3), FGF receptor 4 (FGF-R4), PDGF receptor alpha (PDGF-Ralpha), PDGF receptor beta (PDGF-Rbeta), and heparan sulfate proteoglycan (HSPG), in muscle biopsy specimens from nine facioscapulohumeral muscular dystrophy (FSHD) patients using immunohistochemistry. Two cases of Duchenne-type muscular dystrophy (DMD), two of Becker-type muscular dystrophy (BMD), and one of limb girdle-type muscular dystrophy (LGMD) were also investigated. Widespread immunostaining for FGF-1 and FGF-2 on the sarcolemma and overexpression of FGF-R4 in endomysial and perimysial connective tissue were seen in one patient with a severe clinical phenotype of FSHD who had respiratory failure. Standard histochemistry in this patient revealed marked interstitial fibrosis and lobulated fibers. The overexpression of FGF and FGF-R4 in this severe FSHD case may be associated with the muscle fibrosis and disease severity. PMID- 10716759 TI - Effect of maturation on nerve excitability in an experimental model of threshold electrotonus. AB - Threshold electrotonus (TE) is a new tool for investigating axonal function noninvasively in vivo. To increase its potential clinical value, we developed a rat model of TE, and examined the effects of maturation and pharmacological intervention. We recorded TE in 92 male rats (body weight 90-650 g) by stimulating the motor nerve in the tail, and applying 100-ms conditioning currents. Motor conduction velocities increased up to a body weight of 330 g, and remained constant thereafter. TE in mature rats was similar to that in humans, and two parameters were analyzed: TEd(10-20) or the mean threshold reduction 10 20 ms after the onset of the depolarizing conditioning current at 40% of threshold intensity; and TEh(10-20) or the corresponding threshold decrease on hyperpolarization. Like latency, the absolute value of TEh(10-20) decreased up to 330 g, and then stabilized thereafter, probably reflecting the progressive increase in the axonal diameter and relative reduction in internodal impedance. In contrast, TEd(10-20) gradually decreased up to 330 g, and then jumped to a higher level, which was maintained for animals of >400 g. 4-Aminopyridine, a blocker of fast potassium channels, selectively increased TEd(10-20) only in the immature or young (<330 g) rats. This suggests that, in the mature animals, fast potassium channels become sequestrated from the nodal membrane and not activated in response to nodal depolarization. These findings indicate that mature rats (>400 g) may provide a useful experimental model for interpreting abnormal TE responses in humans, and provide evidence for nonlinear maturation of potassium channel function in myelinated axons. PMID- 10716760 TI - Marked increase in CD44-highly positive cells in hyperplastic thymuses from patients with Myasthenia gravis. AB - To investigate the role of the thymus in the pathogenesis of myasthenia gravis (MG), immunohistochemical expression of CD44, CD45R0, B7-1, and IL-2 was studied in: (1) hyperplastic thymuses of patients with MG whose symptoms markedly improved after thymectomy, (2) remnant thymuses of patients with MG whose symptoms did not respond to thymectomy, and (3) non-MG control thymus. Lymphocytes strongly expressing CD44, a marker for homing lymphocytes and activated memory lymphocytes in adults, were much more frequently observed in hyperplastic MG thymuses than in remnant thymuses and non-MG control thymuses. These CD44-highly positive cells in hyperplastic MG thymuses were for the most part located in the subcapsular and cortical areas but also occasionally in medullary areas. Some of these CD44-highly positive cells coexpress CD45R0. CD44 highly positive cells were located in the vicinity of blood vessels and thus appeared to have migrated directly from extralobular blood vessels. B7-1-positive cells and interleukin (IL)-2-positive cells were also more abundant in the MG patients than in controls and were localized in the proximity of CD44-highly positive cells. These findings suggest that mature T and B cells recirculate into hyperplastic MG thymus via CD44-associated mechanisms and are activated there. PMID- 10716761 TI - Quadriceps activation and perceived exertion during a high intensity, steady state contraction to failure. AB - The ability to sustain a high-intensity, steady-state muscle contraction may have differential effects on neuromuscular activation and perceived exertion. The purpose of this study was to examine changes in neuromuscular activation and perceived exertion at a near-maximal steady-state contraction of the quadriceps in healthy men. Seventeen healthy, college-aged male volunteers were studied during isometric contractions equivalent to 80% of the maximum voluntary contraction (MVC). Perceived exertion was measured with a modified category-ratio scale (CR-10). The CR-10 scale was anchored with one high anchor at 100% MVC and one low anchor at 10% MVC. Subjects then performed an 80% MVC for as long as they could sustain it. Subjects were asked to rate the feelings in their quadriceps every 5 s during the contraction. The results demonstrated significant increases in neuromuscular activation of the vastus medialis and vastus lateralis muscles (P < 0.05) during the 80% MVC, but there were no significant muscle by time interactions. The results also demonstrated a significant increase (P < 0.05) in perceived exertion during the 80% MVC. Neuromuscular activation of both muscles, and perceived exertion, were found to increase in linear (P < 0.05) and quadratic (P < 0.05) trends. Alterations in motor unit discharge properties or impairments in muscle fiber membrane excitability may account for nonlinear increases in vastii muscle activation and perceived exertion. PMID- 10716762 TI - Clenbuterol reduces degeneration of exercised or aged dystrophic (mdx) muscle. AB - Evidence of dystrophic muscle degeneration in the hind limb muscles of young (20 week-old) treadmill-exercised or aged (87-week-old) sedentary mdx mice was greatly reduced by treatment with clenbuterol, a beta(2)-adrenoceptor agonist. Daily treadmill exercise for 10 weeks increased the size of regions within the mdx plantaris but not the soleus or gastrocnemius muscles, in which necrotic muscle fibers or the absence of fibers was observed. Clenbuterol reduced the size of these abnormal regions from 21% of total muscle cross-sectional area to levels (4%) found in sedentary mdx mice. In addition, the muscles obtained from aged clenbuterol-treated mdx or wild-type mice did not display the extensive fibrosis or fiber loss observed in untreated mdx mice. These observations are consistent with a mechanism of dystrophic muscle degeneration caused by work load-induced injury that is cumulative with aging and is opposed by beta(2)-adrenoceptor activation. Optimization of beta(2)-agonist treatment of muscular dystrophy in mdx mice may lead to a useful therapeutic modality for human forms of the disease. PMID- 10716763 TI - Contractile properties of single muscle fibers in myotonic dystrophy. AB - Muscle fiber contractile dysfunction in myotonic dystrophy (MD) is poorly understood. We biopsied the tibialis anterior of two symptomatic and three asymptomatic subjects (aged 21-31 years) with the MD mutation. Biopsies were freeze dried. A total of 103 single muscle fibers were activated with Ca(++), allowing maximal force measurements and specific force (SF) estimates. The slack test was performed to calculate maximum unloaded shortening velocity (V(o)). The myosin heavy chain composition of each fiber was determined using sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). Type I and IIA fibers of all subjects had reduced SF when compared with healthy control subjects (P < 0.001). In addition, the type I fibers of symptomatic subjects generated less SF than those of asymptomatic subjects (P < 0.001). Type I fibers from asymptomatic and symptomatic subjects did not differ in V(o), but V(o) was lower than in control subjects (P < 0.001). There was no significant difference in V(o) of type IIA fibers from symptomatic, asymptomatic, and control subjects. These results indicate that the MD mutation leads to a diminished force-generating capacity of the myofilaments in both symptomatic and asymptomatic individuals. The results further suggest that reduction in force-generating capacity at the cellular level develops prior to clinical weakness. PMID- 10716764 TI - Lipomatosis, proximal myopathy, and the mitochondrial 8344 mutation. A lipid storage myopathy? AB - Multiple symmetric lipomatosis (MSL) has been related in some cases to the 8344 point mutation of the tRNA-lysine gene of the mitochondrial DNA, mainly in the context of families with classic myoclonic epilepsy with ragged-red fibers (MERRF) and exceptionally in patients with proximal myopathy as the only manifestation of mitochondrial disease. We report on two families harboring the 8344 mutation. The patients presented with MSL and myopathy, expressed as limb girdle weakness in index cases and as exercise intolerance in the others. All muscle biopsies performed showed lipid storage apart from RRF and respiratory chain complexes deficiency. A possible explanation for both adipose proliferation and lipid storage myopathy in these cases is a disturbance in intermediary lipid metabolism secondary to mitochondrial respiratory chain deficiency that could be related via carnitine deficiency. PMID- 10716765 TI - Amyotrophic lateral sclerosis IgG-treated neuromuscular junctions develop sensitivity to L-type calcium channel blocker. AB - In order to search for early changes induced by the application of human immunoglobulin G (IgG) on motor nerve terminals, IgG from patients with amyotrophic lateral sclerosis (ALS) and control subjects was injected subcutaneously into the levator auris muscle of mice. A week or a month after the last injection, endplate potentials were recorded. No changes in quantal content of transmitter release were observed. In control and ALS IgG-treated muscles, neurotransmitter release remained sensitive to P/Q-type and insensitive to N-type voltage-sensitive calcium channel (VSCC) blockers as in untreated muscles. In contrast, IgG from 5 of 8 different ALS patients induced a significant reduction in quantal content of the evoked response after incubation with nitrendipine, indicating that a novel sensitivity to this calcium channel blocker appears in these motor nerve terminals. These results indicate that ALS IgG induces plastic changes at nerve terminals. The expression of transmitter release coupled to L type VSCC indicate that ALS IgGs are capable of inducing plastic changes at the nerve terminals that may participate in the process leading to neuronal death. PMID- 10716766 TI - Immunoglobulin treatment in refractory Myasthenia gravis. AB - Failure to induce and maintain remission in severe exacerbations of myasthenia gravis (MG), despite optimal care, is a common problem. We evaluated the efficacy and safety of high-dose intravenous immunoglobulin (IVIg) therapy in an open label study of 10 patients with severe generalized myasthenia and an acute deterioration unresponsive to conventional therapy including high-dose corticosteroids, cyclosporine, and azathioprine. Intravenous Ig at a loading dose of 400 mg/kg was administered daily for 5 consecutive days, with maintenance IVIg treatment at a dose of 400 mg/kg, once every 6 weeks. Significant improvement occurred in all patients, beginning at 6 +/- 2 days of treatment as measured by the Osserman scale, fatigue variables, muscle strength, and respiratory function tests. No side effects were observed during induction of remission. Further IVIg treatments were highly efficacious in maintaining the remission. The severity of the disease decreased by 2.5 +/- 0.8 grades of the Osserman scale over a period of 1 year (P <0.001), in parallel with reduction of immunosuppressive therapy as well as a decrease in acetylcholine receptor antibody titers (P < 0.01). Intravenous Ig therapy seems to be highly potent for inducing rapid improvement in refractory myasthenia during acute deterioration as well as for maintaining remission. PMID- 10716767 TI - Bilateral phrenic neuropathy as a presenting feature of multifocal motor neuropathy with conduction block. AB - Diaphragmatic paralysis has previously been reported as a result of diverse pathologic processes involving the peripheral nervous system. We report the clinical history, physical findings, and antibody profile of an atypical case of multifocal motor neuropathy with conduction block initially presenting with respiratory failure secondary to bilateral phrenic neuropathy. PMID- 10716768 TI - Enhanced migration and fusion of donor myoblasts in dystrophic and normal host muscle. AB - Sliced male C57Bl/10Sn (H2-b) donor muscles were grafted into the female histocompatible muscles of untreated, FK506-treated, and T-cell depleted (with or without thymic tolerization) dystrophic (mdx; H2-b) and normal (C57Bl/10Sn; H2-b) hosts, and also into histoincompatible normal (Balb/c; H2-d) hosts. The fate of male donor nuclei was monitored on tissue sections by in situ hybridization with a Y-chromosome specific probe. The results demonstrate that the dystrophic environment is more conducive than normal muscle to donor myoblast migration, with the distance moved being threefold greater at 12 weeks in dystrophic hosts. T-cell depletion was significantly more effective than FK506 treatment at enhancing donor myoblast emigration in both histocompatible and histoincompatible hosts at 3 weeks. Furthermore, the effects of T-cell depletion were sustained in histoincompatible hosts at 12 weeks. These data endorse the use of host T-cell depletion as a promising long-term strategy to improve myoblast transfer therapy (MTT) in the clinical situation. PMID- 10716769 TI - Benign prognosis in idiopathic hyper-CK-emia. AB - We report on the long-term follow-up in 31 patients with idiopathic hyper-CK emia. At referral, all patients underwent a neurological interview and examination. Ancillary investigations included an open muscle biopsy and electromyography (EMG) in almost all, and other ancillary tests in some patients. After a follow-up period of 7.2 (mean; range 4-18) years, 74% of the patients had a final evaluation. The most common complaints at referral were fatigue and myalgia. EMG and muscle biopsy demonstrated minor, non-diagnostic abnormalities in 30 and 71% of patients, respectively. At follow-up, the pattern and the number of complaints had not changed substantially. One patient developed a sensory polyneuropathy. Neurological abnormalities were absent in all other patients. In conclusion, long-term follow-up of patients with idiopathic hyper-CK-emia does not reveal clinical deterioration. It seems justifiable to refrain from routine long-term follow-up in these patients. PMID- 10716770 TI - Type IV collagen and its degradation in paralyzed human muscle: effect of functional electrical stimulation. AB - The purpose of this study was to evaluate the effects of spinal cord injury (SCI) and functional electrical stimulation (FES) of paralyzed muscles on type IV collagen content and proteins involving its degradation, which is initiated by matrix metalloproteinase (MMP)-2 and -9 and regulated by their tissue inhibitors (TIMPs)-2 and -1. Ten SCI subjects participated in an 18-month program of functional electrical stimulation (FES) of their leg muscles. Needle biopsies were taken from the vastus lateralis muscle before and at various times during the training period, and from able-bodied controls. Type IV collagen concentration was unaltered. ProMMP-2 level of SCI subjects before the training period tended to be higher than able-bodied controls and was significantly above the control level after FES. MMP-9 concentration was unchanged. The results suggest accelerated type IV collagen turnover in skeletal muscle of SCI individuals especially after FES as a part of adaptive process of the muscle. PMID- 10716771 TI - Membrane skeleton of innervated and denervated fast- and slow-twitch muscle. AB - We used confocal microscopy and immunoblotting to study membrane skeletal proteins of fast-twitch (extensor digitorum longus) and slow-twitch (soleus) muscles of the adult rat. In the extensor digitorum longus (EDL), beta-spectrin concentrates in costameres, whereas dystrophin is enriched at costameres but is also present in intercostameric regions. In the soleus, beta-spectrin and dystrophin underlie much of the sarcolemma, and intercostameric regions are difficult to detect. The EDL sarcolemma reorganizes following denervation to resemble soleus sarcolemma, but denervation does not significantly affect the latter. Consistent with these observations, soleus contains similar amounts of dystrophin but more beta-spectrin than EDL. Denervation increases beta-spectrin levels only in the EDL and dystrophin levels in both muscles. Denervation does not affect beta-fodrin, a beta-spectrin homolog expressed in embryonic myofibers. Thus, neuromuscular activity controls sarcolemmal organization and the levels of beta-spectrin and dystrophin, but not postnatal downregulation of beta-fodrin. The differences in organization of the sarcolemma may underlie the differential susceptibility of fast and slow myofibers to dystrophinopathies. PMID- 10716772 TI - Steadiness is reduced and motor unit discharge is more variable in old adults. AB - The purpose of this study was to compare the steadiness and discharge rate of motor units during submaximal contractions performed by young and old adults. Subjects performed isometric and slow shortening and lengthening contractions with the first dorsal interosseous muscle. The steadiness of the isometric and slow anisometric contractions was less for the old subjects compared with young subjects, especially at the lower target forces and with the lightest loads. Furthermore, the steadiness of the lengthening contractions was less compared with the shortening contractions for the old subjects. Although the mean discharge rates of motor units were not different for the two groups of subjects, the variability of the discharge rates was greater for the old subjects during the isometric and anisometric contractions. We conclude that a more variable discharge by single motor units probably contributes to the reduced ability of old adults to perform steady muscle contractions. PMID- 10716773 TI - Post-intubation pulmonary function test in Guillain-Barre syndrome. AB - The value of daily postintubation respiratory function tests in predicting duration of ventilation in 37 mechanically ventilated patients with Guillain Barre syndrome (GBS) was studied. Patients ventilated for less than 3 weeks were compared with those ventilated more than 3 weeks. Daily vital capacity and maximal inspiratory and expiratory pressures were summed to an integrated pulmonary function (PF) score. We calculated the PF ratio, which represents the PF score at day 12 after intubation divided by the PF score on the day of intubation. The PF ratio was greater than 1 in all 10 patients ventilated less than 3 weeks and was less than 1 in 19 of 27 patients ventilated for longer (P = 0.0001, Fisher exact test). The sensitivity of a PF ratio less than 1 for predicting duration of ventilation greater than 3 weeks was 70%; the specificity and positive predictive value were 100%. This study thus suggests that serial postintubation respiratory tests can provide a measure of respiratory status in patients with GBS. These parameters may help predict duration of ventilation and need for tracheostomy. If, at day 12, the PF ratio is less than 1, it is highly unlikely that patients will be weaned within 3 weeks, and tracheostomy should be performed. If the ratio is greater than 1, tracheostomy should be deferred, because a substantial proportion of these patients may be successfully weaned from the ventilator within 3 weeks. PMID- 10716774 TI - Dynamics of nuclei of muscle fibers and connective tissue cells in normal and denervated rat muscles. AB - The mitotic activity in muscles of growing rats and the effect of denervation were studied by means of continuous infusion of 5-bromo-2-deoxyuridine (BRDU). Denervated muscles after 10 weeks contained 20 to 60% fewer muscle nuclei than normal; BRDU labeled about 25% of the nuclei of normal soleus and extensor digitorum longus (EDL) and of denervated EDL muscles but only 5% in the denervated soleus muscle. Labeled nuclei persisted in denervated but not in normal muscles. After the main growth period, the turnover of myonuclei was at most 1 to 2% per week. The behavior of connective tissue nuclei was similar to that in muscle fibers. Infusion of BRDU had no effect on contractile properties. It is suggested that the exceptionally rapid atrophy of the denervated rat soleus associated with loss of satellite cells was due to loss of myonuclei and differentiation and fusion of satellite cells. The cause may possibly be that the phase of postdenervation fibrillation is shorter than in other muscles. PMID- 10716775 TI - Phrenic nerve conduction studies in acute organophosphate poisoning. AB - Phrenic nerve conduction studies were performed within 48 h of admission and subsequently in 29 patients (14 of whom required mechanical ventilation) with acute organophosphate (OP) poisoning. The mean (+/-SD) amplitude of the diaphragmatic compound muscle action potential (CMAP) in patients requiring mechanical ventilation (119.09 +/- 173.85 microV) was significantly lower than in those not requiring mechanical ventilation (461.63 +/- 138.69 microV) (P < 0.0001). Diaphragmatic CMAP amplitudes in ventilated patients increased with time during the course of hospitalization and were normal in 5 (36%) patients and only mildly reduced in another 6 (43%) patients prior to discontinuation of mechanical ventilation, which was undertaken 4-18 days (mean 7 +/- 3 days) after poisoning. Eleven patients (79%) were successfully weaned from mechanical ventilation at the first attempt. In the 3 (21%) remaining patients, mechanical ventilation had to be reestablished because of weaning failure. The mean (+/-SD) diaphragmatic CMAP amplitude, prior to discontinuation of ventilatory assistance, was 242.6 +/- 94.1 microV in these 3 patients. After ventilatory discontinuation, it fell to 95.5 +/ 105.8 microV. Thus, reduced diaphragmatic CMAP amplitudes correlate with the need for mechanical ventilation in acute OP poisoning. PMID- 10716776 TI - FK506 promotes functional recovery in crushed rat sciatic nerve. AB - In this study we examine whether the systemic administration of FK506 or Cyclosporin A (CsA) expedited functional recovery following an axonotmetic nerve injury, and compared their effects in a rat model. Seventy-five adult Buffalo rats received a crush injury to the right posterior tibial nerve and subsequently underwent either no treatment (group I), daily injections of FK506 (group II), or daily injections of CsA (group III). Walking track analysis demonstrated return of hindlimb function by 20 days postoperatively in group I, 14 days in group II, and 18 days in group III. The blood-nerve barrier (BNB) was reconstituted by postoperative day (POD) 7 in both FK506- and CsA-treated animals and by POD 13 in control animals. These results suggest that recovery of function is more rapid with daily administration of FK506 than with CsA or no treatment, perhaps because of earlier restoration of the blood-nerve barrier. Agents that facilitate nerve regeneration have the potential to limit the extent of motor endplate loss and muscle atrophy seen with prolonged denervation, thereby limiting permanent functional loss. PMID- 10716777 TI - McArdle's disease presenting with asymmetric, late-onset arm weakness. AB - McArdle's disease or myophosphorylase deficiency is one of the most common muscle glycogenoses and typically presents in childhood or adolescence with exercise intolerance, myalgia, myoglobinuria, and cramps in exercising muscle. We describe an elderly man who developed asymmetric proximal arm weakness at age 73. He had no history of exercise-induced cramps, myalgias, or myoglobinuria. Creatine kinase levels were elevated, serum lactate did not rise on ischemic exercise testing, and muscle biopsy showed a vacuolar myopathy with absent myophosphorylase activity. This unusual case demonstrates that McArdle's disease may present with fixed, asymmetric proximal weakness at an advanced age and should be considered in this clinical setting, especially when a history of poor exercise tolerance can be elicited. PMID- 10716778 TI - Clinical features of flail arm syndrome. PMID- 10716779 TI - Mitochondriopathy, blepharospasm, and treatment with botulinum toxin. PMID- 10716780 TI - AAEM news and comments PMID- 10716790 TI - Brain and language in the millennium PMID- 10716791 TI - Brain, language, and environment. PMID- 10716792 TI - Language acquisition and language impairment: a comparative psycholinguistic approach. PMID- 10716793 TI - Neuropragmatics: brain and communication. PMID- 10716794 TI - The aphasias: fall and renaissance of the neurological model? PMID- 10716795 TI - Beyond phrenology: brain and language in the next millennium. PMID- 10716796 TI - Toward a theory of therapy for aphasia. PMID- 10716797 TI - Neural systems and language processing: toward a synthetic approach. PMID- 10716798 TI - How to move away from dualism. PMID- 10716799 TI - Neuropsychology in the next century: I wish, I wish. PMID- 10716800 TI - Perseveration in Y2K. PMID- 10716801 TI - Recovery from aphasia: why and how? PMID- 10716802 TI - Back to the future: diversity and coherence in brain and language studies. PMID- 10716803 TI - Not fractionating, converging. PMID- 10716804 TI - Working memory, long-term memory, and language processing: issues and future directions. PMID- 10716805 TI - Attentional modulation of language performance. PMID- 10716806 TI - Task-specificity and similarities in processing numbers and words: available data and future directions. PMID- 10716807 TI - The pursuit of brain-language relationships. PMID- 10716808 TI - It's about time. PMID- 10716809 TI - The developmental cognitive neuroscience of language: a new research domain. PMID- 10716810 TI - The role of learning and memory paradigms in the remediation of aphasic disorders. PMID- 10716811 TI - Brain, language, and linguistics. PMID- 10716812 TI - Frontiers of brain mapping of speech prosody. PMID- 10716813 TI - Aphasia treatment: a key issue for research into the twenty-first century. PMID- 10716814 TI - The neural substrate of the language faculty: suggestions for the future. PMID- 10716815 TI - Investigating the neural basis for language in the twenty-first century. PMID- 10716816 TI - What we shall know only tomorrow. PMID- 10716817 TI - Thinking outside the (black) box. PMID- 10716818 TI - Neuropharmacology and linguistic neuroplasticity. PMID- 10716819 TI - The third millennium: toward an age of psychoneural monism. PMID- 10716820 TI - The ineluctable and interdependent evolution of the concepts of language and aphasia. PMID- 10716821 TI - Voluntary consent for participation in research in the twenty-first century. PMID- 10716822 TI - Multilingual/multiliterate/multicultural studies of aphasia--the rosetta stone of neurolinguistics in the new millennium. PMID- 10716823 TI - Primary progressive aphasia-The future of neurolinguistic and biologic characterization. PMID- 10716824 TI - Inner speech and the inner life. PMID- 10716825 TI - Language studies in the third millennium. PMID- 10716826 TI - Multiple route plasticity. PMID- 10716827 TI - The learning brain. PMID- 10716828 TI - Quality of life with brain damage. PMID- 10716829 TI - From psychogeny to organicity: is the brain going to outgrow the psyche in the third millennium? PMID- 10716830 TI - Languages and aphasia. PMID- 10716831 TI - Planum of the apes: a case study. PMID- 10716832 TI - Word processing and verbal short-term memory: how are they connected and why do we want to know? PMID- 10716833 TI - Reconsidering the hegemony of linguistic explanations in aphasia: the challenge for the beginning of the millennium. PMID- 10716834 TI - It's time to face a simple question: why is canonical form simple? PMID- 10716835 TI - Back to the future: reclaiming aphasia from cognitive neurolinguistics. PMID- 10716836 TI - Advancing brain-language models in the next millennium. PMID- 10716837 TI - Invariance vs variability in aphasic performance. An example: agrammatism. PMID- 10716838 TI - Discourse analyses and dementia. PMID- 10716839 TI - On space, time, and language: for the next century, timing is (almost) everything. PMID- 10716840 TI - The neurolinguistics of bilingualism in the next decades. PMID- 10716841 TI - A century beyond Brodmann: new insights into cortical cytoarchitectonics and function. PMID- 10716842 TI - Paying attention to conversation. PMID- 10716843 TI - Language and reading as specific cases and specializations of auditory scene analysis: neurophysiologic evidence. PMID- 10716844 TI - Syntactic circuits: how does the brain create serial order in sentences? PMID- 10716845 TI - Aphasia treatment. On drugs, machines, and therapies: what will the future be? PMID- 10716846 TI - The organization of semantic memory: in support of a distributed model. PMID- 10716847 TI - Phylogeny, ontogeny, and microgeny in linguistic process: perception and action as progressive specification. PMID- 10716848 TI - The ontogeny of cerebral language dominance. PMID- 10716849 TI - Compound words: a challenge for aphasiology. PMID- 10716850 TI - Charting the time-course of language processing. PMID- 10716851 TI - The future of aphasia treatment. PMID- 10716852 TI - Neuropragmatics in the twenty-first century. PMID- 10716853 TI - Phonemic representation: a twenty-first century challenge. PMID- 10716854 TI - Understanding the behavioral-methodology/language-processing interface. PMID- 10716855 TI - The neurobiology of language recovery in aphasia. PMID- 10716856 TI - Discourse revisited: contributions of lexico-syntactic devices. PMID- 10716857 TI - Use of pharmacotherapy in the treatment of aphasia. PMID- 10716858 TI - Recurring utterances-how, where, and why are they generated? PMID- 10716859 TI - "Exceptional" case: what does it mean? PMID- 10716860 TI - Syntactic and semantic composition. PMID- 10716861 TI - Developmental aspects of verbal fluency and confrontation naming in children. AB - Developmental changes in children's verbal fluency and confrontation naming were explored in this study. One hundred and sixty children (ages 5 years and 11 months to 11 years and 4 months) completed two verbal fluency tasks (phonemic and semantic) and the Boston Naming Test (BNT). Normative data were compiled for the BNT and the phonemic and semantic fluency tasks. With the exception of the phonemic fluency task, all tests showed a linear increase from year-groups I to V, with a significant increase between year-groups I and II. Principal Component Factor Analysis was conducted to determine whether the tests evaluated similar or different functions. Two factors emerged: the first involving all of the measurements and the second explaining exclusively the phonemic fluency. These results make it possible to conclude that children also seem to have different subsystems responsible for the analysis and processing of different aspects of language. PMID- 10716862 TI - Verbal transformation: habituation or spreading activation? AB - Experiment 1 tested the habituation hypothesis of the Verbal Transformation Effect, an auditory illusion in which a repeating verbal stimulus undergoes perceptual transformation, by varying stimulus dimensions which might be expected to retard habituation. Transformations were found to increase as a function of imagery value and word length, failing to support the habituation hypothesis. Experiment 2, in which transformations were found to vary as a function of number of activated semantic representations of a physically invariant homophone stimulus, provided support for a new dual-process explanation of the Transformation effect, based on spreading activation between cognitive representations. PMID- 10716863 TI - Deep dyslexia is right-hemisphere reading. AB - Two views exist concerning the proper interpretation of the form of acquired dyslexia known as deep dyslexia: (a) that it represents reading by a multiply damaged left hemisphere reading system; (b) that it represents reading which relies extensively on right-hemisphere orthographic and semantic processing. Price, Howard, Patterson, Warburton, Friston, and Frackowiak (1998) have recently reported a brain-imaging study whose results, they claim, "preclude an explanation of deep dyslexia in terms of purely right-hemisphere word processing." Their claim conflicts with the conclusions of previous published work, which strongly supports the RH hypothesis, work which they do not mention. Furthermore, I argue that their own results also favor the RH hypothesis (even though they claim otherwise); indeed, their results permit the formulation of a much more detailed version of the RH hypothesis than has hitherto been possible. Hence I conclude that the right-hemisphere interpretation of deep dyslexic reading remains the preferred explanation of deep dyslexia. PMID- 10716865 TI - Predicting child language impairment from too many variables: overinterpreting stepwise discriminant function analysis. AB - Tallal (1999) defends an earlier presentation of 160 variables predicting classification of 59 children for specific language impairment (SLI) from the statistical criticisms of Zhang and Tomblin (1998). I present here 3 additional reasons why it is wrong to conclude from Tallal's original 1985 analysis and presentation that it is specifically temporal processing variables that underlie SLI. On the contrary, these data lend no support to this position. PMID- 10716866 TI - Generalizability of aphasiology studies: nontraditional populations examined. PMID- 10716864 TI - Right and left hemisphere cooperation for drawing predictive and coherence inferences during normal story comprehension. AB - In three experiments, healthy young participants listened to stories promoting inferences and named inference-related test words presented to the right visual field-Left Hemisphere (rvf-LH) or to the left visual field-Right Hemisphere (lvf RH). Participants showed priming for predictive inferences only for target words presented to the lvf-RH; in contrast, they showed priming for coherence inferences only for target words presented to the rvf-LH. These results, plus the fact that patients with RH brain damage have difficulty drawing coherence inferences and do not show inference-related priming, suggest that information capable of supporting predictive inferences is more likely to be initially activated in the RH than the LH, but following coherence breaks these concepts (now coherence inferences) are completed in the LH. These results are consistent with the theory that the RH engages in relatively coarse semantic coding, which aids full comprehension of discourse. PMID- 10716867 TI - Selective uppercase dysgraphia with loss of visual imagery of letter forms: a window on the organization of graphomotor patterns. AB - We report a patient who, after a left parieto-occipital lesion, showed alexia and selective dysgraphia for uppercase letters. He showed preserved oral spelling, associated with handwriting impairment in all written production; spontaneous writing, writing to dictation, real words, pseudowords, and single letters were affected. The great majority of errors were well-formed letter substitutions: most of them were located on the first position of each word, which the patient always wrote in uppercase (as he used to do before his illness). The patient also showed a complete inability to access the visual representation of letters. As demonstrated by a stroke segmentation analysis, letter substitutions followed a rule of graphomotor similarity. We propose that the patient's impairment was at the stage where selection of the specific graphomotor pattern for each letter is made and that the apparent selective disruption of capital case was due to a greater stroke similarity among letters belonging to the same case. We conclude that a visual format is necessary neither for spelling nor for handwriting. PMID- 10716868 TI - Selective attention fails to alter the dichotic listening lag effect: evidence that the lag effect is preattentional. AB - Berlin et al. (1973) reported that either stimulus from a dichotic pair of consonant-vowel syllables is processed preferentially when its presentation is delayed by 30-60 ms. In the first of three experiments with 60 normal right handed adults, we replicated the Berlin et al. "lag effect," but only for asynchronies between 60 and 90 ms. In Experiment 2 subjects focused attention selectively on one ear. The results indicated that focused attention and stimulus asynchrony have additive effects: Performance improved at the attended ear irrespective of stimulus asynchrony, but the lag effect remained unchanged relative to the divided-attention condition. Experiment 3 entailed a signal detection task that allowed separate analysis of detection and localization accuracy. As in previous studies, selective attention to one ear increased the accuracy of localization but not detection at the attended ear. Both dependent measures indicated a lag effect that remained invariant as attention was manipulated. These findings imply that the lag effect is attributable to a preattentional stage of auditory processing. PMID- 10716869 TI - Using fMRI to study recovery from acquired dysphasia. AB - We have used functional magnetic resonance imaging (fMRI) to characterize brain activations associated with two distinct language tasks performed by a 28-year old woman after partial recovery from dysphasia due to a left frontal hemispheric ischemic stroke. MRI showed that her ischemic lesion extended posteriorly from the left inferior frontal to the perisylvian cortex. fMRI scans of both language tasks revealed substantial differences in activation pattern relative to controls. The nature of this difference was task-specific. During performance of a verbal semantic decision task, the patient, in contrast to controls, activated a network of brain areas that excluded the inferior frontal gyrus (in either hemisphere). A second task involving rhyme judgment was designed to place a heavier cognitive load on language production processes and activated the left inferior frontal gyrus (Broca's area) strongly in normal controls. During this task, the most prominent frontal activation in the patient occurred in the right homologue of Broca's area. Subsequent analysis of this data by methods able to deal with responses of changing amplitude revealed additional, less sustained recruitment by the patient of cortex adjacent to the infarct in the region inferior to Broca's area during rhyming. These results suggest that in addition to changes in cognitive strategy, recovery from dysphasia could be mediated by both the preservation of neuronal networks in and around the infarct and the use of homologous regions in the contralateral hemisphere. PMID- 10716871 TI - Syntactic comprehension deficits in Alzheimer's disease. AB - Syntactic comprehension of German patients with dementia of the Alzheimer type was investigated and compared to healthy controls matched with respect to age, sex, and education. Special attention was directed at syntactic structures, which, in contrast to a language like English, are feasible in a grammatically rich language like German. In a sentence picture matching paradigm, only semantically reversible sentences were used. Syntactic complexity ranged from simple active voice sentences to more complex sentences like center-embedded object relative sentences. In comparison to their controls, patients showed a deficit in nearly all categories. Their performance was not influenced by age, but was heavily influenced by the degree of cognitive impairment. Patients with mild cognitive impairment, as defined by a MMSE score of 20 or higher, showed only slight difficulties in syntactic processing, whereas patients with moderate to severe impairment (MMSE < 20) did not perform above chance limits in most syntactic categories. It appears as though syntactic comprehension is only mildly affected in the early stages of Alzheimer's disease and is rather severely impaired in more advanced stages. In the present report, results are discussed in terms of working memory demands for syntactic processing. PMID- 10716870 TI - Online measures of basic language skills in children with early focal brain lesions. AB - Twenty children with early focal lesions were compared with 150 age-matched control subjects on 11 online measures of the basic skills underlying language processing, a digit span task, and 6 standardized measures. Although most of the children with brain injury scored within the normal range on the majority of the tasks, they also had a disproportionately high number of outlier scores on the reaction time tests. This evidence for a moderate impairment of the basic skills underlying language processing contrasts with other evidence suggesting that these children acquire normal control of the functional use of language. Furthermore, these children scored within the normal range on a measure of general cognitive ability, suggesting that there is no particular sparing of linguistic functions at the expense of general cognitive functions. Using the MPD procedure (Valdes-Perez & Pericliev, 1997), we found that the controls and the five clinical groups could be best distinguished with two measures of online processing (word repetition and visual number naming) and one standardized test subcomponent (the CELF Oral Directions subtest). The 12 children with left hemisphere lesions scored significantly lower than the 8 other children on the CELF-RS measure. Within the group of children with cerebral infarct, the nature of the processing disability could be linked fairly well to site of lesion. Otherwise, there was little relation between site or size of lesion and the pattern of deficit. These results support a model in which damage to the complex functional circuits subserving language leads to only minor deficits in process efficiency, because of the plasticity of developmental processes. PMID- 10716872 TI - The role of sublexical graphemic processing in reading. AB - Dual-route models of reading postulate the existence of two separate mechanisms: The lexical route allows words to be recognized in their holistic form, and the sublexical route proceeds by converting the written sublexical entities of a word or a nonword into their corresponding phonological equivalents. Sublexical reading is assumed to require three stages of processing: graphemic parsing, graphophonemic conversion, and phoneme blending. This study provides evidence in favor of the existence of a graphemic parsing process which occurs prior to grapheme-phoneme conversion. A group of normal subjects read nonwords which contained multiletter graphemes significantly more slowly than graphemically simple nonwords. These results, best interpretable in the context of a recent dual-route model of reading, confirm previous data obtained in pathology which suggest the functional independence of this cognitive procedure. PMID- 10716873 TI - The interaction of multiple routes in oral reading: evidence from dissociations in naming and oral reading in phonological dyslexia. AB - During oral reading we hypothesized that lexical representations are activated and selected for output by the simultaneous activation of the semantic, the direct lexical orthography to phonology, and the sublexical grapheme-to-phoneme conversion (GPC) routes (Southwood & Chatterjee, 1999). Serial models of reading argue that the semantic route governs oral reading with minimal influence from the nonlexical direct route and the sublexical GPC route. These models predict that semantic errors should occur in reading when the semantic route and GPC are both impaired. The Simultaneous Activation Hypothesis predicts few semantic errors in oral reading but many during picture naming. Semantic errors are infrequent in reading because information from all three reading routes constrains activation of a phonological entry. By contrast phonological selection in picture naming is constrained primarily by the semantic route and if damaged additional information is unavailable to select the appropriate phonological code. In agreement with the Simultaneous Activation Hypothesis five phonological dyslexics produced semantic errors during picture naming but not when reading aloud. Phonological errors were present during oral reading and minimal during picture naming. PMID- 10716874 TI - The effects of varying attentional demands on the word retrieval skills of adults with aphasia, right hemisphere brain damage, or no brain damage. AB - Adults with mild aphasia, right hemisphere brain damage (RBD), or no brain damage (NBD) provided one-word phrase completions under isolation, focused attention, and divided attention conditions and in response to relatively constrained or unconstrained phrase stems. Despite comparable word retrieval accuracy among groups during the isolation condition, aphasic and RBD groups performed less accurately than the NBD group during focused and divided attention conditions. Across conditions, there were no significant differences between aphasic and RBD groups. Only aphasic subjects demonstrated a significant effect of phrase type, responding more accurately when completing constrained versus unconstrained stimuli. For aphasic and RBD groups, error type analysis indicated that semantic and phonological aspects of word retrieval were influenced by increased attentional demands. These findings suggest that for adults with aphasia or RBD, there is a negative relation between attention impairments and word retrieval abilities. PMID- 10716875 TI - Distinguishing genuine from spurious causes: a coherence hypothesis. AB - Two opposing views have been proposed to explain how people distinguish genuine causes from spurious ones: the power view and the covariational view. This paper notes two phenomena that challenge both views. First, even when 1) there is no innate specific causal knowledge about a regularity (so that the power view does not apply) and 2) covariation cannot be computed while controlling for alternative causes (so that the covariation view should not apply), people are still able to systematically judge whether a regularity is causal. Second, when an alternative cause explains the effect, a spurious cause is judged to be spurious with greater confidence than otherwise (in both cases, no causal mechanism underlies the spurious cause). To fill the gap left by the traditional views, this paper proposes a new integration of these views. According to the coherence hypothesis, although a genuine cause and a spurious one may both covary with an effect in a way that does not imply causality at some level of abstraction, the categories to which these candidate causes belong covary with the effect differently at a more abstract level: one covariation implies causality; the other does not. Given this superordinate knowledge, the causal judgments of a reasoner who seeks to explain as much as possible with as few causal rules as possible will exhibit the properties that challenge the traditional views. Two experiments tested and supported the coherence hypothesis. Both experiments involved candidate causes that covary with an effect without implying causality at some level, manipulating whether covariation that implies causality has been acquired at a more abstract level. The experiments differed on whether an alternative cause explains the effect. PMID- 10716876 TI - In defense of representation. AB - The computational paradigm, which has dominated psychology and artificial intelligence since the cognitive revolution, has been a source of intense debate. Recently, several cognitive scientists have argued against this paradigm, not by objecting to computation, but rather by objecting to the notion of representation. Our analysis of these objections reveals that it is not the notion of representation per se that is causing the problem, but rather specific properties of representations as they are used in various psychological theories. Our analysis suggests that all theorists accept the idea that cognitive processing involves internal information-carrying states that mediate cognitive processing. These mediating states are a superordinate category of representations. We discuss five properties that can be added to mediating states and examine their importance in various cognitive models. Finally, three methodological lessons are drawn from our analysis and discussion. PMID- 10716877 TI - Regulation of CS1 fibronectin expression and function by IL-1 in endothelial cells. AB - VLA-4 is a critical adhesion molecule that regulates mononuclear cell trafficking to sites of inflammation. VCAM-1 is a primary ligand of VLA-4, although alternatively spliced fibronectin (FN) containing the CS1 region (CS1 FN) also binds to VLA-4. CS1 FN is expressed by rheumatoid arthritis (RA) synovial endothelial cells, but the factors that regulate CS1 FN expression are not known. We incubated human umbilical vein endothelial cells (HUVEC) with IL-1 (0.1-10 ng/ml) for 8-48 h and determined total FN and CS1 FN mRNA by Northern blot analysis. Both were constitutively expressed by HUVEC, and IL-1 increased total FN mRNA and the CS1-containing isoform (P < 0.05). IL-1 also increased CS1 FN protein expression on HUVEC as determined by Western blot analysis. An adhesion assay using (51)Cr-labeled Jurkat cells and IL-1-stimulated HUVEC was used to determine if IL-1-induced CS1 FN mediates cell binding. Cyclic CS1 peptide (10 microg/ml) blocked 49 +/- 5% of IL-1-induced Jurkat cell adhesion to HUVEC (P < 0.01), whereas anti-VCAM-1 antibody inhibited binding by only 35 +/- 5% (P < 0.01). CS1 peptide and anti-VCAM antibody treatment were not additive (50 +/- 7% inhibition), and 38 +/- 6% of new VLA-4-mediated adhesion to IL-1-treated HUVEC was due to an increase in CS1 FN. These data show that IL-1 increases CS1 FN expression by HUVEC and increases CS1-mediated cell adhesion. CS1 mimetics might have therapeutic efficacy by blocking recruitment of VLA-4-bearing cells. PMID- 10716878 TI - Effect of allotype on activation of neutrophils by FcgammaRIIIB cross-linking. AB - Human neutrophils constitutively synthesize two receptors for the constant region of IgG, FcgammaRII, and FcgammaRIIIB. Fluo-3-loaded neutrophils were treated with biotinylated Fab fragments of anti-FcgammaR antibodies and cross-linked with streptavidin, and intracellular calcium ([Ca2+](i)) was monitored by flow cytometry. Polymerization of filamentous actin was quantitated by NBD-phallacidin using flow cytometry. Cross-linking of FcgammaRII by monoclonal antibody (mAb) IV.3 induces an increase in [Ca2+](i), superoxide generation, and the polymerization of actin. [Ca2+](i) responses from cross-linking of FcgammaRIIIB by mAb 3G8 varied from minimal to no release. To determine whether discrepancies in 3G8-induced [Ca2+](i) release were due to allotype variation, we selected five donors who were homozygous for the NA1 allotype of FcgammaRIIIB and five who were either heterozygous or homozygous for the NA2 allotype and compared their [Ca2+](i) response and actin polymerization induced by FcgammaRIIIB cross linking. Cross-linking of FcgammaRIIIB by 3G8 produced minimal [Ca2+](i) release and polymerization of actin irrespective of donor allotype. PMID- 10716879 TI - In vitro characterization of five humanized OKT3 effector function variant antibodies. AB - Orthoclone OKT 3 (mOKT3) is a highly effective agent for the reversal of steroid resistant renal allograft rejection. However, its wider use has been limited by the development of a human anti-mouse antibody response (HAMA) and by the "cytokine release syndrome" (CRS). CRS has been associated with T cell/monocyte activation and, secondarily, with activation of the complement cascade. These processes are mediated through Abs' Fc regions by their abilities to cross-link T cells and mononuclear cells and to activate complements. To alleviate these problems, a group of five huIgG1- and huIgG4-based OKT3 wild-type antibodies and their corresponding Fc mutants with altered residues at amino acids 234, 235, and 318, reported to be required for FcgammaRI and FcgammaRII binding and complement fixation, were constructed. Characterization of these humanized OKT3 Abs, denoted huOKT3gamma1, huOKT3gamma4, huOKT3gamma1(A(234), A(235)), huOKT3gamma4(A(234), A(235)), and huOKT3gamma1(A(318)), has demonstrated that huOKT3gamma1(A(234), A(235)) and huOKT3gamma4(A(234), A(235)), and have at least a 100-fold reduced binding to FcgammaRI and FcgammaRII. As expected, they are much less potent in the induction of T cell activation and cytokine release, yet retain in vitro immunosuppressive effects as potent as those of mOKT3. Unexpectedly, while huOKT3gamma1(A(318)) did not show any reduction in its ability to bind C1q and to fix a complement, huOKT3gamma1(A(234), A(235)) was completely inactive. The in vitro characteristics of huOKT3gamma1(A(234), A(235)) are consistent with recent in vivo studies, in which this Ab showed greatly reduced HAMA and CRS with the retention of its ability to reverse ongoing graft rejection in man. PMID- 10716880 TI - B cells and antibodies in the pathogenesis of myelin injury in Semliki Forest Virus encephalomyelitis. AB - To determine the contribution of B cells to brain myelin injury in Semliki Forest Virus (SFV) encephalomyelitis, normal C57BL/6 (B6) and B-cell-deficient (C57BL/6 tm1Cgn) B6 mice were infected with SFV. The peak of clinical disease, i.e., the time at which the greatest proportions of mice had moderate to severe clinical signs, appeared earlier in B6 mice [day 7 postinfection (pi)] than in B-cell deficient mice (day 21 pi). By flow cytometry, no clear differences were found in the percentages of CD3(+)CD4(+) T cells in the brains of B6 and B-cell-deficient mice. However, by day 21 pi, percentages of CD3(+)CD8(+) T cells were greater in brains of B-cell-deficient than in those of B6 mice. On day 21 pi, percentages of CD19(+) B cells were maximal in B6 mice, but B cells were absent in B-cell deficient mice at all time points. Sera obtained from B6 mice showed antibody responses to SFV, to SFV E2 peptides p137-151 and p115-133, and to peptides of myelin oligodendrocyte glycoprotein p18-32 and myelin basic protein (MBP) p64-75. Sera obtained from B-cell-deficient mice showed minimal or no reactivity to SFV, E2, or myelin peptides. CNS inflammatory and PAS-positive macrophage foci were maximal on days 7-14 pi in all mice. Additionally, B6 mice had brain white matter vacuolation, whereas B-cell-deficient mice did not. These data suggest that brain infiltrating B cells and anti-myelin antibodies contribute to myelin injury in SFV encephalomyelitis. PMID- 10716881 TI - Lectin ligands on human dendritic cells and identification of a peanut agglutinin positive subset in blood. AB - As only a few cell surface markers for dendritic cells (DC) have been identified to date, this study examined the expression of ligands for lectin on different human DC populations. The ability of Concanavalin A (Con A), Wheat Germ Agglutinin (WGA), peanut agglutinin (PNA), and Helix pomatia (HPA) to bind to cell lines and PBMC and DC populations was analyzed by flow cytometry and specificity of binding confirmed using inhibitory and noninhibitory sugars. The cell lines showed non-lineage-restricted binding with Con A and WGA, independent of sialidase treatment. HPA and PNA bound to a restricted number of lines, but showed broad reactivity after sialidase treatment. The peripheral blood mononuclear cells (PBMC) and directly isolated blood DC, activated CD83(+) blood DC, epidermal Langerhans cells (LC), and monocyte-derived DC (Mo-DC) showed strong binding of Con A and WGA, both before and after sialidase treatment. No HPA binding ligands were detected on PBMC populations, including directly isolated blood DC. Following sialidase treatment CD3(+), CD16(+), and a subset of CD19(+) lymphocytes bound HPA. The lectin PNA bound weakly to CD14(+) monocytes and a subpopulation of circulating DC that were HLA-DR(hi)CDw123 Dr(hi)CDw123(dim)/(neg)CD11c(+). The HLA-DR(mod)CDw123(hi)CD11c(neg) subpopulation did not bind PNA. Without sialidase treatment LC expressed both HPA and PNA ligands, but these were either absent on activated CD83(+) blood DC or weakly expressed on Mo-DC. Following sialidase treatment PBMC populations, activated CD83(+) blood DC, and Mo-DC became PNA positive. Thus human DC express several lectin ligands and PNA binding identifies a subset of blood DC. That may reflect discrete changes associated with stages of DC development or functional maturation. PMID- 10716882 TI - Differential control of neonatal tolerance by antigen dose versus extended exposure and adjuvant. AB - Ig-PLP1, an immunoglobulin (Ig) chimera carrying the encephalitogenic proteolipid protein (PLP) sequence 139-151 (PLP1), induces neonatal tolerance in mice and confers resistance to experimental allergic encephalomyelitis (EAE) without the need for incomplete Freund's adjuvant (IFA). The mechanism underlying such tolerance involves organ-specific T cell regulation characterized by lymph node deviation and an unusual IFNgamma-dependent splenic anergy. This form of T cell modulation may prove useful for prevention of autoimmunity. However, since the neonatal period is susceptible to regulation, further investigations are necessary to define parameters required to establish regimens suitable for optimal protection against disease. Therefore, studies were carried out to investigate the effect that IFA, the dose of Ig-PLP1, and the number of Ig-PLP1 injections might have on Ig-PLP1-mediated neonatal tolerance and protection against disease. Herein it is reported that as little as 1 microg of Ig-PLP1 supported IFNgamma-dependent splenic anergy but lymph node deviation and protection against disease strengthened as the dose of tolerogen increased. However, when a two-injection regimen was applied, resistance to disease was observed but the mechanism manifested proliferative and cytokine unresponsiveness in both lymphoid organs. Furthermore, the use of IFA along with Ig-PLP1 yielded a suppressive mechanism similar to that of the two-injection regimen. Therefore, the dose of Ig-PLP1 displays a quantitative influence, while the number of injections of Ig-PLP1 and the presence of IFA rather drive qualitative influences on such tolerance. PMID- 10716883 TI - The CD40-inducible Bcl-2 family member A1 protects B cells from antigen receptor mediated apoptosis. AB - CD40 activation is necessary for thymus-dependent humoral immune responses and rescuing both phenotypically immature WEHI-231 B lymphoma cells from B cell antigen receptor-induced cell death and germinal center B cells from spontaneous apoptosis. As some effects of CD40 are probably mediated by differences in gene expression, cDNA expression arrays and RNase protection assays were used to identify the anti-apoptotic Bcl-2 homolog A1 as a CD40-inducible gene in B cell lines and purified germinal center B cells. Sustained CD40-induced A1 upregulation correlated with CD40-mediated rescue of WEHI-231 cells from anti-IgM induced apoptosis. Moreover, overexpression of A1 specifically protected WEHI-231 cells from anti-IgM-induced apoptosis but not cell death triggered by certain other stimuli. PMID- 10716884 TI - Acidosis induces necrosis and apoptosis of cultured hippocampal neurons. AB - Acidosis, hypoxia, and hypoglycemia rapidly and transiently appear after reduction of cerebral blood flow. Acidosis also accompanies head trauma and subarachnoid hemorrhage. These insults result in necrotic and apoptotic loss of neurons. We previously demonstrated that transient acidification of intracellular pH from 7.3 to 6.5 induces delayed neuronal loss in cultured hippocampal slices (49). We now report that acidosis induced both necrotic and apoptotic loss of neurons. Necrosis and apoptosis were distinguished temporally and pharmacologically. Necrosis appeared rapidly and was dose dependent with the duration of the acidosis treatment. Apoptosis was delayed with maximal number of apoptotic cells seen with a 30-min acidosis treatment. Apoptotic neuronal loss was accompanied by DNA fragmentation and was blocked by inhibitors of protein and RNA synthesis, ectopic expression of the anti-apoptotic gene bcl-2, or an inhibitor of caspases, proteases known to be activated during apoptosis. Necrotic neuronal loss was unaffected by these treatments. Hypothermia, a treatment known to attenuate neuronal loss following a variety of insults, blocked both acidosis induced necrosis and apoptosis. These results indicate that acidosis is neurotoxic in vitro and suggest that acidosis contributes to both necrotic and apoptotic neuronal loss in vivo. PMID- 10716885 TI - The effects of bone morphogenetic protein 2 and 4 (BMP2 and BMP4) on gap junctions during neurodevelopment. AB - Nervous system deficits account for the third largest group of fatal birth defects (after heart and respiratory problems) in North America. Although considerable advance has been made in neuroscience research, the early events involved in neurogenesis remain to be elucidated. More specifically, the effects of signaling molecules on intercellular communication during neurodevelopment have not yet been studied. The development of the central nervous system is regulated, at least in part, by signaling molecules such as bone morphogenetic proteins (BMPs). In this study, we have used the embryonal mouse P19 cell line to examine the effects of BMP2 and BMP4 on gap junctional communication as well as neuronal and astrocytic differentiation. The undifferentiated P19 cells show high levels of the gap junction protein, connexin43 (Cx43), and functional intercellular coupling. However, Cx43 expression and dye coupling decrease as these cells differentiate into neurons and astrocytes. In contrast, cells treated with BMP2 or BMP4 lose their capacity to differentiate into neurons but not astrocytes, while they maintain extensive gap junctional communication. The very few neurons that remain in the BMP-treated cultures are coupled (a characteristic not seen in the control neurons). Together, our data suggest that BMPs may play a critical role in morphogenesis of P19 cells while they affect gap junctions. PMID- 10716886 TI - Metallothionein expression is altered in a transgenic murine model of familial amyotrophic lateral sclerosis. AB - Missense mutations in the gene encoding copper zinc superoxide dismutase (SOD1) have been found to cause one form of familial amyotrophic lateral sclerosis (FALS). Although the exact mechanism of disease is unknown, abnormalities in the ability of mutant SOD1 to bind zinc or copper ions may be crucial in the pathogenesis of disease. Because members of the metallothionein (MT) family of zinc and copper binding proteins function as important cellular regulators of metal ion bioavailability in the central nervous system, we used in situ hybridization and immunohistochemistry to study the expression pattern of these molecules in a transgenic mouse model of familial ALS. In adult wild-type mouse spinal cord, expression of MT-I and MT-II is restricted to ependymal cells and a subset of astrocytes located in white matter tracts, while MT-III synthesis is limited to neurons within gray matter. Compared to wild-type littermates, transgenic mice carrying the G93A SOD1 mutation demonstrate markedly increased expression of MT-I and MT-II within astrocytes in both white and gray matter as weakness develops. MT-III synthesis in neurons is also greatly upregulated as G93A SOD1 animals age, with glial cell expression of MT-III evident by later stages of the disease. Changes in MT expression occur before the onset of motor deficits or significant motor neuron pathology in G93A SOD1 mice and remarkably extend beyond ventral horn populations of neurons and glia. These results are consistent with the hypothesis that metallothioneins may serve an early and important protective function in FALS. PMID- 10716887 TI - Abnormal mitochondrial morphology in sporadic Parkinson's and Alzheimer's disease cybrid cell lines. AB - Diseases linked to defective mitochondrial function are characterized by morphologically abnormal, swollen mitochondria with distorted cristae. Several lines of evidence now suggest that sporadic forms of Parkinson's disease (PD) and Alzheimer's disease (AD) are linked to mitochondrial dysfunction arising from defects in mitochondrial DNA (mtDNA). Human neuroblastoma (SH-SY5Y) cells that are deficient in mtDNA (Rho(0)) were repopulated with mitochondria from AD or PD patients or age-matched controls. These cytoplasmic hybrid (cybrid) cell lines differ only in the source of their mtDNA. Differences between cybrid cell lines therefore arise from differences in mtDNA and provide a model for the study of how impaired mitochondrial function alters the mitochondria themselves and how these changes adversely affect the neuronal cells they occupy. Cybrid cell mitochondria were labeled with the mitochondrial membrane potential-sensitive dye, JC-1. Analysis of these JC-1 labeled mitochondria by confocal microscopy revealed that mitochondrial membrane potential was significantly reduced in both PD and AD cybrid cells when compared with controls. Ultrastructural examination showed that control cybrid cells contained small, morphologically normal, round or oval mitochondria with a dark matrix and regular distribution of cristae. PD cybrid cells contained a significant and increased percentage of mitochondria that were enlarged or swollen and had a pale matrix with few remaining cristae (0.26-0.65 microm(2)). AD cybrid cells also contained a significantly increased percentage of enlarged or swollen mitochondria (0.25-5.0 microm(2)) that had a pale matrix and few remaining cristae. Other pathological features such as crystal-like intramitochondrial inclusions and cytoplasmic inclusion bodies were also found in PD and AD cybrids. These observations suggest that transfer of PD or AD mtDNA into Rho(0) cells was sufficient to produce pathological changes in mitochondrial ultrastructure that are similar to those seen in other mitochondrial disorders. These data were reported in abstract form (Trimmer et al., 1998, Soc. Neurosci. Abstr. 24: 476). PMID- 10716889 TI - Diffusion and high resolution MRI of traumatic brain injury in rats: time course and correlation with histology. AB - Although widely employed in studies of cerebral ischemia, the use of diffusion weighted imaging (DWI) for traumatic brain injury (TBI) has been both limited and primarily confined to the first few hours after injury. Therefore, the present study examined the temporal evolution of magnetic resonance imaging (MRI) signal changes from hours to weeks after moderate fluid-percussion TBI in rats. We used isotropic diffusion along three directions and high resolution (HR) spin-echo pulse sequences to visualize DWI and HR MRI changes, respectively. Late changes were compared to histopathological and neurological outcome. A significant decrease (P<0.05) in the apparent diffusion coefficients (ADC) below preinjury levels was found in the left cortex and left hippocampus (ipsilateral to injury) at 1-2 h post-TBI. At 2 weeks post-TBI, ADCs were significantly elevated (P<0.05) above preinjury levels in both cortex and hippocampus. Regions of hypo- and hyperintensity detected in HR MRI scans also showed evidence of tissue damage by histological evaluation. Neurological assessment indicated that such changes were observed at a level of injury which produced moderate impairment 2 weeks after the insult. These results indicate that alterations in DWI and HR MRI signals occur both early (hours) and late (weeks) after lateral fluid-percussion injury. Furthermore, the study showed that DWI was sensitive to MR signal change at 1-2 h post TBI (in select ROIs), whereas HR scans showed MR signal change primarily at later time points (3-4 h and later). Moreover, regions which demonstrate late changes are associated with histological damage and neurological impairment. The study demonstrates the utility of MRI to detect early changes, in some cases, that are predictive of long-lasting damage verified histologically. PMID- 10716888 TI - A possible model of senile plaques using synthetic amyloid beta-protein and rat glial culture. AB - The senile plaque (SP) is one of the pathological hallmarks in the brains of patients with Alzheimer's disease (AD), but the mechanism of its formation and its role in AD progression are not yet fully understood. Synthetic amyloid beta protein (Abeta)1-40 is known to aggregate in vitro, and the aggregated Abeta has been widely used for in vitro experiments, in which its peculiar effects on neuronal and glial cells have been shown. To date, however, the formation of a SP like structure in a culture system using synthetic Abeta has not been demonstrated. In this study, we established a possible SP model using synthetic Abeta1-40 and rat glial cultures as follows: (1) large spherical aggregates of synthetic Abeta (sAmys) were produced from synthetic Abeta1-40 (10-50 microm in diameter), (2) the sAmys were added to a glial culture, and (3) the characteristics of the sAmys and the reactions of glial cells (microglia and astrocytes) around the sAmys were analyzed. We found that the sAmys exhibited the same features as the dense amyloid core in SPs, including the intense green birefringence under polarized light with Congo red, and induced reactive features in glial cells, including induction of major histocompatibility complex class II antigen in the microglia and interleukin-1beta in the astrocytes, similar to those seen in SPs in the brain in AD. Given our findings, we consider that this glial culture system with the sAmys is a possible in vitro SP model and useful for investigating the effects of massive amyloid deposition on neuronal and glial cells. PMID- 10716890 TI - Despite the internucleosomal cleavage of DNA, reactive oxygen species do not produce other markers of apoptosis in cultured neurons. AB - The cell death induced by hydroxyl radicals generated by Cu-phenanthroline and peroxynitrite generated by 3-morpholinosydnonimine hydrochloride (SIN-1) in rat primary cortical neuronal cultures was compared with the apoptotic death induced by staurosporine and the necrotic death induced by glutamate. Both SIN-1 and Cu phenanthroline were capable of generating internucleosomal cleavage of DNA-a hallmark of apoptosis. Other characteristics of this cell death, such as nuclear morphology by light microscopy; DNA breaks by single-cell gel electrophoresis; the effects of the apoptotic inhibitors cycloheximide, aurintricarboxylic acid, and tosyl-l-lysine chloromethyl ketone; the measurement of caspase activity; and the effects of antioxidants, were then analyzed. The conclusion from these hallmarks of apoptosis is that the cell death induced by these reactive oxygen species is not apoptosis. PMID- 10716891 TI - A serum factor enhances production of nitric oxide and tumor necrosis factor alpha from cultured microglia. AB - The pathological activation of microglia has been implicated in ischemic neuronal damage and some neurodegenerative diseases; however, the mechanism of microglial activation is not well understood. Previously, we showed that a serum factor, albumin, increased O(2)(-) production by cultured microglia (Si et al., 1997, Glia 21: 413-418). In the present study, we found that serum also enhanced lipopolysaccharide (LPS)-induced production of nitric oxide and tumor necrosis factor-alpha, which are other important neurotoxins released by activated microglia. In the presence of 0.1% normal rat serum, the half-effective concentration for LPS decreased from 300 to 1 ng/ml. The factor seemed to be a relatively high-molecular-weight protein because the factor was retained after a molecular sieve (50 kDa) membrane separation. The factor was labile to trypsinization and heat treatment at 72 degrees C for 5 min but was stable at 56 degrees C for 60 min. Several purified serum proteins including albumin could not mimic the enhancing effect of serum. Acute-phase serum showed a potent enhancing effect at a 10 times lower concentration than the normal serum. By gel filtration chromatography, the enhancing effect observed was a single peak at about 60 kDa. These results suggest that some serum protein infiltrates into brain parenchyma after blood-brain barrier disruption and such protein may result in neuronal damage by activating microglia to release neurotoxins in some central nervous system diseases. PMID- 10716892 TI - Delayed rat facial nerve repair leads to accelerated and enhanced muscle reinnervation with reduced collateral axonal sprouting during a definite denervation period using a cross-anastomosis paradigm. AB - To establish the influence of prolonged denervation on the recovery of a motor nerve, the rat facial nerve was transected and denervated for 0 to 224 days. Then, the freshly transected hypoglossal nerve was sutured to the predegenerated facial nerve (hypoglossal-facial nerve anastomosis, HFA). Using this nerve cross anastomosis paradigm we analyzed the nerve regeneration and muscle reinnervation 7 to 112 days post-suture operation (DPSO). After HRP injection into the whiskerpad 931+/-27 hypoglossal neurons were labeled at 112 DPSO after immediate HFA. Following 14 to 112 days denervation the number of labeled neurons increased to 138% (14 days delay), 154% (56 days), and 145% (112 days). In contrast, the reinnervation was poorer after 7 days denervation with the number of neurons increasing to 84%, and after long-term denervation of 224 days the number of neurons increased to 81%. The increase in amplitude of evoked electromyography wave after nerve suture correlated with the number of labeled neurons. After immediate HFA each regenerated motoneuron established on average 5.1 myelinated sprouts at 112 DPSO. The number of sprouts remained constant after delayed suture of 14 to 112 days, whereas the slower reinnervation after 7 or 224 days delay was accompanied by a massive sprouting of 9.1 or 8.1, respectively, sprouts per neuron. The muscles showed recovery after any denervation time. The muscle cross sectional area continuously decreased with longer denervation time. This decrease was only significant after 224 days denervation (67% of the normal value). We conclude that motor nerve reconstruction achieves better functional results after a definite period of denervation when using a nerve cross-anastomosis paradigm. PMID- 10716893 TI - Increased expression of mRNA encoding ferritin heavy chain in brain structures of a rat model of absence epilepsy. AB - Differential mRNA display was carried out to find genes that are differentially regulated in the brain of a rat strain with absence epilepsy, the genetic absence epilepsy rats from Strasbourg (GAERS). Among the 32 differentially displayed cDNA fragments actually cloned and sequenced, one shows 100% identity with the rat heavy chain ferritin (H-ferritin) mRNA. Northern blot analysis confirmed the up regulation of the H-ferritin mRNA. Using dot blotting, a 40% increase in expression was reported in the subcortical forebrain of the adult GAERS, while cortex, brain stem, and cerebellum appeared unmodified. This change was not observed in the brain of 25-day-old rats, an age at which the epileptic phenotype is not present. By in situ hybridization, the enhanced expression was localized in the hippocampus. The increase in mRNA encoding H-ferritin was not immunodetected at the protein level by Western blotting. These results are not apparently related to the neural substrate of SWD or to the distribution of local increase in glucose metabolism previously described in the GAERS. It is hypothesized that the up-regulation of the H-ferritin mRNA is part of a mechanism protecting the hippocampus, a seizure-prone area, against a possible overactivation during absence seizures. PMID- 10716894 TI - Role for basic fibroblast growth factor (FGF-2) in tyrosine kinase (TrkB) expression in the early development and innervation of the auditory receptor: in vitro and in situ studies. AB - A previous study showed that basic fibroblast growth factor (FGF-2) promotes the effects of brain-derived neurotrophic factor (BDNF) on migration and neurite outgrowth from the cochleovestibular ganglion (CVG). This suggests that FGF-2 may up-regulate the receptor for BDNF. Thus we have examined TrkB expression during CVG formation and otic innervation in vitro and in the chicken embryo using immunohistochemistry. Following anatomical staging according to Hamburger Hamilton, results were compared with mRNA expression in vitro using in situ hybridization. In the embryo at stage 16 (E2+) clusters of either lightly stained or immunonegative cells occurred within the otocyst and among those migrating to the CVG. By stage 22 (E3.5), immunostaining was concentrated in the CVG perikarya and invaded the processes growing into the otic epithelium but not into the rhombencephalon. Subsequently TrkB expression decreased in the perikarya and became localized in the leading processes of the fibers invading the epithelium and in the structures participating in synapse formation with the hair cells. In vitro there was moderate immunostaining and modest in situ hybridization for trkB in the neuroblasts migrating from the otocyst under control conditions. In contrast, neuroblasts previously exposed to FGF-2 exhibited accelerated migration and differentiation, with increased trkB mRNA expression. Morphological differentiation was associated with more intense immunostaining of processes than cell bodies. Evidently TrkB shifts its expression sequentially from sites engaged in migration, ganglion cell differentiation, axonal outgrowth, epithelial innervation, and synapse formation. FGF-2 may promote the role of BDNF in these developmental events by upregulating the TrkB receptor. PMID- 10716895 TI - Progressive metabolic changes underlying the chronic reorganization of brain circuits during the silent phase of the lithium-pilocarpine model of epilepsy in the immature and adult Rat. AB - The lithium-pilocarpine (Li-Pilo) model of epilepsy reproduces most of the features of human temporal lobe epilepsy. In the present study, we explored the correlation between metabolic changes, neuronal damage, and epileptogenesis during the silent phase following status epilepticus (SE) induced by Li-Pilo in 10- (P10) and 21-day-old (P21) and adult rats. Cerebral metabolic rates for glucose (CMR(glcs)) were measured at 14 and 60 days after SE by the 2 [(14)C]deoxyglucose method and neurodegeneration was assessed by the silver staining and cresyl violet techniques. In P10 rats, there was no damage and no metabolic consequences at any time after SE. In P21 rats, metabolic decreases were recorded at 14 days after SE, mainly in damaged forebrain regions. Conversely at 60 days after SE, P21 rats exhibited metabolic increases in both forebrain-damaged and brain-stem-intact areas. Finally, in adult rats studied at 14 days after SE, CMR(glcs) decreased in damaged forebrain areas involved in the circuitry of spontaneous seizures and increased in nondamaged brain-stem areas involved in the remote control of epilepsy. The increase in CMR(glcs) in damaged forebrain areas of P21 rats at 60 days after SE may reflect the genesis of a new circuitry underlying the occurrence of spontaneous seizures. The metabolic increase recorded in nondamaged brain-stem areas of P21 and adult rats occurs in regions involved in the remote control of seizures and might underlie a process of protection against the occurrence of seizures. PMID- 10716896 TI - Amyloid-beta injection in rat amygdala alters tau protein but not mRNA expression. AB - Previously we demonstrated local and distant changes in tau protein immunoreactivity reminiscent of those seen in Alzheimer's disease (AD) following a unilateral injection of amyloid-beta (Abeta)(25-35) into the rat amygdala. To explore the relevance of these findings to AD, we compared the effects of Abeta(1 42) to those of Abeta(25-35). Injections of both Abeta(1-42) and Abeta(25-35) into rat amygdala resulted in increased tau-2 immunolabeling in neurons. To determine whether these alterations were due to changes in the expression of tau, we measured tau protein expression by Western blotting and tau mRNA isoform expression by the reverse transcription-polymerase chain reaction in the amygdala, hippocampus, and cerebellum following a unilateral injection of Abeta(25-35) or vehicle into the amygdala. The levels of tau proteins were increased bilaterally in the amygdala of Abeta(25-35)- compared to vehicle treated animals 8 and 16 days following treatment. The molecular weights of tau proteins were decreased in the Abeta(25-35)-treated (59-69 kDa) compared to the vehicle-treated (67-72 kDa) animals 8 days following treatment. There were no changes in tau mRNA expression in any brain region examined. In this model, just as in AD, there is an increase in tau protein levels without a change in tau mRNA expression, suggesting that Abeta peptides may influence tau protein stability in both the rat and the human brain. PMID- 10716897 TI - Electrophysiology of sipatrigine: a lamotrigine derivative exhibiting neuroprotective effects. AB - Sipatrigine (BW619C89), a derivative of the antiepileptic agent lamotrigine, has potent neuroprotective properties in animal models of cerebral ischemia and head injury. In the present study we investigated the electrophysiological effects of sipatrigine utilizing intracellular current-clamp recordings obtained from striatal spiny neurons in rat corticostriatal slices and whole-cell patch-clamp recordings in isolated striatal neurons. The number of action potentials produced in response to a depolarizing current pulse in the recorded neurons was reduced by sipatrigine (EC(50) 4.5 microM). Although this drug preferentially blocked action potentials in the last part of the depolarizing current pulse, it also decreased the frequency of the first action potentials. Sipatrigine also inhibited tetrodotoxin-sensitive sodium (Na(+)) current recorded from isolated striatal neurons. The EC(50) for this inhibitory action was 7 microM at the holding potential (V(h)) of -65 mV, but 16 microM at V(h) = -105, suggesting a dependence of this pharmacological effect on the membrane potential. Moreover, although the inhibitory action of sipatrigine on Na(+) currents was maximal during high-frequency activation (20 Hz), it could also be detected at low frequencies. The amplitude of excitatory postsynaptic potentials (EPSPs), recorded following stimulation of the corticostriatal pathway, was depressed by sipatrigine (EC(50) 2 microM). This inhibitory action, however, was incomplete; in fact maximal concentrations of this drug reduced EPSP amplitude by only 45%. Sipatrigine produced no increase in paired-pulse facilitation, suggesting that the modulation of a postsynaptic site was the main pharmacological effect of this agent. The inhibition of voltage-dependent Na(+) channels exerted by sipatrigine might account for its depressant effects on both repetitive firing discharge and corticostriatal excitatory transmission. The modulation of Na(+) channels described here, as well as the previously observed inhibition of high-voltage activated calcium currents, might contribute to the neuroprotective efficacy exerted by this compound in experimental models of in vitro and in vivo ischemia. PMID- 10716898 TI - Differential vulnerabilities of substantia nigra catecholamine neurons to excitatory amino acid-induced degeneration in rat midbrain slices. AB - Although differential vulnerability in different regions of the central nervous system is a characteristic feature of neurodegenerative disorders in vivo, its cellular basis is not well understood. In the present study we investigated whether catecholamine neurons in different regions of the substantia nigra (SN) are differentially vulnerable to excitatory amino acid-induced damage in a midbrain slice preparation. Rats were anesthetized by halothane inhalation and killed, the brain was rapidly removed, and 300-microm-thick midbrain slices were cut horizontally on a vibratome. The slices were incubated at 35 degrees C for 2 h in saline buffer containing either kainic acid (KA) or N-methyl-d-aspartate (NMDA) (10-50 microM). They were then fixed and cut into 30-microm sections that were coplanar with the horizontal slice. Individual catecholamine neurons were identified in these thin sections using an antibody to tyrosine hydroxylase coupled to diaminobenzidine. Catecholaminergic neurons in the dorsal and ventral tiers of the SN were readily identified by reference to an atlas of the distribution of catecholamine neurons in the horizontal plane. Using dendritic degeneration as a sensitive index of damage, and submaximal concentrations of KA and NMDA, we found that catecholamine neurons in the dorsal tier were more vulnerable than those in the ventral tier. For example, KA (10 microM) caused a significant reduction in the proportion of neurons with dendrites in the dorsal tier (from 60 to 34%) without altering the dendritic arbor of ventral tier neurons. After treatment with 50 microM KA, only 11% of dorsal tier neurons retained any dendrites while 45% of ventral tier neurons retained their dendrites. These differences were statistically significant (P<0.001). A similar differential vulnerability was apparent in slices treated with NMDA; neurons in the dorsal tier lost dendrites before detectable damage in the ventral tier. An understanding of the comparative anatomical, neurochemical, and physiological properties of vulnerable (dorsal tier) and resistant (ventral tier) catecholamine neurons in rat SN may provide significant insights into the mechanisms and treatment of neurodegenerative disorders involving catecholamine neurons. PMID- 10716899 TI - Mammalian-cell-produced neurturin (NTN) is more potent than purified Escherichia coli-produced NTN. AB - Neurturin (NTN) is a recently identified homologue of glial-cell-line-derived neurotrophic factor. Both factors promote the survival of dopaminergic (DA) neurons. We investigated the biological activity of mammalian-cell-produced NTN versus purified Escherichia coli-produced NTN. Baby hamster kidney cells were engineered to stably secrete mature human NTN. Mammalian-cell-derived NTN enhanced the activity of embryonic DA neurons in vitro, with greater potency (maximum effect achieved in the picogram range) than purified E. coli-produced NTN. Cell-based delivery of NTN (less than 10 ng/day) was also shown to be biologically active in vivo. These results suggest that mammalian-cell-derived NTN, synthesized de novo and delivered in small quantities to the parenchyma at the target site, may be as active as much larger quantities of purified, E. coli produced NTN, delivered by other means. PMID- 10716900 TI - A two-dimensional gel electrophoretic study of proteins synthesized and released by degenerating adult mouse sciatic nerve. AB - Previous two-dimensional (2-D) gel electrophoretic studies of proteins secreted by degenerating mammalian peripheral nerves (Ignatius et al., 1986, Proc. Natl. Acad. Sci. USA 83: 1125-1129; Muller et al., 1986, J. Cell Biol. 102: 393-402) detected the up-regulation of two proteins of 67-70 and 34-37 kDa, although they failed to resolve proteins smaller than about 15 kDa or with isoelectric points greater than 8. In the present study, we have used 2-D gels that can resolve proteins in the molecular mass range 3.6-200 kDa and isoelectric point range 2.4 10.6. This revealed that the incorporation of radiolabel by three diffusible proteins with apparent molecular mass/isoelectric point values of 38/5-6, 27-31/4 5, and 8/>10 was increased in the distal stumps of sciatic nerves 4 days after lesion, while the radiolabel incorporation by a further two proteins (15/5.3 and 12.5-17.5/6.8-7.5) was increased in the distal nerve stump 15 days after lesion. The possible cellular sources of these proteins were assessed by comparing protein secretion from unoperated nerves with nerve segments maintained in culture for 4 days (in which the contribution from recruited macrophages would be expected to be minimal) and segments of nerve that had been frozen and then replaced in situ for 4 days (in which the contribution from nerve sheath cells would be expected to be minimal). This revealed that three of the proteins up regulated in lesioned nerves (27-31/4-5, 15/5.3, and 12.5-17.5/6.8-7.5) are probably sheath cell products, while the other two (38/5-6 and 8/>10) may be secreted mainly by macrophages (or other cells) that infiltrate the frozen nerve segments. The identity of these proteins and their possible involvement in axonal regeneration remain to be determined. PMID- 10716901 TI - Effects of cortical spreading depression on cortical blood flow, impedance, DC potential, and infarct size in a rat venous infarct model. AB - A cortical venous infarction model has been evaluated as to the degree of regional flow reduction and by studying effects of cortical spreading depression (CSD). Two adjacent cortical veins were occluded photochemically with rose bengal and fiberoptic illumination. Seven rats served to demonstrate effects on regional cortical blood flow using laser Doppler scanning. In 36 rats local CBF, DC potential, and brain tissue impedance were measured continuously for 75 min after vein occlusion. No, 3, or 10 CSD waves were induced by potassium chloride injection during the initial 75 min. Rats were compared for spontaneous CSDs; baseline local CBF, CBF, and impedance response to CSD; and infarct volume. Seventy-five minutes after vein occlusion regional cortical flow in a 3.5x7-mm window was reduced to 34.3+/-13.2%. At 45% of the 840 measured locations in 7 rats flow was <40% baseline and at 27.3% <30%, indicating a widespread penumbra territory. During the initial 75 min 2.1+/-1.1 spontaneous CSDs were observed. There was a positive correlation between the number of spontaneous CSDs seen acutely and infarction volume after 5 days. Moreover, brain injury was significantly increased in the group with 10 KCl-induced CSDs. A reduced 1CBF response and an overshooting tissue impedance change during CSD were predictors of ischemic damage. This study demonstrates a CSD-related growth of the venous infarct. Second, the data indicate that flow after two-vein occlusion resembles that seen under penumbra conditions, allowing for studies of damage mechanisms responsible for infarct growth. PMID- 10716902 TI - Functional activity of zona incerta neurons is altered after nigrostriatal denervation in hemiparkinsonian rats. AB - The cellular expression of cytochrome oxidase subunit I (COI) mRNA as a metabolic marker for neuronal activity has recently been used to examine the effects of nigrostriatal denervation on the functioning of the basal ganglia. However, this technique also allows functional changes to be detected in other cerebral structures in parkinsonian syndromes. Since the zona incerta has been implicated in locomotor activity and has been the site of stereotactic surgery in Parkinson's disease, the aim of our study was to determine whether changes in neuronal activity are observed in this structure during parkinsonism. Using in situ hybridization, we analyzed the expression of COI mRNA in rats with 6 hydroxydopamine unilateral lesion of the substantia nigra and sham-operated animals. A quantitative analysis showed that COI mRNA expression was increased in the zona incerta ipsilateral to the lesion 24 h and 3 days after lesion, but by day 14 had returned almost to the level observed in controls. The hyperactivity of zona incerta neurons was confirmed by single-unit electrophysiological recordings. In contrast to the COI mRNA expression, the increase in electric neuronal activity was still observed 1 month after the lesion. This increase in zona incerta neuronal activity after nigrostriatal denervation might be related to the pathophysiology of parkinsonism, at least in the early stages, in agreement with previous reports suggesting an involvement of the zona incerta in motor functions. PMID- 10716903 TI - Laminar distribution of isocortical neurons projecting to occipital grafts in neonate and adult rats. AB - Physiologically responsive grafts of embryonic (E16) occipital neurons placed into the visual cortex of adult rats were shown previously (Gaillard et al., 1998, Restor. Neurol. Neurosci. 12: 13-25) to receive a predominant (93-97%) cortical input from the infragranular layers V-VI. The present paper examines whether this specific pattern of connections is related to some process of maturation of the host cortex. Pieces of embryonic (E16) occipital cortical tissue were grafted into the visual cortex of neonate (P0), 1-week-old (P7), and adult (P120) subjects. Four months later, graft responsiveness was assessed through field potential recordings and host-to-graft afferents were labeled with a retrograde tracer (cholera toxin subunit B). The data show first that afferents to physiologically active grafts originate about equally from both supra- and infragranular cortical layers in newborn subjects and second that supragranular neurons contribute only 20 and 1.5% of these inputs in P7 and P120 recipients, respectively. This strong upside-down laminar shift of afferents may correlate with the layout of subsets of host neurons that at a given developmental stage would have the intrinsic capacity to regrow an axon. Substantial axogenesis and synaptic stabilization of host-to-graft cortical afferents appear possible only within the critical period for the supragranular neurons but may occur throughout life for the infragranular neurons. PMID- 10716904 TI - Helper T-cell epitope immunodominance associated with structurally stable segments of hen egg lysozyme and HIV gp120. AB - Although many antigen sequences potentially can bind to the MHCII proteins, only a small number of epitopes dominate the immune response. Additional mechanisms of processing, presentation or recognition must restrict the immune response against a large portion of the antigen. A highly significant correlation is found between epitope immunodominance and local structural stability in hen egg lysozyme. Since antigen proteins are likely to retain substantial native-like structure in the processing compartment, protease action may be focused on regions that are most readily accommodated in the protease active sites, and thus, the intervening sequence are preferentially presented. Immunodominance also correlates with sequence conservation as expected from the constraints imposed by structure. These results suggest that the three-dimensional structure of the antigen limits the immune response against some antigen segments. For HIV gp120, a substantial improvement in the accuracy of epitope prediction is obtained by combining rules for MHCII binding with a restriction of the predicted epitopes to well-conserved sequences. PMID- 10716905 TI - The universal ancestor lived in a thermophilic or hyperthermophilic environment. AB - Galtier et al. (Science 1999, 283, 220-221) exploit the correlation between the optimal growth temperature in prokaryotes and the G+C content of rRNAs and establish that the last universal common ancestor (LUCA) lived in a mesophilic environment. This result was achieved by estimating the G+C content of the ancestral sequences of the rRNAs of the LUCA through use of a complex Markov model. I have re-analysed their alignments of the rDNAs with maximum parsimony and I have found that their result is not robust and is, in all likelihood, incorrect. In particular, the rRNA ancestral sequences reconstructed with maximum parsimony from these rDNA alignments as well as those reconstructed after eliminating all the sites that turn out to be ambiguous to the parsimony algorithm and to a site-by-site inspection of these alignments, are such as to suggest that the LUCA lived in a thermophilic or hyperthermophilic environment. This finding is also supported by some tRNA ancestral sequences. The main conclusion of this analysis is that if the LUCA was a progenote then the origin of life might have taken place at a high temperature. PMID- 10716906 TI - A statistical-mechanical model for regulation of long-range chromatin structure and gene expression. AB - In eukaryotic organisms, organization of chromatin is considered to play a role in transcriptional regulation by limiting the accessibility of a gene to the transcription machinery. It is not fully understood, however, how chromatin around a particular locus can be specifically altered to allow transcription. This paper introduces a statistical-mechanical model of chromatin to illustrate a potential mechanism. The model, which is mathematically equivalent to the one dimensional Ising model of magnetic systems, explains how gene regulatory DNA sequences can affect the chromatin structure over a long distance in cis. The main assumption of the model is cooperativity of histone H1 in binding to the nucleosome array. This cooperativity results in a long-range correlation of histone H1 distribution along the chromatin. Due to this long-range correlation, a gene regulatory element, such as a transcriptional enhancer, may lead to depletion of histone H1 over a large region of chromatin thereby increasing the accessibility of the gene. The model provides a mechanism for a sensitive genetic switch and explains several aspects of gene regulation and chromatin structure. PMID- 10716907 TI - Theory for the systemic definition of metabolic pathways and their use in interpreting metabolic function from a pathway-oriented perspective. AB - Cellular metabolism is most often described and interpreted in terms of the biochemical reactions that make up the metabolic network. Genomics is providing near complete information regarding the genes/gene products participating in cellular metabolism for a growing number of organisms. As the true functional units of metabolic systems are its pathways, the time has arrived to define metabolic pathways in the context of whole-cell metabolism for the analysis of the structural design and capabilities of the metabolic network. In this study, we present the theoretical foundations for the identification of the unique set of systemically independent biochemical pathways, termed extreme pathways, based on system stochiometry and limited thermodynamics. These pathways represent the edges of the steady-state flux cone derived from convex analysis, and they can be used to represent any flux distribution achievable by the metabolic network. An algorithm is presented to determine the set of extreme pathways for a system of any complexity and a classification scheme is introduced for the characterization of these pathways. The property of systemic independence is discussed along with its implications for issues related to metabolic regulation and the evolution of cellular metabolic networks. The underlying pathway structure that is determined from the set of extreme pathways now provides us with the ability to analyse, interpret, and perhaps predict metabolic function from a pathway-based perspective in addition to the traditional reaction-based perspective. The algorithm and classification scheme developed can be used to describe the pathway structure in annotated genomes to explore the capabilities of an organism. PMID- 10716908 TI - Assessment of the metabolic capabilities of Haemophilus influenzae Rd through a genome-scale pathway analysis. AB - The annotated full DNA sequence is becoming available for a growing number of organisms. This information along with additional biochemical and strain-specific data can be used to define metabolic genotypes and reconstruct cellular metabolic networks. The first free-living organism for which the entire genomic sequence was established was Haemophilus influenzae. Its metabolic network is reconstructed herein and contains 461 reactions operating on 367 intracellular and 84 extracellular metabolites. With the metabolic reaction network established, it becomes necessary to determine its underlying pathway structure as defined by the set of extreme pathways. The H. influenzae metabolic network was subdivided into six subsystems and the extreme pathways determined for each subsystem based on stoichiometric, thermodynamic, and systems-specific constraints. Positive linear combinations of these pathways can be taken to determine the extreme pathways for the complete system. Since these pathways span the capabilities of the full system, they could be used to address a number of important physiological questions. First, they were used to reconcile and curate the sequence annotation by identifying reactions whose function was not supported in any of the extreme pathways. Second, they were used to predict gene products that should be co-regulated and perhaps co-expressed. Third, they were used to determine the composition of the minimal substrate requirements needed to support the production of 51 required metabolic products such as amino acids, nucleotides, phospholipids, etc. Fourth, sets of critical gene deletions from core metabolism were determined in the presence of the minimal substrate conditions and in more complete conditions reflecting the environmental niche of H. influenzae in the human host. In the former case, 11 genes were determined to be critical while six remained critical under the latter conditions. This study represents an important milestone in theoretical biology, namely the establishment of the first extreme pathway structure of a whole genome. PMID- 10716909 TI - Modeling plasma virus concentration during primary HIV infection. AB - During primary HIV infection the viral load in plasma increases, reaches a peak, and then declines. Phillips has suggested that the decline is due to a limitation in the number of cells susceptible to HIV infection, while other authors have suggested that the decline in viremia is due to an immune response. Here we address this issue by developing models of primary HIV-1 infection, and by comparing predictions from these models with data from ten anti-retroviral, drug naive, infected patients. Applying nonlinear least-squares estimation, we find that relatively small variations in parameters are capable of mimicking the highly diverse patterns found in patient viral load data. This approach yields an estimate of 2.5 days for the average lifespan of productively infected cells during primary infection, a value that is consistent with results obtained by drug perturbation experiments. We find that the data from all ten patients are consistent with a target-cell-limited model from the time of initial infection until shortly after the peak in viremia. However, the kinetics of the subsequent fall and recovery in virus concentration in some patients are not consistent with the predictions of the target-cell-limited model. We illustrate that two possible immune response mechanisms, cytotoxic T lymphocyte destruction of infected target cells and cytokine suppression of viral replication, could account for declines in viral load data not predicted by the original target-cell-limited model. We conclude that some additional process, perhaps mediated by CD8+ T cells, is important in at least some patients. PMID- 10716910 TI - On the genealogy of a population of biparental individuals. AB - If one goes backward in time, the number of ancestors of an individual doubles at each generation. This exponential growth very quickly exceeds the population size, when this size is finite. As a consequence, the ancestors of a given individual cannot be all different and most remote ancestors are repeated many times in any genealogical tree. The statistical properties of these repetitions in genealogical trees of individuals for a panmictic closed population of constant size N can be calculated. We show that the distribution of the repetitions of ancestors reaches a stationary shape after a small number G(c) approximately log N of generations in the past, that only about 80% of the ancestral population belongs to the tree (due to coalescence of branches), and that two trees for individuals in the same population become identical after G(c)generations have elapsed. Our analysis is easy to extend to the case of exponentially growing population. PMID- 10716911 TI - Endogenous MCP-1 influences systemic cytokine balance in a murine model of acute septic peritonitis. AB - Sepsis and septic syndrome represent an intense systemic response with multiple physiologic and immunologic abnormalities, leading to multiple organ failure. Recent investigations suggest that the critical conditions are balanced by endogenous cytokines. In the present study, we examined the involvement of endogenous monocyte chemoattractant protein (MCP)-1 in the regulation of cytokine production in tissue/organs in a murine model of acute septic peritonitis induced by cecal ligation and puncture (CLP). Initial studies showed that CLP induced elevated levels of MCP-1 in tissues, such as liver, lung, and kidney. To neutralize endogenous MCP-1, either anti-MCP-1 antibodies or control antibodies were intraperitoneally administered 2 h prior to CLP. Administration of anti-MCP 1 antibodies resulted in a decrease in the level of interleukin (IL)-13 in tissues, while increasing the level of tumor necrosis factor-alpha, compared to control. In addition, anti-MCP-1 treatment decreased the level of IL-12 and, in contrast, increased the level of IL-10 in specific tissues. These findings suggest that endogenous MCP-1 influences the cytokine balance in tissues in favor of anti-inflammatory and immune-enhancing cytokines, probably protecting the host from tissue/organ damage during sepsis. PMID- 10716912 TI - Cell dynamics and expression of tumor necrosis factor (TNF)-alpha, interleukin-6, and TNF receptors in angioimmunoblastic lymphadenopathy-type T cell lymphoma. AB - Angioimmunoblastic lymphadenopathy (AILD)-type T cell lymphoma is histologically characterized by a mixed infiltrate of atypical T cells and B cells including B immunoblasts and plasma cells as well as eosinophils accompanied by proliferation of high endothelial venules. These morphological peculiarities are widely believed to reflect an abnormal pattern of cytokine expression. To clarify the cell dynamics and cytokine expression pattern in the lymph nodes of AILD-type T cell lymphoma, the frequency of proliferating/apoptotic cells and localization of tumor necrosis factor (TNF)-alpha and interleukin (IL)-6 producing cells were determined. Double staining was performed for (1) cell markers and Ki-67 antigen, (2) cell markers and the terminal deoxytransferase-mediated dUTP nick end labeling (TUNEL) method, or (3) cell markers and cytokines. The proliferating cell ratio in atypical T cells of AILD-type T cell lymphoma determined by Ki-67 labeling was 20.2+/-5.0%, while other peripheral T cell lymphomas (PTL) exhibited a ratio of 32.9+/-2.5%. TUNEL-positive apoptotic cells were 0.8+/-0.1% of total cells in AILD-type T cell lymphoma. They were dominantly atypical cells with positive T cell markers. In contrast, lymphoma cells in other types of PTL or paracortical cells in reactive follicular hyperplasia had only 0.3+/-0.1 or 0.4+/ 0.1% TUNEL-positive cells, respectively. Thus, lymphoma cells in AILD-type T cell lymphoma demonstrated suppressed proliferating activity and enhanced apoptosis when compared to other types of PTL. TNF-alpha-producing cells were observed in all of the lymph nodes from AILD-type T cell lymphoma cases (15/15) and positive staining was obtained in the majority of atypical T cells and scattered macrophages. In contrast, IL-6 was localized to clusters of atypical T cells in some of the cases (9/15). Further, the expression of TNF-alpha, IL-6, and TNF receptors I and II (TNFRI and TNFRII) was examined by RT-PCR. The TNF-alpha message (2/2) and IL-6 message (2/2) was present in the lymph nodes of AILD-type T cell lymphoma by examination using RT-PCR, while both messages were negative in control cases (0/7). As far as an expression of mRNA for TNF receptors in AILD type T cell lymphoma cases, mRNA for TNFRI was definitely expressed in both of the cases (2/2) while TNFRII mRNA was weakly expressed in one case (1/2). Overexpression of TNF-alpha as well as TNFRI may play a role in controlling T cell proliferation through an autocrine (T cell-T cell interaction) and paracrine (macrophage-T cell interaction) fashion. IL-6, which was also expressed by part of lymphoma cells of AILD-type T cell lymphoma, facilitates the proliferation of B cells, plasma cells, and T cells or endothelial cells in the lymph nodes of AILD-type T cell lymphoma. PMID- 10716913 TI - Distribution of disease around the aortocoeliac branch of white carneau pigeons at different ages. AB - The distribution of atherosclerotic lesions in the human aorta changes with age. Lipid deposition tends to occur downstream of branch sites in immature vessels but upstream of them at later ages. Comparable age-related distributions of spontaneous and induced lesions have been demonstrated in rabbit aortas. Spontaneous disease is known to develop upstream of the aortocoeliac branch in mature White Carneau pigeons. Using a frequency mapping technique, we investigated whether the same pattern or a downstream one occurs in immature pigeons. Lesions in hatchlings occurred upstream of the branch, to the left of the midline. By 5 months, these lesions had expanded, and a second upstream area, to the right of the midline, was also affected. At later ages, disease frequencies increased in both of these regions but not elsewhere. Thus, contrary to findings in rabbit and human aortas, there was no evidence for a switch from a downstream to an upstream distribution with age. The two rabbit distributions have been attributed to the similar age-related patterns seen in the permeability of the arterial wall; the mature but not the immature pattern of permeability is NO-dependent. The absence of the juvenile disease pattern in pigeons suggests that they might show the NO-dependent pattern of permeability at all ages. PMID- 10716914 TI - Swelling of free-radical-induced megamitochondria causes apoptosis. AB - Recently, we have found that cultured cells from various sources exposed to free radicals become apoptotic in the presence of megamitochondria (MG). The purpose of the present study is to answer the following two questions: (1) Do functions obtained from the "MG fraction" isolated from normal mitochondria by a routine procedure represent the functions of MG since the fraction consists of enlarged and normal-size mitochondria? (2) What is the correlation between MG formation and apoptotic changes of the cell? In the present study the heavy fraction rich in mitochondria enlarged to varying degrees and the light fraction consisting mainly of normal-size mitochondria were isolated independently from the livers of rats treated with hydrazine for 4 days (4H animals) and 8 days (8H animals), and some functions related to apoptosis were compared. Results were as follows: (1) Mitochondria in both fractions obtained from 8H animals swelled far less in various media than those obtained from the controls, suggesting that the permeability transition pores had been opened before they were exposed to swelling media. (2) The membrane potential of mitochondria in both fractions obtained from 8H animals was distinctly decreased. (3) The rates of reactive oxygen species generation from mitochondria of both fractions in 4H animals were equally elevated, while those in 8H animals were equally decreased compared to those of controls. These results, together with morphological data obtained in the present study, suggest that enlarged and normal-size mitochondria are a part of MG and that the secondary swelling of MG causes the apoptotic changes in the cell. PMID- 10716915 TI - Effect of growth factors on proliferation of normal, borderline, and malignant breast epithelial cells. AB - Growth factors play an important role in the development and growth regulation of normal breast. They are also known to be autocrine or paracrine stimulators of breast cancer cells. However, their role on cells of proliferative breast disease has not been studied so far. This study was undertaken to quantitate the proliferative effect of various growth factors on "normal," borderline, and malignant breast epithelial cells. For this purpose, epithelial cell lines of histologically normal human breast and histologically proven proliferative breast diseases were established. Cell lines MCF-7 and T47D were used as malignant cells. The growth factors under study include epidermal growth factor, fibroblast growth factors acidic and basic, platelet-derived growth factor, and insulin-like growth factors 1 (IGF-1) and 2 (IGF-2). Their proliferative effect was determined by incubating the cells with growth factors for 24 h after achieving basal conditions in serum-free medium for 72 h, followed by quantitating the S-phase fraction by flow cytometry. All of the growth factors were found to be capable of inducing cellular proliferation on the entire range of mammary epithelia. Epidermal growth factor was consistently found to be a potent mitogen. Fibroblast growth factor acidic had a higher effect compared to fibroblast growth factor basic in inducing the cells to move from G(0)/G(1) to S-phase. Platelet-derived growth factor had a moderate proliferative response. In the family of insulin like growth factors, IGF-1 was the dominant mitogen for normal cells and IGF-2 was the dominant proliferative stimulus for the cell line MCF-7. In the cell lines of proliferative breast disease and T47D, both were at par as mitogenic agents. These results suggest that the cells of proliferative breast disease develop some of the biological characteristics of malignancy. PMID- 10716916 TI - EMBO Gold Medal 1999. Waiting periods, instructive signals and positional information. PMID- 10716917 TI - Acetylation: a regulatory modification to rival phosphorylation? AB - The fact that histones are modified by acetylation has been known for almost 30 years. The recent identification of enzymes that regulate histone acetylation has revealed a broader use of this modification than was suspected previously. Acetylases are now known to modify a variety of proteins, including transcription factors, nuclear import factors and alpha-tubulin. Acetylation regulates many diverse functions, including DNA recognition, protein-protein interaction and protein stability. There is even a conserved structure, the bromodomain, that recognizes acetylated residues and may serve as a signalling domain. If you think all this sounds familiar, it should be. These are features characteristic of kinases. So, is acetylation a modification analogous to phosphorylation? This review sets out what we know about the broader substrate specificity and regulation of acetyl- ases and goes on to compare acetylation with the process of phosphorylation. PMID- 10716918 TI - Consequences of manganese replacement of copper for prion protein function and proteinase resistance. AB - The prion protein (PrP) binds copper and has antioxidant activity enhancing the survival of neurones in culture. The ability of the PrP to bind other cations was tested and it was found that only manganese could substitute for copper. Although initially manganese-loaded PrP exhibited similar structure and activity to copper loaded PrP, after aging, manganese-loaded PrP became proteinase resistant and lost function. It was also found that manganese could be incorporated into PrP expressed by astrocytes and that this PrP was partially proteinase resistant. These results show that it is possible to generate proteinase-resistant PrP from cells and suggest a possible mechanism for the formation of the scrapie isoform of the PrP as generated in sporadic prion disease. PMID- 10716919 TI - Secreted cathepsin L generates endostatin from collagen XVIII. AB - Endostatin, an inhibitor of angiogenesis and tumor growth, was identified originally in conditioned media of murine hemangioendothelioma (EOMA) cells. N terminal amino acid sequencing demonstrated that it corresponds to a fragment of basement membrane collagen XVIII. Here we report that cathepsin L is secreted by EOMA cells and is responsible for the generation of endostatin with the predicted N-terminus, while metalloproteases produce larger fragments in a parallel processing pathway. Efficient endostatin generation requires a moderately acidic pH similar to the pericellular milieu of tumors. The secretion of cathepsin L by a tumor cell line of endothelial origin suggests that this cathepsin may play a role in angiogenesis. We propose that cleavage within collagen XVIII's protease sensitive region evolved to regulate excessive proteolysis in conditions of induced angiogenesis. PMID- 10716920 TI - Inhibitor binding induces active site stabilization of the HCV NS3 protein serine protease domain. AB - Few structures of viral serine proteases, those encoded by the Sindbis and Semliki Forest viruses, hepatitis C virus (HCV) and cytomegalovirus, have been reported. In the life cycle of HCV a crucial role is played by a chymotrypsin like serine protease encoded at the N-terminus of the viral NS3 protein, the solution structure of which we present here complexed with a covalently bound reversible inhibitor. Unexpectedly, the residue in the P2 position of the inhibitor induces an effective stabilization of the catalytic His-Asp hydrogen bond, by shielding that region of the protease from the solvent. This interaction appears crucial in the activation of the enzyme catalytic machinery and represents an unprecedented observation for this family of enzymes. Our data suggest that natural substrates of this serine protease could contribute to the enzyme activation by a similar induced-fit mechanism. The high degree of similarity at the His-Asp catalytic site region between HCV NS3 and other viral serine proteases suggests that this behaviour could be a more general feature for this category of viral enzymes. PMID- 10716921 TI - Distinct cellular receptor interactions in poliovirus and rhinoviruses. AB - Receptor binding to human poliovirus type 1 (PV1/M) and the major group of human rhinoviruses (HRV) was studied comparatively to uncover the evolution of receptor recognition in picornaviruses. Surface plas- mon resonance showed receptor binding to PV1/M with faster association and dissociation rates than to HRV3 and HRV16, two serotypes that have similar binding kinetics. The faster rate for receptor association to PV1/M suggested a relatively more accessible binding site. Thermodynamics for receptor binding to the viruses and assays for receptor mediated virus uncoating showed a more disruptive receptor interaction with PV1/M than with HRV3 or HRV16. Cryo-electron microscopy and image reconstruction of receptor-PV1/M complexes revealed receptor binding to the 'wall' of surface protrusions surrounding the 'canyon', a depressive surface in the capsid where the rhinovirus receptor binds. These data reveal more exposed receptor-binding sites in poliovirus than rhinoviruses, which are less protected from immune surveillance but more suited for receptor-mediated virus uncoating and entry at the cell surface. PMID- 10716922 TI - Receptor recognition by a hepatitis B virus reveals a novel mode of high affinity virus-receptor interaction. AB - The duck hepatitis B virus model system was used to elucidate the characteristics of receptor (carboxypeptidase D, gp180) interaction with polypeptides representing the receptor binding site in the preS part of the large viral surface protein. We demonstrate the pivotal role of carboxypeptidase D for virus entry and show its C-domain represents the virus attachment site, which binds preS with extraordinary affinity. Combining results from surface plasmon resonance spectroscopy and two-dimensional NMR analysis we resolved the contribution of preS sequence elements to complex stability and show that receptor binding potentially occurs in two steps. Initially, a short alpha-helix in the C-terminus of the receptor binding domain facilitates formation of a primary complex. This complex is stabilized sequentially, involving approximately 60 most randomly structured amino acids preceding the helix. Thus, hepadnaviruses exhibit a novel mechanism of high affinity receptor interaction by conserving the potential to adapt structure during binding rather than to preserve it per se. We propose that this process represents an alternative strategy to escape immune surveillance and the evolutionary pressure inherent in the compact hepadnaviral genome organization. PMID- 10716923 TI - Structural basis of hyaluronan degradation by Streptococcus pneumoniae hyaluronate lyase. AB - Streptococcus pneumoniae hyaluronate lyase (spnHL) is a pathogenic bacterial spreading factor and cleaves hyaluronan, an important constituent of the extra- cellular matrix of connective tissues, through an enzymatic beta-elimination process, different from the hyaluronan degradation by hydrolases in animals. The mechanism of hyaluronan binding and degradation was proposed based on the 1.56 A resolution crystal structure, substrate modeling and mutagenesis studies on spnHL. Five mutants, R243V, N349A, H399A, Y408F and N580G, were constructed and their activities confirmed our mechanism hypothesis. The important roles of Tyr408, Asn349 and His399 in enzyme catalysis were proposed, explained and confirmed by mutant studies. The remaining weak enzymatic activity of the H399A mutant, the role of the free carboxylate group on the glucuronate residue, the enzymatic behavior on chondroitin and chondroitin sulfate, and the small activity increase in the N580G mutant were explained based on this mechanism. A possible function of the C-terminal beta-sheet domain is to modulate enzyme activity through binding to calcium ions. PMID- 10716925 TI - Hsp90 is a core centrosomal component and is required at different stages of the centrosome cycle in Drosophila and vertebrates. AB - To determine the molecular composition of the centrosome of a higher eukaryote, we carried out a systematic nano-electrospray tandem or MALDI mass spectrometry analysis of the polypeptides present in highly enriched preparations of immunoisolated Drosophila centrosomes. One of the proteins identified is Hsp83, a member of the highly conserved Hsp90 family including chaperones known to maintain the activity of many proteins but suspected to have other essential, unidentified functions. We have found that a fraction of the total Hsp90 pool is localized at the centrosome throughout the cell cycle at different stages of development in Drosophila and vertebrates. This association between Hsp90 and the centrosome can be observed in purified centrosomes and after treatment with microtubule depolymerizing drugs, two criteria normally used to define core centrosomal components. Disruption of Hsp90 function by mutations in the Drosophila gene or treatment of mammalian cells with the Hsp90 inhibitor geldanamycin, results in abnormal centrosome separation and maturation, aberrant spindles and impaired chromosome segregation. This suggests that another role of Hsp90 might be to ensure proper centrosome function. PMID- 10716924 TI - Functional HLA-DM on the surface of B cells and immature dendritic cells. AB - HLA-DM (DM) plays a critical role in antigen presentation through major histocompatibility complex (MHC) class II molecules. DM functions as a molecular chaperone by keeping class II molecules competent for antigenic peptide loading and serves as an editor by favoring presentation of high-stability peptides. Until now, DM has been thought to exert these activities only in late endosomal/lysosomal compartments of antigen-presenting cells. Here we show that a subset of DM resides at the cell surface of B cells and immature dendritic cells. Surface DM engages in complexes with putatively empty class II molecules and controls presentation of those antigens that rely on loading on the cell surface or in early endosomal recycling compartments. For example, epitopes derived from myelin basic protein that are implicated in the autoimmune disease multiple sclerosis are down-modulated by DM, but are presented in the absence of DM. Thus, this novel concept of functional DM on the surface may be relevant to both protective immune responses and autoimmunity. PMID- 10716926 TI - The endocytic protein intersectin is a major binding partner for the Ras exchange factor mSos1 in rat brain. AB - We recently identified intersectin, a protein containing two EH and five SH3 domains, as a component of the endocytic machinery. The N-terminal SH3 domain (SH3A), unlike other SH3 domains from intersectin or various endocytic proteins, specifically inhibits intermediate events leading to the formation of clathrin coated pits. We have now identified a brain-enriched, 170 kDa protein (p170) that interacts specifically with SH3A. Screening of combinatorial peptides reveals the optimal ligand for SH3A as Pp(V/I)PPR, and the 170 kDa mammalian son-of-sevenless (mSos1) protein, a guanine-nucleotide exchange factor for Ras, con- tains two copies of the matching sequence, PPVPPR. Immunodepletion studies confirm that p170 is mSos1. Intersectin and mSos1 are co-enriched in nerve terminals and are co-immunoprecipitated from brain extracts. SH3A competes with the SH3 domains of Grb2 in binding to mSos1, and the intersectin-mSos1 complex can be separated from Grb2 by sucrose gradient centrifugation. Overexpression of the SH3 domains of intersectin blocks epidermal growth factor-mediated Ras activation. These results suggest that intersectin functions in cell signaling in addition to its role in endocytosis and may link these cellular processes. PMID- 10716927 TI - Putative fusogenic activity of NSF is restricted to a lipid mixture whose coalescence is also triggered by other factors. AB - It has recently been reported that N-ethylmaleimide-sensitive fusion ATPase (NSF) can fuse protein-free liposomes containing substantial amounts of 1,2 dioleoylphosphatidylserine (DOPS) and 1, 2-dioleoyl-phosphatidyl-ethanolamine (DOPE) (Otter-Nilsson et al., 1999). The authors impart physiological significance to this observation and propose to re-conceptualize the general role of NSF in fusion processes. We can confirm that isolated NSF can fuse liposomes of the specified composition. However, this activity of NSF is resistant to inactivation by N-ethylmaleimide and does not depend on the presence of alpha SNAP (soluble NSF-attachment protein). Moreover, under the same conditions, either alpha-SNAP, other proteins apparently unrelated to vesicular transport (glyceraldehyde-3-phosphate dehydrogenase or lactic dehydrogenase) or even 3 mM magnesium ions can also cause lipid mixing. In contrast, neither NSF nor the other proteins nor magnesium had any significant fusogenic activity with liposomes composed of a biologically occurring mixture of lipids. A straightforward explanation is that the lipid composition chosen as optimal for NSF favors non-specific fusion because it is physically unstable when formed into liposomes. A variety of minor perturbations could then trigger coalescence. PMID- 10716928 TI - Exocytotic mechanism studied by truncated and zero layer mutants of the C terminus of SNAP-25. AB - The highly conserved SNARE proteins, SNAP-25, syntaxin and synaptobrevin, form a tight ternary complex, which is essential for exocytosis. Crystallization of this complex revealed a four-helix bundle with an unusual hydrophilic layer (zero layer) in its center. In order to evaluate the role of this layer in different kinetic components of secretion, we used the Semliki Forest virus (SFV) system to infect adrenal chromaffin cells with SNAP-25 Q174L, a point mutant in the zero layer. Using combined flash photolysis of caged calcium and membrane capacitance measurements, we investigated its effect on the exocytotic burst and sustained phase of exocytosis with high time resolution. Cells expressing SNAP-25 Q174L displayed a selective reduction in the sustained phase, while the two components of the exocytotic burst remained unaffected. Furthermore, the exocytotic response to the second flash was significantly reduced, indicating a decrease in refilling kinetics. We therefore conclude that the zero layer is critical for the formation of SNARE complexes, but that it plays no role in the dynamic equilibrium between the two exocytosis-competent vesicle pools. PMID- 10716929 TI - BDNF regulates eating behavior and locomotor activity in mice. AB - Brain-derived neurotrophic factor (BDNF) was studied initially for its role in sensory neuron development. Ablation of this gene in mice leads to death shortly after birth, and abnormalities have been found in both the peripheral and central nervous systems. BDNF and its tyrosine kinase receptor, TrkB, are expressed in hypothalamic nuclei associated with satiety and locomotor activity. In heterozygous mice, BDNF gene expression is reduced and we find that all heterozygous mice exhibit abnormalities in eating behavior or locomotor activity. We also observe this phenotype in independently derived inbred and hybrid BDNF mutant strains. Infusion with BDNF or NT4/5 can transiently reverse the eating behavior and obesity. Thus, we identify a novel non-neurotrophic function for neurotrophins and indicate a role in behavior that is remarkably sensitive to alterations in BDNF activity. PMID- 10716930 TI - Molecular determinants that mediate selective activation of p38 MAP kinase isoforms. AB - The p38 mitogen-activated protein kinase (MAPK) group is represented by four isoforms in mammals (p38alpha, p38beta2, p38gamma and p38delta). These p38 MAPK isoforms appear to mediate distinct functions in vivo due, in part, to differences in substrate phosphorylation by individual p38 MAPKs and also to selective activation by MAPK kinases (MAPKKs). Here we report the identification of two factors that contribute to the specificity of p38 MAPK activation. One mechanism of specificity is the selective formation of functional complexes between MAPKK and different p38 MAPKs. The formation of these complexes requires the presence of a MAPK docking site in the N-terminus of the MAPKK. The second mechanism that confers signaling specificity is the selective recognition of the activation loop (T-loop) of p38 MAPK isoforms. Together, these processes provide a mechanism that enables the selective activation of p38 MAPK in response to activated MAPKK. PMID- 10716931 TI - Point mutation in kit receptor tyrosine kinase reveals essential roles for kit signaling in spermatogenesis and oogenesis without affecting other kit responses. AB - The Kit receptor tyrosine kinase functions in hemato- poiesis, melanogenesis and gametogenesis. Kit receptor-mediated cellular responses include proliferation, survival, adhesion, secretion and differentiation. In mast cells, Kit-mediated recruitment and activation of phosphatidylinositol 3'-kinase (PI 3-kinase) produces phosphatidylinositol 3'-phosphates, plays a critical role in mediating cell adhesion and secretion and has contributory roles in mediating cell survival and proliferation. To investigate the consequences in vivo of blocking Kit mediated PI 3-kinase activation we have mutated the binding site for the p85 subunit of PI 3-kinase in the Kit gene, using a knock-in strategy. Mutant mice have no pigment deficiency or impairment of steady-state hematopoiesis. However, gametogenesis is affected in several ways and tissue mast cell numbers are affected differentially. While primordial germ cells during embryonic development are not affected, Kit(Y719F)/Kit(Y719F) males are sterile due to a block at the premeiotic stages in spermatogenesis. Furthermore, adult males develop Leydig cell hyperplasia. The Leydig cell hyperplasia implies a role for Kit in Leydig cell differentiation and/or steroidogenesis. In mutant females follicle development is impaired at the cuboidal stages resulting in reduced fertility. Also, adult mutant females develop ovarian cysts and ovarian tubular hyperplasia. Therefore, a block in Kit receptor-mediated PI 3-kinase signaling may be compensated for in hematopoiesis, melanogenesis and primordial germ cell development, but is critical in spermatogenesis and oogenesis. PMID- 10716932 TI - The pathway for perception and transduction of low-temperature signals in Synechocystis. AB - Low temperature is an important environmental factor that has effects on all living organisms. Various low-temperature-inducible genes encode products that are essential for acclimation to low temperature, but low-temperature sensors and signal transducers have not been identified. However, systematic disruption of putative genes for histidine kinases and random mutagenesis of almost all the genes in the genome of the cyanobacterium Synechocystis sp. PCC 6803 have allowed us to identify two histidine kinases and a response regulator as components of the pathway for perception and transduction of low-temperature signals. Inactivation, by targeted mutagenesis, of the gene for each of the two histidine kinases and inactivation of the gene for the response regulator depressed the transcription of several lowtemperature-inducible genes. PMID- 10716933 TI - Perinatal synthetic lethality and hematopoietic defects in compound mafG::mafK mutant mice. AB - Prior studies exploring the mechanisms controlling erythroid gene regulation implicated MARE (Maf recognition element) cis-elements as crucial to the transcriptional activity of many erythroid genes. Numerous transcription factors can elicit responses through MAREs, including not only the AP-1 family proteins, but also a growing list of factors composed of Cap-N-Collar (CNC)-small Maf heterodimers. While these factors can activate transcription from MAREs in co transfection assays, mouse germline mutations in cnc genes tested to date have failed to reveal primary erythroid phenotypes. Here we report that after combining the mafK and mafG targeted null alleles, mutant animals display several synthetic phenotypes, including erythroid deficiencies. First, compound homozygous small maf gene mutants survive embryogenesis, but die postnatally. Secondly, compound mutant animals develop severe neurological disorders. Thirdly, they exhibit an exacerbated mafG deficiency in megakaryopoiesis, specifically in proplatelet formation, resulting in profound thrombocytopenia. Finally, the compound mutant animals develop severe anemia accompanied by abnormal erythrocyte morphology and membrane protein composition. These data provide direct evidence that the small Maf transcription factors play an important regulatory role in erythropoiesis. PMID- 10716934 TI - Structure of the central core domain of TFIIEbeta with a novel double-stranded DNA-binding surface. AB - Human general transcription factor TFIIE consists of two subunits, TFIIEalpha and TFIIEbeta. Recently, TFIIEbeta has been found to bind to the region where the promoter starts to open to be single-stranded upon transcription initiation by RNA polymerase II. Here, the central core domain of human TFIIEbeta (TFIIEbetac) has been identified by a limited proteolysis. This solution structure has been determined by NMR. It consists of three helices with a beta hairpin at the C terminus, resembling the winged helix proteins. However, TFIIEbetac shows a novel double-stranded DNA-binding activity where the DNA-binding surface locates on the opposite side to the previously reported winged helix motif by forming a positively charged furrow. A model will be proposed that TFIIE stabilizes the preinitiation complex by binding not only to the general transcription factors together with RNA polymerase II but also to the promoter DNA, where double stranded DNA starts to open to be single-stranded upon activation of the preinitiation complex. PMID- 10716935 TI - Three conserved members of the RNase D family have unique and overlapping functions in the processing of 5S, 5.8S, U4, U5, RNase MRP and RNase P RNAs in yeast. AB - The biogenesis of a number of RNA species in eukaryotic cells requires 3' processing. To determine the enzymes responsible for these trimming events, we created yeast strains lacking specific 3' to 5' exonucleases. In this work, we describe the analysis of three members of the RNase D family of exonucleases (Rex1p, Rex2p and Rex3p). This work led to three important conclusions. First, each of these exonucleases is required for the processing of distinct RNAs. Specifically, Rex1p, Rex2p and Rex3p are required for 5S rRNA, U4 snRNA and MRP RNA trimming, respectively. Secondly, some 3' exonucleases are redundant with other exonucleases. Specifically, Rex1p and Rex2p function redundantly in 5.8S rRNA maturation, Rex1p, Rex2p and Rex3p are redundant for the processing of U5 snRNA and RNase P RNA, and Rex1p and the exonuclease Rrp6p have an unknown redundant essential function. Thirdly, the demonstration that the Rex proteins can affect reactions that have been attributed previously to the exosome complex indicates that an apparently simple processing step can be surprisingly complex with multiple exonucleases working sequentially in the same pathway. PMID- 10716936 TI - Distinct roles of two conserved Staufen domains in oskar mRNA localization and translation. AB - Drosophila Staufen protein is required for the localization of oskar mRNA to the posterior of the oocyte, the anterior anchoring of bicoid mRNA and the basal localization of prospero mRNA in dividing neuroblasts. The only regions of Staufen that have been conserved throughout animal evolution are five double stranded (ds)RNA-binding domains (dsRBDs) and a short region within an insertion that splits dsRBD2 into two halves. dsRBDs 1, 3 and 4 bind dsRNA in vitro, but dsRBDs 2 and 5 do not, although dsRBD2 does bind dsRNA when the insertion is removed. Full-length Staufen protein lacking this insertion is able to associate with oskar mRNA and activate its translation, but fails to localize the RNA to the posterior. In contrast, Staufen lacking dsRBD5 localizes oskar mRNA normally, but does not activate its translation. Thus, dsRBD2 is required for the microtubule-dependent localization of osk mRNA, and dsRBD5 for the derepression of oskar mRNA translation, once localized. Since dsRBD5 has been shown to direct the actin-dependent localization of prospero mRNA, distinct domains of Staufen mediate microtubule- and actin-based mRNA transport. PMID- 10716938 TI - Fission yeast switches mating type by a replication-recombination coupled process. AB - Fission yeast exhibits a homothallic life cycle, in which the mating type of the cell mitotically alternates in a highly regulated fashion. Pedigree analysis of dividing cells has shown that only one of the two sister cells switches mating type. It was shown recently that a site- and strand-specific DNA modification at the mat1 locus precedes mating-type switching. By tracking the fate of mat1 DNA throughout the cell cycle with a PCR assay, we identified a novel DNA intermediate of mating-type switching in S-phase. The time and rate of appearance and disappearance of this DNA intermediate are consistent with a model in which mating-type switching occurs through a replication-recombination coupled pathway. Such a process provides experimental evidence in support of a copy choice recombination model in Schizosaccharomyces pombe mating-type switching and is reminiscent of the sister chromatid recombination used to complete replication in the presence of certain types of DNA damage. PMID- 10716937 TI - Cyclin F regulates the nuclear localization of cyclin B1 through a cyclin-cyclin interaction. AB - The key regulator of G(2)-M transition of the cell cycle is M-phase promoting factor (MPF), a complex composed of cdc2 and a B-type cyclin. Cyclin B1 nuclear localization involves phosphorylation within a region called the cytoplasmic retention signal, which also contains a nuclear export signal. The mechanism of MPF nuclear localization remains unclear since it contains no functional nuclear localization signal (NLS). We exploited the yeast two-hybrid screen to find protein(s) potentially mediating localization of cyclin B1 and identified a novel interaction between cyclin B1 and cyclin F. We found that cdc2, cyclin B1 and cyclin F form a complex that exhibits histone H1 kinase activity. Cyclin B1 and cyclin F also colocalize through immunofluorescence studies. Additionally, deletion analysis revealed that each putative NLS of cyclin F is functional. Taken together, the data suggest that the NLS regions of cyclin F regulate cyclin B1 localization to the nucleus. The interaction between cyclin B1 and cyclin F represents the first example of direct cyclin-cyclin binding, and elucidates a novel mechanism that regulates MPF localization and function. PMID- 10716939 TI - Neonatal lethality with abnormal neurogenesis in mice deficient in DNA polymerase beta. AB - DNA polymerase beta (Polbeta) has been implicated in base excision repair in mammalian cells. However, the physiological significance of this enzyme in the body remains unclear. Here, we demonstrate that mice carrying a targeted disruption of the Polbeta gene showed growth retardation and died of a respiratory failure immediately after the birth. Histological examination of the embryos revealed defective neurogenesis characterized by apoptotic cell death in the developing central and peripheral nervous systems. Extensive cell death occurred in newly generated post-mitotic neuronal cells and was closely associated with the period between onset and cessation of neurogenesis. These findings indicate that Polbeta plays an essential role in neural development. PMID- 10716940 TI - Structure, function, and biology of SHIP proteins. PMID- 10716941 TI - The MAR-binding protein SATB1 orchestrates temporal and spatial expression of multiple genes during T-cell development. AB - SATB1 is expressed primarily in thymocytes and can act as a transcriptional repressor. SATB1 binds in vivo to the matrix attachment regions (MARs) of DNA, which are implicated in the loop domain organization of chromatin. The role of MAR-binding proteins in specific cell lineages is unknown. We generated SATB1 null mice to determine how SATB1 functions in the T-cell lineage. SATB1-null mice are small in size, have disproportionately small thymi and spleens, and die at 3 weeks of age. At the cellular level, multiple defects in T-cell development were observed. Immature CD3(-)CD4(-)CD8(-) triple negative (TN) thymocytes were greatly reduced in number, and thymocyte development was blocked mainly at the DP stage. The few peripheral CD4(+) single positive (SP) cells underwent apoptosis and failed to proliferate in response to activating stimuli. At the molecular level, among 589 genes examined, at least 2% of genes including a proto-oncogene, cytokine receptor genes, and apoptosis-related genes were derepressed at inappropriate stages of T-cell development in SATB1-null mice. For example, IL 2Ralpha and IL-7Ralpha genes were ectopically transcribed in CD4(+)CD8(+) double positive (DP) thymocytes. SATB1 appears to orchestrate the temporal and spatial expression of genes during T-cell development, thereby ensuring the proper development of this lineage. Our data provide the first evidence that MAR-binding proteins can act as global regulators of cell function in specific cell lineages. PMID- 10716942 TI - NUC-1, a caenorhabditis elegans DNase II homolog, functions in an intermediate step of DNA degradation during apoptosis. AB - One hallmark of apoptosis is the degradation of chromosomal DNA. We cloned the Caenorhabditis elegans gene nuc-1, which is involved in the degradation of the DNA of apoptotic cells, and found that nuc-1 encodes a homolog of mammalian DNase II. We used the TUNEL technique to assay DNA degradation in nuc-1 and other mutants defective in programmed cell death and discovered that TUNEL labels apoptotic cells only during a transient intermediate stage. Mutations in nuc-1 allowed the generation of TUNEL-reactive DNA but blocked the conversion of TUNEL reactive DNA to a subsequent TUNEL-unreactive state. Completion of DNA degradation did not occur in the absence of cell-corpse engulfment. Our data suggest that the process of degradation of the DNA of a cell corpse occurs in at least three distinct steps and requires activities provided by both the dying and the engulfing cell. PMID- 10716943 TI - An auxiliary mode of apoptotic DNA fragmentation provided by phagocytes. AB - CAD (caspase-activated DNase) can cause DNA fragmentation in apoptotic cells. Transgenic mice that ubiquitously express a caspase-resistant form of the CAD inhibitor (ICAD) were generated. Thymocytes prepared from the mice were resistant to DNA fragmentation induced by a variety of stimuli. However, similar numbers of TUNEL-positive cells were present in adult tissues of transgenic and wild-type mice. Exposure to gamma-irradiation caused a striking increase in the number of TUNEL-positive cells in the thymus of wild-type, but not transgenic, mice. TUNEL positive nuclei in transgenic mice were confined to thymic macrophages. When apoptotic thymocytes from the transgenic mice were cocultured with macrophages, the thymocytes underwent phagocytosis and their chromosomal DNA underwent fragmentation. This DNA fragmentation was sensitive to inhibitors that block the acidification of lysosomes. Hence, we conclude that the DNA fragmentation that occurs during apoptosis not only can result cell-autonomously from CAD activity but can also be attributed to a lysosomal acid DNase(s), most likely DNase II, after the apoptotic cells are engulfed. PMID- 10716944 TI - Rules for DNA target-site recognition by a lactococcal group II intron enable retargeting of the intron to specific DNA sequences. AB - Group II intron homing occurs primarily by a mechanism in which the intron RNA reverse splices into a DNA target site and is then reverse transcribed by the intron-encoded protein. The DNA target site is recognized by an RNP complex containing the intron-encoded protein and the excised intron RNA. Here, we analyzed DNA target-site requirements for the Lactococcus lactis Ll.LtrB group II intron in vitro and in vivo. Our results suggest a model similar to yeast mtDNA introns, in which the intron-encoded protein first recognizes a small number of nucleotide residues in double-stranded DNA and causes DNA unwinding, enabling the intron RNA to base-pair with the DNA for reverse splicing. Antisense-strand cleavage requires additional interactions between the protein and 3' exon. Key nucleotide residues are recognized directly by the intron-encoded protein independent of sequence context, and there is a stringent requirement for fixed spacing between target site elements recognized by the protein and RNA components of the endonuclease. Experiments with DNA substrates containing GC-clamps or "bubbles" indicate a requirement for DNA unwinding in the 3' exon but not the distal 5' exon region. Finally, by applying the target-site recognition rules, we show that the L1.LtrB intron can be modified to insert at new sites in a plasmid borne thyA gene in Escherichia coli. This strategy should be generally applicable to retargeting group II introns and to delivering foreign sequences to specific sites in heterologous genomes. PMID- 10716945 TI - Induction of terminal differentiation by constitutive activation of p38 MAP kinase in human rhabdomyosarcoma cells. AB - MyoD inhibits cell proliferation and promotes muscle differentiation. A paradoxical feature of rhabdomyosarcoma (RMS), a tumor arising from muscle precursors, is the block of the differentiation program and the deregulated proliferation despite MyoD expression. A deficiency in RMS of a factor required for MyoD activity has been implicated by previous studies. We report here that p38 MAP kinase (MAPK) activation, which is essential for muscle differentiation, is deficient in RMS cells. Enforced induction of p38 MAPK by an activated MAPK kinase 6 (MKK6EE) restored MyoD function and enhanced MEF2 activity in RMS deficient for p38 MAPK activation, leading to growth arrest and terminal differentiation. Stress and cytokines could activate the p38 MAPK in RMS cells, however, these stimuli did not promote differentiation, possibly because they activated p38 MAPK only transiently and they also activated JNK, which could antagonize differentiation. Thus, the selective and sustained p38 MAPK activation, which is distinct from the stress-activated response, is required for differentiation and can be disrupted in human tumors. PMID- 10716946 TI - WISP-1 is a Wnt-1- and beta-catenin-responsive oncogene. AB - WISP-1 (Wnt-1 induced secreted protein 1) is a member of the CCN family of growth factors. This study identifies WISP-1 as a beta-catenin-regulated gene that can contribute to tumorigenesis. The promoter of WISP-1 was cloned and shown to be activated by both Wnt-1 and beta-catenin expression. TCF/LEF sites played a minor role, whereas the CREB site played an important role in this transcriptional activation. WISP-1 demonstrated oncogenic activities; overexpression of WISP-1 in normal rat kidney fibroblast cells (NRK-49F) induced morphological transformation, accelerated cell growth, and enhanced saturation density. Although these cells did not acquire anchorage-independent growth in soft agar, they readily formed tumors in nude mice, suggesting that appropriate cellular attachment is important for signaling oncogenic events downstream of WISP-1. PMID- 10716947 TI - The caenorhabditis elegans fate-determining gene mab-9 encodes a T-box protein required to pattern the posterior hindgut. AB - Caenorhabditis elegans mab-9 mutants are defective in hindgut and male tail development because of cell fate transformations in two posterior blast cells, B and F. We have cloned mab-9 and show that it encodes a member of the T-box family of transcriptional regulators. MAB-9 localizes to the nucleus of B and F and their descendents during development, suggesting that it acts cell autonomously in the posterior hindgut to direct cell fate. T-box genes related to brachyury have also been implicated in hindgut patterning, and our results support models for an evolutionarily ancient role for these genes in hindgut formation. PMID- 10716948 TI - The Drosophila shark tyrosine kinase is required for embryonic dorsal closure. AB - Dorsal closure (DC) in the Drosophila embryo requires the coordinated interaction of two different functional domains of the epidermal cell layer-the leading edge (LE) and the lateral epidermis. In response to activation of a conserved c-Jun amino-terminal kinase (JNK) signaling module, the dorsal-most layer of cells, which constitute the LE of the stretching epithelial sheet, secrete Dpp, a member of the TGFbeta superfamily. Dpp and other LE cell-derived signaling molecules stimulate the bilateral dorsal elongation of cells of the dorsolateral epidermis over the underlaying amnioserosa and the eventual fusion of their LEs along the dorsal midline. We have found that flies bearing a Shark tyrosine kinase gene mutation, shark(1), exhibit a DC-defective phenotype. Dpp fails to be expressed in shark(1) mutant LE cells. Consistent with these observations, epidermal specific reconstitution of shark function or overexpression of an activated form of c-Jun in the shark(1) mutant background, rescues the DC defect. Thus, Shark regulates the JNK signaling pathway leading to Dpp expression in LE cells. Furthermore, constitutive activation of the Dpp pathway throughout the epidermis fails to rescue the shark(1) DC defect, suggesting that Shark may function in additional pathways in the LE and/or lateral epithelium. PMID- 10716950 TI - MEMORANDUM FOR: science writers and editors on the journal press list: study of adult twins shows tendency to develop moles has a strong genetic component PMID- 10716951 TI - MEMORANDUM FOR: science writers and editors on the journal press list: new measurements show that a protein involved in vitamin A metabolism is missing in 24% of breast cancers PMID- 10716949 TI - A major surface glycoprotein of trypanosoma brucei is expressed transiently during development and can be regulated post-transcriptionally by glycerol or hypoxia. AB - Differentiation is a means by which unicellular parasites adapt to different environments. In some cases, the developmental program may be modulated by interactions with the host, but the mechanisms are largely unknown. Trypanosoma brucei is transmitted between mammals by tsetse flies. The development of the procyclic form in the tsetse midgut is marked by the synthesis of a new glycoprotein coat, composed of EP and GPEET procyclins, that is important for survival. Here we demonstrate that the composition of the coat changes in response to extracellular signals in vitro and during development in vivo. EP and GPEET are coinduced when differentiation is initiated. Subsequently, EP expression is maintained, whereas GPEET is repressed after 7-9 days. The timepoint at which GPEET is repressed coincides with the appearance of parasites in a new compartment of the fly midgut. In culture, down-regulation of GPEET can be prevented by exogenous glycerol or accelerated by hypoxia. Regulation is post transcriptional, and is conferred by the GPEET 3' untranslated region. The same sequence also regulates expression of a reporter gene in the fly. The finding that GPEET is expressed during a defined window during the establishment of infection suggests that it has a specific function in host-parasite interactions rather than a generalized role in shielding underlying membrane molecules. PMID- 10716952 TI - Aberrant retinoid signaling and breast cancer: the view from outside the nucleus. PMID- 10716953 TI - DNA repair: a double-edged sword. PMID- 10716955 TI - Breast density and cancer risk: what is the relationship? PMID- 10716954 TI - FDA approves system for digital mammography. PMID- 10716957 TI - Breast cancer statistics PMID- 10716956 TI - Inner-city mammography programs aim to make breast cancer "a visible disease". PMID- 10716958 TI - Wanted: senior citizens for cancer treatment trials. PMID- 10716959 TI - Now showing on HBO: cancer documentary makes prime time. PMID- 10716960 TI - New drugs, better treatments lessen side effects of cancer care. PMID- 10716961 TI - Stat bite: Use of mammography and annual household income, 1991-1997. PMID- 10716962 TI - Participation in the cooperative family registry for breast cancer studies: issues of informed consent. PMID- 10716963 TI - Genetics of risk factors for melanoma: an adult twin study of nevi and freckles. AB - BACKGROUND: We sought by use of an adult twin study to investigate the relative contribution of genetic and environmental effects on the expression of nevi and freckles, which are known risk factors for melanoma, and to determine if age and sun exposure influence the heritability of nevi. DESIGN AND METHODS: Total nevus and freckle counts were conducted on 127 monozygotic twin pairs and 323 dizygotic twin pairs. Intraclass correlations were calculated by use of analysis of variance. Model-fitting analyses were performed to quantify the genetic and environmental components of the variance for nevus and freckle counts. RESULTS: The intraclass correlation for total nevus counts was.83 in monozygotic pairs compared with.51 in dizygotic pairs. Quantitative genetic analyses showed that the contribution of genetic factors on nevi expression varied according to age. For twins less than 45 years old, the additive genetic variance on total nevus count was 36% (95% confidence interval [CI] = 0.8%-63%), with 38% (95% CI = 14% 61%) and 26% (95% CI = 16%-42%) of the remaining variance attributed to common environment and unique environmental effects, respectively. In twins aged 45 years or older, common environmental effects on total nevus count became negligible, with the additive genetic variance increasing to 84% (95% CI = 77% 88%). Body site was also found to affect the heritability estimates for nevus counts, with a statistically significant difference between sun-exposed and sun protected sites. The polychoric correlation (i.e., the correlation in liability within twins for more than two categories) for total freckle counts was.91 in monozygotic twin pairs compared with.54 in dizygotic twin pairs. Additive genetic effects explained 91% (95% CI = 86%-94%) of the variance in freckle counts. CONCLUSION: The contribution of genetic factors on the variance for total nevus counts increased with age, and sun exposure appears to influence the expression of nevi. The results of this study highlight the need to take into account the age and site of nevus counts for future genetic linkage or association studies in the search for new melanoma genes. PMID- 10716964 TI - Population-based study of human papillomavirus infection and cervical neoplasia in rural Costa Rica. AB - BACKGROUND: Human papillomavirus (HPV) is the main cause of cervical neoplasia. Because few population-based studies have investigated the prevalence of type specific infection in relation to cervical disease, we studied a high-risk population, estimating the prevalence of HPV infection and the risk associated with various HPV types. METHODS: We screened 9175 women in Guanacaste, Costa Rica, to obtain a referent standard final diagnosis, and tested 3024 women for more than 40 types of HPV with a polymerase chain reaction-based system. RESULTS: Among women with normal cytology, HPV infections peaked first in women younger than 25 years, and they peaked again at age 55 years or older with predominantly non-cancer-associated types of HPV and uncharacterized HPV types. Low-grade squamous intraepithelial lesions (LSILs) (n = 189) decreased consistently with age. The prevalence of high-grade squamous intraepithelial lesions (HSILs) (n = 128) peaked first around age 30 years and again at age 65 years or older. Seventy three percent of LSILs were HPV positive, with HPV16 being the predominant type (16% of positive subjects). HPV was found in 89% of HSILs and 88% of cancers, with HPV16 being strongly predominant (51% and 53% of positive subjects). Virtually all HSILs and cancers had cancer-associated HPV types, with high odds ratios (ORs) and attributable fractions around 80%. Risk for HPV16 was particularly high (OR for HSILs = 320, 95% confidence interval [CI] = 97-1000; OR for cancer = 710, 95% CI = 110-4500). CONCLUSIONS: We confirm the early decline of HPV infection with age but note increased prevalence after menopause, which could be related to a second peak of HSILs, an observation that warrants further investigation. At least 80% of HPVs involved in cervical carcinogenesis in this population have been characterized. Polyvalent vaccines including the main cancer associated HPV types may be able to prevent most cases of cervical disease in this region. PMID- 10716965 TI - Cellular retinol-binding protein expression and breast cancer. AB - BACKGROUND: The biologic activity of vitamin A depends, in part, on its metabolism to active nuclear receptor ligands, chiefly retinoic acid. The cellular retinol-binding protein (CRBP) binds vitamin A with high affinity and is postulated to regulate its uptake and metabolism. In this report, we analyze the expression of CRBP in normal and malignant breast tissues. METHODS: We evaluated CRBP expression by in situ hybridization in six reduction mammoplasty specimens and 49 human breast carcinoma specimens by use of digoxigenin-labeled RNA probes and in nine cultured mammoplasty specimens by northern or western blot analysis. Statistical significance was evaluated with the chi(2) test or Fisher's exact test if the sample sizes were small. All P values are from two-sided tests. RESULTS: CRBP was expressed in all 15 mammoplasty specimens (normal breast tissue) and in 33 of 35 available specimens of normal tissue adjacent to carcinoma. In contrast, 12 (24%) of 49 carcinoma lesions were uniformly negative for CRBP (P =.023 for comparison with adjacent normal breast tissue). The loss of CRBP expression was as frequent in ductal carcinoma in situ (six [27%] of 22) as in invasive lesions (six [22%] of 27), suggesting that it is a relatively early event in carcinogenesis and not associated with patient age, tumor grade, and expression of steroid receptors or c-Myc. Preliminary experiments did not find an association between CRBP and retinoic acid receptor beta loss, but most (four of five) CRBP-negative tumors were also retinoic acid receptor beta negative. CONCLUSION: CRBP is underexpressed in 24% (95% confidence interval = 12.5%-36.5%) of human breast carcinomas, implying a link between cellular vitamin A homeostasis and breast cancer. We hypothesize that the loss of CRBP restricts the effects of endogenous vitamin A on breast epithelial cells. PMID- 10716966 TI - Cytotoxicity and mutagenicity of frameshift-inducing agent ICR191 in mismatch repair-deficient colon cancer cells. AB - BACKGROUND: Deficiency of DNA mismatch repair is a common feature of cancers exhibiting instability of microsatellite DNA sequences. Cancers with microsatellite instability are recognizable by their high rate of spontaneous frameshift mutations within microsatellite sequences, their resistance to killing by cytotoxic agents, and their localization to specific tissues, e.g., the proximal colon and stomach. We hypothesized that the mismatch repair deficiency of these cancers would make them vulnerable to environmental or chemical frameshift-inducing agents. This study was undertaken to test whether exogenous frameshift-inducing agents selectively induce mutations in mismatch repair deficient cells of mutagen-exposed tissues like the colon and whether cytotoxic doses of these agents would preferentially kill those cells. METHODS: Cytotoxicity of the acridine mutagen 6-chloro-9-[3-(2 chloroethylamino)propylamino]-2-methoxy-acridine (ICR191), a DNA frameshift inducer, was determined in the mismatch repair-deficient human colon carcinoma cell line HCT116 versus the repair-reconstituted derivative HCT116+C3. Vulnerability to the mutagenic effects of ICR191 was determined by transfection of HCT116 or HCT116+C3 cells with a frameshift reporter vector, followed by treatment of the cells with ICR191. Alternatively, the reporter vector was reacted ex vivo with ICR191, and the derivatized vector was then transfected into HCT116 or HCT116+C3 cells. RESULTS: ICR191 proved to be fivefold to 10-fold more potent in inducing mutations in mismatch repair-deficient HCT116 cells than in mismatch repair-proficient HCT116+C3 cells. Moreover, at cytotoxic doses of ICR191, repair-deficient HCT116 cells proved to be fivefold more vulnerable to killing than did HCT116+C3 cells. CONCLUSIONS: Frameshift-inducing mutagens can selectively induce mutations in mismatch repair-deficient cells versus mismatch repair-proficient cells. Environmental exposures may, therefore, favor development of cancers with microsatellite instability in tissues like the gut. Frameshift-inducing agents can, however, also preferentially kill mismatch repair deficient cancer cells and, thus, may be promising as model therapeutic compounds. PMID- 10716967 TI - Association of angiogenesis in lymph node metastases with outcome of breast cancer. AB - BACKGROUND: Microvessel density (MVD) is a measure of the extent of new blood vessel growth or angiogenesis, which is required for tumor progression. Increased MVD in primary breast cancers appears to adversely affect disease-free survival and overall survival in patients with breast cancer. However, the clinical implications of angiogenesis in breast cancer metastases have not been well studied. The purpose of this study was to compare intratumoral MVD in primary breast cancer tissues with MVD in axillary lymph node metastases and to evaluate the relationships among primary- and metastatic-tumor MVD, disease-free survival, and overall survival in patients with lymph node-positive, stage II breast cancer who were treated with adjuvant chemotherapy in Cancer and Leukemia Group B Protocol 8082. METHODS: Immunostaining for factor VIII-related antigen was performed on tissue sections from 47 primary tumors and 91 axillary lymph nodes containing metastases from 110 patients with lymph node-positive breast cancer. Sections were examined for the presence or absence of focal areas of relatively intense neovascularization (vascular hot spots), and a quantitative assessment of intratumoral MVD was performed. RESULTS: The presence of vascular hot spots in axillary lymph node metastases, but not primary breast cancers, was associated with statistically significantly decreased disease-free survival (P =.006) and overall survival (P =.004) by univariate analysis. Similarly, increased MVD in metastases, but not in primary tumors, was statistically significantly associated with diminished overall survival in these patients (P =.02). In multivariate analysis, the number of positive axillary lymph nodes and the presence of vascular hot spots in axillary lymph node metastases predicted decreased disease free survival (P =.0001 and.02, respectively) and overall survival (P =.0001 and.007, respectively). All P values were two-sided. CONCLUSION: This pilot study suggests that assessing neovascularization in axillary lymph node metastases may provide clinically useful information regarding survival in patients with primary breast cancer. PMID- 10716969 TI - RESPONSE: re: scientific interest in newcastle disease virus is reviving PMID- 10716968 TI - Re: Scientific interest in newcastle disease virus is reviving. PMID- 10716970 TI - Re: Cytosine deaminase/5-fluorocytosine-based vaccination against liver tumors: evidence of distant bystander effect. PMID- 10716971 TI - More about: cell and molecular biology of simian virus 40: implications for human infections and disease. PMID- 10716974 TI - Eleventh Annual Meeting. AB - A joint meeting with the Behavioral Neurology SocietySunday-Tuesday, February 20 22, 2000Sanibel Harbour Resort and Spa, Fort Myers, FL PMID- 10716972 TI - Response to more about: cell and molecular biology of simian virus 40: implications for human infections and disease. PMID- 10716976 TI - Insights into DNA replication from the third domain of life. PMID- 10716977 TI - The science of literacy: from the laboratory to the classroom. PMID- 10716978 TI - Connecting oxidative stress, auxin, and cell cycle regulation through a plant mitogen-activated protein kinase pathway. PMID- 10716980 TI - Monopoles and fractional vortices in chiral superconductors. AB - I discuss two exotic objects that must be experimentally identified in chiral superfluids and superconductors. These are (i) the vortex with a fractional quantum number (N = 1/2 in chiral superfluids, and N = 1/2 and N = 1/4 in chiral superconductors), which plays the part of the Alice string in relativistic theories and (ii) the hedgehog in the;l field, which is the counterpart of the Dirac magnetic monopole. These objects of different dimensions are topologically connected. They form the combined object that is called a nexus in relativistic theories. In chiral superconductors, the nexus has magnetic charge emanating radially from the hedgehog, whereas the half-quantum vortices play the part of the Dirac string. Each half-quantum vortex supplies the fractional magnetic flux to the hedgehog, representing 1/4 of the "conventional" Dirac string. I discuss the topological interaction of the superconductor's nexus with the 't Hooft Polyakov magnetic monopole, which can exist in Grand Unified Theories. The monopole and the hedgehog with the same magnetic charge are topologically confined by a piece of the Abrikosov vortex. Such confinement makes the nexus a natural trap for the magnetic monopole. Other properties of half-quantum vortices and monopoles are discussed as well, including fermion zero modes. PMID- 10716979 TI - Combinatorial target-guided ligand assembly: identification of potent subtype selective c-Src inhibitors. AB - A method for the rapid and efficient identification of ligands to biological targets is reported. The combinatorial method does not require structural or mechanistic information and is accomplished in four straightforward steps. (i) A set of potential binding elements is prepared wherein each molecule incorporates a common chemical linkage group. (ii) The set of potential binding elements is screened to identify all binding elements that interact even weakly with the biological target. (iii) A combinatorial library of linked binding elements is prepared whereby the binding elements are connected by the common chemical linkage groups through a set of flexible linkers. (iv) The combinatorial library is screened to identify the tightest-binding ligands. The utility of the method was demonstrated by the identification of a potent and subtype-selective small molecule inhibitor of the non-receptor tyrosine kinase c-Src (IC(50) = 64 nM). Because the method relies on connecting two distinct binding elements, the relative contributions of the two binding elements to the potency and selectivity of the inhibitor were readily determined. This information provides valuable insight into the molecular basis of inhibition. PMID- 10716981 TI - Ultrafast dynamics of many-body processes and fundamental quantum mechanical phenomena in semiconductors. AB - The large dielectric constant and small effective mass in a semiconductor allows a description of its electronic states in terms of envelope wavefunctions whose energy, time, and length scales are mesoscopic, i.e., halfway between those of atomic and those of condensed matter systems. This property makes it possible to demonstrate and investigate many quantum mechanical, many-body, and quantum kinetic phenomena with tabletop experiments that would be nearly impossible in other systems. This, along with the ability to custom-design semiconductor nanostructures, makes semiconductors an ideal laboratory for experimental investigations. We present an overview of some of the most exciting results obtained in semiconductors in recent years using the technique of ultrafast nonlinear optical spectrocopy. These results show that Coulomb correlation plays a major role in semiconductors and makes them behave more like a strongly interacting system than like an atomic system. The results provide insights into the physics of strongly interacting systems that are relevant to other condensed matter systems, but not easily accessible in other materials. PMID- 10716982 TI - Different functional domains of TAFII250 modulate expression of distinct subsets of mammalian genes. AB - The TATA box-binding protein-associated factors (TAFs) are thought to play an essential role in eukaryotic RNA polymerase II transcription by mediating the expression of distinct subsets of genes. In hamster ts13 cells, a single amino acid change in TAF(II)250, which disrupts its acetyl-transferase activity at the restrictive temperature, alters the transcription of specific genes involved in cell cycle control. Likewise, disruption of the amino-terminal kinase domain of TAF(II)250 results in transcriptional defects in ts13 cells. However, it was not known whether the acetyl-transferase or kinase domains of TAF(II)250 modulate specific classes of genes and whether these two domains regulate distinct subsets of genes. Here we have used high-density gene-profiling to identify mammalian transcripts that require either the TAF(II)250 acetyl-transferase or protein kinase function for proper expression. We found that transcription of at least 18% of genes are differentially expressed at the restrictive temperature. The promoter region of one of these genes was subsequently characterized, and both upstream elements as well as the core promoter were shown to be TAF(II)250 responsive. We also found that expression of approximately 6% of genes in ts13 cells requires a functional TAF(II)250 amino-terminal kinase domain, but only approximately 1% of these hamster genes also require the TAF(II)250 acetyl transferase activity. Our results suggest that the two TAF(II)250 enzymatic activities are important for regulating largely nonoverlapping sets of genes involved in a wide range of biological functions in vivo. PMID- 10716983 TI - Ribosomal subunit kinase-2 is required for growth factor-stimulated transcription of the c-Fos gene. AB - Ribosomal subunit kinases (Rsk) have been implicated in the regulation of transcription by phosphorylating and thereby activating numerous transcription factors, such as c-Fos, cAMP responsive element binding protein (CREB), and nuclear receptors. Here we describe the generation and characterization of immortalized embryonic fibroblast cell lines from mice in which the Rsk-2 gene was disrupted by homologous recombinant gene targeting. Rsk-2-deficient (knockout or KO) cell lines have no detectable Rsk-2 protein, whereas Rsk-1 expression is unaltered as compared with cell lines derived from wild-type control mice. KO cells exhibit a major reduction in platelet-derived growth factor (PDGF) and insulin-like growth factor (IGF)-1-stimulated expression of the immediate-early gene c-Fos. This results primarily from a reduced transcriptional activation of the ternary complex factor Elk-1 and reduced activation of the serum response factor. The reduced Elk-1 activation in KO cells occurs despite normal activation of the mitogen-activated protein kinase pathway and normal PDGF- and IGF-1 stimulated Elk-1 phosphorylation. By contrast, PDGF- and IGF-1-stimulated phosphorylation and transcriptional activation of CREB is unaltered in KO cells. Thus Rsk-2 is required for growth factor-stimulated expression of c-Fos and transcriptional activation of Elk-1 and the serum response factor, but not for activation of CREB or the mitogen-activated protein kinase pathway in response to PDGF and IGF-1 stimulation. PMID- 10716984 TI - Yeast ribonucleotide reductase has a heterodimeric iron-radical-containing subunit. AB - Ribonucleotide reductase (RNR) catalyzes the de novo synthesis of deoxyribonucleotides. Eukaryotes have an alpha(2)beta(2) form of RNR consisting of two homodimeric subunits, proteins R1 (alpha(2)) and R2 (beta(2)). The R1 protein is the business end of the enzyme containing the active site and the binding sites for allosteric effectors. The R2 protein is a radical storage device containing an iron center-generated tyrosyl free radical. Previous work has identified an RNR protein in yeast, Rnr4p, which is homologous to other R2 proteins but lacks a number of conserved amino acid residues involved in iron binding. Using highly purified recombinant yeast RNR proteins, we demonstrate that the crucial role of Rnr4p (beta') is to fold correctly and stabilize the radical-storing Rnr2p by forming a stable 1:1 Rnr2p/Rnr4p complex. This complex sediments at 5.6 S as a betabeta' heterodimer in a sucrose gradient. In the presence of Rnr1p, both polypeptides of the Rnr2p/Rnr4p heterodimer cosediment at 9.7 S as expected for an alpha(2)betabeta' heterotetramer, where Rnr4p plays an important role in the interaction between the alpha(2) and the betabeta ' subunits. The specific activity of the Rnr2p complexed with Rnr4p is 2,250 nmol deoxycytidine 5'-diphosphate formed per min per mg, whereas the homodimer of Rnr2p shows no activity. This difference in activity may be a consequence of the different conformations of the inactive homodimeric Rnr2p and the active Rnr4p bound form, as shown by CD spectroscopy. Taken together, our results show that the Rnr2p/Rnr4p heterodimer is the active form of the yeast RNR small subunit. PMID- 10716985 TI - Zinc transfer potentials of the alpha - and beta-clusters of metallothionein are affected by domain interactions in the whole molecule. AB - The alpha- and beta-polypeptides of human metallothionein (isoform 2), obtained by chemical synthesis, were converted into their respective zinc/thiolate clusters, and each domain was investigated separately. Proton titration data for the N-terminal beta-domain fit a simple model with three ionizations of the same apparent pK(a) value of 4.9 and a collective binding constant for zinc of 5 x 10( 12) M at pH 7.0. The zinc cluster in the C-terminal alpha-domain is more stable than that in the beta-domain. Its pH titration is also more complex, indicating at least two classes of zinc sites with different affinities. The whole molecule is stabilized with regard to the individual domains. Chemical modification implicates lysine side chains in both the stabilization of the beta-domain cluster and the mutual stabilization of the domains in the whole molecule. The two zinc clusters also differ in the reactivity of their cysteine sulfurs and their potential to donate zinc to an acceptor molecule dependent on its type and characteristics. The isolated beta-domain cluster reacts faster with Ellman's reagent and is a better zinc donor toward zinc-depleted sorbitol dehydrogenase than is the isolated alpha-domain cluster, whereas the reverse is observed when a chelating agent is the zinc acceptor. Thus, although each cluster assembles independently of the other, the cumulative properties of the individual domains do not suffice to describe metallothionein either structurally or functionally. The two-domain structure of the whole molecule is important for its interaction with ligands and for control of its reactivity and overall conformation. PMID- 10716986 TI - Evidence for a requirement for ATP hydrolysis at two distinct steps during a single turnover of the catalytic cycle of human P-glycoprotein. AB - P-glycoprotein (Pgp) is an ATP-dependent hydrophobic natural product anticancer drug efflux pump whose overexpression confers multidrug resistance to tumor cells. The work reported here deals with the elucidation of the energy requirement for substrate interaction with Pgp during the catalytic cycle. We show that the K(d) (412 nM) of the substrate analogue [(125)I]iodoarylazidoprazoin for Pgp is not altered by the presence of the nonhydrolyzable nucleotide 5'-adenylylimididiphosphate and vanadate (K(d) = 403 nM). Though binding of nucleotide per se does not affect interactions with the substrate, ATP hydrolysis results in a dramatic conformational change where the affinity of [(125)I]iodoarylazidoprazoin for Pgp trapped in transition-state conformation (Pgp x ADP x vanadate) is reduced >30-fold. To transform Pgp from this intermediate state of low affinity for substrate to the next catalytic cycle, i.e., a conformation that binds substrate with high affinity, requires conditions that permit ATP hydrolysis. Additionally, there is an inverse correlation (R(2) = 0.96) between 8AzidoADP (or ADP) release and the recovery of substrate binding. These results suggest that the release of nucleotide is necessary for reactivation but not sufficient. The hydrolysis of additional molecule(s) of ATP (or 8AzidoATP) is obligatory for the catalytic cycle to advance to completion. These data are consistent with the observed stoichiometry of two ATP molecules hydrolyzed for the transport of every substrate molecule. Our data demonstrate two distinct roles for ATP hydrolysis in a single turnover of the catalytic cycle of Pgp, one in the transport of substrate and the other in effecting conformational changes to reset the pump for the next catalytic cycle. PMID- 10716987 TI - Extension of transverse relaxation-optimized spectroscopy techniques to allosteric proteins: CO- and paramagnetic fluoromet-hemoglobin [beta (15N valine)]. AB - We present the first steps in applying transverse relaxation-optimized spectroscopy (TROSY) techniques to the study of allosterism. Each beta-chain of the hemoglobin (Hb) tetramer has 17 valine residues. We have (15)N-labeled the beta-chain Val residues and detected 16 of the 17 (1)H-(15)N correlation peaks for beta-chain Val of the R state CO-Hb structure by using the TROSY technique. Sequence-specific assignments are suggested, based mainly on analysis of the (1)H pseudocontact-shift increments produced by oxidizing the diamagnetic R state HbCO to the paramagnetic R state fluoromet form. When possible, we support these assignments with sequential nuclear Overhauser effect (NOE) information obtained from a two-dimensional [(1)H,(1)H]-NOESY-TROSY experiment (NOESY, NOE spectroscopy). We have induced further the R-T conformational change by adding the allosteric effector, inositol hexaphosphate, to the fluoromet-Hb sample. This change induces substantial increments in the (1)H and (15)N chemical shifts, and we discuss the implication of these findings in the context of the tentative sequence assignments. These preliminary results suggest that amide nitrogen and amide proton chemical shifts in a selectively labeled sample are site-specific probes for monitoring the allosteric response of the ensemble-averaged solution structure of Hb. More important, the chemical-shift dispersion obtained is adequate to permit a complete assignment of the backbone (15)N/(13)C resonances upon nonselective labeling. PMID- 10716988 TI - Mechanisms and kinetics of beta-hairpin formation. AB - Thermodynamics and kinetics of off-lattice models with side chains for the beta hairpin fragment of immunoglobulin-binding protein and its variants are reported. For all properties (except refolding time tau(F)) there are no qualitative differences between the full model and the Go version. The validity of the models is established by comparison of the calculated native structure with the Protein Data Bank coordinates and by reproducing the experimental results for the degree of cooperativity and tau(F). For the full model tau(F) approximately 2 micros at the folding temperature (experimental value is 6 micros); the Go model folds 50 times faster. Upon refolding, structural changes take place over three time scales. On the collapse time scale compact structures with intact hydrophobic cluster form. Subsequently, hydrogen bonds form, predominantly originating from the turn by a kinetic zipping mechanism. The assembly of the hairpin is complete when most of the interstrand contacts (the rate-limiting step) is formed. The dominant transition state structure (located by using cluster analysis) is compact and structured. We predict that when hydrophobic cluster is moved to the loop tau(F) marginally increases, whereas moving the hydrophobic cluster closer to the termini results in significant decrease in tau(F) relative to wild type. The mechanism of hairpin formation is predicted to depend on turn stiffness. PMID- 10716989 TI - The retro-GCN4 leucine zipper sequence forms a stable three-dimensional structure. AB - The question of whether a protein whose natural sequence is inverted adopts a stable fold is still under debate. We have determined the 2. 1-A crystal structure of the retro-GCN4 leucine zipper. In contrast to the two-stranded helical coiled-coil GCN4 leucine zipper, the retro-leucine zipper formed a very stable, parallel four-helix bundle, which now lends itself to further structural and functional studies. PMID- 10716991 TI - Mik1 levels accumulate in S phase and may mediate an intrinsic link between S phase and mitosis. AB - Two paradigms exist for maintaining order during cell-cycle progression: intrinsic controls, where passage through one part of the cell cycle directly affects the ability to execute another, and checkpoint controls, where external pathways impose order in response to aberrant structures. By studying the mitotic inhibitor Mik1, we have identified evidence for an intrinsic link between unperturbed S phase and mitosis. We propose a model in which S/M linkage can be generated by the production and stabilization of Mik1 protein during S phase. The production of Mik1 during unperturbed S phase is independent of the Rad3- and Cds1-dependent checkpoint controls. In response to perturbed S phase, Rad3-Cds1 checkpoint controls are required to maintain high levels of Mik1, probably indirectly by extending the S phase period, where Mik1 is stable. In addition, we find that Mik1 protein can be moderately induced in response to irradiation of G(2) cells in a Chk1-dependent manner. PMID- 10716990 TI - Identification and localization of two brefeldin A-inhibited guanine nucleotide exchange proteins for ADP-ribosylation factors in a macromolecular complex. AB - Two brefeldin A (BFA)-inhibited guanine nucleotide-exchange proteins for ADP ribosylation factors, 200-kDa BIG1 and 190-kDa BIG2, were copurified from bovine brain cytosol associated with >670-kDa macromolecular complexes. When observed by immunofluorescence in HeLa S3 and HepG2 cells, endogenous BIG1 and coexpressed BIG2 were distributed in a punctate pattern throughout the cytosol, and also concentrated in the perinuclear region, where endogenous BIG1 and BIG2 each partially colocalized with Golgi-specific 58K protein and gamma-adaptin. On Western blot analysis, both BIG1 and BIG2 were clearly more abundant in the cytosol than in the microsomal fractions. After density gradient centrifugation of a microsomal fraction, BIG1 and BIG2 were recovered in the same fraction as beta-COP, a marker for Golgi membranes. When cytosol from HeLa S3 cells was subjected to gel filtration and fractions were analyzed by Western blotting, the largest percentages of both BIG1 and BIG2 were detected in fractions containing proteins with a molecular mass of >670 kDa. Western blotting using anti-peptide antibodies specific for BIG1 or BIG2 demonstrated that approximately 70% of BIG2 was immunoprecipitated along with 100% of BIG1 by the anti-BIG1 IgG, and approximately 75% of BIG1 was coprecipitated with 100% of BIG2 by the anti-BIG2 IgG. All observations were consistent with the conclusion that significant fractions of BIG1 and BIG2 exist as components of the same macromolecular complexes in bovine brain cytosol and are similarly localized in cultured cells. PMID- 10716992 TI - BAR: An apoptosis regulator at the intersection of caspases and Bcl-2 family proteins. AB - Two major pathways for induction of apoptosis have been identified-intrinsic and extrinsic. The extrinsic pathway is represented by tumor necrosis factor family receptors, which utilize protein interaction modules known as death domains and death effector domains (DEDs) to assemble receptor signaling complexes that recruit and activate certain caspase-family cell death proteases, namely procaspases-8 and -10. The intrinsic pathway for apoptosis involves the participation of mitochondria, which release caspase-activating proteins. Bcl-2 family proteins govern this mitochondria-dependent apoptosis pathway, with proteins such as Bax functioning as inducers and proteins such as Bcl-2 and Bcl X(L) serving as suppressors of cell death. An apoptosis regulator, BAR, was identified by using a yeast-based screen for inhibitors of Bax-induced cell death. The BAR protein contains a SAM domain, which is required for its interactions with Bcl-2 and Bcl-X(L) and for suppression of Bax-induced cell death in both mammalian cells and yeast. In addition, BAR contains a DED-like domain responsible for its interaction with DED-containing procaspases and suppression of Fas-induced apoptosis. Furthermore, BAR can bridge procaspase-8 and Bcl-2 into a protein complex. The BAR protein is anchored in intracellular membranes where Bcl-2 resides. BAR therefore may represent a scaffold protein capable of bridging two major apoptosis pathways. PMID- 10716993 TI - Activin receptor-like kinase 1 modulates transforming growth factor-beta 1 signaling in the regulation of angiogenesis. AB - The activin receptor-like kinase 1 (ALK1) is a type I receptor for transforming growth factor-beta (TGF-beta) family proteins. Expression of ALK1 in blood vessels and mutations of the ALK1 gene in human type II hereditary hemorrhagic telangiectasia patients suggest that ALK1 may have an important role during vascular development. To define the function of ALK1 during development, we inactivated the ALK1 gene in mice by gene targeting. The ALK1 homozygous embryos die at midgestation, exhibiting severe vascular abnormalities characterized by excessive fusion of capillary plexes into cavernous vessels and hyperdilation of large vessels. These vascular defects are associated with enhanced expression of angiogenic factors and proteases and are characterized by deficient differentiation and recruitment of vascular smooth muscle cells. The blood vessel defects in ALK1-deficient mice are reminiscent of mice lacking TGF-beta1, TGF beta type II receptor (TbetaR-II), or endoglin, suggesting that ALK1 may mediate TGF-beta1 signal in endothelial cells. Consistent with this hypothesis, we demonstrate that ALK1 in endothelial cells binds to TGF-beta1 and TbetaR-II. Furthermore, the ALK1 signaling pathway can inhibit TGF-beta1-dependent transcriptional activation mediated by the known TGF-beta1 type I receptor, ALK5. Taken together, our results suggest that the balance between the ALK1 and ALK5 signaling pathways in endothelial cells plays a crucial role in determining vascular endothelial properties during angiogenesis. PMID- 10716994 TI - Defective embryonic neurogenesis in Ku-deficient but not DNA-dependent protein kinase catalytic subunit-deficient mice. AB - Mammalian nonhomologous DNA end joining employs Ku70, Ku80, DNA-dependent protein kinase catalytic subunit (DNA-PKcs), XRCC4, and DNA ligase IV (Lig4). Herein, we show that Ku70 and Ku80 deficiency but not DNA-PKcs deficiency results in dramatically increased death of developing embryonic neurons in mice. The Ku deficient phenotype is qualitatively similar to, but less severe than, that associated with XRCC4 and Lig4 deficiency. The lack of a neuronal death phenotype in DNA-PKcs-deficient embryos and the milder phenotype of Ku-deficient versus XRCC4- or Lig4-deficient embryos correlate with relative leakiness of residual end joining in these mutant backgrounds as assayed by a V(D)J recombination end joining assay. We conclude that normal development of the nervous system depends on the four evolutionarily conserved nonhomologous DNA end joining factors. PMID- 10716995 TI - The genetics of ivermectin resistance in Caenorhabditis elegans. AB - The ability of organisms to evolve resistance threatens the effectiveness of every antibiotic drug. We show that in the nematode Caenorhabditis elegans, simultaneous mutation of three genes, avr-14, avr-15, and glc-1, encoding glutamate-gated chloride channel (GluCl) alpha-type subunits confers high-level resistance to the antiparasitic drug ivermectin. In contrast, mutating any two channel genes confers modest or no resistance. We propose a model in which ivermectin sensitivity in C. elegans is mediated by genes affecting parallel genetic pathways defined by the family of GluCl genes. The sensitivity of these pathways is further modulated by unc-7, unc-9, and the Dyf (dye filling defective) genes, which alter the structure of the nervous system. Our results suggest that the evolution of drug resistance can be slowed by targeting antibiotic drugs to several members of a multigene family. PMID- 10716996 TI - Gene microarray identification of redox and mitochondrial elements that control resistance or sensitivity to apoptosis. AB - Multigenic programs controlling susceptibility to apoptosis in response to ionizing radiation have not yet been defined. Here, using DNA microarrays, we show gene expression patterns in an apoptosis-sensitive and apoptosis-resistant murine B cell lymphoma model system both before and after irradiation. From the 11,000 genes interrogated by the arrays, two major patterns emerged. First, before radiation exposure the radioresistant LYar cells expressed significantly greater levels of message for several genes involved in regulating intracellular redox potential. Compared with LYas cells, LYar cells express 20- to 50-fold more mRNA for the tetraspanin CD53 and for fructose-1,6-bisphosphatase. Expression of both of these genes can lead to the increase of total cellular glutathione, which is the principle intracellular antioxidant and has been shown to inhibit many forms of apoptosis. A second pattern emerged after radiation, when the apoptosis sensitive LYas cells induced rapid expression of a unique cluster of genes characterized by their involvement in mitochondrial electron transport. Some of these genes have been previously recognized as proapoptotic; however others, such as uncoupling protein 2, were not previously known to be apoptotic regulatory proteins. From these observations we propose that a multigenic program for sensitivity to apoptosis involves induction of transcripts for genes participating in mitochondrial uncoupling and loss of membrane potential. This program triggers mitochondrial release of apoptogenic factors and induces the "caspase cascade." Conversely, cells resistant to apoptosis down-regulate these biochemical pathways, while activating pathways for establishment and maintenance of high intracellular redox potential by means of elevated glutathione. PMID- 10716997 TI - Polyclonal antibodies from patients immunized with a globo H-keyhole limpet hemocyanin vaccine: isolation, quantification, and characterization of immune responses by using totally synthetic immobilized tumor antigens. AB - We have previously reported on a carbohydrate-based vaccine program for immunotherapy in cancer patients. One such vaccine, based on the globo H antigen conjugated to the protein keyhole limpet hemocyanin (KLH), has been in clinical evaluation. Although this and other carbohydrate vaccines have been shown to induce antibody responses, there are currently no quantitative data on the antibody levels achieved in immunized patients by these or other anti-cancer vaccines. We report herein an efficient route to complex synthetic oligosaccharides attached to an affinity matrix for identifying and isolating antibodies elicited against such a carbohydrate-based vaccine in humans. Pre- and postvaccination profiles from serum samples of patients immunized with globo H KLH were compared. All anti-globo H antibody activity was efficiently separated from other serum constituents. The isolated antibodies were readily quantified, and their specificities were analyzed. Since no comparable data were available on antibodies resulting from the vaccination of other cancer patients, we compared the observed levels with those quoted in studies with bacterial polysaccharide vaccines that had been quantified. Remarkably, cancer patients immunized with globo H-KLH produce anti-globo H antibody levels often exceeding those formed by immunization with bacterial polysaccharides. In addition, substantial quantities of both IgG and IgM antibodies were elicited, clearly indicating a class switch to IgG. Taken together, these analyses serve to clarify several aspects of the immune response to the vaccine and give several new insights to the carbohydrate based vaccination strategy. Furthermore, antibodies so isolated could well have applications in clinical therapy. PMID- 10716998 TI - Redox regulation of chemokine receptor expression. AB - Cytokines and reactive oxygen intermediates (ROI) are frequent companions at sites of acute inflammation. We have shown previously that in human monocytes, bacterial lipopolysaccharide, IL-1, and tumor necrosis factor-alpha induce a rapid down-regulation of the monocyte chemotactic protein-1 receptor CCR2 (CC chemokine receptor-2). These stimuli also induce production of ROI. In this paper, we investigate the influence of antioxidants and/or ROI on chemokine receptor expression. In human monocytes, the antioxidant pyrrolidine dithiocarbamate (PDTC) rapidly inhibited CCR2 (95-100% of inhibition) and CCR5 (77-100% of inhibition) mRNA expression by strongly decreasing transcript stability. CCR2 half-life was decreased from 1.5 h to 45 min; CCR5 half-life was decreased from 2 h to 70 min. This inhibitory activity also included CXCR4 (CXC chemokine receptor-4) but not CXCR2 receptor and, although to a lesser extent, was shared by the antioxidants N-acetyl-l-cysteine and 2-mercaptoethanol. In contrast, the ROI-generating system xanthine/xanthine oxidase increased CCR5 and CXCR4 mRNA expression and counteracted the inhibitory effect of PDTC. Accordingly, H(2)O(2) and the glutathione-depleting drug buthionine sulfoximine increased to different extents CCR2, CCR5, and CXCR4 mRNA expression. The PDTC mediated inhibition of CCR5 and CXCR4 mRNA expression was associated with decreased chemotactic responsiveness (>90% inhibition) and with a marked inhibition of surface-receptor expression. In contrast, xanthine/xanthine oxidase opposed the bacterial lipopolysaccharide- and tumor necrosis factor-alpha mediated inhibition of CCR5 and CXCR4 mRNA expression and increased both the CCR5 surface expression and the cell migration (3-fold) in response to macrophage inflammatory protein-1beta. These results suggest that the redox status of cells is a crucial determinant in the regulation of the chemokine system. PMID- 10716999 TI - A mutant herpes simplex virus type 1 thymidine kinase reporter gene shows improved sensitivity for imaging reporter gene expression with positron emission tomography. AB - We are developing assays for noninvasive, quantitative imaging of reporter genes with positron emission tomography (PET), for application both in animal models and in human gene therapy. We report here a method to improve the detection of lower levels of PET reporter gene expression by utilizing a mutant herpes simplex virus type 1 thymidine kinase (HSV1-sr39tk) as a PET reporter gene. The HSV1 sr39tk mutant was identified from a library of site-directed mutants. Accumulation (net uptake) of the radioactively labeled substrates [8 (3)H]penciclovir ([8-(3)H]PCV), and 8-[(18)F]fluoropenciclovir (FPCV) in C6 rat glioma cells expressing HSV1-sr39tk is increased by a factor of approximately 2.0 when compared with C6 cells expressing wild-type HSV1-tk. The increased imaging sensitivity of HSV1-sr39tk when FPCV is used is also demonstrated in vivo both with tumor cells stably transfected with either HSV1-tk or HSV1-sr39tk, and after hepatic delivery of HSV1-tk or HSV1-sr39tk by using adenoviral vectors. The use of HSV1-sr39tk as a PET reporter gene and FPCV as a PET reporter probe results in significantly enhanced sensitivity for imaging reporter gene expression in vivo. PMID- 10717000 TI - Glucose stimulates protein modification by O-linked GlcNAc in pancreatic beta cells: linkage of O-linked GlcNAc to beta cell death. AB - The pancreatic beta cell can respond in the long term to hyperglycemia both with an increased capacity for insulin production and, in susceptible individuals, with apoptosis. When glucose-induced apoptosis offsets the increasing beta cell capacity, type 2 diabetes results. Here, we tested the idea that the pathway of glucose metabolism that leads to the modification of intracellular proteins with the O-linked monosaccharide N-acetylglucosamine (O-GlcNAc) is involved in the glucose-induced apoptosis. This idea is based on two recent observations. First, the beta cell expresses much more O-GlcNAc transferase than any other known cell, and second, that the beta cell-specific toxin, streptozotocin (STZ), itself a GlcNAc analog, specifically blocks the enzyme that cleaves O-GlcNAc from intracellular proteins. As a consequence, we now show that hyperglycemia leads to the rapid and reversible accumulation of O-GlcNAc specifically in beta cells in vivo. Animals pretreated with STZ also accumulate O-GlcNAc in their beta cells when hyperglycemic, but this change is sustained upon re-establishment of euglycemia. In concert with the idea that STZ toxicity results from the sustained accumulation of O-GlcNAc after a hyperglycemic episode, we established a low-dose STZ protocol in which the beta cells' toxicity of STZ was manifest only after glucose or glucosamine administration. Transgenic mice with impaired beta cell glucosamine synthesis treated with this protocol are resistant to the diabetogenic effect of STZ plus glucose yet succumb to STZ plus glucosamine. This study provides a causal link between apoptosis in beta cells and glucose metabolism through glucosamine to O-GlcNAc, implicating this pathway of glucose metabolism with beta cell glucose toxicity. PMID- 10717001 TI - Mitochondrial dysfunction during hypoxia/reoxygenation and its correction by anaerobic metabolism of citric acid cycle intermediates. AB - Kidney proximal tubule cells developed severe energy deficits during hypoxia/reoxygenation not attributable to cellular disruption, lack of purine precursors, the mitochondrial permeability transition, or loss of cytochrome c. Reoxygenated cells showed decreased respiration with complex I substrates, but minimal or no impairment with electron donors at complexes II and IV. This was accompanied by diminished mitochondrial membrane potential (DeltaPsi(m)). The energy deficit, respiratory inhibition, and loss of DeltaPsi(m) were strongly ameliorated by provision of alpha-ketoglutarate plus aspartate (alphaKG/ASP) supplements during either hypoxia or only during reoxygenation. Measurements of (13)C-labeled metabolites in [3-(13)C]aspartate-treated cells indicated the operation of anaerobic pathways of alphaKG/ASP metabolism to generate ATP, yielding succinate as end product. Anaerobic metabolism of alphaKG/ASP also mitigated the loss of DeltaPsi(m) that occurred during hypoxia before reoxygenation. Rotenone, but not antimycin or oligomycin, prevented this effect, indicating that electron transport in complex I, rather than F(1)F(0)-ATPase activity, had been responsible for maintenance of DeltaPsi(m) by the substrates. Thus, tubule cells subjected to hypoxia/reoxygenation can have persistent energy deficits associated with complex I dysfunction for substantial periods of time before onset of the mitochondrial permeability transition and/or loss of cytochrome c. The lesion can be prevented or reversed by citric acid cycle metabolites that anaerobically generate ATP by intramitochondrial substrate-level phosphorylation and maintain DeltaPsi(m) via electron transport in complex I. Utilization of these anaerobic pathways of mitochondrial energy metabolism known to be present in other mammalian tissues may provide strategies to limit mitochondrial dysfunction and allow cellular repair before the onset of irreversible injury by ischemia or hypoxia. PMID- 10717002 TI - Intracellular processing of endothelial nitric oxide synthase isoforms associated with differences in severity of cardiopulmonary diseases: cleavage of proteins with aspartate vs. glutamate at position 298. AB - An endothelial nitric oxide synthase (eNOS) polymorphism in exon 7 (894 G/T) resulting in glutamate or aspartate, respectively, at position 298 on the protein is correlated with severity of cardiopulmonary diseases. Because glutamate and aspartate are considered to be conservative replacements, the polymorphism was thought to be a marker for a functional locus elsewhere in the gene. We now show in transfected cells, primary human endothelial cells, and human hearts, that eNOS with aspartate, but not glutamate, at position 298 is cleaved, resulting in the generation of 100-kDa and 35-kDa products. Recombinant or native eNOS was examined by immunoblotting either in lysates (COS7) or after partial purification over 2',5'-ADP-Sepharose and calmodulin-Sepharose. Immunoblotting after SDS/PAGE with a carboxyl-terminal antibody showed a single major protein band in the predicted position for eNOS at 135 kDa. An additional band at approximately 100 kDa was present only in the recombinant 298Asp eNOS and in the eNOS synthesized by primary cells and heart tissue with a G/T genotype. Using an eNOS amino terminal-specific antibody, an immunoreactive band at approximately 35 kDa, corresponding to the residual N-terminal cleavage fragment, was observed in those cells with a T genotype. Thus, eNOS with aspartate but not glutamate at position 298 is cleaved, resulting in the generation of N-terminal 35-kDa and C-terminal 100-kDa fragments. Thus, the eNOS gene with polymorphisms at nucleotide 894 generates protein products with differing susceptibility to cleavage, suggesting that, in contrast to prior predictions, this polymorphism has a functional effect on the eNOS protein. PMID- 10717003 TI - Effects of heat shock, heat shock protein 40 (HDJ-2), and proteasome inhibition on protein aggregation in cellular models of Huntington's disease. AB - Huntington's disease (HD), spinocerebellar ataxias types 1 and 3 (SCA1, SCA3), and spinobulbar muscular atrophy (SBMA) are caused by CAG/polyglutamine expansion mutations. A feature of these diseases is ubiquitinated intraneuronal inclusions derived from the mutant proteins, which colocalize with heat shock proteins (HSPs) in SCA1 and SBMA and proteasomal components in SCA1, SCA3, and SBMA. Previous studies suggested that HSPs might protect against inclusion formation, because overexpression of HDJ-2/HSDJ (a human HSP40 homologue) reduced ataxin-1 (SCA1) and androgen receptor (SBMA) aggregate formation in HeLa cells. We investigated these phenomena by transiently transfecting part of huntingtin exon 1 in COS-7, PC12, and SH-SY5Y cells. Inclusion formation was not seen with constructs expressing 23 glutamines but was repeat length and time dependent for mutant constructs with 43-74 repeats. HSP70, HSP40, the 20S proteasome and ubiquitin colocalized with inclusions. Treatment with heat shock and lactacystin, a proteasome inhibitor, increased the proportion of mutant huntingtin exon 1 expressing cells with inclusions. Thus, inclusion formation may be enhanced in polyglutamine diseases, if the pathological process results in proteasome inhibition or a heat-shock response. Overexpression of HDJ-2/HSDJ did not modify inclusion formation in PC12 and SH-SY5Y cells but increased inclusion formation in COS-7 cells. To our knowledge, this is the first report of an HSP increasing aggregation of an abnormally folded protein in mammalian cells and expands the current understanding of the roles of HDJ-2/HSDJ in protein folding. PMID- 10717004 TI - Requirement of a leucine residue for (apical) membrane expression of type IIb NaPi cotransporters. AB - Type II NaPi cotransporters mediate epithelial phosphate (P(i)) reabsorption. In mammals the type IIb protein is expressed in the small intestinal apical membrane and other epithelia; it is not expressed in the renal proximal tubule where we find the type IIa isoform. To look for molecular determinant(s) involved in apical expression of type IIb cotransporters, we have made deletion mutations within the C-terminal tails of mouse IIb (mIIb) and human IIb (hIIb) transporter proteins. The constructs were fused to the enhanced green fluorescent protein and transiently transfected into intestinal CaCo2-cells. Both mIIb and hIIb were located exclusively in the apical membrane of the cells. For mIIb, the removal of a cysteine cluster or the last three amino acids (TVF) had no effect on the location of the protein. However, truncation at the level of the conserved L691/689 prevented the apical membrane expression of both mIIb and hIIb, respectively, and the mutated proteins were located in endosomal and lysosomal structures. A similar expression pattern of the mIIb and hIIb constructs was found in renal proximal tubular opossum kidney cells. Our data suggest that L691/689 is involved in mechanisms leading to an apical expression of type IIb NaPi cotransporters. PMID- 10717005 TI - The inhibitory effect of hormones associated with stress on Na appetite of sheep. AB - Stress is a large stimulus of Na appetite in rabbits, rats, and mice. This study investigated the influence of some peptides implicated in stress, i.e., adrenocorticotropin (ACTH), corticotropin-releasing factor (CRF), and the recently discovered member of the CRF family, urocortin, on the ingestive behavior of sheep. Intracerebroventricular infusion of these peptides over 4 days decreased the need-free Na intake of Na-repleted sheep. Intracerebroventricular infusion of urocortin, however, did not alter Na intake of Na-depleted sheep. Systemic infusion of ACTH increased, whereas systemic infusion of either urocortin or CRF decreased, Na intake of Na-repleted sheep. The increase in Na intake caused by the peripheral infusion of ACTH was blocked by concurrent i.v. infusion of urocortin, substantiating the inhibitory role of this peptide on Na appetite. Central administration of all peptides and i.v. administration of urocortin or urocortin and ACTH combined decreased food intake. Water intake was not directly influenced by the peptides. Rather, decreased water intake, when observed, was secondary to decreased food intake, as determined by pair-feeding experiments. Whereas systemic infusion of ACTH mimics the increase in Na intake observed in several different stressful situations, CRF and urocortin actually inhibit Na intake, indicating a direct central action overriding any effect of these peptides on ACTH release. Indeed, the inhibition of Na intake by urocortin occurred despite its stimulation of ACTH release and the subsequent increase in peripheral level of cortisol. Thus it would appear that hormones associated with stress have both excitatory and inhibitory influences on Na intake. Presumably, other physiological processes entrained by stress also will be important in determining the quantitative outcome on Na appetite. PMID- 10717006 TI - Rapid desensitization of the nitric oxide receptor, soluble guanylyl cyclase, underlies diversity of cellular cGMP responses. AB - A major receptor for nitric oxide (NO) is the cGMP-synthesizing enzyme, soluble guanylyl cyclase (sGC), but it is not known how this enzyme behaves in cells. In cerebellar cells, NO (from diethylamine NONOate) increased astrocytic cGMP with a potency (EC(50) /=15%-paclitaxel, docetaxel, vinorelbine, gemcitabine, CPT11-are described, and their implications for the design of new clinical trials are discussed. PMID- 10717171 TI - Introduction. PMID- 10717172 TI - The Pathology of Hodgkin's Disease: What's New? AB - From the viewpoint of the pathologist, there are three major issues concerning Hodgkin's disease (HD) that must be addressed. First is making the correct diagnosis; second is identifying any pathological features that may be of prognostic importance; and third is defining the identity of the neoplastic cell. Since Hodgkin's disease (HD) was first described, the origin of the Reed Sternberg cell and the pathogenesis of HD have remained elusive. Recently developed techniques have allowed investigators to analyze the immunophenotype and genotype of Reed-Sternberg cells. The results of these studies include recognition that, in many cases, Reed-Sternberg cells are of lymphoid origin. Thus, although in most cases of HD the diagnosis is straightforward on routinely stained sections, the conceptual dividing line between HD and non-Hodgkin's lymphoma is not as sharp as it once seemed. Occasionally, there is difficulty in distinguishing between HD and non-Hodgkin's lymphoma. Nevertheless, fundamental differences in the clinical features, histology, immunophenotype, and genotype between non-Hodgkin's lymphoma and HD remain. The impact of histological features on prognosis are controversial, largely because of the excellent prognosis of most cases of HD. PMID- 10717173 TI - Diagnostic Procedures and Guidelines for the Evaluation and Follow-up of Hodgkin's Disease. AB - Because of the success of various treatment approaches in most clinical settings of Hodgkin's disease (HD), it is now both possible and necessary to consider late effects and cost in the selection of a treatment plan for a given patient. Staging and therapeutic strategies are inextricably linked, with some treatment approaches requiring more staging information than others. As we evaluate the therapeutic options available, it is also appropriate to examine the roles of both standard and newly developed diagnostic procedures used in HD to determine their value and cost in the overall management approach. This article defines the role of staging in various treatment approaches, reviews the results achieved with available diagnostic procedures, and offers staging guidelines for various clinical settings. PMID- 10717174 TI - Prognostic Factors in Hodgkin's Disease. AB - Prognostic factors in Hodgkin's disease (HD) are reviewed. The Ann Arbor staging classification remains the basis for evaluation of patients with HD. However, subgroups of patients with differing prognoses exist within the individual stages. In pathological stages I and II, the number of involved regions and the tumor mass in each region are important, and an estimate of the total tumor burden has proved significant. B symptoms, histological subtype, age, and gender are also generally significant but less important. Prognostic factors for laparotomy findings in clinical stages I and II are: number of involved regions, disease confined to upper cervical nodes, B symptoms, gender, histology, age, and mediastinal disease (variable influence). In clinical stages I and II, the same prognostic factors apply as for pathological stages I and II and for laparotomy findings, and also some indirect indicators of extent of disease such as erythrocyte sedimentation rate, anemia, and serum albumin. In advanced disease the number of involved nodal and extranodal regions, the total tumor burden, B symptoms, age, gender, histology, and a number of hematologic and biochemical indicators are significant. Research into serum values of certain HD-associated antigens and cytokines may in the future provide valuable tumor markers in HD. PMID- 10717175 TI - Limited Radiation Therapy for Selected Patients With Pathological Stages IA and IIA Hodgkin's Disease. AB - The first definitive reports that early stages Hodgkin's disease (HD) could be cured by radiotherapy were published by Vera Peters in 1950 and by Easson and Russell in 1963. Since then, advances in understanding the natural history, refining the histopathologic classification, and development of a staging system have improved the management of HD. Several studies of early stage HD show greater than 80% actuarial 10- to 15-year freedom from relapse, and less than a 10% mortality from HD after treatment with radiation therapy alone. Radiotherapy has become so successful in the treatment of early stage HD (with combination chemotherapy reserved for relapse) that the risk of dying from other causes approaches the risk of dying from HD itself at 15 to 20 years following treatment. This article outlines the principles of treatment for favorable prognosis clinical stages IA and IIA HD, with emphasis on identifying patients with favorable features who may be treated effectively with mantle irradiation only. PMID- 10717176 TI - Radiation Therapy Techniques in the Treatment of Hodgkin's Disease. AB - Treatment of Hodgkin's disease with radiotherapy requires awareness of risk for subclinical disease and patterns of spread in assisting in the design of treatment volumes. This desire to treat larger volume needs to be balanced against concern for normal tissue toxicity. Traditionally, there have been "standard" field sizes for various disease presentations; however, the more recent trend has been to select patients in such a manner as to minimize long term toxicity while increasing or maintaining disease-free survival. This has most commonly involved elimination of laparotomy and decreasing field sizes and radiation doses in specific situations. This article discusses the appropriate dose and treatment for the treatment of Hodgkin's disease. PMID- 10717177 TI - Combined Modality Treatment for Poor Prognosis Stages I and II Hodgkin's Disease. AB - Recommendations for the treatment of "poor prognosis" stages I and II Hodgkin's disease (HD) depend on the extent of staging. For centers that favor exploratory laparotomy and splenectomy, the standard treatment after negative surgical staging remains subtotal lymphoid irradiation. However, most centers recommend excluding selected patients from surgical staging. In particular, most specialists agree that patients with bulky mediastinal disease (large mediastinal adenopathy) should receive combined chemotherapy and radiotherapy without surgical staging. Centers that have eliminated staging laparotomy rely on clinical staging and have identified a number of poor prognostic factors that influence treatment recommendations. In 1995, the following factors were considered to be associated with an unfavorable prognosis in clinical stages (CS) I and II HD: advanced age, systemic symptoms, high erythrocyte sedimentation rate, large number of individual sites, and presence of bulky disease. For example, these factors have been used in the H7 and H8 European Organization for the Research and Treatment of Cancer trials and have been adopted by almost 100 participant groups. For CS I and II patients with unfavorable presentations, the literature suggests that the optimal treatment is a combination of chemotherapy and radiotherapy. Nevertheless, the optimal delivery of chemotherapy and radiotherapy is still debated and a number of the questions listed below are awaiting answers: 1. The chemotherapy schedule: Mechlorethamine, vincristine, procarbazine, and prednisone (MOPP) alone can no longer be recommended (for toxicity reasons) for the routine treatment of poor prognosis CS I and II HD. Many centers, on theoretical arguments, favor seven or eight drug combinations (MOPP/ABV hybrid), although the advantage of these combinations in toxicity and efficacy over doxorubicin, bleomycin, vinblastine, and dacarbazine (ABVD) alone has not been demonstrated. 2. The number of chemotherapy courses: Most groups use six cycle regimens, but reduction to three to four cycles is currently being investigated both in pilot and randomized studies. 3. Timing of chemotherapy: Chemotherapy is given first in most instances. Whether or not a "sandwiched" scheme is better than all the chemotherapy given upfront remains to be answered. 4. Volumes to be irradiated: Data suggest that the irradiated volumes can be safely limited to initially involved lymph node regions after effective chemotherapy. The question is presently being addressed in a few ongoing randomized trials. 5. Radiation dose to be delivered: The decrease of the radiation dose to 35 to 36 Gy after effective chemotherapy has gained a wide acceptance. Further dose reduction is being investigated. In conclusion, except for selected CS I and II patients still referred for surgical staging, combined modality therapy appears to be the treatment of choice for poor prognosis CS I and II HD. PMID- 10717178 TI - The Role of Adjuvant Radiation Therapy for Stages III and IV Hodgkin's Disease. AB - The treatment for advanced stage Hodgkin's disease was revolutionized approximately 30 years ago with the introduction of combination chemotherapy. This approach has resulted in the cure of approximately 50% of patients with advanced disease. In an effort to improve these results, consolidation of radiotherapy (RT) was introduced approximated 25 years ago. The rationale for adjuvant RT comes from several observations: significant numbers of patients relapse following combination chemotherapy alone, these relapses occur predominantly at initially involved sites, and successful salvage following relapse can be quite difficult to achieve. Data from numerous phase II trials have demonstrated that adjuvant RT significantly reduces the relapse rate compared with what is seen following chemotherapy alone and appears to increase survival as well. Phase III trials have confirmed the reduction in relapse but, as yet, have failed to demonstrate survival benefits. Because definitive phase II trials are lacking, the use of consolidation radiotherapy has gained only partial acceptance and remains controversial. There have been numerous methodological difficulties with the execution of the reported phase III trials, most notably the failure of a substantial portion of patients randomly allocated to the radiotherapy group to actually receive radiation. This review presents updated results of studies using chemotherapy alone including patterns of failure analyses, summarizes the available literature pertaining to both retrospective and prospective trials of combined modality therapy, discusses toxicity issues, and attempts to provide guidelines regarding patient selection, sequencing of chemotherapy and radiotherapy, and radiation dose and field size. It is our conclusion that the available evidence supports the routine use of combined modality therapy for patients with advanced stage Hodgkin's disease. PMID- 10717179 TI - Management of Relapsed and Refractory Hodgkin's Disease. AB - Hodgkin's disease remains highly curable after relapse or even after an ineffective initial therapy. The likelihood of attaining a lasting second remission is particularly high for patients who failed to respond to a primary program of radiation therapy (RT) alone. The preferred treatment for these patients is with a doxorubicin-containing chemotherapy and additional RT (if feasible). The prospect for a "second cure" after failure to respond to initial chemotherapy is smaller and for most cases a program of high-dose therapy followed by stem-cell transplantation is required. Advances in using this modality over the last 15 years have resulted in a disease-free survival of approximately 40% with a follow-up that extends up to 10 years. Most important, a recent dramatic decrease in treatment-related mortality of high-dose therapy as well as a reduction in cost have been documented. Analysis of prognostic factors allows better selection of patients and a sounder choice of salvage options. Although it appears that further dose escalation of chemotherapy is limited by nonhematologic toxicity, the benefit from incorporation of RT into high-dose programs has only recently been recognized. Because of the availability of hematopoietic growth factors and easier mobilization and collection of peripheral blood stem cells, these have become the preferred source for hematopoietic support. The use of peripheral blood stem cells and growth factors was shown to correlate with more rapid recovery of granulocytes and platelets and a shorter hospitalization period. Still, many patients remain refractory to salvage despite intensive therapy. These patients can be identified by their response to re induction chemotherapy before transplant and should be considered for alternative approaches such as allogeneic bone marrow transplantation, sequential bone marrow transplant, and other experimental programs. This article reviews standard-dose salvage as well as current development in high-dose therapy with particular attention to the role of RT. PMID- 10717180 TI - Long-Term Complications of Treatment and Causes of Mortality After Hodgkin's Disease. AB - The majority of newly diagnosed patients are expected to survive Hodgkin's disease because of effective therapies established during past 30 years. Long term observations from large populations of treated patients have disclosed a variety of late effects of the disease and its therapy have contributed morbidity and excess mortality to Hodgkin's disease survivors. Secondary cancers have continued to accrue, and the risk relative to the general population has increased to 6.4 (95% confidence intervals: 5.5 to 7.3) in updated experience at Stanford University. Risks are significantly elevated for leukemia (primarily after chemotherapy regimens containing alkylating agents); non-Hodgkin's lymphoma; and tumors of the lung, breast, soft tissues, bone, stomach, pancreas, salivary gland, thyroid, and cutaneous melanoma. Early cardiovascular disease has also been observed and numerically exceeds second cancers as a cause of death in patients with early stage Hodgkin's disease (49 v 47 cases). Pulmonary dysfunction, thyroid dysfunction, infertility, psychosocial changes, gastrointestinal problems, soft-tissue changes, alterations in immunity, and risks for infection have also affected some treated patients. As these problems have been recognized, treatment approaches have been modified over the last 10 to 15 years, and early data suggest a decrease in some treatment sequellae. PMID- 10717181 TI - Introduction. PMID- 10717182 TI - Clinical Applications of Molecular Biology in Radiation Oncology. AB - As oncologists, our ability to understand and treat cancer will change dramatically over the next few decades. The fields of molecular, cellular, and cancer biology are rapidly changing, bringing forth new concepts regarding the basic mechanisms of oncogenesis, and with them, totally novel opportunities for therapeutic intervention. A general knowledge of the terms and concepts of the new biology is essential to radiation oncologists because these are already in our daily practice in the form of questions from our patients regarding the newest findings, assays of prognostic and predictive factors, and potential new therapies to deliver or augment radiation therapy. The classic radiation biology observations remain true-cell cycle perturbation, hypoxia, variation in cell survival after radiation, the competition model, fractionation effects, normal tissue injury, and so forth. The techniques available from the new biology enable us to elucidate the basic mechanisms underlying these observations. The current classes of radiation modifiers-halopyrimidines, hypoxic modifiers, radioprotectors, and combined modality therapy-will be supplemented by modifiers of signal transduction, cell cycle arrest, apoptosis, angiongenesis, and the stress response, to name a few. PMID- 10717183 TI - Radiation-Mediated Gene Expression in the Pathogenesis of the Clinical Radiation Response. AB - Cells and tissues respond to reactive oxygen intermediates during physiological processes such as inflammation, ischemia, and reperfusion. Ionizing radiation mimics naturally occurring free radicals to produce similar responses. Radiation induces the production of reactive oxygen intermediates and subsequent oxidation of cell membrane, phospholipids, and DNA. This initial event leads to activation of signals (enzymes) within the cells. These activated enzymes produce a cascade of energy transfer within the cell, leading to activation of transcription factors and the transcriptional apparatus. Radiation-inducible genes are then transcribed, leading in part to biological responses to ionizing radiation. Examples of radiation-inducible genes that regulate the biological response include inflammatory mediators and cell cycle regulators. The products of radiation-inducible genes are proteins such as transcription factors, cell adhesion molecules, and cytokines. The importance of identifying the mechanism of radiation-mediated gene expression is that enzyme inhibitors can be used to prevent gene expression. This synopsis of radiation-induced gene expression reviews genes that are induced by x-rays and gamma-rays and categorizes them according to their related biological responses. PMID- 10717184 TI - Signal Transduction and Cellular Responses to Ionizing Radiation. AB - The current understanding of cellular responses to ionizing radiation fuses traditional radiobiological concepts with recent insight into underlying molecular mechanisms. Cell cycle arrest, induction of specific cascades of genes, and activation of DNA repair mechanisms have been reported after radiation exposure. Of particular importance is the mitogen-activated protein (MAP) kinase signaling pathway, in which the ras and raf oncogenes participate. Both of these genes have been implicated in the cellular resistance to killing by ionizing radiation. Furthermore, regulated NF-gammaB transcriptional activation may be important for the pleiotrophic responses of cells to ionizing radiation. Disruption of this signaling cascade may contribute to the radiation sensitivity syndrome seen in ataxia telangiectasia. In this review, these pathways are consolidated into ionizing radiation signal transduction pathways that lead from the cell membrane/cytosol to the nuclear DNA. PMID- 10717185 TI - Modulation of the Apoptotic Response: Potential for Improving the Outcome in Clinical Radiotherapy. AB - The most prevalent mechanism of cell kill by radiation is mitosis dependent and results from lethal DNA double-strand breaks and failure to maintain normal replication. Apoptosis is believed to represent a minor component of the clinical effects of radiation. Apoptosis is a preprogrammed death pathway that is constitutively expressed in many cells, albeit in an inactive form, regulated by antiapoptotic mechanisms. Data are presented to show that in irradiated cells the balance between proapoptotic and antiapoptotic signaling may determine the apoptotic outcome in vitro and in vivo. This balance can be modulated by pharmacological intervention to produce a more proapoptotic phenotype, increasing apoptotic cell kill by radiation. These studies establish the basic principles of signaling-based apoptosis therapy, designed to overcome the relative resistance to radiation-induced apoptosis and to improve the therapeutic ratio in the treatment of human tumors with fractionated radiation. PMID- 10717186 TI - Molecular Biology of the Cell Cycle: Potential for Therapeutic Applications in Radiation Oncology. AB - The cell cycle was first described by radiation biologists more than 40 years ago. Since then, radiation oncologists have used information regarding the cell cycle, in particular cell cycle kinetics, to design various treatment protocols; these have resulted in only modest improvements in patient outcome. Over the past 10 to 15 years there has been an explosion of scientific knowledge regarding the cell cycle, in particular the molecular regulation of the cell cycle checkpoints. In this review we will discuss the genetic events involved in regulating the G1 and G2 checkpoints and how this information may lead to future therapeutic breakthroughs. In addition we will discuss the potential clinical impact of the recent cloning of the gene for ataxia telangiectasia. PMID- 10717187 TI - Repair of DNA Damage Produced by Ionizing Radiation: A Minireview. AB - A knowledge of the biochemical and molecular basis of sensitivity to ionizing radiation can provide useful information regarding carcinogenesis, cancer susceptibility, and patient responses to radiotherapy. Various factors can influence radiosensitivity, including the induction, processing and manifestation of DNA damage. One factor known to strongly affect radiosensitivity is the ability of cells to repair damage to their DNA. Cells have evolved an elaborate array of enzymatic systems to maintain the integrity of their genetic material in the face of numerous agents that alter DNA structure or base sequence. Cellular repair of a macromolecule is known to occur only for DNA. In no other instance is the integrity of a single molecule so vital for the survival of a cell. A number of distinct enzymatic mechanisms are known that result in either the removal of damaged DNA bases or the rejoining of single- and double-strand breaks. In this article, we discuss some of the different types of DNA damage induced by ionizing radiation, including modified bases, sugar damage, strand breaks, and clustered DNA damage. This will be followed by a description of some of the basic mechanisms the cell uses to repair ionizing radiation-induced DNA damage, such as base excision repair. We will also discuss the relationship between DNA repair and other cellular processes, including transcription and the cell cycle. Recent advances, including the cloning and characterization of DNA repair genes implicated in human inherited disease, have provided new insights into the surprising complexity of cellular responses to DNA damage. Finally, the potential for using this information clinically will be discussed. The goal of this review is to inform the clinician not only about recent progress in the field of DNA repair, but also about relationships between the processing of DNA damage and other cellular functions. PMID- 10717188 TI - Cytokine-Induced Radiation Protection and Sensitization. AB - Cytokines, hormone-like proteins that are produced by stimulated cells and tissues, serve as intercellular messengers. The cloning and large-scale production in a recombinant form of an expanding number of cytokines in the past decade has permitted investigations aimed at assessing the benefit they may provide in preserving and restoring functions of tissues compromised by irradiation. This review focuses primarily on the preclinical and clinical findings of a number of radiation and chemotherapy studies in which cytokines were found to protect, restore, or at times harm the hematopoietic and gastrointestinal tissues, as well as lung and liver, against cytotoxic therapies. Included are the myelorestorative effects of cytokines, their application in mobilization of peripheral blood progenitor cells for bone marrow transplantation, and their myeloprotective effect when given before irradiation. Studies indicating the importance of the treatment schedule, and in some cases their contrasting effects on different tissues, are included. The insights gained from such studies into the mechanisms of regulation by cytokines of radiation induced damage are discussed. PMID- 10717189 TI - Modification of the Radiation Response by the Administration of Exogenous Genes. AB - Gene therapy combined with radiotherapy represents a new approach for cancer treatment to selectively radio-sensitize malignant cells or protect normal tissues. Examples of gene therapy strategies that modify the biological response to radiation include cytokine-mediated gene therapy and virally directed enzyme/pro-drug therapy (VDEPT). We have proposed the concept of gene therapy targeted by ionizing radiation, whereby a cDNA encoding a cytotoxin or an immunogenic protein that modifies the cellular radiation response is ligated downstream from a radiation inducible promoter. Protein is thus induced in response to irradiation. In this way, radiation acts as a molecular switch to activate transcription of a therapeutic gene. VDEPT incorporates a viral vector system to deliver a gene encoding a pro-drug activating enzyme to malignant cells. After intracellular expression of the enzyme, a nontoxic pro-drug is converted to an active compound to selectively sensitize tumor cells to radiation. We will review current concepts of gene therapy and the studies that show its potential efficacy in the modification of radiation response. PMID- 10717190 TI - Introduction. PMID- 10717191 TI - RAPID: An Electronic Medical Records System for Radiation Oncology. AB - A electronic text-based medical record system used in a radiation oncology department since 1990 is described. The database is composed of a group of subfiles linked with a powerful relational database. The clinically relevant subfile structures are described and the hardware and software requirements for the system are itemized. Data entry is accomplished primarily by transcriptionists with some additional input by physicians at the time a dictation is reviewed. The database has been the exclusive source of radiation oncology notes since 1990. The flow of information from the dictaphone to the printed copy is described. Compared with a standard transcription service, addition clinical information is collected at the time of the patient encounter, and this information can be abstracted in reports. While not entirely replacing the paper record, the easy and rapid access to text-based digital information from any computer within the department has been invaluable. PMID- 10717192 TI - Designing Radiotherapy Software Components and Systems That Will Work Together. AB - The radiation treatment planning and delivery process requires that many different computerized data sources, software tools, and computer control systems be smoothly integrated. These include computer-controlled accelerators and new digital imaging modalities. At the same time, the applications themselves are becoming increasingly complex. In radiotherapy, the variety of components to integrate is much greater than in other areas of clinical medicine, growing out of more than a 30-year history of use of highly stylized traditional computer applications and many rigid conventions and practices associated with these computer programs. The introduction of new tools and computer-controlled equipment influenced the evolution of radiotherapy planning software, leading us to take a new look at how radiotherapy planning is done. Addressing and solving these integration problems as a serious research undertaking is vital to continued success in deploying computer applications for clinical use. New software design ideas such as object oriented design, behavioral abstraction and mediators can solve these problems. Our experience shows that the time and effort to build high-quality adaptable modular systems can be kept modest and within the reach of a small development team. PMID- 10717193 TI - Design Specifications for a Radiation Oncology Picture Archival and Communication System. AB - The handling of various types of images is an important function in radiotherapy and can be aided significantly by the use of a picture archival and communication system (PACS). The conventional imaging PACS has been well-studies, but the radiotherapy picture archival and communication system (RT-PACS) is relatively unfamiliar to the medical community. While many of the massive data handling requirements of a conventional imaging PACS may not be necessary for an RT-PACS, the RT-PACS must support various image operations that are unique to radiotherapy. This report describes some of the desired functional capabilities of the RT-PACS and the design specifications required to support these capabilities. PMID- 10717194 TI - Integrating the Management of Patient Three-Dimensional Treatment Planning and Image Data. AB - The management of data for three-dimensional conformal radiation therapy (3D CRT) is a multifaceted enterprise that includes image data transfer and format conversion, the safeguarding and distribution of treatment planning data, and the construction of efficient and secure user environments. Because 3D radiation therapy is so closely tied to computational methods for processing and presenting images, attention to these issues is essential for developing practical systems for 3D CRT. PMID- 10717195 TI - Clinical Applications of Picture Archival and Communications Systems in Radiation Oncology. AB - Picture archival and communications systems (PACS) for radiation oncology present an entirely different set of constraints and requirements from systems developed for diagnostic imaging. PACS for radiation oncology aid in organizing the complex, interrelated functions of radiation oncology. Integration of PACS with clinical data management systems will provide the backbone for the comprehensive computer system that has long been sought in radiation oncology. Simulation, geometric and dosimetric treatment planning, field shaping, set-up, verification, and delivery are now all observable and/or controllable from computer systems that can be interfaced with the departmental PACS. Costs are substantially lower than with diagnostic PACS because the systems can be based on desktop computers and the image resolution requirements are not as stringent. Each PACS user will have more information more easily available than under current systems of organization. Vendor support of digital image communications (DICOM) protocols will enable full integration of equipment regardless of manufacturer. Potential increased in productivity will be realized if the systems for handling and evaluating images are fully automated and provide the users with analytic tools that enhance the utility of systems such as electronic portal imagers, multileaf collimators, and clinical data management systems. this report describes our efforts in producing such a system. PMID- 10717196 TI - A Critical Look at Currently Available Radiation Oncology Information Management Systems. AB - Many complicated technical factors are involved in selecting a radiation oncology information management system. This report presents a discussion of some important information systems components and concepts which should be considered when evaluating vendor options. Network operating systems, database management systems, network infrastructure, and client issues are all presented. The current marketplace of products is analyzed from both a features and a systems standpoint. Because selection of an appropriate system is a highly site-specific matter taking into account may important factors, general recommendations are difficult. Nonetheless, current market leadership by certain vendors and a broad sense of the future direction are indicated in the conclusions. PMID- 10717197 TI - Management of Information in Radiation Oncology: An Integrated System for Scheduling, Treatment, Billing, and Verification. AB - An effective information system is an essential prerequisite to delivering quality patient care at competitive costs. From scheduling and billing to complex treatment machine control and verification, the quality of the information system strongly affects the efficiency and accuracy with which patient care is delivered. The standard paper-based information system used in many clinics suffers form inefficiencies and incompleteness in scheduling and billing, no centralized database and the inability to generate routine reports and communicate with other information systems. Many of these problems are resolved by the introduction of an electronic information system. The implementation, gains, and limitations of two electronic information systems are discussed. While limitations such as the lack of complete seamless integration of all information still exist, major improvements have been made in efficiency, accuracy, data integrity, and reporting and billing completeness. PMID- 10717198 TI - Image Storage Requirements for Treatment Planning and Verification. AB - Digital imaging techniques are becoming increasingly accepted in radiation therapy, and they bring significant advantages when compared with film, especially the ease of storage and retrieval. A simple model is proposed for the estimation of digital data storage requirements in a typical department of radiation therapy. The model assumes that 100 patients are undergoing treatment at any given time, and their computed tomography (CT) images, treatment plans, digitized simulator films, and portal images are to be available at short notice ("on-line"). Off-line archival image storage is required after the completion of treatment, for 1,000 patients per year. Reasonable assumptions are postulated regarding image sizes and acquisition rates, and on-line storage requirements are estimated to be up to 300 megabytes (MB) for CT data, 170 MB for treatment plans, 200 MB for digitized simulator films, and 500 MB for digitized port films. If electronic portal imaging is used instead of port films, the on-line requirement becomes about 2 gigabytes (GB) for verification images, and 20 GB if portal movies are to be archived. Off-line storage requirements are a factor of ten larger. It is suggested that data compression may reduce these requirements by up to a factor of 20. PMID- 10717199 TI - Outcome Assessment in Radiation Oncology. AB - Improvements in medical practice most often are based on the analysis of our past performance. With the increased emphasis in the United States on managed care, the examination of our clinical performance has taken on increased importance. In addition to traditional tumor registries and clinical trials, we are being asked to substantiate the quality of our practice for purchasers such as third-party intermediaries and industry. Data collection can often be done on existing departmental and hospital information resources. Industry-wide standards such as HL7 and DICOM further support this effort. Selection of the best computer hardware, networks, and software is critical to effective clinic and patient management. PMID- 10717200 TI - Evaluating Commercially Available Three-Dimensional Radiotherapy Treatment Planning Systems. AB - Over the past year we have developed two similar questionnaires on three dimensional radiotherapy treatment planning (3DRTP) systems and used them to conduct a study of the function and features of commercially available 3DRTP systems. The issues which we reviewed to develop these questionnaires are important to all current and potential users of 3DRTP systems. The items identified in this article should aid readers in developing their own review questionnaire based on the items we discuss as well as their own clinical practice. Because it is not possible to develop a universal questionnaire that will satisfy every clinic because of site-specific treatment techniques and capabilities, this information should aid an institution contemplating the purchase of such a system to develop their own specific set of questions to aid them in their system selection. We believe that a high-level checklist of this nature will prove quite helpful to those institutions preparing to purchase such a system. PMID- 10717201 TI - Introduction. PMID- 10717202 TI - Patterns of Care Studies: Past, Present, and Future. AB - The Patterns of Care Study in Radiation Oncology (PCS) has existed for 25 years. This overview details the basic principles that have guided the study from its inception to the present and defines the future role for the PCS. PMID- 10717203 TI - National Averages for Process and Outcome in Radiation Oncology: Methodology of the Patterns of Care Study. AB - The purpose of this article is to describe the methodology used by the Patterns of Care Study to conduct surveys of the practice of radiation oncology that not only provide national averages for process and outcome measures but also allow comparisons among important subgroups. The article describes how the sample design assures that these analyses are possible and presents methods used for data collection and analysis. PMID- 10717204 TI - The Changing Structure of Radiation Oncology: Implications for the Era of Managed Care. AB - This report presents results from the facilities surveys that are useful for radiation oncology practices facing the challenges of managed care. Facilities surveys collect data from the entire census of facilities practicing megavoltage radiation therapy. Data include equipment, personnel, and patient load. The data presented show that most, but not all, facilities throughout the United States are adequately equipped in terms of highest energy treatment machine, type of treatment planning computer, simulation, and quality assurance programs. The data also present the variation in percentage of new cancer cases receiving radiation therapy and repeat patients as a percentage of new radiation therapy cases by census region. The data show trends in patient load per type of personnel for academic, hospital-based, and freestanding facilities in 1994 show that academic facilities are larger and treat more patients per treatment machine. Academic facilities used more therapists per machine than other facilities. PMID- 10717205 TI - 20 Years of Progress in Radiation Oncology: Prostate Cancer. AB - The Patterns of Care Study in Radiation Oncology has documented United States national averages for the evaluation and treatment of prostate cancer from 1973 to 1994. We report the characteristics of patient populations, treatment characteristics, and outcomes of care. PMID- 10717206 TI - 20 Years of Progress in Radiation Oncology: Prostate Cancer. AB - Carcinoma of the cervix remains of special interest to the Patterns of Care Study (PCS) because of the prominent role that radiotherapy plays in the definitive management of all stages and extents of disease. Five PCS surveys have been conducted for squamous cell cancer of the uterine cervix beginning in 1973 and repeated thereafter at 5-year intervals. The records of over 2,700 women have been reviewed for these surveys. Changes in the pretreatment investigations of cervical cancer patients have occurred during these years, with an increase in the use of computed tomography (CT) and a decrease in the use of intravenous pyelography and cystoscopy. A marked increase in the use of linear accelerators has also occurred, with 98% of all facilities having a linear accelerator as the highest energy treatment machine in the 1988 survey. There has been a change in brachytherapy prescription over the time of the surveys, with most institutions reporting point dose calculations and dose distributions instead of milligram hours. Mean point A dose has increased to approximately 80cGy, reflecting the PCS recommendations of dose intensity to the paracentral point through use of brachytherapy. The outcome for stage I and II cervical cancer has remained stable over the time of the surveys, while the results have improved for stage III disease. This improvement in survival and local control for stage III cervical cancer is due in part to the PCS, which has developed standards of treatment with the goal of radiation dose intensity. A number of patient, tumor, and treatment factors significant in multivariate analysis have been described by the PCS with large cohorts of patients. The most important treatment factors associated with a decrease in pelvic failure and improved survival are the use of brachytherapy and higher paracentral dose. A significant dose response has been described for stage III cervical cancer and optimal pelvic control has been demonstrated with point A doses above 8,500 cGy. Other treatment factors associated with improved outcome include the reduction of overall treatment time, particularly for stage III cervical cancer, and the adequacy of an intracavitary placement. The rate and time course of major complications associated with the radiation for cervix cancer has been described by the PCS. An increase in complications was seen for young patients and those with a history of abdominal surgery. Treatment factors that increased the risk of complications included a fraction size greater than 7,500 cGy, and the use of staging laparotomy via a transperitoneal approach. Through the findings of the PCS, national averages of the process and outcome of radiation for cervical cancer as well as the important patient, tumor, and treatment factors that affect outcome have been reported. These results have positively affected routine clinical practice. Greater emphasis is placed on the use of intracavitary radiation therapy (RT) and dose escalation, as well as on a reduction in overall treatment time for cervical cancer. Future research will focus on the use of new technologies, such as high-dose rate brachytherapy and the impact of CT- and magnetic resonance imaging-directed treatment planning on local tumor control and survival. It will specifically evaluate the radiation treatment and outcome of minority populations. It will also measure the adoption of the results of positive clinical trials and the findings from previous PCS analyses into the national practice of radiation oncology. PMID- 10717207 TI - Over 20 Years of Progress in Radiation Oncology: Hodgkin's Disease. AB - The Patterns of Care Study (PCS) surveys for Hodgkin's disease have documented important correlations between treatment processes and patient outcome. Nationwide improvements in radiotherapy for Hodgkin's disease since 1973, such as routine use of extended fields (subtotal lymphoid irradiation) in patients with early stage disease, individualized blocking, and treatment using linear accelerators, have resulted in greater freedom from relapse and overall survival for patients with stage I/II disease treated with radiotherapy alone. In addition, the introduction of computed tomography along with increased use of chemotherapy in high-risk patients has reduced the use of routine laparotomy and has improved outcome for patients with stage III disease. Overall survival for Hodgkin's disease in the national practice is excellent, reflecting the dissemination of complex treatment programs and radiation therapy technology to the oncologic community at large. Future studies of the national practice will be important in assessing the impact of managed care on workup and other facility practices, as well as evaluating the transfer of new approaches aimed at reducing treatment toxicities. PMID- 10717208 TI - Over 20 Years of Progress in Radiation Oncology: Seminoma. AB - During the past 20 years, significant changes have occurred in the management of seminoma. Survival has improved by approximately 10%, and now 97% of patients are cured. Reductions in the numbers of patients irradiated, the volumes irradiated, and the doses used should reduce morbidity. The 1973 Patterns of Care Study (PCS) and the planned new study proffer statements of consensus on optimal care and evaluate compliance with guidelines. Specific changes in investigation, including measurement of the serum tumor markers beta human choriaonic gonadotropin (betaHCG) and alphafetoprotein (AFP) and computed tomography (CT) or magnetic resonance imaging (MRI) evaluation of the retroperitoneum, better evaluate disease extent. For stage I disease, a reduction in the total dose of infradiaphragmatic irradiation to 2,500 cGy is recommended. An option for surveillance reduces unnecessary therapy in 80% and may improve fertility. The significance of disease bulk in stage II has been recognized, and treatment has been refined. The maximal radiation dose now recommended for stage II disease is 3,500 cGy. CT definition of radiation target volumes minimizes the risk of geographic miss. Prophylactic mediastinal irradiation is no longer recommended. Chemotherapy, usually now bleomycin, etoposide, and cisplatin, produces high cure rates for stage IID, III< and IV disease and has become the standard managemetn. Controversy still surrounds optimal therapy for stage IIC disease. Unresolved questions include cost benefit and quality of life issues surrounding optimal management for stage I disease, inguinal scrotal irradiation in stage I and II disease, and identification of the least toxic but effective chemotherapy for specific subgroups of patients with advanced disease. PMID- 10717209 TI - Quality Assessment in the USA: How the Patterns of Care Study Has Made a Difference. AB - Improving the quality and accessibility of radiation care in the United States has been the primary objective of the Patterns of Care Study (PCS) since its inception. While patient care has two components, technical and interpersonal, the PCS has only studied the quality of technical care. Such assessments of technical quality of radiation oncology, which are representative of the United States as a whole, virtually do not exist outside those of the PCS. The methodology used by the PCS to assess quality in radiation oncology is based on an examination of structure, process, and outcome. Structural elements identified by the PCS to be associated with inferior quality include the use of a Cobalt 60 unit with surface-to-skin distance (SSD) 50,000 and age>9 yrs. define eligibility for high-risk protocols. Infants (<1 yr. old) are high-risk, frequently associated with chromosomal t(4;11) translocation and extramedullary disease. T-cell immunophenotype is usually associated with clinical high-risk features, including older age, high WBC, and extramedullary disease. T-cell immunophenotype is usually associated with clinical high-risk features, including older age, high WBC, and extramedullary disease. Current regimens include induction chemotherapy, consolidation (often including high-dose systemic methotrexate, MTX), and continuation phases; intensification or reinduction is incorporated for high-risk disease. Preventive CNS therapy is a critical component of therapy. Intrathecal (IT) chemotherapy (MTX +/- cytosine arabinoside and hydrocortisone) and systemic agents provide adequate CNS therapy in standard risk patients. The "threshold" for using preventive cranial irradiation (Crl, 18 Gy) varies among protocols, but systematically includes those with T-cell ALL and WBS>50,000; other criteria may include B-progenitor ALL with WBC>100,000,philadelphia chromosome positive ALL, residual marrow disease at day 7, or male gender, For the 5% of children with CNS leukemia at diagnosis (defined as CSF with 5 or greater WBC/uL and positive cytology), Crl is administered at does of 18 to 24 Gy. Recent series show a CNS relapse rate approximating 5% Reinduction chemotherapy, IT therapy, and subsequent craniospinal irradiation ( typically 24, Gy Crl, 15 Gy spine) achieve durable secondary disease control in greater than 60% of cases with isolated CNS relapse. High-dose systemic MTX has virtually eliminated testicular relapse. CNS toxicities (including leukoencephalopathy) relate to systemic and IT MTX as well as Crl. Late neuropsychologicral data following Crl at 18 Gy show little intellectual deficit compared with children receiving MTX- based preventive therapy alone. Neuroendocrine dysfunction is apparent in long-term survivors following Crl at 24 Gy. The relative efficacy and toxicities of Crl versus more intensive MTX-based regimens are yet controversial. Overall disease-free survival in ALL approximates 70% in contemporary series. PMID- 10717214 TI - The Changing Role of Radiation Therapy in the Treatment of Neuroblastoma. AB - Neuroblastoma (NBL) is the fourth most common pediatric malignancy. With a median age at diagnosis of 2 years, it represents half of all malignancies that present in the first month of life and one third of those that are diagnosed in the first year of life. NBL is unique among human cancers in its ability to undergo spontaneous differentiation and permanent tumor regression. This phenomenon is particularly characteristic of disseminated disease with liver, skin, and limited bone marrow involvement and involving a limited primary and presenting in children under 1 year of age. Advances in the understanding of the biologic behavior of NBL coupled with the clinical presentation have led to a risk-based approach to treatment, minimizing the treatment to some patients and supporting the need for more aggressive treatment to others. A new international staging system has been adopted that has allowed better comparisons among treatment reports from varying centers and cooperative groups. This article reviews the risk-related approach to the treatment of NBL and the changing role of radiation therapy. PMID- 10717215 TI - Wilms' Tumor: Changing Tole of Radiation Therapy. AB - Wilms' tumor is a highly curable neoplasm. Greater that 90% of all children with this disease can be expected to become long-term survivors. Although radiation therapy (RT) was once the mainstay of nonsurgical treatment, its use has been reduced both in indications and in dosage because of the chemoresponsiveness of the tumor. In the Third National Wilms' Tumor Study (NWTS 3), patients with stage II tumors were shown not to require postoperative RT, and in patients with stage III tumors, 10 Gy was sufficient. In NWTS 5, patients with stage III favorable histology (FH), stage IV FH (with abdominal stage III), and stage II-IV anaplastic and all patients with clear cell sarcoma receive 10 Gy to to the abdomen (usually given as 1.8 Gy x 6-total doe 10.8Gy). Results from the International Society of Paediatric Oncology, in which downstaged patients had a higher incidence of abdominal relapse, and the United Kingdom Children's Cancer Study Group first Wilms' Tumor Study, in which omission of whole-lung RT led to lowered survival in stage IV patients, suggest caution in further modifications of RT at this time. PMID- 10717216 TI - Rhabdomyosarcoma of Parameningeal Sites. AB - Rhabdomyosarcoma arising in the skull base has a feature of its natural history that is unique in oncology: by virtue of parameningeal location and invasive behavior, it can extend intracranially and produce neoplastic meningitis. The four anatomic sites with this potential are the nasopharynx/nasal cavity, the middle ear, the paranasal sinuses, and the infratemporal fossa/pterygopalatine space. Patients with these primary sites comprise 41% of patients with head and neck sites and 15% of all patients with rhabdomyosarcoma. Most are under 10 years old at diagnosis (72%) and are Intergroup Rhabdomyosarcoma Study (IRS) Clinical Group III (76%). IRS protocols over the last 20 years have lead to a gradual improvement in outcome while refining radiation treatment parameters to allow for restricted indications for (and then the elimination of) whole cranial radiotherapy ports. The current guidelines (Group III) are for a dosage of 1.8 Gy/d up to a total of 50.4 Gy using 2-cm margins around the gross tumor volume at diagnosis. The timing of radiotherapy in relation to combination chemotherapy is based on assessment of three risk factors that predict tumor access to the cranial subarachnoid space: skull base erosion, cranial nerve palsy, and intracranial extension. Patients with one or more factors begin radiotherapy concurrently with chemotherapy. The remaining one third of patients begin radiotherapy after induction chemotherapy (week 9). The 5-year failure-free survival rate for patients with Group III parameningeal rhabdomyosarcoma on IRS III was 71%. In the absence of the three risk factors, 5-year survival was 97%. The local failure rate was 15%. Among patients who achieved a compete or partial response (IRS-III), 5% had contiguous ((nonhematogenous) central nervous system failure. Late effects of radiotherapy in these young patients are protean and include growth inhibition, cataracts, impaired dentition and hearing, and facial bone asymmetry. The promise of improvement of the therapeutic ratio may be realized from treatment refinements, which include chemotherapy response-based portal reduction, hyperfactionation, and conformal radiation therapy. PMID- 10717217 TI - Genitourinary Rhabdomyosarcoma. AB - Rhabdomyosarcoma (RMS) is a malignant tumor with a variety of clinical presentations requiring varied therapeutic approaches. Radiotherapy plays an important role in the treatment of many children with RMS arising in genitourinary (GU) sites. Although the majority of these children present with locally advanced disease, the prognosis is relatively favorable when multimodility treatment strategies are used, with overall survival approaching 70% to 80%. Aggressive surgical procedures for local control are being replaced by bladdersparing treatments involving more intensive chemotherapy and the early use of radiotherapy, resulting in perservation of bladder function in as many as two thirds of children. Good radiotherapy technique is critical for ensuring not only the inclusion of adequate tissue volumes for treatment of tumor, but also for sparing normal, uninvolved structures. Optimal treatment factors such as dose, volume and timing of radiotherapy are still being investigated. This article discusses strategies currently being used for the management of GU RMS as well as research that is underway to answer unresolved questions about the biology and therapy of this disease. PMID- 10717218 TI - The Role of Radiation Therapy in Localized Ewing' Sarcoma. AB - Ewing's sarcoma is a radiosensitive tumor that has historically been treated primarily with radiation therapy (RT). With the introduction of effective systemic therapy in the early 1970s, RT remained the primary local control modality until recently. With the concern about second tumors in the irradiated field and improved surgical techniques, surgical resection is currently playing a more important role in the management of these tumors. There have been no randomized trials comparing radiation with surgery, and in most series, patients with smaller tumors in accessible sites are treated with surgical resection. Retrospective reviews typically show a greater advantage to the use of surgery than do prospective trials. Although there may be slight local control advantage to surgery, the survival advantage is not great. Since the mid-1980s, fields tailored to the tumor volume have been recommended and the use of whole-bone irradiation has been abandoned. Local control has not been adversely effected. The intergroup trials for Ewing's sarcoma have recommended 45 Gy to a larger volume followed by a 10 Gy boost since 1978. Data from other studies suggest that the dose may be decreased in selected situations. Older studies of functional outcome report significant late effects related to radiation, but these seem to be reduced in more recent series. PMID- 10717219 TI - Conformal Therapy for Pediatric Sarcomas. AB - For decades, pediatric oncology has lead the field in the implementation of combined modality therapy and has tested indications for radiation therapy in this patient population. More recently, adult experiences with conformal methods of external beam irradiation and brachytherapy have refined radiation therapy as modality and increased its attractiveness as a means to achieve local control. These refinements come at a time when the current rates of local failure for advanced stage patients with pediatric solid tumors are significant enough to warrant a reexamination of the accepted indications for radiation therapy and its sequencing and dose. Conformal methods of radiation therapy, including brachytherapy, will facilitate efforts to improve local control. These methods have a special role in the pediatric patient where the potential for dose escalation, function preservation, and decreased toxicity can be used to the benefit of these patients, making the majority of them likely to be long-term survivors. PMID- 10717221 TI - Introduction. PMID- 10717222 TI - Mechanisms of Action and Modulation of Fluorouracil. AB - Fluorouracil (5-FU) and 5-fluoro-2'-deoxyuridine (FdUrd) are commercially available fluorinated pyrimidine analogues. 5-FU has antitumor activity against adenocarcinomas arising in the breast, gastrointestinal tract, and ovary, and against squamous cell carcinomas arising in the head, neck, and esophagus, with single-agent response rates of 10% to 30%. FdUrd has mainly been used for hepatic arterial infusions for patients with isolated hepatic metastases, with response rates of 42% to 62% as first-line therapy for colorectal cancer patients and 30% in those failing prior systemic 5-FU-based therapy. An appreciation for the factors influencing the cellular pharmacology of 5-FU has generated interest in combining it with both modulatory agents that enhance its metabolism or cytotoxic effects and other antineoplastic agents or modalities, such as cisplatin, methotrexate, and ionizing radiation, with which it can produce synergistic cytotoxicity. The preclinical and clinical pharmacology of 5-FU is reviewed, and novel approaches to permit oral adminstration as a means of mimicking protracted infusional schedules are discussed. PMID- 10717223 TI - Fluoropyrimidine-Radiation Interactions in Cells and Tumors. AB - The fluoropyrimidines (fluorodeoxyuridine [FdUrd] and fluorouracil [5-FU] are potent radiation sensitizers. This article summarizes recent data concerning potential mechanisms of fluoropyrimidine-radiation interactions at the cellular and tumoral level. Evidence now suggests cells can be radiosensitized when exposed to the fluoropyrimidines either before or after radiation but by different mechanisms. Sensitization is not explained by redistribution of cells into a sensitive phase of the cell cycle. Rather, cells that are exposed to FdUrd before radiation progress in S phase in the presence of fluoropyrimidines and are radiosensitized. Radiation can also increase the anabolism of 5-FU into the toxic metabolite and decrease drug clearance from the tumor. These laboratory data may aid in the design of clinical trials combining radiation and fluoropyrimidines. PMID- 10717224 TI - Irradiation Plus 5-Fluorouracil: Cellular Mechanisms of Action and Treatment Schedules. AB - The use of external irradiation and 5-fluorouracil continues because of the proven benefits of this combination in the treatment of a variety of human malignancies. There are probably several mechanisms of action on the cellular level that account for radiation sensitization. One main feature of both laboratory and applied clinical research in this area is the apparent superiority of prolonged exposure of this S-phase cytotoxic agent when administered with fractionated irradiation. Also reviewed here are schedules using protracted infusional chemoradiation with this agent that are being tested in ongoing clinical trials. PMID- 10717225 TI - Radiation Therapy With 5-fluorouracil in Head and Neck Cancer. AB - 5-Fluorouracil (5-FU) alone or combined with other drugs, most frequently cisplatin, has been used concurrently or as induction or adjuvant therapy with radiotherapy with or without surgery in the treatment of head and neck cancer. Improved local-regional control and disease-free survival or overall survival have been shown in several randomized trials using a concurrent approach. However, acute mucositis is usually increased with simultaneous 5-FU and radiation administration, especially when other drugs are used in addition to 5 FU. Alternating radiotherapy with 5-FU and cisplatin was shown to improve the local-regional relapse-free, progression-free, and overall survival of unresectable squamous cell carcinoma of the head and neck compared with radiotherapy alone in one randomized trial. Further evaluation of the alternating chemotherapy and radiotherapy approach is needed, however, before one can accept this as a standard of practice. Induction chemotherapy with 5-FU infusion and cisplatin followed by definitive radiotherapy in the chemotherapy responders in an alternative treatment option for patients with locally advanced resectable squamous cell carcinoma of the larynx or hypopharynx who wish to preserve organ function. Induction or adjuvant chemotherapy with 5-FU infusion and cisplatin may also decrease or delay the occurrence of distant metastasis. Induction chemotherapy, however, has not been shown to improve local-regional control or overall survival. Further clinical trials combining 5-FU and its biochemical modulators using innovative radiation and drug dose schedules and other treatment modifiers are needed to improve the therapeutic ratio. PMID- 10717226 TI - Combination of 5-Fluorouracil and Radiation in Esophageal Cancer. AB - A review of the trends in the clinical practice of esophageal cancer treatment is presented. The preponderance of evidence indicates that concomitant chemotherapy and radiation is superior to either modality above. Chemotherapy usually, but not invariably, consists of 5-fluorouracil (5-FU) and cisplatin; radiotherapy is usually 50 Gy. Attempts to escalate to higher dose by external-beam boost or brachytherapy is still experimental. Combination treatment with surgery is also successful, but the inclusion of esophagectomy in the treatment is not universally accepted and is unlikely to be tested. A suggestion to base treatment selection on response is proposed. PMID- 10717227 TI - Radiation Therapy and 5-Fluorouracil in Pancreatic Cancer. AB - Combination chemotherapy has not been shown to be superior to single-agent 5 fluorouracil (5-FU) in the treatment of metastatic pancreatic cancer. Initial studies from Duke University suggest a role for radiation in the management of locally unresectable presentations of this disease. In vivo studies showed that 5 FU could act as a radiation sensitizer. This concept was subsequently tested in a randomized trial by the Gastrointestinal Tumor Study Group (GITSG). In their studies, combined modality therapy using radiation and 5-FU-based chemotherapy prolonged survival in both adjuvant and locally unresectable settings and remains as a standard therapeutic option for appropriately selected patients. Variations on this approach designed to potentiate 5-FU efficacy are under active clinical investigation. This review discussed past and current studies involving 5-FU as well as other agents currently of interest. PMID- 10717228 TI - The Role of Radiation Therapy and 5-Fluorouracil in Colon and Rectal Cancer. AB - Since the original report improved survival of patients with locally unresectable gastric cancer treated with irradiation and 5-fluorouacil (5-FU) in 1968, there has been extensive investigation of this treatment combination in a variety of gastrointestinal tumors, including colon and rectal cancer. There are now extensive data supporting the use of 5-FU-based chemotherapy with irradiation in the adjuvant treatment of rectal cancer and increasing experience defining its role in negotiant therapy. The value of adjuvant 5-FU-based chemotherapy and irradiation in colonies cancer is controversial, but there may be subsets of patients who benefit from this treatment. Ongoing studies in rectal and colon cancer are evaluating the role of irradiation with various modes of 5-FU adminstration (bolus v short or continuous infusion) as well as its modulation with other agents. PMID- 10717229 TI - The Role of Radiation Therapy With 5-Fluorouracil in Anal Cancer. AB - The most commonly used initial treatment of primary epidermoid cancer of the anal canal is radiation combined with concurrent 5-fluorouracil (5-FU) and mitomycin. Randomized trials have shown that these drugs, when combined with split-course or moderate-dose radiation, are superior to the same doses of radiation delivered without drugs. In another trial, the addition of mitomycin to 5-FU resulted in a better outcome thand when 5-FU alone was combined with radiation. Studies are in progress to evaluate treatment with radiation plus 5-FU and cisplatin; this combination has also produced high rates of tumor response in preliminary studies. The optimal schedules and doses of 5-FU to combine with radiation are not known-common usage favors 96- or 120-hour infustions of 5-FU at a dose of 750 to 1,000 mg/m(2)/24 hours, generally administered as one or two courses concurrently with conventional once-daily fractionated radiation. It is unclear whether 5-FU in these combinations is acting as a cytotoxic agent, a radiosensitizer, or both. Despite these uncertainties, empiric clinical studies have led to the development of effective treatment regimens that allow conservation of anorectal function in the majority of patients with anal cancer. PMID- 10717230 TI - Clinical Results of the Combination of Radiation and Fluoropyrimidines in the Treatment of Intrahepatic Cancer. AB - Radiation therapy has played a minor role in the conventional management of patients with unresectable primary hepatobiliary malignancies and liver metastases. This can be attributed to normal tissue toxicity, which has limited the dose of radiation that can be safely administered, and imaging limitations, which have prevented focal irradiation of involved regions. Trials of whole-liver irradiation with and without 5-fluorouracil (5-FU) or fluorodeoxyuridine (fur) suggest that the addition of fluoropyrimidines results in a slight increase in the median survival without an increase in radiation-induced liver disease (RILD). The development of three-dimensional radiotherapy techniques has now allowed high-dose focal irradiation to be safely delivered, which, when combined with hepatic arterial fur, produces a much higher rate of response and, in some cases, prolonged survival particularly for patients with primary hepatobiliary cancers. Further investigations into the optimal methods of combining the fluoropyrimidines and radiation to permit higher doses of both agents to be delivered as well as methods of protecting the normal liver are expected to yield significant improvement in the outcome of treatment for patients with intrahepatic malignancies. PMID- 10717231 TI - An Approach to Radiosensitizing Cervical Cancer by Use of Chemical Modulators of Nucleoside Metabolism. AB - Although radiation alone is the treatment of choice for patients with cervical cancer that is not surgically respectable, locoregional failures rates approach 50% for locally advanced stages of disease. Therefore, decades of clinical trials using chemoradiotherapy have been performed in an attempt to enhance cure rates. Unfortunately, the addition of chemotherapy has not been shown to unequivocally improve outcome compared with radiation alone. Reasons for this include inadequate radiation doses, radiation treatment delays caused by higher acute toxicities of combined modality therapy, and insufficient understanding of both the optimal sequencing and mechanisms of radiosensitizers. Some of the chemotherapy agents tested include the fluoropyrmidines (5-fluorouracil [5-FU]), the halogenated thymidine analogues (iododeoyxuridine [IdUrd] and bromodeoyxuridine [BrdUrd] and hydroxyurea (HU). This article focuses on clinical results using these compounds, the evolving understanding of these different types of drug-radiation interactions, and potential new strategies for the use of these radiosensitizers in patients with locally advanced cervical cancer. PMID- 10717232 TI - The c-Jun N-terminal kinase pathway and apoptotic signaling (review). AB - The c-Jun N-terminal kinase (JNK) group of mitogen-activated protein kinases (MAPKs) is activated in mammalian cells by environmental stress, pro-inflammatory cytokines, and mitogenic stimuli. Biochemical and genetic studies demonstrate that JNK regulates the activities of many transcription factors, and that the JNK pathway is required for the regulation of inflammatory responses, cell proliferation, and apoptosis. The involvement of JNK in apoptotic cell death is particularly intriguing, and has been actively studied in recent years. An improved understanding of JNK-mediated apoptotic signaling may provide novel strategies in prevention and treatment of cancers. PMID- 10717233 TI - GSTP1 CpG island DNA hypermethylation in hepatocellular carcinomas. AB - Glutathione S-transferases, enzymes that defend cells against damage mediated by oxidant and electrophilic carcinogens, may be critical determinants of cancer pathogenesis. We report here that the pathogenesis of hepatocellular carcinoma (HCC), one of the most common cancers in the world, frequently involves an accumulation of somatic DNA methylation changes at GSTP1, the gene encoding the pi-class glutathione S-transferase. For our study, Hep3B HCC cells and a cohort of 20 HCC tissue specimens were subjected to analysis for GSTP1 expression and for somatic GSTP1 alterations. GSTP1 DNA hypermethylation in HCC DNA was assessed by Southern blot analysis, via a polymerase chain reaction (PCR) assay, and by using a genomic sequencing approach. Hep3B HCC cells failed to express GSTP1 mRNA or GSTP1 polypeptides. Similarly, HCC cells in 19 of 20 HCC cases were devoid of GSTP1 polypeptides. By Southern blot analysis, DNA from Hep3B HCC cells displayed abnormal GSTP1 hypermethylation. Treatment of Hep3B HCC cells in vitro with the DNA methyltransferase inhibitor 5-aza-deoxycytidine both reversed GSTP1 DNA hypermethylation and restored GSTP1 expression. Using a PCR assay, somatic GSTP1 DNA hypermethylation was also detected in HCC DNA from 17 of 20 HCC cases. Genomic sequencing analyses, undertaken to map 5 methyldeoxycytidine nucleotides located at the GSTP1 transcriptional regulatory region, frequently detected somatic DNA hypermethylation near the gene promoter in HCC DNA. The data indicate that GSTP1 DNA hypermethylation changes appear frequently in human HCC. In addition, the data raise the possibility that somatic GSTP1 inactivation, via hypermethylation, may contribute to the pathogenesis of HCC. PMID- 10717234 TI - Stromal CEA immunoreactivity is correlated with lymphatic invasion of human esophageal carcinoma. AB - Carcinoembryonic antigen (CEA) is a good marker of colorectal cancer. Recent studies have demonstrated that CEA may function as a metastatic potentiator by different pathways; i.e. modulation of immune responses, facilitation of intercellular adhesion and cellular migration. However, expression patterns of CEA have not yet been established in human esophageal carcinomas. In this study, we examined CEA expression in human esophageal squamous cell carcinoma and its clinicopathological significance. CEA immunoreactivity was frequently detected in the cancer cells (cytoplasmic type; 81.1%, 43/53) as well as in the cancer stroma (stromal type; 32.1%, 17/53), regardless of the depth of tumor invasion. Lymphatic invasion of cancer cells was frequently found in the stromal CEA positive esophageal cancer (44.4%, 16/36), compared to stromal CEA-negative cancer (5.9%, 1/17) (p<0.05). These observations suggested that stromal CEA expression plays important roles in lymphatic invasion of human esophageal squamous cell carcinoma. PMID- 10717235 TI - Translocation and coamplification of loci from chromosome arms 8p and 11q in the MDA-MB-175 mammary carcinoma cell line. AB - Rearrangement and coamplification of the 8p12 and 11q13 chromosomal regions occurs in a significant proportion of breast cancers. It usually involves a complex hybrid structure in which the FGFR1 and CCND1 genes are amplified. We report here a different type of 8p12-11q13 rearrangement in the MDA-MB-175 mammary carcinoma cell line. This amplification contains the NRG1/HGL (from 8p12 21) and DOC4 (from 11q13) genes, encoding respectively a ligand for ERBB receptors and a stress-induced protein which is a mammalian ortholog of Drosophila Tenm/Odz. It has been shown previously (Wang et al, Oncogene 18: 5718 5721, 1999) that these two genes are rearranged and fused by a translocation event. This type of event was not found in 30 tumors tested that showed coamplification of the 8p12 and 11q13 regions. PMID- 10717236 TI - Glial cell line-derived neutrotrophic factor and neurturin can act as paracrine growth factors stimulating DNA synthesis of Ret-expressing spermatogonia. AB - Glial cell line-derived neurotrophic factor (GDNF) and neurturin (NTN) are related growth factors which exert trophic effects on several neuronal populations and developing kidney. GDNF-family ligands interact with membrane receptors designated GFRalphas which, in turn, mediate stimulation of the Ret receptor tyrosine kinase. Here we show that Ret, GFRalpha-1 (the GDNF receptor), and GFRalpha-2 (the NTN receptor) are expressed by testicular germ cells, while GDNF and NTN are expressed by Sertoli cells. Both GDNF and NTN stimulate DNA synthesis in spermatogonia. Furthermore, Ret, ligands and co-receptors are expressed in germ cell tumors. Thus, GDNF-family ligands may act as paracrine factors in spermatogenesis and this circuit may be active in germ cell tumors. PMID- 10717237 TI - Enhanced apoptosis in retrovirally transfected osteosarcoma cells after exposure to sodium butyrate. AB - We investigated the effects of sodium butyrate (NaBu) on two retrovirally transfected osteosarcoma cell lines (1547-TK and 1547-LacZ cells) compared to the corresponding untransfected cell line. The first finding was an inhibitory effect only on the proliferation of both transfected cell lines. This antiproliferative effect was associated with apoptosis induction, which was detected using techniques that monitor either characteristic biochemical or morphological processes. Our findings show that 1547-TK and 1547-LacZ cells were much more sensitive to NaBu treatment than untransfected 1547 cells as concerns both proliferation and apoptosis induction. PMID- 10717238 TI - Expression of the OGG1-type 1a (nuclear form) protein in cancerous and non cancerous human cells. AB - The human OGG1 gene encodes 8-hydroxyguanine DNA glycosylase. By RT-PCR analysis, five novel type 1 transcripts, in addition to eight known types (OGG1-types 1a to 1c and 2a to 2e), were identified. Among them, only the type 1a isoform contains both a nuclear localization signal and the entire DNA binding motif, suggesting the involvement of type 1a in chromosomal DNA repair. By Western blot analysis using a monoclonal antibody prepared by immunizing the whole type 1a protein, a 39 kDa type 1a protein was detected in lung cancer cell lines and peripheral lymphocytes. The type 1a protein was expressed at a similar level, irrespective of its polymorphic types characterized by distinct repair activity. By an immunocytochemical study, the majority of type 1a protein was localized in the nucleus. These results indicate that OGG1-type 1a protein is involved in the repair of 8-hydroxyguanine in chromosomal double-stranded DNA and constitutively expressed in cancerous and non-cancerous human cells. PMID- 10717240 TI - alpha6 integrin expression in esophageal carcinoma. AB - One of the genes that the authors have isolated by a differential display of hepatocellular carcinoma compared to adjacent liver is the alpha6 integrin. alpha6 integrin is the major adhesion receptor for laminin and is suggested to be involved in tumor cell invasion and metastasis. To our knowledge, however, there are no reports on alpha6 integrin expression in esophageal carcinoma. We thus conducted a study to determine its clinicopathologic significance in human esophageal carcinoma. The tumor/normal (T/N) ratio of alpha6 integrin expression was calculated by Northern hybridization in 45 cases of esophageal carcinoma. In selected cases the expression of the alpha6 integrin variants A and B was also investigated by reverse transcriptase-polymerase chain reaction, and immunostaining for alpha6 integrin was performed. The expression levels of alpha6 integrin mRNA in the tumor tissue were greater than those of the corresponding normal tissue in 39 of 45 cases (87%). The overexpression of alpha6 integrin was also recognized by immunostaining. Fifteen cases with a high T/N ratio demonstrated a deeper invasion into the esophageal wall than the 30 cases with a low T/N ratio. Although there was no significant difference, the 15 cases with a high T/N ratio had a tendency for a worse prognosis. The ratio of the two variants (alpha6A/alpha6B) did not show any relationship to survival. The findings imply that alpha6 integrin is overexpressed in human esophageal carcinomas and that alpha6 integrin may play an important role in esophageal tumor invasion. PMID- 10717239 TI - Enhancement of the transformed shape phenotype by microtubule inhibitors and reversal by an inhibitor combination. AB - Differences between transformed cells and their normal counterparts were defined by analyzing the cells' three-dimensional distribution of mass. Variables called factors, which explained the covariance of the real variables, were extracted from the data. We found that factors #4 (sharp, tapering projections) and #12 (rounding up), corresponded to G-protein functions. Then, the signature-type mass distribution of transformed cells was defined by factor values. Agents that caused signature-type mimicry could quantitatively shift factor values, for example, those affecting endocytic processing disproportionately reduced values of #4. Signature-type reversal was also observed and may be valuable in predicting the efficacy of chemotherapeutic agents. PMID- 10717241 TI - Analysis of the candidate target genes for mutation in microsatellite instability positive cancers of the colorectum, stomach, and endometrium. AB - Microsatellite instability (MSI) in human carcinoma DNA is a characteristic phenotype observed in hereditary non-polyposis colorectal cancer and also in some human sporadic cancers including multiple primary carcinomas. In this study, we analyzed mutations in the hCHK1, E2F4, hMSH3, and hMSH6 genes in MSI+ human cancers arising in colorectum, stomach and endometrium. The E2F4 and hMSH3 genes were mutated in all tumor types. Interestingly, the hMSH6 gene was mutated in colorectal and gastric cancers but not in endometrial cancer; this is similar to the TGFbetaRII gene. It is notable that the mutation status of the secondary mutators, hMSH3 and hMSH6, did not influence slippage-related frameshift mutations in genes harboring simple tandem-repeats, which suggests that the MSI phenotype may be affected mainly by abnormalities in the primary mutator genes, not by the secondary mutator genes. No mutations were observed in the cell cycle checkpoint gene hCHK1; mutations of this gene are thought to have a limited role, if any, in at least the tumor types analyzed in this study. PMID- 10717242 TI - The effect of p53-function on the sensitivity to paclitaxel with or without hyperthermia in human colorectal carcinoma cells. AB - The importance of p53-function for the sensitivity to paclitaxel with and without hyperthermia (HT) was studied in an isogenic cell line system. The inactivation of p53 decreased sensitivity to paclitaxel (1.1-2.5-fold), which correlated with a lower induction of apoptosis. The magnitude of the G2/M arrest after treatment with paclitaxel was similar in all cell lines. The cytotoxicity of paclitaxel was not enhanced by HT in either wild-type p53 or p53-inactivated cells. In conclusion, cellular sensitivity to paclitaxel depends on p53-function by its ability to induce apoptosis. Irrespective of the p53-function HT was not able to enhance the sensitivity to paclitaxel. PMID- 10717243 TI - Bax overexpression enhances cytochrome c release from mitochondria and sensitizes KATOIII gastric cancer cells to chemotherapeutic agent-induced apoptosis. AB - To evaluate whether overexpression of Bax, an apoptosis-promoting gene, sensitizes KATOIII gastric cancer cells to apoptosis induced by chemotherapeutic agents, three stable cell lines of KATOIII transfected with Bax (KATOIII-Bax), Bcl-2 (KATOIII-Bcl-2), or control pCI-neo expression vector (KATOIII-pCI-neo) were established. The cells were treated with paclitaxel, 5-fluorouracil, or doxorubicin, and the apoptotic response was measured. Our results showed that the sensitivity of the KATOIII-Bax cells to chemotherapeutic agents was enhanced compared with that of the KATOIII-pCI-neo cells, and the KATOIII-Bcl-2 cells were more resistant to these agents. Western blotting revealed that cytochrome c level in the cytosol fraction of the KATOIII-Bax cells was higher than that of the KATOIII-pCI-neo cells. Significant increase of cytochrome c level in the cytosol fraction of the KATOIII-Bax cells was detected 24 h after exposure to chemotherapeutic agents, when apoptotic cells were less than 10%. The cytochrome c level in the cytosol fraction of the KATOIII-Bax cells was higher than that of the KATOIII-pCI-neo cells at all time points examined after exposure to chemotherapeutic agents. Marked activation of caspase-3 in the KATOIII-Bax cells was observed 48 h and 72 h after exposure to chemotherapeutic agents compared with that in the KATOIII-pCI-neo cells. Consistently, zVAD-fmk, a pancaspase inhibitor, repressed the paclitaxel-induced apoptosis. In addition, Bcl-2 overexpression strongly blocked KATOIII cell apoptosis by inhibiting the cytochrome c release from mitochondria and caspase-3 activation. These findings suggest that cytochrome c release is a major mechanism of apoptotic response and Bax overexpression sensitizes KATOIII cells to chemotherapeutic agent-induced apoptosis through enhancing the release of cytochrome c from mitochondria. PMID- 10717244 TI - Increased expression of NIPP-1 mRNA correlates positively with malignant phenotype in rat hepatomas. AB - In order to elucidate an involvement of protein phosphatase type 1 (PP1) in the malignant transformation, we have isolated rat cDNA for NIPP-1, a nuclear inhibitor for PP1. Rat NIPP-1 protein conserved the PP1 binding domain and the nuclear targeting sequence. The mRNA was ubiquitously expressed with the highest expression in bone marrow and thymus. The rat NIPP1 mRNA levels were slightly, if at all, increased in some primary hepatomas, and significantly increased in rat ascites hepatomas. The levels of NIPP-1 mRNA were closely correlated with malignant phenotype of rat ascites hepatomas, suggesting that NIPP-1 is tightly linked to malignant phenotype in hepatomas. PMID- 10717245 TI - Ribosomal protein L7a gene is up-regulated but not fused to the tyrosine kinase receptor as chimeric trk oncogene in human colorectal carcinoma. AB - Ribosomal protein L7a (rp L7a) was identified in a subtractive hybridization screen as a gene up-regulated in human colorectal cancer. Expression of rp L7a was greater than 2-fold higher in tumors compared to adjacent normal mucosa in 72% of the patients studied (n=36). rp L7a was also up-regulated in concomitant polyps. The number of patients with rp L7a T/N ratio of >2 was significantly higher in the female (16/18) than in the male (10/18). rp L7a expression was also significantly higher in females with lymph node involvement compared to males. These results indicate that rp L7a expression is related to tumor growth in colorectal cancer especially in females, where it may also be related to tumor spread. There was no correlation of rp L7a expression with tumor cell differentiation. We also show that rp L7a does not exist as a fusion oncogene (trk-2h) in colorectal cancer. PMID- 10717246 TI - Expression of hnRNP A2/B1 proteins in human cancer cell lines. AB - The overexpression of heterogeneous nuclear ribonucleoprotein (hnRNP) A2/B1 is recently reported informative as a marker of lung cancer for early detection. To examine whether the expression of the proteins is specific for lung cancer, immunological analyses were performed both in lung and non-lung cancer cell lines. In immunostaining and Western blotting, the expression of A2/B1 was observed not only in the lung cancer cells but also in non-lung cancer cells. The expression of hnRNP A2/B1 is not specific for lung cancer and quantitative determination of A2/B1 is required to elucidate their significance in carcinogenesis. PMID- 10717247 TI - Doubling of the epirubicin dosage within the 5-fluorouracil, epirubicin and cyclophosphamide regimen: a prospective, randomized, multicentric study on antitumor effect and quality of life in advanced breast cancer. AB - In metastatic breast cancer (MBC) doubling the epirubicin (EPI) dose intensity (DI) within the FEC (5-fluorouracil, EPI, cyclophosphamide) regimen could increase the response rate (RR) and ameliorate the quality of life (QoL) over standard FEC. From May, 1995, 74 consecutive patients with MBC were randomly treated with 6 courses of two FEC regimens containing 60 (60FEC) or 120 (120FEC, supported by primary G-CSF) mg/m2 of EPI. Drugs were administered every 21 days. The QoL was assessed over and after treatment by the EORTC QLQ-C30 (VER 2.0) and QLQ-BR23 questionnaires, compiled by the patient, and the Spitzer's QL-index, compiled by the physician. The study was prematurely closed in May, 1997, due to RR and QoL data of 4th interim analysis. The delivered EPI DI was 20.0 and 37.9 mg/m2/week in 60- and in 120FEC, respectively. Among the two regimens, there was no statistically significant difference in RR or in improvement of baseline overall QoL. With respect to 60FEC patients, the 120FEC patients had longer time to progression (19.2 vs 13.1 mos, p=0.04). Over baseline, the 120- but not the 60FEC patients had significantly greater pain decrease and lower deterioration of body image. In MBC, both 60- and 120FEC regimens furnished the same RR and improvement in overall baseline QoL. With respect to 60FEC patients, the 120FEC patients experienced longer time to progression. Over baseline, pain decrease and preservation of body image were also greater in these patients. PMID- 10717248 TI - VEDex (vincristine, epirubicin dexamethasone): an effective and well tolerated palliative chemotherapy regimen for non-Hodgkin's lymphoma. AB - An evaluation was carried out of the efficacy and toxicity of a novel weekly palliative chemotherapy regimen comprising vincristine, epirubicin and dexamethasone (VEDex) in 57 patients with non-Hodgkin's lymphoma (NHL) treated at this centre. The age range was 34-88 years and the median age was 67 years. Twenty-three patients (40%) had low grade disease and 4 patients (7%) had transformed NHL. Thirty patients (53%) had high grade NHL; 7 had relapsed after conventional chemotherapy and were not fit for high-dose chemotherapy, 7 were heavily pre-treated, 8 had received prior radiotherapy and 8 had not received any prior therapy. Responding patients received a total of 8 weeks of treatment, but treatment could be repeated at a later stage if required. The overall response rate was 66.6%; 11 patients (19.3%) achieved a complete response and 27 (47.3%) achieved a partial response. A further 11 patients (19.3%) had stable disease. Twenty-four patients (42.1%) reported complete resolution of symptoms and 21 (36.8%) had partial resolution of symptoms. The median survival from the onset of treatment was 6 months. Grade III neutropenia was seen in 9 patients (15.8%). Other toxicity included nausea and vomiting grade II (3.5%), grade III (1.8%) and alopecia grade III (1.8%). There were no treatment related deaths. We conclude that VEDex is an effective palliative treatment in patients with indolent or aggressive lymphoma with poor performance status or who have been heavily pre treated. It is well tolerated in the elderly. PMID- 10717249 TI - Lipids: A key role in multidrug resistance? (Review). AB - Among tumoral resistances, multidrug resistance (MDR) is characterized as cross resistance to a variety of structurally and functionally unrelated drugs such as vinca alkaloids, colchicine, and anthracyclines. Decreased drug cellular influx and increased cellular ability for drug extrusion are the main mechanisms involved in MDR. Two plasma membrane proteins, p-glycoprotein (p-gp) and the multidrug resistance-associated protein (MRP), act as ATP-dependent cellular efflux. Furthermore, protein kinase C (PKC) is also central to MDR. The present study reviews the role of cholesterol and other lipids in the reduction of drug influx and drug binding to cellular membranes. The study also examines the effect of lipid composition on p-gp activity. Concerning the role of PKC in MDR, two phospholipases involved in diacylglycerol (DG) production increase in MDR cells. These are phosphatidylinositol-4, 5-bisphosphate-specific phospholipase C and phosphatidylethanolamine-specific phospholipase D. A positive feedback mechanism for DG production which includes these phospholipases, a phosphatidylcholine specific phospholipase C and a phosphatidylcholine-specific phospholipase A2 has also been suggested. The hypothesis of exocytic involvement in MDR is reviewed, and some lipid changes found in MDR cells are interpreted according to those fusogenic properties normally involved in exocytic transport. Also, the possible role of lipid mediators, such as phosphatidic acid and platelet-activating factor, is examined. PMID- 10717250 TI - Assessment of Stanniocalcin-1 mRNA as a molecular marker for micrometastases of various human cancers. AB - We assessed Stanniocalcin-1 (STC-1) mRNA expression in human normal tissues, various types of human cancer cell lines, and cancer tissues obtained during surgery. Using a reverse transcriptase-polymerase chain reaction (RT-PCR) assay developed to detect STC-1 mRNA, the transcripts were detected in 20 out of 21 cancer cell lines and in all tumor tissues from various types of cancer. Semi quantitative analyses with multiplex RT-PCR showed that STC-1 mRNA tended to be enhanced in cancer tissues of hepatocellular carcinoma and colorectal cancer compared to background cancer-free tissues. Analysis of blood samples obtained from 11 patients with hepatocellular carcinoma before, after and during hepatectomy showed STC-1 mRNA expression in 8 out of 11 patients at least one time. Normal donor blood samples (n=31) were all-negative for STC-1 mRNA expression. Our results indicate that STC-1 mRNA might be a useful molecular marker for detection of tumor cells in blood from patients with various types of malignancies. PMID- 10717252 TI - Effect of low molecular weight heparin (Certoparin) versus unfractionated heparin on cancer survival following breast and pelvic cancer surgery: A prospective randomized double-blind trial. AB - Recent studies suggest that low molecular weight heparin (LMW heparin) therapy in malignancy may improve cancer survival following surgical resection. We studied prospectively whether cancer mortality during follow-up in women with previously untreated breast, and pelvic cancer is reduced in those who randomly received LMW heparin (Certoparin) compared to patients given unfractionated heparin (UF heparin) for thrombosis prophylaxis during primary surgery. In a prospective, randomized, double-blind clinical trial, 160 patients received Certoparin and 164 UF heparin until post-operatively day 7. Survival estimations are based on the outcome data from a subset of 140 LMW heparin - and 147 UF heparin recipients. Long-term survival in the Certoparin group compared to the UF heparin group was significantly improved after 650 days (P=0. 0066) but not thereafter when analysis was performed on all cancer cell types combined. In the probability estimates survival benefit within this time was restricted to patients with pelvic cancer but was not observed in breast cancer. However, in breast cancer patients who received LMW heparin the impact of classical tumor prognostic markers was statistically significant after 1,050 days but not after 650 days. Thus, breast cancer patients with unfavorable prognosis seem to benefit in terms of survival advantage from LMW heparin within the 650 days after surgery. These results suggest that improvement in cancer survival can be achieved after even a short course of treatment with LMWH (compared to UFH) given for DVT prophylaxis in the post-operative period. An effect of UFH on disease outcome is not excluded. Further definitive trials of LMWH vs. placebo for cancer outcome (rather then DVT) using doses and schedules that may be more optimal are indicated. PMID- 10717251 TI - The development of peritumoral stroma required for IL-12 induced tumor regression depends on the T cell/IFN-gamma-involving host-tumor interaction. AB - T cell migration into tumor masses is a critical process in the scenario of IL-12 induced tumor regression. Our previous study showed that this depends on the development of peritumoral stroma prior to IL-12 therapy. The present study investigated the regulation of the development of peritumoral stroma in comparison with tumor-parenchymal stroma. In the OV-HM and CSA1M tumor models, tumor regression associated with T cell migration was induced following IL-12 treatment. Both OV-HM and CSA1M tumor masses growing in syngeneic mice developed peritumoral stroma before IL-12 treatment. However, peritumoral stroma was not observed in these two types of tumor masses generated in nude mice, T cell depleted syngeneic mice, anti-IFN-gamma mAb-treated mice or IFN-gamma-deficient mice. In contrast, parenchymal stroma formation did not appear to be affected because tumors generated in these groups of mice exhibited rather higher growth rates than those of tumors in normal syngeneic mice. Importantly, the lack of peritumoral stroma in tumor masses was associated with the failure of T cells to migrate to these tumor masses: splenic T cells prepared from IL-12-treated tumor bearing mice migrated into the corresponding tumor mass growing in untreated syngeneic recipient mice, whereas portions of the same donor cells failed to migrate into the above stroma-negative tumor masses. These results indicate that the development of peritumoral and parenchymal stroma is differentially regulated; there exist functional differences in the two types of stroma; and the formation of peritumoral stroma requires components of the host's immune system such as IFN-gamma and T cells. PMID- 10717253 TI - Lack of defective expression of the ATM gene in sporadic breast cancer tissues and cell lines. AB - Homozygous mutations of the gene mutated in ataxia telangiectasia (ATM) causes the AT syndrome, a pleiotropic phenotype that includes an increased risk of cancer. Most of the known mutations at the ATM gene lead to truncations which are usually associated with instability of mRNA and protein. A decrease or loss of ATM protein expression is associated with specific lymphoid malignancies in AT and non-AT patients. ATM is located within a region in chromosome 11q22-23 that is frequently undergoing loss of heterozygosity in sporadic breast cancer. Epidemiological studies estimated a 4-fold increase in breast cancer risk in heterozygous women. However, direct mutational analysis failed to clearly support a role for mutant ATM alleles in breast carcinogenesis. If ATM does have a suppressor role in this tissue, one would expect deficient ATM expression. We therefore tested the hypothesis that the expression of the ATM gene is reduced in sporadic breast cancer. We determined ATM transcript levels using competitive RT PCR on 89 randomly selected sporadic breast cancer samples and 29 normal breast tissues. Of these, 11 were matched normal/cancer pairs. We also evaluated 7 breast cancer cell lines. Deficiency in ATM expression was not observed. Of the 11 matched pairs, 7 tumors expressed mildly higher levels, 3 tumors expressed the same amount and only 1 tumor expressed <50% of the normal match. In addition, 3 cancers with tumor-associated LOH of the ATM gene expressed higher mRNA levels in the tumors than in their normal tissue matches, suggesting that no correlation exists between tumors with LOH and decreased ATM expression. In summary, our results do not support a suppressor role for ATM in the development of sporadic breast cancer. PMID- 10717254 TI - Natural history of hepatitis C and the impact of anti-viral therapy. AB - The hepatitis C virus (HCV) infects some 170 million people worldwide and is responsible for approximately 20% of cases of acute hepatitis and 70% of cases of chronic hepatitis. Acute hepatitis is icteric in only 20% of patients and is rarely severe. Eighty five per cent of the infected patients develop chronic infection which is generally asymptomatic. Among the HCV chronic carriers, 25% have persistently normal serum alanine aminotransferase (ALT) levels despite having detectable HCV-ribonucleic acid in serum, 75% have elevated ALT levels. While the majority of patients with mild chronic hepatitis have a slowly progressive liver disease, the patients with moderate or severe chronic hepatitis may develop cirrhosis within a few years. In patients with HCV-related cirrhosis, the incidence of hepatocellular carcinoma is 2-5% per year. HCV-related end-stage cirrhosis is currently the first cause of liver transplantation. Treatment with the combination of interferon-alpha and ribavirin induces a sustained virological response in roughly 40% of the patients. The virological response is associated with a biochemical response and histological improvement. It is believed that the decrease of necroinflammatory liver lesions induced by anti-viral therapy in responders, is associated with a decreased risk of development of cirrhosis and hepatocellular carcinoma PMID- 10717255 TI - Interferon and ribavirin combination therapy: indications and schedules. AB - Treatment outcome for patients with chronic hepatitis C virus infection has greatly improved during the last years with the development of interferon (IFN) and ribavirin combination therapy. The final decision to treat or not, however, is complex and should be based on several factors such as the age of the patient, the general health, the risk of developing cirrhosis and the probability of a cure with treatment. Combination therapy with standard doses (IFN-a 3 x 106 IU three times per week plus ribavirin 1000-1200 mg daily in two divided doses) for six (up to 12) months significantly improves the sustained biochemical and virological response rates 2-3 times as compared to IFN alone given during 12 months. Combination therapy has thus become standard therapy for na ve patients and relapse patients after a prior IFN treatment course. For patients with favourable baseline viral characteristics (genotype 2 and 3 irrespective of viral load) six months combination therapy is sufficient whereas patients with unfavourable viral baseline characteristics (genotype 1 with high baseline viral load) will need 48 weeks combination treatment. In addition, for patients with compensated cirrhosis, combination therapy is superior and better tolerated than IFN monotherapy. For the future better optimised treatment schedules and dosing regimens for IFN in combination with ribavirin need to be worked out and individualised according to genotype to further improve treatment results. Utilisation of new IFN formulas such as pegylated IFN and consensus IFN in combination regimens will probably improve treatment further. PMID- 10717256 TI - Hepatitis C virus: kinetics and quasispecies evolution during anti-viral therapy. AB - The balance of virus production and clearance for untreated patients with chronic hepatitis C changes into a decline of viraemia when initiating effective anti viral treatment. During the first phase of interferon-alpha (IFN-a) therapy, the kinetics of the viral load is characterised by a rapid dose-dependent decline starting after a delay of about eight to nine hours. This early response can be observed for almost all patients treated with IFN-a. After about 24 to 48 hours, the viral decline slows down leading to a second phase with a relatively stable exponential decay. Some non-responding patients show a nearly constant viraemia and some even a rebound throughout this second phase. Kinetic models allow the estimation of rates of viral production and clearance and reveal high turnover rates of hepatitis C virus (HCV) and an in vivo half-life of hepatitis C virions of a few hours, only. Due to the continuous and high replication rate in vivo, the low fidelity of the ribonucleic acid (RNA)-dependent RNA polymerase, and the immune surveillance of the host, HCV exists in an individual patient as a heterogeneous population of related viruses (quasispecies). A high degree of quasispecies variability correlates with a lower response to IFN-a therapy. Changes of the quasispecies population are more pronounced after initiation of treatment with IFN-a or interleukin-12 than during the natural course of disease. Ribavirin, however, has not been found to affect the HCV quasispecies population. Identification of a specific region within an envelope-encoding gene as the most variable region of HCV and as a critical neutralisation domain suggests that viral escape mechanisms are a possible cause for chronification and poses a major challenge for the development of a broadly reactive vaccine against HCV. PMID- 10717257 TI - The molecular basis for responsiveness to anti-viral therapy in hepatitis C. AB - Hepatitis C virus (HCV) infection is an important clinical problem, with a world wide prevalence of approximately 1-2%. HCV infection is associated with an increased risk for the development of severe liver disease. HCV is inherently resistant to anti-viral therapy with interferon (IFN). The virus circulates in infected individuals as a mixture of related, yet genetically distinct variants, or quasispecies. Many studies have implicated HCV quasispecies in IFN responsiveness. Effective containment of HCV quasispecies mutation and selection through more aggressive therapy (e.g. daily induction), combination therapy (e.g. IFN plus ribavirin), or longer lasting therapy (e.g. pegylated IFN) is required for IFN responsiveness. Recently, several HCV proteins including the non structural 5A and envelope gene 2-glycoprotein have been implicated in HCV anti viral resistance. It is likely that multiple HCV genes disrupt IFN-induced anti viral responses at many levels and that these virus-host cell interactions are associated with IFN resistance. Characterisation of HCV-encoded mechanisms of anti-viral resistance has important implications for the development of new anti virals. PMID- 10717258 TI - What to do when standard therapy fails. AB - An important group of patients with chronic hepatitis C do not respond to interferon (IFN) therapy. Compared with untreated patients with chronic hepatitis C, non-responders have a higher percentage of cirrhosis, are more frequently infected by genotype 1 and usually have a viral load above 2 x 106 copies/ml. Also, patients with cirrhosis have lower life expectancy and higher risk of clinical complications, and therefore, are most in need of effective treatment strategies. There is no evidence that the re-treatment of non-responders with a standard regimen of IFN or more prolonged IFN therapy achieves a sustained biochemical or virological response. Between 20% and 40% of non-responder patients treated with IFN therapy for more than two years had an hepatic improvement in liver histology associated with a decrease in hepatitis C virus ribonucleic acid levels. In contrast, combination therapy with IFN and ribavirin for six months now results in sustained response rates between 6% and 29% depending on the viral genotype and the presence or absence of cirrhosis. Patients infected with genotype 2 and 3 have a higher probability of achieving a sustained virological response than those infected by genotype 1. Currently, different studies are underway to determine whether high-dose IFN and/or induction therapy combined with ribavirin for more prolonged periods of time could increase the sustained response rate in non-responders. No other drugs appear to be efficacious in these patients, except the combination of IFN, ribavirin and amantadine which has shown interesting results in a preliminary trial but they need to be confirmed in further studies. These findings suggest that combination therapy is beneficial and can be recommended for some non responder patients until other new therapies are available. PMID- 10717259 TI - Natural rubber latex aeroallergen exposure in rubber plantation workers and glove manufacturers in Thailand and health care workers in a UK hospital. AB - OBJECTIVES: To estimate personal airborne natural rubber latex (NRL) concentrations for three occupational exposure groups; rubber plantation workers and NRL glove manufacturers in Thailand and health care workers in the UK. To utilise these data to classify the populations into appropriate exposure groups for the exposure-response analysis in the epidemiological study on latex allergy. METHODS: Two rubber plantations (110 workers), three NRL glove manufacturing factories (583 workers) in Thailand and one UK hospital (490 workers) were selected for the study. A preliminary workplace survey was carried out at each workplace in order to assign job titles subjectively in to high, moderate or low exposure groups for the purpose of sample selection. Between 5 and 20% of workers from each group for the three populations were then selected randomly for personal measurement of latex airborne allergens. Personal sampling was conducted using a 25 mm PTFE filter loaded in to an IOM sampling head at 2 l. min(-1). NRL aeroallergens were measured by an inhibition assay with NRL-specific IgE antibodies from NRL-sensitised people. RESULTS: A total of twenty-two personal samples were collected from plantation workers, sixty-one samples from the glove manufacturer employees and twenty seven from health care workers. The highest geometric mean (GM) NRL aeroallergen concentration was found in the glove manufacturing factories (7.3 microg m(-3)), followed by the rubber plantations (2.4 microg m(-3)) and the UK hospital (0.46 microg m(-3)). Amongst the NRL glove factories, the NRL aeroallergen concentrations were highest for those conducting the following tasks; glove stripping, glove inspections and packing of powdered gloves. The GM NRL aeroallergen for these tasks were in the range of 12.9 to 17.8 microg m(-3). CONCLUSIONS: In the process from tapping and manufacture of latex gloves through to their use the highest exposure to NRL aeroallergens is likely to occur in the manufacturing factories. Exposure to aeroallergens for the plantation workers was considered to be moderate and that of health care workers to be low. PMID- 10717260 TI - Polycyclic aromatic hydrocarbon exposure in an artificial shooting target factory: assessment of 1-hydroxypyrene urinary excretion as a biological indicator of exposure. AB - Five representative workers and two external observers were monitored by personal air and urinary 1-hydroxypyrene (PyOH) sampling for a four-shift working week in an artificial shooting target factory. The targets (clay pigeons), are made from petroleum pitch and molded at 190 degrees C. No respiratory protective mask was worn. Atmospheric concentrations of pyrene and benzo (a) pyrene (BaP) ranged from 0.66 to 5.05 microg/m(3) and 0.037 to 0.270 microg/m(3) respectively with a mean pyrene/BaP ratio of about 20 and a correlation r = 0.51. Maximum PyOH urinary excretion ranged from 1.84 to 10.9 micromol/molCreat. This occurred at the postshift for the observers but often appeared later for workers: up to 10.75 h for the person with the apparently highest dermal exposure. The apparent PyOH excretion half lives ranged from 1.9 to 12.5 h with an arithmetic mean of 6.1 h. All these data were confirmed by additional measurements taken over a weekend after the postshift. The correlation between atmospheric pyrene and urinary PyOH concentrations (increase over the shift) was poor (r = 0.37). It improve greatly (r = 0.74) if the amount of pyrene inhaled over the shift and the corresponding amount of PyOH excreted were considered. The ratio of urinary excreted PyOH to the pyrene inhaled dose (with assumed retention of 100%), ranged from 0.18 to 0.70 (arithmetic mean = 0.34). This suggests that the respiratory tract is the main entrance route for pyrene (apart from the worker who handled crude targets without gloves). PMID- 10717261 TI - Depletion of glutathione and ascorbate in lung lining fluid by respirable fibres. AB - OBJECTIVE: The use of synthetic vitreous fibres has increased along with a decline in the utilisation of asbestos. There remains concern that these synthetic fibres pose a health risk to workers because of the generation of respirable fibres which can enter the lung and cause adverse health effects. An improved understanding of the mechanism of fibre pathogenicity should allow more rational short-term testing regimes for new fibres as they are developed. We hypothesised that carcinogenic fibres have greater free radical activity compared with non-carcinogenic fibres and that they contribute to disease by causing oxidative stress in the lung. We examined a panel of respirable fibres, designated as being carcinogenic or non-carcinogenic based on previous animal studies for ability to deplete antioxidants from lung lining fluid. METHODS: On the basis of inhalation studies, a panel of fibres was divided into three carcinogenic fibres-amosite asbestos, silicon carbide, and refractory ceramic fibre 1 (RCF1) and three non-carcinogenic fibres-man-made vitreous fibre 10 (a glass fibre MMVF10), Code 100/475 glass fibre, and refractory ceramic fibre 4 (RCF4). We measured the levels of glutathione (GSH) and ascorbate, two antioxidants present in lung lining fluid (LLF) after fibre treatment. All of the experiments were carried out at equal fibre number. RESULTS: Fibres had the ability to deplete both GSH and ascorbate from both LLF and pure solutions, an effect which was fibre number dependent. The greatest depletion of antioxidants was observed with the two non-carcinogenic glass fibres, and this effect was observed when A549 lung epithelial cells were treated with fibres. CONCLUSIONS: Our results show that antioxidant depletion in cell free solution and lung lining fluid solely is not a simple indicator of the ability of fibres to cause lung pathology and that other biological events in the lung are involved. PMID- 10717262 TI - Asbestos content of lung tissue and carcinoma of the lung: a clinicopathologic correlation and mineral fiber analysis of 234 cases. AB - The aim of this study was to investigate the asbestos content of lung tissue in a series of patients with lung cancer and some history of asbestos exposure. This information was then correlated with demographic information, occupational and smoking history, presence or absence of pathologic asbestosis or pleural plaques, and pathologic features of the cancer. The pulmonary concentration of asbestos fibers in 234 cases of primary carcinoma of the lung was determined by means of a tissue digestion technique. Asbestos body counts were performed in 229 cases and fiber analysis by scanning electron microscopy in 221 cases. Asbestos content was recorded as total asbestos fibers, commercial amphibole fibers, noncommercial amphibole fibers, and chrysotile fibers 5 microm or greater in length per gram of wet lung tissue. The study group included 70 patients with asbestosis (Group I), 44 patients with parietal pleural plaques but without asbestosis (Group II), and 120 patients with neither (Group III). The median asbestos body content of Group I was more than 35 times greater than Group II and more than 300 times greater than Group III. The total asbestos fiber count for Group I was nearly 20 times greater than Group II and more than 50 times greater than Group III. The difference was due almost entirely to commercial amphiboles. In a series of primary lung cancer cases with some history of asbestos exposure, a markedly elevated asbestos content was identified among those with pathologic asbestosis as compared with patients with pleural plaques alone or with neither plaques nor asbestosis. PMID- 10717263 TI - Exposure to organic solvents in the offset printing industry in Norway. AB - The purpose of this study was to document the conditions regarding solvent exposure at offset printing offices in Norway at present and to study the variation of exposure between printing office technologies. Measurements were made at seven offset printing offices. The measurements consisted of five to 10 whole day personal exposure measurements at each office performed over a period of 2 months. Variables that may influence the level of exposure were registered by the occupational hygienist at the end of each measuring day using a check list. The influence of the variables on the "additive factor" was examined by linear regression analysis.The main contributor to the "additive factor" was isopropanol. The exposure to isopropanol sometimes exceeded the Norwegian TLV. The exposure decreased when a separate exhaust ventilation was used. The exposure increased when the machine had automatic cleaning. The variables automatic cleaning and separate exhaust ventilation explained 59% of the variation in the "additive factor". The results of this study indicate that the most important source of solvent exposure in printing offices at present is the moisturizer used in the printing machines. We think it is worth giving attention to this exposure and making efforts to reduce it. PMID- 10717264 TI - Influence of evaporation and solvent mixtures on the absorption of toluene and n butanol in human skin in vitro. AB - The influence of forced ventilation on the percutaneous absorption of butanol and toluene was studied in vitro. Human skin was exposed to the neat solvents and the solvents in binary mixtures with each other and in ternary mixtures with chloroform:methanol. The exposure was either unventilated or ventilated with various flow rates. At the ventilated exposure the skin absorption of all solvents and solvent mixtures was markedly reduced compared to unventilated exposure. Exposure with solvent mixtures increased the amounts of solvent absorbed as well as absorption rates. The absorption of the butanol component was most influenced. Increase in absorption was 11 to 9 times depending on whether toluene or chloroform/methanol was cosolvent. There was also an interindividual variation of absorption rate, varying with a factor of 3.5 for toluene and 4.3 for n-butanol within the 3 skin donors used. Skin absorption of volatile organic solvents at continuous ventilated conditions is related to their volatility and to the ventilation rate.A sufficient workplace ventilation is an important occupational hygienic measure not only to reduce exposure via respiration but to reduce absorption via the skin of volatile compounds as well. PMID- 10717265 TI - Microscopic identification of asbestos fibres associated with African clay crafts manufacture. AB - The use of asbestos in manufacturing is a world-wide phenomenon, not just confined to the developed world. The activity described below shows that there are similar problems in the third world which need to be tackled. A sample of white fibrous material used in pot making by women in a village of Botswana was provided for analysis. The identification of fibres was carried out using established analytical and vibrational microspectroscopic methods. The occupational hygiene implications and the measures which may need to be taken in order to improve the safety of the pot making process are discussed in this article. PMID- 10717266 TI - Required response time for variable air volume fume hood controllers. AB - This paper describes results from tests made with the aim of investigating how quickly the exhaust air flow rate through fume hoods needs to be controlled in order to prevent contaminants from leaking out of the fume hood and putting the safety of the laboratory personnel at risk. The measurements were made on a laboratory fume hood in a chemical laboratory. There were no other fume hoods in the laboratory, and the measurements were made without interference from persons entering or leaving the laboratory or walking about in it. A tracer gas method was used with the concentration of dinitrogen oxide (N(2)O) being recorded by a Foxboro Miran 101 infra-red gas analyser. In parallel with the tracer gas measurements, the air velocity through the face opening was also measured, as was the control signal to the damper controlling the air flow rate. The measurements show an increased outward leakage of tracer gas from the fume hood if the air flow rate is not re-established within 1-2 s after the sash is opened. If the delay exceeds 3 s the safety function is temporarily defeated. The measurements were made under virtually ideal conditions. Under more typical conditions, the fume hood could be exposed to various other external perturbations, which means that the control system should re-establish the correct exhaust flow more quickly than indicated by the measurement results obtained under these almost ideal conditions. PMID- 10717267 TI - External fixation in the upper limb. PMID- 10717268 TI - Treatment of pain in trauma patients with injuries of the upper limb. AB - Since preoperative pain therapy of a trauma patient did not play an outstanding role in the past, this article shall give information about the adequate treatment of such patients, which can be mainly divided into three phases: the prehospitalisation phase with stabilisation of the trauma patient, the early phase of hospitalisation with further stabilisation, diagnosis and surgery, and finally the postoperative phase with corresponding treatment. An optimal analgesic in the prehospitalisation phase should guarantee good analgesic effects, rapid onset and good controllability, simple handling and the opportunity to combine it with other medication. In addition, it should prevent a wide therapeutic range and the absence of side effects. Opioids and ketamine are available for acute pain therapy after trauma. The main opioids used are piritramide and pethidine, with piritramide acting as a sedative at the same time and with pethidine preventing the stronger analgesic effect. The intravenous use of ketamine has become established in trauma patients because of its excellent analgesia at subanaesthetic doses. Especially in multiple trauma patients, the indication for general anaesthesia with intubation should be established on a liberal basis. Nevertheless, for some patterns of injury regional techniques may be advantageous; therefore, this article describes the possible regional procedures (such as intravenous regional anaesthesia or block of peripheral nerves). Concerning the postoperative phase, an individual pain management can be guaranteed by systemic pharmacotherapy and regional catheter techniques, for example the brachial plexus blockade that results in a long period of free pain. PMID- 10717269 TI - The anatomical base of unilateral external fixation in the upper limb. AB - Unilateral external fixation requires an anatomically sound implantation of screws into the upper extremity. Detailed knowledge about the anatomical situation in the areas of pin implantation is of great importance. This paper focuses on relevant anatomical landmarks when implanting screws for external fixation in the humerus, the elbow, the forearm and the hand by studying anatomical specimen. PMID- 10717270 TI - The use of upper limb external fixation in paediatric trauma. AB - External fixation is an alternative method of treatment for paediatric fractures of the upper extremity. We report our experience of the management of 23 children with an average age of 10 years 1 month and review the literature. The method is indicated for second and third degree open fractures, open multiply injured patients and severely comminuted fractures. External fixation has proved unequalled for correction of limb deformities and lengthening procedures. PMID- 10717271 TI - Is there a place for external fixation in humeral shaft fractures? AB - There is a good indication for unilateral axial dynamic external fixation in fractures of the humeral shaft when the fracture appears in the distal third or in cases of bilateral fractures. A non-union or a posttraumatic paralysis of the radial nerve may be indications for external fixation as well as fractures associated with multiple injuries. Further indications include osteitis, infected non-union and comminuted fracture. There is maximum protection of the soft tissue with this method of treatment. External fixation combines the advantages of conservative and operative treatment by influencing callus formation by dynamizing, distraction or compression. Minimizing soft tissue damage facilitates the decision for early exploration of the radial nerve in cases of palsy. A safer positioning technique of the distal screws of the fixator is described. PMID- 10717272 TI - Transarticular fixation with the capacity for motion in fracture dislocations of the elbow. AB - Post-traumatic stiffness of the elbow joint is a frequent result of immobilisation leading to severe disability in the use of the upper extremity. Recognition of the tendency to stiffness leads to the assumption that the strong self-healing forces of the capsule and ligament apparatus converts the initial instability of the joint after ligament disrupture, into a high-grade undirected stability following immobilisation. Directed stability as it is produced by the natural ligament apparatus of the joint on the other hand produces a guided movement of the joint in one direction. These theoretical considerations lead to the idea that the self-healing forces of the ligament apparatus under continuous guided movement of the joint will result in a stable and movable joint to allow healing of the compromised soft tissue envelope and moreover to maintain free soft tissue access without compromising the stability. For this a unilateral fixator with motion capacity was developed. The joint bridging application approaches the humerus and ulna from the lateral side. The proximal pin group is inserted into the proximal region of the humerus respecting the radial nerve. The distal pin group is implanted from the dorsal side into the middle third of the ulna. The fixator has a hinge joint. The design of the fixator clamps, bars and the hinge joint allows simple alignment with the rotational axis of the elbow. Pro- and supination of the forearm is unhindered. Flexion and extension can be permitted according to the soft tissue situation. PMID- 10717273 TI - External fixation in forearm shaft fractures. AB - External fixation for uncomplicated forearm fractures is rarely performed. The situation is different in a multiply injured patient or in a fracture with considerable soft tissue damage. In these cases the external fixator confers quick and efficient stabilisation which meets the requirements of adequate nursing and aids recovery of the general and local condition. Later change to an appropriate internal fixation procedure for definitive fracture treatment is recommended. PMID- 10717274 TI - Reduction techniques in distal radius fractures. AB - External skeletal fixation is an important minimal invasive procedure in the management of fractures at the distal end of the radius. Attention to detail is important not only in the recognition of indications and function of the external fixator but also in its specific application. To improve anatomical restoration (e.g. palmar tilt) multi-planar ligamentotaxis is recommended. The demonstrated dynamic fixator is easy to apply and the double ball joint facilitates reduction after mounting by multi-planar ligamentotaxis. This transarticular unilateral external fixation system permits restoration of anatomy with the wrist in neutral or extension, thereby allowing full flexion of the metacarpophalangeal joints, with fingers and wrist extensor tendons relatively relaxed. Multi-planar ligamentotaxis, combined with a limited approach for supplementary procedures (fixation of articular fragments and/or bone grafting) and with early wrist motion, offers an encouraging treatment option in the management of unstable distal radial fractures by providing better anatomical restoration, especially of the palmar tilt and reduces the risk of wrist and finger stiffness. PMID- 10717275 TI - Principles of external fixation and supplementary techniques in distal radius fractures. AB - External fixation for fractures of the distal radius has been used for almost 80 years. The main objective is to gain reduction and maintain the reduction throughout the treatment period. Several fixator concepts are available and selection is based on the complexity of the injury to be treated as well as the surgeon's experience. Periarticular application of the fixator with immediate use of the wrist joint is recommended whenever possible. For intra-articular fractures, transarticular application is advisable. External fixtion in complex fractures has to be supplemented by bone grafting, fixation wires and stabilization of the radioulnar joint. Associated injuries in distal radius fractures need to be identified and treated. The possible complications of external fixation and the means to prevent them are discussed. External fixation of the distal radius has found its place as an established method in treating certain types of this common fracture. PMID- 10717277 TI - Corrective osteotomies in malunited distal radius fractures: external fixation as one stage and hemicallotasis procedures. AB - The correction of distal radius malunion was carried out using a radio-radial fixator. The two techniques used in 14 cases were one stage correction or gradual correction by hemicallotasis. The techniques are described and the patients treated were evaluated. PMID- 10717276 TI - Results of transarticular fixator application in distal radius fractures. AB - From January 1989 until October 1993, 102 patients with 103 distal radius fractures were included in a prospective study with the Pennig wrist fixator (Orthofix, Srl, Italy). 90.5% of these patients were reviewed with a minimum follow-up of 12 months. Using the functional outcome score according to Gartland and Werley, we obtained 41% excellent, 46% good, 10% fair and 3% poor results. Additional procedures such as K-wires, bone grafting or radio-ulnar stabilizations were carried out in 61% of cases. The complications included five fixator displacements, four of which happened with the prototype used until the first half of 1990. Two major pin-track-infections required surgical intervention, three patients in the beginning of the series encountered irritations of the superficial radial nerve, one pin cut out of the second metacarpal bone and one patient sustained algodystrophy. The results show the significance of an anatomical reduction, the restoration of the radial length seems to be of special importance to obtain good functional outcome. This can not always be achieved with the fixator alone; often additional procedures are required in order to obtain an anatomical joint reconstruction. The advantage of this fixator lies in its lightweight and small-size design as well as in its easy application technique and the possibility to carry out the reduction after mounting the fixator with the double ball joint centered on the fracture level and the carpus. PMID- 10717278 TI - The treatment of complex carpal dislocations by external fixation. AB - This article presents the treatment of complex carpal dislocations and fracture dislocations by external fixation. Twenty-four patients were treated with external fixation after complex carpal trauma. Operative technique, aftertreatment, complications and long-term results are presented and discussed. PMID- 10717279 TI - The use of minimally invasive fixation in fractures of the hand--the minifixator concept. AB - The minimally-invasive treatment of metacarpal and phalangeal fractures with a low profile external fixator is described. In most sites fixator pins are inserted percutaneously and closed reduction of the fracture is performed. Due to the special design bridging of adjacent joints in shaft, distal or proximal fractures is not necessary unless the fracture is intraarticular. The results of treatment are described. Advantages and disadvantages of a small external fixator in relation to conservative management and open reduction and internal fixation are discussed. PMID- 10717280 TI - Callus distraction in the hand skeleton. AB - From 1992 to 1996, 27 patients with traumatic amputations or malformations underwent lengthening of thumb and fingers. A total of 36 procedures were carried out. In several cases, deepening of the web space or bone transplantations proved to be necessary to improve general function or to compensate for missing bone structure. Complications included pin-track infections, necrosis of bone, non union and scarring. Overall the results indicate that distraction of the thumb and fingers is a feasible operative technique leading to a promising improvement of function. PMID- 10717282 TI - Avian macrophage: effector functions in health and disease. AB - Monocytes-macrophages, cells belonging to the mononuclear phagocytic system, are considered as the first line of immunological defense. Being mobile scavenger cells, macrophages participate in innate immunity by serving as phagocytic cells. These cells arise in the bone marrow and subsequently enter the blood circulation as blood monocytes. Upon migration to various tissues, monocytes mature and differentiate into tissue macrophages. Macrophages then initiate the 'acquired' immune response in their capacity as antigen processing and presenting cells. While responding to their tissue microenvironment or exogenous antigenic challenge, macrophages may secrete several immunoregulatory cytokines or metabolites. Being the first line of immunological defense, macrophages therefore represent an important step during interaction with infectious agents. The outcome of the macrophage-pathogen interaction depends upon several factors including the stage of macrophage activation, the nature of the infectious agent, the level of genetic control on macrophage function as well as environmental and nutritional factors that may modulate macrophage activation and functions. Research in avian macrophages has lagged behind that in mammals. This has been largely due to the lack of harvestable resident macrophages from the chicken peritoneal cavity. However, the development of elicitation protocols to harvest inflammatory abdominal macrophages and the availability of transformed chicken macrophage cell lines, has enabled researchers to address several questions related to chicken macrophage biology and function in health and disease. In this manuscript the basic profiles of several macrophage effector functions are described. In addition, the interaction of macrophages with various pathogens as well as the effect of genetic and environmental factors on macrophage functional modulation is described. PMID- 10717281 TI - Mannan-binding lectin (MBL) in chickens: molecular and functional aspects. AB - Mannan-binding lectin (MBL) is a serum collectin (i.e. mosaic protein with collagenous and lectin domains) involved in the innate immune defence against various microbes. In vitro studies indicate that MBL exerts its function by binding to the microbial surface through its carbohydrate recognition domains followed by direct opsonization or complement activation via the MBL associated serine proteases MASP-1 and MASP-2. In Aves (i.e. chickens), as in man, only one MBL form has been found, while traditional laboratory animals (i.e. mouse and rat) have two MBL forms in serum. MBL has been extensively studied in mammals but recently also in Aves. This review summarizes the present knowledge of MBL in chickens and compares it to the situation in mammals. PMID- 10717283 TI - Nonspecific cellular defense of the avian respiratory system: a review. AB - The normal, steady-state, avian respiratory system has very low numbers of residing avian respiratory phagocytes (ARP). Birds must rely heavily on the influx of ARP to defend against infectious agents. The system is refractory to elicitation by inert stimulants, but responds efficiently to replicating bacteria, with very rapid influx of large numbers of activated ARP (polymorphonuclear neutrophils, heterophils, and macrophages) with increased phagocytic proportions and capacities. The numbers subside within a few a days. Activated ARP act in a non agent-specific manner: Pasteurella multocida-activated ARP can defend against a severe Escherichia coli airsacculitis. Parenteral routes of stimulation generally are not, respiratory routes are very, efficient in activating ARP. Heterophils are the most efficient in defensive reactions, such as oxidative burst, production of nitric oxide and killing of bacteria. Respiratory viruses may stimulate, but also may diminish some of the defensive functions of ARP. This is also true for attenuated, modified live virus vaccines. These vaccines must be used carefully in the presence of subclinical bacterial, mycoplasmal infections. Published literature on non-specific cellular defense of the avian respiratory system is very limited, particularly about interactions among multiple infectious agents and the system. PMID- 10717284 TI - Immunocytochemical techniques to investigate the pathogenesis of infectious micro organisms and the concurrent immune response of the host. AB - For poultry as well as for mammalian species used for scientific research, many immunocytochemical techniques have been developed to investigate in detail the interaction between infectious micro-organisms and the nonspecific and specific immune systems of the host. In this review three techniques have been described with all technical details necessary to perform them correctly: (1) single immunocytochemical staining to detect the infectious micro-organisms in situ at their site of infection, (2) double immunocytochemical staining to visualize simultaneously the infectious micro-organism and the host cellular response to investigate their interactions, and (3) detection of plasma cells producing antibodies specific to the micro-organism. Of the three techniques the results are described when applied on chicken tissues infected with various micro organisms, such as Marek's disease virus, chicken anemia virus, infectious bursal disease virus and Eimeria tenella. PMID- 10717285 TI - Duck immune responses to Riemerella anatipestifer vaccines. AB - Riemerella anatipestifer (Ra) infection is probably the most economically important infectious disease of farm ducks worldwide but the immune responses to natural infection and vaccines are poorly understood. We have used the lymphocyte transformation test (LTT) to study the expression of cell-mediated immunity (CMI), and the enzyme-linked immunosorbent assay to monitor antibody (Ab) production following administration of formalin-inactivated and live attenuated serotype 2 (= G) Ra vaccines. Lymphocytes (8x10(5) in 200 microl of RPMI + 10% duck serum, in 96 well trays) were stimulated with Ra antigen, prepared by freeze thaw and sonication; optimum responses were obtained with antigen at 6.25 microg/ml. Cells were cultured for 3 days at 41.6 degrees C/5% CO(2), prior to assessing 3H-thymidine uptake. Ra bacterin, incorporating aluminium hydroxide as adjuvant, stimulated strong but transient (about 4 weeks) LTT response; there was some cross-reaction of the LTT to proteins derived from other serotypes of Ra. Revaccination stimulated slightly stronger responses with the same time course. The Ab response to each vaccination was longer-lived than the LTT response. Vaccination with a live, attenuated strain of Ra stimulated weaker but longer lasting LTT responses, but similar Ab responses compared to the bacterin. It is apparent, therefore, that the transient protection reported using Ra bacterins is due to the fact that the CMI response to these vaccines is transient; and that it is possible for ducks to have detectable levels of serum Ab at times when CMI is not detectable by LTT. These observations are important in terms of our understanding of immunopathogenesis, immunoprophylaxis, and immunodiagnosis in Ra. PMID- 10717286 TI - Avian immune responses to Mycobacterium avium: the wildfowl example. AB - Immunological responses of wildfowl (Order Anseriformes: ducks, geese, swans and screamers) to mycobacteria have been investigated as part of studies to develop a vaccine and diagnostic assay for avian tuberculosis. 10(9) killed Mycobacterium vaccae protected the Cairinini (perching ducks) from avian tuberculosis (p<0.02) but did not achieve statistically significant protection in the other taxonomic tribes. The Cairinini includes the threatened, yet highly susceptible, white winged duck (Cairina scutulata).Together, loss of cell-mediated responses to common mycobacterial antigens, increased responsiveness to the species specific antigens of M. avium, and increased antibody production are reminiscent of the T(H1) to T(H2) shift seen in mammalian mycobacterial infections. It is speculated that excessive exposure to environmental mycobacteria prior to vaccination is detrimental and common antigens play an important role in wildfowl immunity to mycobacteria. A new vaccination trial using killed M. vaccae is being undertaken. Antibody responses are a useful ante mortem diagnostic indicator in most taxonomic tribes with the exception of the primitive Dendrocygnini (whistling ducks). PMID- 10717287 TI - Cytotoxic T lymphocytes are critical in the control of infectious bronchitis virus in poultry. AB - Various strains of infectious bronchitis virus (IBV) cause respiratory, kidney, enteric and reproductive illnesses in chickens, especially in newly hatched chicks. Assays have been developed to identify Gray strain IBV-specific cytotoxic T lymphocyte (CTL) responses using viral infected antigen presenting cells (APC) and using the Semliki Forest virus vector infected APC expressing individual viral polypeptides. It was shown that major histocompatibility complex restricted CTL are responsible for early control of IBV infection. The kinetics of viral load observed in the lungs and kidneys correlated with the level of IBV-specific CTL activity of effector cells prepared from spleens of infected chicks. Adoptive transfer of immune T cells to chicks prior to infection demonstrated that IBV primed CD8(+), alphabeta T lymphocytes could protect chicks from acute infection. CTL determinants in the viral particle can be mapped to the spike and nucleocapsid proteins but not to the membrane protein. The carboxyl terminus of the nucleocapsid protein houses an epitope(s) responsible for induction of CTL responses to IBV N protein. Inoculation of DNA plasmid expressing the carboxyl terminus of Gray strain N resulted in induction of CTL that cross-react with two distinct IBV strains. In addition, this potential DNA vaccine resulted in protection of chicks against acute infection. PMID- 10717288 TI - Specific and nonspecific immune responses to Marek's disease virus. AB - Marek's disease (MD) virus (MDV) has provided an important model to study immune responses against a lymphoma-inducing herpesvirus in its natural host. Infection in chickens starts with a lytic infection in B cells, followed by a latent infection in T cells and, in susceptible birds, T cell lymphomas develop. Non specific and specific immune responses are important for the control of virus infection and subsequent tumor development. Interferon-gamma and nitric oxide are important for the control of virus replication during the lytic phase of infection and are also important to prevent reactivation of MDV replication in latently infected and transformed cells. Cytotoxic T cells (CTLs) are the most important of the specific immune responses in MDV. In addition to antigen specific CTL against MDV proteins pp38, glycoprotein B (gB), Meq, and ICP4, ICP27 specific CTL can also be detected as early as 6 to 7 days post infection. The epitope for gB recognized by CTLs from P2a (MHC: B(19)B(19)) chickens has been localized to the Eco47III-BamHI (nucleotides 1515-1800) fragment. A proposed model for the interactions of cytokines and immune responses as part of the pathogenesis of MD is discussed. PMID- 10717289 TI - Infectious bursal disease virus of chickens: pathogenesis and immunosuppression. AB - Infectious bursal disease virus (IBDV) is an important immunosuppressive virus of chickens. The virus is ubiquitous and, under natural conditions, chickens acquire infection by the oral route. IgM+ cells serve as targets for the virus. The most extensive virus replication takes place in the bursa of Fabricius. The acute phase of the disease lasts for about 7-10 days. Within this phase, bursal follicles are depleted of B cells and the bursa becomes atrophic. Abundant viral antigen can be detected in the bursal follicles and other peripheral lymphoid organs such as the cecal tonsils and spleen. CD4(+) and CD8(+) T cells accumulate at and near the site of virus replication. The virus-induced bursal T cells are activated, exhibit upregulation of cytokine genes, proliferate in response to in vitro stimulation with IBDV and have suppressive properties. Chickens may die during the acute phase of the disease although IBDV induced mortality is highly variable and depends, among other factors, upon the virulence of the virus strain. Chickens that survive the acute disease clear the virus and recover from its pathologic effects. Bursal follicles are repopulated with IgM(+) B cells. Clinical and subclinical infection with IBDV may cause immunosuppression. Both humoral and cellular immune responses are compromised. Inhibition of the humoral immunity is attributed to the destruction of immunoglobulin-producing cells by the virus. Other mechanisms such as altered antigen-presenting and helper T cell functions may also be involved. Infection with IBDV causes a transient inhibition of the in vitro proliferative response of T cells to mitogens. This inhibition is mediated by macrophages which are activated in virus-exposed chickens and exhibit a marked enhancement of expression of a number of cytokine genes. We speculate that T cell cytokines such as interferon (IFN)-gamma may stimulate macrophages to produce nitric oxide (NO) and other cytokines with anti-proliferative activity. Additional studies are needed to identify the possible direct immunosuppressive effect of IBDV on T cells and their functions. Studies are also needed to examine effects of the virus on innate immunity. Earlier data indicate that the virus did not affect normal natural killer (NK) cell levels in chickens. PMID- 10717290 TI - Immunopathogenesis of haemorrhagic enteritis virus (HEV) in turkeys. AB - Infection of turkeys with the haemorrhagic enteritis virus (HEV), a type II avian adenovirus, results in varying rates of morbidity and mortality. The disease is characterised by splenomegaly, intestinal haemorrhage, sudden death and immunosuppression. The mechanisms of HEV immunopathogenesis and immunosuppression are not fully understood. Recent studies indicate that immune responses play a central role in disease pathogenesis. HEV infects B cells and macrophages and induces necrosis as well as apoptosis in infected and possibly in by-stander cells. The ability of the infected birds to mount an optimum humoral immune response as well as normal macrophage functions such as phagocytosis may be impaired. Elevated numbers of splenic CD4(+) cells during the acute phase of infection may be associated with viral clearance. Types I and II interferons (IFN) and pro-inflammatory cytokines such as interleukin-6 and tumour necrosis like factors (TNF) are released at the peak of the infection. Cytokines may play a protective as well as a destructive role. While a massive release of proinflammatory cytokines may lead to systemic shock associated with haemorrhagic enteritis and death, release of IFNs may protect turkeys from the disease. Treatment with thalidomide, which is a potent TNF down-regulatory drug, prevented HEV-induced intestinal haemorrhage and treatment with an IFN-inducing chemical prevented HEV-replication and inhibited HEV-induced pathological and histopathological lesions. PMID- 10717291 TI - Immunopathogenesis of chicken anemia virus infection. AB - The immunopathogenesis of chicken anemia virus (CAV) infection is reviewed. The virus causes a disease in young chicks which is characterised by generalised lymphoid atrophy, increased mortality and severe anemia. The virus appears to target erythroid and lymphoid progenitor cells in the bone marrow and thymus respectively. The B cells in the chicken are not susceptible to CAV infection and are not directly affected by the virus. Destruction of erythroid progenitors in bone marrow results in severe anemia, and depletion of granulocytes and thrombocytes. Destruction of precursor T cells results in depletion of mature cytotoxic and helper T cells with consequent effects on susceptibility to, and enhancement of, the pathogenicity of secondary infectious agents, and sub-optimal antibody responses. Apoptosis appears to be a feature of the lymphocyte depletion in the thymic cortex, which may be mediated by one of the non-structural viral proteins, VP3 (apoptin). PMID- 10717292 TI - The avian response to Newcastle disease virus. AB - Newcastle disease virus (NDV) is classified as a member of the superfamily Mononegavirales in the family Paramyxoviridae. This virus family is divided into two subfamilies, the Paramyxovirinae and the Pneumovirinae. In 1993 the International Committee on the Taxonomy of Viruses rearranged the order of the Paramyxovirus genus and placed NDV within the Rubulavirus genus among the Paramyxovirinae. The enveloped virus has a negative sense single-stranded RNA genome of 15,186 kb which codes for an RNA directed RNA polymerase, hemagglutinin neuraminidase protein, fusion protein, matrix protein, phosphoprotein and nucleoprotein in the 5' to 3' direction. The virus has a wide host range with most orders of birds reported to have been infected by NDV. Isolates are characterized by virulence in chickens and are categorized into three main pathotypes depending on severity of disease. Lentogenic isolates are of low virulence while viruses of intermediate virulence are termed mesogenic. Highly virulent viruses that cause high mortality in birds are termed neurotropic or viscerotropic velogenic. Velogenic NDV are List A pathogens that require reporting to the Office of International Epizootics and outbreaks result in strict trade embargoes. The primary molecular determinant for NDV pathogenicity is the fusion protein cleavage site amino acid sequence. Vaccination for NDV is primarily by mass application of live-virus vaccines among commercial poultry. Although protection is measured by presence of antibodies to NDV, vaccinated B cell depleted chickens are resistant to disease. Consequently, immune protection involves responses that are presently incompletely defined. PMID- 10717293 TI - Immunology of avian influenza virus: a review. AB - Avian influenza virus can cause serious disease in a wide variety of birds and mammals, but its natural host range is in wild ducks, gulls, and shorebirds. Infections in poultry can be inapparent or cause respiratory disease, decreases in production, or a rapidly fatal systemic disease known as highly pathogenic avian influenza (HPAI). For the protection of poultry, neutralizing antibody to the hemagglutinin and neuraminidase proteins provide the primary protection against disease. A variety of vaccines elicit neutralizing antibody, including killed whole virus vaccines and fowl-pox recombinant vaccines. Antigenic drift of influenza viruses appears to be less important in causing vaccine failures in poultry as compared to humans. The cytotoxic T lymphocyte response can reduce viral shedding in mildly pathogenic avian influenza viruses, but provides questionable protection against HPAI. Influenza viruses can directly affect the immune response of infected birds, and the role of the Mx gene, interferons, and other cytokines in protection from disease remains unknown. PMID- 10717294 TI - Immune responses to duck hepatitis B virus infection. AB - The duck hepatitis B virus (DHBV) was the first hepatitis B virus identified from an avian host. It is a member of the Hepadnaviridae family of viruses. All members of this family display similar genomic organization and replication strategies and cause species-specific infections that result in either transient (acute) or persistent infection. Hepadnavirus infection occurs primarily in hepatocytes in the liver with release of infectious virions and non-infectious 'empty' surface antigen particles into the bloodstream. Hepadnavirus replication is non-cytopathic and immune responses to viral antigens are thought to be responsible for the liver damage seen in both transient and persistent infection and for the clearance of virus from infected cells. This has provided the basis for the use of vaccines and prophylactic treatments for individuals at high risk of human hepatitis B virus (HBV) infection. It follows that detailed understanding of the immune responses induced during transient and persistent infection may well facilitate the development of more effective approaches to immunotherapy in patients with persistent infection and may also provide a means of reducing the liver damage associated with this infection, without reducing the effectiveness of the immunity required to eliminate the virus. Immune responses to hepadnavirus infection have been studied primarily in humans, following natural infection with HBV, but studies have also been performed with the woodchuck hepatitis virus (WHV) and the DHBV models. This manuscript reviews the recent studies of immune responses to DHBV infection. PMID- 10717295 TI - Intestinal immune responses to coccidiosis. AB - Intestinal parasitism is a major stress factor leading to malnutrition and lowered performance and production efficiency of livestock and poultry. Coccidiosis is an intestinal infection caused by intracellular protozoan parasites belonging to several different species of Eimeria. Infection with coccidia parasites seriously impairs the growth and feed utilization of chickens and costs the US poultry industry more than $1.5 billion in annual losses. Although acquired immunity to Eimeria develops following natural infection, due to the complex life cycle and intricate host immune response to Eimeria, vaccine development has been difficult and a better understanding of the basic immunobiology of pertinent host-parasite interactions is necessary for developing effective immunological control strategies against coccidiosis. Chickens infected with Eimeria produce parasite specific antibodies in both the circulation and mucosal secretions but humoral immunity plays only a minor role in protection against this disease. Rather, recent evidence implicates cell-mediated immunity as the major factor conferring resistance to coccidiosis. This review will summarize current understanding of the avian intestinal immune system and its response to Eimeria as well as provide a conceptual overview of the complex molecular and cellular events involved in intestinal immunity to coccidiosis. It is anticipated that increased knowledge of the interaction between parasites and host immunity will stimulate the birth of novel immunological and molecular biological concepts in the control of intestinal parasitism. PMID- 10717296 TI - Immunity, vaccination and the avian intestinal tract. AB - Defence of the intestinal mucosal surface from enteric pathogens is initially mediated by secretory IgA (SIgA). As oral immunization of non-replicating antigen induces minimal SIgA antibody titers, novel immunization strategies which selectively induce mucosal immune responses in mammals are now being assessed in chickens. The strategies reviewed include the route of antigen delivery, the incorporation of antigenic components in delivery vehicles, the inclusion of immunomodulators in the vaccine formula or in the diet, and manipulation of intestinal microflora. The differences in anatomical organization and immunological mechanisms between birds and mammals must be considered when manipulating avian intestinal immunity with the latest immunotechnologies developed for mammals. Our knowledge of the function and functioning of the avian mucosal system is discussed. Progress in our understanding of this system, the location of precursor IgA B cells and antigen sampling by these sites is not as advanced as knowledge of the mammalian system, highlighting the need for ongoing research into the avian application of novel vaccination strategies. PMID- 10717297 TI - Delivery of avian cytokines by adenovirus vectors. AB - A fowl adenovirus serotype 8 (FAV-8) recombinant was constructed by inserting an expression cassette consisting of the FAV major late promoter/splice leader sequences (MLP/SL), the chicken interferon-gamma (ChIFN-gamma) gene and SV40 polyA into sites in the right hand end of the FAV-8 genome. One recombinant (A3 13) was constructed by an insertion of ChIFN-gamma into a 1.3 kilobase pair (kbp) deletion which removed a putative open reading frame (ORF) with identity to the CELO (FAV serotype 1) 36 kDa homologue. A second recombinant (S4) removed a further 0.9 kbp and a third recombinant (AA1) was constructed in a small 50 base pair (bp) SpeI deletion. The recombinants displayed differing growth characteristics in CK monolayers. A3-13 grew slowly and only attained a titre of 10(5) pfu/ml, S4 had intermediate growth and AA1 showed wild type growth kinetics. These differing growth properties indicated that removal of the 36 kDa homologue had an effect on growth in vitro. Supernatants from CK monolayers infected with the recombinant virus were assayed for the production of ChIFN gamma. Detectable levels of ChIFN-gamma were observed in supernatants as early as 24 h post infection (p.i.), peaked at 48 h p.i. and this level was maintained for at least 10 days. The level of production of ChIFN-gamma correlated with each recombinant's growth characteristics in vitro. Chickens treated with rFAV-ChIFN gamma showed increased weight gains compared to controls and suffered reduced weight loss when challenged with the coccidial parasite Eimeria acervulina. PMID- 10717298 TI - Avian cytokines - the natural approach to therapeutics. AB - While the effective use of antibiotics for the control of human disease has saved countless lives and has increased life expectancy over the past few decades, there are concerns arising from their usage in livestock. The use of antibiotic feed additives in food production animals has been linked to the emergence in the food chain of multiple drug-resistant bacteria that appear impervious to even the most powerful antimicrobial agents. Furthermore, the use of chemical antimicrobials has led to concerns involving environmental contamination and unwanted residues in food products. The imminent banning of antibiotic usage in livestock feed has intensified the search for environmentally-friendly alternative methods to control disease. Cytokines, as natural mediators and regulators of the immune response, offer exciting new alternatives to conventional chemical-based therapeutics. The utilisation of cytokines is becoming more feasible, particularly in poultry, with the recent cloning of a number of avian cytokine genes. Chickens offer an attractive small animal model system with which to study the effectiveness of cytokine therapy in the control of disease in intensive livestock. In this report we will review the status of avian cytokines and focus on our recent studies involving the therapeutic potential of chicken interferon gamma (ChIFN-gamma) as a vaccine adjuvant and a growth promoter. PMID- 10717299 TI - Preliminary profile of the Cryptosporidium parvum genome: an expressed sequence tag and genome survey sequence analysis. AB - Cryptosporidium parvum is a protozoan enteropathogen that infects humans and animals and causes a pronounced diarrheal disease that can be life-threatening in immunocompromised hosts. No specific chemo- or immunotherapies exist to treat cryptosporidiosis and little molecular information is available to guide development of such therapies. To accelerate gene discovery and identify genes encoding potential drug and vaccine targets we constructed sporozoite cDNA and genomic DNA sequencing libraries from the Iowa isolate of C. parvum and determined approximately 2000 sequence tags by single-pass sequencing of random clones. Together, the 567 expressed sequence tags (ESTs) and 1507 genome survey sequences (GSSs) totaled one megabase (1 mb) of unique genomic sequence indicating that approximately 10% of the 10.4 mb C. parvum genome has been sequence tagged in this gene discovery expedition. The tags were used to search the public nucleic acid and protein databases via BLAST analyses, and 180 ESTs (32%) and 277 GSSs (18%) exhibited similarity with database sequences at smallest sum probabilities P(N)< or =10(-8). Some tags encoded proteins with clear therapeutic potential including S-adenosylhomocysteine hydrolase, histone deacetylase, polyketide/fatty-acid synthases, various cyclophilins, thrombospondin-related cysteine-rich protein and ATP-binding-cassette transporters. Several anonymous ESTs encoded proteins predicted to contain signal peptides or multiple transmembrane spanning segments suggesting they were destined for membrane-bound compartments, the cell surface or extracellular secretion. One-hundred four simple sequence repeats were identified within the nonredundant sequence tag collection with (TAA)(> or =6)/(TTA)(> or =6) and (TA)(> or = 10)/(AT)(> or =10 ) being the most prevalent, occurring 40 and 15 times, respectively. Various cellular RNAs and their genes were also identified including the small and large ribosomal RNAs, five tRNAs, the U2 small nuclear RNA, and the small and large virus-like, double-stranded RNAs. This investigation has demonstrated that survey sequencing is an efficient procedure for gene discovery and genome characterization and has identified and sequence tagged many C. parvum genes encoding potential therapeutic targets. PMID- 10717300 TI - Molecular characterization of a thrombospondin-related anonymous protein homologue in Neospora caninum. AB - Thrombospondin-related anonymous protein (TRAP) family members participate in attachment and invasion of host cells by apicomplexan parasites. A TRAP homologue in Neospora caninum strain Nc-1 (NcMIC2) was cloned, sequenced and found to be 61% identical (75% similar) at the amino acid level to Toxoplasma gondii MIC2 (TgMIC2). Similar to TgMIC2, the predicted amino acid sequence of NcMIC2 contains one integrin-like domain (I or A domain), five thrombospondin (TSP) repeats, a putative transmembrane spanning region and intracellular C-terminus, and was localized to micronemes by cryo-immunoelectron microscopy. The secretion of NcMIC2 was temperature dependent and was induced at or above 25 degrees C. The secreted form of NcMIC2 released into the medium was found to be proteolytically processed such that it lacked the C-terminal domain. Secretion of NcMIC2 was regulated by calcium, since several agents which raise intracellular calcium levels were shown to promote NcMIC2 secretion and chelation of [Ca(2+)](i) abrogated release. As a member of the growing family of apicomplexan TRAP proteins, NcMIC2 may play an important role in attachment and invasion by N. caninum into host cells. PMID- 10717301 TI - Cyclin H activation and drug susceptibility of the Pfmrk cyclin dependent protein kinase from Plasmodium falciparum. AB - The eukaryotic cell cycle is regulated by a group of highly conserved cyclin dependent protein kinases (CDKs). Several CDKs have been identified in Plasmodium falciparum, however, their regulatory mechanisms as well as their role in parasite growth and differentiation are not understood fully. To further our understanding of Plasmodium CDK regulation, we have characterized Pfmrk kinase activity. Pfmrk was expressed and purified as a 6xHis tagged recombinant protein from Escherichia coli and assayed for histone H1 kinase activity. Pfmrk has significant histone H1 kinase activity and is autophosphorylated in vitro. Human cyclin H forms a stable complex with Pfmrk and stimulates kinase activity. This is the first indication that Plasmodial CDKs are partially regulated by cyclin subunits, as are human CDKs. CDKs are attractive drug targets due to their role in cellular proliferation. Specific CDK inhibitors were selected to evaluate Pfmrk as a potential drug target. Olomoucine and roscovitine failed to inhibit Pfmrk kinase activity which places Pfmrk with a class of CDKs that are insensitive to these compounds. A molecular model of Pfmrk provides a structural explanation for the failure of these compounds to inhibit Pfmrk. PMID- 10717302 TI - Overexpression of miniexon gene decreases virulence of Leishmania major in BALB/c mice in vivo. AB - During the construction of a physical map for Leishmania major (LV39) chromosome 2 we have rescued and characterized a L. major (LV39) derived genomic clone bearing solely as insert a long stretch of the miniexon gene array. The recombinant was devised as a tool to study the effect of miniexon overexpression on virulence and growth advantage. Such clone, 32D05, contains approximately 40 kb of the miniexon tandem array. We have examined the course of infection in susceptible BALB/c mice inoculated with transfectants carrying 32D05 as an episome. The study was carried out in two different clonal lines of L. major: virulent line LV39 (clone 5) and avirulent LT252 (CC1 clone). The results presented here indicate that high levels of miniexon expression affect negatively the ability of once virulent lines to induce lesions when injected in susceptible mice. PMID- 10717303 TI - The Wuchereria bancrofti orthologue of Brugia malayi SXP1 and the diagnosis of bancroftian filariasis. AB - The gene encoding the Wuchereria bancrofti orthologue of the Brugia malayi derived diagnostic antigen SXP1 was identified from a W. bancrofti L3 cDNA library and characterized. The Wb-sxp-1 cDNA encoded a basic protein with a calculated molecular mass of 20.8 kDa. Wb-SXP-1 was 85% identical to the SXP1 protein described from B. malayi (Bm-SXP-1). The Wb-SXP-1 sequence also showed significant identity with proteins described from B. pahangi, Onchocerca volvulus, Acanthochilonema vitea, Ascaris suum, Loa loa, Litomosoides sigmodontis and Caenorhabditis elegans. The presence of a number of invariant and conserved residues in all of these nematode-derived molecules suggests that Wb-SXP-1 is a member of a new protein family. A recombinant form of Wb-SXP-1 was produced and it was determined that the anti-Wb-SXP-1 antibody response in patients with W. bancrofti infections was restricted to the IgG4 subclass. An anti-Wb-SXP-1 IgG4 ELISA was developed and this assay was found to be 100% sensitive for patients with patent W. bancrofti infection. Sera from individuals experiencing chronic pathology, endemic normals or patients with non-filarial nematode infections had no detectable IgG4 against Wb-SXP-1. While patients with patent Onchocerca volvulus infections were uniformly negative in the Wb-SXP-1 assay, 40% of sera from patent Loa loa infections were positive. When Bm-SXP-1 was used as the antigen under identical conditions, the assay was 88% specific for patent W. bancrofti infections and the antigen was recognized by antibodies from both O. volvulus and L. loa infections. The results strongly suggested that, for certain diagnostic filarial antigens, the use of same-species molecules can enhance the specificity of diagnostic tests. PMID- 10717304 TI - Inhibition of gene expression in Entamoeba by the transcription of antisense RNA: effect of 5' and 3' regulatory elements. AB - Down regulation of gene expression by antisense RNA is one of the ways to investigate the specific contribution of certain components to the physiology and activities of a cell. A successful inhibition of gene expression in Entamoeba trophozoites was achieved in stable transfectants by using hybrid plasmid constructs containing promotors that produce transcripts which do not bind to polysomes. Different promotors were found to be required for Entamoeba histolytica or Entamoeba dispar. In E. histolytica one of the two copies (g34) of the gene coding for ribosomal protein L21 was previously found to be transcribed but not translated. Inhibition of gene expression was obtained by placing in a transfection vector, the amoebapore A gene, in its antisense orientation, under the control of the g34 promotor. Transfectants of E. histolytica were shown to accumulate antisense transcripts and inhibit amoebapore synthesis. In contrast, transfectants with plasmid constructs in which the amoebapore gene was placed under the control of the gLE3 promotor of RP-L21, which is known to be translated, did not accumulate antisense transcript or inhibit gene expression. Maximal inhibition of amoebapore expression was obtained when the antisense construct also included the 5' and 3' untranslated regions of the amoebapore gene. In E. dispar the opposite situation was found, plasmid constructs containing the promotor regions of the gLE3 copy, which were shown to be poorly translated, were more efficient in inhibiting the synthesis of a 30 kDa surface specific antigen than a construct with the g34 promotor element. PMID- 10717305 TI - A microneme protein from Eimeria tenella with homology to the Apple domains of coagulation factor XI and plasma pre-kallikrein. AB - Microneme organelles are present in all apicomplexan protozoa and contain proteins that are critical for parasite motility and host cell invasion. One apicomplexan-wide family of microneme proteins has been identified with members that are characterised by the possession of thrombospondin type I repeats, conserved adhesive motifs which are implicated in binding to glycosaminoglycan chains. In this paper we describe a micronemal glycoprotein, EtMIC 5, from Eimeria tenella which contains eleven cysteine-rich motifs that have striking similarity to the adhesive Apple (A-) domains of blood coagulation factor XI and plasma pre-kallikrein. EtMIC 5 is confined to an intracellular location in resting sporozoites but is translocated to the parasite surface and secreted into the culture supernatant during parasite infection of MDBK cells. During intracellular replication, the protein is switched off in early schizogony and is then re-expressed within the apical tips of newly formed merozoites. A-domain sequences were also found in microneme proteins from Sarcocystis muris and Toxoplasma gondii and in a protein of unknown localisation from Eimeria acervulina. These studies suggest that A-domain containing proteins may comprise a novel apicomplexan-wide family of microneme adhesins. PMID- 10717306 TI - Genomic distribution and functional characterisation of two distinct and conserved Plasmodium falciparum var gene 5' flanking sequences. AB - Approximately 50 highly diverse var genes distributed throughout the haploid genome of the malaria parasite Plasmodium falciparum code for PfEMP1 variants located on the surface of infected erythrocytes. PfEMP1 is involved in cytoadherence of parasitised red blood cells and undergoes antigenic variation through differential expression of var genes. Members of the var gene family are located in chromosome-internal positions on chromosomes 4, 7, 8 and 12, and in subtelomeric regions of all chromosomes. Here we show that there are two distinct and conserved types of 5' upstream regions (var17-type and 5B1-type) of var genes, and suggest that most subtelomeric var genes are flanked by a var17-type 5' upstream sequence. In contrast, 5B1-type 5' upstream are localised to chromosomes that have been shown to contain var genes within chromosome-internal regions. Transcriptional analysis using RT-PCR revealed that var genes flanked by either type of 5' upstream sequence are transcribed in in vitro cultured trophozoite stage parasites. In addition, we have shown that the 5' flanking sequences of four different var genes are able to drive transient expression of the cat reporter gene. Our results suggest that at least the minimal regulatory sequences required for transcription of var genes are conserved among both subgroups of the var gene family. Furthermore, these sequences provide new markers for the investigation of the chromosomal organisation of var genes. PMID- 10717307 TI - A family of galectins from Haemonchus contortus. PMID- 10717308 TI - Cross-species homologous recombination in Leishmania donovani reveals the sites of integration. PMID- 10717309 TI - Cloning and biochemical characterization of genes encoding two isozymes of cysteine synthase from Entamoeba dispar. PMID- 10717310 TI - Trypanosoma brucei RNase HI requires its divergent spacer subdomain for enzymatic function and its conserved RNA binding motif for nuclear localization. PMID- 10717311 TI - Adventures with poxviruses of vertebrates. AB - Because they were the largest of all viruses and could be visualised with a light microscope, the poxviruses were the first viruses to be intensively studied in the laboratory. It was clear from an early date that they caused important diseases of humans and their domestic animals, such as smallpox, cowpox, camelpox, sheeppox, fowlpox and goatpox. This essay recounts some of the early history of their recognition and classification and then expands on aspects of research on poxviruses in which the author has been involved. Studies on the best known genus, Orthopoxvirus, relate to the use of infectious ectromelia of mice as a model for smallpox, embracing both experimental epidemiology and pathogenesis, studies on the genetics of vaccinia virus and the problem of non-genetic reactivation (previously termed 'transformation') and the campaign for the global eradication of smallpox. The other group of poxviruses described here, the genus Leporipoxvirus, came to prominence when the myxoma virus was used for the biological control of Australian wild rabbits. This provided a unique natural experiment on the coevolution of a virus and its host. Future research will include further studies of the many immunomodulatory genes found in all poxviruses of vertebrates, since these provide clues about the workings of the immune system and how viruses have evolved to evade it. Some of the many recombinant poxvirus constructs currently being studied may come into use as vaccines or for immunocontraception. A field that warrants study but will probably remain neglected is the natural history of skunkpox, raccoonpox, taterapox, yabapox, tanapox and other little-known poxviruses. A dismal prospect is the possible use of smallpox virus for bioterrorism. PMID- 10717312 TI - Riding the sulfur cycle--metabolism of sulfonates and sulfate esters in gram negative bacteria. AB - Sulfonates and sulfate esters are widespread in nature, and make up over 95% of the sulfur content of most aerobic soils. Many microorganisms can use sulfonates and sulfate esters as a source of sulfur for growth, even when they are unable to metabolize the carbon skeleton of the compounds. In these organisms, expression of sulfatases and sulfonatases is repressed in the presence of sulfate, in a process mediated by the LysR-type regulator protein CysB, and the corresponding genes therefore constitute an extension of the cys regulon. Additional regulator proteins required for sulfonate desulfonation have been identified in Escherichia coli (the Cbl protein) and Pseudomonas putida (the AsfR protein). Desulfonation of aromatic and aliphatic sulfonates as sulfur sources by aerobic bacteria is oxygen-dependent, carried out by the alpha-ketoglutarate-dependent taurine dioxygenase, or by one of several FMNH(2)-dependent monooxygenases. Desulfurization of condensed thiophenes is also FMNH(2)-dependent, both in the rhodococci and in two Gram-negative species. Bacterial utilization of aromatic sulfate esters is catalyzed by arylsulfatases, most of which are related to human lysosomal sulfatases and contain an active-site formylglycine group that is generated post-translationally. Sulfate-regulated alkylsulfatases, by contrast, are less well characterized. Our increasing knowledge of the sulfur-regulated metabolism of organosulfur compounds suggests applications in practical fields such as biodesulfurization, bioremediation, and optimization of crop sulfur nutrition. PMID- 10717313 TI - Signal transduction mechanisms in Caulobacter crescentus development and cell cycle control. AB - The life cycle of the aquatic bacterium Caulobacter crescentus includes an asymmetric cell division and an obligate cell differentiation. Each cell division gives rise to a motile but replication inert swarmer cell and a sessile, replication competent stalked cell. While the stalked progeny immediately reinitiates DNA replication and cell division, the swarmer cell remains motile and chemotactically active for a constant period of the cell cycle before it differentiates into a stalked cell. During this process, the cell looses motility by ejecting the flagellum, synthesizes a stalk and eventually initiates chromosome replication and cell division. The link of morphogenic transitions to the replicative cycle of Caulobacter implies that the developmental programs which determine asymmetry and cell differentiation must be tightly connected with cell cycle control. This has been confirmed by the recent identification of signal transduction mechanisms, which are involved in temporal and spatial control of both development and cell cycle. Interestingly, the cell has recruited two-component signal transduction systems for this internal control, a family of regulatory proteins which usually are involved in the information transfer between the environment and the inside of a cell. The response regulator protein CtrA controls several key cell cycle events like the initiation of DNA replication, DNA methylation, cell division, and flagellar biogenesis. The activity of this master regulator is subject to complex temporal and spatial control during the C. crescentus cell cycle, including regulated transcription, phosphorylation and degradation. Three membrane bound sensor kinases have been proposed to control the phosphorylation status of CtrA. Two of these, CckA and DivJ, exhibit specific subcellular localization and, in the case of CckA, dynamic rearrangement in the course of the cell cycle. These findings support the idea that the developmental program of C. crescentus is controlled at least in part by localized cues that act as checkpoints for the control of morphological changes and cell cycle progression. PMID- 10717314 TI - Characterisation of bacteria by matrix-assisted laser desorption/ionisation and electrospray mass spectrometry. AB - Chemical analysis for the characterisation of micro-organisms is rapidly evolving, after the recent advent of new ionisation methods in mass spectrometry (MS): electrospray (ES) and matrix-assisted laser desorption/ionisation (MALDI). These methods allow quick characterisation of micro-organisms, either directly or after minimum sample preparation. This review provides a brief introduction to ES and MALDI MS and a discussion of micro-organism characterisation capabilities. Some attention is devoted to the analysis of mixtures of proteins, lipids and other compounds, to the combination of polymerase chain reaction technology and MS, and to the analysis of whole bacteria and their lysates. The review of results produced hitherto is concluded with an outlook on future developments. PMID- 10717315 TI - Purification and partial characterisation of geranylgeranyl diphosphate synthase, from Taxus baccata cell cultures. An enzyme that regulates taxane biosynthesis. AB - Geranylgeranyl diphosphate (GGPP) synthase, an enzyme that regulates taxane biosynthesis, was purified to homogeneity from cell cultures of Taxus baccata. The molecular weight of the native protein was estimated to be 76+/-2 kDa, resulting from the association of two apparently identical subunits having a molecular weight of 38 kDa. Farnesyl diphosphate (K(m) 2.46 uM) in combination with isopentenyl diphosphate (K(m) 1.5 uM) was the most effective substrate. Dimethyl allyl diphosphate was a poorer substrate (K(m) 12.7 uM). Mn(2+) ion at 4 mM in combination with Mg(2+) of 2 mM gave the greatest stimulation of activity. The pI of the enzyme was lower than 4 and the pH optimum was between 6.9 and 7.2. The enzyme activity was found in the 20000xg (centrifugal force) pellet and a non ionic detergent was used for its extraction. The inclusion of detergent was not necessary during subsequent chromatographic steps. PMID- 10717316 TI - Greenhouse-grown conditionally lethal tobacco plants obtained by expression of plastidic glutamine synthetase antisense RNA may contribute to biological safety. AB - A cDNA corresponding to plastidic glutamine synthetase (GS-2), an enzyme involved in photorespiration, was expressed in antisense orientation under the control of a leaf-specific soybean ribulose-1,5-bisphosphate carboxylase/oxygenase small subunit gene promotor in transgenic tobacco to yield conditionally lethal plants. Three transgenic tobacco lines with decreased (at most 64%) foliar GS-2 activity were obtained. These plants grew normally when maintained in an atmosphere with a CO(2) partial pressure sufficiently high (300 Pa CO(2)) to suppress photorespiration. However, when photorespiration was initiated by the transfer of the plants to air (35 Pa CO(2)), ammonium accumulated in the leaves. With time, the transgenic plants exhibited severe chlorotic lesions and, eventually, the plants died. A stable atmosphere containing at least 300 Pa CO(2) can be established easily in the greenhouse but is unlikely to occur in a natural environment. Therefore, the transgenic tobacco plants with decreased leaf GS-2 activity may contribute to biological safety for production of desired proteins. PMID- 10717317 TI - Subcellular localisation of cherry leaf roll virus coat protein and genomic RNAs in tobacco leaves. AB - The in vivo subcellular location of the coat protein and RNAs of cherry leaf roll nepovirus (CLRV) was studied in infected tobacco plants by two different approaches and it was correlated with the cytopathic structures induced by the virus. Subcellular fractions were obtained by differential centrifugation, visualised by electron microscopy and analysed for their viral RNA and coat protein content by Northern blot and Western blot analysis, respectively. Results indicate that viral RNAs accumulated preferentially at the microsomal fraction. Immunocytochemical studies revealed a clear association of the coat protein of CLRV with the virus-induced cytopathological structures. In situ hybridisation studies confirmed the cytoplasmic location of the virus and allowed one to elucidate the distribution of the CLRV genomic RNAs in the different cell types of infected tissue. PMID- 10717318 TI - Characterization and expression of cold-induced glutathione S-transferase in freezing tolerant Solanum commersonii, sensitive S. tuberosum and their interspecific somatic hybrids. AB - Glutathione S-transferases (GST) form a large family of non-photosynthetic enzymes known to function in detoxification of xenobiotics. We have cloned and characterized a novel, low temperature regulated GST, Solanum commersonii glutathione S-transferase (Scgst1), from a cold acclimated wild potato species S. commersonii and studied the level of its transcription in freezing tolerant and sensitive Solanum genotypes. Active oxygen species (AOS) were associated with the early steps of Scgst1 regulation since a strong mRNA signal was detected in hydrogen peroxide and salicylic acid treated plants. In experimental conditions where the formation of AOS is known to accelerate, such as excessive light at low temperature, significant accumulation of the transcript was observed in S. commersonii. Under similar experimental conditions, Scgst1 transcript did not accumulate in freezing sensitive S. tuberosum eventhough a single copy of the Scgst1 sequence was present in both species. Thus, Scgst1 in the S. tuberosum genome did not exhibit the same cold-induction properties as in S. commersonii. In comparison with the parental lines, the somatic hybrid SH9A (S. commersonii (+) S. tuberosum) had an interparental level of Scgst1 accumulation as well as freezing tolerance. The abundance of Scgst1 transcript thus correlated well with the freezing tolerance of the parental lines and the somatic hybrid SH9A. Increased GST enzyme activity was observed in S. commersonii and SH9A after 2 days of cold acclimation whereas the activity declined in S. tuberosum during the same period. Further studies of potato lines (S1) that were derived by selfing the somatic hybrid revealed a more complex relationship between freezing tolerance and Scgst1 expression level. In the S1 genotypes, the regulation of Scgst1 transcription resembled more that of S. tuberosum and was not directly related to their freezing tolerance. This could be due to the interaction of the two genomes in S1 genotypes as well as chromosomal rearrangements during meiosis. PMID- 10717319 TI - Transgenic carrots with enhanced resistance against two major pathogens, Erysiphe heraclei and Alternaria dauci. AB - In vitro assay indicated that the human lysozyme has lytic activity against phytopathogenic fungi and bacteria. A human lysozyme gene was placed under control of the constitutive CaMV 35S promoter and the resulting expression plasmid was introduced into two cultivars (cv.) of carrot, Kurodagosun (K5) and Nantes Scarlet (NS), by Agrobacterium tumefaciens-mediated method. Seven and fourteen transgenic plants of cv. K5 and cv. NS were regenerated, respectively, and the obtained transgenic carrots of T0 generation was tested for disease resistance against Erysiphe heraclei, a pathogenic fungi causing powdery mildew. Among the tested lines, the transgenic plant No. 12-1 and 8-1 of cv. NS showed a fairly strong resistance against E. heraclei. The strong disease resistance was also confirmed in T1 generation. Disease resistance against another pathogen of leaf blight, Alternaria dauci, were also tested using T1 transgenic lines. Significant enhanced resistance was observed in the No. 12-1 of cv. NS. Accumulation of synthesized human lysozyme protein was observed in this line, a finding consistent with observed disease resistance. PMID- 10717320 TI - Antioxidant responses of cucumber (Cucumis sativus) to photoinhibition and oxidative stress induced by norflurazon under high and low PPFDs. AB - Photooxidative damage is exacerbated by norflurazon (NF), which blocks carotenoid biosynthesis. This study examined the influence of photosynthetic photon flux density (PPFD) on the overall responses of both non-enzymatic and enzymatic antioxidants to NF-caused oxidative damage in leaves of cucumber (Cucumis sativus). Seven-day-old cucumber plants were exposed to NF under either low PPFD (30 umol m(-2) s(-1)) or high PPFD (300 umol m(-2) s(-1)) for 3 days. The NF plants exposed at high PPFD had lower levels of F(v)/F(m) ratio, quantum yield of electron transport, and 33-kDa protein of photosystem II as compared with the NF plants at low PPFD. In the NF plants, there was a reduction in total chlorophylls and carotenoids except newly formed zeaxanthin in either PPFD. The NF plants at high PPFD resulted in less level of photochemical quenching, q(P), and Stern Volmer quenching, NPQ, than those of the plants at low PPFD, whereas both plants had similar level of non-photochemical quenching coefficient, q(N). However, the level of PPFD did not significantly affect the NF-caused induction of antioxidant enzymes including peroxidase, superoxide dismutase, glutathione reductase, and ascorbate peroxidase. PMID- 10717321 TI - Effect of spikelet position on rice anther culture efficiency. AB - The potential of anthers from different parts of the panicle to induce callus was investigated with the japonica rice variety Taipei 309. The results showed that the callusing abilities of anthers from different spikelet positions were significantly different. After plating 4483, 4496, 4348 anthers from the basal, middle and top parts, the percentage of anthers forming calli was 20% in the basal part, 12% in the middle part and 8% in the top part. The anthers of basal parts containing pollen at all uninucleate stages, including early, middle and late, showed higher callus induction frequency than those from middle and top parts. The green plantlet regeneration frequencies of top, middle and basal spikelets were around 18% in all three cases. From the results it would appear that anthers from the basal part of the panicle should be used in anther culture of rice in order to obtain higher efficiencies, and thereby optimise the usefulness of this technique in rice breeding programmes. PMID- 10717322 TI - The ability of pea transformation technology to transfer genes into peas adapted to western Canadian growing conditions. AB - Transgenic pea plants can be produced by Agrobacterium-mediated transformation of thin slices from developing embryo axes. To determine if the method is effective for different pea genotypes, seven pea breeding lines adapted to western Canadian growing conditions were tested, using three different Agrobacterium tumefaciens transformation vectors. All vectors contained the gus (uidA) gene coding for the beta-glucuronidase (GUS) protein, but with different chemical selection genes. In total, 323 transgenic plants were recovered from 39 independent transformation events. Transgenic plants were recovered from each genotype and each selection system, but not from all combinations. GUS-positive explants were obtained from seeds harvested between 24 and 31 days after flowering. The mean time from Agrobacterium treatment to planting into soil averaged 186 days. Based on the initial number of seeds used, the transformation frequency was 0.6% (i.e. six independent transgenic events per 1000 axes sliced). The inserted genes were functional and inherited in a Mendelian fashion. Although more plants were recovered by selection on chlorsulfuron, GUS activity was generally greater in plants selected on kanamycin. GUS activity in the leaves of the original plants varied, but GUS activity in the second generation was correlated with that of the original transformants. PMID- 10717323 TI - Empirical investigations into the tunica structuring point of the shoot apex of Pelargonium zonale. AB - A tunica structuring point of 3.85 cell units was formally predicted for the shoot apex. This point is decisive for the genesis of the tunica corpus structure as such and for the number of tunica layers in an apex at any given moment. Here, a method to determine, in empirical investigations, the theoretical number of tunica layers in real apices is demonstrated. Furthermore, results of an empirical investigation into the postulated tunica structuring point are presented and causes for deviations of the observed values from the theoretical values are discussed. PMID- 10717324 TI - A theoretical approach to the genesis of cell layer arrangements in undifferentiated tissues. AB - Cell layer arrangements in undifferentiated tissues result from tissue-internal stress generated by a high accumulation of undifferentiated cells. The development of the tunica corpus structure in higher plants is used as an example to explain this thesis. The conditions on which this structure will develop are described. The crucial mark from which a tunica layer in an idealized apex will develop is calculated. This tunica structuring point of 3.85 cell units is a natural constant. A mathematical formula is deduced to determine the existing number of tunica layers in a shoot apex at any given moment. In recent years, a number of genes seen in causal connection with the meristem development could be identified. However, the assumption that the genesis of the layered structure of the shoot apex is also genetically fixed can be refuted with this theory. PMID- 10717325 TI - Plant electrophysiology: pentachlorophenol induces fast action potentials in soybean. AB - In the present work, we show that pentachlorophenol (PCP) induces ultra fast action potentials and decreases the variation potential in a soybean. The speed of the propagation of action potentials in a soybean induced by PCP reaches up to 30 m/s, similar to excitation propagation in animal nerves. Measured velocities for transmission of action potentials in green plants reported by different authors were registered in the range of a few mm/s. Action potentials induced in soybeans by PCP are many times faster. Seventy hours after adding PCP to soil variation (resting) potential decreases to zero level. This is the first attempt of high-speed automatic measurements of fast action potentials in green plants. PMID- 10717326 TI - An estimate of the cost of conducting phase II trials in lung cancer. AB - OBJECTIVE: Although it is commonly assumed that clinical trials are more costly than standard therapy, there have been no previous studies of the cost of conducting phase II trials in lung cancer. We retrospectively analyzed two National Cancer Institute of Canada phase II trials in previously untreated small cell lung cancer (SCLC) to determine the costs of conducting the trials in a cancer treatment centre. Both studies were clinical trials undertaken as part of the NCIC's Investigational New Drug program: IND 69 and IND 50 evaluated docetaxel (taxotere) and gemcitabine, respectively. METHODS: data management costs in a Canadian cancer treatment centre were determined from the time estimates provided by data managers to complete various protocol related tasks. Nursing and pharmacy personnel measured the time and supplies necessary to prepare and administer the chemotherapy. Physician fees were determined from the type and number of care visits required by the clinical protocols. Laboratory tests and imaging studies were costed according to the Ontario Health Insurance Plan (OHIP) Schedule of Fees and Benefits. To estimate whether phase II trials are more costly than standard treatment, we determined the cost of four cycles of VP-16-cisplatin, a standard treatment for SCLC. RESULTS: The total cost of performing the docetaxel study was $18443 for an average cost per case of $1317 and an average cost per treatment cycle of $683. The gemcitabine study cost more, due to the fact that the drug proved to be active against SCLC and more cycles of therapy were administered to a larger number of patients. Laboratory and administration costs were also higher, because of the drug administration schedule. The total cost of this study was estimated to be $64670 and the average cost per patient entered was $2230 with an average cost per treatment cycle of $898. In comparison, the estimated cost of four cycles of VP-16-cisplatin chemotherapy was $3948 or $987 per treatment cycle. The major cost drivers in the clinical trials were laboratory and imaging tests which made up 17 and 39%, respectively, of the costs of the taxotere study, and 29 and 27%, respectively, for the gemcitabine study. Data management costs contributed 21 and 13% of the total costs, respectively. CONCLUSION: As the main cost drivers in these phase II clinical trials are laboratory and imaging tests, the cost of clinical trials could potentially be reduced by ensuring that only essential tests are required by protocol. Not surprisingly, the cost of conducting a trial of an active agent is greater than for an inactive agent, because more patients are treated and each patient receives more treatment. The implications for the per-case funding of phase II clinical trials are discussed. PMID- 10717327 TI - Economic evaluation in a randomized phase III clinical trial comparing gemcitabine/cisplatin and etoposide/cisplatin in non-small cell lung cancer. AB - INTRODUCTION: Information on the relative cost-effectiveness of treatments for cancer is being increasingly sought as pressure on health care resources increases. The objective of this study was to assess the cost-effectiveness of gemcitabine/cisplatin (GC) versus cisplatin/etoposide (CE) in patients with advanced non-small cell lung cancer (NSCLC), using resource utilization data collected in conjunction with the first randomized clinical trial comparing both combinations. METHODS: Efficacy and medical care resource utilization data were collected prospectively in an open-label, multicenter, randomized, comparative, phase III trial conducted in Spain which compared gemcitabine/cisplatin and cisplatin/etoposide in 135 chemonaive patients with Stage IIIB or IV NSCLC. There were no differences between both regimens when survival was used as primary end point, so a cost-minimization analysis was used to compare them. In addition, cost-effectiveness analyses were conducted when percentage of responses and time to progression were used as secondary end-points. RESULTS: There were no differences between both regimens when survival was selected as the efficacy end point. Despite the higher chemotherapy cost of GC when compared to CE, there were no differences in total direct costs (584523 pts for GC and 589630 pts for CE; P=NS) between both regimens. Potential savings with GC were mainly associated with a decrease in hospitalization rate. There were differences in favor of GC when response rate (40.6% for GC and 21.9% for CE; P<0.05) and time to disease progression (8.7 months for GC and 7.2 months for CE; P<0. 05) were used as clinical end-points. GC presented a favorable cost-effectiveness profile when compared to CE. CONCLUSIONS: This prospective economic evaluation conducted alongside a clinical trial offers valuable preliminary information on the potential efficiency of the combination gemcitabine-cisplatin in NSCLC. Future assessments based on larger clinical trials focused on survival and naturalistic economic studies conducted in real clinical practice settings are necessary to confirm these findings. PMID- 10717328 TI - A phase II trial of the combination of gemcitabine, ifosfamide and cisplatin in the treatment of advanced non-small cell lung cancer. AB - OBJECTIVE: This phase II trial was designed to assess the efficacy and toxicity profile of the combination of gemcitabine, ifosfamide and cisplatin (GIP) in the treatment of patients with locally advanced or metastatic non-small cell lung cancer (NSCLC). PATIENTS AND METHODS: Patients included in the study were those with surgically unresectable or metastatic NSCLC, with bidimensionally measurable disease, a Karnofsky performance status > 60, and who had not received previous chemotherapy. Treatment consisted of 1000 mg/m(2) gemcitabine on days 1 and 8, 3 g/m(2) ifosfamide on day 1, and 50 mg/m(2) cisplatin on day 1, administered in 21 day cycles. A maximum of six cycles were administered. RESULTS: Between March 1996 and December 1997, 60 patients were included in the study (37 stage III and 23 stage IV), of which 59 were evaluated for response. An objective response was obtained in 43% of patients (3% complete and 40% partial responses), whereas 22% had stable disease. The median survival time for the whole group was 52 weeks (65 weeks in patients with stage III, and 35 weeks in stage IV). The most frequent toxicity was haematological, 56% of patients presented grade 3 or 4 myelotoxicity in one of the cycles, although only seven episodes of febrile neutropenia were recorded in the 255 cycles administered. CONCLUSIONS: The GIP regimen attains response rates similar to those obtained with the gemcitabine plus cisplatin combination used in advanced NSCLC, and had an acceptable toxicity profile. PMID- 10717329 TI - Allelotype and loss of heterozygosity around the L-myc gene locus in primary lung cancers. AB - L-myc S-allele was reported to be associated with metastasis of lung cancer, indicating the existence of a putative tumor suppressor gene around the L-myc locus, in linkage disequilibrium. The relationship between the S-allele and inactivation of some tumor suppressor gene should be indicated by allelic loss. Therefore, we examined the association between the L-myc S-allele and loss of heterozygosity at 11 loci around the L-myc locus (1p34.3) in primary lesions or other biological characteristics in lung cancer. No associations between the S allele and allelic loss around the L-myc locus or other characteristics were found. According to the deletion map, three shortest regions of overlap between D1S230 and D1S76 were identified. While loss of heterozygosity at SRO1, between D1S2797 and MYCL1, showed no relationship with the pathological stage, it was more frequently observed in squamous cell carcinoma than adenocarcinoma (P=0.019), and associated with high telomerase activity (P=0.046), an indicator of cellular immortality. In conclusion, we found three shortest regions of overlap (SROs) from D1S2797 to pter, and a tumor suppressor gene, which might be associated with suppression of lung cancer development but not with L-myc S allele, may exist in SRO1. PMID- 10717330 TI - RARbeta2 specificity in mediating RA inhibition of growth of lung cancer-derived cells. AB - Retinoic acid receptor beta is the retinoid receptor most frequently associated with the growth suppressive effects of retinoic acid in various epithelial tumor derived cell lines. In particular, it has been shown that transfection of RARbeta2 in epidermoid lung tumor cells could reduce their in vitro growth rate in the presence of retinoic acid and in vivo tumorigenicity. However, the question remained as to the isoform specificity of this effect. To investigate this, we transfected RARalpha1, RARbeta1 and RARbeta2 into the epidermoid lung cancer cell line Calu-1 and assessed the in vitro growth capacities of the transfected cells. The expression of the fetal RARbeta1 or overexpression of the ubiquitous RARalpha1 isoforms could not mimick the growth suppressive effect of RARbeta2. In addition we analyzed the expression of another RAR isoform, alpha2, in many tumor-derived lines and conclude from its expression pattern that RARalpha2 is unlikely to be involved in retinoic acid growth suppression of lung cancer. Overall our data suggest that the suppressive effect of RARbeta2 is isoform specific. PMID- 10717331 TI - Cellular immunologic parameters related to age, gender, and stage in lung cancer patients. AB - Several immunologic parameters have been reported to correlate with the clinicopathologic status of lung cancer patients. However, these studies were based on relatively small numbers of patients and often yielded conflicting results. We prospectively studied cellular immunologic parameters related to age, gender, and stage in lung cancer patients. We obtained pretreatment peripheral blood samples from 287 lung cancer patients. Lymphocyte subsets (percentage of lymphocytes positive for CD3, CD4, CD8, HLA-DR, or representing FcgammaR IIIa positive T cells), natural killer (NK) cell activity, and lymphoblastogenesis (LB) after stimulation by phytohemagglutinin (PHA) were evaluated. Significant decline was seen in older patients in percentages of cells positive for CD3 or CD4, in the CD4/CD8 ratio and in LB. The percentage of FcgR IIIa-positive T cells increased with age. LB as well as CD4 positivity were significantly greater in women than in men. NK cell activity showed the greatest cytotoxic responses in stage IIIA, with significantly less response in stage IV than in IIIA. Node negative patients showed higher reactivities for LB and lower positivity for HLA DR than node-positive patients. Patients with no distant metastases had a higher level of NK cell activity than patients with distant metastases. Immune parameters are variously related to age, gender, and the stage in lung cancer patients, some may prove to be useful predictors of survival. PMID- 10717332 TI - Lung cancer risk in relation to genetic polymorphisms of microsomal epoxide hydrolase among African-Americans and Caucasians in Los Angeles County. AB - Microsomal epoxide hydrolase participates in the metabolism of benzo[a]pyrene, an important carcinogen in tobacco smoke. Two relatively common polymorphisms of the microsomal epoxide hydrolase gene that influence enzyme activity have been described. An association between genetic variation in microsomal epoxide hydrolase and lung cancer risk has been reported in one of two studies of Caucasians. We examined the relation between these two polymorphisms and lung cancer risk among 337 incident cases and 700 population controls of African American and Caucasian ethnicity enrolled in a case-control study in Los Angeles County. African-Americans, homozygous for the exon 3 variant allele conferring reduced activity, were at decreased risk of lung cancer (odds ratio (OR)=0.08, 95% CI 0.01-0.62). When data from both the exon 3 and exon 4 polymorphisms were combined into indices of predicted microsomal epoxide hydrolase activity, a decreased risk was seen among African-American subjects with very low predicted activity OR=0.10 (95% CI 0.01-0.83). No comparable association was seen among Caucasians. Although the three published results for Caucasians are somewhat variable, the association among African-Americans in these data provides some support for the hypothesis that genetically reduced microsomal epoxide hydrolase activity may be protective against lung cancer. PMID- 10717333 TI - A randomized phase II trial of two schedules of topotecan for the treatment of advanced stage non-small cell lung cancer. AB - We conducted a randomized phase II trial of two different schedules of topotecan in patients with advanced-stage non small lung cancer (NSCLC) without prior cytotoxic chemotherapy. All patients had histologic or cytologic confirmation of stage IV (M1) or III-B NSCLC. Patients were stratified by performance status, stage and weight loss. Patients were randomized to receive topotecan at intravenous doses of 1.5 mg/m(2)/day over 30 min for 5 days every 3 weeks (Arm A) or 1.3 mg/m(2)grade 3 in both arms included leukopenia, thrombocytopenia, malaise, constipation, diarrhea, lethargy, pulmonary, vomiting, infection and myalgia. Severe (> or = grade 3) thrombocytopenia occurred in 15.8% of Arm A patients and 37.8% of Arm B patients and this difference was statistically significant (P=0.03). The median times to progression are 101 and 63 days (P=0. 75) and the median survival times are 257 and 179 days (P=0.83) for Arms A and B, respectively. These differences in time to progression and overall survival are not statistically significant. Topotecan has limited, single agent activity in advanced NSCLC when given as 1. 5 mg/m(2)/day over 30 min for 5 days every 3 weeks. We do not intend to pursue further investigations with topotecan in patients with NSCLC. PMID- 10717334 TI - Endobronchial metastases secondary to solid tumors: report of eight cases and review of the literature. AB - Endobronchial metastases (EBM) secondaries to extrapulmonary solid malignant tumors are rare. Breast, colon and renal adenocarcinomas are the most frequent tumors associated with EBM. Since 1990 we have treated eight patients with EBM secondary to renal adenocarcinoma (three cases), colon adenocarcinoma (two cases), gastric adenocarcinoma (one case), bladder carcinoma (one case) and basal cell carcinoma (one case). Endobronchial lesions were detected by bronchoscopy and their metastatic nature was confirmed histopathologically in all eight cases. We also conducted a review of EBM reporting studies published in English language. The median interval from the diagnosis of the primary tumour was 41 months. Symptoms and radiological findings were indistinguishable from those of primary lung cancer. Five patients were treated with external radiotherapy with symptomatic improvement while two patients had chemotherapy and one patient underwent surgical resection of the metastasis. Systemic treatment was used in six cases with no significant effect on EBM. Median survival after EBM diagnosis was 9 months with one patient surviving 3.5 years and two patients still alive at 1 year. In conclusion, EBM usually represent a late manifestation requiring differential diagnosis from a primary lung cancer. Local treatment may result in symptomatic improvement but prognosis is generally poor averaging 1-2 years in most series. PMID- 10717335 TI - Seroepidemiology of Bordetella pertussis infections in the Spanish population: a cross-sectional study. AB - A study was conducted on a representative sample (n=4084) of the Spanish population to assess the prevalence of antibodies to pertussis toxin (PT) and filamentous hemagglutinin (FHA). A total of 1982 men and 2102 women aged 5-59 years were stratified by sex and age (5-12, 13-19, 20-29, 30-39, 40-49 and 50-59 years). Antibodies to PT were found in 46% samples and to FHA in 74% and increased with age (p<0.0001), ranging from 35% in the 5-12 year age group to 52% in the 50-59 year age group for anti-PT and from 65 to 80% for anti-FHA, respectively. As vaccine induced-immunity wanes over time, the observed age distribution of antibodies suggests that Bordetella pertussis infection is widespread in the Spanish population and that pertussis booster vaccination in adolescents and adults with the newly developed acellular vaccines, should be considered if it is deemed necessary to control the circulation of the organism. PMID- 10717336 TI - A nonionic block co-polymer adjuvant (CRL1005) enhances the immunogenicity and protective efficacy of inactivated influenza vaccine in young and aged mice. AB - The use of adjuvants is one approach to improve influenza vaccine immunogenicity and efficacy, particularly in aged populations. The response of BALB/c mice to subcutaneously administered formalin-inactivated whole influenza virus vaccine in the presence or absence of a nonionic block copolymer adjuvant CRL1005 was evaluated. In young adult naive mice, the copolymer adjuvant significantly enhanced virus-specific IgG and hemagglutination-inhibition (HI) antibody responses and augmented the production of IL-2 following vaccination. Influenza vaccine formulated with 2.5 mg CRL1005 significantly enhanced the protective efficacy of the inactivated vaccine in the upper and lower respiratory tract. In mice previously infected with influenza virus or naive aged mice, inactivated vaccine administered with the copolymer adjuvant substantially enhanced the serum HI antibody response to inactivated influenza vaccine and significantly reduced lung virus titers following subsequent challenge with live virus compared with mice administered vaccine alone. These results suggest that the copolymer adjuvant warrants further investigation as a potential adjuvant for use in human vaccination against influenza. PMID- 10717337 TI - Stability of aluminium-containing adjuvants during aging at room temperature. AB - Aluminium phosphate adjuvant and aluminium hydroxide adjuvant became more ordered during aging at room temperature. The increased degree of order was accompanied by a decrease in protein adsorption capacity. PMID- 10717338 TI - DeltaguaBA attenuated Shigella flexneri 2a strain CVD 1204 as a Shigella vaccine and as a live mucosal delivery system for fragment C of tetanus toxin. AB - The DeltaguaBA Shigella flexneri 2a vaccine candidate, CVD 1204, was evaluated as a delivery system for the non-toxic C-terminal of tetanus toxin (fragment C), either as a polypeptide expressed in the bacteria or as a DNA vaccine. CVD 1204 was transformed with plasmid pTETnir15 which encodes the fragment C gene (tetC) under the control of the inducible prokaryotic nir15 promoter or a DNA vaccine plasmid pcDNA3tetC which encodes tetC under the eukaryotic hCMV promoter. Guinea pigs immunised intranasally (i.n.) with either recombinant strain mounted a secretory immune response against S. flexneri 2a Lipopolysaccharide (LPS) and were protected against ocular challenge with wild-type S. flexneri 2a. Both strains were effective in eliciting a serum IgG response against fragment C in guinea pigs following i.n. immunisation. Furthermore, serum from guinea pigs immunised with CVD 1204(pTETnir15) contained tetanus toxin neutralising antibodies. These results demonstrate that this S. flexneri 2a vaccine candidate can serve as a vehicle for the delivery of foreign antigens to the systemic immune system while retaining its capacity to serve as a mucosal Shigella vaccine. PMID- 10717339 TI - Intranasal immunization with liposome-formulated Yersinia pestis vaccine enhances mucosal immune responses. AB - The induction of mucosal immune responses by a liposome-formulated Y. pestis vaccine (formaldehyde-killed whole cell vaccine; KWC) was evaluated. We demonstrated that intranasal immunization of mice with Y. pestis KWC vaccine, formulated with liposomes, significantly enhanced mucosal immune responses in the lung when compared to the responses induced with KWC vaccine alone. These immune responses were characterized by increased titres of specific IgA and IgG in mucosal secretions (lung and nasal washes), and an increased frequency of specific antibody-secreting cells in the lungs. In addition, antigen-specific proliferative responses and IFN-gamma-secreting cells were also significantly enhanced in the spleens of mice immunized with the KWC vaccine formulated in liposomes. Animals that were immunized intranasally with the KWC vaccine showed significant protection against an intranasal challenge with Y. pestis. These results highlight the importance of mucosal administration of vaccine antigens to stimulate immunity in the respiratory tract and demonstrate that liposome formulations can improve the effectiveness of conventional vaccines. PMID- 10717340 TI - Targeted expression of HTLV-I envelope proteins in muscle by DNA immunization of mice. AB - We have compared two types of plasmids for DNA immunization against HTLV-I envelope glycoproteins. One type of plasmid contains the coding DNA of the complete envelope gene of HTLV-I under the control of the CMV promoter with (CMVenvLTR) or without (CMVenv) the tax/rex genes. The second type contains the coding DNA of the complete env gene of HTLV-I under the control of the human desmin muscle specific promoter (DesEnv). These plasmids were inoculated into mice and the humoral response was studied by flow cytometry, ELISA and neutralization assays. Inoculation of the DesEnv construct elicited a higher humoral response with better neutralization properties than the injection of CMVenvLTR or CMVenv plasmids. The choice of vectors will be important for the design of genetic HTLV-I vaccines. PMID- 10717341 TI - A model to estimate the probability of hepatitis B- and Haemophilus influenzae type b-vaccine uptake into national vaccination programs. AB - Most countries have been slow to adopt new vaccines into national vaccination schedules, despite recommendations from global multi-lateral agencies. Characteristics of countries that have adopted hepatitis B (HB) vaccine were analysed and used to formulate a logistic regression model. The model was applied to country-specific data to predict HB and Haemophilus influenzae type b (Hib) vaccine uptake. The greatest predictors of HB uptake were coverage rates of other vaccines, vaccine cost relative to the economy, and perceived disease burden. The logistic regression model's probability estimate of vaccine uptake agreed well with observed data for HB and Hib, (c-statistic 85 and 82%, respectively). Application of this model to other antigens may aid in predicting potential national markets to better plan new vaccine supply and demand. PMID- 10717342 TI - Protective immunity against foot-and-mouth disease virus induced by a recombinant vaccinia virus. AB - We report the construction of a recombinant vaccinia virus expressing the precursor for the four structural proteins of FMD virus (FMDV) (P1) strain C3Arg85 using a procedure for isolation of recombinant vaccinia viruses based solely on plaque formation. Adult mice vaccinated with this recombinant vaccinia virus elicited high titers of neutralizing antibodies against both the homologous FMDV and vaccinia virus, measured by neutralization assays. Liquid phase blocking sandwich enzyme-linked immunosorbent assays (ELISAs) using whole virus as antigen showed high total antibody titers against homologous FMDV, similar to those induced by the conventional inactivated vaccine. When ELISAs were carried out with heterologous strains A79 or O1Caseros as antigens, sera from animals vaccinated with the recombinant virus cross-reacted. Mice boosted once with the recombinant vaccinia virus were protected against challenge with infectious homologous virus. These results indicate that recombinant vaccinia viruses are efficient immunogens against FMDV when used as a live vaccine in a mouse model. PMID- 10717343 TI - Granulocyte selected live Salmonella enteritidis vaccine is species specific. AB - Selection of Salmonellae from granulocytes may result in safe and effective vaccine strains. We demonstrated that the reduced virulence of such strains is limited to the species in which the selection was made. Repeated (sequential) selection of Salmonella enteritidis from chicken granulocytes yielded an avirulent strain for chickens. Repeated (sequential) selection of Salmonella enteritidis from pig granulocytes (neutrophils) yielded a strain that was comparable to the original wildtype strain. PMID- 10717344 TI - Preparation and characterisation of quillaja saponin with less heterogeneity than Quil-A. AB - Immunisation against pathogens remains one of the most effective ways of preventing or reducing losses due to infectious diseases in animal husbandry. When inactivated vaccines are used, adjuvants are most often required to obtain satisfactory immune responses. One such type of adjuvant is saponin derived from the bark of Quillaja saponaria Molina, a tree of the rose family. A few different commercial sources exist, but due to the structural complexity and heterogeneity of these saponin preparations, it has been difficult to establish exactly which components are responsible for the adjuvant activity. By carefully selecting the bark source, we have succeeded in preparing a much less heterogeneous preparation of quillaja saponin. In this report we describe the preparation, in terms of structural complexity, hemolytic activity, adjuvant activity, and its ability to form ISCOM matrix. This new preparation could have implications for use per se, or as starting material for more effective preparation of pure substances. PMID- 10717345 TI - A recombinant avipoxvirus HIV-1 vaccine expressing interferon-gamma is safe and immunogenic in macaques. AB - Complex recombinant fowlpoxvirus (rFPV) vaccines expressing both HIV-1 antigens and type 1 cytokines could facilitate the induction of cellular immunity against HIV-1. A single rFPV expressing both HIV-1gag/pol and human interferon-gamma (FPVgag/pol-IFNgamma) was constructed and assessed as a therapeutic vaccine for safety and immunogenicity in macaques (Macaca nemestrina) previously infected with HIV-1. FPV gag/pol-IFNgamma vaccinations were safe and enhanced T cell proliferative responses to Gag antigens (but not control tetanus antigens). Enhanced CTL responses to gag/pol antigens were also observed following IFNgamma expressing vaccinations. Since cellular immunity may be critical to controlling or preventing HIV-1 infection, these observations suggest that avipox vectors co expressing IFNgamma should be further evaluated as therapeutic or preventive HIV 1 vaccines. PMID- 10717346 TI - Rotavirus VP7 epitope mapping using fragments of VP7 displayed on phages. AB - cDNA copies of the complete porcine rotavirus CRW-8 VP7 gene were randomly digested to fragments of about 30-60 or 30-500 base pairs by DNase1 in the presence of Mn(2+). The fragments were cloned and expressed in a filamentous phage fd-tet-derived vector to create specific-gene-related peptide libraries. Polyclonal antibodies were then used to pan the SGRP libraries for antibody binding phages. Analysis of the phage isolates revealed that the majority (86%) of them only had a single insert. However, phages displaying composite inserts containing the VP7 antigenic regions A, B, and C, originally defined by neutralising monoclonal antibody escape mutants, were also isolated. Inserts containing A or C region peptide were found to contain extra sequences from the C region, while the B region epitope was linear and had additional sequence from either upstream or downstream. In addition a dominant and possibly non neutralising VP7 epitope was identified around amino acids 263-270. One of the recreated antigenic epitopes has also been fused to the outer membrane protein A (OmpA) of Escherichia coli and shown to maintain its antigenicity. The results in this study may have significant implication for recreation of conformational epitopes and vaccine development. PMID- 10717347 TI - Immunization with a chimeric tobacco mosaic virus containing an epitope of outer membrane protein F of Pseudomonas aeruginosa provides protection against challenge with P. aeruginosa. AB - A chimeric tobacco mosaic virus (TMV) was constructed by inserting sequences representing peptide 9-14mer (TDAYNQKLSERRAN) of outer membrane (OM) protein F of Pseudomonas aeruginosa between amino acids Ser154 and Gly155 of the TMV coat protein (CP). This is the first example of TMV being used to construct a chimera containing a bacterial epitope. Mice immunized with TMV-9-14 produced anti peptide-9-14mer-specific antibodies that reacted in whole-cell ELISA with all seven Fisher-Devlin (FD) immunotype strains of P. aeruginosa, reacted specifically by Western blotting with OM protein F extracted from all seven FD immunotypes, and were opsonic in opsonophagocytic assays. The chimeric TMV-9-14 vaccine afforded immunoprotection against challenge with wild-type P. aeruginosa in a mouse model of chronic pulmonary infection. TMV-9-14 is an excellent candidate for further development as a vaccine for possible use in humans to protect against P. aeruginosa infections. PMID- 10717348 TI - Control of ticks resistant to immunization with Bm86 in cattle vaccinated with the recombinant antigen Bm95 isolated from the cattle tick, Boophilus microplus. AB - The recombinant Bm86-containing vaccine Gavac(TM) against the cattle tick Boophilus microplus has proved its efficacy in a number of experiments, especially when combined with acaricides in an integrated manner. However, tick isolates such as the Argentinean strain A, show low susceptibility to this vaccine. In this paper we report on the isolation of the Bm95 gene from the B. microplus strain A, which was cloned and expressed in the yeast Pichia pastoris producing a glycosylated and particulated recombinant protein. This new antigen was effective against different tick strains in a pen trial, including the B. microplus strain A, resistant to vaccination with Bm86. A Bm95-based vaccine was used to protect cattle against tick infestations under production conditions, lowering the number of ticks on vaccinated animals and, therefore, reducing the frequency of acaricide treatments. The Bm95 antigen from strain A was able to protect against infestations with Bm86-sensitive and Bm86-resistant tick strains, thus suggesting that Bm95 could be a more universal antigen to protect cattle against infestations by B. microplus strains from different geographical areas. PMID- 10717349 TI - Influenza vaccination in children with asthma in health maintenance organizations. Vaccine Safety Datalink Team. AB - We assessed vaccination coverage and predictors of influenza vaccination in asthmatic children in four large Health Maintenance Organizations. We studied 68,839 children with asthma at four Health Maintenance Organizations (HMOs) in the 1995-1996 influenza season and 34,032 children at two HMOs in the 1996-1997 influenza season. In both seasons only 9-10% were vaccinated against influenza. Children who were hospitalized, had an emergency department visit for asthma or a prescription for a beta-agonist prior to the influenza season, were more likely to be vaccinated. Overall, 61% of the unvaccinated asthmatic children had made an outpatient clinic visit during months when influenza vaccination would have been appropriate. Vaccination coverage could be increased by taking advantage of all opportunities to vaccinate children with asthma whenever they make clinic visits in the fall and early winter. PMID- 10717350 TI - Diphtheria antitoxin response to DTP vaccines used in Swedish pertussis vaccine trials, persistence and projection for timing of booster. AB - Data from two Swedish pertussis vaccine trials with various combination vaccines were used to compare anti-diphtheria antitoxin concentrations over time between different vaccines, vaccine lots and vaccine schedules. The immune responses were measured with a validated ELISA method.Results are given for 1326 children, born 1992, that were recruited to the placebo (DT)-controlled Trial I which used a 2, 4, 6 month schedule. Two DTP acellular and one DTP whole cell vaccine were used. No DT boosters were given until 5 years of age. Trial II recruited children born 1993-94 and compared three DTP acellular vaccines with one DTP whole cell vaccine. Results are given for 306 children in a 2, 4, 6 month schedule and for 531 children in a 3, 5, 12 month schedule. The latter schedule gave significantly higher diphtheria antitoxin concentrations post third dose. The various DTP acellular vaccines and an inefficacious DTP whole cell vaccine gave lower antitoxin concentrations than both an efficacious DTP whole cell vaccine and the DT vaccine. The larger differences in antigen response between vaccines was reduced in the course of time. Generally, an initial rapid decline of antitoxin concentration was followed by a slower decline; the change typically occurring when the antitoxin concentration reached 0.13-0.16 EU/ml. The time needed to reach this level was between 6 and 10 months based on the initial vaccine response.A "best-fit" combined exponential regression model was used to predict the optimal timing for booster vaccinations against diphtheria.Our data support a 3, 5, 12 month schedule followed by a fourth dose 4-5 years after the third dose, depending upon the vaccine used. PMID- 10717351 TI - Rapid antibody response to influenza vaccination in "at risk" groups. AB - Persons attending for routine influenza vaccination in an urban practice each provided three specimens of blood for evaluating their immunological response. 138 (67%) of the 206 persons were defined as "at risk" by reason of morbidity as given in the guidelines published by the Chief Medical Officer. The mean age was 67 yr and 65% were aged 65 yr or more. By day 7, 71% of 31 persons had protective H(1)N(1) titres, 61% H(3)N(2) and 42% B. These proportions were similar to those found at day 14 and at day 21 based on 159 persons. These findings suggest that an effective immune response is mounted within seven days of vaccination indicating that the vaccination of persons "at risk" is worthwhile even after an epidemic has established itself. This is not a reason to modify present policy of routine vaccination in early winter well before epidemics are likely to occur. PMID- 10717352 TI - Recombinant expression and neutralizing activity of an MHC class II binding epitope of toxic shock syndrome toxin-1. AB - Toxic shock syndrome (TSS) is caused by the staphylococcal superantigen, TSST-1. The MHC class II binding domain of TSST-1 containing a conserved sequence with other related staphylococcal enterotoxins, comprising TSST-1 residues 47-64 [(T(47-64)], was expressed as a fusion protein with either glutathione-S transferase (GST(47-64)), filamentous phage coat protein (pIII(47-64)), or E. coli outer membrane porin protein (OprF(47-64)), or synthesized as a peptide conjugated to bovine serum albumin, BSA(47-64). GST(47-64), OprF(47-64) and BSA(47-64), but not pIII(47-64), all induced high-titer T(47-64)-specific antibodies in Balb/c mice. However, only anti-GST(47-64) antibodies inhibited (125)I-TSST-1 binding to MHC class II and abrogated TSST-1-induced T cell mitogenesis and TNFalpha secretion in human peripheral blood mononuclear cells. Purified GST(47-64) also inhibited (125)I-TSST-1 binding in a dose-dependent manner. These findings suggest that GST(47-64) may have potential as a recombinant peptide vaccine or TSST-1 receptor inhibitor against TSS. PMID- 10717353 TI - Evaluation of a subchronic (13-week) oral toxicity study, preceded by an in utero exposure phase, with arachidonic acid oil derived from Mortierella alpina in rats. AB - Arachidonic acid oil (ARA-oil) derived from the fungus Mortierella alpina for use in infant nutrition was tested in a subchronic (13-week) oral toxicity study in rats, preceded by an in utero exposure phase. The ARA-oil was administered as admixture to the rodent diet at dose levels of 3000 ppm, 15,000 ppm and 75,000 ppm. An additional high-dose group received 75,000 ppm ARA-oil in combination with 55,000 ppm fish oil containing docosahexaenoic acid (DHA), at a ratio of ARA to DHA, comparable to the ratio in mother's milk of 2:1. The total levels of fat in each diet were kept constant by adding the appropriate amounts of corn oil. A concurrent control group received 130,000 ppm corn oil in the diet. An additional carrier control group was fed unsupplemented rodent diet. Administration of the test substances from 4 weeks prior to mating, throughout mating, gestation, lactation of parental (F(0)) animals and weaning of the F(1) pups did not affect fertility or reproductive performance, nor the general condition of pups, viability, sex ratio or number of pups. Pup weight gain in the ARA/DHA-oil group was lower than the controls administered equal amounts of corn oil. In the subsequent subchronic study survival, clinical signs, body weight gain and food consumption were not adversely affected by the test substances. Ophthalmoscopic examination did not reveal any treatment-related changes. There were no treatment related effects observed up to dietary test substance concentrations of 15,000 ppm. The following statistically significant differences were found in the ARA high-dose group and /or in the ARA/DHA group compared to the corn oil control group: decreased alkaline phosphatase activity, decreases in cholesterol, triglycerides and phospholipids concentrations, increased creatinine and urea concentrations. Furthermore, these groups showed increased adrenal, spleen and liver weights. The incidence of hepatocellular vacuolation was increased in females of the ARA high-dose group and the ARA/DHA group. Oil droplets were observed in the mesenteric lymph nodes and in the intestinal villi in the ARA high-dose group and the ARA/DHA group. In addition, lipogranulomas were observed in the mesenteric lymph nodes in these groups. The observed changes in the high dose groups may be effects of the high intake of high-fat levels, rather than specific effects of the ARA-oil. The no-observed-effect level in this study was placed at 15,000 ppm ARA-oil. This level is equivalent to approximately 970mg ARA oil/kg body weight/day. PMID- 10717354 TI - Identification of ochratoxins and some of their metabolites in bile and urine of rats. AB - The objectives of this study were to develop and evaluate procedures for the confirmation of ochratoxin A (OA), lactone opened OA (OP-OA), ochratoxin B (OB), hydroxy OA (OA-OH) and ochratoxin alpha (Oalpha) and metabolites formed in the rats from these toxins, and to demonstrate that many ochratoxin metabolites can be identified in the bile and urine of rats injected with the different ochratoxins. An esterification procedure in acidified methanol and a lactone hydrolysis procedure in strong base yielded two additional forms of most of the different ochratoxins. The esterification procedure provided a simple, fast and reliable method for the confirmation of the ochratoxins. A total of 20 different metabolites of OA, OP-OA, OB, OA-OH and Oalpha were detected in the urine and the bile of rats of which several were identified. Among these, OA and the recently discovered and toxic form of OA (OP-OA) were readily formed in vivo when either were injected. Procedures developed in this study can be used to confirm and isolate ochratoxins in biological samples and have shown that a new form of OA (OP-OA) along with many other metabolites are formed from OA and related ochratoxins in vivo. PMID- 10717355 TI - Acute oral toxicity and bacterial translocation studies on potentially probiotic strains of lactic acid bacteria. AB - Three potentially probiotic lactic acid bacteria (LAB) strains, Lactobacillus rhamnosus HN001 (DR20(TM)), Lb. acidophilus HN017 and Bifidobacterium lactis HN019 (DR10()), have recently been identified and characterized. The present study was designed to evaluate the acute oral toxicity of these strains to mice, and also to investigate bacterial translocation and gut mucosal pathology in BALB/c mice fed HN019, HN001 or HN017 for 8 consecutive days at a high dose of 10(11)cfu/mouse/day. Results showed that these probiotic strains had no adverse effect on general health status, feed intake, body weight gain and intestinal mucosal morphology (villus height, crypt depth, epithelial cell height and mucosal thickness). No viable bacteria were recovered from blood and tissue samples (mesenteric lymph nodes, liver and spleen) of mice, and no treatment associated illness or death was observed. According to these results, the oral LD(50) of HN019, HN001 and HN017 is more than 50g/kg/day for mice, and their acceptable daily intake (ADI) value is 35g dry bacteria per day for a 70-kg person. This suggests that the probiotic strains HN019, HN001 and HN017 are non pathogenic and likely to be safe for human consumption. PMID- 10717356 TI - Inhibition of N-acetyltransferase activity and DNA-2-aminofluorene adducts by glycyrrhizic acid in human colon tumour cells. AB - Glycyrrhizic acid (GA) was tested for inhibition of arylamine N-acetyltransferase (NAT) activity in a human colon tumour (adenocarcinoma) cell line (colo 205). Two assay systems were performed, one with cellular cytosols (9000g supernatant), the other with intact colon tumour cell cultures. The NAT activity in a human colon tumour cell line was inhibited by GA in a dose-dependent manner in both types of systems examined. The data also indicated that GA decreased the apparent values of K(m) and V(max) of NAT enzymes from human colon tumour cells in both examined systems. The DNA-2-aminofluorene adduct formation in human colon tumour cells were inhibited by GA. This report is the first to demonstrate that GA does inhibit human colon tumour cell NAT activity and DNA adduct formation. PMID- 10717357 TI - Nutritional value of ganoderma extract and assessment of its genotoxicity and antigenotoxicity using comet assays of mouse lymphocytes. AB - The nutritive composition of a hot aqueous extract of wild Ganoderma fruit bodies was determined. This extract was assessed for cytotoxicity and in vivo genotoxicity by both acute and subchronic exposure of mice (given by mouth at a dose equivalent to extract of 220g fresh Ganoderma fruit body/kg body weight). To test any alleged protection against mutagens by Ganoderma treatments, the mice were injected intraperitoneally with the radiomimetic mutagen ethyl methanesulfonate (EMS), and after 24hr of treatment their lymphocytes were examined using the comet assay. Ganoderma extract consisted of Folin-positive material (68.9% of dry weight), but protein comprised only 7.3% of dry weight. Glucose accounted for 11. 1% and metals 10.2% of dry weight (K, Mg and Ca being the major components with Ge (often touted as being of value in sales literature for Ganoderma preparations) having the fifth highest metal concentration at 489 microg/g). In comparison to rodent chow, Ganoderma extract was a modest dietary supplement. No evidence was found for genotoxic chromosomal breakage nor cytotoxic effects by Ganoderma extract in the mouse, nor did it protect against the effects of ethyl methanesulfonate. We found no support in this study for the extract having any value in protecting against the test mutagen. PMID- 10717358 TI - CM3, one of the wheat alpha-amylase inhibitor subunits, and binding of IgE in sera from Japanese with atopic dermatitis related to wheat. AB - The purpose of the present study was to determine whether wheat alpha-amylase inhibitor subunits bind IgE sera from Japanese with atopic dermatitis (AD) related to wheat. IgE in sera from eight out of the 11 patients with RAST value of 3 or 4 against wheat-bound salt-soluble proteins of wheat and the alpha amylase inhibitor proteins separated by gel filtration chromatography. The subunits of tetrameric alpha-amylase inhibitor, CM2, CM3 and CM16 (molecular weight approximately 14 kd), known to be major allergens for baker's asthma, were purified. Then the binding activity to IgE sera from the AD patients was examined by immunoblotting. The IgE sera bound only to CM3, not to CM2 and CM16. These results suggest that CM3 may be involved in both atopic dermatitis and baker's asthma. PMID- 10717359 TI - Lack of carcinogenicity and increased survival in F344 rats treated with 5 fluorouracil for two years. AB - The carcinogenicity of 5-fluorouracil (5-FU), a compound employed as an antineoplastic drug, was investigated in F344 rats of both sexes. 5-FU was administered to groups of 50 male and 50 female rats ad lib. for 104 weeks, added to drinking water at concentrations of 0 (control), 62 and 125 ppm, these dose levels being selected on the basis of results of a 13-week subchronic toxicity study. Body weight gains were slightly depressed in the 125 ppm group of both sexes. While not statistically significant in females, final survival rates at week 111 in the 125 ppm group of both sexes were higher than those in the control group, suggesting an ability of 5-FU to prolong the lifespan. Histopathologically, a decreased incidence of islet cell adenomas in males and increased incidences of pituitary gland adenomas and pheochromocytomas in females were observed in the 62 ppm group without dose dependence. There was no significant induction of any other neoplastic or non-neoplastic lesions. These results indicate a lack of carcinogenicity of 5-FU under the present experimental conditions using rats. PMID- 10717360 TI - A limited three-generation reproduction study on hexachlorocyclohexane (HCH) in rats. AB - Dietary feeding of technical hexachlorocyclohexane (HCH) at 125 and 250 ppm to rats have not shown any adverse effects on reproductive function and were comparable to control animals in a three-generation study. There were no major malformations, while some minor variants found were not compound or dose related. Despite some mild toxicological effects in rats of the P(0) generation, the litters of F(1B), F(2B) and F(3B) generations were devoid of any morphological or teratological changes. The presence of HCH residues in vital tissues of F(3B) pups have indicated transmigration of HCH in preceding generations but not to an extent that produced adverse effects. PMID- 10717361 TI - Characteristics of rat platelets and relative contributions of platelets and blood coagulation to haemostasis. AB - In order to understand some of the haemostatic mechanisms in rats for the interpretation of toxicological data, basic haemostatic parameters with a special emphasis on platelet functions were first measured in vitro. The results of reactions of rat platelets to many aggregating agents suggest that only ADP may be a consistently significant aggregator. The search for physiologic aggregators revealed ADP to be available from erythrocytes. Adhesion reaction also required ADP. Collagen was not considered to be essential for either reaction. Aggregation and adhesion were probably both reversible in flowing blood, while irreversible thrombi were formed in blood at rest ex vivo. Blood coagulation parameters determined revealed that the intrinsic pathway may be more important than the extrinsic one. The rate of intrinsic coagulation reaction was rapid, and plasma coagulation appeared to be of primary importance while the influence of platelet aggregation was minor. A simple model of rat haemostatic mechanism is proposed based on these results. Additionally, to define the relative contribution of platelets versus other cellular and plasma coagulation in vivo, rats were administered antiplatelet drugs (ticlopidine, suprofen and clopidogrel) and an anticoagulant (warfarin) intraperitoneally. Bleeding times (BTs) were significantly increased in all treated groups. ADP-induced platelet aggregations were significantly depressed by the administration of the three antiplatelet drugs, while kaolon-activated partial thromboplastin time and prothrombin time were greatly increased in the warfarin-treated rats. The increase in BT may be due to the inhibition of platelet activity or blood coagulation defect in rats given antiplatelet drugs or warfarin, respectively. These results suggest that platelets play a key role in haemostasis in the rat. Two possible explanations of the disparity between in vitro and in vivo results may be that functional tests used here are not adequate to cover the properties of rat platelets or that mechanisms leading to the formation of platelet thrombi in rats are ADP-dependent adhesion and ADP-induced aggregation. PMID- 10717362 TI - Rodent carcinogenicity tests need be no longer than 18 months: an analysis based on 210 chemicals in the IARC monographs. AB - The IARC Monographs (Vols 1-70) were studied to determine the time of onset of treatment-related tumorigenicity in long-term rodent studies for chemicals classified by IARC as having sufficient evidence of carcinogenicity in animals. The analysis excluded studies on metals and their salts, studies on particulates, studies by parenteral routes of administration that resulted in tumours only at the site of exposure, and studies that did not approximate to the current standard long-term rodent carcinogenicity bioassay, for instance transplacental or multigeneration studies, initiator-promoter studies, lung tumour assays in Strain A mice and studies in newborn animals. Data from a total of 210 chemicals revealed that, overall, evidence of treatment-related tumorigenicity was first apparent within 12 months for 66% of the chemicals and for only 7% were studies of longer than 18 months necessary. All IARC Group 1 chemicals were detected in animals within 18 months, and most within 12 months. Most of the tumour types that required more than 18 months for detection were of dubious relevance to human risk assessment. Termination of rodent carcinogenicity studies at 18 months or earlier would greatly reduce the complications that arise in interpreting the findings in aged animals which often have defective hepatic or renal function and would also markedly reduce the time required for histopathological examination of dozens of tissues taken from the approximately 500 animals routinely employed in these studies. PMID- 10717363 TI - Review of risk factors for osteoporosis with particular reference to a possible aetiological role of dietary salt. AB - Laboratory animal, clinical and epidemiological studies in the published literature have been reviewed in order to establish whether excessive salt intake is an important risk factor for the development of osteoporosis and whether an intervention strategy based on salt restriction would be beneficial in the prevention of osteoporosis. Genetic factors appear to be far more important than the combination of nutritional, hormonal, environmental and lifestyle factors in the pathogenesis of osteoporosis. The most important single non-genetic factor is oestrogen deficiency in postmenopausal women. Preventive measures should be aimed at maximizing peak bone mass at skeletal maturity and retarding bone loss thereafter. Apart from postmenopausal oestrogen deficiency, various factors have been incriminated as risk factors for osteoporosis, and these include age at menarche, age at and years since menopause, insufficient physical exercise, alcohol, smoking, low calcium intake, low or high protein intake and high intake of phosphorus, sodium or caffeine. Many of the risk factors are considered to be weak, although when combined they could impact significantly on bone health. Increased intakes of various nutritional factors (potassium, magnesium, zinc, vitamin C), fibre and alkaline-producing fruit and vegetables favour adult bone health. Calcium homeostasis is normally well regulated such that increased calcium loss via the urine leads to increased calcium absorption from the gut. However, the duration of this adaptive process may be greater than that of many of the studies demonstrating that increased salt intake leads to both increased sodium and calcium in the urine. In any case, higher urinary calcium output appears to be seen only in a minority of humans in response to increased salt intake. As numerous factors-genetic, nutritional, hormonal and lifestyle-are involved in the maintenance of calcium homeostasis, it is difficult to devise human studies which adequately take into account all the important factors. Another difficulty is that many past studies have relied on imprecise methods for the measurement of bone resorption. Nor have studies based on the use of the laboratory rat produced clear answers to the problem because the rat, as a species, is uniquely deficient in its ability to handle the relevant minerals. Limited studies to date indicate that increased sodium intake neither exerts a consistent effect on various biomarkers of bone health nor leads to irreversible changes in the bone modelling process in men or in pre- or postmenopausal women. We conclude from the available evidence that increased sodium (or salt) intake is not an important risk factor for osteoporosis and that a reduction of salt intake from 9 to 6g/day in the diet would not be beneficial as an intervention measure in the prevention of osteoporosis. More research is needed to (i) assess the effects (especially long-term) of various nutrients including sodium on bone health, (ii) assess the long-term value of any intervention strategy involving reduced intake of a particular nutrient such as sodium; and (iii) determine whether subpopulations exist particularly in the elderly (e.g. sodium-responsive subjects) in which adaptation to sodium-induced hypercalciuria may be compromised. General prudence dictates that excessively high levels of dietary salt should be eschewed by those persons with raised blood pressure or a limited range of genetic disorders. However, for the generally healthy person there is no sound evidence that the consumption of salt at the present average level of 9g/day constitutes a risk factor for osteoporosis. PMID- 10717364 TI - Threshold of toxicological concern for chemical substances present in the diet: a practical tool for assessing the need for toxicity testing. AB - The de minimis concept acknowledges a human exposure threshold value for chemicals below which there is no significant risk to human health. It is the underlying principle for the US Food and Drug Administration (FDA) regulation on substances used in food-contact articles. Further to this, the principle of Threshold of Toxicological Concern (TTC) has been developed and is now used by the Joint FAO/WHO Expert Committee on Food Additives (JECFA) in their evaluations. Establishing an accepted TTC would benefit consumers, industry and regulators, since it would preclude extensive toxicity evaluations when human intakes are below such threshold, and direct considerable time and cost resources towards testing substances with the highest potential risk to human health. It was questioned, however, whether specific endpoints that may potentially give rise to low-dose effects would be covered by such threshold. In this review, the possibility of defining a TTC for chemical substances present in the diet was examined for general toxicity endpoints (including carcinogenicity), as well as for specific endpoints, namely neurotoxicity and developmental neurotoxicity, immunotoxicity and developmental toxicity. For each of these endpoints, a database of specific no-observed-effect levels (NOELs) was compiled by screening oral toxicity studies. The substances recorded in each specific database were selected on the basis of their demonstrated adverse effects. For the neurotoxicity and developmental neurotoxicity databases, it was intended to cover all classes of compounds reported to have either a demonstrated neurotoxic or developmentally neurotoxic effect, or at least, on a biochemical or pharmacological basis were considered to have a potential for displaying such effects. For the immunotoxicity endpoint, it was ensured that only immunotoxicants were included in the database by selecting most of the substances from the Luster et al. database, provided that they satisfied the criteria for immunotoxicity defined by Luster. For the developmental toxicity database, substances were selected from the Munro et al. database that contained the lowest NOELs retrieved from the literature for more than 600 compounds. After screening these, substances showing any effect which could point to developmental toxicity as broadly defined by the US were recorded in the database. Additionally, endocrine toxicity and allergenicity were addressed as two separate cases, using different approaches and methodology. The distributions of NOELs for the neurotoxicity, developmental neurotoxicity and developmental toxicity endpoints were compared with the distribution of NOELs for non-specific carcinogenic endpoints. As the immunotoxicity database was too limited to draw such a distribution of immune NOELs, the immunotoxicity endpoint was evaluated by comparing immune NOELs (or LOELs-lowest-observed-effect levels-when NOELs were not available) with non-immune NOELs (or LOELs), in order to compare the sensitivity of this endpoint with non-specific endpoints. A different methodology was adopted for the evaluation of the endocrine toxicity endpoint since data currently available do not permit the establishment of a clear causal link between endocrine active chemicals and adverse effects in humans. Therefore, this endpoint was analysed by estimating the human exposure to oestrogenic environmental chemicals and evaluating their potential impact on human health, based on their contribution to the overall exposure, and their estrogenic potency relative to endogenous hormones. The allergenicity endpoint was not analysed as such. It was addressed in a separate section because this issue is not relevant to the overall population but rather to subsets of susceptible individuals, and allergic risks are usually controlled by other means (i.e. labelling) than the Threshold of Toxicological Concern approach. (ABSTRACT TRUNCATED) PMID- 10717365 TI - ILSI Europe Workshop on Assessing Health Risks From Environmental Exposure to Chemicals: the Example of Drinking Water. Summary Report. International Life Sciences Institute. PMID- 10717366 TI - The epidemiology of chemical contaminants of drinking water. AB - A number of chemical contaminants have been identified in drinking water. These contaminants reach drinking water supplies from various sources, including municipal and industrial discharges, urban and rural run-off, natural geological formations, drinking water distribution materials and the drinking water treatment process. Chemical contaminants for which epidemiologic studies have reported associations include the following: aluminium, arsenic, disinfection by products, fluoride, lead, pesticides and radon. Health effects reported have included various cancers, adverse reproductive outcomes, cardiovascular disease and neurological disease. In evaluating epidemiologic studies for risk assessment, considering whether the study design was qualitative (hypothesis generating) or quantitative (hypothesis testing) is important and whether sufficient epidemiologic data of a quantitative nature exists to determine the dose-response curve. Each of the chemical contaminants mentioned are summarized by study designs (qualitative and quantitative) and whether a dose-response curve based on epidemiologic data has been proposed. Environmental epidemiology studies are driven by environmental exposures of interest. For drinking water contaminants, the design of epidemiologic studies and their interpretation should consider the following exposure issues: the source of the contaminant; other sources of the contaminant; the route of exposure; the frequency, duration and magnitude of exposure; the ability to document an actual internal dose; and the ability to document the dose to the target organ. Health effects of concern have other risk factors that must be measured in the conduct of these studies. In evaluating epidemiologic studies, potential errors and biases that may occur must be considered given the very low magnitude of associations (less than 2.0 for either odds ratio or risk ratio). Given the issues, the next generation of drinking water epidemiologic studies should include a multidisciplinary team beyond traditional epidemiologists and statisticians. Study teams will require toxicologists, chemists, engineers and exposure assessors. Arsenic is briefly discussed as an example of the importance of susceptible populations. Disinfection by-products are discussed as an example of epidemiologic studies of mixtures. PMID- 10717367 TI - Sources of chemical contaminants and routes into the freshwater environment. AB - Drinking water is derived from either surface waters or groundwater. The latter is of enormous importance, with more than 65% of Europe's drinking water needs being supplied in this way. However, water from either source is rarely, if ever, pure. Industrialization and urbanization together with intensified agricultural activity have led to increased demands for water on the one hand but to the potential for large scale release of contaminants on the other. The result is that surface water can be contaminated through direct or indirect emissions and groundwater can be contaminated by leaching from the soil. The diversity and number of existing and potential sources of chemical contamination are quite large. This paper reviews the major sources of chemical emissions and the routes by which contaminants can arise in surface waters and groundwaters intended for use as a supply of drinking water. It is estimated that there are between 90,000 and 100,000 chemicals in regular use but that as few as 3000 account for about 90% of the total mass used. Whether a substance may be found in the air, soil or aqueous environment depends on a number of factors, including how the chemical is released, the volume released, where the chemical is released, its release pattern and its physicochemical properties. Of the major routes of contamination for the aquatic environment, the most significant are directly from treated and untreated waste waters, run-off and atmospheric deposition (including spray drift) and indirectly from leaching. The fate of emissions of contaminants in a particular water body will depend not only on the amount of the substance emitted but also on the transport, dispersion and transformation (biodegradation, hydrolysis, photolysis) processes in the receiving body. The preventative measures (biodegradation testing and sewage treatment) taken to minimize contamination are discussed. PMID- 10717368 TI - Water as consumed and its impact on the consumer--do we understand the variables? AB - Water is the most important natural resource in the world, without it life cannot exist. In 1854 a cholera outbreak in London caused 10, 000 deaths and positively linked enteric disease with bacterial contamination of drinking water by sewage pollution. Since then, adequate water hygiene standards and sewage purification have played the most significant role in disease eradication and public health improvements everywhere. Standards for drinking water have become an extensive range of microbiological and chemical parametric values. Which has not increased consumer, if the media is to be believed. Customers rightly expect that the water they drink is safe and wholesome. Standard setting is perceived as a precise science and meaningful to health. Is this justified and do scientists and regulators who derive and set the standards understand the uncertainties in the system? Water is the universal solvent, therefore it will never be pure; it will contain impurities prior to and after treatment. Knowledge of its potential to become contaminated is necessary to understand the epidemiology associated with waterborne contaminants and their effects. Water use patterns vary considerably and affect assumptions based on toxicology derived from laboratory studies under tightly controlled conditions. Consideration must be given to the model systems used to assess toxicity and translate results from the laboratory to the real world, if sensible scientifically-based water quality standards are to be set and achieved cost effectively. PMID- 10717369 TI - Levels of exposure from drinking water. AB - The relative exposure from drinking water is generally small, although there is a lack of information on total daily intake of individual organic micropollutants. There are, however, a few exceptions. Materials used in domestic distribution systems (lead, copper and plastics) may cause a deterioration of the water quality, especially in stagnant water. The relative exposure to the related compounds may increase considerably. Monitoring data from the tap (with defined sampling techniques) are needed. Also, disinfection/oxidation by-products (bromate, trihalomethanes) can be present in drinking water in considerable amounts and the relative exposure from drinking water may even approach 100%. Especially for volatile organic micropollutants, exposure routes from drinking water other than ingestion must be taken into account (inhalation, percutaneous uptake). When there is a need for detection of substances at very low levels it is important that the measurements are reliable. International interlaboratory comparisons for organic micropollutants are lacking at the moment. PMID- 10717370 TI - Predictive exposure modelling--a case study with a detergent surfactant. AB - Environmental exposure estimations are generally based on a knowledge of how and in what quantity a substance enters the environment and how it may subsequently be distributed and transformed. Once present within the environment, biota (including man) may be exposed. This paper outlines the tools commonly used to estimate environmental exposure to ingredients from detergents and other household products. Such products are typically manufactured in large quantities, used by many people, and disposed of after household use into the environment via the sewer. The vast majority of this waste stream is treated via domestic wastewater treatment plants (WWTPs) as documented in the sewage treatment Directive 91/275/EEC. WWTPs significantly reduce the load of chemical substances to the receiving surface waters, and have become an intrinsic part of exposure and risk assessment of household chemicals. WWTP models are generally first-order (e.g. SIMPLETREAT, WWTREAT) or mixed-order (e.g. Monod) kinetics and can exhibit, potentially, very distinct dependencies on the influent concentration. Thus, the correct representation of xenobiotic behaviour in a WWTP and modeling of their fate has a significant impact on exposure assessment. The Environmental Risk Assessment Steering Committee (ERASM) of the Association Internationale de la Savonnerie et la Detergence, et des Produits d'Entretiens (AISE) and the Comite Europeen de Agents de Surface et Intermediares Organiques (CESIO) has commissioned a joint industry Task Force of the Association to develop and apply specific methodology for the environmental monitoring of surfactants, and verification of fate models. The monitoring programmes have been designed to (1) establish the fate, distribution and concentrations of the major surfactants used in detergents-linear alkylbenzene sulfonate (LAS), alcohol ethoxylates (AE), alcohol ethoxylated sulfates (AES) and soap in relevant environmental compartments and (2) to provide the necessary data for checking the applicability of mathematical models to predict their fate and concentrations in these environmental compartments. The case study detailed here, specifically focuses on the refinement of the LAS exposure assessment for surface waters in The Netherlands. PMID- 10717371 TI - Safe drinking water: the toxicologist's approach. AB - The production of adequate and safe drinking water is a high priority issue for safeguarding the health and well-being of humans all over the world. Traditionally, microbiological quality of drinking water has been the main concern, but over the last decades the attention of the general public and health officials on the importance of chemical quality and the threat of chemical pollutants have increased with the increase of our knowledge on the hazards of chemical substances. There are many sources of contamination of drinking water. Broadly they can be divided into two categories: contaminants originating from surface and groundwater, and contaminants used or formed during the treatment and distribution of drinking water. Contaminants in surface and groundwater can range from natural substances such as arsenic and manganese leaching from soil, to contaminants introduced by human activities, such as run-off from agricultural activities, controlled discharge from sewage treatment works and industrial plants, and uncontrolled discharges or leakage from landfill sites and from chemical accidents. Disinfectants and disinfectant by-products are well known contaminants resulting from the processes used by the drinking water industry for the treatment and distribution of water. The basic question in the production of drinking water is how to rid drinking water of potentially dangerous microorganisms and chemicals without introducing new hazards that might pose new and different threats to human health. It is the responsibility of toxicologists to provide risk assessments for chemical pollutants and to derive guidelines or standards for drinking water quality below which no significant health risk is encountered, to assure consumers that drinking water is safe and can be consumed without any risk. This paper will focus on the toxicological procedures used by the World Health Organization to derive guideline values for chemical compounds in drinking water, and will touch upon some critical differences in the nature of guidelines and legally binding standards. PMID- 10717372 TI - Measuring risks in humans: the promise and practice of epidemiology. AB - Epidemiology has been considered the fundamental science of public health policy. The use of epidemiologic data in environmental health policy has been limited particularly in the environmental regulatory arena. Epidemiologic risk assessment (ERA) is different from risk assessment and the interplay between the two has led to some misconceptions over the promise and practice of epidemiologic data. The current risk assessment process was designed in a time when the need for regulation was great and the epidemiologic information was sparse. There was little time for the consideration of conducting specific studies to improve the information base for environmental health policy. Animal bioassays could be conducted under standardized protocols within defined time periods. The limitations and uncertainties of animal studies also became standardized and risk assessors became comfortable with their models of extrapolation. As the cost of regulations have grown, the economic realities of regulating with little or no data to support actual public health benefit have become a political and legal liability. Major among epidemiology's advantages is that the information is of direct relevance. The majority of epidemiology data are observation and whether the number of studies is broad enough, the data can be generalized to major segments of the population. The uncertainties in animal-based risk assessments are likely to be greater than the uncertainties associated with epidemiologic studies. Another advantage is the range of extrapolation is often smaller. Another advantage is that epidemiologic data include the genetic diversity and variability in other endogenous factors inherent in human populations. The homogeneity of animal studies has often been cited as an advantage but is unrepresentative of the heterogeneity of the human race. Epidemiology does have its limitations. A major limitation is the time needed to obtain a database sufficient for policy-making purposes and the resources needed to conduct the research to develop the database. This has often prompted the conduct of poorly designed studies, forced the use of data collected for other purposes or improper use of existing data. Four situations where epidemiology should be pursued are discussed. Once an environmental health decision has been made, epidemiologic studies should be considered for documenting the reduction of exposure and therefore disease in the population. This traditional use of epidemiology has rarely been applied in the environmental health arena. A final consideration on the use of epidemiologic data is the need to provide a sense of perspective to set priorities in the larger context of public health priorities. The role of environmental pollutants in causing disease may in some cases be minor in comparison to other risk factors and needs to be considered in setting environmental regulations. PMID- 10717373 TI - Mixtures and interactions. AB - Drinking water can be considered as a complex mixture that consists of tens, hundreds or thousands of chemicals of which the composition is qualitatively and quantitatively not fully known. From a public health point of view it is most relevant to answer the question of whether chemicals in drinking water interact in a way that results in an increased overall response as compared to the sum of the responses to the individual chemicals in the mixture, or indeed in an effect that is simply a summation of the expected effects of the individual chemicals. Present methods for risk assessment of mixtures rely heavily on some form of additivity model, unless data are adequate for a direct risk assessment of the mixture of concern in its entirety. The "dose-addition" concept ("simple similar action") is the most common approach to risk assessment of mixtures and it is applicable over the whole range of exposure levels from low non-toxic to toxic levels when all chemicals in the mixture act in a similar way. However, in toxicity studies at environmentally relevant exposure scenarios the mixtures that meet such conditions are the exception rather than the rule. In that case the "effect addition" model has to be followed assuming "independent joint action". For these compounds now experimental data have indicated that the results at low exposure levels are probably difficult to predict based on response additivity found at higher dose levels. Thus, although the additivity models are mathematically simple, they require assumptions about the mechanisms of action and the high-to-low dose extrapolation. Therefore, theoretical considerations in risk assessment of chemical mixtures should be verified by simple case studies. Up till now, the number of environmentally relevant mixtures to which a direct risk assessment has been devoted is limited. Even if toxicity data on individual compounds are available, we are still facing the immense problem of extrapolation of findings obtained at relatively high exposure concentration in laboratory animals to man being exposed to (much) lower concentrations. Therefore the prioritization of compounds for further research and the extrapolation to low doses should be considered as key issues in the assessment of possible health risks from exposure to chemical mixtures such as drinking water. PMID- 10717374 TI - Public health implications of new guidelines for lead in drinking water: a case study in an area with historically high water lead levels. AB - Concern about the neurotoxicity of lead, particularly in infants and young children, has led to a revision of blood lead levels which are considered to involve an acceptable level of human exposure. Drinking water guidelines have also been reviewed in order to reduce this source of population exposure to lead. In the last 20 years, guidelines have been reduced from 100 to 50 to 10 microg/litre. Lead in tap water used to be a major public health problem in Glasgow because of the high prevalence of houses with lead service pipes, the low pH of the public water supply and the resulting high levels of lead in water used for public consumption. Following two separate programmes of water treatment, involving the addition of lime and, a decade later, lime supplemented with orthophosphate, it is considered that maximal measures have been taken to reduce lead exposure by chemical treatment of the water supply. Any residual problem of public exposure would require large scale replacement of lead service pipes. In anticipation of the more stringent limits for lead in drinking water, we set out to measure current lead exposure from tap water in the population of Glasgow served by the Loch Katrine water supply, to compare the current situation with 12 years previously and to assess the public health implications of different limits. The study was based on mothers of young children since maternal blood lead concentrations and the domestic water that mothers use to prepare bottle feeds are the principal sources of foetal and infant lead exposure. An estimated 17% of mothers lived in households with tap water lead concentrations of 10 microg/litre (the [WHO,] guideline) or above in 1993 compared with 49% in 1981. Mean maternal blood lead concentrations fell by 69% in 12 years. For a given water lead concentration, maternal blood lead concentrations were 67% lower. The mean maternal blood lead concentration was 3.7 microg/litre in the population at large, compared with 3.3 microg/litre in households with negligible or absent tap water lead. Nevertheless, between 63% and 76% of cases of mothers with blood lead concentrations of 10 microg/dl or above were attributable to tap water lead. The study found that maternal blood lead concentrations were well within limits currently considered safe for human health. About 15% of infants may be exposed via bottle feeds to tap water lead concentrations that exceed the WHO guideline of 10 microg/litre. In the context of the health and social problems which affect the well-being and development of infants and children in Glasgow, however, current levels of lead exposure are considered to present a relatively minor health problem. PMID- 10717375 TI - Clues and uncertainties in the risk assessment of arsenic in drinking water. AB - On the basis of studies of the prevalence of skin cancer among users of As-rich well water in Taiwan, WHO experts recommended in 1984 a maximum As concentration of 50 microg/litre in drinking water. Since that time, a plethora of non-cancer as well as cancer effects has been observed in several other populations sustaining a chronic exposure to various As concentrations in drinking water. This prompted a revision of the standard and a provisional guideline of 10 microg/litre was recommended in 1993. While the uncertainty linked to the statistical inferences leading to the guideline are reduced by the fact that they are directly estimated from human data and result from extrapolations made relatively close to observed exposure levels, developed guideline depends strongly on the choice of the dose-response model (linear, quadratic, hockey stick) and the accuracy of the exposure data. The potential exposure to As sources other than drinking water, dietary habits and genetic characteristics of the populations may also make more difficult the inference of a recommendation for As concentration in drinking water. Owing to the huge cost of strongly reducing the current As in water standard, many efforts are presently made to clarify the quantitative aspects of As-induced cancers, particularly at low dose levels. New data on the metabolism and carcinogenic mechanism of As in humans along with the results of epidemiological studies presently under way in several countries will help to reduce the uncertainty in the risk assessment of As. PMID- 10717376 TI - Pesticides in drinking water--a case study. AB - Pesticides occupy a unique position among chemicals found in drinking water, since they are deliberately used to control pests in agriculture and public health. They comprise a variety of compounds of various chemical properties (many of which being persistent in the environment), toxic potential, and mechanism of action. Safety assessment for drinking water is conducted by allocating health based guidelines for exposure through this environmental medium. The tolerable daily intake (TDI) approach is used by WHO in the case of pesticides with assumed thresholds of effect, while low-dose extrapolation is used in the case of non threshold carcinogens. Risk management, however, can also be based on the precautionary principle. Risk assessment evaluates the risk emanating from a given exposure on the basis of all available data. PMID- 10717377 TI - Risk assessment case study--chloroform and related substances. AB - Chlorine, used as an important disinfectant for drinking water, can react with naturally occurring organic matter to form chloroform, bromodichloromethane, chlorodibromomethane and bromoform. Chloroform and other trihalomethanes have been shown to increase tumours of the liver, kidney or large intestine in rats or mice. The risk to man from these contaminants must be assessed carefully since there is considerable benefit associated with the use of chlorine. The weight of evidence suggests that chloroform is non-genotoxic and there are data, for each site, to indicate that tumours only occur at high doses where there is also tissue damage. Bromodichloromethane has also been shown to increase liver and kidney tumours but this and bromoform have been shown to increase large intestinal tumours in rats. The weight of evidence is that they are only weak genotoxins and they do not appear to be active in vivo. It is probable that the mechanism for the liver and kidney tumours is the same as for chloroform but the mechanism for the large intestinal tumours is uncertain. However, the toxicity and carcinogenicity of these substances is profoundly affected by dosing in corn oil. New studies suggest that dosing in drinking water would not result in increases in tumours. The evidence suggests that the use of a threshold approach, based on a tolerable daily intake, would be the most appropriate way of determining safe levels in drinking water. PMID- 10717378 TI - Effects of di-n-butyl phthalate (DBP) on male reproductive development in the rat: implications for human risk assessment. AB - The National Toxicology Program (NTP) conducted a continuous breeding study in SD rats with di-n-butyl phthalate (DBP) given via the diet at dose levels of up to 650 mg/kg/day. In the parental generation effects on reproduction were modest (small decreases in litter size and pup weight following treatment). However, the F(1) male offspring had marked decreases in fertility (at 650 mg/kg/day), with reduced sperm counts and reproductive tract malformations on reaching adulthood. A no-observed-adverse-effect level (NOAEL) was not established for the study [lowest-observed-adverse-effect level (LOAEL) 66 mg/kg/day]. In a study conducted at CIIT, the majority of these adverse changes could be reproduced over a similar dose range, but with a much shorter dosing regimen covering a critical window of development (gestation days 12-20). A default risk assessment for DBP indicates a reference dose (RfD) of 66 microg/kg/day, based on a LOAEL of 66 mg/kg/day and default factors of 10 for inter-species and inter-individual differences and the lack of a NOAEL. Human exposure data would indicate worst-case scenarios to infants (via formula) in the dose range of the RfD. A default risk assessment appears to be inappropriate since rodents, unlike primates, metabolize phthalate diesters (including DBP) to monoesters extensively in the gut following oral administration. It is believed that the monoester is the active principle for induction of reproductive and developmental toxicity of specific phthalate esters. Thus, if humans produce very low levels of the monoester from an environmental exposure to the diester, the likelihood of any reproductive or developmental toxicity via the oral route appears extremely remote. PMID- 10717379 TI - Future problems requiring scientific consideration. AB - Water is essential for life, a precept that should control all other considerations. Water and its toxicological safety represent a paradigm for much of what is happening in other areas of society in evolving perceptions of what is 'toxicity', what is 'risk' and what we wish to or are prepared to do about such 'risks' at what cost to ourselves and others. Water is an universal solvent and may contain a wide diversity of substances arising from sources and supply systems and modified by treatment and storage. The problems that are already present, and which are closely linked to others just beginning to be recognisable, arise from the growing sensitivity of analytical techniques showing exposure to novel or ever smaller amounts of known substances in water, how to recognize and evaluate their conventional and possibly new (i.e. previously unrealized) toxic actions, how to measure exposure to water as drunk and as in foods and drinks, in what way can safe exposure levels be set, and is it feasible to people to demonstrate that 'safety' has been achieved? Behind those lie the very real problems of deciding what is 'safety', what level of notional safety do we demand in our assessments, and how do we, the people, come to realise and accept the costs that we shall have to pay in demanding any given level of 'safety'? The latter includes deciding what is an appropriate level of safety for the community as a whole and any specially susceptible groups within it. These questions are deceptively simple to pose and tortuously difficult even to attempt to answer. But they are not all the future problems. Behind them lie the socio political uncertainties of risk and its place in society-what real or perceived risks do we accept and why, and how in a democratic society are we, the people, to be informed about risks, how should we decide what to accept or reject, and how are we to balance our beliefs and wishes against the costs of prevention or acceptance of diseases that unclean or contaminated water can bring. PMID- 10717380 TI - Scientific justification of setting limits. PMID- 10717381 TI - Mechanisms and consequences of replication fork arrest. AB - Chromosome replication is not a uniform and continuous process. Replication forks can be slowed down or arrested by DNA secondary structures, specific protein-DNA complexes, specific DNA-RNA hybrids, or interactions between the replication and transcription machineries. Replication arrest has important implications for the topology of replication intermediates and can trigger homologous and illegitimate recombination. Thus, replication arrest may be a key factor in genome instability. Several examples of these phenomena are reviewed here. PMID- 10717382 TI - Repair and replication of oxidized DNA bases using modified oligodeoxyribonucleotides. AB - Modified oligodeoxyribonucleotides (ODNs) are powerful tools to assess the biological significance of oxidized lesions to DNA. For this purpose, we developed original synthetical pathways for the site-specific insertion of several oxidized bases into DNA fragments. Thus, the chemical solid-phase synthesis of ODNs using original strategies of protection and mild conditions of deprotection, as well as a specific post-oxidation approach of an unique nucleoside residue within the sequence have been applied. These two approaches of preparation allowed us to have access to a set of modified ODNs that contain a single modified nucleoside, i.e., N-(2-deoxy-beta-D-erythro pentofuranosyl)formylamine (dF), 5-hydroxy-2'-deoxycytidine (5-OHdCyd), thymidine glycol (dTg), 5,6-dihydrothymidine (DHdThd), 2,2-diamino-4-[(2-deoxy-beta-D erythro-pentofuranosyl)-amino]-5(2H)- oxazolone (dZ), N-(2-deoxy-beta-D-erythro pentofuranosyl)cyanuric acid (dY), 5',8-cyclo-2'-deoxyguanosine (cyclodGuo) and 5',8-cyclo-2'-deoxyadenosine (cyclodAdo). The substrates were used to investigate recognition and removal of the lesions by bacterial DNA N-glycosylases, including endonuclease III (endo III) and Fapy glycosylase (Fpg). In addition, the DNA polymerase-mediated nucleotide incorporation opposite the damage was determined using modified ODNs as templates. PMID- 10717383 TI - The activity of the DNA-dependent protein kinase (DNA-PK) complex is determinant in the cellular response to nitrogen mustards. AB - The DNA-dependent protein kinase plays a critical role in mammalian DNA double strand break (DSB) repair and in specialized recombination, such as lymphoid V(D)J recombination. Its regulatory subunit Ku (dimer of the Ku70 and Ku80 protein) binds to DNA and recruits the kinase catalytic sub-unit, DNA-PKcs. We show here that three different strains deficient in either the Ku80 (xrs-6) or DNA-PKcs (V-3, scid) component of DNA-PK are markedly sensitive (3.5- to 5-fold) to a group of DNA cross-linking agents, the nitrogen mustards (NMs) (melphalan and mechlorethamine) as compared to their parental cell line. Importantly, the level of hypersensitivity to these drugs was close to the level of hypersensitivity observed for radiomimetic agents that create DSBs in DNA (bleomycin and neocarzinostatin). In addition, sensitivity to NMs was restored to the parental level in the xrs-6 cell line stably transfected with the human Ku80 gene (xrs-6/Ku80), showing unequivocally that DNA-PK is involved in this phenotype. These results indicate that a function of the whole DNA-PK protein complex is involved in the cellular response to NMs and suggest that the repair of DNA interstrand cross-links induced in DNA by NMs involved a DNA-PK dependent pathway that shares common features with DNA DSBs repair. PMID- 10717384 TI - DNA rearrangements at the extremities of the Streptomyces ambofaciens linear chromosome: evidence for developmental control. AB - In Streptomyces, a genomic instability results from frequent recombination events which occur at the ends of the linear chromosomal DNA. These events are believed to be responsible for the variability observed in these regions among Streptomyces species and strains. In order to identify functions able to control this type of genome plasticity, mutators as well as mutants produced at different stages of development have been characterized in S. ambofaciens. Their characterization suggests the existence of a relationship between genomic instability and colony development. PMID- 10717385 TI - The ionophore monensin inhibits mouse polyomavirus DNA replication and destabilizes viral early mRNAs. AB - Monensin is a ionophore compound with different biological activities. It raises the intralysosomal pH, it binds the plasma membranes particularly at the level of the cisternal system of the Golgi apparatus. It causes imbalance in the intramembrane ion traffic and inhibits export of secretory proteins at membrane level. Monensin blocks endocytosis and therefore impedes entry of toxic molecules. The drug also inhibits viral proliferation of RNA and DNA viruses such as vesicular stomatitis, influenza and human polyomaviruses. In this report we show that monensin effectively abolishes viral DNA replication of mouse polyomavirus. Results show that the half life of viral early mRNAs is significantly reduced in the presence of the drug. Therefore we suggest that the reduction of viral DNA synthesis is a consequence of the reduced intranuclear pool of viral early antigens. PMID- 10717386 TI - DNA synthesis by CHO cell extracts on fork-like DNA templates containing the major cisplatin adduct requires a ligation step. AB - In this work we have examined the role of DNA ligation in the in vitro replication catalyzed by CHO crude extracts on fork-like oligonucleotide substrates containing a unique d(GpG) intrastrand cross-link produced by the antitumor drug cisplatin. We show here that this reaction involves a ligation step, which necessitates excision of the flap strand of the forked substrate. By constructing substrates in which the unannealed tail could not be degraded by a 5' exonuclease, we obtained evidence suggesting that this type of activity participates in the removal of the flap strand. Furthermore, we found that the ligation event played a predominant role in the synthesis of fully replicated products from both intact and platinated templates. Finally, we investigated whether translesion synthesis of the cisplatin lesion could occur concomitantly to ligation by monitoring the incorporation of labeled precursors downstream of the adduct. Our results are compatible with the possibility that some translesion syntheses of the Pt-d(GpG) adduct by the extracts also contributed to the generation of full length molecules. PMID- 10717387 TI - Loss of the Fanconi anemia group C protein activity results in an inability to activate caspase-3 after ionizing radiation. AB - Fanconi anemia (FA) is a human genetic disease featuring cancer predisposition, genetic instability and DNA damage hypersensitivity. Although abnormalities in DNA repair and cell cycle checkpoint have been proposed as the underlying defect in this syndrome, these hypotheses did not provide full explanations of the complex phenotype. Although not exclusive of such possibilities, alterations in the control of apoptosis might account for the pleiotropic phenotype of this syndrome. We and others have previously reported a deregulation of the apoptotic response to mitomycin C, suggesting that the products of the Fanconi anemia group C protein (FANCC) contribute to the regulation of apoptosis. To explore the functional importance of the apoptotic alterations in FA we analyzed biochemical steps of the execution phase of apoptosis stimulated by another DNA damaging agent, the gamma-ray using FA cell lines derived from complementation group C (FA C) independent patients. It is shown that the poly(ADP-ribose) polymerase, a target of caspase-3, is not cleaved in FA-C after ionizing radiation (IR). Moreover, caspase-3 is not processed in its active form and, its activity is not increased by IR in FA-C cells compared to normal cells. Altogether, these results demonstrate that loss of the FANCC activity results in a deficiency of the IR induced apoptosis which is due to an inability to activate caspase-3. Our work suggests that apoptosis signaling induced by mitomycin C and IR is subject to common regulation involving the FANCC protein. PMID- 10717388 TI - Expression of the Fpg protein of Escherichia coli in Saccharomyces cerevisiae: effects on spontaneous mutagenesis and sensitivity to oxidative DNA damage. AB - The biological relevance of oxidative DNA damage has been unveiled by the identification of genes such as fpg of E. coli or OGG1 of Saccharomyces cerevisiae. Both Fpg and Ogg1 proteins are DNA glycosylases/AP lyases that excise 7,8-dihydro-8-oxoguanine (8-OxoG) and 2,6-diamino-4-hydroxy-5-N methylformamidopyrimidine (Me-FapyG) from damaged DNA. Although similar, the enzymatic and biological properties of Fpg and Ogg1 proteins are not identical. Furthermore, the Fpg and Ogg1 proteins do not show significant sequence homologies. In this study, we investigated the ability of the Fpg protein of E. coli to complement phenotypes thought to be due to oxidative DNA damage in Saccharomyces cerevisiae. To express Fpg in yeast, the coding sequence of the fpg gene was placed under the control of a strong yeast promoter in the expression vector pCM190 to generate the pFPG240 plasmid. The Ogg1-deficient yeast strain CD138, ogg1::TRP1, was transformed with pFPG240 and the expression of Fpg was measured. Expression of Fpg in yeast harboring pFPG240 was revealed by efficient release of Me-FapyG and cleavage of 8-OxoG-containing duplexes by cell free protein extracts. The production of the Fpg protein in yeast cells was further demonstrated by immunoblotting analysis using anti-Fpg antibodies. Fpg expression suppresses the spontaneous mutator phenotype of ogg1- yeast for the production of canavanin resistant mutants (CanR) and Lys+ revertants. Fpg expression also restores the capacity of plasmid DNA treated with methylene blue plus visible light (MB-light) to transform the yeast ogg1- rad1- double mutant. PMID- 10717389 TI - Structure of an oligonucleotide containing a N-(2-deoxy-beta-D-erythro pentofuranosyl)formamide residue facing a guanine. AB - Formamide residue is a major oxidative DNA damage product from ionizing radiation on thymine residues in DNA. We report NMR and molecular modeling studies on a DNA duplex structure which contains guanine opposite formamide residue. Formamide residue exists as either the cis and trans isomer. For the trans and the cis isomers, we find that guanine and formamide are stacked inside the helix and are hydrogen bonded. The oligonucleotide adopts globally a B form structure for the two isomers. Conformational changes are observed between the two isomers. PMID- 10717390 TI - Characterization of genetic interactions with RFA1: the role of RPA in DNA replication and telomere maintenance. AB - Replication protein A (RPA) is a heterotrimeric single-stranded DNA binding protein whose role in DNA replication, recombination and repair has been mainly elucidated through in vitro biochemical studies utilizing the mammalian complex. However, the identification of homologs of all three subunits in Saccharomyces cerevisiae offers the opportunity of examining the in vivo role of RPA. In our laboratory, we have previously isolated a missense allele of the RFA1 gene, encoding the p70 subunit of the RPA complex. Strains containing this mutant allele, rfa1-D228Y, display increased levels of direct-repeat recombination, decreased levels of heteroallelic recombination, UV sensitivity and a S-phase delay. In this study, we have characterized further the role of RPA by screening other replication and repair mutants for a synthetic lethal phenotype in combination with the rfa1-D228Y allele. Among the replication mutants examined, only one displayed a synthetic lethal phenotype, pol12-100, a conditional allele of the B subunit of pol alpha-primase. In addition, a delayed senescence phenotype was observed in raf1-D228Y strains containing a null mutation of HDF1, the S. cerevisiae homolog of the 70 kDa subunit of Ku. Interestingly, a synergistic reduction in telomere length observed in the double mutants suggests that the shortening of telomeres may be the cause of the decreased viability in these strains. Furthermore, this result represents the first evidence of a role for RPA in telomere maintenance. PMID- 10717391 TI - Heavy ion-induced plasmid DNA damage in aerated or deaerated conditions. AB - Using the plasmid relaxation assay, the induction of single strand breaks (SSB) and base damages was investigated in air-dried plasmid DNA irradiated under air or under vacuum, with two high LET particles. We first observed that an irradiation with 12C5+ ion produced less of both damages when performed in a vacuum rather than in the presence of air. This could be due to the presence of O2 which increases the primary radicalar effects in the latter case. Another explanation is a difference in the degree of hydration of the DNA molecules. Indeed, under vacuum only the water molecules tightly bound to DNA will persist. In contrast, in the presence of air, the outer hydration shell enhances the amount of hydroxyl radicals available for the radiolytic attack. However, no difference in the SSB induction was observed when DNA was irradiated with 36S16+ ion in the presence of air or under vacuum. This is likely due to the LET effect which partly cancels the production of radicals by recombination and increases the formation of superoxide anions in the track. Similarly, the lower induction of damage by 36S16+ irradiation in comparison with the 12C5+ ion is a consequence of the higher ionizing density for 36S16+ than for 12C5+ ions. Meanwhile, for both ions, base damages are not detected when DNA is irradiated under vacuum, whereas they are as frequent as SSB when irradiation is performed in the presence of air. Altogether, these observations support the idea that SSB and base damage are not formed by the same mechanism. PMID- 10717392 TI - The L17 ribosomal protein of Bacillus subtilis binds preferentially to curved DNA. AB - We searched in Bacillus subtilis for proteins that bind preferentially to curved DNA. Two proteins of 9 and 15 kDa displaying this property were purified from exponentially growing cells of B. subtilis strain 168. Sequencing of N-terminal amino acids identified them as the proteins HBsu and L17 respectively. The overproduction of L17 from B. subtilis in Escherichia coli was shown to have a strong effect on nucleoid morphology and segregation. PMID- 10717393 TI - Connections of sympathetic fibres inside the cavernous sinus: a microanatomical study. AB - A microanatomical study has been designed to investigate the pattern of arrangement of the sympathetic fibres inside the cavernous sinus. The course of these fibres has been examined in 60 fresh specimens of parasellar region from autopsy cadavers. Apart from the thin branches arising at different intervals along its course, the sympathetic plexus of the carotid artery gives rise to a large division that usually joins the abducens nerve and leaves it to combine with the ophthalmic branch of the fifth. In 10% of specimens we have found a direct connection between sympathetic fibres and the ophthalmic branch of the trigeminal nerve. We did not recognise similar connections with oculomotor and trochlear nerves. PMID- 10717394 TI - Elimination of phenytoin in toxic overdose. AB - Phenytoin is widely used for the management of seizures. Fortunately overdosage with this drug is rare. We performed a prospective study to investigate the elimination kinetics of phenytoin in toxic overdose. All patients were only on phenytoin and not on other anticonvulsants. Phenytoin toxicity was defined by clinical features and correlated with drug levels. Daily phenytoin levels were obtained until they were less than or equal to 15 mcg/ml. Nine patients with age ranging from 20 to 66-years-old were recruited. Sex ratio was male:female, 5:4. Initial phenytoin levels ranged from 34 to 57.5 mcg/ml. Serum phenytoin levels of three patients remained relatively constant for 2-5 days before declining in a steady fashion. Phenytoin levels of the remaining patients declined in an almost linear manner. Regression analysis of all patients showed that the slope terms were highly significant (with low P-values) and corresponding R(2) values were close to 1. Different patients have different rates of metabolism; in seven of nine patients, levels declined between 4.6 and 5.9 mcg/ml per day. Knowledge of the rate of elimination assists the clinician in deciding on the best time to reinstitute phenytoin therapy. PMID- 10717395 TI - Contrast enhanced color duplex for diagnosis of subtotal stenosis or occlusion of the internal carotid artery. AB - BACKGROUND AND PURPOSE: We initiated this prospective study to investigate the usefulness of contrast enhancement in combination with color Doppler-assisted duplex imaging (CDDI) for the distinction of subtotal internal carotid artery (ICA) stenosis and ICA occlusion. METHODS: During 1 year all patients with a previously unknown subtotal ICA stenosis (>90%) or ICA occlusion on routine CDDI were included in the study. These patients underwent a CDDI with and without intravenous contrast, Levovist 300 mg/ml. RESULTS: The study group consisted of 32 patients, 15 with subtotal stenosis and high velocity at the ICA stenosis, two with subtotal stenosis and minimal residual color flow and relative low velocity at the ICA stenosis and 15 with ICA occlusion. In all patients the diagnosis by CDDI without and with contrast were the same. Image quality was improved with contrast in 13 of the 17 patients at the subtotal ICA stenosis. There was no significant difference in mean velocities at the subtotal ICA stenoses without and with contrast. CONCLUSION: The usefulness of contrast enhancement with CDDI for differentiating subtotal ICA stenosis and ICA occlusion is limited. Possibly it is useful in patients with moderate image quality of the CCA and ICA and in patients with a subtotal stenosis with minimal residual color flow and relative low velocity at the ICA stenosis. PMID- 10717396 TI - Persistent hiccup as presenting symptom in medulla oblongata cavernoma: a case report and review of the literature. AB - A rare case of persistent intractable hiccup as presenting symptom of cavernous angioma in the medulla oblongata is reported. Pathophysiologic hypotheses about the triggering mechanism of hiccup are discussed, with special reference to the causes affecting the central nervous system. A review of the literature concerning medullary lesions presenting with persistent hiccup is also reported. Finally we have included some brief considerations about cavernous angiomas and the patterns of their clinical presentation, focusing on those located in the medulla oblongata. PMID- 10717397 TI - Basilar artery occlusion due to mucormycotic emboli, preceded by acute hydrocephalus. AB - A rare case of brain stem infarction caused by mucormycotic emboli, preceded by acute hydrocephalus, is reported. The patient, who had suffered from leukemia and had undergone bone marrow transplantation several months before, presented initially with seizure and persistent disturbance of consciousness. A head CT scan revealed marked ventricular dilation and diagnosed as acute hydrocephalus. The patient received emergent ventricular drainage. Despite the aggressive treatment, the patient did not survive. Autopsy revealed systemic mucormycosis occluding and invading various arteries including basilar artery and its branches, causing fatal brainstem infarction. Although early diagnosis remains difficult in the cases of systemic mucormycosis, prompt initiation of treatment is mandatory; one must have in mind the possibility of presence of fungal infection when treating patients with acute neurological deterioration who have underlying debilitating diseases, even though fungi themselves are hard to detect in most cases. PMID- 10717398 TI - Predicting outcome from coma: man-in-the-barrel syndrome as potential pitfall. AB - The Glasgow coma scale motor score is often used in predicting outcome after hypoxic-ischemic coma. Judicious care should be exerted when using this variable in predicting outcome in patients with coma following hypotension since borderzone infarction can obscure the clinical picture. We describe a patient who underwent skull base surgery for a schwannoma of the left facial nerve. The operation, which lasted for 10 h, was conducted under controlled hypotension. After the intervention the patient remained comatose with absent arm movements upon painful stimuli. An absent motor score usually carries a poor prognosis. However, magnetic resonance inversion recovery imaging of the brain showed bilateral hyperintense lesions in the arm-hand area indicative of borderzone ischemic damage. The patient received optimal supportive care and after 17 days he regained consciousness with 'man-in-the-barrel syndrome', which also further improved over time. PMID- 10717399 TI - Cerebral herniation after lumbar puncture in sarcoid meningitis. AB - A patient with chronic meningitis due to neurosarcoidosis became comatose within minutes of a lumbar puncture and died 24 h later. The diagnosis of neurosarcoidosis was made post mortem. Development of cerebral herniation may have been exacerbated by lumbar puncture. It was proposed that arachnoid villi dysfunction may have contributed to very high intracranial pressures in this patient, since post mortem examination revealed communication between the ventricles and outlet foramina of the fourth ventricle, and that herniation was in part due to an acute pressure differential caused by lumbar puncture. PMID- 10717400 TI - Subarachnoid haemorrhage following rupture of an ophthalmic artery aneurysm presenting as traumatic brain injury. AB - Head trauma may provoke subarachnoid haemorrhage. The question sometimes arises whether in patients with trauma and subarachnoid haemorrhage the latter is of traumatic or aneurysmal origin. We present a 49-year-old patient who fell from a truck, struck his head and was unconscious immediately. On the brain computed tomography (CT) scan subarachnoid haemorrhage was present, initially diagnosed as of traumatic origin. Four-vessel angiography revealed rupture of a left ophthalmic artery aneurysm. We review the literature and give recommendations for angiography in patients with trauma and subarachnoid haemorrhage. PMID- 10717401 TI - Sagittal sinus thrombosis associated with transient free protein S deficiency after L-asparaginase treatment: case report and review of the literature. AB - Cerebral sinus thrombosis associated with acquired free protein S deficiency is very rare. We report the case of an adult patient with acute lymphoblastic leukemia who presented with repeated transient ischemic attacks followed by a seizure during consolidation treatment with L-asparaginase. Magnetic resonance of the brain showed a small cortical hemorrhagic infarct. Superior sagittal sinus thrombosis was demonstrated by cerebral angiogram. A marked decrease of the free form of protein S was documented. One month later, when the patient was free of symptoms, the follow-up free protein S antigen level was restored to the normal range. We suggest that the sagittal sinus thrombosis in this patient was caused by acquired, transient free protein S deficiency. This case also extends the clinical spectrum of cerebral sinus thrombosis to include recurrent transient ischemic attacks alternating with seizures. PMID- 10717402 TI - Acute cerebellar ataxia of a patient with SLE. AB - We described a 28-year-old woman with systemic lupus erythematosus (SLE) presented with digestive tract, skin and renal symptoms and afterwards developed acute cerebellar ataxia, a paresis of the right inferior rectus muscle, left abducens paralysis and left facial palsy which seemed to be consistent with a brainstem lesion visible on magnetic resonance imaging (MRI). This lesion disappeared within 9 days of corticosteroid treatment. It is suggested that this lesion is focal edema induced by acute changes in the blood brain barrier secondary to a vasculopathy. Other causes, including local infarction, are unlikely. PMID- 10717403 TI - Cellular dumbbell schwannoma of the hypoglossal nerve presenting without hypoglossal nerve palsy. AB - A rare case of cellular schwannoma of the hypoglossal nerve, with intraspinal extension, presenting without any recognisable impairment of the function of the hypoglossal nerve is presented. PMID- 10717404 TI - Metastatic spinal intramedullary germinoma with elevated cerebrospinal fluid chorionic gonadotropin: a case report. AB - We treated a patient whose unusual recurrent germinoma illustrates the diagnostic value of measuring human chorionic gonadotropin beta subunit (HCG-beta) in cerebrospinal fluid (CSF) and serum. A 25-year-old man with a suprasellar germinoma and ventricular dissemination was treated successfully with systemic chemotherapy and cranial irradiation. Six years later he developed progressive numbness and weakness in both upper extremities. Magnetic resonance imaging (MRI) disclosed an intramedullary spinal cord tumor in the cervical region. The CSF concentration of HCG-beta was elevated and exceeded that in serum. After completion of systemic chemotherapy and spinal irradiation, symptoms subsided and the tumor was no longer evident on MRI. Based on the patient's history and the rapid response of the tumor to treatment, the spinal cord tumor was considered a metastatic intramedullary spinal germinoma representing CSF dissemination via the central canal. PMID- 10717405 TI - The treatment with alendronate in hemifacial spasm associated with Paget's disease of bone. AB - The association of Paget's disease of bone and hemifacial spasm has rarely been reported. Hemifacial spasm is often associated with compression of the facial nerve by a vascular loop at the point where the nerve leaves the brainstem before traversing the cerebellopontine angle. It is believed that narrowing of the cerebellopontine angle cistern caused by Paget's disease increases the chance of vascular compression of the facial nerve. Whilst specific antipagetic therapy such as calcitonin has been used with good response in hemifacial spasm associated with Paget's disease, the usefulness of the newer bisphosphonates is not clear. A 65-year-old woman with hemifacial spasm associated with Paget's disease was treated with alendronate, and the hemifacial spasm became very infrequent 4 months after commencement of the therapy. PMID- 10717406 TI - Cerebellar neuroblastoma in an infant. AB - A cerebellar neoplasm in an 8-month-old boy is reported. While this tumour was composed of small cells and had regions resembling desmoplastic medulloblastoma, it showed ultrastructural neuronal characteristics including bundles of microtubules in the cell processes, numerous synaptic vesicles, and occasional abortive or complete synapses. These characteristic features warranted the diagnosis of a neuroblastoma of the cerebellum. The nature of this rare intraparenchymal tumour in infants is also briefly discussed. PMID- 10717407 TI - A cortical motor region that represents the cutaneous back muscles in the macaque monkey. AB - A cortical motor region that represented the cutaneous muscles on the back was identified on the medial wall of the frontal lobe in the macaque monkey. In this region, neurons responded to somatosensory stimuli such as light touch or squeezing of the back skin, and intracortical microstimulation elicited contraction of the back skin. Such a region was located primarily on the dorsal bank of the cingulate sulcus, corresponding to the dorsal cingulate motor area. PMID- 10717408 TI - N-Methyl-D-aspartate receptor NR1 subunit mRNA level was decreased in rat brain during pentobarbital withdrawal. AB - We examined the effect of continuous intracerebroventricular (i.c.v. ) administration of pentobarbital on the level of N-Methyl-D-aspartate receptor NR1 subunit mRNAs in different regions of rat brain by in situ hybridization histochemistry. Animals were rendered tolerant to pentobarbital by continuous i.c.v. infusion (300 microgram/10 microliter/h for 6 days) through pre-implanted cannulae connected to osmotic mini-pumps, and dependent, by abrupt withdrawal from pentobarbital. NR1 subunit mRNA level was significantly decreased in the entorhinal cortex, cingulate, caudate-putamen, septum, and the CA1 of hippocampus in pentobarbital withdrawal rats. The level of NR1 mRNAs was decreased in only CA1 of hippocampus in pentobarbital tolerant rats. These results indicate that withdrawal from i.c.v. infusion with pentobarbital decreases in NR1 subunit expression in the rat brain, suggesting that changes in expression of NR1 subunit may contribute to the development of withdrawal from pentobarbital. PMID- 10717409 TI - The correlation between P300 alterations and regional cerebral blood flow in non demented Parkinson's disease. AB - P300 was evoked by a visual oddball and an S1-S2 task in 22 non-demented Parkinson's disease (PD) patients (13 in the early stage, nine in the late stage) and 18 normal controls. Reaction time was also measured. All patients undertook the (99)Tc-ECD SPECT examination. Quantitative regional cerebral blood flow (rCBF) was obtained by overlying SPECT image on the 3D-magnetic resonance image. In the PD patients in the late stage, P300 latency to S2-same and reaction time were significantly prolonged, while rCBF at bilateral frontal, temporal, and the right parietal lobes was decreased. P300 latency to S2-same was significantly correlated with the rCBF at bilateral temporal lobes. Reaction time was significantly correlated with the rCBF at the right frontal and parietal lobes, as well as the temporal and occipital lobes. The results suggest that P300 changes in non-demented PD in the late stage could be related to the temporal lobe dysfunction. PMID- 10717410 TI - Plasminogen enhances the secretion of plasminogen activator inhibitor-1 from cultured rat astrocytes. AB - To investigate the physiological significance of microglia-derived plasminogen (PGn) in the central nervous system, we determined the effects of rat PGn on the secretion of plasminogen activator inhibitor (PAI) in cultured rat astrocytes by reverse zymography and Western blotting. The cultured astrocytes normally produce only a limited amount of PAI-1. After stimulation with PGn, however, the amount of active PAI-1 in the astrocyte-conditioned medium was significantly increased in a time-dependent manner. PGn was also proved to activate p38 MAP kinase and c Jun N-terminal kinase in these astrocytes. Taken together, these results suggest that microglia-derived PGn increases the secretion of PAI-1 in astrocytes through the activation of MAP kinases, and that enhanced PAI-1 regulates various physiological phenomena including tissue remodeling, neuronal plasticity and neurotoxicity. PMID- 10717411 TI - Intracellular Ca(2+) signaling is required for neurotrophin-induced potentiation in the adult rat hippocampus. AB - Recent studies have demonstrated the importance of neurotrophin function in adult synaptic plasticity. In an effort to characterize the intracellular signaling pathways that couple Trk receptor activation to the final physiological effects of neurotrophins, we have examined the role of intracellular calcium rises in neurotrophin-induced synaptic enhancement in hippocampal slices. Using pharmacological blockers to two different calcium ion (Ca(2+)) sources, voltage gated Ca(2+) channels and intracellular Ca(2+) stores, we show that the potentiating effects of neurotrophins in hippocampal slices are mediated by intracellular Ca(2+) signaling. Although basal synaptic transmission between hippocampal CA3 and CA1 neurons was not affected by nifedipine or thapsigargin, both drugs significantly attenuated brain-derived neurotrophic factor or neurotrophin-3-induced synaptic enhancement. The pharmacological blockade of Ca(2+) signaling is effective only during the initial period of neurotrophin induced potentiation. These data suggest that the minimal requirements for inducing potentiation by neurotrophins involve a transient increase in intracellular Ca(2+) concentration, via voltage-gated Ca(2+) channels and/or intracellular Ca(2+) stores. PMID- 10717412 TI - Neural substrates for depth perception of the Necker cube; a functional magnetic resonance imaging study in human subjects. AB - We have studied the cerebral activity for the depth perception of the Necker cube by functional magnetic resonance imaging. Three types of line drawing figures were used as stimuli, the Necker cube, hidden line elimination cube and overlapping squares. Subjects were instructed to perceive both orientations of the depth of the Necker cube. They were instructed to shift their attention voluntarily during viewing overlapping squares to obtain a control for the attentional shift in perceiving the Necker cube. A hidden line elimination cube was used as a control for monocular stereopsis. The results showed a clear symmetrical activation in premotor and parietal areas during the Necker cube perception compared with other conditions. The present result suggests that a neural process similar to mental image manipulation occurs during depth perception of the Necker cube. PMID- 10717413 TI - Detection of complement factor B in the cerebrospinal fluid of patients with cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy disease using two-dimensional gel electrophoresis and mass spectrometry. AB - Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is a hereditary condition with onset in mid adulthood and is associated with mutations in the Notch-3 gene. (Joutel, A., Corepechot, C., Ducros, A., Vahedi, K., Chabriat, H., Mouton, P., Alamowitch, S., Domenda, V., Cecilion, M., Marechal, J., Vayssiere, C., Cruaud, C., Cabanis, E.A., Ruchoux, M.M. , Weissenvach, J., Bach, J.F., Bousser, M.G. and Tournier Lasserve, E., Notch3 mutations in CADASIL, a hereditary adult-onset condition causing stroke and dementia. Nature, 383 (1996) 707-710) Ultrastructural examination of the pathology of the cerebral infarcts reveals deposits in the vascular smooth muscle cells of the small arteries of the brain, but there is no obvious indication how the Notch-3 mutations give rise the observed pathology, nor is there any information on the exact nature of the deposits. We have investigated cerebrospinal fluid (CSF) from three CADASIL cases with known mutations in Notch-3 using two-dimensional gel electrophoresis. CSF from these patients was compared to that of six controls. We detected a single spot in the protein maps of patients which was absent from all the controls. In-gel tryptic digestion of this protein followed by mass spectrometric analysis of the tryptic fragments and a database search identified the spot as human complement factor B. These preliminary findings suggest that the alternative complement pathway may play a role in the pathogenesis of CADASIL. PMID- 10717414 TI - P2 Purinoceptor expression and functional changes of hypoxia-activated cultured rat retinal microglia. AB - P(2) purinoceptors appear to modulate microglia function, but their role in hypoxic microglia has not been investigated. We examined in postnatal rat retinal microglia cultured under hypoxic (1% oxygen) condition, their P2 expression, proliferation and cytokine release in the presence or absence of the P2 receptor agonists and antagonists. Fura-2 fluorescence measurements of intracellular Ca(2+) rises to P2 receptor agonists and antagonists indicated that both P(2U) and P(2Z) were expressed in hypoxic microglia. Hypoxia induced BrdU incorporation and release of interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF alpha) as well. The P(2U) agonist, UTP, maintained the BrdU incorporation, whereas the P(2Z) agonist, BzATP, suppressed it, but significantly enhanced IL 1beta and TNF-alpha release, suggesting that the P(2U) response may underlie the mitotic activity, and that of P(2Z), the IL-1beta and TNF-alpha release of hypoxia-activated microglia. PMID- 10717415 TI - Neural activity in the horizontal limb of the diagonal band of broca can be modulated by electrical stimulation of the olfactory bulb and cortex in rats. AB - Previously published theoretical models of olfactory processing suggest that cholinergic modulatory inputs to the olfactory system should be regulated by neural activity in the olfactory bulb. We tested these predictions using in vivo electrophysiology in rats. We show that the activity of approximately 20% of neurons recorded in the horizontal limb of the diagonal band of Broca (HDB), which is the source of cholinergic projections to the olfactory system, can be modulated by electrical stimulation of either the lateral olfactory tract or the cell body layer of piriform cortex. These data suggest a possible physiological pathway for the proposed regulation of neural activity in the HDB by activity in the olfactory bulb or cortex. PMID- 10717416 TI - Selective MT(2) melatonin receptor antagonist blocks melatonin-induced antinociception in rats. AB - The present study was undertaken to assess the effects of intracerebroventricular (i.c.v.) luzindole (a selective MT(2) melatonin receptor antagonist) and prazosin (a selective MT(3) melatonin receptor antagonist) on melatonin-induced antinociception, so as to clarify which of melatonin receptor subtypes within the central nervous system (CNS) was mediating antinociception. The pain threshold of rats was measured by the hot water (50 degrees C) tail-flick test. It was found that intraperitoneal (i.p.) melatonin (30, 60, 120 mg/kg) resulted in a dose dependent antinociception. Luzindole (50, 100 microgram) administered intracerebroventricularly antagonized significantly the antinociceptive effect induced by i.p. melatonin (120 mg/kg), whereas prazosin (50 microgram) did not. Neither luzindole (100 microgram, i.c.v.) nor prazosin (50 microgram, i.c.v.) affected the nociceptive threshold when given alone. The results suggest that melatonin-induced antinociception is mediated through the MT(2) melatonin receptor subtype within the CNS. PMID- 10717418 TI - Up-regulation of sigma(1) receptor mRNA in rat brain by a putative atypical antipsychotic and sigma receptor ligand. AB - Sigma(1) (sigma(1)) receptor mRNA expression was studied in the prefrontal cortex, striatum, hippocampus and cerebellum of rat brain by northern blot and in situ hybridization. The effects of a chronic treatment with antipsychotic drugs (haloperidol and clozapine), and with E-5842, a sigma(1) receptor ligand and putative atypical antipsychotic on sigma(1) receptor expression were examined. A significant increase in the levels of sigma(1) receptor mRNA in the prefrontal cortex and striatum after E-5842 administration was observed, while no apparent changes were seen with either haloperidol or clozapine. Our results suggest a long-term adaptation of the sigma(1) receptor at the level of mRNA expression in specific areas of the brain as a response to a sustained treatment with E-5842. PMID- 10717417 TI - Bis(7)-tacrine, a novel acetylcholinesterase inhibitor, reverses AF64A-induced deficits in navigational memory in rats. AB - The novel dimer bis(7)-tacrine (1,7-N-Heptylene-bis-9,9'-amino-1,2,3, 4 tetrahydroacridine), which exhibits higher potency, selectivity and oral activity on acetylcholinesterase inhibition in vivo than tacrine, was evaluated for its ability to reverse AF64A-induced spatial memory impairment in rats using the Morris water maze. The intracerebroventricular injection of AF64A (3 nmol/side) resulted in a substantial increase in the escape latency to find the platform (F(1,7)=30.2, P<0.01). The observed impairment of spatial memory was paralleled by a 47% decrease in choline acetyltransferase activity in the hippocampus. Oral administration of bis(7)-tacrine (0.22-0.89 micromol/kg) dose-dependently reversed the AF64A-induced latency delay to the level of the saline control group (F(4,28)=7.45, P<0. 05). The present study provides additional evidence of bis(7) tacrine as an ideal candidate for the palliative treatment of Alzheimer's disease. PMID- 10717419 TI - Up-regulation of type II adenylyl cyclase mRNA in kindling model of epilepsy in rats. AB - The expression level of type II adenylyl cyclase mRNA (ACII) was analyzed by northern blotting in amygdaloid kindled rats. Remarkable increases in ACII mRNA were observed in the bilateral cerebral cortex and hippocampus at 24 h after the last generalized seizure. The elevated expression level in the hippocampus persisted for 4 weeks on the stimulated side. There were no changes in expression level in single-stimulated and partially-kindled states. These results suggest that the involvement of ACII might have an effect on the mechanisms of seizure generalization and the maintenance of persistent epileptogenesis rather than on the acquisition process. PMID- 10717420 TI - Percentage incidence of gamma-aminobutyric acid neurons in the claustrum of the rabbit and comparison with the cortex and putamen. AB - We describe the incidence of gamma-aminobutyric acid (GABA)ergic neurons after post-embedding immunocytochemistry on semithin sections of the claustrum, putamen and lateral, dorsal and medial cortical areas. Twelve percent of the neurons counted in the claustrum of 11 rabbits were GABAergic. This incidence was significantly higher in the dorsal halves of both the insular and endopiriform claustra than in the ventral (13 vs. 10%). The incidence of GABAergic cells was 4% in the putamen, 14% in the insular cortex, 15% in areas 17 and 18 and 13% in area 29d. Thus, our results indicate that in contrast to the putamen the incidence of GABAergic cells was similar in the claustrum and cortical areas. We interpret this in the light of the pallial origin of the claustrum, which has recently been substantiated. PMID- 10717421 TI - Somatosensory evoked potentials to a threshold air-puff can predict stimulus detection in human subjects. AB - Somatosensory evoked potentials (SEPs) and reaction time (RT) were studied following a threshold intensity air-puff stimulation of the skin in nine subjects. We delivered brief air-puffs, at the threshold intensity of about 50% detection probability, to the volar surface of the right hand at the base of the index finger. The SEPs were averaged separately for detected and undetected stimuli. Prior to the SEP data collection, RTs to the same intensity threshold stimulus were measured. P300 SEP late component was recorded in response to detected stimuli and was not to undetected stimuli. However, in one subject, P300 was associated with undetected stimuli, suggesting errors in the psychophysical task. The peak latency of P300 was 316+/-29 ms (mean+/-SD) and earlier than the average RT (390+/-43 ms). These results demonstrate that SEPs can correctly predict perception of the threshold air-puff stimulus. PMID- 10717423 TI - Quantitative changes in interleukin proteins following focal stroke in the rat. AB - The aim of the present study was to quantitate the temporal changes in protein concentration for interleukin (IL)-1alpha, IL-1beta, IL-1ra, and IL-6 from 1 h to 15 days following focal ischemia. Protein expression was evaluated by enzyme linked immunosorbent assay utilizing newly available rat antibodies. There were no detectable basal levels of IL-1alpha, 1L-1beta, or IL-6 in the sham-operated or non-ischemic control cortex. IL-1beta (increased significantly (P<0.05) as early as 4 h and peaked at 3 to 5 days. IL-1alpha (increased significantly (P<0.05) at 3 days. IL-6 increased early and peaked at 24 h (P<0.05). IL-1ra increased significantly (P<0.05) over basal levels from 12 h to 5 days. The present study provides the first quantitative determination of interleukin protein concentrations in the rat brain following focal stroke and demonstrates that this technology is now available for mechanistic studies in neuroprotection. PMID- 10717422 TI - Adenosine triphosphate-induced peripheral nerve discharges generated from the cat petrosal ganglion in vitro. AB - Since nucleotides have been postulated as transmitters between glomus cells and chemosensory nerve endings in the carotid body, we studied the effects of their application to the petrosal ganglion, where the perikarya of carotid (sinus) nerve are located. Cat petrosal ganglia were superfused in vitro, while electrical activities of their peripheral processes (carotid nerve and glossopharyngeal branch) were recorded simultaneously. Adenosine triphosphate (ATP) evoked dose-dependent bursts of impulses in carotid nerve, while those in glossopharyngeal branch were less intense and consistent. Adenosine monophosphate was less effective than ATP. ATP-induced carotid nerve responses presented no temporal desensitization and persisted after applying P(2Y) receptor blocker Reactive Blue 2 to the ganglion. The results indicate that ATP has an excitatory effect on the perikarya of the population of petrosal ganglion neurons projecting peripherally through the carotid nerve. PMID- 10717424 TI - Neuroimmune relations in patients with fibromyalgia: a positron emission tomography study. AB - To study relations between neural and immune activity in patients with chronic pain, we correlated regional cerebral blood flow measured with [(15)O]butanol positron emission tomography to immune function in five patients with fibromyalgia. Partly replicating previous data in healthy volunteers, natural killer cell activity correlated negatively with right hemisphere activity in the secondary somatosensory and motor cortices as well as the thalamus. Moreover, natural killer cell activity was negatively and bilaterally related to activity in the posterior cingulate cortex. Thus, immune parameters were related to activity in brain areas involved in pain perception, emotion, and attention. Implicated from a small study population, these strong neuro-immune associations are discussed in view of recent findings concerning mechanisms and adaptive values in immuno-cortical communication and regulation. PMID- 10717425 TI - Gender differences in rat neuropathic pain sensitivity is dependent on strain. AB - It is well recognized that gender differences play a major role in pain sensitivity, pain report, analgesic efficacy and prevalence of certain chronic pain disorders. In the present study we sought to determine whether male or female rats of two different outbred strains (Sprague-Dawley and Holtzman) experienced differential pain sensitivity after the same mononeuropathy lesion. Following baseline mechanical allodynia testing, rats of each sex and strain underwent an L5 spinal nerve transection. Mechanical allodynia using 2 and 12 g von Frey filaments was assessed at days 1, 3, 5, 7, and 10 post surgery. There were no statistically significant differences in allodynia between gender in the Holtzman strain or between strains. However, mechanical allodynia was significantly greater in female Sprague-Dawley rats as compared with males following a spinal nerve transection. These data suggest that the choice of rat gender and strain should be considered in experimental neuropathic pain studies, especially in the assessment of potential analgesics. PMID- 10717426 TI - Time of day determines modulation of synaptic transmission by adenosine in the rat hippocampal slices. AB - Adenosine, an endogenous modulator of synaptic transmission, has been implicated in regulation of sleep and arousal. The effect of adenosine on neuronal excitability depends on its concentration in the extracellular space. The present study shows that the state of activity of laboratory rats determines the level of tonic inhibition by adenosine in hippocampal slices prepared from these animals. Thus, slices taken at the end of the active period showed significantly more inhibition by adenosine, as determined by the effects of the A1 receptor blocker 8-CPT, in comparison to slices taken in the inactive state. The results support the proposed role of adenosine in regulation of sleep and arousal and point to the importance of the time of day at which slices for electrophysiological experiments are prepared. PMID- 10717427 TI - The connections of the dopaminergic system with the striatum in rats and primates: an analysis with respect to the functional and compartmental organization of the striatum. AB - This Commentary compares the connections of the dopaminergic system with the striatum in rats and primates with respect to two levels of striatal organization: a tripartite functional (motor, associative and limbic) subdivision and a compartmental (patch/striosome-matrix) subdivision. The topography of other basal ganglia projections to the dopaminergic system with respect to their tripartite functional subdivision is also reviewed. This examination indicates that, in rats and primates, the following observations can be made. (1) The limbic striatum reciprocates its dopaminergic input and in addition innervates most of the dopaminergic neurons projecting to the associative and motor striatum, whereas the motor and associative striatum reciprocate only part of their dopaminergic input. Therefore, the connections of the three striatal subregions with the dopaminergic system are asymmetrical, but the direction of asymmetry differs between the limbic versus the motor and associative striatum. (2) The limbic striatum provides the main striatal input to dopamine cell bodies and proximal dendrites, with some contribution from a subset of neurons in the associative and motor striatum (patch neurons in rats; an unspecified group of neurons in primates), while striatal input to the ventrally extending dopamine dendrites arises mainly from a subset of neurons in the associative and motor striatum (matrix neurons in rats; an unspecified group of neurons in primates). (3) Projections from functionally corresponding subdivisions of the striatum, pallidum and subthalamic nucleus to the dopaminergic system overlap, but the specific targets (dopamine cells, dopamine dendrites, GABA cells) of these projections differ. Major differences include the following. (1) In rats, neurons projecting to the motor and associative striatum reside in distinct regions, while in primates they are arranged in interdigitating clusters. (2) In rats, the terminal fields of projections arising from the motor and associative striatum are largely segregated, while in primates they are not. (3) In rats, patch- and matrix-projecting dopamine cells are organized in spatially, morphologically, histochemically and hodologically distinct ventral and dorsal tiers, while in primates there is no (bi)division of the dopaminergic system that results in two areas which have all the characteristics of the two tiers in rats. Based on the anatomical data and known dopamine cell physiology, we forward an hypothesis regarding the influence of the basal ganglia on dopamine cell activity which captures at least part of the complex interplay taking place within the substantia nigra between projections arising from the different basal ganglia nuclei. Finally, we incorporate the striatal connections with the dopaminergic system into an open-interconnected scheme of basal ganglia-thalamocortical circuitry. PMID- 10717428 TI - Changes in neuronal conductance during different components of cortically generated spike-wave seizures. AB - Neuronal conductance was studied in anesthetized cats during cortically generated spike-wave seizures arising from slow sleep oscillation. Single and dual intracellular recordings from neocortical neurons were used. The changes were similar whether the seizures occurred spontaneously, or were evoked by electrical stimulation or induced by bicuculline. In all seizures, the conductance increased from the very onset of the seizure and returned to control values only at the end of the postictal depression. Simultaneous intracellular recordings from two neurons showed that the neuron leading the other neuron displayed the largest increase in membrane conductance. The changes in neuronal conductance during the two phases of the slow sleep oscillation, i.e. highest during depolarizations and lowest during hyperpolarizations, were similar to those occurring during the "spike" and "wave" components of seizures. (1) Maximal conductance was found during the paroxysmal depolarizing shift corresponding to the electroencephalogram "spike" (median: 252 nS; range: 90 to more than 400 nS). It was highest at the onset of the depolarized plateau and decreased thereafter. (2) During the hyperpolarization corresponding to the electroencephalogram "wave", the conductance was significantly lower (median: 71 nS; range: 41 to 140 nS). (3) The conductance was elevated during the fast runs (median: 230 nS; range: 92 to 350 nS) which occurred in two-thirds of the seizures. (4) The conductance values during postictal depression were situated between those measured during the seizure hyperpolarizations and during sleep hyperpolarizations. The conductance decreased exponentially back to the values of the slow sleep oscillation over the total duration of the postictal depression. The data suggest that the major mechanism underlying the "wave"-related hyperpolarizing component of spike-wave seizures relies mainly not on active inhibition, but on a mixture of disfacilitation and potassium currents. PMID- 10717429 TI - Neuropeptide Y reduces epileptiform discharges and excitatory synaptic transmission in rat frontal cortex in vitro. AB - Neuropeptide Y reduced spontaneous and stimulation-evoked epileptiform discharges in rat frontal cortex slices perfused with a magnesium-free solution and with the GABA(A) receptor antagonist picrotoxin. To investigate the mechanism of that action, effects of neuropeptide Y on intrinsic membrane properties and synaptic responses of layer II/III cortical neurons were studied using intracellular recording. Neuropeptide Y (1 microM) had no detectable effect on the membrane properties of neurons. The evoked synaptic potentials were attenuated by neuropeptide Y. Moreover, the pharmacologically isolated excitatory postsynaptic potentials, mediated by N-methyl-D-aspartate and non-N-methyl-D-aspartate receptors, were reversibly depressed by neuropeptide Y. The most pronounced inhibitory effect of neuropeptide Y was observed on late polysynaptic excitatory postsynaptic potentials. To assess a putative postsynaptic action of neuropeptide Y, N-methyl-D-aspartate was locally applied in the presence of tetrodotoxin. The N-methyl-D-aspartate-evoked depolarizations were unaffected by neuropeptide Y, which suggests that the depression of excitatory postsynaptic potentials was due to an action at sites presynaptic to the recorded neurons. These data show that neuropeptide Y attenuates epileptiform discharges and the glutamate receptor mediated synaptic transmission in the rat frontal cortex. The above results indicate that neuropeptide Y may regulate neuronal excitability within the cortex, and that neuropeptide Y receptors are potential targets for an anticonvulsant therapy. PMID- 10717430 TI - Ultrastructural evidence of fibrillar beta-amyloid associated with neuronal membranes in behaviorally characterized aged dog brains. AB - The aged dog brain accumulates beta-amyloid in the form of diffuse senile plaques, which provides a potentially useful in vivo model system for studying the events surrounding the deposition of beta-amyloid. We used postembedding immunocytochemistry at the electron microscopic level to determine the subcellular distribution of beta-amyloid 1-40 and beta-amyloid 1-42 peptides in the prefrontal and parietal cortex of behaviorally characterized dogs ranging in age from one to 17 years. Immunogold particles signaling beta-amyloid 1-42 occurred over intracellular and extracellular fibrils that were approximately 8 nm in width. Intracellular beta-amyloid 1-42 fibrils were found in close proximity to glial fibrillary acidic protein fibers within astrocytes, but only in cells with signs of plasma membrane disruption. Neuronal labeling of beta amyloid 1-42 appears to be associated with the plasma membrane. Membrane-bound beta-amyloid 1-42 occurs in the form of fine fibrils that are embedded in the dendritic membrane and appear to project into the extracellular space as determined by quantitative analysis of the immunogold particle distribution. Bundles of beta-amyloid 1-42 were also closely associated and/or integrated with degenerating myelin sheaths of axons. In one dog that was impaired on several cognitive tasks, extensive beta-amyloid 1-42 deposition was associated with microvacuolar changes and vascular pathology. The present findings suggest that beta-amyloid 1-42 may be generated at the dendritic plasma membrane as well as in intracellular compartments. The close association between beta-amyloid 1-42 and destroyed myelin suggests one possible new mechanism by which beta-amyloid 1-42 induces neurodegeneration. PMID- 10717431 TI - Developmental change of GABA(A) receptor-mediated current in rat hippocampus. AB - Developmental change of GABA(A)ergic inhibitory postsynaptic current in rat hippocampal CA3 region was examined using patch-clamp recording method. Spontaneous and evoked inhibitory postsynaptic currents were recorded from acute hippocampal slices of neonates (postnatal days 2-4) and adults (days 18-38). Decay kinetics of the spontaneous inhibitory postsynaptic current was slower in neonates than in adults. Application of 500 nM zolpidem increased decay time constants of the inhibitory postsynaptic currents in both groups with a stronger effect on adults. Zinc (50 microM) inhibited the neonatal inhibitory postsynaptic currents but the inhibition was weaker in adults. Modification of the GABA(A)ergic inhibitory postsynaptic currents by furosemide (0.6 mM) or diazepam (100 nM) did not cause marked differences between the neonate and adult groups. These results demonstrate that GABA(A)ergic inhibitory postsynaptic currents in hippocampal CA3 pyramidal cells change developmentally and indicate that different receptor isoforms are functionally expressed between neonates and adults. PMID- 10717432 TI - Morphology of spiny neurons in the human entorhinal cortex: intracellular filling with lucifer yellow. AB - The present study was designed to investigate the morphology of spiny neurons in the human entorhinal cortex. Coronal entorhinal slices (n = 67; 200 microm thick) were obtained from autopsies of three subjects. Spiny neurons (n = 132) filled with Lucifer Yellow were analysed in different subfields and layers of the entorhinal cortex. Based on the shape of the somata and primary dendritic trees, spiny neurons were divided into four morphological categories; (i) classical pyramidal, (ii) stellate, (iii) modified stellate, and (iv) horizontal tripolar cells. The morphology of filled neurons varied more in different layers than in the different subfields of the entorhinal cortex. In layer II, the majority (81%) of spiny neurons had stellate or modified stellate morphology, but in the rostromedial subfields (olfactory subfield and rostral subfield) there were also horizontal tripolar neurons. Dendritic branches of layer II neurons extended to layer I (94%) and to layer III (83%). Unlike in layer II, most (74%) of the filled neurons in layers III, V and VI were classical pyramidal cells. The majority of pyramidal cells in the superficial portion of layer III had dendrites that extended up to layer II, occupying the space between the neuronal clusters. Some dendrites reached down to the deep portion of layer III. Apical dendrites of layer V and VI pyramidal cells traveled up to the deep portion of layer III.Our data indicate that the morphology of spiny neurons in different layers of the human entorhinal cortex is variable. Vertical extension of dendritic branches to adjacent layers supports the idea that inputs terminating in a specific lamina influence target cells located in various entorhinal layers. There appears to be more overlap in the dendritic fields between superficial layers II and III than between the superficial (II/III) and deep (V/VI) layers, thus supporting the idea of segregation of information flow targeted to the superficial or deep layers in the human entorhinal cortex. PMID- 10717433 TI - Hippocampal lesions enhance startle gating-disruptive effects of apomorphine in rats: a parametric assessment. AB - Prepulse inhibition of the startle reflex is an operational measure of sensorimotor gating that is impaired in schizophrenia patients and dopamine agonist-treated rats. Previous reports demonstrated an enhanced sensitivity to the prepulse inhibition-disruptive effects of the D(1)/D(2) agonist apomorphine in adult rats four weeks after cytotoxic lesions of the hippocampus, but left unanswered several important questions regarding the nature of this apparent lesion-induced dopamine supersensitivity. Because of the potential importance of this model to current theories of the pathophysiology of schizophrenia, studies now assessed specific features of this effect of hippocampus lesions on prepulse inhibition in rats. The enhanced prepulse inhibition-disruptive effects of apomorphine in ventral hippocampus-lesioned rats were unaffected by startle pulse intensity, suggesting an independence of this lesion effect from potential ceiling effects of elevated startle magnitude. These lesion effects were observed four weeks post-lesion, but not two weeks post-lesion, suggesting a delayed development of this phenomenon. No enhancement of apomorphine sensitivity was observed in rats four weeks after lesions restricted to the dorsal hippocampus; in contrast, these lesions significantly increased "no-drug" levels of prepulse inhibition. Ventral hippocampus-lesioned rats exhibited a significant reduction in prepulse inhibition after subthreshold doses of either the selective D(2) family agonist quinpirole or the partial D(1) agonist SKF 38393, suggesting that activation of either receptor family is adequate for the expression of this effect of ventral hippocampus lesions. This may be an important paradigm for understanding the contribution of ventral hippocampus dysfunction to the neurobiology of impaired sensorimotor gating in neuropsychiatric populations. PMID- 10717434 TI - Depletion of glutamate GluR2 receptor-containing neurons in the rat neostriatum by specific immunotoxin. AB - In the present study, a novel GluR2 receptor-specific immunotoxin was produced. The immunotoxin was produced by conjugation of molecules of trichosanthin, a ribosome inactivating protein, with goat anti-mouse immunoglobulin molecules. The secondary antibody was then combined with a commercially available GluR2 specific primary antibody to form an immunotoxin. The immunotoxins were unilaterally injected either into the neostriatum or into the lateral ventricle of rats. After one week, ipsilateral turning movements were observed after apomorphine treatments in those animals injected by the striatal route. In perfuse-fixed sections of the neostriatum, immunoreactivity for GluR2 was found to decrease in the striatal-lesioned animals. Most of the GluR2-immunoreactive perikarya in the neostriatum, the presumed medium spiny neurons, were depleted. In addition, immunoreactivity for GluR2/3, GluR5/6/7 and NMDAR1 was found to decrease to a different extent in the lesioned neostriatum. The number of GluR1-immunoreactive perikarya in the neostriatum, a group of striatal interneurons, was not affected by the GluR2 lesion. Ventricular administration of the GluR2 immunotoxin however, was found to be less potent. These results demonstrate for the first time that an indirect immunotoxin is useful for immunolesioning. A difference in potency was also observed in different routes of administration. The depletion of GluR2 containing medium spiny neurons in the neostriatum may upset the balance of the output systems of the basal ganglia and has a profound effect in movement control of the animals. PMID- 10717435 TI - Single-unit analysis of the pallidum, thalamus and subthalamic nucleus in parkinsonian patients. AB - Microelectrode-guided stereotactic operations performed in 29 parkinsonian patients allowed the recording of 86 cells located in the globus pallidus and 563 in thalamic nuclei. In the globus pallidus, the average firing rate was significantly higher in the internal (91+/-52 Hz) than in the external (60+/-21 Hz) subdivision. This difference was further accentuated when the average firing rate in the external subdivision was compared with that of the internal part of the internal subdivision (114+/-30 Hz). A rhythmic modulation in globus pallidus activities was observed in 19.7% of the cells, and this only during rest tremor episodes. In these cases, modulation frequency of unit activities was not statistically different from the rest tremor frequency (average: 4.6+/-0.5 vs 4. 4+/-0.4 Hz, respectively). In the medial thalamus, four types of unit activities could be defined. A sporadic type was mainly found in the parvocellular division of the mediodorsal nucleus (96.8% of the cells recorded) and in the centre median parafascicular complex (74.2%). Two other types of activities characterized by random or rhythmic bursts fulfilling the extracellular criteria of low-threshold calcium spike bursts were concentrated in the central lateral nucleus (62.3%) and the paralamellar division of the mediodorsal nucleus (34.1%). These activities could be recorded independently of the presence of a rest tremor. When a tremor episode occurred, the rhythmic low-threshold calcium spike bursts had an interburst frequency similar to rest tremor frequency, although they were not synchronized with it. The fourth type, the so-called tremor locked, was also characterized by rhythmic bursts which, however, did not display low-threshold calcium spike burst properties. These bursts occurred only when a rest tremor was present and was in-phase with the electromyographic bursts. All tremor-locked cells were located in the centre median-parafascicular complex. In the lateral thalamus, cells exhibiting random or rhythmic low-threshold calcium spike bursts were found preponderantly in the ventral anterior nucleus (53.4%) and in the ventral lateral anterior nucleus (52.7%). Tremor-locked units were confined to the ventral division of the ventral lateral posterior nucleus (35.4%). None of the random or rhythmic low-threshold calcium spike bursting units responded to somatosensory stimuli or voluntary movements, either in the medial or in the lateral thalamus. The presence of low-threshold calcium spike bursts at the thalamic level, together with the paucity (8%) of responses to voluntary movements compared to what is found in normal non-human primates, demonstrate a pathological state of inhibition due to the overactivity of the internal subdivision of the globus pallidus units. Activities of the thalamic cells producing low-threshold calcium spike bursts are not synchronized with each other or with the tremor. However, this does not exclude a causal role of these activities in the generation of tremor. Indeed, it has been demonstrated that even random electrical stimulations of the rolandic cortex in parkinsonian patients induce tremor episodes, probably due to the triggering of rhythmic, low threshold calcium spike-dependent, thalamocortical activities. Similarly, low threshold calcium spike bursts could be at the origin of rigidity and dystonia through an activation of the supplementary motor area and of akinesia when reaching the pre-supplementary motor area. We conclude that the intrinsic oscillatory properties of individual neurons, combined with the dynamic properties of the thalamocortical circuitry, are responsible for the three cardinal parkinsonian symptoms. PMID- 10717436 TI - Release of monoamines and nitric oxide is involved in the modulation of hyperpolarization-activated inward current during acute thalamic hypoxia. AB - Using slices of the dorsal lateral geniculate nucleus, it has been shown that, in the presence of excitatory and inhibitory amino acid antagonists, brief periods of hypoxia (3-4 min of 95% N(2)/5% CO(2)) induce in thalamocortical neurons an increase in instantaneous input conductance (G(N)) accompanied by an inward shift in baseline holding current (I(BH)). These effects have been suggested to be mediated, at least in part, by a positive shift in the voltage-dependence of the hyperpolarization-activated, mixed Na(+)/K(+) current (I(h)) and a change in its activation kinetics which transforms it into an almost instantaneously activated current. In this study, using the whole-cell patch-clamp technique, the contribution of an increased Ca(2+)-dependent transmitter release to the hypoxic response of thalamocortical neurons was further investigated using (i) blockers of calcineurin, a Ca(2+)/calmodulin-activated phosphatase that selectively regulates Ca(2+)-dependent release, and (ii) antagonists of neurotransmitters that are known to modulate I(h). Thalamocortical neurons (n = 23) recorded with electrodes filled with calcineurin autoinhibitory fragment (30-250 microM), a membrane impermeable blocker of calcinuerin, showed no difference either in resting, or in the hypoxia-induced changes in, G(N), I(BH) and I(h), when compared to thalamocortical cells patched with electrodes that did not contain calcineurin autoinhibitory fragment. In contrast, in 18 of these neurons recorded with calcineurin autoinhibitory fragment-filled electrodes, bath application either of cyclosporin-A (20 microM) or tacrolimus (50-100 microM), two membrane permeable blockers of calcineurin, abolished the effects of hypoxia on G(N), I(BH), and I(h). Separate application of noradrenaline, serotonin, histamine and nitric oxide antagonists produced only a small depression of the hypoxic response, while concomitant bath application of these antagonists decreased the hypoxia-induced changes in G(N) and I(BH) by 55 and 42%, respectively (n = 12). Concomitant bath application of 8-bromo-adenosine-3'5'-cyclicmonophosphate and 8 bromo-guanosine-3'5'-cyclicmonophosphate (both 1mM), which are known to mediate the action of these transmitters on I(h), increased G(N) (40%), decreased I(h) time-constant of activation (30%) and significantly occluded (50%) the hypoxia induced effect on G(N) and I(BH). Thalamocortical neurons (n = 6) patched with electrodes filled with 8-bromo-adenosine-3'5'-cyclicmonophosphate and 8-bromo guanosine-3'5'-cyclicmonophosphate (both 1 mM) showed a larger G(N) than the one recorded with the standard internal solution, and a significant depression of the hypoxia-induced changes in G(N) and I(BH). These results indicate that during acute thalamic hypoxia an increased release of noradrenaline, serotonin, histamine and nitric oxide is responsible for transforming I(h) into an instantaneously activating current via cyclic AMP- and cyclic GMP-mediated mechanisms. PMID- 10717437 TI - Atm-deficient mice Purkinje cells show age-dependent defects in calcium spike bursts and calcium currents. AB - Ataxia telangiectasia in humans results from homozygous loss-of-function mutations in ATM. Neurological deterioration is the major cause of death in ataxia telangiectasia patients: in the cerebellum, mainly Purkinje cells are affected. We have generated Atm-deficient mice which display neurological abnormalities by several tests of motor function consistent with an abnormality of cerebellar function, but without histological evidence of neuronal degeneration. Here we performed a more detailed morphological analysis and an electrophysiological study on Purkinje cells from Atm-deficient mice of different ages. We found no histological or immunohistochemical abnormalities. Electrophysiology revealed no abnormalities in resting membrane potential, input resistance or anomalous rectification. In contrast, there was a significant decrease in the duration of calcium and sodium firing. The calcium deficit became significant between six to eight and 12-20 weeks of age, and appeared to be progressive. By voltage-clamp recording, we found that the firing deficits were due to a significant decrease in calcium currents, while inactivating potassium currents seem unaffected. In other mutant mice, calcium current deficits have been shown to be related to cell death.Our experiments suggest that the electrophysiological defects displayed by Atm-deficient mice are early predegenerative lesions and may be a precursor of Purkinje cell degeneration displayed by ataxia telangiectasia patients. PMID- 10717438 TI - Perfusion of the hypothalamic paraventricular nucleus with N-methyl-D-aspartate produces thromboxane A2 and centrally activates adrenomedullary outflow in rats. AB - We applied a microdialysis technique for the measurement of hypothalamic thromboxane B2, a stable metabolite of thromboxane A2, in urethane-anesthetized rats. Perfusion with N-methyl-D-aspartate (1.5 and 2.5mM) of the paraventricular nucleus by microdialysis probe concentration-dependently elevated the levels of thromboxane B2 in this region and plasma levels of catecholamines. The elevation of adrenaline was much more marked than that of noradrenaline. Pretreatment with dizocilpine maleate (0.1 mM), a non-competitive antagonist of N-methyl-D aspartate receptors, of the paraventricular nucleus by microdialysis probe attenuated the N-methyl-D-aspartate (1.5 mM)-induced elevations of both thromboxane B2 and plasma catecholamines. Intracerebroventricular administration of furegrelate (250 microg/animal), a thromboxane A2 synthase inhibitor, also abolished the responses evoked by N-methyl-D-aspartate. These results indicate that N-methyl-D-aspartate applied into the paraventricular nucleus produces thromboxane A2 in this region and elevates plasma levels of catecholamines, especially adrenaline. Thromboxane A2 produced in this hypothalamic nucleus is probably involved in the N-methyl-D-aspartate-induced central adrenomedullary outflow. PMID- 10717439 TI - GABAergic modulation of gap junction communication in slice cultures of the rat suprachiasmatic nucleus. AB - We employed morphological and electrophysiological methods in order to elucidate mechanisms which are responsible for communication between cellular oscillators in the cultured rat suprachiasmatic nucleus, the site of the endogenous biological clock that regulates circadian rhythms in mammals. When a gap junction permeable dye, Lucifer Yellow, was injected into single neurons in the suprachiasmatic nucleus culture, the dye was transferred to neighboring cells in a gap junction blocker-sensitive manner. Optical imaging of neural activity evoked by electrical stimulation in the culture revealed that the spread of depolarization was inhibited by gap junction blockers but not by a blocker of voltage-dependent Na(+) channels. Depolarization propagation was inhibited by muscimol, a GABA(A) receptor agonist, in a dose-dependent manner and the inhibition was reversed by bicuculline, a GABA(A) receptor antagonist. Furthermore, muscimol inhibited dye-transfer between neurons in the suprachiasmatic nucleus culture in a dose-dependent fashion.These independent lines of evidence suggest that the gap junction communication is involved in interneuronal communication in the suprachiasmatic nucleus slice culture and that the coupling state between neurons is not static but dynamically regulated via GABA(A) receptor systems. PMID- 10717440 TI - Short-circuited neuron: a note. AB - Here, we demonstrate in a direct electrophysiological experiment that a neuron can form electrical connections to itself. An isolated identified neuron with a long axon was plated in culture and the axon was looped so that its distal end contacted the cell body. After two days in culture, the cell body and the axon were both impaled with microelectrodes and the axon segment between the recording electrodes was cut. Electrotonic coupling was revealed between the separated cell compartments immediately after axon transection. In contrast to an earlier publication [Guthrie P. B. et al. (1994) J. Neurosci. 14, 1477-1485], no constraints on the formation of the electrical connections between different parts of the same neuron were revealed in our experiments.Thus, these experiments demonstrate that in vitro culture of a single neuron can form reflexive electrical connections which may strongly affect the basic properties of the neuron and should be taken into account in both experimental and model electrophysiological studies. PMID- 10717441 TI - Interactions between brainstem and trigeminal neurons detected by cross-spectral analysis. AB - Cells in the nucleus raphe magnus that are inhibited by noxious skin stimuli (off cells) have been postulated to suppress pain by continuously inhibiting spinal and trigeminal nociceptive neurons. To test this hypothesis, spontaneous activity was simultaneously recorded from off-cells (n = 15) and wide-dynamic range cells (n = 27) of the trigeminal complex (subnucleus interpolaris), in rats anesthetized with pentobarbital. Most off-cells (n = 14) had rhythmic interspike intervals, their modes averaging 106 ms. No trigeminal cell fired rhythmically. Rhythmic firing was defined quantitatively: the autospectrum's peak power had to exceed 1.75 times its asymptote. This formula was obtained by generalizing from a natural cut-off in the theoretical autospectrum for serially uncorrelated, gamma distributed intervals, whose firing can be varied from Poissonian to highly regular by adjusting one parameter. It encompasses the qualitative judgement of autocorrelograms commonly made in neurophysiology. Cross-correlograms (n = 29) appeared noisy and otherwise featureless. However, their power spectra (cross periodograms) sometimes showed significant peaks, compared with simulated non interactive distributions. The latter were generated by interchanging the raphe interval sequences at one random point (as in cutting a deck of cards), thus retaining most of their serial correlation. Of 29 cross-periodograms, 21 were significant at 1 to 13 frequencies (100 points, 0.4 to 39 Hz). These frequencies were often near the peak raphe power, and sometimes near its harmonics too. Furthermore, cross-spectral phase angles at peak power were non-uniform, most falling between 0 and 180 degrees (unit vector sum 60 degrees, n = 20). To understand why the frequency domain gave better detection, cross-spectra and cross-correlations were modeled theoretically by convolving idealized input autocorrelations and synaptic response functions. This demonstrated that rhythmic firing is insufficient for better frequency-domain detection, and that serially correlated input intervals or non-additive synaptic responses are necessary. The conclusion was confirmed by stochastic simulation of a simple non-additive synapse, that required successful input spikes to fall within a specified interval of the preceding spike. Experimentally, serial correlation was found in 12 of the 15 raphe cells, and in 20 of the 27 trigeminal cells. It is proposed that the weak experimental cross-correlograms arise because many asynchronous raphe inputs converge on each trigeminal cell, possibly to optimize the resting suppression of pain. The distribution of cross-spectral phase angles at peak raphe power suggested that raphe spikes arriving at the synapses' preferred interval cause a fall in trigeminal activity. In general, cross-spectral analysis can sometimes uncover influences hidden in cross-correlograms, but the firing of one neuron must be rhythmic and non-renewal, or else certain input intervals must be favored in synaptic transmission. PMID- 10717442 TI - After axotomy, substance P and vasoactive intestinal peptide expression occurs in pilomotor neurons in the rat superior cervical ganglion. AB - Autonomic sympathetic postganglionic neurons normally express distinct combinations of neuropeptides which are often highly correlated with the projection of the neurons. When sympathetic postganglionic neurons are axotomized, they can express quite different neuropeptides, notably substance P, vasoactive intestinal peptide or galanin. In this study, we have examined rat sympathetic postganglionic neurons in the superior cervical ganglion that project to the skin, the vasculature of the skeletal muscle or to the submandibular salivary gland, and assessed whether the neuropeptides that they express after axotomy depend on which target tissue they previously innervated. In all three populations, around half of the postganglionic neurons expressed galanin after axotomy. In contrast, only skin-projecting neurons showed a significant increase in the number of neurons that expressed substance P (22%) and vasoactive intestinal peptide (17%) following axotomy. Within the skin-projecting neurons, as judged on the basis of cell body size, substance P and vasoactive intestinal peptide were expressed predominantly in pilomotor neurons, but only rarely were the two neuropeptides present in the same nerve cell body. In conclusion, we have demonstrated that three different neuropeptides, which can be induced by axotomy in postganglionic neurons, follow quite different patterns of expression when they are viewed in relation to the function of the postganglionic neurons in the superior cervical ganglion. PMID- 10717443 TI - Differential messenger RNA distribution of lactate dehydrogenase LDH-1 and LDH-5 isoforms in the rat brain. AB - The role of lactate in brain energy metabolism has recently received renewed attention. Although blood-borne monocarboxylates such as lactate poorly cross the blood-brain barrier in the adult brain, lactate produced within the brain parenchyma may be a suitable substrate for brain cells. Lactate dehydrogenase is crucial for both the production and utilization of lactate. In this article, we report the regional distribution of the messenger RNAs for lactate dehydrogenase isoforms 1 and 5 in the adult rat brain using in situ hybridization histochemistry with specific [alpha-(35)S]dATP 3' end-labeled oligoprobes. The autoradiographs revealed that the lactate dehydrogenase-1 messenger RNA is highly expressed in a variety of brain structures, including the main olfactory bulb, the piriform cortex, several thalamic and hypothalamic nuclei, the pontine nuclei, the ventral cochlear nucleus, the trigeminal nerve and the solitary tractus nucleus. In addition, the granular and Purkinje cell layers of the cerebellum showed a strong labeling. The neocortex (e.g., cingular, retrosplenial and frontoparietal cortices) often exhibits a marked laminar pattern of distribution of lactate dehydrogenase-1 messenger RNA (layers II/III, IV and VI being most strongly labeled). In contrast, expression of the lactate dehydrogenase-5 messenger RNA generally seemed more diffusely distributed across the different brain regions. Expression was particularly strong in the hippocampal formation (especially in Ammon's horn and dentate gyrus) and in the cerebral cortex, where no laminar pattern of distribution was observed. Overall, these data are consistent with the emerging idea that lactate is an important energy substrate produced and consumed by brain cells. PMID- 10717444 TI - Ontogeny of Rxt1, a vesicular "orphan" Na(+)/Cl(-)-dependent transporter, in the rat. AB - The developmental expression of the orphan Na(+)/Cl(-)-dependent transporter, Rxt1, was studied in the rat using a specific [(35)S]complementary RNA probe and affinity purified antibodies. Western blotting experiments allowed the detection of Rxt1 in brain as early as on embryonic day 16. After birth, the brain levels of Rxt1 increased dramatically up to a maximum around postnatal day 30 and then decreased slightly to the adult value. In situ hybridization experiments allowed the earliest detection of Rxt1 messenger RNA in the brain and spinal cord at embryonic day 14. In embryonic day 18 embryos, Rxt1 messenger RNA was present not only in the nervous system but also in the pituitary, the thymus and the heart. Immunoautoradiograms of whole embryo at embryonic days 16 and 18 showed high amounts of the Rxt1 protein in the spinal cord and brain. Moreover, at embryonic day 18, the orphan transporter was expressed in the thymus, heart and liver. At these ages, Rxt1 immunolabeling was localized in neurons of the subplate and in the ventricular zone of the brain. During early postnatal stages, Rxt1 messenger RNA expression demonstrated dynamic and complex changes until postnatal day 13. In particular, this transcript was relatively abundant in the striatum at postnatal days 3 and 5 and then decreased to very low levels after postnatal day 10. At the same period, Rxt1 immunostaining in the hippocampus and the cerebral cortex was observed all over the gray matter, in cell bodies as well as in the neuropil. Finally, the adult pattern was reached around postnatal day 13 for Rxt1 messenger RNA, but only at postnatal day 20 for the Rxt1 protein. The presence of Rxt1 messenger RNA and protein at embryonic stages and the high expression of the protein during synaptogenesis suggest that this vesicular "orphan" transporter is involved in the brain maturation process. PMID- 10717445 TI - Ultrastructural features of medullary chromaffin cell cultures. AB - The ultrastructural organization on the fourth day of culture of chromaffin cells isolated from the bovine adrenal medulla was characterized based on electron microscopic and morphological analysis. We established that medullary chromaffin cells could be divided into four morphologically different subtypes. Most cells (49.1% of those examined) had a dense cytoplasm and fine dense granules. Cells with dense cytoplasm and large granules represented a second type of chromaffin cell (21.1%). Cells of the third type had a light cytoplasm, granules with a light halo and a well-developed Golgi apparatus (26.3%). The fourth type of chromaffin cell was characterized by moderately dense cytoplasm with well expressed varicose rough endoplasmic reticulum (about 3.5%). Among concomitant cell types, cortical adrenal cells from the zona fasciculata and zona glomerulosa, epithelial cells, fibroblasts, lymphocytes, brown lipoblasts and glial Schwann cells were present. Morphological analysis implies that cells with dense cytoplasm and fine granules and those with light cytoplasm and haloed granules (75.4% in total) are adrenaline-containing cells, whereas the cells with dense cytoplasm and large granules (26.3%) contain noradrenaline. Cells with moderately dense cytoplasm and varicose reticulum share common morphological properties with classical glandular cells and, by their properties, were closer to noradrenaline-containing cells. It is concluded that chromaffin cells, which are the main cell type among cultured cells from adult bovine adrenal medulla, are morphologically quite heterogeneous. Other cell types of different nature may also be present in the culture and can locally influence the properties of the investigated medullary chromaffin cells used in electrophysiological experiments. PMID- 10717446 TI - Standards for the Ph.D. Degree in the Molecular Biosciences. PMID- 10717447 TI - Calculating apparent equilibrium constants of enzyme-catalyzed reactions at pH 7. AB - Apparent equilibrium constants K' of biochemical reactions at pH 7 and standard apparent reduction potentials of half reactions at pH 7 can be calculated using a table of standard transformed Gibbs energies of formation Delta(f)G'(0) at pH 7. A table is provided for 136 reactants at 25 degrees C, pH 7, and ionic strengths of 0, 0.10, and 0.25 M. Examples are given to illustrate the use of the table. PMID- 10717448 TI - Optimization of energy coupling: what is all the argument about? AB - Although it may be asked how effective glycolysis is in retaining the chemical energy in the bonds of glucose during its breakdown in the formation of ATP, the reasons for the coupled pathway of glycolysis having evolved as it has are probably as much as a consequence of the need to find reactions that can lead to formation of phosphoryl groups able to transfer to ADP as to the overall thermodynamics of the pathway. It is not meaningful to talk of optimization of energy coupling solely in terms of free energy changes. PMID- 10717449 TI - The making of a model of Escherichia coli, magnified two million times - the 'Millennium Bug' AB - The author is making for the Millennium a model of Escherichia coli at 2x10(6) with emphasis on its interior composition. At this magnification molecules come into view and metabolic pathways are demonstrable as tangible systems of metabolites and enzymes interacting with one another. The model is intended to have value both for public understanding of science and in teaching of microbiology and biochemistry. PMID- 10717450 TI - Understanding ion transport by quantification of mitochondrial swelling. AB - Suspensions of mitochondria are turbid and scatter light. An increase in the matrix volume (swelling) due to the influx of permeable solutes results in a decrease in the amount of light scattered. This property can be used to study solute fluxes across the mitochondrial inner membrane. A rapid method for isolating mitochondria is presented along with three swelling experiments using energized and non-energized mitochondria to illustrate ion transport across energy transducing membranes. PMID- 10717451 TI - Classifying amino acids as gluco(glyco)genic, ketogenic, or both. AB - Eleven popular and recent biochemistry textbooks were examined and compared concerning the classification of amino acids as being gluco(glyco)genic, ketogenic, or both. The main differences concerned mostly two amino acids, threonine, which in some cases is considered only gluco(glyco)genic and in others both gluco(glyco)-ketogenic, and lysine, which is considered chiefly ketogenic, but in two cases is reported as being gluco(glyco)-ketogenic. Some more mistakes, discrepancies and curiosities among different textbooks are also reported. Biochemistry textbook authors and teachers should be aware of this situation to avoid any confusion, especially among students. PMID- 10717453 TI - Biotechnology education. PMID- 10717452 TI - Problem-based learning in veterinary medicine: protein metabolism. AB - A problem-based approach to teaching biochemistry to veterinary students is described which aims to encourage the finding and learning of biochemical information that is relevant to real-life veterinary problems. PMID- 10717454 TI - Development of biotechnology education in Turkey. AB - For sometime Turkish scientists have been actively involved in biotechnology related research. However, biotechnology educa-tion in Turkey is a relatively recent phenomenon. The subject has not been addressed at the undergraduate level in a serious way until recently. This is evident from the lack of undergraduate degree programmes in biotechnology at Turkish Universities. The Turkish scientific establishment is very much aware of the importance of biotechnology and has identified this subject as one of the priority areas. The Universities are taking positive steps towards enhancing Biotechnology education. This article focuses on the emergence, as well as the problems and prospects of Biotechnology education in Turkey. PMID- 10717455 TI - Plant protoplast isolation - a practical approach. AB - A method for the isolation of plant protoplasts is presented that uses inexpensive sources of enzymes. It is suitable for student use in Biotechnology. PMID- 10717456 TI - Chemical cross-linking analysis of Ca(2+-)ATPase from rabbit skeletal muscle. AB - A student laboratory experiment is described dealing with chemical cross-linking analysis of Ca(2+-)ATPase from rabbit muscle. The experiment introduces students to a wide range of techniques including subcellular fractionation, chemical cross linking, polyacrylamide gel electrophoresis, blotting and immunochemical detection of proteins. PMID- 10717457 TI - A simple and rapid method for screening amylolytic bacteria. AB - A laboratory class was designed for the study of the ecology of amylolytic bacteria in soil, although other sources may be equally suitable for this purpose. Groups of three students carried out the following: (a) preparation and sterilization of medium and plates, (b) collection and preparation of soil samples, spreading the samples on the plates, (c) incubation of the plates at 37 degrees C overnight, a further 1 h incubation at 60 degrees C to observe amylolytic activity due to thermophilic bacteria, and (d) interpretation and discussion of the results. These tasks are accomplished in two periods of 4h on consecutive days. No sophisticated instruments are required for these experiments, which can be carried out in three classes of 4h each. On the first day the students prepare culture media, buffers and reagents, as well as collect and grow soil samples. The second day is spent for both taxonomic identification of colonies and the HAI determination. PMID- 10717458 TI - Investigating plant lipid biopolymers. AB - This paper presents a review of the biological importance and occurrence of plant cutin, the main lipid biopolymer in higher plants. Its chemical composition and structure are reviewed and a short laboratory practical exercise on the extraction, purification and qualitative analysis of fruit cutin is presented. PMID- 10717459 TI - Does succinate oxidation yield FADH(2) or ubiquinol? AB - Most textbooks still show the oxidation of succinate in the tricarboxylic acid (TCA) cycle resulting in the reduction of FADH(2). Such a presentation does not reflect the reaction catalysed by the enzyme in vivo or in vitro, does not simplify the treatment of the reaction, and is unnecessarily misleading and confusing. PMID- 10717473 TI - Role of the transcription factor AML-1 in acute leukemia and hematopoietic differentiation. AB - Chromosomal translocations affecting the AML-1 gene are among the most frequent aberrations found in acute leukemia. Because the AML-1 transcription factor is a critical regulator of hematopoeitic cell development, normal homeostasis is disrupted in cells containing these translocations. In this review we describe the mechanisms of transcriptional activation and repression by AML-1 and how this transcriptional control is disrupted by the chromosomal translocations that affect AML-1. Finally, we discuss how the mechanism of transcriptional repression by these chromosomal translocation fusion proteins is a possible target of therapeutic intervention in acute leukemia. PMID- 10717474 TI - Analysis of the human LHX3 neuroendocrine transcription factor gene and mapping to the subtelomeric region of chromosome 9. AB - The Lhx3 LIM homeodomain transcription factor is critical to pituitary organogenesis and motor neuron development. We determined the genomic structure and chromosomal localization of human LHX3. The gene contains seven coding exons and six introns that span 8.7 kilobases in length. The LHX3 gene codes for two functionally distinct isoforms that differ in their amino termini but share common LIM domains and a homeodomain. The functional domains of the LHX3 proteins are encoded by distinct exons. The alternate amino termini and LIM domains lie within individual exons, and the homeodomain is coded by two exons interrupted by a small intron. Human LHX3 maps to the subtelomeric region of chromosome 9 at band 9q34.3, within a region noted for chromosomal translocation and insertion events. Characterization of the genomic organization and chromosomal localization of LHX3 will enable molecular evaluation and genetic diagnoses of pituitary diseases and central nervous system developmental disorders in humans. PMID- 10717475 TI - Eel WT1 sequence and expression in spontaneous nephroblastomas in Japanese eel. AB - Nephroblastomas spontaneously developing in Japanese eel reared at farms for 5 to 9months after collection from the wild [Masahito et al., Cancer Res., 52 (1992) 2575-2579] were investigated to cast light on the role of Wilms' tumor 1 gene (WT1) in eel kidney tumorigenesis. Cloning of the WT1 counterpart, EWT1, revealed that conservation of an alternative splice II site, located between the third and fourth zinc fingers, was conserved. The zinc finger domain was highly conserved. The transregulator region, sequences corresponding to exons 4 and 5 in WT1, were lacking in EWT1 cDNA. EWT1 was found to be expressed in kidney, testis and spleen and in situ hybridization revealed dark-stained immature cells in elver kidney to be positive. Although no EWT1 gene mutations were found in 38 eel nephroblastomas, 26 polymorphic nucleic acid changes were observed. Aberrant WT1 expression was noted in epithelial (12 out of 27; 44%) and nephroblastic cell histological types (three out of five; 60%) of eel nephroblastomas. On in situ hybridization the EWT1 expressive cells resembled human blastema cells, similar to those in human Wilms' tumor. These data demonstrated strong signals that the EWT1 protein may function in the development of eel kidney and play a role in genesis of nephroblastomas as in mammals. PMID- 10717476 TI - Expression and regulation of a gene encoding neural recognition molecule NB-3 of the contactin/F3 subgroup in mouse brain. AB - NB-3 is a neural recognition molecule which is a member of contactin/F3 subgroup in the immunoglobulin superfamily. We report here the developmental expression pattern and localization of NB-3 mRNA in mouse brain, determination of the NB-3 gene organization and identification of the promoter region. We also describe a splicing isoform of mouse NB-3. Mouse NB-3 exhibited 96% identity with rat NB-3 at the amino acid sequence level. The splicing isoform lacked the amino acid residues between 62 and 78 of the original NB-3, which constituted a part of the first immunoglobulin-like domain. The expression of NB-3 mRNA was evident after birth, reaching a maximum at the postnatal seventh day, and declined thereafter in the cerebrum, whereas the mRNA increased in the cerebellum to adulthood. In situ hybridization demonstrated that NB-3 mRNA was preferentially expressed in the accessory olfactory bulb, layers II/III and V of the cerebral cortex, piriform cortex, anterior thalamic nuclei, locus coeruleus of the pons and mesencephalic trigeminal nucleus, and in Purkinje cells of the cerebellum. The mouse NB-3 gene consisted of 23 exons spanning more than 130kb. The overall organization of the gene was similar to those of the F11, axonin-1 and TAX-1 genes of the subgroup. By reporter gene analysis with the 5'-flanking region of the gene, we found a basal promoter activity in the 1.2kb fragment upstream of the putative transcription initiation site. This study provides a basis for elucidating the biological significance of the contactin/F3 subgroup molecules. PMID- 10717477 TI - Molecular characterization of a mouse heterogeneous nuclear ribonucleoprotein D like protein JKTBP and its tissue-specific expression. AB - The human DNA- and RNA-binding protein JKTBP is a new member of heterogeneous nuclear ribonucleoproteins (hnRNPs) that are involved in mRNA biogenesis. We cloned and characterized a mouse homolog and studied its expression in mouse tissues. The cDNA encoded a 301-residue polypeptide. There is only a single amino acid difference between the mouse and human sequences. Northern blotting indicated ubiquitous but varied expressions of approximately 1.4 and 2.8kb mRNAs in various tissues. Immunoblotting indicated that the amounts of protein of about 38kDa were higher in the brain and testis than in other tissues. An additional protein of about 53kDa was found in the brain and testis. Germ cell-deficient W/W(v) mutant mice and aged mice had the reduced amounts of JKTBP in the testes. Immunohistochemical staining indicated cell type-specific expression of JKTBP in tissues: neurons and spermatocytes displayed strong signal intensities. The signals were confined to the nucleus. The amount of 38kDa JKTBP was estimated to be approximately 1.3x10(7) molecules per HL-60 cell. These results indicate that JKTBP is an abundant, highly conserved nuclear protein. PMID- 10717478 TI - The genomic structure of the scallop, Patinopecten yessoensis, troponin C gene: a hypothesis for the evolution of troponin C. AB - Two cDNAs encoding troponin C (TnC) isoforms are isolated from the scallop, Patinopecten yessoensis, striated adductor muscle. The sequential differences between these isoforms, named TnC(long) and TnC(short), are restricted in several residues of the C-terminal region. TnC(long) is commonly expressed in both the striated and the smooth adductor muscle; however, TnC(short) is only in the striated adductor muscle. The TnC gene is a single copy gene in the scallop, thus they are expressed through the alternative splicing from the same gene. The scallop TnC gene is constructed from five exons and four introns, and positions of introns are identical with chordate TnC genes, although the scallop TnC possesses no corresponding intron to the fourth intron of chordates. The loss of this intron is also observed in Drosophila TnC; these may be remnants of their ancestor, namely the early metazoan TnC gene might be a five exons-four introns structure. In addition, the absence of the corresponding intron is also observed among protostomian calmodulins (CaMs), a molecule closely related to TnC. This suggests that the common ancestor gene of the TnC superfamily might also be a five exons-four introns structure. Assuming this to be true, the discordance of the fourth intron positions observed among members of the family is well explained by the evolutionary independent gain of the intron on each member's lineage. PMID- 10717479 TI - Expression of the chloroplast-localized small heat shock protein by oxidative stress in rice. AB - A rice (Oryza sativa L. cv. Nakdong) cDNA clone, Oshsp26, encoding the chloroplast-localized small heat shock protein (smHSP) was isolated. Southern blot analysis of genomic DNA and the result of screening of a cDNA library indicated that the Oshsp26 gene is encoded by a single gene in the rice genome. The Oshsp26 gene was expressed following heat stress: the transcript level was highest when rice leaves were treated at high temperatures for 2h at 42 degrees C, and the transcripts became detectable after 20min and reached a maximum level after 2h. It was also found that the Oshsp26 gene was expressed following oxidative stress even in the absence of heat stress. Treatment of rice plants with methyl viologen (MV) in the light and treatment with hydrogen peroxide (H(2)O(2)), either in the light or in the dark, both caused a significant accumulation of the transcripts and the protein. Since MV treatment in the light leads to the generation of H(2)O(2) inside the chloroplast, it is likely that H(2)O(2) by itself acts to induce the expression of the Oshsp26 gene. These results suggest that the chloroplast smHSP plays an important role in protecting the chloroplast against damage caused by oxidative stress as well as by heat stress. PMID- 10717480 TI - A functional significance for codon third bases. AB - Most amino acids are specified by more than one trinucleotide codon. Here we show that amino acids of differing functional importance may be distinguished by the pattern of synonymous codon usage. GC-rich genes tend to be of a greater transcriptional (p<0.01) and mitogenic (p<0.0001) significance than AT-rich genes, consistent with GC-->AT mutational drift in methylated genomic regions. Third-base GC retention also identifies critical amino acids within individual proteins, as indicated by non-random patterns of codon variation between gene homologs and also by differential sequelae of site-directed mutagenesis. Sequence analysis of human receptor tyrosine kinase genes confirms that functionally important transmembrane hydrophobic amino acids are specified by codons containing GC third bases more often than are transmembrane neutral amino acids (chi(2)=134.2). Amino acids encoded by GC third bases thus appear more tightly linked to cell function and survival than are those encoded by AT third bases. PMID- 10717481 TI - Evolution of a glucose-regulated ADH gene in the genus Saccharomyces. AB - To determine when a glucose-repressed alcohol dehydrogenase isozyme and its regulatory gene, ADR1, arose during evolution, we surveyed species of the genus Saccharomyces for glucose-repressed ADH isozymes and for ADR1 homologues. Glucose repressed ADH isozymes were present in all species of Saccharomyces sensu strictu and also in Saccharomyces kluyveri, the most distant member of the Saccharomyces clade. We cloned and characterized ADH promoters from S. bayanus, S. douglasii, and S. kluyveri. The ADH promoters from S. bayanus and S. douglasii had conserved sequences, including upstream regulatory elements, and an extended polydA tract. The expression of a reporter gene driven by the S. bayanus promoter was glucose repressed and dependent on the major activator of transcription, ADR1, when it was introduced into S. cerevisiae. One S. kluyveri promoter was also glucose repressed and ADR1-dependent in S. cerevisiae. The other S. kluyveri ADH promoter was expressed constitutively and was ADR1-independent. Although showing little sequence conservation with the S. cerevisiae ADH2 promoter, the glucose-repressed S. kluyveri promoter contains numerous potential binding sites for Adr1. The glucose-repressed ADH from S. kluyveri is a mitochondrial isozyme most closely related to S. cerevisiae ADHIII. ADR1 homologues from S. douglasii and S. paradoxus contain a trinucleotide repeat encoding polyAsn that is lacking in S. cerevisiae and S. bayanus. No ADR1 homologue could be detected in S. kluyveri, suggesting that the potential for Adr1 regulation may have arisen first, before ADR1 evolved. PMID- 10717482 TI - Genomic organization of the human rod photoreceptor cGMP-gated cation channel beta-subunit gene. AB - We previously reported that the CNGB1 locus encoding the rod photoreceptor cGMP gated channel beta-subunit is complex, comprising non-overlapping transcription units that give rise to at least six transcripts (Ardell, M.D., Aragon, I., Oliveira, L., Porche, G.E., Burke, E., Pittler, S.J., 1996. The beta subunit of human rod photoreceptor cGMP-gated cation channel is generated from a complex transcription unit. FEBS Lett. 389, 213-218). To further understand the transcriptional regulation of this extraordinarily complex locus, and to develop a screen for defects in the gene in patients with hereditary disease, we determined its genomic organization and DNA sequence. The CNGB1 locus consists of 33 exons, which span approximately 100kb of genomic DNA on chromosome 16. The beta-subunit comprises two domains, an N-terminal glutamic acid-rich segment (GARP), and a C-terminal channel-like portion. Two additional exons encoding a short GARP transcript and a truncated channel-like transcript have been identified. A major transcription start point was identified 79bp upstream of the initiator ATG. To begin analysis of the basis for the generation of multiple transcripts, and to identify promoters driving expression in retina, approximately 2.5kb of the upstream region were sequenced. Putative cis-elements, which can bind the retina-specific transcription factors Crx and Erx, were found immediately upstream of the transcription start point, and may be important for gene expression in this tissue. From our analysis, a model is reported to account for at least four of the retinal transcripts. PMID- 10717483 TI - A novel Pax-6 binding site in rodent B1 repetitive elements: coevolution between developmental regulation and repeated elements? AB - Pax-6 encodes a transcription factor that is important in the development of eye and CNS. Identification of Pax-6 target genes is crucial for understanding the gene regulatory network in these developmental processes. Using an in-vitro approach of cyclic amplification of the protein binding sequences (CAPBS), we isolated a PAX6 binding sequence from a human single-copy (sc) DNA library. Characterization of this PAX6 binding sequence revealed a 15bp region (hGCalpha1BLs5) that is sufficient for PAX6 specific binding. From a homology search in the GenBank, we found that an hGCalpha1BLs5-like Pax-6 binding site exists in 21 genes (16 from rodent), 15 of which were shown to be able to bind Pax-6 in vitro. Interestingly, some of these sites occur in B1 repetitive elements. Although hGCalpha1BLs5 is highly similar to a region in B1 repetitive elements, PAX6 does not bind to the consensus sequence in B1. However, a single step mutation in some B1 elements can lead to a gain of function for PAX6 binding. This experimental evidence and phylogenetic analysis raise an interesting speculation for the coevolution between PAX6 regulation and repeat elements. Since a (Pax-6-binding) null B1 element can be re-activated by even a single-step mutation, it has the potential to recruit gene targets for Pax-6 if it is inserted into the regulatory region, and therefore may play a role for evolutionary modification of Pax-6 regulation. PMID- 10717484 TI - Identification of drosophila bicoid-interacting proteins using a custom two hybrid selection. AB - Bicoid directs pattern formation in the developing Drosophila embryo, and does so by performing two seemingly unrelated tasks; it activates transcription and represses translation. To understand how Bicoid carries out this dual role, we sought to identify Bicoid-ancillary proteins that might mediate Bicoid's function in transcription or translation. We used a customized version of the two-hybrid method and found two Bicoid-interacting proteins, Bin1 and Bin3, both of which interact with Bicoid in vitro. Bin1 is similar to a human protein (SAP18) involved in transcription regulation, and Bin3, described in this paper, is similar to a family of protein methyltransferases that modify RNA-binding proteins. Given that Bicoid's role as a translation regulator requires RNA binding, we suggest that the Bicoid-interacting methyltransferase might be important for that role. The custom two-hybrid method we used, in which Bicoid is bound to DNA via its own DNA binding domain, rather than via a fusion-protein tether, should be generally applicable to other DNA binding proteins. PMID- 10717485 TI - Oestrogens and wound healing. AB - During the past few decades several studies have documented the deleterious impact of the menopause on bone mass and cardiovascular disease, and the reduction of risk in this area by HRT. However, the possible effects of the postmenopausal deficiency in ovarian hormones on skin and its repair post-injury, are less well documented. This review provides a survey of the literature that is available regarding the involvement and influence of oestrogens on the various phases of cutaneous repair - inflammation, proliferation and remodelling. Research carried out on the effects of oestrogens, both in terms of deficiency and replacement, on the process of wound healing in various animal models is described and discussed, together with the very limited work undertaken in humans. This area of research is of paramount clinical importance both in terms of financial cost and human suffering, since many chronic wounds such as venous ulcers, pressure sores and burns afflict the elderly population, of whom postmenopausal women comprise the majority. Clinically our aim should be to restore the integrity and function of wounded tissue as rapidly as possible after injury and it is generally believed that a better understanding of the effects of oestrogens on wound healing could lead to improved care of cutaneous wounds, and the treatment of not only the wound but of the postmenopausal woman as a whole. PMID- 10717486 TI - Determinants of the intention to adopt hormone replacement therapy among premenopausal women. AB - OBJECTIVE: To identify the psychosocial factors that influence the intention to adopt hormone replacement therapy (HRT) at menopause. METHODS: Random Digit Dialing was used to recruit 644 premenopausal non-hysterectomized women aged 45 54. Data were collected using a telephone questionnaire previously developed according to the theory of planned behaviour. Variables measured were: intention to adopt HRT (INT); attitude towards HRT (Aact); perceived social norm (SN); perceived behavioural control (PBC); and personal normative belief (PNB). Socio demographic data were also obtained. RESULTS: Stepwise multiple regression of INT on the theoretical variables yielded an R(2) of 0.70. The determinants were Aact (beta=0.39, P<0.001), PNB (beta=0.25, P<0.001), PBC (beta=0.23, P<0.001) and SN (beta=0.12, P<0.001). Women with a strong intention to adopt HRT represented 25% of the sample. These women were more likely to believe that adopting HRT would have the following positive consequences: an improvement in general well-being, the prevention of health problems, an improvement in interpersonal relationships, an increase in productivity, the regulation of mood swings and a reduction of hot flashes. They were also more likely to believe in the following negative consequences: side-effects, an increased risk of cancer, the likelihood of weight gain, and interference in the natural course of menopause (all at P<0.001). CONCLUSION: Actions that target behaviourial beliefs regarding HRT and perceived barriers to its adoption are most likely to influence adoption of HRT. PMID- 10717487 TI - Few oligo-amenorrheic athletes have vasomotor symptoms. AB - OBJECTIVE: To assess whether women with athletic oligo-amenorrhea have vasomotor symptoms. MATERIAL AND METHODS: A mailed questionnaire was sent to 252 female athletes about vasomotor symptoms. Identical questions were also mailed to 1523 peri- and postmenopausal women. RESULTS: The prevalence of vasomotor symptoms was low in female athletes with oligo- and amenorrhea and similar to that found in athletes with regular menstruations. The prevalence was significantly lower than in menopausal women. Although more than a third of the menopausal women had hormone replacement therapy, 30% of them still had vasomotor symptoms at least every week compared with only 2% of the oligo-amenorrheic athletes. CONCLUSION: Vasomotor symptoms are very uncommon in oligo-amenorrheic athletes, although many of them are hypoestrogenic. It was suggested that one factor contributing to these symptoms around menopause is low hypothalamic activity of beta-endorphins, which makes the thermoregulatory centre labile. On the other hand, supraphysiological activity in hypothalamic beta-endorphins may cause the oligo amenorrhea in athletes, but may stabilise the thermoregulatory centre and thus prevent hot flushes. PMID- 10717488 TI - Radial scar of the breast: mammographic enigma in pre- and postmenopausal women. AB - OBJECTIVE: The aim of the study was to investigate the incidence and the mammographic features of the lesions suggestive of radial scar (RS). METHODS: We reviewed 31883 mammograms of women in pre and postmenopause and we found 23 (0.072%) images suggestive of RS. Twelve out of 23 (52%) women were in premenopause and 11 out of 23 (48%) in postmenopause, respectively. Histologic diagnosis was made on the surgical biopsy specimen. RESULTS: We described mammographic features of these lesions. On 23 biopsy specimens of mammograms suggestive of RS, histology pointed out 11 (48%) radial scars, 3 (13%) sclerosing adenosis and 9 (39%) carcinomas. CONCLUSIONS: In our case histories we found 11 (0.034%) radial scars among 31883 performed mammographies. Mammographic findings suggestive of RS provide remarkable diagnostic problems because numerous aspects at mammography suggestive of this lesion can be found also both in case of sclerosing adenosis and carcinomas making differential diagnosis impossible. The finding of mammographic features suggestive of RS imposes performance of targeted surgical biopsy for the correct diagnosis. PMID- 10717489 TI - Effect of menopause on low-density lipoprotein oxidation: is oestrogen an important determinant? AB - OBJECTIVES: Significantly increased risk for developing cardiovascular disease in post-menopausal women is linked with the fall of oestrogen. Although supraphysiological levels of oestrogen may inhibit oxygen free radical mediated low-density lipoprotein (LDL) oxidation, the effect of physiological level of oestrogen on LDL oxidation is unknown. METHODS: The present study compared oxidizability of LDL in healthy pre- and post-menopausal women by using a commonly employed copper ion-dependent method. RESULTS: Pre-menopausal women (n=20, mean age 27) had significantly higher serum oestradiol level (576+/-109 pmol/l) in comparison to post-menopausal women (n=23, mean age 51, oestradiol 64+/-18 pmol/l, P<0.001). The oxidation of LDL in two groups was not different by measuring either the lag phase of conjugated dienes formation (54+/-12 vs. 55+/ 14 min, P0.05) or the generation of thiobarbituric acid reactive substances over 4 h of oxidation. The major lipid soluble antioxidant in LDL, vitamin E (determined as alpha-tocopherol) is similar in two groups (2.34+/-0.48 vs. 2.40+/ 0. 56 nmol/mg LDL, pre- and post-menopausal subjects, respectively, P0. 05). Linear regression analysis found a weak but significant correlation between LDL vitamin E level and oxidizability of LDL in both groups but did not show effect of serum oestradiol levels. CONCLUSION: The results suggest that physiological levels of oestrogen may not be able to affect in vitro LDL oxidation. PMID- 10717490 TI - The effects of hormone replacement therapy on plasma lipids in type II diabetes. AB - OBJECTIVES: The effects of hormone replacement therapy on cardiovascular risk factors in postmenopausal women with non-insulin dependent diabetes mellitus (type II diabetes) is uncertain. METHODS: The effects of estrogen replacement therapy (ERT, conjugated equine estrogen0.625mg orally daily), combined estrogen and continuous progestogen therapy (HRT, 0.625 mg of conjugated equine estrogens plus medroxyprogesterone acetate 5 mg daily) or placebo was compared in 20 postmenopausal type II diabetic women and 20 normal postmenopausal women in a double blind, randomised, crossover study. Patients receiving insulin were excluded from the study and all lipid modifying drugs were ceased at least 4 weeks prior to randomisation. Other medication including oral hypoglycaemics was kept constant for the duration of the study. RESULTS: Women with type II diabetes were a similar age (58.7+/-1.3 years) to the non-diabetic women (59.6+/-1.6 years) but they had a significantly greater body mass index, a higher incidence of treated hypertension, higher fasting plasma glucose levels, higher triglycerides and lower HDL cholesterol levels than non-diabetic women. ERT reduced total cholesterol and LDL cholesterol by a similar extent (8.9-12.3%) in normal and type II diabetic women and increased HDL cholesterol to a similar extent in both groups (11.0 and 8.9% respectively). ERT did not significantly alter fasting triglyceride levels in either group. The addition of medroxyprogesterone acetate 5 mg daily abolished the increase in HDL cholesterol associated with ERT in both groups but did not significantly affect any of the other lipid measurements. ERT and HRT did not significantly alter fasting insulin levels nor alter fasting glucose levels in either non-diabetic women or women with type II diabetes. CONCLUSIONS: ERT and HRT have similar effects on lipids in women with type II diabetes and non-diabetic women after 1 month of therapy. PMID- 10717491 TI - Effects of treatment with 17beta-estradiol on the hypercholesterolemic rabbit middle cerebral artery. AB - OBJECTIVE: The effects of acute and long-term treatment with 17beta-estradiol on the vasomotor responses of rabbit middle cerebral artery (RMCA) were investigated. METHODS: For 8 weeks, male rabbits consumed standard chow (control group), standard chow+1% cholesterol (cholesterol group) or 1% cholesterol chow+17beta-estradiol (i.m. injection 700 microg per week) (estradiol group). The RMCA was precontracted with high K(+) solution and exposed to agonists. RESULTS: Acute exposure to 17beta-estradiol strongly induced relaxation of the RMCA isolated from either control or cholesterol groups. This effect was endothelium independent. Incubation with 17beta-estradiol shifted the calcium contraction curve to the right. High cholesterol diet impaired the relaxation induced by acetylcholine and did not alter relaxation to sodium nitroprusside or to papaverine. Chronic treatment with 17beta-estradiol restored this impaired relaxation to acetylcholine. This protective effect of estradiol was significantly reduced in the presence of N(omega) nitro-L-arginine methyl ester, a constitutive nitric oxide-synthase inhibitor and was not modified in the presence of aminoguanidine, an inducible nitric oxide-synthase inhibitor. Neither tetrabutylammonium, a blocker of calcium-activated K(+) channels, nor glibenclamide, a blocker of ATP-sensitive K(+) channels, affected concentration response to acetylcholine in the RMCA of the estradiol group, whereas 4 aminopyridine, a blocker of voltage-dependent K(+) channels strongly inhibited this relaxation. CONCLUSIONS: These results suggest that acute effects of 17beta estradiol in the RMCA is mediated through blockade of calcium entry into vascular smooth muscle cells, while chronic treatment with this hormone seems to be mediated by release of nitric oxide which activates voltage-dependent potassium channels. PMID- 10717492 TI - Difference in the effect of adiposity on bone density between pre- and postmenopausal women. AB - OBJECTIVES: Elevated bone mineral density (BMD) in obese women is partially attributable to the higher circulating estrogen levels derived from extraglandular aromatization in adipose tissue. However, it remains unclear whether there is an effect of overall adiposity on BMD in both pre- and postmenopausal women. The difference in the effect of overall adiposity on BMD between pre- and postmenopausal women was investigated. MATERIALS AND METHODS: Subjects were 296 premenopausal women with regular menstruation and 233 postmenopausal women. Age, age at menarche, years since menopause (YSM, in postmenopausal women), weight, height, and body mass index were recorded. Total fat mass amount, lean mass amount, and percentage of body fat were measured by whole body scanning with dual-energy X-ray absorptiometry (DEXA). Lumbar spine BMD (L2-L4) was measured by DEXA. In each group, significant determinants of BMD were investigated using univariate and stepwise multiple regression analysis. RESULTS: In postmenopausal women, YSM, lean mass amount, total fat mass amount, and height were significant determinants of BMD (R(2)=0.273, P<0.001). In premenopausal women, only two variables including lean mass amount and age at menarche were significant determinants of lumbar spine BMD (R(2)=0.110, P<0.001), but total fat mass amount and percentage of body fat were not significant determinants of BMD. CONCLUSION: The effect of overall adiposity on BMD is more prominent in postmenopausal women than in premenopausal women. PMID- 10717493 TI - Clinical profile of a new hormone replacement therapy containing 2 mg 17 beta estradiol and 10 mg dydrogesterone. AB - OBJECTIVE: Patient's acceptability, compliance, and effectiveness of a new sequential hormone replacement regimen containing 2 mg 17beta-estradiol and 10 mg dydrogesterone, were assessed in a 3-month, open, multicentre study involving 110 menopausal women. METHODS: A specially designed menopause score was used to assess the severity of menopausal symptoms, each symptom being graded at baseline and after 3 months on a four-point scale. Bleeding data were recorded by the patient on a diary card. Serum hormone levels including FSH, LH, E2, P, PRL, DHEA S, T, SHBG were checked at the initial visit and at the end of the study. RESULTS: After 3 months of treatment, all but four of the 34 climacteric symptoms investigated showed a significant improvement. There were no significant changes noted in body weight. The average duration and flow of bleeding showed no significant changes during hormone replacement therapy (HRT). There were no serious adverse events related to treatment. CONCLUSION: The 17beta estradiol/dydrogesterone combination HRT reduced effectively climacteric symptoms, showed no significant changes in endometrial thickness as determined by transvaginal ultrasonography and provided excellent cycle control. PMID- 10717494 TI - Sympathoadrenal response of postmenopausal women prior and during prolonged administration of estradiol. AB - OBJECTIVE: Cardiovascular disease seems to increase after the menopause and is thought to be reduced by estrogen replacement therapy. Among the many studies which have tried to define the multifactorial mechanisms of estrogens cardiovascular prevention, very few have focused on their possible modulation of adrenergic activity. In the present study we investigated whether prolonged estradiol replacement via transdermal patches is able to modulate cardiovascular and adrenergic responses to stimuli. METHODS: Baseline and responses to a cold stimulus and to the upright position of catecholamines (epinephrine and norepinephrine), heart rate, systolic and diastolic blood pressure were investigated in 15 healthy volunteer postmenopausal women both prior to and after 2 months of treatment with patches rated to deliver 50 microg/day of estradiol. RESULTS: Basal norepinephrine levels (P<0.005), as well as their integrated responses to the cold stimulus (P<0.02) were lower during estradiol. By contrast, responses of norepinephrine to the upright test, as well as basal and responses to stimuli of epinephrine and circulatory parameters were not different before and during estradiol. CONCLUSIONS: Estradiol replacement at low doses significantly decreases overall sympathetic output, both in basal conditions and under specific stimuli. These effects whether maintained or magnified in the long term may play a role in the prevention of the postmenopausal cardiovascular risk. PMID- 10717495 TI - 'Towards the magic bullet'. Hamao Umezawa memorial award lecture. PMID- 10717496 TI - The impact of laboratory reporting practice on antibiotic utilisation. AB - Microbiology reports are often misinterpreted by clinicians, which may lead to inappropriate antibiotic prescribing. Restricted release of susceptibilities combined with interpretative comments, can have a positive impact on the level of appropriate antibiotic use. Such a system requires two-way communication between the laboratory and the clinician and the laboratory's reporting practices should encourage such communication. The production and transmission of clinically relevant microbiology reports should be an integral part of infectious disease management programmes in hospitals. PMID- 10717497 TI - Modelling and forecasting antimicrobial resistance and its dynamic relationship to antimicrobial use: a time series analysis. AB - To investigate the relationship between antimicrobial use and resistance in our hospital, we collected antimicrobial susceptibility and use data from existing microbiology laboratory and pharmacy databases for the period July 1st, 1991 December 31, 1998. The data was analyzed as time series and autoregressive integrated moving average (Box-Jenkins) and transfer function models were built. By using this method, we were able to demonstrate a temporal relationship between antimicrobial use and resistance, to quantify the effect of use on resistance and to estimate the delay between variations of use and subsequent variations in resistance. The results obtained for two antimicrobial-microorganism combinations: ceftazidime-gram-negative bacilli and imipenem-Pseudomonas aeruginosa, are shown as examples. PMID- 10717498 TI - Use of antibiotics in an Italian children's hospital, 1993-1995; clinical and economic considerations. AB - Antibiotic consumption and expenditure was studied during 1993 and 1995 in G. Gaslini children's hospital, an Italian 400-bed paediatric hospital, to see if any changes in use had occurred. There was an increase in the cost of antibacterial agents from 1993 to 1995, with a decrease in the daily cost of antibiotics and in the consumption of antiviral, antifungal and antiparasitic agents. There was a notable increase in the use of glycopeptides and carbapenems between 1995 and 1993 especially in specialities such as onco-haematology and intensive care. We suggest a basis for an antibiotic management programme aimed at reducing costs while still providing a high standard of care for patients. PMID- 10717499 TI - Methicillin-resistant Staphylococcus aureus (MRSA) isolated at Fukuoka University Hospital and hospitals and clinics in the Fukuoka city area. AB - Bacteriological and epidemiological studies were carried out on 106 isolates of methicillin-resistant Staphylococcus aureus (MRSA) isolated at our hospital (56 isolates) and from 15 other hospitals and clinics (50 isolates) in the Fukuoka city area. Strains were studied regarding coagulase-type, beta-lactamase production, and antimicrobial susceptibility; genotype studies used pulsed-field gel electrophoresis (PFGE) with cluster analysis. The majority of isolates produced coagulase type II (75.5%) and beta-lactamase (72. 6%); there was high susceptibility to arbekacin (84.9%) but no resistance to vancomycin. Dendrogram analysis of PFGE patterns identified five major clusters that generally correlated with coagulase-type and beta-lactamase production. Though isolates of two clusters were both coagulase type II and beta-lactamase producing, which was the most common circulating strain both in our hospital and other hospitals and clinics, dendrogram analysis of PFGE patterns showed that they were heterogeneous. Four genetically identical isolates from the same hospital suggested the existence of hospital-specific strains. Nine genetically identical isolates from intensive care units (ICU) in our hospital suggested that a unique strain of MRSA was found there. It had not been transmitted from another area. PFGE with cluster analysis seemed to be an essential tool to detect area-specific or hospital-specific strains undifferentiated by phenotyping. These findings confirmed that a combination of PFGE, including cluster analysis along with coagulase-type and beta-lactamase production may provide more detailed information for the epidemiological study of MRSA. PMID- 10717500 TI - Comparative in vitro activity of gatifloxacin, grepafloxacin, levofloxacin, moxifloxacin and trovafloxacin against 4151 Gram-negative and Gram-positive organisms. AB - Gatifloxacin, grepafloxacin, moxifloxacin and trovafloxacin are fluoroquinolones with enhanced Gram-positive activity while retaining broad-spectrum activity against Gram-negative pathogens. Levofloxacin and ciprofloxacin are older quinolones with broad activity against Gram-negative pathogens and borderline activity against some Gram-positive organisms. We compared the in vitro activity of these compounds against 4151 Gram-negative and -positive organisms. Gatifloxacin, grepafloxacin, moxifloxacin and trovafloxacin were highly active against penicillin sensitive and resistant Streptococcus pneumoniae, Staphylococcus aureus, Streptococcus pyogenes and Streptococcus agalactiae. Ciprofloxacin and levofloxacin were active but less potent. All compounds were highly active (overall) against Gram-negative pathogens with ciprofloxacin being the most active agent against Pseudomonas aeruginosa. Our data indicate that the advanced fluoroquinolones will be important compounds for treating infections caused by Gram-positive and Gram-negative pathogens. PMID- 10717501 TI - Isepamicin versus amikacin for the treatment of acute pyelonephritis in children. AB - In this study we compared the efficacy and safety of isepamicin versus amikacin at a dose of 7.5 mg/kg i.v. q12h for 10-14 days in children with pyelonephritis. Sixteen children were enrolled in the study; ten received isepamicin and six amikacin. Urine cultures grew Escherichia coli in all patients. All patients were treated successfully with either isepamicin or amikacin. Clinical and bacteriological response rates were 100% for both groups. No adverse events occurred. Peak serum levels ranged from 9.05 to 30.70 mg/l (median: 16.165) and from 12.20 to 25.90 mg/l (median: 19.05) for isepamicin and amikacin, respectively. Trough serum levels ranged from 0.11 to 3.20 mg/l (median: 0.75) and from 0.1 to 2.1 mg/l (median: 0.655), respectively. Isepamicin was shown to be as effective and safe as amikacin in the treatment of children with pyelonephritis and might prove an advantageous alternative in areas with high incidence of resistance to other aminoglycosides. PMID- 10717502 TI - Risk factors influencing clinical outcome in Staphylococcus aureus bacteraemia in a Turkish University Hospital. AB - A total of 101 episodes of Staphylococcus aureus bacteraemia were evaluated for the factors influencing prognosis. The overall episode mortality rate and the mortality rate due to bacteraemia were 43.6 and 21.8%, respectively. Episodes with methicillin-resistant S. aureus (MRSA) bacteraemia had a significantly higher overall mortality rate (58.7 vs. 30.9%, P<0.01) and mortality rate due to bacteraemia (32.6 vs. 12.7%, P=0.02) when compared with episodes caused by methicillin-sensitive S. aureus (MSSA). The multivariate analysis revealed that the underlying disease, presence of infective endocarditis, septic shock and central intravascular catheter and methicillin resistance of S. aureus were the five independent risk factors associated with a higher mortality rate. PMID- 10717503 TI - Diarrhoea: a significant worldwide problem. AB - Diarrhoea is a problem, not only of the developing world, but also of the Western world. However, the economic implications of diarrhoeal diseases are particularly evident in the poorer countries. The most common worldwide cause of diarrhoea is intestinal infection and infants, pre-school children, the elderly, and those with congenital or acquired immunodeficiency run a high risk of contracting such infections. Diarrhoeal disease can be classified into three major clinical syndromes: acute watery diarrhoea, bloody diarrhoea, and persistent diarrhoea. A number of different micro-organisms can cause infectious diarrhoea, depending on the clinical setting. The development of oral rehydration solution has provided a simple approach to rehydration and maintenance of hydration in patients with acute watery diarrhoea, and has been implemented worldwide under the auspices of the World Health Organization. However, rehydration does not treat the diarrhoea itself, which will persist until the infection resolves. Since the drugs currently used for the treatment of diarrhoea, such as the opiate agents and antibiotics, have limitations, the search continues for a drug that acts predominantly on secretory pathways without affecting gastrointestinal motility. Novel therapeutic approaches include 5-HT(2) and 5-HT(3) receptor antagonists, calcium-calmodulin antagonists, and sigma-receptor agonists. Another approach has concentrated on the antisecretory role of the neurotransmitter, enkephalin, and has resulted in the development of the enkephalinase inhibitor, racecadotril. This drug has true antisecretory activity, and has demonstrated good efficacy and tolerability in clinical trials. PMID- 10717504 TI - Current and future management of childhood diarrhoea. AB - Diarrhoea continues to have a devastating impact in infants and children. It is a major cause of retarded growth. Substantial declines in hospitalization rates and possibly in the mortality due to diarrhoea have occurred following the launch of programmes based on oral rehydration therapy, and yet about 1 million diarrhoea related deaths occur each year in South-East Asia. The World Health Organization currently recommends oral rehydration therapy plus continued breast- and complementary feeding for children with diarrhoea, and antibiotics for dysentery or associated systemic infection. Although oral rehydration therapy has achieved substantial acceptance, physicians and families continue to prescribe and seek drug therapy to reduce diarrhoeal duration and severity. Research is aimed at developing improved oral rehydration salt solutions or identifying adjunct therapy that will provide substantial benefit in reducing stool output together with safety and selectivity of action. It must, however, be recognized that control of malnutrition is a key requirement to reduce the duration and severity of acute diarrhoea. PMID- 10717505 TI - Racecadotril: a new approach to the treatment of diarrhoea. AB - Since enkephalins were discovered in 1975, it has become clear that they play an antisecretory role in the gastrointestinal tract. Hence a rational research programme was directed at the development of a drug that would preserve these neurotransmitter peptides in the gut by preventing their inactivation. This research programme has resulted in the development of the enkephalinase inhibitor, racecadotril. Preclinical studies have demonstrated the efficacy of racecadotril in two models of hypersecretory diarrhoea: infusion of cholera toxin and castor oil-induced diarrhoea. Moreover, unlike loperamide, racecadotril did not prolong transit time in the small intestine or colon. Further experiments have shown that racecadotril does not promote bacterial overgrowth in the small intestine. Racecadotril lacks any potential for neurotoxicity, and radiolabelled studies have demonstrated that the drug does not enter the brain after oral administration. No potential for abuse or physical dependence has been seen. It is concluded that racecadotril demonstrates specificity of antisecretory action on the gastrointestinal tract without any adverse effect on gastrointestinal motility, and that the results of the preclinical studies indicate the potential usefulness in the treatment of hypersecretory diarrhoea in man. PMID- 10717506 TI - An overview of clinical studies with racecadotril in adults. AB - Since preclinical studies had indicated the potential efficacy and tolerability of racecadotril for the treatment of diarrhoea in man, a series of studies was carried out to assess the clinical effects of racecadotril. These studies were also designed to evaluate whether racecadotril possessed the clinical properties that had been previously identified for an ideal agent to treat infectious diarrhoea. The pure antisecretory action of racecadotril was confirmed in these clinical studies, as was the drug's rapid onset of action. The high therapeutic index of racecadotril was combined with a lack of effect on the central nervous system. Finally, racecadotril was found to be effective in treating acute diarrhoea in double-blind studies against both placebo and the mu opiate receptor agonist, loperamide. The efficacy of racecadotril in acute diarrhoea was not associated with adverse gastrointestinal effects, and its adverse events profile was similar to that of placebo. It was concluded that racecadotril offers a new approach to the treatment of diarrhoea via its mechanism of action as a true antisecretory agent. PMID- 10717507 TI - Vasomotor reactivity in the ophthalmic artery: different from or similar to intracerebral vessels? AB - OBJECTIVE: In order to evaluate hemodynamic features of ophthalmic arteries in patients with severe carotid artery stenosis, we assessed and compared vasomotor reactivity in the middle cerebral and ophthalmic arteries. METHODS: Sixty-five patients (25 symptomatic, 40 asymptomatic) with severe (70-99%) internal carotid artery stenosis were studied using transcranial Doppler and the Diamox test. RESULTS: Vasomotor reactivity was found to be similar in the middle cerebral and ophthalmic arteries on the side of severe carotid stenosis in both symptomatic and asymptomatic patients. In contrast, the vasomotor reactivity of the ophthalmic arteries was significantly different from that of the middle cerebral arteries on the side of the normal or the non-significantly stenotic side of the internal carotid artery. CONCLUSIONS: These data suggest a specific autoregulative response of the ophthalmic artery compared to that of the middle cerebral artery and may shed light on the role of the ophthalmic artery in oculovascular hemodynamics. PMID- 10717508 TI - Ultrasonic assessment of variations in thickness of subcutaneous fat during the normal menstrual cycle. AB - OBJECTIVE: To verify the occurrence of natural variations in thigh and abdominal subcutaneous fat thickness related to the phases of the menstrual cycle, to assess the value of ultrasonography as a reliable method for monitoring subcutaneous fat thickness changes and to evaluate their amplitudes. METHODS: This study included 10 women (19-39 years) who menstruated regularly. None had used oral contraceptives or slimming products during the 3 months prior to the study. At cycle day 2 (CD2), CD6, CD14, CD22, CD27 and CD30 days (CD0: beginning of menstruation), the subjects were submitted to: (1) measurement of weight and thigh perimeters, (2) measurements of thigh and abdomen subcutaneous fatty tissue thickness on B-mode images acquired at 10 MHz. A protocol was designed to guarantee a reproducible repositioning during the whole time course of the study and ultrasound examinations (US) were always performed by the same trained person to avoid inter-examiner variability. RESULTS: Subcutaneous fat thicknesses decreased during the first half of the cycle and reached their lowest values at day 22 (-2.0% for the thighs; -3.3% for the abdominal region). Both thigh and abdomen subcutaneous fat reached their maximum thicknesses during menstruation with respective increases of +2.2 and +4.0%. The observed cyclic amplitude variations in the subcutaneous adipose tissue thickness accounted for 7.3% for the abdominal region and 4.1% for the thighs. CONCLUSION: Variations in adipose tissue thickness during the menstrual cycle could be quantified and monitored by US. The thickness of the thigh and abdominal hypodermis was more important during menstruation and decreased in mid-cycle with a minimum occurring 1 week after ovulation. PMID- 10717509 TI - Assessment of middle cerebral artery diameter after aneurysmal subarachnoid hemorrhage by transcranial color-coded duplex sonography. AB - This study examined if vasospasm after aneurysmal subarachnoid hemorrhage could be visualized by middle cerebral artery (MCA) diameter changes in transcranial color-coded duplex sonography (TCCS). Comparative measurements between mean blood velocity (MBV) and MCA diameter were carried out in 17 patients in 76 instances. At two depth ranges (proximal, 60 55 mm: distal, 50-45 mm) two observers assessed the MCA diameter as indicated by the visualized blood flow column. At both points of measurement, the diameter differences between the two observers were within the ? 2 S.D. range of the mean difference indicating interobserver agreement. In 17 instances, MBV was > 120 cm/s indicating vasospasm but MBV did not correlate with absolute or relative diameter changes. MCA diameter assessment in TCCS seems reproducible. Because TCCS imaging is influenced by several factors comparative angiographic studies are necessary to clarify the TCCS findings. PMID- 10717510 TI - Segmental portal hypertension due to a splenic Echinococcus cyst. AB - A 60-year-old Libyan woman developed perihilar splenic varices without other signs of portal hypertension. Plain abdominal X-ray examination showed two calcified structures in the left and right hypochondria. Ultrasound examination disclosed a 3-cm diameter, globally calcified hydatid cyst lodged in a critical location at the hilar region of the spleen. The cyst was compressing the hilar vessels which resulted in dilatation and varix formation. Another hydatid cyst measuring 5 cm in diameter, with extensive wall calcification was visualized in the right lobe of the liver. The splenic size was within normal limits. The liver revealed normal texture and size and the portal vein was of normal caliber. The patient underwent an uneventful splenectomy and was well at discharge. PMID- 10717511 TI - Ultrasonographic assessment of congestion of the choroid plexus in relation to carbon dioxide pressure. AB - Routine cerebral ultrasound examinations of the neonatal brain often show the choroid plexus to be enlarged, without revealing any other structural pathology. This enlargement might be due to congestion of the choroid plexus as a result of increased cerebral blood flow, due to increased partial pressure of carbon dioxide, amongst other factors. In this exploratory study, 76 cerebral ultrasound examinations, performed on 42 newborn infants within the first 10 days after birth, were analysed retrospectively. The ultrasonograms were classified into three diagnostic groups: normal, congestion of the choroid plexus and haemorrhage. The relationship between the diagnostic groups and the estimated mean arterial partial pressure of carbon dioxide (P(a)CO(2)) was investigated. In the first three postnatal days, the estimated mean P(a)CO(2) in the normal group was significantly lower than in the congestion group (P<0.001). No significant differences were found between the P(a)CO(2) in the congestion and the haemorrhage groups. The findings might support a relation between a high P(a)CO(2) and congestion of the choroid plexus in the first three postnatal days and might be a sign of increased risk for a periventricular haemorrhage. PMID- 10717513 TI - Miniaturised ultrasonic aspiration handpiece for increased applicability. AB - OBJECTIVE: At present, ultrasonic aspiration is routinely used in several fields of surgery, especially in brain and spinal micro-surgery for tumour removal. In order to broaden the access to difficult surgical sites, it is important to design highly miniaturised but still efficient handpieces. The internal resonant system, always made of high-grade materials, must be optimally dimensioned. Normally this is done semi-empirically, by successively improving the design during many iterative test steps. This method however involves several additional difficulties when the degree of miniaturisation increases. For example, small transducer weights exacerbate heat-dissipation problems and make design optimisation important. METHODS: To resolve these problems we have produced modelling software that makes it possible to simulate and automatically tune each individual interacting section of the design before it is actually manufactured, thereby assuring optimal efficiency. RESULTS: Using a new mini-handpiece, designed via the software, two cases of dissection of acoustic neurinomas were successfully performed. CONCLUSION: Using conventional physical steps for improving ultrasonic aspiration handpieces, several problems arise when the grade of miniaturisation increases, due to increasing demands. We have designed computer software for handpiece simulation. Using this model it has been possible to manufacture a highly efficient miniaturised handpiece. PMID- 10717512 TI - Endosonography in gastric lymphoma and large gastric folds. AB - To establish a correct preoperative differential diagnosis between gastric lymphoma and cancer is essential but can be difficult as endoscopic biopsies can sometimes provide a low diagnostic yield. By EUS, infiltrative carcinoma tends to show a vertical growth in the gastric wall, while lymphoma tends to show mainly a horizontal extension. EUS provides an accurate staging of gastric lymphoma, showing the exact level of infiltration and the presence of perigastric lymph nodes, thus the physician can obtain an accurate prognosis for each patient and select the best form of treatment accordingly. The response to chemoradiotherapy can also be investigated very accurately by EUS. Large gastric folds are seen in a great number of benign and malignant conditions. Diagnosis represents a clinical challenge because etiology may be extremely varied and standard biopsies are often inconclusive. Different diseases show different levels of infiltration of the gastric wall, thus a characteristic echo-pattern helps for the differential diagnosis. Endosonography, used always in combination with biopsy, allows to rule out malignancies and to select the most appropriate treatment for each patient (medical or surgical). PMID- 10717514 TI - Effects of fetal and maternal breathing on the ultrasonic Doppler signal due to fetal heart movement. AB - Fetal heart rate (FHR) monitoring is widely used to evaluate fetal health and is based on the detection of movements of the fetal heart by Doppler ultrasound. Fetal health can also be evaluated by prolonged observation of body- and pseudo respiratory movements using two-dimensional ultrasound. Fetal breathing movements are in particular considered to be an important indicator of fetal well being. Ultrasonic Doppler signals caused by movements of the fetal heart were analyzed in detail. The signals were acquired from five healthy fetuses. Minor changes in the insonification geometry gave rise to great variations in the aspects of successive heartbeats. The signals are shown to contain information on both maternal and fetal respiratory movements. This may facilitate the development of a simple method for fetal respiration monitoring coupled to standard FHR monitoring. PMID- 10717515 TI - Platelet aggregation and change in intracellular Ca(2+) induced by low frequency ultrasound in vitro. AB - Treatment of platelet-rich plasma and washed platelets with low frequency ultrasound (US, 22 kHz) at intensity range of 1.0-8.8 W/cm(2) resulted in intensity- and time-dependent platelet aggregation. The effect was absent in calcium-free medium and was initiated by adding supernatant from sonicated suspension (16 W/cm(2), 2 min) to non-treated platelets. A marked decrease in the rate of US-induced aggregation was observed in the presence of specific inhibitors of platelet activation dipyridamole, pentoxifillin, aspirin and verapamil. Concentration of intracellular calcium in washed platelets evaluated with fluorescent probe quin-2 acetoxymethyl ester (quin-2) increased upon sonication in both the calcium containing and calcium free media. It is suggested that US increase of [Ca(2+)](i) is involved in platelet aggregation induced by low frequency US. PMID- 10717516 TI - Radiation stress and its bio-effects. European Committee for Medical Ultrasound Safety (ECMUS). PMID- 10717517 TI - Clinical stage I non-seminomas. Europe versus USA. A draw. PMID- 10717518 TI - Radiotherapy of the brain in elderly patients. Pro:. PMID- 10717520 TI - Radiotherapy of the brain in elderly patients. arbiter: PMID- 10717519 TI - Radiotherapy of the brain in elderly patients. Contra:. PMID- 10717521 TI - Measuring comorbidity in older cancer patients. AB - The aim of this article was to provide oncology researchers with adequate tools and practical advice to integrate comorbidity into clinical studies. Open research questions are also discussed. Commonly used comorbidity indexes were identified and a detailed literature search was done by MEDLINE and cross referencing. Expert opinion was sought on each index. A common scheme exploring the description of the index, clinical experience, metrological performance, easiness of use, cross-compatibility and preservation of data was followed. The actual indexes are included in the Appendix. Four commonly used indexes were identified: the Charlson Comorbidity Index (Charlson), the Cumulative Illness Rating Scale (CIRS), the Index of Coexistent Disease (ICED), and the Kaplan Feinstein index. The Charlson is the most commonly used whereas the performance of the first two indexes is best characterised. Most studies are retrospective and focus on mortality as an outcome and a base of grading. All indexes are easy to use and require a maximum of 10 min to be filled. Inter-rater and test-retest reliability is generally good. Little is known about other outcomes and the way various diseases cumulate in influencing prognosis. Thus, several reliable indexes are available to measure comorbidity in cancer patients. They show that globally comorbidity is a strong predictor of outcome. Since little is still known about the importance of individual comorbidities for various outcomes and the way comorbidity cumulates in influencing cancer treatment, a wide integration of comorbidity in prospective studies is essential. PMID- 10717522 TI - Adjuvant bleomycin, etoposide and cisplatin in pathological stage II non seminomatous testicular cancer. the Indiana University experience. AB - Two cycles of bleomycin, etoposide, and cisplatin (BEP) were evaluated as adjuvant chemotherapy for patients with pathological stage II non-seminomatous germ cell tumours. Between 1985 and 1995, 86 patients with pathological stage II non-seminomatous testicular cancer were treated with two cycles of BEP. At retroperitoneal lymph node dissection (RPLND) 49 patients (57%) had pathological stage II(A) (microscopic nodal metastases) and 37 (43%) had stage II(B) (gross nodal metastases). After RPLND, the patients received bleomycin, 30 units weekly for 8 weeks, etoposide (100 mg/m(2)) and cisplatin (20 mg/m(2)) each for 5 days every 28 days for two cycles as adjuvant chemotherapy. 4 patients were lost to follow-up. 10 patients (12%) developed granulocytopenic fever during their chemotherapy. Of the 82 evaluable patients all remained with no evidence of disease except for a single patient with a cervical nodal relapse of teratoma. This was resected and he remains disease free. Median follow-up has been 85 months (range: 42-173 months). In patients with fully resected stage II non seminomatous germ cell tumour, two cycles of BEP were almost universally effective in preventing relapse. PMID- 10717523 TI - Risk of complications from bone metastases in breast cancer. implications for management. AB - A retrospective analysis of 859 patients who developed bone metastases from breast cancer between 1975 and 1991 was performed in order to identify factors that predict for complications from skeletal disease. The patients were divided into four groups based on the sites of disease at diagnosis of skeletal metastases: bone disease only; bone and soft tissue disease; bone and pleuro pulmonary disease; bone and liver disease. Patients with metastatic disease confined to the skeleton were most likely to develop a pathological fracture. The time to long bone fracture was similar for all groups, but the least number of such fractures occurred in patients with bone and liver metastases since their survival was shortest (median: 5.5 months; P<0.001). Patients with bone metastases only were most likely to require radiotherapy to painful osseous deposits (P=0.0001) and most rapidly developed spinal cord compression (P=0.01, data not shown). The results suggest that patients with disease confined to the skeleton at the diagnosis of bone metastases are most likely to develop skeletal related complications from advanced breast cancer. Such patients may benefit most from treatment with bisphosphonates. PMID- 10717524 TI - Accrual rate-limiting factors in a Swedish randomised ductal carcinoma in situ (DCIS) trial - a demographic study. AB - In the last two decades the introduction of mammographic screening in the Western world has increased the number of diagnosed ductal carcinomas in situ (DCIS) considerably. In situ carcinoma of the breast is considered a heterogeneous disease, the natural history of which is not well known. Thus, appropriate treatment needs to be established. For this reason, a randomised trial studying the effect of breast conserving operation with or without postoperative radiotherapy was instituted in Southern Sweden in 1987. The aim of the present study was to assess patient accrual, identify limiting factors, and evaluate possible ways to influence these factors in order to increase patient accrual. Between 1987 and 1992, 331 patients had been registered with DCIS in the Regional Tumour Registry, 96 of which had been randomised. All 331 were subjected to chart review studying clinical data, mammography reports, cytology and pathology reports to identify inclusion and exclusion criteria according to the design of the trial. It was found that 5% (18/331) had an incorrect diagnosis of DCIS. According to the trial protocol 52% were not eligible (162/313). Fifty-eight per cent (n=88) of the 151 eligible patients had been correctly randomised. The most common reason for exclusion was lesion size. In 21% (66/313) the lesion was 'too large'. Several other limiting factors were identified such as in cytological and pathological definitions and reports, lack of information/awareness in certain physicians, patient reluctance to participate, which in turn may be influenced by the previous factor. With increased information to participating hospitals and considering the above given facts it should be possible to increase accrual from the 28% noted in the present consecutive demographic study to at least one-third of the diagnosed cases of DCIS. PMID- 10717525 TI - p53-regulated GML gene expression in non-small cell lung cancer. a promising relationship to cisplatin chemosensitivity. AB - The GML gene (glycosylphosphatidylinositol-anchored molecule-like protein gene) is a novel gene specifically induced by wild-type p53, which may participate in cell cycle control or the cell apoptotic pathway. Recent experiments suggest that the expression of this novel gene in cancer cells is closely associated with sensitivity to certain anticancer drugs. To elucidate the role of the gene expression in cisplatin (CDDP) chemosensitivity of non-small cell lung cancer (NSCLC), 30 surgically resected materials were examined by reverse transcriptase polymerase chain reaction (RT-PCR). GML gene expression was detected in 9 (30%) samples. Its incidence was significantly higher in immunohistochemically p53 negative (P=0.040) or wild-type p53 tissues (P=0.041). On in vitro chemosensitivity testing using 29 primary tissues, six samples with GML gene expression showed good sensitivity to CDDP. In particular, in tissues with immunohistochemically p53-negative accumulation, those with GML gene expression showed significantly better in vitro sensitivity to CDDP (P=0.012). Clinically a good response to CDDP-based chemo(thermo)therapy for NSCLC patients with tumour residue or recurrence, was observed only in those with p53-negative accumulation and GML gene expression, in agreement with in vitro results. Thus, although the number of tested samples was small, GML gene expression is commonly detected in immunohistochemically p53-negative NSCLCs in close association with good sensitivity to CDDP. GML gene expression analysis may serve as a predictor of CDDP-based chemotherapy for patients with NSCLC. PMID- 10717526 TI - Expression of telomerase component genes in hepatocellular carcinomas. AB - The aim of the study was to clarify the role of telomerase component genes in hepatocarcinogenesis and to examine both the relationship between the expression of telomerase component genes and histological differentiation in hepatocellular carcinoma (HCC) and the relationship between expression levels of telomerase component genes and telomerase activity in HCCs. Telomerase is a ribonucleoprotein enzyme composed of a template RNA and several proteins. Recently, three such telomerase component genes have been identified: human telomerase reverse transcriptase (hTERT); human telomerase RNA component (hTERC); and telomerase-associated protein 1 (TEP1). The expression of these components was evaluated in 34 HCCs and 24 non-cancerous liver tissues by reverse transcriptase-polymerase chain reaction (RT-PCR). Expression of hTERT mRNA was detected in most HCCs, but not in the non-cancerous tissues (P<0.01). Expression of hTERC was detected in both HCCs and non-cancerous tissues, but the expression level in HCCs was higher than that in non-cancerous tissues (P<0.01) and tended to increase as histological differentiation became less marked. The expression level of hTERT mRNA correlated with relative telomerase activity (P<0.01). These results suggest that telomerase reactivation during hepatocarcinogenesis might be regulated by only hTERT and an increase in telomerase activity level in tumour progression might be regulated by both hTERT and hTERC. PMID- 10717527 TI - Combining platinum, paclitaxel and anthracycline in patients with advanced gynaecological malignancy. AB - Two meta-analyses have suggested that the addition of an anthracycline to platinum-based chemotherapy may improve survival in advanced ovarian cancer, and two randomised trials have demonstrated superiority of paclitaxel over cyclophosphamide in platinum combinations. A combination of platinum, anthracycline and paclitaxel would, therefore, be a reasonable experimental arm of any future randomised trial in patients with epithelial ovarian carcinoma (EOC). Patients who required chemotherapy for EOC but were ineligible for standard trials or had other gynaecological tumours that required similar platinum-based chemotherapy were considered for this pilot. The platinum/anthracycline/paclitaxel regimen (G-CAT) was given 3-weekly and consisted of doxorubicin 50 mg/m(2) or epirubicin 60 mg/m(2) intravenously (i.v.) bolus, paclitaxel 175 mg/m(2) (i.v.) over 3 h and either cisplatin 75 mg/m(2) (i.v.) or carboplatin AUC 6, with granulocyte colony-stimulating factor (G-CSF) at the neutrophil nadir. Different combinations were used in order to determine the least toxic regimen. Toxicity and response were assessed according to CTC and WHO criteria, respectively. 26 patients entered the study, 13 with EOC and 13 with other gynaecological cancers (peritoneal, fallopian tube, mixed Mullerian). Median age was 49 years (range: 27-67). 8 patients received carboplatin/doxorubicin/paclitaxel, 8 cisplatin/doxorubicin/paclitaxel and 10 carboplatin/epirubicin/paclitaxel. A total of 135 cycles of chemotherapy were delivered, with a median of 6 cycles per patient (range: 2-6). 54 (40%) cycles required G-CSF support and 17 (65%) patients required at least one dose reduction. All patients experienced grade 4 neutropenia and 13 (50%) patients developed grade 3-4 thrombocytopenia (12 of whom had received carboplatin). There were 4 (15%) patients with grade 3/4 infections but no septic deaths. Non haematological toxicities were manageable, lethargy occurred in 75% of cisplatin treated patients. Grade 1/2 cardiotoxicity, as assessed pre- and post-treatment by left ventricular ejection fraction, was observed in 6/13 (46%) patients who had received doxorubicin and 2/7 (29%) epirubicin-treated patients. No clinically detectable cardiac toxicity was encountered. The response rate in 25 evaluable patients was 76% (12 CR, 7 PR). Dose intensity was highest in the carboplatin/epirubicin/paclitaxel combination. G-CAT shows high activity and can be administered safely, but only very fit patients are suitable for this regimen as it is associated with considerable toxicity. Carboplatin/epirubicin/paclitaxel was the best tolerated regimen overall. PMID- 10717528 TI - Allelic imbalance on chromosome 18 in neuroblastoma. AB - We previously demonstrated that chromosome 18 is frequently deleted in neuroblastoma. To further elucidate the role of chromosome 18 deletions in the development of neuroblastomas we examined 82 cases of neuroblastomas for allelic imbalance (AI) at 17 loci on chromosome 18 to define the common region of AI in neuroblastoma. AI at one or more loci on chromosome 18 was detected in 18/82 (22%) cases. AI on 18q was detected in 17/82 (21%) cases, whereas AI on 18p was detected in 4/82 (5%) cases. There was a distinct common region of AI at 18q21.1 between the D18S363 and D18S858 loci. In addition, cases 16 and 53, which did not show AI at 18q21.1, showed AI at 18pter-q12.3 between the D18S52 and D18S36 loci, indicating that another common region of AI may exist on chromosome 18. AI on chromosome 18 did not significantly correlate with any clinicopathological findings of patients with neuroblastoma. The common region of AI at 18q21.1 includes the DCC gene but not the Smad2 and Smad4 genes. However, our previous studies together with the present study indicated that the incidence of DCC mutation is much less than that of AI at 18q21.1 in neuroblastoma. These results indicate that novel tumour suppressor genes involved in the development of neuroblastoma are present at 18q21.1, and possibly at 18pter-q12.3. PMID- 10717529 TI - Earlier detection of breast cancer by surveillance of women at familial risk. AB - A positive family history increases the risk for breast cancer which oft en occurs at a much younger age than in the general population. We stud ied whether surveillance of these women resulted in the detection of bre ast cancer in an earlier stage than in symptomatic patients with a famil y history. Between January 1994 and April 1998, 294 women with 15-25% r isk (moderate), mean age:43.3 (22-75) years, were screened with a yearly physical examination and mammography from 5 years before the youngest ag e of onset in the family and 384 women with >25% risk (high) for breast cancer, mean age: 42.9 (20-74) years were screened with a physical examination every 6 months and yearly mammography. From September 1995 breast magnetic resonance imaging (MRI) was also carried out for 109 high risk women where mammography showed over 50% density. 26 breast cancers detected under surveillance were significantly more often found in an early T1N0 stage than the 24 breast cancers in patients with a family history referred in that period because of symptoms: 81 versus 46% (P=0.018). Patients under surveillance were also less frequently node-positive than the symptomatic group: 19 versus 42% (P=0.12). 20 patients with a family history referred by our national screening programme in that period had 21 breast cancers detected, 81% in stage T1N0 and 5% node-positive, which was comparable to the results in our national screening programme T1N0 66%, N+ 24% resulting in a 30% reduction in mortality. The incidence in women under surveillance was 10.1 per 1000 in the 'high' risk group and 13.3 per 1000 in the 'moderate' risk group. Expected incidence in an average risk population aged 40-50 years is 1.5, expected if the group consisted of only gene carriers 15 per 1000. 23% of the breast cancers in the surveillance group were detected at physical examination, but occult at mammography. 38% were detected at mammography and clinically occult. Breast MRI (in the subgroup) detected 3 occult breast cancers. The results of this study show that women with a family history benefit from surveillance as breast cancer was detected significantly more often in a favourable T1N0 stage and a mortality reduction comparable to that obtained in our national screening programme may be expected also in women <50 years of age. Both physical examination and mammography contribute to this result, but the former in this study only contributed in women before menopause. Starting surveillance some years before the youngest age of onset in the family may result in higher detection rates. Screening with MRI can detect breast cancers, still occult at physical examination and mammography. PMID- 10717530 TI - Population attributable risk for ovarian cancer. AB - Parity, oral contraceptive (OC) use, age at menopause, a family history of the disease and selected aspects of diet have been related to the risk of ovarian cancer. The quantification of their impact on a population level may help focus and rank the importance of potential prevention strategies. Using data from a case-control study conducted in Italy between 1983 and 1991 on 971 ovarian cancer cases and 2758 control women we computed the multivariate relative risk estimates, and population attributable risks (PARs), i.e. the proportion of ovarian cancers that would have been avoided if a given exposure had not been present in the population. Overall, the PARs were 5% for nulliparity, 12% for never OC use and 4% for a family history of breast or ovarian cancer in first degree relatives. Among women aged >/=50 years, later age at menopause accounted for 16% of all ovarian cancer cases. Low intake of green vegetables accounted for 24% of cases and a high fat score for 7%. All these factors together explained 51% of cases. In conclusion, even if the PAR estimates were based on several arbitrary assumptions, available knowledge could, in principle, explain over 50% of all ovarian cancer cases in this Italian population, thus indicating and quantifying the theoretical scope for prevention. PMID- 10717531 TI - Truncated adenomatous polyposis coli (APC) tumour suppressor protein can undergo tyrosine phosphorylation. AB - Numerous mutations in the adenomatous polyposis coli (APC) gene have been described in colorectal cancer. The vast majority introduce nonsense codons leading to the production of truncated N-terminal APC fragments. Mutations occurring before APC codon 158, have been associated with an attenuated form of familial adenomatous polyposis whereas those occurring at codon 168 or beyond lead to the characteristic form of the disease. These 10 amino acid residues of APC contain a YYAQ motif which appears to constitute a potential SH2 binding domain similar to a sequence present in tyrosine kinase receptors that activate STAT 3 when phosphorylated. We have expressed a recombinant, N-terminal APC fragment in bacterial cells, and shown that it can indeed undergo tyrosine phosphorylation in this domain. We used site-directed mutagenesis to confirm the specificity of the reaction. These observations raise the possibility that tyrosine phosphorylation may be another mechanism involved in controlling APC function. PMID- 10717532 TI - The androgen receptor exon 1 trinucleotide repeat does not act as a modifier of the age of presentation in breast cancer. AB - The CAG repeat in exon 1 of the androgen receptor (AR) genes has been postulated as both a susceptibility allele and phenotypic modifier in BRCA1-associated breast cancers. We have analysed this repeat in a set of 178 breast cancer cases who have been selected only for age of presentation at 65 years or less. No effect of repeat length on age of presentation was found and there was no association between repeat length and family history. In combination with the data from other workers, our findings suggest that the androgen receptor repeat does not act as a modifier gene or susceptibility locus outside the context of the hereditary breast/ovarian cancer syndrome. PMID- 10717534 TI - Human papilloma virus status and chromosomal imbalances in primary cervical carcinomas and tumour cell lines. AB - Human papilloma virus (HPV) infection is the crucial step in the initiation of cervical carcinomas. In addition, HPV18 has been implicated in tumour progression and adverse clinical outcome. We determined the HPV types in 12 primary cervical carcinomas and 12 cell lines and compared the findings with the comparative genetic hybridisation (CGH) pattern of chromosomal alterations. The most frequent alteration was the deletion at 3p14 followed by the loss of 2q34-q36 along with 3q gain. High risk HPV types were detected in all samples except one primary tumour. In contrast to the normal distribution, HPV18 was present in 75% of cases including all cell lines. The cell lines carried a higher number of genetic alterations and a different CGH pattern for several chromosomes than the primary tumours, despite microdissection. Purely HPV18 positive cases indicated a high incidence of imbalances at specific loci with peaks of the histogram coinciding with known HPV integration sites. The study suggests that HPV infection is associated with a recurrent pattern of chromosomal changes in cervical carcinomas and that the development and progression of these alterations is triggered by integration into the host genome. PMID- 10717533 TI - Mechanisms of enhancement of cytotoxicity in etoposide and ionising radiation treated cells by the protein kinase inhibitor wortmannin. AB - We have investigated the effects of the protein kinase inhibitor wortmannin (WM) on the cytotoxic mechanisms of etoposide and ionising radiation (IR) in the Chinese hamster ovary K1 (CHO-K1) cell line, and its radiation-sensitive derivative, xrs-6, which is defective in DNA-dependent protein kinase (DNA-PK) function. WM potentiated the cytotoxicity of etoposide and IR in CHO-K1 cells approximately 1.6 and 3-fold, respectively, and this potentiation was abolished in xrs-6 cells, which were themselves more sensitive to etoposide and IR alone. WM partially inhibited the repair of etoposide-induced DNA double-strand breaks. Etoposide treatment caused a biphasic inhibition of DNA synthesis in both cell lines, and this was abrogated by co-incubation with WM. These data suggest that WM inhibits in intact cells both DNA-PK and either or both the ataxia telangiectasia (AT) and AT-related gene products ATM and ATR. PMID- 10717535 TI - In vivo interactions of rabbit enteropathogenic Escherichia coli O103 with its host: an electron microscopic and histopathologic study. AB - A family of human and animal pathogens, including enteropathogenic and enterohemorrhagic Escherichia coli (EPEC and EHEC), trigger formation of 'attaching and effacing' lesions on cultured and intestinal epithelial surfaces. However, our understanding of these events in vivo is incomplete. To further study these interactions in a natural infection model, weaned rabbits were infected with rabbit enteropathogenic E. coli O103 (REPEC O103), followed clinically, and infected tissues were evaluated by electron and light microscopy. Of the 36 rabbits challenged, morbidity and mortality were 65 and 23%, respectively. Twenty-four hours after infection, expression of fimbriae-like organelles was observed on the bacterial surface. Microvilli of ileal Peyer's patches (PP) became disorganized, and intestinal mucus secretion increased which coincided with intraluminal binding of the pathogen in the proximal colon. Forty eight hours after infection, there was conspicuous lack of fimbriae-like organelle expression, while bacterial adherence preferentially occurred at the domed villi of PP. Seventy-two hours after infection, broad morphological heterogeneity was noted within pedestals beneath attached bacteria, including extended pseudopods. We conclude that REPEC O103 express surface organelles during initial exposure to the host, that the initial target sites of adherence are the domed villi of ileal PP, and that increased mucus secretion occurs during REPEC O103 infection. As well, extended pseudopod formation was demonstrated in vivo. PMID- 10717536 TI - Clinical signs, reproduction of attaching/effacing lesions, and enterocyte invasion after oral inoculation of an O118 enterohaemorrhagic Escherichia coli in neonatal calves. AB - Attaching and effacing (AE) lesions are produced among others by enteropathogenic Escherichia coli and enterohaemorrhagic E. coli (EHEC), which differs from the former by the production of cytotoxins active on various cell cultures, the verocytotoxins, or shigacytotoxins. EHEC are associated with diarrhoea and dysentery in humans and in ruminants, mainly calves from two to eight weeks of age. Clinical signs and/or lesions have been reproduced experimentally with EHEC strains belonging to serotypes O5:K4/Nm, O26:K-:H11, O111:Nm, and O157:H7 which are isolated from cattle and/or humans. The purpose of this work was to develop an experimental model of infection in newborn calves with a bovine EHEC strain isolated from a calf which of died of diarrhoea, and belonging to the O118:H16 serotype, which is also common to both cattle and humans. The bovine O118:H16 EHEC strain was able to colonize the gut of three newborn calves, and to induce diarrhoea twenty-four hours after challenge and to produce AE lesions in the small and/or large intestines. AE lesions were detected microscopically and ultrastructurally in the small intestine of one calf and in the whole intestinal track of two calves. Internalization of bacteria and also of pedestal-bacteria complex inside of the enterocyte was observed in two of the three calves. The significance of this stage is unknown but may be related to the invasion of the calf by the bacteria. The challenge strain was isolated from the mesenteric lymph nodes of the same two calves but not from other organs or from heart blood. No blood was observed in the faeces of any of the three calves, nor were any lesions in the internal organs, which may have been related to the production of a verotoxin whose role is still unknown in cattle. PMID- 10717537 TI - Current epidemiology of human plague in Madagascar. AB - From 1996 to 1998, 5,965 patients with suspected plague were identified in 38 districts of Madagascar (40% of the total population are exposed). Using standard bacteriology, 917 of them were confirmed or presumptive (C + P) cases. However, more than 2,000 plague cases could be estimated using F1 antigen assay. Two out of the 711 Yersinia pestis isolates tested were resistant to chloramphenicol and to ampicillin (both isolates found in the harbour of Mahajanga). Urban plague (Mahajanga harbour and Antananarivo city) accounted for 37.4% of the C + P cases. Bubonic plague represented 97.2% of the cases, and the lethality rate was still high (20%). In comparing the exposed population, plague was more prevalent in males (M:F sex ratio 1.3:1) and patients under 20 years (2.7% babies under two years). Buboes were mainly localised in the inguinal/femoral regions (55.8%). The epidemiological risk factors are discussed. PMID- 10717538 TI - Stability-related studies on 17D yellow fever vaccine. AB - Yellow fever (YF) vaccine using the 17D strain of YF attenuated virus has been produced at the Institut Pasteur in Dakar since 1962. Until now, the stabilised YF had an expiry date of utilization of two years from the end of the lot control process under storage at +4 degrees C. We conducted a stability study to assess the three full year validity of this preparation, when correctly stored at +4 degrees C to optimise the conditions of production, storage and availability of such a vaccine. The activity of 19 consecutive batches of vaccines kept for three years at +4 degrees C was compared to that of the same batches that were kept three years at -20 degrees C. Using the in vitro microculture method, we found that three-year storage at +4 degrees C induced a higher loss of activity than storage at -20 degrees C or than the accelerated degradation test of vaccines kept for 14 days at 37 degrees C. Whatever the conditions of storage, in all cases decreases in activity were below the WHO's requirements, i.e., < 1 log PFU/dose, and residual activity of the selected batches was over 1000 mouse LD50 per dose. We demonstrated that the 17D YF vaccine produced in Dakar has a shelf life of three years and that its required potency was maintained at +4 degrees C, after reconstitution with saline diluent, following three-year storage at +4 degrees C. PMID- 10717539 TI - Ebola and Marburg virus antibody prevalence in selected populations of the Central African Republic. AB - With the natural history of the filovirus family seemingly unknown, filovirus ecology in its natural environment remains a rudimentary field of research. In order to investigate the maintenance cycle of filovirus in Central Africa, a study was conducted within the rain forest of the Central African Republic. The epidemiological study determines the frequency and distribution of filovirus seroprevalence in a selected human population. Using an ELISA, serum samples from Pygmy and non-Pygmy populations were tested for Ebola-Zaire virus and Marburg (MBG) virus antibody. Filovirus antibody reacting sera were found in all zones investigated, and in all populations studied (Ebola virus IgG 5.3%; Marburg virus IgG 2.4%). Pygmies appeared to have a significantly higher seroprevalence (P < 0.03) against Ebola-Zaire virus (7.02%) than non-Pygmies (4.2%). MBG virus or related unknown filovirus strains also seem to be present in the western part of Central Africa. MBG virus antibodies were present in different Pygmy groups (ranging from 0.7 to 5.6%, mean 2.05%) and in several non-Pygmy populations (ranging from 0.0 to 3.9%, mean 3.4%) without an overall significant difference between the two groups (P = 0.14). The potentialities of nonpathogenic filovirus strains circulating in the Central African Republic are discussed. PMID- 10717541 TI - Unique mechanism of Helicobacter pylori for colonizing the gastric mucus. AB - Helicobacter pylori is a human gastric pathogen causing chronic infection. Urease and motility using flagella are essential factors for its colonization. Urease of H. pylori exists both on the surface and in the cytoplasm, and is involved in neutralizing gastric acid and in chemotactic motility. H. pylori senses the concentration gradients of urea in the gastric mucus layer, then moves toward the epithelial surface by chemotactic movement. The energy source for the flagella movement is the proton motive force. The hydrolysis of urea by the cytoplasmic urease possibly generates additional energy for the flagellar rotation in the mucus gel layer. PMID- 10717540 TI - Potential effect of cattle diets on the transmission of pathogenic Escherichia coli to humans. AB - Grain feeding seems to promote the growth and acid resistance of Escherichia coli in fattening beef cattle, and acid-resistant E. coli are more likely to survive the human gastric stomach. When cattle were fed hay for only five days, the number and acid resistance of E. coli decreased dramatically. PMID- 10717542 TI - Pneumocystis carinii mutations associated with sulfa and sulfone prophylaxis failures in immunocompromised patients. AB - Recent studies have shown that mutations in two amino acid positions of the Pneumocystis carinii dihydropteroate synthase gene are significantly more common in immunocompromised patients with P. carinii pneumonia who fail sulfa or sulfone prophylaxis. This paper reviews the studies that suggest that these mutations may be responsible for some failures of prophylaxis in P. carinii. PMID- 10717543 TI - Telomeres and HIV disease. AB - Telomere measurement, envisioned as a novel approach to elucidate T-cell dynamics in HIV disease, failed to reveal any consistent pattern in CD4+ T cells. By contrast, significant telomere shortening, as well as other hallmarks suggestive of replicative senescence, was observed within the CD8+ T-cell subset. Telomere studies have thus provided unanticipated insight into a novel facet of memory CD8+ T lymphocyte dynamics that may explain the exhaustion of the protective antiviral immune response. Strategies aimed at manipulating replicative senescence, therefore, offer unique approaches to immune reconstitution. PMID- 10717544 TI - Regulation of cell growth and death by Epstein-Barr virus. AB - Epstein-Barr virus (EBV) efficiently induces growth of human B cells and prevents cell death. Considerable progress has been made in understanding these processes, the role of EBV in human cancer cells and the relationship of viral gene expression to virus persistence and cancer. PMID- 10717545 TI - Modulation of intramacrophage iron metabolism during microbial cell invasion. AB - The mechanisms whereby vertebrate hosts withhold iron from microbial invaders, as well as the methods used in attempts by invaders to capture the growth-essential metal, differ between host extracellular and intracellular environments. This review focuses on procedures employed by host cells and by invaders, respectively, that alter intramacrophage iron metabolism. PMID- 10717546 TI - Microbial metalloproteases and pathogenesis. AB - Zinc metalloproteases produced by human pathogenic microorganisms show a wide variety of pathological actions. In local infections, the proteases cause necrotic or hemorrhagic tissue damage through digestion of structural components of the ground substance, and also form edematous lesions through generation of inflammatory mediators, while in systemic infections, the proteases act as a synergistic virulence factor through disordered proteolysis of many plasma proteins. Clostridial neurotoxins, Bacteroides fragilis enterotoxin and Bacillus anthracis lethal factor are also zinc metalloproteases. PMID- 10717547 TI - A morphological analysis of the macro motor unit potential. AB - The technique of macro EMG is used to investigate the motor unit architecture in a number of pathological conditions. Amplitude and area are the most commonly used criteria, but these parameters alone are not sufficient to assess the complexity of the macro MUP morphology. In an attempt to examine the morphology of the macro MUP in more detail, additional measures were investigated including, (i) average power, (ii) duration, and (iii) number of phases. Macro MUP duration was defined as the time parameter that contains a particular fraction (90%) of the total power of the potential. The above mentioned parameters were evaluated for normal subjects and for patients suffering with motor neuron disease (MND), spinal muscular atrophy (SMA), and Becker's muscular dystrophy (BMD). It is shown that high amplitude and average power macro MUPs give shorter macro MUP duration than macro MUPs with normal amplitude. In contrast, in low amplitude macro MUPs there is a tendency towards a higher duration measure, as compared with the duration of the normal amplitude macro MUPs. Also, t-test results for the duration measure gave a significant difference between the NOR-MND, and no significant difference between the NOR-BMD and NOR-SMA groups at P<0. 05. Significant difference between the NOR and the three disease groups investigated was obtained for the parameters log amplitude, log area, and log average power. The number of phases was not significantly different between the NOR and the rest of the groups. In conclusion, the average power and duration parameters can possibly be used as additional discriminators to detect abnormalities of the macro motor unit potential in both needle and surface EMG but further investigation is necessary. PMID- 10717548 TI - Physiological variability of peak latency in evoked potentials: use of a property of asynchronous averaging. AB - A method for estimating the physiological variability of peak latency in an evoked potential, of whom the peak latency and amplitude varied, by using the property of asynchronous averaging was proposed. A point estimate of the physiological variability was obtained by minimizing the mean square error between a standard waveform and the asynchronously averaged waveform. An interval estimate was obtained by using the relationship between the signal-to-noise ratio and the standard deviation of the point estimate for the variability of peak latency. The proposed method was evaluated by using simulation data, and was successfully applied to actual P300 data of six normal subjects. The method gave accurate estimates for the physiological variability of peak latency of P300, and would also be effectively applicable to any other evoked potential records for estimating the physiological variability. PMID- 10717549 TI - Finite element modeling of the cervical spine: role of intervertebral disc under axial and eccentric loads. AB - An anatomically accurate, three-dimensional, nonlinear finite element model of the human cervical spine was developed using computed tomography images and cryomicrotome sections. The detailed model included the cortical bone, cancellous core, endplate, lamina, pedicle, transverse processes and spinous processes of the vertebrae; the annulus fibrosus and nucleus pulposus of the intervertebral discs; the uncovertebral joints; the articular cartilage, the synovial fluid and synovial membrane of the facet joints; and the anterior and posterior longitudinal ligaments, interspinous ligaments, capsular ligaments and ligamentum flavum. The finite element model was validated with experimental results: force displacement and localized strain responses of the vertebral body and lateral masses under pure compression, and varying eccentric anterior-compression and posterior-compression loading modes. This experimentally validated finite element model was used to study the biomechanics of the cervical spine intervertebral disc by quantifying the internal axial and shear forces resisted by the ventral, middle, and dorsal regions of the disc under the above axial and eccentric loading modes. Results indicated that higher axial forces (compared to shear forces) were transmitted through different regions of the disc under all loading modes. While the ventral region of the disc resisted higher variations in axial force, the dorsal region transmitted higher shear forces under all loading modes. These findings may offer an insight to better understand the biomechanical role of the human cervical spine intervertebral disc. PMID- 10717550 TI - A method for determining the heat transfer and water vapour permeability of patient support systems. AB - The formation of pressure ulcers can be exacerbated by a breakdown in the integrity of the patient's skin caused by poor maintenance of the skin microclimate. Patient support systems (PSSs-specialised beds, mattresses, chairs, cushions and pads) play an important role in the dissipation of heat and moisture away from the skin/support interface which is necessary in order to maintain the physiological skin microclimate. This paper reports a laboratory method and theory for the simultaneous measurement of the heat and water vapour dissipating properties of PSSs. The results demonstrate that the method is extremely selective, exhibiting very significant differences between the PSSs tested. It also shows that assessing PSS covers independently does not necessarily indicate the overall performance of the complete PSS. PMID- 10717551 TI - Classification of normal and pathological tremors using a multidimensional electromagnetic system. AB - A new multidimensional movement analysis system was used to record limb tremor over six degrees-of-freedom, and signal processing techniques were explored to develop a suitable classification method to distinguish between different types of tremor. The specific aims were to investigate the ability of the system to screen for differences between normal subjects and a group of neurological patients, and then to differentiate between three diagnostic groups of patients. Postural tremor at the hand was recorded in normal subjects (n=24) and patients with essential tremor (n=21), multiple sclerosis (n=17) and parkinsonism (n=19). Data were collected using a 3Space Fastrak((R)) (Polhemus, Inc.) over six degrees of-freedom (three translational directions and three rotations). Spectral estimates produced measures of tremor frequency and amplitude. Mathematical models of the data, using autoregressive modelling and K-nearest neighbour classification, produced parameters used to classify, (1) the normal subjects and 24 patients (using the three rotational movements), and (2) the three patient groups (using all six movement directions). Results were given in terms of the probability of each subject belonging to the groups being classified. 70%). The diagnostic classification produced clear differences between the patient groups (60% for essential tremor, 80% for multiple sclerosis and 60% for parkinsonism). The ability of this assessment technique to distinguish between postural tremor in normal subjects and neurological patients suggests that it could be developed as a screening tool. Classification of tremors between the patients groups, with a high degree of sensitivity, indicates the potential for further development of the system as a diagnostic aid. PMID- 10717552 TI - An inexpensive sensor for measuring surface geometry. AB - A technique to measure surface geometry using a conductive ink sensor is described. In the human system distorted cylinders are common, and geometry can be reconstructed from local measures of curvature. An algorithm is presented to reconstruct the shape of a surface from a series of curvature measurements. The Abrams Gentile Entertainment patented bend sensor was evaluated as a curvature transducer. The sensor was tested at the extremes of the likely measurement range, from curvatures below 0.01 mm(-1) up to a curvature of 0.1 mm(-1). The upper curvature limit proved beyond the design specification of the sensor. The technique was applied at the lower curvature range to reconstruct one quadrant of the chest of a volunteer and record breathing movement. The bend sensor is inexpensive and can be applied to obtain an approximate reconstruction of surface geometry in the human system. PMID- 10717553 TI - Filtering of electromyogram artifacts from the electrocardiogram. AB - Electromyogram (EMG) artifacts often contaminate the electrocardiogram (ECG). They are more difficult to suppress or eliminate, compared for example to the power line interference, due to their random character and to the considerable overlapping of the frequency spectra of ECG and EMG signals obtained from the same pair of electrodes. The usually applied low-pass filtering (cutoff frequency of minimum 35 Hz) results in limited suppression of the EMG artifact and considerable reduction of sharp Q, R and S ECG wave amplitudes. A solution to this problem is proposed by applying approximation filtering with dynamically varied number of samples and weighting coefficients, depending on the ECG signal slope. The slope measure used is the absolute value of the product of the tilts of two adjacent 10 ms segments sliding along the signal. The results obtained show a slight widening of some sharper QRS complexes, but a virtual preservation of their amplitudes and a considerable reduction of the EMG artifact. PMID- 10717554 TI - [Best wishes for 2000]. PMID- 10717555 TI - [PubMed Central: a metamorphosis of print-on-paper publishing?]. PMID- 10717556 TI - [Worse late than never]. PMID- 10717557 TI - [Arteriovenous malformations: a study of 200 cases]. AB - OBJECTIVE: The aim of this study was to analyze the natural history of arteriovenous malformations by reviewing 200 consecutive cases observed in all localizations between 1992 and 1996 in a multidisciplinary angioma clinic. PATIENTS AND METHODS: Files concerning 200 arteriovenous malformations were reviewed with a standardized observation sheet applying the severity criteria defined by the International Society for the Study of Vascular Anomalies. We used the Schobinger staging system which includes 4 grades of severity: grade I=dormancy, grade II=expansion, grade III=destruction, grade IV=cardiac decompensation. RESULTS: There was no predominance by gender. Cephalic localizations were the most common. The malformation was present at birth as indicated by history taking in 40 p. 100 of the cases. Progression during childhood (grade II) was observed in 84 p. 100. At the first consultation, the patients generally were more often in grade II than in grade I or III. Bone destruction was observed in 3 cases, signs of cardiac decompensation in 5. Arteriovenous malformations were part of a more complex syndrome in 9 cases. We noted a flare-up in the prepuberty or puberty period in 75 p. 100 of the cases and the possible role of puberty (64 cases), trauma (39 cases) and pregnancy (25 p. 100 of the adult women). Explorations were generally completed (Doppler, arteriography) by grade II, expressing the need for a map of expanding lesions. Finally it was difficult to assess posteriorly the beneficial or deleterious effect of the often multiple treatments prescribed for these patients. DISCUSSION: The review of these 200 cases pointed out the "pediatric" nature of the problem of arteriovenous malformations in the large majority of the cases and the often misleading presentation of these vascular anomalies, particularly grade I malformations. A false aspect of capillary malformation could raise the risk of inappropriate treatment. This review also confirmed known factors of exacerbation (puberty, pregnancy, trauma) and demonstrated the severity of these vascular anomalies which can progress with loco-regional expansion and invasion without a cellular proliferation component. The analysis of treatments used showed that embolization alone cannot definitively and totally control a superficial arteriovenous malformation and that no one treatment, even combined embolization and large excision, can be a guarantee to provide total cure. PMID- 10717558 TI - [Neonatal curettage of giant congenital nevi]. AB - OBJECTIVE: All agree upon the need for early treatment of giant congenital nevi, basically because of the risk of melanoma degeneration, estimated at about 5 p. 100. Another reason is the cosmetic, psychological and social impact of such nevi. The aim of this study was to assess neonatal curettage of giant congenital nevi as an alternative to classical surgery. PATIENTS AND METHODS: Between 1996 and 1999, the curettage technique was used in 14 newborns with giant congenital nevi. Three nevi were located on the scalp, 4 on lower limbs and 7 on the trunk with a jacket configuration in 1 case and a cape configuration in 4. RESULTS: Curettage achieved 70-95 p. 100 clearing of the giant nevi in 10 of the 14 children. Four of the children developed hypertrophic scar tissue which resolved with time. Secondary hair growth was observed in 5 cases. Outcome was better when the curettage was performed very early (before 2 weeks of life). DISCUSSION: Curettage is a surface technique proposed when surgical excision cannot be performed because the surface is too large or the localization is incompatible with surgery. Curettage is a simple low-cost technique which provides particularly satisfactory cosmetic results for very extensive giant congenital nevi. The risk of malignant transformation is greatly reduced although not totally. Regular clinical surveillance under conditions greatly improved by the clearing should help reduce the risk. PMID- 10717559 TI - [Cerebral magnetic resonance imaging (MRI) in the diagnosis of leptomeningeal carcinomatosis in melanoma patients]. AB - OBJECTIVE: Meningeal involvement is frequent in metastatic melanoma, approximately 30% in autopsy series. Functional signs may be misleading and the neurological examination may be normal. Certain diagnosis requires identification of tumor cells in the cerebrospinal fluid. CSF cytology is however sometimes negative and magnetic resonance imaging (MRI) with gadolinium injection may provide the diagnosis. The aim of this retrospective study was to assess the role of imaging in the diagnosis of leptomeningeal carcinomatosis. PATIENTS AND METHODS: The diagnosis of leptomeningeal carcinomatosis was made in 8 patients between 1992 and 1998. All had signs of neurological function impairment, but the neurology examination was abnormal in only 2. RESULTS: Cytology examination of the cerebrospinal fluid provided the diagnosis of leptomeningeal carcinomatosis in 5 patients. One out of 5 brain CT scans were positive, showing meningeal enhancement confirmed by brain MRI. The spinal tap was not contributive in 2 cases and was not done in 1. In these three cases, the brain CT did not provide any diagnostic element while the brain MRI with gadolinium injection confirmed the diagnosis of leptomeningeal carcinomatosis. DISCUSSION: Forty-one percent of patients with autopsy proven leptomeningeal carcinomatosis have a normal ante mortem spinal tap. Brain MRI with gadolinium injection has better sensitivity than brain CT scan. All patients with nonspecific neurological signs and a normal spinal tap should be explored with a brain MRI. PMID- 10717560 TI - [Comparative diffusion of fusidic acid, oxacillin, and pristinamycin in dermal interstitial fluid after repeated oral administration]. AB - OBJECTIVE: The aim of this study was to use the suction bullae technique to compare skin diffusion of 3 antibiotics commonly used for skin infections (fusidic acid, oxacillin, pristinamycin) and to estimate their potential activity at the site of skin infections. SUBJECTS AND METHODS: This comparative open study was conducted in 12 healthy volunteers using a repeated latin square experimental scheme. Antibiotic concentrations in serum and suction bullae fluid were measured by high performance liquid chromatography after 5.5 days of repeated oral administration of fusidic acid (1 g/d), oxacillin (2 g/d), and pristinamycin (2 g/d). RESULTS: Mean antibiotic concentrations in serum and interstitial fluid (suction bullae fluid) were highest for fusidic acid with a Cmax at 91.3 +/- 23.0 mg/l and 45.5 +/- 18.0 mg/l respectively (interstitial fluid/serum ratio=49 +/- 10 p. 100). For oxacillin, Cmax was 8.3 +/- 3.6 mg/l and 0.98 +/- 0.49 mg/l (ratio 13 +/- 5 p. 100). Pristinamycin concentrations were low with a Cmax at 0.51 +/- 0.40 and 0.26 +/- 0.15 mg/l (ratio 73 +/- 57 p. 100). Comparing the area under the interstitial fluid and the serum concentration-time curves showed that the best diffusion was obtained with pristinamycin (114 +/- 61 p. 100), followed by fusidic acid (57 +/- 13 p. 100) and oxacillin (48 +/- 25 p. 100). DISCUSSION: These data were used to calculate indicators of potential efficacy in the interstitial dermal fluid: inhibitor quotient (Cmax/MIC) and AUIC (ASC/MIC), indicator of the time antibiotic concentrations are maintained above the minimal inhibitor concentration (MIC). This showed that fusidic acid was potentially more active against all staphylococci. For streptococci, the observed interstitial concentrations of pristinamycin and of fusidic acid should theoretically inhibit streptococci A growth, but oxacillin was the most adapted antibiotic. PMID- 10717561 TI - [Detection of human papillomavirus in cutaneous extragenital Bowen's disease in immunocompetent patients]. AB - INTRODUCTION: A specific link between human papillomavirus (HPV) types 16, 18, 31, and 33 and genital carcinomas and between HPV type 5 and cutaneous extragenital carcinomas in patients with epidermodysplasia verruciformis and renal transplant has been previously found. The aim of this prospective study was to detect HPV in cases of cutaneous extragenital Bowen's disease (BD) from non immunosuppressed patients. PATIENTS AND METHODS: Twelve cases of cutaneous extragenital BD or Bowen's carcinoma (BC), seen in the period 1994-1996 and confirmed by histologic examination, were included in the study. Tissue sections were studied by in situ hybridization with a mixture of HPV DNA probes and specific HPV DNA probes. In addition, study on fresh materiel from 1995 included: Southern blot hybridization with various usual HPV probes (6, 11, 16, 18, 31, 33, 35, 39, 42), polymerase chain reaction (PCR) with hybridization using consensus HPV probes and probes specific for HPV types 6, 11, 16, 18 and 33. In positive samples with conventional PCR, in situ PCR with probes specific for HPV types 6/11 and 16 was performed on tissue sections. RESULTS: In situ hybridization was negative in all the cases. Southern blot hybridization was negative in our 9 studied cases. Three cases studied by consensus PCR were positive. PCR with specific HPV probes revealed positivity on two of these cases: HPV 6 in one, and HPV 16 in another. In situ PCR was positive with a mixed 6/11 HPV probe in the third positive consensus PCR case. DISCUSSION: Our study revealed the presence of HPV in 3 out of 12 cases of cutaneous extragenital BD and BC. HPV type 16, found in BC of skull, was the most usually found type in the literature. HPV types 6/11, detected in 2 cases, were rarely found in cutaneous extragenital BD and BC and these results are in favor of the oncogenic effect of these virus types. In our study, in situ hybridization and Southern blot hybridization were negative in all the cases; HPV was only found in 3 cases by conventional PCR and in 1 case by in situ PCR. The low range of detection of HPV in cutaneous extragenital BD may be due to the used methods, to difficulties related to sampling and/or to a low number of copies of the HPV genoma. PMID- 10717562 TI - [Ambulatory skin grafting in leg ulcers: a feasibility study of 34 patients]. AB - OBJECTIVES: Despite the advent of modern dressings, management of leg ulcers remains a long costly process, particularly if no etiological treatment is possible. Autologous skin grafting is more and more widely used in this indication. The aim of this open single center noncomparative study was to analyze the feasibility of ambulatory procedures for skin grafting and the incidence of ambulatory care in a medical nursing clinic as an alternative to traditional hospitalization on total cost in this pathological condition. PATIENTS AND METHODS: Thirty-nine grafts were performed in 34 consecutive patients. No selection was made for etiology or duration of the leg ulcers. Three grafting techniques were used after debridement-cleansing: flap grafts for medium sized ulcers (29 cases), mesh grafts for large ulcers (6 cases) and patch grafts for small ulcers or ulcers with irregular contours (4 cases). The dressing was opened on day 5, nursing care was provided every 2 days and daily in case of infection. Percentage of healing was evaluated clinically on days 5, 15 and 30 then at months 3, 6 and 12. Photographs were taken. RESULTS: Four patients were lost to follow-up and one died. Among the 34 grafts assessed at 6 months, we obtained total healing in 56 p. 100, 75 p. 100 healing in 6 p. 100, 50 p. 100 healing in 9 p. 100 and failure in 29 p. 100. Healing rates were those expected for arterial ulcers and necrotic angiodermas. For venous leg ulcers, the rate of total healing was only 30 p. 100 at 6 months and 43 p. 100 at 1 year. Outcome depended on duration of the lesion and not on the type of skin graft or patient age. DISCUSSION: This prospective study reports outcome of ambulatory skin grafting in a large representative sample of patients with leg ulcers of various etiologies. The less favorable outcome for venous ulcers can be explained by the duration of the ulcerations and infection in these often neglected lesions. The risk of graft displacement, contact eczema, and infection must be recognized for early treatment. There were no cases with general complications. This ambulatory technique has the enormous advantage of limiting the risk of hospital-related problems in this elderly population and of reducing overall cost of care for leg ulcers, and finally of limiting the risk of recurrence by regular post-graft follow-up in a specialized center and by treatment of the causal disease. PMID- 10717563 TI - [Alpha interferon-induced eczema in atopic patients infected by hepatitis C virus: 4 case reports]. AB - OBJECTIVES: We report four cases of eczema induced by alpha interferon in atopic patients treated for chronic hepatitis C. CASE REPORTS: Eczema developed in 4 patients with certain (3 cases) or possible (1 case) atopy treated by subcutaneous injections of alpha interferon for hepatitis C virus infections. Delay to onset was 3 weeks to 6 months. Interferon was highly likely the causal agent: lesions started at site of interferon injection, followed the rhythm of interferon injections (three cases), disappeared at interferon withdrawal. In two patients, the lesions diffused to other sites. Both Introna and Roferon were used. Three patients also took ribavirine. The possible role of a contact factor (antiseptic.) was ruled out. Skin tests (patch tests, prick tests, intradermal reactions) were negative for interferon alpha and for a standard battery. DISCUSSION: The role of interferon in the induction of skin diseases or its influence on the course of certain dermatoses is well known. In atopic patients, interferon might induce eczema via an immunomodulator rather than an allergic mechanism since skin tests (performed in one patient) were negative. This observation is similar to that in psoriasis induced by interferon in predisposed subjects who develop skin lesions at injection sites which sometimes diffuse to distant localizations. The role of other factors (hepatitis C virus infection, ribavirine) remains unknown; they might participate in this mechanism by aggravating skin dryness. PMID- 10717564 TI - [Serum S100B protein and stage of cutaneous melanoma: a prospective study]. AB - OBJECTIVE: We searched for a correlation between serum S100B protein and cutaneous malignant melanoma stage, according to the American Joint Committee on Cancer staging system, and between elevation of serum S100B protein and development of metastasis. PATIENTS AND METHODS: We conducted a prospective bicentric study for 20 months in 122 patients with malignant melanoma. LIA-mat(R) assay was used to determine serum S100B at each examination. The optimal cut-off value was determined from the ROC curve. RESULTS: The optimal cut-off value to discriminate patients with metastases from patients in remission was 0.09 microg/l. Sensitivity was 46 p. 100 in patients with stage III and 86 p. 100 in patients with stage IV disease. The positive predictive value for stage III/IV was 77 p. 100 and the negative predictive value was 89 p. 100. Serial measurements were made in 56 patients. The serum S100B protein level was elevated in 69 p. 100 of the patients disclosing disease progression (9/13). In 44 p. 100 of the patients (4/9), serum S100B protein level rose within a delay of 3 months before new metastases were detected. DISCUSSION: Our study confirms that serum S100B protein is a tumor marker for melanoma staging and follow-up. A rise in S100B protein precedes or reveals metastasis. PMID- 10717565 TI - [Ultra-late metastasis of melanoma with secondary dissemination along the venous stripping line]. AB - BACKGROUND: A woman was cured of a melanoma of the leg. She presented an ultra late recurrence 17 years later, then rapid and extensive recurrence immediately after venous stripping. CASE REPORT: In 1977, a 38-year-old woman was operated for a SSM melanoma of the calf. She was then given chemotherapy and BCGtherapy for 2 years. In 1994, she presented a nodular recurrence which was treated surgically. In March 1997, she underwent a stripping of the leg. One month later, a nodular recurrence was discovered along the stripping line. DISCUSSION: The ultra late recurrence (> 15 years) in this case is highly exceptional and, to our knowledge, this is the first case of a recurrence along a vascular course. We discuss metastatic dissemination of SSM melanoma and the role played by stripping in this case of recurrence. PMID- 10717566 TI - [Icthyosiform erythroderma: atypical manifestation of sarcoidosis]. AB - BACKGROUND: Specific cutaneous manifestations of sarcoidosis can be extremely varied. We report one observation of cutaneous sarcoidosis with an exceptional clinical presentation: icthyosiform erythroderma. CASE REPORT: A 33-year-old woman had a dry icthyosiform erythroderma associated with arthralgia and muscular deficit of the pelvic belt in a context of fever and weight loss. Skin biopsy revealed the presence of an epithelioid giant-cell granuloma. This granuloma was also found in the salivary glands and in the bronchial mucosa. The chest X ray showed diffuse reticularis opacities. The diagnosis of sarcoidosis was retained. The patient was treated with oral prednisone (0.75 mg/kg/day). Total skin clearing was obtained after 15 days. DISCUSSION: This observation points out the heterogeneous nature of clinical manifestations of cutaneous sarcoidosis. The icthyosis erythroderma form is exceptional. Correct diagnosis is difficult and must be based on clinical, histological, radiological and biological criteria of sarcoidosis. PMID- 10717567 TI - [Tumoral mycetoma of the buttock]. AB - BACKGROUND: Mycetomas are actinomycosic or fungal infections where the infectious agent produces grains. We report an atypical case of fungal mycetoma presenting as a tumoral formation on the buttock. CASE REPORT: A 56-year-old unemployed man from the Diourbel region of central Senegal consulted in February 1997 for a fistulalized tumor of the right buttock which had developed spontaneously and progressed for 5 years. The patient's general health remained satisfactory. Physical examination showed a voluminous 25 cm tumefaction extending from the right buttock to the perineum. The tumor showed a few areas of fistualization which discharged black grains under pressure. There was a 1.5 cm right inguinal node which did not appear to be inflammatory. The remainder of the physical examination was normal. Pathology reported inflammatory connective tissue surrounding a brownish polycyclic grain composed of spores and mycele filaments. The diagnosis of fungal mycetoma was retained and surgical excision under general anesthesia was performed. DISCUSSION: This is an atypical case of fungal mycetoma because of its tumoral form and gluteal localization. The differential diagnosis was cutaneous neurofibroma, myoma, or Darier-Ferrand dermatofibrosarcoma. The frequency of extrapodal red grain mycetomas has been pointed out by several authors from Senegal, but extrapodal black grain forms as seen in our case are more exceptional. PMID- 10717568 TI - [Localized bullous pemphigoid following radiotherapy]. AB - BACKGROUND: Bullous pemphigoid is the most frequent autoimmune blistering dermatologic disease. Induction of pemphigoid has been observed after administration of certain drugs but also after various irradiation procedures. We report a case of pemphigoid strictly confined to the irradiated area. CASE REPORT: A 66-year-old woman had been irradiated 16 years before because of a breast carcinoma. She presented with a blistering rash strictly confined to the irradiated area. The diagnosis of pemphigoid was confirmed with pathologic examination, direct immunofluorescence, direct immunofluorescence of sodium chloride-separated skin, electron microscopy and immunoelectron microscopy. Indirect immunofluorescence and serum immunoblot were also positive. The patient was successfully treated with topical steroids. DISCUSSION: The location on the area of radiotherapy and the limited character of lesions were unusual. A few cases of pemphigoid strictly localized to the area of radiotherapy have been published. However, in our case, only direct immunofluorescence and immunoelectron microscopy reliably confirmed the diagnosis. The unusual location of this pemphigoid is not due to the regional variation in the expression of the antigen. Other hypotheses concerning the pathogenesis of localized pemphigoid should be examined. PMID- 10717569 TI - [POEMS syndrome revealed by a scleroderma-like skin thickening]. AB - BACKGROUND: POEMS syndrome is a rare form of plasma-cell dyscrasia characterized by the various association of Polyneuropathy, Organomegaly, Endocrinopathy, Monoclonal component and Skin changes. The most frequent skin changes such as hyperpigmentation, hypertrichosis, scleroderma-like skin thickening and angiomas are not pathognomonic but nearly constant. However, they are rarely isolated. CASE REPORT: A 57-year-old Caucasian woman presented with a 6 month history of skin thickening on both hands and feet, Raynaud's phenomenon and facial telangiectasias. Physical examination on presentation revealed hepatomegaly, signs of a sensorimotor peripheral neuropathy which was demyelinating in type on electrophysiological assessment, and ankle edemas. Initial laboratory investigations revealed a platelet count of 900 000/mm(3), a monoclonal IgG lambda gammapathy. Plasma-cells were slightly increased (10 p. 100 of marrow elements) and full skeletal radiographs showed no focal osteosclerotic or lytic lesion. A diagnosis of POEMS syndrome was made. The patient was treated with tamoxifen, methylprednisolone and plasmapheresis without improvement in polyneuropathy or in skin changes. DISCUSSION: Our patient satisfied the criteria for POEMS syndrome. The most typical feature here was the scleroderma-like skin change, which has been recognized by other authors. But, in the present case, Raynaud's phenomenon, skin thickening and facial telangiectasias were present 6 months before the diagnosis, and initially suggestive of systemic scleroderma, confirmed histologically. The pathogenesis of POEMS syndrome might be regarded as the result of a marked activation of the proinflammatory cytokine network, but an increase in serum Vascular Endothelial Growth Factor (VEGF) levels could well account for other manifestations such as skin thickening. PMID- 10717570 TI - [Granuloma faciale: efficacy of cryosurgery in 2 cases]. AB - BACKGROUND: Granuloma faciale is a rare condition of unknown pathogenesis. Treatment often gives less than satisfactory results. We report two cases successfully treated with cryosurgery. CASE REPORT: A 42-year-old woman had a round 65 x 40 mm erythematous violet-colored papulous plaque on the right cheek. Histology reported a dense inflammatory infiltration of the superficial and mid derma around dilated vessels under a normal epidermis. The infiltration was composed of histiocytes, lymphocytes, plasma cells, neutrophils and eosinophils and was compatible with granuloma facial. Surgical treatment was performed but was followed by recurrence within a few months. Class I local steroids, cotton swab cryotherapy and oral dapsone remained ineffective. Cryosurgery however led to cure without recurrence at 1 year follow-up. The second patient was a 52-year old man with an oval reddish-brown papulous plaque on the right preauricular area. The plaque measured 40 mm in diameter and histology favored granuloma facial. Three cotton swab cryotherapy sessions and one surgical excision procedure were followed by recurrence. Cryosurgery provided successful cure without recurrence at 3.5 years follow-up. DISCUSSION: Most destructive treatments for granuloma facial, including pulverization cryotherapy, surgery, and CO(2) or argon laser are unsuccessful in preventing recurrence. Cryosurgery was proposed in 1977 but few cases have been reported since. Medical treatments have been variously successful with most authors reporting unsatisfactory results. In our two cases, surgical excision was rapidly followed by recurrence and in one case dapsone was ineffective. Cryosurgery provided rapid regression of the lesion and recurrence-free cure at 1 and 3.5 years follow-up respectively. Cryosurgery is an effective treatment for readily recurrent eosinophilic granuloma facial and should be proposed as first intention therapy. PMID- 10717572 TI - [Superficial cheilitis and angular cheilitis]. PMID- 10717571 TI - [Eruptive xanthogranulomatosis in a trisomy 21 patient with acute lymphoblastic leukemia]. AB - BACKGROUND: Juvenile xanthogranulomas may be associated with myeloproliferative disorders, usually juvenile chronic granulocytic leukemia. CASE REPORT: We report the case of a 25-year-old man with Down's syndrome who presented eruptive xanthogranulomas. At this time, the patient was being treated for acute lymphocytic leukemia. The eruption increased with an extensive unusual "satellite" arrangement despite apparent remission of the acute leukemia. Leukemia relapse occurred some months later. DISCUSSION: This case had an unusual clinical presentation. It also suggests their might be a link between the clinical course of lymphocytic leukemia and juvenile xanthogranuloma. PMID- 10717573 TI - [Yellow nail syndrome]. PMID- 10717574 TI - [Angiocentric cutaneous lymphoproliferative disorders]. PMID- 10717575 TI - [Lupus and pregnancy]. PMID- 10717576 TI - [Pacemakers and dermatology]. PMID- 10717577 TI - [Topical therapy in dermatology: pharmaceutical forms, excipients and bases for magistral preparations]. PMID- 10717578 TI - [Early treatment of sexual exposure to human immunodeficiency virus]. PMID- 10717580 TI - [Taxane-induced ungual anomalies]. PMID- 10717579 TI - [Pharmacokinetics: elementary notions]. PMID- 10717581 TI - [Centers for Disease Control and Prevention (CDC)]. PMID- 10717582 TI - [ [In Process Citation] PMID- 10717583 TI - [Epidemiology of brain metastases]. AB - The incidence of intracerebral metastasis is largely underestimated. They appear to be the most frequent cause of intracerebral tumors with an average incidence of probably more than 12/100 000 inhabitants per year. Depending on reports, 25 to 35% of cancer patients will develop brain metastasis. In neurosurgical series, the frequency of cerebral metastasis among surgical procedures for brain tumors increased from 3.5 to 12%. We reviewed the literature in order to better know the frequency of intracerebral metastasis depending on the type of primary cancer. The rate of other metastatic lesions, the duration between discovery of the primitive cancer and brain metastasis were found to be useful for selection of patients for neurosurgical treatment. Three types of cancers could be defined, regarding the incidence of brain metastasis. While occurrence of brain metastases is very high, surgery should often be useless; if brain metastases only occur in the late stage of a cancer, then, surgery should be discussed according to the general status of the patient; although brain metastasis is rare in a cancer, surgery will often be required for diagnosis and treatment. PMID- 10717584 TI - [Biology of brain metastases: current concepts]. AB - Brain metastasis results from complex interactions between host cells and primitive tumor cells. An analysis of the molecular pathways at the cellular level is provided in this review of the literature. The principal new therapeutic modalities are directly based on our comprehension of those molecular biology hypothesis. PMID- 10717585 TI - [Neurosurgical multicenter study of cerebral metastases (Neurosurgical departments of Paris Beaujon, Paris Sainte-Anne, Strasbourg)]. AB - We report a multicentric outcome study of cerebral metastases in 174 patients collected consecutively in three neurosurgical centers in France. Age and condition of patients allowed surgical operation; the sex ratio was 1.67. The revelation mode was usual. However tumoral hemorrhage frequency was 3.4%. Primitive cancers were usual cancers as in oncological series with a higher proportion of radioresistant metastases. Mean dimension was 30 mm with mean of 1. 25 metastases per patient. Supratentorial localization was more frequent and the metastasis revealed cancer in 40% of cases. Global median survival was 12.1 months. Factors correlated with the survival in our series of patients with cerebral metastases were: solitary cerebral metastasis, extracerebral extension of cancer, treatment of primitive cancer, complete excision and post operative cerebral radiotherapy. PMID- 10717586 TI - [Surgical management of cerebral metastases]. AB - Surgical treatment of metastatic brain tumors had been controversial until two prospective randomized trials demonstrated that surgery followed by radiation is superior to radiation alone in patients with single metastasis. In this report, we reviewed current perspectives on the neurosurgical treatment of brain metastasis. The goals of surgery is to establish a histologic diagnosis, relieve symptoms and provide local cure through gross total resection. The results which established surgery as the preferred treatment for single brain metastasis were largely a result of advances in neurosurgical techniques of localization and neuroimaging, microdissection, and functional mapping. These modern methods have reduced the operative mortality to 3% and the morbidity to less than 10%. The indications of surgical treatment should be related to clinical and radiological criteria. Surgical treatment is best indicated for single and accessible metastasis and is also accepted for patients with multiple lesions especially when there is a life-threatening lesion or when two lesions are accessible through the same craniotomy or for recurrent brain metastasis. Surgery can also be a good option for recurrent metastasis. Altogether, surgery improves quality of life and the median survival time, all the more as pronostic factors are present such as age>60 years, long interval between diagnosis of the primary tumor and the metastasis, the absence of systemic disease and a Karnofsky score> 70. PMID- 10717587 TI - [Cerebral metastases: radiotherapy and chemotherapy]. AB - Brain metastases are common events in adult patients with solid tumors. The choice of the optimal therapy is still challenging and controversial. Whole brain radiotherapy (WBRT) is a standard practice in most patients with an excellent palliative effect. Boost to gross disease has also been advocated without a clear benefit. Moreover following extended irradiation, a substantial proportion of the long term survivors (>6 months), will present documented cognitive impairments. Patients with favorable prognostic factors can benefit from more aggressive therapy: local resection, mono or multifractionated irradiation with or without radiosensitizing agents, stereotactic radiotherapy, brachytherapy. Although brain metastases of solid tumors occur in the presence of progressive widespread disease, chemotherapy has played a limited role in their treatment. Poor drug penetration across the normal blood-brain barrier of chemotherapy agents is not a limiting factor because of the neovascularization in the tumor. The few prospective studies that have addressed this issue, especially in lung and breast tumors, are reviewed. PMID- 10717588 TI - [Radiosurgery for brain metastases]. AB - This article presents the state of art in radiosurgery as a new therapeutic strategy of brain metastases. Radiosurgery of brain metastases is ten years old and we propose to make an update of the literature. X-rays and gamma-rays are commonly used for radiosurgery and does not make a difference for dosimetry and precision. Selected patients for this treatment have usually good Karnofsky status and the mean size of their metastases is about 20 mm. Mean number of metastases treated in the same procedure is 2. Most frequent primary sites are lung and breast, but also kidney and skin (malignant melanoma) which are supposed to be radioresistant and so often untreatable by other techniques. Usual dose administrated ranges between 15 and 25 Grays in center of target with a 80% isodose in peripheral. Results are quite good for local control (80% to 100%) but survival does not seem to be improved (11 months). Radiation complication rate is 4% and sometimes hemorrhagic complications occur (1 to 2%). For local control, quality of life and cost/benefit ratio, there are strong arguments in favor of radiosurgery, especially for radioresistant metastases in spite of the lack of improvement of survival duration. Moreover published studies do not allow any comparison with efficacy of surgery and radiosurgery, whole brain radiosurgery and radiosurgery of the present metastases. Evaluations are still going on in several centers. Their results will allow more precise indications of this technique. PMID- 10717590 TI - [ [In Process Citation] PMID- 10717589 TI - [Therapeutic strategy]. PMID- 10717591 TI - [Thoracic spinal cord compression by a gouty tophus. Case report. Review of the literature]. AB - An unusual case of thoracic spinal cord compression caused by extradural tophaceous deposits is reported in a 59-year-old female with a long-standing history of gout involving the metatarsophalangeal joints. T1 and T2 magnetic resonance images of the spine illustrated an extradural hyperintense signal extending from T2 to T9. A decompressive laminectomy disclosed a white caseum like material in the extradural space, together with a small organized hematoma. Histologic examination showed areas of amorphous substance containing urate crystals surrounded by inflammatory cells, which was diagnosed as a gouty tophus. The patient made an uneventful recovery after surgery. Fifteen similar cases of the literature are reviewed. Although spinal involvement by gout seems relatively common, a compression of the spinal cord or of the cauda equina in gout patients seems exceptional. The diagnosis should be considered in patients showing a relevant history of gout, but spinal cord compromise may also represent the initial manifestation of the disease. PMID- 10717592 TI - [Intracranial meningiomas revealed by hemorrhage. Report of three cases and literature review]. AB - Three cases of meningioma revealed by hemorrage are reported. The first patient, 65 presented with a temporal hematoma associated with intraventricular contamination. The diagnosis of a tumoral origin of the hemorrhage was made only few weeks later when the total hematoma resorption was ended. The second patient, 49 presented with a sudden right hemiplegia. CTscan and MRI revealed a left frontal tumor surrounded by hematoma. The third patient, 46, presented with a transient aphasia and superior limb dysesthesia. CT scan showed a temporal hematoma; a carotid angiography revealed a typical meningioma blush. A review of the literature collected 54 additional cases of meningioma associated with hemorrage. Clinical findings and mechanisms are discussed. PMID- 10717593 TI - [Recurrent bleeding of thalamic cavernous angioma under hormonal treatment. A case report]. AB - A case of recurrent bleeding from a probable left thalamic cavernoma in a 26 year old woman taking hormonal treatment is reported. Four episodes of bleeding were clinically and radiologically documented, prior to her referral to our institution. Interestingly, each episode occurred three weeks after starting hormonal treatment, dydrogesterone, desogestrel ethinylestradiol, chlormadin, nomegestrel acetate). The patient was not operated because of the deep situation of the cavernoma which was remote from the thalamic surface within the third ventricle. There was no recurrent bleeding after the onset hormonal treatment was discontinued. Although no similar case has been found in the literature, we believe that this case gives further argumentation in favor of a role of hormonal factors influencing the biological behavior of cavernous angiomas which has been previously suggested in pregnant females with bleeding cavernous angiomas. PMID- 10717594 TI - [Cervical spine infection with Streptococcus anginosus. Case report]. AB - A case of cervical spine infection due to Streptococcus anginosus is reported. Streptococcus milleri is encountered in the mouth, gastro-intestinal tract, vagina and nasopharynx. It is an uncommon pathogen responsible of suppurative infections such as brain liver or spleen abscesses, intra-abdominal or soft tissue abscesses and pleural empyema. In rare cases it can cause spondylodiscitis and osteomyelitis. Based on the review of eight cases of spondylodiscitis or osteomyelitis, diagnosis and treatment are discussed. PMID- 10717595 TI - [Growing fracture of the orbital roof. A case report]. AB - We report a case of growing fracture of the orbital roof in a 5-year-old child. The presenting sign was a pulsatile orbital mass. This child had a history of a minor head injury with orbital impact 2 years ago. Cerebral CT scan revealed a diastatic fracture of the right orbital roof. On MRI a leptomeningeal cyst extending in the orbital cavity was shown. Frontal craniotomy with direct repair of the dural and bone defects was performed. The outcome was excellent. In the literature the exact pathophysiology of the growing fractures is still debated but a dural laceration along the fracture line is noted in all the cases. They are mostly located in the cranial convexity, and rarely affect the skull base. Only 5 similar cases were found in the relevant literature. Growing fracture of the orbital roof should be suspected if ocular symptoms appears in childs who have sustained a head injury several months or years ago. PMID- 10717597 TI - [ [In Process Citation] PMID- 10717596 TI - [Multiple cerebral hydatic cysts of cardiac origin. A case report]. AB - The hydatic cyst is a pathology observed in developing countries. Multiple cerebral localizations with a cardiac origin are exceptional and are sometimes diagnosed only after onset of complications. We present the case of a 22-year-old male student: the diagnosis of multiple cerebral hydatic cysts caused by rupture of a cardiac hydatic cyst was established after an episode of ischemia of the limbs with cerebral hemorrhage induced by heparin. One year later, the clinical situation consisted in intracranial hypertension, hemiplegia and convulsive seizures. We observed no cardiac symptoms. The brain CT showed 9 hydatic cysts and echocardiography showed a myxomatous cystic tumor. Abdominal CT detected renal and splenic hydatic cysts. The patient underwent total ablation of the cerebral and abdominal cysts and made a full recovery. After surgical removal of the cardiac cyst, the patient has been lost to follow-up. Cerebral hydatidosis of cardiac origin is highly exceptional and, due to nonspecific symptomatology, may go undiagnosed. In general, prognosis is good in case of a cerebral localization but the cardiac localization is associated with high mortality. PMID- 10717598 TI - Efficiency of donor screening for human parvovirus B19 by the receptor-mediated hemagglutination assay method. AB - BACKGROUND AND OBJECTIVES: Mass screening for parvovirus B19 (B19) by receptor mediated hemagglutination assay (RHA) may be inadequate to eliminate the virus from plasma pools. We characterized B19 carriers detected in blood donor screening by RHA to explain why some carriers were not detected by RHA. MATERIALS AND METHODS: Donor plasma was screened for B19 by RHA, B19 DNA by nested polymerase chain reaction (PCR) assay, and anti-B19 by enzyme immunoassay. RESULTS: B19 DNA-positive specimens (n = 73) screened from 456,665 donors were divided into group A (n = 41) with high DNA and high RHA titers, group B (n = 16) with low DNA and low RHA titers, and group C (n = 16) RHA-negative. Most (37/41) of the group A samples were without anti-B19, while most (15/16) of the group B samples were positive for anti-B19. Group C specimens were screened by PCR from 3, 042 random RHA-negative specimens. Analysis of samples from early infections revealed that the viremic period, corresponding to group A, lasted only 8-10 days after infection. The RHA reactivity fell rapidly with the appearance of anti-B19 and disappeared 28-30 days after infection, thus corresponding to group B. The RHA reactions of group B specimens were often unstable, probably because of the formation of immune complexes. The B19 DNA-positive, RHA-negative state lasted for several months, which corresponded to group C. CONCLUSION: Only group A specimens are reliably eliminated in donor screening by RHA. Therefore, although donors with high B19 DNA could be screened out by testing for B19 by the RHA, most B19 carriers, with low B19 DNA and RHA-negative, will not be eliminated. PMID- 10717599 TI - Clinical experience with a new solvent detergent-treated intravenous immunoglobulin free of hypotensive effects. AB - OBJECTIVE: To see if modifications to the processing of intravenous immunoglobulin to include a virus inactivation stage alter immunoglobulin G (IgG) resulting in hypotension in patients. METHODS: Clinical trials were done involving extensive patient monitoring during infusion: in vitro - testing for markers of hypotension, and in vivo - an animal model which closely simulates clinical use. RESULTS: No hypotensive response was seen in the animal model or clinical trial. CONCLUSIONS: The production process used does not damage IgG or create vaso-active kinins as the preparation was free of hypotensive effects. PMID- 10717600 TI - Clinical tolerance of methylene blue virus-inactivated plasma. A randomized crossover trial in 12 healthy human volunteers. AB - OBJECTIVES: Baxter's methylene blue (MB) photoinactivation of fresh frozen plasma (FFP) is a method that effectively removes any intracellular potentially infectious agent, such as prions, by filtration combined with an improved photochemical virus inactivation on a single unit of FFP. The purpose of this study was to demonstrate clinical and biochemical tolerance of MB-treated plasma in healthy human volunteers. METHODS: The design was a crossover randomized trial. Twelve subjects received alternatively treated and nontreated autologous FFP. Coagulation parameters were tested as well as clinical and biochemical parameters. RESULTS: No clinical or statistically significant difference was detected for any of the parameters. CONCLUSION: Transfusion of FFP treated for viral reduction by an improved MB photoinactivation method was shown to be safe and did not affect the normal function of coagulation proteins. We consider this method as a valuable alternative to other methods described as it combines the photoinactivation with a filtration step removing any intracellular infectious agents, such as prions. PMID- 10717601 TI - Determination of granulocyte-specific antigens on neutrophil FcA receptor IIIb by PCR-preferential homoduplex formation assay, and gene frequencies in the Japanese population. AB - BACKGROUND AND OBJECTIVES: Granulocyte-specific antigens play an important role in provoking immune neutropenia and transfusion reactions. We developed a new DNA typing method, PCR-preferential homoduplex formation assay (PHFA), to determine granulocyte-specific antigens on the neutrophil Fcgamma receptor IIIb (FcgammaRIIIb, CD16b), namely, the NA1, NA2, and SH antigens and their gene frequencies in the Japanese population. MATERIALS AND METHODS: Four hundred unrelated healthy Japanese blood donors were typed using PCR-PHFA. To confirm the accuracy of the results of FcgammaRIIIB genotyping using PCR-PHFA, PCR-sequence specific primer (SSP) typing and PCR-restriction fragment length polymorphism (RFLP) typing were carried out in another 20 samples for comparison. RESULTS: The results of PCR-PHFA typing agreed well with other methods. The frequencies of the FcgammaRIIIB alleles were 62.2, 37.8, 0 and 0% for NA1, NA2, SH, and 'NA-null', respectively. CONCLUSION: The PCR-PHFA method can be semi-automated easily with computer-based assignment and is suitable for typing both small and large numbers of samples. In the Japanese population, the frequency of NA1 is about double that in Caucasians (32.5%), and the SH allele is rare. PMID- 10717602 TI - The direct antiglobulin test: still a place for the tube technique? AB - OBJECTIVES: Antibodies of different immunoglobulin isotypes, or complement, may coat red blood cells in vivo. They are detected by the direct antiglobulin test (DAT), usually performed by the conventional tube technique. The purpose of this study was to compare the latter technique with the gel test. METHODS: Three hundred and ninety-eight consecutive samples, sent to our laboratory for direct antiglobulin testing, were analyzed with the tube technique and the gel test, using reagents from different manufacturers. Eighty-seven samples had been collected from newborns and 23 from fetuses. Results were expressed as positive or negative. RESULTS: In 162 out of 398 cases, the DAT was negative with both methods, whereas in 178 out of 398 cases, the DAT was positive with both techniques using polyspecific antibodies (observed agreement: 84.5%; kappa = 0.71). Discrepancies between the two methods were observed in 58 cases: 51 samples appeared as DAT positive using the tube method and negative with the gel test, whereas only 7 were positive exclusively with the gel test. Among the 178 samples that were positive with both techniques, 93 samples showed discordant results when evaluated with monospecific antisera (11 with anti-IgG and 82 with anti-C3d, respectively). The sensitivity of the DAT performed by the gel test, in comparison with the conventional tube technique, was 75.4% (95% confidence interval (CI): 69.4-80.8). 96. 8% (95% CI: 92.8-99.0), and 16.3% (95% CI: 9.8 24.9) with polyspecific, anti-IgG and anti-Cd3 reagents, respectively. CONCLUSIONS: The gel test appeared to be less sensitive than the conventional tube technique when utilized for DAT, particularly when C3d was present on red blood cells. These results emphasize that before implementing a new technique in the laboratory, comparison with existing techniques, using different reagents, is mandatory. PMID- 10717603 TI - Typing for the human lewis blood group system by quantitative fluorescence activated flow cytometry: large differences in antigen presentation on erythrocytes between A(1), A(2), B, O phenotypes. AB - BACKGROUND: Lewis phenotyping by hemagglutination is an unreliable routine method for Lewis antigen designation. Now genomic typing of the Lewis gene is available. Additionally, flow cytometry has been used for typing. We wanted to compare the results of Lewis typing in healthy individuals using the three methods. MATERIALS AND METHODS: Ninety-three randomly selected plasma donors were genotyped for inactivating Secretor (FUT2) G428A and Lewis (FUT3) T59G, T202C, C314T, G508A and T1067A point mutations. All Le(a+b-) individuals (nonsecretors) were homozygous for the FUT2 G428A mutation and all Le(a-b-) individuals had inactivating mutations on both FUT3 alleles. Fixed erythrocytes were analyzed by fluorescence activated flow cytometry and the results were compared with hem- agglutination and genotypic data. Antigen availability was expressed as median fluorescence intensity and as percentage positive cells with fluorescence intensities > or =10(2). RESULTS: Using an anti-Le(a) reagent a mean of 99% of erythrocytes from Le(a+b-) individuals and 1% of erythrocytes from Le(a-b-) or Le(a-b+) individuals were stained positive. Using an anti-Le(b) reagent, a mean of 71% of erythrocytes from A(1), 95% from B and 99% from O and A(2) Le(a-b+) individuals and less than 10% of erythrocytes from Le(a-b-) or Le(a+b-) individuals were stained positive. After papain treatment 100% of the erythrocytes from A(1) and A(1)B Le(a-b+) individuals stained positive without increase in background staining. The flow cytometric technique revealed large differences in staining intensities, within each ABO Le(a-b+) subgroup which was not directly correlated to plasma donation frequencies nor to Secretor or Lewis genotypes. CONCLUSION: Flow cytometry may prove valuable as a Lewis blood group typing technique but also as a research tool when investigating Lewis phenotypes of human erythrocytes. PMID- 10717604 TI - Malaria antibody ELISA insufficiently sensitive for blood donor screening. PMID- 10717605 TI - Norfloxacin-induced positive direct antiglobulin test. PMID- 10717606 TI - Prostate carcinoma practice patterns: what do they tell us about the diagnosis, treatment, and outcomes of patients with prostate carcinoma? PMID- 10717607 TI - The breast carcinoma screening interval is important. PMID- 10717608 TI - Histopathologic findings predicting lymph node metastasis and prognosis of patients with superficial esophageal carcinoma: analysis of 240 surgically resected tumors. AB - BACKGROUND: If it were possible to elucidate the histopathologic findings predicting lymph node metastasis and prognosis in superficial squamous cell carcinoma of the esophagus (SSCCE), they could be used as markers to identify patients who do not require additional surgical resection after endoscopic mucosal resection (EMR). METHODS: Two hundred forty surgically resected SSCCEs were examined histopathologically. Histopathologic factors including vertical tumor invasion depth in the submucosal layer (VTIDsm), degree of nuclear atypia (low, one point; high, two points), growth pattern (expansive, one point; infiltrative, two points), and histologic grade (calculated by adding the latter two scores to obtain Grade 1, two points; Grade 2, three points; and Grade 3, four points) were evaluated to investigate the associations among these factors, lymph node metastasis, and prognosis. RESULTS: No lymph node metastasis was found in 54 patients with carcinoma limited to the lamina propria. Their 5-year survival rate was 100%. Multivariate analysis of 186 carcinomas invading beyond the lamina propria showed that lymphatic permeation correlated with lymph node metastasis (P<0.0001) and the presence of lymph node metastasis and a high histologic grade were independent factors indicating a poor prognosis (P = 0.0061 and 0.023, respectively). In 53 patients whose tumors had invaded the lamina muscularis or slightly invaded the submucosa (VTIDsm <500 microm), no lymph node metastasis was found in the lymphatic permeation negative and blood vessel permeation negative patients with VTIDsm values <200 microm and histologic Grades 1 or 2. CONCLUSIONS: Lymphatic permeation is a good predictor of lymph node metastasis in patients with SSCCE. Lymph node metastasis and the histologic grade are independent prognostic factors. Vessel permeation, VTIDsm, and histologic grade were found to be important factors for identifying patients who did not require additional surgical treatment after EMR. PMID- 10717609 TI - Quality of life in survivors of colorectal carcinoma. AB - BACKGROUND: Colon carcinoma is a common malignancy that accounts for a substantial share of all cancer-related morbidity and mortality. However, little is known with regard to general and disease specific quality of life in survivors of colorectal carcinoma, particularly from community-based samples of cases across stage and survival times from diagnosis. METHODS: Subjects with colorectal carcinoma were recruited from the National Cancer Institute's Surveillance, Epidemiology, and End Results cancer registry. Subjects completed two self administered surveys: the Functional Assessment of Cancer Therapy Scales for Colorectal Cancer (FACT-C) and the Health Utilities Index (HUI) Mark III. RESULTS: One hundred seventy-three respondents (average age: 70.4 years, 71.4% female) completed the survey. In the first 3 years after diagnosis, quality of life was lower and varied substantially among respondents. After 3 years, respondents in all TNM stages of disease except Stage IV reported a relatively uniform and high quality of life. Pain, functional well-being, and social well being were affected most substantially across all stages and times from diagnosis. Low income status was associated with worse outcomes for pain, ambulation, and social and emotional well-being. Only emotional well-being scores improved significantly over time in both surveys. CONCLUSIONS: Those individuals who achieve a long term remission from colorectal carcinoma may experience a relatively high quality of life, although deficits remain for several areas, particularly in those of low socioeconomic status. Sampling design may have excluded the most severely ill patients. PMID- 10717610 TI - Quantification of telomerase activity in sporadic colorectal carcinoma: association with tumor growth and venous invasion. AB - BACKGROUND: Activation of telomerase, a ribonucleoprotein enzyme complex that synthesizes telomere repeats, is associated with acquisition of unlimited cellular proliferation and is commonly detected in human cancer. Measurement of telomerase activity (TA) may provide important information as a diagnostic marker or a prognostic indicator. The authors studied the quantification of TA and assessed its utility as a prognostic marker in sporadic colorectal carcinoma. METHODS: Sixty surgical specimens, including 30 specimens of cancer tissue and 30 specimens of corresponding normal colorectal mucosa, were examined. TA was measured by a fluorescence-based telomeric repeat amplification protocol assay. The authors determined the telomerase index (TI = log (A-B), where A represented TA of cancer tissues and B represented TA of normal mucosa) and examined the relation between TI and clinicopathologic factors using the Student t test, analysis of variance, the Chi-square test, and the Fisher PLSD as a post hoc test. RESULTS: TA of cancer and corresponding normal mucosa was 51.87+/-27.38 and 7.14+/-9.85, respectively (P<0.0001). The cutoff value was determined to be 26 in a receiver operating characteristic study, with 90% sensitivity, 96.7% specificity, and 96.4% positive predictive value. TI was closely correlated with depth of invasion (P = 0.0129) but not with age, gender, histologic type, location, lymph node metastasis, lymphatic infiltration, or Dukes stage. There was a significant difference in TI between tumors with and without venous invasion (P = 0.0003). Four of five tumors with synchronous liver metastasis showed high TI (1.555 20 years ago. The authors reviewed their recent experience with basosquamous carcinoma to identify prognostic factors influencing recurrence. METHODS: The medical records of all patients with the diagnosis of basosquamous carcinoma treated at the University of Louisville-affiliated hospitals between 1985-1988 were reviewed by a senior pathologist. Prognostic factors were analyzed using Cox regression analysis and the log rank test. RESULTS: Thirty-one cases of basosquamous carcinoma were identified in 28 patients. The median age at diagnosis was 68 years (range, 10-94 years). The median follow-up was 60 months (range, 12-312 months). Seventy-five percent of cases were located on the face, neck, and scalp. One patient had regional lymph node metastasis synchronous with the primary tumor. Patterns of recurrence were: local recurrence only (five patients), local recurrence plus regional lymph nodes (three patients), and pulmonary plus regional lymph nodes (one patient). One patient died of pulmonary metastasis. Significant factors predictive of recurrence (P<0.01) were male gender, positive surgical resection margin, lymphatic invasion, and perineural invasion. Although tumor size was not a statistically significant factor overall (P = 0.076), the 3 patients with lymph node metastases had large tumors (measuring 2 cm, 5 cm, and 5 cm, respectively). CONCLUSIONS: Basosquamous carcinoma is an aggressive epithelial neoplasm with a propensity for local recurrence and potential for distant metastatic spread. This behavior differs substantially from basal cell carcinoma. Complete resection with negative surgical margins is essential. Long term follow-up for the detection of local recurrence and distant metastatic spread is recommended. PMID- 10717619 TI - Evaluation of DNA ploidy and degree of DNA abnormality in benign and malignant melanocytic lesions of the skin using video imaging. AB - BACKGROUND: Making a morphologic distinction between benign and malignant melanocytic tumors of the skin is frequently difficult, especially because "gray zones" between these lesions often exist. DNA image cytometry as an adjuvant method for the diagnosis and prognostic prediction of premalignant lesions and malignant tumors of many other organs is already well established. The aim of this study was to determine whether DNA image cytometry is helpful in distinguishing benign from malignant melanocytic lesions and whether cytometry would give valid information with which to predict the prognoses associated with malignant melanomas. METHODS: DNA image cytometry was performed on 127 benign and 58 primary maligant melanomas of the skin as well as 11 metastatic melanomas, using an enzymatic single cell solution according to a method described by Heiden et al. in Cytometry (1991;12:614-21). RESULTS: DNA aneuploidy was graded by DNA index (DI) and a 2c deviation index (2cDI). In contrast to benign melanocytic lesions (with 16% DNA aneuploidy), primary and metastatic malignant melanomas had significantly higher frequencies of DNA aneuploidy (86% and 73%, respectively). In the degree of DNA aneuploidy, significant differences between benign and malignant melanocytic tumors could be observed. The mean 2cDI of aneuploid benign lesions was 1.0, whereas the primary malignant melanomas had a mean 2cDI of 2.92 and the metastatic melanomas a mean of 6.9. The frequency of DNA aneuploidy increased with Breslow thickness. Twenty-one patients with primary malignant melanoma developed metastases. All metastasizing primary tumors were aneuploid and showed a significantly higher grade of DNA aneuploidy than nonmetastasizing malignant melanomas. Moreover, none of the diploid malignant melanomas developed metastases. CONCLUSIONS: This study reveals that DNA image cytometry is prognostically and diagnostically relevant to the evaluation of melanocytic lesions of the skin. Nevertheless, it cannot be relied on alone to provide enough information for a diagnosis. PMID- 10717620 TI - The expression of Fhit protein is related inversely to disease progression in patients with breast carcinoma. AB - BACKGROUND: The FHIT gene, located at human chromosome 3p14.2, frequently is deleted in a number of human tumors, including breast carcinoma. Its protein product (Fhit) is presumed to have tumor suppressor function. Loss of expression of a tumor suppressor gene is an important step in tumor progression from premalignant, to in situ, to invasive carcinoma. METHODS: In the current study, Fhit expression was examined in invasive carcinomas and in epithelial lesions representing stages of carcinoma progression in 50 mastectomy specimens using immunohistochemical methods. RESULTS: Normal ductal and lobular epithelium consistently and strongly expressed Fhit. A complete loss of or a significant reduction in Fhit expression was observed in 72% of breast carcinomas. A statistically significant, negative correlation in Fhit expression among the stages of disease progression in Fhit negative breast carcinomas was observed (normal epithelium > hyperplasia > atypical hyperplasia and carcinoma in situ > invasive carcinoma), whereas no loss of Fhit expression in precursor lesions was observed in Fhit positive tumors. CONCLUSIONS: These observations are consistent with the observed role of FHIT as a tumor suppressor gene in the pathogenesis of specific subsets of carcinomas. PMID- 10717621 TI - Inhibition of growth of MDA-MB-468 estrogen-independent human breast carcinoma by bombesin/gastrin-releasing peptide antagonists RC-3095 and RC-3940-II. AB - BACKGROUND: The growth of breast carcinoma is promoted by autocrine growth factors such as the bombesin (BN)-like peptides and epidermal growth factor (EGF). The stimulatory action of BN-like peptides can be blocked by the use of BN/gastrin-releasing peptide (GRP) antagonists. METHODS: The authors investigated the effects of synthetic BN/GRP antagonists RC-3095 and RC-3940-II on tumor growth and the expression of mRNA for EGF receptors and three BN receptor subtypes in MDA-MB-468 human breast carcinoma. Athymic nude mice with xenografts of MDA-MB-468 human breast carcinoma were injected subcutaneously for 6 weeks with RC-3940-II at doses of 20 or 40 microg/day. In another study, the effects of RC-3940-II and RC-3095 were compared. RESULTS: RC-3940-II caused a significant and dose-dependent growth inhibition of MDA-MB-468 tumors in nude mice; therapy with either dose of RC-3940-II significantly (P<0.01) reduced the mean final tumor volume and weight compared with controls. RC-3940-II induced a persistent regression of > 50% of all tumors. One of 3 tumors treated with 20 microg of RC 3940-II and 3 of 5 tumors treated with 40 microg were found to have regressed completely by the end of the study. When RC-3940-II and RC-3095 were compared at the dose of 20 microg/day, both powerfully suppressed growth of MDA-MB-468 tumors, with RC-3940-II causing a complete regression of 2 tumors and RC-3095 a complete regression of 1 tumor. Receptor analyses of untreated MDA-MB-468 tumors revealed an overexpression of EGF receptors and two classes of binding sites for BN/GRP. mRNAs for receptors of GRP, neuromedin B, and BN receptor subtype-3 were detected by reverse transcriptase-polymerase chain reaction. CONCLUSIONS: A virtual arrest of growth or regression of MDA-MB-468 human breast carcinoma after therapy with RC-3940-II and RC-3095 indicates that these BN/GRP antagonists could provide a new treatment modality for breast tumors expressing BN and EGF receptors. PMID- 10717622 TI - Frequency of BRCA1/BRCA2 mutations in a population-based sample of young breast carcinoma cases. AB - BACKGROUND: There is a clear and growing need for data regarding BRCA1 and BRCA2 mutation frequencies among breast carcinoma cases not specifically ascertained on the basis of extreme family history profiles. Toward this end, the authors previously reported results with regard to BRCA1 in breast carcinoma patients drawn from a population-based study. In the current study the authors present new findings concerning BRCA2 mutation frequency in this same population, as well as summary data regarding the combined contribution of these two genes. METHODS: Subjects were drawn from two population-based, case-control studies of breast carcinoma in young women conducted in western Washington State and focused on 1) women diagnosed with breast carcinoma before age 35 years (n = 203); and 2) women with a first-degree family history of breast carcinoma who were diagnosed before age 45 years (n = 225). Similarities and differences between BRCA2 carriers and BRCA1 carriers were analyzed in terms of age at diagnosis, family history status, and disease features. RESULTS: Of cases diagnosed before age 35 years, all of whom were unselected for family history, 9.4% carried germline mutations (3.4% for BRCA2 and 5.9% for BRCA1). Of cases diagnosed before age 45 years who had a first-degree family history of breast carcinoma, 12.0% carried germline mutations (4.9% for BRCA2 and 7.1% for BRCA1). Increased frequencies of mutations were observed in cases with a personal or family history of early age at diagnosis and in those with four or more family members affected with breast carcinoma. BRCA2 mutations were less common than BRCA1 mutations in families with any history of ovarian carcinoma. CONCLUSIONS: Overall, given current constraints on health care resources, these data suggest that screening for germline mutations in these breast carcinoma susceptibility genes may have the greatest impact on overall health care if it is prioritized toward high and moderate risk populations. PMID- 10717623 TI - Microinvasive breast carcinoma: clinicopathologic analysis of a single institution experience. AB - BACKGROUND: Microinvasive breast carcinoma (MIC) has a good prognosis but specific definitions have varied in the past, making the clinical significance of MIC a subject of debate. METHODS: Microscopic slides of 59 cases of breast carcinoma originally diagnosed as MIC were reviewed retrospectively. Histologic parameters were correlated with clinical findings and outcome to define diagnostic criteria better. RESULTS: On review, the 59 cases were recategorized as follows: pure DCIS (N = 16), DCIS with foci equivocal for microinvasion (N = 7), DCIS with > or =1 focus of microinvasion (N = 11), T1 invasive carcinomas with > or =90% DCIS (N = 18), and T1 tumors with <90% DCIS (N = 7). The MIC cases in the current study averaged 3 separate foci of early infiltration outside the basement membrane, each one not >1.0 mm. The mean follow-up was 95 months. Six patients (10%) had only local recurrence: 1 case each in patients with equivocal microinvasion, microinvasion, and T1 tumors with <90% DCIS and 3 cases among the patients with T1 tumors with > or = 90% DCIS. Four patients, all with T1 tumors with > or =90% DCIS, had distant failure (7%). In the MIC group, only one patient developed a local recurrence after breast conservation. No patient had axillary lymph node metastasis. For the entire series, factors associated with local recurrence were younger age, breast conservation versus mastectomy, and close surgical margins. The only factor associated with distant failure was the size of the DCIS component. Seven patients with T1 tumors with > or =90% DCIS experienced local or distant failure and 5 of these (71%) developed progressive disease or died of disease. All other patients who developed a recurrence were disease free at last follow-up. In a retrospective series, poorer outcome in carcinomas with > or =90% DCIS may be related to the greater likelihood of missed larger areas of invasive carcinoma. Therefore, meticulous and extensive sampling of these carcinomas is required. CONCLUSIONS: MIC as defined has a good prognosis. It has a different biology than T1 invasive carcinoma with > or =90% DCIS, which may progress and cause death. Large tumors with multiple foci of microinvasion may have metastatic potential. PMID- 10717624 TI - Allelic loss at the 8p22 region as a prognostic factor in large and estrogen receptor negative breast carcinomas. AB - BACKGROUND: Allelic losses of tumor suppressor genes, or the chromosomal regions harboring them, in the DNA of tumor cells may become useful postoperative prognostic indicators. The authors studied frequent loss of heterozygosity (LOH) on chromosome 8p22 and its association with disease progression that occurred later in patients with breast carcinoma. METHODS: To examine whether allelic losses at 8p22 might correlate with postoperative survival during a 5-year period of prospective follow-up, the authors tested tumors from a cohort of 298 breast carcinoma patients informative for 8p22 markers. The tumors were tested for allelic losses of microsatellite markers D8S136 and D8S1106 located at 8p22, a chromosomal region where genetic alterations are frequent in breast carcinomas. RESULTS: Among the 298 breast carcinoma patients, 154 (52%) lost alleles in tumors. Patients whose tumors had lost an allele at 8p22 had a significantly higher risk of postoperative mortality than those whose tumors retained both alleles at those loci; their 5-year mortality rates were 18% (26 patients died among 154 with losses at 8p22) versus 7% (10 patients died among 144 with retentions at 8p22) (P = 0.017). The 8p22 LOH was a significant independent prognostic factor for postoperative survival in a group of patients with large tumors (>2.1 cm) and in a group of patients with estrogen receptor negative tumors in both univariate and multivariate analyses. These data show that 8p22 LOH was a significant prognostic factor for the postoperative survival of certain clinical groups of patients who underwent surgery for breast carcinoma. CONCLUSIONS: Allelic loss on chromosome 8p22 can serve as a negative prognostic indicator to guide the postoperative management of patients, especially patients with large tumors (>2.1 cm) and those with estrogen receptor negative tumors. PMID- 10717625 TI - The feasibility of minimally invasive surgery for stage IIA, IIB, and IIIA breast carcinoma patients after tumor downstaging with induction chemotherapy. AB - BACKGROUND: Induction chemotherapy (IC) has become the standard of care for locally advanced breast carcinoma, frequently downstaging both the primary tumor and the axilla, and making patients eligible for less invasive surgical procedures. The usefulness of IC in earlier stage operable breast carcinoma is now being considered. METHODS: This study involved a subset of 129 patients from a series of 174 with T2-3, N0-1, M0 or T1, N1, M0 breast carcinoma (Stage IIA, IIB, or IIIA ) who were registered in a prospective IC trial using paclitaxel or a combination of fluorouracil, doxorubicin, and cyclophosphamide (FAC). The subset included patients who had received no preoperative radiation therapy but had completed 3-5 cycles of induction chemotherapy and had undergone a Level I-II axillary lymph node dissection. The objective was to evaluate the effectiveness of induction chemotherapy with paclitaxel or FAC in downstaging the primary tumor and axillary metastases in these early stage breast carcinoma patients. RESULTS: The median initial tumor size was 4 cm (range, 0.6-10.0); after IC, tumor size was downstaged to 1.6 cm (range, 0.0-7.0) (P < 0.0001). Clinical response to IC was complete in 24% of patients and partial in 36%. Primary tumor shrinkage was similar with paclitaxel and FAC. Among patients clinically classified as N1, 34% became histologically negative and 38% had only 1-3 positive lymph nodes after induction chemotherapy. CONCLUSIONS: IC with paclitaxel or FAC resulted in effective downstaging of primary tumors and axillary metastases in patients with Stage IIA, IIB, and IIIA breast carcinoma. However, a significant proportion of patients still had residual but low volume microscopic disease; such disease status may allow minimally invasive surgical approaches to locoregional therapy. PMID- 10717626 TI - Analysis of clinicopathologic prognostic factors for 157 uterine sarcomas and evaluation of a grading score validated for soft tissue sarcoma. AB - BACKGROUND: Uterine sarcomas (US) are rare and carry a poor prognosis characterized by high rates of local recurrence and metastasis. The aim of this study was to test, for what the authors believe was the first time with US, the prognostic impact of the histologic grade validated by the French Federation of Anticancer Centers (FNCLCC) for soft tissue sarcomas (STS). The grade is the sum of the scores allocated for three major histologic criteria: tumor differentiation, mitotic count, and tumor necrosis. Other histologic and clinical factors were tested as well. METHODS: The study included 157 patients in whom 78 leiomyosarcomas (LMS), 52 malignant mixed mullerian tumors (MMMT), and 27 endometrial stroma sarcomas (ESS) were documented. RESULTS: The median follow-up was 54 months (range, 6-230 months). The median OS and EFS were 33 and 13 months, respectively. The FNCLCC grade validated in soft tissue sarcomas was not a prognostic factor for survival or relapse for any of the US histologic subtypes. For LMS, stage and mitotic count were the only factors that had an influence on survival and relapse. For MMMT, stage and age were the only prognostic factors, and none of the histologic criteria impacted on the outcome. For ESS, the grade defined by Norris and Taylor was an important prognostic factor, particularly for survival. CONCLUSIONS: The FNCLCC grading score could not be used as a prognostic indicator for uterine sarcomas. The diagnosis of US is in itself an unfavorable prognostic factor, except when the diagnosis is low grade ESS. PMID- 10717627 TI - Clinical significance of serum and ascitic p53 autoantibodies in epithelial ovarian carcinoma. AB - BACKGROUND: Accumulation of mutated p53 in malignant cells can lead to the generation of anti-p53 autoantibodies in the serum and other body fluids of cancer patients. This retrospective study was performed to evaluate the prognostic significance of preoperative serum and ascitic anti-p53 antibodies in advanced ovarian carcinoma. METHODS: In 113 ovarian carcinoma patients who presented with significant amounts of ascites, anti-p53 autoantibodies were determined by a highly specific enzyme-linked immunosorbent assay of blood and ascites. Disease free and overall survival of study patients was estimated by the product limit method of Kaplan and Meier. Differences in survival were examined according to criteria of Mantel and Breslow. A multiple regression analysis based on the Cox proportional hazards model was used to determine the independence of prognostic variables. RESULTS: Serum and ascitic anti-p53 antibodies were found in 28 (25%) and 21 (19%) of the study patients, respectively. In univariate analysis, detection of anti-p53 antibodies in ascites but not in serum was found to be a sign of unfavorable disease free survival (P<0.003) and overall survival (P < 0.01). Multivariate analysis revealed that anti-p53 positivity in ascites retained independent significance only in the prediction of adverse progression free survival (P<0.01). CONCLUSIONS: The generation of a humoral immune response against p53 protein in the close tumor environment, as demonstrated by the occurrence of p53 autoantibodies in the ascitic fluid of ovarian carcinoma patients, is associated with poor disease free survival. PMID- 10717628 TI - Oral estramustine and cyclophosphamide in patients with metastatic hormone refractory prostate carcinoma: a phase II study. AB - BACKGROUND: Nearly all cases of metastatic prostate carcinoma progress, after hormonal ablation, to a hormone refractory status. To the authors' knowledge no standard chemotherapy for patients with hormone refractory prostate carcinoma (HRPC) exists. In a prospective study, the efficacy and toxicity of an oral combination of estramustine and cyclophosphamide were evaluated. METHODS: Between March 1996 and April 1998, 32 consecutive patients (median age 74 years; range, 53-84 years) with metastatic HRPC were treated with oral estramustine (10 mg/kg/day) and oral cyclophosphamide (2 mg/kg/day) for 14 days every 28 days. Inclusion criteria were previous complete androgen blockade, antiandrogen withdrawal evaluation, and clinical or biochemical disease progression. Response assessment was based on a decrease > or =50% in the prostate specific antigen (PSA) level associated with improvement (or no worsening) in Eastern Cooperative Oncology Group (ECOG) performance status (PS) and relief of bone pain (if present). RESULTS: All patients were evaluable for efficacy and toxicity. PSA levels decreased by at least 50% in 14 patients (43.7%) (95% confidence interval, 26.5-60.9), remained stable in 12 patients (37.5%), and rose in 6 patients (18.8%). ECOG PS was 0 in 5 of 14 patients, improved from 1 to 0 in 7 patients, and remained unchanged in 2 patients. Bone pain, present in 8 of 14 patients, disappeared in 7 and was partially relieved in 1. The median duration of response was 30 weeks (range, 8-88+ weeks). An objective partial response was obtained in two cases. Toxicity was mild and mainly gastrointestinal (World Health Organization [WHO] Grade 1). No cases of WHO Grade 3-4 hematologic toxicity occurred. CONCLUSIONS: The oral combination of estramustine and cyclophosphamide appears to be safe and effective in patients with HRPC. In responding patients its use shows a clinical benefit in terms of improvement of ECOG PS and pain control. PMID- 10717629 TI - Prostate carcinoma trends in three counties in Sweden 1987-1996: results from a population-based national cancer register. South-East Region Prostate Cancer Group. AB - BACKGROUND: To detect changes in the incidence rate and management of prostate carcinoma, all cases of the disease diagnosed in the southeast region of Sweden between 1987-1996 were recorded. METHODS: The register is based on Swedish personal registration numbers, thereby minimizing the number of dropouts. All cases of prostate carcinoma detected in the southeast region have been recorded according to a defined protocol that has been updated successively to match recent views regarding the disease. To ensure a high number of presented cases, the National Cancer Register was checked for missing cases. RESULTS: Six thousand seven hundred eighty-two cases of prostate carcinoma were registered in the region between 1987-1996. The age-adjusted incidence rate reached a peak in 1993, followed by a slight decrease. The mean age at diagnosis throughout the period was 74.2 years, with a peak age of 74.8 years in 1992. The number of incidental tumors followed the development of the number of transurethral resections of the prostate performed in the region, with a peak in 1991. The percentage of patients receiving gonadotropin-releasing hormone (GnRH) analogues increased from 3.9% to 37.8% whereas the percentage of patients treated with orchiectomy decreased from 40.0% to 12.8% and the percentage of those treated with radical prostatectomy decreased from 11.1% to 2.5%. CONCLUSIONS: A diminishing pool of latent tumors may explain the decreasing incidence rate and lower age at diagnosis observed after 1993. Orchiectomy is rapidly being superseded by GnRH analogues. In contrast to trends reported in the U.S., the percentage of men with prostate carcinoma undergoing total prostatectomy appears to be declining in Sweden. PMID- 10717630 TI - Quality-of-life outcomes for men with prostate carcinoma detected by screening. AB - BACKGROUND: There is limited information on outcomes of prostate carcinoma treatments given to screened patient populations for whom cancer is usually detected at an earlier stage. METHODS: The authors conducted a cross-sectional evaluation of quality-of-life outcomes for men with prostate carcinoma detected in screening studies at a university center. Of 2234 men diagnosed with prostate carcinoma between 1989 and 1997, 74% responded to the questionnaire. Primary management included radical prostatectomy (76%), radiotherapy (11%), observation (7%), hormonal therapy (4%), and cryoablation (2%). Main outcome measures included validated measurements of quality of life, urinary and sexual functioning, and bother (36-item RAND Health Survey, UCLA Prostate Cancer Index). RESULTS: After controlling for demographic factors, differences among treatment groups were found for all general quality-of-life outcomes, with increased impairment in men who underwent hormonal therapy (all P values <0.05). Urinary and sexual function and bother were also significantly related to treatment. However, among men followed for > or =12 months, only 9% reported a moderate or major problem with urinary control. Sexual functioning was a moderate or major problem following treatment for 58% treated with prostatectomy, 48% treated with radiotherapy, 64% treated with hormonal therapy, 45% treated with cryoablation, and 30% managed with observation. Approximately one-third of the men younger than 70 years who underwent radical prostatectomy maintained adequate sexual functioning posttreatment. CONCLUSIONS: Up to 6 years after diagnosis, the majority of men with prostate carcinoma detected by screening were bothered by their current sexual function, regardless of treatment. In contrast, most men were not bothered by their current urinary function. PMID- 10717631 TI - Human papillomavirus-associated carcinomas in Hawaii and the mainland U.S. AB - BACKGROUND: To the authors' knowledge, human papillomavirus (HPV)-associated carcinomas in Hawaii have not been studied in detail. METHODS: Surveillance, Epidemiology, and End Results data (from 1973-1996) were used to study rate of incidence patterns of squamous cell carcinomas (SCCs) of the uterine cervix, vulva/vagina, anus, penis, and palatine tonsils among Asian/Pacific Islanders and whites in Hawaii and among whites in the U.S. in general. RESULTS: With the exception of invasive cervical SCC, male and female Asian/Pacific Islanders in Hawaii had considerably lower incidence rates of HPV-associated SCCs than Hawaii whites and U.S. whites. Among women, Hawaii whites and U.S. whites had rather similar rates of invasive anogenital and tonsillar SCCs, but in situ SCC of the cervix or vulva/vagina was diagnosed less often among Asian/Pacific Islanders and whites in Hawaii than among whites in the general U.S. Among men, Hawaii whites had higher rates than U.S. whites of both anal and tonsillar, but not penile, SCCs. Among Hawaiian men with anal carcinoma, 43% (15 of 35) had remained unmarried versus 3% (2 of 65) of Hawaiian women with anal carcinoma. CONCLUSIONS: Asian/Pacific Islanders in Hawaii generally have lower incidence rates of HPV associated SCCs than whites. However, low ratios of in situ to invasive cervical SCCs suggest that many Hawaii women, notably Asian/Pacific Islanders, are not diagnosed and treated for cervical neoplasias at a preinvasive stage. The high rate of incidence of anal SCC in male Hawaiian whites and the high proportions of unmarried men among patients with this disease suggest the transmission of HPV through homosexual contact. These men may be targeted in future screening programs for anal carcinoma. PMID- 10717632 TI - Inverse association between age at the time of radiation exposure and extent of disease in cases of radiation-induced childhood thyroid carcinoma in Belarus. AB - BACKGROUND: Increased incidence of childhood thyroid carcinoma, particularly in the youngest children, has been reported from Belarus since the nuclear reactor accident at Chernobyl in 1986. The relation between disease severity and age at the time of the accident, not previously established in this cohort, was analyzed in this study. METHODS: The authors studied the association between disease severity, expressed by TNM classification, and age at radiation exposure in a cohort of 483 patients younger than 8 years at the time of the Chernobyl accident who have been diagnosed with differentiated thyroid carcinoma since 1986 at the Center for Thyroid Cancer in Minsk. The associations between age at radiation exposure and TNM categories were compared among 4 groups of patients who were ages <2, 2.1-4, 4.1-6, and 6.1-8 years at the time of the accident. Multivariate discriminant analysis was performed to examine the effects of age at the time of the accident, gender, histology, tumor stage, and N classification on the frequency of distant metastasis. RESULTS: Younger age at the time of the Chernobyl accident was associated with greater extrathyroidal tumor extension (P<0.01) and more lymph node involvement (P<0.0001) and tended to be associated with more distant metastases (P = 0.09). Compared with patients who were ages 6.1 8 years at the time of the accident, patients who were younger than 2 years had significantly more extrathyroidal tumor invasion (P = 0.004), lymph node involvement (P = 0.004), and distant metastases (P = 0.05). The age at diagnosis increased with older age at the time of radiation exposure (linear regression analysis; correlation coefficient = 0. 67; P<0.001). Multivariate analysis revealed that younger age at the time of the accident (P = 0.001) and advanced locoregional tumor extension (P<0.001) were the only powerful factors influencing the risk for distant metastasis of this malignancy. CONCLUSIONS: The severity of disease was associated inversely with age at the time of radiation exposure in these cases of radiation-induced childhood thyroid carcinoma. PMID- 10717633 TI - Limitations of the World Health Organization classification of childhood supratentorial astrocytic tumors. Children Brain Tumor Consortium. AB - BACKGROUND: In the context of many implied but not rigorously stated histologic feature combinations, the World Health Organization (WHO) classification of astrocytic tumors specifies only the presence or absence of endothelial proliferation, necrosis, and mitosis to distinguish astrocytoma, anaplastic astrocytoma, and glioblastoma multiforme. METHODS: The authors examined the effects of these and other reliably recognized histologic features on survival in the Childhood Brain Tumor Consortium (CBTC) sample of 340 children with supratentorial astrocytic tumors. RESULTS: Overall, the WHO criteria distinguished only two prognostically distinct classes of astrocytomas. When the specific combinations of the three features were unambiguously designated, three diagnostic categories resulted. These revised diagnostic categories are consistent with WHO guidelines and have significantly different survival distributions. However, neither the original WHO diagnoses nor the revised categories adequately separated these tumors prognostically, because histologic features other than those specified by WHO were significantly associated with improved or worsened survival. CONCLUSIONS: Classifications based on small numbers of specified histologic features may not be feasible because they inadequately separate childhood astrocytic tumors into prognostically homogeneous groups. Preferable classification techniques are those that simultaneously account for all reliably recognized histologic features. PMID- 10717634 TI - A new American Joint Committee on Cancer staging system for cutaneous melanoma. AB - The Melanoma Staging Committee of the AJCC has proposed major revisions of the melanoma TNM and stage grouping criteria. The committee members represent most of the major cooperative groups and cancer centers worldwide with a special interest in melanoma; the committee also collectively has had clinical experience with over 40,000 patients. The new staging system better reflects independent prognostic factors that are used in clinical trials and in reporting the outcomes of various melanoma treatment modalities. Major revisions include 1) melanoma thickness and ulceration, but not level of invasion, to be used in the T classification; 2) the number of metastatic lymph nodes, rather than their gross dimensions, the delineation of microscopic versus macroscopic lymph node metastases, and presence of ulceration of the primary melanoma to be used in the N classification; 3) the site of distant metastases and the presence of elevated serum LDH, to be used in the M classification; 4) an upstaging of all patients with Stage I,II, and III disease when a primary melanoma is ulcerated; 5) a merging of satellite metastases around a primary melanoma and in-transit metastases into a single staging entity that is grouped into Stage III disease; and 6) a new convention for defining clinical and pathologic staging so as to take into account the new staging information gained from intraoperative lymphatic mapping and sentinel lymph node biopsy. The AJC Melanoma Staging Committee invites comments and suggestions regarding this proposed staging system before a final recommendation is made. PMID- 10717636 TI - Author reply PMID- 10717635 TI - Increased incidence rates but no space-time clustering of childhood astrocytoma in Sweden, 1973-1992: a population-based study of pediatric brain tumors. PMID- 10717638 TI - Dendritic and axonal targeting of the vesicular acetylcholine transporter to membranous cytoplasmic organelles in laterodorsal and pedunculopontine tegmental nuclei. AB - Autoregulation of cholinergic neurons in the laterodorsal tegmental (LDT) and pedunculopontine (PPT) nuclei has been implicated in many functions, most importantly in drug reinforcement and in the pathophysiology of schizophrenia. This autoregulation is attributed to the release of acetylcholine, but neither the storage or release sites are known. To determine these sites, we used electron microscopy for the immunocytochemical localization of antipeptide antiserum raised against the vesicular acetylcholine transporter (VAchT) that is responsible for the uptake of acetylcholine into storage vesicles. The cellular and subcellular distribution of VAchT was remarkably similar in the two regions by by using each of two methods, immunogold and immunoperoxidase. In both PPT and LDT nuclei, VAchT labeling was seen mainly on membranous organelles including the trans-Golgi network in many somata. VAchT-immunoreactive tubulovesicles resembling saccules of smooth endoplasmic reticulum were often seen near the plasma membrane in dendrites. The VAchT-containing dendrites comprised almost 50% of the labeled profiles (1027/2129) in PPT and LDT nuclei. The remaining VAchT immunoreactive profiles were primarily small unmyelinated axons and axon terminals. In axon terminals, VAchT was densely localized to membranes of small synaptic vesicles. The VAchT-immunoreactive axon terminals formed either symmetric or asymmetric synapses. The postsynaptic targets of these axon terminals included dendrites that were with (36/110) or without (74/110) VAchT immunoreactivity. Our results suggest that dendrites, as well as axon terminals, have the potential for storage and release of acetylcholine in the LDT and PPT nuclei. The released acetylcholine is likely to play a major role in autoregulation of mesopontine cholinergic neurons. PMID- 10717637 TI - Molecular mapping of brain areas involved in parrot vocal communication. AB - Auditory and vocal regulation of gene expression occurs in separate discrete regions of the songbird brain. Here we demonstrate that regulated gene expression also occurs during vocal communication in a parrot, belonging to an order whose ability to learn vocalizations is thought to have evolved independently of songbirds. Adult male budgerigars (Melopsittacus undulatus) were stimulated to vocalize with playbacks of conspecific vocalizations (warbles), and their brains were analyzed for expression of the transcriptional regulator ZENK. The results showed that there was distinct separation of brain areas that had hearing- or vocalizing-induced ZENK expression. Hearing warbles resulted in ZENK induction in large parts of the caudal medial forebrain and in 1 midbrain region, with a pattern highly reminiscent of that observed in songbirds. Vocalizing resulted in ZENK induction in nine brain structures, seven restricted to the lateral and anterior telencephalon, one in the thalamus, and one in the midbrain, with a pattern partially reminiscent of that observed in songbirds. Five of the telencephalic structures had been previously described as part of the budgerigar vocal control pathway. However, functional boundaries defined by the gene expression patterns for some of these structures were much larger and different in shape than previously reported anatomical boundaries. Our results provide the first functional demonstration of brain areas involved in vocalizing and auditory processing of conspecific sounds in budgerigars. They also indicate that, whether or not vocal learning evolved independently, some of the gene regulatory mechanisms that accompany learned vocal communication are similar in songbirds and parrots. PMID- 10717639 TI - Serotonergic systems in the spinal cord of the amphibian urodele Pleurodeles waltl. AB - The role of the monoamine serotonin (5-HT) in modulating the neural networks underlying axial locomotor movements was studied in an adult amphibian urodele, Pleurodeles waltl. 5-HT was applied to an in vitro brainstem-spinal cord preparation of P. waltl, which displayed fictive axial locomotor patterns following bath application of N-methyl-D-aspartate (5 microM) with D-serine (10 microM). Our results showed that 5-HT (1-25 microM) produces a reversible increase in the cycle duration and the duration of rhythmic bursting activity recorded extracellularly from ventral roots innervating the axial musculature. When applied alone, 5-HT does not trigger axial locomotor activity. The distribution pattern of 5-HT immunoreactive (5-HT-ir) cells along the spinal cord was investigated both in intact and in chronic spinal animals. The number of 5-HT ir cell bodies is higher at brachial levels and decreases through crural levels. Sparse oval or fusiform 5-HT-ir somata are present within the gray matter, just ventrolateral to the central canal. Longitudinal fibers were detected throughout the entire white matter, except in the medial part of the dorsal funiculi. Two columns of intensely labeled and profusely branching thick and thin fibers associated with numerous varicosities run continuously along the ventrolateral surface of the spinal cord. Three weeks following full spinal cord transection at the level of the second spinal root, all longitudinal processes had disappeared, indicating their supraspinal origin, whereas the ventrolateral plexes remained, suggesting that they originated from intraspinal 5-HT-ir cell bodies. Our data showing that spinal 5-HT is organized according to a rostrocaudal gradient suggest that the 5-HT systems of P. waltl are not related to the presence of limb motor pools but more likely are related to axial central pattern generators (CPGs) networks down the length of the spinal cord. The possible involvement of these two sources (descending vs. intraspinal) of 5-HT innervation in the modulation of the axial CPGs is discussed. PMID- 10717640 TI - Neurochemical and connectional organization of the dorsal pulvinar complex in monkeys. AB - To investigate the organization of the dorsal pulvinar complex, patterns of neurochemical staining were correlated with cortico-pulvinar connections in macaques (Macaca mulatta). Three major neurochemical subdivisions of the dorsal pulvinar were identified by acetylcholinesterase (AChE) histochemistry, as well as immunostaining for calbindin-D(28K) and parvalbumin. The dorsal lateral pulvinar nucleus (PLd) was defined on histochemical criteria as a distinct AChE- and parvalbumin-dense, calbindin-poor wedge that was found to continue caudally along the dorsolateral edge of the pulvinar to within 1 mm of its caudal pole. The ventromedial border of neurochemical PLd with the rest of the dorsal pulvinar, termed the medial pulvinar (PM), was sharply defined. Overall, PM was lighter than PLd for AChE and parvalbumin and displayed lateral (PMl) and medial (PMm) histochemical divisions. PMm contained a central "oval" (PMm-c) that stained darker for AChE and parvalbumin than the surrounding region. The neurochemically defined PLd was labeled by tracer injections in the inferior parietal lobule (IPL) and dorsolateral prefrontal cortex but not the superior temporal gyrus (STG). Label within PMl was found after prefrontal and IPL and, to a lesser extent, after STG injections. The PMm was labeled after injections of the IPL and STG, but only sparsely following prefrontal injections. The histochemically distinct subregion or module of PMm, PMm-c, was labeled only by STG injections. Overlapping labeling was found in dorsal pulvinar divisions PMl and PLd following paired IPL/prefrontal, but not IPL/STG or these particular STG/prefrontal, injections. Thus, PLd may be a visuospatially related region whereas PM appears to contain several types of territories, some related to visual or auditory inputs, and others that receive directly converging input from posterior parietal and prefrontal cortex and may participate in a distributed cortical network concerned with visuospatial functions. PMID- 10717641 TI - Topographic organization of inferior olive cells projecting to translational zones in the vestibulocerebellum of pigeons. AB - In the nodulus and ventral uvula of pigeons, there are four parasagittal zones containing Purkinje cells responsive to patterns of optic flow that results from self-translation along a particular axis in three-dimensional space. By using a three-axis system to describe the preferred direction of translational optic flow, where +X, +Y, and +Z represent rightward, upward, and forward self-motion, respectively, the four cell types are: +Y, -Y, -X-Z, and -X+Z (assuming recording from the left side of the head). The -X-Z zone is the most medial, followed in sequence by the -X+Z, -Y zone, and the +Y zones. In this study, we injected the retrograde tracer cholera toxin subunit B into each of the four translational zones to determine the origin of the climbing fiber inputs in the inferior olive. Retrograde labeling in the inferior olive was found in the ventrolateral margin of the medial column from injections into all four translational zones; however, there was a clear functional topography. Retrograde labeling from -Y zone injections was found most rostrally in the medial column, whereas retrogradely labeled cells from -X-Z zone injections were found most caudally in the medial column. Labeling from +Y and -X+Z zone injections were found between the labeling from -Y zones and -X-Z zones, with +Y labeling located slightly caudal to -X+Z labeling. PMID- 10717642 TI - Patterns of calretinin, calbindin, and tyrosine-hydroxylase expression are consistent with the prosomeric map of the frog diencephalon. AB - This paper re-examines a previously published segmental map of the frog diencephalon (Puelles et al. [1996] Brain Behav.Evol. 47:279-310) by means of immunocytochemical mapping of calretinin, calbindin, and tyrosine hydroxylase. The distribution of neuronal populations, axon tracts, and neuropils immunoreactive for these markers was studied in adult specimens of Rana perezi and Xenopus laevis sectioned sagittally or horizontally. Emphasis was placed on study of the relationship of observed chemoarchitectural boundaries with the postulated overall prosomeric organization and the schema of nuclear subdivisions we reported previously, based on acetylcholinesterase histochemistry and Nissl pattern in Rana. The data reveal a large-scale correspondence with the segmental map in both species, although some differences were noted between Rana and Xenopus. Notably, retinorecipient neuropils were generally immunoreactive for calretinin only in Rana. Importantly, calretinin immunostaining underlines particularly well the transverse prosomeric boundaries of the dorsal thalamus. A number of nuclear subdivisions noted before with AChE were corroborated, and some novel subdivisions became apparent, particularly in the anterior nucleus of the dorsal thalamus and in the habenular complex. The mapping of tyrosine hydroxylase clarified the segmental distribution of the catecholaminergic cell groups in the frog forebrain, which is comparable to that observed in other vertebrates. PMID- 10717644 TI - Corrigendum AB - What's in a Citation Impact Factor? A Journal by Any Other Name. J. Comp. Neurol. 411:1-2 (1999). PMID- 10717643 TI - Neurones in the adult rat anterior medullary velum. AB - The presence of neurones in the rat anterior medullary velum (AMV) has been investigated by using antibodies to the calcium-binding proteins, parvalbumin (PV), calretinin (CR), and calbindin-D28k (CB). Disparate populations of mainly GABAergic neurones were located in the rostral and caudal regions of the AMV. The rostral region of the AMV was characterised by GABAergic CR-labelled or PV labelled neurones. CR-labelled neurones were bipolar or multipolar with round to ovoid somata (diameters between 8 and 12 microm), and rostrocaudally running dendrites forming a network. PV-labelled neurones had round somata (diameters between 6 and 10 microm) and were bi-tufted, with beaded dendrites. Both CR labelled and PV-labelled dendrites formed punctate pericellular associations with unlabelled somatic profiles. In the caudal region of the AMV, PV-labelled neurones were GABAergic, multipolar cells, having round somata (diameters between 9 and 12 microm), with either beaded or nonbeaded dendrites forming a network of interconnecting dendrites. PV-labelled pericellular associations were made around both PV-labelled and unlabelled somatic profiles. CR labelled unipolar brush cells (UBCs) were not GABAergic. UBCs were characterised by a round to oval somata (10-15 microm in diameter) from which a single primary dendrite emerged to form a distal expansion having small terminal dendrites. From the distal expansion, there also appeared to be CR-labelled processes emanating and extending for up to 250 microm. CB occasionally labelled "Purkinje-like cells" (PLCs). The rat AMV is a more complex structure than first envisaged with the presence of predominantly inhibitory neurones expressing different calcium binding proteins. Functional and anatomic aspects of this circuitry are further discussed. PMID- 10717645 TI - Simple and effective gas chromatographic mass spectrometric procedure for the speciation analysis of organotin compounds in specimens of marine mussels. An evaluation of the organotin pollution of the lagoon of venice. AB - This paper describes a simple, effective analytical procedure, based on a gas chromatographic mass spectrometric technique, for the speciation analysis of organotin compounds (OTC) in mussel samples. The direct alkylation reaction of the organotin chlorides in the aqueous digestion solution by NaBEt(4) allowed a short analysis time and a good recovery. The evaluation of the yield of each step constituting the analytical procedure indicated that the alkylation step is the most critical one. The proposed method was advantageously utilised to monitor the level of OTC pollution of the Lagoon of Venice. All the sites examined, both near to and far from anthropogenic activities, revealed significant levels of OTC pollution. PMID- 10717646 TI - Determination of novel aromatic amines in environmental samples by gas chromatography/mass spectrometry. AB - A number of compounds belonging to five chemical classes of aromatic amines were identified by electron impact mass spectrometry, both in the vapor phase and in suspended particulate matter collected in an area subjected to heavy traffic. This paper presents the mass spectra of new aromatic amines, proposed mechanisms for formation of the major ions obtained by electron impact, concentrations of each class in 13 environmental samples, and the distribution of the individual compounds of the major chemical classes (greater than 80% by weight). PMID- 10717647 TI - Analysis of oxosteroids by nano-electrospray mass spectrometry of their oximes. AB - A method for the analysis of neutral oxosteroids by electrospray mass spectrometry is described. The oxosteroids are converted into their oximes by treatment with hydroxyammonium chloride in aqueous methanol. Intense peaks corresponding to protonated oxime molecules are observed in nano-electrospray mass spectra. The detection limits for the oximes of progesterone, pregnenolone and dehydroepiandrosterone were 2.5, 5 and 25 pg/microL, respectively, approximately 20 times lower than for the underivatised steroids. The signal intensities were proportional to the concentration of the steroids in the range of 500 to 2.5 pg/microL. Fragmentation by collision-induced dissociation (CID) was studied using oximes of 28 model steroids carrying an oxo group at C-3, C-17 or C-20. Some of the steroid oximes were labelled with deuterium or (15)N. Fragment ions were observed which yielded useful structural information. Upon CID, protonated oximes of 3-oxo-Delta(4)-steroids produced abundant ions by cleavage through the B-ring and by loss of the side chain, while protonated oximes of saturated 3-oxosteroids did not give abundant ions by cleavage through the B-ring. Protonated oximes of 20-oxosteroids unsubstituted at C-21, C-17 or C 16 produced a characteristic ion at m/z 86 containing the side chain, C-16 and C 17. Protonated oximes of steroids containing only a 17-oxo group gave fewer ions of diagnostic value. Coupled with the selective isolation of steroid oximes from a biological matrix this method of derivatisation and CID may be used for the analysis of neutral oxosteroids in biological samples. PMID- 10717648 TI - Trace analysis of estrogenic chemicals in sewage effluent using liquid chromatography combined with tandem mass spectrometry. AB - A rapid, selective, sensitive, and reproducible method for the analysis of environmental estrogens, either natural or synthetic, present in samples from sewage treatment works (STW) has been developed. Isolation of these compounds from STWs was performed by applying a simple extraction procedure using an ENVI CARB cartridge (a graphitized non porous carbon black) as the solid-phase extraction (SPE) system. Liquid chromatography coupled with atmospheric pressure chemical ionization (APCI) and tandem mass spectrometry was used for detection. For the multiple reaction monitoring (MRM) mode, the [M + H](+) ion of estrone and the [M + H-H(2)O](+) ions of 17beta-estradiol, estriol and 17alpha ethynylestradiol were selected as the precursors for collisionally induced dissociation (CID). The average recoveries from sewage final effluent samples ranged from 84 to 93% for low levels, and from 89 to 95% for high levels. The precision of the method ranged from 11 to 8% for low level and from 9 to 7% for high level samples. The lower level of quantitation for these estrogens in STW samples was determined at 0.5 ng/L for 17beta-estradiol and 17alpha ethynylestradiol, and 1 ng/L for estrone and estriol, based on 1-L aliquots of sewage treatment works water, using the optimum tuning parameters for each individual selected precursor ion/product ion transition. Compared to a previous gas chromatography/mass spectrometry (GC/MS) method for the analysis of ten STW samples, this method was shown to provide higher sensitivity and lower time consumption. PMID- 10717649 TI - Dimethyl ether chemical ionization of arylalkylamines. AB - The gas-phase reactions of dimethyl ether (DME) ions with a number of biologically active arylalkylamines of the general formula R(1)R(2)C(6)H(3)CHR(3)(CH(2))(n)NR(4)R(5), where R(1) = H or OH, R(2) = H, F, NO(2), OH or OCH(3), R(3) = H or OH, R(4) and R(5) = H or CH(3), have been studied by means of chemical ionization mass spectrometry. Under the experimental conditions used, the most abundant DME ion is the methoxymethyl cation (CH(3)OCH(2)(+), m/z 45). The unimolecular metastable decompositions of the [M + 45](+), [M + 13](+) and [M + 15](+) adducts formed have been interpreted in terms of the initial site of reaction with the amines and the presence of different functional groups in the molecule. This has permitted establishment of general fragmentation patterns for the adducts, and their correlation with structural features of the molecules. The main site of reaction of the ion CH(3)OCH(2)(+) with the amines seems to be the amino group, particularly if the amine is primary, although a competition with attack on the aromatic ring and especially on the benzylic hydroxy group is observed. In a few cases the reaction mechanisms have been elucidated through the use of deuterated amines obtained by H/D exchange with D(2)O. PMID- 10717650 TI - Proton mobility in protonated peptides: a joint molecular orbital and RRKM study. AB - The mobile proton model was critically evaluated by using purely theoretical models which include quantum mechanical calculations to determine stationary points on the potential energy surface (PES) of a model compound, and Rice Ramsperger-Kassel-Marcus (RRKM) calculations to determine the rate constants of various processes (conformational changes, proton transfer reactions) which occur during mass analysis of protonated peptides. Extensive mapping of the PES of protonated N-formylglycinamide resulted in various minima which were stabilized by one or more of the following types of interaction: internal hydrogen bond, charge transfer interaction, charge delocalization, and ring formation. The relative energies of most of the investigated minima are less then 20 kcal mol( 1) compared with the most stable species. More importantly, the relative energies of the transition structures connecting these minima are fairly low, allowing facile transitions among the energetically low-lying species. It is demonstrated that a path can be found leading from the energetically most stable species, protonated on an amide oxygen, to the structure from which the energetically most favorable fragmentation occurs. It is also shown that the added proton can sample all protonation sites prior to fragmentation. The RRKM calculations applied the results of ab initio computations (structures, energetics, vibrational frequencies) to the reactions (internal rotations, proton transfers) occurring in protonated N-formylglycinamide, and clearly lend additional evidence to the mobile proton model. Based on the results of the PES search on protonated N formylglycinamide, we also comment on the mechanism proposed by Arnot et al. (Arnot D, Kottmeier D, Yates N, Shabanowitz J, Hunt D F. 42(nd) ASMS Conference on Mass Spectrometry, 1994; 470) and Reid et al. (Reid G E, Simpson R J, O'Hair R A J. J. Am. Soc. Mass Spectrom. 1998; 9:945) for the formation of b(2)(+) ions. According to the high level ab initio results, the mechanism relying on amide oxygen protonated species seems to be less feasible than the one which involves N protonated species. PMID- 10717651 TI - Quasi-linear gradients for capillary liquid chromatography with mass and tandem mass spectrometry. AB - Gradient elution, capillary liquid chromatography mass spectrometry was performed with linear, static gradients constructed by laminar flowing ten, 1.5 microL volume steps of decreasing organic concentration into tubing of small internal diameter. Sample loading, gradient formation, and sample elution were accomplished entirely by means of a commercially available micro-autosampler and single-syringe drive pump. The procedure was simple, fast, stable, and reproducible. Essentially linear gradients were produced without the use of additional valves, mixers, pumps or software. It took less than 10 minutes to form a gradient and less than 30 minutes to construct the set of individual buffer vials. The gradients were shown to be stable to storage. One hour after forming, peak retention times were reproduced to +/-0.5%. Long-term retention time reproducibility was found to vary by +/-2%. Chromatographic resolution was comparable or superior to that obtained by gradient elution with conventional dynamic mixing and split flow. The procedure was adapted with a 'peak parking' method which extended the time for generating peptide fragmentation data up to 10 minutes per peptide with the triple quadruple mass spectrometer. Using this technique, collision data were collected at the 25 femtomole level on nine of ten tryptic peptides in a single run. PMID- 10717652 TI - Matrix effects on selectivities of poly(ethylene glycol)s for alkali metal ion complexation in matrix-assisted laser desorption/ionization. AB - The effect of the matrix on poly(ethylene glycol) (PEG) cationization was investigated using an equimolar mixture of CsCl and LiCl as cationizing agents. It was observed that for both carboxylic acid and non-carboxylic acid matrices used, PEG cationization by a given alkali metal ion depends on the matrix used. For example, cesiated PEG ion intensities were much stronger than those of the corresponding lithiated PEG ions when equimolar concentrations of Cs(+) and Li(+) were present and IAA (indolacrylic acid) was the matrix. However, an opposite result was obtained when the same experimental conditions were utilized, but with HABA (2-(4-hydroxyphenylazo)benzoic acid) in place of IAA as the matrix. PMID- 10717653 TI - Mass spectrometric study of tricarbonyl (eta(6)-phenyl methanols) of chromium(0). AB - A mass spectrometric study of several tricarbonyl (eta(6)-phenyl methanols) of chromium(0) was performed. Electron ionization (EI), chemical ionization (CI) and fast atom bombardment (FAB) mass spectra were acquired for each molecule, and compared in order to establish a general fragmentation pattern. The suggested pathways were investigated and confirmed by means of constant b/e linked scans and high resolution data. In addition a Hammett-McLafferty correlation for some peaks derived from the molecular ions was accomplished. PMID- 10717654 TI - Preparation of hydrogen from water by reduction with lithium aluminium hydride for the analysis of delta(2)H by isotope ratio mass spectrometry. AB - An off-line technique is described for the preparation of H(2) from water prior to analysis of delta(2)H by dual-inlet isotope ratio mass spectrometry. H(2) is produced from sample water by reaction with LiAlH(4). This provides a rapid and inexpensive method for the analysis of delta(2)H in small (10 microL) samples of water. Precision was +/- 4.2 to 8.0 (1sigma(n), n = 8) delta(2)H(VSMOW) for samples between 428 and 1500 delta(2)H(VSMOW), +/- 14.5 delta(2)H(VSMOW) for water enriched to 3750 delta(2)H(VSMOW) and +/- 26.0 delta(2)H(VSMOW) for water enriched to 6100 delta(2)H(VSMOW). Accuracy was +/- 1.1 to 4.2 delta(2)H(VSMOW) for water standards from natural abundance to 1000 delta(2)H(VSMOW) (the highest enrichment at which water of accepted delta(2)H is currently available). This method for delta(2)H determination is most appropriate for use with small (<50 microL) samples of high delta(2)H enrichment such as those produced from doubly labelled water studies of small animals. The levels of measurement precision of delta(2)H would contribute 2.6-3.8% to the precision error in estimates of small animal energy expenditure made using the doubly labelled water technique when duplicate analyses are performed. PMID- 10717655 TI - Three-dimensional monopole - a new ion trap mass analyser with one-dimensional ion sorting. AB - This article describes an axially symmetric ion trap with one-dimensional ion sorting ('three-dimensional (3D) monopole'). The mass spectrometer can be created by substituting one hyperboloid endcap electrode with a cone-shaped electrode, the vertex of the cone coincides with the cross-over point of the asymptotes of the ion trap electrodes. The potential applied to this cone-shaped electrode is equal to the potential at the centre of the electrode system. The stability zone of the 3D monopole has a shape of a narrow band extending along the z boundary; the mass resolution and the quality factor (the product of resolution and sensitivity) are constant within this stability band which also lowers the requirements for the driving voltage stability. The mass resolution obtained 1s 100 measured at 50% peak height and the relative sensitivity is 3 x 10(-5). PMID- 10717656 TI - Mass spectrometric determination of appearance energies for ions formed from CoF(4) and CoF(3) molecules. AB - Knudsen cell mass spectrometry was applied to the evaluation of the ionization efficiency curves for the ions originating from CoF(4) molecules. Cobalt tetrafluoride was obtained in the gas phase over the CoF(3)(s)-TbF(4)(s) system in the temperature range from 640 to 690 K. From the ionization efficiency curves the appearance energies of the ions formed from the CoF(4) molecules were determined by means of Vogt's deconvolution method. Clausius-Clapeyron plots for the ions from CoF(4) molecules were measured. Evaporation of pure CoF(3)(s) was carried out, and the appearance energies of the ions formed from CoF(3) molecules were determined. The ionization energies for CoF(4) and CoF(3) molecules were found to be (14.3 +/- 0.2) and (13.3 +/- 0.1) eV, respectively. PMID- 10717657 TI - Gas phase thermal denaturation of an oligonucleotide duplex and its complexes with minor groove binders. AB - Electrospray ionization with in-source collisionally induced dissociation has been used to probe the gas phase stability of an oligonucleotide duplex and its complexes with some minor groove binding drugs. On the basis of the arguments developed in detail by Drahos et al. (J. Mass Spectrom. 1999; 34:1373), this type of experiment can also be described as 'thermal denaturation in the gas phase'. We found that the gas phase denaturation curves were very similar to the solution phase denaturation curves determined by the traditional UV spectrophotometric method and, by analogy with the melting temperature T(m) which characterizes the stability in solution, we define a melting voltage V(m) to characterize the stability in the gas phase. A comparison of the T(m) and V(m) relative values suggests that the structure of the complexes is conserved during the electrospray process which transfers the ions from the solution to the gas phase. PMID- 10717658 TI - Analysis of a commercial preparation of erythromycin estolates by tandem mass spectrometry and high performance liquid chromatography/electrospray ionization tandem mass spectrometry using an ion trap mass spectrometer. AB - Because of the lack of a UV chromophore and their much smaller abundances in comparison with the major component, the minor components in erythromycin estolate preparations are difficult to analyze by high performance liquid chromatography ultraviolet (HPLC-UV). Tentative assignment of the major and minor components can be achieved with the combination of full scan and ZoomScan using an ion trap mass spectrometer. Tandem mass spectrometry (MS/MS) provided an effective method to quickly identify most components without chromatographic separation, and all the related compounds, except the isobaric pair ECE and PdMeEA, could be identified in this way. The best result was obtained by using liquid chromatography/tandem mass spectrometry (LC/MS/MS) operated in selected reaction monitoring mode. The major compound, the estolate of erythromycin A (EAE), and seven other minor components, could be separated and identified, with semiquantitative estimates of relative concentrations. PMID- 10717659 TI - Trace-level quantitation of iralukast in human plasma by microbore liquid chromatography/tandem mass spectrometry. AB - Iralukast (CGP 45715A) is a potent peptido-leukotriene antagonist that is active in various in vitro and animal models for the treatment of asthma. An analytical challenge was to develop a sensitive liquid chromatography/tandem mass spectrometry (LC/MS/MS) method with a lower limit of quantitation (LLOQ) of 10 pg/mL for the analysis of iralukast when administered at low doses during clinical trials. Several issues had to be addressed in order to devise a LC/MS/MS assay for the above compound. First, iralukast appeared to be light sensitive and unstable at room temperature under acidic conditions. Second, a LLOQ of 10 pg/mL was needed to support several clinical trials. Third, positive electrospray ionization of iralukast did not yield the necessary sensitivity required for studies in humans. Consequently, LC/MS/MS conditions were optimized for the negative ion mode of detection. Fourth, sample preparation steps proved to be critical to reduce the possibility of microbore HPLC column (50 mm x 1.0 mm i.d.) obstruction, chromatographic deterioration, and matrix-mediated electrospray ion suppression. While our validated method addressed the above challenges, its major drawback was limited sample throughput capability. Nonetheless, plasma concentration-time profiles for patients with moderate asthma after oral administration of 200, 500, 1000, and 5000 microgram/kg/day of iralukast were successfully obtained. PMID- 10717660 TI - Qualitative and quantitative end-group analysis of a small molecular weight polyester by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. AB - Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry was used for qualitative and quantitative end-group analysis of a small molecular weight polyester, poly(2-butyl-2-ethyl-1,3-propylene phthalate). The presence of carboxyl-terminated linear and cyclic polyester oligomers was confirmed with the help of simple sample preparation methods. The presence of carboxyl end-groups in the polyester chains was verified through their formation of carboxylate salts with alkali metal cations. Cyclic oligomers were identified through deuterium exchange of the exchangeable protons of the polyester. Various inorganic salts were tested for salt formation of the carboxyl end-groups, but only the alkali metal salts proved effective. The influence of the alkali metal salts on the results of the quantitative end-group analysis was also studied. The relative amounts of differently terminated and cyclic oligomers were calculated when the alkali metal salts were used with different matrices. The results showed that both the salts and the matrices used in sample preparation can have a marked effect on the quantitative results of the end-group analysis. The measurements were carried out using 2,5-dihydroxybenzoic acid (DHB), 1,8, 9 trihydroxyanthracene (dithranol), and 2-(4-hydroxyphenylazo)benzoic acid (HABA) as matrix compounds. Dithranol and HABA repeatably exhibited similar results, and these results differed from those obtained with DHB probably because of the different ionization mechanisms in the MALDI process. PMID- 10717661 TI - Analysis of missed cleavage sites, tryptophan oxidation and N-terminal pyroglutamylation after in-gel tryptic digestion. AB - Peptide mass fingerprinting is a powerful tool for the identification of proteins. Trypsin is the most widely used enzyme for this purpose. Therefore, 104 protein digests from human Jurkat T cells and Mycobacterium were analyzed considering missed cleavage sites, tryptophan oxidation and N-terminal pyroglutamylation. About 90% of the matched peptides with missed cleavage sites could be classified into three groups: (i) lysine and arginine with a neighbouring proline on the carboxy-terminal side, (ii) neighboring lysines/arginines, and (iii) lysines and arginines with an aspartic acid or glutamic acid residue on either the amino- or carboxy-terminal side. The first group is already accounted for by search programs. The number of missed cleavage sites can be increased without reducing the precision of the database search by taking the other two groups into consideration. Peptides with tryptophan were observed in non, singly (+16 Da) and doubly (+32 Da) oxidized forms. The higher oxidized form was only observed with lower intensity in the presence of the lower oxidized form. Peptides with N-terminal glutamine were found always as pyroglutamate (-17 Da), and in the majority of cases in pairs with unmodified glutamine. These data can be used for the refinement of protein searches by peptide mass fingerprinting. PMID- 10717662 TI - Characterization of dansylated glutathione, glutathione disulfide, cysteine and cystine by narrow bore liquid chromatography/electrospray ionization mass spectrometry. AB - A method using reversed phase high performance liquid chromatography/electrospray ionization-mass spectrometry (RP-LC/ESI-MS) has been developed to confirm the identity of dansylated derivatives of cysteine (C) and glutathione (GSH), and their respective dimers, cystine (CSSC) and glutathione disulfide (GSSG). Cysteine, GSH, CSSC and GSSG are present at low concentrations in rainbow trout (Oncorhynchus mykiss) liver cells. Initially, hepatic cells were sampled from a suspension culture and disrupted upon addition of 10% perchloric acid. The reduced thiols present in the cell extracts were acetylated to prevent dimerization and then the C and GSH species were derivatized with dansyl chloride for fluorescence detection. An LC system using a weak anion exchange column (AE) with fluorescence detection (FLD) was used for sensitive routine analysis; however, it produced peaks of unknown origin in addition to the expected analytes. Analytes were then separated on a C18 RP-LC system using a water/acetonitrile gradient with 0.2% formic acid, and detected using LC/ESI-MS at 3.5 KV which produced an intense ion with a minimum limit of detection of less than 0.5 pmole injected (>10:1 signal-to-noise (S/N). Subsequently, fractions of effluent from the AE-LC/FLD system were analyzed by LC/ESI-MS to confirm the presence of the target analytes in routine cell extracts. Monodansylated GSSG was identified as a product that could possibly affect the quantification of GSH and GSSG. PMID- 10717663 TI - Identification of ethyl pyruvate and dihydrocinchonidine adducts by electrospray ionization mass spectrometry. AB - Several ethyl pyruvate and dihydrocinchonidine adducts, formed by non-covalent interactions with alkali cations, have been identified for the first time using electrospray ionization mass and tandem mass spectrometry. This type of adduct may have an important role in asymmetric reactions of pyruvates in the presence of cinchonas. PMID- 10717665 TI - Negative fast atom bombardment ionization of aromatic sulfonic acids: unusual sample-matrix interaction. AB - The most intense ion(s) in negative ion fast atom bombardment (FAB) mass spectra of 2- and 4-benzaldehyde sulfonic acid (BSA) in glycerol or 3-nitrobenzyl alcohol matrix corresponds to a covalent association of the analyte with one or two matrix molecules accompanied by the elimination of a molecule of water. The molecular ion [M - H](-), however, is of low abundance. The identity of the resulting ions [M + nA - H(2)O - H](-) (where M is the analyte and A is the matrix) was confirmed by exact mass measurement using the peak matching technique. These covalent matrix-analyte complexes were not observed when the sulfonic acid functionality in BSA was substituted with COOH, NO(2), and OH or when the sulfonic acid was in salt form. These observations indicate that the free sulfonic acid group in BSA is responsible for the covalent adduct formation. To our knowledge, analyte-matrix covalent association in negative ion FAB spectra of BSA has not been reported previously. PMID- 10717664 TI - A comparative study of a liquid and a solid matrix in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and collision cross section measurements. AB - We present experimental matrix-assisted laser desorption/ionization time-of flight (MALDI-TOF) results comparing a liquid (glycerol/K(4)[Fe(CN)(6)]) and a solid matrix (2,5-dihydroxybenzoic acid, DHB) with respect to analyte signal stability and initial ion velocity. For applications requiring stable production of analyte ions over a long period of time, the liquid matrix is superior to the solid matrix. The stable analyte ion signal obtained from a liquid matrix allowed the measurement of collision cross sections of small poly(ethylene glycol) (PEG(n)) adduct ions in the flight tube with good resolution. The initial velocity of these adduct ions was measured. It was found that analyte molecules from the liquid matrix have initial ion velocities significantly smaller than those from the solid matrix. MALDI-TOF measurements for large molecules using a liquid matrix are therefore likely to result in smaller systematic errors in mass calibrations due to initial ion velocity. PMID- 10717666 TI - High-throughput caco-2 cell permeability screening by cassette dosing and sample pooling approaches using direct injection/on-line guard cartridge extraction/tandem mass spectrometry. AB - A method for high-throughput Caco-2 permeability screening of drug candidates has been developed using thirteen generic drugs as test compounds. The high throughput was achieved by either a sample pooling or a cassette dosing approach, along with the use of a rapid, simple and sensitive direct injection/on-line guard cartridge extraction/tandem mass spectrometric assay that was also developed in this study. It was of concern that possible drug-drug interactions (e.g., inhibition of P-glycoprotein-mediated transport of a drug by another, and/or competition of the drugs for transport pathways), when the cassette dosing regimen was implemented, may give rise to inconsistent results compared with those attained by a traditional single-drug dosing approach. However, the apparent permeability coefficients of the test drugs across Caco-2 monolayers measured by the sample pooling or cassette dosing (up to five drugs co administered in this study) strategy were in good conformity with the data obtained by single-drug dosing followed by discrete sample analysis. PMID- 10717667 TI - In vitro mimicry of metabolic oxidation reactions by electrochemistry/mass spectrometry. AB - The aim of these studies was to investigate the scope and limitations of electrochemistry on-line with mass spectrometry as a quick and convenient way to mimic phase I oxidative reactions in drug metabolism. A compound with previously reported in vitro and in vivo metabolism, the dopamine agonist 2-(N-propyl-N-2 thienylethylamino)-5-hydroxytetralin, was examined in an electrochemistry/mass spectrometry (EC/MS) system. The previously reported N-dealkylation was mimicked by the electrochemical cell while the oxidation of the phenol function was not fully mimicked by the EC/MS system, since the catechol and p-hydroquinone formed were immediately oxidized to the corresponding quinones. Since cytochrome P450 isoenzymes are the most important enzymes in phase I oxidative metabolism, two standard substrates used for the characterization of those enzymes, lidocaine and 7-ethoxycoumarin, were tested in the EC/MS system. The electrochemical cell was capable of mimicking the N-dealkylation of lidocaine but, under the conditions used in our experiments, the O-deethylation of 7-ethoxycoumarin could not be simulated in the electrochemical cell. PMID- 10717668 TI - In vitro exposure of human hemoglobin to the antineoplastic drug thiotepa. PMID- 10717669 TI - Reactions of perfluorotri-n-butylamine fragment ions in the quadrupole ion trap: the origin of artefacts in the perfluorotri-n-butylamine calibration spectrum. PMID- 10717670 TI - Actin and the agile spine: how and why do dendritic spines dance? AB - Since early anatomical descriptions, the existence of dendritic spines has stimulated intense curiosity and speculation about their regulation and function. Research over the past three decades has described an impressive mutability in dendritic-spine number and morphology under a variety of physiological circumstances. Current evidence favors a proposed model in which two pools of actin filaments, one stable and the other dynamic, support both persistent spine structure and rapid spine motility. Potential functions of spine motility and dynamic actin include regulated protein scaffolding, retrograde signaling and synapse stabilization. PMID- 10717671 TI - The bicentennial of the Voltaic battery (1800-2000): the artificial electric organ. AB - Alessandro Volta invented the electric battery at the end of 1799 and communicated his invention to the Royal Society of London in 1800. The studies that led him to develop this revolutionary device began in 1792, after Volta read the work of Luigi Galvani on the existence of an intrinsic electricity in living organisms. During these studies, Volta obtained a series of results of great physiological relevance, which led him to anticipate some important ideas that marked the inception of modern neuroscience. These results have been obscured by a cultural tradition that has seen Volta exclusively as a physicist, lacking interest for biological problems and opposed in an irreversible way to the physiologist, Luigi Galvani. PMID- 10717672 TI - Functions of the pontine nuclei in cerebro-cerebellar communication. PMID- 10717673 TI - Reply PMID- 10717674 TI - The neurobiology of magnetoreception in vertebrate animals. AB - Diverse vertebrate animals can sense the earth's magnetic field, but little is known about the physiological mechanisms that underlie this sensory ability. Three major hypotheses of magnetic-field detection have been proposed. Electrosensitive marine fish might sense the geomagnetic field through electromagnetic induction, although definitive evidence that such fish actually do so has not yet been obtained. Studies with other vertebrates have provided evidence consistent with two different mechanisms: biogenic magnetite and chemical reactions that are modulated by magnetic fields. Despite recent progress, however, primary magnetoreceptors have not yet been identified unambiguously in any animal. PMID- 10717675 TI - From worm to man: three subfamilies of TRP channels. AB - A steadily increasing number of cDNAs for proteins that are structurally related to the TRP ion channels have been cloned in recent years. All these proteins display a topology of six transmembrane segments that is shared with some voltage gated channels and the cyclic-nucleotide-gated channels. The TRP channels can be divided, on the basis of their homology, into three TRP channel (TRPC) subfamilies: short (S), long (L) and osm (O). From the evidence available to date, this subdivision can also be made according to channel function. Thus, the STRPC family, which includes Drosophila TRP and TRPL and the mammalian homologues, TRPC1-7, is a family of Ca2+-permeable cation channels that are activated subsequent to receptor-mediated stimulation of different isoforms of phospholipase C. Members of the OTRPC family are Ca2+-permeable channels involved in pain transduction (vanilloid and vanilloid-like receptors), epithelial Ca2+ transport and, at least in Caenorhabditis elegans, in chemo-, mechano- and osmoregulation. The LTRPC family is less well characterized. PMID- 10717676 TI - Mitochondrial membrane potential and neuronal glutamate excitotoxicity: mortality and millivolts. AB - In the past few years it has become apparent that mitochondria have an essential role in the life and death of neuronal and non-neuronal cells. The central mitochondrial bioenergetic parameter is the protonmotive force, Deltap. Much research has focused on the monitoring of the major component of Deltap, the mitochondrial membrane potential Deltapsim, in intact neurones exposed to excitotoxic stimuli, in the hope of establishing the causal relationships between cell death and mitochondrial dysfunction. Several fluorescent techniques have been used, and this article discusses their merits and pitfalls. PMID- 10717677 TI - A new concept in neurodegeneration: TNFalpha is a silencer of survival signals. AB - The p55 receptor for the pro-inflammatory cytokine tumor necrosis factor alpha (TNFalpha) is best characterized by its ability to induce signals that trigger cell death. However, this is not the only way in which this TNF receptor kills neurons. A new view of neurodegeneration has recently emerged in which a TNF receptor induces death through the 'silencing of survival signals' (SOSS), such as phosphatidylinositol 3' kinase (PI3 kinase), that are activated by the insulin like growth factor 1 receptor. This mechanism of intracellular crosstalk is the most pathophysiologically relevant action of TNFalpha in the brain and is applicable to a broad number of receptors that are localized on the same cell. Treatment of the more-devastating and costly neurodegenerative diseases of our time might be best promoted by increasing the efficacy of neuronal survival factors using new approaches aimed at inhibiting the SOSS. PMID- 10717678 TI - Synchronicity: differential responses to vaccination illuminate dynamics. PMID- 10717679 TI - Ecotone: speciation-prone. PMID- 10717680 TI - Corrigendum. PMID- 10717681 TI - Do edge effects occur over large spatial scales? PMID- 10717682 TI - Generalization versus specialization in plant pollination systems. AB - The long-standing notion that most angiosperm flowers are specialized for pollination by particular animal types, such as birds or bees, has been challenged recently on the basis of apparent widespread generalization in pollination systems. At the same time, biologists working mainly in the tropics and the species-rich temperate floras of the Southern hemisphere are documenting pollination systems that are remarkably specialized, often involving a single pollinator species. Current studies are aimed at understanding: (1) the ecological forces that have favoured either generalization or specialization in particular lineages and regions; (2) the implications for selection on floral traits and divergence of populations; and (3) the risk of collapse in plant pollinator mutualisms of varying specificity. PMID- 10717683 TI - Molecular evolution of flower development. AB - Flowers, as reproductive structures of the most successful group of land plants, have been a central focus of study for both evolutionists and ecologists. Recent advances in unravelling the genetics of flower development have provided insight into the evolution of floral structures among angiosperms. The study of the evolution of genes that control floral morphogenesis permits us to draw inferences on the diversification of developmental systems, the origin of floral organs and the selective forces that drive evolutionary change among these plant reproductive structures. PMID- 10717684 TI - Why are female birds ornamented? AB - Sexual selection is now widely accepted as the main evolutionary explanation of extravagant male ornaments. By contrast, ornaments occurring in females have received little attention and often have been considered as nonadaptive, correlated effects of selection on males. However, recent comparative evidence suggests that female ornaments have evolved quite independently of male showiness. Also, new theoretical models predict that both male mate choice and female contest competition will occur under certain circumstances. This is supported by recent experimental studies. Thus, selection acting on females might be a widespread cause of female ornaments. PMID- 10717685 TI - Stress and the evolution of condition-dependent signals. AB - Stressful events are known to initiate a cascade of physiological mechanisms that are potentially costly for metabolic processes. Although these mechanisms are well understood, the long-term costs and the potential implications for individual condition and behaviour have been considered only recently. Combining information from physiological, ecological and behavioural studies can help us to understand the implications of stress for individual life history strategies. Furthermore, the concept of individual variation in stress tolerance has implications for the immunocompetence handicap hypothesis and the evolution of secondary sexual signals. PMID- 10717687 TI - Sir Ronald Fisher and natural selection. PMID- 10717686 TI - Cognitive ecology: field or label? PMID- 10717688 TI - Reply from E.G. Leigh, Jr. PMID- 10717689 TI - What does sexual trait FA tell us about stress? AB - There is widespread acceptance that fluctuating asymmetry (FA) increases under environmental and genetic stress. Fluctuating asymmetry in sexual traits is thought to be particularly sensitive to stress and to reflect genetic and phenotypic quality. Recent experimental studies show that the relationship between FA and stress is inconsistent, and there is little evidence that sexual traits are especially responsive to stress. PMID- 10717690 TI - Polymorphism of attack and defense. AB - Coevolution of attack and defense occurs in host-parasite systems and various forms of genomic conflict. Attackers benefit when their specific molecules allow entry past host defenses. Defenders gain when their matching biochemical specificities aid recognition. Selection continually favors new attack specificities that avoid matching defense and, in turn, new defense specificities that match novel attackers. The introduction of novel specificities strongly influences the spatial and temporal dynamics of conflict. Lack of reciprocally matching diversity in a particular system suggests biochemical constraints that prevent diversification. New work on cytoplasmic male sterility, B chromosomes and meiotic drive suggests that varying biochemical constraints on recognition cause varying patterns of diversity and spatiotemporal dynamics PMID- 10717691 TI - Liver transplantation in children. PMID- 10717692 TI - Need for liver transplantation in Indian children. AB - BACKGROUND: Liver transplantation (LT) is the most successful and accepted mode of therapy for failing liver in children. Pediatric LT has neither been widely attempted nor its need objectively assessed in our country. OBJECTIVE: To assess requirement of LT in children at a tertiary care hospital. METHODS: Data of children admitted to pediatric GE services (January 1992 to June 1997) were retrospectively analyzed. Subgroups of children with acute liver disease (ALD), chronic liver disease (CLD), neonatal cholestasis syndrome (NCS) and other etiology were evaluated for need for LT according to established criteria. RESULTS: Of the total 301 inpatients with liver diseases assessed at our center, ALD constituted 26% (n=79), CLD 35% (n=106), NCS 27% (n=82) and miscellaneous 11% (n=34). Among ALD, 19% (n=15) had FHF and 67% (n=10) qualified for LT (INR>4.0). Of CLD, LT was warranted in 13% (2/15) cases of Wilson's disease (Wilson's score > 6) and 60% of cirrhotics (n=40/66) with decompensation. NCS comprised extrahepatic biliary atresia (EHBA) in 43, choledochal cyst in 2, paucity of intralobular bile duct (PILBD) in 2, neonatal hepatitis in 23, and was of indeterminate etiology in 12 cases. Of NCS groups, LT was the only therapeutic option in 45% (n=36) of cases (EHBA 34, choledochal cyst 2). Of 34 cases of EHBA requiring LT, 32 presented after 4 months of age and other 2 children had decompensation before four months of age. Both children with choledochal cysts had decompensated liver disease. One patient of Crigler Najjar syndrome type I had kernicterus and qualified for LT. CONCLUSION: Our data shows need for LT in 30% of children with liver diseases constituted by cirrhosis (45%), biliary atresia (38%) and FHF (11%). PMID- 10717693 TI - Cellular and humoral factors in colostrum of HIV infected and uninfected lactating mothers. AB - OBJECTIVE: To compare the cellular and humoral factors in colostrum from HIV infected and uninfected lactating mothers. DESIGN: Cross sectional study. SETTING: Maternity Ward. METHODS: Colostrum was collected from 130 mothers (62 HIV seropositives and 68 HIV seronegatives). These colostrum samples were tested for total cell count, cell viability, differential count, phagocytic activity of macrophages, 'T' cell counts, IgA, IgM and IgG levels. RESULTS: There was a statistically significant decrease in the phagocytosis and 'T' cell number (p <0.001) and in the IgA and IgG levels (p<0. 05) in the colostrum obtained from HIV seropositive mothers as compared to HIV seronegative ones. CONCLUSION: Some of the cellular and humoral factors are reduced in colostrum samples obtained from HIV seropositives as compared to normals. PMID- 10717694 TI - Mission statement of the indian academy of pediatrics PMID- 10717695 TI - Medical management of birth asphyxia. PMID- 10717696 TI - Papuloeruptive xanthomatosis associated with chronic cholestasis. PMID- 10717697 TI - Early experiences with high frequency ventilation in neonates. PMID- 10717698 TI - Correlation of tomographic liver density with serum ferritin levels in multiple transfused children with thalassemia major. PMID- 10717699 TI - Role of flexible fiberoptic bronchoscopy in the diagnosis of tracheobronchial foreign bodies in children. PMID- 10717700 TI - Perinatal mortality in a teaching hospital. PMID- 10717701 TI - Idiopathic primary pulmonary hemosiderosis. PMID- 10717702 TI - Frontonasal dysplasia with corpus callosum lipoma. PMID- 10717704 TI - Intracerebral hydatid cyst in a child with atrial septal defect. PMID- 10717703 TI - Mondini dysplasia of the inner ear with CSF leak-A rare cause of recurrent meningitis. PMID- 10717705 TI - How to make sure that vaccines are properly refrigerated? PMID- 10717706 TI - How to make sure that vaccines are properly refrigerated - reply PMID- 10717707 TI - Defrosting a refrigerator used to store vaccine. PMID- 10717709 TI - A status report on polio eradication in India. PMID- 10717708 TI - Defrosting a refrigerator used to store vaccine - reply PMID- 10717710 TI - Pediatrician as family planning counsellor. PMID- 10717711 TI - Deaths due to poisoning in children. PMID- 10717712 TI - Serum cortisol [correction of control] following replacement therapy in Triple A syndrome. PMID- 10717713 TI - Serum control following replacement therapy in triple A syndrome - reply PMID- 10717714 TI - Healthy baby contests - beyond show, beyond objectivity towards health education. PMID- 10717715 TI - Healthy baby contests. PMID- 10717716 TI - Global developments and the Hospice Information Service. PMID- 10717717 TI - The use of steroids in the management of inoperable intestinal obstruction in terminal cancer patients: do they remove the obstruction? AB - This multicentre, randomized double-blind study was undertaken to assess the efficacy of corticosteroids as a palliative treatment of intestinal obstruction due to advanced and incurable cancer. Thirty-one French palliative care units agreed to participate in the study and 12 actually recruited at least one patient. To be included, patients had to have an advanced cancer with a surgically inoperable bowel obstruction and to have received no specific anticancer therapy within the preceding 28 days. They had to fulfil at least three of the following criteria: vomiting at least twice a day; colicky abdominal pain; no flatus for 12 h or more; no stool for at least 4 days, faecal impaction being excluded; intestinal distension; air-fluid levels or absence of gas in the colon on an abdominal radiograph. Patients were randomized in three groups to receive either a placebo for 3 days (group A), or methylprednisolone 240 mg daily for 3 days (group B) or methylprednisolone 40 mg daily for 3 days (group C). Symptoms were assessed daily but success or failure of the treatment was assessed on day 4, according to the disappearance or persistence of symptoms. Fifty-eight patients were randomized, of whom 52 were able to be evaluated. Details of symptoms and associated treatments are described below. Of 40 patients without a nasogastric tube, symptoms were relieved in 68% of cases versus 33% among placebo treated patients (P = 0.047). In 12 patients who had a nasogastric tube already in place, the results are less significant (60% versus 33% with P = 0.080). Because of the small sample size, no conclusions can be reached about the relative efficacy of low versus high-dose treatment regimes. PMID- 10717718 TI - Do pain and disability differ in depressed cancer patients? AB - Seventy consecutively admitted Chinese patients with advanced cancer and pain (mean age 62 years) were evaluated with the Chinese version of the Hospital Anxiety and Depression Scale (HADS), and with the Geriatric Depression Scale (GDS) Short Form (for patients 65 years old or over) in a prospective study. The HADS and GDS had good concordance (kappa = 0.53). By these depression screening tests, the prevalence of probable depression was 41-49%, and the prevalence of definite depression (HADS > or = 11) was 29%. There was no difference in age, gender and educational level; no difference in nature and severity of pain; and no difference in the level of disability between depressed (using HADS > or = 11) and nondepressed patients with advanced cancer. The study suggests that depression does not correlate with the severity of pain in patients with advanced cancer. It also suggests that impaired activity of daily living (ADL) in these patients is not related to depression. PMID- 10717719 TI - Risk factors for death rattle in terminally ill cancer patients: a prospective exploratory study. AB - Death rattle is frequently observed in cancer patients whose death is impending and may contribute to the severe distress of patients and their family members. To identify risk factors for development and persistency of death rattle, a prospective study was performed on 245 hospice inpatients. One-hundred-and-seven patients (44%) developed death rattle, 71% of whom achieved satisfactory symptom palliation until death. A multiple regression analysis identified neoplasms of brain and lung as independent risk factors for development of death rattle, while refractory symptoms were significantly associated with pulmonary neoplasms and infection/oedema. In conclusion, development of death rattle was influenced by both brain and lung malignancies, while its persistency was mainly determined by pulmonary pathology. A clinical classification of death rattle based on these factors would be established by a further confirmatory study. PMID- 10717720 TI - Measuring patient outcomes in palliative care: a reliability and validity study of the Support Team Assessment Schedule. AB - This study reports the process and results of a psychometric evaluation of a clinical audit tool, the Support Team Assessment Schedule (STAS), used to measure outcomes of palliative care. The STAS was developed in London, UK to audit community palliative care services provided by a support team. The purpose of this study was to evaluate the reliability and validity of the STAS when introduced in a different setting and with different populations from those for which it had been designed. Evaluation of the STAS was completed using multidisciplinary team members, patients and families from a palliative care unit and an oncology unit of a large urban Canadian teaching hospital. The results from the reliability tests revealed a lack of consistency in the use of the tool by team members with simulated patients in clinical scenarios. The validity analysis highlighted the differences between patients, families and health care professionals' perceptions of the same clinical situation. This study provided a valuable perspective on using a previously developed clinical audit tool in different patient populations and clinical settings. Recommendations for future use of the tool are offered. PMID- 10717721 TI - A survey of education and research facilities for palliative medicine trainees in the United Kingdom. AB - In June 1997 a postal questionnaire was sent to all palliative medicine trainees in the United Kingdom to ascertain the resources available to facilitate education and research, and the experiences of trainees who had undertaken research. Questions posed related to protected study time, library facilities, information technology (IT) support, postgraduate educational activities, details of research projects undertaken, level of senior supervision and overall impressions. The response rate was 85%. Thirty-six per cent of respondents were undertaking postgraduate educational courses and 30% of those remaining planned to do so. Protected time for study was allocated to almost all trainees, but access to educational resources was more variable. The availability of several journals and of information technology support was lower than their perceived importance. Satisfaction scores for the availability of books covering areas related to clinical practice were high, but were lower for availability of the internet and of books covering specific training issues. Ninety-one per cent of respondents had been involved in research/audit projects, two-thirds had published at least one article and half had presented an oral or written paper. Details of 162 projects were received, together with problems encountered. The results indicate that many of the foundations for specialist education in palliative medicine are in place and that trainees are highly motivated towards conducting research. The aspects in need of improvement on the educational side include IT facilities and the availability of specific speciality related journals. There is a need for more accessible supervision from senior colleagues and training in research methodology. PMID- 10717722 TI - Dying for care: the experiences of terminally ill cancer patients in hospital in an inner city health district. PMID- 10717723 TI - Are postbereavement research interviews distressing to carers? Lessons learned from palliative care research. PMID- 10717724 TI - Nebulized and intranasal fentanyl in the management of cancer-related breakthrough pain. PMID- 10717725 TI - The importance of low magnesium in palliative care: two case reports. PMID- 10717726 TI - Paracentesis--an effective method of symptom control in the palliative care setting? PMID- 10717727 TI - Palliative care in Romania. PMID- 10717728 TI - Why are trials in palliative care so difficult? PMID- 10717729 TI - Cardiopulmonary resuscitation in Wales. PMID- 10717730 TI - Absence of cross-tolerance and the situational specificity of tolerance. PMID- 10717731 TI - Thalidomide for night sweats in patients with advanced cancer. PMID- 10717732 TI - Treatment of biliary ascariasis in China. PMID- 10717733 TI - Insecticide-treated materials for malaria control in Latin America: to use or not to use? AB - Studies on the protective efficacy of insecticide-treated materials (ITMs) in Plasmodium vivax endemic areas of Latin America have not yielded sufficient evidence for recommendation of their extensive use in the region. Therefore 2 randomized community trials have been conducted on the Pacific Coast of Nicaragua which analysed the minimum coverage of ITMs needed to be effective against malaria. For the characterization of the study area, epidemiological and entomological baseline surveys and household interview surveys were undertaken. Thereafter the communities were paired (6 pairs in the 1st year and 13 pairs in the 2nd year) according to 4-monthly reported malaria incidence rates, population size and bednet coverage, and then randomly allocated to intervention and control groups. In the intervention groups, bednets were impregnated with lambdacyhalothrin; in the control groups, people received general health education. Anopheles albimanus was found to be the main vector with marked indoor biting behaviour late in the evening. P. vivax (99%) clearly outweighed P. falciparum (1%) with low parasite prevalence rates in the asymptomatic general population (8%) and low parasite densities. The protective efficacy of ITMs varied according to the coverage achieved: protective efficacy was 68% in communities with an average ITM coverage of 50% (10 pairs); 31% in communities with an ITM coverage of 16-30% (4 pairs); and no protective efficacy in communities with ITM coverage below 16% (5 pairs). The comparison with other P. vivax endemic areas in Latin America showed that the vector's late biting behaviour and the indoor preference (where ITMs have a repellent effect) probably led to the favourable results in the study. In malaria endemic areas of Latin America, where P. vivax is predominant, studies on vector behaviour should be conducted in order to predict the impact of ITMs on malaria transmission. PMID- 10717734 TI - Are insecticide-treated nets affordable? Relating costs of two sizes of nets and net re-treatment with basic household expenditures. PMID- 10717735 TI - Comparative susceptibility of different members of the Anopheles culicifacies complex to Plasmodium vivax. AB - Three colonized species of the Anopheles culicifacies complex (species A, B and C) were compared with Anopheles stephensi (control) for susceptibility to Plasmodium vivax. In feeding experiments involving over 400 paired comparisons, the mean number of oocysts, oocyst rate and sporozoite rate were found to be significantly different. Of the test groups, species A had the highest percentage of mosquitoes with oocysts (> 60%) and sporozoites (> 50%). An. culicifacies species B were least susceptible, less than 5% had oocysts and less than 2% had sporozoites in the salivary glands. The results are discussed in the light of the vectorial potential of the members of the An. culicifacies complex observed in field studies. PMID- 10717736 TI - Susceptibility of Anopheles quadriannulatus Theobald (Diptera: Culicidae) to Plasmodium falciparum. AB - Anopheles quadriannulatus, the cattle-feeding member of the An. gambiae complex, was fed human blood which contained cultured gametocytes of Plasmodium falciparum, using a membrane feeding system. After 7 days, 33-80% of the mosquitoes that took a blood meal contained developing oocysts. In 7 out of 12 females sporozoites were found in the salivary glands 14 days after the infectious blood meal. Control groups of An. gambiae s.s. and An. stephensi became readily infected with > 90% developing oocysts. All of the An. gambiae dissected 14 days after the infectious blood meal had sporozoites in their salivary glands. The results demonstrate that An. quadriannulatus is susceptible to infections with P. falciparum. PMID- 10717737 TI - Aedes aegypti in Ho Chi Minh City (Viet Nam): susceptibility to dengue 2 virus and genetic differentiation. AB - Aedes aegypti is the principal vector of dengue viruses, responsible for a viral infection that has become a major public health concern in Asia. In Viet Nam, dengue haemorrhagic fever was first detected in the 1960s and is now a leading cause of death in childhood. We studied the variability in competence of Ae. aegypti as a vector for dengue 2 virus and genetic differentiation in this mosquito species. Twenty mosquito samples collected in 1998 in Ho Chi Minh City were subjected to oral infection and isoenzyme polymorphism analysis by starch gel electrophoresis. Ae. aegypti populations from the centre of Ho Chi Minh City were genetically differentiated and their infection rates differed from those of populations from the commuter belt. These results have implications for insecticidal control during dengue outbreaks. PMID- 10717738 TI - Genetic analysis of Plasmodium falciparum infections on the north-western border of Thailand. AB - Genetic characterization of Plasmodium falciparum infections in north-western Thailand, a region of low transmission intensity (1 infection/person each year), has found a comparable number of parasite genotypes per infected person to regions with hyperendemic malaria. Clone multiplicity and parasite diversity were found to be homogeneous across 129 infected individuals comprising a range of age groups (1.32 parasite genotypes; n = 98), patients (aged 2-16 years) with recrudescent infections (1.54; n = 13), and pregnant women (1.61; n = 18). Individuals belonging to groups with a high risk of infection, as deduced by clinical epidemiology, did not harbour a higher number of clones per infection, nor greater parasite diversity than low-risk groups. In fact, multiple genotype infections were as common in low-risk groups, suggesting that there is frequent transmission of polyclonal infections from a single inoculum, rather than superinfection. Such a polyclonal transmission system would enable generation of extensive parasite diversity by recombination, despite the low level of transmission. However, co-infection with P. vivax was associated with fewer P. falciparum genotypes per infection. PMID- 10717739 TI - Palm trees as ecological indicators of risk areas for Chagas disease. PMID- 10717740 TI - The Vellore Vibrio watch 1996-98. PMID- 10717741 TI - Detection and identification of the aetiological agent of Mediterranean spotted fever (MSF) in two genera of ticks in Cyprus. PMID- 10717742 TI - Possible increased risk of hospital admission for asthma in women in Zaria, northern Nigeria. PMID- 10717743 TI - Excess mortality from hepatocellular carcinoma in an HCV-endemic township of an HBV-endemic country (Taiwan). AB - Taiwan is an endemic area of hepatitis B virus (HBV). All previous studies have concluded that HBV is the major cause of hepatocellular carcinoma (HCC) in Taiwan. An HBV- and hepatitis C virus (HCV)-endemic township, Tzukuan, in southern Taiwan has been identified with the prevalence of 24% for HB surface antigen (HBsAg) and 37% for anti-HCV antibodies. To elucidate the aetiology of HCC and impact of HCV in this township, we conducted a case-control study and compared HBV-related liver cancer mortality in Tzukuan and Taiwan as a whole. Based on cancer registration datasets of 2 medical centres from 1991 to 1995, we recruited 18 male and 9 female HCC cases from the study township. Their mean age (+/- standard deviation) was 60.3 (+/- 7.3) years. Randomly sampled from a community-based survey, 4 age- (+/- 2 years) and sex-matched residents were selected as community controls for each HCC case. The HBsAg carrier rate was 40.7% in cases and 25.0% in controls (P = 0.1). Anti-HCV positive rate was 88.9% in cases and 53.7% in controls (P = 0.008). Age-adjusted liver cancer mortality in Tzukuan (36.5 per 10(5)) was significantly higher than that of Taiwan as a whole (20 per 10(5)). Based on the HBsAg-positive rate among HCC patients (40.7% in Tzukuan and 77.4-86.6% in Taiwan), the estimated HBV-related liver cancer mortality was similar in Tzukuan (14.9 per 10(5)) and Taiwan (15.8-17.3 per 10(5)). We concluded that HCV was the major risk factor for excess liver cancer mortality in this HCV-endemic township of the HBV-endemic country. PMID- 10717744 TI - Evaluation of a synthetic tripeptide as antigen for detection of IgM and IgG antibodies to Trypanosoma cruzi in serum samples from patients with Chagas disease or viral diseases. AB - An enzyme-linked immunosorbent assay (ELISA) has been developed to detect IgG and IgM antibodies in human sera against a synthetic tripeptide derived from a hybrid peptide containing 3 specific epitopes from Trypanosoma cruzi. This assay was compared in Brazil with one using conventional antigen, the alkaline crude extract. Serum samples were divided into positive (40 samples) or negative (107 samples) for Chagas disease. Positive samples included 9 serum samples from patients with acute Chagas disease, while negative samples included 57 samples from patients suffering from viral diseases. The total percentages of IgG positive samples from patients with chronic Chagas disease for alkaline extract and synthetic tripeptide were 93.5% and 100%, respectively. All samples from patients with acute Chagas disease were confirmed positive for IgM antibodies by using both the tripeptide and the alkaline extract. However, the results for anti T. cruzi IgM in the group of chronic Chagas disease patients demonstrated that 41.9% were positive for IgM with the alkaline extract, while the synthetic peptide showed a significantly lower number of positive samples (12.9%). The serum samples from healthy people showed similar results for both antigens. However, 40% of the serum samples from patients presenting with viral diseases were IgM positive for Chagas disease when assayed with conventional antigen; with the synthetic tripeptide as antigen, 100% of this group of samples were found to be negative. Thus, as the results of ELISA with synthetic tripeptide showed higher rates of sensitivity and specificity than ELISA with conventional antigen, the former should be included as a laboratory tool in the serodiagnosis of Chagas disease. PMID- 10717745 TI - Comparison of PCR results using scrape/exudate, syringe-sucked fluid and biopsy samples for diagnosis of cutaneous leishmaniasis in Ecuador. PMID- 10717746 TI - A study on the provocative day test effect of ivermectin and albendazole on nocturnal periodic Wuchereria bancrofti microfilaraemia. AB - We conducted a randomized double-blind placebo-controlled study, in the Ahanta West District of Ghana, on the provocative day test effect of ivermectin and albendazole alone and in combination on nocturnal periodic Wuchereria bancrofti microfilaraemia. Sixty-three individuals with high night-time microfilaria (mf) intensities were identified in 1997 or 1998 and randomized into 4 groups. Blood samples for mf were then collected from the same individuals in the daytime (between 09:00 and 15:00) immediately before and 30-50 min after treatment. Groups 1-4 were treated with ivermectin alone (150-200 micrograms/kg), albendazole alone (400 mg), the combination of ivermectin and albendazole, and placebo, respectively. Intensities of mf in daytime samples were considerably lower than in night-time samples. Neither ivermectin or albendazole alone nor their combination provoked significant liberation of W. bancrofti mf into the peripheral circulation after the daytime treatment. PMID- 10717747 TI - Field diagnosis of Echinococcus granulosus infection among intermediate and definitive hosts in an endemic focus of human cystic echinococcosis. AB - Human, canine and ovine echinococcosis prevalence was determined in a highland community located in the central Peruvian Andes during 1997 and 1998. Human echinococcosis was determined using portable ultrasonography, chest X-ray examination, and an enzyme-linked immunoelectrotransfer blot (EITB) assay. Canine echinococcosis was determined using microscopy stool examinations and a coproantigen detection enzyme-immunoassay (EIA) for Echinococcus granulosus. Ovine echinococcosis was determined by an EITB assay for sheep echinococcosis and necropsy examination of viscera from domestic slaughtered animals. An abdominal ultrasound, a chest X-ray examination and an EITB for echinococcosis were performed on 214 subjects (45% of the village population). The frequency of presumptive liver/abdominal, lung and liver-lung hydatid cysts was 5.1% (11/214), 3.7% (8/214) and 0.5% (1/214), respectively. The overall prevalence of human cystic echinococcosis was 9.3% (20/214). The frequency of canine echinococcosis was 46% (23/50) and 32% (16/50) by the coproantigen EIA test and arecoline purging, respectively. The frequency of sheep echinococcosis was 65% (22/34) by the EITB and 38% (13/34) by necropsy. We demonstrated a high prevalence of human and animal echinococcosis in this Peruvian village. In remote areas where echinococcosis is endemic, both the coproantigen EIA and arecoline purging may be used for the study of canine echinococcosis; the EITB is useful in establishing the diagnosis of echinococcosis in sheep prior to necropsy. PMID- 10717748 TI - Plasma glutamine levels and falciparum malaria. AB - Glutamine deficiency is associated with increased rates of sepsis and mortality, which can be prevented by glutamine supplementation. Changes in glutamine concentration were examined in Ghanaian children with acute falciparum malaria and control cases. The mean (SD) plasma glutamine concentration was lower in patients with acute malaria (401 (82) mumol/L, n = 50) than in control patients (623 (67) mumol/L, n = 7; P < 0.001). Plasma glutamine concentrations all rose in convalescence. The mean (SD) increase in plasma glutamine was 202 (123) mumol/L (n = 18; P < 0.001) compared with acute infection. We conclude that acute falciparum malaria is associated with large decreases in plasma glutamine and these falls may increase susceptibility to sepsis and dyserythropoeisis. PMID- 10717749 TI - A review of the spectrum of clinical ocular fundus findings in P. falciparum malaria in African children with a proposed classification and grading system. AB - Ocular fundus pathology in Plasmodium falciparum malaria is common and has prognostic significance. We have made a collaborative effort to document the ocular features in several populations. Based on examination of 735 patients in Malawi, Kenya and The Gambia by direct and indirect ophthalmoscopy with dilated pupils, we have determined that the 5 distinct clinical features (in order of frequency) include retinal whitening, haemorrhages, unique vessel abnormalities, papilloedema, and cotton wool spots. Photographs and descriptions of these are presented, along with a proposed grading scheme. PMID- 10717750 TI - Anaemia caused by asymptomatic Plasmodium falciparum infection in semi-immune African schoolchildren. AB - A cohort of 250 Ghanaian schoolchildren aged 5-15 years was followed clinically and parasitologically for 4 months in 1997/98 in order to study the effect of asymptomatic Plasmodium falciparum infections on haematological indices and bone marrow responses. Of the 250 children 65 met the predefined study criteria. Thus, 14 children were parasite-free throughout (group 1), 44 had P. falciparum in all blood samples collected but no symptoms of malaria (group 2), and 7 had 1 malaria attack during the study period (group 3). At the end of the study the mean haemoglobin (Hb) level in group 1 was 123 g/L, significantly higher than the value of 114 g/L in groups 2 and 3 (P < 0.02, adjusted for age and splenomegaly). The low Hb in group 2 was associated with subnormal plasma iron. Low Hb was associated with elevated erythropoietin (EPO) levels, and there was a positive correlation between EPO and reticulocyte counts. However, the reticulocyte response to EPO was more pronounced in uninfected than in infected children, suggesting a partial interference with erythropoiesis in asymptomatic infections. Children with asymptomatic infections had significantly higher plasma levels of tumour necrosis factor than uninfected children (geometric means 50 ng/L and 27 ng/L, respectively, P < 0.001) and this cytokine may contribute to bone-marrow suppression and disturbed iron metabolism. We suggest that asymptomatic malaria leads to a homeostatic imbalance in which erythrocyte loss due to parasite replication is only partially compensated for by increased erythropoiesis. The consequences of the reduced Hb levels on the development and cognitive abilities of children with asymptomatic infections, and the risk of precipitation of iron deficiency, deserve further study and should be considered in malaria control programmes that aim at reducing morbidity rather than transmission. PMID- 10717751 TI - Preliminary observations on the intestinal pathology of mice infected with Trypanosoma brucei brucei. PMID- 10717752 TI - Reactivation of Chagas disease manifested by skin lesions in a patient with AIDS. PMID- 10717753 TI - Bancroftian filariasis in a paediatric population: an ultrasonographic study. AB - Little is known about lymphatic filariasis or the anatomical location of adult Wuchereria bancrofti in children. Seventy-eight children from Greater Recife, 23 microfilaria-positive and 55 microfilaria-negative in approximately 60 microL blood, underwent ultrasound examinations of the major superficial lymphatic vessels of the limbs, scrotal area (boys), and breast area (girls). The characteristic movements of adult worms, known as the filaria dance sign (FDS), were detected in 11 (14.1%) children. In 9 boys, the FDS was detected in lymphatic vessels of the scrotal area (8, ages 14-16) and the inguinal cord (1, age 11). In girls, the FDS was detected in a crural lymphatic vessel and an axillary lymph node. FDS detection was more common in boys (P = 0.06), older children (P = 0.001), and children with microfilaraemia (P = 0.05). Diffuse lymphangiectasia was visualized in 4 boys (ages 14-16) and 2 children had clinical signs of filariasis. These ultrasonographic findings associate W. bancrofti with both infection and disease in children. PMID- 10717754 TI - Atovaquone and proguanil hydrochloride followed by primaquine for treatment of Plasmodium vivax malaria in Thailand. AB - Chloroquine-resistant Plasmodium vivax malaria has been reported in several geographical areas. The P. vivax life-cycle includes dormant hepatic parasites (hypnozoites) that cause relapsing malaria weeks to years after initial infection. Curative therapy must therefore target both the erythrocytic and hepatic stages of infection. Between July 1997 and June 1998, we conducted an open-label study in Thailand to evaluate the efficacy and tolerability of a sequential regimen of combination atovaquone (1000 mg) and proguanil hydrochloride (400 mg), once daily for 3 days, followed by primaquine (30 mg daily for 14 days) for treatment of vivax malaria. All 46 patients who completed the 3-day course of atovaquone-proguanil cleared their parasitaemia within 2-6 days. During a 12-week follow-up period in 35 patients, recurrent parasitaemia occurred in 2. Both recurrent episodes occurred 8 weeks after the start of therapy, consistent with relapse from persistent hypnozoites rather than recrudescence of persistent blood-stage parasites. The dosing regimen was well tolerated. Results of this trial indicate that atovaquone-proguanil followed by primaquine is safe and effective for treatment of vivax malaria. PMID- 10717755 TI - Randomized controlled trials of 5- and 14-days primaquine therapy against relapses of vivax malaria in an Afghan refugee settlement in Pakistan. AB - Primaquine is the only drug available that can eliminate hypnozoites from the liver and prevent relapses of vivax malaria. The World Health Organization recommends a course of 14-21 days depending on region and strain. The National Malaria Control and Eradication Programmes of Pakistan and India have adhered to a 5-day course as their standard as it is deemed more practical to implement and because facility for detecting glucose 6-phosphate dehydrogenase (G6PD) deficiency is seldom available at the periphery. Evidence for the efficacy of the 5-day regimen is controversial or lacking. Two, year-long, randomized controlled trials were undertaken in an Afghan refugee camp in north-western Pakistan using a process of passive case detection and treatment at the camp's clinic: the first trial compared treatment with chloroquine alone versus chloroquine plus 5-days primaquine, the second trial compared chloroquine alone versus chloroquine plus 14-days primaquine. Chloroquine is not hypnozoitocidal and was administered to eliminate the erythrocytic stages and to alleviate clinical symptoms. The daily primaquine dose was 0.25 mg/kg bodyweight and the total chloroquine dose was 25 mg/kg over 3 days. During the first trial 52% (129/250) of the non-primaquine group recorded a 2nd clinical-parasitaemic episode and 23% recorded a 3rd, whereas 51% (128/250) of the 5-days primaquine group reported a 2nd episode and 21% recorded a 3rd. During the second trial 49% (49/100) of the non-primaquine group recorded a 2nd episode and 25% recorded a 3rd, whereas only 32% (32/100) of the 14-days primaquine group recorded a 2nd and only 2% recorded a 3rd. The 5 days primaquine regimen has no value as an anti-relapse therapy and should be abandoned. In extended tests in vivo in which vivax cases (n = 31) were treated with chloroquine 25 mg/kg and 14-days primaquine, there was no parasite recrudescence within 28 days and hence no evidence of resistance to chloroquine. PMID- 10717756 TI - Sulfadoxine-pyrimethamine effectiveness against Plasmodium falciparum malaria in Mpumalanga Province, South Africa. PMID- 10717757 TI - Amodiaquine remains effective for treating uncomplicated malaria in west and central Africa. AB - Many countries in Africa are now confronted with the dilemma of shifting drug policies for uncomplicated falciparum malaria from chloroquine, which has become largely ineffective, to a new first-line drug and amodiaquine is one of the possible options. A multicentre, open-label randomized controlled trial of amodiaquine 30 mg/kg vs chloroquine 25 mg/kg over 3 days was performed in Senegal, Cameroon, Gabon, and Burkina Faso between 1996 and 1998 and patients were followed-up for 14 days. Sensitivity of isolates in vitro and whole blood levels of chloroquine and amodiaquine were also measured. The primary efficacy parameter was parasitological clearance on day 14 (parasitological success). The secondary efficacy parameter was absence of signs/symptoms of malaria on day 14 (clinical success). Among the 364 patients randomized and receiving the assigned treatment (chloroquine n = 185, amodiaquine n = 179), 137 and 139, respectively, reached the primary endpoint. Amodiaquine proved significantly more effective than chloroquine. The summary odds ratio (95% CI) was 7.79 (4.54-13.35) for parasitological success, and 6.3 (3.4-11.68) for clinical success. Sensitivity in vitro and chloroquine blood levels were good predictors of chloroquine failure. Amodiaquine remains effective for treating uncomplicated falciparum malaria in areas of West and Central Africa where chloroquine resistance is prevalent. However, measures should be taken to protect the lifespan of amodiaquine where the drug is introduced for use. PMID- 10717758 TI - Assessment of pyronaridine activity in vivo and in vitro against the hepatic stages of malaria in laboratory mice. PMID- 10717759 TI - Alternatives to bodyweight for estimating the dose of praziquantel needed to treat schistosomiasis. AB - Data on age, height and mid upper-arm circumference (MUAC) from nearly 6000 schoolchildren in Ghana, Tanzania and Malawi (not MUAC) were used to examine their power to predict bodyweight and thus the dosage of praziquantel required to treat schistosomiasis. Height was found to provide a simple and reasonably accurate estimate of weight, and about 75% of children would have been given a dosage of praziquantel within the range normally given using bodyweight at a dosage of 40 mg/kg bodyweight. The upper and lower ranges in dosage did not exceed dosages of praziquantel which have been used before or are currently recommended to treat schistosomiasis. A pole marked with the number of tablets could thus be used as a simple way to determine the dose of praziquantel to treat children in school-based health programmes. PMID- 10717760 TI - Antimicrobial susceptibility test of Helicobacter pylori isolated from Jos, Nigeria. AB - Fifty-five strains of Helicobacter pylori isolated from November 1997 until October 1998 from 33 female and 22 male adults attending for endoscopy at the Evangel Hospital, Jos, Nigeria were assayed for antibiotic susceptibility to amoxycillin, clarithromycin, metronidazole and tetracycline by the E-test strip method. Minimum inhibitory concentration (MIC) within the attainable peak serum concentrations for each drug was used as the parameter to determine the susceptibility of H. pylori. The results showed 100% susceptibility for amoxycillin, 89.0% for tetracycline, 87.3% for clarithromycin and 60% for metronidazole. The MIC50 and MIC90 values were: 0.016 microgram/mL and 0.75 microgram/mL for amoxycillin, 0.016 microgram/mL and 2 micrograms/mL for clarithromycin, 0.094 microgram/mL and 12 micrograms/mL for tetracycline, and 2 micrograms/mL and > 48 micrograms/mL for metronidazole. The MIC90 values for metronidazole (> 48 micrograms/mL) and tetracycline (12 micrograms/mL) were in each case higher than the break-point value (peak serum concentrations) of 8 micrograms/mL for metronidazole and 3 micrograms/mL for tetracycline. This pattern of resistance to metronidazole and tetracycline has to be considered when therapeutic regimens against H. pylori contain either or both drugs. PMID- 10717761 TI - Nitric oxide in murine malaria: divergent roles in blood and brain suggested by voltametric measures. PMID- 10717762 TI - Inflammatory cytokines following diethylcarbamazine (DEC) treatment of different clinical groups in lymphatic filariasis. AB - In an earlier study in Indonesia we reported on adverse reactions to diethylcarbamazine (DEC) in brugian filariasis patients identified as microfilaraemics (n = 26), endemic normals (n = 11) and elephantiasis patients (n = 17). To assess the link between adverse reactions and cytokines we have now analysed an array of inflammatory mediators in plasma samples collected during the same study. Pre-treatment levels of interleukin (IL)-6 and soluble tumour necrosis factor receptor 75 (sTNF-R75) were higher in elephantiasis patients compared to microfilaraemics and endemic normals, indicating the presence of an ongoing inflammation in patients with chronic disease. After initiation of treatment, the levels of IL-6 and LPS-binding protein (LBP) were consistently and significantly higher in microfilaraemics who suffered most from adverse reactions compared with endemic normals and elephantiasis patients. In microfilaraemics the levels of sTNF-R75 increased after treatment to reach levels recorded in elephantiasis patients. IL-6 increased early, concurrent with the development of adverse reactions and peaked by 24 h post treatment. The levels of LBP and sTNF R75 in microfilaraemics also increased to peak, later than IL-6, at 32 h post DEC therapy. Although changes were recorded in IL-8 and IL-10 levels in some individuals, no significant differences were found between the 3 clinical groups. These results demonstrate that intake of DEC leads to an increase in a selected number of inflammatory mediators in the group of filarial patients who suffer most from adverse systemic reactions. PMID- 10717763 TI - Delayed culture of Leishmania in skin biopsies. AB - Between January 1997 and October 1998, 16 skin biopsies collected from 13 patients with cutaneous leishmaniasis in French Guiana were inoculated in culture medium after travel for 3-17 days from the place of biopsy to the culture laboratory in France. Each biopsy fragment was introduced near the flame of a Bunsen burner into the transport medium (RPMI medium supplemented with 10% fetal calf serum) which was maintained at ambient temperature during postal delivery to France. In France the biopsies were ground in sterile saline before being inoculated into NNN culture tubes. The cultures were incubated at 25 degrees C and subcultured every week until the 5th week. The cultures were positive in 9 cases, remained negative in 4, and were contaminated in 3 cases. Positive results were obtained at all seasons and for 3 different Leishmania species. The study indicates that delayed culture can yield useful results from biopsies taken in field conditions. PMID- 10717764 TI - 'Tropical' cases admitted to the Albert Dock Hospital in the early years of the London School of Tropical Medicine. PMID- 10717765 TI - The tuberculosis pandemic: implications for health in the tropics. PMID- 10717766 TI - Risk factors for severe malaria: importance of careful study design. PMID- 10717767 TI - Adaptation of human skeletal muscle to training and anabolic steroids. PMID- 10717768 TI - Objectivity and reliability in qualitative analysis: realist, contextualist and radical constructionist epistemologies. AB - The effect of the individual analyst on research findings can create a credibility problem for qualitative approaches from the perspective of evaluative criteria utilized in quantitative psychology. This paper explicates the ways in which objectivity and reliability are understood in qualitative analysis conducted from within three distinct epistemological frameworks: realism, contextual constructionism, and radical constructionism. It is argued that quality criteria utilized in quantitative psychology are appropriate to the evaluation of qualitative analysis only to the extent that it is conducted within a naive or scientific realist framework. The discussion is illustrated with reference to the comparison of two independent grounded theory analyses of identical material. An implication of this illustration is to identify the potential to develop a radical constructionist strand of grounded theory. PMID- 10717769 TI - A new family handedness sample with findings consistent with X-linked transmission. AB - A family handedness study of 2632 families and 8605 offspring was conducted. Of the 2632 parental couples, there were 2123 in which both parents were right handed (RR), 232 in which the mother was left-handed and the father right-handed (LR), 254 in which the mother was right-handed and the father was left-handed (RL), and 23 in which both parents were left-handed (LL). Results showed some important differences from the composite results of four earlier large scale studies that had also employed the same criterion of handedness (writing hand). These had collectively found that the incidence of left-handed offspring, of both sexes, was significantly lower for RR couples than for LR or RL couples, but not lower than for LL couples. Present results, however, suggest an X-linked pattern of genetic influence on handedness. The LR parents produced significantly more left-handed offspring than did RR couples, and this was particularly the case for sons; but while RL couples produced a higher incidence of left-handed daughters than did RR parents, they failed to produce a higher incidence of left-handed sons than did RR couples. Additionally, the present sample showed a significantly greater incidence of left-handed offspring of LL couples than of RR couples, a finding that, while predicted by genetic-influence theories of handedness, was not found in the composite of previous comparison samples. The finding of sex linkage is potentially important, but will require replicative studies that take special care to preclude possible biasing factors such as selective volunteering of participants and inaccurate offspring ascriptions of parental (particularly paternal) handedness. PMID- 10717770 TI - Diurnal variations in the mood and performance of highly practised young women living under strictly controlled conditions. AB - The diurnal variation in a range of psychological functions and core body temperature were investigated in a series of studies involving a total of 24 highly practised young women who lived in a controlled environment and on a strictly regimented 24-hour routine for 6 or 7 days. Ten participants were exposed to the natural light/dark cycle (L/Dc) through windows, whereas the 14 remaining participants saw no daylight, but all had access to normal clock time. A battery of mood and performance tests was completed every 2 hours whilst awake (08:00-00:00), resulting in nine equally spaced measures per waking day. Average time of day (ToD) functions were calculated from the last 5 or 6 days spent in the controlled environment. Significant ToD effects were found for many of the variables taken although the nature of these effects differed across measures, with a 'post-lunch dip' being observed at 16:00 in some variables. Analysis of the standardized data established that all variables presented reliably different ToD functions to core body temperature, whilst factor analyses indicated possible relationships between the variables. It was concluded that those variables that exhibited diurnal variation showed trends that did not parallel those in core body temperature. PMID- 10717771 TI - Dogs as catalysts for social interactions: robustness of the effect. AB - It is known that pet dogs can act as catalysts for human social interactions, and it has been suggested that this may enhance feelings of well-being. Two studies were carried out to establish the robustness of this effect. In Study 1, a highly trained dog was used to ensure that the dog itself did not solicit attention from passers-by, and data were collected across a range of normal daily activities in which a dog could be included, not confined to conventional dog walking areas as in previous studies. Being accompanied by a dog increased the frequency of social interactions, especially interactions with strangers. In Study 2, also using a trained dog, a different (male) participant observer was dressed either smartly or scruffily. Although there were significantly more interactions when he was smartly dressed, the greatest effect was between the Dog present and No Dog conditions irrespective of the handler's dress. It is concluded that the social catalysis effect is very robust, which opens the way for investigating possible consequences of the effect for well-being and health. PMID- 10717772 TI - The ability to detect unseen staring: a literature review and empirical tests. AB - There is evidence to suggest that individuals not only believe in their ability to detect an unseen gaze, but may genuinely be able to do so. The present study reviewed past research and sought to determine whether such a phenomenon was empirically demonstrable. In Expt 1, 12 participants responded to 12 sequences (with feedback in the last nine) of 20 trials each, with staring or non-staring episodes based on Sheldrake's random number sequences. No effects were obtained when no feedback was given. With feedback, more accurate than mean chance expectation (MCE) results were obtained on staring, but no difference on non staring trials. However other 'normal' explanations of ESP phenomena discuss the possibility of matching in bias between experimental sequences and participants' representations of randomness. Tests of the sequences found more alternations than expected, a feature typical of subjective randomness, but the increase in accuracy found on staring trials only was not consistent with this explanation. It was concluded that the improvement in accuracy with feedback is likely to be due to implicit learning, given the structure in non-random sequences. This hypothesis was supported in Expt 2 where 12 participants responded to 12 'genuinely random' sequences, and no differences in accuracy from MCE were obtained. PMID- 10717773 TI - Orienting to exogenous cues and attentional bias to affective pictures reflect separate processes. AB - On the basis of psychophysiological research it has been argued that pleasant and unpleasant pictures inhibit orienting to abrupt startle stimuli. Converging psychophysical evidence was sought for this finding using a modified version of the spatial precueing paradigm (Posner, 1980). Specifically, exogenous cues were presented to the left and right of neutral, pleasant and unpleasant picture stimuli. In contrast to the experimental hypothesis, pleasant and unpleasant pictures failed to affect attention to exogenous cues despite slowing overall reaction times in comparison to neutral pictures. This null finding was replicated in a further experiment where rate arousal was varied within the pleasant and unpleasant picture groups. However, unpleasant pictures produced more task interference than pleasant pictures and highly arousing pictures produced more interference than pictures rated low in arousal. PMID- 10717774 TI - Facial distinctiveness: its measurement, distribution and influence on immediate and delayed recognition. AB - It is conventionally assumed that many faces are relatively typical and few are distinctive (e.g. Valentine, 1991), producing a highly skewed distribution. However, Burton and Vokey (1998) argue that the distribution will be normal, and our review of previous research suggested this is true. In three studies we explored the distributions using different techniques to estimate distinctiveness. Both traditional ratings and pairwise selection produced normal distributions. However, ratings emphasizing the degree of deviation from a typical face were skewed towards the distinctive end of the scale. The instructions given when distinctiveness is rated may not necessarily oppose typicality with distinctiveness: a face that is relatively typical might also stand out in a crowd because of some particular feature, familiarity or a host of other reasons. In our fourth study, recognition memory was predicted by all of the distinctiveness measures, with the relationship being stronger after a 5-week delay than in the immediate test. PMID- 10717775 TI - Sex-typicality and attractiveness: are supermale and superfemale faces super attractive? AB - Many animals find extreme versions of secondary sexual characteristics attractive, and such preferences can enhance reproductive success (Andersson, 1994). We hypothesized, therefore, that extreme versions of sex-typical traits may be attractive in human faces. We created supermale and superfemale faces by exaggerating all spatial differences between an average male and an average female face. In Expt 1 the male average was preferred to a supermale (50% exaggeration of differences from the female average). There was no clear preference for the female average or the superfemale (50% exaggeration). In Expt 2, participants chose the most attractive face from sets of images containing feminized as well as masculinized images for each sex, and spanning a wider range of exaggeration levels than in Expt 1. Chinese sets were also shown, to see whether similar preferences would occur for a less familiar race (participants were Caucasian). The most attractive female image was significantly feminized for faces of both races. However, the most attractive male image for both races was also significantly feminized. These results indicate that feminization, rather than sex exaggeration per se, is attractive in human faces, and they corroborate similar findings by Perrett et al. (1998). PMID- 10717776 TI - Effect of low-dose oral contraceptives on androgenic markers and acne. AB - Oral contraceptives (OC) suppress excess androgen production; however, different progestins in combination with low-dose estrogens produce divergent effects on sex hormone-binding globulin (SHBG) and testosterone that may influence clinical outcomes. This multicenter, open-label, randomized study compared biochemical androgen profiles and clinical outcomes associated with two OC containing the same amounts of ethinyl estradiol (EE, 20 micrograms) but different progestins, levonorgestrel (LNG, 100 micrograms), and norethindrone acetate (NETA, 1000 micrograms). Fifty-eight healthy women (18-28 years old) received three cycles of treatment with LNG/EE (n = 30) or NETA/EE (n = 28). The results showed that LNG reduced androgen levels in three compartments--adrenal, ovarian, and peripheral. NETA reduced only adrenal and peripheral androgens. Despite a 2.2-fold greater relative increase in SHBG with NETA than LNG, bioavailable testosterone (T) was reduced by the same amount with LNG and NETA. Both treatments improved acne and were well tolerated. Low-dose OC (EE, 20 micrograms) are effective in reducing circulating androgens and acne lesions without causing weight gain. Although LNG and NETA affected secondary markers differently, both OC formulations produced an equivalent decrease in bioavailable. PMID- 10717777 TI - The trimonthly combination oral contraceptive regimen: is it cost effective? AB - The extended use of combination oral contraceptive pills (COCPs) to decrease the frequency of withdrawal bleeding can be convenient and beneficial to women. We conducted a cost-effective analysis comparing the standard regimen (21 days of estrogen/progestin) to a trimonthly regimen (84 days of estrogen/progestin) followed by a pill-free week for 1-year. The economic savings for patient out-of pocket expenses from decreased sanitary product usage as a result of nine fewer withdrawal bleeding episodes is offset by the cost of three extra packages of COCPs from the trimonthly regimen. On the basis of an average use of 18 tampons per month, the trimonthly regimen is cost effective when the patient cost per package of pills is less than $9.45. The trimonthly regimen is also cost effective when the sanitary product usage is in the higher range; an above average use of 48 tampons per month is cost effective when the patient cost per package of pills is less than $25.20. Therefore, the trimonthly regimen may be useful for women with menorrhagia, but for the average women, the qualitative benefits of less frequent withdrawal bleeding need to be weighed against an increase in cost. PMID- 10717778 TI - Comparison of efficacy, cycle control, and tolerability of two low-dose oral contraceptives in a multicenter clinical study. AB - This study compares the contraceptive reliability, cycle control, and tolerability of two oral contraceptive preparations containing 20 micrograms of ethinyl estradiol combined with either 75 micrograms of gestodene (EE/GSD) or 150 micrograms of desogestrel (EE/DSG). Women received the trial preparations daily for 21 days, followed by a 7-day pill-free interval. Contraceptive efficacy, cycle control, and tolerability were evaluated over a period of 12 cycles. Efficacy data of 14,700 treatment cycles (EE/GSD: 7299; EE/DSG: 7401) were obtained from 1476 women (EE/GSD, n = 740; EE/DSG, n = 736). Both preparations provided effective contraception and good cycle control with a similarly low incidence of both spotting and breakthrough bleeding. The spotting rates in both treatment groups decreased from 35.1% (EE/GSD) and 37.5% (EE/DSG) in the first treatment cycle to approximately 10% in the fourth treatment cycle. The spotting incidence as percent of the total number of cycles was 12.7% for EE/GSD and 14.3% for EE/DSG. The breakthrough bleeding incidence was 5.2% of all cycles for EE/GSD and 6.0% of all cycles for EE/DSG. For 84.7% of the cycles in the gestodene group and for 82.5% of the cycles in the desogestrel group, neither spotting nor breakthrough bleeding were recorded. Overall, the spotting and breakthrough bleeding incidence tended to be lower with EE/GSD than with EE/DSG. However, the difference was not statistically significant. Amenorrhea was recorded in 2.7% of the cycles with EE/GSD and in 2.9% with EE/DSG. Both preparations were well tolerated and showed a similar pattern of adverse events. More than 83% of the women in both groups either did not gain weight or lost more than 2 kg. Both preparations had a beneficial effect on dysmenorrhea. Both regimens provided reliable contraception and good cycle control. The incidence of adverse events was relatively low and both preparations were well tolerated. PMID- 10717779 TI - Ovulatory potential of preovulatory sized follicles during oral contraceptive treatment. AB - The ovulatory potential of preovulatory follicles was studied in five women taking monophasic gestodene pills containing 20 micrograms of ethinyl estradiol. After one normal pill cycle, follicles were allowed to grow to 16 mm in diameter by deliberate extension of the pill-free period. Once the size of the leading follicle reached 16 mm, the women resumed oral contraceptives for the following 21 days to investigate whether ovulation can be inhibited by late onset of the pill. In addition, 100 micrograms of gonadotropin releasing hormone analog was given intravenously on the third pill day to induce ovulation. Follicular growth and activity were monitored by ultrasonography and by serum concentrations of ethinyl estradiol, progesterone, luteinizing hormone, and follicle stimulating hormone from the last pill day of the first cycle until the end of the second pill intake of 21 days. An increase in luteinizing hormone secretion started before intravenous administration of a gonadotropin releasing hormone analog in all women, eventually leading to ovulation in four of five women. One woman developed an unruptured follicle. Thus, the ovulatory potential of a 16-mm functional follicle cannot be inhibited by reintroduction of pills containing 20 micrograms ethinyl estradiol and 75 micrograms of gestodene. PMID- 10717780 TI - The effects of Implanon on lipid metabolism in comparison with Norplant. AB - This open, randomized study was intended to assess the effects of the new single rod contraceptive implant (Implanon) containing etonogestrel on lipid metabolism in Indonesian women, in comparison with the six-rod implant Norplant, containing levonorgestrel. The effects of both products were compared with a control group using an intrauterine device (IUD) over a 3-year period. A total of 135 healthy volunteers of childbearing potential, aged 22 to 41 years, were enrolled in Jakarta, Indonesia. Ninety volunteers were randomized to use Implanon (n = 45) or Norplant (n = 45), and a nonrandomized group of 45 Multiload Cu 250 IUD users served as a control. Serum concentrations of total cholesterol, triglycerides, HDL cholesterol, LDL cholesterol, apolipoprotein AI, apolipoprotein AII, and apolipoprotein B were measured. The ratios of HDL cholesterol to total cholesterol, of HDL to LDL cholesterol, and of apolipoprotein AI to apolipoprotein B were determined. Lipid and lipoprotein determinations were done at screening and after 3, 6, 12, 18, 24 and 36 months. Contraceptive efficacy and insertion and removal times were also recorded. Mean changes from baseline in the lipid and apolipoprotein parameters, although occasionally statistically significant, were small in all groups (less than 1 standard deviation of the mean concentration at baseline) and clinically not significant. Statistically significant differences between the Implanon and Norplant groups were only occasionally observed. In both implant groups, total mean cholesterol, LDL cholesterol, and apolipoprotein AI concentrations tended to decrease during the study, but statistically significant changes from baseline were only occasionally observed, suggesting that drug-related factors are not involved. The mean ratios of HDL/total cholesterol and the HDL/LDL cholesterol showed very little change over time in both implant groups; slight and statistically nonsignificant increases were noted at most time points. Similar changes were seen in the group of IUD users. It can be concluded that changes from baseline in the lipid and apolipoprotein parameters tested were generally small and did not differ between Implanon and Norplant. PMID- 10717781 TI - Evaluation of prostate-specific antigen as a quantifiable indicator of condom failure in clinical trials. AB - The ability of condoms to retain all elements of semen during intercourse has been assessed by postcoital visual inspection and in vitro permeability studies. Yet, these observations may not be sufficiently precise or realistic. This pilot study evaluated prostate-specific antigen (PSA) as a semen marker of inapparent failure of the condom barrier under conditions of actual use. Twelve couples collected samples from the vagina and surfaces of the condom using sterile cotton swabs. We obtained precoital and postcoital samples for 24 acts of unprotected intercourse, 54 acts of intercourse using intact condoms, and 40 acts of intercourse using condoms that had been deliberately punctured. We used electrophoresis to determine the amount of PSA present in the samples. PSA was detected in 100% (24/24) of vaginal samples collected immediately after unprotected intercourse and in none of the vaginal samples collected more than 24 h after intercourse (0/90). PSA was also present in 98% (83/85) of the samples collected from the inside of the condom that had failed during intercourse. Excluding uses where the condom failed during intercourse, PSA was detected in 2% (1/47) of the postcoital vaginal samples collected after use of intact condoms and in 41% (14/34) of the samples collected after use of condoms with known 1-mm punctures. We conclude that PSA shows great promise as a semen biomarker in clinical trials of barrier methods. We recommend that future studies further investigate the ability of this biomarker to identify condom failures and quantify the extent of semen exposure associated with various types of condom failures. PMID- 10717783 TI - Interceptive effect of Lapachol in rats. AB - Lapachol is a naphtoquinone with therapeutic potential against Chagas disease and is also used as an antimalarial agent. To study the reproductive toxicity potential of Lapachol, pregnant Wistar rats were treated with 0.5 mL of distilled water (control group), 0.5 mL of hydroalcoholic solution (vehicle group), or 20 mg of Lapachol in 0.5 mL of hydroalcoholic solution (treated group) by oral gavage from the 8th to the 12th day of pregnancy. The following variables were observed: maternal body weight on days 1, 6, 15, and 21; food intake on days 2, 6, 15, and 21 of pregnancy. The number of live and dead fetuses and the sites of resorptions were counted. The ovaries were weighed and the corpora lutea were counted. Data were analyzed by ANOVA one-way Dunnett test and chi 2 test. Results showed that mothers were uneffected but there was 100% fetal/embryo mortality, indicative of a strong interceptive effect of Lapachol in rats. PMID- 10717782 TI - Pharmacokinetics of 7 alpha-methyl-19-nortestosterone (MENT) delivery using subdermal implants in healthy men. AB - We studied the pharmacokinetics of 7 alpha-methyl-19-nortestosterone (MENT), a potent synthetic androgen, administered by subdermal implants. The implants contained 112 +/- 4 mg of MENT acetate in a polyethylene vinyl acetate copolymer. MENT acetate released from the implants is rapidly hydrolyzed to MENT in vivo. Fifteen healthy Finnish men were randomized to have either one, two, or four implants inserted in the medial aspect of the upper arm. The implants remained in place for 4 weeks. Blood samples were obtained before implant insertion, 1, 2, 3, and 4 weeks after insertion, and 1 and 2 weeks after removal. Serum MENT concentrations were determined by gas chromatography with mass selective detection. The MENT levels attained in each implant group remained at a steady level during the 4 weeks of implant use. The mean steady state MENT concentrations in the one, two, and four implant groups were 0.6, 1.4, and 2.3 nmol/L, respectively. Serum MENT concentrations during implant use were clearly dose dependent; the between-subject effect of implants as well as the differences between each pair of groups were all statistically significant. The release rate of MENT from one, two, and four implants was calculated to be approximately 0.3, 0.8, and 1.3 mg/day, respectively. This study suggests that MENT acetate implants are a promising method for long-term androgen administration in hypogonadism and male contraception. PMID- 10717784 TI - Mifepristone regulation of leukemia inhibitory factor and uterine receptivity in rabbits. AB - The effects of mifepristone on production of leukemia inhibitory factor (LIF) and uterine receptivity in rabbits was studied. In ovariectomized rabbits, LIF protein was at an undetectable level in control (score = 0), and upregulated by progesterone alone (score = 4). Estrogen had no additive effect, and may even have had a slightly negative effect when the rabbits were treated with both estrogen and progesterone (score = 3). Meanwhile, mifepristone obviously inhibited the stimulation of progesterone on the production of LIF in rabbit uterus (score = 1). The results also showed that LIF protein has a beneficial effect on uterine receptivity and mifepristone prevents this effect. The transfer of embryos to LIF-treated recipients significantly increased pregnancy (70%) and implantation rate (31%) as compared with control (pregnant rate = 50% and implantation rate = 17%). The transfer of embryos to LIF and mifepristone-treated recipients significantly decreased pregnancy (30%) and implantation rate (9%). The results of this study suggest that mifepristone prevented the effects of progesterone on LIF production and the beneficial effect of LIF on uterine receptivity. PMID- 10717785 TI - What do reflex and voluntary mean? Modern views on an ancient debate. AB - Are the words reflex and voluntary useful scientific concepts, or are they prescientific terms that should be discarded? Physiologists use these words routinely in their publications, in laboratory experiments and, indeed, like most lay people, in their daily lives. The tacit assumption is that we all know, more or less, what they mean. However, the issue has a rich history of philosophical and scientific debate; and, as this article demonstrates, present-day researchers still cannot reach a consensus on the meaning of the words and on whether it is possible to draw a scientific distinction between them. The five authors present five quite different analyses. In broad terms, they split into two camps: those who equate voluntary behaviours with consciousness and suppressibility and those who view all behaviours as sensorimotor interactions, the complexity of which determines whether they are reflexive or voluntary. According to the first view, most movements of daily life are neither purely reflex nor purely voluntary. They fall into the middle ground of automatic motor programs. According to the second view, as neuroscience advances the class of reflex behaviours will grow and the class of voluntary behaviours will shrink. PMID- 10717786 TI - Testing quasi-visual neurons in the monkey's frontal eye field with the triple step paradigm. AB - To look successively at sites where several spots of light have appeared in the dark, we cannot simply rely on the image left by these targets on our retina. Our brain has to update target coordinates by taking into account each gaze movement that has taken place. A particular type of brain cell--the quasi-visual (QV) neuron--is assumed to play an important role in this updating by combining target coordinates and eye displacement signals. However, what is exactly this role? Is a QV neuron an element of a working memory that encodes the location of a potential target, or is it pointing to the location of the single goal selected for the next saccade? The two roles theoretically correspond to successive stages of processing: the locations of the optional targets would be stored at one stage, whereas the location of the next selected target would be stored at the subsequent stage. With a task that imposes a choice of goals--the triple-step paradigm--we found evidence that several groups of QV neurons can become simultaneously activated in the monkey's frontal eye field (FEF), suggesting that each group represents a different target location. This supports the hypothesis that the FEF itself contains the spatial information about not yet selected targets. PMID- 10717787 TI - Causes of left-right ball inaccuracy in overarm throws made by cerebellar patients. AB - Cerebellar patients throw inaccurately in the left-right direction but the cause of this multijoint ataxia is unclear. We tested whether it was due, as originally proposed, to variable left-right directions of the hand path, or, alternatively, to variable timing of ball release occurring on a right to left curved hand path. We also examined the cause of the variability in hand path direction per se. Six right-handed cerebellar patients and six control subjects were instructed to throw tennis balls at a slow, medium and fast speed from a seated position while angular positions in 3D of five arm segments were recorded at 1000 Hz with the search-coil technique. Compared to controls, cerebellar patients threw slower and less accurately, had more variable timing of ball release occurring on a right to left curved hand path and had more variable left-right directions of hand paths at a fixed point in front of the sternum. In all cerebellar patients, ball left right inaccuracy was related both to timing of ball release and to hand path direction at the fixed point. The cause of the increased variability in hand path direction varied between patients and could not be explained by disorder in a single joint rotation. No evidence was found that it resulted from variable stabilization at the shoulder during elbow extension. Instead, the more variable left-right direction of the hand path was related to the initial pattern of joint rotations occurring early in the throw before the onset of elbow extension, and to the amplitudes of radioulnar pronation and wrist abduction occurring late in the throw. The results emphasize that in the presence of a cerebellar lesion, ball left-right inaccuracy in overarm throws cannot be explained by a single disorder. Rather ball inaccuracy was likely due to disorders in central commands to proximal joint rotations that produced the hand path and in central commands to distal joints that controlled the timing of finger opening. PMID- 10717788 TI - Classically conditioned withdrawal reflex in cerebellar patients. 1. Impaired conditioned responses. AB - The role of the cerebellum in the classically conditioned, human lower-limb withdrawal reflex was studied in ten patients with pure cerebellar diseases (CBL), ten patients showing additional extracerebellar symptoms (CBL+), and in 11 sex- and age-matched normal controls (CTRL). Where conditioning was successful, the electrically evoked, unconditioned response was preceded by a tone conditioned response (CR). CR incidence was variable, with best results in the CTRL, significantly less in CBL, and lowest in CBL+. Although CRs could be established in subjects in all groups, a continuous increase in the CR incidence in the course of the recording session was observed primarily in CTRL. In CBL and CBL+, such a characteristic reflex acquisition was rather the exception. CR onsets in CBL were within the range of those in CTRL, but CR amplitude was significantly lower in CBL. Cerebellar patients with circumscribed lesions behaved differently in our motor-learning paradigm, depending on the lesion site. Patients suffering from pathology of the posterior inferior cerebellum showed a mean CR incidence within the lower range of CTRL. In contrast, if the anterior and superior cerebellum was affected, few or even no CRs were observed. Our findings thus provide evidence that the human cerebellum is required for the acquisition and the retention of this specific conditioned limb-withdrawal reflex. In particular, anterior and superior parts of the cerebellum appear to be involved. Thus, an expansion of the current concept of clinically based, functional compartmentalization is suggested, such that anterior and superior cerebellar regions must be intact to establish plastic changes required for the acquisition of the conditioned withdrawal response. PMID- 10717789 TI - Classically conditioned withdrawal reflex in cerebellar patients. 2. Impaired unconditioned responses. AB - The study addresses the issue of the role of the cerebellum in human withdrawal reflex conditioning by comparing data from patients with pure cerebellar diseases (CBL, n = 10) and from cerebellar patients showing additional extracerebellar symptoms (CBL+, n = 10) with those from 11 control subjects (CTRL). During recording sessions, the standard delay-conditioning paradigm with paired-trials was used with tone as the conditioned stimulus (CS). Parameters of the conditioned muscle responses are analyzed in an accompanying paper. Here, we focus on the unconditioned muscle response. A train of current pulses (unconditioned stimulus, US) evoked a lower-limb withdrawal reflex (unconditioned response, UR), which was recorded electromyographically from leg muscles. During the recording sessions with CTRL subjects, UR amplitudes decayed from initially 100% to approximately 50% at the end of the session. This type of decay was clearly less pronounced in the CBL group and minimal in the CBL+ group. Furthermore, the CBL group exhibited UR onsets that were delayed by 20 ms compared with those from CTRL subjects. Although the ranges of measurements characterizing the URs of a given cerebellar patient tested in the paired-trial paradigm overlapped with those of control subjects, the statistically significant differences observed at the group level suggest deficits in the performance of the reflex responses. The delayed URs in patients and the different type of decay of UR amplitudes in repetitively evoked withdrawal reflexes constitute evidence that the cerebellum is critically involved in the control of these UR parameters. PMID- 10717790 TI - Initial vestibulo-ocular reflex during transient angular and linear acceleration in human cerebellar dysfunction. AB - During transient, high-acceleration rotation, performance of the normal vestibulo ocular reflex (VOR) depends on viewing distance. With near targets, gain (eye velocity/head velocity) enhancement is manifest almost immediately after ocular rotation begins. Later in the response, VOR gain depends on both head rotation and translation; gain for near targets is decreased for rotation about axes anterior to the otoliths and augmented for rotation about axes posterior to the otoliths. We sought to determine whether subjects with cerebellar dysfunction have impaired modification of the VOR with target distance. Eleven subjects of average age 48 +/- 16 years (mean +/- standard deviation, SD) with cerebellar dysfunction underwent transients of directionally unpredictable whole-body yaw rotation to a peak angular acceleration of 1000 or 2800 degrees/s2 while viewing a target either 15 cm or 500 cm distant. Immediately before onset of head rotation, the lights were extinguished and were relit only after the rotation was completed. The axis of head rotation was varied so that it was located 20 cm behind the eyes, 7 cm behind the eyes (centered between the otoliths), centered between the eyes, or 10 cm anterior to the eyes. Angular eye and head positions were measured with magnetic search coils. The VOR in subjects with cerebellar dysfunction was compared with the response from 12 normal subjects of mean age 25 +/- 4 years. In the period 35-45 ms after onset of 2800 degrees/s2 head rotation, gain was independent of rotational axis. In this period, subjects with cerebellar dysfunction had a mean VOR gain of 0.5 +/- 0.2, significantly lower than the normal range of 1.0 +/- 0.2. During a later period, 125-135 ms after head rotation about an otolith-centered axis, subjects with cerebellar dysfunction had a mean VOR gain of 0.67 +/- 0.46, significantly lower than the value of 1.06 +/- 0.14 in controls. Unlike normal subjects, those with cerebellar dysfunction did not show modification of VOR gain with target distance in the early response and only one subject showed a correct effect of target distance in the later response. The effect of target distance was quantitatively assessed by subtracting gain for a target 500 cm distant from gain for a target 15 cm distant. During the period 35-45 ms after the onset of 2800 degrees/s2 head motion, only two subjects with cerebellar loss demonstrated significant VOR gain enhancement with a near target, and both of these exhibited less than half of the mean enhancement for control subjects. During the later period 125-135 ms after the onset of head rotation, when VOR gain normally depended on both target location and otolith translation, only one subject with cerebellar dysfunction consistently demonstrated gain changes in the normal direction. These findings support a role for the cerebellum in gain modulation of both the canal and otolith VOR in response to changes in distance. The short latency of gain modification suggests that the cerebellum may normally participate in target distance-related modulation of direct VOR pathways in a manner similar to that found in plasticity induced by visual-vestibular mismatch. PMID- 10717791 TI - Inactivation of the ventral premotor cortex biases the laterality of motoric choices. AB - The visual, tactile, and motor properties of neurons in the ventral premotor cortex (PMv) suggest that the PMv plays an important role in the interaction of the face and upper extremities with visual objects, a function that might be disrupted by inactivation of the PMv. The behavior of three rhesus monkeys was, therefore, examined while the PMv was reversibly inactivated by intracortical injection of muscimol. Unilateral PMv inactivation produced no overt deficit in a monkey's ability to reach out and grasp a food morsel with either hand, nor did the monkey have difficulty in extracting a food morsel from a narrow well or in performing a visually cued individuated finger movement task. Unilateral PMv inactivation did bias the laterality of the monkeys' motoric choices, however. When two equivalent food morsels were presented simultaneously to the monkey's right and left, the likelihood that the monkey would make motoric responses contralateral to the inactivated PMv was reduced. After PMv muscimol injections, a monkey was less likely to initially turn its head contralaterally to inspect food morsels, less likely to reach for the food morsel with its contralateral hand, and less likely to take the morsel on its contralateral side. Catch trials in which a food morsel was present only on one side showed that the monkey was aware of the contralateral food morsels and was able to turn its head contralaterally and to use its contralateral arm and hand promptly and accurately. These observations suggest that, when equivalent visual objects for behavioral interaction are present bilaterally, the PMv plays a role in choosing the side to which motoric responses will be directed and the body part that will be deployed as the response effector. PMID- 10717792 TI - Behavior of hypoglossal inspiratory premotor neurons during the carbachol induced, REM sleep-like suppression of upper airway motoneurons. AB - In individuals with compromised upper airway anatomy, genioglossus (GG), the main protruder muscle of the tongue, is an important upper airway dilator which helps prevent upper airway obstructions. During rapid eye movement (REM) sleep, both the tonic and inspiratory-modulated components of GG activity are suppressed in parallel with the characteristic postural atonia. We tested whether the REM sleep related reduction in the respiratory activity of GG may, in part, result from a reduced inspiratory drive relayed to hypoglossal (XII) motoneurons from their premotor medullary inspiratory neurons. In 15 urethane-anesthetized, paralyzed, vagotomized and artificially ventilated rats, we recorded XII nerve activity and the extracellular activity of medullary inspiratory-modulated neurons antidromically activated with latencies of 0.8 ms +/- 0.3(SD) from within (n = 19) or adjacent to (n = 11) the XII nucleus. Carbachol (10-20 nl, 10 mM), a cholinergic agonist, was microinjected into the dorsomedial pons. Such injections trigger a REM sleep-like state in chronically instrumented, intact animals and, in anesthetized rats, produce respiratory and electrocortical changes similar to those of REM sleep. Following the injections, the respiratory component of XII nerve activity was depressed by 51 +/- 22%, while the mean inspiratory firing rate of the neurons decreased by only 7.4 +/- 13.8% (from 69 +/- 34 Hz to 65 +/- 37 Hz; P < 0.02; n = 30). The activity of ventral respiratory group (VRG) and reticular formation inspiratory neurons with axons within the XII nucleus was reduced by 10 +/- 14% (P < 0.005; n = 19), whereas the activity of neurons located near the VRG that had axons passing below the XII nucleus did not change (n = 5). Thus, the depressant effect of carbachol on medullary inspiratory neurons was slightly more pronounced in reticular formation and VRG cells premotor to XII motoneurons than in other medullary inspiratory cells. For all cells, the magnitude of the decrease of cell activity was not related to the magnitude of depression of XII nerve activity, the simultaneously occurring decrease in respiratory rate or the cell's control firing rate. Since the magnitude of this depressant effect on all cell types was disproportionately small when compared with the depression of XII nerve activity, the REM sleep-like decrease in GG activity must be mainly mediated by non-respiratory premotor pathways. PMID- 10717793 TI - Regional alterations in an excitatory amino-acid transporter, blood flow, and glucose metabolism after middle cerebral artery occlusion in the rat. AB - A rise in the extracellular concentration of excitatory amino acids (EAA) plays a pivotal role in ischemic brain injury. EAA concentrations are regulated by uptake mechanisms through high-affinity EAA transporters. Since EAA uptake is energy dependent, it is a matter of interest to explore the relationship between the EAA transporter and derangement of flow-metabolism during ischemia. We examined the regional changes in EAA transporters after permanent occlusion of the middle cerebral artery in rats by in vitro autoradiography using [3H]-D-aspartate as a ligand, and correlated these changes to the local cerebral blood flow (LCBF) and local cerebral glucose metabolism (LCMRglc) determined by in vivo double-labeled autoradiography. The values of specific binding of [3H]-D-aspartate decreased maximally by 20% in the ischemic core. The magnitude of the reduction in specific binding correlated well with the changes in LCBF and LCMRglc. In half of the regions with LCMRglc between 80 and 120% of the intact side, the values of the specific binding were relatively preserved, while in the remainder of the regions in the ischemic hemisphere, with LCMRglc ranging from 40 to 160% of the intact side, there was a reduction in specific binding. These results suggest that energy failure and the related perturbation caused by ischemia can decrease EAA uptake capacity, leading to further deterioration. PMID- 10717794 TI - Potentiating and fatiguing cortical reactions in a voluntary fatigue test of a human hand muscle. AB - Fatigue-associated changes in the excitability of central motor mechanisms were investigated using transcranial magnetic stimulation (TMS) of the motor cortex. Test stimuli were applied before, during and after a voluntary fatigue test of the first dorsal interosseus muscle (FDI). Subjects were required to maintain 50% of their maximum voluntary force (MVC) for at least 2 min (1/2-MVC test) and electromyographic (EMG) reactions of FDI were measured with surface electrodes. Prior to the test, TMS pulses of 70% maximum output (about 1.4 T) produced muscle evoked potentials (MEPs) of widely different amplitudes in different subjects, ranging from 13% to 55% of the maximum compound action potential (M-wave) evoked by ulnar nerve stimulation. During the test, MEPs of all subjects showed a potentiation; this effect was markedly greater in subjects with a small initial MEP. After the test, the differential degrees of contraction-evoked potentiation still influenced the MEP amplitudes; small pre-test MEPs showed a post-test net potentiation and larger pre-test MEPs showed a net post-test depression. The results underline that the net outcome of motor activation on motor cortex excitability, as studied with TMS, depends on a complex balance of fatiguing and potentiating effects. PMID- 10717795 TI - Redundant-signals effects on reaction time, response force, and movement-related potentials in Parkinson's disease. AB - Studies using transcranial magnetic stimulation have established that patients with Parkinson's disease have increased motor cortex excitability. Relying on current evidence that the redundant-signals effect has its source in the motor system, we investigated whether, as a result of cortical hyperexcitability, Parkinson's disease patients demonstrate an enhancement of this effect. Eight patients with moderately severe Parkinson's disease and nine healthy control subjects participated in a task requiring simple manual responses to visual, auditory, and combined auditory-visual signals. During the task, motor cortex activation was recorded by means of movement-related EEG potentials, while responses were measured via isometric force recordings. The movement-related potentials and the force measures both yielded support for the view that the redundant-signals effect is partially caused in the motor system. However, the facilitatory effect of bimodal as compared to unimodal stimulation (i.e. the redundant-signals effect) was of the same size in Parkinson's disease patients and control subjects, as expressed in latency measures of the movement-related potentials and the force signals. We conclude that the redundant-signals effect is not enhanced in Parkinson's disease and that the mechanisms underlying this effect are probably not influenced by the increased motor cortex excitability found in this disease. PMID- 10717796 TI - Control of voluntary and reflexive saccades. AB - The latency of 'reflexive' saccades (made in response to peripheral visual stimuli) was compared to that of 'voluntary' saccades performed in anti-saccade and symbolically cued paradigms. Manipulation of visual events at fixation was carefully controlled across all conditions. Reflexive saccade latency was significantly faster than the latency of all forms of voluntary saccades. Importantly, the latency of saccades made after presentation of a symbolic cue at central fixation (voluntary arrow-cue condition) was greater than that made in the anti-saccade paradigm that requires suppression of a reflexive response. It is suggested that the increase in latency of saccades made in the voluntary arrow cue condition may reflect differences in programming a 'When' trigger signal for saccades made in the absence of a peripheral stimulus. PMID- 10717797 TI - Pantomimed actions may be controlled by the ventral visual stream. AB - Several studies have demonstrated that while perceptual judgements of object size can be biased by visual illusions, actions remain more closely scaled to true object properties. This dissociation is often cited in support of a two-stream model of visual processing, in which visual perception is thought to be mediated by a ventral stream, while goal-directed actions are controlled by a dorsal stream. Evidence suggests that pantomimed actions (i.e., actions directed toward remembered targets) are controlled differently to natural actions; indeed, it has been proposed that pantomimed actions are mediated by the ventral rather than the dorsal stream. To test this hypothesis, we examined the effect of a visual size illusion (a variation of the Muller-Lyer figure) on manual aperture formation during natural and pantomimed prehension (i.e., action) and aperture scaling (i.e., perception). As found in earlier studies, mean peak aperture (MPA) was significantly affected by the illusion in the perception task but not the natural action task. In the pantomime condition, action and perception were equally affected by the illusion as reflected by MPA. These results provide support for the hypothesis that pantomimed actions are mediated by the ventral visual processing stream, while natural actions depend on the dorsal stream. PMID- 10717798 TI - Proceedings of the Annual Scientific Meeting of the Australian and South East Asian Tissue Typing Association (ASEATTA). New Delhi, India, 1998. PMID- 10717799 TI - Unrelated donors selected prospectively by block-matching have superior bone marrow transplant outcome. AB - Previous retrospective studies have demonstrated improved outcome in patients whose donors were matched for non-HLA markers in the MHC as well as for HLA genes. Forty patients receiving transplants from unrelated donors were typed prospectively for HLA and non-HLA markers. Non-HLA markers near HLA-B (beta-block markers) and in the DRB1 introns (delta-block markers) were used to assess MHC match between donors and recipient. Patients whose donors were matched at the beta- and delta-blocks had improved event free survival (63%) compared to patients whose donors were mismatched at one or both blocks (25%) (p < 0.05). Patients whose donors were matched at the beta-block had significantly less severe acute graft versus host disease (p < 0.05). In order to investigate the basis for improved outcome block matching was correlated with HLA matching as determined by DNA sequencing. Beta-block matching was highly correlated with matching for exons 2 and 3 of HLA-B but less so for HLA-C. Delta-block matching was highly correlated with matching for exon 2 of HLA DRB1. It is concluded that matching for non-HLA markers in the MHC improves matching for HLA genes. Further studies are required to determine whether matching for non-HLA markers improves outcome to a greater extent than matching for the HLA genes alone. PMID- 10717800 TI - HLA matching in hematopoietic cell transplantation. AB - Progress in hematopoietic cell transplantation has been greatly facilitated by our increasing knowledge of the HLA system, as well as by improved therapies for achieving sustained engraftment, preventing graft-versus-host disease, and protecting the patient from infection. Disparity for HLA genes can cause graft rejection and graft-versus-host disease and decrease survival in patients receiving grafts from both related and unrelated donors. The presence of patient alloantibodies against donor antigens demonstrated by a positive crossmatch is a strong risk factor for graft rejection. The availability of matched donors for patients lacking a genotypically HLA-matched sibling has been greatly improved by the establishment of international registries of HLA-typed volunteer donors. The development of accurate and reproducible high-resolution DNA-based typing methods has significantly improved the prospects for identifying unrelated donors who are well matched with the patient for HLA. The use of these methods to optimize donor selection will improve both donor identification and the success of unrelated donor transplants. PMID- 10717801 TI - Immunological importance of chimerism in transplantation: new conditioning protocol in BMT and the development of chimeric state. AB - Chimerism is an exceptional immunogenetic state, characterized by the survival and collaboration of cell populations originated from two different individuals. The prerequisits to induce chimerism are immuno-suppression, myeloablation, or severe immunodeficiency of the recipients on the one side and donor originated immuno-hematopoietic cells in the graft on the other. The pathologic or special immunogenetic conditions to establish chimerism are combined with bone marrow transplantation, transfusion, and various kinds of solid organ grafting. Different types of chimerism are known including complete, mixed and mosaic, or split chimerism. There are various methods used to detect the type of chimera state, depending on the immunogenetic differences between the donor and recipient. The induction of complete or mixed chimerism is first determinated by the effect of myeloablative therapy. The chimera state seems to be one of the leading factors to influence the course of the post-transplant period, the frequency and severity of GVHD, and the rate of relapse. However, the most important contribution of the chimeric state is in development of graft versus leukemia effect. A new conditioning protocol (DBM/Ara-C/Cy) for allogeneic BMT in CML patients and its consequence on chimera state and GVL effect is demonstrated. PMID- 10717802 TI - Cord blood serum does not increase lymphocyte responses in comparison to adult serum. AB - To date, over 1000 cord blood (CB) transplants have been reported from different centers worldwide and it is generally agreed that CB represents an encouraging alternative to bone marrow (BM) transplantation. There are a variety of reasons for this, however, possibly the two most controversial aspects are (a) whether there is less graft versus host disease (GVHD) with CB compared to BM transplantation, and (b) whether we can use more HLA mismatches with CB transplantation. The major theory regarding the reduced immunological response of CB lymphocytes is that CB T and NK cells are naive and, therefore, not primed for activation. However, the naive phenomena that has been noted in vitro may be bypassed in vivo by unforeseen factors. We show evidence that there are differences in the soluble factors present in CB and adult serum and that these differences play a role in T cell function. Thus, adult serum will enhance both mitogen and IL-2 specific T cell growth whereas CB serum has no effect, suggesting that there is an activation/growth factor present in adult sera, which is absent in CB sera. This work could enable us to identify the molecular mechanisms which are associated with a lower GVHD in CB compared to BM transplanted individuals. PMID- 10717803 TI - HLA matching in Asian recipients of kidney grafts from unrelated living or cadaveric donors. The Collaborative Transplant Study. AB - The success rate of kidney transplants in Asian recipients was reported in the literature to be exceptionally high. This raises the question whether HLA incompatibilities do not elicit allograft rejection in Asians. Asian recipients, 3,721, of kidney transplants from unrelated living or cadaveric donors were analyzed. The transplants were part of the Collaborative Transplant Study database. HLA typing data and clinical follow-up information were provided by the participating centers. The survival rate of cadaver kidney transplants was associated statistically significantly with the number of HLA-DR mismatches (p < 0.01) as well as with the number of HLA-A + B + DR mismatches (p < 0.01). A significant association of HLA mismatches with graft outcome was also found in a subanalysis of cadaver transplants from Asian donors into Asian recipients (p = 0.03). Among transplants from unrelated living donors, both HLA-DR mismatches and HLA-A + B + DR mismatches were associated with graft outcome (p < 0.01). The success rate of 123 grafts with 2 HLA-DR mismatches was a poor 69 +/- 4% at 3 years, suggesting that such transplants should be avoided if possible. Kidney transplants from unrelated donors into Asians benefit from improved HLA compatibility. The results, therefore, suggest that prospective HLA matching strategies for Asian patients should be pursued. PMID- 10717804 TI - Endogenous and exogenous factors contributing to the surface expression of HLA B27 on mutant APC. AB - We have examined the expression of HLA B*2705 in the mutant cell line 721.220, which lacks endogenous HLA A and B alleles and expresses a defective tapasin molecule. Several peptide sensitive mAbs distinguish between HLA B*2705 expressed on the surface of 721.220 cells (B27.220) and 721.220 cells co-transfected with human tapasin (B27.220.hTsn). This differential staining defines subtle differences in the conformation of HLA B27, which most likely reflect changes in the repertoire of antigenic peptides bound to B27 in the presence and absence of wild type tapasin. HLA B27 molecules expressed on the surface of 721.220 display increased levels of "free" B27 heavy chain (HC-10 staining), an epitope that is dependent on TAP-translocated peptides. The conformation and stability of B27 molecules was examined by investigating the integrity of mAb epitopes and the half-lives of these complexes on cells cultured with and without serum. The decay of surface B27 epitopes occurred more rapidly in B27.220 and this effect was exaggerated in serum free media. Importantly, the decay of surface B27 molecules in B27.220.hTsn cells was characterized by an early increase in HC-10 staining when the cells were grown in serum free media. This decay of B27 molecules via HC 10 reactive intermediates was not observed in B27.220 cells, implying molecules on these cells may already have passed through this stage prior to surface expression. Taken together these observations indicate that tapasin has a significant contribution to the composition and stability of the B27-bound peptide repertoire. PMID- 10717805 TI - HLA-B27 alone rather than B27-related class I haplotypes contributes to ankylosing spondylitis susceptibility. AB - Characterization of non-B27 susceptibility genes will be required to know the pathogenesis of AS. The aim of this study was to examine whether ankylosing spondylitis (AS) susceptibility is controlled by B27 alone rather than B27 haplotypes and, whether other closely related class I loci, such as MICA and TNFA genes might play a role in AS. Three hundred eleven B27-positive samples from Caucasoid, Asian, and African populations were selected and genotypes were carried out by PCR/SSOP (HLA-B27 and HLA-C), PCR/SSP (MICA-TM polymorphism in the transmembrane region), PCR/SSCP (MICA alleles), and PCR-RFLP (TNF-alpha). Of these, 161 were AS patients, chosen in order to investigate the contribution of TNFA and MICA loci to AS in HLA-B27 positive individuals. Some findings can be concluded from the study: (a) No significant differences of TNF-alpha promoter alleles at position -308 and -238 (A/G) were found between AS patients and B27 matched alleles from healthy controls; (b) strong linkage disequilibrium was found between the B27 and the MICA alleles. The MICA-A4 was found to be in association with B*2705,02,03 and 08; MICA-A5 with B*2704 and B*2707 and MICA A.5.1 with B*2706; (c) no significant differences of MICA alleles were found between AS and controls carrying the B27-associated alleles, and therefore no evidence is provided for an additional role of MICA gene in AS susceptibility; (d) there are a striking correlations between the structure of B27 extended haplotypes (from MICA region to HLA-C) and the ethnic distribution of these subtypes. The results of differential linkage disequilibrium with HLA-B27 subtypes suggest that B27 itself remains the primary gene for AS susceptibility, and TNFA and MICA are not involved in the pathogenesis of the disease. PMID- 10717807 TI - HLA-DQ6-mediated protection in IDDM. AB - Insulin-dependent diabetes mellitus is positively associated with DQ8, DQ2, and DQ6 (DQB1*0604), and negatively associated with DQ6 (DQB1*0602), DQ6 (DQB1*0603), and DQ7 in Swedish caucasians. The protection conferred by DQ6 (DQB1*0602) is stronger in younger individuals and there is decrease in the effect of protection with increasing age. Three-dimensional modeling of the susceptible DQ6 (DQB1*0604) and protective DQ6 (DQB1*0602), which share the same DQA chain (DQA1*0102) but differ in the DQB chain at 6 residues, identifies residue 57 and 70 to be important for protection. Three-dimensional models of the DQ8 molecules were constructed from the coordinates of the DR1 crystal structure and other susceptibility and resistance molecules were made by homology modeling. The positively associated DQ molecules had weakly negative to significantly positive surface electrostatic potentials over the peptide binding and T cell recognition areas, whereas the negatively associated molecules had distinctly more negative areas over the relevant surface. This suggests that the variation in the physicochemical properties such as molecular electrostatic potentials among different DQ molecules are important. PMID- 10717806 TI - HLA-B27 transgenic mice are susceptible to collagen-induced arthritis: type II collagen as a potential target in human disease. AB - HLA-B27 is highly linked with a group of human diseases called spondyloarthropathies (SpA). Many of these disorders begin after an infection with an enterobacteria. The symptoms seen in patients with spondyloarthropathies are inflammatory pain in the spine and asymmetrical arthritis of lower limbs. Additional symptoms related to SpA include inflammation in the eyes, bowel, and skin. The autoantigen(s) in SpA are not known. Proteins such as collagen and proteoglycans have been thought to be potent autoantigens in arthritidis including B27-associated human diseases. Type II collagen is a common denominator among eyes and joints, affected tissues in B27-linked diseases. Moreover, a few reports indicated CII specific T cells and antibodies in patients with spondyloarthropathies. We and others have previously described development of spontaneous arthritis and nail disease in HLA-B27 transgenic animals. To determine whether CII may be a target antigen in the B27-linked diseases, B27 + m beta 2 m% (HLA-B27) transgenic mice lacking mouse beta 2m with and without human beta 2m) mice were immunized with type II collagen inside the barrier facility. Male HLA-B27 transgenic mice developed collagen-induced arthritis compared to transgene negative littermates or female counterparts. There was no difference in the incidence of arthritis in HLA-B27 transgenic mice with and without human beta 2m. Our data suggest that beta 2m free heavy chain of HLA-B27 may present soluble antigens such as type II collagen to trigger specific T cells contributing in the development of arthritis. Our data also suggest that CII may be a potential target antigen in the cartilage during the disease process. PMID- 10717808 TI - Worldwide differences in the incidence of insulin autoimmune syndrome (Hirata disease) with respect to the evolution of HLA-DR4 alleles. AB - The relationship between the geographic distribution of susceptibility genes to insulin autoimmune syndrome (IAS) and the incidence of insulin autoimmune syndrome was investigated in order to examine the distribution of the genetic background to susceptibility to certain diseases. The HLA-DR4 allele, DRB1*0406, is associated with increased susceptibility to IAS among Japanese, while the DRB1*0403 and DRB1*0407 alleles are not (the odds ratio of which are 1.6 and 1.1, respectively). The worldwide geographic distribution of the three DR*04 alleles showed that the distribution of DRB1*0403 encompassed that of DRB1*0406 and DRB1*0407. Taken together with the findings that Glu at position 74 in the DRB1 molecule is shared by the three DRB1*04 alleles, there are only a few differences between the DRB1 molecule-nucleotide sequences of DRB1*0403, DRB1*0406 and DRB1*0407, and that all the three DRB1*04 alleles are carried by the same class II haplotype, DQA1*0301/DQB1*0302, it may be considered that DRB1*0403 is the ancestral allele of DRB1*0406 and DRB1*0407. Therefore, populations with a higher prevalence of DRB1*0406 have a higher risk of developing IAS. The extremely low prevalence of IAS among Caucasians can be explained by the low prevalence of DRB1*0406 in this population. This is a good example of the association between the predisposition to risk of development of certain diseases and the evolution of susceptibility genes. PMID- 10717809 TI - HLA class I expression on human cancer cells. Implications for effective immunotherapy. AB - Early studies demonstrated the role of cytotoxic T cells as an immune defence mechanism against tumour cells. The demonstration of tumour antigen peptides and their presentation to T cells on major histocompatibility complex class I molecules highlighted the importance of these molecules in effective anti-tumour responses. It is well established that many tumours escape T cell recognition by loss or down regulation of class I molecule expression on the cell surface of tumour cells. Tumours which have lost class I expression are immunoselected and as a result have a propensity for growth and metastatic spread. With the development of cancer vaccine strategies for clinical use, there will be a future role for histocompatibility laboratories in determining class I expression on tumour cells in individual patients. These studies of expression will require not just the demonstration of total class I expression but the demonstration of locus and allele specific class I molecules involved in the relevant tumour peptide presentation. These studies will be pivotal in tailoring individual patient therapies. The identification of appropriate monoclonal antibody reagents for class I expression and techniques used on different kinds of tissue sections will be a component of the forthcoming 13th International Histocompatibility Workshop. PMID- 10717810 TI - HLA-restricted immune response to mycobacterial antigens: relevance to vaccine design. AB - Identification of mycobacterial antigens that are recognized by CD4+ Th1 cells in HLA-nonrestricted manner or in association with multiple allelic products is required to develop universally effective vaccines against mycobacterial diseases. Our studies in this direction have shown that several recombinant mycobacterial antigens of cytosolic and culture filtrate origin are recognized by CD4+ Th1 cells. Mapping of T cell epitopes with overlapping synthetic peptides covering the entire sequence of these antigens identified peptide sequences stimulatory for Th1 cells. HLA-restriction analysis showed that in addition to HLA-DRB1 products (serologically defined HLA-DR1 to HLA-DR10), the HLA molecules encoded by HLA-DRB3 (HLA-DR52) and HLA-DRB4 (HLA-DR53) are important in presentation of mycobacterial antigens and epitopes to T cells. Depending on the T cell donor, the presentation of a given antigen or peptide could be restricted by HLA-DRB1, HLA-DRB3, and/or HLA-DRB4 products. In addition, stimulation of Th1 cells by some antigens and peptides in the presence of autologous and HLA-DR mismatched allogeneic APC suggested promiscuous presentation. These results taken together suggest that from HLA-restriction perspective, several mycobacterial antigens qualify as candidates for subunit or recombinant vaccine design against mycobacterial diseases. PMID- 10717811 TI - HLA and other host factors in transfusion-acquired HIV-1 infection. AB - The host and viral factors that underlie infection with HIV-1 vary considerably with some individuals progressing to AIDS within 3 to 5 years after infection, whereas others remain clinically asymptomatic for over 10 years. Host factors that may contribute to disease progression include HLA and allelic variants of the chemokine receptors CCR5 and CCR2, which have been shown to influence both long-term survival and rapid progression. In this study, we have examined the contribution of HLA and polymorphisms in CCR5 and CCR2 to long-term survival in transfusion-acquired HIV-1-infected individuals. We have found a higher number of HLA-A32 and -A25 alleles but a lower number of the HLA-B8 allele in the study group compared with the frequencies seen in the HIV-1-negative Australian caucasian population. However, there was no apparent contribution by allelic variants of CCR5 and CCR2 to long-term survival and the combined influence of HLA and CCR polymorphisms could not be evaluated in this relatively small (n = 20) group of study subjects. The results of this work support a role for HLA in long term nonprogression though the presence in the Sydney Blood bank Cohort of nef defective HIV-1 may confound associations between certain HLA alleles and long term survival in the face of infection with HIV-1. PMID- 10717812 TI - Natural variation in immune responsiveness, with special reference to immunodeficiency and promoter polymorphism in class II MHC genes. AB - This review deals with natural selection operating on heterozygotes as a key factor controlling (a) the frequency of immunodeficiencies, and (b) promoter polymorphism in MHC class II genes. The known difference in frequency distribution of X-linked and autosomal deficiencies lend support to this possibility, and suggest that the frequency of neonatal defect may rise as old established equlibria between entry and exit of deleterious mutations change. MHC class II gene promoters differ in their capacity to favor Th1 (or reciprocally Th2) responses, thus suggesting that promoter polymorphism is sustained by the greater flexibility in response that this confers on heterozygotes. PMID- 10717813 TI - Plasma endothelin-1 and mean arterial pressure in the prediction of pre eclampsia. AB - OBJECTIVE: To determine whether increased first trimester plasma endothelin-1 and/or increased midtrimester mean arterial blood pressure detected in pregnant women who are free of symptoms can predict the subsequent development of pre eclampsia. METHOD: Eighty pregnant women were successfully followed from 10 weeks gestation until delivery. Pre-eclampsia and eclampsia developed in 29 and 2 women, respectively, whereas 49 women remained normotensive. Plasma endothelin-1 was determined in the first trimester (10-12 weeks gestation) by a competitive radioimmunoassay. RESULT: First trimester plasma endothelin-1 levels in pregnant women who subsequently developed mild, severe pre-eclampsia and eclampsia were significantly higher than those of pregnant women who remained normotensive. The release of endothelin-1 increases with the severity of the disease, age, body mass index and mean arterial blood pressure. The predictive values of plasma endothelin-1 for pre-eclampsia were: sensitivity 96.8%, specificity 51%, positive predictive value 55.5% and negative predictive value 91%, whereas those of MAP were 48.4, 45, 35.7 and 58%, respectively. CONCLUSION: Determination of first trimester plasma endothelin level may be a valuable marker to identify 55.5% of individuals at high risk of developing pre-eclampsia, if combined with midtrimester MAP, the positive predictive value increases to 68.2%. PMID- 10717814 TI - A population-based cohort study of birth and neonatal outcome in older primipara. AB - OBJECTIVES: To examine the risk of adverse birth outcome in older primiparous women. METHODS: We identified 14,676 primiparae of 20 years of age or more from 1991 to 1996 using the Birth Registry in the North Jutland County, Denmark. We evaluated the risk of adverse birth outcome in the primiparous women aged 30-34 years and above 35 years using the primiparae aged 20-29 years at time of birth as reference. RESULTS: The risks of induced labor, perineotomy, stimulating contraction and vacuum extraction were significantly higher (adjusted odds ratio: 1.3 to 1.7) in the primiparae of 35 years or more. The odds ratio for cesarean section delivery was 2.1 (95% confidence interval: 1.7-2.6) and the odds ratio for delivering a low birth weight child among the primiparae of 35 years or more was 2.2 (95% confidence interval: 1.4-3.3) compared with the primiparae of 20-29 years of age. These risk estimates were independent of women's infertility treatment history. CONCLUSIONS: A negative effect of maternal age on birth and neonatal outcome may be seen even after 30 years of age and is partly related to chronic diseases. However, it is impossible to rule out selection bias, but the actual risk must be taken into consideration in antenatal care. PMID- 10717815 TI - Relationship between duration of preterm premature rupture of membranes and pulmonary maturation. AB - OBJECTIVE: The purpose of this study was to evaluate the relationship between duration of preterm premature rupture of membranes (PPROM) before delivery and development of respiratory distress syndrome (RDS). METHOD: One hundred and fifty nine cases of PPROM with gestational ages between 24 and 37 weeks and birth weights between 1300 and 2100 g were studied retrospectively, out of which, 61 infants developed RDS. Subjects with factors known to affect RDS such as: maternal diabetes; hemorrhage; hypertension; fetal asphyxia; multifetal pregnancies; abnormal presentations; or cesarean deliveries were excluded from this study. RESULTS: Out of 94 neonates, who had rupture of membranes (ROM) in less than 12 h, 41 cases (43.6%) developed RDS. Durations of ROM in 19 neonates were between 12 and 24 h and 6 of them (31.6%) developed RDS. In 21 cases whose ROM were between 24 and 48 h, RDS was seen in 4 (19%). However, in 25 subjects whose ROM was greater than 48 h, RDS presented in 10 cases. Statistical analysis, using Bartholomew's test, showed that there exists a reverse linear relationship between duration of ROM and RDS in the first 48 h. However, after 48 h, the risk of RDS increases, which may represent the effect of complications such as: chorioamnionitis; sepsis; and pulmonary hypoplasia on RDS. CONCLUSION: Increasing the duration of PPROM, in the first 48 h, decreases the risk of RDS with a linear pattern. PMID- 10717816 TI - Delayed luteo-placental shift of progesterone production in IVF pregnancy. AB - OBJECTIVE: To observe absolute and relative levels of progesterone, 17 alpha hydroxyprogesterone (17-OHP) and human chorionic gonadotrophin (hCG) in in vitro fertilization (IVF) pregnancies after withdrawal of luteal support. METHOD: Single blood samples were obtained from 41 pregnant women following IVF treatment and 43 normal pregnant women at various weeks gestation within the first trimester. Progesterone, 17-OHP and hCG were measured by immunoassay. RESULTS: Serum levels of progesterone, but not of hCG, in IVF pregnancies were significantly greater than in normal pregnancies up to 8 weeks post-conception, despite discontinuing luteal support 2 weeks after conception. The ratio of progesterone to 17-OHP, a predominantly ovarian product, in normal pregnancies rose between 4 and 9 weeks but did not change over the same period in IVF pregnancies. CONCLUSION: The luteal contribution to maternal serum levels of progesterone is much higher in IVF pregnancies compared with normal pregnancies. This is sustained throughout the first trimester without the need for luteal support and obscures the placental contribution of progesterone for much longer than in normal pregnancies. Progesterone or hCG supplements may therefore be unnecessary in IVF pregnancy. PMID- 10717817 TI - The urinary incontinence score in the diagnosis of female urinary incontinence. AB - OBJECTIVE: Our purpose was to determine whether the urinary incontinence (UI) score is significantly useful in evaluating the clinical status of UI. METHOD: The questionnaire was administered to 198 UI patients (27-73 years of age) diagnosed by conventional procedures. It consisted of 15 questions, and the answers were assigned points divided into a stress score (s-s) and urge score (u s) according to severity. RESULTS: The patients were classified into a stress incontinence group (SI; 125 cases), urge incontinence group (URI; 29 cases), mixed incontinence group (MI; 41 cases), and overflow incontinence group (3 cases). Classification by questionnaire yielded 110 SI cases, 31 URI cases, and 46 MI cases, accuracy of 83.2%, 86.2%, and 61.0%, respectively. A significant correlation was observed with s-s of SI (r = 0.669, P < 0.001) and u-s of URI (r = 0.583, P < 0.005). CONCLUSION: The UI score will be a simple, clinically effective diagnostic procedure for UI for use by general gynecologists. PMID- 10717818 TI - Incidence of spontaneous abortion in Bahrain before and after the Gulf War of 1991. AB - OBJECTIVES: To determine the incidence of spontaneous abortions in the 5 years before and 5 years after the Gulf War of 1991 and to explore the possible causes that may have affected these changes. To analyze the clinical types, associated medical problems, morbidity, length of hospital stay and mortality rate of abortions. DESIGN: Retrospective study for the period starting on 1 January 1987 31 December 1996. SUBJECTS: The study involved 14,850 cases of abortions admitted into Salmaniya Medical complex during this period. SETTING: The Salmaniya Medical Complex (SMC) is the main referral hospital in Bahrain. METHODS: Analysis of medical records of patients admitted with diagnosis of abortion during this period. RESULTS: By comparing the incidence of abortions in the 5 years before (1 January 1987-31 December 1991) and the 5 years after (1 January 1992-31 December 1996) the Gulf War a significant rise was observed--starting from 1992, reaching a peak in 1994, which then began to decline in 1996. CONCLUSION: Several published reports from Iraq, Kuwait and now from Bahrain are suggestive of an increase in the incidence of abortion and adverse outcome of pregnancy after the Gulf War of 1991. The mechanism is not clear, i.e. whether this is affected by toxicity acquired through the food chain, the oil spillage, smoke pollution resulting from the burning of the Kuwaiti oil fields or stress and anxiety caused by the war. PMID- 10717819 TI - Successful pregnancy outcome with cardiac pacemaker after complete heart block. PMID- 10717820 TI - Screening for gestational diabetes in a high-risk population using fasting plasma glucose. PMID- 10717821 TI - Triple bubble sign: a marker of proximal jejunal atresia. PMID- 10717822 TI - Breast-fed infants, possibly exposed to dioxins in milk, have unexpectedly lower incidence of endometriosis in adult life. PMID- 10717823 TI - Acute pelvic inflammatory disease in a gynecological casualty setting. PMID- 10717824 TI - Actinomycotic tubo-ovarian abscess mimicking advanced ovarian malignancy in a woman with tubal ligation. PMID- 10717825 TI - High serum follicle stimulating hormone (FSH) during perimenopause at high altitude. PMID- 10717826 TI - Maternal mortality and 'near-miss' in rural north India. PMID- 10717827 TI - ACOG practice bulletin. Management of herpes in pregnancy. Number 8 October 1999. Clinical management guidelines for obstetrician-gynecologists. AB - Genital herpes simplex virus (HSV) infection during pregnancy poses a significant risk to the developing fetus and newborn. In the United States, the incidence of this sexually transmitted disease (STD) has increased significantly since 1970 (1). Because many women of childbearing age are infected or are becoming infected, the risk of maternal transmission of this virus to the fetus or newborn is a major health concern. The purpose of this document is to define the stages of herpetic infection, outline the spectrum of maternal and neonatal infection, including rates of transmission and risks, and provide management guidelines that have been validated by appropriately conducted outcome-based research. Additional guidelines based on consensus and expert opinion also are presented to permit a review of most clinical aspects of HSV. PMID- 10717828 TI - ACOG practice bulletin. Antepartum fetal surveillance. Number 9, October 1999 (replaces Technical Bulletin Number 188, January 1994). Clinical management guidelines for obstetrician-gynecologists. AB - The goal of antepartum fetal surveillance is to prevent fetal death. Antepartum fetal surveillance techniques based on assessment of fetal heart rate patterns have been in clinical use for almost three decades. More recently, real-time ultrasonography and Doppler velocimetry have been used to evaluate fetal well being. Antepartum fetal surveillance techniques are now routinely used to assess the risk of fetal death in pregnancies complicated by preexisting maternal conditions (e.g., type 1 diabetes mellitus) as well as those in which complications have developed (e.g., intrauterine growth restriction). This document will review the current indications for and techniques of antepartum fetal surveillance and outline management guidelines for antepartum fetal surveillance, consistent with the best contemporary scientific evidence. PMID- 10717829 TI - Proposal for modification of the TNM staging classification for cancer of the oral cavity. DOSAK. AB - AIM: The prognostic value of the TNM and pTNM classifications currently used for tumours of the oral cavity is unsatisfactory. A better classification should be aimed at as today's definition of T4 leads to overclassification of many tumours and today's definition of N3 results in too few lymph nodes in this group. Until 1987 the grade of fixation of lymph-nodes was part of the N-classification for oral cancer as it is currently used in the N-classification of breast cancer. METHODS: From 1987 to 1991 the DOSAK tumour registry has stored 1532 primary cases of cancer of the oral cavity from 23 hospitals. Crosstables were applied to outline the classification rule for clinical and histopathological T and N based on important factors (T: tumour diameter and thickness; N: lymph node diameter and grade of fixation; pT: histopathological tumour diameter and thickness; pN: number of lymph nodes involved by the tumour). A Cox model was calculated and combinations of similar prognostic estimates were summarized to the same clinical and histopathological T and N. It was aimed at separating categories and achieving equivalent clinical and histopathological T classifications and group frequencies. In a final step a clinical and histopathological stage grouping can be proposed. RESULTS: The gradation of the survival rates shows a marked separation between the T, N and stage categories. The distribution of T, N and stage categories was more uniform when applying the new classification. PMID- 10717830 TI - Technical refinements in surgical treatment of benign parotid tumours. AB - Over the last 15 years, a number of techniques have been developed that make it possible to treat benign parotid tumours surgically with virtually no morphofunctional sequelae. Given that the main objectives of the intervention are the complete removal of the lesion as well as isolation and preservation of the facial nerve and its branches, the authors recommend the following procedure: a face-lifting type of incision in order to disguise the cutaneous scar better; preservation of the posterior branch of the great auricular nerve, in order to maintain the sensitivity of the ear lobe; coverage of the residual defect by means of a flap composed of superficial and deep temporalis fascia in order to reduce the postoperative depression in the parotid region and the onset of Frey's syndrome. PMID- 10717831 TI - Burkitt's lymphoma of the oral cavity in Israel. AB - The clinical presentation of Burkitt's lymphoma in the maxillofacial area is variable. The objective of this study is to review and analyse all cases of Burkitt's lymphoma with oral or maxillofacial involvement diagnosed in our department. A retrospective review of patients with Burkitt's lymphoma in the facial area between the years 1978 and 1997 was undertaken. The patients, 8 male and 5 female, were from 7 to 50 years old (mean 15.3 years). Six patients were Stage I and 7 were Stage II. Five had abdominal involvement and 2 lymph node involvement besides the maxillofacial presentation. Three tumours were in the mandible, 3 in both mandible and maxilla, 2 in the palate, and 5 in the maxilla alone. In 5 patients the tumour presented as a facial swelling, in 3 as an exophytic mass, in 2 as an ulcer, 1 case presented as a hyperplastic lesion, and 2 were periapical lesions. Complaints included pain (7), swelling (5), and sensory disturbance (2). EBV titres were positive in 4 patients. Abdominal involvement was only seen in patients under 12 years old. All patients were treated with chemotherapy, while adjuvant radiotherapy was indicated in 3 cases. Follow-up of 1-20 years revealed a 2-year survival rate of 61.5%. It seems that the Israeli disease is between that of the African and American types, when considering age distribution. PMID- 10717832 TI - Reconstruction of contour and anterior wall defects of frontal bone with a porous polyethylene implant. AB - Frontal bone contour defects cause marked facial deformity, which is instantly obvious to the observer. The aetiology is usually post-traumatic either following a traffic accident or a gunshot injury. The contour deformity of the frontal bone was reconstructed using Medpor porous polyethylene in 12 consecutive patients during a period of 2 years. In four of the patients, we used a coronal approach, whilst using the old incision scar and laceration for access in the remainder. In two of the patients it was not necessary to fix the implant at all, but the remainder were fixed with lag screws because of implant mobility. The aetiology, the technique used, and the results obtained are presented. PMID- 10717833 TI - Intraarterial chemotherapy as neoadjuvant treatment of oral cancer. AB - Neoadjuvant chemotherapy in patients with primary squamous cell carcinomas of the oral cavity should lead to high remission rates whilst having low morbidity. Efficacy can also be enhanced by treating small tumour stages. As part of a multi modality therapy of all stages of primary oral cavity carcinoma, 103 patients were treated with neoadjuvant intraarterial (i.a.) chemotherapy. After regimen A with 100 mg/m2 i.a. cisplatin followed by 5 day continuous intravenous infusion of 5-fluorouracil (1 g/m2 per day) in 36 patients, an i.a. high pressure chemo perfusion with a dose of 150 mg/m2 cisplatin was used with simultaneous intravenous infusion of 9 g/m2 sodium thiosulfate (regimen B, 67 patients). Subsequent treatment comprised radical surgery and simultaneous radiochemotherapy with docetaxel. Partial and complete remissions were found in 80.6% (regimen A) and 67.2% (regimen B) of cases, tumour growth was inhibited in 11.1% and 31.3%. Very low toxicity could be shown especially in regimen B. 66.7% and 74.6% of patients could be operated on radically. Survival rate was 61.1% (regimen A, 22.7 months of mean observation time) and 79.1% (regimen B, 8.4 months). Patients with high-grade remissions seemed to have a survival advantage. Neoadjuvant i.a. chemotherapy with cisplatin, especially in its high dose variant, is a practical therapeutic tool for the treatment of all stages of primary oral cavity carcinoma. PMID- 10717834 TI - Experimental study of bone formation from autogenous periosteal graft following insulin-like growth factor I administration. AB - The present study evaluates the effect of insulin-like growth factor I (IGF-I) administration on bone formation following grafted periosteum. Twenty-four Japanese white rabbits were randomly assigned into one IGF-I administered group (1 mg for 14 days) and a second control group. Grafted periosteum taken from the tibia was placed under the submandibular muscles. At 14, 21, and 28 days following the operation, the grafted periosteum was extirpated and examined. Throughout all stages of the experiment, active bone formation was confirmed histologically and radiographically in both control and experimental groups. In addition, a micro-CT scan was used to observe three-dimensional micro structures in newly formed bone and to measure the trabecular bone thickness as a marker of bone development. As a result, a significant increase in bone formation in the IGF-I group was observed when compared with the findings in the control group. Trabecular bone thickness in the IGF-I group was significantly greater when compared with the control group at 14 days and 21 days following grafting (P < 0.01). At 28 days following grafting, there were no significant differences, suggesting that administration of IGF-I may play an important role in inducing bone formation from grafted periosteum in the early stages. PMID- 10717835 TI - The effect of pentosan polysulphate on bone healing of rat cranial defects. AB - The purpose of the study was to determine the efficacy of pentosan polysulphate, used in combination with guided bone regeneration on rat skull defects. The study was conducted on 45 adult Wistar rats. On each animal two symmetrical 6 mm wide, full-thickness, skull defects were created in the parietal regions. The right defect was chosen as the experimental site and the left one was left empty to provide a control. Each experimental site was covered by an inner and outer polytetrafluoroethylene membrane. The 45 rats were divided into 3 groups: in group I (n = 15), carboxymethyl cellulose, used as a delivery vehicle, was injected between the two membranes; in group II (n = 15), 1 mg of pentosan polysulfate was added to the carboxymethyl cellulose vehicle; in group III (n = 15), purified micronized eggshell powder was added to the mixture of pentosan polysulfate and carboxymethyl cellulose between the two membranes. In each group, the animals were sacrificed at 42 days. The harvested specimens were processed for contact radiography and standard histological examination. The results were assessed by a Fisher's exact test. All animals, except one, healed uneventfully. In group I, partial bone healing was observed in 14 out of 15 animals. In group II, partial bone healing was observed in 13 out of 15 animals, and complete bone healing in 1 out of 15 cases. In group III, partial resorption of the eggshell implant was observed with a partial bone healing in only 2 cases (P < 0.001). In conclusion, significant bone regeneration was observed with the membranes alone. The use of pentosane polysulphate did not result in additional bone gain. The use of particulate material as a space maintainer is also questionable. PMID- 10717836 TI - Surgical treatment of choanal atresia in CHARGE association: case report with long-term follow-up. AB - A patient affected by a multisystem malformation, the CHARGE association, is described. The choanal atresia is one of the stigmata characterizing this syndrome. The main anomalies of the association, the surgical correction (choanoplasty with endoscopic transnasal approach followed by the application of nasal stenting) and a comparison with different techniques are reported. PMID- 10717837 TI - Hydro-jet cutting: a method for selective surgical dissection of nerve tissue. An experimental study on the sciatic nerve of rats. AB - The aim of this study was to answer the question: is it possible to save motor nerves when dissecting tissue with the hydro-jet dissector? In order to study the influence of the hydro-jet on motor nerves the function of the sciatic nerves of 10 Wistar rats was evaluated. The sciatic nerves were dissected bilaterally and only the left one was exposed to the hydro-jet. The water-jet emerged from a nozzle with a diameter of 0.1 mm and was applied to the nerve for 2, 5 or 10 s and with jet pressures of 80, 85 and 90 bar, respectively. After the operation the animals were observed for 5 months in order to monitor the degree of limping using a scale with 10 clinical grades of function. Five months postoperatively the animals were sacrificed and the sciatic nerves were studied by light and electron microscopy. It was found that hydro-jet pressures of 80 bar and exposure times of 2 s had already lead to irreversible damage to the sciatic nerve. Therefore further studies with lower pressures or shorter exposure times are required before considering hydro-jet cutting for parotid gland surgery. It must be confirmed as harmless to motor nerves before applying this method in humans. PMID- 10717838 TI - Effects of sodium fertilizers and supplements on milk production and mammary gland health. AB - A series of experiments was conducted to investigate the effects of sodium fertilizers and supplements on the milk production and mammary health of dairy cows. In Expt 1, where sodium fertilizer was applied to productive pastures consisting mainly of the natrophile perennial rye-grass, the herbage sodium content and the milk yield of cows was increased and milk somatic cell count (SCC) reduced. In Expt 2, which used pastures containing less productive, natrophobic grasses and broad-leaved plants in Estonia, sodium fertilizer did not increase herbage sodium content and did not affect milk production or composition. In Expt 3 the sodium content of the diet of individually tethered cows was increased from 1 to either 6 or 11 g/kg dry matter (DM) by adding salt to their restricted feed allowance, and the cows' milk yield was increased by the high level of sodium supplement and milk SCC were reduced by both levels of sodium supplement. The calcium and magnesium status of cows was improved by the sodium supplement. In Expt 4 a low level of supplementary salt was included in the ration of tethered cows to increase the sodium content of the diet from 2 to 3.6 g/kg DM. No effects on milk yield or SCC were found, but the sodium supplement reduced Staphylococcus aureus contamination of the milk, but not the proportion of milk samples infected with Escherichia coli. It was concluded that the optimum dietary sodium concentration for maximum milk yield was greater than the published requirements, and that substantial increases in sodium intake above current requirements also reduced milk SCC. PMID- 10717839 TI - Effects of morphine and naloxone on the release oxytocin and on milk ejection in dairy cows. AB - The aim of this study was to investigate the action of opioids (the mu receptor agonist morphine) and the antagonist naloxone on inhibition of oxytocin release and milk let-down in response to milking in dairy cows. In the first experiment, cows were injected with 0, 21, 70 and 210 mg morphine 10 min before milking on four successive days. Plasma oxytocin levels after 1 min manual stimulation of the udder were reduced by 70 and 210 mg morphine, and milk let-down was inhibited at the latter dose. In the second experiment, cows were injected after a control milking with 210 mg morphine (or 350 mg at 10 min before milking the following day if not effective) to inhibit milk flow. On the following day the inhibiting dose of morphine was given with 210 mg naloxone. Naloxone injection given before morphine had no effect on plasma oxytocin concentrations, but abolished the inhibition of oxytocin release by morphine and potentiated oxytocin release in response to milking. Naloxone alone injected the day after control milking increased oxytocin levels during milking, suggesting involvement of the opioid system in milking. A model has been developed for the control of opioid effects during milking. Morphine suppressed oxytocin release during milking in a dose dependent manner and the effect was reversible by naloxone. PMID- 10717840 TI - Heterogeneity of cationic amino acid transport systems in mouse mammary gland and their regulation by lactogenic hormones. AB - The mechanism of cationic amino acid transport in lactating mouse mammary gland was investigated. Two Na(+)-independent systems of arginine transport were discriminated on the basis of their sensitivity to leucine. The leucine-sensitive uptake of arginine (Km 0.4 mM) was through a broad specificity system that interacted with both cationic and neutral amino acids, and was inhibited by preloading mammary tissue with neutral amino acids. The leucine-insensitive uptake was identified as the y+ system (Km 0.76 mM). Preloading mammary tissue with cationic amino acids increased the uptake of arginine by the y+ system. Decreasing the pH of the external medium to 6.0 suppressed the y+ system-mediated uptake by approximately 25%, whereas the broad specificity system remained unaffected. Lactogenic hormones upregulated the y+ system-mediated uptake of arginine in pregnant mouse mammary tissue cultured in vitro, although the broad specificity system remained unaffected. The y+ system-mediated uptake increased 2 fold with insulin alone and 4-fold with the combination of insulin, cortisol and prolactin. PMID- 10717841 TI - High-pressure treatment of milk: effects on casein micelle structure and on enzymic coagulation. AB - High isostatic pressures up to 600 MPa were applied to samples of skim milk before addition of rennet and preparation of cheese curds. Electron microscopy revealed the structure of rennet gels produced from pressure-treated milks. These contained dense networks of fine strands, which were continuous over much bigger distances than in gels produced from untreated milk, where the strands were coarser with large interstitial spaces. Alterations in gel network structure gave rise to differences in rheology with much higher values for the storage moduli in the pressure-treated milk gels. The rate of gel formation and the water retention within the gel matrix were also affected by the processing of the milk. Casein micelles were disrupted by pressure and disruption appeared to be complete at treatments of 400 MPa and above. Whey proteins, particularly beta-lactoglobulin, were progressively denatured as increasing pressure was applied, and the denatured beta-lactoglobulin was incorporated into the rennet gels. Pressure treated micelles were coagulated rapidly by rennet, but the presence of denatured beta-lactoglobulin interfered with the secondary aggregation phase and reduced the overall rate of coagulation. Syneresis from the curds was significantly reduced following treatment of the milk at 600 MPa, probably owing to the effects of a finer gel network and increased inclusion of whey protein. Levels of syneresis were more similar to control samples when the milk was treated at 400 MPa or less. PMID- 10717842 TI - Purification of goat beta-lactoglobulin from whey by an ultrafiltration membrane enzymic reactor. AB - This paper presents a novel contribution to the purification of goat beta lactoglobulin by using an ultrafiltration membrane enzymic reactor. The basis of the purification process was the enzymic hydrolysis of contaminating proteins, alpha-lactalbumin and traces of serum albumin, by pepsin at 40 degrees C and pH 2, conditions under which beta-lactoglobulin is resistant to peptic digestion. Simultaneously, beta-lactoglobulin and peptides were separated by ultrafiltration. beta-Lactoglobulin was retained in the reactor while peptides generated by hydrolysis from alpha-lactalbumin and serum albumin permeated through the membrane. The process was made continuous by the addition of fresh whey to replace the lost permeate. Three mineral membranes with 10, 30 and 50 kDa molecular mass cut-off were tested and the 30 kDa membrane was selected for the continuous process. The simultaneous purification and concentration of beta lactoglobulin from clarified goats' whey was achieved in a single step. The ultrafiltration membrane enzymic reactor could treat eight reactor volumes of clarified whey. The recovery of beta-lactoglobulin was 74%, its purity was 84% and its concentration 6.6-fold that in the initial clarified whey. PMID- 10717843 TI - Angiotensin I-converting enzyme inhibitory properties of whey protein digests: concentration and characterization of active peptides. AB - The aim of this study was to identify whey-derived peptides with angiotensin I converting enzyme (ACE) inhibitory activity. The bovine whey proteins alpha lactalbumin and beta-lactoglobulin were hydrolysed with pepsin, trypsin, chymotrypsin, pancreatin, elastase or carboxypeptidase alone and in combination. The total hydrolysates were fractionated in a two step ultrafiltration process, first with a 30 kDa membrane and then with a 1 kDa membrane. Inhibition of ACE was analysed spectrophotometrically. The peptides were isolated by chromatography and identified by mass and sequencing analysis. The most potent inhibitory peptides were synthesized by the 9-fluorenylmethoxycarbonyl solid phase method. Inhibition of ACE was observed after hydrolysis with trypsin alone, and with an enzyme combination containing pepsin, trypsin and chymotrypsin. Whey protein digests gave a 50% inhibition (IC50) of ACE activity at concentration ranges within 345-1733 micrograms/ml. The IC50 values for the 1-30 kDa fractions ranged from 485 to 1134 micrograms/ml and for the < 1 kDa fraction from 109 to 837 mg/ml. Several ACE-inhibitory peptides were isolated from the hydrolysates by reversed-phase chromatography, and the potencies of the purified peptide fractions had IC50 values of 77-1062 microM. The ACE-inhibitory peptides identified were alpha-lactalbumin fractions (50-52), (99-108) and (104-108) and beta-lactoglobulin fractions (22-25), (32-40), (81-83), (94-100), (106-111) and (142-146). PMID- 10717844 TI - Inability of dairy propionibacteria to grow in milk from low inocula. AB - Growth of propionibacteria in complex media was independent of the initial number of cells; in contrast, growth of propionibacteria in milk and whey did not occur if the initial level of cells was < 10(6) cfu/ml. Addition of vitamins, minerals or complex nitrogen sources to the milk or whey, or incubation under anaerobic conditions had no effect on the lack of growth. Addition of freeze-dried whey, prepared from skim milk reconstituted from powder, to a complex medium prevented growth from low inocula in the complex medium, demonstrating the presence of an inhibitor or inhibitors in the whey. The inhibitor(s) was heat stable, had a low molecular mass and retained its activity for at least 4 weeks at 20 degrees C. Pregrowth of some lactic acid bacteria, used as starter cultures in Swiss-type cheese manufacture, in milk for 2 weeks at 20 degrees C removed the inhibition, which explains how propionibacteria develop in Swiss-type cheese from low numbers even though they are inhibited in milk. PMID- 10717845 TI - Physicochemical, molecular and immunological characterization of camel calf rennet: a comparison with buffalo rennet. AB - Camel calf rennet (CCR) and buffalo calf rennet (BCR) were prepared from dried abomasa to study their physicochemical properties and electrophoretic behaviour and to carry out an immunological characterization of the rennet proteins. CCR was more thermostable than BCR. The milk clotting activity of both rennets increased as pH decreased. The optimum temperatures for CCR and BCR were 50 and 45 degrees C respectively. CCR was more sensitive to increased CaCl2 in milk than BCR. Addition of NaCl to milk in the range 0-100 g/l resulted in a marked decrease in the clotting activity of both rennets. When the rennets were treated with acetone, the activity of BCR was completely destroyed, but that of CCR was unaffected. The proteolytic activity of CCR was higher than that of BCR and pepsin towards both camel and cows' milk caseins at pH 6.0. SDS-PAGE electrophoretic patterns of CCR and BCR proteins gave two major bands with molecular masses estimated as 52 and 39 kDa for CCR and 50 and 35 kDa for BCR. Immunodiffusion and immunoelectrophoresis using anti-CCR serum demonstrated immunological cross reactivity between CCR and BCR. PMID- 10717846 TI - Method of quantifying the loss of acidification activity of lactic acid starters during freezing and frozen storage. AB - We have developed a method to quantify the resistance to freezing and frozen storage of lactic acid starters, based on measuring the time necessary to reach the maximum acidification rate in milk (tm) using the Cinac system. Depending on the operating conditions, tm increased during the freezing step and storage. The loss of acidification activity during freezing was quantified by the difference (delta tm) between the tm values of the concentrated cell suspension before and after freezing. During storage at -20 degrees C, linear relationships between tm and the storage time were established. Their slope, k, allowed the quantitation of the decrease in acidification activity during 9-14 weeks of frozen storage. The method was applied to determine the resistance to freezing and frozen storage of four strains of lactic acid bacteria and to quantify the cryoprotective effect of glycerol. PMID- 10717847 TI - Application of capillary electrophoresis to the characterization of processed cheeses. AB - Capillary electrophoresis using a hydrophilically coated capillary and a low pH buffer containing urea has been used to characterize processed cheeses. Different electrophoretic patterns were obtained depending on the ingredients used in the blend such as acid casein, rennet casein, sodium and calcium caseinates and skim milk powder. Isoelectric casein, and sodium and calcium caseinates were shown to contain intact non-glycosylated kappa-casein (kappa-CN), while rennet casein contained only trace amounts of kappa-CN and mainly para-kappa-CN. Therefore, the addition of casein or caseinate to processed cheeses has been detected by analysing the intact non-glycosylated kappa-CN. Quantitation of intact non glycosylated kappa-CN in processed cheeses of known and unknown composition was carried out using a regression curve from standard mixtures of 150-550 g isoelectric casein/kg total rennet casein. This capillary electrophoresis method successfully confirmed the addition of isoelectric casein or caseinate to processed cheeses of known composition. The quantitative determination range was 0.605-3.688 mg kappa-CN/ml. This method cannot be used for measuring additions of rennet casein or any caseinates that have been exposed to chymosin. PMID- 10717848 TI - Ultrasound technique for measuring mammary cistern size of dairy ewes. PMID- 10717849 TI - Expression polymorphism of kappa-casein gene in Holstein cows. PMID- 10717850 TI - Distribution of nitrogen in goats' milk and use of capillary electrophoresis to determine casein fractions. PMID- 10717851 TI - Microbial resistance of caseinate films crosslinked by gamma irradiation. PMID- 10717852 TI - In vitro digestion of caseinophosphopeptide-iron complex. PMID- 10717853 TI - Influence of ions on growth and production of exopolysaccharides by Lactobacillus delbrueckii subsp. bulgaricus NCFB 2772. PMID- 10717854 TI - Acute elbow injuries in the National Football League. AB - We performed a retrospective review to evaluate acute medial collateral ligament injuries of the elbow in professional football players from 1991 to 1996 (5 seasons). There were 5 acute medial collateral ligament injuries in 4 players (1 player with bilateral involvement). All injuries occurred with the hand planted on the playing surface while a valgus or hyperextension force was applied to the elbow. There were 2 centers, both involved with long-snapping situations, 1 running back, and 1 quarterback. All elbows had valgus instability on physical examination. Despite this instability, all players were able to function without operative reconstruction of the medial collateral ligament. No evidence of valgus instability was seen at the time of follow-up (average, 3.4 years). Next, we reviewed all acute elbow injuries in the National Football League from the same 5 season period. Ninety-one acute elbow injuries were reviewed. Overall, there were 70 (76.9%) elbow sprains, 16 (17.6%) dislocation/subluxation patterns, 4 (4.4%) fractures, and 1 (1.1%) miscellaneous injury. Review of the acute elbow sprains revealed 39 (55.7%) hyperextension injuries, 14 (20%) medial collateral ligament injuries, 2 (2.9%) lateral collateral ligament sprains, and 15 (21.4%) nonspecific sprains. The epidemiology of the 14 medial collateral ligament injuries was studied in more detail. The 2 most common mechanisms of injury were blocking at the line of scrimmage (50%) and the application of a valgus force with the hand planted on the playing surface (29%). There were 8 linemen, 4 receivers, 1 running back, and 1 quarterback. All injuries were managed with nonoperative treatment. The average time lost was 0.64 games (range, 0 to 4). We report 19 acute medial collateral ligament injuries of the elbow in elite football players, 2 of whom are considered overhead throwing athletes, who were able to function at a competitive level without surgical repair or reconstruction, in contrast to baseball players, in whom the mechanics and demands may differ. PMID- 10717855 TI - Glenohumeral motion in patients with rotator cuff tears: a comparison of asymptomatic and symptomatic shoulders. AB - The purpose of this study was to determine whether there was a relationship between altered scapular plane glenohumeral kinematics end shoulder pain. Subjects were divided into 3 groups: normal volunteers (n = 10), patients with symptomatic rotator cuff tears severe enough to warrant surgery (n = 10), and subjects with no symptoms who had tears documented on magnetic resonance imaging and normal examination (n = 10). Humeral kinematics were observed with a computer enhanced modification of the Poppen and Walker technique. Scapular plane x-ray films were obtained at 0 degree, 30 degrees, 60 degrees, 90 degrees, 120 degrees, and 150 degrees of elevation. Measurements were made by 3 independent observers blinded to the diagnosis, and data interpretation was performed based on mean values for independent observers. Results showed a high degree of interobserver and intraobserver reliability (coefficients = 0.96 and 0.95, respectively). The symptomatic and asymptomatic groups showed progressive superior translation of the humeral head on the glenoid with increasing arm elevation. The normal group, in contrast, maintained a constant center of rotation along the geometric center of the glenoid. Symptomatic and asymptomatic rotator cuff tear groups showed superior head migration from 30 degrees to 150 degrees, which was significantly different from those seen in the normal group. No significant difference between the symptomatic and asymptomatic groups was demonstrated with the small numbers used in this study. The presence of a rotator cuff tear was associated in a disruption of normal glenohumeral kinematics in the scapular plane. Because significant superior migration of the humeral head was seen in both the asymptomatic and symptomatic rotator cuff groups, painless and normal shoulder motion is possible in the presence of abnormal glenohumeral kinematics. Abnormal glenohumeral kinematics alone was not an independent factor, which could explain the occurrence of symptoms. PMID- 10717856 TI - Shoulder impingement syndrome: preoperative health status. AB - Eighty-one patients with chronic shoulder impingement resistant to conservative treatment completed a generic quality-of-life questionnaire (SF-36) and shoulder specific questionnaire (Simple Shoulder Test [SST]). SF-36 data were compared with those of an Australian normative data set. Patients with chronic shoulder impingement were found to be significantly lower in all health dimensions of the SF-36 than the normal population. Results from the SST test indicated that patients were functionally very limited, particularly in being unable to work full time at their usual job and being unable to lift a weight above the head. Our results indicate that chronic shoulder impingement results in significant functional disability and a reduction in quality of life. Baseline descriptive data of this nature are important, because they provide a point of comparison for the effect of different conditions and for determining the effect of surgical treatment. PMID- 10717857 TI - Frozen shoulder: a 12-month clinical outcome trial. AB - A prospective study was undertaken of 73 patients with frozen shoulder syndrome who were treated with an arthroscopic capsulotomy. All of the patients were assessed for pain, function, and range of motion before surgery and were monitored through to 1-year follow up. Improvement in all parameters was achieved, with pain taking an average of 2.24 weeks to diminish and range of motion improving to within 10% of the other side at an average of 5.5 weeks after surgery. Patients were discharged with a full range of motion and without pain at an average of 8.9 weeks. There was, however, some mild reaggravation of most patients' pain within the postoperative period (mean 4.5 weeks). This pain usually settled with appropriate massage within a 2-week period. In 37% of cases, however, an injection of corticosteroid was required as part of the postoperative management. These cases were usually in that subgroup of patients who still had significant night pain and were in stage 2 or 3 of the disease process at the time of surgery. The postoperative results continued to the 12-month follow-up, with 11% of patients having a recurrence of pain or stiffness. This study has demonstrated that arthroscopic capsulotomy is an effective technique in the management of the frozen shoulder. It also has enabled the authors to document postoperative recovery times, which has given prospective patients realistic time frames of functional expectation in their postoperative recovery. PMID- 10717858 TI - Manipulation under anesthesia for primary frozen shoulder: effect on early recovery and return to activity. AB - Frozen shoulder is still an enigma of shoulder surgery. It is reported that at 2 years from onset, most patients will have recovered whether treated or not. However, the duration of morbidity has major implications for patient function and satisfaction. In view of this fact, we have focused on the early effect of manipulation under anesthesia on shoulder function. We prospectively assessed 39 shoulders in 37 patients who were given the diagnosis of primary frozen shoulder between June 1997 and June 1998 and were treated with manipulation under anesthesia of the affected shoulder. The median preoperative Constant score rose from 24 of 100 to 63 of 100 at 3 to 6 weeks and to 69 of 100 at 3 months. Improvement was maintained at a mean follow-up of 11 months after surgery (range 6 to 18 months). Overall, 94% of patients were satisfied with the procedure. At 3 months 59% (23 shoulders) were rated as having no or mild disability only, 28.2% (11 shoulders) as having a moderate degree of disability, and 12.8% (5 shoulders) as having a severe degree of disability. Of the 5 cases scoring less than 50 of 100 (mean 40) at 3-month follow-up, 1 had unmasked symptoms of a subacromial impingement syndrome that has required further treatment. There was no relationship between the initial Constant score or the initial range of movement after manipulation and the eventual result. We recommend the use of manipulation under anesthetic in primary frozen shoulder to restore early range of movement and to improve early function in this often protracted and frustrating condition. PMID- 10717859 TI - Rotator cuff repair as an outpatient procedure. AB - The purpose of this study was to evaluate outpatient rotator cuff repair on the basis of patient satisfaction, pain control, early postoperative complications, and cost control. Patients were considered good candidates for an outpatient repair if they were in good health and had adequate support at home. Seventy-five rotator cuff repairs were performed on an outpatient basis. The average age of the patients was 58 years. Patients with tears smaller than 2 cm in diameter were excluded. Postoperative pain was managed effectively in 74 of 75 outpatients. There were no cases of deltoid origin compromise, deep infection, or early failure of repair, and no outpatient required readmission to the hospital. This study demonstrates that outpatient rotator cuff repair is possible in the appropriately selected patient and can be performed safely and effectively with a 43% reduction in overall cost. PMID- 10717860 TI - Cause of long thoracic nerve palsy: a possible dynamic fascial sling cause. AB - Long thoracic nerve palsy can result from sudden or repetitive external biomechanical forces. This investigation describes a possible dynamic cause from internal forces. Six fresh cadaveric shoulders (3 female, 3 male, 4 left, 2 right) with full range of motion were systematically dissected to evaluate the anatomic course of the long thoracic nerve. In all specimens a tight fascial band of tissue arose from the inferior aspect of the brachial plexus, extended just superior to the middle scalene muscle insertion on the first rib, and presented a digitation that extended to the proximal aspect of the serratus anterior muscle. With progressive manual abduction and external rotation, the long thoracic nerve was found to "bow-string" across the fascial band. Medial and upward migration of the superior most aspect of the scapula was found to further compress the long thoracic nerve. Previous investigations have reported that nerves tolerate a 10% increase in their resting length before a stretch-induced neuropraxia develops. Previous studies postulated that long thoracic nerve palsy resulted from the tethering effect of the scalenus medius muscle as it actively or passively compressed the nerve; however, similar neuromuscular relationships occur in many other anatomic sites without ill effect. We propose that the cause of long thoracic nerve palsy may be this "bow-stringing" phenomenon of the nerve across this tight fascial band. This condition may be further exacerbated with medial and upward migration of the superior aspect of the scapula as is commonly seen with scapulothoracic dyskinesia and fatigue of the scapular stabilizers. Rehabilitation for long thoracic nerve palsy may therefore benefit from special attention to scapulothoracic muscle stabilization. PMID- 10717861 TI - Anatomy of provocative tests for impingement syndrome of the shoulder. AB - The purpose of this study was to describe the extra- and intra-articular anatomic relationships present during the Neer and Hawkins tests. Nine fresh-frozen cadaveric shoulders were positioned in the impingement position described by Neer (n = 5) or that described by Hawkins (n = 4), embedded in polyurethane, and studied with the use of a cross-sectional technique. All shoulders placed in the Neer position demonstrated soft tissue contact with the medial acromion and contact between the articular surface of the rotator cuff tendons and the anterosuperior glenoid rim. Shoulders placed in the Hawkins position demonstrated consistent contact between soft tissues and the coracoacromial ligament. In all Hawkins positioned shoulders, contact between the articular surface of the rotator cuff tendons and the anterosuperior glenoid was observed. The subscapularis tendon was deformed by the coracoid in 1 of the Hawkins positioned specimens. Although factors inherent to human subjects such as edema and muscle tone may influence the anatomy, these provocative tests for subacromial impingement appear to elicit contact consistent with impingement. PMID- 10717862 TI - Electromyographic analysis of deltoid and rotator cuff function under varying loads and speeds. AB - The purpose of this study was to compare the effect of increasing loads and doubling speed on the deltoid and rotator cuff muscles during isotonic scapular plane abduction (scaption) with neutral humeral rotation. These muscles were studied in 16 volunteers with asymptomatic shoulders with the use of fine wire electromyography. The addition of load to the arm during scaption caused an increase in electromyographic activity during the first 90 degrees of motion. Furthermore electromyographic activity decreased during the final 30 degrees of motion with each increase in load. Doubling the speed caused an increase in electromyographic activity during the first 60 degrees of motion while causing a decrease in activity in the final 60 degrees. This study demonstrates the response of the rotator cuff and deltoid muscles to varying loads and speeds during the most basic shoulder motion. With the data obtained in this study, rehabilitation exercises and experimental shoulder models can be refined to reflect this more physiologic situation. PMID- 10717863 TI - Avulsion fracture of the medial and lateral epicondyles of the humerus. AB - It is still controversial whether fresh avulsion fractures of the medial or lateral epicondyle of the humerus in adults should be treated conservatively or surgically. We monitored 12 patients to consider treatment selection. The patients were examined for site and size of bone fragment, degree of displacement, dislocation of the elbow joint, method of treatment, and bone union and elbow stability. The treatment results were scored according to the Elbow Assessment Score System, and the scores ranged from 86 to 100 points. The scores did not differ significantly among any of the patients, whether treated conservatively or surgically. Although surgery produces good results, conservative treatment can be selected for patients in whom the maximum diameter of bone fragment is 13 mm or less or the displacement of the bone fragment is 9 mm or less. PMID- 10717864 TI - Recurrence of a "primary frozen shoulder": a case report. PMID- 10717865 TI - Subluxation of the annular ligament as a cause of elbow clicking. PMID- 10717866 TI - Bilateral suprascapular nerve entrapment syndrome associated with rotator cuff tear. PMID- 10717868 TI - Dynamic stabilization of winging scapula by direct split pectoralis major transfer: a technical note. AB - A new technique of split pectoralis major tendon transfer (sternal head) for symptomatic scapular winging is shown. Whereas other authors use a lengthening with autogenous grafts, we prefer a direct attachment of the split pectoralis major tendon. With the use of an anterior and a posterior incision, the tendon of the sternal head of the pectoralis major is mobilized and transferred directly to the inferior angle of the scapula. An anatomic study shows that the pectoralis major muscle usually seems to be suitable for this procedure. Direct transosseous fixation of the transferred split pectoralis major tendon appears to be an excellent operation for correcting winging scapula without the necessity of an autogenous graft and concern over stretching or tearing of the graft extension. PMID- 10717867 TI - Subacromial bursitis mimicking a soft tissue tumor. PMID- 10717869 TI - The millennium in medicine. PMID- 10717870 TI - Muddled by the media. PMID- 10717871 TI - Measuring clinical outcome in asthma. PMID- 10717872 TI - The scope of adolescent medicine. PMID- 10717873 TI - The adolescent with disability. PMID- 10717874 TI - Epilepsy and the adolescent. PMID- 10717875 TI - Chronic fatigue in adolescents. PMID- 10717876 TI - The adolescent with diabetes. PMID- 10717877 TI - Malignant disease and the adolescent. PMID- 10717878 TI - Telephone access to health care: the role of NHS direct. PMID- 10717879 TI - Congestive heart failure--can the nephrologist help? PMID- 10717880 TI - Changing concepts of health and disease during the 20th century. AB - Sir Christopher Booth looks at the major medical achievements of the 20th century, from the discovery of vitamins to the eradication of smallpox, and asks where scientific development might lead physicians in the new millennium. PMID- 10717881 TI - The accelerating need for pharmacovigilance. AB - The search for new drugs takes on greater complexity with increasing knowledge, allowing more sophisticated therapeutic interventions. At the same time there is increasing commercial pressure for the pharmaceutical industry to find 'blockbuster' drugs which will be marketed globally to maximise profit in the shortest possible time. Other changes in the industry--shortened times for drug development and increasing outsourcing of functions--make for an environment where some pre-marketing safety issues may go unnoticed. The increasing challenge to pharmacovigilance is not only to be able to find early signals of drug problems, but to rapidly determine the true benefits and risks. We may not have adequate systems to prevent unnecessary harm from globally marketed drugs. PMID- 10717882 TI - National benchmarking as a support system for clinical governance. AB - Audit of the management of acute asthma in hospital has developed in tandem with guidelines produced and updated by the British Thoracic Society (BTS), on the principle that agreed guidelines combined with systematic review of practice by periodic audit are more likely to result in improvements in practice than guidelines alone. A short audit data set was distilled from previous experience with more elaborate tools and made available nationally to audit departments and through letters to consultant members of the BTS. Hospitals have been able to contribute since 1990. The data set reflects key items of the process of care: peak flow measured on admission and twice daily during the hospital stay; blood gases on admission; systemic corticosteroids as an inpatient; discharged with inhaled and oral corticosteroids; written self-management plans; follow-up arrangements. Data from 4,741 admissions over a seven year period are presented. The proportion of patients nationally receiving these items of asthma care is given. The median values for hospital performance improved significantly over the seven years, although there is potential for further improvement. If these data represent the national picture, they could form the basis upon which to set national standards for the care of patients with acute asthma in hospital. A further result of the developing audit has been the recognition of the value of external benchmarking in providing a context for the interpretation of local audit results. This audit system provides hospitals with a quick and easy method of obtaining an overview of local performance, with comparative national data for the same year. This has potential as a tool for clinical governance with much wider applicability, providing the data are handled carefully, particularly as the variability between hospitals diminishes over time. PMID- 10717883 TI - PACES: Practical Assessment of Clinical Examination Skills. The new MRCP(UK) clinical examination. AB - Following an extensive review of the current MRCP(UK) Examination, the Federation of Royal Colleges of Physicians of the United Kingdom has accepted the recommendation of the MRCP(UK) Policy Committee, namely, that the existing MRCP(UK) Part 2 Clinical and Oral Examination be replaced by PACES. This recommendation will not be implemented before June 2001. PMID- 10717884 TI - Diagnosing permanent vegetative state. PMID- 10717885 TI - Organ grafting: from the laboratory to the clinic. PMID- 10717886 TI - The function and fate of neutrophils at the inflamed site: prospects for therapeutic intervention. AB - Neutrophils play a key role in the immediate non-specific immune response, and defects in their function increase host susceptibility to a range of infective agents. However, excess activation and/or delayed clearance of these cells from an inflamed site can lead to significant tissue damage. Neutrophil priming by agents such as endotoxin, granulocyte macrophage colony stimulating factor (GM CSF), platelet activating factor (PAF) and tumour necrosis factor-alpha (TNF alpha) may play a pivotal role in modulating the adhesive and secretory properties of these cells. Priming also appears to affect the survival of neutrophils by delaying constitutive apoptosis. The unique signal transduction events that control neutrophil priming and apoptosis, and particularly the importance of the phospholipase C and phosphoinositide 3-kinase pathways, suggest opportunities for selective pharmacological intervention. PMID- 10717887 TI - The quality of care in hospitals. AB - In recent years there has been an increase in the regulation of the medical profession. In the past there have been problems. The GMC can act only when things go seriously wrong. It has, however, introduced the health and performance procedures, increased the proportion of lay members, is working on revalidation and has introduced Good Medical Practice which makes very clear what is expected of a doctor and will be relevant to doctors' contracts. The medical Royal Colleges can be influential in raising general standards but the activities of the different colleges are not well co-ordinated and they cannot compel doctors to take part in continuing medical education, although this is an aim. Without statutory powers to introduce changes they have to carry their members with them. Audit has its problems and these are understandable because of the natural defensiveness which can occur if there is a threat of possible litigation. The Department of Health has had no proper system for measuring the quality of the care for which it is responsible and largely sees this as the responsibility of individual doctors. Responsibility for the quality of care is shared in a confusing way between different groups. But there is change in the air. There are moves for a 'patient led NHS'. The Government has a new emphasis on quality of care, there is greater sophistication in the methods used for surveying patients' experiences. Measurement of hard outcome data such as adjusted death rates can reveal underlying system failures. Finally, there is a growing realisation that within medicine, as within other complex organisations, doctors are not perfect and will always make errors. Blaming individuals will not in itself make much contribution to the improvement of the overall system: we have to work towards ways of reducing system failures. PMID- 10717888 TI - Stroke services: the good, the bad and the.... AB - The introduction of clinical governance has increased the emphasis placed on measuring the quality of health services. This article addresses the problems associated with assessing the structure, process and outcomes of care of stroke services. The main problem in assessing the structure of stroke services is the difficulty in defining each component and applying criteria which ensure that each component, e.g. stroke unit, stroke physician, is actually in place and meets minimum standards. The structures of stroke services inevitably also vary to meet local needs and conditions. Audits of process and outcome are usually based on aggregated patient data and often include a review of case notes. They are therefore prone to case selection bias, variation in case-mix, measurement error, and random variation. These factors limit the conclusions which can be drawn from most audits. The adverse effects of audit must also be taken into account. It may lead to distortion of health service priorities, divert resources from patient care and encourage individuals responsible for particular services to cheat. Any quality assurance framework should be designed to minimise these effects and to encourage real improvement in the organisation of services and patient care. PMID- 10717889 TI - Stroke. AB - Clearly, progress is being made in the investigation and evaluation of therapies for stroke, and the last two decades have seen stroke medicine come of age. The challenge now is to maintain this momentum with government support, transfer research findings into routine clinical practice and increase public awareness. This conference should have inspired its audience to do so, with clear evidence and good humour. PMID- 10717890 TI - The legacy of Thomas Hodgkin (1798-1866). PMID- 10717891 TI - Careers in academic medicine. PMID- 10717892 TI - GIM and specialty medicine. PMID- 10717893 TI - GIM and specialty medicine. PMID- 10717894 TI - Physicians as educators. PMID- 10717895 TI - How to intervene against smoking. PMID- 10717896 TI - The detection of myocardial viability and hibernation. PMID- 10717897 TI - Reproducibility and repeatability of 99Tcm-HMPAO rCBF SPET in normal subjects at rest using brain atlas matching. AB - The aim of this study was to assess regional cerebral blood flow (rCBF) in normal subjects at rest using 99Tcm-HMPAO single photon emission tomography (SPET). Analysis of reproducibility and repeatability was performed both before and after normalization of flow data. Six healthy volunteers were examined, three times each, according to a routine rCBF protocol. A computerized brain atlas was used to evaluate flow data in eight selected regions. The overall reproducibility of rCBF was evaluated from two scans performed at an average interval of 3 months. Repeatability was evaluated from two scans, 3 h apart and without re-injection of 99Tcm-HMPAO. For the normalized (relative) flow data, the reproducibility was +/- 1.3% and the repeatability +/- 2.2% (i.e. methodological errors dominate). For the non-normalized flow data, the corresponding values were +/- 14.8% and +/- 5.9%. rCBF SPET with 99Tcm-HMPAO is highly reproducible provided that the flow data are normalized. The variation in flow between individuals at one point in time and 3 months later was less than +/- 5% for all brain regions. PMID- 10717898 TI - Influence of diabetes mellitus on regional cerebral glucose metabolism and regional cerebral blood flow. AB - Previous studies have shown both increased and decreased regional cerebral glucose metabolism-blood flow (rMRGlu-rCBF) values in diabetes. We sought to elucidate the influence of diabetes on rMRGlu-rCBF in 57 patients with pure cerebral microangiopathy. Sixteen of 57 patients had diabetes requiring therapy (11 NIDDM, 5 IDDM). Using a special head-holder for exact repositioning, rMRGlu (PET) and rCBF (SPET) were imaged and measured in slices, followed by MRI. White matter and cortex were defined within regions of interest taken topographically from MRI (overlay). Diabetic and non-diabetic microangiopathy patients were compared to 19 age-matched controls. The diabetic patients showed significantly lower rMRGlu-rCBF values in all regions than controls, whereas non-diabetic patients did not. There were no significant NIDDM-IDDM differences. rMRGlu-rCBF did not depend on venous blood glucose levels at the time of the PET examination. However, analysis of variance with the factors diabetes, atrophy and morphological severity of microangiopathy showed that lowered rMRGlu-rCBF in the diabetic group was due to concomitant atrophy only (P < 0.005), while neither diabetes nor microangiopathy had any influence on rMRGlu-rCBF (all P > 0.2). These results were confirmed by multivariate factor analysis. It can thus be concluded that a supposed decrease in rMRGlu-rCBF in diabetes mellitus is in fact only an artefact produced by the concomitant atrophy. All previous studies failed to correct for atrophy, and a critical reappraisal is required. PMID- 10717899 TI - Cerebral blood flow in Sjogren's syndrome using 99Tcm-HMPAO brain SPET. AB - Neuropsychiatric disturbances are frequent in connective tissue diseases. Little is known about the cerebral pathophysiology of Sjogren's syndrome, including cerebral blood flow disturbances. 99Tcm-HMPAO brain SPET was performed in 21 Sjogren's syndrome patients. We also studied 77 patients with systemic lupus erythematosus and 27 healthy individuals. Our results demonstrate the high rate of alterations in cerebral blood flow in Sjogren's syndrome, both psychoneurologically symptomatic and asymptomatic. Focal interhemispherical perfusion deficits were seen in 17 of 21 patients (80.9%) with Sjogren's syndrome: 13/15 symptomatic (86.6%) and 4/6 asymptomatic (66.6%). These changes were mostly localized in the prefrontal and frontal areas, occipital lobes and occipitoparietal area, and less frequently so in the temporal, parietal and central areas. Diffuse hypoperfusion of the frontal lobes (bilateral hypofrontality) was seen in 29% of patients in the Sjogren's group. An acetazolamide stress test was performed in seven patients. There was an increase in perfusion deficits in two patients, no change in two patients, and hypoperfusion decreased in three patients compared with baseline. The results indicate that most Sjogren's syndrome patients experience alterations in cerebral blood flow that are consistent with systemic lupus erythematosus, with heterogeneous reactivity to acetazolamide-induced hypercapnia. These alterations present as focal perfusion deficits and bilateral diffuse hypoperfusion of the lobes. The mechanism of cerebral blood flow alterations is unknown, although it might be the result of diffuse cerebral vasculitis. PMID- 10717900 TI - Changes in the rCBF images of patients with Alzheimer's disease receiving Donepezil therapy. AB - Alzheimer's disease is associated with a loss in presynaptic cholinergic function. It has been suggested that cholinergic inhibitors such as donepezil hydrochloride (Donepezil) could restore this function and improve some of the symptoms of Alzheimer's disease. Previous work has shown that Donepezil improves cognitive and global function in patients with mild to moderate Alzheimer's disease. This study reviewed retrospectively 12 patients who had previously had a 99Tcm-hexamethylpropylene amine oxime (99Tcm-HMPAO) single photon emission tomography (SPET) regional cerebral blood flow (rCBF) examination and had gone on to receive Donepezil therapy. These patients were recalled for a further 99Tcm HMPAO SPET rCBF examination and the image data sets were compared. The results showed an overall increase in global cerebral blood flow (P = 0.04) averaged over the group with a percentage change in blood flow ranging from -1.8% to 6.4%. However, some patients showed a slight decrease in blood flow. When the data were analysed in terms of regional cerebral blood flow, we found that the most significant increase in blood flow occurred in the frontal lobes (P = 0.02). PMID- 10717901 TI - A comparison of 99Tcm-MIBI myocardial SPET and electron beam computed tomography in the assessment of coronary artery disease in two different age groups. AB - The aim of this study was to compare the clinical value of 99Tcm-MIBI single photon emission tomography (SPET) and electron beam computed tomography (EBCT) in the assessment of coronary artery disease (CAD) in different age groups. 99Tcm MIBI SPET (stress-rest), EBCT and coronary angiography studies were performed in 64 consecutive patients with suspected CAD. The patients were classified into two groups: Group A = 40 patients > 45 years of age and Group B = 24 patients < or = 45 years of age. There were 31 and 14 patients with coronary stenosis > or = 50% as determined by coronary angiography in Groups A and B, respectively. All patients (30 cases) with abnormal 99Tcm-MIBI myocardial SPET and coronary calcification detected by EBCT had significant coronary artery disease, and 93.3% of the patients with normal 99Tcm-MIBI SPET and normal EBCT had normal coronary angiography or < 50% lumen narrowing of the coronary arteries. In Group B, the sensitivity of SPET for detecting CAD was significantly higher than that of EBCT (92.9 vs 42.9%, P < 0.01); the specificity of SPET was comparable to that of EBCT. In Group A, there was no significant difference between SPET and EBCT in terms of sensitivity (93.6 vs 90.3%) or specificity (88.9 vs 55.6%). However, in the detection of individual coronary artery disease, the specificity of SPET was significantly higher than that of EBCT in Group A (94.1 vs 66.7%, P < 0.001). The sensitivity of SPET was again significantly higher than that of EBCT (85.7 vs 38.1%, P < 0.005) in Group B. The accuracy of SPET was higher than that of EBCT in both groups (82.5 vs 67.5%, P < 0.01 in Group A; 93.1 vs 76.4%, P < 0.01 in Group B, respectively). We conclude that 99Tcm-MIBI myocardial perfusion SPET has a higher sensitivity than EBCT in the detection of CAD in patients < or = 45 years old and a higher specificity in patients > 45 years of age. A combination of SPET and EBCT may assess CAD more accurately. PMID- 10717902 TI - Effect of trimetazidine on 99Tcm-tetrofosmin uptake in patients with coronary artery disease. AB - Myocardial uptake of 99Tcm-tetrofosmin in vivo is determined by a combination of flow and metabolic status of myocytes. The accumulation of tetrofosmin in the mitochondria is related to their ability to transduce metabolic energy into electronegative membrane potential. Trimetazidine (TMZ), an anti-ischaemic drug, appears to have a metabolic cytoprotective effect related to mitochondrial function, since it does not induce systemic or coronary haemodynamic changes. In this study, we evaluated the effects of TMZ on tetrofosmin uptake in hypoperfused myocardial regions in patients with coronary artery disease (CAD). Twenty-two patients, 14 with previous myocardial infarction (group A) and eight with a history of angina (group B), with angiographically documented CAD were studied. All patients underwent two tetrofosmin SPET studies at rest, before (baseline) and 1 week after TMZ administration (post-TMZ). On quantitative analysis, 131 segments showed less tetrofosmin uptake at baseline. In these segments, tetrofosmin uptake was 51 +/- 13% at baseline and 55 +/- 15% post-TMZ (P < 0.001 vs control). In the 86 hypoperfused segments of group A, tetrofosmin uptake was 48 +/- 14% at baseline and 52 +/- 17% post-TMZ (P < 0.001 vs control). In the 45 hypoperfused segments of group B, tetrofosmin uptake was 56 +/- 9% at baseline and 60 +/- 10% post-TMZ (P < 0.001 vs control). In the remaining 309 segments, no significant difference in tetrofosmin uptake before and after TMZ was observed. In conclusion, our results suggest that TMZ administration may increase myocardial uptake of tetrofosmin in hypoperfused regions at rest in patients with CAD, based on its metabolic effect. PMID- 10717903 TI - The effect of respiration on left ventricular diastolic filling as assessed by radionuclide ventriculography. AB - Left ventricular function is modified by respiration and pericardial constraint. The aim of this study was to compare left ventricular systolic and diastolic function during inspiration and expiration in four patient groups: patients (1) without cardiac disease, (2) with severe pulmonary disease, (3) with cardiac amyloid and (4) with pericardial constriction (before and after pericardiectomy). Using blood-pool left ventriculography with modified gating, we obtained time activity curves at the onset of inspiration and expiration. On inspiration and expiration, patients with pericardial constriction and patients with cardiac amyloid were significantly different from those without cardiac disease and those with severe pulmonary disease, in that left ventricular ejection fraction (LVEF) was less, peak filling rate was greater, time to peak filling rate was shorter, and rapid filling fraction was increased. When inspiration and expiration were compared, time to left ventricular peak filling rate was shorter (P = 0.05) on inspiration (118 +/- 48 ms) than on expiration (168 +/- 35 ms) in patients with pericardial constriction. No other measures differed between inspiration and expiration in pericardial constriction, and left ventricular function was unaffected by respiration in the other groups. Time to left ventricular peak filling rate was 49 +/- 69 ms less on inspiration than on expiration in pericardial constriction and this difference was significantly different (P = 0.04) from that in patients with cardiac amyloid (34 +/- 58 ms greater), patients without cardiac disease (2 +/- 69 ms greater) and patients with severe pulmonary disease (19 +/- 63 ms less). In pericardial constriction, pericardial resection caused an increase in LVEF without a change in left ventricular diastolic filling but abolished the differences present between inspiration and expiration in time to left ventricular peak filling rate. This respiratory response in time to left ventricular peak filling rate may be valuable in the diagnosis of pericardial constriction. PMID- 10717904 TI - Simplified algorithms for the estimation of 99Tcm-MAG3 clearance. AB - The aim of this study was to evaluate two formulae allowing the determination of MAG3 clearance by means of a single blood sample, namely Bubeck's formula and Russell's formula. As a first step, a simulation study was performed with the two single-sample algorithms to predict MAG3 clearance as a function of plasma concentration, using various times for blood sampling and various body surface areas. As a second step, a validation study on 47 adult patients with varying renal function allowed a clinical comparison between the reference technique, namely the multiple blood sample technique, and the two simplified techniques. The simplified algorithms were calculated using the fitted value at 44 min. In the simulation study, whatever the time of blood sampling or the level of correction introduced for body surface area, the results obtained by means of Bubeck's algorithm diverged significantly from those of Russell's algorithm, for low clearance values as well as for high clearance values. The curve of the differences between the two methods had a typical boomerang shape. In the clinical study, the difference between Russell's algorithm and the reference method was generally within 20 ml.min-1, with no systematic bias; with Bubeck's algorithm there was a marked underestimation, both in the low and high clearance ranges. We suggest Russell's single-sample method is the method of choice. PMID- 10717905 TI - 99Tcm-MAG3: problems with radiochemical purity testing. AB - The radiochemical purity (RCP) of 99Tcm-MAG3 was determined using solid-phase extraction (SPE), high-performance liquid chromatography (HPLC) and thin-layer chromatography (TLC). The difference between the HPLC and SPE methods was highly significant (P < 0.001), yielding values for RCP of 94.4 +/- 1.4% and 86.0 +/- 5.1% [corrected] respectively (mean +/- s). Further qualitative analysis of the SPE fractions obtained, was carried out using HPLC and TLC. The unexpected presence of 99Tcm-MAG3 in one of the fractions was observed together with the appearance of hydrophilic impurities in the hydrophobic extract. This lack of specificity may be the reason for the discrepancy between the SPE and HPLC methods. Use of the SPE method leads to an underestimation of the RCP of 99Tcm MAG3 and, indeed, had we been relied solely on this method of analysis, we would have had to reject most kits we prepared. In a separate study, we compared a TLC method with HPLC. Differences were found to be highly significant (P < 0.001), yielding values of 98.3 +/- 0.6% and 95.8 +/- 0.9% respectively. Comparison of the data points showed that TLC gave consistently higher RCP yield than HPLC. This elevated value was found to be due to the inability of the TLC method to separate 99Tcm-lipophilic impurity, seen on HPLC, from the 99Tcm-MAG3. Therefore, use of this TLC method leads to an overestimation of the RCP of 99Tcm-MAG3. PMID- 10717906 TI - Contribution of 99Tcm-DMSA scintigraphy to aetiological diagnosis in renal transplant recipients with impaired renal function. AB - Routine 99Tcm-dimercaptosuccinic acid (DMSA) scintigraphy was performed in a series of 24 kidney transplant recipients with impaired renal function. Diagnostic findings on planar and tomoscintigraphic acquisitions obtained 3 and 4 h after the injection of 130-140 MBq 99Tcm-DMSA were compared with the diagnosis established by fine-needle biopsy in 13 patients and by clinical course and other examinations (ultrasonography, bacteriology) in 11 patients. Renal scintigraphy demonstrated segmental defects in patients with rejection (n = 2/6), immunosuppressor nephrotoxicity (n = 2/6), acute pyelonephritis (n = 3/3), renal artery stenosis (n = 1/1) and obstructive lymphocele (n = 1/1). Diffuse lack of uptake was observed in one patient with severe renal failure. The scintigram was normal in 14 patients, including three with lesions histologically compatible with graft rejection or immunosuppressor nephrotoxicity. 99Tcm-DMSA was thus found to contribute little to the differential diagnosis between graft rejection and immunosuppressor nephrotoxicity. However, it may be useful for identifying specific disease states, particularly acute pyelonephritis, seen as well delimited systematized defects. 99Tcm-DMSA scintigraphy could also be used in late follow-up after pyelonephritis in renal transplant recipients. PMID- 10717907 TI - Evaluation of skeletal muscle metabolism and response to erythropoietin treatment in patients with chronic renal failure using 99Tcm-sestamibi leg scintigraphy. AB - It is well known that uraemia affects skeletal muscle metabolism. This has been attributed to a variety of causes, including anaemia, vitamin D, carnitine deficiency and hyperparathyroidism. The aim of this study was to ascertain whether 99Tcm-sestamibi leg scintigraphy is useful in the evaluation of skeletal muscle metabolism and the monitoring of treatment response in uraemic myopathy. Forty patients with chronic renal failure and 24 normal controls underwent examination. Fifteen patients with chronic renal failure received erythropoietin treatment. 99Tcm-sestamibi leg scintigraphy was performed in all subjects and in 15 patients after therapy. The calf-to-ankle uptake ratio was calculated by semi quantitative analysis and normalized to lean body mass. The normalized uptake ratios were significantly different between patients and controls. After erythropoietin therapy, there was a significant increase in the normalized uptake ratios compared with pre-therapy. Our results suggest that 99Tcm-sestamibi leg scintigraphy is useful in the assessment of muscle metabolic abnormalities and the effect of treatment in uraemic myopathy. PMID- 10717908 TI - Pre-targeted radioimmunotherapy of human colon cancer xenografts in athymic mice using streptavidin-CC49 monoclonal antibody and 90Y-DOTA-biotin. AB - Radioimmunotherapy of solid malignancies using directly labelled 90Y-monoclonal antibodies has been associated in clinical trials with dose-limiting bone marrow toxicity. Our objective in this study was to evaluate the efficacy and toxicity of an alternative two-step pre-targeted radioimmunotherapy protocol for the treatment of human colon cancer. The two-step protocol consisted of administration of the tumour-associated glycoprotein (TAG-72) monoclonal antibody CC49 conjugated to streptavidin, followed by administration of 90Y-DOTA-biotin. Swiss nu/nu athymic mice bearing subcutaneous LS174T human colon cancer xenografts were injected intraperitoneally with streptavidin-CC49 (250 micrograms), followed 40 h later by an intravenous injection of 90Y-DOTA-biotin (900 microCi, 40 micrograms). Tumour growth was measured over 25 days and compared with that in control mice receiving no treatment. Bone marrow and normal tissue toxicity was determined by peripheral blood leucocyte counts and by monitoring the body weight of the animals. Pre-targeted radioimmunotherapy resulted in a modest (30-40%) decrease in the mean tumour growth rate in treated mice compared to control animals. There was no change in body weight following treatment and peripheral blood leucocyte counts remained within the normal range. Pre-targeted radioimmunotherapy was safe at administered amounts of 90Y radioactivity, which were at least nine-fold higher than those previously determined to be lethal using directly labelled 90Y-monoclonal antibodies. The results of this study are promising for the application of pre-targeted radioimmunotherapy using streptavidin-CC49 and 90Y-DOTA-biotin for the treatment of advanced colorectal cancer in humans. PMID- 10717909 TI - Toxicity of high-activity 111In-Octreotide therapy in patients with disseminated neuroendocrine tumours. AB - Disseminated neuroendocrine tumours are difficult to treat and are generally not responsive to radiotherapy or chemotherapy. Nuclear medicine techniques using a radiolabelled somatostatin analogue, 111In-Octreotide, have been used for the diagnosis of neuroendocrine tumours. It has been suggested that high activities of such an agent may have a therapeutic effect. The aims of this study were to assess toxicity and to determine if there had been evidence of efficacy. Eight patients with known disseminated neuroendocrine tumours were enrolled in the study; six had carcinoid tumours, one had a medullary cell carcinoma of the thyroid and one patient had a malignant gastrinoma. Between 1.3 and 4.6 GBq of 111In-Octreotide were administered to each patient for up to five administrations over 12 months. A total of 23 administrations were given. Tests of vital signs, renal, liver and endocrine function as well as haematological markers were taken before and after treatment. The treatment was well tolerated with only one patient suffering from a sensation of flushing during the infusion but no changes in vital sings. There was a transient (up to 48 h) drop in circulating lymphocytes in four patients and platelets in two patients; no supportive therapy was needed. One patient with severe renal impairment had a slight reduction in glomerular filtration rate. We conclude that high-activity 111In-Octreotide is well tolerated with low toxicity and can be considered for use in patients with disseminated neuroendocrine tumours. Further work is now being performed to assess efficacy. PMID- 10717910 TI - Usefulness of 99Tcm-MDP knee SPET for pre-arthroscopic evaluation of patients with internal derangements of the knee. AB - The aim of this study was to ascertain whether knee SPET can localize lesion sites in patients with internal derangements of the knee. We performed knee SPET as a pre-arthroscopic examination in 63 consecutive patients. SPET imaging was performed with a triple-headed SPET camera 4 h after the injection of 99Tcm methylene diphosphonate. Arthroscopic diagnoses were as follows: 28 medial meniscus injuries, 24 lateral meniscus injuries, 31 anterior cruciate ligament (ACL) injuries, three posterior cruciate ligament injuries and one medial collateral ligament injury. Of 30 patients with crescent-shaped increased activity at the medial tibial plateau, 22 had medial meniscus injuries (positive predictive value: PPV 73%); of 17 patients with crescent-shaped activity at the lateral tibial plateau, 13 had lateral meniscus injuries (PPV 76%). Of 18 patients with increased activity at ACL attachment sites (primary sign), 17 had ACL injuries (PPV 94%). Of 27 patients with increased activity at bone impaction sites of ACL injury (secondary sign), 22 had ACL injuries (PPV 81%). Of 32 patients who had either a primary or secondary sign, 26 had ACL injuries (PPV 81%). We conclude that knee SPET is very useful in the management of internal derangements of the knee, particularly in determining the need for arthroscopy by localizing lesion sites. PMID- 10717912 TI - Body surface area correction in single-sample methods of glomerular filtration rate estimation. PMID- 10717913 TI - [Gastroenterology in the past, at present and in future]. PMID- 10717911 TI - 67Ga-citrate and 99Tcm-MDP for estimating the severity of vertebral osteomyelitis. AB - The aim of this study was to evaluate the roles of 67Ga-citrate and 99Tcm methylene diphosphonate (99Tcm-MDP) planar and single photon emission tomographic (SPET) imaging in patients with vertebral osteomyelitis. Thirty patients (22 females, 8 males) aged 62.7 +/- 16.4 years (mean +/- s) were enrolled prospectively between May 1995 and May 1998. The patients had been on antibiotics for 7 +/- 4 weeks prior to the study. Histology was available for all but nine patients with mild infections, who were evaluated by a combination of magnetic resonance imaging (MRI), clinical and laboratory tests. 67Ga-citrate (185 MBq) and three-phase bone (555 MBq 99Tcm-MDP) planar and SPET imaging were performed in all patients, together with MRI as a comparison. In total, 67 infectious foci were detected. Based on histology, there were four cases of severe, 13 cases of moderate and four cases of mild osteomyelitis; nine mild infections were also classified by the combination of MRI, clinical and laboratory results. Combined MRI and 67Ga-citrate SPET correctly classified all patients; MRI detected all 67 infectious foci, whereas 67Ga-citrate SPET identified 54 only. False-negative results were seen with all other modalities, especially in cases of mild and moderate infection. 67Ga-citrate SPET identified unsuspected cases of endocarditis (n = 2), paravertebral abscess (n = 1), subaxillary soft tissue abscess (n = 1) and rib osteomyelitis (n = 1). For 67Ga-citrate SPET, the target to-background ratio was 2.24 +/- 0.31, 1.76 +/- 0.07 and 1.30 +/- 0.18 for severe, moderate and mild osteomyelitis, respectively. Significant differences were noted between severe and moderate infection (P = 0.0051) and between severe and mild infection (P < 0.0001); that between moderate and mild infection was non significant. For 99Tcm-MDP planar and SPET imaging, and for planar 67Ga-citrate imaging, there was no correlation with severity. We conclude that 67Ga-citrate SPET is able to identify vertebral osteomyelitis and detect additional sites of infection. It can also aid in determining the severity of infection and, potentially, the response to therapy. PMID- 10717914 TI - [24-hour monitoring of gastric juice pH in assessment of anti-ulcer treatment]. AB - AIM: Comparison of various schemes of three-component eradication of duodenal ulcer (DU) using 24-h monitoring of gastric juice pH. MATERIALS AND METHODS: 62 DU patients in aggravation period have undergone 24-h monitoring of gastric juice pH and esophagogastroduodenoscopy with biopsy and H.pylori test of the treatment beginning and end. Patients of group 1 (n = 28) received ranitidin, metronidazole and amoxicillin while patients of group 2 (n = 34) took quamatel, metronidazole and amoxicillin. RESULTS: 24-h monitoring of pH of the gastric juice demonstrated that three-component therapy consisting of quamatel (20 mg/day, metronidazole (500 mg 4 times a day), amoxicillin (750 mg 3 times a day) maintains the level of intragastric acidity optimal for quick scarring of duodenal ulcer defect and led to eradication of H.pylori in 91.2% of cases. CONCLUSION: 24-h monitoring of gastric juice pH is a significant method of assessment of antiulcer treatment efficacy. PMID- 10717915 TI - [Long-term results of gastric ulcers chemotherapy]. AB - AIM: Determination of criteria of choice of gastric ulcer (GU) treatment. MATERIALS AND METHODS: 904 GU patients given conservative drug therapy in 1986 1996 were followed up. RESULTS: Recurrences of GU can be of three types: type 1- rare recurrences (once in 10-11 years), type 2--recurrences occur twice for 10-11 years), type 3--recurrences occur every 1.5 years). CONCLUSION: A simple criterion of assessing effectiveness of conservative treatment of uncomplicated ulcers is proposed. It is duration of remission. If the recurrence occurs within 1.5 years after the treatment, the patient is at risk of frequent recurrence. In such cases surgical treatment is justified. PMID- 10717916 TI - [Complaints as reflection of psychic status in patients with duodenal ulcer exacerbation]. AB - AIM: Study of relationship between leading complaints of patients with duodenal ulcer (DU) exacerbation and their psychic status, attitude to the disease and quality of life. MATERIALS AND METHODS: Psychological questionnaire surgery enrolled 94 males with endoscopically revified uncomplicated recurrence of DU. RESULTS: Patients complaining of dull epigastric pains only demonstrated minimal psychological changes. Their attitude to the disease is primarily ergopathic, sensitive and anosognosic. Nausea and vomiting complains come along with neurotization, maladaptation (anxiety, neurasthenia, egocentric attitude) and deterioration of quality of life. CONCLUSION: Patients with nausea and vomiting complaints need correction of psychic status to achieve psychological compensation in remission of DU. PMID- 10717917 TI - [Chronic hepatitis: indicators of disease activity]. AB - AIM: To assess correlations between activity of chronic hepatitis (CH) by morphological data on the liver and alaninaminotransferase (AlAT) serum levels in patients with viral and alcoholic liver lesions. MATERIALS AND METHODS: Serological, biochemical blood tests and histology of hepatic biopsy were made in 47 patients with chronic hepatitis. RESULTS: The Knodell index characterizing the activity of chronic hepatitis by morphological evidence correlated with AlAT activity in the serum. Normal values may be associated with 1 to 5 scores by the Knodell index. AsAT/AlAT was higher in patients with CH free of virus markers. CONCLUSION: Morphological data (Knodell's index), AlAT activity and AsAT/AlAT indicate CH activity. AsAT/AlAT may help in elucidation of the presence or absence of alcohol abuse. PMID- 10717918 TI - [Hepatobiliary alternations in leptospirosis convalescents]. AB - AIM: To study hepatic function in leptospirosis convalescents. MATERIALS AND METHODS: Clinical laboratory and device examinations were performed in 121 leptospirosis convalescents. RESULTS: The majority of the examinees retained biochemical signs of cholestatic, mesenchymal-inflammatory and moderate cytolytic syndromes secondary to hepatic impairment. Ultrasound and radionuclide investigations detected biliary disorders and secondary chronic hepatitis in many of leptospirosis convalescents. CONCLUSION: It is shown that leptospirosis rehabilitation must be targeted. Reasons of development of chronic hepatic and biliary lesions in population of the north Caucus and Kuban in the absence of viral hepatitis markers are suggested. PMID- 10717919 TI - [Prevalence of and risk factors for gallstones in female population of Novosibirsk]. AB - AIM: To investigate the prevalence of gallstones and associated factors in female population of Novosibirsk (Western Siberia). MATERIALS AND METHODS: A representative sample of 870 women aged 25-64 years was drawn from general population according to WHO "MONICA" protocol. The subjects were screened for the presence of gallstones by gallbladder ultrasonography, completed a questionnaire relating to food and alcohol consumption, smoking, gastrointestinal symptoms and obstetric history. They also underwent physical examination and blood chemistry tests. Age-adjusted prevalence of cholelithiasis was 9.5%. Increasing age, obesity, diabetes mellitus, consumption of animal fat, pregnancies and opisthorchiasis positively correlated with gallstones in univariate analysis. Serum lipids, family history of gallstones, consumption of alcohol and tobacco were not predictors of gallstones. Only association with age and obesity was significant in multivariate analysis. Among subjects with cholelithiasis 52.1% were not aware of having gallstones. Subjects with gallstones more frequently suffered from biliary colics and non-specific dyspeptic symptoms. However, their predictive value was poor. Cholecystectomized patients revealed more often upper abdominal pain and dyspeptic symptoms. CONCLUSION: Prevalence and risk factors for gallstones in female population of Novosibirsk are similar to those reported in Western European countries. Cholecystectomy is not recommended in patients with symptomless disease. PMID- 10717920 TI - [Paramagnetic centers of some biological media in cholecystitis patients]. AB - AIM: To study paramagnetic centers (PMC) of some biological media in cholecystitis patients. MATERIALS AND METHODS: PMC of 81 patients with chronic acalculous cholecystitis and 29 patients with acute calculous cholecystitis were examined using 3-cm radiospectrometer "Rubin" employing low-temperature fixation of biological tissues (77K). RESULTS: Most informative for diagnosis of cholecystitis exacerbation were the following PMC: in plasma--transferrin Fe3+ and free radicals; in duodenal portion B--Cu2+, free radicals and Mn2+. CONCLUSION: Electronic paramagnetic resonance is a perspective method in diagnosis of chronic cholecystitis exacerbation and in investigations of its pathogenesis. PMID- 10717922 TI - [Apudocytes and mast cells in chronic inflammation of colon: clinicomorphological correlations]. AB - AIM: To study functional morphology of a total population of endocrine cells of colon mucosa, mast and enterochromaffine cells. MATERIALS AND METHODS: Light and electrone microscopy, immunohistochemical methods, morphometry were used to study endocrine and mast cells of the sigmoid colon in inflammation. RESULTS: Changes of functional morphology and size of endocrine and mast cell population as well as apudocytes producing serotonin, melatonin, vasointestinal peptides were stated. Apudocyte and mass cell functional morphology, clinical symptoms and mucosal structural changes correlated. Specificity of some parameters in Chron's disease is shown. CONCLUSION: The results may provide additional criteria in diagnosis of different variants of chronic colitis. PMID- 10717921 TI - [Role of bioantioxidants depression and deficiency of protease inhibitor alpha 1 antitrypsin in mechanisms activating free radical oxidation and proteolysis in chronic pancreatitis]. AB - AIM: To elucidate the reason and mechanisms of neutralization of protease inhibitors and antioxidants by proteolytic enzymes and oxidants. MATERIALS AND METHODS: The trial included 92 patients with exacerbation of chronic pancreatitis. 47 of them had chronic recurrent pancreatitis (CRP), 45 patients had chronic fibrozing pancreatitis (CFP). Measurements were made of blood catalase and ceruloplasmin (according to P. Hubl, R. Breschneider and O. Houchin, respectively), alpha1-antitrypsin (by Reiderman), schiff bases (by B. Fletcher et al.), dienic conjugates (by Z. Placer), serum acid phosphatase (by Bodansky), acid phosphatase of polynuclear cells (by R. Nartsissiv), NBT-test was made according to B. Park. RESULTS: Exacerbation of CRP was associated with enhancement of free radical lipid peroxidation (FPOL), release of proteolytic and lysosomal enzymes from acinar cells, a fall in catalase level. Catalase depression depends on the level of blood lysosomal enzymes and partially on FPOL activity. In CFP moderate activity of FPOL and trypsin is associated with normal levels of lysosomal enzymes and catalase. In both pancreatitis forms, alpha1 antitrypsin levels are low. This lowering is primary and unrelated with inflammatory process in the pancreas. A trypsin rise in both forms depends on lowering of alpha1-antitrypsin which via trypsin inhibits formation of lysosomal enzymes in polynuclear cells. Inability of protease inhibitor to block proteolytic (lysosomal) enzymes manifests in initial intraacinar activation of trypsin from trypsinogen, in inflammatory focus under polynuclear cells release of lysosomal enzymes and in proteolytic enzymes release from the affected acinar structures. CONCLUSION: Lack of alpha1-antitrypsin--protease inhibitor--and depression of antioxidant catalase are main and intermediate elements in activation of mechanisms of proteolytic aggression and FPOL. PMID- 10717923 TI - [Chronic abdominal ischemia: clinical manifestations, diagnostic potential and treatment policy]. AB - AIM: To define variants of clinical course of chronic abdominal ischemia (CAI) and treatment policy depending on functional class of the disease. MATERIALS AND METHODS: The examination of 236 patients with CAI included clinicolaboratory, endoscopic, x-ray and ultrasonic investigations, ultrasonic dopplerography, x-ray contrast aortoarteriography, histological analysis of gastric, duodenal and colon mucosa biopsies. RESULTS: 6 variants of CAI clinical course were distinguished. Most frequent CAI manifestations were erosive-ulcerative gastroduodenal lesions, primarily in men, which combined with ischemic heart disease and atherosclerosis of lower limb vessels. Women often presented with pseudopancreatic variant which was associated with hypertension and dyslipoproteinemia. By severity of clinical symptoms, 3 functional classes of the diseases were recognized. They are decisive for the treatment policy. Biopsy specimens of the mucosa exhibited dystrophy in weak inflammation. CONCLUSION: Today's diagnostic techniques and recognition of functional classes of the disease contribute to correct decision on the treatment policy. PMID- 10717925 TI - [Peculiar course of paraneoplastic syndrome in esophageal cancer]. PMID- 10717924 TI - [Clinical course of hepato-gastro-duodenal diseases]. AB - AIM: To characterize clinical course of hepatogastroduodenal diseases (HGDD). MATERIALS AND METHODS: 897 patients with HGDD were examined aged 15 to 82 years. RESULTS: Three groups of patients were recognized: with predominant symptoms of pains in epigastric and hepatic area; with motor-evacuatory disturbances of the stomach and duodenum; with prevailing symptoms of asthenovegetative and depressive disorders. CONCLUSION: Distribution of the patients by type of clinical course provides algorithm of examination and helps to formulate optimal therapeutic policy. PMID- 10717926 TI - [Systemic enzyme therapy in chronic glomerulonephritis]. AB - AIM: To try polyenzyme drug vobenzim in chronic glomerulonephritis (CGN). MATERIALS AND METHODS: 174 CGN patients were randomized into 2 groups. The study group of 39 patients received conventional pathogenetic treatment plus vobenzim (initial dose 18-24, maintenance 12-15 dragees a day). Control group of 135 patients were given only pathogenetic therapy. 67 patients appeared ineligible. RESULTS: 3-6 months after the treatment the response was seen in 61.5% patients of the study and 17.8% patients of the control group. Side effects of vobenzim in CGN treatment were absent. CONCLUSION: Enzyme therapy with vobenzim in CGN is safe and effective. PMID- 10717927 TI - [Blood flow in affected major arteries of head in hypertensive patients]. AB - AIM: To assess blood flow in intracranial internal carotid and vertebral arteries in patients with essential hypertension (EH) and hemodynamically insignificant atherosclerotic lesions and deformities of major arteries of the head (MAH). MATERIALS AND METHODS: The blood flow was assessed in 55 untreated patients (mean age 53 +/- 1 years) with mild, moderate or severe hypertension. 20 healthy patients served control. Duplex scanning was performed using ACUSON unit. RESULTS: Total blood flow (Q) in hypertensive subjects appeared significantly subnormal. It was less in patients with hemodynamically insignificant atherosclerotic stenosis and flexures of the carotid and vertebral arteries than in patients with affection of the carotid arteries only. Q in MAH was lower in smokers than in non-smokers. PMID- 10717928 TI - [Ednit effects on activity of renin-angiotensin-aldosterone system and renal function in hypertensive subjects with diabetes mellitus]. AB - AIM: To evaluate effects of enalapril (ednit) therapy on activity and reactivity of renin-angiotensin-aldosteron system (RAAS) and renal function. MATERIALS AND METHODS: Ednit was given in a dose 5-10 mg/day to 30 patients with mild or moderate arterial hypertension (AH) in combination with moderate or severe non insulin-dependent diabetes mellitus (NIDDM) in compensation or subcompensation of carbohydrate metabolism. RESULTS: Ednit lowered blood pressure after 6-7 days of treatment. Normal blood pressure was achieved on the treatment week 4-6. Antihypertensive effect of ednit resulted from a significant reduction in a total peripheral vascular resistance under unchanged cardiac output. Adequate hypoglycemic and antihypertensive therapy normalized carbohydrate metabolism. Renal elimination of nitrogen did not change much. Glomerular filtration rate in patients with hyper- and hypofiltration returned to normal. Proteinuria reduced, microalbuminuria was not registered. RAAS activity normalized. CONCLUSION: Ednit has both high antihypertensive activity and marked renoprotective effect with minor influence on carbohydrate metabolism in AH patients with NIDDM. PMID- 10717929 TI - [Main causes of hyperuricemia in diabetes mellitus]. AB - AIM: Investigation of hyperuricemia (HU) causes in diabetes mellitus (DM). MATERIALS AND METHODS: Purin, lipid and carbohydrate metabolisms, renal excretion of uric acid (UA) were examined in 535 patients with DM. Non-insulin-dependent DM (NIDDM) and insulin-dependent DM (IDDM) were diagnosed in 369 and 166 patients, respectively. RESULTS: Hyperglycemia and glucosuria in DM enhance renal excretion of UA. However, HU in DM occurs significantly more frequently than in the population (22.5 +/- 3.0% in IDDM and 29.9 +/- 2.5% of cases in NIDDM). CONCLUSION: In NIDDM, HU genesis is strongly related with renal lesions. In IDDM, HU genesis is associated with metabolic factors (high activity of xantinoxidase, lipid peroxidation, obesity). However, combination of factors is, as a rule, involved in HU development: renal dysfunction and hyperproduction of UA. PMID- 10717930 TI - [Case of infection with hepatitis TT and G viruses in female patient with alcoholic liver cirrhosis]. PMID- 10717931 TI - [Primary hepatoportal sclerosis]. PMID- 10717932 TI - [Pharmacoeconomics in helicobacter pylori infection (lecture)]. PMID- 10717933 TI - [Barrier function of gastrointestinal tract]. PMID- 10717934 TI - [Primary biliary cirrhosis and CREST syndrome: a rare classical combination (literature review and case report)]. PMID- 10717935 TI - [Spontaneous bacterial peritonitis as a problem of current medicine]. PMID- 10717936 TI - [Left ventricular hypertrophy. Pathogenetic issues]. PMID- 10717937 TI - [Ethic problems of randomized trials]. PMID- 10717938 TI - The efficacy of the vaginal plug formation after mating for pregnancy diagnosis, and embyonic resorption in utero in the greater cane rat (Thryonomys swinderianus, Temminck). AB - The efficacy of the detection of vaginal plug formation after mating for pregnancy diagnosis, and the degree of embryonic resorption were studied in 67 wild greater cane rats (Thryonomys swinderianus) at Esiam in the Ekumfi District, Ghana, over a period of 3 months. Vaginal plug formation was first observed on day 59 of gestation, and could be used for pregnancy diagnosis on or after that date. However, the vaginal orifice subsequently opened a couple of times prior to day 105 of gestation and further checks for pregnancy after day 59 of gestation are suggested. Animals with unplugged vaginas 105 days after mating could, however, be considered as not pregnant. The mean (+/- SE) number of implantation sites and litter size in the greater cane rat were 7.2 +/- 0.18 and 3.4 +/- 0.29, respectively, the embryonic resorption rate being 42.7 +/- 6.66%. The significant positive correlation between the number of implanted embryos and the bled-out carcase weight suggests a positive role of maternal nutrition in increasing the litter size in the greater cane rat. The incidence of post-partum oestrus during the study period was 42.1%, which suggests that the greater cane rat can be re bred immediately after parturition. PMID- 10717939 TI - Reproduction and mortality in a colony of captive greater cane rats, Thryonomys swinderianus, Temminck. AB - The reproductive performance and mortalities in a colony of captive greater cane rats, Thryonomys swinderianus, were monitored from 1992 to 1998 at the Grasscutter Domestication Centre, Achimota, Ghana. The animals were kept in cages and exposed to constant lighting from a 100 W electric light bulb during the night. The diet consisted mainly of freshly cut Panicum maximum (guinea grass) fed ad libitum, with occasional supplements of cassava and cane sugar. The results indicate that the mean litter size and litter weight were 2.9 +/- 0.51 and 439.4 +/- 81.23 g, respectively. These figures are low compared to those reported elsewhere. However, the mean birth weight was 151.2 +/- 11.08 g, higher by 12% than previously reported values. It is considered that poor nutrition, excessive exposure to light and stress were responsible for the relatively poor reproductive performance reported in these animals. The main causes of death were traumatic injuries (32%) and pulmonary congestion (16%). PMID- 10717940 TI - Passive haemagglutination test in the serology of Newcastle disease virus. PMID- 10717941 TI - Effect of environmental factors and of the proportion of Holstein blood on the milk yield and lactation length of crossbred dairy cattle on smallholder farms in north-east Tanzania. AB - A study was carried out on the lactation performance of crossbred dairy cattle in a smallholder farming system in north-east Tanzania. Data were collected from the records for 6 years and the factors considered were district, proportion of Holsteins, season of calving, year of calving and herd size. The data were considered separately for animals with a single lactation record. The least square means for first lactation length and yield were 331 days (SD 77.0) and 2332 L (SD 283.0), respectively, while for cows with data on more than one lactation record the yield was 2477 L (SD 840.1) in 324 days (SD 74.0). First lactation yield was significantly affected by year of calving. For repeated records, the lactation yield was significantly affected by district, proportion of Holsteins and herd size, while lactation length was significantly affected by district and herd size. The calculated repeatabilities for lactation yield and length were 0.27 and 0.12, respectively. For the pooled data, the correlation between lactation length and yield gave r = 0.569 (p < 0.0001). PMID- 10717942 TI - Genetic and environmental effects on the productivity of Holstein-Friesian cattle under the climatic conditions of central Sudan. AB - A herd of 370 Holstein-Friesian cows were maintained in Central Sudan, Khartoum for intensive dairy production during the period 1990-1996. The area is characterized by high temperatures during the day and cools down at night, with an average thermal-humidity index (THI) of 74.8. The average adjusted lactation milk yield, milk yield per day, milk yield per day of calving interval and lactation length of the animals in the herd were 5117 +/- 123 (SE) kg, 14.7 +/- 0.25 kg, 11.3 +/- 0.36 kg and 350 +/- 8.0 days, respectively. Heifers calved at 25.2 +/- 2.3 (SD) months of age. The variation was large, which suggested large environmental fluctuations. Sire variance contributed 1.35% and 19.6% to the total variance in cows and heifers, respectively, while cow within sire contributed 19%. The heritabilities of total lactation yield, 305-day yield, milk per day of lactation, milk per day of calving interval, and lactation length in heifers were 0.78 +/- 0.24, 0.36 +/- 0.34, 0.39 +/- 0.24, 0.23 +/- 0.23, and 0.23 +/- 0.22, respectively. In cows, the heritability estimates were 0.05 +/- 0.24, 0.06 +/- 0.24, 0.08 +/- 0.24, 0.00 and 0.00 for the same traits, respectively. The repeatabilities of total lactation yield, 305-day yield, milk per day, milk per day of calving interval and lactation length were 0.02 +/- 0.03, 0.22 +/- 0.03, 0.17 +/- 0.02, 0.05 +/- 0.02 and 0.00, respectively. Regression analysis showed that, for each unit increase in THI, milk yield decreased by 0.29 +/- 0.04 kg. The stress usually caused by the combined effects of temperature and humidity was not severe in this area. PMID- 10717943 TI - The feeding of leaf meal of Calliandra calothyrsus to laying hens. AB - A 67-day feeding experiment was conducted to study the effects of inclusion of 5%, 7.5% or 10% leaf meal of Calliandra calothyrsus (calliandra) in the diets of laying hens on feed intake, egg production, egg weights, yolk colour and the birds' weights. While no significant effects were seen on either egg numbers or egg size, feed intake increased and the efficiency of feed utilization decreased with increasing inclusion of the foliage. Absolute initial and final body weights did not show significant treatment differences but live weight changes over the course of the experiments were statistically significant, weight gains decreasing with increasing calliandra levels. The strength of colour of the yolks increased within 3 days of offering the calliandra, irrespective of the level of inclusion. The persistence of the colour change after withdrawal of the leaf meal ranged from 3 days at the 5% inclusion to over 10 days at the 10% level. While it may be possible to include calliandra leaf meal in poultry rations along with other, local, low-cost components, there would appear to be little advantage in using it in conjunction with commercial layers meal at levels higher than those necessary to provide the desired pigmentation level in the yolks (5% or less). PMID- 10717944 TI - Injection of a boar sperm factor causes calcium oscillations in oocytes of the marsupial opossum, Monodelphis domestica. AB - At fertilisation, the sperm triggers an abrupt and transient increase in intracellular calcium ([Ca2+]i) in the oocyte cytoplasm. In eutherian mammals, oocytes exhibit multiple [Ca2+]i transients which are necessary for egg activation. We investigated [Ca2+]i in the marsupial opossum, Monodelphis domestica. Embryo development in this therian mammal is quite distinct from that in most Eutheria. Oestrus was induced in an adult female opossum by introduction of a male into her cage. Injection of a boar sperm extract induced repetitive increases in [Ca2+]i. Each oscillation travelled across and around the periphery of the egg in a wave-like manner. A control injection of KCl elicited no change in [Ca2+]i. This is the first observation of [Ca2+]i oscillations in the oocyte of a marsupial. The repetitive nature of the [Ca2+]i changes were more similar to those in oocytes of Eutheria than those in oocytes of non-mammalian vertebrates. PMID- 10717945 TI - Persistence of human mitochondrial DNA throughout the development to the blastocyst of mouse zygotes microinjected with human mitochondria. AB - The conditions for transfer of human mitochondria into fertilised mouse ova were elaborated. Species-specific primers were designed to discriminate human mitochondrial DNA (mtDNA) and the endogenous mtDNA in the preimplantation embryos. Human mitochondria isolated from the HepG2 cell line were microinjected into murine zygotes, and the latter cultured for 96 h to the blastocyst stage. The polymerase chain reaction allowed the detection of human mtDNA at every stage of embryo cleavage. In some cases a clear disparity in distribution of human mtDNA among blastomeres was evident. PMID- 10717946 TI - Microinjection of cyclic ADP-ribose triggers a regenerative wave of Ca2+ release and exocytosis of cortical alveoli in medaka eggs. AB - Medaka (Oryzias latipes) eggs microinjected with the Ca(2+)-mobilising messenger cyclic adenosine diphosphate ribose (cADPR) underwent a wave of exocytosis of cortical alveoli and were thus activated. The number of eggs activated was sharply dependent on the concentration of cADPR in the pipette, the threshold concentration was approximately 60 nM. After injection, a pronounced latency preceded the onset of cortical alveoli exocytosis; this latency was independent of the concentration of cADPR but decreased markedly with increasing temperature. Heat-treated cADPR, which yields the inert non-cyclised product ADP-ribose, was ineffective in activating eggs. When cADPR was injected into aequorin-loaded eggs, a wave of luminescence arose at the site of cADPR injection and then swept out across the egg with a mean velocity of approximately 13 microns/s; the velocity was independent of the concentration of injected cADPR. In such a large cell (diameter of around 1 mm), this is considerably faster than that possible by simple diffusion of cADPR, which unambiguously demonstrates that cADPR must activate a regenerative process. cADPR has been demonstrated to modulate Ca(2+) induced Ca2+ release (CICR) via ryanodine receptors (RyRs) in many cell types, and consistent with this was the finding that microinjection of the pharmacological RyR modulator, ryanodine, also activated medaka eggs. These results suggest that a cADPR-sensitive Ca2+ release mechanism is present in the medaka egg, that cADPR is the most potent activator of medaka eggs described to date, and that it activates eggs by triggering a wave of CICR from internal stores that in turn stimulates a wave of exocytosis. PMID- 10717947 TI - Control of duration of the first two mitoses in a mouse embryo. AB - The duration of M-phase is largely determined by the time necessary for the formation of a functional metaphase spindle and the correct alignment of all chromosomes on the metaphase plate. The spindle assembly checkpoint prevents the exit from M-phase before the proper alignment of all chromosomes on a metaphase plate in many cell types. In the present paper we show that the first mitotic M phase of the mouse embryo lasts about 119 min, while the second embryonic M-phase lasts only about 70 min. Histone H1 kinase is activated rapidly during nuclear envelope breakdown in both mitoses. Its maximum, however, is followed by a plateau only during the first mitosis. In the second mitosis, the inactivation of histone H1 kinase activity follows its maximum directly. Histone H1 kinase is more stable in the cytoplasts obtained from mouse embryos during the first embryonic M-phase than during the second one. The stability of histone H1 kinase is greatly increased by the presence of the mitotic apparatus in both M-phases. The mitotic spindle assembly during the first and the second mitoses differs and the first metaphase spindle is stabilised during the period of maximum histone H1 kinase activity. These data show that an unknown developmentally regulated mechanism controls the duration of the two first mitoses in the mouse embryo. PMID- 10717948 TI - Changes in intracellular content of glutathione and thiols associated with gamma glutamyl cycle during sperm penetration and pronuclear formation in rat oocytes. AB - The content of glutathione and other thiols in rat eggs was examined during sperm penetration and pronuclear formation by high-performance liquid chromatography with fluorescence detection. Reduced glutathione (GSH) content was higher in unfertilized oocytes (8.50 +/- 0.29 pmol/egg) and penetrated eggs with a decondensed sperm nucleus (DSH eggs; 7.72 +/- 0.56 pmol/egg) than eggs at the pronuclear stage (PN eggs; 5.93 +/- 0.10 pmol/egg). The content of oxidised glutathione (GSSG) was not different among experimental groups (152.6 +/- 74.1 nmol/egg in unfertilized eggs, 146.0 +/- 50.0 nmol/egg in DSH eggs and 39.7 +/- 17.3 nmol/egg in PN eggs). The GSSG/GSH ratio did not change during fertilization. Although the reduced cysteinylglycine content of eggs did not change among experimental groups, the oxidised form of cysteinylglycine increased (p < 0.025) between sperm decondensation (6.9 +/- 1.5 nmol/egg in unfertilized oocytes and 10.1 +/- 2.1 nmol/egg in DSH eggs) and pronuclear formation (40.5 +/- 11.5 nmol/egg in PN eggs). Low contents of cystine were detected during fertilization but cysteine and gamma-glutamylcysteine were not detected in any treatment groups. These results demonstrate that GSH content in rat eggs decreases between sperm decondensation and pronuclear formation, probably due to the increased activity of gamma-glutamyl transpeptidase. PMID- 10717949 TI - Ultrastructural observations on in vivo fertilisation in the brushtail possum, Trichosurus vulpecula, following PMSG/LH superovulation and artificial insemination. AB - Information on the dynamics of gamete interaction in marsupials is very limited and not available for any species from the major Australian Order Diprotodontia which includes most of the more familiar animals such as kangaroos, possums and the koala. This study addressed this deficiency by examining the ultrastructure of in vivo fertilised eggs from common brushtail possums (Trichosurus vulpecula). Females were superovulated by treatment with 15 IU PMSG and then 4 mg porcine LH 3 days later, and inseminations were performed 910-13 h after LH) using epididymal spermatozoa. Between 33 and 39 h after LH injection females were killed, reproductive tracts excised and the oviduct ampulla segment flushed for eggs. Three of the six eggs examined were fertilised as judged by the presence of sperm remnants in the cytoplasm. On the basis of these eggs it was found that sperm penetration left a large hole in the zona pellucida (ZP), suggesting that sperm zona penetration occurs primarily by the enzymatic action of acrosomal enzymes. Sperm lying within the perivitelline space were lacking both an outer acrosomal membrane and the associated acrosomal contents, while both these structures were found on sperm embedded within the mucoid layer, which is consistent with induction of the acrosome reaction by binding to the ZP. Once inside the egg cytoplasm, the sperm head travelled only a short distance before chromatin decondensation occurred. Fertilised eggs showed signs of cytoplasmic activation including cytoskeleton association with apparently dividing mitochondria and prominent rough endoplasmic reticulum. Unfertilised eggs appeared to be undergoing degenerative changes and lacked any evidence of activation. This study was demonstrated that superovulation and laparoscopic intravaginal artificial insemination provide a system through which perifertilisation events in the possum and other monovular Australian marsupials can be examined experimentally. PMID- 10717950 TI - Dithiothreitol induces sperm nuclear decondensation and protects against chromosome damage during male pronuclear formation in hybrid zygotes between Chinese hamster spermatozoa and Syrian hamster oocytes. AB - The present study was undertaken to investigate whether a time lag in sperm nuclear decondensation and male pronuclear formation in the course of development of eggs is associated with any occurrence of structural chromosome aberrations in male genomes of hybrid zygotes between Chinese hamster spermatozoa and zona-free Syrian hamster oocytes. Shortly after insemination, hybrid zygotes were treated with dithiothreitol (DTT) at different concentrations (0.1-10.0 mM) for 30 min to reduce protamine disulphide (S-S) bonds and thereby accelerate sperm nuclear decondensation and male pronuclear formation. The incidence of sperm nuclear decondensation and male pronuclear formation increased with increasing DTT concentrations, indicating that a reduction in S-S bonds effectively induces these cytological events. Chromosomes of male genomes in hybrid zygotes generated by treatment with 1.0 mM, 2.5 mM and 10.0 mM DTT were analysed at the first cleavage metaphase. Incidence of structural chromosome aberrations in each treatment was 34.5%, 27.1% and 24.7%, respectively. There was a significant difference between the incidences with 1.0 mM and 10.0 mM DTT treatment. As the time lag in nuclear decondensation and male pronuclear formation was greatest in the 1.0 mM treatment condition, followed in order by 2.5 mM and 10.0 mM, it is suggested that the lag in sperm nuclear development behind egg development is responsible for structural chromosome aberrations in male genomes of hybrid zygotes. PMID- 10717951 TI - Initiation of sperm motility in the newt, Cynops pyrrhogaster, is induced by a heat-stable component of egg-jelly. AB - Sperm motility in amphibians is thought to be initiated by a decrease in environmental osmolarity. However, fertilisation in the newt, Cynops pyrrhogaster, is achieved in an environment without osmotic change. We show here that sperm motility initiating activity is present in jelly layer extract (JE). JE was gel-filtrated and a single peak with sperm motility initiating activity was detected in the fraction corresponding to about 50 kDa. The activity was strengthened by heat treatment of JE at 100 degrees C for 30 min. This suggests that JE includes the inactive form of sperm motility inducing substance (SMIS) in addition to active substance. Thus JE was fractionated before and after the heat treatment. When JE was fractionated first and then each fraction was heated, the activity was detected in the fraction both above 500 kDa and below 500 kDa. When heat-treated JE was fractionated, the activity was detected only in the fraction below 500 kDa. These results suggest that JE includes the inactive form of SMIS of more than 500 kDa in molecular weight. A regulatory mechanism for the initiation of sperm motility in C. pyrrhogaster is proposed according to the results of the present study. PMID- 10717952 TI - Mannose-binding molecules of rat spermatozoa and sperm-egg interaction. AB - We have previously reported the occurrence and partial characterisation of an alpha-D-mannosidase activity on plasma membranes of rat, mouse, hamster and human spermatozoa. A soluble isoform of the rat sperm surface mannosidase was purified and polyclonal antibody raised. Since several reports have suggested that mannosyl residues on the rat, mouse and human zona pellucida may be involved in sperm-zona binding, studies were undertaken to examine the receptor-like role of mannose-binding molecules on rat spermatozoa. Sprague-Dawley rats (25-30-days old) were superovulated and eggs collected from the oviduct were treated with 0.3% hyaluronidase to remove the cumulus cells. Spermatozoa, collected from the cauda epididymis were capacitated for 5 h at 37 degrees C in 5% CO2 in air. The sperm-zona binding assay was performed in the presence of increasing concentrations of several sugars as well as preimmune and immune (anti mannosidase or anti-mannose binding protein) IgG. Data from these studies show that: (1) significantly fewer sperm bound per egg in the presence of competitive inhibitors of mannosidase; (2) among the sugars examined, D-mannose was the most potent inhibitor causing 70% reduction in the number of sperm bound per egg; (3) anti-mannosidase or anti-mannose binding protein (but not preimmune) IgG showed a dose-dependent reduction in the number of sperm bound per egg; (4) anti mannosidase IgG (but not anti-mannose binding protein IgG) showed a dose dependent inhibition of sperm surface mannosidase activity; (5) the competitive inhibitors of mannosidase or the immune IgG had no effect on sperm motility or the sperm acrosome reaction. These result suggest that mannose-binding molecule(s) such as alpha-D-mannosidase or mannose-binding protein on the spermatozoa may recognise mannosyl residues on zona pellucida, and play a receptor-like role in sperm-egg interaction in the rat. PMID- 10717953 TI - The kinetics of oocyte activation and dynamics of polar body formation in Calomys musculinus and Calomys laucha (Rodentia-Sigmodontinae). AB - The objective of this study was to determine whether Calomys laucha and Calomys musculinus superovulated oocytes undergo parthenogenetic activation following activation stimuli. Cumulus-intact or denuded oocytes were treated with medium containing ethanol (7%), medium containing strontium chloride, or medium alone. They were then incubated for 6-8 h to allow for activation. A group of oocytes was fixed immediately after maturation to serve as a control. The nuclear status of the oocytes was examined after staining with Hoechst 33342, to determine the timing of pronuclear progression from metaphase II to anaphase II or telophase II or to the pronuclear stage. The proportion of oocytes that underwent activation was higher for oocytes treated with ethanol or strontium chloride than in those incubated in medium alone, for the two species studied (p < 0.001). There was little evidence of spontaneous activation occurring in oocytes during the treatments. Most of the activated oocytes contained a single haploid pronucleus, but it was possible to find immediate cleavage and two pronuclei. The different classes of activated oocytes were cultured for 5 days. The type of activating treatment had a marked effect on the ability of the resulting C. musculinus and C. laucha parthenogenetic embryos to develop to the preimplantation stages. Incubation with ethanol produced only 8-cell embryos while the embryos induced with strontium chloride reached the blastocyst stage. This is the first report of parthenogenesis in C. musculinus and C. laucha. The ability of strontium ions to induce matured secondary oocytes to initiate parthenogenesis and obtain further development of Calomys provides opportunities to use Calomys oocytes in vitro and, therefore, to study the genetics, cell biology and virology of development. PMID- 10717954 TI - Farewell to a unique institution. PMID- 10717955 TI - The UK Tobacco White Paper: beacon of hope or white elephant? PMID- 10717956 TI - Drug transitions: uncoupling drug and route. PMID- 10717957 TI - A confidential enquiry into methadone-related deaths. AB - AIM: To assess the acceptability and usefulness of the "confidential enquiry" process in examining methadone-related deaths. DESIGN: An audit of patient care. SETTING: Glasgow, Scotland, UK (population 915,000) Participants. All doctors who, in the final 14 days of the patient's life, had attended a patient who suffered a methadone-related death. MEASUREMENTS: The medical care of each case was assessed by peer review and the results of these assessments returned to the responsible clinician(s). FINDINGS: (1) The audit cycle was completed in 32 of the 34 reported cases (94%). (2) Twenty-eight of 33 doctors (85%) found the audit to be helpful. (3) As a result of the enquiry, the majority of doctors whose patient management had attracted criticism intended to amend their practice. (4) Shortcomings in clinical care were identified in 18 cases (56%) and problems in the organization of services in 22 (69%). CONCLUSIONS: (1) The model of audit piloted here was found to be highly acceptable to participants. (2) The episodes of substandard care that were uncovered provided useful opportunities to improve the future management of patients who were being prescribed methadone. PMID- 10717958 TI - Transitions between the injection of heroin and amphetamines. AB - AIMS: To assess in current injecting heroin and amphetamine users to what extent their history of injecting represents a transition from amphetamine to heroin. DESIGN: Cross-sectional survey. SETTING: Sydney, Australia. PARTICIPANTS: One hundred and fifty-one primary heroin injectors and 145 primary amphetamine injectors recruited through advertisements, needle exchanges, methadone maintenance clinics and by word of mouth. FINDINGS: Six major transition patterns were detected: heroin-heroin (n = 61), amphetamines-heroin (n = 60), heroin amphetamines-heroin (n = 30), amphetamines-amphetamines (n = 80), amphetamines heroin-amphetamines (n = 41) and heroin-amphetamines (n = 24). A logistic regression analysis predicting presence or absence of a transition from the original primary drug indicated that length of injecting career, years of education and original drug injected were independent predictors. Thus, the longer the injecting career, the greater the likelihood of a transition. If the original drug injected was amphetamine, the greater the likelihood of transition; and the more prior years of education, the lower the chances of a transition. CONCLUSIONS: While there was a small preponderance of movement from primary amphetamine injecting to primary heroin injecting, there was also movement in the other direction. Heroin use is not necessarily a stable endpoint for injecting careers. PMID- 10717959 TI - Dual diagnosis patients in substance abuse treatment: relationship of general coping and substance-specific coping to 1-year outcomes. AB - AIMS: This study examined general and substance-specific coping skills and their relationship to treatment climate, continuing care and 1-year post-treatment functioning among dual diagnosis patients (i.e. co-occurrence of substance use and psychiatric disorders). DESIGN: In a prospective multi-site study, dual diagnosis patients participating in substance abuse treatment were assessed at intake, discharge and at a 1-year follow-up. SETTING: Patients were recruited from 15 substance abuse treatment programs, which were selected from a larger pool of 174 inpatient treatment programs in the Department of Veterans Affairs Health Care System. PARTICIPANTS: A total of 981 male dual diagnosis patients participated in the study. MEASUREMENTS: Assessments included general and substance-specific coping skills, treatment climate, continuing outpatient care, abstinence and clinically significant psychiatric symptoms. FINDINGS: Dual diagnosis patients modestly improved on general and substance-specific coping skills over the 1-year follow-up period. Patients who were in programs with a 'dual diagnosis treatment climate' and who participated in more 12-Step self-help groups showed slightly more gains in adaptive coping. Both general and substance specific coping were associated with abstinence, but only general coping was associated with freedom from significant psychiatric symptoms. CONCLUSIONS: Enhancing general and substance-specific coping skills in substance abuse treatment may reduce dual diagnosis patients' post-treatment substance use and improve their psychological functioning. PMID- 10717960 TI - Reinforcing, subjective, and psychomotor effects of sevoflurane and nitrous oxide in moderate-drinking healthy volunteers. AB - AIMS: To characterize the reinforcing, subjective and psychomotor effects of sevoflurane, a volatile anesthetic, across a range of subanesthetic concentrations in non-drug-abusing humans. In addition, a concentration of nitrous oxide was included in the design in order to compare and contrast behavioral effects of a gaseous to a volatile anesthesic. DESIGN: Repeated measures, double-blind, placebo control experiment. SETTING: Human psychopharmacology laboratory. PARTICIPANTS: Fourteen moderate-drinking healthy volunteers. INTERVENTION: In each of four sessions, subjects first sampled placebo-oxygen and an active drug (end-tidal concentrations of 0.2, 0.4, 0.6% sevoflurane and 30% nitrous oxide in oxygen) and then chose between the two MEASUREMENTS: Mood and psychomotor performance during the sampling trials, and choice of drug or placebo-oxygen during choice trial. FINDINGS: Nitrous oxide was chosen by 71% of the subjects, and 0.2, 0.4 and 0.6% sevoflurane were chosen by 50%, 57% and 50% of the subjects, respectively. Neither drug was chosen at levels that exceeded that of chance. Sevoflurane and nitrous oxide both impaired psychomotor performance and produced changes in mood. There were several differences in subjective effects between sevoflurane and nitrous oxide at concentrations which were considered to be equivalent in anesthetic effect. Finally, although sevoflurane did not function as a reinforcer in the majority of individuals tested, there was evidence that sevoflurane functioned as a reinforcer in some volunteers: subjects who chose to inhale sevoflurane over placebo-oxygen tended to report a positive spectrum of subjective effects during the sevoflurane sampling trial, relative to those subjects who chose placebo oxygen over sevoflurane. CONCLUSIONS: Although sevoflurane did not function as a reinforcer in the majority of subjects tested, the correspondence between positive subjective effects of sevoflurane and subsequent sevoflurane choice suggests that the volatile anesthetic drug can function as a reinforcer in some moderate drinkers. PMID- 10717961 TI - HIV risk behaviours among gay men who use anabolic steroids. AB - AIMS: To examine the injecting and sexual risk behaviours of gay men in London who use anabolic steroids or other fitness-enhancing substances (referred to as AS). DESIGN: Cross-sectional survey using self-administered questionnaire. SETTING: Five gyms in central London. PARTICIPANTS: 1004 gay men using the gyms during September-October 1997. MEASUREMENTS: Proportion of men who report (i) injecting AS, (ii) sharing needles and (iii) HIV status-unknown unprotected and intercourse (UAI). FINDINGS: Of 1004 men, 136 (13.5%) were current users of AS (range across the five gyms, 2.7-21.2%, p < 0.001), and 81 (8.1%) injected AS (range 0.4-17.5%, p < 0.001). None said they shared a needle with other users and more than 90% said they always used disposable units from sealed packets. Among current AS users, 20.9% (28/136) reported status-unknown UAI compared with 12.9% (107/827) of never-users (p = 0.02), a differential which remained significant after adjusting for confounding factors (adjusted odds ratio = 1.75, 95% CI 1.05, 2.91, p = 0.03). CONCLUSIONS: While nearly one in 10 gay men in this study injected anabolic steroids or other fitness-enhancing substances, none reported sharing needles. Steroid users were, however, more likely to report status unknown UAI than other men, a differential that remained after adjusting for confounding factors. Since any change in injecting practice could dramatically increase the risk of HIV transmission in this population, behavioural surveillance to monitor risk behaviours among gay men using anabolic steroids is recommended, as are targeted HIV prevention programmes. PMID- 10717962 TI - Narratives of recovery from addictive behaviours. AB - AIMS: The purpose of this study was to look for the ways in which people who have recovered from various addictions understand and present their change process. MATERIALS: The research material consisted of 51 autobiographical stories of people who had been able to quit their addiction to alcohol, multiple drugs, binge eating, smoking, sex and gambling. METHODS: The basic logic of each narrative was first defined. The narratives were then categorized according to what they presented as the key to recovery. Composite stories were then constructed and analysed with regard to their emotional, causal, moral and ethical meanings. FINDINGS: The analysis revealed five different story types among these self-narratives: the AA story, the growth story, the co-dependence story, the love story and the mastery story. All of them helped to make the addiction and recovery understandable, they released the protagonist from guilt and had a happy ending by which the values of the story were realized. Each story type was told predominantly by representatives of a particular gender and addiction. CONCLUSIONS: As there are several ways out of addictive behaviours there are also several ways to construe the change. People who try to quit addictive behaviours could be encouraged to make full use of the cultural stock of stories in creating an account that fits their own experience of defeating their particular addiction. PMID- 10717963 TI - The Alberta Interlock Program: the evaluation of a province-wide program on DUI recidivism. AB - AIMS: This study was designed to determine the efficacy of alcohol safety interlocks in reducing recidivism among first and second driving-under-the influence (DUI) offenders. It also evaluates the overall effectiveness of interlock programs where typically only a small portion of DUI offenders elect to install interlocks. DESIGN: The driving records of DUI offenders participating in interlock programs for 6 months for first offenders and 2 years for second offenders were compared with similar offenders who chose not to participate. SETTING: A province-wide program in Alberta, Canada. PARTICIPANTS: Records of 35,132 drivers convicted of DUI between 1 July 1998 and 30 September 1996 were analyzed MEASUREMENTS: Repeat DUI offenses during and after the interlock period. FINDINGS: While the offenders had interlocks on their vehicles, DUI recidivism was substantially reduced. Once the interlock had been removed and the participants had been reinstated, their DUI rate was the same as other offenders indicating that the interlock reduced recidivism while in place. Because only 8.9% of eligible drivers elected to participate in the interlock program, the program did not significantly increase the overall effectiveness of the province's management of DUI offenders. CONCLUSIONS: Interlocks are associated with a major reduction in DUI recidivism while on the vehicle of the offender. However, because few offenders elect to participate, the program produces only a small (5.9%) overall reduction in the recidivism rate of all DUI offenders. PMID- 10717964 TI - Behavioral monitoring of DUI offenders with the Alcohol Ignition Interlock Recorder. AB - AIMS: To evaluate patterns of blood alcohol concentration (BAC) and driving logged on the ignition interlock recorder and to assess whether this event record is a useful outcome measure for a behavioral intervention. DESIGN: Descriptive analyses of recorder data and multivariate analysis of the predictors of high BACs associated with a motivational intervention for driving-under-the-influence (DUI) offenders using an interlock. SETTING: Two interlock service centers in Alberta, Canada. PARTICIPANTS: 1309 first-time and multiple DUI offenders who agreed to participate during interlock installation. INTERVENTION: A human services (supportive guidance) intervention based on motivational interviewing and pragmatic counseling was delivered to interlock users in Calgary, but not to interlock users in Edmonton, Canada. MEASUREMENTS: This report summarizes the patterns and predictors of BAC warnings (0.02-0.039%) (20-39 mg/dl) and failures (> or = 0.04) (> or = 40 mg/dl) from more than 3 million in-vehicle breath tests. Data come from three sources: driver records, questionnaires and the interlock. FINDINGS: From the beginning to the end of the interlock use period, there was a significant linear decline in the proportion of positive BAC driving to total driving. After controlling for prior offenses, demographics and reported drinking levels, offenders in the intervention site (Calgary) were less likely to have recorded fail BACs than were offenders in the control site (Edmonton). The temporal patterns of BAC fails with the interlock mimic the high-risk periods for DUI arrests and alcohol-involved fatal crashes. CONCLUSIONS: The interlock successfully blocks drinking and driving during high-risk periods. Preliminary recorder data suggest the services intervention may be affecting DUI behavior. PMID- 10717966 TI - A comparison of two measures of stage of change for smoking cessation. AB - AIMS: To compare two questionnaires used to identify the stages of change of current and former smokers: a conventional five-item questionnaire and an alternative one-item questionnaire. DESIGN AND SETTING: Mail surveys of 1167 ever smokers in Geneva (Switzerland), conducted in 1997. Participants were classified into five stages: precontemplation, contemplation, preparation, action and maintenance. Other questions covered smoking-related behaviours, attitudes and self-efficacy. FINDINGS: Only 62% of participants were classified as being in the same stage by the two questionnaires (weighted kappa = 0.69). The five-item questionnaire produced more missing data (8%) than the one-item questionnaire (2%, p < 0.001). Using the conventional questionnaire, the precontemplation stage included a group of smokers who had absolutely no intention of stopping smoking and a group more prone to change, and the preparation stage included only 43% of people who had made a "firm decision" to quit smoking in the next 30 days. Using the alternative questionnaire, the contemplation stage was also quite heterogeneous. The action stage included over 35% of people who were still smoking occasionally, whichever questionnaire was used. CONCLUSIONS: The single item questionnaire was better at avoiding missing responses. However, both staging questionnaires classified smokers in heterogeneous groups, and both misclassified many occasional smokers and ex-smokers, which suggests that a discrete five-stage model does not fit reality well. This may reflect underlying conceptual issues, notably that the classic definition of stages incorporates separate dimensions, albeit incompletely (current behaviour, quit attempts, intention to change, time). Both theoretical and methodological developments are needed to overcome these problems. PMID- 10717965 TI - Do the rich really die young? Alcohol-related mortality and social class in Great Britain, 1988-94. AB - AIMS: To determine whether social class is a major influence on alcohol-related mortality in the general, economically active population of Great Britain. DESIGN AND PARTICIPANTS: Poisson regression of rates of mortality known to be directly caused by alcohol consumption by age, sex and social class in England, Wales and Scotland. MEASUREMENTS: The measure of alcohol-related mortality is total deaths from ICD-9 categories 291; 303; 357.5; 425.5; 535.3; 305; 790.3; and 571.0-571.3 over the 7-year period 1988-94. (It excludes deaths for which alcohol attributable fractions would need to be calculated.) The measure of social class is the British Registrar General's six-fold occupational classification, used to code census and death certification data. FINDINGS: Alcohol-related mortality rates are higher for men in the manual occupations than in the non-manual occupations, but the relative magnitude depends on age. Men aged 25-39 in the unskilled manual class are 10-20 times more likely to die from alcohol-related causes than those in the professional class, whereas men aged between 55 and 64 in the unskilled manual class are only about 2.5-4 times more likely to die. For women in paid employment there is no consistent class gradient; younger women in the manual classes are more likely to die from alcohol-related causes, but for older women it is those in the professional class who suffer elevated mortality. CONCLUSIONS: Social class is a risk factor for alcohol-related mortality in Britain, although it is mediated by age and sex. Alcohol appears to be similar to other psychoactive substances, therefore, in that problem use is linked to social structural factors such as poverty, disadvantage and social class. This suggests that social interventions aimed at reducing poverty and inequality have the potential to reduce current levels of alcohol-related harm among the poorest groups in the community. PMID- 10717967 TI - Charles Dickens and the neurological consequences of alcoholism. PMID- 10717968 TI - Who talks? The social psychology of illness support groups. AB - More Americans try to change their health behaviors through self-help than through all other forms of professionally designed programs. Mutual support groups, involving little or no cost to participants, have a powerful effect on mental and physical health, yet little is known about patterns of support group participation in health care. What kinds of illness experiences prompt patients to seek each other's company? In an effort to observe social comparison processes with real-world relevance, support group participation was measured for 20 disease categories in 4 metropolitan areas (New York, Chicago, Los Angeles, and Dallas) and on 2 on-line forums. Support seeking was highest for diseases viewed as stigmatizing (e.g., AIDS, alcoholism, breast and prostate cancer) and was lowest for less embarrassing but equally devastating disorders, such as heart disease. The authors discuss implications for social comparison theory and its applications in health care. PMID- 10717969 TI - Contemporary research on parenting. The case for nature and nurture. AB - Current findings on parental influences provide more sophisticated and less deterministic explanations than did earlier theory and research on parenting. Contemporary research approaches include (a) behavior-genetic designs, augmented with direct measures of potential environmental influences; (b) studies distinguishing among children with different genetically influenced predispositions in terms of their responses to different environmental conditions; (c) experimental and quasi-experimental studies of change in children's behavior as a result of their exposure to parents' behavior, after controlling for children's initial characteristics; and (d) research on interactions between parenting and nonfamilial environmental influences and contexts, illustrating contemporary concern with influences beyond the parent child dyad. These approaches indicate that parental influences on child development are neither as unambiguous as earlier researchers suggested nor as insubstantial as current critics claim. PMID- 10717970 TI - Boulder at 50. Introduction to the section. PMID- 10717971 TI - Clinical psychology training. A history of ideas and practices prior to 1946. PMID- 10717972 TI - The affirmation of the scientist-practitioner. A look back at Boulder. AB - In the aftermath of World War II, several influences were paramount in forcing academic psychology to recognize, albeit reluctantly, the coming professionalization of psychology. The federal government, wishing to avoid a repeat of blunders following World War I that led to significant dissatisfaction among veterans, took proactive steps to ensure that mental health needs of the new veterans would be met. The USPHS and the VA were mandated to expand significantly the pool of mental health practitioners, a direction that led not only to the funding of the Boulder conference but also to the development of APA's accreditation program, funded practical and internship arrangements with the VA, and the USPHS grants to academic departments for clinical training. The GI Bill, amended to include payment for graduate education, created tremendous interest in graduate programs in psychology. As a result, psychology programs were inundated with funded applicants, most of whom were interested in the application of psychology to clinical and other applied fields. Graduate psychology departments were mixed in their views of this "blessing." The reality of a separate curriculum for professional training in psychology was a bitter pill for some academic psychologists to swallow. Graduate departments feared that control of their programs would be taken over by external forces and that they would lose their right to determine their own curriculum. Further, they feared the domination of clinical training within their own departments and the effects of such educational emphasis on their traditional experimental programs. The Boulder conference brought together these disparate needs and concerns, although one can argue about how well some points of view were represented with respect to others. It was a time of high anticipation and fear. The conference could easily have ended in failure, with such diverse interests being unable to reach any consensus. There are many letters in the correspondence of committee members that suggest disagreements serious enough to prevent the development of any single model of training. Instead, by most yardsticks that one could apply, the conference succeeded, perhaps beyond the dreams of many of those in attendance who were most invested in a model for professional training. In evaluating the legacy of Boulder, several points are apparent. First, the conference succeeded because 73 individuals were able to agree to some 70 resolutions in 15 days, creating the scientist-practitioner model of professional training. Such consensus was arguably a remarkable achievement. The endorsement of the model by academic units followed with little evident resistance, although it is clear that some Boulder-model programs were developed that bore little resemblance to the model's insistence on significant training in both research and practice. Second, as a response to social and political needs, the conference was clearly a success. The cooperation of the APA, the USPHS, and the VA benefited all three entities. Clinical psychology was given the financial support and backing to advance it as a profession, and the federal government was able to begin the process of securing the personnel needed to address the mental health needs of the nation. The architects of Boulder were clear that their vision of training for professional psychology should be viewed as dynamic and experimental rather than fixed and prescribed. Certainly there are several variants of professional training extant today, yet the overwhelming majority of currently accredited programs in psychology label themselves as "Boulder-model" programs or "scientist practitioner" programs. Still, new national conferences on professional training in psychology occur with some regularity as participants seek to resolve many of the same concerns debated by those at Boulder. The grand experiment goes on. PMID- 10717973 TI - The Boulder model's fatal flaw. PMID- 10717974 TI - Scientist-practitioner not equal to science + practice. Boulder is bolder. PMID- 10717975 TI - The Boulder model. A dream deferred--or lost? PMID- 10717976 TI - Scientist-practitioner or scientific practitioner? PMID- 10717977 TI - The scientist-practitioner model. Gandhi was right again. PMID- 10717978 TI - Snapshots of psychology circa 1900. PMID- 10717979 TI - Psychology without p values. Data analysis at the turn of the 19th century. AB - Although the fledgling psychology of 100 years ago was assertively empirical, there were no inferential statistics to guide psychologists' data analyses. However, 19th-century developments had left psychology with a rich array of techniques for analyzing and presenting data, some of which remain underutilized today. These include comparisons across replications, within-subject designs, reanalysis of data, analyses of factorial designs, and especially the use of tables and graphs. As the merits of hypothesis-testing statistics are debated at the turn of the 21st century, the history of data-handling practices can remind psychologists that there are many ways to overcome the current uniformity of statistical practice. PMID- 10717980 TI - From deception trials to control reagents. The introduction of the control group about a century ago. AB - This is the story of the remarkable psychologist John E. Coover, who, in the early 1900s, was the first to advocate the comparison of experimental and control groups as a methodological necessity. Moreover, the author raises the issue of why control groups were launched about a century ago, and why psychology was comparatively early in codifying group comparison as a methodological routine. In dealing with these questions, the author discusses the relations between turn-of the-century science and society as well as between psychophysical research and educational experimentation. Furthermore, the mystery is solved of how Coover's rightful place in the received history of experimental controls could be taken by precisely the authors whom he criticized for the lack of controls. PMID- 10717982 TI - Theta H. Wolf (1904-1997) PMID- 10717981 TI - Richard W. Husband (1904-1995) PMID- 10717983 TI - Neuroscience and mainstream psychology. PMID- 10717984 TI - Corroboration. PMID- 10717985 TI - Difficulties in analyzing trends in psychology. PMID- 10717986 TI - On cognitivism and behaviorism. PMID- 10717987 TI - Questionable validity, not vitality. PMID- 10717988 TI - There's more neuroscience. PMID- 10717989 TI - Trends in psychology: an empirical issue. PMID- 10717990 TI - Enculturative continuity and adolescent stress. PMID- 10717991 TI - Violence against women and the impending AIDS crisis in Russia. PMID- 10717992 TI - Genetic tests mark new era for dentistry. PMID- 10717993 TI - Lead and dental decay. PMID- 10717994 TI - Ribbon dentistry. PMID- 10717995 TI - Dentists' awareness of bias questioned. PMID- 10717996 TI - Saliva substitutes. PMID- 10717997 TI - Allergy to local anaesthetic: the importance of thorough investigation. AB - A case report is presented which highlights the importance of a good history in arriving at the correct diagnosis in cases where allergy to local anaesthetic is suspected. Management of the patient is discussed and the topic of 'adverse reaction' briefly reviewed. PMID- 10717998 TI - Personal Dental Services--a practitioner's point of view. AB - NHS dentistry has seen many changes recently. The latest has been the introduction of Personal Dental Services. This article describes the experiences of two general dental practitioners in their practice who entered as a first wave pilot. The article explains the steps involved in generating a proposal and preparing a practice to run as a pilot. The authors have highlighted areas of particular concern for others to consider before embarking on a similar journey. PMID- 10717999 TI - Dental care for the patient with a cleft lip and palate. Part 2: The mixed dentition stage through to adolescence and young adulthood. AB - This is the second of two articles looking at dental care for the patient with a cleft lip and palate. Part 1 looked at the needs of the child from birth through to the mixed dentition stage. Part 2 looks at dental care from the mixed dentition stage through to adolescence and young adulthood. PMID- 10718000 TI - Referral patterns and the referral system for oral surgery care. Part 1: General dental practitioner referral patterns. AB - OBJECTIVE: To investigate current GDP oral surgery referral patterns given the anticipated change since the introduction by the General Dental Council of the specialty of surgical dentistry. DESIGN: Postal questionnaire. SETTING: 400 GDPs in Greater Manchester. RESULTS: 84% participation rate. 69% of dentists made a referral because of anticipated difficulty of surgery and 49% because of the complex nature of the patients' medical history. Practitioners who had undergone some oral surgery postgraduate training were more likely to undertake more surgery in their practices (P < 0.01) and to refer more patients for specialist care (P < 0.05). While female practitioners rated their own surgical confidence less highly than male practitioners (P < 0.001), and younger practitioners less than their older colleagues (P < 0.05), there was no significant difference in the number of referrals made. CONCLUSION: The most common reasons for referral were the anticipated difficulty of surgery and patient medical compromise. There was a wide variation between practitioners in the number of patients referred for specialist care. Postgraduate oral surgery training was identified as a factor contributing to this variation. Other practitioner variables, such as sex, experience and type of practice were not found to contribute. PMID- 10718001 TI - Prevalence of caries and developmental defects of enamel in 9-10 year old children living in areas in Brazil with differing water fluoride histories. AB - AIMS: To assess the prevalences of caries, of developmental defects of enamel and their interrelationship in Brazilian 9-10-year-olds from areas of contrasting fluoridation histories. METHODS: Systematic random sampling procedures were used to select children from an area where water had been fluoridated in 1963 and from a second area where water had been fluoridated since 1998. Clinical examinations for caries were carried out using the DMFT index and WHO diagnostic criteria. Developmental defects of enamel on upper incisors were diagnosed using the DDE index. RESULTS: A difference of 40% in DMFT was observed, with a lower prevalence of disease in the area fluoridated since 1963. Diffuse opacities affected 14.3% of the children from the area fluoridated since 1963 compared with only 2.4% in the area fluoridated in 1998. Children living in the area fluoridated in 1963 who had diffuse defects had twice the chance of being free from caries compared with those living in the same area who had no defects or who had only demarcated or hypoplastic defects. CONCLUSIONS: This study confirms previous ones in showing the benefits of water fluoridation. Diffuse opacities of upper incisors affected relatively few subjects in either of the two areas. PMID- 10718002 TI - A survey of hygienists qualifying from the Liverpool School of Dental Hygiene 1977-1998. AB - OBJECTIVES: To analyse the working patterns of all those who qualified from the Liverpool School of Dental Hygiene over a 20-year period. To assess the proportion who give up practice, the degree of part-time work, career breaks, job satisfaction, availability of continuing professional education etc. METHOD: A questionnaire sent to all 226 hygienists who qualified from the School between 1997 and 1998, whether still enrolled as dental hygienists or not. RESULTS: Responses were received from 83% of whom 89% were still working as hygienists, the majority in general practice. 46% had taken an employment break, mostly for maternity reasons but a significant number for other reasons. Around 80% expressed good job satisfaction. Although there is a high level of part-time work, especially after career breaks, few had experienced difficulty in finding employment. One third of respondents considered that the availability of continuing professional education was 'poor' or 'very poor'. CONCLUSIONS: The great majority of hygienists enjoy good job satisfaction and work in more than one general dental practice. Comparisons suggest that they continue to work in their chosen career at least as much as female dentists do in theirs. The reasons for this are thought to be that hygienists tend to be recruited from the ranks of highly motivated, mature dental nurses and that the profession lends itself to part-time work which can be combined with family commitments. There are perceived deficiencies in the availability of continuing professional education, which may be remedied by the development of distance learning packages and Section 63 type courses designed specifically for them. PMID- 10718004 TI - Methods of corporate branding: what's happening? PMID- 10718003 TI - Views of academic dentists about careers in academic dentistry in the United Kingdom. AB - The aim of this paper is to report the views of academic dentists about careers in academic dentistry assessed by method of a postal questionnaire survey. The subjects of the survey were dentists in academic posts in the United Kingdom. The incentives in pursuing an academic career which respondents rated most highly were the opportunity to teach and the variety of work in an academic career. The greatest disincentives were competing pressures from service work, teaching and research, and the difficulty of getting research grants. Many would like to spend more time on research and less on service work and teaching. The length of time required for training, and the quality of training, was a concern, particularly for junior academics. Most respondents rated the enjoyment of their job highly but scored much lower on satisfaction with the time their job left for domestic and leisure activities. By contrast with academic medicine, in academic dentistry there is typically greater emphasis on teaching and less on research. In conclusion, the balance of activities in academic posts, particularly between service work, teaching and research, needs to be regularly reviewed. The development of a more structured training programme for junior academics, which does not disadvantage academic dentists when compared with their NHS colleagues, may be required. PMID- 10718005 TI - Three cases of toxigenic Corynebacterium ulcerans infection. PMID- 10718006 TI - Fatal neonatal Salmonella meningitis linked to pet reptile. PMID- 10718007 TI - Malaria in Mexico and the Dominican Republic. PMID- 10718008 TI - Entanglement of guide wires by vena cava filters during central venous catheter insertion: report of three cases and a review of the literature. AB - Since 1993, 14 cases of central line guide wires becoming entangled with vena cava filters have been reported. We present three additional cases and review the 14 cases in the literature. Obtaining a detailed patient history is important in identifying patients with a vena cava filter. A low threshold of suspicion is needed and immediate radiograph obtained. Entangled guide wires required fluoroscopic manipulation and or retrieval of the dislodged filter. Of all reported cases, only one sustained an arrhythmia. With no signs and symptoms, conservative management of the dislodged filter is a viable option. PMID- 10718010 TI - The fading art of the physical examination. PMID- 10718009 TI - Dalteparin sodium injection treatment in patients with immunologic recurrent pregnancy loss. AB - Dalteparin sodium injection in patients with an immunologic component to RPL resulted in an increase in compliance without any adverse early pregnancy outcomes. PMID- 10718011 TI - Aberfan and the management of trauma. AB - This paper is a case study of the management of post-disaster trauma in 1960s Britain. It explores the traumatic aftermath of the 1966 Aberfan disaster (where 116 children and 28 adults were killed when a colliery spoil heap collapsed on top of a school in a small Welsh mining community) which had a devastating impact on the village. The professional and voluntary services made available to help the bereaved, survivors and wider community are documented and assessed. The paper demonstrates how limited finance and the popular and professional contemporary understanding of trauma and disasters hindered those services, and how the actions of government and media had a negative impact on the community's recovery. This case study of disaster management in the 1960s illustrates many of the pitfalls that continue to haunt the response to man-made tragedies in the UK. PMID- 10718012 TI - Cyclone mitigation, resource allocation and post-disaster reconstruction in south India: lessons from two decades of research. AB - This paper opens with a history of development and disaster-prevention strategies in a cyclone-prone area of the east coast of India and traces the evolution in the area of British and Indian governments' programmes and policy over a century. Research over the last 20 years has shown, however, that the programmes and policies have failed to balance economic growth with safety. Resources intended for the benefit of all have been diverted by alliances of powerful people to a small minority, and recent developments have reduced the physical protection of the area. The result is that increasing numbers of people are vulnerable to the effects of cyclones and floods. The findings suggest that the best way to reduce vulnerability is to improve the socio-economic standing of the most vulnerable and for this to happen these people must have an assured income based on assets that will enable them to acquire social and economic credit-worthiness within the local economy. This paper presents evidence that suggests that non-governmental organisation (NGO)-supported co-operatives are the best way to achieve this through self-help and self-employment schemes. It also suggests that NGOs should be encouraged to take up environmentally and ecologically beneficial activities involving the poorest groups in the communities, in this way combining sustained self-employment with environmental protection. PMID- 10718013 TI - Gender and drought: experiences of Australian women in the drought of the 1990s. AB - A unique collaborative, sociological study undertaken during 1995-7, explored the social construction of drought as a disaster, looking at farm families in two Australian states: Queensland (beef producers) and New South Wales (sheep/wheat producers). A decision was made to interview the women and men separately to test our hypothesis that there would be gender issues in any analysis of a disaster, but particularly one which has had so much long-term impact on individuals, families and communities, such as drought. Interviews were conducted with over 100 individuals male and female. We conclude that drought as a disaster is a gendered experience. The paper draws on the narratives of some women involved in the study to identify 'themes of difference' which confirm the necessity to maintain gender as a variable in all studies of the social impacts of disaster. PMID- 10718014 TI - Female adolescents and their sexuality: notions of honour, shame, purity and pollution during the floods. AB - This paper explores the experiences of female adolescents during the 1998 floods in Bangladesh, focusing on the implications of socio-cultural norms related to notions of honour, shame, purity and pollution. These cultural notions are reinforced with greater emphasis as girls enter their adolescence, regulating their sexuality and gender relationships. In Bangladeshi society, adolescent girls are expected to maintain their virginity until marriage. Contact is limited to one's family and extended relations. Particularly among poorer families, adolescent girls tend to have limited mobility to safeguard their 'purity'. This is to ensure that the girl's reputation does not suffer, thus making it difficult for the girl to get married. For female adolescents in Bangladesh, a disaster situation is a uniquely vulnerable time. Exposure to the unfamiliar environment of flood shelters and relief camps, and unable to maintain their 'space' and privacy from male strangers, a number of the girls were vulnerable to sexual and mental harassment. With the floods, it became difficult for most of the girls to be appropriately 'secluded'. Many were unable to sleep, bathe or get access to latrines in privacy because so many houses and latrines were under the water. Some of the girls who had begun menstruation were distressed at not being able to keep themselves clean. Strong social taboos associated with menstruation and the dirty water that surrounded them made it difficult for the girls to wash their menstrual cloths or change them frequently enough. Many of them became separated from their social network of relations, which caused them a great deal of anxiety and stress. Their difficulty in trying to follow social norms have had far reaching implications on their health, identity, family and community relations. PMID- 10718015 TI - Predictors for people's response to a tornado warning: Arkansas, 1 March 1997. AB - On 1 March 1997, powerful tornadoes touched down in Arkansas (USA) on a Saturday afternoon. Twenty-six fatalities and 400 non-fatal injuries were reported. We performed a population-based cross-sectional study to determine factors associated with appropriate responses to tornado warnings. Of 146 survey participants, 140 (96 per cent) knew the difference between 'tornado watch' and 'tornado warning' and were aware of when the warning was announced. Of those 140 participants, 64 (45.7 per cent) responded to the warning by seeking shelter, and 58 (90.6 per cent) of those 64 acted within five minutes of hearing the warning. Four factors were positively associated with those seeking shelter: having graduated from high school (OR = 4.2, 95 per cent CI = 1.1-15.5); having a basement in one's house (OR = 3.8, 95 per cent exact CI = 1.1-17.1); hearing a siren (OR = 4.4, 95 per cent CI = 1.3-18.9); and having prepared a household plan of response when tornadoes occur (OR = 2.6, 95 per cent CI = 1.1-6.3). On the basis of these findings, we recommend: first, that people who live in tornadoprone areas have a personal plan of action to help them respond immediately to warnings; second, public-health education officials in areas with frequent tornadic activity should do more to educate the public about what they can do to protect themselves from a tornado; and third, that emergency-management officials planning protection measures for vulnerable communities should consider that most people have limited time (our study documented five minutes) in which to respond to a tornado warning. Thus, shelters in tornado-prone areas should be quickly accessible by residents. PMID- 10718016 TI - Perpetual problem to plague those who study and mitigate disaster. PMID- 10718017 TI - Medicare & drugs: the elusive prize revisited. PMID- 10718018 TI - The Medicare prescription drug benefit: how will the game be played? AB - Most recent proposals to add a prescription drug benefit to the Medicare program suggest using pharmacy benefit managers (PBMs) to control costs and promote quality. However, the proposals give little detail on the institutional arrangements that would govern PBM operations and drug procurement. The recent Congressional Budget Office cost estimate of the Clinton administration's proposal reflects this lack of detail on how PBMs would function. We sketch an approach for structuring PBM operations that focuses on competition among PBMs, manufacturers, and distributors; incentive pricing; and risk sharing with PBMs. PMID- 10718019 TI - Pharmaceutical benefit management: an alternative approach. PMID- 10718020 TI - Designing a Medicare prescription drug benefit: issues, obstacles, and opportunities. AB - We review the policy concerns underlying some of the most contentious issues that must be resolved prior to the enactment of a Medicare drug benefit. We consider critical issues both in benefit design-targeted versus universal eligibility, benefit subsidies, and benefit comprehensiveness--and in benefit administration, focusing especially on issues involving the administration of the drug benefit in traditional Medicare. Despite the apparent contentiousness of the drug benefit debate, alternative proposals may not be so far apart on these issues. PMID- 10718021 TI - Managing the pharmacy benefit in Medicare HMOs: what do we really know? AB - An estimated five million Medicare beneficiaries received outpatient prescription drug benefits through Medicare + Choice in 1999. However, little is known about how these benefits are managed or about their effects on costs and quality of care. This exploratory study applies a case-study methodology to four large Medicare health maintenance organizations (HMOs) to identify and assess drug-use management strategies. It also poses a number of important issues for consideration by both policymakers and health services researchers, as the debate rages on over the creation and administration of a Medicare outpatient drug benefit, especially in light of the predilection for the use of private pharmacy benefit managers (PBMs) in many of these proposals. PMID- 10718022 TI - Medicare pharmacy coverage: ensuring safety before funding. PMID- 10718023 TI - Prescription drugs and managed care: can 'free-market detente' hold? AB - The rapid rise in pharmaceutical benefits costs, often cited as a major contributor to the resurgence in health care cost growth, is beginning to strain the relationship between the pharmaceutical and the managed care industries in the United States. In interviews conducted in 1999, executives from both industries maintained a continued preference for a market-based resolution of these tensions. There is evidence, however, that this private-sector detente may give way in the face of the rising business and political pressures that both industries face. Active leadership will be required to prevent deterioration of the prevailing political climate toward economic controls. PMID- 10718024 TI - Just what the HMO ordered: the paradox of increasing drug costs. AB - Drug companies argue that newer, more expensive drugs offset other medical costs; health plans counter that they increase pharmacy costs more than they offer a "pharmacoeconomic" benefit. Neither side is universally right or wrong, and neither has the data to support its case. Increasing drug costs for selective therapeutic classes represent the fulfillment of managed care's original promise. Certain therapeutic classes of drugs offer pharmacoeconomic benefit, while others represent induced costs in excess of this benefit. Health maintenance organizations (HMOs) should determine one from the other and incorporate these findings into their plan designs; multitier drug coverage is the best method to achieve this. PMID- 10718025 TI - Are pharmaceuticals cost-effective? A review of the evidence. AB - The argument that prescription drugs are cost-effective has been made both by the pharmaceutical industry to support rising drug prices and expenditures, and by advocates of expanded drug coverage for elderly and low-income persons. A new database of 228 published cost-utility analyses sheds light on the issue. According to published data, some drugs do save money or are cost-effective, but the issue depends critically on the context in which the drug is used and the intervention with which it is compared. Cost-utility analyses funded by the drug industry tend to report more favorable results than do those funded by nonindustry sources. Cost-effectiveness analysis can help policymakers to determine whether drugs and other interventions offer value for money. PMID- 10718026 TI - Direct-to-consumer prescription drug advertising: trends, impact, and implications. AB - We provide an overview of what is known about the impact of direct-to-consumer (DTC) advertising of prescription drugs. Specifically, we explore the historical trends that led to the industry's increasing use of this form of promotion. Then, using the published literature to date, we review the impact of DTC advertising on the consumer, the medical profession, and the health care system. We conclude by offering policy suggestions for how the pharmaceutical industry can promote its products more responsibly, how the Food and Drug Administration (FDA) can regulate DTC advertising more effectively, and how the medical and public health communities can educate the public about drug therapies more constructively. PMID- 10718027 TI - The industrialization of clinical research. AB - Recent controversies over the protection of human subjects, payment of physicians for recruiting patients to clinical trials, Food and Drug Administration (FDA) removal of approved drugs from the market, and reporting of results of clinical trials have highlighted important facets of clinical research. Less visible has been the industrialization of clinical research, and especially of clinical trials, that is, its emergence as a "line of business" of substantial magnitude and rapid growth. The growth of drug-industry outsourcing of clinical trials and the concomitant rise of a contract research industry are described in this paper, which argues for greater transparency in the conduct of both publicly and privately sponsored clinical trials. PMID- 10718028 TI - The economics of Viagra. PMID- 10718029 TI - Selling teaching hospitals to investor-owned hospital chains: three case studies. PMID- 10718030 TI - Public hospitals: privatization and uncompensated care. PMID- 10718031 TI - The geography of health insurance regulation. AB - The health insurance market consists of three distinct segments--individual, small group, and large group--each governed by different economic and regulatory structures. A number of border-crossing techniques have arisen for avoiding the burdens of one segment and capitalizing on the benefits of others. Drawing from extensive qualitative research into the functioning of existing market structures, this paper describes these techniques and their purposes and effects. This road map helps to identify which reform proposals seek to produce true economic efficiencies and which have the potential to undermine previous reform objectives. PMID- 10718032 TI - Kaiser Permanente's prescription drug benefit. PMID- 10718033 TI - Voices from the clinic: AIDS then and now. PMID- 10718034 TI - Beyond survey data: a claims-based analysis of drug use and spending by the elderly. AB - Previous estimates of Medicare beneficiaries total and out-of-pocket spending on outpatient prescription drugs have largely been based on data from the 1995 Medicare Current Beneficiary Survey and have focused on how expenditures vary among beneficiaries with different demographic characteristics. This paper reports the results of an analysis of prescription claims from 1998 for more than 375,000 elderly persons whose prescription benefit was managed by Merck-Medco Managed Care. In addition to examining how total and out-of-pocket drug spending in a well-insured population varies by age and sex, we report how total and condition-specific drug spending varies for elderly persons with ten common chronic diseases. Our results illustrate the highly skewed nature of prescription drug spending, even among those with drug coverage, and underscore the particularly high cost burden that pharmaceuticals place on elderly people with chronic diseases. PMID- 10718035 TI - The costs of a Medicare prescription drug benefit. AB - This paper describes a preliminary cost estimate, prepared by the Congressional Budget Office (CBO), of President Clinton's 1999 prescription drug benefit proposal. The CBO estimated that the new benefit would increase net Medicare outlays by $136 billion between 2002 and 2009, although these estimates are highly uncertain. Because the proposal included an annual cap on the amount of the benefit, it did not require consideration of an important effect of a more comprehensive benefit: higher prices for some drugs. Estimates of future proposals for a Medicare prescription drug benefit may require consideration of that pricing effect. PMID- 10718036 TI - Drug coverage and drug purchases by Medicare beneficiaries with hypertension. AB - Research has shown that older Americans with prescription drug coverage purchase more medications; however, it is unclear whether they are more likely to purchase essential medications. This study addresses that question by examining the relationship between drug coverage and medication purchases among older Americans with hypertension. It finds that drug coverage has a significant impact: It lowers the likelihood that persons with hypertension will go without antihypertensive drugs, and it raises the number of tablets purchased among those who purchase these essential drugs. PMID- 10718037 TI - Explaining drug spending trends: does perception match reality? AB - Several recent studies have made clear that drug expenditures are rising more rapidly than other health care spending. What has not been clear, however, is how much drug spending is driven by price rather than volume and whether volume increases are appropriate. This DataWatch takes a closer look at the components and drivers of drug spending using large claims databases from managed care and employer-sponsored health benefit plans. In both environments this study found volume, not price, to be the largest driver of drug spending for seven diseases studied. For four of the diseases, we review the clinical issues that may have influenced volume growth. PMID- 10718038 TI - Drug coverage decisions: the role of dollars and values. AB - Given the increasing costs of pharmaceuticals today, it is important to understand how pharmacy benefits decisions are made and the role of cost and values in these decisions. This study examines what coverage decisions insurers make and the information and processes used in making these decisions. Fifty three organizations, differing in size, tax status, and region, were asked about their policies for four new and controversial drugs: Viagra, Enbrel, Zyban, and Celebrex. Enbrel and Celebrex were much more likely to be covered than Viagra and Zyban. In addition, coverage of Enbrel and Celebrex was limited through strategies such as prior authorization, to encourage medically appropriate use of these agents, whereas coverage of Viagra and Zyban was limited predominantly through generalized exclusion or through restrictions on quantity or duration of use. Value judgments, rather than cost, seem to play a central, though largely unspoken, role in these coverage decisions. PMID- 10718039 TI - Medicare beneficiaries and drug coverage. AB - Whether or not to add a prescription drug benefit to the basic Medicare package is at the forefront of congressional debate. Using data from the 1996 Medicare Current Beneficiary Survey (MCBS), we examine changes in drug insurance coverage levels from 1995 to 1996 and compare drug use and spending data for Medicare beneficiaries with and without drug coverage. The data show the enrollees without drug insurance consistently use fewer prescriptions, spend more out of pocket, and have less in total drug expenditures than their insured peers. PMID- 10718040 TI - A closer look: profiling foundations created by health care conversions. PMID- 10718041 TI - PBMs: a key player in Medicare drug coverage. PMID- 10718042 TI - Patient education and protection through PBMs. PMID- 10718043 TI - Public spending for health care approaches 60 percent. PMID- 10718044 TI - Role of retiree benefits in health insurance's future. PMID- 10718045 TI - Taking a giant leap forward in promoting quality. PMID- 10718046 TI - Emergency care as safety net. PMID- 10718047 TI - Experiments with CHRELMS patient-driven stimulator controllers for the restoration of function to paralysed legs. AB - This paper examines the control of electrical stimulators for restoring the standing function to paraplegics. Patient-driven controllers are based on the knowledge that standing, using support handles, is guided by the neurologically intact natural motor control system, acting through the arms. One category of patient-driven controller is CHRELMS (Control by Handle Reactions of Leg Muscle Stimulation). The first experiments with patient-driven controllers are reported. Experiments in the CHRELMS control of one volunteer are described. Proportional control did not allow the subject to stand up because of the strategy he used, but integral control was successful. With it, he could stand up and sit down slowly, entirely guided by himself, as well as making other manoeuvres. Without changing controller mode, and by leaning forwards and backwards, he could 'posture switch' his support muscles. These are significant achievements and justify a larger study of patient-driven controllers. The subject's control strategies are discussed. PMID- 10718048 TI - Biological reactions to wear debris in total joint replacement. AB - The vast majority of total hip prostheses currently implanted consist of a hard metal or ceramic femoral head articulating against an ultra-high molecular weight polyethylene (UHMWPE) acetabular cup. Over the last 10 years, evidence has accumulated to show that these prostheses are prone to failure due to late aseptic loosening and few survive beyond 25 years. With an increasing need to implant hip prostheses in the younger, more active patient the need to understand the mechanisms of failure and to develop artificial hip joints using alternative materials have become major issues in the orthopaedic community. This review focuses initially on our current understanding of the biological reactions to UHMWPE prosthetic wear debris in vivo and in vitro since this is believed to be the main cause of late aseptic loosening. While the precise mechanisms of osteolysis induced by UHMWPE wear debris have not been elucidated, the major message to emerge is that it is not the wear volume that determines the biological response to the debris, but the concentration of the wear volume that is within the critical size range (0.2-0.8 micron) for macrophage activation. The review then considers whether the problem of wear-debris-induced osteolysis may be overcome with the use of new generation metal-on-metal or ceramic-on-ceramic prostheses. For metal-on-metal prostheses, the prospects for increasing the osteolysis free life of the implant are good but additional biological problems associated with the nanometre size and reactivity of the wear particles in vivo may emerge. For the ceramic-on-ceramic prostheses, although initial prospects are encouraging, more data are needed on the characteristics of the wear particles generated in vivo before predictions can be made. It is concluded that the pre clinical testing of any new materials for joint replacement must include an analysis of the wear particle characteristics and their biological reactivity in addition to the usual assessment of wear. PMID- 10718049 TI - A comparative joint simulator study of the wear of metal-on-metal and alternative material combinations in hip replacements. AB - While total hip replacement represents the major success story in orthopaedic surgery in the twentieth century, there is much interest in extending even further, early in the twenty first century, the life of implants. Osteolysis has been identified as a major factor limiting the life of prostheses, with indications that fine polyethylene wear debris, generated primarily at the interface between the femoral head and the acetabular cup, promotes the process. There is therefore considerable interest in the introduction of alternative wear resistant systems to limit the deleterious effects of wear. These alternatives include ceramic-on-ceramic and metal-on-metal configurations and the present paper is primarily concerned with the latter. Some six pairs of new metal-on metal implants of 36 mm diameter and four pairs of existing metal-on-metal implants of 28 mm diameter were tested in a ten-station hip joint simulator in the presence of a 25 per cent bovine serum solution. The implants were tested in the anatomical position to 5 x 10(6) cycles. The new heads and cups were manufactured from CoCrMo alloy with careful attention being paid to sphericity and surface finish of both components. The wear performance of the new and existing metal-on-metal total hip replacements have been evaluated and compared. The overall wear rates have then been compared with previously reported wear rates for a zirconia-on-polyethylene prosthesis of 22 mm diameter tested on the same simulator. The comparison is taken further by recalling published penetration data for metal-on-polyethylene implants of 22 and 28 mm diameter and converting these to volumetric wear rates. It was found that the heads and cups in metal-on-metal joints wore by almost equal amounts and that the opposing surfaces converged to similar surface roughness as the testing time increased. Steady state wear rates were generally achieved after 1-2 x 10(6) cycles. The mean long-term wear rates for the metal-on-metal prostheses were very low, being 0.36 mm3/10(6) cycles and 0.45 mm3/10(6) cycles for the new implants of 36 mm diameter and established implants of 28 mm diameter respectively. These wear rates compare with 6.3 mm3/10(6) cycles for zirconia-on-ultra-high molecular weight polyethylene tested on the same simulator and representative clinical values for metal-on-polyethylene of 36 mm3/year for heads of 22 mm diameter and a reported range of 60-180 mm3/year for 28 mm heads. These values do not translate directly into numbers of particles, since the metallic debris from metal-on-metal joints is very fine. The number of metallic particles may exceed the number of polyethylene wear particles from an otherwise similar metal-on-polyethylene joint by a factor of 10(3). A detailed discussion of the size and morphology of wear debris and tissue reaction to various forms of debris is beyond the scope of this paper, but the biological response to polymeric, metallic and ceramic wear debris forms a major subject for further study. The present investigation nevertheless confirms the potential of carefully designed and manufactured metal-on-metal total replacement joints for the treatment of diseased and damaged hips. PMID- 10718050 TI - Comparison of friction and lubrication of different hip prostheses. AB - It is well documented that an important cause of osteolysis and subsequent loosening of replacement hip joints is polyethylene wear debris. To avoid this, interest has been renewed in metal-on-metal and ceramic-on-ceramic prostheses. Various workers have assessed the lubrication modes of different joints by measuring the friction at the bearing surfaces, using different lubricants. Measurements of friction factors of a series of hip prostheses were undertaken using carboxymethyl cellulose (CMC) fluids, silicone fluids, synovial fluid and different concentrations of bovine serum as the lubricant. The experimental results were compared with theoretical predictions of film thicknesses and lubrication modes. A strong correlation was observed between experiment and theory when employing CMC fluids or silicone fluids as the lubricant. Mixed lubrication was found to occur in the metal-on-metal (CoCrMo/CoCrMo) joints with all lubricants at a viscosity within the physiological range. This was also the case for the metal-on-plastic (CoCrMo/ultra-high molecular weight polyethylene) joints. The ceramic-on-ceramic (Al2O3/Al2O3) joints, however, exhibited full fluid film lubrication with the synthetic lubricants but mixed lubrication with the biological lubricants. Employing a biological fluid as the lubricant affected the friction to varying degrees when compared with the synthetic lubricants. In the case of the ceramic-on-ceramic joints it acted to increase the friction factor tenfold; however, for the metal-on-metal joints, biological fluids gave slightly lower friction than the synthetic lubricants did. This suggests that, when measuring friction and wear of artificial joints, a standard lubricant should be used. PMID- 10718051 TI - Development of artificial articular cartilage. AB - Attempts have been made to develop an artificial articular cartilage on the basis of a new viewpoint of joint biomechanics in which the lubrication and load bearing mechanisms of natural and artificial joints are compared. Polyvinyl alcohol hydrogel (PVA-H), 'a rubber-like gel', was investigated as an artificial articular cartilage and the mechanical properties of this gel were improved through a new synthetic process. In this article the biocompatibility and various mechanical properties of the new improved PVA-H is reported from the perspective of its usefulness as an artificial articular cartilage. As regards lubrication, the changes in thickness and fluid pressure of the gap formed between a glass plate and the specimen under loading were measured and it was found that PVA-H had a thicker fluid film under higher pressures than polyethylene (PE) did. The momentary stress transmitted through the specimen revealed that PVA-H had a lower peak stress and a longer duration of sustained stress than PE, suggesting a better damping effect. The wear factor of PVA-H was approximately five times that of PE. Histological studies of the articular cartilage and synovial membranes around PVA-H implanted for 8-52 weeks showed neither inflammation nor degenerative changes. The artificial articular cartilage made from PVA-H could be attached to the underlying bone using a composite osteochondral device made from titanium fibre mesh. In the second phase of this work, the damage to the tibial articular surface after replacement of the femoral surface in dogs was studied. Pairs of implants made of alumina, titanium or PVA-H on titanium fibre mesh were inserted into the femoral condyles. The two hard materials caused marked pathological changes in the articular cartilage and menisci, but the hydrogel composite replacement caused minimal damage. The composite osteochondral device became rapidly attached to host bone by ingrowth into the supporting mesh. The clinical implications of the possible use of this material in articular resurfacing and joint replacement are discussed. PMID- 10718052 TI - Analytical modelling of the elastomeric layer in soft layer hip replacements. AB - A mechanical analysis is carried out for a flat deformable layer, firmly anchored to a rigid substrate, and frictionlessly compressed by a rigid spherical indenter, with reference to the design of hip replacements whose cup is covered by an elastomeric layer. An available perturbation solution of differential character, developed for a layer subject to plane strain conditions, indented by a cylinder approximated as parabolic in shape and valid for high contact widths, is here extended to the axisymmetric problem of a layer compressed by a paraboloidal indenter. In addition to the paraboloidal idealization, an accurate mathematical description of the indenter spherical profile is incorporated into the modelling of this contact problem. Perturbed solutions up to the second order are developed for both compressible and incompressible layers. Selected numerical examples addressing actual joint geometries are included to assess the validity of the proposed analytical modelling, to explore the accuracy in describing the spherical profile and to compare the contact pressure profiles activated by paraboloidal and spherical indenters. PMID- 10718053 TI - Boundary lubrication in vivo. AB - Evidence is reviewed for the concept that the body employs essentially the same lubrication system in many sites in the body where tissues slide over each other with such ease. This system consists of fluid adjacent to surfaces coated with an oligolamellar lining of surface-active phospholipid (SAPL) acting as a back-up boundary lubricant wherever the fluid film fails to support the load--a likely event at physiological velocities. Particular attention is paid to the load bearing joints, where the issue of identifying the vital active ingredient in synovial fluid is reviewed, coming down--perhaps predictably--in favour of SAPL. It is also explained how Lubricin and hyaluronic acid (HA) could have 'carrier' functions for the highly insoluble SAPL, while HA has good wetting properties needed to promote hydrodynamic lubrication of a very hydrophobic articular surface by an aqueous fluid wherever the load permits. In addition to friction and wear, release is included as another major role of boundary lubricants, especially relevant in environments where proteins are found, many having adhesive properties. The discussion is extended to a mention of the lubrication of prosthetic implants and to disease states where a deficiency of boundary lubricant is implicated, particular attention being paid to osteoarthritis. PMID- 10718054 TI - Tissue engineering: confronting the transplantation crisis. AB - Tissue engineering is the development of biological substitutes and/or the fostering of tissue regeneration/remodelling. It is emerging as a technology which has the potential to confront the crisis in transplantation caused by the shortage of donor tissues and organs. With the development of this technology, ther is emerging a new industry which is at the interface of biotechnology and the traditional medical implant field. For this technology and the associated industry to realize their full potential, there are core, enabling technologies that need to be developed. This is the focus of the Georgia Tech/Emory Center for the Engineering of Living Tissues, newly established in the United States, with an Engineering Research Center Award from the National Science Foundation. With the development of these core technologies, tissue engineering will evolve from an art form to a technology based on science and engineering. PMID- 10718055 TI - Design forms of total knee replacement. AB - The starting point of this article is a general design criterion applicable to all types of total knee replacement. This criterion is then expanded upon to provide more specifics of the required kinematics, and the forces which the total knee must sustain. A characteristic which differentiates total knees is the amount of constraint which is required, and whether the constraint is translational or rotational. The different forms of total knee replacement are described in terms of these constraints, starting with the least constrained unicompartments to the almost fully constrained fixed and rotating hinges. Much attention is given to the range of designs in between these two extreme types, because they constitute by far the largest in usage. This category includes condylar replacements where the cruciate ligaments are preserved or resected, posterior cruciate substituting designs and mobile bearing knees. A new term, 'guided motion knees', is applied to the growing number of designs which control the kinematics by the use of intercondylar cams or specially shaped and even additional bearing surfaces. The final section deals with the selection of an appropriate design of total knee for specific indications based on the design characteristics. PMID- 10718056 TI - A new method for quantitative evaluation of perceived sounds from mechanical heart valve prostheses. AB - Closing clicks from mechanical heart valve prostheses are transmitted to the patient's inner ear mainly in two different ways: as acoustically transmitted sound waves, and as vibrations transmitted through bones and vessels. The purpose of this study was to develop a method for quantifying what patients perceive as sound from their mechanical heart valve prostheses via these two routes. In this study, 34 patients with implanted mechanical bileaflet aortic and mitral valves (St Jude Medical and On-X) were included. Measurements were performed in a specially designed sound insulated chamber equipped with microphones, accelerometers, preamplifiers and a loudspeaker. The closing sounds measured with an accelerometer on the patient's chest were delayed 400 ms, amplified and played back to the patient through the loudspeaker. The patient adjusted the feedback sound to the same level as the 'real-time' clicks he or she perceived directly from his or her valve. In this way the feedback sound energy includes both the air- and the bone-transmitted energies. Sound pressure levels (SPLs) were quantified both in dB(A) and in the loudness unit sone according to ISO 532B (the Zwicker method). The mean air-transmitted SPL measured close to the patient's ear was 23 +/- 4 dB(A). The mean air- and bone-transmitted sounds and vibrations were perceived by the patients as an SPL of 34 +/- 5 dB(A). There was no statistically significant difference in the perceived sound from the two investigated bileaflet valves, and no difference between aortic and mitral valves. The study showed that the presented feedback method is capable of quantifying the perceived sounds and vibrations from mechanical heart valves, if the patient's hearing is not too impaired. Patients with implanted mechanical heart valve prostheses seem to perceive the sound from their valve two to three times higher than nearby persons, because of the additional bone-transmitted vibrations. PMID- 10718057 TI - A review of robotics in surgery. AB - A brief introduction is given to the definitions and history of surgical robotics. The capabilities and merits of surgical robots are then contrasted with the related field of computer assisted surgery. A classification is then given of the various types of robot system currently being investigated internationally, together with a number of examples of different applications in both soft-tissue and orthopaedic surgery. The paper finishes with a discussion of the main difficulties facing robotic surgery and a prediction of future progress. PMID- 10718058 TI - Associative priming in Pavlovian conditioning. AB - Any occasion on which an animal is placed in an experimental setting or context and receives pairings of one event with another provides the opportunity for a variety of associative structures to be acquired. These structures range from simple associations, which allow the presentation of one event to activate or prime a memory of the other, to hierarchical associations, which allow a simple association to be primed by some other event (e.g. the context in which the simple association was acquired). Experiments with rats that reveal priming effects consistent with both of these putative associative structures are reviewed. PMID- 10718059 TI - Recent and remote extinction cues reduce spontaneous recovery. AB - Five appetitive conditioning experiments with rats examined the ability of extinction cues (ECs) to reduce spontaneous recovery after extinction procedures that varied the temporal relation between the ECs and the conditioned stimulus (CS) and included presentations of additional events (e.g. other stimuli correlated with extinction, CSs, and the US). In extinction, two different ECs were presented either closely in time before (i.e. recent to), or more distant (i.e. remote) from a nonreinforced CS. Both recent and remote ECs reduced spontaneous recovery to the CS when present during testing (Experiments 1-4). Each EC reduced recovery despite the addition during extinction of a second EC and CS (Experiments 1, 3, and 4), and the US (Experiment 3). Experiments 4 and 5 investigated the recent and remote ECs' tendency to control a serial occasion setting discrimination involving the target CS under explicit training conditions. Neither EC gained such discriminative control. Possible explanations of the results are discussed, including configural learning, occasion setting, and contextual cue control. PMID- 10718060 TI - Super-learning of causal judgements. AB - In three experiments, participants learned which of a variety of foods were capable of causing an allergic reaction in a hypothetical patient during training in which a compound of a treatment and a target food cue was paired with the reaction. In Experiment 1 the causal ratings of the target cue were increased if the treatment cue was pretrained as a preventative cause of the reaction. Experiments 2 and 3 demonstrated that the magnitude of this super-learning is unaffected by the order of compound and treatment cue training. The final study also showed that forward super-learning is not induced solely by simple exposure to the treatment cue prior to compound training but, rather, depends upon training the treatment cue as a preventative cause, whereas retrospective super learning may be due merely to exposure of the treatment cue. These results are problematic for contingency-based accounts of causal induction but accord with modified and extended associative theories. PMID- 10718061 TI - The word-length effect and disyllabic words. AB - Three experiments compared immediate serial recall of disyllabic words that differed on spoken duration. Two sets of long- and short-duration words were selected, in each case maximizing duration differences but matching for frequency, familiarity, phonological similarity, and number of phonemes, and controlling for semantic associations. Serial recall measures were obtained using auditory and visual presentation and spoken and picture-pointing recall. In Experiments 1a and 1b, using the first set of items, long words were better recalled than short words. In Experiments 2a and 2b, using the second set of items, no difference was found between long and short disyllabic words. Experiment 3 confirmed the large advantage for short-duration words in the word set originally selected by Baddeley, Thomson, and Buchanan (1975). These findings suggest that there is no reliable advantage for short-duration disyllables in span tasks, and that previous accounts of a word-length effect in disyllables are based on accidental differences between list items. The failure to find an effect of word duration casts doubt on theories that propose that the capacity of memory span is determined by the duration of list items or the decay rate of phonological information in short-term memory. PMID- 10718062 TI - Effects of gender marking in pronominal coindexation. AB - The naming latency of a pronoun was measured when a single previously presented name in a discourse either agreed or did not agree with the pronoun in gender and person. An effect of agreement was found both under conditions in which subjects were likely to have engaged in strategic processing of the pronoun (Experiment 1) and under conditions in which this was unlikely (Experiment 3). The effect of gender agreement was also investigated when two noun phrases were present in the discourse. The results continued to show an immediate effect of gender agreement (naming latencies increased when a pronoun did not agree with one of two previously presented nouns) under experimental conditions likely to engender strategic processing (Experiment 2). This last effect was not significant under experimental conditions that were not likely to engender strategic processing (Experiment 3). The results are discussed in terms of models of the process of identifying the referent of a pronoun in a discourse. PMID- 10718063 TI - Interference in visual working memory. AB - This paper uses the theoretical distinction that has recently developed between the passive visual store and the active spatial rehearsal mechanism of the visuo spatial component of working memory (VSSP). It examines the circumstances under which visual fields gain functional access to the passive visual store and seeks to cast light on the circumstances under which irrelevant visual fields interfere with concurrent visual processing. Experiment 1 contrasts a dynamic visual noise field with a static noise field and shows that the static field, in contrast to the dynamic noise field, causes no interference when presented concurrently with a visual task. Experiment 2 investigates the reason for this contrast and concludes that the static field is susceptible to decay and so fails to cause interference. Experiment 3 investigates further the circumstances under which dynamic visual noise causes interference and shows that manipulation of the number of changes within the noise field is also of importance in causing interference. The results allow further consideration of the characteristics of the VSSP. PMID- 10718064 TI - Proprioception and stimulus-response compatibility. AB - Sixteen subjects pressed a left or right key in response to lateralized visual stimuli, in uncrossed (left index finger on left key, right finger on right key) and crossed conditions (left finger on right key and vice versa), with varying finger separations. Visual, tactile, or "efference copy" cues about relative finger positions were unavailable. Subjects had to press the key on the same side as (compatible group) or opposite side to the stimulus (incompatible group). Separate proprioceptive judgements of the relative finger positions were obtained. Findings of an overall reaction time (RT) advantage for compatible instructions and for uncrossed hands were replicated. With decreasing finger separation the RT advantage for compatible instructions decreased, and the probability of responding with either hand increased. The compatibility effect disappeared completely at the 6-cm crossed position, not at the position that was hardest to judge proprioceptively. This suggests that two forms of neural activation are summed: automatic activation of the anatomically same-side limb, and an integrated, rule-based activation. The results further demonstrate that independent proprioceptive cues from each limb, unassociated with skin contact between the limbs, can mediate the determination of relative position for response selection in stimulus-response compatibility tasks. PMID- 10718065 TI - Hierarchical knowledge influences stimulus-response compatibility effects. AB - The influence of spatial stimulus grouping on stimulus-response compatibility effects was investigated in three experiments. Stimuli were grouped as part of a superordinate unit BY (1) perceptually organizing them (Experiment 1), (2) organizing them on the basis of semantic links (Experiment 2), or (3) arbitrary links (Experiment 3). In some instances the arrangement of the stimuli resulted in a conflict between two types of spatial relationship: one between stimulus and response and the other between superordinate unit and response. The experiments indicated that it was the latter relationship that mainly determined performance in the experiments. Reaction time analyses showed that responses were fastest if they spatially corresponded to the relative location of the superordinate unit of which the stimulus was part. The results are discussed with reference to hierarchical accounts of coding stimulus information. PMID- 10718066 TI - Interactions between exogenous auditory and visual spatial attention. AB - Six experiments were carried out to investigate the issue of cross-modality between exogenous auditory and visual spatial attention employing Posner's cueing paradigm in detection, localization, and discrimination tasks. Results indicated cueing in detection tasks with visual or auditory cues and visual targets but not with auditory targets (Experiment 1). In the localization tasks, cueing was found with both visual and auditory targets. Inhibition of return was apparent only in the within-modality conditions (Experiment 2). This suggests that it is important whether the attention system is activated directly (within a modality) or indirectly (between modalities). Increasing the cue validity from 50% to 80% influenced performance only in the localization task (Experiment 4). These findings are interpreted as being indicative for modality-specific but interacting attention mechanisms. The results of Experiments 5 and 6 (up/down discrimination tasks) also show cross-modal cueing but not with visual cues and auditory targets. Furthermore, there was no inhibition of return in any condition. This suggests that some cueing effects might be task dependent. PMID- 10718067 TI - Interference from distractors in reach-to-grasp movements. AB - Descriptions of interference effects from non-relevant stimuli are extensive in visual target detection and identification paradigms. To explore the influence of features of non-relevant objects on reach-to-grasp movements, we instructed healthy normal controls to reach for and pick up a cylinder (target) placed midsagittally 30 cm from the starting position of the hand. In Experiment 1, the target was presented alone, or accompanied by a narrower, wider, or same-size distractor positioned to the left or right of the target. In Experiment 2, the target was presented alone or accompanied by a distractor, which was slanted at a different orientation to the target. Reflective markers were placed on the wrist, thumb, and index finger of the right hand, and infra-red light-detecting cameras recorded their displacement through a calibrated 3-dimensional working space. Kinematic parameters were derived and analysed. Consistent changes in the expression of peak velocity, acceleration, and deceleration were evident when the distractor was narrower or wider than the target. The impact of the orientation of the distractor, conversely, was not marked. We discuss the results in the context of physiological findings and models of selective attention. PMID- 10718068 TI - Imitation of gestures in children is goal-directed. AB - The view that the motor program activated during imitation is organized by goals was investigated by asking pre-school children to imitate a set of hand gestures of varying complexity that were made by an experimenter sitting in front of them. In Experiments 1 and 3, children reached for the correct object (one of their own ears or one of two dots on a table) but preferred to use the ipsilateral hand. This ipsilateral preference was not observed when hand movements were made to only one ear (Experiment 2), or when movements were directed at space rather than physical objects (Experiment 3). The results are consistent with the notion that imitation is guided by goals and provide insights about how these goals are organized. PMID- 10718069 TI - The differential effects of simultaneous and successive cueing on the detection of bilateral symmetry in dot patterns. AB - Corballis and Roldan (1975) obtained speeded judgements of whether dot patterns were bilaterally symmetrical about, or translated across, a line. Reaction times (RTs) were ordered V (vertical) > D (diagonal) > H (horizontal) where ">" means faster than. Similar results occurred with blocked axis orientations, suggesting subjects cannot prepare by rotating a mental frame of reference. Blocking trials may have been ineffective because blocking cannot provide incremental benefits over those already provided by axis lines. Four experiments show that the usual axis orientation ordering of V > H > D is markedly attentuated by simultaneous but not successive axis lines. Also, axis cue lines and axis blocking are not equivalent treatments. Instead, unblocked line cues require finite processing time whereas, under blocking, subjects can prepare for the expected orientation. There was no suggestion anywhere of the V > D > H axis ordering that Corballis and Roldan reported. Successive axis line cues may only direct attention to the orientation being cued, but simultaneous line cues may change the stimulus itself, thus providing an additional means of visual processing that facilitates symmetry detection at non-vertical axis orientations. PMID- 10718070 TI - Interference from multi-dimensional objects during feature and conjunction discriminations. AB - Feature discrimination performance within an attended object and interference from irrelevant, multi-dimensional objects (distractors) were examined in a two choice, response compatibility paradigm. Results showed that the amount of interference by multi-dimensional distractors was dependent on three factors: (1) the discriminability of the incompatible, task-relevant distractor features; (2) the number of incompatible, task-relevant distractor features; and (3) whether the task-relevant, incompatible features matched the task goals. The most interesting finding was that additive priming effects were found for multiple, task-relevant features that matched the task goals, whether these features were present in the attended object or in the ignored object. Models that assume that each task-relevant feature primes its corresponding decision/response asynchronously and that this priming is combined to meet a decision/response criterion (at least when attended) can account for distractor interference during conjunction discriminations. Implications of these findings for feature integration models, template models, and a response selection model are discussed. PMID- 10718071 TI - Low reliability of perceptual priming: consequences for the interpretation of functional dissociations between explicit and implicit memory. AB - In this study, three experiments are presented that investigate the reliability of memory measures. In Experiment 1, the well-known dissociation between explicit (recall, recognition) and implicit memory (picture clarification) as a function of age in a sample of 335 persons aged between 65 and 95 was replicated. Test retest reliability was significantly lower in implicit than in explicit measures. In Experiment 2, parallel-test reliabilities in a student sample confirmed the finding of Experiment 1. In Experiment 3, the reliability of cued recall and word stem completion was investigated. There were significant priming effects and a dissociation between explicit and implicit memory as a function of levels of processing. However, the reliability of implicit memory measures was again substantially lower than in explicit tests in all test conditions. As a consequence, differential reliabilities of direct and indirect memory tests should be considered as a possible determinant of dissociations between explicit and implicit memory as a function of experimental or quasi-experimental manipulations. PMID- 10718072 TI - Is there cross-format transfer in implicit invariance learning? AB - Three experiments investigated cross-form transfer in the invariance learning paradigm introduced by McGeorge and Burton (1990). The results suggest that the transfer observed by McGeorge and Burton depended on subjects' ability to use a response strategy discovered by Wright and Burton (1995). When that strategy was denied to subjects (Experiments 1 and 2), no cross-form transfer was observed; when the strategy was made available (Experiment 3), cross-form transfer re emerged. These results suggest that this form of learning, like many other forms of implicit learning and memory, is hyperspecific. PMID- 10718073 TI - Set-size and frequency-of-occurrence judgements in young and older adults: the role of the availability heuristic. AB - Two experiments examined the cognitive processes underlying judgements of set size and judgements of frequency of occurrence in young (Experiments 1 and 2) and older (Experiment 2) adults. Previous research has implicated the availability heuristic in set-size judgements, whereas an automatic processing mechanism has been implicated in judgements of frequency of occurrence. In the current experiments, path analysis was employed to investigate the role of an availability bias in performance on the judgement tasks. In Experiments 1 and 2, both types of judgement were influenced by repetition frequency of words independent of the availability (recall) of specific exemplars. Experiment 2 extended the investigation to include age differences. Although older adults' recall performance was poorer overall, the availability bias was age invariant, and there were no age differences in either set-size or frequency-of-occurrence judgements. Our results indicate that both set-size and frequency-of-occurrence judgements are independent of the availability bias evident in recall, and they support the notion that an automatic processing mechanism underlies both types of judgement. PMID- 10718074 TI - Sleep loss and temporal memory. AB - Historical evidence suggests that sleep deprivation affects temporal memory, but this has not been studied systematically. We explored the effects of 36 hr of sleep deprivation on a neuropsychological test of temporal memory. To promote optimal performance, the test was short, novel, and interesting, and caffeine was used to reduce "sleepiness". A total of 40 young adults were randomized into four groups: control + caffeine (Cc), control + placebo (Cp), sleep deprived + caffeine (SDc), and sleep deprived + placebo (SDp). Controls slept normally. Caffeine (350 mg) or placebo were given just prior to testing. The task comprised colour photographs of unknown faces and had two components: recognition memory (distinction between previously presented and novel faces), and recency discrimination (temporal memory), when a previously shown face was presented. An interpolated task, self-ordered pointing, acted as a distraction. Caffeine had no effects within control conditions, but significantly reduced subjective sleepiness in SDc. Recognition was unaffected by sleep deprivation, whereas for recency, sleep deprivation groups scored significantly lower than controls. There was no significant improvement of recency with caffeine in the SDc group. Both sleep deprivation groups had poorer insight into their performance with recency. Self-ordered pointing remained unchanged. In conclusion, sleep deprivation impairs temporal memory (i.e. recency) despite other conditions promoting optimal performance. PMID- 10718075 TI - Taking humor seriously. PMID- 10718076 TI - Measuring and monitoring children's well-being across the world. AB - An international initiative to measure and monitor the status of children beyond survival is an effort to use tools of the information age to promote understanding of children's life perspectives and an action to improve their condition. An interdisciplinary group proposes widespread consensus on the selection and monitoring of cross-cultural indicators to cover the following children's life domains: social connectedness, civil life skills, personal life skills that enable children to contribute to their own well-being, safety and physical status, and children's subculture. Social workers can contribute substantially to the design, collection, interpretation, and use of indicators in various arenas ranging from local to global levels. PMID- 10718077 TI - Late entry into prenatal care: the neighborhood context. AB - This study investigated individual and neighborhood factors associated with late entry into prenatal care. Data from 220,694 New York City birth certificates were linked with data from the 1990 census and other secondary sources to determine the effect of women's individual characteristics and their neighborhood context on timing of prenatal care entry. Results indicate that 15 percent of New York City's pregnant women entered prenatal care late and that residence in a distressed urban neighborhood significantly increased the risk of late initiation, even in a model controlling for individual risk factors. Implications for social workers include the importance of outreach and case management for pregnant women, the value of health and social policies targeting distressed urban neighborhoods, and the need to ensure that Medicaid managed care implementation fosters use of prenatal care. The findings also underscore the importance of continuing to strive for a policy that ensures lifelong universal access to health care. PMID- 10718078 TI - Access barriers and the use of prenatal care by low-income, inner-city women. AB - An important public health agenda in the United States is improving access to prenatal care, particularly for low-income women. The study discussed in this article was designed to determine which social, environmental, and psychological barriers are most likely to interfere with the early and regular use of prenatal health services. Low-income adult women hospitalized on the postpartum unit of a large urban medical center were interviewed about the barriers they experienced gaining access to prenatal care. Access barriers involving family and friends significantly increased the odds of receiving inadequate care, particularly not wanting anyone to know about the pregnancy and not having help getting to clinic appointments. Other important barriers included those related to the health care system and intrapersonal issues. Social workers are in an ideal position to help women eliminate barriers to access to prenatal care through clinical expertise in assessment, advocacy, outreach, and case management. PMID- 10718079 TI - Family caregiving responsibilities among lesbians and gay men. AB - This study examines the full range of family care responsibilities among lesbians and gay men, including caring for children and adults with an illness or disability. Thirty-two percent of the gay men and lesbians in this study were providing some type of caregiving assistance. Lesbians, compared with gay men, were significantly more likely to be caring for children and elderly people, whereas gay men were more likely to be assisting working-age adults with an illness or disability. After controlling for the sociodemographic characteristics of the caregivers, having child care responsibilities was a significant predictor of not being openly identified as gay or lesbian, but child care and adult care responsibilities were not significant predictors of degree of support received from biological family members or of harassment experienced. These findings have implications for the development of human services practices and policies that are responsive to the unique needs of lesbians and gay men and their families. PMID- 10718080 TI - The effects of nonresident father involvement on single black mothers and their young children. AB - Using data from an ongoing study of current and former welfare recipients and their preschool children, the study discussed in this article examined the influence of the presence of nonresident fathers on the well-being and development of 188 low-income, employed and nonemployed single black mothers and their three- and four-year-old children. There is evidence that involvement by nonresident fathers has positive effects on maternal depression symptoms and child problem behaviors. The data show that maternal employment status seems to affect nonresident fathers' relations with single black mothers and their young children. It also seems to make a difference in the mothers' psychological well being. Policy implications are discussed. PMID- 10718081 TI - Adoption and race: implementing the Multiethnic Placement Act and the Interethnic Adoption Provisions. AB - Passage of the Multiethnic Placement Act of 1994 (MEPA) and the 1996 provisions on Removal of Barriers to Interethnic Adoption (Interethnic Adoption Provisions) provides an ideal opportunity to examine what can be done to increase adoptions of foster children, particularly children of color. To assist states and child welfare professionals in serving children better and meeting legal obligations, this article discusses racial matching, MEPA, and the Interethnic Adoption Provisions. Implications for practice and challenges in implementing the law also are discussed. The article suggests principles for adoption practice, recruitment, and placement that provide a framework for simultaneously meeting the requirements of the law and serving the best interests of children. Recommendations for practice, policy, and research are offered. PMID- 10718082 TI - "Friends in need": designing and implementing a psychoeducational group for school children from drug-involved families. AB - Despite widespread recognition of the risks that parental drug use pose to children, few resources are available to help such children. Using a developmental intervention approach, the authors designed and tested a model curriculum for use with groups of latency-aged children in schools located in communities where drug use is pervasive. In implementing this curriculum, the authors documented the need that children affected by family drug use have for workable strategies and skills for coping with aversive environments. The responsiveness of group participants to structure, predictability, and affirmation in the groups was remarkable. Measurable changes occurred in classroom behavior and feelings of self-worth. Obstacles to implementing and testing such an intervention are discussed. PMID- 10718083 TI - [Medical malpractice (I)]. PMID- 10718084 TI - [Society of siblings]. PMID- 10718085 TI - [Malignant melanoma of the skin. Pathogenesis, clinical aspects and prognosis]. AB - Cutaneous malignant melanoma is not among the most frequent cancer types at an incidence of 10-12/100,000 persons/year in Central Europe. However, the number of melanomas has doubled within the last 10-15 years. This increase is higher than that of most other malignancies. It has been attributed to an increased UV-light exposure, whereby the latency period probably reaches decades. Persons at high risk for melanoma were identified in various studies: those from families with other melanoma patients and those who have more than 50 benign nevi, atypical nevi and actinic freckles. The majority of melanomas develops de novo and not from preexisting nevi. Treatment of choice is complete surgical excision of the tumors. Prognosis of these patients is mainly determined by tumor thickness. Metastatic spread occurs most frequently within lymph vessels, local recurrences as well as regional metastases significantly impairing the prognosis. From long term experience in Australia it is evident that the dilemma "melanoma" can only be addressed effectively by rigorous prophylaxis and early diagnosis of the tumors. PMID- 10718086 TI - [New developments in surgery of the knee joint (II)]. AB - Due to the recent fitness craze and increased interest in leisure activities, there has also been a dramatic rise in injuries to the anterior cruciate ligament (ACL) of the knee. Thanks to the introduction of arthroscopic surgery techniques, the number of ACL operations is increasing, frequently with insufficient results caused by technical errors during the operation. New methods are therefore necessary in order to assure quality. Traumatic or degenerative lesions to the cartilage are relevant causes of loss of working hours and permanent disability. The development of new therapeutic methods is focused on interest in orthopaedic research. An important cause for lesion of the cartilage is a partial or total rupture of the meniscus as a result of traumatic or degenerative processes. Today refixation of a torn meniscus is unfortunately not performed enough. New fixation techniques have been developed to facilitate preservative meniscal surgery. Finally, accelerated and functional rehabilitation programs have significantly contributed to improve the outcome of knee surgery. PMID- 10718087 TI - [Epidemiology and socioeconomic consequences of migraine and headache diseases]. AB - The paper describes the situation of traditional headache therapy on the basis of up-to-date empirical data on headache epidemiology, the habits of the persons affected, and the economic consequences for employers, health insurance funds and society in general. It also describes for the first time the direct costs of in patient headache therapy in a traditional treatment context without specially organized therapy on the basis of actual invoicing between hospitals and health insurance funds. PMID- 10718088 TI - [Does combination therapy of chronic viral hepatitis C with interferon-alpha and ribavirin approach financial limits?]. AB - Until April 1999, interferon alpha was the only licensed medication in Germany for chronic hepatitis C virus infection with proven effectiveness. In two large placebo-controlled studies the combination therapy with interferon alpha and ribavirin confirmed a major improvement of the sustained response compared to the treatment with interferon alone. These results led to the approval in Germany of the combination therapy for patients with chronic hepatitis C without prior interferon therapy and for patients who relapse after an initial response to therapy with interferon alone. The expenses for the two drugs of the combination therapy are, however, very high (about 40,000 DM for one year of treatment). Considering the large number of patients with chronic hepatitis C in Germany (about half a million), it is suggested that more detailed recommendations for the indication of the combination therapy should be elaborated by a group of experts. Using data from the published literature it is shown for two subgroups of patients with chronic hepatitis C that a more differentiated indication for the use of the combination therapy is possible and justified and might help to avoid a more dramatic increase of drug expenses without shortcomings for the patients. PMID- 10718089 TI - [Bicycle accidents with and without bicycle helmets]. AB - Traffic accidents with bicycles are very frequent, since bicycling has become increasingly popular as a means of transportation and a sports instrument. Therefore, there are many bicycling victims in casualty hospitals. The head being exposed and unprotected, head and brain injuries are the most frequent ones. Just as in professional and sports activities, protection of the head and neck also when bicycling has been urgently recommended and ist effect repeatedly confirmed by recent statistical results. Wearing helmets by motorcyclists has clearly reduced the severity and frequency of head injuries since it became compulsory. A legislative initiative for wearing bicycle helmets as well ist therefore necessary. PMID- 10718090 TI - [Subarachnoid hemorrhage as life-threatening complication of neural therapy. Case report]. AB - In Germany, neural therapy is a frequently used method of complementary medicine. It is based on the hypothesis, that pathologically altered regions may cause symptoms in other, distant localizations and that this process can be interrupted by the local injection of anesthetic drugs. Since the pathogenetic acting regions are supposed to occur in any part of the body, neural therapy is not limited to superficial injections, but also includes injections into deep structures and internal organs. Supporters of this therapeutic approach always claim that these procedures are free of risk if carried out by an experienced therapist. We report the case of a young woman, who suffered subarachnoidal bleeding caused by an attempt to infiltrate the tonsilla pharyngea during a neural therapeutic treatment. The analysis of this incident shows that severe and even life threatening complications cannot be ruled out in all neural therapeutic injections, even if they are performed strictly according to the rules given in the textbooks of neural therapy. PMID- 10718091 TI - [Comment on X. Baur, C. Sieger: Evaluation of residual working capacity with lung function studies]. PMID- 10718092 TI - [Comment on S. Gabius, H.-J. Gabius: Immune modulating mistletoe therapy by lectin standardization: a double-edged sword?]. PMID- 10718093 TI - Age-specific excess deaths associated with stroke among racial/ethnic minority populations--United States, 1997. AB - Stroke was the third leading cause of death in the United States in 1997. During 1950-1996, age-standardized stroke death rates declined 70% for the entire U.S. population; however, the decline varied among racial/ethnic populations. The estimated number of stroke deaths by race/ethnicity and age illustrate the differences in stroke mortality that may be used to direct prevention efforts. This report presents an analysis of stroke mortality by age and racial/ethnic group; the findings indicate that for persons aged 35-64 years, excess stroke deaths and higher risk for stroke mortality occurred among members of U.S. racial/ethnic minority populations than among the non-Hispanic white population. PMID- 10718094 TI - Elevated blood lead levels among internationally adopted children--United States, 1998. AB - Lead poisoning has been reported recently among Chinese children adopted by U.S. citizens. However, little is known about the prevalence of elevated blood lead levels (BLLs) among adoptees from China and other countries. Persistent sources of lead exposure outside the United States include leaded gasoline exhaust; industrial emissions; cottage industries (e.g., battery breaking and recycling plants); traditional medicines; and some cosmetics, ceramic ware, and foods. In 1998, approximately 15,000 orphans from countries outside the United States who were adopted abroad or were to be adopted in the United States by U.S. citizens were issued U.S. immigrant visas-a nearly two-fold increase over 1988 (L. Lewis, Immigrant and Visa Control and Reporting Division, VISA Office, Bureau of Consular Affairs, U.S. State Department, personal communication, August 1999). Some orphans have been abandoned for extended periods and have no obtainable medical history. Immigrants aged <15 years are not required to have serologic or blood tests either in their country of origin or on entry into the United States unless exposure to syphilis or human immunodeficiency virus is suspected. To obtain reports on the prevalence of elevated BLLs (> or =10 microg/dL) among international adoptees, CDC contacted 12 international adoption medical specialists identified through the Joint Council on International Children's Services and two collaborating medical specialists. This report summarizes the results of that investigation, which suggest that international adoptees may arrive in the United States with elevated BLLs. PMID- 10718095 TI - Vaccination coverage among adolescents 1 year before the institution of a seventh grade school entry vaccination requirement--San Diego, California, 1998. AB - In 1996, the Advisory Committee on Immunization Practices, the American Academy of Pediatrics, the American Association of Family Physicians, and the American Medical Association recommended routine health-care visits for children aged 11 12 years, emphasizing vaccination with hepatitis B vaccine; measles, mumps, and rubella vaccine (MMR); tetanus and diphtheria toxoids (Td); and varicella vaccine. Because no national data exist regarding vaccination coverage among adolescents, the impact of these recommendations is unknown. In October 1997, California enacted Assembly Bill 381 (AB381) that requires students entering the seventh grade on or after July 1, 1999, to have received three doses of hepatitis B vaccine and two doses of MMR. To assist in planning and implementing AB381, the San Diego County Health Department expanded its 1998 infant and adult vaccination survey to include fifth and sixth graders. This report summarizes the findings from that survey, which indicate that most fifth and sixth graders lacked required and recommended vaccinations. PMID- 10718096 TI - HMG-CoA reductase inhibitors in the prevention of stroke. AB - Stroke is a heterogeneous disorder, with the definition including both haemorrhagic and ischaemic stroke. Although these subtypes of stroke have different underlying pathophysiological mechanisms, atherosclerosis plays a pivotal role in both. Most risk factors for cardiovascular disease are also risk factors for stroke. Patients with a history of cardiovascular events are at an increased risk of stroke. Although hypercholesterolaemia is the most characteristic risk factor for atherosclerotic diseases, recent data suggest that the correlation between cholesterol levels and either ischaemic or haemorrhagic stroke is weak. However, the interpretation of these results is hampered by the inconsistent use of classifications of the various subtypes of stroke in studies. Pooled data on the effect of HMG-CoA reductase inhibitors show a 30% risk reduction in strokes. These beneficial effects are obtained from studies in middle aged patients with ischaemic heart disease, the interpretation being that the effects of HMG-CoA reductase inhibitors on stroke are mediated via (i) cholesterol-lowering effects on the coronary vasculature or (ii) cholesterol independent effects of these agents. The results cannot be extrapolated to the elderly, among whom stroke most frequently occurs. PMID- 10718097 TI - Oxazolidinones: a review. AB - The oxazolidinones represent a novel chemical class of synthetic antimicrobial agents. They exhibit an unique mechanism of protein synthesis inhibition and generally display bacteriostatic activity against many important human pathogens, including methicillin-resistant Staphylococcus aureus, vancomycin-resistant enterococci, and penicillin- and cephalosporin-resistant Streptococcus pneumoniae. Linezolid, the oxazolidinone which has been selected for clinical development, has near complete oral bioavailability plus favourable pharmacokinetic and toxicity profiles. Results from experimental models of infection and phase II trials reveal linezolid to be highly active in vivo against infections due to many common gram-positive pathogens. The role of linezolid remains to be determined in phase III clinical trials, but it shows great promise as an alternative to glycopeptides and streptogramins to treat serious infections due to resistant gram-positive organisms. Further modification of the oxazolidinone nucleus may yield agents with even greater potency and with novel spectra of activity. PMID- 10718098 TI - Advances in non-nicotine pharmacotherapy for smoking cessation. AB - Progress in understanding the pharmacological nature of tobacco addiction, along with the modest success rates achieved by the nicotine replacement therapies, has provided the major impetus for the development of non-nicotine drugs as smoking cessation aids. This article reviews evidence from controlled trials of several non-nicotine medications for the treatment of nicotine dependence. Clonidine was the first non-nicotine medication to show efficacy for smoking cessation in multiple studies, but its effect was found to be limited at best. Positive results across several trials have been consistently demonstrated for amfebutamone (bupropion). Encouraging results have also been observed for nortriptyline and moclobemide. Studies of combined treatments using non-nicotine medications (amfebutamone, mecamylamine, oral dextrose) with nicotine replacement therapy suggest increased efficacy relative to treatments using one or the other treatment strategy alone. Thus, available evidence indicates that non-nicotine drug treatments offer a promising panoply of therapeutic strategies for the addicted smoker. PMID- 10718100 TI - An evaluation of pharmacological strategies for the prevention and treatment of acute renal failure. AB - Acute renal failure (ARF) occurs frequently in hospitalised patients, and is associated with significant morbidity and mortality. The most common and generalised forms of acute renal failure are pre-renal conditions and intra-renal acute tubular necrosis (ATN). Pre-renal ARF in its pure state should be entirely reversible by restoring renal perfusion, but in some cases ATN has already occurred. ATN remains a more vexing problem, and is seen most often with hypotension, perioperative or systemic inflammatory stresses, radiocontrast administration, and exposure to nephrotoxins. Among the available pharmacological options for prevention or treatment of ATN, there is a remarkable lack of definitive evidence supporting specific therapy in any setting. Although loop diuretics, mannitol, and dopamine are frequently used for prevention and/or treatment of ATN, clinical studies have failed to prove value. Other drugs with theoretical value, specifically atrial natriuretic peptide analogues, adenosine blockers, and calcium antagonists, have been insufficiently studied to recommend use. Other pharmacological options may arise in the future. Ensuring adequate intravascular fluid volume remains the only approach to managing ATN which can be considered relatively effective and safe. Given the abundant theoretical basis for the prevention and treatment of ATN with drugs, well conducted clinical studies with relevant outcome measures are clearly warranted. PMID- 10718101 TI - Pituitary disorders. Drug treatment options. AB - Pituitary diseases are relatively common entities in the general population. They include pituitary adenomas and hypopituitarism. Pituitary tumours can cause symptoms of mass effect and hormonal hypersecretion that can be reversed with surgical resection or debulking of the adenoma, radiotherapy, or medical treatment. Transsphenoidal adenomectomy is the treatment of choice for acromegaly, Cushing's disease, gonadotropin-secreting tumours; and thyrotropin (TSH)-secreting adenomas. Pituitary irradiation and medical therapy are secondary options. Conversely, medical treatment is the primary choice for prolactinomas. Dopamine agonists are very effective in the treatment of prolactin (PRL) secreting tumours, with rates of control as high as 80 to 90% for microprolactinomas (< 10 mm) and 60 to 75% for macroprolactinomas (> or = 10 mm). Somatostatin analogues have also shown efficacy in patients with acromegaly who have not responded to surgery or in patients with TSH-secreting adenomas who have not improved with surgery and radiotherapy. In patients with Cushing's disease, who are not cured surgically or who relapse after pituitary adenomectomy and irradiation, steroidogenic inhibitors can be an efficient method of controlling the hypercortisolism. Pituitary insufficiency is the partial or complete loss of the anterior hypophyseal function, which is due to hypothalamic or pituitary disease. Although the classic sequence of loss of pituitary secretion is growth hormone (GH), gonadotropins, TSH, and corticotropin (ACTH), the order to begin the replacement therapy of the deficient hormone(s) is cortisol, thyroxine, androgens/estrogens and, if necessary, GH. There are multiple preparations that can be used to achieve clinical and biochemical improvement. In general, the hormone replacement therapy is lifelong. PMID- 10718102 TI - Ganirelix. AB - Ganirelix is a synthetic third generation gonadotropin-releasing hormone (GnRH) antagonist that is administered via the subcutaneous route. The drug competitively blocks GnRH receptors in the anterior pituitary gland, preventing endogenous GnRH from inducing luteinising hormone (LH) and follicle stimulating hormone release. Ganirelix effectively inhibited LH surges during controlled ovarian stimulation in a large, multicentre clinical trial in women undergoing in vitro fertilisation. A vital pregnancy rate per embryo transfer of 40.3% was achieved at weeks 5 to 6 after treatment with the 0.25 mg/day dosage. Subcutaneous ganirelix has been generally well tolerated in clinical trials. The most common adverse events were local injection site events, asthenia, nausea, malaise, headache and fatigue. PMID- 10718103 TI - Moxifloxacin: a review of its clinical potential in the management of community acquired respiratory tract infections. AB - Moxifloxacin is an extended-spectrum fluoroquinolone which has improved coverage against gram-positive cocci and atypical pathogens compared with older fluoroquinolone agents, while retaining good activity against gram-negative bacteria. The antibacterial spectrum of moxifloxacin includes all major upper and lower respiratory tract pathogens; it is one of the most active fluoroquinolones against pneumococci, including penicillin- and macrolide-resistant strains. In in vitro studies, emergence of bacterial resistance was less common with moxifloxacin than with some other fluoroquinolones, but this requires confirmation in large-scale clinical studies. As with other fluoroquinolones, moxifloxacin achieves good penetration into respiratory tissues and fluids. It shows a low potential for drug interactions and dosage adjustment is not required for patients of advanced age or those with renal or mild hepatic impairment. The efficacy of oral moxifloxacin has been demonstrated in large, well-designed clinical trials in patients with community-acquired pneumonia, acute exacerbations of chronic bronchitis or acute sinusitis. Moxifloxacin 400 mg once daily achieved bacteriological and clinical success rates of approximately 90% or higher. It was as effective as, or more effective than, comparators including clarithromycin, cefuroxime axetil and high dose amoxicillin in these trials. The most commonly reported adverse events in patients receiving moxifloxacin are gastrointestinal disturbances. Moxifloxacin is also associated with QTc prolongation in some patients; there are, as yet, no data concerning the possible clinical sequelae of this effect in high-risk patients. Moxifloxacin has a low propensity for causing phototoxic reactions relative to other fluoroquinolones, and animal data suggest that it has a low potential for causing excitatory CNS and hepatotoxic effects. CONCLUSIONS: As an extended-spectrum fluoroquinolone, moxifloxacin offers the benefits of excellent activity against pneumococci, once daily administration and a low propensity for drug interactions. Although studies are needed regarding its tolerability in at-risk patients with QT interval prolongation, available data suggest that moxifloxacin is likely to become a first-line therapy option for the treatment of community-acquired lower respiratory tract infections, particularly in areas where drug-resistant S. pneumoniae or H. influenzae are common. PMID- 10718105 TI - Is administration of n-3 fatty acids by mucosal enema protective against trinitrobenzene-induced colitis in rats? AB - We investigated the protective role of fish oil (FO-source of n-3 FA) enriched diet (in the first protocol) in 20 rats and FO administration intrarectally (in the second protocol) in 40 rats with trinitrobenzene (TNB) colitis. All colonic specimens were pathologically evaluated, myeloperoxidase enzyme activities were measured, leukotriene B4 (LTB4) and LTC4 levels were determined by radioimmunoassay. In the first protocol 10 rats (group A1) were fed with 8% sunflower and cotton oil enriched diet and (group A2) with 8% FO enriched diet for 6 weeks. At the end of this period, TNB (30 mg in 0.25 ml of 30% ethanol) were intrarectally administered. After 2 weeks, rats were sacrificed. MPO activities (2.47 versus 30.17), LTB4 (34.5 versus 903.3) and LTC4 (77.7 versus 456.0) levels were significantly reduced in group A2 compared with group A1 (P<0.005). There was also a significant difference in pathologic scores (1.55 versus 2.12, P<0.002) between two groups. In the first part of the second protocol, 20 male rats were randomized into two equal groups (B1 and B2) and TNB colitis was induced. After 1 day, 1 ml of saline (group B1) or n-3 FA enemas (group B2) were administered every day for 2 weeks. At the end of this period, rats were sacrificed and evaluated as done for previous groups. Although there was no significant difference between the two groups in comparison with MPO enzyme activities and pathologic scores, the LTB4 (130.1 versus 971.0) and LTC4 (126.0 versus 532.0) levels of FO group were significantly reduced (P<0.005). In the second part of the second protocol, 20 male rats were randomized into two groups. One millilitre of saline (group B3) or FO enemas (group B4) were administered to rats every day for 3 days. At the fourth day, TNB-colitis was induced and after 24 h rats were sacrificed. We could not find any significant difference in MPO activities, pathologic scores, LTB4 and LTC4 levels between groups B3 and B4. In conclusion, FO enriched diet decreased both pathologic damage and tissue LT levels. The second protocol of our study revealed that the long-term FO enemas decreased the LTB4 and LTC4 levels; however, did not have any beneficial effect on the tissue lesions. Short periods of FO enemas did not have a protective role in the occurrence of experimental colitis. The present study showed that FO enemas significantly decreased LT levels. The protective effect of FO (oral and enema) in TNB colitis may open a new insight into the treatment of inflammatory bowel disease. PMID- 10718104 TI - Felodipine/metoprolol: a review of the fixed dose controlled release formulation in the management of essential hypertension. AB - The main objective of fixed dose combination therapy for hypertension is to improve blood pressure (BP) control with lower, better tolerated dosages of 2 antihypertensives rather than higher dosages of a single agent. Felodipine and metoprolol lower BP via different, but complementary, mechanisms and controlled release formulations of these 2 drugs are available as a fixed dose combination, felodipine/metoprolol. In clinical trials in patients with hypertension, felodipine/metoprolol was significantly more effective than placebo and the respective monotherapies administered at the same dosages. Mean BP was reduced to < 155/90 mm Hg in patients treated with combination therapy and controlled in approximately 70% of patients. In one study that titrated dosages to effect, fewer felodipine/metoprolol than felodipine or metoprolol monotherapy recipients required dosage increases to achieve BP control (45 vs 60 and 67%, respectively). Data from double blind comparative studies show that the antihypertensive efficacy of felodipine/metoprolol 5 to 10/50 to 100 mg/day is significantly greater than that of enalapril monotherapy or captopril plus hydrochlorothiazide and equivalent to nifedipine/atenolol and amlodipine. In comparisons with enalapril, fewer felodipine/metoprolol than enalapril recipients required dosage titration to achieve BP control. Compared with amlodipine, felodipine/metoprolol significantly reduced mean 24-hour average BP (8.9/5.5 vs 14.4/9.5 mm Hg after 6 weeks; p < 0.001). Both treatments preserved diurnal rhythm. Long term follow-up studies show that the antihypertensive effect of felodipine/metoprolol occurs mostly during the first month of treatment with small additional decreases in BP being observed in the second and third months, and a relatively constant effect thereafter. According to a validated questionnaire, quality of life was relatively similar during 12 weeks treatment with felodipine/metoprolol, enalapril or placebo. In a retrospective pharmacoeconomic analysis conducted in Sweden, felodipine/metoprolol was more cost effective than enalapril as initial treatment for hypertension. Peripheral oedema, headache and flushing were the most commonly reported adverse events with felodipine/metoprolol and felodipine monotherapy, whereas dizziness, fatigue, headache and respiratory infection were more frequent with metoprolol monotherapy. Dose-dependent adverse events such as oedema may occur less often in patients taking lower dosages in combination than in those taking higher dosages of felodipine monotherapy. Thus, patients with hypertension treated with felodipine/metoprolol experience greater control of BP, with less need for dosage titration, than those treated with felodipine, metoprolol or enalapril monotherapy. Importantly this greater efficacy does not appear to be associated with a higher incidence of adverse events relative to monotherapy. Additionally, in short term studies felodipine/metoprolol had a similar (minimal) effect on QOL to enalapril monotherapy but was more cost effective. PMID- 10718106 TI - Decreased production of interleukin-6 and prostaglandin E2 associated with inhibition of delta-5 desaturation of omega6 fatty acids in mice fed safflower oil diets supplemented with sesamol. AB - The differences in the immune responses in mice fed sesame oil diets and those fed sesamin may be attributed to the presence of other lignans in the non-fat portion of the oil. The fatty acid composition (mean +/- SD mol. %) of liver membrane phospholipids and the levels of endotoxin-induced prostaglandin (PG) E2, interleukin (IL)-6, IL-10, IL-12 and tumor necrosis factor (TNF)-alpha were determined in mice fed diets supplemented with 5% safflower oil (SO) in the absence or presence of 1% sesamol. The levels of dihomo-gamma-linolenic acid (20:3omega6) were markedly higher (P<0.025) in the livers from mice fed sesamol supplemented SO diets (1.6 +/- 0.1) compared to the controls (1.4 +/- 0.1). These data suggest that sesamol or its metabolite could inhibit the in vivo delta-5 desaturation of omega6 fatty acids. Further, in animals fed sesamol supplemented SO diets, the levels of PGE2 (228 +/- 41 pg/ml) were markedly lower (P<0.01) compared to those fed SO diet alone (1355 +/- 188 pg/ml). Concomitantly, the concentrations of IL-6 were also lower (P<0.01) in mice fed sesamol diet (63 +/- 11 ng/ml) compared to the controls (143 +/- 22 ng/ml). A marked reduction in the levels of PGE2 in animals fed sesamol diets suggests that sesamol or its metabolite could inhibit the activity of cyclooxygenase enzyme. PMID- 10718107 TI - The effect of epidermal growth factor on prostaglandin synthesis of oestrogenized rat uterus is mediated by nitric oxide. AB - We examined the possible relationship between cytokines, nitric oxide and prostaglandins (PGs) in the estrogenized rat uterus. Results indicate that epidermal growth factor (EGF) enhances the synthesis of prostaglandins in estrogenized rat uteri and induces the augmentation of nitric oxide (NO) production in this tissue by stimulating iNOS. While the effect of EGF is abolished by L-NMMA, an NO antagonist, the NS-398, a cyclooxygenase-II (COX-II) inhibitor, prevents the augmentation of prostanoids induced by EGF. These results suggest that there is an interaction among EGF, NO and PGs and that in this interrelationship are involved COX-II and iNOS. This mechanism might be important during implantation and labor. PMID- 10718108 TI - The enzymes responsible for the metabolism of prostaglandins in bovine placenta. AB - The following review presents some data on the enzymes of synthesis and catabolism of prostaglandins in bovine placenta. The available literature confirms the anabolic and catabolic ability of bovine placenta to metabolise prostaglandins, but there is little direct information about the pathways involved. The aim of the review is to describe what is known about the physico chemical properties and activity of enzymes involved in prostaglandin metabolism in bovine placenta. PMID- 10718109 TI - Endogenous prostaglandin formation in the field-stimulated guinea-pig vas deferens: comparison of the inhibitory effects of indomethacin and NS-398. AB - Prostaglandin (PG) E2 inhibited both phases of contraction produced by electrical field stimulation of the guinea-pig vas deferens. PGF2alpha and PGD2 were without effect on this preparation. Carbacyclin (a PGI2) analogue inhibited the first phase of contraction at higher concentrations, whereas U46619 (a thromboxane mimetic) potentiated both phases of contraction. As exogenous arachidonic acid inhibits both phases of contraction of the electrically field-stimulated guinea pig vas deferens, it is likely that the arachidonic acid is converted to PGE2 in the vas deferens. Indomethacin, a non-specific inhibitor of prostaglandin H synthase (PGHS), attenuated the inhibitory actions of exogenous arachidonic acid when examined on the first phase of contraction. NS-398, a relatively specific inhibitor of PGHS-2, also prevented the inhibitory action of exogenous arachidonic acid. However, NS-398 was much less effective than indomethacin in this respect even though NS-398 and indomethacin inhibit PGHS-2 with similar potencies. Consequently, the findings suggest that exogenous arachidonic acid is converted to PGE2 in the guinea-pig vas deferens by the actions of PGHS-1 and, to a lesser extent, by PGHS-2. PMID- 10718110 TI - Vascular effects of isoprostanes after endothelial damage. AB - The isoprostanes, 8-iso-PGF2alpha and 8-iso-PGE2, are powerful vasoconstrictors in vitro and in vivo. Increased formation of 8-iso-PGF2alpha was detected in atherosclerotic patients. In this disease endothelial injuries appear, followed by a reduced release of nitric oxide (NO). Our results show that the vasoconstrictor effects of 8-iso-PGF2alpha as well as of 8-iso-PGE2 were amplified after endothelial damage of the vasculature of the isolated perfused rabbit ear. Also vasoconstrictions induced by both isoprostanes were amplified by perfusions with the NO-synthase blocker N(G)-nitro-L-arginine methylester (L NAME) at a final concentration of 100 micromol/l. Therefore, we can assume that the amplification of the vasoconstrictor effects of the isoprostanes used after damage of endothelium might be mainly due to the reduced formation of NO. PMID- 10718111 TI - Characterization of the amnion-derived AV3 cell line for use as a model for investigation of prostaglandin-cytokine interactions in human amnion. AB - Increased production of prostaglandins and cytokines by amnion, particularly prostaglandin (PG) E2, interleukin (IL)-6 and IL-8, is thought to be an important event in infection-associated preterm labour. We characterized the amnion-derived AV3 cell line to determine its appropriateness as a model for investigation of the regulation of amnion cytokine and PG production. Amnion-derived AV3 cells were treated with tumour necrosis factor-alpha (TNF-alpha, interleukin-1beta (IL 1beta), epidermal growth factor (EGF) and phorbol 12-myristate 13-acetate (PMA) and IL-6, IL-8 and prostaglandin production was determined by immunoassay. Production of IL-6 and IL-8 rose dramatically with all treatments. PGE2, but not PGF2alpha or 6-keto-PGF1alpha, biosynthesis was also increased in a concentration dependent manner with all treatments. A rapid increase in PGHS-2 (but not PGHS-1) mRNA expression was observed in response to TNF-alpha and IL-1beta. We conclude that the AV3 cell line inflammatory response profile is similar to those observed in primary amnion and other amnion-derived cell lines, and is an appropriate model for human amnion. PMID- 10718112 TI - Oestrogen and essential fatty acid supplementation corrects bone loss due to ovariectomy in the female Sprague Dawley rat. AB - Essential fatty acid deficient animals develop osteoporosis. Eicosapentaenoic acid and gamma-linoleic acid have been reported to have positive effects on bone metabolism in both the growing male rat and the ovariectomized (OVX) female rat. These effects have been further investigated using a novel gamma linolenic/eicosapentaenoic acid diester together with an oestrogen implant in the ovariectomized, female Sprague Dawley rat. Rats were sham-operated or ovariectomized at age 11 weeks. Two groups of OVX rats received an oestrogen implant at ovariectomy. Animals received fatty acids, linoleic acid (control) or a diester with gamma-linolenic acid and eicosapentaenoic acid as part of a semi synthetic diet. Bone calcium content and excretion of deoxypyridinolines as marker of bone degradation were measured at 14 weeks. Oestrogen, as well as diester alone, increased calcium/femur to sham levels. Oestrogen plus diester potentiated the effect of oestrogen on bone calcium (P < 0.05 vs OVX). At the same time, oestrogen alone and the combination of oestrogen plus diester significantly reduced (P < 0.05 vs OVX) urinary deoxypyridinoline and hydroxyproline excretion. Again, the diester potentiated the effect of oestrogen. The effects of the diester alone, together with the potentiated effects of oestrogen by the essential fatty acids on osteoporosis, are novel findings. PMID- 10718113 TI - Inflammatory mediators induce sequestration of VE-cadherin in cultured human endothelial cells. AB - The mechanisms through which inflammatory mediators modify endothelial junctional structure are not well understood. Endothelial cells exposed to 1 mM H2O2, 0.1 mM histamine or 4 mM EDTA displayed decreased amounts of VE-cadherin on the cell surface in a time-dependent manner. H2O2 and EDTA-treated cells showed a sustained reduction in surface VE-cadherin, but histamine (0.1 mM) decreased cell surface VE-cadherin only at 5 and 15 min, not at 30 and 60 min. Sequestering of VE-cadherin could also be visualized as a decrease in immunofluorescent labeling of endothelial junctions in fixed, non-extracted monolayers. However, junctional staining was observed in these cells after membrane extraction. This decreased surface expression of VE-cadherin was actin-filament, but not PKC/MAP kinase dependent. VE-cadherin binding to the cytoskeleton was decreased by EDTA, but was not diminished by histamine or H2O2. Therefore, by promoting sequestration of junctional cadherins, inflammatory mediators may decrease adhesive bonds between apposed endothelial cells and increase solute permeability. PMID- 10718099 TI - Pharmacology of AMPA/kainate receptor ligands and their therapeutic potential in neurological and psychiatric disorders. AB - It has been postulated, consistent with the ubiquitous presence of glutamatergic neurons in the brain, that defects in glutamatergic neurotransmission are associated with many human neurological and psychiatric disorders. This review evaluates the possible application of ligands acting on glutamate alpha-amino-3 hydroxy-5-methyl-4-isoxazole propionate (AMPA) and kainate (KA) receptors to minimise the pathology and/or symptoms of various diseases. Glutamate activation of AMPA receptors is thought to mediate most fast synaptic neurotransmission in the brain, while transmission via KA receptors contributes only a minor component. Variants of the protein subunits forming these receptors greatly extend the pharmacological and electrophysiological properties of AMPA/KA receptors. Disease and drug use can differentially affect the expression of the subunits and their variants. Ligands bind to AMPA receptors by competing with glutamate at the glutamate binding site, or non-competitively at other sites on the proteins (allosteric modulators). Ligands showing selective competitive antagonist actions at the AMPA/ KA class of glutamate receptors were first reported in 1988, and the systemically active antagonist 2,3-dihydroxy-6-nitro-7 sulphamoyl-benzo(F)quinoxaline (NBQX) was first shown to have useful therapeutic effects on animal models of neurological diseases in 1990. Since then, newer antagonists with increased potency, higher specificity, increased water solubility, and a longer duration of action in vivo have been developed. Negative allosteric modulators such as the prototype GYKI-52466 also block AMPA receptors but have little action at KA receptors. Positive allosteric modulators enhance glutamatergic neurotransmission at AMPA receptors. Polyamines and adamantane derivatives bind within the ion channel of calcium-permeable AMPA receptors. The latest developments include ligands selective for KA receptors containing Glu-R5 subunits. Evidence for advantages of AMPA receptor antagonists over N-methyl-D aspartate (NMDA) receptor antagonists for symptomatic treatment of neurological and psychiatric conditions, and for minimising neuronal loss occurring after acute neurological diseases, such as physical trauma, ischaemia or status epilepticus, have been shown in animal models. However, as yet AMPA receptor antagonists have not been shown to be effective in clinical trials. On the other hand, a limited number of clinical trials have been reported for AMPA receptor ligands that enhance glutamatergic neurotransmission by extending the ion channel opening time (positive allosteric modulators). These acute studies demonstrate enhanced memory capability in both young and aged humans, without any apparent serious adverse effects. The use of these allosteric modulators as antipsychotic drugs is also possible. However, the long term use of both direct agonists and positive allosteric modulators must be approached with considerable caution because of potential adverse effects. PMID- 10718114 TI - Regulation of ICAM-1 expression in mouse macrophages. AB - In a mouse model of silica (SI) induced lung injury, SI exposure increases expression of intercellular adhesion molecule-1 (ICAM-1) on lung (alveolar/interstitial) macrophages and alveolar type II epithelial cells. To investigate the regulation of SI induced ICAM-1 expression on mouse macrophages, freshly isolated macrophages (alveolar, peritoneal) and macrophage cell lines (MH S, RAW 264.7) were evaluated for ICAM-1 expression elicited by the particle silica (alpha quartz; 20 microg/ml; 6 microg/cm2) or the inflammatory cytokine, TNFalpha (20 ng/ml). TNFalpha significantly increased ICAM-1 expression in all cell types whereas SI elicited an increase in peritoneal macrophages (PM) and the cell line, MH-S. This pattern of increased expression was confirmed by immunocytochemistry. To investigate the regulation of ICAM-1 expression, PM were incubated with SI, TNFalpha or media concomitantly with anti-TNFalpha antibody, the antioxidant, NAC, or the iNOS synthase inhibitor, L-NAME. Both anti-TNFalpha and NAC, but not L-NAME, inhibited elicited (TNFalpha, SI) as well as constitutive (media) ICAM-1 expression. These data demonstrate that both inflammatory cytokines and inorganic particles can increase ICAM-1 expression on mouse macrophages and that this expression is mediated, in part, by TNFalpha and reactive oxygen species. PMID- 10718115 TI - Expression and regulation of macrophage inflammatory protein-2 gene by vanadium in mouse macrophages. AB - Environmental and occupational exposure to vanadium (V) dusts results in inflammation mainly confined to the respiratory tract. Macrophages apparently play an important role in mediating the inflammation via the production of many chemokines. In the current study, we investigated whether vanadium can regulate the gene expression of a CXC chemokine macrophage inflammatory protein-2 (MIP-2), and to determine the molecular mechanisms controlling MIP-2 gene expression. A mouse macrophage cell line RAW 264.7 was treated with sodium metavanadate (NaVO3) at the dose of 0.5, 5, or 10 microg/mi V. Northern blot analysis showed that induction of MIP-2 mRNA expression was in a dose-dependent manner. To define the time course of the inflammatory response, RAW 264.7 cells were exposed to 5 microg/ml V, MIP-2 mRNA in macrophages increased markedly as early as 1 h after treatment, maximally induced at 4 h and reduced to 2-fold above control levels by 6 and 8 h. The protein levels of MIP-2 in conditioned media, measured by enzyme linked immunosorbent assay (ELISA), was well correlated with the levels of MIP-2 mRNA following all of the treatments in the study. In addition, the increase in MIP-2 mRNA expression by vanadium was attenuated by co-treatment with the antioxidant N-acetylcysteine (NAC), at the doses of 10 and 20 mM, suggesting that the induction of MIP-2 mRNA is mediated via the generation of reactive oxygen species (ROS). To further investigate transcriptional regulation of the MIP-2 gene expression by vanadium, we performed RNA decay assay by measuring the half life of MIP-2 mRNA. Co-treatment of macrophages with the transcriptional inhibitor actinomycin D at 5 microg/ml following exposure to 5 microg/ml V for 4 h revealed complete stabilization of vanadium-induced MIP-2 mRNA and no sign of mRNA degradation, at least, for 6 h, in comparison to the half-life of MIP-2 mRNA was approximately 2.5 h by bacterial lipopolysaccharide (LPS) treatment, supporting post-transcriptional stabilization as the predominant role of MIP-2 gene expression. In conclusion, these observations demonstrate that in vitro vanadium can induce MIP-2 mRNA expression, mediating, at least in part, via the production of ROS. In addition, the increase in MIP-2 mRNA level involves, most likely, post-transcriptional control via increased mRNA stability. PMID- 10718116 TI - Carrageenan-induced arthritis in the rat. AB - This study documents a model of carrageenan-induced chronic inflammatory arthritis in the rat, using quantitative histomorphometric assessment. Ten Sprague-Dawley female rats were randomly assigned to one of two groups. Arthritis was induced in the right tibiofemoral joint by 7 intra-articular injections of 0.02 mL of 1% carrageenan in the arthritic group over 24 days. The control (normal) group was injected with 0.02 mL of saline in the right tibiofemoral joint. Sagittal sections of the right knee joint (distal femur and proximal tibia) were assessed by histomorphometry using the LECO 2001 image analysis system. Articular cartilage thickness, epiphyseal plate thickness, subchondral bone plate thickness, trabecular bone volume and thickness of the synovial lining cell layer were measured. Differences between normal and arthritic groups were statistically significant for articular cartilage thickness of the femur, epiphyseal plate thickness of both the femur and tibia, subchondral bone plate thickness of the tibia and the thickness of the synovial lining cell layer. These findings demonstrate that carrageenan-induced arthritic changes are similar to other, established models of arthritis in the rat. PMID- 10718119 TI - Fenamates and the potent inhibition of human liver phenol sulphotransferase. AB - 1. The inhibition of the human liver phenol sulphotransferase (HL-PST) and catechol sulphotransferase (HL-CST) by five fenamates has been studied and the activities of HL-PST and HL-CST were measured with 4-nitrophenol and dopamine as substrates, respectively. 2. The IC50 for inhibition of HL-PST were 0.02 microM (mefenamic acid); 0.12 microM (tolfenamic acid); 0.28 microM (niflumic acid); 0.87 microM (meclofenamic acid) and 1.50 microM (flufenamic acid). 3. HL-CST was less susceptible than HL-PST to the inhibition by fenamates and the IC50 for HL CST were 36 microM (tolfenamic acid); 70 microM (flufenamic acid); 76 microM (mefenamic acid); 180 microM (niflumic acid) and 185 microM (meclofenamic acid). 4. The ratios of the IC50 for HL-CST:HL-PST were drug-dependent and ranged from 47 (flufenamic acid) to 3800 (mefenamic acid). Mefenamic acid is a relatively potent and selective inhibitor of HL-PST. 5. The IC50 for HL-PST obtained with mefenamic acid was three orders of magnitude lower than the peak plasma concentration of this drug after an oral dose of 0.5 g. Accordingly, mefenamic acid should impair sulphation in vivo. PMID- 10718118 TI - Bradykinin and alpha-thrombin increase human umbilical vein endothelial macromolecular permeability by different mechanisms. AB - Bradykinin and alpha-thrombin both increase endothelial macromolecular permeability, however the mechanism for this effect is unclear. Human umbilical vein endothelial cell (HUVEC) permeability to human serum albumin was increased by 1 microM alpha-thrombin (AT) or bradykinin (BK), but the kinetics of the permeability response were different. Intracellular calcium mobilization of HUVEC by AT was increased, yet BK had no effect on intracellular calcium. Distribution of F-actin and content was increased by AT as early as 10 minutes after administration, yet BK had no affect on F-actin when compared to control. We hypothesized that BK may increase HUVEC permeability by producing matrix metalloproteinase-2 (MMP-2). The AT-treated HUVEC produced an intermediate 64 kDa MMP-2, whereas BK-treated HUVEC increased the intermediate 64 kDa MMP-2 and also an active 62 kDa MMP-2. Pre-treatment of the HUVEC with tissue inhibitor of matrix metalloproteinase-2 slightly decreased the AT-induced increase in macromolecular permeability and completely inhibited the BK-induced increase in macromolecular permeability. PMID- 10718117 TI - Integrin expression on neutrophils in a rabbit model of Group B Streptococcal meningitis. AB - Products released by polymorphonuclear cells (PMNs) during an acute inflammatory response can result in diffuse tissue injury. Integrins are cell surface adhesion proteins that play a pivotal role in inflammation by allowing PMNs to adhere to the endothelium and migrate through the extracellular matrix. We examined the expression of beta1 and beta2 integrins on neutrophils from blood and cerebrospinal fluid (CSF) in an animal model of Group B Streptococcal meningitis. We further evaluated whether integrin expression correlates with pathophysiologic markers of central nervous system inflammation. Our data demonstrate that beta3 and beta2 integrin expression on circulating neutrophils does not significantly increase as a consequence of meningitis. In extravesated CSF neutrophils, a significant increase in expression of both beta1 and beta2 integrins is noted. Furthermore, a majority of the beta1 integrins on extravesated neutrophils have undergone affinity modulation. Using regression analysis, we demonstrated that increasing beta1 integrin expression correlates with decreasing CSF glucose concentration and serum/CSF glucose ratio. Regression analysis approached significance when CSF protein was compared to PMN beta1 integrin expression. Polymorphonuclear leukocytes beta1 integrin expression also showed a direct correlation to myeloperoxidase activity in brain tissue. Beta2 expression on CSF PMNs did not correlate with these markers of inflammation/sequestration. These data demonstrate integrin expression on extravesated neutrophils markedly increases during meningitis and support a role for beta1 integrins on neutrophils in the pathophysiologic consequences of meningitis. PMID- 10718120 TI - Comparative in vitro metabolism of indinavir in primates--a unique stereoselective hydroxylation in monkey. AB - 1. The in vitro metabolism of indinavir (CRIXIVAN, MK-0639, L-735,524), an HIV protease inhibitor, was evaluated using liver microsomes from cynomolgus monkey, rhesus monkey, chimpanzee and human. Indinavir exhibited marked species differences in metabolism. The overall rate of indinavir metabolism varied > 4 fold among primates (84 pmol/min/mg protein in cynomolgus monkey versus 20.4 pmol/min/mg protein in human) and followed the rank order: cynomolgus monkey > rhesus monkey > chimpanzee > human. 2. The cis-(indan)hydroxylated metabolite of indinavir was formed only in cynomolgus and rhesus monkey livers, whereas trans (indan)hydroxylation and N-dealkylation were observed as the major metabolites in all primates tested. Inhibition studies with P450-selective inhibitors (ketoconazole, quinine, quinidine) and monoclonal antibodies (against CYP2D6 or CYP3A4) indicated that a cytochrome P450 isoform of the CYP2D subfamily is involved in the formation of the unique cis-(indan) hydroxylated metabolite in monkey, whereas all other oxidative metabolites, including the trans (indan)hydroxylated metabolite, are formed by CYP3A isoform(s). 3. The present study has demonstrated that monkeys were unique in their abilities to form the stereoselective metabolite and were not appropriate surrogates for the qualitative prediction of indinavir metabolism in human. PMID- 10718122 TI - Automated high throughput human CYP isoform activity assay using SPE-LC/MS method: application in CYP inhibition evaluation. AB - 1. A high throughput screening (HTS) method for the evaluation of the seven major human hepatic CYP isoform activities was developed on a 96-well format, with automation. The method utilized pooled human liver microsomes and seven probe substrates, generic conditions for incubation, reaction termination and metabolite extraction with solid phase extraction (SPE) plates. Metabolites from the seven reactions were pooled and quantified using a generic liquid chromatography and tandem mass spectrometry (LCMS/MS) method. 2. The HTS method was validated based on Km values obtained, which were in agreement with literature data. 3. The isoform inhibition profiles of ketoconazole, quinidine, sulfaphenazole, tranylcypromine, alpha-naphthoflavone, and 4-methylpyrazole against CYPs 3A4, 2D6, 2C9, 2A6 land 2C19), 1A2 and 2E1, respectively, were obtained by this HTS method. Graphically obtained IC50 values are in agreement with literature reported values. 4. The HTS method represents a significant efficiency and selectivity improvement over traditional methods, and can be used for CYP inhibition assay and can be extended for liver activity profiling. PMID- 10718121 TI - Metabolite-intermediate complexation and inhibition of microsomal CYP3A in rat liver by diltiazem. AB - 1. The formation of metabolite intermediate (MI)-complexes between cytochrome P450 (CYP) and the alkylamine-substituted drugs diltiazem (DTZ) and desipramine (DES) and their effect on CYP activities was investigated in rat liver. 2. Dexamethasone and phenobarbitone pretreatment enhanced MI-complexation by DTZ (36% and 11% of total CYP complexed, respectively), whereas beta-naphthoflavone induction was without effect. All three treatments decreased MI-complexation produced by DES. 3. After a preincubation step in NADPH-supplemented microsomes DTZ and DES were effective inhibitors of the activities of CYPs 3A and 2C11 (testosterone 6beta- and 16alpha-hydroxylations, respectively). 4. Although MI complexation by DTZ was more extensive in microsomes from dexamethasone-induced rats, the apparent inhibition potency of the drug toward CYP activity was unchanged. By comparison, inhibition of CYP activity by DES was less pronounced than in control liver. 5. These findings indicate that drug-mediated MI complexation of CYPs does not necessarily potentiate the inhibitory effect on monooxygenase activity, although the duration of inhibition is longer. The extent of inhibition produced by stable drug metabolites may be similar to that from MI complexation. but their duration of action is limited by diffusion from the active site of the enzyme. PMID- 10718123 TI - DNA arrays: technology, options and toxicological applications. AB - The human genome contains an estimated 3 billion bases of DNA making up some 100000 genes, and the variation within this genome accounts for human diversity and, in many cases, disease. Defining and understanding the expression profile of given genotypes is essential to understanding adverse effects from acute or chronic exposure to environmental toxicants or other stimuli. DNA array technology could help researchers understand how organisms function in response to exposure by elucidating the molecular mechanisms that underlie them. DNA arrays have been developed and refined over the past 5 years and matured into a relatively accessible and affordable technology. They vary in design from membrane-based filters with a few hundred cDNAs, to glass-based 'chips' with tens of thousands of genetic elements. Mammalian DNA arrays will soon allow expression analysis on a genome-wide scale, similar to that already accomplished in some lower organisms (e.g. S. cerevisiae, E. coli). These whole-genome arrays will be powerful tools for identifying and characterizing toxicants in environmental and pharmaceutical science. This review discusses the technology behind the production of DNA arrays, the options available to those interested in applying them to their own research, and the possible toxicological applications of this exciting new technology. PMID- 10718124 TI - Metabolic fate of delmopinol in man after mouth rinsing and after oral administration. AB - 1. The metabolism of delmopinol, an inhibitor of plaque formation and gingivitis. has been studied after mouth rinsing or an oral dose of 14C-delmopinol to healthy male volunteers. The metabolite pattern was studied in urine and plasma samples (unhydrolysed or hydrolysed with conjugate cleaving enzymes) by liquid chromatography (LC) with radio detection. Metabolite identifications were carried out by gas chromatography-electron-impact mass spectrometry (GC-MS) and by liquid chromatography-thermospray mass spectrometry (LC-MS). 2. The metabolic clearance of delmopinol was high, and < 0.2% of the dose was excreted as intact delmopinol in the urine. The main metabolites were, for both administration routes, glucuronide conjugates of delmopinol and of (omega-1-hydroxy) delmopinol. These metabolites were predominant and accounted for nearly the entire urinary radioactivity and most of the plasma radioactivity. After mouth rinsing, parent delmopinol was also one of the main compounds in plasma. 3. Several other products of oxidation of the aliphatic side-chain were present in minor amounts in urine. These metabolites also appeared to be excreted as glucuronic acid conjugates. 4. Glucuronidated (omega-1-hydroxy) delmopinol separated into three peaks by the LC system used. This could be due to different chromatographic properties of conjugate isomers, positional or optical. PMID- 10718125 TI - Pharmacodynamic analysis between plasma level and inhibition of acid output after administration of a new histamine H2-receptor antagonist (Z-300) in dog. AB - 1. The relationship between plasma levels of a new H2-receptor antagonist, Z-300, and an active sulphoxide metabolite, and inhibitory effects on gastric acid secretion after intravenous or oral administration to dog have been examined. After both routes of administration, Z-300 in plasma eliminated with two phases, and the terminal half-lives were approximately 2 h. The level of sulphoxide in plasma reached a maximum at 0.6-0.7 h after administration, then subsequently eliminated with a half-life of approximately 6 h. 2. The maximal pharmacological effect of inhibition of gastric acid secretion (90-91%) was observed at 2 h (i.v.) and 6 h (p.o.), and the action persisted until 7 h after administration. 3. Since there was no correlation between plasma levels of Z-300 and pharmacological effects, the pharmacological effects were calculated by pharmacodynamic model including the effect compartment. The inhibition of acid output could be calculated by the amounts of Z-300 and sulphoxide corrected by pharmacological efficacy in the effect compartment. 4. These findings suggest that Z-300 contributes to the rapid appearance of the pharmacological effects in dog, whereas the sulphoxide, which eliminates slowly in plasma, contributes to duration. PMID- 10718126 TI - Metabolism of levormeloxifene, a selective oestrogen receptor modulator, in the Sprague-Dawley rat, Cynomolgus monkey and postmenopausal woman. AB - 1. The metabolic fate of levormeloxifene in the Sprague-Dawley rat, Cynomolgus monkey and postmenopausal volunteer has been investigated. 2. Two doses of [14C]levormeloxifene, 0.7 and 50 mg/kg, were given to the male and female rat and monkey, and a single 20-mg dose to the postmenopausal volunteer. 3. The primary route of excretion in all three species was the faeces. Metabolism was similar in all three species, with demethylation forming the major metabolite in the rat and postmenopausal volunteer. One of the major metabolites in the monkey involved an oxidative ring opening of a pyrrole ring. 4. The main site of metabolism of levormeloxifene is the liver and the majority of the drug and its metabolites is excreted via the faecal route. Metabolic pathways appear to be similar in the three species studied. PMID- 10718127 TI - Acetophenone-based linker for solid-phase peptide synthesis. AB - A new and cost-effective linker for the generation of carboxylic acid end groups on Multipin supports (SynPhase crowns) has been developed. Synthesis of the linker was based on modification of grafted polystyrene (PS) crowns to generate a hydroxyethyl moiety which is acid labile in 10-20% trifluoroacetic acid (TFA) in dichloromethane (DCM). Solid-phase syntheses of model decapeptides using this linker are described. PMID- 10718128 TI - CD, 1H NMR and molecular modeling studies of the interaction of Ca2+ with substance P and Ala7-substance P in a non-polar solvent. AB - The biologically relevant conformation of substance P is likely to be dictated by the lipid milieu wherein the hormone would interact with its receptor. Assuming that specific constraints to the hormone structure may be imparted by its interaction with Ca2+ ions in the low dielectric lipid medium, the interaction of substance P and its inactive analog, Ala7-substance P, has been characterized in a lipid-mimetic solvent. Circular dichroism (CD) and NMR spectral methods were employed to study the conformation of the free and Ca2+-bound forms of the peptides and the conformational changes that occur on Ca2+ binding. The results show that both peptides assume a helical structure in the non-polar solvent used, a mixture of acetonitrile and trifluoroethanol. The N-terminal region is, however, less ordered in the analog peptide compared with the native hormone. Ca2+ addition causes significant conformational changes in both the peptides. However, while substance P binds two Ca2+ ions in a cooperative manner, Ala7 substance P binds only one Ca2+ ion with a relatively weaker affinity. Computations of the minimum-energy conformations of the free and Ca2+-bound peptides were performed using interproton distances derived from nuclear Overhauser enhancement spectra of the two peptides, as well as the information provided by changes in proton chemical shifts caused by Ca2+ addition. Taken together, the results of this study suggest that differences in the interaction of substance P and Ala7-substance P with Ca2+ in the non-polar milieu, which in turn leads to differences in their Ca2+-bound conformations, may be the basis for the differences in their biological potencies. PMID- 10718129 TI - Combined solid-phase and solution approach for the synthesis of large peptides or proteins. AB - In the synthesis of large peptides or proteins, highly homogeneous segments are indispensable for a convergent strategy either on a solid-phase resin or in solution. Employing Boc/Bzl chemistry to prepare fully protected segments with a free alpha-carboxyl group from the solid support, base-labile linkers are profitable for practical peptide synthesis since they require no special equipment. For this purpose, an N-[9-(hydroxymethyl)-2-fluorenyl]succinamic acid (HMFS) linker was adopted. Consequently, there must be high compatibility between the protecting groups of the segment and the anchoring group which is cleavable by treatment with morpholine or piperidine in DMF. Instead of using the 2 bromobenzyloxycarbonyl (BrZ) group for the Tyr residue and the formyl (For) group for the Trp residue, both of which are the most susceptible protecting groups under these base-catalysed conditions, the base-resistant 3-pentyl (Pen) and cyclohexyloxycarbonyl (Hoc) groups were introduced to the respective side-chain functional groups. By applying the present strategy, the authors were able to rapidly synthesize homogeneous protected segments for use in the subsequent segment coupling in solution. In the present paper, the utility of the combined solid-phase and solution approach is demonstrated by synthesizing muscarinic toxin 1 (MTX1) which binds to the muscarinic acetylcholine receptors. PMID- 10718130 TI - Standards for image quality and radiation exposure impede the pursuit of optimized quality/dose ratios in radiology. PMID- 10718131 TI - Performance analysis of a new semiorthogonal spline wavelet compression algorithm for tonal medical images. AB - Lossy image compression is thought to be a necessity as radiology moves toward a filmless environment. Compression algorithms based on the discrete cosine transform (DCT) are limited due to the infinite support of the cosine basis function. Wavelets, basis functions that have compact or nearly compact support, are mathematically better suited for decorrelating medical image data. A lossy compression algorithm based on semiorthogonal cubic spline wavelets has been implemented and tested on six different image modalities (magnetic resonance, x ray computed tomography, single photon emission tomography, digital fluoroscopy, computed radiography, and ultrasound). The fidelity of the reconstructed wavelet images was compared to images compressed with a DCT algorithm for compression ratios of up to 40:1. The wavelet algorithm was found to have generally lower average error metrics and higher peak-signal-to-noise ratios than the DCT algorithm. PMID- 10718132 TI - Strategies to improve the signal and noise performance of active matrix, flat panel imagers for diagnostic x-ray applications. AB - A theoretical investigation of factors limiting the detective quantum efficiency (DQE) of active matrix flat-panel imagers (AMFPIs), and of methods to overcome these limitations, is reported. At the higher exposure levels associated with radiography, the present generation of AMFPIs is capable of exhibiting DQE performance equivalent, or superior, to that of existing film-screen and computed radiography systems. However, at exposure levels commonly encountered in fluoroscopy, AMFPIs exhibit significantly reduced DQE and this problem is accentuated at higher spatial frequencies. The problem applies both to AMFPIs that rely on indirect detection as well as direct detection of the incident radiation. This reduced performance derives from the relatively large magnitude of the square of the total additive noise compared to the system gain for existing AMFPIs. In order to circumvent these restrictions, a variety of strategies to decrease additive noise and enhance system gain are proposed. Additive noise could be reduced through improved preamplifier, pixel and array design, including the incorporation of compensation lines to sample external line noise. System gain could be enhanced through the use of continuous photodiodes, pixel amplifiers, or higher gain x-ray converters such as lead iodide. The feasibility of these and other strategies is discussed and potential improvements to DQE performance are quantified through a theoretical investigation of a variety of hypothetical 200 microm pitch designs. At low exposures, such improvements could greatly increase the magnitude of the low spatial frequency component of the DQE, rendering it practically independent of exposure while simultaneously reducing the falloff in DQE at higher spatial frequencies. Furthermore, such noise reduction and gain enhancement could lead to the development of AMFPIs with high DQE performance which are capable of providing both high resolution radiographic images, at approximately 100 microm pixel resolution, as well as variable resolution fluoroscopic images at 30 fps. PMID- 10718133 TI - An interlaboratory measurement of screen-film speed and average gradient according to ISO 9236-1. AB - The speed and average gradient of a conventional screen-film system was measured at four European laboratories. This is the first interlaboratory comparison in which the measurement conditions described in ISO 9236-1 were applied. The four laboratories used calibrated measurement equipment. The values obtained by the four laboratories were within a range of 14% for the speed and within a range of 8% for the average gradient. These variations are consistent with the expected measurement uncertainty. PMID- 10718134 TI - Photon scatter in portal images: physical characteristics of pencil beam kernels generated using the EGS Monte Carlo code. AB - Pencil beam kernels describing scattered photon fluence behind homogeneous water slabs at various air gap distances were generated using the EGS Monte Carlo code. Photon scatter fluence was scored in separate bins based on the particle's history: singly scattered, multiply scattered, and bremsstrahlung and positron annihilation photons. Simultaneously, the mean energy and mean angle with respect to the incident photon pencil beam were tallied. Kernels were generated for incident photon pencil beams exhibiting monoenergetic spectra of 2.0 and 10.0 MeV, and polyenergetic spectra representative of 6 and 24 MV beams. Reciprocity was used to generate scatter fractions on the central axis for various field sizes, phantom thicknesses, and air gaps. The scatter kernels were further characterized by full width at half-maximum estimates. Modulation transfer functions were calculated, providing theoretical estimates of the limit of performance of portal imaging systems due to the intrinsic scattering of photon radiation through the patient. PMID- 10718135 TI - Automatic on-line electronic portal image analysis with a wavelet-based edge detector. AB - A fully automatic method for on-line electronic portal image analysis is proposed. The method uses multiscale edge detection with wavelets for both the field outline and the anatomical structures. An algorithm to extract and combine the information from different scales has been developed. The edges from the portal image are aligned with the edges from the reference image using chamfer matching. The reference is the first portal image of each treatment. The matching is applied first to the field and subsequently to the anatomy. The setup deviations are quantified as the displacement of the anatomical structures relative to the radiation beam boundaries. The performance of the algorithm was investigated for portal images with different contrast and noise level. The automatic analysis was used first to detect simulated displacements. Then the automatic procedure was tested on anterior-posterior and lateral portal images of a pelvic phantom. In both sets of tests the differences between the measured and the actual shifts were used to quantify the performance. Finally we applied the automatic procedure to clinical images of pelvic and lung regions. The output of the procedure was compared with the results of a manual match performed by a trained operator. The errors for the phantom tests were small: average standard deviation of 0.39 mm and 0.26 degrees and absolute mean error of 0.31 mm and 0.2 degrees were obtained. In the clinical cases average standard deviations of 1.32 mm and 0.6 degrees were found. The average absolute mean errors were 1.09 mm and 0.39 degrees. Failures were registered in 2% of the phantom tests and in 3% of the clinical cases. The algorithm execution is approximately 5 s on a 168 MHz Sun Ultra 2 workstation. The automatic analysis tool is considered to be a very useful tool for on-line setup corrections. PMID- 10718136 TI - Monte Carlo studies of the exit photon spectra and dose to a metal/phosphor portal imaging screen. AB - The energy spectra and the dose to a Cu plate/Gd2O2S phosphor portal imaging detector were investigated for monoenergetic incident beams of photons (1.25, 2, and 5 MeV). The Monte Carlo method was used to characterize the influence of the patient/detector geometry, detector material and design, and incident beam energy on the spectral distribution and the dose, at the imaging detector plane, of a photon beam scattered from a water phantom. The results show that radiation equilibrium is lost in the air gap and that, for the geometries studied, this effect led to a reduction in the exit dose of up to 40%. The finding that the effects of the air gap and field size are roughly complementary has led to the hypothesis that an equivalent field size concept may be used to account for intensity and spectral changes arising from air gap variations. The copper plate preferentially attenuates the low-energy scattered photons incident on it, while producing additional annihilation, bremsstrahlung, and scattered photons. As a result, the scatter spectra at the copper surface entrance of the detector differs significantly from that at the Cu/phosphor interface. In addition, the mean scattered photon energy at the interface was observed to be roughly 0.4 MeV higher than the corresponding effective energy for 2 MeV incident beams. A comparison of the dose to various detector materials showed that exit dosimetry errors of up to 24% will occur if it is assumed that the Cu plate/Gd2O2S phosphor detector is water equivalent. PMID- 10718137 TI - Dose calculation for asymmetric photon fields with independent jaws and multileaf collimators. AB - We have developed a simple method for dose calculation in dual asymmetric open and irregular fields with four independent jaws and multileaf collimators. Our calculation method extends the scatter correction method of Kwa et al. [Med. Phys. 21, 1599-1604 (1994)] based on the principle of Day's equivalent-field calculation. The scatter correction factor was determined by the ratio of the derived doses of a smaller asymmetric open field or irregular field to a larger symmetric field. The algorithm with the scatter correction method can be calculated from output factors, tissue maximum ratios, and off-axis ratios for conventional symmetric fields. The doses calculated by this method were compared with the measured doses for various asymmetric open and irregular fields. The agreement between the calculated and measured doses for 4 and 10 MV photon beams was within 0.5% at the geometric center of the asymmetric open fields. For the asymmetric irregular fields with the same geometrical center, agreement within 1% was found in most cases. PMID- 10718138 TI - Breathing-synchronized radiotherapy program at the University of California Davis Cancer Center. AB - In this paper we present a complete description of the breathing synchronized radiotherapy (BSRT) system, which has been jointly developed between the University of California Davis Cancer Center and Varian Associates. BSRT is a description of an emerging radiation oncology procedure, where simulation, CT scan, treatment planning, and radiation treatment are synchronized with voluntary breath-hold, forced breath-hold, or breathing gating. The BSRT system consists of a breathing monitoring system (BMOS) and a linear accelerator gating hardware and software package. Two methods, a video camera-based method and the use of wraparound inductive plethysmography (RespiTrace), generate the BMOS signals. The BMOS signals and the synchronized fluoroscopic images are simultaneously recorded in the simulation room and are later analyzed to define the ideal treatment point (ITP) where organ motion is stationary. The BMOS signals at ITP can be used to gate a CT scanner or a linear accelerator to maintain the same organ configuration as in the simulation. The BSRT system allows breath-hold or gating. This dual role allows the system to be applicable for a variety of patients, i.e., the breath-hold method for those patients who can maintain and reproduce the ITP, and the forced breath-hold or gating method for those who are not ideal for voluntary breath-hold. PMID- 10718139 TI - A computerized remote table control for fast on-line patient repositioning: implementation and clinical feasibility. AB - A computerized remote control for a Siemens ZXT treatment couch was implemented and its characteristics were investigated to establish its feasibility for on line setup corrections, using portal imaging. Communication with the table was obtained by connecting it via a serial line to a work station. The treatment couch enables "goto" commands in the three main directions and around the isocenter. The accuracy of the movements after giving such a command was checked and the time for each movement was recorded. First, the movements into a single direction were studied (range of -4 to +4 cm and -4 degrees to +4 degrees). Each command was repeated four times. Second, the table was moved into the three main directions simultaneously. For this experiment a clinically relevant three dimensional (3-D) normal distribution of shifts was used [N = 200, standard deviation (SD) 5 mm in the three main directions]. This latter experiment was done twice: without and with rotations (a distribution with SD 1 degrees). During the first experiment, with shifts into one direction, no systematic deviations were found. The overall accuracy of the shifts was 0.6 mm (1 SD) in each direction and 0.04 degrees (1 SD) for the rotations. The time required for a translation ranged between 4 and 13 s and for the rotation between 8 and 20 s. The second experiment with the 3-D distribution of setup errors yielded an error in the 3-D vector length equal to 0.96 mm (1 SD), independent of rotations. Shifts were performed in less than 11 s for 95% of the cases without rotations. When rotations were also performed, 95% of the movements finished in less than 16 s. In conclusion, the table movements are accurate and enable on-line setup corrections in daily clinical practice. PMID- 10718140 TI - Modification of the University of Washington Neutron Radiotherapy Facility for optimization of neutron capture enhanced fast-neutron therapy. AB - A modified neutron production target assembly has been developed to provide improved performance of the proton-cyclotron-based neutron radiotherapy facility at the University of Washington for applications involving neutron capture enhanced fast-neutron therapy. The new target produces a neutron beam that yields essentially the same fast-neutron physical depth-dose distribution as is produced by the current UW clinical system, but that also has an increased fraction of BNCT enhancement relative to the total therapeutic dose. The modified target is composed of a 5-millimeter layer of beryllium, followed by a 2.5-millimeter layer of tungsten, with a water-cooled copper backing. Measurements of the free-field neutron spectrum of the beam produced by the new target were performed using activation foils with a direct spectral unfolding technique. Water phantom measurements were performed using a tissue-equivalent ion chamber to characterize the fast-neutron depth-dose curve and sodium activation in soda-lime glass beads to characterize the thermal-neutron flux (and thus the expected neutron capture dose enhancement) as a function of depth. The results of the various measurements were quite consistent with expectations based on the design calculations for the modified target. The spectrum of the neutron beam produced by the new target features an enhanced low-energy flux component relative to the spectrum of the beam produced by the standard UW target. However, it has essentially the same high-energy neutron flux, with a reduced flux component in the mid-range of the energy spectrum. As a result, the measured physical depth-dose curve in a large water phantom has the same shape compared to the case of the standard UW clinical beam, but approximately twice the level of BNCT enhancement per unit background neutron dose at depths of clinical interest. In-vivo clinical testing of BNCT enhanced fast-neutron therapy for canine lung tumors using the new beam was recently initiated. PMID- 10718141 TI - Multi-layer energy filter for realizing conformal irradiation in charged particle therapy. AB - A new type of filter for charged particle radiotherapy is developed to reduce unwanted dose transfer to the normal tissues around a tumor. The new filter can make a static irradiation field where the width of the spread-out Bragg peak (SOBP) is two-dimensionally adjusted. That makes the field conformal to the tumor three-dimensionally. The filter is made of many layers produced by using stereolithography. The layer has a miniaturized structure that has geometrical similarity to the conventional ridge filter. Shapes of cone and pyramid are also usable for the unit-cell constructing the layer. The spread of the field in the depth direction is decided by the thickness of the filter, or by the number of layers. The experimental result of the irradiation using the ridge-type construction shows a good agreement with an estimate by the Monte Carlo calculation. By combining this technique with intensity modulation that has lateral position dependence, the conformal irradiation can be achieved by a simple procedure. PMID- 10718142 TI - Dosimetric perturbations of linear array of beta-emitter seeds and metallic stent in intravascular brachytherapy. AB - The radiation treatment with catheter-based beta-emitter sources is under clinical trials to prevent restenosis following interventional coronary procedures. There are still large uncertainties in the dose calculation due to the complicated treatment geometry. We present the Monte Carlo simulations to account for the dosimetric perturbations due to neighboring trained seeds, proximal/distal gold markers, and a stainless steel stent. A catheter-based beta emitter system is modeled using the Monte Carlo code, MCNP4B. Dose distributions and dose rates are calculated in voxels (0.64x0.64x0.5 mm3) around the long cylindrical trains of 90Sr/Y source with and without the stent (at 1.92 mm from the source axis). For the total activity of 70 mCi (2.59x10(9) Bq), the dose around most of the source length (except for edge seeds and gold markers) varies from 40 to 0.23 cGy/s as the radial distance from the source axis (r) increases from 0.64 to 6.4 mm. At the prescription range of r = 1.5-4.0 mm, the dose gradient is very steep and the contribution of neighboring seeds to the dose is significant. The dose enhancement due to neighboring seeds (the so-called "train effect") varies from 9% to 64% as r increases from 0.64 to 5.2 mm. The doses at r = 2 mm from the last edge seed and the gold marker are about 80% and 40% of that of the nonedge seed (8.7 cGy/s), respectively. The dose enhancement due to the secondary electrons and the primary electrons scattered with the stent is shown to be about 9.3% in the voxel including the stent. However, as r increases beyond the stent (r = 2.0-6.4 mm), the dose is slightly reduced by 4%-12%, compared to that without the stent. PMID- 10718143 TI - Comparison of seed loading approaches in prostate brachytherapy. AB - Since uniform seed loading in prostate brachytherapy can produce an intolerably high dose along the urethra, some form of peripheral loading is commonly employed. We define three variants of peripheral loading and compare them in a small, medium, and large prostate in terms of coverage of the planning target volume (PTV), homogeneity, and ability to spare critical structures of excessive dose. Modified uniform loading has at least 2/3 of the seeds occupying sites on a 1 cm cubic grid keyed to the prostate base and the posterior border of the prostate. Nonuniform loading explicitly spares the urethra by using only basal and apical seeds in at least two centrally located needles. Peripheral loading uses higher activity seeds with the posterior implant plane 5 mm anterior to the posterior border of the prostate. The three prostate volumes (18.7, 40.7, and 60.2 cm3 by ultrasound) were expanded to planning volumes (32.9, 60.0, and 87.8 cm3, respectively). The planning volumes (PTVs) were loaded with a 125I seed distribution and activity sufficient to cover 99.7+/-0.3% of the PTV with the prescribed minimal peripheral dose (mPD) of 145 Gy. Activities used ranged from 0.32 to 0.37 mCi/seed (0.41-0.47 U/seed) for the first two approaches and from 0.57 to 0.66 mCi (0.72-0.84 U) for peripheral loading. Modified uniform loading produced the most uniform distribution based on dose-volume histograms and the volume receiving >150% of prescribed dose. All the approaches are capable of constraining the superior-inferior dose profile (the urethral path) to less than 150% of the mPD, but the nonuniform approach with explicit urethral sparing kept the urethral dose below 120% of the mPD. Dose profiles for the three approaches along the posterior-anterior midline axis are comparable near the urethra, but peripheral and nonuniform approaches have extended regions where the dose is >150% of mPD. These regions approach within 10 mm of the rectum or urethra, so these two approaches require greater accuracy in intraoperative execution of the plan. Although each of the three planning approaches can achieve the treatment goals of adequate coverage and critical structure sparing, modified uniform loading has a more homogeneous dose distribution. This approach may be more forgiving of systematic errors in seed placement. PMID- 10718144 TI - Modeling dose response in the presence of spatial variations in dose rate. AB - Nonuniform dose rates are an inevitability in treatments involving internal sources, arising from electronic disequilibrium effects as well as nonuniformity in activity distribution. These dose-rate nonuniformities are of consequence for protracted treatments (when dose-delivery times are of the order of cell-repair times). The influence of nonuniform dose rates on tumor control probability (TCP) has thus been considered. A model for TCP has been developed by merging established (linear-quadratic based) TCP models for dose nonuniformity, with dose rate effects as influenced by cell repair and proliferation capacities. This model has been examined by considering treatment of spherical tumors of varying sizes filled with uniform distributions of several beta-emitting isotopes. Dose (or dose-rate) volume histograms (DVHs) were calculated for the combinations of tumor size and isotope, and applied to the developed TCP model. Comparison of the results identified several characteristics of the effect of nonuniform dose rate, including the balance between minimum dose and cell number as they vary with tumor size, the dominance of minimum dose (dose rate) on TCP, and the influence of cell-proliferation effects on effective delivered dose (and the effective DVH). The model was also used to determine TCPs for simulated 90Y-labeled microsphere treatments of liver metastases using both uniform and clustered microsphere models for activity distributions, and for varying tumor size. Despite significantly higher doses being achieved via clustered (nonuniform) activity distributions, the minimum dose for clustered distributions is consistently lower than that of the corresponding uniform distributions, and TCP is always higher for the uniform distributions. PMID- 10718145 TI - A model for photon detection and dosimetry with superheated emulsions. AB - A model is presented for the analysis and prediction of the photon response of detectors based on superheated emulsions of light halocarbons in tissue equivalent gels. It is shown that on the basis of a nondimensional thermodynamic quantity, called reduced superheat, it is possible to identify the degree of superheat, or the operating temperature, corresponding to the photon sensitization of the emulsions. Moreover, on the basis of the mass energy absorption coefficients, it is possible to determine the energy dependence of the photon response. The vaporization energy necessary for bubble nucleation is estimated by means of the thermal spike theory developed for bubble chambers. The energy deposition requirements are consistent with the energy transferred by secondary electrons at the end of their range in the halocarbons. These findings provide design criteria for photon detectors based on superheated emulsions. It is shown that light halocarbons of low effective atomic numbers present the best dosimetric properties. In particular, by manufacturing superheated emulsions with octafluoropropane, or halocarbon R-218, photon sensitivity is achieved at room temperature along with a fairly constant air-kerma response. PMID- 10718146 TI - Oximetry based on diffuse photon density wave differentials. AB - The quantification of tissue optical properties for calculating blood saturation and hemoglobin concentration using measurements of diffuse photon density waves at some distance away from an intensity-modulated light source, generally requires the determination of the amplitude and phase of this light source. This determination may become a severe impediment for measurements performed in the clinical environment. In this work we extend a self-calibrating methodology developed for constant wave and modulation depth-phase measurements, to include amplitude and phase measurements of diffuse photon density waves. The method uses amplitude and phase changes of intensity modulated light, under the assumption of known index of refraction and invariant reduced scattering coefficient mu's, to quantify the absorption coefficient mu(a) without requiring initial amplitude and phase knowledge. Quantification of the mu(a) at selected time points during a measurement can then be employed to calibrate numerical solutions of the diffusion equation and compute the mu(a) for the remaining time points of the experiment. It is shown that the method is quite insensitive to the knowledge of the exact mu's value so that an assumption on the average mu's value for the tissue measured may be employed. The sensitivity of calculating blood saturation and hemoglobin concentration, as a function of the deviation of the mu's used in the calculation versus the real mu's value is investigated using simulated data. It is also demonstrated that the saturation calculation is especially insensitive to the mu's guess. The performance of the method to quantify blood oxygen saturation and the concentrations of oxy- and deoxy-hemoglobin is examined with experimental measurements at two wavelengths on specially constructed blood model phantoms. To validate the method the measurements are monitored by a time resolved spectrometer. The method is shown to be accurate to within +/-5% in calculating blood saturation and to within +/-10% in calculating hemoglobin concentration compared to the results obtained with the time-resolved spectrometer and the expected theoretical values. PMID- 10718147 TI - The beta-amyloid precursor protein and its derivatives: from biology to learning and memory processes. AB - Intensive investigation towards the understanding of the biology and physiological functions of the beta-amyloid precursor protein (APP) have been supported since it is known that a 39-43 amino acid fragment of APP, called the beta-amyloid protein (Abeta), accumulates in the brain parenchyma to form the typical lesions associated with Alzheimer's disease (AD). It emerges from extensive data that APP and its derivatives show a wide range of contrasting physiological properties and therefore might be involved in distinct physiological functions. Abeta has been shown to disrupt neuronal activity and to demonstrate neurotoxic properties in a wide range of experimental procedures. In contrast, both in vitro and in vivo studies suggest that APP and/or its secreted forms are important factors involved in the viability, growth and morphological and functional plasticity of nerve cells. Furthermore, several recent studies suggest that APP and its derivatives have an important role in learning and memory processes. Memory impairments can be induced in animals by intracerebral treatment with Abeta. Altered expression of the APP gene in aged animals or in genetically-modified animals also leads to memory deficits. By contrast, secreted forms of APP have recently been shown to facilitate learning and memory processes in mice. These interesting findings open novel perspectives to understand the involvement of APP in the development of cognitive deficits associated with AD. In this review, we summarize the current data concerning the biology and the behavioral effects of APP and its derivatives which may be relevant to the roles of these proteins in memory and in AD pathology. PMID- 10718149 TI - Neural mechanisms of memory retrieval: role of the prefrontal cortex. AB - In the primate brain, long-term memory is stored in the neocortical association area which is also engaged in sensory perception. The coded representation of memory is retrieved via interactions of hierarchically different cortical areas along bottom-up and top-down anatomical connections. The functional significance of the fronto-cortical top-down neuronal projections has been relevantly assessed in a new experimental paradigm using posterior-split-brain monkeys. When the splenium of the corpus callosum and the anterior commissure were selectively split, the bottom-up visual signal originating from the unilateral striate cortex could not reach the contralateral visual cortical areas. In this preparation, long-term memory acquired through visual stimulus-stimulus association learning was prevented from transferring across hemispheres. Nonetheless, following the presentation of a visual cue to one hemisphere, the prefrontal cortex could instruct the contralateral hemisphere to retrieve the correct stimulus specified by the cue. These results support the hypothesis that the prefrontal cortex can regulate memory recall in the absence of bottom-up sensory input. In humans, functional neuroimaging studies have revealed activation of a distributed neural network, including the prefrontal cortex, during memory retrieval tasks. Thus, the prefrontal cortex is consistently involved in retrieval of long-term memory in primates. PMID- 10718148 TI - Sensory and memory properties of hippocampal place cells. AB - The rat hippocampus contains place cells whose firing is location-specific. These cells fire only when the rat enters a restricted region of the environment called the firing field. In this review, we examine the sensory information that is fundamental to the place cell system for producing spatial firing. While visual information takes precedence in the control of firing fields when it is available, local (olfactory and/or tactile) cues combined with motion-related cues can permit stable spatial firing. Motion-related cues are integrated by hippocampal place cells, but in the absence of external cues do not support stable firing over long periods. While firing fields are based on a variety of sensory cues, they do not strictly depend on such cues. Rather, sensory information is important for activating the representation appropriate to the current environment as reflected by the firing properties of place cell ensembles. Specific sensory channels as well as the memory properties of place cells can support ongoing firing under manipulations of the environment. These memory features raise the question of the role of the place cell system in the acquisition, storage and retrieval of spatial information. Based on the existing literature about the effects of hippocampal lesions and about the metabolic activations in spatial memory tasks, we suggest that a function of the place cell system is to automatically provide the organism with information about its current location so as to allow for the rapid acquisition of novel information. PMID- 10718150 TI - The control of low-level information flow in the visual system. AB - Visual information processing needs to be error free and efficient. Our visual system tries to achieve the first goal by accommodating a wide variety of visual algorithms for the extraction of the relevant features in the scene, while at the same time the second goal is addressed by controlling the amount of visual information flow in the network employing selective attention. Attentional or pre attentional mechanisms are found throughout many visual areas and these processes may start as early as in the visual thalamus (lateral geniculate nucleus, LGN). In this review we pay particular attention to experimental and theoretical findings which indicate that even low-level structures, such as LGN and V1, can play a major role in the flow-control of visual information. PMID- 10718151 TI - Proton magnetic resonance spectroscopy of the human brain in schizophrenia. AB - In vivo proton magnetic resonance spectroscopy (1H MRS) has been utilized by neuroimaging laboratories in recent years to reliably measure compounds such as N acetylaspartate (NAA), choline (Cho), creatine (Cr), and to a lesser extent glutamate and glutamine in the human brain. To date, the most consistently replicated findings in schizophrenia are reduced NAA measures in the hippocampal regions. Since NAA is thought to be a neuronal/axonal marker and a measure of neuronal/axonal integrity, hippocampal NAA reductions have been interpreted as strong evidence for neuronal/axonal loss or dysfunction in this brain region. The evidence for neuronal loss or dysfunction based on NAA is less consistent for the frontal cortex and white matter, temporal cortex, basal ganglia, cingulate region, and thalamus in schizophrenia. Furthermore, there are no consistently replicated findings for choline or creatine alterations in any of the brain regions examined in schizophrenia. Finally, significant technical difficulties make reliable measurement of glutamine and glutamate problematic at the present time. PMID- 10718152 TI - Regulation of quantal size by presynaptic mechanisms. AB - Quantal size is often modeled as invariant, although it is now well established that the number of transmitter molecules released per synaptic vesicle during exocytosis can be modulated in central and peripheral synapses. In this review, we suggest why presynaptically altered quantal size would be important at social synapses that provide extrasynaptic neurotransmitter. Current techniques used to measure quantal size are reviewed with particular attention to amperometry, the first approach to provide direct measurement of the number of molecules and kinetics of presynaptic quantal release, and to CNS dopamine neuronal terminals. The known interventions that alter quantal size at the presynaptic locus are reviewed and categorized as (1) alteration of transvesicular free energy gradients, (2) modulation of vesicle transmitter transporter activity, (3) modulation of fusion pore kinetics, (4) altered transmitter degranulation, and (5) changes in synaptic vesicle volume. Modulation of the number of molecules released per quantum underlies mechanisms of drug action of L-DOPA and the amphetamines, and seems likely to be involved in both normal synaptic modification and disease states. Statistical analysis for examining quantal size and data presentation is discussed. We include detailed information on performing nonparametric resampling statistical analysis, the Kolmogorov-Smirnov test for two populations, and random walk simulations using spreadsheet programs. PMID- 10718153 TI - Paradigm shifts in stem-cell biology. AB - Hematopoiesis is a physiologic process that can be transplanted by intravenous infusion of stem and progenitor cells. Because these cells contribute to blood production over a lifespan, they are attractive targets for cell-based therapies of hematologic malignancies and genetic defects. A more complete understanding of the basic biology of hematopoiesis will accelerate our progress toward the clinical goal of improved stem-cell-based therapies. Many advances in recent years have brought us closer to that goal and have, in addition, challenged a number of dogmatic notions about hematopoiesis. Three of these advances are briefly addressed here: (1) an emerging appreciation of the complex relationship between cell-cycle status, engraftment potential, and self-renewal in the hematopoietic system; (2) the demonstration of new progenitor populations and lineage relationships in early hematopoietic development; and (3) a reanalysis of the embryonic origins of hematopoiesis. These and other advances are allowing the mysteries of hematopoiesis to be unlocked at a pace that was unimaginable just a few years ago. PMID- 10718154 TI - Mechanisms of stem-/progenitor-cell mobilization: the anti-VLA-4 paradigm. AB - Since the introduction of mobilized peripheral blood stem cells for transplantation purposes, many studies have been performed using mobilized cells with different mobilization schemes, primarily to optimize mobilization protocols. Studies aiming at mechanisms of mobilization have been few, but have provided useful insights. However, conclusions about mobilization mechanisms were largely inferential. We have attempted to analyze the mobilization process Involving the VLA-4/VCAM-1 pathway. Our findings are summarized and an attempt is made to put our experience into a general model of mobilization. PMID- 10718155 TI - The role of metalloproteinases and adhesion molecules in interleukin-8-induced stem-cell mobilization. AB - The chemokine interleukin-8 (IL-8) is a potent chemoattractant and activator of neutrophils. Upon systemic injection, IL-8 induces an immediate neutropenia followed by a rebound granulocytosis. In this report, we discuss the effects of IL-8 on the mobilization of hematopoietic stem cells. Within 20 minutes following a single intraperitoneal injection in mice, IL-8 induces the mobilization of hematopoietic progenitor cells (HPC) with colony-forming, radioprotective, and long-term lymphomyeloid resubpopulating ability. Mobilization can be specifically prevented by pretreatment with antibodies against the beta2 integrin LFA-1 (CD11a). In monkeys, IL-8 Induces the rapid release of the metalloproteinase gelatinase-B concurrent with the mobilization of HPC. The latter effect can be prevented by blocking gelatinase-B activity using specific monoclonal antibodies, suggesting the involvement of gelatinase-B as a mediator of HPC mobilization. These results are consistent with the hypothesis that neutrophils are major regulators of stem-cell mobilization through the release of metalloproteinases (MMPs) that cleave extracellular matrix molecules to which HPC are attached. PMID- 10718156 TI - Mechanisms of granulocyte colony-stimulating factor-induced hematopoietic progenitor-cell mobilization. AB - Hematopoietic progenitor cells (HPC) can be mobilized from the bone marrow into the peripheral circulation in response to diverse stimuli, including hematopoietic growth factors, cytotoxic agents, and certain chemokines. Despite significant differences in their biologic activities, these stimuli result in the mobilization of HPC with a similar phenotype, suggesting that a common mechanism for mobilization may exist. To explore the mechanisms of granulocyte colony stimulating factor (G-CSF)-induced mobilization, we examined HPC mobilization in mice that are genetically deficient for the G-CSF receptor (G-CSFR). Their response was determined to each of three major types of mobilizing stimuli: cytotoxic agents (cyclophosphamide), chemokines (Interleukin-8[IL-8]), and hematopoietic growth factors (G-CSF, fit-3 ligand, and IL-12). These studies demonstrate that the G-CSFR is required for mobilization in response to cyclophosphamide and IL-8, but not fit-3 ligand or IL-12, and suggest that the G CSFR may play an important and previously unexpected role in HPC migration. PMID- 10718157 TI - Innovations in allogeneic stem-cell transplantation. AB - Allogeneic bone marrow transplantation (BMT) is associated with prolonged periods of neutropenia and thrombocytopenia, which can lead to severe infections and bleeding complications. Transplantation-related side effects might be ameliorated by use of cytokine-mobilized peripheral blood progenitor cells (PBPC) Instead of bone marrow. We have studied PBPC mobilization and transplantation in more than 150 patients with high-risk hematologic malignancies. Normal donors can be sufficiently mobilized with granulocyte colony-stimulating factor (G-CSF), with 91% of G-CSF-stimulated normal donors producing more than 2 x 10(6) CD34+ cells/kg by a single apheresis. The combination of G-CSF plus granulocyte macrophage colony-stimulating factor (GM-CSF) was more effective than mobilization with G-CSF alone. A clear relationship was seen between numbers of resting CD34+ cells premobilization and numbers of PBPC collected by apheresis, indicating that resting CD34+ cells might be used to predict mobilization results and identify donors who could benefit from more effective mobilization regimens. Transplantation of G-CSF-mobilized PBPC was associated with a more rapid engraftment than that observed for BMT. While engraftment was safe and acute graft-versus-host disease (aGvHD) rates were not increased over BMT, chronic GvHD rates were higher after PBPC transplantation. An additional PBPC infusion on day +3 resulted in a further shortening of neutropenia and thrombocytopenia. Incorporation of these innovative approaches with "minimal" conditioning regimens has resulted in near-complete elimination of fever, neutropenia, thrombocytopenia, and the need for antibiotics and RBC and platelet transfusions after allogeneic transplantation. PMID- 10718158 TI - Ex vivo manipulation of hematopoietic stem and progenitor cells. AB - Ex vivo manipulations of hematopoietic stem and progenitor cells are Increasingly used in the context of autologous and allogeneic stem-cell transplantation. These manipulations include the positive selection of CD34+ cells for tumor-cell reduction and/or T-cell depletion, the ex vivo expansion of hematopoietic progenitor and stem cells under appropriate cytokine-stimulated culture conditions, and the ex vivo generation of myeloid or megakaryocytic postprogenitor cells and Immune effector cells. This article summarizes both the preclinical data on the ex vivo expansion of hematopoietic progenitor and stem cells from purified CD34+ cells and the Initial clinical studies with ex vivo expanded stem and progenitor cells for hematopoietic support after high-dose chemotherapy. PMID- 10718159 TI - Exposure to environmental risk factors and parenting attitudes among substance abusing women. AB - This study examined the amount of exposure to negative environmental risks and their association with parenting attitudes among a group of inner city substance abusing women. Mothers (N = 198) were recruited at delivery and were part of a randomized longitudinal intervention study for substance-abusing women and their infants. When the infants were 18 months old, a cumulative environmental risk score was calculated for each mother based on nine factors: violence (both domestic and environmental), depression, homelessness, incarceration, number of children, life stress, psychiatric problems, and absence of significant other. Based on their cumulative scores, mothers were placed in a low (N = 106) or high environmental risk group (N = 92). Mothers in the high-risk group had fewer years of education and were younger when their first child was born. Multivariate analyses indicate that mothers in the high-risk group had significantly worse scores on parenting attitude scales. Given the current state of welfare reform, it is important to determine which factors besides maternal substance abuse place these mothers at risk for poor parenting. PMID- 10718160 TI - Detecting bipolar disorder among treatment-seeking substance abusers. AB - Bipolar disorder is increasingly recognized to have frequent comorbidity with substance use disorders, but may be difficult to diagnose among patients with active substance use. The purpose of this paper is to describe a brief, self report form for the efficient detection of bipolar disorder. The 19-item form was piloted in 373 consecutive applicants for substance abuse treatment at an urban Veterans Affairs (VA) medical center. Results show reasonable internal consistency (alpha = .850) and high rates of manic symptomatology (36%), previous bipolar diagnosis (30%, 51% of whom report prior psychiatric hospitalization), and exposure to mood stabilizers (20%, 66% of whom reported therapeutic benefit). Comparison of nine different scoring algorithms with chart diagnosis as the validating criterion found that self-report of bipolar diagnosis was optimally sensitive. Either self-report of bipolar diagnosis with hospitalization or self report of exposure to mood stabilizers with therapeutic response was optimally specific. Symptom self-report items had significantly poorer sensitivity and specificity (F = 7.60, p < .01). We conclude that questions pertaining to diagnostic and treatment history (especially hospitalization or therapeutic medication response) are considerably superior to symptom-based screening for clinically diagnosed bipolar disorder. Further work using structured interview as the diagnostic criterion is under way to validate this instrument. PMID- 10718161 TI - Venlafaxine treatment of cocaine abusers with depressive disorders. AB - OBJECTIVE: There appears to be a link between depression and cocaine that is both complex and elusive. The purpose of this study was to examine the effect of venlafaxine, a broad spectrum antidepressant, in the treatment of 13 patients who were diagnosed with cocaine dependence and comorbid major depressive disorder (MDD). METHOD: The majority of the patients in the study were part of a larger double-blind trial using desipramine. This subgroup consisted of people who had failed to respond to desipramine or could not tolerate its side effects. Thirteen patients were enrolled, 10 men and 3 women. Of the patients, 11 completed the 12 week study. All of the patients had a Hamilton Depression (HAM-D) score greater than 14 at baseline, and each had used at least $20 worth of cocaine per week in the 4 weeks prior to entering the study. In addition, all of the patients received weekly relapse prevention therapy throughout the study. The median dose of venlafaxine was 150 mg/day. RESULTS: The 11 patients who completed the study had significant reductions in mood symptoms by the end of the study. The average total HAM-D score at baseline was 18.0 +/- 3.2; at Week 2, it was 1.9 +/- 0.94; and at the end of the study, it was 1.4 +/- 1.8. The majority of patients reported reductions of cocaine use short of abstinence. All subjects reported a greater than 75% reduction in cocaine use compared to baseline. There were no serious side effects. CONCLUSIONS: The results of this small study indicate that venlafaxine may be a safe, well-tolerated, rapidly acting, and effective treatment for patients with a dual diagnosis of depression and cocaine dependence. PMID- 10718162 TI - Motivation for treatment in a prison-based therapeutic community. AB - Current research concludes that participation in postprison aftercare is critical to the effectiveness of prison-based therapeutic community (TC) treatment. This conclusion makes it imperative to understand the client determinants of retention in prison treatment, particularly continuance in postprison aftercare. Currently, however, little data exist as to client predictors of seeking and remaining in prison-based TCs or entering postrelease aftercare. In the present study, significant relationships were obtained between initial motivation (i.e., Circumstances, Motivation, Readiness [CMR] scores), retention, aftercare, and outcomes in a sample of substance abusers treated in a prison-based TC program. Implications are discussed for theory, research, and treatment policy. PMID- 10718164 TI - Abortion and subsequent substance abuse. AB - A statistical association between a history of substance abuse and a history of abortion has been identified in several studies, but this association has not yet been thoroughly analyzed. This study draws on a subset of data from a reproductive history survey that included a nonparametric self-assessment of past substance abuse distributed to a random sample of American women. Analysis of this substance abuse variable showed that a report of substance abuse following a first pregnancy was associated significantly with (a) abortion for all women, (b) abortion for adolescents, and (c) abortion for women over 19 years of age. Women who aborted a first pregnancy were five times more likely to report subsequent substance abuse than women who carried to term, and they were four times more likely to report substance abuse compared to those who suffered a natural loss of their first pregnancy (i.e., due to miscarriage, ectopic pregnancy, or stillbirth). Women with a history of abortion or a history of substance abuse were significantly more likely to feel discomfort in responding to the survey. The findings of this study have important implications for the design of future studies examining substance abuse, adolescents, and women. These findings may also have clinical and counseling implications. PMID- 10718163 TI - Effects of lamotrigine on behavioral and cardiovascular responses to cocaine in human subjects. AB - We evaluated the effects of acute pretreatment with lamotrigine, a putative glutamate release inhibitor, on the physiological and behavioral responses to intranasal cocaine in cocaine-dependent volunteers (N = 8). The study employed a double-blind, placebo-controlled, within-subject design. Subjects participated in six experimental sessions. On each study day, placebo, lamotrigine 125 mg, or lamotrigine 250 mg was administered orally in the morning, followed 2 hours later by intranasal cocaine 120 mg/70 kg or placebo. Measurements of heart rate and blood pressure were acquired, and subjects responded to mood state questionnaires at predetermined time intervals. Cocaine alone produced increases in heart rate, blood pressure, and several measures of pleasurable mood and drug effects. Lamotrigine alone produced a mild relaxing effect. Lamotrigine pretreatment altered neither the physiological responses nor the subjective ratings of cocaine's pleasurable or aversive mood effects. PMID- 10718165 TI - Symptoms of dependence, multiple substance use, and labor market outcomes. AB - The prevalence and costs of alcohol and drug disorders pose a serious social concern for policymakers. In this paper, we use data from the National Household Surveys on Drug Abuse (NHSDA) to estimate simple descriptive statistics and analysis of variance (ANOVA) models of the relationship between symptoms of dependence and labor market outcomes for alcohol, cigarettes, marijuana, and other illicit drugs. For men, we find that substance use with symptoms of dependence is associated with both lower employment rates and fewer hours of work. For women, we find that substance use with symptoms of dependence is associated with lower employment rates, but we find no consistent evidence of a relationship between symptoms of dependence and the number of hours worked. Finally, all of our point estimates are smaller in magnitude when we control for multiple substance use, suggesting that comorbidities play a critical role in the relationship between substance use and labor market outcomes. Our results suggest that policymakers and researchers should consider the full spectrum of substance use and dependence rather than focusing on the simple use of a single substance. PMID- 10718166 TI - Racial identity and its assessment in a sample of African-American men in treatment for cocaine dependence. AB - Substance abuse treatment studies frequently include subjects from different ethnic and racial groups, but many investigations limit the examination of race and ethnicity to the use of nominal labels. This approach reveals little about the social or psychological significance of racial and ethnic group membership to the subjects of study or about the potential effects of these factors on substance-involved behaviors. In this study, a principal components analysis (PCA) with varimax rotation was conducted on the 50-item long form of the Racial Identity Attitude Scale (RIAS) (1) in a sample of 294 African-American men in treatment for cocaine dependence. The RIAS was developed to measure attitudes about race and racial status among blacks, but it has not been utilized widely in substance abuse research. Our findings provide evidence for the structural validity of this instrument in this sample of substance abusers. We discuss how recent advances in racial identity theory and its measurement may provide an important avenue for understanding the psychological consequences of racial group membership and for examining the potential effects of these factors on treatment response in studies of substance misuse. PMID- 10718167 TI - Adolescent risk taking and self-reported injuries associated with substance use. AB - OBJECTIVE: To examine the incidence of adolescent substance use at the time of injury and its relation to risk-taking behavior. METHOD: A total of 643 male and 782 female 9th through 12th grade students at three high schools anonymously completed surveys on any injuries that had occurred in the prior 6 months associated with substance use and risk-taking behavior. RESULTS: Males reported a higher incidence of injuries related to alcohol or other drugs than females (17.3% vs. 13%). The 17 year olds reported more injuries related to substance use than 14 or 15 year olds (20.2% vs. 14.4% and 15%, respectively). A logistic regression analysis revealed that the odds of a substance use-related injury increased approximately sixfold when adolescents reported engaging in risk-taking behavior. CONCLUSION: A significant portion of adolescents (approximately 15%) reported injuries associated with substance use. Adolescents who reported a history of risk-taking behaviors were much more likely to report substance use related injuries. PMID- 10718168 TI - Determinants of youth tobacco use in West Virginia: a comparison of smoking and smokeless tobacco use. AB - PURPOSE: To identify and compare the determinants of different types of tobacco use among rural youths and discuss the implication of these differences for youth tobacco use cessation. METHODS: Ninth grade participants (n = 883) were 95% white, between 13 and 19 years old with a mean age of 14.6 years. Students were classified into four exclusive groups: non-tobacco use, smoking only, smokeless tobacco (ST) use only, and conjoint smoking and ST use. The influences of 14 specific risk factors on tobacco use were investigated for each group using separate multiple logistic regression analyses. RESULTS: Among participants, 20% were smokers only, 6% were ST users only, and 10% were conjoint users. Students who had more friends (odds ratio [OR] =] 2.75) and siblings (OR = 1.96) who smoke, family problems (OR = 1.70), and favorable attitudes toward tobacco use (OR = 1.12) were more likely to smoke than were other students. Among students who used only ST, gender was a primary determinant (95% were male). Excluding gender, sibling ST use (OR = 4.28), friends' ST use (OR = 1.71), and favorable attitudes (OR = 1.11) were the most significant risk factors. Male students were also more likely to use both cigarettes and ST (OR = 8.62). In addition, among students who used both tobacco products, siblings' and friends' ST use were significant (OR = 3.09 and 2.13, respectively), as well as family problems (OR = 2.41) and attitude (OR = 1.15). Unlike smokers only or ST users only, lack of knowledge about tobacco was a significant determinant among conjoint users (OR = 1.39). CONCLUSIONS: Findings revealed that 7 of 14 factors were significant predictors of tobacco use. Some factors predicted smoking only, ST only, and conjoint use; however, the pattern of predictors varied for these three categories. Implications for these findings as they relate to tobacco use interventions are discussed. PMID- 10718169 TI - Deconstructing contexts of binge drinking among college students. AB - This paper examines the contextual characteristics common to binge drinking occasions reported by college students. In addition, the study examines the influence of such contextual characteristics on alcohol-related problems experienced by students. Using random sampling and telephone interview methodology, 401 college students attending a large southern California university were surveyed by trained research staff. The interview protocol was based on the Core Survey and included context of use questions from the College Risk Assessment Guide. Results of stepwise multiple logistic regressions indicate that drinking with friends and events with food available protect against alcohol problems, while drinking events in which illicit drugs are available present higher risk for problems. Implications for future research and prevention are discussed. PMID- 10718170 TI - Divalproex in the treatment of alcohol withdrawal. AB - The present study represents an open-label clinical trial comparing treatment with a benzodiazepine (lorazepam) to divalproex in 11 inpatients with uncomplicated alcohol withdrawal syndrome. The trial used the Clinical Institute Withdrawal Assessment for Alcohol-Revised (CIWA-Ar) scale. There were no significant differences in demographics or substance use parameters between the divalproex group (n = 6) or the lorazepam group (n = 5). A significant Group x CIWA-Ar score interaction [F(8,72) = 2.57, p < or = .01] was confirmed and further substantiated by a quadratic trend component for the interaction [F(1,9) = 24.9, p < or = .001]. This preliminary study supports further investigation of divalproex in the treatment of alcohol withdrawal. PMID- 10718171 TI - Reflections and proposals for the standardization of lymphadenectomy for gastric carcinoma. AB - In this paper, I consider: the value of various histological procedures; the feasibility and reproducibility of lymphadenectomies according to the Japanese Research Society for Gastric Cancer; the minimal number of lymph nodes (LNs) which should be resected for each type of lymphadenectomy; and the best way to present the results concerning the LN status. A reproducible lymphadenectomy is proposed for simplified anatomical level I (anterior plane, along the gastric curves) and level II (intermediate plane, along the gastric arteries), without spleno-pancreatectomy for the majority of cases. The LN status must be based on the total number of involved nodes, as recommended by the 1997 UICC classification. These simple, reproducible guidelines offer a basis for the standardization of procedures used for the treatment and classification of gastric carcinomas. PMID- 10718172 TI - Variations in prognostic factors in primary breast cancer throughout the menstrual cycle. AB - AIMS: We investigated whether menstrual cycle dependent variations in prognostic factors are detectable in malignant breast tissue. METHODS: Since 1977 the Danish Breast Cancer Cooperative Group has collected population-based information about primary clinical data, treatment regimens and follow-up status on Danish women with breast cancer. Information about last menstrual periods prior to surgery was obtained from files recorded at the time of admission for primary surgery. Included in this study were 1060 patients self-reported to be regularly menstruating and with a menstrual period within 6 weeks of surgery and who were operated in a single-step procedure. None of the patients were current users of exogenous hormones at the time of surgery. Variations of prognostic factors throughout the menstrual cycle were evaluated. RESULTS: Overall, no significant correlation between endogenous hormone fluctuations and oestrogen receptor (ER) status and progesterone receptor (PgR) status were found. Furthermore, we observed no cycle-dependent variation for mitotic index, lymph node involvement or tumour size. CONCLUSIONS: The classical prognostic factors in breast cancer did not differ significantly throughout the menstrual cycle in the present study. PMID- 10718173 TI - The psychological impact of immediate rather than delayed breast reconstruction. AB - AIMS: A retrospective analysis of the psychological advantages of immediate reconstruction (IR) against delayed reconstruction (DR). METHODS: A total of 121 patients who underwent different types of breast reconstruction were seen in the follow-up clinic and assessed for: anxiety, depression, body image, self-esteem, sexuality and satisfaction. RESULTS: Ninety-five percent of the patients who had IR preferred this technique and 76% of the DR group would have preferred IR. Anxiety and depression were decreased and body image, self-esteem and sexual feeling of attractiveness and satisfaction were significantly superior in the IR group compared with that of the DR group. CONCLUSION: Patients who had immediate reconstruction recalled less distress and had better psychosocial well being than those who had delayed reconstruction. PMID- 10718174 TI - Quality of life in patients with early and advanced carcinoma of the breast. AB - AIM: Emotional disturbances are known to occur in patients suffering from malignant diseases even after treatment. This is mainly because of a fear of death which modifies quality of life (QOL). QOL has gained an important place in the management of cancer in industrialized nations, with the increase in survival. However, in developing countries like India, very little attention has been paid to this issue. Developing countries have poor infrastructure and lack proper treatment facilities at most centres, this leads to poor survival rates and hence much emphasis is on attaining quantity of life rather than quality. This study was carried out to assess the quality of life determinant in patients with breast cancer and the impact of treatment on quality of life indices. METHODS: We carried out QOL assessment in 50 patients with breast cancer using the modified linear analogue scale for self assessment (LASA). RESULTS: Significant deterioration was seen in health-related parameters in terms of recreation (P=0.01), social life (P=0.002), mobility (P=0.03), physical activity (P=0.4) and sleep and appetite (P=0.05). Treatment related parameters deteriorated in both early and advanced carcinoma. Similarly, weight loss was seen in both the groups, however, this was not statistically significant. Self care and recreation were found to be the most important parameters influencing the QOL in breast cancer patients. CONCLUSIONS: Breast cancer detection programs, health education and better awareness among women in industrialized nations has helped in downstaging of the disease, thus improving overall survival. It has not been so in developing countries, where the majority of patients present with advanced disease (T3 and T4). These are usually managed with surgery and adjuvant chemotherapy, which duly interferes with general health-related parameters and the social life of these patients, thereby adversely affecting the QOL. PMID- 10718175 TI - Integrin beta1 upregulation in MCF-7 breast cancer cells by angiotensin II. AB - AIMS: Integrins are a major family of cell adhesion molecules whose function is perturbed in tumour invasion and metastasis. Angiotensin II (A II) is well-known in the systemic control of water and electrolyte homeostasis and haemodynamics, but recent evidence points to an additional local renin-angiotensin system (RAS) with possible long-term trophic effects including carcinogenesis. METHODS: The effect of angiotensin II on MCF-7 human breast cancer cell line integrin expression was evaluated with immunocytochemistry (ICC) and immunoprecipitation (IP). RESULTS: The experiments demonstrated a 1.40 +/- 0.14-fold increase in beta, integrin expression on MCF-7 cells following treatment with A II. CONCLUSIONS: These findings report the first evidence of an association between integrins and the RAS in human breast cancer cells and suggest a novel research avenue for future anti-metastatic strategies, through the manipulation of cell adhesion mechanics, in the management of invasive human breast cancer. PMID- 10718176 TI - Telomerase activity and lymphovascular invasion in breast cancer. AB - BACKGROUND: Telomerase is a ribonucleoprotein enzyme that plays an important role in cell immortalization and carcinogenesis. Lymphovascular invasion (LVI) is a fundamental step in the process of breast cancer metastasis and is recognized as an important prognostic factor in patients with breast cancer. METHODS: Using a PCR-based assay, telomerase activity was determined in 34 prospectively collected infiltrating breast carcinomas. Adjacent sections of the specimens were examined histologically by two experienced breast pathologists using light microscopy and haematoxylin & eosin staining. RESULTS: Telomerase activity was detected in 24 (71%) of 34 breast tumours. Two (20%) of 10 telomerase-negative tumours had LVI compared with 14 (58.3%) of 24 telomerase-positive tumours. This association was statistically significant (P<0.05). Telomerase activity was also significantly associated with nodal metastases but not with tumour grade, tumour size or menopausal status. CONCLUSIONS: Telomerase reactivation is significantly associated with LVI in breast cancer and may reflect the metastatic potential of the disease. PMID- 10718177 TI - The analysis of prognostic factors in stage III-B non-inflammatory breast cancer. AB - AIMS: To evaluate factors predicting disease recurrence in patients treated for stage III-B breast cancer by neoadjuvant chemotherapy followed by surgery. METHODS: A retrospective study of 52 patients who responded to neoadjuvant chemotherapy followed by modified radical mastectomy was carried out. The parameters studied included pre-treatment tumour size, clinical axillary status, grade, lymphatic-vascular invasion, pathological axillary status, number of metastatic lymph nodes, menopausal status and oestrogen receptor status. RESULTS: In the univariate analysis, number of metastatic lymph nodes, primary tumour size, pathological axillary status and histological grade were statistically significant factors associated with recurrence of disease. Multivariate analysis demonstrated that the number of metastatic lymph nodes (relative hazard 6.1) and primary tumour size (related hazard 2.5) were the most important independent prognostic factors for recurrence. CONCLUSIONS: These results indicate that the number of involved lymph nodes after neoadjuvant chemotherapy, independent of the clinical response of primary tumour, had a most significant impact on disease free survival. Additionally primary tumour size had a marked prognostic significance in spite of clinical changes in tumours following chemotherapy. PMID- 10718178 TI - Expression of p53 and p21waf1/cip1 in gastric carcinoma: lack of inter relationship or correlation with prognosis. AB - AIMS: The cell cycle regulators p53 and p21waf1/cip1 are expressed variably in human cancers. We investigated their expression in gastric carcinoma and determined their inter-relationship and prognostic significance. METHODS: Immunohistochemistry was used to determine their expression in material from 100 resected specimens of gastric carcinoma, and comparison was then made of the degree of expression between each, with conventional clinicopathological indices and with survival. RESULTS: Positivity was found with p53 (40%) and p21 (75%). There was no significant correlation between the expression of each individual marker, nor between each marker and 5-year survival. There appeared to be an association between p53 expression and lymph node metastases, and a higher frequency of p21waf1/cip1 expression in males. CONCLUSIONS: The expression of p53 and p21waf1/cip1 as detected by immunohistochemistry were of no value in predicting the prognosis of patients with gastric carcinoma. PMID- 10718179 TI - Urinary tissue factor levels in patients with bladder and prostate cancer. AB - AIMS: Coagulation activation is a recognized complication of cancer in which increased tissue factor (TF) is implicated. TF can be detected in urine (uTF). This study assesses uTF levels in benign and malignant urological disease and correlates the results with conventional markers of tumour progression. METHODS: Using a simple and reproducible kinetic chromogenic assay, we determined uTF levels in controls (normal volunteers (n = 57) and patients with renal stones (n = 30)), benign and malignant bladder (n = 75) or prostate (n = 106) disease and in patients with or without recurrent bladder cancer (n=30). Each benign disease group was stratified as inflammatory (cystitis or prostatitis) or non inflammatory (negative cystoscopy following haematuria or benign prostatic hypertrophy). RESULTS: The controls and the benign non-inflammatory results were indistinguishable. The malignant and inflammatory groups showed raised uTF levels over controls (P<0.001 bladder and P<0.01 prostate). The difference between malignant and benign inflammatory disease was only significant for the bladder group. uTF levels were significantly related to histological tumour grading, prostate serum specific antigen, static bone scan images and recurrence status. CONCLUSIONS: uTF levels can distinguish, statistically but not without overlap, patients with malignancy from normal controls and benign non-inflammatory conditions. Discrimination between inflammatory and malignant disease has only been demonstrated in the bladder. uTF levels showed a significant association with markers of tumour progression or metastasis and may be useful in predicting bladder tumour recurrence. PMID- 10718180 TI - Sarcoma metastases due to iatrogenic implantation. AB - AIM: To demonstrate the ability of extremity soft tissue sarcomas (STSs) to implant into tissues exposed during surgery. METHODS: We use two cases to illustrate that wounds created during surgery for STSs, including donor sites for skin grafts, pedicled and free flaps and blood vessels used in reconstruction, should be regarded as potential sites of quasi-local recurrence. CONCLUSIONS: This report reinforces the need for meticulous surgical technique and planning of procedures to avoid contamination of clean areas that might not be included in adjuvant radiotherapy fields. The cases also highlight the pivotal importance of the correct initial management of these uncommon tumours. PMID- 10718181 TI - Osteosarcoma of the pelvis--oncological results of 40 patients registered by The Netherlands Committee on Bone Tumours. AB - AIM AND METHODS: We reviewed the oncological outcome in 40 consecutive patients with an osteosarcoma of the pelvic region, registered in the files of the Netherlands Committee on Bone Tumours (NCBT) between 1978 and 1995. RESULTS: Six patients had distant metastases at initial presentation (Enneking stage IIIB), 33 patients had stage IIB osteosarcoma and one patient stage IB osteosarcoma. Patients with metastases were treated with chemotherapy (four) or palliative procedures (two). Patients with non-metastatic osteosarcoma were treated with surgical procedures with (14) or without (four) neoadjuvant chemotherapy, chemotherapy without surgical resection (nine), or palliative procedures (seven). The median survival of stage IIB and IIIB osteosarcoma was 14 months (2-175) and 7.5 months (2-16), respectively. Survival in patients with stage IIB osteosarcoma treated with curative procedures was significantly better (P<0.0006) compared with stage IIB patients treated with palliative intent. Two and 5-year survival for patients with curatively treated stage IIB osteosarcoma was 35% and 26%, respectively; distant metastases had developed in 65% of these patients. On univariate analysis, positive prognostic factors for patients with stage IIB osteosarcoma were complaints of 3 months or less before initial presentation, tumour size of 8 cm or less, osteoblastic subtype, surgical resection of the primary tumour and limb salvage procedures. CONCLUSION: In conclusion, the prognosis of pelvic osteosarcoma remained poor despite modern multimodality treatment regimens, including neoadjuvant chemotherapy. PMID- 10718182 TI - 16q heterozygosity loss in Wilms' tumour in children and its clinical importance. AB - INTRODUCTION: The loss of heterozygosity (LOH) of 16q is a structural change detected in about 20-30% of Wilms' tumour cases. Aberrations which result in deletion of 16q are also found in breast cancer, prostate cancer and liver cancer, where they are connected with a worse prognosis. The hypothesis of a bad prognosis in nephroblastomas with LOH 16q was first formulated by scientists from NWTS (National Wilms Tumor Study) on the basis of 232 cases of Wilms' tumour. However, SIOP studies (International Society of Paediatric Oncology) which included 28 cases of Wilms' tumour, did not show any clinico-pathological correlations with LOH 16q. Therefore, we aimed to evaluate the importance of LOH 16q in relation to clinico-pathological factors in a group of children, treated according to the SIOP criteria. AIMS: The aim of this work was to evaluate the frequency of LOH 16q in sporadic unilateral Wilms' tumour and to study the relationship between LOH 16q and selected patho-clinical parameters. The study comprised 66 children (31 girls and 35 boys) aged from 2 days to 13 years. METHODS: LOH 16q was studied by the examination of polymorphism of marker sequences in the region 16q24. DNA was isolated from paraffin sections of tissue for routine microscopic examination by the microdissection method. The method of study involved the amplification of polymorphic sequences from the 16q24 region by polymerase chain reaction (PCR) and separation of the products of amplification by polyacrylamide gel electrophoresis. The results were the subject of statistical analysis in relation to gender, age of child at first diagnosis, stage of clinical advancement and histological type of tumour. The connection between LOH 16q and recurrences, metastases and death, and failure free survival and absolute survival of children followed-up for over 24 months after nephrectomy were studied. RESULTS: The study revealed a lack of correlation between LOH 16q and gender, however LOH 16q was more frequent in children with Wilms' tumour aged >24 months, P<0.05. Also, LOH 16q was more frequent in tumours classified as clinical stage (CS) II or III than in CS I, P<0.05, but there were no differences in the occurrence of LOH 16q in tumours classified as CS II and CS III. We have found no correlation between LOH 16q and the histological type of tumour. However, LOH 16q has been found three times as frequently in tumours from children who died than in tumours of children who survived, P<0.0024. PMID- 10718183 TI - Multimodality treatment for patients with hepatocellular carcinoma: a single institution retrospective series. AB - BACKGROUND: The main therapeutic options for hepatocellular carcinoma (HCC) are hepatic resection, transcatheter arterial embolization (TAE), percutaneous ethanol injection therapy (PEIT) and regional chemotherapy (RC). METHODS: This study retrospectively examined the results of primary treatment of 600 patients with hepatocellular carcinoma selected according to the treatment guidelines of our facility and the results of various combination therapies for recurrent cases. The selection criteria of therapeutic options included the number and size of tumours and hepatic function. RESULTS: The selected primary treatment was hepatic resection for 53.7% of the cases, TAE for 31.5%, PEIT for 8.2% and RC for 6.6%,. The treatment for post-resection recurrence was TAE alone for 62.4% of the cases, TAE + RC for 4.0%, PEIT for 15.2%, TAE + PEIT alone for 4.8%, RC for 8.0% and hepatic resection for 5.6%. The treatment for post-TAE recurrence was TAE alone for 83% of the cases, TAE + PEIT for 9%, TAE + RC for 3%, RC alone for 3% and PEIT alone for 2%. For post-PEIT, therapy was PEIT alone for 71.4% of the cases and PEIT + TAE for 28.6%. For post-RC, RC alone was used for 92.5% and RC + PEIT for 7.5%. The cumulative 3 and 5-year survival rates were 84.4% and 70.6%, respectively for stage I; 61.5% and 48.6% for stage II; 52.7% and 20.5% for stage III; and 22.8% and 17.1% for stage IVA. The cumulative 5 and 7-year survival rates after the primary treatments were 52.% and 40.1%, respectively, for hepatic resection; 46.5% and 38.7%, for TAE; 49.6% and 33.1% for PEIT; and 16.7% and 8.3% for RC. CONCLUSIONS: To improve the treatment results for HCC, early detection is essential and various modalities of treatments in combination should be used for recurrence after primary treatment. PMID- 10718185 TI - A technique for dealing with the short gastric vessels during gastric surgery. PMID- 10718184 TI - Lattice intrahepatic doxorubicin with and without in-flow occlusion: a pharmacokinetic study of direct liver injection. AB - AIM: To assess possible improvements in drug delivery to unresectable liver tumours we investigated the pharmacokinetics of intrahepatic doxorubicin in the dog liver with and without inflow occlusion. METHODS: Using a lattice template, doxorubicin was injected into 16 sites (over a 9 cm2 area) in each of three lobes of the liver for a total of 48 sites. The total doses of doxorubicin used were 4.8 mg and 96 mg (0.1 and 2 mg/site). Dogs with intravenous and intra-arterial delivery of the same total doses of doxorubicin were used as controls. Experiments using intrahepatic injection were performed with and without a 30 min occlusion of the hepatic artery, common bile duct and portal vein (inflow occlusion). The studies with vascular stasis were performed to determine if drug clearance from the injection sites and their plasma levels were reduced. Also, it was observed that blood and drug loss along the needle tract was reduced when inflow occlusion was used. Plasma and liver samples were harvested over a 90-min period and analysed by high pressure liquid chromatography (HPLC). RESULTS: In plasma mean peak levels and mean area under the curve (AUC) were significantly lower with intrahepatic doxorubicin (P<0.05) than with intravenous or intra atrial delivery. In the liver AUCintrahepatic/AUCintravenous ratios were 3.45 and 3.6 with 4.8 mg and 96 mg of doxorubicin respectively. The AUCintrahepatic/AUCintraarterial ratios were 1.97 and 1.65 respectively. The liver extraction ratio (AUCliver/AUCplasma) after intravenous administration with 4.8 mg and 96 mg of doxorubicin was 31.9 with both doses of doxorubicin. The corresponding extraction ratios were 107.6 and 51 for intra-arterial administration and 425.2 and 237.7 for intrahepatic administration for the two doses of doxorubicin. Intrahepatic injection with inflow occlusion minimized the leakage back along the needle-tracts. The liver visibly retained the injected drug more completely. However, there was a decrease in systemic clearance resulting in a reduction of the pharmacological advantage. CONCLUSION: These results indicate that lattice-intrahepatic administration of doxorubicin into the liver using a lattice template was associated with a significant increase in local and decrease in systemic exposure as compared to intravenous or intra arterial administration of the same doses. Simultaneous occlusion of arterial and portal venous inflow was not shown to improve regional drug delivery. These pharmacokinetic studies may constitute a basis for palliative treatment of liver tumours by lattice intralesional injection when these cancers are found to be unresectable at the time of exploratory surgery. PMID- 10718186 TI - Primary intraosseous carcinoma of the mandible--a case report and review of the literature. AB - BACKGROUND: Squamous cell carcinoma arising within bone is a rare lesion and is only seen essentially in the jaw bones. METHODS: A case of primary intraosseous carcinoma arising in the mandible is reported in a 60-year-old female patient. Twenty-eight cases of primary intraosseous carcinoma published in the literature, till date, are reviewed. RESULTS: The mean age of the patients at the time of diagnosis was 53 years and the male: female ratio was 2.2:1. The posterior mandible was the predominant site. Fourteen of 28 patients presented with routine dental disorders, while eight patients complained of swelling, four of severe pain and three had sensory disturbances. The incidence of lymphadenopathy was 10 (34.481%) out of the 29 cases reviewed here. Wide surgical excision is accepted as the treatment of choice. CONCLUSION: The most common presenting symptom of these tumours is swelling and persistent pain in the jaw. Hence, in making a diagnosis one is likely to consider benign dental conditions. The importance of considering intraosseous carcinoma as a possibility in all cases of persistent pain and swelling in the jaw is emphasized so that suitable treatment can be instituted early. PMID- 10718187 TI - Modelling in tumour biology part 1: modelling concepts and structures. AB - Our strategies for the treatment of cancer are constrained by our incomplete understanding of tumour biology and behaviour, and by the enormous complexity and resilience to therapeutic perturbation found in the biological world. We are obliged to simplify this complexity through the use of models and mechanistic explanations. In the first of these papers, we consider the nature of modelling mechanisms available to clinical researchers and the extent to which we rely upon them in our understanding of the nature and behaviour of tumours. In the second part, we will consider specifically how models help us to develop more effective strategies for cancer therapy. PMID- 10718188 TI - Port-site metastasis following laparoscopic cholecystectomy: a review of the literature and a case report. AB - Port-site metastasis following laparoscopic cholecystectomy with unsuspected gallbladder carcinoma is a serious problem. We reviewed 45 such cases reported in the English literature to date, and add another case which occurred in a 72-year old female 13 months after a laparoscopic cholecystectomy for gallstones, who also had an unapparent gallbladder carcinoma. Pre-operative diagnosis of gallbladder carcinoma is possible in less than 10% of cases, with a high index of suspicion. If detected during laparoscopy early conversion to open procedure is recommended. If diagnosed later, however, to contemplate further radical operation depending on histopathology would be unwise as a universal approach, because of increased associated morbidity and mortality. The prospect of cure is also very unrealistic in this condition. PMID- 10718189 TI - Disappearing microcalcification after neoadjuvant chemotherapy--a case report. PMID- 10718190 TI - Stromal sarcoma of the prostate. AB - Sarcomas of the prostate are very rare. This article describes the radiological and histopathological findings of a case of prostatic stromal sarcoma, with the appearance of a phyllodes tumour with adenoid basal cell hyperplasia. Management and follow-up of this tumour are discussed. PMID- 10718191 TI - Solitary drain-site metastasis from Hurthle-cell carcinoma of the thyroid. PMID- 10718192 TI - Neural science: a century of progress and the mysteries that remain. PMID- 10718193 TI - Tnat1 and Tnat2 from Arabidopsis thaliana: novel transposable elements with tandem repeat sequences. AB - A computer-aided homology search of databases found that the nucleotide sequences flanking ATLN44, a non-LTR retrotransposon (LINE) from Arabidopsis thaliana, are repeated in the A. thaliana genome. These sequences are homologous to flanking sequences of 664 bp with terminal inverted repeat sequences of about 70 bp. The 664-bp sequence and most of the 14 homologues identified were flanked by direct repeat sequences of 9 bp. These findings indicate that the repeated sequence, named Tnat1, is a transposable element that duplicates a 9-bp sequence at the target site on transposition and that ATLN44 is inserted in one Tnat1 member. Interestingly, all of the Tnat1 members had tandem repeats comprised of several units of a 60-bp sequence, the number of repeats differing among Tnat1 members. Of the Tnat1 members identified, one was inserted into another sequence repeated in the A. thaliana genome: that sequence is about 770 bp long and has terminal inverted repeat sequences of about 110 bp. The sequence is flanked by direct repeats of a 9-bp sequence, indicating that it is another transposable element, named Tnat2, from A. thaliana. Moreover, Tnat2 members had a tandem repeat about 240 bp long. Tnat1 and Tnat2 with tandem repeats in their internal regions show no homology to each other or to any of the elements identified previously; therefore they appear to be novel transposable elements. PMID- 10718194 TI - Tagged mutagenesis and gene-trap in the moss, Physcomitrella patens by shuttle mutagenesis. AB - The moss, Physcomitrella patens has been used as a useful material in many fields, because of its simple body plan, ease of gene targeting, and other reasons. Although many mutants have been reported, no method to isolate the corresponding genes was reported. We developed a gene tagging and gene-trap system in P. patens by using the shuttle mutagenesis technique, which has been used in the budding yeast. In 5264 tagged lines, 203 mutants with altered developmental or morphological phenotypes were obtained. In 129 of 4757 gene-trap lines, beta-glucuronidase (GUS) activity was detected in some tissue. Although multiple copies of a tag were detected in many tagged lines by Southern analyses, most copies are likely integrated at the same locus according to PCR analyses. PMID- 10718195 TI - Random PCR-based genome sequencing: a non-divide-and-conquer strategy. AB - We propose a genome sequencing strategy, which is neither divide-and-conquer (clone by clone) nor the shotgun approach. Random PCR-based and PCR relay sequencing constitute the basis of this novel strategy. Most of the genome is sequenced by the former process that requires only a set of non-specific primers and a template DNA. Random PCR-based sequencing reduces redundancy in sequencing by exploiting known sequence information. The number of primers required for random PCR was significantly diminished by using a combination of primers. The former process can be partially replaced by the shotgun method, if necessary. The gap-filling process can be effectively performed by way of PCR relay. The feasibility of this strategy was demonstrated using the Escherichia coli genome. This strategy enhances the global effort towards genome sequencing by being available through the Internet and by allowing the use of preexisting sequence data. PMID- 10718196 TI - mRNAs encoding zinc finger protein isoforms are expressed by alternative splicing of an in-frame intron in fission yeast. AB - We report here that a gene encoding a protein with three zinc fingers is expressed predominantly to produce a protein containing only two zinc fingers in the fission yeast Schizosaccharomyces pombe. A third zinc finger resides within the in-frame intron that is normally spliced out. By RT-PCR analysis, we detected a minor transcript encoding a protein with three zinc fingers. Such alternative splicing for assortment of zinc finger domains have been reported in animals and implicated in switching of the target genes expressed specifically during development. This is the first report of the occurrence of such zinc finger assortment in lower eucaryotes. PMID- 10718197 TI - Structural analysis of Arabidopsis thaliana chromosome 5. X. Sequence features of the regions of 3,076,755 bp covered by sixty P1 and TAC clones. AB - In our ongoing project to deduce the nucleotide sequence of Arabidopsis thaliana chromosome 5, non-redundant P1 and TAC clones have been sequenced on the basis of the fine physical map, and as of January, 2000, the sequences of 16.6 Mb representing approximately 60% of chromosome 5 have been accumulated and released at our web site. Along with the sequence determination, structural features of the sequenced regions have been analyzed by applying a variety of computer programs, and we already predicted a total of 2697 potential protein coding genes in the 11,166,130 bp regions, which are covered by 159 P1 and TAC clones. In this paper, we describe the structural features of the 3,076,755 bp regions covered by newly analyzed 60 P1 and TAC clones. A total of 715 potential protein coding genes were identified, giving an average density of the genes identified of 1 gene per 4001 bp. Introns were observed in 80% of the genes, and the average number per gene and the average length of the introns were 4.5 and 147 bp, respectively. These sequence features are nearly identical to those in our latest report in which the data were compiled based on a new standard of gene assignment including the computer-predicted hypothetical genes. The regions also contained 12 tRNA genes when searched by similarity to reported tRNA genes and the tRNA scan-SE program. The sequence data and information on the potential genes are available through the World Wide Web database KAOS (Kazusa Arabidopsis data Opening Site) at http://www.kazusa.or.jp/kaos/. PMID- 10718198 TI - Prediction of the coding sequences of unidentified human genes. XVI. The complete sequences of 150 new cDNA clones from brain which code for large proteins in vitro. AB - We have carried out a human cDNA sequencing project to accumulate information regarding the coding sequences of unidentified human genes. As an extension of the preceding reports, we herein present the entire sequences of 150 cDNA clones of unknown human genes, named KIAA1294 to KIAA1443, from two sets of size fractionated human adult and fetal brain cDNA libraries. The average sizes of the inserts and corresponding open reading frames of cDNA clones analyzed here reached 4.8 kb and 2.7 kb (910 amino acid residues), respectively. From sequence similarities and protein motifs, 73 predicted gene products were functionally annotated and 97% of them were classified into the following four functional categories: cell signaling/communication, nucleic acid management, cell structure/motility and protein management. Additionally, the chromosomal loci of the genes were assigned by using human-rodent hybrid panels for those genes whose mapping data were not available in the public databases. The expression profiles of the genes were also studied in 10 human tissues, 8 brain regions, spinal cord, fetal brain and fetal liver by reverse transcription-coupled polymerase chain reaction, products of which were quantified by enzyme-linked immunosorbent assay. PMID- 10718199 TI - Relationship between functional and neuropsychological performance in early Alzheimer disease. AB - Twenty-four subjects with Alzheimer disease underwent cognitive and functional assessment. Functional assessment by caregivers consisted of a 25-item bipolar analog scale measuring activities of daily living and social behaviors divided into four functional domains: memory, attention/executive abilities, everyday skills, and self-care. Cognitive assessment consisted of standardized neuropsychological tests designed to evaluate five cognitive domains: episodic memory, attention/executive function, semantic memory, visuospatial function, and auditory-verbal short-term (working) memory. Functional assessment correlated well with overall severity as measured by Mini Mental State Examination (r = 0.733). Analysis of individual cognitive and functional domains revealed no significant correlation between episodic memory and functional performance. By contrast, functional ability correlated strongly with the cognitive domains of visuospatial function and semantic memory, being significantly greater than the correlation of functional ability with any other cognitive domain. These results were supported by multiple regression analyses that showed visuospatial function to be the sole cognitive predictor of functional abilities. These findings have implications for the evaluation of drug therapies in Alzheimer disease, in particular the effect of current cholinergic therapies on activities of daily living. PMID- 10718200 TI - Premorbid personality predicts level of rated personality change in patients with Alzheimer disease. AB - Multiple studies of individuals with Alzheimer disease have substantiated significant levels of informant-rated change in several domains and facets of the Neuroticism-Extraversion-Openness Personality Inventory, including increases in Neuroticism and decreases in Extraversion and Conscientiousness relative to premorbid personality traits. Decline in Openness was cited in some reports, and replicable changes were identified in several facets. Current and premorbid personality of 50 patients with Alzheimer disease were rated by informants using the Neuroticism-Extraversion-Openness Personality Inventory. Multiple regression analysis was used to assess possible relationships of levels of reported change with covariates, including premorbid rating, education, duration of dementia, age, gender, and Mini-Mental State Examination score. Premorbid rating was the only significant predictor of reported change for Neuroticism, Extraversion, Conscientiousness, and the facets Anxiety (N1), Assertiveness (E3), and Activity (E4). Rated change in Depression was also found to be related to duration of dementia, change in Vulnerability was influenced by gender, and reported change in both Openness and Ideas showed a relationship to level of education. PMID- 10718201 TI - Research involving persons with cognitive impairments: results of a survey of Alzheimer disease research centers in the United States. AB - Research involving persons with cognitive impairments presents ethical and practical challenges, including how to obtain valid informed consent. We asked the directors of the 29 U.S. research centers funded by the National Institute on Aging as "Alzheimer Disease Centers" to provide us with policies or guidelines used in their centers or associated institutions with regard to research involving cognitively impaired subjects. Twenty-four of the 29 centers (83%) responded. Five institutions (21%) had authored their own institutional policies, seven (29%) used guidelines issued by the Department of Health and Human Service's Office for Protection from Research Risks, and 12 (50%) had no policy or guidelines. The five institutional policies addressed a variety of issues, including obtaining consent from cognitively impaired subjects or their authorized representatives, subjects' assent to research participation, and guidance concerning determination of subjects' intellectual capacity. A well written policy on the protection of cognitively impaired research subjects is one way a research institution demonstrates that it gives serious attention to the rights and welfare of these vulnerable persons. We recommend that all institutions conducting such research author written policies articulating appropriate safeguards for these vulnerable subjects. To promote the protection of cognitively impaired subjects, federal agencies and other funding groups may want to consider requiring written institutional policies as one condition of receiving funds to conduct such research. PMID- 10718202 TI - Professional environmental assessment procedure for special care units for elders with dementing illness and its relationship to the therapeutic environment screening schedule. AB - The Professional Environmental Assessment Procedure (PEAP) was developed as a global quality-assessment measure for use by trained professionals in special care units for older people in dementia units of nursing homes. The PEAP consists of nine ratings whose relationship to another assessment device, the Therapeutic Environment Screening Schedule (TESS), is reported. Although designed to be multidimensional, the PEAP as tested in 43 special care units seems to reflect primarily a single evaluative dimension. It correlates highly with the TESS and may be used either separately or in combination with the TESS. PMID- 10718203 TI - Randomized, double-blind, placebo-controlled, multicenter study to evaluate the safety and tolerability of metrifonate in patients with probable Alzheimer disease. The Metrifonate Study Group. AB - A randomized, double-blind, placebo-controlled, parallel-group study was undertaken to evaluate the safety and tolerability of a once-daily oral administration of metrifonate in patients with probable mild to moderate Alzheimer disease. Metrifonate was given as a loading dose of 125-225 mg based on weight (2.5 mg/kg) for 2 weeks, followed by a maintenance dose of 50-90 mg based on weight (1.0 mg/kg) for 4 weeks. Twenty-nine patients received metrifonate, and 10 patients received placebo. Metrifonate produced a mean erythrocyte acetylcholinesterase inhibition at the end of treatment of 86.3%. The proportion of patients who experienced at least one adverse event was comparable between the metrifonate (76%) and placebo (80%) groups. Selected adverse events in disfavor of metrifonate (defined as those for which the incidence in the metrifonate and placebo groups differed by at least 10%) were diarrhea, nausea, leg cramps, and accidental injury. Adverse events were predominantly mild in intensity and transient. No severe adverse events were experienced by any patient. The most notable hemodynamic change observed during metrifonate treatment was a clinically insignificant mean decrease in the heart rate (by electrocardiogram) of approximately 9 beats/min, compared with an approximate 3-beats/min decrease for the placebo group. No muscle weakness was observed in this study. No clinically relevant laboratory abnormalities, such as liver toxicity, or changes in exercise tolerance or pulmonary function tests were found with metrifonate treatment. This metrifonate dose provided a high level of acetylcholinesterase inhibition, which was associated in these patients with a favorable safety and tolerability profile. Indeed, the magnitude of the peripheral acetylcholinesterase inhibition is the highest tolerable inhibition level yet observed. PMID- 10718204 TI - Cerebral blood flow in corticobasal degeneration and progressive supranuclear palsy. AB - To compare brain perfusion between corticobasal degeneration (CBD) and progressive supranuclear palsy (PSP), we investigated regional cerebral blood flow (rCBF) semiquantitatively with single-photon emission computed tomography and [123I]iodoamphetamine in six patients with CBD and five with PSP. Compared with 12 age-matched control subjects, the average of the left and right rCBF values for the CBD patients was significantly reduced in the inferior prefrontal, anterior cingulate, medial premotor, sensorimotor, posterior parietal, and superior temporal cortices as well as in the basal ganglia and thalamus, whereas only the medial premotor cortex was significantly hypoperfused in the PSP patients. Compared with the PSP patients, the CBD patients showed significantly decreased rCBF in the inferior prefrontal, sensorimotor, and posterior parietal cortices, but not in the subcortical regions. Compared with the controls, interhemispheric differences of rCBF were significant in the inferior prefrontal, sensorimotor, and posterior parietal cortices of the CBD patients but in only the medial prefrontal cortex of the PSP patients. These results indicate that rCBF reductions are more extensive and asymmetric in CBD than in PSP, although the two diseases share medial frontal involvement. PMID- 10718205 TI - Densitometric analysis of Galphao protein subunit levels from postmortem Alzheimer disease hippocampal and prefrontal cortical membranes. AB - An immunoblotting method using prefrontal cortical and hippocampal membranes from control and Alzheimer disease postmortem brains was employed to detect three subtypes of Galphao protein. In the membranes from control subjects, the density of Galphao1 in hippocampus and cortex was the highest, whereas the density of Galphao2 was the lowest and that of Galphao3 was intermediate. In the Alzheimer disease membranes from hippocampus, the density of total Galphao and all three subtype forms was not changed significantly when compared with control values. There were statistically significant alterations in Galphao in cortical membranes from Alzheimer disease when compared with controls. The density of Galphao1 was decreased by approximately 85%, density of Galphao3 was decreased by approximately 95%, and total Galphao density was decreased by approximately 84% of control value. However, Galphao2 density was decreased by approximately 44% but was found not to be statistically different from controls. PMID- 10718206 TI - Strong association of SYT-SSX fusion type and morphologic epithelial differentiation in synovial sarcoma. AB - Synovial sarcoma is characterized by a specific recurrent translocation t(X; 18), resulting in either the SYT-SSX1 or SYT-SSX2 gene fusion. Because this is the primary genetic alteration in these tumors, we sought to identify the impact of molecular heterogeneity of the t(X;18) on cell proliferation, apoptosis, and epithelial differentiation in synovial sarcoma. Seventy-three patients with synovial sarcoma (18 biphasic, 55 monophasic) were selected on the basis of availability of tumor material for molecular and immunohistochemical analysis. Tumors were classified as biphasic on the basis of morphologic glandular differentiation. SYT-SSX fusion transcripts were examined by reverse transcriptase polymerase chain reaction using tumor RNA extracted from frozen or paraffin-embedded tissue. Cell proliferation was assessed immunohistochemically by the Ki-67 labeling index. Apoptosis was analyzed immunohistochemically with BAX and BCL2 antibodies and by the TUNEL method. Immunohistochemical evidence of epithelial differentiation was assessed using antibodies to cytokeratins and epithelial membrane antigen. Approximately two thirds of the tumors had an SYT SSX1 and one third had an SYT-SSX2 fusion transcript. There was a strong association between SYT-SSX fusion type and histologic subtype. All biphasic synovial sarcomas had the SYT-SSX1 fusion, whereas all tumors with SYT-SSX2 were of monophasic morphology. There was, however, no association between SYT-SSX fusion type and expression of cytokeratins and epithelial membrane antigen among monophasic tumors. Tumors with SYT-SSX2 had a significantly higher mean and median Ki-67 labeling index than those with SYT-SSX1, but a comparison of Ki-67 according to fusion type, histologic type, and sample source suggested that the main determinants of proliferation rate were the latter two factors. Specifically, monophasic tumors and metastatic tumors showed significantly higher Ki-67 scores. Apoptosis (by TUNEL) was rarely observed, consistent with prominent expression of the anti-apoptotic protein BCL2 in almost all cases. TUNEL, BCL2, and BAX results did not correlate with SYT-SSX fusion type. These data confirm the strong association of SYT-SSX fusion transcript type with morphologic but not immunophenotypic epithelial differentiation in synovial sarcoma. PMID- 10718208 TI - Accumulation of chromosomal imbalances from intraductal proliferative lesions to adjacent in situ and invasive ductal breast cancer. AB - Carcinoma of the breast is thought to evolve through a sequential progression from normal to proliferative epithelium and eventually into carcinoma. Here lumpectomy specimens from five patients were studied, selected for the presence of ductal hyperplasia without atypia, atypical ductal hyperplasia, ductal carcinoma in situ, and invasive ductal carcinoma. Laser microdissection of tissue allowed precise sampling and direct correlation of phenotypic and genotypic changes. Analyses of the samples revealed an increasing mean number of chromosomal changes occurring with increasing histologic severity, and for the first time chromosomal abnormalities were demonstrated in ductal hyperplasia without atypia. Chromosomal changes found in each of the four histologic entities included gains on 10q, 12q, 16p, and 20q and loss on 13q. In ductal hyperplasia without atypia, gain on 20q as well as loss on 13q was detected with high frequency (four of five samples). Alterations identified in more than 50% of atypical ductal hyperplasia samples included gains on 3p, 8q, 15q, and 22q and loss on 16q. In ductal carcinoma in situ, gain of DNA on 1q and 17q and loss on 4q were additionally found, and in invasive ductal carcinoma, further gains on 6p, 10q, 11q13, and 17p were identified. The chromosomal alterations occurring in the different histopathologic lesions strongly suggest that these regions harbor tumor suppressor genes or oncogenes significant for the development of ductal carcinoma of the breast. PMID- 10718207 TI - Clinical relevance of molecular diagnosis in childhood rhabdomyosarcoma. AB - Rhabdomyosarcoma may be divided into three subtypes--embryonal, alveolar, and undifferentiated sarcoma--which can be distinguished by molecular analysis. The authors applied reverse transcriptase-polymerase chain reaction analysis (RT-PCR) to analyze tumor samples from 14 children with rhabdomyosarcoma for the presence of the chimeric PAX3-FKHR transcript resulting from the translocation t(2;13)(q35,q14). Both fresh and paraffin-embedded tissues were used. In only nine specimens was the RNA intact for the analysis. The chimeric transcript was identified in seven samples: four alveolar type, one embryonal type, and two undifferentiated sarcoma. Histologic review was performed in the three samples with discordance between the molecular and histologic findings. A sample from a patient with a diagnosis of embryonal rhabdomyosarcoma on presentation and expression of PAX3-FKHR fusion transcript yielded a small focus of alveolar rhabdomyosarcoma and was reclassified as alveolar rhabdomyosarcoma. One of the samples from a patient with undifferentiated sarcoma was redefined as alveolar subtype; the diagnosis of the second undifferentiated sarcoma remained unchanged, in accordance with the histologic diagnosis. These findings further support the recommendation that molecular analysis be included in the diagnostic workup of childhood small round cell tumors to reach a more accurate diagnosis for tailoring of specific treatment. PMID- 10718209 TI - Routine analysis of p53 mutation in clinical breast tumor specimens using fluorescence-based polymerase chain reaction and single strand conformation polymorphism. AB - Improved prognostic and predictive markers in breast cancer management would help considerably in therapeutic decision making, particularly in patients with early stage breast cancer. Tumor factors currently used for prognostication and management decisions are tumor size, histologic type and grade, axillary lymph node status, and estrogen receptor content. The discovery of various somatic genetic alterations in breast cancer has raised the possibility that these may provide additional and independent prognostic and predictive information. Alterations of the p53 tumor suppressor gene in particular have received the most attention as potential prognostic and predictive factors. In multivariate analysis, p53 gene mutation is consistently associated with a two- to threefold increased risk of relapse and death from breast cancer. One of the major reasons preventing the introduction of p53 mutation as a routine marker to assist in therapeutic decision making is the lack of a simple, reproducible, and inexpensive assay. In the present study the authors optimized a polymerase chain reaction-based mutation screening method, fluorescence-single strand conformation polymorphism (F-SSCP), that allows p53 status to be assessed accurately and reproducibly in routinely handled, formalin-fixed and paraffin-embedded tumor specimens. The frequency of p53 mutation observed using F-SSCP in a consecutive series of invasive ductal breast carcinomas was 17% (28/164). The authors propose that the prognostic and predictive values of p53 mutation in breast cancer should be further evaluated in prospective, randomized studies using this standardized technique. PMID- 10718210 TI - Tumor-associated overexpression of the soluble T1-S receptor in lymph node negative breast cancer. AB - The oncogene-inducible secreted T1-S glycoprotein is overexpressed in invasive breast carcinomas in mice. As yet, nothing is known about the expression of T1-S in spontaneously occurring human cancers. A report follows on the overexpression of T1-S mRNA in 67% of primary invasive lymph node-negative breast carcinomas (31 of 46 patients) as determined by quantitative reverse transcriptase polymerase chain reaction. Overexpression of T1-S mRNA was independent of the tumor size, the histologic tumor type, and the estrogen-and progesterone-receptor status but was associated with high to moderate differentiation of the tumors (G , G2). T1-S mRNA levels were low to nondetectable in resting normal mammary tissue and benign fibrocystic disease of the breast. Immunohistochemistry confirmed a low to moderate T1 immunoreactivity in epithelial cells of resting mammary tissue and benign fibrocystic disease and highly variable levels of T1 immunoreactivity in breast carcinoma cells. Kaplan-Meier analysis of disease-free survival during a median observation period of 61 months revealed a trend toward a reduced relapse rate and an extended relapse-free survival period for T1-S mRNA--overexpressing breast carcinomas. It is concluded that overexpression of T1-S receptor in lymph node-negative breast cancer may be a potential indicator for tumors with a low metastatic potential. PMID- 10718211 TI - A proposal for the integration of immunohistochemical staining and DNA-based techniques for the determination of TP53 mutations in human carcinomas. AB - The p53 protein plays an important role in the control of the cell cycle and DNA repair. Mutations in the TP53 gene may be a prognostic factor for certain forms of human cancer, with specific mutation sites being associated with significantly worse prognosis, particularly for colorectal and breast cancer. Thus, standardization of accurate, rapid, and cost-effective techniques for the detection of TP53 mutations is a high priority. At present, the only widely available technology that reliably detects and defines all mutations is DNA sequencing. However, the routine sequencing of the entire TP53 gene in all breast and colorectal cancer cases in hospital laboratories is prohibitively costly, complex, and time consuming. In order for the analytical power of DNA to be accessed by the routine laboratory, initial screening using immunohistochemistry, which is widely used as a test for detection of accumulated, mutated protein, followed by heteroduplex analysis of exons 4 to 9 to detect frameshift mutations in immunohistochemistry-negative cases, is proposed. To illustrate the effectiveness of this approach, 28 cases of head and neck squamous-cell carcinomas that were known to contain TP53 mutations were retrospectively analyzed. All missense mutations stained positive on immunohistochemistry using the monoclonal antibody DO7, and all insertions and deletions, even those involving a single nucleotide, were positive using an extremely simple heteroduplex analysis. Only rare nonsense mutations were not detected by this strategy. Nevertheless, application of these results to published data suggests that the prescreening would detect 80% of mutations but would result in a 75% reduction in the sequencing load of the laboratory. PMID- 10718212 TI - TP53 mutations and mdm2 protein overexpression in cholangiocarcinomas. AB - Tumor suppressor protein p53 is a positive regulator of MDM2 gene expression and the mdm2 protein can bind to p53, preventing the transactivation of p53 responsive genes, thus mimicking TP53 mutation. The authors looked for alterations that could affect, directly and indirectly, p53 function in 13 patients with extrahepatic cholangiocarcinoma. Molecular analysis by single strand conformation polymorphism and DNA sequencing revealed that TP53 gene mutations occurred in only 2 of 13 cholangiocarcinomas. High levels of mdm2 protein were found, by immunohistochemical staining, in 61% of the cholangiocarcinomas and in almost all specimens (70%) displaying stabilized p53 protein in the absence and in the presence of TP53 mutations. The finding of co overexpressed mdm2 and p53 proteins in cholangiocarcinomas indicates that they can upregulate the expression of mdm2 protein to a level sufficient for binding and accumulating p53 in a presumably inactive complexed form. The presence of TP53 mutations or upregulation of MDM2 gene expression in 9 of the 13 cholangiocarcinomas strongly supports that the impairment of the p53 pathway is an important and specific step in cholangiocarcinoma pathogenesis. At variance with other authors, no alteration of p16ink4/CDKN2 gene was observed in all 13 cholangiocarcinomas. PMID- 10718213 TI - Correlative immunohistochemical and reverse transcriptase polymerase chain reaction analysis of somatostatin receptor type 2 in neuroendocrine tumors of the lung. AB - Somatostatin receptors type 2 (sst2) have been frequently detected in neuroendocrine tumors and bind somatostatin analogues, such as octreotide, with high affinity. Receptor autoradiography, specific mRNA detection and, more recently, antisst2 polyclonal antibodies are currently employed to reveal sst2. The aim of the present study was to investigate by three different techniques the presence of sst2 in a series of 26 neuroendocrine tumors of the lung in which fresh frozen tissue and paraffin sections were available. It was possible, therefore, to compare, in individual cases, RNA analysis studied by reverse transcriptase polymerase chain reaction (RT-PCR), in situ hybridization (ISH), and immunohistochemistry. A series of 20 nonneuroendocrine lung carcinoma samples served as controls. RT-PCR was positive for sst2 in 22 of 26 samples, including 15 of 15 typical carcinoids, 5 of 6 atypical carcinoids, and 2 of 5 small-cell carcinomas. The sst2 mRNA signal obtained by RT-PCR was strong in the majority (87%) of typical carcinoids and of variable intensity in atypical carcinoids and small-cell carcinomas. A weakly positive signal was observed in 5 of 20 control samples. In immunohistochemistry, two different antibodies (anti-sst2) were employed, including a monoclonal antibody, generated in the Department of Pathology, University of Turin. In the majority of samples a good correlation between sst2 mRNA (as detected by RT-PCR) and sst2 protein expression (as detected by immunohistochemistry) was observed. However, one atypical carcinoid and one small-cell carcinoma had focal immunostaining but no RT-PCR signal. ISH performed in selected samples paralleled the results obtained with the other techniques. A low sst2 expression was associated with high grade neuroendocrine tumors and with aggressive behavior. It is concluded that 1) neuroendocrine tumors of the lung express sst2, and there is a correlation between the mRNA amount and the degree of differentiation; 2) immunohistochemistry and ISH are reliable tools to demonstrate sst2 in these tumors; and 3) sst2 identification in tissue sections may provide information on the diagnostic or therapeutic usefulness of somatostatin analogues in individual patients with neuroendocrine tumors. PMID- 10718214 TI - Genetic imbalances in primary gastric diffuse large B-cell lymphomas: comparison of comparative genomic hybridization, microsatellite, and cytogenetic analysis. AB - Extranodal malignant non-Hodgkin's lymphomas account for about 40% of lymphoid neoplasms, but few data are available concerning the genetic background of primary gastric diffuse large B-cell lymphoma (DLBCL). A study was performed of 27 primary gastric DLBCLs and 5 gastric DLBCLs with a concomitant low grade component of mucosa-associated lymphoid tissue-type lymphoma using comparative genomic hybridization (CGH), microsatellite studies, classic cytogenetics, and fluorescence in situ hybridization (FISH) to search for specific genetic aberrations. The most frequent aberrations were losses of material on chromosome 6q and gains of parts of chromosome 3. In three cases, a total of six high level DNA amplifications were detected, with five of them involving chromosomal regions not having been reported before in gastric DLBCL. A high overall concordance of 91.4% between microsatellite analysis and CGH was observed using DNA extracted from the same tissue block. The concordance achieved using DNA from different tissue blocks of the same patient was 85%. Microsatellite studies, CGH, FISH, and classic cytogenetics represent complementary techniques that facilitate a comprehensive view of genetic alterations in malignancies such as primary gastric DLBCL. PMID- 10718215 TI - More than one. PMID- 10718216 TI - Of golden proportions. PMID- 10718217 TI - Passion of practice: the intuition of treatment. PMID- 10718218 TI - Looking to the past for the future. PMID- 10718219 TI - Using the coefficient of variation to detect sincerity of effort of grip strength: a literature review. AB - Many clinicians use the coefficient of variation (CV) to assess sincerity of effort, without understanding the premise on which it is based or its physiological and mathematical bases. Clinicians who use computerized evaluation systems that calculate the CV may not even be aware of the formula used to derive it. The wide use of the CV in detecting sincerity of effort of grip strength is puzzling, since it lacks empirical support in the literature. This paper examines the physiological rationale for using measures of variability to detect sincerity of effort, the mathematical basis on which the CV is founded, and the reliability and validity of the CV. The conclusions based on this literature review are that the CV is not an appropriate method for determining whether an effort is sincere and that CV values may be inflated in injured patients with compromised hand strength. PMID- 10718220 TI - Elbow positioning for maximum grip performance. AB - Numerous studies have shown that elbow positioning influences grip measurements. These studies have had various and contradictory results. The intent of this study was to clarify the discrepancies and determine which elbow position yields maximal grip strength. Sixty-four men and 64 women were tested on a Greenleaf Medical EVAL system, once with the elbow in full extension and once with the elbow in 90 degrees of flexion. With larger subject groups and computerized data collection, results indicate that-for both the dominant and nondominant hands and regardless of the gender of the subject-grip strength is significantly greater when measured with the elbow in the fully extended position. PMID- 10718221 TI - Validity and reliability of the dexter hand evaluation and therapy system in hand injured patients. AB - This study compares measurements obtained using the Dexter Hand Evaluation and Therapy System with those obtained using manual goniometers and grip and pinch dynamometers. Intrarater and inter-rater reliabilities for each tool were also examined. Repeated digit range of motion and strength measurements were performed on 30 subjects who had prior upper extremity injuries. Three therapists performed all measurements on each subject three times, using both Dexter and manual instruments. Analyses of variance (ANOVAs) and intraclass correlation coefficients (ICCs) were used to assess concurrent validity and therapist reliability. The findings show that the computerized Dexter finger goniometer and grip and pinch dynamometers provide measurements that are statistically similar to those of their manual counterparts. The ANOVA results for concurrent validity showed no significant differences between mean measurement values across therapists for the Dexter tools compared with the manual tools (p> or =0.54). In addition, no significant differences were found among or between the therapists' measurements. The range of ICCs for intrarater and inter-rater reliability for all four tests was 0.86 to 0.99. However, a two-way ANOVA revealed a therapist effect during pinch strength measurements (p< or =0.05), suggesting the need for same-therapist and same-tool measurements until the Dexter pinch dynamometer has been further evaluated. PMID- 10718222 TI - Validity of the Dexter Evaluation System's Jamar dynamometer attachment for assessment of hand grip strength in a normal population. AB - There are several instruments available to measure grip strength, but some instruments are costly, time-consuming to use, or have questionable reliability. The purpose of this study is to examine the concurrent validity of the Dexter Evaluation System with Jamar dynamometer attachment (Dexter) compared with the reference-based criterion of the Jamar adjustable hand dynamometer (Jamar) for measurement of maximal hand grip strength among normal subjects. Sixty-two subjects between the ages of 20 and 50 years, who had no history of hand, arm, shoulder, or neck injuries, were tested with the Jamar in the second handle position and, during the same visit, with the Dexter in the identical position. The Jamar was found to be highly reliable (ICC [3,1] = 0.98) and valid (ICC (2,K) = 0.99) for measuring hand grip strength. In this study, the Dexter was shown to be valid when compared to the Jamar dynamometer for measuring hand grip strength. PMID- 10718223 TI - The treatment of interphalangeal joint flexion contractures with reinforced lycra finger sleeves. PMID- 10718224 TI - Evaluating research studies using the analysis of variance (ANOVA): issues and interpretations. PMID- 10718225 TI - Review of the role of non-invasive ventilation in the emergency department. PMID- 10718226 TI - Discharge instructions for emergency department patients: what should we provide? AB - Effective communication between the physician and patient is required for optimum post-emergency department management. Written emergency department discharge instructions, when used to complement verbal instructions, have been shown to improve communication and patient management. This review examines the purpose, advantages, and disadvantages of three commonly used types of discharge instruction. The desirable features of discharge instructions are described. It is recommended that structured, pre-formatted instruction sheets be provided to all patients discharged to home, that emergency departments establish uniform policies to promote best practice in communication, and that the use of discharge instructions be considered as an emergency department performance indicator. PMID- 10718227 TI - Foreign bodies in the nose and ear: a review of techniques for removal in the emergency department. PMID- 10718228 TI - Survey of the use of rapid sequence induction in the accident and emergency department. AB - OBJECTIVES: To determine the current position regarding the use of rapid sequence induction (RSI) by accident and emergency (A&E) medical staff and the attitudes of consultants in A&E and anaesthetics towards this. METHODS: A questionnaire was designed that was distributed to consultant anaesthetists and A&E physicians in hospitals receiving over 50,000 new A&E patients per year. RESULTS: A total of 140 replies were received (a response rate of 72%). The breakdown of results is shown. There was wide difference of opinion between anaesthetists and A&E consultants as to who performs RSI at present in their A&E departments, however two thirds of anaesthetists thought A&E staff with appropriate training and support should attempt RSI either routinely or in certain circumstances. CONCLUSIONS: A&E staff in several hospitals routinely undertake RSI and the majority of A&E consultants thought that RSI would be undertaken by A&E staff if an anaesthetist were unavailable. There is disagreement regarding the length of anaesthetic training required before A&E medical staff should undertake RSI. PMID- 10718229 TI - Characteristics of children and adolescents presenting to accident and emergency departments with deliberate self harm. AB - OBJECTIVES: The aim of this study was to provide a description of the characteristics of children and adolescents presenting to the accident and emergency (A&E) department with deliberate self harm. METHODS: Descriptive analysis of data collected by reviewing the notes of all children and adolescents aged 16 years and under, presenting during the period of study (1 January to 31 December) with a history of deliberate self harm. RESULTS: A total of 100 children (18 boys, 82 girls) were responsible for 117 episodes of deliberate self harm. Nine repeaters were responsible for 22% of the attendances; 38% had made use of emergency ambulance service and 6% were referred by their general practitioner (GP). Sixty nine per cent were accompanied by immediate family and 21% children presented alone. Seventy four per cent presented within three hours of the attempt and 37% presented between 6 pm and midnight; 77% presented during weekdays and 30% of attempts had occurred during spring. Ninety two per cent had used a pharmaceutical drug. Sixty five per cent had made the attempt at home and 12% in a public place. Twenty five per cent had prior or current contact with the child psychiatric services and a similar proportion had prior or current contact with social services. CONCLUSIONS: Few of the children and adolescents presenting with deliberate self harm to the A&E department have been referred by their GP. They frequently present alone or are accompanied by people who are not family members making assessment and treatment difficult. Many already have other services involved in their care and thus the gathering and dissemination of information can become quite lengthy. The time of presentation is usually out of hours, further complicating this process. A small number of young people present with repeated self harm, who are known to be most vulnerable for completing suicide. PMID- 10718230 TI - Triage nurse requested x rays--are they worthwhile? AB - OBJECTIVE: To study an established triage nurse x ray requesting system to determine whether sending defined groups of patients for radiography before assessment by doctors or emergency nurse practitioners (ENPs) resulted in shorter waiting times for patients without compromising quality of care. METHODS: Prospective randomised controlled study of "walking wounded" patients attending a district general hospital. Data were collected over two separate two week periods, six months apart, in the middle of two senior house officer appointment periods. A total of 675 patients were entered into the study. Analysis of results was achieved using standard statistical methods. RESULTS: Altogether 335 patients were in the nurse x ray group and 340 in the control group. The triage categories of the groups were similar. A 36% mean time reduction of 37.2 min (95% confidence interval 30.2 to 44.2, p=0.000) from time of triage to time of treatment decision was achieved in the nurse requested group. Triage nurses requested 8% (p=0.002) fewer x rays than doctors or ENPs and had a 6% higher positive "hit" rate (p=0.03). In 7.8% (26 cases), patients in the triage nurse group were judged to require radiographs or further views by the doctor or ENP; of these, 11 cases showed a positive finding on radiography. The time from triage to assessment by doctor or ENP was not lengthened by prior requesting of radiography (nurse x ray group 64.4 min, control group 63.7 min, p=0.79). CONCLUSIONS: A triage nurse x ray requesting system speeds up the progress of walking wounded patients through the department without compromising service quality. Further benefits are staff and patient satisfaction and a greater sense of team working for all staff. PMID- 10718231 TI - Triage nurse requested x rays--the results of a national survey. AB - OBJECTIVE: To ascertain the prevalence and experiences of triage nurse requested x ray systems among accident and emergency (A&E) departments in the UK. METHOD: A descriptive study of a postal survey of 225 major A&E departments listed in the British Association for Accident and Emergency Medicine directory. RESULTS: Altogether 165 (73%) questionnaires were returned. Fifty nine (35%) departments indicated that they currently had a triage nurse requested radiology system. Of those departments that did not have such a system, the main reasons were that it was not necessary, radiologists or A&E consultants were opposed to the idea, that nurses were not capable/did not want the system, or that it would delay triage. Of those departments that do operate a nurse requesting system, most have started doing so in the last three years, and allow nurses of E grade and above who have completed an in house training course and radiation protection certificate to request x rays. Protocols vary, but usually allow requests for limb radiology in patients over 5 years old. Many departments have audited their system, with positive results. In all departments that currently operate the system, staff and patients felt that the system was either good or excellent. One department abandoned the system, after a trial, because they felt that x ray requesting was not a nursing role. CONCLUSION: The system of triage nurse requested x rays is generally well received and departments considering adopting this system can be reassured. Pitfalls and possible protocols for A&E departments intending to start triage nurse requested x rays are suggested. PMID- 10718232 TI - Complications of tube thoracostomy in trauma. AB - OBJECTIVE: To assess the complication rate of tube thoracostomy in trauma. To consider whether this rate is high enough to support a selective reduction in the indications for tube thoracostomy in trauma. METHODS: A retrospective case series of all trauma patients who underwent tube thoracostomy during a 12 month period at a large UK teaching hospital with an accident and emergency (A&E) department seeing in excess of 125,000 new patients/year. These patients were identified using the hospital audit department computerised retrieval system supplemented by a hand search of both the data collected for the Major Trauma Outcome Study and the A&E admission unit log book. The notes were assessed with regard to the incidence of complications, which were divided into insertional, infective, and positional. RESULTS: Fifty seven chest drains were placed in 47 patients over the 12 month period. Seven patients who died within 48 hours of drain insertion were excluded. The commonest indications for tube thoracostomy were pneumothorax (54%) and haemothorax (20%); 90% of tubes were placed as a result of blunt trauma. The overall complication rate of the procedure was 30%. There were no insertional complications and only one (2%) major complication, which was empyema thoracis. CONCLUSION: This study reveals no persuasive evidence to support a selective reduction in the indications for tube thoracostomy in trauma. A larger study to confirm or refute these findings must be performed before any change in established safe practice. PMID- 10718233 TI - Chemical protective clothing; a study into the ability of staff to perform lifesaving procedures. AB - OBJECTIVE: To investigate the ability of medical and nursing staff to perform certain tasks while wearing a chemical protection suit with a respirator. Tasks chosen were those that would be required before decontamination. METHODS: Ten experienced accident and emergency doctors (middle grade and consultants) and 10 nurses were asked to perform certain tasks that were judged to be life saving, relevant to triage, or necessary to confirm death, on an advanced life support manikin, while wearing a TST-Sweden chemical protection suit. The operators were objectively assessed by one of the authors for achieving each task, then asked to make a subjective assessment of the difficulty experienced. RESULTS: Medical staff were asked to ventilate the manikin using a bag-valve-mask, intubate within 30 seconds, apply monitor electrodes and cables and check cardiac rhythm, apply gel pads and defibrillate safely, and finally, fold the cruciform triage card to show "RED", and attach it to the manikin. All the doctors completed these tasks, except for one, who could only intubate the manikin after several attempts. Nursing staff were asked to open and apply an oxygen mask, adjust oxygen flow, size and insert an oropharyngeal airway, ventilate the manikin using a bag-valve mask, apply a pressure bandage to a limb, and fold the cruciform triage card to show "YELLOW", and attach it to the manikin. All the nurses completed these tasks. Operators reported varying degrees of difficulty, the most difficult tasks were those requiring fine movements or delicate control. Generally, operators found the butyl rubber gloves cumbersome. Communication difficulties were frequently reported. Although only intubation was formally timed, tasks were perceived to take longer. Some operators found the suits too warm and uncomfortable. CONCLUSION: Should the need arise, the TST-Sweden chemical protection suits would enable experienced doctors and nurses to perform lifesaving measures effectively, without significant impairment to their skills. Tasks would be easier to accomplish with better fitting gloves. PMID- 10718234 TI - Public understanding of medical terminology: non-English speakers may not receive optimal care. AB - INTRODUCTION: Many systems of telephone triage are being developed (including NHS Direct, general practitioner out of hours centres, ambulance services). These rely on the ability to determine key facts from the caller. Level of consciousness is an important indicator after head injury but also an indicator of severe illness. AIMS: To determine the general public's understanding of the term unconscious. METHODS: A total of 700 people were asked one of seven questions relating to their understanding of the term unconscious. All participants were adults who could speak sufficient English to give a history to a nurse. RESULTS: Correct understanding of the term unconscious varied from 46.5% to 87.0% for varying parameters. Those with English as their first language had a better understanding (p<0.01) and there was a significant variation with ethnicity (p<0.05). CONCLUSIONS: Understanding of the term unconscious is poor and worse in those for whom English is not a first language. Decision making should not rely on the interpretation of questions using technical terms such as unconscious, which may have a different meaning between professional and lay people. PMID- 10718235 TI - Activities of accident and emergency consultants--a time and motion study. AB - The work of an accident and emergency (A&E) consultant is not clearly defined. There is difficulty in fixing a job plan due to the unpredictable workload. This study shows the daily activities of nine consultants in A&E over a one month period. The results suggest that A&E consultants vary tremendously in the content of their working day, although a large proportion of hours is spent on administrative duties in all cases. Predictable variations occur between a single handed clinical director who spends 60% of his time in management and the consultant in a multiconsultant department who spends 74% of his time in clinical care and teaching. None of the consultants studied spent more than 48% of their time in clinical contact. A&E remains a specialty with no consistency between activities of consultants, and where opportunities exist to pursue special interest. Training must ensure adequate attention to management and methods of support for new consultants in their management role must be found. PMID- 10718236 TI - Towards evidence based emergency medicine: best BETS from the Manchester Royal Infirmary. Signs and symptoms of oesophageal coins. PMID- 10718237 TI - Towards evidence based emergency medicine: best BETS from the Manchester Royal Infirmary. Immobilisation of suspected scaphoid fractures. PMID- 10718238 TI - Towards evidence based emergency medicine: best BETS from the Manchester Royal Infirmary. Activated charcoal in tricyclic antidepressant overdose. PMID- 10718239 TI - Towards evidence based emergency medicine: best BETS from the Manchester Royal Infirmary. Analgesia and assessment of abdominal pain. PMID- 10718240 TI - Mentoring--the trainee's perspective. PMID- 10718241 TI - Discovery of the intraosseous route for fluid administration. AB - One of the many problems in the resuscitation of the shocked patient is how to gain access to the circulation to provide fluids or drugs. Since the 1830s fluids have been administered intravenously. Intravenous access is not always possible in the very shocked patient. An alternative, used in the first world war, was the rectal route. This has rarely been used on a large scale since. Just before the outbreak of the second world war a chance discovery resulted in the development of intraosseous infusions of fluid and drugs. From its discovery it was used in adults and children. For many years it seemed to be ignored in adult resuscitation, but there are now signs of renewed interest in the technique. This brief review traces the discovery of the intraosseous route to put the current developments into a historical context. PMID- 10718242 TI - Domestic violence: the shaken adult syndrome. AB - A case of domestic violence is reported. The patient presented with the triad of injuries associated with the shaking of infants: retinal haemorrhages, subdural haematoma, and patterned bruising; this has been described as the shaken adult syndrome. This case report reflects the difficulties in diagnosing domestic violence in the accident and emergency setting. PMID- 10718243 TI - Airbag associated fatal head injury: case report and review of the literature on airbag injuries. AB - Airbags have been shown to significantly reduce mortality and morbidity in motor vehicle crashes. However, the airbag, like the seat belt, produces its own range of injuries. With the increasing use of airbags in the UK, airbag associated injuries will be seen more often. These are usually minor, but in certain circumstances severe and fatal injuries result. Such injuries have been described before in the medical literature, but hitherto most reports have been from North America. This is the first case report from the UK of serious injury due to airbag deployment and describes the case of a driver who was fatally injured when her airbag deployed in a moderate impact frontal collision where such severe injury would not normally have been anticipated. The range of airbag associated injuries is described and predisposing factors such as lack of seat belt usage, short stature, and proximity to airbag housing are discussed. The particular dangers airbags pose to children are also discussed. PMID- 10718244 TI - Anaphylaxis and monoamine oxidase inhibitors--the use of adrenaline. AB - A 67 year old woman taking a monoamine oxidase inhibitor (MAOI) presented to the accident and emergency department with an anaphylactic reaction to flucloxacillin. This case highlights the uncertainty regarding the use of adrenaline (epinephrine) in the context of concurrent MAOI use. A summary of the evidence is presented to clarify this. PMID- 10718245 TI - Prolonged coma due to cerebral fat embolism: report of two cases. AB - Fat embolism syndrome remains a rare, but potentially life threatening complication of long bone fractures. The true incidence is difficult to assess as many cases remain undiagnosed. Cerebral involvement varies from confusion to encephalopathy with coma and seizures. Clinical symptoms and computed tomography are not always diagnostic, while magnetic resonance imaging is more sensitive in the detection of a suspected brain embolism. Two cases of post-traumatic cerebral fat embolism, manifested by prolonged coma and diffuse cerebral oedema, are presented. The clinical course of the disease as well as the intensive care unit management are discussed. PMID- 10718246 TI - Air rifle injury to the oropharynx. The essential role of computed tomography in deciding on surgical exploration. AB - Gunshot wounds specific to the oropharynx are extremely rare with no reported cases of such injury in the world literature. The importance of such cases rests on the use of modern imaging techniques including computed tomography, magnetic resonance imaging, and vascular imaging in the making of management decisions and particularly in deciding the need for exploration of such an injury. In our case a conservative approach was adopted in view of the computed tomography finding and the stable clinical condition of the patient. PMID- 10718247 TI - A useful form of glue ear. AB - Children and adults commonly present to the emergency department with a foreign body lodged in the ear. Over the past 15 years techniques for cyanoacrylate ("superglue") assisted foreign body removal have been described, but are not widely employed. Two cases of successful and one of unsuccessful removal using this technique are reported, and some advice is offered to aid others. PMID- 10718248 TI - Asymptomatic cerebellar medulloblastoma unmasked by minor head injury. PMID- 10718249 TI - Intramuscular or intravenous adrenaline in acute, severe anaphylaxis? PMID- 10718250 TI - Medical treatment of anaphylaxis. PMID- 10718251 TI - Future inpatient management of patients with minor head injuries. PMID- 10718252 TI - Cycle helmets. PMID- 10718253 TI - Playing in the back seat. PMID- 10718254 TI - Oesophageal rupture. PMID- 10718255 TI - Carbon monoxide poisoning. PMID- 10718256 TI - Evidence based and guideline based medicine. PMID- 10718257 TI - Weekly web review. PMID- 10718258 TI - The effect of dopamine depletion from the caudate nucleus of the common marmoset (Callithrix jacchus) on tests of prefrontal cognitive function. AB - This study examined the effects of depletion of dopamine from the caudate nucleus of the common marmoset (Callithrix jacchus), on tasks sensitive to prefrontal damage (attentional set-shifting and spatial delayed response). There was a marked impairment in performance on the spatial delayed response task, but performance on the attentional set-shifting task was relatively preserved except for an impairment in re-engagement of a previously relevant perceptual dimension. This pattern of impairment is distinct from that seen after excitotoxic lesions of the prefrontal cortex and in patients with Parkinson's disease. Though it is not possible to identify specific cognitive functions that are independent of dopaminergic modulation of the caudate nucleus, due to the partial nature of the lesion, the results do provide insight into those cognitive processes that appear most dependent on caudate dopamine. PMID- 10718259 TI - Perirhinal cortex ablation in rats selectively impairs object identification in a simultaneous visual comparison task. AB - In Experiment 1, rats discriminated among computer-generated visual displays (scenes) comprising 3 different shapes (objects). One constant scene (unrewarded) appeared on every trial together with a trial-unique variable scene (rewarded). Four types of variable scene were intermingled: (a) unfamiliar objects in different positions from the constant; (b) unfamiliar objects in same positions as the constant; (c) same objects as the constant in different positions; (d) same objects and positions, recombined. Aspiration lesions of perirhinal cortex impaired performance with type (b) only. Experiment 2 tested spatial delayed nonmatching-to-sample. The perirhinal group were impaired nonsignificantly, and less than fornix-transected rats in an earlier study. Rats' perirhinal cortex, like monkeys', subserves object identification in the absence of memory requirement but does not contribute substantially to hippocampal system spatial memory function. PMID- 10718260 TI - Muscarinic cholinergic neuromodulation reduces proactive interference between stored odor memories during associative learning in rats. AB - Previous electrophysiological studies and computational modeling suggest the hypothesis that cholinergic neuromodulation may reduce olfactory associative interference during learning (M. E. Hasselmo, B. P. Anderson, & J. M. Bower, 1992; M. E. Hasselmo & J. M. Bower, 1993). These results provide behavioral evidence supporting this hypothesis. A simultaneous discrimination task required learning a baseline odor pair (A+B-) and then, under the influence of scopolamine, a novel odor pair (A-C+) with an overlapping component (A) versus a novel odor pair (D+E-) with no overlapping component. As predicted by the model, rats that received scopolamine (0.50 and 0.25 mg/kg) were more impaired at acquiring overlapping than nonoverlapping odor pairs relative to their performance under normal saline or methylscopolamine. These results support the prediction that the physiological effects of acetylcholine can reduce interference between stored odor memories during associative learning. PMID- 10718261 TI - Disconnection of the anterior cingulate cortex and nucleus accumbens core impairs Pavlovian approach behavior: further evidence for limbic cortical-ventral striatopallidal systems. AB - The nucleus accumbens (NAcc) has been implicated in a variety of forms of reward related learning, reflecting its anatomical connections with limbic cortical structures. After confirming that excitotoxic lesions of the anterior cingulate cortex (Ant Cing) impaired the acquisition of appetitive Pavlovian conditioning in an autoshaping procedure, the effects of excitotoxic lesions to the NAcc core or shell on autoshaping were also assessed. Only selective core lesions impaired Pavlovian approach. A subsequent experiment studied the effects of a disconnection of the Ant Cing and NAcc core, using an asymmetric lesion procedure, to determine whether these structures interact sequentially as part of a limbic corticostriatal system. Such lesioned rats were also significantly impaired relative to controls at autoshaping. These results demonstrate that the NAcc core and Ant Cing are "nodes" of a corticostriatal circuit involved in stimulus-reward learning. PMID- 10718262 TI - Disruption of contextual freezing, but not contextual blocking of fear potentiated startle, after lesions of the dorsal hippocampus. AB - The role of the dorsal hippocampus in contextual fear conditioning was investigated with a contextual blocking paradigm. In Experiment 1, rats were given pairings of a light conditioned stimulus (CS) and footshock after preexposure either to footshock or to the context alone. The group preexposed to footshock showed poorer fear conditioning to the light CS, as measured by the fear-potentiated startle reflex. In Experiment 2, a group preexposed to footshock in the same context showed poorer fear conditioning to the light CS than did a group preexposed to footshock in a different context, indicating contextual blocking of fear-potentiated startle. In Experiment 3, lesions of the dorsal hippocampus had no effect on contextual blocking, even though contextual freezing was disrupted. The sparing of contextual blocking indicated that contextual memory was intact following hippocampal lesions, despite the disruption of contextual freezing. PMID- 10718263 TI - The development of declarative memory in human infants: age-related changes in deferred imitation. AB - In 2 experiments, deferred imitation procedures were used to trace age-related changes in declarative memory by human infants over the first 2 years of life. An adult modeled 3 actions with an object, and infants' ability to reproduce those actions was assessed 24 hr later. Some infants were tested with a new object or in a new context relative to the original demonstration. Changes in the context disrupted the performance of 6-month-olds but had no effect on the performance of 12- and 18-month-olds. Changes in the object disrupted the performance of 6- and 12-month-olds but had no effect on the performance of 18-month-olds. This age related increase in representational flexibility may account for the decline of childhood amnesia during the 3rd year of life. PMID- 10718264 TI - N-methyl-D-aspartate receptor-dependent plasticity within a distributed corticostriatal network mediates appetitive instrumental learning. AB - The effect of microinfusion of the N-methyl-D-aspartate (NMDA) antagonist 2-amino 5-phosphonopentanoic acid (AP-5) into the amygdala, medial prefrontal cortex, and dorsal and ventral subiculum on acquisition of a lever-pressing task for food in rats was examined. Serial transmission between the basolateral amygdala and nucleus accumbens core was also examined in an asymmetric infusion design. AP-5 administered bilaterally into either the amygdala or medial prefrontal cortex markedly impaired learning, whereas administration into the dorsal or ventral subiculum had no effect. Unilateral infusion of AP-5 into either the nucleus accumbens core or amygdala was also sufficient to impair learning. These data provide novel evidence for NMDA receptor-dependent plasticity within corticostriatal networks in the acquisition of appetitive instrumental learning. PMID- 10718265 TI - The induction of c-Fos in the NTS after taste aversion learning is not correlated with measures of conditioned fear. AB - The induction of c-Fos-like immunoreactivity (c-FLI) in the nucleus of the solitary tract (NTS) has been shown to be correlated with behavioral expression of a conditioned taste aversion (CTA). However, because this cellular response is also dependent on an intact amygdala, it may represent the activation of a stress related autonomic response. The present experiments addressed this possibility by evaluating the correlation between c-FLI in the intermediate division of the NTS (iNTS) and 2 measures of conditioned fear: freezing and changes in mean arterial pressure (MAP) and heart rate (HR). Exposure to the taste conditioned stimulus (CS) resulted in a marked induction of c-FLI in the iNTS, whereas exposure to a fear CS did not. Further, exposure to a taste CS did not selectively lead to increases in MAP or HR. Results suggest that induction of c-FLI in the iNTS may reflect the activation of a cell population whose function is unique to the CTA paradigm. PMID- 10718266 TI - Amygdala lesions do not impair shock-probe avoidance retention performance. AB - The present experiment used the shock-probe paradigm, a procedure usually used to assess anxiolytic processes, to assess memory in amygdala-lesioned rats. Rats were placed in a chamber that contained a probe protruding from 1 of 4 walls and were kept there for 15 min after they contacted the probe. For half the rats, the probe was electrified (2 mA). Four days later, sham or neurotoxic amygdala lesions were induced. Retention performance was assessed 8 days later by measuring the latency to contact the probe and the number of contact-induced shocks. The results indicated that, although shock-naive amygdala-lesioned rats were impaired on the 2nd shock-probe test, shock-experienced amygdala-lesioned rats were not. These data indicate that the memory of a shock experience, as indexed with a shock-probe avoidance response, is spared in rats with large amygdala lesions. PMID- 10718267 TI - Selective associations in day-old chicks: when do CS traces become available for sickness-conditioned learning? AB - Barber and colleagues (T. A. Barber, A. M. Klunk, P. D. Howorth, M. F. Pearlman, & K. E. Patrick, 1998; T. A. Barber, M. F. Pearlman & K. E. Patrick, 1995) showed that male domestic chicks would not peck a bead presented 15 or more min before a lithium chloride (LiCl) injection, but would peck a bead presented less than 15 min before a LiCl injection. In 4 experiments, the authors (a) confirmed their initial observation, (b) showed that the effect is not due to retrograde amnesia produced by LiCl, and (c) confirmed that memories less than 15 min old are available for other types of learning. PMID- 10718268 TI - Mu opioid receptors in the ventrolateral periaqueductal gray mediate stress induced analgesia but not immobility in rat pups. AB - Rat pups become immobile and analgesic when exposed to an adult male rat. The aim of this study was to determine whether these reactions are under the control of endogenous opioids and to determine the role of the midbrain periaqueductal gray (PAG), which mediates stress-induced immobility and analgesia in adult animals. In Experiment 1, 14-day-old rats were injected systemically with the general opioid receptor antagonist naltrexone (1 mg/kg), which blocked male-induced analgesia to thermal stimulation but did not affect immobility. In Experiment 2, the selective mu opioid receptor antagonist D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr NH2 (CTOP; 50 or 100 ng/200 nl) was microinjected into the ventrolateral and lateral PAG. CTOP suppressed male-induced analgesia when injected into the ventrolateral PAG. Male-induced immobility was not affected by CTOP. Male proximity therefore seems to induce analgesia in rat pups by releasing endogenous opioids that bind to mu opioid receptors in the ventrolateral PAG. PMID- 10718269 TI - A comparison of the effects of bilateral and unilateral infusions of muscimol into the basal forebrain on cued detection of visual targets in rats. AB - This study investigated the role of the basal forebrain cholinergic system (BFCS) in rats' performance of a visuospatial attention task. Muscimol was infused bilaterally and unilaterally into the BFCS to inhibit cholinergic projections to the cortex. Muscimol slowed responding without significantly affecting side-bias. Bilateral infusions increased accuracy for all targets, whereas unilateral infusions reduced accuracy for targets contralateral to the infusion and increased accuracy for targets ipsilateral to the infusion. After a low unilateral dose of muscimol, invalid cues impaired detection of contralateral targets and spared detection of ipsilateral targets. A high unilateral dose of muscimol impaired detection of contralateral targets independently of cueing. These results suggest that interhemispheric imbalance in cortical activity by pharmacological manipulation of the BFCS can impair the detection of lateralized visual stimuli. PMID- 10718270 TI - Anticipatory errors after unilateral lesions of the subthalamic nucleus in the rat: evidence for a failure of response inhibition. AB - The nature of anticipatory responding in rats with unilateral subthalamic nucleus lesions was examined. Rats were trained to respond toward visual targets that were preceded by 1 of 4 different cues. For normal rats, a cue invokes an involuntary attentional orienting that enhances processing of the target at the location of attention. The cue is also a salient stimulus to which a response must be suppressed. Therefore, this task was used to investigate possible attentional impairments, as well as the ability of a lesioned rat to suppress competing motor programs. Responding under target control was not affected by the lesion. There was an increase in anticipatory responses before target onset, which could be accounted for by a failure to inhibit contralateral responses. PMID- 10718271 TI - Plasticity in the maternal circuit: effects of experience and partum condition on brain astrocyte number in female rats. AB - Female rats that have received a maternal experience undergo enhanced c-fos expression in a number of brain sites when reexposed to pups. The present 2 studies examined changes in the expression of another brain protein, glial fibrillary acidic protein (GFAP), which is a major unit of the astrocytic cytoskeleton. In both experiments, primiparous and multiparous female rats were given varying amounts of postpartum contact with pups and overdosed after varying intervals, with no pups. Brains were prepared for GFAP immunohistochemical analysis. In both studies, Day 5 postpartum multiparous subjects given additional postpartum contact with pups, when compared with pup-exposed primiparous subjects, were found to have significantly higher numbers of GFAP positive cells in the medial preoptic area of the hypothalamus, an area critical for the expression of maternal behavior, but not in control sites. In Experiment 2, an opposite effect of parity was found in the medial amygdala and habenula. PMID- 10718272 TI - Dopamine D2 receptors in the nucleus accumbens are important for social attachment in female prairie voles (Microtus ochrogaster). AB - The prairie vole (Microtus ochrogaster), a monogamous rodent that forms long lasting pair bonds, has proven useful for the neurobiological study of social attachment. In the laboratory, pair bonds can be assessed by testing for a partner preference, a choice test in which pair-bonded voles regularly prefer their partner to a conspecific stranger. Studies reported here investigate the role of dopamine D2-like receptors (i.e., D2, D3, and D4 receptors) in the nucleus accumbens (NAcc) for the formation of a partner preference in female voles. Mating facilitated partner preference formation and associated with an approximately 50% increase in extracellular dopamine in the NAcc. Microinjection of the D2 antagonist eticlopride into the NAcc (but not the prelimbic cortex) blocked the formation of a partner preference in mating voles, whereas the D2 agonist quinpirole facilitated formation of a partner preference in the absence of mating. Taken together, these results suggest that D2-like receptors in the NAcc are important for the mediation of social attachments in female voles. PMID- 10718273 TI - The role of the hippocampal system in social odor discrimination and scent marking in female golden hamsters (Mesocricetus auratus). AB - Female golden hamsters (Mesocricetus auratus) received aspiration lesions of the parahippocampal region (PARA) or electrolytic lesions of the fimbria-fornix (FNX) and were tested for their (a) discrimination between odors of individual males in a habituation-discrimination task, (b) preference for male over female odors, and (c) scent-marking in response to conspecific odors. Both lesion groups habituated to repeated presentations of a male's odor. However, only FNX females discriminated between scents of individual males, whereas PARA females did not. Neither lesion eliminated female preferences for male odors. Females with FNX lesions showed decreased levels of scent marking, but those with PARA lesions had more subtle deficits. Thus, the PARA, but not the subcortical connections of the hippocampus, is critical for discrimination of the odors of individuals. PMID- 10718274 TI - The medial preoptic area, necessary for adult maternal behavior in rats, is only partially established as a component of the neural circuit that supports maternal behavior in juvenile rats. AB - To determine whether the neurons of the medial preoptic area (MPOA) are necessary for pup-induced maternal behavior (MB) in juvenile and adult rats, subjects received bilateral injections of the neurotoxin N-Methyl-D-Aspartic acid into the MPOA. Controls were intact or were sham treated by surgical placement of the syringe barrel. The rats were then induced into MB by constant pup exposure. Starting at 27 (juvenile) or 60 (adult) days of age, rats were tested for MB for 12 consecutive days. After histological analysis, rats were categorized as having either large or small lesions of the MPOA. In juveniles, large lesions of the MPOA blocked retrieval and impaired nest-building, but crouching behavior was unaffected; small lesions had no effect on MB. In contrast, in adults, large or small lesions severely impaired all components of MB. The results suggest that in juvenile rats, the role of the MPOA neurons in MB is only partially established, whereas by 60 days of age, the unsubstitutable role of the MPOA in the neural circuit that mediates MB is fully established. PMID- 10718275 TI - The effects of combined lesions of the subicular complex and the entorhinal cortex on two forms of spatial navigation in the water maze. AB - The role of the subicular complex and entorhinal cortex (SUB-EC) in spatial learning was examined in 2 water maze experiments. In Experiment 1, rats had to locate a hidden platform that was always a fixed distance and direction from an intramaze landmark. Each day, the landmark and platform were moved to a new location. Both control and SUB-EC-lesioned rats learned to locate the platform equally readily during training. However, the control group was impaired in locating the platform when the visual extramaze cues were concealed, whereas the lesioned group was unaffected by this manipulation. In Experiment 2, the lesioned rats were impaired in finding a hidden platform that was in a fixed place in the water maze and showed no evidence of having learned its location in a probe test. These results suggest that damage to the SUB-EC impairs the integration of geometric information but spares a more general navigational-directional strategy. PMID- 10718276 TI - Rationales for treating IgA nephropathies. AB - An outline is given of the pathophysiology of IgA nephropathy (IgA) in order to emphasize the role of eicosanoids, angiotensin II, and reactive oxygen species. ACE inhibitors and early corticosteroid usage are prime therapies. Tonsillectomy is to be considered, certainly for individual cases. It is logical that other components of a cocktail could be (i) thromboxane antagonists, (ii) leukotriene antagonists, or (iii) PAF antagonist. In theory there should be benefit from antioxidants. Fish oils have not come up to expectation. PDGF aptamers look promising for the prevention of mesangial cell proliferation. Heparins are not used in the way that they could be. Various other agents could help reduce decline. PMID- 10718277 TI - Antiproteinuric effect of calcium antagonists on puromycin-induced experimental nephrosis. AB - Calcium antagonists have a potential for beneficial effects on kidney function unrelated to their antihypertensive action. In this study we have investigated the efficacy of calcium antagonists compounds (verapamil, nifedipine and diltiazem) on reversible acute renal insufficiency, proteinuria and interstitial nephritis induced by the puromycin ammonucleoside (PAN). An increase in blood pressure (BP) was detected on day 14, with no statistical differences in the response to calcium antagonists. Serum creatinine concentration increased to 1.2 mg/dL on day 7 after PAN and decreased to 0.7 mg/dL at 14 days, calcium antagonists shortened the time required to reach baseline or control levels. Calcium antagonists also reduced proteinuria in the PAN-treated animals, in both day 7 and day 14. Differential effects of the antagonists were observed. Verapamil caused a greater reduction (p < 0.01) in proteinuria than nifedipine or diltiazem in day 7. Moreover, verapamil (p < 0.01) and nifedipine (p < 0.01) reduced the total number of interstitial infiltrating leukocytes from 690 to 120 and 425 positive cells/20 high power fields (x63) respectively, by contrast, diltiazem had no effect. We conclude that in this model of PAN nephropathy verapamil is more effective in reducing both proteinuria and the severity of acute interstitial nephritis than either nifedipine or diltiazem. The possible clinical implications of these results remain to be elucidated. PMID- 10718278 TI - Chloroquine inhibits arginine vasopressin production in isolated rat inner medullary segments induced cAMP collecting duct. AB - Previous studies showed that acute chloroquine administration increases plasma arginine vasopressin (AVP) concentration in the rat without influencing urine flow rate. The present study was designed to investigate whether chloroquine inhibits the AVP-induced cAMP production that mediates the antidiuretic effects of vasopressin. Single inner medullary collecting duct (IMCD) segments were pre incubated at 35 degrees C for 10 min followed by 4 min at 37 degrees C with combinations of AVP and/or chloroquine with 1 mM 3-isobutyl-I-methylxanthine (IBMX) and cAMP concentrations were measured by radioimmunoassay. To establish the possible site of interference in cAMP production IMCD segments were incubated in the presence of chloroquine and forskolin. Chloroquine at concentrations ranging from 10(-9) M to 10(-6) M did not affect cAMP production by comparison with control. However, AVP (10(-8) M) and forskolin (10(-6) M) significantly (p < 0.01) increased cAMP accumulation. Chloroquine at all concentrations significantly suppressed the AVP stimulated cAMP production (e.g., chloroquine (10(-8) M) + AVP (10(-8) M) 41 +/- 12 fmol/4 mm (n = 9 tubules) vs. AVP (10(-8) M) alone 82 +/- 9 fmol/4 min/mm (n = 37 tubules). Chloroquine at all concentrations tested did not have any effect an forskolin-induced cAMP production. The data suggest that chloroquine inhibits the AVP induced cAMP production at the level of hormone/receptor complex. This possibly explains the previously reported lack of the normal antidiuretic responses of AVP in rats following chloroquine administration. PMID- 10718279 TI - Total dose iron infusion: safety and efficacy in predialysis patients. AB - Iron deficiency anemia is not uncommon in predialysis patients. Oral iron often cannot maintain adequate iron stores. Hence we evaluated the safety and efficacy of total dose infusion (TDI) of iron in these patients. Anemic predialysis patients were screened and those with Hb < 7.0 g/dL and serum ferritin < 200 ng/mL were selected. Patients with active bleeding and acute liver disease were excluded. All patients were on oral iron 100 mg/day. None of the patients were on erythropoeitin. 11 patients (6 males and 5 females), aged 45.9 +/- 15 yrs, were suitable. Hb was 5.9 +/- 1.0 g/dL and serum ferritin was 89.5 + 50 ng/mL. The preparation used was iron dextran. A test dose of 25 mg in 100 mL normal saline was administered over 1 hr to all patients. One patient had fever and chills during the test dose and was not given TDI. 10 patients received TDI. None of these patients had any problem during the infusion. The dose of iron administered was 900 + 316.2 mg. One patient who received 1600 mg had arthralgia-myalgia and another patient had thrombophlebitis following TDI. One month after TDI, Hb was 8.0 + 1.0 g/dL and serum ferritin was 362 ng/mL. We feel that TDI is a safe and effective method of correcting iron deficiency in predialysis patients. PMID- 10718280 TI - Effects of prolonged infusion of the natriuretic peptides Escherichia coli enterotoxin and atrial natriuretic peptide on the outcome of acute ischemic renal failure in the rat. AB - Atrial Natriuretic Peptide (ANP), has protective effect on the outcome of acute renal failure in animals when infused over short periods of time. Escherichia Coli heat-stable enterotoxin (STa) acts on a particulate guanylate cyclase receptors leading to an increase in cytosolic cGMP similar to ANP. STa has longer duration of action and induces significant natriuresis in the healthy kidney, however. Therefore the effects of prolonged infusion of STa and ANP on the outcome of ischemic Acute Renal Failure (ARF) was assessed in the rat. Three groups of rats were used in the experiment, control group (n = 6), had ischemic ARF induced by 30 minutes of left renal pedicle clamping and right nephrectomy. SG group (n = 7) had similar surgery to the control group followed by 1 microg intraperitoneal (i.p.) loading dose of STa and placement of mini-osmotic pumps delivering 0.1 microg/hour of STa for 72 hours. AG group (n = 8) also had similar surgery to the control group followed by i.p. injection of 10 microg of ANP and placement of mini-osmotic pumps delivering 1 microg/hour of ANP for 72 hours. 72 hours post the induction of ARF, there were no statistical differences among the three groups' creatinine levels, hematocrit and body weights. ANP level was significantly higher in the AG group compared to controls, 5387.6 +/- 878.8 and 36.2 +/- 10.0 pg/mL respectively, p < 0.001. Urinary sodium (U(Na)) on the other hand reached highest levels in the SG group compared to controls 16.4 +/- 5.0 and 5.0 +/- 0.0 meq/L respectively, p = 0.028. In conclusion Sta induces significant natriuresis in ischemic ARF without changing the course of renal impairment. Likewise prolonged infusion of ANP does not alter the course of ischemic ARF. PMID- 10718281 TI - Long term treatment of IgA nephropathy with cyclosporine A. AB - 20-50% of patients with IgA nephropathy (IgAN) reach end-stage renal failure. Yet a standard treatment for those with progressive course and/or great proteinuria is lacking. We treated 6 patients with biopsy proven IgAN, proteinuria over 3.5 g/24 h and S-creatinine less than 200 micromol/L non-responding to corticosteroids administered for 3 months. They were given cyclosporine A (CsA) 5 mg/kg bw/day then titrated aiming at a serum concentration of 70-150 ng/mL for one year tapered to discontinuation in 9 months. Prednisone 5-10 mg on alternate days was given with CsA. Proteinuria (g/day) decreased from 4.66 +/- 0.43 to 1.38 +/- 0.29 (p < 0.01) after 1 month and to 0.59 +/- 0.14 (p < 0.001) after 1 year of treatment and remained lower than baseline 2 years from the beginning (1.44 +/ 0.27, p < 0.001). GFR (creatinine clearance) did not change during the first month (1.25 +/- 0.21 mL/s vs 1.38 +/- 0.29 mL/s), but decreased after 1 year (1.05 +/- 0.14 mL/s, p < 0.05). After two years it increased to 1.17 +/- 0.16, NS from baseline. We also calculated the ratio of proteinuria to the GFR (mg/L) to assess the role of hemodynamic changes in the decrease of proteinuria. This ratio was 53.80 + 6.47 before therapy, it decreased after 1 month (11.56 +/- 1.7, p < 0.05) and further after 1 year (6.78 + 1.45, p < 0.01). Three months after discontinuation it was still 14.32 +/- 1.00, p < 0.05 from baseline. In conclusion, CsA significantly lowered moderate to high proteinuria in 6 patients with IgAN. Significant decrease of the proteinuria/GFR ratio suggests some non hemodynamic mechanism of CsA action. The therapy was well tolerated and side effects were not so severe as to require CsA withdrawal. PMID- 10718282 TI - Lipoprotein profiles at different stages of chronic renal insufficiency. AB - BACKGROUND: Lipoprotein abnormalities characteristic of renal dyslipoproteinemia are significantly associated with different stages of chronic renal insufficiency. The renal dyslipoproteinemia may contribute not only to accelerated development of atherosclerosis but also to progression of human chronic renal insufficiency. METHODS: The purpose of the studies was to estimate the lipid and lipoprotein profiles in 52 not dialysed patients with various renal insufficiency advancement. Basing on creatinine level the patients were divided into 3 groups. CR1-A--serum creatinine 2-5 mg/dL (n = 16), CR1-B--serum creatinine 5-10 mg/dL (n = 19), CR1-C--serum creatinine > 10 mg/dL (n = 17). RESULTS: In CR1-A and CR1-B dyslipoproteinemia was found at different stages of renal insufficiency which was manifested by the significant increase of TG, TC, LDL-C, apo B levels and TC/HDL-C, LDL-C/HDL-C ratios and significant decrease of HDL-C level and apo AI/apoB, HDL-C/apoAI ratios in comparison with controls. We also observed decreased TG, TC, LDL-C, apo AI, apo B levels and TC/HDL-C, LDL C/HDL-C ratios and unchanged HDL-C level and apo AI/apoB, HDL-C/apoAI ratios in cm-c in comparison to CR1-A. The decrease of the lipoprotein parameters in CR1-C might result from malnutrition (statistically decreased albumin level) and metabolism disturbances connected with the renal insufficiency advancement. Negative correlation between IG, HDL-C levels (r = -0.43, p < 0.001) and TG, IIDL C/apoAI (r = -0.56; p < 0.001) were found, which confirmed the abnormal composition of HDL molecules and indicated a high risk of atherosclerosis. CONCLUSION: Our results may indicate that of atherosclerosis in CR1 patients is connected with dyslipoproteinemia and disturbances in HDL molecular composition and with different stages of chronic renal insufficiency. PMID- 10718283 TI - Effect of L-carnitine supplementation on red blood cells deformability in hemodialysis patients. AB - Anemia is a serious problem in hemodialysis patients, the main cause of which is erythropoietin deficiency. After the discovery of recombinant human erythropoietin (rHuEpo) at the end of the last decade, the hematological profile of hemodialysis patients improved significantly but at considerable expense. The deformability of red blood cells (RBC) influences their microcirculation and tissue oxygen delivery along with their life span. We investigated the deformabilty of RBCs in 15 hemodialysis patients before and after three months on L-carnitine supplementation (30 mg/Kg body wt/dialysis session). We excluded from the study all patients who received blood transfusions three months before or during the study, patients who had hemorrhagic episodes, those with hyperparathyroidism or infections, and any who required surgical intervention during the study. The serum iron, folic acid and vitamin B-12 levels were kept normal during the duration of the study. The erythropoietin dose taken before the beginning of L-cartnitine supplementation was not changed. The deformability of RBCs before and after dialysis, prior to and following three months on L carnitine was determined and compared to the deformability of RBCs from a control group. Hematocrit levels were measured before entry into the study and every month for three months. We found that the deformability of RBCs before the dialysis session was significantly greater than that found in the control group (t-test, p < 0.00001), and that there was a further increase after the end of the dialysis session. Three months following L-carnitine supplementation, we found a significant reduction of RBCs deformability (paired t-test, p < 0.004), and a significant increase in the hematocrit (ANOVA, p < 0.0001). We concluded that abnormalities in the deformability of RBCs improved after L-carnitine and that this was responsible for the increase in the hematocrit. This may allow a substantial reduction in rHuEpo dose. PMID- 10718284 TI - Non-traumatic rhabdomyolysis with acute renal failure. AB - A clinical profile of non-traumatic rhabdomyolysis with acute renal failure is presented. Myoglobinuric renal failure is treatable and hence a high index of suspicion is warranted in the etiologies discussed. PMID- 10718285 TI - Etiology, prognosis, and outcome of post-operative acute renal failure. AB - A Multivariate analysis was done in all patients who developed post operative ARF, during the period 1990-1995 to determine the etiological spectrum and to identify various variables affecting the outcome. Of 140 patients (110 operated at SGPGI and 30 operated outside) 116 underwent elective surgery. The different types of surgery leading to ARF were urosurgery (3.5%), open heart surgery (32.9%), gastrosurgery (16.4%), pancreatic surgery (9.3%), obstetrical surgery (3.6%) and others (2.8%). The incidence of ARF in SGPGI patients was highest in pancreatic surgery group (8.2%) followed by open heart surgery (3%). The different etiological factors responsible for ARF were perioperative hypotension (67.1%), sepsis (63.6%) and exposure to nephrotoxic drugs (29.3%). Sixty-four patients (45.7%) required dialysis. The overall mortality was 45%. The mortality was highest in patients who underwent open heart surgery (89.1%) followed by pancreatic surgery (84.6%). The factors associated with high mortality, other than the type of surgery, were preoperative hypotension (p < 0.05), oliguria (p < 0.01), need for dialysis (p < 0.05) and multiorgan failure (p < 0.001). AM following emergency surgery had poor outcome, though not statistically significant. Perioperative sepsis (p < 0.05) and preoperative use of aminoglycoside (p < 0.05) were significantly higher in patients operated outside SGPGI. This was associated with higher incidence of ARF. Thus we conclude that presence of multiorgan failure, oligoanuria, preoperative hypotension and need far dialysis are poor prognostic markers in ARF following surgery. PMID- 10718286 TI - Prediction of mortality in patients with bacteremia: the importance of pre existing renal insufficiency. AB - Pre-existing renal insufficiency serves as a common risk factor in the development of acute renal failure. Acute renal failure is a common finding in patients with bacteremia and is associated with poor prognosis. A total of 2722 consecutive patients 18 years old or older, fulfilling strike criteria of bacteremia or fungemia were prospectively evaluated to establish the prognostic importance of pre-existing renal insufficiency in bacteremic patients. They were classified according to serum creatinine levels upon admission into three groups. 915 patients had normal creatinine levels (< or = 1.0 mg/dL), 1528 had mild to moderate renal failure (creatinine 1.1-3 mg/dL) and 279 patients had severe renal failure upon admission (creatinine > 3.0 mg/dL). Mild to severe renal failure upon admission was associated with old age, male gender, diabetes mellitus, ischemic heat disease, hypertension and congestive heart failure. The serum albumin in patients with severe renal failure was significantly low, with a mean of 2-9 mg/dL. Urinary tract infections were more prevalent in patients with mild to severe renal failure, while intravenous line infections, bacterial endocarditis and soft and skin tissue infections were more common in patients with normal renal function. In the 279 patients with severe renal failure the mortality rate was significantly higher (50%) compared to patents with mild to moderate renal failure and patients with normal renal function (21% and 26% respectively, p = 0.0001). Multiple regression analysis revealed that pre existing serum creatinine > 3 mg/dL was significantly associated with death attributable to bacteremia (OR = 1.7, 95% CI 1.0-2.7). In conclusion, adult bacteremic patients with pre-existing serum creatinine above 3 mg/dL upon admission are at increased risk of mortality due to bacteremia than patients with normal or mild to moderate renal failure. PMID- 10718287 TI - Hair dye poisoning--a case. PMID- 10718288 TI - 'Overlearning'--vista into the nature of conscious processes. PMID- 10718289 TI - Dynorphin A-(1-13) and (2-13) improve beta-amyloid peptide-induced amnesia in mice. AB - The anti-amnesic effects of dynorphins on beta-amyloid peptide-(25-35)-induced impairment of learning and/or memory in mice were investigated using a Y-maze task and a passive avoidance test. Administration of beta-amyloid peptide-(25-35) (betaAP(25-35); 8.2 nmol, i.c.v.) 7 and 14 days before behavioral tests induced a decrease in both alternation behavior and latency in passive avoidance tests. Dynorphin A-(1-13) and A-(2-13) (0.5 and/or 1.5 nmol, i.c.v.) 30 min before behavioral tests improved the beta-amyloid peptide-(25-35)-induced impairment of alternation performance and shortened the step-down latency. Nor-binaltorphimine (4.9 nmol, i.c.v.) partially blocked the effects of dynorphin A-(1-13), but not A (2-13). These results indicate that dynorphin A-(1-13) and A-(2-13) improve amnesia induced by betaAP-(25-35) via not only kappa opioid receptors, but also by non-opioid mechanisms. PMID- 10718290 TI - c-Fos expression in the cholinergic basal forebrain after enforced wakefulness and recovery sleep. AB - The present study used c-Fos expression to examine cellular activity in cholinergic regions in the basal forebrain (BF) following enforced waking and recovery sleep. Cholinergic cells within the vertical and horizontal limbs of the diagonal band of Broca (VDB and HDB, respectively) showed significantly higher c Fos immunoreactivity after prolonged waking than after recovery sleep. Cholinergic cells within the medial septal nucleus (MS), however, showed no change in c-Fos expression under these conditions. Consistent with our previous findings, c-Fos immunoreactivity in the ventral lateral preoptic area (VLPO) was increased after 1-2h of recovery sleep compared with enforced waking. These results indicate state-specific effects on transcription and subsequent protein expression in cell populations associated with behavioral state and further show that the HDB, VDB and VLPO are good candidates for the further study of intracellular events associated with sleep and wakefulness. PMID- 10718291 TI - High frequency stimulation of the subthalamic nucleus influences striatal dopaminergic metabolism in the naive rat. AB - High frequency stimulation (HFS) of the subthalamic nucleus (STN) can partially alleviate motor symptoms in patients with Parkinson's disease (PD). However, the mechanism of action of HFS is incompletely understood. We investigated the effect of HFS (130 Hz) and low frequency stimulation (LFS, 20 Hz) of the STN on striatal dopaminergic transmission and metabolism using in vivo microdialysis in anaesthetized and freely moving rats. While LFS had no effect, HFS of the STN produced a delayed, stable and intensity-dependent increase of extracellular dopamine metabolites. Striatal extracellular levels of dopamine and 5-HIAA were not influenced by HFS or LFS in the present experimental paradigm. We conclude that HFS of the STN influences striatal dopaminergic metabolism in naive, nonlesioned rats. PMID- 10718292 TI - Brain myo-inositol level is elevated in Ts65Dn mouse and reduced after lithium treatment. AB - The segmental trisomy Ts65Dn mouse is a novel model of Down syndrome (DS). The purpose of this study was to measure brain levels of myo-inositol (ml), N acetylaspartate (NAA), and other metabolites in Ts65Dn mice using in vivo 1H magnetic resonance spectroscopy (MRS), and to determine whether lithium (Li) treatment alters brain ml level. The ratio of ml over total creatine (Cr), ml/Cr, was significantly elevated (mean change +38%), while NAA/Cr was significantly decreased (mean change -18%) in Ts65Dn mice (n=5) compared with control mice (n= 7). This is consistent with 1H MRS findings in DS human adults. Brain ml/Cr of the entire sample group (n= 12) was reduced (mean change -15%) following Li treatment, supporting the Li-induced ml depletion hypothesis. PMID- 10718293 TI - Computation of high safety factor impulse propagation at axonal branch points. AB - The propagation of single impulses at axonal branch points and widenings was computed numerically. Previous computational studies have held that an action potential propagates across an unmyelinated axonal branch point with diameter dependent lowered likelihood, such that increasingly complex arborizations could eliminate propagating information. This result is counter-intuitive to the principle of information divergence within neuronal circuits. The present study re-examined this result. The boundary conditions at a branch point were extracted from a physical analog circuit with actual branches. The main results were that impulse propagation was reliable past branch points and widenings, and that conduction velocity changed spatially as a function of fiber geometrical inhomogeneity. PMID- 10718294 TI - Developmental and tissue-specific expression of DEAD box protein p72. AB - Retinoic acid (RA) is a vitamin A derivative that has been well documented to be involved in cell differentiation. Using RNA fingerprinting by arbitrarily primed PCR, we have previously identified a number of transcripts that are regulated during RA-induced neuronal differentiation of embryonal carcinoma NT2/D1 cells. DEAD box protein p72 is one of the clones found to be down-regulated following treatment with RA. To further investigate the regulation of p72, the mRNA expression of p72 in various neuronal cell lines and primary neuronal cultures was examined. Transcripts of p72 were reduced in differentiated PC12 and IMR-32 cells but not in SH-SYSY cells. Partial cDNA fragments of p72 were isolated from rat and chick for the systematic analysis of p72 expression in different adult tissues and developmental stages. While prominent expression of p72 was observed in brain and testis, the expression was down-regulated in brain, muscle and liver during development. Taken together, our findings provide the first demonstration on the spatial and temporal expression profile of p72 in rat and chick tissues which is consistent with a role of p72 during early development. PMID- 10718295 TI - Opening of K(ATP) channels is mandatory for acquisition of ischemic tolerance by adenosine. AB - Binding of adenosine to neuronal adenosine receptors activates a signal transduction pathway (the adenosine mechanism), leading to a temporary ischemic tolerance. We have demonstrated before that induction of this mechanism in primary rat neuronal cultures, by activation of adenosine receptors, or by activation of protein kinase C (PKC), confers a wide time window of ischemic tolerance, lasting up to 72h, the early (immediate) part of which depends on opening of K(ATP) channels (glibenclamide sensitive). Here we demonstrate that the entire duration of the ischemic tolerance conferred by activation of the adenosine mechanism depends on opening of the K(ATP) channels. Thus, opening of the K(ATP) channels appears to be a mandatory step in the adenosine mechanism, leading to the creation of the wide time window of ischemic tolerance. PMID- 10718296 TI - Limits to the capacity of transplants of olfactory glia to promote axonal regrowth in the CNS. AB - Olfactory bulb ensheathing cell (OBEC) transplants promoted axonal regeneration in the spinal cord dorsal root entry zone and in the corticospinal tract. However, OBECs failed to promote abducens internuclear neuron axon regeneration when transplanted at the site of nerve fibre transection. In experiments performed in both cats and rats, OBECs survived for up to 2 months, lining themselves up along the portion of the regrowing axons proximal to the interneuron cell body. However, OBECs migrated preferentially towards abducens somata, in the direction opposite to the oculomotor nucleus target. OBECs seem to promote nerve fibre regeneration only where preferred direction of glial migration coincides with the direction of axonal growth towards its target. PMID- 10718297 TI - Circadian rhythm of serotonin levels in planarians. AB - In the freshwater planarian Dugesia japonica, which belongs to the most primitive metazoan phylum, serotonin (5-hydroxytryptamine) was detectable by both immunohistochemistry and high-performance liquid chromatography (HPLC) with fluorometric detection. Immunohistochemical studies showed that serotonin was localized primarily in the cephalic ganglion (brain), in the main nerve cords extending posteriorly from the brain and in the commissure axons connecting the main nerve cords. HPLC with fluorometric detection analysis revealed that the serotonin levels of planarians maintained under a 12:12h light:dark cycle showed significant diurnal variations with a trough in the middle of the dark phase. In constant darkness, the serotonin levels fluctuated with a circadian rhythm. These results demonstrate the existence of a circadian timekeeping mechanism in the planarian. PMID- 10718298 TI - Cluster of proliferating cells in rat vomeronasal sensory epithelium. AB - We investigated the properties of small cells in the vomeronasal sensory epithelium of adult rats. The sensory neurons in the sensory epithelium were stained by antibodies to G(i2alpha) and G(oalpha) in their cell bodies and dendrites, while the small cells, which formed a cluster in the epithelium, were not stained at all. Voltage-activated inward currents were not detected by patch clamp recordings, but outward currents were induced by the application of voltage step pulses. These results suggest that the small cells are different from the vomeronasal sensory neurons. Bromodeoxyuridine (BrdU) labeling indicated that dividing cells existed in the cluster of small cells. PMID- 10718299 TI - Role of nitric oxide in the control of the heart rate within the nucleus ambiguus of rats. AB - The aim of this study was to determine whether NO plays a role in the control of heart rate (HR) within the nucleus ambiguus (NA). Experiments were performed in 29 male Wistar rats anaesthetized with urethane. Microinjections of the NO-donor sodium nitroprusside (SNP; 5 mmol) as well as of L-arginine (L-arg; 50 mmol) into functionally identified cardioinhibitory sites within the NA significantly decreased HR (-57.7 +/- 8.4 and -53.8 +/- 3.2 bpm, respectively), whereas the NO synthase inhibitor N(omega)-nitro-L-arginine methyl ester (L-NAME) significantly increased HR (+40 +/- 2.7 bpm). Bilateral vagotomy and i.v. injection of atropine (0.5mg/kg) always abolished the HR decrease induced by SNP and L-arg, whereas propranolol did not affect the HR responses. These results demonstrated that NO mechanisms within the NA play a role in the parasympathetic control of the HR. PMID- 10718300 TI - Comparison of visual--vestibular interaction in insulin-dependent and non-insulin dependent diabetes mellitus. AB - We compared various measures of visual-vestibular interaction in subjects with insulin-dependent diabetes mellitus (IDDM) or non-insulin-dependent diabetes mellitus (NIDDM), as well as non-diabetic controls. Subjects with IDDM exhibited significantly greater postural sway than controls on those conditions in the Clinical Test of Sensory Interaction and Balance (CTSIB) which require greater reliance on the vestibular system (p < 0.005). The IDDM group also exhibited significantly worse gaze-holding in darkness and a significantly higher mean slow phase eye velocity (SPV) of optokinetic nystagmus (OKN; p<0.05 for both comparisons). However, there were no significant differences in latency to circularvection (CV). The NIDDM group showed a significant increase in postural sway across all 12 conditions compared with the controls, as well as a significant decrease in gaze-holding in darkness (p < 0.05 and p < 0.0005, respectively). However, they showed no significant difference in OKN SPV and a significant decrease in latency to CV for anticlockwise trials only (p < 0.05). These results suggest that IDDM and NIDDM are both associated with specific but different changes in visual-vestibular interaction. PMID- 10718301 TI - Effects of frequent marijuana use on brain tissue volume and composition. AB - To investigate CNS effects of frequent marijuana use, brain tissue volume and composition were measured using magnetic resonance imaging (MRI) in 18 current, frequent, young adult marijuana users and 13 comparable, non-using controls. Automated image analysis techniques were used to measure global and regional brain volumes, including, for most regions, separate measures of gray and white matter. The marijuana users showed no evidence of cerebral atrophy or global or regional changes in tissue volumes. Volumes of ventricular CSF were not higher in marijuana users than controls, but were, in fact, lower. There were no clinically significant abnormalities in any subject's MRI. Sex differences were detected in several global volume measures. PMID- 10718302 TI - Peripheral glutamate release in the hindpaw following low and high intensity sciatic stimulation. AB - The present study demonstrates that following A and/or C fiber stimulation of the sciatic nerve, glutamate levels increase significantly in the hindpaw extracellular space. In hindpaw dialysate, electrical stimulation (5 min) of the sciatic nerve at 2x, 20x, 50x or 200x threshold current required to produce a muscle twitch resulted in peak glutamate increases of 120.8 +/- 9%, 134.3 +/- 5%, 153.9 +/- 10% and 150.5 +/- 5% of basal levels, respectively. Application of 1% capsaicin to the sciatic nerve (10 min) to selectively activate C fibers resulted in a peak glutamate increase of 130.8 +/- 8% of basal levels. Aspartate levels did not change significantly in either paradigm. These data indicate that low and high intensity stimulation can result in peripheral release of glutamate, providing a major source of ligand for the glutamate receptors localized on peripheral primary afferents. PMID- 10718303 TI - L-DOPA-induced neurotoxic and apoptotic changes on cultured chromaffin cells. AB - Adrenal chromaffin cell (ACC) transplantation has been considered as one of the therapeutic strategies for Parkinson disease (PD). This strategy involves the administration of L-DOPA, although in reduced doses, to ACC-transplanted patients. Using cytochemical and morphological methods, we examined the effects of clinically applicable concentrations of L-DOPA on cultured chromaffin cells. We found an increase of cell death in both necrotic and apoptotic patterns. These data suggest that therapeutic preventive measures during ACC transplantation processes for PD should be taken. PMID- 10718304 TI - Kainic acid-induced seizures cause neuronal death in infant rats pretreated with lipopolysaccharide. AB - A major controversy in human epilepsy is whether severe seizures in infants or young children cause brain damage and subsequent epilepsy. Kainic acid (KA) produces severe seizures in infant rats, but hippocampal neuronal death and mossy fibre sprouting have not been previously demonstrated. There are similarities between lipopolysaccharide (LPS) pretreatment and KA-induced seizures in rats and the febrile convulsion of young children, in that both processes are associated with an immune stimulus and seizures. Infant rats, co-treated with LPS and KA, showed hippocampal neuronal death and mossy fibre sprouting. Taken together, our results suggest that severe febrile convulsion of young children may cause hippocampal damage and synaptic reorganization. PMID- 10718305 TI - Identification of potential compounds promoting BDNF production in nigral dopaminergic neurons: clinical implication in Parkinson's disease. AB - Parkinson's disease (PD) is characterized by the selective loss of dopamine (DA) neurons in the substantia nigral brain region. Currently, there is no cure or treatment that prevents such neuronal loss. Brain-derived neurotrophic factor (BDNF) has been found to support the survival of DA neurons in animal models and in primary cell cultures. However, the large molecular size of BDNF, coupled with the blood brain barrier, prevents its delivery to DA neurons to promote cell survival in the PD brain. The nigral DA neurons have the ability to produce BDNF for neuroprotection via either autocrine or paracrine mechanisms. Low mol. wt compounds were tested to see whether they could increase the production of BDNF in the DA neurons. The compounds tested include neurotransmitters, neuropeptides, intracellular signaling agents, known neuroprotective agents and growth factors. Our results demonstrate that salicyclic acid, cGMP analog, okadaic acid, IBMX, dipyridamole and glutamate significantly enhance BDNF production in DA neuronal cells. PMID- 10718306 TI - Galanin-(1-16) modulates 5-HT1A receptors in the ventral limbic cortex of the rat. AB - The aim of the present study was to evaluate whether galanin-(1-16) of the rat and porcine type and rat galanin-(1-29) can modulate the 5-HT1A receptors, using [3H]8-OH-DPAT as a radioligand, in membrane preparations from the ventral limbic cortex of the rat. Galanin-(1-16) produced a concentration dependent increase in the Kd value of [3H]8-OH-DPAT binding sites with a maximal effect of approximately 61% at 30 nM without changing the Bmax values. The galanin antagonist M35 blocked these effects. Rat galanin produced the same pattern of response but was less potent and effective. These results indicate the existence of a galanin receptor subtype in the ventral limbic cortex mainly recognizing N terminal galanin fragments and capable of more strongly modulating 5-HT1A receptors than cloned galanin receptors. PMID- 10718307 TI - Immature rat convulsions and long-term effects on hippocampal cholinergic neurotransmission. AB - Generalized tonic-clonic convulsions were induced on 2 consecutive days by pentylenetetrazol (PTZ) in immature rats (postnatal days 10 and 20), and hippocampal slices were prepared at different intervals post-injection. The anticholinesterase eserine provoked interictal-like discharges in the CA3 area of PTZ-injected rats (19/33), but not in controls (0/15), an effect mimicked by carbachol and reversed by atropine. This enhanced response to eserine was recorded in slices from 25-100% of the PTZ-injected rats, the percentage varying with the age at injection and post-injection interval. These results suggest that seizures in immature brain may have long-term consequences in cholinergic neurotransmission, converting a rise in endogenous ACh into an epileptogenic stimulus, which in turn would presumably facilitate the recurrence of seizures. PMID- 10718308 TI - LTP in the rat basal amygdala induced by perirhinal cortex stimulation in vivo. AB - The present study examined the effects of ventral perirhinal cortex (vPRC) stimulation on evoked field potentials (EPs) in the basal amygdala (BN), using extracellular recording techniques. Single pulse stimulation of the vPRC reliably evoked a negative field potential in the BN that was missing in the lateral nucleus. Paired-pulses delivered to the vPRC induced a short-lasting facilitation at intervals between 15 and 120 ms. Application of brief theta burst stimulation to the vPRC produced an enduring long-term potentiation (LTP) that reached 150% of control values. The induction of LTP was not accompanied by a decrease in paired-pulse facilitation. These results suggest that emotional learning involving complex stimulus representations may be mediated by cortico-amygdalar projections amenable to LTP. PMID- 10718309 TI - Alteration of the expression of the hypocretin (orexin) gene by 2-deoxyglucose in the rat lateral hypothalamic area. AB - Following an i.p. injection of 2-deoxyglucose (2DG), a nonmetabolizable analogue of glucose known to induce intracellular glucopenia, a progressive decrease in the level of hypocretin (Hcrt)/orexin mRNA was observed in the rat lateral hypothalamus while the melanin-concentrating hormone (MCH) expression in neighbouring neurons remained unaffected. This result together with the previously reported stimulation of Hcrt expression by insulin confirms that Hcrt neurons, but not MCH neurons, are sensitive to glucose availability and suggests that they respond through different mechanisms and/or different pathways to intracellular glucopenia and hypoglycemic conditions. PMID- 10718310 TI - An IP3-assisted form of Ca2+-induced Ca2+ release in neocortical neurons. AB - Calcium increases induced by single action potentials in rat visual cortex layer II/III pyramidal neurons were shown to be augmented by muscarinic acetylcholine receptor (mAchR) stimulation. This augmentation was drastically reduced by intracellular injection of heparin but not ruthenium red, therefore involving inositol-1,4,5-trisphosphate (IP3)-sensitive rather than ryanodine-sensitive calcium stores. Only the calcium increase induced by the second or later spike of a spike train, but not that induced by the first spike, was augmented, indicating the requirement of both spike-induced calcium increase and mAchR activation. The calcium store depletor thapsigargin abolished this augmentation use-dependently. These findings suggest a neocortical occurrence of calcium-induced calcium release from IP3-sensitive calcium stores that have been sensitized beforehand by IP3 through mAchR-mediated mechanisms. PMID- 10718311 TI - Propionic and L-methylmalonic acids induce oxidative stress in brain of young rats. AB - The in vitro effects of propionic and L-methylmalonic acids on some parameters of oxidative stress were investigated in the cerebral cortex of 21-day-old rats. Chemiluminescence, thiobarbituric acid-reactive substances (TBA-RS) and total radical-trapping antioxidant capacity (TRAP) were measured in brain tissue homogenates in the presence of propionic or L-methylmalonic acids at concentrations ranging from 1 to 10mM. Both acids significantly increased chemiluminescence and TBA-RS and decreased TRAP, indicating a simulation of lipid peroxidation and a reduction of tissue antioxidant potential. Other organic acids tested which accumulate in some organic acidemias (suberic, sebacic, adipic, 3 methylglutaric and 4-hydroxybutyric acids) did not affect these parameters. This study provides evidence that free radical generation may participate in the neurological dysfunction of propionic and methylmalonic acidemias. PMID- 10718312 TI - Brain functional reorganization in early blind humans revealed by auditory event related potentials. AB - Visually challenged individuals often compensate for their handicap by developing supra-normal abilities in their remaining sensory systems. Here, we examined the scalp distribution of components N1 and P3 of auditory evoked potentials during a sound localization task in four totally blind subjects who had previously shown better performance than sighted subjects. Both N1 and P3 waves peaked at their usual positions while blind and sighted individuals performed the task. However, in blind subjects these two components were also found to be robust over occipital regions while in sighted individuals this pattern was not seen. We conclude that deafferented posterior visual areas in blind individuals are recruited to carry out auditory functions, enabling these individuals to compensate for their lack of vision. PMID- 10718313 TI - Effect of atypical antipsychotics on phencyclidine-induced expression of arc in rat brain. AB - The effect of atypical antipsychotics on the immediate-early gene, arc (activity regulated cytoskeleton-associated gene), expression was investigated in phencyclidine (PCP)-treated rats using RT-PCR. Administration of PCP (10 mg/kg) increased arc mRNA levels in the prefrontal cortex, nucleus accumbens and posterior cingulate cortex. Pretreatment with clozapine (20 mg/kg), olanzapine (10 mg/kg) and risperidone (2 mg/kg), but not haloperidol (2 mg/kg), prevented PCP-induced arc expression in the prefrontal cortex and nucleus accumbens. Pretreatment of haloperidol increased the striatal arc mRNA levels. Clozapine, olanzapine and haloperidol inhibited the PCP-induced arc expression in the posterior cingulate cortex. These results suggest that the effects of antipsychotic drugs on PCP-induced arc expression in the prefrontal cortex and nucleus accumbens are useful for distinguishing atypical antipsychotic properties of the drugs. PMID- 10718314 TI - Does frontal lobe activation during retrieval reflect complexity of retrieved information? AB - Neuroimaging studies of memory have consistently shown that episodic retrieval is associated with right frontal activation, whereas semantic retrieval is associated with left frontal activation. Various hypotheses have been proposed to account for this lateralization in terms of underlying psychological processes. Alternatively, this lateralization may reflect the complexity of information retrieved: retrieval of complex, contextual information accompanying episodic retrieval invokes right-lateralized processes preferentially. We tested this hypothesis by manipulating the type and complexity of information retrieved. Initial increase in complexity of both episodic and semantic information was associated with right inferior frontal activation; further increase in complexity was associated with left dorsolateral activation. We conclude that frontal activation during retrieval is a non-linear function of the complexity of retrieved information. PMID- 10718315 TI - Interacting biological and electronic neurons generate realistic oscillatory rhythms. AB - Small assemblies of neurons such as central pattern generators (CPG) are known to express regular oscillatory firing patterns comprising bursts of action potentials. In contrast, individual CPG neurons isolated from the remainder of the network can generate irregular firing patterns. In our study of cooperative behavior in CPGs we developed an analog electronic neuron (EN) that reproduces firing patterns observed in lobster pyloric CPG neurons. Using a tuneable artificial synapse we connected the EN bidirectionally to neurons of this CPG. We found that the periodic bursting oscillation of this mixed assembly depends on the strength and sign of the electrical coupling. Working with identified, isolated pyloric CPG neurons whose network rhythms were impaired, the EN/biological network restored the characteristic CPG rhythm both when the EN oscillations are regular and when they are irregular. PMID- 10718316 TI - Time course of c-FOS expression in status epilepticus induced by amygdaloid stimulation. AB - Decaying of c-FOS immunoreactivity (FIR) was studied in adult rats with 1 h self sustaining limbic status epilepticus (SSLSE) induced by amygdaloid electrical stimulation. Rats that failed to enter SSLSE showed localized FIR in the ipsilateral limbic cortex, neocortex, and amygdala. FIR became bilaterally extensive, including the hippocampal formation 0.5 h after SSLSE. It decreased gradually between 2 and 6 h and returned to basal levels around 1 day. Neocortical FIR in clonic SSLSE persisted longer than in other types of SSLSE. We demonstrate for the first time that FIR in SSLSE lasts much longer than several hours, its decaying is related to the seizure behavior, and absent or weak FIR at the hippocampal formation is associated with failed SSLSE entry. PMID- 10718317 TI - Neocortical ectopias are associated with attenuated neurophysiological responses to rapidly changing auditory stimuli. AB - Developmental dyslexia has been separately associated with the presence of ectopic collections of neurons in layer I of neocortex (ectopias) and with alterations in processing rapidly changing stimuli. We have used BXSB/MpJ-Yaa mice, some of which have neocortical ectopias, to directly test the hypothesis that ectopias may alter auditory processing. Auditory event related potentials (AERPs) were elicited by pairs of 10.5 kHz tones separated by silence, 0.99 kHz, or 5.6 kHz tones of variable duration. Half of the mice tested had 1-3 ectopias in frontal or parietal cortex, and half had no ectopias. Mice with ectopias showed a reduced response to the second 10.5 kHz stimuli only when it was preceded by short duration 5.6 kHz tones. These results indicate that BXSB mice are an excellent model for determining how focal neocortical anomalies alter sensory processing. PMID- 10718318 TI - Co-localization of patched and activated sonic hedgehog to lysosomes in neurons. AB - We demonstrate co-localization of the Patched 1 (Ptc1) receptor and its ligand sonic hedgehog (Shh) in lysosomes suggests an intracellular sorting mechanism for this receptor and its ligand. Treatment of murine brain primary cultures and a human teratoma cell line with the N-terminal activated form of Shh (ShhNT), a Ptc1-Shh complex was observed in lysosomes. Consistent with this interaction, Western immunoblot analysis revealed intracellular localization of native Ptc1 and ShhNT. Examination of the topological model of the Ptc1 receptor revealed a number of Yxxphi lysosomal targeting sequences consistent with our observations for Ptc1 sorting. PMID- 10718319 TI - The development of the nociceptive responses in neurokinin-1 receptor knockout mice. AB - An important yet unanswered question is how neonates respond to painful stimuli, given the immaturity of their neural pathways. We examined the development of the neurokinin system using a novel approach, examining changes of this system by observing the pain responses of mice lacking the NK1 receptor at different stages of development We show that the NK1 receptor is not involved in nociception to heat, mechanical or chemical stimuli, at 3 days. In contrast, the NK1 receptor is involved in nociceptive responses to high intensity heat and mechanical stimuli, and mediates the second phase of the formalin response in 21-day-old mice. This indicates that nociception in neonates does not require the NK1 receptor and that the functional maturation of the NK1 receptor allows diversity in both the type of stimuli that activate the pain system and the types of responses elicited by nociceptive stimuli. PMID- 10718320 TI - Expression and regulation of CNTF receptor-alpha in the in situ and in oculo grafted adult rat adrenal medulla. AB - Cultured and transplanted adrenal medullary cells respond to ciliary neurotrophic factor (CNTF) with neurite formation and improved cell survival although the presence of the CNTF receptor-alpha (CNTFRalpha) has been unclear. This study show that CNTFRalpha mRNA was expressed in the postnatal day 1 as well as in the adult rat adrenal medulla. The highest CNTFRalpha mRNA signal was found in the ganglion cells of the adrenal medulla. After transplantation of adrenal medullary tissue the CNTFRalpha mRNA levels were down-regulated in the chromaffin cells. CNTF treatment of grafts did not normalize the receptor levels, but treatment with nerve growth factor (NGF) did. Thus, we demonstrate that CNTFRalpha mRNA is expressed in adrenal medulla, the levels becomes down-regulated after transplantation, but normalized after treatment with NGF. PMID- 10718321 TI - Transgenic expression of TNF by astrocytes increases mechanical allodynia in a mouse neuropathy model. AB - It has been hypothesized that increased expression of proinflammatory cytokines mediate a variety of central nervous system disorders such as multiple sclerosis, Alzheimer's disease, cerebral ischemia, spinal cord injury, HIV encephalopathy and chronic pain. In order to further examine the central role of TNF in neuropathic pain, transgenic mice were used in which expression of murine TNF was targeted to astrocytes using a glial fibrillary acidic protein (GFAP)-TNF fusion gene. Spinal nerve (L5) transection was performed in either the GFAP-TNF transgenic or wild type mice. Mechanical allodynia was significantly enhanced in the GFAP-TNF transgenic mice compared with the wild type mice. These data support a central role of glial expression of TNF in the generation of neuropathic pain. PMID- 10718322 TI - Behavioral deficits in APP(V717F) transgenic mice deficient for the apolipoprotein E gene. AB - Both the beta-amyloid precursor protein (APP) and the apoliprotein E (apoE) genes are involved in the pathogenesis of Alzheimer's disease (AD). We previously showed that mice over-expressing a human mutated form of APP (APP(V717F)) display age-dependent recognition memory deficits associated with the progression of amyloid deposition. Here, we asked whether 10- to 12-month-old APP(V717F) mice lacking the apoE gene, which do not present obvious amyloid deposition, differ from APP(V717F) mice in the object recognition task. The recognition performance is decreased in both transgenic mouse groups compared to control groups. Moreover, some behavioral disturbances displayed by APP mice lacking apoE are even more pronounced than those of APP mice expressing apoE. Our results suggest that the recognition memory deficits are related to high levels of soluble Abeta rather than to amyloid deposits. PMID- 10718323 TI - Cerebellar volume in humans related to magnitude of classical conditioning. AB - Neural circuits in the cerebellum are essential for eyeblink classical conditioning, and hippocampal activation is also present during acquisition. Anatomical (volumetric) brain MRI, delay eyeblink conditioning and neuropsychological tests were administered to eight healthy older subjects. The correlation between cerebellar volume (corrected for total cerebral volume) and conditioned response percentage was 0.81 (p < 0.02), but neither hippocampal nor total cerebral volume correlated with conditioning or any neuropsychological test scores. There was no relationship between age and cerebellar volume, but the correlation between hippocampal volume and age was -0.80 (p < 0.02). These volumetric results add to the increasing evidence in humans demonstrating a relationship between the integrity of the cerebellum and eyeblink classical conditioning. PMID- 10718324 TI - Picture naming without Broca's and Wernicke's area. AB - Lexical and semantic retrieval was investigated in normal volunteers with PET by comparing picture confrontation naming and verb generation related to the same pictures. Conjunction analysis of the naming and verb generation uncovered a common network including the occipito-temporal ventral pathway for object recognition, and the bilateral anterior insula, SMA and precentral gyrus for coordination, planning and overt word production. Naming and verb generation highlighted two different patterns: verb generation showed specific implication of Broca and Wernicke's areas, whereas naming specifically relied on the primary visual areas, the right fusiform and parahippocampal gyri and the left anterior temporal region. These results indicate that speech does not necessarily involve the Wernicke-Broca's language network and testify that naming relies on an early developmental language network. PMID- 10718325 TI - Reversible cisplatin ototoxicity in the albino guinea pig. AB - Guinea pigs implanted with round window electrodes received daily doses (2.0 mg/kg) of cisplatin until a profound hearing loss occurred (> 40 dB at 8 kHz). Afterwards, pronounced recovery occurred. Recovery progressed over intervals up to 3 weeks before it saturated. Loss and recovery involved both the compound action potential and, less pronounced, the cochlear microphonics. Cochlear potentials evoked by lower frequencies recovered more fully than those evoked by higher frequencies. Loss and recovery was found also in the endocochlear potential. Outer hair cell counts did not change over the recovery period. These findings confirm our previously reported results on the reversibility of cisplatin damage. Further, they implicate the vascular stria as an important target for cisplatin in the cochlea. PMID- 10718326 TI - Sleep in the Wistar-Kyoto rat, a putative genetic animal model for depression. AB - The Wistar-Kyoto (WKY) rat exhibits several behavioral and hormonal abnormalities often associated with depression. One of the hallmarks of depression consists of alterations in the sleep-wake cycle, particularly in rapid eye movement (REM) sleep. If the WKY rat is indeed an animal model for depression, we hypothesized that it should also show sleep abnormalities relative to the control strain, the Wistar (WIS) rat Under baseline conditions, WKY rats showed a 50% increase in total REM sleep time during the 12 h light phase and an increase in sleep fragmentation during both the light and dark phase. The WKY rats also exhibited lower EEG power densities over the entire frequency range (0.2-25.0 Hz) during REM sleep. After a 6 h sleep deprivation, the REM sleep rebound was more pronounced during the dark but not the light phase in the WKY rats. Since the WKY rat represents a genetic model for depression with altered EEG sleep patterns, this strain may be particularly useful for investigating the relationship between depression and sleep abnormalities. PMID- 10718327 TI - Functional brain imaging of tinnitus-like perception induced by aversive auditory stimuli. AB - Tinnitus is an aversive auditory percept of unknown origin. We tested the speculation that tinnitus may share neuronal processing mechanisms with aversive auditory percepts of known origin. This study revealed the functional neuroanatomy of the perception of aversive auditory stimuli. The stimuli were presented to 12 healthy volunteers so as to mimic the psychoacoustical features of tinnitus and its affective response in tinnitus sufferers. The regional cerebral blood flow distribution was measured by PET during four auditory processing conditions and one control condition. The aversive auditory stimuli activated primary and secondary auditory areas bilaterally, dorsolateral prefrontal attention areas, and structures in the limbic system which subserve emotional processing. Based on these results and findings from other functional neuroimages of tinnitus, we hypothesize that the perception of tinnitus may involve the functional linkage of these brain areas: secondary auditory cortex, dorsolateral prefrontal cortex, and limbic system. PMID- 10718328 TI - Genetic factors modulate effects of C-section birth on dopaminergic function in the rat. AB - Genetic predisposition and environmental factors such as perinatal complications are believed to contribute to the etiology of schizophrenia, a disorder involving enhanced CNS dopaminergic activity. This study used a rat model to test whether genetic factors and a minor birth complication, i.e. Caesarean section (C section) birth, interact in producing longterm effects on dopamine-mediated behavior. For this, we compared the effects of vaginal and C-section birth on amphetamine (AMPT)-induced locomotor activity in strains of rats differing in genetic composition. In Sprague-Dawley rats, C-section birth increased AMPT induced locomotion compared with vaginal birth. By contrast in Lewis rats, C section birth reduced AMPT-induced locomotion compared with vaginal birth. In Fischer rats, AMPT-induced locomotion was increased by C-section under maternal anesthesia but decreased by C-section after maternal decapitation, compared with vaginal birth. It is concluded that a minor birth complication like C-section can have differing long-term effects on dopaminergic function in the rat, depending on the genetic composition of the individual. PMID- 10718329 TI - Which nerve fibers mediate the axon reflex flare in human skin? AB - Axon reflex vasodilatation due to transcutaneous electrical stimulation in human skin was measured by laser Doppler imaging. Constant current pulses of 10 mA, 0.2 ms, delivered at 1 or 10 Hz for 2 min through a probe of 30 mm2 surface area did not induce a significant flare response, though this stimulus previously has been found supra-maximal for cutaneous polymodal (mechano-heat responsive) C nociceptors in microneurography experiments. Pulses of the same strength from a pointed probe yielding a higher current density induced an extended and persistent flare. This type of stimulus previously has been proven to recruit mechano-insensitive C-units in microneurography experiments, in contrast to stimuli from the 30 mm2 probe. It is concluded that mechano-insensitive C nociceptors and not polymodal C-units mediate the axon reflex flare in human skin. PMID- 10718330 TI - Increased sensitivity to ET-1 in rat cerebral arteries following organ culture. AB - Endothelin-1 (ET-1) is recognized as being involved in the pathophysiology of cerebrovascular diseases. Using organ culture as a model for possible pathological changes we studied changes in ET(A) and ETB receptor function using a sensitive in vitro method. We observed an up-regulation of the ET(B) receptor and an amazingly increased sensitivity to ET-1 by 3 log units in pEC50; pEC50(fresh) was 8.7 +/- 0.1, and pEC50(cultured) was 11.7 +/- 0.3. pA2 for FR139317 in the fresh vessel was 7.0 +/- 0.2 whereas it could not be obtained for the cultured vessel, indicating a possible cross-talk between the ET(A) and ET(B) receptors. The increased sensitivity to ET-1 could also take place during cerebrovascular disease such as stroke or haemorrhage rendering the vessels considerably more sensitive to ET-1. PMID- 10718331 TI - Texaphyrins: new drugs with diverse clinical applications in radiation and photodynamic therapy. AB - The texaphyrins are quintessential metal-coordinating expanded porphyrins. They constitute a new series of synthetic porphyrin analogues that show promise as drugs for use in a range of medical therapies. Currently, two different water solubilized lanthanide(III) texaphyrin complexes, namely the gadolinium(III) and lutetium(III) derivatives 1 and 2 (Gd-Tex and Lu-Tex, respectively), are being tested clinically. The first of these, XCYTRIN, is in a pivotal Phase III clinical trial as a potential enhancer of radiation therapy for patients with metastatic cancers to the brain receiving whole brain radiation therapy. The second, in various formulations, is being tested as a photosensitizer for use in: (i) the photodynamic treatment of recurrent breast cancer (LUTRIN; Phase II clinical trials complete), (ii) photoangioplastic reduction of atherosclerosis involving peripheral arteries (ANTRIN; now in Phase II testing), and (iii) light based treatment of age-related macular degeneration (OPTRIN; currently in Phase I clinical trials), a vision-threatening disease of the retina. Taken in concert, these two metallotexaphyrins provide a powerful new class of experimental drugs whose diverse potential utility is abetted by a combination of well-optimized physical features, favorable tissue biolocalization characteristics, and novel mechanisms of action. Interestingly, these mechanisms may alter conventional wisdom regarding mechanisms of radiation therapy and the pathophysiology of atherosclerosis. PMID- 10718332 TI - 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD)-induced changes in activities of nuclear protein kinases and phosphatases affecting DNA binding activity of c-Myc and AP-1 in the livers of guinea pigs. AB - To study the effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on nuclear protein phosphorylation activities, male guinea pigs were treated in vivo with a single 1 microg/kg i.p. injection of TCDD, and the state of protein kinases and phosphatases in the nuclei of the hepatocytes was examined after 1, 10, and 40 days. TCDD was found to cause a rise in nuclear protein tyrosine kinase on day 1, and to a lesser extent on day 10, but this effect diminished almost completely on day 40. TCDD also caused a reduction in nuclear casein kinase II (CKII) activity at all time points. To study the biochemical events taking place at the early stage of the action of TCDD, a short-term in vitro model system was established using explant liver tissues maintained in tissue culture medium. It was found that TCDD caused a rapid reduction of the activity of nuclear CKII with an accompanying increase in the cytosol. Such changes in protein phosphorylation activities were also accompanied by an increase in the DNA binding activity of activator protein 1 (AP-1). The effect of TCDD on nuclear proteins binding to the c-Myc response element DNA was, on the other hand, biphasic: an initial increase of protein binding to the c-Myc response element was followed by suppression. To test the hypothesis that some of the above changes were caused by TCDD-induced changes in protein kinase activity, nuclear proteins isolated from hepatocytes of in vivo treated guinea pigs were incubated with exogenously added Mg2+ and ATP under cell-free conditions. The results showed that this in vitro phosphorylation treatment exacerbated this tendency of increased AP-1 and decreased c-Myc binding to their respective response element DNAs, indicating that kinases and phosphatases present in the isolated nuclear protein preparation were active and capable of modifying protein binding to DNA. Such effects of Mg2+ and ATP on AP-1 were blocked by heparin, indicating that CKII plays an important role in transducing the signal of TCDD into the nucleus. PMID- 10718333 TI - Anti-stress effects of dehydroepiandrosterone: protection of rats against repeated immobilization stress-induced weight loss, glucocorticoid receptor production, and lipid peroxidation. AB - In the present study, we have (i) examined the biological effects of repeated immobilization stress, and (ii) tested the hypothesis that the adrenal steroid hormone dehydroepiandrosterone (DHEA) is an anti-stress hormone, using male Sprague-Dawley rats. Rats (N = 6) were divided into the following four groups: (i) control, (ii) repeated immobilization stress (2 hr daily, for 60 days), (iii) repeated immobilization stress (2 hr daily, for 60 days) plus daily i.p. administration of 5 mg DHEA/0.1 mL DMSO, and (i.v.) daily i.p. administration of 5 mg DHEA/0.1 mL DMSO alone. Results obtained showed that repeated immobilization stress resulted in a significant (25%) inhibition in body weight gain, a significant increase in adrenal weight, an increase in glucocorticoid receptor (GR) in the liver, thymus, and spleen, decreased plasma triglyceride levels, and increased lipid peroxidation in the liver and heart as compared with control unstressed animals. Interestingly, DHEA administration resulted in a significant reversal in stress-induced inhibition in body weight gain, adrenal weight, GR levels in liver, thymus, and spleen, and lipid peroxidation levels in the liver and heart. In addition, animals treated with DHEA alone without stress showed a significant (15%) inhibition in body weight gain and an almost 60% decrease in plasma triglyceride levels as compared with control unstressed animals. It is concluded that DHEA acts as an anti-stress hormone in rats, as shown in its antagonizing the effects of repeated immobilization stress on total body weight, adrenal weight, GR levels, and free radical generation. PMID- 10718334 TI - Selectivity of the diacylglycerol kinase inhibitor 3-[2-(4-[bis-(4 fluorophenyl)methylene]-1-piperidinyl)ethyl]-2, 3-dihydro-2-thioxo 4(1H)quinazolinone (R59949) among diacylglycerol kinase subtypes. AB - Diacylglycerol kinases (DGKs) attenuate diacylglycerol-induced protein kinase C activation during stimulated phosphatidylinositol turnover. This reaction also initiates phosphatidylinositol resynthesis. Two agents, 3-(2-(4-[bis-(4 fluorophenyl)methylene]-1-piperidinyl)ethyl)-2,3-dihydro -2-thioxo 4(1H)quinazolinone (R59949) and 6-(2-(4-[(4-fluorophenyl)phenylmethylene]-1 piperidinyl)ethyl)-7-m ethyl-5H-thiazolo(3,2-a)pyrimidin-5-one (R59022), inhibit diacylglycerol phosphorylation in several systems. To examine the mechanism of this effect, we developed a mixed micelle method suitable for in vitro study of DGK inhibition. Animal cells express multiple DGK isoforms. In a survey of DGK isotypes, these agents selectively inhibited Ca2+-activated DGKs. R59949 was the more selective of the two. To map the site of interaction with the enzyme, a series of DGKalpha deletion mutants were prepared and examined. Deletion of the Ca2+-binding EF hand motif, which is shared by Ca2+-activated DGKs, had no effect on inhibition. Consistent with this observation, inhibition kinetics were noncompetitive with Ca2+. A construct expressing only the catalytic domain was also inhibited by R59949. Studies of substrate kinetics demonstrated that MgATP potentiated R59949 inhibition, indicating synergy of inhibitor and MgATP binding. These results indicate that R59949 inhibits DGKalpha by binding to its catalytic domain. PMID- 10718335 TI - Isolation and characterization of a cDNA encoding a horse liver butyrylcholinesterase: evidence for CPT-11 drug activation. AB - Butyrylcholinesterases (BuChEs; acylcholine acylhydrolase; EC 3.1.1.8) have been demonstrated to convert the anticancer agent CPT-11 (irinotecan, 7-ethyl-10-[4-(1 piperidino)-1-piperidino]carbonyloxycamptothecin) into its active metabolite SN 38 (7-ethyl-10-hydroxycamptothecin). In addition, significant differences in the extent of drug metabolism have been observed with BuChEs derived from different species. In an attempt to understand these differences, we have isolated the cDNA encoding a horse BuChE. Based upon the NH2-terminal amino acid sequence of a purified horse BuChE, we designed degenerate primers to amplify the coding sequence from horse liver cDNA. Following polymerase chain reaction and rapid amplification of the cDNA ends, we generated an 1850-bp DNA fragment, containing an 1806-bp open reading frame. The cDNA encodes a protein of 602 amino acid residues, including a 28-amino-acid NH2-terminal signal peptide. Furthermore, the DNA sequence and the deduced amino acid sequence revealed extensive homology to butyrylcholinesterase genes from several other species. In vitro transcription translation of the cDNA produced a 66-kDa protein, identical to the size of native horse serum BuChE following removal of carbohydrate residues with endoglycosidase F. Additionally, transient expression of the cDNA in Cos-7 cells yielded extracts that exhibited cholinesterase activity and demonstrated a Km value for butyrylthiocholine of 106+/-9 nM. This extract converted the anticancer drug CPT-11 into SN-38, demonstrating that this drug can be activated by enzymes other than carboxylesterases. PMID- 10718336 TI - The expression and secretion of atrial natriuretic factor and brain natriuretic peptide by rat proximal tubular cells. AB - We examined the expression of both the natriuretic peptides and natriuretic peptide receptors (NPR) in primary cultures of rat proximal tubular (RPT) cells using Northern blot assay for peptides and receptors and radioimmunoassay and immunohistochemical analysis for atrial natriuretic factor (ANF), brain natriuretic peptide (BNP), and C-type natriuretic peptide. Freshly isolated cells expressed mRNA coding for ANF, BNP, and the NPR-C. The presence of ANF and BNP in freshly isolated cells was confirmed by immunocytochemical staining. As cells approached confluence, there was a marked increase in mRNA expression for ANF and BNP. Immunocytochemical analysis and radioimmunoassay confirmed that both these peptides were co-localised in RPT cells and present in the cell supernatant. These changes in peptide expression were associated with a concurrent decrease in the expression of the NPR-C and the appearance of the NPR-A and -B. These results confirm that freshly isolated RPT cells possess the components of an autocrine natriuretic peptide system and that growth in primary culture is associated with changes in both peptide system and that growth in primary culture is associated with changes in both peptide and receptor subtype expression, raising the possibility that the endogenous production of ANF and BNP may be involved in the control of control cell growth. PMID- 10718337 TI - A chimeric rat brain P2Y1 receptor tagged with green-fluorescent protein: high affinity ligand recognition of adenosine diphosphates and triphosphates and selectivity identical to that of the wild-type receptor. AB - We tested how the green fluorescent protein (GFP) tag affects signaling of the nucleotide-activated P2Y1 receptor. Therefore, we generated stably transfected human embryonic kidney 293 cells expressing the rat P2Y1 wild-type receptor (rP2Y1-wt) or the receptor tagged at the C-terminus with the enhanced GFP (rP2Y1 eGFP). The chimeric rP2Y1-eGFP receptor is localized mainly to the plasma membrane as revealed by Western blotting of subcellular fractions. Both receptors were analyzed by measuring Ca2+ responses to short pulses of the agonists in single cells by continuous superfusion with medium. The rP2Y1-eGFP receptor was coupled to Ca2+ release as was the rP2Y1-wt receptor. 2-Methylthio adenosine 5' diphosphate and -triphosphate (2-MeSATP and 2-MeSADP) were the most potent agonists at the heterologously expressed receptors, with EC50 values of 50 to 70 nM for rP2Y1-eGFP and 0.06 to 0.4 nM for rP2Y1-wt. These potencies of the two P2Y selective agonists at rP2Y1-wt receptor-expressing cells are the highest values reported so far. This increase is probably due to a receptor reserve. In both rP2Y1-wt- and in rP2Y1-eGFP-expressing cells, the effect of 2-MeSATP was inhibited equally by the antagonist pyridoxal phosphate-6-azophenyl-2',4' disulfonic acid. We established that ATP as well as adenosine 5'-O-(1 thiotriphosphate) (ATPalphaS) are full agonists at the rP2Y1 receptor at both transfected cell lines. The rP2Y1-eGFP receptor has the same ligand selectivity as the rP2Y1-wt receptor (2-MeSADP = 2-MeSATP > ADP > ATPalphaS, ATP >> UTP). Thus, the GFP-tagged P2Y1 receptor is fully active and shows regular signal transduction coupling. It provides the means for biochemical characterization, since it can be solubilized and is a tool for further physiological analysis. PMID- 10718338 TI - Beta-glucuronidase latency in isolated murine hepatocytes. AB - The physiological function of microsomal beta-glucuronidase is unclear. Substrates may be either glucuronides produced in the lumen of endoplasmic reticulum (ER) or those taken up by hepatocytes. In the latter case, efficient inward transport of glucuronides at the plasma membrane and the ER membrane would be required. Therefore, the potential role of beta-glucuronidase in ER was investigated. Isolated mouse hepatocytes and mouse and rat liver microsomal vesicles were used in the experiments. Selective permeabilization of the plasma membrane of isolated hepatocytes with saponin or digitonin resulted in an almost 4-fold elevation in the rate of beta-nitrophenol glucuronide hydrolysis, while the permeabilization of plasma membrane plus ER membrane by Triton X-100 caused a further 2-fold elevation. In microsomal vesicles, the p-nitrophenol glucuronide or phenolphthalein glucuronide beta-glucuronidase activity showed about 50% latency as revealed by alamethicin or Triton X-100 treatment. A light-scattering study indicated that the microsomes are relatively impermeable to both glucuronides and to glucuronate. On the basis of our results, the role of liver microsomal beta-glucuronidase in the deconjugation of glucuronides taken up by the liver seems unlikely. Hydrolysis of the glucuronides produced in the ER lumen may play a role in substrate supply for ascorbate synthesis or in "proofreading" of glucuronidation. PMID- 10718339 TI - F 11782, a novel epipodophylloid non-intercalating dual catalytic inhibitor of topoisomerases I and II with an original mechanism of action. AB - F 11782, a novel epipodophylloid, proved a potent inhibitor of the catalytic activities of both topoisomerases I and II. Unlike classical inhibitors such as camptothecin or etoposide, F 11782 did not stabilise cleavable complexes induced by either topoisomerases I or II nor did it preferentially inhibit the religation step of the catalytic cycle of either enzyme. F 11782 neither intercalated DNA nor bound in its minor groove, and showed only weak inhibition of the ATPase activity associated with topoisomerase II. F 11782 appeared to act by inhibiting the binding of topoisomerases I and II to DNA in a manner dependent both on drug and enzyme concentrations, via a mechanism not previously described or shared by other known topoisomerase 'poisons' or inhibitors. In contrast, F 11782 had only a weak effect or none at all on various other DNA-interacting enzymes. In conclusion, F 11782, as a non-intercalating, specific catalytic inhibitor of both topoisomerases I and II with an original mechanism of action, may be considered to represent the first of a new class of topoisomerase-interacting agents. PMID- 10718340 TI - N-butyldeoxygalactonojirimycin: a more selective inhibitor of glycosphingolipid biosynthesis than N-butyldeoxynojirimycin, in vitro and in vivo. AB - N-Butyldeoxynojirimycin (NB-DNJ) inhibits the ceramide glucosyltransferase which catalyses the first step in glycosphingolipid (GSL) biosynthesis. It has the potential to be used for the treatment of the GSL lysosomal storage diseases and is currently in clinical trials for the treatment of type 1 Gaucher's disease. However, NB-DNJ is also a potent inhibitor of other enzymes, including alpha glucosidase I and II, which could potentially cause side effects in patients receiving life-long therapy. Wetherefore evaluated a potentially more selective GSL biosynthesis inhibitor, N-butyldeoxygalactonojirimycin (NB-DGJ), in vitro and in vivo. The distribution and degree of GSL depletion in the liver of mice treated with NB-DGJ or NB-DNJ were equivalent. Mice treated with NB-DGJ had normal body weights and lymphoid organ sizes, whereas NB-DNJ-treated mice showed weight loss and partial lymphoid organ shrinkage. NB-DNJ inhibited glycogen catabolism in the liver, whereas NB-DGJ did not. NB-DNJ was also a potent inhibitor of sucrase and maltase in vitro but not of lactase, while NB-DGJ inhibited lactase but not sucrase or maltase. NB-DGJ is therefore more selective than NB-DNJ, and deserves to be evaluated for human therapy. PMID- 10718341 TI - Pharmacological properties of a new aziridinylbenzoquinone, RH1 (2,5-diaziridinyl 3-(hydroxymethyl)-6-methyl-1,4-benzoquinone), in mice. AB - RH1 (2,5-diaziridinyl-3-(hydroxylmethyl)-6-methyl-1,4-benzoquinone) has shown preferential activity against human tumour cell lines which express high levels of DTD (EC 1.6.99.2; NAD(P)H:quinone oxidoreductase, NQO1, DT-diaphorase) and is a candidate for clinical trials. EO9 (3-hydroxy-5-aziridinyl-1-methyl-2-[1H indole-4,7-dione]prop-beta-en-alpha-ol) is a known substrate for DTD but clinical trials were disappointing, as a result of rapid plasma clearance and reversible dose-limiting kidney toxicity. It is an obvious concern that RH1 does not exhibit the same limitations. We therefore describe the antitumour activity and pharmacology of RH1 in mice and compare its pharmacological characteristics to those of EO9. Significant antitumour activity (P = 0.01) was seen for RH1 (0.5 mg/kg, i.p.) against the high DTD-expressing H460 human lung carcinoma. Pharmacokinetic analysis of RH1 in mice showed a t1/2 of 23 min with an area under the curve of 43.0 ng hr mL(-1) resulting in a calculated clearance of 5.1 mL min(-1), 10-fold slower than EO9. RH1 was also more stable than EO9 in murine blood, where the breakdown was thought to be DTD-related. NADH-dependent microsomal metabolism of RH1 and EO9 in both liver and kidney was slow (<100 pmol/min/g tissue), reflecting the low microsomal DTD expression (<35 nmol/mg/min). Liver cytosol metabolism was rapid for both compounds (>4500 pmol/min/g tissue), although DTD levels were low (21.4+/-0.6 nmol/mg/min). DTD activity in the kidney cytosol was high (125+/-8.2 nmol/mg/min) and EO9 was rapidly metabolised (4396+/-1678 pmol/min/g), but the metabolic rate for RH1 was 7-fold slower (608+/-86 pmol/min/g), even though RH1 was shown to be an excellent substrate for DTD (Vmax = 800 micromol/min/mg and a Km of 11.8 microM). The two DTD substrates RH1 and EO9 are clearly metabolised differently, suggesting that RH1 may have different pharmacological properties to those of EO9 in the clinic. PMID- 10718342 TI - Involvement of Hg2+-sensitive sulfhydryl groups in regulating noradrenaline release induced by S-nitrosocysteine in rat brain slices. AB - Nitric oxide has been shown to regulate neurotransmitter release. Previously, we reported that S-nitrosothiols such as S-nitrosocysteine (SNC) stimulate noradrenaline (NA) release in rat hippocampus in vivo and in vitro. To examine the role of sulfhydryl groups in SNC-induced NA release, the effects of metal ions such as Hg2+ and N-ethylmaleimide (NEM, a sulfhydryl alkylating agent) on [3H]NA release from labeled rat brain slices (hippocampus and cerebral cortex) were studied and compared with the effects of SNC. The addition of 200 microM HgCl2, but not Pb2+, Zn2+, or Cd2+, stimulated [3H]NA release from both types of slices in the presence of extracellular CaCl2. p-Chloromercuribenzoic acid (p CMBA) also stimulated [3H]NA release. NEM stimulated [3H]NA release from both types of slices in the presence and absence of extracellular CaCl2. The effect of 200 microM NEM was enhanced, but the effect of 200 microM SNC was inhibited by co addition of 200 microM p-CMBA in the absence of extracellular CaCl2. The concentration-response curve of SNC shifted to the right after co-addition of 200 microM p-CMBA or 100 microM HgCl2, although the effect of 200 microM NEM was additive to the effect of SNC. These findings demonstrate that SNC acts as a sulfhydryl agent on proteins that regulate NA release, and that SNC may share the same sulfhydryl groups with Hg compounds. The effect of T-588 ?(R)-(-) (benzo[b]thiophen-5-yl)-2-[2-(N,N-diethylamino)ethoxy]eth anol hydrochloride?, a novel cognitive enhancer and a stimulator of NA release, was compared with the effects of sulfhydryl reagents. PMID- 10718343 TI - Hepatocyte growth factor: a regulator of extracellular matrix genes in mouse mesangial cells. AB - The potential role of hepatocyte growth factor (HGF) in regulating extracellular matrix in mouse mesangial cells (MMC) was evaluated. Functional HGF receptors were deed in MMC by HGF-induced extracellular acidification, a response that was inhibited by the HGF inhibitor HGF/NK2, a splice variant expressing the N terminal domain through the second kringle domain HGF also increased fibronectin and collagen alpha1 (IV) mRNA levels in these cells; the increases were associated with a concentration-dependent increase in transcriptional activity of the collagen alpha1 (IV) gene. HGF also stimulated fibronectin and collagen alpha1 (IV) mRNA levels in primary rabbit proximal tubule epithelial cells To evaluate the potential consequences of chronic elevation of HGF on renal fuction, HGF was administered continuously for 18 days to normal and diabetic C57BLKS/J lepr(db) mice. In the diabetic mice, HGF reduced creatinine clearance and increased microalbuminuria, indicating that chronic exposure to HGF impairs renal function. Thus, chronically elevated HGF may contribute to the progression of chronic renal disease in diabetes by decreasing the glomerular filtration rate and possibly promoting the accumulation of extracellular matrix. PMID- 10718344 TI - Regulation of phosphatidylserine exposure at the cell surface by the serine--base exchange enzyme system during CD95-induced apoptosis. AB - Early in the apoptotic process, CD95 induces a translocation of phosphatidylserine (PtdSer) from the inner to the outer leaflet of the cellular plasma membrane. In mammalian cells, PtdSer is only synthesized through a calcium dependent exchange of the polar head group of pre-existing phospholipids, either phosphatidylcholine or phosphatidylethanolamine, by a serine. Using a pharmacological approach, we examined the influence of PtdSer synthesis on CD95 induced PtdSer exposure at the surface of Jurkat cells. We found that CD3/TCR triggering or thapsigargin treatment of Jurkat cells was accompanied both by a decreased PtdSer synthesis and by a strong reduction of CD95-induced PtdSer at the cell surface, as monitored by fluorescence-activated cell sorting (FACS) analysis of annexin V-fluorescein isothiocyanate (FITC)-labeled cells. PtdSer synthesis through the serine-base exchange enzyme system thus appeared as one of the mechanisms implicated in the recently discovered CD3/TCR-induced down regulation of CD95-induced apoptosis. Conversely, increasing the activity of the serine-base exchange enzyme system with different drugs, either the K+ channel blocker quinine, the cationic amphiphil stearylamine, or three different calmodulin antagonists, chlorpromazine, trifluoperazine, and N-(6-aminohexyl)-5 chloro-1-naphthalene sulfonamide (W7), resulted in an increased appearance of PtdSer at the surface of CD95-treated cells. Both PtdSer synthesis and CD95 induced annexin V-FITC reactivity were abrogated in ATP-depleted cells. Also, modifying the membrane potential with valinomycin (hyperpolarization) or either gramicidin or KCl (depolarization) demonstrated a strong relationship between PtdSer synthesis and annexin V-FITC reactivity in CD95-treated cells. Together, our results indicate that CD95-induced exposure of PtdSer at the cell surface could be regulated by the activity of the serine-base exchange enzyme system. PMID- 10718345 TI - Effect of resveratrol, a natural polyphenolic compound, on reactive oxygen species and prostaglandin production. AB - Resveratrol is a natural molecule with antioxidant action. Moreover, resveratrol is also considered to be a molecule with anti-inflammatory action, an effect attributed to suppression of prostaglandin (PG) biosynthesis. The aim of the present study was to investigate the effects of resveratrol, a polyphenol present in most red wines, on reactive oxygen species formation as well as on arachidonic acid (AA) release, cyclooxygenase expression, and PG synthesis in murine resident peritoneal macrophages. Results show that resveratrol exerted a strong inhibitory effect on superoxide radical (O2-) and hydrogen peroxide (H2O2) produced by macrophages stimulated by lipopolysaccharides (LPS) or phorbol esters (PMA). Resveratrol also significantly decreased [3H]AA release induced by LPS and PMA or by exposure to O2- or H2O2. Resveratrol treatment caused a significant impairment of cyclooxygenase-2 (COX-2) induction stimulated by LPS and PMA or by O2- or H2O2 exposure. These effects of resveratrol on [3H]AA release and COX-2 overexpression were correlated with a marked reduction of PG synthesis. Our results indicate that the antioxidant action of resveratrol affects AA mobilization and COX-2 induction. PMID- 10718346 TI - Prevention of halothane-induced hepatotoxicity by hemin pretreatment: protective role of heme oxygenase-1 induction. AB - Reductive metabolism of halothane in phenobarbital-pretreated rats is known to increase free radical formation that results in hepatotoxicity. It also is associated with a marked induction of microsomal heme oxygenase-1 (HO-1), suggesting that there is an alteration in heme metabolism. In this study, we examined heme metabolism in rats pretreated with phenobarbital, followed by exposure to halothane-hypoxia. In this model, there was a significant decrease in microsomal cytochrome P450 content in the liver, followed by a rapid increase in free heme concentration and a decrease in the level of mRNA for the nonspecific delta-aminolevulinate synthase. A transient but dramatic induction of HO-1 mRNA and a prolonged induction of heat shock protein 70 mRNA also occurred. The HO-1 protein was detected principally in the hepatocytes around the central vein. Serum alanine transaminase (ALT) activity, an indicator of hepatic dysfunction, increased continuously throughout the experiment. Hemin pretreatment induced hepatic HO-1 with abrogation of the halothane-induced hepatotoxicity in this model, as judged by ALT activity and normal histology. Our findings in this study thus indicate that halothane-induced hepatotoxicity is due not only to its reductive metabolite formation, but also to an increase in hepatic free heme concentration, which is a potent prooxidant; HO-1 induction is an important protective response against such changes. This is also the first study to demonstrate that hemin pretreatment, which induces HO-1 prior to exposure to halothane, effectively prevents halothane-induced hepatotoxicity. PMID- 10718347 TI - Inhibition of H+,K+ -ATPase by hinesol, a major component of So-jutsu, by interaction with enzyme in the E1 state. AB - Hinesol, a major component of the crude drug "So-jutsu" (Atractylodis Lanceae Rhizoma), strongly inhibited H+,K+-ATPase activity with a IC50 value of 5.8x10( 5) M. It also inhibited Na+,K+-ATPase, Mg2+-ATPase, Ca2+-ATPase, and H+-ATPase activities, although the inhibition rate was lower. No effects on alkaline or acid phosphatase activities were observed. The mechanism by which hinesol inhibited H+,K+-ATPase activity was studied in detail. The inhibition was uncompetitive with respect to ATP, and it increased as the Mg2+ concentration was raised, whereas it was not affected by the K+ concentration. The activity of K+ dependent p-nitrophenyl phosphatase (K+-pNPPase), a partial reaction of H+,K+ ATPase, was inhibited by hinesol noncompetitively with respect to pNPP (IC50 value of 1.6x10(-4) M), and competitively with respect to K+, whereas it was not affected by the Mg2+ concentration. These results suggest that hinesol is a relatively specific inhibitor of H+,K+-ATPase. It appears that hinesol reacts with enzyme in the E1 state in the presence of ATP and Mg2+ and forms the complex hinesol-H+ E1-ATP or hinesol x E1-P, blocking the conformational change to the E2 state. Furthermore, hinesol enhanced the inhibitory effect of omeprazole on H+,K+ ATPase, and the inhibitory site of hinesol was different from that of omeprazole. The effect of So-jutsu as an anti-gastric ulcer agent may be ascribed to the inhibitory effect of hinesol on H+,K+-ATPase activity. PMID- 10718348 TI - Inhibitory effect of metformin on intestinal glucose absorption in the perfused rat intestine. AB - To investigate the effect of metformin on intestinal glucose absorption, a perfusion study of the intestine was performed in the rat. Male Wistar albino rats (8 weeks old) were used in the present study. The glucose absorption by the perfused intestine (788.1+/-81.9 micromol/30 min) was not changed significantly by the direct addition of metformin (90 microg/mL) to the perfusing medium (737.0+/-118.2 micromol/30 min) or by intraduodenal metformin (250 mg/kg in saline solution) infusion (772.8+/-106.3 micromol/30 min). In rats orally administered metformin (250 mg/kg) for 5 days, glucose absorption by the perfused intestine (375.0+/-164.3 micromol/30 min) was significantly (P<0.001) lower than that in control rats (811.0+/-83.1 micromol/30 min). These results indicate that metformin had a significant effect on the digestive tract, and that metformin treatment exerted an inhibitory effect on intestinal glucose absorption in the rat. PMID- 10718349 TI - Medical malpractice: treating the causes instead of the symptoms. PMID- 10718350 TI - Negligent care and malpractice claiming behavior in Utah and Colorado. AB - BACKGROUND: Previous studies relating the incidence of negligent medical care to malpractice lawsuits in the United States may not be generalizable. These studies are based on data from 2 of the most populous states (California and New York), collected more than a decade ago, during volatile periods in the history of malpractice litigation. OBJECTIVES: The study objectives were (1) to calculate how frequently negligent and nonnegligent management of patients in Utah and Colorado in 1992 led to malpractice claims and (2) to understand the characteristics of victims of negligent care who do not or cannot obtain compensation for their injuries from the medical malpractice system. DESIGN: We linked medical malpractice claims data from Utah and Colorado with clinical data from a review of 14,700 medical records. We then analyzed characteristics of claimants and nonclaimants using evidence from their medical records about whether they had experienced a negligent adverse event. MEASURES: The study measures were negligent adverse events and medical malpractice claims. RESULTS: Eighteen patients from our study sample filed claims: 14 were made in the absence of discernible negligence and 10 were made in the absence of any adverse event. Of the patients who suffered negligent injury in our study sample, 97% did not sue. Compared with patients who did sue for negligence occurring in 1992, these nonclaimants were more likely to be Medicare recipients (odds ratio [OR], 3.5; 95% CI [CI], 1.3 to 9.6), Medicaid recipients (OR, 3.6; 95% CI, 1.4 to 9.0), > or =75 years of age (OR, 7.0; 95% CI, 1.7 to 29.6), and low income earners (OR, 1.9; 95% CI, 0.9 to 4.2) and to have suffered minor disability as a result of their injury (OR, 6.3; 95% CI, 2.7 to 14.9). CONCLUSIONS: The poor correlation between medical negligence and malpractice claims that was present in New York in 1984 is also present in Utah and Colorado in 1992. Paradoxically, the incidence of negligent adverse events exceeds the incidence of malpractice claims but when a physician is sued, there is a high probability that it will be for rendering nonnegligent care. The elderly and the poor are particularly likely to be among those who suffer negligence and do not sue, perhaps because their socioeconomic status inhibits opportunities to secure legal representation. PMID- 10718351 TI - Incidence and types of adverse events and negligent care in Utah and Colorado. AB - BACKGROUND: The ongoing debate on the incidence and types of iatrogenic injuries in American hospitals has been informed primarily by the Harvard Medical Practice Study, which analyzed hospitalizations in New York in 1984. The generalizability of these findings is unknown and has been questioned by other studies. OBJECTIVE: We used methods similar to the Harvard Medical Practice Study to estimate the incidence and types of adverse events and negligent adverse events in Utah and Colorado in 1992. DESIGN AND SUBJECTS: We selected a representative sample of hospitals from Utah and Colorado and then randomly sampled 15,000 nonpsychiatric 1992 discharges. Each record was screened by a trained nurse-reviewer for 1 of 18 criteria associated with adverse events. If > or =1 criteria were present, the record was reviewed by a trained physician to determine whether an adverse event or negligent adverse event occurred and to classify the type of adverse event. MEASURES: The measures were adverse events and negligent adverse events. RESULTS: Adverse events occurred in 2.9+/-0.2% (mean+/-SD) of hospitalizations in each state. In Utah, 32.6+/-4% of adverse events were due to negligence; in Colorado, 27.4+/-2.4%. Death occurred in 6.6+/-1.2% of adverse events and 8.8+/-2.5% of negligent adverse events. Operative adverse events comprised 44.9% of all adverse events; 16.9% were negligent, and 16.6% resulted in permanent disability. Adverse drug events were the leading cause of nonoperative adverse events (19.3% of all adverse events; 35.1% were negligent, and 9.7% caused permanent disability). Most adverse events were attributed to surgeons (46.1%, 22.3% negligent) and internists (23.2%, 44.9% negligent). CONCLUSIONS: The incidence and types of adverse events in Utah and Colorado in 1992 were similar to those in New York State in 1984. Iatrogenic injury continues to be a significant public health problem. Improving systems of surgical care and drug delivery could substantially reduce the burden of iatrogenic injury. PMID- 10718352 TI - Tolerance of uncertainty of medical students and practicing physicians. AB - BACKGROUND: Tolerance of uncertainty is believed to be an important attribute of practicing physicians. This study attempts to (1) estimate how medical students perceive physicians' tolerance of uncertainty and (2) measure the tolerance of uncertainty of practicing physicians. RESEARCH DESIGN: Cross-sectional. SETTING AND SUBJECTS: Medical students (n = 113) and practicing physicians (n = 151) at the Faculty of Health Sciences, Ben-Gurion University, Israel. MEASURES: A self administered, Hebrew version of an instrument developed in the United States. INDEPENDENT VARIABLES: Age, gender, seniority (year of study for students or years in practice for physicians), country of birth for students or of graduation for physicians, and physicians' specialty. DEPENDENT VARIABLES: Two dimensions, which were identified by factor analysis: reluctance to disclose uncertainty and stress from uncertainty. RESULTS: The estimates of physicians' stress from uncertainty by first-year students aged <22 years were higher than those by first year students aged > or =22 years. There were no significant differences in the way junior and senior medical students perceived physicians' tolerance of uncertainty. Stress from uncertainty was higher in female physicians (P = 0.028) and in graduates of the former Soviet Union (P = 0.044) than among male physicians and Israeli graduates, respectively. Reluctance to disclose uncertainty was higher among graduates of the former Soviet Union (P = 0.003) and among psychiatrists (P = 0.021) than among Israeli graduates and other specialties, respectively. CONCLUSIONS: The reliability and factor structure of the instrument were replicated. The previously reported differences in tolerance of uncertainty between women and men and between local and foreign graduates were confirmed. Physicians' tolerance of uncertainty appeared to be higher than that attributed to them by students. The expected age-related differences in perception of clinical uncertainty were not detected between junior and senior medical students. PMID- 10718353 TI - Underutilization of mammography in older breast cancer survivors. AB - BACKGROUND: Annual mammography is recommended for all breast cancer survivors. OBJECTIVES: To elucidate mammography use among older survivors of breast cancer and to explore determinants of such use. RESEARCH DESIGN: Retrospective cohort study using data from the Surveillance, Epidemiology, and End Results (SEER) registry linked to Medicare claims. SUBJECTS: A cohort of 3885 breast cancer survivors aged > or =65 years diagnosed with early-stage breast cancer in the United States in 1991. MEASURES: Medicare mammogram claims during the 2-year period following initial breast cancer treatment. RESULTS: Overall, 62% of the cohort underwent annual mammography, 23% underwent mammography in 1 of 2 years, and 15% had no mammography claim in the 2 years evaluated. Twenty-two percent of the women who underwent breast-conserving surgery (BCS) without radiotherapy had no mammogram in the 2-year period evaluated, compared with 17% of those who underwent mastectomy and 4% of those who underwent BCS with radiotherapy. In multivariate analyses controlling for age, cancer stage, and other patient factors, the use of annual mammography was significantly lower among women treated with mastectomy or BCS without radiotherapy than among women treated with BCS with radiotherapy. CONCLUSIONS: Mammography is underused in the follow-up care of older breast cancer survivors. Underuse is of particular concern in women treated with BCS without radiotherapy because of the high risk of local disease recurrence. It is unknown whether poorer follow-up care contributes to the previously described lower rate of long-term survival among women who received this therapy. PMID- 10718354 TI - Health Utilities Index Mark 3: evidence of construct validity for stroke and arthritis in a population health survey. AB - BACKGROUND: The Health Utilities Index Mark 3 (HUI3) is a comprehensive, compact health status classification and health state preference system. The HUI3 system has been included in 4 Canadian population health surveys and numerous clinical trials. OBJECTIVES: To evaluate the construct validity of the HUI3 for the measurement of health-related quality of life (HRQL) and attribute-specific morbidity in respondents to the 1990 Ontario Health Survey reported to have arthritis or stroke. The authors assessed (1) whether those with stroke, arthritis, and both conditions had lower HRQL scores than those with neither condition and (2) whether HUI3 detects morbidity in specific health attributes affected by arthritis and stroke. Stroke (but not arthritis) were expected to affect speech and cognition; arthritis (but not stroke) to affect pain; both to affect mobility, dexterity, and emotion; and neither to affect vision and hearing. RESEARCH DESIGN: Linear regression models of HRQL and attribute-specific utilities were estimated as a function of 3 indicator variables of health problem (stroke only, arthritis only, both) and variables included to reduce confounding. RESULTS: Subjects with stroke, arthritis, and both conditions had substantially lower HRQL than those with neither condition. Stroke subjects had greater morbidity in speech and cognition than arthritis subjects; somewhat surprisingly, pain morbidity was only slightly higher among arthritis subjects; neither condition affected vision or hearing. These associations were robust to various model specifications. CONCLUSIONS: The HUI3 system appears valid for measuring health status and HRQL for stroke and arthritis in the context of a noninstitutionalized population health survey. PMID- 10718355 TI - Racial and ethnic differences in a patient survey: patients' values, ratings, and reports regarding physician primary care performance in a large health maintenance organization. AB - BACKGROUND: Few studies have investigated the influence of race and/or ethnicity on patients' ratings of quality of care. None have incorporated patients' values and beliefs regarding medical care in assessing these possible differences. OBJECTIVES: We explored whether patients' values, ratings, and reports regarding physicians' primary care performance differed by race and/or ethnicity. RESEARCH DESIGN: This was a cross-sectional, mailed patient survey. SUBJECTS: The study subjects were adult primary care patients in a large health maintenance population (7,747 whites, 836 blacks, 710 Latinos, and 1,007 Asians). MEASURES AND METHODS: Ratings of the following dimensions of primary care were measured: technical competence, communication, accessibility, prevention and health promotion, and overall satisfaction. Patients' values regarding these dimensions and their confidence in medical care were measured. Multivariate analyses yielded associations of race/ethnicity with satisfaction and with reports of prevention services received. RESULTS: For 7 of the 10 dimensions of primary care measured, Asians rated physician performance significantly less favorably than did whites, including differences among Asian ethnic subgroups. Latinos rated physicians' accessibility less favorably than did whites. Blacks rated physicians' psychosocial and lifestyle health promotion practices higher than did whites. No differences were found in patient reports of prevention services received, except Pacific Islanders reported receiving significantly more prevention services than whites. CONCLUSIONS: In a large HMO population, significant differences were found by race and ethnicity, and among Asian ethnic subgroups, in levels of patient satisfaction with primary care. These findings may represent actual differences in quality of care or variations in patient perceptions, patient expectations, and/or questionnaire response styles. More research is needed to assess, in accurate and culturally appropriate ways, whether health plans are meeting the needs of all enrollees. PMID- 10718356 TI - Is provider capitation working? Effects on physician-hospital integration and costs of care. AB - BACKGROUND: Capitation holds health providers fiscally responsible for the services they deliver or arrange and thus provides strong motivation for physicians and hospitals to integrate activities and reduce costs of care. OBJECTIVES: The objective of this study was to assess 2 potential effects of capitation: (1) its effects on the integration of functional, financial, and clinical processes between hospitals and physicians and (2) its effects, in conjunction with process integration, on hospital costs. STUDY DESIGN: We studied a 1995 American Hospital Association (AHA) special survey that has information on 44 different physician-hospital integrative activities and on global capitation contracts held by management service organizations, physician-hospital organizations, and other similar entities. These data were combined with the AHA's Annual Survey of Hospitals, InterStudy HMO data, the area resource file, and state regulation data. Multivariate analysis was used to assess the relationship between capitation and integration and then to examine the influence of these factors and others on hospital costs. We studied 319 urban hospitals with complete data. FINDINGS: Provider capitation was found to promote integration between hospitals and physicians in relation to administrative/practice management, physician financial risk sharing, joint ventures to create new services, computer linkages, and an overall measure of physician-hospital integration. However, anticipated effects of integration and capitation on hospital costs were not evident. CONCLUSIONS: Global capitation is motivating tighter integration between physicians and hospitals in a number of respects. Although capitation is currently having the intermediate effect of encouraging process integration, it is not yet having the ultimate anticipated effect of lowering hospital costs. PMID- 10718357 TI - Patient satisfaction with hospital care: effects of demographic and institutional characteristics. AB - BACKGROUND: There are a growing number of efforts to compare the service quality of health care organizations on the basis of patient satisfaction data. Such efforts inevitably raise questions about the fairness of the comparisons. Fair comparisons presumably should not penalize (or reward) health care organizations for factors that influence satisfaction scores but are not within the control of managers or clinicians. On the basis of previous research, these factors might include the demographic characteristics of patients (eg, age) and the institutional characteristics (eg, size) of the health care organizations where care was received. OBJECTIVES: The goal of this study was to examine the extent to which a patient's satisfaction scores are related to both his/her demographic characteristics and the institutional characteristics of the health care organization where care was received. METHODS: We conducted an analysis of secondary data from the Veterans Health Administration (VHA), US Department of Veterans Affairs. The database contained patient responses to self-administered satisfaction questionnaires and information about demographic characteristics. Additional data from VHA were obtained regarding the institutional characteristics of the hospitals where patients received their care. RESULTS: Among demographic characteristics, age, health status, and race consistently had a statistically significant effect on satisfaction scores. Among the institutional characteristics, hospital size consistently had a significant effect on patient satisfaction scores. CONCLUSIONS: Study results can be interpreted as justifying the need to adjust patient satisfaction scores for differences in patient population among health care organizations. However, from a policy perspective, such adjustments may ultimately create a disincentive for health care organizations to customize their care. PMID- 10718358 TI - Patient preferences for medical decision making: who really wants to participate? AB - OBJECTIVES: To identify the determinants of patient preferences for participation in medical decision making. METHODS: Data were analyzed for 2,197 patients from the Medical Outcomes Study, a 4-year observational study of patients with chronic disease (hypertension, diabetes, myocardial infarction, congestive heart failure, and depression). Multivariate logistic regression models estimated the effects of patients' sociodemographic, clinical, psychosocial, and lifestyle characteristics on their decision-making preferences. RESULTS: A majority of the patients (69%) preferred to leave their medical decisions to their physicians. The odds for preferring an active role significantly decreased with age and increased with education. Women were more likely to be active than men (odds ratio [OR] = 1.44, P < 0.001). Compared with patients who only suffered with unsevere hypertension, those with severe diabetes (OR = 0.62, P = 0.04) and unsevere heart disease (OR = 0.45, P = 0.02) were less likely to prefer an active role. Patients with clinical depression were more likely to be active (OR = 1.64, P = 0.01). Patients pursuing active coping strategies had higher odds for an active role than "passive" copers, while those who placed higher value on their health were less likely to be active than those with low health value (OR = 0.59, P < 0.001). CONCLUSIONS: Although a majority of patients prefer to delegate decision making to physicians, preferences vary significantly by patient characteristics. Approaches to enhancing patient involvement will need to be flexible and accommodating to individual preferences in order to maximize the benefits of patient participation on health outcomes. PMID- 10718359 TI - An English and Spanish Pediatric Asthma Symptom Scale. AB - BACKGROUND: Pediatric asthma survey measures have not been adequately tested in non-English-speaking populations. OBJECTIVES: To test the reliability and validity of an English and Spanish symptom scale to measure asthma control in children. SUBJECTS: Parents (54% Spanish-speaking; 61% not high school graduates) of 234 children seen in the emergency department for an asthma exacerbation. MEASURES: Parent report of frequency and perceived severity of child asthma symptoms during the beginning and after resolution of the exacerbation. RESULTS: An 8-item scale composed of reports of cough, wheezing, shortness of breath, asthma attacks, chest pain, night symptoms, and overall perceived severity had very good psychometric properties in both English and Spanish. The reliability (Cronbach's alpha) of the scale ranged from 0.81 to 0.87 for both languages and time frames. In both languages, the validity of the scale was supported by responsiveness to changes in clinical status (lower symptom score after resolution of the exacerbation, P < 0.001) and by moderate to strong correlations (P < 0.001) with other asthma morbidity measures (parent report of child bother: r = 0.59-0.65; school days lost: r = 0.38-0.67; and activity days lost: r = 0.41 0.59). There were no statistically significant differences in the reliability or construct validity of the summary symptom scale by language, although Spanish speakers reported a lower frequency of some symptoms than did English speakers. CONCLUSIONS: A reliable and valid 8-item scale can be used to measure control of asthma symptoms in Spanish-speaking populations of low literacy. Additional research to evaluate language equivalency of asthma measures is necessary. PMID- 10718360 TI - Self-regulation of type I collagen degradation by collagen-induced production of matrix metalloproteinase-1 on cholangiocarcinoma and hepatocellular carcinoma cells. PMID- 10718361 TI - Transfection of mouse mammary epithelial cells. PMID- 10718362 TI - Development of myofibroblasts from human bone marrow mesenchymal stem cells cocultured with human colon carcinoma cells and TGF beta 1. PMID- 10718363 TI - Relative sensitivity of undifferentiated and cyclic adenosine 3',5'-monophosphate induced differentiated neuroblastoma cells to cyclosporin A: potential role of beta-amyloid and ubiquitin in neurotoxicity. AB - Cyclosporin A is routinely used in transplant therapy following allogeneic or xenogeneic tissue transplantation to prevent rejection. This immunosuppressive drug is also neurotoxic; however, its mechanisms of action for neurotoxicity are poorly understood. Undifferentiated and cyclic adenosine 3',5'-monophosphate (cAMP)-induced differentiated neuroblastoma (NB) cells were used as an experimental model to study the toxicity of cyclosporin A. Results showed that cyclosporin A promoted the outgrowth of neurites and inhibited the growth of undifferentiated NB cells. When cyclosporin A was added simultaneously with RO20 1724, an inhibitor of cyclic nucleotide phosphodiesterase, or with prostaglandin E1, a stimulator of adenylate cyclase, it markedly enhanced the growth inhibitory and differentiation effects of these cAMP-stimulating agents. In addition, cyclosporin A added to cAMP-induced differentiated NB cells caused dose-dependent degeneration of these cells as evidenced by the vacuolization of cytoplasm and the fragmentation of nuclear and cytoplasmic materials; however, neurites remained intact. Cyclosporin A alone did not alter the intensity of cell immunostaining for ubiquitin or beta-amyloid peptide (amino acids 1-14) (Abeta1 14); however, it enhanced the intensity of staining for both ubiquitin and Abeta in cells that were treated with cAMP-stimulating agents. The intensity of staining of amyloid precursor protein (amino acids 44-63) (APP44-66) did not change in any treated group, suggesting that the increase in Abeta staining is due to increased processing of APP to Abeta. We propose that one of the mechanisms of cyclosporin A-induced neurotoxicity involves increased levels of Abeta and ubiquitin. PMID- 10718364 TI - In vitro teratogenic potential of two antifungal triazoles: triadimefon and triadimenol. AB - The teratogenic potential of two antifungal triazoles (Triadimefon and Triadimenol) has been investigated in vitro by the rat postimplantation whole embryo culture method. Rat embryos 9.5 d old were cultured for 48 h in rat serum with Triadimefon (12.5-250 microM) or Triadimenol (6.25-125 microM) and then examined. Some embryos exposed to Triadimenol (6.25-125 microM) were cultured for 12 extra hours in control serum to improve their developmental degree and then immunostain cranial nerves and ganglia. The exposure to the highest doses of triazoles only moderately reduced some morphometrical developmental parameters. By contrast, 25-250 microM Triadimefon and 25-125 microM Triadimenol induced specific concentration-related teratogenic effects at the level of first and second branchial arches. After immunostaining, embryos exposed to 12.5-125 microM Triadimenol showed specific cranial nerve and ganglia abnormalities. The possible implication of neural crest cell alterations on triazole-related abnormalities is discussed. PMID- 10718365 TI - Multistep production of bioengineered skin substitutes: sequential modulation of culture conditions. AB - Many studies are being conducted to define the role of growth factors in cutaneous physiology in order to add cytokines in a timely fashion for optimal tissue engineering of skin. This study is aimed at developing a multistep approach for the production of bioengineered skin substitutes, taking into account the effects of various growth factors according to the culture time. The use of a serum-supplemented medium throughout the whole culture period of skin substitutes was compared to the sequential use of specific additives at defined culture steps. Histological analysis revealed that serum was necessary for keratinocyte proliferation and migration on dermal substitutes during the first 2 d after their seeding. However, the serum-free medium presented some advantages when supplemented with different additives at specific culture steps. Interestingly, ascorbic acid added to the dermal substitutes before and after keratinocyte seeding maintained their cuboidal morphology in the basal epidermal layer. In the absence of serum, collagen matrix degradation slowed down, and a better multilayered epidermal organization was obtained, notably with retinoic acid. Stratum corneum formation was also enhanced by fatty acids. Thus, sequential addition of exogenous factors to the medium used to produce skin substitutes can improve their structural features and functional properties in vitro. PMID- 10718366 TI - High level of CA19-9, CA50, and CEA-producible human cholangiocarcinoma cell line changes in the secretion ratios in vitro or in vivo. AB - The ascites of a 78-yr-old Japanese woman with cholangiocarcinoma was used for a primary culture. An established new cell line (designated TK from the Japanese description of cholangiocarcinoma; Tankan-gann) showed conspicuous tumor marker production. A high level of circulating serum tumor markers; carbohydrate antigen (CA) 19-9, 32,000 U/ml; CA50, 6900 U/ml; and carcinoembryonic antigen (CEA), 300 ng/ml (on an average from 10(6) cells/ml for 3 d culture) were detected in the tissue culture supernatant. With an inoculum of 2 x 10(7) TK cells, nude mice progressively developed tumors. The histological features of the tumors forming in nude mice showed well-differentiated adenocarcinoma. Within the tumor mass, large amounts of extra cellular fluid retained approximately 590,000 U/ml of CA19 9, 200,000 U/ml of CA50, and 2000 ng/ml of CEA. Alpha-feto-protein was undetectable in the TK culture supernatant. There are few cholangiocarcinoma cell lines producing stable human tumor markers. Newly established TK cells derived from cholangiocarcinoma showed a stable production of serum tumor markers in vivo and in vitro. The changes in the tumor marker secretion ratios were shown to be dependent upon type of tumor cells, i.e., whether they are in vitro or in vivo. These features make TK cells a valuable tool for studying tumor markers. PMID- 10718367 TI - Transfer of SV40 temperature-sensitive early gene into human epidermal keratinocytes by the recombinant adenovirus vector. AB - We constructed a recombinant adenovirus vector that contained the origin defective SV40 early gene, coding temperature-sensitive T antigen. This vector transferred the SV40 early gene into human epidermal keratinocytes with high efficiency. T antigen conferred the ability of keratinocytes to grow with limited differentiation in the presence of serum and high calcium concentration at the permissive temperature (34 degrees C), although normal keratinocytes were induced to differentiate and stop growing under the same conditions. The serum/Ca++ resistant cells did not proliferate at the nonpermissive temperature (40 degrees C), indicating that they depended on T antigen for their proliferation. The temperature-sensitive T antigen dissociated from the tumor suppressor gene products, p53, at 40 degrees C. The serum/Ca++-resistant cells still had the ability to proceed to terminal differentiation when injected into SCID mice as cultured keratinocytes. However, they did not form an apparent basal layer. This indicated that the tissue remodeling process in the serum/Ca++-resistant keratinocytes was abnormal. All of these epidermoid cysts disappeared within 8 wk and no tumor developed for 6 mo. We consider that deltaE1/SVtsT is a useful tool to examine multistep carcinogenesis of human epithelial cells in vitro. PMID- 10718368 TI - Adaptation of an insect cell line of Spodoptera frugiperda to grow at 37 degrees C: characterization of an endodiploid clone. AB - Sf21 and Sf9 cell lines established from the lepidoptera Spodoptera frugiperda do not display major induction of heat shock proteins when exposed to a temperature of 37 degrees C. After some months of adaptation at 37 degrees C we obtained two new cell lines, Sf21-HT and Sf9-HT, which have now been established for several years in our laboratory. The Sf9-HT line displays a slightly shorter doubling time at 37 degrees C than the wild type at 28 degrees C, but cell lethality gives rise to an earlier growth arrest. The composition of total lipid extract from heat-adapted cells reveals a higher sphingomyelin to phosphatidylcholine ratio and a higher percentage of saturated fatty acids, which are expected for the lower membrane fluidity, required for thermotolerance. The cell volume of Sf9-HT is doubled, and by flow cytometry we showed that the DNA content is twice that in the parental cell line. Karyotypic examination of metaphasic cells achieved under epifluorescence microscopy revealed a doubled chromosome number in Sf9-HT. PMID- 10718369 TI - Human macrovascular endothelial cells: optimization of culture conditions. AB - The purpose of this study is to identify optimal culture conditions to support the proliferation of human macrovascular endothelial cells. Two cell lines were employed: human saphenous vein endothelial cells (HSVEC) and human umbilical vein endothelial cells (HUVEC). The influence of basal nutrient media (14 types), fetal bovine serum (FBS), and mitogens (three types) were investigated in relation to cell proliferation. Additionally, a variety of extracellular matrix (ECM) substrate-coated culture dishes were also tested. The most effective nutrient medium in augmenting cell proliferation was MCDB 131. Compared to the more commonly used M199 medium, MCDB 131 resulted in a 2.3-fold increase in cell proliferation. Media containing 20% FBS increased cell proliferation 7.5-fold compared to serum-free media. Among the mitogens tested, heparin (50 microg/ml) and endothelial cell growth supplement (ECGS) (50 microg/ml) significantly improved cell proliferation. Epithelial growth factor (EGF) provided no improvement in cell proliferation. There were no statistical differences in cell proliferation or morphology when endothelial cells were grown on uncoated culture plates compared to plates coated with ECM proteins: fibronectin, laminin, gelatin, or collagen types I and IV. The culture environment yielding maximal HSVEC and HUVEC proliferation is MCDB 131 nutrient medium supplemented with 2 mM glutamine, 20% FBS, 50 microg/ml heparin, and 50 microg/ml ECGS. The ECM substrate-coated culture dishes offer no advantage. PMID- 10718370 TI - Inhibition of myogenesis by ouabain: effect on protein synthesis. AB - Ouabain, a specific inhibitor of the sodium- and potassium-activated adenosine triphosphatase, causes reversible inhibition of the fusion of myoblasts to form myotubes. We further examined this observation to investigate whether control of Na/K-ATPase activity may normally contribute to the regulation of myogenesis. In control cultures, fusion was preceded by a small decrease in intracellular sodium concentration, but intracellular sodium and potassium increased significantly during fusion. Levels of ouabain that produce prolonged inhibition of fusion (400 microM) virtually eliminated sodium and potassium gradients. However, lower ouabain levels (10-100 microM) also produced significant changes in intracellular potassium and/or sodium along with little apparent decrease in the eventual extent of fusion. The effect of ouabain on protein synthesis was also examined. Low levels of ouabain (<50 microM) that did not affect myogenesis also did not affect incorporation of radiolabeled amino acids, while higher concentrations produced a decline in protein synthesis that paralleled decreases in the rate of myoblast fusion. Levels of metabolic labeling were reduced 90% in cultures treated with 400 microM ouabain. Inhibition of protein synthesis would prevent membrane remodeling required for fusion and other events in myogenesis. Thus, our results do not support any specific role for the sodium- and potassium-activated adenosine triphosphatase in regulating myogenesis. PMID- 10718371 TI - Function of 90-kDa heat shock protein in cellular differentiation of human embryonal carcinoma cells. AB - Heat shock proteins (HSPs) have been recognized as molecules that maintain cellular homeostasis during changes in the environment. Here we report that HSP90 functions not only in stress responses but also in certain aspects of cellular differentiation. We found that HSP90 showed remarkably high expression in undifferentiated human embryonal carcinoma (EC) cells, which were subsequently dramatically down-regulated during in vitro cellular differentiation, following retinoic acid (RA) treatment, at the protein level. Surprisingly, heat shock treatment also triggered the down-regulation of HSP90 within 48 h at the protein level. Furthermore, the heat treatment induced cellular differentiation into neural cells. This down-regulation of HSP90 by heat treatment was shifted to an up-regulation pattern after cellular differentiation in response to RA treatment. In order to clarify the functions of HSP90 in cellular differentiation, we conducted various experiments, including overexpression of HSP90 via gene transfer. We showed that the RA-induced differentiation of EC cells into a neural cell lineage was inhibited by overexpression of the HSP90alpha or -beta isoform via the gene transfer method. On the other hand, the overexpression of HSP90beta alone impaired cellular differentiation into trophoectoderm. These results show that down-regulation of HSP90 is a physiologically critical event in the differentiation of human EC cells and that specific HSP90 isoforms may be involved in differentiation into specific cell lineages. PMID- 10718372 TI - Ex vivo T-lymphocyte derived cytokine production in SIRS patients is influenced by experimental procedures. AB - Ex vivo production of interleukin-2 (IL-2), IL-4, IL-5, and IL-10 by peripheral blood mononuclear cells (PBMC) was studied in 13 septic patients with infectious systemic inflammatory response syndrome (SIRS) and 13 patients undergoing cardiac surgery with cardiopulmonary bypass (CPB) (noninfectious SIRS). We have investigated the levels of cytokines after activation by either concanavalin A (ConA), phytohemagglutinin (PHA), or anti-CD3 antibodies. In whole blood assays, ConA-induced IL-10 was significantly reduced in both groups of patients compared with healthy controls. In sepsis patients, IL-2, IL-5, and IL-10 productions by isolated PBMC were diminished on ConA-induced activation but not in response to PHA and anti-CD3; in CPB patients, only anti-CD3-induced IL-10 production was significantly reduced. Our data indicate that subtle modifications of the reactivity of circulating cells occur during infectious and noninfectious SIRS. Production of both Th1 and Th2 cytokines can be down-regulated; however, the nature of the SIRS, of the cell population, and of the activator may influence the observation. PMID- 10718373 TI - Differential regulation of Ca2+ influx by fMLP and PAF in human neutrophils: possible involvement of store-operated Ca2+ channel. AB - Calcium (Ca2+) influx into human polymorphonuclear cells (PMNs) in response to N formyl-Met-Leu-Phe (fMLP) and platelet-activating factor (PAF) stimulation was studied. Whole blood was taken by venous puncture from healthy human volunteers. PMNs were isolated, diluted, and incubated with 2 microM fura-2 AM. The cytosolic free calcium concentration, [Ca2+]i, in human neutrophils was determined by microfluorometry. We found that the net area under the fMLP- or PAF-induced [Ca2+]i rise curve in Ca2+-free medium decreased to 75% or 30% of the area under the curve in Ca2+ medium. Treatment of PMNs with phorbol myristate acetate (PMA), a protein kinase C activator, completely abolished the intracellular Ca2+ level stimulated by PAF, but not the intracellular Ca2+ level stimulated by fMLP. Treatment of PMNs with PAF did not abolish the intracellular Ca2+ level elevation stimulated by fMLP. In addition, treatment of PMNs with fMLP did not abolish intracellular Ca2+ level elevation stimulated by PAF. Loperamide, a positive modulator for store-operated calcium (SOC) channels, elicited an increase in intracellular calcium after the activation of SOC channels stimulated by fMLP or PAF. After the addition of guanosine 3',5'-cyclic monophosphate, N2,2'-O Dibutyryl-, sodium salt (db-cGMP), the initial increase of PAF- or fMLP-induced PMNs intracellular Ca2+ fluorescences was well preserved, but the slope and the peak height of fluorescence curves declined compared with the curves without db cGMP. In conclusion, we found that PAF and fMLP regulate the Ca2+ influx of PMNs in different ways. Most of the PAF-induced [Ca2+]i rise resulted from Ca2+ influx, and most of the fMLP-induced [Ca2+]i elevation resulted from intracellular stores release. The initial mobilization of intracellular Ca2+ stores in PAF-stimulated signals is mediated by protein kinase C (PKC) phosphorylation, but not in fMLP-stimulated route. SOC channels are present and important in the fMLP- or PAF-induced PMNs Ca2+ influx. There was no apparent cross-regulation between PAF- and fMLP-stimulated intracellular Ca2+ influx. PMID- 10718374 TI - Interactions of calcium/calmodulin-dependent protein kinases (CaMK) and extracellular-regulated kinase (ERK) in monocyte adherence and TNFalpha production. AB - The circulating monocyte possesses a markedly different functional phenotype relative to the macrophage (Mphi). The adhesive interactions encountered by the monocyte, en route to the inflammatory focus, generate signals that culminate in the expression of a pro-inflammatory Mphi phenotype, marked by enhanced cytokine production. Previously, we demonstrated that calcium and calmodulin are essential for maximal Mphi activation and, in particular, TNFalpha production. These effects are likely to be mediated through signal transduction kinases that require the calcium/calmodulin complex. Here, we investigated the effect of adherence on calcium/calmodulin-dependent protein kinase (CaMK) II and IV activation of the extracellular-signal regulated kinase (ERK) 1/2 cascade and on lipopolysaccharide (LPS)-induced TNFalpha production by human monocytes. Adherence activated ERK 1/2 and led to an 8-fold potentiation in LPS-induced TNFalpha production over similarly stimulated non-adherent cells. Inhibition of CaMK II prior to adherence prevented ERK 1/2 activation and attenuated by up to 40%, the TNFalpha response to subsequent LPS stimulation. CaMK II inhibition after adherence, however, failed to modify cytokine release. Inhibition of CaMK IV, both after adherence and in non-adherent monocytes, significantly inhibited LPS-induced ERK 1/2 activation and abrogated TNFalpha production by up to 75%. These data suggest that the function of CaMK II in TNFalpha production by adherent monocytes occurs during adhesion, is mediated in part by activation of ERK 1/2, and appears to "prime" the monocyte for enhanced cytokine production. CaMK IV, through activation of ERK 1/2, appears to have a direct role in the LPS signal transduction for TNFalpha production. PMID- 10718375 TI - Differential effects of short-term ace- and AT1-receptor inhibition on postischemic injury and leukocyte adherence in vivo and in vitro. AB - There is recent evidence that angiotensin-converting enzyme (ACE) inhibition reduces postischemic injury and angiotensin II receptor inhibition may have similar effects. We therefore further characterized the role of ACE- vs. AT1 receptor inhibition on cell injury and temporal association of leukocyte endothelial interaction in response to ischemia-reperfusion. A combined in vivo and in vitro study comparing the ACE inhibitor enalapril and the AT1-receptor antagonist losartan was performed. The extent and temporal correlation of cellular damage (propidium-iodide staining), microvascular perfusion failure and leukocyte-endothelial interaction (leukocyte adherence) were investigated by means of intravital microscopy, after the application of hemodynamically ineffective doses of enalapril and losartan (5 mg/kg). A hamster dorsal skinfold model with a 4-h tourniquet ischemia was used. In vitro, the effect of enalapril and losartan on polymorphonuclear cell (PMN) adherence, as well as adhesion molecule expression (ICAM-1, VCAM-1), on hypoxia- or IL-1beta-stimulated endothelial cells (HUVEC) was assessed using a PMN-adhesion assay and flow cytometry, respectively. Ischemia-reperfusion responses revealed a biphasic pattern, comprised of an early phase (30 min) of acute cellular damage and microvascular perfusion failure, followed by a late increase (240 min) in leukocyte adherence in vivo. Enalapril significantly reduced early cellular damage, microvascular perfusion failure, and leukocyte adherence in response to ischemia-reperfusion. Conversely, AT1 receptor inhibition with losartan proved to be ineffective at attenuating postischemic microcirculatory disorders (leukocyte endothelial interactions, microvascular perfusion failure) and aggravated cellular injury. In vitro, enalapril reduced PMN adherence and ICAM-1 and VCAM-1 expression, while losartan was ineffective in the same respect. Following ischemia-reperfusion injury, ACE- versus AT1-receptor inhibition induce differential effects concerning the extent and temporal association of cell injury and leukocyte-endothelial interaction. The use of enalapril combines the beneficial effects of preventing cell and vascular injury immediately after reperfusion, with a delayed inhibition of the inflammatory response. Since the AT1-receptor inhibitor losartan did not mimic effects obtained with ACE inhibition, it is conceivable that the responses in ischemia-reperfusion are mediated by a non-angiotensin II-AT1 receptor-dependent mechanism. PMID- 10718376 TI - Role of hyperbaric oxygen exposure in reduction of lipid peroxidation and in multiple organ failure induced by zymosan administration in the rat. AB - The aim of the present study was to evaluate the effects of hyperbaric oxygen (HBO) therapy on multiple organ failure induced by zymosan. Administration of zymosan (500 mg/kg) in the rat induced neutrophil infiltration in the lung, liver, and intestine as evaluated by increase in myeloperoxidase (MPO) activity. Therefore, lipid peroxidation was significantly increased in zymosan-treated rats. This inflammatory process coincided with the damage of lung, liver, and small intestine. Immunohistochemical examination demonstrated a marked increase in the immunoreactivity to nitrotyrosine in the lung, liver, and small intestine of zymosan-shocked rats. HBO (2 absolute Atmosphere) exposure attenuates the increase in the tissue levels of MPO and malondialdehyde (MDA) caused by zymosan in the lung, liver, and intestine. In addition, HBO (2 absolute Atmosphere) was effective in preventing the development of lung, liver, and intestine injury. Taken together, the present results demonstrate that HBO may also be an efficacious treatment in multiple organ failure induced by zymosan. PMID- 10718377 TI - VMN hypothalamic dopamine and serotonin in anorectic septic rats. AB - During sepsis, catabolism of proteins and associated changes in plasma amino acids occur. Tryptophan and tyrosine, and their derivatives serotonin (5-HT) and dopamine (DA), influence hypothalamic feeding-related areas and are associated with the onset of anorexia. We hypothesized that anorexia of sepsis is associated with changes in serotonin and dopamine in the ventromedial nucleus (VMN) of the hypothalamus. The aim of this study was to test our hypothesis by measuring intra VMN changes of these two neurotransmitters at the onset of anorexia during sepsis. Fischer 344 male rats had an intracerebral guide cannula stereotaxically implanted into the VMN. Ten days later, in awake, overnight-food-deprived rats, a microdialysis probe was inserted through the in situ VMN cannula. Two hours thereafter, serial baseline serotonin and dopamine concentrations were measured. Then cecal ligation and puncture to induce sepsis or a control laparotomy was performed under isoflurane anesthesia. VMN microdialysis samples were serially collected every 30 min for 8 h after the surgical procedure to determine 5-HT and DA changes in response to sepsis. During the hypermetabolic response to sepsis, a strong association occurred between anorexia and a significant reduction of VMN dopamine concentration (P < 0.05; constant rate of dopamine decrease in the Study group of 0.99 pg per 2 h); no changes occurred in 5-HT in association with anorexia of sepsis. Six hours after operation, a single meal was offered for 20 min to assess the response of neurotransmitters to food ingestion. Food intake was minimal in anorectic septic rats (mean size of the after food-deprived meal in the Septic group was 0.03+/-0.01 g, that of the Control group was 1.27+/-0.14 g; P = 0.0001), while Control rats demonstrated anticipated changes in neurotransmitters in response to eating. We conclude that the onset of anorexia in septic rats is associated with a reduction in VMN dopamine. PMID- 10718378 TI - Phagocytic and intestinal endothelial and epithelial barrier function during the early stage of small intestinal ischemia and reperfusion injury. AB - The effects of intestinal ischemia and reperfusion (I/R) on small intestinal mucosal endothelial and epithelial barrier integrity and phagocytic function were assessed in rats subjected to 20- or 40-min mesenteric ischemia and a 3-h reperfusion. The results showed that human serum albumin (125I-HSA) flux through the endothelial layer to the interstitial space increased as did 125I-HSA clearance from blood to the gut lumen and 131I-HSA flux from the gut lumen to the interstitial space in rats with I/R. E. coli adhering to microvilli, invading and passing into the microvessels, were noted on the small intestinal mucosa in animals subjected to 40-min ischemia and a 3-h reperfusion. Phagocytic function increased, especially in the small intestinal wall, lungs, liver, and spleen in the groups with I/R, correlating with the length of ischemia. The results imply that both endothelial and epithelial barrier integrity is impaired in the early phase after I/R and that the epithelial barrier more effectively restricts macromolecular leakage compared with the endothelial barrier. I/R impairs the intestinal barrier not only by causing tissue hypoxia but also by activating the phagocytic system and aggravating barrier damage, which finally may result in bacterial translocation and remote organ dysfunction. PMID- 10718379 TI - Role of endogenous nitric oxide in TNF-alpha and IL-1beta generation in hepatic ischemia-repefusion. AB - In the present study, we examined the role of nitric oxide (NO) in early-response cytokine production by using a rat model of hepatic ischemia-reperfusion (HI/R). The left and median lobes of the liver were subjected to 30 min of ischemia, followed by 4 h of reperfusion. Group I and II rats were sham-operated controls that received saline (vehicle) or N(W)-nitro-L-arginine methylester (L-NAME) (10 mg/kg, iv); group III and IV rats were subjected to HI/R and received vehicle or L-NAME (10 mg/kg, iv, 10 min before reperfusion), respectively. Administration of L-NAME to rats subjected to I/R resulted in a fourfold decrease in plasma NO levels, accompanied by a marked increase of plasma alanine aminotransferase (ALT) activity relative to group III. These changes in group IV were associated with elevation of superoxide generation in ischemic liver lobes by 2.1-fold and circulating leukocyte number by 1.42-fold, compared with group III. Normalized for expression of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) messenger ribonucleic acid (mRNA), expression of tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta) mRNA in ischemic liver of group IV was augmented by 207% and 175% compared with Group III. The expression of (iNOS) mRNA was also increased (223%) relative to group III. Moreover, in group IV, plasma TNF-alpha levels at 4 h of reperfusion and IL-1beta levels at 90 min and 4 h of reperfusion were significantly increased compared with group III. No statistically significant changes were observed between groups I and II in plasma ALT activity, plasma NO levels, circulating leukocyte counts, superoxide generation in the ischemic lobes of liver, and plasma TNF-a and IL-1beta concentrations. The observed enhancement of I/R injury by L-NAME is consistent with the hypothesis that endogenous NO down-regulates TNF-alpha and IL1beta generation, thereby decreasing HI/R injury. PMID- 10718380 TI - Cardiopulmonary and splanchnic blood flow during 48 hours of a continuous infusion of endotoxin in conscious pigs: a model of hyperdynamic shock. AB - Hyperdynamic shock can be modeled in pigs by chronic administration of a continuous, low-dose infusion of endotoxin. Lipopolysaccharide (LPS, E. coli 0111:B4, 80 ng/kg/min i.v.) initially resulted in a hypodynamic shock state with significant decreases in mean arterial pressure (112+/-3 mmHg at baseline to 94+/ 4 mmHg at 8 h) and cardiac index (5.35+/-0.32 L/min/m2 at baseline to 4.07+/-0.32 L/min/m2 at 4 h). Eight hours after the initiation of the LPS infusion, cardiac index rose to above baseline values (5.82+/-0.4 L/min/m2 at 8 h) and remained elevated for the duration of the 48-h study (6.54+/-0.39 L/min/m2 at 48 h). Similarly, systemic vascular resistance fell significantly by 8 h (1640+/-100 dyne sec cm(-5) at baseline to 1239+/-80 dyne sec cm(-5) at 8 h) and remained decreased for the duration of the study. Blood flow in major abdominal vessels, including the left renal artery, the cranial mesenteric artery, and the portal vein, paralleled changes in the cardiac index. Serum concentrations of tumor necrosis factor were increased after 2 h of LPS infusion, but did not remain elevated above baseline concentrations for more than about 4 h despite continuous LPS challenge. Concentrations of tumor necrosis factor did not differ between arterial and portal venous samples. This model of hyperdynamic shock should be useful to assess potential therapies for septic shock. PMID- 10718381 TI - Crystalloid or colloid resuscitation of uncontrolled hemorrhagic shock after moderate splenic injury. AB - Using a standardized moderate splenic injury (MSI) model of uncontrolled hemorrhagic shock, we studied the effect of vigorous crystalloid or colloid fluid resuscitation on the hemodynamic response and survival time in rats. The animals were randomized into 6 groups: group 1 (n = 8) sham-operated, group 2 (n = 10) MSI untreated, group 3 (n = 10) MSI treated with 41.5 mL/kg Ringer's lactate (large-volume Ringer's lactate [LVRL]), group 4 (n = 10) MSI treated with 5 mL/kg 7.5% NaCl (hypertonic saline [HTS]), group 5 (n = 10) MSI treated with 7.5 mL/kg hydroxyethyl starch (HES-7.5), group 6 (n = 10) MSI treated with 15 mL/kg hydroxyethyl starch (HES-15). After MSI, mean arterial pressure (MAP) in group 2 decreased from 105.0+/-5.6 to 64.0+/-12.7 mmHg (P < 0.001) after 60 min. Mean survival time was 157.4+/-28.9 min, and total blood loss was 24.0+/-5.4% of blood volume. LVRL infusion resulted in an early rise in MAP from 75.2+/-8.7 to 96.7+/ 9.0 mmHg (P < 0.01), which then rapidly dropped to 43.0+/-9.7 mmHg (P < 0.001) after 60 min. The mean survival time was 140.7+/-22.3 min, and total blood loss was 41.4+/-4.8% (P < 0.05). Total blood loss following HTS infusion was 24.7+/ 3.7%, and mean survival time was 177.5+/-18.9 min. HES-7.5 infusion was followed by bleeding of 25.6+/-5.1%, and mean survival time was 181+/-16.1. HES-15 infusion resulted in an increase in blood loss to 48.2+/-7.3% (P < 0.05), and mean survival time of 133.0+/-27.7 min. Large-volume Ringer's lactate (LVRL) or hydroxyethyl starch (HES-15) infusion in uncontrolled hemorrhagic shock after moderate splenic injury resulted in a significant increase in intra-abdominal bleeding, but survival time remained unchanged compared with untreated, small volume HTS-, or HES-7.5-treated animals. The hemodynamic response to large-volume crystalloid or colloid infusion was similar to moderate large-vessel injury. PMID- 10718382 TI - Endotoxin inducible transcription is repressed in endotoxin tolerant cells. AB - Stimulation of the human promonocytic cell line, THP-1, with endotoxin results in a rapid and transient increase in interleukin 1beta expression. Endotoxin pretreatment of THP-1 cells results in tolerance, characterized by decreased levels of endotoxin-induced interleukin 1beta expression due to decreased transcription of the interleukin 1beta gene. We hypothesized that tolerant cells could not activate transcription factors necessary to express the interleukin 1beta gene. This hypothesis was tested in tolerant THP-1 cells by using stable and transiently transfected reporter genes containing the interleukin 1beta promoter. We found decreased endotoxin-induced transcription of all reporter genes tested; however, individual transcription factors, such as NFkappaB, retain normal, CD14-dependent, nuclear translocation and DNA binding. Tolerance is specific for endotoxin, because phorbol ester is still able to activate transcription of the endogenous interleukin 1beta gene and transfected reporter genes. A constitutively active reporter gene that is not inducible by endotoxin is unaffected. We further show that nuclear extracts of tolerant cells show transcription inhibitor activity that is specific for promoter sequences of the interleukin 1beta gene. These results support a mechanism of endotoxin tolerance that is independent of transcription factor DNA binding and appears to be associated with the inability of DNA-bound transcription factors to activate transcription, perhaps through the activity of a repressor. PMID- 10718383 TI - CXC chemokine suppression of polymorphonuclear leukocytes apoptosis and preservation of function is oxidative stress independent. AB - Interleukin 8 (IL-8) and growth-related oncogene alpha (Gro-alpha) delay neutrophil apoptosis, which is thought to be important for the resolution of inflammation. We hypothesized that (IL-8) and Gro-alpha interfere with extracellular death receptor signaling or intracellular caspase activation to suppress neutrophil apoptosis. In addition, we sought to determine if prolonged neutrophil half-life was associated with preservation of function. Polymorphonuclear leukocytes (PMN) were cultured with IL-8 or Gro-alpha (0-100 ng/mL) in normoxia or hypoxia, and the extent of apoptosis was assessed by histology and TdT-mediated dUTP nick end labeling (TUNEL). Subsequently, to determine the role of apoptotic-associated receptors, PMN were cultured with IL-8 and neutralizing monoclonal antibody to Fas (CD95), TNFR55, and TNFR75. To establish the effect of IL-8 or Gro-alpha on pro-apoptotic caspase activity, the cleavage of specific colorimetric substrates was assessed. Functional changes in PMN included the capacity to produce superoxide anion and phagocytosis of Escherichia coli. At the 100 ng/mL dose, the addition of IL-8 and Gro-alpha maximally suppressed PMN apoptosis from 54% (untreated) to 5% and 6%, respectively. The addition of neutralizing antibodies to Fas, TNFR55, or R75 caused no change in IL-8 suppression of apoptosis. Caspase 3 activity was markedly suppressed at 24 h by the inclusion of either IL-8 and Gro-alpha. IL-8 and Gro-alpha-stimulated PMN released more superoxide anion and had an increased phagocytic index vs. control PMN. IL-8 and Gro-alpha suppress neutrophil apoptosis to a similar level that is not influenced by oxygen tension at high doses. The effect of IL-8 and Gro-alpha does not depend on activation of the Fas, TNFR55, or R75 receptor pathways but involves suppression of caspase 3 activity. IL-8 or Gro-alpha extends the functional half-life of neutrophils and may explain their role in disease states such as acute respiratory distress syndrome. PMID- 10718384 TI - Variceal bleeding and portal hypertension. AB - Within the short span of half a century, the treatment of variceal bleeding has become highly differentiated, with multiple treatment options. Pharmacological therapy with beta-blockers is well established for preventing the first variceal bleeding. The utility of adding a vasodilator to beta-blockers needs to be studied further. Octreotide is widely used as an adjuvant to standard endoscopic treatment to prevent variceal rebleeding, and the utility of this approach has been validated in several randomized controlled trials. Band ligation is well established, and its popularity has increased with the introduction of multiple ligation devices. The technical simplicity and safety of band ligation has sparked interest in using this technique for primary prophylaxis of variceal bleeding. However, randomized trials have not shown any advantage for band ligation over beta-blocker therapy, and the high variceal recurrence rate after band ligation may eliminate any theoretical advantage. A synchronous combination of band ligation and sclerotherapy has not been shown to improve the results of band ligation alone, but a metachronous approach using sclerotherapy to treat recurrent varices after band ligation has shown beneficial results. Histoacryl remains the best treatment option for gastric varices, but band ligation and loop ligation have shown promising results, and should be considered when Histoacryl is not available. Balloon-occluded retrograde transvenous obliteration is a new radiological modality for gastric varices, and one that sounds promising. TIPS is well established as an alternative to elective endoscopic treatment. Compared with endoscopic treatment, TIPS has been shown to improve the survival rate in one randomized trial. However, the cost and complications of TIPS have restricted its use. The use of endoscopic ultrasound for Doppler studies of blood flow in portal hypertension is currently investigational, but it may gain a role in selecting the optimal treatment approach for the individual patient. PMID- 10718385 TI - Diagnostic endoscopic retrograde cholangiopancreatography. AB - The importance of diagnostic endoscopic retrograde cholangiography (ERCP) has dramatically decreased owing to the development of less invasive techniques such as ultrasonography, computed tomography, endoscopic ultrasonography, and finally magnetic resonance cholangiopancreatography (MRCP). MRCP is becoming the gold standard in the diagnostic work-up of the pancreaticobiliary duct. However, MRCP cannot solve all the problems that occur, and still has inadequate resolution for small stones and tiny pancreatic and bile duct lesions. ERCP continues to be useful in difficult cases and when the diagnosis is uncertain, particularly when fluid collection and tissue sampling are necessary. However, several alternatives to sphincter of Oddi manometry have been proposed. Finally, ERCP is always the first step before endoscopic treatment, which in contrast to diagnostic ERCP is still widely used. PMID- 10718386 TI - Therapeutic biliary endoscopy. AB - An analysis of the recent medical literature on therapeutic biliary endoscopy shows continuing developments in technique, as well as an effort to obtain objective data on the long-term outcomes with different clinical applications. There are still too few studies analyzing costs, and these should be encouraged in the immediate future. PMID- 10718387 TI - Therapeutic pancreatic endoscopy. AB - A number of endoscopic interventions have expanded the range of treatment options in symptomatic pancreatic diseases. Early endoscopic retrograde cholangiopancreatography (ERCP) and endoscopic sphincterotomy (ES) appear to be beneficial in patients with severe acute biliary pancreatitis. Endoscopic pancreatic sphincterotomy can be safely performed with high technical success rates in patients with chronic pancreatitis. Routine additional biliary ES or drainage procedures are not apparently necessary. Stenting can be limited to the treatment of dominant pancreatic duct strictures. In patients with sphincter of Oddi dysfunction, temporary placement of pancreatic stents reduces the morbidity associated with ES. Transpapillary or transmural endoscopic drainage achieves resolution of pancreatic pseudocysts in the majority of selected patients. Drainage procedures with endoscopic ultrasound guidance can potentially expand the indications and reduce the procedure-related morbidity. Therapeutic endoscopy should be considered in symptomatic patients with pancreas divisum, as well as in selected children with pancreatic diseases. Most of the published studies on therapeutic pancreatic endoscopy have been conducted retrospectively. Additional prospective controlled trials are warranted to allow further evaluation of the impact of these methods on the clinical outcome in comparison with alternative treatment strategies. PMID- 10718388 TI - Virtual endoscopy--comparison with colonoscopy in the detection of space occupying lesions of the colon. AB - BACKGROUND AND STUDY AIMS: A new technique has been described which combines abdominal helical computed tomography (CT) scanning and virtual reality computer technology, known as virtual colonoscopy (VC); the reconstructed images provide a simulation of the interior of the colon as viewed by endoscopy. We compared VC with conventional colonoscopy in patients with suspected or known colonic neoplasia. PATIENTS AND METHODS: A total of 38 patients, in whom there was a high likelihood of colonic polyps or cancer, underwent a noncontrast helical CT scan of the abdomen and pelvis after regular colonoscopy bowel preparation. The images were reconstructed into a VC presentation and compared with the subsequent conventional colonoscopy in a blinded manner. RESULTS: Conventional colonoscopy identified a total of 24 polyps 5 mm or greater. VC correctly identified five of 13 polyps 5-9 mm in size, and ten of 11 lesions greater than or equal to 10 mm in diameter. The reasons for four missed lesions were identified as being secondary to a collapsed rectum in two patients and stool in the right colon in two patients. The sensitivity and specificity per patient of VC for lesions greater than or equal to 5 mm were 66.7% and 75.0% respectively, and for lesions greater than 1 cm were 90.0% and 82.1%, respectively. CONCLUSIONS: Virtual colonoscopy is feasible, well tolerated, and capable of detecting most lesions greater than 10 mm in diameter. This technique is continuing to be developed and warrants further evaluation as a diagnostic and screening tool in colorectal neoplasia. PMID- 10718389 TI - Improved sedation in diagnostic and therapeutic ERCP: propofol is an alternative to midazolam. AB - BACKGROUND AND STUDY AIMS: Adequate sedation of the patient is required for diagnostic and therapeutic endoscopic retrograde cholangiopancreatography (ERCP). The anesthetic propofol, with its shorter half-life, affording better control, offers an alternative to the benzodiazepine midazolam. The aim of this randomized, controlled, unblinded study was to compare prospectively the quality of sedation under propofol and midazolam in patients undergoing ERCP. PATIENTS AND METHODS: A total of 80 patients were randomized to sedation with propofol alone (n = 40) or midazolam alone (n = 40). Blood pressure, pulse, and oxygen saturation were measured. Midazolam was given by the endoscopist and titrated to the patients' response during ERCP, to a maximum dose of 15 mg per patient. In the propofol group an anesthetist was present to administer the propofol and to observe the patient. Standardized testing procedures (Steward score, Trieger test) were used to determine the length of postendoscopy recovery time. Efficacy of sedation was assessed by investigators and patients, using scoring systems. RESULTS: Complete ERCP and adequate sedation was possible in 80% of patients (32 out of 40) with midazolam, and in 97.5% of patients (39 out of 40) with propofol (P<0.01). The average propofol induction dose was 1.24 mg/kg body weight, with maintenance requiring a mean dose of 9 mg/kg body weight per hour, or the equivalent of 354 mg in total. The average dose of midazolam administered was 0.12 mg/kg body weight; the total dose averaged 8 mg. Recovery time in the propofol patients was significantly shorter (P<0.01). The investigators (P<0.01) and the patients (P<0.05) both judged the quality of sedation to be better in the propofol group. There were no differences in blood pressure, pulse, or oxygen saturation between the two groups. One patient in the propofol group (79 years old) suffered a protracted apneic phase accompanied by hypotension that was managed by manual ventilation and drug therapy, and led to no complications. CONCLUSIONS: Propofol proves to be an excellent sedative for ERCP and shows a shorter recovery time than midazolam. Because of the narrow therapeutic window, we recommend close patient monitoring. PMID- 10718390 TI - Synergistic sedation with low-dose midazolam and propofol for colonoscopies. AB - BACKGROUND AND STUDY AIMS: Patients undergoing colonoscopy are often sedated with benzodiazepines and long-acting opiates. Since low-dose midazolam also acts synergistically with short-acting propofol, we compared this synergistic sedation with a standard combination of midazolam and the opioid nalbuphine for colonoscopies. PATIENTS AND METHODS: A total of 79 patients presenting for colonoscopies were randomly assigned to the following protocols. Patients in group I (n = 32) received a median dose of 9 mg midazolam (interquartile range [IQR] 6 to 12); 20 patients (59%) needed additional nalbuphine (median 20 mg, IQR 10 to 20). Patients in group II (n = 47) received 2 mg midazolam and repeated injections of propofol (median 100 mg, IQR 53 to 145) with a maximal bolus of 50 mg. RESULTS: Patients treated with the synergistic sedation (group II) recovered remarkably sooner after the procedure compared with those in group I, with a median time to discharge of 17 minutes vs. 93 minutes (P<0.001). Of the patients treated with analgosedation (group I), 28 % were unable to take part in a reaction time measurement and attention awareness test 1 hour after the procedure. All patients treated with the synergistic sedation were able to participate (P=0.002), and performed better. Despite a lower proportion of complete amnesia, patients treated with synergistic sedation more often rated the procedure as comfortable (81% vs. 50 %). Quality of sedation from the point of view of the endoscopist, and cardiorespiratory parameters, were similar in both groups. CONCLUSIONS: Low-dose midazolam combined with propofol is an effective and economic alternative to benzodiazepine-based analgosedation. It is associated with a high degree of patient comfort and rapid recovery times, and has a potential cost benefit concerning nursing care and bed facilities. PMID- 10718391 TI - Mesalazine changes apoptosis and proliferation in normal mucosa of patients with sporadic polyps of the large bowel. AB - BACKGROUND AND STUDY AIMS: Regular intake of nonsteroidal anti-inflammatory drugs (NSAIDs) may reduce the occurrence of colorectal adenoma and carcinoma, possibly by inducing apoptosis and/or decreasing proliferation in colorectal epithelial cells. Mesalazine is widely used in the treatment of patients with inflammatory bowel disease, and well tolerated. We investigated its effect on apoptosis and proliferation of colorectal mucosa in 21 patients with sporadic polyps of the large bowel. PATIENTS AND METHODS: In total, 17 patients with sporadic colorectal polyps (> or = 5 mm) underwent polypectomy and biopsy of uninvolved mucosa before and after treatment with 1 g/d mesalazine for 1, 3, 7 or 14 days. Four additional patients served as untreated controls. Apoptotic index (AI) was measured by terminal deoxynucleotidyl transferase-mediated d-uridine triphosphate nick-end labeling (TUNEL) assay; proliferation index (PI) was measured by immunohistochemical examination with anti-Ki67 antibody. RESULTS: AI was significantly increased 1 and 3 days after initiation of treatment with mesalazine compared with controls (P= 0.0107 for the 1-day treatment group and P=0.0142 for the 3-day treatment group), and seemed to remain largely unchanged after longer treatment duration. Proliferation appeared to be decreased by mesalazine in all treatment groups, while proliferation in controls did not change (P=0.0107 for the 1-day treatment group and P= 0.0142 for the 3-day treatment group compared with controls. CONCLUSIONS: Mesalazine significantly induces apoptosis and decreases proliferation in colorectal mucosa in patients with sporadic polyps of the large bowel. This may be clinically relevant in that it may lower the rate of polyp recurrence after polypectomy, thereby possibly contributing to the chemoprevention of sporadic colorectal carcinoma. PMID- 10718392 TI - Endoscopic ultrasound-guided one-step transmural drainage of cystic abdominal lesions with a large-channel echo endoscope. AB - BACKGROUND AND STUDY AIMS: Transmual endoscopic drainage of peripancreatic fluid collections under endoscopic ultrasound (EUS) control has been reported. We evaluated a facilitated technique of one-step puncture and drainage using a new stenting device and a large-channel echo endoscope. PATIENTS AND METHODS: EUS guided transumural drainage of cystic lesions was attempted in six male patients. The drainage sites were duodenal in two instances and gastric in four. The lesions were pseudocysts, arising from chronic pancreatitis (n=2), and following acute pancreatitis (n=3), and one abscess. The median size of the pseudocysts was 50mm (range, 25 to 90). The punctures were carried out under direct EUS guidance, using a new echo endoscope with a 3.2-mm working channel. Transmural drainage was done using modified 7-F stents. Stents were inserted directly over a 1-mm puncture needle and their position was optimized using a 7-F pusher connected to the stent by a special construction. The stents were released by withdrawing the needle. RESULTS: EUS-guided one-step drainage was technically successful in all 6 patients. It was possible to position the stents as desired, regardless of the location, size or pathogenesis of the target lesions. There were no complications associated with the endoscopic interventions. All the lesions, including two with putrid contents, had collapsed by the day following drainage. The stents were removed after a median period of 2 weeks (range 2-12). The cysts had completely resolved in four patients at follow-ups of 3-13 months (range). The patient with the gastric abscess underwent gastrectomy 2 weeks after stent extraction, because of a coincidental adenocarcinoma at the cardia, the abscess being resolved. One patient with necrotizing pancreatitis, who refused surgical treatment, died because of septic complications. CONCLUSIONS: New large-channel echo endoscopes allow more aggressive endoscopic interventions to be carried out safely under direct EUS control. One-step drainage using the new 7-F stent-over-needle device was effective for six cystic lesions of variable origin. PMID- 10718393 TI - Virtual colonoscopy: time for some tough questions for radiologists and gastroenterologists. PMID- 10718394 TI - Colonoscopy--is sedation necessary and is there any role for intravenous propofol? PMID- 10718395 TI - An 18-year-old woman with recurrent pancreatitis. PMID- 10718396 TI - Management and endoscopic techniques in cases of ingestion of foreign bodies. PMID- 10718397 TI - Patient acceptance of virtual colonoscopy. PMID- 10718398 TI - Fast method of jejunal biopsy. PMID- 10718399 TI - Aspects of cell proliferation in oral epithelial dysplastic lesions. AB - There is a need for objective methods of assessment of oral epithelial precancerous lesions and reliable markers for the prediction of malignant change in these lesions. Cell proliferation was examined in 20 dysplastic lesions from the tongue and floor of mouth using bromodeoxyuridine (BrdU) and Ki-67, and a histological compartment analysis was performed. Half of a fresh biopsy from each case was incubated in BrdU for 15 min, the other half was routinely processed and used for Ki-67 analysis. Sections from each block were immunohisto chemically stained with antibodies against BrdU and Ki-67. Dysplasia was graded according to the method of Smith & Pindborg. The BrdU labelling index (LI) and the growth fraction (GF), assessed by the use of Ki-67, was quantified and expressed as units per millimetre basement membrane length (BL) and per 100 total nucleated cells (TNC). The mean LI/TNC was 10.87 (SD 3.65) and the mean LI/BL was 51.55 (SD 20.75). The mean GF/TNC was 26.66 (SD 17.78) and GF/BL was 157.07 (SD 125.84). The mean epithelial thickness was 229.09 microm (SD 104.73). The LI/BL correlated with the atypia score and with the GF/BL. The progenitor compartment sizes also correlated with the atypia scores. The BrdU labelling index provides a further objective measurement of oral epithelial dysplasia and the progenitor compartments were large, implying that basal cell hyperplasia is a significant component of the dysplasia. PMID- 10718400 TI - Association of aberrant p53 and p21(WAF1) immunoreactivity with the outcome of oral verrucous leukoplakia in Taiwan. AB - The expression of p53 and p21WAF1 in 53 oral verrucous leukoplakias (OVLs), mostly non-dysplastic lesions, was investigated to ascertain the role of such events in malignant conversion. Immunohistochemical analysis revealed aberrant p53 and p21WAF1 immunoreactivity in 51% (27 cases) and 75% (40 cases), respectively. After an average follow-up period of three and a half years, histopathological examination revealed that 22 (42%) cases had developed oral squamous cell carcinoma (OSCC), 14 (26%) cases had undergone recurrence, and 17 (32%) cases were free of disease. The oncogenic potential of this subset of premalignant lesions warrants attention. A significant difference in the frequency of OSCC progression/recurrence was noted in lesions bearing aberrant immunoreactivity of either p53 (93% vs 42%; P=0.00008) or p21WAF1 (80% vs 32%; P=0.002) in comparison with lesions without immunoreactivity. This study suggested that the aberrant immunoreactivity of p53 and p21WAF1 may represent important alterations of OVL and could affect the outcome of this lesion. PMID- 10718401 TI - Dysregulated expression of bcl-2 and bax in oral carcinomas: evidence of post transcriptional control. AB - A study was conducted to investigate the gene expression of bcl-2 and bax in oral squamous cell carcinomas. We used reverse-transcriptase-polymerase chain reaction (RT-PCR) to evaluate the expression of bcl-2 and bax mRNAs and the ratio of bcl 2/bax mRNA, and employed immunohistochemistry to investigate the bcl-2- and bax encoded proteins. It was observed that the expression level of bcl-2 mRNA or bax mRNA was not consistent with their protein level in some cases. Higher expression of bcl-2 mRNA and stronger immunostaining of bcl-2 protein were found in oral squamous cell carcinomas than in the adjacent histologically normal oral epithelium. These findings were more prominent in poorly differentiated carcinomas. No significant differences in bax mRNA and protein were observed between carcinomas and the adjacent histologically normal oral epithelium. However, poorly differentiated carcinomas showed very weak immunostaining for bax. The ratio of bcl-2/bax mRNA was higher in carcinomas than in the adjacent histologically normal oral epithelium, and higher ratios were seen in most of poorly differentiated carcinomas. This study supplies indirect evidence of post transcriptional control of bcl-2 and bax expression, and suggests that dysregulated expression of bcl-2 and bax may be related to the differentiation of oral squamous cell carcinomas. PMID- 10718402 TI - Comparative study of oral squamous cell carcinoma in Okinawa, Southern Japan and Sapporo in Hokkaido, Northern Japan; with special reference to human papillomavirus and Epstein-Barr virus infection. AB - In Okinawa, a subtropical island in Southern Japan, the incidence of oral squamous cell carcinoma is 1.5 times higher than that in mainland Japan. Sixty cases of oral squamous cell carcinoma from 1993 to 1996 in Okinawa and 42 cases over the same period in Sapporo were examined histologically. Human papillomavirus (HPV) and Epstein-Barr virus (EBV) were detected by polymerase chain reaction (PCR) amplification with primers specific for HPV and EBV. In situ hybridisations of the viruses were also carried out. In the case of Epstein-Barr virus, in situ PCR was also performed. Thirty-five (58.3%) Okinawan tumours were well-differentiated in type, but in Sapporo, 18 (42%) were of such type. In Okinawa, tumours of the mouth floor (10 cases, 16.7%) and oropharynx (12 cases, 20%) were frequently observed, whereas in Sapporo only five cases (12%) of each were found. HPV was demonstrated in 78% of Okinawan cases and 26.2% of Sapporon cases by PCR or non-isotopic in situ hybridisation (NISH). There were 76.6% (46 cases) of Okinawan and 38.1% (16 cases) of Sapporo cases positive for EBV by PCR. In only 12 Okinawan cases and 4 Sapporon cases, were positive signals demonstrated by in situ PCR on the cancer cells themselves. EBV was demonstrated in the large number of infiltrating lymphocytes, most of which were CD3+, and a few were CD19+. In Okinawa, HPV might be an important causative factor of oral squamous cell carcinoma and EBV a less important factor, whereas in Sapporo HPV and EBV might play only a small part in the aetiology of the tumour. PMID- 10718403 TI - Penetration of N-nitrosonornicotine (NNN) across oral mucosa in the presence of ethanol and nicotine. AB - The effects of ethanol concentrations of 5, 15, 20, 25, 27, 30 and 50% on the penetration of the tobacco-specific carcinogen, nitrosonornicotine (NNN), across porcine oral mucosa were examined using an in vitro perfusion system. Concentrations of ethanol of 25% and above significantly increased the permeability of oral mucosa to NNN, although this increase ceased with 50% ethanol, possibly due to a fixative effect. Nicotine is a consistent component of smoked and smokeless tobacco; the presence of 0.2% nicotine significantly increased the permeability of oral mucosa to NNN and 2% nicotine caused a further increase. Combined use of nicotine and ethanol significantly increased the penetration of NNN across oral mucosa over that of ethanol alone until the concentration of ethanol reached 50%. The results of this study suggest that the synergy between tobacco and alcohol in the etiology of oral cancer may be explained, at least in part, by the local permeabilizing effects of alcohol on the penetration of tobacco-specific (and other) carcinogens across oral mucosa. PMID- 10718404 TI - Isolation of C. dubliniensis from insulin-using diabetes mellitus patients. AB - The identification of the novel candidal species, C. dubliniensis, from oral swab studies of HIV-seropositive and -seronegative individuals has led to speculation that such a species may also reside in the oral cavity of other patient groups. In this study involvement of the newly described species, C. dubliniensis, was investigated in oral carriage and disease in 414 insulin-using diabetes mellitus patients. Seventy-seven percent of the diabetic patients carried candidal species in the oral cavity. C. albicans was the most commonly identified candidal species. This was followed by C. dubliniensis, which was isolated on 64 occasions. Colonisation with multiple candidal species was common, and C. dubliniensis was present in both carriage and disease states. Seven patients without signs of oral disease had C. dubliniensis isolated as the sole candidal species, while the same species was associated with various forms of oral candidosis in six patients. PMID- 10718405 TI - Bacillary angiomatosis affecting the oral cavity. Report of two cases and review. AB - Bacillary angiomatosis (BA) is an infectious disease characterized by proliferative vascular lesions; it mainly affects HIV-positive patients. Multiple cutaneous nodular lesions together with fever, chills, malaise, anorexia, vomiting and headache are the most important clinical manifestations. It may also involve the heart, liver, spleen, bones, lung, muscles, lymph nodes, central nervous system and other organs. Erythromycin, 500 mg four times a day, is the drug of choice. The importance of this lesion lies in its clinical and histological similarity with other diseases. Cutaneous and oral lesions of BA clinically resemble Kaposi's sarcoma (KS). Histopathologically, BA may be confused with angiosarcoma, pyogenic granuloma and epithelioid hemangioma. We report two HIV-positive men with BA lesions in the oral mucosa. Diagnosis was confirmed by biopsy and Warthin-Starry silver staining. PMID- 10718406 TI - Quality of life in head and neck cancer. AB - Because treatments for patients with cancer of the head and neck can have major impact on physical, social, and psychological function, the collection of quality of life (QOL) data in this group of patients is critical for our specialty. The University of Washington Quality of Life data have been collected and analyzed on three subsets of cancer patients. Information learned from these patients is summarized and strategies for future projects are outlined. PMID- 10718407 TI - Clinimetrics of Meniere's disease. AB - Two clinically useful measures to quantify the morbidity of Meniere's disease are the daily vertigo diary card and the Meniere's Disease Patient-Oriented Severity Index (MD POSI). The development and application of these outcomes instruments for patients with vertigo are discussed. PMID- 10718408 TI - Pediatric outcomes research: development of an outcomes instrument for tonsil and adenoid disease. AB - We describe the design and validation process of a disease-specific health status instrument for use in children with tonsil and adenoid disease. This instrument is reliable and valid and should be useful in future outcomes research on tonsil and adenoid disease. Some of the unique issues and challenges regarding outcomes research in pediatric patients are also discussed. PMID- 10718409 TI - Outcomes research and obstructive sleep apnea. AB - Sleep disorders in general and obstructive sleep apnea syndrome in particular are prevalent health problems. This report describe the methodology and findings from a prospective multicenter outcomes research study on obstructive sleep apnea syndrome that was conducted by the American Academy of Otolaryngology-Head and Neck Surgery. Other outcome measures available for outcomes research in obstructive sleep apnea syndrome are also summarized. PMID- 10718410 TI - Community-based outcomes research: the Project Solo experience. AB - One of the critical issues in outcomes research is physician participation. The lack of such participation is one of several factors that have made widespread adoption of outcomes measurement slow to materialize. This article focuses on the 6-year experience of Project Solo, a multi-site community-based outcomes research effort. PMID- 10718411 TI - Clinical outcomes in patients with chronic sinusitis. AB - Although sinusitis is one of the most common chronic illnesses in this country, relatively little is known about the effect of this disease or its treatment on quality of life. In a series of studies utilizing both disease-specific and general health instruments, patients with chronic sinusitis were found to have significant decrements in several subscales of general health, including bodily pain and social functioning (P<.05), compared with the general US population. Surgery for sinus disease was shown to result in significant reduction in both symptoms and medication usage (P<.05) after 12 months. These same outcome instruments can be used by health care providers to document clinical outcomes in similar populations of patients with chronic sinusitis. PMID- 10718412 TI - Objective and subjective outcomes in surgery for chronic sinusitis. AB - Endoscopic and radiological findings in patients with chronic sinusitis do not always correlate with symptoms. Studies suggest that postoperative endoscopic examination of the sinonasal cavity provides prognostic information regarding the potential for future episodes of sinusitis and the need for revision surgery. It is recommended that findings on nasal endoscopy be included in future outcomes studies on sinusitis. PMID- 10718413 TI - Outcomes assessment for chronic otitis media: the Chronic Ear Survey. AB - Analysis of outcomes in chronic otitis media has in the past been limited to audiological measurement or physical examination. The Chronic Ear Survey (CES) is an instrument to measure the impact of chronic otitis media and its treatment. The survey provides information regarding total ear-specific health, as well as subscore information regarding activity restriction, symptoms, and medical resource usage attributable to chronic otitis media Application of the CES to a prospective, nonrandomized series of 147 patients revealed that patients with chronic otitis media have significantly decreased CES scores compared with unaffected controls and that surgical intervention provides a significant improvement in ear-specific outcomes. PMID- 10718414 TI - Role of the specialty society in outcomes assessment: the development of COG*ENT. AB - The Cooperative Outcomes Group for ENT (COG*ENT), a clinical outcomes project of the American Academy of Otolaryngology--Head and Neck Surgery Foundation, was established to assess clinical outcomes of otolaryngological care of patients with two diseases, otitis media and rhinosinusitis. COG*ENT has produced an assessment tool with many potential applications. The development and early experience of the Foundation with this project are described, emphasizing the factors the authors believe are important for success. PMID- 10718415 TI - Corticosteroid treatment for idiopathic facial nerve paralysis: a meta-analysis. AB - OBJECTIVE: A meta-analysis was designed to evaluate facial recovery in patients with complete idiopathic facial nerve paralysis (IFNP) by comparing outcomes of those treated with corticosteroid therapy with outcomes of those treated with placebo or no treatment. STUDY DESIGN: Meta-analysis of prospective trials evaluating corticosteroid therapy for idiopathic facial nerve paralysis. METHODS: A protocol was followed outlining methods for trial selection, data extraction, and statistical analysis. A MEDLINE search of the English language literature was performed to identify clinical trials evaluating steroid treatment of IFNP. Three independent observers used an eight-point analysis to determine inclusion criteria. Data analysis was limited to individuals with clinically complete IFNP. The endpoints measured were clinically complete or incomplete facial motor recovery. Effect magnitude and significance were evaluated by calculating the rate difference and Fisher's Exact Test P value. Pooled analysis was performed with a random effects model. RESULTS: Forty-seven trials were identified. Of those, 27 were prospective and 20 retrospective. Three prospective trials met the inclusion criteria Tests of heterogeneity indicate the trial with the smallest sample size (RD = -0.19; 95% CI, -0.58-0.20), to be an outlier. It was excluded from the final analysis. Analyses of data from the remaining two studies indicate corticosteroid treatment improves complete facial motor recovery for individuals with complete IFNP. Rate difference demonstrates a 17% (990% CI, 0.01-0.32) improvement in clinically complete recovery for the treatment group based on the random effects model. CONCLUSIONS: Corticosteroid treatment provides a clinically and statistically significant improvement in recovery of function in complete IFNP. PMID- 10718416 TI - Adenoidectomy with laser or incisional myringotomy for otitis media with effusion. AB - OBJECTIVE: To compare the effectiveness of CO2 laser myringotomy to incisional myringotomy at the time of adenoidectomy for refractory otitis media with effusion (OME). STUDY DESIGN: Controlled retrospective consecutive case series. METHODS: All children undergoing myringotomy and adenoidectomy for OME in the spring of 1999 had 1.7-mm-diameter perforations created in their tympanic membranes using a CO2 laser and conventional microslad. Their ears were evaluated at first postoperative visit (mean, 16.65 days after surgery) by a validated otoscopist to determine the presence or absence of perforations and middle ear effusions. These patients were compared with historical controls comprising all children undergoing incisional myringotomy and adenoidectomy in 1998. A chi2 analysis was performed to compare the results of these two myringotomy techniques. RESULTS: Twenty-three children (39 ears) underwent laser myringotomy and adenoidectomy in 1999, compared with 26 children (48 ears) who underwent incisional myringotomy and adenoidectomy in 1998. In the laser myringotomy group, 8 of the 39 ears had a persistent opening at first follow-up; 4 of the 39 ears showed evidence of effusion. In the incisional myringotomy group, all 48 ears had healed; 7 of these ears showed evidence of effusion. CONCLUSION: Myringotomies created using the CO2 laser are more likely to be patent at first postoperative visit than those made with incisional technique (P < .01). However, this prolonged middle ear ventilation does not significantly decrease the prevalence of effusion (P > .1). PMID- 10718417 TI - Early arytenoid adduction for vagal paralysis after skull base surgery. AB - OBJECTIVES: To evaluate the efficacy of early arytenoid adduction in the management of vagal paralysis after skull base surgery. STUDY DESIGN: Retrospective evaluation at a tertiary care skull base center. METHODS: Aggressive surgical management of skull base lesions has become increasingly popular owing to advances in surgical technique and intraoperative monitoring. Temporary and permanent lower cranial neuropathies occur frequently, especially after the surgical management of lesions involving the vertebrobasilar system and the jugular foramen. An injury to the proximal vagus nerve is usually associated with dysphonia and swallowing dysfunction. An early arytenoid adduction has been employed in 26 patients with a vagal paralysis after skull base surgery. Most commonly, the neurosurgical patient underwent an arytenoid adduction under general anesthesia on postoperative day 2. RESULTS: Videostroboscopy after arytenoid adduction demonstrated 76% of patients had complete glottic closure. Of those with inadequate glottic closure, all demonstrated a well-medialized posterior glottis with a persistent anterior glottal gap. These patients were easily treated with a secondary type I thyroplasty under local anesthesia with sedation resulting in complete glottic closure. Despite excellent voice outcomes, 66% of these patients had dysphagia requiring enteral feedings for nutritional support. CONCLUSIONS: An early arytenoid adduction is an excellent medialization technique that can be performed safely in the early postoperative period under general anesthesia after skull base surgery. PMID- 10718418 TI - Technical modifications of the latissimus dorsi pedicled flap to increase versatility and viability. AB - OBJECTIVE: Describe the elevation and insetting of the pedicled latissimus dorsi musculocutaneous flap. Review history of this flap's evolution and personal series of 68 consecutive cases since 1984. STUDY DESIGN: Retrospective review. SETTING: Tertiary, referral, academic center. METHODS: Retrospective review of 68 consecutive patient records in which the pedicled latissimus dorsi musculocutaneous flap was used to reconstruct head and neck defects. Overall flap survival and postoperative complications were used as outcome measures. RESULTS: Thirty-one women and 37 men underwent reconstruction with the latissimus dorsi pedicled flap between 1984 to 1998. The mean age was 61 years. Sixty-three cases followed postoncologic ablation and 5 cases addressed traumatic tissue loss. Forty-three patients had prior radiotherapy and 26 patients had undergone prior reconstructive surgery. The overall flap survival rate was 67/68 (98.5%), with one case of complete flap necrosis. Six cases of partial flap necrosis occurred. There were 8 other minor complications including fistula, wound dehiscence, hematoma and cerebrospinal fluid accumulation. Fifty-six donor sites were closed primarily resulting in 2 dehiscences and 17 seromatas. Three of 12 skin grafts to the donor sites were compromised. CONCLUSION: The excellent flap survival rate (98.5%) is the result of proper patient selection and adherence to three technical fundamentals: skin paddle design, pedicle dissection, and pedicle stabilization. The minimal donor site morbidity also demonstrated in this series supports the continued use of the latissimus dorsi pedicled flap for reconstruction of head and neck defects. PMID- 10718419 TI - Increased female authorship in otolaryngology over the past three decades. AB - OBJECTIVE: To identify changing trends in female authorship and publication in the otolaryngology literature. METHODS: All articles published in the four major otolaryngology journals in each of the years 1978, 1988, and 1998 were reviewed. The authorship panel of each article was examined for number of authors, gender, educational degree category, and subspecialty area of publication. Data were analyzed for trends in female authorship and the association of gender with the other design variables. RESULTS: A total of 2,463 articles were analyzed. The average percentage of female authorship increased from 4.1% in 1978 to 8.7% in 1988 and 12.4% in 1998, and the percentage of articles with a female "first author" increased from 3.2% to 7.4% and 11.4% for the same years, respectively. Each of these increases was statistically significant (P < .001). The weighted rank of female authorship also increased from 0.063 to 0.164 and 0.243 for the same years, respectively (P < .001). With respect to subspecialty publication, women were first authors of 14.7% of articles concerning pediatric otolaryngology but accounted for 9.9% or less of the first authors in the other subspecialty areas (P < .001). Female authors were also much more likely to be nonphysicians (P < .001) than men. CONCLUSIONS: There has been a significant trend toward increased female authorship in the otolaryngology literature. A significant portion of this is accounted for by nonphysician female authors, and female authorship tends to be concentrated in pediatric otolaryngology. PMID- 10718420 TI - Otolaryngological manifestations of velocardiofacial syndrome: a retrospective review of 35 patients. AB - OBJECTIVE: Because many patients with velocardiofacial syndrome (VCFS) are first examined by otolaryngologists for ear or speech problems before being diagnosed with VCFS, we describe a series of patients with this genetic disorder, which is associated with multiple anomalies, including velopharyngeal insufficiency, cardiac defects, characteristic facial features, and learning disabilities. STUDY DESIGN: We retrospectively analyzed the medical charts and available nasoendoscopic observations for 35 patients who were diagnosed with VCFS and who had a microscopic deletion in chromosome 22q11 as shown by DNA probe and fluorescence in situ hybridization. RESULTS: For most patients, the medical chart documented cardiac anomalies, velopharyngeal insufficiency with hypernasal speech, and characteristic facial features including nasal, auricular, craniofacial, and ocular abnormalities. Incidence of middle ear infection with associated conductive hearing loss was also high and necessitated early placement of pressure equalization tubes. Some patients were treated with adenoidectomy for chronic otitis media; consequently, velopharyngeal insufficiency and hypernasal speech worsened. Nasoendoscopic examination as documented in the medical chart showed occult cleft palate, a small adenoid pad, and pulsation in the muscular wall. CONCLUSION: Otolaryngologists have an important role in diagnosis and treatment of persons with VCFS and therefore should familiarize themselves with the typical history and most frequent head and neck manifestations of this syndrome. PMID- 10718421 TI - A pilot study of autofluorescent endoscopy for the in vivo detection of laryngeal cancer. AB - OBJECTIVES: To determine the advantage of autofluorescent endoscopy for the identification of laryngeal cancer. STUDY DESIGN: This is a prospective, multicenter clinical study. We investigated whether autofluorescent endoscopy using the Lung Imaging Fluorescent Endoscopy (LIFE)-Lung System (Xillix, Olympus) is capable of identifying early cancer of the larynx, especially in comparison with conventional white-light endoscopy and microscopic laryngoscopy. Benign lesions as well as microinvasive and invasive squamous cell carcinoma of the larynx were investigated. For logistic reasons and because of the pilot character of this study, the number of patients was limited. METHODS: Sixteen patients having 24 laryngeal lesions of both benign or malignant character were subsequently examined by autofluorescent endoscopy, white-light endoscopy, and microscopic laryngoscopy. Based on optical appearance, and for each method separately, the lesions were classified as malignant or not. The visual results were documented and histologically verified. RESULTS: The sensitivity of autofluorescent endoscopy for laryngeal cancer detection was more than 90% and therefore higher than that of white-light endoscopy and microscopic laryngoscopy. However, as far as laryngeal cancer is concerned, the specificity of autofluorescent endoscopy was very low. Many of the false-positive results were due to inflammation, hypervascularization, and edema. CONCLUSION: Autofluorescent endoscopy is advantageous only in the hands of an experienced ENT specialist. Although it does not replace the combination of white-light endoscopy and a critical evaluation of the clinical symptoms of the individual disease, it can profitably complement them. Autofluorescent endoscopy can help in determining whether microscopic laryngoscopy performed with general anesthesia should be recommended urgently to the patient. Microscopic laryngoscopy remains the best method for the identification of malignant lesions, if it is combined with obtaining taking multiple biopsy specimens. Confirmation of the results of this pilot study with a larger series of patients is desirable. PMID- 10718423 TI - Percutaneous magnetic resonance image-guided biopsy and aspiration in the head and neck. AB - OBJECTIVE/HYPOTHESIS: To test the hypotheses that 1) magnetic resonance imaging (MRI)-guided biopsy and aspiration with an open 0.2-T system (Magnetom Open, Siemens, Erlangen, Germany) in the head and neck is feasible and successful and 2) procedure times can be sufficiently short to be well tolerated by the patient. METHODS: Sixty-one MRI-guided procedures were performed in 47 patients (ages, 6 mo-88 y) in the head and neck, including the mucosal sites and masticator and parapharyngeal spaces (n = 23), parotid space (n = 6), submandibular space (n = 2), cervical vertebral column/paraspinal tissues (n = 8), skull base (n = 3), larynx or hypopharynx (n = 3), and infrahyoid nodal chains and surrounding tissues (n = 16). A clinical C-arm imaging system was used, supplemented by an in room radiofrequency-shielded liquid crystal monitor, rapid gradient echo sequences for needle guidance, and MRI-compatible anesthesia, monitoring, and surgical lighting equipment. Tissue sampling included fine-needle aspiration (n = 58) and cutting-needle core biopsy (n = 27), with 24 patients undergoing both procedures. Procedures were evaluated for success of needle placement, procedure time, and complications. RESULTS: Successful needle placement was accomplished in all cases without complication, with tissue sufficient for pathological diagnosis obtained for all but five patients with an average of 2.1 passes per patient. For fine-needle aspiration, average instrument time was 7.8 minutes per pass, and average cutting-needle core biopsy time was 9.2 minutes. CONCLUSIONS: Interactive MRI guidance for needle biopsy and aspiration of deep head and neck lesions is feasible, successful, and safe. Procedure times are sufficiently short to be well tolerated by the patient. PMID- 10718422 TI - Identification of genes overexpressed in head and neck squamous cell carcinoma using a combination of complementary DNA subtraction and microarray analysis. AB - OBJECTIVES/HYPOTHESIS: To discover unique genes specific for squamous cell carcinoma of the head and neck for eventual development as tumor markers and vaccine candidates. STUDY DESIGN: Molecular biological analysis of fresh-frozen head and neck squamous cell cancer (HNSCC). METHODS: A subtractive library was made from two HNSCC and six normal tissues using a polymerase chain reaction (PCR)-based approach. Genes from this library were PCR amplified and placed on a microarray glass slide. RNA was prepared or obtained from 16 fresh-frozen HNSCC and 22 normal tissue sources. Fluorescent probes were made from the polyA+ RNA derived from the tumor and normal tissues. The probes were hybridized to the glass slides and excited by a tuneable laser. One hundred seven of the genes showing the highest differential fluorescence value between tumor and normal tissue were identified by sequence analysis. RESULTS: Thirteen independent genes were found to be overexpressed in tumor tissues. Of these, nine were previously known: keratins K6 and K16, laminin-5, plakophilin-1, matrix metalloproteinase-2 (MMP), vascular endothelial growth factor, connexin 26, 14-3-3 sigma, and CaN19. The level of polyA+ RNA of these genes in the tumors was significantly different from the levels in normal tissue (P < .05). Four previously unidentified genes were also discovered to have increased expression in tumor tissue. Comparing the total tumor group (n = 16) to the normal group (n = 22), only one of these genes showed significant overexpression. CONCLUSION: We report the identification of nine known genes that are significantly overexpressed in HNSCC as compared to normal tissue using subtractive and microarray technology. In addition, we present four previously unidentified genes that are overexpressed in a subset of tumors. These genes will be developed as tumor markers and vaccine candidates. PMID- 10718424 TI - Assessment of carotid artery invasion in patients with head and neck cancer. AB - PURPOSE: Define radiological and histological features in which patients with head and neck cancer would benefit from a carotid artery resection. Resection of the carotid artery has been advocated for local control of advanced squamous cell carcinoma of the head and neck. To provide appropriate preoperative counseling and optimize the utilization of resources, the criteria for patient selection has to be defined. METHODS: Thirty-four patients underwent carotid artery resection based on the clinical impression of tumor fixation. Eighteen and 28 patients were evaluated using computed tomography (CT) and histological analysis, respectively. The distance between the tumor cells and external elastic lamina was measured. CT scans were examined to determine the circumference of tumor attachment around the carotid artery. RESULTS: Clinical assessment predicted tumor within 1.8 mm of the carotid artery in 68% of cases. The overall survival for patients with tumor greater than 1.8 mm (N = 9) was better than that of patients with less (N = 19) than 1.8 mm (33.3% vs. 5.3%; median 24 versus 9 mo, P = .0899). Three of six patients (50%) with less than 180 degrees circumference tumor attachment had tumor within 1.8 mm from the external elastic lamina. Eight of twelve patients (67%) with tumors encompassing more than 180 degrees of the artery wall had tumor within 1.8 mm from the external elastic lamina. The overall survival rates for patients with tumor attachment greater and less than 180 degrees were 8.3% and 33%, respectively. DISCUSSION: Tumor invasion into the carotid artery was the strongest predictor of outcome. Clinical assessment was as predictive as CT for tumor invasion. If tumor involvement of the carotid artery is less than 180 degrees, peeling the tumor is an alternative to carotid artery resection. PMID- 10718425 TI - Telomerase activity is upregulated in laryngeal squamous cell carcinoma. AB - OBJECTIVE/HYPOTHESIS: The immortalizing enzyme telomerase has been linked to carcinogenesis and is being targeted as a novel molecular marker. This study investigated telomerase expression in patients with laryngeal squamous cell carcinoma and correlated telomerase activity with conventional prognostic parameters. STUDY DESIGN: A consecutive series of patients with laryngeal squamous cell carcinoma undergoing surgical salvage for persistent or progressive disease after failed radiation therapy. METHODS: Twenty patient samples of laryngeal squamous cell carcinoma and 20 adjacent histologically normal mucosal samples were assayed using the telomeric repeat amplification protocol (TRAP) method for detection of telomerase activity. The leukemic cell line, K562, acted as a positive control and the human fibroblast line, Hs21Fs, as a negative control. A sample was classified as telomerase positive when an RNase-sensitive hexameric repeat ladder was observed. Absence of laddering was considered a negative result. RESULTS: Seventeen of 20 (85%) tumor samples and 4 of 20 (20%) adjacent histologically normal samples were telomerase positive. No statistically significant difference was observed when densitometric readings were compared by T category, tumor grade, or site (by ANOVA). CONCLUSIONS: Although telomerase activity is present in laryngeal cancer, levels of activation do not correlate with conventional parameters used for prognostication. Our study indicates that the marker may be a useful adjunctive method in the diagnosis of malignancy after radiation failure. PMID- 10718426 TI - Screening for distant metastases in patients with head and neck cancer. AB - OBJECTIVES: The detection of distant metastases at initial evaluation may alter the selection of therapy in patients with head and neck squamous cell carcinoma (HNSCC). In this study the value of screening for distant metastases is evaluated. STUDY DESIGN: Retrospective analysis. METHODS: The results of screening for distant metastases were retrospectively analyzed in 101 consecutive HNSCC patients with high-risk factors who were scheduled for major surgery. All patients had computed tomography (CT) scan of the thorax, bone scintigraphy, examination of the liver by ultrasound and/or CT scan, and blood tests. RESULTS: Distant metastases were found in 17% of the patients. Patients with four or more clinical lymph node metastases or low jugular lymph node metastases had the highest incidence of distant metastases (33%). CT scan of the thorax detected in 12 patients, lung metastases; in 4, mediastinal lymph node metastases; and in 2, primary lung tumors. Bone scintigraphy detected in four patients bone metastases; in all four patients lung or mediastinal lymph node metastases were also found. Ultrasound and/or CT scan of the liver revealed one patient with metastases. Blood tests did not show any significant difference between patients with or without bone or liver metastases. CONCLUSIONS: Screening in patients with three or more lymph node metastases, bilateral lymph node metastases, lymph nodes of 6 cm or larger, low jugular lymph node metastases, locoregional tumor recurrence, and second primary tumors revealed distant metastases in 10% or more. CT scan of the thorax is currently the single most important diagnostic technique for screening of distant metastases. PMID- 10718427 TI - Peripheral blood T-lymphocyte and monocyte function and survival in patients with head and neck carcinoma. AB - OBJECTIVES/HYPOTHESIS: To determine if the T-lymphocyte and monocyte functions of peripheral blood mononuclear cells (PBMCs) from patients with head and neck squamous cell carcinomas (HNSCC) are predictive factors for outcome. STUDY DESIGN: A prospective study describing the outcome, as to total survival and death by disease after at least 40 months observation, of 81 previously untreated male HNSCC patients in relation to PBMC T-lymphocyte and monocyte function. METHODS: T-lymphocyte mitogenesis and the cytokine level in culture supernatants of PBMC as well as monocytes were analyzed. These parameters were related to survival by Cox regression and Kaplan-Meier survival analysis. RESULTS: When patients with high versus low T-lymphocyte mitogen-stimulated proliferations were compared, a decreased proliferation was seen to be related to worse outcome. The predictive value of T-lymphocyte proliferation was shown to be an independent prognostic factor when adjusted for stage and stratified for anatomic location in survival analysis. The predictive value was also retained when the serum values of the major serum proteins and hormones and scores based on the smoking and alcohol history were added to the survival analysis with lymphocyte proliferation. Supernatant levels of gamma-interferon, interleukin (IL)-2, or IL 4 in PBMC cultures were not related to outcome. Monocyte function measured by endotoxin-stimulated IL-1beta, IL-6, IL-12, and tumor necrosis factor-alpha secretion did not relate to outcome of the patients. CONCLUSION: The PBMC T lymphocyte-stimulated proliferation is an independent prognostic factor for male HNSCC patients. PMID- 10718428 TI - Influence of tobacco and alcohol on the stage of laryngeal cancer at diagnosis. AB - OBJECTIVE: This study sought to examine the influence of cigarette smoking and alcohol consumption on the stage of laryngeal cancer at diagnosis. STUDY DESIGN: A retrospective review of 499 laryngeal cancer patients diagnosed between 1978 and 1997 was conducted. METHODS: Parameters that included smoking history, history of alcohol consumption, and the tumor stage and location at diagnosis were analyzed using the proportional odds model, correlation coefficient, and Student t test. RESULTS: Three hundred sixteen patients met the inclusion criteria, and 180 (56%) had advanced-stage disease at the time of presentation. The statistical model demonstrated a small but significant relationship between tobacco and alcohol on the stage of laryngeal cancer at diagnosis. Patients diagnosed with an advanced-stage tumor (stage III or IV) smoked a significantly greater amount and were more likely to be heavy drinkers than those diagnosed with a localized laryngeal cancer. CONCLUSIONS: Our results demonstrate that for every incremental increase in pack years of smoking, there is a small but measurable increase in the odds that a patient's laryngeal cancer will be stage III or IV at diagnosis. Likewise, being a "heavy" drinker as opposed to a "social" drinker raises the likelihood of an advanced tumor. Given the preventable nature of these risk factors, the moderation of alcohol consumption and cessation of smoking is prudent advice that should be conveyed to all patients. PMID- 10718429 TI - Conservative surgery in T3-T4 pharyngolaryngeal squamous cell carcinoma: an alternative to radiation therapy and to total laryngectomy for good responders to induction chemotherapy. AB - OBJECTIVE: To evaluate the possibility of preservation of the larynx after neoadjuvant chemotherapy by performing a conservative surgery instead of total laryngectomy initially planned, in patients with previously untreated laryngeal and piriform sinus squamous cell carcinoma (SCC). STUDY DESIGN: Retrospective study. METHODS: A total of 115 patients treated at Tenon Hospital with induction chemotherapy from 1985 to 1995, all with initial indication of radical surgery, were available for the study. The clinical tumor response was evaluated after three cycles of chemotherapy. According to this response, to preserve laryngeal functions, some patients had a modification of the treatment initially planned: radiation therapy essentially for complete responders, and conservative surgery for some partial responders. RESULTS: Of 69 patients with laryngeal cancer, 14 were treated by partial laryngectomy and 19 by radiation therapy; of 46 patients with piriform sinus cancer, 8 were treated by partial surgery and 12 by radiation therapy; the other patients were treated as was initially planned (total laryngectomy with partial pharyngectomy). Overall survival rates, estimated by the Kaplan-Meier method, were not statistically different between the three treatment groups. The laryngeal functions were preserved in 54% of the patients who were alive at 3 years. CONCLUSION: This report is a retrospective study, but these results suggest the possibility of using conservative surgery, instead of initially planned total laryngectomy, for good responders to induction chemotherapy with a small residual tumor. PMID- 10718430 TI - Anterior distribution of human olfactory epithelium. AB - OBJECTIVES/HYPOTHESIS: To functionally investigate the distribution of the olfactory epithelium in humans by means of the electro-olfactogram (EOG) and anatomically located biopsy specimens. STUDY DESIGN: Prospective, nonrandomized, investigational. METHODS: Supra-threshold EOG recordings were made on 12 healthy, trained volunteers (6 women, 6 men; age range, 21-48 y). Vanillin was used as the stimulus, since it exclusively excites olfactory receptor neurons. The EOG was recorded with tubular electrodes that were placed using thin-fiber endoscopic guidance. Biopsy specimens were obtained of anterosuperior nasal cavity mucosa in the same regions as the positive EOGs in 15 smell-tested patients (7 women, 8 men; age range, 22-60 y) during routine nasal and sinus surgery. This biopsied tissue was histologically processed and stained for olfactory and neural proteins. RESULTS: Viable responses to EOG testing were obtained in 7 of 12 subjects. In these seven subjects it was possible to identify nine sites above or below the anterior middle turbinate insertion where EOGs were obtained. The biopsy results showed mature olfactory receptor neurons in this same area. CONCLUSIONS: Human olfactory epithelium appears to be distributed more anteriorly than previously assumed. PMID- 10718432 TI - The nose as bacterial reservoir: important differences between the vestibule and cavity. AB - The difference between the spectra of potential bacterial pathogens (PBPs) in the nasal vestibule and cavity has not been taken into account in clinical studies. PURPOSE: Since one can anticipate different flora in different kinds of mucosae, the authors compared bacterial species in the vestibule with those of the cavity. SUBJECTS AND METHOD: A total of 534 healthy male clerical workers in a downtown Lucerne office building were examined with fractionated swabs. RESULTS: PBPs, notably Staphylococcus aureus, were found in 412 subjects and surprisingly, differences in flora of the two sites were noted in 130 of them: PBPs were observed in the vestibule and not in the cavity in 85 of the subjects, and in 45 of them, the reverse was true. CONCLUSION: The practical implications of these findings are considerable regarding infection control in patients at increased infection risk. PMID- 10718431 TI - HLA-DRB1, -DQA1, and -DQB1 genotypes in patients with nasal polyposis. AB - OBJECTIVES/HYPOTHESIS: Genetic etiology is suspected in the development of nasal polyposis on the basis of familial aggregation. This study investigated whether there is an association between HLA-DRB1, -DQA1, and -DQB1 alleles and developing nasal polyposis. STUDY DESIGN: Data from 50 polypectomized patients were compared with data from 50 healthy randomly selected controls. Polyp score, possible asthma, aspirin sensitivity, and ASA triad were also recorded. METHODS: Genotyping of HLA-DRB1 alleles was carried out with the Dynal RELI SSO HLA-DRB, a direct DNA probe test that utilizes a polymerase chain reaction (PCR) and nucleic acid hybridization for the differentiation of 70 HLA-DRB alleles and 9 supertypes. For DQA1* genotyping PCR-RFLP (restriction fragment length polymorphism) was used, differentiating eight alleles. The DQB1* typing was carried out using a INNO-LiPA DQB PCR-reverse hybridization kit, allowing the discrimination of 30 alleles. RESULTS: People carrying the HLA-DR7-DQA1*0201, and -DQB1*0202 haplotype were found to have a two to three times higher odds ratios (ORs) for developing the disease, compared with controls. Patients with ASA triad carried the above-mentioned DR7 allele with the linked alleles significantly more often (P < .001). Subjects carrying HLA-DR5 allele and the linked alleles had lower odds ratio values. CONCLUSION: These results underline that allergy is not conditional for the formation of nasal polyps as thought before. Nasal polyposis associated with asthma and aspirin sensitivity is probably a unique form of nasal polyps. The authors plan further investigations in this field. PMID- 10718433 TI - Hypoxia depresses nitric oxide output in the human nasal airways. AB - OBJECTIVES: The role of oxygen in the nasal air on nasal nitric oxide (NO) output was studied in 13 adult volunteers. METHODS: Nasal NO was measured while air containing oxygen (0%-100% in nitrogen) was aspirated through the nasal airway before and after the topical application of xylometazoline. RESULTS: The mean nasal NO output of the untreated nose was 507.8 +/- 161.9 nL/min (mean +/- SD) when 21% oxygen was aspirated through the nasal cavities in series and remained unaltered by 100% O2 (P = .79). Below 10% oxygen the reduction in nasal NO output correlated positively and significantly with the decrease in oxygen concentration (r2 = 0.14). NO output was 245.2 +/- 153.4 nL/min at 0% oxygen, a significant decline from 21% oxygen (P < .0001). Nasal vasoconstriction induced by xylometazoline and alterations in the blood oxygen content by a maximal breath holding or breathing 100% oxygen did not alter nasal NO in hypoxia (P = .41). CONCLUSIONS: Nasal NO output is markedly depressed in hypoxia and is oxygen dependent at concentrations of less than 10%. Approximately 50% of nasally generated NO is produced from oxygen in nasal air or regulated by it. PMID- 10718434 TI - Role of laryngeal movement and effect of aging on swallowing pressure in the pharynx and upper esophageal sphincter. AB - OBJECTIVES: Describe contribution of laryngeal movement to pressure changes at the upper esophageal sphincter (UES) and the effect of aging on the swallowing function. STUDY DESIGN: Manofluorography on 56 nondysphagic adults divided into three age groups: the 21- to 31-year-old group (n = 32), the 61- to 74-year-old group (n = 12) and the 75- to 89-year-old group (n = 12). Analyses of the bolus transit time, the amplitudes and durations of pharyngeal pressures, the timing of a pressure fall at the UES and the laryngeal movements. METHODS: Intraluminal strain-gauge sensors recorded pressure changes in the oropharynx, hypopharynx and the UES. Motion pictures of the videotapes were fed into a personal computer, and movements of the hyoid bone were measured in both the horizontal and vertical directions as an indication of laryngeal movement. RESULTS: In 26- and 70-year old men with calcification of the thyroid cartilage, it was determined that the larynx and hyoid bone moved in consonance until the end of the rapid hyoid movements in both the superior and anterior directions. In the 21- to 31-year-old group, the magnitude of the pressure fall at the UES was maximal before or almost at the same time as the bolus arrival, in preparation for smooth passage of the bolus from the pharynx to the esophagus. The rapid superior movements of the hyoid bone started significantly early as compared with its anterior movements (P = .0001). The rapid anterior movements of the hyoid bone started simultaneously with the pressure fall at the UES. In the elderly, all segmental transit times were significantly increased. The timing of the pressure fall at the UES was significantly delayed and the UES pressure reached its minimum value after arrival of the bolus at the UES. The minimum pressure at the UES increased to a significantly positive value. The rapid anterior movements of the hyoid were significantly delayed, suggesting that this delay causes the delay in the pressure fall at the UES. CONCLUSIONS: The rapid superior and anterior movements of the hyoid bone are considered to start at the same time as those of the larynx. In the young group, it is suggested that superior laryngeal movement protects the lower airway prior to the anterior laryngeal movement, causing the pressure fall at the UES to enable the passage of a bolus into the UES. In the elderly, smooth passage of the bolus from the pharynx to the esophagus is hindered and the system that prevents aspiration is rendered inefficient by changes in the swallowing pressures and laryngeal movements with aging. PMID- 10718435 TI - Two-dimensional analysis of vocal fold vibration in unilaterally atrophied larynges. AB - OBJECTIVE: Considerable inconsistencies regarding the vibratory pattern of the vocal fold among patients with unilateral vocal fold paralysis (UVFP) have been reported. These differences are derived from differences in the position, stiffness, and atrophy of the paralyzed vocal fold and other factors among patients. The purpose of this study was to assess the effects of unilateral atrophy of the vocal fold on vocal fold vibration. METHODS: Seven excised canine larynges were studied. The unilateral recurrent laryngeal nerve was severed to cause vocal fold atrophy in four of the seven. The lateral and vertical displacements were monitored simultaneously with photoglottography and a laser Doppler vibrometer, respectively. Videostroboscopy of each larynx was also performed before and after the experiment to translate photoglottographic output into absolute lengths. Atrophy of the unilateral vocal fold was confirmed histologically. RESULTS: The lateral amplitude was significantly greater than the vertical amplitude in all larynges. The Lissajous trajectories in the normal larynges were shaped like a reverse crescent. Vibration in the unilaterally atrophied larynges was periodical and symmetrical in phase when the thyroid ala on the atrophied side was pressed medially. The lateral and vertical amplitudes on the atrophied side were significantly greater than those on the normal side. The Lissajous trajectories differed from those of the normal larynges. CONCLUSIONS: In the absence of a prephonatory glottal gap, periodical vibration occurs in unilaterally atrophied larynges and the amplitude of vibrations of the atrophied vocal fold is greater in the lateral and vertical directions than that of the normal fold. This implies that phonosurgical procedures aiming at closure of the prephonatory glottal gap may have a beneficial effect on hoarseness in UVFP patients, although displacements of the vocal folds during vibration are not symmetrical. PMID- 10718436 TI - Lateral thyrotomy approach on the paraglottic space for laryngocele resection. AB - OBJECTIVE: To report on the results of using a lateral thyrotomy approach on the paraglottic space to gain greater access for laryngocele resection under direct vision. STUDY DESIGN: A 26-year prospective and retrospective study. The study was conducted on 10 adult patients (5 men and 5 women) who had laryngocele of varying size on the paraglottic space. Six of the patients had internal laryngocele and four had exteriorized laryngocele. Five laryngoceles were left sided, three were right-sided, and two were bilateral. METHODS: A V-shaped, full thickness thyroid lamina resection with the triangle base at the superior border and the apex at a point midway of the thyroid lamina vertical extent was performed. RESULTS: A V-shaped lateral thyrotomy made exposure to the paraglottic space possible for direct submucosal laryngocele dissection. This approach has presented no complications to date. Postoperative minor edema or hematoma was found in the aryepiglottic and ventricular folds, but this disappeared within a few days. There was no recurrence; the minimum follow-up was 1 year. CONCLUSION: The triangular lateral thyrotomy approach provided access to the paraglottic space and superb visibility for resection of laryngocele of any size under direct vision, thus avoiding recurrence, morbidity, and complications. PMID- 10718437 TI - Characterization of an experimentally induced inner ear immune response. AB - OBJECTIVE: To determine the effects of a sterile immune response on the structure and function of the cochlea. METHODS: An immune response was created in guinea pigs by systemically sensitizing the animals to keyhole limpet hemocyanin and subsequently challenging the inner ear with the protein. Animals were allowed to survive for 1 to 5 weeks, after which the cochlea was evaluated histologically. Hearing was measured by auditory brainstem response before the inner ear challenge, during the survival period, and prior to sacrifice. RESULTS: Inflammatory cells infiltrated the cochlea from the circulation. Surface preparations and plastic sections of the organ of Corti 1 and 2 weeks after the initiation of the inflammation demonstrated degeneration of the sensory and supporting cells in cochlear turns containing inflammatory cells. Good preservation of structures was seen in the more apical cochlear turns with little or no inflammatory cells. In cochleas from animals that survived 5 weeks, most of the infiltrated cells were cleared after undergoing apoptosis and the inflammatory matrix in the scala tympani began to calcify. Hearing loss was moderate to severe depending on the amount of inflammation. CONCLUSION: Although in general the immune response serves to protect an organism from infection, these results demonstrate that bystander injury associated with local immune responses in the cochlea, an organ incapable of regeneration, causes permanent cochlear destruction and hearing loss. PMID- 10718438 TI - Hearing loss in the nonocular Stickler syndrome caused by a COL11A2 mutation. AB - OBJECTIVE: Evaluation of hearing impairment as a feature of the nonocular Stickler syndrome (type II) linked to COL11A2. STUDY DESIGN: Family study. METHODS: General, orthopaedic, ophthalmologic, and otorhinolaryngologic examinations were performed on 15 affected persons in a Dutch family. Audiograms were obtained and/or retrieved from elsewhere. Cross-sectional and longitudinal analyses were conducted on the hearing threshold (sensorineural component) in relation to the patient's age to evaluate whether hearing impairment was progressive. RESULTS: Mixed hearing loss, i.e., including a substantial air-bone gap of up to 20 to 60 dB, was present in six cases, concomitantly with a submucous or overt cleft palate in five of them. The audiograms in 14 evaluable cases showed the following types of threshold: U-shaped (n = 3), flat (n = 2), flat or gently (downward) sloping (n = 3), gently sloping (n = 3), or steeply sloping (n = 3). Cross-sectional analysis did not reveal any significant effect of age on sensorineural hearing impairment. CONCLUSION: In contrast to the classic Stickler syndrome (type I) with high myopia, this nonocular type shows a high prevalence of sensorineural hearing impairment. The mean sensorineural hearing threshold in our patients was about 40 dB HL (95% CI, 15-65 dB) and was liable to increase (presumably by presbycusis) by several tens of decibels at the highest frequencies. Given the tendency for otitis media to develop in many of these patients, appropriate otologic care is of major importance. PMID- 10718439 TI - Modified particle repositioning procedure. AB - OBJECTIVES: To evaluate the efficacy of modifications to traditional particle repositioning maneuvers in the treatment of benign paroxysmal positional vertigo. STUDY DESIGN: Prospective trial of 118 patients with cupolocanalithiasis of the posterior canal treated with three different canal-repositioning techniques. METHODS: Results were compared with the maneuvers employed and the statistical importance of rotating patients by 360 degrees along their longitudinal axis and head shaking on reaching each single position were evaluated. RESULTS: Treatment of patients with our maneuver, which, in comparison with traditional repositioning maneuvers, was modified by breaking the procedure up into seven positions and rotating patients by 360 degrees along their longitudinal axis, gives a higher, but not statistically significant, number of treatment successes (84.5%) than the traditional Parnes maneuver (60%) (P = .154); treatment of a third group of patients with our modified particle repositioning maneuver with the addition of head-shaking on reaching each single position gives a higher (95.6%), statistically significant number of treatment successes than traditional Parnes maneuver (P = .00011). CONCLUSIONS: The success rates achieved from modified particle repositioning maneuvers are statistically significant. Onset or persistence of dizziness, which patients frequently complain of after liberatory maneuvers, affects only 5.6% of the patients treated. This low incidence is statistically correlated to head-shaking. PMID- 10718440 TI - Long-term results of uvulopalatopharyngoplasty for obstructive sleep apnea syndrome. AB - OBJECTIVES: Assessment of the long-term effect of uvulopalatopharyngoplasty (UPPP) on snoring, excessive daytime sleepiness, and nocturnal oxygen desaturation index (ODI) in patients with obstructive sleep apnea syndrome. STUDY DESIGN: Evaluation of snoring, excessive daytime sleepiness, and ODI in patients treated by UPPP earlier. MATERIALS AND METHODS: Patients (n = 58) with a follow up period of 11 to 74 months (median, 34 mo) were included in this study. Snoring and excessive daytime sleepiness were scored on specially designed semiquantitative scales. In all patients ODI was calculated from pulse-oximetry combined with polysomnography at base line and by polygraphy (MESAM 4) during follow-up in 38 patients. Long-term response was compared with 6-month response in the same cohort. RESULTS: There was a long-term improvement of snoring in 63% of patients, no change in 23%, and a deterioration in 14% (P < .00001). Overall snoring increased slightly between 6 months and long-term follow-up. There was an improvement of excessive daytime sleepiness in 38%, no change in 27%, and a deterioration in 35% (P = .80). Excessive daytime sleepiness showed a relapse to preoperative levels between 6 months and long-term follow-up. The median improvement of ODI was -1 (95% interpercentile range, 73-51) and was not significant (P = .35). In 5 of 13 patients in whom ODI at baseline exceeded 20, ODI was reduced to less than 20. In 4 of the 38 patients ODI was reduced to less than 5. The improvement of ODI decreased significantly between 6 months and long term follow-up (P = .03). No relation was found between body mass index, Mueller maneuver, X-cephalometry, and long-term outcome. An additional finding was that the ODI decreased after UPPP in combination with tonsillectomy, compared with a slight increase after UPPP alone; the difference was significant (P = .008). CONCLUSION: The response to UPPP for obstructive sleep apnea syndrome decreases progressively over the years after surgery. UPPP in combination with tonsillectomy was more effective than UPPP alone. PMID- 10718442 TI - Endoscopic postcricoid advancement flap for posterior glottic stenosis. PMID- 10718441 TI - A clinicopathologic series of 22 cases of tonsillar granulomas. AB - OBJECTIVES: Tonsils are uncommonly affected by granulomatous inflammation, often with an obscure cause. This study attempts to elucidate the nature of tonsillar granulomatous inflammation. DESIGN: Retrospective clinicopathologic review. METHODS: Twenty-two cases of tonsillar granulomas diagnosed between 1940 and 1999 were retrieved from the files of the Armed Forces Institute of Pathology. H&E slides and a series of histochemical stains were reviewed, and patient follow-up was obtained. RESULTS: There were 11 males and 11 females, aged 7 to 64 years (mean, 29.9 y). Most of the cases presented bilaterally (n = 19) with sore throat, dysphagia, and/or nasal obstruction. The clinical differential included chronic tonsillitis, tuberculosis, nonspecific infection, sarcoidosis, and a neoplasm. Histologically, the granulomas were focal and scattered, or diffuse, identified in the interfollicular zones (n = 16) and/or the germinal centers (n = 13), and occasionally associated with necrosis (n = 6). Based on histochemical and clinical follow-up information, the etiology of the granulomas included sarcoidosis (n = 8), tuberculosis (n = 3), Hodgkin's lymphoma (n = 2), toxoplasmosis (n = 1), squamous cell carcinoma (n = 1), and no specific known cause (n = 7). Twelve patients were either alive at last follow-up or had died with no evidence of disease (mean, 12.4 y), and 9 were either alive at last follow-up or had died with disease (mean, 24.9 y). One patient was alive with unknown disease status (lost to follow-up after 13.3 y). CONCLUSIONS: Although a cause for tonsillar granulomas is frequently identified, a number may not develop an identifiable etiology, with the granulomas probably representing an exaggerated immune response to chronic tonsillitis. However, a careful work-up must be conducted to exclude specific causes and avoid clinical mismanagement. PMID- 10718443 TI - Use of chalazion forceps to ease biopsy of minor salivary glands. PMID- 10718444 TI - Hiatus hernia: a review of evidence for its origin in esophageal longitudinal muscle dysfunction. AB - The axial forces exerted on the esophagus by the swallowing-induced contraction of its two longitudinally oriented muscle layers should, if unopposed, herniate the cardia through the diaphragm. A mathematical model of esophageal contraction shows that the magnitude of such a force becomes maximal just above the cardia, which is consistent with anatomic evidence of the existence of an inhibitory innervation in this same region. We propose that the inhibition exerted by these nerves when they discharge in swallowing prevents the supracardiac esophagus from shortening, allowing it to stretch and dissipating the pulling force of longitudinal muscle contraction above the diaphragm. Thus, hiatal hernia could originate from nerve disease, alteration in the viscoelastic properties of distal esophagus, or increased strength of the longitudinal muscle layers. PMID- 10718445 TI - Reflex-mediated enhancement of airway protective mechanisms. AB - The upper esophageal sphincter (UES) and lower esophageal sphincter (LES) comprise the basal mechanisms against entry of gastric content into the aerodigestive tract and the airway. There are, however, other mechanisms referred to here as "response mechanisms" that become activated after certain stimulation, such as distention of the esophagus or tactile/pressure stimulation of the pharyngeal wall, and result either in fortification of the UES barriers--i.e., esophago-UES, pharyngo-UES, and laryngo-UES contractile reflexes--or closure of the glottis--i.e., esophagoglottal and pharyngoglottal closure reflexes. In addition, there are other reflexes included among the response mechanisms--such as pharyngeal swallow and secondary peristalsis induced by pharyngeal stimulation by liquid and esophageal distention by refluxate--that result in pharyngeal and esophageal volume clearance, thus reducing the chance for contact of refluxate with the tracheal, bronchial, and glottal structures. PMID- 10718446 TI - Response of the lungs to aspiration. AB - Aspiration of acid from the stomach and water from the mouth can cause significant lung injury. Animal experiments suggest that acid entering the lungs is normally neutralized by bicarbonate derived from the plasma. It is hypothesized that this process may be impaired in patients with cystic fibrosis and that some of the airway injury that they experience may be related to this defect. This disease is characterized by abnormalities in the cystic fibrosis transmembrane conductance regulator, which normally conducts bicarbonate and chloride exchange. Evidence is discussed regarding the role of water channels (aquaporins) in transporting water from the airspaces into the vasculature. PMID- 10718447 TI - Esophageal body motor response to reflux events: secondary peristalsis. AB - The esophageal body is a major component of the antireflux mechanism. Disruption of esophageal peristalsis affects both volume clearance and delivery of swallowed saliva to the distal esophageal body. The esophageal body responds to reflux by an increase in primary peristalsis through stimulation of swallowing and secondary peristalsis through esophageal distension. Primary peristalsis is the most common motor event after reflux and accounts for up to 90% of initial and subsequent motor activity. Secondary peristalsis is uncommon but may be important during sleep when swallowing is relatively suppressed. Some patients with reflux disease, particularly those with severe esophagitis, exhibit impaired esophageal responses to reflux. It is likely that this impairment prolongs acid clearance and may also influence the proximal extent of the refluxate within the esophageal body. PMID- 10718448 TI - Functional anatomy and physiology of the upper esophageal sphincter. AB - Upper esophageal sphincter (UES) refers to the high-pressure zone located in between the pharynx and the cervical esophagus. The physiological role of this sphincter is to protect against reflux of food into the airways as well as prevent entry of air into the digestive tract. UES is a musculocartilaginous structure with its anterior wall being formed by the full extent of the posterior surface of the cricoid cartilage and arytenoid and interarytenoid muscles in the upper part. Posteriorly and laterally the cricopharyngeus (CP) muscle is a definitive component of the UES. CP has many unique characteristics: it is tonically active, has a high degree of elasticity, does not develop maximal tension at basal length, and is composed of a mixture of slow- and fast-twitch fibers, with the former predominating. These features enable the cricopharyngeus to maintain a resting tone and yet be able to stretch open by distracting forces, such as a swallowed bolus and hyoid and laryngeal excursion. CP, however, constitutes only the lower one third of the entire high-pressure zone. The thyropharyngeus (TP) muscle accounts for the remaining upper two thirds of the UES. The UES pressure is not entirely the result of myogenic activity, as a component of the pressure is the result of passive elasticity of the tissues. The opening of the UES involves relaxation of CP and TP muscles and forward movement of the larynx by the contraction of hyoid muscles. The UES function is controlled by a variety of reflexes that involve afferent inputs to the motorneurons innervating the sphincter. These physiological reflexes elicit either contraction or opening of the UES. Inability of the sphincter to open leads to difficulty in swallowing. Opening of the sphincter without associated CP relaxation leads to the clinical syndrome of cricopharyngeal bar. PMID- 10718449 TI - Understanding the motor innervation of the human cricopharyngeus muscle. AB - A comprehensive understanding of the swallow mechanism continues to suffer from an incomplete appreciation of basic morphology and function. This article is intended to underscore our currently incomplete and pending understanding of cricopharyngeus motor innervation in humans. Given the critical function of this sphincteric muscle in the control of the proximal alimentary tract, a focused effort to unravel its motor innervation would go a long way to reduce the mystery of its overall role in normal and disordered function in patients. PMID- 10718450 TI - Electromyographic recording of the cricopharyngeus muscle in humans. AB - Electromyography of the cricopharyngeus muscle is helpful in the study of normal swallowing and in the evaluation of various conditions leading to dysphagia. This article describes the technical aspects of the studies and the findings in normal controls and in various disease states. PMID- 10718451 TI - Management of upper esophageal sphincter disorders: indications and complications of myotomy. AB - Since 1951, when it was first used as a treatment for post-poliomyelitis dysphagia, cricopharyngeal myotomy (CPM) has been used in the treatment of various neurogenic, myogenic, structural, and idiopathic disorders. Yet, the efficacy of CPM in treating patients with upper esophageal sphincter (UES) disorders remains controversial. Despite favorable reports regarding its success, too few studies about indications, complications, and outcomes of CPM have been conducted to quell the controversy. Swallowing is accomplished when three primary conditions exist: (1) the cricopharyngeus muscle (CP) relaxes--that is, it is not tonically contracted, (2) the laryngo-hyoid complex elevates in an anterior superior direction to open the sphincter, and (3) pharyngeal pressure is sufficient to propel a bolus through the open sphincter. CPM is indicated when the second and third conditions are "adequate" but the first is inadequate, thus resulting in pharyngeal dysphagia associated with a defective opening of the UES. UES dysfunction is determined most often through patient history, physical examination, and testing. Patients with Zenker's (pharyngoesophageal) diverticulum, oculopharyngeal dystrophy, or inclusion body myositis are among those reported to have the most positive responses to CPM. Modified barium swallow is the most common measurement of UES dysfunction; manometry also is used, but to a lesser degree because of catheter motion during swallowing. There are four approaches to CPM, including: (1) the external technique, which is indicated when a muscle biopsy or neck exploration is needed; (2) the endoscopic approach, which is reported to work best with patients with Zenker's diverticulum and offers the choice of electrocautery, laser, or the surgical stapler--the last option being the best choice for high-risk patients; (3) balloon dilatation of the UES, a low-risk option that reportedly works best in patients with fibrosis of the CP; and (4) botulinum toxin injection of the CP transcervically or endoscopically, which offers low risk and minimal or no anesthesia. This approach best serves cases of failed relaxation of the CP. Each approach has potential complications, but reports of such are few and rarely severe. In all cases, massive reflux should be controlled before CPM and the patient should be medically stable. Patient selection for CPM remains inadequate. To assess the efficacy of CPM, more multi-institutional outcome studies need to be conducted. In the meanwhile, clinical judgment and selective testing via modified barium swallow are the best methods for identifying patients who may derive the most benefit from CPM. PMID- 10718452 TI - Sensory receptors of the larynx. AB - The larynx is a highly reflexogenic area, and stimulation with mechanical and chemical stimuli results in a number of protective reflexes. Investigators have used anatomical, behavioral, and neurophysiological techniques to examine the receptors responsible for initiating these reflex responses. Histologic examination has revealed the presence of free nerve endings, Merkel cells, Meissner corpuscles, and taste buds. Mechanoreceptors have been classified in several different ways and are located either in the superficial mucosa or in muscles and laryngeal joints. Recordings from afferent fibers innervating laryngeal mechanoreceptors have revealed that some of them are spontaneously active whereas others are silent until stimulated. Laryngeal mechanoreceptors respond to stimulation with either a rapidly adapting or a slowly adapting response pattern. Often the mechanoreceptors respond to respiratory movement of the larynx, giving bursts of action potentials during inspiration. A large number of taste buds that are anatomically similar to lingual taste buds populates the laryngeal surface of the epiglottis. Taste buds of the larynx respond to a number of chemical stimuli and to water. They do not respond to NaCl solutions close to physiological concentrations (0.154 M) but do respond at both a lower and higher concentration. When water is the solvent for the chemical stimuli, most chemicals initiate a response in laryngeal taste buds. However, when 0.154 M saline is used as a solvent, chemicals that taste bitter or sweet when applied to the tongue are ineffective stimuli. Taste buds of the larynx tend to be stimulated by the pH and tonicity of the stimulating solution and not by the gustatory properties. These results reveal a fundamental difference between the chemoreceptors of the oral cavity and larynx and result in the conclusion that chemoreceptors of the larynx do not play a role in gustation but are adapted to detect chemicals that are not saline-like in composition. PMID- 10718454 TI - Central integration of swallow and airway-protective reflexes. AB - The relationship between the timing of respiration and swallowing has been proven not to be random. Using pseudorabies virus (PRV) as a transsynaptic neural tracer, a basis for the central integration of swallowing and airway-protective reflexes can be located in the neural circuits projecting to swallowing-related muscles. The premotor neurons (PMNs) that constitute the swallowing central pattern generators, interneuronal networks able to initiate repetitive rhythmic muscle activity independent of sensory feedback, connect with multiple areas of the brainstem and other areas of the central nervous system. Those PMNs that project to muscles used in swallowing have been localized within the nucleus of the solitary tract (NTS) and its adjacent reticular formation, and they are synaptically linked both to peripheral afferents and to cortical swallowing areas. Bartha PRV, an attenuated vaccine strain of swine alpha-herpesvirus with a long postinjection survival rate and the ability to produce controlled infections that spread in a hierarchical manner within synaptically linked neurons, can specifically label neurons projecting to PMNs of a given circuit. Thus, it has been used to isolate two neuroanatomically distinct subnetworks of PMNs involved in the buccopharyngeal and esophageal phases of swallowing. Use of PRV as a neural tracer shows that during the buccopharyngeal phase of swallowing, vagal afferents from the pharynx and larynx and from the superior laryngeal nerve terminate in the intermediate and interstitial subnuclei of the NTS. Motoneurons projecting to the pharynx and larynx are located in the semicompact and loose formations of the nucleus ambiguus (NA). Neural tracing with PRV also shows that esophageal PMNs have direct synaptic contact with esophageal motoneurons in the compact formation of the NA. Moreover, esophageal PMNs are localized exclusively to the central subnucleus of the NTS, a site that also is the sole point of termination of esophageal vagal afferents. Using PRV, one can identify third order (neurons projecting to PMNs) esophageal neurons in sites where pharyngeal PMNs have been noted. Injection of PRV into the esophagus and subsequent detection using immunofluorescence found a subpopulation of neurons in the intermediate and interstitial subnuclei of the NTS. This subpopulation projects to pharyngeal motoneurons and buccopharyngeal PMNs, and it is synaptically linked to esophageal PMNs. The synaptic link between buccopharyngeal and esophageal PMNs provides a potential anatomic substrate within the NTS for the central integration of esophageal peristalsis with the pharyngeal phase of swallowing and airway-protective reflexes. Human studies and animal models investigating esophagoglottal closure and pharyngo-upper esophageal sphincter (pharyngo-UES) contractile reflexes have located the neural pathways that mediate airway protective reflexes. Similar studies and models using two PRV strains injected simultaneously into different swallowing and respiration-related muscle groups may identify synaptic connectivity between laryngeal, esophageal, and pharyngeal PMNs and, thus, may help to demonstrate the central integration of swallowing and airway-protective reflexes. PMID- 10718453 TI - Sensory innervation of the pharynx and larynx. AB - To shed light on supraesophageal complications of reflux disease, sensory innervation--particularly, distinct distribution, area, and density of sensory fibers--of the feline pharyngolaryngeal mucosa was reported. The investigations were performed by means of histochemistry (tracer techniques) and immunohistochemistry. The pharyngeal mucosa from the Eustachian cushion to the middle level of aryepiglottic fold, except the laryngeal surface of epiglottis, was supplied by the glossopharyngeal sensory fibers, whereas the laryngeal sensory fibers innervated between the apex of epiglottis and the level of the first tracheal ring in the larynx and between the middle level of aryepiglottic fold and the caudal end of piriform sinus in the pharynx. Most areas of the mucosa, except the subglottis, had unilateral innervation. The subglottis, including the caudal aspect of vocal fold and the posterior glottis, had bilateral supply with ipsilateral predominance. The density of sensory fibers in the vestibule of larynx involving the posterolateral aspect of arytenoid eminence was much heavier than the other parts. Sensory nerve fibers around the caudal pole of palatine tonsil, and in the root of tongue and the hypopharyngeal wall also were dense. Regional distribution and density of substance P and calcitonin gene-related peptide immunoreactive fibers showed almost the same pattern as did the sensory fibers. PMID- 10718455 TI - Clinical assessment of pharyngolaryngeal sensitivity. AB - The purpose of this article is to review the ongoing clinical research on assessment of laryngeal and pharyngeal sensitivity with particular emphasis on the technique of endoscopic air pulse stimulation of the laryngopharyngeal mucosa. Studies of laryngopharyngeal sensation in healthy controls and in stroke patients with dysphagia are presented initially. What then follows is a detailed description of a study comparing modified barium swallow and pharyngolaryngeal sensory testing as predictors of aspiration pneumonia after stroke. Finally, the combination of laryngopharyngeal sensory testing with endoscopic swallowing evaluations, termed flexible endoscopic evaluation of swallowing with sensory testing, and its implications in the office or bedside evaluation of the patient with dysphagia are discussed. PMID- 10718456 TI - The cough reflex and its relation to gastroesophageal reflux. AB - Each cough involves a complex reflex arc beginning with the stimulation of sensory nerves that function as cough receptors. There is evidence, primarily clinical, that the sensory limb of the reflex exists in and outside of the lower respiratory tract. Although myelinated, rapidly adapting pulmonary stretch receptors (RARs), also known as irritant receptors, are the most likely type of sensory nerve that stimulates the cough center in the brain, afferent C-fibers and slowly adapting pulmonary stretch receptors (SARs) also may modulate cough. RARS, C-fibers, and SARs have been identified in the distal esophageal mucosa; however, studies have not been performed to determine whether they can participate in the cough reflex. Although gastroesophageal reflux disease can potentially stimulate the afferent limb of the cough reflex by irritating the upper respiratory tract without aspiration and by irritating the lower respiratory tract by micro- or macroaspiration, there is evidence that strongly suggests that reflux commonly provokes cough by stimulating an esophageal bronchial reflex. Theoretically, the pathways of this reflex may be modeled in a variety of ways, and these are speculated upon in this article. The predominant role of acid in triggering cough by means of this reflex is unclear because of conflicting results from provocative challenge studies. It is interesting to speculate that a distal esophageal-bronchial reflex evolved as an early warning defense so that coughing could be started, just in case the refluxate were to reach the inlet of the lower respiratory tract. That is, thinking teleologically, it is possible that an esophageal-bronchial reflex evolved as one of several mechanisms designed to protect the lungs from aspiration of gastric contents. PMID- 10718457 TI - Brainstem viscerotopic organization of afferents and efferents involved in the control of swallowing. AB - Cholera toxin horseradish peroxidase (CT-HRP), a sensitive antegrade and retrograde tracer, is effective at labeling swallowing motoneurons and their dendritic fields within the nucleus ambiguus (NA), nucleus of the solitary tract (NTS), dorsal motor nucleus of the vagus nerve, and hypoglossal nucleus. Using this tracer to label motoneurons within the NTS demonstrates that palatal, pharyngeal, and laryngeal afferents overlap considerably within the interstitial and intermediate subnuclei. These afferents have a pattern of distribution within the NTS similar to the labeling observed after application of the same tracer to the superior laryngeal nerve. Esophageal afferents, however, terminate entirely within the central (NTScen) subnucleus and do not overlap their distribution with palatal, pharyngeal, or laryngeal afferents. Within the nodose ganglion (NG), sensory neurons projecting to the soft palate and pharynx are located superiorly, and those projecting to the esophagus and stomach are located inferiorly, an organization that indicates rostrocaudal positioning along the alimentary tract. Sensory neurons within the NG and NTS contain, among others, the major excitatory and inhibitory amino acid neurotransmitters glutamate (Glu) and gamma aminobutyric-acid (GABA). Both Glu and GABA help to coordinate esophageal peristalsis. Using pseudorabies virus as a transsynaptic tracer demonstrates the role of GABA and Glu as mediators of synaptic transmission within the swallowing central pattern generator, a fact further supported by the presence of specific receptors for each neurotransmitter within the NTScen. Anatomic studies using CT HRP have been effective in revealing the total extent of extranuclear dendritic projections and the organization of dendrites within the confines of a nucleus; further studies have produced the following data. Motoneurons innervating the soft palate, pharynx, larynx, and cervical esophagus have extensive dendrites that extend into the adjacent reticular formation with a distinct pattern for each muscle group. Motoneurons of the musculature active during the buccopharyngeal phase of swallowing (soft palate, pharynx, cricothyroid, and cervical esophagus) have extensive dendritic arborizations that terminate within the adjacent reticular formation of the NA. Swallowing premotor neurons located in the reticular formation surrounding the NA are active during the buccopharyngeal phase of swallowing. These data provide an anatomic basis for interaction of swallowing motoneurons with premotor neurons located in this area. Motoneurons innervating all levels of the esophagus are confined to the compact formation (NAc), whereas those motoneurons projecting to the pharynx and cricothyroid muscle are located in the semicompact formation (NAsc). The intrinsic laryngeal muscles were represented within the loose formation (NAI) and the heart within the external formation. In contrast, the dendrites of motoneurons projecting to the thoracic and subdiaphragmatic esophagus are confined to the NAc. Both the NAsc and NAc have extensive longitudinal bundling of dendrites within the confines of the nucleus, resulting in the formation of a rostrocaudal dendritic plexus where dendrites crisscross between bundles. Intranuclear bundling of dendrites is evident in the soft palate, pharynx, and esophagus and is lacking only for the cricothyroid muscle. Moreover, ventrolateral- and dorsomedial-oriented dendritic bundles are present within the NAsc. In contrast to the longitudinal dendritic bundles, the ventrolateral- and dorsomedial-oriented dendritic bundles exit the NAsc and penetrate the adjacent reticular formation. The extensive bundling of motoneuronal dendrites within the NA supports the hypothesis that these structures serve as networks for the generation of complex motor activities, such as swallowing. PMID- 10718458 TI - An overview of esophageal sensory receptors. AB - The neurophysiological basis of esophageal pain and discomfort is not well known. Functional disorder, such as noncardiac chest pain, is thought to be associated with hypersensitivity of primary afferents innervating the esophagus and/or sensitization of spinal dorsal horn cells receiving input from the organ. Although we have accumulated a large body of information about the morphologic structure and neuropeptide contents of the extrinsic primary afferents, we lack a full understanding of its integrative function in esophageal pain. The esophagus is innervated dually by vagus and spinal nerves. The majority of sensory afferents in the vagal and spinal pathway are pseudounipolar cells, with their cell bodies (soma) located in the nodose and dorsal root ganglia, respectively. These afferent fibers innervate serosa (adventitia), longitudinal and circular muscles, and mucosa of the esophagus. Afferents innervating the muscle are sensitive to intraluminal distension. In the vagus, these afferents exhibit low threshold for response, whereas the spinal afferents, including the splanchnic nerve afferents, have either low or high thresholds for response. In addition, these afferents are chemosensitive. Both vagal and spinal afferents also innervate the mucosa of the esophagus. These fibers are exquisitely sensitive to light touch of the mucosa and are sensitive to pH and chemicals. The spinal afferents, including splanchnic nerve afferents, project to the spinal cord, spanning from upper cervical (C1) to upper lumbar (L2) segments. A majority of the spinal dorsal horn neurons receiving input from the esophageal spinal afferents also receives somatic converging input. The somatic receptive fields are distributed mainly ipsilaterally over the chest and forearm area. These spinal dorsal horn neurons exhibit either excitatory, inhibitory, or biphasic (i.e., excitation followed by inhibition) responses to esophageal distension. PMID- 10718459 TI - Central control of lower esophageal sphincter relaxation. AB - The lower esophageal sphincter is innervated by both parasympathetic (vagus) and sympathetic (primarily splanchnic) nerves; however, the vagal pathways are the ones that are essential for reflex relaxation of the lower esophageal sphincter (LES), such as that which occurs during transient LES relaxations. Vagal afferent sensory endings from the distal esophagus and LES terminate in the hindbrain nucleus tractus solitarius. The preganglionic motor innervation of the LES arises from the dorsal motor nucleus of the vagus. Together these nuclei comprise the dorsal vagal complex within which there is a neural network coordinating reflex control of the sphincter. Vagal efferent preganglionic neurons to the gastrointestinal tract are organized viscerotopically in the dorsal motor nucleus of the vagus. Stimulation of the dorsal motor nucleus of the vagus caudal to the opening of the fourth ventricle results in relaxations, whereas stimulation in the rostral portion of the nucleus evokes contractions of the LES. Few details are known about the neural circuitry that links sensory information from the stomach and esophagus within the nucleus tractus solitarius to these separate populations of neurons within the dorsal motor nucleus of the vagus. The motor vagal preganglionic output is primarily cholinergic, which ultimately stimulates excitatory or inhibitory motor neurons that control the smooth muscle tone. Excitatory neurons evoke muscarinic receptor-mediated muscle contraction. Inhibitory neurons evoke nitric oxide or vasoactive intestinal polypeptide mediated relaxation of the lower esophageal sphincter. However, other neurotransmitters are found in vagal preganglionic neurons, including norepinephrine/dopamine and nitric oxide. A subpopulation of nitric oxide synthase-containing vagal preganglionic neurons innervate the upper gastrointestinal tract and mediate relaxation. The neurotransmitters and circuitry controlling lower esophageal sphincter pressure are important to characterize, because part of the dorsal vagal complex is outside of the blood brain barrier and is a potential target for pharmacologic intervention in the treatment of such disorders as gastroesophageal reflux disease. PMID- 10718460 TI - Epidemiology of esophageal and supraesophageal reflux injuries. AB - Gastroesophageal reflux disease (GERD) is a common gastrointestinal disorder. Despite its frequent occurrence, only a minority of patients seek medical attention, making it difficult to ascertain the true epidemiologic distribution of the disorder. A causal association between GERD and esophageal complications such as esophagitis, esophageal ulcer, and esophageal stricture is well accepted. Recent epidemiologic evidence suggests that GERD may likewise represent a risk factor for the development of supraesophageal conditions, such as asthma, chronic bronchitis, laryngitis, and even laryngeal cancer. Although epidemiologic associations do not establish a cause-and-effect relationship, they may indicate potential etiologic risk factors. Nevertheless, confirmation of the causal role of GERD in supraesophageal disorders awaits further investigation. PMID- 10718461 TI - Mechanisms of reflux-induced epithelial injuries in the esophagus. AB - The esophagus is lined by a moist stratified squamous epithelium. In humans, this epithelium, as evidenced by the Bernstein test, has considerable capacity to resist damage even upon direct contact of high concentrations of luminal acid. This intrinsic property of the epithelium to defend itself against luminal acid is called "tissue resistance." Tissue resistance is comprised of a number of important structures and functions, the individual functions of which are described in this review. Moreover, when luminal acidity is sufficiently noxious, tissue resistance can be overcome, leading to heartburn and ultimately cell necrosis. Herein are described the mechanisms by which acid overcomes the epithelial defense to produce these typical signs and symptoms of gastroesophageal reflux disease. PMID- 10718462 TI - Histopathology of reflux-induced esophageal and supraesophageal injuries. AB - As many as half of patients who have symptoms and objective evidence of gastroesophageal reflux disease (GERD) will have normal mucosa or only hyperemia at endoscopy. Because inflamed esophageal mucosa may appear normal endoscopically, and because hyperemia may or may not reflect histologic espophagitis, biopsy to document tissue injury in symptomatic patients with these minimal endoscopic findings may be helpful. Reflux may induce inflammation in the squamous mucosa of the esophagus, but in many patients only hyperplasia of the epithelium is seen. This hyperplasia is defined by a basal zone that exceeds 15% of the thickness of the mucosa and subepithelial papillae that exceed 67% of the thickness of the mucosa. Because these changes may be present normally in the distal 2.5 cm of the esophagus, and because they may be distributed over the distal 8 cm in a patchy fashion, multiple biopsies taken more than 2.5 cm above the esophagogastric junction are necessary to detect them reliably. Supraesophageal complications of GERD include posterior laryngitis, inflammatory polyp of the larynx (contact ulcer or laryngeal granuloma), subglottic stenosis and laryngeal squamous cell carcinoma. PMID- 10718463 TI - Diagnosis and management of chronic laryngitis associated with reflux. AB - Chronic laryngitis symptoms are commonly seen in otherwise healthy people. This article reviews recent progress in our understanding and effective treatment of chronic laryngitis. Clinical experience and prospective treatment and outcome studies have demonstrated objective evidence of the efficacy of treating patients with chronic laryngitis symptoms with nocturnal antireflux precautions and acid suppressing medications. The role of pH testing and most common errors in treatment are reviewed. PMID- 10718464 TI - Obstructive sleep apnea and gastroesophageal reflux. AB - A number of recent studies have described the presence of significant gastroesophageal reflux (GER) in patients with obstructive sleep apnea (OSA). The aims of our studies were to determine the prevalence of this in a controlled population and to investigate the potential for a causal relationship between the two entities by determining whether therapy of OSA altered GER parameters, and vice versa. All patients presenting to our sleep laboratory for screening polysomnography underwent distal esophageal pH monitoring simultaneously with polysomnography. Control subjects were selected if the apnea-hypopnea index (AHI) was <5.0, and patients were selected if AHI was >15.0. Fourteen subjects with OSA undertook a second polysomnographic study including distal esophageal pH monitoring, with nasal continuous positive airway pressure (nCPAP) intervention. Twelve subjects with proven OSA took part in a randomized, placebo-controlled, double-blinded, parallel group study of the effect of antireflux therapy (nizatidine) on OSA parameters. In 63 patients and 41 controls, we found that patients with OSA had significantly more GER events than controls as measured by number of reflux events over 8 hours (115 vs 23; P <0.001), and percent of time spent at pH <4.0 (21.4% vs 3.7%; P <0.001). In patients with proven OSA, 53.4% of GER episodes were temporally related to apneas or hypopneas. Less than half (46.8%) of all apneas were temporally related to acid reflux, and only 43.8% of arousals were related to reflux events. In the therapeutic trials, nCPAP reduced GER parameters in both patients with OSA and without OSA, suggesting a nonspecific effect. Antireflux therapy (nizatidine) reduced arousals but not apnea-hypopnea index in patients with OSA. Patients with OSA have a higher prevalence of GER than matched control subjects. Nasal CPAP reduces GER parameters nonspecifically, and thus the role of OSA in the pathogenesis of GER remains unclear. GER, however is likely to be important in the pathogenesis of arousals, but there is no evidence that it is involved in the pathogenesis of apneas. PMID- 10718465 TI - Anatomical diagnostic protocol in evaluating chronic cough with specific reference to gastroesophageal reflux disease. AB - Using the anatomic, diagnostic protocol, the cause of chronic cough can be determined 88% to 100% of the time, leading to specific therapy with success rates of 84% to 98%. Gastroesophageal reflux disease (GERD), along with postnasal drip syndrome (PNDS) and asthma, is one of the three most common causes of chronic cough in all age groups. When GERD is the cause of chronic cough, there may be no gastrointestinal (GI) symptoms up to 75% of the time, and, in these cases, the term "silent GERD" is used. The most sensitive and specific test for GERD is 24-hour esophageal pH monitoring. In interpreting this test, it is essential not only to evaluate the duration and frequency of the reflux episodes but also to determine the temporal relationship between reflux and cough events. Patients with normal standard reflux parameters still may have reflux diagnosed as the likely cause of cough if a temporal relationship exists. The definitive diagnosis of cough resulting from GERD can only be made if cough goes away with antireflux therapy. When 24-hour esophageal pH monitoring cannot be done, an empiric trial of antireflux medical therapy is appropriate when GERD is a likely cause of chronic cough. It is likely in the following settings: patients with prominent GI symptoms consistent with GERD and/or those with no GI complaints and normal chest x-rays, who are not taking angiotensin-converting enzyme inhibitors and who are not smoking, and in whom asthma and PNDS have been excluded. However, if empiric treatment fails, it cannot be assumed that GERD has been ruled out as a cause of chronic cough; rather, objective investigation for GERD is recommended, because the empiric therapy may not have been intensive enough or it may have failed. In treating patients with chronic cough resulting from GERD, cough has been reported to resolve with medical therapy 70% to 100% of the time. Mean time to recovery may take as long as 161 to 179 days, and patients may not start to get better for 2 to 3 months. In patients who fail to respond to maximal medical therapy, antireflux surgery can be successful. PMID- 10718466 TI - Esophageal biopsy for the diagnosis of gastroesophageal reflux-associated otolaryngologic problems in children. AB - Recently, gastroesophageal reflux (GER) has been found to contribute to many types of otolaryngologic pathology in infants and children. The complaints may be intermittent and unresponsive to usual therapies, such as antimicrobial treatments. A high index of suspicion for GER and for the concept of "silent" GER (GER without overt symptoms) is necessary for accurate diagnosis and treatment of otolaryngologic manifestations of GER in these patients. In this prospective historical cohort study, the records were reviewed from 101 children who underwent esophagoscopy and biopsy as a diagnostic test for GER at the time of other otolaryngologic procedures. Significant associations were found between the presence of histologic esophagitis and asthma, recurrent croup, cough, apnea, sinusitis, stridor, laryngomalacia, subglottic stenosis, posterior glottic erythema, and posterior glottic edema. There were no complications. Esophageal biopsy is a rapid, safe and effective diagnostic test for GER that should be considered at the time of other procedures in children with selected GER associated problems. PMID- 10718467 TI - Management of supraesophageal complications of gastroesophageal reflux disease in infants and children. AB - Therapy of supraesophageal manifestations of gastroesophageal reflux disease (GERD) in infants and children nearly always includes "lifestyle modifications" (conservative or nonpharmacologic therapy). Depending on the severity of the GERD manifestation, pharmacotherapy is often added. Although data to support the practice are not abundant, it is rational to begin with prokinetic pharmacotherapy and to add acid suppression if pathologic effects of acid contact with the esophagus or airway are suspected. Pathologic effects of acid produce most forms of supraesophageal GERD; the exception is infantile regurgitation, the most common example of supraesophageal GERD, which is often unaccompanied by either esophagitis or evidence of acid entry into the airway. Currently, fundoplication is rarely required for pediatric GERD, but the supraesophageal complications of GERD are more common indications for this surgery than the esophageal complications in children. Other management options for supraesophageal symptoms in children include delivery of nutrients by tube feeding slowly and continuously into the stomach or, better, small intestine. Short-term or trial tube feeding uses a transnasal tube, for example, for nasojejunal feeding; longer-term tube feeding is simplified by a gastrostomy, which can be placed relatively noninvasively using endoscopy or fluoroscopy. PMID- 10718468 TI - Sudden infant death syndrome: can gastroesophageal reflux cause sudden infant death? AB - Although gastric contents in the airways and lungs of sudden infant death syndrome (SIDS) victims is commonly found during postmortem examination, its significance as a sole or contributory cause of death has long been controversial. Currently, most authorities view such aspiration as resulting from "agonal" processes and, therefore, irrelevant to cause of death. Recent clinical and experimental evidence indicates that infants who are near death because of a variety of conditions, frequently "autoresuscitate," which produces rapid and complete recovery. In animal models, aspiration of water or saline into the airways has been shown to prevent autoresuscitation. In light of these findings, aspiration of gastric contents should be reconsidered as a contributory cause of many SIDS deaths. PMID- 10718469 TI - Gastroesophageal reflux and laryngitis: a skeptic's view. AB - Gastroesophageal reflux disease (GERD) and laryngitis are common problems in the community. To prove a causal relationship between GERD and laryngitis, one must show the mechanism for acid-induced laryngitis, establish the frequency of association, have a gold standard for diagnosis of the condition, and show that treatment that reduces acid reduces laryngitis in double-blind, randomized, placebo-controlled trials. This article examines each of these components and finds that all are wanting as proof for reflux-induced laryngitis. The best approach to prove the association between GERD and laryngitis will likely lie in well-designed treatment trials. PMID- 10718470 TI - Gastroesophageal reflux disease and asthma: the two are directly related. AB - GERD and asthma have met the three criteria set out to prove a relationship between the two diseases. Patients with GERD have a higher prevalence of asthma, and there are several pathophysiologic mechanisms by which acid reflux can cause bronchospasm. Furthermore, aggressive antireflux therapy in patients with asthma and GERD results in improvements in asthma outcome in as many as 70% to 80% of patients treated in both medical and surgical series. Nevertheless, there are design flaws in many of the outcome studies performed to date. To further clarify this issue, future studies should be multicentered and placebo controlled using acid-suppressive therapy for at least 3 to 6 months with documentation of asthma outcome, cost analysis, and quality-of-life assessment. As with many things in medicine, all the data are not consistent. However, I strongly believe that the available data support the aggressive search for GERD and treatment in any patient with difficult-to-control asthma. PMID- 10718471 TI - Why do the published data fail to clarify the relationship between gastroesophageal reflux and asthma? AB - The relationship between gastroesophageal reflux (GER) and asthma has troubled physicians for centuries and has been a source of debate among pulmonologists, allergists, and gastroenterologists for decades. Attempting to tie together the pieces of the puzzle, numerous investigators have struggled to show that in patients with asthma, GER symptoms occur too frequently, gastric acid dwells for too long in the esophageal lumen, and refluxed gastric acid injures the esophageal mucosa more than expected. Unfortunately, all of the work done by these fine investigators has failed to demonstrate a "cause and effect" relationship. Although they have succeeded in convincing us that GER occurs more frequently in asthmatics than in nonasthmatics, they still must continue until we all know how to predict which patients have gastroesophageal-induced or gastroesophageal-exacerbated asthma. PMID- 10718472 TI - Medical therapy of supraesophageal gastroesophageal reflux disease. AB - Medical therapy of supraesophageal gastroesophageal reflux disease (GERD) is based on the principals for treating patients with heartburn and erosive esophagitis, observations from the few available clinical trials, and clinical experience. In general, patients with supraesophageal GERD require higher doses of antireflux therapy, principally with proton pump inhibitors, for longer periods of time to effectively relieve symptoms compared with patients with heartburn and/or erosive esophagitis. This article reviews the current literature and discusses a suggested approach to medical management of these often complex patients. PMID- 10718473 TI - Surgical therapy for supraesophageal reflux complications of gastroesophageal reflux disease. AB - Supraesophageal complications of gastroesophageal reflux can be successfully treated by antireflux surgery. Careful preoperative testing, including 24-hour esophageal pH, manometry, and endoscopy, will help to identify appropriate patients who will benefit from surgery. The best results are achieved in patients with nocturnal asthma, the onset of reflux before pulmonary symptoms, laryngeal inflammation, and a good response to medical therapy. Cough is more responsive to surgical therapy than is asthma. The benefits of minimally-invasive surgery are evident in patients with pulmonary disease, who have a faster recovery with fewer complications than after open surgery. PMID- 10718474 TI - Life after antireflux surgery. AB - Laparoscopic fundoplication technique has become the operative modality of choice for antireflux surgery. An increasing number of patients and physicians have enthusiastically embraced this "minimally-invasive" technologic development for treatment of gastroesophageal reflux disease (GERD). However, laparoscopic fundoplication has been frequently advertised as the therapeutic solution for all GERD patients. Subsequently, the number and severity of complications resulting from laparoscopic surgery--often performed indiscriminately--has increased dramatically. This article reviews the efficacy of the laparoscopic fundoplication operation for GERD based on current, relatively short-term reports from centers specializing in this treatment modality. The majority of these reports are very positive. Unfortunately, the results of fundoplication operations performed by community surgeons are unknown. There are a number of disturbing factors relating to laparoscopic treatment for GERD that should raise a red flag of caution to the medical community, particularly primary-care physicians and their patients. The central portion of this report devotes itself to discussing the problems associated with this new minimally-invasive technique for fundoplication operations. These problems include the selectivity of current reports on outcomes of the laparascopic fundoplication operation and the lack of uniform data acquisition associated with these postoperative studies. The technical difficulties of the laparascopic fundoplication surgery are discussed, and the need for operator expertise and appreciation of esophageal physiology and anatomy are stressed. Finally, the long-term durability of the fundoplication wrap is questioned and the morbidity associated with the operation--particularly dysphagia--is addressed. In the final segment, the complications encountered after laparoscopic fundoplication operations are detailed and the techniques for evaluating the symptomatic postfundoplication patient are discussed. Laparoscopic fundoplication operation is good therapy in an appropriate clinical setting when performed by a well-trained and experienced surgeon. However, the operation should not be first-line therapy for the majority of GERD patients. An esophagus disabled by an inappropriate or dysfunctional fundoplication wrap is a terrible price to pay for control of acid reflux. PMID- 10718475 TI - In vitro studies of the interaction of poly(NIPAm/MAA) nanoparticles with proteins and cells. AB - The pH- and temperature-responsive poly(N-isopropylacrylamide-co-methacrylic acid) (PNIPAm/MAA) nanoparticles are of potential application in targeted drug delivery. Their responsive properties in the presence of human serum albumin were investigated using dynamic light scattering (DLS), protein assay, and electron spin resonance (ESR) spectroscopy. Their interaction with human monocytes and polymorphonuclear leukocytes (PMNLs) was studied using scanning electron microscopy (SEM) and oxygen consumption method. The nanoparticles exhibited a volume phase transition at 35-40 degrees C in Hanks balanced salt solution (HBSS) and in phosphate buffer solution (PBS) of pH 7.4. The diameter of the nanoparticles decreased slightly in the presence of HSA at 25 degrees C at neutral pH, whereas an increase in the diameter in pH 6 PBS at 40 degrees C was revealed. The amount of albumin adsorbed onto the nanoparticles decreased with increasing temperature. The ESR spectra of spin labeled HSA indicated a more restricted environment in the nanoparticles at elevated temperatures. The stimulation of PMNL oxygen consumption by PNIPAm based nanoparticles, an indication of phagocytosis of the particles, was not observed regardless whether the nanoparticles were incubated in plasma or serum. In contrast, the more hydrophobic polystyrene (PSt) particles induced a significant increase in the rate of oxygen consumption after the incubation. PNIPAm/MAA-grafted-PSt particles behaved similarly to the PNIPAm/MAA nanoparticles, suggesting that surface properties dictate the recognition of colloids by PMNLs. PMID- 10718476 TI - Growth of L929 cells on polymeric films prepared by Langmuir-Blodgett and casting methods. AB - The growth and spreading of fibroblast, L929 cells, on various polymeric films prepared by the Langmuir-Blodgett (LB) and casting methods were investigated. L929 cells, which were cultivated on collagen and synthetic polymeric films prepared by the LB method, adhered and spread much more than those on synthetic films prepared by the casting method. This is explained by the fact that cell growth and cell spreading are suitable for L929 cells on the films having serum proteins that contain a high alpha-helix content, because LB films adsorbed those serum proteins estimated from the circular dichroism measurements of the films immersed in cell culture medium. An exponential relationship was observed from the plot of the cell density vs root mean square of roughness of the films, which is estimated by atomic force microscopy, whereas a linear relationship was observed from the plot of the spreading ratio vs the root mean square of roughness. It is suggested that the correlation between the cell growth or spreading ratio and surface roughness of the films where L929 cells were cultivated is considered to be more important than the correlation between the cell growth or spreading ratio and the contact angle of the films. PMID- 10718477 TI - Surface modification of poly(tetrafluoroethylene) films via grafting of poly(ethylene glycol) for reduction in protein adsorption. AB - Poly(tetrafluoroethylene) (PTFE) films with surface grafted poly(ethylene glycol) (PEG) chains were prepared by two methods: (1) UV-induced graft copolymerization of methoxy poly- (ethylene glycol) monomethacrylate (PEGMA) onto the plasma pretreated PTFE films; and (2) coupling of the hydroxyl groups of PEG via ester linkages with the carbonyl chloride groups which were introduced onto the acrylic acid (AAc) graft-copolymerized PTFE surface through reaction with thionyl chloride (SOCl2). The UV-induced graft copolymerization of PEGMA onto the plasma pretreated PTFE film was explored with different macromonomer concentrations and different UV graft copolymerization time. The coupling reaction, on the other hand, was explored with PEG of different molecular weights. The surface microstructures and compositions of the PEG-modified PTFE films from both processes were characterized by contact angle, X-ray photoelectron spectroscopy (XPS), and atomic force microscopy (AFM) measurements. In general, higher macromonomer concentration and longer UV graft copolymerization time led to a higher graft yield for the UV-induced graft copolymerization with PEGMA. Contact angle measurements revealed that the hydrophilicity of the PTFE film surface was greatly enhanced by the grafting of the PEG chains. The PTFE surface with a high density of grafted PEG was very effective in preventing bovine serum albumin adsorption. PMID- 10718479 TI - The in vitro blood compatibility of poly(ethylene oxide)-grafted polyurethane/polystyrene interpenetrating polymer networks. AB - Poly(ethylene oxide) (PEO)-grafted polyurethane (PU)/polystyrene (PS) interpenetrating polymer networks (IPNs) were synthesized. The effects of the mobile pendant PEO chains with their microphase separated structure on blood compatibility were investigated. The morphology of both the fracture surface as well as the top surface indicate that the size of the dispersed domains of the PS rich phase decreased as the grafting with the PEO was increased. The swelling ratio also decreased as the grafting with the PEO was increased. However, the dynamic contact angle and the interfacial energy between IPN surface and water decreased, due to the structural reorganization of the pendant PEO chains. PU/PS IPNs have an excellent mechanical property as compared with PU homopolymers. The adsorption of bovine plasma fibrinogen (BPF) onto the PU/PS IPNs and PU homopolymers was effectively suppressed by the PEO-grafting. In the platelet adhesion test, the amount of platelets adsorbed, activated, and/or coagulated upon the PEO-grafted PU/PS IPNs were reduced when compared to the ungrafted PU homopolymers. PMID- 10718478 TI - In vitro toxicity of spiroorthocarbonate monomers designed for non-shrinking dental restoratives. AB - In development of photopolymerized expanding monomers with epoxy resin systems, there is a need for reactive expanding monomers that exert a good biocompatibility profile. The objective of this study was to evaluate the in vitro toxicology of new spiroorthocarbonates designed to be expanding monomers. The expanding monomers investigated were: trans/trans-2,3,8,9-di(tetramethylene) 1,5,7,11-tetraoxaspiro[5,5] undecane (DTM-TOSU), 5,5-diethyl-19-oxadispiro-[1,3 dioxane-2,2'-1,3-dioxane-5',4'-bicy clo[4.1.0]heptane] (DECHE-TOSU); 3,9-diethyl 3,9-dipropionyloxy methyl-1,5,7,11-tetraoxaspiro[5.5]undecane (DEDPM-TOSU); and 3,9-diethyl-3,9-diacetoxy methyl-1,5,7,11-tetraoxaspiro[5.5]undecane (DAMDE TOSU). The in vitro toxicology of these monomers measured their cytotoxicity and mutagenicity potential. Succinic dehydrogenase (SDH) activity in the MTT assay was used to assess the toxic dose that kills 50% of cells (TC50) for all the monomers. Their mutagenic potential was measured in the Ames Salmonella assay with and without metabolic activation. Two solvents, DMSO and acetone, were used to validate effects. Appropriate controls included the solvents alone. All the expanding monomers in this study were less cytotoxic than BISGMA (p < 0.01), a commercial component of dental restoratives. The relative cytotoxicity of the expanding monomers in DMSO was defined in the following order: DTM-TOSU (more toxic) > DECHE-TOSU > DEDPM-TOSU > DAMDE-TOSU. Each was significantly different from the other (p < 0.05). Overall, the TC50 values of all expanding monomers were significantly greater in DMSO than in acetone (p < 0.05). However, for BISGMA this trend was opposite. For mutagenicity results, the expanding monomers were non-mutagenic and there was no solvent effect on this outcome. The non mutagenicity and low cytotoxicity profile of these expanding monomers suggests their potential for development of biocompatible non-shrinking composites. PMID- 10718480 TI - In situ self hardening bioactive composite for bone and dental surgery. AB - A new biomaterial is presented which consists of a cellulose derivative- silanised hydroxyethylcellulose (HEC-SIL) and biphasic calcium phosphate (BCP). Rheological properties of the polymer itself and its mixture with BCP are pH dependent. At pH 10-12 HEC-SIL is liquid and undergoes quick gellation at pH < 9. Similarly, the paste of HEC-SIL and BCP is fluid and injectable at higher pH and solidifies in biological solutions. The rate of this solidification can be easily controlled by the degree of substitution of hydroxyethylcellulose with silicoalkoxy groups. PMID- 10718481 TI - SUM-159PT cells: a novel estrogen independent human breast cancer model system. AB - Breast cancer remains one of the most common malignant diseases in women in North America and Western Europe, yet therapies for the more aggressive estrogen independent tumors are limited and few model systems are available for the study of this type of breast cancer. In these studies, we characterized a novel estrogen independent breast cancer cell line, SUM-159PT. SUM-159PT cells are epithelial in origin, demonstrated by expression of cytokeratin 18. SUM-159PT cells are estrogen independent, demonstrated by lack of estrogen receptor (ER) protein and ER ligand binding studies. Furthermore, SUM-159PT cells injected subcutaneously or orthotopically are tumorigenic in ovariectomized athymic nude mice in the absence of estradiol supplementation. SUM-159PT cells are capable of invading through an 8 microm Matrigel membrane and display a stellate morphology in Matrigel, indicative of a metastatic phenotype. Correlating with this phenotype, we have detected secondary tumors upon inoculation of SUM-159PT cells into the mammary fat pad. To further investigate the metastatic potential of the SUM-159PT cells, we examined the expression of two proteins, vimentin and E cadherin, implicated in the transition of carcinoma cells to a metastatic phenotype. Western blot and immunohistochemical analysis demonstrated that both SUM-159PT cells and xenografts express vimentin. No expression of E-cadherin was detected in SUM-159PT cells. Our data indicate that despite estrogen independence, SUM-159PT cells are growth inhibited in vitro by compounds such as 1,25(OH)2D3, transforming growth factor beta (TGF-beta), and the phorbol ester TPA. These studies indicate that SUM-159PT cells represent a good model system for the study of late stage estrogen independent, invasive breast cancer. PMID- 10718482 TI - The effect of estrogen usage on the subsequent hormone receptor status of primary breast cancer. AB - In order to determine if prior use of exogenous estrogens was related to the estrogen receptor (ER) content of primary breast cancers, a retrospective analysis was performed from 536 patients with invasive breast cancer. The patient's age, menopausal status, oral contraceptive or estrogen replacement therapy usage, and the ER and progesterone receptor (PR) content of the breast cancer were recorded for all patients. Hormone usage in premenopausal and postmenopausal patients was compared to ER and PR levels in primary breast cancers using non-parametric testing. Complete information was available from 508 (193 premenopausal and 315 postmenopausal) patients. Breast cancers were ER positive in 72% of postmenopausal patients and 57% of premenopausal patients. The majority of patients received 'Some' form of hormone therapy (111 of 193 premenopausal patients and 233 of 315 postmenopausal patients). Significantly more estrogen receptors were detected in tumors from patients receiving 'some' estrogen therapy compared to 'never' users. Postmenopausal patients 'never receiving estrogen therapy had a lower rate of ER positive tumors (62%) compared to 'some' users (75%, chi2 = 4.99, p < 0.05). The same relationship was seen for PR ('never' users 44% positive, 'some' users 58% positive, chi2 = 5.19, p < 0.05). We conclude that postmenopausal patients who received 'some' estrogen therapy are more likely to have breast cancers that are estrogen receptor and progesterone receptor positive. PMID- 10718483 TI - Quantitation of erbB-2 gene copy number in breast cancer by an improved polymerase chain reaction (PCR) technique, competitively differential PCR. AB - A new method of measuring gene copy number in small samples of DNA was used to measure amplification of the erbB-2 gene and a reference gene in breast cancers. The method, termed 'competitively differential polymerase chain reaction' (CD PCR), combines the advantages of two other techniques for measuring amplification by PCR, namely differential PCR (D-PCR) and competitive PCR (C-PCR). The CD-PCR methodology was evaluated for sensitivity and specificity by comparing amplification measured by CD-PCR with that obtained by fluorescence in situ hybridization (FISH), C-PCR, and Southern blotting analysis. CD-PCR analysis proved to be an accurate predictor of amplification. CD-PCR also overcomes the problems involved in variation of PCR efficiencies and DNA concentrations in tumor samples, and the problems caused by the plateau effect in PCR. PMID- 10718484 TI - Influence of estrogen receptor variants on the determination of ER status in human breast cancer. AB - Determination of estrogen receptor alpha (ER) status in breast cancer is an important predictive factor for clinical response to endocrine therapy. We have recently shown that discrepancies in ER status determined by immunohistochemical assay (ER-IHA) can occur between amino-terminal (1D5) and carboxyl-terminal (AER 311) targeted ER antibodies and that those tumors which demonstrate discordance are associated with increased expression of truncated ER variant mRNAs. In this study, we have explored this observation to examine if ER variant expression can exert a direct effect on ER-IHA or whether this association is attributable to the characteristics of the antibodies. ER negative cos-1 cells were transfected with expression vectors containing wild type ER (wt-ER) and/or a frequently expressed truncated variant, ER-clone-4 variant. We found that ER-IHA performed with the same N- and C-terminal targeting ER antibodies on cos-1 cells expressing wt-ER alone demonstrated no difference in signals by western blot (P > 0.1). However, co-expression of wt-ER and the truncated ER-clone-4 variant, resulted in discordant IHA results with relatively higher ER-IHA H-scores from N-terminal antibodies (P < 0.03). Furthermore, re-examination of a subset of breast tumors previously studied by ER-IHA showed persistent concordance in 4/5 cases and persistent differences in 3/5 cases with a different pair of ER antibodies. We conclude that the presence of truncated ER variant proteins can interfere with the interpretation of ER status determined by IHA and that this may account for some of the inconsistencies between ER status and response to endocrine therapy. PMID- 10718485 TI - Sex hormone-induced mammary carcinogenesis in female Noble rats: expression of TGF-beta1 and its receptors, TGF-alpha, and EGF-R in mammary carcinogenesis. AB - We have established a Noble rat model to explore the mechanisms of hormonal mammary carcinogenesis, in which the role of androgen in promoting mammary carcinogenesis was highlighted. We have also established that stromal-epithelial interactions may be responsible for the promotional effects of testosterone in mammary carcinogenesis. Based on these understandings, in the present study we examined the expression of transforming growth factor beta-1 (TGF-beta1) and its receptors (TGF-beta RI, TGF-beta RII), transforming growth factor alpha (TGF alpha), and epidermal growth factor receptor (EGF-R) in 'pre-malignant' mammary glands treated with different protocols of sex hormones, as well as in mammary cancers. We observed that TGF-beta1 was strongly expressed in most mammary tumors, whereas TGF-beta RI and TGF-beta RII were negative in most mammary tumor cells. The results from comparative study of 'pre-malignant' glands further showed that when the animals were treated with testosterone, either alone or in combination with 17beta-estradiol, the mammary gland epithelial cells expressed high levels of TGF-beta1. This over-expression of TGF-beta1 can be blocked by flutamide, indicating that testosterone may be responsible for the expression of TGF-beta1 in mammary glands. TGF-beta RI and TGF-beta RII were also expressed strongly in testosterone-treated mammary epithelial cells and only weakly detectable in 17beta-estradiol treated and control mammary epithelial cells. Furthermore, TGF-beta RI and TGF-beta RII were also expressed in stromal cells, both in mammary tumors and in hormone-treated mammary glands. These observations indicate that the mechanism of testosterone in mammary carcinogenesis may be through its regulation of expression of TGF-beta1 and its receptors. On the other hand, TGF-alpha was also expressed in all 39 mammary cancers, while only 81% of the cancers were EGF-R positive. TGF-alpha was also strongly expressed in stromal cells in all three experimental groups, but only moderately expressed in epithelial cells when treated with a combination of testosterone and 17beta estradiol. By contrast, EGF-R was strongly expressed in epithelial cells in the three experimental groups but negative in stromal cells. Flutamide or tamoxifen was unable to block the expression of TGF-alpha induced by the combined sex hormone treatment. However, they were effective in blocking the expression of TGF alpha when the animals were treated with testosterone or 17beta-estradiol alone, respectively. These results suggest that both testosterone and 17beta-estradiol may be required for the over-expression of TGF-alpha in the mammary carcinogenesis induced by sex hormones. To our knowledge, this is the first experimental study to explore the regulation of TGF-beta1, TGF-alpha, and their receptors by testosterone and 17beta-estradiol in mammary carcinogenesis. PMID- 10718486 TI - Changes associated with delay of mammary cancer by retinoid analogues in transgenic mice bearing c-neu oncogene. AB - Breast cancer is one of the common cancers and is a leading cause of cancer mortality in women. The TG.NK transgenic mouse line on FVB strain background expresses the c-neu oncogene under the control of a MMTV promoter in mammary tissue and appears to be a useful animal model for evaluation of strategies to delay or prevent mammary cancer. Fiber-rich nonpurified diet (NTP-2000) and some retinoid analogues have delayed mammary cancer in the TG.NK model. Four week old hemizygous TG.NK female mice with MMTV/c-neu (erbB2) activated oncogene were fed NTP-2000 diet containing the retinoid analogue 4-hydroxyphenylretinamide (4-HPR) at 7 mmol/kg or the arotinoid Ro 40-8757 at 1.5 and 2.5 mmol/kg for 26 weeks. The 4-HPR at 7 mmol/kg diet delayed the development of palpable tumors up to 24 weeks, but by 26 weeks, the incidence markedly increased and was closer to the NTP-2000 diet control group. However, the 4-HPR diet markedly decreased the average weight of the tumors at 26 weeks with no decrease in multiplicity. The 4 HPR also caused significant increase in liver weights without an effect on body weight. Arotinoid Ro 40-8757 caused marked decrease in the number and branching of mammary ducts, and inhibited mammary tumor development with significant decrease in the incidence, multiplicity, and tumor weights compared to the NTP 2000 diet control. Arotinoid also caused a significant dose-related increase in liver weights without a significant effect on body weights. At the doses tested, the arotinoid but not 4-HPR decreased the circulating levels of IGF-1. However, there was no association between the IGF-1 levels and the size, incidence, or absence of tumors when evaluated for any treatment group or for all mice in the study irrespective of treatment. The oncogene erbB2 (c-neu) and the growth factor EGF expression were more prominent in the small tumors of the mice treated with arotinoid than in the larger tumors of the control group. PCNA staining was observed in areas where there was high erbB2 and EGF staining. The delay in onset of mammary tumors by the above retinoid analogues may be related to the delay in development of mammary glands. PMID- 10718487 TI - Heterogeneity of MUC1 expression by human breast carcinoma cell lines in vivo and in vitro. AB - Increased expression of the epithelial mucin MUC1 has been linked to tumor aggressiveness in human breast carcinoma. Recent studies have demonstrated that overexpression of MUC1 interferes with cell-substrate and cell-cell adhesion by masking cell surface integrins and E-cadherin. Additionally, the cytoplasmic tail of MUC1 is involved in signal transduction and interactions with catenins. In the present study, we have examined the in vitro expression of MUC1 mRNA and protein in a panel of 14 human breast cancer cell lines using northern blotting, western blotting, immunocytochemistry, and flow cytometry. Considerable variability of expression was noted not only between cell lines but also within several individual lines. Many cell lines such as BT 20, KPL-1, and T47D expressed abundant MUC1 whilst others such as MDA-MB-231 and MCF-7 showed intermediate expression, and MDA-MB-435 and MDA-MB-453 expressed very low levels. Low levels of MUC1 expression were associated with decreased expression of cytokeratin and increased expression of vimentin. Additionally, 12 of the cell lines were established as xenografts in immunocompromised (SCID) mice, and MUC1 expression in both the primary tumors as well as metastases was assessed immunohistochemically. In general, in vivo expression mirrored in vitro expression, although there was reduced in vivo expression in T47D and ZR-75-1 xenografts. Although we showed no correlation between tumorigenicity or metastasis and MUC1 expression, this study will assist development of experimental models to assess the influence of MUC1 of on breast cancer progression. PMID- 10718488 TI - Expression of the estrogen receptor-associated immunophilins, cyclophilin 40 and FKBP52, in breast cancer. AB - The estrogen receptor alpha (ER alpha) is implicated in the development of breast cancer. The immunophilins, cyclophilin 40 (CyP40) and FKBP52, are associated with ER alpha and other steroid receptors in mutually exclusive heterocomplexes and may differentially modulate receptor activity. Since previous studies have not assessed the levels of these immunophilins in breast cancer, we examined 10 breast cancer cell lines for mRNA and protein expression of CyP40 and FKBP52 and for amplification of the CyP40 gene. In addition, 26 breast carcinomas, including seven with matched normal breast tissue, were examined for mRNA expression of both immunophilins. CyP40 and FKBP52 were ubiquitously expressed in breast cancer cell lines, but there were significant differences in their pattern of expression. FKBP52 protein levels were generally an order of magnitude greater than those for CyP40. FKBP52 mRNA expression correlated strongly with protein expression and was significantly higher in ER alpha-positive compared with ER alpha-negative cell lines. However, CyP40 mRNA expression did not correlate with protein expression, nor did expression of this immunophilin correlate with ER alpha status. Relatively high expression of CyP40 in one cell line (BT-20) could be attributed to amplification of the CyP40 gene. Both immunophilins were also ubiquitously expressed in breast carcinomas, and we demonstrate for the first time that both CyP40 and FKBP52 mRNA are overexpressed in breast tumors compared to matched normal breast controls. The overexpression of CyP40 and FKBP52, coupled with relative differences in their expression in tumors, may have important functional implications for ER alpha and other steroid receptors in breast cancer. PMID- 10718489 TI - Symptoms associated with oophorectomy and tamoxifen treatment for breast cancer in premenopausal Vietnamese women. AB - There are very few data about the efficacy and toxicity of adjuvant systemic therapies for breast cancer in non-western populations. In 1993 in Vietnam we began a randomized controlled clinical trial on premenopausal women with operable breast cancer comparing adjuvant surgical oophorectomy plus tamoxifen with observation and this same combined hormonal treatment on recurrence. We evaluated the symptoms reported at regular follow-up visits by the first 482 premenopausal women entered in this clinical trial and treated with surgical oophorectomy plus tamoxifen or observation. Hot flash frequency and intensity, vaginal discharge, and genital pruritus were the only symptoms to occur more frequently in oophorectomy and tamoxifen-treated subjects. Seventy-seven percent of oophorectomy/tamoxifen subjects reported grade 1 or more and 44% grade 2 or more hot flash frequency symptoms in the first 12 months, versus 9% and 1% of observation subjects, respectively. Twenty percent of oophorectomy/tamoxifen subjects had grade 2 or greater intensity of hot flashes some time in the first 12 months versus 0% in observation subjects. Through three years, vasomotor symptoms were reported more frequently in oophorectomy/tamoxifen-treated women (in 23% vs. 3% at three years, mostly grade 1 toxicities). While noted and persistent vasomotor symptoms were found with oophorectomy plus tamoxifen in this population of Vietnamese women, these were of lower grades and tolerable. This adjuvant treatment may be widely accepted if it is demonstrated to be effective in this population. PMID- 10718490 TI - MMP-2 positivity and age less than 40 years increases the risk for recurrence in premenopausal patients with node-positive breast carcinoma. AB - Node-positive breast carcinoma is associated with a poor prognosis. Some patients benefit from adjuvant chemotherapy but new treatment modalities should still be developed in order to further increase the cure rate in this patient group. Prognostic factors are needed to define patients for such studies. Here, the prognostic value of matrix metalloproteinase-2 (MMP-2) and age was evaluated in 108 premenopausal, node-positive breast carcinoma patients treated with an adjuvant chemotherapy. Expression of MMP-2 protein was studied in paraffin embedded tissue sections from primary tumors by using specific MMP-2 monoclonal antibody in an immunohistochemical staining. Age less than 40 years predicted 5 year recurrence free survival (RFS) as unfavorable, being 74% in patients 41-49 years of age and 54% in those under age 40 (p = 0.02). The 5-year RFS rate was 85% in patients with an MMP-2 negative primary tumor while it was 65% in the MMP 2 positive patient group. This difference was not, however, statistically significant (p = 0.07). Correlation between hematogenous metastasis and MMP-2 positivity in breast carcinoma was demonstrated for the first time (p = 0.03). A risk group for a relapse was identified using MMP-2 immunohistochemistry and age. The RFS rate in patients less than 40 years with an MMP-2 positive primary tumor was only 50% while it was 74% in other premenopausal patients (p = 0.007). Young age and MMP-2 positivity may, thus, associate with early relapse in node-positive breast carcinoma. PMID- 10718491 TI - Reproductive factors in hereditary breast cancer. AB - BACKGROUND: An early age at menarche, a short menstrual cycle length, and a high age at first full term pregnancy or nulliparity are known risk factors for breast cancer. These risk factors have previously been reported to differ between breast cancer patients with and without a family history of breast cancer and also between breast cancer patients and controls. METHODS: Self-administered questionnaires were filled out by 95 women belonging to 24 families with known BRCA1 mutations, 16 women belonging to nine families with known BRCA2 mutations, and 95 women belonging to 65 families with hereditary breast cancer where no BRCA1 or BRCA2 mutations could be detected. Thirty-nine women were BRCA1 mutation carriers and 56 women were BRCA1 negative, 11 women were BRCA2 carriers and five BRCA2 negative. All women were born between 1905 and 1979. RESULTS: Age at menarche, physiological menstrual cycle length at age 30 or at current age in younger women (when not using oral contraceptives), age at first full term pregnancy, and nulliparity did not significantly differ between BRCA1 mutation carriers and BRCA1 negative women. Too few women were BRCA2 negative to serve as a control group. BRCA2 mutation carriers were therefore compared with BRCA1 negative and BRCA2 negative women. None of the above reproductive factors did significantly differ between BRCA2 mutation carriers and from BRCA1 and BRCA2 families. Women from non-BRCA1/BRCA2 hereditary breast cancer families had a higher age at menarche, but this was no longer significant after adjustment for other factors in a multivariate model. CONCLUSION: Our results suggest that reproductive risk factors of breast cancer are not related to BRCA1 or BRCA2 carrier status. There was also no indication that these factors differ in carriers of unknown susceptibility genes compared with non-carriers from BRCA1 and BRCA2 families. PMID- 10718492 TI - Analysis of time-space clustering in veterinary epidemiology. AB - Techniques useful for investigating time-space interaction in the clustering of events in veterinary epidemiology--the Mantel test, Barton's method, nearest neighbour test and Knox's test--are described. The use of these techniques is demonstrated by the analysis of a data set (containing blowfly catches on a commercial sheep property located in Tasmania, Australia, during the period December 1997-May 1998) in which clustering was apparent. Significant (P<0.05) clustering of blowfly catches in the data set was only detected by Knox's test. The use of Knox's test provided an insight into the possible clustering of blowfly catches over short (< or = 3 km) spatial and temporal (< or = 1 month) distances. Results demonstrate that several techniques should be used when attempting to identify and describe whether events are clustered in time and space (to maximise the power of the analysis). Also, it is important to consider the spatial and temporal models implicit in techniques chosen when interpreting results of analysis. PMID- 10718493 TI - Natural transplacental infection of dairy calves with bovine immunodeficiency virus and estimation of effect on neonatal health. AB - Many experimental infection studies with bovine immunodeficiency virus (BIV) have been conducted, but neither virus transmission under natural conditions nor longitudinal clinical effects of naturally occurring infections in non experimental populations are well explored. We tested the hypotheses that BIV is transmitted across the placenta during gestation and that intragestionally infected calves are at increased risk of neonatal disease. A cohort of 59 dairy cows on one farm were enrolled at parturition and the BIV serostatus of the cows and their pre-colostral calves determined with an indirect fluorescent-antibody assay. Moreover, the enrolled calves were monitored thrice weekly for specific clinical signs through the duration of the 30 day neonatal period and the occurrence of clinical signs analyzed for association with calf pre-colostral BIV serostatus and dam BIV serostatus. Confounding due to calf passive immunity and season of birth were also explored. Forty percent of seropositive cows (14/35) gave birth to seropositive calves but no seropositive calves (0/19) were born to seronegative dams (estimated relative risk 16, 95% exact confidence interval 2.6 5.8 x 10(29)). Calf pre-colostral BIV serostatus was not associated with the occurrence or frequency of clinical signs--but dam BIV serostatus was associated with the odds of occurrence of calf hyperthermia and with the frequency of occurrence of calf hyperthermia and hyperventilatory events. This study is inconclusive about the effects of prenatal BIV infection on neonatal health--but it does provide evidence for the natural occurrence of transplacental BIV infection. PMID- 10718494 TI - Cryptosporidia on dairy farms and the role these farms may have in contaminating surface water supplies in the northeastern United States. AB - The prevalence and risk factors for shedding of cryptosporidia by dairy cattle and calves and the prevalence and risk factors for cryptosporidia in surface waters associated with dairy farms were determined for a well-defined watershed in the northeastern United States. Eleven dairy farms were enrolled in the study and subjected to monthly sampling over a 6-month period. Animal-, water-, and manure-management practices were determined by survey and fecal, on-farm water, and stream samples were obtained monthly and evaluated for the presence of cryptosporidia. Ninety-one percent of the dairy farms in our study had Cryptosporidium on their premises. Fifteen percent of the sampled calves 0-3 weeks of age were shedding cryptosporidia. The risk factors for calves shedding cryptosporidia were contact between calves and frequent bedding changes. The probability of shedding cryptosporidia decreased with age. Nine percent of farm associated stream samples were cryptosporidia-positive. The single risk factor for detecting cryptosporidia in surface water was increasing frequency of spreading of manure on fields. The probability of detecting cryptosporidia in streams decreased as 5-day cumulative precipitation increased. There were no animal-associated or barnyard-management features associated with detecting cryptosporidia in farm-impacted streams. PMID- 10718495 TI - Mortality and recovery of runt white sturgeon (Acipenser transmontanus) in a commercial farm in California, USA. AB - We investigated the effect of raising runt white sturgeon (Acipenser transmontanus) separately from dominant fish during the initial stages of grow out in a commercial farm. Runt fish are poor-growers, have underdeveloped muscle mass, swim slowly and are more-frequently found at the top of the water column. The objective of the study was to describe the mortality and recovery rates (and their determinants) of white-sturgeon runts after separating them from dominant fish. Runt white sturgeon were stocked into twelve 2 m x 2 m rectangular tanks and graded periodically during a follow-up of 46-102 days. Overall mortality rates ranged from 0.3 to 7 dead fish per 1000 sturgeon-days at risk and overall recovery rates from 3.9 to 13.5 recovered fish per 1000 sturgeon-days at risk. Period-specific mortality and recovery rates increased over time. The period specific mortality rates for all three periods were significantly higher for tanks of runts originating from grow-out tanks with high mortality (p-values: first period = 0.06; second period = 0.09; third period = 0.03), but were similar for tanks of runts of high- and low-mean initial weight. The period-specific recovery rates were significantly higher in runts originating from high-mortality grow-out tanks only for the third period (p = 0.05) but not the first and second periods (p-values = 0.33 and 0.25, respectively). Recovery rates were significantly higher in the higher-mean-weight runts tanks for the first and third period but not for the second (p-values: first period = 0.02; second period = 0.65; third period = 0.06). We concluded that the proportion of runts that recover during a 46-89 day period is substantial (16-58%); therefore, it might be worthwhile growing such fish separately in a fish farm for about three months. Financial analysis showed that this practice was profitable, if the value of white sturgeon fish for the farm exceeded $2.05 per kg. PMID- 10718496 TI - Growth of white sturgeon (Acipenser transmontanus) following recovery from the stunted stage in a commercial farm in California, USA. AB - Runt white sturgeon (Acipenser transmontanus) develop during grow-out and are characterized by atrophied muscles and decreased growth. Our first objective was to compare the growth (and body condition) of previously-runt white sturgeon after they recovered from the runt state and sturgeon that had never been runts. On 12 occasions, recovered runts and age- and size-matched controls that had never been runts were tagged and put in a tank that already contained fish of similar age and size. Tagged groups were followed for 119-134 days. Median relative growth rates (RGRs) of the recovered runts were significantly (p < or = 0.05) higher than those of the controls in three tanks. Multiple linear regression was used to model final weight as a function of initial weight and status (recovered runt or control). Status was not significantly related (p = 0.71) to final weight, after adjusting for initial weight, "tank" and time of follow-up. Our second objective was to determine factors that influenced the loss of tags by white sturgeon during the follow-up period. Logistic regression analysis indicated that higher initial weight and being a control fish might have been associated with losing both tags. We concluded that once white sturgeon runts recovered and started growing, they grew at least as well as fish that had never been runts. PMID- 10718497 TI - Associations between genetics, farm characteristics and clinical disease in field outbreaks of porcine reproductive and respiratory syndrome virus. AB - Porcine reproductive and respiratory syndrome (PRRS) is a disease of domestic swine characterized by exceptionally high clinical variability. This study addresses the question of whether clinical variability in PRRS results from (a) genetic variation among viral isolates and/or (b) variation in management practices among farms on which isolates are found. Genetic data (open reading frame 5 gene sequences) and data on farm characteristics and associated clinical disease signs were collected for 62 PRRS virus (PRRSV) field isolates, representing 52 farms. Clinical disease signs were interrelated--confirming that a true reproductive syndrome exists (involving abortions, infertility in sows, deaths of sows and preweaning mortality). Pairs of farms experiencing deaths in their sow populations also tended to share viral isolates which were more similar to one another than expected by chance alone. This implies that sow death (one of the more-severe manifestations of PRRS) is under genetic influence. Large herd size was a significant risk factor for the death of sows and for respiratory disease in nursery pigs. All-in-all-out management practices in the nursery were protective against reproductive signs in the sow herd. All-in-all-out management practices in the finishing stages of production were protective against respiratory disease in nursery pigs--but were paradoxically associated with an increased risk of infertility in sows. These results suggest that farm-management practices can also influence which PRRS clinical signs are manifested during an outbreak. In general, signs associated with PRRS appear to result from a combination of genetic factors and herd-management characteristics. The relative contributions of these two influences differ depending on the specific clinical sign in question. PMID- 10718498 TI - What physicians don't know about their female patients. PMID- 10718499 TI - Increasing breast and cervical cancer screening among women with disabilities. PMID- 10718500 TI - Toward optimal health: the experts discuss cosmetic surgery. PMID- 10718501 TI - Baseline characteristics of participants in the Women's Health Study. AB - The Women's Health Study (WHS) is a randomized, double-blind, placebo-controlled trial designed to evaluate the balance of benefits and risks of low-dose aspirin and vitamin E in the primary prevention of cardiovascular disease and cancer in women. A total of 39,876 female health professionals, age 45 years or older and without a history of cardiovascular disease or cancer (other than nonmelanoma skin cancer), were randomized in a 2x2 factorial design to one of four treatment groups: active aspirin and vitamin E placebo, aspirin placebo and active vitamin E, both active agents, or both placebos. The process of randomization was successful, as evidenced by the equal distribution of a large number of baseline demographic, lifestyle, and health history characteristics among the four treatment groups. Similar distribution of known potential confounders, as well as the large sample size, provides reassuring evidence that unmeasured or unknown potential confounders are also equally distributed. As expected in a clinical trial, the women in the study are healthier in some respects than the general population, but they have very comparable rates of obesity, hypertension, and elevated cholesterol. With adequate duration of treatment and follow-up, this trial will provide important and relevant information on the balance of benefits and risks of aspirin and vitamin E supplementation in the primary prevention of cardiovascular disease and cancer in women. PMID- 10718502 TI - Coronary heart disease: sexual bias in referral for coronary angiogram. How does it work in a state-run health system? AB - This study examined the effects of a state-run health system on the gender specific differences in cardiology worldwide, taking coronary angiography as an example. In a prospective study, 476 angiographed patients (155 female, 321 male) were enrolled in consecutive order over a study period of 9 months and asked to answer a standardized questionnaire. The responses showed a discrepancy in the heart death statistics (52.7% female, 47.3% male) and the demographic statistics (51.8% female, 48.2% male). This was true for all age groups. The duration of complaints before undergoing a coronary angiogram was reported to be acute for 4.5% of the women and 13.7% of the men, <1 year for 27.1% of the women and 34% of the men, and >1 year for 68.4% of the women and 52.3% of the men. Women take longer to access coronary angiogram. This is confirmed by New York Heart Association (NYHA) classes I (1.9% female, 7.8% male), II (46.5% female, 65.4% male), III (41.9% female, 21.8% male), and IV (9.7% female, 5.0% male). Prior to angiogram, all of the women and most of the men (98.4%) were under treatment for heart complaints, more women (87.1%) than men (78.8%) took heart medication, but fewer women (29.7%) than men (37.1%) had been referred to a cardiologist. Major differences were seen in the social situation; that is, 68.4% of the women but 93.5% of the men lived with their family, 30.3% of the women but only 5% of the men lived alone, and 1.3% of the women together with 1.6% of the men lived in a care-giving facility. The results of our study show that even in a state-run health system with free access to high-tech medicine at no charge and no age limits, there is a marked gender bias in access to high-tech medicine. PMID- 10718503 TI - Menstrual patterns during the inception of perimenopause: what are the predictors and what do they predict? AB - Using data from a British national cohort of women born in 1946, this study aims to identify menstrual patterns during the first year of perimenopause (based on the frequency of periods, the numbers of days bled each month, and menstrual flow) to see if they are related to health and behaviors earlier in adult life and if they predict entry into menopause and hormone replacement therapy (HRT) use. Three groups of women were identified using cluster analysis: those who experienced more of these characteristics, those who experienced less, and those who experienced few changes. In polychotomous logistic regression models, the likelihood ratio tests indicated that parity and body mass index (BMI) were significant at the 5% level. The odds ratios from the parity models showed a gradient, with women from the Less cluster being most likely to have no children and those from the More cluster most likely to have at least one child. A similar gradient was detected for BMI, with the Less cluster tending to be underweight. The Less cluster came into menopause significantly faster than the Same and the More groups, where the estimated hazard ratios (HR) (95% confidence interval [CI]) were, respectively, 0.61 (0.37-0.99) and 0.24 (0.11-0.52). There was no association between the clusters and later HRT use. The findings suggest that menstrual characteristics should be more carefully studied in population studies of the climacteric. PMID- 10718504 TI - Successful management of a viable cervical pregnancy by single-dose methotrexate. AB - Cervical pregnancy is very uncommon and carries a high risk for hysterectomy with surgical treatment. Prior reports of medical treatment included various regimens of high-dose systemic methotrexate (MTX) with citrovorum rescue and local injection. This is the first report of successfully treating a viable cervical pregnancy with single-dose i.m. MTX, followed by a vaginal delivery. PMID- 10718505 TI - Acceptability of human papillomavirus immunization. AB - The purpose of this study was to examine the attitudes about hypothetical human papillomavirus (HPV) vaccines in two groups of women in clinical settings. Twenty adolescent women attending an urban community adolescent health clinic and 20 adult women attending a city health department sexually transmitted disease (STD) clinic were recruited to participate in individual interviews. Adolescents were 14-18 years of age (mean 15.6), 75% nonHispanic white, and 75% sexually experienced. Adults were 20-50 years of age (mean 33.6), 95% African American, and all were sexually experienced. As part of the interview, participants ranked nine hypothetical HPV vaccines in order of acceptability. Each vaccine was uniquely defined as a function of cost ($150, $50, or free), efficacy (50% or 90%), disease targeted (genital warts, cervical cancer, or both), and physician recommendation (not mentioned by a physician or specifically recommended). Rankings by adolescents and adults were highly concordant (Spearman rho = 0.9). Efficacy, physician's recommendation, and cost influenced rankings most strongly. Ranking decisions were often based on complex decision making, in which all characteristics were considered simultaneously. These findings suggest that certain features of an HPV vaccine might significantly affect vaccine acceptability. Vaccine efficacy, physician endorsement, and cost were particularly salient issues. PMID- 10718507 TI - Knowledge of menopause and hormone replacement therapy use in low-income urban women. AB - Hormone replacement therapy (HRT) can have significant long-term health benefits in postmenopausal women, yet rates of HRT use are low, especially in low-income urban women. Previous research has revealed that knowledge of menopause is a key predictor of HRT use in this population. A descriptive cross-sectional survey of 215 perimenopausal and postmenopausal low-income urban women was carried out to characterize knowledge of menopause and HRT and factors associated with knowledge level. Sociodemographic characteristics, patterns of HRT use, and knowledge about menopause and HRT were collected through a structured interview. Results revealed a general lack of knowledge about menopause and HRT, particularly relative to heart disease and the role of HRT in prevention. Major independent predictors of increased knowledge (R2 = 0.31) were having talked with a healthcare provider about HRT, having at least a high school education, and being less than 60 years of age. These findings emphasize the key role of providers in educating this vulnerable population about menopause and HRT and the potential subsequent impact on HRT use. PMID- 10718506 TI - Comprehensive medical care among HIV-positive incarcerated women: the Rhode Island experience. AB - Our objective was to characterize the clinical presentation of human immunodeficiency virus (HIV) infection among incarcerated women in a program that provides HIV testing and primary care to all state prisoners in Rhode Island. A retrospective medical chart review on all HIV-seropositive women who were incarcerated between 1989 and 1994 and had at least two medical visits with an HIV medical care provider was used. At the Rhode Island Adult Correctional Institution (ACI), under mandatory testing laws between 1989 and 1994, 28% (172 of 623) of all women were identified with HIV infection. Of the 172 women who tested seropositive in prison, 110 were included in the study. Of the 110 women followed, 84% reported injection drug use (IDU) as their primary risk factor, and 30% reported both IDU and sex work. The median CD4 count was 596/mm3, with 60% having a CD4 count >500 cells/mm3. The most common medical conditions were vaginal candidiasis, oral candidiasis, and bronchitis. Antiretroviral therapy was well accepted and followed community standards. Continuity of medical care after release was facilitated by the same physician caring for the patient in the community setting, with 83% of women following up for HIV care after release. The medical conditions noted reflect that these women are early in the course of their HIV disease when they are initially diagnosed. This comprehensive program in Rhode Island's state prison plays a central role in the diagnosis of HIV seropositive women and provides counseling, primary medical and gynecological care, and linkage to community resources after release. PMID- 10718508 TI - Moderate intensity exercise training improves cardiorespiratory fitness in women. AB - Among women, there is an increased prevalence of sedentary lifestyle and less participation in physical activity at levels recommended by the Surgeon General. As a result, women have been identified as a target group in public health initiatives to increase physical activity. The health-related benefits of habitual, moderate intensity physical activity are well documented in the epidemiological literature, but less is known about the effect of such physical activity on cardiorespiratory fitness. Our hypothesis was that moderate and vigorous exercise training regimens of similar estimated energy expenditure would result in similar changes in cardiorespiratory fitness. Eighteen sedentary premenopausal women with the following baseline characteristics [x +/- SE]: maximal oxygen consumption (Vo2max) = 29.5+/-1.5 ml x kg(-1) x min(-1); age = 33+/-1 years; height = 162.6+/-0.9 cm; mass = 62.7+/-2.3 kg, were randomly assigned to either vigorous (HI, 80% Vo2max, n = 10) or moderate intensity (MOD, 40% Vo2max, n = 8) cycle ergometer training groups. Exercise training was conducted 3-4 (3.37+/-0.05) days/week for 12 weeks in a supervised and progressive manner, with estimated exercise energy expenditure equated across both training groups. Vo2max and time to exhaustion increased significantly in both groups (p<0.05), with no difference between groups. Both groups had lower (p<0.05) posttraining submaximal heart rates (HR), respiratory exchange ratios (RER), and ratings of perceived exertion (RPE) during graded exercise testing, with no significant differences between the groups in posttraining values. Women participating in moderate intensity exercise training as recommended in basic public health guidelines demonstrate an increase in cardiorespiratory fitness similar to that elicited by vigorous training. PMID- 10718509 TI - Mandatory reporting of intimate partner violence: safety or retaliatory abuse for women? AB - We wished to ascertain abused women's perspectives on mandatory reporting of intimate partner violence. A consecutive sample of 161 abused women accessing the criminal justice system were asked six questions. Most women (81%) thought there should be a law making the nurse or doctor report the abuse, with less than half of the women (45%) reporting they would have been at greater risk for abuse following a mandatory report. Although these 161 abused women support mandatory reporting of partner abuse, it remains unclear if such a law would deter further abuse. Outcome and evaluation studies of mandatory reporting laws are urgently needed. PMID- 10718510 TI - New generation search engines. PMID- 10718511 TI - Gene transfer into hippocampal slice cultures with an adenovirus vector driven by cytomegalovirus promoter: stable co-expression of green fluorescent protein and lacZ genes. AB - Virus-mediated gene transfer into identified neurons of organotypic hippocampal slice cultures offers a great potential for studying the cellular and molecular mechanisms of synaptic plasticity. We describe here a new adenovirus vector Ad GFP-lacZ carrying an early cytomegalovirus (CMV) gene promoter that efficiently co-transferred the beta-galactosidase (lacZ) and green fluorescent protein (GFP) genes in rat organotypic hippocampal slice cultures. Monitoring of GFP fluorescence and immuno-histochemical staining for beta-galactosidase showed that the expression of the transferred genes was widespread in the glial cells and neurons of CA1, CA3/4, and dentate gyrus regions. Immunoblot analyses showed that the expression of gamma-galactosidase and GFP was maximal about 48 h after infection of hippocampal slices with the adenovirus vector and the expression levels were maintained for several weeks. Also, immunoblot analyses showed no significant differences in the MAP-2 and glial fibrillary acidic protein levels in the adenovirus vector infected and uninfected hippocampal slices. In addition, we found that the infection of hippocampal slices with the adenovirus vector caused no significant increase in the induction of heat shock protein (HSP)-70 and showed no change in their electrophysiological properties as measured by stable field synaptic potentials in CA1 region and its reactivity to high frequency stimulation. Our data suggest that this adenovirus vector can be exploited to transfer multiple genes into neurons and may have implications for developing strategies for gene therapy. PMID- 10718512 TI - Co-grafts of muscle cells and mesencephalic tissue into hemiparkinsonian rats: behavioral and histochemical effects. AB - Extracts from skeletal muscle cell cultures have been shown to increase levels of the enzyme tyrosine hydroxylase (TH) and promote survival of different types of developing neurons in vitro. To determine the effect of muscle cell co-grafts on the survival of dopamine neurons in a rat model of Parkinson's disease, we transplanted an embryonic day (ED)-15 rat mesencephalic cell suspension alone or with neonatal muscle cells into 6-hydroxydopamine (6-OHDA) denervated rat striatum. In parallel experiments conducted in vitro, we cultured ED-15 rat mesencephalon or rat striatum in conditioned medium from neonatal rat muscle cultures (MC-CM). Our results showed that: (A) in vitro, MC-CM increased the number of TH-immunoreactive (TH-IR) neurons in embryonic mesencephalic cultures but did not induce expression of TH in embryonic striatal cultures; (B) in vivo, animals with co-grafts of muscle cells and ED-15 mesencephalon had more TH-IR in the grafted striatum compared to animals that received mesencephalic cells grafts alone, although the graft-induced reversal of circling behavior in response to methamphetamine was the same in both transplanted groups; and (C) grafts of muscle cells alone did not induce TH-IR in the denervated striatum and did not reduce methamphetamine-induced circling. These findings suggest that in vivo, neonatal muscle cells secrete factors that promote survival and/or outgrowth of fetal midbrain dopamine cells and improve the levels of TH-IR in grafted striatum. PMID- 10718513 TI - Cholinergic enhancement improves performance on working memory by modulating the functional activity in distinct brain regions: a positron emission tomography regional cerebral blood flow study in healthy humans. AB - Previously, we have shown that physostigmine, an acetylcholinesterase inhibitor, improved performance on a working memory for faces task, as reflected by reduced reaction time (RT), and reduced task-specific regional cerebral blood flow (rCBF) in right prefrontal cortex and, further, that these reductions in RT and right frontal rCBF were significantly correlated. Here we investigated the relation between the effects of physostigmine on task performance and task-specific functional brain response throughout the cortex by examining correlations between physostigmine-related changes in rCBF in all brain areas and changes in RT. In subjects who received an infusion of physostigmine, reduced RT correlated (p<0.001) positively with reduced rCBF in right frontal cortex, left temporal cortex, anterior cingulate, and left hippocampus; and correlated with increased rCBF in medial occipital visual cortex. In subjects who received a placebo infusion of saline, no significant correlations between changes in RT and cortical rCBF were observed. The results show that cholinergically induced improvements in working memory performance are related to alterations in neural activity in multiple cortical regions, including increased neural activity in regions associated with early perceptual processing and decreased neural activity in regions associated with attention, memory encoding, and memory maintenance. PMID- 10718514 TI - Efferent connections of the anterior hypothalamic nucleus: a biocytin study in the cat. AB - The efferent connections of the anterior hypothalamic nucleus (AH) were examined using biocytin as anterograde tracer in the cat. The results provide several new findings in addition to confirming earlier observations. In the hypothalamus, the AH projections terminated mainly in the medial regions which are related to the defensive, reproductive and feeding behaviors, and autonomic functions. Moreover, we found dense patches of the AH terminals in the medial preoptic area and ventromedial hypothalamic nucleus, which suggests the existence of modular connections between sub-regions of each nucleus. In addition, the AH projected to regions which may be related to the emotional and autonomic responses, i.e., such regions in the amygdala, midline thalamus, septum, subthalamus, and midbrain. The data suggest that the AH may play an important role in the autonomic functions and behaviors between animals, and thus may play a key role in the defensive behavior elicited in the medial preoptic area and ventromedial hypothalamic nucleus. PMID- 10718515 TI - Cellular electrophysiological changes in the hippocampus of freely behaving rats during local microdialysis with epileptogenic concentration of N-methyl-D aspartate. AB - N-methyl-D-aspartate (NMDA) receptor dysfunctions are thought to be involved in the pathophysiology of seizures of hippocampal origin. While the cellular effects of excessive NMDA receptor stimulation have been widely studied in vitro, no data are available on the sequence of cellular electrophysiological events that follow the overstimulation of hippocampal NMDA receptors in awake, behaving subjects. Therefore, the present study addressed this problem. Intrahippocampal microdialysis with 500 microM NMDA was performed in freely behaving rats, and the electrical activity of single neurons in the dialysis area were monitored. In all recorded neurons (n = 9), regardless of their type, NMDA induced a long-lasting electrical silence preceded in most cells by a brief but robust firing rate increase. During these firing rate increases, place cells lost the spatial selectivity of their discharges, and a gradual reduction in the amplitude of the action potentials was also observed. Remarkably, electroencephalographic (EEG) seizures developed exclusively after the appearance of cellular electrical silence in the recording/dialysis site. The NMDA-induced electrophysiological changes were reversible. This study demonstrates that the combined single-cell recording-intracerebral microdialysis technique can be readily used for inducing focal epileptiform events in the hippocampus and monitoring the induced cellular electrophysiological events in behaving animals. PMID- 10718516 TI - Anticonvulsive effect of AMPA receptor antagonist GYKI 52466 on 4-aminopyridine induced cortical ictal activity in rat. AB - The effect of GYKI-52466 (1-(4-aminophenyl)-4-methyl-7,8-methylenedioxy-5H-2,3 benzodiazepine), a selective antagonist of AMPA receptor was investigated on the generation and manifestation of 4-aminopyridine-induced cortical epileptiform activity. In vivo experiments were carried out on pentobarbital-anaesthetised adult rats. Ictal epileptiform activity was induced by local application of 4 aminopyridine (4-Ap) to the surface of somatosensory cortex. In one group of animals, GYKI 52466 was administered intraperitoneally before 4-Ap application, in another group, the already active primary focus was treated locally by GYKI 52466. Different parameters of epileptic activity were measured and compared in GYKI 52466-treated and control animals. The results demonstrate that GYKI 52466 exerts anticonvulsive effects on both the induction and the expression of epileptiform activity, by delaying the onset of the first ictal event, decreasing the numbers and duration of ictal periods, as well as the amplitudes of epileptiform discharges both in the primary and mirror foci. However, seizure propagation to other cortical areas seemed to be facilitated. The anticonvulsive effect of GYKI 52466 was stronger in pretreatment than in treatment of ongoing epileptiform activity. As a conclusion, it is supposed that AMPA receptors are probably more dominant in the induction of epileptiform activity than in the maintenance of it, mainly through the activation of corticothalamo-cortical networks. It is also supposed that the cortical inhibition which blocks the propagation of epileptiform process might be activated mainly through non-N methyl-D-aspartate receptors. PMID- 10718517 TI - Individual variability of dopamine release from nucleus accumbens induced by nicotine. AB - Effects of subcutaneous administration of vehicle, amphetamine (1 mg/kg) or nicotine (0.4 mg/kg, injected twice, 90 min apart) on extracellular dopamine (DA) concentration in the nucleus accumbens (ACC) and ventral tegmental area (VTA) of the Sprague-Dawley rat were studied using microdialysis. Experiments were conducted at least 10 days following implantation of guide cannulae, and at least 2 h following insertion of microdialysis probes into the guides on the morning of each experiment. Probes were perfused at 2.5 microl/min and several fractions were collected every 10 min before and after the two test injections. Samples were analyzed by high-performance liquid chromatography with electrochemical detection for the major neurotransmitters and their metabolites. Significant DA release following nicotine administration was observed in ACC but not in VTA. By classifying ACC DA responses of individual rats, three major subgroups were identified which exhibited more robust responses. Nicotine appeared to be acting as a modulator of ACC DA, increasing DA output if baseline was <5 nM, but slowing release when the baseline exceeded 5 nM. These data are consistent with previous reports of modulation of arousal level by nicotine via DA. PMID- 10718518 TI - Dendritic organization of neurons of the superior colliculus in animals with different visual capability. AB - The aim of the study was to compare several morphological characteristics of neurons in the superficial layers of the superior colliculus in diurnal and nocturnal mammals with different visual specialization. Thus, we investigated the rat (Rattus norvegicus), a nocturnal animal; the tree shrew (Tupaia glis), a diurnal animal, and the Mongolian rodents, Microtus brandti (nocturnal) and Alticola barakshin (diurnal). The investigation was focused on the study of the organization and extent of dendrites of Golgi-impregnated projection neurons, which were divided in two classes: narrow-field and wide-field cells. We determined that the ratios between the volumes of dendritic fields of the investigated neuronal types and the total volume of the superior colliculus differed to a great extent between the different species. The tree shrew had the largest superior colliculus and the smallest wide-field neurons, while the rat had the largest wide-field neurons. As for the Mongolian rodents, we provided the first description of superior colliculus neurons. The day-active animal Alticola barakshin was found to have a 50% larger volume of the superior colliculus than that of the night-active animal Microtus brandti, and the size of the dendritic field of both wide-field neurons and narrow-field neurons was smaller than that of Microtus brandti. Electron microscopic investigation of wide-field neurons performed in the rat revealed only a few symmetric synaptic contacts on the arborizations of distal and terminal dendrites and numerous asymmetric synapses on the dendritic stem. Our findings support the hypothesis that whereas the narrow-field neurons are relay neurons in the retino-tecto-thalamic pathway of the visual system, the wide-field neurons may play additional roles in the retino tecto-reticulo-spinal system. PMID- 10718519 TI - Prenatal morphine exposure differentially alters TH-immunoreactivity in the stress-sensitive brain circuitry of adult male and female rats. AB - Previously, we demonstrated that exposure to morphine during gestation increases hypothalamic norepinephrine (NE) content and turnover rate in adult male rats and decreases these measures in adult females. To investigate the basis of these alterations, the present study examined the effects of prenatal exposure to morphine on tyrosine hydroxylase immunoreactivity (TH-IR) in the brains of adult male and female progeny. In male rats, prenatal morphine exposure significantly increased the density of TH-IR in cells and fibers in the caudal paraventricular nucleus of the hypothalamus (PVN) and locus coeruleus (LC), but had no effects in the lateral hypothalamus (LH). In female rats that were ovariectomized (OVX), prenatal morphine exposure significantly decreased the density of TH-IR in cells and fibers in the LC. Interestingly, an injection of estrogen in OVX control females reduced the mean optical density of TH-IR in the LC, but it was ineffective in drug-exposed females in the same brain region. Estrogen injections also reduced the mean optical density of TH-IR in the LH but not in the PVN of females, regardless of prenatal drug exposure. Thus, the present study suggests that prenatal morphine exposure induces long-term, sex-specific alterations in TH IR in the PVN and LC of adult progeny. PMID- 10718520 TI - Intraventricular 2-deoxy-D-glucose induces Fos expression by hypothalamic vasopressin, but not oxytocin neurons. AB - The glucostatic theory supports the role of central and peripheral substrate "sensors" in monitoring cellular glucose metabolism. Induction of hyperphagia and hyperglycemia by intracerebroventricular (i.c.v.) delivery of drugs inhibiting glucose uptake or oxidation suggests that glucose "sensors" are accessible from the cerebroventricular system. Although glucopenia elevates neurohypophyseal vasopressin (VP) and oxytocin (OXY) secretion and induces c-fos expression by hypothalamic paraventricular (PVN) and supraoptic (SON) neurons, the origin of glucoprivic regulatory signals impinging upon these cell populations is unclear. The following study evaluated immunolabeling of hypothalamic VP and OXY neurons for the nuclear transcription factor, Fos, following systemic vs. i.c.v. delivery of the glucose antimetabolite, 2-deoxy-D-glucose (2DG). Intraperitoneal drug treatment resulted in Fos expression by a high proportion of AVP- and OXY-ir neurons in the PVN and SON, whereas i.c.v. antimetabolite administration resulted in immunostaining of a smaller proportion of AVP neurons and a lack of colabeling of OXY neurons in both sites. These results suggest that decreased glucose metabolism within the periventricular CNS is a stimulus for central mechanisms that activate the Fos stimulus-transcription cascade in a discrete subpopulation of VP neurons in the PVN and SON. Alternatively, the absence of demonstrable Fos expression by OXY neurons in the same structures suggests that the functional status of these cells is regulated by glucoprivic stimuli of peripheral and/or nonperiventricular central origin. PMID- 10718521 TI - The brain tumor of George Gershwin and the legs of Cole Porter. AB - George Gershwin died in 1937 of a glioblastoma of the right temporal lobe. He had been in psychoanalytical care for some time and was hospitalized a few weeks before his death, when he was thought to have a functional illness. The controversies about George Gershwin's death, duration of neurologic symptoms, and problems in diagnosis are discussed. Cole Porter fell off a horse he was riding in 1937 and sustained multiple open fractures of both legs. There probably was some nerve injury in the right leg, at least, from this fall. Despite intensive pain, many hospitalizations, and 33 operations on his legs, Porter continued to write music and lyrics until his amputation in 1958. After the amputation, all creative activities ceased. The explanations for this are discussed. PMID- 10718522 TI - Glenn Gould: insights into the cause of death of the great pianist. PMID- 10718523 TI - Robert Schumann. AB - Robert Schumann, one of the giants of early romantic music, was born in Saxony in 1810 and died in an asylum shortly after his 46th birthday. Early in life, he demonstrated extraordinary skills in both music and journalism; he remained active in both areas until his final illness. His marriage to the remarkable pianist, Clara Wieck, provided him with both much-needed emotional support and a highly effective champion of his music throughout her lengthy career. Schumann's plans to be a concert pianist were thwarted at least partially by an injury to his right hand, the nature of which has been the subject of much speculation. After considering what few facts are available, the author concludes that this may have represented focal dystonia. His compositional output waxed and waned dramatically over his professional life, reflecting to some degree his emotional state. It is considered most likely that he suffered from a major affective disorder, bipolar type. This ultimately led to a suicide attempt in February 1854, and to his eventual death in July 1856. Despite wide-spread and reasonable suspicion that he may have died from neurosyphilis, severe malnutrition from self starvation seems more likely. PMID- 10718524 TI - Musical tradition, insurrection, and resurrection: the life and legacy of composer/bassist Charles Mingus. AB - Charles Mingus was perhaps the foremost straight-ahead jazz upright bassist and composer of his generation, blending the inspirational influences of gospel, jazz improvisation, and art music leanings into a unique style all his own. His demise from amyotrophic lateral sclerosis (ALS) in his fifth decade robbed the world of one of the great creative voices of American music. Aspects of Mingus' life, his career as a bassist, bandleader, and composer, and his neuromuscular illness are discussed, emphasizing his legacy for the disparate fields of jazz and neurology. PMID- 10718525 TI - The painful perils of a pair of pianists: the chronic pain of Clara Schumann and Sergei Rachmaninov. AB - Clara Wieck Schumann (1819-1896) and Sergei Vassilievich Rachmaninov (1873-1943) were two of the greatest pianists that ever lived. They had full lives composing and performing music. Each also had more than his or her fair share of hardships. In addition to all the pressures that are part of a performer's life, both Clara Schumann and Sergei Rachmaninov also suffered from chronic pain. This article discusses the pain syndromes that plagued these great musicians and the effect of chronic illness on their music. PMID- 10718526 TI - Neurosyphilis in musicians and composers. PMID- 10718527 TI - Musicians' dystonia: the case of Gary Graffman. AB - A professional pianist developed career-ending focal dystonia. There is a possible relationship between the pianist's syndrome and his past playing history. Although the patient has derived very little benefit from various treatment modalities, his candor regarding his impairment indirectly led to the establishment of performing arts medicine as a recognized subspecialty of occupational medicine. PMID- 10718528 TI - The life, legacy, and premature death of Felix Mendelssohn. AB - Felix Mendelssohn is one of the great classical composers of all time. During his short lifetime in the first half of the nineteenth century, he reached enormous heights as a composer, conductor, and leader in the world of music. Nearly one hundred years after his death, the Nazi regime attempted, unsuccessfully, to erase his music and his memory from history. Since the end of World War II, there has been a resurgence in interest in the life and music of Felix Mendelssohn and that of his sister, Fanny. Felix Mendelssohn died in 1947 at the age of 38. Both of his sisters died suddenly at the ages of 42 and 45. There is insufficient laboratory or post-mortem data to make a medical diagnosis with certainty. However, based on the information available to us, we speculate that Mendelssohn suffered a subarachnoid or intracerebral hemorrhage. The differential diagnosis of familial stroke syndrome is discussed. PMID- 10718529 TI - Ravel's neurological illness. AB - In the last 10 years of his life, Maurice Ravel (1875-1937) experienced a gradually progressive decline in neurological function. Dr. Alajouanine examined Ravel, noting the presence of aphasia and apraxia with relative preservation of comprehension and memory. The exact diagnosis remains unclear, but the likelihood of a progressive degenerative disorder, such as frontotemporal dementia, is herein discussed. PMID- 10718530 TI - The tragedy of Sergei Prokofiev. AB - Sergei Sergeevich Prokofiev (1891-1953) was one of the great Russian composers. He died, at age 61, of a supposed intracerebral brain hemorrhage. During the 8 years before his death, he suffered from episodic headaches, nausea and dizziness. This article reviews the historical information about Prokofiev's illness leading to a plausible underlying etiology for his death. PMID- 10718531 TI - Shostakovich and amyotrophic lateral sclerosis. AB - Dmitri Shostakovich (1906-1975) is one of the Twentieth Century's greatest composers. Beginning in the late 1950s, he experienced gradual progressive asymmetric weakness of the limbs, and was eventually diagnosed with motor neuron disease, which is the subject of this review. PMID- 10718532 TI - Epidemiology of human immunodeficiency virus infection and associated neurologic illness. AB - Worldwide, 30 million people are infected with the human immunodeficiency virus (HIV), with almost 6 million new infections in 1997. Recent therapeutic advances, which have altered the natural history of HIV infection, and changes in the AIDS case definition, both complicate evaluations of temporal changes in AIDS incidence and prevalence, and lesson the utility of AIDS incidence as a proxy for monitoring the HIV epidemic. The highest AIDS incidence rates in the United States are in black men. Rates are increasing most quickly in women, minorities, and adolescents and young adults, largely due to heterosexual transmission and intravenous drug abuse. Survival after diagnosis of AIDS is associated most strongly with the initial AIDS-defining diagnosis, and patients with neurologic opportunistic infections or primary central nervous system (CNS) lymphoma have shorter survival periods. Neurological illnesses are the initial manifestation of AIDS in 7% to 20% of patients, but the frequency of neurologic complications increases over the course of the illness. The most common AIDS-defining opportunistic illnesses are HIV encephalopathy, CNS toxoplasmosis, cytomegalovirus retinitis, and primary CNS lymphoma. Primary prevention of HIV infection is accomplished by changing factors that enable transmission, through behavioral changes, utilization of antiretroviral agents to prevent vertical transmission, and through securing the safety of the blood supply. Secondary prevention, involving early detection and prompt treatment, has become important in developed countries since the introduction of powerful antiretroviral therapies and has contributed to the 46% decline in AIDS deaths in the United States from 1996 to 1997. Inequality of access to effective therapies and emergence of multi-drug-resistant strains of HIV have raised serious concerns. PMID- 10718533 TI - Pathobiology of human immunodeficiency virus dementia. AB - The pathobiology of dementia that accompanies infection with the human immunodeficiency virus involves complex interactions of the virus with the host. The virus enters the brain either as free viral particles or hidden in infected monocytes (the "Trojan Horse" mechanism). Within the brain it infects microglial cells, causing a productive and cytopathic infection, and infects astrocytes, causing a latent or restricted infection. The brain thus acts as an important reservoir for the virus. These infected cells release several viral proteins, some of which are toxic to neurons and are called "virotoxins." These virotoxins activate glial cells to release a number of soluble factors that are either toxic to neurons or cause chemotaxis of monocytes into the brain. Because the glial cells outnumber the neurons by 10:1, this is an important mechanism by which the virotoxins amplify their toxic potential and initiate a self-perpetuating cascade of events, resulting in a "domino effect" on the brain. Only a transient exposure to virotoxins is necessary to initiate these positive feedback loops. Thus, a "hit and run" phenomenon may be operative within the brain. Therapeutic approaches are based on decreasing the viral burden in the brain and blocking the actions of the key neurotoxic substances at various levels within the various cascades. PMID- 10718534 TI - Human immunodeficiency virus-associated dementia. AB - HIV-associated dementia will eventually develop in 15-20% of individuals with advanced HIV disease. It has become one of the leading causes of dementia in the young, with 10,000 new cases annually in the USA. The clinical syndrome includes progressive development of psychomotor slowing and memory impairment, eventually with brain atrophy and neurol loss. The pathology is characterized by infection of macrophages and microglia, marked activation of macrophages, and release of a variety of postinflammatory cytokines into the parenchyma. Antiretroviral therapy has impacted positively on the incidence rates, and at least partial reversal of neurologic deficits can be achieved in established dementias. PMID- 10718535 TI - Diseases of the spinal cord in human immunodeficiency virus infection. AB - The most common disease of the spinal cord in human immunodeficiency virus (HIV) infection is vacuolar myelopathy. Pathology studies have demonstrated that vacuolization in the thoracic spinal cord is present in more than a third of patients with AIDS. The disease, however, manifests clinically only when the vacuolization in the spinal cord has become severe, with prominent myelin loss in the lateral and posterior columns. Vacuolar myelopathy presents usually with slowly progressing spastic paraparesis, accompanied by loss of vibratory and position sense and urinary frequency and urgency. In males, erectile dysfunction can be an early manifestation of the disease. The pathogenesis of vacuolar myelopathy is unknown but may be related to abnormal trans-methylation mechanisms induced by the HIV virus and cytokines. There is no known treatment for the disease, although therapy with methylating agents is being investigated. There are other rarer causes of spinal cord disease in AIDS, including a number of infectious myelitis and neoplastic and vascular myelopathies. PMID- 10718536 TI - Neuromuscular complications of the human immunodeficiency virus type 1 infection. AB - Neuromuscular disorders are the most frequent neurologic complications associated with human immunodeficiency virus (HIV) infection and AIDS. Although neurologic disorders are frequently overlooked, they add considerable morbidity and mortality to patients with HIV infection. It is critically important to properly diagnose and treat these neuromuscular complications, which leads to substantial improvement in patients' quality of life. Distal symmetric polyneuropathy is the most common form of peripheral neuropathy in HIV infection. It occurs mainly in patients with advanced immunosuppression and may also result from the neurotoxicity of several antiretroviral agents. Myopathy may occur at any stage of HIV disease and has also been described as a toxic side effect of zidovudine. Here we review the clinical features, diagnostic approach, and pathogenetic mechanisms of the neuromuscular complications of HIV infection. We also discuss management strategies and the results of clinical trials for the treatment of these disorders. PMID- 10718537 TI - Clinical features and treatment interventions for human immunodeficiency virus associated neurologic disease in children. AB - HIV-1 infection in children and adolescents can cause progressive neurologic disease, affective brain growth, motor function, and neurodevelopment. In addition, myelopathies, neuropathies, myopathies, strokes, and psychiatric or behavioral manifestations can be a result of HIV-1 infection, OI, or toxicities of treatment interventions. CNS OI are important causes of morbidity and mortality, often mimicking the HIV-1 associated neurologic syndromes. Psychometric, clinical, neuroradiologic, and laboratory testing are valuable for diagnostic and treatment decisions. The cornerstone of treating HIV-1-associated neurologic disease is providing an effective regimen of antiretroviral drugs to reduce the viral burden. It is also necessary to provide rehabilitation, optimize nutrition, supply appropriate antimicrobial prophylaxis against OI, minimize pain, and treat neurobehavioral or psychiatric complications. Efforts at preventing HIV-1 infection are important for diminishing and allaying the growth of this international pandemic. PMID- 10718538 TI - Bacterial and fungal brain infections in AIDS. AB - Many bacterial and fungal pathogens can infect the central nervous system (CNS) in patients also infected with human immunodeficiency virus (HIV). Most of these organisms cause meningitis, and this is sometimes accompanied by focal parenchymal infection. More virulent pathogens, such as Mycobacterium tuberculosis hominis, can cause disease in individuals with mild degrees of HIV associated immunosuppression. Less virulent pathogens, such as Cryptococcus neoformans, tend to affect individuals with more advanced immunosuppression. This review focuses on the clinical manifestations, diagnosis, and treatment of the more common bacterial and fungal CNS infections. PMID- 10718539 TI - Opportunistic viral infections in the setting of human immunodeficiency virus. AB - Human immunodeficiency virus (HIV) infection has provided a setting in which additional neurologic problems develop. The mechanism of these complications varies from agent to agent, but the added spectrum of diseases encountered has challenged diagnosticians and provided unparalleled opportunities to develop a deeper understanding of these conditions and their treatments. This review addresses the most prominent viral-associated complications, except for progressive multifocal leukoencephalopathy, which is addressed in a separate review. The complications of greatest importance both due to their frequency and severity are caused by cytomegalovirus, so these are discussed in greater depth. However, the association of Epstein-Barr virus with induction of central nervous system lymphoma represents an important viral linked complication of great importance. In addition, the increased activity of varicella zoster virus has been notable in the setting of HIV. Finally, human herpesvirus type 6 is an emerging virus of interest that has been identified in the setting of HIV infection, whose role in pathophysiology is only now being investigated. PMID- 10718541 TI - Neurologic manifestations of toxoplasmosis in AIDS. AB - Central nervous system (CNS) toxoplasmosis is the most common cause of cerebral mass lesions in AIDS patients. Toxoplasma gondii is commonly acquired through ingestion of contaminated meats resulting in latent infection. With the onset of immunosuppression, it may preferentially infect the CNS, resulting in a wide range of clinical presentations. Effective antibiotic therapy is available and capable of producing rapid remission of active infection but must be continued throughout life to prevent recurrence. Characteristic presentations and rapid therapeutic response permit presumptive diagnosis and initiation of specific antibiotics in many cases; however, appropriate clinical and radiographic monitoring to detect alternative or mixed pathologies is necessary. Unusual presentations may hinder rapid diagnosis and should be considered in AIDS patients with cryptic CNS symptoms. Despite increasing attention to primary prophylaxis, the worldwide distribution of this parasite, its potential to be the presenting illness in previously unidentified human immunodeficiency virus infected individuals, and failures of prophylaxis are likely to make toxoplasmosis an important continuing source of neurologic morbidity in AIDS. PMID- 10718540 TI - Progressive multifocal leukoencephalopathy. AB - Before the AIDS epidemic, progressive multifocal leukoencephalopathy (PML) was a rare disorder occurring most often in association with leukemia and lymphoma. Current estimates indicate that PML ultimately develops in up to 5% of all patients with AIDS. This demyelinating disease results from infection with JC virus, a papova virus, that most of the world's population is exposed to prior to adulthood. Although PML commonly occurs in the setting of advanced immunosuppression, it may be observed in patients with CD4 lymphocyte counts in excess of 200 cells/mm3. Focal neurological symptoms and signs coupled with hyperintense signals abnormalities of the white matter on T2-weighted cranial magnetic resonance imaging are highly suggestive of the disease. In this setting, a positive CSF polymerase chain reaction for JCV DNA has been felt to be sufficiently diagnostic to eliminate the need for brain biopsy. Survival of AIDS associated PML is poor with median survivals averaging just 6 months. However, as many as 10% of AIDS patients with PML will have prolonged (>12 months) survival and partial recovery. Highly active antiretroviral therapy (HAART) has been demonstrated to have a salutary effect on survival. PMID- 10718542 TI - Lymphoma of the central nervous system in AIDS. AB - Central nervous system (CNS) lymphoma is a common complication of patients with HIV infection occurring in as many as 20% of patients with AIDS. This article reviews current observations on primary CNS lymphoma and systemic AIDS-related lymphoma with CNS involvement. Clinical features, diagnosis, differential diagnosis, clinical course, and therapeutic options are herein reviewed. PMID- 10718543 TI - Cerebrospinal fluid analysis for the diagnosis of human immunodeficiency virus related neurologic diseases. AB - Although early attempts demonstrated increased amounts of p24 antigen in the cerebrospinal fluid (CSF) of AIDS patients with dementia (HADC), gene amplification based assays have recently shown a correlation between the HI virus load in CSF and dementia. Although these correlations are valid for a large population, current evidence does not favor the use of CSF HI virus load measurement for the diagnosis of HADC. By polymerase chain reaction, opportunistic infections of the central (CNS) and peripheral nervous system due to cytomegalovirus, JC virus, and herpes simplex virus types 1 and 2 and primary lymphoma caused by Epstein-Barr virus can be diagnosed with an overall sensitivity ranging from 76% to 98% and a specificity between 98% and 99.5%. In contrast, bacterial, protozal, and fungal infections of the CNS are still better diagnosed by conventional assays such as culture, antigen detection, and empirical therapy. Brain biopsy still remains the ultimate means of obtaining a specific diagnosis in selected cases. PMID- 10718544 TI - Hors d'oeuvres for neurology. AB - From time to time, in the setting of lectures, rounds, or casual conversation, there is a need for hors d'oeuvres; small pieces, spices, and artifacts that generate a bit of thought and interest with a neurological twist. A potpourri of neurological trivia is herein presented for the purpose of stimulating the reader and serving as a brief reserve of questions and topics for use on rounds. PMID- 10718545 TI - The ATA, the Endocrine Society, and AACE confuse endocrinologists on thyroid disease in pregnancy. American Thyroid Association. American Association of Clinical Endocrinology. PMID- 10718546 TI - In situ hybridization and immunohistochemistry study of thyroid peroxidase expression in thyroid tumors. AB - Malignant thyroid tumors reportedly exhibit an anomaly in thyroid peroxidase (TPO) resulting in a lower affinity for monoclonal antibody 47 (mAb 47) in immunohistochemistry studies. The purpose of the present study was to compare TPO immunostaining in normal, benign, and malignant thyroid tissue with expression of mRNA sequences in four exons of the molecule including the epitope of mAb 47. TPO immunostaining was performed using mAb 47 and a polyclonal antibody (pAb). Messenger RNA expression was investigated by in situ hybridization using probes specific for mRNA sequences in exons 2, 12 (epitope of mAb), 15, and 17. As expected, pAb immunostaining was significantly positive on all benign tumors and 50% of carcinomas. With mAb 47, little or no immunostaining was observed in 16 of 17 carcinomas while significantly positive immunostaining was found in normal tissue and benign tumors. In situ hybridization showed a decrease and heterogeneity in the expression of all mRNA sequences in carcinomas as compared to normal tissue and benign tumors. Unlike the other three probes, the probe specific for exon 12 hybridized strongly with benign tumors but poorly with most carcinomas. Poor hybridization was usually correlated with defective mAb 47 immunostaining. These results confirm that TPO is expressed in thyroid carcinomas but in smaller amounts than in normal tissue and benign tumors. In malignant tumors, qualitative changes in TPO may also impede mAb 47 immunostaining. In situ hybridization showed a concomitant decrease in the corresponding TPO mRNA sequence. These changes could be due to abnormalities in the maturation of TPO mRNA leading to a different splicing variant. PMID- 10718547 TI - In situ apoptosis in the thyroid. AB - Recent evidence has emphasized the importance of programmed cell death, or apoptosis, in the maintenance of tissue homeostasis and pathogenesis of tumors. This study analyzed the significance of apoptosis in relation to the expression of p53 and bcl-2 proteins, tissue proliferation defined by Ki-67 expression, and tissue histology in thyroid tissue. Extent of apoptosis was defined by morphological criteria and the terminal deoxynucleotidyl transferase-mediated deoxy uridine triphosphate (dUTP) biotin nick end labeling (TUNEL) assay. Immunocytochemistry was performed for p53, bcl-2, and Ki-67 expression. There was good correlation between TUNEL-reactive cells and morphological evaluation criteria for apoptosis. The extent of apoptosis was significantly associated with the type of thyroid lesion (r = 0.66990, p = 0.000012), both proliferative (namely multinodular goiter) and neoplastic (benign and malignant). A higher extent of apoptosis was evident in medullary and anaplastic carcinomas. Apoptosis also correlated to p53 protein accumulation (r = 0.485, p = 0.00041) and Ki-67 immunoreactivity (r = 0.435, p = 0.001). An inverse correlation was observed between bcl-2 expression and the extent of apoptosis (r = -0.33369, p = 0.01912). A direct correlation was also observed between p53 expression and Ki-67 immunoreactivity (r = 0.623, p = 0.0002). By inhibiting apoptosis, bcl-2, may cause a shift in tissue kinetics toward the preservation of genetically aberrant cells, thereby facilitating tumor progression. These results imply that rapidly proliferating tumors appear to have a high cell turnover state in which there may be increased chance of apoptosis among the proliferating cells. The ability of apoptosis to occur in the presence of a possibly mutant p53 protein suggest the existence of at least two p53 dependent apoptotic pathways, one requiring activation of specific target genes and the other independent of it. However, keeping in mind the limited number of subjects studied in each subgroup and the rather low correlation coefficients, these possibilities would have to be substantiated in a larger study population. PMID- 10718548 TI - Excess iodine induces apoptosis in the thyroid of goitrogen-pretreated rats in vivo. AB - Previously, we observed that excess iodide rapidly suppressed the elevated ornithine decarboxylase activity in the thyroid of propylthiouracil (PTU) pretreated rats. Excess iodide also induces involution of goitrous thyroids. These findings led us to study effects of excess iodide on apoptosis of rat thyroids. When given to PTU-pretreated rats, excess potassium iodide (KI) (13 mg/kg body weight, 10 mg as iodine) induced DNA fragmentation in the thyroid at the first 3 hours after its treatment. The percentage of DNA fragmentation was also maximal at 3 hours after KI treatment. In methimazole-pretreated rats, the kinetic of DNA fragmentation was nearly the same; apoptosis increased for the first 6 hours and then decreased at 12 hours after KI administration. Other iodinated compounds such as amiodarone and diiodotyrosine have also shown apoptosis-inducing activity, but their effect was observed later than KI. Iopanoic acid had no such effect. Apoptotic changes were also observed with the use of flow cytometry. PTU or methimazole alone had some stimulatory effect on thyroid apoptosis. Iodine effect was not observed in rats treated with either perchlorate or thiocyanate. These results suggest that excess iodine induces thyroid involution in goitrogen-treated rats at least partially by apoptosis. PMID- 10718549 TI - Characterization of autonomous thyroid adenoma: metabolism, gene expression, and pathology. AB - Fifty-one in vivo characterized autonomous single adenomas have been studied for functional parameters in vitro, for gene and protein expression and for pathology, and have been systematically compared to the corresponding extratumoral quiescent tissue. The adenomas were characterized by a high level of iodide trapping that corresponds to a high level of Na+ /iodide symporter gene expression, a high thyroperoxidase mRNA and protein content, and a low H2O2 generation. This explains the iodide metabolism characteristics demonstrated before, ie, the main cause of the "hot" character of the adenomas is their increased iodide transport. The adenomas spontaneously secreted higher amounts of thyroid hormone than the quiescent tissue and in agreement with previous in vivo data, this secretion could be further enhanced by thyrotropin (TSH). Inositol uptake was also increased but there was no spontaneous increase of the generation of inositol phosphates and this metabolism could be further activated by TSH. These positive responses to TSH are in agreement with the properties of TSH stimulated thyroid cells in vitro and in vivo. They are compatible with the characteristics of mutated TSH receptors whose constitutive activation accounts for the majority of autonomous thyroid adenomas in Europe. The number of cycling cells, as evaluated by MIB-1 immunolabeling was low but increased in comparison with the corresponding quiescent tissue or normal tissue. The cycling cells are observed mainly at the periphery; there was very little apoptosis. Both findings account for the slow growth of these established adenomas. On the other hand, by thyroperoxidase immunohistochemistry, the whole lesion appeared hyperfunctional, which demonstrates a dissociation of mitogenic and functional stimulations. Thyroglobulin, TSH receptor, and E-cadherin mRNA accumulations were not modified in a consistent way, which confirms the near-constitutive expression of the corresponding genes in normal differentiated tissue. On the contrary, early immediate genes expressions (c-myc, NGF1B, egr 1, genes of the fos and jun families) were decreased. This may be explained by the proliferative heterogeneity of the lesion and the previously described short, biphasic expression of these genes when induced by mitogenic agents. All the characteristics of the autonomous adenomas can therefore be explained by the effect of the known activating mutations of genes coding for proteins of the TSH cyclic adenosine monophosphate (cAMP) cascade, all cells being functionally activated while only those at the periphery multiply. The reason of this heterogeneity is unknown. PMID- 10718551 TI - Use of thyroid ultrasound volume in calculating radioactive iodine dose in hyperthyroidism. AB - One hundred twenty-one patients treated with 131I had a thyroid ultrasound to measure thyroid volume precisely. This volume measurement was used to determine the radioactive iodine dose. The average size (+/-SEM) of the thyroid glands measured in this manner was 39.7 cm3 +/- 1.9 cc. A significant correlation was found in the estimated size of the gland by the endocrinologists and the ultrasound volume. Of the 121 patients, 89 patients had the same 131I microcurie per gram of tissue factor to determine the radioactive iodine dose. This group of patients was further evaluated in this study. The average 131I dose (+/-SEM) given was 13.2 mCi +/- 0.5 mCi. The average time until hypothyroidism was achieved 2.85 +/- 0.14 months. Ultrasound provides a safe and precise way to determine actual thyroid size when calculating 131I doses. PMID- 10718550 TI - The thyroxine-binding proteins. AB - The slow clearance, prolonged half-life, and high serum concentration of thyroxine (T4) are largely due to strong binding by the principal plasma thyroid hormone-binding proteins, thyroxine-binding globulin (TBG), transthyretin (TTR), and albumin. These proteins, which shield the hydrophobic thyroid hormones from their aqueous environment, buffer a stable free T4 concentration for cell uptake. Free rather than bound T4 is subject to homeostatic control by the hypothalamic pituitary thyroid axis. Although it is not a protease inhibitor, sequence analysis identifies TBG as a member of the serine protease inhibitor (serpin) family of proteins. Proteolytic cleavage of TBG appears to be a mechanism for site-specific release of T4 independently of homeostatic control. TBG probably facilitates the transport of maternal T4 and iodide to the fetus, although this remains to be proven. High-affinity cellular binding sites for TTR have been described; however, their function and that of choroid plexus synthesis of TTR and transport of T4 into the cerebrospinal fluid remain unclear. Albumin, with the lowest T4 affinity and fastest T4 release of the major T4-binding proteins may promote quick exchange of T4 with tissue sites. The affinity of albumin for T4 is increased by histidine substitution for arginine 218 in the most common form of dysalbuminemic hyperthyroxinemia. However, proline and alanine substitutions at the same site have a similar effect, suggesting that arginine 218 interferes with T4 binding. PMID- 10718552 TI - Criticizing evidence-based medicine. PMID- 10718553 TI - Increased lead excretion correlates with desoxypyridinoline crosslinks in hyperthyroid patients. AB - Lead is a widespread toxic metal that accumulates predominantly in human bone. Altered bone metabolism in hyperthyroidism is characterized mainly by bone resorption. Thus, we speculated that lead excretion could be increased in hyperthyroid patients. In 12 hyperthyroid patients (43.3 +/- 16.1 years) who were not previously occupationally exposed to lead, lead concentrations in blood (PbB), spot urine samples corrected by urine creatinine (PbUs), and in 24-hour urine samples (PbU24) were determined in the hyperthyroid state and after euthyroidism had been induced by therapy. Serum osteocalcin (OC) and desoxypyridinoline crosslinks (Pyr) served as specific markers for bone metabolism. After induction of euthyroidism (duration of antithyroid therapy: mean 17.3 +/- 6.9 weeks) PbB was reduced (3.7 +/- 2.6 vs. 5.7 +/- 4.7 microg/dL, p = 0.041) as was PbUs (0.39 +/-0.27 vs. 0.61 +/- 0.32 microg/mg Cr, p = 0.005). A fourfold decrease of PbU24 was associated with a 3.3-fold decrease of Pyr, and moreover there was a significant correlation between Pyr and PbUs (r = 0.58, p = 0.047). Concentration of total triiodothyronine correlated with Pyr (r = 0.66, p = 0.018), but not with PbB or PbUs. OC showed only a tendency to be increased before antithyroid medication, and did not correlate with either thyroid hormone or Pyr. Our results indicate that in hyperthyroid patients, even when not previously exposed to lead, lead excretion is increased due to bone resorption. PMID- 10718554 TI - Serum thyroglobulin levels after thyroxine withdrawal in patients with low risk papillary thyroid carcinoma. AB - We hypothesized that elevated levels of serum thyroglobulin (Tg) are frequently found as the only index of residual neoplasm in patients with low-risk papillary thyroid carcinoma. The records of patients operated on for papillary thyroid carcinoma over a 2-year period were reviewed, and the patients were allocated to risk groups by a validated staging method that does not include Tg levels. Of the 35 patients who manifested a low-risk carcinoma, 9 (26%) exhibited elevated Tg concentrations (11-53 ng/mL) during thyroxine withdrawal after therapies, while clinical, scintigraphic, and radiographic studies at least 1 year later showed no evidence of tumor. Prior scintigraphic imaging of therapeutic doses of 131I in 8 of 9 patients demonstrated no distant metastases, further confirming the low-risk status of this group. The staging method predicts that only 0.9% of patients with low-risk papillary carcinoma will have a cause specific death in 20 years. Elevated Tg concentrations have not been shown to forecast independently the survival of patients with low-risk papillary carcinoma. Thus, although frequently encountered, elevated Tg concentrations are unlikely to predict shortened survival in patients with papillary carcinoma for whom low risk has been determined from other data. PMID- 10718555 TI - Recurrent differentiated thyroid cancer without elevation of serum thyroglobulin. AB - Thyroglobulin (Tg) is a reliable tumor marker in patients with well differentiated thyroid cancer (WDTC). We identified 11 patients who had undetectable serum Tg and no thyroglobulin antibody (TgAb) in the presence of clinical disease. Three had residual disease after ablation of the thyroid by surgery plus radioiodine and 8 relapsed after a disease-free interval. Histologic review confirmed that 7 of the tumors were papillary carcinomas and 4 were follicular carcinomas. Immunohistochemical staining for Tg was positive in 6 of 7 papillary and in 3 of 4 follicular carcinomas. There were no identifiable histologic or clinical features that could be used to predict further patients who may relapse with absence of this serum marker. Negative serum Tg did not appear to be an adverse prognostic feature. During follow-up, measurement of Tg and TgAb should be supplemented by radioiodine scanning and radiological imaging in patients in whom recurrence is likely or suspected. PMID- 10718556 TI - Inhibition of metabolic activity in papillary thyroid carcinoma by a somatostatin analogue. AB - Two patients with widely metastatic papillary thyroid cancer demonstrated progressive growth of diffuse pulmonary lesions. One patient had no apparent response to high doses of 131I and the other hand no 131I uptake. 111In pentetreotide scans revealed that many of the metastatic lesions expressed somatostatin receptors. The baseline metabolic activity and three-dimensional volume of the lesions were determined by 18F-fluoro-de-oxyglucose positron emission tomography (FDG-PET). After 3 or 4 months of octreotide (Sandostatin LAR Depot; Novartis Pharmaceutical, East Hanover, NJ) therapy, repeat FDG-PET scans revealed reductions in tumor volume and decreases in the standard uptake values of FDG. We conclude that octreotide therapy can change the biological activity of metastatic thyroid cancer lesions that exhibit somatostatin receptors. PMID- 10718557 TI - Tall cell papillary carcinoma of the thyroid: metastatic to the pancreas. AB - We present the case of a 53-year-old man with tall cell variant of papillary thyroid carcinoma that metastasized to the pancreas. The pancreas is a unique site of metastasis for differentiated thyroid cancer. The lesion in the neck and lung concentrated radioiodine, whereas the intrapancreatic lesions did not. Discordance between radioiodine studies and serum thyroglobulin in follow-up of patients with differentiated tall cell is reviewed and discussed. PMID- 10718561 TI - The British Olympic Association's position statement on athlete confidentiality. PMID- 10718560 TI - Clear fluid from a thyroid cyst. PMID- 10718558 TI - Orbital metastasis as primary manifestation of thyroid carcinoma. AB - A 59-year-old woman with unknown primary tumor developed progressive painless left upper eyelid swelling and exophthalmos. Computed tomography (CT) showed a well-circumscribed left orbital mass producing bone lysis. Immunohistologic staining of the incisional biopsy specimen was positive for thyroglobulin, suggesting an orbital metastasis from thyroid carcinoma. At this time, thyroglobulin was high (1400 ng/dL). Total thyroidectomy with lymph node dissection disclosed a follicular carcinoma with microscopic foci of papillary variant follicular carcinoma. Two months after radioiodine treatment, the CT showed a regression of the orbital tumor mass with concomitant decrease in thyroglobulin (428 ng/dL). Although orbital metastases of thyroid carcinoma are uncommon, thyroid carcinoma has to be considered as a potential primary tumor in a patient with an orbital metastasis. PMID- 10718562 TI - Fluid ingestion and exercise hyperthermia: implications for performance, thermoregulation, metabolism and the development of fatigue. AB - The development of fatigue during exercise and the subsequent onset of exhaustion occur earlier in the heat than in cooler environments. The underlying mechanisms responsible for the premature development of fatigue in the heat have yet to be clearly identified. However, the proposed mechanisms include metabolic, cardiovascular and central nervous system perturbations, together with an elevated core temperature. Fluid ingestion is one of three strategies that have been shown to be successful in enhancing the performance of endurance exercise in the heat, with the other interventions being precooling and acclimatization. However, like the development of fatigue in the heat, the mechanisms by which fluid ingestion allows for improved exercise performance remain unclear. We propose that fluid ingestion enhances exercise performance in the heat by increasing the heat storage capacity of the body. We suggest that the thermoregulatory, metabolic and cardiovascular alterations that occur as a result of this increased heat storage capacity contribute to performance enhancement when fluid is ingested during exercise heat stress. PMID- 10718559 TI - Is there a "difference" in thyroglobulin testing. PMID- 10718563 TI - Acute effects of intense interval training on running mechanics. AB - The aims of this study were to determine if there are significant kinematic changes in running pattern after intense interval workouts, whether duration of recovery affects running kinematics, and whether changes in running economy are related to changes in running kinematics. Seven highly trained male endurance runners (VO2max = 72.3+/-3.3 ml x kg(-1) x min(-1); mean +/- s) performed three interval running workouts of 10 x 400 m at a speed of 5.94+/-0.19 m x s(-1) (356+/-11.2 m x min(-1)) with a minimum of 4 days recovery between runs. Recovery of 60, 120 or 180 s between each 400 m repetition was assigned at random. Before and after each workout, running economy and several kinematic variables were measured at speeds of 3.33 and 4.47 m x s(-1) (200 and 268 m x min(-1)). Speed was found to have a significant effect on shank angle, knee velocity and stride length (P < 0.05). Correlations between changes pre- and post-test for VO2 (ml x kg(-1) x min(-1)) and several kinematic variables were not significant (P > 0.05) at both speeds. In general, duration of recovery was not found to adversely affect running economy or the kinematic variables assessed, possibly because of intra-individual adaptations to fatigue. PMID- 10718566 TI - Correlates of simulated hill climb cycling performance. AB - The aim of this study was to assess the relationship between several commonly used aerobic and anaerobic cycle ergometer tests and performance during a treadmill cycling hill climb. Eight competitive cyclists (age 27+/-7 years; body mass 73.2+/-5.2 kg; height 177+/-6 cm; mean +/- s) completed six tests in random order: a lactate minimum test; a Wingate anaerobic power test; and two 6-km climbs at 6% and two 1-km climbs at 12% gradient performed on a motorized treadmill. The mean times and power outputs for the 6-km and 1-km climbs were 16:30+/-1:08 min: s and 330+/-17.8 W, and 4:19+/-0:27 min: s and 411+/-24.4 W, respectively. The best individual predictor of 6-km and 1-km performance times was the time for the corresponding climb at the other distance (r = 0.97). The next strongest predictor of both hill climb performances was the average power produced during the Wingate test divided by body mass. Stepwise regression analysis showed that the two variables contributing most to the prediction equation for both climbs were the Wingate average power per unit of body mass and maximal aerobic power divided by total mass (rider + bike), which together accounted for 92 and 96% of the variability in the 6-km and 1-km climbs. In conclusion, among competitive cyclists, the Wingate average power per unit of body mass was the best single predictor of simulated cycling hill climb performance at the distance and gradient used. PMID- 10718567 TI - From novice to no know-how: a longitudinal study of implicit motor learning. AB - The aim of this study was to ascertain whether the performances of implicit and explicit learners would converge over an extended period of learning. Participants practised a complex motor skill--golf putting--for 3000 trials, either with a concurrent secondary, tone-counting task (implicit learning) or without such a task (explicit learning). The cognitive demands of the secondary task were predicted to prevent the accumulation of verbalizable rules about the motor task. The implicit group reported significantly fewer rules than the explicit group on subsequent verbal protocols. The performance of the implicit group remained below that of the explicit group throughout the learning phase. However, no significant differences were found between groups during a delayed retention test. Additionally, for the participants in the explicit group only, a Reinvestment Scale score correlated positively with the number of rules accrued and negatively with overall putting performance during the learning phase. We use the results to argue against the excessive use of verbal instruction during skill acquisition, which might be unnecessary and ultimately might hamper performance under stressful conditions. PMID- 10718564 TI - The influence of environmental factors on rugby football injuries. AB - The aim of this study was to establish the influence of weather and pitch conditions on the frequency and nature of rugby injuries. Observers at 26 senior rugby clubs in the Borders District of the Scottish Rugby Union reported all injuries to 1169 (96%) registered players at Saturday home and away matches during the 1993-94 season (August 1993 to April 1994). Weather and pitch conditions at 112 grounds were recorded on 605 occasions; 1268 Borders teams played at these grounds, with 344 injury episodes being sustained. Matches were played in dry weather for three-quarters of these occasions. There was a moderate association between weather and the state of the pitch (Spearman's rank correlation, r = -0.46, P < 0.001), with heavier pitches occurring in wetter weather. Logistic regression revealed that there were significant month-of-season (P = 0.003), wind strength (P = 0.008) and temperature (P = 0.011) effects on the risk of injury. On four of five occasions when matches were played in a downpour of rain, at least one injury episode occurred. Our results show that the month of the season and the weather may influence the occurrence of rugby injuries, but that the state of the pitch does not. Further studies are required to investigate these factors in more detail. PMID- 10718565 TI - The Loughborough Intermittent Shuttle Test: a field test that simulates the activity pattern of soccer. AB - The aims of this study were to describe and determine the test-retest reliability of an exercise protocol, the Loughborough Intermittent Shuttle Test (the LIST), which was designed to simulate the activity pattern characteristic of the game of soccer. The protocol consisted of two parts: Part A comprised a fixed period of variable-intensity shuttle running over 20 m; Part B consisted of continuous running, alternating every 20 m between 55% and 95% VO2max, until volitional fatigue. Seven trained games players (age 21.5+/-0.9 years, height 182+/-2 cm, body mass 80.1+/-3.6 kg, VO2max 59.0+/-1.9 ml x kg(-1) x min(-1); mean +/- s(x)) performed the test on two occasions (Trial 1 and Trial 2), at least 7 days apart, to determine the test-retest reliability of the sprint times and running capacity. The physiological and metabolic responses on both occasions were also monitored. The participants ingested water ad libitum during the first trial, and were then prescribed the same amount of water during the second trial. The 15 m sprint times during Trials 1 and 2 averaged 2.42+/-0.04 s and 2.43+/-0.04 s, respectively. Run time during Part B was 6.3+/-2.0 min for Trial 1 and 6.1+/-1.3 min for Trial 2. The 95% limits of agreement for sprint times and run times during Part B were -0.14 to 0.12 s and -3.19 to 2.16 min respectively. There were no differences between trials for heart rate, rating of perceived exertion, body mass change during exercise, or blood lactate and glucose concentrations during the test. Thus, we conclude that the sprint times and the Part B run times were reproducible within the limits previously stated. In addition, the activity pattern and the physiological and metabolic responses closely simulated the match demands of soccer. PMID- 10718568 TI - Coping with pre- and in-event fluctuations in competitive state anxiety: a longitudinal approach. AB - The direction of anxiety--the positive or negative nature of competition-related cognitions--is an important distinguishing variable that accounts for significant individual differences. However, very little is known about the pre- and in-event patterning of the construct, specifically how it changes over time. The aims of this study were to establish the extent to which the direction of anxiety is a dynamic or stable longitudinal response to stress and whether this patterning appears to be related to coping. Pre-event, state anxiety intensity and direction data were obtained from 22 youth sport participants in two training and two competition environments within the same season, with the in-event anxiety and coping data being obtained from the 'high-stress' competition condition. The predispositional coping strategies of the participants were assessed by use of the trait version of the 'COPE' scale. The findings revealed significant differences in the patterning of anxiety direction between and within the facilitative and debilitative groups. This was accompanied by distinct qualitatively and quantitatively reported differences in the strategies of coping adopted by each group. Facilitators appeared to use problem- and emotion-focused coping characteristics in response to stress, whereas debilitators appeared limited in their use of coping constructs. Conceptually, the direction of anxiety would appear to be a mechanism that may, in itself, exist as a strong indicator that effective coping is taking place. PMID- 10718569 TI - The British Olympic Association's position statement on athlete confidentiality. PMID- 10718570 TI - Duplicate sampling reproducibility of atmospheric residues of herbicides for paired pan and high-volume air samplers. AB - The reproducibility of collection of atmospheric residues of the herbicides 2,4-D and triallate as bulk (wet plus dry) deposition samples by paired pan samplers and as particulate (filter) and vapour (PUF/XAD-2 resin cartridge) samples by paired high-volume air samplers was determined. Variability of herbicide concentrations in paired bulk deposition samples was within 25% for 65 and 80% of the samples for 2,4-D and triallate, respectively, with approximately 90% of the paired samples being within a factor of 2 for both herbicides. The vapour samples of 2,4-D and triallate showed similar reproducibilities. The highest reproducibility was observed for the filter samples with 92% of the paired data sets for 2,4-D being within 25% variability. No triallate was detected in the filter samples. PMID- 10718571 TI - Bioaccumulation of polychlorinated biphenyls in ranid frogs and northern water snakes from a hazardous waste site and a contaminated watershed. AB - Livers of bullfrogs (Rana catesbeiana) from a polychlorinated biphenyl (PCB) contaminated watershed and hazardous waste site located in Pickens County, South Carolina, contained significantly higher concentrations of PCBs (2.33 and 2.26 ppm, respectively) than those from a reference site (0.05 ppm). Green frogs (R. clamitans) from the two contaminated sites also accumulated higher levels of PCBs (2.37 and 3.88 ppm, respectively) than those from the reference site (0.02 ppm). No temporal variation was observed in PCB concentrations of bullfrogs or green frogs from the contaminated sites between 1992 and 1993. Levels of PCBs in the livers of northern water snakes (Nerodia sipedon) were significantly higher in snakes from the contaminated watershed (13.70 ppm) than in those from the waste site (2.29 ppm) and two reference sites (2.50 and 1.23 ppm). When compared to frogs, significantly higher bioaccumulation occurred in water snakes from the contaminated watershed. No significant differences in PCB levels were found with respect to sex or body size (snout-vent length (SVL) or body mass) for frogs or snakes. PCBs were detected also in eggs of both frogs and snakes. Results of this study provide baseline data and document the bioaccumulation of PCB residues in frog and snake tissues; however, the significance of these tissue residues to reproduction, survival, growth/development, and population dynamics in contaminated habitats is unknown. PMID- 10718572 TI - Selective removal of organic contaminants from sediments: a methodology for toxicity identification evaluations (TIEs). AB - Aqueous slurries of a test sediment spiked with dibenz[a,h]anthracene, 2,4,5,2',4',5'-hexachlorobiphenyl, p,p'-DDE, or phenanthrene were subjected to decontamination experimentation. The spiked sediments were agitated at elevated temperatures for at least 96 h in the presence of either of the two contaminant absorbing media: clusters of polyethylene membrane or lipid-containing semipermeable membrane devices (SPMDs). The effects of treatment temperature and surface area of media on the removal of contaminants were explored. This work is part of a larger methodology for whole-sediment toxicity identification evaluation (TIE). A method is being sought that is capable of detoxifying sediments with respect to organic contaminants while leaving toxicity attributable to inorganic contaminants unaffected. PMID- 10718573 TI - Bioassays with Vibrio fischeri for the assessment of delayed toxicity. AB - The standardized bioluminescence assay with Vibrio fischeri underestimates the aquatic toxicity of chemicals which interfere with metabolic pathways supporting long term processes like growth and reproduction due to its short incubation time (30 min). Therefore this short term assay was compared with two alternative bioassays with prolonged incubation times using the same test organism: the growth inhibition assay (7 h) and the long term bioluminescence assay (24 h). Two sets of compounds were selected to reflect acute and delayed toxicity. The first group comprised pentachlorophenol, dodecylpyridiniumbromide and 3,4 dichloroaniline and the second nalidixic acid, chloramphenicol and streptomycinsulfate. The effects of compounds with acute toxicity are determined with similar sensitivity in all bioassays. Substances with delayed toxicity show only minor or no toxicities in the standardized short term bioassay but severe effects in both long term bioassays independent of the parameter used. It is concluded that the standardized short term bioluminescence assay exhibits serious limitations for the assessment of aquatic toxicity. The long term bioassays, however, may help to overcome these limitations. PMID- 10718574 TI - Urease inhibition: a tool for toxicity identification in sediment elutriates. AB - A method for the detection and confirmation of heavy metal toxicants in sediment elutriates based on a urease inhibition assay, ICP-AES analysis and EDTA chelation in the frame of toxicity identification evaluation (TIE) is presented. Zinc was identified as the major toxicant in pHstat elutriates of sediments of the river Saale (Germany). Implications of natural and anthropogenic chelating agents, which are frequently present in environmental samples, on toxicity confirmation of heavy metals based on the toxic unit approach are discussed. PMID- 10718575 TI - Surfactant (Genapol OX-80) toxicity to Selenastrum capricornutum. AB - A non-ionic surfactant (Genapol OX-80) was proposed as a means of reducing destructive crayfish activity in rice. In order to study the toxicity of this product on algae, a test in accordance to the ISO 8692 protocol (1989) was performed. The growth inhibiting effect of the pure formulation of Genapol OX-80 was studied on Selenastrum capricornutum in the concentration range of 0.01-1.0 mg/l. The result indicated a 72 h EC50-value of 0.5 mg/l. The suggested field concentration of approximately 50 mg/l is therefore two orders of magnitude above the algal EC50-value. From our findings it is expected that the impact on the rice field algae will not be negligible. PMID- 10718576 TI - Relative lipid content as the sole mechanistic determinant of the adipose tissue:blood partition coefficients of highly lipophilic organic chemicals. AB - The adipose tissue:blood partition coefficient (PCat:b) refers to the ratio of chemical concentration or solubility in adipose tissue and blood. The solubility of a chemical in adipose tissue or whole blood is equal to the sum total of its solubility in lipid and water fractions of these matrices. For highly lipophilic organic chemicals (HLOCs, i.e., chemicals with log n-octanol:water partition coefficients (PCo:w) greater than four), their solubility in the water fractions of both tissue and blood is negligible, and therefore their solubility in lipid fractions of tissue and blood alone determines PCat:b. Since the numerical value representing chemical solubility in lipids is likely to be the same for both blood lipids and adipose tissue lipids, the PCat:b values should be hypothetically, equal to the ratio of lipid content of adipose tissue and blood. The objective of the present study was therefore to verify whether the PCat:bs of HLOCs (volatile organics, dioxins, PCBs, PBBs, DDT) are equal to the ratio of adipose tissue and blood lipid levels. The data on lipid content of rat and human blood and adipose tissues were obtained from the literature. The calculated tissue:blood lipid ratios were comparable to the human and rat PCat:b of volatile organic chemicals, dioxins, PCBs, PBBs and/or DDT obtained from the literature. These results then suggest that, regardless of the identity and PCo:w of HLOCs, their PCat:b is equal to the ratio of lipid in adipose tissues and blood. PMID- 10718577 TI - Microcalorimetric studies on the metabolism of Chlorella vulgaris. AB - The heat output of the non-growth metabolism of Chlorella vulgaris has been determined using an LKB-2277 BioActivity Monitor. The experimental results indicate that there is a turn-point on the metabolic thermogenic curves, which can be divided into two parts, an early phase and a later phase. For the early phase, there is a linear relationship between the metabolic power (P) and the cell concentration (C), and that the heat output produced by a single cell's metabolism (P0) depends on the cell concentration (C) and is inhibited by the cell density, the thermokinetic equation of their metabolism is dP/dt = k0, k0 = 0. PMID- 10718579 TI - Effects of chromium and nickel on germination and growth in tolerant and non tolerant populations of Echinochloa colona (L.) Link. AB - The tolerance of populations of a grass, Echinochloa colona, growing abundantly on chromite minewaste dumps, was tested in two separate experiments. Seed-based experiments indicate that the populations growing naturally on uncontaminated sites, germinated better in nutrient solutions without metal than those collected from minewaste dumps. Metal tolerance indices were greater in the plant populations derived from metal contaminated sites and better growth of these plants was noted on mine spoil soil-mix in the ratio of 1:1; the percentage of seed germination and the rate of seedling growth, however, declined in a soil compost containing 25% mine spoil and 75% uncontaminated (control) soil. Populations of Echinochloa colona occurring naturally on chromite mine spoils, therefore, appear to have developed metal tolerance. It is maintained by a balanced and stable genetic system built up and adjusted by natural selection. Such material is very suitable to be used in restoration work designed to produce an effective vegetation cover to improve the derelict land and to reduce erosion. This finding might be useful in revegetation programmes on metalliferous minewastes. PMID- 10718578 TI - Microcalorimetric studies of the toxic action of La3+ in mitochondria isolated from Star-cross 288 chicken heart tissue cells. AB - The thermogenic curves of the metabolism of mitochondria isolated from the heart of chicken Star-cross 288 and the effect of La3+ on it were studied by using an LKB-2277 BioActivity Monitor, ampoule method, at 37 degrees C. After isolation from the chicken heart tissue, mitochondria still have metabolic activity and can live for a long time by using the stored nutrients. From the thermogenic curves, we obtained the thermokinetic equations under different conditions. The kinetics show that La3+ has changed the metabolism completely. PMID- 10718580 TI - Biomarker responses at different levels of biological organisation in crabs (Carcinus aestuarii) experimentally exposed to benzo(alpha)pyrene. AB - The aim of this study was to validate a multi-trial biomarker approach for the evaluation of toxicological risk due to benzo(alpha)pyrene. Carcinus aestuarii, exposed to increasing concentrations of B(alpha)P in the water, was used as the bioindicator organism. A set of biomarkers were tested in order to: identify biological materials for biomarker and residue analysis; determine a group of sensitive techniques for the assessment of PAH contamination; investigate correlation between responses at different levels of biological organisation. The results underlined that BPMO activities in hepatopancreas and gills were a good biomarker of exposure to PAH-type compounds. B esterases activities in hemolymph and porphyrin patterns in excreta could be proposed as a non-destructive approach for evaluating chemical exposure in this species. PMID- 10718581 TI - Classifying environmental pollutants: Part 3. External validation of the classification system. AB - In order to validate a classification system for the prediction of the toxic effect concentrations of organic environmental pollutants to fish, all available fish acute toxicity data were retrieved from the ECETOC database, a database of quality-evaluated aquatic toxicity measurements created and maintained by the European Centre for the Ecotoxicology and Toxicology of Chemicals. The individual chemicals for which these data were available were classified according to the rulebase under consideration and predictions of effect concentrations or ranges of possible effect concentrations were generated. These predictions were compared to the actual toxicity data retrieved from the database. The results of this comparison show that generally, the classification system provides adequate predictions of either the aquatic toxicity (class 1) or the possible range of toxicity (other classes) of organic compounds. A slight underestimation of effect concentrations occurs for some highly water soluble, reactive chemicals with low log K(ow) values. On the other end of the scale, some compounds that are classified as belonging to a relatively toxic class appear to belong to the so called baseline toxicity compounds. For some of these, additional classification rules are proposed. Furthermore, some groups of compounds cannot be classified, although they should be amenable to predictions. For these compounds additional research as to class membership and associated prediction rules is proposed. PMID- 10718582 TI - A risk assessment of selected phthalate esters in North American and Western European surface waters. AB - Potential risks to aquatic organisms by four commercial phthalate esters, dimethyl (DMP), diethyl (DEP), di-n-butyl (DBP), and butylbenzyl (BBP), were assessed using measured and calculated concentrations in North American and Western European surface waters. Predicted no effect concentrations (PNECs) were calculated using statistical extrapolation procedures and the large aquatic toxicity database. Surface water concentrations of DMP, DEP, DBP, and BBP were calculated using reported emissions to US surface waters from the toxics release inventory (TRI). Monitoring data obtained from the US EPA STORET database and literature surveys from North America and Western Europe show that DMP, DEP, DBP, and BBP are infrequently detected in surface water. Calculated and measured concentrations of DMP, DEP, DBP, and BBP are typically several orders of magnitude below their respective PNECs, indicating that these phthalate esters do not pose a ubiquitous threat to aquatic organisms in North American and Western European surface waters. PMID- 10718583 TI - Identification of the dimethylbenzyl mercapturic acid in urine of rats administered with 1,2,4-trimethylbenzene. AB - A study was undertaken of the mercapturic acid metabolism of 1,2,4 trimethylbenzene in the rat. Of three regioisomeric dimethylbenzyl mercapturic acids, i.e. 2,4-, 2,5- and 3,4-dimethylbenzyl isomers, the third isomer was not found in the urinary mercapturic acid isolated by preparative HPLC, from the comparison of NMR spectrum of the isolate with those of authentic specimens. The urinary mercapturate was then assigned to 2,4- and/or 2,5-dimethylbenzyl isomers. The excretion rate of the mercapturic acid was 14-20% of dose as 2,4 dimethylbenzyl isomer. PMID- 10718584 TI - Aquatic toxicity of two Corexit dispersants. AB - The oil spill dispersants, Corexit 9500 and Corexit 9527 have low to moderate toxicity to most aquatic species in laboratory tests. Toxicity estimates are significantly affected by test variables such as species, lifestage, exposure duration, and temperature. Aquatic toxicity data generated from spiked, declining exposures (107 min half-life) are more reflective of actual dispersant use conditions. Decisions to use oil spill response chemicals should not be based solely on aquatic toxicity. Factors to consider include product effectiveness, toxicity of dispersed oil, species/habitats requiring priority protection, and recovery potential of sensitive habitats and populations. An environmental risk assessment approach is recommended where dispersant toxicity data generated under environmentally relevant exposures are compared to estimated environmental concentrations of dispersants. PMID- 10718585 TI - Geothermal power plants at Mt. Amiata (Tuscany-Italy): mercury and hydrogen sulphide deposition revealed by vegetation. AB - At Mt. Amiata (Italy) geothermal energy is used, since 1969, to generate electricity in five plants with a nominal capacity of 88 MW. Anomalous levels of mercury characterise geothermal fluids of Mt. Amiata, an area renowned for its vast cinnabar deposits and for the mercury production carried out in the past. Mercury emission rates range from 300 to 400 g/h, or 3-4 g/h per MW electrical installed capacity. These emissions are coupled with a release of 7-8 kg/(h MW) of hydrogen sulphide (H2S). Mercury is discharged as Hg0 gaseous species and reaches the atmosphere with the non-condensable gas fraction. In this fraction, CO, is the major component (94-98%), H2S is around 1% and mercury concentration is as high as 1-10 mg/Nm3. Leaves of a spontaneous grass (Avena sterilis), at the end of the vegetative cycle, were used as mercury bioconcentrators to map deposition near geothermal power plants and to calculate the corresponding average levels of Hg0 in the air. Direct measurements of mercury and hydrogen sulphide vapours in the air reached by power plant emissions showed a ratio of about 1-2000. This ratio was applied to calculate average levels of hydrogen sulphide starting from mercury deposition mapping: typical concentrations of mercury and hydrogen sulphide were of the order of 10-20 ng/m3 and 20-40 microg/m3, respectively. PMID- 10718586 TI - Transfer of responsibilities to the FSA. Food Standards Agency. PMID- 10718587 TI - MOD guidance on livestock claims. PMID- 10718588 TI - Hunting inquiry seeks factual evidence. PMID- 10718589 TI - Production factors that influence the hygienic condition of finished beef cattle. AB - A survey was conducted at five UK abattoirs to trace the source of dirty beef cattle and identify factors in the production chain that contributed to their dirtiness. The Meat Hygiene Service Clean Livestock score categories were used and the animals' histories were traced back to their farm of origin. Comprehensive information was collected relating to the farm, transport and lairage phases for 675 cattle from 85 batches. The mean score of the animals on arrival at the abattoir was 1.57, with 2.8 per cent in categories 3 and 4, and none in category 5. Regression analysis, blocking the data by farm of origin, revealed that age, feed type, coat length, clipping, journey distance and time, and abattoir were the six principal factors that affected the scores. Cattle under 20 months of age were cleaner than older cattle. Dry diets resulted in lower scores than wet diets. 'Shorthair' cattle were cleaner than 'medium' or 'longhair' cattle, and cattle which had been clipped were cleaner than unclipped animals. The cattle which had travelled over 150 miles (15 per cent) were dirtier than the others. Within each abattoir there were variations in score between 1 and 4, and the mean scores of the five abattoirs ranged from 1.19 to 1.76. A significant part of this variation could only be accounted for by unmeasured variables such as abattoir management practice, assessors' categorisations and climate. PMID- 10718590 TI - Ultrasonographic diagnosis of pericardial effusion and atrial dilation in a spur thighed tortoise (Testudo graeca). AB - Ultrasonography was used to diagnose pericardial effusion, atrial dilatation and liver masses in a spur-thighed tortoise which was more than 80 years old and suffering from posthibernation anorexia, lethargy, oedema and pneumonia. The tortoise was treated twice with frusemide and ceftazidime for the pneumonia, resulting each time in a temporary remission for about a month. After a further recurrence, the animal was euthanased and the lesions predicted by ultrasound were confirmed postmortem. It is suggested that ultrasound may be useful for the differentiation of cardiac problems from other causes of posthibernation lethargy in the tortoise. PMID- 10718591 TI - Detection of granulocytic Ehrlichia species DNA by PCR in persistently infected dogs. AB - Three female beagle dogs inoculated with granulocytic Ehrlichia species were monitored for four to six months to determine whether there was evidence that the organisms persisted. The dogs were inoculated intravenously with blood containing an Ehrlichia species closely related to Ehrlichia equi and Ehrlichia phagocytophila, and identical to the human granulocytic ehrlichiosis agent with respect to its 16S rRNA gene sequence. The clinical signs were evaluated, and blood samples were collected for haematology, serum biochemistry and serology. Ehrlichial inclusions in the blood were monitored by microscopy, and ehrlichial DNA was detected by the polymerase chain reaction (PCR). Two of the dogs were injected with prednisolone on days 54 to 56 and days 152 to 154 after infection, and the other was injected with prednisolone on days 95 to 97 after infection. The dogs were euthanased and examined postmortem. Ehrlichial inclusions were demonstrated in the neutrophils and seroconversion occurred shortly after inoculation. Two of the dogs developed acute disease with rectal temperatures above 39.0 degrees C, after which no further clinical signs were observed. The administration of corticosteroids seemed to facilitate the detection of ehrlichial inclusions. Ehrlichial DNA was detected intermittently by PCR in blood samples from two of the dogs throughout the study. Persistent infection was demonstrated up to five-and-a-half months after inoculation. PMID- 10718592 TI - Avian infectious bronchitis virus: isolation of an apparently new variant in Italy. PMID- 10718593 TI - Novel localised variant of canine epidermolysis bullosa acquisita. PMID- 10718594 TI - Control of paratuberculosis (Johne's disease) in goats by vaccination of adult animals. PMID- 10718595 TI - Student debt and socioeconomic mix. PMID- 10718596 TI - Deaths in captive penguins. PMID- 10718597 TI - Triclabendazole-resistant Fasciola hepatica in southwest Wales. PMID- 10718598 TI - Chronic uveitis in cats. PMID- 10718599 TI - The therapeutic alliance. Introduction. PMID- 10718600 TI - The therapeutic relationship: from transference to alliance. AB - A variety of conceptualizations of the relationship between therapist and client, as well as the impact the quality of this relationship has on the client, are reviewed. Different hypotheses about bow the alliance might influence therapeutic progress are also presented. The review pays attention to both the historical and conceptual dimensions of this issue, and provides a summary of the empirical bases of the claims made of the value of a good alliance. The article concludes with a discussion of an integrated model of the alliance and a discussion of the clinical and training implications of our current understanding of this concept. PMID- 10718601 TI - Alliance ruptures and repairs in experiential therapy. AB - The formation and maintenance of a positive working alliance is fundamental to the success of experiential therapy. Experiential therapists face a special challenge insofar as they must constantly consider whether to direct the therapeutic process more actively or remain more closely within the clients' own frame of reference. This requires that experiential therapists be acutely aware of the alliance with their clients on a moment-to-moment basis during the session. In this article ruptures to the alliance, comprising breakdowns In the agreement between clients and therapists as to the goals and tasks of therapy during the early and middle phases of treatment, are identified and explored. As ruptures are often covert processes, methods of detecting their occurrence during the session are presented. Finally, experiential techniques for forging, maintaining, and repairing the alliance between clients and therapists are discussed and illustrated. PMID- 10718602 TI - Two examples of strains in the therapeutic alliance in an integrative cognitive therapy. AB - Two case examples are presented to illustrate methods of recognizing and working with strains in the therapeutic alliance in an integrative cognitive therapy. I argue that developmental, personality, and therapy-readiness constructs are useful in understanding strains in the alliance and whether to confront such strains directly. Attachment theory was relevant to understanding the therapeutic alliance in the first case, when the client reacted to the possibility of termination in an insecure manner. Direct discussion of the alliance strain was useful in this case. The client in the second case was resistant to interpersonal influence. I did not address strains directly because the client would probably not have seen this work as relevant to his goals. In this case, a focus on strains could have weakened our alliance rather than strengthened it. PMID- 10718603 TI - Troubles in the therapeutic relationship: a pluralistic perspective. AB - From a pluralist perspective, there is no universally correct therapeutic attitude and no set way to deal with troubled therapeutic relationships. A three factor model of therapeutic impasse is presented in terms of hopeless narratives, failing strategies, and ineffective interactions. This triple mapping allows for the development of a working hypothesis and a critical intervention in order to set therapy back on track. Although a one-shot event, the critical intervention is not a single-session magical cure, but an attempt to restore a productive therapeutic routine based on a better working alliance. This article illustrates this approach in three separate cases. PMID- 10718604 TI - The therapeutic alliance in couples and family therapy. AB - The therapeutic alliance is central to couples and family therapy. Although the formal concept of therapeutic alliance has not been used widely within the family therapy field, virtually every prominent clinical theorist has addressed the importance of establishing and maintaining a positive therapeutic relationship with the family. The alliance in couples and family treatment differs from the alliance in individual psychotherapy in that the couples and family therapist must establish and maintain multiple alliances. They also must adopt a conceptual framework that accounts for the interactions within triangles or three-person systems, recognize the influence of the system operating on him or her, and appreciate how different models of family therapy define the position of the therapist in relation to the couple or family. An integrative review of the therapeutic alliance in couples and family therapy is followed by a discussion of problems in the therapeutic relationship. Two general clinical strategies for managing difficulties in the alliance then are illustrated through case vignettes. PMID- 10718605 TI - Impasses in the psychoanalytic relationship. AB - In this article, I outline a psychoanalytic perspective on therapeutic impasses based on more recent developments in psychoanalytic therapy and on Winnicott's (1971) work in particular. This perspective suggests that many impasses result from clients' inability to participate in their own experience in a subjectively meaningful and affectively vital way. I argue that, in such cases, the therapeutic task becomes one of helping clients relate to events in a more personal and affectively engaged fashion through a type of therapeutic play in which the boundary between what is real and unreal becomes temporarily ambiguous. PMID- 10718606 TI - Resolving therapeutic alliance ruptures: diversity and integration. AB - This article reviews and synthesizes the diverse contributions of the authors in this issue of In Session: Psychotherapy in Practice. It presents a schematization of direct and indirect interventions that therapists typically implement to address problems related to the tasks and goals of treatment, or the affective bond between therapist and client. We then present an additional perspective on the resolution of therapeutic alliance ruptures, emerging out of our own research program. PMID- 10718607 TI - Determination of a molecular torsional angle in the metarhodopsin-I photointermediate of rhodopsin by double-quantum solid-state NMR. AB - We present a solid-state NMR study of metarhodopsin-1, the pre-discharge intermediate of the photochemical signal transduction cascade of rhodopsin, which is the 41 kDa integral membrane protein that triggers phototransduction in vertebrate rod cells. The H-C10-C11-H torsional angles of the retinylidene chromophore in bovine rhodopsin and metarhodopsin-I were determined simultaneously in the photo-activated membrane-bound state, using double-quantum heteronuclear local field spectroscopy. The torsional angles were estimated to be [phi] = 160+/-10 degrees for rhodopsin and phi = 180+/-25 degrees for metarhodopsin-I. The result is consistent with current models of the photo induced conformational transitions in the chromophore, in which the 11-Z retinal ground state is twisted, while the later photointermediates have a planar all-E conformation. PMID- 10718608 TI - Assessment of zinc finger orientations by residual dipolar coupling constants. AB - Residual dipolar coupling constants measured in anisotropic solution contain information on orientations between internuclear vectors and the magnetic field, providing long-range information that may help determine the relative orientations of distinct domains in biomolecules. Here we describe the measurement and use of residual dipolar coupling restraints in the refinement of the structure of the complex of DNA with three zinc fingers of transcription factor IIIA (TFIIIA), measured in a DMPC/DHPC bicelle solution. These dipolar restraints were applied on a variety of orientations of the zinc finger domains (derived from crystallography, previous NMR studies, and systematic modeling) in order to examine the validity and sensitivity of using residual dipolar splittings to study interdomain orientations. The spread in interdomain angles between zinc fingers is reduced from 24 degrees to 9 degrees upon incorporation of dipolar restraints. However, the results also show that the ability to determine relative orientations is strongly dependent on the structural accuracy of the local domain structures. PMID- 10718609 TI - Efficient analysis of macromolecular rotational diffusion from heteronuclear relaxation data. AB - A novel program has been developed for the interpretation of 15N relaxation rates in terms of macromolecular anisotropic rotational diffusion. The program is based on a highly efficient simulated annealing/minimization algorithm, designed specifically to search the parametric space described by the isotropic, axially symmetric and fully anisotropic rotational diffusion tensor models. The high efficiency of this algorithm allows extensive noise-based Monte Carlo error analysis. Relevant statistical tests are systematically applied to provide confidence limits for the proposed tensorial models. The program is illustrated here using the example of the cytochrome c' from Rhodobacter capsulatus, a four helix bundle heme protein, for which data at three different field strengths were independently analysed and compared. PMID- 10718610 TI - Intensity modulated HSQC and HMQC: two simple methods to measure 3J(HNH)alpha in proteins. AB - Two methods for the measurement of homonuclear 3J(HNH)alpha coupling constants are described. Both HSQC- and HMQC-type experiments employ 'quantitative J correlation', in which the coupling constant of interest is obtained from the intensity ratio of cross peaks of two spectra. The first spectrum is acquired with 3J(HNH)alpha evolution and the second with alpha-proton decoupling. The resolution of these methods in the F1-domain is not restricted. PMID- 10718611 TI - Determination of h2J(NN) and h1J(HN) coupling constants across Watson-Crick base pairs in the Antennapedia homeodomain-DNA complex using TROSY. AB - This paper describes NMR measurements of 15N-15N and 1H-15N scalar couplings across hydrogen bonds in Watson-Crick base pairs, h2J(NN) and h1J(HN), in a 17 kDa Antennapedia homeodomain-DNA complex. A new NMR experiment is introduced which relies on zero-quantum coherence-based transverse relaxation-optimized spectroscopy (ZQ-TROSY) and enables measurements of h1J(HN) couplings in larger molecules. The h2JNN and h1J(HN) couplings open a new avenue for comparative studies of DNA duplexes and other forms of nucleic acids free in solution and in complexes with proteins, drugs or possibly other classes of compounds. PMID- 10718612 TI - Refinement of the protein backbone angle psi in NMR structure calculations. AB - Cross-correlated relaxation rates involving the Calpha-Halpha dipolar interaction and the carbonyl (C') chemical shift anisotropy (CSA) have been measured using two complementary 3D experiments. We show that the protein backbone angle psi can be directly refined against such cross-correlated relaxation rates (gammaHalphaCalpha,C') and the three-bond H/D isotope effect on the Calpha chemical shifts (3 deltaCalpha(ND)). By simultaneously using both experimental parameters as restraints during NMR structure calculations, a unique value for the backbone angle psi is defined. We have applied the new refinement method to the alpha-Spectrin SH3 domain (a beta-sheet protein) and to the Sgs1p HRDC domain (an alpha-helical protein) and show that the quality of the NMR structures is substantially improved, judging from the atomic coordinate precision and the Ramachandran map. In addition, the psi-refined NMR structures of the SH3 domain deviate less from the 1.8 A crystal structure, suggesting an improved accuracy. The proposed refinement method can be used to significantly improve the quality of NMR structures and will be applicable to larger proteins. PMID- 10718613 TI - Conformational analysis of the Sd(a) determinant-containing tetrasaccharide and two mimics in aqueous solution by using 1H NMR ROESY spectroscopy in combination with MD simulations. AB - The conformational behaviour of the spacer-linked synthetic Sd(a) tetrasaccharide beta-D-GalpNAc-(1-->4)-[alpha-Neu5Ac-(2-->3)]-beta-D-Galp-(1-->4)- beta-D-GlcpNAc (1-->O)(CH2)5NH2 (1) and the two mimics beta-D-Galp-(1-->4)-[alpha-Neu5Ac-(2- >3)]-beta-D-Galp-(1-->4)-bet a-D-GlcpNAc-(1-->O)(CH2)5NH2 (2) and beta-D-GlcpNAc (1-->4)-[alpha-Neu5Ac-(2-->3)]-beta-D-Galp-(1-->4)- beta-D-GlcpNAc-(1- >O)(CH2)5NH2 (3) were investigated by 1H NMR spectroscopy in combination with molecular dynamics (MD) simulations in water. Experimental 2D 1H ROESY cross-peak intensities (ROEs) of the tetrasaccharides were compared with calculated ROEs derived from MD trajectories using the CROSREL program. Analysis of these data indicated that the oligosaccharidic skeletons of the compounds 1-3 are rather rigid, especially the beta-D-Hex(NAc)-(1-->4)-[alpha-Neu5Ac-(2-->3)]-beta-D-Galp fragments. The alpha-Neu5-Ac-(2-->3)-beta-D-Galp linkage occurred in two different energy minima in the three-dimensional structure of the compounds 1-3 in aqueous solution. Experimental data and dynamics simulations supported the finding that the higher energy rotamer (CHEAT forcefield) was abundant in compounds 1 and 3 due to the existence of a hydrogen bond between the carboxyl group of the sialic acid and the acetamido group of the terminal monosaccharide (GalNAc or GlcNAc) unit. The conformational similarity between 1 and 3 leads to the suggestion that also their activities will be alike. PMID- 10718614 TI - Backbone NMR assignments of a high molecular weight protein (47 kDa), cyclic AMP receptor protein (apo-CRP) PMID- 10718615 TI - Sequence-specific 1H, 15N and 13C resonance assignments for the third EH domain of Eps15. PMID- 10718616 TI - Backbone NMR assignment of the 19 kDa translationally controlled tumor-associated protein p23fyp from Schizosaccharomyces pombe. PMID- 10718617 TI - 1H, 13C, and 15N assignments of un-myristoylated Ca2+-frequenin, a synaptic efficacy modulator. PMID- 10718618 TI - Synergistic effects of 8-Cl-cAMP and retinoic acids in the inhibition of growth and induction of apoptosis in ovarian cancer cells: induction of retinoic acid receptor beta. AB - Both cAMP and retinoids play a role in cell differentiation and the control of cell growth. A site-selective cAMP analog, 8-Cl-cAMP and retinoic acid synergistically inhibit growth and induce apoptosis in certain cancer cells. In advanced or recurrent malignant diseases, retinoic acid (RA) is not effective even at doses that are toxic to the host. The objective of our present study was to examine the mechanism(s) of synergistic effects of retinoic acid (9-cis, 13 cis or all-trans RA) and 8-Cl-cAMP on apoptosis in human ovarian cancer NIH: OVCAR-3 and OVCAR-8 cells. RA induced growth inhibition and apoptosis in OVCAR-3 and OVCAR-8 cells. 8-Cl-cAMP acted synergistically with RA in inducing and activating retinoic acid receptor beta (RARbeta) which correlates with growth inhibition and apoptosis in both cell types. In addition, induction of apoptosis by RA plus 8-Cl-cAMP requires caspase-3 activation followed by cleavage of anti poly(ADP-ribose) polymerase. Furthermore, mutations in CRE-related motif within the RARbeta promoter resulted in loss of both transcriptional activation of RARbeta and synergy between RA and 8-Cl-cAMP. RARbeta expression appears to be associated with induction of apoptosis. Introduction of the RARbeta gene into OVCAR-3 cells resulted in gain of RA sensitivity. Loss of RARbeta expression, therefore, may contribute to the tumorigenicity of human ovarian cancer cells. Thus, combined treatment with RA and 8-Cl-cAMP may provide an effective means for inducing RARbeta expression leading to apoptosis in ovarian cancer cells. PMID- 10718619 TI - The effect of divalent cations on bovine retinal NOS activity. AB - The divalent cation requirements of NOS activity in bovine retina homogenate supernatant were investigated. Supernatants were assayed under standard conditions (in mM: EDTA 0.45, Ca2+ 0.25, Mg2+ 4.0). In order to investigate the enzyme's dependence on divalent cations, the tissue homogenate was depleted of di and trivalent cations by passing it over a cation-exchange column (Chelex 100). Surprisingly, NOS activity was 50-100% higher in this preparation. However, addition of either EDTA (33 microM) or EGTA (1 mM) almost fully inhibited NOS activity, suggesting a requirement for residual divalent metal cation(s). Phenanthroline or iminodiacetic acid at low concentrations had little effect on activity, suggesting no requirement for Fe2+, Zn2+ or Cu2+. Ca2+ had a moderate stimulatory effect, with an optimum activity around 0.01 mM. Mg2+ or Mn2+ had little effect at concentrations < 0.25 mM. However, in the presence of EDTA, Mn2+ or Ca2+ markedly stimulated NOS activity with the optimum at 0.1 mM. At high concentrations (> 0.1-0.2 mM), all divalent cations tested (Ba2+, Zn2+, Co2+, Mn2+, Mg2+, Ca2+), as well as La3+, dose-dependently inhibited NOS activity. We propose that retinal NOS requires low concentrations of naturally occurring divalent metal ions, most probably Ca2+, for optimal activity and is inhibited by high di- and trivalent metal concentrations, probably by competition with Ca2+. PMID- 10718620 TI - Deferiprone (L1) induced conformation change of hemoglobin: A fluorescence and CD spectroscopic study. AB - The interaction of deferiprone (1,2-dimethyl-3-hydroxy-pyrid-4-one) L1, the first clinically available oral iron chelator, with the tetrameric allosteric protein hemoglobin from human red blood cells has been investigated spectrofluorometrically and by circular dichroism spectroscopy. The interaction is hydrogenbond like electrostatic in nature, the binding constant being 4.54 x 10(3) M(-1) in 0.15 M NaCl. Circular dichroism studies indicate a conformational change of hemoglobin in presence of deferiprone, helicity of hemoglobin being reduced in presence of increasing concentration of the drug L1. PMID- 10718621 TI - Altered transarcolemmal Ca transport modifies the myofibrillar ultrastructure and protein metabolism in cultured adult ventricular cardiomyocytes. AB - The present study was designed to investigate how prolonged (24-72 h) exposure to modifiers of Ca transarcolemmal transport affects the myofibrillar structure, protein turnover and content of myofibrillar proteins in adult guinea pig cardiomyocytes maintained beating synchronously in long-term cultures. First we established the functional responses (the contractile activity and [Ca]i transients) of the cultured myocytes to acute exposures to several drugs used in this study. The ultrastructural characteristics of these cultures under the various treatments were determined using immunohistochemistry and confocal scanning laser microscopy, and their biochemical properties were evaluated using analysis of total cellular protein content, myofibrillar protein content and SDS PAGE electrophoretic examination. We compared the effects of 24, 36 and 72 h-long exposures to the various specific Ca-flux modifiers. Increased Ca influx via CaL channel agonist (Bay K 8644) or via the reversed-mode of the Na/Ca exchanger (veratrine) did not alter the myofibrillar structure or the specific protein profiles or proteosynthesis. However, when cytosolic Ca was increased by three different types of inhibitors of Ca extrusion from the cells via Na/Ca exchange, (Na-free solution, 5 mM NiCl2 and 10(-6) M ouabain), very significant changes in all investigated parameters occurred almost immediately. Twenty-four h-long exposure to Na-free did not affect significantly the total cellular protein (TCP), but the protein synthesis was decreased by 87% and the total myofibrillar protein (TMP) content was decreased by 38%. The myofibrils were heavily fragmented. Similarly, 24 h-long exposure to 5 mM NiCl2 did not affect the TCP, but it reduced protein synthesis by about 90% and decreased the total myofibrillar protein content by 30%. These effects were even more pronounced at 72 h of exposure and they were accompanied with a complete disassembly of myofilaments. Exposure to 10(-6) M ouabain over 72 h resulted in > 80% inhibition of protein synthesis, a 45% decrease in TCP content and a 53% in TMP content. In contrast, 10(-7) M ouabain did not produce any such changes. The changes produced by the Na/Ca-exchange inhibitors were accompanied by only minor changes in DNA content, indicating that the myocytes remained viable. Moreover, these effects were not due to the associated contractile arrest, since exposure to CaL-channel antagonists (5-20 microM nifedipine or 10 microM verapamil) produced only very minor changes in the myofibrillar structure and in protein profiles. Our data demonstrate that short-term (up to 72 h) increased Ca influx or contractile arrest of well-interconnected, spontaneously beating adult cardiomyocytes does not affect their ultrastructural characteristics or their myofibrillar protein turnover greatly, while any situations leading to Ca accumulation (via inhibition of Na/Ca exchange) affect cardiomyocyte function and ultrastructure almost immediately. These data are in sharp contrast to those previously reported from immature, neonatal myocytes. PMID- 10718622 TI - Transcriptional regulation of the human Na/K ATPase via the human mineralocorticoid receptor. AB - The mineralocorticoid receptor (MR) and the glucocorticoid receptor (GR) are members of the steroid/thyroid hormone receptor superfamily of ligand inducible transcription factors and have been shown to bind the glucocorticoid response element (GRE). Sodium-potassium ATPase (Na/KATPase) is a major target of mineralocorticoids. Both aldosterone and glucocorticoids activate the human Na/K ATPase alpha1 subunit and beta1 subunit genes transcriptionally. However, the mechanisms of corticosteroid regulation of mammalian Na/K ATPase subunit gene expression are not known. In this investigation, we report for the first time that cell lines (T-84 and 293) express endogenous MR by RT-PCR message expression. However, the protein product was not expressed as determined by western blot analyses. In transactivation studies of MR with GRE31, we detected MR expression at low concentrations of aldosterone. We also performed Northern blot and nuclear run-off transcription assays to further confirm that the regulation is transcriptional. We conclude that the transcriptional regulation of the human Na/K ATPase alpha1 and beta1 subunits by aldosterone occurs via the involvement of the MR. PMID- 10718623 TI - Effect of different whole body hyperthermic sessions on the heat shock response in mice liver and brain. AB - We examined by Western blots the effect of variations of the heating sessions, such as duration and intensity on the following aspects: 70-kDa heat shock protein (HSP70) and HSP72 induction. Protein ubiquitination PLCgamma , PKCepsilon and PKCalpha levels in murine liver and brain were also studied. Results demonstrated that maximal induction of HSP72 was obtained after heat shock at 43.5 degrees C in both organs. Preconditioning at lower temperatures (either acclimation to 39 degrees C or induction of thermotolerance to 43.5 degrees C with a single exposure to 39 degrees C) attenuated the heat shock response. Hepatic HSP72 induction was elicited only as a consequence of hyperthermia since either fasting or restraint were unable to trigger its synthesis. On the contrary, a ubiquitination decrease of a 31 kDa protein was obtained both after hyperthermia and fasting This indicates that the latter is a more generic response of hepatic cells to noxious stimuli. Analysis of the above mentioned enzymes showed that in liver of naive mice PKCalpha is barely present while PKCepsilon is quite abundant. All hyperthermic treatments caused a general decrease of the latter, except for the heat shock at 43.5 degrees C that caused an increase. PLCgamma decreased after all heating sessions. It is known that hyperthermia in the range of 41-45 degrees C induces apoptotic death in many cell types. Therefore we analyzed the presence of the typical apoptotic DNA ladder. Our data strongly suggest that both hyperthermia and restraint induce necrosis in liver while apoptosis and necrosis become evident in brain. All these effects are still present 24 h from the last heating session: This indicates that in vivo, hyperthermia produces long term modifications of the hepatic cell. PMID- 10718625 TI - Membrane changes in rat erythrocyte ghosts on ghee feeding. AB - Alterations in membrane lipid composition is known to result in functional and structural changes in the membrane, and dietary lipids play an important role in this change. It was of interest to study the influence of ghee feeding to the rat on membrane structure and function. The activities of membrane bound enzymes Na+ K+ ATPase and Acetylcholinesterase were studied as an index of membrane changes. Male weanling rats were fed 2.5% fresh or thermally oxidized ghee in the diet for a period of 8 weeks. The control rats were fed groundnut oil. A decrease of 28% in the membrane fluidity of erythrocyte ghost membranes was observed in the oxidized ghee fed group at 37 degrees C, by fluorescence polarization measurements using 1,6-Diphenyl-1,3,5-hexatriene as a probe. The activities of Na+ K+ ATPase and Acetylcholinesterase showed an increase of 65 and 200% respectively after feeding oxidized ghee (2.5%). Also changes in Na+, K+ and ATP kinetics were observed in these rats. Increased membrane lipid peroxidation (80%) and C/PL ratio (11%) in the oxidized ghee fed group was observed. Marginal changes in the fatty acid composition were also seen. Further, an increase in the osmotic fragility of erythrocytes was observed in the oxidized ghee fed rats. It is inferred from these experiments that consumption of oxidized ghee with the diet affects the erythrocyte ghost membrane structure and function at 2.5% level, whereas consumption of fresh ghee has no effect on the erythrocyte membrane. PMID- 10718624 TI - Selenoprotein W accumulates primarily in primate skeletal muscle, heart, brain and tongue. AB - The human selenoprotein W coding region with the selenocysteine codon (TGA) changed to a cysteine codon (TGT) was fused to six histidine codons (at its 3' end), cloned into a prokaryotic expression vector (pTrc99a), and the corresponding mutated selenoprotein W was expressed in bacteria. The protein was purified by Ni-NTA agarose column and reverse phase HPLC. Polyclonal antibodies raised against this protein were used in Western blots to determine tissue distribution of selenoprotein W from rhesus monkeys fed a commercial chow. Selenoprotein W was found in several tissues with highest amounts in skeletal muscle and heart (muscle 6 fold greater than liver) and lowest levels in liver, but selenium concentrations were highest in kidneys (10 fold greater than muscle) and lowest in skeletal muscle. Northern blots using a human selenoprotein W cDNA probe indicated that mRNA levels were highest in monkey skeletal muscle and heart (2-2.5 fold greater than in liver), which is similar to the pattern found with a human multiple tissue Northern blot. However, as in the monkey, selenium concentrations were highest in human kidney and lowest in skeletal muscle and heart. Thus, selenoprotein W protein levels correlated with selenoprotein W mRNA levels but not with tissue selenium concentrations. PMID- 10718627 TI - Interactions between Panax quinquefolium saponins and vitamin C are observed in vitro. AB - Inasmuch as the oxidation of low-density lipoprotein (Ox-LDL) may play a key role in the initiation and progression of atherosclerosis, it has become increasingly important to identify potential antioxidants. Panax quinquefolium saponins (PQS) are extracted from the stems and leaves of the North American form of ginseng, Panax quinquefolium. Our previous studies have indicated that PQS (0.25-1 mg/ml) can protect against oxidation of LDL in vitro. The purpose of the current work was to investigate the potential interaction of lower concentrations of PQS (1 100 microg/ml) with vitamin C on the reduction of LDL oxidation. LDL was isolated from the plasma of healthy human donors by sequential ultracentrifugation. Native LDL (0.05 or 0.2 mg/ml) was incubated with PQS and/or vitamin C for 30 min at 20 degrees C. Oxidative modification was initiated with 2 microM or 5 microM CuSO4 at 37 degrees C for (0-24 h. Pretreatment with PQS (100 microg/ml) reduced alterations in phospholipids, lipid peroxide levels and relative electrophoretic mobility of Ox-LDL. The presence of vitamin C (1-10 microM) significantly enhanced the protective effects of PQS. Pretreatment with PQS (1-100 microg/ml) resulted in concentration-dependent inhibition of LDL oxidation and prolongation of lag time as determined from measurements of conjugated lipid hydroperoxide content in Ox-LDL samples. Interestingly, the inhibitory actions of lower amounts of PQS (1 and 10 microg/ml) on the formation of conjugated dienes were significantly increased when vitamin C (0.1 or 1 microM) was present. In conclusion, our results suggest that PQS not only have direct antioxidant property but at low concentrations, their actions can be enhanced by vitamin C. PMID- 10718626 TI - Stabilization of collagen-tailed acetylcholinesterase in muscle cells through extracellular anchorage by transglutaminase-catalyzed cross-linking. AB - A component of collagen-tailed acetylcholinesterase (asymmetric; A-AChE) in muscle forms a metabolically-stable pool which can be released from the cell surface only by collagenase, suggesting that part of the enzyme is covalently bound by its tail (COLQ) subunits. We have investigated whether this insoluble pool forms through covalent cross-linking of A-AChE to extracellular matrix glycoproteins by tissue transglutaminase (Tg; type 2 transglutaminase). Tg catalyzed the incorporation of the polyamine substrate 3[H]-putrescine into the collagen tail of affinity-purified avian A12-AChE. Complexes between A12-AChE and cellular fibronectin were also formed in vitro by Tg. In quail myotubes, retinoic acid, which stimulates the formation of epsilon(gamma-glutamyl)lysine isodipeptide bonds by Tg in myotubes, increased the proportion of extraction resistant (er) A-AChE. Following irreversible inactivation of AChE by diisopropylfluorophosphate, entry of newly-synthesized A-AChE into the extraction resistant pool was inhibited by a competitive Tg inactivator RS48373-007. The quantity of exogenously-added A 12 AChE incorporated into the extraction resistant pool in living myotubes was increased by Tg in the presence of calcium. The inhibition of cross-bridge formation in fibrillar collagen by beta aminopropionitrile, and pre-exposure of myotubes to a monoclonal antibody to fibronectin, resulted in a reduction in the size of the erA-AChE pool present on the cell-surface. The evidence supports the hypothesis that a component of insoluble collagen-tailed AChE, once subject to clustering influences mediated via reversible docking to proteoglycans and their receptors, is anchored at the cell surface through covalent cross-linking by Tg. The high stability of the epsilon(gamma-glutamyl)lysine isopeptide bond is likely to contribute to the observed low turnover of the erA-AChE fraction. PMID- 10718628 TI - Effect of regulated expression of the fragile histidine triad gene on cell cycle and proliferation. AB - The mechanism of tumor suppressor action of the fragile histidine triad (FHIT) gene is unknown. Disruption of cell cycle regulation leads to the tumor formation and many tumor suppressor genes suppress tumorigenesis through their effect on cell cycle regulation. We examined the expression of FHIT during the cell cycle, and determined whether overexpression of FHIT affects cell cycle kinetics and apoptosis. The FHIT cDNA was cloned into the ecdysone-inducible expression vector in both the sense and antisense orientations. Overexpression of the sense or antisense construct did not affect cell proliferation, cell cycle distribution or apoptosis in human 293T cells. Analysis of the FHIT expression in 293T cells collected at various cell cycle phases showed that the expression of FHIT is not under cell cycle regulation. These results indicate that the tumor suppressor activity of the FHIT gene may be independent of an effect on the cell cycle and apoptosis mechanisms. PMID- 10718629 TI - Oxidative stress in the human heart is associated with changes in the antioxidative defense as shown after heart transplantation. AB - The study was designed to demonstrate--for the first time in humans--that oxidative stress in the heart indicated by lipid peroxidation is associated with time-dependent changes in the enzymatic antioxidative defense. For this purpose, we analyzed the oxygen radical metabolism in 69 myocardial biopsies (taken between the fifth day and 6 years after transplantation) of 31 heart transplant recipients who were suspected of suffering from increased formation of oxygen radicals in the allograft. The levels of lipid peroxides (LPO), glutathione peroxidase (GSH-Px), total-, copper/zinc- and manganese superoxide dismutase (t SOD, CuZnSOD, MnSOD) were compared in 3 post-transplantation periods (5-90 d vs. 91-365 d vs. >1 y). Significantly increased LPO levels were found (0.27+/-0.04 vs. 0. 13+/-0.02 vs. 0.27+/-0.04 nmol/mg protein) in the first and third period. Increased activities of GSH-Px (39.8+/-3.8 vs. 30.2+/-4.1 vs. 76.7+/-6.5 mU/mg protein), t-SOD (1.57+/-0.10 vs. 1.30+/-0.14 vs. 2.44+/-0.23 U/mg protein) and CuZnSOD (1.09+/-0.08 vs. 0.93+/-0.13 vs. 2.05+/-0.21 U/mg protein) occurred only in the third period. For calculation of time courses more precisely, the single data with respect to time were analyzed with a curve fitting program. Except for the first period, the allograft LPO and GSH-Px levels rose for up to 6 years after transplantation. However, the t-SOD and CuZnSOD activities switched from increase to decrease in the third period. The study provided indication for: first, the potency of the human heart to time-limited increase of the enzymatic antioxidative defense, and secondly, the inability of human heart allografts- despite this adaptation--for complete prevention of myocardial oxidative stress. PMID- 10718630 TI - Role of high-energy phosphate metabolism in hydrogen peroxide-induced cardiac dysfunction. AB - This study was undertaken to clarify the role of high-energy phosphate metabolism in hydrogen peroxide-induced cardiac dysfunction using phosphorus and fluorine nuclear magnetic resonance spectroscopy. The exposure of a Langendorff-perfused heart to hydrogen peroxide (200-400 micromol/L, 8 min) provoked biphasic contractile dysfunction characterized by a transient depression of left ventricular developed pressure during the administration of hydrogen peroxide and a delayed elevation of left ventricular end-diastolic pressure after the washout of hydrogen peroxide. The initial phase of cardiac dysfunction correlated well with the accumulation of sugar phosphates (r = 0.89, p < 0.01). Furthermore, we demonstrated that glibenclamide, a potent inhibitor of the ATP-sensitive K+ channel, attenuated the initial depression of developed pressure. On the other hand, the delayed elevation of end-diastolic pressure correlated well with the total ATP depletion (r = 0.96, p < 0.01). However, ATP loss was supposed to be a mere result from the increased ATP consumption corresponding to a rise in intracellular free Ca2+ (from the control value of 315+/-23 nmol/L to 708+/-104 after the administration of hydrogen peroxide, p < 0.01), which also paralleled the elevation of end-diastolic pressure. Thus glycolytic inhibition and intracellular Ca2+ overload are independently responsible for the biphasic contractile dysfunction induced by hydrogen peroxide. PMID- 10718631 TI - Intestinal mucins: the binding sites for Salmonella typhimurium. AB - Mucus-bacterial interactions in the gastrointestinal tract and their impact on subsequent enteric infections are poorly delineated. In the present study, we have examined the binding of Salmonella typhimurium to rat intestinal mucus and characterized a mucus protein (Mucus-Rs) which specifically binds to S. typhimurium. Both virulent (1402/84), and avirulent (SF 1835) S. typhimurium were observed to bind to crude mucus, however, the virulent strain showed 6 fold more binding as compared to avirulent strain. Fractionation of crude mucus on sepharose CL-6B resolved it into three major peaks. Maximal bacterial binding was observed with a high mol. wt. glycoprotein corresponding to neutral mucin. SDS PAGE of purified protein (termed Mucus-Rs) under non reducing conditions showed it to be a homogenous glycoprotein (mol. wt. 250 kDa), while under reducing conditions, three bands corresponding to mol. wt. of 118,75 and 60 kDa were observed. Pretreatment of Mucus-Rs with pronase, trypsin and sodium metaperiodate markedly inhibited bacterial binding. GLC analysis of Mucus-Rs showed it to contain Mannose, Glucose, Galactose, Glucosamine, Galactosamine and Sialic acid as main sugars. Competitive binding in the presence of various sugars and lectins indicated the involvement of mannose in the mucus-bacterial interactions. The Mucus-Rs binding was highly specific for S. typhimurium; no significant binding was seen with E. coli and V. cholerae. Thus, we conclude that S. typhimurium specifically binds to a 250 kDa neutral mucin of intestinal tract. This binding appears to occur via specific adhesin-receptor interactions involving bacterial pili and mannose of neutral mucin. PMID- 10718632 TI - Pirfenidone inhibits NADPH-dependent microsomal lipid peroxidation and scavenges hydroxyl radicals. AB - Pirfenidone (Pf), a new broad-spectrum anti-fibrotic agent, is known to offer protection against lung fibrosis in vivo in laboratory animals, and against mitogenesis and collagen formation by human lung fibroblasts in vitro. Because reactive oxygen species are thought to be involved in these events, we investigated the mechanism(s) by which Pf ameliorates oxidative stress and its effects on NADPH-dependent lipid peroxidation. Pf has been shown to cause inhibit NADPH-dependent lipid peroxidation in sheep liver microsomes in a dose-dependent manner. The concentration of Pf required to cause 50% inhibition of lipid peroxidation was approximately 6 mM. Pf was found to be ineffective as a superoxide radical scavenger. Pf was also ineffective in decomposing H2O2 and chelating iron. In deoxyribose degradation assays, Pf was a potent scavenger of hydroxyl radicals with a rate constant of 5.4 x 10(9) M(-1) sec(-1). EPR spectroscopy in combination with spin trapping techniques, using a Fenton type reaction and DMPO as a spin-trapping agent, Pf scavenged hydroxyl radicals in a dose-dependent manner. The concentration of Pf required to inhibit 50% signal height was approximately 2.5 mM. Because iron was used in the Fenton reaction, the ability of Pf in chelating iron was verified in a fluorescent competitive assay using calcein as the fluorescent probe. Pf up to 10 mM concentration was ineffective in chelating either Fe2+ or Fe3+ in this system. We propose that Pf exerts its beneficial effects, at least in part, through its ability to scavenge toxic hydroxyl radicals. PMID- 10718633 TI - Purification and characterization of intestinal adenosine deaminase from mice. AB - Adenosine deaminase (ADA) was isolated from small intestine of mice and purified to utmost homogeneity. SDS-PAGE of purified ADA gave a molecular weight of 41 kDa. Western blot analyses gave a single reactive band at 41 kDa and the other band was an associated ADA binding protein. The purified enzyme was more stable in the alkaline pH. The optimum pH and the pI values were about 7.0 and 4.96, respectively. Km values of the small intestinal ADA for adenosine and 2' deoxyadenosine were 23 and 16 microM, respectively. Purine riboside was a competitive inhibitor with Ki of 5 microM, whereas 2'-3'-o-isopropylidene adenosine acted as an uncompetitive inhibitor (Ki 66 microM). Activity of ADA was inhibited by the presence of theophylline (-40%), caffeine (-30%), and L-cysteine (-50%). Significantly, Hg2+ (100 microM) inhibited 98% of the initial ADA activity. In addition, various purine analogs such as inosine, purine, alpha adenosine and adenine showed variable inhibitions on the activity of ADA. Relative ADA activity towards 3'-deoxyadenosine and 6-chloropurine riboside was lower by 30% and 40%, respectively. However, the activity towards 2'-o-methyl adenosine was higher (30%) compared to the activity obtained using adenosine. PMID- 10718637 TI - 4-Dimethylaminobenzylamine as a sensitive chemiluminescence derivatization reagent for 5-hydroxyindoles and its application to their quantification in human platelet-poor plasma. AB - A selective and sensitive high-performance liquid chromatographic method with chemiluminescence detection for the determination of 5-hydroxyindoles is described, based on the reaction of 5-hydroxyindoles with 4 dimethylaminobenzylamine. Serotonin, 5-hydroxyindole-3-acetic acid, 5 hydroxytryptophol, 5-hydroxyindole-3-acetamide and N-acetyl-5-hydroxytryptamine were used as model compounds to optimize the derivatization and chemiluminescent reaction. The reagent reacts with 5-hydroxyindoles in slightly alkaline media in the presence of potassium hexacyanoferrate(III) to give the corresponding derivatives, which can be separated on a reversed-phase column, Wakosil-II 5C18RS, with aqueous acetonitrile as an eluent. The derivatives were detected by peroxyoxalate chemiluminescence detection. The detection limits are in the range of 0.5-1.2 fmol per 100-microl injection. The method was applied to the simultaneous determination of serotonin and 5-hydroxyindole-3-acetic acid in human platelet-poor plasma. PMID- 10718634 TI - N-myristoyltransferase. AB - Myristoylation refers to the co-translational addition of a myristoyl group to an amino-terminal glycine residue of a protein by an ubiquitously distributed enzyme myristoyl-CoA:protein N-myristoyltransferase (NMT, EC 2.3.1.97). This review describes the basic enzymology, molecular cloning and regulation of NMT activity in various pathophysiological processes such as colon cancer and diabetes. PMID- 10718636 TI - Regulation of heat shock protein 72 kDa and 90 kDa in human breast cancer MDA-MB 231 cells. AB - It has been shown that expression of HSPs can negatively regulate the effectiveness of cytotoxic drugs. In this study, we conducted experiments to study the regulation of expression of heat shock proteins (HSPs) in human breast cancer MDA-MB-231 cells. Using [35S]methionine incorporation and Western immunoblots, we established that heat shock increased production of HSP-72 and 90. Cells exposed to 44 degrees C for 20 min displayed increased expression of HSP-72 and -90, that reached a maximum 3-7 h later and returned to baseline levels within 24 h. The synthesis of both HSP-72 and -90 was attenuated when cells were exposed to heat shock in medium devoid of Ca2+ or pretreated with the calcium chelator BAPTA for 30 min prior to heat shock. Similarly, synthesis of HSP-72 and -90 was inhibited when cells were treated with the protein kinase A inhibitor, H89. These data indicate that Ca2+ and PKA are involved in the regulation of HSP-72 and -90 protein synthesis. Levels of HSP-72 mRNA in cells exposed to heat shock increased, suggesting that the heat-induced increase in HSP 72 occurs at the transcriptional level. Also, heat shock caused phosphorylation and translocation from the cytosol to the nucleus of heat shock factor 1 (HSF 1), a transcription factor for heat shock protein synthesis. Removal of external Ca2+ or treatment with a PKA inhibitor prevented the phosphorylation and the translocation of HSF 1. Cells overexpressing HSP-72 and -90 induced by exposure to a sublethal temperature displayed cytoprotection from thermal injury. Removal of external Ca2+ and treatment with BAPTA or H89 prior to exposure to sublethal heat shock that reduced the amount of HSP-72 and -90 production still protected cells from subsequent thermal injury. The intracellular free calcium concentration ([Ca2+]i) in resting fura-2-loaded MDA-MB-231 cells was 175+/- nM. Heat shock increased [Ca2+]i in a time-and temperature-dependent manner. Exposure of cells to 44 degrees C for 20 min increased [Ca2+]i by 234+/-13%, which subsequently returned to baseline levels within 30 min. Removal of external Ca2+ eliminated the increase, indicating that the increase in [Ca2+]i was due to Ca2+ influx. Pretreatment of the cells with H89 but not GF-109203X for 30 min led to an attenuation of the increase in [Ca2+]i by a subsequent heat shock. The results suggest that HSP-72 and -90 are regulated by [Ca2+]i and PKA activity in MDA-MB 231 cells. Kiang JG, Gist ID, Tsokos GC: Regulation of Heat Shock Protein 72 kDa and 90 kDa in Human Breast Cancer MDA-MB-231 Cells. PMID- 10718635 TI - Ethanol and arachidonic acid produce toxicity in hepatocytes from pyrazole treated rats with high levels of CYP2E1. AB - Ethanol and polyunsaturated fatty acids such as arachidonic acid were shown to be toxic and cause apoptosis in HepG2 cells which express CYP2E1 but not in control HepG2 cell lines. The goal of the current study was to extend the observations made with the HepG2 cells to non-transformed, intact hepatocytes. Rats were treated with pyrazole to increase CYP2E1 levels, hepatocytes were isolated and placed into culture and treated for varying time points with ethanol or arachidonic acid. Comparisons were made to hepatocytes from saline-treated rats, with low CYP2E1 content. Incubation with ethanol (100 mM) or especially arachidonic acid (60 microM) resulted in loss of viability of hepatocytes from the pyrazole-treated rats, without any effect on the hepatocytes from the saline treated rats. The toxicity appeared to be apoptotic in nature and was prevented by diallyldisulfide, an inhibitor of CYP2E1. Toxicity was reduced by trolox, an antioxidant. The treatment with ethanol or arachidonic acid resulted in release of cytochrome c into the cytosol fraction, and activation of caspase 3 (but not caspase 1) in hepatocytes from the pyrazole-treated rats but not hepatocytes from the saline-treated rats. The activation of caspase 3 was prevented by diallyldisulfide, by trolox, and by DEVD-fmk. The latter also prevented the toxicity produced by ethanol or arachidonic acid. These results extend previous observations found with HepG2 cells expressing CYP2E1 to intact hepatocytes and suggest that release of cytochrome c and activation of caspase 3 play a role in the overall pathway by which CYP2E1 contributes towards the hepatotoxic actions of ethanol and polyunsaturated fatty acids. PMID- 10718638 TI - Modelling alterations in the partition coefficient in in vitro biological systems using headspace gas chromatography. AB - Headspace gas chromatography was used to determine the physiological media-air partition coefficient (K) of four volatile organic solvents of industrial importance. The experimental conditions were those likely to be used in in vitro metabolic and toxicological studies on volatile compounds. The addition of solvent to the liquid phase from a stock solution in ethanol, or the presence of organic material at concentrations normally seen in in vitro studies, did not significantly alter the K value. Binary solvent addition resulted in a dose dependent decrease in K for each solvent that was also influenced by the solvent solubility and the constituents of the liquid matrix. The aromatic solvents exerted the greatest effect and showed the greatest change in K value. PMID- 10718639 TI - Immobilization of antibodies onto glass wool. AB - The immobilization of antibodies onto solid phases in an efficient and activity retaining form is an important goal for both research and industry. Methods have been developed for the site-directed attachment of antibodies to agarose by oxidation of the carbohydrate moieties in their Fc region. Similar attachment to silianized supports have not been as successful. Here we describe a novel combination protocol for the site-directed attachment of periodate oxidized, goat polyclonal antibodies to glass wool fibers activated with 3 aminopropyltriethoxysilane. The study demonstrates that this procedure results in effective immobilization of polyclonal antibodies that retain their antigen binding capacity. This protocol should prove useful in the development of more efficient and effective glass-based immunosupports. PMID- 10718640 TI - Two reproducible and sensitive liquid chromatographic methods to quantify atenolol and propranolol in human plasma and determination of their associated analytical error functions. AB - Two liquid chromatography (LC) methods with fluorimetric detection have been developed to measure atenolol and propranolol in human plasma. The same 5 microm Nucleosil RP-18 column, extraction procedure and mobile phase (containing acetonitrile, water, triethylamine and phosphoric acid, pH 3) were used. The linearity ranges were 25-800 ng/ml for atenolol and 3.13-100 ng/ml for propranolol. The coefficients of variation for validation assays were lower than 15% at the concentration assayed. The functions of the analytical error were linear: SD (ng/ml)=7.698+0.037C for atenolol and SD (ng/ml)=0.126+0.036C for propranolol. PMID- 10718641 TI - Determination of zearalenone and its metabolites alpha- and beta-zearalenol in beer samples by high-performance liquid chromatography-tandem mass spectrometry. AB - A fast, robust and sensitive LC-MS-MS method for the determination of zearalenone (ZON) and its metabolites alpha-zearalenol (alpha-ZOL) and beta-zearalenol (beta ZOL) in beer samples is described. Sample preparation was performed by direct RP 18 solid-phase extraction of undiluted beer samples followed by selective determination of analytes by LC-MS-MS applying an atmospheric pressure chemical ionization (APCI) interface. Using the negative ion mode limits of determination of 0.03-0.06 microg l(-1) beer and limits of quantification of 0.07-0.15 microg l(-1) beer were achieved, which was distinctly more sensitive than in the positive ion mode. Twenty-three beer samples from different countries, produced from different grains and under different brewing conditions, were investigated by this method, but only in one sample could beta-ZOL and ZON be detected. Independently of the type of beer, relative standard deviations between 2.1% and 3.3%, a linear working range of 0.15 microg l(-1) to 500 microg l(-1) beer and recovery rates around 100% could be achieved when zearalanone (ZAN) was used as internal standard. PMID- 10718642 TI - Effect of temperature on the separation of DNA fragments by high-performance liquid chromatography and capillary electrophoresis: a comparative study. AB - This study investigates the effect of experimental temperature on the separation of DNA fragments, 21-587 bp, by both high-performance liquid chromatography (HPLC) and capillary electrophoresis (CE). The results show that the temperature plays an important role in the HPLC separation of DNA fragments. The optimum temperature was found to be between 40 and 50 degrees C for HPLC, while 25 degrees C was the optimum temperature for the CE separation. Also, although CE migration times became shorter, efficiency and resolution decreased with an increase in temperature from 25 to 50 degrees C, but the separation was not significantly affected. Also, the optimum HPLC temperature might be different depending on the fragment sizes to be resolved. PMID- 10718643 TI - Measurement of plasma uracil using gas chromatography-mass spectrometry in normal individuals and in patients receiving inhibitors of dihydropyrimidine dehydrogenase. AB - A sensitive gas chromatographic-mass spectrometric method is described for reliably measuring endogenous uracil in 100 microl of human plasma. Validation of this assay over a wide concentration range, 0.025 microM to 250 microM (0.0028 microg/ml to 28 microg/ml), allowed for the determination of plasma uracil in patients treated with agents such as eniluracil, an inhibitor of the pyrimidine catabolic enzyme, dihydropyrimidine dehydrogenase. Calibration standards were prepared in human plasma using the stable isotope, [15N2]uracil, to avoid interference from endogenous uracil and 10 microM 5-chlorouracil was added as the internal standard. PMID- 10718644 TI - Novel sample preparation method facilitating identification of urinary drug metabolites by liquid chromatography-tandem mass spectrometry. AB - A simple, efficient procedure was developed for the preparation of urine samples, which greatly facilitated the identification of the urinary metabolites of a new antifungal agent SYN-2836. The urine samples following dilution with acetonitrile (ACN) formed distinct upper (ACN) and lower (aqueous) solution phases. The SYN 2836 metabolites were concentrated in the upper solution except that two glucuronides were concentrated in the lower solution. The upper solutions, containing concentrated metabolites and significantly reduced endogenous polar species, were ideally suitable for the metabolite identification. This novel sample preparation procedure would be applicable in identification of urinary metabolites of other drugs and drug candidates. PMID- 10718645 TI - Determination of estramustine phosphate and its metabolites estromustine, estramustine, estrone and estradiol in human plasma by liquid chromatography with fluorescence detection and gas chromatography with nitrogen-phosphorus and mass spectrometric detection. AB - Bioanalytical methods for the determination of estramustine phosphate by liquid chromatography and its four main metabolites estromustine, estramustine, estrone and estradiol by gas chromatography are described. For the estramustine phosphate assay the plasma was purified by protein precipitation followed by a C18 solid phase extraction. For the metabolite assay the plasma samples were purified by a C18 solid-phase and liquid-liquid extraction procedure and derivatised by silanization. Thereafter, estramustine and estromustine were quantified by gas chromatography with nitrogen-phosphorus detection and estradiol and estrone were quantified by gas chromatography with selected ion monitoring. The methods were validated with respect to linearity, selectivity, precision, accuracy, limit of quantitation, limit of detection, recovery and stability. The limit of quantitation was 2.3 micromol/l for estramustine phosphate, 30 nmol/l for estromustine and estramustine, 12 nmol/l for estrone and 8 nmol/l for estradiol. The results showed good precision and accuracy for estramustine phosphate and the four metabolites. The intermediate precision was 6.2-13.5% (C.V.) and the accuracy was 91.8-103.9%. PMID- 10718646 TI - Simultaneous determination of four anthracyclines and three metabolites in human serum by liquid chromatography-electrospray mass spectrometry. AB - A sensitive and very specific method, using liquid chromatography-electrospray mass spectrometry (LC-ES-MS), was developed for the determination of epirubicin, doxorubicin, daunorubicin, idarubicin and the respective active metabolites of the last three, namely doxorubicinol, daunorubicinol and idarubicinol in human serum, using aclarubicin as internal standard. Once thawed, 0.5-ml serum samples underwent an automated solid-phase extraction, using C18 Bond Elut cartridges (Varian) and a Zymark Rapid-Trace robot. After elution of the compounds with chloroform-2-propanol (4:1, v/v) and evaporation, the residue was reconstituted with a mixture of 5 mM ammonium formate buffer (pH 4.5)-acetonitrile (60:40, v/v). The chromatographic separation was performed using a Symmetry C18, 3.5 microm (150 x 1 mm I.D.) reversed-phase column, and a mixture of 5 mM ammonium formate buffer (pH 3)-acetonitrile (70:30, v/v) as mobile phase, delivered at 50 microl/min. The compounds were detected in the selected ion monitoring mode using, as quantitation ions, m/z 291 for idarubicin and idarubicinol, m/z 321 for daunorubicin and daunorubicinol, m/z 361 for epirubicin and doxorubicin, m/z 363 for doxorubicinol and m/z 812 for aclarubicin (I.S.). Extraction recovery was between 71 and 105% depending on compounds and concentration. The limit of detection was 0.5 ng/ml for daunorubicin and idarubicinol, 1 ng/ml for doxorubicin, epirubicin and idarubicin, 2 ng/ml for daunorubicinol and 2.5 ng/ml for doxorubicinol. The limit of quantitation (LOQ) was 2.5 ng/ml for doxorubicin, epirubicin and daunorubicinol, and 5 ng/ml for daunorubicin, idarubicin, doxorubicinol and idarubicinol. Linearity was verified from these LOQs up to 2000 ng/ml for the parent drugs (r > or = 0.992) and 200 ng/ml for the active metabolites (r > or = 0.985). Above LOQ, the within-day and between-day precision relative standard deviation values were all less than 15%. This assay was applied successfully to the analysis of human serum samples collected in patients administered doxorubicin or daunorubicin intravenously. This method is rapid, reliable, allows an easy sample preparation owing to the automated extraction and a high selectivity owing to MS detection. PMID- 10718647 TI - Determination by capillary zone electrophoresis of berenil, phenamidine, diampron and dibromopropamidine in serum and urine. AB - A quick, simple and reliable analysis method has been developed in order to determine berenil, phenamidine, diampron and dibromopropamidine by capillary zone electrophoresis in samples of serum and urine. In order to define the operation parameters in CZE, we have carried out a study on how the apparent electrophoretic mobility (mu(app)) varies when pH, buffer concentration, voltage and temperature are modified. Ohm's law plot has been studied, too. With the data obtained from this study we have determined the optimum work conditions, which are: citrate buffer 25 mM, pH=3.70, 14 kV, 30 degrees C, wavelength of the UV detector: 200 nm, capillary tube: 570 mm x 75 microm. Under these conditions, all the products appear in times between: 7.6 min phenamidine and 8.8 min dibromopropamidine, limits of detection being: berenil: 0.50, phenamidine: 0.25, diampron: 0.40 and dibromopropamidine: 0.80 microg ml(-1). We have carried out a recovery study with three kinds of extraction cartridges: Sep-pak C-18 plus, Sep pak C-8 plus and Oasis HBL for each one of the products in blood and urine. PMID- 10718648 TI - Determination of finasteride in human plasma by liquid-liquid extraction and high performance liquid chromatography. AB - A high-performance liquid chromatographic method for the quantitation of finasteride in human plasma is presented. The method is based on liquid-liquid extraction with hexane-isoamylalcohol (98:2, v/v) and reversed-phase chromatography with spectrophotometric detection at 210 nm. The mobile phase consists of acetonitrile-15 mM potassium dihydrogenphosphate (40:60, v/v). Clobazam is used as the internal standard. The limit of quantitation is 4 ng/ml and the calibration curve is linear up to 300 ng/ml. Within-day and between-day precision expressed by relative standard deviation is less than 5% and inaccuracy does not exceed 8%. The assay was used for pharmacokinetic studies. PMID- 10718649 TI - Fast quantification of the urinary marker of oxidative stress 8-hydroxy-2' deoxyguanosine using solid-phase extraction and high-performance liquid chromatography with triple-stage quadrupole mass detection. AB - Exogenous and endogenous oxidants constantly cause oxidative damage to DNA. Since the reactive oxidants itself are not suitable for analysis, oxidized bases like 8 hydroxy-2'-deoxyguanosine (8OHdG) are used as biomarkers for oxidative stress, either in cellular DNA or as elimination product in urine. A simple, fast and robust analytical procedure is described for urinary 8OHdG as an indicator of oxidative damage in humans. The adduct was purified from human urine by applying a single solid-phase extraction step on LiChrolut EN. After evaporation of the eluate, the residue was resolved and an aliquote was injected into a HPLC system with a triple quadrupole mass spectrometer. The limit of detection was 0.2 ng ml( 1) (7 fmol absolute) when using one product ion as quantifier and two further product ions as qualifier. The coefficient of variation was 10.1% (n=5 at 2.8 ng ml(-1) urine). The sample throughput was about 50 samples a day. Thus, this method is more sensitive and much faster than the common method using HPLC with electrochemical detection. The results of a study with nine volunteers investigated at six time-points each over 5 days are presented. The mean excretion of 8OHdG was 2.1 ng mg(-1) creatinine (range 0.17-5.9 ng mg(-1) creatinine; 4 of 53 samples were below the LOD). A relatively large intra- (relative SD 66%) and inter-individual (relative SD 71%) variation in urinary 8OHdG excretion rates was found. PMID- 10718650 TI - Determination of norepinephrine in small volume plasma samples by stable-isotope dilution gas chromatography-tandem mass spectrometry with negative ion chemical ionization. AB - A stable-isotope based gas chromatography-tandem mass spectrometry-negative ion chemical ionization method was developed for the determination of norepinephrine (NE) levels in small volumes (25-100 microl) of plasma. NE was stabilized in plasma by the addition of semicarbazide and spiked with deuterium-labeled norepinephrine internal standard. The analytes were isolated from the plasma by solid-phase extraction using phenylboronic acid columns and derivatized using pentafluoropropionic anhydride. The derivatized analytes were chromatographed on a capillary column and detected by tandem mass spectrometry with negative ion chemical ionization. Unparalleled sensitivity and selectivity were obtained using this detection scheme, allowing the unambiguous analysis of trace levels of NE in small-volume plasma samples. Linear standard curves were obtained for NE over a mass range from 1 to 200 pg per sample. The method had a limit of quantitation of 10 pg NE/ml plasma when using a 100-microl sample aliquot (1 pg/sample). Accuracy for the analysis of plasma samples spiked with 10 to 200 pg NE/ml typically ranged from 100+/-10%, with RSD values of less than 10%. The methodology was applied to determine the effect of clonidine on plasma NE levels in conscious spontaneously hypertensive rats. Administration of clonidine (30 microg/kg) produced an approximately 80% reduction in plasma NE accompanied by a 30% reduction in heart and mean arterial pressure that persisted >90 min after drug administration. The ability to take multiple samples from individual rats allowed the time course for the effect of clonidine to be mapped out using only one group of animals. PMID- 10718651 TI - Semi-automatic liquid chromatographic analysis of olpadronate in urine and serum using derivatization with (9-fluorenylmethyl)chloroformate. AB - The semi-automatic bioanalytical assays for olpadronate [(3-dimethylamino-1 hydroxypropylidene)bisphosphonate] involves a protein precipitation with trichloroacetic acid and a double co-precipitation with calcium phosphate for serum samples and a triple calcium co-precipitation for urine samples. These manual procedures are followed by an automated solid-phase extraction on a cation exchange phase. The procedure is continued either directly, at high olpadronate levels in urine, or after off-line evaporation under nitrogen and reconstitution in water on the same robotic workstation. The continued automatic procedure comprehends derivatization with (9-fluorenylmethyl)chloroformate, ion-pair liquid liquid extraction and ion-pair HPLC with fluorescence detection at 274/307 nm. The intra- and inter-day precisions for urine and serum samples are typically in the 5-8% range for different olpadronate concentrations [levels near the lower limit of quantification (LLQ) excluded]. The LLQ is 5 ng/ml olpadronate for a 2.5 ml urine sample and 10 ng/ml for a 1-ml serum sample, respectively. PMID- 10718652 TI - Chromatographic investigation on the binding site characteristics of quail egg white riboflavin binding protein as a chiral stationary phase. AB - Recently we described the use of riboflavin binding protein extracted from quail egg-white, as a new HPLC chiral stationary phase. In this study we show the further results obtained with the use of high-performance affinity chromatography to provide a better understanding of the chiral recognition mechanism for the observed enantioselectivity and to gain a deeper knowledge into the binding site that has been recently characterised by X-ray crystallography for chicken egg white. High-performance affinity chromatography provides information on the potential protein structural changes occurring upon its immobilisation and enables competitive binding studies as well as the assessment of binding constants through frontal analysis experiments. PMID- 10718653 TI - High-performance liquid chromatographic assay for minocycline in human plasma and parotid saliva. AB - A sensitive and specific high-performance liquid chromatographic (HPLC) method with UV detection was developed for the determination of minocycline in human plasma and parotid saliva samples. Samples were extracted using an Oasis HLB cartridge and were injected into a C8 Nucleosil column. The HPLC eluent contained acetonitrile-methanol-distilled water-0.1% trifluoroacetic acid (25:2:72.9:0.1, v/v). Demeclocycline was used as internal standard. The assay showed linearity in the tested range of 0.1-25 microg/ml. The limit of quantitation was 100 ng/ml. Recovery from plasma or parotid saliva averaged 95%. Precision expressed as %CV was in the range 0.2-17% (limit of quantitation). Accuracy ranged from 93 to 111%. In the two matrices studied at 20 and 4 degrees C, rapid degradation of the drug occurred. Frozen at -30 degrees C, this drug was stable for at least 2 months, the percent recovery averaged 90%. The method's ability to quantify minocycline with precision, accuracy and sensitivity makes it useful in pharmacokinetic studies. PMID- 10718654 TI - Simultaneous determination of 6beta- and 6alpha-hydroxycortisols and 6beta hydroxycortisone in human urine by stable isotope dilution mass spectrometry. AB - A capillary gas chromatographic-mass spectrometric method for the simultaneous determination of 6beta-hydroxycortisol (6beta-OHF, 6beta,11beta,17alpha,21 tetrahydroxypregn-4-ene- 3,20-dione), 6alpha-hydroxycortisol (6alpha-OHF, 6alpha,11beta,17alpha,21-tetrahydroxypregn-4-ene -3,20-dione) and 6beta hydroxycortisone (6beta-OHE, 6beta,17alpha,21-trihydroxypregn-4-ene-3,11,20 trione) in human urine is described. Deuterium-labelled compounds, 6beta [1,1,19,19,19-2H5]OHF (6beta-OHF-d5), 6alpha-[1,1,19,19,19-2H5]OHF (6alpha-OHF d5) and 6beta-[1,1,19,19,19-2H5]OHE (6beta-OHE-d5) were used as internal standards. Quantitation was carried out by selected-ion monitoring of the characteristic fragment ions ([M-31]+) of the methoxime-trimethylsilyl (MO-TMS) derivatives of 6beta-OHF, 6alpha-OHF and 6beta-OHE. The sensitivity, specificity, precision and accuracy of the method were demonstrated to be satisfactory for measuring 6beta-OHF, 6alpha-OHF and 6beta-OHE in human urine. PMID- 10718655 TI - Determination of celecoxib in human plasma by normal-phase high-performance liquid chromatography with column switching and ultraviolet absorbance detection. AB - A method is described for the determination of celecoxib in human plasma. Samples were extracted using 3M Empore membrane extraction cartridges and separated under normal-phase HPLC conditions using a Nucleosil-NO2 (150x4.6 mm, 5 microm) column. Detection was accomplished using UV absorbance at 260 nm. The HPLC method included a column switching procedure, in which late eluting compounds were diverted to waste, to reduce run-time to 12 min. The assay was linear in the concentration range of 25-2000 ng/ml when 1-ml aliquots of plasma were extracted. Recoveries of celecoxib were greater than 91% over the calibration curve range. Intraday precision and accuracy for this assay were 5.7% C.V. or better and within 2.3% of nominal, respectively. The assay was used to analyze samples collected during human clinical studies. PMID- 10718656 TI - Reversed-phase liquid chromatographic column switching for the determination of N methylcarbamates and some of their main metabolites in urine. AB - A column switching system for the determination of some polar pesticides and their main metabolites, such as aldicarb, aldicarb sulphoxide, aldicarb sulphone, carbofuran and 3-hydroxicarbofuran, in human urine has been developed. The limits of detection were between 0.3 and 1 microg/l. We used a simple solid-phase extraction with graphite carbon and a RPLC-LC analysis with UV detection yielding average recoveries between 84 and 110% (N=5) with RSD between 4 and 8%. PMID- 10718657 TI - Analysis of an adenine nucleotide-containing metabolite of clodronate using ion pair high-performance liquid chromatography-electrospray ionisation mass spectrometry. AB - Clodronate belongs to the family of bisphosphonates, which are synthetic analogues of pyrophosphate. Bisphosphonates are widely used in the treatment of metabolic bone diseases. Some bisphosphonates, including clodronate, can be metabolized in cells into non-hydrolysable nucleotide analogues. In this paper, we describe a new method for extraction and quantitation of the clodronate metabolite in cell lysates by using ion-pairing HPLC method that is compatible with negative ion electrospray ionization mass spectrometry (ESI-MS). The method was used for detection of the metabolite of clodronate in extracts from RAW 264 macrophage cells after treatment with clodronate. PMID- 10718658 TI - Determination of erythromycin, clarithromycin, roxithromycin, and azithromycin in plasma by high-performance liquid chromatography with amperometric detection. AB - In this study, a high-performance liquid chromatographic method was developed for the quantitative determination of erythromycin (EM), roxithromycin (RXM), and azithromycin (AZM) in rat plasma with amperometric detection under a standardized common condition using clarithromycin (CAM) as an internal standard. This method was also proved to be applicable for the determination of CAM by employing RXM as an internal standard. Each drug was extracted from 150 microl of plasma sample spiked with internal standard under an alkaline condition with tert.-butyl methyl ether. The detector cell potential for the oxidation of the drugs was set at +950 mV. The linearity of the calibration curves were preserved over the concentration ranges of 0.1-10 microg/ml for EM and RXM, and 0.03-3.0 microg/ml for CAM and AZM. Coefficients of variation and relative error were less than 9% and +/-7%, respectively. The analytical method presented here was proved to be useful for the investigation of the pharmacokinetic characteristics of EM, CAM, RXM, and AZM in rats. PMID- 10718659 TI - High-performance liquid chromatographic determination of quercetin and isorhamnetin in rat tissues using beta-glucuronidase and acid hydrolysis. AB - Quercetin is a plant polyphenol which is present in the diet as an aglycone and as sugar conjugates. Despite potent vasodilatory and antioxidant effects in vitro, destruction by intestinal organisms has been assumed to limit its nutritional relevance in the rat. However, we have refined extraction techniques using beta-glucuronidase followed by acid hydrolysis. Following this with HPLC methodology with post-column derivatisation, we have detected significant concentrations of quercetin and its metabolite, isorhamnetin, in tissues of rats maintained on quercetin-rich diets. Percentage recoveries are greater than 95% and intra-batch variation does not exceed 7% suggesting that the method may be useful in further studies of the biological role of this flavonoid. PMID- 10718660 TI - Simultaneous stereoselective high-performance liquid chromatographic determination of 10-hydroxycarbazepine and its metabolite carbamazepine-10,11 trans-dihydrodiol in human urine. AB - An enantioselective HPLC method for the simultaneous determination of the concentration of the enantiomers of the oxcarbazepine metabolites 10 hydroxycarbazepine (MHD) and carbamazepine-10,11-trans-dihydrodiol (DHD) in human urine is described. The method is based on extraction with tert.-butylmethyl ether-dichloromethane (2:1, v/v) under alkaline conditions, separation and evaporation of the organic phase and dissolution of the residue in the mobile phase. Enantiomers are resolved on a Diacel Chiralcel OD column (250 mm x 4.6 mm I.D.) under isocratic conditions using as mobile phase n-hexane-ethanol-2 propanol (18:2:1, v/v/v) with addition of glacial acetic acid (0.1%). The enantiomers are detected by UV at 215 nm. The method allows reliable determination of the MHD and DHD enantiomers in human urine with limits of quantification of 0.2 mg/l and 0.4 mg/l, respectively. PMID- 10718661 TI - Gas chromatographic method for the determination of toluidines in spiked urine samples. AB - A capillary gas-chromatographic method was developed for the analysis of a mixture of toluidines in urine. The method is based on the extraction of toluidines with toluene and derivatisation with heptafluorobutyric anhydride to form a product for electron capture detection. The procedure gave a linear response at concentrations of 0.02-0.20 microg/ml with sufficient reproducibility. The method is simple, requires little sample pretreatment and is being considered for biomonitoring workers exposed to toluidines. PMID- 10718662 TI - Development and validation of a new high-performance liquid chromatographic estimation method of meloxicam in biological samples. AB - A simple, HPLC method was developed to estimate meloxicam (COX-2 inhibitor) using piroxicam as the internal standard. The mobile phase containing methanol, acetonitrile and an aqueous solution of diammonium hydrogenorthophosphate (50 mM) in the ratio of 4:1:5 was pumped at the rate 1 ml/min. Lichrocart RP-18 (125 x 4 mm) was used as an analytical column and the analytes were detected at 364 nm using a UV detector. Acidified plasma samples were extracted with chloroform, evaporated to dryness, reconstituted in the mobile phase and then a volume of 10 microl of the prepared sample was injected in the column. The retention time of meloxicam and piroxicam was found to be 2.7 and 1.9, respectively. This method showed an accuracy of 102.3% at 0.52 microg/ml and was capable of detecting a minimum concentration of 0.029 microg/ml meloxicam from biological samples. The analytical method was successfully utilized for estimating meloxicam in biological samples. PMID- 10718663 TI - Validated method for the therapeutic drug monitoring of flunitrazepam in human serum using liquid chromatography-atmospheric pressure chemical ionization tandem mass spectrometry with an ion trap detector. AB - An HPLC-MS-MS method for the quantitative analysis of flunitrazepam in human serum was established. The method features a very simple liquid-liquid extraction, the use of a standard 4-mm HPLC column, isocratic elution using a buffer-free solvent, short retention times in connection with good peak resolution and the sensitivity and selectivity of an ion trap MS-MS detector. The procedure enables unambiguous identification of analytes by their product ion spectra, as well as sensitive quantitation (limit of quantitation for flunitrazepam=0.5 ng/ml). This feature was used for the confirmation of HPLC-UV results for nitrazepam. PMID- 10718664 TI - Accuracy verification of the photostereometric system KKN/1B developed for intraoperative measurement of knee movement immediately after total knee arthroplasty. AB - The function and integrity of the knee joint following total knee arthroplasty (TKA) is determined at first by the design and implantation of the prosthesis, and later by the tension of soft tissues surrounding it. Accurate post-TKA motion data obtained intraoperatively could be used not only to optimize implantation techniques from a kinematic standpoint, but also to improve prosthetic design. We therefore developed a system specifically geared to photostereometric measurement of 6 d.o.f. knee motion. A total of eight LEDs are mounted on the prosthetic components in two sets of four by means of connecting measuring-bows. The positions of the LEDs are detected in three-dimensions by two sets of three linear CCD cameras, located bilaterally relative to the knee. The position and orientation of the femoral component relative to the tibial one are estimated from the positions of all LEDs in the sense of least-squares. Based upon results of various accuracy validation experiments performed after precise camera calibration, static overall accuracy and spatial resolution were considered to lie within 0.52 and 0.11 mm, respectively, at any point on the femoral articular surface. PMID- 10718665 TI - A needle-type immunotherapeutic system incorporating laser light and platonin in combination with ethanol injection in the treatment of cancer growing in deep organs. AB - Platonin is a potent cell-activating agent and a photosensitizing dye characterized by a typical absorption peak at 590 nm. It has been already clarified that macrophage activity is enhanced synergistically by platonin administration together with laser light irradiation. This method is considered to be useful in cancer immunotherapy. In this study, a needle-type therapeutic system incorporating this method was applied for the therapy of a homogeneous tumor grafted onto mice. The tumor was strongly cicatrized by collagen fibers at an early stage of the treatment. A high T/B cell ratio of lymphocytes was observed in the peripheral blood of the treated mice. In addition, a high survival ratio was seen, especially in the group of concomitant therapy with ethanol injection (PEI method). It was determined that the induction of high immunopotentiation of mice was triggered by ethanol injection at an early stage. These results demonstrated that the system is reliable for the treatment of cancer in deep organs such as a liver, requiring no major surgical intervention. PMID- 10718666 TI - A newly designed underwater antenna and its application to underwater radio telemetry for measuring electroencephalographic activity from the rainbow trout freely swimming in natural environments. AB - A novel underwater antenna (which we named an 'aquaerial') for telemetering the biological signals from freely swimming fish in freshwater natural environments is presented. It is designed for receiving a 90-100 MHz carrier wave and consists of plural unit receiving antennas (UAs). The plural UAs are placed underwater to cover the area where the target fish carrying the transmitter is swimming. The UAs are equally spaced and have a directional coupling amplifier to supply the signals received to the coaxial cable. The optimal length of the UA was found to be 16.5 cm (a half wavelength in water) and optimal spacing was 2 m (one wavelength along coaxial cable) when 95 MHz was used as the carrier frequency. Using this 'aquaerial', long-term monitoring of EEG signals from the olfactory bulb of the rainbow trout (Oncorhynchus mykiss) swimming freely in natural environments was achieved. PMID- 10718667 TI - Three-shell head model constructed from scalp geometry for electroencephalogram dipole localization. AB - One of the most sophisticated methods of dipole localization from (electroencephalograms) is based on a multi-compartment head model, which consists of the scalp, skull, cerebrospinal fluid (CSF) and brain cortex. The CSF layer in normal subjects is thin and can be ignored in dipole localization to good approximation. This multi-shell model is called the SSB (scalp-skull-brain) head model. The SSB head model is constructed from magnetic resonance imaging (MRI) or X-ray computed tomography slices of the head. These image slices, however, are not easy to obtain. The present paper presents a simple method for constructing the SSB head model only from the geometrical shape of the scalp, which is easily measured with a three-dimensional digitizer. We have derived a relationship between the head structure and the geometry of the head from statistical analysis of MRI slices of 12 normal subjects. The average estimation errors of the outer and inner boundaries of the skull were 1.2 and 2.2 mm, respectively. The head model thus estimated is called the sSSB (statistical SSB) model. PMID- 10718668 TI - Muscle activity-torque-velocity relations in human elbow extensor muscles. AB - With the aid of an artificial neural network technique, we investigated relationships between the torque and extending velocity of an elbow at constant muscle activation in healthy volunteers. Each subject sat on a chair and was able to move his upper- and forearm on a shoulder-high horizontal plane. First, with the gravitational force of a weight hanging from a pulley, the subject's wrist was pulled to flex the elbow. Next, the subject was instructed to extend his elbow joint at a constant velocity. Integrated electromyograms (IEMGs), elbow joint angle and torque were measured while the elbow was being extending. Then the relationships among these three variables were modeled using an artificial neural network where IEMGs, joint angle and velocity were the inputs, and torque was the output. After back propagation learning, we presented various combinations of IEMGs, elbow joint angle and velocity to the model, and estimated the elbow joint torque to obtain the torque-velocity relationship for constant muscle activation. The torque decreased in a nearly linear manner as the velocity increased. This was caused by slow extending velocity and was explained by Hill's equation at slow velocity. PMID- 10718669 TI - A low-distortion filter method to reject muscle noise in multi-lead electrocardiogram systems. AB - This paper proposes a low-distortion filter method for rejection of muscle noise in multilead electrocardiogram (ECG) systems. This approach combines low-pass filtering with a modified version of source consistency filtering. The low-pass filtering splits the raw ECG signal into a muscle-noise-free part and a muscle noise-overlapping part. The modified source consistency filtering then extracts the signal components from the muscle-noise-overlapping part. The extracted signal components are restored to the muscle-noise-free part as the output. The performance of our method was verified with simulated and clinically recorded ECG signals. The simulated ECG signals were created from computer simulation using three-dimensional, realistically shaped heart and torso models. The ideal signals are superimposed with white noise to simulate muscle noise and with a low frequency sine wave to simulate baseline drift. For verification, our method was compared with Butterworth low-pass filters. The results show that our method can effectively reduce muscle noise with less distortion of the QRS wave than conventional low-pass filters. PMID- 10718670 TI - Effect of local anesthetics on red blood cell deformability. AB - The deformability of human red blood cells under the influence of local anesthetics is studied in vitro. Red blood cells obtained from blood samples of healthy adult volunteers are treated with two local anaesthetics, procain and tetracaine, at varied concentrations. Deformability of red blood cells, in terms of its elongation index, were then examined by laser light shear stress diffractometer both at low and high shear forces. Results show that the deformability of local anesthetic-treated red blood cells is decreased when compared to that of control samples. At low shear forces and at low concentrations of the anesthetic agent, the decrease in deformability is not significant, whereas it is significant for high shear forces and at high concentrations. These effects are similar for both anesthetic agents studied. PMID- 10718672 TI - Double blind comparison of three hearing aid circuits with new hearing aid users. AB - This study was a double blind comparison of three types of hearing aid circuits: Class A linear peak clipping, Class D compression limiting and K-Amp wide dynamic range compression. Subjective ratings, speech perception tests, real ear measurements and questionnaire data were obtained from a group of 17 new hearing aid users with mild to moderate sensorineural hearing loss. The results indicate a similar performance for all three circuits. We saw no evidence of performance degradation due to saturation distortion, even in the presence of high levels of speech and noise. Our primary conclusions include recommending K-Amps to new hearing aid users with mild to moderate hearing loss, mostly on the basis of battery life, while cautioning about the use of compression knee-point controls and recognizing that Class A and Class D amplifiers are virtually equivalent in every performance measurement. PMID- 10718671 TI - Epidemiology of hereditary hearing disorders in childhood: a retrospective study in Germany with special regard to ethnic factors. AB - Recent studies have revealed marked differences in the prevalence of hearing impairment in childhood. It remains unknown whether this is caused by geographic, sociocultural or ethnic factors. In order to provide consistent data, 314 children in residence at a school for hearing impaired children of a well-defined German area were investigated retrospectively. We estimated an overall prevalence of .43/1000 of hearing impairment for this area. A striking variability between different nationalities could be observed. For German children, the prevalence was .36/1000, for Turkish 1.04/1000 and for Italian children 1.37/1000. Hereditary hearing losses and, in particular, autosomal recessive caused hearing disorders were the most common cause of hearing impairment in foreign but not in native children. One reason might be the frequency of marriages between relatives resulting from the social isolation of the foreign groups in German society. PMID- 10718673 TI - Optimization of TEOAE recording protocols: a linear protocol derived from parameters of a time-frequency analysis: a pilot study on neonatal subjects. AB - Linear and non-linear TEOAE protocols were compared in terms of nine parameters in order to define the protocol producing recordings with the highest signal quality (lowest noise and highest signal-to-noise ratio). The pilot project acquired data using ILO-92 apparatus from 220 neonates (397 ears) at the second/third day after birth in three European laboratories. A Gabor spectrogram time-frequency representation of the recordings showed considerable frequency dispersion in TEOAE latencies >4.0 ms. The data, analysed with a Wilcoxon test, indicated that a linear TEOAE protocol: (i) generates recordings of a lower noise and a higher signal-to-noise (S/N) ratio in the 2.0, 3.0, 4.0 kHz TEOAE bands; (ii) the increase in the S/N ratio can result in a decrement of the required number of TEOAE sweeps; (iii) the higher values of S/N can be used in the estimation of more robust pass-fail criteria, minimizing the percentage of false positives and negatives. PMID- 10718674 TI - Hyperacusis: case studies and evaluation of electronic loudness suppression devices as a treatment approach. AB - Hyperacusis, as defined here, is a relatively rare condition in which the patient, with or without hearing loss, experiences severe loudness discomfort to everyday environmental sound levels. The case studies of 14 patients with severe hyperacusis are described; all wore passive attenuators (earplugs and/or earmuffs) in an attempt to alleviate their discomfort, frequently producing communication difficulties. These subjects were fitted binaurally with experimental electronic loudness suppression devices housed in in-the-ear casings. The devices supplied low-level amplification followed by an extreme form of amplitude compression for moderate or high-level inputs in an attempt to reduce loudness discomfort without reducing audibility. Many of the subjects were found to function with a wider dynamic range with the active devices compared with passive attenuators or the unoccluded ear, and most reported that they benefited from the devices in at least some listening situations. PMID- 10718675 TI - Reliability and validity of a Danish adaptation of the Tinnitus Handicap Inventory. AB - The objective of this study was to determine the reliability and validity of a Danish translation of the Tinnitus Handicap Inventory (THI), a self-report measure of perceived tinnitus handicap. The Danish version of the THI was administered to 50 patients reporting tinnitus as their primary complaint or secondary to hearing loss. Construct validity was assessed using tinnitus symptom rating scales, the Beck Depression Inventory (BDI), the State-Trait Anxiety Inventory (STAI), the Tinnitus Coping Style Questionnaire (TCSQ), the Eysenck Personality Questionnaire (EPQ), and perceived tinnitus loudness and pitch. The Danish translation of the THI and its subscales showed good internal consistency reliabilities (c = 0.93 to alpha = 0.74) comparable to those of the original version. High to moderate correlations were observed between THI and psychological distress, tinnitus symptom ratings, neuroticism and maladaptive tinnitus coping. A confirmatory factor analysis failed to validate the three subscales of THI, and high intercorrelations found between the subscales question whether they represent three distinct factors. The results suggest that the Danish THI-Total scale may be a reliable and valid measure of general tinnitus related distress that can be used in a clinical setting to quantify the impact of tinnitus on daily living. PMID- 10718676 TI - Objective detection of auditory brainstem potentials: comparison of statistical tests in the time and frequency domains. AB - Newborn hearing screening with auditory brainstem potentials (ABR) requires objective ABR detection by a statistical test procedure with high performance. Statistical testing can be performed in the time or frequency domain. The aim of the present study was to compare the performance of three tests in the time domain (Friedman test, variance ratio F(SP), Cochran's Q-test) with that of a test in the frequency domain (modified q-sample uniform scores test) that, in a former investigation, was shown to be the best test in the frequency domain. To compare the performance of the four tests, the test power was calculated and receiver operating characteristics (ROCs) were constructed from the probability density functions estimated using Monte Carlo simulations. In addition, a comparison on the basis of real near-threshold ABR data was carried out. The modified q-sample uniform scores test appeared to be the most powerful one. Some aspects of practical application are discussed. PMID- 10718677 TI - Database for a hearing conservation program. AB - We have developed a database and an analysis program (NoiseScan) for noise induced hearing loss (NIHL). The exposure data are based on the evaluation of the noise immission level, which includes duration, frequency content, and the use of, and the attenuation performance of, hearing protectors. The input data can handle an unlimited number of exposure periods. If the noise exposure level is not known, the program lists noise levels of comparable work places, and thus provides an estimate of exposure. Confounding medical factors that may contribute to NIHL, such as elevated serum cholesterol level, hypertension, and extensive use of pain killers, are collected. Combined exposure to agents that clearly contribute to NIHL, such as hand-arm vibration, tobacco smoking, use of aminoglycosides and exposure to solvents are also assessed. An unlimited number of audiograms can be stored, and all the data can be completed and edited following collection. The program gives the predicted hearing loss according to the ISO 1999 model based on total exposure. At present, our NoiseScan program (under continuous development in an EU research program) is suitable for the data collection of various risk factors. It can be used to determine whether the hearing loss is occupational in origin and to estimate the efficiency of hearing conservation measures. NoiseScan also predicts the development of hearing loss in individuals in 5-year periods. The goal is to improve and validate the rules by which single and combined risk factors contribute to HIHL, thus leading to more precise prediction of individual hearing loss, and for the evaluation of success of the hearing conservation programs. PMID- 10718678 TI - Sleep organization and regulation. AB - Sleep is a vital, complex state with as yet unknown functions. It is active and highly organized and is regulated by homeostatic, circadian, and ultradian processes. It consists of two distinct sleep states, rapid eye movement (REM) and non-rapid eye movement (NREM), both of which have a dramatic impact on many aspects of physiology and behavior. The significance and consequences of the REM NREM organization of sleep are not known. On the other hand, the sleep state and its organization are quite fragile and dynamic. Any number of factors can disrupt sleep and its expression, and its nature changes over the life span. What is certain is that any reduction and/or disruption of sleep hinders an organism's ability to navigate through the waking state. PMID- 10718679 TI - Diagnosis and treatment of sleep disorders caused by co-morbid disease. AB - Sleep disorders, both insomnia and hypersomnia, are commonly associated with various co-morbid conditions, including general medical and neurologic disorders, psychiatric illnesses, and secondary or symptomatic restless legs syndrome/periodic limb movements in sleep. Diagnosis of the co-morbidity is the first step in treatment, followed by an assessment of the sleep disturbance. This begins with a complete history and physical examination, followed by laboratory testing such as polysomnography, multiple sleep latency testing, and actigraphy. The treatment of sleep disorders caused by co-morbid conditions is discussed under seven categories, including the use of general measures (e.g., sleep hygiene, encouraging patients to develop good sleep habits, medication), treatment of the co-morbid condition, treatment of insomnia, treatment of excessive daytime somnolence, treatment of parasomnias, treatment of sleep-wake schedule disorders, and treatment of secondary or symptomatic restless legs syndrome. PMID- 10718680 TI - Effect of anticonvulsants on sleep. AB - The intensive study of seizures via serial 24-hour EEG-video monitoring has allowed enhanced observation of sleep patterns among epilepsy patients and has revealed that disturbed rest is common in this population. Previous work has shown that sleep deprivation of any type can exacerbate seizure activity, leading to speculation that the intrinsically poor sleep in these patients may serve as a threshold-lowering factor, and that this factor might be partially reversed by effective antiepileptic drugs (AEDs). However, to better understand this interaction, it is necessary to know whether the sleep disorder of epilepsy is caused by repetitive ictal events or whether it is part of a process that causes epilepsy to emerge in the first place. In addition, to separate and analyze the sleep effects of AEDs, one must compare studies of normal controls, which have only rarely been accomplished, with studies of drug-free patients, which are difficult to achieve. As little as is known about the detailed effects of AEDs on sleep architecture, even less is known about the mechanisms by which AEDs might cause such effects. Nevertheless, there is great potential for those undertaking such work, due to the wealth of basic science accumulating in the field of sleep mechanisms and the prodigious amount of information already amassed in the area of anticonvulsant mechanisms. PMID- 10718681 TI - Effect of anticonvulsants on nocturnal sleep in epilepsy. AB - Our objective was to determine, in three separate studies, the effects of controlled-release carbamazepine (CBZ-CR), lamotrigine (LTG), and gabapentin (GBP) on nocturnal sleep in epilepsy. Antiepileptic drugs (AEDs) control seizures and also modify hypnic structure. Despite widespread clinical use, their effects on sleep are not well known. PSG was performed in all three studies as follows: CBZ-CR: at baseline, after initial administration of CBZ-CR 400 mg, and after 1 month of CBZ-CR treatment (400 mg BID) in a sample of seven temporal lobe epileptic (TLE) patients. Results were compared with those of nine healthy volunteers; LTG: at baseline, after 3 months of stable treatment with LTG (300 mg/day); GBP: at baseline, after 3 months of stable treatment with GBP (1800 mg/day). Significant findings are as follows for each study. The acute administration of CBZ-CR increased number of stage shifts, reduced REM sleep, and increased REM sleep fragmentation. In the TLE group, these effects were almost completely reversed after chronic treatment. LTG increased REM sleep, reduced number of entries into REM sleep, decreased number of phase shifts, and decreased percentage of slow-wave sleep. GBP increased REM sleep percentage, increased mean duration of REM periods, reduced number of awakenings, and reduced stage 1 sleep percentage. We conclude that CBZ-CR disrupts REM sleep, but only during acute administration. LTG and GBP improve sleep stability while reducing seizures. PMID- 10718682 TI - Importance of sleep restoration in co-morbid disease: effect of anticonvulsants. AB - Over the past two to three decades, sleep medicine has emerged as an important discipline as it strives to meet the challenges of some of the most prevalent disorders among humans. Among the 110 disorders listed in the International Classification of Sleep Disorders, two of the most prevalent and treatable have only recently begun to receive significant attention: sleep apnea and restless legs syndrome with sleep-related periodic limb movements disorder. It is becoming clear that the sleep disruption caused by such disorders has ramifications beyond the usually associated daytime sleepiness, and may include: exacerbation of seizures, headaches, short-term memory deficits, and other cognitive problems. Sleep apnea has also been correlated with hypertension and cardiovascular/cerebrovascular disease. Animal studies have taken this one step further by demonstrating that total sleep deprivation is consistently fatal, usually within 1 month, although the precise mechanism remains to be discovered. The most compelling finding in the animal studies is that "rescuing" the animals with sleep, before the irreversible stage, is associated with rebound amounts of deep sleep and rapid eye movement (REM) sleep ("dream sleep"). This same response is seen after initiating treatment of sleep apnea with nasal continuous positive airway pressure (CPAP), and can also occur in patients with other sleep disorders in response to particular medications, such as valproate or gabapentin. PMID- 10718683 TI - Celecoxib for rheumatoid arthritis. PMID- 10718684 TI - Exercise as an effective treatment option for major depression in older adults. PMID- 10718685 TI - Noninvasive glucose monitoring. PMID- 10718686 TI - Outcomes for new anti-hypertensives in the elderly. PMID- 10718687 TI - Improving prevention systems in primary care practices: the Health Education and Research Trial (HEART) AB - BACKGROUND: The Health Education and Research Trial (HEART) was a multicenter clinical trial designed to test methods to improve primary care practice systems for heart disease prevention services. We present the trial methodology, the practices' use of medical record tools, and changes in documentation of cardiovascular risk factor screening and management. METHODS: Primary care practices were recruited from 4 Midwestern states. The factorial design resulted in 4 study groups: conference only, conference and quality improvement consultations, conference and prevention coordinator, and all interventions combined. Medical record audits and physician, staff, and patient surveys assessed practice change in cardiovascular disease risk factor documentation. RESULTS: Practices participated fully in this project, set goals to improve preventive services, and implemented recommended medical record tools. The number of goals set and the increase in the use of medical record tools were greatest in the combined intervention group, with improvements noted in all groups. The use of patient history questionnaires, problem lists, and flow sheets was significantly higher in the combined intervention group when compared with the conference-only group. Documentation of risk factor screening in a recommended medical record location improved in all intervention groups, with significant sustained improvements in the practices that received the combined intervention. Documented risk factor management significantly improved in all intervention groups compared with the conference-only control. CONCLUSION: Primary care practices are interested in improving prevention systems and can change these systems in response to supportive external interventions. Promoting organizational change to produce sustained improvement in preventive service clinical outcomes is a complex process that requires further research. PMID- 10718688 TI - Changing physician practice behavior: the merits of a diagnostic approach. PMID- 10718689 TI - Should children be in the room when the mother is screened for partner violence? AB - BACKGROUND: The goal of our study was to understand the important issues to consider when screening women for intimate partner violence in front of their children. METHODS: Interviews and focus groups were conducted with experienced family physicians and pediatricians and family violence experts (child psychologists, social workers, and domestic violence agency directors). Session transcripts were coded and categorized. RESULTS: Experts disagreed on the appropriateness of general screening for intimate partner violence in front of children older than 2 to 3 years. The majority thought that general questions were appropriate, if the in-depth questioning of the abused parent was done in private. Screening for child abuse when domestic violence is identified (and for domestic violence when child abuse is discovered) was recommended. Documentation about intimate partner violence in the child's medical chart raises questions about confidentiality, since the person committing the abuse may have access, if he or she is a legal guardian. Physicians need more education on the symptoms of children who are exposed to violence between adults. CONCLUSIONS: More research is needed to understand appropriate questions and methods of screening for intimate partner violence in front of children. The tension is between practical recommendations for routine screening and preserving the safety of the parent and the children. Intimate partner violence screening by physicians is important. Interrupting the cycle of violence may give a child a better chance at maturing into a healthy adult. PMID- 10718690 TI - Screening for family violence: overcoming the barriers. PMID- 10718691 TI - Low-income women's priorities for primary care: a qualitative study. AB - BACKGROUND: Because of their challenging social and economic environments, low income women may find particular features of primary care uniquely important. For this qualitative study we explored which features are priorities to women fiumi low-income settings and whether those priorities fit into an established primary care framework. METHODS: We performed a qualitative analysis of 4 focus groups of women aged 40 to 65 years from 4 community health clinics in Washington, DC. Prompted by semistructured open-ended questions, the focus groups discussed their experiences with ambulatory care and the attributes of primary care that they found important. The focus groups were audiotaped, and the tapes were transcribed verbatim and coded independently by 3 readers. RESULTS: The comments were independently organized into 5 content areas of primary care service delivery plus the construct of patient-provider relationship in the following order of frequency: accessibility (37.4%), the physician-patient relationship (37.4%), comprehensive scope of services (11.5%), coordination between providers (6.8%), continuity with a single clinician (3.7%), and accountability (3.2%). Commonly reported specific priorities included a sense of concern and respect from the clinicians and staff toward the patient, a physician who was willing to talk and spend time with them (attributes of the physician-patient relationship), weekend or evening hours, waiting times (attributes of organizational accessibility), location in the inner city and on public transport routes (an attribute of geographic accessibility), availability of coordinated social and clinical services on-site; and, availability of mental health services on-site (attributes of comprehensiveness and of coordination). CONCLUSIONS: All attributes of care that were priorities for low-income women fit into 1 of 6 content areas. Specific features within the content areas of accessibility, physician-patient relationship, and comprehensiveness were particularly important for these women. PMID- 10718692 TI - Gender differences in the utilization of health care services. AB - BACKGROUND: Studies have shown that women use more health care services than men. We used important independent variables, such as patient sociodemographics and health status, to investigate gender differences in the use and costs of these services. METHODS: New adult patients (N = 509) were randomly assigned to primary care physicians at a university medical center. Their use of health care services and associated charges were monitored for 1 year of care. Self-reported health status was measured using the Medical Outcomes Study Short Form-36 (SF-36). We controlled for health status, sociodemographic information, and primary care physician specialty in the statistical analyses. RESULTS: Women had significantly lower self-reported health status and lower mean education and income than men. Women had a significantly higher mean number of visits to their primary care clinic and diagnostic services than men. Mean charges for primary care, specialty care, emergency treatment, diagnostic services, and annual total charges were all significantly higher for women than men; however, there were no differences for mean hospitalizations or hospital charges. After controlling for health status, sociodemographics, and clinic assignment, women still had higher medical charges for all categories of charges except hospitalizations. CONCLUSIONS: Women have higher medical care service utilization and higher associated charges than men. Although the appropriateness of these differences was not determined, these findings have implications for health care. PMID- 10718693 TI - Patient beliefs about the characteristics, causes, and care of the common cold: an update. AB - BACKGROUND: Many people seek medical care for cold symptoms. The cold-related knowledge and beliefs of adults seeking medical care for themselves or their children may not correspond with current medical opinion. METHODS: A total of 249 parents of symptomatic children and 257 symptomatic adults who sought medical advice in the spring of 1997 from 1 of 3 primary care clinics in the Minneapolis St. Paul, Minnesota, area were surveyed by telephone 48 to 96 hours after contact with the medical system. RESULTS: Of the adults seeking care for a child or themselves, 44% believed viruses alone cause the common cold: an additional 42% believed both viruses and bacteria play a role. Most thought rest (97%) and nonprescription medications (63%) were helpful for colds, which was consistent with published reports. Contrary to medical reports, however, most felt vitamin C (67%) and the inhalation of steam (70%) reduced cold symptoms, and 44% believed antibiotics help colds (chi2=19.57; P=.0002). But 85% believed colds could resolve on their own. CONCLUSIONS: Those adults seeking medical care for uncomplicated colds are misinformed about the primary cause of the common cold, the use of prescription medications for treating cold symptoms, and the effectiveness of some palliative care techniques. Care providers should address these perceptions rather than enabling overuse of antibiotics. PMID- 10718694 TI - Review of primary care-based physical activity intervention studies: effectiveness and implications for practice and future research. AB - OBJECTIVE: To summarize the literature on primary care-based interventions for increasing physical activity and make recommendations for future research and for integrating successful strategies into practice. SEARCH STRATEGIES: We searched MEDLINE (1980 to 1998), psychological abstracts, ERIC and HealthStar databases, the WeB site for The Journal of Family Practice, bibliographies of selected studies, and previous reviews for relevant articles. The search was limited to the English language. Three experts in the field of physical activity were contacted for leads on unpublished trials. SELECTION CRITERIA: Inclusion criteria were: randomized controlled trial or quasiexperimental study using a comparison group, intervention delivered or initiated in a primary care setting, and reported results on at least 1 measure of physical activity. Studies that focused solely on patients with cardiovascular disease were excluded. MAIN RESULTS: Primary care-based physical activity counseling is moderately effective in the short term, although there is considerable variability across studies. Studies in which the interventions were tailored to participant characteristics and which offered written materials to patients produced stronger results. Unlike many types of health promotion, the reach of primary care-based physical activity interventions is high. Questions remain about the consistency of implementation and long-term maintenance of outcomes. CONCLUSIONS: Despite the need for further research, enough is known to recommend integration of key strategies of physical activity counseling into routine practice. We recommend incorporating these strategies into primary care and prioritizing them for further research. PMID- 10718695 TI - Age-specific patterns of prostate-specific antigen testing among primary care physician visits. AB - BACKGROUND: Early detection of prostate cancer is thought to be effective, and indirect evidence suggests that men aged 50 to 69 years will benefit most while those aged 70 and older will benefit least from it. The goal of our study was to describe usual care patterns for prostate-specific antigen (PSA) testing by primary care physicians in the United States. METHODS: We analyzed office visits made by adult men to family physicians, general internists, general practitioners, and geriatricians recorded by the 1995 and 1996 National Ambulatory Medical Care Surveys. Our outcome measure was the probability of a primary care physician ordering a PSA test during a visit. RESULTS: Seventeen percent of the tests reported were among men aged younger than 50 years, 50% were for men aged 50 to 69 years, and 33% were for men aged 70 years and older. The frequency of PSA testing was highest during visits by men aged 60 to 64 years (7.1%), 65 to 69 years (7.0%), 70 to 74 years (7.0%), and 75 to 79 years (6.3%) but lower for men aged older than 80 years (3.1%). CONCLUSIONS: Our findings suggest that during the mid-1990s prostate cancer screening decisions by primary care physicians were not sensitive to patients' ages. PMID- 10718696 TI - Best vasectomy technique? PMID- 10718697 TI - Best vasectomy technique? PMID- 10718698 TI - Oseltamivir for flu prevention. PMID- 10718699 TI - Screening for intracranial aneurysms in high-risk relatives. PMID- 10718700 TI - Best treatment for single-vessel coronary artery disease. PMID- 10718701 TI - The use of tocolytics in preterm labor. PMID- 10718702 TI - Some current obstetric and gynaecological problems. AB - It has never been right to practise obstetrics and gynaecology in a purely mechanistic style. Medical practitioners have always discussed and argued, often driven by their patients, both the ethical framework and the individual circumstantial detail of practice within this specialty. This chapter starts by discussing the place of ethics and minimum core values for practice. Their integration into every doctor-patient relationship must be taught from the earliest training. It is not possible to catalogue every circumstance within the specialty where there are ethical problems to be addressed. Principles are outlined and many examples within the context of routine general specialty practice including aspects of pregnancy and fertility control are considered. PMID- 10718703 TI - New reproductive technologies. AB - The ethical calculus is described. There are four entities that have a stake in all ethical decisions. These are the individual, family, community and society at large. Values assigned to these entities can be modified by experience, cultural background, religious authority and the law. In the simple case of in vitro fertilization (IVF), there seems to be general international acceptance, except for the official position of the Roman Catholic Church, the origin of which dissent is extensively discussed. There is concern about the number of pre zygotes/pre-embryos to be transferred, the disposition of any unfertilized egg and the reporting of results. Clear ethical considerations arise as IVF is applied in more complex situations, such as unmarried couples, the use of donor gametes and donor pre-embryos, the use of cryopreservation, intracytoplasmic sperm injection (ICSI), prenatal genetic diagnosis, surrogate gestational motherhood and research on the pre-embryo. The ethical, religious and legal acceptance of these depends on the weight given to elements of the ethical calculus as described in the chapter. PMID- 10718704 TI - Ethics in fetal medicine. AB - In this chapter we provide an overview of current ethical issues in fetal medicine. We begin with an introduction to the language and concepts of ethics. We then show how ethical principles can be applied to fetal medicine. In this part of the chapter we introduce the concept of the fetus as a patient and explain its implications for directive versus non-directive counselling. We then take up two current clinical controversies, routine obstetric ultrasound and termination of third trimester pregnancy. PMID- 10718705 TI - Neonatal care: withholding or withdrawal of treatment in the newborn infant. AB - Life-sustaining treatment may be ethically withdrawn or withheld in critically ill or dying newborns if the action is genuinely in the best interests of the patient. This may occur in situations where life-sustaining treatment is futile because of a hopeless prognosis, or if the burdens of intensive treatment clearly outweigh its likely benefits. There is no fundamental ethical difference between the withholding of resuscitation and the withdrawing of life-sustaining treatment once it has commenced. However the actions may have different emotional and psychological implications. A decision to withdraw treatment should only be taken with the consensus of experienced staff caring for the baby and with the unpressurized agreement of the parents. Palliative care and symptomatic relief should always continue after life-support has been withdrawn. Emotional and practical support should be provided for parents and adequate training and support is essential for obstetric and neonatal unit staff. PMID- 10718706 TI - Practical clinic problems. Part I: Conclusions. PMID- 10718707 TI - Alternative regulatory perspectives of obstetrics and gynaecology. AB - In addressing whether the field of obstetrics and gynaecology demands internal self-regulation or external state intervention, the authors look at the distinct yet at once interrelated notions of ethics, deontology and law. The distinctness of these concepts relates not only to their optional and/or obligatory natures, but also to the group and/or groups they influence and/or regulate. The interrelated nature of these concepts suggests that they are complimentary paradigms in as much as they collectively form a framework of individual, professional and societal norms. Given the emergence of both legal pluralism and self-regulation over the last two decades, it is believed that neither self regulation nor state intervention is in itself appropriate. Rather, it is contended that only a pluralistic system--incorporating ethics, deontology (self regulatory models) and law (state regulatory model) can effectively and comprehensively regulate the medical community so that it may efficaciously and appropriately respond to the continually emerging issues raised by contemporary medicine. PMID- 10718708 TI - From state eugenics to private eugenics. AB - Eugenics--or 'the cultivation of a race'--is a concept dating from the latter part of the 19th century. It preceded the new science of genetics by merely 25 years. Negative eugenics stressed especially the exclusion of negative characteristics and was associated with the practice and theory of radical eugenics between the two World Wars. In order to redress 'the decline of the race', reinforcement by positive eugenics was also advocated. After the atrocities committed by the Nazis there was a lull in the practice and discourse of eugenics. More recent technical advances in assisted reproduction techniques and the genome project, however, have revived the eugenics debate. State eugenics and eugenics as an individual choice ought to be distinguished. PMID- 10718709 TI - Economics and justice: the ethical aspects of inequity or inequality in health care. AB - Inequality in access to health care exists under a variety of aspects in different parts of the world. In the USA, the absence of universal health insurance leaves 15% of the population, including 12 million children, unprotected. Managed care and health maintenance organisations (HMOs) tend to further deepen this inequality. In Europe, where state-regulated Social Security covers most of the population, welfare policies affect income-related inequalities leaving others untouched, namely inequalities related to family history, education and occupation. In developing countries, poverty is the main cause of inaccessibility to health care and both internal structural reforms and international support could contribute to alleviating such an appalling injustice. PMID- 10718710 TI - Medical practice and society. Part II: Conclusions. PMID- 10718711 TI - International aspects of ethical problems in obstetrics and gynaecology. AB - Until the middle of the twentieth century, family planning was commonly condemned as an immoral enterprise. This chapter argues that ethical disapproval made access to fertility regulation difficult, and that while the rich and educated were often able to access the services they needed, typically the poor could not. Thus, ethical interpretations caused much of the differential fertility observed between social classes, ethnic groups and ultimately much of the explosion in population that has characterized the last 50 years. Since the 1960s human rights have been interpreted to encompass the right to decide the number and spacing of children and as a result access to contraception has improved. Where several methods of contraception (backed up by safe abortion) are available, fertility always falls. But, unfortunately, consensus has not been reached on how to specify particular sets of rights (e.g. decision-making for adolescents), and no consensus exists on how to balance the opposing rights of the mother and fetus in the case of abortion. The ethics of the one-child family in China is also noted. PMID- 10718712 TI - Will we be taught ethics by our clones? The mutations of the living, from endocrine disruptors to genetics. AB - Considering the worldwide threat to health and reproduction related to endocrine disruptors (by-products of the chemical industry); considering the untrammelled development of the industrialization and engineering of the living, ethics and gynaecology/obstetrics itself is at a crossroads. Endocrine disruptors (derived from organochlorines and persistent organic pollutants such as PCBs, dioxins and furans, and pesticides such as aldrin, chlordane and DDT), are prime suspects in the deterioration of fertility and intellectual faculties and possibly a key factor in endometriosis, breast cancer and prostate cancer. The long-term and pernicious impacts of endocrine disruptors show our poor understanding of the complexities of life's mechanisms. Paradoxically, with our short-term perspectives and predilection for a technological fix, the problem posed by endocrine disruptors may accelerate the use of reproductive technologies such as ICSI and even cloning, as well as the dissemination of genetically modified organisms. The cure could be worse than the disease. Given the gravity of the challenge to humanity related to the chemical erosion of human health, the mutation of human conception introduced by reproductive technologies and by the drive to genetically modify nature and even human nature, we must urgently re evaluate the direction in which our societies are headed and the reliance on profit-oriented technology to save us from ourselves. In these circumstances, the collective exercise of wisdom, prudence and responsibility towards the essence and integrity of humanity has become, more than ever, an ethical, and perhaps even a survival, imperative. PMID- 10718713 TI - General conclusion: back from law, economics, politics and philosophy to clinical practice. What future, what role, what responsibility for the practising obstetrician gynaecologist? PMID- 10718714 TI - Academic medicine into the millennium: the UK scene. AB - Clinical academic medicine has a vital role to play in the introduction and development of new therapies. The clinical scientist is essential in asking for answers to questions that arise from clinical practice. However, clinical academics should not be overburdened with clinical service or teaching commitments as these functions are shared by National Health Service staff. The exciting developments in molecular genetics and the study of the interaction between environmental and genetic factors emphasize the importance of clinical academic medicine and the clinician scientist into the millennium. PMID- 10718715 TI - The recruitment and retention of academic staff in the UK. AB - This paper deals with a detailed survey of medical and dental schools in the UK and a poll of the views and experiences of specialist registrars. Between 1991 1992 and 1995-1996 the total number of academic staff in the medical schools increased roughly in line with the increase in the number of medical students. In 1995-1996 there were 19 professorships and 27 other academic posts vacant through lack of suitable candidates. The major incentive to young doctors to follow an academic career is the challenge of research, but research is the first to suffer when they are under pressure from demand for more patient care. The balance of activity between patient care, teaching and training, and research has been disturbed by pressures for more service and revisions of the medical curriculum. Suggestions are made for the protection of research time. Aspects of training are discussed at length and recommendations made for improvements. It is suggested that efforts should be made to align more closely the management of medical schools and of the trusts with which they are associated. PMID- 10718716 TI - Factors affecting career choices in medicine. AB - The choice of medicine as a career is often limited by lack of knowledge concerning the wide range of options facing the applicant. Once enrolled, many students regret the choice. Over half the entrants to medical schools are women and career choice is viewed as limited. Careers advice is often patchy and career choices are usually made late and well into the pre-registration year. The choice of obstetrics and gynaecology as a career is low and the choice of an academic career does not register at all. Given that many students have high academic aspirations, it is difficult to understand why this should be the case. PMID- 10718717 TI - The impact of structured training on academic medicine in the UK. AB - Introduction of structured training has been a recent event. The programmes have been modelled on curricula produced by the medical royal colleges. Regular assessment throughout the training has helped to achieve the designed goals. This reform encourages research for up to 1 year. However, the research year would not be funded by the normal National Health Service channels. The period of research can be extended by a year without losing the national training number. If the specialist registrars take this towards the end of year 4 then they can continue in research and acquire the certificate of 'Completion of Specialist Training' yet continue with the research for a degree by thesis. Clinical competence needs new ways of measurement without adhering to time periods of training. This will enable clinicians not to turn away from academic medicine because of longer periods of training needed prior to being appointed to a substantive academic post, compared with a colleague who is pursuing a clinical career. PMID- 10718718 TI - Factors influencing recruitment and retention of academic staff in the European Union. AB - Academic medicine plays an essential role in the organization of health care in the European Union. Academic hospitals have been the Mecca for advanced medicine. Academic hospitals differ from country to country. Some are public institutions and others are largely private hospitals. Financial pressures on patient care are now threatening research activities in the university hospitals and universities have a responsibility to safeguard the structure of clinical academic obstetrics and gynaecology. In many European teaching hospitals, little importance is placed on research and the academic posts are a mechanism for developing private practice. PMID- 10718719 TI - Factors that may influence recruitment into academic obstetrics and gynaecology in European Union countries. AB - Factors that may influence recruitment into academic obstetrics and gynaecology in European Union countries were studied by a questionnaire method. The survey attracted responses from 15 professors and nine clinical obstetricians and gynaecologists. The majority considered a clinical academic to be one who worked in public hospitals. Although 15 of 16 respondents did not think that the academics had a high status in society, 75% felt that young obstetricians and gynaecologists wished to pursue an academic career. Despite the fact that there was a reasonable prospect of promotion and reasonable salary many did not pursue academic medicine because of several factors. They were no governmental support, relatively lower pay compared with clinical colleagues, long hours of work, much severe competition because of few posts and poor condition of work. PMID- 10718720 TI - Factors influencing recruitment and retention of academic staff in South-East Asia. AB - South-East Asia has a number of developing and developed countries. The economic standings of these countries are different and influence the number of staff in each academic department. The quantum of clinical work they are expected to perform and the remuneration vary enormously. The retirement age, facilities for research and criteria for promotion are different. The health care delivery system, the financial status and outlook of the universities also differ. These factors operate differently in the South-East Asian countries and determine the recruitment and retention of academic staff. PMID- 10718721 TI - Profile and recruitment in Australasia. AB - There are 12 medical schools in Australia and New Zealand producing a total of 1600 graduates per year. Academic recruitment is poor as a result of lack of career paths, poor financial remuneration, problems with lack of research funding and the small size of departments. Increased demands from the clinical service and lack of 'start-up' research funds act as a deterrent to recruitment of junior academics. Clinical academics need to convince governments that, without strong and continuing academic leadership, there will be a deterioration in patient care, teaching and research in obstetrics and gynaecology. PMID- 10718722 TI - Profile of recruitment in Sweden. AB - Sweden enjoys a good doctor and specialist obstetrician and gynaecologist to population ratio. The medical education for 5.5 years is state funded. The specialist training period is flexible but 5 years is a minimum. The head of department takes the responsibility of assessing and certifying the physician based on completion of the training objectives. There is a voluntary examination by Swedish Society of Obstetrics and Gynaecology. The university hospitals accept those who combine specialist training with research. The research programme has to be approved by the faculty and should lead to a doctoral degree by a thesis. Those who are trained in a non-university hospital are virtually guaranteed continued employment while those from the university hospitals have to apply for a position. There are many applicants to each post and they are judged by an internal and external panel based on the attributes of research, teaching and administrative skills. PMID- 10718723 TI - Academic obstetrics and gynaecology in the UK. AB - An investigation into the current structure of academic obstetrics and gynaecology was performed by circular to 33 departments in the UK. All departments responded. Of the 217 clinical academic staff, 32% were funded by National Health Service trusts and charities. Between 1 January 1995 and 1 January 1997, 84 posts were advertised eliciting a total of 302 applicants. Of the 73 staff that resigned over the same period, only 11 remained in academic obstetrics and gynaecology. The most commonly listed concern affecting career choice was listed as pressures from research and clinical practice. PMID- 10718724 TI - Training of academic staff in obstetrics and gynaecology in the USA. AB - As we enter a new century and millennium, departments of obstetrics and gynaecology in American medical schools face a number of challenges. Of primary concern among these is the relative dearth of physician-scientists to explore and connect the basic mechanisms and clinical aspects of the diseases of women. This report reviews some aspects of three programmes designed to educate such investigators: The Reproductive Scientist Development Program, The American Gynecological and Obstetrical Society--American Association of Obstetricians and Gynecologists Foundation Fellowship Program and the Women's Reproductive Health Research Career Development Centers. Based on a decade of experience with the first two of these programmes, the prospects for the relatively new third programme are promising. Nonetheless, the need for and opportunities for obstetrician-gynaecologist reproductive scientists far exceed the supply. PMID- 10718725 TI - Subspecialty training and academic careers. AB - Subspecialty training in obstetrics and gynaecology in the UK was formulated following a report published in 1982 by a working party established by the Royal College of Obstetricians and Gynaecologists. One of the aims of subspecialization was to improve knowledge, practice, teaching and research. The three major subspecialties were maternal-fetal medicine, reproductive medicine and gynaecological oncology. The two minor subspecialties were urogynaecology and community gynaecology. Some of the requirements for training centres are adequate clinical workload, a programme which embodies teaching, training, research and audit, adequate staffing to support training; collaboration with related disciplines and a programme director responsible for supervision and training. Each programme has a well-defined syllabus. The recruitment is based on expected needs. A minimum 7 years is needed to complete the general and subspecialty training. There is a strong academic input in this training and it includes structured research training. Many of the individuals who complete the subspecialty training will take on national responsibilities and hence their clinical and research training is of the highest quality. PMID- 10718726 TI - Academic obstetrics and gynaecology in the 21st century. AB - The future of academic departments of obstetrics and gynaecology will be determined by a number of sometimes conflicting forces. On the present funding model, departments may become entirely research centred with the major undergraduate teaching being provided by the National Health Service. Such departments could be run by non-clinical staff. An alternative scenario may be teaching-only departments with minimal research commitment. A third option is the integration of clinical research, teaching and practice in a university hospital structure where the head of department is the head of service. This model has proven to be very successful but is an expensive option as it functions well only with high funding and staffing levels. PMID- 10718727 TI - Lifetime Reproductive Success and Heritability in Nature. AB - The observation that traits closely related to fitness ("fitness traits") have lower heritabilities than traits more distantly associated with fitness has traditionally been framed in terms of Fisher's fundamental theorem of natural selection-fitness traits are expected to have low levels of additive genetic variance due to rapid fixation of alleles conferring highest fitness. Subsequent treatments have challenged this view by pointing out that high environmental and nonadditive genetic contributions to phenotypic variation may also explain the low heritability of fitness traits. Analysis of a large data set from the collared flycatcher Ficedula albicollis confirmed a previous finding that traits closely associated with fitness tend to have lower heritability. However, analysis of coefficients of additive genetic variation (CVA) revealed that traits closely associated with fitness had higher levels of additive genetic variation (VA) than traits more distantly associated with fitness. Hence, the negative relationship between a trait's association with fitness and its heritability was not due to lower levels of VA in fitness traits but was due to their higher residual variance. However, whether the high residual variance was mainly due to higher levels of environmental variance or due to higher levels of nonadditive genetic variance remains a challenge to be addressed by further studies. Our results are consistent with earlier suggestions that fitness-related traits may have more complex genetic architecture than traits more distantly associated with fitness. PMID- 10718728 TI - Disturbance Frequency and Community Stability in Native Tallgrass Prairie. AB - Ecological communities are spatially and temporally variable in response to a variety of biotic and abiotic forces. It is not always clear, however, if spatial and temporal variability leads to instability in communities. Instability may result from strong biotic interactions or from stochastic processes acting on small populations. I used 10-15 yr of annual data from the Konza Prairie Long Term Ecological Research site to examine whether plant, breeding bird, grasshopper, and small mammal communities in tallgrass prairie exhibit stability or directional change in response to different experimentally induced fire frequencies. Based on ordination and ANOVA, plant and grasshopper communities on annually burned sites differed significantly from plant and grasshopper communities on less frequently burned sites. Breeding birds and small mammals differed among sites as well, but these differences were not clearly related to disturbance frequency. A modified time series analysis indicated that plant communities were undergoing directional change (unstable) on all watersheds, regardless of fire frequency. Contrary to expectations, directional change was greatest on the annually burned sites and lowest on the infrequently burned sites. Unlike the plant communities, breeding bird, grasshopper, and small mammal communities were temporally stable, despite high-compositional variability from 1 yr to the next. Stability among the consumer communities within these dynamic plant communities occurs because three-dimensional vegetation structure does not change over time, despite changes in plant species composition. Evidence suggests that instability in the plant community results from strong biotic interactions among temporally persistent core species and stochastic dynamics among infrequent satellite species. Overall, community stability cannot be assessed if the pattern of temporal dynamics is unknown. Long-term empirical studies of different taxa under different disturbance regimes are needed to determine over what time frames and spatial scales communities may be stable. Such studies are essential for the development of generalities regarding the relationship between disturbance frequency and community stability in terrestrial and aquatic systems. PMID- 10718729 TI - Parental Care: The Key to Understanding Endothermy and Other Convergent Features in Birds and Mammals. AB - Birds and mammals share a number of features that are remarkably similar but that have evolved independently. One of these characters, endothermy, has been suggested to have played a cardinal role in avian and mammalian evolution. I hypothesize that it is parental care, rather than endothermy, that is the key to understanding the amazing convergence between mammals and birds. Endothermy may have arisen as a consequence of selection for parental care because endothermy enables a parent to control incubation temperature. The remarkable ability of many birds and mammals to sustain vigorous exercise may also have arisen as a consequence of selection for parental care because provisioning of offspring often requires sustained vigorous exercise. Because extensive parental care encompasses a wide range of behaviors, morphology, and physiology, it may be a key innovation that accounts for the majority of convergent avian and mammalian characters. PMID- 10718730 TI - Effect of Predator-Prey Phylogenetic Similarity on the Fitness Consequences of Predation: A Trade-off between Nutrition and Disease? AB - A largely neglected aspect of foraging behavior is whether the costs and benefits of predation vary as a function of phylogenetic (i.e., genetic) similarity between predator and prey. Prey of varying phylogenetic similarities to predators might differ in value because both the risk of pathogen transmission and the nutritional quality of prey typically decline with decreasing phylogenetic similarity between predator and prey. I experimentally evaluated this hypothesis by feeding omnivorous spadefoot toad tadpoles (Spea bombifrons, Spea multiplicata, and Scaphiopus couchii) either conspecific tadpoles or an equal mass of three different species of heterospecific prey, all of which contained naturally occurring bacteria. I also examined which prey species Spea tadpoles preferred. I found that all three species of tadpoles performed best on, and preferred to eat, prey that were of intermediate phylogenetic similarity to the predators. Prey of intermediate phylogenetic similarity may provide the greatest fitness benefits to predators because such prey balance the nutritional benefits of closely related prey with the cost of parasite transmission between closely related individuals. PMID- 10718731 TI - Using the Past to Predict the Present: Confidence Intervals for Regression Equations in Phylogenetic Comparative Methods. AB - Two phylogenetic comparative methods, independent contrasts and generalized least squares models, can be used to determine the statistical relationship between two or more traits. We show that the two approaches are functionally identical and that either can be used to make statistical inferences about values at internal nodes of a phylogenetic tree (hypothetical ancestors), to estimate relationships between characters, and to predict values for unmeasured species. Regression equations derived from independent contrasts can be placed back onto the original data space, including computation of both confidence intervals and prediction intervals for new observations. Predictions for unmeasured species (including extinct forms) can be made increasingly accurate and precise as the specificity of their placement on a phylogenetic tree increases, which can greatly increase statistical power to detect, for example, deviation of a single species from an allometric prediction. We reexamine published data for basal metabolic rates (BMR) of birds and show that conventional and phylogenetic allometric equations differ significantly. In new results, we show that, as compared with nonpasserines, passerines exhibit a lower rate of evolution in both body mass and mass-corrected BMR; passerines also have significantly smaller body masses than their sister clade. These differences may justify separate, clade-specific allometric equations for prediction of avian basal metabolic rates. PMID- 10718732 TI - A Transactional Theory of Within-Group Conflict. AB - Transactional models of social evolution emphasize that dominant members of the society can be favored to donate parcels of reproduction to subordinate members in return for cooperation. I construct a formal theory of intragroup conflict within the framework of transactional models by determining the maximum extent to which colony members can be selfish without destabilizing the group. The difference between the maximum value of the subordinate's fraction of group reproduction that the dominant can tolerate before ejecting the subordinate and the minimum value required by the subordinate to stay and cooperate peacefully in the group defines the "window of selfishness," which in turn predicts the frequency of within-group conflict. The window of selfishness tends to increase with increasing group reproductive output, increasingly harsh ecological constraints on solitary breeding, and, counterintuitively, increasing relatedness between subordinate and dominant. Increasing fighting ability of the subordinate can either widen or narrow the window of selfishness, the latter being most likely when ecological constraints on group living are strong. Although increasing relatedness is predicted to increase the rate of within-group aggression, the mean intensity of an aggressive act should decline, as predicted by the general theory of honest signaling between relatives and the tug-of-war models of within-group selfishness. In the bidding game, in which multiple dominants bid for the services of a subordinate, the window of selfishness is predicted to have zero width. A zero-width window of selfishness and low conflict also are predicted for saturated N-person groups, that is, groups whose total output is a concave function of group size and in which the dominant is not favored to admit additional subordinates. The model's predictions are compared to empirical evidence and to predictions of alternative models of intragroup aggression, including the value-aggression model and the pure tug-of-war model. PMID- 10718733 TI - Inbreeding Depression and Genetic Rescue in a Plant Metapopulation. AB - While migration of individuals has been shown to increase the persistence of small isolated populations through a process known as the "rescue effect," the demographic effects that pollen-mediated gene flow may have in plant populations are not known empirically. This study investigates the role that inbreeding depression plays in newly colonized populations of a common, dioecious, weedy species, Silene alba. Experimental greenhouse studies presented here show that S. alba displays high levels of inbreeding depression (expressed as lowered germination success) in progeny produced with inbreeding coefficients of 0.125 (half-sib mated), 0.250 (full-sib mated), and 0.375 (second-generation sib mated). In addition, it is shown that the degree of inbreeding depression in 12 natural colonies varies with the degree of isolation from other established populations. Significantly, data from experimental populations showed that gene flow into patches comprised of full sibs was higher than those observed into patches comprised of unrelated individuals and may serve to mitigate the effects of inbreeding depression. It is suggested that population connectivity through pollen-mediated gene flow may have substantial effects on the persistence of isolated colonies and on the spatial structure of a metapopulation in general. PMID- 10718734 TI - The Evolution of Intraspecific Brood Parasitism in Birds and Insects. AB - Many species of birds and insects engage in intraspecific brood parasitism (IBP), when a female lays eggs in the nest of a conspecific and leaves without providing parental care. These visiting females may also act to cooperate with a primary female, staying to provide parental care. Therefore, IBP and cooperative breeding can be considered extremes on a continuum of parental care provided by a secondary female. When a secondary female abandons a nest, she creates an asymmetry in parental care between herself and the host. While models of asymmetry in reproductive allocation have focused directly on relatedness between females, we lack an appropriate theoretical framework that addresses the effects of relatedness on parental care asymmetry. Here, I present an evolutionarily stable strategy (ESS) model that predicts the conditions under which IBP is favored over cooperation and solitary breeding. Intraspecific brood parasitism is less likely to evolve (relative to cooperation and solitary breeding) as the relatedness between a host and parasite increases. It can evolve, however, if parasites achieve a high overall fecundity relative to solitary females. Constraints on solitary breeding can further promote IBP under some circumstances. Cooperation is favored when relatedness is high and reproductive skew is low. This model makes several predictions regarding the conditions under which IBP may evolve, motivating a variety of experimental approaches. PMID- 10718735 TI - Power Struggles, Dominance Testing, and Reproductive Skew. AB - Models of reproductive skew are concerned with the partitioning of reproduction between dominant and subordinate members of a group. In an interesting extension of these models, Reeve and Ratnieks briefly considered whether it might benefit subordinates to engage in aggressive behavior to test the fighting ability of a dominant. Their analysis suggested that such testing should be more probable in groups that feature high skew and, hence, perhaps among closer relatives (because high relatedness favors high skew). Here we explore in more detail the possibility of dominance testing. Three models that differ in the outcome of fights over dominance are presented: in the first model, the loser of the challenge is killed; in the second model, the loser is evicted from the nest; and, in the third model, the loser becomes (or remains) subordinate. In each case we consider the independent effects of the parameters that determine skew (namely, relatedness, group productivity, and ecological constraints) on the predicted level of dominance testing. We then construct an amalgamated model to examine situations where fights may lead to any one of the three outcomes. Our analysis reveals that, in the majority of cases, higher relatedness will in fact lead to lower levels of aggression. Moreover, dominance testing need not be associated with high skew. Rather, the relationship between skew and dominance testing will depend on which factor (relatedness, group productivity, or level of ecological constraints) is principally responsible for variation in the distribution of reproduction. PMID- 10718736 TI - Voltage-dependent Ca2+ release from the SR of feline ventricular myocytes is explained by Ca2+-induced Ca2+ release. AB - 1. Direct voltage-gated (voltage-dependent Ca2+ release, VDCR) and Ca2+ influx gated (Ca2+-induced Ca2+ release, CICR) sarcoplasmic reticulum (SR) Ca2+ release were studied in feline ventricular myocytes. The voltage-contraction relationship predicted by the VDCR hypothesis is sigmoidal with large contractions at potentials near the Ca2+ equilibrium potential (ECa). The relationship predicted by the CICR hypothesis is bell-shaped with no contraction at ECa. 2. The voltage dependence of contraction was measured in ventricular myocytes at physiological temperature (37 C), resting membrane potential and physiological [K+]. Experiments were performed with cyclic adenosine 3',5'-monophosphate (cAMP) in the pipette or in the presence of the beta-adrenergic agonist isoproterenol (isoprenaline; ISO). 3. The voltage-contraction relationship was bell-shaped in Na+-free solutions (to eliminate the Na+ current and Na+-Ca2+ exchange, NCX) but the relationship was broader than the L-type Ca2+ current (ICa,L)-voltage relationship. 4. Contractions induced with voltage steps from normal resting potentials to -40 mV are thought to represent VDCR rather than CICR. We found that cAMP and ISO shifted the voltage dependence of ICa,L activation to more negative potentials so that ICa,L was always present with steps to -40 mV. ICa,L at -40 mV inactivated when the holding potential was decreased (VL = -57.8 +/- 0.49 mV). 5. ISO increased inward current, SR Ca2+ load and contraction in physiological [Na+] and a broad bell-shaped voltage-contraction relationship was observed. Inhibition of reverse-mode NCX, decreasing ICa,L and decreasing SR Ca2+ loading all decreased contractions at strongly positive potentials near ECa. 6. The voltage-contraction relationship in 200 microM cadmium (Cd2+) was bell shaped, supporting a role of ICa,L rather than VDCR. 7. All results could be accounted for by the CICR hypothesis, and many results exclude the VDCR hypothesis. PMID- 10718737 TI - Capacitative Ca2+ entry is graded with degree of intracellular Ca2+ store depletion in bovine vascular endothelial cells. AB - 1. In endothelial cells, release of Ca2+ from endoplasmic reticulum (ER) Ca2+ stores activates Ca2+ influx via the capacitative Ca2+ entry (CCE) pathway. In cultured bovine pulmonary artery endothelial cells, we investigated the relationship between intracellular Ca2+ store load and CCE activity, as well as the kinetics of CCE activation and deactivation, by simultaneously measuring changes in [Ca2+]i and unidirectional manganese (Mn2+) entry through the CCE pathway. 2. Submaximal concentrations of ATP caused quantal release of Ca2+ from the ER, resulting in a dose-dependent depletion of Ca2+ stores and acceleration of Mn2+ entry. Mn2+ entry rate, as a measure of CCE activity, was graded with the amount of released Ca2+. Maximal activation of CCE did not require complete store depletion. 3. Slow depletion of the ER by exposure to the ER Ca2+ pump inhibitor cyclopiazonic acid resulted in a delayed activation of CCE, revealing a temporal dissociation between release of Ca2+ from intracellular stores and activation of CCE. 4. During [Ca2+]i oscillations, at frequencies higher than 0.5 spikes min-1, each Ca2+ spike resulted in a progressive acceleration of CCE without leading to oscillations of Ca2+ entry. In contrast, low frequency [Ca2+]i oscillations were paralleled by transient CCE that was activated and deactivated with each Ca2+ spike, resulting in an oscillatory pattern of Ca2+ entry. 5. It is concluded that CCE is a rapidly activating process which is graded with store depletion and becomes fully activated before complete depletion. The duration of CCE activation correlates with the degree of store depletion and the time that is required to refill depleted stores. Overall, a mechanism of graded CCE prevents exhaustion of intracellular Ca2+ reserves and provides an efficient way to respond to variable degrees of intracellular store depletion. PMID- 10718738 TI - Activation of the cAMP-protein kinase A pathway facilitates Na+ translocation by the Na+-K+ pump in guinea-pig ventricular myocytes. AB - 1. The effects of the adenylyl cyclase activator forskolin on steady-state and transient currents generated by the Na+-K+ pump were studied in guinea-pig ventricular myocytes by means of whole-cell voltage clamp at 30 C. 2. In external solution containing 144 mM Na+ (Na+o) and 10 mM K+ (K+o), steady-state Na+-K+ pump current (Ip) activated by 5 mM pipette Na+ (Na+pip) at -20 mV was reversibly augmented by forskolin (4 microM) to 133 +/- 4 % of the control current (n = 15). The forskolin analogue 1, 9-dideoxyforskolin (10 microM), which does not activate adenylyl cyclases, did not increase Ip (n = 2). Application of the protein kinase A (PKA) inhibitor H-89 (10 microM) in the continued presence of forskolin reversed the forskolin-induced elevation of Ip (n = 3). 3. The forskolin effect on Ip persisted in the presence of 50 mM Na+pip which ensured that the internal Na+-binding sites of the Na+-K+ pump were nearly saturated. Under these conditions, the drug increased Ip to 142 +/- 3 % of the control Ip when the pipette free Ca2+ concentration ([Ca2+]pip) was 0.013 nM (n = 5) and to 138 +/- 4 % of the control Ip when free [Ca2+]pip was 15 nM (n = 9). 4. In Na+-free external solution, Ip activated by 50 mM Na+pip and 1.5 mM K+o was likewise increased by forskolin but to a lesser extent than in Na+-containing medium (116 +/- 3 % of control, n = 10). 5. In order to investigate exclusively partial reactions in the Na+ limb of the pump cycle, transient pump currents under conditions of electroneutral Na+-Na+ exchange were studied. Transient pump currents elicited by voltage jumps displayed an initial peak and then decayed monoexponentially. Moved charge (Q) and the rate constant of current decay varied with membrane potential (V). The Q-V relationship followed a Boltzmann distribution characterized by the midpoint voltage (V0.5) and the maximum amount of movable charge (DeltaQmax). Forskolin (2-10 microM) shifted V0.5 to more negative values while DeltaQmax was not affected (n = 11). The effects of forskolin on transient pump currents were mimicked by 8-bromo-cAMP (500 microM; n = 2) and abolished by a peptide inhibitor of PKA (PKI, 10 microM; n = 5). 6. We conclude that activation of the cAMP-PKA pathway in guinea-pig ventricular myocytes increases Na+-K+ pump current at least in part by modulating partial reactions in the Na+ limb of the pump cycle. Under physiological conditions, the observed stimulation of the cardiac Na+-K+ pump may serve to shorten the action potential duration and to counteract the increased passive sarcolemmal Na+ and K+ fluxes during sympathetic stimulation of the heart. PMID- 10718739 TI - Regulation of transient Na+ conductance by intra- and extracellular K+ in the human delayed rectifier K+ channel Kv1.5. AB - 1. Significant Na+ conductance has been described in only a few native and cloned K+ channels, but has been used to characterize inactivation and K+ binding within the permeation pathway, and to refine models of K+ flux through multi-ion pores. Here we use Na+ permeation of the delayed rectifier K+ channel Kv1.5 to study extra- and intracellular K+ (K+o and K+i, respectively) regulation of conductance and inactivation, using whole-cell recording from human embryonic kidney (HEK) 293 cells. 2. Kv1.5 Na+ currents in the absence of K+o and K+i were confirmed by: (i) resistance of outward Na+ currents to dialysis by K+-free solutions; (ii) tail current reversal potential changes with Na+o with a slope of 55.8 mV per decade; (iii) block by 4-aminopyridine (50 % at 50 microM), and resistance to Cl- channel inhibition. 3. Na+ currents were transient followed by a small sustained current. An envelope test confirmed that activated Kv1.5 channels conducted Na+, and that rapid current decay reflected C-type inactivation. Sustained currents ( approximately 13 % of peak) represented Na+ flux through inactivated Kv1.5 channels. 4. K+o could modulate the maximum available Na+ conductance in the stable cell line while channels were closed. Before the first pulse of a train, increasing K+o concentration increased the subsequent Na+ conductance from approximately 15 (0 mM K+o) to 30 nS (5 mM K+o), with a Kd of 23 microM. Repeated low rate depolarizations in Na+i/Na+o solutions induced a use-dependent loss of Kv1.5 channel Na+ conductance, distinct from that caused by C-type inactivation. K+o binding that sensed little of the electric field could prevent this secondary loss of available Kv1.5 channels with a Kd of 230 microM. These two effects on conductance were both voltage independent, and had no effect on channel inactivation rate. 5. K+o concentrations >= 0.3 mM slowed the inactivation rate in a strongly voltage-dependent manner. This suggested it could compete for binding at a K+ site or sites deeper in the pore, as well as restoring the Na+ conductance. K+i was able to modulate the inactivation rate but was unable to affect conductance. 6. Mutation of arginine 487 in the outer pore region of the channel to valine (R487V) greatly reduced C-type inactivation in Na+ solutions, caused loss of channel use dependence, and prevented any conductance increase upon the addition of 0.1 mM K+o. Our results confirm the existence of a high affinity binding site at the selectivity filter that regulates inactivation, and also reveals the presence of at least one additional high affinity outer mouth site that predominantly regulates conductance of resting channels, and protects channels activated by depolarization when they conduct Na+. PMID- 10718740 TI - The sustained inward current and inward rectifier K+ current in pacemaker cells dissociated from rat sinoatrial node. AB - 1. Myocytes were dissociated from the sinoatrial (SA) node of rat heart using a new enzymatic dissociation technique. Only a small number of isolated SA node myocytes showed regular rhythmic contractions and spontaneous action potentials, and these were used in the present study. 2. The spontaneous action potential was resistant to TTX, and the action potential parameters were similar to those of rabbit and guinea-pig pacemaker cells. Major time- and voltage-dependent currents were the delayed rectifier K+ current IKr, the L-type Ca2+ current ICa,L and the sodium current INa. The hyperpolarization-activated cation current (If) was recorded from approximately 50 % of the cells with hyperpolarization beyond -90 mV. 3. The instantaneous current jump at the onset of a hyperpolarizing pulse showed inward rectification and was largely blocked by Ba2+. This Ba2+-sensitive current corresponded well to the inward rectifier K+ current (IK1), although it was much smaller in amplitude than in the ventricle. 4. A sustained inward current was activated on depolarization from -80 mV to the voltage range of slow diastolic depolarization. The current was blocked by nicardipine, enlarged by isoprenaline and was insensitive to removal of external Ca2+. These characteristics were similar to the sustained inward current, Ist, previously described in the rabbit and guinea-pig SA node cells. 5. The role of Ist was considered by constructing empirical equations, which were applied to the experimental record of the action potential. It is demonstrated that the voltage dependent activation of Ist constitutes a positive feedback loop with the depolarization of the membrane. PMID- 10718741 TI - Stretch-activated and background non-selective cation channels in rat atrial myocytes. AB - 1. Stretch-activated channels (SACs) were studied in isolated rat atrial myocytes using the whole-cell and single-channel patch clamp techniques. Longitudinal stretch was applied by using two patch electrodes. 2. In current clamp configuration, mechanical stretch of 20 % of resting cell length depolarised the resting membrane potential (RMP) from -63.6 +/- 0.58 mV (n = 19) to -54.6 +/- 2.4 mV (n = 13) and prolonged the action potential duration (APD) by 32.2 +/- 8.8 ms (n = 7). Depolarisation, if strong enough, triggered spontaneous APs. In the voltage clamp configuration, stretch increased membrane conductance in a progressive manner. The current-voltage (I-V ) relationship of the stretch activated current (ISAC) was linear and reversed at -6.1 +/- 3.7 mV (n = 7). 3. The inward component of ISAC was abolished by the replacement of Na+ with NMDG+, but ISAC was hardly altered by the Cl- channel blocker DIDS or removal of external Cl-. The permeability ratio for various cations (PCs:PNa:PLi = 1.05:1:0.98) indicated that the SAC current was a non-selective cation current (ISAC,NC). The background current was also found to be non-selective to cations (INSC,b); the permeability ratio (PCs:PNa:PLi = 1.49:1:0.70) was different from that of ISAC,NC. 4. Gadolinium (Gd3+) acted on INSC,b and ISAC,NC differently. Gd3+ inhibited INSC,b in a concentration-dependent manner with an IC50 value of 46.2 +/- 0.8 microM (n = 5). Consistent with this effect, Gd3+ hyperpolarised the resting membrane potential (-71.1 +/- 0.26 mV, n = 9). In the presence of Gd3+ (0.1 mM), stretch still induced ISAC,NC and diastolic depolarisation. 5. Single channel activities were recorded in isotonic Na+ and Cs+ solutions using the inside-out configuration. In NMDG+ solution, outward currents were abolished. Gd3+ (100 microM) strongly inhibited channel opening both from the inside and outside. In the presence of Gd3+ (100 microM) in the pipette solution, an increase in pipette pressure induced an increase in channel opening (21.27 +/- 0.24 pS; n = 7), which was distinct from background activity. 6. We concluded from the above results that longitudinal stret in rat atrial myocytes induces the activation of non-selective cation channels that can be distinguished from background channels by their different electrophysiology and pharmacology. PMID- 10718742 TI - Distinct synaptic and extrasynaptic NMDA receptors identified in dorsal horn neurones of the adult rat spinal cord. AB - 1. Using patch-clamp recordings, properties of single-channel and synaptic currents mediated by N-methyl-D-aspartate receptors (NMDARs) were examin ed in substantia gelatinosa (SG) neurones of adult rat spinal cord slices. 2. In somatic outside-out patches, high- and low-conductance NMDAR channels were present. The low-conductance channels exhibited asymmetrical transitions between the main (44 pS) and subconductance (19 pS) levels, suggesting that they arise from NR2D subunit-containing receptors. The high-conductance channels (main conductance, 57 pS) were blocked by ifenprodil, an NR2B subunit selective blocker. 3. Ifenprodil had no effect on NMDA-EPSCs. The double-exponential decay time course and the apparent Kd for Mg2+ of NMDA-EPSCs suggested the expression of NR2A subunit-containing receptors at the synapse. 4. These results indicate that different NMDAR subtypes are expressed in subsynaptic and extrasynaptic regions of adult SG neurones, which may have differential roles in nociception. PMID- 10718743 TI - GABA release from mouse axonal growth cones. AB - 1. Using developing hypothalamic neurons from transgenic mice that express high levels of green fluorescent protein in growing axons, and an outside-out patch from mature neuronal membranes that contain neurotransmitter receptors as a sensitive detector, we found that GABA is released by a vesicular mechanism from the growth cones of developing axons prior to synapse formation. 2. A low level of GABA release occurs spontaneously from the growth cone, and this is substantially increased by evoked action potentials. 3. Neurotransmitters such as acetylcholine can enhance protein kinase C (PKC) activity even prior to synapse formation; PKC activation caused a substantial increase in spontaneous GABA release from the growth cone, probably acting at the axon terminal. 4. These data indicate that GABA is secreted from axons during a stage of neuronal development when GABA is excitatory, and that neuromodulators could alter GABA release from the growing axon, potentially enabling other developing neurons of different transmitter phenotype to modulate the early actions of GABA. PMID- 10718744 TI - Opposite changes in synaptic activity of organotypic rat spinal cord cultures after chronic block of AMPA/kainate or glycine and GABAA receptors. AB - 1. The well-developed cytoarchitecture of rat organotypic spinal cord culture makes it a suitable model to explore how persistent suppression of certain synaptic inputs might be compensated by increased synaptic efficacy (homeostatic plasticity). 2. Spontaneous or electrically evoked synaptic transmission of patch clamped ventral horn interneurons was studied in control solution after blocking, for the second week in culture, AMPA/kainate receptors with CNQX or glycine and GABAA receptors with strychnine and bicuculline, or indiscriminately removing inputs with tetrodotoxin (TTX). 3. In untreated cells, spontaneous postsynaptic currents (PSCs) had fast (tau < 5 ms) or slow (tau > 10 ms) decay. A similar separation was observed when recording miniature currents (mPSCs). Slow decay PSCs were suppressed by strychnine plus bicuculline while fast decay events were eliminated by CNQX. 4. After chronic CNQX treatment the frequency of spontaneous, fast PSCs (of larger amplitude) or mPSCs was almost doubled with respect to control. These events were blocked by acutely applied CNQX, which unmasked slow PSCs. 5. After chronic TTX treatment neither the frequency nor the amplitude of spontaneous events was changed. 6. After chronic strychnine and bicuculline treatment the frequency and amplitude of all PSCs was decreased in most cells. mPSCs were also decreased in frequency. Spontaneous or electrically evoked currents acquired a larger component mediated by NMDA receptor activity. 7. The developing spinal network thus operated distinct homeostatic processes which led to strong enhancement in glutamatergic transmission after CNQX block or to broad downregulation of synaptic activity following chronic exposure to strychnine and bicuculline. PMID- 10718745 TI - Kainate receptor-mediated presynaptic inhibition at the mouse hippocampal mossy fibre synapse. AB - 1. The presynaptic action of kainate (KA) receptor activation at the mossy fibre CA3 synapse was examined using fluorescence measurement of presynaptic Ca2+ influx as well as electrophysiological recordings in mouse hippocampal slices. 2. Bath application of a low concentration (0.2 microM) of KA reversibly increased the amplitude of presynaptic volley evoked by stimulation of mossy fibres to 146 +/- 6 % of control (n = 6), whereas it reduced the field excitatory postsynaptic potential (EPSPs) to 30 +/- 4 %. 3. The potentiating effect of KA on the presynaptic volleys was also observed in Ca2+-free solution, and was partly antagonized by (2S, 4R)-4-methylglutamic acid (SYM 2081, 1 microM), which selectively desensitizes KA receptors. 4. The antidromic population spike of dentate granule cells evoked by stimulation of mossy fibres was increased by application of 0.2 microM KA to 160 +/- 10 % of control (n = 6). Whole-cell current-clamp recordings revealed that the stimulus threshold for generating antidromic spikes recorded from a single granule cell was lowered by KA application. 5. Application of KA (0.2 microM) suppressed presynaptic Ca2+ influx to 78 +/- 4 % of control (n = 6), whereas the amplitude of the presynaptic volley was increased. 6. KA at 0.2 microM reversibly suppressed excitatory postsynaptic currents (EPSCs) evoked by mossy fibre simulation to 38 +/- 9 % of control (n = 5). 7. These results suggest that KA receptor activation enhances the excitability of mossy fibres, probably via axonal depolarization, and reduces action potential-induced Ca2+ influx, thereby inhibiting mossy fibre EPSCs presynaptically. This novel presynaptic inhibitory action of KA at the mossy fibre-CA3 synapse may regulate the excitability of highly interconnected CA3 networks. PMID- 10718746 TI - Contrasting molecular composition and channel properties of AMPA receptors on chick auditory and brainstem motor neurons. AB - 1. Neurons in the brainstem auditory pathway exhibit a number of specializations for transmitting signals reliably at high rates, notably synaptic AMPA receptors with very rapid kinetics. Previous work has not revealed a common structural pattern shared by the AMPA receptors of auditory neurons that could account for their distinct functional properties. 2. We have used whole-cell patch-clamp recordings, mRNA analysis, immunofluorescence, Western blots and agonist-evoked cobalt uptake to compare AMPA receptors on the first-, second- and third-order neurons in the chick ascending auditory pathway with those on brainstem motor neurons of the glossopharyngeal/vagal nucleus, which have been shown to have very slow desensitization kinetics. 3. The results indicate that the AMPA receptors of the cochlear ganglion, nucleus magnocellularis and nucleus laminaris share a number of structural and functional properties that distinguish them from the AMPA receptors of brainstem motor neurons, namely a lower relative abundance of glutamate receptor (GluR)2 transcript and much lower levels of GluR2 immunoreactivity, higher relative levels of GluR3 flop and GluR4 flop, lower relative abundance of the C-terminal splice variants GluR4c and 4d, less R/G editing of GluR2 and 3, greater permeability to calcium, predominantly inwardly rectifying I-V relationships, and greater susceptibility to block by Joro spider toxin. 4. We conclude that the AMPA receptors of auditory neurons acquire rapid kinetics from their high content of GluR3 flop and GluR4 flop subunits and their high permeability to Ca2+ from selective post-transcriptional suppression of GluR2 expression. PMID- 10718747 TI - One-way cross-desensitization between P2X purinoceptors and vanilloid receptors in adult rat dorsal root ganglion neurones. AB - 1. Capsaicin and ATP can activate ligand-gated cation channels in nociceptive rat dorsal root ganglion (DRG) neurones. We have studied cross-desensitization between these two agents in rat isolated DRG neurones using the whole-cell voltage-clamp technique. 2. ATP (10 microM) activated an inward current in DRG neurones at a holding potential of -60 mV. ATP evoked 'fast' responses that underwent rapid activation and desensitization, 'slow' responses that activated and desensitized more slowly, or responses that displayed a mixture of these two characteristics. The time course of the response to ATP was not related obviously to capsaicin sensitivity. 3. Prior application of capsaicin (0.5 microM) increased the proportion of cells displaying only fast responses to ATP (10 microM) suggesting that cross-desensitization had occurred between capsaicin and the slow component of the ATP response. Prior desensitization to ATP had no apparent effect on the inward current response to capsaicin (0.5 microM). 4. Cross-desensitization between capsaicin and ATP was Ca2+ dependent. 5. Changing the membrane holding potential (Vh) to +40 mV for brief period before applying ATP at -60 mV had a similar effect to capsaicin, i.e. the proportion of cells displaying only fast responses to ATP was increased significantly. This effect of depolarization was not Ca2+ dependent. 6. The heterogenity of responses to ATP is probably due to co-expression of homomeric P2X3 receptors and heteromeric receptors comprising P2X3 subunits with other P2X subunits. We propose that the change in time course of the ATP response produced by prior desensitization to capsaicin is due to selective cross-desensitization with the heteromeric P2X receptors. PMID- 10718748 TI - Mutation of histidine 286 of the human P2X4 purinoceptor removes extracellular pH sensitivity. AB - 1. Effects of external pH on the human P2X4 purinoceptor, an ATP-activated ion channel, were studied using the Xenopus oocyte expression system. 2. Changing the external pH from 7.4 to 6.5 significantly reduced, whilst an increase to pH 8 enhanced, maximum ATP-activated current amplitude, without changing the current- voltage relationship of the ATP-activated current. 3. Diethyl pyrocarbonate (DEPC; 10 mM) treatment of P2X4-injected oocytes had no effect on the pH sensitivity of the ATP-activated current. 4. Site-directed mutagenesis of histidine 286 (H286) to alanine completely abolished the pH sensitivity of the P2X4 receptor at all agonist concentrations. ATP potency showed a small (fourfold) leftward shift. Mutagenesis of the other three histidines present in the P2X4 sequence had no effect on pH sensitivity. 5. The results show that pH modulation of P2X4 in the pathophysiological range is mediated by protonation of H286. This provides direct confirmation that pH sensitivity resides in the P2X4 channel protein rather than the agonist species. PMID- 10718749 TI - Intracellular cyclic AMP inhibits native and recombinant volume-regulated chloride channels from mammalian heart. AB - 1. ClC-3 encodes a volume-regulated Cl- channel (ICl,vol) in heart. We studied the regulation of native and recombinant cardiac ICl,vol by intracellular cyclic AMP (cAMPi). 2. Symmetrical high Cl- concentrations were used to effectively separate outwardly rectifying ICl,vol from other non-rectifying Cl- currents, such as the cystic fibrosis transmembrane conductance regulator (CFTR) and Ca2+ activated Cl- currents (ICl,CFTR and ICl,Ca, respectively), which are concomitantly expressed in cardiac myocytes. 3. 8-Bromo-cyclic AMP (8-Br-cAMP) significantly inhibited ICl,vol in most guinea-pig atrial myocytes. In approximately 30 % of the atrial myocytes examined, 8-Br-cAMP increased macroscopic Cl- currents. However, the 8-Br-cAMP-stimulated difference currents exhibited a linear current-voltage (I-V ) relation, consistent with activation of ICl,CFTR, not ICl,vol. 4. In canine atrial myocytes, isoprenaline (1 microM) consistently reduced ICl,vol in Ca2+-free hypotonic bath solutions with strong intracellular Ca2+ (Ca2+i) buffering. In Ca2+-containing hypotonic bath solutions with weak Ca2+i buffering, however, isoprenaline increased net macroscopic Cl- currents. Isoprenaline-stimulated difference currents were not outwardly rectifying, consistent with activation of ICl,Ca, not ICl, vol. 5. In NIH/3T3 cells transfected with gpClC-3 (the gene encoding ICl,vol), 8-Br-cAMP consistently inhibited ICl,ClC-3. These effects were prevented by a protein kinase A (PKA) inhibitor, KT5720, or by mutation of a single consensus protein kinase C (PKC) phosphorylation site (S51A) on the N-terminus of ClC-3, which also mediates PKC inhibition of ICl,ClC-3. 6. We conclude that cAMPi causes inhibition of ICl,vol in mammalian heart due to cross-phosphorylation of the same PKC consensus site on ClC-3 by PKA. Our results suggest that contamination of macroscopic ICl,vol by ICl,CFTR and/or ICl,Ca may account for some of the inconsistent and controversial effects of cAMPi on ICl,vol previously reported in native cardiac myocytes. PMID- 10718750 TI - Effects of hypoxia and dithionite on catecholamine release from isolated type I cells of the rat carotid body. AB - 1. Amperometric recordings were conducted to investigate the ability of hypoxia and anoxia to evoke quantal catecholamine secretion from isolated type I cells of the rat carotid body. 2. Hypoxia (PO2 8-14 mmHg) consistently failed to evoke catecholamine secretion from type I cells, when cells were perfused either at room temperature (21-24 C) or at 35-37 C, and regardless of whether Hepes- or HCO3-/CO2-buffered solutions were used. 3. Elevating extracellular [K+] caused concentration-dependent secretion from individual type I cells, with a threshold concentration of approximately 25 mM. In the presence of this level of extracellular K+, hypoxia (PO2 8-14 mmHg) caused a marked enhancement of secretion which was fully blocked by 200 microM Cd2+, a non-specific blocker of voltage-gated Ca2+ channels. 4. Anoxia (N2-equilibrated solution containing 0.5 mM dithionite) evoked exocytosis from type I cells when extracellular [K+] was 5 mM. This secretion was completely inhibited by removal of extracellular Ca2+, but was not significantly affected by Cd2+ (200 microM), Ni2+ (2 mM), Zn2+ (1 mM) or nifedipine (2 microM). Secretion was also observed when 0.5 mM dithionite was added to air-equilibrated solutions. 5. Anoxia also evoked secretion from chemoreceptive phaeochromocytoma (PC12) cells, which was wholly Ca2+ dependent, but unaffected by Cd2+ (200 microM). 6. Our results suggest that hypoxia can evoke catecholamine secretion from isolated type I cells, but only in the presence of elevated extracellular [K+]. This may be due to the cells being relatively hyperpolarized following dissociation. In addition, we have shown that dithionite evokes catecholamine release regardless of PO2 levels, and this release is due mainly to an artefactual Ca2+ influx pathway activated in the presence of dithionite. PMID- 10718751 TI - Recurrent inhibitory circuitry in the deep layers of the rabbit superior colliculus. AB - 1. Local inhibition in the deep layers of the superior colliculus plays a crucial role in sensorimotor integration. Using intracellular and extracellular recording techniques, we studied the organization of inhibitory circuits in the deep layers of the superior colliculus in anaesthetized rabbits. 2. We identified a new cell type in the deep superior colliculus that showed a characteristic burst response to stimulation of both the predorsal bundle and optic chiasm. The response had a jittering latency and failed to follow high frequency stimuli, indicating trans synaptic (orthodromic) events. Moreover, the predorsal bundle stimulation-evoked orthodromic response could be made to collide with the response to a preceding stimulation of the optic chiasm, suggesting that burst-firing cells received excitatory inputs from the axonal collaterals of predorsal bundle-projecting cells. 3. Stimulation of the predorsal bundle could evoke an IPSP in predorsal bundle-projecting cells. The latency of the IPSP was 0.5-1.0 ms longer than the orthodromic response in burst-firing cells. Simultaneous recordings showed that the IPSP in predorsal bundle-projecting cells was preceded by a burst of extracellular spikes from burst-firing cells with short latency ( approximately 0.9 ms), indicating an inhibitory monosynaptic connection from burst-firing cells to predorsal bundle-projecting cells. 4. Burst-firing cells exhibited a prolonged depression after the predorsal bundle or optic chiasm stimulation due to an inhibitory postsynaptic potential. Latency analysis implies that burst-firing cells may form mutual inhibitory connections. 5. Together our results suggest that burst-firing cells and predorsal bundle-projecting cells form reciprocal excitatory and inhibitory connections and burst-firing cells may function as the recurrent inhibitory interneurons in the deep layers of the rabbit superior colliculus. PMID- 10718752 TI - Non-linear membrane properties of sacral sphincter motoneurones in the decerebrate cat. AB - 1. Responses to pudendal afferent stimulation and depolarizing intracellular current injection were examined in sacral sphincter motoneurones in decerebrate cats. 2. In 16 animals examined, 2-10 s trains of electrical stimulation of pudendal afferents evoked sustained sphincter motoneurone activity lasting from 5 to >50 s after stimulation. The sustained response was observed in: 11 animals in the absence of any drugs; two animals after the intravenous administration of 5 hydroxytryptophan (5-HTP; <= 20 mg kg-1); one animal in which methoxamine was perfused onto the ventral surface of the exposed spinal cord; and two animals following the administration of intravenous noradrenergic agonists. 3. Extracellular and intracellular recordings from sphincter motoneurones revealed that the persistent firing evoked by afferent stimulation could be terminated by motoneurone membrane hyperpolarization during micturition or by intracellular current injection. 4. Intracellular recordings revealed that 22/40 sphincter motoneurones examined displayed a non-linear, steep increase in the membrane potential in response to depolarizing ramp current injection. The mean voltage threshold for this non-linear membrane response was -43 +/- 3 mV. Five of the 22 cells displaying the non-linear membrane response were recorded prior to the administration of 5-HTP; 17 after the intravenous administration of 5-HTP (<= 20 mg kg-1). 5. It is concluded that sphincter motoneurones have a voltage sensitive, non-linear membrane response to depolarization that could contribute to sustained sphincter motoneurone firing during continence. PMID- 10718753 TI - Visual physiology of the lateral geniculate nucleus in two species of new world monkey: Saimiri sciureus and Aotus trivirgatis. AB - 1. Visual responses were recorded from neurones in the magnocellular and parvocellular layers of the lateral geniculate nucleus (LGN) of the thalamus in two species of New World monkeys - the diurnal squirrel monkey (Saimiri sciureus) and the nocturnal owl monkey (Aotus trivirgatis). Recording sites were reconstructed in postmortem tissue and comparisons were made between the response properties of magnocellular and parvocellular neurones. 2. Receptive fields were characterized with both white noise and drifting gratings. We found that most of the differences between magnocellular and parvocellular neurones that have been described in the macaque monkey hold for the squirrel monkey and owl monkey. In squirrel monkey and owl monkey, receptive fields of magnocellular neurones were larger than those of parvocellular neurones at similar eccentricities. Although visual responses in the owl monkey were significantly slower than in the squirrel monkey, in both species magnocellular neurones differed from parvocellular neurones in that their responses (1) had higher contrast gains, (2) tended to peak at higher temporal frequencies (but with considerable overlap), (3) had shorter response latencies, and (4) were more transient. 3. The strength of a neurone's receptive-field surround was assessed by comparing neuronal responses to gratings of optimal spatial frequency with responses to gratings of low spatial frequency. Using this approach, receptive-field surrounds were found to be equally strong on average for magnocellular and parvocellular neurones. 4. Spatial summation, as measured by a null test, was linear for all magnocellular and parvocellular cells tested; that is, Y cells were not observed in either species. Finally, most magnocellular neurones showed a contrast gain control mechanism, although this was not seen for parvocellular neurones. PMID- 10718754 TI - Surround inhibition of mammalian AII amacrine cells is generated in the proximal retina. AB - 1. Intracellular recordings were obtained from neurons in the superfused retina eyecup preparation of the rabbit under dark-adapted conditions. Neurotransmitter agonists and antagonists were applied exogenously via the superfusate to dissect the synaptic pathways pharmacologically and thereby determine those pathways responsible for the generation of the on-centre/off-surround receptive fields of AII amacrine cells. 2. Application of the metabotropic glutamate receptor agonist, APB, reversibly blocked both the on-centre and off-surround responses of AII cells. These data were consistent with the idea that both the centre- and surround-mediated responses are derived from inputs from the presynaptic rod bipolar cells. 3. Whereas rod bipolar cells showed on-receptive fields approximately 100 microm across, we found no evidence for an antagonistic off surround response using light stimuli which effectively elicited the off surrounds of AII amacrine cells. These results indicated that the surrounds of AII cells are not derived from rod bipolar cell inputs. 4. Application of the ionotropic glutamate receptor antagonists CNQX or DNQX enhanced the on-centre responses of AII cells but attenuated the off-surround responses. These data indicated that the centre- and surround-mediated responses could not both be derived from signals crossing the rod bipolar-to-AII cell synapse. 5. Application of the glycine antagonist, strychnine, had only minor and variable effects on AII cell responses. However, the GABA antagonists picrotoxin and bicuculline enhanced the on-centre response but attenuated or completely blocked the off-surround response of AII cells. The GABA antagonists had no effect on the responses of horizontal cells indicating that their effects on AII cell responses reflected actions on inner retinal circuitry rather than feedback circuitry in the outer plexiform layer. 6. Application of the voltage-gated sodium channel blocker TTX enhanced the on-centre responses of AII cells but attenuated or abolished their off-surround responses. 7. Taken together, our results suggest that the on-centre responses of AII cells result from the major excitatory drive from rod bipolar cells. However, the surround receptive fields of AII cells appear to be generated by lateral, inhibitory signals derived from neighbouring GABAergic, on-centre amacrine cells. A model is presented whereby the S1 amacrine cells produce the surround receptive fields of AII amacrine cells via inhibitory, feedback circuitry to the axon terminals of rod bipolar cells. PMID- 10718755 TI - Changes in the mechanisms involved in uterine contractions during pregnancy in guinea-pigs. AB - 1. The mechanisms involved in contraction in guinea-pig myometrium were compared at mid- and late pregnancy. Tension was recorded simultaneously with either membrane potential or cytoplasmic calcium ([Ca2+]i) in strips exposed briefly to prostaglandin F2alpha (PGF). 2. PGF-induced increases in tension were underpinned by action potentials followed by sustained depolarization and biphasic increases in [Ca2+]i at mid- (peak, 879 +/- 199 nM; sustained, 298 +/- 35 nM, n = 11) and late pregnancy (peak, 989 +/- 302 nM; sustained 178 +/- 33 nM, n = 8). 3. At mid- and late pregnancy, nifedipine (10-6 M) reduced (a) the PGF-induced increase in tension to 84 and 35 %, (b) the level attained during the depolarization by 2 and 12 mV and (c) the peak rise in [Ca2+]i to 42 and 17 %. The sustained rises in [Ca2+]i were resistant to nifedipine. 4. In Ca2+-free solution (containing 1 mM EGTA), PGF elicited an increase in tension that was 26 % of that in 2.5 mM Ca2+ and an increase in [Ca2+]i (24 % of the sustained level) at mid-pregnancy but no increase in tension or [Ca2+]i at term. 5. At both stages of pregnancy, PGF decreased the level of [Ca2+]i required to elicit increases in tension comparable to those evoked by high K+o. The slope of the tension-[Ca2+]i curves were steeper in mid- than in late pregnancy. 6. In conclusion, at mid-pregnancy, the contractile response of the guinea-pig myometrium to PGF involves Ca2+ influx through L-type voltage-operated Ca2+ channels (VOCCs) and by receptor-operated mechanisms, release of Ca2+ from intracellular stores, and an increase in the sensitivity of the contractile apparatus to Ca2+. At term the situation is different: a modest increase in the sensitivity of the contractile apparatus to Ca2+ persists and there is a major reliance on Ca2+ influx through VOCCs. PMID- 10718756 TI - The canine parasternal and external intercostal muscles drive the ribs differently. AB - 1. In the dog, the elevation of the ribs during inspiration results from the combined actions of the parasternal and external intercostal muscles. In the present studies, the hypothesis was tested that co-ordinated activity among these two sets of muscles reduces the distortion of the rib cage. 2. During spontaneous inspiration before or after section of the phrenic nerves, the ribs moved cranially and outward in the same way as they did during passive inflation. However, whereas the sternum moved cranially during passive inflation, it was displaced caudally during spontaneous inspiration. 3. When the parasternal intercostal muscles were selectively denervated, both the sternum and the ribs moved cranially, but the rib outward displacement was markedly reduced. In contrast, when the external intercostals were excised and the parasternal intercostals were left intact, the sternum continued to move caudally and the outward displacement of the ribs was augmented relative to their cranial displacement. 4. These observations establish that the external intercostal muscles drive the ribs primarily in the cranial direction, whereas the parasternal intercostals drive the ribs both cranially and outward. They also indicate, in agreement with the hypothesis, that co-ordinated activity among these two sets of muscles displaces the ribs on their relaxation curve. 5. However, this co-ordinated activity also displaces the sternum caudally. Although this distortion requires an additional energy expenditure, it enhances the outward component of rib displacement which is more effective with respect to lung expansion. PMID- 10718757 TI - Ischaemic changes in refractoriness of human cutaneous afferents under threshold clamp conditions. AB - 1. A technique was developed to counteract the changes in threshold to electrical stimuli of large myelinated cutaneous afferents in the human median nerve induced by ischaemia for 13 min. Intermittent application of polarizing currents was used in five subjects, in whom refractoriness, supernormality and the strength duration time constant (tauSD) were tracked to determine whether compensating for the ischaemia-induced changes in threshold also controlled the ischaemic changes in these excitability parameters. 2. The threshold compensation prevented the ischaemic changes in tauSD, an excitability parameter dependent on nodal Na+ channels. Threshold compensation did not prevent the changes in refractoriness and supernormality, whether the compensation began 10, 100 or 200 ms prior to the test stimuli. 3. In three subjects, continuous polarizing current was injected for 13 min to compensate for the ischaemic change in threshold, thus clamping threshold at the pre-ischaemic level. Again, tauSD was effectively controlled, but there were still ischaemic changes in refractoriness and supernormality. 4. The effective control of tauSD suggests that both the intermittent threshold compensation and the continuous threshold clamp effectively controlled membrane potential at the node of Ranvier. 5. The ischaemic increase in refractoriness when threshold was kept constant could be due to interference with the processes responsible for refractoriness by a metabolic product of ischaemia. The ischaemic change in supernormality during effective compensation probably results from the intrusion of refractoriness into the conditioning-test intervals normally associated with maximal supernormality. 6. The present results indicate that ischaemia has effects on axonal excitability that cannot be readily explained by changes in membrane potential. Specifically, it is suggested that ischaemic metabolites interfere with the recovery of Na+ channels from inactivation. PMID- 10718759 TI - DALI apheresis in hyperlipidemic patients: biocompatibility, efficacy, and selectivity of direct adsorption of lipoproteins from whole blood. AB - Recently, the first apheresis technique for direct adsorption of low-density lipoprotein (LDL) and lipoprotein(a) [Lp(a)] from whole blood (DALI) was developed that does not require a prior plasma separation. That markedly simplifies the extracorporeal circuit. The aim of the present study was to test the acute biocompatibility, efficacy, and selectivity of DALI apheresis. In a prospective clinical study, 6 hypercholesterolemic patients suffering from angiographically proven atherosclerosis were treated 4 times each by DALI. 1.3 patient blood volumes were treated per session at blood flow rates of 60-80 ml/min using 750 or 1,000 ml of polyacrylate/polyacrylamide adsorber gel. The anticoagulation consisted of an initial heparin bolus followed by a citrate infusion. The sessions were clinically essentially uneventful. Mean corrected reductions of lipoproteins amounted to 65% for LDL-cholesterol, 54% for Lp(a), 28% for triglycerides, 1% for HDL-cholesterol, and 8% for fibrinogen. The selectivity of lipoprotein removal was high. Cell counts remained virtually unchanged and no signs of hemolysis or clotting were detected. Cell activation parameters elastase, beta-thromboglobulin, interleukin-1beta, and IL-6 showed no significant increase. Complement activation was negligible. There was significant, but clinically asymptomatic, bradykinin activation in the adsorber with mean maxima of 12,000 pg/ml in the efferent line at 1,000 ml of treated blood volume. In conclusion, DALI proved to be safe, selective, and efficient for the adsorption of LDL-C and Lp(a), which simplifies substantially the extracorporeal therapy in hypercholesterolemic patients. PMID- 10718758 TI - Major role for sensory feedback in soleus EMG activity in the stance phase of walking in man. AB - 1. Sensory feedback plays a major role in the regulation of the spinal neural locomotor circuitry in cats. The present study investigated whether sensory feedback also plays an important role during walking in 20 healthy human subjects, by arresting or unloading the ankle extensors 6 deg for 210 ms in the stance phase of gait. 2. During the stance phase of walking, unloading of the ankle extensors significantly (P < 0.05) reduced the soleus activity by 50 % in early and mid-stance at an average onset latency of 64 ms. 3. The onset and amplitude of the decrease in soleus activity produced by the unloading were unchanged when the common peroneal nerve, which innervates the ankle dorsiflexors, was reversibly blocked by local injection of lidocaine (n = 3). This demonstrated that the effect could not be caused by a peripherally mediated reciprocal inhibition from afferents in the antagonist nerves. 4. The onset and amplitude of the decrease in soleus activity produced by the unloading were also unchanged when ischaemia was induced in the leg by inflating a cuff placed around the thigh. At the same time, the group Ia-mediated short latency stretch reflex was completely abolished. This demonstrated that group Ia afferents were probably not responsible for the decrease of soleus activity produced by the unloading. 5. The findings demonstrate that afferent feedback from ankle extensors is of significant importance for the activation of these muscles in the stance phase of human walking. Group II and/or group Ib afferents are suggested to constitute an important part of this sensory feedback. PMID- 10718760 TI - Use of clinical guidelines for treatment of anemia among hemodialysis patients. AB - Changing financial incentives have strongly influenced dosing patterns of recombinant human erythropoietin (rHuEPO) since its introduction in 1989. Although guidelines for prescribing rHuEPO exist, the extent to which they are adhered to is unknown. Using a retrospective cohort observational study design, the factors influencing the initial dosing of rHuEPO prescribed to 413 hemodialysis patients in 1994 were examined. Patient weight, the only recommended guideline, was not found to be a significant predictor of dosing of rHuEPO after controlling for selected patient demographic and clinical characteristics. The strongest predictor for initial rHuEPO dosing was hematocrit followed by White race (p < 0.05). Finally, each subsequent month was associated with a significantly larger initial rHuEPO dose, reflecting the general trend in increasing dose since 1991 (p < 0.001). In conclusion, despite the recent DOQI guidelines for treatment of anemia among persons with chronic renal failure, providers are not using patient weight as an independent criterion for determining dosing of rHuEPO. PMID- 10718761 TI - In vitro evaluation study of the membrane autotransfusion system experimental prototype: MATS-I. AB - Membrane Autotransfusion System (MATS) utilizing plasmapheresis technology has been developed in our laboratory. A specially designed polyethylene hollow fiber membrane was utilized. This study was conducted to evaluate performance of the first experimental prototype, MATS-I. The results of this study showed that the MATS-I could concentrate diluted blood at 10% of the initial hematocrit concentration (HCTi) into over 40% after passing through the system at a transmembrane pressure of 70 mm Hg. Moreover, the MATS-I can continuously treat 10,000 ml of diluted blood at various HCTi levels without deteriorating its performances. Even though the MATS-I met all required performances as an autotransfusion system, several areas of improvement of the system were necessary to meet various clinical needs. The next prototype, MATS-II, can be designed based on experiences obtained from the MATS-I. The MATS is smaller, more atraumatic and continuous, and is a faster system when compared to the currently available centrifugal autotransfusion devices. PMID- 10718762 TI - Removal of immunoglobulins by a protein A versus an antihuman immunoglobulin G based system: evaluation of 602 sessions of extracorporeal immunoadsorption. AB - Elimination of IgG can be achieved by extracorporeal immunoadsorption (IA) based on specific binding to either staphylococcal protein A (Excorim) or sheep polyclonal antibodies directed against human IgG (Therasorb). In 602 analyzed sessions of IA, elimination of IgG was 60% through 80% depending on the treated plasma volume, with no significant difference between the mentioned systems. However, the decrease of IgM and IgA was approximately 50% in the anti-IgG compared to 20-40% in the protein A system. Plasma albumin concentration decreased by 20% in the anti-IgG system compared to 15% in the protein A system, and hemoglobin values increased by 2% in the anti-IgG system and decreased by 6% in the protein A system. In conclusion, a clinical relevance for these findings cannot be ruled out, and the individual choice might depend on the clinical situation and laboratory findings. PMID- 10718763 TI - Preprimed artificial lung for emergency use. AB - We investigated the possibility of preprimed storage of an artificial lung (AL), aiming at facilitating its emergency use. Test ALs, consisting of a special microporous hollow fiber membrane made of polyolefin in which direct blood-gas contact was completely eliminated, were preprimed with saline solution, sterilized by gamma-ray irradiation, and evaluated after 1-3 months of storage at room temperature. A small amount of bubble was noted in the priming solution after storage in some ALs, which most likely originated from the air dissolved in the priming solution or persisted in the liquid compartment at priming. Although the preprimed solution contained several polyolefin-breakdown products due to irradiation, including ethyl alcohol, n- and t-butyl alcohol, acetone, and carbon dioxide, the levels of these substances were at concentrations known to be not toxic. Endotoxin concentration was negligible. In SEM observation, no perceptible microstructural change was observed in the hollow fibers after preprimed storage. Maximum tensile stress and ultimate elongation of the hollow fiber in the test ALs were reduced by approximately 20% and 3%, respectively, from those of the control AL. The influence of preprimed storage on gas-exchange function was examined in a venoarterial bypass animal study using a goat. Oxygen transfer function was well preserved whereas carbon dioxide removal function was slightly lowered according to the storage term in the stored ALs compared with those of a nonpreprimed control AL. On the basis of these results, we conclude that preprimed storage of the AL with gamma-ray sterilization is basically feasible and realistic. PMID- 10718764 TI - Validation of a model for flow-dependent carbon dioxide exchange in artificial lungs. AB - The exchange rate of CO2 in artificial lungs depends on the sweep gas flow rate. Control of the amount of CO2 removed by an artificial lung requires quantitative knowledge of the flow dependence. A simple model of the dependence of CO2 exchange on sweep gas flow rate in artificial lungs has been previously presented (1). For a given partial pressure of CO2 in the blood phase, sweep gas flow rate, and CO2 exchange rate, the model indicates how close the CO2 exchange rate is to the maximum level attainable by the artificial lung. The focus of this study was to validate the model experimentally by testing 2 commercial artificial lungs in an in vitro test loop. The CO2 exchange rate for each artificial lung was measured over a range of sweep gas flow rates. Linear regression was used to fit the data to the model and estimate the maximum possible CO2 exchange rate and the average water-side PCO2 (PCO2w). The difference between the measured and regressed values of PCO2w was used as an indicator of the ability of the model to quantitatively predict the dependence of CO2 exchange on gas flow rate. This difference was less than 5% for each experiment, indicating that the model can be used to guide control of CO2 exchange rates in artificial lungs. PMID- 10718765 TI - Comparative In vitro evaluation of two different preparations of small diameter polyurethane vascular grafts. AB - Polyurethane (PU) frequently has been used to manufacture small diameter vascular grafts due to its good bicompatibility. In this study, sponge PU small diameter vascular grafts were fabricated from Pellethane 2103, as well as a self synthesized PU, by utilization of a salt casting technique and by varying the salt/polymer ratios. Two types were made; one of them had a thin solid layer on the outer surface. The inner surface was identical and was coated with gelatin crosslinked by epoxide. Tensile properties, compliance, platelet activation, and endothelial cell attachment were evaluated in vitro. It was found that for the grafts with an outer nonporous coating, compliance could not be estimated from modulus due to anisotropy. The most compliance-matched grafts were made of self synthesized polyurethane with a salt/polymer ratio 4 to 8 or Pellethane with a ratio 6 to 8 without the outer nonporous coating. The former had better elongation. Self-synthesized PU had lower platelet adhesion as well as more endothelial attachment than Pellethane. Porosity activated platelets strongly and reduced endothelial adhesion unless the surface was modified by the crosslinked gelatin layer. It was concluded that PU synthesized in our lab with a salt/polymer ratio of at least 4 and coated with epoxy crosslinked gelatin was a better substrate for preparation of small diameter vascular grafts. PMID- 10718766 TI - Synergistic effect of released aspirin/heparin for preventing bovine pericardial calcification. AB - Calcification is a frequent cause of the clinical failure of bioprosthetic heart valves fabricated from glutaraldehyde pretreated bovine pericardium (GATBP). Aspirin, a potent antiplatelet drug, and heparin, an anticoagulant, are commonly used for postimplant complications such as thrombosis and thromboembolism. Aspirin and heparin were embedded in chitosan/polyethylene vinylacetate co-matrix to develop a prolonged release form. The effect of these drugs towards the bioprosthetic calcification was investigated by in vitro and in vivo models. In vitro and in vivo evaluation suggest that the released aspirin/heparin from the co-matrix had a synergistic effect in inhibiting GATBP calcification. In vivo subcutaneous co-implantation was performed with PEG-20,000 grafted bovine pericardium (PEG-GABP), aspirin, and heparin. Biochemical, histological, and scanning electron microscopic evaluation of retrieved samples demonstrated a significant reduction in calcium deposition and alkaline phosphatase activity on PEG-GABP compared to GATBP. It seems that the aspirin/heparin combination synergistically inhibits the pericardial calcification in addition to their antithrombotic function. PMID- 10718767 TI - Biodesign of a skeletal muscle flap as a model for cardiac assistance. AB - In using autologous muscles for cardiac assistance, it is crucial to reduce ischemia-reperfusion injury in the surgically traumatized skeletal muscle. In adult sheep, we developed a simple model of surgically designed 2 latissimus dorsi muscle leaflets by modifying the vascular supply to these leaflets. Three pockets with graded injury were established, and muscle morphology and vascular remodeling were monitored in 3 experimental groups: muscle leaflets without any treatment (Group 1, n = 6) that served as controls; muscle leaflets integrated with a fibrin interlayer (Group 2, n = 6); and leaflets integrated with fibrin and entrapped pyrrolostatin (Group 3, n = 6). We applied the fibrinogen and thrombin solutions, which polymerize to form a three-dimensional meshwork joining the tissues, creating a provisional matrix for angiogenesis, and acting as a delivery depot for agents aimed at minimizing ischemia-reperfusion lesion formation. After 2 months, the muscle leaflets biointegrated with the fibrin interface showed none of the signs of necrosis or ischemia-reperfusion lesions seen in the controls. Although no angiogenic factors were incorporated, the fibrin interlayer rapidly (<2 weeks) became a densely vascularized tissue replete with a voluminous capillary network. In contrast, controls showed poor bonding between the tissues, muscle fiber deterioration, and a compromised vascular network. Muscle structure was best preserved and angiogenesis was greatest when pyrrolostatin, a free radical scavenger, was added to the fibrin meshwork to reduce damage caused by overproduction of free radicals. This newly designed model will be useful to study many current approaches in cardiovascular biology, from pharmaceuticals to gene therapy, which might prove advantageous in muscle designed cardiac assistance. PMID- 10718768 TI - Potentialities and problems of a novel bilateral ventricular assist system without thoracotomy. AB - Mechanical cardiac assistance can be critical in saving the lives of patients with acute cardiac failure. However, currently used methods of ventricular assistance require advanced technical knowledge and equipment, and only small numbers of patients can be provided with them. Our aim was to develop new cannulas to construct a less invasive biventricular assist system that would permit easy application without thoracotomy. We prepared 2 centrifugal pumps and 4 uniquely shaped cannulas and conducted experiments to investigate the potential and problems of the system. In the first experiment, the system was attached to 6 dogs with ventricular fibrillation to confirm whether our system could maintain cardiac output. In the second experiment, the system was installed for 3 days in 3 goats, and changes in aminotransferases, BUN, creatinine, and plasma free hemoglobin levels were examined. In Experiment 1, it was demonstrated that the system was able to maintain circulation in dogs. In Experiment 2, although the flow rate of the pumps was maintained over 3 days, increased hemolysis and deteriorating renal function were observed. Although these problems need to be solved, the system was still helpful in the management of acute biventricular failure for short periods and may be clinically useful in the near future. PMID- 10718769 TI - Easy aortic cannulation: a transxyphoid approach. AB - An easy aortic cannulation technique in minimally invasive pediatric cardiac surgery is described. We have developed a dilator which fits an aortic perfusion cannula. The aortic cannula over the dilator with a hole for passage of a guide wire is inserted into the ascending aorta by the Seldinger technique. Using this technique, the cannula never slips off, even at a slant angle to the aorta owing to the guidance of the guide wire. We recommend this safe and reliable technique for insertion of an aortic cannula into the ascending aorta in minimally invasive pediatric cardiac surgery. PMID- 10718770 TI - A novel in vitro assessment of tissue valve calcification by a continuous flow type method. AB - A dynamic flow type testing to study calcification was self-designed to investigate calcification in bioprosthetic heart valves. The apparatus consists of a container into which leaflets from a porcine aortic valve are placed, a chamber that contains calcium solution, and a peristaltic pump that provides a continuous supply of the solution toward the container. Efficacy of the apparatus was compared with the conventional batch type calcification testing at 37 degrees C through measuring the amount of calcium and phosphate deposited by inductively coupled plasma (ICP) and scanning electron microscope (SEM). After 14 days, calcium levels detected from the calcified deposit on leaflets were 470.4 +/- 37.0 microg/cm3 in the flow type testing whereas in the batch type testing levels were 81.0 +/- 6.7 microg/cm3. Though the calcium level on the leaflet increased as the exposure time to calcium solution increased in both testings, the rate and the tendency of calcification could be assessed very rapidly by flow type testing in comparison with batch type testing. [Ca]/[P] molar ratio decreased over time, and after 14 days, the ratio was close to 1.83 +/- 0.18 in the flow type testing. The ratio could not be determined in the batch type testing because the deposit was too small to assess. The descending rate of [Ca]/[P] molar ratio demonstrates that deposited calcium-complex at the earliest stage may interact with inorganic phosphate ions to create a calcified deposit mineral precursor. This in vitro dynamic flow type calcification testing was a favorable tool for rapid investigation of calcification. PMID- 10718771 TI - Development of a centrifugal pump with thick blades. AB - We have developed a centrifugal blood pump with thick impeller blades (60% of pitch) to obtain a small tip clearance. An unshrouded impeller with 6 backward curved thick blades was used to reduce the dead zone between the shroud and upper casing. A streamline angle in volute was uniform in circumferential direction by continuity and angular momentum conservation. To prove the effectiveness of small tip clearance, performance and hemolysis tests were conducted on pumps with a tip clearance of 0.5, 1.5, and 2.0 mm at exit with the blade thickness of 60% of pitch, and with that of 1.0, 2.0, and 2.5 mm at exit with the thickness of 40% of pitch. The results showed that the smaller the tip clearance, the better the hydrodynamic and hemolytic performance. The best result was seen in the pump with tip clearance of 0.5 mm with a blade thickness of 60% of pitch. These results suggest that a centrifugal pump with thick blades and a small tip clearance can be a promising alternative as a cardiopulmonary bypass pump. PMID- 10718772 TI - Low density lipoprotein (LDL) apheresis is currently recognized as a reliable tool for removal of LDL cholesterol directly from blood in patients with severe hypercholesterolemia. PMID- 10718774 TI - Pharmacokinetics of bambuterol during oral administration of plain tablets and solution to healthy adults. AB - AIMS: The pharmacokinetics of orally administered bambuterol were investigated in healthy adult subjects, with particular regard to time to steady state, pharmacokinetic linearity, intraindividual variability for the parent drug and its active beta2-adrenergic metabolite terbutaline and bioequivalence between tablet and solution. METHODS: Twenty-six healthy Caucasian subjects were included and 23 (12 women) completed this open, randomised, crossover study. Racemic bambuterol hydrochloride was administered orally as 10 mg, 20 mg, and 10 + 20 mg tablets, and as a solution once daily for 2 weeks at about 19.00 h. Plasma concentrations and urinary recoveries of bambuterol and terbutaline were measured after single doses and during repeated treatments. RESULTS: Absorption of bambuterol was biphasic. The initial rate could not be assessed directly, but it was faster than that during the second phase where absorption was rate-limiting for elimination (mean terminal half-life: 16 h). Steady-state AUC(0,24 h) of bambuterol, reached within 1 week, was not dose-linear. Mean terminal half-life of terbutaline was 22 h and steady-state was reached within one week of bambuterol treatment. Contrary to bambuterol, overall pharmacokinetics of terbutaline indicated dose-linearity. Day-to-day intraindividual variation in AUC(0,24 h) of terbutaline, 15% with the tablet, was half that of bambuterol. Urine data indicated that intraindividual variability was slightly smaller with the solution. Tablets were bioequivalent with the solution with regard to terbutaline (90% confidence interval: 87-100%). CONCLUSIONS: With oral bambuterol steady state was reached within 1 week. Regarding generated terbutaline, pharmacokinetics judged to be were linear, intraindividual variability of AUC at steady state was on average 15% with the tablet, and tablets were bioequivalent with the solution. PMID- 10718773 TI - Ion channels and the control of blood pressure. AB - Ion channels exist in all cells and are enormously varied in structure, function and regulation. Some progress has been made in understanding the role that ion channels play in the control of blood pressure, but the discipline is still in its infancy. Ion channels provide many different targets for intervention in disorders of blood pressure and exciting advances have been made in this field. It is possible that new drugs, as well as antisense nucleotide technology or gene therapy directed towards ion channels, may form a new class of treatments for high and low blood pressure in the future. PMID- 10718775 TI - The pharmacokinetics of morphine and morphine glucuronide metabolites after subcutaneous bolus injection and subcutaneous infusion of morphine. AB - AIMS: To investigate the pharmacokinetics of morphine, morphine-6-glucuronide (M6G) and morphine-3-glucuronide (M3G) in healthy volunteers after the administration of morphine by subcutaneous bolus injection (s.c.b.) and subcutaneous infusion (s.c. i.) over 4 h, and to compare the results with the intravenous bolus (i.v.) administration of morphine. METHODS: Six healthy volunteers each received 5 mg morphine sulphate by i.v., s.c.b. and short s.c.i. over 4 h, on three separate occasions, in random order, each separated by at least 1 week. Plasma samples were assayed for morphine, M6G and M3G. RESULTS: After i.v. morphine, the concentrations of morphine, M6G and M3G and their pharmacokinetic parameters were similar to those we have observed previously, in other healthy volunteers (when standardized to nmol l- 1, for a 10 mg dose to a 70 kg subject). After s.c.b. morphine, similar results were obtained except that the median tmax values for morphine and M3G were significantly longer than after i.v. morphine (P< 0.001 and P< 0.05, respectively), with a trend to a longer tmax for M6G (P = 0. 09). The appearance half-lives after s.c.b. morphine for M6G and M3G were also significantly longer than after i.v. morphine (P = 0.03 and P< 0.05, respectively). Comparison of log-transformed AUC values indicated that i.v. and s.c.b. administration of morphine were bioequivalent with respect to morphine, M6G and M3G. In comparison with i.v. morphine, morphine by s.c.i. was associated with significantly longer median tmax values for morphine (P< 0.001), M6G (P< 0.001) and M3G (P< 0.05), and the mean standardized Cmax values significantly lower than after both i.v. and s.c.b. morphine (morphine P< 0.001, M6G P< 0.001 and M3G P< 0.01 for each comparison). Comparison of log-transformed AUC values after i.v. and s.c.i. morphine indicated that the two routes were not bioequivalent for morphine (log-transformed AUC ratio 0.78, 90% CI 0.66-0.93), M6G (0.72, 90% CI 0.63-0.82), or M3G (0.65, 90% CI 0.54-0.78). A small stability study indicated no evidence of adsorptive losses from morphine infused over 4 h using the infusion devices from the study. CONCLUSIONS: Although bioequivalence was demonstrated between the s. c.b. and i.v. routes of morphine administration, the bioavailabilities of morphine, M6G and M3G after s.c.i. were significantly lower than after i.v. administration. However, despite this, the study demonstrates that the subcutaneous route is an effective method for the parenteral administration of morphine. PMID- 10718776 TI - Effect of renal function on risedronate pharmacokinetics after a single oral dose. AB - AIMS: To determine the relationship between risedronate pharmacokinetics and renal function. METHODS: Risedronate was administered to adult men and women (n=21) with various degrees of renal function (creatinine clearance 15-126 ml min 1 ) as a single oral dose of 30 mg. Serum samples were obtained for 72 h after dosing, and urine samples were collected for 72 h after dosing and then periodically for 6 weeks. Risedronate concentrations were determined using an enzyme-linked immunosorbent assay (ELISA). Risedronate serum concentration-time and urinary excretion rate-time profiles were analysed simultaneously using nonlinear regression. RESULTS: Renal clearance and volume of distribution were linearly related to creatinine clearance (r2=0.854, P<0.001; and r2=0.317, P<0.01, respectively). Decreases in predicted renal clearance and volume of distribution of 82 and 69%, respectively, were observed when creatinine clearance decreased from 120 to 20 ml min-1. A 64% decrease in predicted oral clearance was observed when creatinine clearance decreased from 120 to 20 ml min-1 (P=0.064). Iohexol clearance, a predictor of renal function, produced similar results to those observed with creatinine clearance. Risedronate was well tolerated by the study population. CONCLUSIONS: Risedronate renal clearance was significantly related to a decrease in renal function. There was a consistent reduction in oral clearance with a decrease in creatinine clearance. However, based on the regression analysis, generally no dosage adjustment appears to be necessary for most patients with mild or moderate renal impairment (creatinine clearance >20 ml min-1 ). PMID- 10718777 TI - Pharmacokinetics and pharmacodynamics of gliclazide in Caucasians and Australian Aborigines with type 2 diabetes. AB - AIMS: Gliclazide pharmacokinetics and pharmacodynamics were assessed in 9 Caucasians and 10 Australian Aborigines with uncomplicated type 2 diabetes. METHODS: Subjects were on a stable dose of 80 mg gliclazide twice daily, took 160 mg on the morning of study and had a standard breakfast. No further gliclazide was given over the next 48 h. Regular blood samples were drawn for serum glucose, insulin and gliclazide assay. Gliclazide was measured using h.p.l.c. Noncompartmental analysis was used to describe primary data. A multicompartment model incorporating entero-hepatic recirculation was fitted to group mean serum gliclazide profiles. RESULTS: The Caucasians were older than the Aborigines (mean +/- s.d. age 53.4 +/- 12.2 vs 40.3 +/- 6.9 years, P < 0.05) but had similar diabetes duration, body mass index and glycated haemoglobin. Noncompartmental analysis revealed no between-group differences in gliclazide kinetics. Post breakfast serum glucose and insulin responses were also similar apart from a longer time to maximum concentration (tmax) for glucose amongst the Aborigines (2.6 +/- 0.4 vs 2.2 +/- 0. 3 h in Caucasians; P = 0.024). Gliclazide tmax exhibited a skewed unimodal distribution and was not associated with gliclazide maximum concentration, or glucose or insulin responses. Most patients had a serum gliclazide profile suggestive of enterohepatic recirculation and/or biphasic absorption. Model-derived estimates of the extent of putative enterohepatic recirculation were 30% and 20% of dose in Caucasians and Aborigines, respectively. CONCLUSIONS: Gliclazide is equally effective in Caucasian and Aboriginal diabetic patients. The pharmacokinetics of oral gliclazide appear more complex than previously thought. Gliclazide pharmacodynamics are unrelated to rate and extent of absorption, consistent with a threshold concentration for hypoglycaemic effect. PMID- 10718779 TI - The effect of NAT2 genotype and gender on the metabolism of caffeine in nonsmoking subjects. AB - AIMS: To establish whether gender or N-acetyltransferase 2 (NAT2) genotype influence the urinary 17 U+17X/137X ratio after dosing with caffeine. METHODS: Ninety-two nonsmoking individuals underwent caffeine phenotyping. NAT2 genotype was determined by the polymerase chain reaction followed by a restriction digest (PCR-RFLP). RESULTS: The median ratio for urinary 17 U+17X/137X was 6.7 (range 1.45-18. 65). 55% of subjects were slow acetylators. Gender did not affect the metabolic ratio or NAT2 genotype. Mean 17 U+17X/137X ratio differed between fast (6.75) and slow (8.69) acetylators (95% CI for the difference, 0.32-3.56). CONCLUSIONS: The findings are further evidence that the 17 U+17X/137X urinary ratio is not a robust measure of CYP1A2 activity. A possible mechanism by which the ratio might be influenced by NAT2 genotype is suggested. PMID- 10718778 TI - Pharmacokinetics and pharmacodynamics of sibrafiban alone or in combination with ticlopidine and aspirin. AB - AIMS: The purpose of this clinical study was to evaluate the effects of a ticlopidine/aspirin combination on the pharmacokinetics and pharmacodynamics of sibrafiban and the tolerability of the combination therapy METHODS: Thirty-eight healthy male volunteers were randomized to receive one of the following treatments for 7 days: sibrafiban (n = 12), ticlopidine/aspirin (n = 12), or the combination treatment sibrafiban/ticlopidine/aspirin (n = 14). Concentrations of the active metabolite of sibrafiban, Ro 44-3888, in plasma and urine were determined by column-switching liquid chromatography combined with tandem mass spectrometry. The pharmacodynamics of sibrafiban and ticlopidine/aspirin were examined by measuring the inhibition of ADP- or collagen-induced platelet aggregation. RESULTS: The addition of ticlopidine/aspirin to sibrafiban did not significantly alter the pharmacokinetic parameters of Ro 44-3888. the geometric mean ratio for AUC(0,12h) was 110 (95% CI 0.82, 1.22). Separately, sibrafiban and ticlopidine/aspirin inhibited ADP-and collagen-induced platelet aggregation and the effects of the two treatments were additive. For example, the average inhibition of ADP-induced platelet aggregation over 12 h was 42% in the sibrafiban treated group, 55% in the ticlopidine/aspirin group and 69% in the sibrafiban/ticlopidine group. The bleeding time was prolonged in the treatments with ticlopidine/aspirin (8.1 min) and sibrafiban/ticlopidine/aspirin (8. 6 min) compared with sibrafiban alone (3.5 min). CONCLUSIONS: This study shows a significant pharmacodynamic interaction between sibrafiban and ticlopidine/aspirin. Consequently, the simultaneous administration of sibrafiban and ticlopidine/aspirin should be carefully monitored to ensure the patient's coverage with an antiplatelet drug without exposure to an excessive bleeding risk. PMID- 10718780 TI - Inhibitory effects of amiodarone and its N-deethylated metabolite on human cytochrome P450 activities: prediction of in vivo drug interactions. AB - AIMS: To predict the drug interactions of amiodarone and other drugs, the inhibitory effects and inactivation potential for human cytochrome P450 (CYP) enzymes by amiodarone and its N-dealkylated metabolite, desethylamiodarone were examined. METHODS: The inhibition or inactivation potency of amiodarone and desethylamiodarone for human CYP activities were investigated using microsomes from B-lymphoblastoid cell lines expressing CYP1A1, CYP1A2, CYP2A6, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and CYP3A4. The in vivo drug interactions of amiodarone and desethylamiodarone were predicted in vitro using the 1+Iu/Ki values. RESULTS: Amiodarone weakly inhibited CYP2C9, CYP2D6, and CYP3A4-mediated activities with Ki values of 45.1-271.6 microm. Desethylamiodarone competitively inhibited the catalytic activities of CYP2D6 (Ki=4.5 microm ) and noncompetitively inhibited CYP2A6 (Ki=13.5 microm ), CYP2B6 (Ki=5.4 microm ), and CYP3A4 (Ki=12.1 microm ). The catalytic activities of CYP1A1 (Ki=1.5 microm, alpha=5.7), CYP1A2 (Ki=18.8 microm, alpha=2.6), CYP2C9 (Ki=2.3 microm, alpha=5.9), and CYP2C19 (Ki=15.7 microm, alpha=4.5) were inhibited by desethylamiodarone with mixed type. The 1+Iu/Ki values of desethylamiodarone were higher than those of amiodarone. Amiodarone inactivated CYP3A4, while desethylamiodarone inactivated CYP1A1, CYP1A2, CYP2B6, and CYP2D6. CONCLUSIONS: The interactions between amiodarone and other drugs might occur via the inhibition of CYP activities by its N-dealkylated metabolite, desethylamiodarone, rather than by amiodarone itself. In addition, the inactivation of CYPs by desethylamiodarone as well as by amiodarone would also contribute to the drug interactions. PMID- 10718781 TI - Dutch hospital drug formularies: pharmacotherapeutic variation and conservatism, but concurrence with national pharmacotherapeutic guidelines. AB - AIMS: This research examines current hospital drug formularies (HDFs) of all Dutch general hospitals. It assesses the extent to which they recommend the same drugs, the breadth of their coverage in terms of therapeutic areas, drug groups incorporated and individuals drugs included, and their extent of conservatism by considering the year of introduction of the drugs included within groups. Furthermore, it considers the extent to which their recommendations concur and comply with those of national pharmacotherapeutic guidelines and the WHO Essential Drugs List (EDL). METHODS: Seventy-eight (81%) out of all 96 current Dutch HDFs were received of which 62 were suitable for study. Differences between HDFs and eventual associations with hospital characteristics were researched by statistical testing and case-control studies. To evaluate HDFs' concurrence with national guidelines and compliance with the WHO EDL, nine drug groups were studied in detail: benzodiazepines, calcium channel blockers, beta-adrenoceptor blocking agents, ACE-inhibitors, angiotensin-II inhibitors, NSAIDs, H2-receptor antagonists, 5HT3-antagonists, and H+-pump inhibitors. Concurrence and compliance with national guidelines and the WHO EDL was defined as inclusion of recommended drugs. Non-concurrence was defined as inclusion of nonrecommended drugs. RESULTS: The total number of indications addressed and drug groups incorporated within HDFs varied from 28 to 72 (median 56) and from 30 to 123 (median 97), respectively. The total number of individual drug entities (pharmacological substances) included ranged from 239 to 658 (median 430) and the total number of drug products, including all different dosage forms, from 412 to 1121 (median 655). Within drug groups, drug entities first marketed were most frequently included. Teaching hospitals were most likely to include recently marketed drugs. Depending on the drug group, HDFs' concurrence and compliance with national guidelines and the WHO EDL ranged from 35% to 100%. CONCLUSIONS: Findings indicate that Dutch HDFs are rather uniform in the indications addressed and the drug groups incorporated. However, the number of individual drug entities and drug products included within groups varies considerably. Furthermore, Dutch HDFs are considered rather conservative, as older drugs are favoured over more recent drugs. Generally, with some drug exceptions, Dutch HDFs concur and comply with recommendations in national pharmacotherapeutic guidelines and with the WHO EDL over 90%. PMID- 10718782 TI - Birth outcome following maternal use of metoclopramide. The Euromap study group. AB - AIMS: Metoclopramide is an antiemetic drug used widely during pregnancy for nausea and vomiting. Because of its frequent use any adverse effects on infant health would have major public health implications. We therefore examined the safety of metoclopramide during pregnancy. METHODS: Using the Pharmaco Epidemiological Prescription Database of North Jutland County, we identified 309 women with singleton pregnancies who had prescriptions for metoclopramide fom 1 January 1991 to 31 December 1996. Information on malformations, birth weight, preterm deliveries and stillbirth were compared with 13 327 references who did not receive prescriptions of any kind during pregnancy. RESULTS: Mean birth weight among exposed women was 3480 g compared with 3470 g among nonexposed. Based on logistic regression models no major differences in the risk were found concerning malformations (OR= 1.11; 95% confidence interval (CI): 0.6-2.1); low birth weight (OR= 1.79; 95% CI: 0.8-3.9) or preterm delivery (OR= 1.02; 95 CI: 0.6-1.7). CONCLUSIONS: We could not document any association between the use of metoclopramide during pregnancy and adverse pregnancy outcome. Because of the limited power of our study further research is required. PMID- 10718783 TI - Single dose methodology to assess the influence of an alpha1-adrenoceptor antagonist on uroflowmetric parameters in patients with benign prostatic hyperplasia. AB - AIMS: To establish methodology which rapidly and reliably assesses the effect of an alpha1-adrenoceptor antagonist on peak urine flow rates in men with benign prostatic hyperplasia (BPH). This methodology could then be applied to screening new drugs to treat BPH. METHODS: Twenty-five patients with BPH enrolled in a double-blind, placebo-controlled, two-period crossover study. Patients were either withdrawn from their current alpha1-adrenoceptor antagonist therapy (n = 22) or were untreated prestudy (n = 3) and all met prespecified uroflowmetric criteria including: (1) a peak urine flow rate (Qmax) < 12 ml s-1 off therapy (or < 10 ml s-1 if untreated prestudy) and (2) a decrease in peak urine flow rate (Qmax) of > 2 ml s-1 after withdrawal from therapy. Study treatment consisted of tamsulosin 0.4 mg (or matching placebo) once daily for 8 days in a two-period crossover. Uroflowmetry was performed predose and once postdose (4.5-5.5 h postdose) on day 1, and once postdose (4.5-5.5 h postdose) on day 8 of each treatment period. RESULTS: After a single dose of tamsulosin, the least-square mean difference between tamsulosin and placebo in the change from baseline Qmax was 2.8 ml s-1 (P = 0.017 vs placebo). After 8 days dosing of tamsulosin, the least-square mean difference between tamsulosin and placebo in the change from baseline Qmax was also 2.8 ml s-1 (P = 0.044 vs placebo). Additionally, there was no significant difference observed between the single and multiple dose results (P > 0.200 for between group difference). CONCLUSIONS: Both single and multiple doses of tamsulosin 0.4 mg increased Qmax in men with BPH. A single dose produced a comparable response to multiple dose administration. The magnitude of the effect was greater than the effect generally seen in longer term clinical trials, but this difference may be explained by the patient population in this study which was preselected for 'responsiveness' to an alpha1-adrenoceptor antagonist. These results support the utility of single dose uroflowmetric measurements in rapidly providing preliminary data on new investigational agents, specifically agents which act to increase urine flow in men with BPH. However, clinical efficacy would still need to be confirmed with longer term clinical trials. PMID- 10718784 TI - Gastric decontamination performed 5 min after the ingestion of temazepam, verapamil and moclobemide: charcoal is superior to lavage. AB - AIMS: The aim was to study the efficacy of gastric lavage and activated charcoal in preventing the absorption of temazepam, verapamil and moclobemide when gastric decontamination was performed immediately after ingestion of the drugs. METHODS: Nine healthy volunteers took part in a randomized cross-over study with three phases. The subjects were administered single oral doses of 10 mg temazepam, 80 mg verapamil and 150 mg moclobemide. Five minutes later, they were assigned to one of the following treatments: 200 ml water (control), 25 g activated charcoal as a suspension in 200 ml water or gastric lavage. Plasma concentrations and the cumulative excretion into urine of the three drugs were determined up to 24 h. RESULTS: The mean AUC(0,24 h) of temazepam, verapamil and moclobemide was reduced by 95.2% (P < 0.01), 92.8% (P < 0.01) and 99. 7% (P < 0.01), respectively, by activated charcoal compared with control. Gastric lavage did not reduce significantly the AUC(0,24 h) of these drugs. The 24 h cumulative excretion of temazepam, verapamil and moclobemide into urine was reduced significantly (P < 0.05) by charcoal but not by gastric lavage. Charcoal reduced the AUC(0,24 h), Cmax and urinary excretion of all three drugs significantly more than lavage. CONCLUSIONS: Activated charcoal is very effective and gastric lavage can be rather ineffective in preventing the absorption of temazepam, verapamil and moclobemide when the treatment is given very rapidly after ingestion of the drugs, before tablet disintegration has occurred. PMID- 10718786 TI - Effect of a single oral dose of Fansidar on the pharmacokinetics of halofantrine in healthy volunteers: a preliminary report. PMID- 10718785 TI - Comparative oral bioavailability of azimilide dihydrochloride in the fed and fasted states. AB - AIMS: This study investigated the relative oral bioavailability of azimilide dihydrochloride following administration in the fed (high-fat meal) and fasted states. METHODS: This was a single-dose, randomized, two-way crossover study in 30 healthy, Caucasian, male subjects. Following oral administration, blood samples were collected over 27 days and analysed for azimilide using h.p.l.c. with u.v. detection. Pharmacokinetic parameters were determined using 'noncompartmental' analysis and compared using an ANOVA and 90% or 95% confidence intervals. RESULTS: The extent of absorption was equivalent in the fed and fasted states (ratio = 96.2%; 90% CI=90.5% -102.4%). However, Cmax was decreased 19% following a high-fat meal (ratio=81.4%; 90% CI= 76.2% -87.0%). No difference in tmax or t(1/2),z was observed. CONCLUSIONS: Azimilide dihydrochloride may be orally administered to patients without regard to the prandial state. PMID- 10718787 TI - Genetic polymorphism of CYP2D6 in a keralite (South India) population. PMID- 10718788 TI - Excretion of fluvoxamine into breast milk. PMID- 10718789 TI - Evidence-based assessment of severity and management of acute pancreatitis. PMID- 10718790 TI - Imaging of soft tissue tumours. PMID- 10718792 TI - COPE: committee on publication ethics PMID- 10718791 TI - Where can surgeons publish? PMID- 10718793 TI - Warfarin induced skin necrosis. AB - BACKGROUND: Warfarin induced skin necrosis is a rare complication with a prevalence of 0.01-0.1 per cent. It was first described in 1943. METHODS: A literature review was undertaken using Medline; all relevant papers on this rare compli-cation of warfarin therapy were used. RESULTS: There are several adverse skin manifestations associated with the use of oral anticoagulants, ranging from ecchymoses and purpura, haemorrhagic necrosis, maculopapular vesicular urticarial eruptions to purple toes. This article concentrates mainly on warfarin induced skin necrosis. The syndrome typically occurs during the first few days of warfarin therapy, often in association with the administration of a large initial loading dose of the drug. Although the precise nature of the disease is still unknown, advances in knowledge about protein C, protein S and antithrombin III anticoagulant pathways have led to a better understanding of the mechanisms involved in pathogenesis. Differential diagnosis between warfarin induced skin necrosis and necrotizing fasciitis, venous gangrene and other causes of skin necrosis may be difficult; the disease may also be confused with other dermatological entities. CONCLUSION: Warfarin induced skin necrosis, while rare, is an important complication. All surgeons should be aware of its existence. PMID- 10718794 TI - Pain after laparoscopic cholecystectomy. AB - BACKGROUND: Although laparoscopic cholecystectomy (LC) results in less pain than open chole-cystectomy, it is not a pain-free procedure. Many methods of analgesia for pain after laparoscopy have been evaluated. METHODS: Forty-two randomized controlled trials assessing interventions to reduce pain after LC are reviewed, as are the mechanisms and nature of pain after this procedure. RESULTS: Non steroidal anti-inflammatory drugs, wound local anaesthetic, intraperitoneal local anaesthetic, intraperitoneal saline, a gas drain, heated gas, low-pressure gas and nitrous oxide pneumo-peritoneum have been shown to reduce pain after LC. The clinical significance of this pain reduction is questionable. CONCLUSION: Pain after LC is multifactorial. Although many methods of analgesia produce short-term benefit, this does not equate with earlier discharge or improved postoperative function. However, single trials evaluating low-pressure insufflation, heated gas and multimodal analgesia suggest that clinically relevant benefits can be achieved. PMID- 10718795 TI - Randomized clinical trial of granisetron, droperidol and metoclopramide for the treatment of nausea and vomiting after laparoscopic cholecystectomy. AB - BACKGROUND: Patients undergoing laparoscopic cholecystectomy (LC) may be especially at risk of experiencing postoperative nausea and vomiting (PONV). This study was undertaken to evaluate the efficacy of granisetron, droperidol and metoclopramide for the treatment of PONV after LC. METHODS: After experiencing PONV during the first 3 h after recovery from anaesthesia, 120 patients (78 women) received, in a randomized double-blind manner, granisetron 40 microg/kg, droperidol 20 microg/kg or metoclopramide 0.2 mg/kg (n = 40 per group) intravenously. Patients were then observed for 24 h after administration of the study drug. RESULTS: Complete control of established PONV, defined as no emetic symptoms and no need for another rescue antiemetic medication, was achieved in 88 per cent of patients with granisetron, 60 per cent with droperidol and 55 per cent with metoclopramide (P < 0.05). No clinically adverse events were observed in any of the groups. CONCLUSION: A high dose of granisetron (40 microg/kg) was more effective than droperidol 20 microg/kg or metoclopramide 0.2 mg/kg for the treatment of established PONV after LC. PMID- 10718796 TI - Late results of a randomized clinical trial comparing total fundoplication versus calibration of the cardia with posterior gastropexy. AB - BACKGROUND: The aim was to perform a prospective randomized study in patients with chronic gastro-oesophageal reflux treated either by total fundoplication or calibration of the cardia with posterior gastropexy. Late follow-up considered subjective and objective parameters, and related outcome to the presence of Barrett's oesophagus. METHODS: A total of 164 patients were randomized to fundoplication (n = 76) or calibration of the cardia (n = 88). They were evaluated by clinical questionnaire, upper gastrointestinal endoscopy with biopsies, oesophageal manometry and gastro-oesophageal reflux studies, including scintigraphy and 24-h oesophageal pH monitoring. RESULTS: There were no operative deaths. There was 95 per cent follow-up at a mean of 85 months. The mean recurrence rate for both operations was near 40 per cent at 10 years, but patients without Barrett's oesophagus had a recurrence rate after both operations of around 23 per cent compared with 83 per cent after 10 years for those with Barrett's oesophagus (P < 0.0001). Low-grade dysplasia developed in 13 per cent of the patients with Barrett's oesophagus. There were significant differences in all objective parameters in a comparison of patients with Visick I or II and those with Visick III or IV disease at the late assessment. CONCLUSION: Both total fundoplication and calibration of the cardia with posterior gastropexy had similar subjective and objective late results. However, results were significantly worse in patients with Barrett's oesophagus. PMID- 10718797 TI - Treatment of recurrent colorectal liver metastases by interstitial laser photocoagulation. AB - BACKGROUND: Hepatic resection improves survival in selected patients with colorectal liver metastases. The treatment of recurrent hepatic metastases after resection is controversial. Interstitial laser photocoagulation, performed under local anaesthesia, offers a minimally invasive option to repeat resection. The first series of patients with recurrent colorectal liver metastases treated with photo- coagulation is reported. METHODS: Nineteen patients (five women and 14 men, median age 57 (range 44-71) years) who developed recurrent colorectal liver metastases after hepatectomy (five with bilateral disease) were treated with photocoagulation between 1993 and 1997. Fifteen patients also received chemotherapy (14 systemic, one hepatic arterial) before photocoagulation. RESULTS: There were no major complications or deaths related to the treatment. Six patients developed minor complications related to the procedure but did not require any form of intervention. Median survival from commencement of photocoagulation was 16 (range 4-36) months. CONCLUSION: Photocoagulation is a safe, minimally invasive therapy that may be used as an adjunct to chemotherapy and repeat resection in the treatment of recurrent colorectal liver metastases, and may lead to improved survival. PMID- 10718798 TI - Relationship of carotenoid and vitamins A and E with the acute inflammatory response in acute pancreatitis. AB - BACKGROUND: Inflammation and oxidative stress are believed to be important in the development of the systemic complications of acute pancreatitis. The fat-soluble vitamins A and E, and the carotenoids have antioxidant properties. The aim of this study was to assess the effect of acute pancreatitis on serum concentrations of vitamin antioxidants and to relate such changes to the degree of the inflammatory response. METHODS: Thirteen consecutive patients with predicted severe acute pancreatitis were compared with 26 matched healthy controls. Five patients developed severe acute pancreatitis and three of these died. Vitamin antioxidant and C-reactive protein (CRP) levels were measured daily for up to 7 days. RESULTS: Patients had significantly lower levels of antioxidants throughout the course of the study (P < 0.017). In patients there was a significant correlation between peak CRP and trough antioxidant levels (P < 0. 01). In patients with mild acute pancreatitis, the concentrations of retinol and beta carotene at final review were significantly higher than those in patients with severe acute pancreatitis (P < 0.05). This coincided with a reduction in CRP level. CONCLUSION: In acute pancreatitis, circulating concentrations of vitamin antioxidants are reduced and are inversely related to the rise in CRP level. PMID- 10718799 TI - Radical resection of hilar bile duct carcinoma and predictors of survival. AB - BACKGROUND: Patients with carcinoma of the main hepatic duct have a poor prognosis. This study attempted to identify clinicopathological predictors of survival after resection. METHODS: A retrospective review was performed of 114 patients who presented with hepatic ductal carcinoma between 1976 and 1998. Of the 114 patients, 98 had a radical resection, three underwent palliative resection and 13 were not treated surgically. Forty-six patients with stage IVA disease had microscopic tumour residue after resection. Of these, 28 patients were treated with radiotherapy and the remaining 18 had resection alone. RESULTS: The overall operative morbidity and mortality rates were 14 and 4 per cent respectively. The overall 5-year survival rate after resection was 28 per cent. Nineteen patients survived for more than 5 years, including ten with stage IVA disease. The main prognostic factors were performance status; jaundice; tumour location; gross appearance; histological grade; T, N and M categories in tumour node metastasis (TNM) classification; TNM stage; and residual tumour. Adjuvant radiotherapy, tumour extension into the hepatic ducts, histological grade, N and residual tumour were independent predictive factors by multivariate Cox analysis. CONCLUSION: This study suggests that radical resection provides the best survival rate for patients with hilar bile duct carcinoma. For patients with stage IVA disease, following complete gross resection radiotherapy improved treatment outcome. PMID- 10718800 TI - Influence of allograft size to recipient body-weight ratio on the long-term outcome of renal transplantation. AB - BACKGROUND: The critical nephron mass needed to meet the metabolic demands of an individual depends on the body-weight. This study evaluated the effect of the kidney transplant ultrasonographic size to recipient body-weight ratio (Tx/W) on the outcome of kidney transplantation. METHODS: A consecutive series of 104 cadaveric renal transplants was studied. Transplant cross-sectional area (TXSA) was measured ultrasonographically in the first week after transplantation as an index of renal size. A 'nephron dose' index (Tx/W) was calculated by dividing TXSA by recipient weight and was used to define three groups of patients, with high (more than 0.45), medium (0.3-0.45) or low (less than 0.3) Tx/W ratios. Isotope glomerular filtration rate (GFR) measurements were made at 1, 6 and 12 months after transplantation. RESULTS: The serum creatinine level was significantly lower in the first 5 years after transplantation in patients with a high Tx/W ratio than in those with a medium or low ratio. GFR measurements were marginally higher in the groups with a high and medium Tx/W ratio compared with the low Tx/W group. A statistically significant association between Tx/W ratio and graft survival was not found. CONCLUSION: The renal transplant size to recipient weight ratio was an important determinant of long-term renal allograft function in this study. Extreme mismatching between allograft and recipient size should be avoided where possible, but the findings presented require confirmation in larger studies before clear recommendations can be made about size matching and kidney allocation. PMID- 10718801 TI - Carotid duplex imaging: variation and validation. AB - BACKGROUND: Duplex imaging is increasingly used as the only investigation before carotid endarterectomy, but many different criteria exist in the literature for the detection of a severe (70-99 per cent) carotid stenosis. This study aimed to investigate current practice in carotid duplex imaging in Great Britain and Ireland. METHODS: A postal questionnaire was sent to 86 vascular surgical units. RESULTS: The median number of scans performed per year was 450 (range 60-4500). Thirty-six per cent of units who responded used peak systolic : end diastolic velocity ratio to calculate carotid stenosis. Overall, nine different major duplex criteria were used to grade carotid stenosis in 14 different systems of percentage bands. Only 51 per cent of units verified their duplex criteria against angiography. Eighteen per cent of units used two or more different types of duplex scanner and applied the same diagnostic criteria to each machine. CONCLUSION: A wide variation in diagnostic duplex criteria and methods of grading stenosis exists among vascular units. Internal validation is not performed routinely. Standardization of duplex criteria would ensure greater consistency, but would not replace the need for validation of results within each unit. PMID- 10718802 TI - Cost-effective carotid endarterectomy. AB - BACKGROUND: Although carotid endarterectomy is increasing in the UK, there is evidence that the procedure is still underused. Methods of reducing cost in a single vascular unit have been assessed using a continuous audit including outcome measures. METHODS: A consecutive series of 333 patients admitted over 7 years under a single consultant surgeon were studied. Outcome measures included the rate of perioperative neurological complication of any kind, and death. The length of hospital stay and the number of readmissions within 30 days were recorded prospectively by computerized audit. RESULTS: Over the interval of the study, the number of preoperative investigations was reduced; angiography and cerebral computed tomography were reserved for specific indications. The median duration of hospital stay decreased from 7 to 2 days. There was no change in the stroke and death rate (3 per cent) during the study and only two patients required readmission within 30 days. CONCLUSION: Carotid endarterectomy can be performed cost-effectively using non-invasive preoperative investigations for the majority of patients. In-hospital stay has been reduced and the routine use of intensive care replaced by a 2-h stay in theatre recovery. These changes have been achieved without compromising patient safety. PMID- 10718803 TI - Epidemiology of lower extremity amputation in centres in Europe, North America and East Asia AB - BACKGROUND: This study was established to enable a comparison of lower extremity amputation incidence rates between different centres around the world. METHODS: Ten centres, all with populations greater than 200 000, in Japan, Taiwan, Spain, Italy, North America and England collected data on all amputations done between July 1995 and June 1997. Patients were identified from at least two data sources (to allow checks on ascertainment); denominator populations were based on census figures. RESULTS: The highest amputation rates were in the Navajo population (43.9 per 100 000 population per year for first major amputation in men) and the lowest in Madrid, Spain (2.8 per 100 000 per year). The incidence of amputation rose steeply with age; most amputations occurred in patients over 60 years. In most centres the incidence was higher in men than women and the incidence of major amputations was greater than that of minor amputations. Diabetes was associated with between 25 and 90 per cent of amputations. CONCLUSION: Apart from the Navajo centre, differences in the known prevalence of diabetes could not account for the differences in overall incidence of amputation. Differences in the prevalence of peripheral vascular disease are likely to be important, but this and the role of other factors, including availability of health care, are worthy of further investigation. PMID- 10718804 TI - Prospective study of the effect of laparoscopic hemifundoplication on motor and sensory function of the proximal stomach. AB - BACKGROUND: Some 30 per cent of patients develop dyspeptic symptoms following antireflux surgery. These symptoms may result from alterations in the motor and sensory function of the proximal stomach. METHODS: Proximal gastric motor and sensory function was studied with an electronic barostat in 12 patients with reflux who underwent laparoscopic hemifundoplication. In addition, 24-h pHmetry, gastric emptying (scintigraphy) and vagus nerve integrity (pancreatic polypeptide response to hypo-glycaemia) were assessed. Fifteen healthy volunteers served as controls. RESULTS: Laparoscopic hemifundoplication significantly decreased total acid exposure time (P < 0.05). Vagus nerve function remained intact in all but one patient. The mean(s.e.m.) lag phase for emptying of solids was significantly shorter after operation than before (15(3) versus 21(3) min; P < 0.05). Proximal gastric compliance was not significantly different before and after fundoplication. However, mean(s.e.m.) postprandial relaxation was significantly reduced (P < 0.05) after hemifundoplication compared with the value before operation (3341(1105) versus 12 763(3616) ml over 90 min) and in controls (14 567(2358) ml over 90 min). Postprandial fullness was significantly increased after hemi-fundoplication (P < 0.05). Postprandial gastric relaxation correlated with the lag phase for emptying of solids (r = 0.55, P < 0.02). CONCLUSION: After hemifundoplication, proximal gastric compliance is not altered but postprandial relaxation is impaired and associated with sensations of fullness and shorter duration of the lag phase for emptying of solids. PMID- 10718805 TI - Diagnostic accuracy of needle-localized open breast biopsy for impalpable breast disease. AB - BACKGROUND: Needle-localized open breast biopsy (NLBB) is considered the gold standard procedure for the diagnosis of impalpable breast disease. In an observational follow-up study the sensitivity and negative predictive value of this procedure was determined in a clinical population with long-term follow-up. METHODS: Some 199 consecutive patients with a benign histological diagnosis on NLBB were followed for the occurrence of breast cancer, using information from the Dutch National Morbid-Anatomical Record Department. Based on a review of mammograms and histological slides, an expert panel decided whether the carcinomas detected during follow-up were newly developed, or were present already at the time of the NLBB. RESULTS: After a median follow-up of 60.5 months, seven carcinomas were detected. At panel review, six appeared to have been missed by NLBB. The sensitivity of NLBB was 99 per cent after 2 years of follow-up, but dropped to 96 per cent after 5 years. Similarly, the negative predictive value dropped from 99 per cent after 2 years to 94 per cent after 5 years of follow-up. CONCLUSION: NLBB is an accurate diagnostic procedure for the evaluation of impalpable breast disease. However, with longer follow-up the accuracy becomes lower than generally reported. PMID- 10718806 TI - Intraperitoneal polypropylene mesh repair of incisional hernia is not associated with enterocutaneous fistula. AB - BACKGROUND: Incisional hernia repair with prosthetic material is followed by fewer recurrences than primary repair. Polypropylene is the most commonly used prosthetic material but may cause entero- cutaneous fistulas. The aim of this study was to determine whether enterocutaneous fistulas developed after incisional hernia repair with polypropylene mesh and to evaluate clinical outcome after incisional hernia repair. METHODS: A retrospective analysis of the outcome of incisional hernia repair with polypropylene mesh between 1982 and 1998 was conducted. Follow-up data were obtained from medical records and questionnaires. RESULTS: Polypropylene incisional hernia repair was performed in 136 patients. Median follow-up was 34 months. No enterocutaneous fistulas developed. Wound infection occurred in 6 per cent. Wound sinus formation occurred in two patients. No mesh was removed because of infection and no persisting infection of the mesh occurred. CONCLUSION: Enterocutaneous fistula formation appears to be very rare after incisional hernia repair with polypropylene mesh, regardless of intraperitoneal placement, omental coverage or closing of the peritoneum. PMID- 10718807 TI - Postoperative outcome and sites of recurrence in patients following curative resection of gastric cancer. AB - BACKGROUND: Recurrence occurs in a variety of forms and in different organs after 'curative resection' of gastric cancer. This study investigated the postoperative prognosis for each type of recurrence. METHODS: From 1969 to 1988, 939 patients with gastric cancer underwent curative resection; data on 130 of 207 patients who died with recurrence were analysed. Attention was focused on the site of recurrence and the postoperative outcome. RESULTS: Haematogenous recurrence was evident in 54 per cent (70 of 130 patients), peritoneal recurrence in 43 per cent (56 of 130), lymph node recurrence in 12 per cent (16 of 130) and local recurrence in 22 per cent (29 of 130). Thirty-three patients (25 per cent) had recurrences at multiple sites. Peritoneal and local recurrences were related to infiltrative growth, in contrast to haematogenous and lymphatic recurrences. There were no statistical differences in survival time among each type of recurrence and survival was not related to the number of sites of recurrence. Survival did not depend on factors of sex, age, tumour location, tumour size, depth of invasion, tissue differentiation, histological growth pattern, lymphatic and vascular involvement, lymph node metastasis and extent of lymph node dissection. CONCLUSION: The clinicopathological characteristics of gastric cancer determine the type of recurrence, although the clinical outcome is the same for each type of tumour and is not related to the number of sites of recurrence. PMID- 10718808 TI - Changing patterns in the management of gastric volvulus over 14 years. AB - BACKGROUND: Gastric volvulus is an uncommon condition, which can be difficult to diagnose and treat. This study represents a large series of patients with the condition. METHODS: All patients presenting with gastric volvulus over a 14-year period were reviewed. RESULTS: Some 36 patients (median age 75 years) were identified. Volvulus, usually secondary to a hiatus hernia, presented acutely in 29 patients. The major symptoms were abdominal pain, vomiting and upper gastrointestinal haemorrhage. The most useful investigations were barium contrast studies and upper gastrointestinal endoscopy, which were helpful in 21 of 25 and 11 of 18 patients respectively. Treatment was conservative in five patients, by open surgery in 13 and laparoscopic repair in 18 (three converted to open operation). There were no major complications and no deaths. Median hospital stay was shorter in patients treated by laparoscopic rather than open surgery (6 (range 4-36) versus 14 (7-50) days; P < 0.05). CONCLUSION: Acute and chronic gastric volvulus can be treated successfully by either open or laparoscopic surgery. However, laparoscopic surgery now represents a safe and acceptable approach, with minimal morbidity and a significantly shorter hospital stay. This is likely to be of considerable benefit for the treatment of a predominantly elderly population, often with significant co-morbidity. PMID- 10718809 TI - Laparoscopic splenectomy: a suitable technique for children and adults AB - Aims: Splenectomy retains an important role in the management of certain haematological conditions that fail to respond to conventional medical therapy, and has traditionally been performed through a midline or left subcostal incision with patients requiring 5-7 days in hospital. The well recognized benefits of laparoscopic surgery should also apply to splenectomy. This study aimed to develop a safe and effective technique suitable for all age ranges and without the requirement for expensive stapling devices. METHODS: An operative technique evolved over the 5-year period from 1994, from an initial six-port approach with the patient supine, to a four-port approach in a modified right lateral position, with locking surgical clips applied down a 5-mm port to vessels in the hilum, and removal of the spleen within a retrieval bag through a 4-6-cm Pfannanstiel incision. Data were collected prospectively for all patients undergoing laparoscopic splenectomy at Leicester Royal Infirmary, including demographic details, indication for surgery, duration of surgery, length of inpatient stay, transfusion requirement, postoperative complications and the response of the original condition to surgical intervention. RESULTS: A total of 40 patients underwent laparoscopic splenectomy (14 children, 26 adults) for a variety of conditions (idiopathic thrombocytopenia (ITP) (n = 24), haemolytic anaemia (n = 9) or malignancy (n = 7)) with a median operating time of 180 min for the first 20 patients and 100 min for the second 20 (P < 0.0001), and median inpatient stay of 3 days for the first 20 patients and 2 days for the second 20 (P < 0.0003). None of the operations was converted to open surgery, five patients required blood and/or platelet transfusion perioperatively, none of the patients had major postoperative complications, 23 of the 24 patients with ITP developed normal platelet counts after operation, and all nine patients with haemolytic anaemia maintained a normal haemoglobin concentration after operation. CONCLUSION: Laparoscopic splenectomy can be performed safely and effectively in adults and children without the need for stapling devices. PMID- 10718810 TI - Association of tumour necrosis factor microsatellite haplotypes with chronic pancreatitis AB - AIMS: Recent evidence implicates the immune system in having a primary pathogenic role in the development of chronic pancreatitis (CP). This study examined the relationship between the tumour necrosis factor (TNF) gene and susceptibility to CP. METHODS: DNA was extracted from the blood of 40 patients with CP, 107 ethnically matched normal controls and 33 patients with alcohol-induced liver disease (alcoholic controls). Three microsatellite polymorphisms at the TNF locus were examined by the polymerase chain reaction, gel electrophoresis and autoradiography. RESULTS: The TNFa6 allele was significantly over-represented in patients with CP compared with normal controls (chi2 = 4.09, P = 0.043), and in those with alcoholic CP (n = 24) compared with alcoholic controls (chi2 = 4.63, P = 0.031). The three locus haplotypes 6-5-1 and 2-5-2 were significantly over represented (chi2 = 4.14, P = 0.042) and under-represented (chi2 = 4.63, P = 0.042) respectively in patients with alcoholic CP compared with alcoholic controls. CONCLUSIONS: A TNF haplotype has been defined that is associated with CP. This is consistent with a genetically determined role for TNF in the immunopathogenesis of CP. PMID- 10718811 TI - Mismatch between trainees' subspecialist interest and advertised jobs; worrying implications for upper gastrointestinal trainees AB - AIMS: The aim of this study was first to assess the primary subspecialist interests of general surgical specialist registrars who were accredited and still looking for a consultant position, or who were within 3 years of their CCST (certificate of completion of surgical training). These interests were then compared with subspecialist interests declared in consultant vacancies advertised in the British Medical Journal over the preceding 16 months. METHODS: All trainees in general surgery holding a national training number in six regions (Mersey, North West, Trent, Yorkshire, Northern, West Midlands) were identified, and those accredited or within 3 years of their CCST (n = 136) were telephoned to ascertain their primary subspecialist interest, whether they had a higher degree and what their desired consultant job would be. The consultant vacancies advertised in the British Medical Journal between 3 January 1998 and 8 May 1999 were assessed according to the required subspecialist interest. RESULTS: Upper gastrointestinal surgery is the second most popular subspecialty (n = 37; 27 per cent of trainees) after colorectal surgery (n = 40; 29 per cent of trainees). However, there were fewer consultant vacancies in upper gastrointestinal surgery (31 of 226; 14 per cent of jobs) than in any of the other three main subspecialist areas of general surgery (colorectal, vascular and breast/endocrine). The ratio of percentage of jobs to percentage of trainees was lowest in upper gastrointestinal surgery (0.50), compared with colorectal (0.77), vascular (0.89) and breast/endocrine (2.23) surgery. CONCLUSIONS: Upper gastrointestinal surgery appears to be the most competitive of the general surgical subspecialties at present, having by far the lowest ratio of jobs to trainees. In addition, 87 per cent of upper gastrointestinal trainees have or are completing a higher degree, and 43 per cent expressed a desire to work in a teaching hospital. PMID- 10718812 TI - Prospective 5-year audit for day-case laparoscopic cholecystectomy AB - AIMS: Laparoscopic cholecystectomy can be performed as a day-case procedure but success depends on adequate recovery time, patient motivation and physical condition. To determine the proportion of patients suitable for a day-case procedure, 23-h stay and inpatient treatment, a prospective audit was introduced in 1994. METHODS: A 5-year (1994-1999) prospective annual audit was performed for all patients undergoing laparoscopic cholecystectomy (537 patients). In the first 3 years patients were either classified as day-case or inpatient stay; the establishment of a 23-h stay ward allowed a third option for these patients. RESULTS: Day-case laparoscopic cholecystectomy, including 23-h stay in the final 2 years of the audit, increased in each of the 5 years from 16 to 80 per cent. The audit loop was closed when the successful allocation rate for day-case patients was reaudited; this increased from 56 per cent in the first year to 88 per cent in the final year. CONCLUSIONS: This audit has identified the need for extended recovery for patients undergoing more major procedures on an ambulatory basis such as laparoscopic cholecystectomy. Further refinements in selection criteria and improved analgesia control are needed to reduce the unexpected extended stay for day-case and ambulatory patients. PMID- 10718813 TI - Current practice in the management of acute cholecystitis AB - AIMS: Several recent papers have advocated emergency cholecystectomy for patients with acute cholecystitis, stating that it is safe, cost effective and leads to less time off work. This study was designed to assess current practice in the management of acute cholecystitis in the UK. METHODS: A postal questionnaire was sent to 357 consultant surgeons who were thought to be involved in a general surgical on-call rota, to ascertain their current management of patients with acute cholecystitis. Replies were received from 250 consultants (70 per cent) of whom 242 (68 per cent) were involved in a general surgical take. Sixteen of these consultants, however, handed their patients with acute cholecystitis on to a different team the following day for further management. RESULTS: Twenty-seven consultants (12 per cent) routinely treat their patients by emergency cholecystectomy whenever possible, with 24 stating that they would do this within 72 h. Limiting factors to this practice were stated to be availability of surgical staff (15), theatre space (nine) and radiological investigations (four). The remaining consultants (n = 199) routinely manage their patients conservatively initially and providing they settle, either (1) book directly for cholecystectomy (n = 94, 47 per cent), (2) reassess as an outpatient (n = 65, 33 per cent), (3) either of above (n = 21; 11 per cent) or (4) refer on to a colleague (n = 19, 10 per cent). The commonest indications for acute cholecystectomy stated by consultants whose initial treatment policy is conservative are spreading peritonitis due to bile leak (93 per cent), empyema (89 per cent), unexpected space on a theatre list (28 per cent) and failure of an acute episode to settle (21 per cent). The laparoscopic method is the commonest for both elective and emergency cholecystectomy, but the percentage of consultants using an open method rises dramatically from 9 per cent in the elective situation to 48 per cent for emergency cholecystectomy. CONCLUSIONS: Despite evidence to support the increased use of emergency cholecystectomy, this practice is routinely carried out by only 12 per cent of consultants. However, of the consultants who treat their patients conservatively, 28 per cent are prepared to undertake emergency cholecystectomy if an unexpected space appears on the theatre list. PMID- 10718814 TI - Extending the indications for curative liver resection by portal vein embolization AB - AIMS: The aim of ipsilateral portal vein embolization is to induce hypertrophy of normal tissue when resection of a cancerous portion of the liver is contraindicated only by the volume of liver that would remain following surgery. This study reports its use in primary and metastatic liver tumours. METHODS: Eight patients with inoperable liver tumours (three women and five men of median age 68. 5 years; three colorectal hepatic metastases, two cholangiocarcinomas and three hepatocellular cancers) were selected for portal vein embolization. Selected portal branches were occluded distally with microbeads and proximally with coils. Liver volumes were determined by magnetic resonance imaging before embolization and again before surgery, 6-8 weeks later. RESULTS: Embolization was performed successfully in seven patients by the percutaneous-transhepatic route; one further patient required an open cannulation of the inferior mesenteric vein. Management was altered in six patients, who proceeded to 'curative' surgery. The projected remaining (predominantly left lobe) liver volumes increased significantly from a median of 350 to 550 ml (P < 0.05, Wilcoxon matched pairs test). Two patients had disease progression such that surgery was no longer indicated. One patient, whose disease progressed, had the left portal branch occluded unintentionally by a misplaced coil that was successfully retrieved, although the left portal branch remained occluded. CONCLUSIONS: Portal vein embolization produced significant hypertrophy of the normal liver and extended the option of 'curative' surgery to six of the eight patients in whom it was attempted. It appears to be equally effective for primary and metastatic liver tumours in selected patients. PMID- 10718816 TI - Author's reply PMID- 10718815 TI - Evaluation of pylorus-preserving pancreato-duodenectomy with the imanaga reconstruction by hepatobiliary and gastrointestinal dual scintigraphy. PMID- 10718817 TI - Surgical management of severe secondary peritonitis. PMID- 10718818 TI - Author's reply PMID- 10718819 TI - Randomized controlled trial to examine the influence of thoracic epidural analgesia on postoperative ileus after laparoscopic sigmoid resection. PMID- 10718820 TI - Authors' reply PMID- 10718821 TI - Non-surgical management of erectile dysfunction. AB - Erectile dysfunction is a common and distressing medical condition that is now highly amenable to treatment almost irrespective of the cause. Safe, non-surgical treatments with unequivocal efficacy are psychological therapy, intracorporeal injection of vasoactive drugs, transurethral vasodilators and oral sildenafil, all of which have been reported to have a 50-70% overall response rate. Vacuum constriction devices are acceptable for some, usually older patients and oral yohimbine is thought to have marginal efficacy. Local creams to induce or enhance erectile function are currently being investigated. There is no place for androgen supplementation unless the patient is profoundly hypogonadal. Treatment of hyperprolactinaemia is very effective but is a rare cause of erectile dysfunction. As intercourse may entail an unfamiliar level of physical activity, it is sensible to ensure that the patient is able to climb a flight or two of stairs comfortably without provoking undue breathlessness or chest pain and to provide suitable advice about technique before commencing treatment. Once it is clear to the patients that erectile dysfunction can be satisfactorily overcome, the long-term use of treatments to do so tends to wane. Thus, although the prospect of effective treatment for what had been for many a distressing life sentence has the potential to place new demands on the health service, there is no evidence that restrictions on prescribing will prove economically rational. PMID- 10718822 TI - Clinical features and pathogenesis of thyrotoxicosis in adult Melanesians in Papua New Guinea. AB - OBJECTIVE: To investigate the cause of an apparent increase in the incidence of thyrotoxicosis in the Melanesian population of the coastal areas around Port Moresby, Papua New Guinea. DESIGN: Consecutive patients attending the thyroid clinic at Port Moresby General Hospital were included in the study which collected clinical and immunogenetic data. PATIENTS: One hundred and ninety-four patients with thyrotoxicosis were studied. The mean age was 31 years. The clinical features were typical for thyrotoxicosis apart from increased pigmentation, which occurred in the majority of patients and which reversed with treatment with antithyroid drugs and resolution of the thyrotoxicosis. MEASUREMENTS: Antibodies to thyroid peroxidase, thyroglobulin and the thyrotropin receptor were present in 99%, 52% and 69% of patients, respectively, confirming that all cases were caused by autoimmune thyroid disease. Fine needle aspirations in 42 of 43 patients were consistent with Graves' disease. CONCLUSIONS: The cause of this apparent sudden increase in Graves' disease in the coastal region around Port Moresby, an apparently iodine-replete area, remains unexplained. PMID- 10718823 TI - Association of thyrotrophin receptor antibodies with the clinical features of Graves' ophthalmopathy. AB - OBJECTIVE: Graves' ophthalmopathy (GO) and Graves' hyperthyroidism are closely associated diseases and thought to be caused by the same autoimmune process. An obvious explanation for this would be the presence of autoantibodies reacting with an autoantigen present in the orbit and the thyroid gland. The TSH-Receptor (TSH-R) antibodies are a likely candidate, because they cause Graves' hyperthyroidism and the TSH-R appears to be present also in orbital tissues. If TSH-R antibodies are responsible for the ophthalmopathy one would expect their titres to correlate with clinical characteristics of the eye disease. The aim of the present study is to see whether TSH-R antibodies are related to the activity and severity of the thyroid-associated ophthalmopathy. DESIGN AND PATIENTS: TSH-R antibody levels were measured as TBII (TRAK assay), and TSI (cAMP response of a TSH-R transfected cell line) in serum of 63 patients with untreated moderately severe GO, accompanying Graves' thyroid disease; all patients had been euthyroid for > 2 months. RESULTS: TBII and TSI titres were strongly related to each other. TBII or TSI titres did not correlate with thyroidal or orbital disease duration, nor with TPO antibody levels. In contrast, we found a striking and highly significant correlation between the Clinical Activity Score (CAS) of the eye disease, and both TBII (r = 0.54; P < 0.0001) and TSI (r = 0.50; P < 0.0001). In addition, a weaker but significant relation was found between proptosis (in mm) and TBII (r = 0.36; P = 0.004) and TSI (r = 0.49; P = 0.0001). No correlation was found with eye muscle motility. CONCLUSION: TSH-R antibody levels correlate directly with clinical features of Graves' ophthalmopathy. The results support the hypothesis of a pathogenetic role of TSH-R antibodies and the TSH-R in the orbit of Graves' ophthalmopathy patients. PMID- 10718825 TI - A novel mutation in the pendrin gene associated with Pendred's syndrome. AB - OBJECTIVE: Pendred's syndrome is an autosomal recessive disorder characterized by goitre, sensorineural deafness and iodide organification defect. It is one of the most frequent causes of congenital deafness, accounting for about 10% of hereditary hearing loss. It is caused by mutations in the pendrin (PDS) gene, a 21 exon gene located on chromosome 7. The aim of this study was to examine an Italian family affected with Pendred's syndrome at the molecular level. PATIENTS: Thirteen subjects belonging to a family from Southern Italy were evaluated for the clinical and genetic features of Pendred's syndrome. MEASUREMENTS: Exons 2-21 of the PDS gene were amplified from peripheral leucocytes by the polymerase chain reaction; mutation analysis was performed by single strand conformation polymorphism, direct sequencing and restriction analysis. RESULTS: The index patient had the classical triad of the syndrome and harboured two mutations in the PDS gene in the form of compound heterozygosity. He was found to be heterozygous for a cytosine to adenosine mutation at nucleotide 1523 in exon 13 and for a IVS 1001 + 1G --> A mutation. The former is a novel mutation which results in a change of 508 threonine to asparagine in the putative eleventh transmembrane domain. The latter mutation in the donor splice site has already been described in other patients and is thought to lead to aberrant splicing and premature protein truncation. Three subjects who were heterozygous for one mutation had normal phenotypes. Two subjects had sensorineural deafness and were heterozygous for a single mutation. Goitre was found only in patients with Pendred's syndrome and was absent in all other individuals, albeit residing in an iodine-deficient area. CONCLUSIONS: We have identified a novel mutation in the pendrin gene causing Pendred's syndrome, and confirm that molecular analysis is a useful tool for a definitive diagnosis. This is particularly relevant in cases such as in the subjects of our family in which the clinical features might be misleading and other genetics factors might be responsible for deafness. PMID- 10718824 TI - Determinants of thyroid volume in healthy French adults participating in the SU.VI.MAX cohort. AB - OBJECTIVE: To study the relative importance of determinants of thyroid volume. DESIGN: Cross-sectional study on a sample of subjects issued from the SU.VI.MAX cohort. SUBJECTS: 2987 French subjects (1713 women aged 35-60 years and 1274 men aged 45-60 years). None of them had previous or present thyroid disease. MEASUREMENTS: Thyroid volume was determined by ultrasound. Serum TSH and free thyroxine (fT4) were measured in duplicate. Urinary iodine and urinary thiocyanate were assayed in random morning urine samples. RESULTS: For both sexes, thyroid volume (ml) was positively correlated with weight, height, body mass index and body surface area (P = 0.0001) and negatively with age for females (P = 0.0009). When the urinary iodine concentration was adjusted for urinary thiocyanate concentration and their interaction, the thyroid volume was negatively correlated with urinary iodine (males P = 0.02, females P = 0.006) and positively correlated with urinary thiocyanate (males P = 0.0001, females P = 0.004). Mean thyroid volume was greater among active smokers than non-smokers (males P < 0.0001, females P = 0.0004) and was greater among former smokers than among non-smokers (males P = 0.0001, females = 0.004). Free T4 and thyroid volume were positively correlated for both sexes (P = 0. 0001). TSH was negatively correlated with thyroid volume for both groups (P = 0.0001). Female users of oral contraception (aged 35-45 years) had a smaller thyroid volume than non-users (P = 0.0009). CONCLUSIONS: The state of borderline iodine deficiency observed in France, in association with a slightly goitrogenic environment, may result in sustained stimulation of the thyroid, independently of TSH level, and is of paramount importance in the formation of goitre. Smoking may affect the thyroid, inducing marked long-lasting thyroid enlargement. PMID- 10718826 TI - Reduction of the pituitary GH releasable pool in short children with GH neurosecretory dysfunction. AB - OBJECTIVES: The classical 'GH neurosecretory dysfunction' (GHNSD) refers to slowly growing children with normal GH responses to classical provocative tests but impaired spontaneous GH secretion over 24 h frequently leading to low IGF-I levels. Thus it has been assumed that these subjects have insufficiency of spontaneous GH secretion due to neuroendocrine abnormalities in spite of a normal releasable pool of GH. However, classical provocative tests do not reliably assess the maximal somatotroph capacity; thus it is still unclear if the GH pool is really preserved or not. GHRH + arginine test is more potent than the classical tests and evaluates the maximal secretory capacity of somatotroph cells. The GH response to this stimulus is reproducible and also independent of age and puberty. DESIGN AND PATIENTS: We studied the GH response to GHRH (1 microgram/kg iv) + arginine (ARG, 0.5 g/kg iv) in 19 short children with GHNSD (14 boys and 5 girls, age: 12.1 +/- 0.7 years, pubertal stages I-III, HV-SDS between -1.6 and -4.9; GH peak > 10 micrograms/l after classical stimuli but mean GH concentration (mGHc) < 3 micrograms/l). The results in GHNSD were compared with those in 38 short children with idiopathic or organic severe GHD (GHD, 29 boys and 9 girls, age: 11.2 +/- 0.6 years, pubertal stages I-III, HV-SDS between 1.8 and -4.4; GH peak < 10 micrograms/l after 2 classical provocative tests) and in 83 children with normal or familial short stature (NC, 59 boys and 24 girls, age: 11.5 +/- 0.3 years., pubertal stages I-III; HV-SDS > 25th centile, normal IGF-I levels). RESULTS: Mean IGF-I levels in GHNSD (121.9 +/- 20.3 micrograms/l) were lower (P < 0.001) than those in NC (270.3 +/- 13.8 micrograms/l) but higher (P < 0.001) than those in GHD (72.0 +/- 4.0 micrograms/l). The mean GH concentration (mGHc) in GHNSD (2.1 +/- 0.1 micrograms/l) was lower (P < 0.01) than that in NC (4.9 +/- 0.5 micrograms/l) but higher (P < 0.01) than that in GHD (1.5 +/- 0.2 micrograms/l). On the other hand, the mean peak GH response to GHRH + ARG in GHNSD (43.7 +/- 3.7 micrograms/l) was markedly higher (P < 0.001) than that in GHD (8.2 +/- 0.9 micrograms/l) but significantly lower (P < 0.01) than that in NC (60. 4 +/- 2.7 micrograms/l). All GHD patients had peak GH responses to GHRH + ARG below the 3rd centile limit of normality (20 micrograms/l), while all GHNSD patients had peak GH responses within the normal range. No significant correlation was found between GH peak after GHRH + ARG, mGHc and IGF-I levels in each group. CONCLUSION: Our study demonstrates that short children with 'GH neurosecretory dysfunction' show reduction in the GH releasable pool evaluated by the provocative and potent GHRH + arginine test. However, the peak GH response to a single GHRH + arginine test in GH neurosecretory dysfunction is always within the normal range indicating that this test as well as classical stimuli does not distinguish normal subjects from GH neurosecretory dysfunction. PMID- 10718827 TI - A therapeutic trial of growth hormone in hypopituitary adults and its influence upon continued prescription by general practitioners. AB - OBJECTIVES: Adult GH deficiency (GHD) is associated with profound alterations in body composition, lipid profiles and quality of life which frequently improve after GH therapy. However, the beneficial effects of treatment are not derived by all and consequently some scepticism persists with regard to the use of GH therapy in adults. We assessed whether a 3-month therapeutic assessment with GH therapy could be used to determine which GHD adults should be treated over the longer term. We also assessed the continued prescription of GH by general practitioners (GPs) following the initial therapeutic assessment. DESIGN: A three month open therapeutic trial of GH in GHD adults. Patients were treated with GH at an initial dose of 0.01 iU/kg/d, increased after 1 month to 0.015 iU/kg/d for males and 0.02 iU/kg/d for females. After completion of the three months the continued prescription of GH by the GPs was assessed. PATIENTS: All adult GHD patients were considered for GH therapy. Thirty-nine GHD adults wanted GH therapy (group 1) and their baseline characteristics such as age, duration of GHD, and IGF-1 concentration were compared with 24 subjects who declined to receive GH (group 2). MEASUREMENTS: Measurements of body composition using bioelectrical impedance analysis, lipids and quality of life measured using a dedicated questionnaire were made before and after GH therapy. The response of the general practitioners to continued GH therapy after the initial therapeutic assessment was also noted. RESULTS: Compared with subjects who declined GH therapy (group 2), subjects of group 1 were younger (46.4 +/- 14.4 vs. 54.2 +/- 15.7 years, P < 0.05) and had lower peak GH responses to provocative testing (1.4 +/- 2.1 vs. 2.9 +/- 2.7 mU/l, P < 0.001), though there were no differences between IGF-1 concentration (11.7 +/- 6.2 vs. 14. 2 +/- 7.9 nmol/l). Following three months of GH therapy, there were significant improvements in all measured parameters including increased free fat mass (50.2 vs. 52.4 kg, P < 0.005) and total body water (37 vs. 38.7 l, P < 0.005), reduced fat mass (31.6 vs. 29.8 kg, P < 0.005), reduced AGHDA score (7 vs. 4, P < 0.001) and reduced cholesterol (6.3 vs. 5.8 mmol/l, P < 0.001), LDL (4 vs. 3.33 mmol/l, P < 0.001) and cholesterol/HDL ratio (5.57 vs. 4.67, P < 0.001). IGF-1 concentrations were significantly increased following treatment (12 vs. 32.4 nmol/l). Six subjects decided to discontinue GH therapy, 2 before the end of the study due to potential drug-related side-effects and 4 subjects derived no benefit from treatment. Despite the demonstrable benefits of treatment for the remaining 33 GHD adults, 6 GPs refused to continue to prescribe GH therapy for reasons of lack of familiarity with the drug or advice from their health authority. CONCLUSION: Patients who wanted GH therapy were usually younger and more severely GHD than counterparts who elect not to be treated. However, a therapeutic trial of GH therapy is required to distinguish those subjects who derive benefit from treatment. We have shown that three months of low dose GH therapy is a sufficient period to elicit significant beneficial responses in quality of life, body composition parameters and lipids for the majority of patients and appears to be a sufficient period for patients to decide whether they want longer term therapy. The initial therapeutic trial also provides the objective evidence for the general practitioners to decide upon the continued prescription of therapy. Despite the positive evidence provided by this study, a small minority of general practitioners still refuse to prescribe GH therapy. PMID- 10718828 TI - Effects of growth hormone administration on protein dynamics and substrate metabolism during 4 weeks of dietary restriction in obese women. AB - OBJECTIVE: Treatment of obesity with very low calorie diet (VLCD) is complicated by protein loss. We evaluated the effects of coadministration of GH on protein turnover, substrate metabolism, and body composition in VLCD treated obesity. DESIGN AND PATIENTS: Fifteen obese women underwent 4 weeks of very low calorie diet (VLCD) in parallel with GH treatment (n = 7) or placebo (n = 8). MEASUREMENTS: Protein metabolism and total glucose turnover were isotopically assayed. Plasma concentrations of amino acids were determined by an HPLC system. Estimated rates of lipid and glucose oxidation were obtained by indirect calorimetry. Fat free mass was determined by DEXA-scan. RESULTS: Protein breakdown decreased in both groups (tyrosine flux micromol/h): -12% +/- 3 (GH) vs. - 9% +/- 3 (placebo)). Phenylalanine degradation in relation to phenylalanine concentration decreased by 9% in the GH group, whereas an increase of 8% was observed in the placebo group (P = 0.1). Plasma concentrations of several amino acids were significantly decreased in the placebo group, while urea excretion decreased in the GH group. A decrease in FFM was found in placebo treated patients (2.14% +/- 1.9 (GH) vs. - 3.54% +/- 1.6 (placebo), P < 0.05). Rates of lipid oxidation tended to be increased by GH treatment (lipid oxidation (mg/minutes): 79.7 +/- 5.9 (GH) vs. 64.6 +/- 5.9 (placebo), P = 0.1). CONCLUSION: During dietary restriction GH primarily seems to conserve protein by a reduced hepatic degradation of amino acids. PMID- 10718829 TI - Insulin-like growth factor (IGF)-1 and IGF-binding protein-1 concentrations in serum of normal subjects after alcohol ingestion: evidence for decreased IGF-1 bioavailability. AB - OBJECTIVE: Both insulin-like growth factor (IGF)-1 and insulin-like growth factor binding protein (IGFBP)-1 are synthesized by the liver. It is well known that chronic alcohol abuse impairs liver function, but less is known about how an acute intake of moderate quantities of alcohol affects hepatic production of IGF 1 and IGFBP-1. The objective of the present investigation was to study this issue by measuring serum levels of IGF-1 and IGFBP-1 in normal subjects before, during and after ingestion of a moderate dose of ethanol. SUBJECTS AND DESIGN: Eight healthy subjects were tested on two occasions. On one occasion (experiment A), three 150-ml doses of ordinary drinking-water were given at 08.00, 09.30, and 11.00 h. On another occasion (experiment B), three 150-ml drinks of diluted ethanol were given instead of water. Each drink contained 0.45 g ethanol/kg b.w. Experiments A and B were performed in random order, 1 week apart. Blood samples were collected before, during and after the drinks over a period of 7 h (08.00 15.00 h). One blood sample was also drawn at 08.00 h the following day. MEASUREMENTS: Blood glucose and serum concentrations of ethanol, insulin, growth hormone (GH), IGF-1 and IGFBP-1 were determined. RESULTS: When alcohol had been ingested, the serum ethanol concentration rose to a peak value of 28.6 +/- 0.9 mmol/l (mean +/- SEM). The serum IGF-1 level declined significantly toward the end of the 7-h period. The serum IGFBP-1 level increased promptly in response to ethanol, and reached a maximum 2.7 times above basal as the ethanol level peaked. Neither IGF-1, nor IGFBP-1 levels changed significantly after water intake. The IGF-1 : IGFBP-1 ratio declined markedly after ethanol (from 15.7 +/- 3.9 to 3.9 +/- 0.6; P < 0.01), but not after water intake. The blood glucose and serum levels of insulin and GH were unaffected by both ethanol and water. CONCLUSIONS: Ingestion of moderate amounts of alcohol by healthy individuals results in an acute and profound increase in the serum IGFBP-1 level and a protracted and less powerful decline in the IGF-1 level. The mechanism behind the IGFBP-1 increase is probably a direct effect on the liver, since neither insulin, nor glucose or GH concentrations changed significantly in response to the ethanol challenge. PMID- 10718830 TI - Prolactinomas in adolescents: persistent bone loss after 2 years of prolactin normalization. AB - OBJECTIVE: To evaluate the effect of hyperprolactinaemia and its treatment with dopamine-agonists on bone mass and turnover in adolescent patients compared to adults. PATIENTS: Forty patients with hyperprolactinaemia (20 with disease onset during adolescence and 20 during adulthood) and 40 healthy control subjects. DESIGN: Open transverse (in patients and controls) and open longitudinal (in the patients). MEASUREMENTS: Bone mineral density (BMD) at lumbar spine and femoral neck, serum osteocalcin (OC) and urinary cross-linked N-telopeptides of type-1 collagen (Ntx) levels were evaluated in patients and controls. In the 40 patients, bone mass and turnover were re-evaluated after 12 and 24 months of treatment with bromocriptine (BRC, dose 2.5-10 mg daily), quinagolide (CV, dose 0.075-0.3 mg daily) or cabergoline (CAB, dose 0.5-1.5 mg weekly). RESULTS: Transverse study: BMD values were significantly lower in hyperprolactinaemic patients than in controls, both at lumbar spine (0.81 +/- 0.01 vs. 1.010 +/- 0.01 g/cm2; P < 0.001) and femoral neck (0.71 +/- 0.01 vs. 0.873 +/- 0.03 g/cm2; P < 0.001). Thirty-two patients (80%) had osteoporosis and/or osteopenia at one or both skeletal sites. A significant inverse correlation was found between T score values measured at lumbar spine and femoral neck and the estimated disease duration. BMD was significantly lower in young than adult patients both at lumbar spine (T score, -2.4 +/- 0.1 vs. -1.4 +/- 0.3, P < 0.01) and at femoral neck (T score, -2.1 +/- 0.05 vs. -1.5 +/- 0.2, P < 0.05). Similarly, serum OC levels were significantly lower (2.0 +/- 0.11 vs. 9.1 +/- 2.4 micrograms/l, P < 0. 01) while Ntx levels were significantly higher in patients than in controls (129.2 +/- 1.7 vs. 80.7 +/- 2.9 nmol Bone collagen equivalent (BCE)/mmol creatinine; P < 0.001). A significant inverse correlation was found between prolactin (PRL) levels and OC levels, lumbar and femoral T score values, as well as between disease duration and OC levels, lumbar and femoral T score values. A significant direct correlation was also found between Ntx levels and PRL levels and disease duration. Longitudinal study: Normalization of serum PRL levels was obtained in all patients after 6-12 months of treatment. A significant increase of serum OC levels together with a significant decrease of Ntx levels was observed after 12 and 24 months of treatment (P < 0.01). Urinary and serum calcium, phosphorus, creatinine, and serum alkaline phosphatase and parathyroid hormone levels did not change during the study period in all patients. After 12 months of therapy OC and Ntx concentrations were restored to normal. A slight but not significant increase of BMD values was recorded after 12 and 24 months of treatment. After 12 months of treatment the percent increment of BMD values in the whole group of patients was 1.13 +/- 0.6% at lumbar spine and 1.2 +/- 0.4% at femoral neck level, whereas after 24 months, it was 2.8 +/- 0.7% at lumbar spine and 3.5 +/- 0.7% at femoral neck level. After 12 months of treatment, the percent increment of BMD values was 0.7 +/- 0.2% and 1.6 +/- 1.1% at lumbar spine and 0.9 +/- 0.5% and 1.6 +/- 0.5% at femoral neck level in the young and adult patients, respectively, whereas after 24 months, it was 2.1 +/- 0.8% and 3.4 +/- 1.3% at lumbar spine and 2.6 +/- 0.8% and 4.4 +/- 1.0% at femoral neck level in the young and adult patients, respectively. CONCLUSIONS: Adolescents with prolactinoma have osteopenia or osteoporosis, a finding that strengthens the need for a prompt diagnosis. Since normalization of PRL concentrations by dopamine agonist therapy is unable to restore the bone mass, other therapeutic approaches should be considered in order to prevent further long-term problems. PMID- 10718831 TI - The effects of age and gender on parathyroid hormone dynamics. AB - BACKGROUND AND AIMS: Hyperparathyroidism is a risk factor for bone loss. An age related increase in parathyroid hormone (PTH) level has been demonstrated in several studies. It has been suggested that the type II osteoporotic syndrome, a condition of increased prevalence among elderly women, may be at least partially caused by elevations in intact parathyroid hormone (iPTH) levels. To date, however, the effects of age and gender per se on PTH dynamics in healthy subjects independent of other risk factors such as vitamin D deficiency and/or impaired renal function that can impact on parathyroid function, remain unknown. In this study, we used citrate and calcium (Ca) infusions to characterize the impact of age and gender on PTH dynamics in normal subjects. SUBJECTS AND METHODS: Twelve young women with mean age +/- SD of 26.4 +/- 1.6 years, 12 young men with mean age of 26.6 +/- 1.3 years, 12 older women with mean age of 68.6 +/- 1.3 years and 12 older men with mean age of 67.2 +/- 1.6 years were studied. The sigmoidal curves relating serum iPTH to serum levels of ionized Ca (Cai) were characterized by maximal and minimal iPTH levels, the set-points (levels of Cai causing half maximal suppression of iPTH), and the slopes of the curves at the set-points. RESULTS: Baseline serum Ca, Cai, 25 hydroxyvitamin D [25(OH)D] and 1,25 dihydroxyvitamin D [1,25(OH)2D3] levels, as well as the set-points, slopes and minimal values of the sigmoidal curves relating Cai to iPTH, did not differ among the four groups. iPTH levels at baseline were slightly but not significantly higher in the older age groups (P = 0.18). The maximal iPTH level was 25% higher in the older women than in the younger women, although this difference was not significant (P = 0.29). However, the integrated iPTH responses calculated from the areas under the curves (AUC) of iPTH levels vs. time during the calcium and citrate infusions were significantly higher in postmenopausal women than in young women during both infusions and in older men than in young men during the calcium infusion. There was no effect of gender on serum iPTH levels. CONCLUSIONS: In both women and men, ageing per se, independent of changes in vitamin D economy or renal function, is associated with an increase in integrated PTH secretory response to changes in serum calcium. No alterations in the Cai/iPTH set-point were present. The biological relevance of these modest increments in integrated iPTH levels during dynamic testing in older healthy men and women remain uncertain. PMID- 10718832 TI - Calcium-sensing receptor expression and signalling in human parathyroid adenomas and primary hyperplasia. AB - OBJECTIVE: Both in vivo and in vitro evidence indicates that primary hyperparathyroidism is characterized by a reduced sensitivity to extracellular calcium ([Ca2+]o). The existence of alterations in the expression and signalling of calcium sensing receptor (CaSR) in parathyroid neoplasia is still uncertain. In order to clarify the role of CaSR in the reduced [Ca2+]o sensing of parathyroid neoplasia we investigated PTH secretion and intracellular effectors triggered by CaSR activation as well as the levels of expression of CaSR and CaSR coupled G proteins (Gq/G11) in parathyroid adenomas and primary hyperplasia. MATERIALS AND METHODS: The study included 27 parathyroid adenomas, 4 cases of primary hyperplasia and pools of normal parathyroid biopsies. Tissues were either snap frozen in liquid nitrogen or placed in sterile medium for cell dispersion. The effects of increasing [Ca2+]o on in vitro PTH release, intracellular cAMP levels and intracellular calcium ([Ca2+]i) in cells loaded with the Ca2 + indicator fura-2 were evaluated. CaSR mRNA levels were assessed by semiquantitative RT-PCR analysis, using GAPDH as internal standard, while CaSR protein was detected by western blot analysis using a specific polyclonal antibody. Purified antisera selective for G11alpha and Gqalpha were used to detect this class of proteins. RESULTS: In basal conditions (at 0.5 mM [Ca2+]o) in vitro PTH released ranged from 29.4 to 1186 pg/well/60 minutes. Increasing [Ca2+]o from 0.5 to 1, 2.5 and 5 mM caused a variable effect. One group (n = 7) showed a significant but partial reduction of PTH release (of 17 to 60% of basal levels) that occurred at physiological [Ca2+]o concentrations (1 mM) while the remainder showed either inhibition detectable only at 2.5 mM (n = 15) or total (n = 9) resistance to [Ca2+]o. In the responsive cells, [Ca2+]o (1-5 mM) caused a pertussis toxin-insensitive [Ca2+]i rise (ranging from 10% to 260%), due to Ca2+ release from intracellular stores, and an inhibition of forskolin-stimulated cAMP levels. By RT-PCR almost all tumours tested showed a substantial reduction in CaSR mRNA levels when compared to the normal tissue (CaSR/GAPDH ratio: 3.1 +/- 0.5 vs. 15.5 +/- 3.1; P < 0.001), which was confirmed by immunoblotting analysis demonstrating low levels of CaSR protein in tumour tissues. Moreover, low amounts of G11alpha and Gqalpha, the G proteins involved in CaSR coupling, were observed in the majority of pathological tissues. CONCLUSIONS: The study shows that the activation of the calcium sensing receptors expressed in adenomatous parathyroid glands modulates intracellular effectors in a similar way to those operating in the normal parathyroid. Although a reduction of calcium sensing receptor expression is probably involved in the poor inhibition of PTH release induced by [Ca2+]o, this is not the only factor altering [Ca2+]o sensing in parathyroid adenomas, since tumours characterized by different in vitro sensitivity to [Ca2+]o showed similar CaSR levels. The low content of G proteins of the Gq subfamily might represent an additional alteration leading to a defective [Ca2+]o sensing. PMID- 10718833 TI - Increased urinary albumin excretion in Cushing's syndrome: remission after correction of hypercortisolaemia. AB - OBJECTIVES: Increased urinary albumin excretion (UAE) in diabetic and nondiabetic subjects is frequently associated with insulin resistance syndrome and central obesity. Cushing's syndrome is also characterized by central obesity and insulin resistance. This study was undertaken to see whether increased UAE is found in Cushing's syndrome. DESIGN: Cross-sectional study. PATIENTS: Thirteen consecutive patients with Cushing's syndrome. MEASUREMENTS: Patients collected three overnight urine samples for the measurement of UAE by radioimmunoassay. UAE was also measured in 479 nondiabetic subjects who comprised the control population for this study. In the patients who had initial microalbuminuria, UAE was remeasured 2 months after successful removal of pituitary or adrenal tumours. Kidney biopsy was performed in three patients during adrenalectomy. RESULTS: Eleven out of 13 patients (84.6%) had increased UAE (> 9.6 micrograms/min), and eight patients (61.5%) had microalbuminuria or overt proteinuria (> 20 micrograms/min). Kidney biopsy revealed apparently normal glomerular structures without evidence of diabetic nephropathy. After correction of hypercortisolaemia, UAE declined profoundly in all of the patients. CONCLUSIONS: More than 80% of patients with Cushing's syndrome had increased UAE. This was almost completely reversed after successful treatment of hypercortisolaemia. These results indicate that endogenous hypercortisolaemia increases UAE by a mechanism that is presently unknown. PMID- 10718834 TI - The application of a combined stimulation with CRH and desmopressin during bilateral inferior petrosal sinus sampling in patients with Cushing's syndrome. AB - BACKGROUND: Bilateral inferior petrosal sinus sampling (BIPSS) is a useful investigative technique in the differential diagnosis of ACTH-dependent Cushing's syndrome (CS). The diagnostic sensitivity of this procedure is improved by the administration of CRH to stimulate ACTH secretion. It has been reported recently that the combined administration of CRH and desmopressin is a more potent stimulus for ACTH release from corticotroph adenomas. We therefore hypothesized that the combined stimulation of ACTH secretion with CRH plus desmopressin may further improve the diagnostic outcome of this procedure. AIMS: To report our experience of the application of combined stimulation with CRH and desmopressin during BIPSS in patients with ACTH-dependent Cushing's syndrome, and to compare these results to those obtained in patients who have undergone BIPSS with CRH stimulation alone. PATIENTS: We studied 34 patients with ACTH-dependent CS: 30 with Cushing's disease (CD) and four with occult ectopic ACTH syndrome (oEAS). A combined stimulation with CRH (100 micrograms i.v.) plus desmopressin (10 micrograms i.v.) during BIPSS was performed in 15 patients with CD, while in a different group of 15 patients with CD, BIPSS was performed with CRH stimulation alone (100 micrograms i.v.). In the patients with oEAS, BIPSS was performed with CRH stimulation in three and CRH plus desmopressin in one patient. RESULTS: In patients with CD the mean peak ACTH levels from the dominant petrosal sinus samples were significantly higher in the group given a combined stimulus than in the group who had only CRH stimulation (mean +/- SD: 1649 +/- 938 vs. 692 +/- 561 ng/l, P < 0. 05). Dominant inferior petrosal sinus/peripheral (IPS/P) ACTH ratios greater than 2 were observed in 15/15 (100%) patients following the combined stimulation with CRH and desmopressin and 13/15 (87%) patients undergoing stimulation with CRH alone. No patient with oEAS had an IPS/P ratio greater than 2. It is of note that the single patient with oEAS studied following a combined stimulation during BIPSS had a IPS/P ratio of less than 2, despite a significant peripheral ACTH and cortisol response. CONCLUSIONS: A combined stimulus using CRH and desmopressin appears to induce a higher ACTH output from pituitary corticotroph adenomas during BIPSS, which may improve the diagnostic sensitivity of this procedure. PMID- 10718835 TI - Blood pressure and Turner syndrome. AB - INTRODUCTION: Elevated blood pressure (BP) is an important predictor of morbidity and mortality from cardiovascular disease. Patients with Turner syndrome (TS) have a higher morbidity and mortality in middle age than the normal population. As BP in childhood or early adulthood is predictive of BP later in adult life, we assessed manual and 24 h ambulatory BP in patients with TS to determine whether the BP pattern is altered at an early stage in these patients who are known to be at risk of cardiovascular disease. PATIENTS AND METHODS: We studied manual and 24 h ambulatory BP profiles in 75 girls with Turner syndrome, age range 5.4-22.4 years. A monitor with an oscillometric device (SpaceLabs model 90207) and an appropriate sized cuff was used. BP was measured during the day-time (0800-2000 h) and the night-time periods (2200-0800 h). The BP measured were compared with population standards. The effect of different growth promoting agents on BP was also evaluated. RESULTS: Mean manual and 24 h ambulatory BP measurements were 118/77 mmHg (range 95/60-140/102) and 115/70 mmHg (range 93/57-154/99), respectively. There was minimal difference between the two methods with a positive bias of 2.4 mmHg for diastolic BP and a negative bias of 2.1 mmHg for systolic BP. The mean standard deviation scores (SDS) corresponding to the mean BP recordings were 24 h systolic + 0. 81 (range - 1.26 to + 4.45), 24 h diastolic + 0.43 (range - 0.85 to + 3.42), day-time systolic + 1.08 (range - 0.95 to + 4.72), day-time diastolic + 0.70 (range - 0.94 to + 3.71), night-time systolic + 0. 22 (range -2.2 to + 3.64) and night-time diastolic - 0.18 (range -2. 0 to + 2.43). The SDS for both the mean 24 h and day-time systolic and diastolic BP were shifted to the right of the normal distribution. 57% of the girls had less than the normal 10% reduction in nocturnal systolic blood pressure. 17% had diastolic and 21% had systolic blood pressure above the 95th percentile for age and sex. There was no significant difference in the BP SDS between girls on no treatment and those receiving treatment. CONCLUSION: Over 50% of girls with Turner syndrome have an abnormal BP circadian rhythm, which is similar to adult patients with secondary hypertension. Patients with Turner syndrome have higher blood pressure measurements compared to published population standards, as evidenced by the shift to the right of both the systolic and diastolic BP SDS. These findings suggest that girls with Turner syndrome should be carefully monitored in childhood and adulthood for blood pressure and other cardiovascular risk factors. PMID- 10718836 TI - The influence of renal and cardiovascular abnormalities on blood pressure in Turner syndrome. AB - INTRODUCTION: Patients with Turner syndrome (TS) are at an increased risk of morbidity and mortality from cardiovascular disease. This study was undertaken to establish the prevalence of hypertension in patients with TS and to establish to what extent cardiovascular or renal abnormalities contribute to the measured blood pressure. PATIENTS AND METHODS: 62 patients with TS, age 5.4-22.4 years, had 24 h-ABPM (ambulatory blood pressure monitoring), echocardiography, renal imaging and measurement of recumbent plasma renin activity (PRA). Blood pressure was compared with population standards. RESULTS: 21% of the TS study population had mean systolic and 17% mean diastolic 24 h-ABPM measurements above the 95th percentile for age and sex (i.e. mild hypertension). Borderline blood pressure (i.e. 90th to 95th percentile) was found in another 17% of the patients. 57% of the patients had a blunted (i.e. less than 10%) fall in the night-time blood pressure. 24% of the patients had a detectable cardiac abnormality, 42% a detectable renal abnormality and 52% were found to have raised plasma renin activity. The presence of a cardiac or renal abnormality had no significant effect on blood pressure. Blood pressure of patients on growth and/or pubertal therapy was not different from those patients on no such treatment. CONCLUSION: Over 30% of patients with Turner syndrome were found to be mildly hypertensive and over 50% had an abnormal diurnal blood pressure profile. In this study we were unable to demonstrate that the presence of renal or cardiac abnormalities had an effect on recorded blood pressure. The use of growth hormone and oestrogen to manage growth failure and pubertal delay did not seem to affect blood pressure. This study suggests that there is a high prevalence of raised blood pressure in Turner syndrome patients. The 24 h-ambulatory blood pressure monitoring profile suggests that this may be secondary in origin, but we were unable to demonstrate an underlying mechanism with the renal and cardiac investigations performed. PMID- 10718837 TI - Relapse of hirsutism following long-term successful treatment with oestrogen progestogen combination. AB - OBJECTIVE: While several forms of treatment have been reported to be successful in relieving hirsutism over periods of 6-12 months, there is little if any information on the long-term outcome of hirsutism following the withdrawal of successful treatment. The combination of ethinyl oestradiol, 35 micrograms, and cyproterone acetate, 2 mg (EE-CA) for 21 days followed by 7 days without treatment is widely used in a cyclical manner in the treatment of hirsutism. The present study was undertaken to evaluate the outcome of withdrawal of long-term successful treatment of hirsutism with EE-CA. DESIGN, PATIENTS AND MEASUREMENT: In this retrospective study the clinical records of 57 patients with idiopathic hirsutism or polycystic ovary syndrome who had been treated with EE-CA were reviewed. The degree of hirsutism had been assessed by the Ferriman and Gallwey scoring system (FG). The testosterone/sex hormone binding globulin ratio (T/SHBG), was derived prior to and following the introduction of treatment with EE-CA. RESULTS: Fifty-two of the 57 patients achieved a satisfactory clinical response. In the group of patients who were satisfied with the outcome of treatment, FG decreased from 12.9 +/- 3.6 to 5.5 +/- 2.5 and T/SHBG ratio decreased from 11.3 +/- 9.5 to 1 +/- 0.8 (reference range: 1-5.2). The duration of treatment prior to its withdrawal in this group was 28.2 +/- 13.7 months. The five patients who were not satisfied with the response abandoned treatment after 16 +/- 2.8 months; the pretreatment FG was 16.2 +/- 8.3, while the T/SHBG decreased from 6.1 +/- 3.1 to 1.1 +/- 0.6 in these patients. Subsequent follow-up data, after withdrawal of treatment, were available on 34 of these 52 patients. Twenty-eight of the 34 patients exhibited relapse of hirsutism after 6.15 +/- 2.8 months. Six patients did not relapse during a follow-up period of 18.8 +/- 7. 8 months. The six patients who did not relapse were treated for a significantly longer period than the group who relapsed, 40 +/- 6.9 and 26.1 +/- 8.3 months, respectively, P < 0.01. However, the groups did not differ significantly when examined for pretreatment FG, 11.5 +/- 3.8 and 13.2 +/- 3.6 and pretreatment T/SHBG 8.9 +/- 5 and 13.4 +/- 11.9. CONCLUSION: These data indicate that ethinyl oestradiol and cyproterone acetate achieved a satisfactory clinical outcome in the treatment of hirsutism in 90% of patients. However, on withdrawal of treatment after a mean duration of over 2 years, relapse occurred in 80% of these patients after a mean of 6 months. If it is assumed that the successfully treated patients lost to follow-up all maintained long-term remission, the relapse rate is still an unsatisfactory 65% at 6 months. These are disappointing results which indicate for the first time that successful outcome requires that treatment be maintained for several years. Patients embarking upon treatment for hirsutism should be advised that maintenance of reduced hair growth requires long-term treatment, probably for at least 3-4 years. PMID- 10718838 TI - Identification of missense mutations in the SRD5A2 gene from patients with steroid 5alpha-reductase 2 deficiency. AB - BACKGROUND AND OBJECTIVE: Mutations of the steroid 5alpha-reductase type 2 (SRD5A2) gene in karyotypic males result in a spectrum of external genitalia phenotypes ranging from complete female to nearly complete male. Here we performed genomic DNA analyses from individuals bearing the enzyme deficiency in order to detect the molecular abnormalities. PATIENTS: Four unrelated 46,XY patients of Mexican origin with ambiguous external genitalia were studied. A fertile, phenotypically normal male was also included. MEASUREMENTS: Coding sequence abnormalities of the SRD5A2 gene were assessed by exon-specific polymerase chain reaction, single-stranded conformational polymorphism and sequencing analysis. RESULTS: Five different missense mutations (two of them novel mutations) were identified. Three subjects presented homozygous single base mutations. These were located at exon 2 (G115D), exon 4 (P212R) and exon 5 (R246Q), and such changes have been described previously. The fourth patient was a compound heterozygote who presented two mutations located in exons 1 and 2. We found a hitherto unreported G --> A transition at the second nucleotide of codon 85 in exon 1 (GGC --> GAC), substituting glycine for aspartic acid (G85D). This patient also presented an identical alteration at codon 115 of exon 2, which was carried by his father (G115D). Finally, in another subject who was included originally as a control, we found a C --> A transversion (yet undescribed) at codon 245 in exon 5 (S245Y). CONCLUSIONS: Four different single base mutations that cause amino acid substitutions were detected in the steroid 5alpha-reductase type 2 gene of affected individuals. One patient and a normal control had two previously undescribed mutations. Although in the latter individual we cannot exclude the possibility that the base change is a genetic polymorphism, the molecular screening of 100 chromosomes suggests strongly that the change at codon 245 does represent a heterozygous mutation. Further studies, including the recreation of the mutations, will help to reveal the biochemical consequences resulting from these changes. PMID- 10718839 TI - Pseudohypoparathyroidism--another monogenic obesity syndrome. AB - Obesity is a common feature of pseudohypoparathyroidism (PHP) type 1a, but is usually associated with short stature. We describe two children referred because of hyperphagia and excessive weight gain from early infancy. Tall stature in both children initially confounded the diagnosis of PHP, but on follow-up both children developed the typical hormonal abnormalities and Case 2 developed typical skeletal features of Albright hereditary osteodystrophy. PHP type 1a is caused by germline loss of function mutations in the alpha subunit of GS, the ubiquitously expressed G protein that couples many hormone receptors to the adenylate cyclase second messenger system. Recent evidence suggest that the hypothalamic GS protein coupled melanocortin-4 receptor (MC4R) may mediate the central effects of leptin on inhibition of satiety. Similar patterns of infancy onset hyperphagia, excessive weight gain and tall stature are seen in subjects with congenital leptin deficiency and in subjects with MC4R mutations. We suggest that the genetic mutations in GSalpha which underlie PHP type 1a may also directly result in severe obesity. This diagnosis should be considered in any child with a history of hyperphagia and early onset morbid obesity. PMID- 10718840 TI - Mast cell matrix interactions. PMID- 10718841 TI - Occupational latex allergy: the magnitude of the problem and its prevention. PMID- 10718842 TI - Nedocromil sodium: a review of the evidence for a dual mechanism of action. PMID- 10718843 TI - Clara cell secretory protein (CC16): characteristics and perspectives as lung peripheral biomarker. AB - Clara cell protein (CC16) is a 15.8-kDa homodimeric protein secreted in large amounts in airways by the non-ciliated bronchiolar Clara cells. This protein increasingly appears to protect the respiratory tract against oxidative stress and inflammation. In vitro, CC16 has been shown to modulate the production and/or the activity of various mediators of the inflammatory response including PLA2, interferon-gamma and tumour necrosis factor-alpha. CC16 has also been found to inhibit fibroblast migration or to bind various endogenous or exogenous substances such as polychlorobiphenyls (PCBs). This protective role is confirmed by studies on transgenic mice, showing that CC16 deficiency is associated with an increased susceptibility of the lung to viral infections and oxidative stress. In humans, a polymorphism of the CC16 gene, localized to a region linked to airway diseases, has recently been discovered in association with an increased risk of developing childhood asthma. Finally, CC16 also presents a major interest as a peripheral marker for assessing the integrity of the lung epithelium. The determination of CC16 in serum is a new non-invasive test to detect Clara cell damage or an increased epithelial permeability in various acute and chronic lung disorders. PMID- 10718844 TI - Putative virulence factors of Aspergillus fumigatus. AB - Various putative virulence factors of Aspergillus fumigatus have been studied over the past decades. A. fumigatus gliotoxin is a potent inhibitor of the mucociliary system. Several fungal metabolites interfere with phagocytosis and opsonization including toxins, 'conidial inhibitory factor', 'A. fumigatus diffusible product' and 'complement inhibitory factor'. A. fumigatus can bind specifically to different host tissues components, whereas toxins give a general and significant immunosuppressive effect on host defences. Circumstantial evidence links the production of elastinolytic proteases with the ability to cause disease. However, none of the reports demonstrates conclusively a decisive role for any of the virulence factors described thus far. It is conceivable that proteolytic enzyme activities such as those expressed by AFAlp are one of a number of factors, each with a minor effect, that combine to facilitate disease progression. PMID- 10718845 TI - Activation of fibroblasts in collagen lattices by mast cell extract: a model of fibrosis. AB - BACKGROUND: Mast cells are resident connective tissue cells able to secrete numerous inflammatory mediators in response to tissue aggression and might be implicated in the fibrotic processes. OBJECTIVES: To study the effects of mast cell products on fibroblast activity in connective tissues. METHODS: Mast cell extract was prepared by sonication of pure mast cell preparations obtained by peritoneal lavage of rats and added to the culture medium of fibroblast-populated collagen lattices. RESULTS: Mast cell extract was able to decrease the contraction of the collagen lattices, to stimulate total protein and collagen synthesis, and to increase the expression and activation of gelatinase A/MMP-2. CONCLUSION: These data are consistent with the hypothesis that mast cells in connective tissue may be responsible for fibroblast activation at the early phases of tissue repair and fibrosis. PMID- 10718846 TI - Localization of interleukin (IL) -4 but not IL-5 to human mast cell secretory granules by immunoelectron microscopy. AB - BACKGROUND: Human mast cells synthesize and secrete many cytokines of relevance to the pathogenesis of allergic diseases such as asthma and rhinitis. In particular, interleukin (IL) -4 and IL-5 are likely to play key roles in the development of the inflammatory response that characterizes these diseases. Immunohistochemical studies on human nasal and bronchial mucosal biopsies suggest that IL-4 and IL-5 may be stored preformed in mast cells. OBJECTIVE: To identify whether IL-4 and IL-5 are stored within mast cell secretory granules. METHODS: We used immunogold electron microscopic analysis on bronchial mucosa and lung parenchyma from resected lung specimens, and a nasal mucosal biopsy from a patient with active allergic rhinitis. Samples were fixed in 4% paraformaldehyde plus 0.5% glutaraldehyde and processed into Lowicryl K4M resin by the 'Progressive Lowering of Temperature' technique. Ultrathin sections were stained immunohistochemically by an indirect immunogold method. RESULTS: Immunoreactivity for IL-4, but not IL-5, was localized to the granules of mast cells in all tissue samples. IL-5 was localized to the matrix of eosinophil granules in these samples, but neither cytokine was detected in T cells. IL-4 immunoreactivity increased in the granules of mast cells 24 h after immunoglobulin (Ig) E dependent activation (mean 17.5 +/- 1.4 gold particles per granule) compared with nonactivated mast cells (mean 6.8 +/- 0.8 gold particles per granule, P < 0.001), suggesting replenishment of stores by newly generated protein. Immunoreactive IL 5 remained undetectable in mast cells 24 h after activation, a time point at which they are known to secrete large quantities of this cytokine. CONCLUSION: Human mast cells store IL-4 within the matrix of their granules. Very few, if any, lung or nasal mast cells store IL-5. A store of preformed IL-4 within mast cell granules is likely to have an important influence during the initiation and maintenance of the allergic immunological response. PMID- 10718847 TI - Effects of luteolin, quercetin and baicalein on immunoglobulin E-mediated mediator release from human cultured mast cells. AB - BACKGROUND: Flavonoids have a variety of activities including anti-allergic activities, and are known to inhibit histamine release from human basophils and murine mast cells. OBJECTIVE: The effects of luteolin, a flavone, on the immunoglobulin (Ig) E-mediated allergic mediator release from human cultured mast cells (HCMCs) were investigated and compared with those of baicalein and quercetin. METHODS: HCMCs were sensitized with IgE, and then treated with flavonoids before challenge with antihuman IgE. The amount of released mediators was determined as was mobilization of intracellular Ca2+ concentration, protein kinase C (PKC) translocation and phosphorylation of intracellular proteins were detected after anti-IgE stimulation. RESULTS: Luteolin, baicalein and quercetin inhibited the release of histamine, leukotrienes (LTs), prostaglandin D2 (PGD2), and granulocyte macrophage-colony stimulating factor (GM-CSF) from HCMC in a concentration-dependent manner. Additionally, the three flavonoids inhibited A23187-induced histamine release. As concerns Ca2+ signalling, luteolin and quercetin inhibited Ca2+ influx strongly, although baicalein did slightly. With regard to PKC signalling, luteolin and quercetin inhibited PKC translocation and PKC activity strongly, although baicalein did slightly. The suppression of Ca2+ and PKC signallings might contribute to the inhibition of mediator release. The activation of extracellular signal-regulated kinases (ERKs) and c-Jun NH2 terminal kinase (JNK), that were activated just before the release of LTs and PGD2 and GM-CSF mRNA expression in IgE-mediated signal transduction events, were clearly suppressed by luteolin and quercetin. In contrast, the flavonoids did not affect the activation of p38 mitogen-activated protein kinase (p38 MAPK) pathway. CONCLUSION: These results indicate that luteolin is a potent inhibitor of human mast cell activation through the inhibition of Ca2+ influx and PKC activation. PMID- 10718848 TI - Prevention of latex allergy by selection of low-allergen gloves. AB - BACKGROUND: In recent years the prevalence of type I allergy to latex has continuously increased, in particular among healthcare workers, to about 10%. While most forms of type I allergy caused by other environmental allergens can be treated by pharmacotherapy or specific immunotherapy, minimizing exposure to latex proteins may represent an effective preventive measure for latex allergy. OBJECTIVE: To investigate whether it is possible to select by in vitro and in vivo testing low-allergen latex gloves for prevention of latex allergy. METHODS: We obtained separate extracts by standard aqueous extraction from the inner and outer surfaces of 15 different commonly used (10 examination, five surgical) glove brands. The extracts were analysed by quantitative (bicinchoninic protein assay, immunoglobulin [Ig] E-ELISA, ELISA competition) and qualitative (SDS-PAGE, silver staining, IgE immunoblotting) methods for their protein and allergen contents. In addition, the glove extracts were analysed for their capacity to induce basophil histamine release and immediate skin reactions. RESULTS: Extracts from different glove brands contained cross-reactive IgE epitopes. However, IgE binding studies, basophil histamine release and skin testing showed that different glove brands and their inner and outer surfaces contained widely varying protein and allergen contents. While the determination of total protein contents was not sufficient to identify low-allergen gloves, IgE measurements, basophil histamine release and skin testing were in good agreement and allowed to select low-allergen products. CONCLUSION: We suggest the use of low-allergen latex products identified by IgE binding, basophil histamine release assays and skin testing as a feasible preventive measure for latex allergy. PMID- 10718849 TI - Ethnicity, childhood environment and atopic disorder. AB - BACKGROUND: The International Study of Asthma and Allergies in Childhood (ISAAC) has demonstrated large differences in the prevalence of atopic disorders in children between different regions in the world. Populations with a higher standard of living and a more westernized lifestyle tend to have higher rates of atopy and asthma. Many hypotheses regarding environmental causes of atopic disorder focus on the early childhood environment. OBJECTIVE: To study the influence of ethnicity and country of birth for the prevalence of atopic disorders. METHODS: The prevalence of atopic disorders in Swedish residents born in Turkey and Chile, who settled in Sweden as adults in the 1980s, was compared with their own Swedish-born children and a sample of Swedish-born parents and their children in interview data from the Survey of Living Conditions in 1996. The study group included 1734 adults 27-60 years of age and their 2964 children aged 3-15. RESULTS: The Chilean-born parents and their children had the highest risk for allergic asthma; adjusted odds ratios (ORs) 2.2 (1.2-4.0) and 2.7 (1.6 4.5), respectively, and allergic rhino-conjunctivitis; OR 1.6 (1.1-3) and 1.6 (1.1-2.5) in both groups, when compared with the Swedish-born parents and their children. The Turkish-born parents and their children had the lowest risk for allergic rhino-conjunctivitis; both groups had OR 0.6 (0. 4-0.9) and the children in this group also had the lowest risk for eczema; OR; 0.4 (0.3-0.7). The risk for all atopic disorders was lower in the Turkish group compared with the Chileans. CONCLUSION: This study demonstrates that ethnicity is an important determinant of atopic disorder independent of the external childhood environment. The value of international comparisons of environment and risk for atopic disorders can be questioned until more is known about factors related to ethnicity, such as genetic susceptibility and diet, for the development of atopy. PMID- 10718850 TI - Exposure and allergic sensitization to cockroach allergen in East Germany. AB - BACKGROUND: Some studies suggest that the prevalence of sensitization to cockroach allergens may be higher in the United States than in Europe, but there are no comparable data from population-based studies. OBJECTIVES: To determine the prevalence of allergic sensitization to German cockroach (GCR) in German schoolchildren and to assess its clinical relevance; and to determine the exposure to the major GCR allergen Bla g 2 in non-selected homes and nurseries. METHODS: The prevalence of allergic sensitization to GCR and other allergens was determined by measurement of specific IgE and skin-prick tests in a cross sectional study of 2993 children aged 5-11 years in Dresden, Germany. The prevalence of atopic disease was determined by questionnaire, and pulmonary function and bronchial hyperresponsiveness to hypertonic saline were measured. Bla g 2 exposure was determined on floors of 187 kitchens and 47 nurseries by a commercial sandwich ELISA. RESULTS: One hundred and twenty-seven (4.2%) of the children had specific IgE (> 0.7 kU/L) against GCR. Among children with current wheeze, 8.4% were GCR-sensitized. Compared to data from the United States, the prevalence of sensitization to cockroach was similar in children without asthma (3.9%), but less frequent in asthmatic children from Dresden (6.1%). After adjustment for positive reactions to other allergens (SX1 test) no significant impact of GCR sensitization on wheeze or other symptoms and diagnoses was found. Bla g 2 was detected in 29% of the kitchens and 43% of the nurseries. None of these sites had exposure levels above the proposed threshold for causing disease of 80 ng/g dust. CONCLUSION: The data suggest that allergic sensitization to GCR is less frequent in asthmatics from Dresden, Germany than in US cities. The data indicate that GCR sensitization is not an independent risk factor for asthma and other atopic diseases in 5-11-year-olds from this city. PMID- 10718851 TI - HLA DPB1*0201 allele is negatively associated with immunoglobulin E responsiveness specific for house dust mite allergens in Taiwan. AB - BACKGROUND: House dust mite (HDM) Dermatophagoides pteronyssinus is the most important source of indoor allergens that cause allergic diseases in Taiwan. We prepared purified HDM allergens (Der p 1, Der p 2 and Der p 5) to detect allergen specific immunoglobulin (Ig) E responsiveness among a large number of test subjects. The robust genetic typing system for HLA class II genes also facilitated the study on association of HLA and allergic response toward HDM. OBJECTIVE: This study intended to investigate the association between HLA class II alleles and the IgE responsiveness to the major allergens from HDM, D. pteronyssinus. METHODS: Two hundred and forty-eight subjects were selected for HLA association study. Plasma HDM allergen (Der p 1, Der p 2, Der p 5) -specific IgE and Der p 2-specific IgG antibodies were detected by ELISA, while HLA class II -DRB1, -DQA1, -DQB1, -DPB1 genetic polymorphism was determined by polymerase chain reaction/sequence-specific oligonucleotide probe hybridization (PCR/SSOPH). Statistical comparison of the allelic distribution of each HLA class II genes among the individuals with/without HDM allergen-specific IgE and IgG antibodies were performed. RESULTS: There was no significant association between HLA DRB1, DQB1, DQA1 alleles and HDM-specific IgE responsiveness noted. Only DRB1*0803 and the linked DQA1*0103 alleles showed positive association with Der p 5-specific IgE responsiveness. However, we found that HLA-DPB1*1301 predisposed subjects to IgE responsiveness to HDM Der p 5. HLA DPB1*0501 was weakly associated with the IgE responsiveness to HDM Der p 1 and Der p 5. There was a strong negative association between the HLA-DPB1*0201 allele with IgE responsiveness to Der p 1 (OR: 0.30, P 0.05) but there were four postoperative deaths. There was no significant difference in the length of stay. CONCLUSIONS: Conservative management of postoperative adhesional intestinal obstruction is safe and 65 per cent settle. In those who fail to settle there is no significantly increased risk of bowel strangulation. There is no way of identifying those who will not settle from the history or initial investigations on admission. PMID- 10718952 TI - Early outcome after laparoscopic and open floppy nissen fundoplication AB - AIMS: There remains debate regarding the best operative procedure for gastro oesophageal reflux. For those who advocate a full 'floppy' Nissen fundoplication there is some scepticism that this can be completed effectively laparoscopically with division of all the short gastric vessels. This study therefore compared the results of patients operated on in two different hospitals in which floppy Nissen fundoplication was carried out either laparoscopically (hospital 1) or by open surgery (hospital 2). METHODS: All patients undergoing antireflux surgery in both hospitals were recorded prospectively and perioperative data were collected retrospectively from the case records. A postoperative questionnaire was sent to each patient to determine the modified DeMeester score and the impact of surgery on daily activities, which were then analysed anonymously by an independent clinician. RESULTS: Operating time was significantly longer (median 150 versus 75 min; P < 0.001), but postoperative stay (2 versus 4 days; P < 0.001) and return to normal activities (14 versus 25 days; P < 0.002) were shorter for the laparoscopic approach. There was no difference in gas bloat (both groups median 1.0, range 0-3), dysphagia (median 1.0 versus 0.0, range 0-3) or modified DeMeester score (both groups median 1.0, range 0-9) between the two techniques (median follow-up 10 and 15 months). CONCLUSIONS: As the postoperative results were comparable this study confirms that a 'floppy' Nissen fundoplication with complete mobilization of the gastric fundus can be performed laparoscopically. The advantages of a shorter hospital stay and earlier return to normal activities are offset by a longer operating time. PMID- 10718953 TI - Telementoring in laparoscopic cholecystectomy: a useful adjunct in training and assessment of higher surgical trainees AB - AIMS: This study assessed the feasibility, safety and utility of telementoring as a training tool in laparoscopic cholecystectomy (LC) for higher surgical trainees (HST). Telementoring in LC was developed as a technique for objective assessment of performance and progress in order to help decide when an HST is competent to perform LC unsupervised. METHODS: This was a prospective study of patients attending for LC, with surgery performed by an HST, in which the laparoscopic image was relayed live to an adjoining room where a supervising consultant observed the procedure. The trainee or supervisor sought or proffered advice/assistance as appropriate, and safety, complication rates, operating time, difficulty of procedure, intervention rate and type were recorded. RESULTS: LC was accomplished in 33 of 34 patients enrolled; there was one conversion to an open procedure. Interaction between the HST and trainer occurred in 11 cases, by way of advice being sought or offered, and in two of these the supervisor scrubbed up and took over the operation. The rate of interaction and duration of the procedure increased with procedure difficulty, with rates of interaction for difficulty grades 1, 2 and 3 of 15 per cent (two of 13), 41 per cent (seven of 17) and 50 per cent (two of four), and overall median operating times of 35 (23 50), 45 (28-75) and 92 (45-110) min respectively. CONCLUSIONS: Telementoring for trainees capable of performing LC is feasible and appears to be safe with a supervising surgeon situated in the immediate vicinity. Telementoring in LC is a promising tool that may provide objective assessment of a trainee's insight, skill and progress in operative performance. PMID- 10718954 TI - Increase in the incidence of oesophagogastric carcinoma in the south thames region: an epidemiological study AB - Aims: There is some evidence to indicate that the incidence of gastric and oesophageal carcinoma is changing rapidly in the West. However, to study the true magnitude of the problem (to help in planning health and cancer strategies over the next decades), this epidemiological study was performed. METHODS: The Thames Cancer Registry database for the years 1960-1995 was searched for all oesophageal and gastric cancers in the South Thames region, and was analysed for all subsites for both sexes separately. Statistical analysis using the age-standardized ratio (ASR) (world) and chi2 test for trend for each subgroup was performed. RESULTS: The overall incidence of carcinoma of the oesophagus for both sexes saw a marked rise in the period studied, while the overall incidence of gastric cancer fell. In the upper two-thirds of the oesophagus, the increase in incidence was more dramatic for women (from 0.30 in 1960 to 0.70 in 1996; chitrend2 = 18.7, P = 0.00015) than men (from 0.59 in 1960 to 1.09 in 1996; chitrend2 = 4.4, P = 0.0354). In the lower third of the oesophagus, there was a rise in men (ASR from 0.93 in 1960 to 3. 43 in 1996; chitrend2 = 466.3, P < 0.0001) as well as in women (ASR from 0.42 in 1960 to 0.91 in 1996; chitrend2 = 51.4, P < 0.0001). For the gastric cardia, the incidence in men increased (ASR from 1. 24 in 1960 to 2.41 in 1996; chitrend2 = 109.8, P < 0.0001), while in women it remained unchanged (ASR from 0.33 in 1960 to 0.55 in 1996; chitrend2 = 0.0, P = 0.8281). The incidence of distal gastric carcinoma has decreased in men (ASR from 2.47 in 1980 to 0.63 in 1996; chitrend2 = 264.7, P < 0.0001) and women (from 1.14 in 1980 to 0.43 in 1996, chitrend2 = 131.8, P < 0.0001). For cancers of other subsites of the stomach (fundus, lesser and greater curvature, and body), there was a fall in incidence in both sexes. CONCLUSION: There has been a twofold to threefold increase over 36 years in the incidence of cancer of the oesophagogastric junction with a decreasing incidence in the distal stomach. There was also an increase in the incidence of upper oesophageal cancers, and a decrease in the incidence of cancers of other subsites of the stomach. Studies to cope with this increase and resources to cope with the treatment of these patients are required. PMID- 10718955 TI - Management of acute pancreatitis: a comparative audit of clinical practice against the recommendations of the british society of gastroenterology AB - AIMS: In 1998 the British Society of Gastroenterology (BSG) published national guidelines for the management of acute pancreatitis (AP) in an attempt to improve diagnosis and reduce mortality rates. A comparative audit was undertaken of the management of AP against the BSG recommendations. METHODS: A retrospective analysis of 53 patients (median age 61 (range 24-95) years) admitted with AP during 1998 was undertaken and a comparison was made with the BSG guidelines. RESULTS: Some 70 per cent (n = 37) of the patients were admitted with mild AP and 30 per cent (n = 16) with severe AP. The BSG recommendations are shown in parentheses in the following text. The overall mortality rate was 17 per cent (less than 10 per cent), zero in mild AP and 56 per cent in patients with severe AP (less than 30 per cent). A correct diagnosis of AP was made within 48 h of admission in all patients (100 per cent). Severity stratification within 48 h using the Glasgow criteria and C-reactive protein (CRP) level, as suggested by the BSG, was done in approximately 80 per cent of patients. Table 1. CRP was measured in 51 per cent of the patients within the first 4 days (100 per cent) and at the end of the first week (100 per cent). Gallstones and alcohol counted for 72 per cent of causes of AP and the aetiology was determined in 77 per cent (75-80 per cent). All patients with severe AP were managed in a high-dependency or intensive therapy unit with central venous pressure monitoring and prophylactic intravenous antibiotics, and underwent dynamic computed tomography within 3-10 days of admission (100 per cent). Some 86 per cent of patients with suspected common bile duct (CBD) stones (jaundice, deranged liver function tests, dilated CBD) underwent endoscopic retrograde cholangiopancreatography with or without duct drainage and clearance. CONCLUSIONS: Management of AP closely mirrors the BSG guidelines, but fails to fully address severity stratification with respect to LDH, CRP and PaO2 at 48 h. A thorough analysis of patients with severe AP (i.e. extent of pancreatic necrosis, onset of infection) has been undertaken to explain the mortality rate and it is proposed to prospectively audit admissions with AP in 1999 in order to close the audit loop. PMID- 10718956 TI - Symptoms are a poor indication of severity of reflux in Barrett's oesophagus AB - AIMS: Patients with Barrett's oesophagus have increased acid and duodenogastric reflux and impaired motility compared with non-Barrett's patients with reflux disease. Impaired sensitivity to acid infusion and distension have also been described, but the relationship of this visceral response to symptoms is unclear. A symptom index was used to compare Barrett's and non-Barrett's patients with reflux. METHODS: Patients with reflux (DeMeester score above 14) were studied with 24-h pH monitoring and manometry. An event marker recorded symptom events. An event was positive if it corresponded to a period greater than 10 s within 2 min either side of the drop in pH. The symptom index was calculated as the number of symptoms with pH less than 4/total number of symptoms x 100. RESULTS: Eighteen patients with Barrett's oesophagus were compared with 58 non-Barrett's patients with significant reflux. CONCLUSIONS: Patients with Barrett's oesophagus have a low symptom index compared with non-Barrett's patients with reflux disease. This occurs despite a near 100 per cent increase in acid exposure in the Barrett's group. Symptoms are thus no guide to the severity of reflux in patients with Barrett's oesophagus. Proof of efficacy of therapeutic modalities may need physiological rather than symptom-based confirmation. PMID- 10718957 TI - Acute severe pancreatitis and multiple organ failure: total parenteral nutrition is still required in a proportion of patients AB - AIMS: Enteral nutrition (EN) is increasingly advocated as the favoured means of nutritional support in patients with multiple organ failure resulting from acute severe pancreatitis. In a proportion of patients, however, EN may not be feasible either because of gastrointestinal stasis and high nasogastric aspirates or because of superseding complications of pancreatitis such as fistulation. In these cases total parenteral nutrition (TPN) is indicated. METHODS: Patients with acute severe pancreatitis admitted to the intensive care unit (ICU) were commenced on either nasogastric or, where established, jejunostomy EN. Rates of EN were controlled by protocol and volumes of nasogastric aspirate. TPN was commenced when volumes of EN were persistently inadequate or when this route was contraindicated (e.g. high-volume small bowel fistula). Physiological and outcome data were collected prospectively and included severity of illness scoring (Acute Physiology and Chronic Health Evaluation (APACHE) II), length of stay, duration of EN and TPN, and hospital outcome. RESULTS: Of 69 patients with acute severe pancreatitis, 31 were transferred from other ICUs. The median APACHE II score was 18 (range 4-40) and the overall hospital mortality rate was 39 per cent. Seventeen patients (25 per cent) were managed with EN alone, ten (14 per cent) with TPN alone and 19 (28 per cent) with a combination of both. Twenty-three (33 per cent) did not have any nutritional support during their ICU stay. The mortality rate was worse among patients who received TPN only than in those who had EN (60 versus 24 per cent). CONCLUSIONS: EN is likely to remain the route of choice for nutritional support in the ICU. However, in high-risk patients with acute severe pancreatitis EN may not be feasible and TPN is often required. This may reflect a greater severity of illness. PMID- 10718958 TI - Long-term symptomatic follow-up after lind fundoplication AB - AIMS: Few published studies have detailed the long-term results of antireflux surgery. The aim of this study was to assess the long-term success of open Lind fundoplication in controlling the symptoms of gastro-oesophageal reflux disease. METHODS: One hundred and thirty-two patients with reflux symptoms underwent a primary Lind fundoplication between April 1986 and February 1994; all operations were supervised or performed by one surgeon. The median age at operation was 47 (range 17-77) years. All patients attended for follow-up in the early postoperative period. It was possible to conduct a telephone interview to assess long-term symptom control, at a median time of 9.5 (range 5-13) years following operation, in 112 of the 124 patients who were still alive. RESULTS: Ninety-one patients underwent oesophageal pH studies before and soon after operation. The oesophageal pH was less than 4 for a mean 14.9 per cent of the time before operation, falling to 2.4 per cent in the early postoperative period (P < 0.001, Wilcoxon test). At early postoperative assessment, two patients complained of mild reflux symptoms and 44 (33 per cent) complained of postfundoplication symptoms (dysphagia, epigastric bloating and early satiety). At telephone interview, 106 patients (95 per cent) were symptom free with regard to heartburn and regurgitation. Six patients have developed recurrent reflux symptoms, in four of whom symptoms are controlled by a proton pump inhibitor. Two patients have required further antireflux surgery, one within 2 months of the first procedure for severe dysphagia and the other for recurrent reflux. Significant postfundoplication symptoms persist (dysphagia with or without gas bloat) in three patients (3 per cent). CONCLUSIONS: Open Lind fundoplication appears to be effective in the long-term control of gastro-oesophageal reflux in 95 per cent of patients and represents a standard against which the long-term results of laparoscopic surgery will need to be compared. PMID- 10718959 TI - Benign anastomotic stricture following transthoracic subtotal oesophagectomy and stapled oesophago-gastrostomy: risk factors and management AB - AIMS: Benign anastomotic stricture (BAS) is a common cause of dysphagia following oesophagectomy. The aim of this study was to assess the incidence of BAS, identify risk factors for its development and evaluate the results of postoperative endoscopic dilatation. METHODS: A consecutive series of 234 patients undergoing oesophagectomy with a stapled intrathoracic oesophagogastric anastomosis (Autosuture CEEA gun) between April 1990 and April 1999 were studied. BAS was defined as dysphagia with anastomotic narrowing (XQ200 endoscope) and no suspicion of recurrence. Statistical analysis was by the chi2 and Mann-Whitney U tests. RESULTS: The postoperative mortality rate was 5 per cent (12 of 234) and the anastomotic leak rate 3 per cent (six of 234). One-third of patients (70 of 222) who survived surgery re-presented with dysphagia; 5 per cent (11 of 222) were found to have proven local recurrence and 27 per cent (59 of 222) BAS. The median time to development of BAS was 92 (range 24-210) days. BAS formation was significantly related to the size of the staple gun employed: 21 mm, 80 per cent (eight of ten); 25 mm, 32 per cent (25 of 78); 28 mm, 23 per cent (19 of 81); 31 mm, 19 per cent (seven of 37) and 34 mm, 0 per cent (none of five) (chi2 = 18.3, 4 d.f., P < 0.01). All patients underwent radiographically controlled endoscopic dilatation with no complications. Recurrent BAS occurred in over half of these patients (32 of 59), who had a median of 2 (range 2-17) recurrences all resolving within 532 (mean interval 68) days. Recurrent BAS formation was also significantly related to the size of the staple gun employed (chi2 = 11.6, 4 d.f., P < 0.05). Following the introduction of the Autosuture 'tilt-top' device in July 1995 the median size of gun used rose from 25 to 28 mm with an overall decrease in the incidence of BAS from 33 to 21 per cent (chi2 = 4.0, 1 d.f., P < 0.05). All patients with anastomotic leaks survived and none subsequently developed BAS. Similarly, no association was found between the development of BAS and the anastomotic site (measured from incisors), tumour subtype, resection margin length, sex, age or preoperative cardiorespiratory status. CONCLUSIONS: Staple gun size is an important risk factor for BAS formation and 'tilt-top' devices enable the use of a larger head with a subsequently lower incidence of BAS. Endoscopic dilatation is an effective treatment for BAS which rarely recurs and always resolves within 18 months. PMID- 10718960 TI - Biliary complications associated with laparoscopic cholecystectomy: analysis of common misconceptions AB - AIMS: General surgeons often express the view that the majority of biliary complications following laparoscopic cholecystectomy are caused by trainee surgeons; complications occur most often in the presence of difficult anatomy or pathology; biliary injuries occur more proximally in the biliary tree than at open cholecystectomy; most injuries are recognized at the time of surgery; and most complications can be managed non-operatively. The aim of this study was to determine if these views are substantiated in clinical practice. METHODS: Thirty consecutive patients were referred to a specialist hepatobiliary unit over a 7 year period with biliary complications following laparoscopic cholecystectomy. The mode of presentation, management and outcome of these patients were analysed. RESULTS: In 21 cases the initial operator was a consultant. Four of the 30 complications occurred in the presence of an anatomical variation or unusually difficult pathology. Only patients in whom a previous attempt at repair had been made had injuries at or proximal to the bile duct confluence; the only two deaths occurred in this group. Only 41 per cent of injuries were detected at the time of surgery and 89 per cent required further surgical intervention, hepaticojejunostomy being the most common procedure performed (75 per cent). CONCLUSIONS: The majority of bile duct injuries are not caused by trainees, do not occur because of unusual anatomy or pathology, do not occur in the proximal biliary tree, are not recognized at the time of the initial operation and often require major reconstructive procedures for their management. PMID- 10718961 TI - Cholecystectomy: does subspecialization alter workload? AB - AIMS: Cholecystectomy is a common operation. This study reviewed the changes in workload and practice in a teaching hospital over a 4-year period, during which a hepatobiliary subspecialist unit was developed. METHODS: Computerized demographic data, and details of operations and inpatient events were reviewed for all patients undergoing cholecystectomy in a single teaching hospital from 1993 to 1997. For statistical analysis the consultants were grouped into those with a hepatobiliary interest (n = 3) and those with other primary interests (n = 6); and the workload for the first 12 months of the study was compared with that of the last 12-month period. RESULTS: Between April 1993 and April 1997, 1121 cholecystectomies were performed, of which 75 were excluded because they were performed with other simultaneous procedures. Of the remaining operations, 911 involved cholecystectomy alone (mean patient age 52. 9 years), and 135 (12.9 per cent) comprised cholecystectomy with exploration of the common bile duct (ECBD) (mean age 60.1 years). Between the first and last years studied, the rate of ECBD rose significantly from 7.4 to 14.9 per cent (P < 0.01, chi2 test), and the proportion of ECBD procedures being performed by hepatobiliary specialists rose from 31.6 to 52.7 per cent, but this was not statistically significant (P = 0.13). However, for cholecystectomy in the same period the proportion performed by hepatobiliary surgeons rose from 40.4 to 58.5 per cent, representing a highly significant trend (P < 0.001). Following cholecystectomy alone there was a significantly shorter stay associated with patients treated by hepatobiliary surgeons (P = 0.002, F test), although the median postoperative hospital stay was 2 days for both groups of surgeons (interquartile range 1-3 days for hepatobiliary and 1-4 days for non-hepatobiliary surgeons). CONCLUSIONS: Although cholecystectomy is not viewed as a specialist procedure, the trend in this teaching hospital reveals a steady increase in the proportion of cholecystectomies being performed by teams with a biliary interest. The data indicate that this practice is associated with a shorter hospital stay. PMID- 10718962 TI - Investigating the proximal limit of lymphadenectomy in patients with adenocarcinoma of the oesophagus in the mid-thoracic region AB - AIMS: The benefit of extended lymphadenectomy in patients with squamous cell carcinoma of the oesophagus is established, but there is little evidence to support this in patients with adenocarcinoma. The aim of this study was to investigate the extent of lymphatic spread of oesophageal adenocarcinomas, and particularly the proximal spread in tumours located in the mid thorax. METHODS: Twenty-six consecutive patients with tumours arising between 29 and 35 cm from the incisor teeth underwent three-stage oesophagectomy with two-field lymphadenectomy, including nodes in the recurrent laryngeal chains. The proximal extent was measured by endoscopic ultrasonography and confirmed at operation, with division of the lymph node harvest into anatomical sites according to the Japanese classification of oesophageal cancer. RESULTS: There were 21 men and five women, with a mean age of 64 (range 42-78) years; seven patients were lymph node negative in both the mediastinal and abdominal fields. Six patients had nodal metastases more than 2 cm above the tumour and all had extensive involvement of other nodes at the level of the tumour or below, with 7, 7, 9, 12, 15 and 18 nodes positive. There were no patients in whom nodes above the tumour contained metastases while those at the level or below were clear. CONCLUSIONS: Dissection of proximal lymph nodes along the recurrent laryngeal nerve chains in patients with adenocarcinoma of the oesophagus is not warranted. Lymphatic spread above the level of the tumour occurs in association with extensive lymph node involvement elsewhere and removal of proximal nodes from difficult locations is not warranted as a means of improving staging or survival. PMID- 10718963 TI - Need for staging laparoscopy in patients with gastric cancer AB - AIMS: The aim of preoperative staging in gastric cancer is to correctly identify patients with advanced disease who should not be subjected to surgery and to allow the treatment of those who are admitted for operation to be planned accurately. This study assessed the impact of a policy of selective staging laparoscopy in patients deemed suitable for curative or palliative surgery, to ascertain whether the selective approach increased the frequency of abandoned or unplanned surgical procedures. METHODS: Fifty consecutive patients with gastric or type III gastro-oesophageal junction cancer staged by computed tomography (CT) alone, in whom surgery was felt to be appropriate for 'cure' or palliative symptom control, were studied. Specific CT, endoscopic and biochemical criteria were applied prospectively to select out a subgroup of 18 patients who also underwent preoperative staging laparoscopy. The overall accuracy of staging and operative outcomes were assessed. RESULTS: Using this selective approach the resection rate was 98 per cent, although three patients in each group had their planned procedure altered to a less radical (two in each group) or more radical (one in each group) resection (P = 0.23). Overall, 41 of 50 patients were staged correctly (accuracy 82 (95 per cent confidence interval 69-90) per cent) and 86 per cent of patients underwent the planned surgical procedure. The only abandoned operation occurred in the staging laparoscopy group. CONCLUSIONS: It is possible to plan a patient's operation accurately without the need for a staging laparoscopy in all cases. PMID- 10718964 TI - Angiographic embolization for major haemorrhage after upper gastrointestinal surgery AB - AIMS: Severe postoperative haemorrhage after upper gastrointestinal surgery is a serious complication. This study examined the effectiveness of selective mesenteric angiography (SMA) in localizing a bleeding point and the ability of angiographic haemostatic methods to control bleeding. METHODS: The case notes of a consecutive series of nine patients undergoing urgent SMA during 1996-1998 were analysed. Examination of angiography suite records confirmed the accuracy of patient identification. SMA was performed 18 times with 13 embolizations in nine individuals (seven men; median age 54 (27-73) years). Patients underwent the following operations: Whipple pancreaticoduodenectomy (four patients), pancreatic necrosectomy (two), total gastrectomy (one), cholecystectomy (one) and splenectomy (one). The median interval from surgery to haemorrhage was 15 (2-49) days. Six patients presented with haematemesis/melaena and three with bleeding from drains. Seven had evidence of shock (systolic blood pressure less than 100 mmHg, pulse more than 100 per min); the mean preprocedure haemoglobin concentration was 59 g/l. A median of 8 (4-14) units of blood were transfused before embolization and 4 (2-9) units after. Ten initial endoscopies were performed in six patients, seven of which revealed a source of bleeding. Endoscopic haemostasis was attempted in five and achieved temporary control of bleeding in two. RESULTS: Angiography revealed a discrete bleeding point in 13 of 18 procedures in eight patients. Where a bleeding point was identified, angiographic embolization using 3-8-mm stainless steel coils (ten) or a combination of coils and gelatin sponge (three) achieved radiological evidence of haemostasis in all cases. Periprocedural complications occurred in one patient with unintentional partial embolization of the right hepatic artery during embolization of an actively bleeding left hepatic artery pseudoaneurysm. Rebleeding occurred in six patients within 48 h. Three rebleeds were successfully managed with repeat SMA and embolization (one patient required a third embolization); the remaining three required surgery. Definitive radiological haemostasis was achieved in six patients. Five of the nine patients died in hospital, two of whom had been successfully embolized. CONCLUSIONS: In this group of patients, endoscopy contributed relatively little to treatment of postoperative haemorrhage. In contrast, SMA identified a bleeding point in eight of nine patients and achieved definitive control of bleeding in six. SMA and embolization appears to have a useful role in patients with this infrequent but potentially lethal complication. PMID- 10718965 TI - Current nature and management of cancer of the oesophagus and cardia AB - Aims: To inform the debate about upper gastrointestinal cancer care in the UK, the incidence of cancer of the oesophagus and cardia (OGJ) was determined in the West Midlands, a region covering 10 per cent of England and Wales, with particular reference to the methods of treatment. METHODS: The case-notes of 2776 patients diagnosed with oesophageal and OGJ cancer in the 5 years from 1 January 1992 to 31 December 1996 were scrutinized by one experienced surgeon. Tumour types were classified by histology and site, and treatment modalities assessed for 30-day mortality rate together with life-table analyses. RESULTS: Oesophageal cancer was identified in 2188 patients (61 per cent lower, 34 per cent middle, 4 per cent upper), including 999 squamous carcinomas (27 per cent lower, 64 per cent middle, 9 per cent upper) and 995 adenocarcinomas (97 per cent lower, 3 per cent middle), while there were 588 cases of OGJ cancer (94 per cent adenocarcinomas). Resection was the commonest treatment (865 cases; 31 per cent), with a mortality rate of 10 per cent for oesophageal and 4 per cent for OGJ cancer. Palliative resection had a higher mortality rate than radiotherapy (9 versus 3 per cent), compared with 22 per cent for endoscopic palliation, while the 30-day mortality rate was 30 per cent for the 308 patients given no treatment. CONCLUSIONS: Squamous carcinomas and adenocarcinomas of the oesophageal body are now equally common; lower-third and OGJ tumours are predominantly adenocarcinomas. This study provides baseline data for critical appraisal of potential changes in the delivery of upper gastrointestinal cancer in the UK. PMID- 10718966 TI - Specialized intestinal metaplasia: analysis of prevalence, risk factors and association with gastro-oesophageal reflux disease AB - AIMS: There is an increasing awareness that short (less than 3 cm) segments of Barrett's epithelium and macroscopically normal cardia epithelium may harbour specialized intestinal metaplasia (SIM), a premalignant phenotype. This was a prospective study of both the prevalence of SIM in an unselected population of patients attending for endoscopy, and the association of SIM with symptoms, lifestyle, medication, endoscopic oesophagitis and carditis. METHODS: Two hundred consecutive patients underwent endoscopy. Biopsies taken from just below the squamocolumnar junction were stained for SIM, and were analysed for carditis and Helicobacter pylori infection. A detailed questionnaire of symptoms, tobacco consumption and the use of proton pump inhibitors was completed. RESULTS: Forty two patients (21 per cent) had SIM, 19 of 126 (15 per cent) in an endoscopically normal oesophagus, 15 of 63 (24 per cent) in a short segment of Barrett's epithelium and eight of 11 in classical Barrett's oesophagus. Comparative analysis between the SIM positive and negative groups with respect to potential risk factors is outlined below. Table 1. CONCLUSION: SIM is prevalent in patients undergoing endoscopy, does not correlate with symptoms or with H. pylori infection, but is significantly associated with endoscopic and pathological markers of gastro-oesophageal reflux. PMID- 10718967 TI - Early results of laparoscopic swedish adjustable gastric banding for morbid obesity AB - AIMS: Gastric bypass and vertical banded gastroplasty (VBG) are currently the most commonly performed bariatric procedures but neither are ideal. The results of the first 56 patients who had laparoscopic Swedish adjustable gastric banding (SAGB) are presented. METHODS: All patients referred for bariatric surgery were considered for SAGB. Each was given the alternative of gastric bypass. Patients with a body mass index of less than 35, large hiatus hernia, under 18 years of age, gastric pathology and significant psychiatric illness were excluded. Preoperative gastroscopy, ultrasonographic examination of the gallbladder and specialist anaesthetic assessment were arranged. All but five patients had attempted laparoscopic procedures. Patients were discharged when they were mobile and could tolerate 1500 ml fluid per day. Patients were given a liquidized diet for 6 weeks. Assessments were made at 6 weeks and 3, 6, 9 and 12 months. RESULTS: Some 56 consecutive patients were followed for up to 12 months. The conversion rate fell from 52 per cent for the first 25 patients to 20 per cent for the last 20. Conversion rates were higher in men and in superobese patients. The duration of a laparoscopic operation fell significantly with experience but was still significantly longer than that of an open procedure (P < 0.004). The length of hospital stay was significantly shorter for laparoscopic procedures (P < 0.001). There was no death and little morbidity. Two bands had to be removed (easily) by open surgery, one for infection and one for a recurrence of gastric herniation. The mean excess weight loss was 60 per cent at 12 months. Greater than 50 per cent of excess body-weight was lost by 48 per cent of patients at 6 months and 69 per cent at 12 months. Band adjustments in most patients achieved further weight loss to compensate for late pouch dilatation. Most failures were in patients who failed to attend. CONCLUSION: SAGB appears to have many advantages over gastric bypass and VBG; it avoids stapling the stomach, should not cause any malabsorption, can be performed laparoscopically, is adjustable, is more readily reversed (if necessary) and, therefore, has the potential for lower associated morbidity and mortality rates. In terms of excess weight loss, the early results are certainly as good, if not better, than those of gastric bypass and VBG. PMID- 10718968 TI - Pattern of recurrence following subtotal oesophagectomy with two field lymphadenectomy AB - Aims: Despite increasingly radical surgery for oesophageal cancer many patients continue to represent with recurrent disease. This study aimed to evaluate the pattern of failure following attempted curative oesophagectomy with mediastinal and upper abdominal lymphadenectomy. METHODS: Some 212 consecutive patients undergoing R0 resection for malignancy between 1 April 1990 and 1 April 1999 were followed up for evidence of recurrence. Clinical evaluation was supported by ultrasonography, computed tomography, isotope scan, endoscopy and laparotomy with biopsy assessment if appropriate. Patients were excluded if recurrence was diagnosed on clinical grounds alone. Statistical analysis was performed using chi2 and log rank tests. RESULTS: Some 142 patients with adenocarcinoma and 70 with squamous carcinoma (SCC) were followed up for a median of 14 (range 1-108) months. Sex and age distribution were similar for both histological subtypes (men : women 3 : 1; median age 64 (30-79) years). Twenty patients died from non-cancer related causes, including 11 (5 per cent) from postoperative complications. Some 89 patients (42 per cent) developed proven recurrent disease of which seven are alive and 82 dead. The median time to recurrence was 11 (2-40) months with a median time to death thereafter of 3 (1-21) months. The pattern of recurrence was locoregional in 23 per cent (oesophageal bed 15 per cent, upper abdominal 3 per cent, upper mediastinal 3 per cent, cervical 2 per cent) and haematogenous in 18 per cent (comprising liver 8 per cent, bone 4 per cent, cerebral 3 per cent, lung 2 per cent, skin 1 per cent) with peritoneal dissemination in 1 per cent. While there was no difference in the overall pattern of dissemination for each histological subtype, the incidence of cervical and upper mediastinal recurrence was significantly higher for adenocarcinoma compared with SCC (chi2 = 5. 9, 1 d.f., P < 0.02). The timing of recurrence was similar for both histological subtypes: 60 per cent of all recurrence occurred within 12 months of surgery, with distant and locoregional recurrence occurring at a median of 10 (2-40) and 11 (2-32) months respectively. CONCLUSIONS: The low incidence of upper mediastinal and cervical recurrence suggests that more extensive lymphadenectomy is unlikely to impact upon survival. Improved staging modalities are required to identify the significant number of patients who develop early recurrence in the first year following surgery in order to offer them multimodality therapies of non-surgical palliation. PMID- 10718969 TI - Management of spontaneous rupture of the oesophagus AB - AIMS: Spontaneous rupture of the oesophagus (SRO) is a rare and often fatal event. The aim of this study was to evaluate the presentation, management and outcome of SRO in a single unit. METHODS: Data were collected on all patients presenting with SRO over a 5-year period with respect to presenting features, diagnostic investigations and subsequent management. Statistical analysis was by Student's t test, chi2 and Fisher's exact tests. RESULTS: Fourteen patients were identified, 12 men and two women with a median age of 64 (range 18-78) years; eight were tertiary referrals. Thirteen of 14 patients presented with chest or upper abdominal pain following vomiting or retching and 13 had an abnormal initial chest radiograph; only one presented with Mackler's triad of pain, vomiting and surgical emphysema. The median delay to diagnosis was 21 (range 1 84) h; this delay did not significantly affect outcome (P = 0.16). An endoscopic assessment and contrast swallow were performed in all patients. Nine of ten patients with a demonstrable leak and full-thickness tear were managed surgically and the four patients with no leak were managed conservatively (P = 0.005); surgical management consisted of thoracotomy, lavage, repair of the perforation and a feeding jejunostomy. Seven patients had a repair over a T tube and two had a primary repair. All conservatively managed patients had contained, controlled or intramural perforations and two also required a feeding jejunostomy. Patients requiring surgery had a longer hospital stay (mean(s.d.) 57.9(34.8) versus 22.2(30.7) days; P = 0.081) and a significantly longer intensive care unit stay (P = 0.044). The overall mortality rate from SRO was 14 per cent (two patients); no deaths occurred in the conservatively managed group. CONCLUSIONS: SRO continues to be diagnosed late despite a classical history and/or abnormal chest radiograph. Endoscopic assessment of perforations is safe and in combination with a contrast swallow can confidently predict patients with contained or controlled rupture in whom non-operative management is successful. PMID- 10718970 TI - Achalasia: management, outcome and surveillance in a specialist unit AB - AIMS: The management and surveillance of achalasia remains controversial at the present time. The aim of this study was therefore to evaluate the results of endoscopic management and subsequent surveillance of patients with achalasia presenting to a specialist unit. METHODS: A prospective cohort of 40 patients with a radiological and manometric diagnosis of achalasia who presented to this unit between 1991 and 1998 were studied; the male : female ratio was 1 : 1 and the median age 38 (range 15-84) years. Twenty-one patients presented de novo, seven had previously undergone cardiomyotomy and 12 were referred following unsuccessful dilatation. RESULTS: Some 36 patients were treated with balloon dilatation (Microvasive achalasia balloon, 35/40 mm). Results were graded 1-4 (1, asymptomatic; 2, symptomatic but significantly improved; 3, symptomatic with no change; and 4, symptomatic but worse); 29 of 36 patients were grade 1 at subsequent follow-up and the remaining seven were grade 2 (median follow-up 17 (range 5-96) months). There was a single complication of oesophageal perforation which was treated conservatively with full recovery. Following intervention, patients were enrolled in a prospective surveillance programme of chromoendoscopy at 2-year intervals; in a total of 74 patient-years' follow-up, two superficial squamous cell carcinomas (SCCs) and one adenocarcinoma (following cardiomyotomy) were detected, giving a relative risk of one cancer in 25 patient-years. CONCLUSIONS: Balloon dilatation is a safe and effective treatment for achalasia even in patients who have had previous unsuccessful dilatations or cardiomyotomy. There is a high risk of SCC and the adenocarcinoma may have resulted from previous refluxogenic therapy, so all patients with achalasia should be followed up with surveillance endoscopy. PMID- 10718971 TI - Prognostic value of quality of life scores in patients with oesophageal cancer AB - AIMS: Quality of life (QL) data are useful in evaluating treatment, in screening or psychosocial morbidity and there is accumulating evidence to show that they predict survival. This study investigated if baseline QL scores are prognostic for patients with oesophageal cancer. METHODS: Between 1993 and 1995, 89 consecutive new patients with oesophageal cancer completed baseline QL assessments with the EORTC QLQ-C30 questionnaire and the dysphagia scale from the oesophageal cancer module. Cox's proportional hazards models were used to assess the impact of QL variables on survival (82 patients have died). RESULTS: Univariate analyses revealed that better baseline physical and role function scores were significantly associated with increased survival (P 90% of the total genetic component in both species considered individually or combined. This indicated restricted gene flow, consistent with the limited dispersal abilities of Begonia generally and the ancient separation of isolated forest patches. Genetic distances among populations are much higher than usually found within species. Allozyme data provide no support for the recognition of B. dregei and B. homonyma as distinct species. PMID- 10719005 TI - Artocarpus (Moraceae)-gall midge pollination mutualism mediated by a male-flower parasitic fungus. AB - A previously undescribed pollination system involving a monoecious tree species, Artocarpus integer (Moraceae), pollinator gall midges, and fungi is reported from a mixed dipterocarp forest in Sarawak, Borneo. The fungus Choanephora sp. (Choanephoraceae, Mucorales, Zygomycetes) infects male inflorescences of A. integer, and gall midges (Contarinia spp., Cecidomyiinae, Diptera) feed on the fungal mycelia and oviposit on the inflorescence. Their larvae also feed on the mycelia and pupate in the inflorescence. The gall midges are also attracted by female inflorescences lacking mycelia, probably due to a floral fragrance similar to that of male inflorescences. Because of the sticky pollen, dominance of Contarinia spp. in flower visitors, and pollen load observed on Contarinia spp. collected on both male and female inflorescences, Artocarpus integer is thought to be pollinated by the gall midges. Although several pathogenic fungi have been reported to have interactions with pollinators, this is the first report on a pollination mutualism in which a fungus plays an indispensable role. The pollination system described here suggests that we should be more aware of the roles fungi can play in pollinations. PMID- 10719006 TI - Respiratory pathogens: assessing resistance patterns in Europe and the potential role of grepafloxacin as treatment of patients with infections caused by these organisms. AB - Although most respiratory tract infections (RTI) are caused by viruses, various bacteria, particularly Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis, are common causes of community-acquired pneumonia, acute exacerbations of chronic bronchitis, otitis media and sinusitis. Empirical antibiotic therapy of patients with RTI must take account of the increasing prevalence of resistance among the predominant pathogens. Europe-wide susceptibility surveillance studies have revealed that resistance to penicillin and macrolides is highly prevalent among isolates of S. pneumoniae from France and Spain. Uniquely, in Italy, macrolide resistance is highly prevalent while the prevalence of penicillin resistance is low. Resistance to other antibiotic classes, including chloramphenicol, doxycycline and, in particular, co trimoxazole, is associated with penicillin resistance in pneumococci, but resistance to the fluoroquinolones is rare. beta-Lactamase production is the principal mechanism of resistance in isolates of H. influenzae and M. catarrhalis, with fluoroquinolone resistance being detected rarely in these pathogens. In 1998 a surveillance study involving 15 European countries determined the susceptibilities of many respiratory pathogens to a range of antimicrobials, including grepafloxacin. The MIC(90) of grepafloxacin for 1251 isolates of S. pneumoniae was 0.25 mg/L, the MICs for only five strains being >2 mg/L, and 99.4% of all of the isolates tested were inhibited by concentrations MIC) were calculated and serum inhibitory titres (SITs) were determined. Haemophilus spp. were the predominant potential bacterial pathogens and were recovered from the sputa of 24 patients. Strains of Streptococcus pneumoniae were isolated from two patients in the grepafloxacin group and a strain of Moraxella catarrhalis was isolated from one patient in the clarithromycin group. Haemophilus spp. isolates were eradicated from the sputa of 13 of 14 (93%) patients given grepafloxacin, but from only two of 10 (20%) patients given clarithromycin (P < 0.05). In the other eight (80%) patients who received clarithromycin, the sputum cultures remained positive throughout the 10 day course. Grepafloxacin eliminated potential bacterial pathogens more quickly than clarithromycin (median T(erad) 4 h versus 76 h). The S. pneumoniae strains were eradicated by grepafloxacin within 4 h and the single M. catarrhalis strain was eradicated by clarithromycin within 1 h. The greater efficacy of grepafloxacin, compared with that of clarithromycin, in terms of the incidence and speed of eradication of the Haemophilus spp. isolates, was associated with higher median values of AUIC(24) (169 SIT(-1)*h versus 8.1 SIT( 1)*h), C(max):MIC ratio (23.6 versus 0.7) and %tau >MIC (100% versus 0%). A Hill type model adequately described the relationship between the percentage probability of eradicating potential bacterial pathogens from sputa and the plasma grepafloxacin concentration. PMID- 10719008 TI - Optimizing economic outcomes in antibiotic therapy of patients with acute bacterial exacerbations of chronic bronchitis. AB - The social, medical and economic effects of acute bacterial exacerbations of chronic bronchitis on individual patients and the resource implications of this disease for the healthcare sector are considerable. Optimizing the selection of patients who should receive antibiotics according to stringent clinical criteria is the first step in promoting good clinical practice and cost-effectiveness. Antibiotic efficacy is then the major driver of cost, especially when it reduces the need for hospitalization. Resistance to first-line antibiotics can be expected to increase the risk of treatment failure. Other drivers of cost include non-compliance, which predisposes to therapeutic failure, and the selection of resistant strains. Treatment regimens of short duration, once-daily dosing and good tolerability are determinants of good compliance and cost savings. The expenses of first-line antibiotics typically account for only a small proportion of the overall costs of healthcare and the cheapest antibiotics are not necessarily the most cost-effective. The clinical success rate of first-line therapy is the primary determinant of the overall expenditure on healthcare because of the high costs associated with treatment failure, especially if it leads to hospitalization. Factors such as poor patient compliance and high antibiotic resistance rates, which undermine the clinical efficacies of first line therapy, will increase the overall costs of treatment. PMID- 10719010 TI - Israel (Sol) penn (1930-1999) PMID- 10719009 TI - The patient's burden: physical and psychological effects of acute exacerbations of chronic bronchitis. AB - In this prospective qualitative study we evaluated the subjective perspectives of the quality of life of patients with chronic bronchitis. Individuals with diagnoses which fulfilled the clinical criteria of chronic bronchitis, i.e. daily production of sputum for at least three consecutive months in two consecutive years, were recruited into four focus groups from general practices in two industrial cities. Younger patients (those of pre-retirement age) of both sexes-a significant, but frequently 'invisible' minority in this patient population-were targeted. The groups were constituted with the aim of stimulating variation in the discussions. Twenty sufferers (10 males and 10 females, ranging in age from 30 to 86 years) were eventually included in the study; there were five in each group. Group discussions were recorded and transcribed verbatim and the data were analysed thematically. It was evident from the discussions that chronic bronchitis had led to a high degree of psychological distress in the participants, particularly in relation to dependency on medication, and disruption of social and family relationships. Acute exacerbations of chronic bronchitis (AECB) were met with dread. They brought about further reductions in quality of life, increased anxieties about breathlessness, fear of atmospheric pollution and of changes in and extremes of temperature, embarrassment about coughing up phlegm in public and suspicion of medical practitioners' motives if they were unwilling to prescribe antibiotics on request. Patients' health-related behaviour and beliefs were often contradictory. For example, AECB in some patients led to increased smoking. There were also gender and age differences; for example, it was the perception of males that they received more support from their partners than did females. Younger participants appeared more distressed by AECB than older ones. The results of this study suggest that raising the standard of care for patients with chronic bronchitis requires that greater attention be paid to patients' subjective experiences of the disease. PMID- 10719011 TI - Fetal microchimerisms in the mother: immunologic implications. AB - The previously held concept that the fetus is completely separated from the mother, especially by trophoblasts that line the outer layer of the placenta, has recently been questioned. It has recently been shown that fetal cells are detectable not only in the peripheral blood, but also in maternal skin and liver. Although the migration of fetal cells into the maternal circulation has been given a great deal of attention because of its implication in the prenatal diagnosis of genetic diseases, the potential role of such placental transfer of fetal cells in the pathogenesis of autoimmune diseases has only recently been considered. In patients with scleroderma, fetal cell-derived DNA was detected more frequently in the peripheral blood of patients than controls. This finding of a limited number of fetal cells in maternal tissues leading to microchimerism has been proposed to have a role in the induction of scleroderma. Although evidence for microchimerism has also been confirmed in a high frequency of liver tissues from patients with primary biliary cirrhosis (PBC), a similar high frequency was noted in control patients, which suggests that microchimerism by itself is unlikely to fully account for the pathogenesis of PBC. The finding of such a high frequency of fetal microchimerism in the liver suggests that this is a very common event, raising the possibility that the migration of fetal cells may be important in the induction and subsequent maintenance of tolerance against the fetus during pregnancy. In addition, it is clearly possible that such microchimerism could contribute to the pathogenesis of select autoimmune diseases. PMID- 10719012 TI - Management of cytomegalovirus infection after liver transplantation. PMID- 10719013 TI - Ascites after liver transplantation. AB - Massive ascites after liver transplantation, although uncommon, usually represents a serious adverse event. The pathogenesis of this complication has not been adequately investigated. To determine the incidence, characteristics, and pathogenic factors of massive ascites after liver transplantation (ascitic fluid > 500 mL/d for >10 days), the charts of 378 liver transplant recipients were reviewed. Massive ascites occurred in 25 patients (7%). Mean ascitic fluid production was 960 mL/d (range, 625 to 2,350 mL/d), and the duration of ascites was 77 days (range, 15 to 223 days). The ascitic fluid had a high protein content (36 +/- 7 g/L; range, 25 to 50 g/L). When patients who did and did not develop massive ascites were compared, significant differences were found in receptor sex (men, 88% v 60%, respectively; P <.01) and surgical technique (inferior vena cava preservation with piggyback technique, 72% v 41%; P <.01). Significantly increased wedged and free hepatic venous pressures and gradients between hepatic vein and right atrial pressures were found in patients who developed ascites, suggesting a difficulty in graft blood outflow. Massive ascites was associated with renal impairment, increased incidence of abdominal infection, prolonged hospitalization, and a tendency toward reduced survival. In conclusion, massive ascites after liver transplantation is relatively uncommon but associated with increased morbidity and mortality and is predominantly related to difficulties of hepatic venous drainage. Measurement of hepatic vein and atrial pressures to detect a significant gradient and correct possible alterations in hepatic vein outflow should be the first approach in the management of these patients. PMID- 10719014 TI - Acute liver failure: clinical features, outcome analysis, and applicability of prognostic criteria. AB - Acute liver failure (ALF) is an uncommon condition associated with high morbidity and mortality. We performed a retrospective analysis of patients evaluated for ALF. The aim of our study is to determine the clinical features and outcome of such patients and to assess the validity of King's College Hospital (KCH) prognostic criteria. One hundred seventy-seven patients were evaluated for ALF during a period of 13 years. Mean age was 39 years, and 63% were women. The causes included viral hepatitis (31%), acetaminophen toxicity (19%), idiosyncratic drug reactions (12%), miscellaneous causes (11%), and an indeterminate group (28%). Twenty-five patients (14%) recovered with medical therapy (group I), 65 patients (37%) died without orthotopic liver transplantation (OLT; group II), and 87 patients (49%) underwent OLT (group III). Patients in group II were older and often had advanced encephalopathy, whereas those in group I had less hyperbilirubinemia and often had hyperacute failure. KCH criteria had high specificity and positive predictive value but low negative predictive value for a poor outcome. We conclude that early prognostication is needed in patients with ALF to assist decision making regarding OLT. The fulfillment of KCH criteria usually predicts a poor outcome, but a lack of fulfillment does not predict survival. PMID- 10719015 TI - Use and validation of selection criteria for liver transplantation in acute liver failure. PMID- 10719016 TI - Retransplantation for late liver graft failure: predictors of mortality. AB - As patient survival after orthotopic liver transplantation (OLT) improves, late complications, including late graft failure, more commonly occur and retransplantation (re-OLT) is required more often. Survival after re-OLT is poorer than after primary OLT, and given the organ shortage, it is essential that we optimize our use of scarce donor livers. We sought to identify variables that predict poor outcome after late re-OLT. Among adults who underwent OLT between September 1989 and October 1997, we identified transplant recipients who survived greater than 6 months (n = 964) and analyzed those who required late re-OLT (>/=6 months after primary OLT). We recorded the indication for the initial OLT and interval from OLT to re-OLT. We also analyzed data collected at the time of re OLT, including age, sex, indications for primary OLT and re-OLT, United Network for Organ Sharing status, preoperative laboratory values (white blood cells, platelets, hemoglobin, albumin, bilirubin, creatinine, and prothrombin time), Child-Pugh-Turcotte score, number of rejection episodes before re-OLT, and interval between OLT and re-OLT. In addition, we analyzed surgical factors (including procedure performed and use of packed red blood cells, fresh frozen plasma, and platelets), postoperative immunosuppression, and donor factors (age, ischemic time). Forty-eight patients (5%) underwent late re-OLT at a median of 557 days (range, 195 to 2,559 days) post-OLT. Survival rates after re-OLT at 90 days, 1 year, and 5 years were 71%, 60%, and 42%, respectively. Patients surviving 90 days or greater after re-OLT had an 85% chance of surviving to 1 year. Sepsis was the leading cause of death (15 of 25 deaths; 60%). Recipient age older than 50 years (P =.04), preoperative creatinine level greater than 2 mg/dL (P =.004), and use of intraoperative blood products (packed red blood cells, P =.001; fresh frozen plasma, P =.002; platelets, P =.004) had significant impacts on survival. Late re-OLT was associated with increased mortality. Careful patient selection, with particular attention to recipient age and renal function, may help improve results and optimize our use of scarce donor livers. PMID- 10719017 TI - Rapid method for the analysis of peripheral chimerism in suspected graft-versus host disease after liver transplantation. AB - The effects of microchimerism and possible tolerance have been well studied in orthotopic liver transplantation. In some patients, greater levels of donor cells persist in the periphery. These cells were characterized and their effects on clinical outcome were studied. Peripheral blood was obtained from patients at various times posttransplantation. HLA class II typing was performed by the polymerase chain reaction-sequence-specific primer method on unfractionated blood and lymphocyte subpopulations. Relative levels of amplification of donor and recipient alleles were compared. All patients studied had a low degree of chimerism that was most apparent in the CD8(+)T/natural killer (NK) cell population. One patient with persistently high levels of donor alleles in his CD8(+)T/NK cell population was diagnosed with severe graft-versus-host disease (GVHD) and died of opportunistic infections. Another patient with biopsy-proven GVHD was chimeric in several cell populations. On resolution of her symptoms, donor alleles were reduced to levels undetectable by this assay. These results suggest that persistently elevated levels of donor CD8(+)T/NK cells in the periphery may indicate GVHD in liver transplant recipients. This technique aids in rapid diagnosis, which facilitates appropriate treatment and thus may improve clinical outcome. PMID- 10719018 TI - Prevalence of hepatitis C virus-associated mixed cryoglobulinemia after liver transplantation. AB - Hepatitis C virus (HCV) infection is associated with mixed cryoglobulinemia and membranoproliferative glomerulonephritis. After orthotopic liver transplantation (OLT), isolated cases of HCV-associated mixed cryoglobulinemia have been reported. We determined the prevalence and clinical characteristics of mixed cryoglobulinemia in HCV-infected liver transplant recipients at our institution. Between January 1991 and February 1998, a total of 191 OLTs were performed in 178 patients. Among these transplant recipients, 53 patients (29.8%) had positive serological test results for HCV infection by second-generation enzyme-linked immunosorbent assay. We studied 31 HCV-positive (HCV+) and 21 HCV-negative (HCV-) transplant recipients (control group). Renal and liver function studies were performed, and cryoglobulin, rheumatoid factor, C3, C4, and serum HCV RNA levels and genotype were determined. Results were compared using unpaired Student's t test for continuous variables and Fisher's exact test for categorical variables. Baseline characteristics were similar between the groups. Six patients in the HCV+ group (19%) had mixed cryoglobulins present at the time of evaluation compared with none in the HCV- group (P =. 036). The only parameter associated with cryoglobulins in the HCV+ group was rheumatoid factor (P <.01). In 3 HCV+ patients with cryoglobulins, extrarenal signs of cryoglobulinemia were present. Glomerulonephritis was found in 4 HCV+ patients. Two patients with purpura and cryoglobulinemia had reduced clinical manifestations after antiviral therapy. In conclusion, mixed cryoglobulinemia was found in approximately 20% of the HCV+ liver transplant recipients. The presence of purpura or glomerulonephritis suggests HCV-associated mixed cryoglobulinemia, a clinical syndrome that may respond favorably to antiviral therapy. PMID- 10719019 TI - Hepatitis A antibodies in liver transplant recipients: evidence for loss of immunity posttransplantation. AB - Liver transplant recipients frequently have chronic liver diseases and should be considered for vaccination against hepatitis A virus (HAV). However, persistence of protective antibodies after orthotopic liver transplantation (OLT) has not been shown in this population, which may have implications for future vaccine recommendations. We evaluated the prevalence and epidemiological significance of immunoglobulin G (IgG) antibody to HAV (anti-HAV) in a nonvaccinated population before OLT (immunity from previous exposure) and determined the persistence of IgG anti-HAV at 1 and 2 years after OLT. One hundred consecutive patients were identified who underwent OLT and had at least 2 years of follow-up post-OLT. They were not vaccinated against HAV infection at any time. Clinical data were summarized from medical records, and stored sera were tested for IgG anti-HAV before OLT and at 1 and 2 years after OLT by a commercially available enzyme immunoassay. Of 100 patients, 24 had IgG anti-HAV before OLT. No epidemiological differences were noted between those with or without detectable IgG anti-HAV before OLT. Among patients with detectable IgG anti-HAV before OLT, 4 of 22 patients (18%) and 7 of 24 patients (29%) became negative for IgG anti-HAV at 1 and 2 years post-OLT, respectively. None of the patients with undetectable IgG anti-HAV before OLT became positive at any time. Most of our patients with end stage liver disease had no serological evidence for immunity against HAV. A significant proportion of patients with detectable protective antibodies before OLT lost their antibodies at 2 years after OLT. PMID- 10719020 TI - Adenosine A2 receptor stimulation protects the predamaged liver from cold preservation through activation of cyclic adenosine monophosphate-protein kinase A pathway. AB - The shortage of organ donors has led to reconsideration for the use of non-heart beating donors (NHBDs). However, graft injury caused by warm ischemia in livers from NHBDs strongly affects posttransplantation outcome. The aim of the present study is to investigate the role of adenosine A2 receptor with regard to hepatic viability after cold preservation of NHBD livers. Cardiac arrest was induced in Wistar rats by phrenotomy of the anesthetized nonheparinized animal. After 60 minutes, the livers were excised and flushed with 60 mL of histidine-tryptophan ketoglutarate (HTK) and stored submerged in HTK at 4 degrees C for 24 hours. Reperfusion was performed in vitro after all livers were incubated at 22 degrees C in saline solution to account for the period of slow rewarming during surgical implantation in vivo. Addition of the selective A2-receptor agonist (CGS 21680; 30microg/100 mL) to the preservation solution resulted in a significant reduction to one quarter of the parenchymal enzyme release of alanine aminotransferase or lactate dehydrogenase on reperfusion and promoted a 2-fold increase in hepatic bile production. This salutory effect was accompanied by a significant increase (40%) in the activity ratio of protein kinase A (PKA) in the liver tissue and could be abrogated in large part by the PKA inhibitor, Rp-cAMPs. Stimulation of the adenosine A2 receptor during harvest and storage of the graft improves maintenance of tissue integrity in liver grafts. A major part of this effect, which may represent a promising approach for the use of NHBD grafts, seems to be mediated through activation of PKA. PMID- 10719021 TI - Results of choledochojejunostomy in the treatment of biliary complications after liver transplantation in the era of nonsurgical therapies. AB - Advances in radiological and endoscopic techniques have allowed many biliary complications after orthotopic liver transplantation (OLT) to be managed without surgery. The influence of nonsurgical management on the outcome of patients requiring surgical revision has not been addressed. We reviewed our 10-year experience (October 1988 to January 1998) of Roux-en-Y choledochojejunostomy (CDJ) to treat biliary complications after OLT. Forty-six patients underwent CDJ for biliary complications (32 men, 14 women; age, 22 to 65 years; median, 60 years). Biliary reconstruction at the time of OLT was duct to duct in 41 patients, primary CDJ in 3 patients, and gall bladder conduit in 2 patients. T tubes were used only in patients with gallbladder conduit. The indication for CDJ was biliary leak (23 patients), stricture (20 patients), biliary stones (2 patients), and biliary sludge (1 patient). Two patients (4.3%) had associated hepatic artery thrombosis. The bile leaks were diagnosed at a median of 29 days post-OLT (range, 2 to 65 days) and strictures at a median of 2 years (range, 33 days to 6.5 years) post-OLT. Before surgery, 25 patients (54%) underwent an attempt at radiological or endoscopic therapeutic intervention that failed. Median follow-up was 5 years (range, 9 months to 10 years). Early complications occurred in 12 patients (26%); the most common was chest infection (4 patients). There were 3 perioperative deaths (6%); 1 death was directly related to surgery. Late complications, mainly anastomotic strictures, occurred in 10 patients (22%), half of which were successfully treated by biliary balloon dilatation. The complication rate post-CDJ was less in those who underwent a failed nonsurgical approach than those proceeding straight to surgery (9 of 25 patients; 36% v 13 of 21 patients; 62%; P =.21, not significant). The procedure-related mortality for surgical revision of biliary complications after OLT is low, but early and late complications are common. A failed attempt at nonsurgical management does not increase the complications of reconstructive surgery. Strictures after CDJ should be considered for biliary balloon dilatation. PMID- 10719022 TI - Hydrophilic bile salts protect bile duct epithelium during cold preservation: a scanning electron microscopy study. AB - Prolonged graft preservation is associated with postoperative bile duct strictures after liver transplantation. We previously showed that hydrophilic bile salts mitigate bile duct preservation injury in a pig model. Because this injury occurs at the epithelial level, scanning electron microscopy was performed to further characterize this effect in vitro. Swine livers were harvested after the intravenous infusion of 1 of 3 solutions: saline (n = 7), tauroursodeoxycholate ([TUDC] hydrophilic; n = 4), or taurodeoxycholate ([TDC] hydrophobic; n = 4). Livers were perfused with University of Wisconsin solution. The bile ducts were flushed retrograde, and the liver was stored at 0 degrees C to 1 degrees C for 20 hours. Bile duct samples were obtained at the time of harvest and 8, 12, 16, and 20 hours thereafter. In saline-infused controls at time 0, the epithelium was intact and composed of uniform cuboidal cells covered with fine regular microvilli. There were no spaces between individual cells. After 8 to 12 hours of preservation, cells were more irregular in shape, with loss of cell-cell contact. The cell surfaces showed fewer microvilli. Surface erosions suggested loss of cell-wall integrity. TUDC was protective, evidenced by normal-appearing cells with uniform microvilli after 16 hours. In contrast, TDC accelerated the injury process, causing cell-surface erosions, blebs, and loss of microvilli as early as time 0. Scanning electron microscopy is an excellent tool to study injury to bile duct epithelium. This study supports the hypothesis that hydrophilic bile salts protect bile ducts during preservation. To determine whether treatment with hydrophilic bile salts can prevent postoperative stricture, in vivo transplantation studies are needed. PMID- 10719023 TI - Maternal hemodynamics and pregnancy outcome in women with prior orthotopic liver transplantation. AB - The aim of this study is to evaluate the hemodynamics and pregnancy outcome of women with prior orthotopic liver transplantation. Hemodynamic measurements by Doppler technique were performed on pregnant subjects with prior orthotopic liver transplantation. Maternal characteristics, renal function, pregnancy complications, delivery indications, delivery mode, and neonatal outcomes were evaluated. Six pregnancies occurred in 5 women after orthotopic liver transplantation at the University of Washington Medical Center (Seattle, WA) between 1991 and 1999. Four of the 6 pregnancies were complicated by chronic hypertension, fetal growth restriction, and preterm delivery. Two pregnancies had worsening hypertension characterized by vasoconstriction in the second trimester despite antihypertensive therapy. These 2 subjects were administered cyclosporine for maintenance immunosuppression and had greater mean arterial pressures preconception and in the first trimester than the other subjects. One of these pregnancies resulted in fetal demise at 25 weeks' gestation. The other subject was delivered at 28 weeks' gestation for nonreassuring fetal status and superimposed preeclampsia. All pregnancies were complicated by renal insufficiency; however, the 2 subjects with poor obstetric outcome had preconception serum creatinine levels greater than 1.5 mg/dL and creatinine clearances less than 40 mL/min. Pregnancies complicated by second-trimester vasoconstriction and moderate renal insufficiency are at risk for preeclamspia, fetal growth restriction, and fetal demise. Good obstetric outcome can occur in women with mild renal insufficiency and well-controlled chronic hypertension. Improved hypertensive control preconception may decrease the risk for preeclampsia and poor obstetric outcome. PMID- 10719024 TI - Cytokine profile of liver- and blood-derived nonspecific T cells after liver transplantation: T helper cells type 1/0 lymphokines dominate in recurrent hepatitis C virus infection and rejection. AB - Orthotopic liver transplantation (OLT) is a successful treatment in patients with hepatitis C virus (HCV)-associated end-stage liver disease worldwide. T lymphocytes and their cytokines are believed to have a pivotal role in the defense against HCV and in allograft rejection. An immunosuppressive drug regimen may cause a cytokine imbalance toward a T helper (TH) cell type 2 profile that is associated with the persistence of infection and acceptance of the graft. The aim of this study is to assess whether cytokine imbalances toward a TH1- or TH2-type response may have a role in recurrent HCV infection and rejection after OLT. Twenty-one intrahepatic T-cell lines of 15 patients with recurrent HCV infection after OLT (group A) and 15 lines of 11 patients with rejection (group B) were studied. Both patient groups had received liver allografts because of HCV associated end-stage liver disease. Patients with HCV-induced liver disease who did not undergo OLT served as controls: 17 patients with chronic hepatitis C (CH C) and 8 patients with cirrhosis. At the time of liver biopsy, 14 blood-derived T cell lines of 12 patients with recurrent HCV infection, 7 of 10 patients with rejection, and 18 of 18 control patients were also investigated. Regardless of the underlying disease (recurrent HCV infection, rejection, HCV-induced hepatitis, and cirrhosis), all liver tissue-derived T-cell lines produced interferon-gamma; some additionally produced interleukin-4 (IL-4), but none produced IL-10, indicating that the TH0/1 cytokine profile dominates. T-cell lines showing a TH1 cytokine profile derived from intrahepatic T cells could be established significantly more often in recurrent HCV infection and rejection than in controls with CH-C (Fisher's exact test, P <.05). The cytokine profile of intrahepatic T cells did not differ from that obtained in peripheral blood. TH0/1 cytokine profile dominates the intrahepatic and blood-derived immune response in recurrent HCV infection and rejection after OLT regardless of the mode of immunosuppression. The lymphokine profile of immunocompromised patients with recurrent HCV infection or rejection does not differ principally from that of patients with HCV-induced hepatitis and cirrhosis, but seems to show a TH1 profile significantly more often. PMID- 10719025 TI - Increased late hepatic artery thrombosis rate and decreased graft survival after liver transplants with zero cross-reactive group mismatches. AB - The use of broad-specificity cross-reactive groups (CREGs) at the A and B HLA loci has been proposed as a means to improve both access and outcome for renal transplantation but has not yet been studied for liver transplantation. We retrospectively analyzed our results for all adult liver transplantations performed at our center between 1989 and 1996 for which HLA typing and complete 3 year follow-up data were available. Two hundred eight transplantations were studied, with a mean follow-up of 66 +/- 2 months (range, 36 to 110 months); A and B loci were converted to CREGs by the method of Rodey. Thirteen percent of the patients with 0 CREG mismatches had hepatic artery thrombosis versus 2% for those with 1 or more mismatches (odds ratio, 6.7; 95% confidence interval, 2.6 to 17.3 by univariate analysis; odds ratio, 3.5; 95% confidence interval, 1.1 to 11.7 by logistic regression analysis). These events occurred significantly later in the 0-CREG mismatch group (21 +/- 7 v 4 +/- 2 months posttransplantation; P =.04 by Student's t-test). Graft survival rates were significantly lower for patients with 0 CREG mismatches (56% v 68% at end of study; P =.05 by Cox-Mantel test). The number of CREG mismatches had no effect on the frequency of rejection, steroid-resistant rejection, or infectious complications, including cytomegalovirus. Neither total nor individual A, B, or DR HLA matching had an effect on outcome. There appears to be little evidence that intentional CREG matching would improve outcomes for liver transplantation and, under some circumstances, could be deleterious. PMID- 10719026 TI - Liver transplantation complicated by embedded transjugular intrahepatic portosystemic shunt: a new method for portal anastomosis- a surgical salvage procedure. AB - A transjugular intrahepatic portosystemic shunt (TIPS) is an increasingly used method of treating variceal bleeding from portal hypertension. Many patients are subsequently listed for transplantation, which may be complicated by malposition of the inferior end of the TIPS stent. This report details such a case and offers a surgical technique to salvage this situation. PMID- 10719027 TI - Transplantation of three adult patients with one cadaveric graft: wait or innovate. AB - Graft shortage continues to prolong waiting times for adults requiring liver transplantation. Living related donor transplantation is possible for only a small minority of adults. The techniques for in situ splitting of the liver used for right and left hepatectomies in living donors were adapted to a combined split-liver-domino procedure to obtain right and left hemiliver grafts from a patient undergoing total hepatectomy with liver transplantation for a metabolic disorder. The two grafts were adequate in size and function for transplantation to two adults with low priority for regular cadaver grafts. More frequent use of split-liver techniques in cadaver donors could considerably reduce the graft shortage and waiting time for adult liver recipients. PMID- 10719028 TI - A spectrum of pulmonary vascular pathology in portopulmonary hypertension. PMID- 10719029 TI - Genotype 1b and severity of posttransplant recurrence of hepatitis C infection- unconvictable felon or wrongly accused? PMID- 10719030 TI - Hypothermia for fulminant hepatic failure: a cool approach to a burning problem. PMID- 10719031 TI - Vancomycin-resistant enterococci in liver transplant recipients. PMID- 10719032 TI - Combined effects of gamma-radiation and ethylene oxide in human diploid fibroblasts. AB - Human diploid VH-10 fibroblasts were pre-exposed to gamma-rays and then treated with ethylene oxide (EtO). In the reverse experiment, the cells were pretreated with EtO and then exposed to gamma-rays. Two different dose rates of gamma-rays were used: a low dose rate (LDR, 0.66 Gy/min) and a high dose rate (HDR, 10 Gy/min). Cell killing, mutagenicity and DNA double-strand breakage were studied in both types of experiment. The induction of mutations in the HPRT locus was studied by selection in medium containing 6-thioguanine. DNA double-strand breakage, measured as fraction of activity released (FAR), was investigated using pulsed field gel electrophoresis. Concerning mutagenesis, it was found that pre exposure of the cells to gamma-radiation (1 Gy) followed by treatment with EtO (2.5 mMh) led to an additive co-interaction, irrespective of dose rate. On the other hand, the reverse experimental procedure (pretreatment with EtO followed by gamma-ray exposure) resulted in an antagonistic effect, which was most pronounced when the HDR was applied. In the latter case, the resultant mutant frequency was two times lower than the sum of the mutant frequencies after the individual treatments. However, the effect of the combined treatment on FAR was different: FAR increased with both combinations of agents used compared with the separate and hypothetically expected effects. Moreover, the HDR exposure led to an additional increase in FAR compared with the LDR one. PMID- 10719033 TI - Altered splicing of the ATDC message in ataxia telangiectasia group D cells results in the absence of a functional protein. AB - The ATDC gene was cloned using functional complementation and complements the radiosensitivity of ataxia telangiectasia (AT) group D cells. Although a number of transcripts have been detected, only a 3.0 kb cDNA found in a HeLa cell cDNA library has been cloned. Since AT group D cells express only a 2.4 kb transcript, efforts were made to clone and sequence this transcript. Using a biotinylated oligonucleotide probe, mRNA preparations were enriched in ATDC-related sequences. After this enrichment, 2.4 kb clones were obtained from the resulting library. The 2.4 kb transcript appears to be untranslated, since no protein from this transcript has been detected in AT group D cells, and this transcript is probably non-functional, since a splicing variation has positioned part of intron 1 near the first methionine codon in exon 1, eliminating most of exon 1 and important functional regions from this transcript. This transcript now has a stop codon located 33 bp in front of the first methionine, which would stop translation after the eleventh amino acid. As a result of these changes, the AT group D cell line (AT5BI) expresses no functional ATDC protein. PMID- 10719034 TI - Induction of genotoxicity by cadmium chloride inhalation in several organs of CD 1 mice. AB - In recent years, the concentration of metals in the environment has increased significantly. Metal compounds, as a group, are among the best-documented human carcinogens, but the mechanisms by which they act are not completely understood. In the present study a cadmium inhalation model in mice was implemented in order to detect the induction of genotoxic damage as single-strand breaks and alkali labile sites in several organs (nasal epithelial cells, lung, whole blood, liver, kidney, bone marrow, brain and testicle) using the single cell gel electrophoresis (SCGE) or Comet assay. We found differences among the studied organs after a single and subsequent inhalations: in the organs analyzed we observed that major DNA damage was induced after a single inhalation; in subsequent inhalations there was a tendency to maintain the same magnitude of damage or in some cases it decreased. A correlation between length of exposure, DNA damage and metal tissue concentration was found. These results suggest that cadmium chloride inhalation induces systemic DNA damage; some organs showed less damage than others (liver, brain, etc.) and this finding could be as a consequence of the capacity to remove the damage induced by long periods of exposure, possibly because of the induction of detoxifying mechanisms such as induction of metallothionein. PMID- 10719035 TI - Characteristics of UV-induced repair patches relative to the nuclear skeleton in human fibroblasts. AB - We have tried to characterize the nucleotide excision repair (NER) events associated with the nuclear skeleton in both repair-proficient and repair deficient human cell lines following UV irradiation. The repair patches were labelled with biotin-16-dUTP and the repair sites were visualized by fluorescence microscopy using fluorescence-conjugated antibodies to biotin. The intensities of repair labelling measured for the three human cell lines of normal, xeroderma pigmentosum group C (XP-C) and Cockayne syndrome group B (CS-B) are in good agreement with their known repair capabilities. Digestion of nuclei with DNase I markedly solubilized the repair patches in normal (3-fold reduction after 1 h post-UV incubation) and transcription-coupled repair (TCR)-defective Cockayne syndrome cells (6-fold reduction after 1 h post-UV incubation). The intensity of repair labelling remained the same in TCR-proficient XP-C cells after DNase I digestion, indicating that the repair events mediated by the TCR pathway are tightly associated with the nuclear skeleton. Treatment with ammonium sulphate after DNase I digestion further reduced the intensity of repair patches in both normal and Cockayne syndrome cells, but not in XP-C cells. The tight association of repair patches generated by the TCR pathway with the nucleoskeleton in XP-C cells reinforces the concept of functional compartmentalization of the nucleus, where NER is highly heterogeneous. PMID- 10719036 TI - Spontaneous and osmium tetroxide-induced mutagenesis in an Escherichia coli strain deficient in both endonuclease III and endonuclease VIII. AB - Thymine glycol, uracil glycol, 5-hydroxycytosine and 5-hydroxyuracil are common base lesions produced by cellular metabolism as well as ionizing radiation and environmental carcinogens. Escherichia coli DNA glycosylase, endonuclease III and endonuclease VIII recognize and remove these lesions from DNA. In this study, we assessed the mutagenic potential of these lesions in the supF gene as a forward mutation target in double-stranded plasmid DNA using an E.coli strain deficient in both endonuclease III and endonuclease VIII. These lesions were introduced into pTN89 DNA by the chemical oxidant osmium tetroxide. Spontaneous supF mutations occurred at a frequency of 3.03x10(-7) and osmium tetroxide-induced at a frequency of 8. 25x10(-7). Sequence analysis of supF mutants revealed that mutations occurred at cytosine sites rather than thymine sites, suggesting that thymine glycol is not the principal premutagenic lesion. In contrast, G:C-->A:T transitions were dominantly detected in the spontaneous and osmium tetroxide induced mutations in the endonuclease III and endonuclease VIII double defective host. In this case, products of cytosine oxidation such as 5-hydroxycytosine, which are the substrate for endonuclease III and endonuclease VIII, were the principal mutagenic lesions. PMID- 10719037 TI - p53 mutations experimentally induced by 8-methoxypsoralen plus UVA (PUVA) differ from those found in human skin cancers in PUVA-treated patients. AB - 8-Methoxypsoralen (8-MOP) plus UVA irradiation (PUVA therapy) has been used for the treatment of psoriasis. PUVA therapy has been associated with an increased risk of developing skin squamous cell carcinoma (SCC). In order to determine the PUVA-induced p53 mutation spectrum, a yeast expression vector harbouring a human wild-type p53 cDNA was incubated with 8-MOP, and UVA irradiated in vitro. PUVA damaged and undamaged DNA was transfected into a yeast strain containing the ADE2 gene regulated by a p53-responsive promoter. An 8-MOP concentration-dependent decrease in survival and increase in mutant frequency were observed. At a fixed 8 MOP concentration, survival decreased and mutant frequency increased as UVA irradiation increased. Eleven mutant clones contained 11 mutations: 10 were single base pair substitutions, the remaining one being a complex mutation. All eight T:A-targeted mutations were at 5'-TpA sites, hallmark mutations of PUVA mutagenesis. Through a rigorous statistical test, the PUVA-induced p53 mutation spectrum appears to differ significantly (P < 0.0002) from that observed in SCC in PUVA-treated patients. The present work demonstrates that a specific PUVA induced mutational fingerprint could be obtained and recognized on human p53 cDNA. This result may suggest that PUVA therapy can be a risk factor for the development of SCC in psoriasis patients through a mechanism not involving the induction of p53 mutations. PMID- 10719038 TI - 1-Aminobenzotriazole inhibits acrylamide-induced dominant lethal effects in spermatids of male mice. AB - Acrylamide (AA) is a germ cell mutagen and induces clastogenic effects predominantly in spermatids of mice. The mechanism of AA clastogenicity has been a matter of dispute. Since the reactivity of AA with DNA is low but is high with proteins containing SH groups, it was suggested that protamine alkylation could be the mechansim of clastogenicity by AA in spermatids. This was substantiated by the observation that the time course of protamine alkylation and dominant lethal effects in spermatids of mice induced by AA was strictly parallel. Another suggestion was that AA may be metabolized by cytochrome P-450 to the epoxide glycidamide (GA), which is then the ultimate DNA-reactive clastogen. This suggestion was based on the similarity of the stage specificity pattern for dominant lethality and heritable translocation induction by AA and GA. To test this latter assumption, 1-aminobenzotriazole (ABT), an inhibitor of P-450 metabolism, was used in the present experiments. Male mice were pretreated with ABT (3x50 mg/kg) on three consecutive days followed by AA treatment (125 mg/kg) on day 4. Parallel groups of animals were treated with AA (125 mg/kg), ABT (3x50 mg/kg) or with the solvent double-distilled water. The experiment was repeated once with slightly varied mating parameters. The results of both experiments showed that ABT inhibited or significantly reduced the AA-induced dominant lethal effects. Thus, the present data support the hypothesis that the AA metabolite GA is the ultimate clastogen in mouse spermatids. PMID- 10719040 TI - Cytokinesis-block micronucleus assay in primary human liver fibroblasts exposed to griseofulvin and mitomycin C. AB - Primary liver fibroblasts were applied in a cytokinesis-block micronucleus assay in combination with fluorescence in situ hybridization (FISH) using two protocols. In protocol A (Prot. A), cytochalasin B (Cyt B) was added at the end of the treatment time directly to the medium containing the standard compounds, whereas in protocol B (Prot. B) the chemical-containing medium was removed and fresh medium with Cyt B was added. The study was performed using the aneugen griseofulvin (GF) and the clastogen mitomycin C (MMC) as standard compounds. With both protocols GF induced a significant increase in MN frequency over controls in a dose-related manner at the lower concentrations tested (7.5 and 15 microg/ml). At the highest dose (30 microg/ml) the aneugen effect was substantially reduced. MN induction obtained with Prot. A was significantly higher ( approximately 3 fold) than with Prot. B at the most effective concentration. The aneugen effect induced by GF did not change when different cell densities were used, but again with Prot. A we obtained the highest effect. MN induced by MMC showed a dose- and time-dependent increase in both protocols. In contrast to GF, the greater clastogenic response induced by MMC in human liver fibroblasts was obtained with Prot. B, approximately 3-fold higher than Prot. A at the most effective concentration and approximately 2-fold with 24 h treatment at 0.17 microg/ml MMC. With GF, the FISH data in human liver fibroblasts (80% C+MN) were fairly consistent with those obtained in the rodent cell lines. In human whole blood cultures, the same dose used in our experiment produced a relatively higher percentage of C+MN. FISH analysis showed that MMC induced mainly MN containing acentric fragments rather than whole chromosomes. In conclusion we have demostrated that chemically induced genetic effects are strongly dependent on the cell culture employed, treatment schedule and intra- and post-treatment experimental conditions. PMID- 10719039 TI - Evaluation of antimutagenic effect of todralazine in cultured lymphocytes. AB - Todralazine, an antihypertensive drug from the hydrazinophthalazine group, significantly decreased the activities of benzo[a]pyrene and mitomycin C in three short-term genotoxicity tests in human lymphocyte cultures. The thioguanine resistance test, the cytokinesis-blocked micronucleus assay and the sister chromatid exchange test were used to demonstrate the antimutagenicity of todralazine. Todralazine lowered the level of free radicals generated by human granulocytes in vitro in the presence of benzo[a] pyrene and also in the presence of the granulocyte activator and tumor promoter phorbol myristate acetate. These results, together with our previous data obtained in the standard bacterial Ames test, strongly suggest that todralazine is a good antimutagen in vitro and deserves further research on its inhibitory action on mutagenesis and carcinogenesis. PMID- 10719041 TI - Pathways of heterocyclic amine activation in the breast: DNA adducts of 2-amino-3 methylimidazo[4,5-f]quinoline (IQ) formed by peroxidases and in human mammary epithelial cells and fibroblasts. AB - Most human mammary carcinomas originate in the epithelial cells of the breast ducts. A potential role of heterocyclic amines (HAs) in the aetiology of this disease has led us to investigate peroxidase-catalysed and stromal (non epithelial) activation of the HA 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), which may subsequently lead to DNA damage in the adjacent human mammary epithelial cells (HMECs). HAs are formed when proteinaceous foods are cooked at high temperature and some, but not all, can cause mammary tumours in rats. Myeloperoxidase (MPO) and lactoperoxidase (LPO) are peroxidase enzymes present in breast secretions. (32)P-post-labelling analysis showed that IQ-DNA adducts were formed after co-incubation of IQ (500 microM) with calf thymus DNA, hydrogen peroxide and either bovine LPO or horseradish peroxidase (HRP). The major HRP mediated IQ-DNA adduct co-migrated on TLC and HPLC with the major adduct formed in HMECs, suggesting a common reactive intermediate (nitrenium ion). IQ-DNA adducts were also formed in extracellular DNA when phorbol myristate acetate stimulated neutrophils (which activate IQ via MPO) were co-incubated with IQ (500 microM) and extracellular plasmid (4 +/- 1 adducts/10(8) nucleotides) or calf thymus DNA (6 +/- 2). Mean adduct formation was five to seven times greater in neutrophil DNA (31 +/- 20). Primary cultures of human mammary fibroblasts or epithelial cells isolated from reduction mammoplasty tissues (n = 4 individuals) were incubated with IQ (500 microM) and formed 2.5 and 14.8 adducts/10(8) nucleotides (mean values), respectively. Our results indicate the possible contribution of stromal cells and breast peroxidases to the metabolic activation of carcinogens in the mammary gland. PMID- 10719042 TI - In vivo cytokinesis blocked micronucleus assay with carbendazim in rat fibroblasts and comparison with in vitro assays. AB - A successful in vivo application of the cytokinesis blocked micronucleus assay for the detection of aneuploidy induced by carbendazim (CARB) was carried out in the granuloma pouch assay. This was performed in two ways: (i) in vivo exposure of the skin fibroblasts to cytochalasin B (cytB) and CARB, by simultaneous injection of both substances into the pouch; (ii) in vivo exposure to CARB followed by in vitro culturing of the fibroblasts in the presence of cytB. Only the first assay was successful. Injection of cytB (with or without the test compound) into the pouch resulted in the induction of binucleate cells in vivo, up to a maximum of 5% at 1 mg cytB/pouch. After injection of CARB (0-50 or 0-10 mg/pouch) and cytB (1 mg) into the pouch, aneuploidy was determined in the isolated binucleate fibroblasts by fluorescence in situ hybridization with a general centromeric probe and combinations of chromosome-specific probes (19p + 19q, 4q + Yq). With all probes, the induction of chromosome loss and/or non disjunction by CARB was very pronounced; at 10 mg CARB/pouch the total malsegregation frequency of chromosomes 4, 19 and Y was approximately 300/1000 binucleate cells. In an in vitro cytokinesis block assay with CARB (0-2.5 microg/ml) in primary skin fibroblasts the induced aneuploidy frequencies were as high as observed in the in vivo assay. The use of two probes for chromosome 19, which enabled the scoring of chromosome breaks in addition to aneuploidy, revealed no significant induction of chromosome breaks by CARB. The frequency of polyploid mononucleate and binucleate cells was decreased after CARB treatment, in both the in vivo and in vitro assays. However, in an additional in vitro assay without cytB a major induction of polyploidy from 2.5 microg/ml CARB and above was observed, showing that cytB may interfere with polyploidy induction. PMID- 10719043 TI - Isolation of DNA probes specific for rat chromosomal regions 19p, 19q and 4q and their application for the analysis of diethylstilbestrol-induced aneuploidy in binucleated rat fibroblasts. AB - DNA probes specific for rat chromosomes 19p, 19q and 4q were isolated, characterized and used for the detection and analysis of diethylstilbestrol(DES) induced aneuploidy. By denaturing and partially reassociating total genomic DNA a new rat repetitive DNA family was isolated, which was located on chromosome 19p21. Sequencing of a number of subclones from cos76-1 and other clones of this so-called 76-family revealed that the repeat units are interrupted with large areas of other (unique) DNA. Consequently, after fluorescence in situ hybridization (FISH) the signals in interphase nuclei are large and spread out. The other two probes, cos25 (chromosome 4q) and cos42-47 (chromosome 19q), were isolated by screening cosmid libraries with probes isolated previously in our laboratory. The repeat unit of cos25 is a 2174 bp long EcoRI unit that contains three Sau3A sites and is tandemly organized. Sequencing of subclones of cos42-47 revealed that this probe was in fact the 5S RNA gene, located on 19q12. In order to determine if these probes were suitable probes for aneuploidy detection, two series of dual colour FISH with the combinations cos25/cos76-1 (4q/19p) and cos42 47/cos76-1 (19q/19p) were carried out on slides from an in vitro micronucleus assay with DES. With all three probes used, an increase in binucleated cells with non-disjunction or chromosome loss was observed in the DES-treated cultures. Scoring of additional micronucleated cells on slides hybridized with the cos25/cos76-1 (4q/19p) probes revealed that the hybridization signal of probe cos25 (4q) was over-represented in the micronuclei of the control cultures. The simultaneous use of the 19q and 19p probes is a particularly valuable tool for the detection of aneuploidy, since it allows distinction between aneugenic and clastogenic events in binucleated cells. Results of this analysis showed that apart from aneuploidy, DES also induced structural chromosome aberrations, although to a lesser extent. PMID- 10719044 TI - Cytogenetic genotoxicity of anti-herpes purine nucleoside analogues in CHO cells expressing the thymidine kinase gene of herpes simplex virus type 1: comparison of ganciclovir, penciclovir and aciclovir. AB - The three anti-herpes nucleoside analogues ganciclovir, penciclovir and aciclovir were investigated as to their recombinogenic [sister chromatid exchange (SCE) inducing] and clastogenic activity in CHO cells expressing the thymidine kinase gene of HSV-1, which is a precondition of therapeutic activity of these drugs. The compounds were applied for the duration of one cell cycle and cytogenetic end points were measured between 0 and 42 h after exposure. Although the nucleoside analogues are quite similar with respect to chemical structure, they differ basically in their genotoxic potency, aberration types induced as well as the time course of chromosomal damage. Aciclovir induced SCEs and chromosomal aberrations immediately after exposure but only in a concentration range much higher than that reached in blood plasma during anti-herpes therapy. The direct genotoxic activity is explained by the obligate chain terminating property of aciclovir upon incorporation into genomic DNA. On the other hand, genotoxic damage caused by ganciclovir and penciclovir is of the delayed type requiring at least one post-exposure cell cycle for its expression. Unlike aciclovir, ganciclovir is an extremely potent inducer of SCEs and chromosome breaks and translocations at concentrations far below those impairing the proliferative activity and triggering apoptosis of the target cells (as shown by our previous investigation). Penciclovir is essentially devoid of genotoxic activity. It induces SCEs only at cytotoxic/apoptotic concentrations, is only weakly clastogenic and induces premature chromosome condensation which appears to result from uncoupling of karyokinesis and cytokinesis. The genotoxic activity of ganciclovir is explained as due to repair processes triggered in the second post exposure replication cycle at the sites of nucleoside analogue incorporation into genomic DNA. The findings have considerable implications with respect to the use of ganciclovir or other antiviral drugs in suicide gene therapy of malignant diseases. PMID- 10719045 TI - Lack of chemopreventive efficacy of DL-selenomethionine in colon carcinogenesis. AB - Epidemiologic observations and laboratory research have suggested that dietary selenium reduces the risk of colon cancer. Selenium-enriched brewer's yeast as a dietary supplement reduces the incidence of and mortality from cancer of the colon in humans. It is not clear whether the observed inhibitory effect is due to selenomethionine, or to other forms of selenium, or to a mixture of the selenium compounds present in selenium-enriched brewer's yeast. Therefore, bioassay described in this study examined the chemopreventive efficacy of 10 and 15 ppm selenomethionine, equivalent to 3.6 and 5.4 ppm as selenium, against azoxymethane (AOM)-induced colon carcinogenesis. At five weeks of age, groups of male F344 rats were fed diets containing 0 (control diet), 10 or 15 ppm selenomethionine. At seven and eight weeks of age, all rats except those in vehicle-treated groups received s.c. injections of AOM at a dose rate of 15 mg/kg body wt. The rats were maintained on their respective diets for 52 weeks and were then sacrificed. Colon tumors were processed and evaluated histopathologically. Colon tumor incidence and multiplicity were analyzed statistically. No obvious toxic effects were observed following dietary administration of 10 or 15 ppm selenomethionine as indicated by body weight gain. Administration of 10 or 15 ppm selenomethionine had no significant effect on colon tumor incidence and multiplicity. This study suggests that i) selenomethionine lacks chemopreventive efficacy against AOM induced colon carcinogenesis and ii) other forms of selenium or a mixture of selenium compounds present in selenium-enriched brewer's yeast need to be evaluated for their chemopreventive efficacy. PMID- 10719046 TI - Metabolism of D-[1,2-13C]glucose and alpha-D-[1,2-13C]glucose pentaacetate in tumoral pancreatic islet cells. AB - Tumoral pancreatic islet cells of the RINm5F line were incubated, in groups of 25x106 cells each, for 120 min at 37 degrees C in media (5. 0 ml) containing either alpha-D-[1,2-13C]glucose pentaacetate (1.7 mM) or both D-[1,2-13C]glucose (1.7 mM) and acetate (8.5 mM). In both cases, the amounts of 13C-enriched metabolites (D-glucose, L-lactate and acetate) and non-enriched metabolites (acetate) recovered in the incubation medium after incubation were close to the initial amount of esterified or non-esterified D-[1, 2-13C]glucose and acetate, respectively. The 13C-enriched metabolites corresponded mainly to double-labelled D-[1, 2-13C]glucose, L-[2,3-13C]lactate and [1,2-13C]acetate. The output of L [2,3-13C]lactate and [1,2-13C]acetate was about 3-4 times lower in the cells exposed to alpha-D-[1,2-13C]glucose pentaacetate than in those incubated with unesterified D-[1,2-13C]glucose. These findings indicate that, despite extensive hydrolysis of alpha-D-[1, 2-13C]glucose pentaacetate in the RINm5F cells, the hexose moiety of the ester is less efficiently metabolized than unesterified D [1, 2-13C]glucose tested at the same molar concentration (1.7 mM) in the presence of 8.5 mM acetate. Thus, a higher utilization of the hexose moiety of alpha-D glucose pentaacetate than that of unesterified D-glucose, as previously documented in isolated pancreatic islets, represents a far-from-universal situation. PMID- 10719047 TI - RGS domain in the amino-terminus of G protein-coupled receptor kinase 2 inhibits Gq-mediated signaling. AB - We have previously shown that not only G protein-coupled receptor kinase (GRK) 2, but also a catalytically inactive Lys220Trp GRK2 decreases endothelin (ET)-1 induced inositol 1,4,5-trisphosphate (IP3) formation, and demonstrated the presence of phosphorylation-independent desensitization mechanism. To clarify the role of GRK2 other than that as a kinase, we characterized an RGS (regulator of G protein signaling)-like domain in the amino-terminus of GRK2. Both GRK2(1-181) and GRK2(54-174) suppressed Ca2+ responses induced by angiotensin II (Ang II) and ET-1, and bound directly with Galphaq but not Galphas nor Galphai3 in the presence of GDP and AlF4-. These results demonstrate that GRK2 regulates Gq mediated signaling negatively by direct interaction between its RGS domain and the transitional state of Galphaq, as well as through phosphorylation of activated receptors by its kinase domain. PMID- 10719048 TI - Bryostatin 1-induced modulation of nucleoside transporters and 2 chlorodeoxyadenosine influx in WSU-CLL cells. AB - WSU-CLL cells, a fludarabine resistant B-cell chronic lymphocytic leukemia cell line, has been shown to exhibit enhanced sensitivity to 2-chlorodeoxyadenosine (2 CdA) following 48-72 h exposure to bryostatin 1. For 2-CdA to manifest its chemotherapeutic activity, it must first enter the cell through one of several specific nucleoside transporter systems. We present data to show that bryostatin 1-induced enhanced influx of 2-CdA is in part the result of bryostatin 1-induced modulation of nucleoside transporters in WSU-CLL cells. The bi-directional equilibrative NBMPR sensitive transporters in WSU-CLL cells were significantly down-regulated 90 min post-exposure to 1-200 nM bryostatin 1. This down regulation was evident up to 144 h. In contrast, WSU-CLL cells exhibited a transient increase in Na+-dependent concentrative 2-CdA influx from 48 to 96 h after bryostatin 1 exposure which was evident for a longer duration than that accounted for by the increase in deocycytidine kinase activity. These data may, in part, explain the enhanced efficacy of 2-CdA seen in WSU-CLL cells following 48-72 h exposure to bryostatin 1. It may raise questions as to the importance of the bi-directional transporters in determining the resistance or sensitivity of CLL cells to 2-CdA or other nucleoside analogues. PMID- 10719049 TI - Increased responsiveness of ventromedial hypothalamic neurons to norepinephrine in obese versus lean mice: relation to the metabolic syndrome. AB - Studies of the effects of acute and chronic norepinephrine (NE) infusion into the ventromedial hypothalamus (VMH) of rodents indicate important roles for VMH NE activities in the development of the obese-glucose intolerant state. Moreover, elevated endogenous levels of NE and/or its metabolites have been observed in a variety of obese-glucose intolerant animal models. We therefore investigated the VMH neuronal electrophysiologic responsiveness to iontophoretically applied NE in lean-euglycemic and obese-hyperglycemic mice. Additionally, the effect of dopamine agonist treatment (which reduces obesity and hyperglycemia) on VMH responsiveness to NE was examined in obese-hyperglycemic mice. Obese (ob/ob) mice were treated daily for 14 days with either bromocriptine (BC, D2 agonist) (10 mg/kg) plus SKF38393 (SKF, D1 agonist) (20 mg/kg) or vehicle. Lean mice were also similarly treated with vehicle. Twenty-seven hours following the final treatment, mice were anesthetized to obtain electrophysiologic responses of glutamate activated VMH neurons to local NE administration. In all three study groups, NE administration inhibited glutamate evoked neuronal activity in the majority (90%) of recorded neurons. No response to NE was observed in the remaining 10% of neurons. Also within all three groups there existed two patterns of response to NE; a) long duration (>2 min) and low threshold (<20 nA) and b) short duration and high threshold. Relative to lean mice, obese mice exhibited a significant 70% increase in average duration of response, 3-fold increase in percent neurons with long duration of response, and 2-fold increase in percent neurons with low threshold of response. BC/SKF treatment of obese mice significantly reduced the percent VMH neurons with long duration and low threshold of response to NE to resemble the VMH neuronal responsiveness to NE observed in lean mice. Increased VMH responsiveness to NE is part of the endogenous neurophysiology of obese hyperglycemic ob/ob mice. Taken together with previous findings mentioned above, the present results suggest that this increased VMH responsiveness to NE contributes to the pathophysiology of the obese-hyperglycemic state. PMID- 10719050 TI - Experimental postoperative adjuvant chemotherapy by UFT using primary tumor amputation model. AB - We evaluated the postoperative adjuvant chemotherapy by UFT using the primary tumor amputation-pulmonary metastasis model. When Lewis lung carcinoma (LLC) primary tumors on the hind foot pad grew palpable, they were amputated on two different days. In experiment (A) (earlier amputation model), micrometastases were detected on the day of amputation only by the histopathological examination. In the experiment (B) (later amputation model), nodules could be determined even by necropsy. Long-term (60-day) consecutive administration of UFT (22 mg/kg/day), which produced no body weight loss, markedly prolonged the survival period in experiment (A) (ILS: over 118%), 1 of the 15 mice being cured. UFT had a relatively weak but significant effect (67% of ILS) in schedule (B). Using the same model, we examined the inhibitory effect of UFT (2-week administration) on the number of metastatic nodules. A significant decrease of metastatic nodules was observed by UFT with both amputation schedules, but its effect was superior with schedule (A). In the same model using Colon 26 PMF-15, UFT markedly prolonged the survival period of mice (150% of ILS) and significantly decreased the metastatic nodules (86% inhibition). The dose of UFT used was relatively low, and did not significantly inhibit the growth of large tumors. However, the sensitivity to the micrometastases was high. These findings suggest that the postoperative adjuvant chemotherapy by the long-term consecutive administration of UFT would be effective for clinical cancer especially in curatively resected cases. PMID- 10719051 TI - The Epstein-Barr virus is rarely associated with esophageal cancer. AB - The Epstein-Barr virus is an agent that causes African Burkitt's lymphoma, infectious mononucleosis, and Hodgkin's disease. It is also related to nasopharyngeal carcinoma and gastric carcinoma. The aim of this study was to evaluate the prevalence of the Epstein-Barr virus in esophageal cancer. Polymerase chain reaction and in situ hybridization were used to detect the Epstein-Barr virus. We detected 103 Epstein-Barr virus positive cells out of 107 of KYSE 273 cells using first standard-PCR. Epstein-Barr virus DNA could not be detected in 30 of the esophageal squamous cell carcinoma cell lines and 2 of the Barrett's esophageal adenocarcinoma cell lines. Out of 77 esophageal cancer patients, 3 cases were found positive for Epstein-Barr virus DNA using polymerase chain reaction. However, by in situ hybridization we found signals in only 1 of the 3 cases, the signal was located in the infiltrating lymphocytes. The Epstein Barr virus is rarely associated with esophageal cancer. PMID- 10719052 TI - Endogenous lectins (galectins-1 and -3) as probes to detect differentiation dependent alterations in human squamous cell carcinomas of the oropharynx and larynx. AB - Expression of glycan determinants for in situ binding is the prerequisite for a productive protein (lectin)-carbohydrate recognition. Labeled tissue lectins as tools are preferable to plant lectins to assess this parameter, because plant and animal lectins with identical saccharide specification can well differ in their profiles of oligosaccharide binding pattern. Due to their relevance in growth control and matrix adhesion the family of galectins (galactoside-binding metal ion independent animal lectins) is receiving increasing utilization in human biology. Employing biotinylated galectin-1 and galectin-3 we studied the expression of binding sites for these galectins in normal human squamous epithelium and human carcinomas from the oropharyngeal region and larynx in relation to the expression of LP-34+ cytokeratins by the procedure of double labeling. Tissue sites accessible for galectin-1 were located in all layers of normal epithelium and in tumor cells. In contrast, galectin-3 binding was suprabasal in the normal epithelium and in tumor cells exhibiting signs of keratinization. These results reveal differences in the localization of accessible sites for the two galectins. Relating to cell development galectin-3 appeared to display affinity to areas with increased extent of differentiation. PMID- 10719053 TI - Expression of vascular endothelial growth factor and thrombospondin-1 in colorectal carcinoma. AB - Solid tumors require neovascularization for growth and metastasis. Vascular endothelial growth factor (VEGF) is a well-characterized inducer of angiogenesis, while, thrombospondin-1 (TSP-1) is thought to be an antiangiogenic factor. In this study, we examined the expressions of these antigens and their relationship with microvessel density, and determined their prognostic significance. One hundred specimens resected from patients with colorectal adenocarcinoma were examined using immunohistochemical methods. Microvessel density, determined by immunostaining for factor VIII-related antigen, was significantly higher in tumors that were VEGF-positive and TSP-1-negative than in other tumors. Patients with VEGF-positive tumors had a significantly worse prognosis than did those with VEGF-negative tumors, and TSP-1 expression was inversely correlated with prognosis. The frequency of hepatic recurrence was significantly higher in patients with tumors that were VEGF-positive and TSP-1-negative than in all other patients. In conclusion, VEGF and TSP-1 are important regulators of tumor angiogenesis, and combined analysis of VEGF and TSP-1 may be useful for predicting recurrence in patients with colorectal adenocarcinoma. PMID- 10719054 TI - The p73 gene is less involved in the development but involved in the progression of neuroblastoma. AB - We performed expression, mutation, loss of heterozygosity (LOH) and fluorescence in situ hybridization (FISH) analyses of the p73 gene in neuroblastomas (NBs). Reverse transcription-polymerase chain reaction (RT-PCR) using primers which can detect both the p73alpha and p73beta transcripts was performed on 30 fresh NBs and 22 NB cell lines. Aberrant expression of the p73 gene was found in 4 (25%) of 16 primary tumors found by mass screening and in 10 (71.4%) of 14 primary tumors found clinically. The rates of expression in these two types of tumors were significantly different (p=0.026, Fisher's exact test). The incidence of aberrant expression of the p73 gene was significantly higher in stage IV patients than in stages I, II, III plus IVS patients (p=0.0236, Fisher's exact test). No homozygous deletions or rearrangements of the p73 gene were found in any samples examined. In addition to the polymorphism in exon 2, a silent mutation (codon 336 GCC/GCT) was found in one primary tumor. LOH of the p73 gene was detected in 5 (15%) of 33 primary NBs using PCR-LOH analysis. FISH analysis showed that all 17 NB cell lines used in this study revealed allelic loss of the p73 gene, while most of them expressed the p73 gene. These results suggest that the p73 gene is not monoallelically expressed in NB. We conclude that the p73 gene is less involved in the development but involved in the progression of neuroblastoma. PMID- 10719055 TI - Impairment by cytochalasin B of the inhibitory action of D-mannoheptulose upon D glucose metabolism in rat pancreatic islets. AB - D-mannoheptulose was recently found to inhibit D-glucose metabolism in hepatocytes and pancreatic islets, whilst failing to do so in parotid cells, erythrocytes and the exocrine pancreas. In the latter three systems, however, the hexaacetate ester of D-mannoheptulose efficiently inhibits D-glucose metabolism. It was proposed, therefore, that the transport of unesterified D-mannoheptulose into cells may be mediated by GLUT2. Since cytochalasin B is known to inhibit D glucose transport into pancreatic islet cells, it was now investigated whether the mould metabolite (0.02 mM) also impairs the inhibitory action of D mannoheptulose (1.0 mM) upon D-glucose metabolism in rat pancreatic islets. The relative extent of D-mannoheptulose inhibitory action on D-[5-3H]glucose utilization and D-[U-14C]glucose conversion to 14CO2, as well as radioactive amino acids and acidic metabolites, was indeed much less marked in the presence of cytochalasin B (13+/-4% inhibition) than in its absence (40+/-3% inhibition). A comparable situation was not observed, however, in the case of glucose stimulated insulin secretion, cytochalasin B augmenting insulin output to the same relative extent in the absence or presence of D-mannoheptulose. These findings support the view that the entry of D-mannoheptulose into cells may be mediated by a cytochalasin B-sensitive transport system, such as the GLUT2 carrier. PMID- 10719056 TI - Interleukin-10 expression in intestine of Crohn disease. AB - Inflammatory bowel diseases are considered to be related to dysregulation of pro- and anti-inflammatory cytokines in the intestinal wall. We investigated the levels of TNFalpha, IFNgamma, and IL-10 mRNA expression in intestinal tissues resected from the patients with Crohn disease (CD) (n=29), ulcerative colitis (UC) (n=8), and controls (n=8) using reverse transcription-polymerase chain reaction (RT-PCR). In addition, we examined the relationship between the expression of these cytokine mRNA and their clinical conditions using CD activity index (CDAI) and Nutritional Surgical Risk Index (NSRI). Compared with controls, tissues in CD showed high levels of TNFalpha and IFNgamma mRNA expression both in inflamed and non-inflamed tissues, and showed high levels of IL-10 mRNA expression in inflamed tissues. In UC, high levels of IL-10 mRNA expression were detected both in inflamed and non-inflamed UC tissues, while those of TNFalpha and IFNgamma were not. In 80% of CD tissues (n=23), levels of IL-10 and TNFalpha expression were interrelated. While the remaining tissues (n=6) showed low levels of IL-10 expression despite high levels of TNFalpha expression in inflamed CD tissues, and 4 of these 6 patients had high CDAI and low NSRI. Furthermore, in low nutritional CD patients (NSRI <40, n=13), the levels of IL-10 mRNA to inhibit pro-inflammatory cytokines were poorer than in good nutritional patients (NSRI >/=40, n=16). These findings suggest the overexpressions of TNFalpha and IFNgamma in CD, and less producibility of IL-10 against these cytokine might lead to development of severe CD. PMID- 10719057 TI - Quantitation of macrophage migration inhibitory factor (MIF) using the one-step sandwich enzyme immunosorbent assay: elevated serum MIF concentrations in patients with autoimmune diseases and identification of MIF in erythrocytes. AB - We raised monoclonal antibodies against human macrophage migration inhibitory factor (MIF), and developed a one-step sandwich enzyme-linked immunosorbent assay (ELISA) method highly specific for human MIF. The ELISA system utilizes a solid phase monoclonal antibody as a capture antibody and a horseradish peroxidase conjugated monoclonal antibody as a detector antibody. We used this ELISA method to evaluate the serum level of MIF in 240 healthy volunteers (140 males and 100 females). We found no significant difference in MIF concentration with respect to age. A significant difference was found with respect to sex, with the mean value (+/- SD) for male subjects of 5.3+/-2.3, and that for female subjects of 4.6+/ 2.3 ng/ml (p<0.05). We next measured the serum MIF contents of patients with autoimmune diseases, and found that MIF levels were significantly elevated in patients with systemic lupus erythematosus and rheumatoid arthritis, 20.0+/-11.0 ng/ml and 21. 7+/-11.2 ng/ml, respectively. Using anti-MIF antibody-immobilized sepharose column chromatography, we discovered for the first time that MIF was present in erythrocytes. Taken together these results suggest that MIF plays a major role in autoimmune diseases and, moreover, potentially induces various patho-logical outcomes in cases of hemolytic disorders. PMID- 10719058 TI - P53 codon 72 polymorphism as a risk factor in the development of HPV-associated non-melanoma skin cancers in immunocompetent hosts. AB - Human papilloma virus (HPV) has been implicated in skin cancer. Also, in human populations, the p53 gene is polymorphic at amino acid 72 of the protein that it encodes. The association between p53 polymorphisms and HPV-associated skin cancer risk has been examined, but the results were conflicting. It was revealed that the arginine form of p53 is more susceptible to degradation by the HPV E6 protein than the proline form and that patients with the arginine form have a higher risk of developing cancer than those with the proline form. The purpose of this study was to examine whether p53 Arg at the polymorphic position 72 could represent a risk factor for patients with high risk HPV-associated malignant skin lesions. The study was conducted on 29 high risk HPV-related skin lesions from Greece. Blood samples from 61 healthy individuals were used as controls. HPV-8 was the most frequent type. There was a difference in the distribution of p53 genotypes between high risk HPV-skin lesions and the controls, and the allele frequency of p53 Arg/Arg was much higher than the controls (65.5% versus 20%). Therefore, it is suggested that p53 Arg homozygosity could represent a potential risk factor for tumorigenesis of the skin. PMID- 10719059 TI - Late effects of anthracycline therapy in childhood on signal-averaged ECG parameters. AB - The aim of this study was to investigate the long-term effects of anthracycline cytostatics upon the frequency-domain characteristics of the signal-averaged electrocardiogram (SAECG) and to evaluate the differences in the frequency content according to gender. At mean follow-up period of 4 years 188 SAECGs were repeatedly performed in 62 childhood cancer survivors, who were in complete remission 1-14 years following anthracycline therapy (mean dose 256 mg/m2). No patient had an abnormal end-of-therapy echocardiogram. The control group consisted of 100 healthy children and young adults. 23% patients vs 5% controls had abnormal area ratio (AR) values (over the 97th percentil of normal controls). Abnormalities in AR remained persistent in 15% of cancer survivors. Frequency domain analysis revealed significantly higher AR 40-100/0-40 Hz in patients after anthracycline therapy than in controls. Within the patient group significantly higher AR were observed in females than in males. Permanent altered frequency components in SAECGs from cancer survivors, evident particularly in female patients, might signal an increased electrical instability in these patients. PMID- 10719060 TI - Secretion of complement components of the alternative pathway (C3 and factor B) by the human alveolar type II epithelial cell line A549. AB - The complement system participates in the local immune system of the lung. In this study, we investigated the secretion of complement components of the alternative pathway (C3 and factor B) in the alveolar type II epithelial cell line A549. The levels of C3 and factor B protein in the culture medium were determined by enzyme-linked immunosorbent assay (ELISA). The C3 and factor B mRNA expression was assessed by reverse transcription polymerase chain reaction (RT PCR). The addition of interleukin (IL)-1beta or tumor necrosis factor (TNF)-alpha induced a dose- and time-dependent increase in C3 and factor B secretion. The addition of IL-6 or interferon (IFN)-gamma also induced a weak but significant increase in C3 and factor B secretion. These responses at the protein level were also observed at the mRNA level. Furthermore, the combination of IL-1beta plus TNF-alpha induced a marked increase in C3 and factor B secretion. Similarly, IL-6 and IFN-gamma potently enhanced IL-1beta- or TNF-alpha-induced C3 and factor B secretion, respectively. In this study, we demonstrated C3 and factor B secretion in A549 cells, and showed that the proinflammatory cytokines, IL-1beta, IL-6, TNF alpha and IFN-gamma, acted as potent inducers of C3 and factor B secretion. It is likely that alveolar type II epithelial cells are the local sites of complement biosynthesis, and that various cytokines act as regulators of this local immune protection system. PMID- 10719061 TI - Leptin expression in colorectal and breast cancer patients. AB - Leptin is a hormone which controls fat metabolism. Leptin plasma levels and adipose tissue mRNA expression were measured in cancer patients. Plasma levels were correlated with TNM staging, cachexia parameters, tumour markers and hormones. Breast and colorectal cancer patients showed blood plasma levels of insulin, TNF-alpha and tumour markers higher than controls. Breast cancer patients, but not colorectal cancer patients, had plasma levels and adipose tissue expression of leptin significantly higher than controls associated with elevated values of estrogen- and progesterone-receptors. These data suggest the possible use of leptin as a clinical marker. PMID- 10719062 TI - Ultra fast identification of Aspergillus species in pulmonary cytology specimens by in situ hybridization. AB - The cytologic diagnosis of pulmonary aspergillosis infection is typically made on a presumptive basis and later confirmed by fungal culture, which may take up to one week to complete. An in situ hybridization (ISH) probe specific for Aspergillus for use in surgical pathology specimens has been developed which has not been used on cytology preparations. We describe a supra-threshold adapted testing (STAT) in situ hybridization test for cytology specimens, which takes less than one hour to finish. We performed ISH on three cases of culture-proven pulmonary aspergillosis and one case with Aspergillus fungal forms but negative cultures to test the feasibility of using this same Aspergillus probe on cytology specimens. Four patients with pulmonary aspergillosis were initially diagnosed by cytologic examination of their respective specimens. The presumptive diagnosis was confirmed by culture to be Aspergillus fumigatus on three cases. ISH on both cytology cytospin and Thin-Prep specimens was performed using an rRNA Aspergillus specific probe. All four cytology specimens exhibited positive staining with the Aspergillus probe. Most, but not all, fungal hyphae were stained with the probe. Even though ISH is more expensive than culture, in situ hybridization can be performed in less than one hour on cytology specimens and may be beneficial for patients in selected clinical circumstances. PMID- 10719063 TI - Binding properties of [3H]gacyclidine (cis(pip/me)-1-[1-(2-thienyl)-2 methylcyclohexyl]piperidine) enantiomers in the rat central nervous system. AB - Gacyclidine (cis(pip/me)-1-[1-(2-thienyl)-2-methylcyclohexyl]piperidine) is a TCP derivative, which exhibits potent neuroprotective properties against glutamate induced neurotoxicity in vitro and in vivo. In order to better understand gacyclidine pharmacological properties, the binding parameters of its enantiomers ((-) and (+)[3H]GK11) were determined in the rat central nervous system (CNS). An autoradiographic study has shown that their binding distributions are correlated with those of N-methyl-D-aspartate (NMDA) receptors throughout the CNS. Globally, the labeling was the highest with (-)[3H]GK11. In the cerebellum, both radioligands similarly labeled the molecular layer. For both radioligands, on telencephalic, cerebellum and spinal cord homogenates, the association and dissociation kinetics were accounted for by multiphasic process. In all regions, (-)[3H]GK11 exhibited the highest affinity in the nanomolar range. The pharmacological study revealed that both enantiomers labeled both high and low affinity sites in all regions. The pharmacological profile of high affinity sites was correlated with those of NMDA receptors. Those of low affinity sites were different in telencephalic and cerebellar homogenates. Overall, this study showed that low affinity sites might constitute a heterogeneous population, which could include sigma receptors in the cerebellum. The autoradiographic study has shown that these sites may be located in the molecular layer. The contribution of low affinity sites to the neuroprotective properties of gacyclidine remains to be investigated. PMID- 10719064 TI - Timecourse and corticosterone sensitivity of the brain, pituitary, and serum interleukin-1beta protein response to acute stress. AB - Activation of peripheral immune cells leads to increases of interleukin-1beta (IL 1beta) mRNA, immunoreactivity, and protein levels in brain and pituitary. Furthermore, IL-1beta in brain plays a role in mediating many of the behavioral, physiological, and endocrine adjustments induced by immune activation. A similarity between the consequences of immune activation and exposure to stressors has often been noted, but the potential relationship between stress and brain IL-1beta has received very little attention. A prior report indicated that exposure to inescapable tailshocks (IS) raised levels of brain IL-1beta protein 2 h after IS, but only in adrenalectomized (and basal corticosterone replaced) subjects. The studies reported here explore this issue in more detail. A more careful examination revealed that IL-1beta protein levels in hypothalamus were elevated by IS in intact subjects, although adrenalectomy, ADX (with basal corticosterone replacement) exaggerated this effect. IL-1beta protein increases were already present immediately after the stress session, both in the hypothalamus and in other brain regions in adrenalectomized subjects, and no longer present 24 h later. Furthermore, IS elevated levels of IL-1beta protein in the pituitary, and did so in both intact and adrenalectomized subjects. IS also produced increased blood levels of IL-1beta, but only in adrenalectomized subjects. Finally, the administration of corticosterone in an amount that led to blood levels in adrenalectomized subjects that match those produced by IS, inhibited the IS-induced rise in IL-1beta in hypothalamus and pituitary, but not in other brain regions or blood. PMID- 10719065 TI - Limitations in the neuroprotective potential of gene therapy with Bcl-2. AB - Considerable attention has focused on the therapeutic transfer of genes with viral vectors into neurons for the purpose of protecting against neurological insults. A number of papers have reported that overexpression of the anti apoptotic protein Bcl-2 can protect neurons both in vitro and in vivo against a variety of necrotic insults. An emerging literature suggests that the availability of energy tends to modulate a neuron towards dying apoptotically, rather than necrotically, in the aftermath of an insult. This suggests that an anti-apoptotic protein such as Bcl-2 should be minimally protective, at best, against purely energetic insults. In support of this idea, we report that overexpression of Bcl-2 with a herpes simplex viral vector fails to protect hippocampal neurons, either in vitro or in vivo, against the electron transport uncoupler 3-acetylpyridine (3AP). As a positive control, the same vector significantly protected against the excitotoxin kainic acid. This finding supports the view that neurotoxicity induced by 3AP is likely to have only minimal apoptotic facets. On a broader level, it suggests some limitations in the neuroprotective potential of gene therapy with Bcl-2. PMID- 10719066 TI - NMDA and non-NMDA receptor-stimulated IkappaB-alpha degradation: differential effects of the caspase-3 inhibitor DEVD.CHO, ethanol and free radical scavenger OPC-14117. AB - The excitotoxic response of striatal neurons to NMDA and non-NMDA receptor agonists involves the nuclear translocation of transcription factor nuclear factor-kappa B (NF-kappaB) due to IkappaB-alpha degradation. Resultant augmentation in c-Myc, p53 and cyclin D1 expression presages the apoptotic-like destruction of these cells in vivo. To differentiate molecular events triggered by intrastriatally injected quinolinic acid (QA, 60 nmol) and kainic acid (KA, 2.5 nmol), we compared the effects of a caspase-3 inhibitor (DEVD.CHO, 8 microgram intrastriatally), a free radical scavenger (OPC-14117; 600 mg/kg, orally) and ethanol (2.14-8.6 micromol, intrastriatally or 25-100 mmol/kg, orally) on changes induced by these glutamatergic agonists on NF-kappaB cascade components and the apoptotic death of rat striatal neurons in vivo. The results indicated that the QA-induced degradation of IkappaB-alpha is almost totally mediated by a caspase-3-dependent mechanism, while KA-induced IkappaB-alpha degradation is only partially dependent on caspase-3. OPC-14117 attenuated the effects of QA but not KA on IkappaB-alpha degradation, suggesting that oxidative stress contributes to the QA- but not the KA-induced degradation of IkappaB alpha. In contrast, ethanol inhibited the KA- but not the QA-induced degradation of IkappaB-alpha and the ensuing DNA fragmentation and loss of striatal GABAergic neurons. It would now appear that NF-kappaB activation in striatal neurons induced by NMDA or KA receptor stimulation involves different biochemical mechanisms. Since excitotoxicity associated with NF-kappaB activation may contribute to neuronal degenerative disorders such as Huntington's disease, a more detailed understanding of biochemical events underlying ionotrophic glutamate receptor-stimulated cell death may assist in the discovery of alternative approaches to interdicting the deleterious consequences of excitotoxic insult. PMID- 10719068 TI - Dorsal raphe nuclear stimulation of SCN serotonin release and circadian phase resetting. AB - Serotonin (5-HT) is strongly implicated in the regulation of mammalian circadian rhythms. However, little is known of the functional relationship between the circadian clock located in the suprachiasmatic nucleus (SCN) and its source of serotonergic innervation, the midbrain raphe nuclei. In previous studies, we reported that electrical stimulation of the dorsal or median raphe nuclei (DRN and MRN, respectively) induced 5-HT release in the SCN. Notably, DRN- but not MRN stimulated 5-HT release was blocked by the 5-HT(1,2,7) antagonist, metergoline, suggesting that the DRN signals to the SCN indirectly via the activation of a 5 HT-responsive multisynaptic pathway. In the present study, pretreatment with the 5-HT(2,7) antagonist, ritanserin, also significantly inhibited DRN-electrically stimulated SCN 5-HT release. However, pretreatment with the 5-HT(1A) antagonist, NAN-190, or the 5-HT(2) antagonists ketanserin and cinanserin had little suppressive effect on this DRN-stimulated 5-HT release. In complementary behavioral trials, electrical stimulation of the DRN during subjective midday caused a 1.3-h advance in the free-running circadian activity rhythm under constant darkness, which was inhibited by metergoline. Collectively, these results are evidence that: (1) DRN-stimulated 5-HT release in the SCN requires the activation of an intermediate target with receptors having 5-HT(7) pharmacological characteristics; (2) electrical stimulation of the DRN induces phase-resetting of the circadian activity rhythm; and (3) activation of 5-HT receptors is necessary for this DRN-stimulated circadian phase-resetting. In view of the dynamic changes in DRN neuronal activity incumbent with the daily sleep activity cycle, and its functional linkages to the SCN and intergeniculate leaflet, the DRN could serve to provide behavioral/arousal state information to various sites comprising the brain circadian system. PMID- 10719067 TI - Chronic fentanyl treatments induce the up-regulation of mu opioid receptor mRNA in rat pheochromocytoma cells. AB - Chronic activation of adenylate cyclase-cAMP-cAMP-dependent protein kinase (PKA) systems by administration of opioid receptor agonists has been considered as one of the mechanisms of opioid tolerance and dependence. Although analysis of the micro opioid receptor (MOR) gene suggests that cAMP-related signal transduction systems regulate the expression of this gene, which transcription factors affect the MOR gene expression in brain and neural cells has not been clarified. This study deals with the effects of fentanyl on MOR mRNA levels in the rat pheochromocytoma cell line (PC12 cells). PC12 cells were cultured in medium with clinically relevant concentrations of fentanyl. The quantitative reverse transcription and polymerase chain reaction (RT-PCR) method was used for determination of MOR mRNA. Treatment of PC12 cells with fentanyl induced the MOR mRNA up-regulation in a concentration- and time-dependent manner. A cAMP analogue also up-regulated MOR mRNA. The intracellular cAMP level increased after fentanyl treatment. A PKA inhibitor blocked the MOR mRNA up-regulation by fentanyl and the cAMP analogue. Expression of a dominant inhibitory Ras also inhibited the MOR mRNA up-regulation. Fentanyl-induced up-regulation of MOR mRNA via activation of cAMP signaling may be important in compensating for the MOR reduction during long term treatment of PC12 cells with fentanyl. The present study could be relevant to understanding the molecular mechanisms of opioids in a state of drug tolerance or dependence, and in patients under anesthesia or being treated for pain. PMID- 10719069 TI - Distinct pattern of neuronal degeneration in the fetal rat brain induced by consecutive transplacental administration of methylmercury. AB - The transplacental neurotoxicity of methylmercury (MeHg) on the fetal rat brain was studied. Adult female rats were administered 1, 2 or 3 mg/kg/day methylmercury chloride (MMC) orally for either 5 or 12 days, and were then mated. They were subsequently administered MMC in the same manner until the end of gestation. On embryonic day 22, a proportion of the fetal brains were histologically examined. Neuronal degeneration of varying degree was detected consistently in the brain stem, cingulate cortex, thalamus and cerebral basal area, including the hypothalamus. The distribution pattern of neuronal damage was different from those in rats treated with MeHg in the postnatal or adult stages. This finding suggests that pathomechanisms in MeHg intoxication operate distinctively in the fetal brain. The offspring derived from dams treated with 1 mg/kg/day MMC for 5 pregestational days and throughout pregnancy survived with inherent brain lesions. This experimental model could be a useful tool for research on the neurotoxicity of MeHg in the human fetal brain. PMID- 10719070 TI - Infarct tolerance induced by intra-cerebral infusion of recombinant brain-derived neurotrophic factor. AB - Neuronal expression of brain-derived neurotrophic factor (BDNF) has been implicated in the mechanism of infarct tolerance (resistance to stroke) (H. Yanamoto et al., Infarct tolerance accompanied enhanced BDNF-like immunoreactivity in neuronal nuclei, submitted to Brain Res.), a process that takes more than 7 days following a preconditioning of repetitive cortical spreading depression (CSD). To investigate whether an elevated level of BDNF protein in the brain solely protects neurons against temporary focal ischemia, recombinant (r)BDNF was infused into the rat neocortex. Recombinant BDNF (or vehicle: saline) was administered into the left neocortex via an implanted osmotic minipump for 2.5, 7, 10 or 14 days pre-ischemia, during ischemia and for 2 days post-ischemia (8 microgram in total) in male Sprague-Dawley rats (n=6 each). Temporary focal ischemia was induced in the left middle cerebral artery (MCA) territory by three-vessel occlusion of bilateral common carotid arteries (CCAs) and MCA for 2 h, and the cerebral infarct volume was analyzed 2 days after ischemia using TTC staining. Regional cerebral blood flow (rCBF) of the left neocortex was monitored after 14 days of intracerebral administration of BDNF or vehicle (n=10 each). The distribution of BDNF following different periods of rBDNF or vehicle-infusion was analyzed using immunohistochemical techniques (n=5 each). In the groups treated with 8 microgram of rhBDNF for 7, 10, or 14 days pre ischemia, there were significant reductions of neocortical infarct volume compared to in the control or vehicle-treated groups (p<0.05). In the rCBF study, there was no significant change after the infusion of 8 microgram rhBDNF for 14 days. In the histological study, a wide distribution of BDNF-like immunoreactivity in the neuronal nuclei in the ipsilateral neocortex was demonstrated after the infusion of 8 microgram rhBDNF for 14 days. The BDNF-like immunoreactivity in the neuronal nuclei was enhanced at the time that the resistance to stroke was achieved by direct intra-cerebral infusion of exogenous rBDNF. Elucidating the function of the BDNF-like protein located in the neuronal nuclei should reveal a new strategy for neuroprotection against ischemic brain attack in humans. PMID- 10719071 TI - Conditioned fear and inescapable shock modify the release of serotonin in the locus coeruleus. AB - The aim of the present study was to investigate the importance of the serotonergic transmission in the locus coeruleus (LC) to conditioned fear. Rats were conditioned to fear by exposing them to noise signal (N), light signal (L) and electric foot shock (S) for 4 days. Control rats were exposed to the same events without receiving S. The LC was superfused with artificial cerebrospinal fluid (aCSF) through a push-pull cannula, and the release of 5-hydroxytryptamine (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) was determined in the superfusate. Motility, blood pressure (BP) and heart rate (HR) were telemetrically recorded. (1) The process of moving animals from their home cage into the grid-floor chamber transiently increased the release rate of 5-HT and the outflow of 5-HIAA in control and naive rats. In conditioned rats, 5-HT release was similarly increased during transfer but was permanently decreased in the grid-floor chamber. Control rats showed phases of enhanced motility in the chamber, while conditioned animals displayed continuous immobility. In naive rats, enhanced motility persisted in the novel environment. (2) Exposure of rats to N+L+S increased the release of 5-HT and the outflow of 5-HIAA to the same extent in conditioned and naive rats. These changes were associated with elevated motility, rise in BP and tachycardia. (3) In conditioned subjects, exposure to N+L in the fifth day led to a pronounced and sustained decrease in the release rate of 5-HT and to tachycardia, while no effects were observed in control rats or naive rats. The findings suggest that conditioned fear attenuates serotonergic neurotransmission within the LC. Telemetric recording of HR proves to be a valuable index for fear and stress processes. PMID- 10719072 TI - Anticonvulsant actions of nefiracetam on epileptic EL mice and their relation to peripheral-type benzodiazepine receptors. AB - Anticonvulsant actions of the nootropic drug nefiracetam were studied using EL mice, an animal model of epilepsy, in which peripheral-type benzodiazepine receptors (PBRs) might be involved in their epileptogenesis. Nefiracetam, when administered orally t o EL mice, inhibited convulsions induced by the PBR agonist, Ro 5-4864, with an ED(50) of 17.2 mg/kg, whereas it did not inhibit the drug-induced convulsions in control DDY mice. When administered intravenously (i.v.) to DDY mice, nefiracetam and other piracetam-like nootropics inhibited the Ro 5-4864-induced convulsions in the sequence of nefiracetam>aniracetam>>oxiracetam, piracetam. Spontaneous EL mouse seizures were also inhibited by these nootropics with a similar rank order of potencies. Binding studies for PBRs, performed on crude membranes of brain tissues of these mice, revealed that [3H]Ro 5-4864 and [3H]PK 11195 bindings were both inhibited by micromolar concentrations of nootropic agents in the sequence of nefiracetam> aniracetam>>oxiracetam, piracetam. The results suggest that nefiracetam may exert an anticonvulsant action through interacting with a low-affinity type of PBR in the brain, and could be developed as a promising therapeutic drug for neurological disorders including epilepsies. PMID- 10719073 TI - Combined c-fos and 14C-2-deoxyglucose method to differentiate site-specific excitation from disinhibition: analysis of maternal behavior in the rat. AB - On the basis of evidence that 14C-2-deoxyglucose (2-DG) autoradiography indicates activity at axonal terminals, whereas c-fos immunocytochemistry indicates activity of neuronal cell bodies, we combined these techniques in adjacent histological brain sections to assess excitatory and disinhibitory synaptic relations in selected sites in female rats in which maternal behavior was elicited by natural parturition, sensitization (7- to 10-day cohabitation with foster pups), or hysterectomy. All individuals in these three groups expressed maternal behavior immediately before 2-DG injection. Controls were non-maternal virgins. Parturient and Hysterectomized groups: elevation (compared with controls) in both 2-DG and c-fos activity in medial preoptic area (MPOA) indicated an increase in its input and output activity, i.e., an excitatory interaction; the MPOA was previously shown to be critical for maternal behavior. Sensitized group: a decrease in 2-DG activity of vomeronasal nuclei (bed nucleus of the accessory olfactory tract, BAOT, and medial amygdala, ME, replicating our previous study) and an elevation in c-fos activity, jointly indicate disinhibition of these nuclei, that were previously shown to modulate pup chemostimulation-induced sensitization. All other sites showed evidence of excitatory input-output relationships (i.e., joint increase in both 2-DG and c fos activity), e.g., bed nucleus of the stria terminalis (BNST), lateral habenula (LHAB), central gray (CG), thalamus (THAL), septum (SEPT), and ventral tegmental area (VTA). The present study demonstrates the feasibility of measuring 2-DG and c-fos activity jointly in adjacent sections of the same brain, thereby providing evidence to distinguish between localized excitation and disinhibition. PMID- 10719074 TI - Spermine is neuroprotective against anoxia and N-methyl-D-aspartate in hippocampal slices. AB - Polyamines were implicated as either neurotoxic or neuroprotective in several models of stroke. Spermine augments the excitotoxicity mediated by the N-methyl-D aspartate (NMDA) receptor because this receptor is activated at micromolar spermine concentrations. However, at higher concentrations, spermine could be neuroprotective because it blocks the NMDA receptor and voltage-activated Ca(2+) channels. In this work, acute hippocampal slices were exposed to 1 mM spermine and either 10 min of anoxia or 0.5 mM NMDA. The percent recovery of population spikes was the measure of neuroprotection. One millimolar spermine was robustly neuroprotective; however, 0.1 mM spermine and 1 mM putrescine were not. The neuroprotective concentration of spermine was higher than the physiological concentration of free spermine. However, during an excitotoxic episode, extracellular Ca(2+) is decreased, enabling the inhibitory activity of lower spermine concentration. In addition, several noxious stimuli trigger the release of intracellular spermine and could raise local levels of spermine. Therefore, it is possible that spermine has a neuroprotective role in vivo. PMID- 10719075 TI - Excitability changes of dorsal root axons following nerve injury: implications for injury-induced changes in axonal Na(+) channels. AB - Electrophysiological recordings were obtained from rat dorsal roots in a sucrose gap chamber to study changes in Na(+) currents following nerve injury. Application of 4-aminopyridine unmasks a prominent and well-characterized depolarization (delayed depolarization) following the action potential. In our previous studies, this potential, which is only present in cutaneous afferent axons, has been shown to correlate with activation of a slow Na(+) current. The delayed depolarization in the dorsal root was reduced 1 week after sciatic nerve ligation, suggesting a reduction in the kinetically slow Na(+) currents on dorsal root axons [control: 44. 2+/-7.3% (n=5); injury: 7.3+/-4.7% (n=5), P<0.001]. The refractory period of the action potential was reduced following nerve injury, in agreement with biophysical studies indicating faster "repriming" of fast Na(+) currents on cutaneous afferent cell bodies. Dorsal root ligation near the spinal cord also results in a reduction in the delayed depolarization. These results indicate that changes in Na(+) channel organization occur on dorsal root axons following either central or peripheral target disconnection, suggesting trophic support can be derived from either the CNS or the PNS. PMID- 10719076 TI - N-terminal tripeptide of IGF-1 (GPE) prevents the loss of TH positive neurons after 6-OHDA induced nigral lesion in rats. AB - The effect of the N-terminal tripeptide of insulin-like growth factor (IGF)-1, glycine-proline-glutamate (GPE), as a neuroprotective agent for nigro-striatal dopaminergic neurons was examined in the present study using a rat model of Parkinson's disease. A unilateral nigro-striatal lesion was induced in rats by injecting 6-hydroxydopamine (6-OHDA) into the right medial forebrain bundle (MFB). GPE (3 microgram) or its vehicle was administered intracerebroventricularly (i.c.v.) 2 h after the 6-OHDA lesion. Tyrosine hydroxylase (TH) immunohistochemistry in the substantia nigra compacta (SNc) and the striatum were examined 2 weeks after the lesion. Following 6-OHDA injection, the number of TH immunopositive neurons in the ipsilateral SNc was reduced. The density of TH immunostaining was also reduced in the ipsilateral SNc and the striatum. Treatment with a single dose of GPE (n=9) significantly prevented the loss of TH immunopositive neurons (p<0. 001) and restored the TH immunoreactivity in both the SNc and the striatum compared with the vehicle control group (n=9, p<0.001). The results suggest that GPE showed promise as a potential treatment for Parkinson's disease. PMID- 10719078 TI - Des-tyrosine(1) dynorphin A-(2-13) improves carbon monoxide-induced impairment of learning and memory in mice. AB - The effects of des-tyrosine(1) dynorphin A-(2-13) (dynorphin A-(2-13)) on carbon monoxide (CO)-induced impairment of learning and memory in mice were investigated using a Y-maze task and a passive avoidance test. The lower percentage alternation and shorter step-down latency of the CO-exposed group indicated that learning and/or memory impairment occurred in mice 5 and 7 days after CO exposure, respectively. Administration of dynorphin A-(2-13) (1.5 and/or 5.0 nmol/mouse, intracerebroventricularly (i.c.v.)) 30 min before behavioral tests improved the CO-induced impairment in alternation performance and the CO-induced shortened step-down latency. We previously reported that dynorphin A-(1-13) improved the impairment of learning and/or memory via kappa opioid receptor mediated mechanisms. To determine whether the effect of dynorphin A-(2-13) was also mediated via kappa opioid receptors, we attempted to block its action using a selective kappa opioid receptor antagonist, nor-binaltorphimine (nor-BNI). Nor BNI (4.9 nmol/mouse, i.c.v.) did not block the effects of dynorphin A-(2-13) on the CO-induced impairment of learning and/or memory. These results indicate that dynorphin A-(2-13) improves impairment of learning and/or memory via a non-opioid mechanism. PMID- 10719077 TI - Modulation of glutamate transporters (GLAST, GLT-1 and EAAC1) in the rat cerebellum following portocaval anastomosis. AB - Glutamate transporters have the important function of removing glutamate released from synapses and keeping extracellular glutamate concentrations below excitotoxic levels. Extracellular glutamate increases in portocaval anastomosis (PCA), so we used a portacaval anastomosis model in rats to analyze the expression of glutamate transporters (GLAST, GLT-1 and EAAC1) in rat cerebellum, 1 and 6 months after PCA, using immunohistochemical methods. In controls, EAAC1 immunoreactivity in Purkinje cells and glial GLAST and GLT-1 immunoreactivities in the molecular layer (ML) increased from young to old rats. One month after PCA, Purkinje cell bodies were not immunostained for neuronal EAAC1 glutamate transporter, whereas glial glutamate transporter expressions (GLAST and GLT-1) were decreased when compared to young controls. In rats with long-term PCA (6 months post-PCA), neuronal and glial glutamate transporter expressions were increased. The expression of the neuronal glutamate transporter EAAC1 was less intense than old controls, whereas glial glutamate transporters (GLAST and GLT-1) increased more than their controls. Since the level of the neuronal glutamate transporter (EAAC1) in long-term PCA did not reach that of the controls, GLAST and GLT-1 glutamate transporters seemed to be required to ensure the glutamate uptake in this type of encephalopathy. EAAC1 immunoreactivity also was expressed by Bergmann glial processes in long-term PCA, but this increase did not suffice to reverse the alterations caused at the early stage. The present findings provide evidence that transitory alteration of glutamate transporter expressions could be a significant factor in the accumulation of excess glutamate in the extracellular space in PCA, which probably makes Purkinje cells more vulnerable to glutamate effect. PMID- 10719079 TI - Phenytoin administration reveals a differential role of pontine reticular formation and periaqueductal gray neurons in generation of the convulsive behaviors of audiogenic seizures. AB - The ventrolateral periaqueductal gray (PAG) and pontine reticular formation (PRF) are implicated in the neuronal network for audiogenic seizures (AGS). The AGS of genetically epilepsy-prone rats (GEPR-9s) culminate in tonic hindlimb extension (TE), and elevated acoustically evoked neuronal firing and burst firing, immediately preceding TE, have been observed in PAG and PRF. This study examined changes in PAG and PRF neuronal firing and behavior in GEPR-9s, following phenytoin administration. Recordings involved 16 PAG and nine PRF neurons in GEPR 9s. Phenytoin in doses (mean, 6. 3 mg/kg) that suppressed TE selectively did not consistently alter PAG neuronal firing. However, these doses of phenytoin resulted in significant (51.6% of control) suppression of PRF neuronal firing. Doses of phenytoin (mean, 8.3 mg/kg), which completely blocked AGS, significantly reduced PAG neuronal firing (64.6% of control), and more greatly suppressed PRF firing (25.8% of control). These results are consistent with a critical role for PRF neurons in generation of TE not evident for PAG. The suppression of PAG and PRF neuronal firing induced by phenytoin with complete seizure blockade is consistent with vital roles for both structures in the seizure network. The differential effects of phenytoin on structures requisite to the seizure network indicate that this experimental approach may be able to identify the most sensitive therapeutic target for anticonvulsant drugs, which could be critical to pharmacological suppression of specific seizure behaviors manifest in various types of convulsions, potentially including human epilepsy. PMID- 10719080 TI - Long-lasting behavioral alterations following a hypoxic/ischemic brain injury in neonatal rats. AB - The characterization of motor and cognitive dysfunctions following a neonatal ischemic injury is a prerequisite to investigate putative pharmacological interventions. To this end, in the present study, we evaluated the long-lasting behavioral alterations occurring after a hypoxic/ischemic injury obtained by the combination of monolateral carotid ligation and exposure to 8% oxygen for 3 h in 7-day-old rats. These animals show a different degree of damage in the side ipsilateral to the occluded artery. Motor coordination, tested both before and after weaning, was not affected, whereas spontaneous activity was increased at weaning but not in the adult age. When tested in an open field after apomorphine administration, most ischemic animals showed a marked turning behavior ipsilateral to the lesioned side. They also had a reduced rate of spontaneous alternation and a marked tendency to visit the arm of the T-maze ipsilateral to the lesion. Injured rats were deficient in performing water maze and T-maze acquisition tests but, when evaluated in a passive avoidance paradigm, no difference from controls was observed. These data indicate that an ischemic insult in neonatal rats causes long-lasting learning deficits and motor behavior asymmetry. These behavioral alterations may represent a useful endpoint for studying the efficacy of potential pharmacological treatments that may improve the behavioral consequences of a perinatal hypoxic/ischemic insult in humans. PMID- 10719081 TI - Acute and long-term effects of 17beta-estradiol on G(i/o) coupled neurotransmitter receptor function in the female rat brain as assessed by agonist stimulated [35S]GTPgammaS binding. AB - Estrogens exert effects on mood, mental state, memory and other central nervous system (CNS) functions by modulating neurotransmitter receptor systems in the brain. Studies were designed to investigate the effect of 17beta-estradiol (E(2)) on agonist-stimulated [35S]GTPgammaS binding in membranes to assess the first step in the intracellular signal transduction cascade in a functional assay following: (1) an acute, one-time bolus subcutaneous injection, or (2) 14-day continuous exposure by a slow-release pellet implanted subcutaneously. In rats treated with E(2) acutely, the maximal response produced by activation of serotonin(1A) (5-HT(1A)) receptors was decreased approximately 25% in the hippocampus, cortex, and amygdala. Similarly, acute E(2) administration desensitized 5-HT(1B) and GABA(B) receptors in hypothalamus and cerebellum, respectively, and cannabinoid receptors in hippocampus and cortex. Although the maximal responses were decreased, acute E(2) treatment did not alter the EC(50) of any of the aforementioned receptors. The incubation of membranes prepared from the cortex of ovariectomized (OVX) rats with E(2) (1 microM) in vitro did not alter 5-HT(1A) or cannabinoid receptor-mediated [35S]GTPgammaS binding. By contrast to acute treatment in vivo, 14-day E(2) administration to OVX rats did not alter the maximal responses produced by activation of 5-HT(1A), 5-HT(1B), GABA(B), or cannabinoid receptors in any of the brain regions examined. Thus, it is concluded that acute E(2) administration in vivo modulates multiple G(i/o) coupled receptors in various regions of the female rat brain. Because these effects are observed only in vivo, it is concluded that cytosolic, nuclear and/or extraneuronal factors are required. PMID- 10719082 TI - Combined treatment with alphaMSH and methylprednisolone fails to improve functional recovery after spinal injury in the rat. AB - To date, relatively little progress has been made in the treatment of spinal cord injury (SCI)-related neurological impairments. Until now, methylprednisolone (MP) is the only agent with clinically proven beneficial effect on functional outcome after SCI. Although the mechanism of action is not completely clear, experimental data point to protection against membrane peroxidation and edema reduction. The melanocortin melanotropin is known to improve axonal regeneration following sciatic nerve injury, and to stimulate corticospinal outgrowth after partial spinal cord transection. Recently, we showed that intrathecally administered alphaMSH had beneficial effects on functional recovery after experimental SCI. Since both drugs have shown their value in intervention studies after (experimental) spinal cord injury (ESCI), we decided to study the effects of combined treatment. Our results again showed that alphaMSH enhances functional recovery after ESCI in the rat and that MP, although not affecting functional recovery adversely by itself, abolished the effects observed with alphaMSH when combined. Our data, thus, suggest that the mechanism of action of MP interferes with that of alphaMSH. PMID- 10719084 TI - Relationship between brain zinc and transient learning impairment of adult rats fed zinc-deficient diet. AB - The relationship between brain zinc and learning behavior was studied based on the data of 65Zn localization in the hippocampal formation. Learning behavior, tested by passive avoidance performance, of 6-week-old rats improved significantly compared to that of 4-week-old rats and it was maintained at 20 weeks of age. When 8-week-old rats were fed zinc-deficient diet for 4 weeks, the learning behavior was significantly impaired. However, it was recovered to almost normal level by feeding with control (zinc-adequate) diet for 5 weeks. These results demonstrate that a proper zinc supply to the brain is necessary for improvement and maintenance of learning ability. Although an appreciable decrease in brain zinc was not observed in the rats fed zinc-deficient diet for 4 weeks, significant decrease of hippocampal zinc was observed in rats fed zinc-deficient diet for 12 weeks. Moreover, synaptosomal zinc in the hippocampal formation and cerebral cortex was significantly decreased by the 12 weeks of zinc deprivation. These results suggest that the decrease of vesicular zinc in the hippocampal formation and cerebral cortex is involved in the transient learning impairment of adults rats. PMID- 10719083 TI - Characterization of strychnine-sensitive glycine receptors in acutely isolated adult rat basolateral amygdala neurons. AB - Large concentrations of the beta-amino acid, taurine, can be found in many forebrain areas such as the basolateral amygdala, a portion of the limbic forebrain intimately associated with the regulation of fear/anxiety-like behaviors. In addition to its cytoprotective and osmoregulatory roles, taurine may also serve as an agonist at GABA(A)- and strychnine-sensitive glycine receptors. In this latter context, the present study demonstrates that application of taurine to acutely isolated neurons from the basolateral amygdala of adult rats causes significant alterations in resting membrane current, as measured by whole-cell patch clamp electrophysiology. Using standard pharmacological approaches, we find that currents gated by concentrations of taurine /=12 weeks to report having a nonsevere event in the year before symptom discovery (p < 0.001). This appeared however, to be an artifact, because the women presenting promptly were recalling events that were closer in time to the research interview. Patient delay was not related to prevalence of depression (p = 0.2) or anxiety (p = 0.8) in the year preceding symptom discovery. CONCLUSION: This study suggests that neither adverse life experiences nor mood disorder in the year before symptom discovery increase the risk of patients with symptoms of breast cancer delaying their presentation to a general practitioner. PMID- 10719135 TI - Cobalamin level is related to self-reported and clinically rated mood and to syndromal depression in bereaved HIV-1(+) and HIV-1(-) homosexual men. AB - OBJECTIVE: An examination of the relationship of plasma cobalamin (vitamin B(12)) level to overall psychological distress, specific mood states, and major depressive disorder was conducted in 159 bereaved men (90 HIV-1(+) and 69 HIV-1( )). METHODS: The relationship of a continuous measure of cobalamin level to psychological distress was examined, while controlling for HIV-1 serostatus, life stressors, social support, and coping styles. RESULTS: Of this sample, 23.9% were either overtly or marginally cobalamin deficient; however, the deficiency rate was not significantly different by HIV-1 serostatus. Cobalamin level was inversely related to self-reported overall distress level and specifically to depression, anxiety, and confusion subscale scores, as well as to clinically rated depressed and anxious mood. Lower plasma cobalamin levels also were associated with the presence of symptoms consistent with major depressive disorder. CONCLUSION: These findings suggest that cobalamin level may be physiologically related to depressed and anxious mood level, as well as to syndromal depression. PMID- 10719136 TI - A presurgical psychosocial intervention for breast cancer patients. psychological distress and the immune response. AB - OBJECTIVE: The present study evaluated the feasibility and potential immunological benefit of a presurgical intervention for breast cancer patients. METHODS: Forty-one newly diagnosed breast cancer patients were randomized into control (standard care) and intervention groups. In addition to standard care, intervention group members received a two-session psychosocial intervention. Blood was drawn at three timepoints: (1) at preintervention; (2) at postintervention/presurgery; and (3) at postsurgery. RESULTS: Examination of the immunological data revealed evidence of suppression of interferon-gamma (IFN gamma) in the control group over time, but not in the intervention group. Secondary findings related to psychological assessment generally paralleled the IFN-gamma results. CONCLUSION: The relevance and applicability of these findings to future breast cancer intervention research is detailed. PMID- 10719137 TI - Stress, dietary restraint and food intake. AB - OBJECTIVE: The purpose of this study was to examine the associations between work stress and nutritional status in relation to dietary restraint in a community sample of adults. METHODS: The design included a cross-sectional and a longitudinal study element. Ninety staff members (58 women and 32 men) of a large department store were assessed on four occasions over a 6-month period with measures of diet, weight, and perceived stress. Work stress was indexed in terms of the hours of work over the past 7 days, which provided an objective indicator of demand. RESULTS: Participants worked an average of 47 hours on the high-work stress session compared with 32 hours on the low-work-stress session. The highest work-stress session was compared with the lowest work-stress session in the longitudinal analyses, and the moderating effects of gender and restrained eating were examined. High-workload periods were associated with higher energy and saturated fat and sugar intake. There was a significant moderating effect of restrained eating, with a hyperphagic response to work stress in restrained eaters, compared with no effect in unrestrained eaters. CONCLUSION: The results indicate that the associations between restraint and stress-induced eating that have been observed in the laboratory extend to the real-life setting. They raise the possibility that restrained eaters are particularly vulnerable to adverse effects of stress on health, through influences on food intake. PMID- 10719138 TI - Schizophrenia: the fundamental questions. AB - Identifying the correct phentotype of schizophrenia is perhaps the most important goal of modern research in schizophrenia. This identification is the necessary antecedent of indentifying the pathophysiology and etiology. A working model is proposed, which suggests that the phenotype should be defined on the basis of abnormalities in neural circuits and a fundamental cognitive process. This type of unitary model may be more heuristic than early ones that were based on heterogeneous signs and symptoms. PMID- 10719139 TI - Familial and genetic mechanisms in schizophrenia. AB - A distinction should be made between genetic aspects of schizophrenia and familial aspects. Genetic aspects of the disease have been reviewed and found to be deficient in many respects. Until recent years, familial factors were assumed to be psychological in origin, but this assumption is now discredited. Research efforts should focus on familial factors that are biological, especially infectious agents that may be transmitted within the family. Most infectious agents are influenced by predisposing genes. The etiology of schizophrenia, then, may turn out to involve biological familial infectious agents that are influenced by susceptibility genes governing the infectious process and the clinical expression of the disease. PMID- 10719140 TI - Schizophrenia as the price that homo sapiens pays for language: a resolution of the central paradox in the origin of the species. AB - The central paradox of schizophrenia is that the condition, apparently genetic in origin, persists in spite of a substantial fecundity disadvantage. The hypothesis is proposed that the predisposition to schizophrenia is a component of Homo sapiens-specific variation associated with the capacity for language. A genetic change (the 'speciation event', predicted to be related to the Xq21.3 to Yp chromosomal transposition that separates Homo sapiens from the great apes) allowed the hemispheres to develop with a 'cerebral torque', reflected particularly in association cortex, from right frontal to left occipital. Variations in the dimension of lateralization are associated with differences in the rate at which verbal and non-verbal ability develops. The nuclear symptoms of schizophrenia can be understood as a failure to establish dominance for a key component - the phonological sequence - of language in one hemisphere, with consequent disruption of the mechanism of 'indexicality' that allows the speaker to distinguish his thoughts from the speech output that he generates and the speech input that he receives and decodes from others. PMID- 10719141 TI - Early and late environmental risk factors for schizophrenia. AB - Although a high proportion of liability to schizophrenia is under genetic control, a number of environmental risk factors have been identified. The earliest of these are complications of pregnancy and birth, though whether these cause or reflect disturbed brain development is not absolutely clear. Neurodevelopmental deviance is also indicated by neurological dysfunction, social, behavioural and cognitive deficits during childhood. Immigrant status is a significant risk factor, especially prominent among the African Caribbean population in England, though the mechanism is unknown. Later environmental risk factors include adverse life events and substance abuse. An additive model of multiple genetic and environmental risk factors of small effect may be too simplistic and an interactive model where genetic predisposition is compounded by environmental effects is more in keeping with current evidence. The nature of such interactions can be explored more fully when susceptibility genes for schizophrenia are identified. PMID- 10719142 TI - Evidence for a compromised dorsolateral prefrontal cortical parallel circuit in schizophrenia. AB - Evidence is reviewed that one of the cognitive-affective parallel circuits in the brain, the dorsolateral prefrontal circuit, is compromised at the level of anatomical, neuropathological and transmitter-related molecules in a subgroup of schizophrenic patients. The dorsolateral prefrontal cortex (DLPFC) comprises a key structure in this circuit. Data supporting a compromised DLPFC includes cognitive deficits, decreased regional metabolism and blood flow activation; disruption of cortical subplate activity (inferred from maldistribution of neurons from the cortical subplate which are required for the orderly neuronal migration during the second trimester and for connectivity of the thalamocortical neurons); decrease in major components of the cortical inhibitory neurotransmitter system; and alterations in the molecules critical for NMDA receptor mediated neural transmission. Thus a great deal of evidence accumulated over the last decade has definitively implicated the dorsolateral prefrontal cortex in the pathophysiology of schizophrenia. Emerging data also confirms neuropathology in the mediodorsal nucleus of the thalamus that projects to the DLPFC. There is currently a consensus that schizophrenia involves epigenetic factors interacting with genetic information in the cells to produce abnormal molecules which when they are associated with abnormal circuits such as the DLPFC, may result in abnormal behavior. Thus, abnormal cortical connections and or altered neurotransmitter related molecules in the DLPFC could explain some of the prominent frontal cognitive disruptions seen in schizophrenia. PMID- 10719143 TI - Lessons from childhood-onset schizophrenia. AB - Childhood-onset schizophrenia (with an onset of psychosis by age 12) is a rare and severe form of the disorder which is clinically and neurobiologically continuous with the adult-onset disorder. Very early onset diseases provide an opportunity to look for more salient or striking risk or etiologic factors in a possibly more homogenous patient population. For the 47 patients with very early onset schizophrenia studied to date, there were more severe premorbid neurodevelopmental abnormalities, more cytogenetic anomalies, and potentially greater family histories of schizophrenia and associated spectrum disorders than later onset cases. There was no evidence for relatively increased obstetrical complications or environmental stress. These data, while preliminary, suggest a very early age of onset of schizophrenia may be secondary to greater genetic vulnerability. It is anticipated that future genetic studies of these patients may provide important etiologic information. PMID- 10719144 TI - Neurobiological findings in early phase schizophrenia. AB - This paper summarises the available information on MRI-determined hippocampal morphometry in first-episode patients as an illustration of the value and interpretation of findings in the neurobiology of early phase schizophrenia. We report a thin slice (1.5 mm) study of 32 first episode and 39 high risk patients which demonstrated significantly smaller hippocampi (right -9%, left -11%) in first episode patients that were of a similar magnitude to those found in chronic patients (right -10%, left -11%) but non-significant volume reductions in high risk individuals, including the 15 subjects who subsequently developed psychoses. Consideration is given to the implications of these findings, including the possible role of early and later neurodevelopmental influences. We present animal data showing that chronic placental insufficiency, as elicited by uterine artery ligation can give rise to substantial reduction (31%) in hippocampal volumes and reflect on other potentially relevant pathophysiological mechanisms, including those that may occur during the early phases of psychotic illnesses, including their prodromes. Greater attention needs to be paid to the study of early phase psychosis in order to obtain a clearer understanding of the nature and time course of neurobiological changes associated with it. Although there is a growing literature on first episode psychosis, there is a striking dearth of information on the neurobiology of the prodrome. PMID- 10719145 TI - Obstetric complications and congenital malformation in schizophrenia. AB - Recent years have witnessed increasingly intense research activity concerning early life somatic trauma and dysmorphogenesis which are associated with the later development of schizophrenia. The two somatic factors that have received the most extensive scientific attention as antecedents of schizophrenia are obstetric complications (OCs) and the congenital malformations termed 'minor physical anomalies' (MPAs). Head circumference (HC) at birth has also been studied as a measure of prenatal cerebral development. A great number of studies indicate clearly that schizophrenia patients have a significantly increased history of OCs, representing many different OCs from pregnancy, labor-delivery and the neonatal period. The probable common denominator of these OCs is oxygen deprivation. Especially labor-delivery OCs relate strongly to brain structure abnormality in ill twins from monozygotic pairs discordant for schizophrenia. Schizophrenia patients very consistently have evidenced an increased frequency of MPAs in the global head, eyes, mouth, ears, hands, feet and limbs. Specific MPAs occur with considerable frequency even among normal comparison subjects, but combination models for specific MPAs efficiently discriminate most patients from comparison subjects. Schizophrenia patients also have significantly reduced HC at birth, independently of gestational age, suggesting a disturbance in prenatal cerebral development, and most frequently observed in female patients. Evidence has thus accumulated, increasingly, for the role of various forms of early trauma and dysmorphogenesis in subsequent schizophrenia, and efforts continue to determine the manner in which these early trauma influence both the early developing brain and the brain of the adult patient with manifest schizophrenia. PMID- 10719146 TI - Molecular genetic studies of schizophrenia. PMID- 10719147 TI - Critical overview of current approaches to genetic mechanisms in schizophrenia research. AB - A genetic etiology to schizophrenia was recognized a century ago by E. Kraepelin [Ein Lehrbuch fur studirende und aerzte, Vol II. Leipzig, Verlagvon, Barth (1899)], yet no clear inherited pattern or mechanism has been established. In the last decade, a new wave of molecular genetic studies of families with schizophrenia has yielded unconvincing evidence for the involvement of multiple putative loci. The task of the next century will be to use genomics to define the true nature of deviant brain growth and development throughout the lifetime of an individual that could result in the perceptual disturbances characterized as schizophrenia. PMID- 10719148 TI - Endogenous retroviruses and schizophrenia. AB - Retroviruses are biologically complex infectious agents which are capable of cellular infection and subsequent integration into the host genome. Retroviruses can exist in an endogenous form in which viral sequences are integrated into the human germline and are vertically transmitted in a Mendelian fashion. The transcriptional activation of these viral sequences in cells within the central nervous system can affect the transcriptional regulation of adjacent genes and result in alterations of neural functioning. This report discusses evidence for a possible role of endogenous retroviruses in the etiopathogenesis of schizophrenia and other human brain diseases. Evidence of endogenous retrovirus activity is manifested by the identification of viral sequences in the brains and cerebrospinal fluids of affected individuals. In addition, affected individuals display evidence of increased activity of virally-encoded reverse transcriptase. The identification of a retroviral component of schizophrenia would be consistent with genetic, environmental, and neurodevelopmental aspects of the disease process. The delineation of a role for retroviruses in disease pathogenesis might lead to new methods for the diagnosis and treatment of schizophrenia. PMID- 10719149 TI - Stereological studies of the schizophrenic brain. AB - Stereological studies have contributed with important results to the understanding of brain abnormalities in schizophrenia. The data obtained from stereological studies of brains from schizophrenic patients, including studies of the thalamus, hippocampus, and cortex, are discussed and suggest a central role of the thalamic nuclei in the etiology of the disease. The basic stereological tools are presented and possible biases in quantification studies are discussed. PMID- 10719150 TI - Basal forebrain in the context of schizophrenia. AB - The human basal forebrain has been notoriously difficult to analyze, and it was only in the last part of the twentieth Century that its various components came into sharper focus. It has now been demonstrated that the main parts of what was previously referred to as the 'substantia innominata' (a neurological equivalent of the geographer's 'terra incognita') belong to nearby and better defined anatomical systems. These include the ventral aspects of the basal ganglia, i.e. the ventral striatopallidal system, extensions of the centromedial amygdala that links it via subpallidal cell columns to the bed nucleus of stria terminalis, i.e. the extended amygdala, and a more or less continuous collection of aggregated and non-aggregated, predominantly large, hyperchromatic projections neurons referred to as the basal nucleus of Meynert. Following a pictorial survey of the basal forebrain, the anatomy of these three systems are described with special emphasize on clinically relevant details. Sections devoted to clinical anatomical correlations emphasize the potential significance of the basal forebrain in the context of schizophrenia. The functional-pathological importance of the cortico-subcortical re-entrant circuits through the ventral striatopallidal system is well recognized in the field of neuropsychiatry. Less appreciated is the fact that both the ventral striatum and the extended amygdala contain prominent collections of islands with small neurons, which have collectively been referred to as 'interface islands'. The abundance of neuroactive substances in the interface islands, and the potential for significant postnatal development of the neurons in these islands make them especially intriguing in the context of the developmental hypothesis of schizophrenia. The basal nucleus of Meynert is also of special interest. Involvement of the basal nucleus of Meynert and its related circuits may well be one of the main reasons for attentional dysfunction and cognitive symptoms in this complex disorder. Finally, we emphasize that changes in the neuronal circuits related to the ventral striatopallidal system, extended amygdala and basal nucleus of Meynert in all likelihood provide the anatomical substrate through which pathologic activities in the medial temporal lobe and prefrontal orbitofrontal structures are translated into disruption of a number of functions ranging from motor activities and basic drives, to personality changes involving stress, mood and higher cognitive functions. PMID- 10719151 TI - Basal ganglia and cerebellar loops: motor and cognitive circuits. AB - The traditional view that the basal ganglia and cerebellum are simply involved in the control of movement has been challenged in recent years. One of the pivotal reasons for this reappraisal has been new information about basal ganglia and cerebellar connections with the cerebral cortex. In essence, recent anatomical studies have revealed that these connections are organized into discrete circuits or 'loops'. Rather than serving as a means for widespread cortical areas to gain access to the motor system, these loops reciprocally interconnect a large and diverse set of cerebral cortical areas with the basal ganglia and cerebellum. The properties of neurons within the basal ganglia or cerebellar components of these circuits resembles the properties of neurons within the cortical areas subserved by these loops. For example, neuronal activity within basal ganglia and cerebellar loops with motor areas of the cerebral cortex is highly correlated with parameters of movement, while neuronal activity within basal ganglia and cerebellar loops with areas of the prefrontal cortex is more related to aspects of cognitive function. Thus, individual loops appear to be involved in distinct behavioral functions. Studies of basal ganglia and cerebellar pathology support this conclusion. Damage to the basal ganglia or cerebellar components of circuits with motor areas of cortex leads to motor symptoms, whereas damage of the subcortical components of circuits with non-motor areas of cortex causes higher order deficits. In this report, we review some of the new anatomical, physiological and behavioral findings that have contributed to a reappraisal of function concerning the basal ganglia and cerebellar loops with the cerebral cortex. PMID- 10719152 TI - Emerging principles of altered neural circuitry in schizophrenia. AB - This paper presents an overview of recent microscopic studies that have sought to define how limbic circuitry may be altered in postmortem schizophrenic brain. The discussion is organized around several basic questions regarding the manner in which interconnections within and between the anterior cingulate cortex and hippocampal formation and involving the glutamate, GABA and dopamine systems may contribute to the pathophysiology of this disorder. The answers to these questions are used to derive several conclusions regarding circuitry changes in schizophrenia: 1) Schizophrenia is not a 'typical' degenerative disorder, but rather it is one in which excitotoxicity may contribute to neuronal pathology, whether or not cell death occurs; 2) Three or more neurotransmitter systems may be simultaneously altered within a single microcircuit; 3) Each transmitter system may show circuitry changes in more than one region, but such changes may vary on a region-by-region basis; 4) The pathophysiology of schizophrenia may involve 'mis-wirings' in intrinsic circuits (microcircuitry) within a given region, but significant changes are probably also present at the level of interconnections between two or more regions within a network (macrocircuitry); 5) While some microscopic findings appear to be selectively present in schizophrenia and be related to a susceptibility gene for this disorder, others may also be present in patients with bipolar disorder; 6) Although some of the circuitry changes seen in schizophrenia and bipolar disorder seem to be associated with neuroleptic exposure, most are not and may reflect the influence of non-specific environmental factors such as pre- and/or postnatal stress; 7) Normal postnatal changes at the level of both macro- and microcircuitry within the limbic system may serve as 'triggers' for the onset of schizophrenia during adolescence. Taken together, these emerging principles can provide a framework for future postmortem studies of schizophrenic brain. PMID- 10719153 TI - GABAergic local circuit neurons and prefrontal cortical dysfunction in schizophrenia. AB - The pathophysiology of schizophrenia involves dysfunction of the dorsolateral prefrontal cortex, and this dysfunction may be related to alterations in GABA neurotransmission. Determining the causes and consequences of altered GABA neurotransmission in schizophrenia requires knowledge of which subpopulations of cortical GABA neurons are affected. The chandelier class of GABA neurons are of interest in this regard because their axon terminals form distinctive vertical arrays (termed 'cartridges') which synapse exclusively with the axon initial segments of pyramidal neurons, the principal class of cortical excitatory neurons. We evaluated the integrity of chandelier neuron cell bodies and axon cartridges in PFC areas 9 and 46 of schizophrenic subjects using immunocytochemical techniques and antibodies against parvalbumin and the GABA membrane transporter GAT-1. Schizophrenic subjects did not differ from matched control subjects in the relative density, laminar distribution or size of parvalbumin-containing neurons. In contrast, the density of GAT-1-immunoreactive chandelier neuron axon cartridges was decreased by 40% in schizophrenic subjects compared to both normal controls and subjects with other psychiatric disorders. The axon terminals of other subclasses of GABA neurons did not appear to be similarly affected. These findings suggest that disturbed GABA neurotransmission in the PFC of schizophrenic subjects may be due to a selective alteration of GAT 1 protein in the axon terminals of chandelier neurons. PMID- 10719154 TI - Possible implications of the dopamine D(3) receptor in schizophrenia and in antipsychotic drug actions. AB - The D(3) receptor may represent an important target for antipsychotic drugs which all bind with high affinity and do not induce upon repeated administration either tolerance or receptor upregulation. The D(3) receptor is localized in brain areas, namely the nucleus accumbens and cerebral cortex, implicated in neural circuits believed to display defective functioning in schizophrenia. Overexpression of the D(3) receptor, which accounts for the behavioral sensitization to levodopa in a rodent model of Parkinson's disease, might also be responsible for the sensitization to dopamine agonists observed in schizophrenia. The appearance of the D(3) receptor during brain development, early in proliferating neuroepithelia and later in neurons from limbic areas, suggests further studies to assess its participation in the neurodevelopmental disorders of schizophrenia. Finally, meta-analysis of approximately 30 studies comprising over 2500 patients indicate that a polymorphism in the coding sequence of the D(3) receptor is associated with a small but significant enhancement of vulnerability to the disease. PMID- 10719155 TI - Glutamate receptor expression in schizophrenic brain. AB - Glutamatergic dysfunction has been suggested as a possible substrate of the pathophysiology of schizophrenia. Of the multiple glutamate receptors, those most commonly implicated in schizophrenia are the ionotropic subtypes, the NMDA, AMPA, and kainate receptors. The expression of the glutamate receptors has been determined at multiple levels of gene expression in postmortem brain samples from schizophrenics and controls; while results have not been entirely consistent from study to study, several generalizations have emerged from this literature: (1) The AMPA receptor is abnormally decreased in expression in the schizophrenic hippocampus, involving decreased levels of subunit transcripts and protein levels, as well as binding sites, (2) similar changes are seen for kainate receptor expression in the hippocampus, and (3) the obligate NMDA receptor subunit, NMDAR1, may be abnormally expressed in some cortical regions in schizophrenia. These data support the hypothesis of abnormal glutamatergic neurotransmission involving the ionotropic glutamate receptors in schizophrenia. PMID- 10719156 TI - D(1) receptors in prefrontal cells and circuits. PMID- 10719157 TI - Serotonin model of schizophrenia: emerging role of glutamate mechanisms. AB - The serotonin (5-HT) hypothesis of schizophrenia arose from early studies on interactions between the hallucinogenic drug LSD (D-lysergic acid diethylamide) and 5-HT in peripheral systems. More recent studies have shown that the two major classes of psychedelic hallucinogens, the indoleamines (e.g., LSD) and phenethylamines (e.g. , mescaline), produce their central effects through a common action upon 5-HT(2) receptors. This review focuses on two brain regions, the locus coeruleus and the cerebral cortex, where the actions of indoleamine and the phenethylamine hallucinogens have been shown to be mediated by 5-HT(2A) receptors; in each case, the hallucinogens (via 5-HT(2A) receptors) have been found to enhance glutamatergic transmission. In the prefrontal cortex, 5-HT(2A) receptors stimulation increases the release of glutamate, as indicated by a marked increase in the frequency of excitatory postsynaptic potentials/currents (EPSPs/EPSCs) in the apical dendritic region of layer V pyramidal cells; this effect is blocked by inhibitory group II/III metabotropic glutamate agonists acting presynaptically and by an AMPA/kainate glutamate antagonist, acting postsynaptically at non-NMDA glutamate receptors. A major alternative drug model of schizophrenia, previously believed to be entirely distinct from that of the psychedelic hallucinogens, is based on the psychotomimetic properties of antagonists of the NMDA subtype of glutamate receptor (e.g., phencylidine and ketamine). However, recently it has been found that many of the effects of the NMDA antagonists may also (1) involve 5-HT(2A) receptors and (2) be mediated through excess activity at non-NMDA (i.e., AMPA/kainate) glutamate receptors. Moreover, pharmacological manipulations of glutamate transmission (e. g., by inhibitory metabotropic glutamate agonists) provide unexpected parallels between the actions of these two classes of drugs. Given an emerging recognition of the importance of alterations in glutamatergic transmission in the actions of both psychedelic hallucinogens an NMDA antagonists, this review concludes with of implications for the pathophysiology and therapy of schizophrenia. PMID- 10719158 TI - The DARPP-32 knockout mouse. PMID- 10719159 TI - Dysfunctional brain dopamine systems induced by psychotomimetic NMDA-receptor antagonists and the effects of antipsychotic drugs. AB - Clinical studies utilizing imaging techniques demonstrate that classical antipsychotic drugs, such as haloperidol, in clinically effective doses display around 75% dopamine (DA)-D(2) receptor occupancy in the brain. In contrast, the atypical antipsychotic drug clozapine is even more effective at only about 45% D(2)-receptor occupancy. Yet at this D(2)-receptor occupancy classical antipsychotics are not effective, raising the question of which other receptors may be involved in mediating the atypical antipsychotic profile of clozapine and other atypical antipsychotics. The present paper describes experimental work aimed at elucidating this critical question, utilizing the phencyclidine (PCP) model of schizophrenia in combination with studies of typical and atypical antipsychotics as well as various specific receptor blocking agents. Both electrophysiological methods, i.e. single cell recording from DA neurons in the ventral tegmental area (VTA), and biochemical analysis of biogenic amines such as DA following microdialysis in difference DA terminal areas in the brain, were used. In addition, behavioural measurements using the conditioned avoidance response (CAR) paradigm and assessments of locomotor activity were utilized. Experiments with functional inactivation of the medial frontal cortex (mPFC) in the rat as well as with MK-801 and other antagonists at central NMDA-receptors revealed that following systemic administration of schizophrenomimetic NMDA receptor antagonists a profound dysregulation of the mesocorticolimbic DA system occurs, severely impairing the dynamic physiological response range of the neurons. Specifically, DA neurons which largely project to the mPFC showed a profound loss of burst firing, whereas VTA-DA neurons, which mainly project subcortically, showed an increased monotonous high-frequency firing with increased DA output from nerve terminals and concomitant behavioural activation. Significantly, drugs with a prominent 5-HT(2A)-receptor blocking action could effectively restore the burst firing mode, i.e. phasic responsivity, in mesocortically projecting DA neurons, and also potentiate the CAR suppressant effect of the selective D(2)/D(3)-receptor antagonist raclopride without increasing catalepsy scores. The selective alpha(1)-adrenoreceptor antagonist prazosin effectively suppressed both the stereotyped, high-frequency firing of subcortically projecting DA neurons following systemic MK-801 and the concomitant behavioural, i.e. locomotor, activation. In addition, the MK-801 evoked DA release in the nucleus accumbens was suppressed. A similar effect was seen also with AMPA-receptor antagonists when applied locally into the VTA and, in addition, systemic administration of chemically different AMPA-receptor antagonists caused a CAR-suppressant effect similar to both classical and atypical antipsychotic drugs. These results and other data showing a clearcut difference between typical and atypical antipsychotic drugs on DA output in the shell and core, respectively, of the nucleus accumbens, suggest that both the 5 HT(2A)- and the alpha(1)-adrenoreceptor blocking effects of a number of atypical antipsychotic drugs in all probability contribute to their antipsychotic effect. Moreover, our results indicate that AMPA-receptor antagonists may possess an atypical antipsychotic profile. PMID- 10719160 TI - Gating of information flow within the limbic system and the pathophysiology of schizophrenia. AB - Although first thought of as a dopaminergic disorder, there is little direct evidence to support a primary pathology in the dopamine system as the etiological factor in schizophrenia. In contrast, evidence is amassing in support of a cortical disturbance in this disorder; one consequence of which is a disruption in the cortical regulation of subcortical dopamine systems. Our studies show that the hippocampus plays a major role in this interaction, in that, along with the dopamine system, it provides a gating influence over information flow from the prefrontal cortex at the level of the nucleus accumbens. Moreover, chemically induced disruption of the development of the hippocampus and entorhinal cortex were found to lead to pathophysiological changes in these interactions in the limbic system of adult rats. Therefore, schizophrenia is proposed to be a developmentally-related disorder, in which disruption of the hippocampal influence over the limbic system during ontogeny results in a pathological alteration of corticoaccumbens interactions in the adult organism. PMID- 10719161 TI - Network interactions in schizophrenia - therapeutic implications. AB - Research into the role of neurotransmitters and neural networks in the pathogenesis of schizophrenia has been remarkably successful in recent years. The hypothesis postulating a dopamine dysfunction, which has for a long time been supported only by indirect evidence, has received direct support by means of sophisticated imaging techniques. Interactions between dopamine and several other neurotransmitters in complex neural networks have been revealed, largely thanks to the advent of an array of new pharmacological probes. Two major pharmacological models of schizophrenia, based on hyperdopaminergia and hypoglutamatergia, respectively, are ready for clinical testing. In addition, the hypothesis of network stabilization as a major therapeutic strategy in psychiatry and neurology has now reached the 'proof-of-concept' level. From a therapeutic perspective, several ongoing and forthcoming clinical trials, using drugs acting on dopaminergic, serotonergic and glutamatergic receptors, give rise to optimism. PMID- 10719162 TI - Eye movements and the search for the essence of schizophrenia. AB - Clinical Schizophrenia has eluded precise description because of its protean phenotypic manifestations and variable clinical course, a variability that also makes it difficult to discover genetic linkages. Co-familial traits have higher recurrence risk rates than schizophrenia itself and might serve as pointers to the underlying physiology of schizophrenic illness. A dysfunction of smooth pursuit eye movements is one such co-familial trait that occurs in about 40 to 80% of schizophrenic patients and about 25 to 40% of their first degree relatives. The eye movement abnormality appears only when the subject tracks a moving target. We have traced this abnormality to a deficit in velocity sensitivity, a function that is regulated by a specific central nervous system network that includes the middle temporal and medial superior temporal areas of the extra-striate cortex. The higher familial recurrence risk of abnormal eye tracking, compared with that of clinical schizophrenia (about 5 to 8%), suggests that schizophrenic psychosis may be the rare form of a more prevalent disorder whose symptoms are much milder and more benign than the cognitive and behavioral disturbances of the clinical psychosis. From this vantage point, abnormal eye tracking can be viewed as one pleiotropic manifestation of schizophrenia, considered broadly, just as the cafe-au-lait spots of neurofibromatosis are a more benign and more frequent manifestation of that disease than are the neurofibromata. PMID- 10719163 TI - Explaining the symptoms of schizophrenia: abnormalities in the awareness of action. AB - We propose that the primary cognitive deficit associated with delusions of control is a lack of awareness of certain aspects of motor control. This problem arises because of a failure in the mechanism by which the predicted consequences of an action are derived from a forward model based on the intended sequence of motor commands. This problem leads to a number of behavioural consequences, such as a lack of central error correction, many of which have been observed in patients with delusions of control and related symptoms. At the physiological level, delusions of control are associated with over-activity in parietal cortex. We suggest that this over-activity results from a failure to attenuate responses to sensations of limb movements even though these sensations can be anticipated on the basis of the movements intended. The lack of attenuation may arise from long range cortico-cortical disconnections which prevent inhibitory signals arising in the frontal areas which generate motor commands from reaching the appropriate sensory areas. PMID- 10719164 TI - The limbic cortex in schizophrenia: focus on the anterior cingulate. PMID- 10719165 TI - The role of endogenous sensitization in the pathophysiology of schizophrenia: implications from recent brain imaging studies. AB - Long-term sensitization is a process whereby exposure to a given stimulus such as a drug or a stressor results in an enhanced response at subsequent exposures. Sensitization of mesolimbic dopamine systems has been postulated by several authors to underlie the development of dopaminergic abnormalities associated with schizophrenia. In this review, core features of stimulant-induced sensitization of dopamine systems in rodents are briefly reviewed, as well as the behavioral and clinical evidence suggesting the relevance of this process to drug-induced psychosis and schizophrenia. Results of recent brain imaging studies relevant to the question of sensitization in schizophrenia are then discussed. These studies indicate that schizophrenia is associated with increased amphetamine-induced dopamine release. This exaggerated response was detected in patients experiencing an episode of clinical deterioration but not in clinically stable patients. Since increased stimulant-induced dopamine release is a hallmark of sensitization, these results support the view that schizophrenia is associated with a process of endogenous sensitization. Based on the preclinical evidence that dopamine projection to the prefrontal cortex acts as a buffer that oppose the development of sensitization in subcortical dopamine projections, we propose that, in schizophrenia, neurodevelopmental abnormalities of prefrontal dopaminergic systems might result in a state of enhanced vulnerability to sensitization during late adolescence and early adulthood. It is also proposed that D(2) receptor blockade, if sustained, might allow for an extinction of this sensitization process, with possible re-emergence upon treatment discontinuation. A better understanding of the neurocircuitry associated with endogenous sensitization and its consequence in schizophrenia might be important for the development of better treatment and relapse prevention strategies. PMID- 10719166 TI - Development of novel antipsychotic drugs. AB - The development of the current generation of antipsychotics has depended on animal, biochemical and pharmacological models of the action of drugs discovered serentipidously. They have focused on the dopamine hypothesis of schizophrenia. More recent studies of the structural abnormalities in schizophrenia, coupled with an understanding of the genetics and developmental biology of brain development, point to a new generation of novel medicines and therapies for patients with this disorder. PMID- 10719167 TI - Schizophrenia and the mechanisms of conscious integration. AB - This article considers the possibility that defective interactions among distributed brain areas may underlie certain dysfunctions of conscious integration such as those seen in schizophrenia. Recent experimental evidence obtained using whole-head magnetoencephalography during binocular rivalry is first reviewed. The results outline a cortical network that underlies conscious integration in the normal brain. This network is not localized to a small part of the brain but it is distributed over frontal, parietal, temporal, and occipital areas. Large-scale simulations of the dynamics of thalamocortical integration are then examined. These studies indicate that several factors can affect the rapid integration of the activity of distributed thalamocortical regions and the resulting behavioral performance. These simulations show that an altered dynamics of corticothalamic and corticocortical re-entrant circuits can result from increased conduction delays, blockade of voltage-dependent connections, reduced synaptic density, and disruptions of the local connectivity within a single cortical area. It can also result from alterations in the activity of diffuse ascending systems that lead to defective reinforcement of integrated activity patterns. Finally, the article briefly reviews theoretical measures of the integration of multiple brain areas, such as measures of functional clustering. These measures have been applied to PET data obtained from schizophrenic subjects and controls while performing cognitive tasks. The results show a change in the functional interactions among distributed brain areas in schizophrenics despite the absence of a change in activation patterns. The possibility is raised that disruption of re-entrant interactions among cortical areas may contribute to the pathophysiology of schizophrenia. PMID- 10719168 TI - Schizophrenia: pathophysiological mechanisms--a synthesis. AB - A Nobel Symposium, 'Schizophrenia: Pathophysiological Mechanisms' was held in Stockholm, October 1-3, 1998. The topics ranged from etiology, genetics, neuropathology, neurotransmitter, brain imaging, to integrative aspects including cognition and language. The collective amount of information is already enormous and will rapidly increase with advances in molecular pathology, gene transcript profiling and non-invasive imaging techniques. Probably reflecting the complexity is the absence of a disease model that can accommodate the data and explain its roots. In the process of moving from simplistic mechanisms involving single or a few interacting neurotransmitters or circuits and turning to more complex interactive models, there is a need for improved data storage and retrieval. Stochastic search in chemical libraries for potential new drug candidates coupled with rational approaches in their final design may improve therapeutic efficacy. PMID- 10719169 TI - Cloning of the bovine leukemia virus proteinase in Escherichia coli and comparison of its specificity to that of human T-cell leukemia virus proteinase. AB - The proteinase of bovine leukemia virus (BLV) was cloned into pMal-c2 vector with N-terminal or with N- as well as C-terminal flanking sequences, and expressed in fusion with maltose binding protein. The proteinase self-processed itself from the fusion protein during expression and formed inclusion bodies. The enzyme was purified from inclusion bodies by cation-exchange chromatography followed by gel filtration. Specificity of the enzyme was compared to that of human T-cell leukemia proteinase type 1. Although the two viruses belong to the same subfamily of retroviruses, the differences in their proteinase specificity, based on kinetics with oligopeptide substrates representing naturally occurring cleavage sites as well as on inhibition pattern, appear to be pronounced. PMID- 10719170 TI - A new cytolysin from the sea anemone, Heteractis magnifica: isolation, cDNA cloning and functional expression. AB - We purified a new cytolysin (HMgIII) from the sea anemone, Heteractis magnifica. HMgIII, which has a molecular mass of approximately 19 kDa, functions as both a cytolysin and a hemolysin. The full-length HMg III cDNA was obtained by reverse transcriptase-polymerase chain reaction, using primers designed from its N terminal amino acid sequence and an internal conserved region of two other sea anemone cytolysins: equinatoxin II (EqT II) and cytolysin III. The cDNA contained an open reading frame of 633 bp, which encodes a protein of 211 amino acids. The nascent HMg III protein contained a prepropeptide of 34 amino acids, which includes a signal peptide of 19 amino acids. The mature HMg III has a predicted molecular mass of 19 kDa and a pI of 9.1, and shares 91%, 89%, 65% and 63% amino acid sequence similarity with cytolysin III, cytolysin ST I, tenebrosin-C and equinatoxin (EqT II), respectively. The predicted secondary structure of the mature HMg III comprises 16% alpha-helix, 23% extended strand and 60% random coils. The characteristic amphiphilic alpha-helix of cytolysins is located at the N-terminus of the processed HMg III. Recombinant HMg III (rHMg III) was expressed in Escherichia coli as a fusion protein containing a 6xHisTag at the N-terminus. The hemolytic and cytotoxic activities of the purified rHMg III were comparable to those of the native HMg III. The hemolytic activities of both proteins were similarly potentiated with 8-anilino-1-naphthalenesulfonate (ANS). Increasing the length of the peptide tag on the N-terminal of rHMg III correlated with decreasing hemolytic activity, thus confirming the importance of the N-terminal amphiphilic alpha-helix for its cytolytic activity. PMID- 10719171 TI - Cysteinyldopaenkephalins: synthesis, characterization and binding to bovine brain opioid receptors. AB - The reaction of opioid peptides with mushroom tyrosinase in the presence of an excess of a thiol compound gives rise to cysteinyldopaenkephalins (CDEnks). The major product is represented by the 5-S-CDEnk (80%) and the minor one by the isomer 2-S-CDEnk (20%). The adducts between leucine-enkephalin (Leu-enk) and cysteine have been isolated by high performance liquid chromatography (HPLC) and identified by amino acid analysis and electrospray ion mass spectrometry. 5-S CDEnk is able to bind to opioid receptors in bovine brain membranes. Its binding affinity is higher for delta than for mu receptors and about 8-fold lesser than that exploited by Leu-enk. In the presence of the peroxidase/H(2)O(2) system, CDEnks can be converted into the corresponding pheo-opiomelanins. PMID- 10719172 TI - Disulfide bond structure of the atrial natriuretic peptide receptor extracellular domain: conserved disulfide bonds among guanylate cyclase-coupled receptors. AB - The disulfide bond structure of the extracellular domain of rat atrial natriuretic peptide (ANP) receptor (NPR-ECD) has been determined by mass spectrometry (MS) and Edman sequencing. Recombinant NPR-ECD expressed in COS-1 cells and purified from the culture medium binds ANP with as high affinity as the natural ANP receptor. Reaction with iodoacetic acid yielded no S carboxymethylcysteine, indicating that all six Cys residues in NPR-ECD are involved in disulfide bonds. Electrospray ionization MS of NPR-ECD deglycosylated by peptide-N-glycosidase F gave a molecular mass of 48377.5+/-1.6 Da, which was consistent with the presence of three disulfide bonds. Liquid chromatography MS analysis of a lysylendopeptidase digest yielded three cystine-containing fragments with disulfide bonds Cys(60)-Cys(86), Cys(164)-Cys(213) and Cys(423) Cys(432) based on their observed masses. These bonds were confirmed by Edman sequencing of each of the three fragments. No evidence for an inter-molecular disulfide bond was found. The six Cys residues in NPR-ECD, forming a 1-2, 3-4, 5 6 disulfide pairing pattern, are strictly conserved among A-type natriuretic peptide receptors and are similar in B-type receptors. We found that in other families of guanylate cyclase-coupled receptors, the Cys residues involved in 1-2 and 5-6 disulfide pairs are conserved in nearly all, suggesting an important contribution of these disulfide bonds to the receptor's structure and function. PMID- 10719173 TI - A helix initiation motif, XLLRA, is stabilized by hydrogen bond, hydrophobic and van der Waals interactions. AB - Five partially overlapping synthetic peptides containing the N-terminal portion of the leucine zipper (LZ)-like domain of human immunodeficiency virus envelope glycoprotein gp41 were used to deduce the helix initiation site. Circular dichroism (CD) data suggested a strong helix-inducing motif, LLRA. The coupling constant and nuclear Overhauser effect (NOE) results obtained from nuclear magnetic resonance experiments in 20% trifluoroethanol aqueous solution at 280 K for the four decapeptides under study suggested that the motif XLLRA, where X is a group or an amino acid residue capable of forming hydrogen bond to arginine, constitutes a helix nucleation core. A similar conclusion was reached for a pentadecapeptide in water, suggesting that the result was not dependent on both chain length and the helix promoting medium. Detailed analysis of NOE and CD data from the four decapeptides indicated that the acetyl group and asparagine had a strong tendency to be helix N-capping, in confirmation of previous studies. Molecular modeling using restraints derived from NOE data showed that van der Waals, hydrophobic interactions and hydrogen bonds contribute synergetically to the stability of the core structure. The concept of nucleation core consisting of a few amino acids may be generally applied in proton design and folding studies. PMID- 10719174 TI - Matrix metalloproteinase inhibition by green tea catechins. AB - We have investigated the effects of different biologically active components from natural products, including green tea polyphenols (GTP), resveratrol, genistein and organosulfur compounds from garlic, on matrix metalloproteinase (MMP)-2, MMP 9 and MMP-12 activities. GTP caused the strongest inhibition of the three enzymes, as measured by fluorescence assays using gelatin or elastin as substrates. The inhibition of MMP-2 and MMP-9 caused by GTP was confirmed by gelatin zymography and was observed for MMPs associated with both various rat tissues and human brain tumors (glioblastoma and pituitary tumors). The activities of MMPs were also measured in the presence of various catechins isolated from green tea including (-)-epigallocatechin gallate (EGCG), (-) epicatechin gallate(ECG), (-)-epigallocatechin (EGC), (-)-epicatechin (EC) and (+)-catechin (C). The most potent inhibitors of these activities, as measured by fluorescence and by gelatin or casein zymography, were EGCG and ECG. GTP and the different catechins had no effect on pancreatic elastase, suggesting that the effects of these molecules on MMP activities are specific. Furthermore, in vitro activation of proMMP-2 secreted from the glioblastomas cell line U-87 by the lectin concanavalin A was completely inhibited by GTP and specifically by EGCG. These results indicate that catechins from green tea inhibit MMP activities and proMMP-2 activation. PMID- 10719175 TI - Interaction of taxol with human serum albumin. AB - Taxol (paclitaxel) is an anticancer drug, which interacts with microtuble proteins, in a manner that catalyzes their formation from tubulin and stabilizes the resulting structures (Nogales et al., Nature 375 (1995) 424-427). This study was designed to examine the interaction of taxol with human serum albumin (HSA) in aqueous solution at physiological pH with drug concentrations of 0.0001-0.1 mM, and HSA (fatty acid free) concentration of 2% w/v. Gel electrophoresis, absorption spectra and Fourier transform infrared (FTIR) spectroscopy with self deconvolution and second-derivative resolution enhancement were used to determine the drug binding mode, binding constant and the protein secondary structure in the presence of taxol in aqueous solution. Spectroscopic evidence showed that taxol-protein interaction results into two types of drug-HSA complexes with overall binding constant of K=1.43 x 10(4) M(-1). The molar ratios of complexes were of taxol/HSA 30/1 (30 mM taxol) and 90/1 (90 mM taxol) with the complex ratios of 1.9 and 3.4 drug molecules per HSA molecule, respectively. The taxol binding results in major protein secondary structural changes from that of the alpha-helix 55 to 45% and beta-sheet 22 to 26%, beta-anti 12 to 15% and turn 11 to 16%, in the taxol-HSA complexes. The observed spectral changes indicate a partial unfolding of the protein structure, in the presence of taxol in aqueous solution. PMID- 10719176 TI - Destabilase from the medicinal leech is a representative of a novel family of lysozymes. AB - Intrinsic lysozyme-like activity was demonstrated for destabilase from the medicinal leech supported by (1) high specific lysozyme activity of the highly purified destabilase, (2) specific inhibition of the lysozyme-like activity by anti-destabilase antibodies, and (3) appreciable lysozyme-like activity in insect cells infected with recombinant baculoviruses carrying cDNAs encoding different isoforms of destabilase. Several isoforms of destabilase constitute a protein family at least two members of which are characterized by lysozyme activity. The corresponding gene family implies an ancient evolutionary history of the genes although the function(s) of various lysozymes in the leech remains unclear. Differences in primary structures of the destabilase family members and members of known lysozyme families allow one to assign the former to a new family of lysozymes. New proteins homologous to destabilase were recently described for Caenorhabditis elegans and bivalve mollusks suggesting that the new lysozyme family can be widely distributed among invertebrates. It remains to be investigated whether the two enzymatic activities (isopeptidase and lysozyme like) are attributes of one and the same protein. PMID- 10719177 TI - Characterization of isoperoxidase-B2 inactivation in etiolated Lupinus albus hypocotyls. AB - One basic peroxidase isoenzyme, with a pI of 8.8, is present in the intercellular washing fluid in the aerial part of 6-day-old Lupinus albus hypocotyl seedlings. This isoenzyme, called LuP-B2, is the principal soluble component secreted into the apoplastic space and it is a constitutive enzyme along the whole length of etiolated hypocotyl. The enzymatic inactivation process which this apoplastic peroxidase undergoes is described for the first time. The kinetic constants which describe its inactivation by H(2)O(2) in the absence of reductant substrates are determined. LuP-B2 is inactivated in situ and in vitro in a time- and concentration-dependent manner. H(2)O(2) acts as a suicide substrate according to a model previously proposed by us. The constant values calculated are similar to those calculated for the basic isoenzyme of horseradish roots, HRP-C. LuP-B2 presents a k(inact) value of 7.5 x 10(-3) s(-1) and a k(cat) of 6.7 s(-1). This isoenzyme makes 889 catalytic cycles for each inactivation event. The similarity in behavior and the constant values, together with other situations (both are excreted, soluble and constitutive isoenzymes) suggest that the inactivation process could play an important role in plant development and stress situations. PMID- 10719178 TI - Stratum corneum protein mobility as evaluated by a spin label maleimide derivative. AB - The molecular dynamics in the vicinity of sulfhydryl groups of stratum corneum (SC) proteins has been studied by electron paramagnetic resonance (EPR) spectroscopy of maleimide spin labels covalently bound to the proteins. The total amount of bound maleimide was around 4 nmol per mg of SC. We have interpreted the coexistence of two spectral components in the EPR spectra by a two-state model with a fraction of label hydrogen bonded to proteins and another fraction exposed to the aqueous environment. We showed that the relative populations among these two states, determined by spectral simulation, are in thermodynamic equilibrium. The calculated energetic gain for the nitroxide to form hydrogen bond with SC proteins rather than to be dissolved in the buffer was approximately 12 kcal/mol in the temperature range of 2-30 degrees C and approximately 5 kcal/mol in the range of 30-86 degrees C. Temperature profiles of other EPR parameters related to the rotational diffusion of the probe also showed changes in the temperature interval of 26-42 degrees C, suggesting alterations in the vibration modes of SC proteins which are sensitive to higher motional freedom above 26-42 degrees C. We also compared samples of intact and lipid-depleted SC and we found that the delipidization process does not alter significantly the backbone mobility in the SH group regions, but the data suggest that the protein cavity is more open in the case of the delipidized samples. These results contribute to the understanding of the protein participation in the barrier function of SC, and can be useful to improve the spectral analysis of site-directed spin labeling, particularly for a more quantitative description of the dynamic modes of the nitroxide side chains. PMID- 10719179 TI - Identification of oxidized protein hydrolase of human erythrocytes as acylpeptide hydrolase. AB - Partial amino acid sequence of 80 kDa oxidized protein hydrolase (OPH), a serine protease present in human erythrocyte cytosol (Fujino et al., J. Biochem. 124 (1998) 1077-1085) that is adherent to oxidized erythrocyte membranes and preferentially degrades oxidatively damaged proteins (Beppu et al., Biochim. Biophys. Acta 1196 (1994) 81-87; Fujino et al., Biochim. Biophys. Acta 1374 (1998) 47-55) was determined. The N-terminal amino acid of diisopropyl fluorophosphate (DFP)-labeled OPH was suggested to be masked. Six peptide fragments of OPH obtained by digestion of DFP-labeled OPH with lysyl endopeptidase were isolated by use of reverse-phase high-performance liquid chromatography, and the sequence of more than eight amino acids from the N terminal position of each peptide was determined. Results of homology search of amino acid sequence of each peptide strongly suggested that the protein was identical with human liver acylpeptide hydrolase (ACPH). OPH showed ACPH activity when N-acetyl-L-alanine p-nitroanilide and N-acetylmethionyl L-alanine were used as substrates. Glutathione S-transferase (GST)-tagged recombinant ACPH (rACPH) was prepared by use of baculovirus expression system as a 107-kDa protein from cDNA of human erythroleukemic cell line K-562. rACPH reacted with anti-OPH antiserum from rabbit. rACPH showed OPH activity when hydrogen peroxide-oxidized or glycated bovine serum albumin was used as substrates. As well as the enzyme activities of OPH, those of rACPH were inhibited by DFP. The results clearly demonstrate that ACPH, whose physiological function has not yet been well characterized, can play an important role as OPH in destroying oxidatively damaged proteins in living cells. PMID- 10719180 TI - Mechanisms for the enhanced thermal stability of a mutant of transcription factor 1 as explained by (1)H, (15)N and (13)C NMR chemical shifts and secondary structure analysis. AB - A variant of the bacteriophage SPO1-encoded transcription factor 1 (TF1) with two site-specific mutations (E15G and T32I) was shown to be more thermally stable and bind DNA more tightly compared to the wild-type protein. In order to understand the biochemical mechanisms underlying these properties, we are engaged in determining the solution structures of this mutant alone and in complex with DNA using nuclear magnetic resonance (NMR) spectroscopy. The first phase of this project is reported here, as we have completed most of the backbone and sidechain sequential NMR assignments of the mutant protein, TF1-G15/I32. Insights derived from the (1)H, (15)N and (13)C chemical shifts and from the secondary structure analysis provide us with an explanation for the noted increase in thermal stability of TF1-G15/I32. Compared to the structure of the wild-type protein, the beta-sheet and the C-terminal helix remain largely unaffected whereas the mutations cause great changes in the first two helices and their enclosed loop. Specifically, we have found that the second helix is extended by one residue at its N-terminus and rotated in a way that allows Ala-37 to interact with Tyr-94 of the C-terminal helix. The loop has been found to become more rigid as a result of hydrophobic interactions between the flanking second and first helices and also between the second helix and the loop itself. Furthermore, the T32I mutation allows tighter packing between the second helix and the beta-sheet. Collectively, these changes contribute to a more tightly associated dimer and hence, to a greater thermal stability. PMID- 10719181 TI - Mutant residues suppressing rho(0)-lethality in Kluyveromyces lactis occur at contact sites between subunits of F(1)-ATPase. AB - Characterisation of 35 Kluyveromyces lactis strains lacking mitochondrial DNA has shown that mutations suppressing rho(0)-lethality are limited to the ATP1, 2 and 3 genes coding for the alpha-, beta- and gamma- subunits of mitochondrial F(1) ATPase. All atp mutations reduce growth on glucose and three alleles, atp1-2, 1-3 and atp3-1, produce a respiratory deficient phenotype that indicates a drop in efficiency of the F(1)F(0)-ATP synthase complex. ATPase activity is needed for suppression as a double mutant containing an atp allele, together with a mutation abolishing catalytic activity, does not suppress rho(0)-lethality. Positioning of the seven amino acids subject to mutation on the bovine F(1)-ATPase structure shows that two residues are found in a membrane proximal region while five amino acids occur at a region suggested to be a molecular bearing. The intriguing juxtaposition of mutable amino acids to other residues subject to change suggests that mutations affect subunit interactions and alter the properties of F(1) in a manner yet to be determined. An explanation for suppressor activity of atp mutations is discussed in the context of a possible role for F(1)-ATPase in the maintenance of mitochondrial inner membrane potential. PMID- 10719182 TI - Proteolytic cleavage of actin within the DNase-I-binding loop changes the conformation of F-actin and its sensitivity to myosin binding. AB - Effects of subtilisin cleavage of actin between residues 47 and 48 on the conformation of F-actin and on its changes occurring upon binding of myosin subfragment-1 (S1) were investigated by measuring polarized fluorescence from rhodamine-phalloidin- or 1, 5-IAEDANS-labeled actin filaments reconstructed from intact or subtilisin-cleaved actin in myosin-free muscle fibers (ghost fibers). In separate experiments, polarized fluorescence from 1, 5-IAEDANS-labeled S1 bound to non-labeled actin filaments in ghost fibers was measured. The measurements revealed differences between the filaments of cleaved and intact actin in the orientation of rhodamine probe on the rhodamine-phalloidin-labeled filaments, orientation and mobility of the C-terminus of actin, filament flexibility, and orientation and mobility of the myosin heads bound to F-actin. The changes in the filament flexibility and orientation of the actin-bound fluorophores produced by S1 binding to actin in the absence of ATP were substantially diminished by subtilisin cleavage of actin. The results suggest that loop 38-52 plays an important role, not only in maintaining the F-actin structure, but also in the conformational transitions in actin accompanying the strong binding of the myosin heads that may be essential for the generation of force and movement during actin-myosin interaction. PMID- 10719183 TI - Isolation and cDNA-derived amino acid sequences of hemoglobin and myoglobin from the deep-sea clam Calyptogena kaikoi. AB - The heterodont clam Calyptogena kaikoi, living in the cold-seep area at a depth of 3761 m of the Nankai Trough, Japan, has abundant hemoglobins and myoglobins in erythrocytes and adductor muscle, respectively. Two types of hemoglobins (Hb I and Hb II) were isolated, and the complete amino acid sequences of Hb I (145 residues) and Hb II (137 residues) were obtained with combination of cDNA and protein sequencing. The amino acid sequences of C. kaikoi Hbs I and II differed from homologous chains of the congeneric clam Calyptogena soyoae in eight and five positions, respectively. The distal (E7) His, one of the functionally important residues in hemoglobin and myoglobin, was replaced by Gln in hemoglobins of C. kaikoi. A phylogenetic analysis of clam hemoglobins indicates that the evolutionary rate of Calyptogena hemoglobins is rather faster than those of other clams, suggesting that the mutation rate might be accelerated in the deep-sea animals around the areas of cold seeps or hydrothermal vents. On the other hand, it was found unexpectedly that two myoglobins Mbs I and II, isolated from the red adductor muscle, are identical in amino acid sequence Hbs I and II, respectively. Thus it was assumed that genes for Hbs I and II are also expressed in the muscle of C. kaikoi in substitution for myoglobin gene. This suggests that the major physiological role of globins in C. kaikoi is storage of oxygen under the low oxygen conditions, rather than circulating of oxygen. PMID- 10719184 TI - Sequence and expression of seven new tetraspans. AB - The tetraspans are components of large molecular complexes that include also non tetraspan molecules, in particular integrins. We have identified and sequenced several new members of the tetraspan superfamily, called NET-1 to NET-7 (new EST tetraspan). Sequence analysis of the NET reveals a structure typical for tetraspans, with the presence of four transmembrane domains delimiting two extracellular regions as well as conserved amino acid residues. The NET are differentially expressed in human cell lines. PMID- 10719185 TI - Cattle use visual cues to track food locations. AB - We tested the hypothesis that cattle aided by visual cues would be more efficient than uncued animals in locating and consuming foods placed in either fixed or variable locations within a 0.64-ha experimental pasture. Eight yearling steers were randomly selected and trained to associate traffic barricades and traffic cones with high- (oat-barley mixture) and low- (straw) quality foods, respectively. Initially steers were randomly assigned to 1 of 4 food location/visual cue treatments: fixed locations/with cues (F/C), variable locations/with cues (V/C), fixed locations/no cues (F/NC), or variable locations/no cues (V/NC). High- and low-quality foods and their respective cue (or no cue) were placed in the experimental pasture. Individual animals were allowed to explore the pasture for 10 min twice per day every other day for 1 week. Minutes until feeding, first feed type consumed (i.e., high-quality, low quality, or no food consumed), animal location and activity (i.e., feeding, standing, or moving), and total intake of high- and low-quality feed were recorded during each 10-min trial. At the end of each week, location/visual cue treatments were randomly assigned to another 2 steers, which permitted an independent test of each animal in each treatment over a 4-week period. Animals in the F/C and V/C treatments took about 2 min to initially locate and consume a food, compared to F/NC and V/NC animals who took nearly 4 and 6 min, respectively. The high-quality food was the first food located and consumed by F/C, V/C, F/NC, and V/NC animals during 79, 77, 67, and 54% of sampling occasions, respectively. Cued animals typically spent more time feeding (P=0.0004) and less time standing (P=0.005) than uncued animals. Cued animals had a higher mean intake than uncued animals of high- (P=0.001) and low- (P=0.04) quality food. Mean high-quality intake for F/C, V/C, F/NC, and V/NC treatments was 266, 245, 214, and 126 (+/-22) g, respectively; mean low-quality intake for the same treatments was 36, 32, 12, and 10 (+/-10) g. Cued animals also consumed more food per distance traveled than uncued animals (P=0.005). Animals located food quicker (P=0.03) and consumed more high-quality food (P=0.02) when food locations were constant than when they were variable. Our data strongly indicate that cattle can learn to associate visual cues with disparate food qualities and use this information to forage more efficiently in both fixed and variable foraging environments. PMID- 10719186 TI - Vocal behaviour in cattle: the animal's commentary on its biological processes and welfare. AB - The vocalizations of cattle provide conspecifics with meaningful information about the caller. If we can learn how to interpret this information correctly, it could be used to improve management and welfare assessment. Vocalization may be viewed as a subjective commentary, by an individual, on its own internal state. The vocal behaviour of cattle is potentially a useful indicator of their physiological and psychological functioning.In the first part of this article we ask what information is exchanged using auditory cues. Vocalizations provide information on the age, sex, dominance status and reproductive status of the caller. Calves can recognize their mothers using vocal cues but it is not clear whether cows recognize their offspring in this way. Vocal behaviour may play a role in estrus advertisement and competitive display by bulls. Under experimental conditions involving pain or social isolation, vocal response is useful as an indicator of welfare, if properly used. Unlike commonly used physiological measures, it can be recorded non-invasively and varies on a number of quantitative and qualitative dimensions.In the second part we review methodological approaches to the study of vocal behaviour and their application in cattle welfare research. Methods may focus on the actions of the vocalizing animal and the conditions which elicit vocal behaviour, the response of an animal to hearing another's vocalizations, or interactions between sender and receiver.We argue that vocal behaviour in cattle may be valuable in welfare studies if the endogenous, exogenous and developmental factors influencing its expression can be more thoroughly investigated and understood. PMID- 10719187 TI - Play behaviour in group-housed dairy calves, the effect of space allowance. AB - In dairy calves kept in pens, lack of sufficient space may inhibit the performance of play behaviour. The present study investigated, firstly, if an increase in space allowance increases the occurrence of play behaviour, and secondly, if calves kept at a low space allowance perform more locomotor play when released individually in a large novel area. A total of 96 dairy calves in six repetitions were housed in groups of four, in pens of either 4, 3, 2.2 or 1.5 m(2) per calf from 2 weeks of age. The occurrence of play behaviour in the home environment was recorded continuously for each individual calf during 24 h at 5, 7 and 9 weeks of age. Locomotor play decreased over the weeks (54, 29 and 19 s for weeks 5, 7 and 9, respectively; F(2,40)=17.98; P<0.001), and the interaction between space allowance and week tended to be significant (F(6,40)=1.96; P<0.10). At 5 weeks of age, calves kept at 4 or 3 m(2) per calf performed more locomotor play in the home environment than calves at 2.2 or 1.5 m(2) per calf (68, 74, 38 and 39 s for 4, 3, 2.2 and 1.5 m(2) per calf, respectively; F(3,15)=3.40; P<0.05), but in weeks 7 and 9, no effects of space allowance were found. In addition, the duration of locomotor play was recorded for all calves during an individual 10-min open-field test in a 9.6x4.8 m arena at 4 and 10 weeks of age. During the open-field test at 10 weeks of age, calves from pens with 1.5 m(2) per calf performed more locomotor play than calves on the remaining treatments (10, 9, 12 and 25 s for 4, 3, 2.2 and 1.5 m(2) per calf, respectively; F(3,15)=4.05; P<0.05). The present study shows that an increase in the available space increases the occurrence of locomotor play in the home environment at 5 weeks of age. It also shows that calves kept in pens with the smallest space allowance performed more locomotor play behaviour when released in a large arena at 10 weeks of age. PMID- 10719188 TI - Effect of age of calf on suckling behaviour and other behavioural activities of Zebu and crossbred calves during restricted suckling periods. AB - The objective of this study was to investigate the effect of age of calf on the behaviour of Zebu and crossbred calves during restricted suckling (RS) periods. The behaviours of 20 Zebu and 16 crossbred calves were recorded during two 30-min sessions each day after milking when the calves and their dams were brought together in a group for suckling. This was made for a time period of 2 weeks/month for 6 months postpartum.The total suckling duration was significantly longer in Zebu calves (11.8+/-0.19 min) compared to the crossbred calves (9.4+/ 0.19 min), but decreased significantly in both breeds with increasing age from 1 to 6 months. The number of suckling bouts decreased from a mean of 3.8 at 1 month of age to 1.1 at 6 month (P<0.05). The duration of each suckling bout decreased significantly from a mean of 3.5+/-0.15 min at 1 month of age to 1.6+/-0.01 min at 6 months (<0.05). The frequency of crossbred calves cross-suckling (3.7%) was significantly higher than that of the Zebu calves (1.9%; P<0.05). The frequency of calves cross-suckling decreased significantly from 4.2% at 1 month of age to 2.3% at 6 months. The duration and number of bouts of cross-suckling was significantly higher in the crossbred calves (duration 0.9+/-0.06 min; bouts 3, 7) than in the Zebu calves (duration 0.5+/-0.06 min; bouts 2, 7) and decreased with increasing age of calf. The duration and number of bouts of inter-sucking was significantly higher in the crossbred calves (duration 0.6+/-0.07 min; bouts 1, 6) than in the Zebu calves (duration 0.1+/-0.04 min; bouts 0, 5) and decreased with increasing age of calf. Exploration increased in duration as the calves increased in age from 1 to 6 months (P<0.05). The duration of play increased significantly with the increase in age of calf from 1 to 6 months, and occurred mainly after nursing. PMID- 10719189 TI - Floor temperature preference of sows at farrowing. AB - A preference testing apparatus was used to provide sows with continuous access to three identical farrowing crates, each with a different floor temperature. The concrete floor under each crate contained copper pipe through which temperature controlled water was circulated to achieve unoccupied floor temperatures of 22 degrees C (+/-3.5), 29 degrees C (+/-1) and 35 degrees C (+/-1). Eighteen sows were tested in the apparatus. Video recording was used to determine sow position from 7 days before farrowing (Days -7 to -1) to 14 days after (Days 1 to 14). On Days -7 to -1, sows showed no significant preference among the three temperatures when selecting a resting area. Once farrowing had begun, there was a significant increase (P<0.01) in the use of the 35 degrees C floor and it became the most preferred resting area for Days 1 to 3. After this interval, use of the 35 degrees C floor declined significantly (P<0.01), and use of the cooler floors increased, resulting in no significant thermal preference during Days 4 to 6. There was a further decline in the use of the 35 degrees C floor after Days 4 to 6 (P<0.01) to the extent that the coolest floor (22 degrees C) became the most preferred from Days 7 to 14. In summary, sows showed a pronounced increase in preference for a warm floor during the 3 days after the start of farrowing. This change in preference may explain how free-living sows select a suitable thermal environment for their young, and why sows try to avoid metal flooring at the time of farrowing. PMID- 10719190 TI - The aversiveness of carbon dioxide stunning in pigs and a comparison of the CO(2) stunner crate vs. the V-restrainer. AB - Using aversion learning techniques, the relative aversiveness of CO(2) to pigs in comparison to a shock with an electric prodder, and the aversiveness of a CO(2) stunner crate in comparison to the aversiveness of a V-belt restrainer used for electric stunning were examined. The results showed that 90% CO(2) was considerably less aversive than an electric shock with a prodder. However, during exposure to 90% CO(2) all pigs lost conscious, which may have affected their memory of the procedure. The pigs remained conscious after exposure to 60% CO(2) and again showed virtually no aversion towards the stunner crate, while an electric shock with a prodder appeared highly aversive. The aversion to the V restrainer belt and the CO(2) crate were similar. PMID- 10719191 TI - Behavioural interactions between West African dwarf nanny goats and their single born kids during the first 48 hours post-partum. AB - West African dwarf nanny goats and their single-born kids were tested to find out their behavioural response to separation and their mutual recognition during the first 48 h post-partum. The tests were conducted at 18, 24, 36 or 48 h post partum. Chi-square analyses were used to determine whether age, sex and birth weight of kids as well as h partum and parity of dams had an effect on post partum bahaviours.In a preliminary study where the kids were not prevented from sucking prior to the tests, the majority of kids (93%) and does (86%) exhibited apparant minimal concern to separation (i.e., had scores of less than 3) and also failed to seek each other. When the kids were prevented from sucking 2 h prior to the tests, the couple showed some response to separation and made attempts to seek each other. This suggests that in goats the state of the neonate's nourishment and the fullness of the dam's udder may be important factors that determine the willingness of the kid and the dam to seek each other when separated. The behavioural response of 48-h-old kids to separation from their dams when they had not sucked for 2 h was significantly higher (P<0.05) than that of 18-h-old kids. The sex and birth-weight of 48-h-old kids did not affect their response to separation from their dams. Hours post-partum and parity of does did not affect their response to separation from their kids even though the proportion of does exhibiting the highest response was much higher in primiparous then multiparous animals (44% vs. 13%). Recognition of dams by kids was poor at 18 and 24 h with more than half (71%) the kids failing to recognize their dams. Dam recognition ability improved with age and at 48 h the majority of kids (88%) were able to recognize their dams. Does tested at 48 h post partum had better (P0.05) kid recognition than those tested at 18 and 24 h. The sex and birth weight of kids and the parity of dams had no effect on the mutual recognition of kids and their dams at 48 h post-partum. PMID- 10719192 TI - Mating behavior of bucks and does in goat operations under range conditions. AB - The objectives of this study were to characterize the mating capacity of bucks under range condition and assess the effect of mating frequency on flock fertility. Two adjacent flocks of crossbred goats (Criolloxdairy breeds; n=70 does, 5 bucks and 141 does, 4 bucks) were used in this study. The mating period was 16 and 20 days for each flock, respectively, in January and February, 1996. Mating activity of bucks and does was recorded day and night during the first 11 days of the breeding period. The combined data of both flocks showed a strong relationship between number of goats in estrus and the average services of bucks (r=0.95; P<0.01). With an excess of estrous does, bucks on average copulated 9.1 times daily during the first 11 days of the mating period. Considering both flocks combined, does copulated on average 4+/-1.8 times through the estrus period, and they did so with an average of 2.2+/-1.3 different bucks. Sexual activity of bucks was greatest from sunset to midday. Number of services from different bucks did not affect pregnancy rate (81.4% for goats serviced by 1-2 bucks and 77.8% for goats serviced by 3-4 different bucks; chi(2)=0.25; P7 services, respectively; chi(2)=0.67; P0.05). Mean+/-S.D. daily liveweight loss of bucks during the entire mating period was 547+/-197 g. It was concluded that, with buck percentages higher than 3%, the number of ejaculations of bucks is linearly and positively related to number of females in estrus. Also, these findings indicate that neither number of copulations nor number of services from different bucks affected kidding rates. PMID- 10719193 TI - Responses of adult laying hens to abstract video images presented repeatedly outside the home cage. AB - Previous studies reported that domestic chicks showed progressively greater attraction towards biologically neutral video images (screensavers) with repeated exposure [Jones, R.B., Carmichael, N., Williams, C., 1998. Social housing and domestic chicks' responses to symbolic video images. Appl. Anim. Behav. Sci. 56, 231-243; Jones, R.B., Larkins, C., Hughes, B.O., 1996. Approach/avoidance responses of domestic chicks to familiar and unfamiliar video images of biologically neutral stimuli. Appl. Anim. Behav. Sci. 48, 81-98]. The potential existence of an adult parallel was examined here by studying the responses of laying hens to similar screensaver videos when these were presented repeatedly in front of their home cage. In Experiment 1, individually housed, 31-week-old laying hens were exposed to either the video image of a computer screensaver (SS) programme (Fish), a blank but illuminated television monitor (B), or a black plastic hide (H) presented approximately 50 cm in front of their home cages for 10 min/day on each of 5 consecutive days. The bird's position in the cage and the orientation of its head were recorded every 15 s during the-10 min exposure period in order to assess approach and interest, respectively. Interest was scored by summing the numbers of observations at which the hen was either facing the front or had its head out of the cage. Birds in the B and H treatment groups showed few deviations from neutrality in their approach or interest. Conversely, although SS birds avoided the video stimulus on the first day their responses had reached neutrality (neither approach nor avoidance) by the third day and they showed significantly more approach than would be expected by chance at the fifth presentation. They also showed significantly more interest than chance on each test day; this score increased progressively and showed no sign of waning even at the fifth presentation. To determine whether or not such interest would be maintained indefinitely, the responses of previously untested hens were examined when the same video (Fish) was presented for 10 min/day on each of 20 consecutive days (Experiment 2). A blank, lit television (B) was again used a control. An unfamiliar video (Doodles) was presented to the SS birds on day 21 to determine the effects of stimulus change. After avoiding the stimuli upon their first presentation, both SS and B birds achieved neutrality by day 3. Approach scores then fell in B birds but rarely deviated from neutrality in SS ones. The SS video attracted markedly more interest than did the blank screen. On this occasion, SS hens showed significantly greater interest than would be expected by chance as early as the third presentation and this was still evident upon the eighth presentation; thereafter it waned gradually. However, interest was reinstated fully when the unfamiliar SS image was shown on day 21. The present findings clearly demonstrate that abstract video images, presented in front of the home cage for 10 min on consecutive days, reliably attracted and sustained the interest of individually housed laying hens for as long as 8 days. These results are consistent with those obtained when chicks were repeatedly exposed to similar screensaver videos, i.e., this phenomenon is not dependent on the stage of development. Our results also confirm the importance of considering the environment outside as well as inside the cage in future environmental enrichment programmes. PMID- 10719194 TI - The behaviour of broiler chickens and its modification by lameness. AB - The behaviour of six replicates of broilers obtained from commercial farms, fed ad-libitum and housed on 23-h light:1-h dark schedule at 20 lx was observed using scan sampling. Comparisons were made between sound birds and those of varying degrees of lameness between 39 and 49 days of age. Sound broilers averaged 76% of their time lying and this increased significantly to 86% in lame birds (gait score 3). Lying also increased with age. Although sound broilers spent only a minor part of the day on their feet, they spent significantly more time standing idle (7%), standing preening (3.5%) and standing eating (4.7%) than lame birds. Walking declined with age, but occupied an average 3.3% of the time of a slaughter-weight broiler. Again, lameness significantly reduced this to a minimal 1.5% in the worst affected birds. Sound birds predominantly chose the usual standing posture for eating, whereas, lame birds lay down to eat for almost half their feeding time. Detailed observations using video records revealed that lameness altered the feeding strategy of broilers. Whereas sound birds fed over 50 times in 24 h, the number of visits to the feeder was reduced with increasing lameness to an average of around 30 in the lamest broilers. However, meal duration was adjusted to give no overall differences in time spent feeding per day. Time spent drinking was also the same for all birds, averaging 3% of the day. The alterations of the time budget, in particular the reductions in activities performed whilst standing, and the different feeding strategies adopted, are consistent with lameness imposing a cost on the affected broilers to the detriment of their welfare. PMID- 10719195 TI - Preference for various nest box designs in farmed silver foxes (Vulpes vulpes) and blue foxes (Alopex lagopus). AB - Nest box choice experiments were carried out outside the breeding season on adult silver and blue fox vixens with no previous permanent nest box experience. Nest boxes were varied in height of placement, number of rooms, presence of entrance room or platform and light conditions. Only one parameter was varied in any one experiment. Both fox species clearly preferred an elevated multi-room nest box; while silver foxes showed preference for boxes supplied with a platform, blue foxes preferred boxes with an entrance room. There was no significant box preference with respect to light conditions. The possible welfare implications of the preferences are discussed. PMID- 10719196 TI - Seasonal foraging behavioural compensation in reproductive wapiti hinds (Cervus elaphus canadensis). AB - Captive wapiti hinds were observed during seven periods between June 1996 and November 1997. We investigated their activity and foraging behaviour on two pastures, heavily and lightly grazed, during four phases of the reproductive cycle (early gestation, late gestation, peak lactation, and late lactation). Within season, differences in forage availability between pastures had little apparent effect on wapiti feeding behaviour (p0.05). However, within each pasture, hinds tended to select locations of higher phytomass than the pasture average. Among seasons, effects of forage availability on feeding behaviour were pronounced. Hinds grazed longest in late lactation (12.8 h/day), when they spent 94% of their active time foraging, whereas during early gestation they foraged fewer hours (8.2 h/day) and less intensively (66% of active time). The longest foraging bouts also occurred in late lactation (100 min) and decreased linearly as the number of bouts per day increased (R(2)=0.88). The annual peak bite rate (BR) was in late lactation (62 bites/min), whereas the annual nadir occurred in early gestation (37 bites/min). Smallest bite sizes (BS) (mg) were recorded in late gestation (127 mg), and increased linearly with forage availability (R(2)=0.46), with largest BS occurring during peak lactation (280 mg). This study demonstrated how seasonal modifications in activity and foraging behaviour enabled gestating and lactating wapiti hinds to satisfy their changing nutritional requirements on seasonal pastures. Knowledge of behavioural compensation in response to nutritional demand and pasture conditions will be useful in designing supplementation program for farmed wapiti. PMID- 10719198 TI - Erratum to "Individual behavioural characteristics in pigs and their impact on production" PMID- 10719197 TI - Effects of management at weaning on behaviour and weight gain of farmed red deer calves. AB - Two studies were performed to determine whether there were behavioural or productive differences arising from two contrasting weaning practices for red deer; proximate vs. distant separation of dams and calves. In Experiment 1, 80 calves across two replicates were used. For each replicate, calves were separated from their mothers, weighed and allocated to one of two treatments (n=20 calves), either confinement in an unfamiliar paddock 100 m from their mothers for 2 weeks following weaning (treatment N), or transportation 2 km to a different farm (treatment F). Groups were observed during the following 6 days and weighed 14 days after weaning. In Experiment 2, the same procedure was performed out on two commercial farms, but using 40 calves per treatment group, without replication or weight recording. In Experiment 1 running, fenceline pacing and vocalising declined following weaning, with steeper declines for F than N calves for running and vocalising (P<0.01). Similar trends, with an indication of less vocalising and movement overall, were seen in Experiment 2. In contrast, mean post-weaning weight gains for N calves were higher than for F calves (3.0 compared with 2.4 kg, SED 0.48 kg; P<0.05). Weather variables (cloud, temperature and wind) were associated with behaviour in both studies (P<0.05), with a trend for pacing, calling and running to increase as conditions deteriorated (cloud cover and wind speed increased, and temperature decreased). It was concluded that distant separation appeared beneficial to the calves but more research was required to determine optimal weaning management. The study supported previous evidence that weaning should be carried out in good weather. PMID- 10719199 TI - Interspecific combative interactions between wood-decaying basidiomycetes. AB - Competition is the most common type of interaction occurring between wood decaying higher fungi. Since competition for nutrients in organic resources is effectively brought about by competition for space, the common division into interference and exploitation competition is not very appropriate. Fungal competition can be divided into primary resource capture (obtaining uncolonized resources) and secondary resource capture (combat to obtain resources already colonized by other fungi). Combative mechanisms include antagonism at a distance, hyphal interference, mycoparasitism and gross mycelial contact. Interactions can result in deadlock or replacement, and a hierarchy of combative ability can be discerned amongst fungi that inhabit particular resources, but within this hierarchy there exists intransitivity, modification of outcome by other species and abiotic variables. Interactions can dramatically alter mycelial function, and have potential as biological control agents of fungal pathogens of trees and in service timber. PMID- 10719200 TI - Three types of phenol and p-cresol catabolism in phenol- and p-cresol-degrading bacteria isolated from river water continuously polluted with phenolic compounds. AB - A total of 39 phenol- and p-cresol-degraders isolated from the river water continuously polluted with phenolic compounds of oil shale leachate were studied. Species identification by BIOLOG GN analysis revealed 21 strains of Pseudomonas fluorescens (4, 8 and 9 of biotypes A, C and G, respectively), 12 of Pseudomonas mendocina, four of Pseudomonas putida biotype A1, one of Pseudomonas corrugata and one of Acinetobacter genospecies 15. Computer-assisted analysis of rep-PCR fingerprints clustered the strains into groups with good concordance with the BIOLOG GN data. Three main catabolic types of degradation of phenol and p-cresol were revealed. Type I, or meta-meta type (15 strains), was characterized by meta cleavage of catechol by catechol 2,3-dioxygenase (C23O) during the growth on phenol and p-cresol. These strains carried C23O genes which gave PCR products with specific xylE-gene primers. Type II, or ortho-ortho type (13 strains), was characterized by the degradation of phenol through ortho fission of catechol by catechol 1,2-dioxygenase (C12O) and p-cresol via ortho cleavage of protocatechuic acid by protocatechuate 3,4-dioxygenase (PC34O). These strains carried phenol monooxygenase gene which gave PCR products with pheA-gene primers. Type III, or meta-ortho type (11 strains), was characterized by the degradation of phenol by C23O and p-cresol via the protocatechuate ortho pathway by the induction of PC34O and this carried C23O genes which gave PCR products with C23O-gene primers, but not with specific xylE-gene primers. In type III strains phenol also induced the p-cresol protocatechuate pathway, as revealed by the induction of p-cresol methylhydroxylase. These results demonstrate multiplicity of catabolic types of degradation of phenol and p-cresol and the existence of characteristic assemblages of species and specific genotypes among the strains isolated from the polluted river water. PMID- 10719201 TI - Molecular analysis of ammonia-oxidising bacteria in soil of successional grasslands of the Drentsche A (The Netherlands). AB - Changes in the community structure of chemolitho-autotrophic ammonia-oxidising bacteria of the beta-subgroup Proteobacteria were monitored during nutrient impoverishment management of slightly acidic, peaty grassland soils, which decreased in pH with succession. Specific PCR, cloning and sequence analysis, denaturing gradient gel electrophoresis (DGGE) and probe hybridisation were used to analyse rDNA sequences directly recovered from successional soils. Four previously characterised ammonia oxidiser sequence clusters were recovered from each soil, three associated with the genus Nitrosospira and one with the genus Nitrosomonas. All samples were dominated by Nitrosospira-like sequences. Nitrosospira cluster 3 was the most commonly recovered ammonia oxidiser group in all fields, but a greater representation of Nitrosospira clusters 2 and 4 was observed in older fields. Most probable number (MPN) counts were conducted using neutral and slightly acid conditions. Neutral pH (7.5) MPNs suggested a decrease in ammonia oxidiser numbers in later successional fields, but this trend was not observed using slightly acid (pH 5.8) conditions. Analysis of terminal MPN dilutions revealed a distribution of sequence clusters similar to direct soil DNA extractions. However, an increased relative recovery of Nitrosospira cluster 2 was observed for acid pH MPNs compared to neutral pH MPNs from the most acidic soil tested, in agreement with current hypotheses on the relative acid tolerance of this group. PMID- 10719202 TI - Francisella tularensis does not manifest virulence in viable but non-culturable state. AB - Francisella tularensis is a small Gram-negative bacterium that causes tularemia in animals and man. The disease can be transmitted by handling of infected animals, by contaminated dust, by insect vectors, or by drinking contaminated water. In the present study cells of F. tularensis were subjected to extended storage in cold water devoid of carbon sources. Total cell counts remained constant throughout a 70-day period and beyond, while plate counts decreased to an undetectable level after 70 days. Attempts to resuscitate the cells were unsuccessful. Quantitative PCR targeting the 16S rDNA of F. tularensis showed an increase in variability after 25 days and the signal was lost after 45 days. Metabolic activity, measured by accumulation of rhodamine 123, declined to approximately 35% after a 140-day period. Analyses of substrate responsiveness of cells stored for 140 days in cold water showed that approximately 30% of the population increased in size after incubation in rich medium in the presence of nalidixic acid. Approximately 10(5) of these cells were injected intraperitoneally into mice. No signs or symptoms of tularemia were observed during 3 weeks. In addition, there was no evidence of stimulation of lymphocytes with F. tularensis as recall antigen. In conclusion, viable but non-culturable cells of F. tularensis are avirulent in mice, giving new insight into the ecological niche of this bacterium. PMID- 10719203 TI - Growth kinetics of thermophilic Methanosarcina spp. isolated from full-scale biogas plants treating animal manures. AB - This study determines the growth kinetics of thermophilic strains of Methanosarcina spp. from full-scale thermophilic biogas plants. The complete set of kinetic parameters, including maximum specific growth rate u(max), half saturation constant K(S), acetate threshold concentration and cell growth yield Y(X/S), were determined for six Methanosarcina strains newly isolated from full scale reactors and the type strain Methanosarcina thermophila TM-1(T). The kinetic experiments were performed in media supplemented with acetate and activated carbon at the optimum growth temperatures of the individual strains, 50 55 degrees C. The u(max) values of the isolates were in the range of 0.044-0.064 h(-1), the K(S) ranged from 6.5 to 24.7 mM acetate and the threshold for acetate utilization from 0.11 to 0.40 mM. The cell growth yields of the strains were between 0.78 and 2.97 g dry weight cells mol(-1) acetate. The six isolates exhibited significantly higher u(max) and had higher affinity to acetate than the type strain M. thermophila TM-1(T). Generally, the affinities of thermophilic Methanosarcina strains tested in this study cover a similar range to those reported in the literature for mesophilic Methanosarcina spp. with acetate as substrate. The strains isolated from plants treating mixtures of animal manures and industrial organic wastes had higher affinity for acetate and lower thresholds than strains isolated from reactors operating solely on manures. PMID- 10719204 TI - Archaeaplankton in the Columbia River, its estuary and the adjacent coastal ocean, USA. AB - PCR-amplified 16S rRNA genes from particle-attached and free-living Archaea in the Columbia River estuary, particle-attached Archaea in the river, and Archaea in the adjacent coastal ocean were cloned, and 43 partial sequences were determined. There was a high diversity of Archaea in the estuary, especially among the particle-attached Archaea, with representatives from four major phylogenetic clusters. Eighteen of 21 estuarine clones were closely related to clones from the river and the coastal ocean or to clusters of marine and soil clones identified in other studies. This contrasts with a similar study of the estuarine bacterial community that found 62% of bacterial 16S rRNA clones to be unique to the estuary. Archaea in the estuary were primarily allochthonous, and therefore, unlike the bacteria, probably do not form a native estuarine community. PMID- 10719205 TI - Effects of yoghurt intake on plasmid transfer and colonisation with transconjugants in the digestive tract of mice associated with human faecal flora. AB - This study deals with the effects of yoghurt intake on wild-type and recombinant plasmid transfer from an exogenous Escherichia coli K12-derivative donor strain to an endogenous recipient strain in the digestive tract of mice associated with human faecal flora. We showed that the self-transmissible plasmid R388 was efficiently transferred to recipient strain PG1 in mice associated with human faecal flora (HFF-PG1) and that the resulting transconjugants (PG1-R388) became established at a high and maximal population level without any selective pressure. Using HFF-PG1 mice made it possible to determine whether yoghurt consumption decreases R388 transfer efficiency and the ability of transconjugant PG1-R388 to successfully colonise the digestive tract. Results indicated that yoghurt consumption had two effects: it reduced the efficacy of plasmid transfer 10-fold and decreased the transconjugant PG1-R388 population density 100-fold, compared to the control group. We also describe another HFF mouse model in which recipient PG1 was replaced by EM0 with which no plasmid transfer was observed. This model made it possible to demonstrate the potential promoting effect of yoghurt intake on transconjugant formation and establishment. Our results indicated no yoghurt effect; no transconjugants appeared in the digestive tract of HFF-EM0 mice fed on yoghurt or on standard food. In both mouse models, HFF-PG1 and HFF-EM0, yoghurt intake did not promote the mobilisation of either the non self-transmissible plasmid pUB2380 or the recombinant plasmid pCE325. PMID- 10719206 TI - Transformation of 1,1-dichloro-2,2-(4-chlorophenyl)ethane (DDD) by Ralstonia eutropha strain A5. AB - Evidence is presented demonstrating the ability of Ralstonia eutropha A5 to degrade 1,1-dichloro-2,2-bis(4-chlorophenyl)ethane (DDD) aerobically. Strain A5 was able to effect significant transformation of [(14)C]DDD: the hexane extractable radioactivity decreased to approximately 50% of the controls while more than 25% of the total radioactivity became associated with the acidified culture supernatant. There was also an increase in the amount of radioactivity associated with the cell pellet when compared to the biotic control. A meta fission pathway for the degradation of DDD is proposed based on the recovery of seven chlorinated metabolites identified by gas chromatography-mass spectrometry analysis. PMID- 10719207 TI - Survival and conjugation of Bacillus thuringiensis in a soil microcosm. AB - The survival and conjugation ability of sporogenic and asporogenic Bacillus thuringiensis strains were investigated in broth, in non-amended sterile clay soil monoculture and in mixed soil culture. The 75 kb pHT73 plasmid carrying an erythromycin resistance determinant and a cry1Ac gene was transferred in mating broth and soil microcosm. Survival of strains was assessed in soil monoculture and in mixed soil culture for up to 20 days. Sporogenic strains rapidly formed viable spores which were maintained until the end of the experiment. The asporogenic strains were no longer recovered after 8 days of incubation. This study shows that the environmental impact of asporogenic B. thuringiensis strains is lower than that of sporogenic B. thuringiensis strains. Thus, the use of asporogenic strains may significantly reduce any potential risk (gene transfer, soil and plant contamination) due to the dissemination of B. thuringiensis-based biopesticides in the environment. PMID- 10719208 TI - Identification of spores in the polycentric anaerobic gut fungi which enhance their ability to survive. AB - Two new isolates of the gut fungi were obtained from the rumen digesta and faeces of a cow. These isolates, designated Anaeromyces following rDNA typing, displayed a polycentric growth habit but differed from all other gut fungi in that they were able to survive in the laboratory for considerable periods without the need for sub-culture. Light microscopy of preparations from old liquid-grown cultures revealed the presence of DNA-containing spores with two or four chambers. A comparative evaluation of the growth produced when fresh media were inoculated with a sample originating from young or old cultures revealed that active growth was delayed with the inoculum from the older culture. We propose that the chambered spores observed in these cultures provide an alternative path in the life cycle of these fungi and may function as a resting stage within the anaerobic environment of the herbivore gut. PMID- 10719209 TI - Distribution of orexin/hypocretin in the rat median eminence and pituitary. AB - We determined the distribution of orexin-A and orexin-B (also known as hypocretin 1 and hypocretin-2) and their receptors in the rat median eminence and pituitary using sensitive radioimmunoassays coupled with high-performance liquid chromatography, immunohistochemistry, and in situ hybridization. Orexin-A and -B were present in the median eminence, adenohypophysis, and neurohypophysis. Orexin fibers were abundant in the median eminence, and a few fibers projected to the neurohypophysis. Both the orexin(1)- and orexin(2)-receptor mRNAs were expressed robustly in the pituitary intermediate lobe, whereas in the anterior lobe, the orexin(1) receptor was more markedly expressed than the orexin(2) receptor. These two receptor mRNAs were also found in the posterior lobe. These findings may implicate orexin's involvement in additional as yet undefined physiological functions in the hypothalamo-hypophyseal tract. PMID- 10719210 TI - Prolonged c-Jun expression in the basolateral amygdala following bulbectomy: possible implications for antidepressant activity and time of onset. AB - Olfactory bulbectomy is a well established animal model of depression. Neurochemical and behavioral alterations observed following olfactory bulbectomy, are due, in part, to the neurodegeneration of specific brain structures. Amygdaloid dysfunction in particular, is known to play a substantial role in the syndrome of the olfactory bulbectomized rat. The present study examined both short- and long-term alterations in immediate early gene expression, tyrosine hydroxylase and serotonin immunoreactivity, and classical silver staining, following olfactory bulbectomy in the basolateral amygdala. The results indicated no consistent change in Fos expression observed over the experimental period. Following bulbectomy, long term (up to 64 days post-lesion) Jun expression, not coincident with silver staining, was observed in the basolateral nucleus. The basolateral nucleus was also intensely immunoreactive for serotonin at this timepoint post-bulbectomy. Thus, following bulbectomy long term alterations in Jun expression occurs in the serotonin rich basolateral amygdala. As a site of action for antidepressant compounds, alterations at the immediate early gene level in this region may have implications both for the model, and antidepressant drug action therein. PMID- 10719211 TI - NGFI-B and nor1 mRNAs are upregulated in brain reward pathways by drugs of abuse: different effects in Fischer and Lewis rats. AB - The two inbred Fischer and Lewis rat strains display differences in acquisition of drug self-administration, suggesting genetic factors controlling the vulnerability to drugs of abuse. In this study, we analyzed the effects of acute and chronic cocaine and morphine on mRNAs encoding the NGFI-B/Nur77 family of nuclear orphan receptors in reward pathways in Fischer and Lewis rats. After a single injection of cocaine, a similar upregulation of NGFI-B mRNA in striatal subregions and cortex cinguli was seen in both Fischer and Lewis rats. In contrast, Nor1 mRNA was only significantly upregulated by cocaine in the Fischer rats. Morphine increased NGFI-B mRNA in medial caudate putamen and cortex cinguli in Lewis rats and Nor1 mRNA in medial caudate putamen in Fischer rats. Chronic cocaine upregulated NGFI-B mRNA in nucleus accumbens core, lateral caudate putamen and cingulate cortex in Fischer rats, whereas no effect was seen in Lewis rats. In contrast, Nor1 mRNA levels were upregulated in Lewis rats in medial caudate putamen and cingulate cortex after chronic cocaine and in cingulate cortex after chronic morphine. No effect on Nor1 mRNA levels was seen in Fischer rats after chronic treatments. Our results demonstrate different responses in addiction-prone Lewis rats as compared to the less addiction-prone Fischer rats with respect to NGFI-B and Nor1 mRNA regulation after acute and repeated administration of cocaine and morphine. Thus, we suggest that the transcription factors NGFI-B and Nor1 might be involved in the control of behaviors such as sensitized locomotor response, craving and aversion that appears after repeated administration of abused drugs. PMID- 10719212 TI - Expression of the GDNF family members and their receptors in the mature rat cochlea. AB - The GDNF family comprises glial cell line-derived neurotrophic factor (GDNF) and the related proteins neurturin, artemin and persephin, which form a subgroup of the TGF-beta superfamily of growth factors. All four neurotrophic factors provide neuronal cell protection and cell survival. GDNF expression was found in the cochlea, and GDNF has been shown to be effective for inner ear protection from drugs and noise-induced insults. As the other members of the GDNF family also provide protective effects on neuronal cells, they may play important roles in the inner ear. We used RT-PCR to examine the expression of GDNF, neurturin, artemin, persephin and their receptors GFRalpha-1, GFRalpha-2, GFRalpha-3 and c ret in whole rat cochlea as well as in functionally different subfractions (modiolus and sensorineural epithelium/lateral wall) and compared the levels of neurotrophin and receptor mRNAs in the cochlea to those in substantia nigra brain region. Our results demonstrate the expression of all GDNF family members and their receptors in cochlea and substantia nigra. However, the relative levels of mRNA were different for several genes tested in subfractions of the cochlea and/or compared to expression levels in substantia nigra. The presence of mRNA for all four members of the GDNF family and their preferred receptors in the rat cochlea suggests potential functional importance of these neurotrophic factors as protection and survival factors in the inner ear. PMID- 10719213 TI - Learning-associated activation of nuclear MAPK, CREB and Elk-1, along with Fos production, in the rat hippocampus after a one-trial avoidance learning: abolition by NMDA receptor blockade. AB - It is widely accepted that the formation of long-term memory (LTM) requires neuronal gene expression, protein synthesis and the remodeling of synaptic contacts. From mollusk to mammals, the cAMP/PKA/CREB signaling pathway has been shown to play a pivotal role in the establishment of LTM. More recently, the MAPK cascade has been also involved in memory processing. Here, we provide evidence for the participation of hippocampal PKA/CREB and MAPK/Elk-1 pathways, via activation of NMDA receptors, in memory formation of a one-trial avoidance learning in rats. Learning of this task is associated with an activation of p44 and p42 MAPKs, CREB and Elk-1, along with an increase in the levels of the catalytic subunit of PKA and Fos protein in nuclear-enriched hippocampal fractions. These changes were blocked by the immediate posttraining intra hippocampal infusion of APV, a selective blocker of glutamate NMDA receptors, which renders the animals amnesic for this task. Moreover, no changes were found in control-shocked animals. Thus, inhibitory avoidance training in the rat is associated with an increase in the protein product of an IEG, c-fos, which occurs concomitantly with the activation of nuclear MAPK, CREB and Elk-1. NMDA receptors appear to be a necessary upstream step for the activation of these intracellular cascades during learning. PMID- 10719214 TI - Influence of subunit configuration on the interaction of picrotoxin-site ligands with recombinant GABA(A) receptors. AB - We have assessed the interaction of picrotoxin and a putative picrotoxin-site ligand [4-dimethyl-3-t-butylcarboxyl-4,5-dihydro (1, 5-a) quinoxaline] (U-93631) with varying configurations of recombinant GABA(A) receptors, using the whole cell patch clamp technique. In alpha2beta2gamma2 GABA(A) receptors, coapplication of picrotoxin with GABA had minimal effects on initial GABA-activated Cl(-) current amplitude, and subsequently enhanced decay of GABA-activated Cl(-) currents. The half-maximal inhibitory concentration (IC(50)) for picrotoxin in alpha2beta2gamma2 receptors was 10.3+/-1.6 microM. The alpha subunit isoform did not affect picrotoxin-induced inhibition, as IC(50) values for alpha3beta2gamma2 (5.1+/-0.7 microM) and alpha6beta2gamma2 receptors (7.2+/-0.4 microM) were comparable to those obtained in alpha2beta2gamma2 receptors. Interestingly, in receptors lacking an alpha subunit (beta2gamma2 configuration), picrotoxin had a markedly lower IC(50) (0.5+/-0.05 microM) compared to alpha-containing receptors. The inhibitory profile was generally similar for the presumed picrotoxin-site ligand U-93631, i.e., IC(50) values were comparable in all alphabetagamma containing receptors, but the IC(50) in beta2gamma2 receptors was greater than 10 fold lower. In addition, a modest but significant initial stimulation of GABA activated current by U-93631 was observed in alpha2beta2gamma2 and beta2gamma2 receptors. A mutation in the second transmembrane domain, shown previously to abolish picrotoxin sensitivity, also greatly attenuated sensitivity to U-93631. Moreover, incubation of receptors with excess U-93631 hindered picrotoxin's ability to gain access to its binding site; both results indicate that U-93631 interacts at the picrotoxin site of the receptor. Our results indicate the presence of an alpha subunit hinders the ability of picrotoxin to block the GABA(A) receptor, and thus provides additional insight into the site of action of picrotoxin. In addition, we have shown that domains important for the actions of picrotoxin also affect U-93631. Thus, this compound should prove to be a useful ligand for analysis of the convulsant site of this receptor. PMID- 10719215 TI - Isolation, characterization and expression of a human brain mitochondrial glutaminase cDNA. AB - Various cDNAs that encode overlapping portions of the full-length human brain glutaminase (GA) cDNA were cloned and sequenced. The overall nucleotide sequence of hGA has a very high degree of identity with that of the rat kidney-type GA cDNA (77.4%) and the known portion of the cDNA that encodes the 5.0-kb porcine GA mRNA (81.1%). The identity is even more remarkable at the amino acid level, particularly in the C-terminal half where the three proteins share a 99.7% sequence identity. The hGA cDNA encodes a 73,427-Da protein that contains an N terminal mitochondrial targeting signal and retains the primary proteolytic cleavage site characterized for the cytosolic precursor of the rat renal mitochondrial glutaminase. The entire coding region was assembled through the use of unique restriction sites and cloned into a baculovirus. Sf9 cells infected with the recombinant virus express high levels of properly processed and active glutaminase. Thus, expression of the isolated hGA cDNA should provide a means to purify large amounts of the mitochondrial glutaminase, a protein that catalyzes a key reaction in the metabolism of glutamine and the synthesis of important excitatory and inhibitory neurotransmitters. PMID- 10719216 TI - mu Opioid receptor: role for the amino terminus as a determinant of ligand binding affinity. AB - The importance of the amino-terminal domain of the mu opioid receptor (MOR) as a component of the high affinity ligand-binding pocket was evaluated. A deletion mutant lacking 64 amino acids from the amino-terminus of MOR (DeltaN64) was constructed and expressed in HEK 293 cells. The affinities of bremazocine and cyclazocine were similar for the truncated and full-length MORs. Affinities of the mu receptor antagonist, naloxone, and the mu receptor agonist, morphine, were decreased 3.5-fold and 6-fold, respectively, for the truncated receptor relative to the wild-type MOR. Similarly, the affinities of the opioid peptide agonists, DAMGO (Tyr-D-Ala-Gly-MePhe-Gly-ol), beta-endorphin, and DADL (Tyr-D-Ala-Gly-Phe-D Leu), for the DeltaN64 receptor were decreased from 3- to 8-fold as a result of the deletion. In contrast, the affinities of the alkaloid agonists, methadone and fentanyl, and the peptide agonists, endomorphin 1 and endomorphin 2, for the truncated receptor relative to MOR were reduced dramatically by 20- to 60-fold. MOR is glycosylated when expressed in HEK 293 cells; however, analysis of N glycosidase F-treated membranes indicated that N-glycan chains within the amino terminal domain of MOR do not contribute significantly to ligand affinities. These results indicate that amino acid residues within the amino-terminal domain of MOR play a crucial role in the composition of the binding pocket for a select group of agonists. PMID- 10719217 TI - Selective transport of SC1 mRNA, encoding a putative extracellular matrix glycoprotein, during postnatal development of the rat cerebellum and retina. AB - The selective transport of mRNA species into peripheral processes of cells is an important aspect of gene expression in the nervous system. In this study, we report the transport of SC1 mRNA into the distal processes of Bergmann glial (BG) cells at particular stages of development. SC1 is a putative anti-adhesive extracellular matrix (ECM) glycoprotein that is expressed not only in the developing central nervous system (CNS) but also in the adult brain. The intracellular distribution of SC1 mRNA was examined in two highly laminated neural structures, the cerebellum and retina, during postnatal development and in the adult rat. Our results indicate that SC1 mRNA expression is both spatially and temporally regulated. SC1 message was localized to BG cell bodies at postnatal day 5 (P5) and P10. However, by P15 through to the adult, SC1 mRNA was transported to distal processes of BG cells in the synapse-rich molecular layer (ML) of the cerebellum. In the developing rat retina, SC1 mRNA was expressed in specific neuronal populations by P10, however, transport of SC1 message to the dendrites of these retinal neurons was not detected during development or in the adult. These results indicate neural mechanisms which control the timing and cell type in which selective transport of SC1 mRNA is observed. The localization of SC1 mRNA to the distal processes of BG cells in the synapse-rich ML of the cerebellum could facilitate local control of SC1 protein synthesis which may play roles in synapse formation during development and in synaptic plasticity in the adult. PMID- 10719218 TI - Cellular prion protein binds laminin and mediates neuritogenesis. AB - Laminin (LN) plays a major role in neuronal differentiation, migration and survival. Here, we show that the cellular prion protein (PrPc) is a saturable, specific, high-affinity receptor for LN. The PrPc-LN interaction is involved in the neuritogenesis induced by NGF plus LN in the PC-12 cell line and the binding site resides in a carboxy-terminal decapeptide from the gamma-1 LN chain. Neuritogenesis induced by LN or its gamma-1-derived peptide in primary cultures from rat or either wild type or PrP null mice hippocampal neurons, indicated that PrPc is the main cellular receptor for that particular LN domain. These results point out to the importance of the PrPc-LN interaction for the neuronal plasticity mechanism. PMID- 10719219 TI - Localization of calcitonin receptor mRNA in the mouse brain: coexistence with serotonin transporter mRNA. AB - To elucidate the sites of and mechanisms of analgesic effect of centrally injected calcitonin, we examined expression of calcitonin receptor mRNA in the mouse brain by in situ hybridization techniques. Calcitonin receptor mRNA was expressed in various brain regions, including the preoptic area, dorsomedial hypothalamic nucleus, lateral hypothalamic area, periaqueductal gray, dorsal raphe nucleus, locus coeruleus, lateral parabrachial nucleus, gigantocellular reticular nucleus alpha part, lateral paragigantocellular nucleus, raphe magnus nucleus and solitary tract nucleus, which are known to play important roles in pain modulation. In addition, a double in situ hybridization technique demonstrated the intense expression of calcitonin receptor mRNA on serotonergic neurons in some raphe nuclei and the lateral paragigantocellular nucleus, suggesting the involvement of central serotonergic pathways in analgesic effect of calcitonin. PMID- 10719220 TI - Structural and functional characterization of 20S and 26S proteasomes from bovine brain. AB - Two proteins were isolated, in a stable form, from bovine brain by ion exchange chromatography, gel filtration and ultracentrifugation on glycerol gradient. They were identified as 20S and 26S proteasomes on the basis of molecular mass, migration velocity on non-denaturing gels, immunoreactivity, multipeptidase activity and the 26S proteasome also for dependence on ATP for the degradation of short peptides and ubiquitinylated proteins. However, the 26S proteasome has some properties not yet described for its counterpart of other tissues and from brain of this and other species. In particular, the ATP concentration required by the 26S proteasome to reach maximal peptidase activity was approximately 40-fold lower than the one required for maximal proteolytic activity on polyubiquitinylated substrates. Moreover, plots of substrate concentration vs. velocity gave a saturation curve for the 26S proteasome only, which, for the trypsin-like and post-glutamyl peptide hydrolase activities fitted the Michaelis Menten equation, whereas for the chymotrypsin-like activity indicated multibinding site kinetics with positive cooperativity (n = 2.32+/-0.38). As concerns the 20S proteasome, its electrophoretic pattern on native gel revealed a single protein band, a feature, to our knowledge, not yet described for the brain particle of any species. PMID- 10719221 TI - Amyloid beta peptides activate the phosphoinositide signaling pathway in oocytes expressing rat brain RNA. AB - Amyloid beta peptides (Abetas) of 39-43 amino acids constitute the major protein component of the amyloid plaques found in Alzheimer's disease brain. The generation of Abetas is regulated by the phosphoinositide (PI) pathway, which commonly couples to transmitter receptors. This study reports evidence for the activation of the PI pathway by Abetas in Xenopus oocytes expressing rat brain RNA. The naturally occurring peptides Abeta1-40 and Abeta1-42 were both active, whereas the cytotoxic fragment Abeta25-35 and the reverse peptide Abeta40-1 did not stimulate the PI pathway. Abetas rapidly lost potency in solution, suggesting that they were active only in their non-aggregated form. The Abeta response was saturable and not reduced by a substance P antagonist. This pharmacology excludes the participation of known Abeta binding proteins. The results indicate that a PI coupled receptor for non-aggregated Abeta may be present in brain. PMID- 10719222 TI - GFAP mRNA positive glia acutely isolated from rat hippocampus predominantly show complex current patterns. AB - Electrophysiologically complex glial cells have been widely identified from different regions of the central nervous system and constitute a dominant glial type in juvenile mice or rats. As these cells express several types of ion channels and neurotransmitter channels that were thought to be only present in neurons, this glial cell type has attracted considerable attention. However, the actual classification of these electrophysiologically complex glial cells remains unclear. They have been speculated to be an immature astrocyte because, although these cells show positive staining for the predominantly astrocytic marker S 100beta, it has not been possible to show staining for the commonly accepted mature astrocytic marker, glial fibrillary acidic protein (GFAP). To address the question of whether these cells might express GFAP at the transcript level, we combined patch-clamp electrophysiological recording with single cell RT-PCR for GFAP mRNA in glial cells acutely isolated from 4 to 12 postnatal day rats. In fresh cell suspensions from the CA1 region, complex glial cells were found to be the dominant cell type (65% total cells). We found that the majority of these electrophysiologically complex cells (74%) were positive for GFAP mRNA. We also showed that the complex cells responded to AMPA and GABA application, and these were also GFAP mRNA positive. We also fixed and stained the preparations for GFAP without electrophysiological recording to better preserve GFAP immunoreactively. In agreement with other studies, only 1.5% of these presumed electrophysiologically complex cells, based on morphology, showed immunoreactivity for GFAP. The expression of GFAP at the transcript level indicates GFAP (-)/GFAP mRNA (+) glial cells have an astrocytic identity. As single cell RT-PCR is able to detect both GFAP (-)/GFAP mRNA (+) and GFAP (+)/GFAP mRNA (+) astrocytic subtypes, the present study also suggests it is a feasible approach for astrocytic lineage studies. PMID- 10719223 TI - New dopamine receptor, D2(Longer), with unique TG splice site, in human brain. AB - Brain dopamine receptor agonists alleviate the signs of Parkinson's disease, while dopamine receptor antagonists alleviate hallucinations and delusions in psychosis. The dopamine type 2 receptor (or D2) is blocked by antipsychotic drugs, including even the "atypical" drugs such as clozapine or remoxipride, in direct relation to their clinical potencies. Compared to the long form of the D2 receptor (D2(Long)), the short form (D2(Short)) may be three times more sensitive to benzamide antipsychotic drugs. Hence, it is essential to identify additional variants of dopamine receptors for which more selective antipsychotic drugs can be found. Although no family linkage has been found between the D2 receptor and schizophrenia, there can be brain region abnormalities in the RNA transcript expression of dopamine receptors. Therefore, in order to identify variant dopamine D2 receptors, we searched for mutations in the RNA transcripts for the dopamine D2 receptor in the striatum of post-mortem brains from individuals who died with psychosis, including schizophrenia. A new splice variant of the D2 receptor, D2(Longer), with a unique TG splice site, was found in one control brain and in two psychotic brains. PMID- 10719224 TI - Mutant mice lacking ryanodine receptor type 3 exhibit deficits of contextual fear conditioning and activation of calcium/calmodulin-dependent protein kinase II in the hippocampus. AB - As it is known that ryanodine receptor type 3 is expressed in the hippocampus, we examined the contribution of this receptor to contextual fear conditioning behavior and to the activation of Ca(2+)/calmodulin-dependent protein kinase II using mice lacking the receptor. Ryanodine receptor type 3-deficient mice exhibited impairments of performance in the contextual fear conditioning test, passive avoidance test, and Y-maze learning test. Both the activities of Ca(2+)/calmodulin-dependent protein kinase IIbeta and Ca(2+)/calmodulin-dependent protein kinase IIalpha were significantly increased in the experimental group compared to the control group in the hippocampus, but not in the cingulate cortex on the testing day 24 h after contextual fear training. However, the activities of Ca(2+)/calmodulin-dependent protein kinase IIbeta and alpha were almost the same in the experimental and control groups in the hippocampus on the training day. Ryanodine receptor type 3-deficient mice did not show the increment of Ca(2+)/calmodulin-dependent protein kinase IIbeta and alpha activities in the hippocampus on the testing day. In addition, these mutant mice showed the reduction of fear response in the elevated plus-maze test. The present results suggest that calcium-induced calcium release through the activation of ryanodine receptor type 3 in the hippocampus is important to the expression of the performance of contextual learning through the elevation of Ca(2+)/calmodulin dependent protein kinase IIbeta and alpha activities. PMID- 10719225 TI - Immunocytochemical localization of a neuron-specific thrombospondin-1-like protein, NELL2: light and electron microscopic studies in the rat brain. AB - We have studied the cellular and intracellular localization of NELL2, a neural thrombospondin-1-like protein. NELL2 protein was detected as doublet bands of 140 and 90 kDa with the use of the specific antibodies raised against the C-terminal region of NELL2 and was recognized only in the brain but not in the peripheral tissues. Within the brain, NELL2 was abundantly present in the hippocampus and cerebral cortex, found moderately in the olfactory bulb and hypothalamus, and at a low level in the thalamus, cerebellum, and medulla. Immunocytochemically, NELL2 was seen only in neurons but not in glial cells or in the white matter. NELL2 immunoreactive cells were distributed throughout the brain with the highest density in the hippocampus and cerebral cortex. NELL2 was mainly found in the cell bodies of neurons and the immunoreactivity was often seen as dots in the perikarya. The distribution of NELL2 immunoreactivity did not completely correspond to that of any subtypes of protein kinase C (PKC). Under electron microscopy, NELL2 protein was associated with the endoplasmic reticulum (ER), especially with rough ER. NELL2 immunoreactivity was found in the restricted parts of the ER and found commonly inside the ER. These results suggest that NELL2 protein is synthesized by neurons and may be secreted from the neurons involved in certain neuronal functions. PMID- 10719226 TI - PLGamide characterization and role in osmoregulation in leech brain. AB - Neurons immunoreactive to an antiserum specifically directed against the Prolyl Leucyl-Glycinamide peptide (PLGamide: Melanocyte Inhibiting Factor: MIF) were detected in the brain of the leech Theromyzon tessulatum. Radioimmunoassay titrations of the PLGamide-like material at different physiological stages of the life cycle indicated a maximal amount at stage 3B, which is correlated to phase of both maximal water uptake and coelomic vitellogenin accumulation. In vivo experiments demonstrate that this cerebral PLGamide-like material is an anti diuretic factor that would act at stage 3B in order to permit a water uptake leading to water retention allowing coelomic yolk protein accumulation. In brains of the Gnatobdellid leech Hirudo medicinalis and the Pharyngobdellid leech Erpobdella octoculata, anti-PLGamide material was also detected with an amount not differing with the degree of sex maturation of the animals, confirming the link between osmoregulation and ovogenesis in rhynchobdellid leeches. Using a combination of biochemical techniques including high-pressure gel permeation chromatography followed by reversed-phase HPLC on brain extracts and Edman degradation, we demonstrated the presence of an authentic MIF-1 peptide in leech brain. Finally, since in vertebrates MIF-1 belongs to the non-classical opioid peptide family, we studied its binding displacement, in contrast to morphine, on mu-receptors and on nitric oxide (NO) release experiments in leech brain. PLGamide did not bind to mu-alkaloid opioid receptors and did not stimulate NO release. PMID- 10719228 TI - Enhanced levels of scrapie responsive gene mRNA in BSE-infected mouse brain. AB - The expression of the mRNA of nine scrapie responsive genes was analyzed in the brains of FVB/N mice infected with bovine spongiform encephalopathy (BSE). The RNA transcripts of eight genes were overexpressed to a comparable extent in both BSE-infected and scrapie-infected mice, indicating a common series of pathogenic events in the two transmissible spongiform encephalopathies (TSEs). In contrast, the serine proteinase inhibitor spi 2, an analogue of the human alpha-1 antichymotrypsin gene, was overexpressed to a greater extent in the brains of scrapie-infected animals than in animals infected with BSE, reflecting either an agent specific or a mouse strain specific response. The levels of spi 2 mRNA were increased during the course of scrapie prior to the onset of clinical signs of the disease and the increase reached 11 to 45 fold relative to uninfected controls in terminally ill mice. Spi 2, in common with four of the other scrapie responsive genes studied, is known to be associated with pro-inflammatory processes. These observations underline the importance of cell reactivity in TSE. In addition, scrg2 mRNA the level of which is enhanced in TSE-infected mouse brain, was identified as a previously unrecognized long transcript of the murine aldolase C gene. However, the level of the principal aldolase C mRNA is unaffected in TSE. The increased representation of the longer transcript in the late stage of the disease may reflect changes in mRNA processing and/or stability in reactive astrocytes or in damaged Purkinje cells. PMID- 10719227 TI - An autoradiographic study in mu-opioid receptor knockout mice. AB - An autoradiographic study was carried out to study the opioid binding sites in a mu-opioid receptor knock-out mouse line whose exon 2 and 3 were deleted. Mu opioid binding sites were undetectable in this knock-out mouse line while the binding of the other two types of receptors were unaltered. Our results suggest destroying functional mu-opioid receptor does not affect the expression of the other two opioid receptors. PMID- 10719230 TI - Oxidative stress status-the second set. PMID- 10719229 TI - Constraints on proper folding of the amino terminal domains of group III metabotropic glutamate receptors. AB - The glutamate binding site of the G-protein coupled metabotropic glutamate receptors (mGluRs) is contained within the large extracellular amino terminal domain (ATD) of the receptor. In this study, we examined the ligand binding properties and cellular dispositions of the membrane-bound mGluR4 and mGluR8 subtypes of mGluRs, and a series of truncated versions of these receptors. Truncation of the ATDs of mGluR4 and mGluR8 40 amino acids upstream of the first transmembrane domain produced soluble proteins that were secreted into the cell culture media of transfected human embryonic kidney cells. The soluble receptors retained ligand binding capabilities. Additional constructs of the ATDs of mGluR4 and mGluR8 were assessed for their ability to bind the agonist [(3)H]L-AP4 and for secretion from cells. A shorter mGluR4 construct truncated 98 amino acids upstream from the first transmembrane domain failed to bind [(3)H]L-AP4, while the analogous mGluR8 construct displayed a low level of binding. Unlike the full length receptors, which were expressed on the cell surface, or the soluble constructs which were secreted, the shorter constructs were primarily associated with intracellular membranes. These observations suggest that the cysteine-rich region may be important for efficient secretion, but not absolutely obligatory for ligand binding. Surprisingly, longer constructs encoding the entire ATDs of mGluR4 and mGluR8 failed to bind ligand and were localized intracellularly. Together, these findings demonstrate that there are strict limitations on the proper folding of truncated versions of the ATDs of mGluR4 and mGluR8. Specifically, all of the leucine-isoleucine-valine binding protein homology region, and part of the cysteine-rich region is required for optimal secretion in a soluble form that retains ligand binding activity. PMID- 10719231 TI - Measurement of F(2)-isoprostanes as an index of oxidative stress in vivo. AB - In 1990 we discovered the formation of prostaglandin F(2)-like compounds, F(2) isoprostanes (F(2)-IsoPs), in vivo by nonenzymatic free radical-induced peroxidation of arachidonic acid. F(2)-IsoPs are initially formed esterified to phospholipids and then released in free form. There are several favorable attributes that make measurement of F(2)-IsoPs attractive as a reliable indicator of oxidative stress in vivo: (i) F(2)-IsoPs are specific products of lipid peroxidation; (ii) they are stable compounds; (iii) levels are present in detectable quantities in all normal biological fluids and tissues, allowing the definition of a normal range; (iv) their formation increases dramatically in vivo in a number of animal models of oxidant injury; (v) their formation is modulated by antioxidant status; and (vi) their levels are not effected by lipid content of the diet. Measurement of F(2)-IsoPs in plasma can be utilized to assess total endogenous production of F(2)-IsoPs whereas measurement of levels esterified in phospholipids can be used to determine the extent of lipid peroxidation in target sites of interest. Recently, we developed an assay for a urinary metabolite of F(2)-IsoPs, which should provide a valuable noninvasive integrated approach to assess total endogenous production of F(2)-IsoPs in large clinical studies. PMID- 10719232 TI - Methods for measuring ethane and pentane in expired air from rats and humans. AB - Numerous studies in animals and humans provide evidence that ethane and pentane in expired air are useful markers of in vivo lipid peroxidation. The measurement of breath hydrocarbons, being noninvasive, is well suited for routine use in research and clinical settings. However, the lack of standardized methods for collecting, processing, and analyzing expired air has resulted in the use of a wide variety of different methods that have yielded highly disparate results among investigators. This review outlines the methods that we have developed and validated for measuring ethane and pentane in expired air from rats and humans. We describe the advantages of these methods, their performance, as well as potential errors that can be introduced during sample collection, concentration, and analysis. A main source of error involves contamination with ambient-air ethane and pentane, the concentrations of which are usually much greater and more variable than those in expired air. Thus, it appears that the effective removal of ambient-air hydrocarbons from the subject's lungs before collection is an important step in standardizing the collection procedure. Also discussed is whether ethane or pentane is a better marker of in vivo lipid peroxidation. PMID- 10719233 TI - Measurement of 3-nitrotyrosine and 5-nitro-gamma-tocopherol by high-performance liquid chromatography with electrochemical detection. AB - Nitric oxide (NO) is a lipophilic gaseous molecule synthesized by the enzymatic oxidation of L-arginine. During periods of inflammation, phagocytic cells generate copious quantities of NO and other reactive oxygen species. The combination of NO with other reactive oxygen species promotes nitration of ambient biomolecules, including protein tyrosine residues and membrane-localized gamma-tocopherol. The oxidative chemistry of NO and derived redox congeners is reviewed. Techniques are described for the determination of 3-nitro-tyrosine and 5-nitro-gamma-tocopherol in biological samples using high-performance liquid chromatography with electrochemical detection. PMID- 10719234 TI - Detection of oxidative DNA damage in human sperm and its association with sperm function and male infertility. AB - The expanding research interest in the last two decades on reactive oxygen species (ROS), oxidative stress, and male infertility has led to the development of various techniques for evaluating oxidative DNA damage in human spermatozoa. Measurement of 8-hydroxydeoxyguanosine (8-OHdG) offers a specific and quantitative biomarker on the extent of oxidative DNA damage caused by ROS in human sperm. The close correlations of 8-OHdG level with male fertility, sperm function and routine seminal parameters indicate the potential diagnostic value of this technique in clinical applications. On the other hand, single cell gel electrophoresis (SCGE or comet assay) and terminal deoxynucleotidyl transferase (TdT) mediated dUTP nick end labeling (TUNEL) assay have also been demonstrated to be sensitive, and reliable methods for measuring DNA strand breaks in human spermatozoa. As certain technical limitations were inherent in each of these tests, it is believed that a combination of these assays will offer more comprehensive information for a better understanding of oxidative DNA damage and its biological significance in sperm function and male infertility. PMID- 10719235 TI - Genotoxic potential of porphyrin type photosensitizers with particular emphasis on 5-aminolevulinic acid: implications for clinical photodynamic therapy. AB - Photodynamic therapy (PDT) uses exogenously administered photosensitizers activated by light to induce cell death or modulation of immunological cascades, presumably via formation of reactive oxygen species (ROS). 5-Aminolevulinic acid (ALA) mediated photosensitization is increasingly used for the treatment of nonmelanoma skin cancer and other indications including benign skin disorders. Long-term side effects of this investigational modality are presently unknown. Just as tumor treatments such as ionizing radiation and chemotherapy can cause secondary tumor induction, PDT may potentially have a carcinogenic risk. Evaluation of the biological effects of ALA in absence of activating light and analysis of the mechanism of ALA-PDT and porphyrin-type photosensitizers mediated photosensitization indicate that this therapy has a pro-oxidant and genotoxic potential. However, porphyrin type molecules also possess antioxidant and antimutagenic properties. ALA-PDT delays photocarcinogenesis in mice, and topical ALA alone does not increase skin cancer incidence in these animals. Patients with increased tissue levels of ALA have an increased incidence of internal carcinoma, however, it is not clear whether this relationship is casual or causal. There is no evidence indicating higher rates of skin cancer in patients with photosensitivity diseases due to presence of high protoporphyrin IX (PP) levels in skin. Overall, the presently available data indicate that the risk for secondary skin carcinoma after topical ALA-PDT seems to be low, but further studies must be carried out to evaluate the carcinogenic risk of ALA-PDT in conditions predisposed to skin cancer. PMID- 10719236 TI - Peroxidation of linoleate at physiological pH: hemichrome formation by substrate binding protects against metmyoglobin activation by hydrogen peroxide. AB - Peroxidation by metmyoglobin, MbFe(III), by metmyoglobin/hydrogen peroxide, MbFe(III)/H(2)O(2), to yield the myoglobin ferryl radical (*MbFe(IV)=O), or by ferrylmyoglobin, MbFe(IV)=O, was investigated at physiological pH (7.4) in oil-in water linoleate emulsions. Linoleate peroxidation was followed using second derivative ultraviolet (UV)-spectroscopy for monitoring formation of conjugated dienes and quantitative determination of specific linoleate hydroperoxides by liquid chromatography with photodiode absorption detection. Modifications of myoglobins during lipid peroxidation were followed simultaneously by changes in the Soret absorption band (410 or 424 nm), and in the visible absorption region (from 450 to 700 nm), combined with electron spin resonance (ESR) spectroscopy for direct detection of changes in the spin state of the iron center. In contrast to MbFe(IV)=O, MbFe(III) and MbFe(III)/H(2)O(2) were not able to initiate linoleate peroxidation in oil-in-water emulsions, and MbFe(III) was converted, by binding of linoleate (but not methyl linoleate), to a low-spin hemichrome derivate, HMbFe(III), with the distal histidine reversibly bound to the iron center. HMbFe(III) is ineffective in initiating lipid peroxidation and cannot be activated to *MbFe(IV)=O or MbFe(IV)=O by addition of moderate amounts of H(2)O(2). Addition of MbFe(III) to linoleate emulsions containing H(2)O(2) results in the competitive formation of *MbFe(IV)=O and HMbFe(III) in favor of HMbFe(III), and little linoleate peroxidation is detected, demonstrating the inherent protection, at physiologic pH, against peroxidation by reversible binding of the substrate to the potential myoglobin catalyst. PMID- 10719237 TI - Evaluation of autoantibodies against oxidized LDL and antioxidant status in top soccer and basketball players after 4 months of competition. AB - Antioxidant status and titers of autoantibodies against oxidized low-density lipoproteins (ox-LDL-Ab) were investigated in top soccer (S; n = 21, age 24.6 +/- 4.3 years) and basketball (B; n 3,000 mIU/ml) in ox-LDL-Ab were found in half the players (12S and 4B) with a maximum reaching 6000 mIU/ml (normal range: 200-600 mIU/ml), showing an in vivo LDL oxidation. There was no correlation between ox LDL-Ab titers and chlolesterol, LDL cholesterol, or antioxidant levels. Nevertheless, plasma vitamin C concentration was lower in athletes having high levels of ox-LDL-Ab when compared with those with normal levels (8.49 +/- 3.14 mirogram/ml vs. 10.39 +/- 2.55 microgram/ml), but this difference was not statistically significant. In conclusion, our data suggest that potential atherogenic and cardiovascular risks as reflected by high titers in ox-LDL-Ab may exist in some top athletes despite a nonaltered antioxidant status. PMID- 10719238 TI - Age-associated decline in gamma-glutamylcysteine synthetase gene expression in rats. AB - Although glutathione (GSH) concentration has been reported to diminish with age, the mechanism underlying such age-associated decline in the GSH content is not well understood. In this study, we compared the gene expression of both subunits of gamma-glutamylcysteine synthetase (GCS), the rate-limiting enzyme in de novo GSH synthesis, in young, adult, and old Fisher 344 rats. It was found that GCS activity was significantly decreased with increased age in liver, kidney, lung, and red blood cells (RBC). Parallel with the decreased enzyme activity, the protein and mRNA contents of both GCS subunits also changed inversely with age in liver, kidney, and lung, implying a decreased GCS gene expression during aging. Such a reduced GCS gene expression was accompanied by a decline in total GSH content without any change in cysteine concentration. Furthermore, the decreased GCS gene expression in old rats was not associated with a decline in the plasma insulin or cortisol level. This study showed, for the first time, that the expression of both GCS subunit genes was decreased in some organs of old rats, which would result in a reduced rate of GSH biosynthesis. Such decline in GSH synthetic capacity may underlie the observed decrease in GSH content during aging. PMID- 10719239 TI - Dominant-negative Jun N-terminal protein kinase (JNK-1) inhibits metabolic oxidative stress during glucose deprivation in a human breast carcinoma cell line. AB - Signal transduction pathway involved in glucose deprivation-induced oxidative stress were investigated in human breast carcinoma cells (MCF-7/ADR). In MCF 7/ADR, glucose deprivation-induced prolonged activation of c-Jun N-terminal kinase (JNK1) as well as cytoxicity and the accumulation of oxidized glutathione. Glucose deprivation also caused significant increases in total glutathione, cysteine, gamma-glutamylcysteine, and immunoreactive proteins corresponding to the catalytic as well as regulatory subunits of gamma-glutamylcysteine, and immunoreactive proteins corresponding to the catalytic as well as regulatory subunits of gamma-glutamylcysteine synthetase, suggesting that the synthesis of glutathione increased as an adaptive response. Expression of a catalytically inactive dominant negative JNK1 in MCF-7/ADR inhibited glucose deprivation- induced cell death and the accumulation of oxidized glutathione as well as altered the duration of JNK activation from persistent (> 2 h) to transient (30 min). In addition, stimulation of glutathione synthesis during glucose deprivation was not observed in cells expressing the highest levels of dominant negative protein. Finally, a linear dose response suppression of oxidized glutathione accumulation was noted for clones expressing increasing levels of dominant negative JNK1 during glucose deprivation. These results show that expression of a dominant negative JNK1 protein was capable of suppressing persistent JNK activation as well as oxidative stress and cytotoxicity caused by glucose deprivation in MCF-7/ADR. These findings support the hypothesis that JNK signaling pathways may control the expression of proteins contributing to cell death mediated by metabolic oxidative stress during glucose deprivation. Finally, these results support the concept that JNK signaling-induced shifts in oxidative metabolism may provide a general mechanism for understanding the diverse biological effects seen during the activation of JNK signaling cascades. PMID- 10719240 TI - In vitro study of the cytotoxicity of isolated oxidized lipid low-density lipoproteins fractions in human endothelial cells: relationship with the glutathione status and cell morphology. AB - Toxic effects of oxidized lipid compounds contained in oxidized LDL to endothelial cells are involved in the pathogenesis of atherosclerosis. Glutathione (GSH) plays an important role in the redox status of the cell and in the protective effect against oxidant injuries. However, little is known about the respective effect of these different oxidized lipid compounds toward cytotoxicity and GSH status of the cell. In this report, we isolated by high performance liquid chromatography oxidized lipid compounds from low-density lipoproteins (LDL) oxidized by copper and we examined their effects on cultured endothelial cells. Cytotoxicity and GSH status were determined after incubation of endothelial cells with crude LDL or isolated lipid fractions derived from cholesterol, phospholipids, or cholesteryl esters. Their effects on cell morphology were also assessed. Oxidized lipids coming from cholesteryl esters (hydroperoxides or short-chain polar derivatives) induced a slight but significant GSH depletion without inducing cytotoxicity. The same species coming from phospholipids induced a more pronounced GSH depletion and a cytotoxic effect which is only present for the more polar compounds (short-chain polar derivatives) and corresponding to a total GSH depletion. In contrast, fractions containing oxysterols had a larger cytotoxic effect than their effect on GSH depletion suggesting that their cytotoxic effects are mediated by a GSH independent pathway. All together, these data suggest that LDL-associated oxidized lipids present in copper-oxidized LDL exert cytotoxicity by an additional or synergistic effect on GSH depletion, but also by another mechanism independent of the redox status of the cell. PMID- 10719241 TI - Atypical effect of some spin trapping agents: reversible inhibition of acetylcholinesterase. AB - N-tert-butyl-alpha-phenylnitrone (PBN), a widely used nitrone-based free radical trap was recently shown to prevent acetylcholinesterase (AChE) inhibitors induced muscle fasciculations and brain seizures while being ineffective against glutamergic or cholinergic receptor agonist induced seizures. In the present study we compared the effects on AChE activity of four free radical spin traps PBN, alpha-(4-pyridil-1)-N-tert-butyl nitrone (POBN), N-tert-butyl-alpha-(2 sulfophenyl)-nitrone (S-PBN) and 5-diethoxyphosphoryl-5-methyl-1-pyrroline-N oxide (DEPMPO). The kinetics of AChE inhibition were studied in vitro using a spectrophotometric kinetic assay with AChE from rat brain, diaphragm, electric eel and mouse brain. Spin trapping compounds S-PBN and DEPMPO, in concentrations up to 3 mM did not inhibit hydrolysis of ACh, while PBN and POBN inhibited hydrolysis of ACh in a reversible and concentration-dependent manner. Double reciprocal plots of the reaction velocity against varying ACh concentrations at each inhibitor concentration were linear and generally indicated mixed type inhibition. PBN was the most potent inhibitor of mouse AChE with Ki and Ki' of 0.58 and 2.99 mM, respectively, and the weakest inhibitor of electric eel AChE. In contrast, POBN showed the highest affinity for electric eel enzyme, with Ki and Ki' values of 1.065 and 3.15 mM, respectively. These findings suggest that the effect of PBN and POBN on AChE activity does not depend on trapping of damaging reactive oxygen and that in addition to their antioxidant action other pharmacological effects of these compounds should be considered when neuroprotective actions of PBN or POBN are investigated. PMID- 10719242 TI - Alpha-phenyl-tert-butylnitrone (PBN) inhibits NFkappaB activation offering protection against chemically induced diabetes. AB - Alpha-phenyl-tert-butylnitrone (PBN) is an effective spin trapping agent by reacting with and stabilizing free radical species. Reactive oxygen species (ROS) have been implicated in pancreatic beta cell death and the development of insulin dependent diabetes mellitus (IDDM). We speculate that treatment with the PBN, will protect against diabetes development in two distinct chemically induced models for IDDM. Pretreatment with PBN (150 mg/kg ip) significantly reduced the severity of hyperglycemia in both alloxan- and streptozotocin (STZ) induced diabetes. To determine the mechanism by which PBN prevents hyperglycemia, we examined the ability of PBN to inhibit NFkappaB activation and to stabilize alloxan- and STZ-induced radicals. Both alloxan and STZ induced NFkappaB activation in the pancreas 30 min after their injection (50 mg/kg iv). PBN pretreatment inhibited both alloxan- and STZ-induced activation of NFkappaB and nitric oxide production. EPR studies showed that PBN could effectively trap alloxan-induced free radicals. It is clear that PBN can inhibit NFkappaB activation in the pancreas and reduce hyperglycemia in two distinct diabetogenic compounds. This research indicates that NFkappaB activation may be a key signal leading to beta cell death and IDDM. Understanding the cellular pathways leading to beta cell death may help in developing effective preventive or therapeutic targets for IDDM. PMID- 10719243 TI - Comparative effects of oxygen on indoleamine 2,3-dioxygenase and tryptophan 2,3 dioxygenase of the kynurenine pathway. AB - Indoleamine 2,3-dioxygenase (IDO) reacts with either oxygen or superoxide and tryptophan (trp) or other indoleamines while tryptophan 2,3-dioxygenase (TDO) reacts with oxygen and is specific for trp. These enzymes catalyze the rate limiting step in the kynurenine (KYN) pathway from trp to quinolinic acid (QA) with TDO in kidney and liver and IDO in many tissues, including brain where it is low but inducible. QA, which does not cross the blood-brain barrier, is an excitotoxin found in the CNS during various pathologies and is associated with convulsions. We proposed that HBO-induced convulsions result from increased flux through the KYN pathway via oxygen stimulation of IDO. To test this, TDO and IDO of liver and brain, respectively, of Sprague Dawley rats were assayed with oxygen from 0 to 6.2 atm HBO. TDO activity was appreciable at even 30 microM oxygen and rose steeply to a maximum at 40 microM. Conversely, IDO had almost no detectable activity at or below 100 microM oxygen and maximum activity was not reached until about 1150 microM. (Plasma contains about 215 microM oxygen and capillaries about 20 microM oxygen when rats breathe air.) KYN was 60% higher in brains of HBO convulsed rats compared to rats breathing air. While the oxygen concentration inside cells of rats breathing air or HBO is not known precisely, it is clear that the rate-limiting, IDO-catalyzed step in the brain KYN pathway (but not liver TDO) can be greatly accelerated in rats breathing HBO. PMID- 10719244 TI - Redox state of glutathione in human plasma. AB - Thiol and disulfide forms of glutathione (GSH) and cysteine (Cys) were measured in plasma from 24 healthy individuals aged 25-35 and redox potential values (E(h)) for thiol/disulfide couples were calculated using the Nernst equation. Although the concentration of GSH (2.8 +/- 0.9 microM) was much greater than that of GSSG (0.14 +/- 0.04 microM), the redox potential of the GSSG/2GSH pool (-137 +/- 9 mV) was considerably more oxidized than values for tissues and cultured cells (-185 to -258 mV). This indicates that a rapid oxidation of GSH occurs upon release into plasma. The difference in values between individuals was remarkably small, suggesting that the rates of reduction and oxidation in the plasma are closely balanced to maintain this redox potential. The redox potential for the Cys and cystine (CySS) pool (-80 +/- 9 mV) was 57 mV more oxidized, showing that the GSSG/2GSH and the CySS/2Cys pools are not in redox equilibrium in the plasma. Potentials for thiol/disulfide couples involving CysGly were intermediate between the values for these couples. Regression analyses showed that the redox potentials for the different thiol/disulfide couples within individuals were correlated, with the E(h) for CySS-mono-Gly/(Cys. CysGly) providing the best correlation with other low molecular weight pools as well as protein disulfides of GSH, CysGly and Cys. These results suggest that E(h) values for GSSG/2GSH and CySS-mono-Gly/(Cys. CysGly) may provide useful means to quantitatively express the oxidant/antioxidant balance in clinical and epidemiologic studies. PMID- 10719245 TI - Phosphine-induced oxidative damage in rats: attenuation by melatonin. AB - Phosphine (PH(3)), from hydrolysis of aluminum, magnesium and zinc phosphide, is an insecticide and rodenticide. Earlier observations on PH(3)-poisoned insects, mammals and a mammalian cell line led to the proposed involvement of oxidative damage in the toxic mechanism. This investigation focused on PH(3)-induced oxidative damage in rats and antioxidants as candidate protective agents. Male Wistar rats were treated ip with PH(3) at 2 mg/kg. Thirty min later the brain, liver, and lung were analyzed for glutathione (GSH) levels and lipid peroxidation (as malondialdehyde and 4-hydroxyalkenals) and brain and lung for 8 hydroxydeoxyguanosine (8-OH-dGuo) in DNA. PH(3) caused a significant decrease in GSH concentration and elevation in lipid peroxidation in brain (36-42%), lung (32 38%) and liver (19-25%) and significant increase of 8-OH-dGuo in DNA of brain (70%) and liver (39%). Antioxidants administered ip 30 min before PH(3) were melatonin, vitamin C, and beta-carotene at 10, 30, and 6 mg/kg, respectively. The PH(3)-induced changes were significantly or completely blocked by melatonin while vitamin C and beta-carotene were less effective or inactive. These findings establish that PH(3) induces and melatonin protects against oxidative damage in the brain, lung and liver of rats and suggest the involvement of reactive oxygen species in the genotoxicity of PH(3). PMID- 10719246 TI - In vitro supplementation with different tocopherol homologues can affect the function of immune cells in old mice. AB - Alpha-tocophorel (T) is the most common form of vitamin E inplasma and tissues. Alpha-T is also believed to be superior to its homologues beta-T, gamma-T, and delta-T in antioxidant activity. Biological activity of alpha-T has been intensively studied in a number of bodily systems. In contrast, the other homologues have received little attention beyond the evaluation of their relative antioxidant activity. We as well as others have previously shown that alpha-T can enhance cell- mediated immune function of aged animals and humans. Gamma-T is a principal form of vitamin E in the American diet and some cooking oils contain substantial amount of beta-T and delta-T. Thus it is of public health interest to compare their biological effects with than of alpha-t in various systems. In this study, we used an in vitro supplementation protocol to determine immunologic effects of these T homologues on murine splenocytes. The results showed that all four T homologues enhance both spontaneous and mitogen-stimulated lymphocyte proliferation (LP) and the maximal enhancement produced by them was of the same magnitude. The dose range to produce maximal enhancement varied with different homologues. The efficiency was in the order of beta-T approximately delta-T > alpha-T. Interestingly, at 50 (optimal for alpha-T) and 150 micromol/L, while alpha-T enhanced LP, all the other homologues inhibited LP. This inhibition was found to be due to their cytotoxicity at these levels. T homologues had a differential effect on interleukin (IL)-2 and prostaglandin (PG)E(2) production. IL-2 production by mouse splenocytes was not affected by alpha-T or beta-T, but was increased by gamma-T and delta-T. All T homologues, except for beta-T, inhibited PGE(2) alpha-T. Thus, all the T homologues enhance LP. However, the dose required to reach maximal enhancement varies among the homologues. On the other hand, they have a differential effect on IL-2 and PGE(2) production. The difference in nature and magnitude of the effect on immune function does not correlate with their reported relative antioxidant activity and might be due to minor differences in their structure important to their other biological activities. PMID- 10719247 TI - Glycochelates and the etiology of diabetic peripheral neuropathy. AB - People with diabetes are prone to develop peripheral vascular and nerve abnormalities which, in extreme cases, can lead to limb amputations. Although numerous theories have been advanced for these complications, no firm explanation is yet available. Recently, evidence has appeared suggesting that these vascular and nerve abnormalities may involve transition metals; administration of chelators such as desferrioxamine has been shown to prevent or actually reverse slowed peripheral nerve conduction and neuronal blood flow, as well as impaired endothelium-dependent arterial relaxation. Here, we argue that (i) the heavily glycated proteins known to accumulate in people with diabetes gain an increased affinity for transition metals such as iron and copper, (ii) as a result, proteins such as elastin and collagen within the arterial wall-which are known to be particularly heavily glycosylated in diabetes-may accumulate bound metal, especially copper, (iii) the bound metal causes the catalytic destruction of endothelium derived relaxing factor (nitric oxide or a derivative thereof), thereby engendering a state of chronic vasoconstriction. The resulting impairment of blood flow to peripheral nerves restricts the delivery of oxygen and nutrients and, in extremis, nerve death eventuates. If this hypothesis is proved correct, there are important implications for the development of novel pharmaceuticals for the treatment of diabetic peripheral neuropathy. PMID- 10719259 TI - Making features relevant: learning faces and event-related potentials recording using an analytic procedure. AB - We present a procedure designed for the learning of faces (represented by realistic drawings) and a task for the subsequent recording of Event-Related Potentials (ERPs), with the aim of investigating the psychobiological mechanisms involved in the recognition of familiar (or known) faces. The learning procedure consisted in a series of six sessions in which subjects familiarized themselves with the faces through a forced-choice features task: after the study of a set of 40 faces, each of these was presented at random in an incomplete way (without the eyes/eyebrows fragment), together with two sets of eyes and eyebrows for the subject to decide which of them corresponded to each face. This procedure is thought to facilitate for subjects the acquisition of the information related to the structural description of the faces (that is, without associated verbal/semantic information) using preferentially analytic processing strategies. The parameter d' of the Theory of Signal Detection allowed us to evaluate in the course of the learning sessions the degree of familiarization achieved with the studied faces. Subsequently, an ERPs' recording session was carried out, during which subjects executed a face-feature matching task. In this task, similar in structure to that carried out in the learning sessions, the subject had to decide whether the automatic completion of the faces that was made was correct or incorrect. ERPs' effects related to mismatching features were obtained, which indicate the existence of specific cerebral mechanisms involved in the recognition of faces. PMID- 10719260 TI - Exchange transfusion with fluorocarbon for studying synaptically evoked optical signal in rat cortex. AB - Optical imaging of intrinsic signal is a powerful technique for studying the functional organization of the brain [T. Bonhoeffer, D. S. Kim, D. Malonek, D. Shoham, A. Grinvald, Optical imaging of the layout of functional domains in area 17 and across the area 17/18 border in cat visual cortex, Eur. J. Neurosci. 7 (1995) 1973-1988; M. Hubener, D. Shoham, A. Grinvald, T. Bonhoeffer, Spatial relationships among three columnar systems in cat area 17, J. Neurosci. 17 (1997) 9270-9284; D. Malonek, A. Grinvald, Interactions between electrical activity and cortical microcirculation revealed by imaging spectroscopy: implications for functional brain mapping, Science 272 (1996) 551-554; A. Shmuel, A. Grinvald, Functional organization for direction of motion and its relationship to orientation maps in cat area 18, J. Neurosci. 16 (1996) 6945-6964] [1] [10] [14] [22]. Three components of intrinsic optical signal can be distinguished. Two of these components can be attributed either to changes in blood volume or to changes in oxygen consumption [R.D. Frostig, E.E. Lieke, D.Y. Ts'o, A. Grinvald, Cortical functional architecture and local coupling between neuronal activity and the microcirculation revealed by in vivo high resolution optical imaging of intrinsic signals, Proc. Natl. Acad. Sci. U. S. A. 87 (1990) 6082-6086] [7]. The origin of the third component is not yet clear but the component seems to be based on scattered light [H.U. Dodt, G. D'Arcangelo, E. Pestel, W. Zieglgansberger, The spread of excitation in neocortical columns visualized with infrared-dark field videomicroscopy, NeuroReport 7 (1996) 1553-1558; K. Holthoff, O.W. Witte, Intrinsic optical signals in rat neocortical slices measured with near-infrared dark-field microscopy reveal changes in extracellular space, J. Neurosci. 16 (1996) 2740-2749; B.A. MacVicar, D. Hochman, Imaging of synaptically evoked intrinsic optical signals in hippocampal slices, J. Neurosci. 11 (1991) 1458-1469; L. Trachsel, H.U. Dodt, W. Zieglgansberger, The intrinsic optical signal evoked by chiasm stimulation in the rat suprachiasmatic nuclei exhibits GABAergic day-night variation, Eur. J. Neurosci. 8 (1996) 319-328] [3] [9] [13] [24]. A spectral fitting method with three components is used for the analysis of intrinsic optical signal [M. Nemoto, Y. Nomura, C. Sato, M. Tamura, K. Houkin, I. Koyanagi, H. Abe, Analysis of optical signals evoked by peripheral nerve stimulation in rat somatosensory cortex: dynamic changes in hemoglobin concentration and oxygenation, J. Cereb. Blood Flow Metab. 19 (1999) 246-259] [17]. In order to validate the analysis, we need the knowledge on contribution of signal resulted from hemoglobin to total intrinsic optical signal. The exchange transfusion with fluorocarbon has the advantage that can change the spectral contribution of hemoglobin [M. Ferrari, M.A. Williams, D.A. Wilson, N.V. Thakor, R.J. Traystman, D.F. Hanley, Cat brain cytochrome-c oxidase redox changes induced by hypoxia after blood-fluorocarbon exchange transfusion, Am. J. Physiol. 269 (1995) H417-H424; A.L. Sylvia, C.A. Piantadosi, O(2) dependence of in vivo brain cytochrome redox responses and energy metabolism in bloodless rats, J. Cereb. Blood Flow Metab. 8 (1988) 163-172] [6] [23]. Here we describe a new method of the reduction of hemoglobin signal from somatosensory evoked optical intrinsic signal in rat cortex by the combination of exchange transfusion with fluorocarbon and imaging system of thinned skull cranial window. The method allows for the study of the synaptically evoked changes in light scattering as well as fluorescence of calcium indicator or voltage-sensitive dye without absorption of hemoglobin. PMID- 10719261 TI - Microdialysis in the mouse nucleus accumbens: a method for detection of monoamine and amino acid neurotransmitters with simultaneous assessment of locomotor activity. AB - Microdialysis has been extensively used to characterize the effects of drugs of abuse on extracellular levels of various neurotransmitters in nucleus accumbens (NAc) of the rat brain. However, recent advances in mouse genetics have prompted the need for studying the in vivo neurochemical correlates of drug intake in genetically engineered mice. While an earlier study has shown the feasibility of measuring monoamines in the NAc of behaving transgenic mice [I. Sillaber, A. Montkowski, R. Landgraf, N. Barden, F. Holsboer, R. Spanagel, Enhanced morphine induced behavioural effects and dopamine release in the nucleus accumbens in a transgenic mouse model of impaired glucocorticoid (type II) receptor function: influence of long-term treatment with the antidepressant moclobemide, Neuroscience, 85 (1998) 415-425 [16] ], in this protocol we demonstrate a method for measuring both monoamine and amino neurotransmitters from the NAc of freely moving mice combined with open field locomotor activity monitoring. Mice were implanted with guide cannulae aimed at the NAc and allowed 4 days of recovery before being implanted with microdialysis probes equipped with 1-mm cuprophane membranes. On the following day, mice were placed in plexiglass chambers equipped with infrared photobeams, where microdialysis samples and locomotor activity data were collected in 10-min intervals. Immediately after collection, microdialysis samples were split into two equal aliquots for separate analysis of monoamine and amino acid neurotransmitter content. High performance liquid chromatography (HPLC) analysis revealed that norepinephrine, dopamine, serotonin, aspartate, glutamate, glycine, taurine, and gamma-aminobutyric acid (GABA) could be detected in each microdialysis sample. Thus, we have shown it is feasible to monitor extracellular levels of multiple neurotransmitters with simultaneous measurement of locomotor behavior in the mouse, making this model suitable for studying differential neurochemical and behavioral responses to drugs of abuse in genetically engineered mice. PMID- 10719262 TI - Neurophysiological evaluation of tactile space perception deficits through transcranial magnetic stimulation. AB - We describe a procedure useful to investigate the contralateral space perception deficits frequently encountered in patients with unilateral right brain damage. In particular, we focused on the phenomenon of extinction, i.e., the failure to perceive a contralesional stimulus only when a symmetrical contralateral stimulus is simultaneously applied. Fifteen right brain- and 15 left brain-damaged patients were examined. Somatosensory perception was evaluated by using a dedicated electronic device able to provide electrical stimuli of variable intensity to digits of one or both hands. The electrical stimulator was able to trigger a magnetic brain stimulator connected with a focal figure of eight coil. Threshold electrical stimuli were delivered to one or both hands of the patients, who were asked to indicate whether they perceived the stimulus (i) and to localise it (them). The electrical stimulator was connected with a magnetic stimulator with an interstimulus interval (ISI) of 40 msec (electrical stimulation preceding the transcranial one). Focal threshold transcranial magnetic stimulation (TMS) was applied to frontal and parietal scalp sites of the unaffected hemisphere. At each interpulse interval we found that TMS of the unaffected hemisphere was associated to a decrease in the level of contralesional extinction. Our method demonstrates that a basic deficit underlying neglect and extinction of contralateral space in unilaterally brain damaged patients is the interhemispheric imbalance between the two hemispheres in directing contralateral attention. A transient interference with the function of the unaffected hemisphere can improve these deficits, suggesting a possible application of TMS in the daily clinical practice for speeding up recovery from neglect. PMID- 10719263 TI - Gold toning preserves integrity of silver enhanced immunogold particles during osmium tetroxide treatment for demonstration of a biogenic amine. AB - We describe a protocol that enhances immunolabelling of nervous tissue for ultrastructural study. Insect tissue is fixed, sectioned, and labelled with a polyclonal antiserum against serotonin and a secondary antibody conjugated with 1 nm colloidal gold. The gold particles are silver-enhanced to ease detection and then protected by gold toning. Finally, the tissue is post fixed in glutaraldehyde fixative followed by osmium tetroxide and further processed for electron microscopy. We demonstrated on insect nervous tissue that gold toning protects marker particles from the influence of osmium tetroxide. Use of buffered solutions throughout the protocol led to well preserved ultrastructural details, and marker particle size was not reduced with a short gold toning time. We also suggest use of this protocol for vertebrate or other invertebrate tissue. PMID- 10719264 TI - Immunocytochemical detection of two nuclear proteins within the same neuron using light microscopy. AB - We developed a method of double immunocytochemistry (ICC) that can be used with conventional light microscopy for localizing two different nuclear proteins. The procedure involves two sequential rounds of ICC that both employ the avidin and biotin conjugated enzyme (ABC) amplification method, separated by an Avidin D and biotin blocking step to reduce non-specific avidin-biotin reactions. Round one of ICC employs the use of avidin and biotin conjugated alkaline phosphatase (ABC-AP) and the Vector Red (VR) substrate, which produces a red colorimetric reaction product. The second round of ICC makes use of avidin and biotin conjugated peroxidase (ABC-HRP) and the Vector(R) SG substrate, which produces a gray colorimetric reaction product. Neuronal nuclei that are double-labeled for both proteins appear red with a gray core. This protocol allows the simultaneous detection of two proteins within the same subcellular compartment of a single neuron, without the need for epifluorescence or scanning confocal laser microscopy. PMID- 10719265 TI - Immunocytochemical detection of fos protein combined with anterograde tract tracing using biotinylated dextran. AB - The present report deals with an axonal tract-tracing procedure in rat enabling visualization of anterogradely transported biotinylated dextran amine (BDA) combined with immnunocytochemical detection of Fos protein following electrical stimulation of the brain. This method allows us to evaluate whether a given structure, receiving both injection of BDA and electrical stimulation, elicits neuronal activation in another part of the brain via direct or indirect projections. We have used the method at the light microscopic level to determine the connectivity of the sensorimotor cortex in the rat. In various parts of the forebrain and brainstem, BDA-labeled fibers originating from the cortex were observed in close apposition to Fos-like immunoreactive cells (FLI) activated by stimulation. This result suggests a direct (probably monosynaptic) projection. On the contrary, FLI neurons were observed in areas devoid of direct afferents, indicating a cascade of activations. The method described in this protocol is applicable for functional anatomy purposes elsewhere within the central nervous system. It constitutes a preliminary step in identifying the validity of a pathway before examination of the reality of the monosynaptic relationship at the electron microscopic level. PMID- 10719266 TI - Using event-related fMRI to assess delay-period activity during performance of spatial and nonspatial working memory tasks. AB - Event-related experimental design and analysis techniques for functional magnetic resonance imaging (fMRI) take advantage of the intrinsic temporal resolution of fMRI to permit investigation of complex human behaviors on the time scale over which they can occur. The protocol described in this report permits the effective isolation and assessment of variance in the fMRI signal that is attributable solely to the delay portion of delayed-response tasks. It permits, therefore, evaluation of the purely mnemonic portions of working memory tasks without requiring the "cognitive subtraction" of nonmnemonic components of such tasks, such as visual processing and motor output. Features of this event-related fMRI technique include the empirical derivation of an impulse response function (IRF) from each subject participating in the experiment, single-subject and random effects group analyses, use of t-values of dependent measures, and the use of regions of interest (ROI) to improve the sensitivity of a priori contrasts. This report provides a detailed exposition of the research methodology of our event related fMRI technique, the rationale behind many of its critical features, and examples of its application to two empirical datasets. PMID- 10719267 TI - Using intracranial electrical stimulation to study the timing of prepulse inhibition of the startle reflex. AB - Due to the short latency and briefness of the startle reflex, event-related inhibition of startle has high temporal resolution and is useful for studying the hierarchical organization of sensorimotor gating and motive-motor gating. In this article, we describe methods for measuring the inhibitory effects of electrically stimulating each of the following four brain structures on startle in awake rats: the inferior colliculus (IC), the deeper layers of the superior colliculus (SC), the pedunculopontine tegmental nucleus (PPTg), and the ventral pallidum (VP). These four brain structures have been reported to be important in mediating sensorimotor or motive-motor gating. Startle responses are elicited by either intense noise bursts or electrical stimulation of the principal trigeminal nucleus. The time course of the IC-inhibited startle reflex is used as a standard for estimating timing of the neural transfer of startle-inhibitory information to motor outputs. We also discuss how these methods can be used in combination with neuropharmacology. PMID- 10719268 TI - Delivering drugs, via microdialysis, into the environment of extracellularly recorded hippocampal neurons in behaving primates. AB - Hippocampal neurons in primates have been extensively studied with electrophysiological and neuroanatomical methods. Much less effort has been devoted to examining these cells with contemporary pharmacological techniques. Therefore, we modified a recently developed integrative technique (N. Ludvig, P.E. Potter, S.E. Fox, Simultaneous single-cell recording and microdialysis within the same brain site in freely behaving rats: a novel neurobiological method, J. Neurosci. Methods 55 (1994) 31-40 [9] ) for cellular neuropharmacological studies in behaving monkeys. A driveable microelectrode microdialysis probe guide assembly was implanted stereotaxically into the left hippocampus of squirrel monkeys (Saimiri sciureus) under isoflurane anesthesia. The assembly was covered with a protective cap. After 3 weeks of postsurgical recovery and behavioral training, the experimental subject was seated in a primate chair. For 4-5 h, single-cell recording and microdialysis were simultaneously performed in the hippocampal implantation site. The technique allowed the recording of both complex-spike cells and fast-firing neurons without the use of head restraint. The control microdialysis solution, artificial cerebrospinal fluid (ACSF), was replaced with either 1 M ethanol or 500 microM N methyl-D-aspartate (NMDA) for 10-30 min intervals. The ethanol perfusions principally suppressed the firing of the neurons in the dialysis area. The NMDA perfusions initially increased the firing of local neurons, then caused electrical silence. These drug delivery/cell recording sessions were performed with 1-4 day intersession intervals over a 1-month period. The described method provides a tool to elaborate the pharmacology of primate hippocampal neurons during behavior and without the confounding effects of systemic drug administrations. PMID- 10719269 TI - Isolation and characterization of adult microglial cells and oligodendrocytes derived from postmortem human brain tissue. AB - The present study provides a detailed description of the simultaneous establishment and immunocytochemical characterization of highly enriched human adult microglial cell cultures as well as of oligodendrocyte cultures. For this study, brain tissue specimens were collected at autopsy with relatively short postmortem times (3-9 h) from various regions of the CNS of Alzheimer's disease, Pick's disease and non-demented control cases. Although methods to isolate viable glial cells from human adult brain tissue have been described, these human brain specimens were often derived from surgical resections, i.e., in order to treat intractable epilepsy, brain tumors or cardiovascular diseases involving the brain. However, for the study of many neurological disorders, surgical material is not available. Furthermore, for obvious reasons, there is a limit to the number of central nervous system (CNS) regions from which (enough) tissue can be obtained at surgery. The adherent primary microglial cells, isolated according to the here described procedures consisted of proliferating, phagocytotic cells that expressed various microglia/macrophage-specific markers as judged by immunocytochemical analysis. Non-adherent cells isolated from the same brain tissue samples expressed oligodendrocyte-specific markers. The current described culture system may provide a valuable tool in studying human CNS biology and disease. PMID- 10719270 TI - Improved recovery of highly enriched mitochondrial fractions from small brain tissue samples. AB - The investigation of mitochondrial abnormalities in brain commonly requires isolation of these organelles from small tissue samples. We have modified a mitochondrial isolation procedure based on Percoll density gradient centrifugation to increase the proportion of the total mitochondrial pool recovered while reducing contamination with synaptosomes and related structures containing cytoplasm. Initially, myelin was removed by centrifugation in 12% Percoll in isotonic buffer. The pellet was resuspended, treated with digitonin to break up synaptosomes and similar structures and subjected to discontinuous Percoll density gradient centrifugation. The mitochondrial fraction obtained from this procedure was highly metabolically active and well coupled, exhibiting respiratory control ratios above 5. The recovery of mitochondrial markers using a single rat forebrain as starting material was approximately 18% to 21%. When small tissue samples (approximately 50 mg wet weight) were used as starting material the recovery of the mitochondrial marker was approximately 16%. The ratio of recovery of a mitochondrial marker to the cytoplasmic marker lactate dehydrogenase exceeded 200 in preparations from a single rat forebrain. This is substantially greater than values reported for previously published procedures reflecting both an improved yield of mitochondria and a reduction in cytoplasmic contamination. PMID- 10719271 TI - Methods for analysis of Ca(2+)/H(+) antiport activity in synaptic vesicles isolated from sheep brain cortex. AB - The involvement of Ca(2+)-storage organelles in the modulation of synaptic transmission is well-established [M.K. Bennett, Ca(2+) and the regulation of neurotransmitter secretion, Curr. Opin. Neurobiol. 7 (1997) 316-322 [1]; M.J. Berridge, Neuronal calcium signaling, Neuron 21 (1998) 13-26 [2]; Ph. Fossier, L. Tauc, G. Baux, Calcium transients and neurotransmitter release at an identified synapse, Trends Neurosci. 22 (1999) 161-166 [7] ]. Various Ca(2+) sequestering reservoirs (mitochondria, endoplasmic reticulum and synaptic vesicles) have been reported at the level of brain nerve terminals [P. Kostyuk, A. Verkhratsky, Calcium stores in neurons and glia, Neuroscience 63 (1994) 381-404 [18]; V. Mizuhira, H. Hasegawa, Microwave fixation and localization of calcium in synaptic terminals using X-ray microanalysis and electron energy loss spectroscopy imaging, Brain Res. Bull. 43 (1997) 53-58 [21]; A. Parducz, Y. Dunant, Transient increase of calcium in synaptic vesicles after stimulation, Neuroscience 52 (1993) 27-33 [23]; O.H. Petersen, Can Ca(2+) be released from secretory granules or synaptic vesicles?, Trends Neurosci. 19 (1996) 411-413 [24] ]. However, the knowledge of the specific contribution of each compartment for spatial and temporal control of the cytoplasmic Ca(2+) concentration requires detailed characterization of the Ca(2+) uptake and Ca(2+) release mechanisms by the distinct intracellular stores. In this work, we described rapid and simple experimental procedures for analysis of a Ca(2+)/H(+) antiport system that transport Ca(2+) into synaptic vesicles at expenses of the energy of a DeltapH generated either by activity of the proton pump or by a pH jumping of the vesicles passively loaded with protons. This secondary active Ca(2+) transport system requires high Ca(2+)100 microM) for activation, it is dependent on the chemical component (DeltapH) of the proton electrochemical gradient across the synaptic vesicle membrane and its selectivity is essentially determined by the size of the transported cation [P.P. Goncalves, S.M. Meireles, C. Gravato, M.G. P. Vale, Ca(2+)-H(+)-Antiport activity in synaptic vesicles isolated from sheep brain cortex, Neurosci. Lett. 247 (1998) 87-90 [10]; P.P. Goncalves, S.M. Meireles, P. Neves, M.G.P. Vale, Ionic selectivity of the Ca(2+)/H(+) antiport in synaptic vesicles of sheep brain cortex, Mol. Brain Res. 67 (1999) 283-291 [11]; P.P. Goncalves, S.M. Meireles, P. Neves, M.G.P. Vale, Synaptic vesicle Ca(2+)/H(+) antiport: dependence on the proton electrochemical gradient, Mol. Brain Res. 71 (1999) 178-184 [12] ]. The protocols described here allow to ascertain the characteristics of the Ca(2+)/H(+) antiport in synaptic vesicles and, therefore, may be useful for clarification of the physiological role of synaptic vesicles in fast buffering of Ca(2+) at various sites of the neurotransmission machinery. PMID- 10719272 TI - Technique for collecting cerebrospinal fluid in the cisterna magna of non anesthetized rats. AB - We developed a technique for collecting cerebrospinal fluid (CSF) from the cisterna magna in non-anesthetized adult and young pup rats. In the adults, CSF was collected through a previously implanted guide cannula without previous disruption of the cisterna magna. In the pups, CSF was directly aspirated through a syringe from the cisterna in awake animals without previous surgery. In the adults, the volume of CSF collected varied from 50 to 120 microl, and in pups 7 to 10 days old, it was approximately 25 microl. The technique can easily be done by anyone who is familiar with stereotaxic surgery, and the material needed is cheap and easy to obtain commercially. A simple procedure to calculate the parameters for the implantation of guide cannula in rats other than Wistar ones is also presented. PMID- 10719273 TI - Cytochemical and immunocytochemical methods for electron microscopic detection of glucose-6-phosphate dehydrogenase in brain areas. AB - This paper reports on protocols for the cytochemical and immunocytochemical determination of the glucose-6-phosphate dehydrogenase (G6PD) in brain areas by electron microscopy (EM). The cytochemical assay consists of a pre-embedding staining of small and flat tissue blocks, which were first mildly fixed and then floated in a staining mixture based on the reduction of tetrazolium salts by NADPH. Tissue blocks were then washed, post-fixed in OsO(4), dehydrated through graded ethanol concentrations and embedded in resin. Ultrathin sections were then obtained and observed at the EM. The immunocytochemical technique was performed on completely fixed tissues of perfused animals. After the tissue embedding in resin, ultrathin sections were obtained and treated with a primary anti erythrocyte G6PD antibody, produced and purified in our laboratory. The immunostaining was performed with secondary gold-conjugated antibody. Gold grains were well evident by EM analysis thus revealing the G6PD protein in the subcellular compartments. These protocols are useful to detect peculiar populations of neurons which express high levels of G6PD to sustain processes of neural plasticity in some brain areas. PMID- 10719274 TI - Effects of orbital spaceflight on human osteoblastic cell physiology and gene expression. AB - During long-term spaceflight, astronauts lose bone, in part due to a reduction in bone formation. It is not clear, however, whether the force imparted by gravity has direct effects on bone cells. To examine the response of bone forming cells to weightlessness, human fetal osteoblastic (hFOB) cells were cultured during the 17 day STS-80 space shuttle mission. Fractions of conditioned media were collected during flight and shortly after landing for analyses of glucose utilization and accumulation of type I collagen and prostaglandin E(2) (PGE(2)). Total cellular RNA was isolated from flight and ground control cultures after landing. Measurement of glucose levels in conditioned media indicated that glucose utilization occurred at a similar rate in flight and ground control cultures. Furthermore, the levels of type I collagen and PGE(2) accumulation in the flight and control conditioned media were indistinguishable. The steady-state levels of osteonectin, alkaline phosphatase, and osteocalcin messenger RNA (mRNA) were not significantly changed following spaceflight. Gene-specific reductions in mRNA levels for cytokines and skeletal growth factors were detected in the flight cultures using RNase protection assays. Steady-state mRNA levels for interleukin (IL)-1alpha and IL-6 were decreased 8 h following the flight and returned to control levels at 24 h postflight. Also, transforming growth factor (TGF)-beta(2) and TGF-beta(1) message levels were modestly reduced at 8 h and 24 h postflight, although the change was not statistically significant at 8 h. These data suggest that spaceflight did not significantly affect hFOB cell proliferation, expression of type I collagen, or PGE(2) production, further suggesting that the removal of osteoblastic cells from the context of the bone tissue results in a reduced ability to respond to weightlessness. However, spaceflight followed by return to earth significantly impacted the expression of cytokines and skeletal growth factors, which have been implicated as mediators of the bone remodeling cycle. It is not yet clear whether these latter changes were due to weightlessness or to the transient increase in loading resulting from reentry. PMID- 10719275 TI - Differential involvement of matrix vesicles during the initial and appositional mineralization processes in bone, dentin, and cementum. AB - The distribution of matrix vesicles and its role in biological mineralization were examined in bone and dental hard tissues of the rat after daily administrations of 1-hydroxyethylidene-1, 1-bisphosphonate (HEBP), a potent inhibitor of mineralization, for 7 or 14 days. Newly formed, nonmineralized matrices of the HEBP-affected bone and mesodermal dental hard tissues other than circumpulpal dentin contained numerous mineral-filled matrix vesicles (MV), randomly distributed throughout the collagenous matrix. The distribution density of the mineral-filled MV in the HEBP-affected matrices of calvaria, metaphyseal trabecular bone, alveolar bone, and cellular cementum ranged from 60 to 70 per 100 microm(2), and no statistically significant differences were noted among the values. In the HEBP-affected dentin, however, MV were located only in the nonmineralized matrix of mantle dentin and totally absent in the circumpulpal dentin layers. Instead, the HEBP-affected circumpulpal dentin contained a dense meshwork of noncollagenous matrix enriched with calcium and phosphorus. Comparable meshwork structures were undetectable in nonmineralized matrices of the other hard tissues affected by HEBP. These observations suggest that a certain population of MV (60-70 per 100 microm(2)) is involved in the process of appositional mineralization in most of the mesodermal hard tissues, in addition to their well-known role in initial mineral induction in these tissues. Circumpulpal dentin appears to be an exception, where MV are not required for the appositional mineralization process. Exclusive localization of dentin phosphoproteins in circumpulpal dentin layers must take place to facilitate appositional mineralization at the calcification front, in the absence of MV. PMID- 10719276 TI - Characterization of the trabecular rat bone mineral: effect of ovariectomy and bisphosphonate treatment. AB - Bisphosphonates, potent inhibitors of bone resorption, have been used clinically to correct the continued loss of bone mass in osteoporosis and in other conditions. However, there has been some concern that long-term treatment with these compounds, as well as more recently developed drugs, may also decrease the rate of bone formation. Bisphosphonates, which are strongly bound to hydroxyapatite crystals, may alter the structure and reactivity of the crystals, interfere with new crystal nucleation and growth, as well as alter the short range order of newly formed crystals. We have investigated the chemistry and structure of the solid calcium-phosphate mineral phase of lumbar vertebrae of ovariectomized, 6.5-month-old rats treated with bisphosphonates for 1 year after onset of osteopenia. Appropriate control groups were used for comparison. The techniques used to assess the mineral phase were chemical analyses, Fourier transform-infrared (FT-IR) and FT-Raman spectroscopy, FT-IR microspectroscopy, and phosphorus-31 magic-angle-sample spinning nuclear magnetic resonance spectroscopy ((31)P MAS NMR). The (31)P MAS NMR spectra of trabecular bone of lumbar vertebrae of control, ovariectomized, and treated animals were similar. However, there were several significant differences in the results obtained by FT IR spectroscopy of the whole tissue samples, FT-IR microspectroscopy of sections of bone, and chemical analyses. For example, whereas chemical analyses demonstrated that the CO(3) content of the mineral phase of the ovariectomized animals was decreased compared with controls, FT-IR microspectroscopy of bone sections showed no changes in the relative CO(3) content, but some changes in the environment of the CO(3) groups. However, chemical analyses of the crystals, combined with data from all three spectroscopic methods and with data from serum analysis, did indicate small changes in the mineral phase after ovariectomy, corrected after treatment with bisphosphonates. In any event, the chemical and structural data in the present studies demonstrate that the bisphosphonate, tiludronate, does not significantly alter the mineral components of bone after 1 year of treatment during the course of which bone loss was reversed. PMID- 10719277 TI - Structural alterations in rat skin and bone collagen fibrils induced by ovariectomy. AB - In this study, the influence of ovariectomy in rat skin and bone (trabecular and cortical) collagen fibrils is examined using electron microscopy. Structural changes (fibril architecture and diameter) were detected, at the ultrastructural level, in skin and bone specimens from ovariectomized rats. The overall collagen fibril architecture was disturbed as compared with normal animals. Treated collagen fibrils' mean diameter values were significantly smaller than those from controls, in all tissues examined. The banding patterns of fibrils were normal in all cases; however, measurements by a computerized method of measuring axial periodicity of fibrils indicated significantly lower values for treated samples than untreated samples. Our results show a correlation between the effects induced by ovariectomy in skin and bone collagen. But, the question of whether these changes play a role in the pathogenesis of ovarian hormone deficiency in osteoporosis remains to be demonstrated. PMID- 10719278 TI - Effects of hormonal conditions and drugs on both muscle and bone strength can be assessed in a single rat test. AB - Strength of both muscles and bone are important for fracture prevention in osteoporotic individuals. Therefore, drugs that are preclinically tested in animals for preventing or treating osteoporosis, and reducing fracture risk, should not only be checked for their effects on bone strength, but also for those on muscle strength. We developed a rat model to measure both in the same animal, using a single test. The model is based on an in vivo, ventral three-point bending test of the lower leg (Nordsletten L. and Ekeland A. J Orthop Res 11:299 304; 1993). This model was developed to test the contribution of triceps surae muscle contraction to the strength of the tibia. We hypothesized that this same test can be applied to determine bone and muscle strength independently, in an absolute sense. To investigate this possibility, the muscle contribution to bone stresses was estimated from mechanical analyses, based on direct assessment of muscle strength in a separate test. Sixteen mature female Wistar rats were used, half of which were ovariectomized. After 12 weeks, the rats were tested in vivo in three-point bending of the right lower leg during muscle contraction, and then the isolated triceps surae muscle strength in the left lower leg was measured separately, in another model. The rats were then killed, and the left nude shafts were tested mechanically in three-point bending in vitro to determine structural strength of the bone alone. Ultimate external bending moments of the in vivo and in vitro tests, maximal muscle force, and geometrical parameters formed the basis for the analysis. While contracting, the triceps surae loads the tibia in axial compression and bending. We found that the axial compressive stress on the bone due to muscle contraction was less than 2.5% of the bending stress this produced. This indicates that muscle contribution to lower leg strength is due almost entirely to the bending moment it produces, counteracting the external moment put on the leg by the testing device. Thus, the difference between the in vivo (lower leg) and in vitro (nude tibia) failure bending moments is approximately equal to the maximal muscle bending moment. This information can be applied to test the effects of hormonal conditions and drugs on both muscle and bone strength independently, in a single rat test, using the aforementioned procedure. PMID- 10719279 TI - Aging-induced osteopenia in avian cortical bone. AB - Cortical bone loss contributes substantially to the degradation of skeletal integrity associated with aging. However, animal models that closely mimic age related alterations in cortical bone are limited. The objective of this study was to determine if aged rooster cortical bone demonstrates phenotypic alterations similar to those observed in aged human cortical bone (i.e., expansion of the endocortical and periosteal envelopes and elevated cortical porosity). When compared with young adult roosters, aged roosters demonstrated significant expansion of the endocortical (16%) and periosteal (10%) envelopes, resulting in significantly increased cross-sectional moments of inertia. In addition, aged rooster bone demonstrated significantly elevated cortical porosity (51%) and average area of porosity (83%). We conclude that rooster bone demonstrates age related adaptations similar to those of humans at both tissue and cellular levels, and may therefore represent a relatively useful, inexpensive animal model for investigating the mechanisms of age-related bone loss. PMID- 10719280 TI - Immunochemical characterization of assay for carboxyterminal telopeptide of human type I collagen: loss of antigenicity by treatment with cathepsin K. AB - The assay for the cross-linked carboxyterminal telopeptide of type I collagen (ICTP) has been shown to reflect increased type I collagen degradation in such pathological conditions as bone metastases and rheumatoid arthritis, but to be rather insensitive to the changes in physiological bone collagen turnover (e.g., induced by estrogen or bisphosphonate treatment). To determine the reasons for this discrepancy we localized the antigenic determinant recognized by the ICTP assay and studied the effects of two major osteoclastic proteinases, cathepsin K (EC 3.4.22.38) and matrix metalloproteinase-9 (MMP-9; gelatinase B; EC 3.4.24.35), on immunoreactivity. The antigenic determinant was shown to reside within the hydrophobic phenylalanine-rich regions of the carboxyterminal telopeptides of the two alpha1 chains of human type I collagen, situated between the triple helical domain and the lysine-derived trivalent cross-link. This conclusion was based on differences between the amino acid sequences and cross reactivities of the corresponding human and bovine antigens before and after proteolytic treatments with chymotrypsin. A trivalent cross-link is necessary for providing such a structure, because the divalently cross-linked and monomeric natural and synthetic peptides from the same region, but containing only one phenylalanine-rich sequence, showed poor immunoreaction. Recombinant human cathepsin K cleaved the trivalently cross-linked ICTP structure at two sites between the phenylalanine-rich region and the cross-link, destroying the reactivity with ICTP antibodies. On the contrary, the treatment of isolated ICTP by the matrix metalloproteinases MMP-9 (gelatinase B), MMP-1 (collagenase 1), or MMP-13 (collagenase 3) had no effect on the immunoreaction. Our results indicate that the increased circulating concentrations of ICTP found in several clinical situations are most likely produced by matrix metalloproteinases, whereas cathepsin K-mediated, osteoclastic bone resorption destroys ICTP antigenicity. PMID- 10719281 TI - Decline in osteocyte lacunar density in human cortical bone is associated with accumulation of microcracks with age. AB - Despite osteocytes' ideal position to sense the local environment and thereby influence bone remodeling, the function of osteocytes in bone remains controversial. In this study, histomorphometric examination of male and female femoral middiaphyseal cortical bone was conducted to determine if bone's remodeling response, indicated by tissue porosity and accumulation of damage, is associated with osteocyte lacunar density (number of osteocyte lacunae per bone area). The results support the sensory role of the osteocyte network as the decline in osteocyte lacunar density in human cortical bone is associated with the accumulation of microcracks and increase in porosity with age. Porosity and microcrack density increased exponentially with a decline in osteocyte lacunar density indicating that a certain minimum number of osteocytes is essential for an "operational" network. No gender-related differences were found in the relationship of osteocyte lacunar density to age, porosity, or microcrack density. The coefficient of variation of osteocyte lacunar density increased linearly with age, indicating that aging bone tissue is characterized by increased heterogeneity in the spatial organization of osteocytes. Osteocyte lacunar density, porosity, and microcrack density exhibited the same exponential probability density distribution in the donor population, indicating their regulation by similar biological phenomena. PMID- 10719282 TI - Annual skeletal balance and metabolic bone marker changes in healthy early postmenopausal women: results of a prospective study. AB - The aim of this study was to establish the duration and annual rate of menopause related bone loss and to investigate the relationship between bone turnover and bone loss in early healthy postmenopausal women. The rate of change in bone mineral density (BMD) at the lumbar spine and in bone turnover was measured twice at the exact interval of 12 months by dual-energy X-ray absorptiometry (DXA) and by the determination of plasma alkaline phosphatase levels (ALP) and fasting urinary hydroxyproline/creatinine ratio (OHPr/Cr), respectively, in 123 healthy premenopausal and postmenopausal women 45-60 years of age. The subjects were divided into nine groups according to their menstrual status and years since menopause (YSM). Annual bone loss at the lumbar spine of women who were menopausal for 1, 2, 3, 4, and 5 years was -2.62 +/- 0.37 (95% confidence interval -3.66, -1.58), -3.87 +/- 0.96 (-6.02, -1.73), -2.50 +/- 0. 37 (-3.29, 1.70), -2.86 +/- 0.73 (-4.44, -1.27), and -1.54 +/- 0.41 (-2.42, -0.66), respectively, and was significantly less than zero. But, the annual bone loss of women who were premenopausal or menopausal for 6, 7, and 8 years was -0.76 +/- 0.60 (-2.04, +0.53), -1.16 +/- 0.68 (-2.61, +0.29), 0.24 +/- 0.48 (-0.78, +1.26), and 0. 16 +/- 0.63 (-1.18, -1.49), respectively, and was not significantly different from zero. These results demonstrate that the early hormone-dependent bone loss commences in the first year after menopause and is arrested within 6 years after the onset of menopause. The overall bone loss for this phase is estimated to be approximately 15%. Annual change in ALP and OHPr/Cr seems to indicate that bone resorption prevails on bone formation in the first 2 YSM, whereas osteoblastic activity relatively prevails from YSM 3 to YSM 5, which explains the progressive repairing of the imbalance between bone resorption and formation. PMID- 10719283 TI - Prediction of fracture from low bone mineral density measurements overestimates risk. AB - There is a well-established relationship between bone mineral density (BMD) and fracture risk. Estimates of the relative risk of fracture from BMD have been derived mainly from short-term studies in which the correlation between BMD at assessment and BMD in later life ranged from 0.8 to 0.9. Because individuals lose bone mineral at different rates throughout later life, the long-term predictive value of low BMD is likely to decrease progressively with time. This article examines and formalizes the relationship between current BMD, correlation coefficients, and long-term risk. The loss of predictive value has important implications for early assessment and supports the view that measurements should be optimally targeted at the time interventions are contemplated and, when necessary, repeated in later life. PMID- 10719284 TI - "how do anti-osteoporotic agents prevent fractures?" abstracts from the round table held at the XVI annual meeting of the argentine association of osteology & mineral metabolism (AAOMM) city of bahia blanca, october 29, 1999 PMID- 10719286 TI - The role of lymphocytes in allergic disease. AB - In the last few years strong evidence has accumulated to suggest that allergen reactive type-2 T helper (T(H)2) cells play an important role in the induction and maintenance of the allergic inflammatory cascade. First, cytokines and chemokines produced by T(H)2 cells (GM-CSF, IL-4, IL-5, IL-6, IL-9, IL-10, IL-13, macrophage-derived chemokine) and those produced by other cell types in response to T(H)2 cytokines or as a reaction to T(H)2-related tissue damage (eotaxin, transforming growth factor-beta, IL-11) account for most pathophysiologic aspects of allergic disorders (production of IgE antibodies; recruitment or activation of mast cells, basophils, and eosinophils; mucus hypersecretion; subepithelial fibrosis; and tissue remodeling). The T(H)2 hypothesis may also explain the complex genetic background responsible for allergic disorders. Several genes are involved in the development and regulation of T(H)2 cells and may provide the reason why the prevalence of atopic allergy is increasing in Western countries. Indeed, a dramatic change has occurred in the last several decades in the "microbial" environment of children, thus probably altering the balance between T(H)1 and T(H)2 responses to "innocuous" antigens (allergens) in favor of T(H)2 responses. Finally, the T(H)2 hypothesis offers exciting opportunities for the development of novel immunotherapeutic strategies targeted to address allergen specific T(H)2 cells or T(H)2-derived effector molecules in atopic individuals. PMID- 10719287 TI - The diagnostic evaluation, treatment, and prevention of allergic contact dermatitis in the new millennium. AB - Identifying the etiology of allergic contact dermatitis is a rewarding yet challenging endeavor. Not all allergic contact reactions are eczematous in appearance. The most reliable clinical clue to the allergic nature of the dermatitis is its geographic distribution. Once a list of culprit allergens has been identified by patch testing, the practitioner must identify the relevant allergen(s) and counsel the patient in avoidance. For most individuals, allergen avoidance results in resolution of the dermatitis; however, some patients will require continuing symptomatic therapy despite avoidance. For those patients unable to avoid known allergens, immunosuppressant therapies (including phototherapy) or barriers can be beneficial. Currently, hyposensitization is not a viable alternative for the treatment of allergic contact dermatitis. PMID- 10719288 TI - Regulation of immunologic homeostasis in peripheral tissues by dendritic cells: the respiratory tract as a paradigm. AB - Dendritic cells are now recognized as the gatekeepers of the immune response, possessing a unique potential for acquisition of antigens at extremely low exposure levels and for efficient presentation of these in an immunogenic form to the naive T-cell system. Dendritic cell populations throughout the body exhibit a wide range of features in common that are associated with their primary functions, and these are considered in the initial section of this review. In addition, it is becoming evident that the properties and functions of these cells are refined by microenvironmental factors unique to their tissues of residence, a prime example being mucosal microenvironments such as those in respiratory tract tissues, and the latter represents the focus of the second section of this review. PMID- 10719289 TI - IL-13 genetics: pale rider or horse of a different color? PMID- 10719290 TI - Autoallergy: a pathogenetic factor in atopic dermatitis? AB - Long before the discovery of IgE it was reported that human dander extract can elicit immediate-type skin reactions in patients with severe atopy and that this skin sensitivity can be passively transferred with serum. Several recent findings have rekindled the interest in this phenomenon and led to the concept that IgE autoreactivity may play a pathogenetic role in severe and chronic forms of atopy. The elucidation of the nature of several environmental allergens has revealed striking structural and immunologic similarities with human proteins. It was also reported that patients predominantly with severe and chronic manifestations of atopy (eg, atopic dermatitis) contain IgE autoantibodies against a wide variety of proteins expressed in histogenetically unrelated human cell types and tissue specimens. Last, complementary DNAs coding for autoallergens were isolated from human expression complementary DNA libraries and recombinant autoallergens were produced. The autoallergens characterized to date represent mainly intracellular proteins, but some of them could be detected as IgE immune complexes in sera of sensitized patients. We suggest that at least two pathomechanisms could play a role in autoallergy. First, autoallergens may cross-link effector cell-bound IgE autoantibodies and, by release of inflammatory mediators, lead to immediate-type symptoms. Second, IgE-mediated presentation of autoallergens may activate autoreactive T cells to release proinflammatory cytokines, contributing to the magnitude of the allergic tissue reaction. PMID- 10719291 TI - Endogenous nitric oxide in allergic airway disease. AB - There has been intense research into the role nitric oxide (NO) plays in physiologic and pathologic mechanisms. The presence of NO in exhaled breath and the high concentrations in nasal airways stimulated many studies examining exhaled and nasal NO as potential markers of airway inflammation, enabling repeated monitoring of airway inflammation not possible with invasive tests (eg, bronchoscopy). In airway inflammation, NO is not merely a marker but may have anti-inflammatory and proinflammatory effects. Nasal NO measurement may be used in the noninvasive diagnosis and monitoring of nasal disease. This review was compiled by speakers who gave presentations on NO at the annual meeting of the American Academy of Allergy, Asthma, and Immunology in 1999 on exhaled and nasal NO, in vitro studies of NO, the chemistry of airway NO formation, and standardized measurement of exhaled mediators. PMID- 10719292 TI - Relationship of nasal carriage of Staphylococcus aureus to pathogenesis of perennial allergic rhinitis. AB - BACKGROUND: Several studies have previously shown some factors that modify the pathogenesis of perennial allergic rhinitis (PAR). However, the association between bacterial colonization and PAR has not been well understood. OBJECTIVE: We sought to study the association between superantigen-producing Staphylococcus aureus and PAR. METHODS: S aureus colonization in the nasal cavity and its superantigen production were studied in 65 patients with PAR and 45 nonallergic control subjects. The nasal symptom scores of the patients were evaluated. Furthermore, we examined the response to staphylococcal enterotoxin B or toxic shock syndrome toxin 1 of peripheral blood lymphocytes from both patients and control subjects by measuring proliferative responses and production of cytokines (IFN-gamma, IL-4, and IL-5). RESULTS: The rate of nasal carriage of S aureus in the patients (44%) was significantly higher than that of the control subjects (20%, P <.01). Moreover, the rate of nasal carriage of superantigen-producing S aureus in the patients (22%) was significantly higher than that of the control subjects (6.7%, P <.05). The nasal symptom scores of the S aureus -positive patients were significantly higher than those of the S aureus -negative patients (P <.05), although there was no significant association between symptom scores and superantigen production. Peripheral blood lymphocytes from the patients showed significantly higher proliferative responses and were more likely to produce T(H2 )type cytokines in response to superantigens (P <.01), but the responses were not different among the patients with S aureus carriage and superantigen production. CONCLUSION: This study suggests that PAR leads to a higher carriage rate of S aureus, and nasal carriage of S aureus may aggravate PAR. PMID- 10719293 TI - Demonstration of mast cell chemotactic activity in bronchoalveolar lavage fluid collected from asthmatic patients before and during pollen season. AB - BACKGROUND: Mast cells are versatile effector cells of primary importance in asthma and airway inflammation. During inflammation mast cells accumulate in the bronchial epithelium. The mechanism for this increase in mast cell number has not been defined. OBJECTIVES: The aim of this study was to examine the presence of mast cell chemotactic activity in bronchoalveolar lavage (BAL) fluid taken before and at the end of 2 pollen seasons from patients with allergic asthma. METHODS: To measure mast cell chemotactic activity, we used a modified Boyden chamber and the human mast cell line HMC-1 or in vitro-developed mast cells as responder cells. RESULTS: A total of 27 patients were investigated, of which 8 exhibited mast cell chemotactic activity in their BAL fluid collected before season. A significant increase in the activity was found in 18 of 27 BAL fluids sampled at the end of the pollen season. No difference was found between patients treated with immunotherapy or placebo. The presence of stem cell factor could be detected in all BAL fluids analyzed. Blocking antibodies against stem cell factor or transforming growth factor-beta partially blocked the activity in some of the BAL fluids. Treatment of the responder cells with pertussis toxin reduced the migratory activity in 13 of 14 BAL fluids collected during pollen season. CONCLUSION: This study demonstrates the presence of mast cell chemotactic activity in BAL fluids from patients with allergic asthma, with a significant increase in activity during pollen season. The major part of this activity consisted of factors mediating their effect through G(i)-protein coupled receptors. This activity may be responsible for the mast cell accumulation in the intraepithelial layer seen in allergic asthmatic patients. PMID- 10719294 TI - Alcoholic drinks: important triggers for asthma. AB - BACKGROUND: Although anecdotally alcoholic drinks seem to be common triggers for asthma, little is known of the prevalence, the characteristics, or the mechanisms underlying these reactions. OBJECTIVES: The primary aim of this study was to determine the frequency and characteristics of asthmatic reactions triggered by alcoholic drinks in a community-based cohort of asthmatic subjects. Investigations of other food sensitivities were also completed to explore some of the possible components of alcoholic drinks that may be responsible for these asthmatic responses. METHODS: A validated food allergy questionnaire was used to assess the characteristics of alcoholic drink-induced asthma in 366 adult patients recruited from the Asthma Foundation of Western Australia. The food allergy questionnaire was sent out by mail and self-administered by recipients. RESULTS: Thirty-three percent of respondents indicated that alcoholic drinks had been associated with the triggering of asthma on at least 2 occasions. Wines were the most frequent triggers, with responses being rapid in onset (<1 hour) and of mild to moderate severity. Logistic regression analysis indicated that wine induced asthmatic reactions were reported more often by women (P =.032), by those taking oral steroids (P =.021), by individuals who had reported their first asthma attack at a younger age (P <. 001), and by those who had previously visited an alternative health practitioner for asthma (P =.041). A significant association between wine-induced asthma and asthma triggered by sulfite containing foods (P <.001) and by aspirin and nonsteroidal anti-inflammatory medicines (P =.01) was also observed. CONCLUSION: Alcoholic drinks, and particularly wines, appear to be important triggers for asthmatic responses. Sensitivity to the sulfite additives in wines seems likely to play an important role in many of these reactions. Sensitivities of individuals to salicylates present in wines may also play a role. PMID- 10719295 TI - The determination of equivalent doses of standardized allergen vaccines. AB - BACKGROUND: Standardized allergen vaccines are tested for potency by manufacturers by using assays proposed in their license applications and approved by the Center for Biologics Evaluation and Research, which reviews and verifies the test results before lot release. The current lot-release limits for mite and grass pollen allergen vaccines impose statistical equivalence to the national reference extract; thus the limits are primarily based on assay variability. OBJECTIVE: We sought to establish a clinical basis for lot-release limits for the relative potency of allergen vaccines and to evaluate alternative specifications. METHODS: We performed literature selection and review, linear and logistic regression analyses of selected studies, and analysis of lots submitted to the Food and Drug Administration for approval since 1995. RESULTS: Therapeutic equivalence is achieved over a 10-fold range of allergen concentration. Safety equivalence is more difficult to assign, but on the basis of injection data, a 4 fold increase in allergen concentration is associated with a 5% to 10% increase in adverse reaction rates. The SD in log relative potency for the submitted allergenic products was determined to be 0.090 for grasses and 0.061 for mites, compared with 0.079 for competition ELISA. CONCLUSIONS: Current lot-release limits are well within literature-based estimates of therapeutic, diagnostic, and safety equivalence ranges for the clinical use of allergen vaccines. In addition, the aggregate consistency of the submitted products is comparable with the precision of the assay that is used to assess the products. These results support expanded release limits for verification of relative potency, provided the submitted lots of material remain at their present level of consistency. PMID- 10719296 TI - Allergen provocation augments endotoxin-induced nasal inflammation in subjects with atopic asthma. AB - BACKGROUND: Recent epidemiologic and in vivo studies have suggested that inhaled endotoxin plays an important role in asthma pathogenesis. OBJECTIVE: The present study examines the effect of nasal allergen provocation on subsequent endotoxin challenge in subjects with atopic asthma. METHODS: By using a split-nose randomized crossover design, individual nares of 12 asthmatic subjects underwent challenge and lavage as follows. Immediately after a baseline nasal lavage, one nares received normal saline, and the other received dust mite antigen. Four hours later, both nares were exposed to either saline or endotoxin. Dust mite antigen (Dermatophagoides farinae) and endotoxin (Escherichia coli 026:B6) doses were 100 AU and 1000 ng, respectively. Postchallenge lavages were done at 8 and 24 hours after the initial challenge. The subjects then returned a minimum of 3 weeks later for crossover to the study arm. Nasal lavage fluid was analyzed for total and differential cell counts, IL-8, IL-6, intercellular adhesion molecule 1, GM-CSF, eosinophil cationic protein, myeloperoxidase, and soluble CD14. RESULTS: A significant increase in the total inflammatory cell count was seen at 8 hours for the dust mite/endotoxin exposure compared with the saline/saline and saline/endotoxin exposures. Differential cell counts revealed a similar neutrophilic and eosinophilic inflammation for the dust mite/endotoxin exposure at 8 hours. CONCLUSIONS: These data demonstrate an interaction between allergen and endotoxin exposure in asthmatic subjects, suggesting that a prior allergen challenge significantly augments the endotoxin-induced inflammation. Moreover, these data provide further evidence that concomitant exposure to allergen and endotoxin may be an important factor in asthma pathogenesis. PMID- 10719297 TI - The stability of house dust mite allergens in glycerinated extracts. AB - BACKGROUND: Mite allergen vaccines are important diagnostic and immunotherapeutic reagents. Previous studies on mite allergen stability under different storage conditions have yielded contradictory results. OBJECTIVE: We sought to compare, over a 12-month period, the stability of mite allergens reconstituted in 50% glycerol and stored at different temperatures and to examine the role of protease inhibitors in enhancing allergen stability. METHODS: Lyophilized allergen extracts were reconstituted in 50% glycerol, with and without protease inhibitors, and stored at -70 degrees C, -20 degrees C, 4 degrees C, or 37 degrees C for 12 months. At 6 and 12 months, the extracts were compared with freshly dissolved extracts by competition ELISA with pooled allergic sera, 2-site ELISA with mite-specific mAbs, and immunoblot analyses. RESULTS: The overall potencies of the stored extracts measured by competition ELISA were stable at -20 degrees C and 4 degrees C. As determined by means of the immunoblot and 2-site ELISA, Der f 1 levels decreased at 4 degrees C. Levels of Der f 2, Der p 1, and Der p 2 decreased in at least one of the allergen-specific assays. Storage at 37 degrees C led to overall loss of potency and allergen content, whereas storage at -70 degrees C was associated with a moderate loss of potency that increased with multiple freeze-thaw cycles. Protease inhibitors had no effect on allergen stability. CONCLUSION: Although overall potency of the extracts, as measured by competition ELISA, was preserved at -20 degrees C and 4 degrees C, allergen specific assays indicated loss of allergens. These findings suggest that the competition ELISA is insensitive to decreases in the concentrations of individual allergens. PMID- 10719298 TI - Onset of action of intranasal budesonide (Rhinocort aqua) in seasonal allergic rhinitis studied in a controlled exposure model. AB - BACKGROUND: Intranasal budesonide aqueous nasal spray (BANS) is recognized as an efficacious treatment for seasonal allergic rhinitis (SAR), but the time to onset of action is not known. OBJECTIVE: The primary objective was to evaluate the time at which the onset of action of BANS in the symptomatic relief of seasonal allergic rhinitis becomes evident within 12 hours after a single dose in a controlled ragweed pollen exposure setting. METHODS: The study was of a double blind, randomized, parallel-group design, testing BANS (64 microgram and 256 microgram) and placebo on ragweed-sensitive subjects with symptoms for at least 1 year by using a controlled pollen challenge system (Environmental Exposure Unit). The efficacy variables were the combined nasal score (the sum of blocked nose, runny nose, and sneezing-itchy nose), individual nasal symptoms, overall evaluation of treatment efficacy reported by participants on diaries, and peak nasal inspiratory flow (PNIF). RESULTS: A total of 217 participants were treated with BANS or placebo. At 7 to 12 hours, BANS was better than placebo in reducing combined nasal and blocked nose symptoms. For PNIF, the time to onset of action was shortest for 256 microgram of BANS relative to placebo (3 hours, P =.003). BANS 64 microgram was better than placebo in reducing the individual scores of blocked nose, runny nose, and sneezing-itchy nose from 3 to 5 hours after administration. Treatment efficacy was higher for those receiving BANS compared with placebo starting at 5 hours. All treatments were well tolerated, and no specific adverse events occurred. CONCLUSIONS: The onset of action of intranasal BANS was 7 hours according to combined nasal and blocked nose symptom scores. Evidence of earlier response was observed at 3 hours for runny nose and PNIF. PMID- 10719299 TI - Induced sputum: comparison of postinfectious cough with allergic asthma in children. AB - BACKGROUND: Cough persisting after a respiratory infection is common in children and is often managed as asthma. However, little is known about the pathophysiologic mechanisms of such cough and how it compares with asthma. OBJECTIVE: We used the technique of induced sputum to examine the inflammatory index values associated with persistent cough or allergic asthma in children. We hypothesized that the sputum from children with persistent postinfectious cough would differ from that of children with allergic asthma in that the former would lack eosinophils compared with the latter. STUDY DESIGN: Sputum production was induced with hypertonic saline solution in 34 children: 12 with cough persisting for 1 month or more after an apparent respiratory tract infection, not treated with corticosteroid; 11 with untreated atopic asthma, not using inhaled corticosteroid; and 11 with treated atopic asthma using inhaled corticosteroid. RESULTS: The percentage of eosinophils in the sputum of children with cough was significantly lower than in the sputum of children with untreated allergic asthma (median 0.5% vs 14.5%, P <.0001). Similarly, the percentage of eosinophils in the sputum of children with asthma treated with inhaled steroids was significantly lower compared with untreated asthmatic children (1.5% vs 14.5%, P <.0001). The peripheral blood eosinophils, serum eosinophil cationic protein, and nasal percent eosinophils of the patients with cough were also significantly lower than those from patients with untreated asthma. Methacholine challenge in 6 of the 11 cough patients tested showed mild-to-moderate hyperresponsiveness, whereas the other 5 had a negative methacholine challenge. CONCLUSIONS: Children with persistent postinfectious cough do not have airway eosinophilia typical of untreated asthma. Despite the absence of eosinophilic inflammation, some of the patients with chronic cough had reactive airways. These results suggest that postinfectious cough in children has different pathophysiologic features than allergic asthma and probably represents a different disease. PMID- 10719300 TI - Dependence of mast cell IgE-mediated cytokine production on nuclear factor-kappaB activity. AB - BACKGROUND: The transcription factor nuclear factor-kappaB (NF-kappaB) has been implicated in the regulation of a number of inflammatory cytokines and has been the proposed target for anti-inflammatory therapeutics. OBJECTIVE: Our purpose was to explore the role of NF-kappaB in the regulation of allergic inflammation. METHODS: To determine whether NF-kappaB is activated during IgE-mediated reactions and what types of mediators it regulates, a mutant form of IkappaB was used to block the ability of NF-kappaB to translocate to the nucleus and promote the transcription of selected genes. RESULTS: Mouse bone marrow-derived mast cells stimulated by IgE receptor cross-linking exhibited an activation of NF kappaB as assessed by electrophoretic mobility shift assays. Transfected mast cells expressing the mutant IkappaB showed very little NF-kappaB activation. Both control and transfected cells released beta-hexosaminidase after specific antigen challenge, and this release could be potentiated by exogenous adenosine. Transfected mast cells that failed to develop NF-kappaB activation did not produce IL-6 messenger RNA or protein after IgE-mediated stimulation, but these cells retained the ability to produce transcripts for IL-4 and IL-5 in spite of the suppression of NF-kappaB activity. CONCLUSIONS: It appears that NF-kappaB is activated during IgE-mediated allergic inflammation and that this activity is necessary for the production of IL-6, but not IL-4 or IL-5. When considering the use of agents that target NF-kappaB to reduce inflammatory processes, it is important to know precisely which cytokines are under its control. PMID- 10719301 TI - A cluster of seven tightly linked polymorphisms in the IL-13 gene is associated with total serum IgE levels in three populations of white children. AB - BACKGROUND: Increased levels of total serum IgE are a strong risk factor for the development of asthma. IgE is also involved in host defenses against parasites and fungi. Linkage of total serum IgE with markers located close to the 3 Mb cluster of cytokine genes in chromosome 5q31 has been reported. IL-4 or IL-13 are regarded as essential for IgE synthesis. OBJECTIVE: We tested whether polymorphisms in the IL-13 gene might explain the linkage between chromosome 5q31 and total serum IgE levels. METHODS: We used denaturing HPLC to detect polymorphisms in overlapping PCR fragments of the IL-13 gene including promoter and 3' untranslated regions. After sequencing was performed to identify the locations of the polymorphisms, PCR and primer-induced restriction site assays were used to genotype subjects in 3 unselected populations. RESULTS: We report here 7 polymorphisms (6 novel) in IL-13. Four of these polymorphisms are tightly linked to a variant in the terminal portion of the coding region of the gene that results in a predicted amino acid change in residue 130 (Arg130Gln). The Gln form is strongly associated (P =.000002) with increased serum IgE levels in 3 different populations comprising a total of 1399 children. Two additional polymorphisms in the promoter region of IL-13 are more loosely linked to Arg130Gln and are also less significantly associated with total serum IgE levels. CONCLUSION: These data suggest that the Arg130Gln polymorphism in IL-13, or others in close linkage with it, is associated with the development of the elevated serum IgE phenotype. PMID- 10719302 TI - Peptides obtained by tryptic hydrolysis of bovine beta-lactoglobulin induce specific oral tolerance in mice. AB - BACKGROUND: Oral tolerance against food proteins has been achieved in different animal models with use of native or moderately hydrolyzed proteins as inducers. However, native proteins remain highly allergenic, although it has been demonstrated that protein hydrolyzates and resulting peptides can lose their allergenicity. OBJECTIVE: This study was designed to evaluate the ability of beta lactoglobulin hydrolyzate and peptides to induce oral tolerance to native beta lactoglobulin and to identify tolerogenic beta-lactoglobulin peptides with low allergenicity. METHODS: beta-Lactoglobulin was hydrolyzed by trypsin and fractionated by ion exchange chromatography. Peptide enrichment of fractions was evaluated. Balb/c mice were fed beta-lactoglobulin hydrolyzate or fractions by single gavage at day 1. Five days later animals were challenged intraperitoneally with native beta-lactoglobulin. At day 27 delayed-type hypersensitivity was performed. Twenty-four hours later mice were bled, and intestinal contents and spleens were collected. Oral tolerance was measured by titrating specific IgE in sera and intestinal samples. Specific T-cell responses were analyzed by splenocyte proliferation. Antigenicity of hydrolyzate and fractions was evaluated by specific ELISA inhibition. RESULTS: Mice fed either beta-lactoglobulin hydrolyzate or 2 fractions of the hydrolyzate were tolerized against beta lactoglobulin. Specific serum and intestinal IgE were suppressed. Delayed-type hypersensitivity and proliferative responses were inhibited. One tolerogenic fraction was found to be 50 times less antigenic than the total beta lactoglobulin hydrolyzate was. CONCLUSION: These findings support the strategy of inducing oral tolerance in "at-risk" patients by means of tolerogenic cow's milk peptides or hydrolyzate. PMID- 10719303 TI - CD40 and adenosine A2 receptor agonist-cyclic adenosine monophosphate rescue B cell antigen receptor-induced apoptosis through independent pathways and converge to prevent caspase activation. AB - BACKGROUND: Antigen receptor ligation induces apoptosis of B lymphocytes, but the molecular mechanisms underlying induction of apoptosis remain unclear, although the growing family of IL-1beta-converting enzyme cysteine proteases (caspases) are recognized to be major effectors of cellular death. OBJECTIVE: We sought to delineate and compare the rescue of B-cell apoptosis through CD40 ligand-CD40 interaction and cyclic adenosine monophosphate (cAMP)-dependent protein kinase A in human B cells. METHODS: By using tonsillar B cells and the B-lymphoblastoid cell line Ramos, rescue from B-cell apoptosis was compared, as were signaling pathways after activation of cells through CD40 and the adenosine A2 receptor. RESULTS: Both CD40 ligand-CD40 interaction and activation of intracellular cAMP rescue B cells from apoptosis after antigen receptor ligation. Although these pathways do not overlap, they converge by preventing the anti-IgM-induced activation of CPP32 (caspase 3), a member of the IL-1beta-converting enzyme protease family. CONCLUSION: These data indicate that the cAMP-protein kinase A dependent and CD40-signaling pathways regulate B-cell survival and converge at a common point, the inhibition of antigen receptor-induced activation of caspases. PMID- 10719304 TI - Ganglioside GQ1b enhances anti-double-stranded DNA antibody and IgG production of PBMCs from patients with systemic lupus erythematosus. AB - BACKGROUND: Previously, we reported that ganglioside GQ1b greatly enhanced spontaneous immunoglobulin production in vitro by PBMCs from normal human subjects. OBJECTIVE: We examined in vitro effects of GQ1b on anti-double-stranded DNA (anti-dsDNA) antibody production by PBMCs from patients with systemic lupus erythematosus (SLE). METHODS: PBMCs from patients with SLE were cultured with GQ1b. IgG anti-dsDNA antibody, total IgG, and cytokine amounts in the culture supernatants and protein kinase C (PKC) activity of T cells were measured by using ELISA. RESULTS: GQ1b enhanced both anti-dsDNA and total IgG production of PBMCs from patients with SLE who were seropositive for anti-dsDNA. Among the seropositive patients, the active patients were more responsive to GQ1b in anti dsDNA production than the inactive patients. GQ1b also enhanced total IgG production of PBMCs from patients with SLE who were seronegative for anti-dsDNA but did not induce their anti-dsDNA production. In contrast to PBMCs, GQ1b did not affect the antibody production either of purified CD5(+) or of CD5(-) B cells. Anti-IL-6 or anti-IL-10 antibody each partially blocked the GQ1b-induced enhancement of antibody production in PBMCs, and the addition of both antibodies completely blocked the enhancement. GQ1b increased IL-6 and IL-10 production of T cells. The supernatant from GQ1b-treated T cells enhanced antibody production both of CD5(+) and of CD5(-) B cells to a greater extent than that from medium treated T cells. Exogenous IL-6 and IL-10 additively increased the antibody production both of CD5(+) and CD5(-) B cells. GQ1b-induced increases in IL-6 and IL-10 production of T cells were both blocked by PKC inhibitors, calphostin C and staurosporine. GQ1b enhanced PKC activity of T cells. CONCLUSION: These results suggest that GQ1b may polyclonally increase the production of IgG, including IgG anti-dsDNA antibody, in PBMCs from patients with SLE by promoting IL-6 and IL-10 production of T cells through the enhancement of their PKC activity. PMID- 10719305 TI - Detection of C1 inhibitor mutations in patients with hereditary angioedema. AB - BACKGROUND: Hereditary angioedema (HAE) results from a deficiency in the functional level of C1 inhibitor caused by mutations in the C1 inhibitor gene. The mutations responsible for HAE have been shown to be heterogeneous. OBJECTIVE: Because the identification of C1 inhibitor mutations may depend, in part, on the technique used to screen for mutations, we screened the entire C1 inhibitor coding region to identify mutations in a cohort of patients with HAE. METHODS: By using single-stranded conformational polymorphism analysis, 24 subjects with HAE from 16 different kindreds were screened for C1 inhibitor polymorphisms. C1 inhibitor mutations were identified by sequencing the exons containing identified polymorphisms. RESULTS: All 24 subjects with HAE had identifiable polymorphisms, involving exons 2, 3, 4, 5, or 8. Fourteen different C1 inhibitor mutations were identified: 8 missense, 1 nonsense, 4 frameshift, and 1 small deletion mutations. No large deletions or duplications were found. Nine of the 14 mutations represent newly recognized C1 inhibitor mutations, 6 of which involve exon 4. CONCLUSIONS: Single-stranded conformational polymorphism is an effective approach for identifying new mutations in HAE. Elucidation of the range of C1 inhibitor mutations causing HAE is important for both defining which residues are required for C1 inhibitor secretion or function and providing the basis for future studies to define the relationship between the C1 inhibitor genotype and disease severity. PMID- 10719306 TI - Trichophyton-specific IgE in patients with dermatophytosis is not associated with aeroallergen sensitivity. AB - BACKGROUND: It has been proposed that Trichophyton infection is associated with atopy and allergy. OBJECTIVES: Our purpose was (1) to confirm whether atopy predisposes to chronic dermatophytosis and (2) to investigate whether Trichophyton infection induces atopic disease. METHODS: Patients attending dermatology clinics and suspected of having dermatomycosis underwent in a prospective manner fungal culture and Trichophyton and inhalant skin tests, and blood serum was collected for total IgE and Trichophyton radioallergosorbent testing. Personal and family history of atopic diseases was also investigated. RESULTS: According to mycologic culture, atopic history, and inhalant skin test results, patients were classified into 4 groups: (1) atopy plus mycosis (n = 28), (2) atopy (n = 26), (3) mycosis (n = 35), and (4) no atopy, no mycosis (n = 33). Patients with active mycosis (groups 1 and 3) demonstrated significantly increased positivity of Trichophyton skin tests compared with patients without fungal infection (groups 2 and 4), regardless of their atopic status, whereas atopic patients (those in groups 1 and 2) had significantly increased levels of total serum IgE compared with nonatopic subjects. Trichophytosis was not more prevalent in atopic than in nonatopic subjects, and atopic diseases were not more frequent in culture-positive than in culture-negative patients. CONCLUSIONS: Our results indicate that Trichophyton -specific IgE is observed in patients with trichophytosis regardless of atopy. PMID- 10719307 TI - Chronic urticaria serum induces histamine release, leukotriene production, and basophil CD63 surface expression--inhibitory effects ofanti-inflammatory drugs. AB - BACKGROUND: A role of potential histamine-releasing autoantibodies against the high-affinity IgE receptor on the surface of basophils and mast cells is discussed in the pathogenesis of chronic urticaria. This so-called autoimmune urticaria may be diagnosed by a positive intracutaneous autologous serum skin test, which is found in about 30% of patients with chronic urticaria. OBJECTIVE: Our purpose was, first, to compare the effect of complement-inactivated sera of 20 patients with chronic urticaria and positive autologous serum skin tests, 20 patients with chronic urticaria and negative skin tests, and 20 control subjects without chronic urticaria (10 atopic and 10 nonatopic subjects) and, second, to analyze the effect of anti-inflammatory drugs on the serum activity. METHODS: The following assay systems were used: release of histamine in whole blood samples, surface expression of the activation marker CD63 on basophils, and sulfidoleukotriene de novo production in leukocyte suspensions. Whole blood, basophils, and leukocyte suspensions were obtained from a nonatopic and an atopic donor. RESULTS: Sera of patients with autologous serum skin test positive chronic urticaria resulted not only in significantly increased histamine release compared with skin test-negative chronic urticaria sera but also in a significant higher induction of basophil CD63 surface expression and sulfidoleukotriene de novo production. However, serum activity was neither characteristic for chronic urticaria nor for chronic urticaria with a positive autologous serum skin test. Preincubation with dapsone, chloroquine, and lidocaine dose dependently resulted in a significant reduction of all histamine release, CD63 expression, and sulfidoleukotriene production. In addition, mizolastine was able to inhibit serum induced sulfidoleukotriene production. CONCLUSION: Further studies investigating the in vivo effect of these drugs will have to clarify their role in the management of the subset of patients with chronic urticaria demonstrating serum induced inflammatory effects. PMID- 10719308 TI - In situ localization of latex allergens in 3 different brands of latex gloves by means of immunogold field emission scanning and transmission electron microscopy. AB - BACKGROUND: Latex proteins represent relevant allergens, particularly for those persons who are frequently exposed to latex products (eg, health care workers and patients with chronic disorders). Although several latex allergens have been characterized by biochemical and molecular biologic techniques, little information is available concerning the in situ localization of allergenic proteins in latex products. OBJECTIVE: The objective of the present study was the in situ localization of latex allergens. METHODS: Serum IgE from patients with latex allergy reacting with a broad range (5-200 kd) of latex allergens was used for the in situ localization of latex allergens. One surgical and 2 examination latex glove brands were investigated by using immunogold field emission scanning and transmission electron microscopy. RESULTS: Allergens were detected on the inner and outer surface of the gloves, particularly near the edges, crests, or folds of the bleb-like structures visible on the surface of the latex material at high magnifications. In ultrathin cross-sections, latex allergens were found throughout the sections. CONCLUSIONS: Latex allergens were localized on the outer and inner surface but also in the interior of latex gloves. The occurrence of latex allergens on the surface of latex products may be related to their potential to induce local reactions and, perhaps, to sensitize individuals by means of contact. PMID- 10719309 TI - Sensitization to soybean hull allergens in subjects exposed to different levels of soybean dust inhalation in Argentina. AB - BACKGROUND: Soybean hulls (SHs) have been identified as the source of aeroallergens responsible for soybean asthma outbreaks. However, the prevalence of sensitization to SH allergens in subjects from Argentina, a country where soybeans are produced, is unknown. OBJECTIVE: The purpose of this study was to determine the prevalence of sensitization to SH by in vivo and in vitro tests in subjects with asthma or allergic rhinitis and in control subjects from Argentina who have been exposed to different levels of soybean dust inhalation (SDI). METHODS: Exposure to SDI is defined as follows: (1) direct = occupational, (2) indirect = proximity to soybean fields or grain elevators, and (3) urban = urbanized areas without a known source of SDI. Two groups were studied. Group 1 consisted of 365 subjects with asthma or allergic rhinitis and group 2 (control group) of 50 healthy individuals. Subjects from both groups were classified according to their exposure to SDI. All subjects completed standard questionnaires. Prick skin tests (STs) with an SH extract and with common allergens were performed on all subjects. Specific IgE and IgG4 to SH were measured in sera of 51 of 56 subjects from group 1 who had a positive ST to SH and in all sera from group 2. RESULTS: Fifty-six (15.3%) subjects from group 1 and no subjects from group 2 had a positive ST to SH (wheal SH/wheal histamine >/=0.5). In group 1, positive STs to SH were 38.7%, 20.3%, and 8.2% in subjects with direct, indirect, and urban exposures, respectively (P <.001). Monosensitization to SH is absent in all subjects from group 1. The percent of subjects with positive STs to mites, pollen, and molds was highest in those with a positive ST to SH versus those with a negative ST to SH (P <.01). Asthmatic patients with a positive ST to SH, compared with those exclusively sensitized to mites, had a higher frequency of daily or weekly symptoms (59.4% vs 25.7%, respectively, P <.001) and a higher percent of glucocorticoid dependence (52.8% vs 34%, respectively, P <.01). Percent positive IgE in group 1 and group 2 were 39.2% and 10% (P <.001) and percents positive IgG4 are 27.4% and 12%, respectively (not significant). In subjects from group 1 and group 2 with direct exposure percents positive IgE are 58.3% and 13.3% (P < .001) and percents positive IgG4 were 75% and 20%, respectively (P < .02). IgG4 in group 1 was significantly higher in subjects with direct exposure compared with subjects with indirect or urban exposure. CONCLUSION: This study demonstrated that there was (1) a high prevalence of sensitivity to SH in subjects with asthma or allergic rhinitis from Argentina and (2) an association between sensitivity to SH and severity of asthma and level of exposure to SDI. PMID- 10719310 TI - Hazelnut allergy: a double-blind, placebo-controlled food challenge multicenter study. AB - BACKGROUND: Tree nuts are a common cause of food allergy in Europe. However, few studies deal with real food allergy to hazelnuts in subjects believed to be allergic to this food. OBJECTIVE: We sought to select subjects with a history of allergic reactions on ingestion of hazelnut and determine how many of these have true allergy by means of the double-blind, placebo-controlled food challenge (DBPCFC). METHODS: Eighty-six subjects with a history of symptoms after hazelnut ingestion were recruited from 3 allergy centers (Milan, Zurich, and Copenhagen). All subjects underwent skin prick tests (SPTs) with aeroallergens and hazelnut, as well as having their specific hazelnut IgE levels determined. Diagnosis of clinical relevant food allergy was made on the basis of the DBPCFC. RESULTS: Sixty-seven (77.9%) of 86 subjects had a positive DBPCFC result; 8 were placebo responders, and 11 were nonresponders. Of the 11 nonresponders, 4 had positive open-challenge test results. Of the DBPCFC-positive subjects, 87% also had positive skin test responses to birch pollen extract. Specific IgE determination for hazelnut (positive CAP response >/=0.7 kU/L [ie, class 2]) showed a sensitivity of 0.75, a positive predictive value (PPV) of 0.92, a specificity of 0.16, and a negative predictive value (NPV) of 0.05. Skin tests with commercial hazelnut extract produced a sensitivity of 0.89, a PPV of 0.92, a specificity of 0.05, and an NPV of 0.05. Skin tests with natural food produced a sensitivity of 0.88, a PPV of 0.94, a specificity of 0.27, and an NPV of 0.15. CONCLUSION: This study shows that hazelnut is an allergenic source that can cause real food allergy, as confirmed by DBPCFC. Skin and IgE tests demonstrated reasonable sensitivity and PPV but a very low specificity and NPV, thus implying that these should not be used to validate the diagnosis of food allergy to hazelnut. PMID- 10719311 TI - Dose-response in double-blind, placebo-controlled oral food challenges in children with atopic dermatitis. AB - BACKGROUND: Double-blind, placebo-controlled oral food challenges (DBPCFCs) are considered the "gold standard" for diagnosing food hypersensitivity, but the dose that elicits positive challenges, or determinants that may predict dose-response relationships, have not been reported. OBJECTIVE: Our purpose was to determine the quantity of food that elicits reactions during DBPCFCs and to evaluate parameters that may predict the provocative dose and severity of reaction. METHODS: We reviewed challenge data for all positive challenges to 6 common allergenic foods in children with atopic dermatitis evaluated for food allergy over a 13-year period. Challenge food was generally administered in 6 doses at 10 to 15-minute intervals beginning with 400 to 500 mg and completing with a total of 8 to 10 g of food. An open feeding of a larger portion followed negative challenges. At the physician's discretion, a lower starting dose was occasionally used (100 mg, 250 mg). Food-specific IgE antibody concentrations (radioallergosorbent test [RAST]) were determined on stored sera of 20% of the challenges selected randomly and 99.6% had prick skin tests (PSTs) performed to the challenged food. RESULTS: A total of 196 children (45% male; median age 5 y 9 mo; atopic dermatitis 98%, asthma 62%) had 513 positive challenges distributed as follows: egg 267, milk 117, soy 53, wheat 40, peanut 24, fish 12. The percentage of children reacting at the first dose (500 mg or less) was as follows: egg 49%, milk 55%, soy 28%, wheat 25%, peanut 26%, and fish 17%. Twenty-six milk challenges and 22 egg challenges were positive at a first dose of 250 mg; 3 milk challenges and 7 egg challenges were positive at a first dose of 100 mg. Eleven percent of the reactions that occurred on the first dose were severe. The percentage reacting after the final dose of the DBPCFC (or during open challenge) were egg 11%, milk 12%, soy 19%, wheat 12.5%, peanut 8.7%, and fish 25%. There was not a strong correlation between PST absolute wheal size or score (adjusted for histamine controls) and dose at reaction or severity of reaction (R(s) range 0.22 to 0.39 for particular foods). Serum concentration of food-specific IgE did not correlate well with the dose causing a reaction or with severity (R(s) range 0.40 to 0.55 for particular foods). CONCLUSIONS: This food-allergic population may react to as little as 100 mg of food, possibly less, and the dose causing a reaction and the severity of reaction is not predicted by PST or RAST. Lower doses (100 mg or less) should be investigated for their appropriateness in initiating DBPCFCs. PMID- 10719312 TI - Anaphylactic reactions to raw eggs after negative challenges with cooked eggs. PMID- 10719313 TI - Trimethoprim-sulfamethoxazole-graded challenge in HIV-infected patients: long term follow-up regarding efficacy and safety. PMID- 10719315 TI - Evaluation of cat skin test sensitivity. PMID- 10719317 TI - First-trimester diagnosis of Morquio disease type A. AB - Since the introduction in 1990 of a novel fluorogenic substrate for galactose-6 sulphate sulphatase we have used this substrate for prospective prenatal diagnosis in 10 pregnancies at risk for Morquio disease type A. Chorionic villi were analysed in five cases. The results indicated an affected fetus in one pregnancy which represents the first case of first-trimester diagnosis of this disorder; heterozygosity was demonstrated in two cases. Following amniocentesis, two affected fetuses and one heterozygote were diagnosed. The results of the present prospective prenatal analyses confirm our previous retrospective studies and demonstrate the reliability and convenience of the 4-methylumbelliferyl substrate. PMID- 10719318 TI - Co-variables in first trimester maternal serum screening. AB - The objective of this study was to determined the influence of maternal weight, maternal smoking habits, gravidity, parity and fetal gender on the level of maternal serum marker used in first trimester screening for Down syndrome. A total of 2449 singleton unaffected pregnancies from two centres were studied. Maternal serum free beta-human chorionic gonadotrophin (hCG) and alpha fetoprotein (AFP) concentrations had been measured in all pregnancies, and pregnancy associated plasma protein (PAPP)-A levels had been measured in 924. All results were expressed as multiples of the gestation specific median (MoM) values after regression, using each centre's own medians. Information on maternal weight was available in 2259 pregnancies, on self-reported current cigarette smoking in 1364 (of whom 117 (8.6%) were smokers), on gravidity in 1371, parity in 1303 and fetal gender in 253. All three markers showed a statistically significant negative association with maternal weight (p<0.0005) and in the subsequent analyses MoM values were weight adjusted using standard methods. The median PAPP A level in smokers was 0.81 MoM, a significant reduction (p<0.005); free beta-hCG was also reduced (median 0.89 MoM) but not significantly (p=0.17), and AFP was unaltered. The median AFP level in primagravidas was highly significantly greater than that in gravid women (p<0.0005). In PAPP-A the reverse effect was seen but it did not reach statistical significance (p=0.15) and there was no effect for free beta-hCG. Results of a similar magnitude and direction were found for parity. The median level of free beta-hCG was higher (p=0.0005), and the median AFP lower in female pregnancies. Maternal weight and, for PAPP-A, maternal smoking are important first trimester screening co-variables. Gravidity, parity and fetal gender also seem to influence one or more first trimester markers. PMID- 10719319 TI - Fetoplacental chromosomal discrepancy. AB - Chorionic villus sampling is now an acceptable alternative to second trimester amniocentesis. Several reports have raised concerns about the occurrence of discrepancies between the chorionic villi and fetal chromosomal constitution, which adds multiple diagnostic complexities to the process of prenatal genetic counselling. We report on a series of 26 cases in which fetoplacental discrepancies have occurred. The chromosomal aberration was exclusively confined to the placenta in 21 cases. Twice the identical aberration was also observed in amniotic fluid with variant range of aneuploidy. The chromosomal abnormality was found in amniotic fluid and fetal blood samples in two cases, and was absent in chorionic villi. One case had very unusual cytogenetic findings as two different non-mosaic chromosomal abnormalities were identified separately in the placenta and amniocytes. Among the 21 gestations with confined placental abnormal karyotype, three cases of intrauterine growth retardation were identified. Of six cases evaluated for uniparental disomy, four demonstrated biparental inheritance. These findings support a positive correlation between placental aneuploidy and abnormal fetal development. They also emphasize the importance of further DNA analysis whenever discrepant karyotype findings between the placenta and amniocytes are identified. PMID- 10719320 TI - Prenatal diagnosis of beta-thalassaemia and other haemoglobinopathies in India. AB - This paper reports prenatal diagnosis of 787 fetuses of beta-thalassaemia and other haemoglobinopathies in Indian high-risk communities. DNA based diagnosis was offered in the first, as well as the second trimester, in 489 pregnancies (with five twins) on fetal tissues such as chorionic villus (CV) and amniocytes using the amplification refractory mutation system (ARMS) and restriction fragment length polymorphism (RFLP) techniques. Two hundred and ninety-two women (with one twin), who either presented late in the second trimester or whose DNA diagnosis was not informative, were offered prenatal diagnosis using globin chain synthesis (GCS) on fetal blood cells. Maternal contamination of fetal DNA was ruled out by variable number tandem repeat (VNTR) analysis using sites in four different genes (Apo-B, D1S-80, Ig-JH and Ha-ras), while contamination of fetal blood was checked by a particle size distribution channelyzer. Using both techniques we were able to offer complete diagnosis in 99.8% cases. Out of 494 fetuses tested by DNA analysis, 135 were found to be normal, 201 were carriers, whereas 146 were affected. Out of 293 fetuses analysed by GCS, 215 were unaffected and 71 were affected. In this study, both fetuses were tested in twin pregnancies, of which three required selective termination of one fetus. Because of social, religious taboos and family influences, genetic counselling was found to be an important guideline for couples selecting options for prenatal diagnosis. Our experience suggests that because of late presentation by many couples to the diagnostic centres, in developing countries like India, both the techniques of DNA analysis and GCS should be made available at major referral centres for maximum benefit to couples. PMID- 10719321 TI - Iniencephaly: prenatal diagnosis and management. AB - Iniencephaly is a rare malformation characterized by the triad of occipital bone defect, cervical dysraphism and fixed retroflexion of the fetal head. Because of its almost invariable lethal prognosis, termination of pregnancy is commonplace when this condition is diagnosed before viability. In this report we describe eight cases of iniencephaly prenatally diagnosed by ultrasound between 18 and 28 weeks of gestation and discuss the subsequent obstetric management in a country where elective abortion is illegal. Prenatal karyotyping was performed in seven cases, revealing a normal complement in all fetuses. One pregnancy miscarried at 24 weeks. Uneventful vaginal delivery was accomplished in six of the remaining seven cases, one delivered spontaneously at 29 weeks and five were induced between 28-32 weeks due to increasing polyhydramnios. In the remaining case the pregnancy progressed to 35 weeks, at which time spontaneous labour began and an emergency Caesarean section was performed because of malpresentation. There were no survivors in this series. We conclude that, in countries were elective abortion is not allowed, women carrying an iniencephalic fetus may benefit from preterm induction of labour in order to avoid labour dystocia, maternal trauma during delivery and the risks of a Caesarean section. PMID- 10719322 TI - A study of mild fetal pyelectasia - outcome and proposed strategy of management. AB - Mild fetal pyelectasia, defined as a renal pelvic anteroposterior (AP) diameter of 4-10 mm, has become a frequent finding on fetal ultrasonography. The natural history of such dilatation is unclear, resulting in confusion as to appropriate postnatal investigation and management. The aim of this study was to examine the urinary tract outcome of a series of infants with mild fetal pyelectasia demonstrated on routine morphology ultrasonogram between 16 and 21 weeks' gestation. Of the 37 cases identified, 13 (35%) went on to require medical or surgical intervention for significant urinary tract anomalies. These anomalies included pelvi-ureteric junction obstruction, dysplastic kidney, vesicoureteric reflux and posterior urethral valves. On initial scan all cases had an AP diameter of 4-8 mm and did not predict those infants who would go on to require intervention. An AP diameter of greater than 7 mm on repeat scans performed after 27 weeks' gestation had a positive predictive value of 0.92 and a negative predictive value of 0.76 for significant urinary tract anomaly requiring intervention. The specificity was 0. 94 and sensitivity 0.70. A protocol of one repeat antenatal ultrasound at 28-34 weeks' gestation would be able to identify those infants who would require postnatal investigation, using a measurement of >/=7 mm. The fetus with a normal repeat ultrasound would not require postnatal follow-up. PMID- 10719323 TI - Critical reappraisal of the utility of sonographic fetal femur length in the prediction of trisomy 21. AB - Measurement of femur length (FL) has been advocated as part of a genetic sonogram for the prediction of Down syndrome (DS). However its predictive ability has been inconsistent. We have studied the diagnostic value of this sonographic parameter in a prospective cohort of women with singleton gestations undergoing genetic sonogram between 14 and 22 weeks because of advanced maternal age or family history of aneuploidies. Genetic sonograms were performed at a mean gestational age of 17.0 weeks (range 14-22). DS was diagnosed in 30 fetuses, while 888 were euploid. Mean+/-SD observed/expected (O/E) values of FL (1.00+/-0.10 versus 0.97+/-0.01, p=0.07) were not significantly different between euploid and DS fetuses. Comparison of the regression equations of FL versus biparietal diameter revealed that while the intercepts were not significantly different between euploid and DS fetuses, the difference in slopes reached significance (p=0.04) suggesting that the predictive ability of FL may increase with advancing gestational age. In addition, a MEDLINE search (National Library of Medicine) was conducted for articles published between 1985 and 1998 on fetal femur length in the prediction of trisomy 21. Review of the published literature on the subject suggests that FL is not a consistent or reliable sonographic predictor of DS. Published thresholds of FL should not be used outside of the Institution from which they originated, and each Institution should establish whether this parameter has predictive ability in its own population. PMID- 10719324 TI - The clinical application of interphase FISH in prenatal diagnosis. AB - Fluorescence in situ hybridization (FISH) for five chromosomes (13, 18, 21, X and Y) detected 87 of 107 (81%) of the chromosome aberrations identified by conventional chromosome analysis applied to fetal interphase cells obtained by chorionic villus sampling or amniocentesis. The choice of FISH was solely determined by prospective parents after formal genetic counselling concerning the advantages and disadvantages of FISH analysis. Excluding known familial chromosome aberrations, if FISH analysis revealed normal signals, there was an overall residual risk of 1 in 149 for an undetectable chromosome aberration. This risk varied according to the indication for prenatal diagnosis: 1 in 177 for women of advanced maternal age; 1 in 60 for women at increased risk for Down syndrome based on maternal serum screening; and, 1 in 43 for women whose ultrasound examination revealed fetal anomalies. There were 20 cases of discordance between the FISH results and standard karyotype analysis: four were the outcome of a failure to apply the appropriate FISH probe; 16 were not detectable by the available FISH probes. Of these 16, nine were chromosome abnormalities with clinical significance and seven were familial. If FISH is to become a standard part of prenatal genetic diagnosis, genetic counselling that is sensitive to patient health needs must be based on accurate information about the biological and obstetrical implications of the results of FISH analysis. PMID- 10719325 TI - Maternal midtrimester serum AFP and free beta-hCG levels in in vitro fertilization twin pregnancies. AB - We aimed to compare the levels of alpha-fetoprotein (AFP) and free beta-human chorionic gonadotrophin (beta-hCG) levels as multiples of the median (MoM) values between spontaneous and in vitro fertilized (IVF) twin pregnancies. The control group of spontaneous singleton pregnancies was used for calculating the gestational age specific median levels of the values. Within a cohort of 19 310 pregnancies, 145 twin pregnancies were identified. The data were collected from Down syndrome (DS) screening programmes in four University catchment areas in Finland between 1994-98. Maternal midtrimester serum marker levels were measured across gestational weeks 14-18. There were no fetal chromosome anomalies in either of the twin groups or the singleton group. Serum AFP of 145 and beta-hCG values of 39 spontaneous twin pregnancies were compared to the values of 6548 singleton pregnancies. In IVF twins 30 AFP and 29 beta-hCG values were compared to the levels of the control group. Both AFP and beta-hCG values were twice as high in the spontaneous twin pregnancies (medians 2.18 and 1.83 MoM respectively) as in the singleton group (medians 1.00 and 1.00 MoM respectively). In IVF twin pregnancies beta-hCG levels were higher (median 2.20 MoM) than in spontaneous twins (p=0.08), whereas no significant difference was found in AFP levels (2.30 MoM). In conclusion, the higher levels of beta-hCG levels in IVF twin pregnancies should be considered in DS screening to avoid high false positive rates. PMID- 10719326 TI - Cordocentesis at 16-24 weeks of gestation: experience of 1,320 cases. AB - The objective of this study was to assess the safety and efficacy of diagnostic cordocentesis at midpregnancy. 1,320 singleton pregnancies with no obvious congenital anomalies, a gestational age of 16-24 weeks, and proper indications underwent cordocentesis using the freehand technique. The results of each procedure was prospectively collected and subsequently analysed for the results and pregnancy outcomes. The mean maternal age was 31.1 years and the mean gestational age at the time of cordocentesis was 19.8 weeks. The most common indication was the risk of severe thalassaemia syndrome (69.8%) and was followed by rapid karyotyping. Of 1,320 cordocenteses, 1,281 (97%) were done successfully at the first attempt. The mean duration of the procedure was 10.5 min and was significantly longer in the first 50 cases of practice for each operator. The maternal blood contamination rate was higher when the cord insertion was targeted. The procedure-related complications included transient bleeding at puncture site (20.2%), transient fetal bradycardia (4. 3%), chorioamnionitis (two cases), and cord haematoma (one case). Of 1,281 successful cases, 184 fetuses had severe disease. The total fetal loss rate was 3.2% and the procedure-related loss was 1%. The other obstetric complications were comparable with those in the general population. We conclude that cordocentesis at midpregnancy is a useful, relatively safe, and effective procedure for prenatal diagnosis. PMID- 10719327 TI - Prenatal control of severe thalassaemia: Chiang Mai strategy. AB - Prenatal diagnosis of severe thalassaemia is conventionally diagnosed by fetal DNA analysis but it can not be widely used due to its drawbacks of high cost and technical effort. This prospective study describes a new prenatal strategy in preventing severe thalassaemia by a more simple and inexpensive way. The strategy included: (1) genetic counselling; (2) identification of pregnancies at risk by retrospective screening (history of known risk) and prospective screening for asymptomatic women; (3) cordocentesis at 16-22 weeks' gestation; (4) fetal blood analysis with high performance liquid chromatography (HPLC); (5) termination of affected pregnancy. The prospective screening consisted of 2 min osmotic fragility (OF) and HbE screening test in women with no risk, and testing the husbands of the women with a positive result. If both of the couple had a positive result, the diagnostic test (HbA(2) level and PCR alpha-thal 1) for the carrier was needed. A pregnancy in which both of the couple were carriers was considered at risk. This strategy identified 181 and 108 couples at risk by prospective (from 7954 pregnancies) and retrospective screening, respectively. Two hundred and forty-two underwent cordocentesis, 108 from retrospective screening and 134 from prospective screening, and 62 were proven to have severe thalassaemia (29 and 33 in retrospective and prospective screening, respectively). The strategy identified nearly all, if not all, fetuses with severe thalassaemia without false positives among the screened couples. In conclusion, the strategy proves to be highly effective in the control of severe thalassaemia. PMID- 10719328 TI - Female gamete segregation in two carriers of translocations involving 2q and 14q. AB - FISH, using a combination of whole-chromosome painting and telomeric probes, was used to study the gamete segregation of two female carriers of translocations involving the same chromosome arms, 2q and 14q. Preimplantation genetic diagnosis of the first polar bodies of these oocytes permitted selecting normal embryos for replacement. PMID- 10719329 TI - Fetal demise with Smith-Lemli-Opitz syndrome confirmed by tissue sterol analysis and the absence of measurable 7-dehydrocholesterol Delta(7)-reductase activity in chorionic villi. AB - Smith-Lemli-Opitz syndrome (SLOS), an autosomal recessive condition with multiple malformations, mental retardation, and growth failure, results from markedly reduced activity of the final enzyme in the cholesterol biosynthetic pathway, 7 dehydrocholesterol Delta(7)-reductase (DHCR7). We diagnosed SLOS in a fetus following intrauterine demise at 32 weeks' gestation. Chorionic villus (CV) sampling had been performed at 30 weeks because oligohydramnios and atrioventricular septal defect were noted on fetal ultrasound. On fetal post mortem examination, a midline U-shaped soft palate cleft, micrognathia, postaxial polydactyly of the fingers with single transverse palmar creases bilaterally, and cutaneous syndactyly of toes two-three bilaterally suggested SLOS. We hypothesized that SLOS could be confirmed by analysis of tissue sterols despite extensive autolysis, and by measurement of enzyme activity in CV cells. Measurement of DHCR7 activity in CV cells was undertaken using ergosterol as a substrate. CV cells were unable to convert any ergosterol to brassicasterol after a 72 h incubation period while control CV cells reduced 12.6-71.8% of ergosterol to brassciasterol in a 72 h period. SLOS was confirmed by measurement of elevated 7-dehydrocholesterol (7-DHC) in the CV cells. Measurements of sterols were made in multiple fetal tissues. All tissues analysed showed elevated 7-DHC with markedly increased 7-DHC/cholesterol ratios. PMID- 10719330 TI - Learning in medicine: chorionic villus sampling. AB - Operator experience is considered to influence the safety and success of medical procedures. We performed a retrospective survey to assess learning curves in chorionic villus sampling (CVS). Data of 2081 consecutive women, in whom CVS was carried out in a tertiary care university hospital for prenatal diagnosis, were available for analysis. Endpoints of the analysis were fetal loss, failure of the procedure and need for several needle insertions. Frequencies of each endpoint were calculated and plotted for consecutive series of 50 samplings per operator. Logistic regression analysis was used to quantify the effect of operator experience. We observed a statistically significant learning effect for the unit as a whole as assessed by fetal loss figures and the same tendency for need to do several insertions. We did not observe a (collective) learning effect for failure of the procedure. Individual operators showed a significant learning profile for some endpoints. The individual learning profile predominantly depended on previous experience in amniocentesis. Our study substantiates the presence of a learning curve for CVS. This supports the view that centralization and restriction of CVS to a limited number of experienced operators within centres, is likely to have a positive influence on safety and success of the procedure. PMID- 10719331 TI - Genetic amniocentesis in women 20-34 years old: associated risks. AB - The aim of this retrospective controlled study is to evaluate the impact of predisposing factors on amniocentesis-related fetal loss. It comprises 3910 consecutive cases of women, aged 20-34 years, who had genetic amniocentesis during the years 1992-97 (study group). The control group comprised 5324 women under 35, at low risk for Down syndrome, during the same period. The fetal losses in both groups were analysed, in respect of: (a) maternal historical conditions; and/or (b) bleeding during current pregnancy. The leading indication for amniocentesis in women 20-34 years was maternal anxiety, mainly for marginal age (33-34 years), which accounted for a remarkable 34.4% of the study group. Total fetal loss rate up to the 28th week was 2.1% in the study group versus 1. 5% in controls. A history of previous spontaneous or induced abortions, as well as bleeding during the current pregnancy, was associated with a substantial rise of fetal loss in both groups. In cases with no predisposing factors, the added fetal loss rate was 0.03%. Previous abortions and bleeding during the current pregnancy are associated with the most fetal losses after amniocentesis. In the absence of these, the added fetal loss rate (0.03%) is non-significant. PMID- 10719332 TI - Oesophageal atresia. PMID- 10719333 TI - Prenatal diagnosis of Apert syndrome with widely separated cranial sutures. AB - A 35-year-old Taiwanese woman visited us for prenatal evaluation in the 20th week of pregnancy. Both clinical observation of the mother and analysis of maternal DNA indicated a diagnosis of Apert syndrome. Sonographic examination of the fetus demonstrated findings that were consistent with this diagnosis; however, no prematurely fused cranial sutures were observed. The pregnancy was terminated after genetic counselling and fetal DNA analysis showed the same mutation as found in the mother. Autopsy of the abortus revealed the same findings as were detected by sonography; however, all cranial sutures were widely separated. These findings indicate that, in Apert syndrome, craniosynostosis and syndactyly may develop asynchronously up to 20 weeks of pregnancy. PMID- 10719334 TI - Choice of anticoagulant can influence the analysis using fluorescence in situ hybridization of fetal cells enriched from maternal blood. AB - A notable degree of research attention is being focused on the use of fetal cells enriched from the blood of pregnant women as a non-invasive means of prenatal diagnosis. By using magnetic activated cells sorting (MACS) and fluorescence in situ hybridization (FISH), we have examined the efficacy of enriched fetal cells in determining fetal sex. An unexpected finding of this investigation was that the sensitivity of this analysis was influenced by the anticoagulant used to treat the maternal blood samples. As such, samples treated with heparin showed significantly lower detection rates than samples chelated with EDTA. PMID- 10719335 TI - Prenatal diagnosis of cystic bladder distension secondary to obstructive uropathy. AB - We report the perinatal findings of a huge midline posterior cystic bladder distension secondary to lower urinary tract obstruction and prune-belly syndrome in a male fetus. A 40-year-old woman, gravida 3, para 0, was referred at 21 gestation weeks with sonographic findings of anhydramnios and a fetus with a 9.5 x 6.0 cm intra-abdominal cystic mass containing two chambers. The in utero ultrasound-guided fetal bladder drainage using a single needle aspiration and the ultrasound follow-ups of fetal bladder filling provided a diagnostic aid. This method helped to show the position of the bladder and the cystic bladder mass as well as the status of communication in response to decompression or filling of the fetal bladder. Cytogenetic analysis revealed a 46,XY karyotype. Autopsy showed agenesis of the posterior urethra, prominent megacystis, a cystically distended mass arising from the lower posterior bladder, hydronephrosis, megaureters, and anorectal agenesis with an intestinal blind end adherent to the posterior wall of the uterus. There were no urogenital duplication, hindgut duplication, or urachal abnormalities. The contracted bladder had a full thickness muscular wall with a trigone and two ureteral orifices while the cystically distended bladder did not have any opening and was lined by a very thin wall. Histology of the cystic bladder wall demonstrated typical urothelium, lamina propria and muscularis propria. The pathogenesis and differential diagnosis of cystic bladder distension are discussed. PMID- 10719336 TI - Prenatal diagnosis on fetal cells from maternal peripheral blood in cases of confined placental mosaicism. PMID- 10719337 TI - Comparison of miscarriage rates between early and late amniocentesis. PMID- 10719338 TI - Current awareness in prenatal diagnosis. PMID- 10719353 TI - Gliogenesis in the central nervous system. AB - Multipotential neuroepithelial stem cells are thought to give rise to all the differentiated cells of the central nervous system (CNS). The developmental potential of these multipotent stem cells becomes more restricted as they differentiate into progressively more committed cells and ultimately into mature neurons and glia. In studying gliogenesis, the optic nerve and spinal cord have become invaluable models and the progressive stages of differentiation are being clarified. Multiple classes of glial precursors termed glial restricted precursors (GRP), oligospheres, oligodendrocyte-type2 astrocyte (O-2A) and astrocyte precursor cells (APC) have been identified. Similar classes of precursor cells can be isolated from human neural stem cell cultures and from embryonic stem (ES) cell cultures providing a non-fetal source of such cells. In this review, we discuss gliogenesis, glial stem cells, putative relationships of these cells to each other, factors implicated in gliogenesis, and therapeutic applications of glial precursors. PMID- 10719354 TI - Developmental dynamics of peripheral glia in Drosophila melanogaster. AB - To study the roles of peripheral glia in nervous system development, a thorough characterization of wild type glial development must first be performed. We present a developmental profile of peripheral glia in Drosophila melanogaster that includes glial genesis, developmental morphology, the establishment of transient cellular contacts, migration patterns, and the extent of nerve wrapping in the embryonic and larval stages. In early embryonic development, immature peripheral glia that are born in the CNS seem to be intermediate targets for neurites that are migrating into the periphery. During migration to the PNS, peripheral glia follow the routes of pioneer neurons. The glia preferentially adhere to sensory axonal projections, extending cytoplasmic processes along them such that by the end of embryogenesis peripheral glial coverage of the sensory system is complete. In contrast, significant lengths of motor branch termini are unsheathed in the mature embryo. During larval stages however, peripheral glia further extend and elaborate their cytoplasmic processes until they often reach to the neuromuscular junction. Throughout the embryonic and larval developmental stages, we have also observed a number of similarities of peripheral glia to vertebrate Schwann cells and astrocytes. Peripheral glia seem to have dynamic and diverse roles and their similarities to vertebrate glia suggest that Drosophila may serve as a powerful tool for analysis of glial roles in PNS development in the future. PMID- 10719356 TI - Intermediate filament protein synemin is transiently expressed in a subset of astrocytes during development. AB - Synemin, a developmentally regulated protein first described in muscle cells, has recently been recognized as an intermediate filament (IF) protein. Because IF proteins are invaluable markers of cell origin within the nervous system, we were interested in determining the expression pattern of synemin in the brain. Our results show that, during development of the rat cortex, synemin is expressed only in a subpopulation of astrocytic cells expressing GFAP as well as vimentin and nestin. Unlike GFAP, however, synemin is not expressed in mature astrocytes and, unlike vimentin and nestin, synemin is not present in astrocytic precursors before GFAP expression. Taken together with morphological evidence, the time course of synemin expression, as determined by Western blotting, suggests that synemin is expressed in radial glial cells undergoing morphological transformation into astrocytes. Studies of synemin expression in vitro demonstrate that, early in primary culture, the majority of polygonal astrocytes are derived from synemin(+) radial glial cells. With time in culture, however, polygonal astrocytes either stop expressing synemin or are overgrown by cells not expressing synemin. The unique pattern of synemin expression, both in vivo and in vitro, suggests that the use of synemin as a marker will add a new dimension to studies of astrocytic differentiation. PMID- 10719355 TI - Interleukin-10, interleukin-4, and transforming growth factor-beta differentially regulate lipopolysaccharide-induced production of pro-inflammatory cytokines and nitric oxide in co-cultures of rat astroglial and microglial cells. AB - The pro-inflammatory cytokines interleukin-1beta (IL-1beta), IL-6, tumor necrosis factor-alpha (TNF-alpha), and nitric oxide (NO) can be produced by activated glial cells and play a critical role in various neurological diseases. Using primary co-cultures of rat microglial and astroglial cells, we investigated the effects of the anti-inflammatory cytokines transforming growth factor-beta1 (TGF beta1)/beta2, IL-4, and IL-10 on the production of (pro-) inflammatory mediators after stimulation of the cells with lipopolysaccharide (LPS; 0.1 micrograms/ml, 24 h). IL-10 (10 and 100 ng/ml) and IL-4 (5 and 50 U/ml) suppressed the LPS induced production of NO, IL-6, and TNF-alpha in a dose-dependent manner, whereas TGF-beta1/beta2 (2 and 20 ng/ml) only suppressed NO production. LPS-induced levels of IL-1beta were suppressed by IL-10, but not by IL-4 and TGF-beta1/beta2. Conversely, co-incubation of the glial cells with LPS and antibodies to TGF beta1/beta2 selectively enhanced LPS-induced NO production, whereas co-incubation with antibody to IL-10 enhanced LPS-induced production of all pro-inflammatory cytokines and NO. This finding strongly suggests that effective concentrations of TGF-beta1/beta2 and IL-10 are produced by LPS-stimulated glial cell co-cultures. Production of IL-10 in these co-cultures was confirmed by measurement of rat IL 10 by radioimmunoassay. We conclude that anti-inflammatory cytokines affect the production of inflammatory mediators in LPS-activated co-cultures of microglial and astroglial cells differentially. PMID- 10719357 TI - Astrocytes express specific variants of CaM KII delta and gamma, but not alpha and beta, that determine their cellular localizations. AB - Multiple isoforms of type II Ca(2+)-calmodulin-dependent kinase (CaM KII) are composed of two major neuron-specific subunits, designated alpha and beta, and two less well-characterized subunits that are also expressed in non-neuronal tissues, designated delta and gamma. Regulated expression of these 4 gene products, and several variants produced by alternative splicing, shows temporal and regional specificity and influences intracellular targeting. We used immunoblotting and RT-PCR to analyze subunit and variant expression and distribution in cultured cerebellar astrocytes and neurons, and whole cerebellar cortex from rodent brain. The data indicate that: (i) astrocytes express a single splice variant of delta, namely delta(2); (ii) like neurons, astrocytes express two forms of CaM KII gamma; gamma(B) and gamma(A); (iii) these CaM KII variants are enriched in the supernate fraction in astrocytes, and the particulate fraction in neurons; (iv) unlike neurons, astrocytes do not express detectable levels of alpha or beta subunits or their respective splice variants. The results indicate that neurons and astrocytes express distinct CaM KII subunits and variants that localize to distinct subcellular compartments and, by inference, exert distinct cellular functions. PMID- 10719359 TI - Expression of nitric oxide synthase-2 (NOS-2) in reactive astrocytes of the human glaucomatous optic nerve head. AB - Inducible nitric oxide synthase (NOS-2) is abundantly present in the optic nerve heads of glaucoma patients. To determine the regulation of NOS-2 expression in the glaucomatous optic nerve head, the specific cells that express NOS-2 in the optic nerve heads of patients with primary open-angle glaucoma were studied by immunohistochemical double-labeling of NOS-2 and one of the characteristic cell markers for different cell types. Most of the labeling for NOS-2 was identified in reactive astrocytes that were clustered in the areas of nerve damage in the prelaminar and lamina cribrosa regions of the glaucomatous optic nerve heads. In vitro, the expression of GFAP and NOS-2 by reactive astrocytes of human optic nerve heads was demonstrated by immunocytochemistry and Western blot. In primary cultures of human lamina cribrosa astrocytes, stimulation by interferon-gamma and interleukin-1beta upregulated GFAP and induced expression of NOS-2 protein. At 24, 48 and 72 h of stimulation, NOS-2 appeared first in the Golgi body and then was sent out into the cytoplasm in granules. These results demonstrated that the astrocytes of human optic nerve head are capable of inducing the expression of NOS-2. Reactive astrocytes in the glaucomatous optic nerve heads apparently play an important role in local neurotoxicity to the axons of the retinal ganglion cells by producing excessive nitric oxide in glaucomatous optic neuropathy. PMID- 10719358 TI - Density dependent regulation of human Schwann cell proliferation. AB - Cessation of division is prerequisite for Schwann cell differentiation but regulation of this critical function is poorly understood. Heregulin/forskolin induced growth of human Schwann cells (HSCs) in vitro was found to be strongly regulated by cell density and thus could model some aspects of negative growth regulation in vivo. To better understand this phenomenon, the production of an autocrine growth-inhibitor and the role of contact-inhibition were investigated. The possible involvement of two membrane proteins, contactinhibin (CI) and peripheral myelin protein 22 (PMP22) in regulating growth was studied. Thymidine labeling of HSCs on collagen-coated dishes was inhibited at cell densities less than one tenth of the density at maximal growth-inhibition. Medium from high density cultures did not inhibit the thymidine-labeling of HSCs at low density, a result that argues against the production of a soluble inhibitor. The expression of CI and PMP22 in nerve and HSCs, and the effect of a function-blocking antibody to CI on HSC growth, were determined. CI was detected in fresh nerve by western blotting, and could easily be detected by immunocytochemistry in cultured HSCs by five days and for several weeks thereafter. Twenty-four hour treatment with anti CI antibody did not increase the thymidine-labeling of HSCs at any density but a significant increase in HSC number was observed in cultures treated with anti-CI for 20 days. This increase was not related to decreased cell death. PMP22, unlike other myelin proteins, was not down-regulated after nerve dissociation and by seven days nearly all HSCs were PMP22 positive. These results provide evidence for a contact-mediated mechanism of growth-regulation in HSCs and suggest that CI is involved in this mechanism. PMID- 10719360 TI - Terminal complement complexes concomitantly stimulate proliferation and rescue of Schwann cells from apoptosis. AB - The consequences of sublytic terminal complement complex (TCC) assembly on Schwann cell proliferation and apoptosis were examined by using purified complement proteins (C5*-9) or antibody-sensitized Schwann cells in the presence of a serum that was depleted of the seventh component of complement (C7dHS) and reconstituted with purified C7. Stimulation of cultured Schwann cells with antibody plus 10% C7dHS and C7 or C5*-9 induced DNA synthesis over antibody plus 10% C7dHS alone or in Schwann cells in which C5*-9 insertion was inhibited by heat inactivation, respectively. Cell cycle analysis with propidium iodide showed that, at 24 h, viable Schwann cells in defined medium were synchronized in G1/G0 phase. C5*-9 shifted 64% of these cells into S or G2/M phases in a manner similar to beta-neuregulin (beta-NRG), a known Schwann cell mitogen. Furthermore, antibody with 10% C7dHS and C7 or purified C5*-9 induced proliferation of viable Schwann cells. These effects were mediated by signal-transduction pathways involving p44 ERK1 (extracellular-regulated kinase 1), Gi proteins, and protein kinase C. Culturing in defined medium for 24 h resulted in apoptosis of up to 50% of Schwann cells that was prevented by treatment with beta-NRG or TCC. Sublytic C5*-9 significantly inhibited apoptosis 41% by 24 h, as determined by a terminal deoxyuridine triphosphate-biotin nick end labeling assay, and also decreased annexin-V binding at 4 h. Collectively, these data suggest that sublytic TCC, like beta-NRG, is a potent Schwann cell trophic factor that is capable of stimulating mitogenesis and apoptotic rescue. TCC assembly on Schwann cells during inflammatory demyelination of peripheral nerves may promote survival of mature cells to enhance repair and remyelination processes. PMID- 10719361 TI - Differential regulation of tissue inhibitor of metalloproteinase mRNA expression in response to intracranial injury. AB - Injury to the CNS induces complex cellular and molecular interactions referred to as reactive gliosis. Alterations in the extracellular microenvironment associated with the gliotic response are believed to be the primary cause of regenerative failure of the mature CNS. For injured neurons to reestablish severed connections their processes must explore the extracellular milieu. Thus far, experiments have focused on extracellular matrix (ECM) proteins whose expression is upregulated after CNS injury and that exert inhibitory effects on neurite outgrowth. An intricate balance between ECM synthesis and degradation must be maintained during the tissue remodeling associated with injury. Matrix metalloproteinases (MMPs) are believed to be the main mediators of ECM degradation. MMP activity is tightly regulated by interaction with tissue inhibitors of metalloproteinases (TIMPs). To determine whether TIMPs are expressed during injury-induced matrix remodeling, TIMP expression was examined during reactive gliosis. A stab injury to the mature rat brain leads to the differential regulation of TIMP mRNA expression. Timp-1 and Timp-2 mRNA are significantly upregulated after injury, while the expression of Timp-3 and Timp-4 is unaltered. The expression of Timp-1 in reactive astrocytes and Timp-2 in microglia and neurons suggests these TIMPs may serve distinct functions in response to injury. PMID- 10719362 TI - Novel genomic imbalances in embryonal rhabdomyosarcoma revealed by comparative genomic hybridization and fluorescence in situ hybridization: an intergroup rhabdomyosarcoma study. AB - A comparative genomic hybridization (CGH) approach provides identification of genomic gains and losses in a tumor specimen in a single experiment. Only 11 embryonal rhabdomyosarcomas (E-RMS) have previously been subjected to CGH. The underlying genetic events in this histologic subtype are not well defined. In this investigation, 12 E-RMS specimens from 10 patients entered into Intergroup Rhabdomyosarcoma Study (IRS) I-IV and two local patients were analyzed by CGH and fluorescence in situ hybridization (FISH). Gains of chromosomes or chromosomal regions 2 (50%), 7 (42%), 8 (67%), 11 (42%), 12 (58%), 13q21 (33%), and 20 (33%) and losses of 1p35-36.3 (42%), 6 (33%), 9q22 (33%), 14q21-32 (25%), and 17 (25%) were most prominent. Chromosomal regions 1p35-36.3 and 9q22 represent novel regions of loss. Importantly, loss of 9q22 corresponds to the locus of a putative tumor suppressor gene (PTCH), which has been shown to play a role in rhabdomyosarcoma in a mouse model of Gorlin syndrome. Loss of 1p36 corresponds to the locus for PAX7, a paired box containing gene characteristically altered in alveolar rhabdomyosarcoma. Moreover, loss of 1p36 is prominent in another common pediatric soft tissue tumor, neuroblastoma. Gains of 2, 7, 8, 12, and 13 and loss of 14 were seen in the sole prior E-RMS CGH series; thus, these data provide important confirmatory results. In contrast to this previous study, however loss, not gain, of chromosome 17 was observed in the current study. Chromosome 17 loss correlates well with previous descriptions of frequent allelic loss of 17p (TP53) in E-RMS. In summary, CGH and FISH analyses of 12 E-RMS specimens revealed novel genomic imbalances that may be useful in directing further molecular studies for the determination of E-RMS critically involved genes. PMID- 10719363 TI - Isolation and characterization of a novel TP53-inducible gene, TP53TG5, which suppresses growth and shows cell cycle-dependent transition of expression. AB - Through a strategy of direct cloning of TP53-binding DNA sequences from human genome DNA, we have identified a novel TP53-target gene, termed TP53TG5 (TP53 target gene 5). This gene, localized to chromosome band 20q13.1 by fluorescence in situ hybridization, encodes a 290-amino-acid peptide with no significant homology with any known proteins in the public database. A colony-formation assay using human glioblastoma cell line T98G, which lacks wild-type TP53 and expresses no endogenous TP53TG5, revealed a growth-suppressive effect of the TP53TG5 gene product. Furthermore, immunohistochemical studies, following transfection of T98G with plasmid designed to express green fluorescent protein-fused TP53TG5, revealed cell cycle-dependent intracellular localization of this protein. Our results suggest that functional studies of TP53TG5 may provide new insights into the complex physiological activities of TP53. PMID- 10719364 TI - Homozygous deletions inactivate DCC, but not MADH4/DPC4/SMAD4, in a subset of pancreatic and biliary cancers. AB - Loss of heterozygosity (LOH) of chromosome arm 18q is frequent in gastrointestinal cancers. Over 90% of pancreatic carcinomas have 18q LOH. Bi allelic inactivation of the MADH4/DPC4/SMAD4 gene at 18q21.1 is seen in about half of pancreatic carcinomas with 18q LOH. In the remaining tumors with 18q LOH, MADH4 is not mutated and its expression is unaffected, and no alterations in MADH2/SMAD2, a MADH4-related gene at 18q12.3, have been found. A controversial candidate tumor-suppressor gene at 18q21.2 is DCC (deleted in colorectal carcinoma), which encodes a netrin-1 receptor component with functions in cell migration and apoptosis. Reduced or absent DCC expression has been observed in many cancers, but few somatic mutations that would clearly inactivate DCC function have been reported. We studied a panel of 115 pancreatic and 14 biliary cancers for homozygous deletions of DCC exons and flanking 18q regions. Seven homozygous deletions were seen in the region that includes the DCC gene. In two tumors, the deletions inactivate DCC but not MADH4. A physical and transcript map of the deleted regions was constructed, and DCC was the only known gene affected by all seven deletions. These data are the strongest mutational evidence presented yet in support of the hypothesis that DCC or another gene in the region distal to MADH4 is inactivated, playing a causal role in cancer development. PMID- 10719365 TI - Patients with both pancreatic adenocarcinoma and melanoma may harbor germline CDKN2A mutations. AB - Germline mutations of the CDKN2A tumor suppressor gene have been identified in melanoma kindreds linked to 9p21, and pancreatic adenocarcinoma is the second most common malignancy in some of these families. We hypothesized that unselected patients with both primary cancers, i.e., pancreatic cancer and malignant melanoma, have a genetic predisposition to tumor development, and that this susceptibility may be due to germline CDKN2A mutations. Fourteen patients, with both pathologically verified pancreatic adenocarcinoma and melanoma, were assessed for germline CDKN2A mutations by polymerase chain reaction amplification and sequencing of six overlapping fragments encompassing exons 1alpha and 2. A yeast two-hybrid assay was used to assess the functional consequences of CDKN2A variants. Germline CDKN2A mutations were identified in 2/14 patients: I49S, a novel substitution in exon 1alpha, and M53I, a previously reported missense mutation in exon 2. Both variants lead to compromised CDKN2A function. We conclude that the occurrence of both pancreatic cancer and melanoma, in the same patient, signals an inherited susceptibility to cancer, and that this predisposition is, in some cases, due to germline CDKN2A mutations. This finding has important implications not only for the proband, but also for other family members. PMID- 10719366 TI - A dual-color FISH framework map for the characterization of the Sai1 tumor suppression region on rat chromosome 5. AB - The analysis of cell hybrids between malignant mouse hepatoma cells and normal rat fibroblasts has previously demonstrated the critical role of a deletion in rat chromosome 5 (RNO5) that was related to an anchorage independent phenotype. Those hybrids that were anchorage independent displayed loss of the entire RNO5 or an interstitial deletion in RNO5. These findings suggested that a putative tumor suppressor gene, Sai1 (suppression of anchorage independence 1), was located within the deleted region. To explore the molecular basis of the tumor suppressor activity of the Sai1 region, we analyzed the RNO5q23-q36 region with several genes and microsatellite markers that could be assigned to the region, as well as with new markers derived by representational difference analysis (RDA) or by microdissection. Dual-color FISH was used to construct a detailed physical map of the entire RNO5. These new data can be used to connect the physical and linkage maps in the rat, as well as to identify the details of the comparative map with other mammalian species including humans and mice. Using as FISH reagents genomic YAC, P1, or phage lambda clones corresponding to RNO5 markers, the order and unique positions of 18 markers could be established. The map provided a framework for the detailed characterization of the deletion found in anchorage independent hybrids. All markers within the bands RNO5q31.3-q35 were shown to be lost, including known cancer-related genes such as Ifna (5q32), Cdkn2a, -b (5q32), Jun (5q34), and Cdkn2c (5q35). However, the aberration in the deletion chromosome turned out to be more complex than originally thought in that we detected the presence of a paracentric inversion in addition to a deletion. The inversion led to the juxtaposition of the gene markers Tal2 (5q24.1) and Cd30lg (5q24.3). The framework map will provide the basis for the detailed physical YAC clone contig mapping of this region, and facilitate the identification and characterization of the Sai1 locus. PMID- 10719367 TI - Cervical cancer suppressor gene is within 1 cM on 6p23. AB - We previously showed loss of heterozygosity at 6p to be a common genetic alteration in cervical cancer and cervical intraepithelial neoplasia. To characterize this critical area of deletion in chromosome 6, we evaluated 107 invasive cervical cancers, using 23 polymorphic markers. Genomic DNA from microdissected frozen or paraffin-embedded cervical tumors and corresponding normal tissue was analyzed. Fifty-three percent (57/107) of the cervical tumors showed loss in 6p. Deletions were found in all stages and histologic types. Ninety-one percent (52/57) of these tumors had a loss at 6p23. One tumor defined the distal area of deletion at marker D6S429. Two tumors defined the proximal area of deletion at marker D6S1578. Genotyping of parental DNA was done on 16 cases to evaluate the origin of chromosomal loss. The deletion occurred in the paternal chromosome in 10 tumors and in the maternal in six. Within each tumor, the same parental chromosome was lost at all tested heterozygous 6p markers. The order of the polymorphic markers and estimate of distances in the critical region were confirmed by generation of a yeast artificial chromosome (YAC) contig and pulse-field gel electrophoresis. Our data strongly suggest that a gene important in cervical cancer tumorigenesis is located within a 1-cM region of 6p23, and it is not imprinted. PMID- 10719368 TI - Amplification of the MLL gene on double minutes, a homogeneously staining region, and ring chromosomes in five patients with acute myeloid leukemia or myelodysplastic syndrome. AB - Gene amplification is one of the mechanisms for activating proto-oncogenes resulting in an enhanced expression of the corresponding gene product. By fluorescence in situ hybridization (FISH), amplification of the proto-oncogene MLL has been described only in seven patients with acute myeloid leukemia (AML). We report five new patients (four had de novo AML, one had a de novo myelodysplastic syndrome) displaying different mechanisms of MLL amplification, suspected by G-banding and confirmed by FISH analysis. In two patients, MLL was amplified on double-minute chromosomes (dmins). In both cases, an interstitial deletion in 11q23 including the MLL gene was associated with the occurrence of the dmins containing MLL. As a rarely described mechanism, MLL amplification in the form of size-variable ring chromosomes was observed in two patients. Remodeling of the ring chromosomes leads to multiple copies of MLL and obviously provided a selective growth advantage. In one of the two cases with ring chromosomes, the centromeric alpha-satellite DNA of the ring chromosome was not detectable. Our fifth patient showed the unique finding of MLL amplification within a uniformly (homogeneously?) stained region in interaction with amplified ribosomal DNA sequences. Also, one of the patients with ring chromosomes exhibited the amplification of ribosomal DNA on the ring chromosomes. The transcriptionally active genes for ribosomal RNA could probably enhance the expression of MLL. In one of our five patients, we found the new combination of concomitant amplification of the proto-oncogenes MLL and MYC. PMID- 10719369 TI - High-resolution deletion mapping of chromosome 14 in stromal tumors of the gastrointestinal tract suggests two distinct tumor suppressor loci. AB - DNA copy number losses at chromosome arm 14q are the most frequently occurring aberrations in gastrointestinal stromal tumors (GISTs). To characterize the deletion at 14q, we performed comparative genomic hybridization (CGH) and high resolution deletion mapping using a panel of 32 polymorphic microsatellite markers in 30 GISTs. The GISTs were classified according to their metastatic potential and mitotic counts into 15 low-risk and 15 high-risk tumors. Losses with a minimal common overlapping region at 14q12-q24 were detected by CGH in 16 tumors (53) (nine low-risk and seven high-risk). Investigation with microsatellite markers was informative in 690 analyses (72%). Loss of heterozygosity (LOH) with at least one marker was detected in 279 analyses in 24 tumors (80%). Deletions were equally frequent in low-risk and high-risk GISTs. Two common deletion regions were identified at 14q11.1-q12 and 14q23-q24.3. The highest frequencies of deletions were seen in regions corresponding to markers D14S283 (20/28, 71%) at 14q11.1-q12 and D14S258 (17/27, 63%) at 14q23-q24, suggesting that these are two tumor suppressor loci. PMID- 10719370 TI - Genetic modeling of human urinary bladder carcinogenesis. AB - We developed a model of human urinary bladder cancer progression from in situ precursor lesions to invasive carcinoma using whole organ histologic and genetic mapping. The model represents a high-density and detailed analysis regarding allelic losses on chromosomes 4, 8, 9, 11, and 17 as revealed by testing of 234 samples obtained from five cystectomy specimens. The samples corresponded to microscopically identified intraurothelial precursor conditions ranging from dysplasia to carcinoma in situ and invasive cancer. The initial analysis of paired normal and tumor DNA samples disclosed allelic losses in 72 of 225 tested hypervariable DNA markers. Subsequent use of these markers on all mucosal samples revealed that 47 had alterations with a statistically significant relation to urothelial neoplasia. The allelic losses clustered in 33 distinct chromosomal regions, indicating the location of putative tumor suppressor genes involved in the development and progression of urinary bladder cancer. Some of the markers with statistically significant allelic losses mapped to the regions containing well-characterized tumor suppressor genes but many were located in previously unknown loci. The majority of statistically significant allelic losses (70%) occurred early in low-grade intraurothelial dysplasia, and some of them involved adjacent areas of morphologically normal mucosa preceding the development of microscopically recognizable precursor lesions. The remaining 30% of markers developed allelic losses in the later phases of urothelial neoplasia, implicating their involvement in progression to invasive disease. Markers exhibiting allelic losses in early phases of urothelial neoplasia could be used for detection of occult preclinical or even premicroscopic phases of urinary bladder cancer, whereas markers that showed allelic losses in the later phases of the process could serve as indicators of progression to invasive disease. The approach used in this study facilitates genome-wide modeling of cancer progression and provides important chromosomal landmarks for more specific studies of multistep urinary bladder carcinogenesis. PMID- 10719371 TI - Distinct clinicopathologic and genetic profiles in sporadic gastric cancer with different mutator phenotypes. AB - A subset of sporadic gastric cancers (GC) exhibits microsatellite instability (MSI). To define the precise role of MSI in GC, a total of 100 patients with sporadic GC were classified into three groups, i.e., high-frequency MSI (MSI-H), low-frequency MSI (MSI-L), and microsatellite stable (MSS), based on 10 microsatellite markers. Mutational analyses of TGFbetaRII, IGFIIR, BAX, MSH3, MSH6, E2F4, MSH2, MLH1, and TP53 genes, and methylation and protein expression of MLH1 and MSH2 were performed and correlated. Twenty-seven percent of GC showed MSI at least in one locus and could be further graded as MSI-H (14%) and MSI-L (13%). No clinicopathologic difference was noted between GC with MSI-L and MSS. Compared with GC with MSI-L or MSS, GC with MSI-H had a significantly higher frequency of antral location, intestinal subtype, H. pylori seropositivity, but a lower incidence of lymph node metastasis, and displayed a higher frequency of frameshift mutations of TGFbetaRII, IGFIIR, BAX, MSH3, and E2F4 genes but a lower incidence of TP53 mutations. Furthermore, hypermethylation of the MLH1 promoter was responsible for the loss of protein function in 13 of 14 MSI-H tumors. It was concluded that a specific phenotype and a distinct profile of genetic alterations exist in MSI-H GC. We speculate that epigenetic inactivation of MLH1 by methylation plays a crucial role in initiating such a pathway of carcinogenesis. In contrast, GCs with MSS and MSI-L exhibit clinicopathologic features that are distinct from MSI-H tumors and have a higher frequency of TP53 mutations, suggesting that they may evolve through an entirely different pathway. PMID- 10719373 TI - Spectral karyotyping combined with locus-specific FISH simultaneously defines genes and chromosomes involved in chromosomal translocations. AB - Genes that play roles in malignant transformation have often been found proximate to cancer-associated chromosomal breakpoints. Identifying genes that flank chromosomal reconfigurations is thus essential for cancer cytogenetics. To simplify and expedite this identification, we have developed a novel approach, based on simultaneous spectral karyotyping and fluorescence in situ hybridization (FISH) which, in a single step, can identify gross chromosomal aberrations as well as detect the involvement of specific loci in these rearrangements. Signals for specifically queried genes (FISH probe) were easily detectable in metaphase cells, together with the signals from painted chromosomes (spectral karyotyping probes). The concentration and size of the FISH probes could cover a wide range and still be used successfully. Some of the nucleotide-bound dyes used for the labeling, as Cy3, Spectrum Orange, Alexa 594, Texas Red, and Rhodamine 110, were particularly efficient. More than one gene can be queried in the same metaphase, because multiple FISH probes could be hybridized simultaneously. To demonstrate this technique, we applied it to the myeloma cell line Karpas 620, which has numerous chromosomal rearrangements. The approach that we present here will be particularly useful for the analysis of complex karyotypes and for testing hypotheses arising from cancer gene expression studies. Published 2000 Wiley Liss, Inc. PMID- 10719372 TI - Molecular analysis of the rearranged genome and chimeric mRNAs caused by the t(6;11)(q27;q23) chromosome translocation involving MLL in an infant acute monocytic leukemia. AB - Chromosomal analysis of acute monocytic leukemia cells in a female infant revealed a t(6;11)(q27;q23) translocation. Southern blot analysis with a cDNA probe of the MLL gene at chromosome band 11q23 indicated that the breakpoint was in an 8.3-kb BamHI fragment that contained exons 5-11 of the MLL gene. Northern blot analysis showed a faint band corresponding to the MLL chimeric transcript. Structural analysis of a genomic clone with the rearranged MLL gene from der(11) chromosome demonstrated the breakpoint to be localized between exons 6 and 7 of the the MLL gene and to lie in an Alu sequence of this region. The partner gene fused to the 3' part of MLL was shown to be the AF6 gene on chromosome 6q27 by in situ chromosome hybridization and nucleotide sequencing of chimeric MLL cDNA clones. However, it was shown that MLL exon 5 was fused to AF6 in one clone, whereas most clones were MLL exon 6/AF6 chimeric cDNA clones. These findings indicate that exon 6 of MLL is spliced out in the process of transcription in a variant MLL/AF6. In addition, we were able to detect the splicing of exon 6 in either this chimeric MLL/AF6 or MLL transcripts from untranslocated chromosomes by reverse transcriptase-polymerase chain reaction. The detailed genetic map of AF6 was determined by in situ chromosome hybridization and radiation hybrid mapping. PMID- 10719374 TI - Genomic instability and target gene mutations in colon cancers with different degrees of allelic shifts. AB - Two grades (high and low) of microsatellite instability (MSI) are known, depending on the number of mutated markers and the amount of allelic shifts. Forty-two colorectal tumors, previously found to have high-degree MSI at dinucleotidic repeat loci, were revisited with BAT26, a mononucleotide marker, and the number of shifted bases were counted. Seven tumors, all with local stages at diagnosis, had < or =6-bp deletions and consistently displayed shorter shifts also with other intronic mononucleotide markers. Analysis of mononucleotide tracts in the coding regions of MSH3, MSH6, BAX, and TGFbetaRII in the groups with large (>6 bp) and short (< or =6 bp) allelic shifts showed specific patterns of involvement for the individual genes: TGFbetaRII displayed a uniformly high rate of mutations, while MSH3, MSH6, and BAX were less frequently altered in tumors with short shifts. Our findings suggest that microsatellite instability arises gradually, evenly involving loci with similar features of length and repetition. However, target genes have a specific timing of mutation in this process: TGFbetaRII is involved in the early phases, while BAX and MSH6 are frequently associated with big size shifts and tumors with more advanced stages. PMID- 10719375 TI - How are antibodies involved in the protective mechanism of susceptible mice infected with T. cruzi? AB - Host resistance to Trypanosoma cruzi is dependent on both natural and acquired immune responses. During the acute phase of the infection the presence of IFN gamma, TNF-alpha, IL-12 and GM-CSF has been closely associated with resistance, whereas TGF-ss and IL-10 have been associated with susceptibility. Several investigators have demonstrated that antibodies are responsible for the survival of susceptible animals in the initial phase of infection and for the maintenance of low levels of parasitemia in the chronic phase. However, how this occurs is not yet understood. Our results and other data in the literature support the hypothesis that the protective role of antibodies in the acute phase of infection is dependent mostly on their ability to induce removal of bloodstream trypomastigotes from the circulation in addition to other concomitant cell mediated events. PMID- 10719376 TI - Activity and functional interaction of alternative oxidase and uncoupling protein in mitochondria from tomato fruit. AB - Cyanide-resistant alternative oxidase (AOX) is not limited to plant mitochondria and is widespread among several types of protists. The uncoupling protein (UCP) is much more widespread than previously believed, not only in tissues of higher animals but also in plants and in an amoeboid protozoan. The redox energy dissipating pathway (AOX) and the proton electrochemical gradient energy dissipating pathway (UCP) lead to the same final effect, i.e., a decrease in ATP synthesis and an increase in heat production. Studies with green tomato fruit mitochondria show that both proteins are present simultaneously in the membrane. This raises the question of a specific physiological role for each energy dissipating system and of a possible functional connection between them (shared regulation). Linoleic acid, an abundant free fatty acid in plants which activates UCP, strongly inhibits cyanide-resistant respiration mediated by AOX. Moreover, studies of the evolution of AOX and UCP protein expression and of their activities during post-harvest ripening of tomato fruit show that AOX and plant UCP work sequentially: AOX activity decreases in early post-growing stages and UCP activity is decreased in late ripening stages. Electron partitioning between the alternative oxidase and the cytochrome pathway as well as H+ gradient partitioning between ATP synthase and UCP can be evaluated by the ADP/O method. This method facilitates description of the kinetics of energy-dissipating pathways and of ATP synthase when state 3 respiration is decreased by limitation of oxidizable substrate. PMID- 10719377 TI - Signal transduction induced in Trypanosoma cruzi metacyclic trypomastigotes during the invasion of mammalian cells. AB - Penetration of Trypanosoma cruzi into mammalian cells depends on the activation of the parasite's protein tyrosine kinase and on the increase in cytosolic Ca2+ concentration. We used metacyclic trypomastigotes, the T. cruzi developmental forms that initiate infection in mammalian hosts, to investigate the association of these two events and to identify the various components of the parasite signal transduction pathway involved in host cell invasion. We have found that i) both the protein tyrosine kinase activation, as measured by phosphorylation of a 175 kDa protein (p175), and Ca2+ mobilization were induced in the metacyclic forms by the HeLa cell extract but not by the extract of T. cruzi-resistant K562 cells; ii) treatment of parasites with the tyrosine kinase inhibitor genistein blocked both p175 phosphorylation and the increase in cytosolic Ca2+ concentration; iii) the recombinant protein J18, which contains the full-length sequence of gp82, a metacyclic stage surface glycoprotein involved in target cell invasion, interfered with tyrosine kinase and Ca2+ responses, whereas the monoclonal antibody 3F6 directed at gp82 induced parasite p175 phosphorylation and Ca2+ mobilization; iv) treatment of metacyclic forms with phospholipase C inhibitor U73122 blocked Ca2+ signaling and impaired the ability of the parasites to enter HeLa cells, and v) drugs such as heparin, a competitive IP3-receptor blocker, caffeine, which affects Ca2+ release from IP3-sensitive stores, in addition to thapsigargin, which depletes intracellular Ca2+ compartments and lithium ion, reduced the parasite infectivity. Taken together, these data suggest that protein tyrosine kinase, phospholipase C and IP3 are involved in the signaling cascade that is initiated on the parasite cell surface by gp82 and leads to Ca2+ mobilization required for target cell invasion. PMID- 10719378 TI - Morphologic and biochemical changes in male rat lung after surgical and pharmacological castration. AB - The morphology of the rat lung was studied by light microscopy in different situations: after surgical and pharmacological castration and after administration of testosterone to the castrated rat to determine if the androgen is required to maintain the normal morphology of the lung. We also determined the effect of flutamide on the phospholipid composition of both the surfactant and microsomes of the lung. Rats were separated into five groups: I - control non castrated rats, II - castrated rats sacrificed 21 days after castration, III - castrated rats that received testosterone daily from day 2 to day 21 after castration, IV - castrated rats that received testosterone from day 15 to day 21 after castration, and V - control rats injected with flutamide for 7 days. The amount of different phospholipids in the surfactant and microsomes of the lung was measured in group I and V rats. At the light microscopy level, the surgical and pharmacological castration provoked alterations in the morphology of the lung, similar to that observed in human lung emphysema. The compositions of surfactant and microsomes of the lung were similar to those previously reported by us for the surgically castrated rats. These results indicate that androgens are necessary for the normal morphology as well as for some metabolic aspects of the lung. PMID- 10719379 TI - Retinol-induced changes in the phosphorylation levels of histones and high mobility group proteins from Sertoli cells. AB - Chromatin proteins play a role in the organization and functions of DNA. Covalent modifications of nuclear proteins modulate their interactions with DNA sequences and are probably one of the multiple factors involved in the process of switch on/off transcriptionally active regions of DNA. Histones and high mobility group proteins (HMG) are subject to many covalent modifications that may modulate their capacity to bind to DNA. We investigated the changes induced in the phosphorylation pattern of cultured Wistar rat Sertoli cell histones and high mobility group protein subfamilies exposed to 7 microM retinol for up to 48 h. In each experiment, 6 h before the end of the retinol treatment each culture flask received 370 KBq/ml [32P]-phosphate. The histone and HMGs were isolated as previously described [Moreira et al. Medical Science Research (1994) 22: 783 784]. The total protein obtained by either method was quantified and electrophoresed as described by Spiker [Analytical Biochemistry (1980) 108: 263 265]. The gels were stained with Coomassie brilliant blue R-250 and the stained bands were cut and dissolved in 0.5 ml 30% H2O2 at 60oC for 12 h. The vials were chilled and 5.0 ml scintillation liquid was added. The radioactivity in each vial was determined with a liquid scintillation counter. Retinol treatment significantly changed the pattern of each subfamily of histone and high mobility group proteins. PMID- 10719380 TI - Antimicrobial resistance patterns of Haemophilus influenzae isolated from patients with meningitis in Sao Paulo, Brazil. AB - From 1989 to 1995, a total of 391 Haemophilus influenzae isolates were recovered from the cerebrospinal fluid (CSF) of hospitalized patients in Sao Paulo, Brazil. The majority of strains were isolated from infants aged less than 5 years. Strains belonging to biotype I (64.7%), biotype II (34.5%) and biotype IV (0.76%) were detected. Ninety-nine percent of these strains were serotype b. Minimal inhibitory concentration (MIC) was determined for ampicillin, chloramphenicol and ceftriaxone. The ss-lactamase assay was performed for all strains. The rate of ss lactamase producer strains ranged from 10 to 21.4% during a period of 7 years, with an overall rate of 13.8%. Of the 391 strains analyzed, none was ss-lactamase negative ampicillin resistant (BLNAR). A total of 9.7% of strains showed resistance to both ampicillin and chloramphenicol; however, 4% of them were resistant to ampicillin only and 2% to chloramphenicol. All strains were susceptible to ceftriaxone and the MIC90 was 0.007 microg/ml, suggesting that ceftriaxone could be an option for the treatment of bacterial meningitis in pediatric patients who have not been screened for drug sensitivity. PMID- 10719381 TI - HFE gene mutations in coronary atherothrombotic disease. AB - Although iron can catalyze the production of free radicals involved in LDL lipid peroxidation, the contribution of iron overload to atherosclerosis remains controversial. The description of two mutations in the HFE gene (Cys282Tyr and His63Asp) related to hereditary hemochromatosis provides an opportunity to address the question of the association between iron overload and atherosclerosis. We investigated the prevalence of HFE mutations in 160 survivors of myocardial infarction with angiographically demonstrated severe coronary atherosclerotic disease, and in 160 age-, gender- and race-matched healthy control subjects. PCR amplification of genomic DNA followed by RsaI and BclI restriction enzyme digestion was used to determine the genotypes. The frequency of the mutant Cys282Tyr allele was identical among patients and controls (0.022; carrier frequency, 4.4%), whereas the mutant His63Asp allele had a frequency of 0.143 (carrier frequency, 27.5%) in controls and of 0.134 (carrier frequency, 24.5%) in patients. Compound heterozygotes were found in 2 of 160 (1.2%) controls and in 1 of 160 (0.6%) patients. The finding of a similar prevalence of Cys282Tyr and His63Asp mutations in the HFE gene among controls and patients with coronary atherothrombotic disease, indirectly questions the possibility of an association between hereditary hemochromatosis and atherosclerosis. PMID- 10719382 TI - Biomechanical and histological evaluation of hydrogel implants in articular cartilage. AB - We evaluated the mechanical behavior of the repaired surfaces of defective articular cartilage in the intercondylar region of the rat femur after a hydrogel graft implant. The results were compared to those for the adjacent normal articular cartilage and for control surfaces where the defects remained empty. Hydrogel synthesized by blending poly(2-hydroxyethyl methacrylate) and poly(methyl methacrylate-co-acrylic acid) was implanted in male Wistar rats. The animals were divided into five groups with postoperative follow-up periods of 3, 5, 8, 12 and 16 weeks. Indentation tests were performed on the neoformed surfaces in the knee joint (with or without a hydrogel implant) and on adjacent articular cartilage in order to assess the mechanical properties of the newly formed surface. Kruskal-Wallis analysis indicated that the mechanical behavior of the neoformed surfaces was significantly different from that of normal cartilage. Histological analysis of the repaired defects showed that the hydrogel implant filled the defect with no signs of inflammation as it was well anchored to the surrounding tissues, resulting in a newly formed articular surface. In the case of empty control defects, osseous tissue grew inside the defects and fibrous tissue formed on the articular surface of the defects. The repaired surface of the hydrogel implant was more compliant than normal articular cartilage throughout the 16 weeks following the operation, whereas the fibrous tissue that formed postoperatively over the empty defect was stiffer than normal articular cartilage after 5 weeks. This stiffness started to decrease 16 weeks after the operation, probably due to tissue degeneration. Thus, from the biomechanical and histological point of view, the hydrogel implant improved the articular surface repair. PMID- 10719383 TI - Zymosan phagocytosis by mouse peritoneal macrophages is increased by apoHDL- and not by intact HDL-covered particles. AB - The uptake of lipids and lipoprotein particles by macrophages undergoes phagocytic activation and the formation of foam cells are key events in atherosclerosis. In this study we determined how intact high density lipoproteins (HDL) and apolipoproteins-HDL (removal of the lipid component from HDL, i.e., apoHDL) influence the phagocytosis of zymosan by mouse peritoneal macrophages. Zymosan particles preincubated together with lipoproteins or alone (control) were incubated with the macrophages. Phagocytosis activity was reported as the percent of macrophages that internalized three or more zymosan particles. HDL co incubated with zymosan did not influence the over-all uptake of zymosan particles compared to apoHDL, which greatly enhanced the ability of the particle to be phagocytized (P<0.001). Part of this effect might be related to a greater binding of apoHDL to the particles compared to that of HDL (P<0.05). We conclude that this can be a useful method to study the ability of lipoproteins, including modified lipoproteins obtained from subjects with genetic forms of hyperlipidemia, to opsonize particles such as red blood cells and thus to investigate the processes that control the formation of foam cells and the mechanisms of atherogenesis. PMID- 10719384 TI - Detection of early apoptosis and cell death in T CD4+ and CD8+ cells from lesions of patients with localized cutaneous leishmaniasis. AB - Human localized cutaneous leishmaniasis (LCL), induced by Leishmania braziliensis, ranges from a clinically mild, self-healing disease with localized cutaneous lesions to severe forms which can present secondary metastatic lesions. The T cell-mediated immune response is extremely important to define the outcome of the disease; however, the underlying mechanisms involved are not fully understood. A flow cytometric analysis of incorporation of 7-amino actinomycin D and CD4+ or CD8+ T cell surface phenotyping was used to determine whether different frequencies of early apoptosis or accidental cell death occur at different stages of LCL lesions. When all cells obtained from a biopsy sample were analyzed, larger numbers of early apoptotic and dead cells were observed in lesions from patients with active disease (mean = 39.5 +/- 2.7%) as compared with lesions undergoing spontaneous healing (mean = 17.8 +/- 2.2%). Cells displaying normal viability patterns obtained from active LCL lesions showed higher numbers of early apoptotic events among CD8+ than among CD4+ T cells (mean = 28.5 +/- 3.8 and 15.3 +/- 3.0%, respectively). The higher frequency of cell death events in CD8+ T cells from patients with LCL may be associated with an active form of the disease. In addition, low frequencies of early apoptotic events among the CD8+ T cells were observed in two patients with self-healing lesions. Although the number of patients in the latter group was small, it is possible to speculate that, during the immune response, differences in apoptotic events in CD4+ and CD8+ T cell subsets could be responsible for controlling the CD4/CD8 ratio, thus leading to healing or maintenance of disease. PMID- 10719385 TI - Compound 48/80, a histamine-depleting agent, blocks the protective effect of morphine against electroconvulsive shock in mice. AB - We have shown that morphine has an anticonvulsive effect against maximal electroconvulsive shock (MES) in mice, and this effect is antagonized by histamine H1-receptor antagonists. Brain histamine is localized both in neurons and in mast cells, and morphine is known to enhance the turnover of neuronal histamine and to release histamine from mast cells. In the present experiments, compound 48/80 was injected chronically (0.5 mg/kg on day 1, 1 mg/kg on day 2, 2 mg/kg on day 3, 3 mg/kg on day 4, and 4 mg/kg on day 5, twice daily, ip) to deplete mast cell contents. Morphine (0.001-10 mg/kg, ip; N = 20) produced a dose dependent anticonvulsive effect against MES seizure in mice with non-depleted mast cells, whereas it did not exert any anticonvulsive effect in mice with depleted mast cells. These results indicate that morphine produces its anticonvulsive effect against maximal electroconvulsive shock in mice by liberating histamine from mast cells. PMID- 10719386 TI - Quantitative studies of the vasculature of the carotid body in the chronically hypoxic rat. AB - The carotid bodies of rats made chronically hypoxic by breathing 12% O2 in a normobaric chamber (inspired PO2 91 mmHg) were compared with those of controls. Serial 5-microm sections of the organs were examined using an interactive image analysis system. The total volume of the carotid bodies was increased by 64%. The total vascular volume rose by 103% and was likely due to an increase in size of the large vessels (>12 microm lumen diameter) because the small vessel (5-12 microm lumen diameter) volume did not increase significantly while the small vessel density tended to decrease. The extravascular volume was increased by 57%. Expressed as a percentage of the total volume of the organ, the total vascular volume did not change, but the small vessel volume was significantly decreased from 7.83 to 6.06%. The large vessel volume must therefore have been increased. The proportion occupied by the extravascular volume was virtually unchanged (84 vs 82%). In accordance with these findings, the small vessel endothelial surface area per unit carotid body volume was diminished from 95.2 to 76.5 mm-1, while the extravascular area per small vessel was increased from 493 to 641 microm(2) or by 30%. In conclusion, the enlargement of the carotid body in chronic hypoxia is most likely due to an increase in total vascular volume, mainly involving the "large" vessels, and to an increase in extravascular volume. This is in contrast to our previously published findings indicating that in the spontaneous insulin dependent diabetic rat the enlargement of the carotid body is due solely to an increase in extravascular volume. PMID- 10719387 TI - A pulsatile flow model for in vitro quantitative evaluation of prosthetic valve regurgitation. AB - A pulsatile pressure-flow model was developed for in vitro quantitative color Doppler flow mapping studies of valvular regurgitation. The flow through the system was generated by a piston which was driven by stepper motors controlled by a computer. The piston was connected to acrylic chambers designed to simulate "ventricular" and "atrial" heart chambers. Inside the "ventricular" chamber, a prosthetic heart valve was placed at the inflow connection with the "atrial" chamber while another prosthetic valve was positioned at the outflow connection with flexible tubes, elastic balloons and a reservoir arranged to mimic the peripheral circulation. The flow model was filled with a 0.25% corn starch/water suspension to improve Doppler imaging. A continuous flow pump transferred the liquid from the peripheral reservoir to another one connected to the "atrial" chamber. The dimensions of the flow model were designed to permit adequate imaging by Doppler echocardiography. Acoustic windows allowed placement of transducers distal and perpendicular to the valves, so that the ultrasound beam could be positioned parallel to the valvular flow. Strain-gauge and electromagnetic transducers were used for measurements of pressure and flow in different segments of the system. The flow model was also designed to fit different sizes and types of prosthetic valves. This pulsatile flow model was able to generate pressure and flow in the physiological human range, with independent adjustment of pulse duration and rate as well as of stroke volume. This model mimics flow profiles observed in patients with regurgitant prosthetic valves. PMID- 10719388 TI - Effect of inhibition of nitric oxide synthase on blood pressure and renal sodium handling in renal denervated rats. AB - The role of sympathetic nerve activity in the changes in arterial blood pressure and renal function caused by the chronic administration of N G-nitro-L-arginine methyl ester (L-NAME), an inhibitor of nitric oxide (NO) synthesis, was examined in sham and bilaterally renal denervated rats. Several studies have demonstrated that sympathetic nerve activity is elevated acutely after L-NAME administration. To evaluate the role of renal nerve activity in L-NAME-induced hypertension, we compared the blood pressure response in four groups (N = 10 each) of male Wistar Hannover rats weighing 200 to 250 g: 1) sham-operated vehicle-treated, 2) sham operated L-NAME-treated, 3) denervated vehicle-treated, and 4) denervated L-NAME treated rats. After renal denervation or sham surgery, one control week was followed by three weeks of oral administration of L-NAME by gavage. Arterial pressure was measured weekly in conscious rats by a tail-cuff method and renal function tests were performed in individual metabolic cages 0, 7, 14 and 21 days after the beginning of L-NAME administration. L-NAME (60 mg kg-1 day-1) progressively increased arterial pressure from 108 +/- 6.0 to 149 +/- 12 mmHg (P<0.05) in the sham-operated group by the third week of treatment which was accompanied by a fall in creatinine clearance from 336 +/- 18 to 222 +/- 59 microl min-1 100 g body weight-1 (P<0. 05) and a rise in fractional urinary sodium excretion from 0.2 +/- 0. 04 to 1.62 +/- 0.35% (P<0.05) and in sodium post proximal fractional excretion from 0.54 +/- 0.09 to 4.7 +/- 0.86% (P<0.05). The development of hypertension was significantly delayed and attenuated in denervated L-NAME-treated rats. This was accompanied by a striking additional increase in fractional renal sodium and potassium excretion from 0.2 +/- 0.04 to 4.5 +/- 1.6% and from 0.1 +/- 0.015 to 1.21 +/- 0.37%, respectively, and an enhanced post-proximal sodium excretion compared to the sham-operated group. These differences occurred despite an unchanged creatinine clearance and Na+ filtered load. These results suggest that bilateral renal denervation delayed and attenuated the L-NAME-induced hypertension by promoting an additional decrease in tubule sodium reabsorption in the post-proximal segments of nephrons. Much of the hypertension caused by chronic NO synthesis inhibition is thus dependent on renal nerve activity. PMID- 10719389 TI - Thyroid peroxidase activity is inhibited by amino acids. AB - Normal in vitro thyroid peroxidase (TPO) iodide oxidation activity was completely inhibited by a hydrolyzed TPO preparation (0.15 mg/ml) or hydrolyzed bovine serum albumin (BSA, 0.2 mg/ml). A pancreatic hydrolysate of casein (trypticase peptone, 0.1 mg/ml) and some amino acids (cysteine, tryptophan and methionine, 50 microM each) also inhibited the TPO iodide oxidation reaction completely, whereas casamino acids (0.1 mg/ml), and tyrosine, phenylalanine and histidine (50 microM each) inhibited the TPO reaction by 54% or less. A pancreatic digest of gelatin (0.1 mg/ml) or any other amino acid (50 microM) tested did not significantly decrease TPO activity. The amino acids that impair iodide oxidation also inhibit the TPO albumin iodination activity. The inhibitory amino acids contain side chains with either sulfur atoms (cysteine and methionine) or aromatic rings (tyrosine, tryptophan, histidine and phenylalanine). Among the amino acids tested, only cysteine affected the TPO guaiacol oxidation reaction, producing a transient inhibition at 25 or 50 microM. The iodide oxidation inhibitory activity of cysteine, methionine and tryptophan was reversed by increasing iodide concentrations from 12 to 18 mM, while no such effect was observed when the cofactor (H2O2) concentration was increased. The inhibitory substances might interfere with the enzyme activity by competing with its normal substrates for their binding sites, binding to the free substrates or reducing their oxidized form. PMID- 10719391 TI - [Comparison of hypoglycemia perception and hormonal counterregulation in controlled hypoglycemia. A contribution to diagnosis of hypoglycemia unawareness syndrome in type 1 diabetic patients]. AB - BACKGROUND AND OBJECTIVE: Diabetics with abnormal hypoglycaemia awareness cannot recognize hypoglycaemic symptoms early and adequately enough to respond, thus endangering in everyday life not only themselves but also others. Thus it is important to diagnose such symptoms early and assess the extent to which the diabetic is at risk of severe hypoglycaemia. This study was designed to contribute to the evaluation of hypoglycaemia awareness in type 1 (IDDM) diabetics. PATIENTS AND METHODS: In 57 diabetics (IDDM) awareness of typical hypoglycaemic warning symptoms and of hormonal counterregulation were tested during controlled induced hypoglycaemia. A questionnaire was used to measure at fixed time intervals, along a four-point scale, the severity of 6 autonomic and 6 cerebral warning symptoms, and compare them with synchronously obtained plasma levels of glucagon and epinephrine. RESULTS: The cohort was divided into three groups according to the rise of epinephrine levels induced by the hypoglycaemia. Group A (n = 22) showed a significant rise in epinephrine level at the onset of hypoglycaemia; group B (n = 22): significantly increased epinephrine only after 30 min of hypoglycaemia; group C (n = 15): no epinephrine rise at any time during hypoglycaemia. None of the patients had a significant rise in glucagon levels. There were significant differences between the three groups regarding HbA1c levels and the duration of diabetes. Low levels of HbA1c and duration of diabetes were commonly associated with a loss of hormonal counterregulation. In only 40% of patients was it possible to draw any conclusion about epinephrine-induced hypoglycaemia from symptoms revealed in the past history. CONCLUSIONS: The past history of hypoglycaemic symptoms is an unreliable and inadequate guide for objectively assessing the actual risk of hypoglycaemia in type 1 diabetics. The results of this study indicate that when severe hypoglycaemic attacks (requiring outside help etc.) are suspected, standardized diagnostic tests of hypoglycaemia awareness should be performed in specialized centres, particularly if an official medical report ist required. PMID- 10719390 TI - [Effectiveness and efficiency of ambulatory diabetes education programs. A comparison of specialty practice and general practice]. AB - BACKGROUND AND OBJECTIVE: The increasing incidence of NIDDM requires adequate proof of the effectiveness and efficiency of out-patient programmes for diabetics. This study aimed at examining which of the two methods may promise the better results under controlled expenses--modifications of the present standardized diabetes education program in the general practice or specifically developed diabetes education programme in specialist diabetes practice. PATIENTS AND METHODS: 75 diabetics took part in the different out-patient diabetes education programmes at one special practice for diabetology or one of seven general practitioner practices, respectively. The self-developed intensive diabetes education programme of the special practice led by two specialists in diabetology and a dietician was compared with the diabetes education programme of the general practices performed by a doctor's assistant. 38 diabetics in eight training groups at the specialist for diabetology and 37 patients in seven training groups at the general practices were instructed. 32 patients in total (18 patients at the specialist practice and 14 patients at the general practices) additionally received an evaluated support programme which addressed psychosocial impediments related to the topics of a structured diabetes therapy. All patients were asked to complete a questionnaire before, right after, 3 and 6 months after the programme. Weight and glycosylated haemoglobin (HbA1c) were measured before, 3 and 6 months after the programme. RESULTS: The mean weight and glycosylated hemoglobin of all groups decreased as expected. However, the patients of the general practices achieved more lasting reduction of these objective parameters. All patients stated impairments of their quality of life, but the patients of the specialist practice felt more impaired. The motivational support programme achieved only few positive results. CONCLUSION: The standardized diabetes education programme of the general practices and the more intensive and expensive diabetes education programme achieved equally valuable results. Regarding time and expenses, the standardized diabetes education programme may be the more efficient method of NIDDM out-patient education. PMID- 10719392 TI - [Retractile mesenteritis after resection of Meckel's diverticulum]. AB - HISTORY AND ADMISSION FINDINGS: A 22-year-old man who had never been seriously ill previously was admitted because of epigastric pain and vomiting of bile. INVESTIGATIONS: He had slight pain on pressure over the epigatric region and decreased intestinal peristaltic sounds. There was evidence of ileus of the small intestine both by ultrasound and radiologically. TREATMENT AND COURSE: As the patient's condition deteriorated on conservative treatment, an exploratory laparotomy was performed. It revealed an invaginated Meckel's diverticulum, Ileus of the small intestine recurred postoperatively, requiring relaparotomy. A glued together volvulus of the small intestine had to be resected, even though there was no sign of an anastomotic leak. But there was no postoperative improvement. A third operation revealed a clearly shortened and 3 cm-thick mesentery which showed a stage III retractile mesenteritis. Histological re-examination of the specimens resected at the previous operations revealed stage I and II retractile mesenteritis. The patient's condition slowly improved on high doses of corticosteroids and he ultimately became symptom-free. CONCLUSIONS: Retractile mesenteritis is a very rare benign disease of the mesentery, almost always causing abdominal pain and diagnosed histologically by exploratory laparotomy. Administration of corticosteroids is the treatment of choice. PMID- 10719393 TI - [Disruption of cardiac pacemaker function by magnetic buttons on clothing]. PMID- 10719394 TI - [Therapy and after-care of familial adenomatous polyposis and hereditary colorectal carcinoma without polyposis]. PMID- 10719395 TI - [Clinical relevance of phospholipids in the gastrointestinal tract]. PMID- 10719396 TI - [Model for certification in specialty gastroenterology practice according to DIN ISO 9001]. PMID- 10719397 TI - [Fatal liver failure after corticosteroid therapy in hepatitis B carrier status]. PMID- 10719398 TI - The preconception question: nutritional foundations of fertility in women and men. PMID- 10719399 TI - FDA approves health claim labeling for foods containing soy protein. PMID- 10719400 TI - National nutrition month: planning an effective multicultural program. PMID- 10719401 TI - Healthy People 2010 targets healthy diet and healthy weight as critical goals. PMID- 10719402 TI - National Nutrition Month: more than just a month. PMID- 10719403 TI - Accuracy of energy intake data estimated by a multiple-pass, 24-hour dietary recall technique. AB - OBJECTIVE: This study examined the accuracy of a multiple-pass, 24-hour dietary recall method for estimating energy intakes of men and women by comparing it with energy intake required for weight maintenance. DESIGN: Three-day, multiple-pass, 24-hour recalls were obtained on randomly selected days during a self-selected diet period when subjects were preparing their own meals and during a controlled diet period when all meals were provided by the study. During the dietary intervention, weight was maintained; body weight and dietary intake were monitored closely, thereby allowing estimation of the energy intake required for weight maintenance. SUBJECTS/SETTING: Seventy-eight men and women (22 to 67 years old) from the Dietary Effects on Lipoprotein and Thrombogenic Activity (DELTA) study participated in this study. All 24-hour recalls were collected using a computer-assisted, interactive, multiple-pass telephone interview technique. Energy requirements for each individual were determined by the energy content of the DELTA study foods provided to maintain weight. STATISTICAL ANALYSIS: Paired and independent t tests were conducted to examine differences among study variables. Agreement between recalled energy intake and weight maintenance energy intake was analyzed using the Bland-Altman technique. RESULTS: Compared with weight maintenance energy intake, during the self-selected diet period men and women underestimated energy intake by 11% and 13%, respectively. During the controlled diet period, men underestimated energy intake by 13%, whereas women overestimated energy by 1.3%. APPLICATIONS/CONCLUSIONS: Men had a tendency to under-estimate energy intake irrespective of the recording period. The accuracy of the recalled energy intake of women may be influenced by recording circumstances. Researchers should examine the factors influencing underreporting and overreporting by individuals and their impact on macronutrient and micronutrient intakes. Also, strategies need to be developed to minimize underreporting and overreporting. PMID- 10719404 TI - Pecans lower low-density lipoprotein cholesterol in people with normal lipid levels. AB - OBJECTIVE: To compare serum lipid profiles and dietary intakes of people with normal lipid levels who consumed pecans and those who did not consume nuts. DESIGN: Eight-week, randomized, controlled study of pecan treatment group vs control group. SUBJECT: Nineteen people with normal lipid levels completed the study; 10 had been randomly assigned to the pecan treatment group (7 women, 3 men, mean age = 45 +/- 10 years) and 9 to the control group (8 women, 1 man, mean age = 37 +/- 12 years). INTERVENTION: The pecan treatment group consumed 68 g pecans per day for 8 weeks plus self-selected diets. The pecans contributed 459 kcal and 44 g fat daily. The control group avoided nuts and consumed self selected diets. MAIN OUTCOME MEASURES: Total serum cholesterol, low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), and total triglyceride levels were measured at the time of entrance to the study (baseline), week 4, and week 8. Computer analyses were done on five 3-day food records. STATISTICAL ANALYSIS: Comparisons were made using analysis of variance or paired t test. RESULTS: LDL-C was lowered in the pecan treatment group from 2.61 +/- 0.49 mmol/L at baseline to 2.35 +/- 0.49 at week 4 (P < .05) and to 2.46 +/- 0.59 at week 8 (P < .05). At week 8, total cholesterol and HDL-C in the pecan treatment group were significantly lower (P < .05) than in the control group (total cholesterol: 4.22 +/- 0.83 vs 5.02 +/- 0.54 mmol/L; HDL-C: 1.37 +/- 0.23 vs 1.47 +/- 0.34 mmol/L). Dietary fat, monounsaturated fat, polyunsaturated fat, insoluble fiber, magnesium, and energy were significantly higher in the pecan treatment group than in the control group. Body mass indexes and body weights were unchanged in both groups. APPLICATIONS: Pecans can be included in a healthful diet when energy intake and potential weight gain are addressed. PMID- 10719405 TI - Fat and sugar levels are high in snacks purchased from student stores in middle schools. AB - OBJECTIVE: Children consume about one third of their daily energy at school, mostly from cafeteria food and bag lunches. Students also shop at student-run stores that generate revenue for extracurricular activities; yet the nutritional value of snacks sold at student stores has not been documented to our knowledge. DESIGN: Cross-sectional study of foods sold at student stores in middle schools. SUBJECTS/SETTING: Twenty-four San Diego County (Calif) public middle schools, grades 6 through 8 (age 11 to 13), from 9 school districts. The schools represent a diversity of ethnic groups and socioeconomic levels. STATISTICAL ANALYSES PERFORMED: Descriptive statistics, Pearson product moment correlations, analysis of variance. RESULTS: Snacks averaged 8.7 g fat and 23.0 g sugar. Overall, 88.5% of store inventory was high in fat and/or high in sugar. Sugar candy accounted for one third of store sales. Chocolate candy was highest in fat content: 15.7 g. Fourteen of the 24 schools had stores that sold food and were run by student organizations. Stores were open daily for about 90 minutes; half sold food during lunch. CONCLUSIONS: Adolescents need opportunities to supplement main meals; however, student stores in middle schools sell primarily high-fat, high-sugar snacks. Key intervention possibilities include limiting sales of chocolate candy and substituting low-fat varieties of cakes, cookies, chips, and crackers. Competition with cafeterias for sales at lunchtime should be addressed. PMID- 10719406 TI - Development of an instrument to assess nutritional risk factors for children infected with human immunodeficiency virus. AB - OBJECTIVE: To produce a simple and effective instrument to evaluate and monitor the nutritional risk of children infected with the human immunodeficiency virus (HIV). DESIGN: The test instrument was developed in consultation with 5 physicians, 5 nutritionists, and 5 social workers with expertise in caring for HIV-infected children. Patient information was collected through medical record review for 19 sociodemographic, 10 anthropometric, 4 biochemical, 6 dietary intake, and 19 medical factors. As a part of routine nutrition care, anthropometric data were obtained and the caregiver was asked to complete a 3-day diet record. Also recorded were the most recent CD4+ T-cell numbers and serum HIV p24 antigen and plasma HIV RNA levels. SUBJECTS/SETTING: Thirty-nine HIV-infected children were selected using quota sampling; that is, subjects were stratified by clinical class as defined by the Centers for Disease Control and Prevention. STATISTICAL ANALYSIS: The severity or degree of potential nutritional risk in each section (anthropometric, biochemical, dietary intake, and medical data) was graded (0 to 4, 0 = low risk) and summed. Reliability of internal consistency was determined through covariance matrixes. Validity was determined through Pearson product moment correlation coefficients to measure convergent and divergent validity; predictive validity was determined using analysis of variance. Correlation for validity was compared to 6 selected dependent variables: weight for height, weight growth velocity, lean body mass, serum albumin level, CD4+ T cell numbers, and quantitative plasma HIV RNA levels. RESULTS: Of the 38 factors that were analyzed for reliability, 11 fell in the strongly reliable range: height for age, weight for age, clinical class, somatic protein stores, mid-arm circumference, weight for height, serum albumin, immunologic status, body mass index, energy intake, and opportunistic infection. CONCLUSIONS: Anthropometric, dietary intake, and medical data were reliable indicators of nutritional risk. The entire instrument was reliable after 8 of the weakest items were removed. The instrument was found to be valid and a good predictor of nutritional risk in HIV infected children. PMID- 10719408 TI - Nutrition intervention group program based on preaction-stage-oriented change processes of the Transtheoretical Model promotes long-term reduction in dietary fat intake. AB - OBJECTIVE: To develop and evaluate the long-term effectiveness of an intervention program, based on preaction-stage-oriented change processes of the Transtheoretical Model of Behavior Change, that could be delivered in a group setting to help participants lower dietary fat intake. DESIGN: An enhanced version of the nonequivalent control group experimental design was used. Entire sections of an undergraduate introductory nutrition science course were assigned to an experimental, pretest/posttest control, or posttest-only control group. Daily fat intake and stage of change of the experimental and pretest/posttest control groups were determined at the pretest and posttest and 1-year later at a follow-up test. Every 1 to 2 weeks during the study, stage of change of the experimental group was assessed. Daily fat intake of the experimental group was assessed at study midpoint. Daily fat intake and stage of change of the posttest only control group was determined at the posttest. Pretest results were used to place participants of the experimental and pretest/posttest control groups in either the preaction stage (i.e., precontemplation, contemplation, or preparation) or the action/maintenance stage. SUBJECTS/SETTING: The sample consisted of 38, 30, and 42 undergraduate students who were assigned to the experimental, pretest/posttest control, and posttest-only control groups, respectively. INTERVENTION: The experimental group participated in a group-based, dietary fat intake intervention that included a series of 11 lessons taught over a 14-week period. Each lesson was based on 1 or 2 of the preaction-stage-oriented change processes of the Transtheoretical Model. MAIN OUTCOME MEASURES: Data were evaluated to determine the effects of the intervention program on long-term dietary fat reduction and stage of change progression. STATISTICAL ANALYSIS PERFORMED: Analysis of variance, repeated-measures analysis of variance, and paired t tests. RESULTS: For pretest and posttest dietary fat intake scores, stage and time were significant, and there was a significant time-by-stage interaction. Time was significant for pretest and posttest stage scores. Subjects in the preaction-stage experimental group significantly increased their mean stage of change and reduced their fat intake between the pretest and posttest; these changes persisted for 1 year. Pretest/posttest control group participants who began in a preaction stage also significantly increased their mean stage and reduced fat intake by the posttest, but these changes did not endure until the follow-up test. APPLICATIONS/CONCLUSIONS: This intervention program produced an enduring, significant reduction in mean dietary fat consumption and a significant progression in mean stage of change of subjects in the experimental group who were in the preaction stage. It may be appropriate to design group interventions to use preaction stage processes rather than the more traditionally used action and maintenance stages change processes. PMID- 10719407 TI - Impact of acute illness on nutritional status of infants and young children with sickle cell disease. AB - OBJECTIVE: To evaluate changes in growth, nutritional status, body composition, and energy and nutrient intakes during illness and usual state of health in infants and young children with sickle cell disease. DESIGN: Sixteen children, aged 0.4 to 5.6 years, with SS type sickle cell disease (SCD-SS) were assessed at the time of hospital admission for an acute illness episode and during an 18-hour overnight follow-up visit 2 to 6 weeks after the acute illness episode when in a state of usual health. Main outcome measures included growth in height and weight compared with reference standards, body composition determined by the skinfold thickness technique and total body electrical conductivity, and dietary intake determined by 24-hour recall during hospital admission and at follow-up. RESULTS: Height, weight, and weight-for-height z scores did not differ from national reference data; triceps skinfold thickness and arm fat area z scores were less. Mean +/- standard error body fat was 15.6 +/- 2.1% at the time of hospital admission, as measured by total body electrical conductivity, and was not significantly different from the follow-up value (16.2 +/- 2.2%). Mean energy intake was 44 +/- 9% of Recommended Dietary Allowances at the time of admission and differed significantly from the follow-up value of 90 +/- 9% (P < .05). APPLICATIONS: Infants and children with sickle cell disease appear to be at nutritional risk during an acute illness episode, as indicated by body fat measures and inadequate intakes of energy and macronutrients. Energy intake may be suboptimal for several days surrounding an admission for an acute illness in children with sickle cell disease. Physicians and other health practitioners should be alert to inadequate nutrient intakes of their patients during this time period and may consider supplemental energy to avoid a potential net negative energy balance. PMID- 10719409 TI - Nutrition and multifetal pregnancy. AB - Largely because of assisted reproduction, the rate of multifetal pregnancy is rising rapidly in the United States. Accordingly, dietitians are increasingly being called upon to provide nutrition services for these high-risk pregnancies. This article gives an overview of the incidence of and risks associated with multifetal pregnancy and reviews studies that contribute to our knowledge of nutrition and multifetal pregnancy. Practice guidelines for promoting healthy outcomes based on the best available scientific data are suggested. Guidelines for weight gain for twin and triplet pregnancy, dietary intake, and supplement use are included. Suggested practice guidelines for multifetal pregnancy include a positive rate of weight gain early in pregnancy, the use of prepregnancy weight status to determine total weight gain goals in twin pregnancy, a 50-lb weight gain goal for triplet pregnancy, and higher minimal number of servings of foods from several of the Food Guide Pyramid groups. The need for additional information on the effects of nutritional status on the course and outcome of multifetal pregnancy is critical. Preliminary evidence of the benefits of nutrition services suggests that both the incorporation of dietetics services into care programs and additional research on nutrition and multifetal gestation are warranted. PMID- 10719410 TI - The 5 A day Virtual Classroom: an on-line strategy to promote healthful eating. AB - Communications technology can help stimulate youth to become involved in health promotion. This article reports on an innovative, Internet-based nutrition program that encouraged children to be advocates for policies that promote eating more fruits and vegetables, the 5 A Day Virtual Classroom. Through this program, students from across the United States discussed the recommendation of 5 A Day at the same time in a classroom without walls. In September 1997 children were asked, "If you were President Clinton, how would you get kids across the country to eat 5 A Day?" Based on content analysis of responses, this article suggests strategies that policymakers could use to encourage children to consume more fruits and vegetables. Approximately 2,600 students participated; 635 entries and 910 suggestions were received. The suggestion categories cited most often were mass media (19.8%), economic issues (15.4%), and social influence (13.8%). The most frequently mentioned specific ideas were to reward children for eating fruits and vegetables and to use presidential authority. Some regional, age, and gender patterns were found. Findings support the potential impact on health education of the 5 A Day Virtual Classroom and of interventions based on communications technology. PMID- 10719411 TI - Psychosocial and lifestyle factors associated with insufficient and excessive maternal weight gain during pregnancy. PMID- 10719412 TI - Micronutrient supplementation does not attenuate seasonal decline of immune system indexes in well-nourished elderly women: a placebo-controlled study. PMID- 10719413 TI - Nutrition messages in a sample of children's picture books. PMID- 10719414 TI - Adolescent health and nutrition: a survey of perceived knowledge and skill competencies and training interests among dietitians working with youth. PMID- 10719415 TI - Serum lipid response to n-3 fatty acid enriched eggs in persons with hypercholesterolemia. PMID- 10719416 TI - Assessment of general nutrition knowledge of nurse practitioners in New England. PMID- 10719417 TI - Zinc: what role might supplements play? PMID- 10719418 TI - [Mono-polychemotherapeutic treatment? The opportunities and limits]. PMID- 10719419 TI - [The prevention of vomiting due to antineoplastic chemotherapy in the elderly cancer patient]. PMID- 10719421 TI - [Something has changed in the treatment of non-small-cell bronchogenic carcinoma in the elderly]. PMID- 10719420 TI - [The medical treatment of advanced non-small-cell lung carcinoma (adv NSCLC). The correlation with multidimensional assessment]. PMID- 10719422 TI - [Lung tumors in the geriatric patient]. PMID- 10719423 TI - [Serum chromogranin A in prostate cancer. Preliminary results]. PMID- 10719424 TI - [The multidisciplinary treatment of colorectal cancer in the elderly patient]. PMID- 10719425 TI - [The radiotherapeutic treatment of rectal tumors in the elderly patient over 70]. PMID- 10719426 TI - [The adjuvant therapy of colonic carcinoma in old age]. PMID- 10719427 TI - [The chemotherapy of advanced colorectal carcinoma in the elderly patient. The new oral fluoropyrimidines]. PMID- 10719429 TI - [The development of the first hospice organization in central Italy one year after its starting to operate]. PMID- 10719428 TI - [The prognostic factors in cancer patients with the disease in an advanced phase]. PMID- 10719430 TI - Pain from bone metastases: recent therapeutics positioning. PMID- 10719431 TI - [Domiciliary care for cancer patients in old age]. PMID- 10719432 TI - [The net clinical benefit]. PMID- 10719433 TI - [Nutritional therapy]. PMID- 10719434 TI - [The role of the nurse in the hospice]. PMID- 10719435 TI - [Infusion systems]. PMID- 10719436 TI - [Bioethics: what is the role of the physician and the professional nurse in the approach to the elderly cancer patient?]. PMID- 10719437 TI - [The handling of antineoplastic agents and Dlgs 626/94. Decreto legislativo (legislative decree)]. PMID- 10719438 TI - Treatment of arterial hypertension in the elderly using enalapril. AB - BACKGROUND: Arterial pressure increases with age and is the most common chronic disease in the elderly, men and women alike. At present arterial hypertension is considered a social disease as it poses great health, economic and social problems. METHODS: A group of 50 patients of both sexes, between 66 and 83 years of age, suffering from essential arterial hypertension and treated with Enalapril administered at doses varying from 10 to 20 mg/day, for a period of 18 months, has been studied. RESULTS AND CONCLUSIONS: The results of this study showed that blood pressure values of almost all the patients treated normalised and left ventricular hypertrophy decreased in all patients with this complication. PMID- 10719439 TI - [TTV: a new hepatitis virus? Transfusion-transmitted virus]. AB - Following the discovery of hepatitis C virus (HCV), it was clear that a proportion of cases with acute and chronic hepatitis are still negative for all known viral markers, which prompted investigations to search for new hepatitis agents. In 1997 investigators in Japan isolated a DNA clone of a novel human virus using a representation difference analysis from serum samples from a patient (TT) with post-transfusion hepatitis of unknown etiology and designated TTV for transfusion-transmitted virus. TTV DNA is common in populations at increased risk for infection with hemophilia or maintenance hemodialysis and abusers of intravenous drugs. As a nonenveloped virus with a linear, single stranded genomic DNA, TTV resembles the Parvoviridae among known animal viruses. TTV has been reported in association with post-transfusion and acute and chronic hepatitis of unknown etiology. Phylogenetic analysis indicated six different genotypes of TTV with distinct subtypes [correction of sottotypes]. Viral DNA can be detected in plasma but also in liver tissue of infected subjects, suggesting that it is hepatotropic. TTV can be transmitted parenterally by blood and blood products and probably also non-parenterally by a fecal-oral route. TTV DNA is present in a large proportion of patients with different forms of non-A-G hepatitis. A new simple genotyping assay based on a restriction fragment length polymorphism of TTV was developed. This system will provide the framework for future detailed epidemiological and clinical investigations. PMID- 10719441 TI - [The Brugada syndrome. A predicted sudden juvenile death]. AB - This review deals with the clinical, basic and genetic aspects of recently highlighted form of idiopathic ventricular fibrillation known as the Brugada syndrome. The available data suggest that the Brugada syndrome is a primary electrical disease resulting in abnormal electrophysiological activity in the right ventricular epicardium. Diagnosis is based on the presence of an ST elevation in the anteroseptal territory and a right branch block. No underlying dysarrhythmic condition or arrhythmogenic heart disease can be detected. The available data suggest that less of the action potential dome in the right ventricular epicardium but not endocardium underlies the ST segment elevation seen in the Brugada syndrome and that electrical heterogeneity within right ventricular epicardium leads to the development of closely coupled premature ventricular contractions via a phase 2 reentrant mechanism that then precipitates ventricular tachycardia/ventricular fibrillation. Brugada syndrome is a recently discovered hereditary condition with a probably underestimated prevalence. Systematic family studies have demonstrated an autosomal dominant inheritance. The characteristic electrocardiographic anomalies can be transitory and may be unmasked by sensitization tests. The only currently treatment is the implantable defibrillator programmed to prevent sudden death by ventricular fibrillation and it is indicated in symptomatic patients and should be considered in asymptomatic patients in whom ventricular tachycardia/ventricular fibrillation is inducible by electrophysiologic study. PMID- 10719440 TI - [Doping and sports]. AB - Doping is widely known as the use of banned substances and practices by athletes in an attempt to improve sporting performances. The term doping likely derives from "dope", an ancient expression referred to a primitive alcoholic drink that was used as a stimulant in South African ceremonial dances; gradually, the term was extended and finally adopted his current significance. There are at least two essential reasons to support the fight against doping: the potential harmful effects on athletes and the depth corruption of the fair competition. An exhaustive list of banned substances and methods has been drawn by the International Olympic Committee and further accepted by other International Sport Authorities and Federations. This list, regularly updated, is basically divided into doping substances (stimulants, narcotic analgesics, anabolic agents, diuretics, peptide and glycoprotein hormones and analogues), doping methods (blood doping, pharmacological, chemical and physical manipulation) and drugs subjected to certain restrictions (alcohol, marijuana, local anesthetics, corticosteroids and beta-blockers). Although there might be some medical conditions, which could legitimate the need of these substances or methods, there is no place for their use in sport. Thus, an athlete's consume of any of these substances or methods will result in disqualification. Aim of the present review is to provide a synthetic description of both the desirable effects and the potentially harmful consequences of the use of some of the major doping substances and methods. PMID- 10719442 TI - [Hyperhomocysteinemia and arterial or venous thrombosis. Retrospective study of 75 cases]. AB - OBJECTIVE: Previous studies suggest that hyperhomocysteinemia may be a risk factor for arterial and venous thrombosis. We retrospectively analyzed data from 75 patients with thrombosis. PATIENTS AND METHODS: Thirty-four patients had arterial thrombosis, 22 venous thrombosis and 19 venous and arterial thrombosis. Of the 75 patients (49 men and 26 women, mean age 49 +/- 15 years) about two thirds had recurrent episodes of thrombosis. RESULTS: Hyperhomocysteinemia was defined as serum homocysteine level above 14.1 mumol/l (mean + 2.7 SD in healthy subjects) and was found in 67 patients (89%, CI95% = 80-95). Mean total homocysteine concentration was 21.6 +/- 13.6 mumol/l for the 75 patients. About half of the patients were smokers, 35% had hypertension and 25% high serum cholesterol. There was no significant relationship between serum homocysteine level and smoking status, hypertension or serum cholesterol level. Ten patients (13%, CI95% = 7-23) had low serum cobalamin (< 150 pmol/l). Serum folates were < or = 10 nmol in 41% of the patients in the arterial thrombosis group (CI95% = 25 59), in 27% in the venous thrombosis group (CI95% = 11-50), and in 31% in the arterial and venous thrombosis group (CI95% = 13-57). Thirteen patients received vitamin B supplementation. Hyperhomocysteinemia decreased in 12/13 patients (CI95% = 64-100) and returned to normal values in 9/13 patients (69%, CI95% = 38 91). CONCLUSION: Our data show that hyperhomocysteinemia is frequently found in arterial and venous thrombosis. Further studies are needed to determine the clinical impact of homocysteine lowering therapy. PMID- 10719443 TI - [Allergy to macrolides. 21 cases]. AB - OBJECTIVE: Allergic drug reactions to macrolides are extremely rare and there is little information in the literature concerning relevant diagnostic tests. PATIENTS AND METHODS: Twenty-one patients were recently seen for assumed allergies (principally urticaria) to diverse macrolides. Skin tests (prick and intradermal tests) were performed with injectable forms of spiramycin and erythromycin. Seventeen out of 21 patients were provoked under strict hospital surveillance. RESULTS: Only 3 patients had a positive provocation test and were thus truly allergic (to spiromycin). They had positive skin tests to both macrolides tested. CONCLUSION: Most hypersensitivity reactions to macrolides are therefore diagnosed with provocation tests. PMID- 10719444 TI - [Amyotrophic lateral sclerosis manifesting as cognitive disorders. Value of brain perfusion scintigraphic tomography in intensive care]. AB - BACKGROUND: Cognitive disorders have been described in amyotrophic lateral sclerosis, but usually after the diagnosis has ben established. CASE REPORT: A 57 year-old man was intubated for acute respiratory distress subsequent to pneumonia and diaphragm palsy. He had a 2-year history of drug-resistant depression and deterioration of cognitive functions. A pyramidal syndrome associated with biopsy proven chronic neurogenic atrophy led to the diagnosis of amyotrophic lateral sclerosis. The electromyogram did not contribute to diagnosis. Brain MRI only evidenced moderate bilateral frontal-temporal atrophy. Brain SPECT demonstrated major perfusion defects in the frontal lobes. DISCUSSION: This patient had amyotrophic lateral sclerosis and frontal-temporal dementia with an unusually late onset clinical presentation: cognitive disorder was the inaugural sign. Brain SPECT and muscle biopsy enabled us to identify the cortical and peripheral motor neurone involvement in this uncooperative intensive care patient totally dependent on mechanical ventilation. PMID- 10719445 TI - [Imputable cutaneous eruption caused by ticlopidine after implantation of a coronary endoprosthesis: why not continue treatment?]. AB - BACKGROUND: The decision to interrupt a ticlopidine regimen in a patient who develops an adverse effect can be particularly difficult when discontinuing the drug could lead to a high risk situation. CASE REPORT: A 64-year old patient developed a skin rash after taking ticlopidine for coronary artery stenting. Stopping ticlopidine could have led to stent occlusion and no alternative therapy seemed to be suitable. We therefore decided to carry on the treatment under close clinical surveillance. The skin signs rapidly resolved. DISCUSSION: This cases shows that ticlopidine may be continued in patients who develop an adverse skin reaction. The rapid involution of the cutaneous signs in our patient demonstrated that the risk of discontinuing treatment can be greater than that of continuing. PMID- 10719446 TI - [Systemic amyloidosis: localization in the gallbladder]. PMID- 10719447 TI - [Torsade de pointes after poisoning with fluoxetine alone]. PMID- 10719449 TI - [Medenox study]. PMID- 10719448 TI - [Value of immunoglobulins in Schulman fasciitis]. PMID- 10719450 TI - [Quality of life measure in cancerology]. AB - CONCEPT: Quality of life assessment in cardiology has become an important point in the overall evaluation of anticancer treatments. Quality of life is a multidimensional subjective concept usually assessed by self-administered questionnaires with a linear analogous design. Multiple choice questions can also be used to explore several dimensions of quality of life. QUESTIONNAIRES: No one questionnaire is ideal. Two appear to be most widely used worldwide. The main body of these modular questionnaires is completed by specific sections dealing as needed with designated organs or treatment toxicity. Data interpretation is particularly difficult and must take into consideration the fact that most currently used questionnaires measure health in general rather than the philosophical concept of quality of life. CLINICAL APPLICATION: Use of quality of life questionnaires varies greatly. These tools can be used to estimate the impact of public health measures or treatments in general. They are most widely used to date as assessment tools for therapeutic trials or to provide secondary criteria in trials where survival is the main outcome criteria. In this case, they provide a means of comparing symptomatic treatment and anticancer treatments or two different anticancer treatments. Studies are under way to use quality of life scales in everyday patient care. PERSPECTIVES: Certain authors have attempted to combine quality and quantity of life in a single tool in order to obtain a global index which could be used to compare large groups of patients. Different tools are still in the development phase. PMID- 10719451 TI - [Drug addiction: sentinel and partnership role of the hospital]. PMID- 10719452 TI - [Allergy to macrolide antibiotics. Review of the literature]. AB - MACROLIDE CLASSES: Macrolides are characterized by their basic structure made up of a lactonic cycle with 2 osidic chains. They are classified according to the number of carbon atoms in the cycle: 14-membered macrolides (erythromycin, troleandomycin, roxithromycin, dirithromycin, clarithromycin), 15-membered macrolides (azithromycin) and 16-membered macrolides (spiramycin, josamycin, midecamycin). MACROLIDE ALLERGY: Allergy to macrolides is extremely rare (0.4% to 3% of treatments). The little information available in the literature is insufficient to establish the usefulness of diagnostic tests. An immediate IgE dependent hypersensitivity has been shown with erythromycin in some cases but the mechanism remains unknown and skin tests are quite often negative. Clinical manifestations are the same as those encountered with beta-lactams. It would appear that macrolide allergies are unlikely to be class allergies. This is important as eviction advice could be limited to the single causal macrolide. PMID- 10719453 TI - [Neurologic forms of sarcoidosis]. AB - BACKGROUND: Neurosarcoidosis is an uncommon but severe, sometimes life threatening, manifestation of sarcoidosis. Signs of neurological involvement usually are seen in patients known to have active disease. Strictly neurological forms are seen in less than 10% of cases. CLINIC: Neuropsychic manifestations are the most common clinical signs, independent of corticosteroid therapy or neuroendocrine involvement, epileptic seizures, and signs related to hypocephalia. DIAGNOSIS: Diagnosis of systemic sarcoidosis is confirmed on the basis of clinical arguments and laboratory findings favoring neurosarcoidosis. Key investigations include angiotensin converting enzyme assay in cerebrospinal fluid, and brain stem magnetic resonance imaging with gadolinium injection. Nerve biopsies may be needed in certain cases. TREATMENT: Corticosteroid therapy is given as first line treatment with a satisfactory effect in most cases. Immunosuppressors may be added in case of failure. PMID- 10719454 TI - [Characteristics of pain and pain management in the elderly]. AB - A COMMON SYMPTOM: Pain is a common and underestimated problem in older people who are likely to suffer from many acute and chronic conditions. DIFFICULT ASSESSMENT: Clinical assessment of pain often depends on the patient's ability to communicate his or her experience. If self-assessment of pain is not possible, behavioral hetero-evaluation instruments have been built for easy pain assessment by caregivers. ADJUSTED PAIN MANAGEMENT: As for younger patients, the most common strategy for pain management is the use of analgesic drugs. Special care should be taken however when such drugs are initiated in the elderly because increased sensitivity, prolonged drug half-life and adverse effects and drug interactions are more likely. PMID- 10719456 TI - [Diagnostic and interventional radiology in the 21st century]. PMID- 10719455 TI - [Posture and aging. Current fundamental studies and management concepts]. AB - FUNDAMENTAL IMPORTANCE OF POSTURE: In the elderly subject, preservation of posture is fundamental to maintaining functional independence. In recent years, there has been much progress in our understanding of the mechanisms underlying strategies used to control equilibrium in the upright position. Physiological aging, associated with diverse disease states, dangerously alters the postural function, particularly anticipated adjustments which allow an adaptation of posture to movement. CLINICAL ASSESSMENT OF POSTURE: Several tests have been developed to assess posture in the elderly subject, particularly the time it takes to start walking. We selected certain tests which can be used in everyday practice to predict falls: the stance test, the improved Romberg test, the "timed get up and go test", measurement of walking cadence, assessment of balance reactions, sitting-standing and standing-sitting movements and capacity to get up off the floor. PATIENT CARE: Elderly patients with equilibrium disorders can benefit from specific personalized rehabilitation protocols. Different techniques have been developed for multiple afferential stimulation, reprogramming postural strategies, and correcting for deficient motor automatisms. PMID- 10719457 TI - [Rational diagnostic imaging of pelvic and acetabulum injuries]. AB - In spite of the widespread availability of CT scanners, conventional X-ray radiographs remain the basic imaging modality in patients with pelvic and/or acetabular trauma. However, the extent of their use will depend on local utilities (e.g., availability of CT scanners) and on the patient's clinical condition. Regarding the inaccuracy of conventional radiography in the diagnosis of injuries of the dorsal pelvic ring and of the acetabulum, computed tomography represents the most important imaging modality in the clinically stable patient. CT provides an exact staging of the extent of trauma and allows for differentiation of pelvic instabilities. CT clearly demonstrates the severity of acetabular trauma and is superior in the detection of local complicating factors, i.e., impressions fractures and (sub-)luxations of the femoral head as well as free intraarticular fragments. CT findings provide the basis for definite treatment regimens of the injured patient. By extension of the examination, all relevant organs and systems (craniospinal, cardiovascular, gastrointestinal, respiratory, genitourinary) can be imaged during one session. The speed of spiral CT scanners and their diagnostic accuracy will play a major role in the management of, especially, polytraumatized patients. The indication for angiography with the option of therapeutic embolization exists if a pelvic bleeding persists even after reposition and operative fixation of the injury. PMID- 10719458 TI - [Percutaneous interstitial thermotherapy of malignant liver tumors]. AB - PURPOSE: Description of local ablative techniques such as laser-induced thermotherapy (LITT) and radiofrequency ablation in the percutaneous interstitial thermotherapy for malignant liver tumors. PATIENTS AND METHODS: MR-guided LITT is currently performed by means of implantable percutaneous catheter systems. CT is used to control the insertion of the catheter. Irrigated administration systems are available both for LITT and for radiofrequency therapy. RESULTS: At present LITT enables a local tumor control of 97.2% for localized liver metastases without extrahepatic spreading patterns. In a group of 381 patients the average survival times were 45.7 months for patients with liver metastasis, 42.7 months for those with colorectal liver metastases, and 32 months for HCC tumors. The data for radiofrequency ablation confirm the high value for tumor control of hepatocellular carcinomas with poorer results for liver metastases. CONCLUSIONS: Percutaneous, MR-guided LITT permits good tumor control of liver metastases smaller than 5 cm and less than 5 in number. For HCC, at present percutaneous injection of alcohol, radiofrequency ablation, and LITT are equally effective. PMID- 10719459 TI - [Left ventricular heart volume determination with fast MRI in breath holding technique: how different are quantitative heart catheter, quantitative MRI and visual echocardiography?]. AB - GOAL: Comparison of fast MRI, echocardiography (Echo), and ventricular angiography (Cath) in the assessment of left ventricular global function. METHODS: Fast MRI in short axis plane, biplane Cath, and 2D Echo were performed in 62 patients [35 coronary artery diseases, 16 acquired valvular diseases (VD), 9 dilated cardiomyopathies (DCM), 1 congenital heart disease and 1 heart transplantation]. Enddiastolic (EDV), endsystolic (ESV), stroke volumes (SV), cardiac output (CO), and ejection fraction (EF) were compared in MRI and Cath. EF was visually estimated in 2D Echo by an experienced observer. RESULTS: In comparison to MRI, Cath overestimated EF by 8.4%, and Echo underestimated EF by 5.6%. The limits of agreement between MRI and Cath in EF were +/- 23.8%, between MRI and Echo +/- 18%, and between Echo and Cath +/- 19.4%. Significant differences were found between Cath and MRI in EDV, SV, and CO, but not for ESV. The best agreement in EF was found in the group with DCM, the worst in the group with VD. CONCLUSION: Important systemic and random errors were found in the comparison of MRI, Echo, and Cath. For therapy decision and follow-up, the methods should not be exchanged unscrupulously. PMID- 10719460 TI - [Spondylolysis in the developmental stage: diagnostic contribution of MRI]. AB - PURPOSE: To assess the value of MR imaging in demonstrating ongoing spondylolysis in adolescents. METHODS: MRI was performed in 9 juvenile patients (3 female, 6 male aged 8-16 years; mean 12.5 y) with pain during hyperextension. In 6 patients a CT scan and in 5 a plain film was available. RESULTS: In all patients bone marrow edema was found in the pars interarticularis and the pedicle, which was bilateral in 4 patients. In 7/9 cases the L5 vertebra was affected, in 2/9 cases spondylolysis was found in L4. In 3 cases the edema reached the middle third of the vertebral body and a tumor was suspected. In all CT scans a bilateral incomplete or complete cleft in the pars inter-articularis was found. In 4/6 CT scans a sclerosis was seen in the area of the bone marrow edema. Only in 1/5 plain films was there a suspicion for a spondylolysis, four examinations were completely normal. CONCLUSIONS: To eliminate underlying causal conditions of spondylolysis and to install specific therapy, early diagnosis is mandatory. MR imaging should be the first and only imaging modality in young patients with low back pain during and after exercise and pain with hyperextension. Bone scans and CT scans should be avoided due to irradiation, plain films usually do not reveal pathological findings in developing sponylolysis. PMID- 10719461 TI - [Perfusion parameters in MRI diagnosis of pancreas transplants]. AB - PURPOSE: Evaluation of the role of perfusion parameters in the detection of circulatory disturbance and chronic rejection in patients after pancreas transplantation. MATERIALS AND METHODS: 70 examinations of 39 patients after pancreas transplantation were performed. Using a dynamic gadolinium-enhanced Turbo-FLASH-sequence, we evaluated the perfusion parameters in a group of patients with chronic rejection, with circulatory disturbance, and in a control group with normal organ function. RESULTS: There were statistically significant differences of the perfusion parameters in patients with chronic rejection and circulatory disturbance compared to the control group. CONCLUSION: Dynamic MRI can help detect patients with chronic rejection and circulatory disturbance and should therefore be part of the routine follow-up in patients after pancreas transplantation. PMID- 10719462 TI - [Multi-sequential arterial chemoembolization of advanced hepatocellular carcinomas: computerized tomography follow-up parameters for evaluating effectiveness of therapy]. AB - PURPOSE: An evaluation of clinical and computed tomography parameters for the transarterial chemoembolization (TACE) of hepatocellular carcinoma (HCC). MATERIAL AND METHODS: Retro- and prospective analysis of 123 TACE procedures in 37 patients (mean age: 63.3 years). RESULTS: Overall mean survival rate for all treated patients was 785 days with a mean value of 387 days. The 1 year survival rate was 62%. Quantitative tumor volumetry revealed a significant difference in survival rate with a mean value of 678 days for patients with a reduction of tumor volume between 0 and 50%, 976 days for a volume reduction of 51-100% and 277 days for an increase in tumor volume during therapy. Patients with a high lipiodol retention presented improved survival data (902 days) versus patients with a low lipiodol retention (513 days). Homogeneous retention of lipiodol was a positive factor on survival as compared to an inhomogenous form of retention. CONCLUSION: In patients with a positive therapy response after TACE the prognosis can be improved by repeated embolization. PMID- 10719463 TI - [Superparamagnetic iron oxide enhanced MR angiography of the portal vein system]. AB - PURPOSE: Superparamagnetic iron oxide (SPIO) is used for the detection of focal liver lesions. The current study examines whether the blood-pool effect of an approved substance is adequate for contrast-enhanced MR angiography of the portal venous system. METHODS AND MATERIALS: Fifteen patients [9 women, 6 men, mean age 57 (34-73)] were examined with a breathhold 3D-FISP sequence (TR/TE/FA; 5 ms/2 ms/25 degrees) at 1.5 Tesla (Vision; Siemens, Erlangen) during the last third of a half-hour Infusion of a standard dose (15 mumol Fe/kg) of AMI-25 (Endorem, Guerbet Co., France). The image quality of source images and 3D reconstructions was evaluated by two examiners using a 3-point scale. RESULTS: Large vessels (portal vein, left and right branch of the portal vein, superior mesenteric vein, splenic vein) are visualized well or adequately in 80-93% of the cases, while this was achieved in only 53-60% of the small veins (vessels of the first order emptying into the superior mesenteric vein and the spleen). Venous arcades of the mesenterium are not visible by this technique. CONCLUSION: SPIO-supported MRA of the portal venous system yields usable results in the majority of patients. A possible indication is preinterventional angiography together with an SPIO examination for the detection of focal liver lesions. PMID- 10719464 TI - [Value of negative oral contrast media in MR cholangiopancreatography (MRCP)]. AB - PURPOSE: Evaluation of the use of a negative oral contrast material in MR cholangiopancreatography (MRCP). MATERIAL AND METHODS: We performed MRCP in single-shot technique (TSE, TR = 2800 ms, TE = 1100 ms, ETL = 64) in 38 patients before and 20-30 min after oral administration of 300-600 ml of a negative oral contrast material. The visualization ducts and details important for the diagnosis was evaluated in a blinded manner. Ductal diameters were measured. Both sets of images were evaluated qualitatively. RESULTS: The ductal diameters did not change after administration of oral contrast material. In 1/3 of all cases the ductal structures were superimposed by a high signal intensity of fluid in the gastrointestinal tract, especially in the tail of the pancreas. After administration of oral contrast material only in 3 patients could a complete visualization of the ducts not be achieved. In 5 cases, details relevant for the diagnostic decision could be seen only on post-contrast images. The anatomic orientation was not compromised by the absence of signal in the gastrointestinal tract. CONCLUSION: Negative oral contrast material should be given before performing a MRCP to provide non-superimposed visualization of the bile and pancreatic ducts. There is no negative influence of the oral contrast material on the diameter of the ducts. PMID- 10719465 TI - [Stimulated acoustic emissions with the ultrasound contrast medium levovist: a clinically useful contrast effect with liver-specific properties]. AB - PURPOSE: The purpose of this study was systematically to investigate stimulated acoustic emission (SAE) with the microbubble contrast agent Levovist (Schering AG, Berlin) in vivo with regards to reproducibility, distribution in various organs over time, dependence on technical factors, and influence on the delineation of focal liver lesions. PATIENTS AND METHODS: 2 intravenous injections of 1 g of Levovist were given to 2 dogs and 1-6 injections of 2.5 g Levovist to 5 healthy volunteers and 37 patients. The liver, spleen, large abdominal vessels, and kidney were intermittently scanned for up to 30 min. Studies were evaluated for the presence of SAE signals by 2 observers. In 20 patients with focal liver lesions (15 with metastases, 4 haemangiomata, 1 hepatocellular carcinoma, and 1 cyst) the influence on lesion visualization was also assessed. RESULTS: SAE effects, lasting up to 30 minutes, were seen in all subjects in the liver and spleen. Vascular and renal SAE signals were noted shortly after injection, lasting up to 6 minutes. SAE was absent or markedly reduced in focal liver lesions, which were seen as colour voids. This increased the conspicuity of focal lesions, and in 5 patients additional metastases were detected that could not be delineated on B-mode alone. CONCLUSION: A liver- and spleen-specific late phase of Levovist can be consistently demonstrated using SAE and the effect increases the conspicuity of focal liver lesions. PMID- 10719466 TI - [Radiographic image magnification as quality control of microcalcification imaging within the scope of histopathological control of breast biopsy tissue]. AB - PURPOSE: Evaluation of specimen radiography as a quality control for the detection of microcalcifications by the pathologist during histopathological examination of breast biopsies. MATERIALS AND METHODS: 29 breast biopsy specimen with microcalcifications were radiographed using the magnification (x4) mammography System DIMA Plus MI I before and after the histopathological examination. Tissue with detected microcalcifications after the histopathological examination was reexamined histologically. RESULTS: In 55% (16/29) of all specimen we could identify missed microcalcifications. In 5 specimens the microcalcifications were missed completely. The radiologically detected areas of residual microcalcifications were reexamined histologically and in one case invasive parts of a carcinoma were detected, which primarily had been diagnosed as an in-situ carcinoma. In 6 cases the radiographs of the intraoperative specimen radiography were sent to the pathologist together with the biopsy. In these cases the rate of complete detection of the microcalcifications increased from 39% to 66%. CONCLUSION: Specimen radiography should be performed not only intraoperatively but also as part of the histopathological examination. PMID- 10719467 TI - [Prognostic value of hepatobiliary functional scintigraphy in diagnosis and after care of biliary atresia]. AB - AIM: To investigate the prognostic relevance of hepatobiliary scintigraphy (HBS) in newborns suffering from biliary atresia (BA) for establishing the primary diagnosis and in the postoperative follow-up after portoenterostomy (Kasai). METHODS: Twenty newborns with direct hyperbilirubinemia and 6 children after operative treatment of BA (Kasai) underwent HBS with Tc-99m-DEIDA. In patients without intestinal drainage, hepatocellular extraction was estimated visually and calculated semiquantitatively by means of liver/heart-ratio 5 min p.i. RESULTS: 10/20 patients with hyperbilirubinemia did not display biliary drainage; 6 had BA, 3 intrahepatic hypoplasia, and one showed a bile plug syndrome. 4/6 with BA but none of the 4 children with diagnoses other than BA presented with a good extraction. All of the 4 children with BA, who had either pre- or postoperatively a bad extraction, needed liver transplantation due to liver failure. Both of the two newborns with BA and favourable outcome after Kasai had a good extraction in the preoperative HBS and demonstrated good intestinal drainage in the postoperative scan. CONCLUSION: HBS rules out BA with high accuracy by demonstrating drainage of bile into the intestine. In newborns without drainage a good extraction favours the diagnosis of BA. In newborns with BA a bad extraction seems to indicate a poor postoperative prognosis after Kasai operation. HBS might therefore help to select those children who will not benefit from portoenterostomy. Postoperatively, HBS can easily and quickly confirm the successful hepatobiliary anastomosis by demonstrating biliary drainage into the intestine. PMID- 10719468 TI - [Decision analysis in radiology using Markov models]. AB - Markov models (Multistate transition models) are mathematical tools to simulate a cohort of individuals followed over time to assess the prognosis resulting from different strategies. They are applied on the assumption that persons are in one of a finite number of states of health (Markov states). Each condition is given a transition probability as well as an incremental value. Probabilities may be chosen constant or varying over time due to predefined rules. Time horizon is divided into equal increments (Markov cycles). The model calculates quality adjusted life expectancy employing real-life units and values and summing up the length of time spent in each health state adjusted for objective outcomes and subjective appraisal. This sort of modeling prognosis for a given patient is analogous to utility in common decision trees. Markov models can be evaluated by matrix algebra, probabilistic cohort simulation and Monte Carlo simulation. They have been applied to assess the relative benefits and risks of a limited number of diagnostic and therapeutic procedures in radiology. More interventions should be submitted to Markov analyses in order to elucidate their cost-effectiveness. PMID- 10719469 TI - [Indications and rate of repeat studies in the color duplex department of a radiology university clinic]. AB - PURPOSE: Who refers patients for a color duplex ultrasonography (CDUS) in the routine work of a radiological university department and how frequent are repeat examinations? MATERIAL AND METHODS: 1110 patients had at least one CDUS examination between 5/97 and 5/98. The specialities of the referring physicians/primary examiners and the inpatient or outpatient status of the patients were documented at the first consultation. Additionally the patients were asked by the medical staff, whether, how often, and by whom ultrasonographic, CT, or MR examinations of the same organ system had been carried out during the last 4 weeks without evidence of any clinical changes. RESULTS: 97% of the 1118 patients were referred by one of the university departments. The arterial system was exclusively investigated in 58% and the venous system in 29% of the cases. 81% of the 651 arterial examinations were requested by the Departments of Vascular/Cardiac Surgery and Radiology. 75 repeated ultrasonographic examinations were documented in 67 (6%) of the 1118 consultations. Repetitions were documented in 8% of the arterial and in 3% of the venous examinations. X-ray angiographies were already done or planned in 105 of the 625 arterial CDUS (16%). Ultrasonographic referrals with parallel phlebographies (1% of 320 venous indications) as well as parallel CT and MR examinations (1% of the 1118 consultations, respectively) were the exception. CONCLUSIONS: In the described setting, CDUS was mainly used to assess the arterial vascular system prior to vascular surgery and radiological interventions. Repeat ultrasonographies alone (6%) and parallel examinations altogether (14%) were observed less frequently than expected. PMID- 10719470 TI - [Stent angioplasty of pelvic artery stenosis with MRI control: initial clinical results]. AB - PURPOSE: To investigate the feasibility of MR-guided stent angioplasty of iliac artery stenoses under passive visualization. MATERIAL AND METHODS: Three patients with short, concentric stenoses of the iliac arteries were enrolled. The vascular interventions were performed on a 1.5 T MR scanner (Magnetom Symphony, Siemens, Erlangen, Germany). Stents, guidewires, and balloon catheters were visualized on the basis of susceptibility artifacts. Contrast-enhanced MR angiography (ceMRA) was used to localized the stenosis prior to stent deployment. Nitinol stents were placed under MR-guidance using a fast 2D gradient echo technique. Balloon dilatiation was performed with an angioplasty catheter inflated with diluted gadolinium-DTPA. Postinterventional results were evaluated by ceMRA, DSA, and Doppler indices. RESULTS: Position of the stent, stent deployment, and balloon dilatation were depicted by MR. All stents were correctly placed within the stenosis. Stent positions as monitored by MRI were identical to those seen on DSA images. All patients were treated successfully by the MR-guided intervention. CONCLUSION: An MR-guided stent angioplasty of simple iliac artery stenosis is feasible under passive visualization. PMID- 10719471 TI - [Triple umbilical cord wrap-around as etiology of generalized sinus thrombosis]. PMID- 10719472 TI - [Adrenal vein thrombosis in heparin-induced thrombocytopenia]. PMID- 10719473 TI - [Adenoma of the middle ear: CT and MRI findings]. PMID- 10719474 TI - A computer-assisted training/monitoring system for TURP structure and design. AB - A generic framework for a computer-assisted system for both soft tissue endoscopic surgery and surgical training is being researched and developed. The concept demonstrator is a specific system for transurethral prostatic resection (TURP). The main novelty of the research is that it is not confined to an in vitro trainer system. An in vivo monitoring version of the system, for use in the operating theater, is also being researched. This paper presents the framework's structure and design using the United Modeling Language. It also discusses and justifies the underlying information technologies chosen to implement this approach. Object-oriented concepts and well-proven mathematical tools have been adopted as the foundation of this research and development. The rationale for having chosen such tools is presented. The objectives are to arrive at a system which is modular, general, and reusable. PMID- 10719475 TI - CRIGOS: a compact robot for image-guided orthopedic surgery. AB - The CRIGOS (compact robot for image-guided orthopedic surgery) project was set up for the development of a compact surgical robot system for image-guided orthopedic surgery based on user requirements. The modular system comprises a compact parallel robot and a software system for planning of the surgical interventions and for supervision of the robotic device. Because it is not sufficient to consider only technical aspects in order to improve in clinical routine the therapeutic outcome of conventional interventions, a user-centered and task-oriented design process has been developed which also takes human factors into account. The design process for the CRIGOS system was started from requirement analysis of various orthopedic interventions using information gathered from literature, questionnaires, and workshops with domain experts. This resulted in identification of conventional interventions for which the robotic system would improve the medical and procedural quality. A system design concept has been elaborated which includes definitions of components, functionalities, and interfaces. Approaches to the acquisition of calibrated X-rays will be presented in the paper together with design and evaluation of a first human computer interface. Finally, the first labtype parallel robot based on low-cost standard components is presented together with the first evaluation results concerning positioning accuracy. PMID- 10719476 TI - Techniques for development of safety-related software for surgical robots. AB - Regulatory bodies require evidence that software controlling potentially hazardous devices is developed to good manufacturing practices. Effective techniques used in other industries assume long timescales and high staffing levels and can be unsuitable for use without adaptation in developing electronic healthcare devices. This paper discusses a set of techniques used in practice to develop software for a particular innovative medical product, an endoscopic camera manipulator. These techniques include identification of potential hazards and tracing their mitigating factors through the project lifecycle. PMID- 10719477 TI - Estimation of the stapes-bone thickness in the stapedotomy surgical procedure using a machine-learning technique. AB - Stapedotomy is a surgical procedure aimed at the treatment of hearing impairment due to otosclerosis. The treatment consists of drilling a hole through the stapes bone in the inner ear in order to insert a prosthesis. Safety precautions require knowledge of the nonmeasurable stapes thickness. The technical goal herein has been the design of high-level controls for an intelligent mechatronics drilling tool in order to enable the estimation of stapes thickness from measurable drilling data. The goal has been met by learning a map between drilling features, hence no model of the physical system has been necessary. Learning has been achieved as explained in this paper by a scheme, namely the d-sigma Fuzzy Lattice Neurocomputing (d sigma-FLN) scheme for classification, within the framework of fuzzy lattices. The successful application of the d sigma-FLN scheme is demonstrated in estimating the thickness of a stapes bone "on-line" using drilling data obtained experimentally in the laboratory. PMID- 10719478 TI - Interactive 3-D registration of ultrasound and magnetic resonance images based on a magnetic position sensor. AB - The use of stereotactic systems has been one of the main approaches for image based guidance of the surgical tool within the brain. The main limitation of stereotactic systems is that they are based on preoperative images that might become outdated and invalid during the course of surgery. Ultrasound (US) is considered the most practical and cost-effective intraoperative imaging modality, but US images inherently have a low signal-to-noise ratio. Integrating intraoperative US with stereotactic systems has recently been attempted. In this paper, we present a new system for interactively registering two-dimensional US and three-dimensional magnetic resonance (MR) images. This registration is based on tracking the US probe with a dc magnetic position sensor. We have performed an extensive analysis of the errors of our system by using a custom-built phantom. The registration error between the MR and the position sensor space was found to have a mean value of 1.78 mm and a standard deviation of 0.18 mm. The registration error between US and MR space was dependent on the distance of the target point from the US probe face. For a 3.5-MHz phased one-dimensional array transducer and a depth of 6 cm, the mean value of the registration error was 2.00 mm and the standard deviation was 0.75 mm. The registered MR images were reconstructed using either zeroth-order or first-order interpolation. The ease of use and the interactive nature of our system (approximately 6.5 frames/s for 344 x 310 images and first-order interpolation on a Pentium II 450 MHz) demonstrates its potential to be used in the operating room. PMID- 10719479 TI - Analysis of trabecular bone structure using Fourier transforms and neural networks. AB - Hip fracture due to osteoporosis (OP) and hip osteoarthritis (OA) are both important causes of locomotor morbidity in the elderly population. In osteoporosis, bone mass gradually decreases until the skeleton is too fragile to support the body and a fracture occurs, typically in the femur, wrist, or spine. In osteoarthritis, there is a proliferation of bone, leading to a stiffening of the tissue. Current clinical methods for assessment of bone changes in these disorders largely depend on assessing bone mineral density. However, this does not provide any information about bone structure which is considered to be an equally important factor in assessing bone quality. This paper presents a novel approach for computer analysis of trabecular (or cancellous) bone structure. The technique uses a Fourier transform to generate a "spectral fingerprint" of an image. Principal components analysis is then applied to identify key features from the Fourier transform and this information passed to a neural network for classification. Testing this on a series of 100 histological sections of trabecular bone from patients with OP and OA and a normal group correctly classified over 90% of the OP group with an overall accuracy of 77%-84%. Such high success rates on a small group suggest that this may provide a simple, but powerful, method for identifying alterations in bone structure. PMID- 10719480 TI - Telemedical information systems. AB - Telemedical information systems (TIS's) form the basis for telemedicine services as well as for health information services. This paper gives an introduction to the wide scope of TIS's and discusses examples for the different types of TIS's: patient-related TIS's, knowledge-related TIS's, and meta-TIS's. It concludes that for patient-related TIS's there is the need for a better integration of TIS's with other patient-centered information systems. For the other types of TIS's, more sophisticated retrieval techniques and better user interfaces for ordinary users are required. Furthermore, short descriptions of our own projects are given: the patient-related TIS of the project TECSAC (distributed electronic patient record for cardiology) and the domain-specific knowledge-related TIS's ODITEB (radiological Internet text-book) and ENDOTEL-EIS (endoscopy information system). PMID- 10719481 TI - Wavelet image compression--the quadtree coding approach. AB - Perfect reconstruction, quality scalability, and region-of-interest coding are basic features needed for the image compression schemes used in telemedicine applications. This paper proposes a new wavelet-based embedded compression technique that efficiently exploits the intraband dependencies and uses a quadtree-based approach to encode the significance maps. The algorithm produces a losslessly compressed embedded data stream, supports quality scalability, and permits region-of-interest coding. Moreover, experimental results obtained on various images show that the proposed algorithm provides competitive lossless/lossy compression results. The proposed technique is well suited for telemedicine applications that require fast interactive handling of large image sets, over networks with limited and/or variable bandwidth. PMID- 10719482 TI - Virtual surgery in a (tele-)radiology framework. AB - This paper presents telemedicine as an extension of a teleradiology framework through tools for virtual surgery. To classify the described methods and applications, the research field of virtual reality (VR) is broadly reviewed. Differences with respect to technical equipment, methodological requirements and areas of application are pointed out. Desktop VR, augmented reality, and virtual reality are differentiated and discussed in some typical contexts of diagnostic support, surgical planning, therapeutic procedures, simulation and training. Visualization techniques are compared as a prerequisite for virtual reality and assigned to distinct levels of immersion. The advantage of a hybrid visualization kernel is emphasized with respect to the desktop VR applications that are subsequently shown. Moreover, software design aspects are considered by outlining functional openness in the architecture of the host system. Here, a teleradiology workstation was extended by dedicated tools for surgical planning through a plug in mechanism. Examples of recent areas of application are introduced such as liver tumor resection planning, diagnostic support in heart surgery, and craniofacial surgery planning. In the future, surgical planning systems will become more important. They will benefit from improvements in image acquisition and communication, new image processing approaches, and techniques for data presentation. This will facilitate preoperative planning and intraoperative applications. PMID- 10719483 TI - A pacemaker working status telemonitoring algorithm. AB - To extend the application of a previously developed home ECG and blood pressure telemonitoring system for pacemaker users, a specially designed pacemaker working status analysis algorithm is developed in this paper. This paper includes four sections. First, a pacing beat and spontaneous beat recognition algorithm are established in which a FIR filter and three threshold methods are used to pick up the pacing spikes and a dynamic threshold modification method is proposed for error elimination. Second, a beat classification algorithm is developed based on decision rules for six commonly used pacemaker types. Third, a pacemaker malfunction recognition algorithm is established to evaluate the pacemaker working status. Fourth, the statistics of the analysis and the report are also provided. To validate the algorithm, clinical data were collected and manually classified by specialists. Then, the classification and recognition results were obtained with the clinical data. Results show that the algorithm can achieve a recognition rate of 98.4%. Results also indicate that the algorithm is capable of working in real-time speed. The algorithm has been incorporated into the telemonitoring system, and trial application cases are also reported. PMID- 10719484 TI - Design and implementation of a novel compression method in a tele-ultrasound system. AB - This paper represents a novel compression method for ultrasound images in a tele ultrasound system. The encoder chooses the ultrasound scan line signals as the object to be compressed and transmitted other than the standard video images in conventional methods. Furthermore, nonuniform discrete wavelet transform is proposed considering the different resolutions in axial and lateral directions of ultrasound images. Experimental results show that the new method provides better compression performance than conventional methods in terms of a peak signal-to noise ratio and processing time. PMID- 10719486 TI - Information technology applications in biomedical functional imaging. AB - In parallel with rapid advances in computer technology, biomedical functional imaging is having an ever-increasing impact on healthcare. Functional imaging allows us to see dynamic processes quantitatively in the living human body. However, as we need to deal with four-dimensional time-varying images, space requirements and computational complexity are extremely high. This makes information management, processing, and communication difficult. Using the minimum amount of data to represent the required information, developing fast algorithms to process the data, organizing the data in such a way as to facilitate information management, and extracting the maximum amount of useful information from the recorded data have become important research tasks in biomedical information technology. For the last ten years, the Biomedical and Multimedia Information Technology (BMIT) Group and, recently, the Center for Multimedia Signal Processing have conducted systematic studies on these topics. Some of the results relating to functional imaging data acquisition, compression, storage, management, processing, modeling, and simulation are briefly reported in this paper. PMID- 10719485 TI - Three-dimensional facial model reconstruction and plastic surgery simulation. AB - Facial model reconstruction and surgical simulation are essential to plastic surgery in today's medicine. Both can help surgeons to design appropriate repair plans and procedures prior to actual surgery. In this paper, we exploit a metamorphosis technique in our new design. First, using metamorphosis and vision techniques, we can establish three-dimensional facial models from a given photo. Second, we design several morphing operators, including augmentation, cutting, and lacerating. Experiments show that the proposed algorithms can successfully create acceptable facial models and generate realistically visual effects of surgical simulation. PMID- 10719487 TI - Context-based lossless and near-lossless compression of EEG signals. AB - In this paper, we study compression techniques for electroencephalograph (EEG) signals. A variety of lossless compression techniques, including compress, gzip, bzip, shorten, and several predictive coding methods, are investigated and compared. The methods range from simple dictionary-based approaches to more sophisticated context modeling techniques. It is seen that compression ratios obtained by lossless compression are limited even with sophisticated context based bias cancellation and activity-based conditional coding. Though lossy compression can yield significantly higher compression ratios while potentially preserving diagnostic accuracy, it is not usually employed due to legal concerns. Hence, we investigate a near-lossless compression technique that gives quantitative bounds on the errors introduced during compression. It is observed that such a technique gives significantly higher compression ratios (up to 3 bit/sample saving with less than 1% error). Compression results are reported for EEG's recorded under various clinical conditions. PMID- 10719488 TI - Neuroinformatics as a megascience issue. PMID- 10719489 TI - Evaluation of telemedical services. AB - With the rapidly increasing development of telemedicine technology, the evaluation of telemedical services becomes more and more important. However, professional views of the aims and methods of evaluation are different from the perspective of computer science and engineering or from medicine and health policy. We propose that a continuous evaluation strategy should be chosen which guides the development and implementation of telemedicine technologies and applications. The evaluation strategy is divided into four phases in which the focus of evaluation is shifted from technical performance of the system in the early phases to medical outcome criteria and economical aspects in later phases. We review the study design methodology established for clinical trials assessing therapeutic effectiveness and diagnostic accuracy and discuss how it can be adapted to evaluation studies in telemedicine. As an example, we describe our approach to evaluation in a teleconsultation network in ophthalmology. PMID- 10719490 TI - Telemedicine and terminology: different needs of context information. AB - Traditionally, communication between healthcare providers, including the provision of physicians with information and knowledge, works quite well because there are implicitly common conventions and assumptions in the collaboration of all involved actors. The basic prerequisite of the communication process between humans is a mutual agreement on syntax and semantics of oral and written language, i.e., of the medical sublanguage. With telemedicine, i.e., the use of telecommunication and informatics in medicine, this prerequisite is no longer sufficient. First, various professionals from different clinical communities, characterized by specialty, nationality, school, etc., share the management of the patient's health. Second, electronically communicated information is more and more intended for processing by computer applications rather than for direct interpretation by human users. Third, the interconnection of distributed heterogeneous software systems in medicine raises the issue of semantic interoperability, especially the problem of data integration. A faithful communication in such a scenario must be based on explicit assumptions "behind" a message. Interpreting an usually highly context-dependent utterance demands for mechanisms on a pragmatic level in natural language processing. The need for additional processing and integration of the transferred data by the receiving system demands for standardization and mediation, taking into consideration contextual knowledge that cannot entirely be explained and processed with linguistic and terminological approaches. This paper describes different categories of contexts and also different needs of applications concerning "context awareness." PMID- 10719491 TI - Telecommunication in healthcare for a better coordination between hospitals and GP's: routine application of the "ISAR-Telematics" project. AB - With the development of new information and communication technologies, the nature of data exchange between healthcare professionals is modified by the use of telematics tools. With the implementation of the "Reseau Sante Social" of the smart cards CPS and Sesam-Vitale, hospitals and private clinics are installing their hospital information systems, while general practitioners are computing their patient records. The environment is therefore suitable for the installation and diffusion of telematics services, allowing a better exchange of information between the physicians of a geographical sector. The RITHME intercommunication platform was developed in the ISAR-Telematics and IRISI European projects. It ensures several communication functions, such as movements of patients (in/out/inside) the hospital, management and mailing of letters and reports, specific information on prescribed treatments, or data concerning the hospital structure. This platform is tested in the town of Armentieres in the large city of Lille (North of France). It is currently being routinely used to accelerate and secure data exchanges between all the physicians working in this area. PMID- 10719492 TI - Tactical audio and acoustic rendering in biomedical applications. AB - Complexity of biomedical data requires novel sophisticated analysis and presentation methods. Sonification is used as a new information display in augmented reality systems to overcome problems of existing human-computer interface (e.g., opaque or heavy head-mounted displays, slow computer graphics, etc.). A novel taxonomy of sonification methods and techniques is introduced. We present our experience with tactical audio and acoustic rendering in biomedical applications. Tactical audio as an audio feedback is used as support for precise manual positioning of a surgical instrument in the operating room. Acoustic rendering is applied as an additional information channel and/or warning signal in biomedical signal analysis and data presentation. PMID- 10719493 TI - Alignment of adjacent picture frames captured by a CLSM. AB - Mosaicking a picture from its adjacent parts (called picture frames or tiles) is encountered in different fields of research and technology, e.g., photogrammetry, remote sensing, microscopy, etc. It is applied whenever the object of investigation is too large for the field of view of the sensor, e.g. a microscope. We describe mosaicking with a confocal laser-scanning microscope (CLSM) Bio-Rad, MRC 600 (U.K.). Aligning neighboring picture tiles was accomplished by registering the overlapped border areas of these tiles. Such registration procedures are constrained by: 1) the limited size of the registered samples (windows); 2) anisotropy of the form of the windows (usually narrow rectangles); and 3) the content of the windows, including changes of their intensity scale. Focusing on the latter problem, methods of registration were discussed and the robustness of the following three similarity based methods was studied with regard to the distortions of the intensity scales of the tiles to be registered: 1) the sum of absolute valued differences (SAVD); 2) normalized correlation coefficient (NCC); and 3) the mutual information function (MIF). Pilot experiments were extended to three-dimensional (3-D) stacks of pictures encountered in the framework of 3-D object rendering and visualization. MIF was found in most cases to be the most robust; however, it also demanded the most computational power. It is discussed how to choose a cost-effective method of the registration with regard to the content (texture, contrast, intensity scale distortion) of the tiles. PMID- 10719494 TI - Rapid automated tracing and feature extraction from retinal fundus images using direct exploratory algorithms. AB - Algorithms are presented for rapid, automatic, robust, adaptive, and accurate tracing of retinal vasculature and analysis of intersections and crossovers. This method improves upon prior work in several ways: 1) automatic adaptation from frame to frame without manual initialization/adjustment, with few tunable parameters; 2) robust operation on image sequences exhibiting natural variability, poor and varying imaging conditions, including over/under-exposure, low contrast, and artifacts such as glare; 3) does not require the vasculature to be connected, so it can handle partial views; and 4) operation is efficient enough for use on unspecialized hardware, and amenable to deadline-driven computing, being able to produce a rapidly and monotonically improving sequence of usable partial results. Increased computation can be traded for superior tracing performance. Its efficiency comes from direct processing on gray-level data without any preprocessing, and from processing only a minimally necessary fraction of pixels in an exploratory manner, avoiding low-level image-wide operations such as thresholding, edge detection, and morphological processing. These properties make the algorithm suited to real-time, on-line (live) processing and is being applied to computer-assisted laser retinal surgery. PMID- 10719495 TI - Interactive 3-D virtual colonoscopy system. AB - We describe a low-cost three-dimensional (3-D) virtual colonoscopy system that is a noninvasive technique for examining the entire colon and can assist physicians in detecting polyps inside the colon. Using the helical CT data and proposed techniques, we can three-dimensionally reconstruct and visualize the inner surface of the colon. We generate high resolution of video views of the colon interior structures as if the viewer's eyes were inside the colon. The physicians can virtually navigate inside the colon in two different modes: interactive and automatic navigation, respectively. For automatic navigation, the flythrough path is determined a priori using the 3-D thinning and two-pass tracking schemes. The whole colon is spatially subdivided into several cells, and only potentially visible cells are taken into account during rendering. To further improve rendering efficiency, potentially visible cells are rendered at different levels of detail. Additionally, a chain of bounding volume in each cell is used to avoid penetrating through the colon during navigation. In comparison with previous work, the proposed system can efficiently accomplish required preprocessing tasks and afford adequate rendering speeds on a low-cost PC system. PMID- 10719496 TI - AIM: attentionally based interaction model for the interpretation of vascular angiography. AB - We propose a model to interpret neurovascular X-ray angiogram (XRA) images interactively. This attentionally based interactive model (AIM) exploits human interaction as part of the solution. AIM posits two channels of interaction: context ("what to look for") and focus-of-attention ("where to look") as the locus of spatial information exchange between the user and the machine. In an AIM system, the user specifies a context (e.g., a carotid vessel) and directs the attentional spotlight to focus machine processing. AIM involves the user with the computer as integral partners and facilitates varying degrees of human intervention in the process. A hierarchy of context abstractions permits the system to function more autonomously (doing high-level tasks like extracting an arterial vessel) in routine interpretation and to require more user intervention (e.g., locating arterial wall boundaries) as the image complexity increases. This is especially important in medical imaging where the medical professional must have ultimate control and confidence in the system. Such technology can have a significant impact on the design of radiological systems. PMID- 10719497 TI - Telemedicine in the United Kingdom: current status and future prospects. AB - The objective of this correspondence is to present an overview of some of the current development and some of the ongoing telemedicine programs in the United Kingdom. The issues of the future integration of telemedicine activities within the National Health Service that promises better access to healthcare with higher efficiency, mobility, and lower cost are also discussed. PMID- 10719498 TI - Biomedicine in the 21st century: impact of information technology. PMID- 10719499 TI - Designing and implementing the transition to a fully digital hospital. AB - The increase in the number of examinations performed in modern healthcare institutions in conjunction with the range of imaging modalities available today have resulted in a tremendous increase in the number of medical images generated and has made the need for a dedicated system able to acquire, distribute, and store medical image data very attractive. Within the framework of the Hellenic R&D program, we have designed and implemented a picture archiving and communication system for a high-tech cardiosurgery hospital in Greece. The system is able to handle in a digital form images produced from ultrasound, X-ray angiography, gamma-camera, chest X-rays, as well as electrocardiogram signals. Based on the adoption of an open architecture highly relying on the DICOM standard, the system enables the smooth transition from the existing procedures to a fully digital operation mode and the integration of all existing medical equipment to the new central archiving system. PMID- 10719500 TI - Pain relief using smart technology: an overview of a new patient-controlled analgesia device. AB - A new adaptive patient-controlled analgesia (PCA) system designed to improve PCA through the use of a variable bolus dose and a variable background infusion is outlined here. The handset allows patients to rate their pain on a scale of 1-10. Data derived from the handset signals are used by an expert algorithm to repeatedly adapt the drug dosage of the bolus and the background infusion according to both pain intensity and patient response to previous dosages. A feasibility study of the system consisted of a small number of randomized, double blind, crossover clinical trials. The new system was alternated with a conventional system every 12 h. When the new system was active, 12-h questionnaire pain scores were significantly lower and a trend toward fewer bolus requests was found in alternating intervals. In addition, when the new system was used to commence trials, the number of bolus requests were significantly lower and there was a trend toward lower handset scores, lower questionnaire pain scores, and lower verbal pain scores over the entire trial period. The new adaptive PCA system was well accepted by both patients and clinical staff. The trials successfully established the feasibility of the new system. Further development is being carried out as a result. PMID- 10719501 TI - Computer-supported collaborative work (CSCW) in biomedical signal visualization and processing. AB - A collaborative extension to a platform dedicated to signal visualization and processing is presented. The platform now allows geographically distant users to work together, in real-time, on the same data (electroencephalographic signals). The extension is based on a client-server collaborative architecture using remote procedure call programming. Through this first implementation, general issues of computer-supported collaborative work are presented. PMID- 10719502 TI - Coherence of multiscale features for enhancement of digital mammograms. AB - Mammograms depict most of the significant changes in breast disease. The primary radiographic signs of cancer are related to tumor mass, density, size, borders, and shape, and local distribution of calcifications. We show that each of these features can be well described by coherence and orientation measures and provide visual cues for radiologists to identify possible lesions more easily without increasing false positives. In this paper, an artifact-free enhancement algorithm based on overcomplete multiscale representations is presented. First, an image was decomposed using a fast wavelet transform algorithm. At each level of analysis, energy and phase information are computed via a set of separable steerable filters. Then, a measure of coherence within each level was obtained by weighting an energy measure with the ratio of projections of local energy within a specified window. Each projection was computed onto the central point of a window with respect to the total energy within that window. Finally, a nonlinear operation, integrating coherence and orientation information, was applied to modify transform coefficients within distinct levels of analysis. These modified coefficients were then reconstructed, via an inverse fast wavelet transform, resulting in an improved visualization of significant mammographic features. The novelty of this algorithm lies in the detection of directional multiscale features and the removal of aliased perturbations. Compared to existing multiscale enhancement techniques, images processed with this method appeared more familiar to radiologists due to localized enhancement of features. PMID- 10719503 TI - Automatic retinal image registration scheme using global optimization techniques. AB - Retinal image registration is commonly required in order to combine the complementary information in different retinal modalities. In this paper, a new automatic scheme to register retinal images is presented and is currently tested in a clinical environment. The scheme considers the suitability and efficiency of different image transformation models and function optimization techniques, following an initial preprocessing stage. Three different transformation models- affine, bilinear and projective--as well as three optimization techniques- downhill simplex method, simulated annealing and genetic algorithms--are investigated and compared in terms of accuracy and efficiency. The registration of 26 pairs of Fluoroscein Angiography and Indocyanine Green Chorioangiography images with the corresponding Red-Free retinal images, showed the superiority of combining genetic algorithms with the affine and bilinear transformation models. A comparative study of the proposed automatic registration scheme against the manual method, commonly used in the clinical practice, is finally presented showing the advantage of the proposed automatic scheme in terms of accuracy and consistency. PMID- 10719504 TI - DNA ploidy and cell cycle distribution of breast cancer aspirate cells measured by image cytometry and analyzed by artificial neural networks for their prognostic significance. AB - Chromosomal abnormalities are commonly associated with cancer, and their importance in the pathogenesis of the disease has been well recognized. Also recognized in recent years is the possibility that, together with chromosomal abnormalities, DNA ploidy of breast cancer aspirate cells, measured by image cytometric techniques, may correlate with prognosis of the disease. Here, we have examined the use of an artificial neural network to predict: 1) subclinical metastatic disease in the regional lymph nodes and 2) histological assessment, through the analysis of data obtained by image cytometric techniques of fine needle aspirates of breast tumors. The cellular features considered were: 1) DNA ploidy measured in terms of nuclear DNA content as well as by cell cycle distribution; 2) size of the S-phase fraction; and 3) nuclear pleomorphism. A further objective of the study was to analyze individual markers in terms of impact significance on predicting outcome in both cases. DNA ploidy, indicated by cell cycle distribution, was found markedly to influence the prediction of nodal spread of breast cancer, and nuclear pleomorphism to a lesser degree. Furthermore, a comparison between histological assessment and artificial neural network prediction shows a closer correlation between the neural approach and the development of further metastases as indicated in subsequent follow-up, than does histological assessment. These data demonstrate that artificial neural networks are capable of providing powerful and reliable indicators of possible lymph node metastasis, using measurements of cellular features alone. PMID- 10719505 TI - The display of photographic-quality images on the Web: a comparison of two technologies. AB - Downloading medical images on the Web creates certain compromises. The tradeoff is between higher resolution and faster download times. As resolution increases, download times increase. High-resolution (photographic quality) electronic images can potentially play a key role in medical education and patient care. On the Internet, images are typically formatted as Graphics Interchange Format (GIF) or the Joint Photographic Experts Group (JPEG) files. However, these formats are associated with considerable data loss in both color depth and image resolution. Furthermore, these images are available in a single resolution and have no capability of allowing the user to adjust resolution as needed. Images in the photo compact disc (PCD) format have higher resolutions than GIF or JPEG, but suffer the disadvantage of large file sizes leading to long download times on the Web. Furthermore, native web browsers are not currently able to read PCD files. The FlashPix format (FPX) offers distinct advantages over the PCD, GIF, and JPEG formats for display of high-resolution images on the Web. A Java applet can be easily downloaded for viewing FPX images. FPX images are higher resolution than JPEG and GIF images. FPX images offer rich resolutions comparable to PCD images with shorter download times. PMID- 10719506 TI - A systems approach to achieving CarerNet--an integrated and intelligent telecare system. AB - The system requirements for an integrated telecare system are presented by using a modification of the CORE methodology. The resulting system model uses viewpoint analysis to establish the needs and requirements of both the client and the care providers within a system context and discusses the implications for a user centered, technology-based solution referred to as "CarerNet." The implementation of CarerNet is considered in terms of the enabling technologies required within the local environment of the client and identifies the monitoring, networking requirements, and system intelligence necessary to the provision of a comprehensive telecare service. A hypothetical case study based on patient aftercare following discharge from the hospital is presented to illustrate the points discussed. PMID- 10719507 TI - Extracting isovolumes from three-dimensional torso geometry using PROLOG. AB - Three-dimensional (3-D) finite element torso models are widely used to simulate defibrillation field quantities, such as potential, gradient, and current density. These quantities are computed at spatial nodes that comprise the torso model. These spatial nodes typically number between 10(5) and 10(6), which makes the comprehension of torso defibrillation simulation output difficult. Therefore, the objective of this study is to rapidly prototype software to extract a subset of the geometric model of the torso for visualization in which the nodal information associated with the geometry of the model meets a specified threshold value (e.g., minimum gradient). The data extraction software is implemented in PROLOG, which is used to correlate the coordinate, structural, and nodal data of the torso model. A PROLOG-based environment has been developed and is used to rapidly design and test new methods for sorting, collecting, and optimizing data extractions from defibrillation simulations in a human torso model for subsequent visualization. PMID- 10719508 TI - A three-generation model for teleradiology. AB - This paper proceeds from the definition of teleradiology. It identifies three different generations of teleradiology systems and includes those systems that are not regarded as teleradiology systems by the authors. A list of requirements pertinent to users of first-generation teleradiology systems is introduced. Most of the requirements have been realized in a new generation teleradiology system called CHILI. PMID- 10719509 TI - Experience with PACS in an ATM/Ethernet switched network environment. AB - Legacy local area network (LAN) technologies based on shared media concepts are not adequate for the growth of a large-scale picture archiving and communication system (PACS) in a client-server architecture. First, an asymmetric network load, due to the requests of a large number of PACS clients for only a few main servers, should be compensated by communication links to the servers with a higher bandwidth compared to the clients. Secondly, as the number of PACS nodes increases, the network throughout should not measurably cut production. These requirements can easily be fulfilled using switching technologies. Here asynchronous transfer mode (ATM) is clearly one of the hottest topics in networking because the ATM architecture provides integrated support for a variety of communication services, and it supports virtual networking. On the other hand, most of the imaging modalities are not yet ready for integration into a native ATM network. For a lot of nodes already joining an Ethernet, a cost-effective and pragmatic way to benefit from the switching concept would be a combined ATM/Ethernet switching environment. This incorporates an incremental migration strategy with the immediate benefits of high-speed, high-capacity ATM (for servers and high-sophisticated display workstations), while preserving elements of the existing network technologies. In addition, Ethernet switching instead of shared media Ethernet improves the performance considerably. The LAN emulation (LANE) specification by the ATM forum defines mechanisms that allow ATM networks to coexist with legacy systems using any data networking protocol. This paper points out the suitability of this network architecture in accordance with an appropriate system design. PMID- 10719510 TI - Deployable teleradiology: Bosnia and beyond. AB - The United States military has been an effective proponent of digital imaging and teleradiology for the past 15 years [1]. A digital imaging network that eliminates the use of x-ray film makes military medicine requirements simpler. X ray film requirements include storage of new, unexposed films, storage and use of chemicals and water for processing, and disposal of chemicals. In some deployed situations, the chemical discharge needs to be collected and shipped out of the area. Therefore, the ability to implement electronic imaging and eliminate or greatly reduce the dependence on film, chemicals, and water are intrinsically important to military medicine. In December 1995, the United States government began deployment of 20,000 United States troops to Bosnia-Herzegovina as part of NATO's peacekeeping implementation force (IFOR) operation. A full complement of military medical support facilities was established in Bosnia. An army base in Hungary was the location from which the deployment was staged. The project to deploy telemedicine and teleradiology capabilities to the medical treatment facilities (MTF) in Bosnia and Hungary became known as PrimeTime III [2]. This paper deals with the deployable teleradiology (DEPRAD) system that was installed by the Imaging Science and Information Systems (ISIS) Center, Department of Radiology, Georgetown Medical Center, Washington, DC, at a number of facilities to implement filmless radiology and teleradiology services in support of PrimeTime III. PMID- 10719511 TI - Interactive DICOM image transmission and telediagnosis over the European ATM network. AB - The European High-Performance Information Infrastructure in Medicine, n(o)B3014 (HIM3) project of the Trans-European Network--Integrated Broadband Communications (TEN-IBC) program, started on March 1996 and finished on February 1997, aimed to test the medical usability of the European asynchronous transfer mode (ATM) network in medical image transmission. The Department of Radiology, University of Pisa, Pisa, Italy, and St-Luc University Hospital, Brussels, Belgium, involved in the project as healthcare partners in the radiological domain, established several connection sessions finalized to test the usability of Digital Imaging and Communication (DICOM) image transmission and interactive telediagnosis tools in the daily radiological practice. The Pisa site was connected to the Italian ATM pilot (Sirius Network) through the Tuscany metropolitan area network (MAN), while St-Luc University Hospital was connected to Belgium ATM network through the Brussels MAN. By means of international connections provided by the European JAMES project, a link between the two sites was established, connecting both national ATM networks. Due to the large variety of hardware present in the medical centers, multiplatform software tools were used and tested: central test node (CTN) release 2.8 [3], VAT [6], NV-3.3 [7], and IDI (UCL homemade multiplatform teleradiology tool for interactive visualization and processing of DICOM images). During the telediagnosis session, lead by radiologists in both hospitals, each site submitted neuroradiological clinical cases to the other for remote consultation. The connection, available for a period of two weeks, at 2 Mbit/s bandwidth, allowed the transmission of MR images (256 x 256 x 12 bit) and simultaneous multimedia interactive discussion of the cases. Both off-line transmission and review of the images, using the CTN DICOM transfer routines, and on-line interactive image discussion, using the IDI telediagnosis software, were tested successfully from the technical and medical point of view. PMID- 10719512 TI - PACS/information systems interoperability using Enterprise Communication Framework. AB - Interoperability among healthcare applications goes beyond connectivity to allow components to exchange structured information and work together in a predictable, coordinated fashion. To facilitate building an interoperability infrastructure, an Enterprise Communication Framework (ECF) was developed by the members of the Andover Working Group for Healthcare Interoperability (AWG-OHI). The ECF consists of four models: 1) Use Case Model, 2) Domain Information Model (DIM), 3) Interaction Model, and 4) Message Model. To realize this framework, a software component called the Enterprise Communicator (EC) is used. In this paper, we will demonstrate the use of the framework in interoperating a picture archiving and communication system (PACS) with a radiology information system (RIS). PMID- 10719513 TI - Time and space results of dynamic texture feature extraction in MR and CT image analysis. AB - Texture feature extraction is a fundamental part of texture image analysis. Therefore, the reduction of its computational time and storage requirements should be an aim of continuous research. The Spatial Grey Level Dependence Method (SGLDM) is one of the most important statistical texture description methods, especially in medical image analysis. Co-occurrence matrices are employed for the implementation of this method; however, they are inefficient in terms of computational time and memory space, due to their dependency on the number of gray levels (gray-level range) in the entire image. Since texture is usually measured in a small image region, a large amount of memory is wasted while the computational time of the texture feature extraction operations is unnecessarily raised. Their inefficiency puts up barriers to the wider utilization of SGLDM in a real application environment, such as a clinical environment. In this paper, the memory space and time efficiency of a dynamic approach to texture feature extraction in SGLDM is investigated through a pilot application in the analysis of magnetic resonance (MR) and computed tomography (CT) images. PMID- 10719514 TI - A VRML-based anatomical visualization tool for medical education. AB - The advent of the Virtual Reality Modeling Language (VRML) as a portable file format for describing three-dimensional (3-D) scenes has enabled researchers, educators, and students to share anatomical models on the World Wide Web (WWW). The implication for medical teaching is that students can interactively examine anatomical structures and their 3-D spatial relationships by using current personal computer (PC) technology. This paper describes the creation of 3-D anatomical models that are accessible on the WWW, using high-resolution middle ear data as an example. The 3-D models are created by interactive segmentation of the source images (histological and MRI sections) and 3-D surface reconstruction. The resulting models are translated into VRML format. Section images can be superimposed on the model, allowing students to view a section in its 3-D context. To enhance the viewing of these scenes, a VRML browser was modified to support transparent rendering of surfaces. Finally, a WWW interface was designed to allow users to choose the model structures, section images, and associated viewing parameters to build their own 3-D scenes. PMID- 10719515 TI - Teleworks: a CSCW application for remote medical diagnosis support and teleconsultation. AB - The present paper describes methods for the design of both synchronous and asynchronous computer-supported cooperative work (CSCW) procedures suitable for the medical application area and specifically for the purpose of medical teleconsultation and remote diagnosis support. The experimental implementation of a CSCW system built upon a PC/Windows platform is detailed as an example of a low cost system suitable for adoption in a wide range of medical teleconsultation applications. PMID- 10719516 TI - Design and development of an interactive medical teleconsultation system over the World Wide Web. AB - The objective of the medical teleconsultation system presented in this paper is to demonstrate the use of the World Wide Web (WWW) for telemedicine and interactive medical information exchange. The system, which is developed based on Java, could provide several basic Java tools to fulfill the requirements of medical applications, including a file manager, data tool, bulletin board, and digital audio tool. The digital audio tool uses point-to-point structure to enable two physicians to communicate directly through voice. The others use multipoint structure. The file manager manages the medical images stored in the WWW information server, which come from a hospital database. The data tool supports cooperative operations on the medical data between the participating physicians. The bulletin board enables the users to discuss special cases by writing text on the board, send their personal or group diagnostic reports on the cases, and reorganize the reports and store them in its report file for later use. The system provides a hardware-independent platform for physicians to interact with one another as well as to access medical information over the WWW. PMID- 10719517 TI - Clinical decision support, systems methodology, and telemedicine: their role in the management of chronic disease. AB - In this paper, the design and evaluation of decision support systems, including those incorporating a telematic component, are considered. It is argued that effective design and evaluation are dependent upon the adoption of appropriate methodology set firmly within a systemic framework. Systems modeling is proposed as an approach to system design, with evaluation adopting an approach incorporating evaluability analysis and formative and summative evaluation, including the use of stakeholder matrix analysis. The relevance of such systemic methodology is demonstrated in the context of diabetes and end-stage renal disease as examples of the generic clinical problem of the management of chronic disease. PMID- 10719518 TI - On the integration of healthcare emergency systems in Europe: the WETS project case study. AB - The European Union (EU) is characterized by a large number of different emergency healthcare (EHC) systems. In this situation, a common policy for healthcare emergency handling is largely prevented and is a cause of an increase of the costs associated with such systems all over Europe. There is, hence, a need for a homogenization and integration of healthcare emergency systems in Europe. This turns out to be difficult because of the ethical, political, legal, and technological differences and peculiarities of the European scenario and the large investments that would be needed in this sector. The process of integration passes through the identification of the main functionalities--driven by the user needs in their real life conditions--along with the technologies that are best fitted for supporting them. In this paper, several aspects related to these problems are analyzed and a real case study, drawn from the Project "Worldwide Emergency Telemedicine Services" (WETS), supported by the European Commission (DG XIII), is presented. Within WETS, several pilot sites (in Italy, Spain, Greece, Denmark, and Iceland) consider different aspects of the integration of healthcare emergency systems with particular focus on the sharing of solutions that "traditionally" belong to different environments (i.e., land, air, and sea). The involvement of important hospitals, ship companies, airlines, and emergency health institutions allows us to devote a large part of this two-year project (1998-1999) to validate and demonstrate the results of the development phase in real-life conditions. Some more concrete details are given for the Italian pilot site, where the authors operate. PMID- 10719519 TI - The implementation of a quality-net as a part of the European project DIABCARE Q Net. AB - DIABCARE Q-Net is a European project with a consortium of partners in healthcare, industry, and research, which has the overall target of improvement in diabetes care by aggregation, evaluation, and feedback of anonymized patient data with the tools of modern telematics, resulting from the initiative of the St. Vincent Declaration, St. Vincent, Italy. Based on standardized tools for quality improvement in diabetes care, i.e., the Basic Information Sheet (BIS) and recently developed data entry and feedback software (DIABCARE Data for Windows), DIABCARE Q-Net as a part of the Telematics Applications Program of the European Commission will improve diabetes care and disease management by the implementation of a quality network. Therefore, the project implements regional, national, and central nodes for processing of diabetes quality indicators. All participating centers (GP's and clinics in Europe) get feedback by standardized benchmarking. The pilot testing and the state of implementation of our network confirm the importance of improving the quality of life of diabetic patients in all participating countries. PMID- 10719520 TI - Special issue on emerging health telematics applications in Europe. PMID- 10719521 TI - Combining a scientific approach and prototyping in the design of EHCR systems. AB - In this paper, it is emphasized that electronic medical record systems cannot totally be developed in the traditional way. The underlying process of how physicians or nurses are searching for information is not fully understood. Therefore, a method that combines a scientific approach and prototyping is advocated. With the help of this advocated approach, these questions could be answered in a way that was also scientifically sound. In this contribution, two examples of the use of this method are presented. One concerns the determination of the optimum granularity of the narrative parts of the electronic healthcare record (EHCR) and the other concerns the use and impact of stand-alone protocol systems. PMID- 10719522 TI - A CORBA-based integration of distributed electronic healthcare records using the synapses approach. AB - The ability to exchange in a meaningful, secure, and simple fashion relevant healthcare data about patients is seen as vital in the context of efficient and cost-effective shared or team-based care. The electronic healthcare record (EHCR) lies at the heart of this information exchange, and it follows that there is an urgent need to address the ability to share EHCR's or parts of records between carers and across distributed health information systems. This paper presents the Synapses approach to sharing based on a standardized shared record, the Federated Healthcare Record, which is implemented in an open and flexible manner using the Common Object Request Broker Architecture (CORBA). The architecture of the Federated Healthcare Record is based on the architecture proposed by the Technical Committee 251 of the European Committee for Standardization. PMID- 10719523 TI - A workflow-based approach to virtual patient record security. AB - Virtual patient records provide a means for integrated access to patient information that may be scattered around different healthcare organizations (or hospital departments). As Intranets provide, among others, secure access to medical information, they constitute an appropriate technological infrastructure for a virtual patient record implementation. In such cases, a security policy can be enforced by combining the security features of the Intranet with the security features of the intraorganizational systems. However, when a workflow system is implemented to automate interorganizational healthcare processes, an additional security layer is needed. An authorization architecture that serves this purpose is presented in this paper. PMID- 10719524 TI - A broadband multimedia collaborative system for advanced teleradiology and medical imaging diagnosis. AB - This paper presents a new telemedicine system currently in routine clinical usage, developed within the European Union (EU) ACTS BONAPARTE project (1). The telemedicine system is developed on an asynchronous transfer mode (ATM) multimedia hardware/software platform comprising the following set of telemedicine services: synchronous cooperative work, high-quality video conference, multimedia mail, medical image digitizing, processing, storing and printing, and local and remote transparent database access. The medical information handled by the platform conforms to the Digital Imaging and Communications in Medicine (DICOM) 3.0 medical imaging standard. The telemedicine system has been installed for clinical routines in three Spanish hospitals since November 1997 and has been used in an average of one/two clinical sessions per week. At each clinical session, a usability and clinical evaluation of the system was carried out. Evaluation is carried out through direct observation of interactions and questionnaire-based subjective data. The usability evaluation methodology and the results of the system usability study are also presented in this article. The experience gained from the design, development, and evaluation of the telemedicine system is providing an indepth knowledge of the benefits and difficulties involved in the installation and clinical usage of this type of high usability and advanced multimedia telemedicine system in the field of teleradiology and collaborative medical imaging diagnosis. PMID- 10719526 TI - Augmented-reality communication for diagnostic tasks in cardiology. AB - In the CardiAssist project, a teleconsultation module has been implemented that uses both ultrasound images and three-dimensional (3-D) animated heart models as common artifacts to support communication between medical professionals. Real images and conceptual models are linked as "augmented reality" so that the models provide orientation for diagnostic tasks in cardiology. Hypotheses or results can be documented on the image or the abstract level, as the heart model can be modified to include pathological details. In teleconsultation, pointing references accelerate communication between physicians of different backgrounds. PMID- 10719525 TI - The EASI project--improving the effectiveness and quality of image-guided surgery. AB - In recent years, advances in computer technology and a significant increase in the accuracy of medical imaging have made it possible to develop systems that can assist the clinician in diagnosis, planning, and treatment. This paper deals with an area that is generally referred to as computer-assisted surgery, image directed surgery, or image-guided surgery. We report the research, development, and clinical validation performed since January 1996 in the European Applications in Surgical Interventions (EASI) project, which is funded by the European Commission in their "4th Framework Telematics Applications for Health" program. The goal of this project is the improvement of the effectiveness and quality of image-guided neurosurgery of the brain and image-guided vascular surgery of abdominal aortic aneurysms, while at the same time reducing patient risks and overall cost. We have developed advanced prototype systems for preoperative surgical planning and intraoperative surgical navigation, and we have extensively clinically validated these systems. The prototype systems and the clinical validation results are described in this paper. PMID- 10719527 TI - The construction of a simulation-based system for the development of powerful and realistic models and practicals for healthcare professionals. AB - The aim of this paper is to examine the importance of computer-based simulation by the demonstration and study of complex systems and the presentation of essential tools and applications that can help health professionals deliver good quality practicals, which is now impeded by cost and/or technical constrains. The tools that have been developed in the framework of the Courseware Authoring for Scientific Training (COAST) project are the "modeler environment," which is used to describe the different tools and mathematical functions available for building models, and the "simulation author environment," which is used for building simulation sequences and providing the required tools and functions. This effort provides scientists with new technological and cost-effective means, specifically based on multimedia simulation, for preparing educational material, so as to gradually replace laboratory practicals that are gradually becoming more expensive, and improves student's understanding of complex systems. PMID- 10719528 TI - A unified multimedia database system to support telemedicine. AB - A unified approach to managing multimedia medical databases in a telemedicine system is proposed. In order to manage, search, and display patient information more efficiently, we define a patient information package (PIP) as a concise data set of a patient's medical information from each visit. By means of PIP's, both patient-oriented and problem-oriented query strategies, which are most frequently used in daily clinical practice and medical education, can be accommodated. We also provide a unified methodology for accessing various types of patient medical records as well as design two types of user interfaces, high-quality data display and web-based interface, for different medical service purposes. The PIP-based management of databases has been successfully implemented between the National Taiwan University (NTUH), Taipei, and the Chinshan health care center, Chinshan, Taiwan, for teleconsultation, telediagnosis, and tele-education. PMID- 10719529 TI - Black Sea TeleDiab: diabetes computer system with communication technology for Black Sea region. AB - BlackSea TeleDiab (BSTD) is a multidisciplinary research project whose aim is to promote the exchange of healthcare information between clinicians and scientists in countries of the Black Sea area to provide a means by which the care of patients with diabetes may be enhanced. The project is built on an existing organizational framework provided by the WHO Diabcare Quality Network (Q-Net) and the Black Sea Diab Action Project. The aim is to develop a standardized software package (in the national languages of the CCE/NIS Black Sea partners) for the storage and transfer of medical information and healthcare data between participating institutions. It will utilize a standard format for medical records based on the Good European Health Record (GEHR), a project within the Advanced Informatics in Medicine (AIM) program, which aims to develop and propagate a common architecture for computerized health records across Europe that can be used across clinical domains, countries, and computer systems. PMID- 10719530 TI - Microscopic image analysis for quantitative measurement and feature identification of normal and cancerous colonic mucosa. AB - The development of an automated algorithm for the categorization of normal and cancerous colon mucosa is reported. Six features based on texture analysis were studied. They were derived using the co-occurrence matrix and were angular second moment, entropy, contrast, inverse difference moment, dissimilarity, and correlation. Optical density was also studied. Forty-four normal images and 58 cancerous images from sections of the colon were analyzed. These two groups were split equally into two subgroups: one set was used for supervised training and the other to test the classification algorithm. A stepwise selection procedure showed that correlation and entropy were the features that discriminated most strongly between normal and cancerous tissue (P < 0.0001). A parametric linear discriminate function was used to determine the classification rule. For the training set, a sensitivity and specificity of 93.1% and 81.8%, respectively, were achieved, with an overall accuracy of 88.2%. These results were confirmed with the test set, with a sensitivity and specificity of 93.1% and 86.4%, respectively, and an overall accuracy of 90.2%. PMID- 10719531 TI - Real-time separation of discontinuous adventitious sounds from vesicular sounds using a fuzzy rule-based filter. AB - The separation of pathological discontinuous adventitious sounds (DAS) from vesicular sounds (VS) is of great importance to the analysis of lung sounds since DAS are related to certain pulmonary pathologies. An automated way of revealing the diagnostic character of DAS, by isolating them from VS, based on their nonstationarity, is presented in this paper. The proposed algorithm uses two adaptive network-based fuzzy inference systems to compose a generalized fuzzy rule-based stationary-nonstationary filter (GFST-NST). The training procedure of the fuzzy inference systems involves the outputs of the wavelet transform-based stationary-nonstationary filter (WTST-NST), proposed by Hadjileontiadis and Panas [1]. The basic idea of the GFST-NST was initially proposed by the authors with the introduction of the fuzzy rule-based stationary-nonstationary filter (FST NST) [2], tested with the separation of crackles from VS. The main contribution of this paper is the modification of the structure of the FST-NST filter to a serial-type fuzzy filter that, unlike the parallel operation of the FST-NST filter, sends a predicted stationary signal (VS) into the predictor of the nonstationary (DAS). Applying the GFST-NST filter to fine-coarse crackles and squawks, selected from three lung sound databases, the coherent structure of DAS is revealed and they are separated from VS. The separation performance of the GFST-NST filter was evaluated through quantitative and qualitative indexes that proved its efficiency and superiority against the FST-NST filter. When compared to the WTST-NST filter, the GFST-NST filter performed similarly in accuracy and objectiveness, but in a faster way. Thus, the GFST-NST filter combines the separation accuracy of the WTST-NST filter with the real-time implementation of the FST-NST filter, so it can easily be used in clinical medicine as a module of an integrated intelligent patient evaluation system. PMID- 10719532 TI - Decision support and disease management: a logic engineering approach. AB - This paper describes the development and application of PROforma, a unified technology for clinical decision support and disease management. Work leading to the implementation of PROforma has been carried out in a series of projects funded by European agencies over the past 13 years. The work has been based on logic engineering, a distinct design and development methodology that combines concepts from knowledge engineering, logic programming, and software engineering. Several of the projects have used the approach to demonstrate a wide range of applications in primary and specialist care and clinical research. Concurrent academic research projects have provided a sound theoretical basis for the safety critical elements of the methodology. The principal technical results of the work are the PROforma logic language for defining clinical processes and an associated suite of software tools for delivering applications, such as decision support and disease management procedures. The language supports four standard objects (decisions, plans, actions, and enquiries), each of which has an intuitive meaning with well-understood logical semantics. The development toolset includes a powerful visual programming environment for composing applications from these standard components, for verifying consistency and completeness of the resulting specification and for delivering stand-alone or embeddable applications. Tools and applications that have resulted from the work are described and illustrated, with examples from specialist cancer care and primary care. The results of a number of evaluation activities are included to illustrate the utility of the technology. PMID- 10719533 TI - Reconciling users' needs and formal requirements: issues in developing a reusable ontology for medicine. AB - A common language, or terminology, for representing what clinicians have said and done is an important requirement for individual clinical systems, and it is a pre requisite for integrating disparate applications in a distributed telematic healthcare environment. Formal representations based on description logics or closely related formalisms are increasingly used for representing medical terminologies. GALEN's experience in using one such formalism raises two major issues, as follows: how to make ontologies based on description logics easy to use and understand for both clinicians and applications developers; what features are required of the ontology and description logic if they are to achieve their aims. Based on our experience we put forward four contentions: two relating to each of these two issues, as follows: that natural language generation is essential to make a description logic based ontology accessible to users; that the description logic based ontology should be treated as an "assembly language" and accessed via "intermediate representations" oriented to users and "perspectives" adapting it to specific applications; that independence and reuse are best supported by partitioning the subsumption hierarchy of elementary concepts into orthogonal taxonomies, each of which forms a pure tree in which the branches at each level are disjoint but nonexhaustive subconcepts of the parent concept; that the expressivity of the description logic must include support for transitive relations despite the computational cost, and that this computational cost is acceptable in practice. The authors argue that these features will be necessary, though by no means sufficient, for the development of any large reusable ontology for medicine. PMID- 10719534 TI - Integration of clinical information across patient records: a comparison of mechanisms used to enforce semantic coherence. AB - Semantic coherence about clinical information is the bottleneck for true interoperability among applications in health telematics. Healthcare records are in principle made of statements about patient's health and activities performed, organized within attested transactions or messages. Various mechanisms were developed to optimally represent details of statements into a record system, creating de facto three subdivisions: 1) "containers" of clinical information, i.e., section headings, data elements in local records; segments and data fields in messages; 2) their "contents," i.e., coding systems and terminologies; and 3) "transaction context," i.e., circumstances related to document production and message exchange, typically represented in their headers. Details rely on a common semantic background and should therefore, be seen in a continuum; nevertheless, design methodologies and tools for the three subdivisions evolved independently and assignment of details to subdivisions is not predetermined by principles, but depends on implementation issues. Recent developments within the European Committee for Standardization (CEN/TC251/WG II) and in the European Project GALEN-IN-USE provide a new insight on semantics in healthcare. In order to guide harmonization of semantic aspects in the different series of standards- in information models, messages, document markup, terminology systems--we present here a comparison of the various mechanisms they use to enforce semantic coherence on clinical information. PMID- 10719535 TI - Relief for maritime medical emergencies through telematics. AB - MERMAID is a European Union (EU)-financed maritime telemedicine project with global reach and 24-h multilingual capability, so as to serve multinational crews working in the isolation of the world's oceans. It provides a model for the provision of healthcare services based on the electronic transmission of medical information via ISDN-based video conferencing. This model is not limited to medical diagnostics, but it encompasses all cases in which the actual delivery of healthcare services involves a patient who is not located where the provider is. Its implementation requires the commissioning of an extensive telecommunications infrastructure that includes both satellite transmission for ship to shore communication and an extensive ground-based network for summoning expert medical help from around the world so as to meet the project's multilinguality requirements and, therefore, the exploration of a number of solutions. In fact, all categories of telemedical applications (audio and video conferencing, multimedia communications, flat file and image transfer with low-, medium-, and high-bandwidth data requirements) are considered, while the full range of network choices (digital land lines, cellular/wireless, satellite, and broadband) are being tested in terms of cost/performance tradeoffs that are inherent to them and the developmental stage each of these options occupies in their lifecycle. Finally, out of that, MERMAID utilizes advanced land-based line transmission technologies to aid the remote patient by making available the specialist care that is best suited in the particular case. PMID- 10719536 TI - A novel emergency telemedicine system based on wireless communication technology- AMBULANCE. AB - Recent studies conclude that early and specialized prehospital management contributes to emergency case survival. We have developed a portable medical device that allows telediagnosis, long distance support, and teleconsultation of mobile healthcare providers by expert physicians. The device allows the transmission of vital biosignals and still images of the patient from the emergency site to the consultation site using the GSM mobile telephony network. The device can telematically "bring" an expert specialist doctor at the site of the medical emergency, allow him/her to evaluate patient data, and issue directions to the emergency personnel on treatment procedures until the patient is brought to be hospital. Legal reasons mandated the inclusion at the consultation site of a multimedia database able to store and manage the data collected by the system. The performance of the system has been validated in four different countries using a controlled medical protocol and a set of 100 patients per country treated has been collected and analyzed. PMID- 10719537 TI - ViVa: the virtual vascular project. AB - The aim of the virtual vascular project (ViVa) is to develop tools for the modern hemodynamicist and cardiovascular surgeon to study and interpret the constantly increasing amount of information being produced by noninvasive imaging equipment. In particular, we are developing a system able to process and visualize three dimensional (3-D) medical data, reconstruct the geometry of arteries of specific patients, and simulate blood flow in them. The initial applications of the system will be for clinical research and training purposes. In a later stage, we will explore the application of the system to surgical planning. ViVa is based on an integrated set of tools, each dedicated to a specific aspect of the data processing and simulation pipeline: image processing and segmentation; real-time 3-D volume visualization; 3-D geometry reconstruction; 3-D mesh generation; and blood flow simulation and visualization. PMID- 10719538 TI - Virtual environments in neuroscience. AB - Virtual environments (VE's) let users navigate and interact with computer generated three-dimensional (3-D) environments in real time, allowing for the control of complex stimuli presentation. These VE's have attracted much attention in medicine, especially in remote or augmented surgery, and surgical training, which are critically dependent on hand-eye coordination. Recently, however, some research projects have begun to test the possibility of using VE's for the study and rehabilitation of human cognitive and functional activities. This paper highlights recent and ongoing research related to the applications of VE's in the neuroscience arena. In particular, it focuses on the American and European initiatives in this field, including a description of the European Commission (EC)-funded VREPAR projects. Finally, the paper provides a general introduction to virtual reality (VR), as it relates to its impact on cognitive and functional abilities. PMID- 10719539 TI - A system for medical consultation and education using multimodal human/machine communication. AB - Recent developments in networking and computing have enabled collaborative biomedical engineering research by geographically separated participants. One of the most promising goals is to use these technologies to extend human intellectual capabilities in medical decision making. These emerging technologies are poised to drastically reduce healthcare cost by providing service at remote locations. This also increases diagnosis capacity since information is made available to experts at any location. In this paper, we propose a novel application of a recently developed interactive and distributed system in medical consultation and education. Our approach builds on the notion that interactive and distributive capabilities of the system are crucial for medical consultation and education. The presented application uses a multiuser, collaborative environment with multimodal human/machine communication in the dimensions of sight, sound, and touch. The experimental setup, consisting of two user stations, and the multimodal interfaces, including sight (eye-tracking), sound (automatic speech), and touch (microbeam pen), were tested and evaluated. The system uses a collaborative workspace as a common visualization space. Users communicate with the application through a fusion agent by eye-tracking, speech, and microbeam pen. The audio/video teleconferencing is also included to help the radiologists to communicate with each other simultaneously while they are working on the mammograms. The system used in this study has three software agents: a fusion agent, a conversational agent, and an analytic agent. The fusion agent interprets multimodal commands by integrating the multimodal inputs. The conversational agent answers the user's questions and detects human-related or semantic errors and notifies the user about the results of the image analysis. The analytic agent enhances the digitized images using the wavelet denoising algorithm if requested by the user. To show how well the system performs in practice, we used the system for medical consultation on mammograms. Results also show that the relevant information about the region of interest (ROI) of the mammograms chosen by the users is extracted automatically and used to enhance the mammograms. PMID- 10719540 TI - [Risk of inducing resistance upon stopping and restarting lithium after long-term usage]. AB - Three man, aged 66, 60 and 26 years with bipolar disorder and long periods (20-5 years) of effective lithium prophylaxis had relapses on lithium discontinuation. Once the drug was reinstituted, it was no longer effective. Combinations of lithium with a classical MAO inhibitor or a second mood stabilizer (valproic acid and carbamazepine, respectively) ultimately proved to be successful. The risk of refractoriness should be kept in mind before considering to stop lithium therapy. A better compliance can be achieved by informing patients about the risks of discontinuing lithium, by maintaining a minimal maintenance dose and by adequately coping with possible side effects. PMID- 10719541 TI - ['Cholesterol' guideline (first revision) of the Dutch College of Family Physicians; response from the family practice]. AB - The revised guidelines of the Dutch College of General Practitioners on cholesterol are more in accordance with the consensus guidelines of the Dutch Institute for Health Care Improvement. The number of people with a history of cardiovascular disease who will receive medication is probably underestimated. The guidelines attach little value to diet measurements; however, in individual cases a diet can be a realistic possibility for treatment. Screening for hypercholesterolaemia is now focused more on other risk factors. This and stopping medication in people who have been told in the past they have a treatable disease can turn out to be a problem, for which a national campaign of the public might be helpful. PMID- 10719542 TI - ['Cholesterol' guidelines (first revision) of the Dutch College of Family Physicians; response from internal medicine]. AB - The deviations of the revised standard 'Cholesterol' of the Dutch College of General Practitioners from other recent Dutch and European consensus guidelines are disputable in that it advises, only to measure serum lipids if the person will be eligible for treatment with statins according to the risk tables and to omit repeated measurements, since there exist more reasons for serum cholesterol assay: adequate risk estimation for primary prevention without drug treatment, relation between degree of lipid lowering and reduction in risk, estimation of response and compliance during drug treatment and estimation of type of lipid disturbance. Also, repeated serum cholesterol determination improves detection of persons with hereditary hyperlipaemia. Finally, by refraining in advance from treating older persons the possible gains in individual cases are lost. PMID- 10719543 TI - [New developments in the treatment of diabetic foot ulcers]. AB - Diabetic foot ulcers are frequent: 12,000 Dutch diabetes patients have such an ulcer. The ulcers have a multifactorial aetiology: polyneuropathy, biomechanical stress, infection, deficient footwear and to a less extent ischaemia are the major factors. The principles of ulcer treatment are relief of pressure, restoration of skin perfusion, treatment of infection, intensive wound care, metabolic control, treatment of comorbidity, and instruction of the patient. Wound healing is slow. The impaired wound healing is probably caused by deficiencies in local growth factors, changes in the extracellular matrix, diminished fibroblast function, decreased antimicrobial activity of leukocytes and disturbances in the macro- and microcirculation. In recent years several new treatment strategies have been developed to stimulate wound healing in diabetic foot ulcers. These (partly experimental) treatments include: topical growth factors, extracellular matrix products, bioengineered human skin, granulocyte colony stimulating factor and hyperbaric oxygen therapy. In particular recombinant human platelet derived growth factor (becaplermin) has proved to be clinically effective in chronic neuropathic foot ulcer and has been approved in the Netherlands. PMID- 10719544 TI - [CBO guidelines on diagnosis, treatment, and prevention of complication in diabetes mellitus: retinopathy, foot ulcers, nephropathy and cardiovascular diseases. Dutch Institute for Quality Assurance]. AB - Early detection and adequate treatment of complications of diabetes mellitus (DM) are important for many patients in maintaining independence and ability to work. Diabetic retinopathy cannot be prevented. Limitation of damage is possible by aiming for normoglycaemia and normotension. While exudative as well as proliferative retinopathy can occur without any visual symptom, regular ophthalmological examination is necessary for timely laser coagulation. Fundus photography for screening is applicable under certain conditions; fluorescence angiography can be useful in patients with understood deterioration of visual acuity or diabetic maculopathy. In many patients foot disease can be prevented by simple measures: examining the foot at least once a year, recognition of the foot with a high level of risk, education of patient and family, adapted shoes and preventive foot care. Treatment of a foot ulcus consists of relief of mechanical pressure, repair of disturbed skin circulation, treatment of infection and oedema, optimal metabolic control, frequent local wound care and education. Patients with a diabetic foot have to be thoroughly followed up for the rest of their lives. For patients with diabetic nephropathy cardiovascular complications are the main causes of morbidity and mortality. Of all patient with DM older than 10 years urine has to be examined for loss of albumin at least once a year. Treatment of nephropathy consists of non-smoking, sufficient physical exercise, reduction of overweight, well-composed nutrition and particularly treatment of hypertension. Diagnosing cardiovascular diseases in patients with DM is in principle the same as for other patients. Treatment of hypercholesterolaemia has to be based on an absolute risk of 20% for cardiovascular disease in the following 10 years. The limit for treatment will be reached earlier in the presence of microalbuminuria, persistent high HbA1c > 8.5%, triglyceride concentration > 2.0 mmol/l, or a positive family history with myocardial infarction < 60 years. In proven cardiovascular disease one needs to strive for optimalization of the glucose metabolism, non-smoking and if necessary drug therapy. PMID- 10719545 TI - [Diabetic peripheral neuropathy: international guidelines for prevention, diagnosis, and treatment]. AB - Recently, international guidelines for the diagnosis and management of diabetic polyneuropathy were issued. The aim of this initiative was to develop simple and practical guidelines for general practitioners and hospital physicians in day-to day practice. Regular assessment of peripheral nerve function can be performed in diabetic patients using simple diagnostic tools. In such a way, patients will become more aware of the risks associated with diabetic neuropathy. Early detection of neuropathy and other risk factors for the development of foot ulcers may lead to avoidance of amputations. The guidelines emphasize foot care education of diabetic patients with or without neuropathy and early referral of patients with ulcerations to specialised outpatient foot clinic. PMID- 10719546 TI - [Summary of 'Cholesterol' guideline (first revision) of the Dutch Society of Family Physicians]. AB - The revised guidelines on cholesterol of the Dutch College of General Practitioners (DCGP), which closely follow the consensus of the Dutch Institute for Health Care Improvement, provide thresholds for treatment with statins in patients with elevated risks for coronary heart disease (CHD): patients with a history of cardiovascular disease, with an annual CHD risk larger than 2.5-3%, or with a (suspected) hereditary lipid disorder. Unlike the consensus the DCGP guideline advises only to determine a total cholesterol/HDL cholesterol ratio if the accompanying risk table indicates that the patient might fall in the range where drug treatment is indicated. For this purpose an extra column has been added to the table. In patients with a possible hereditary lipid disorder a higher threshold for referral to a specialist is used because moderately raised levels are common in the population and indicate a familial lipid disorder in only part of the cases. PMID- 10719547 TI - [Patients without referral treated in the emergency room: patient characteristics and motives]. AB - OBJECTIVES: To determine the motives of patients to directly visit the Emergency Department for medical treatment without seeing their family practitioner first and to establish the characteristics of this group. DESIGN: Prospective, descriptive. METHOD: Of the surgical patients attending the Emergency Room (ER) of Utrecht University Hospital, the Netherlands, between 1st April and 25th June 1997, the demographic and diagnostic data of the non-referred patients were compared with those of patients referred by the general practitioner (GP). The non-referred patients were asked about their reasons to attend without referral. RESULTS: A total of 1026 patients visited the ER at their own initiative: 603 males and 423 females. The most frequent reasons for the direct visits were 'had not thought of their family practitioner' (48%) and 'wants help that can only be provided in a hospital' (30%). The self-referring patients differed from the 962 referred patients in severity of the trauma: 76% versus 39% had a relatively minor trauma; sort of trauma: 19% versus 4% had a sport trauma; place of residence: 57% versus 45% lived in the near vicinity of the hospital. Of the self referring patients 57% visited the hospital out of office hours, versus 39% of the referred patients. CONCLUSION: A large part of the non-referred patients visited the ER unnecessarily and should actually have consulted their GP. These were mainly young adults with minor injuries or sports injuries who lived close to the hospital. Important factors were unfamiliarity with or lack of understanding of the existing regulations. PMID- 10719548 TI - [Information needs of men with prostate cancer and their partners]. AB - OBJECTIVE: To determine the need for information of men with prostate cancer and their spouses. DESIGN: Descriptive. METHOD: In February and March 1998 three questionnaires, especially developed for this study, were administered to 40 men treated for prostate cancer and 24 spouses in the Antoni van Leeuwenhoek hospital, Amsterdam, the Netherlands. The questionnaires concerned need for information and knowledge of prostate cancer. RESULTS: In general, patients and spouses indicated to want as much information as possible about prostate cancer and its treatment. Such needs decreased with increasing age. The need for medical technical information was found to be higher than the need for psychosocial information. Spouses had significantly more need for information than patients, especially psychosocial information. This difference was partly related to age. The patients' needs for information were largely satisfied, while spouses indicated that their needs were often not met. General knowledge of prostate cancer of both patients and spouses was sufficient. CONCLUSION: Men with prostate cancer and their spouses had a substantial need for especially medical-technical information. Their needs decreased with increasing age. The need of spouses was higher than that of patients. Patients' needs were largely satisfied, whereas those of spouses were not always met. Psychosocial information in particular was inadequate. PMID- 10719549 TI - [Diabetes mellitus after treatment with clozapine]. AB - In a 40-year old patient with schizophrenia treated with sustained-action perfenazine for schizophrenia, diabetes mellitus type 2 and hypertension developed when clozapine was added to the regimen. The blood sugar values and the blood pressure normalized when clozapine was discontinued two weeks later. When he was 44 years old, clozapine was resumed as the psychotic symptoms did not subside. Diabetes mellitus reappeared and did not disappear after discontinuation of the clozapine. Insulin therapy was necessary. An association between the emergence of diabetes mellitus and the use of clozapine is suggested. Previously 15 cases of diabetes mellitus emerging during treatment with clozapine were described in the international literature. Monitoring of blood glucose is advised in patients with a positive family history of diabetes mellitus or with a personal history of impaired glucose tolerance. PMID- 10719550 TI - [1999, the year of fired editors-in-chief]. PMID- 10719552 TI - [CBO guidelines for diagnosis and treatment of acute ankle injury]. PMID- 10719553 TI - [CBO guidelines for diagnosis and treatment of acute ankle injury]. PMID- 10719554 TI - [Revised CBO guideline 'Urinary tract infections']. PMID- 10719555 TI - [Revised CBO guidelines 'Urinary tract infections']. PMID- 10719556 TI - [Truth after death; autopsies as a valued investigative tool]. PMID- 10719557 TI - [Less medicalization of obstetrics in West Friesian islands than on the mainland]. PMID- 10719558 TI - [Less medicalization of obstetrics in West Friesian islands than on the mainland]. PMID- 10719559 TI - [Less medicalization of obstetrics in West Friesian islands than on the mainland]. PMID- 10719560 TI - [Blood group immunization during pregnancy in Netherlands]. PMID- 10719561 TI - The effect of lactase and formula reconstitution on milk osmolality. AB - These experiments investigated the reaction rate of lactase on milk lactose by measuring milk osmolality; and explored the effect of formula reconstitution on milk osmolality. The investigations measured milk osmolality with the Fiske Os, freezing-point osmometer. Lactase (Lactaid) incubated with pure lactose solutions established the validity of the method. Lactase was incubated for 24 hours with four reconstituted milk formulas (Milumil, and Cow and Gate Nutrilon Plus, Farley's First Milk, SMA Gold). Milk osmolality increased most rapidly in the first 4 hours after the addition of lactase. The lactase enzyme completed over 90% of the reaction within 12 hours. The milk osmolalities ranged from 487 to 591 mosm/kg after 24 hours with 2-4 drops of lactase in 240 ml of formula. A clinical guideline osmolality of 400 mosm/kg was reached in 240 ml of formula at 1 to 12 hours depending on the dose of lactase. High milk osmolalities due to prolonged enzyme incubation, or high lactase doses could be reduced to around 400 mosm/kg by dilution of 240 ml of formula with an extra 60 ml of water. The initial osmolality of formula after reconstitution by paediatric nurses varied widely and usually exceeded the manufacturer's quoted osmolality. This initial osmolality was a further influence on the final osmolality reached after the addition of lactase. It is concluded that the recommended incubation time for Lactaid of 24 hours is unnecessary as lactase exerts the majority of its effect in less than 12 hours. Adjustment of Lactaid dose and incubation times will maintain milk formula osmolality within standard guidelines. Dilution with extra water will correct inadvertent high enzyme doses and prolonged incubation times. The normal method of reconstituting milk formulas from powder may be unreliable as the manufacturer's quoted osmolality was not reproduced when milk formulas were reconstituted by paediatric nurses. PMID- 10719562 TI - Adsorption of direct-acting and indirect-acting mutagens by various dietary fibers. AB - The protective effect of dietary fiber on human cancer has received great attention during the last decades. Because dietary fiber constitutes a large group of complex polysaccharides with various solubilities, degrees of lignification, chemical compositions and structural arrangements, several mechanisms for their effects have been proposed. In this study, in vitro binding capacities of various dietary fibers (potato fiber and glucomannan) and dietary fiber constituents (pectic acid and cellulose) against indirect mutagen 2-amino-3 methyl-3H-imidazo (4,5-f) quinoline (IQ) and direct-acting mutagen sodium azide were investigated. Direct-acting mutagen sodium azide was not adsorbed to the dietary fiber and dietary fiber constituents of 0 degree C, pH 4.5 and 37 degrees C, pH 7.0. However, indirect-acting mutagen 2-amino-3-methyl-3H-imidazo (4,5-f) quinoline (IQ) were sorbed by them in variable ratios at 0 degree C, pH 4.5 and 37 degrees C, pH 7.0. The differences between the in vitro binding capacities of the samples at two experimental conditions were found to be statiscially significant (P < 0.01). IQ was not released from the dietary fibers and constituents in distilled water. PMID- 10719563 TI - Chemical composition, nutritionally valuable minerals and functional properties of benniseed (Sesamum radiatum), pearl millet (Pennisetum typhoides) and quinoa (Chenopodium quinoa) flours. AB - The chemical composition, nutritionally valuable minerals and functional properties of benniseed (Sesamum radiatum), pearl millet (Pennisetum typhoides) and quinoa (Chenopodium quinoa) were studied. The results showed that the samples contained crude protein in the range of 11.4 to 22.5% with benniseed having the highest value of 2.5%, and pearl millet with the lowest value 11.4%. Ether extracts fall within the range of 6.3-44.3%. The moisture contents ranged from 5.2 to 11.2% while the ash contents were found to be in the range of 1.2 to 4.1% and the crude fibre ranged between 3.1 and 9.6%. The flours were relatively higher in maltose and D-ribose which were found to be in the range of 1.28-5.08 mg sugar in 5 ml sample. They also have low contents of glucose and fructose which ranged between 0.70 and 1.46 mg sugar in 5 ml sample. The predominant mineral was potassium which varied between 5150 and 7140 mg per kg sample while the samples were significantly low in manganese and copper. The protein solubility of the flours were found to have minimum solubility at pH 5 for benniseed, about pH 6 for pearl millet and quinoa. The seed flours also have good gelation property, water absorption capacity, emulsion capacity and stability. The oil absorption capacity and foaming capacity were low but the foams were relatively stable. PMID- 10719564 TI - Nutritional quality and the presence of anti-nutritional factors in leaf protein concentrates (LPC). AB - Leaf protein concentrates were prepared from 25 different plant species. The concentrates contained acceptable levels of all essential amino acids; except methionine which was limiting with chemical scores ranging between 13.5-96%. LPCs from sesbania, nebergrass, lufa and sweet pepper were lethal to the rats. PER ranged between 0.28 for safflower LPC up to 1.95 for rape. Positive nitrogen balance was obtained with diets based on LPCs with mean values ranging between 6.4 and 22.42 mg N per day. True nitrogen digestibility fluctuated between 72.4% for sweet potato LPC up to 97.9% for LPC prepared from safflower. BV ranged between 60.5 for sunflower LPC to 97.26 for carrot LPC. NPU values of LPCs were 49.4 for sunflower LPC up to 86.5% for rape LPC. There was a positive correlation coefficient between BV and the respective NPU. Highest utilizable protein was obtained with turnip LPC. Tested LPC's contain 0.6-6.8% tannin. Saponin was absent in 11 LPCs. Lufa LPC showed the highest saponin content followed by nebergrass and sesbania LPCs. Trypsin inhibitory activity was quite low. Prelimenary results show that rats consumed diet based on lufa, nebergrass and sesbania LPCs died within the first few days of the experiment. Low growth rates were obtained for drum-dried samples prepared from carrot, but this was not the case for drum-dried samples prepared from chard. The activity of liver alanine amino transferase was also determined and indicated toxicity of some LPCs to liver. PMID- 10719565 TI - Nutritive value of various breads in Saudi Arabia. AB - The nutritive value of nine Saudi breads prepared from wheat, millet and corn were measured chemically by proximate, minerals and vitamins analyses. On fresh weight basis, the bread contained 26.4-44.7% moisture, 6.6-10.4% protein, 0.4 2.4% fat, 40.2-60.6% available carbohydrates, 1.8-5.7% dietary fibre, 0.6-2.4% ash and 190-273 Kcal (metabolizable) per 100 g. All the breads were low in Ca (2.2-12.5 mg/100 g), P ranged from 41.9-320.8, Na 83.2-794.6, K 0.7-224.2 and Fe 1.6-7.8 mg/100 g. The contents of vitamin A (RE), thiamin and riboflavin ranged from 0-145 micrograms, 0.01-0.26 mg, 0.02-0.13 mg/100 g respectively. The bread contributed 12-18, 2-8 and 77-84% of the total food energy from protein, fat and carbohydrates respectively. Wheat bread (355 g/head/day) provided 45 and 61% of energy and protein requirements respectively at national level per person per day. PMID- 10719566 TI - Calcium bioavailability of selected Egyptian foods with emphasis on the impact of fermentation and germination. AB - The bioavailability of calcium (Ca) was assessed in 11 foodstuffs of plant or animal origin using rat feeding experiments and the criteria used for assessing the bioavailability were femur Ca and calcium efficiency. The bioavailability of Ca was found to be highest in fishes (Melouha) and cheeses (Mesh) fermented under local processing techniques. Germination of faba beans also enhanced the bioavailability of calcium to a mean value quite comparable to those of some dairy products, such as cottage cheese. The present study clearly demonstrates that the processes of fermentation and germination of selected foods are associated with an enhancement in the bioavailability of calcium. It is suggested that the breakdown of complex proteins under the fermentation or germination process is the underlying mechanism of action. PMID- 10719567 TI - Fatty acid composition of six varieties of dehulled African yam bean (Sphenostylis stenocarpa) flour. AB - The proximate and fatty acids compositions of dehulled African yam bean (Sphenostylis stenocarpa, Hochst ex A. Rich. Harms) flour were reported. The crude protein values ranged from 20.18 +/- 0.02 to 25.78 +/- 0.05 g/100 g; the ether extract values ranged from 1.93 +/- 0.05 to 10.18 +/- 0.04 g/100 g; crude fibre values ranged from 1.61 +/- 0.02 to 2.38 +/- 0.00 g/100 g; total ash ranged from 2.06 +/- 0.03 to 2.36 +/- 0.05 g/100 g and carbohydrate values ranged from 58.46 +/- 0.04 to 63.34 +/- 0.05 g/100 g. The values of the moisture content ranged from 3.20 +/- 0.03 to 7.10 +/- 0.02 g/100 g. Significant differences were found (P < 0.05) among the samples in the proximate compositions. The most concentrated fatty acids were palmitic acid (18.18 to 19.78%) < linoleic acid (28.33 to 35.16%) < stearic acid (29.01 to 36.71%). Caprylic, capric, lauric, myristic, palmitoleic, oleic, eicosenoic and erucic acids were present in small quantities with none of them recording up to 2.00% in any of the samples. However, the values of alpha-linolenic acid ranged from 2.01 to 2.96%. Significant differences were observed in the fatty acid compositions among the African yam bean dehulled seed cultivars. PMID- 10719568 TI - Effect of insulin-like growth factor-1 (IGF-1) on muscle and bone growth in experimental models. AB - The effect of protein restriction on the growth of muscle, bone and body tissue of rats fed a diet containing 5% protein (experimental group) and 12% protein (control group) was studied. The following parameters were analyzed: body, muscle and cartilage growth, tibial length, plasma insulin IGF-1 concentrations, and tissue proteoglycan, protein, RNA and insulin-like growth factor-1 concentrations. Protein synthesis in tissues was also determined. Except for protein concentration in the two tissues and of insulin-like growth factor-1 in cartilage, all other values differed significantly between groups. The lower ration and protein ingestion by group 1 receiving a 5% case in diet provoked a reduced secretion of the anabolic hormones insulin and IGF-1 and a lower synthesis of proteoglycans, RNA and protein, impairing total body growth and growth of the tissues analyzed. PMID- 10719569 TI - Prevalence of nutritional anaemia among primary school girls in Riyadh City, Saudi Arabia. AB - The prevalence of iron deficiency anaemia was investigated among 1,210 school girls in Riyadh, Saudi Arabia. Anthropometric measurements were recorded. A dietary and socio-economic status questionnaire was administered and heamoglobin (Hb), serum iron and ferritin were estimated. Wasting and stunting is common among the 7- and 14-year-old girls. A total of 8.5% of children showed Hb level below 10 g/dL while 55.4% showed Hb level below 12 g/dL. It was found that 26.3% of the girls have serum iron below 10 mumol/dL while 16.1% of subsample of the girls showed serum ferritin level of less than 12 micrograms/dL. The most affected ones are those in the age group of 7-14 years old. The dietary questionnaire revealed that 16.5% of the girls did not take breakfast at home and depend on snacks offered in the school canteen which consist mostly of biscuits, chocolate bars, potato chips and carbonated cola drinks. No association between education of mother and father and breakfast consumption at home was detected. Tea drinking is common among these girls while fresh fruits and vegetable consumption is infrequent. Iron deficiency anaemia is highly prevalent among these schoolgirls which seriously affects the growth of 7- and 14-year-old girls. An in-depth investigation of the etiological factors of iron deficiency is urgently needed and meanwhile a suitable iron supplementation program is recommended. PMID- 10719570 TI - Intestinal degradation of dietary fibre in green beans--effects of microwave treatments. AB - Intestinal degradation of dietary fibre in blanched and microwaved green beans was studied by using a rat experimental model. Content and composition of dietary fibre as well as molecular weight distribution of water-soluble polysaccharides (WSP) were analysed. There was a solubilization and a shift towards lower Mw of mainly uronic acid-containing polysaccharides with repeated microwave treatment in the raw material. Thus, the apparent Mw of water-soluble polysaccharides decreased from approximately 1,550,000 to approximately 300,000. After the beans had been digested the Mw of the WSP was significantly reduced, to approximately 100,000, and the differences in Mw seen between various processed raw materials had been evened out. After fermentation the Mw of the WSP decreased further approximately 10 times. Fibre fermentability was high (approximately 90%) and similar for the various processed beans. PMID- 10719571 TI - Fatty acid composition and arachidonic acid intake of selected Saudi foods. AB - Twenty-eight Saudi Arabian common foods were analyzed for their fatty acid contents by gas-liquid chromatography using capillary column. The predominant fatty acids were oleic, palmitic, linoleic and stearic acids. The polyunsaturated to saturated fats ratio (P:S) as well as the n-3:n-6 ratio were generally not compatible with recommended values. Assessment of C20:4 (n-6) content in those foods showed a noticeable variation ranging from 9 mg/100 gm in beef sirloin to 256 mg/100 g in kannad fish. Eicosapentaenoic acid (C20:5) n-3 content was highest in kannad (925 mg/100 g). Applying C20:4 (n-6) values determined in this work to Saudi Balanced Sheet, the mean C20:4 (n-6) intake for a Saudian was estimated as approximately 115 mg/day. PMID- 10719572 TI - Fluoride concentration in foods from Iran. AB - Knowledge of fluoride intake is important in optimizing the caries-preventive role of fluoride, and the measurement of fluoride intake usually requires information on the fluoride concentration in foods and drinks. Most information comes from developed countries and there are no data on fluoride concentration in foods in Middle Eastern countries including Iran. The aim of the study was to: (a) describe a modification to the silicon-facilitated diffusion method for determining fluoride concentration, (b) provide information on fluoride content of foods in Iran to supplement food tables for the Middle East, and (c) determine the effect of variation in the fluoride concentration of drinking water on the fluoride concentration of prepared foods. Five hundred and ten samples of 84 popular foods and drinks were collected from three areas of Iran where water fluoride concentrations were 0.32, 0.58 and 4.05 mg/L. The mean recovery of fluoride added to food samples before diffusion was 98 +/- 5%. Values for duplicate analysis of 20 food samples were within 0.03 microgram F/g. Most of the samples of foods and drinks came from the area with 0.32 mg F/L in water supplies. For 30 of the 84 items, fluoride concentration was below 0.1 microgram/g. Fluoride concentrations in the cereals group (which constituted much of the diet) were mainly between 0.2 and 0.3 microgram/g, when prepared for consumption. It was concluded that: (a) modification of a published method for determining fluoride concentration of foods appeared to be an advance, (b) values for fluoride concentration of foods commonly consumed in Iran showed variation between groups but were in broad agreement with published data, and (c) concentration of fluoride in water influences positively fluoride concentration in foods cooked in water, but the increase in foods was less than the increase in fluoride concentration in water. PMID- 10719573 TI - Evaluation of the nutritional quality and acceptability of fingermillet-based tempe as potential weaning foods in Tanzania. AB - Six types of fingermillet (Eleusine coracana)-based tempe were developed by incorporating either commonbeans, groundnuts, cowpeas, mungbeans, chickpeas, sesame and/or their mixtures and fermented by Rhizopus oligosporus. The proximate and mineral composition was not changed significantly by fermentation. The protein content of tempe ranged from 13.3 to 15.7%; the total energy, 374.1 to 434.3 Kcal/100 g; the protein energy, 13.0 to 15.6%; the net dietary protein energy, 7.2 to 8.7%; and the chemical score, 61 to 71%. Tempe processing reduced the tannin and hydrogen cyanide content by 55.2 to 75.7, and 71.0 to 86.2%, respectively. It increased the content of reducing sugars, amino nitrogen, total acidity by 4 to 15, 3 to 6.9, and 4.3 to 12-fold, respectively. The in vitro protein digestibility was also improved. The developed tempe had protein quality and energy content recommended for weaning foods. The deep-fried tempe snacks were fairly organoleptically acceptable. PMID- 10719574 TI - Carbohydrate availability of arroz caldo with lambda-carrageenan. AB - Total available carbohydrate (sugars and starches) and total dietary fiber (soluble and insoluble) make up the total carbohydrate content of a food. Soluble fiber decreases the availability of glucose by delaying its absorption in the proximal small intestine, thus reducing the postprandial glucose levels (Jenkins et al., 1978; Schneeman, 1987a). Carrageenan, a seaweed extract, is a good source of soluble fiber (Montano et al., 1985). This study aimed to determine the effect of carrageenan incorporation into arroz caldo on carbohydrate availability by monitoring the postprandial blood glucose levels of normal subjects. Control and experimental arroz caldo samples were prepared and subjected to proximate analysis and feeding studies. The total dietary fiber (TDF) content of the experimental (2.03%) was about thrice that of the control (0.68%). Using randomized crossover design, preweighed 55 g available carbohydrate serving portions of control and experimental arroz caldo samples, with 3.45 and 14.84 g TDF, respectively, were fed to ten fasting normal subjects then their postprandial blood glucose levels were determined at 15, 30, 45, 60 and 90 min intervals. Results of the short-term in vivo study showed that the mean postprandial glycaemic responses of subjects after consuming the experimental sample were significantly lower than the levels after consuming the control at 15, 45, and 90 min (P < or = 0.05) and at 30 min (P < or = 0.001). Likewise, the mean glucose area under the curve was significantly lower (P < or = 0.01) after consumption of experimental (69.22 +/- 32.94) arroz caldo than control (147.29 +/ 53.34). The hypoglycaemic effect of carrageenan may prove useful in the prevention and management of metabolic conditions such as diabetes. PMID- 10719575 TI - Studies on the antioxidant activity of milk caseins. AB - The antioxidant properties of milk casein subunits (alpha-casein, beta-casein and kappa-casein) were evaluated in liposomal models. All the subunits of casein are able to inhibit Fe-induced peroxidation of arachidonic acid inserted into multilamellar liposomes of dipalmitoylphosphatidylcholine (0.2 mM and 0.8 mM, respectively). The peroxidation was monitored as thiobarbituric acid reactive substances, and the strongest inhibitory effect occurred when 500 micrograms of alpha-casein were added to 0.5 ml of liposomal suspension. At this concentration, peroxidation was completely inhibited in our experimental conditions (incubation for 2 h at room temperature, with a mixture of ferrous sulfate and ascorbate, 50 and 500 microM final concentration, respectively). The mechanisms of antioxidant action are complex, but the strongest effect is achieved by modifying the Fe2+/Fe3+ equilibrium; in fact, caseins seem to favour the autoxidation of iron, and thus inhibit lipid peroxidation. PMID- 10719576 TI - Reduction of the bioavailability of fluoride from Antarctic krill by calcium. AB - The bioavailability of fluoride from krill exoskeleton and the effect of additional calcium on the bioavailability of fluoride from krill paste were evaluated using young rats. Fluoride from the exoskeleton showed a high apparent absorption of 80%. Approximately 3.6% of this fluoride was deposited in the femur, and 44% in the rest of the carcass. The presence of 1.5% and 2.5% calcium (CaCO3) in the diet--1.0 and 2.0% above the minimum recommended content respectively--significantly reduced the bioavailability of fluoride from krill paste. The apparent absorption of fluoride from the paste was reduced by 33.8 and 45.8%, deposition in the femur by 48.1 and 58%, and retention in the rest of the carcass by 44.4 and 55.6%, respectively. The need for further studies using lower amounts of fluoride and calcium and other chemical forms of calcium is indicated. PMID- 10719577 TI - Assessing the contribution of soybean utilisation on the nutritional status of children from low-income families: Lagos State, Nigeria as a case study. AB - Questionnaires were administered to mothers from 120 randomly selected households. Anthropometric measurement of 258 children from these households were taken, to assess the contribution of soybean in the diet of the infants from low income families in Lagos State (Imota, Idimu and Badagry), Nigeria. The result showed that majority of the mothers either have no formal education or were primary-school leavers. Most mothers were categorised into low-income earners because they earn about N500 (US$6) per month. The frequency of soybean consumption in Idimu and Imota is significantly higher (P < 0.05) than that of Badagry. Although the frequency of animal protein consumption in Badagry households is significantly higher (P < 0.05), it is found to be inadequate to meet their daily requirements. The protein intakes were between 92-110, 97-100 and 75-103% of FAO requirements for the children in Idimu, Imota and Badagry respectively. The weight and height of children from Idimu and Imota are higher than the children from Badagry. About 26, 32 and 71% of the children studied in Idimu, Imota and Badagry respectively fell below the WHO 5th centile bracket of weight for age. These differences are attributed to the introduction and contribution of soybean (a cheaper source of protein) to Imota and Idimu. PMID- 10719578 TI - Street foods and dietary intakes of Nigerian urban market women. AB - The food and nutrient intake of 197 market women aged 19-66 years (mean 41 years) have been determined using questionnaire and 24-hour dietary recall. The contributions of street foods (SF) to total intake of food and nutrients were determined in order to quantify the importance of SF as a source of food and nutrients for these women. Eighty-three of the women were of child-bearing age (< 49 years), Group I, while 114 were not (> 49 years). Overall, SF provided 63% of the weight of total food consumed. The mean energy intake was 11.5 MJ with the older women Group II consuming significantly (P < 0.05) higher than the younger age group (Group I). The contribution of SF to energy intake was similar (59%) for both groups. The older women (Group II) also consumed significantly (P < 0.05) more total protein than the younger women (110 vs 80 g) although SF contributed similar percentages (58 vs 59%). Mean calcium intake was high for the subjects (618 mg) but no significant differences were found between the two age cohorts. The proportion of calcium intake supplied by SF was 79% for Group I and 81% for Group II. There was a wide variation (8.1-16.5 mg) in total daily iron intake. Overall SF provided 57% of the pooled mean intake (12 mg). Mean iron intake for Group II was significantly (P < 0.05) higher than that of Group I. The contribution of SF to the intakes of vitamins was above 50% except for thiamin (47%) in the younger age cohort and vitamin A (46%) in the older age group. The results suggest therefore that street foods constitute the major sources of dietary energy and other nutrients for market women in Nigeria. PMID- 10719579 TI - Efficacy of a preschool breakfast program in reducing refined sugar intake. AB - The objective of this study was to measure nutritional differences between breakfasts eaten at home and those eaten in a preschool setting, which were prepared and served following the guidelines of the School Breakfast Program (SBP). During the fall semester, baseline data on children's breakfasts were obtained from families of 3- and 4-year-old children attending our laboratory school. Two weeks later, we began providing breakfasts for the children at preschool, before they began their school day. Breakfast menus varied but were always in compliance with SBP guidelines. Six weeks after initiating the SBP, we recorded breakfasts eaten at school, and the entire procedure was repeated during the spring semester. Children's intake of macronutrients from the breakfast meal was altered through participation in the SBP. During each intervention period, the consumption of starch-rich foods and fibre increased while the intake of simple sugars decreased. Protein, fat, and micronutrient intake were not affected. The implementation of the SBP appeared to be an effective way to increase the intake of starch-rich foods and fibre in a low-risk sample of preschool-aged children who regularly are breakfast prior to their participation in this investigation. PMID- 10719580 TI - Consumer preference optimization of scallopini made from boneless, skinless chicken thighs. AB - Scallopini is a general name derived from scallopine, an Italian preparation of veal, which describes thinly pounded cuts of meat which are either used as a cutlet or rolled around other foods and cooked. Concept development was accomplished through use of focus groups, consumer ranking studies, and mailed surveys. The objective of this study was to assess consumer acceptability of chicken scallopini at various thicknesses (4, 8 and 12 mm) and concentrations (0, 1, and 2%) of added sodium tripolyphosphate (STPP). A 3 x 3 factorial design with three replications was adopted for use in this study. To assess consumer acceptability of packaged products, 53 participants were presented with three packages of the raw scallopini packaged on white styrofoam trays and overwrapped with film. Because STPP produces no visual change in the product at the levels used, only product thickness was varied. The mean price suggested by panelists from the sensory evaluation was $1.28/lb. The average price per pound suggested for the raw, packaged product was $1.62. For the attributes of overall acceptability, tenderness, juiciness, flavor, size, and texture, the mean ratings were high, indicating an acceptable product in any of the nine treatments. Sensory attributes were affected by STPP concentration but not by scallopini thickness. As STPP concentration increased, hedonic ratings for tenderness, texture, juiciness, flavor, overall acceptability, and purchase likehood increased. Purchase likelihood was rated highest for scallopini containing 2% STPP. Purchase likelihood was verified through use of a simulated supermarket simulation test. PMID- 10719581 TI - Comparison of solubility of pea protein hydrolysate by three analytical methods. AB - Pea protein hydrolysate was obtained by enzymatic hydrolysis with trypsin. The degree of hydrolysis (DH) was controlled by using the pH-stat method. Solubility of the trypsin-treated hydrolysate was tested at nine different pH values starting from 2 up to 10. Protein determinations were carried out using Kjeldahl, Lowry and modified Lowry methods. The results revealed that samples analysed with either the Kjeldahl or Lowry method gave similar values. However, systematic consistent differences existed for those results obtained by the Kjeldahl and the modified Lowry as well as between those results obtained by the Lowry and the modified Lowry. PMID- 10719582 TI - Allied studies on the effect of Rosmarinus officinalis L. on experimental hepatotoxicity and mutagenesis. AB - The hepatoprotective and antimutagenic effects of the rosemary essential oil and the ethanolic extract were investigated using carbon tetrachloride and cyclophosphamide as hepatotoxic and mutagenic compounds, respectively. Our results revealed that i.g. administration of the rosemary ethanolic extract (0.15 g/100 g BW) to rats for 3 weeks produced the most pronounced hepatoprotective effect compared to silymarin (reference compound) due to the amelioration of most of the studied serum and liver parameters and confirmed by histopathological examination of the liver tissue. Pretreatment of mice for 7 days with the rosemary essential oil (1.1 mg/g BW) followed by i.p. injection with cyclophosphamide reduced significantly the induced mitodepression in the bone marrow cells of the animals. The potential hepatoprotective and antimutagenic activities of the rosemary ethanolic extract and essential oil, respectively, are attributed to the presence of a relatively high percentage of phenolic compounds with high antioxidant activity (according to our chemical studies). PMID- 10719584 TI - Development of a screening tool for detecting undernutrition and dietary inadequacy among rural elderly in Malaysia: simple indices to identify individuals at high risk. AB - Undernutrition and the consumption of poor diets are prevalent among elderly people in developing countries. Recognising the importance of the early identification of individuals at high nutritional risk, this study aimed to develop a simple tool for screening. A cross-sectional study was conducted on 11 randomly selected villages among the 62 in Mersing District, Malaysia. Undernutrition was assessed using body mass index, plasma albumin and haemoglobin on 285 subjects. Dietary inadequacy (a count of nutrients falling below two thirds of the Recommended Dietary Allowances) was examined for 337 subjects. Logistic regression analysis was performed to identify significant predictors of undernutrition and dietary inadequacy from social and health factors, and to derive appropriate indices based on these predictions. The multivariate predictors of undernutrition were 'no joint disease', 'smoker', 'no hypertension', 'depended on others for economic resource', 'respiratory disease', 'perceived weight loss' and 'chewing difficulty', with a joint sensitivity of 56% and specificity of 84%. The equivalent predictors of dietary inadequacy were 'unable to take public transport', 'loss of appetite', 'chewing difficulty', 'no regular fruit intake' and 'regularly taking less than three meals per day', with a joint sensitivity of 77% and specificity of 47%. These predictions, with minor modification to simplify operational use, led to the production of a simple screening tool. The tool can be used by public health professionals or community workers or leaders as a simple and rapid instrument to screen individual at high risk of undernutrition and/or dietary inadequacy. PMID- 10719583 TI - Heavy metal accumulation in Mullus barbatus, Merluccius merluccius and Boops boops fish from the Aegean Sea, Turkey. AB - The accumulation behaviour of copper, zinc, cadmium and lead concentrations in flesh, gills, liver and gonads of three commercial fish Mullus barbatus, Merluccius merliccius and Boops boops from the Aegean Sea in Turkey have been studied. Copper, zinc and lead concentrations in flesh were found low, while in gonads Cd was found high. Liver showed higher concentration of Cu than other fish organs. Gonads and liver recorded high concentrations of Cd. Liver and gills revealed high levels of Pb as compared to other fish organs. The organs of Boops boops revealed high accumulation of heavy metals as compared to other fishes. Heavy metals in the southern Aegean Sea were found to be higher around Cesme and Fethiye than the northern part. Concentration of heavy metals in the tested organs were within the allowable limit of world levels. PMID- 10719585 TI - Breakfast cereal consumption and subjective reports of health. AB - The aim of the present study was to attempt to replicate and extend a recent result which showed that breakfast cereal consumption was associated with better mental health. The general population sample studied here (262 volunteers aged between 21 and 85 years, mean age: 60.9 years) was older than the sample in the previous study. The results showed that those who consumed breakfast cereal every day reported better mental and physical health than those who consumed it less frequently. This association was still present when demographic factors, indicators of lifestyle, such as smoking, or other aspects of diet were covaried. Further research is now required to elucidate the mechanisms underlying this robust association between daily breakfast cereal consumption and well-being. PMID- 10719586 TI - Survival of some species of Salmonella and Shigella in mukumbi, a traditional Zimbabwean wine. AB - Mukumbi is a traditional Zimbabwean wine prepared from a fruit called mapfura by the Shona people of Zimbabwe and amaganu by the Ndebele. The majority of people in Africa call the tree marula (Sclerocarya birrea subspecies caffra). The survival of Salmonella group B, Salmonella enteritidis, Shigella sonnei and Shigella flexneri in unfermented and fermented mapfura (marula) juice (mukumbi) was investigated. It was found that within 30 min of inoculation, there were no longer any viable pathogens in the fermented mapfura juice whilst in the unfermented juice, more than 10(4) cfu/ml were still viable after 8 h. When lactic acid (0.25 mg/ml) was added to the unfermented mapfura juice, more than 10(4) cfu/ml were also still viable after 8 h but none were viable after 24 h. The fermented product, mukumbi, has a rapid antimicrobial effect against the pathogens as compared to the unfermented juice and is therefore safe from contamination with these pathogens. It appears thus that the death of the pathogens was due to other compounds besides the low levels of lactic acid. PMID- 10719587 TI - Radiotherapy in pediatric head and neck tumours. AB - According to international protocols, radiotherapy remains a part of the treatment of several pediatric ear, nose and throat tumours. The role of radiation therapy in the treatment of rhabdomyosarcomas, non-Hodgkin's lymphomas, nasopharyngeal carcinomas, osteosarcomas and juvenile nasopharyngeal angiofibromas is reviewed. The main complications of this type of treatment in children, as well as their management, are described. Finally, we discuss how several technical advances (increased fractionation of the dose, various types of stereotactic radiotherapy, use of tridimensional treatment planning systems) help the radiation oncologist to minimize the toxicity of the treatment for healthy tissues. PMID- 10719588 TI - Measuring young children's language abilities. AB - This article deals with the new challenges put on language diagnosis, and the growing need for good diagnostic instruments for young children. Particularly for Dutch, the original English Reynell Developmental Language Scales were adapted not only to the Dutch idiom, but some general ameliorations and changes in the original scales resulted in a new instrument named the RTOS. The new instrument was standardized on a large population, and psychometrically evaluated. In communicating the experiences with such a language/cultural/psychometric adaptation, we hope that other language-minority groups will be encouraged to undertake similar adaptations. PMID- 10719589 TI - Transplantation of tracheal autografts: is a two-stage procedure necessary? AB - BACKGROUND: Tracheal autotransplantation has been shown to be a reliable technique for repairing the hemilaryngectomy defect that includes the hemicricoid cartilage and results from resection of unilateral laryngeal cancer with significant subglottic extension. The technique involves a two-stage procedure of cervical tracheal revascularization by wrapping the trachea in a vascularized radial forearm flap and subsequent tracheal transplantation on a newly created vascular pedicle consisting of the radial artery and vein (1, 2). OBJECTIVES: To experimentally (rabbits) investigate if a one-stage tracheal revascularization and transplantation procedure could be a viable option. METHODS: Tracheal patch autografts (1.5 cm x 1 cm) were excised and reimplanted at the anterior cervical trachea with four different patterns of vascular supply: group I: tracheal patch wrapped with vascularized fascia 14 days before excision of the patch (two-stage procedure); group II: tracheal patch wrapped with vascularized fascia at the time of patch excision; group III: tracheal patch without tissue wrapping; and group IV: tracheal patch wrapped with a sheet of Gore-Tex after reimplantation. After orthotopical reimplantation, the patches showed decreasing vascular contacts from group I to group IV. The patches were evaluated morphologically at the moment the animal became dyspnoeic or two weeks after reimplantation in asymptomatic animals. RESULTS: Group I patches fully preserved their viability (median percentage mucosal viability of 100%). Autografts in group II and III showed varying degrees of necrosis and graft take with a median percentage graft viability of 59 and 69% respectively. All group IV tracheal patches showed full thickness necrosis over the majority of their surface area (median percentage graft viability of 10%). CONCLUSION: Fascia enwrapped tracheal autografts show reliable revascularization through the intercartilaginous ligaments only when a 2 stage revascularization technique is used. PMID- 10719591 TI - Cutting forceps in functional endoscopic sinus surgery. AB - Twenty years of Functional Endoscopic Sinus Surgery have brought about a large amount of innovations and a variety of instruments, all claiming to result in better endoscopic procedures and/or better postoperative outcome. An example of such an innovation is the cutting forceps, which has been claimed to result in better wound healing. However, like several other instruments, it has never been proven to be superior to conventional non-cutting forceps techniques. A prospective, randomised double-blind study in 100 consecutive patients, undergoing bilateral FESS, compared the short term clinical outcome of surgery using cutting instruments to that of non-cutting instruments. Symptoms and endoscopic findings were evaluated at four time points during the first three postoperative weeks. Both types of surgery resulted in reduction of symptoms and in endoscopically visible healing over time, but no significant difference between the two methods was found, neither in symptom relief, nor in endoscopic impression of improvement. In conclusion, endoscopic sinus surgery with cutting instruments can be considered effective in resolving sinus disease. However, these instruments do not generate a better healing process than non-cutting instruments in the first weeks postoperatively. PMID- 10719590 TI - Short term effects of antibiotics (Zinnat) after endoscopic sinus surgery. AB - The effects of antibiotics after endoscopic sinus surgery (ESS) were investigated in a prospective, randomized, double blind, placebo controlled study in 202 patients. Hundred and one patients received cefuroxime axetil (Zinnat, 2 x 250 mg/d). The other half received placebo. Patients received no nasal packing and were treated with nasal washes and steroids. Ten subjective symptoms (headache, maxillary pressure, nasal obstruction, nasal secretions, blood, post nasal drip, sneezing, cough, smell disturbances and illness) were recorded daily and at weekly postoperative visits (including suction cleaning when indicated) seven endoscopic parameters were evaluated. There were no statistical differences in the total and individual symptom and endoscopic scores. There was no significant difference in the incidence of postoperative infections between both groups. In conclusion, antibiotics do not influence the immediate postoperative clinical evolution after ESS. PMID- 10719592 TI - Orbital edema resulting from Haller's cell pathology: 3 case reports and review of literature. AB - Sinuspathology must be considered when rapid onset of unilateral orbital edema is found in the absence of ophtalmologic signs. Urgent medical treatment is necessary in these patients when headache, fever, facial pain and vision problems are present. However, symptoms may be more subtle. Three female patients with unilateral orbital edema and facial pain are presented. Inflammation of an ipsilateral Haller's cell should be considered as one of the potential causes of unilateral orbital edema and it can be the only cause. To our knowledge this is the first report of Haller's cell disease resulting in an orbital complication. PMID- 10719593 TI - Sinonasal lymphomas. Case report. AB - In the field of Otorhinolaryngology sinonasal lymphomas are relatively uncommon and represent less than 1% of all head and neck malignancies. Nowadays, they are regarded as consisting of two distinct subgroups, characterised by phenotype, location, prognosis and treatment. Lymphomas of the B-Cell phenotype are the most frequent type found in the paranasal sinuses. They are less aggressive and carry a relatively better prognosis. T/NK-Cell lymphomas are mostly found in the nasal cavity. They are more aggressive and carry a relatively worse prognosis. We present a case of a 65-year-old patient, who complained with unilateral right sided nasal obstruction associated with a sensation of right aural fullness. CT scan demonstrated opacity of the posterior ethmoid and sphenoid sinuses on the right side, with evidence of erosion of the anterior wall of the sphenoid. Nasal endoscopy revealed a smooth purple mass, arising from the right superior meatus, which bled on contact, and which was subsequently shown to be, on histological assessment, a malignant high grade lymphoma, non-Hodgkin's B-cell phenotype. Following a discussion of the case we will present a review of these tumours, which have a poor overall prognosis, focusing on epidemiology, sites of origin, symptoms, investigation and management. PMID- 10719594 TI - [The difficulty of determining a prognosis in skull-brain injuries]. PMID- 10719595 TI - [Principles of catecholamine therapy. 1. Characterization of clinically relevant sympathomimetics]. AB - All involuntary innervated structures of the body are controlled by the sympathetic and parasympathetic nervous system. Adrenaline, noradrenaline and dopamine are endogenous catecholamines binding to adrenergic and dopaminergic receptors, respectively, to mediate their clinical effects. Adrenoceptors are classified as alpha 1, alpha 2, beta 1 and beta 2 subtypes which were even further subcharacterized the recent years. Adrenoceptors are membrane proteins interacting with the agonist and, thus, inducing G-protein mediated intracellular effects. Adrenaline induces an extensive increase of heart rate and stroke volume mediated by beta-adrenoceptors and significantly enhances peripheral vascular resistance by alpha-adrenoceptor stimulation, when administered beyond 0.1 microgram/kg.min. In contrast, the clinical effects of noradrenaline are predominantly characterized by alpha-adrenoceptor stimulation resulting in a less pronounced increase of heart rate. Dopamine, less potent on adrenoceptors, shows additional effects on renal as well as on splanchnic circulation mediated by dopaminergic receptors. Dobutamine, primarily acting on beta-adrenoceptors, results in positive inotropic effects without an increase in vascular resistance. Dopexamine, a synthetic catecholamine, induces vasodilation via beta 2 adrenoceptor stimulation and potentially increases splanchnic blood flow by additional effects on dopaminergic receptors. Isoproterenol, the classical beta adrenoceptor agonist, mediates positive inotropic effects and causes a major increase in heart rate and a significant decrease of systemic vascular resistance. Independent on adrenoceptors, phosphodiesterase-III-inhibitors exert positive inotropic and vasodilating activity by an increase in intracellular cAMP concentration induced by inhibition of cAMP hydrolysis. PMID- 10719596 TI - [In Process Citation] AB - A main basis of this habilitation thesis is the development of a portable, mains free measurement device for determining breath ethanol concentrations (BrEC) both during spontaneous breathing and in mechanically ventilated patinets undergoing inhalation anaesthesia, for detecting and quantifying irrigation fluid absorbed during endourological surgical interventions. Here a respired gas has to be measured which compared to other usually measured gases 1) is present in substatially smaller concentrations, 2) is subject to significant influences by temperature and humidity in the expired air, and 3) can only be measured discontinuously in the alveolar air due to technical limitations of the sensor. The basis for investigating accuracy of measurement was not just evaluation of the device using different lung models, but also a comparison with the target parameter "absorbed irrigation fluid" using another further-developed reference procedure, i.e. direct measurement of sorbitol and mannitol concentrations in serum. This has the added advantage that even when ethanol monitoring is not available, another laboratory procedure is indeed available for directly evaluating the absorption of irrigation fluid. In the clinical aspects of this thesis, ethanol monitoring helped show that during transurethral resection of the prostate (TURP) in spontaneously breathing patients undergoing regional anaesthesia, irrigation fluid was intravascularly absorbed not more often, but more rapidly and in larger quantities than it was in mechanically ventilated patients undergoing inhalation anaesthesia. A possible reason for this was the significantly reduced central venous pressure observed in the group of patients undergoing regional anaesthesia. The diagnosis of a delayed extravascular absorption of irrigation fluid during percutaneous nephrolithotripsy (PNL), made with the assistance of the ethanol monitoring, was associated with a significantly prolonged period of hospitalization in the clinic and an increase in opioid requirements. This finding can be explained pathophysiologically by an increased trauma to the kidney with injury to bordering organ structures. As a result of close co-operation with several departments of the Medical University of Lubeck, we now have at our disposal an excellent BrEC-measuring device for detecting and quantifying absorbed irrigation fluid during urosurgical procedures. The results of the clinical studies presented here underscore the value of the newly-developed AlcoMed 3011. PMID- 10719597 TI - [Median nerve evoked potentials and explicit memory function during recovery from propofol-sufentanil anesthesia]. AB - OBJECTIVE: In a prospective study rnidlatency somatosensory evoked Potentials (SEP) were investigated in relation to explicit memory function during recovery from propofol/sufentanil anaesthesia. METHODS: Anaesthesia was maintained in 20 patients with 8 mg kg-1 h-1 propofol and supplemented with sufentanil. SEPs (N20, P25, N35, P45 N50) elicited by median nerve stimulation were recorded at the day before surgery (AWAKE) and during recovery from anaesthesia, when patients were able to identify a shown object (RECOVERY). The day after surgery patients were interviewed about their memory of the recovery period. STATISTICS: Hotellings' T2. RESULTS: One day after anaesthesia 9 patients could remember events during the recovery period. At RECOVERY SEP-latencies P45 and N50 were significantly shorter in the patients with recall for the wakeup-phase (p < 0.05). CONCLUSION: SEP latencies P45 and N50 indicated impaired explicit memory function during recovery from propofol/sufentanil anaesthesia. PMID- 10719598 TI - [Outcome factors in severe skull-brain trauma. A retrospective analysis of 228 (161) patients]. AB - OBJECTIVE: To study outcome from severe head injury (SHI: GCS < or = 8) and to investigate impact of prehospital factors and clinical intensive care parameters on outcome. To compare with former study results (1980-88) of our clinical setting. METHODS: Retrospectively, the history of 228 patients with SHI treated between 1988 and 1995 was looked into. The outcome was measured with the Glasgow Outcome Scale (GOS) post intensive care (median 9, min-max 2-77 days) and 6 months after trauma by a questionnaire. The GOS was related to age, Glasgow Coma Scale (GCS on the scene), prehospital hypotension and hypoxia (HH), intracranial pressure (ICP), cerebral perfusion pressure (CPP), intensive therapy including Tromethamine and/or Thiopentone. The rate of infections was determined. RESULTS: Increasing age influences outcome negatively. Prehospital GCS and HH were significantly correlated with outcome. GOS of 30 patients with HH: GOS 1: 53%, GOS 2 + 3: 27%, GOS 4 + 5: 20%. GOS of 40 patients without HH: GOS 1: 25%, GOS 2 + 3: 10%, GOS 4 + 5: 65%. During intensive care the level of CPP (not ICP) as well as tromethamine and/or thiopentone treatment for control of elevated ICP were significantly correlated with outcome. Mortality rate in 32 patients with CPP < 50 mmHq was 69%, in 29 patients with CPP > 50 mmHg only 20%. Patients treated additionally with Tromethamine and Thiopentone because of uncontrollable intracranial hypertension showed a significantly worse outcome: GOS 1: 66%, GOS 2 + 3: 6%, GOS 4 + 5: 28%, compared to those who needed neither Tromethamine nor Thiopentone: GOS 1: 27%, GOS 2 + 3: 18%, GOS 4 + 5: 55%. Thiopentone treatment was not associated with an increased rate of pulmonary and other infections. In comparison to our former outcome study, covering the years 1980-88, we have not seen any improvements in outcome, despite modifications in intensive care protocols. CONCLUSIONS: Prehospital hypotension and hypoxia have a significant negative impact on outcome by causing secondary brain damage. Despite various modifications in intensive care therapy an unchanged portion of secondary brain damage will not prove treatable. Therefore, prevention or early aggressive treatment of hypotension and hypoxia is the most promising way of improving outcome after severe head injury at the moment. PMID- 10719599 TI - [Pharmacologic aspects of endotracheal intubation]. PMID- 10719600 TI - Pharmacokinetic aspects of the onset of action of neuromuscular blocking agents. PMID- 10719601 TI - [Effects of muscle relaxants on various muscles: significance for intubation]. PMID- 10719602 TI - [The effect of anesthesia and muscle relaxation on intubation conditions]. PMID- 10719603 TI - [The effect of hypnotics on intubation conditions and onset of action of muscle relaxants]. PMID- 10719604 TI - [Intubation without muscle relaxants: options and limitations]. PMID- 10719605 TI - [Rapid sequence induction with succinylcholine: an option to reduce side effects]. PMID- 10719606 TI - [Rapid sequence intubation with non-depolarizing muscle relaxants: priming, timing, megadose]. PMID- 10719607 TI - [Rapacuronium: a new, non-depolarizing muscle relaxant for rapid intubation?]. PMID- 10719608 TI - [Heparin-induced thrombocytopenia and hemofiltration: regional anticoagulation with sodium citrate]. AB - The symptoms, diagnosis, and therapy for a septic patient with acute renal failure on continuous hemofiltration and heparin-anticoagulation who developed heparin-induced thrombocytopenia type II are described. Different alternatives for anticoagulation of extracorporal circulation are discussed. The principles of anticoagulation with sodium citrate are presented. It is necessary to closely monitor and, if need be, correct the serum calcium levels and acid base metabolism. We show that regional anticoagulation with sodium citrate is an effective and safe treatment. PMID- 10719610 TI - [HP Anesthesiology Village. A commercial subject server]. PMID- 10719609 TI - [The effect of anesthesia on the incidence of postoperative venous thromboembolism]. PMID- 10719611 TI - Pharmacokinetics of tramadol and bioavailability of enteral tramadol formulations. 4th communication: drops (without ethanol). AB - In a balanced two-period cross-over study with 12 (6 male, 6 female) healthy young subjects the pharmacokinetics and absolute bioavailability of tramadol (tramadol hydrochloride, CAS 36282-47-0) after oral administration of Tramal drops (without ethanol) were to be determined in comparison with a 30-min i.v. infusion. Each fasting volunteer received the two single doses of 50 mg tramadol HCl each in the morning; the time interval between the administrations was one week. Serum and urine concentrations of tramadol were analysed by gas chromatography. The pharmacokinetic evaluation was carried out model-dependently after previous selection of the optimal model by means of the Akaike information criterion; solely the extent of bioavailability was calculated model independently. The study population results were presented descriptively as geometric means with standard deviation [xg (SDg)] or as medians with range [x (min, max)]. Model-dependently and model-independently calculated areas under the serum concentration curves (AUC and AUC, resp.) differed only minimally. The extent of the absolute bioavailability (F) of tramadol in the drops, based on AUC data, was 70.6 (1.13)% with a 90% confidence interval of 65.9-75.6% (ANOVAlog). The p.o. serum concentration peaks were reached after tmax = 1.2 (0.74, 1.5) h and amounted to Cmax = 136 (1.33) ng/ml, the half-life of absorption was t1/2,ka = 0.23 (1.89) h and the lag time t0 = 0.23 (0.20, 0.49) h. The dose-normalised p.o. and i.v. results for all pharmacokinetic parameters agreed well with those of a previous study with ethanol-containing drops. In summary, it may be concluded that the active ingredient is rapidly absorbed after oral administration of the drops without ethanol. Rate and extent of the absolute bioavailability of tramadol in drops without ethanol were about the same as after administration of drops with ethanol. The results of this study gave no indication of a therapeutically relevant gender difference in the pharmacokinetics of tramadol. PMID- 10719612 TI - Efficacy of orally administered extract of red vine leaf AS 195 (folia vitis viniferae) in chronic venous insufficiency (stages I-II). A randomized, double blind, placebo-controlled trial. AB - Red vine leaf extract (RVLE) is a herbal medicine containing several flavonoids, with quercetin-3-O-beta-glucuronide and isoquercitrin (quercetin-3-O-beta glycoside) as the main components. OBJECTIVE: To assess the efficacy and safety of once-daily doses of 360 and 720 mg RVLE (pharmaceutical extract code AS 195; Antistax Venenkapseln) compared to placebo in patients with stage I and incipient stage II chronic venous insufficiency (CVI). DESIGN: A 12-week, randomized, double-blind, placebo-controlled, parallel-group, multi-center study. PATIENTS: Male and female outpatients aged 25 to 75 years with stage I to stage II CVI (i.e. without extensive trophic changes), not having any other significant medical conditions and not treated with compression stockings, diuretics or other drugs affecting fluid balance. INTERVENTION: Patients were randomly assigned to a double-blind treatment with placebo, 360 mg AS 195 or 720 mg AS 195 once daily for 12 weeks, preceded and followed by a single-blind 2-week placebo treatment for baseline run-in and end-of-trial washout, respectively. Study criteria were evaluated at baseline, after 6 and 12 weeks of treatment and 2 weeks after discontinuation of treatment. RESULTS: Of the 260 patients enrolled and randomized, 219 completed the study in accordance with the protocol. In the intention-to-treat analysis (N = 257), the mean (+/- SD) lower leg volume (measured by water displacement plethysmography) of the patients treated with placebo (N = 87) increased by 15.2 +/- 90.1 g (displaced water mass) and by 33.7 +/- 96.1 g after 6 and 12 weeks compared to baseline. In contrast, for patients treated with AS 195, lower leg volume decreased, and after 12 weeks of treatment, the difference in mean lower leg volume between the active treatment groups and the placebo group was -75.9 g (95% CI: -106.1 to -45.8 g) and -99.9 g (95% CI: 130.3 to -69.6 g) for the group treated with 360-mg AS 195 (N = 86) and 720-mg AS 195 (N = 84), respectively. The changes in calf circumference showed a similar pattern: in patients treated with AS 195, both the higher dose (720 mg) and, albeit to a lesser extent, the lower dose (360 mg) resulted in a clear reduction in circumference over time, whereas, circumference remained largely unchanged in patients treated with the placebo (95% CI of the estimated treatment effects vs. placebo after 12 weeks: -1.40 to -0.56 cm and -1.73 to -0.88 cm for 360 and 720 mg AS 195, respectively). These differences were statistically significant (p < 0.001). The reductions in ankle circumference were qualitatively similar but quantitatively less marked. Subjectively, there was an improvement in key CVI symptoms (VAS) at 6 weeks with all treatments, but a further improvement at week 12 was seen only in the active treatment groups; at 12 weeks, the changes compared to baseline were significantly greater (p < 0.001) in both active treatment groups than in the placebo group. The treatments were well tolerated; Adverse events were rare and usually mild. Two adverse events (AEs) during treatment with the placebo led to hospitalization and were hence labeled as 'serious'. Three further patients were withdrawn because of AEs which occurred during treatment with the placebo. CONCLUSION: Once-daily doses of 360 and 720 mg AS 195 were confirmed to be safe and effective in the treatment of mild CVI, reducing significantly lower leg edema and circumference whilst improving key CVI related symptoms to a clinically relevant extent. The edema reduction is at least equivalent to that reported for compression stockings and/or other edema-reducing agents. The higher dose was as well tolerated as the lower dose but resulted in a slightly greater and more sustained improvement. PMID- 10719613 TI - Effects of NO-1886, a lipoprotein lipase promoting agent, on homozygous and heterozygous Watanabe heritable hyperlipidaemic rabbits. AB - The novel compound NO-1886 ([4-(4-bromo-2-cyano-phenylcarbamoyl)-benzyl] phosphonic acid diethyl ester, CAS 133208-93-2) is a lipoprotein lipase (LPL) activator, and long term administration of NO-1886 protects against the development of experimental atherosclerosis in rats and rabbits. In the present experiments, the effects of this compound were examined in Watanabe heritable hyperlipidaemic (WHHL) rabbits, an animal model for familial hypercholesterolemia lacking low density lipoprotein (LDL) receptors. NO-1886 increased postheparin plasma LPL activity, resulting in a reduction of plasma triglycerides with concomitant elevation of HDL cholesterol in heterozygous WHHL rabbits. However, the compound did not cause any changes in plasma lipids and postheparin plasma LPL activity in homozygous WHHL rabbits. The different responses suggest that the effects of NO-1886 may be either mediated by LDL receptors, or that persistent exposure to extreme hypercholesterolemia might affect the cellular response to this particular compound in homozygous WHHL rabbits. PMID- 10719614 TI - Effects of the neoflavonoid coutareagenin, one of the antidiabetic active substances of Hintonia latiflora, on streptozotocin-induced diabetes mellitus in rats. AB - In the present study, diabetes mellitus was induced with streptozotocin in Wistar rats from the inbred strain F > 28, and a blood sugar lowering effect due to the oral or intragastric administration of a copalchi native extract (Hintoniae latiflorae cortex) or intragastric administration of the pure substance, coutareagenin (neoflavonoid) (5-hydroxy-7-methoxy-4-(3,4-dihydroxyphenyl)-2H benzo-1-pyran-2-on), was demonstrated with statistically significant results. This study enlarged and confirmed the results of earlier studies by other authors with regard to the antidiabetic effect of the copalchi bark extract in various animal species with hyperglycemia induced by different methods. By the use of the synthetically produced neoflavonoid, coutareagenin, the present series of tests provided evidence that coutareagenin, one of the active substances contained in Hintonia latiflora bark (Copalchi bark), produces a reduction of diabetic elevated blood sugar levels. The native extract as a whole, however, has to be regarded as the active substance (active substance complex) of Hintonia latiflora (Copalchi) bark. PMID- 10719615 TI - Evaluation of ursodeoxycholic acid bioavailability from immediate- and sustained release preparations using gas chromatography-mass spectrometry and high performance liquid chromatography. AB - An improved procedure is presented for the determination of ursodeoxycholic acid (CAS 128-13-2, UDCA) in human plasma and bile after oral administration of UDCA containing dosage forms. The plasma samples after solid-phase extraction with silica-based C18- and strong anion exchange cartridges were assayed by gas chromatography-mass spectrometry (GC-MS) using selected-ion monitoring. The hexafluoroisopropyl trifluoroacetate ester derivative of UDCA was selected for GC analysis since it is easily and rapidly prepared by a one-step reaction. Biliary UDCA levels were determined by a rapid and simple high-performance liquid chromatographic (HPLC) method with on-line sample purification. This analytical protocol was used to investigate the pharmacokinetic of a new sustained-release capsule of UDCA in comparison with a reference immediate-release preparation after single oral administration. Statistical evaluation of the area under the plasma concentration-time curves indicated that two formulations are equivalent with regard to the amount of drug absorbed. However, pharmacokinetic data showed that with the sustained-release preparation a significantly delayed mean peak plasma level was reached compared with the reference preparation. Moreover, the immediate- and extended-release capsules were found to achieve a comparable degree of biliary enrichment with UDCA. PMID- 10719616 TI - Antioxidant properties of essential oils. Possible explanations for their anti inflammatory effects. AB - Pathogenesis and symptoms of inflammatory processes are accompanied and/or initiated by the production of reactive oxygen species (ROS). The effects of essential oils on these processes have been studied with the aid of biochemical model reactions simulating these pathological events. It can be shown that Myrtol Standardized and Eucalyptus oil ameliorate inflammatory processes by interacting with aggressive oxygen radicals of the OH.-type and interfere with leukocyte activation. These activities partially allow attenuation of oxidative attack and damage introduced by infections or environmental impacts. PMID- 10719618 TI - Effects of chondroitin sulfate-C on articular cartilage destruction in murine collagen-induced arthritis. AB - The effects of chondroitin sulfate-C (CAS 25322-46-7, Chondroitin ZS Tab) on type II collagen (CII)-induced arthritis (CIA) in mice were evaluated. DBA/1J mice were immunized with bovine CII emulsified in Freund's complete adjuvant, followed by a booster injection 21 days later. Chondroitin sulfate-C at doses of 100, 300 and 1000 mg/kg was administered orally once daily beginning 14 days before initial immunization. An arthritis index and hind paw edema were examined from day 0 to day 49, when the mice were killed by ether anesthesia for histopathological examination. The delayed-type hypersensitivity (DTH) reaction, serum anti-CII antibody titer, and histopathologic characteristics of both synovitis and destruction of articular cartilage were analyzed. Both the arthritis index and the serum anti-CII antibody titer were reduced by treatment with chondroitin sulfate-C in a dose-dependent manner. Chondroitin sulfate-C (1000 mg/kg) significantly inhibited hind paw edema, synovitis and destruction of the articular cartilage, but not DTH reaction. PMID- 10719617 TI - Synthesis and pharmacological screening of novel non-acidic gastroprotective antipyretic anti-inflammatory agents with anti-platelet properties. 5 Alkyl/cycloalkylamino substituted 2-amino-5H-[1]benzopyrano[4,3-d] pyrimidines. AB - The preparation and screening of antipyretic, anti-inflammatory, analgesic, gastroprotective and antiplatelet activities of original non-acidic aminobenzo pyranopyrimidine derivatives are described. Major and dose-dependent analgesic and antipyretic properties were detected in all the compounds after oral administration (6.25-100 mg kg-1) in the mouse writhing test and in the E. coli lipopolysaccharide-induced fever in the rat, respectively. Unlike the reference drug indometacin (ED50 ulcer = 9.1 mg kg-1), no gastrolesivity was displayed by all the new compounds up to 150 mg kg-1 so that therapeutical indices equal or higher than that of indometacin were calculated. Furthermore, at 100 mg kg-1 they were able to prevent ethanol-induced gastric mucosal injury in rats. At a 1 mmol/l concentration the compounds under study were as effective as acetylsalicylic acid in inhibiting in vitro platelet aggregation induced by adenosine diphosphate. PMID- 10719619 TI - Synthesis of 3-substituted phenacyl-5-[2-phenyl-4H-4-oxo-1-benzopyran-6- yl)methylenyl]-thiazolidine-2,4-diones and evaluation of their antimicrobial activity. AB - Synthesis and physico-chemical properties of two 3-substituted phenacyl thiazolidine-2,4-diones (2c-d) and four 3-substituted phenacyl-5-[2-phenyl-4H-4 oxo-1-benzopyran-6-yl)methyl-enyl]-thiazolidin e- 2,4-diones (4a-d) are described. These products were synthesized by Knoevenagel reaction from flavone-6 carboxaldehyde (3) and 3-(substituted phenacyl)- thiazolidine-2,4-diones (2a-d). Antibacterial and antifungal activities were investigated for these compounds. PMID- 10719620 TI - Pharmacokinetics, metabolism and excretion of megazol in a Trypanosoma brucei gambiense primate model of human African trypanosomiasis. Preliminary study. AB - The pharmacokinetics of megazol (2-amino-5-(1-methyl-5-nitro-2-imidazolyl)-1,3,4 thiadiazol, CAS 19622-55-0) was investigated after a 100 mg/kg oral administration to six primates infected with Trypanosoma brucei gambiense. The plasma levels of megazol were between 0.2 microgram/ml and 46 micrograms/ml 24 h after dosing in all animals. In animals with prolonged infection, megazol absorption was accelerated (Tmax was 4 h compared with 8 h, for day 53 and day 39 post inoculation) but the amount absorbed was not modified. The megazol concentrations in the cerebrospinal fluid represented between 5.5% and 10.6% of the plasma levels at the same times. Unchanged megazol was eliminated predominantly via the kidneys: 46-96% of the ingested dose was recovered in the urine, compared with 0-5% in the faeces. Furthermore, this urinary elimination of megazol was altered in animals with prolonged infections. In the urine, 4 unknown metabolites were observed, unchanged megazol was characterized by LC-MS/MS. This study indicates that megazol crosses the blood-brain barrier after oral administration. Prolonged infections affect the absorption of megazol and its urinary elimination. PMID- 10719621 TI - Inhibition of cryptosporidium parvum in vitro by 9-(alkylthio)acridine derivatives. AB - The synthesis of several acridinic thioethers is described. Compounds prepared were tested in vitro as potential drugs against the opportunistic infection known as cryptosporidiosis. With a view to predict activity, the quantitative structure activity relationships were investigated. Correlations between experimental data and either log P or pKa are discussed. PMID- 10719622 TI - 4-(3-coumarinyl)-4-thiazolin-2-one benzylidenehydrazones with antituberculosis activity. AB - In this study a new series of 4-(3-coumarinyl)-4-thiazoline-2-one benzylidenehydrazones (3a-v) were synthesized by condensation of 3-(omega bromoacetyl)coumarins (1a and 1b) with 1-substituted benzylidene-4-substituted thiosemicarbazides (2a-1). Structures of the title compounds were elucidated by elemental analyses and spectrometric data (UV, IR, 1H-NMR and EIMS). These new compounds and some previously reported compounds (1a-d) were evaluated for antituberculosis activity against Mycobacterium tuberculosis H37Rv. The compounds exhibited varying degrees of inhibition in the in vitro primary screening that was conducted at 12 micrograms/ml against M. tuberculosis H37Rv in BACTEC 12B medium using the BACTEC 460 radiometric system. 1b, 3b, 3e, 3h, 3o and 3p demonstrating activity in the primary screen were re-tested at lower concentrations against M. tuberculosis H37Rv to determine the actual minimum inhibitory concentration (MIC) in CABTEC 460 and Alamar Blue assay (MABA). The most active compound was found to be 1b. The structure-activity relationships of the derivatives were investigated. PMID- 10719623 TI - Correlation between infection intensity, serum immunoglobulin profile, cellular immunity and the efficacy of treatment with praziquantel in murine schistosomiasis mansoni. AB - This study was conducted to evaluate the efficacy of praziquantel (CAS 55268-74 1, EMBAY 8440, Biltricide) in different grades of Schistosoma mansoni infection. Moreover, the relationship between the post treatment worm burden, hepatic granuloma volume, and serum immunoglobin profile was also investigated. Four groups of Swiss albino mice infected with Schistosoma mansoni cercariae were used: Highly infected untreated control mice (infected with 120 Schistosoma mansoni cercariae) and their corresponding praziquantel treated group. Lightly infected untreated control mice (infected with 60 Schistosoma mansoni cercariae) and their corresponding praziquantel treated group. Praziquantel was given seven weeks post infection in a dose of 500 mg/kg body weight for two consecutive days. Animals were sacrificed two weeks post treatment. Praziquantel achieved better cure rates in mice with heavy infection than in less intensely infected animals. The drug reduced the hepatic granuloma in animals with light intensity infection. This reduction was more accentuated in highly infected animals. The serum immunoglobulin profile (immunoglobulin G and immunoglobulin M) showed a higher level in highly infected treated mice (1.2 +/- 0.6 optical density unit and 1.1 +/- 0.5 optical density unit, respectively) and was reduced in animals with low intensity infection (1.18 +/- 0.6 optical density unit and 0.7 +/- 0.6 optical density unit, respectively). This study may be of value in tropical regions, where schistosomiasis with low worm burden is a common occurrence. PMID- 10719624 TI - Effects of a spleen peptide preparation as supportive therapy in inoperable head and neck cancer patients. AB - Patients with inoperable head and neck cancer were treated with a spleen peptide preparation (Polyerga) in a phase III randomized, placebo-controlled double-blind study during chemotherapy (cisplatin/carboplatin, 5-fluorouracil) to investigate further the efficacy of this peptide preparation as supportive treatment under chemotherapy. Immunological changes as well as quality of life aspects were examined. Forty patients were included in this study. The peptide preparation had a significant stabilizing effect on the peripheral blood lymphocyte status during chemotherapy cycles (Student t-test, p = 0.05) and tended to stabilize the shift of granulocyte count (Student t-test, p = 0.18). In addition, the group receiving the verum showed a remarkable stabilization of body weight (Mann-Whitney U-test, p = 0.17) during chemotherapy treatment and the generally observed increase of fatigue-inertia during the chemotherapy cycles was significantly reduced (Student t-test, p = 0.01). PMID- 10719625 TI - Transgenic potato plants expressing osmotin gene inhibits fungal development in inoculated leaves. AB - Osmotin, a 24-KD protein isolated from Nicotiana tabacum 'Xanthi nc' was very toxic to Phytophthora infestans in vitro assay. Transgenic potato plants expressing wild osmotin protein delayed the emergence of symptoms of disease. Two osmotin mutants have been produced by PCR mutagenesis. One mutant, delta C18 osmotin, had a nonsense mutation near the C-terminus that deleted the carboxyl terminal prepropeptide (CTPP) of 18 amino acids. Another mutant, osmotin-P, had amber mutation at 96th amino acid. Both mutant genes driven by CaMV d35S promoter were introduced in potato by Agrobacterium mediated genetic transformation. The expression and osmotin protein accumulation in transgenic potato were detected by Southern, Northern, and Dot-blot ELISA assay. Disease resistance test using P.infestens complex race showed that expression of both osmotin mutants in transgenic potato reduced the lesion growth rate in inoculated leaves. The results showed that accumulation of osmotin protein in intercellular spaces (delta C18osmotin) inhibited the development of fungal disease in inoculated leaves. Based on the results we suggest that antifungal activity was possibly located at N end of osmotin protein (osmotin-P). PMID- 10719626 TI - Replacement vectors for localized gene cloning in the specified region of Streptomyces lividans 66 and model method for the screening of desired recombinants via counter-selection. AB - Two DNA fragments (3.5 kb and 3.8 kb in size) flanking both ends of the phi HAU3R gene in the genome of Streptomyces lividans 66 were determined and cloned in their natural relative orientations in pIJ653, a cosmid vector derived from the multi-copy Streptomyces plasmid pIJ101, resulting in pHZ806. After insertion of spectinomycin/streptomycin (spc/str) resistance gene into the pIJ101 replication region in pHZ806 and insertion of a hygromycin (hyg) resistance gene between 3.5 kb and 3.8 kb DNA fragments, a new vector with a non-functional Streptomyces replicon, pHZ808, was obtained. In principle, any DNA fragment cloned between 3.5 kb and 3.8 kb fragments of this vector can be stably integrated between the two corresponding regions after introduction into the wild-type S.lividans strains, with synchronous replacement of the DNA between the two regions of the chromosome within which phi HAU3R gene is located. The resultant recombinant strains will thus become phi HAU3-sensitive (phi HAU3S). This phenotype could serve as a good indication that the desired gene replacement has occurred. This principle was demonstrated to be successful using pHZ808 as vector. phi HAU3-resistance gene (phi HAU3R) from the genome of S.lividans 66 was substituted by the hygromycin resistance gene (hyg) from pHZ808. An additional advantage of using pHZ808 as the vector to clone foreign genes is that hyg could be served as a reportor gene to imply that foreign DNA fragment has been co-integrated with hyg. The recombinants will have both hygR and phi HAU3S. PMID- 10719627 TI - Purification and characterization of a novel fibrinolytic enzyme from Streptomyces spp. AB - A novel protease with fibrinolytic activity, designated as SW-1, was isolated and purified from the fermentation broth of Streptomyces sp. strain Y405, a soil isolate. The purification procedure involved ammonium sulphate fractionation, decolorization on 290 resin, gel filtration on Sephadex G75, anion-exchange chromatography on DEAE Sephadex A25, and affinity chromatography on Lysine Sepharose 4B. About 4.2 mg purified enzyme was obtained from a liter of fermentation broth and the recovery yield was 12.0%. The purified enzyme showed the specific activity of 2952.3 urokinase units per milligram, which was increased by 230.6 fold over the fermentation broth. The purity determined by HPLC was 83.5%. SW-1 is a single chain polypeptide with a predicted molecular weight of 30 kDa in SDS-PAGE and an isoelectric point of 8.5. The N-terminal sequence of SW-1 is R/N/F-P/D-G-M-T-M-T-A-I-A-N-Q-N-T-Q-I-N. There may be nonhomogeneity in the first and second amino acid residue of its N-terminal sequence. The analysis of amino acid composition showed that SW-1 consisted of 262 amino acids. The fibrinolytic activity of SW-1 was entirely inhibited by 10 mmol/L PMSF, 1 mmol/L EDTA and 1 mol/L lysine, respectively, suggesting that SW-1 is a serine protease and metalloprotease, and that the lysine binding site might play a role in the activity. The fibrinolytic activity of SW-1 is stable between 4-37 degrees C and pH 4.0-9.0, and the optimum pH is 8.0. On plasminogen-free fibrin plates, SW-1 showed the same fibrinolytic activity as the mixture of SW-1 with plasminogen, indicating that SW-1 is a fibrinolytic enzyme which affects fibrin directly, but not a plasminogen activator which affects fibrin by activating plasminogen. PMID- 10719628 TI - Cloning and expression of truncated PTHrP receptor in Escherichia coli. AB - To evaluate the effects of recombinant soluble parathyroid hormone related protein receptor(sPTHrPR), the RNA prepared from renal cell carcinoma was reverse transcribed and amplified by a polymerase chain reaction(PCR). A 543 base pair fragment corresponding to extracellular domain of PTHrPR was obtained and cloned. Sequencing was performed by the dideoxy chain-termination method. Comparison of the nucleic acid sequence with the published data revealed that only one nucleotide had changed. A high level expression vector referred to as pET 3a/sPTHrPR was constructed. The binding specificity of the expression products was proved by indirect ELISA. Our preliminary experiment also showed that the expression products can block the biological activity of PTHrP. PMID- 10719629 TI - Construction of La-tPA vector and expression in the mammary gland of transgenic mice. AB - Expression vectors of long acting human tissue plasminogen activator (La-tPa) in the mammary gland was constructed using the promoters of sheep beta-lactoglobulin gene (BLG, 5 kb) containing the first and the second introns obtained by PCR. Transgenic mice were established by microinjection. 540 fertilized eggs were injected and 6 transgenic mice were screened out from the offspring by PCR and Southern Blot analysis. The foreign gene integrating rate was 32%. The expression of La-tPA in mammary glands of transgenic mice was confirmed by milk assay and Northern blot analysis. Expression level of La-tPA reached 6 ug/ml in milk of transgenic mice. PMID- 10719630 TI - Construction of a stable bioengineered strain of biotechmycin. AB - A stable bioengineered strain of Biotechmycin (Streptomyces spiramyceticus WSJ-1) was constructed by integrating the 4"-isovaleryltransferase gene (ist) through homologous recombination into the chromosome of spiramycin-producing strain S.spiramyceticus F21. In this construction, a Streptomyces/E.coli shuttle plasmid pKC1139 (AmR) was used as the vector and tsr gene was inserted as the marker for selection of homologous recombination. This constructed strain, S.spiramyceticus WSJ-1, was genetically very stable in production titer and in the production of biotechmycin as well as in carrying tsr selective marker when grown without pressure. The fermentation of S.spiramyceticus WSJ-1 was also improved compared with the original strain harboring unintegrated plasmid. Southern hybridization confirmed the integrated status of the ist gene in the host genome. PMID- 10719631 TI - Characterization of the ecdysteroid UDP-glucosyltransferase gene of Spodoptera litura multinucleocapsid nucleopolyhedrovirus. AB - The ecdysteroid UDP-glucosyltransferase (EGT) gene of Spodoptera litura multinucleocapsid nucleopolyhedrovirus (SpltMNPV), Guangzhouisolate was sequenced. The open reading frame of the gene is 1530 nucleotides long, encoding a putative protein of 509 amino acids with Mr of 58,500. The 5' noncoding region has a TATA box. A polyadenylation signal, AATAAA, is contained downstream of the translation stop codon. A putative signal peptide is present at the N-terminus of the protein. Homology comparison with other baculovirus egt genes showed that these genes are highly conserved. The SpltMNPV egt gene is closely related to that of Spodoptera littoralis nucleopolyhedrovirus (SliNPV) with the identities of 84% in nucleotide and 91% in amino acid, respectively. In this study, an alignment of amino acid sequences has been made and a tentative phylogenetic tree of twelve baculovirus egts is presented. PMID- 10719632 TI - Studies on production of pullulan by the feed batch fermentation. AB - The feed-batch fermentation of pullulan was investigated based on the conditions of batch fermentation. The optimal conditions of pullulan fermentation were determined by the investigation on the effect of feed mode, initial feed time, intermittent time, and composition of the feed solution on parameters such as productivity, conversion yield, biomass, dissolved oxygen (DO), pH of the fermentation broth, viscosity of the fermentation broth and molecular weight of the product. Above 70% of conversion yield and 100 kD of average molecular weight of the product was obtained at the optimal conditions. PMID- 10719633 TI - Immobilization of sorghum leaf oxalate oxidase onto alkylamine and arylamine glass. AB - An oxalate oxidase (EC 1.2.3.4) purified from grain sorghum leaves was immobilized onto alkylamine and arylamine glass beads through glutaraldehyde coupling and diazotization with a conjugation yield of 10.8 mg/g support and 9.2 mg/g support, respectively. The enzyme retained 67.5% and 34.1% of its initial specific activity after immobilization onto alkylamine and arylamine glass, respectively. The enzyme exhibited an increase in optimum pH, temperature for maximum activity, energy of activation (Ea) and time for linearity but decrease in thermal stability at 60 degrees C after immobilization on both types of beads. The K(m) value for oxalate was increased by 9- to 10-fold but Vmax remained unaltered after immobilization. Both alkyl and arylamine glass bound enzyme was unaffected by physiological concentrations of Cl- and NO3-. The analytic importance of this work is demonstrated. PMID- 10719634 TI - Detailed differential coefficients of anomalous dispersion terms to derivatives in least-squares refinement for X-ray crystallography AB - In X-ray crystallographic analysis, correct trial structures are refined by least squares method. The detailed differential coefficients for the parameters of the calculated structure factor Fc(h) can not be found in any papers, textbooks, nor user's manual of several crystallographic computer systems. We describe, firstly, the basic expression of the structure factors, Fo(h) and Fc(h), anomalous dispersion terms of the structure factor, and then least-squares refining of magnitude of Fo(h) and Fo(h)2 for use in teaching, education and computer programming. Secondly we derive the differential terms of the structure factors, solve the normal equation by diagonal, block and full-matrix approximations, and apply these to the revised computer program LSBF in DS*SYSTEM. Several practical tests are performed for these differentiations. PMID- 10719635 TI - Pattern formation in a Turing's type model with minimal reactional complexity AB - Turing's original reaction network is systematically studied, particularly in what concerns: (a) Its ability to produce patterns in a predictable way. (b) The feasibility of its concentration-independent sink term. Despite the widely accepted view that Turing's original model presents some inherent inability to produce regular structures, the pattern formation properties of this model are found to obey the predictions of the corresponding Linear Stability Analysis in the one-dimension and in 'small' two-dimensional systems. An 'Enzymatic' variation of the original Turing's Model is introduced, where the unrealistic sink term is substituted by an enzymatic degradation. It seems that reaction networks of this type can inspire a promising search for chemical or biochemical experimental systems with pattern formation properties, even in the absence of high non-linearities. It is pointed out that temporal oscillations, impossible for the original Turing's Model, are stable and persistent in its Enzymatic variation. PMID- 10719636 TI - Use of artificial neural networks to predict the gas chromatographic retention index data of alkylbenzenes on carbowax-20M AB - Quantitative structure-activity relationships (QSARs) quantify the connection between the structure and properties of molecules and allow the prediction of properties from structural parameters. Models of relationships between structure and retention index of alkylbenzenes were constructed by means of a multilayer neural network using extended delta-bar-delta (EDBD) algorithms. The 165 group data belong to 129 alkylbenzenes at different temperatures on carbowax-20M. We proposed a new method to describe the structure of the alkylbenzene with a simple set of six numeric code depending on its molecular formula. A set of six numbers and the temperature were used as input parameters to predict the retention indices. The performance of different order of structural coding was investigated. The networks' architecture and the learning times were optimized. The optimum ANNs could give excellent prediction results. In addition, the multiple linear regression (MLR) and nonlinear multivariate regression were applied. We have shown in our studies that, based on the structural numeric codes, ANNs give more accurate predictions of retention index data of alkykbenzenes than regression analysis. PMID- 10719637 TI - A comparative QSAR study of benzamidines complement-inhibitory activity and benzene derivatives acute toxicity. AB - A novel QSAR study of benzamidines complement-inhibitory activity and benzene derivatives acute toxicity is reported and a new efficient method for selecting descriptors is used. Complement-inhibitory activity QSAR models of benzamidines contain from one to five descriptors. The best, according to fitted and cross validated statistical parameters, is shown to be the five-descriptor model. Models with a higher number of indices did not improve over the five-descriptor model. The benzene derivatives structure-toxicity models involve up to seven linear descriptors. Multiregression models, containing up to ten nonlinear descriptors, are also reported for the sake of comparison with previously obtained additivity models. Comparison with benzamidine complement-inhibitory activity models and with benzene derivatives toxicity models from the literature favors our novel approach. PMID- 10719638 TI - A computer-based approach to describe the 13C NMR chemical shifts of alkanes by the generalized spectral moments of iterated line graphs AB - A recently introduced strategy to quantitative structure-property relationship studies is used to model the sum of 13C NMR chemical shifts of alkanes. This graph theoretical approach is based on the spectral moments of the iterated line graph sequence (ILGS). The statistical quality of the model developed by using the present approach is compared to those obtained by using spectral moments of the bond matrix or the embedding frequencies of alkanes as independent variables. The embedding frequencies were computed on the basis of the spectral moments of the ILGS. The emphasis of this work is on the structural interpretation of the results. Consequently, the contributions of the different structural fragments of alkanes to the 13C NMR chemical shift sum are computed by the three theoretical approaches. The advantages and disadvantages of the use of these approaches are analyzed on the basis of computational expenditure, i.e. computer time and memory, and the statistical quality, complexity, and structural interpretation of the models developed. PMID- 10719639 TI - Diffusion and kinetics of reaction over bidispersive reforming catalyst AB - The objective of the study was to construct a mathematical model for optimizing the porous structure of the reforming catalyst. Using a layered model of the catalyst grain and the kinetic scheme adopted for the process, equations were derived to describe the mass transport in the catalyst grain under conditions of reforming. Particular consideration was given to the problem of how the parameters of the porous structure affected the efficiency of the process. Substantiated was the favourable influence of an appropriate macropore (transport pore) proportion in the catalyst grain on the course of the process. The optimum quantitative ratio of wide to narrow pores in the catalyst grain was established for determined pore radii. PMID- 10719640 TI - An algorithm for the calculation of the hyper-Wiener index of benzenoid hydrocarbons AB - An algorithm for the calculation of the hyper-Wiener index (WW) of benzenoid hydrocarbons (both cata- and pericondensed) is described, based on the consideration of pairs of elementary cuts of the corresponding benzenoid graph B. A pair of elementary cuts partitions the vertices of B into four classes. WW is expressed as a sum of terms of the form n11n22 + n12n21, each associated with a pair of elementary cuts; nrs, r, s = 1, 2 are the numbers of vertices in the respective four classes. The algorithm proposed enables a relatively easy calculation of WW, finding expressions for WW of homologous series of benzenoid hydrocarbons, and envisaging the relations between WW and molecular structure. PMID- 10719642 TI - Vestibulopathic gait: you're better off running than walking. PMID- 10719641 TI - SPAC: identification of polypeptides using their amino-acid composition. AB - We describe here new software able to retrieve from protein or nucleic acid databases, the sequence corresponding to a protein or peptide whose only amino acid composition and molecular weight are known. This algorithm is particularly devoted to the retrieval of partial sequences, a task that other available software performs poorly. Its accuracy for the attribution of a protein fragment to a sequence could represent an easy and economical first tool upstream the use of more sophisticated and expensive methods in proteomic research. SPAC (Sequence Protein Alignment with Composition) is a shareware available software on web site http://www.univ-tln.fr/ approximately grillas/SPAC/. This web site also presents help and a detailed description of the algorithm and interface. PMID- 10719643 TI - Disorders of higher visual function and hemi-spatial neglect. AB - Lesions of the extrastriate cerebral cortex result in disorders of vision, which reveal two cortical processing streams extending ventrally and dorsally from the primary visual cortex. Current theories emphasize the visual perceptual and visuomotor specialization of the ventral and dorsal streams, respectively. Recent investigations of hemi-spatial neglect suggest that the inferior parietal region may lie at the junction of the visual and motor processing pathways. PMID- 10719644 TI - Brain activation studies on visual-vestibular and ocular motor interaction. AB - Despite their different sensorimotor functions, saccades, pursuit eye movements, small-field optokinetic nystagmus and visual motion stimulation with the eyes stationary evoke a common complex pattern of activation in various cortical, basal ganglia, brain-stem and cerebellar areas. On closer inspection, however, typical subregions can be delineated that allow differentiation of adjacent but separate loci for specific functions (e.g. the separation of the two parallel corticocortical systems to control saccades and pursuit in the cortical eye fields). It is becoming increasingly clear that stimulation of one sensory system affects other sensory systems, and generally this is via an inhibitory reciprocal mode of interaction. For example, vestibular stimulation deactivates the visual cortex and visual stimulation deactivates the vestibular cortex. PMID- 10719645 TI - Motion sickness. AB - The number of recently published papers on motion sickness may convey the impression that motion sickness is far from being understood. The current review focusses on a concept which tends to unify the different manifestations and theories of motion sickness. The paper highlights the relations between ergonomic principles to minimise motion sickness and the concept predictions. The clinical management of sufferers from motion sickness in terms of selection, pharmacological measures and desensitisation courses is treated as well. PMID- 10719646 TI - Vestibular compensation and substitution. AB - This is a very brief update on the major papers since August 1998. Unilateral vestibular loss causes oculomotor, postural and sensory symptoms, all of which would be appropriate responses in a healthy person to a strong maintained angular and linear acceleration stimulus directed towards the healthy side. Within hours or days these static symptoms (so called because they are present without any externally imposed vestibular stimulation) reduce, and their progressive disappearance is called 'vestibular compensation'. However, careful testing with natural vestibular stimuli shows that the dynamic vestibular response after unilateral vestibular loss to passively imposed vestibular stimuli does not recover; it is usually asymmetric and functionally ineffective. Major recent developments are: (1) the permanent asymmetrical and functionally ineffective dynamic rotational vestibulo-ocular reflex responses to passive natural vestibular stimulation after unilateral vestibular loss and canal blocks in human patients; (2) evidence for the substitution of other sensory input and responses during vestibular compensation; (3) perceptual testing using visual perception of a horizontal line to confirm permanent otolith dysfunction; (4) the clear and substantial differences in post-unilateral vestibular loss vestibulo-ocular reflex responses between passive and active head turning; and (5) new results in brainstem physiology explaining the disappearance of static symptoms. PMID- 10719647 TI - Pharmacology of the vestibular system. AB - In the past year significant advances have been made in our understanding of the neurochemistry and neuropharmacology of the peripheral and central vestibular systems. The recognition of the central importance of excitatory amino acids and their receptors at the level of the hair cells, vestibular nerve and vestibular nucleus has progressed further, and the role of nitric oxide in relation to activation of the N-methyl-D-aspartate receptor subtype is becoming increasingly clear. Increasing evidence suggests that excessive N-methyl-D-aspartate receptor activation and nitric oxide production after exposure to aminoglycoside antibiotics is a critical part of hair cell death, and new pharmacological strategies for preventing aminoglycoside ototoxicity are emerging as a result. Conversely, the use of aminoglycosides to lesion the peripheral vestibular system in the treatment of Meniere's disease has been studied intensively. In the vestibular nucleus, new studies suggest the importance of opioid, nociceptin and glucocorticoid receptors in the control of vestibular reflex function. Finally, the mechanisms of action and optimal use of antihistamines in the treatment of vestibular disorders has also received a great deal of attention. PMID- 10719648 TI - Vestibular rehabilitation of patients with vestibular hypofunction or with benign paroxysmal positional vertigo. AB - Since the initial introduction of exercises as a treatment for patients with vestibular deficits, there have been numerous clinical reports on the benefits of treatment. Clinical reports, however, are of limited use as a basis for treatment because, without a control group, they offer only interesting descriptions of the patient populations. Fortunately, several prospective, randomized studies on the treatment of patients with vestibular hypofunction or with benign paroxysmal positional vertigo have been published recently, adding to the small number of previous publications. This review will examine the information provided by those studies. Advances in the use of outcome measures, assessment of otolith function and treatment of related balance problems are also presented. PMID- 10719649 TI - Diffusion and perfusion magnetic resonance imaging for the evaluation of acute stroke: potential use in guiding thrombolytic therapy. AB - Recent data indicate that diffusion/perfusion weighted imaging could eventually play a significant role in acute stroke management, particularly in determining the suitability of acute stroke patients for thrombolytic therapy. The evidence supporting these uses is reviewed, and the future role of diffusion and perfusion weighted imaging in acute stroke management is discussed. PMID- 10719650 TI - Intra-arterial thrombolysis in acute stroke. AB - A relatively small number of studies were published on the intra-arterial therapy of stroke in the past year. Those reports, however, do offer strong evidence for the feasibility and safety of this therapeutic approach. Partly on the basis of these data it has become evident that the intra-arterial thrombolytic therapy of acute ischemic stroke is at least as effective as intravenous thrombolysis. There remain, however, many unresolved issues before such therapy can become a part of the standard of care. PMID- 10719651 TI - Biological basis for statin therapy in stroke prevention. AB - Hypercholesterolemia has not been considered an important risk factor for stroke; however, statin therapy reduces stroke in coronary heart disease patients. Statins may provide cerebrovascular protection through various mechanisms that include a reduction in the incidence of embolic stroke from cardiac, aortic and carotid sites, stabilization of vulnerable carotid atherosclerotic plaque, and improvement in cerebral blood flow. PMID- 10719652 TI - New concepts in adult brain arteriovenous malformations. AB - Brain arteriovenous malformations are currently attracting increasing attention among clinicians as modern brain imaging techniques facilitate both diagnostic and follow-up evaluation. Their frequent presentation in young individuals, at times with flagrant clinical effects caused by cerebral hemorrhages or seizure disorders, keeps clinicians alert to any improvement in treatment strategies. Recent technical advances in surgical, endovascular, and radiation therapy add to the constantly accumulating data on clinical features, natural course, and treatment outcome in adult arteriovenous malformation patients. This review focuses on new concepts in arteriovenous malformation etiology, classification, treatment, and study approaches. PMID- 10719653 TI - Magnetic resonance imaging of cerebral microbleeds. AB - Magnetic resonance imaging of patients with primary intracerebral haemorrhage has drawn attention to focal areas of signal loss, which were suggested to indicate hemosiderin deposition from earlier bleeds. Correlative histopathologic data have recently confirmed this assumption and support a strong association between the occurrence of microbleeds and various types of small vessel disease, such as hypertensive lipofibrohyalinosis and cerebral amyloid angiopathy. Therefore, microbleeds that are detectable by magnetic resonance imaging could be viewed as markers for vessel wall disorders with a higher tendency for intracerebral bleeding. This finding appears to be of diagnostic importance, but could also help to predict a patient's risk for spontaneous rebleeding or bleeding complications after anticoagulation. PMID- 10719654 TI - Recent advances in magnetic resonance angiography of carotid and vertebral arteries. AB - New magnetic resonance angiographic sequences using gadolinium infusion allow high-quality images of supra-aortic vessels. Raw data may be obtained in a short scan time of less than 30 s, with a large acquisition volume from the aortic arch to the circle of Willis. After computed reconstruction of the vascular tree, angiograms appear similar to those obtained with conventional catheter angiography. Parameters of the sequence must be carefully chosen, however, with trade-offs between spatial resolution, scan time, acquisition volume and contrast of image. New developments have been proposed to improve the image quality with different acquisition strategies. These recent advances will probably be useful to assess the carotid and the vertebral arteries with more accuracy. They will require high-performance gradient systems and sophisticated software that is not yet available on all machines, however. PMID- 10719656 TI - Bibliography. Current world literature. Cerebrovascular disease. PMID- 10719655 TI - Bibliography. Current world literature. Neuro-ophthalmology and neuro-otology. PMID- 10719657 TI - [Self-certitude and self-being in schizophrenic delusional patients]. AB - When asking how a schizophrenic delusional patient can achieve a new, psychotically determined entity of his existence, we can differentiate the following aspects: 1. A quasi-transcendental organization of the delusional experience establishing the patient's singular self-certitude and the absoluteness of the delusion. 2. A reflexive self-confrontation and self disclosure within his psychotic experience as the basis of the patient's new, psychotically different self-being. 3. The directly intrusive experience of becoming psychotic as a fundamental transformation of oneself may trigger the awareness of a specific new-beginning and therefore of a radical change in personal self-being. This means as well an existential self-seclusion. PMID- 10719658 TI - [Prognostic and therapeutic relevance of cognitive characteristics for the long term course of schizophrenic illness following psychoeducational psychotherapeutic treatment]. AB - In a prospective, randomized clinical trial cognitive characteristics of schizophrenic patients were examined as predictors of the efficacy of a psychoeducational psychotherapeutic intervention. The aim of this study was to select adequate cognitive predictors. The reduction of the selected cognitive deficits by means of a psychoeducational psychotherapy was measured. Additionally, the prophylactic effects of the improvement of cognitive deficits were examined. Predictors of the course of illness were basic cognitive deficits and metacognitive constructs of 106 schizophrenic outpatients. Additionally, the modification of the cognitive skills of these patients was taken into account. Relevant factors of the course of illness representing the therapeutic effect of the intervention were investigated within a five-year follow-up. By means of logistic regression analyses thought disorders (AMDP system) and idiosyncratic and fatalistic assumptions (KK-scale) were obtained as appropriate cognitive predictors of the long-term course of illness. Thought disorders and attentional deficits could not be improved significantly. Though, there was a correlation between the therapeutic improvement of idiosyncratic and fatalistic assumptions and the rehospitalization rate within the follow-up. PMID- 10719659 TI - [Vocational rehabilitation in chronic mental illness. The current position]. AB - Vocational rehabilitation has long been of central importance in the comprehensive treatment of the psychiatrically disabled. This is reflected by the creation of a broad spectrum of vocational rehabilitation programs, ranging from inpatient and outpatient work therapy and sheltered employment to supported employment programs within the competitive labour market. Evaluation studies have shown that although sheltered vocational rehabilitation programs effect a significant rise in the work activity, rate of employment, job tenure and income of people with chronic mental illness in the alternative labour market, these programmes do not substantially increase job placement in the competitive labour market. By contrast, supported employment programs have proven more successful in achieving a higher integration rate in the competitive labour market for the chronically mentally ill. As opposed to the well-developed alternative labour market, supported employment programs are not broadly disseminated in German speaking countries. Furthermore, vocational rehabilitation in general suffers from a lack of systematic evaluation and thus from inadequate scientific foundation. PMID- 10719660 TI - [Motor assymetry]. AB - Although the study of handedness and its association with hemispheric specialisation represents the prevailing focus of motor dominance research, recent inquiry into footedness and other motoric asymmetries has stimulated several interesting propositions. The present review show the variability of assessing motoric asymmetries. Asymmetry in hand use takes two forms: differential hand preference and differential dexterity between the hands. Motoric asymmetries are strongest and most manifest for handedness (hand preference, performance and dexterity), descending through footedness and other less known asymmetries (tonguedness, chewing preference). Handedness is the most easily observed expression of cerebral lateralization. The alpha and omega of relating handedness to other neuropsychological variables or indices of lateral specialization lies in the classification of handedness. A vast range of testing techniques have been used to assess handedness. Writing hand and self-report are two of the most popular techniques. Other preference measures include observation of how people use tools and questionnaires. Performance tests assess speed and accuracy in tasks stressing manipulative dexterity. Although questionnaires are generally thought to be reliable and valid instruments, there is a disagreement as to the nature, the number and weighting of the items to be included. The least stable results will be obtained if a categorization into "right-handers" and "non right-handers" is made on the basis of exclusive "right" answers. Slightly more stable is a classification that is based on "right-handers", "mixed preference handers" and "left-hander" based on extreme choices in either direction and intermediate choices. We present shortly a possible inventory assessing motoric asymmetries. PMID- 10719661 TI - [Tobacco use and smoking cessation in the psychiatric patient]. AB - Smoking is far more prevalent among psychiatric patients than with healthy individuals. Particularly patients with substance-related disorders and schizophrenia often are addicted smokers. This is of great importance, as heavy smoking is clearly accompanied by a higher morbidity and mortality. Up to now there is no convincing explanation for the high prevalence of smoking. Numerous studies point to the fact that smoking is practised as a form of self-medication by psychiatric patients. The influences on cerebral dopaminergic or cholinergic transmission or even psychopharmacological treatment may reinforce smoking behaviour. Efforts to reduce the frequency of smoking in case of psychiatric patients have mostly proved fruitless. Prevention strategies as well as smoking cessation therapies seldom achieve success due to the poor motivation. Besides that smoking cessation often is complicated due to the simultaneous psychopharmacological treatment. The latest investigations, however, confirm the efficacy of cessation strategies, also in case of schizophrenia, mood or substance related disorders. Intensive behavioural treatment strategies as well as high dose nicotine replacement achieve encouraging long-term abstinence rates. Nicotine replacement by patch, gum or nasal spray might be a kind of causal treatment, assuming biological mechanisms to be responsible for heavy smoking. PMID- 10719662 TI - [The relationship between sleep position and therapeutic effect of the Esmarch Scheine appliance in sleep apnea syndromes]. AB - Recently an oral appliance is being used increasingly for the treatment of sleep apnea syndrome. But the success rate of oral appliance therapy shows large interindividual difference, and which factors influence its efficiency remain uncertain. To elucidate the influence of the sleep posture on the therapeutic effect of the Esmarch device, 58 patients with sleep apnea syndrome were investigated polysomnographically before and after insertion of the device. The sleep position during each apnea was classified into three types; supine, lateral and prone. The mean apnea index (25.6 +/- 18.7) decreased significantly (p < 0.0001) after insertion of the device (11.5 +/- 12.6). The number of apneas (in the supine and prone) positions was significantly reduced from 18.0 +/- 16.7 and 2.0 +/- 3.6 to 5.0 +/- 11.2, p < 0.001, and 0.3 +/- 0.4, p < 0.005, respectively, but that in the lateral position was slightly increased from 5.6 +/- 9.4 and 6.2 +/- 8.9. The percent of apneas was 70.3% for supine, 21.9% for lateral, and 7.8% for prone before therapy and 43.5%, 53.9% and 2.6%, respectively after therapy. The results indicated that the effectiveness of oral appliance therapy can differ greatly with the sleep posture. The sleep posture recorded polysomnography may be important for choice of oral appliance therapy and its prognosis. PMID- 10719663 TI - Innate immunity and the normal microflora. AB - This paper discusses the following ten subtitles with the contents indicated. 1. To meet a microbe: discusses the four alternatives in host-microbe interactions. 2. Receptors and signal transduction giving gene activation: discusses the lipopolysaccharide receptor and the limitations of cell cultures versus use of live animals. 3. Effector molecules--antimicrobial peptides with and without cysteines. A data base exists with over 500 sequences. This paper gives a general overview of five classes of gene-encoded effector molecules, based on the absence or presence of cysteines. These molecules are peptide antibiotics with wide spectra against different microbes. They are synthesized as propeptides and post translational modifications are common. 4. Effectors of innate immunity--lethal action without host damage: evaluates current opinions about the mode of action of peptide antibiotics and the fact that these effectors do not create host damage. 5. Genes, introns and movable elements. Two cecropin genes containing movable elements and the human cathelicidin gene for proFALL-39/hCAP18 are discussed. 6. The natural microflora. Hippos or frogs as model systems. This section includes the isolation of bacteria from the normal flora of frogs; Aeromonas hydrophila, the bacterium found on all five frog species studied; arguments and selected examples of frog-microbe interactions in vivo and in vitro; and the use of glucocorticoids as control for nuclear factor-kappa B/I kappa B alpha regulation of effector genes. 7. The use of germ-free mice--hard facts from hard work: summarizes new findings which indicate that germ-free mice are born with a set of antibacterial peptides in their small intestine. The intestine of germ-free mice monoinfected with A. hydrophila have peptide patterns that differ depending on a pretreatment with cortisone. 8. Looking back--an evolutionary perspective on innate immunity: arguments for an early evolutionary need for gene-encoded antibacterial factors. Caenorhabditis elegans should provide some answers. The finding of cecropin-like peptides in Helicobacter pylori and the indications that cecropins are derived from ribosomal protein L1. 9. What about viruses? Arguments for the lack of innate immunity against viruses. 10. Five questions floating in the pond of immunology. The normal microflora, its size and control are too often left out from immunological thinking. Animal model systems may sometimes invite misinterpretation. Which animal species are more equal than others? PMID- 10719664 TI - The role of nitric oxide in innate immunity. AB - Type 2 nitric oxide synthase (iNOS or NOS2) was originally described as an enzyme that is expressed in activated macrophages, generates nitric oxide (NO) from the amino acid L-arginine, and thereby contributes to the control of replication or killing of intracellular microbial pathogens. Since interferon (IFN)-gamma is the key cytokine for the induction of NOS2 in macrophages and the prototypic product of type 1 T-helper cells, high-level expression of NOS2 has been regarded to be mostly restricted to the adaptive phase of the immune response. In this review, we summarize data that demonstrate a prominent role of NOS2/NO also during innate immunity. During the early phase of infection with the intracellular pathogen Leishmania major, focally expressed NOS2/NO not only exerts antimicrobial activities but also controls the function of natural killer cells and the expression of cytokines such as IFN-gamma or transforming growth factor-beta. Some of these effects result from the function of NOS2/NO as an indispensable co factor for the activation of Tyk2 kinase and, thus, for interleukin-12 and IFN alpha/beta signaling in natural killer cells. PMID- 10719665 TI - The innate immune response of the respiratory epithelium. AB - The respiratory epithelium maintains an effective antimicrobial environment to prevent colonization by microorganisms in inspired air. In addition to constitutively present host defenses which include antimicrobial peptides and proteins, the epithelial cells respond to the presence of microbes by the induction two complementary parts of an innate immune response. The first response is the increased production of antimicrobial agents, and the second is the induction of a signal network to recruit phagocytic cells to contain the infection. Inflammatory mediators released by the recruited cells as well as from the epithelium itself further induce the expression of the antimicrobial agents. The result is an effective prevention of microbial colonization. The epithelial cells recognize the pathogen-associated patterns on microbes by surface receptors such as CD14 and Toll-like receptors. Subsequent signal transduction pathways have been identified which result in the increased transcription of host defense response genes. Diseases such as cystic fibrosis, or environmental exposures such as the inhalation of air pollution particles, may create an environment that impairs the expression or activity of the host defenses in the airway. This can lead to increased susceptibility to airway infections. PMID- 10719666 TI - Innate immunity and pulmonary host defense. AB - The lung, in order to facilitate gas exchange, represents the largest epithelial surface area of the body in contact with the external environment. As normal respiration occurs, the upper and lower airways are repeatedly exposed to a multitude of airborne particles and microorganisms. Since these agents are frequently deposited on the surface of the respiratory tract, an elaborate system of defense mechanisms is in place to maintain the sterility of the lung. Innate defenses are primarily responsible for the elimination of bacterial organisms from the alveolus. Early bacterial clearance is mediated by a dual phagocytic system involving both alveolar macrophages and polymorphonuclear leukocytes. The recruitment and activation of inflammatory cells at a site of infection involves the orchestrated expression of leukocyte and vascular adhesion molecules, as well as the establishment of chemotactic gradients via the generation of proinflammatory cytokines and chemokines. Immunologic manipulation of innate immunity may serve as an important adjuvant therapy in the treatment of both immunocompromised and immunocompetent patients with severe lung infections. As the complexities of the host-pathogen interaction are further dissected and elucidated, it is likely that the therapeutic benefits from these approaches will be realized. PMID- 10719667 TI - Collectins and pulmonary innate immunity. AB - The surfactant-associated proteins SP-A and SP-D are members of a family of host defense lectins, designated collectins. There is increasing evidence that these pulmonary, epithelial-derived proteins are important components of the innate immune response to microbial challenge and participate in other aspects of immune and inflammatory regulation within the lung. Both proteins bind to glycoconjugates and/or lipid moieties expressed by a wide variety of microorganisms, and to certain organic particles, such as pollens. SP-A and SP-D have the capacity to modulate leukocyte function and, in some circumstances; to opsonize and enhance the killing of microorganisms. The biologic activity of cell wall components, such as Gram-negative bacterial polysaccharides, or viral glycoproteins, such as the hemagglutinin of influenza viruses, may be altered by interactions with collectins. In addition, complementary or cooperative interactions between SP-A, SP-D and other host defense lectins could contribute to the efficiency of this defense system. Collectins could play particularly important roles in settings of inadequate or impaired specific immunity, and acquired alterations in the levels of active collectins within the airspaces and distal airways may increase susceptibility to infection. PMID- 10719668 TI - The roles of surfactant proteins A and D in innate immunity. AB - Research over the last decade on the surfactant proteins SP-A and SP-D suggests roles beyond surfactant lipid homeostasis, involving their participation in innate immune defence. SP-A and SP-D bind and agglutinate an impressive array of non-self structures, ranging from bacteria and fungi to allergens and environmental inorganic substrates. Complementing binding. SP-A and SP-D initiate and enhance immune cell ingestion and killing of targets. Recently, some exciting developments have extended and clarified their contributions to innate immunity. Knockout mice for SP-A and SP-D have been developed. The SP-A knockout confirms that SP-A plays a key role in defence against lung pathogens and reveals the underlying defense mechanisms that require SP-A. These surfactant proteins have also been shown to have important roles in modulating the immune response, instructing, yet quenching, the immune reactions in the lung. The crystal structure of SP-D plus functional studies with recombinantly altered forms of SP A and SP-D has begun to characterise the structural motifs responsible for mediating their immune functions. Linkage and polymorphism analysis is explaining the role these genes may play in lung diseases and infection. PMID- 10719669 TI - Carbohydrate-mediated recognition systems in innate immunity. AB - There is growing interest in carbohydrate-recognizing receptors of the innate immune system. Among them are members of the C-type lectin family, which include the collectins and the selectins and which operate by ligating exogenous (microbial) or endogenous carbohydrates. De novo assignments of the sequences of ligands for carbohydrate-recognizing receptors are among the most challenging topics in cell biology. This is because of the heterogeneity of oligosaccharides on proteins and lipids, and their availability only in limited amounts. To address the need for a microprocedure for direct binding studies with oligosaccharides derived from glycoproteins, we introduced the neoglycolipid technology for generating solid phase oligosaccharide probes for binding experiments. The technology has enabled assignments of unsuspected oligosaccharide ligands for the selectins and given valuable insights into those for the collectins. The ligands so far identified appear not to be unique for a given receptor system; there are considerable cross-reactions. Specificity can be created, however, through different modes of oligosaccharide presentation on macromolecular carriers, or the expression of a particular oligosaccharide sequence on a selected cell type in a given body compartment, and the regulated expression of the receptor protein at the desired location. The existence of unique ligand structures is not ruled out, however. Co-ligation of a receptor may also occur to a second carbohydrate or even to a non-carbohydrate ligand to create a unique assembly. A further group of C-type lectin-like proteins occurs on natural killer (NK) cells and NK T cells, and is associated with activation or inhibition of the cell effector functions. An important challenge is to determine whether carbohydrates are among physiological ligands for this important group of receptors. PMID- 10719670 TI - Innate immune recognition: mechanisms and pathways. AB - The innate immune system is an evolutionarily ancient form of host defense found in most multicellular organisms. Inducible responses of the innate immune system are triggered upon pathogen recognition by a set of pattern recognition receptors. These receptors recognize conserved molecular patterns shared by large groups of microorganisms. Recognition of these patterns allows the innate immune system not only to detect the presence of an infectious microbe, but also to determine the type of the infecting pathogen. Pattern recognition receptors activate conserved host defense signaling pathways that control the expression of a variety of immune response genes. PMID- 10719671 TI - The importance of gamma delta T cells in the resolution of pathogen-induced inflammatory immune responses. AB - The aim of our research is to determine the biological function of gamma delta lymphocytes and in particular the role they play in microbial immunity. Although evidence of gamma delta T-cell activation and expansion has been obtained from numerous infectious diseases, how they contribute to pathogen-induced immune responses is still not clear. Based upon extensive studies of gamma delta T-cell involvement in the immune response to viral and bacterial pathogens in both mice and humans, we have uncovered evidence of their direct involvement in terminating host immune responses to infection and preventing chronic disease. We have identified an interaction between peripheral gamma delta T cells and a population of activated, proinflammatory macrophages elicited by infection that occurs late in the course of infection during or after pathogen clearance. As a result of this interaction, activated gamma delta T cells acquire cytotoxic activity and kill the stimulatory macrophages, leading us to propose a model for gamma delta T cell-macrophage interactions that contributes to macrophage homeostasis, the resolution of inflammatory immune responses, and prevention of chronic inflammatory disease. PMID- 10719672 TI - Gamma delta T cells as mediators of mucosal tolerance: the autoimmune diabetes model. AB - Mucosal delivery of soluble antigen induces systemic tolerance and has been applied to the prevention of autoimmune diseases. We have studied mucosal tolerance in autoimmune diabetes using the non-obese diabetic mouse model. Treatment of prediabetic mice with the pancreatic islet autoantigen insulin, by aerosol or intranasal delivery, reduces the incidence of diabetes and is associated with induction of CD8 (alpha alpha) gamma delta T cells, small numbers of which prevent adoptive transfer of diabetes. We examine the evidence for gamma delta T cells in mucosal tolerance and discuss possible mechanisms underlying the induction and action of insulin-induced CD8 gamma delta regulatory T cells. CD8 gamma delta cells constitute the most abundant subpopulation of intraepithelial lymphocytes (IELs), the major lymphoid cell compartment and first line of cellular immune defence in the mucosa. Induction of regulatory CD8 gamma delta T cells requires conformationally intact but not biologically active insulin. In contrast, intranasal (pro)insulin peptide, or oral insulin which is degraded in the gut, induces CD4 regulatory cells. Regulatory gamma delta T cells secrete interleukin-10 in pancreatic lymph nodes, which could account for the antidiabetic and bystander suppressor effect of naso-respiratory insulin. The physiological role of gamma delta IELs in maintaining peripheral self-tolerance deserves further study. PMID- 10719673 TI - Negative selection of B lymphocytes: a novel role for innate immunity. AB - Evidence has accumulated that strongly supports a role for innate immunity in B cell tolerance. Specific recognition proteins, such as natural antibody and collectins, are present in serum that identify highly conserved self-antigens such as nuclear proteins and activate the classical pathway of complement. The direct localization of these types of antigens to the lymphoid compartment in a complement-receptor-dependent manner provides a mechanism to deal with immature, self-reactive B cells developing daily in the bone marrow. Since it would be disadvantageous for the organism to maintain self-reactive B cells, which cross react with microbial antigens, the innate immune system provides an efficient pathway for their removal or silencing. A possible outcome of a breakdown in this pathway, as found in individuals bearing natural deficiencies in complement C1q or C4, is autoimmunity such as systemic lupus erythematosus. PMID- 10719674 TI - Mast cells in innate immunity. AB - Mast cells are known to be the main effector cells in the elicitation of the IgE mediated allergic response. The specific location of mast cells within tissues that interface the external environment, and the extent of their functional capacity, including the ability to phagocytose and to produce and secrete a wide spectrum of mediators, have led investigators to propose a potential role for mast cells in innate immune responses. Certain microorganisms have been found to interact either directly or indirectly with mast cells. This interaction results in mast cell activation and mediator release which elicit an inflammatory response or direct killing leading to bacterial clearance. The in vivo relevance of these in vitro observations has been demonstrated by the use of complement deficient and/or mast cell-deficient and mast cell-reconstituted mice. In thus has been shown that both C3 and mast cell- and tumor necrosis factor-alpha dependent recruitment of circulating leukocytes with bactericidal properties are crucial to a full response in certain models of acute infection. Modulation of mast cell numbers in vivo was also found to affect the host response against bacterial infection. Thus, mast cells do have a role in innate immunity in defined animal models of bacterial infection. Whether mast cells participate in innate immune responses in the protection of the human host against bacteria remains to be determined. PMID- 10719675 TI - Innate immunity and graft rejection. AB - Although innate immunity evolved to combat pathogens, increasing awareness of a pivotal role in driving and shaping adaptive immunity has prompted this review on the role of innate immunity in graft rejection. We present evidence that grafts, especially xenografts, elicit innate responses, required for adaptive immunity. Particular attention is paid to studies by ourselves and others demonstrating the important role of innate immunity in T-cell trafficking. The mechanisms by which grafts elicit innate immunity are a fertile subject for further investigation and an important target for therapeutic intervention. PMID- 10719676 TI - [Surveillance of nosocomial infections: prospective study in a pediatric intensive care unit. Background, patients and methods]. AB - BACKGROUND, PATIENTS AND METHODS: From November 1997 through May 1998, the incidence of nosocomial infections was studied prospectively in a 10-bed multidisciplinary pediatric intensive care unit in Germany. A standardized surveillance [SEKI] system based on the National Nosocomial Infection Surveillance [NNIS] System of the Centers for Disease Control and Prevention [CDC] was used. The CDC definitions for nosocomial infections were adapted to the current practice of pediatric intensive care in Germany. Infection rates were calculated as infections per 100 patients, per 1000 patient-days, and per 1000 device-days (central venous catheters, urinary-catheters, and mechanical ventilation). RESULTS: Fifteen nosocomial infections were recorded in 201 patients during 1035 patient-days. The overall nosocomial infection rates were 7.5/100 patients and 14.5/1000 patient-days. Device-associated nosocomial infection rates for urinary-catheters and mechanical ventilation were 7.2/1000 utilization-days and thus below the 75th percentile of the last NNIS report. Central line infection rates were 10.7/1000 utilization days and therefore above the 75th percentile of the NNIS data (10.2/1000). The median length-of-stay was 5.1 days. CONCLUSIONS: Surveillance data are indispensable for internal and external quality control, and prospective surveillance of nosocomial infections should become an essential component of hospital infection control programs in pediatric intensive care in Germany. The standardized calculation of (device utilization ratios and) device-specific infection rates yields results which can be compared with national and international surveillance data. SEKI meets the criteria of a practice oriented, prospective and standardized surveillance system. Considerable efforts for collecting and interpreting the required data should be balanced against the benefit of prevention of nosocomial infections in this population of critically ill persons. PMID- 10719677 TI - [Procalcitonin in comparison to C-reactive protein as markers of the course of sepsis in severely immunocompromised children after bone marrow transplantation]. AB - BACKGROUND: PCT has recently drawn attention as a quite specific marker for bacterial, fungal, and parasitic origin of severe sepsis-syndrome. These specific properties could make PCT to an important tool for sepsis monitoring in severely immunocompromised children. The clinical value of PCT in comparison to CrP was investigated in children after bone marrow transplantation (BMT). METHODS: PCT was measured in the serum of 48 children (median age 12.4 years) after BMT in a prospective study. Results were correlated with the clinical findings and compared to the C-reactive protein (CrP). RESULTS: PCT showed a sensitivity for diagnosing a sepsis-syndrome of 56%, a specificity of 87%, a positive predictive value of 69%, and a negative predictive value of 80%. Regarding CrP they were 100%, 41%, 46% and 100% respectively. The relative risk to die due to sepsis syndrome was 26.4 for PCT levels over 10 ng/ml and 4.0 for CrP levels over 200 mg/l. It could be shown furthermore that there can be a significant liberation of PCT even during hematological aplasia. CONCLUSION: (1) Measuring PCT levels in the sera of children undergoing BMT improves the possibility of diagnosing severe infection and gives an important prognostic tool. (2) Measuring PCT can be recommended if severe sepsis-syndrome is suspected and there is an additional need for differential diagnosis and prognostic evaluation. PMID- 10719678 TI - [Brainstem acoustic evoked potentials in children with developmental disabilities -a useful tool for differential diagnosis]. AB - In a retrospective study brainstem acoustic evoked potentials (BAEP) were evaluated in 222 children with psychomotor retardation or dysmorphic signs. Registrations were done, when no clear response to acoustic stimuli of medium intensity (60-80 dBA) could be obtained during clinical examination. Only 118 children (53%) had normal BAEP. 50 patients (22%) suffered from hearing impairment. 39 children (17%) showed disturbances of neuronal conduction. In 15 cases (7%) a combination of both conditions occurred. The mean age of our children with hearing impairment was 33.1 months, no case having been diagnosed before. In 57% the impairment was of the conductive type with an amount of less than 40 dB nHL This type was predominant in children with skeletal dysplasias (43%), chromosomal aberrations (43%) and malformation syndromes (40%). Severe hearing deficits of the sensorineural type with more than 69 dB nHL were found in children with malformation syndromes (28%), perinatal injuries (23%) and cns malformations (16%). As far as reference data were available, the hearing impairment in the BAEP was confirmed in 92% by our pedaudiologists. As a consequence hearing aids were first prescribed in 10 children, their medium age being 33.6 months. In 18 cases grommets were inserted. 9 children required paracentesis and 4 adenotomy. Disturbances of neuronal conduction with increased interpeak latencies and deformed potentials were predominantly found in the group of children with neurometabolic diseases (67%) and cns malformations (32%). Early diagnosis of hearing impairment in children with psychomotor retardation remains a problem as it is in the general population. More attention in clinical examination and appropriate screening is necessary. BAEP provide a powerful tool for hearing screening and additional information for differential diagnosis especially in children with neurometabolic diseases. PMID- 10719679 TI - [Diabetes mellitus type I, celiac disease and Imerslund-Grasbeck syndrome: only an incidental combination of rare diseases?]. AB - We describe the clinical course of a girl with onset of type I diabetes mellitus at the age of 3 years. At the age of 10, coeliac disease and shortly thereafter a vitamin B12 deficiency anemia (Imerslund-Grasbeck-syndrome) was diagnosed. Her younger sister also suffered from Imerslund-Grasbeck-syndrome when she was 11 year old. This unusual, so far not described association of rare diseases suggests a common autoimmune etiology. PMID- 10719680 TI - [Pseudoarthrosis in neurofibromatosis type 1]. AB - Neurofibromatosis type 1 (NF1) is the most common neurocutaneous disease. The clinical manifestations are diverse. Some of the skeletal changes are most relevant to the patient. We report on 9 patients with NF1 who presented with typical pseudarthrosis. In 8 of these children the lower extremity was involved. In 2 cases lesions of both tibia and fibula were found, in one case even over long segments with a fully instable leg. One child had a complete pseudarthrosis of the radius and ulna. This report analyses the bony changes, the operations performed and the possible technical improvements to be made. The present study as well as other recent studies suggest that bony lesions should be operated early using microsurgical methods. PMID- 10719681 TI - [Retinitis pigmentosa, terminal renal insufficiency and Caroli syndrome: new associations with Opitz trigonocephaly syndrome]. AB - We report on a new patient with Opitz trigonocephaly syndrome. In addition to the findings typical of this mental retardation syndrome, the present patient has retinitis pigmentosa, Caroli's syndrome and renal failure, which is undergoing hemodialysis. This association is never observed before in patients with Opitz trigonocephaly syndrome. This case demonstrate, that with increased survival of patients with mental retardation syndromes, the phenotypes possible are modified. PMID- 10719682 TI - [Trisomy of the short arm of chromosome 10p; description of a female patient with de novo duplication 10p11.2-15]. AB - Trisomy 10p is a rare chromosomal syndrome, characterized by craniofacial abnormalities, malformations of organs and skeleton, and impaired psychomotor development. In most of the cases partial trisomy 10p results of a balanced translocation or inversion, the mother being carrier of the structural abnormality. Only eight of 63 patients with trisomy 10p found in a literature survey present a de novo trisomy. 17 cases show a pure trisomy 10p without an associated deficiency of any other chromosome segment. We report a female patient with an interchromosomal de novo duplication 10p11.2-->15, demonstrating typical clinical signs like craniofacial abnormalities, oral cleft, club foot, seizures, and a severe delay of psychomotor development. PMID- 10719683 TI - [Turkish infant with hypoelectrolytemia and metabolic alkalosis as the sole manifestations of a mild form of cystic fibrosis (mutation D110H)]. AB - We report the history of an infant who presented with hypotonic dehydration and metabolic alkalosis, in whom the diagnosis of cystic fibrosis was made on the basis of investigations for rare cystic fibrosis mutations. Since no other signs and symptoms of the CF disease were present, the finding of the rare mutation D110H on exon 4 of the CFTR-gene was paramount in the delineation of his underlying illness. He is now thriving well with a daily oral substitution of 1-2 grams of sodium chloride. A mild variant of cystic fibrosis has to be considered in infants presenting with unexplained hypoelectrolytemia and metabolic alkalosis. Like in our child, typical signs and symptoms of cystic fibrosis like maldigestion may not be present. The search has to be extended into "mild" mutations of the disease like D110H which was found in our case. PMID- 10719684 TI - [Brain SPECT in children and adolescents suffering from migraine]. PMID- 10719685 TI - [Clinical characteristics of migraine in children according to the age]. AB - We investigated 100 children with migraine admitted to the Developmental Neurology Department, Medical University of Gdansk between 1988-1997. The diagnosis was made on IHS Criteria (1988). There were 53 children with migraine with aura and 47--without aura. The children were divided into three groups according to the onset of migraine attacks (3-16 years of age). To reveal the relation between clinical manifestation of migraine and the age, statistical analysis was made. The results showed that a higher frequency of migraine with aura was observed in the oldest group of children. Boys were more predisposed to develop migraine earlier. The headaches most often were pulsatile and located in the forehead, rarely-lateralized. There was no relation between location of headache, age, sex and type of migraine. In 63% of children the duration of headache was above 2 hours, in 37%--from few minutes to 2 hours. Vomiting was noticed in two thirds of patients, phonophobia and photophobia were observed in 20 children with migraine without aura and in 5 children with migraine with aura. The frequency of migraine attacks was higher in migraine without aura. Aura symptoms were mainly sensory and visual. CT and MRI were negative. PMID- 10719686 TI - [Headache as the consequence of brain concussion and contusion in closed head injuries in children]. AB - The most frequent type of head injury in children is closed head trauma with brain concussion or contusion, and headache is the dominant complaint of early and late postinjury period. Because of scant number of studies on the problem of occurrence, characteristics and persistence of posttraumatic headache this study was undertaken in a group of 100 children (29 girls and 71 boys), aged 3-14 years old, 90 after brain concussion and 10 after contusion. Children with a history of injuries, central nervous system infections, with headaches before injury and chronic diseases were excluded. In 9 cases linear skull fracture was present after injury. The material was examined within one week after trauma, and then after 3, 6, and 12 months. After 3 months EEG was performed and repeated after 6 and 12 months in children with persistent headache or those without headache but with abnormal EEG results in the first examination. According to the additional diagnostic examinations, I excluded other causes of headache, besides head injury. In my observation 83% of children had headache after brain concussion and contusion. The majority--56%--had acute posttraumatic headache, but 27% of children complained of chronic headache, mainly tension type headache. Only 3% had migraine. In 21% of all group of 100 children, I noticed the headache persisting during the whole year of my observation. The important risk factor for the occurrence of posttraumatic headache were the age of child at the moment injury and the period of unconsciousness. The electroencephalographic recording still remains the important additional examination of posttraumatic consequences. PMID- 10719687 TI - [Night headache: report of 2 cases]. AB - The pathogenesis of hypnic headache is still unknown. Some authors supposed that the genesis of hypnic disorder is a decrease of melatonin secretion. It is a rare, moderately severe headache that occurs in middle-aged or elderly patients and affects both sexes. It appears exclusively during sleep and often with alarm clock regularity. The attack lasts from two to 6 hours, it may be unilateral or diffuse, pulsating or boring, without autonomic system features. Case 1. A 49 year old man reported the history of two week nocturnal headache that awakened him every night from sleep. The headache lasted two hours. There were no autonomic system features. Case 2. A 52 year old man suffered from nocturnal headaches for 6 weeks. The pain occurred between 2 and 5 almost every night and woke the patient. He suffered from diabetes mellitus too. Both patients were treated with flunarizine with good results. PMID- 10719688 TI - [Allergy effect on migraine course in older children and adolescents]. AB - The interaction between allergy and migraine has been discussed since many years. The aim of the present study was the evaluation of: 1. allergy prevalence in the studied children with migraine, 2. allergy effect on the clinical course of migraine. MATERIAL AND METHOD: The studied group comprised 30 children and adolescents aged 11-17 years with the diagnosis of migraine with or without aura by IHS criteria. In each case the migraine index was determined and symptoms of allergy were sought. Skin prick test and tests for immunoglobulin E in serum were done. The analysis of results showed the presence of allergy in 12 cases (40%), and on this basis two groups were isolated: with and without allergy. The allergic children were given antiallergic treatment for 6 months (pharmacological or diet restriction). RESULTS: In the allergy group the migraine index decreased significantly from 2.45 to 0.33. Headache intensity decreased as well. CONCLUSIONS: 1. In 40% of cases migraine was associated with allergy. 2. Treatment results suggest that allergy and antiallergic treatment may influence the course of migraine attacks. PMID- 10719690 TI - [The quality of life of school children and adolescents with headaches]. AB - One of the most common children's ailments are headaches--based on literature- about 75% of the population under the age of 15 suffers from them. The aim of the study was to evaluate the influence of headache on the quality of life in children and adolescents. The study group was made up of 86 persons between the age of 12 and 18. The study was conducted with the help of an anonymous questionnaire in the form of inquiry that was identical for all the respondents. The questions, contained in it, played a multidimensional role since, on one hand they were related to the mental, physical and social problems and on the other hand they informed us about the headaches frequency, intensity and localisation. The questionnaire was worked out on the basis of the Questionnaire Quality de Vie Migraine--QVM and it was modified for the needs of the conducted studies. The respondents were the patients of Chair and Department of Developmental Neurology at the University of Medical Sciences in Poznan and their outpatient clinic. The studies also took place in the region of Koscian, the primary and secondary school in Poznan, in school infirmaries where children and adolescents where because of headaches. Changes in the bio-psycho-social existence in the examined groups were found. Negative emotional states and the feeling of diminished self value in children and adolescents were estimated. High sickness absences at school, which didn't influence school records were also observed. A correlation was found between the experienced stress and the frequency of headache. PMID- 10719689 TI - [Prophylactic treatment in children with migraine presenting changes in electrophysiological and cerebral blood flow examinations: preliminary report]. AB - Idiopathic headache is common in children, but lack of specific tests makes diagnosis and treatment of migraine difficult. It has been proved, that in some migrainous children paroxysmal changes in eeg records can be found. The aim of the study was to examine the influence of prophylactic treatment on clinical course and bioelectric brain activity in children. The group examined consisted of 50 migrainous children (29 girls and 21 boys aged 6-18 years, mean 11.5 y.). In every patient routine eeg and ecg were recorded, and in 30 of them cerebral blood flow was examined. Some ecg abnormalities were observed in 6 children (12%) and cerebral blood flow disturbances--in 23 (77%). In 15 patients, in whom paroxysmal changes in routine eeg were found, an average 6-month prophylactic antimigrainous treatment (with Hydacorn in 14 and with Sermion in 1 patient) was performed and then control routine eeg and 24-hours eeg were recorded. In all children clinical improvement was observed after prophylactic medication. Paroxysmal changes in eeg records persisted in 6 children. The coexistence of electroencephalographic changes with disturbed cerebral blood flow migrainous children can indicate some connection between migraine and epilepsy, and/or reflect an influence of angiospasm resulting in hypoxia on the incidence of epileptiform changes in eeg records. The obtained results show clinical efficiency of combined treatment in child migraine and they reflect its good influence on bioelectrical brain activity. PMID- 10719691 TI - [Feverfew as a prophylactic treatment of migraine]. AB - Feverfew has been used in traditional medicine in the treatment of migraine for a long time. In 1985 E.S. Johnson et al. and later J.J. Murphy et al., 1988 and B.K. Vogler et al., 1994 published positive results of prophylactic use of this herb in migraine. Since 1994 through 1996 we studied in our Centre of Migraine Therapy the efficacy of feverfew in migraine treatment. We had 24 patients (women 19-61 years old) in our group. The drug was administered once daily (5 ml of the sap) for 30-60 days. We observed significant reduction of Migraine Index in 8 patients, less significant in additional 5. Our results confirm that feverfew may be beneficial in migraine prophylaxis as an additive drug. Further controlled studies need to be done, especially to establish the optimal dose of the drug. PMID- 10719692 TI - [Spontaneous headaches among secondary school students]. AB - The aim of the work was to evaluate the prevalence of headache in secondary school students. The methods of the study were based on the inquiry investigations of the students of 5 schools recruited randomly from 44 secondary schools in Lodz, following informed consent given by the School Supervision Authorities of the Lodz province, head-masters of the schools, parents, and by the students themselves. Personal questionnaires contained a number of questions referring to headaches that met the criteria of IHS Classification of Headaches. The students were inquired about localization of pains, their radiation, duration, frequency, time of occurrence (day or night), year and age of first onset of pains, associated symptoms, behaviour during pains, circumstances under which pains come on, family history relating to headaches. Headache subjects were then passed on for verification procedures consisting of questioning and physical examination. The results obtained were statistically elaborated basing on the independence chi-square test and F-Fisher's exact test. We studied 2351 students (1500 females, 851 males) aged 15-19 years. Headache in the past year was found in 2059 subjects: migraine in 511 (426 females, 85 males), of which 302 had migraine attacks more than once a month, and 67 had migrainous state; tension type headache (T-TH) in 1346 (847 females, 499 males), of which 414 had headache more than 12 days per year; chronic T-TH in 21; mixed headache in 148 (113 females, 35 males). Mixed headache frequency was determined on the basis of migraine attacks. A total of 125 students reported more than 12 attacks. Sixteen subjects had migrainous state. Idiopathic stabbing headache occurred in 54 students (19 females, 35 males) of which 12 suffered from more than 12 attacks per year. PMID- 10719693 TI - [Headaches in the population of school children in Poznan]. AB - A total of 2353 children and youths, 6-19 years old attending three primary and one secondary school situated in Poznan were examined. It was found that the overall prevalence of all types of headache (primary and secondary) in the general childhood population was 75%--1759 cases during the last 12 months. The medical examination of these school-children comprised full systematic physical and detailed neurological examination with visual acuity. Some laryngological examinations and laboratory tests also were made. The diagnosis of primary headaches was made according to the operational diagnostic criteria of the IHS classification. We was found 675 (28.69%) cases of tension type headache, 198 (8.42%) migraine, and only one case of cluster headache (0.04%). The prevalence of headache increased progressively with increasing age. Obtained data confirm an important part of positive family history and small influence of birth weight and living conditions. The other findings indicate that sleeping problems, dizziness, motion sickness, abdominalgia, belong to important risk factors for headache in children. At the end, it was made a comparison between "migrenous" and tension headache group. The conclusion is that headache in children and youth is serious social and medical problem. PMID- 10719694 TI - A randomized study of the use of a customized immobilization system in the treatment of prostate cancer with conformal radiotherapy. AB - PURPOSE: To evaluate the impact of a customized immobilisation system on field placement accuracy, simulation and treatment delivery time, radiographer convenience and patient acceptability. PATIENTS AND METHODS: Thirty men receiving radical radiotherapy for prostate cancer were randomised using a cross over trial design to have radiotherapy planning and treatment given either in a conventional treatment position (CTP) or using an immobilisation system (IMS). The randomisation was to have either the CTP or IMS for the initial 3 weeks of radiotherapy after which patients were replanned and changed to the alternative treatment set-up. Treatment accuracy was measured using an electronic portal imaging device. Radiographers and patients completed weekly questionnaires. RESULTS: Median simulation time was 22.5 min (range 20-30 min) in the CTP and 25 min (range 15-40 min) for the IMS (P < 0.001). Median treatment time was 9 min for CTP (range 8-10 min), and 10 min (range 8.5-13.5 min) for IMS (P < 0.001). Median isocentre displacement for anterior fields was 1.7 mm from the simulated isocentre for the CTP compared to 2.0 mm for IMS (P = 0.07). For left lateral fields values were 1.8 and 1.8 mm (P = 0.98), and for right lateral fields 2.1 and 1.7 mm (P = 0.06), respectively. No clinically significant reduction in either systematic or random field placement errors was demonstrated. Radiographers reported that patients found the IMS more comfortable than CTP (P < 0.001), but when using the IMS, they noticed greater difficulty in patient positioning (P < 0.001), and alignment to skin tattoos (P < 0.001). CONCLUSIONS: Although IMS may have been more comfortable, treatment accuracy was not improved compared to the CTP in our department. In addition, treatment took longer and patient set-up was more difficult. PMID- 10719695 TI - Endoscopic scoring of late rectal mucosal damage after conformal radiotherapy for prostatic carcinoma. AB - PURPOSE: To describe rectal mucosal damage in an endoscopic study after conformal radiotherapy of prostate cancer and to correlate this with clinical outcome. MATERIALS AND METHODS: Flexible rectosigmoidoscopy was performed on 44 patients who voluntarily accepted the examination. The median follow-up was 29 months (20 41 months) after 3-D-planned conformal radiotherapy of prostate cancer (66 Gy at the ICRU Reference point, 2 Gy per fraction). To enable a systematic topographic description of endoscopic findings the rectum was divided into four sections. Additionally we differentiated between anterior, posterior, right and left lateral rectal wall. Due to the lack of an existing valid graduation system for radiation induced proctitis, we introduced a six-scaled rectoscopy score for describing and reporting endoscopic findings based on the standardization of the endoscopic terminology published by the ESGE (European Society for Gastrointestinal Endoscopy). Endoscopic findings were compared to the EORTC/RTOG morbidity score. In addition, since 3-D dose distribution of organs at risks was available, a correlation could be made between the location of the rectal lesions and the absorbed dose at that level. RESULTS: In general, endoscopic findings increased from the proximal rectum to the anorectal transition, as well as from the posterior to the anterior rectum wall. Telangiectasia grade 1 and 2 were observed at the whole circumference, only telangiectasia grade 3 were limited to the high dose region at the anterior rectum wall. Similar results were found for congested mucosa (reddening and edematous mucosa). Correlation with symptoms, 7/9 patients who suffered from intermittent rectal bleeding (EORTC/RTOG grade 2) had multiple telangiectasia grade 2-3 and/or congested mucosa grade 3 and microulcerations. However, the same extent of mucosal damage (rectoscopy score 2 3) was found in seven out of 35 patients who have never developed a period of macroscopic rectal bleeding. CONCLUSION: Rectoscopy offers the possibility of detecting signs of tissue dysfunction below the level of subjective symptoms. Systematic analytic examinations such as rectoscopy, in addition to clinical examinations, as already foreseen in the LENT-SOMA-score, will be necessary due to the fact that even telangiectatic lesions have been observed for asymptomatic patients. For the opportunity of sharing and comparing data collected from endoscopy after radiotherapy a graduation system as proposed based on a standardisation of the endoscopic terminology will be necessary. PMID- 10719696 TI - Real-time three-dimensional motion analysis for patient positioning verification. AB - BACKGROUND AND PURPOSE: This paper describes the technology and methods involved in a system for automatically checking the position of patients at radiotherapy units. This is proposed for improving the accuracy in the irradiation geometrical set-up, which is a crucial factor in radiotherapy quality control. MATERIALS AND METHODS: The system is based on real-time opto-electronics and close-range photogrammetry and detects multiple passive markers placed on selected patient skin landmarks. Patient alignment and position monitoring is carried out by comparing the current three-dimensional positions of the markers with those of an initial reference position acquired during the simulation procedure and/or the first irradiation session. The system was used to measure the accuracy of conventional laser centering techniques for patient repositioning. Inaccuracies due to breathing and random movements were also taken into account. Professional technicians were asked to reposition three volunteer subjects carefully using a traditional laser centering procedure. RESULTS: The results revealed significant repositioning errors even in highly controlled conditions, affecting particularly body areas relatively far from the skin reference points used for laser alignment. CONCLUSIONS: The outcome of the experimental application of this technology confirms its potential as a tool for patient repositioning and automatic detection of any errors caused by breathing or other unpredictable movements. The real-time feedback on the patient's position given by the system provides operators with appropriate visual indices and allows them to take suitable countermeasures in case of significant failures. In addition, the use of the system output for automatic position control is envisaged. PMID- 10719697 TI - Variation of relative transit dose profiles with patient-detector distance. AB - BACKGROUND AND PURPOSE: In view of using portal images for exit dosimetry, an experimental study is performed of relative transit dose profiles at different distances behind patients (and phantoms) and of their relation to the exit dose profile. MATERIALS AND METHODS: Irregular, homogeneous polystyrene phantoms with a variable thickness to simulate head and neck (H&N) treatments (6-MV photon beam) are investigated by ionization chamber measurements performed close to the exit surface and at various distances behind the phantom (10, 20 and 30 cm). Similar measurements are performed for a rectangular phantom with large inhomogeneities (A1 and air). For one irregular homogeneous phantom and an irregular phantom containing an A1 inhomogeneity, ionization chamber measurements are performed at the exit surface, and a portal film image is taken at 30 cm behind the phantom. Portal films of a patient treated for a head and neck malignancy are evaluated for different air gaps behind the patient. RESULTS: For the irregular phantoms, deviations up to 15% and more are observed between the exit dose profile (along the shaped surface of the phantom) and the transit profile close to the phantom (perpendicular to the beam axis). There is, however, a good agreement--within 3%--between the exit profile and the transit profile at 30 cm. For the rectangular, inhomogeneous phantom, the deviation between the exit profile and the transit dose profile at 30 cm does not exceed 5%; transit dose profiles overestimate the exit dose for the air cavity and underestimate the dose for the A1 inhomogeneity. Measurements on portal films of a H&N patient for different air gaps confirm the order of magnitude of the difference observed between transit dose profiles close to the patient and transit dose profiles at some distance behind the patient. CONCLUSIONS: For 6-MV photon beam treatments with significant thickness variations (H&N), large variations (> 10%) are observed in transit dose profiles as a function of the air gap between the patient and the portal film. For this energy, a good agreement is found between the exit profile and the transit profile at about 30 cm behind the patient. PMID- 10719698 TI - A method to estimate the transit dose on the beam axis for verification of dose delivery with portal images. AB - PURPOSE AND BACKGROUND: A feasibility study is performed to evaluate the possibility of using the transit dose of portal images on the beam axis to measure the accuracy in dose delivery. The algorithm and the method are tested on a breast phantom and on patients with a breast disease. MATERIALS AND METHODS: To estimate the transit dose at various air gaps behind the patient, a method is proposed which applies, for a given air gap, the inverse square law to the primary component of the exit dose and an experimentally determined function for the scatter component of the exit dose. It is assumed that the primary component and the scattered component of the exit dose are given by the treatment planning system. The experimental function for the variation of the scattered component with the air gap, determined by phantom measurements, is modelled by an analytical function which contains only field size, air gap and one energy dependent parameter. RESULTS: The measurements on the breast phantom yield a maximum deviation between measured and estimated transit doses of 4.5%. The mean deviation is 0.9% with a standard deviation of the distribution of 2.3%. In vivo diode measurements on the same phantom yield a maximum deviation of 2.7%. Transit dose measurements on the beam axis for 45 portal images of breast patients show a mean deviation of 0.0% between the measured transit dose and the estimated transit dose. The standard deviation of the distribution is 4.4%. The method seems to be very sensitive to patient positioning and to discrepancies in breast thicknesses used for treatment planning. CONCLUSION: Preliminary results on breast patients show that the method proposed to evaluate transit doses on the beam axis from portal images may be a valuable alternative to conventional in vivo exit dosimetry. The method can be implemented in a simple way and does not require additional time during the irradiation session, as exit dosimetry with diodes does. The transit dose is only considered in one point. Nevertheless, in the framework of quality assurance of treatment delivery, this study is an example of the possibilities of monitoring at the same time the visual evaluation of the irradiated volume as well as the dosimetric control (i.e. in Gy) of treatment delivery with portal images. PMID- 10719699 TI - Observation study of electronic portal images for off-line verification. AB - PURPOSE: The goals of this study were to evaluate the use of electronic portal imaging device (EPID) paper images as off-line verification tools and to assess the feasibility of replacing portal films by EPID printed images. MATERIALS AND METHODS: Electronic portal images were acquired using a video-based imaging system. After contrast enhancement, these images were printed and compared to portal films when prescribed, and judged about their usefulness for off-line verification. A total of 2025 images were acquired from 322 fields on 137 patients. The images were shown to eight radiation oncologists and two senior residents in radiation oncology, each one of them judging fields relevant to his (her) daily practice. The questions asked were related to the choice of important anatomical structures and the visibility of such structures, the usefulness of the printed images, the comparison with portal films and the possible replacement of such films by paper images. RESULTS: Answers to the different questions were treated as quantitative scores. For the visibility question, means and standard deviations were calculated for each individual structure, then a global score was obtained for a given treatment site. Means and standard deviations were also computed for the comparison question. Proportions and confidence intervals were used for the other questions. The results show that EPID paper images are useful for some treatment sites such as breast, thorax, prostate, abdomen, pelvis (other than rectum) and axilla. The image quality remains insufficient for some other sites such as head and neck and spine. CONCLUSION: Although global anatomical landmarks scores are good, the usefulness score is not always as high because some essential anatomical structures scores must be taken into account. There is also a strong habit factor related to acceptance of EPID printed images as verification tools. As long as they see more and more images, radiation oncologists can more easily visualize anatomical structures and are less stringent when evaluating the efficiency of EPID paper images as off-line verification tools. PMID- 10719700 TI - The dosimetric consequences of inter-fractional patient movement on conventional and intensity-modulated breast radiotherapy treatments. AB - BACKGROUND AND PURPOSE: A method has been developed to enable a comparison to be made between the effects of movement on conventional tangential breast treatments and intensity-modulated treatments delivered using compensators. MATERIALS AND METHODS: The effects of set-up error and organ motion were studied for a set of six patients. Images were taken of these patients over the course of their treatment and conventional wedged and compensated treatment plans were designed for each. Dose-volume statistics were used to evaluate each of the treatment plans by examining the volume outside the dose range 95-105%. To assess the effects of movement alone, the volume change from day 1 was also calculated. RESULTS: Thirty-six estimated CT-sets were available for evaluation. Measurements of breast volume showed the volume to increase to a peak between fraction 4 and 8 and then decrease back below the initial volume. The standard treatment was found to yield 29/36 plans (81%) with greater than 5% volume outside the dose range 95 105%. For the compensated plans this dropped to 11/36 plans (31%). The analysis of the volume changes from day 1 showed that for both standard and compensated treatments 7/30 plans (23%) had an increase in volume outside the dose range 95 105% of greater than 5% of the total planning target volume. CONCLUSIONS: The compensated treatment is more susceptible to patient movement. However, the actual volume of tissue outside 95-105% dose is less when compared to standard treatment implying the compensated treatment is still superior. PMID- 10719701 TI - The IAEA/WHO TLD postal programme for radiotherapy hospitals. AB - BACKGROUND AND PURPOSE: Since 1969 the International Atomic Energy Agency (IAEA), together with the World Health Organization (WHO), has performed postal TLD audits to verify the calibration of radiotherapy beams in developing countries. MATERIALS AND METHODS: A number of changes have recently been implemented to improve the efficiency of the IAEA/WHO TLD programme. The IAEA has increased the number of participants and reduced significantly the total turn-around time to provide results to the hospitals within the shortest possible time following the TLD irradiations. The IAEA has established a regular follow-up programme for hospitals with results outside acceptance limits of +/- 5%. RESULTS: The IAEA has, over 30 years, verified the calibration of more than 3300 clinical photon beams at approximately 1000 radiotherapy hospitals. Only 65% of those hospitals who receive TLDs for the first time have results within the acceptance limits, while more than 80% of the users that have benefited from a previous TLD audit are successful. The experience of the IAEA in TLD audits has been transferred to the national level. The IAEA offers a standardized TLD methodology, provides guidelines and gives technical back-up to the national TLD networks. CONCLUSION: The unsatisfactory status of the dosimetry for radiotherapy, as noted in the past, is gradually improving; however, the dosimetry practices in many hospitals in developing countries need to be revised in order to reach adequate conformity to hospitals that perform modern radiotherapy in Europe, USA and Australia. PMID- 10719702 TI - Radiotherapy of advanced mycosis fungoides: indications and results of total skin electron beam and photon beam irradiation. AB - BACKGROUND: The goals of this retrospective study of advanced mycosis fungoides are (1) to describe the indications of a combination of total skin electron beam and photon beam irradiation and (2) to analyze the results of total body or segmental photon irradiation for patients with extension beyond the skin. METHODS: From January 1975 to December 1995, 45 patients with pathologically confirmed mycosis fungoides or Sezary syndrome received a combination of TSEB and photon beam irradiation for advanced disease: 34 males and 11 females, mean age 61 years (range 27-87 years). The mean follow-up was 111 months (range 18-244 months, median 85 months). Whole-skin irradiation treatment to a depth of 3-5 mm with a 6-MeV electron beam was produced by a linear accelerator to a total dose of 24-30 Gy in 8-15 fractions, 3-4 times a week. In cases of thick plaques or tumors that were beyond the scope of low energy electron beams or for treating nodal areas (especially in the head and neck area or axilla involvement), regional irradiation (RRT) with Co-60 photon beams was followed by whole-skin electron beam irradiation (15 patients). In cases of diffuse erythrodermia, Sezary syndrome, nodal or visceral involvement, total body irradiation was delivered with a 25-MV photon beam using a split-course regimen to prevent hematological toxicity (22 patients). The first course consisted of 1.25 Gy delivered in ten fractions and 10 days. Subsequently, patients received TSEB. Four to 6 weeks after TSEB, they received a second course of 1.25 Gy. The cumulative TBI dose ranged from 2.5 to 3 Gy in about 3 months. Hemi-body irradiation (HB) with Co-60 (and a bolus) was given in cases of multiple regional tumors with large and thick infiltration of the skin to a dose of 9-12 Gy (using fractions of 1-1.5 Gy/day) which, once flattened, were boosted with whole-skin electron beam therapy (8 patients). RESULTS: At 3 months, the overall response rate was 75% with 23/45 (51%) patients in complete response and 24% in partial response; one patient had stable lesions and 1 patient presented progressive disease. The overall response rate was 81% for T3 patients, 61% for T4, 79% for N1 and 70% for N3. The complete response rate was 67% for T3 and 28% for T4. Sixty-four percent of N1 patients and 41% of N3 had a complete response. The 5 year actuarial overall survival was 37% for T3 and 44% for T4 (P = 0.84). Patients with clinically abnormal lymph nodes that were pathologically negative (N1) presented a 5-year survival of 63%. Patients with pathologically positive lymph nodes (N3) experienced a 5-year survival rate of 32% (P = 0.040). CONCLUSIONS: TSEB provides an excellent quality of life by reducing itching and discharge from the skin. Patients with more advanced disease may be treated and cured by the addition of photon beams in combination with TSEB. A selection of patients with advanced skin disease and regional extension may be cured by a combination of TSEB and photon beam irradiation. The regional treatment allows the use of electrons after the reduction of the plaques or thick tumors and a prophylactic irradiation of the adjacent nodal area. PMID- 10719703 TI - Sweat gland function as a measure of radiation change. AB - BACKGROUND: Radiotherapy may result in dryness of the skin even when no other change can be detected. We describe a system for recording the electrical conductance of skin as a measure of sweat gland function. PATIENTS AND METHODS: In 22 normal volunteers close agreement was obtained between measurements obtained from comparable sites on both sides of the chest. Measurements were subsequently made in 38 patients treated by radiotherapy to one side of the chest for tumours of the breast or lung using one of five different fractionation schedules. Simultaneous readings were obtained from both sides of the chest with the non irradiated side acting as a control. RESULTS: A dose response relationship was demonstrated: five patients who received the equivalent total dose of 15 Gy in 2-Gy fractions showed no change in conductance. Sixteen out of 23 who received an equivalent total dose of 42-46 Gy in 2-Gy fractions had a greater than 22% reduction in mean skin conductance compared with that of the control areas despite the skin appearing normal in the large majority. Marked changes in skin conductance were seen after higher total doses. In a prospective study 18 women receiving breast irradiation underwent weekly readings during treatment. A mean reduction of 40% in skin conductance was noted by the end of the second week of treatment prior to any clinical evidence of radiation change. Skin conductance returned to normal in 44% of patients by 6 months. In the remainder, those patients who showed the greatest reduction in skin conductance during treatment demonstrated the least recovery. CONCLUSIONS: Changes in sweat gland function can be detected and quantified in skin which may otherwise appear normal. Differences may so be demonstrated between areas treated using different fractionation schedules and the method may be applied to the detection during radiotherapy of unusually sensitive patient. PMID- 10719704 TI - Variation in parotid gland size, configuration, and anatomic relations. AB - PURPOSE: To examine the variation in the anatomy of parotid glands discerned by magnetic resonance imaging. METHODS: Head and neck magnetic resonance scans of 16 patients (representing 32 glands) whose studies consisted of 5 mm contiguous sections were selected at random. The T1 weighted scans were thresholded and outlined to only encompass the parotid tissue. A volumetric analysis program (ISG Technologies, Inc.) was used to compute the parotid volume in cubic millimeters. Each of the 32 glands was measured independently by two observers. RESULTS: The difference between observers averaged 4.8%. The median volume was 25,262 mm3, range 9225-54,080 mm3. In four patients there were considerable differences in the volumes of the right and left parotid glands, with variations of 9, 10, 14 and 29%. In nine patients, (18 glands) the depth from the medial edge of the gland to the spinal cord ranged from 19-32 mm. However, the maximum variation between the two sides in a single patient was 4 mm. Observations made include: (1) parotid glands extending anterior to the masseter muscle, or posterior to the posterior margin of this muscle; (2) parotid glandular tissue extending above the zygoma and the external auditory canal; (3) parotid tissue extending posteriorly to overlap the spinal cord; (4) parotid glands extending below or remaining above the angle of the mandible; and (5) wide variation of the transverse dimension of the parotid glands, with one measuring 4.8 cm. DISCUSSION: To ensure that the entire parotid is or is not in a treatment field a computerized tomography or magnetic resonance scan is necessary. If a specific portion of the gland must be in the field a volume histiogram must be available. PMID- 10719705 TI - Quality assurance into the next century. PMID- 10719706 TI - [Environmental health standards in Germany. Comparison of methods and results]. PMID- 10719707 TI - [Criteria for using epidemiological data in risk assessment and permissible exposure level determination]. PMID- 10719708 TI - [Adverse effects as a human toxicological problem]. PMID- 10719709 TI - [Uncertainty analysis of quantitative risk assessment in temporally variable exposures: model observations based on biological and epidemiological risk models]. AB - Unit risks used for quantitative cancer risk assessment are defined for constant lifetime exposures. The condition of temporal stability, however, usually is not fulfilled in environmental health applications. In practice, cancer risks for time-dependent exposures are often estimated by calculating lifetime average exposure, assuming a mean life expectancy of 70 years. In the present paper we discuss the question whether this is an appropriate procedure considering various variants of multi stage and epidemiological relative risk models. For this purpose, lifetime risks for time dependent exposures as calculated according to the respective model assumptions, were compared with lifetime risks estimated by the lifetime average exposure approach. As typical exposure histories in environmental health applications we studied exposures either limited to the first 5 years of life (children scenario) or limited to duration of employment (30th to 65th year of age; occupational scenario). The consideration of multistage models (Armitage-Doll- and Moolgavkar-Venzon-Knudson model) in general would not induce serious bias in risk estimation when exposures are limited to middle ages (occupational scenario). On the other hand, when exposures occur only in very young ages or only in very old ages the risk estimated by using lifetime average exposure is not comparable with the predictions of multistage models. Whereas the degree of possible underestimation is bounded by factors well below 10, the amount of possible overestimation is unbounded and may become arbitrarily high, when exposures concentrate in extreme ages. In a second part of the study we investigated different relative risk models, taking lung cancer as an example. The models differed with respect to assumptions on latent periods and moderating effects of age at exposure and age at risk. The simulations showed that the unit risk concept is appropriate for the occupational scenario. For the children scenario results strongly depend on the assumptions made. Whereas the degree of possible underestimation is acceptable, in some models the degree of possible overestimation may become arbitrarily high. Both parts of the study showed that bias induced by using lifetime average exposure is acceptable when exposures are limited to middle ages. On the other hand, the unit risk concept should not uncritically be applied to exposures limited to early childhood (e.g., in kindergartens or due to mouthing activities). Depending on the assumptions made, lifetime risk may either be moderately underestimated or grossly overestimated. Without additional knowledge on mechanisms or latency period risk estimations are of questionable value. With respect to exposures in childhood regulation should concentrate on initiating substances or substances known to have long latent periods, respectively. With respect to cancers which occur relatively frequent already in childhood specific considerations are recommended. PMID- 10719710 TI - [Assessment of absorbing capacity of hazardous substances in contaminated materials and soil by an in vitro digestion test system]. PMID- 10719711 TI - [Limit values between toxicology and legal guidelines]. PMID- 10719712 TI - [Height-induced vertigo and its medical interpretation: Goethe and the Strassburger Munster]. AB - An analysis combining medicine and literature challenges the methodology of both disciplines. This essay on the vertigo Goethe suffered on the tower of the Strasbourg Minster attempts to trace the vicissitudes of interpreting an emblem, like vertigo, burdened by cultural meaning and implications. Thus, Goethe's own report of this event 40 years after the fact, in his "Dichtung und Wahrheit", has to be related to another, hidden chronology of vertigo and fear in his account which, at first glance, conveys quite different implications. The first part of this paper refers to a medical interpretation of Goethe's dread of high places and his way of coping with it which, today, could be defined as a typical example of a behaviourist approach. In the second part, Goethe's vertigo is linked to psychoanalytic, literary, and historical reflections on the meanings of symptoms we connect today with medical terms like anxiety, phobia, and vertigo. Goethe's vertigo is shown as a complex problem--not only for himself but also for its interpreters: on the one hand, it tells its own story-within-a-story; on the other, it depends on the tools it was written with. Traditional approaches of medical history try to find symptoms and traces of diseases known to us today in literary texts, an approach which is as dubious as taking today's tools of medical analysis, such as psychoanalytic terms and concepts, to explain specific phenomena in literature without first carefully analysing these methods themselves, and only then subjecting the text to an analysis based on them. Nevertheless, this essay does not contest the justification of interpreting literary texts in the light of today's medical knowledge, but postulates that it should be clear which type of medical knowledge is applied. It is quite possible to read Goethe's account only as an old tale of acrophobia, but how will this help us? It seems more interesting to look at the link between the feeling of dizziness he experienced on top of the tower and the "cultural feeling" of the Sturm and Drang period, and then to trace the perception of this feeling at that time within its cultural context and in the light of prevalent medical theory. PMID- 10719713 TI - [Drug treatment of chronic inflammatory bowel diseases: current status and prospects]. AB - The medical treatment of inflammatory bowel disease is dependent on disease activity and bowel involvement. Severe Crohn's disease and ulcerative colitis are primarily treated with corticosteroids. Alternatively, if inflammation is localised in the right colon and ileum, budesonide may be used in view of its low systemic side effects. In distal colitis and perianal disease, topical therapy with steroids is very effective. In moderate disease preparations containing 5 aminosalicylate (5-ASA) may be used. The latter are highly effective applied locally in distal disease. Steroids should be tapered down and whenever possible not used to maintain remission. In patients with ulcerative colitis, 5-ASA is effective in maintaining remission, whereas this medication plays only a limited role in Crohn's disease. Refractory disease or patients with multiple flare-ups should be treated with azathioprine. Infliximab, an anti-TNF-alpha antibody, is a potent therapy for fistulising Crohn's disease or steroid-refractory disease. Other approved and experimental treatments are discussed. PMID- 10719715 TI - [Significance of cytokines for pathophysiology and clinical aspects of Helicobacter pylori]. AB - H. pylori is the cause of one of the most common infectious diseases worldwide. Infection with H. pylori leads to exaggerated synthesis of several inflammatory cytokines. Only a minority of infected patients develops peptic ulcer disease. H. pylori strains associated with peptic ulcer disease induce strong mucosal infiltration with inflammatory cells and increased expression of several cytokines. Cytokines may contribute to ulcer development by different mechanisms, including stimulation of gastrin and pepsinogen release, suppression of somatostatin synthesis and activation of inflammatory cells. PMID- 10719714 TI - [Portal hypertension and angiogenesis]. AB - The pathophysiological background of the haemodynamic changes in chronic portal hypertension is still only partly understood. An important factor is the so called "forward theory", explaining the systemic and especially splanchnic vasodilation observed. Nitric oxide (NO) is an important mediator of this vasodilatory state. Structural as well as vascular changes seem to be causative factors in this vasodilation. These alterations can be studied by a newly developed in vivo angiogenesis assay. Rats with portal hypertension show increased angiogenesis and NO appears to be a mediator of this finding. The interaction(s) of NO with other angiogenic molecules is now being explored. A better understanding of these structural vascular changes may help to develop new therapeutic strategies in the future. PMID- 10719716 TI - [Upper gastrointestinal hemorrhage?]. PMID- 10719717 TI - [Do physicians interpret therapy outcome differently than students?]. AB - Results are presented in the most varied ways in medical publications and advertising material. The reader should be able to interpret the results of such studies accurately. In the present investigation our interest focused on whether a physician learns to analyse results correctly in medical school or in his subsequent clinical career. We also investigated whether the interpretation of therapeutic data differs as between students in the 4th year of medical school, students of the 6th year and physicians. In a questionnaire, one result from the Helsinki Heart Study was presented in for different ways: (a) absolute risk reduction, (b) relative risk reduction, (c) NNT (number needed to treat) and (d) percentage of patients without a cardiovascular event during the monitoring period. To each group we assigned a different drug. We interviewed students in the 4th and 6th years of medical school, physicians during a continuing education course and physicians before and after an evidence-based medicine (EBM) course. They replied by stating which drug they would select for therapy. Evaluation of the questionnaire showed that the majority regarded the drug for relative risk reduction as the most effective. Physicians taking part in the EBM course analysed the statistical data better than the other groups interviewed, and their answers even improved significantly after the EBM course. There was no difference in the interpretation of therapeutic results between students and physicians who had not taken part in an EBM course. The study also found that in all the groups interviewed the presentation of the results of scientific studies has a marked influence on their interpretation. We conclude that interpretation of medical studies should form part of every student's and physicians's training, either in medical school or continuing education courses. On the other hand, the results of studies should be uniformly expressed in absolute figures, to make it easier to compare different results. PMID- 10719718 TI - [Does a psychosocial group intervention program alter the quality of life of cancer patients?]. AB - BACKGROUND: Structured, multicentre psychosocial support programmes are rare in Switzerland. The Swiss Cancer League has therefore designed a programme of this kind for cancer patients, and the effect on quality of life (QL) is investigated in the present study. METHODS: The programme consisted of 10 weekly 2-hour sessions and was carried out, on the basis of a manual, in 10 cities of German speaking Switzerland. QL was assessed by standardised tools (HAD, EORTC, QLQ-C30, SELT-M, LASA) before, immediately after, and 9 months post baseline. Patients' personal impressions of the programme were gathered by telephone. RESULTS: 134 patients (25 males, 109 females), mean age 51.9 years, took part in the programme. 50% had breast cancer and the other half a variety of tumours. 25 patients attended only the educative part of the programme, while of the remaining 109, 41% attended all 10 sessions, 50% 7-9 and 9% 4-6. 80-90% had a favourable impression of the individual sessions and their content. Significant QL changes in the desired direction were observed in regard to emotional functioning (Wilcoxon p = 0.011), anxiety (p = 0.0005), depression (p < 0.0001), general life orientation (p = 0.006) and spiritual QL (p = 0.012). More optimistic patients showed a more positive change in physical well-being (Kurskall-Wallis p = 0.0005) and QL in general (p = 0.0006). CONCLUSION: A psychosocial support programme is well received by cancer patients after adjuvant treatment and leads to favourable QL changes, although independent time effects cannot be ruled out. Similar programme should be considered more often in standard oncological treatment regimens. PMID- 10719719 TI - [Extra-intestinal salmonella infections in kidney transplant patients]. AB - Immunosuppression to prevent rejection of solid organ transplants is accompanied by an increased risk of infections. The most frequent diseases include cytomegalovirus as well as bacterial infections of the urinary tract and the lung. The rate of enteric salmonella infections is increased in transplant patients. We report on four cases of extraintestinal salmonellosis after kidney allotransplantation. PMID- 10719720 TI - [Spectrum of effectiveness of insulin-like growth factors (IGF)]. AB - The insulin-like growth factors (IGF) I and II mediate endocrine and auto /paracrine effects of growth hormone (GH) on cell growth, cell differentiation and tissue repair. Despite their insulin-like actions in vitro, they do not play a significant role in the acute regulation of substrate metabolism, as compared to insulin and growth hormone. In contrast, administration of IGF I in vivo causes a broad spectrum of effects: it increases insulin sensitivity, improves the lipid profile, supports protein anabolism, enhances kidney function and bone turnover, and exerts beneficial effects on the cardiovascular system. Understanding of the mechanisms underlying these effects provides the basis for potential therapeutic use of IGF I. PMID- 10719721 TI - [[Booster vaccine injection]. PMID- 10719722 TI - Buerger's disease. PMID- 10719723 TI - Genomic alterations (LOH, MI) on chromosome 17q21-23 and prognosis of sporadic colorectal cancer. AB - Genomic alterations at the long arm of chromosome 17, and in particular at the nm23 locus, are still controversial in colorectal cancer (CRC). Our aim was to investigate the possible relationship of loss of heterozygosity (LOH) and microsatellite instability (MI), at 4 microsatellite loci spanning the 17q21-23 region, to the risk of liver metastasis and nm23 protein expression. Genomic DNA extracted from 58 primary and 54 liver secondary formalin-fixed and paraffin embedded CRCs was obtained from 82 patients. A fluorescent PCR coupled with an automated DNA sequencer was applied. Increasing fraction of loci showing LOH was positively associated with risk of liver metastases (logrank test for trend, p = 0.005); this remained independent after adjusting to T-stage (Cox regression, p = 0.022), N-stage (p = 0.007), or Dukes' stage (p = 0.012). Conversely, increasing frequency of MI was associated with a reduced risk of liver metastases in Dukes' B tumours (logrank test for trend, p = 0.032). When comparing 30 primary and matched liver secondary lesions, we found concordant genomic alteration in 72% (NME1) to 43% (D17S579). Finally, we observed a trend in association between the proportion of loci with LOH and nm23 positivity (chi2 test for trend, p = 0.024). Our findings suggest that genomic alterations in the 17q21-23 region may affect prognosis of CRC as well as regulation of the nm23 protein expression via an unknown underlying mechanism. PMID- 10719724 TI - Molecular detection of blood-borne epithelial cells in colorectal cancer patients and in patients with benign bowel disease. AB - In colorectal cancer (CRC), a proportion of patients with early stage disease still die of metastatic or recurrent disease within 5 years of "curative" resection. Detection of carcinoma cells in the peripheral circulation at presentation may identify a subgroup of patients with micro-metastatic disease who may benefit from adjuvant chemotherapy or radiotherapy. Our aim was to determine the presence and clinical significance of colon carcinoma cells in peripheral blood at the time of surgery. Preoperative peripheral blood samples were collected from 94 patients with CRC and 64 patients undergoing bowel resection for benign conditions (adenoma, diverticular disease or Crohn's colitis). Blood was also obtained from 20 normal donors not undergoing bowel surgery. Immunomagnetic beads were used to isolate epithelial cells followed by reverse transcription-polymerase chain reaction (RT-PCR) analysis of expression of cytokeratin (CK) 19, CK 20, mucin (MUC) 1 and MUC 2. Nineteen of 94 (20%) CRC patients were positive for epithelial cells in preoperative blood, including 6 with early stage disease. Kaplan-Meier survival analysis showed that detection of epithelial cells in preoperative blood was associated with reduced disease-free and overall survival (log-rank test, p = 0.0001). Surprisingly, circulating epithelial cells were detected in 3/30 (10%) patients resected for adenoma, and in 4/34 (12%) patients resected for benign inflammatory conditions, suggesting that cells from nonmalignant colonic epithelium may also gain entry into the bloodstream in the presence of bowel pathology. All 20 normal control bloods were negative for epithelial cells. PMID- 10719725 TI - Prognostic impact of p21/waf1/cip1 in colorectal cancer. AB - In addition to the tumor suppressor gene p53, Cyclin Dependent Kinases (CDK) are well known to influence the cell cycle in normal human tissues and various neoplasias as well. The purpose of our present study was to evaluate the expression of the CDK-inhibitor p21/waf1/cip1 in colorectal cancer with special emphasis on the prognostic impact. Between 1985 and 1991, 294 patients (median age, 65 years) underwent surgical operative therapy for colorectal cancer. Formalin-fixed and paraffin-embedded tumor specimens were investigated. For immunohistochemistry the Catalysed Reporter Deposition (CARD) technique was performed. The survival probability was calculated and possible prognostic risk factors were tested using multivariate analysis. The p21/ waf1/cip1 staining pattern was positive in 197 (67%) specimens and negative in 97 (33%) samples. No significant correlation could been calculated between p21/waf1/cip1 expression and other variables such as age, sex, WHO-Classification, localisation, grading, TNM-classification or UICC-stage. Patients with a positive staining reaction had a significantly better survival (p < 0.0052). Moreover, p21/waf1/cip1 was shown to be an independent prognostic parameter by multivariate analysis (p < 0.022). In contrast with these findings, the p53 tumor status had no impact on survival. P21/ waf1/cip1 appears to be an independent prognostic parameter in colorectal cancer and is associated with a favorable survival. This feature may be related to a cell cycle arrest in the G1 phase induced by p21/waf1/cip1, resulting in lower tumor cell proliferative activity. PMID- 10719726 TI - Amplotyping of microdissected, methanol-fixed lung carcinoma by arbitrarily primed polymerase chain reaction. AB - The arbitrarily primed polymerase chain reaction (AP-PCR) was used to detect somatic genetic alterations in lung carcinomas. DNA fingerprints generated by a single arbitrary primer were compared between normal and tumor tissues of the same individuals. We adapted the technique to the use of tissue fixed with methanol, which allowed the analysis of small areas of tissue by microdissection. This improvement of the fingerprinting technique permitted the study of tumors at early stages of progression. Loss of sequences from chromosome 7 was detected in 41.7% of adenocarcinomas and from chromosome 22 in 84.6% of small-cell carcinomas. Gains of sequences from chromosomes 1, 8 and 13 were detected in more than 40% of adenocarcinomas and in chromosome 2 in 63.3% of squamous-cell carcinomas. Our results indicate that allelic imbalances at these chromosomal regions are common genetic abnormalities in lung carcinomas. Loss of sequences from chromosome 22q13.3, found in 11 of 13 small-cell carcinomas, were confirmed by microsatellite PCR analysis. We show that the use of our improved AP-PCR fingerprinting permits the detection of both losses and gains of novel chromosomal regions early during lung cancer development. Our results indicate that early-stage tumors tend to have more allelic imbalances than relatively advanced tumors, suggesting a high tumor genetic heterogeneity in the early stages of lung tumor progression. PMID- 10719727 TI - Cell cycle proteins do not predict outcome in grade I infiltrating ductal carcinoma of the breast. AB - Infiltrating ductal mammary carcinomas are histologically graded according to their extent of differentiation. Well-differentiated, grade I, tumours have low proliferative activity, usually form tubules and exhibit little nuclear pleomorphism. Despite an apparently reassuring morphology, 10-15% of grade I ductal carcinomas metastasize, albeit after a prolonged period. Recent evidence supports the view that evolution to higher grade malignancies occurs rarely and that grade I tumours are biologically distinct from grade III tumours. We have examined a series of 148 grade I ductal carcinomas in order to ascertain whether information about the level of expression of cyclin D1, p27, p53, oestrogen receptor status (ER) or proliferative activity could be used to identify those patients with a poor outcome. The majority of tumours expressed high levels of cyclin D1, p27 and ER, low levels of p53 and had low Ki-67 expression and mitotic counts. Cyclin D1, p27 and ER expression were all significantly correlated with each other but not with p53 (cyclin D1 correlation with ER, p = 0.01; cyclin D1 correlation with p27 and ER correlation with p27 both p < 0.0001). Cyclin D1 and ER were also both correlated with Ki-67 (p = 0.01 and p < 0.0001) but not with mitotic count. Our results suggest that cyclin D1, ER and p27 are all markers of well-differentiated tumours and that their detection is related to proliferative activity in a manner reflecting their functional role within the normal cell cycle. However, none of the proteins or markers of proliferative activity were sensitive enough to predict which patients were likely to have a poor outcome. PMID- 10719728 TI - A prospective trial of midwest breast cancer patients: a p53 gene mutation is the most important predictor of adverse outcome. AB - Several retrospective studies have suggested p53 gene mutation as an adverse prognostic indicator in breast cancer patients, based on a selective growth advantage of p53 mutant cancer cells and their presumed resistance to current adjuvant therapy regimens. A cohort of 90 Caucasian midwestern breast cancer patients was analyzed prospectively (60 months of follow-up) with a rigorous mutation detection methodology. The presence of a p53 gene mutation was the single most adverse prognostic indicator for recurrence (p = 0.0032) and death (p = 0.0001), and was associated with poor response to both adjuvant (p = 0.0001) and palliative (p = 0.006) therapy. Analysis of the p53 gene with appropriate mutation detection methodology markedly improves the prediction of early recurrence, treatment failure, and death in breast cancer patients. PMID- 10719729 TI - Correlation of cyclin D1 MRNA levels with clinico-pathological parameters and clinical outcome in human breast carcinomas. AB - In order to evaluate the prognostic significance of cyclin D1 mRNA expression in mammary neoplasia, its levels were measured in 97 breast cancers by reverse transcription-polymerase chain reaction (PCR) using fluorescent primer and standard RNA along with estrogen receptor (ER). The median value of cyclin D1 mRNA was 1.60 amol/microg RNA (range, 0.01 to 5.63 amol/microg RNA). ER mRNA was detectable in 70 breast cancer samples (72.2%) and cyclin D1 mRNA levels were significantly higher in ER mRNA-positive than in ER mRNA-negative tumors (p = 0.009). Furthermore, cyclin D1 mRNA levels were significantly (p = 0.001) lower in patients who experienced a recurrence during the follow-up period (mean of 40.8 months, median of 39 months) compared with those with no evidence of recurrent disease (mean of 49.2 months, median of 48 months), and in those who died from disease (mean follow-up period of 30.5 months, median of 26 months) than in the survivors (mean of 50.5 months and median of 48 months) (p = 0.005). Setting the median value (= 1.60 amol/microg RNA) as the cutoff point, expression was significantly associated with relapse-free survival (p = 0.002). Similarly, a significant correlation was observed between the cyclin D1 mRNA level and overall survival (p = 0.015). The expression was found to be an independent factor for predicting relapse-free survival using multivariate analysis. PMID- 10719730 TI - Predictive genetic testing for hereditary non-polyposis colorectal cancer: uptake and long-term satisfaction. AB - The aim of this prospective study was to assess the uptake of predictive genetic testing for hereditary non-polyposis colorectal cancer (HNPCC) and its associations with sociodemographic and other factors, and long-term satisfaction with taking the test. The test was offered to all high-risk members (n = 446) of 36 Finnish HNPCC families in which the mutation was known. The procedure comprised an educational counselling session, a period for reflection, and a test disclosure session. Data were collected by questionnaires sent before the educational counselling and 1 month and 1 year after the test disclosure. Of those eligible, 85% (n = 381) completed the first questionnaire study. Non participation was more common among men living alone who had not participated in the clinical cancer surveillance programme. Of the 347 subjects who attended counselling, 334 (75% of all subjects) were actually tested. After logistic regression analysis, the only significant factor predicting test acceptance proved to be employment status: those employed were more likely than others to accept the test (odds ratio = 2.25; 95% confidence intervals, 1.09 to 4.6 1). At follow-up, over 90% of the subjects were fully satisfied with the decision to take the test. In conclusion, acceptance of the test was considerably higher than in previously reported studies. We attribute this to our careful face-to-face individualized counselling, our health care system, and to attitudes of the Finnish population, which are generally favourable towards health care and disease prevention. PMID- 10719731 TI - p53 and vascular-endothelial-growth-factor (VEGF) expression predicts outcome in 833 patients with primary breast carcinoma. AB - The angiogenic factor vascular endothelial growth factor (VEGF) predicts outcome in primary breast carcinoma. Alteration of the p53 gene causes down-regulation of the expression of thrombospondin-1, a natural inhibitor of angiogenesis. This study was conducted to investigate the association between mutant p53 protein and VEGF expression, and the prognostic value of these factors. VEGF165 and p53 protein were measured in tumour cytosols by enzyme immunoassays. Recurrence-free survival (RFS) and overall survival (OS) were estimated in 833 consecutive patients, 485 node-negative (NNBC) and 348 node-positive (NPBC) with primary invasive breast cancer. A significant association was found between mutant p53 protein and VEGF expression. Univariate analysis showed both p53 and VEGF to be significant predictors of survival. Similar correlation was seen when p53 was combined with VEGF. Univariate analysis of NNBC showed significant prognostic value of p53 for OS, also when combined with VEGF expression; for NPBC, significant reductions in RFS and OS were seen for p53-positive patients, and these findings were enhanced when combined with VEGF, also in the sub-group receiving adjuvant endocrine treatment. Multivariate analysis showed both p53 and VEGF as independent predictors of OS in all groups. When the 2 factors were combined, an increased relative risk of 2.7 was seen for OS in the group with both p53 positivity and high VEGF content, as compared with 1.7 in the group with one risk factor. The results suggest an association between loss of wt-p53 and increased VEGF expression, indicating that angiogenic activity may depend, at least partly, on altered p53-protein function. Combination of these 2 biological markers appears to give additional predictive information of survival. A high risk group of patients was associated with p53 positivity and higher VEGF content. PMID- 10719732 TI - High telomerase activity and high HTRT mRNA expression differentiate pure myxoid and myxoid/round-cell liposarcomas. AB - Molecular markers characterizing the transition of a myxoid to a more round-cell liposarcoma have not been described. To examine whether telomerase activity, hTRT and hTR mRNA expression were associated with tumor progression in myxoid liposarcoma, we investigated a total of 28 myxoid liposarcomas (13 pure myxoid tumors, 14 mixed-type tumors, and 1 pure round-cell variant) from 19 patients. Telomerase activity was detected by using the fluorescent PCR-based TRAP-assay. Expression of hTRT and hTR mRNAs was determined by the semi-quantitative RT-PCR. On the basis of only one tumor sample per patient, telomerase activity was found in 9 of 9 myxoid/round-cell liposarcomas and in 3 of 10 pure myxoid tumors. Elevated hTRT expression was found in 13 of 17 liposarcomas. All telomerase positive tumors showed hTRT expression, whereas there were 3 cases showing hTRT expression without telomerase activity. HTR mRNA expression was elevated in all 19 liposarcomas. Thus, only the levels of telomerase activity and of hTRT mRNA expression differentiated pure myxoid liposarcoma and myxoid/round-cell liposarcoma (p < 0.003 and p = 0.029, respectively). We believe that high levels of telomerase activity and of hTRT expression are associated with tumor progression from low-grade pure myxoid to higher-grade malignant round-cell liposarcoma, and may consequently represent a useful prognostic marker for this histological sub-type of soft-tissue tumors. PMID- 10719733 TI - Prognosis of breast-carcinoma lymphagenesis evaluated by immunohistochemical investigation of vascular-endothelial-growth-factor receptor 3. AB - Very few studies have yet addressed the question of the existence and role of lymphagenesis in tumor growth; it is generally overshadowed by the greater emphasis placed on the blood vascular system. Monoclonal antibodies against vascular endothelial-growth-factor receptor 3 (VEGFR3) have been shown to provide a specific antigenic marker for lymphatic endothelium. By comparison with the microvascular count (MVC), we investigated the prognostic value of the microlymphatic count (MLC) in a series of 60 cases of 2-cm-diameter breast carcinomas. The mean value of MVC was 72.5 and of MLC, 40.5. There was no quantitative correlation between these 2 parameters. The MVC but not the MLC had a prognostic value in overall survival. Neither the MLC nor the MVC had any correlation with axillary-lymph-node invasion. PMID- 10719734 TI - Tumor microvessel density and prognosis in node-negative breast cancer. AB - Microvessel density (MVD) of breast cancer is widely regarded as a prognostic factor, but results from studies on the most important case series have produced conflicting results. The present study was performed with confirmatory intent to define the prognostic relevance of MVD on a series of 378 node-negative-breast cancer patients, much larger than any other series previously analyzed. Microvessels were stained using Factor-VIII antibody and an immunoperoxidase reaction. MVD was determined independently by 2 observers according to Weidner's methods. In parallel, cell proliferation was evaluated as S-phase fraction and determined according to the 3H-thymidine-labeling index method (TLI). Estrogen and progesterone receptors were quantitatively assessed using the dextran charcoal technique. Tumor MVD varied greatly from tumor to tumor (2 to 232 MV/mm2) and was unrelated to patient age and menopausal status, or to tumor size, histology and steroid-receptor status. A significant (p = 0.004) but weak inverse correlation (rs = -0.188) was observed with cell proliferation. Univariate analysis using 40 MV/mm2 as cut-off showed an inverse relation with 5-year relapse-free survival (82% vs. 71%, p = 0.018). This finding was limited to very small tumors, slowly proliferating tumors and ER-negative tumors. Multivariate analysis identified tumor size and TLI, but not MVD, menopausal status or ER as independent prognostic factors. PMID- 10719735 TI - Prognostic significance of serum p53 antibodies in patients with limited-stage small cell lung cancer. AB - p53 tumour suppressor gene alterations are one of the most frequent genetic events in lung cancer. A subset of patients with p53 mutation and cancer exhibited circulating serum anti-p53 self-antibodies (p53-Ab). The prevalence of these antibodies in lung cancer is currently being analysed in a multicentric study. In a group of homogeneous SCLC patients, p53-Ab were detected in 20/97 (20.6%) individuals. In this group of patients, Cox's multivariate analysis identified disease extent (p = 0.022), WHO initial performance status greater than 0 (p = 0.005), and the absence of a complete response after 6 months of treatment (p < 0.0001) as independent prognostic variables, with p53-Ab being of borderline significance (p = 0.051). In the subset of limited-stage SCLC patients, Cox's multivariate analysis found p53-Ab (p = 0.033), WHO initial performance status greater than 0 (p = 0.028), and absence of a complete response (p < 0.001) to be independent prognostic variables. Thus, actuarial analysis showed that patients with limited-stage SCLC and p53-Ab had a median survival time of 10 months, whereas limited-stage SCLC patients without p53-Ab had a 17 month median survival time (p = 0.014).Therefore, serum assay of p53-Ab could help to identify a population of SCLC patients with an especially poor prognosis. This population could represent patients with tumours harboring aggressive p53 mutations. PMID- 10719736 TI - Microsatellite instability in colorectal-cancer patients with suspected genetic predisposition. AB - Hereditary non-polyposis colorectal cancer (HNPCC) is a dominantly inherited syndrome linked to DNA-mismatch-repair (MMR) gene defects, which also account for microsatellite instability (MSI) in tumour tissues. Diagnosis is based mainly on family history, according to widely accepted criteria (Amsterdam Criteria: AC). Aim of this work was to assess MSI in colorectal-cancer patients with suspected genetic predisposition, and to verify whether MSI represents a tool to manage MMR gene (hMSH2 and hMLH1) mutation analysis. We investigated 13 microsatellites (including the 5 NCI/ICG-HNPCC markers) in 45 patients with suspected hereditary predisposition (including 16 subjects from HNPCC families fulfilling the AC). We found MSI-H (high frequency of instability, i.e., in > or =30% of the markers) in 85% of the HNPCC patients and in 16% of the non-HNPCC subjects. The 5 NCI/ICG HNPCC microsatellites proved to be the most effective in detecting MSI, being mononucleotide repeats the most unstable markers. We investigated the association between hMSH2- and hMLH1 gene mutations and MSI. Our results indicate that AC are highly predictive both of tumour instability and of MMR-gene mutations. Therefore, as the most likely mutation carriers, HNPCC subjects might be directly analyzed for gene mutations, while to test for MSI in selected non-HNPCC patients and to further investigate MMR genes in MSI-H cases, appears to be a cost effective way to identify subjects, other than those from kindred fulfilling AC, who might benefit from genetic testing. PMID- 10719737 TI - Distinct prognostic values of p53 mutations and loss of estrogen receptor and their cumulative effect in primary breast cancers. AB - A total of 76 primary breast cancers were screened for p53 mutations using the yeast p53 functional assay, and the mutations were determined by DNA sequencing. Clonal mutations of p53 were detected in 30 tumors (39%). Immunohistochemical staining for nuclear p53 accumulation performed on the yeast assay-positive cases clearly differentiated missense mutations in the DNA binding domain (contact mutant; 17 cases) as positive stain and nonsense-type mutations or missense mutations that may affect 3D-structure of p53 protein (structural mutant; 13 cases) as negative stain. Enzyme immunoassay revealed loss of estrogen receptor in 36 tumors (50%). Prognostic values of p53 mutation and loss of estrogen receptor were evaluated after a median follow-up period of 44 months. p53 mutations were associated with a short overall survival (log rank test, p = 0.0319), whereas it was not related to disease-free (recurrence-free) survival. Contact mutants were associated with slightly shorter survival compared with structural mutants. Inversely, loss of estrogen receptor was associated with early recurrence (p = 0.0461) but not with short overall survival. The patients with tumors harboring both p53 mutation and loss of estrogen receptor had the poorest outcome (p = 0.0019 and 0.0075 for overall and disease-free survivals, respectively), suggesting independent and additive effects of the 2 factors. The independent role of the 2 factors was confirmed by a multivariate analysis using the Cox proportional hazard model stratified according to clinical tumor stages. Although preliminary, due to the small number of patients studied and the relatively short follow-up time, our results suggest that p53 mutations and loss of estrogen receptor cooperatively affect the prognosis of primary breast cancer patients. PMID- 10719738 TI - Tumorigenic pathways in low-stage bladder cancer based on p53, MDM2 and p21 phenotypes. AB - Our aim was to determine whether the pattern of expression of the interrelated proteins p53, MDM2 and p21 could shed light on the etiopathogenic mechanisms of superficial bladder tumors. Protein expression was detected by immunohistochemistry (IHC) using monoclonal antibodies (MAbs) Pab 1801 for p53, IF2 for MDM2 and EA10 for p21 on 269 newly diagnosed bladder tumors (214 pTa and 55 pT1; 93 grade I, 145 grade 2 and 31 grade 3). While no p21 immunoreactivity was found in normal urothelium, 85% of tumors were strongly p21-positive. MDM2 was overexpressed in 44% of tumors, almost all being also positive for p21. p53 was overexpressed in 20% of cases: 66% of p53-positive tumors were also MDM2 positive, compared with only 38% of p53-negative tumors. p53 mutations were studied by direct DNA sequencing in a subset of 24 high-grade tumors. Both MDM2 and p21 were less frequently expressed in p53 mutated tumors compared with tumors with a wild-type gene. Distinct phenotypes were correlated with the frequency of poorly differentiated (grade 3) tumors. The most common phenotypes were p21+/MDM2 /p53- and p21+/MDM2+/p53- observed in 38% and 29% of tumors, respectively. Grade 3 tumors were found in 4% and 8% of these 2 groups, in contrast with 30% frequency in p21+/p53+ tumors (p = 0.002) regardless of their MDM2 phenotype. Four of the 5 (80%) tumors that were p53-positive but negative for p21 were grade 3. Our data suggest that several tumorigenic pathways for superficial bladder tumors can be reflected by the expression pattern of these 3 proteins. PMID- 10719739 TI - Heartburn and reflux oesophagitis. PMID- 10719740 TI - Autoimmune disease overlaps and the liver: two for the price of one? PMID- 10719741 TI - Hepatitis E: an overview and recent advances in clinical and laboratory research. AB - Hepatitis E virus (HEV) is a non-enveloped RNA (7.5 kb) virus that is responsible for large epidemics of acute hepatitis and a proportion of sporadic hepatitis cases in southeast and central Asia, the Middle East, parts of Africa and Mexico. Hepatitis E virus infection spreads by the faecal-oral route (usually through contaminated water) and presents after an incubation period of 8-10 weeks with a clinical illness resembling other forms of acute viral hepatitis. Clinical attack rates are the highest among young adults. Asymptomatic and anicteric infections are known to occur. Chronic HEV infection is not observed. Although the mortality rate is usually low (0.07-0.6%), the illness may be particularly severe among pregnant women, with mortality rates reaching as high as 25%. Recent isolation of a swine virus resembling human HEV has opened the possibility of zoonotic HEV infection. Studies of pathogenetic events in humans and experimental animals reveal that viral excretion begins approximately 1 week prior to the onset of illness and persists for nearly 2 weks; viraemia can be detected during the late phase of the incubation period. Immunoglobulin M antibody to HEV (anti-HEV) appears early during clinical illness but disappears rapidly over a few months. Immunoglobulin G anti-HEV appears a few days later and persists for at least a few years. There is no specific treatment available for hepatitis E virus infection. Ensuring a clean drinking water supply remains the best preventive strategy. Recombinant vaccines are being developed that may be particularly useful for travellers to disease-endemic areas and for pregnant women. PMID- 10719742 TI - Percutaneous endoscopic gastrostomy: a review of indications, complications and outcome. AB - Percutaneous endoscopic gastrostomy (PEG) was first described in 1980 as an effective method of feeding via the stomach in situations where oral intake is not possible. Its simplicity has led to its potential use in areas of dubious clinical benefit. Our unit has faced a major increase in referrals for PEG insertion over the last 2 years. For this reason we decided to audit our PEG insertion procedures with regard to indications, complications, outcome and follow up. We studied 168 patients who had an initial PEG insertion during the period 1 February 1996-31 January 1998. The medical records of these patients were reviewed with regard to the procedure, antibiotic use and complications. All patients (or carers or next of kin) were contacted by telephone to provide details regarding late complications and follow up. There were 87 females and 81 males (aged 16-98 years, median age 70 years). At 2 years, 67% were alive. The most frequent indication for PEG insertion was a neurological condition, the commonest being stroke. Most patients received either ticarcillin/clavulanic acid or cephazolin antibiotic prophylaxis before and after the procedure. In six patients (3.6%) infection at the PEG site required intravenous antibiotics. Four of these six patients did not have antibiotic prophylaxis. Only two deaths could be directly related to the procedure. Three died within 7 days of the procedure due to unrelated medical complications. Sixteen patients died within 1 month, the majority of these patients did not leave hospital. One-fifth of the patients (35/168) had their PEG removed due to the re-establishment of oral feeding, with median time of use, 4.3 months. It is a safe, effective feeding method in the elderly, but experience with case selection, the procedure and careful follow up remain essential. The use of prophylactic antibiotics resulted in few significant infections of the PEG site. Up to one-fifth of patients will require their PEG only for a short term. PMID- 10719743 TI - Dilatation of benign oesophageal strictures: 10 years' experience with Celestin dilators. AB - BACKGROUND: Endoscopic dilatation is the first line of treatment for benign oesophageal strictures. There are limited data available on the use of Celestin dilators. METHODS: The efficacy and safety of Celestin dilators was evaluated retrospectively in 61 patients with benign oesophageal strictures. Three hundred and ninety-three dilatations using Celestin dilators were performed over a period of 10 years on an outpatient basis in patients with corrosive, peptic and other causes of benign oesophageal strictures. RESULTS: Initial success was achieved in all patients in the peptic and miscellaneous group and in 91% in the corrosive group of patients. Patients with corrosive strictures required significantly more dilatations for initial success compared with the peptic group (mean 5.82 vs 1.62 P < 0.1). At 6 months follow up after the initial success, 29% of the patients had an excellent response, 56% a good response and 15% a fair response. No patient had a poor response. During the long-term follow up of 10 years, overall dilatation requirement decreased significantly. (72 vs 27 vs 14% of patients requiring dilatation at 1, 5 and 10 years P < 0.05). The dilatation requirement also decreased significantly within the groups (P < 0.05). Patients with corrosive stricture required more frequent dilatations on follow up compared with the other two groups. Complications in the form of oesophageal perforation occurred in only two patients. There was no mortality. CONCLUSION: Oesophageal dilatation with Celestin dilators is an effective and safe modality for managing patients with benign oesophageal strictures. PMID- 10719744 TI - Secondary oesophageal peristalsis in gastro-oesophageal reflux disease. AB - BACKGROUND AND AIMS: To evaluate the status of secondary oesophageal peristalsis in gastro-oesophageal reflux disease (GORD) and the effect of healing of oesophagitis on these abnormalities. METHODS: Twenty-one patients diagnosed with GORD and 10 control subjects in the same age group were studied. Primary peristalsis was elicited by 10 5 mL water boluses and secondary peristalsis by 10 20 mL boluses of air injected 15 cm above the lower oesophageal sphincter. RESULTS: The pattern of primary peristalsis was normal in a significantly lower number of patients compared with control subjects, six patients (28.6%) versus seven controls (70%), (P<0.05). Similarly, the number of subjects with a normal pattern of secondary peristalsis was also lower in the patient group (zero vs three; P<0.05). A normal primary peristaltic response occurred with 71 (33.8%) of the 210 water boluses in the patients and 73 (73%) of the 100 water boluses in the control subjects, respectively (P<0.001). A normal secondary peristaltic response was seen with 15 (7.1%) of 210 air boluses in patients and 32 (32%) of 100 air boluses in the control subjects (P<0.001). The amplitude of secondary peristaltic waves and the duration of contraction (mean+/-SEM) were significantly lower in patients compared with the control subjects (43.5+/-4.7 vs 89.0+/-13.1 and 3.4+/-0.8 vs 3.9+/-0.3, respectively; P=<0.05). In the 13 patients in whom repeat evaluation was performed after healing of oesophagitis, there was no significant change in the number of patients with normal peristaltic response, number of normal responses to air and water boluses, or amplitude, duration and velocity of peristalsis. CONCLUSION: Significant abnormalities of secondary oesophageal peristalsis occur in patients with GORD and these are not reversed by healing of oesophagitis. PMID- 10719745 TI - Heartburn: community perceptions. AB - BACKGROUND AND AIMS: To determine the prevalence of heartburn in the Australian community, and document factors precipitating it and medications used in treatment. METHODS: Telephone interviews with 1200 individuals aged 18 years or more were conducted one weekend in 1996. Each respondent was asked four questions about heartburn, its severity, factors causing it and current therapy. RESULTS: Fifty-six per cent of respondents reported that they had suffered from heartburn at some time in the past and 37% had symptoms at least once every 4-6 months. The frequency of heartburn increased with age and was more common in men (40.7%) than women (32.6%). There was no difference in frequency between city and rural dwellers, or between white- and blue-collar workers. Almost half the individuals experienced mild pain or discomfort, one-third had moderate discomfort and 15% reported severe pain or discomfort. Women were more likely to report greater problems than men. Aggravating factors included spicy foods, greasy/rich foods, stress, alcohol, overeating, pregnancy, smoking, food allergy and coffee. More than half the respondents relied on antacids to control symptoms, 20% used prescription medications and a similar number did not use any medication. CONCLUSION: Heartburn is common in the Australian community and sufferers attribute symptoms to various lifestyle events, including diet and stress. Antacid usage is the commonest mode of therapy. PMID- 10719746 TI - Evaluation of autonomic nervous function in patients with essential hypertension complicated with peptic ulcer. AB - BACKGROUND AND AIMS: The relationship between peptic ulcer, autonomic activity and the incidence of Helicobacter pylori infection in untreated hypertensive patients complicated with peptic ulcer were evaluated. METHODS: Ten hypertensive patients with peptic ulcer (HT-PU group), 15 untreated essential hypertensive patients without peptic ulcer (HT group) and 10 normal subjects (N group) were enrolled, and a power spectral analysis was performed in each subject. A biopsy urease test was used to detect infection by H. pylori. RESULTS: No significant differences were observed in the values of mean low-frequency (LF) power between the three groups. However, the mean high-frequency (HF) power in the HT-PU group was significantly greater than those of the HT and N groups (P<0.01). The mean LF/HF ratios in the HT-PU and HT groups were significantly greater than that of the N group (P<0.01). With respect to H. pylori infection, no significant differences between the three groups were observed. Sympathetic activity (LF power) was increased in the HT and HT-PU groups. Furthermore, parasympathetic activity (HF power) was increased in the HT-PU group. CONCLUSIONS: These findings suggest the participation of increased parasympathetic activity in peptic ulcer patients. Therefore, it is suggested that new techniques, such as spectral analysis of heart rate variability, as used in this study, will clarify the relationship between peptic ulcer and autonomic nervous function. PMID- 10719747 TI - Augmentative effects of L-cysteine and methylmethionine sulfonium chloride on mucin secretion in rabbit gastric mucous cells. AB - BACKGROUND: Our previous study showed that L-cysteine (Cys) and methylmethionine sulfonium chloride (MMSC) inhibited ethanol-induced gastric mucosal damage and increased the amount of surface mucin in rats. This study examined whether Cys and MMSC augmented mucin secretion and changed distribution of mucin vesicles ultrastructurally in mucous cells by using primary cultured mucous cells from rabbit glandular stomach. Changes in intracellular cyclic adenosine 3',5' monophosphate (cAMP) and in levels of cytosolic free Ca2+ were investigated by treatment with Cys and MMSC. METHODS: Mucin content was measured by an enzyme linked lectin assay. Transmission electron micrography was used to examine ultrastructural distribution of mucin granules. The amount of cAMP or levels of free Ca2+ were measured by enzyme immunoassay or by fura-2. 16,16-Dimethyl prostaglandin E2 (dmPGE2) or ATP was used as the positive control. RESULTS: L Cysteine and MMSC increased mucin secretion and decreased cellular mucin content. The same was noted for dmPGE2. Accelerated mucin granule movements toward the plasma membrane were shown by these agents. Intracellular cAMP increased with exposure to dmPGE2 for 20 min, while neither Cys nor MMSC increased cAMP. No increase in cytosolic free Ca2+ levels occurred after treatment with Cys or MMSC, but an increase was induced 10 s after the addition of ATP. CONCLUSIONS: The present findings indicate that the increase in mucin secretion by Cys and MMSC was not mediated through the cAMP or Ca2+ signal transduction pathway, but might occur through non-receptor-mediated mechanisms. PMID- 10719748 TI - Genetically distinct strains of Candida albicans with elevated secretory proteinase production are associated with diarrhoea in hospitalized children. AB - BACKGROUND: Candida albicans has been implicated as the aetiological agent in a significant percentage of children with diarrhoea. The virulence properties of C. albicans strains associated with acute and chronic diarrhoea in hospitalized children were investigated. METHODS: The genotypic relationships between the isolates were determined using restriction enzyme analysis and hybridization with a C. albicans-specific DNA probe, 27A. RESULTS AND CONCLUSION: In patients with acute and chronic diarrhoea, there is evidence for selection of specific, genetically distinct strains of C. albicans. Higher levels of secretory Candida acid proteinase produced by isolates from patients with acute diarrhoea may account for the more severe symptoms. However, the lower adherence of these isolates may predispose to the rapid (within 2 to 4 days) resolution of the condition. In patients with chronic diarrhoea the lower levels of proteinase produced correlate with the less severe symptoms, while the increased adherence may account for the persistence of the infection. PMID- 10719749 TI - Characterization of antibody responses against rectal mucosa-associated bacterial flora in patients with ulcerative colitis. AB - BACKGROUND: Previous reports on faecal microflora have demonstrated that the total number of aerobes and coliforms was increased in patients with ulcerative colitis (UC). Based on the hypothesis that the pathogenesis of UC may be closely associated with the mucosal microflora, we investigated alterations in the mucosa associated microflora of UC patients. METHODS AND RESULTS: The bacterial counts for both aerobes and anaerobes increased in UC patients. In particular, we detected the highest bacterial counts of Bacteroides vulgatus and these bacteria were isolated most frequently. In addition, we also investigated the serum antibody responses against the bacteria isolated from the affected mucosa by serum bacterial agglutination tests and immunoblotting. A high agglutination titre against B. vulgatus, Bacteroides fragilis, and Clostridium ramosum was detected in most UC patients, and the percentage of positive immunoreactivity was much higher in UC patients than in healthy controls. From the results of the immunoblotting, a unique antigenic determinant of B. vulgatus (BV43-26), a 26-kDa protein from the outer membrane, was discovered. The serum immunoreactivity (immunoglobulin (Ig) G) against this 26-kDa protein was much higher in UC patients (53.8%) than in the control sera (9.1%). The serum immunoreactivity (IgG) against a 50-kDa protein isolated from the whole cell protein of Escherichia coli (EC48-1) was also higher in UC patients (29.2%) than in normal controls (6.3%). CONCLUSIONS: These results suggest that B. vulgatus and a specific antibody response directed against it may play an important role in the pathogenesis of UC. PMID- 10719750 TI - Increase in CD95 (Fas/APO-1)-positive CD4+ and CD8+ T cells in peripheral blood derived from patients with autoimmune hepatitis or chronic hepatitis C with autoimmune phenomena. AB - BACKGROUND: The expressions of CD95 (Fas/APO-1) and Bcl-2 are determinants of apoptosis in normal lymphocytes, and abnormalities in their expressions might contribute to the induction of autoimmunity. In this study, we examined the expressions of CD95 and Bcl-2 on freshly isolated T and B cells from patients with autoimmune hepatitis (AIH) or chronic hepatitis C associated with autoimmune phenomena (CH-C(AI)). METHODS: The CD95 and Bcl-2 expressions within CD4+ T, CD8+ T, and CD19+ B cell subsets were analysed by two-colour flow cytometry. RESULTS: The surface expression of CD95 was significantly high in both the CD4+ T and CD8+ T cell subsets derived from the patients with AIH and those with CH-C(AI), compared with expression in patients with CH-C and normal subjects. The increase in CD95 expression was associated with the phenotypic conversion of naive CD45RO- to primed CD45RO+ CD4+ T cells. Bcl-2 was detected in the vast majority of peripheral T and B cells. There was no significant difference in the percentage of Bcl-2-positive cells in the CD4+ T cell, CD8+ T cell and CD19+ B cell subsets among the patient groups and normal subjects. CONCLUSIONS: These results indicate that an increase in CD4+ T cells expressing CD45RO and CD95 marks an important subset of AIH and CH-C(AI) patients. These expanded CD95+ CD45RO+ primed T cells most likely reflect a continuous antigen-specific or non-specific activation of T lymphocytes, and/or the persistent presence of activated lymphocytes as a consequence of abnormalities in the peripheral deletion of activated lymphocytes. These persistently activated lymphocytes might play a role in the induction of autoimmunity in AIH and CH-C(AI). PMID- 10719752 TI - Hepatocyte growth factor/scatter factor enhances the thrombopoietin mRNA expression in rat hepatocytes and cirrhotic rat livers. AB - BACKGROUND: Although thrombopoietin (TPO) is mainly produced in the liver, the regulatory mechanism of TPO gene expression in hepatocytes remains unclear. The role of TPO in thrombocytopenia associated with liver cirrhosis has not been identified. METHODS: We examined the effects of various growth factors and cytokines on TPO mRNA expression in adult rat hepatocytes in primary cultures using a semiquantitative reverse transcription-polymerase chain reaction assay. RESULTS: Among them, only hepatocyte growth factor/scatter factor (HGF/SF) enhanced TPO mRNA expression; other growth factors (epidermal growth factor and transforming growth factor-beta) and cytokines (erythropoietin, granulocyte colony stimulating factor, granulocyte-macrophage-colony stimulating factor, interleukin (IL)-3, IL-6 and interferon-gamma) did not. Next, we examined TPO mRNA expression in the livers of rats with CCl4-induced cirrhosis, the effects of HGF/SF on hepatic TPO mRNA expression and peripheral platelet and bone marrow megakaryocyte counts in the cirrhotic rats. In the cirrhotic rats, both the peripheral platelet count and TPO mRNA expression in the livers were markedly decreased compared with those of the normal rats. The administration of HGF/SF to the cirrhotic rats stimulated TPO mRNA expression in the livers and resulted in significant increases of peripheral platelets and bone marrow megakaryocytes. CONCLUSIONS: These results suggest that HGF/SF is a possible regulatory factor for TPO gene expression and that HGF/SF increases platelet production through an enhancement of TPO mRNA expression in the livers of cirrhotic rats. PMID- 10719751 TI - Plasma human hepatocyte growth factor concentrations in patients with biliary obstruction. AB - BACKGROUND: It has been suggested that human hepatocyte growth factor (hHGF) maintains the growth and viability of hepatocytes and biliary epithelial cells. The purpose of this study was to determine plasma hHGF concentrations in patients with obstructive jaundice and to correlate these findings with clinical outcome. METHODS: The study included 22 patients who had biliary obstruction and underwent percutaneous transhepatic biliary drainage. The plasma concentrations of hHGF, standard liver function tests, daily bile flow and the half-life of serum total bilirubin were measured following the drainage. RESULTS: Plasma hHGF concentrations were significantly higher in patients with biliary obstruction compared with a control group (P<0.01). The plasma hHGF concentrations correlated with white cell count, prothrombin time and bilirubin half-life (P<0.05), but not with the values from other liver function tests. Seven patients who died within 3 months after biliary drainage had significantly higher concentrations of plasma hHGF than the 15 patients who survived for at least 3 months (P<0.05). The patients who experienced a poor outcome also had lower bile flows and prolonged bilirubin half-lives compared with the survivors (P<0.05). The plasma hHGF concentrations decreased significantly after biliary drainage in the survivors (P<0.01), but not in the patients with a poor outcome. CONCLUSIONS: These results suggest that systemic inflammation and the hepatic dysfunction caused by obstructive jaundice cause an increase in the plasma concentrations of hHGF. In addition, the plasma concentrations of hHGF may be a predictor of poor outcome in jaundiced patients. PMID- 10719753 TI - Hepatobiliary and pancreatic: malaise and jaundice. PMID- 10719754 TI - Gastrointestinal: choledochoduodenal fistula. PMID- 10719755 TI - Autoimmune cholangitis with features of autoimmune hepatitis: successful treatment with immunosuppressive agents and ursodeoxycholic acid. AB - We report a 42-year-old Chinese female with elevated serum levels of liver aminotransferases, alkaline phosphatase, gamma-glutamyl transpeptidase, cholesterol and immunoglobulin M. Serum antimitochondrial antibody was negative, but antinuclear antibody was strongly positive. Liver histology showed features of both autoimmune cholangitis and autoimmune hepatitis. Combination therapy with immunosuppressive (prednisone and azathioprine) and choleuretic agents (ursodeoxycholic acid) was given. Serum aminotransferases and biliary enzymes showed much improvement after treatment. A follow-up liver biopsy showed improvement of both hepatic necroinflammation and bile duct damage. Biliary enzymes rose after withdrawal of the immunosuppressive agents and declined again with reinstitution of prednisone. This case demonstrates that a combination of immunosuppressive agents and ursodeoxycholic acid may effectively treat patients with features of both autoimmune cholangitis and autoimmune hepatitis. PMID- 10719756 TI - Burkitt's lymphoma associated with Helicobacter pylori. AB - The association between Helicobacter pylori infection and low grade mucosa associated lymphoid tissue lymphoma is now widely accepted. In this report, we describe the concurrent development of Burkitt's lymphoma in the stomach of a 53 year-old male with perforated duodenal ulcer and positive H. pylori serology. The temporal relationship between these two events raises the possibility of a causal link between H. pylori infection and this lymphoproliferative disease. In describing this rare case of gastric Burkitt's lymphoma, we consider the evidence that supports this possibility. PMID- 10719757 TI - Ubiquitous overexpression of CuZn superoxide dismutase does not extend life span in mice. AB - Oxidative damage has been implicated in the aging process and in a number of degenerative diseases. To investigate the role of oxygen radicals in the aging process in mammals, the life spans of transgenic mice on a CD-1 background expressing increased levels of CuZn superoxide dismutase (CuZnSOD), the enzyme that metabolizes superoxide radicals, were determined. Homozygous transgenic mice with a two- to five-fold elevation of CuZnSOD in various tissues showed a slight reduction of life span, whereas hemizygous mice with a 15- to 3-fold increase of CuZnSOD showed no difference in life span from that of the nontransgenic littermate controls. The results suggest that constitutive and ubiquitous overexpression of CuZnSOD alone is not sufficient to extend the life spans of transgenic mice. PMID- 10719758 TI - Vitality index in survival modeling: how physiological aging influences mortality. AB - We investigated the relation of the age trajectory of physiological indicators of the average metabolic activity of organisms in a population to the age-specific population mortality rate. We show that a metabolic rate indicator (MRI) can be estimated using traditional physiological measures, such as homeostatic serum glucose concentration, vital capacity, and such. Estimates of the MRI were made from data collected in the Multiple Risk Factor Intervention Trial (MRFIT) study. The relation of the empirical mortality rate and MRI was also tested using MRFIT data. The age trajectory of MRI was evaluated using Swedish mortality data. The mortality results reproduce the "Strehler and Mildvan effect." The average rate of decline of MRI with age coincides with estimates predicted by Strehler using other methods. Possible extensions of the method are discussed. PMID- 10719759 TI - Soluble receptors and other substances that regulate proinflammatory cytokines in young and aging humans. AB - Relatively little is known about changes in soluble receptors and other agonists/antagonists that may regulate cytokine actions in aging humans. We have studied age-associated changes in human subjects of (a) the plasma levels of interleukin-1 soluble receptor (IL-1sRII), interleukin-1 receptor antagonist (IL 1ra), tumor necrosis factor soluble receptor-II(75kDa; TNFsRII), and interleukin 6 soluble receptor (IL-6sR) and (b) the ability of their blood mononuclear cells to produce those soluble factors spontaneously and after phytohemagglutinin (PHA) stimulation. Aging subjects (50-67 years) had significantly higher plasma levels of IL-1ra, significantly lower levels of TNFsRII and IL-6sR than young subjects (25-35 years), and no significant change in the level of IL-1sRIL There was less spontaneous output of IL-1ra and TNFsRII by peripheral blood mononuclear cells (PBMC) of aging than of young subjects, but equivalent output of both factors in response to PHA stimulation. Thus, the basal (homeostatic) output of those two factors declined with age, but the potential of the PBMC to produce the factors on stimulation did not. PHA stimulation of PBMC of either age group significantly inhibited the output of IL-6sR. These differences between the young adult and aging subjects, along with reported changes in the corresponding cytokines, presumably foreshadow changes that become more marked with further aging Therefore, immunological processes that depend on, or are modulated by, proinflammatory cytokines may differ between young and aged subjects as a consequence of the availability of regulatory soluble receptors and related agonists or antagonists. The results of this study highlight the need for further studies of the roles played by soluble receptors, and similar agonists/antagonists, in the immune responses of aged adults. PMID- 10719760 TI - Effect of age and testosterone on the vasopressin and aquaporin responses to dehydration in Fischer 344/Brown-Norway F1 rats. AB - To determine if the aging-associated decline in testosterone results in attenuated vasopressin (VP) responses to dehydration, testosterone implants were given to aged male Fischer 344Brown-Norway F1(F344BNF1) rats. Water deprivation caused comparable dehydration, increased plasma VP (pVP), and decreased posterior pituitary (PP) VP content in 4-, 15-, and 28-month-old rats. Dehydration increased VP mRNA content of supraoptic nuclei only at 4 months, whereas VP mRNA length was increased at both 4 and 15 months of age. The elevated pVP in the water-deprived aged rats indicates that even without an increase in VP mRNA content, PP VP storage was adequate to maintain elevated pVP. Dehydration increased aquaporin-2 content at 4, but not at 15 or 28 months of age, suggesting decreased renal responsiveness to VP. Testosterone replacement did not produce dehydration-induced increases in VP mRNA or aquaporin-2. Therefore, testosterone deficiency does not result in altered VP responses to dehydration in aged F344BNF1 rats. PMID- 10719761 TI - Left ventricular response to beta-adrenergic stimulation in aging rats. AB - The incidence of heart failure in the population increases steeply among older people. Overactivation of the sympathetic nervous system is associated to and responsible for worsening of heart failure. This study describes the influence of aging on short-term left ventricular (LV) adaptation to b-adrenergic stimulation in Wistar rats. In controls at 18 mo, interstitial fibrosis was increased with respect to 3-mo-old rats, whereas myocytes dimension and the messenger RNA (mRNA) abundance of atrial natriuretic peptide (ANP), a2(I) procollagen, transforming growth factor (TGF-b1, TGF-b3), and secreted protein, acidic and rich in cysteine (SPARC) were not different. To determine how aging affects LV adaptation to adrenergic stimulation, two groups of animals received isoproterenol (ISO, 1 mg/kg/d) for 3 days. There was no significant difference between young and older rats with respect to increase in LV weight, myocytes dimension, and mRNA abundance of all the genes considered, except a1(III) procollagen. These findings indicate that despite limited compensatory hypertrophy and higher fibrosis, LV from aged nonsenescent rats preserves the capacity to adapt to b-adrenergic stimulation through the upregulation of several genes encoding extracellular matrix-related proteins. PMID- 10719762 TI - Effect of long-term caloric restriction on GLUT4, phosphatidylinositol-3 kinase p85 subunit, and insulin receptor substrate-1 protein levels in rhesus monkey skeletal muscle. AB - Caloric restriction (CR), a reduction in calorie intake without malnutrition, improves insulin sensitivity in various species, including mice, rats, rhesus and cynomolgus monkeys, and humans. Skeletal muscle is quantitatively the most important tissue for blood glucose clearance. Therefore, we assessed the effect of 6 years of CR (30% reduction in calorie intake) in male rhesus monkeys (14-20 years old) on muscle expression of several proteins involved in insulin action. Whole body insulin sensitivity (assessed by Modified Minimal Model) was significantly increased in CR relative to Control monkeys. CR did not alter the expression of GLUT4 glucose transporter or phosphatidylinositol-3 kinase p85 subunit (PI3K). Insulin receptor substrate-1(IRS-1) abundance tended to be greater for CR compared to Control monkeys (p = .051), but correlational analysis revealed no association between IRS-1 and insulin sensitivity (r2 = .075, p = .271). These findings indicate that the CR-induced increase in insulin sensitivity in rhesus monkeys is unrelated to alterations in GLUT4, P13K, and IRS 1 abundance. PMID- 10719763 TI - The contribution of genetic influences to measures of lower-extremity function in older male twins. AB - Tests of balance, gait, and endurance were administered to 95 monozygotic (MZ) and 92 dizygotic (DZ), white male twins aged 68 to 79 years who had been born in the United States. Within-twin-pair correlations were calculated for each individual task and for an overall summary performance score. These were subjected to structural equation modeling to determine the contributions of genetic and environmental influences to individual differences in performance scores. MZ intraclass correlations were significant and greater than DZ correlations for the 8-foot walk and the repeated chair stands task, but not for the standing balance task. The heritability of the lower-extremity summary score was 57%, of which 39% was due to additive genetic effects and 18% due to nonadditive effects. In addition, we found that genetic influences contributed primarily to twin similarity in the poorest quartile of performance, whereas shared environmental influences contributed to twin similarity in the best quartile. PMID- 10719764 TI - Mortality oscillations induced by periodic starvation alter sex-mortality differentials in Mediterranean fruit flies. PMID- 10719765 TI - "Effects of food restriction on mechanical properties of the arterial system in adult and middle-aged rats". PMID- 10719766 TI - Complementary and alternative medicine use among elderly persons: one-year analysis of a Blue Shield Medicare supplement. AB - BACKGROUND: Large scale surveys in the United States and abroad suggest that 35 60% of adults have used some form of complementary/alternative medicine (CAM). However, no studies to date have focused on predictors and patterns of CAM use among elderly persons. METHODS: The population surveyed were Californians enrolled in a Medicare risk product that offers coverage for acupuncture and chiropractic care. Surveys were mailed to 1597 members in 1997 and responses received by 728 (51% response rate). Health risk assessment data were also obtained at baseline and 12-15 months following enrollment in the plan. Multiple logistic regression analyses were carried out to examine predictors of CAM use. RESULTS: Forty-one percent of seniors reported use of CAM. Herbs (24%), chiropractic (20%), massage (15%), and acupuncture (14%) were the most frequently cited therapies. CAM users tended to be younger, more educated, report either arthritis and/or depression/anxiety, not be hypertensive, engage in exercise, practice meditation, and make more frequent physician visits. Use of CAM was not associated with any observed changes in health status. Respondents also expressed considerable interest in receiving third-party coverage for CAM. Although 80% reported that they had received substantial benefit from their use of CAM, the majority (58%) did not discuss the use of these therapies with their medical doctor. CONCLUSIONS: Findings suggest that there is significant interest in and use of complementary/alternative medicine among elderly persons. These results suggest the importance of further research into the use and potential efficacy of these therapies within the senior population. PMID- 10719767 TI - Attentional demands and postural control: the effect of sensory context. AB - BACKGROUND: This study used a dual task design to examine the effect of sensory context on postural stability during the concurrent performance of an attentionally demanding cognitive task in young and older adults with and without a history of imbalance and falls. METHODS: A choice reaction time auditory task was used to produce changes in attention during quiet stance in six different sensory conditions that changed the availability of accurate visual and somatosensory cues for postural control. Postural stability was quantified by using forceplate measures of center of pressure in 18 young adults, 18 healthy older adults, and 18 older adults with balance impairments and a history of recent falls. Reaction time and accuracy of verbal response to the auditory task were quantified by using a repeated measures analysis of variance. RESULTS: In young adults the auditory task did not affect postural stability in any of the sensory conditions. However, in the older adults the effect of the auditory task depended on sensory context. For healthy older adults, the addition of an auditory tone task significantly affected sway only when both visual and somatosensory cues for postural control were removed. In the balance-impaired older adults, the addition of the auditory task significantly affected postural stability in all sensory conditions. In addition, as sensory conditions became more difficult, older adults who had been able to maintain stability in a single task context lost balance when performing a secondary task. CONCLUSION: Results suggest that with aging, attentional demands for postural control increase as sensory information decreases. In addition, the inability to allocate sufficient attention to postural control under multitask conditions may be a contributing factor to imbalance and falls in some older adults. PMID- 10719768 TI - New approach to risk determination: development of risk profile for new falls among community-dwelling older people by use of a Genetic Algorithm Neural Network (GANN). AB - BACKGROUND: Falls risk in older people is multifactorial and complex. There is uncertainty about the importance of specific risk factors. Genetic algorithm neural networks (GANNs) can examine all available data and select the best nonlinear combination of variables for predicting falls. The aim of this work was to develop a risk profile for operationally defined new falls in a random sample of older people by use of a GANN approach. METHODS: A random sample of 1042 community-dwelling people aged 65 and older, living in Nottingham, England, were interviewed at baseline (1985) and survivors reinterviewed at a 4-year follow-up (n = 690). The at-risk group (n = 435) was defined as those survivors who had not fallen in the year before the baseline interview. A GANN was used to examine all available attributes and, from these, to select the best nonlinear combination of variables that predicted those people who fell 4 years later. RESULTS: The GANN selected a combination of 16 from a potential 253 variables and correctly predicted 35/114 new fallers (sensitivity = 31%; positive predictive value = 57%) and 295/321 nonfallers (specificity = 92%; negative predictive value = 79%); total correct = 76%. The variables selected by the GANN related to personal health, opportunity, and personal circumstances. CONCLUSIONS: This study demonstrates the capacity of GANNs to examine all available data and then to identify the best 16 variables for predicting falls. The risk profile complements risk factors in the current literature identified by use of standard and conventional statistical methods. Additional data about environmental factors might enhance the sensitivity of the GANN approach and help identify those older people who are at risk of falling. PMID- 10719769 TI - Exploratory study of incident vehicle crashes among older drivers. AB - BACKGROUND: As the number of older adult drivers increases, distinguishing safe from unsafe older adult drivers will become an increasing public health concern. We report on the medical and functional factors associated with vehicle crashes in a cohort of Alabama drivers, 55 years old and older. METHODS: This prospective study involved 174 older adults, on whom demographic, medical, functional, and physical performance data were collected in 1991. Subjects were then followed through 1996 for incident vehicle crashes. RESULTS: Sixty-one subjects experienced between one and four police-reported vehicle crashes during the study period. Following adjustment for age, race, days driven per week, and gender, Cox proportional-hazards models showed the following variables to be associated with crash involvement: reported difficulty with yardwork or light housework (relative risk [RR] = 2.1; 95% confidence interval [CI] 1.1, 4.0; p = .02), or opening ajar (RR = 3. 1; 95% CI 1.4, 6.7; p = .004); at least one crash before 1991 (RR = 2.1; 95% CI 1.2, 3.7; p = .008); using hypnotic medication (RR = 2.9; 95% CI 1.3, 6.6; p = .01); self-reported stroke or transient ischemic attack (RR = 2.7; 95% CI 1.1, 6.6; p = .03); scoring within the depressed range on the Geriatric Depression Scale (RR = 2.5; 95% CI 1.1, 6.0; p = .03), and failing the useful field-of-view test (RR = 1.9; 95% CI 1.0, 3.5; p = .05). CONCLUSIONS: Variables related to function, medication use, affect, neurological disease, and visuocognitive skills were associated with vehicle crash involvement in this cohort. Our findings suggest that multifactorial assessments are warranted to identify at-risk older drivers. PMID- 10719770 TI - Gender differences in functioning for older adults in rural Bangladesh. The impact of differential reporting? AB - BACKGROUND: The purpose of this study is to examine gender differences in functional ability among older adults in rural Bangladesh in terms of both self reported activities of daily living and observed physical performance and to evaluate the extent to which differential reporting by gender contributes to disparities between the two measures. METHODS: In 1996, the Matlab Health and Socio-Economic Survey collected data on self-reported activities of daily living (ADLs) and observed physical performance for 1,893 men and women aged 50 and older in the Matlab Surveillance area in rural Bangladesh. Gender differences were examined in both self-reported ADLs and physical-performance measures. With physical-performance measures as the gold standard, logistic regression was used to determine how much of the gender difference in the self-reported function was explained by physical-performance ability controlling for age. RESULTS: Older women in this study population consistently had more limitations than men in both self-reported ADLs and observed physical performance. For the same level of observed physical performance, however, older women were more likely than men were to report a higher level of ADL limitation. This reported female health disadvantage varied considerably depending on the nature of the ADLs being examined and the type of scoring system used for the ADLs. CONCLUSIONS: One has to be somewhat cautious in interpreting gender differences in self-reported ADL limitations, as they are affected by the gender-specific nature of the reported activity and by gender differences in the perception of response categories. PMID- 10719771 TI - Longitudinal influence of age, menopause, hormone replacement therapy, and other medications on parotid flow rates in healthy women. AB - BACKGROUND: Recent investigations have demonstrated that parotid salivary dysfunction is not a normal process of aging, but may be the consequence of systemic conditions and their treatment, including medications and menopause. The purpose of this study was to assess longitudinally the influence of age, menopausal status, hormone replacement therapy, and other medications on stimulated parotid flow rates (SPFRs) in healthy women. METHODS: Medical diagnoses, menopausal status, medication utilization, and 2% citric acid stimulated parotid salivas were collected from 396 women, aged 21 to 96 years, from the Baltimore Longitudinal Study of Aging (National Institute on Aging, National Institutes of Health) over a 17-year span by three investigators. RESULTS: There was no overall longitudinal effect of time on SPFR. Age at first visit was a significant predictor of a decrease in SPFR when adjusted for time and xerostomic medications. However, the deleterious effect of taking one xerostomic medication was equivalent to approximately 14 years of aging. Menopausal status and hormone replacement therapy were not consistently associated with diminished SPFR. CONCLUSIONS: These results suggest that menopause and hormone replacement therapy are not associated with parotid salivary dysfunction. Aging may have a statistically significant yet small deleterious influence on SPFR; however, the adverse influence of xerostomic medications is much larger. PMID- 10719772 TI - Preclinical mobility disability predicts incident mobility disability in older women. AB - BACKGROUND: Physical disability and dependency are serious, and frequent, adverse health outcomes associated with aging and resulting from chronic disease. Reasoning has suggested that there might be a preclinical, intermediate phase of disablement which might develop in parallel with progression of underlying disease and precede and predict disability. Definition of this stage could provide a basis for screening and early intervention to prevent disability. The objective of this study was to determine preclinical functional predictors of incident mobility difficulty and provide evidence for a preclinical stage of disability. METHODS: A prospective, population-based cohort study was carried out in Baltimore, Maryland, with two evaluations 18 months apart. The participants were 436 community-dwelling women, 70-80 years of age at baseline, not cognitively impaired, and reporting difficulty in no areas, or only one area, of physical function (primarily mobility), who were participating in the Women's Health and Aging Study II. Participants were recruited from a population-based, age-stratified random sample. Incident mobility disability was studied in the subset without such disability at baseline. The main outcome measure was self reported incident difficulty walking 1/2 mile or climbing up 10 steps. RESULTS: At baseline, 69.3% of the cohort reported no difficulty with mobility. After 18 months, 16.0 and 11.7% of this group reported incident difficulty walking 1/2 mile or climbing up 10 steps, respectively. Those reporting baseline task modification due to underlying health problems, our measure of preclinical disability, were at three- to fourfold higher odds of progressing to difficulty than were those without such modification. In multivariate logistic regression analyses, this self-report measure, task modification without difficulty, and objective measures of performance were independently and jointly predictive of incident mobility difficulty. Specifically, for incident difficulty walking 1/2 mile, self-reported task modification odds ratio (OR) = 3.67, walking speed (.5 m/s difference) OR = 2.16; for incident difficulty climbing up 10 stairs, OR for task modification = 3.84, for stair climb speed (1/3 step/s difference) = 2.08 (95% CI did not include 1 for any). Covariates, age, living alone, number of chronic diseases, depression score, knee strength, and balance by functional reach, were not significant predictors in either model. CONCLUSIONS: Two indicators of functional changes in older women without mobility difficulty, self report of modification of method of doing a task in the absence of difficulty and performance measures, are independent and strong predictors of risk of incident mobility disability. The self-report measure provides substantial strength in predicting risk of incident disability across the full range of performance, and may identify a vulnerable point at which other risk factors act to cause transitions to disability. Together, the preclinical indicators identify a subset of high-functioning older women who are at high risk of mobility disability, and provide a potential basis for screening for disability risk and targeting interventions to prevent mobility disability. PMID- 10719773 TI - Allergy to natural rubber latex. PMID- 10719774 TI - Role for cysteinyl leukotriene receptor antagonist therapy in asthma and their potential role in allergic rhinitis based on the concept of "one linked airway disease". AB - OBJECTIVE: This review focuses on the shared pathophysiology of asthma and allergic rhinitis. The similarities illustrate the "one linked airway disease" concept, a unifying theory of these upper and lower airway inflammatory disorders. Since leukotrienes are mediators in both conditions, studies have been performed to assess the potential therapeutic role of cysteinyl leukotriene antagonists. The purpose of this paper is to provide an overview of the accumulating data concerning these agents in treating asthma and allergic rhinitis. DATA SOURCES: Relevant publications obtained from a literature review. STUDY SELECTION: Relevant publications on the topics of leukotrienes, leukotriene receptor antagonists, asthma, and allergic rhinitis were critically evaluated. RESULTS AND CONCLUSIONS: Studies to date have documented the efficacy of cysteinyl leukotriene receptor antagonists for asthma. The pathophysiology of allergic rhinitis and its similarities to asthma suggest that these agents could play a significant therapeutic role in managing this upper airway disorder. Because the leukotriene antagonists are oral agents, they may be valuable in treating not only either condition but also both at the same time when they coexist. They appear to be beneficial when prescribed as the initial medicine and when used in conjunction with other therapies. PMID- 10719775 TI - Orofacial edema: a diagnostic and therapeutic challenge for the clinician. AB - BACKGROUND AND CONCLUSION: A case of a 41-year-old patient with a 5-year history of chronic recurrent angioedema, refractory to conservative treatment is presented. The results of the case report suggest that in differential diagnosis of angioedema, in addition to usual causes, the allergist-immunologist needs to consider Melkersson-Rosenthal syndrome, which can present with a variety of symptom-combinations of the classic triad. The distinguishing characteristics of the Melkersson-Rosenthal syndrome are its refractoriness to the usual anti inflammatory therapy and the need to consider corrective cosmetic surgery, which may benefit some patients. PMID- 10719776 TI - A blinded, multi-center evaluation of two commercial in vitro tests for latex specific IgE antibodies. AB - PURPOSE: To compare the diagnostic value of two commercial in vitro tests for the detection of latex-specific IgE antibodies. METHODS: Serum samples were collected from latex-allergic and nonlatex-allergic individuals. Persons were classified as latex allergic if they had a positive clinical history and a positive skin prick test with a latex extract. Persons with no latex-related symptoms and negative skin tests were classified as nonlatex allergic. The serum samples were tested in a blinded fashion by a laboratory using the CAP (Pharmacia-Upjohn) and AlaSTAT (Diagnostic Products Company) assays. Values of 0.35 kA U/L or greater were considered positive in both tests. RESULTS: The 143 sera studied came from 83 latex allergic and 60 nonallergic persons. The in vitro tests were found to have sensitivities of 79.5% and 73.8%, and specificities of 90.2% and 91.7%, for CAP and AlaSTAT, respectively. The positive predictive values were 91.7% and 92.5%, while the negative predictive values were 76.4% and 71.4% for the CAP and AlaSTAT, respectively. CONCLUSION: In individuals classified by the combination of clinical history and skin test results, both the Pharmacia CAP and the DPC AlaSTAT demonstrated acceptable sensitivities, specificities, and predictive values for detection of antilatex IgE antibodies. These findings suggest that both assays can be useful adjuncts to the diagnosis of latex allergy. PMID- 10719777 TI - Prevalence of IgE to natural rubber latex in unselected blood donors and performance characteristics of AlaSTAT testing. AB - BACKGROUND: The prevalence of IgE to natural rubber latex (NRL) proteins in the general population remains unsettled, both because of the difficulty of obtaining an unbiased population representative of the general population of the United States and because of concerns about the reproducibility of tests for anti-latex IgE antibodies. Establishing the prevalence in the population is important toward defining the potential risks of persons entering areas where latex exposure may occur. OBJECTIVE: The purposes of this study were to determine the prevalence of IgE to latex in a general population and to assess the performance characteristics of the AlaSTAT microtiter plate test for anti-latex IgE when performed independently by different laboratories. METHODS: One thousand nine hundred and ninety-seven consecutive blood samples obtained from the Oklahoma Blood Institute were assayed independently in three laboratories for IgE to NRL using the FDA-approved AlaSTAT ELISA for IgE to NRL. The group consisted of 56% men and 44% women. Ninety percent were Caucasian, 4% African American, and 6% were "other." RESULTS: The prevalence IgE to NRL between the 3 laboratories varied from 5.4% to 7.6% at the designated cut off of 0.35 kU/L. Examination of results for specific individuals demonstrated >90% agreement between the three sites with the most reproducible results at the Class II cutoff of > or =0.7 kU/L. There was no difference in the percent of positive values at the three laboratories. CONCLUSIONS: There is good agreement between laboratories as to NRL IgE reactive and non-reactive sera using the AlaSTAT test. This report of the largest sample of blood donors confirms earlier reports as to the prevalence of IgE NRL in blood donors. PMID- 10719778 TI - Latex sensitization in children with spina bifida: follow-up comparative study after two years. AB - BACKGROUND: Previous findings suggest that sensitization to latex in children with spina bifida is a dynamic process. OBJECTIVE: To study if changes appear in the sensitization status after withdrawal of latex. METHODS: We studied a consecutive sample of 68 children with spina bifida, by means of latex skin prick tests and quantification of serum latex-specific IgE on two separate occasions two years apart. RESULTS: Forty-four (65%) were classified as nonsensitized, 6 (9%) showed indeterminate results, and 18 (26%) were sensitized to latex, six of whom had clinical reactions to latex. They were instructed to avoid latex. In a second evaluation, 2 years later, 38 (56%) were classified as nonsensitized, 3 (4%) as indeterminate, and 27 (40%) as sensitized to latex, 11 of whom had presented latex symptoms. This meant 22% of spina bifida children demonstrated progressive sensitization, in spite of having adopted a latex-free environment at our hospital. It illustrates the progressive character of latex sensitization in these patients. CONCLUSION: Latex avoidance measures both in the medical and home settings must be stressed. We recommend that children with spina bifida should be periodically evaluated regarding latex sensitization. PMID- 10719779 TI - Assessment of the readability and comprehensibility of a CFC-transition brochure. AB - BACKGROUND: In anticipation of public confusion about the availability of medications during the metered-dose inhaler CFC phaseout period, a multidisciplinary effort has resulted in the development of a brochure designed to educate patients and health care providers about the health consequences of ozone depletion and the transition to CFC-free inhaled products. This brochure is the subject of this assessment. OBJECTIVES: The primary purpose of this study was to estimate the grade of reading difficulty of a brochure designed to educate patients about the change to CFC-free inhalation products. A secondary objective was to assess baseline knowledge of patients concerning CFC transition and their comprehension of this issue after reading the brochure. METHODS: Standard readability formulae were used to assess the grade level of the CFC transition brochure. In addition, baseline knowledge of the CFC transition process and comprehensibility of the brochure were measured via a 2-page questionnaire. RESULTS: The SMOG, Rix, and Flesch-Kincaid tests yielded readability at grade levels of 14, 10, and 10.4, respectively. The survey indicated that even after reading the brochure, many patients had concerns about the transition process. CONCLUSIONS: These results suggest that the readability of the brochure entitled Your Metered-Dose Inhaler Will Be Changing... Here Are the Facts...may not be appropriate for a large segment of the population for whom it is intended. Further, the comprehensibility assessment suggests that many patients are either unaware of or unable to understand the impending changes to their inhaled therapies. PMID- 10719780 TI - Effect of age on response to zafirlukast in patients with asthma in the Accolate Clinical Experience and Pharmacoepidemiology Trial (ACCEPT). AB - BACKGROUND: The Accolate Clinical Experience and Pharmacoepidemiology Trial (ACCEPT), evaluated zafirlukast in a wide spectrum of patients from a variety of clinical practices. OBJECTIVE: To determine the effect of age on the response to zafirlukast 20 mg twice daily in 3759 patients with mild, moderate, or severe asthma. METHODS: Patients received open-label administration of zafirlukast 20 mg twice daily (bid) for 4 weeks. Pulmonary function was measured twice daily, and overall asthma symptom scores, number of nighttime awakenings, severity of morning asthma symptoms, and beta2-agonist use were recorded daily. Trial results were analyzed to compare the efficacy of zafirlukast in 263 adolescent (12 to 17 years old), 2602 adult (18 to 65 years old), and 321 elderly (66 years old and older) patients (the evaluable population). RESULTS: After 4 weeks of zafirlukast therapy, improvements in pulmonary function decreased with age and were significant for all measures in adolescents and adults and for morning peak expiratory flow in elderly patients. Improvements in symptom response were statistically significant across age groups. Reduction in beta2-agonist rescue was similar in adolescents and adults but significantly less in elderly patients. CONCLUSIONS: Zafirlukast is an effective treatment for asthma in all patients, regardless of age. In elderly patients, improvement in asthma symptoms rather than pulmonary function may represent a primary marker for efficacy with zafirlukast. PMID- 10719781 TI - Serologic markers for Chlamydia pneumoniae in asthma. AB - BACKGROUND: Chlamydia pneumoniae infection has been reported as a possible etiologic agent in asthma, which in primary care settings often appears to be initiated by acute respiratory infections. OBJECTIVE: To determine if serologic markers for C. pneumoniae are associated with adult asthma that first became symptomatic after an acute respiratory illness (asthma associated with infection: AAWI). METHODS: Serum samples from 164 primary care outpatients, mean age 44 years, (68 with AAWI; 36 with atopic, occupational or exercise-induced asthma (non-AAWI); 16 nonasthmatic patients with acute bronchitis; and 44 asymptomatic nonasthmatic controls) were tested for the presence of C. pneumoniae-specific IgG and IgA antibodies. Levels of chlamydial heat shock protein 60 (CHSP60) antibody were also measured. Those positive for CHSP60 were tested for C. pneumoniae specific IgE antibodies by immunoblotting. RESULTS: Statistically significant differences in IgG and IgA seroreactivity were noted between groups: acute bronchitis and AAWI had the highest levels (93% to 94% IgG seroreactivity, 69% to 75% IgA seroreactivity) whereas non-AAWI and asymptomatic controls had the lowest levels (61% to 84% IgG seroreactivity, 31% to 43% IgA seroreactivity, P < .02 after adjustment for age, sex and smoking). CHSP60 antibodies were significantly more prevalent in AAWI than in non-AAWI (19% versus 3%, P = .02). IgE antibodies against C. pneumoniae 60, 62, and/or 70 kD antigens were detected in 5 of 13 CHSP60 positive AAWI patients. Persistent IgG, IgA, and CHSP60 seroreactivities were noted in all seropositive asthma patients with serial serum samples. CONCLUSIONS: Serologic markers of C. pneumoniae infection were associated with acute bronchitis and with asthma that first became symptomatic following respiratory illness. Serologic responses to C. pneumoniae may be useful in the classification and diagnosis of asthma. PMID- 10719782 TI - Altered allergen binding capacities of Amb a 1-specific IgE and IgG4 from ragweed sensitive patients receiving immunotherapy. AB - BACKGROUND: The mechanisms for the effectiveness of allergen immunotherapy (IT) are not well understood. The binding potential for immunoglobulins is a function of both antibody concentration and affinity (K(A)). PURPOSE: The purpose was to perform a cross-sectional preliminary study to investigate any differences in allergen-specific antibody affinity and concentration following ragweed immunotherapy by introducing a new concept of antibody binding capacity ([Ig] X K(A)). METHODS: The binding capacity of allergen-specific IgE and IgG4 was determined for ragweed-allergic individuals undergoing ragweed immunotherapy and compared with the capacity of ragweed-specific IgE and IgG4 for allergic individuals not receiving immunotherapy. RESULTS: The mean binding capacity for IgG4 after long-term immunotherapy was 1.6 log units higher (P < .0001) than for individuals not receiving IT. The binding capacity for allergen-specific IgE was 1.2 log units lower following long-term immunotherapy (P < .0001) compared with individuals not receiving ragweed IT. CONCLUSIONS: We hypothesize that a primary effect of immunotherapy is to increase IgG4 binding capacity and concomitantly decrease IgE binding capacity. PMID- 10719783 TI - Study of the effects of vacuuming on the concentration of dust mite antigen and endotoxin. AB - BACKGROUND: Dust mite antigens are major sources of allergens in house dust and together with endotoxin, a proinflammatory component of gram negative bacteria also found in house dust, are important causes of tissue injury involved in the pathogenesis and severity of allergic diseases, eg, asthma. OBJECTIVE: To determine the effectiveness of vacuuming in reducing the quantity of dust mite antigens, Dermatophagoides pteronyssinus (Der p and Der p2) and endotoxin using a quantitative ELISA assay and to correlate results with those obtained using a qualitative rapid dipstick method for Der p2. METHODS: Four specimens of house dust were collected using a Kirby Model G5 vacuum cleaner with a Micron Magic Filtration system from an approximately 54" x 18" standardized area of rug from each of 20 homes at 4 time intervals over a 6-week period, ie, a baseline specimen #1 at 0 week; specimen #2 at 1 week; specimen #3 at 5 weeks (1 month after specimen #2); specimen #4 at 6 weeks (1 week after specimen #3). Three intervals were compared, ie, period 1-2 (1 week), period 2-3 (1 month), and period 3 to 4 (1 week). The concentrations of Der p1 and Der p2 were determined in dust samples using a standard ELISA assay and the concentration of endotoxin was detected using a limulus amebocyte lysate assay. Concentrations of Der p2, determined by the standard ELISA assay, were compared to those in the same samples determined by a rapid dipstick method. RESULTS: A wide range of values for total weight of unprocessed dust (0.3 to 59 g, X = 8.7) and finely sieved dust (0.1 to 19 g, X = 3) from all specimens were found. In finely sieved dust specimens the mean concentrations of Der p1, Der p2 and endotoxin were 775, 1310, and 3836 ng/g of dust, respectively. Following weekly vacuuming there was an increase in concentration of Der p1, Der p2, and endotoxin in 20%, 35%, and 63% of the houses, respectively, compared to in monthly vacuuming in which increases were seen in 65%, 50%, and 63% of the houses, respectively. In contrast, there was a decrease in concentration of Der p1, Der p2 and endotoxin with weekly vacuuming in 43%, 60%, and 37% of the houses respectively versus in monthly vacuuming in 15%, 35%, and 37% of the houses respectively. A correlation coefficient of 0.7 was found for the concentration of Der p2 in 37/40 samples tested detected using the ELISA method compared with rapid dipstick assay. CONCLUSION: The results of this study support the effectiveness of vacuuming on the reduction of dust mite antigens (Der p1 and Der p2) ie, Der p2 > Der p1. This reduction was more pronounced with weekly compared with monthly vacuuming. No reduction in the concentration of endotoxin was found. A good correlation was found between results obtained by ELISA and rapid dipstick assay for Der p2. PMID- 10719784 TI - Comparison of three commercial ultrasonic nebulizers. AB - BACKGROUND: The clinical acceptance of the initial ultrasonic nebulizers was impeded by their production of significant quantities of droplets larger than the respirable range that could have resulted in poor pulmonary deposition of nebulized medications. Subsequent modifications in the design of ultrasonic nebulizers have occurred. Overall nebulizer performance characteristics of the newer ultrasonic devices have not been evaluated. OBJECTIVE: Three commercially available ultrasonic nebulizers (DeVilbiss-Pulmosonic, Omron-Microair, Rhone Poulenc-Rorer-Fisoneb) were studied to compare the aerosol output characteristics. METHODS: The parameters studied were total volume output (TVO), time to nebulize total output (TTO), percent of droplets with volume diameters in the respirable range (PDVRR, 1 to 5 microm), albuterol concentration during nebulization, and the total drug delivered. All nebulizers were filled with 2.5 mL of saline and 0.5 mL of albuterol nebulizer solution. Three units from each manufacturer, each from a different lot, were evaluated in duplicate. RESULTS: The nebulizer with the largest volume output was the Omron (mean 2.94 mL), which also demonstrated the longest nebulization time (mean 10.3 min). The DeVilbiss and Rhone Poulenc-Rorer units delivered smaller volumes (mean 2.5 mL, 2.4 mL, respectively) but nebulized more rapidly (mean 2.21 min, 3.54 min, respectively). The Omron nebulizer generated the highest PDVRR with a mean of 38%. The DeVilbiss had a mean PDVRR of 16% and the Rhone Poulenc-Rorer a mean PDVRR of 21%. The majority of droplets from all three machines had a volume diameter smaller than the respirable range, ie, in the 0.5 to 1.0 microm range (Omron-60%, DeVilbiss 83%, Rhone Poulenc-Rorer-79%). For all three nebulizers there appeared to be no concentrating or diluting effect during nebulization implying that equal quantities of albuterol and diluent were delivered. The Rhone Poulenc-Rorer units demonstrated the greatest unit-to-unit variability with respect to TVO while the Omron units demonstrated the greatest unit to unit variability with respect to TTO. CONCLUSION: We conclude that several improvements in the design of ultrasonic nebulizers have resulted in the reduction of the size of the droplets generated. Our evaluation of the three commercially available ultrasonic nebulizers revealed that the majority of droplets generated were within or below the respirable range. There was no concentrating or diluting effect during nebulization for all three nebulizers. The output characteristics of the three devices differ and this will effect the delivery time as well as amount of drug delivered to the lungs. PMID- 10719785 TI - Successful administration of iron dextran in a patient who experienced a life threatening reaction to intravenous iron dextran. PMID- 10719786 TI - Allergy to grapes. PMID- 10719787 TI - Allergy to grapes. PMID- 10719788 TI - Measurement of quality of life in alcohol-dependent subjects by a cancer symptoms checklist. AB - There are few studies of Quality of Life measures (QoL) in alcohol-misusing patients. The present study addresses this deficiency. The sample consisted of 60 (39 men, 21 women) alcohol dependent subjects defined by DSM-IV criteria (American Psychiatric Association, 1994). At baseline (4-5 days after admission and detoxification) sociodemographic data were collected, and three questionnaires were administered: the Rotterdam Symptoms Checklist (RSCL), the Severity of Alcohol Dependence Questionnaire (SADQ), and Alcohol Problems Questionnaire (APQ). QoL scores for dependent alcoholics both for physical and psychological measures were significantly worse (higher) than those reported for a variety of cancer patients. Psychological symptom scores were higher than physical symptoms at baseline. Correlations of RSCL scores to both SADQ and APQ were greater for RSCL physical compared to psychological symptom scores. The subjects were followed up at 12 weeks when the RSCL was re-administered and relapse status ascertained. Fifty-eight (97%) subjects were successfully contacted at 12 weeks of whom 36 (62%) had relapsed. After a repeated measures ANOVA psychological and physical symptom subscores were statistically significantly improved as a result of not relapsing to heavy drinking. There was no significant change in scores in the relapse group when baseline and week 12 scores were compared. The RSCL measure is a useful QoL assessment tool in alcohol dependent subjects. PMID- 10719789 TI - Ethyl glucuronide: a marker of recent alcohol consumption with clinical and forensic implications. AB - A marker with a specific time spectrum of detection and both high sensitivity and specificity is required to diminish the clinically as well as forensically important gap on the time axis between short- and long-term markers of alcohol consumption like ethanol and CDT, GGT or MCV, respectively. Ethyl glucuronide (EtG) is a non-volatile, water-soluble, stable upon storage, direct metabolite of ethanol with a molecular weight of 222 g/mol that can be detected in body fluids for an extended time period after the complete elimination of alcohol from the body. We investigated 107 urine and 78 serum samples of a total of 107 inpatients in 4 groups: (1) 33 inpatients in acute alcohol withdrawal and long term treatment; (2/3) 29 and 15 addicted forensic psychiatric inpatients (#64 StGB, penal code); (4) 30 recently detoxified inpatients of a station for long term treatment by LC/MS-MS with the internal standard d5-EtG and additionally in the fourth group of patients also by gas chromatography/mass spectrometry (GC/MS). In 2 out of 33 inpatients of the first group, EtG could be determined 3 days after hospitalization; in an other subject, a relapse could be detected. In 2 out of 29 and in 1 out of 15 forensic inpatients of group 2 and 3, respectively--where neither clinical impression nor routine laboratory findings gave an indication for relapse--concentrations of EtG ranged between 0.1 and 18 mg/l in urine. For the serum samples of the 30 inpatients of group 4, we could demonstrate a total agreement for the results of the GC/MS and the LC/MS-MS method as to whether a sample was found to be positive or negative for EtG. We suggest that these results strengthen our earlier findings that ethyl glucuronide is a marker of alcohol consumption in general that can be detected for an extended time period after the complete elimination of alcohol from the body and a marker for relapse control with a specific time frame of detection intermediate between short- and long-term markers. PMID- 10719790 TI - Linear CT-scan measurements in alcohol-dependent patients with and without delirium tremens. AB - The aims of the present study were to examine whether chronic alcohol dependence and the development of delirium tremens are characterized by changes in linear CT measurements of brain liquor spaces and intracranial distances indicative of prefrontal atrophy, and frontal (sub)cortical or temporal (sub)cortical atrophy. Toward this end linear measurements were performed in 47 alcohol dependent patients with and without a history of delirium tremens and in 10 healthy volunteers using CT-scanning. The following linear measurements were calculated: (1) the Evans ratio; (2) the cella media index (CMI); (3) the maximum width of the third ventricle; (4) the maximum width of the fourth ventricle; (5) the maximum frontal subarachnoid space (MFSS); (6) the maximum width of the anterior interhemispheric fissure (MIF), and (7) the maximum width of the Sylvian fissure (MSF). The alcoholics were divided into subgroups according to the Munchner Alkoholismus Test (MALT) and the presence of delirium tremens. The MFSS of the alcohol-dependent patients was significantly larger than that of the controls. The MIF and MSF of high MALT scorers were significantly larger than those of low scorers and controls. Alcohol-dependent patients with a known history of delirium tremens had significantly larger MIF and MSF than did patients without delirium tremens and controls. The results suggest that alcohol dependence is characterized by prefrontal atrophy, and that frontal cortical and temporal (sub)cortical atrophy may be related to the development of delirium tremens. PMID- 10719791 TI - Effects of chronic ethanol consumption on SER of Purkinje neurons in old F344 rats. AB - The purpose of this study was to determine whether the observed swelling of smooth endoplasmic reticulum (SER) profiles in Purkinje dendrites in our old ethanol-fed F344 rats: (1) represented measurable dilatation, (2) was present in dendritic shafts and spines, and (3) was reversed following recovery from ethanol. Of the 45 rats in 3 treatment groups (chow-fed, pair-fed, and ethanol fed), 30 rats were euthanized after 40 weeks, and 15 were maintained on rat chow for an additional 20-week recovery period. Electron microscopy of cerebellar preparations was used to analyze morphological alterations in SER profile size within the dendritic shafts and spines of Purkinje neurons. Results showed significant SER dilatation following 40 weeks of ethanol consumption, which disappeared after ethanol withdrawal. PMID- 10719792 TI - Effects of ethanol, acetaldehyde, and acetic acid on histamine secretion in guinea pig lung mast cells. AB - We studied the direct effects of ethanol and its metabolites on the guinea pig lung mast cell, and the alterations caused in the histamine release induced by different stimuli. Guinea pig lungs cells dispersed by collagenase were used throughout. High concentrations of ethanol (100 mg/ml), acetaldehyde (0.3-3 mg/ml) and acetic acid (3 mg/ml) induced histamine release that was not inhibited by sodium cyanide (0.3 mM). Lower concentration of ethanol (10 mg/ml) and acetic acid (0.3 mg/ml), but not acetaldehyde, inhibited the histamine release induced by antigen and ionophore A23187. The histamine release induced by phorbol 12 miristate 13-acetate (1 microM) was also inhibited by ethanol (10 mg/ml). Changes in the levels of calcium, glucose and phosphatidic acid did not influence the effect of ethanol. We conclude that high doses of ethanol, acetaldehyde, and acetic acid cause a cytotoxic histamine release by independent mechanisms. Low concentrations of acetic acid inhibit the histamine release by pH reduction. Ethanol acts by a generalized effect that is independent of calcium and glucose suggesting a nonspecific effect that, nevertheless, is not cytotoxic since it can be reversed by washing the cells. PMID- 10719793 TI - Synaptic reorganization in the hippocampal formation of alcohol-fed rats may compensate for functional deficits related to neuronal loss. AB - We have examined the behavioral and neuroanatomical effects of long-term alcohol intake in rats ingesting a 20% solution of ethanol for 30 weeks. Previous studies have shown that this treatment provokes neuronal degeneration in the hippocampal formation, which occurs in parallel with remodeling processes. Spatial reference and working memory of alcohol-fed rats were evaluated during last 4 weeks of treatment by comparison of their performance with age-matched controls on the Morris water maze. Alcohol consumption did not affect the performance of rats in the reference memory task as indicated by the measures derived from the acquisition trials and from the probe-trial, which were highly similar for alcohol-fed and control animals. Also, performance in the working memory task was not significantly altered in alcohol-treated animals. No treatment-related changes in swim speed or impairments of sensorimotor abilities, tested in the visible platform task, were detected. Stereological methods were applied to evaluate the damage inflicted by alcohol intake in the structure of the hippocampal formation. In the alcohol-treated animals, there was a noticeable cell loss in the granular layer of the dentate gyrus (10%), and in CA3 (18%) and CA1 (19%) hippocampal subdivisions. In spite of the neuronal loss, the total number of synapses between mossy fibers and CA3 pyramids was unaffected by alcohol treatment suggesting that new synaptic contacts were formed between the surviving neurons. We show that, regardless the marked hippocampal cell loss in rats exposed to chronic alcohol intake, the reorganization that takes place at the synaptic level may alleviate the expected functional deficits. PMID- 10719794 TI - Effects of chronic alcohol consumption on spatial reference and working memory tasks. AB - The aim of this work was to determine the spatial memory impairments induced by chronic alcohol consumption in rats. The alcoholization process began on the 21st postnatal day and alcohol concentrations were gradually increased to reach a concentration of 20% that was maintained for 4 mon. Behavioral tests were performed in the Morris Water Maze (MWM). The first study assessed the effects of chronic alcohol intake on two reference memory tasks (a place learning with multiple trials and a new place learning carried out in the same experimental context). Alcohol-treated animals presented no overall impairment in their ability to process spatial information. Deficits were restricted to reduced behavioral flexibility in spatial strategies. The second study assessed working memory in two tasks in which information about platform location was only valid for one trial. In the first working memory task, the animals had to perform one trial per day and in the second task they were submitted to four trials per day. At the end of the second experiment, all animals were trained in a visual-cued task. In the second experiment, the most important deficits in alcohol-treated animals occur in spatial working memory tasks, and this impairment was independent of the intertrial interval used. In the second spatial working memory task, performance of the alcohol-treated animals in the earlier trials affected their performance in subsequent trials, suggesting that a process of proactive interference had taken place. The visual-cued task demonstrated that these behavioral impairments were produced without visuoperceptive impairments. PMID- 10719795 TI - Mapping of quantitative trait loci for ethanol preference in quasi-congenic strains. AB - Ethanol preference, a component of alcoholism, has been known for four decades to differ greatly between C57BL/6 and BALB/c inbred mouse strains. For mapping quantitative trait loci (QTLs) that affect ethanol preference, we used a set of B6.C Recombinant QTL Introgression (RQI) strains, which carry about 5% of the donor BALB/cJ (C) genome on a C57BL/6ByJ (B6) background. After characterizing males of the progenitor and RQI strains for variations in ethanol preference, we scanned their genome for polymorphisms at 244 dinucleotide-repeat marker loci known to differ between B6 and C. Because of the introgression of BALB/c-type QTLs onto the B6 background, some strains showed ethanol preference significantly lower or higher than that of the background strain, suggesting that genetic interaction between ethanol preference QTLs and the background can be operative. The genomic region showing the strongest influence on ethanol preference was on mouse chromosome 15, and corresponds to human chr.12q11-q13. PMID- 10719796 TI - Effects of chronic ethanol exposure on sleep in rats. AB - Sleep disturbance is a common complaint in alcoholics. When polysomnographic studies are performed in alcoholics, reductions in slow wave sleep are a common finding; however, few studies have evaluated the effects of chronic alcohol exposure on sleep in animal models. In the present study, the sleep EEG was evaluated in 40 Wistar rats who were exposed to chronic alcohol or control conditions in vapor chambers. Rats were exposed to ethanol vapors or control chambers for 6 weeks and then withdrawn. Sleep EEG was recorded before exposure (baseline), immediately following exposure, and 5 weeks after withdrawal from the ethanol/control chambers. In the ethanol-exposed animals, blood ethanol levels averaged 192 mg/dL over 6 weeks of exposure. Chronic ethanol exposure and withdrawal was not found to affect either slow wave sleep latency or slow wave sleep duration; however, overall spectral power as well as power in the delta, theta, and beta frequencies were significantly reduced following chronic exposure (2-4 Hz, [F(1, 17) = 18.11, p = 0.001], 4-6 Hz, [F(1, 17) = 15.98, p = 0.001], 6 8 Hz [F(1, 17) = 15.52, p = 0.001], 8-16 Hz band [F(1, 17) = 18.73, p < 0.0001], 16-32 Hz [F(1, 17) = 10.13, p = 0.005], and 1-50 Hz [F(1, 17) = 17.03, p = 0.001]. After 5 weeks of withdrawal, significant decreases still persisted in the delta and theta frequencies (2-4 Hz [F(1, 16) = 6.21, 0.024], 4-6 Hz [F(1, 16) = 6.26, 0.024], and 6-8 Hz [F(1, 16) = 4.84, p = 0.043]). These findings suggest that spectral analysis of the EEG is a highly sensitive measure of the effects of ethanol on sleep. These findings additionally demonstrate that chronic ethanol exposure can produce persistent diminution in the systems that generate cortical slow waves in the rat and thus may provide a model for understanding the mechanisms underlying sleep disturbances associated with alcoholism. PMID- 10719797 TI - Recurrent detoxification may elevate alcohol craving as measured by the Obsessive Compulsive Drinking scale. AB - Research has demonstrated a relationship between the number of previous alcohol detoxifications and increased severity of the alcohol withdrawal syndrome (AWS) that is hypothesized to be similar to an electrophysiologic "kindling process." Application of a "kindling" model to AWS suggests that neuroadaptation of the central nervous system to repeated detoxifications may also cause neurobehavioral alterations that may affect "craving." This study examined craving as assessed by the Obsessive Compulsive Drinking Scale (OCDS) in 67 adult outpatients meeting DSM-IV criteria (American Psychiatric Association, 1994) for alcohol dependence and AWS having either < 2 and > or = 2 previous detoxifications. Results of ANCOVA revealed that patients with > or = 2 previous detoxifications had higher scores on a scale that measures obsessive thoughts about alcohol, drinking urges and behaviors, and a composite of these scores after controlling for alcohol dependence severity, depressive symptoms and number of drinks 2 weeks prior to the study. Findings emphasize the need to address craving and other psychological variables with respect to treatment of AWS. PMID- 10719798 TI - Effects of acute ethanol on indices of cognitive-behavioral performance in rats. AB - The present experiment examined the effects of ethanol on several complex operant behaviors in rats. Tasks included: temporal response differentiation (TRD) to assess timing behavior; differential reinforcement of low response rates (DRL) to assess timing and response inhibition; incremental repeated acquisition (IRA) to assess learning; conditioned position responding (CPR) to assess auditory, visual, and position discrimination; and progressive ratio (PR) to assess motivation. Ethanol (0.0, 0.5, 1.0, 1.5, 2.0, and 3.0 g/kg via orogastric gavage) reduced accuracy and/or percent task completed for the TRD, DRL, and CPR tasks. For CPR, this reduction was accompanied by a reduction in response rates. Ethanol also reduced response rates on the PR task. There were no effects of ethanol on IRA performance. These data suggest that ethanol can selectively impair performance on cognitive-behavioral tasks and that these effects can occur at doses that do not affect the subjects' ability to respond. PMID- 10719800 TI - Lack of relationship between plasma thrombomodulin and portal hypertension in alcoholic liver disease. AB - The present study was performed to analyze the relationship between portal hypertension and alterations of the endothelium-derived proteins thrombomodulin, plasminogen activator (t-PA) and plasminogen activator inhibitor (PAI-1), which were determined in plasma samples of 28 alcoholic cirrhotic patients and 46 controls. Although cirrhotics showed lower levels of PAI-1, but higher thrombomodulin and t-PA levels than controls, no relationship was observed between thrombomodulin, t-PA or PAI-1 and portal pressure. Therefore, the hypothesis that splachnic endothelial damage secondary to portal hypertension leads to altered thrombomodulin, t-PA and PAI-1 levels in alcoholic cirrhosis is not supported by the results of this study. PMID- 10719799 TI - Acute alcohol intoxication and gadolinium chloride attenuate endotoxin-induced release of CC chemokines in the rat. AB - This work tests the hypotheses that Kupffer cells are a major source of CC chemokines (MIP-1alpha, MCP-1, RANTES) during acute endotoxemia and that acute ethanol intoxication modulates Escherichia coli lipopolysaccharide (LPS, 1 mg/Kg, i.v.)-induced chemokine release in the rat. LPS stimulated the release of CC chemokines into the circulation, hepatic sequestration of leukocytes and liver injury. LPS-induced serum chemokines peaked at 1-3 h and could not be detected at 24-h posttreatment. Splenectomy significantly suppressed LPS-induced RANTES release, but not MIP-1alpha and MCP-1. Kupffer cell depletion by gadolinium chloride or acute ethanol intoxication significantly attenuated LPS-induced CC chemokine release and hepatic injury. Hepatic sequestration of leukocytes during endotoxemia was also suppressed by acute ethanol. LPS downregulated the expression of MIP-1alpha and MCP-1 mRNAs and upregulated RANTES mRNA in Kupffer cells at 3-h post endotoxin. The expression of mRNAs was further suppressed in ethanol plus the LPS-treated group. Ethanol also suppressed the LPS-mediated priming of Kupffer cells for enhanced CC-chemokine release in vitro. Ethanol alone significantly upregulated the expression of CC-chemokine mRNA, and primed the Kupffer cells for enhanced RANTES release. CC-chemokine release and mRNA expression in hepatic sinusoidal endothelial cells were not significantly altered by ethanol, except for MCP-1 release. These data show that acute ethanol may be beneficial in tissue injury during acute endotoxemia. PMID- 10719801 TI - Research in Korean medical colleges. PMID- 10719802 TI - Scientific publication productivity of Korean medical colleges: an analysis of 1988-1999 MEDLINE papers. AB - To identify where the quality research activity has been and is carried out in Korea, and to examine to what extents Korean medical colleges play leading roles in the production of international research papers, we investigated the publication productivity of Korean medical colleges and their medical departments as measured by the number of papers published in foreign journals indexed in MEDLINE. The 12-year period from 1988 to 1999 is covered. A total of 4,881 papers is published in MEDLINE foreign journals by the researchers in Korean medical colleges during the period. The production of MEDLINE papers are concentrated in a few universities. More than 60% of MEDLINE foreign journal papers is published by top five universities 25% by Seoul National University, and 15% by Yonsei University. The newly established medical colleges at the University of Ulsan and Sungkyunkwan University produced outstanding numbers of papers in less than ten years. Radiology has led the internationalization of Korean medical papers. It was the most productive specialty identified in this study. The productivity of Internal medicine is on the rise from the mid-1 990s, and the field began to produce the most number of papers since then. PMID- 10719803 TI - CT interpretation of gastrointestinal tract diseases. AB - Most inflammatory, neoplastic and vascular disorders manifest bowel wall thickening on computed tomography (CT). Therefore, it is very important to understand the patterns of bowel wall involvement (degree, length, symmetry and contrast enhancement patterns) in each category to make a correct diagnosis. Observing extraluminal changes also help to classify the primary causes of pathological conditions involving the gastrointestinal tract. Adequate CT examinations with optimal opacification of the gastrointestinal tract are essential not only to avoid false positive findings but also to detect subtle or minimal lesions. If findings for establishing a diagnosis are equivocal, the use of combined findings increases the diagnostic accuracy of CT. PMID- 10719804 TI - Prediction on lengths of stay in the postanesthesia care unit following general anesthesia: preliminary study of the neural network and logistic regression modelling. AB - The length of stay in the postanesthesia care unit (PACU) following general anesthesia in adults is an important issue. A model, which can predict the results of PACU stays, could improve the utilization of PACU and operating room resources through a more efficient arrangement. The purpose of study was to compare the performance of neural network to logistic regression analysis using clinical sets of data from adult patients undergoing general anesthesia. An artificial neural network was trained with 409 clinical sets using backward error propagation and validated through independent testing of 183 records. Twenty-two inputs were used to find determinants and to predict categorical values. Logistic regression analysis was performed to provide a comparison. The neural network correctly predicted in 81.4% of situations and identified discriminating variables (intubated state, sex, neuromuscular blocker and intraoperative use of opioid), whereas the figure was 65.0% in logistic regression analysis. We concluded that the neural network could provide a useful predictive model for the optimization of limited resources. The neural network is a new alternative classifying method for developing a predictive paradigm, and it has a higher classifying performance compared to the logistic regression model. PMID- 10719805 TI - All-trans-retinoic acid attenuates neointima formation with acceleration of reendothelialization in balloon-injured rat aorta. AB - Retinoic acids may inhibit vascular smooth muscle cell proliferation, but may promote endothelial cell proliferation in cell culture. However, little data are available about the effects of all-trans-retinoic acid (ATRA) on endothelial regeneration and functional recovery in an experimental model of vascular injury. Accordingly, we investigated whether ATRA may attenuate neointima formation and accelerate endothelial regeneration with functional recovery in balloon-injured rat aorta. Twelve-week-old male Sprague-Dawley rats underwent endothelial denudation of the thoracic aorta by balloon injury. Fourteen rats were fed a standard rat pellet diet. Another 14 rats were fed ATRA (1.5 mg/day) for 2 weeks. The animals were killed on day 14 for organ chamber study and morphometric analysis. Rats in the ATRA group had a significantly improved acetylcholine induced relaxation response than those in control group. However, endothelial independent response was not significantly different between the two groups. The extent of reendothelialization was markedly superior in the ATRA group compared with control group (p<0.05). Furthermore, neointima area and the ratio of neointima to medial area were significantly less in ATRA group than in control group (p<0.05). In conclusion, ATRA may accelerate endothelial regeneration with functional recovery, and attenuate neointima formation in balloon-injured rat aorta. PMID- 10719806 TI - Outcome of adults with repaired tetralogy of Fallot. AB - Outcome of adult patients with repaired tetralogy of Fallot (TOF) was studied with emphasis on postrepair problems. A retrospective review of clinical, echocardiographic, catheterization, and surgical data was performed for 48 patients who underwent corrective repair of TOF after 15 years of age. All patients survived total repair and have been followed up from 3 months to 11 years (median 4.6 years). Postoperatively, 81.3% of patients were in functional class I and 85.4% had normal right ventricular function. One patient (2.1%) died during follow-up. There were 6 reoperations (12.5%) in 5 patients. The indications for reoperation included residual ventricular septal defect (VSD) (n=1), right ventricular outflow obstruction with VSD (n=4), and pulmonary regurgitation (n=1). The 10-year actuarial survival rate was 97.1%, and the 10 year freedom from reoperation was 81.3%. Aortic regurgitation was seen preoperatively in 6 patients (12.5%) and there were 2 newly developed aortic regurgitations after operation, one of which was caused by infective endocarditis. Corrective repair of TOF can be recommended in this patient group since the survival rate, postrepair functional status and hemodynamics are acceptable. Continued close follow-up, however, is essential for early identification and correction of post-repair problems. PMID- 10719807 TI - Morphologic change of the internal elastic lamina in Buerger's disease. AB - Morphologic features and pathogenesis of arterial changes occurring in Buerger's disease (thromboangiitis obliterans) are still controversial. This study describes histopathologic features of medium sized arteries from patients with Buerger's disease, particularly of the internal elastic lamina in relation to the immunologic mechanism of the injury. Seventeen segments of occluded arteries (femoral or popliteal arteries) from 17 patients with Buerger's disease were analyzed by histopathological and immunohistochemical methods. The most characteristic features were total luminal obliteration, together with a varying degree of recanalization and deposition of hemosiderin pigments. Detailed analysis, however, showed marked undulation and multiplication of the internal elastic lamina (100%) associated with basophilic degeneration and delicate linear calcification (47%). Lymphocytic infiltration along the internal elastic lamina was seen in 71% and was associated with localized edema. Lymphocytes along the lamina were consistently positive for T cell marker. Mild to moderate fibrosis was present at the media in 24%. Adventitial changes included mild, nonspecific and irregular fibrosis seen in 53%. Immunologic injury to the internal elastic lamina associated with T-lymphocytic infiltration might be the initial morphogenetic mechanism of the thrombotic occlusion and organization of medium sized arteries in Buerger's disease. PMID- 10719808 TI - The possible cost effectiveness of peripheral blood stem cell mobilization with cyclophosphamide and the late addition of G-CSF. AB - The purpose of this study was to develop a cost-effective protocol for the mobilization of peripheral blood stem cells (PBSC) in patients with malignancy. Thirty consecutive patients were randomized to mobilize PBSC with the late addition of a standard 250 microg dose of G-CSF (Neutrogen) from day 8 or early addition of the same dose of G-CSF from day 2, following cyclophosphamide (CY) 4 g/m2. The median yield of CD34+ cells from evaluated patients was 7.87 x 10(6)/kg (range, 2.06-27.25), collected in a median of four apheresis (range, 2-9). Target CD34 + cell doses > or = 2.0 x 10(6)/kg were achieved in all patients able to be evaluated. There were no statistically significant differences in CD34+ cell yields or toxicities. Overall engraftment occurred with median days to neutrophils > or = 0.5 x 10(9)/L or platelets > 20 x 10(9)/L of 11 and 17 days, respectively. However, the duration of G-CSF administration was markedly shorter in the late use of G-CSF group than in the early use of G-CSF group, with a median of 9 days compared with 15 days (p<0.001). PBSC harvesting after priming with CY plus delayed use of G-CSF made it a safe and cost-effective procedure. PMID- 10719809 TI - Mutations of hepatitis C virus 1b NS5A 2209-2248 amino acid sequence is not a predictive factor for response to interferon-alpha therapy and development of hepatocellular carcinoma. AB - Genetic changes between codons 2209 and 2248 of NS5A of genotype 1b hepatitis C virus (HCV-1b) have been reported to be associated with the sensitivity to interferon-alpha (IFN-alpha). The present study was performed to analyze such relationship in Korean patients with chronic hepatitis C and HCV-1b (n=19), including 12 chronic hepatitis C patients treated with IFN-alpha, 3 chronic hepatitis C patients without treatment as controls, and 4 patients with hepatocellular carcinoma (HCC). Two serum samples, before and after the treatment, were analyzed for the mutations by reverse transcription-polymerase chain reaction, cloning and sequencing. The mutations were identified in 32% (6/19), including five intermediate type (1-3 mutations) and one mutant type (4 or more). In 12 patients treated with IFN-alpha, the number of amino acid substitutions in NS5A2209-2248 was not associated with outcome of the treatment. Two HCV isolates with NS5A2209-2248 mutations from HCC patients were intermediate type. These results do not support that the NS5A2209-2248 determines interferon sensitivity of HCV-1b and that the mutations is associated with development of HCC. PMID- 10719810 TI - p53 protein expression and its prognostic importance in patients with nodal non Hodgkin's lymphoma. AB - To determine whether the p53 expression might be a predictor for treatment response and overall survival in nodal non-Hodgkin's lymphoma (NHL), we analyzed the expression of p53 in 69 NHL patients. p53 protein expression was analyzed by immunohistochemistry with long-term follow up (1-148 months: median 12.2). p53 expression was noted in 23/69 (33.3%) patients. Complete response (CR) rate to systemic chemotherapy was correlated with stage (I/II) (p=0.038), but not with p53 expression (p=0.2856). Poor overall survival was associated with stage (p=0.0010) or IPI score (p=0.0076), but not with p53 expression (p=0.8601). From stratification analysis by stage, in stage III/IV patients, the p53 positive group had a trend to be associated with poor overall survival than the p53 negative group. Multivariate analysis revealed that p53 positive group was associated with less CR rate compared to the p53 negative group (p=0.046), whereas overall survival was correlated with stage (p=0.0320), not with p53 status. p53 expression was associated with less CR rate in patients with DLBL. Further studies with large numbers of samples and homogenous group of NHL are needed to determine the prognostic value of cell cycle regulator, p53 in NHL. PMID- 10719811 TI - p53 codon 72 polymorphism and risk of cervical carcinoma in Korean women. AB - A common polymorphism of the wild type p53 is known at codon 72 of exon 4, with 2 alleles encoding either arginine (CGC, p53Arg) or proline (CCC, p53Pro). A recent study suggested that this polymorphism affects the susceptibility of p53 protein to human papillomavirus E6 oncoprotein mediated degradation and that individuals homozygous for p53Arg are seven times more susceptible to HPV-associated carcinogenesis of the cervix than heterozygotes. To examine whether the p53Arg genotype could be a risk factor for HPV-associated cervical carcinomas in the Korean population, we analyzed the p53 codon 72 polymorphism status of HPV positive invasive cervical carcinomas from 52 Korean women and 103 healthy control samples. The proportion of individuals homozygous for p53Arg, homozygous for p53Pro, and heterozygous for the two alleles were 40%, 19%, and 41% in normal healthy controls; 42%, 17%, and 40% in women with HPV-positive invasive cervical carcinoma. There were no significant differences in the distribution of p53 genotypes between controls and cervical carcinomas. This finding indicates that the p53Arg genotype is not associated with an increased susceptibility to cervical carcinoma in Korean women. PMID- 10719812 TI - Effects of general and locoregional anesthesia on reproductive outcome for in vitro fertilization: a meta-analysis. AB - The objective of this meta-analysis was to evaluate prospective trials of general or locoregional anesthesia on reproductive outcomes (cleavage and pregnancy rate) for in vitro fertilization (IVF). Of 115 published studies retrieved from a search of articles indexed on MEDLINE from 1966 to February 1999, four studies with distinct general and locoregional anesthesia were deemed eligible for meta- analysis. The pooled relative risk and odds ratios were calculated. A test for homogeneity was also performed. The pooled log odds ratio was 1.03 (95% CI 0.90 1.18) in cleavage rate and 0.71 (95% CI 0.47-1.08) in pregnancy rate. Heterogeneity was negative. Cleavage and pregnancy rates were not significantly different in both the general anesthesia and locoregional anesthesia groups. Both anesthetic techniques were favorable to IVF procedure by available published evidence when anesthesia was needed. PMID- 10719813 TI - Detecting p53 gene mutation of breast cancer and defining differences between silver staining PCR-SSCP and immunohistochemical staining. AB - This study detects and defines the patterns of p53 gene mutations in breast cancers. We analyse p53 gene mutations through comparing the results of single strand-conformation-polymorphism (SSCP) and immunohistochemistry (IHC), and we try to define the differences between the results of SSCP and IHC. Twenty-seven fresh primary breast cancer tissues and eight normal breast tissues were studied. The IHC was done with the usual streptavidin-biotin peroxidase complement method by using monoclonal antibody DO-7. The results of staining was scored. The SSCP method was done by using Cold SSCP Electrophoresis System. Overexpressions of p53 protein were seven (25.9%) among 27 cancer cases on IHC. Four (57.1%) of seven cases were positive in SSCP. In SSCP, the mutations were detected in 10 (37%) among 27 cancer cases. The mutations were two in exon 5, one in exon 8, and seven cases in exon 7. All of 10 mutations were proved by sequencing analysis. Of them, only four (40%) were positive in IHC. We consider the IHC as a screening method for p53 gene mutations. PMID- 10719814 TI - Foot screening technique in a diabetic population. AB - Foot complications are a well known factor which contribute to the morbidity of diabetes and increases the chance of amputation. A total of 126 consecutive diabetic patients were evaluated by diabetic foot screening. Forty-one patients showed an impaired protective sense when tested with Semmes-Weinstein monofilament 5.07 (10 g), and 92% of them showed peripheral polyneuropathy in nerve conduction study (NCS). The mean vibration score of the Rydel-Seiffer graduated tuning fork in patients with peripheral polyneuropathy in nerve conduction (NCV) study was 5.38+/-2.0, which was significantly different from that of patients without polyneuropathy in NCS. Among the deformities identified on examination, callus, corn, and hallux valgus were the greatest. While checking the ankle/ brachial index (ABI), we also evaluated the integrity of vasculature in the lower extremities. After extensive evaluation, we classified the patients into eight groups (category 0,1,2,3,4A,4B,5,6). The result of this study suggested that the Semmes-Weinstein monofilament test, Rydel-Seiffer graduated tuning fork test, and checking the ankle/brachial index were simple techniques for evaluating pathologic change in the diabetic foot by office screening, and that this screening based on treatment-oriented classification helps to reduce pedal complications in a diabetic population. PMID- 10719815 TI - NRAMP1 gene polymorphisms in patients with rheumatoid arthritis in Koreans. AB - Natural resistance-associated macrophage protein 1 (Nramp1) is a genetic locus associated with innate resistance or susceptibility of murine hosts to infection with intracellular pathogens such as Salmonella, Leishmania and Mycobacterium. The human homologue of the Nramp1 gene, designated NRAMP1, has been investigated as a candidate gene for genetic susceptibility to autoimmune diseases as well as infections. This study tries to determine whether NRAMP1 polymorphisms are associated with susceptibility to rheumatoid arthritis in Koreans. The nine NRAMP1 polymorphisms (1 microsatellite, 1 variation in 3' UTR, 5 silent substitution, 2 amino acid substitution) were typed by PCR-RFLP in 74 patients with rheumatoid arthritis (RA) and 53 healthy controls in Koreans. The distribution of allele and genotype frequencies were compared between patients and controls. Three NRAMP1 polymorphisms (823C/T, D543N and 1729+55del4) were significantly associated with RA. In addition, there were significant differences in the genotype frequencies for 823C/T, D543N and 1729+ 55del4 polymorphisms between RA patients and controls. Genotypes of A/A homozygote for D543N and TGTG deletion homozygote for 1729+55del4 were only detected in the patient group. These data indicate that genetic polymorphisms of NRAMP1 might be associated with the susceptibility to rheumatoid arthritis in Koreans. PMID- 10719816 TI - Expression of cyclooxygenase-1 and -2 in rheumatoid arthritis synovium. AB - The aim of this study was to investigate the expression and localization of cyclooxygenase-1 and -2 (COX-1 and COX-2) in synovial tissues from patients with rheumatoid arthritis (RA). Synovial tissues from 9 patients with RA and 5 patients with osteoarthritis (OA) were examined for COX-1 and COX-2 expressions by immunohistochemical staining using 2 polydonal COX-1 and COX-2 antibodies. In RA synovia, synovial lining cells showed intense immunostaining for COX-1, whereas slight to moderate staining was observed in inflammatory cells, stromal fibroblast-like cells and vascular endothelial cells. There was no significant difference in COX-1 expression between RA and OA synovia. The localization of COX 2 expression dearly differed from that of COX-1 expression, being most intense in inflammatory cells. However, there was no difference in COX-1 and COX-2 expressions between RA and OA synovial tissues. Our observations support that inflammatory mechanisms modulated by COX-1 and COX-2 in chronic RA synovium might be similar to those in chronic OA synovium. PMID- 10719817 TI - Deletion of SMN and NAIP genes in Korean patients with spinal muscular atrophy. AB - Childhood-onset proximal spinal muscular atrophies (SMAs) are an autosomal recessive, clinically heterogeneous group of neuronopathies characterized by selective degeneration of anterior horn cells. The causative genes to be reported are survival motor neuron (SMN) and neuronal apoptosis inhibitory protein (NAIP) genes. The deletion of telomeric copy of SMN (SMN(T)) gene was observed in over 95% of SMAs. The deletion rate of NAIP gene is 20-50% according to disease severity. The objective of this article is to genetically characterize the childhood-onset spinal muscular atrophy in Koreans. Five Korean families (14 constituents containing 5 probands) with SMA were included in this study. Polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) were used for the deletion analysis of SMN(T). Multiplex PCR method was used for NAIP analysis. Four probands showed deletion of SMNT gene. Deletion of SMN(C) (centromeric SMN) gene was found in one proband who did not show the deletion of SMN(T) gene and in the father of one proband who showed the deletion of SMN(T) gene. The deletion of NAIP gene was not found among all the studied individuals. The extent of deletion in Koreans was smaller than that in other studied population. PCR-RFLP deletion analysis can be applied to diagnose SMA and make a prenatal diagnosis. PMID- 10719818 TI - A case of Goodpasture's syndrome with massive pulmonary hemorrhage. AB - We present a typical case of Goodpasture's syndrome with massive pulmonary hemorrhage and acute deterioration of renal function. A 20-year-old male was admitted due to severe azotemia (blood urea nitrogen 214.7 mg/dL, serum creatinine 30.2 mg/dL) and was treated with emergency hemodialysis. On the 4th hospital day, a sudden onset of pulmonary hemorrhage developed. The circulating level of anti-glomerular basement membrane antibody was then elevated highly, and the kidney biopsy showed crescentic glomerulonephritis and linear deposition of IgG along the glomerular capillary. The patient was treated with intravenous high dose-steroid, oral cyclophosphamide and plasma exchanges. The pulmonary hemorrhage improved with the therapy, however, his renal function did not improve. He is currently on a regular schedule of hemodialysis. PMID- 10719819 TI - Successful allogeneic bone marrow transplantation for childhood-onset refractory anemia with ringed sideroblasts. AB - Refractory anemia with ringed sideroblasts (RARS) is an extremely rare type of myelodysplastic syndrome in children. We describe a 10-year-old boy with RARS presented with pancytopenia. He remained relatively stable with only a few transfusions until age of 20 years, when he underwent an allogeneic bone marrow transplantation (BMT) because of increased transfusion requirements. He remains in complete chimeric state at 20 months posttransplant with normal hematologic parameters. To our knowledge, this is the first description of successful BMT in a patient with childhood-onset RARS. The indication of BMT for this rare disorder in children is discussed. PMID- 10719820 TI - Three cases of pancreas allograft dysfunction. AB - We present dincopathologic features of three cases of biopsy-proven pancreas allograft dysfunction in Korea. All patients had advanced insulin-dependent diabetes mellitus (IDDM). Case 1 was a 30-year-old woman who underwent a simultaneous pancreas-kidney transplantation. Urinary infection developed 6 days after the operation, which remitted and reappeared, when urine amylase level was normal. Since the 55th day after the operation, intermittent hematuria has persisted. Cytomegalovirus inclusions were detected on the urinary bladder and grafted duodenal mucosa. The graft was removed due to perforation of the grafted duodenum and panperitonitis. Case 2 was a 27-year-old man undergoing pancreas transplantation alone (PTA). Ten days after the transplatation, the level of 24 urine amylase decreased and the graft was not delineated by 99mTc DTPA scintigraphy. Allograft needle biopsy revealed multiple acinar cell necrosis and mild lymphocytic infiltration which were compatible with mild acute rejection. Case 3 was a 25-year-old man undergoing cadevaric PTA. Three months after the transplantation, graft was removed due to gastric perforation associated with cytomegalovirus and angiodestructive fungal infection. Various causes of pancreas allograft dysfunction can be diagnosed by needle biopsy, thus appropriate biopsy specimen should be taken using improved biopsy technique. PMID- 10719821 TI - Fibrosing cholestatic hepatitis: a report of three cases. AB - Fibrosing cholestatic hepatitis is an aggressive and usually fatal form of viral hepatitis in immunosuppressed patients. We report three cases of fibrosing cholestatic hepatitis in various clinical situations. Case 1 was a 50-year-old man who underwent a liver transplant for hepatitis B virus (HBV)-associated liver cirrhosis. Two and a half years after the transplant, he complained of fever and jaundice, and liver enzymes were slightly elevated. Serum HBsAg was positive. Case 2 was a 30-year-old man in an immunosuppressed state after chemotherapy for acute lymphoblastic leukemia. He was a HBV carrier. Liver enzymes and total bilirubin were markedly elevated. Case 3 was a 50-year-old man who underwent renal transplantation as a known HBV carrier. One year after the transplant, jaundice developed abruptly, but liver enzymes were not significantly elevated. Microscopically lobules were markedly disarrayed, showing ballooning degeneration of hepatocytes, prominent pericellular fibrosis, and marked canalicular or intracytoplasmic cholestasis. Portal inflammation was mild, but interphase activity was definite and cholangiolar proliferation was prominent. Hepatocytes were diffusely positive for HBsAg and HBcAg in various patterns. Patients died of liver failure within 1 to 3 months after liver biopsy in spite of anti-viral treatment. PMID- 10719822 TI - Primary Sjogren's syndrome manifested as multiple sclerosis and cutaneous erythematous lesions: a case report. AB - Sjogren's syndrome is a chronic autoimmune disorder characterized by lymphocytic infiltration of the lacrimal and salivary glands, leading to dryness of eyes (kerato-conjunctivitis sicca) and mouth (xerostomia). The skin lesions in Sjogren's syndrome are usually manifested as xeroderma, but sometimes appear as annular erythema or vasculitis. Central nervous system symptoms may be presented as one of extraglandular manifestations, though rare in incidence, and need differential diagnosis from multiple sclerosis. We report a case of a 45-year-old woman diagnosed as multiple sclerosis at first but later as neurologic manifestation of primary Sjogren's syndrome, showing signs of multiple sclerosis and cutaneous erythematous lesions. PMID- 10719823 TI - Unusual association of pulmonary artery sling with right aortic arch and aberrant left subclavian artery. AB - We present an unusual case of vascular sling, tracheal stenosis by complete cartilaginous ring, and aberrant left subclavian artery with right aortic arch that underwent successful surgical repair for the sling. These abnormalities were suspected from unusual multiple indentations found on esophagogram. Complete preoperative diagnosis was established with chest computerized tomogram combined with angiography. PMID- 10719824 TI - Saline-enhanced radiofrequency ablation of breast tissue: an in vitro feasibility study. AB - RATIONALE AND OBJECTIVES: The feasibility of radiofrequency (RF) ablation for the treatment of breast tumors was investigated in vitro. The best parameters for ablation of breast tissue were chosen. METHODS: Saline-enhanced RF ablation was performed in human breast tissue specimens and cow udder tissue. Temperature profiles were measured depending on RF power (20, 28, 36 W) and NaCl infusion rate (15, 30, 60 mL/h) using eight thermocouples. Lesion development was monitored by ultrasound. Thermolysis efficiency was measured by tissue weight determinations before and after ablation. RESULTS: After RF ablation of tissue samples, 73.6% turned into a fat/saline emulsion. Ultrasound monitoring showed a cone-shaped hyperechoic area during the first 2 minutes of RF ablation, followed by an irregular expansion of the area. Time-dependent spatial temperature curves were more homogeneous at low infusion rates (15 mL/h). Peak temperatures up to 160 degrees C were measured. CONCLUSIONS: Controlled RF ablation of breast tissue is feasible. The irregular expansion of RF lesions in fatty breast tissue is due to liquefied fat. Low saline interstitial infusion rates result in better control of lesioning. PMID- 10719825 TI - Polyiodinated triglyceride lipid emulsions for use as hepatoselective contrast agents in CT: effects of physicochemical properties on biodistribution and imaging profiles. AB - RATIONALE AND OBJECTIVES: A novel lipid emulsion (LE) was developed for hepatoselective delivery of a polyiodinated triglyceride (ITG) with potential for use in CT. This work assessed the effects of mean particle size, total administered dose, and formulation composition on the in vivo biodistribution and imaging profiles of the ITG-LE in rats. METHODS: The concentration of radioactivity derived from intravenously administered 125I-ITG-LE was determined as a function of time after injection. CT imaging studies of the abdomen evaluated the extent of hepatic enhancement after administration of ITG-LE. RESULTS: Mean emulsion particle diameter and total administered dose exerted the greatest effect on ITG-LE biodistribution profiles. In the optimal delivery scenario, >70% of the administered dose localized to the liver 30 minutes after injection. Liver enhancement profiles in CT imaging studies were consistent with biodistribution profiles. CONCLUSIONS: These results suggest that an appropriately formulated and administered dose of ITG-LE provides tissue selective localization of contrast material for use in CT. PMID- 10719827 TI - MRI study of transient cerebral ischemia in the gerbil: interest of T2 mapping. AB - RATIONALE AND OBJECTIVES: The aim of this study was to evaluate the diagnostic use of MRI and, more precisely, the use of quantitative T2 imaging at 7 T for the early detection of neuronal cerebral alterations after transient ischemia in the gerbil. METHODS: One hundred forty-seven Mongolian gerbils were separated into four groups for which a bicarotid artery occlusion lasted for 4, 6, 8, or 10 minutes, respectively. The animals were scanned before carotid artery occlusion and at 3, 6, 10, 24, and 48 hours and 5 days after the ischemic incident. MR images were acquired on a Bruker Avance DRX300 mini-imaging system. RESULTS: Our results show that T2 mapping is able to localize brain damage induced by transient ischemia and to detect early perturbations in water content (as early as 6 hours after ischemia). CONCLUSIONS: T2 measurements in the striata are correlated with the severity of the ischemic incident, since the changes observed on the T2 images are directly proportional to the duration of occlusion. PMID- 10719826 TI - Effects of magnetic thermoablation in muscle tissue using iron oxide particles: an in vitro study. AB - RATIONALE AND OBJECTIVES: To study the effects of magnetic thermoablation in muscle tissue from cow to assess interrelations that might be relevant for a minimally invasive therapy system in the long term. METHODS: Magnetite particles (50-180 mg) were placed in muscle tissue. Temperature elevations as a function of time and distance from the center of the magnetite deposition area were measured during the exposure (up to 304 seconds) to an alternating magnetic field (frequency 400 kHz, amplitude 6.5 kA/m) generated by a circular coil (diameter 90 mm). Measured curves were reproduced by numerical calculations. Tissue alterations, macroscopically visible as light-brown discoloration, were recorded by volume estimations and histopathologic studies. RESULTS: Significant temperature elevations (up to 87 degrees C) were reported within a distance of less than 15 mm from the magnetite deposition area. High initial heating rates were observed during the first 150 seconds of heating. The reproduction of the measured curves by numerical calculations was good (SD = 0.7 degrees C). The theoretical simulation was verified and applied to situations beyond the range of experimental conditions. Damaged tissue comprised pyknotic cell nuclei and degenerated myofibrils. Corresponding volumes were found to be up to 10 times higher than the volume of iron oxide dispersion. CONCLUSIONS: The data demonstrate the applicability of local magnetic thermoablation for therapy of muscle lesions in the long term. PMID- 10719828 TI - Treatment of postinterventional pseudoaneurysms by ultrasound-guided compression. AB - RATIONALE AND OBJECTIVES: This 3-year study was performed to evaluate the effectiveness and safety of ultrasound-guided compression (UGC) in the treatment of postinterventional pseudoaneurysms (PAs). METHODS: One hundred ten PAs were sonographically diagnosed after peripheral or cardiac interventions. In 98 patients (65 men and 33 women; age range, 44-79 years), UGC was performed. The PAs were related to the common femoral artery (n = 78), the superficial femoral artery (n = 26), the profound femoral artery (n = 2), and the distal external iliac artery (n = 4). The PAs showed diameters ranging from 0.8 to 9.86 cm (mean, 4.8 cm) and volumes between 0.6 and 109 mL (mean, 15.6 mL). Follow-up examinations including color Doppler-coded ultrasound and peripheral Doppler were performed after 18 hours +/-6 and 28 days +/-4. RESULTS: Complete closure of the PA and its neck was achieved by UGC in 96 of 98 cases (98%). In 86 of 98 cases (87.8%), UGC was successful during the first session; a second treatment was needed in 10 cases. The compression time varied from 12 to 85 minutes (mean, 35.6 minutes). Except for medically controllable vagal reactions in 4 of 98 cases (4.1%) and 1 easily controlled PA rupture, no treatment-related complications were observed. CONCLUSIONS: According to the effectiveness and safety results, we conclude that UCG is the method of choice in the treatment of postinterventional PAs. Diagnosis and UGC treatment should be performed as early as possible to minimize symptoms and hospitalization time. PMID- 10719829 TI - Advanced mitral annular calcification is associated with severe coronary calcification on fast dual spiral computed tomography. AB - RATIONALE AND OBJECTIVES: Mitral annular calcification (MAC) may be a form of atherosclerosis and can lead to serious clinical consequences. The possible linkage between the presence and extent of MAC and coronary calcium score on CT is unknown. The goal of the present study was to investigate whether an association between MAC and coronary calcification (CC) exists in hypertensive patients with increased cardiovascular risk. METHODS: Five hundred twenty-two patients (284 men and 238 women, age range 52-80 years, mean 65+/-6 years), who were recruited to the INSIGHT study in the authors' region, underwent fast spiral CT of the heart as well as an echo Doppler examination. MAC was defined as advanced when the thickness of the calcium deposit was 5 mm or more; it was defined as trivial otherwise. RESULTS: The advanced MAC group comprised 62 patients, the trivial MAC group 215 patients, and the control group (without MAC) 245 patients. The prevalence of nonsevere CC was similar among the study groups, whereas the prevalence of severe CC (total calcium score >300) and the prevalence of proven coronary artery disease were associated with the presence and extent of MAC: respectively, 12% and 15% in control patients, 18% and 20% in patients with trivial MAC, and 29% and 29% in patients with advanced MAC. Multivariate analysis identified advanced MAC as an independent variable associated with severe CC and proved coronary artery disease. CONCLUSIONS: The results of this study demonstrated an association of advanced MAC and severe CC on spiral CT and proved coronary artery disease on the clinical level. Thus, advanced but not trivial MAC makes the noninvasive diagnosis of coronary atherosclerosis more likely and presumably could be considered as a new indication for further coronary evaluation in high-risk patients. PMID- 10719830 TI - Long-term effect of irradiation on lymph node uptake of interstitially delivered nanoparticulate contrast media. AB - RATIONALE AND OBJECTIVES: To characterize the long-term effects of therapeutic doses of ionizing radiation on the uptake and distribution of percutaneously delivered particulate contrast media in normal lymph nodes. METHODS: Two milliliters of an iodinated nanoparticle suspension (76 mg I/mL) was injected subcutaneously or submucosally into nine normal adult beagles. Region of interest analysis was used to estimate the volume, attenuation, and iodine concentration of opacified targeted lymph nodes and nonopacifled contralateral nodes on 24-hour postinjection CT images. All lymph nodes were then irradiated with 50 Gy in 25 fractions of 2 Gy/d. Contrast-enhanced quantitative CT was repeated 12 months after irradiation. RESULTS: Contrast-enhanced nodes averaged 2.3+/-0.8 times the volume of nonenhanced contralateral nodes before irradiation. The mean attenuation of contrast-enhanced nodes increased to 305 to 380 Hounsfield units from a pre-enhancement value of approximately 25 Hounsfield units. Opacified node volumes after irradiation averaged 61% to 86% of preirradiation volumes but were generally not statistically different. Contrast uptake assessed by average attenuation and iodine concentration decreased significantly by an average of 17% to 22% after irradiation and was significantly less than preirradiation uptake. Qualitatively, irradiated nodes generally appeared smaller than nonirradiated nodes, but the distribution pattern of contrast media did not appear to be appreciably altered. CONCLUSIONS: Lymph node irradiation resulted in only minimal decreases in contrast media uptake and node volume at 12 months. These effects presumably would not appreciably alter the potential clinical value of indirect lymphography for evaluating patients undergoing radiation therapy. PMID- 10719832 TI - Introduction to editorials on crises in academic medicine: diagnoses and treatments PMID- 10719831 TI - First experiences in using a new ultrasound mode and ultrasound contrast agent in the diagnosis of blunt renal trauma: a feasibility study in an animal model. AB - RATIONALE AND OBJECTIVES: To evaluate the diagnostic value of the new ultrasound mode "wide-band harmonic" (WBH) using an ultrasound contrast agent in blunt renal trauma in an animal model. METHODS: A defined blunt renal trauma was induced in 10 rabbits according to published standards. Ultrasound (B-mode, color and power Doppler, WBH) was performed before and after trauma, with and without using a contrast agent (Levovist). Ultrasound features were compared with histologic findings. RESULTS: In 2 of the 10 rabbits, three focal renal intraparenchymal lesions with diameters ranging from 1.0 to 1.8 mm were found that could be identified only using WBH with contrast. Six of the 10 rabbits developed a subcapsular hematoma with a thickness of up to 1.5 mm, which was identified by conventional B-mode as well as WBH. Histologic workup confirmed these findings of intraparenchymal hematomas and did not reveal further lesions. CONCLUSIONS: Only 20% of the experimental subjects developed parenchymal lesions with diameters of 1.0 mm or larger. All these lesions were identified only using WBH. These results indicate the potential to use WBH plus contrast for the diagnosis of blunt renal trauma. PMID- 10719833 TI - Medical education without physician scientists: answers without questions. PMID- 10719834 TI - Vitamin D and autoimmunity: is vitamin D status an environmental factor affecting autoimmune disease prevalence? AB - The environment in which the encounter of antigen with the immune system occurs determines whether tolerance, infectious immunity, or autoimmunity results. Geographical areas with low supplies of vitamin D (for example Scandinavia) correlate with regions with high incidences of multiple sclerosis, arthritis, and diabetes. The active form of vitamin D has been shown to suppress the development of autoimmunity in experimental animal models. Furthermore, vitamin D deficiency increases the severity of at least experimental autoimmune encephalomyelitis (mouse multiple sclerosis). Targets for vitamin D in the immune system have been identified, and the mechanisms of vitamin D-mediated immunoregulation are beginning to be understood. This review discusses the possibility that vitamin D status is an environmental factor, which by shaping the immune system affects the prevalence rate for autoimmune diseases such as multiple sclerosis, arthritis, and juvenile diabetes. PMID- 10719835 TI - Transport of toxic heavy metals across cell membranes. AB - Membrane transport of nonessential toxic heavy metals (type D heavy metals) not only controls their access to intracellular target sites but also helps determine their uptake, distribution, and excretion from the body. The critical role of membranes in the toxicology of class D metals has attracted the attention of many investigators, and extensive information has been collected on the mechanism(s) of metal transfer across membranes. Characteristics of metal transport in different cells, or even on opposite sides of the same cell, or under different physiological conditions, are not identical, and no unitary hypothesis has been formulated to explain this process in all cells. However, it seems possible that the mechanisms proposed for different cells represent variations on a few common themes. PMID- 10719836 TI - Tumor necrosis factor (TNF)-alpha and TNF receptors in viral pathogenesis. AB - Tumor necrosis factor-alpha (TNF-alpha) and TNF receptors (TNFR) are members of the growing TNF ligand and receptor families that are involved in immune regulation. The present report will focus on the role of the prototypic ligand TNF and its two receptors, TNFR1 and TNFR2, in viral pathogenesis. Although TNF was reported years ago to modulate viral infections, recent findings on the molecular pathways involved in TNFR signaling have allowed a better understanding of the molecular interactions between cellular and viral factors within the infected cell. The interactions of viral proteins with intracellular components downstream of the TNFR have highlighted at the molecular level how viruses can manipulate the cellular machinery to escape the immune response and to favor the spread of the infection. We will review here the role of TNF and TNFR in immune response and the role of TNF and TNFR signaling in viral pathogenesis. PMID- 10719837 TI - Serum leptin, lipids, free fatty acids, and fat pads in long-term dehydroepiandrosterone-treated Zucker rats. AB - The obese Zucker rat has a genetically flawed leptin system and is a model of hyperphagia, obesity, hyperlipidemia, and markedly elevated leptin levels. Dehydroepiandrosterone (DHEA) administration reduces hyperphagia, hyperlipidemia, and obesity in Zucker rats. Since serum leptin levels are associated with body fat, we wondered what the effects of fat pad weight reduction from DHEA administration would have on leptin levels. This experiment investigated the effects of DHEA on intra-abdominal fat pads, serum lipids, and peripheral leptin in male lean and obese Zucker rats that were administered DHEA in their food from 4 weeks of age to 20 weeks. Lean and obese rats received plain chow or chow containing DHEA. Additional chow-fed groups of lean and obese weight-matched controls and obese pair-fed rats helped to control for the reduced body weight, food intake, and fat pad weights seen with DHEA administration. DHEA administration to lean Zucker rats reduced body weight and fat pad weights, but leptin levels showed a lower trend. Among obese rats, both DHEA treatment and pair-feeding reduced body weight and fat pad weights, but only DHEA lowered leptin levels. The weight-matched controls had reductions in fat pad weights similar to the DHEA-treated group, but with increased leptin levels. Thus, DHEA may exert a small, independent effect on leptin levels in this animal model, but the reduction is less than what would be expected. PMID- 10719838 TI - Nitroprusside attenuates myocardial stunning through reduced contractile delay and time. AB - We hypothesized that myocardial stunning would be reversed through increased cyclic GMP caused by nitroprusside, and that this would be accomplished through a decreased proportion of regional work during diastole. Hearts were instrumented to measure left ventricular pressure, and regional myocardial mechanics were recorded using a miniature force transducer and ultrasonic dimension crystals in eight open-chest anesthetized dogs. Following baseline (CON), the left anterior descending coronary artery (LAD) was occluded for 15 min, followed by a 30-min recovery (STUN). Then intracoronary LAD infusion of sodium nitroprusside (NP) (4 microg/kg/ min) was begun. The time delay (msec) to regional shortening increased significantly from 18+/-13 to 73+/-13 following stunning, but was reduced to 49+/ 18 by NP. Total regional work (g*mm/min) at baseline (1368+/-401 CON) was unchanged with stunning (1320+/-333 STUN), but reduced (961+/-240) following NP. Time to peak force development (msec) increased significantly with stunning from 284+/-13 (CON) to 333+/-11 (STUN), but was reduced to 269+/-12 following NP. The percentage work during systole was reduced from 96%+/-2% (CON) to 77%+/-7% (STUN), but returned to 98%+/-1% with NP. Regional O2 consumption was unaffected by either treatment. Cyclic GMP was unchanged by stunning (2.9+/-0.3-2.9+/-0.4 pmol/g) but increased significantly with NP (4.6+/-0.6). These data indicated that regional myocardial stunning could be attenuated by nitroprusside, which increased cyclic GMP, decreased contractile delay, increased the proportion of work done during systole, and reduced time of shortening. PMID- 10719839 TI - Peptide YY stimulates the expression of apolipoprotein A-IV gene in Caco-2 intestinal cells. AB - The effect of peptide YY, a gastrointestinal hormone, on the expression of the apolipoprotein A-IV gene in the intestinal epithelial cell line Caco-2 was examined by semiquantitative RT-PCR followed by Southern hybridization with an inner oligonucleotide probe. Apolipoprotein A-IV mRNA levels were increased in response to peptide YY in a dose- and time-dependent fashion. Western blotting revealed that the exogenous peptide YY increased the intracellular concentration of apolipoprotein A-IV. In contrast, apolipoprotein A-I, B, and C-III mRNA did not respond to peptide YY. Differentiated Caco-2 cells expressed Y1- but not Y2- and Y5-receptor subtype mRNA. The present results suggest that peptide YY modulates apolipoprotein A-IV gene expression, likely via the Y1-receptor subtype in intestinal epithelial cells. PMID- 10719840 TI - Intestinal fat suppresses protein-induced exocrine pancreatic secretion in chronically bile-pancreatic juice-diverted rats. AB - Previously, we showed that the increase in pancreatic enzyme secretion was lower after feeding a casein diet containing fat than that after feeding a fat-free casein diet in chronically bile-pancreatic juice (BPJ)-diverted rats. In the present study, we determined whether the suppressive effects of fats on flow volume of BPJ and pancreatic enzyme secretion depend on delaying gastric emptying and examined the characteristics of the suppression with intraduodenal instillation of soybean oil or lecithin in BPJ-diverted rats. The study was conducted as three separate experiments using conscious rats with chronic BPJ diversion by means of a common bile-pancreatic duct catheter. The flow volume of BPJ and the secretion of pancreatic amylase and trypsin were determined after intraduodenal instillation of the test solution. Exocrine pancreatic secretion was strongly stimulated by administration of guanidinated casein hydrolysate (HGC, 150 mg/ml) in chronic BPJ-diverted rats. However, pancreatic secretion after administration of an emulsion containing HGC with either soybean oil (100 mg/ml) or mixed fat (50 mg/ml soybean oil + 50 mg/ml lecithin) was much lower than that after administration of HGC alone. In contrast, administration of the soybean oil emulsion without HGC resulted in a small, but significant increase in the volume of BPJ. The suppressive effects of soybean oil (100 mg/ml) on the increases in the BPJ flow and enzyme secretion were similar to those of sodium taurocholate (10 mg/ml), and there was no additive effect of soybean oil on taurocholate suppression. In conclusion, duodenally instilled soybean oil suppressed increases in flow volume of BPJ and pancreatic enzyme secretion induced by HGC in chronic BPJ-diverted rats, showing that the suppressive effect of the fat does not depend on delaying gastric emptying. PMID- 10719841 TI - Alterations in hypertrophic gene expression by dietary copper restriction in mouse heart. AB - Dietary copper (Cu) restriction causes a hypertrophic cardiomyopathy similar to that induced by work overload in rodent models. However, a possible change in the program of hypertrophic gene expression has not been studied in the Cu-deficient heart. This study was undertaken to fill that gap. Dams of mouse pups were fed a Cu-deficient diet (0.35 mg/kg diet) or a Cu-adequate control diet (6.10 mg/kg) on the fourth day after birth, and weanling mice continued on the dams' diet until they were sacrificed. After 5 weeks of feeding, Cu concentrations were dramatically decreased in the heart and the liver of the mice fed the Cu deficient diet. Corresponding to these changes, serum ceruloplasmin concentrations and hepatic Cu,Zn-superoxide dismutase activities were significantly (P<0.05) depressed. The size of the Cu-deficient hearts was greatly enlarged as estimated from the absolute heart weight and the ratio of heart weight to body weight. The abundances of mRNAs for atrial natriuretic factor, beta-myosin heavy chain, and alpha-skeletal actin in left ventricles were all significantly increased in the Cu- deficient hearts. Furthermore, Cu deficiency activated the expression of the c-myc oncogene in the left ventricle. This study thus demonstrated that a molecular program of alterations in embryonic genes, similar to that shown in the work-overloaded heart, was activated in the hypertrophied heart induced by Cu deficiency. PMID- 10719842 TI - Effects of oral administration of tamoxifen, toremifene, dehydroepiandrosterone, and vorozole on uterine histomorphology in the rat. AB - Tamoxifen, toremifene, DHEA, and vorozole inhibit tumor growth in rodent mammary carcinoma models and are promising chemotherapeutic agents for use against breast cancer development. In the present study, the effect of these agents on uterine histomorphology following oral administration to mature ovary-intact rats (n = 380) was examined. Animals received diet only (control), tamoxifen (0.4 and 1 mg/kg of diet; 10 mg/kg BW by daily gavage), toremifene (3-30 mg/kg of diet), DHEA (24-2000 mg/kg of diet), or vorozole (0.08-1.25 mg/kg BW by daily gavage) for 28 days and were either sacrificed or returned to a basal diet and then sacrificed 21 days later. Treatment with toremifene (all doses) or tamoxifen (1 and 10 mg/kg) for 28 days produced a decrease (P<0.05) in overall uterine size and myometrial thickness; however, uterine luminal and glandular epithelia cell height increased (P<0.05) compared with control. These compartmentalized uterotrophic and antiestrogenic effects of toremifene and tamoxifen were still apparent after 21 days post-treatment. Administration of DHEA (2000 mg/kg of diet) for 28 days had dramatic uterotrophic effects, increasing (P<0.05) overall uterine size and stimulating all three uterine compartments (epithelia, stroma, and myometrium). The other doses of DHEA, however, were not uterotrophic. Interestingly, after removal of DHEA from the diet, uterine weight and myometrial thickness decreased (P<0.05). Vorozole (1.25 mg/kg) administration for 28 days had differential, compartmentalized uterine effects, producing an increase (P<0.05) in epithelial cell height, a decrease (P<0.05) in stromal size, but no change in myometrial thickness. After 21 days postadministration of vorozole, luminal epithelial cell height was increased (P<0.05) compared with control. The data suggest that oral administration of tamoxifen, toremifene, DHEA, and vorozole results in differential, compartmentalized effects in the uterus that are highly dependent on treatment dose. The data may have implications for risk assessment of these agents prior to administration to healthy, cancer-free women. PMID- 10719843 TI - The effect of a special herbal tea on obesity and anovulation in androgen sterilized rats. AB - A special herbal tea has been used to treat clomiphene-resistant anovulatory disease and obesity effectively, especially in polycystic ovary syndrome (PCOS) cases with hyperinsulinemia. The effect of the herbal tea on obesity and anovulation was investigated in androgen-sterilized rats (ASR). The ASR model was established by subcutaneous injection of 1.25 mg testosterone propionate to Sprague-Dawley female rats at the age of 9 days. Rats were sacrificed around 112 days of age. ASR manifested with PCO, anovulation, high food intake, elevated body weight, and obesity. Immunocytochemistry demonstrated that estrogen receptors (ER) were predominantly distributed in the cytoplasm of neuropeptide Y (NPY)-containing neurons in the preoptic area (POA), and the coexpression was also found in the nuclei and fibers of NPY-synthesizing neurons in the arcuate nucleus (ARC). Compared with that in normal control rats, NPY expression was increased, the numbers of ER in hypothalamic ARC-median eminence (ME) decreased, gonadotropin-releasing hormone (GnRH) levels in ME was decreased, serum estrogen (E2) and leptin were elevated, and follicular stimulating hormone (FSH) and luteinizing hormone (LH) levels were reduced significantly in ASR. Significantly negative correlations between NPY and ER or GnRH, and between leptin and FSH or LH were observed. A positive correlation existed between serum leptin and body weight. These metabolic-endocrine changes in ASR were normalized after feeding the herbal tea. Both obesity and hypogonadotropin were expressed in ASR. The abnormal ovarian hormone milieu (elevated E2 levels) may have enhanced NPY expression and resulted in less GnRH and gonadotropin secretion. The herbal tea reduced body weight and induced ovulation in ASR. PMID- 10719844 TI - Beta-adrenergic agonist hyperplastic effect is associated with increased fibronectin gene expression and not mitogen-activated protein kinase modulation in C2C12 cells. AB - Beta-adrenergic agonists (beta-AA) enhance protein accretion in skeletal muscles. This stimulation is characterized by increased protein synthesis, increased expression of myofibrillar protein genes and a depression in protein degradation in animals, and increased proliferation and DNA synthesis in muscle cells in vitro. The mechanism or signal path in muscle whereby beta-AA would elicit these physiological effects upon binding to the G protein-coupled beta-adrenergic receptor (beta-AR) is unclear. C2C12 myoblasts were used to determine beta-AR ligand binding characteristics, cyclic AMP synthesis in response to isoproterenol (ISO) stimulation, and effects of ISO on DNA synthesis, mitogen activated protein kinase (MAPK), and fibronectin (FN) gene expression. Results showed that C2C12 cells possess beta-AR which are specific, saturable, and of high affinity (Kd = 0.2 nM). Forskolin and ISO stimulated cAMP production by = 20-fold (P<0.001) and 17-fold (P<0.001), respectively. ISO and the cAMP analog, 8-bromo-cAMP (8-BC) stimulated DNA synthesis in proliferating cells by 150% (P<0.05) and 200% (P<0.01), respectively, without modulating MAPK activity, whereas addition of fetal bovine serum to culture resulted in a 500% increase (P<0.01) in DNA synthesis and MAPK activation. DNA synthesis in C2C12 cells treated with ISO, 8 BC, or FBS was abolished in the presence of 25 microM PD098059, an MAPK-kinase inhibitor, suggesting that an MAPK-dependent pathway is likely involved in C2C12 proliferation. During cAMP elevating agent stimulation, basal MAPK activity may be sufficient, in the presence of other putative signaling molecules, to support proliferation in these cells. ISO or 8-BC treatment increased FN mRNA by three- and seven-fold, respectively, in growing C2C12 cells implying a connection between increased DNA synthesis and FN gene expression. PMID- 10719845 TI - Response of rat exocrine pancreas to high-fat and high-carbohydrate diets. AB - Intake of diets with high fat content is a risk factor for acute pancreatitis and pancreatic cancer. The underlying mechanisms leading to the development of these diseases due to high fat intake are currently unknown. The current study was designed in rats to determine the physiologic and pathological consequences of a highfat diet that contained excess amounts of cottonseed oil or a high carbohydrate diet that contained high amounts of sucrose on the exocrine pancreas. Rats were maintained on the diets for 4 weeks, and a cannula was inserted into the right jugular vein and one into the pancreatic duct for collection of pancreatic juice. Volume of the pancreatic juice and concentrations of amylase, lipase, and trypsinogen in the pancreatic juice were measured before and after infusions of CCK-8. Results showed that basal and CCK-stimulated pancreatic outputs of volume, amylase and lipase but not trypsinogen, were significantly elevated in intact rats given a high-fat diet when compared with rats given a high-carbohydrate diet. Forty-eight hours later, rats were sacrificed, and parts of the pancreas were removed for isolation of pancreatic acinar cells and for histopathologic studies. Pancreatic acini isolated from rats on a high-fat diet showed significantly lower basal and CCK-stimulated amylase release when compared with those on a high-carbohydrate diet. Histology of the pancreas of rats on a high-carbohydrate diet appeared normal; however, the pancreas of rats on high-fat diet showed significant alterations in exocrine pancreas. These results showed abnormalities in the exocrine pancreas of rats on a high-fat diet, that were not found in rats on a high-carbohydrate diet; further, they support the contention that a high-fat diet has a deleterious effect on the pancreas. PMID- 10719846 TI - Genomic changes and HPV type in cervical carcinoma. AB - To identify chromosomal regions that may include the loci of abnormally expressed cellular genes and may be specifically altered depending on the histological subtype of the tumor, we studied primary cervical carcinoma using CGH and HPV genotyping. Eighty-seven percent of the primary tumors were positive for DNA of a "high-risk" HPV type (e.g., 16 or 18). In the cervical carcinomas, without reference to histologic subtype, overrepresentation of chromosome 3q was the most consistent chromosomal aberration with underrepresentation of chromosome 3p also a frequent finding. Chromosome arms 1q, 5p, 20q, and Xq were overrepresented in many tumors and 3p loss and 5p, 8q, and 16q gain were only associated with squamous cell carcinoma in this series. PMID- 10719847 TI - Molecular keys to the problems of cerebral vasospasm. AB - The mechanisms responsible for subarachnoid hemorrhage (SAH)-induced vasospasm are under intense investigation but remain incompletely understood. A consequence of SAH-induced vasospasm, cerebral infarction, produces a nonrecoverable ischemic tissue core surrounded by a potentially amenable penumbra. However, successful treatment has been inconsistent. In this review, we summarize the basic molecular biology of cerebrovascular regulation, describe recent developments in molecular biology to elucidate the mechanisms of SAH-induced vasospasm, and discuss the potential contribution of cerebral microcirculation regulation to the control of ischemia. Our understanding of the pathogenesis of SAH-induced vasospasm remains a major scientific challenge; however, molecular biological techniques are beginning to uncover the intracellular mechanisms involved in vascular regulation and its failure. Recent findings of microvascular regulatory mechanisms and their failure after SAH suggest a role in the development and size of the ischemia. Progress is being made in identifying the various components in the blood that cause SAH-induced vasospasm. Thus, our evolving understanding of the underlying molecular mechanism may provide the basis for improved treatment after SAH induced vasospasm, especially at the level of the microcirculation. PMID- 10719849 TI - Targeted toxin therapy for malignant astrocytoma. AB - The poor prognosis associated with malignant astrocytoma has led investigators to seek new, innovative methods of treatment. Targeted toxins represent a unique form of therapy that has two components, a carrier molecule with high specificity for tumor-associated antigens and a potent protein toxin. These compounds are extremely cytotoxic to malignant astrocytoma cell lines in vitro. Animal studies have shown prolongation of survival and complete tumor regression when targeted toxins were administered by a variety of routes. The promising results seen in vivo have formed the basis for proceeding with clinical trials in humans with leptomeningeal neoplasia and malignant brain tumors, in which these agents are administered intrathecally or directly into tumor, respectively. To date, in these clinical trials, targeted toxins have been delivered safely without significant neurological toxicity, and cytological analysis of cerebrospinal fluid and radiological findings have shown evidence of a therapeutic response. These studies have confirmed the existence of a therapeutic window between normal brain tissue and malignant cells that can be exploited with targeted therapy directed against the transferrin receptor. The successful delivery of targeted toxins directly into malignant brain tumors has established this route of administration as practical and feasible. Identification of other receptors that are preferentially expressed on brain tumors, such as the interleukin-4 receptor, has resulted in the creation of a fusion protein against this receptor that contains a modified toxin from the bacteria Pseudomonas aeruginosa. This chimeric fusion toxin is currently under investigation in a Phase I clinical trial with patients with recurrent malignant astrocytoma, and other targeted toxins are under development for the treatment of these uniformly fatal tumors. Owing to these recent advances in targeted toxin therapy for malignant primary brain tumors, a review of the development of these agents for practicing neurosurgeons seems timely. PMID- 10719848 TI - Radiosurgery: where we were, are, and may be in the third millennium. AB - Radiosurgery will celebrate its Golden Jubilee in the year 2001. More than 100,000 patients throughout the world have undergone radiosurgery since Lars Leksell first described the technique in 1951. Rapid developments in neuroimaging and even robotic technology in the past decade have contributed to improved outcomes and wider applications for radiosurgery. A variety of different radiosurgical techniques have been developed in the past two decades. Numerous studies have examined the benefits and risks of radiosurgery performed with various devices. The long-term results of radiosurgery are now available, and these results have established radiosurgery as an effective noninvasive treatment method for intracranial vascular malformations and many tumors. Additional applications of radiosurgery for the treatment of malignant tumors and functional disorders are being assessed. Radiosurgery is an impressive combination of minimally invasive technologies administered by a multidisciplinary team of surgeons, oncologists, medical physicists, and engineers. PMID- 10719850 TI - Transsphenoidal surgery for Cushing's disease: outcome in patients with a normal magnetic resonance imaging scan. AB - OBJECTIVE: Transsphenoidal surgery for Cushing's disease from a pituitary adenoma is an effective and safe treatment. Definitive preoperative diagnosis of Cushing's disease caused by a pituitary adenoma is often difficult, particularly in patients with normal imaging studies and a normal sella turcica. We present the outcome of transsphenoidal surgery in patients with presumed Cushing's disease and a normal pituitary magnetic resonance imaging scan. METHODS: Between January 1992 and December 1997, 105 patients underwent transsphenoidal surgery for Cushing's disease at our institution. The criteria for inclusion in this study were clinical and biochemical studies strongly suggestive of Cushing's disease, a normal magnetic resonance imaging scan with normal sella and sellar contents, no previous pituitary surgery, and transsphenoidal surgery performed at this institution. Eighteen patients fulfilled these criteria, and their results were analyzed retrospectively. RESULTS: The average age of the patients was 47.8 years; there were 13 women and 5 men. Inferior petrosal sinus sampling with and without corticotropin-releasing hormone stimulation was performed in 16 patients with correct localization of the lesion in 13 (81%). During surgery, the surgeon identified and removed 17 pituitary tumors; 15 patients had selective adenomectomies, one had a hemihypophysectomy, and two had total hypophysectomies. Thirteen discrete adrenocorticotropic hormone-secreting adenomas were proven histologically, and one pituitary gland had diffuse involvement with tumor. Complications occurred in five patients. Sixteen patients who were followed up for an average of 21.6 months had sustained remission, 12 of whom were profoundly hypocortisolemic immediately after surgery. CONCLUSION: In patients with Cushing's disease and a normal magnetic resonance imaging scan, an experienced surgeon can perform transsphenoidal surgery resulting in effective removal of very small microadenomas, with clinical and biochemical remission in the majority. Inferior petrosal sinus sampling is helpful in localizing the adenoma. PMID- 10719851 TI - Cystic schwannomas of the jugular foramen: clinical and surgical remarks. AB - OBJECTIVE: The goals of this report were to outline the clinical presentation, radiological characteristics, surgical techniques, postoperative morbidity, and long-term follow-up results for cystic jugular foramen (JF) schwannomas and to describe their differences, compared with solid schwannomas involving the JF. METHODS: A retrospective analysis of radiological studies and surgical records identified five primarily cystic tumors among 21 cases of JF schwannomas that had been surgically treated at our institution. RESULTS: Two types of cystic JF schwannomas were observed, i.e., Type 1 lesions, which are single large cysts with thin ring-like enhancement of the tumor wall, and Type 2 lesions, which are multiple cysts with very irregular, thick enhancement of the cyst wall. The most common symptoms were hearing loss, ataxia, and headaches. Total surgical removal could be performed in all cases. The immediate postoperative findings indicated hearing improvement in three cases. No deterioration of lower cranial nerve function was observed. All patients were independent in the immediate postoperative period and in the long-term follow-up period (Karnofsky Performance Scale score, 90). CONCLUSION: Surgical treatment of cystic JF schwannomas can be very demanding because of generally stronger adhesion of the tumor capsule to the surrounding structures, fragile tumor capsules, and difficulty in identification of the arachnoidal planes in some cases. Early identification of the arachnoidal planes without opening of the cyst and sharp dissection may be useful. Careful intradural opening of the JF should be performed to achieve total removal of the last tumor portion within the JF. A comparison of these lesions with solid schwannomas involving the JF indicated that cystic tumors affected a younger population, with less preoperative swallowing impairment (P < 0.05). The immediate postoperative course in both types of cystic JF schwannomas was usually better than for solid lesions, because of minor postoperative cranial nerve morbidity, especially involving lower cranial nerve function, in the latter cases. Long-term follow-up data failed to demonstrate any significant differences in final patient outcomes, however. PMID- 10719852 TI - Long-term outcome and growth rate of subtotally resected petroclival meningiomas: experience with 38 cases. AB - OBJECTIVE: To evaluate the long-term outcome of a subtotally resected residual tumor and to assess its growth rate, we analyzed the records of 38 patients with residual petroclival meningioma. METHODS: Clinical records and radiological findings of 38 cases of petroclival meningioma that were diagnosed and subtotally resected at Seoul National University Hospital between 1981 and 1997 were carefully reviewed. Follow-up imaging studies were reviewed, and Karnofsky performance scale scores at the last follow-up were recorded. The duration of follow-up ranged from 6 to 141 months (mean, 47.5 mo; median, 30 mo). Tumor progression and progression-free survival rates were assessed. The growth rate of a residual tumor was evaluated by measuring the equivalent diameter and the tumor volume serially; the tumor doubling time was calculated, and the predictive factors for determining the growth pattern in residual tumors and the prognosis were analyzed. RESULTS: In 33 (87%) of the 38 patients, Karnofsky performance scale scores at the last follow-up were 80 or above. The median progression-free survival time among patients with subtotally resected tumors was 66 months, and the 5-year progression-free survival rate was 60%. The growth rate of residual tumors was low (volume increase, 4.94 cc/yr; diameter increase, 0.37 cm/yr). The mean tumor doubling time was 8 years. Although there were no significant predictive factors, age and extent of tumor resection seemed to influence the progression-free survival rate. Significant factors affecting the growth rate were age and occurrence of menopause. CONCLUSION: Subtotal resection with or without radiation or radiosurgery should be considered as a suitable treatment option for patients with petroclival meningiomas, especially the elderly, because the growth rate of residual tumors is low. PMID- 10719853 TI - Stereotactic radiosurgery for tumor-related trigeminal pain. AB - OBJECTIVE: Between 1 and 6% of patients who are diagnosed with facial pain syndromes have tumors that involve the trigeminal nerve. We report the effects of stereotactic radiosurgery on tumor-related trigeminal pain. METHODS: We reviewed results, from a prospective database, for 24 consecutive patients with cranial base tumors and either trigeminal neuralgia (n = 9) or painful trigeminal neuropathy (n = 15) who underwent stereotactic radiosurgery during an 8-year period. The tumor was the radiosurgical target for these patients (not the trigeminal nerve or ganglion). The median clinical follow-up period after radiosurgery was 45 months (range, 12-90 mo); the median neuroimaging follow-up period was 36 months (range, 5-86 mo). RESULTS: There were 20 women and 3 men, with an average age of 57 years (range, 33-79 yr). One patient had bilateral facial pain and underwent staged radiosurgery. Pathological classification indicated 16 meningiomas and 8 malignant cranial base tumors (adenoid cystic carcinoma, n = 6; squamous cell carcinoma, n = 2). Twelve of 24 patients (50%) were initially free of pain, and another 11 patients (46%) reported that they experienced significant improvements in their trigeminal pain syndromes after radiosurgery. The tumor histological type, quality of facial pain, preexisting facial numbness, and marginal and maximal radiation doses were not related to postradiosurgical facial pain outcomes. Three patients with malignant cranial base carcinomas developed recurrent facial pain, 1 to 9 months after radiosurgery, which was related to tumor progression outside the irradiated volume. One patient (4%) developed new partial V2 numbness after radiosurgery. CONCLUSION: Radiosurgery proved to be effective in improving tumor-related trigeminal pain for the majority of patients with either benign or malignant cranial base tumors. Recurrence of trigeminal pain is frequent for patients with malignant cranial base carcinomas and is related to tumor progression. PMID- 10719854 TI - Quadrigeminal variant of perimesencephalic nonaneurysmal subarachnoid hemorrhage. AB - OBJECTIVE: Perimesencephalic nonaneurysmal subarachnoid hemorrhage (PNSH) is a benign entity with a low risk of rebleeding. The most widely accepted definition emphasizes the presence of blood ventral to the midbrain or pons on early computed tomography. We sought to determine the frequency of PNSH with blood centered in the quadrigeminal cistern. METHODS: We reviewed a prospectively collected database of all patients admitted to our institution over a 2.5-year period with subarachnoid hemorrhage (SAH) and identified PNSH patients from early computed tomographic scans and negative four-vessel angiograms. RESULTS: Of 220 SAH patients, we identified 9 with PNSH. Two (22%) of these patients had SAH centered in the quadrigeminal cistern without pretruncal blood, negative repeat angiograms, and an uncomplicated clinical course. CONCLUSION: Quadrigeminal SAH is a variant of PNSH that is not well described in the literature. It may comprise up to one-fifth of PNSH cases and carries a similar benign prognosis. PMID- 10719855 TI - Posttreatment sequelae of palliatively treated cerebral arteriovenous malformations. AB - OBJECTIVE: Posttreatment sequelae of palliatively treated cerebral arteriovenous malformations (AVMs) were studied to evaluate the significance of these nonradical treatments. METHODS: Between 1987 and 1997, 46 patients with cerebral AVMs were managed with treatments such as partial embolization, radiosurgery, subtotal resection, or feeder ligation alone. Their AVMs were not radically resected because of difficulties in radical treatment, hesitance to treat eloquent area lesions, or residual nidi after subtotal obliteration. The patients' posttreatment sequelae were evaluated. The duration of follow-up ranged from 0.5 to 169 months (mean, 49.4+/-39.8 mo). RESULTS: Twenty-six bleeding episodes from AVMs were documented in 18 patients. The annual risk of bleeding in this palliatively treated group was 14.6%. Persistent progressive neurological deficit was observed in one patient. Major neurological deficits occurred in 10 patients (23.3%), and the mortality rate was 9.3%. CONCLUSION: Palliative treatments cannot prevent bleeding and may even worsen the posttreatment course compared with the natural history of cerebral AVMs. A more conservative indication is required in recommending palliative treatment alone. PMID- 10719856 TI - Safety and feasibility of continuous infusion of remifentanil in the neurosurgical intensive care unit. AB - OBJECTIVE: Remifentanil is a selective mu-opioid agonist with a context-sensitive half-time of 3 to 5 minutes, independent of dose or administration duration. Other desirable effects include decreased cerebral metabolism and intracranial pressure (ICP) with minimal cerebral perfusion pressure changes. We present six cases illustrating indications for the use of remifentanil in the neurosurgical intensive care unit. METHODS: Patients received bolus doses of remifentanil of 0.05 to 1.0 microg/kg, followed by continuous infusions of 0.03 to 0.26 microg/kg/min, titrated to effect. When infusions were discontinued for neurological examinations, another bolus dose preceded infusion reinstitution. Indications for the use of remifentanil included mean arterial pressure and cerebral perfusion pressure decreases with the use of other agents (e.g., codeine or propofol) for ICP control, elevated ICP that was refractory to propofol/mannitol treatment, agitation that was unresponsive to standard therapies, and coughing that caused ICP increases after subarachnoid hemorrhage. RESULTS: Three patients experienced spontaneous intracranial bleeding (two cases of subarachnoid hemorrhage and one case of intraventricular hemorrhage), and three patients exhibited severe traumatic subdural hemorrhage. All patients recovered from the effects of remifentanil within 3 minutes after discontinuation of infusion, which allowed frequent rapid neurological assessments. Procedures for pulmonary toilet (i.e., endotracheal suctioning, postural drainage, and bronchoscopy) were performed without deleterious ICP increases or mean arterial pressure or cerebral perfusion pressure decreases during remifentanil infusions. CONCLUSION: The ultrashort duration of action of remifentanil allowed easy performance of frequent neurological examinations in the neurosurgical intensive care unit. No patient experienced deleterious hemodynamic or neurological effects as a result of remifentanil use. PMID- 10719857 TI - Intrathecal baclofen for intractable cerebral spasticity: a prospective placebo controlled, double-blind study. AB - OBJECTIVE: To conduct a placebo-controlled prospective study of the effectiveness of intrathecal bolus injections and continuous administration of baclofen on functional parameters in patients with severe spasticity of cerebral origin. To compare this functional evaluation with spasticity scores in different muscle groups. METHODS: In 11 patients with spasticity of cerebral origin (mainly cerebral palsy), double-blind scoring of spasticity (Ashworth scale score and visual analog score), spasms, pain, and functional abilities was performed during tests with bolus injections including a placebo control. Eight patients were considered good responders and received a subcutaneous device for intrathecal drug delivery. Six of these patients were followed up for 2 years, during which they underwent the same scoring procedures as after their bolus injections. These patients were subjected to a blinded dose reduction test. RESULTS: There was a noticeable placebo effect on spasticity scores during tests with bolus injections. Eight patients demonstrated a significant beneficial effect of intrathecal bolus injections compared with this placebo effect. Functional improvements were noted in most patients. During continuous infusion, Ashworth scale scores were less favorable but still significantly lower than at baseline. Subjective evaluation (visual analog scores) remained positive, functional improvements were maintained, and patient comfort was invariably and significantly improved. CONCLUSION: Intrathecal administration of baclofen is a safe and effective treatment for spasticity of cerebral origin. Functional improvement was demonstrated. The presence of a placebo effect on the spasticity scores suggests the need for double-blind screening in each patient. PMID- 10719858 TI - Pallidotomy microelectrode targeting: neurophysiology-based target refinement. AB - OBJECTIVE: Microelectrode recording can refine targeting for stereotactic radiofrequency lesioning of the globus pallidus to treat Parkinson's disease. Multiple intraoperative microelectrode recording/stimulating tracks are searched and assessed for neuronal activity, presence of tremor cells, visual responses, and responses to kinesthetic input. These physiological data are then correlated with atlas-based anatomic data to approximate electrode location. On the basis of these physiological properties, one or more tracks are selected for lesioning. This study analyzes the track physiological factors that seem most significant in determining the microelectrode recording track(s) that will be chosen for pallidal lesioning. METHODS: Thirty-six patients with Parkinson's disease underwent microelectrode-guided pallidotomy. Between one and five microelectrode recording tracks were made per patient. Usually, one (n = 23) or two (n = 12) of these tracks were lesioned. Electrode positions in the x (mediolateral) and y (anteroposterior) axes were recorded and related to track neurophysiological findings and final lesion location. The stereotactic location and sequence of microelectrode tracks were recorded and plotted to illustrate individual search patterns. These patterns were then compared with those noted in other patients. Neurophysiological data obtained from recording tracks were analyzed. A retrospective analysis of track electrophysiology was performed to determine the track characteristics that seemed most important in the surgeon's choice of the track to lesion. Track physiological properties included general cell spike amplitude, tremor synchronous neuronal firing, kinesthetically responsive neuronal firing, and optic track responses (either phosphenes reported by the patient during track microstimulation or neuronal firing in response to light stimulus into the patient's eyes). Orthogonally corrected postoperative magnetic resonance images were used to confirm the anatomic lesion locations. RESULTS: In patients who had a single mapped track lesioned, specific track electrophysiological characteristics identified the track that would be lesioned most of the time (20 of 24 patients). Tracks that exhibited a combination of tremor synchronous firing, joint kinesthesia, and visual responsivity were lesioned 17 (85%) of 20 times. Analysis of intraoperative electrode movement in the x and y axes indicated a significant subset of moves but did not result in microelectrode positioning closer to the subsequently lesioned track. Accuracy of initial electrode movement in the x and y axes was most highly correlated with a measure of first-track electrophysiological activity. The number of microelectrode recording tracks did not correlate with clinical outcome. Anatomic analysis, using postoperative magnetic resonance imaging, revealed that all lesions were placed in the globus pallidus. Most patients (35 of 36) improved after surgery. CONCLUSION: The level of electrophysiological activity in the first track was the best predictive factor in determining whether the next microelectrode move would be closer to the ultimately lesioned track. The analysis of electrode track location and neurophysiological properties yields useful information regarding the effectiveness of microelectrode searching in the x and y axes. Within an institution, the application of this modeling method may increase the efficiency of the microelectrode refinement process. PMID- 10719859 TI - Visual loss after spine surgery: a survey. AB - OBJECTIVE: To survey a large number of neurosurgical spine surgeons for data regarding the presence of risk factors in patients experiencing visual loss after spine surgery. METHODS: A survey was sent to current members (as of 1997) of the American Association of Neurological Surgeons/Congress of Neurological Surgeons, Section on Disorders of the Spine and Peripheral Nerves, with questions focusing on intraoperative factors that may predispose patients to perioperative visual loss. RESULTS: Two hundred ninety surveys were returned, and 24 patients with visual loss after spine surgery were reported by 22 surgeons. Although many of these patients had probable causative factors for visual loss after surgery (e.g., hypotension, low hematocrit level, coexisting disease), some did not (n = 8). CONCLUSION: These results suggest the necessity of a high index of suspicion for evolving perioperative visual loss even in the absence of risk factors. PMID- 10719860 TI - Safety, efficacy, and functionality of high-field strength interventional magnetic resonance imaging for neurosurgery. AB - OBJECTIVE: Interventional magnetic resonance imaging (MRI) allows neurosurgeons to interactively perform surgery using MRI guidance. High-field strength (1.5-T) imaging permits exceptional observation of intracranial and spinal pathological features. The development of this technology and its application to a variety of neurosurgical procedures are described. METHODS: We report on the first 101 cases that were treated in the interventional MRI unit (between January 1997 and September 1998). These cases included 39 brain biopsies, 30 tumor resections, 9 functional neurosurgical cases, 8 cyst drainages, 5 laminectomies, and 10 miscellaneous cases. Patients ranged in age from 14 months to 84 years (median, 43 yr); 61 patients were male and 40 were female. Intraoperative functional techniques that were used to influence surgical decision-making included magnetic resonance spectroscopy, functional MRI, magnetic resonance angiography and venography, chemical shift imaging, and diffusion-weighted imaging. All surgery was performed using MRI-compatible instruments within the 5-gauss line and conventional instruments outside that line. RESULTS: All 39 brain biopsies yielded diagnostic tissue. Of the 30 tumor resections, 24 (80%) were considered radiographically complete. The incidence of serious complications was low and was comparable to that associated with conventional operating rooms. One patient developed a Propionibacterium acnes brain abscess 6 weeks after surgery and another patient experienced Staphylococcus aureus scalp cellulitis after a brain biopsy, yielding an infection rate of less than 2%. No clinically significant hemorrhage was observed in immediate postoperative imaging scans, although one patient developed a delayed hematoma after a thalamotomy. One patient experienced a stroke after resection of a hippocampal tumor. No untoward events were associated with MRI-compatible instrumentation or intraoperative patient monitoring. CONCLUSION: High-field (1.5-T) interventional MRI is a safe and effective technology for assisting neurosurgeons in achieving the goals of surgery. Preliminary results suggest that the functional capabilities of this technology can yield data that can significantly influence intraoperative neurosurgical decision-making. The rates of serious complications, such as infection, associated with this new technology were low. PMID- 10719861 TI - Magnetic resonance imaging-guided neurosurgery in the magnetic fringe fields: the next step in neuronavigation. AB - OBJECTIVE: We describe the development of an alternative approach to intraoperative magnetic resonance imaging (iMR)-guided neurosurgery and report our initial experience with 22 craniotomies and 16 brain biopsies. The advantages and disadvantages of each approach are examined. METHODS: An iMR suite houses a 0.2-T open configuration system (Siemens Medical Systems, Erlangen, Germany) and is equipped with anesthetic gases and a magnetic resonance imaging (MRI) compatible anesthesia machine and monitor. Standard operating instruments and equipment were tested for safety and compatibility in the magnetic fringe fields surrounding the open MRI system. We then performed brain biopsies and craniotomies in the iMR suite. RESULTS: Standard operating equipment functioned properly in the 0.5- to 10-mT zone and was not affected by the magnet's attractive force. Twenty-two craniotomies and 16 brain biopsies were performed in the interventional suite, using serial intraoperative MRI guidance, without injury to patients or operating room staff. CONCLUSION: Full neurosurgical procedures may be performed in the weak fringe fields surrounding an MRI system, using standard operating room equipment. This approach to iMR-guided neurosurgery offers a significant cost advantage over retrofitting an entire operative suite with "MRI-compatible" surgical equipment. The surgeon's familiarity with standard equipment and the reliability of the equipment are additional advantages. Neurosurgery in the fringe fields allows the neurosurgeon to utilize serial MRI with a minimum of inconvenience, disruption, and change to the standard neurosurgical procedure. Serial intraoperative imaging to visualize the changes in the brain that are associated with neurosurgical intervention seems to enhance the ability to safely and effectively accomplish neurosurgical goals. PMID- 10719862 TI - Endoscopic colloid cyst surgery. AB - OBJECTIVE: The purpose of this report is to discuss the technical aspects of operating on colloid cysts through a transventricular approach, with rigid endoscopes. METHODS: Twelve patients underwent 14 endoscopic operations in attempts to treat their colloid cysts. All patients were symptomatic, with headache being the most common complaint (8 of 12 patients). Six patients in this series exhibited enlarged ventricles associated with their colloid cysts. Using rigid endoscopes of < or =3.5-mm diameter, the cysts were inspected and fenestrated. Both hard and soft cyst contents were evacuated, and then the walls of the cysts were coagulated inside and outside. External ventriculostomy tubes were usually placed. Technical obstacles to successful completion of endoscopic colloid cyst surgery are discussed. RESULTS: For 11 of the 12 patients, the colloid cysts could be treated via an endoscopic approach. The mean follow-up time was 173 weeks, and the median follow-up time was 125 weeks. For the 12th patient, bilateral scarring of the foramina of Monro precluded direct surgery; therefore, a septostomy was performed and a ventriculoperitoneal shunt was placed. CONCLUSION: Endoscopic transventricular surgery should be considered for the treatment of colloid cysts. PMID- 10719863 TI - Multicolumn infusion of gene therapy cells into human brain tumors: technical report. AB - OBJECTIVE: Effective gene therapy for brain tumors may require saturation of the tumors with tumoricidal doses of the therapeutic gene. Safe, precise, and efficient delivery of gene therapy vectors is required. Most reported cases of and published protocols for gene therapy for brain tumors involve freehand injection of retroviral vector-producing cells (VPCs) into the brain. Major disadvantages of this method include the inaccuracy of hand-guided needle placement and limited control of injection parameters. These factors can result in failure to deliver the viral vectors to specifically targeted sites within the brain, extensive tissue disruption resulting from excessively forceful injection, and reflux of the injectate along the needle tract. METHODS: We describe a novel stereotactic strategy for saturating tumor volumes with tumoricidal doses of gene therapy vectors and a new, more precise method of infusing VPCs. With our new instrument, the multicolumn stereotactic infusion system, needle placement is stereotactically guided and both VPC infusion and needle withdrawal are mechanically controlled. RESULTS: This technique, which has been used effectively for six patients, permits precise deposition of columns of VPCs throughout the targeted tumor volume. CONCLUSION: This technique should facilitate saturation of tumors with tumoricidal doses of gene therapy vectors and should improve the results of gene therapy protocols that rely on intraparenchymal injection for delivery. PMID- 10719864 TI - Microsurgical anatomic features and nomenclature of the paraclinoid region. AB - OBJECTIVE: We describe the detailed microsurgical anatomic features of the clinoid (C5) segment of the internal carotid artery (ICA) and surrounding structures, clarify the anatomic relationships of structures in this region, and emphasize the clinical relevance of these observations. Furthermore, because the nomenclature of the paraclinoid region is confusing and lacks standardization, this report provides a glossary of terms that are commonly used to descibe the anatomic features of the paraclinoid region. METHODS: The region surrounding the anterior clinoid process was observed in 70 specimens from 35 formalin-fixed cadaveric heads. Detailed microanatomic dissections were performed in 10 specimens. Histological sections of this region were obtained from the formalin fixed cadaveric specimens. RESULTS: The clinoid segment of the ICA is the portion that abuts the clinoid process. This portion of the ICA can be directly observed only after removal of the clinoid process. The dura of the cavernous sinus roof separates to enclose the clinoid process. The clinoid segment of the ICA exists only where this separation of dural layers is present. Because the clinoid process does not completely enclose the ICA in most cases, the clinoid segment is shaped more like a wedge than a cylinder. The outer layer of the dura (dura propria) is a thick membrane that fuses with the adventitia of the ICA to form a competent ring that separates the intradural ICA from the extradural ICA. The thin inner membranous layer of the dura loosely surrounds the ICA throughout the entire length of its clinoid segment. The most proximal aspect of this membrane defines the proximal dural ring. The proximal ring is incompetent and admits a variable number of veins from the cavernous plexus that accompany the ICA throughout its clinoid segment. CONCLUSION: The narrow space between the inner dural layer and the clinoid ICA is continuous with the cavernous sinus via an incompetent proximal dural ring. This space between the clinoid ICA and the inner dural layer contains a variable number of veins that directly communicate with the cavernous plexus. Given the inconstancy of the venous plexus surrounding the clinoid ICA, we think that categorical labeling of the clinoid ICA as intracavernous or extracavernous cannot be justified. PMID- 10719865 TI - Intracerebral clysis in a rat glioma model. AB - OBJECTIVE: Intracerebral clysis (ICC) is a new term we use to describe convection enhanced microinfusion into the brain. This study establishes baseline parameters for preclinical, in vivo, drug investigations using ICC in a rat glioma model. METHODS: Intracranial pressure was measured, with an intraparenchymal fiber-optic catheter, in male Fischer rats 10, 15, 20, and 25 days after implantation of C6 glioma cells in the right frontal lobe (n = 80) and in control rats without tumor (n = 20), before and during ICC. A 25% albumin solution (100 microl) was infused through an intratumoral catheter at 0.5, 1.0, 2.0, 3.0, and 4.0 microl/min. Infusate distribution was assessed by infusion of fluorescein isothiocyanate dextran (Mr 20,000), using the aforementioned parameters (n = 36). Brains were sectioned and photographed under ultraviolet light, and distribution was calculated by computer analysis (NIH Image for Macintosh). Safe effective drug distribution was demonstrated by measuring tumor sizes and apoptosis in animals treated with N,N'-bis(2-chloroethyl)-N-nitrosourea via ICC, compared with untreated controls. Magnetic resonance imaging noninvasively confirmed tumor growth before treatment. RESULTS: Intracranial pressure increased with tumor progression, from 5.5 mm Hg at baseline to 12.95 mm Hg on Day 25 after tumor cell implantation. Intracranial pressure during ICC ranged from 5 to 21 mm Hg and was correlated with increasing infusion volumes and increasing rates of infusion. No toxicity was observed, except at the higher ends of the tumor size and volume ranges. Fluorescein isothiocyanate-dextran distribution was greater with larger infusion volumes (30 microl versus 10 microl, n = 8, P < 0.05). No significant differences in distribution were observed when different infusion rates were compared while the volume was kept constant. At tolerated flow rates, the volumes of distribution were sufficient to promote adequate drug delivery to tumors. N,N' Bis(2-chloroethyl)-N-nitrosourea treatment resulted in significant decreases in tumor size, compared with untreated controls. CONCLUSION: The C6 glioma model can be easily modified to study aspects of interstitial delivery via ICC and the application of ICC to the screening of potential antitumor agents for safety and efficacy. PMID- 10719866 TI - Calcium channel antagonist effect on in vitro meningioma signal transduction pathways after growth factor stimulation. AB - OBJECTIVE: We have previously demonstrated that calcium channel antagonists inhibit the growth of human meningiomas in culture after stimulation with growth factors. This study examined the effects of these drugs on signaling transduction pathways in an attempt to elucidate potential mechanisms by which this growth inhibition is mediated. METHODS: Primary cell cultures from patients with intracranial meningiomas were established. Cell growth studies were performed with inhibitors and stimulators of tyrosine kinase signal transduction. Intracellular calcium changes and inositol phosphate production were measured after growth factor exposure, with or without pretreatment by calcium channel antagonists. RESULTS: The growth of meningiomas in culture can be inhibited by tyrosine kinase receptor inhibitors. Inhibitors and stimulators of phospholipase C can stimulate or inhibit the growth of in vitro meningiomas, respectively. Calcium channel antagonists inhibit intracellular calcium changes induced by serum and epidermal growth factor. Inositol phosphate production is increased after growth factor stimulation, and calcium channel antagonists potentiate this effect. CONCLUSION: Calcium channel antagonists interfere with intracellular signaling pathways of cultured meningioma cells. This inhibition is unrelated to voltage-sensitive calcium channels. The findings of this project may aid in the understanding of the signal transduction mechanisms involved in growth factor mediated meningioma proliferation and may lead to clinically relevant strategies for growth inhibition. PMID- 10719867 TI - Enhanced delivery improves the efficacy of a tumor-specific doxorubicin immunoconjugate in a human brain tumor xenograft model. AB - OBJECTIVE: To evaluate dose intensification with osmotic blood-brain barrier disruption (BBBD) and the potential use of drug targeting with monoclonal antibody (MAb) BR96 conjugated to doxorubicin (BR96-DOX, now called SGN15) for treatment of intracerebral and subcutaneous human LX-1 small cell lung carcinoma xenografts in rats. METHODS: LX-1 tumors with high, low, or heterogeneous levels of the Lewis(y) antigen for BR96 were evaluated. Rats were treated with intracarotid or intravenous BR96-DOX, with or without osmotic BBBD. RESULTS: Both BR96-DOX and MAb BR96 treatment resulted in significant regression of subcutaneous tumors, in contrast to control groups including doxorubicin alone, saline, or nonbinding doxorubicin immunoconjugate. BR96-DOX delivered with BBBD to brain tumors with low antigen expression resulted in significantly (P < 0.001) increased rat survival time compared with animals that received intravenous or intra-arterial BR96-DOX. CONCLUSION: The combination of an effective drug such as doxorubicin with a MAb to facilitate tumor-selective localization and osmotic BBBD to increase tumor delivery may have practical application in the clinic, because an increased delivery of drug to tumor can be obtained without increasing the dose of systemic drug. PMID- 10719868 TI - AR-R15896AR reduces cerebral infarction volumes after focal ischemia in cats. AB - OBJECTIVE: The use of competitive and noncompetitive N-methyl-D-aspartate receptor antagonists to prevent neuronal death during ischemia has been comprehensively studied. This study was performed to examine the neuroprotective effects and pharmacokinetics of the noncompetitive N-methyl-D-aspartate receptor channel blocker (S)-alpha-phenylpyridine-ethanamine dihydrochloride, AR-R15896AR (formerly designated ARL 15896AR), using a gyrencephalic cat middle cerebral artery occlusion model. METHODS: In a separate experiment, three cats were used for pharmacokinetic analysis, thus establishing the optimal dose of AR-R15896AR. Focal cerebral ischemia was induced in 21 cats. After 30 minutes of a 90-min ischemic insult, the cats received an intravenous infusion (total volume, 3 ml), in a 15-minute period, of either AR-R15896AR or normal saline solution (control). Physiological data were obtained after 40 and 80 minutes of ischemia and at 2, 4, and 6 hours after ischemia. At 6 hours after ischemia, each cat was positioned for both T2- and diffusion-weighted scans (eight slices, 5-mm thick). At 8 hours after ischemia, the animals were perfusion-fixed for histopathological analysis. RESULTS: Pharmacokinetic studies indicated that AR-R15896AR remained in the blood at elevated levels for the 6 hours studied, with a calculated half-life of approximately 6 hours. AR-R15896AR rapidly entered the brain and exhibited a brain/plasma ratio of approximately 8:1. The infarction volumes for the AR R15896AR-treated group were 1138.5+/-363.1, 651.3+/-428.9, and 118.6+/-50.1 mm3, as calculated using diffusion- and T2-weighted MRI and histopathological data, respectively. The infarction volumes for the control group were 3866.3+/-921, 3536+/-995.7, and 359.9+/-80.2 mm3, as calculated using diffusion- and T2 weighted MRI and histopathological data, respectively. No significant changes were observed in the physiological parameters measured (mean arterial blood pressure, pH, arterial carbon dioxide pressure, arterial oxygen pressure, sodium, potassium, chloride, and glucose levels, hematocrit, and temperature) for either the control or AR-R15896AR-treated group. Postischemic calcium levels returned to normal in the AR-R15896AR-treated cats, whereas they decreased in the control cats. CONCLUSION: When administered after ischemia, AR-R15896AR was effective in significantly reducing infarction volumes, as measured using diffusion- or T2 weighted magnetic resonance imaging data or quantitative histopathological data. This study also demonstrated that infarction volumes were greater in the diffusion- and T2-weighted magnetic resonance imaging scans than in the qualitative histopathological analyses, with the diffusion-weighted scans exibiting the largest infarction volumes. PMID- 10719869 TI - Neurosurgical notes: World War II. AB - This concerns my activities as a neurosurgeon in the European Theater of Operations and the North African, Tunisian campaign, during World War II. Action during the Battle of the Bulge came later. Our mobile tent hospital, the 9th Evacuation Hospital, was similar to that depicted in the television show M*A*S*H. To lend flavor to these comments, I have referred to medical and surgical matters in other units as well as our own, mentioned global aspects of the war, and included vignettes of life off-duty. The story begins after induction into the Army Medical Corps as a volunteer in July 1942 and ends with honorable discharge in April 1946. PMID- 10719870 TI - Stereotactic radiosurgery for tentorial dural arteriovenous fistulae draining into the vein of Galen: report of two cases. AB - OBJECTIVE AND IMPORTANCE: Treatment of tentorial dural arteriovenous fistulae (DAVFs) primarily draining into the vein of Galen remains a therapeutic challenge. We present two cases of ruptured galenic DAVFs that were successfully treated with gamma knife radiosurgery. CLINICAL PRESENTATION: Patient 1, a 66 year-old woman, experienced a sudden onset of headache and loss of consciousness. Neuroimaging studies revealed intraventricular hemorrhage and a DAVF with aneurysmal dilation of the vein of Galen. The DAVF was supplied by tentorial branches of the right meningohypophyseal artery and bilateral supracerebellar arteries, which drained directly into the vein of Galen. Patient 2, a 64-year-old woman, experienced subarachnoid hemorrhage. Cerebral angiography revealed a galenic DAVF at the falcotentorial junction, which was supplied by bilateral supracerebellar arteries. This patient had an aneurysm at the origin of the left supracerebellar artery. INTERVENTION: Both patients were treated with gamma knife radiosurgery. In each case, the fistula was exclusively targeted and a dose of more than 20 Gy was delivered. Complete obliteration of the fistula was confirmed 27 and 29 months after radiosurgery for Patients 1 and 2, respectively, whereas the normal venous structures of the galenic system were preserved. CONCLUSION: Gamma knife radiosurgery is an effective treatment modality for DAVFs primarily draining into the vein of Galen. Irradiation doses of more than 20 Gy, strictly limited to the fistulae, seem to be sufficient for successful obliteration of these high-risk vascular lesions, with minimal invasiveness. PMID- 10719871 TI - Hypophyseal tuberculoma: direct radiosurgery is contraindicated for a lesion with a thickened pituitary stalk: case report. AB - OBJECTIVE AND IMPORTANCE: Hypophyseal tuberculomas are extremely rare lesions. The recognition of hypophyseal tuberculomas in the differential diagnosis of pituitary tumors is important, even with no evidence of systemic tuberculosis. CLINICAL PRESENTATION: A 27-year-old female patient presented with continuous, dull, generalized headaches and amenorrhea, with no history of visual diminution, galactorrhea, or endocrinological abnormalities and no evidence of systemic tuberculosis. The patient exhibited a normal water balance, without polyuria or polydipsia. A gynecological examination, including an endometrial biopsy for amenorrhea, did not reveal any abnormalities. Perimetric and endocrinological examination results were normal. Contrast magnetic resonance imaging revealed a dense enhancing intrasellar mass, with thickening of the pituitary stalk. INTERVENTION: Sublabial rhinoseptal transsphenoidal decompression of the lesion was performed. The histopathological features were consistent with a diagnosis of tuberculoma, and acid-fast bacilli were demonstrated in the surgically removed tissue with Ziehl-Neelsen staining. As soon as the histopathological features were known, the patient underwent a lumbar puncture for cerebrospinal fluid analysis, which indicated normal findings. An intradermal tuberculin test yielded negative results. The patient was treated with medical therapy for 18 months, and complete resolution of the lesion was observed in follow-up examinations. CONCLUSION: Hypophyseal tuberculomas are often mistaken for pituitary adenomas. The finding of a thickened pituitary stalk in contrast magnetic resonance imaging scans may be useful for the differentiation of these lesions from pituitary adenomas. Direct radiosurgery is not an appropriate primary treatment method for pituitary adenomas and is principally restricted to elderly, medically unfit patients with microadenomas and patients with residual or recurrent tumors after microsurgery. It is contraindicated for patients who exhibit a thickened pituitary stalk in contrast magnetic resonance imaging scans. PMID- 10719872 TI - Intractable epilepsy associated with multiple cavernous malformations: case report. AB - OBJECTIVE AND IMPORTANCE: Epilepsy associated with cavernous malformations is often effectively controlled by lesionectomy alone. Detailed preoperative evaluation is necessary if the lesions are multiple and the seizures are medically intractable. We report on a patient with multiple cavernous malformations associated with medically intractable seizures who became seizure free after a single gyrus resection. The importance of electroencephalography with video monitoring is emphasized. CLINICAL PRESENTATION: The patient was a 25 year-old man with a 10-year history of complex partial and generalized convulsions. Magnetic resonance imaging revealed more than 10 cavernous malformations. Video-electroencephalographic monitoring indicated that seizures originated from either the frontal or temporal lobe of the right hemisphere. INTERVENTION: Subdural electrodes were implanted, covering both frontal and both temporal lobes. The seizures originated in the right frontal lobe. The gyrus, including a calcified cavernous malformation, was removed, and multiple subpial transections of the surrounding cortices were performed. The patient has been free of seizures for 22 months after surgery. CONCLUSION: Medically intractable seizures associated with multiple cavernous angiomas can be controlled by a single resective procedure. PMID- 10719873 TI - An intrasylvian "fibroma" in a child with cystic fibrosis: case report. AB - OBJECTIVE AND IMPORTANCE: Intracranial fibrous tumors are uncommon during childhood. An unusual case of benign intrasylvian "fibroma" that has remained clinically and radiographically stable more than 3 years after a subtotal resection is described. CLINICAL PRESENTATION: A 9-year-old girl with cystic fibrosis presented with new-onset focal seizures referable to a large calcified left sylvian fissure mass. INTERVENTION: An open biopsy with subtotal resection of the lesion revealed a benign process characterized by exuberant fibrocollagenous tissue intermeshed with chronic inflammatory cells and foreign body giant cells, encompassing islands of gliotic brain tissue. Immunohistochemical analysis showed staining for epithelial membrane antigen and reticulin within some of the spindle cells, although the majority were nonreactive. The majority of tumor cells exhibited staining for laminin; CD34 staining was absent. Ultrastructural studies were also suggestive of a fibroblastic rather than a meningothelial origin of the lesion, with elongated cells separated by abundant extracellular collagen. Although dense adherence of the mass to the pial surface and the middle cerebral artery vessels precluded a complete resection, the patient remains seizure-free without anticonvulsant therapy more than 3 years postoperatively with no evidence of growth of the lesion. CONCLUSION: The lesion in this patient bears morphological similarity to a rare group of tumors referred to as "intracerebral fibromas," although a variety of other rare mesenchymal neoplasms were also considered within the differential diagnosis. However, the absence of any definite neoplastic features, the finding of chronic inflammatory changes, and the lack of growth of the residual tumor during an extended follow-up interval indicate that the mass may represent either an extremely indolent neoplasm or a nonneoplastic process. The differential diagnosis of intracranial fibrous tumors is contrasted with that of the reported case. PMID- 10719874 TI - Cerebral angiofibroma: case report. AB - OBJECTIVE AND IMPORTANCE: Intracranial fibromatous tumors are very rare lesions, with few reported cases. CLINICAL PRESENTATION: We report the case of a 34-year old male patient who experienced seizures resulting from a cystic lesion in the left occipital region, which remained unchanged for 11 years. After the seizures increased in number, magnetic resonance imaging revealed a large cyst with a tumor nodule. INTERVENTION: A left occipital craniotomy was performed, and the tumor was removed. Pathological studies, including immunohistochemical and ultrastructural analyses, indicated that this neoplasm was composed of fibrous and angiomatous components, and a diagnosis of cerebral angiofibroma was established. CONCLUSION: Cerebral and meningeal fibromas are rare neoplasms that differ from solitary fibrous tumors and fibrous meningiomas. When a number of prominent blood vessels are present in a cerebral or meningeal fibroma, a diagnosis of angiofibroma can be considered. It is possible that some nodular brain tumors that were previously described as meningioangiomatosis could be reclassified as cerebral or meningeal angiofibromas. PMID- 10719875 TI - Familial colloid cysts of the third ventricle: case report. AB - OBJECTIVE AND IMPORTANCE: Familial colloid cysts of the third ventricle are very rare. This is the largest family reported and the first in which all affected members are female and all members have been screened. Screening led to the diagnosis of an asymptomatic case of a colloid cyst of the third ventricle, and the management of that lesion is discussed. CLINICAL PRESENTATION: A mother and two daughters who were diagnosed with colloid cysts of the third ventricle, from a family containing four sisters, three brothers, and the father, are presented. INTERVENTION: The index patient (Patient 2) underwent computed tomographic scanning-guided stereotactic transcallosal excision of her colloid cyst. Her siblings and her father were screened using magnetic resonance imaging as well as computed tomographic scanning. Cytogenetic analysis of blood samples obtained from the patient and her family revealed no chromosomal abnormalities. CONCLUSION: Screening is of value for families in which two or more members are affected. The management of asymptomatic cases is influenced by the lesion size and the age and fitness of the patient. PMID- 10719876 TI - Cranial base surgical techniques for large sphenocavernous meningiomas: technical note. AB - OBJECTIVE: Large meningiomas arising from the dura covering the sphenoid ridge present surgical challenges because of frequent involvement of the carotid artery and its branches, the optic nerve and tract, the superior orbital fissure, and cavernous sinus structures. To circumvent the inherent difficulties of a traditional approach strategy, cranial base approaches were applied to: 1) isolate and interrupt the major blood supply as an initial step, 2) minimize brain retraction, and 3) isolate the neurovascular structures exiting the tumor at the cranial base to protect and better separate them. METHODS: Six patients were treated with such a strategy in the past 2 years (five women and one man, ages 34-69 yr). All tumors measured at least 5 cm in their greatest diameter and arose at the sphenoid ridge. All tumors extended posteriorly to involve the cavernous sinus to varying degrees. In two patients a frontotemporal bone flap was used; in two patients, a transzygomatic approach was used; and in the remaining two patients, an orbitozygomatic strategy was used. Extensive bone removal at the cranial base was performed in all cases. RESULTS: Four patients had gross total resections, and two were subtotal owing to invasion of the cavernous sinus or the middle cerebral artery. There were no permanent cranial nerve deficits; however, two patients sustained transient IIIrd nerve paresis. Two patients postoperatively developed transient cerebral edema that required intensive treatment. All six patients had good outcomes, resuming independent activity by 3 months after surgery. CONCLUSION: Contemporary cranial base surgical techniques have a role in the treatment of large sphenoid ridge meningiomas. These strategies result in safe resection with low morbidity and obviate the need in most cases for preoperative embolization. The anatomic foundation for using these approaches is discussed. PMID- 10719877 TI - Capturing the spirit of a bygone age: the Treasury of Siphnos. PMID- 10719878 TI - Center stage for NGF in peripheral (but not central) sensory neuron outgrowth. PMID- 10719879 TI - Genetic evidence for a Nova regulator of alternative splicing in the brain. PMID- 10719880 TI - Adult neurogenesis in songbirds: a tale of two neurons. PMID- 10719881 TI - What's the matter? White matter? PMID- 10719882 TI - RNA helicase participates in the editing game. PMID- 10719883 TI - Of mice and MEN: what transgenic models tell us about hypothalamic control of energy balance. PMID- 10719884 TI - Synapse elimination and indelible memory. PMID- 10719885 TI - Maturation and maintenance of the neuromuscular synapse: genetic evidence for roles of the dystrophin--glycoprotein complex. AB - The dystrophin-glycoprotein complex (DGC) links the cytoskeleton of muscle fibers to their extracellular matrix. Using knockout mice, we show that a cytoplasmic DGC component, alpha-dystrobrevin (alpha-DB), is dispensable for formation of the neuromuscular junction (NMJ) but required for maturation of its postsynaptic apparatus. We also analyzed double and triple mutants lacking other cytoskeletal DGC components (utrophin and dystrophin) and myotubes lacking a alpha-DB or a transmembrane DGC component (dystroglycan). Our results suggest that alpha-DB acts via its linkage to the DGC to enhance the stability of postsynaptic specializations following their DGC-independent formation; dystroglycan may play additional roles in assembling synaptic basal lamina. Together, these results demonstrate involvement of distinct protein complexes in the formation and maintenance of the synapse and implicate the DGC in the latter process. PMID- 10719886 TI - Roles for ephrins in positionally selective synaptogenesis between motor neurons and muscle fibers. AB - Motor axons form topographic maps on muscles: rostral motor pools innervate rostral muscles, and rostral portions of motor pools innervate rostral fibers within their targets. Here, we implicate A subfamily ephrins in this topographic mapping. First, developing muscles express all five of the ephrin-A genes. Second, rostrally and caudally derived motor axons differ in sensitivity to outgrowth inhibition by ephrin-A5. Third, the topographic map of motor axons on the gluteus muscle is degraded in transgenic mice that overexpress ephrin-A5 in muscles. Fourth, topographic mapping is impaired in muscles of mutant mice lacking ephrin-A2 plus ephrin-A5. Thus, ephrins mediate or modulate positionally selective synapse formation. In addition, the rostrocaudal position of at least one motor pool is altered in ephrin-A5 mutant mice, indicating that ephrins affect nerve-muscle matching by intraspinal as well as intramuscular mechanisms. PMID- 10719887 TI - Essential roles of Drosophila RhoA in the regulation of neuroblast proliferation and dendritic but not axonal morphogenesis. AB - The pleiotropic functions of small GTPase Rho present a challenge to its genetic analysis in multicellular organisms. We report here the use of the MARCM (mosaic analysis with a repressible cell marker) system to analyze the function of RhoA in the developing Drosophila brain. Clones of cells homozygous for null RhoA mutations were specifically labeled in the mushroom body (MB) neurons of mosaic brains. We found that RhoA is required for neuroblast (Nb) proliferation but not for neuronal survival. Surprisingly, RhoA is not required for MB neurons to establish normal axon projections. However, neurons lacking RhoA overextend their dendrites, and expression of activated RhoA causes a reduction of dendritic complexity. Thus, RhoA is an important regulator of dendritic morphogenesis, while distinct mechanisms are used for axonal morphogenesis. PMID- 10719888 TI - Sonic hedgehog--regulated oligodendrocyte lineage genes encoding bHLH proteins in the mammalian central nervous system. AB - During development, basic helix-loop-helix (bHLH) proteins regulate formation of neurons from multipotent progenitor cells. However, bHLH factors linked to gliogenesis have not been described. We have isolated a pair of oligodendrocyte lineage genes (Olg-1 and Olg-2) that encode bHLH proteins and are tightly associated with development of oligodendrocytes in the vertebrate central nervous system (CNS). Ectopic expression of Olg-1 in rat cortical progenitor cell cultures promotes formation of oligodendrocyte precursors. In developing mouse embryos, Olg gene expression overlaps but precedes the earliest known markers of the oligodendrocyte lineage. Olg genes are expressed at the telencephalon diencephalon border and adjacent to the floor plate, a source of the secreted signaling molecule Sonic hedgehog (Shh). Gain- and loss-of-function analyses in transgenic mice demonstrate that Shh is both necessary and sufficient for Olg gene expression in vivo. PMID- 10719889 TI - Identification of a novel family of oligodendrocyte lineage-specific basic helix loop-helix transcription factors. AB - Basic helix-loop-helix (bHLH) transcription factors have been identified for neurons and their precursors but not for glial cells. We have identified two bHLH factors, Oligo1 and Oligo2, that are specifically expressed in zones of neuroepithelium from which oligodendrocyte precursors emerge, as well as in the precursors themselves. Expression of Oligo2 in the spinal cord precedes that of platelet-derived growth factor receptor alpha (PDGFRalpha), the earliest known marker of oligodendrocyte precursors, by several days. Ectopic expression of Oligo2 in vivo causes ectopic expression of Sox10, an HMG-box transcription factor expressed in oligodendrocyte and other glial precursors. These data identify Oligo genes as the earliest known markers of oligodendrocyte lineage determination and suggest they play a causal role in this process. PMID- 10719890 TI - Development of sensory neurons in the absence of NGF/TrkA signaling in vivo. AB - The neurotrophin survival dependence of peripheral neurons in vitro is regulated by the proapoptotic BCL-2 homolog BAX. To study peripheral neuron development in the absence of neurotrophin signaling, we have generated mice that are double null for BAX and nerve growth factor (NGF), and BAX and the NGF receptor TrkA. All dorsal root ganglion (DRG) neurons that normally die in the absence of NGF/TrkA signaling survive if BAX is also eliminated. These neurons extend axons through the dorsal roots and collateral branches into the dorsal horn. In contrast, superficial cutaneous innervation is absent. Furthermore, rescued sensory neurons fail to express biochemical markers characteristic of the nociceptive phenotype. These findings establish that NGF/TrkA signaling regulates peripheral target field innervation and is required for the full phenotypic differentiation of sensory neurons. PMID- 10719891 TI - Nova-1 regulates neuron-specific alternative splicing and is essential for neuronal viability. AB - We have combined genetic and biochemical approaches to analyze the function of the RNA-binding protein Nova-1, the paraneoplastic opsoclonus-myoclonus ataxia (POMA) antigen. Nova-1 null mice die postnatally from a motor deficit associated with apoptotic death of spinal and brainstem neurons. Nova-1 null mice show specific splicing defects in two inhibitory receptor pre-mRNAs, glycine alpha2 exon 3A (GlyRalpha2 E3A) and GABA(A) exon gamma2L. Nova protein in brain extracts specifically bound to a previously identified GlyRalpha2 intronic (UCAUY)3 Nova target sequence, and Nova-1 acted directly on this element to increase E3A splicing in cotransfection assays. We conclude that Nova-1 binds RNA in a sequence-specific manner to regulate neuronal pre-mRNA alternative splicing; the defect in splicing in Nova-1 null mice provides a model for understanding the motor dysfunction in POMA. PMID- 10719892 TI - Neuronal and glial glycine transporters have different stoichiometries. AB - A neurotransmitter transporter can potentially mediate uptake or release of substrate, and its stoichiometry is a key factor that controls the driving force and thus the neurotransmitter flux direction. We have used a combination of electrophysiology and radio-tracing techniques to evaluate the stoichiometries of two glycine transporters involved in glycinergic or glutamatergic transmission. We show that GlyT2a, a transporter present in glycinergic boutons, has a stoichiometry of 3 Na+/Cl-/glycine, which predicts effective glycine accumulation in all physiological conditions. GlyT1b, a glial transporter, has a stoichiometry of 2 Na+/Cl-/ glycine, which predicts that glycine can be exported or imported, depending on physiological conditions. GlyT1b may thus modulate glutamatergic synapses by increasing or decreasing the glycine concentration around N-methyl-D aspartate receptors (NMDARs). PMID- 10719893 TI - A novel targeting signal for proximal clustering of the Kv2.1 K+ channel in hippocampal neurons. AB - The discrete localization of ion channels is a critical determinant of neuronal excitability. We show here that the dendritic K+ channels Kv2.1 and Kv2.2 were differentially targeted in cultured hippocampal neurons. Kv2.1 was found in high density clusters on the soma and proximal dendrites, while Kv2.2 was uniformly distributed throughout the soma and dendrites. Chimeras revealed a proximal restriction and clustering domain on the cytoplasmic tail of Kv2.1. Truncations and internal deletions revealed a 26-amino acid targeting signal within which four residues were critical for localization. This signal is not related to other known sequences for neuronal and epithelial membrane protein targeting and represents a novel cytoplasmic signal responsible for proximal restriction and clustering. PMID- 10719894 TI - TASK-1, a two-pore domain K+ channel, is modulated by multiple neurotransmitters in motoneurons. AB - Inhibition of "leak" potassium (K+) channels is a widespread CNS mechanism by which transmitters induce slow excitation. We show that TASK-1, a two pore domain K+ channel, provides a prominent leak K+ current and target for neurotransmitter modulation in hypoglossal motoneurons (HMs). TASK-1 mRNA is present at high levels in motoneurons, including HMs, which express a K+ current with pH- and voltage-dependent properties virtually identical to those of the cloned channel. This pH-sensitive K+ channel was fully inhibited by serotonin, norepinephrine, substance P, thyrotropin-releasing hormone, and 3,5-dihydroxyphenylglycine, a group I metabotropic glutamate receptor agonist. The neurotransmitter effect was entirely reconstituted in HEK 293 cells coexpressing TASK-1 and the TRH-R1 receptor. Given its expression patterns and the widespread prevalence of this neuromodulatory mechanism, TASK-1 also likely supports this action in other CNS neurons. PMID- 10719895 TI - A localized interaction surface for voltage-sensing domains on the pore domain of a K+ channel. AB - Voltage-gated K+ channels contain a central pore domain and four surrounding voltage-sensing domains. How and where changes in the structure of the voltage sensing domains couple to the pore domain so as to gate ion conduction is not understood. The crystal structure of KcsA, a bacterial K+ channel homologous to the pore domain of voltage-gated K+ channels, provides a starting point for addressing this question. Guided by this structure, we used tryptophan-scanning mutagenesis on the transmembrane shell of the pore domain in the Shaker voltage gated K+ channel to localize potential protein-protein and protein-lipid interfaces. Some mutants cause only minor changes in gating and when mapped onto the KcsA structure cluster away from the interface between pore domain subunits. In contrast, mutants producing large changes in gating tend to cluster near this interface. These results imply that voltage-sensing domains interact with localized regions near the interface between adjacent pore domain subunits. PMID- 10719896 TI - Functional recovery of paraplegic rats and motor axon regeneration in their spinal cords by olfactory ensheathing glia. AB - Axonal regeneration in the lesioned mammalian central nervous system is abortive, and this causes permanent disabilities in individuals with spinal cord injuries. In adult rats, olfactory ensheathing glia (OEG) transplants successfully led to functional and structural recovery after complete spinal cord transection. From 3 to 7 months post surgery, all OEG-transplanted animals recovered locomotor functions and sensorimotor reflexes. They presented voluntary hindlimb movements, they supported their body weight, and their hindlimbs responded to light skin contact and proprioceptive stimuli. In addition, relevant motor axons (corticospinal, raphespinal, and coeruleospinal) regenerated for long distances within caudal cord stumps. Therefore, OEG transplantation provides a useful repair strategy in adult mammals with traumatic spinal cord injuries. Our results with these cells could lead to new therapies for the treatment of spinal cord lesions in humans. PMID- 10719897 TI - Analysis of clock proteins in mouse SCN demonstrates phylogenetic divergence of the circadian clockwork and resetting mechanisms. AB - The circadian clock in the suprachiasmatic nuclei (SCN) is comprised of a cell autonomous, autoregulatory transcriptional/translational feedback loop. Its molecular components include three period and two cryptochrome genes. We describe circadian patterns of expression of mPER2 and mPER3 in the mouse SCN that are synchronous to those for mPER1, mCRY1, and mCRY2. Coimmunoprecipitation experiments demonstrate in vivo associations of the SCN mPER proteins with each other and with the mCRY proteins, and of mCRY proteins with mTIM, but no mPER/mTIM interactions. Examination of the effects of weak and strong resetting light pulses on SCN clock proteins highlights a central role for mPER1 in photic entrainment, with no acute light effects on either the mCRY or mTIM proteins. These clock protein interactions and photic responses in mice are divergent from those described in Drosophila. PMID- 10719898 TI - Interactions between distinct GABA(A) circuits in hippocampus. AB - Synchronous activity among synaptically connected interneurons is thought to organize temporal patterns such as gamma and theta rhythms in cortical circuits. Interactions between distinct interneuron circuits may underlie more complex patterns, such as nested rhythms. Here, we demonstrate such an interaction between two groups of CA1 interneurons, GABA(A,slow) and GABA(A,fast) cells, that may contribute to theta and gamma rhythms, respectively. Stratum lacunosum moleculare (SL-M) stimuli that activate GABA(A,slow) inhibitory postsynaptic currents (IPSCs) in pyramidal cells simultaneously depress the rate and amplitude of spontaneous GABA(A,fast) IPSCs for several hundred milliseconds. This suppression has a similar pharmacological profile to GABA(A,slow) IPSCs, and SL-M stimuli elicit GABA(A,slow) IPSCs in interneurons. We conclude that GABA(A,slow) cells inhibit both pyramidal cells and GABA(A,fast) interneurons and postulate that this interaction contributes to nested theta/gamma rhythms in hippocampus. PMID- 10719899 TI - Diversity and cell type specificity of local excitatory connections to neurons in layer 3B of monkey primary visual cortex. AB - In the primary visual cortex of macaque monkeys, laminar and columnar axonal specificity are correlated with functional differences between locations. We describe evidence that embedded within this anatomical framework is finer specificity of functional connections. Photostimulation-based mapping of functional input to 31 layer 3B neurons revealed that input sources to individual cells were highly diverse. Although some input differences were correlated with neuronal anatomy, no 2 neurons received excitatory input from the same cortical layers. Thus, input diversity reveals far more cell types than does anatomical diversity. This implies relatively little functional redundancy; despite trends related to laminar or columnar position, pools of neurons contributing uniquely to visual processing are likely relatively small. These results also imply that similarities in the anatomy of circuits in different cortical areas or species may not indicate similar functional connectivity. PMID- 10719900 TI - Formation of temporal memory requires NMDA receptors within CA1 pyramidal neurons. AB - In humans the hippocampus is required for episodic memory, which extends into the spatial and temporal domains. Work on the rodent hippocampus has shown that NMDA receptor (NMDAR) -mediated plasticity is essential for spatial memory. Here, we have examined whether hippocampal NMDARs are also needed for temporal memory. We applied trace fear conditioning to knockout mice lacking NMDARs only in hippocampal CA1 pyramidal cells. This paradigm requires temporal processing because the conditional and unconditional stimuli are separated by 30 s (trace). We found that knockout mice failed to memorize this association but were indistinguishable from normal animals when the trace was removed. Thus, NMDARs in CA1 are crucial for the formation of memories that associate events across time. PMID- 10719901 TI - Targeted neuronal death affects neuronal replacement and vocal behavior in adult songbirds. AB - In the high vocal center (HVC) of adult songbirds, increases in spontaneous neuronal replacement correlate with song changes and with cell death. We experimentally induced death of specific HVC neuron types in adult male zebra finches using targeted photolysis. Induced death of a projection neuron type that normally turns over resulted in compensatory replacement of the same type. Induced death of the normally nonreplaced type did not stimulate their replacement. In juveniles, death of the latter type increased recruitment of the replaceable kind. We infer that neuronal death regulates the recruitment of replaceable neurons. Song deteriorated in some birds only after elimination of replaceable neurons. Behavioral deficits were transient and followed by variable degrees of recovery. This raises the possibility that induced neuronal replacement can restore a learned behavior. PMID- 10719902 TI - Microstructure of temporo-parietal white matter as a basis for reading ability: evidence from diffusion tensor magnetic resonance imaging. AB - Diffusion tensor magnetic resonance imaging (MRI) was used to study the microstructural integrity of white matter in adults with poor or normal reading ability. Subjects with reading difficulty exhibited decreased diffusion anisotropy bilaterally in temporoparietal white matter. Axons in these regions were predominantly anterior-posterior in direction. No differences in T1-weighted MRI signal were found between poor readers and control subjects, demonstrating specificity of the group difference to the microstructural characteristics measured by diffusion tensor imaging (DTI). White matter diffusion anisotropy in the temporo-parietal region of the left hemisphere was significantly correlated with reading scores within the reading-impaired adults and within the control group. The anisotropy reflects microstructure of white matter tracts, which may contribute to reading ability by determining the strength of communication between cortical areas involved in visual, auditory, and language processing. PMID- 10719903 TI - Rapid determination of furanocoumarins in creams and pomades using SPE and GC. AB - A solid-phase extraction and chromatography-flame ionization detection (GC-FID) method has been developed for the routine analysis of psoralen, bergapten, isopimpinellin and pimpinellin in creams and pomades employed in Brazil for the treatment of vitiligo. The calibration curve for psoralen was linear in the range 10-100 microg ml(-1), for bergapten 5-90 microg ml(-1), for pimpinellin 10-90 microg ml(-1) and for isopimpinellin 5-100 microg ml(-1). The best recoveries of the furanocoumarins in the creams analysed were 94-97%, whereas in the pomades, recoveries were 94-96%. The R.S.D. of the quantitative-analysis of the furanocoumarins in the products analyses were within 5%. PMID- 10719904 TI - First derivative spectrophotometric and LC determination of benoxinate hydrochloride and its degradation products. AB - Two methods are presented for the determination of benoxinate HCI and its acid and alkali-induced degradation products using first derivative (1D) spectrophotometry with zero-crossing measurements and liquid chromatography. Benoxinate HCl was determined by measurement of its first derivative amplitude in mcllvaine's-citric acid phosphate buffer pH 7.0 at 268.4 and 272.4 nm in the presence of its alkali- and acid-induced degradation products, respectively. The acid- and alkali-induced, degradation products were determined by measurement of their first derivative amplitude in the same solvent at 307.5 nm. The LC method depends upon using a mu bondapak CN column with a mobile phase consisting of acetonitrile-water triethylamine (60:40:0.01, v/v) and adjusted to apparent pH 7. Quantitation was achieved with UV detection at 310 nm based on peak area. The proposed methods were utilized to investigate the kinetics of the acidic and alkaline degradation processes at different temperatures. The pH-rate profile of degradation of benoxinate HCl in Britton-Robinson buffer solutions was studied. PMID- 10719905 TI - Indirect atomic absorption spectrometric determination of pindolol, propranolol and levamisole hydrochlorides based on formation of ion-associates with ammonium reineckate and sodium cobaltinitrite. AB - A new simple, accurate, precise and sensitive indirect method for the determination of pindolol HCl (1), propranolol HCl (2) and levamisole HCl (3) using atomic absorption spectrometry has been developed. The method is based on precipitation of the ion-associates formed from the reaction of (1), (2) or (3) with ammonium reineckate and/or sodium cobaltinitrite. The solubility of the solid complexes at the optimum conditions of pH and ionic strength values have been studied. Saturated solutions of each ion-associate were prepared under the optimum conditions and the metal ion content in the supernatant was determined. The method has been used for the determination of 1.14-17.07, 1.18-17.75 and 1.08 16.24 microg/ml of (1), (2) and (3), respectively, using ammonium reineckate, and 1.71-25.60, 1.77-26.62 and 1.62-24.36 microg/ml of (1), (2) and (3), respectively, using sodium cobaltinitrite. The method developed was applied for analysis of bulk drugs and some of their pharmaceutical preparations. PMID- 10719906 TI - Liquid chromatographic assay of nifedipine in human plasma and its application to pharmacokinetic studies. AB - A highly sensitive, selective and reproducible reversed-phase high-performance liquid chromatographic method has been developed for the determination of nifedipine in human plasma with minimum sample preparation. The method is sensitive to 3 ng/ml in plasma, with acceptable within- and between-day reproducibilities and linearity (r2 > 0.99) over a concentration range from 10 200 ng/ml. Acidified plasma samples were extracted using diethyether containing diazepam as internal standard and chromatographic separation was accomplished on C18 column using a mobile phase consisting of acetonitrile, methanol and water (35:17:48, v/v). The within-day precision ranged from 2.22 to 4.64% and accuracy ranged from 102.4-106.4%. The day-to-day precision ranged from 2.34-7.07% and accuracy from 95.1-100.1%. The relative recoveries of nifedipine from plasma ranged from 91.0-107.3% whereas extraction recoveries were 88.6-93.3%. Following eight 6-week freeze-thaw cycles, nifedipine in plasma samples proved to be stable with accuracy ranging from 0.64 to 3.0% and precision ranging from 3.6 to 4.15%. Nifedipine was also found to be photostable for at least 120 min in plasma, 30 min in blood and for 60 min in aqueous solutions after exposure to light. The method is sensitive and reliable for pharmacokinetic studies and therapeutic drug monitoring of nifedipine in humans after the oral administration of immediate release capsules and sustained-release tablets to five healthy subjects. PMID- 10719907 TI - Validated HPLC method for determination of PAT-5A, an insulin sensitizing agent, in rat plasma. AB - A high performance liquid chromatographic method for the determination of PAT-5A (a potent insulin sensitizer) using DRF-2095 (a thiazolidinedione) as internal standard (I.S.) is described. A 1:1 v/v ethylacetate and dichloromethane solvent mixture was used for extraction of PAT-5A from plasma. A Kromasil KR100-5C18 250A, 5 microm, 4.6 x 250 mm SS column was used for the analysis. Mobile phase consisting of sodium dihydrogen phosphate (pH 4.0, 0.05 M) and methanol mixture (25:75, v/v) was used at a flow rate of 1.0 ml/min. The eluate was monitored using a UV detector set at 345 nm. Ratio of peak area of analyte to I.S. was used for quantification of plasma samples. Using this method the absolute recovery of PAT-5A from rat plasma was > 90% and the limit of quantification was 0.05 microg/ml. The intra-day relative standard deviation (RSD) ranged from 2.19 to 4.98% at 1.0 microg/ml, 1.05 to 3.68% at 10.0 microg/ml and 3.14 to 5.08% at 50 microg/ml. The inter-day RSD were 1.6, 2.24 and 1.54% at 1, 10 and 50 microg/ml, respectively. The method was applied to measure the plasma concentrations of PAT 5A in pharmacokinetic and bioavailability studies in male Wistar rats. PMID- 10719908 TI - Sensitive spectrofluorimetric and spectrophotometric methods for the determination of thonzylamine hydrochloride in pharmaceutical preparations based on coupling with dimethylbarbituric acid in presence of dicyclohexylcarbodiimide. AB - Two sensitive and selective spectrophotometric and spectrofluoimetric procedures were developed for the determination of thonzylamine hydrochloride (THAH) in tablets and nasal drops. The methods are based on Konig reaction which resulted in an orange-yellow fluorescent product. The orange-yellow product of the interaction between the dicyclohexylcarbodiimide (DCC), THAH and dimethylbarbituric acid (DMBA) showed an absorption maximum at 492 nm, a first derivative signal at 494 nm and a fluorescence emission peak at 518 nm (lambda(ex)=492 nm). The orange-yellow color was found to be stable for at least 2 h. The reaction conditions were studied and optimized. The reaction obeys Beer's law over the ranges 8-20 and 0.2-2.0 microg ml(-1) for the derivative spectrophotometric and fluorimetric measurements, respectively. The detection limits were found to be 0.29 and 0.018 microg ml(-1) for the spectrophotometric and fluorimetric measurements, respectively. The proposed methods were applied to the analysis of pharmaceutical formulations containing THAH, either alone or in combination with naphazoline nitrate. PMID- 10719909 TI - A quantitative minimally invasive assay for the detection of metals in the stratum corneum. AB - A quantitative, minimally invasive tape-stripping assay for the detection of metals on and in skin that also has application to the detection of metallic elements on dry surfaces (where human contact could occur) has been developed. This development included construction, using commercial products, of an approximately 25 microm thick, low-metal content tape suitable both for tape stripping and elemental analysis. Individual tapes were sequentially applied to the skin surface and then removed, taking with them a sample of the dead outer layer of the skin (stratum corneum). Analysis of such tape strip samples by particle induced X-ray emission (PIXE)--a well-characterized, sensitive, analytical technique based on X-ray spectrometry--identified and accurately quantified the metals in the sample. The assay had elemental sensitivities of approximately 1 ng/cm2 for many metals and analysis of elemental contents could be performed in as little as 5 min. The feasibility of the assay for measuring metals in the stratum corneum was demonstrated on the forearms of healthy human volunteers. Samples from approximately half the subjects were found to contain zirconium, possibly arising from the use of roll-on antiperspirants. The assay has potential as a tool: (1) for risk assessment, (2) to identify exposure levels following possible contact with a hazardous metal, and (3) to determine the effectiveness of cleanup or removal measures. PMID- 10719910 TI - Sensitive assay for the determination of cefazolin or ceftriaxone in plasma utilizing LC. AB - A rapid, specific and very sensitive liquid chromatographic assay using standard ultraviolet detection has been developed to measure cefazolin (CFZ) or ceftriaxone (CFX) in small samples (200 microl) of plasma using either drug as the internal standard for measurement of the other. A rapid extraction was performed using C18 bonded Sep Pak cartridges with high extraction efficiency for both drugs. The chromatographic system employed the use of a Nova-Pak C18 4 microm cartridge with a radial compression system preceded by a Guard-Pak with a C18 insert. The mobile phase consisted of an aqueous solution containing 10 mM of dibasic potassium phosphate and 10 mM cetyltrimethylammonium bromide (pH 6.5) with acetonitrile (73:27 v/v). The drug and internal standard (CFZ/CFX) were detected using a UV detector set at a wavelength of 274 nm. Assay results were linearly related to the concentration (r > 0.997) for the wide range which was examined (0.005-120 microg/ml) for either drug. We report the precision, accuracy, recovery, linearity, sensitivity and specificity of this assay. The intra-run and inter-run CV was less than 9.02%. This method is currently being used for clinical therapeutic monitoring and pharmacokinetic studies of CFZ and CFX in patients undergoing cesarean section. PMID- 10719911 TI - Comparison of UV spectrophotometric and LC methods for the determination of nortriptyline hydrochloride in polysorbate 80 based oil/water (o/w) microemulsions. AB - A new rapid, reliable and specific UV spectrophotometric method was developed for the determination of nortriptyline hydrochloride formulated into o/w microemulsions. The UV spectra of nortriptyline standard solution in methanol and placebo (microemulsion without nortriptyline) were recorded over the wavelength range 200-600 nm and the spectra for placebo and nortriptyline loaded microemulsion were recorded over the range 260-400 nm in order to determine the overlapping that might appears, and hence to set the wavelength that could be used for the quantitative analysis. This method was validated and compared with a liquid chromatography (LC) procedure used for the quantitative analysis of the drug. Both methods showed excellent precision and accuracy with RSD values of 2.37 and 1.41%, respectively, for the LC method, and values of 1.24 and 2.88%, respectively, for the UV spectrophotometric method. The established linearity range was 10-50 microg ml(-1) (r2 = 0.9985) and 20-60 microg ml(-1) (r2 = 0.9979) for the HPLC and UV spectrophotometric methods respectively. The recoveries of nortriptyline from spiked placebos were > 95% for both methods over the linear range. The methods have been successfully used for determining the nortriptyline content of microemulsions and for evaluating the chemical stability of the drug in nortriptyline-loaded microemulsions. PMID- 10719912 TI - The determination of 3-nitrophenol and some other aromatic impurities in 4 nitrophenol by reversed phase HPLC with peak suppression diode array detection. AB - In this work the peak suppression technique is used for the determination of 3 nitrophenol and some other aromatic impurities in 4-nitrophenol by reversed phase HPLC with diode array detection. Taking into account the differences between the absorption spectra of the two compounds, two wavelengths were selected in order to obtain the maximum difference between the spectral contribution for 3 nitrophenol and to maintain a small, similar spectral contribution for 4 nitrophenol (the main compound). Then we used the wavelength corresponding to a small spectral contribution of 3-nitrophenol as the reference wavelength. It was shown that taking lambda(an) = 266 nm and lambda(ref) = 364 nm, a broad elution peak of 4-nitrophenol was suppressed deconvoluting the peak of 3-nitrophenol. Moreover, quantitation of 3-nitrophenol was achieved without chemometric tools. Under the proposed conditions the detection limits for 3-nitrophenol and other common impurities of 4-nitrophenol used in the pharmaceutical industry (4 chlorophenol, 4-nitrophenol, 1-chloro-2-nitrobenzene, 1-chloro-4-nitrobenzene, 4,4'-bisfenilether, and 4,4'-dichloroazobenzene) were not significantly affected as compared with respective detection limits evaluated in the absence of 4 nitrophenol and using standard detection conditions (lambda(an) = 280 nm and lambda(ref) = 420 nm). PMID- 10719913 TI - Analytical methodologies for atomic spectrometric determination of metallic oxides in UV sunscreen creams. AB - In this study, methodologies for determining titanium oxide, zinc oxide and iron oxide are proposed and assayed in commercial sunscreen products. The proposed methodology for TiO2, determination in sunscreens is based on a microwave assisted treatment for digesting the organic components in a closed teflon reactor in presence of HNO3 and HCl. Titanium is determined by inductive coupled plasma emission spectrometry (ICP-AES). The proposed methodologies for measuring ZnO and Fe2O3 are based on a sample emulsification in water with a non ionic tensioactive and IBMK, followed by Zn and Fe determination by flame atomic absorption spectrometry (FAAS). The methodologies allow a precise and accurate determination of metallic oxides in UV sunscreen creams, where the sample treatment is less time-consuming than in the classic methods. To our knowledge this is the first study focused to the determination of metallic oxides in commercial sunscreen products. PMID- 10719914 TI - LC determination of indinavir in biological matrices with electrochemical detection. AB - A high performance liquid chromatographic (HPLC) method with electrochemical detection for the quantification of Indinavir in cell culture is described. The sample pre-treatment involved a protein precipitation procedure using acetonitrile. Chromatography was carried out on a base-deactivated reversed-phase column with an isocratic mobile phase. The method was validated with regard to specificity, linearity, limits of detection and quantitation, precision and accuracy, recovery and ruggedness. The proposed HPLC assay was utilised to directly evaluate the capability of P-glycoprotein expressing multidrug resistant cells in mediating the transport and efflux of protease inhibitor (PI) Indinavir, a basic compound in AIDS care. PMID- 10719915 TI - Voltammetric investigation of oxidation of zuclopenthixol and application to its determination in dosage forms and in drug dissolution studies. AB - The oxidative voltammetric behaviour of zuclopenthixol (ZPT) at a glassy carbon has been studied using cyclic, linear sweep and differential pulse voltammetry. Oxidation of the drug produced three pH dependent anodic steps (representing an irreversible oxidation). Using differential pulse voltammetry, the drug yielded a well-defined voltammetric response in phosphate buffer, pH 5.2 at + 0.82 V (vs. Ag/AgCl). This process could be used to determine ZPT concentrations in the range 8 x 10(-7)-2 x 10(-4) M. The method was applied, without any interferences from the excipients, to the determination of the drug in tablets and oral drops, and in drug dissolution studies. PMID- 10719916 TI - Analysis of film coating thickness and surface area of pharmaceutical pellets using fluorescence microscopy and image analysis. AB - A method is presented which enables geometrical characterisation of pharmaceutical pellets and their film coating. It provides a high level of details on the single pellet level. Image analysis was used to determine the coating thickness (h) applied on the pellets and the surface area (A) of the pellet cores. Different definitions of A and h are evaluated. Hierarchical analysis of variance was used to resolve different sources contributing to the total variance. The variance within pellets and the variance between pellets were found as significant sources of variation. Special emphasis was put on evaluation of A/h due to its influence on the release rate of an active drug substance from the pellet core. The pellet images were thus used to predict variations in the release rate using a mathematical model as a link between the image data and the release rate. General aspects of image analysis are discussed. The method would be useful in calibration of near infrared spectra to h in process analytical chemistry. PMID- 10719917 TI - Validation of a LC method for the determination of 5-aminosalicylic acid and its metabolite in plasma and urine. AB - The choice of a proper analytical method for the quantification of drugs and/or their metabolites in biological samples plays a significant role in the evaluation and interpretation of bioavailability, bioequivalence and pharmacokinetic data. The aim of this study was validation of a method for the identification and quantitative determination of 5-aminosalicylic acid (5-ASA) and its metabolite N-acetyl-5-aminosalicylic acid in human plasma and urine. According to previous studies on the disposition of 5-ASA (mesalazine) in a patient with inflammatory bowel diseases, we have developed a rapid, sensitive method for the determination of 5-ASA and its acetylated metabolite, N-acetyl-5 aminosalicylic acid (Ac-5-ASA). The advantage of this method is that it measures both compounds, and is more rapid, reproducible and credible than the previous studies [C. Fischer, K. Maier, U. Klotz, J. Chromatogr. Biomed. Appl. 225 (1981) 498-503; P.N. Shaw, A.L. Sivner, L. Aarons, J.B. Houston, J. Chromatogr. Biomed. Appl. 274 (1983) 393-397; B. Norlander, R. Gotthard, M. Strom, Aliment. Pharmacol. Ther. 3 (1989) 333-342; U. Klotz, G.L. Stracciari, Arzneim.-Forsch. Drug Res. II 43 (12) (1993) 1357-1359]. 5-ASA was quantitatively determined in human plasma and urine samples by liquid chromatography following prior derivatization to its acetylated metabolite (Ac-5-ASA). N-Acetyl-anthranilic acid was used as the internal standard. The detection was performed with a spectrofluorimetric detector, excitation at 311 nm, cut-off at 449 nm. The method was validated for the following parameters: linearity, recovery, sensitivity, precision, accuracy, selectivity and stability, limits of quantification and of detection. It showed good linearity (r2 > or = 0,996) in the range 0.1 ng/ml to 8 microg/ml using a Lichrospher 60 RP-select B column. The lower limit of detection was 20 ng/ml in plasma and urine. The within-run relative standard deviations (R.S.D.) were below 6.7% at all concentration levels and the between-run R.S.D. were below 25.4% at all concentration levels. PMID- 10719918 TI - Cleaning validation procedure eased by using overpressured layer chromatography. AB - In the manufacturing plants of many pharmaceutical companies the reaction apparatus is suitable to produce different active pharmaceutical ingredients. After completing the production of a compound the equipment should be cleaned in order to avoid the cross contamination in the next lot of the other products. In the authors' laboratory several chromatographic methods were introduced to measure the amount of the residual substances remaining on the surface of the apparatus after the cleaning procedure. A sensitive and fairly rapid overpressured layer chromatographic (OPLC) procedure--suitable to separate and control five steroid hormone compounds (allylestrenol, estradiol, ethynodiol diacetate, levonorgestel, norethisterone) produced in the same equipment at different times--was developed and validated. PMID- 10719919 TI - Voltammetric determination of lactate dehydrogenase using a carbon paste electrode. AB - A voltammetric study was performed by linear sweep voltammetry using a carbon paste electrode, in -0.1 to +1.3 V potential range, with the view of elaborating an assay for lactate dehydrogenase (LDH) in different enzymatic preparations and biological fluids. There have been performed studies concerning pH influence upon the enzymatic reaction, as well as the electrochemical behavior of LDH in the presence of modified carbon paste electrodes saturated with sodium pyruvate and/or NADH. PMID- 10719920 TI - Analysis of some antifungal drugs by spectrophotometric and spectrofluorimetric methods in different pharmaceutical dosage forms. AB - Simple spectrophotometric and spectrofluorimetric methods are suggested for the determination of antifungal drugs; clotrimazole, econazole nitrate, ketoconazole, miconazole and tolnaftate. Spectrophotometric one depends on the interaction between imidazole antifungal drugs as n-electron donor with the pi acceptor 2,3 dichloro-5,6-dicyano-1,4-benzoquinone (DDQ) in methanol or with p-chloranilic acid (p-CA) in acetonitrile. The produced chromogens obey Beer's law at lambda(max) 460, and 520 nm in the concentration range 22.5-200 and 7.9-280 microg ml(-1) for DDQ, and p-CA, respectively. Spectrofluorimetric method is based on the measurement of the native fluorescence of ketoconazole at 375 nm with excitation at 288 nm and or the induced fluorescence after alkaline hydrolysis of tolnaftate with 5 M NaOH solution at 420 nm with excitation at 344 nm. Fluorescence intensity versus concentration is linear for ketoconazole at 49.7-800 ng ml(-1) while for tolnaftate, it is in the range of 20.4-400 ng ml( 1). The proposed methods were applied successfully for the determination of all the studied drugs in their pharmaceutical formulations. PMID- 10719921 TI - Determination of metoprolol enantiomers in human urine by coupled achiral-chiral chromatography. AB - Achiral chiral column switching HPLC assay was developed to allow the separation and quantitation of the enantiomers of metoprolol in human urine by means of fluorescence detection. Urine samples were prepared by liquid liquid extraction, followed by HPLC. The racemic metoprolol and internal standard were separated from the interfering components in urine and quantified on the silica column, and the enantiomers were determined on a Chiralcel OD chiral stationary phase. The two columns were connected by a switching valve equipped with a silica trap column. Detection limit was 25 ng/ml for each enantiomer. The intra-day variation ranged between 0.38 and 4.94% in relation to the measured concentration and the inter-day variation was 0.15-3.13%. It has been applied to the determination of (R)-(+)-metoprolol and (S)-(-)-metoprolol in urine from healthy volunteers dosed with racemic metoprolol tartrate. PMID- 10719922 TI - Quantitative determination of some thiazole cephalosporins through complexation with palladium (II) chloride. AB - A simple and sensitive spectrophotometric method has been developed for the determination of five cephalosporins namely cefpodoxime (CFPD), ceftizoxime (CTIZ), ceftazidime (CZD), ceftriaxone (CTRX), and cefixime (CXIM). This method is based on the formation of yellow to yellowish brown complex between palladium (II) chloride and the investigated cephalosporins in the presence of sodium lauryl sulphate (SLS) as surfactant. The reaction conditions were studied and optimized. The procedure was validated. For each drug, the composition of this complex as well as its stability constant were also investigated. The proposed method was used for the determination of the above-mentioned drugs in their commercial preparations. The results were compared statistically with either official or published methods and showed no significant difference between the two methods. PMID- 10719923 TI - LC analysis of benzophenone-3 in pigskin and in saline solution: application to determination of in vitro skin penetration. PMID- 10719924 TI - Anatomy and innervation of the rhabdosphincter of the male urethra. AB - The rhabdosphincter of the male urethra and its innervation are still a subject of controversy. Essentially, two concepts of its anatomy can be found in the literature. Some authors describe the rhabdosphincter as part of the urogenital diaphragm caudal to the prostate, others as a striated muscle that extends from the base of the bladder to the "urogenital diaphragm." In anatomic histological studies, the rhabdosphincter and its innervation were examined by means of anatomical dissections and serial anatomical as well as histological sections of 19 male pelves, including 8 fetal specimens. The rhabdosphincter presents as a vertical structure extending from the bulb of the penis to the region of the bladder neck along the prostate and the membranous urethra. Inserting dorsally in the perineal body via a broad tendinous raphe, the striated muscle fibers form an omega-shaped loop around the anterior and lateral aspects of the membranous urethra. The existence of a "urogenital diaphragm" and a strong, circular, striated "external sphincter urethrae" completely encircling the urethra caudal to the apex of the prostate cannot be confirmed by anatomical and histological investigations. The rhabdosphincter is supplied by branches of the pudendal nerve after leaving the pudendal canal. PMID- 10719925 TI - Anatomic basis for the continence-preserving radical retropubic prostatectomy. AB - The technique of continence-preserving anatomic radical retropubic prostatectomy focuses on the preservation of the following anatomic components of the external striated urethral sphincteric complex: (1) the entire circumference of the rhabdosphincter musculature, (2) the periurethral fascial investments (the pubourethral ligaments anterolaterally and median fibrous raphe posteriorly), and (3) the innervation of both the rhabdosphincter by way of the intrapelvic branch of the pudendal nerve (somatic) and the mucosal and smooth muscle components by way of the urethral branch of the inferior hypogastric plexus (autonomic). The clinical impact of preserving the external striated urethral sphincter and its innervation by performing a continence preserving anatomic retropubic prostatectomy is a shorter time to achieve urinary continence. PMID- 10719926 TI - Minilaparotomy radical retropubic prostatectomy: updated technique and results. AB - The purpose of this article is to reduce the incisional morbidity associated with standard radical retropubic prostatectomy using the minilaparotomy incision developed for pelvic lymph-node dissection, which was applied to radical retropubic prostatectomy. More than 522 patients underwent minilaparotomy radical retropubic prostatectomy from 1991 to 1997. Preoperative evaluation included history, physical examination, prostate-specific antigen (PSA), and Gleason's grade. Postoperative follow-up included serial PSA measurements and a determination of continence. The surgical technique is described in detail. Two hundred sixty-five patients responded to the mailed questionnaire out of a total 522 patients. Satisfactory continence, defined as 0 to 1 pad per day, was achieved in 85% of patients, and 83% of patients had a PSA < 0.2 at an average follow-up of 2.6 years. There was no operative mortality, and overall complication rate was similar to other surgeons. The typical patient was discharged home 3 days postoperatively. Minilaparotomy radical retropubic prostatectomy compares favorably with standard radical retropubic prostatectomy. PMID- 10719927 TI - Puboprostatic ligament sparing radical retropubic prostatectomy. AB - Prostate cancer is the most common solid malignancy and the second most common cause of cancer death in man. Radical prostatectomy is the therapeutic modality that currently provides the best long-term biochemical relapse-free survival rate. Yet many patients select alternative forms of therapy or no therapy at all because of fears that treatment will significantly alter quality of life. Urinary incontinence following radical prostatectomy has a significant deleterious effect on quality of life and, unfortunately, is much more prevalent following surgery compared with other treatment modalities, such as radiation therapy. Many efforts have been undertaken to avoid this complication with only modest success achieved. These include creation of a neobladder neck, bladder neck preservation, periurethral injection of bulking agents, and anterior urethropexy. A technique for radical retropubic prostatectomy that spares the puboprostatic ligaments, which preserves the normal anterior support of the urethra, is described herein. The outcome following this procedure demonstrates more rapid return of full urinary continence following radical prostatectomy in a controlled study. However, the "Holy Grail" of complete eradication of urinary incontinence following radical prostatectomy has not been achieved. PMID- 10719928 TI - Management of the dorsal vein complex during radical retropubic prostatectomy. AB - An understanding of the prostatic dorsal venous anatomy allows dissection of the prostatic apex in a manner that results in minimal bleeding while preserving the rhabdosphincter, urethra, and neurovascular bundles during radical retropubic prostatectomy. This article reviews the pertinent venous anatomy of the prostatic apex. A surgical technique is described that allows secure venous control and that has resulted in consistently low blood loss and an allogeneic transfusion rate of less than 1% of patients. PMID- 10719929 TI - Initial release of the lateral pelvic fascia. AB - Initial incision of the lateral pelvic fascia before division of the dorsal vein complex allows posterior displacement of the neurovascular bundles and development of the proper prostatorectal plane. This technique may decrease positive surgical margins while preserving the neurovascular bundles. This technique also completely preserves the posterior attachments to the urethra, allowing preservation of maximal urethral length. PMID- 10719930 TI - Technique for nerve dissection. AB - Nerve-sparing radical retropubic prostatectomy is a reasonable treatment option in localized prostate cancer with minimal morbidity. Recent techniques in neurovascular bundle preservation could lead to an overall improvement in postoperative quality of life without compromising cancer control in the appropriately selected patient. Different techniques for neurovascular bundle preservation have been described by most major centers. This brief article describes the updated technique of nerve-sparing radical retropubic prostatectomy for clinically localized prostate cancer. Our technique of nerve dissection starts at the lateral aspect of the prostate with secondary urethral dissection. We believe this technique is easy to learn and decreases dissection around the striated sphincter. PMID- 10719931 TI - Neurostimulation during radical prostatectomy: improving nerve-sparing techniques. AB - Considerable variation exists in the reported results of nerve-sparing prostatectomy with respect to potency preservation. This may reflect differences in surgical technique. The Cavermap is a device that uses intraoperative nerve stimulation with real time tumescence monitoring to permit identification of the course of the cavernous nerve fibers during radical prostatectomy. Results of a single center phase 2 and a blinded multicenter phase 3 study have demonstrated an improved outcome compared with conventional nerve sparing with respect to Rigiscan-measured nocturnal erection at 1 year after prostatectomy. The device permits evaluation of the success of nerve-sparing during surgery. Further studies of the effectiveness of the Cavermap device are warranted. PMID- 10719932 TI - Bladder-neck preservation during radical retropubic prostatectomy. AB - The goal of the urologic surgeon performing total prostatectomy for prostate cancer is to eliminate the cancer and minimize the side effects associated with treatment. We believe that careful dissection of the prostate from the bladder can be performed in such a manner as to preserve most of the circular fibers of the bladder neck. This so-called bladder-neck preservation technique appears to reduce the risk of an anastomotic stricture and accelerate the return of urinary continence. An analysis of 676 consecutive prostatectomies revealed that 4.3% of the men had tumor touching the inked bladder neck margin. Only 1% had this as the only positive margin. Most of these patients had a preoperative prostate-specific antigen > 10 and a Gleason score of 7 or greater suggesting that bladder-neck preservation did not compromise the outcome of surgery. A more extensive resection of the bladder neck is not likely to be curative. PMID- 10719933 TI - Early catheter removal following radical retropubic prostatectomy. AB - Indwelling urethral catheters are bothersome for patients following radical retropubic prostatectomy. At Indiana University, to alleviate postoperative discomfort, early removal of urethral catheters has become commonplace. In our series of patients, complications were infrequent and removal improved patient mobility and reduced discomfort. PMID- 10719934 TI - Radical prostatectomy and collaborative care pathways. AB - The purpose of this study was to describe the development, implementation, and evaluation of a collaborative care pathway for radical retropubic prostatectomy. The experience of Vanderbilt University Medical Center is described, and the literature was reviewed. In addition, an example of the radical prostatectomy care pathway is provided for an illustration of a care path. The experience of using collaborative care pathways at Vanderbilt Medical Center has resulted in decreased length of stays, decreased blood utilization, and decreased overall costs. Also, staff satisfaction has improved all without compromising patient care. Collaborative care pathways have been shown to be a cost effective way of standardizing patient care without sacrificing the quality of patient care. In addition, they allow for an easier way to critically evaluate outcomes, costs, and patient and staff satisfaction. PMID- 10719935 TI - Urinary incontinence after radical prostatectomy. AB - Despite improvements in knowledge and technique, a growing number of patients experience incontinence after radical prostatectomy. This may be the result of damage to sphincteric structures, bladder dysfunction, an obstructive stricture, or some combination of these. After an appropriate interval to allow for improvement, the patient should undergo a thorough evaluation to assess the contribution of the various causes and should then be managed using a sequential treatment approach. Following restoration of adequate emptying, bladder dysfunction should be controlled first, if present, and persistent stress incontinence should then be managed according to its severity. Many patients with significant persistent incontinence after radical prostatectomy will need to consider placement of an artificial urinary sphincter. PMID- 10719936 TI - Erectile function after radical prostatectomy. AB - The early detection of prostate cancer through the use of prostate-specific antigen screening has resulted in the performance of many more radical prostatectomy procedures as a curative treatment for this disease. Many patients who are candidates for this procedure already suffer from erectile dysfunction, and the incidence of inadequate erections following radical prostatectomy is certainly high. Nerve-sparing procedures during performance of this operation are encouraged as the incidence of erectile dysfunction is lower if one or both nerves are spared. If the patient is already impotent before the procedure, medical treatments with oral agents, intraurethral compounds, or intracorporally injected medications may be more effective with the nerves intact. Early institution of medical therapy, specifically intracorporal injections, after 2 months postoperatively has resulted in a higher incidence of spontaneous return of erections at 1 year. Vacuum erection devices may be successful in restoring erections but extensive practice in their use is necessary, and they may be unappealing to many patients. A penile prosthesis will restore erections if the patient is so motivated for implantation of such a device. These are expensive and require invasive surgery, but satisfaction rates among patients and partners who have used them have been in the range of 85%, the highest satisfaction rate among all of the treatments of erectile dysfunction. PMID- 10719937 TI - Comparison of patients' and physicians' rating of urinary incontinence following radical prostatectomy. AB - One of the most important endpoints following a radical prostatectomy focuses on the recovery of urinary continence; however, the reported incontinence rates have been quite variable. In men with prostate cancer, it has been found that the physician's assessment of a patient's symptom does not correlate with the patient's own assessment. To further explore the differences in the reported outcomes between physicians and patients, we evaluated the assessment of urinary incontinence in a cohort of men undergoing radical prostatectomy. A total of 145 individuals completed a brief urinary continence questionnaire postoperatively at the 1-year anniversary of their operation and also had the physicians' assessment of incontinence documented in the medical record. Patient-reported incontinence rates varied from 13% to 65% depending on the definition of incontinence applied and the greatest agreement was seen when the physicians' assessment of incontinence was compared with the patient's report of pad use and urinary bother. These comparisons resulted in only moderate to good levels of agreement, which suggests that a more reliable and accurate means to evaluate urinary incontinence following radical prostatectomy needs to be developed. PMID- 10719938 TI - Hemoglobin solutions come of age. PMID- 10719939 TI - Hemoglobin-based oxygen-carrying solutions: close but still so far. PMID- 10719940 TI - Location, location, location. PMID- 10719941 TI - How do we measure (the cost of) pain relief? PMID- 10719942 TI - Randomized trial of diaspirin cross-linked hemoglobin solution as an alternative to blood transfusion after cardiac surgery. The DCLHb Cardiac Surgery Trial Collaborative Group. AB - BACKGROUND: Risks associated with transfusion of allogeneic blood have prompted development of methods to avoid or reduce blood transfusions. New oxygen-carrying compounds such as diaspirin cross-linked hemoglobin (DCLHb) could enable more patients to avoid allogeneic blood transfusion. METHODS: The efficacy, safety, hemodynamic effects, and plasma persistence of DCLHb were investigated in a randomized, active-control, single-blind, multicenter study in post-cardiac bypass surgery patients. Of 1,956 screened patients, 209 were determined to require a blood transfusion and met the inclusion criteria during the 24-h post cardiac bypass period. These patients were randomized to receive up to three 250 ml infusions of DCLHb (n = 104) or three units of packed erythrocytes (pRBCs; n = 105). Further transfusions of pRBCs or whole blood were permitted, if indicated. Primary efficacy end points were the avoidance of blood transfusion through hospital discharge or 7 days postsurgery, whichever came first, and a reduction in the number of units of pRBCs transfused during this same time period. Various laboratory, physiologic, and hemodynamic parameters were monitored to define the safety and pharmacologic effect of DCLHb in this patient population. RESULTS: During the period from the end of cardiopulmonary bypass surgery through postoperative day 7 or hospital discharge, 20 of 104 (19%) DCLHb recipients did not receive a transfusion of pRBCs compared with 100% of control patients (P < 0.05). The overall number of pRBCs administered during the 7-day postoperative period was not significantly different. Mortality was similar between the DCLHb (6 of 104 patients) and the control (8 of 105 patients) groups. Hypertension, jaundice/hyperbilirubinemia, increased serum glutamic oxalo-acetic transaminase, abnormal urine, and hematuria were reported more frequently in the DCLHb group, and there was one case of renal failure in each group. The hemodynamic effects of DCLHb included a consistent and slightly greater increase in systemic and pulmonary vascular resistance with associated increases in systemic and pulmonary arterial pressures compared with pRBC. Cardiac output values decreased more in the DCLHb group patients after the first administration than the control group patients. At 24 h postinfusion, the plasma hemoglobin level was less than one half the maximal level for any amount of DCLHb infused. CONCLUSIONS: Administration of DCLHb allowed a significant number (19%) of cardiac surgery patients to avoid exposure to erythrocytes postoperatively. PMID- 10719943 TI - Changes in intravascular volume during acute normovolemic hemodilution and intraoperative retransfusion in patients with radical hysterectomy. AB - BACKGROUND: Changes in blood volume during acute normovolemic hemodilution (ANH) and their consequences for the perioperative period have not been investigated sufficiently. METHODS: In 15 patients undergoing radical hysterectomy, preoperative ANH to a hematocrit of 24% was performed using 5% albumin solution. Intraoperatively, saline 0.9% solution was used for volume substitution, and intraoperative retransfusion was started at a hematocrit of 20%. Plasma volume (indocyanine green dilution technique), hematocrit, and plasma protein concentration were measured before and after ANH, before retransfusion, and postoperatively. Red cell volume (labeling erythrocytes with fluorescein) was determined before and after ANH and postoperatively. RESULTS: Mean normal plasma volumes (1,514 +/- 143 ml/m2) and reduced red cell volumes (707 +/- 79 ml/m2) were measured preoperatively. Blood (1,150 +/- 196 ml) was removed and replaced with 1,333 +/- 204 ml of colloid. Blood volume before and after ANH was equal and amounted to 3,740 ml. Intraoperatively, plasma volume did not increase until retransfusion despite infusing 3,389 +/- 1,021 ml of crystalloid (corrected for urine output) to compensate for an estimated surgical blood loss of 727 +/- 726 mi. Postoperatively, after retransfusion of all autologous blood, blood volume was 255 +/- 424 ml higher than preoperatively before ANH. Despite mean calculated blood loss of 1,256 +/- 892 ml, only one patient received allogeneic blood. CONCLUSIONS: During ANH, normovolemia was exactly maintained. After surgical blood loss of 1,256 +/- 892 ml, crystalloid and colloid supplies of 5,752 +/- 1,462 ml and 1,667 +/- 548 ml, respectively, and complete intraoperative retransfusions of autologous blood in every patient, mean blood volume was 250 ml higher than preoperatively before ANH. PMID- 10719944 TI - Multiple dose pharmacokinetics of oral transmucosal fentanyl citrate in healthy volunteers. AB - BACKGROUND: Oral transmucosal fentanyl citrate (OTFC) is a solid form of fentanyl that delivers the drug through the oral mucosa. The clinical utility of multiple doses of OTFC in the treatment of "breakthrough" cancer pain is under evaluation. The aim of this study was to test the hypothesis that the pharmacokinetics of OTFC do not change with multiple dosing. METHODS: Twelve healthy adult volunteers received intravenous fentanyl (15 microg/kg) or OTFC (three consecutive doses of 800 microg) on separate study sessions. Arterial blood samples were collected for determination of fentanyl plasma concentration by radioimmunoassay. The descriptive pharmacokinetic parameters (maximum concentration, minimum concentration, and time to maximum concentration) were identified from the raw data and subjected to a nonparametric analysis of variance. Population pharmacokinetic models for all subjects and separate models for each subject were developed to estimate the pharmacokinetic parameters of fentanyl after multiple OTFC doses. RESULTS: The shapes of the profiles of plasma concentration versus time for each dose of OTFC were grossly similar. No change was noted for maximum concentration or time to maximum concentration over the three doses, while minimum concentration did show a significantly increasing trend. Terminal half lives for intravenous fentanyl and OTFC were similar. A two-compartment population pharmacokinetic model adequately represented the central tendency of the data from all subjects. Individual subject data were best described by either two- or three-compartment pharmacokinetic models. These models demonstrated rapid and substantial absorption of OTFC that did not change systematically with time and multiple dosing. CONCLUSIONS: The pharmacokinetics of OTFC were similar among subjects and did not change with multiple dosing. Multiple OTFC dosing regimens within the dosage schedule examined in this study can thus be formulated without concern about nonlinear accumulation. PMID- 10719945 TI - Comparison of blood-conservation strategies in cardiac surgery patients at high risk for bleeding. AB - BACKGROUND: Aprotinin and tranexamic acid are routinely used to reduce bleeding in cardiac surgery. There is a large difference in agent price and perhaps in efficacy. METHODS: In a prospective, randomized, partially blinded study, 168 cardiac surgery patients at high risk for bleeding received either a full-dose aprotinin infusion, tranexamic acid (10-mg/kg load, 1-mg x kg(-1) x h(-1) infusion), tranexamic acid with pre-cardiopulmonary bypass autologous whole-blood collection (12.5% blood volume) and reinfusion after cardiopulmonary bypass (combined therapy), or saline infusion (placebo group). RESULTS: There were complete data in 160 patients. The aprotinin (n = 40) and combined therapy (n = 32) groups (data are median [range]) had similar reductions in blood loss in the first 4 h in the intensive care unit (225 [40-761] and 163 [25-760] ml, respectively; P = 0.014), erythrocyte transfusion requirements in the first 24 h in the intensive care unit (0 [0-3] and 0 [0-3] U, respectively; P = 0.004), and durations of time from end of cardiopulmonary bypass to discharge from the operating room (92 [57-215] and 94 [37, 186] min, respectively; P = 0.01) compared with the placebo group (n = 43). Ten patients in the combined therapy group (30.3%) required transfusion of the autologous blood during cardiopulmonary bypass for anemia. CONCLUSIONS: The combination therapy of tranexamic acid and intraoperative autologous blood collection provided similar reduction in blood loss and transfusion requirements as aprotinin. Cost analyses revealed that combined therapy and tranexamic acid therapy were the least costly therapies. PMID- 10719946 TI - Extrahepatic metabolism of sevoflurane in children undergoing orthotopic liver transplantation. AB - BACKGROUND: Sevoflurane is metabolized by cytochrome P450 and produces inorganic fluoride. The anhepatic phase of liver transplantation provides a useful tool to study the extrahepatic metabolism of drugs. The authors therefore studied the extrahepatic metabolism of sevoflurane by measuring the fluoride production in children receiving sevoflurane solely during the anhepatic phase of orthotopic liver transplantation. METHODS: Children with end-stage liver disease undergoing orthotopic liver transplantation were studied. Anesthesia was provided with isoflurane, sufentanil, and pancuronium. In one group, isoflurane was replaced by sevoflurane as soon as the liver was removed from the patient and maintained until reperfusion of the new liver. Arterial blood samples were drawn at induction, before removal of the liver, 15 min and 30 min after the beginning of the anhepatic phase, at the unclamping of the new liver, and finally 60 and 120 min after the unclamping. Plasma fluoride concentrations were determined by ion selective electrode. RESULTS: No differences between the two groups (n = 10) regarding age, weight, duration of the anhepatic phase, or basal level of inorganic fluoride were found. The fluoride concentration increased significantly as soon as sevoflurane was introduced; it remained stable in the group receiving isoflurane. The peak fluoride concentration was also significantly higher in the first group (mean +/- SD: 5.5 +/- 0.8 microM (sevoflurane group) versus 1.4 +/- 0.5 microM (isoflurane group) P < 0.05). CONCLUSIONS: These results demonstrate the existence of an extrahepatic metabolism of sevoflurane at least in children with end-stage liver disease. PMID- 10719947 TI - Comparison of conventional surgical versus Seldinger technique emergency cricothyrotomy performed by inexperienced clinicians. AB - BACKGROUND: Cricothyrotomy is the ultimate option for a patient with a life threatening airway problem. METHODS: The authors compared the first-time performance of surgical (group 1) versus Seldinger technique (group 2) cricothyrotomy in cadavers. Intensive care unit physicians (n = 20) performed each procedure on two adult human cadavers. Methods were compared with regard to ease of use and anatomy of the neck of the cadaver. Times to location of the cricothyroid membrane, to tracheal puncture, and to the first ventilation were recorded. Each participant was allowed only one attempt per procedure. A pathologist dissected the neck of each patient and assessed correctness of position of the tube and any injury inflicted. Subjective assessment of technique and cadaver on a visual analog scale from 1 (easiest) to 5 (worst) was conducted by the performer. RESULTS: Age, height, and weight of the cadavers were not different. Subjective assessment of both methods (2.2 in group 1 vs. 2.4 in group 2) and anatomy of the cadavers (2.2 in group 1 vs. 2.4 in group 2) showed no statistically significant difference between both groups. Tracheal placement of the tube was achieved in 70% (n = 14) in group 1 versus 60% (n = 12) in group 2 (P value not significant). Five attempts in group 2 had to be aborted because of kinking of the guide wire. Time intervals (mean +/- SD) were from start to location of the cricothyroid membrane 7 +/- 9 s (group 1) versus 8 +/- 7s (group 2), to tracheal puncture 46 +/- 37s (group 1) versus 30 +/- 28s (group 2), and to first ventilation 102 +/- 42s (group 1) versus 100 +/- 46s (group 2) (P value not significant). CONCLUSIONS: The two methods showed equally poor performance. PMID- 10719948 TI - Heterogenous patterns of sensory dysfunction in postherpetic neuralgia suggest multiple pathophysiologic mechanisms. AB - BACKGROUND: Postherpetic neuralgia (PHN) is considered by some investigators to be predominantly a deafferentation-type central pain syndrome; others suggest that activity of remaining peripheral nociceptors plays a critical role. The authors investigated the sensory dysfunction in subjects with PHN of varying duration and at different sites to gain further insight into the mechanisms responsible for the clinical features of neuropathic pain. In addition, the relationships between ongoing pain and pain evoked by mechanical and thermal stimuli were compared in patients with trigeminal and truncal PHN, to determine if the pathophysiologic mechanisms differed among subjects. METHODS: In 63 subjects with PHN, quantitative sensory testing was performed in the region of maximum allodynia or ongoing pain and the corresponding contralateral site. The intensity of ongoing pain was recorded. Sensory thresholds for warmth, coolness, heat pain, and cold pain were determined. Pain induced by various mechanical stimuli (dynamic, static, punctate) was rated using a numerical rating scale of 0 10. RESULTS: The mean rating of ongoing PHN pain was 7.3 +/- 2.0 (mean +/- SD). Allodynia induced by one or more mechanical stimuli was observed in 78% of subjects. A smaller subset (40%) had hyperalgesia to heat or cold stimuli. In subjects with duration of PHN of < or = 1 yr duration, but not in those with duration of > 1 yr, the intensity of ongoing pain correlated with intensity of allodynia induced by dynamic stimuli. Deficits in thresholds for heat and cold pain were observed in the affected region of subjects with PHN in the thoracic dermatomes (P < 0.005), but not in the trigeminal distribution. No relationship was observed between the thermal deficits and ongoing pain or mechanical allodynia in the groups of subjects with either trigeminal or thoracic PHN. CONCLUSION: Despite a common cause, the patterns of sensory abnormalities differ between subjects. Particular differences were noted between groups with facial or truncal PHN and between groups with recent or more chronic PHN. The observations suggest that the relative contributions of peripheral and central mechanisms to the pathophysiology of pain differ among subjects and may vary over the course of PHN. PMID- 10719949 TI - Effect of ambient temperature on human pain and temperature perception. AB - BACKGROUND: Animal studies show reduced nociceptive responses to noxious heat stimuli and increases in endogenous beta-endorphin levels in cold environments, suggesting that human pain perception may be dependent on ambient temperature. However, studies of changes in local skin temperature on human pain perception have yielded variable results. This study examines the effect of both warm and cool ambient temperature on the perception of noxious and innocuous mechanical and thermal stimuli. METHODS: Ten subjects (7 men and 3 women, aged 20-23 yr) used visual analog scales to rate the stimulus intensity, pain intensity, and unpleasantness of thermal (0-50 degrees C) and mechanical (1.2-28.9 g) stimuli applied on the volar forearm with a 1-cm2 contact thermode and von Frey filaments, respectively. Mean skin temperatures were measured throughout the experiment by infrared pyrometer. Each subject was tested in ambient temperatures of 15 degrees C (cool), 25 degrees C (neutral), and 35 degrees C (warm) on separate days, after a 30-min acclimation to the environment. Studies began in the morning after an 8-h fast. RESULTS: Mean skin temperature was altered by ambient temperature (cool room: 30.1 degrees C; neutral room: 33.4 degrees C; warm room: 34.5 degrees C; P < 0.0001). Ambient temperature affected both heat (44-50 degrees C) and cold (25-0 degrees C) perception (P < 0.01). Stimulus intensity ratings tended to be lower in the cool than in the neutral environment (P < 0.07) but were not different between the neutral and warm environments. Unpleasantness ratings revealed that cold stimuli were more unpleasant than hot stimuli in the cool room and that noxious heat stimuli were more unpleasant in a warm environment. Environmental temperature did not alter ratings of warm (37 and 40 degrees C) or mechanical stimuli. CONCLUSIONS: These results indicate that, in humans, a decrease in skin temperature following exposure to cool environments reduces thermal pain. Suppression of Adelta primary afferent cold fiber activity has been shown to increase cold pain produced by skin cooling. Our current findings may represent the reverse phenomenon, i.e., a reduction in thermal nociceptive transmission by the activation of Adelta cutaneous cold fibers. PMID- 10719950 TI - Effect of edrophonium and neostigmine on the pharmacokinetics and neuromuscular effects of mivacurium. AB - BACKGROUND: Previous studies demonstrated that both edrophonium and neostigmine affect mivacurium's pharmacokinetics, thereby potentially affecting its recovery profile. However, those studies were not clinically relevant because mivacurium was still infused after the antagonists were given. In the present study, the authors gave antagonists (or placebo) after discontinuing a mivacurium infusion, thereby obtaining data that are more clinically relevant. METHODS: In 18 patients, mivacurium was infused at 10 microg kg(-1) x min(-1) for 40 min, the infusion was discontinued for 15 min and then restarted at the same rate for another 40 min. Patients were randomized to receive 500 microg/kg edrophonium, 50 microg/kg neostigmine, or saline at discontinuation of the second infusion; all subjects received 1 mg atropine. Plasma was sampled during the final 10 min of each infusion to determine steady state mivacurium concentrations and for 15 min after each infusion. Twitch tension was recorded. Mivacurium concentrations after each of the two infusions were compared. RESULTS: After discontinuation of the second infusion, mivacurium concentrations were larger than those after the first infusion at 2 min with edrophonium and at 2, 4, and 7 min with neostigmine. With both neostigmine and edrophonium, twitch tension recovered after infusion #2 more rapidly than after infusion #1; however, the magnitude of this effect was small CONCLUSION: Edrophonium transiently slows the rate at which mivacurium concentrations decrease; this is consistent with our previous findings. Neostigmine has a similar, although longer, effect. Despite altering mivacurium's elimination characteristics, both drugs facilitate neuromuscular recovery, although their benefit is small. PMID- 10719951 TI - Approximate entropy as an electroencephalographic measure of anesthetic drug effect during desflurane anesthesia. AB - BACKGROUND: The authors hypothesized that the electroencephalogram (EEG) during higher anesthetic concentrations would show more "order" and less "randomness" than at lower anesthetic concentrations. "Approximate entropy" is a new statistical parameter derived from the Kolmogorov-Sinai entropy formula which quantifies the amount of regularity in data. The approximate entropy quantifies the predictability of subsequent amplitude values of the EEG based on the knowledge of the previous amplitude values. The authors investigated the dose response relation of the EEG approximate entropy during desflurane anesthesia in comparison with spectral edge frequency 95, median frequency, and bispectral index. METHODS: Twelve female patients were studied during gynecologic laparotomies. Between opening and closure of the peritoneum, end-tidal desflurane concentrations were varied between 0.5 and 1.6 minimum alveolar concentration (MAC). The EEG approximate entropy, median EEG frequency, spectral edge frequency 95, and bispectral index were determined and the performance of each to predict the desflurane effect compartment concentration, obtained by simultaneous pharmacokinetic-pharmacodynamic modeling, was compared. RESULTS: Electroencephalogram approximate entropy decreased continuously over the observed concentration range of desflurane. The performance of the approximate entropy (prediction probability PK = 0.86 +/- 0.06) as an indicator for desflurane concentrations is similar to spectral edge frequency 95 (PK = 0.86 +/- 0.06) and bispectral index (PK = 0.82 +/- 0.06) and is statistically significantly better than median frequency (PK = 0.78 +/- 0.06). CONCLUSIONS: The amount of regularity in the EEG increases with increasing desflurane concentrations. The approximate entropy could be a useful EEG measure of anesthetic drug effect. PMID- 10719952 TI - Population pharmacokinetics of propofol: a multicenter study. AB - BACKGROUND: Target-controlled infusion is an increasingly common type of administration for propofol. This method requires accurate knowledge of pharmacokinetics, including the effects of age and weight. The authors performed a multicenter population analysis to quantitate the effects of covariates. METHODS: The authors analyzed 4,112 samples of 270 individuals (150 men, 120 women, aged 2-88 yr, weighing 12-100 kg). Population pharmacokinetic modeling was performed using NONMEM (NONMEM Project Group, University of California, San Francisco, CA). Inter- and intraindividual variability was estimated for clearances and volumes. The effects of age, weight, type of administration and sampling site were investigated. RESULTS: The pharmacokinetics of propofol were best described by a three-compartment model. Weight was found to be a significant covariate for elimination clearance, the two intercompartmental clearances, and the volumes of the central compartment, the shallow peripheral compartment, and the deep peripheral compartment; power functions with exponents smaller than 1 yielded the best results. The estimates of these parameters for a 70-kg adult were 1.44 l/min, 2.25 l/min, 0.92 l/min, 9.3 l, 44.2 l, and 266 l, respectively. For patients older than 60 yr the elimination clearance decreased linearly. The volume of the central compartment decreased with age. For children, all parameters were increased when normalized to body weight. Venous data showed a decreased elimination clearance; bolus data were characterized by increases in the volumes of the central and shallow peripheral compartments and in the rapid distribution clearance (Cl2) and a decrease in the slow distribution clearance (Cl3). CONCLUSIONS: Pharmacokinetics of propofol can be well described by a three compartment model. Inclusion of age and weight as covariates significantly improved the model. Adjusting pharmacokinetics to the individual patient should improve the precision of target-controlled infusion and may help to broaden the field of application for target-controlled infusion systems. PMID- 10719953 TI - Comparative spinal distribution and clearance kinetics of intrathecally administered morphine, fentanyl, alfentanil, and sufentanil. AB - BACKGROUND: Despite widespread use, little is known about the comparative pharmacokinetics of intrathecally administered opioids. The present study was designed to characterize the rate and extent of opioid distribution within cerebrospinal fluid, spinal cord, epidural space, and systemic circulation after intrathecal injection. METHODS: Equal doses of morphine and alfentanil, fentanyl, or sufentanil were administered intrathecally (L3) to anesthetized pigs. Microdialysis probes were used to sample cerebrospinal fluid at L2, T11, T7, T3, and the epidural space at L2 every 5-10 min for 4 h. At the end of the experiment, spinal cord and epidural fat tissue were sampled, and each probe's recovery was determined in vitro. Using SAAM II pharmacokinetic modeling software (SAAM Institute, University of Washington, Seattle, WA), the data were fit to a 16-compartment model that was divided into four spinal levels, each of which consisted of a caternary arrangement of four compartments representing the spinal cord, cerebrospinal fluid, epidural space, and epidural fat. RESULTS: Model simulations revealed that the integral exposure (area under the curve divided by dose) of the spinal cord (i.e., effect compartment) to the opioids was highest for morphine because of its low spinal cord distribution volume and slow clearance into plasma The integral exposure of the spinal cord to the other opioids was relatively low, but for different reasons: alfentanil has a high clearance from spinal cord into plasma, fentanyl distributes rapidly into the epidural space and fat, and sufentanil has a high spinal cord volume of distribution. CONCLUSIONS: The four opioids studied demonstrate markedly different pharmacokinetic behavior, which correlates well with their pharmacodynamic behavior. PMID- 10719954 TI - Relationship between local cerebral blood flow and metabolism during mild and moderate hypothermia in rats. AB - BACKGROUND: Hypothermia may interfere with the relationship between cerebral blood flow (CBF) and metabolism. Because this conclusion was based on the analysis of global values, the question remains whether hypothermic CBF/metabolism uncoupling exists on a local cerebral level. This study investigated the effects of hypothermic anesthesia on local cerebral blood flow (LCBF) and local cerebral glucose utilization (LCGU). METHODS: Thirty-six rats were anesthetized with isoflurane (1 minimum alveolar concentration) and artificially ventilated to maintain normal arterial carbon dioxide partial pressure (pH-stat). Pericranial temperature was maintained as normothermic (37.5 degrees C, n = 12) or was reduced to 35 degrees C (n = 12) or 32 degrees C (n = 12). Pericranial temperature was maintained constant for 60 min until LCBF or LCGU were measured by autoradiography. Twelve conscious rats served as normothermic controls. RESULTS: Compared with conscious animals, mean CBF remained unchanged during normothermic anesthesia. Mean CBF significantly increased during mild hypothermia but was unchanged during moderate hypothermia. During normothermic anesthesia, mean CGU was 45% lower than in conscious controls (P < 0.05). No further CGU reduction was found during mild hypothermia, whereas CGU further decreased during moderate hypothermia (48%; P < 0.05). Local analysis showed a linear LCBF/LCGU relationship in conscious (r = 0.94) and anesthetized (r = 0.94) normothermic animals, as well as in both hypothermic groups (35 degrees C: r = 0.92; 32 degrees C: r = 0.95; P < 0.05). The LCBF-to-LCGU ratio increased from 1.4 (conscious controls) to 2.4 (normothermic isoflurane) and 3.6 ml/micromol (mild and moderate hypothermia, P < 0.05). CONCLUSIONS: Decrease of mean CGU at unchanged or increased mean CBF during hypothermic anesthesia may not indicate uncoupling. Local analysis shows a maintained linear relationship that is reset to a higher CBF/CGU ratio. PMID- 10719955 TI - Effect of barbiturates on hydroxyl radicals, lipid peroxidation, and hypoxic cell death in human NT2-N neurons. AB - BACKGROUND: Barbiturates have been shown to be neuroprotective in several animal models, but the underlying mechanisms are unknown. In this study, the authors investigated the effect of barbiturates on free radical scavenging and attempted to correlate this with their neuroprotective effects in a model of hypoxic cell death in human NT2-N neurons. METHODS: Hydroxyl radicals were generated by ascorbic acid and iron and were measured by conversion of salicylate to 2,3 dihydroxybenzoic acid. The effect of barbiturates on lipid peroxidation measured as malondialdehyde and 4-hydroxynon-2-enal was also investigated. Hypoxia studies were then performed on human NT2-N neurons. The cells were exposed to 10 h of hypoxia or combined oxygen and glucose deprivation for 3 or 5 h in the presence of thiopental (50-600 microM), methohexital (50-400 microM), phenobarbital (10 400 microM), or pentobarbital (10-400 microM), and cell death was evaluated after 24 h by lactate dehydrogenase release. RESULTS: Pentobarbital, phenobarbital, methohexital, and thiopental dose-dependently inhibited formation of 2,3 dihydroxybenzoic acid and iron-stimulated lipid peroxidation. There were significant but moderate differences in antioxidant action between the barbiturates. While phenobarbital (10-400 microM) and pentobarbital (10-50 microM) increased lactate dehydrogenase release after combined oxygen and glucose deprivation, thiopental and methohexital protected the neurons at all tested concentrations. At a higher concentration (400 microM), pentobarbital also significantly protected the neurons. At both 50 and 400 microM, thiopental and methohexital protected the NT2-N neurons significantly better than phenobarbital and pentobarbital. CONCLUSIONS: Barbiturates differ markedly in their neuroprotective effects against combined oxygen and glucose deprivation in human NT2-N neurons. The variation in neuroprotective effects could only partly be explained by differences in antioxidant action. PMID- 10719956 TI - Isoflurane increases the apparent agonist affinity of the nicotinic acetylcholine receptor by reducing the microscopic agonist dissociation constant. AB - BACKGROUND: Isoflurane increases the apparent agonist affinity of ligand-gated ion channels. This action reflects a reduction in the receptor's agonist dissociation constant and/or the preopen/open channel state equilibrium. To evaluate the effect of isoflurane on each of these kinetic constants in the nicotinic acetylcholine receptor, the authors analyzed isoflurane's actions on (1) the binding of the fluorescent agonist Dns-C6-Cho to the nicotinic acetylcholine receptor's agonist self-inhibition site and (2) the desensitization kinetics induced by the binding of the weak partial agonist suberyldicholine. METHODS: The dissociation constant for Dns-C6-Cho binding to the self-inhibitory site was determined using stopped-flow fluorescence spectroscopy. The values of the kinetic constants for agonist binding, channel gating, and desensitization were determined by modeling the suberyldicholine concentration-dependence of the apparent rate of desensitization. RESULTS: Isoflurane did not significantly alter the dissociation constant for Dns-C6-Cho binding to the self-inhibitory site even at a concentration as high as 1.5 mM, the highest concentration studied. At this concentration, isoflurane substantially reduced the dissociation constant for suberyldicholine binding to its channel opening site by 97% from 17 +/- 5 microM to 0.5 +/- 0.2 microM, whereas the preopen/open channel state equilibrium was reduced only from 19.1 to 5 +/- 1. CONCLUSIONS: Isoflurane increases the apparent agonist affinity of the nicotinic acetylcholine receptor primarily by reducing the agonist dissociation constant of the site responsible for channel opening rather than altering channel gating kinetics. PMID- 10719957 TI - Complete prevention but stimulus-dependent reversion of morphine tolerance by the glycine/NMDA receptor antagonist (+)-HA966 in neuropathic rats. AB - BACKGROUND: Tolerance to the analgesic effect of morphine complicates the management of chronic pain states. The authors studied the ability of the glycine/N-methyl-D-aspartate receptor antagonist (+)-HA966 to modify morphine tolerance in a rat model of neuropathic pain. METHODS: Mononeuropathy was induced by placing four ligatures around the common sciatic nerve. The 4-day pretreatment regimens with two daily subcutaneous injections of saline and saline, saline and morphine (10 mg/kg), (+)-HA966 (2.5 mg/kg) and morphine, or (+)-HA966 and saline were begun on post-operative day 12 to test the ability of (+)-HA966 to prevent the development of tolerance. Behavioral experiments were performed on postoperative day 16, when the pain-related behavior reached a stable maximum. The effect of an acute dose of morphine (1 mg/kg intravenously) was tested against both mechanical (vocalization threshold to paw pressure) and thermal (struggle latency to hind paw immersion into a 46 degrees C hot-water bath) stimuli. In addition, to test the ability of a single injection of (+)-HA966 to reverse established morphine tolerance, groups of morphine-pretreated rats received injections of either (+)-HA966 (2.5 mg/kg subcutaneously) and morphine (1 mg/kg intravenously), saline and morphine, or (+)-HA966 and saline. RESULTS: Baseline vocalization thresholds and struggle latencies did not differ in the various pretreatment groups, indicating that the pretreatments had no effect on pain-related behavior. Coadministration of (+)-HA966 prevented the reduction of the effect observed with morphine alone in both the mechanical test and the thermal test. (+)-HA966 reversed morphine tolerance in the thermal but not in the mechanical test. CONCLUSION: (+)-HA966 prevented morphine tolerance in both mechanical and thermal tests but reversed established morphine tolerance in the thermal test only. PMID- 10719958 TI - An experimental itch model in monkeys: characterization of intrathecal morphine induced scratching and antinociception. AB - BACKGROUND: The most common side effect of spinal opioid administration is pruritus, which has been treated with a variety of agents with variable success. Currently, there are few animal models developed to study this side effect. The aim of this study was to establish a nonhuman primate model to pharmacologically characterize the effects of intrathecal administration of morphine. METHODS: Eight adult rhesus monkeys were used. Scratching responses were videotaped and counted by observers who were blinded to experimental conditions. Antinociception was measured by a warm-water (50 degrees C) tail-withdrawal assay. The dose response of intrathecal morphine (1-320 microg) for both scratching and antinociception in all subjects was established. An opioid antagonist, nalmefene, was administered either intravenously or subcutaneously to assess its efficacy against intrathecal morphine. RESULTS: Intrathecal morphine (1-32 microg) increased scratching in a dose-dependent manner. Higher doses of intrathecal morphine (10-100 microg) produced thermal antinociception in a dose-dependent manner. On the other hand, nalmefene (10-32 microg/kg intravenously) attenuated maximum scratching responses among subjects. Pretreatment with nalmefene (32 microg/kg subcutaneously) produced approximately 10-fold rightward shifts of intrathecal morphine dose-response curves for both behavioral effects. CONCLUSIONS: These data indicate that intrathecal morphine-induced scratching and antinociception are mediated by opioid receptors. The magnitude of nalmefene antagonism of intrathecal morphine is consistent with microL opioid receptor mediation. This experimental itch model is useful for evaluating different agents that may suppress scratching without interfering with antinociception. It may also facilitate the clarification of mechanisms underlying these phenomena. PMID- 10719959 TI - Isoflurane and pentobarbital reduce AMPA toxicity in vivo in the rat cerebral cortex. AB - BACKGROUND: Isoflurane and pentobarbital can reduce alpha-amino-d-hydroxy-5 methyl-4-isoxazole-propionate (AMPA) receptor-mediated toxicity in vitro. However, their effect on AMPA toxicity in vivo is not known. The present study was undertaken to evaluate the effects of isoflurane and pentobarbital on the in vivo neurotoxicity produced by AMPA. METHODS: Wistar-Kyoto rats were allocated to one of seven groups (n = 8 per group): isoflurane 1 minimum alveolar concentration, isoflurane electroencephalogram burst suppression (EEG-BS), low dose pentobarbital, pentobarbital EEG-BS, NBQX, conscious, and sham groups. AMPA 30 nm was injected into the cortex. An equivalent volume of cerebrospinal fluid was injected into the cortex in the sham group. In the NBQX group, 200 nm NBQX was injected into the cortex with the AMPA. In the isoflurane and pentobarbital groups, anesthesia was maintained for a period of 5 h. Animals in the conscious, NBQX, and sham groups were allowed to awaken immediately after the AMPA injection. Injury to the cortex was evaluated 48 h later. RESULTS: Isoflurane reduced AMPA-induced cortical injury (4.5 +/- 1.9 mm3 and 1.7 +/- 0.8 mm3 in the 1 minimum alveolar concentration and EEG-BS groups, respectively) in comparison to the conscious group (7.2 +/- 0.8 mm3). Pentobarbital reduced cortical injury when administered in EEG-BS doses (2.2 +/- 0.7 mm3) but not when administered in sedative doses (8.6 +/- 0.9 mm3). NBQX reduced AMPA-induced cortical injury (1.2 +/- 0.5 mm3). CONCLUSIONS: Isoflurane and pentobarbital reduced cortical AMPA excitotoxicity. The magnitude of injury reduction was similar to that produced by NBQX when the anesthetics were administered in EEG-BS doses. These results are consistent with the previously demonstrated ability of isoflurane and pentobarbital to inhibit AMPA receptor responses. PMID- 10719960 TI - Role of K+ channels in augmented relaxations to sodium nitroprusside induced by mexiletine in rat aortas. AB - BACKGROUND: A class Ib antiarrhythmic drug, mexiletine, augments relaxations produced by adenosine triphosphate (ATP) sensitive K+ channel openers in isolated rat aortas, suggesting that it produces changes in the vasodilation mediated by ATP-sensitive K+ channels. Nitric oxide can induce its vasodilator effect via K+ channels, including ATP-sensitive K+ channels, in smooth muscle cells. Effects of mexiletine on arterial relaxations to nitric oxide donors, have not been studied. Therefore, the current study in isolated rat aortas was designed to (1) evaluate whether mexiletine augments relaxation in response to nitric oxide donors, including sodium nitroprusside, and (2) determine the role of K+ channels in mediating effects of mexiletine on such nitric oxide-mediated relaxation. METHODS: Rings of rat aortas without endothelia were suspended for isometric force recording. Concentration-response curves of sodium nitroprusside (10(-10) to 10(-5) M) and 1-hydroxy-2-oxo-3-(N-methyl-3-aminopropyl)-3-methyl-1-triazene (NOC-7; 10(-9) to 10(-5) M) were obtained in the absence and in the presence of mexiletine, in combination with a soluble guanylate cyclase inhibitor, 1H [1,2,4]oxadiazolo [4,3,-a]quinoxaline-1-one (ODQ), or inhibitors for ATP sensitive K+ channels (glibenclamide), inward rectifier K+ channels (BaCl2), delayed rectifier K+ channels (4-aminopyridine), large conductance Ca2+-dependent K+ channels (iberiotoxin), or small conductance Ca2+-dependent K+ channels (apamin). RESULTS: Mexiletine (10(-5) or 3 x 10(-5) M) augmented relaxations to sodium nitroprusside and NOC-7. In arteries treated with glibenclamide (10(-5) M), mexiletine (3 x 10(-5) M) did not affect relaxations to nitric oxide donors, whereas mexiletine augmented relaxations to sodium nitroprusside despite the presence of BaCl2 (10(-5) M), 4-aminopyridine (10(-3) M), iberiotoxin (5 x 10(-8) M) and apamin (5 x 10(-8) M). Relaxations to sodium nitroprusside were abolished by ODQ (5 x 10(-6) M), whereas these relaxations were augmented by mexiletine (3 x 10(-5) M) in arteries treated with ODQ (5 x 10(-6) M). CONCLUSIONS: These results suggest that ATP-sensitive K+ channels in vascular smooth muscle, contribute to the augmented vasodilator effect of a nitric oxide donor, sodium nitroprusside induced by mexiletine, and that the vasodilator effect is produced, at least in part, via the guanylate cyclase-independent mechanism. PMID- 10719961 TI - Additive inhibition of nicotinic acetylcholine receptors by corticosteroids and the neuromuscular blocking drug vecuronium. AB - BACKGROUND: Neuromuscular disorders associated with muscular weakness and prolonged paralysis are common in critically ill patients. Acute myopathy has been described in patients receiving a combination therapy of corticosteroids and nondepolarizing neuromuscular blocking drugs for treatment of acute bronchospasm. The cause of this myopathy is not fully established and may involve drug interactions that perturb neuromuscular transmission. To investigate the interaction of corticosteroids with neuromuscular blocking drugs, the authors determined the effects of methylprednisolone and hydrocortisone alone and in combination with vecuronium on fetal (gamma-subunit containing) and adult (epsilon-subunit containing) subtypes of the muscle-type nicotinic acetylcholine receptor. METHODS: Functional channels were expressed in Xenopus laevis oocytes and activated with 1 microM acetylcholine. The resulting currents were recorded using a whole cell two-electrode voltage clamp technique. RESULTS: Both forms of the muscle-type acetylcholine receptor were potently inhibited by methylprednisolone and hydrocortisone, with concentrations producing 50% inhibition in the range of 400-600 microM and 1-2 mM, respectively. The corticosteroids produced noncompetitive antagonism of the muscle-type nicotinic acetylcholine receptor at clinical concentrations. Both receptor forms were also inhibited, even more potently, by vecuronium, with a concentration producing 50% inhibition in the range of 1-2 nM. Combined application of vecuronium and methylprednisolone showed additive effects on both receptor forms, which were best described by a two-site model, with each site independent. CONCLUSIONS: The enhanced neuromuscular blockade produced when corticosteroids are combined with vecuronium may augment pharmacologic denervation and contribute to the pathophysiology of prolonged weakness observed in some critically ill patients. PMID- 10719962 TI - Isoflurane-sevoflurane adminstration before ischemia attenuates ischemia reperfusion-induced injury in isolated rat lungs. AB - BACKGROUND: The effects of volatile anesthetics on ischemia-reperfusion (IR) induced lung injury are not clear. The authors investigated the effects of preadministration of isoflurane and sevoflurane on IR-induced lung injury in an isolated buffer-perfused rat lung model. METHODS: Isolated rat lungs were designated into four groups: control group (n = 6): perfusion for 120 min without ischemia; IR group (n = 6): interruption of perfusion and ventilation for 60 min followed by reperfusion for 60 min; sevoflurane (SEVO)-IR (n = 6) and isoflurane (ISO)-IR (n = 6) groups: 1 minimum alveolar concentration (MAC) isoflurane or sevoflurane was administered for 30 min, followed by 60 min ischemia, then 60 min reperfusion. The authors measured the coefficient of filtration (Kfc) of the lung, lactate dehydrogenase (LDH) activity, tumor necrosis factor alpha, and nitric oxide metabolites (nitrite + nitrate) in the perfusate and the wet-to-dry lung weight ratio. RESULTS: IR caused significant increases in the coefficient of filtration (approximately sevenfold at 60 min of reperfusion compared with baseline; P < 0.01), the wet-to-dry lung weight ratio, the rate of increase of lactate dehydrogenase activity, and tumor necrosis factor a in the perfusate, and caused a significant decrease in nitric oxide metabolites in the perfusate. Administration of 1 MAC isoflurane or sevoflurane before ischemia significantly attenuated IR-induced increases in the coefficient of filtration and the wet-to dry lung weight ratio, inhibited increases in the rate of increase of lactate dehydrogenase activity and tumor necrosis factor alpha in the perfusate, and abrogated the decrease in nitric oxide metabolites in the perfusate. No difference was found between the SEVO-IR and ISO-IR groups. CONCLUSION: Isoflurane and sevoflurane administered before ischemia can attenuate IR-induced injury in isolated rat lungs. PMID- 10719963 TI - Analgesia for labor pain: a cost model. AB - BACKGROUND: Epidural analgesia and intravenous analgesia with opioids are two techniques for the relief of labor pain. The goal of this study was to develop a cost-identification model to quantify the costs (from society's perspective) of epidural analgesia compared with intravenous analgesia for labor pain. Because there is no valid method to assign a dollar value to differing levels of analgesia, the cost of each technique can be compared with the analgesic benefit (patient pain scores) of each technique. METHODS: The authors created a cost model for epidural and intravenous analgesia by reviewing the literature to determine the rates of associated clinical outcomes (benefit of each technique to produce analgesia) and complications (e.g., postdural puncture headache). The authors then analyzed data from their institution's cost-accounting system to determine the hospital cost for parturients admitted for delivery, estimated the cost of each complication, and performed a sensitivity analysis to evaluate the cost impact of changing key variables. A secondary analysis was performed assuming that the cost of nursing was fixed (did not change depending on the number of nursing interventions). RESULTS: If the cesarean section rate equals 20% for both intravenous and epidural analgesia, the additional expected cost per patient to society of epidural analgesia of labor pain ranges from $259 (assuming nursing costs in the labor and delivery suite do not vary with the number of nursing interventions) to $338 (assuming nursing costs do increase as the number of interventions increases) relative to the expected cost of intravenous analgesia for labor pain. This cost difference results from increased professional costs and complication costs associated with epidural analgesia. CONCLUSIONS: Epidural analgesia is more costly than intravenous analgesia. How the cost of the anesthesiologist and nursing care is calculated affects how much more costly epidural analgesia is relative to intravenous analgesia. Published studies have determined that epidural analgesia provides relief of labor pain superior to intravenous analgesia, quantified in one study as 40 mm better on a 100-mm scale during the first stage of labor and 29 mm better during the second stage of labor. Patients, physicians, and society need to weigh the value of improved pain relief from epidural analgesia versus the increased cost of epidural analgesia. PMID- 10719964 TI - How much labor is in a labor epidural? Manpower cost and reimbursement for an obstetric analgesia service in a teaching institution. AB - BACKGROUND: Some anesthesiologists avoid provision of obstetric analgesia services (OAS) because of low reimbursement rates for the work involved. This study defines the manpower costs of operating an OAS in a tertiary referral center and examines reimbursement for this cost. METHODS: The time spent providing OAS in a total of 55 parturients was studied prospectively using a modification of classic time and motion studies. RESULTS: Mean duration of OAS in our population was 412 +/- 313 min. Mean bedside anesthesia staff time was 90 +/- 40 min, and mean number of visits to each patient's bedside was 6.3 +/- 2.0 visits. Assuming staffing on demand for service (intermittent staffing), a minimum of 2.5 full-time equivalent (FTE) attending anesthesiologists was required to meet demand. With intermittent staffing, labor cost was $325 per patient. Actual practice at Duke University Medical Center is around-the-clock (dedicated) staffing, which requires 4.4 FTEs at a cost of $728 per patient. Neither average indemnity reimbursement ($299) nor Medicaid reimbursement ($204) covered the cost per OAS patient. Breaking even is possible under indemnity reimbursement because operating room reimbursement subsidizes OAS costs. Breaking even cannot occur with Medicaid reimbursement under any circumstances. CONCLUSIONS: Obstetric analgesia services requires a minimum of 2.5 FTE attending anesthesiologists at Duke University Medical Center. With the current payer mix, positive-margin operating room activities associated with the obstetric service are not sufficient to compensate for the losses incurred by an OAS. Around-the clock dedicated obstetric staffing (4.4 FTEs) cannot operate profitably under any reasonable circumstances at our institution. PMID- 10719965 TI - New York State guidelines on the topical use of phenylephrine in the operating room. The Phenylephrine Advisory Committee. PMID- 10719966 TI - Xenon anesthesia. PMID- 10719967 TI - Another possible indication for the combined spinal-epidural technique in obstetrics. PMID- 10719968 TI - Combined spinal-epidural analgesia in labor. PMID- 10719969 TI - Postoperative ventricular fibrillation and undiagnosed primary amyloidosis. PMID- 10719970 TI - Fatal perioperative myocardial infarction in four patients with cardiac amyloidosis. PMID- 10719971 TI - Intrathecal baclofen is analgesic in patients with chronic pain. PMID- 10719972 TI - Pulmonary embolism as a consequence of applying sequential compression device on legs in a patient asymptomatic of deep vein thrombosis. PMID- 10719973 TI - Reversal of intraoperative myocardial ischemia with a hemoglobin-based oxygen carrier. PMID- 10719974 TI - Acute myocardial ischemia after administration of ondansetron hydrochloride. PMID- 10719975 TI - Massive airway edema after azathioprine. PMID- 10719976 TI - Successful use of hypnosis as an adjunctive therapy for weaning from mechanical ventilation. PMID- 10719978 TI - Tense diaphragm in tension pneumothorax. PMID- 10719977 TI - Recurrent postoperative stridor requiring tracheostomy in a patient with spasmodic dysphonia. PMID- 10719979 TI - The position of aspect. PMID- 10719980 TI - Sevoflurane and lumbar cerebrospinal fluid. PMID- 10719981 TI - Informed consent issues. PMID- 10719982 TI - Baroreflex activity in hypertensive patients. PMID- 10719983 TI - Hemodilution: fewer keystrokes, greater accuracy. PMID- 10719984 TI - Bispectral index increases and decreases are not always signs of inadequate anesthesia. PMID- 10719985 TI - A method for minimizing leakage during positive pressure ventilation after intubation through laryngeal mask airway. PMID- 10719986 TI - Neuron-specific enolase as a marker of fatal outcome in patients with severe sepsis or septic shock. PMID- 10719987 TI - Uncoiling of wire in arrow flextip epidural catheter on removal. PMID- 10719988 TI - Enterococcus faecalis meningitis after spinal anesthesia. PMID- 10719989 TI - Cessation of paroxysmal atrial fibrillation during acute intravenous propofol administration. PMID- 10719990 TI - Gram-negative rod contamination of an Ohmeda anesthesia machine. PMID- 10719991 TI - Submental orotracheal intubation for maxillofacial surgery. PMID- 10719992 TI - From something old something new. PMID- 10719993 TI - A comparison of modified Howland circuits as current generators with current mirror type circuits. AB - Multi-frequency electrical impedance tomography (EIT) systems require stable voltage controlled current generators that will work over a wide frequency range and with a large variation in load impedance. In this paper we compare the performance of two commonly used designs: the first is a modified Howland circuit whilst the second is based on a current mirror. The output current and the output impedance of both circuits were determined through PSPICE simulation and through measurement. Both circuits were stable over the frequency ranges 1 kHz to 1 MHz. The maximum variation of output current with frequency for the modified Howland circuit was 2.0% and for the circuit based on a current mirror 1.6%. The output impedance for both circuits was greater than 100 kohms for frequencies up to 100 kHz. However, neither circuit achieved this output impedance at 1 MHz. Comparing the results from the two circuits suggests that there is little to choose between them in terms of a practical implementation. PMID- 10719994 TI - High fidelity imaging and high performance computing in nonlinear EIT. AB - We show that nonlinear EIT provides images with well defined characteristics when smoothness of the image is used as a constraint in the reconstruction process. We use the gradient of the logarithm of resistivity as an effective measure of image smoothness, which has the advantage that resistivity and conductivity are treated with equal weight. We suggest that a measure of the fidelity of the image to the object requires the explicit definition and application of such a constraint. The algorithm is applied to the simulation of intra-ventricular haemorrhaging (IVH) in a simple head model. The results indicate that a 5% increase in the blood content of the ventricles would be easily detectable with the noise performance of contemporary instrumentation. The possible implementation of the algorithm in real time via high performance computing is discussed. PMID- 10719995 TI - Real-time three-dimensional electrical impedance imaging. AB - Electrical impedance tomography is a technology for producing images of internal body structures based upon electrical measurements made from electrodes on the body surface. Typically a single plane of electrodes is used, seeking to reconstruct a cross section of the body. Yet the majority of image reconstruction algorithms ignore the three-dimensional (3D) characteristics of the current flow in the body. Actually, a substantial amount of current flows out of the electrode plane, creating distortions in the resulting images. This paper describes a reconstruction algorithm, ToDLeR, for solving a linearized 3D inverse problem in impedance imaging. The algorithm models the body as a homogeneous cylinder and accounts for the 3D current flow in the body by analytically solving for the current flow from one or more layers of electrodes on the surface of the cylinder. The algorithm was implemented on the ACT3 real-time imaging system and data were collected from a 3D test phantom using one, two and four layers of electrodes. By using multiple planes of electrodes, improved accuracy in any particular electrode plane was obtained, with decreased sensitivity to out-of plane objects. A cylindrical target located vertically more than 8 cm below a single layer of 16 electrodes, and positioned radially midway between the centre and the boundary, produced an image that had 35% of the value obtained when the target was in the electrode plane. By adding an additional layer of 16 electrodes below the first electrode plane, and using 3D current patterns, this artefact was reduced to less than 10% of the peak value. We conclude that the 3D algorithm, used with multiple planes of electrodes, reduces the distortions from out-of plane structures in the body. PMID- 10719996 TI - A real-time volumetric visualization system for electrical impedance tomography. AB - Three dimensional (3D) electrical impedance tomography (EIT) presents many additional challenges over and above those associated with two dimensional EIT systems. With present two dimensional (2D) systems, tomographs can be reconstructed and displayed on a PC with a standard computer monitor. In addition, using appropriate data acquisition hardware and simple image reconstruction algorithms, it is possible to collect, reconstruct and display volumetric EIT images in real time using parallel processing architectures. The advantages of this 'real-time' capability are many and include the ability to immediately assess the correct functioning of the system and the ability to track patient events and the effect of procedures in real time. Whilst 3D EIT boundary datasets can be collected in real time, their real-time image reconstruction and display presents some computational challenges. This explains why, to date, no real-time solutions have been presented. In addition the use of a standard computer monitor to display 3D volumes is unsatisfactory since not all depth cues are preserved when using this type of 2D display device. We present a system which is capable of displaying 3D EIT datasets in real time and allows interactive modification of the user's viewpoint. This allows the user to fly around (and through) the EIT volumetric dataset. PMID- 10719997 TI - Algorithms for parametric images in MEIT systems. AB - In this work we show the algorithms developed to extract the Cole parameters from multi-frequency EIT. With these parameters it is possible to obtain information about various different tissues and their pathologies. The algorithms developed obtain the Cole-model parameters from the real and imaginary parts of impedance, or using only the real part, without problems of convergence. A study of the influence of noise is performed with simulations. We find a correct solution in all cases with signal to noise ratio in the data higher than 40 dB. Finally, we show parametric images of the human abdomen obtained with these algorithms. PMID- 10719998 TI - Two-dimensional finite element modelling of the neonatal head. AB - Electrical impedance tomography (EIT) could allow the early diagnosis of infant brain injury following birth asphyxia. The purpose of this work was to determine the effect of variations in skull, scalp or cerebrospinal fluid (CSF) resistivity, as these vary in clinical conditions and could degrade image quality. These factors were investigated using finite element models of the adult and neonatal head. The results suggest that there is a wide range over which the resistivity of the neonatal skull has little effect on the sensitivity to a central impedance change. The scalp and CSF appear to shunt current away from the brain; when their resistivity was decreased from normal values, this shunting effect increased and caused a decrease in sensitivity to a central resistance change. The resistivity of neonatal skull has not, to our knowledge, been directly measured and will anyway vary within and between individuals; this work suggests that EIT will be relatively insensitive to variations in neonatal skull impedance. PMID- 10719999 TI - A simple method to check the dynamic performance of electrical impedance tomography systems. AB - The test concept as well as the design of a simple resistor phantom suitable for the evaluation of the properties of electrical impedance tomographic (EIT) systems is presented. Input and transfer impedance of the phantom are matched with those of the human thorax. Amplitude of the local impedance variations similar to in vivo conditions (ventilation) can be intentionally set to perform measurements on different states. The theoretical potential differences between the electrodes are calculated. The evaluation procedure is performed in terms of the local amplitude of the relative impedance change as well as the local distribution of noise. The whole procedure can be applied either to compare quantitatively the performance of different EIT data acquisition systems or to determine the amount of measurement disturbance caused by the external electrical environment in clinical settings. PMID- 10720000 TI - Sensitivity matrix and reconstruction algorithm for EIT assuming axial uniformity. AB - In electrical impedance tomography (EIT) two-dimensional models continue to be applied despite their known inability to provide correct reconstruction. In this paper, a reconstruction algorithm that assumes a translationally invariant conductivity distribution is described. A more precise forward solver is obtained by taking off-slice currents into consideration. An appropriate sensitivity matrix is derived. Numerical evidence for the improvement in precision compared to two-dimensional reconstruction is given. PMID- 10720001 TI - Electrical impedance imaging at multiple frequencies in phantoms. AB - We have recently built and tested a 32 channel, multi-frequency (1 kHz to 1 MHz) voltage mode system to investigate electrical impedance spectroscopy (EIS) imaging. We completed a series of phantom experiments to define the baseline imaging performance of our system. Our phantom consisted of a plastic circular tank (20 cm diameter) filled with 0.9% aqueous NaCl solution. Conductors and nonconductors of decreasing width (W5: 3.4 cm, W4: 2.54 cm, W3: 0.95 cm, W2: 0.64 cm and WI: 0.32 cm) were positioned at various distances from the tank edge (1 cm, 2 cm, 4 cm and 8 cm). The results suggest that the detection of objects less than 1 cm in width is limited to the first 1 to 2 cm from the tank edge for absolute images, but this depth can extend to 8 cm in difference images. Larger 3.4 cm wide objects can be detected in absolute images at depths up to 8 cm from the tank edge. Generally, conductor images were clearer than their nonconductor counterparts. Not only did electrode artefacts lessen as the frequency increased, but the system's maximum resolution was attained at the highest operating frequencies. Although the system recovered the value of the electrical conductivity at the correct order of magnitude, it tended to smooth out large property discontinuities. The calculated electrical permittivity in these phantom studies was inconclusive due to the presence of electrode artefacts. PMID- 10720002 TI - Movement artefact rejection in impedance pneumography using six strategically placed electrodes. AB - In this paper, we have proposed a technique for reducing movement artefacts in impedance pneumography by placing six electrodes at appropriate locations and suitably combining the measurements obtained. The strategy for electrode placement was based on the observation that the electrodes appeared to slide over the rib cage along with the skin, during movement. A volume conductor model of the thoracic cavity was developed and movement artefacts were simulated by shifting the electrodes to a different location on the surface. The impedance changes due to movement in one of the measurements of a 'symmetrical pair' were 180 degrees out of phase with respect to those observed in the other measurement of that pair. However, the impedance changes due to breathing were in phase in both these measurements. Thus, it was possible to reduce movement artefacts by taking a mean of these measurements without affecting the breathing related changes. The six electrodes could be configured into two such symmetrical pairs. The same observation was made in experimental data recorded from human subjects. This indicated that movement artefacts were caused by sliding of electrodes along with the skin and could be reduced by using the six-electrode configuration. PMID- 10720003 TI - Magnetic induction tomography: experimental realization. AB - Magnetic induction tomography (MIT) is a new non-contacting technique for visualization of the electrical impedance distribution inside inhomogeneous media. A measuring system for MIT has been developed. An oscillating magnetic field is applied in the system as a sounding agent. The system is designed mainly for biomedical applications. Experiments demonstrate that with proper selection of measurement conditions it is possible to use the phase shifts between inductor and detector signals for image reconstruction by filtered backprojection along magnetic lines. Measurements with saline filled phantoms having various spatial distributions of conductivity were carried out and images were reconstructed. The experiments have demonstrated the applicability of MIT for medical imaging and diagnostics. PMID- 10720004 TI - A one step image reconstruction algorithm for electrical impedance tomography in three dimensions. AB - The move from two to three dimensions in the study of electrical impedance tomography (EIT) has generated a great increase in computational demands. It is therefore interesting to investigate ways in which this demand can be reduced, and in this paper we have presented some results of one such approach. The NOSER algorithm was introduced some years ago and we have extended it to include more realistic electrode models. The main feature of the method is that by starting from a uniform conductivity distribution many quantities can be pre-calculated. PMID- 10720005 TI - Multifrequency electrical impedance imaging: preliminary in vivo experience in breast. AB - We have deployed a recently completed spectroscopic electrical impedance tomography (EITS) imaging system in a small series of women (13 participants accrued to date) in order to investigate the feasibility of delivering EITS breast examinations on a routine basis. Hardware is driven with sinusoidally varying spatial patterns of applied voltage delivered to 16 electrodes over the 10 kHz to 1 MHz spectral range using a radially translating interface which couples the electrodes to the breast through direct contact. Imaging examinations have consisted of the acquisition of multi-channel measurements at ten frequencies on both breasts. Participants lie prone on an examination table with the breast to be imaged pendant in the electrode array that is located below the table. Examinations were comfortable and easy to deliver (about 10 minutes per breast including electrode-positioning time). Although localized near-surface electrode artefacts are evident in the acquired images, several findings have emerged. Permittivity images have generally been more informative than their conductivity counterparts, except in the case of fluid-filled cysts. Specifically, the mammographically normal breast appears to have characteristic absolute EITS permittivity and conductivity images that emerge across subjects. Structural features in the EITS images have correlated with limited clinical information available on participants with benign and malignant abnormality, cysts and scarring from previous lumpectomy and follow-up radiation therapy. Several cases from this preliminary experience are described. PMID- 10720006 TI - Systematic errors in multi-frequency EIT. AB - Systematic errors have been measured with a multi-frequency data-collection system operating between 10.24 and 81.92 kHz. The errors were present even though a conventional background measurement on a uniform saline phantom had already been subtracted. Errors due to changes in transimpedance between the calibration and the tissue measurements, cable movement and electrode-skin contact impedance were simulated giving a total systematic error estimate equivalent to a 9% change in tissue conductivity. It was shown that more than 89% of the image was above the total error magnitude, indicating that most of the image revealed true changes in tissue conductivity. In three human subjects, the largest conductivity changes were in two regions, located posteriorly on either side of the midline, and were interpreted as due to the erector spinae muscles. These regions showed increases in conductivity of 73-104%. Identification of other anatomical features was difficult because of the poor spatial resolution of the images. PMID- 10720007 TI - 3D simulation of EIT for monitoring impedance variations within the human head. AB - A preliminary analysis is presented concerning the use of EIT for detecting impedance inhomogeneities within the human brain. The work to date is centred around the monitoring of two distinct impedance variations: those associated with the application of a carotid clamp during surgery and changes caused by the redistribution of blood flow during auditory stimuli. Using the commercially available Ansoft Maxwell package, a 3D finite element model of the human head has been developed to solve the forward problem. The model is hemispherical in shape and comprises regions of brain, cerebrospinal fluid, skull and skin and includes 16 scalp electrodes each of area 1 cm2. Results from simulations using the model suggest that an EIT system, incorporating diametric current excitation, would require a voltage measurement sensitivity of 100-120 dB in order to detect the impedance variations in the above cases. PMID- 10720008 TI - Errors due to the truncation of the computational domain in static three dimensional electrical impedance tomography. AB - In electrical impedance tomography (EIT), an approximation for the internal resistivity distribution is computed based on the knowledge of the injected currents and measured voltages on the surface of the body. The currents spread out in three dimensions and therefore off-plane structures have a significant effect on the reconstructed images. A question arises: how far from the current carrying electrodes should the discretized model of the object be extended? If the model is truncated too near the electrodes, errors are produced in the reconstructed images. On the other hand if the model is extended very far from the electrodes the computational time may become too long in practice. In this paper the model truncation problem is studied with the extended finite element method. Forward solutions obtained using so-called infinite elements, long finite elements and separable long finite elements are compared to the correct solution. The effects of the truncation of the computational domain on the reconstructed images are also discussed and results from the three-dimensional (3D) sensitivity analysis are given. We show that if the finite element method with ordinary elements is used in static 3D EIT, the dimension of the problem can become fairly large if the errors associated with the domain truncation are to be avoided. PMID- 10720009 TI - Cole equation modelling to measurements made using an impulse driven transfer impedance system. AB - Electrical impedance measurements are used to obtain information about a subject, tissue sample or tissue model under test. There are several ways of obtaining these impedance data and thereafter analysing the data to obtain relevant parameters. This paper shows how a completely isolated drive and receive system using current pulses, as opposed to sine waves, achieves good fitted results with resistor-capacitor Cole phantoms. PMID- 10720010 TI - Transthoracic electrical impedance during external defibrillation: comparison of measured and modelled waveforms. AB - The transthoracic electrical impedance is an important defibrillation parameter, affecting the defibrillating current amplitude and energy, and therefore the defibrillation efficiency. A close relationship between transthoracic impedance and defibrillation success rate was observed. Pre-shock measurements (using low amplitude high frequency current) of the impedance were considered a solution for selection of adequate shock voltages or for current-based defibrillation dosage. A recent approach, called 'impedance-compensating defibrillation' was implemented, where the pulse duration was controlled with respect to the impedance measured during the initial phase of the shock. These considerations raised our interest in reassessment of the transthoracic impedance characteristics and the corresponding measurement methods. The purpose of this work is to study the variations of the transthoracic impedance by a continuous measurement technique during the defibrillation shock and comparing the data with results obtained by modelling. Voltage and current impulse waveforms were acquired during cardioversion of patients with atrial fibrillation or flutter. The same type of defibrillation pulse was taken from dogs after induction of fibrillation. The electrodes were located in the anterior position, for both the patients and animals. PMID- 10720011 TI - Diuretic induced change in lung water assessed by electrical impedance tomography. AB - Monitoring patients with left ventricular failure can be difficult. Electrical impedance tomography (EIT) produces cross-sectional images of changes in the impedance of the thorax. We measured changes in the electrical impedance of the lung in nine volunteers following a diuretic challenge. The hypothesis was that lung impedance would increase with diuretic induced fluid loss. Heart rate, blood pressure and urine output were also recorded. After diuretic the mean urine output was 1220 ml compared with 187 ml after placebo. Following diuretic administration, mean thoracic impedance increased by 13.6% (p < 0.01) and lung impedance increased by 7.8% (p < 0.05). Taken as a group there was a correlation between overall impedance change and total urine output. However, for each individual, the time course of change in impedance and urine output did not correlate significantly. Our findings show that EIT may offer a better guide to the response of the lung to diuretic treatment than simply measuring urine output. The urine output is neither specific nor sensitive in the assessment of lung water. Mean lung impedance, however, is largely determined by lung water. The study showed that lung impedance can be recorded at supra-normal values. EIT may help in the management of patients with excess lung water. PMID- 10720012 TI - Field-by-field evaluation of intraoperative transoesophageal echocardiography interpretative skills. AB - A quality assurance system is essential for the credibility and structured growth of anaesthesiology-based transoesophageal echocardiography (TEE) programmes. We have developed software (Q/A Kappa), involving a 400-line source code, capable of directly reporting kappa correlation coefficient values, using external reviewer interpretations as the 'gold standard', and thereby allowing systematic assessment of the validity of intraoperative echocardiographic interpretation. This paper presents assessment of the validity of 240 intraoperative anaesthesiologists' echocardiographic interpretations, and, in addition, the results of field testing of this prototypical software. Data, derived from consecutive cardiac surgery patients, consisted of standardized two-dimensional transoesophageal echocardiographic, colour flow and Doppler imaging sequences. Intraoperative and off-line 'gold standard' TEE interpretations were compared for 19 fields or variables using the Q/A Kappa program. The kappa correlation coefficients were highly variable and dependent on the examination field, ranging from 0.08 for apical regional wall motion scores to 1.00 for tricuspid regurgitation grade, left atrial measurement, aortic valve anatomy and left ventricular long axis and short axis global function. The correlation coefficients were also operator dependent. These data (480 interpretations) were also manually integrated into the equation required for calculation of values of the variable kappa correlation coefficient. The relationship between Q/A Kappa derived values and manually calculated values was highly significant (p < 0.001; r = 1.0). The implications and possible explanations of the results for particular examination fields are discussed. This study also demonstrates successful seamless functioning of this software program from data entry, segmentation into tables and valid statistical analysis. These findings suggest that it is practical to provide sophisticated continuous quality improvement TEE data on a routine basis. PMID- 10720014 TI - A real time programmable digital filter for biomedical signal enhancement incorporating a high-level design interface. AB - A simple but highly integrated digital signal processing system is described for real time filtering of biomedical signals. It includes the necessary processing and communications hardware, the processing code itself and a high-level software interface that enables the user to design and download arbitrary finite impulse response filters to the run-time system. The filter coefficients are calculated using the frequency sampling method. Since the filters are realized using a finite impulse response, no phase distortion is introduced into the processed signals. A unique feature of the design is the manner in which the software and hardware components have been organized as an intelligent system, obviating on the part of the user a detailed knowledge of filter design theory or any abilities in processor architecture and assembly code programming. PMID- 10720013 TI - Evaluation of respiratory influences on left ventricular function parameters extracted from echocardiographic acoustic quantification. AB - This study was designed to assess, using the echocardiographic acoustic quantification technique, the influence of respiration on left ventricular (LV) function and its modifications connected with the ageing process, quantifying in a non-invasive way the respiratory contribution to the LV volume variability. An automated algorithm is applied to extract the beat-to-beat measurements of LV function parameters from the LV volume signal, obtained from recordings lasting a few minutes. Mean values, amount of variability and spectral content were studied in a population of 17 normal young (mean age 25 +/- 1 years) and 12 normal old (mean age 64 +/- 2 years) subjects. Mean values of the beat-to-beat measurements of LV function parameters were able to point out alterations connected with the ageing process in peak filling rate, peak atrial filling rate and peak ejection rate. Spectral analysis, applied to the extracted variability series, displayed a predominance of the high-frequency (HF) component corresponding to respiration in all LV function parameters; moreover, age related changes of HF variability were observed in peak ejection rate. The HF power spectrum component of beat to beat series extracted from the LV signal can provide a non-invasive assessment of the fluctuations in ventricular parameters associated with respiration. PMID- 10720015 TI - System for measuring the transmission spectrum of 'in vitro' corneas. AB - We have developed a spectroscopy system dedicated to eye banks in order to standardize the evaluation of the donated corneas, with respect to their transparency. The system for measuring the corneal transmission spectrum basically consists of a conventional spectroscopy single beam optical apparatus, with particularities in order to attend to the needs of the eye banks, having a linear CCD (2048 sensors) as a detector. The range of evaluation of the system is from 400 to 700 nm, which is the range of interest for these kinds of sample (corneas). Dedicated software has been developed in order to acquire the data from the system and to provide the graphical interface for the user. The software is quite easy to manipulate and provides the diagnostic with respect to the transparency of the cornea, which is based on research done in association with the clinicians of the Eye Bank of Hospital das Clinicas de Ribeirao Preto (Brazil). The system presents a resolution of 9 nm (which is good enough for this kind of measurement, that presents large bandwidths) and it is in agreement with the spectra obtained from commercial spectrophotometers (the correlation factor between our system and a Beckman DU-70 is 0.995 43 for samples of well defined bandwidths, and 0.989 73 for corneas--large bandwidth). PMID- 10720016 TI - Compressed time and frequency recording of the electrogastrogram by individual wave detection. AB - Electrogastrography (EGG) is a method for recording and analysis of the bioelectrical activity of the stomach, acquired by cutaneous electrodes. The problem of obtaining an EGG signal as free of artefact as possible has recently found some solutions. Good quality recordings of several hours duration can be acquired, often needed, especially if clinical applications are envisaged. Therefore, a next problem to be considered would be time-compressed presentation of the EGG signal without loss of relevant signal characteristics. Relative amplitude changes can be of clinical value, while frequency variations are traditionally regarded as more important. A method for simple compressed presentation of long term EGG recordings, without losing information about the real amplitude and/or frequency changes or transients, is presented. It makes use of detection of successive wave extrema in an EGG. Successive wave amplitudes and durations are measured and momentaneous frequencies are derived. Then a compressed two-trace plot is generated, representing amplitude and frequency variations with respect to time. The method is virtually insensitive to baseline drift, as only the peak amplitudes are detected and followed. A compression factor with respect to the traditional signal recording (1 cm min(-1) or 2 cm min(-1) paper speed) of 5 up to 15 per trace can be obtained. PMID- 10720017 TI - Clinical review 112: Does serum growth hormone (GH) binding protein reflect human GH receptor function? AB - Previous observations raised the possibility that circulating GH-binding protein (GHBP) may serve as a useful index for tissue GH receptor (GHR) responsiveness in humans. Indeed, there are many examples to indicate that across a wide scope of comparative studies, ontogenic data, experimental systems, physiological conditions, nutritional states, and diseases there is a close relationship between the concentration of GHR and the level of serum GHBP. In the present review, we discuss various aspects that might affect differentially cellular GHR and circulating GHBP, based on species and tissue divergence, regulation of cell surface GHR turnover, GHR cleavage mechanism, GHR mRNA splicing, and GH insensitivity (GHI) syndrome patients with normal or high serum GHBP levels. Most previous experimental data were collected through comparative analysis of human GHBP against GHR and GHBP determinations in animal models. Yet, GHBPs possess species-specific properties, and the mechanism for their generation and regulation display evolutionary divergence. Another important aspect is tissue divergence, in terms of GHR regulation and its cleavage to GHBP. Although GHBP is generated mainly from the liver GHR, many other tissues express GHRs and probably also contribute to the total GHBP level. Human GHBP is generated by proteolytic cleavage of GHR at the cell-surface and, thus, occupancy or modulation of GHR turnover/internalization would impact the level of cell-surface GHR that are available for proteolysis. An additional degree of complexity arises from recent reports, implicating a protein kinase C-regulated metalloprotease activity in GHBP generation. This suggests that the proteolytic system, which controls the specific cleavage mechanism and switch between GHR proteolysis and GHBP shedding, is a regulated process. Finally, differential splicing regulation to the full length, active human GHR (hGHR) and the inactive truncated hGHRtr isoform messenger RNA transcripts might regulate both the production of GHBP and GHR bioactivity, as hGHRtr generates large amounts of GHBP but has a dominant negative effect on GH signaling. Several clinical GH-resistant conditions, such as liver cirrhosis, renal insufficiency, insulin-dependent diabetes mellitus, hypothyroidism, malnutrition, or critical illness are associated with reduced GHBP levels. However, this is not universally true, as in other conditions (e.g. early childhood, acromegaly) decreased GHBP levels are not associated with GHI. Divergence between serum GHBP and insulin-like growth factor I, such as which occur during puberty or obesity, also questions whether GHBP levels reflect GHR function. Even in patients with GHI syndrome, serum GHBP cannot be relied on to detect all GHR mutations. The correct assessment of GHR expression and GH functionality in an individual patient will require, in parallel to measurements of serum GHBP, additional detailed diagnostic screening of the entire GH-insulin like growth factor I axis. PMID- 10720018 TI - Treatment of growth hormone deficiency in adults. AB - In analogy with other hormonal replacement therapy GH treatment should be commenced with a low starting dose, independent of body weight or body surface area. Hormonal replacement should mimic the normal physiology to minimize the risk of side effects in the life-long replacement of adults. We should, therefore, consider individual responsiveness and also be aware of the difference between pattern of GH under normal condition and during s.c. administration. The safety and monitoring of GH replacement therapy in adults have been addressed in the Growth Hormone Research Society Consensus Guidelines for Diagnosis and Treatment of Adults with GH Deficiency from the Port Stephens Workshop, April 1997. Besides finding better and more accurate biochemical markers for choosing correct GH replacement dose, future research should address the long-term benefits and safety with GH replacement in adults, with special emphasize on incipient risks in terms of cardiovascular disease and of neoplasia, in particular. PMID- 10720019 TI - Severe hypertension induced by the long-acting somatostatin analogue sandostatin LAR in a patient with diabetic autonomic neuropathy. AB - A 26-yr-old woman with type 1 diabetes and severe symptomatic autonomic neuropathy was treated with the long-acting somatostatin analogue Sandostatin LAR for intractable diarrhea. Her diarrhea had previously been successfully managed with three daily injections of octreotide without adverse consequences. She was given a single dose of Sandostatin LAR and within 2 weeks reported the development of increasingly frequent and severe headaches. Three weeks after the injection, she was admitted to hospital with severe hypertension, which eventually resolved with the administration of antihypertensive agents. No other underlying cause of the hypertension was discovered. Rechallenge of the patient with octreotide resulted in a transient hypertensive episode, which lasted 3 h. Severe hypertension, therefore, seems to be a possible adverse effect of treatment of diabetic diarrhea with somatostatin analogues, which should be used with great caution in subjects with severe autonomic dysfunction. PMID- 10720020 TI - Acute ischemic stroke in a young woman with the thiamine-responsive megaloblastic anemia syndrome. PMID- 10720021 TI - Extraocular muscle antibodies positivity as the only serum marker of euthyroid Graves' ophthalmopathy following subacute thyroiditis: case report. PMID- 10720022 TI - Recurrent acute suppurative thyroiditis in an adult due to a fourth branchial pouch fistula. PMID- 10720023 TI - Respiratory fitness, free living physical activity, and cardiovascular disease risk in older individuals: a doubly labeled water study. AB - The objective of this study was to examine the importance of cardiorespiratory fitness vs. physical activity energy expenditure on selected cardiovascular disease risk factors in older individuals. One hundred and seventeen older individuals, 53 men (68 +/- 9 yr) and 63 women (67 +/- 7 yr), participated in the study. This cohort was divided into 4 groups: 1) high cardiorespiratory fitness and high physical activity, 2) high cardiorespiratory fitness and low physical activity, 3) low cardiorespiratory fitness and high physical activity, and 4) low cardiorespiratory fitness and low physical activity. Cardiorespiratory fitness (VO2max) was determined from a graded exercise test, physical activity energy expenditure was measured by doubly labeled water and indirect calorimetry, body composition was determined by dual energy x-ray absorptiometry, and dietary practices were determined by a 3-day recall. Cardiorespiratory fitness exerted greater effects on the cardiovascular disease risk profile than physical activity. That is, older individuals with higher levels of cardiorespiratory fitness, regardless of their physical activity levels, showed lower levels of fasting insulin (P < 0.01), triglycerides (P < 0.05), total cholesterol (P < 0.05), total to high density lipoprotein cholesterol ratio (P < 0.05), low density lipoprotein (P < 0.05), and lower waist circumference (P < 0.01). Moreover, individuals with a high cardiorespiratory fitness but low physical activity energy expenditure displayed a more favorable cardiovascular disease risk profile than individuals with low cardiorespiratory fitness and high physical activity energy expenditure. The results suggest that higher levels of cardiorespiratory fitness have greater cardioprotective effects than higher levels of free living physical activity in older individuals. Although these findings do not discount the health benefits of being physically active, it is possible that greater emphasis should be placed on aerobic exercise to increase cardiorespiratory fitness in the elderly. PMID- 10720024 TI - Pharmacokinetics of transdermal testosterone gel in hypogonadal men: application of gel at one site versus four sites: a General Clinical Research Center Study. AB - Testosterone (T) in a hydroalcoholic gel has been developed as an effective and convenient open system for transdermal delivery of the hormone to men. Because the gel can be applied either to small or large areas of skin, it was important to assess whether the skin surface area on which the gel was applied was an important determinant of serum T levels. To answer this question, the pharmacokinetics of a transdermal 1% hydroalcoholic gel preparation of T was studied in nine hypogonadal men. The subjects applied in random order a 25-mg metered dose of T gel either four times at one site (left arm/shoulder) or at four different sites (left and right arms/shoulders and left and right abdomen) once daily (6-8 min) for 7 consecutive days. After 7 days of washout, each subject was then crossed over to the opposite regimen for another 7 days of treatment. Serum samples were collected for measurements of T, 5alpha dihydrotestosterone (DHT), and estradiol before, during (days 1, 2, 3, 5, and 7), and after (days 8, 9, 11, 13, and 15) application of T gel. Multiple blood samples were drawn on the 1st and 7th day after gel application; single samples were obtained just before the next T gel application on other days (24 h after the previous gel application). The T gel dried in less than 5 min, left no residue, and produced no skin irritation in any of the subjects. Mean serum T levels, irrespective of application at one site or four sites followed the same pattern: rising to 2- to 3- and 4- to 5-fold above baseline at 0.5 and 24 h after first application, respectively. Thereafter, serum T levels reached steady state and remained at 4- to 5-fold above baseline (at the upper limit of the normal adult range) for the duration of gel application and returned to baseline within 4 days after stopping application. The application of T gel at four sites (application skin area approximately four times that of one site) resulted in a mean area under the curve (AUC0-24h) for serum T levels on the 7th day (868 +/- 72 nmol*h/L, mean +/- SEM), which was 23% higher but not significantly different (P = 0.06) than repeated application at one site (706 +/- 59 nmol*h/L). This could be due to the limited number of subjects studied (n = 9). Mean serum DHT levels followed the same pattern as serum T, achieving steady-state levels by 2 days. The mean concentration of serum DHT on the 7th day was significantly higher after application at four sites (9.15 +/- 1.26 nmol/L, P < 0.05) than at one site (6.9 +/- 0.77 nmol/L). These serum DHT levels were at or above the normal adult male range. Serum DHT:T ratio was not significantly altered by T gel application. Serum estradiol levels followed the same pattern as serum T and showed no significant difference between the one- or four-site application. We conclude that transdermal daily application of 100 mg T gel resulted in similar steady levels of serum T. The surface area of the skin to which the gel was applied had only a modest impact on serum T and DHT levels. Mean serum levels of T and DHT was higher by 23% and 33%, respectively, despite application of the gel to four times the skin area in the four sites compared with the one site group. Because of the greater dosage flexibility provided, hydroalcoholic T gel application over multiple sites seems to be an effective and nonskin-irritating method of transdermal T delivery for hypogonadal men. Dose-ranging studies are required to determine dosage regimens for T gel application as a replacement therapy in hypogonadal men. PMID- 10720025 TI - Withdrawal of long-term physiological growth hormone (GH) administration: differential effects on bone density and body composition in men with adult-onset GH deficiency. AB - Adults with acquired GH deficiency (GHD) have been shown to have osteopenia associated with a 3-fold increase in fracture risk and exhibit increased body fat and decreased lean mass. Replacement of GH results in decreased fat mass, increased lean mass, and increased bone mineral density (BMD). The possible differential effect of withdrawal of GH replacement on body composition compartments and regional bone mass is not known. We performed a randomized, single blind, placebo-controlled 36-month cross-over study of GH vs. placebo (PL) in adults with GHD and now report the effect of withdrawal of GH on percent body fat, lean mass, and bone density, as measured by dual energy x-ray absorptiometry. Forty men (median age, 51 yr; range, 24-64 yr) with pituitary disease and peak serum GH levels under 5 microg/L in response to two pharmacological stimuli were randomized to GH therapy (starting dose, 10 microg/kg x day, final dose 4 microg/kg x day) vs. PL for 18 months. Replacement was provided in a physiological range by adjusting GH doses according to serum insulin-like growth factor I levels. After discontinuation of GH, body fat increased significantly (mean +/- SEM, 3.18 +/- 0.44%; P = 0.0001) and returned to baseline. Lean mass decreased significantly (mean loss, 2133 +/- 539 g; P = 0.0016), but remained slightly higher (1276 +/- 502 g above baseline; P = 0.0258) than at study initiation. In contrast to the effect on body composition, BMD did not reverse toward pretreatment baseline after discontinuation of GH. Bone density at the hip continued to rise during PL administration, showing a significant increase (0.0014 +/- 0.00042, g/cm2 x month; P = 0.005) between months 18-36. Every bone site except two (radial BMD and total bone mineral content), including those without a significant increase in BMD during the 18 months of GH administration, showed a net increase over the entire 36 months. Therefore, there is a critical differential response of the duration of GH action on different body composition compartments. Physiological GH administration has a persistent effect on bone mass 18 months after discontinuation of GH. PMID- 10720026 TI - Effect of weight loss with reduction of intra-abdominal fat on lipid metabolism in older men. AB - How weight loss improves lipid levels is poorly understood. Cross-sectional studies have suggested that accumulation of fat in intra-abdominal stores (IAF) may lead to abnormal lipid levels, increased hepatic lipase (HL) activity, and smaller low density lipoprotein (LDL) particle size. To determine what effect loss of IAF would have on lipid parameters, 21 healthy older men underwent diet induced weight loss. During a period of weight stability before and after weight loss, subjects underwent studies of body composition, lipids, measurement of postheparin lipoprotein and HL lipase activities, cholesteryl ester transfer protein activity, and insulin sensitivity (Si). After an average weight loss of 10%, reductions in fat mass, IAF, and abdominal s.c. fat were seen, accompanied by reductions in levels of triglyceride, very low density lipoprotein cholesterol, apolipoprotein B, and HL activity. High density lipoprotein-2 cholesterol and Si increased. In those subjects with pattern B LDL at baseline, LDL particle size increased. Cholesteryl ester transfer protein activity did not change. Changes in IAF and Si correlated with a decrease in HL activity (although not independently of each other). In summary, in men undergoing diet-induced weight loss, only loss of IAF was found to be associated with a reduction in HL, which is associated with beneficial effects on lipid levels. PMID- 10720027 TI - Slow release lanreotide in combination with interferon-alpha2b in the treatment of symptomatic advanced medullary thyroid carcinoma. AB - Somatostatin analogs are promising agents in the treatment of medullary thyroid carcinoma. We have evaluated the effects of the slow release somatostatin analog lanreotide in combination with interferon-alpha2b in seven patients with advanced and symptomatic medullary thyroid carcinoma. The frequency and intensity of daily flushing episodes and bowel movements, the intensity of fatigue, weight, performance status, calcitonin levels, and change in tumor masses were recorded before and during treatment. No objective complete or partial responses were recorded. However, disease stabilization and minor tumor regression were observed in three of seven and two of seven patients, respectively. The number and intensity of bowel movements and flushing episodes decreased in five of six and two of two patients, respectively. Decrease in fatigue and improvement in performance status were observed in five of seven and six of seven patients, respectively. Weight gain was recorded in three of four patients. Plasma levels of calcitonin decreased significantly in six of seven patients. Clinical benefit, evaluated by a structured algorithm, was achieved in six of seven patients and was coupled with a decrease of 50% or more in serum calcitonin levels in three of seven patients. In conclusion, the combination of lanreotide with interferon had a major impact on clinical symptoms and was well tolerated. PMID- 10720028 TI - Embolization for vertebral metastases of follicular thyroid carcinoma. AB - The technique of selective embolization has been applied for years in the treatment of vascular anomalies, severe hemorrhage and benign or malignant tumors, notably vertebral metastases of renal cell carcinoma. Because this technique is relatively easy to perform and offers immediate relief of symptoms, it is an attractive option for patients with vertebral metastases of thyroid carcinoma with signs of spinal cord compression. In these patients, other treatment modalities like radioactive iodine, external irradiation, or surgery are more cumbersome or less effective in the short term. We describe four patients with metastasized follicular thyroid carcinoma, presenting with neurological symptoms due to vertebral metastases. All patients had undergone total thyroidectomy, ranging from 1 month to 4 yr before embolization. Embolization was combined with iodine-131 therapy when appropriate. Selective catheterization of the arteries feeding the metastases was performed, followed by infusion of polyvinyl alcohol particles (Ivalon). The procedure was technically successful in all patients without adverse effects. In the patients described, embolization resulted in rapid resolution of neurological symptoms, sometimes within hours. The therapeutic effect lasted from months to years. We conclude that embolization of vertebral metastases of follicular thyroid carcinoma is an attractive palliative therapeutic option that may offer rapid relief of symptoms. PMID- 10720029 TI - Normal ovulatory women with polycystic ovaries have hyperandrogenic pituitary ovarian responses to gonadotropin-releasing hormone-agonist testing. AB - Women with polycystic ovary syndrome (PCOS) have chronic anovulation and hyperandrogenism and frequently have abnormalities in their lipid profiles and insulin/insulin-like growth factor axis that increase their lifetime risk for cardiovascular disease. Normal ovulatory women may have polycystic ovaries on ultrasonography and yet lack the clinical features of PCOS. To further explore whether ovulatory women without clinical/biochemical hyperandrogenism but with polycystic appearing ovaries (ov-PAO) have subclinical features of PCOS, we prospectively characterized 26 ov-PAO women and matched them by age and body mass index to 25 ovulatory women with normal appearing ovaries (ov-NAO) and to 22 women with PCOS. After an overnight fast, all women had baseline endocrine and metabolic assessments. In addition, a subset of each group of women underwent GnRH-agonist (leuprolide acetate 1 mg s.c.) testing, ACTH stimulation, and an insulin tolerance test (ITT). At baseline, ov-PAO and ov-NAO women had similar endocrine profiles (LH, LH:FSH, androstenedione, and DHEAS). Compared with ov NAO, 31% of ov-PAO women had reduced glucose responses after insulin (K(itt)), suggesting mild insulin resistance, and 35% had high density lipoprotein levels below 35 mg/dL, a level considered to represent significant cardiovascular risk. After GnRH-agonist, ov-PAO women had response patterns in LH, total testosterone, and 17-hydroxyprogesterone (17-OHP) that were intermediate between ov-NAO and women with PCOS. Ovarian responses were above the normal range in 30-40% of women with ov-PAO. In ov-PAO, peak responses of LH after leuprolide correlated with triglyceride levels (P < 0.05) and peak responses of 17-OHP correlated inversely with Kitt values (P < 0.05). No significant differences were noted with ACTH testing. In conclusion, occult biochemical ovarian hyperandrogenism may be uncovered using GnRH-agonist in ovulatory women with ov-PAO, while adrenal responses remain normal. Subtle metabolic abnormalities may also be prevalent. PMID- 10720030 TI - Congenital hypothyroidism with impaired thyroid response to thyrotropin (TSH) and absent circulating thyroglobulin: evidence for a new inactivating mutation of the TSH receptor gene. AB - Congenital hypothyroidism due to impaired thyroid response to TSH was originally described by Stanbury. A diagnosis of congenital hypothyroidism with thyroid unresponsiveness to TSH is accepted if the patient has congenital hypothyroidism, the thyroid gland is in the normal position in the neck, the size of the thyroid is either normal or atrophic, the serum TSH level is increased, the bioactivity of TSH is intact, and the response of the thyroid gland to TSH stimulation is decreased. In all originally described cases serum thyroglobulin was undetectable. We describe a 22-yr-old female patient who was severely hypothyroid and mentally retarded. Serum T4 and T3 concentrations were below the sensitivity of the methods, with elevated serum TSH levels. Serum thyroglobulin was undetectable. A normally shaped hypoplastic gland located in the appropriate anatomical position in the neck was found at scintiscan. The gland did not respond after administration of bovine TSH in terms of 131I uptake, serum thyroid hormones, and thyroglobulin secretion. A diagnosis of congenital hypothyroidism due to TSH unresponsiveness was formulated. Genetic analysis in the propositus showed a homozygous inactivating mutation of the TSH receptor that had not been previously described. The mutation consisted of the substitution of an isoleucine in place of a highly conserved threonine at position 477 in the first extracellular loop of the receptor (T477I). The brother, one sister of the father (whose DNA was not available), the mother of the propositus, one sister, and the brother were heterozygous for T477I. All the heterozygous persons were unaffected. After transfection in COS-7 cells, the mutant receptor displayed an extremely low expression at cell surface. At variance with cells transfected with the wild-type TSH receptor, cells transfected with the mutant T477I did not show constitutive activity for the adenylyl cyclase pathway. A dramatic reduction in the amount of cAMP accumulation after bovine TSH challenge was observed in cells transfected with the mutant T477I receptor. A structural defect in the mutant TSH receptor protein was probably responsible for the poor routing of the receptor to the cell membrane. This is the first time that a loss of function mutation of the TSH receptor is described in a patient with severe congenital hypothyroidism and absent circulating thyroglobulin due to TSH unresponsiveness and the first time that an inactivating mutation of the TSH receptor is described in the first extracellular loop. PMID- 10720031 TI - Activin effects on neoplastic proliferation of human pituitary tumors. AB - Factors underlying growth regulation in human pituitary tumors are largely unknown. Activin functions as an antiproliferative cytokine in a number of cell types and is endogenously expressed in normal and neoplastic human pituicytes. We investigated the effect of activin on proliferation in 16 clinically nonfunctioning pituitary adenomas in primary culture. Treatment for 24 h with activin (0-10 ng/mL) significantly inhibited cell proliferation in 5 tumors (P < 0.05), as determined by [3H]thymidine incorporation. In 9 tumors, we studied regulation of the cyclin-dependent kinase inhibitor p21WAF1/cip1 as a potential activin mediator. In tumors with activin-inhibited proliferation, p21WAF1/cip1 gene expression was up-regulated after 4 h in a dose-dependent manner (0-100 ng/mL). We also investigated tumor expression of follistatin messenger ribonucleic acid, an activin-binding protein with two isoforms of different potencies. In contrast to normal pituitary tissue, only four tumors expressed both follistatin isoforms, and three tumors expressed only the less potent form. Tumors in which activin induced antiproliferative responses showed diminished or no follistatin messenger ribonucleic acid expression compared to normal pituitary. These data indicate that activin has an antiproliferative effect in a subgroup of human pituitary tumors. PMID- 10720032 TI - Hemodynamic, hormonal, and renal effects of short-term adrenomedullin infusion in healthy volunteers. AB - The actions of adrenomedullin (ADM), a 52-amino acid peptide, are not well defined in man. We, therefore, studied eight normal volunteers aged 1832 yr in a placebo-controlled crossover study. On the 2 study days, subjects received, in random order, ADM in "low" and "high" dose (2.9 pmol/kg x min and 5.8 pmol/kg x min for 2 h each) or vehicle (hemaccel) infusion on day 4 of a metabolic diet (Na+ 80 mmol/day, K+ 100 mmol/day). Achieved plasma ADM levels were in the pathophysiological range, and plasma cAMP values rose 5 pmol/L during the higher dose. Compared with time-matched vehicle infusion, high-dose ADM increased peak heart rate by 10 beats per minute (P < 0.05) and lowered diastolic (by 5 mm Hg, P < 0.01) blood pressure. Cardiac output increased in both phases of ADM (low dose, 7.6 L/min; high dose, 10.2 L/min; vehicle, 6 L/min; P < 0.05 for both). Despite a 2-fold rise in PRA during high-dose ADM (P < 0.01), aldosterone levels were unaltered. Norepinephrine levels increased by 50% during high-dose ADM (P < 0.001), but epinephrine levels were unchanged. Plasma PRL levels increased during high-dose ADM (P = 0.014). ADM had no significant effect on urine volume and sodium excretion. Infusion of ADM to achieve pathophysiological plasma levels produced significant hemodynamic effects, stimulated renin but inhibited the aldosterone response to endogenous angiotensin II, and activated the sympathetic system and PRL without altering urine sodium excretion in normal subjects. PMID- 10720033 TI - Rapid maturation of the reproductive axis during perimenarche independent of body composition. AB - The development of the reproductive axis is thought to be a gradual process, but our understanding of the complex endocrine changes that accompany the transition from premenarche to reproductive life in women has been hampered by the paucity of longitudinal studies. We studied 112 premenarchal Caucasian females at 6-month intervals over 4 yr and obtained a detailed reproductive and dietary history. We quantified reproductive hormones in 24-h urine collections as a measure of daily output and measured body composition biometrically and with the use of dual energy x-ray absortiometry scans. The percent body fat did not change appreciably in the study period (range, 21-24%) and was unrelated to menarche. Sex steroid and gonadotropin levels changed exponentially in the year approaching menarche. FSH levels peaked at menarche and then progressively declined thereafter. Estradiol output increased rapidly in the year approaching menarche and then plateaued thereafter. The frequency of menstrual bleeding increased rapidly and plateaued at 1 yr postmenarche. At 1 yr, 65% of these adolescent women had established a pattern of 10 or more menstrual episodes/yr, and by 3 yr postmenarche this figure exceeded 90%. There were no significant changes in dietary intake of protein, carbohydrate, or fat in the same period. Menarche occurs as a result of rapid maturation of the reproductive axis and heralds the reestablishment of a negative sex steroid feedback loop that parallels the adult threshold. These events appear to develop independent of changes in body composition and diet, but may reflect the improved nutrition and socioeconomic status of the late 20th century. PMID- 10720034 TI - Contribution of body fatness and adipose tissue distribution to the age variation in plasma steroid hormone concentrations in men: the HERITAGE Family Study. AB - Obesity has been associated with alterations in plasma steroid hormone concentrations in men. Older men present an altered steroid hormone profile compared to younger individuals, and an increase in body fatness and changes in adipose tissue (AT) distribution are noted with advancing age. Thus, there is a need to examine the relative importance of increased body fatness and changes in AT distribution with advancing age to plasma steroid hormone and sex hormone binding globulin levels in men. We, therefore, investigated the relationships among age, body fatness, AT distribution, and the plasma steroid hormone profile in a group of 217 Caucasian men (mean age +/- SD, 36.2 +/- 14.9 yr) who covered a wide age range (17-64 yr). Compared to young adult men, older men were characterized by increased adiposity (P < 0.0001) expressed either as body mass index or total body fat mass assessed by underwater weighing. Differences in AT distribution were also noted with a preferential accumulation of abdominal fat as indicated by a larger waist girth (P < 0.0001) and higher visceral AT accumulation (P < 0.0001), measured by computed tomography, in older subjects. Age was associated with decreases (P < 0.0001) in C19 adrenal steroid levels, namely reduced dehydroepiandrosterone (DHEA), DHEA fatty acid ester, DHEA sulfate, as well as androstenedione levels. Androgens, i.e. dihydrotestosterone and testosterone, were also affected by age, with lower levels of both steroids being found in older individuals (P < 0.0005). When statistical adjustment for body fatness and AT distribution was performed, differences in C19 adrenal steroids between the age groups remained significant, whereas differences in androgens and sex hormone-binding globulin concentrations were no longer significant. The present study suggests that age-related differences in plasma steroid hormone levels, especially androgens, are partly mediated by concomitant variation in adiposity in men. PMID- 10720035 TI - The contribution of testosterone to skeletal development and maintenance: lessons from the androgen insensitivity syndrome. AB - Although androgen status affects bone mass in women and men, an androgen requirement for skeletal normalcy has not been established. Women with androgen insensitivity syndrome (AIS) have 46,XY genotypes with androgen receptor abnormalities rendering them partially or completely refractory to androgen. Twenty-eight women with AIS (22 complete and 6 high grade partial), aged 11-65 yr, responded to questionnaires about health history, gonadal surgery, and exogenous estrogen use and underwent bone mineral density (BMD) assessment by dual energy x-ray absortiometry. BMD values at the lumbar spine and proximal femur were compared to age-specific female normative values and listed as z scores. Average height for adults in this cohort, 174 cm (68.5 in.), was moderately increased compared with the average height of adult American women of 162.3 cm, with skewing toward higher values: 5 women exceeded 6 ft in height, and 30% of the 18 adult women with complete AIS exceeded 5 ft, 11 in. in height. The average lumbar spine and hip BMD z-scores of the 6 women with partial AIS did not differ from population norms. In contrast, the average lumbar spine BMD z-score of women with complete AIS was significantly reduced at -1.08 (P = 0.0003), whereas the average value for hip BMD did not differ from normal. When BMD was compared between women who reported good estrogen replacement therapy compliance and those who reported poor compliance, there was a significantly greater deficit at the spine for women with poor compliance (z = -2.15 +/- 0.15 vs. -0.75 +/- 0.28; P < .0001). Furthermore, hip BMD was also significantly reduced in the noncompliant group (z = -0.95 +/- .40). Comparison of BMD values to normative male standards gave z-score reductions (z = -1.81 +/- 0.36) greater than those observed with female standards. Because of the high prevalence of tall stature in this study sample, we calculated bone mineral apparent density, a variable that adjusts for differences in bone size. Even for the estrogen-compliant group, bone mineral apparent density z-scores were subnormal at both the spine (z = -1.3 +/- 0.43; P < 0.01) and the hip (z = -1.38 +/- 0.28; P = 0.017). Six women with complete AIS had sustained cortical bone fractures, of whom 3 reported multiple (>3) fractures. We conclude that even when compliance to exogenous estrogen use is excellent, women with complete AIS show moderate deficits in spine BMD, averaging close to 1 SD from normative means, and that with correction of BMD for bone size, skeletal deficits are magnified and include the proximal femur. The results suggest that severe osteopenia in some women with AIS probably reflects a component of inadequate estrogen replacement rather than androgen lack alone. PMID- 10720036 TI - Age and gender predict the outcome of treatment for Graves' hyperthyroidism. AB - The response to treatment in Graves' hyperthyroidism is unpredictable, and factors postulated to predict outcome have not generally proved clinically useful or been widely adopted in clinical practice. We audited outcome in 536 patients with Graves' hyperthyroidism presenting consecutively to determine whether simple clinical features predict disease presentation and response to treatment. At presentation males had slightly more severe biochemical hyperthyroidism [free T4: males, 64.3 +/- 3.0 pmol/L (mean +/- SE); females, 61.3 +/- 1.7 (P = 0.45); free T3: males, 24.3 +/- 1.5 pmol/L; females, 21.0 +/- 0.6, (P = 0.04)]. Patients less than 40 yr at diagnosis had more severe hyperthyroidism than patients more than 40 yr old [free T4: <40 yr, 64.3 +/- 2.0; >40 yr, 56.7 +/- 2.3 (P = 0.02); free T3: <40 yr, 22.8 +/- 0.8; >40 yr, 19.0 +/- 0.9 (P = 0.003)]. Males had a lower remission rate than females after a course of antithyroid medication [19.6% vs. 40%; odds ratio, 0.37; 95% confidence interval (CI), 0.17-0.79; P < 0.01]. Similarly, patients aged less than 40 yr had a lower remission rate than older patients (32.6% vs. 47.8%; odds ratio, 0.53; 95% CI, 0.32-0.87; P = 0.01). One dose of radioiodine cured hyperthyroidism in fewer males than females (47% vs. 74%; P < 0.0001). Logistic regression analysis demonstrated male sex (odds ratio, 2.80; 95% CI, 1.31-5.98; P = 0.008), serum free T4 concentration at diagnosis (odds ratio, 1.02; 95% CI, 1.0-1.04; P = 0.01), and dose of radioiodine administered (odds ratio, 0.99; 95% CI, 0.99-1.00; P = 0.001) were contributing factors associated with failure to respond to a single dose of radioiodine. As males and younger patients are more likely to fail to respond to medical treatment, and male patients are likewise less likely to respond to a single dose of radioiodine, we suggest that those groups with low remission rates should be offered definitive treatment with radioiodine or surgery soon after presentation and that the value of higher initial doses of radioiodine in males be evaluated. PMID- 10720037 TI - Antidiabetogenic action of cholecystokinin-8 in type 2 diabetes. AB - Cholecystokinin (CCK) is a gut hormone and a neuropeptide that has the capacity to stimulate insulin secretion. As insulin secretion is impaired in type 2 diabetes, we explored whether exogenous administration of this peptide exerts antidiabetogenic action. The C-terminal octapeptide of CCK (CCK-8) was therefore infused i.v. (24 pmol/kg x h) for 90 min in six healthy postmenopausal women and in six postmenopausal women with type 2 diabetes. At 15 min after start of infusion, a meal was served and ingested during 10 min. On a separate day, saline was infused instead of CCK-8. In both healthy subjects and subjects with type 2 diabetes, CCK-8 reduced the increase in circulating glucose after meal ingestion and potentiated the increase in circulating insulin. The ratio between the area under the curves for serum insulin and plasma glucose during the 15- to 75-min period after meal ingestion was increased by CCK-8 by 198 +/- 18% in healthy subjects (P = 0.002) and by 474 +/- 151% (P = 0.038) in subjects with type 2 diabetes. In contrast, the increase in the circulating levels of gastric inhibitory polypeptide (GIP), glucagon-like peptide-1 (GLP-1), or glucagon after meal ingestion was not significantly affected by CCK-8. The study therefore shows that CCK-8 exerts an antidiabetogenic action in both healthy subjects and type 2 diabetes through an insulinotropic action that most likely is exerted trough a direct islet effect. As at the same time, CCK-8 was infused without any adverse effects, the study suggests that CCK is a potential treatment for type 2 diabetes. PMID- 10720038 TI - Plasma total homocysteine levels during short-term iatrogenic hypothyroidism. AB - Hypothyroidism is associated with increased cardiovascular morbidity, which cannot be fully explained by the atherogenic lipid profile observed in these patients. We have previously found elevated levels of the cardiovascular risk factor, plasma total homocysteine (tHcy), in hypothyroidism. We conducted a longitudinal study on 17 patients who had undergone total thyroidectomy for thyroid cancer. During 6 weeks of discontinued T4 substitution before radioscintigraphy (phase I), they attained a hypothyroid state, which was reversed by resupplementation (phase II). Plasma tHcy, serum creatinine, serum and red blood cell folate, serum cobalamin, and serum cholesterol were determined at 2-week intervals throughout phases I and II. There was a progressive and parallel increase in tHcy (mean, 27%), serum creatinine (37%), and serum cholesterol (100%) during phase I, and these values returned to the original level within 4-6 weeks after reinitiating T4 therapy. Serum and red blood cell folate levels showed only minor, but statistically significant, changes. In a bivariate model, serum creatinine and serum cholesterol were strongly associated with the changes observed in tHcy during short term hypothyroidism. In conclusion, we found a transient increase in both plasma tHcy and serum cholesterol during short term iatrogenic hypothyroidism, and the tHcy response is probably mainly explained by concurrent changes in renal function. The increase in both plasma tHcy and serum cholesterol may confer increased cardiovascular risk in hypothyroid patients. PMID- 10720040 TI - Predicting phenotype in steroid 21-hydroxylase deficiency? Comprehensive genotyping in 155 unrelated, well defined patients from southern Germany. AB - Congenital adrenal hyperplasia (CAH) is a group of autosomal recessive disorders. CAH is most often caused by deficiency of steroid 21-hydroxylase. The frequency of CYP21-inactivating mutations and the genotype-phenotype relationship were characterized in 155 well defined unrelated CAH patients. We were able to elucidate 306 of 310 disease-causing alleles (diagnostic sensitivity, 98.7%). The most frequent mutation was the intron 2 splice site mutation (30.3%), followed by gene deletions (20.3%), the I172N mutation (19.7%) and large gene conversions (7.1%). Five point mutations were detected that have not been described in other CAH cohorts. Genotypes were categorized in 4 mutation groups (null, A, B, and C) according to their predicted functional consequences and compared to the clinical phenotype. The positive predictive value for null mutations (ppv(null)) was 100%, as all patients with these mutations had a salt-wasting phenotype. In mutation group A (intron 2 splice site mutation in homozygous or heterozygous form with a null mutation), the ppv(A) to manifest with salt-wasting CAH was 90%. In group B predicted to result in simple virilizing CAH (I172N in homozygous or compound heterozygous form with a more severe mutation), ppv(B) was 74%. In group C (P30L, V281L, P453S in homozygous or compound heterozygous form with a more severe mutation), ppv(C) was 64.7% to exhibit the nonclassical form of CAH, but 90% when excluding the P30L mutation. Thus, in general, a good genotype-phenotype relationship is shown in patients with either the severest or the mildest mutations. A considerable degree of divergence is observed within mutation groups of intermediate severity. As yet undefined factors modifying 21-hydroxylase gene expression and steroid hormone action are likely to account for these differences in phenotypic expression. PMID- 10720039 TI - Effect of vitamin D nutrition on parathyroid adenoma weight: pathogenetic and clinical implications. AB - In primary hyperparathyroidism, adenoma size is a major determinant of disease severity and manner of presentation, but the reason for the large variation in size (>100-fold) is unknown. One factor could be the level of vitamin D nutrition, because in India, where vitamin D deficiency is endemic, adenomas are larger and the disease more severe than in the U.S. Accordingly, we determined the relationship between vitamin D nutrition, as measured by serum levels of 25 hydroxyvitamin D (25OHD), and parathyroid gland weight, expressed on a logarithmic scale, in 148 U.S. patients with primary hyperparathyroidism. A significant inverse relationship was found between log gland weight as dependent variable and serum 25OHD as independent variable (r = -0.365; P < 0.0001). The only other influence on gland weight was a weak inverse correlation with age. Log gland weight as an independent variable was significantly related to adjusted calcium, PTH, and alkaline phosphatase (AP) as dependent variables. In 51 patients with serum 25OHD levels less than 15 ng/mL, gland weight, PTH, AP, and adjusted calcium were each significantly higher than in 97 patients with 25OHD levels of 15 ng/mL or more, but 1,25-dihydroxyvitamin D levels were similarly increased in both groups. In the former group the response of adjusted calcium to PTH was blunted, and the response of AP was enhanced, based on significant differences in regression slopes (P = 0.0004 and 0.0022, respectively). Suboptimal vitamin D nutrition stimulates parathyroid adenoma growth by a mechanism unrelated to hypocalcemia or 1,25-dihydroxyvitamin D deficiency and reduces the calcemic response to PTH, so that a higher PTH level and more parathyroid cells are needed to raise the patient's serum calcium to the level corresponding to the increased set-point that is characteristic of the disease. Improved vitamin D nutrition in the population is partly, perhaps largely, responsible for the historical changes in disease severity and manner of presentation that have occurred over the last 50 yr. PMID- 10720041 TI - Endocrine response to high-intensity exercise: dose-dependent effects of dexamethasone. AB - We recently reported that in 30-50% of healthy men and women the release of ACTH and cortisol stimulated by exercise is not suppressed by prior administration of a 4-mg dose of dexamethasone (DEX). We now explore other potential differences between these subjects and those whose exercise response was suppressed by examining the effect of a smaller, 1-mg, dose of DEX on exercise-stimulated ACTH and cortisol. Men (n = 15) and women (n = 9) were studied during three high intensity exercise tests: one after taking placebo, one after taking 1 mg DEX, and one after taking 4 mg DEX. Before participation, subjects underwent a test for classification as either a high (HR; n = 10) or low (LR; n = 14) reactor and a maximal exercise test to assess maximal aerobic capacity. Distinct dose-related reductions in plasma concentrations of ACTH, cortisol, and dehydroepiandrosterone (DHEA) were noted for HR under the treatment conditions, whereas both doses of DEX blocked ACTH, cortisol, and DHEA release in LR. Furthermore, basal plasma cortisol, DHEA, and DHEA sulfate were significantly higher in HR compared to LR. Thus, there are inherent basal and stress-reactive differences in HR and LR, and these differences may be useful in constructing a model for the mechanisms and physiological regulation of hypothalamic-pituitary-adrenal axis activation. The question of whether these differences in reactivity of the ACTH-cortisol axis between the HR and LR groups have implications for individual short term function or long term health remains to be answered. PMID- 10720042 TI - Diurnal rhythms of luteinizing hormone, follicle-stimulating hormone, testosterone, and estradiol secretion before the onset of female puberty in short children. AB - To investigate hormonal changes before the onset of female puberty, we measured LH and FSH in serum samples drawn every 20 min for 24 h and measured testosterone and estradiol hourly for 24 h. Seventeen girls (13 prepubertal and 4 early pubertal) of short stature, from 5.1-11.4 yr of age, participated in this study. LH and FSH were measured using a time-resolved immunofluorometric assay, and testosterone and estradiol were measured using a sensitivity RIA capable of detecting testosterone and estradiol concentrations of 10 and 2 pg/mL, respectively. Diurnal rhythms of LH, FSH, and testosterone were apparent in all subjects, including those aged 5-6 yr. Serum LH and FSH concentrations showed night-day variation in a pulsatile fashion. The serum testosterone concentration was elevated in the early morning in all subjects. The serum estradiol concentration was elevated in the early morning in 4 of 13 prepubertal subjects and all 4 early pubertal subjects. The diurnal pattern of the serum estradiol concentration was similar to that of the serum testosterone concentration. Mean 24-h LH and testosterone concentrations in prepubertal subjects who did not attain puberty for at least 1 yr were 0.07 U/L and 65 pg/mL, respectively, whereas those in prepubertal subjects who attained puberty within 1 yr (0.14 U/L and 106 pg/mL, respectively) were significantly higher. Furthermore, mean 24-h LH, FSH, testosterone, and estradiol concentrations increased with the onset of puberty. In conclusion, the diurnal rhythms of LH, FSH, and testosterone already exist at 5-6 yr of age, and serum LH and testosterone levels increase before the onset of puberty. These results suggest that preparation for the onset of female puberty may begin in 5- to 6-yr-old girls. PMID- 10720043 TI - Synergistic effects of nateglinide and meal administration on insulin secretion in patients with type 2 diabetes mellitus. AB - This study assessed the synergistic effects of nateglinide (a non-sulfonylurea D phenylalanine derivative) and meals on insulin secretion in 24 patients with type 2 diabetes. Oral doses of 60 and 180 mg or 120 and 240 mg were administered to two cohorts of subjects 10 min before meals (or fasting) three times daily for 7 days, with washout intervals between treatment periods. Dose-dependent increases in plasma insulin occurred, with the peak effect within 2 h after treatment. Significantly greater insulin secretion was observed when nateglinide was taken before a meal compared to nateglinide given in the fasted state or in response to just the meal. Nateglinide lowered plasma glucose concentrations significantly vs. placebo at all doses, and doses of 120 and 240 mg were more effective than 60 mg (P < 0.05). Adverse event rates were similar for nateglinide and placebo, and no hypoglycemic episodes or serious adverse events were reported during the study. Nateglinide (120 mg) was the maximum effective dose in this study and was shown to be a safe and well tolerated therapy for control of mealtime glucose excursions in patients with type 2 diabetes. Results indicate that a synergistic interaction occurs between nateglinide and elevated mealtime plasma glucose concentrations to stimulate insulin secretion. PMID- 10720044 TI - Energy expenditure, fat oxidation, and body weight regulation: a study of metabolic adaptation to long-term weight change. AB - Relatively low rates of energy expenditure and fat oxidation predict body weight gain. Weight gain, in turn, is associated with increases in energy expenditure and fat oxidation that may oppose further weight change. In response to experimental weight gain induced by overfeeding, increases in energy expenditure and fat oxidation are overcompensatory, i.e. greater than predicted for the change in body composition. To determine whether such metabolic adaptation occurs in response to spontaneous long term weight change, we conducted a longitudinal study in which 24-h energy expenditure (24-EE) and 24-h respiratory quotient (24 RQ; i.e. fat to carbohydrate oxidation) were repeatedly measured in 102 Pima Indians at baseline and after a mean follow-up of 3.6 +/- 2.7 yr, during which changes in body weight varied widely (-21 to +28 kg). We found that changes in 24 EE and 24-RQ in response to weight change were related to the amount of weight change, even after adjustment for body composition (partial r = 0.23 and -0.30, respectively; both P < 0.05). For a 15-kg weight gain, the increases in 24-EE (+244 Cal/day) and 24-h fat oxidation (+152 Cal/day) were 33 and 53 Cal/day greater than predicted from the cross-sectional relationship between both measures and body weight. Changes in 24-EE and 24-RQ varied substantially among individuals. Thus, on the average, spontaneous long term weight changes are accompanied by small metabolic adaptations in both energy expenditure and fat oxidation. The metabolic responses to weight changes are highly variable among individuals, however. PMID- 10720045 TI - Influence of puberty on muscle area and cortical bone area of the forearm in boys and girls. AB - The aim of the current study is to analyze the interaction of the muscle and bone system (muscle-bone unit) during puberty in males and females by computed tomography of the nondominant forearm. The data presented here are the first results from 318 healthy children (159 boys and 159 girls), aged 6-22 yr, and 336 adults (parents) participating in the DONALD Study (Dortmund Nutritional and Anthropometric Longitudinally Designed Study). Cortical area (CA) of the radius representing bone strength and muscle area (MA) representing muscle strength were measured with peripheral quantitative computed tomography (XCT 2000; Stratec, Pforzheim, Germany). A single slice measurement at a site corresponding to 65% of the ulnar length proximal to the radial endplate was used. MA and CA of the radius have been determined by a built-in software algorithm using density differences. There was a strong correlation between MA (x) and CA (y) in all children, adolescents, and adults (y = 0.019x + 10.93; r2 = 0.77). Before puberty, boys and girls displayed a similar relation between MA and CA. CA in relation to MA was greater in girls than in boys during puberty. Analysis of covariance was performed investigating the dependency of CA on MA, five pubertal stages, sex, and interaction of sex and pubertal stages. MA representing muscle strength was the strongest predictor of CA (P < 0.001) representing bone mass. Pubertal stage (P < 0.001) and interaction of pubertal stage*sex (P = 0.002) also had a significant influence on CA. r2 of the model was 0.85. These data suggest that in pubertal girls and women rather than in pubertal boys and men an additional factor shifts the relationship between MA and CA to higher values of cortical area. The present data confirm previous studies of the influence of puberty and estrogens or related factors on the muscle-bone interaction. PMID- 10720046 TI - Changes in parameters of bone and mineral metabolism during therapy for hyperthyroidism. AB - Hyperthyroid patients have high bone turnover and negative calcium and phosphorus balance often associated with mild osteopenia. Early during antithyroid treatment bone turnover decreases, the mineral balance is converted to positive, and sometimes hypocalcemia occurs. The aim of this investigation was to study the mechanisms of the changes in some parameters of bone and mineral metabolism after treatment of thyrotoxicosis. Thirteen newly diagnosed patients with Graves' disease (seven postmenopausal women, four premenopausal women, and two men) were studied longitudinally, every 6 weeks, for 1 yr after commencing antithyroid treatment with methimazole. Mean serum calcium and phosphorus were both slightly above the normal mean at week 0 and decreased significantly (by 10% and 24%, respectively) during treatment. Fasting urinary calcium was 236 +/- 4 (mean +/- SEM) mg/g creatinine, and the fractional excretion of Ca was 2.0 +/- 0.33% before treatment; both fell significantly to minimums of 61 +/- 20 mg/g and 0.6 +/- 0.16%, respectively. Urinary phosphorus was 282 +/- 60 mg/g creatinine, and the fractional excretion of phosphorus was 3.3 +/- 0.6% before treatment; both increased significantly to 452 +/- 40 mg/g and 8.4 +/- 1.0%, respectively, during treatment. The z-scores were calculated from the mean and SD ofthe respective control groups. The z-score of urinary N-telopeptides of type I collagen (U.NTx) was 9.3 +/- 1.3 at week 0 and declined exponentially, but failed to normalize after 1 yr of antithyroid treatment. The serum alkaline phosphatase (ALP) z-score was initially 2.2 +/- 0.2, increased to 6.0 +/- 1.0 at week 6, and declined slowly there after to 1.0 +/- 1.1 at week 54. The serum osteocalcin (OC) z-score showed a temporal pattern similar to that of ALP. It was initially 2.2 +/- 0.2, increased to 4.0 +/- 0.6 at week 6, and later declined slowly to 0.7 +/- 0.5 at week 54. The failure of the markers of bone turnover to normalize after 1 yr of therapy indicates an on-going high rate of bone turnover despite the attained euthyroidism. The uncoupling index (UI = z-score of U.NTx minus z-score of OC) was 7.1 +/- 1.2 before treatment, indicating unbalanced bone turnover in favor of bone resorption, and fell close to zero at week 30 of treatment. Pretreatment plasma PTH was suppressed slightly to 2.17 +/- 0.47 pmol/L and rose significantly during treatment, reaching a plateau of 5.27 +/- 0.78 at week 12. In all postmenopausal women PTH increased above the upper limit of normal (6.84 pmol/L). Pretreatment serum 25-hydroxyvitamin D was normal and remained unchanged during treatment, whereas 1,25-dihydroxyvitamin D was initially subnormal and rose to normal level after treatment. There was a significant positive linear correlation between PTH and U.NTx after week 12. PTH was also significantly correlated with ALP, but not with OC. ALP and OC were significantly correlated. A significant positive correlation was found between T3 and U.NTx, and a negative correlation was found between T3 and each of the formation markers (ALP and OC) over the 0- to 12-week interval. The latter correlations and the very high pretreatment UI indicate some inhibitory effect of the high thyroid hormone levels on the osteoblasts. The marked and sustained elevation of PTH, more pronounced in the postmenopausal women, during the first year of treatment of hyperthyroidism seems to play a pivotal role in maintaining a relatively high rate of bone turnover despite euthyroidism, and in the conservation of calcium by reducing renal calcium excretion and increasing calcium absorption (via 1,25-dihydroxyvitamin D). It may also account in part for the additional rise of the bone formation markers by an anabolic effect on the osteoblasts. Endogenous PTH may be important in the restoration of bone mineral density of treated hyperthyroid patients. PMID- 10720047 TI - Prognostic value of [18F]fluorodeoxyglucose positron emission tomographic scanning in patients with thyroid cancer. AB - Poorly differentiated thyroid cancer lesions often lose the ability to concentrate radioactive [131I]iodine (RAI) and exhibit increased metabolic activity, as evidenced by enhanced glucose uptake. We incorporated [18F]fluorodeoxyglucose (FDG) positron emission tomography (PET) scanning into the routine follow-up of a cohort of thyroid cancer patients undergoing annual evaluations. One hundred and twenty-five patients who had previous thyroidectomies were included. They had diagnostic RAI whole body scans, serum thyroglobulin measurements, and additional imaging studies as clinically indicated. During 41 months of follow-up, 14 patients died. Univariate analysis demonstrated that survival was reduced in those with age over 45 yr, distant metastases, PET positivity, high rates of FDG uptake, and high volume of the FDG avid disease (>125 mL). Survival did not correlate with gender, RAI uptake, initial histology, or grade. Multivariate analysis demonstrated that the single strongest predictor of survival was the volume of FDG-avid disease. The 3-yr survival probability of patients with FDG volumes of 125 mL or less was 0.96 (95% confidence interval, 0.91, 1.0) compared with 0.18 (95% confidence interval, 0.04, 0.85) in patients with FDG volume greater than 125 mL. Only 1 death (of leukemia) occurred in the PET-negative group (n = 66). Of the 10 patients with distant metastases and negative PET scans, all were alive and well. Patients over 45 yr with distant metastases that concentrate FDG are at the highest risk. Once distant metastases are discovered in patients with differentiated thyroid carcinoma, FDG-PET can identify high and low risk subsets. Subjects with a FDG volume greater than 125 mL have significantly reduced short term survival. PMID- 10720048 TI - Flutamide, testolactone, and reduced hydrocortisone dose maintain normal growth velocity and bone maturation despite elevated androgen levels in children with congenital adrenal hyperplasia. AB - Treatment outcome in congenital adrenal hyperplasia is often sub-optimal due to hyperandrogenism, treatment-induced hypercortisolism, or both. We previously reported better control of linear growth, weight gain, and bone maturation in a short term cross-over study of a new four-drug treatment regimen containing an antiandrogen (flutamide), an inhibitor of androgen to estrogen conversion (testolactone), reduced hydrocortisone dose, and fludrocortisone, compared to the effects of a control regimen of hydrocortisone and fludrocortisone. Twenty-eight children have completed 2 yr of follow-up in a subsequent long term randomized parallel study comparing these two treatment regimens. During 2 yr of therapy, compared to children receiving hydrocortisone, and fludrocortisone treatment, children receiving flutamide, testolactone, reduced hydrocortisone dose (average of 8.7 +/- 0.6 mg/m2 x day), and fludrocortisone had significantly (P < or = 0.05) higher plasma 17-hydroxyprogesterone, androstenedione, dehydroepiandrosterone, dehydroepiandrosterone sulfate, and testosterone levels. Despite elevated androgen levels, children receiving the new treatment regimen had normal linear growth rate (at 2 yr, 0.1 +/- 0.5 SD units), and bone maturation (at 2 yr, 0.7 +/- 0.3 yr bone age/yr chronological age). No significant adverse effects were observed after 2 yr. We conclude that the regimen of flutamide, testolactone, reduced hydrocortisone dose, and fludrocortisone provides effective control of congenital adrenal hyperplasia with reduced risk of glucocorticoid excess. A long term study of this new regimen is ongoing. PMID- 10720049 TI - Gender difference in insulin-like growth factor I response to growth hormone (GH) treatment in GH-deficient adults: role of sex hormone replacement. AB - GH production in healthy women is about thrice that in men. Yet insulin-like growth factor I (IGF-I) levels are similar, suggesting a lower responsivity to GH in women. In untreated GH-deficient adults, basal IGF-I levels are reportedly lower in females than in males, and the therapeutic recombinant human GH (rhGH) dose required to achieve optimal IGF-I levels is higher in the former, suggesting a pivotal role of estrogens on rhGH requirement in GH-deficient patients. We, therefore, analyzed our 2-yr data on the effect of rhGH on serum IGF-I in 77 GH deficient patients (33 men, mean +/- SD age, 37.2 +/- 13.8 yr; 44 women, mean +/- SD age, 36.9 +/- 11.9 yr) with due attention to gender differences and to the effects of sex hormone replacement. Of the 44 women, 33 had estrogen substitution. Of the 33 men, 23 were on androgen replacement. Patients (11 premenopausal women and 10 men) not on hormonal replacement were eugonadal. Basal IGF-I levels in untreated GH-deficient women were significantly lower than in men (8.8 +/- 0.7 nmol/L vs. 12.2 +/- 0.9 nmol/L; P < 0.01), despite similar basal GH levels. The daily rhGH dose per kg body weight required to normalize IGF-I in women was higher than in men, the difference being statistically significant at all time points (P < 0.05-0.01). The IGF-I increase (delta) per IU GH/day x kg over the 24-month period was about twice higher in men than in women. Also calculated on a weight basis, rhGH responsivity (rhGH responsivity = (deltaIGF1(nmol/L)/dose (IU/day/kg)) was higher in men than in women at all time intervals (P < 0.05-0.01). Estrogen replacement in women significantly increased rhGH requirement. The rhGH dose per kg body weight required in estrogen substituted women was significantly higher than in nonestrogen-substituted women (P < 0.01 at t = 18 and 24 months, respectively). In women on estrogen substitution, rhGH responsivity plateaued from 6 months on, whereas in eugonadal women without estrogen substitution the responsivity for rhGH increased over time. In men, the reverse was true; rhGH responsivity increased over time in men on androgen substitution, but plateaued in men without androgen substitution. The mechanisms underlying this gender difference are not known. Differential influences of estrogens and androgens on the expression of the GH receptor gene and IGF-I messenger RNA may be operative. The present study confirms short-term data published in the literature on a sex difference in rhGH dose requirement in GH-deficient patients. It furthers extends the data by demonstrating that this sex difference in GH responsivity persists and changes during the 24 months of the study. Moreover, it shows that estrogen replacement blunts the IGF-I response to rhGH in women, whereas in men with androgen substitution the responsivity increases over time, thus bearing a risk of undertreatment in women and overtreatment in men. PMID- 10720050 TI - Disease-associated autoantibodies as surrogate markers of type 1 diabetes in young children at increased genetic risk. Childhood Diabetes in Finland Study Group. AB - To evaluate the emergence of diabetes-associated autoantibodies in young children and to assess whether such antibodies can be used as surrogate markers of type 1 diabetes in young subjects at increased genetic risk, we studied 180 initially unaffected siblings (92 boys and 88 girls) of children with newly diagnosed type 1 diabetes. All siblings were younger than 6 yr of age at the initial sampling, and they were monitored for the emergence of islet cell antibodies (ICA), insulin autoantibodies (IAA), glutamate decarboxylase antibodies (GADA), and IA-2 antibodies (IA-2A) up to the age of 6 yr and for progression to clinical type 1 diabetes up to the age of 10 yr. All 160 siblings with DNA samples available were typed for susceptible (DQB1*02 and *0302) and protective (DQB1*0301 and *0602-03) HLA DQB1 alleles. Twenty-two siblings (12.2%) tested positive for ICA in their first antibody-positive sample before the age of 6 yr, 13 (7.2%) tested positive for IAA, 15 (8.3%) tested positive for GADA, and 14 (7.8%) tested positive for IA 2A. There were 16 siblings (8.9%) who had 1 detectable autoantibody, 5 (2.8%) had 2, and 12 (6.7%) had 3 or more. In the group of 82 siblings with increased human leukocyte antigen-defined genetic susceptibility [DQB1*02/*0302, *0302/x (x = other than *02 or a protective allele), *02/y (y = other than *0302 or a protective allele)], 18 (22.0%) tested positive for ICA in their first antibody positive sample, 10 (12.2%) tested positive for IAA, 14 (17.1%) tested positive for GADA, and 12 (14.6%) tested positive for IA-2A. One antibody was detectable in 6 siblings (7.3%), 2 were detectable in 5 (6.1%), and 3 or more were detectable in 12 (14.6%). Fifteen siblings (18.3%) presented with clinical type 1 diabetes before the age of 10 yr. All of the progressors showed increased human leukocyte antigen-defined genetic susceptibility. Thirteen of those 15 siblings, who presented with clinical type 1 diabetes before the age of 10 yr, had at least 2 antibodies detectable before the age of 6 yr (disease sensitivity, 87%; 95% confidence interval, 60-98%). Thirteen of the 17 siblings who tested positive for 2 or more autoantibodies before the age of 6 yr developed type 1 diabetes before the age of 10 yr (positive predictive value, 76%; 95% confidence interval, 50 93%). These observations suggest that disease-associated autoantibodies can well be used as surrogate markers of clinical type 1 diabetes in primary prevention trials targeting young subjects with increased genetic disease susceptibility. PMID- 10720051 TI - Increased diurnal plasma concentrations of cortisone in depressed patients. AB - The enzyme 11-beta-hydroxysteroid dehydrogenase (11-beta-HSD) regulates glucocorticoid activity by converting cortisol into cortisone and vice versa. Frequent signs of major depression are elevated concentrations of circulating cortisol and ACTH. However, no information is available about the activity of 11 beta-HSD in this disorder. Therefore, we compared diurnal plasma concentrations of cortisol and cortisone and their ratios, reflecting 11-beta-HSD activity, in 25 severely depressed patients (Hamilton Depression Scale, 29 +/- 6; 14 men, 11 women, age 22-77 yr; mean, 47 +/- 16) and 30 control persons (20 men, 10 women age 23-85 yr; mean, 51 +/- 19). Cortisol and cortisone were measured at 0900 h, 1100 h, 1300 h, 2000 h, 2200 h, 0100 h, 0300 h, and 0700 h with specific RIAs after extraction. Both cortisol and cortisone concentrations were significantly increased in patients compared with controls (cortisol, 251.7 +/- 113.1 vs. 160 +/- 96.6 nmol/L; cortisone, 32.8 +/- 10.9 vs. 21.9 +/- 10.9 nmol/L). The calculated ratios of cortisol to cortisone were similar in controls and patients. Similar to cortisol, the circadian variation of cortisone was flattened in patients with the ratio of maximal cortisone to minimal cortisone being 1.9-fold higher in controls than in patients. There was no gender-specific difference in cortisone values neither in patients nor in controls. We conclude that in major depression increased cortisol is not due, at least partly, to an altered 11-beta HSD activity or to a decrease in cortisone. PMID- 10720052 TI - HMG-CoA reductase inhibitors increase BMD in type 2 diabetes mellitus patients. AB - Recently, it was reported that 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors increased bone mineral density (BMD) in mice. We studied the effect of HMG-CoA reductase inhibitors on BMD of type 2 diabetes mellitus by a retrospective review of medical records. Sixty-nine type 2 diabetic patients were included. The control group (n = 33) did not take HMG-CoA reductase inhibitors. The treatment group (n = 36) was administered either lovastatin, pravastatin, or simvastatin. BMD of the spine, femoral neck, femoral trochanter, and total hip were measured by dual-energy X-ray absorptiometry. There were no significant differences between control and treatment groups in age, sex, body mass index, glycemic control, and serum insulin levels. In the control group, BMD of the spine significantly decreased (from 1.116 +/- 0.165 to 1.081 +/- 0.178 g/cm2) after 14 months. In the treatment group, BMD of the femoral neck significantly increased (from 0.853 +/- 0.139 to 0.878 +/- 0.147 g/cm2) after 15 months. In male subjects treated with HMG-CoA reductase inhibitors, there was a significant increase in BMD of the femoral neck and femoral trochanter (from 0.899 +/- 0.139 to 0.934 +/- 0.139 and from 0.801 +/- 0.145 to 0.833 +/- 0.167 g/cm2, respectively), but in female subjects, only BMD of the femoral neck increased (from 0.819 +/- 0.132 to 0.834 +/- 0.143 g/cm2). Percentage increments of BMD of the femoral neck, femoral wards triangle, femoral trochanter, and total hip in the treatment group were significantly higher than in the control group (2.32% vs. -0.99, 1.77% vs. -1.25%, 1.40% vs. -1.21%, 0.88% vs. -1.03%, respectively). The proportion of subjects who had an increase in BMD of the spine and total hip more than two percentages was significantly larger in the treatment group than in the control group (30.6% vs. 15.2% and 30.6% vs. 9.1%, respectively). The increased increment in BMD of the treatment group was significantly greater than those in the control group after adjustment for age and body mass index (P < 0.05). These results suggest that HMG-CoA reductase inhibitors may increase BMD of the femur in male patients with type 2 diabetes mellitus. PMID- 10720053 TI - Placental growth hormone (GH), GH-binding protein, and insulin-like growth factor axis in normal, growth-retarded, and diabetic pregnancies: correlations with fetal growth. AB - We previously described significant changes in GH-binding protein (GHBP) in pathological human pregnancy. There was a substantial elevation of GHBP in cases ofnoninsulin-dependent diabetes mellitus and a reduction in insulin-dependent diabetes mellitus. GHBP has the potential to modulate the proportion of free placental GH (PGH) and hence the impact on the maternal GH/insulin-like growth factor I (IGF-I) axis, fetal growth, and maternal glycemic status. The present study was undertaken to investigate the relationship among glycemia, GHBP, and PGH during pregnancy and to assess the impact of GHBP on the concentration of free PGH. We have extended the analysis of specimens to include measurements of GHBP, PGH, IGF-I, IGF-II, IGF-binding protein-1 (IGFBP-1), IGFBP-2, and IGFBP-3 and have related these to maternal characteristics, fetal growth, and glycemia. The simultaneous measurement of GHBP and PGH has for the first time allowed calculation of the free component of PGH and correlation of the free component to indexes of fetal growth and other endocrine markers. PGH, free PGH, IGF-I, and IGF-II were substantially decreased in IUGR at 28-30 weeks gestation (K28) and 36 38 weeks gestation (K36). The mean concentration (+/-SEM) of total PGH increased significantly from K28 to K36 (30.0 +/- 2.2 to 50.7 +/- 6.2 ng/mL; n = 40), as did the concentration of free PGH (23.4 +/- 2.3 to 43.7 +/- 6.0 ng/mL; n = 38). The mean percentage of free PGH was significantly less in IUGR than in normal subjects (67% vs. 79%; P < 0.01). Macrosomia was associated with an increase in these parameters that did not reach statistical significance. Multiple regression analysis revealed that PGH/IGF-I and IGFBP-3 account for 40% of the variance in birth weight. IGFBP-3 showed a significant correlation with IGF-I, IGF-II, and free and total PGH at K28 and K36. Noninsulin-dependent diabetes mellitus patients had a lower mean percentage of free PGH (65%; P < 0.01), and insulin dependent diabetics had a higher mean percentage of free PGH (87%; P < 0.01) than normal subjects. Mean postprandial glucose at K28 correlated positively with PGH and free PGH (consistent with the hyperglycemic action of GH). GHBP correlated negatively with both postprandial and fasting glucose. Although GHBP correlated negatively with PGH (r = -0.52; P < .001), free PGH and total PGH correlated very closely (r = 0.98). The results are consistent with an inhibitory function for GHBP in vivo and support a critical role for placental GH and IGF-I in driving normal fetal growth. PMID- 10720054 TI - Sleep apnea and daytime sleepiness and fatigue: relation to visceral obesity, insulin resistance, and hypercytokinemia. AB - Sleep apnea and associated daytime sleepiness and fatigue are common manifestations of mainly obese middle-aged men. The onset of sleep apnea peaks in middle age, and its morbid and mortal sequelae include complications from accidents and cardiovascular events. The pathophysiology of sleep apnea remains obscure. The purpose of this study was to test three separate, albeit closely related, hypotheses. 1) Does sleep apnea contribute to the previously reported changes of plasma cytokine (tumor necrosis factor-alpha and interleukin-6) and leptin levels independently of obesity? 2) Among obese patients, is it generalized or visceral obesity that predisposes to sleep apnea? 3) Is apnea a factor independent from obesity in the development of insulin resistance? Obese middle-aged men with sleep apnea were first compared with nonapneic age- and body mass index (BMI)-matched obese and age-matched lean men. All subjects were monitored in the sleep laboratory for 4 consecutive nights. We obtained simultaneous indexes of sleep, sleep stages, and sleep apnea, including apnea/hypopnea index and percent minimum oxygen saturation. The sleep apneic men had higher plasma concentrations of the adipose tissue-derived hormone, leptin, and of the inflammatory, fatigue-causing, and insulin resistance-producing cytokines tumor necrosis factor-alpha and interleukin-6 than nonapneic obese men, who had intermediate values, or lean men, who had the lowest values. Because these findings suggested that sleep apneics might have a higher degree of insulin resistance than the BMI-matched controls, we studied groups of sleep-apneic obese and age- and BMI-matched nonapneic controls in whom we obtained computed tomographic scan measures of total, sc, and visceral abdominal fat, and additional biochemical indexes of insulin resistance, including fasting plasma glucose and insulin. The sleep apnea patients had a significantly greater amount of visceral fat compared to obese controls (<0.05) and indexes of sleep disordered breathing were positively correlated with visceral fat, but not with BMI or total or sc fat. Furthermore, the biochemical data confirmed a higher degree of insulin resistance in the group of apneics than in BMI-matched nonapneic controls. We conclude that there is a strong independent association among sleep apnea, visceral obesity, insulin resistance and hypercytokinemia, which may contribute to the pathological manifestations and somatic sequelae of this condition. PMID- 10720055 TI - Angiogenesis in pituitary adenomas and the normal pituitary gland. AB - Angiogenesis is essential for tumor growth beyond a few millimeters in diameter, and the intratumoral microvessel count that represents a measure of angiogenesis has been correlated with tumor behavior in a variety of different tumor types. To date no systematic study has assessed pituitary tumors of different secretory types, correlating vascular count with tumor size. The vascular densities of pituitary tumors and normal anterior pituitary were therefore assessed by counting vessels labeled using the vascular markers CD31 and ulex europaeus agglutinin I. One hundred and twelve surgically removed pituitary adenomas (30 GH secreting, 25 prolactinomas, 15 ACTH-secreting, and 42 nonfunctioning tumors) were compared with 13 specimens of normal anterior pituitary gland. The vascular counts in the normal anterior pituitary gland were significantly higher (P < 0.05) than those in the tumors using both CD31 and ulex europaeus agglutinin I. In addition, microprolactinomas were significantly less vascular (P < 0.05) than macroprolactinomas, although there was no such difference between vascular densities of microadenomas and macroadenomas producing GH. ACTH-secreting tumors were, like microprolactinomas, of much lower vascular density than the normal pituitary and other secreting and nonsecreting tumor types. In marked contrast to other tumors, pituitary adenomas are less vascular than the normal pituitary gland, suggesting that there may be inhibitors of angiogenesis that play an important role in the behavior of these tumors. PMID- 10720056 TI - Proteolysis of insulin-like growth factor-binding protein-3 is increased in urine from patients with diabetic nephropathy. AB - The insulin-like growth factor (IGF) system has been implicated in the development of experimental diabetic nephropathy. IGF-binding protein-3 (IGFBP-3) modulates IGF actions, and proteolysis decreases its binding affinity for IGFs. The aim of this study was to explore the possibility that proteolysis of IGFBP-3 may be altered in diabetic nephropathy and may therefore modify the intrarenal effects of IGFs. IGFBP-3 proteolysis in urine from diabetic patients with normo- [albumin excretion rate (AER), <20 microg/min], micro- (AER, 20-200 microg/min), and macroalbuminuria (AER, >200 microg/min) was studied in 34 patients with noninsulin-dependent diabetes mellitus (NIDDM), 14 patients with insulin dependent diabetes mellitus, and 9 controls. Urine samples were analyzed by Western ligand blotting and IGFBP-3 immunoblotting. Protease activity was quantitated using [125I]IGFBP-3 as a substrate. WLB showed three main bands (40 46, 35, and 26 kDa) in control urine and a fainter 18-kDa band. All but the 35 kDa band were immunoreactive with the IGFBP-3 antiserum. The same pattern of IGFBPs was seen in urine from normoalbuminuric diabetic patients. However, the urine of diabetic patients with micro- and macroalbuminuria contained little or no intact 40- to 46-kDa IGFBP-3. In patients with noninsulin-dependent diabetes mellitus, urinary IGFBP-3 protease activity in micro- (n = 13) and macroalbuminuric patients (n = 12; mean +/- SD[SCAP], 75 +/- 25% and 84 +/- 24%) was significantly higher than that in normoalbuminuric patients (29 +/- 9%; P = 0.0001). Similar results were observed in patients with insulin-dependent diabetes mellitus. Proteolytic activity in diabetic urine was due to a serine protease. In conclusion, diabetic nephropathy was associated with IGFBP-3 proteolysis in urine. As similar changes were not observed in patients' sera, this is likely to reflect changes in the kidney or urinary tract, resulting in increased local IGF bioavailability, and therefore may contribute to the structural changes of diabetic nephropathy. PMID- 10720057 TI - The ret/PTC mutations are common in sporadic papillary thyroid carcinoma of children and young adults. AB - The ret/PTC rearrangements (PTC-1, PTC-2, and PTC-3) are characteristic of papillary thyroid cancer (PTC). In adults, PTC-1 is common and may be associated with an aggressive clinical course. The incidence and significance of ret/PTC mutations are less well understood in children. We examined spontaneous PTC from 33 patients (23 females and 10 males) with a median age of 18 yr (range, 6-21 yr) and a median follow-up of 3.5 yr (range, 0-13.4 yr). The ret/PTC mutations were identified in 15 tumors (45%), including 8 PTC-1 (8 of 15, 53%), 2 PTC-2 (2 of 15, 13%), 2 PTC-3 (2 of 15, 13%), and 3 (3 of 15, 20%) combined PTC mutations (PTC-1 and PTC-2). This distribution is significantly different (P = 0.001, by chi2 analysis) from that reported for children with radiation-induced PTC. There was no correlation between the presence or type of ret/PTC mutation and patient age, tumor size, focality, extent of disease at diagnosis, or recurrence. We conclude that ret/PTC mutations are 1) common in sporadic childhood PTC, 2) predominantly PTC-1, 3) frequently multiple, and 4) of different distribution than that reported for children with radiation-induced PTC. PMID- 10720058 TI - Binding of human thyrotropin receptor peptides to a Graves' disease-predisposing human leukocyte antigen class II molecule. AB - There are many reports that Graves' disease (GD) is associated with certain human leukocyte antigen (HLA) molecules, in particular DR3. Here we examined the characteristics of binding of human TSH receptor (TSHR) peptides to this disease associated HLA class II molecule. DR3 molecules bind TSHR immunodominant peptide epitopes with intermediate affinity. On the contrary, DR3 binds nonimmunogenic peptides either with poor affinity or not at all, with one exceptional peptide that has extremely high affinity. These results suggest that susceptibility to GD associated with inheritance of a specific HLA class II gene is due to the influence of the HLA molecule-TSHR peptide complex on the T cell repertoire. PMID- 10720059 TI - A human pituitary tumor-derived folliculostellate cell line. AB - Pituitary cells have been used for the study of hormone synthesis, secretion, and regulation. However, the lack of human cell lines of pituitary origin has made such studies in humans very difficult. Activin, a member of the transforming growth factor-beta cytokine family, is secreted by the pituitary and serves, in addition to regulating hormone biosynthesis, as a regulator of cell growth and differentiation. In the human pituitary, folliculo-stellate cells secrete an activin-binding and -neutralizing protein, follistatin. However, the role of these cells in the autocrine/paracrine regulatory mechanisms of activin is poorly understood. We describe a human pituitary-derived folliculostellate cell line, designated PDFS, that was developed spontaneously from a clinically nonfunctioning pituitary macroadenoma. PDFS cells showed an epithelial-like morphology with long cytoplasmic processes. Electron microscopy revealed frequent intercellular junctions, including desmosomes, and cytogenetic analysis showed clonal characteristics with chromosomal abnormalities. These cells express vimentin and the nervous tissue-specific S-100 protein, specific markers of folliculostellate cells in the anterior pituitary, but no secretory pituitary cell markers. PDFS cells formed large colonies in an anchorage-independent transformation assay. They express follistatin and activin A and have an intact activin intracellular signaling pathway as determined by reporter assays. Therefore, this human cell line provides a useful model for studying the regulation of cell growth and cytokine production by factors endogenously produced in pituitary folliculostellate cells. PMID- 10720060 TI - Serum withdrawal-induced apoptosis in thyroid cells is caused by loss of fibronectin-integrin interaction. AB - In some cell types, including a fetal thyroid cell line, denial of adhesion to extracellular matrix induces a type of apoptosis called anoikis. Serum withdrawal in dog and transformed rat thyroid cells also induces programmed cell death. Because serum can stimulate cells to produce some components of the extracellular matrix, it was of interest to determine the role of the matrix in the apoptosis induced by serum withdrawal in normal human thyroid cells in primary culture. The present report demonstrates that thyroid cells selectively produce and deposit insoluble fibronectin (FN) only when stimulated by serum. Adhesion in the presence of serum is dependent upon integrin-FN interaction. Serum withdrawal determines a degradation of the insoluble FN deposited and a detachment of the cells from the plates. In these conditions, cells undergo anoikis, demonstrated by DNA fragmentation and annexin V staining. Apoptosis was prevented by exogenous FN immobilized onto the plates. These results indicate that serum withdrawal induces apoptosis in human thyroid cells, determining FN degradation and loss of cell-matrix adhesion. PMID- 10720061 TI - Cytokine profiles in eye muscle tissue and orbital fat tissue from patients with thyroid-associated ophthalmopathy. AB - Eye muscle (EM) and retroorbital fat tissue are two major sites of involvement in thyroid-associated ophthalmopathy (TAO). Lymphocytic infiltration in these tissues is a prominent histological feature of TAO. We have investigated the cytokine gene profiles in EM and orbital fat (OF) tissues from patients with TAO. Total RNA was isolated from EM tissue of 14 patients and from OF tissues of 29 patients with TAO. Cytokine gene expression was assessed by RT-PCR using paired primers for interferon gamma (IFNgamma), tumor necrosis factor alpha (TNFalpha), interleukin (IL)-1beta, IL-2, IL-4, IL-6, IL-10, CD4, CD8, and glyceraldehyde-3 phosphate dehydrogenase. IFNgamma, TNFalpha, IL-1beta, and IL-6 messenger RNA (mRNA) were mainly detected in EM tissue, whereas IL-4 and IL-10 mRNA were detected in only one patient. On the other hand, in OF tissue, IL-4 and IL-10 mRNA were detected in 24% and 38% of the patients, respectively, and IFNgamma, IL 1beta, and IL-6 mRNA were less often detected compared with EM tissue. The enlargement of EM tissue as assessed by computed tomography correlated significantly with TNFalpha mRNA expression in EM tissue. The orbital volume was positively correlated with IL-6 mRNA expression and negatively correlated with IL 4 mRNA and IL-10 mRNA expression in OF tissue. These results suggest that T helper (Th) 1-like cytokines predominate in EM tissue in most patients and that the predominant cytokine profile in OF tissue varies from patient to patient. Both Th1-like and Th2-like immune responses may play roles in the development of two components of ophthalmopathy. PMID- 10720062 TI - Estrogen receptor isoform gene expression in ovarian stromal and epithelial tumors. AB - The factors involved in the pathogenesis of ovarian cancers remain unclear, and the response of these tumors to hormonal therapy is limited. The identification of a second estrogen receptor gene (ERbeta), expressed predominantly in ovarian granulosa cells, led us to explore its possible role in ovarian cancer, particularly in granulosa cell tumors (GCT). Several isoforms of ERbeta have been identified. We sought to define the patterns of both ERalpha and ERbeta gene expression in a panel of ovarian tumors consisting of GCT and serous and mucinous cystadenocarcinomas as well as in normal ovary. Expression was determined by RT PCR using gene- and isoform-specific primers and probes combined with Southern blot analysis of the PCR products. Widespread expression of ERalpha was observed in all tumor types, but at relatively low levels. ERbeta is expressed predominantly in GCT, with lower levels in mucinous tumors and very low levels in serous tumors. The ERbeta2 splice variant previously reported in rodents was not observed. Only very low levels of the exon 5, exon 6, and exon 5/6 deletion variants were detected. The C-terminal truncation variant ERbeta(cx), however, exhibited widespread expression across all the tumor types. As ERbeta(cx) has been shown to be a ligand-independent antagonist of ERalpha action, the relative ratios of ERbeta(cx), ERalpha, and ERbeta may influence the response of a tumor to antiestrogen therapy. PMID- 10720063 TI - Timing of prenatal androgen excess determines differential impairment in insulin secretion and action in adult female rhesus monkeys. AB - This study determined whether timing of prenatal androgen excess resulted in differential impairment of insulin-glucose homeostasis in adult female rhesus monkeys. Ten female rhesus monkeys exposed to testosterone propionate starting on gestational day 40 (early treated), 9 females exposed to testosterone propionate starting between gestational days 100-115 (late treated), and 15 control females were studied. The modified minimal model was used to examine various measures derived from an i.v. glucose tolerance test, with regression analysis performed between these variables and body mass index. In addition, the disposition index (DI) and the hyperbolic relationship between insulin sensitivity (S(I)) and acute insulin response to glucose were examined. Early treated females demonstrated impaired pancreatic beta-cell function, as shown by diminished DI and decreased percentile ranking for the hyperbolic relationship between S(I) and acute insulin response to glucose. In contrast, late treated females exhibited both an increase in DI and a negative relationship between body mass index and S(I). These results suggest that prenatal androgen excess in female rhesus monkeys, regardless of gestational timing, perturbs insulin-glucose homeodynamics, with androgen excess in early and late gestation impairing pancreatic beta-cell function and altering insulin sensitivity, respectively. PMID- 10720064 TI - Origin of an ovarian steroid cell tumor causing isosexual pseudoprecocious puberty demonstrated by the expression of adrenal steroidogenic enzymes and adrenocorticotropin receptor. AB - Ovarian steroid cell tumors are rare neoplasms composed of typical steroid hormone-secreting cells. Most ovarian steroid cell tumors, however, cannot be appropriately classified on a morphological basis, because the neoplastic cells closely resemble adrenal cortical cells. Nevertheless, the true adrenal origin of such tumors has been difficult to demonstrate. Here we report a 3-yr-old girl with isosexual pseudoprecocious puberty due to an ovarian steroid tumor whose adrenal cell origin was determined by the presence of messenger ribonucleic acid (mRNA) of adrenal-specific steroidogenic P450 enzymes (P450c11 and P450c21) and ACTH receptor (ACTHR). Her height was +2.3 SD, and she had Tanner stage III breast development, Tanner stage II pubic hair, and a normal clitoris. Bone age was 5 yr. Basal gonadotropin levels were undetectable (<0.6 U/L for LH and <1.0 U/L for FSH) and remained undetectable after stimulation with 100 microg GnRH, i.v. Basal serum testosterone and 17-hydroxyprogesterone levels were slightly elevated, whereas basal serum androstenedione, estradiol, and dehydroepiandrosterone sulfate levels were clearly elevated. Pelvic ultrasound disclosed an enlarged uterus and an adnexal multicystic mass in the right ovary, and pathological studies disclosed an ovarian steroid cell tumor. To establish the cellular origin of the tumor we determined the presence of mRNA for P450c11, P450c21, and ACTHR in tumor tissue and normal adrenal and ovarian tissue. Detection of ACTHR, P450c21, and P450c11 mRNAs isoforms was achieved in tumoral and adrenal control tissue, but not in the ovary control tissue. The RT-PCR products of P450c11 from adrenal control tissue were composed by both BglI sensitive and -resistant complementary DNAs, indicating the presence of both P450c11AS and P450c11beta, whereas RT-PCR product from the tumor was resistant to BglI digestion, indicating only the presence of P450c11beta. We conclude that the histological origin of so-called adrenal rest tumor could be reliably determined by assessing the expression of specific genes in the tumor as P450c11beta and P450c21. The use ofthese molecular tools will allow a more precise classification of an important subset of the ovarian steroid cell tumors and can help to identify ectopic adrenal tissue in ovary and testis. PMID- 10720065 TI - Close correlation between estrogen treatment and renal phosphate reabsorption capacity. AB - To determine the influence of estrogen on the activity of renal proximal tubular reabsorption of inorganic phosphate (Pi) in women, we examined the changes of the renal threshold phosphate concentration (also denoted as TmP/GFR), as well as the changes in the concentrations of mineral components in the circulation in two groups of women--one receiving hormone replacement therapy (HRT) and one receiving gonadotropin-releasing hormone agonists (GnRH-a) therapy. We also examined the changes in the concentrations of serum PTH in the GnRH-a group. The patients in the HRT group were continuously treated with 0.625 mg conjugated equine estrogens plus 2.5 mg medroxyprogesterone acetate per day. The patients in the GnRH-a group were treated with a monthly injection of 3.75 mg leuprolide acetate depot for 6 months. The values of TmP/GFR decreased in all of the patients who received HRT. The mean percentage change in TmP/GFR was -14.5% (range, -24.3% to -9.6%). In contrast, in all of the patients treated with GnRH a, the values of TmP/GFR increased after 6 months of treatment (the mean percentage change was 28.5%; range, 18.2-78.3%) and returned to the preadministration level at 12 weeks after stopping therapy. In these patients, both the values of TmP/GFR and the concentrations of serum Pi correlated significantly with circulating estradiol levels (r = -0.767, P < 0.01 and r = 0.797, P < 0.01, respectively), but the concentrations of serum corrected calcium did not correlate. Moreover, in the same patients, the levels of serum intact PTH decreased significantly (P < 0.05) after 6 months of treatment, but at 12 weeks after stopping therapy the trends of these levels varied among individual patients. These results suggest that estrogen could act directly to suppress sodium-dependent Pi reabsorption in the renal proximal tubules. PMID- 10720066 TI - Expression of corticotropin-releasing hormone messenger ribonucleic acid in human pituitary corticotroph adenomas associated with proliferative potential. AB - Among the factors that promote the growth of human pituitary corticotroph adenomas (hPCAs), the proliferative potential of CRH secreted by hPCAs on these tumors is not well known. In this study, the CRH messenger ribonucleic acid (mRNA) transcripts were demonstrated on paraffin sections using the quantitative in situ hybridization method in 37 of 43 hPCAs, including 17 of 22 microadenomas, 15 of 15 macroadenomas, and 5 of 6 locally invasive adenomas according to Hardy's classification of pituitary adenomas. The more important findings were that CRH mRNA signal intensity in pituitary corticotroph adenoma cells was linearly correlated with Ki-67 tumor growth fractions (r = 0.802; P < 0.0001), and in macroadenoma and locally invasive adenoma cells it was significantly higher than in microadenoma cells (P = 0.035). On the other hand, CRH mRNA transcript accumulation was absent or negligible in 10 normal pituitary glands (P = 0.005). This is the first report of the frequent expression of CRH mRNA localized in human pituitary corticotroph adenoma cells. These results indicate that CRH from a local source of corticotroph adenoma cells not only has autocrine/paracrine functions in corticotroph adenomatous tissue, but also is an important factor associated with a proliferative potential of hPCAs. PMID- 10720067 TI - 17alpha-hydroxylase/17,20-lyase deficiency as a model to study enzymatic activity regulation: role of phosphorylation. AB - Cytochrome P450 17alpha-hydroxylase (CYP17) is a single gene-encoded protein with two activities: 17alpha-hydroxylase and 17,20-lyase. The two catalytic activities are differentially regulated in health and disease. We took advantage of naturally occurring human mutations to understand the molecular bases of this differential regulation. We identified eight novel mutations in the CYP17 gene, different in nature and spread throughout the gene. As posttranslational modifications appear to be important for activity control, we investigated the phosphorylation state of wild-type and mutant CYP17 proteins. Although phospholabeled protein was seen when the wild-type and most mutant proteins were expressed, no phosphorylation was detected for the F417C mutant. F417C is the only 17,20-lyase deficiency case confirmed at the molecular level and represents the first phosphorylation CYP17-deficient mutant. In search of the physiological agents involved in this process, the effect of cAMP was tested on activity and phosphorylation state of our mutant CYP17 proteins. cAMP stimulates activity and phosphorylation in all cases, except in the F417C and R35L mutants. The lack of response to the physiological second messenger might explain the different phenotypes. The F417C mutant protein, which is already shown to be associated with the lack of electron transfer, provides for the first time a link between the electron transfer system and the phosphorylation state of the CYP17 enzyme in the control of 17,20-lyase activity. PMID- 10720068 TI - The intracellular mechanism of insulin resistance in pancreatic cancer patients. AB - The diabetes that frequently occurs in pancreatic cancer patients is characterized by profound peripheral insulin resistance. The intracellular mechanism of this insulin resistance was investigated in skeletal muscle biopsies from pancreatic cancer patients with or without diabetes and control subjects. Insulin receptor (IR) binding, tyrosine kinase activity, IR messenger RNA (mRNA), IR substrate-1 content, GLUT-4, and GLUT-4 mRNA content were all normal in pancreatic cancer patients. In contrast, multiple defects in glycogen synthesis were found in pancreatic cancer patients, especially in those with diabetes. Glycogen synthase I activity, total activity, and mRNA levels were significantly decreased in pancreatic cancer patients compared with controls. The fractional velocity of glycogen synthase was decreased only in the diabetic pancreatic cancer group. Glycogen phosphorylase a and b activities were increased in diabetic pancreatic cancer patients, but glycogen phosphorylase mRNA levels were not significantly different. The insulin resistance associated with pancreatic cancer is associated with a post-IR defect, which impairs skeletal muscle glycogen synthesis and glycogen storage. PMID- 10720069 TI - Corticotropin-releasing hormone gene expression in primary placental cells is modulated by cyclic adenosine 3',5'-monophosphate. AB - CRH, the principal neuropeptide regulator of pituitary ACTH secretion, is also expressed in placenta. Placental CRH has been linked to the process of human parturition. However, the mechanisms regulating transcription of the CRH gene in placenta remain unclear. cAMP signaling pathways play important roles in regulating the expression of a diverse range of endocrine genes in the placenta. Therefore, we have explored the effect of cAMP on CRH promoter activity in primary cultures of human placental cells. Both forskolin and 8-bromo-cAMP, activators of protein kinase A, can increase CRH promoter activity 5-fold in transiently transfected human primary placental cells, in a manner that parallels the increase in endogenous CRH peptide. Maximal stimulation of CRH promoter activity occurs at 500 micromol/L 8-bromo-cAMP and 10 micromol/L forskolin. Electrophoretic mobility shift assay and mutation analysis combined with transient transfection demonstrate that in placental cells cAMP stimulates CRH gene expression through a cAMP regulatory element in the proximal CRH promoter region and involves a placental nuclear protein interacting specifically with the cAMP regulatory element. PMID- 10720070 TI - Tissue iodine content and serum-mediated 125I uptake-blocking activity in breast cancer. AB - In the thyroid, active transport of iodide is under control of the TSH-dependent Na+/I- symporter (NIS), whereas in the breast such control is less well understood. In this study, NIS expression was demonstrated by RT-PCR in 2 of 2 fibroadenomata and 6 of 7 breast carcinoma messenger ribonucleic acid isolates. In addition, mean total tissue iodine levels of 80.9 +/- 9.5 ng I/mg protein in 23 benign tumors (fibroadenomata) were significantly higher than those in 19 breast cancers taken from either the tumor (18.2 +/- 4.6 ng I/mg) or morphologically normal tissue taken from within the tumor-bearing breast (31.8 +/ 4.9 ng I/mg; P < 0.05 in each case). Inhibition of 125I uptake into NIS transfected CHO cells was observed in serum from 20 of 105 (19.0%) breast carcinoma, 8 of 49 (16.3%) benign breast disease, and 27 of 86 (31.4%) Graves' patients, but in only 1 of 33 (3.0%) age-matched female controls. IgG purified from serum of patients showing positive 125I uptake inhibition also inhibited iodide uptake, suggesting that such inhibition was antibody mediated. 125I uptake inhibition was significantly associated with thyroid peroxidase antibody positivity (P < 0.05) in sera from breast cancer patients, but not in those with benign breast disease, once again suggesting an association between thyroid autoimmunity and breast carcinoma. PMID- 10720071 TI - The relationship between glucose disposal in response to physiological hyperinsulinemia and basal glucose and free fatty acid concentrations in healthy volunteers. AB - This study was initiated to see if defects in the ability of physiological hyperinsulinemia (approximately 60 microU/mL) to stimulate glucose uptake in healthy, nondiabetic volunteers are associated with increases in concentrations of plasma glucose and free fatty acid (FFA) when measured at basal insulin concentrations (approximately 10 microU/mL). We recruited 22 volunteers (12 women and 10 men) for these studies, with a (mean +/- SEM) body mass index of 24.8 +/- 0.5 kg/m2. Resistance to insulin-mediated glucose disposal during physiological hyperinsulinemia was determined by suppressing endogenous insulin and determining the steady-state plasma glucose (SSPG) and steady-state plasma insulin (SSPI) concentrations at the end of a 3-h infusion, period during which glucose (267 mg/m2 x min) and insulin (32 mU/m2 x min) were infused at a constant rate. Glucose, insulin and FFA concentrations were also measured in response to infusion rates of glucose (50 mg/m2 x min) and insulin (6 mU/m2 x min). The SSPI concentration (mean +/- SEM) during physiological hyperinsulinemia was 64 +/- 3 microU/mL), in contrast to 12 +/- 0.4 microU/mL during the basal insulin study. The results demonstrated a significant relationship between SSPG concentration in response to physiological hyperinsulinemia (SSPG60) and SSPG(Basal) (r = 0.57, P < 0.01) and FFA(Basal) (r = 0.73, P < 0.001). Furthermore, FFA(Basal) and SSPG(Basal) were significantly correlated (r = 0.47, P < 0.05). Comparison of the seven most insulin-resistant and seven most insulin sensitive individuals (SSPG60 values of 209 +/- 16 vs. 64 +/- 8 mg/dL) revealed that the insulin-resistant group also had significantly higher SSPG(Basal) (105 +/- 5 vs. 78 +/- 7 mg/dL, P < 0.01) and FFA(Basal) (394 +/- 91 vs. 104 +/- 41, P < 0.02) concentrations. However, random fasting plasma glucose and FFA concentrations of the two groups were not different. The results presented demonstrate that individual differences in the ability of elevated insulin concentrations to stimulate muscle glucose disposal are significantly correlated with variations in insulin regulation of plasma glucose and FFA concentrations at basal insulin concentrations. PMID- 10720073 TI - Amino acid residue 147 of human aldosterone synthase and 11beta-hydroxylase plays a key role in 11beta-hydroxylation. AB - A number of amino acids differ between aldosterone synthase and 11beta hydroxylase. To assess their importance in determining the different functional specificities, we substituted aldosterone synthase-specific (aspartate D147, isoleucine I248, glutamine Q43, and threonine T493) with 11beta-hydroxylase specific amino acids (glutamate E147, threonine T248, arginine R43, and methionine M493), respectively. I248T, Q43R, and T493M had no effect on steroid production compared to wild-type aldosterone synthase. However, CYP11B2-D147E caused a significant increase in corticosterone production and a smaller increase in aldosterone production from 11-deoxycorticosterone (DOC). This appeared to be predominantly due to an increase in the 11beta-hydroxylation of DOC to corticosterone mediated by a decrease in Km, which was 1.4 micromol/L for the mutant compared with 5 micromol/L for the wild-type enzyme. CYP11B2-D147E had no effect on the conversion of 11-deoxycortisol to cortisol. The reverse construct (CYP11B1-E147D), substituting the 11beta-hydroxylase residue with the aldosterone synthase equivalent, decreased the conversion of DOC to corticosterone, which was mediated by an increase in Km that was 7.5 micromol/L for the mutant compared with 2.5 micromol/L for the wild-type enzyme. Again, the conversion of 11 deoxycortisol to cortisol was unimpaired. Thus, amino acid 147 is involved in the transformation of the 17-deoxysubstrate, but not the 17alpha-hydroxysubstrate. The results demonstrate that a conservative change in amino acid, even at some linear distance from known active centers, can significantly affect enzyme substrate affinity and subsequent steroid hormone production. PMID- 10720072 TI - Islet cell antibody-positive relatives with human leukocyte antigen DQA1*0102, DQB1*0602: identification by the Diabetes Prevention Trial-type 1. AB - The presence of human leukocyte antigen (HLA) haplotype DQA1*0102, DQB1*0602 is associated with protection from type 1 diabetes. The Diabetes Prevention Trial type 1 has identified 100 islet cell antibody (ICA)-positive relatives with this protective haplotype, far exceeding the number of such subjects reported in other studies worldwide. Comparisons between ICA+ relatives with and without DQB1*0602 demonstrated no differences in gender or age; however, among racial groups, African-American ICA+ relatives were more likely to carry this haplotype than others. The ICA+ DQB1*0602 individuals were less likely to have additional risk factors for diabetes [insulin autoantibody (IAA) positive or low first phase insulin release (FPIR)] than ICA+ relatives without DQB1*0602. However, 29% of the ICA+ DQB1*0602 relatives did have IAA or low FPIR. Although half of the ICA+ DQB1*0602 relatives had a high risk second haplotype, this was not associated with the additional risk factors for diabetes. Hispanic ICA+ individuals with DQB1*0602 were more likely to be IAA positive or to have low FPIR than other racial groups. In conclusion, the presence of ICA in the relatives described here suggests that whatever the mechanism that protects DQB1*0602 individuals from diabetes, it is likely to occur after the diabetes disease process has begun. In addition, there may be different effects of DQB1*0602 between ethnic groups. PMID- 10720074 TI - Glucose metabolism rather than insulin is a main determinant of leptin secretion in humans. AB - Circulating plasma insulin and glucose levels are thought to be major regulators of leptin secretion. There is evidence from in vitro and animal experiments that glucose metabolism rather than insulin alone is a main determinant of leptin expression. Here, we tested the hypothesis that in humans also leptin secretion is primarily regulated by glucose uptake and only secondarily by plasma insulin and glucose. In 30 lean and healthy men we induced 4 experimental conditions by using the blood glucose clamp technique. A total of 60 hypoglycemic and euglycemic clamps, lasting 6 h each, were performed. During these clamps insulin was infused at either high (15.0 mU/min x kg) or low (1.5 mU/min x kg) rates, resulting in low-insulin-hypo, high-insulin-hypo, low-insulin-eu, and high insulin-eu conditions. Serum leptin increased from 0-360 min by 20.5 +/- 4.1% in the low-insulin-hypo, 33.6 +/- 7.6% in the high-insulin-hypo, 39.6 +/- 6.0% in the low-insulin-eu, and 60.4 +/- 7.6% in the high-insulin-eu condition. Multiple regression analysis revealed a significant effect of circulating insulin (low vs. high insulin; P = 0.001) and blood glucose (hypoglycemia vs. euglycemia; P = 0.001) on the rise of serum leptin. However, when the total amount of dextrose infused during the clamp (grams of dextrose per kg BW) was included into the regression model, this variable was significantly related to the changes in serum leptin (P = 0.001), whereas circulating insulin and glucose had no additional effect. These findings in humans support previous in vitro data that leptin secretion is mainly related to glucose metabolism. PMID- 10720075 TI - Heritability of prostate-specific antigen and relationship with zonal prostate volumes in aging twins. AB - Both benign prostatic hyperplasia and prostate-specific antigen (PSA) have been shown to increase with age and with prostate volume in men, but the influence of heredity on these relationships is not completely understood. This study has two aims: 1) to investigate the inter-relationships of age, PSA, and various zonal measurements in the prostate; and 2) to assess the impact of heritable influences on total PSA. Eighty-four monozygotic twin pairs and 83 dizygotic twin pairs were studied, and serum total PSA, free PSA, and PSA-alpha1-antichymotrypsin were measured. Their prostate volumes [total (TV), transition zone (TZ), and peripheral zone) were quantitated using transrectal ultrasound. Total PSA is significantly correlated with all zonal prostate measurements (TZ, peripheral zone, TV, and TZ/TV) and with age. When linear regression was applied, only age and TZ were retained in the final model. The proportion of variability in total PSA explained by these two factors, however, is below 24%. In contrast, estimates of heritability show that approximately 45% of the variability in total PSA can be explained by inherited factors. Whereas age and TZ are linearly related to total PSA, their influence is much less than that of familial and genetic factors. It is uncertain whether these factors predispose also to prostate cancer or if they are independent of those, whether they confound the accuracy of using total serum PSA level as a diagnostic tool. PMID- 10720076 TI - Presence of a 5-HT7 receptor positively coupled to adenylate cyclase activation in human granulosa-lutein cells. AB - Although serotonin (5-HT) has been shown to stimulate progesterone production by human granulosa-lutein cells (hGLC), the receptor type and associated signaling pathway remain uncharacterized. We report here that 5-HT receptors in these cells are positively coupled to adenylate cyclase activity. Formation of cAMP was stimulated by 5-HT and its agonists in a dose- and time-dependent manner. Mianserin, amoxapine, and loxapine were equipotent in antagonizing 5-HT-induced cAMP formation. For both cAMP formation in cells and adenylate cyclase assay using membrane fractions, the rank order of potency for agonists of 5-HT were: 5 carboxy-aminotryptamine >5-HT> or =5-methoxytryptamine, consistent with a typical pharmacological profile of human 5-ht7 (h5-ht7) receptor. Sequence data of amplified complementary DNA fragments reverse transcribed from hGLC RNA revealed complete identity with published sequence of h5-ht7 receptor complementary DNA. Northern analysis showed the presence of 2.8-kb h5-ht7 transcripts in hGLC. The three variants h5-ht7A, h5-ht7B, and h5-ht7D were also detected in hGLC. Preincubation of hGLC with 5-HT (10(-8)-10(-6) M) resulted in a marked reduction in the cAMP response when the cells were subsequently stimulated with gonadotropin, and this heterologous desensitization could be reversed by 5-ht7 receptor antagonist clozapine. These data demonstrate that h5-ht7 receptor is present and stimulate cAMP formation in hGLC. In addition, the h5-ht7 receptor seems to be implicated in the heterologous down-regulation hCG-stimulated cAMP response in hGLC, with a possible ramification for luteal insufficiency. PMID- 10720077 TI - Octreotide may act as a radioprotective agent in acromegaly. AB - Clinical experience shows that an increasing number of patients undergoing radiation treatment for recurring acromegaly or acromegaly persisting after surgery are treated with octreotide. We, therefore, performed a follow-up study of patients undergoing stereotactic radiosurgery (Gamma Knife) to determine whether this medication has an influence on the ultimate result of radiation therapy in either a positive or negative sense. It has been suggested that the combination of radiation with antisecretory drugs may increase the effectiveness of radiation. A follow-up study of 31 patients suffering from recurrent acromegaly and acromegaly persisting after surgery, and who had been treated with stereotactic radiosurgery, showed that patients treated with octreotide at the time of radiation application simultaneously reached a normal level of growth hormone and insulin-like growth factor-I only after a significantly longer interval than patients who did not receive the drug. The two groups of patients did not demonstrate significant differences in the main clinical findings (age, sex, target volume, radiation dose, baseline growth hormone, and baseline insulin like growth factor-I). PMID- 10720079 TI - Adenovirus-mediated herpes simplex virus type-1 thymidine kinase gene therapy suppresses oestrogen-induced pituitary prolactinomas. AB - We tested the hypothesis that gene transfer using recombinant adenovirus vectors (RAds) expressing herpes simplex virus type 1 thymidine kinase (HSV1-TK) might offer an alternative therapeutic approach for the treatment of pituitary prolactinomas that do not respond to classical treatment strategies. HSV1-TK converts the prodrug ganciclovir (GCV) to GCV monophosphate, which is in turn further phosphorylated by cellular kinases to GCV triphosphate, which is toxic to proliferating cells. One attractive feature of this system is the bystander effect, whereby untransduced cells are also killed. Our results show that RAd/HSV1-TK in the presence of GCV is nontoxic for the normal anterior pituitary (AP) gland in vitro, but causes cell death in the pituitary tumor cell lines GH3, a PRL/GH-secreting cell line, and AtT20, a corticotrophic cell line. We have used sulpiride- and oestrogen-induced lactotroph hyperplasia within the rat AP gland as an in vivo animal model. Intrapituitary infection of rats bearing oestrogen induced lactotroph hyperplasia, with RAd/ HSV1-TK and subsequent treatment with GCV, decreases plasma PRL levels and reduces the mass of the pituitary gland. More so, there were no deleterious effects on circulating levels of other AP hormones, suggesting that the treatment was nontoxic to the AP gland in situ. In summary, our results show that suicide gene therapy using the HSV1-TK transgene could be further developed as a useful treatment to complement current therapies for prolactinomas. PMID- 10720078 TI - Identification of novel human GH-1 gene polymorphisms that are associated with growth hormone secretion and height. AB - Height, which is partially determined by GH secretion, is genetically influenced. The purpose of this study was to identify polymorphisms in the GH-1 gene, which are associated with altered GH production. The subjects included prepubertal short children with GH insufficiency without gross pituitary abnormalities (n = 43), short children with normal GH secretion (n = 46), and normal adults (n = 294). A polymorphism in intron 4 (P-1, A or T at base 1663) was identified. Two additional polymorphic sites (P-2, T or G at base 218, and P-3, G or T at base 439) in the promoter region of the GH-1 gene were also identified and matched with the P-1 polymorphism (A or T, respectively) in more than 90% of the subjects. P-1, P-2, and P-3 were considered to be associated with GH production, and the results of P-2 are explained as a representative in this abstract. For example, the allele frequency of T at P-2 in prepubertal short children with GH insufficiency without gross pituitary abnormalities (58.1%) was significantly different from that in short children with normal GH secretion and normal adults (37.0% and 43.5%, respectively; P < 0.001). Furthermore, significant differences were observed in maximal GH peaks in provocative tests (11.1 vs. 18.2 ng/mL, P = 0.006), insulin-like growth factor I SD scores (SDS) (-2.4 vs. -0.8, P < 0.0001), and height (Ht) SDS (-3.7 vs. -3.0, P = 0/001) in children with the T/T or G/G genotypes at P-2, respectively. In the entire study group, significant differences in insulin-like growth factor SDS (T/T, -0.9; G/G, -0.2; P = 0.0009) and Ht SDS (T/T, -1.0; G/G, -0.4; P = 0.022) were observed between the T/T and G/G genotypes at P-2. These data indicate that GH secretion is partially determined by polymorphisms in the GH-1 gene, which explain some of the variations in GH secretion and Ht. PMID- 10720080 TI - 11Beta-hydroxysteroid dehydrogenase type II and mineralocorticoid receptor in human placenta. AB - In mineralocorticoid target organs, 11beta-hydroxysteroid dehydrogenase type II (11beta-HSD2) confers specificity on the mineralocorticoid receptor (MR) by converting biologically active glucocorticoids to inactive metabolites. Placental 11beta-HSD2 is also thought to protect the fetus from high levels of circulating maternal glucocorticoid. In this study, we examined the immunoreactivity of 11beta-HSD2 and MR in human placenta from 5 weeks gestation to full term using immunohistochemistry, 11beta-HSD2 messenger RNA (mRNA) expression using Northern blot analysis, and MR mRNA expression using RT-PCR analysis. Marked 11beta-HSD2 immunoreactivity was detected in placental syncytiotrophoblasts at all gestational stages. MR immunoreactivity was moderately detected in syncytiotrophoblasts, some cytotrophoblasts, and interstitial cells of the villous core. Marked mRNA expression of 11beta-HSD2 was detected in placenta by Northern analysis. RT-PCR analysis of MR in placental tissues showed an amplified product consistent in length with the primers selected. These results suggest that placental 11beta-HSD2 is involved in not only regulating the passage of maternal active glucocorticoids into the fetal circulation but also in regulation of maternal-fetal electrolyte and water transport in the placenta, as in other mineralocorticoid target organs. PMID- 10720081 TI - Growth hormone (GH) responses to GH-releasing hormone alone or combined with arginine in patients with adrenal incidentaloma: evidence for enhanced somatostatinergic tone. AB - Spontaneous and stimulated GH secretion is blunted in hypercortisolemic states due to increased hypothalamic somatostatinergic tone. However, no data are available on the characteristics of GH secretion in patients with incidentally discovered adrenal adenomas. They represent an interesting model for studying GH secretion, as a slight degree of cortisol excess may frequently be observed in such patients who do not present with any clear Cushingoid sign. In the present study, 10 patients (3 men and 7 women, aged 48-63 yr) with an adrenal mass discovered serendipitously underwent, on separate occasions, a GHRH injection alone or combined with an infusion of the functional somatostatin antagonist, arginine. Thirteen age-matched healthy volunteers served as controls. Briefly, arginine (30 g) was infused from -30 to 0 min, and GHRH (100 microg) was injected as a bolus at 0 min, with measurement of serum GH [immunoradiometric assay (IRMA)] every 15 min for 150 min. Plasma IGF-I (RIA after acid-ethanol extraction) was measured in a morning sample. The diagnosis of cortical adenoma was based on computed tomography features and pattern of uptake on adrenal scintigraphy. Patients with obesity and/or diabetes were excluded. The study design included also an endocrine work-up aimed to study the hypothalamic pituitary-adrenal axis [urinary free cortisol (UFC) excretion, serum cortisol at 0800 h, plasma ACTH at 0800 h, morning cortisol after overnight 1 mg dexamethasone]. Five of 10 patients showed abnormalities of the hypothalamic pituitary-adrenal axis, including borderline or increased UFC excretion in 4 of them accompanied by blunted ACTH in 2 cases and failure of cortisol to suppress after dexamethasone in 1; the fifth patient displayed low ACTH and resistance to dexamethasone suppression. However, all patients had a unilateral uptake of the tracer on the side of the mass with suppression of the contralateral normal adrenal gland. As a group, the patients displayed greater UFC excretion and lower ACTH concentrations than the controls. GH release after GHRH treatment was blunted in patients bearing adrenal incidentaloma compared with controls (GH peak, 5.7 +/- 5.2 vs. 18.0 +/- 7.0 microg/L; P < 0.0001), whereas GHRH plus arginine was able to elicit a comparable response in the 2 groups (GH peak, 33.5 +/- 20.3 vs. 33.7 +/- 17.5 microg/L; P = NS). The ratio between GH peaks after GHRH plus arginine and after GHRH plus saline was significantly greater in patients than in controls (751 +/- 531% vs. 81 +/- 45%; P = 0.0001). Similar data were obtained when comparing GH area under the curve after GHRH plus saline or GHRH plus arginine between the 2 groups. In summary, the present data suggest that in patients with incidental adrenal adenomas the GH response to GHRH is blunted due to increased somatostatinergic tone, as it can be restored to normal by pretreatment with the functional somatostatin antagonist arginine. The blunted GH release to GHRH may be an early and long lasting sign of autonomous cortisol secretion by the adrenal adenoma. PMID- 10720082 TI - No increased insulin sensitivity after a single intravenous administration of a recombinant human tumor necrosis factor receptor: Fc fusion protein in obese insulin-resistant patients. AB - Inhibition of tumor necrosis factor (TNF)-alpha results in a marked increase in insulin sensitivity in obese rodents. We investigated the influence of a TNF antagonist [Ro 45-2081, a recombinant fusion protein that consists of the soluble TNF-receptor (p55) linked to the Fc portion of human IgG1] on insulin sensitivity of patients with android obesity. Seven patients (five women and two men; mean +/ SD age, 41 +/- 4 yr; body mass index, 36.1 +/- 4.7 kg/m2; waist to hip ratio, 0.99 +/- 0.11) were studied (three patients with normal glucose tolerance and four patients with impaired glucose tolerance or mild diabetes; all were hyperinsulinemic). Each patient underwent two consecutive euglycemic hyperinsulinemic glucose-clamp tests: 48 h after injection of placebo and 48 h after a single i.v. injection of 50 mg Ro 45-2081. In both tests, steady-state plasma glucose and insulin levels were similar. Insulin-mediated glucose disposal (2.23 +/- 0.74 vs. 2.38 +/- 0.99 mg/kg(-1) x min(-1)) and glucose metabolic clearance rate (2.28 +/- 0.85 vs. 2.48 +/- 1.03 mL/kg(-1) x min(-1)) were similar after placebo and after the drug. Indirect calorimetry showed no difference in substrate oxidation rates between the two experimental conditions. In conclusion, under the conditions of this study, no improvement in insulin sensitivity was observed in obese insulin-resistant patients following a single i.v. administration of a recombinant TNF receptor: Fc fusion protein. PMID- 10720083 TI - A heterozygous deletion of the autoimmune regulator (AIRE1) gene, autoimmune thyroid disease, and type 1 diabetes: no evidence for association. AB - Autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) is a rare monogenic autoimmune disease with endocrine components including type 1 diabetes, adrenal failure, and thyroid dysfunction, with major autoantibodies directed against adrenal, pancreas, and thyroid tissue. A 13-bp deletion in exon 8 of the autoimmune regulator (AIRE1) gene on chromosome 21q22.3 accounts for more than 70% of mutant alleles in United Kingdom subjects with APECED. To determine whether this polymorphism contributes to disease susceptibility in subjects with autoimmune disease in general, we screened 302 patients with Graves' disease, 154 patients with autoimmune hypothyroidism, 235 patients with type 1 diabetes, and 318 control subjects for the 13-bp deletion of the AIRE1 gene. The mutation was present in only 1 (0.33%) patient with Graves' disease, 1 patient with autoimmune hypothyroidism (0.6%), and 1 (0.315) of the control subjects. No patients with type 1 diabetes were found to carry the mutation. We conclude, therefore, that the 13-bp deletion of the AIRE1 gene is not a susceptibility locus for the more common autoimmune endocrinopathies in the United Kingdom. PMID- 10720084 TI - Missense mutations in the human insulin promoter factor-1 gene and their relation to maturity-onset diabetes of the young and late-onset type 2 diabetes mellitus in caucasians. AB - Increasing evidence suggests that defects in genes encoding transcription factors that are expressed in the pancreatic beta-cells may be important contributors to the genetic basis of type 2 diabetes mellitus. Maturity-onset diabetes of the young (MODY) now exists in five subtypes (MODY1-5), four of which are caused by mutations in transcription factors hepatocyte nuclear factor-4alpha (HNF-4alpha), HNF-1alpha, insulin promoter factor-1 (IPF-1), and HNF-1beta (MODY1, -3, -4, and 5). Recent evidence from the British population even suggested that IPF-1 may be a predisposing gene for type 2 diabetes. Thus, highlighting the potential role of this transcription factor in the genetic basis of Danish and Italian MODY as well as in Danish patients with late-onset type 2 diabetes mellitus, we have examined the human IPF-1 gene for mutations by single strand conformation polymorphism and heteroduplex analysis in 200 Danish patients with late-onset type 2 diabetes and in 44 Danish and Italian MODY patients. In the patients with late-onset type 2 diabetes we identified a noncoding G insertion/deletion polymorphism at nucleotide -108, a silent G54G, and a rare missense D76N variant. Moreover, a Danish MODY patient was carrier of an A140T variant. Neither the D76N nor the A140T segregated with diabetes, and their transcriptional activation of the human insulin promoter expressed in vitro was indistinguishable from that of the wild type (115 +/- 21% and 84 +/- 12% vs. 100%). We conclude that variants in IPF-1 are not a common cause of MODY or late-onset type 2 diabetes in the Caucasian population, and that in terms of insulin transcription both the N76 and the T140 mutations are likely to represent functionally normal IPF-1 variants with no direct role in the pathogenesis of MODY or late-onset type 2 diabetes mellitus. PMID- 10720085 TI - An unusual kindred of the multiple endocrine neoplasia type 1 (MEN1) in Japanese. PMID- 10720086 TI - Interleukin-6: the endocrine cytokine. PMID- 10720087 TI - Interleukin-6 gene polymorphism and lipid abnormalities in healthy subjects. AB - Several lines of evidence indicate that interleukin-6 (IL-6) is involved not only in the hepatic acute phase response but also in adipose tissue metabolism, lipoprotein lipase activity, and hepatic triglyceride secretion. A polymorphism in the IL-6 gene, associated with differences in IL-6 transcription rate, has been recently described. We aimed to study whether this IL-6 gene polymorphism leads to differences in fasting and postglucose load plasma lipids in healthy subjects. Subjects with G at position -174 of the IL-6 gene were similar in age, sex, body mass index, and waist to hip ratio in comparison with carriers of the C allele. However, G carriers showed almost twice plasma triglycerides (1.5 +/- 0.9 vs. 0.90 +/- 0.37 mmol/L; P = 0.01), very low-density lipoprotein (VLDL) triglycerides (0.97 +/- 0.69 vs. 0.42 +/- 0.2 mmol/L; P = 0.002), higher fasting (881 vs. 458 micromol/L; P = 0.01), and postglucose load free fatty acids (299 vs. 90.5 micromol/L; P = 0.03), slightly lower high-density lipoprotein-2 cholesterol (0.25 +/- 0.14 vs. 0.39 +/- 0.26 mmol/L; P = 0.058), and similar cholesterol and LDL-cholesterol levels than carriers of the C allele. Serum IL-6 levels correlated positively with fasting triglycerides, VLDL-triglycerides, and postload free fatty acids (r = 0.61, 0.65 and 0.60, respectively; P < 0.001) and negatively with high-density lipoprotein-cholesterol (r = -0.42, P < 0.05). A tendency toward higher serum IL-6 levels was observed among G carriers (9.9 +/- 6.9 vs. 6.85 +/- 1.7 pg/mL; P = 0.09). The -174G construct was recently reported to show higher expression of IL-6 in He La cells and was associated with higher plasma IL-6 levels than the -174C allele. Thus, the results of the present study suggest that subjects with the G allele, associated to higher IL-6 secretion, are prone to lipid abnormalities. Whether this polymorphism contributes to lipid alterations associated with other metabolic disorders awaits additional studies. PMID- 10720089 TI - Circulating iodide concentrations during and after pregnancy. PMID- 10720088 TI - Hormone replacement therapy and interrelation between serum interleukin-6 and body mass index in postmenopausal women: a population-based study. AB - Postmenopausal women are at increased risk to develop osteoporosis, coronary artery disease, heart failure, and hypertension. Interleukin-6 (IL-6) may be a pathogenetic element in these disorders. Serum IL-6 levels increase during aging and seem to be related to increased body fat mass. In the present retrospective study we aimed to investigate the role of hormone replacement therapy (HRT) on serum IL-6 levels and the interrelation of IL-6 and body fat mass. Parameters were assessed in a population-based sample of postmenopausal women (n = 302) and, for comparison, 245 men of the same age. Women with HRT (n = 92) had significantly lower serum IL-6 levels compared to subjects without HRT, which was independent of age, antihypertensive therapy, smoking habits, and blood pressure (1.5 +/- 0.1 vs. 2.9 +/- 0.6 pg/mL; P = 0.017). In women without HRT, the body mass index (BMI) was correlated with serum IL-6 levels (P < 0.001). Multivariate analysis controlling simultaneously for the effects of blood pressure and heart rate confirmed the positive correlation (P = 0.001). However, in subjects with HRT no such correlation between IL-6 and BMI was demonstrated, which was confirmed after controlling covariates. In male subjects, BMI correlated with serum IL-6 (P = 0.009), which was, however, blunted after controlling for blood pressure and heart rate, probably indicating an influence of the sympathetic nervous system on this interrelation. In conclusion, women receiving HRT display lower serum IL-6 levels and a blunted interrelation of IL-6 and BMI. As IL-6 may be a pathogenetic factor in age-related diseases, HRT-related inhibition of IL-6 secretion could be an important element for the favorable effects of HRT in postmenopausal women. PMID- 10720090 TI - Growth hormone treatment in patients with intrauterine growth retardation. PMID- 10720091 TI - Role of cytochrome b5 in the 17,20-lyase activity of P450c17. PMID- 10720092 TI - Comment on menstrual irregularity in women with acromegaly. PMID- 10720093 TI - Intracranial arachnoid cysts. PMID- 10720094 TI - The arterial supply of the carpal joint in the Bactrian camel (Camelus bactrianus). AB - The arterial supply of the six carpal joints was studied in Bactrian camels. The arterial branches supplying the carpal joints were the proximal medial dorsal, middle medial dorsal, distal medial dorsal, proximal lateral dorsal, distal lateral dorsal, proximal medial palmar, distal medial palmar, lateral palmar, proximal palmar and dorsal palmar carpi and the dorsal interosseous antebrachium branches. Some small unnamed branches supplied the diverticulum and the antebrachiocarpal joint capsule. All these arteries arose from the lateral and medial branches of the radial artery. PMID- 10720095 TI - A retrospective evaluation of the causes of death of 448 insured French horses in 1995. AB - Epidemiological studies should allow comparisons to be made of the prevalence of disease in populations from different countries, but the population characteristics and health problems in French horses are not well established. We have conducted a retrospective evaluation of the causes of death and vital characteristics of insured horses in France for the year 1995, with a view to comparison with published data from other countries. Files on 448 deceased horses were provided by nine insurance companies. Most of the animals were used for breeding (60%), followed by leisure (20%), eventing and racing (10% each). Physical characteristics were associated significantly with occupational categories. The overall mortality rate was 2.47%, and was due, in decreasing order, to foaling (24%), colic (21%) or locomotor (21%), cardiovascular (9%), neurological (8%), respiratory (5%) or infectious (4%) disease. Infectious disease was more frequent in younger animals (p < 0.05) and locomotor disease in racehorses (p < 0.01). Horses aged over 15 years had a lower incidence of colic (p <0.05). The cause of death was not significantly linked to breed, insurance value or season. Despite some selection bias, the study provides useful information about mortality in the French equine population. PMID- 10720096 TI - A comparison of three methods for extracting IgY from the egg yolk of hens immunized with Sendai virus. AB - Hens were immunized with partially purified Sendai virus that had been grown on chicken embryos. The titres of specific antibodies to Sendai virus varied from log2 13.0 to 15.5 during the 4 months after immunization and the immunoglobulin Y (IgY) concentration varied from 2 to 10 mg per ml of egg yolk. A method has been developed employing dextran blue to isolate IgY from the egg yolk. The dextran blue method was compared with two other methods (poly(ethylene glycol) and chloroform) in terms of yield, total protein content, IgY concentration, specific activity of IgY and SDS-PAGE analysis. The specific activity and the IgY content obtained by these three methods proved to be identical, in the range log2 12.9 14.1, and 4.6-12.8 mg/egg, respectively. However, the total protein content when purified by chloroform was 2-fold to 4-fold higher than that of the others. The analysis of IgY by SDS-PAGE demonstrated that IgY purified with dextran blue contained three major protein components with molecular weights of 34.7 kDa, 41 kDa and 66 kDa and one minor protein of 45 kDa. IgY that was extracted with chloroform contained two major proteins of 45.7 kDa and 75.2 kDa and that extracted with PEG-6000 contained only one major protein of 66 kDa. The IgY obtained by these latter methods also contained several minor proteins in the range 41-80 kDa. PMID- 10720097 TI - Absence of p21 WAF1 and p27 kip1 gene mutations in various feline tumours. AB - The coding regions of tumour suppressor and cell cycle regulatory genes p21 WAF1 and p27 Kip1 were investigated in 101 feline tumours of various types. No damaging mutations were present in the analysed areas of the genes. PMID- 10720098 TI - Effects of mixed grazing of first- and second-year calves on trichostrongylid infections in Lithuania. AB - The objective of this study was to examine whether susceptible calves grazing together with second-year resistant heifers are less exposed to trichostrongylid infection than are calves grazing on their own. Two groups of animals representing each age category were turned out onto pasture on 24 May 1997 and grazed at comparable stocking rates. The grazing of calves and heifers together was compared to groups of each age category grazing separately. The results indicated that herbage larval counts were significantly reduced in the second part of the grazing season on the plot grazed by the mixed group compared to the plot grazed by the first-season calves only. The mixed grazing strategy protected the young calves and no clinical signs were observed in this group, while most of the calves that grazed alone exhibited clinical signs. The availability of herbage was reduced towards the end of the season, with subsequent competition for the grass forcing all the animals to graze the tufts around the faecal pats, where the quality of the grass is poor and the numbers of infective larvae are high. The effect of this was visible in the form of increased parasite burdens in the calves that were grazed together with the heifers, confirmed by increased blood serum pepsinogen concentrations and reduced daily weight gains in the second part of the grazing season. The lower numbers of infective larvae on the pasture were probably achieved through the heifers ingesting many of the larvae but subsequently depositing relatively few eggs, since they had acquired some degree of resistance against trichostrongylid infections during their first grazing season. Thus they did not suffer any parasitological ill-effects during mixed grazing with first-season calves. PMID- 10720100 TI - Peptide nucleic acids: on the road to new gene therapeutic drugs. AB - Peptide nucleic acid (PNA) is a DNA mimic based on a pseudopeptide (polyamide) backbone. PNA oligomers bind strongly and with high sequence specificity to complementary targets in RNA (or DNA), and they show very high biological stability. Furthermore, studies in cell free systems have demonstrated potent antisense (inhibition of translation) and antigene (inhibition of transcription) activity of PNA. Recently, several studies reporting methods for cellular delivery of PNA as well as antisense effects of PNA in cells ex vivo and in rats have appeared. The potential of developing PNA derived gene therapeutic drugs is discussed. PMID- 10720099 TI - Characterization of Newcastle disease viruses isolated from chickens and ducks in Tamilnadu, India. AB - During 1993, outbreaks of Newcastle disease occurred on many farms in Tamilnadu, India. Six Newcastle disease virus (NDV) isolates were obtained from the chickens on five different farms and from the birds on one duck farm during outbreaks of the disease. All the isolates were characterized as velogenic, based on the mean death time, intravenous pathogenicity index, intracerebral pathogenicity index (ICPI), stability of haemagglutinin at 56 degrees C, agglutination of equine erythrocytes, haemagglutination elution pattern and adsorption of haemagglutinin by chick brain cells. The isolate obtained from ducks resembled a group D strain, based on its ICPI and its reaction with a panel of monoclonal antibodies. The other five NDV isolates obtained from chickens were placed in groups B(1), C1(2) and D(2) on the basis of their binding patterns with the panel of monoclonal antibodies. In challenge experiments, it was found that LaSota vaccine provided 100% protection against each of these field isolates and against a local NDV strain obtained from the Institute of Veterinary Preventive Medicine, Tamilnadu, India, while unvaccinated chickens succumbed to challenge. The possible origin of epizootic viruses causing outbreaks in vaccinated flocks is discussed. PMID- 10720101 TI - The influence of repeated prestressors on single stress response in rats. AB - The effect of various stressors of different intensity applied in random order before the final single immersion restraint stress was tested in two inbred rat strains, isoprenaline-sensitive and isoprenaline-resistant. The isoprenaline sensitive strain revealed higher incidence of heart lesions after this single acute stress and low incidence of gastric lesions. The isoprenaline-resistant strain had the opposite characteristics. These differences were constantly reproducible when this strong stressor was used. After prestress by different stressors (tail-flick, ether anaesthesia, Porsolt swimming stress) at different time schedules, the incidence of gastric ulcer lesions, the weight of organs (heart, adrenals, spleen) changed substantially in isoprenaline-sensitive rats only. The most important result was reversal of the extent of gastric lesions. The isoprenaline-sensitive strain revealed more lesions than the isoprenaline resistant one. The repeated different prestressors mainly changed the reactivity of animals, isoprenaline-sensitive rats becoming more similar to isoprenaline resistant rats. These findings urged us to interpret carefully the results obtained in stress research with different and multiple stressors both in animals and humans. PMID- 10720102 TI - Modulation of capsaicin-sensitive nerve activation by low pH solutions in guinea pig lung. AB - We have studied the stimulation of airways sensory nerves by low pH solutions and concomitantly induced bronchoconstriction. The effect of low pH buffer and lactic acid solutions at the same pH (5 and 6) were compared and the influence of low pH on the capsaicin effect was recorded. We have used the isolated guinea-pig perfused lung model taking the insufflation pressure as an indicator of bronchial smooth muscle tone while the calcitonin gene-related peptide-like immunoreactivity measured in the lung perfusate represented sensory nerves activation. Low pH buffer and lactic acid solution (3 and 4.1 mM) at the same pH of 5 and 6 induced pH-dependent bronchoconstriction and peptides release which were completely abolished after systemic pretreatment with capsaicin. Both responses were significantly inhibited after Ca2+-free infusion. Capsazepine (10( 6) M), a selective capsaicin antagonist, significantly reduced the calcitonin gene-related peptide-like immunoreactivity overflow evoked by all the solutions studied. Diclofenac (10(-5) M), a cyclooxygenase blocker, inhibited pH 5, pH 6 and lactic acid 3 mM (pH 6)-evoked peptide release, but not lactic acid 4.1 mM (pH 5). The functional response was not significantly modified after diclofenac while only the lactic acid 3 mM response was significantly reduced by capsazepine. There was a synergistic interaction between capsaicin and low pH buffer on calcitonin gene-related peptide-like immunoreactivity release and an additive effect on bronchoconstriction. It is concluded that in the isolated perfused guinea-pig lung, lactic acid and low pH buffer induced calcitonin gene related peptide-like immunoreactivity release and bronchoconstriction by stimulation of capsaicin-sensitive C fibres via a pathway partly dependent of extracellular Ca2+. The mechanism of calcitonin gene-related peptide-like immunoreactivity release seems to be the same at pH 6, while differences are evident at pH 5 between low pH buffer and lactic acid. Our results also suggest that proton activity could exert a modulatory role on the capsaicin-sensitive sensory nerves by a mechanism which remains to be clarified. PMID- 10720103 TI - Endocrine-disrupting agents on healthy human tissues. AB - A vast number of substances have been suggested as possibly contributing to perturbation of the endocrine system. Several have been tested with different approaches ranging from yeast expression system of human oestrogenic receptors to human breast cancer cells assays. Surprisingly, no inhibition-binding experiments to steroid receptors on healthy human tissue have been performed so far. Our study provides inhibition binding experiments to oestrogens, progesterone, testosterone and retinoic acid receptors in prostate and uterine human tissue of organochlorine pesticides, phthalate esters, oestrogenic constituents derived from plants and phenol derivates. Affinities of significant extent of phthalates on oestrogenic, progestinic and androgenic receptors have not been detected. As for retinoic acid receptors, mono(2-ethylexyl)phthalate provokes a notable reduction of the binding of the tritiated retinoic acid, phtalic acid ethyl-n butyl ester and 4-octylphenol show an affinity comparable to that of isoflavonoid genistein, whereas 4-nonylphenol reduces the binding of retinoic acid in prostate. PMID- 10720104 TI - Effect of omega-conotoxin GVIA on noradrenaline release from postganglionic sympathetic neurones in rabbit aorta. AB - The aim of the present work was to examine the effect of the selective N-type calcium blocking agent omega-conotoxin GVIA on stimulation-evoked release of noradrenaline from sympathetic nerves in rabbit isolated aorta with regard to stimulation frequency, extracellular Ca2+ concentration, and transmitter uptake. Rings of rabbit isolated aorta were preloaded with (-)-3H-noradrenaline and the fractional 3H-overflow evoked by electrical-field stimulation was determined by liquid scintillation spectrometry. Omega-conotoxin GVIA (3 x 10(-10)-3 x 10(-8) M) did not alter the spontaneous 3H-outflow. Omega-conotoxin GVIA (3 x 10(-10)-3 x 10(-8) M) caused a slowly developing reduction of stimulation-evoked 3H overflow at 1 and 30 Hz. The Emax for the omega-conotoxin-induced inhibition was less (70%) at 30 Hz than that (96%) seen at 1 Hz. Short-term incubation with omega-conotoxin GVIA caused a subsequent steady-state inhibition. The inhibitory action of omega-conotoxin GVIA (3 x 10(-10)-3 x 10(-9) M) was inversely related to the extracellular Ca2+ concentration (6.5 x 10(-4)-2.7 x 10(-3) M). Cocaine (3 x 10(-5) M) plus corticosterone (4 x 10(-5) M), neuronal and extraneuronal uptake inhibitors, respectively, did not alter the inhibitory effect of omega-conotoxin GVIA (3 x 10(-9) M) on 3H-overflow evoked by stimulation at a frequency of either 1 or 30 Hz. It is concluded that omega-conotoxin GVIA acts on prejunctional N type calcium channels to inhibit stimulation-evoked noradrenaline release from sympathetic neurone terminals in rabbit aorta. At a high frequency, another subtype calcium channel may possibly be involved. The action of omega-conotoxin GVIA is independent of neuronal and extraneuronal uptake mechanisms for noradrenaline, but dependent on the amount of Ca2+ to be transported across the neurilemma from the extracellular space into the neurone. PMID- 10720105 TI - Arterial responses in vitro and plasma digoxin immunoreactivity after losartan and enalapril treatments in experimental hypertension. AB - Treatment with the angiotensin-converting enzyme inhibitor, quinapril, has been shown to normalize increased dihydropyridine sensitivity and impaired potassium relaxation, characteristic features of arterial smooth muscle in spontaneously hypertensive rats, and also reduce the concentration of plasma digoxin-like immunoreactivity in these animals. However, whether angiotensin II receptor blocker therapy can beneficially influence these variables is not known. Therefore, we compared the effects of 10-week losartan and enalapril treatments (15 and 4 mg/kg/day, respectively) on functional responses of mesenteric arterial rings in spontaneously hypertensive rats and Wistar-Kyoto rats. Both losartan and enalapril normalized blood pressure, cardiac mass, and media to lumen ratio without significantly changing the media cross-sectional area in the mesenteric artery of spontaneously hypertensive rats (i.e. induced outward remodelling). The inhibitory effect of the calcium entry blocker nifedipine on calcium-evoked contractions was similar and less marked in arterial preparations from Wistar Kyoto rats and losartan- and enalapril-treated spontaneously hypertensive rats than in those from untreated spontaneously hypertensive rats. Furthermore, the relaxations of arterial rings induced by the return of potassium to the organ bath (upon precontractions elicited by potassium-free solution) were used to evaluate the function of vascular Na+,K+-ATPase. The rate of potassium relaxation was faster in losartan- and enalapril-treated spontaneously hypertensive rats and all Wistar-Kyoto groups than in untreated spontaneously hypertensive rats, and the response was effectively inhibited by the sodium pump inhibitor ouabain. Both treatments especially augmented the ouabain-sensitive part of the potassium relaxation in spontaneously hypertensive rats, indicating the involvement of the sodium pump in this response. However, no significant changes in plasma digoxin like immunoreactivity were observed. In conclusion, the outward remodelling following long-term AT1-receptor blockade and angiotensin-converting enzyme inhibition in spontaneously hypertensive rats was associated with normalization of the increased dihydropyridine sensitivity of arteries. Both losartan and enalapril treatments also augmented arterial potassium relaxation in spontaneously hypertensive rats, suggesting enhanced function of Na+,K+-ATPase, but this effect could not be attributed to changes in circulating sodium pump inhibitor concentration. PMID- 10720107 TI - Primary enteric neuropathies underlying gastrointestinal motor dysfunction. PMID- 10720106 TI - Pharmacological characterisation of the enantiomers of BM-5, a muscarinic partial agonist with opposed enantioselectivity between affinity and efficacy. AB - The interaction of (R) and (S) enantiomers of the chiral oxotremorine analogue BM 5 with muscarinic acetylcholine receptors was studied in vitro using radioligand binding and isolated tissue preparations. The in vivo effects of (R)-BM-5 were also studied in anaesthetised cat. No receptor or tissue selectivity was found for either enantiomer in radioligand binding studies in cells expressing human muscarinic receptors (M1-M5) or in guinea pig tissues. The affinity of (R)-BM-5 was about 40 times, or 15-60 times higher than that of (S)-BM-5 in recombinant cells or in guinea pig tissues, respectively. Both enantiomers induced contraction of the guinea pig isolated urinary bladder and ileum. (R)-BM-5 was more potent than (S)-BM-5 in bladder (EC50 590 and 3500 nM, respectively) and in ileum (EC50 39 and 2600 nM, respectively). The maximal agonist effect was lower for (R)-BM-5 than for (S)-BM-5 in bladder (2.7% and 6.6%, respectively) and in ileum (32% and 48%, respectively). Contractions were completely inhibited by atropine (1 microM). In vivo, (R)-BM-5 induced bladder contraction and salivation after intravenous administration in the anaesthetised cat (ED50 4.1 and 6.2 microg kg(-1), respectively). In conclusion, (R)- and (S)-BM-5 act as partial muscarinic agonists in the isolated bladder and ileum. (R)-BM-5 was the more potent enantiomer but had a lower maximal agonist effect giving an opposed enantioselectivity for affinity and efficacy. (R)-BM-5 showed agonist activity in vivo, confirming in vitro findings. From affinity and efficacy data it can be concluded that the effects of racemic BM-5 are mediated by the (R)-enantiomer. PMID- 10720108 TI - Human papillomavirus involvement in esophageal carcinogenesis in the high incidence area of China. A study of 700 cases by screening and type-specific in situ hybridization. AB - BACKGROUND: Human papillomavirus (HPV) DNA has been identified in esophageal precancerous lesions and carcinomas. However, there are marked variations in the prevalence of HPV infection reported in different studies. Most previous studies on HPV and esophageal carcinomas have been based on a limited number of biopsy samples studied by different HPV detection methods with highly variable sensitivity and specificity, making systematic studies of larger series clearly warranted. METHODS: A series of 1876 surgical specimens (primary tumor, adjacent epithelium, regional lymph nodes, resection margins) from 700 patients surgically resected for an invasive squamous cell carcinoma of the esophagus in the high incidence area of China was analyzed for the presence of HPV DNA with screening in situ hybridization (ISH) using biotinylated HPV DNA probes and followed by type-specific ISH for HPV 6, 11, 16, 18, 30, and 53. RESULTS: Of the 700 esophageal carcinomas, 118 (16.9%) were shown to contain HPV DNA sequences by screening ISH. Positive signals were most frequent in the cancer cells (16.6%), more rare in the surrounding hyperplastic and dysplastic epithelia (5.6%), and infrequently present in the resection margins (0.2%). HPV signals were also detected in cancer cells in 6.9% of the lymph node metastases. HPV types 6, 11, 16, 18, and 30 account for 39.8% of the HPV-positive lesions, of which the high risk types HPV 16 and 18 were present in 27.1% (32 of 118). Notably, 60.2% of the HPV-positive lesions contained DNA sequences other than HPV types 6, 11, 16, 18, 30, and 53. CONCLUSIONS: This study reports the largest series of esophageal cancers ever analyzed for the presence of HPV DNA. Our results confirm the presence of common mucosal HPV types in esophageal carcinomas but also suggest the involvement of other (novel?) HPV types that are unusually detected in genital cancers in a significant proportion of these lesions. The results further indicate that HVP has an etiologic role in esophageal carcinogenesis, at least in the high-incidence area of northern China. PMID- 10720110 TI - Diagnosis of Helicobacter pylori infection in patients with atrophic gastritis: comparison of histology, 13C-urea breath test, and serology. AB - BACKGROUND: Atrophic gastritis, a risk factor for gastric cancer, is a late consequence of Helicobacter pylori infection in approximately one-third of the infected patients. It has been suggested that gastric cancer would develop less frequently if H. pylori were eradicated. However, the prevalence of H. pylori infection may be underestimated in patients with atrophic gastritis and intestinal metaplasia if only biopsy-based diagnostic methods are used. METHODS: We compared histology, 13C-urea breath test (13C-UBT), and serology in H. pylori diagnostics in 50 male patients with atrophic corpus gastritis. RESULTS: H. pylori was detected in 15 (30%) patients by histology and in 14 (28%) by 13C-UBT, whereas increased serum antibody levels indicating H. pylori infection were found in 41 (82%) patients (P < 0.0001 between serology and both histology and 13C UBT). H. pylori infection was associated with atrophic corpus gastritis in 84% of the present patients (in one patient with normal antibody titres H. pylori was defined histologically). CONCLUSIONS: H. pylori infection would have been missed in most patients with atrophic gastritis without the analysis of H. pylori antibodies. Therefore, in patients with atrophic gastritis, the use of serology is encouraged in diagnosing H. pylori infection. PMID- 10720109 TI - Relationship between dysplasia, p53 protein accumulation, DNA ploidy, and Glut1 overexpression in Barrett metaplasia. AB - BACKGROUND: There is a need for molecular markers of malignant progression in Barrett metaplasia (BM). The aim of this study is to determine the relationship between dysplasia, p53 protein accumulation, DNA ploidy, and Glut1 in BM. METHODS: Sections of esophageal biopsy specimens from 120 patients with BM were evaluated for dysplasia, p53 protein, and Glut1 expression by immunohistochemistry, and DNA ploidy by Feulgen stain and image analysis. In cases with diploid DNA histograms, the percentage cells in the G0G1 and G2M phases of the cell cycle were determined. RESULTS: Of 108 diploid cases 19 (28%) of 69 cases with G0G1 > or = 90% or G2M > or = 8.33% were p53-positive, in contrast to only 1 (3%) of 39 cases with lower G0G1 or G2M (P = 0.0008). Of 32 p53-positive cases 11 (32%) were aneuploid, in contrast to none (0%) of 88 p53 negative cases (P < 0.0001). Ten (91%) of 11 aneuploid cases were high-grade dysplasial adenocarcinoma (HGD/CA), compared with only 1 (1%) of 109 diploid cases (P < 0.0001). Five (45%) of 11 cases with HGD/CA were Glut1-positive, in contrast to none (0%) of 109 cases without HGD/CA (P < 0.0001). CONCLUSIONS: Our data strongly suggest that in BM, after oxidative DNA damage, as a result of gastroesophageal reflux, there is an increase in the percentage of cells in the G0G1 or G2M phases of the cell cycle to enable repair of damaged DNA; in some of these cases this is followed sequentially by p53 gene mutation and protein accumulation, DNA aneuploidy, HGD, and CA with or without Glut1 overexpression. These events can be detected in routinely processed biopsy samples. PMID- 10720111 TI - Identifying dyspepsia and irritable bowel syndrome: the value of pain or discomfort, and bowel habit descriptors. AB - BACKGROUND: The utility of current diagnostic criteria for dyspepsia and irritable bowel syndrome (IBS) in clinical practice is largely unknown. We aimed to compare the diagnostic value of different definitions and questionnaires in a population. METHODS: The Abdominal Symptom Questionnaire (ASQ) was mailed to a representative sample (n = 1506, 20-87 years old), and every fifth person (n = 302) concomitantly received the Bowel Disease Questionnaire (BDQ). The diagnostic agreement for dyspepsia and for IBS, defined in accordance with the Manning and the modified Rome criteria and a new simple definition, was analysed. RESULTS: In the ASQ the agreement on the IBS status for the three IBS definitions was > or = 88%, and in the BDQ > or = 93%. Agreement for the three definitions on the two questionnaires was > or = 88% regardless of which definition of IBS was applied. Agreement between questionnaires was similar (88%) for dyspepsia. For both IBS and dyspepsia the kappa coefficients indicated moderate to substantial concordance. 'Pain or discomfort' did not cover all linguistic aspects of dyspepsia. Prevalence rates of dyspepsia were comparable in the ASQ and BSQ, whereas higher prevalences of IBS with the ASQ was related to the cutoff levels for reporting abdominal pain or discomfort. CONCLUSION: It is possible to identify IBS more simply by self-report questionnaires. PMID- 10720112 TI - Upper gastrointestinal symptoms in a U.S. national sample of adults with diabetes. AB - BACKGROUND: Individuals with diabetes may be particularly susceptible to motility related upper gastrointestinal (UGI) symptoms such as abdominal pain or discomfort, bloating, early satiety, nausea, and vomiting. We estimated the prevalence of UGI symptoms in a population-based sample of individuals with diabetes and determined whether cases and population controls differed in prevalence of UGI symptoms and in symptom features. METHODS: Individuals with diabetes (n = 483) and matched controls (n = 422) were recruited from a prior U.S. national health survey for a telephone interview on UGI symptoms. To confirm self-reported diabetes status, cases provided information on clinical management measures. Subjects were asked about UGI symptoms in the month before interview. Affirmative responses to initial questions triggered detailed questions about symptom frequency, timing, duration, and severity. Differences between cases and controls were evaluated. RESULTS: Cases not only had a significantly (P < 0.05) higher overall prevalence of one or more UGI symptoms in the past month (50%) than controls (38%), but they also reported a significantly greater number of UGI symptoms than controls. Almost 10% of cases reported three or more UGI symptoms in the past month compared with 2% of controls. Our study also identified UGI symptom features that were more relevant to cases and showed that one UGI symptom, heartburn, co-occurred significantly more often with UGI symptoms in cases than in controls. CONCLUSIONS: Upper GI symptoms are common in individuals with diabetes and more prevalent than in controls. The symptoms are non-specific and may reflect disruptions in motility or perception. PMID- 10720113 TI - Effects of oral Saccharomyces boulardii on bacterial overgrowth, translocation, and intestinal adaptation after small-bowel resection in rats. AB - BACKGROUND: Small-bowel resection in animals results in alterations of the morphology and functional adaptation in the remaining intestine. The aim of our study was to study the effect of Saccharomyces boulardii versus placebo in rats after 50% small-bowel resection. METHODS: Sixty-three rats were assigned to one of three groups: small-bowel resection (n = 31), transected surgery controls (n = 16), or non-surgical controls (n = 16). Of the 31 rats with small-bowel resection, 15 were given S. boulardii (140 mg/dl), and 16 were given placebo. Intestinal markers measured included bacterial overgrowth (BO) on days 4 and 8 and translocation into mesenteric lymph nodes, liver, and spleen. Markers of small-bowel adaptation included histomorphology of the mucosa, protein content, and various brush-border enzymes (sucrase, glucoamylase, n-aminopeptidase). RESULTS: In the jejunal mucosal samples on day 8, S. boulardii-treated rats showed a significant increase in protein content (58.3 +/- 12 mg/10 cm) compared with placebo-treated rats (29.2 +/- 1.8) or non-surgery controls (18.3 +/- 1.2; P < 0.001). S. boulardii-treated rats also had significantly higher levels of all three brush-border enzymes. A significant increase of enzyme-specific activities was observed in the ileum of S. boulardii resected rats compared with the placebo resected group on day 4, and no significant differences were seen in the remnant ileum except an increase in protein content in S. boulardii-treated rats on day 8. Histomorphometric studies showed no differences in ileal villus height or translocation frequencies by day 8 in S. boulardii or placebo resected rats. CONCLUSIONS: These data indicate that, after resection, S. boulardii does not modify bacterial overgrowth or translocation frequency but does significantly enhance the functional adaptation of the remaining intestinal segments. PMID- 10720114 TI - Role of circulating peptide YY in the inhibition of gastric acid secretion by dietary fat in humans. AB - BACKGROUND: Intestinal fat inhibits gastric acid secretion and induces release of peptide YY (PYY) into the circulation. The aim of this study was to further establish the role of circulating PYY in the inhibition of gastric acid secretion by intraduodenal fat. METHODS: Plasma PYY concentrations and gastrin-stimulated gastric acid output were measured in response to intravenous infusion of PYY in eight healthy men. The results were compared with those obtained after intraduodenal administration of dietary fat. RESULTS: Plasma PYY concentrations increased by 8.1 +/- 1.8 pmol/l (P < 0.005) in response to the lower and by 13.5 +/- 2.5 pmol/l (P < 0.005) in response to the higher PYY dose. These increments were comparable to those observed after intraduodenal fat (10.3 +/- 2.4 pmol/l). Intraduodenal fat significantly inhibited (P < 0.005 versus control) gastrin stimulated gastric acid secretion by 74% +/- 6%, but neither the lower (3% +/- 7%; NS) nor the higher PYY dose (1% +/- 10%; NS) induced any change in gastric acid output. PYY was biologically active, as reflected by a significant delay (P = 0.04) of orocaecal transit time. CONCLUSION: Release of PYY into the circulation is not responsible for inhibition of gastrin-stimulated gastric acid secretion by dietary fat. PMID- 10720115 TI - Calcaneal ultrasound attenuation and vitamin-D-receptor genotypes in celiac disease. AB - BACKGROUND: Osteopenia is common in patients with celiac disease and is believed to result from malnutrition. Osteoporosis in otherwise healthy individuals is related to genetically determined polymorphisms within the vitamin-D-receptor (VDR) gene. We hypothesized that in celiac patients particular genes of the VDR enhance the susceptibility for malnutrition-associated low-bone density. METHODS: We determined allelic frequencies within the VDR gene by restriction fragment length polymorphism analysis in 92 patients with celiac disease (age, 15-83 years). Thirty-eight patients were on a gluten-free diet; 54 patients did not adhere to a diet. The determined VDR polymorphisms in 111 unrelated newborns served as controls. Osteopenia was determined by means of ultrasound measurements of the calcaneus (n = 78). Bone turnover was estimated by osteocalcin determination (n = 60). RESULTS: There was no difference in the frequency of the VDR gene polymorphisms in patients with celiac disease compared with controls. Adjusted ultrasound measures of the calcaneus were low in 47% of patients, but there was no difference of the VDR gene frequencies in these patients compared with those with normal ultrasound results or controls. Bone turnover was higher in patients without a gluten-free diet (P = 0.02). Again, there was no association with any particular VDR gene. CONCLUSIONS: Patients with celiac disease frequently have osteopenia, which is not related to any of the determined genes within the VDR. PMID- 10720116 TI - Coeliac disease and Down syndrome: associations not due to genetic linkage on chromosome 21. AB - BACKGROUND: Individuals with Down syndrome have an increased prevalence of coeliac disease (CD). The HLA region accounts for only 30% of the heritability of CD, and segregation analyses have suggested the involvement of at least one other non-HLA gene. Distribution of known HLA susceptibility types in Down syndrome and normal populations are similar and do not explain the difference in disease frequency. This study tests the hypothesis that the association between these disorders is due to a susceptibility gene for coeliac disease being present on chromosome 21. METHODS: We studied 21 families multiply affected with CD, none of whom had Down syndrome. The typing information of six microsatellite markers across chromosome 21 was used to test linkage. RESULTS: Negative results from lod score and model-free linkage analysis were obtained, providing no support for genetic linkage of coeliac disease to chromosome 21 in this population. CONCLUSIONS: The high prevalence of coeliac disease in Down syndrome is not due to an increased copy number of a polymorphic susceptibility gene on chromosome 21. PMID- 10720117 TI - Reliance on serum endomysial antibody testing underestimates the true prevalence of coeliac disease by one fifth. AB - BACKGROUND: Although IgA endomysial antibody (EmA) is currently the serologic test of choice in selecting suspected coeliac patients for duodenal biopsies, false-negative cases have been reported and may be more common than previous studies suggest. We assessed the sensitivity of EmA for patients with biopsy confirmed villous atrophy (VA). METHODS: We studied 89 patients without IgA deficiency for whom biopsy had not been primarily prompted by a positive EmA result. VA was graded as partial, subtotal, or total (PVA, STVA, TVA). Serum EmA was assayed with indirect immunofluorescence. RESULTS: The sensitivity of EmA for VA was 78% (69 of 89) and was similar for PVA (79%) and ST/TVA (77%). Only 4 of the 20 EmA-negative patients had increased serum IgA-class antigliadin antibody levels as measured with enzyme-linked immunosorbent assay. All seronegative patients who complied with dietary gluten exclusion responded clinically, with histologic improvement after 12 months in 8 (67%) of 12 patients who had follow up biopsies. CONCLUSIONS: EmA-negative coeliac disease is common. Reliance on EmA testing to select patients for biopsy will result in significant underdiagnosis. PMID- 10720118 TI - Local short-chain fatty acids supplementation without beneficial effect on inflammation in excluded rectum. AB - BACKGROUND: Rectal instillation of short-chain fatty acids (SCFA), important nutrients for the colorectal mucosa, has been suggested to be of therapeutic value in distal intestinal inflammation. METHODS: In this study nine patients with Hartmann-closed rectum after colectomy for acute colitis were investigated. In a double-blind crossover trial an enema containing SCFA or a placebo solution was administered twice daily for 3 weeks. Before entry into the protocol, after each treatment period, and 6 weeks after the study period the patients' symptoms were evaluated, rectal endoscopy was performed, histologic samples were scored, and microbiologic analyses were carried out. RESULTS: No significant differences in symptoms, in mucosal inflammation, in histologic scoring, or in microbiologic studies were found between SCFA and placebo periods. Unexpectedly, all but one patient entirely lacked coliform bacteria in the rectum. CONCLUSIONS: In this study SCFA enemas had no beneficial effect on inflammation in excluded rectum in patients earlier submitted to colectomy for colitis. However, a different rectal flora was detected in these patients. PMID- 10720119 TI - Plasma exudation, hyperaemia, and epithelial permeability in rats with oxazolone induced colitis: modulatory effects of budesonide. AB - BACKGROUND: Oxazolone-induced colitis in the rat is an immune-driven model of human colitis. The aim of the present study was to measure the changes in the absorptive and exudative permeabilites, oedema formation, and local blood flow in this model during the development of inflammation. We also assessed the effects of acute (<1 h), topical glucocorticosteroid (GCS) treatment on these factors. METHODS: Colitis was induced by local instillation of oxazolone in previously sensitized animals. Calculating the 40-min plasma-equivalent extravascular volume quantitated the plasma exudation rate. This was determined by using labelled albumin as marker for total tissue content of plasma and Evans blue content as marker for the intravascular volume. Absorptive permeability was simultaneously measured as uptake of rectally administered (51Cr)-labelled ethylenediaminetetraacetic acid (EDTA). In separate experiments regional blood flows were measured by means of the labelled microsphere method. RESULTS: At both 3 and 24 h after challenge marked enhancements of both exudative and absorptive permeabilities were found. At 24 h there was also an increase in local blood flow. GCS treatment abolished all of the hyperaemia and the main part of the exudative response but had no significant effect on the absorptive permeability. CONCLUSIONS: In this model immunologic mechanisms induce permeability and blood flow changes similar to those in the human disease. It seems suitable for the study of GCS and other anti-inflammatory or immune-modulating drugs. PMID- 10720120 TI - Brown pigment stones in the common bile duct: reduced bilirubinate diconjugate in bile. AB - BACKGROUND: Bilirubin is the main component of most common bile duct stones. Normally, almost all bilirubin in bile is conjugated to glucuronic acid or some other sugar moiety. These conjugates are unstable and liable to deconjugation. Unconjugated bilirubin is insoluble and may precipitate as the calcium salt found in brown pigment stones. The pattern of bilirubin conjugates in common duct bile of patients with choledocholithiasis has been unknown. METHODS: In a clinical series of 55 patients with choledocholithiasis common-duct bile was aspirated, and the bilirubin conjugates analyzed with high-performance liquid chromatography. One stone from each patient was analyzed for cholesterol and bilirubin content to determine stone type. RESULTS: Sixteen patients had cholesterol stones, 38 patients had brown pigment stones, and 1 patient had a black stone. Patients with pigment stones had a lower percentage of bilirubin diglucuronide (median, 60.3%; interquartile range, 49.7%-67.3%) than patients with cholesterol stones (64.0%; 60.2%-73.3%) (Mann-Whitney, P=0.015). No significant difference was found for the other bilirubin conjugates, total bilirubin, or biliary pH when pigment and cholesterol stone patients were compared. The time of bile sampling in relation to papillotomy and treatment of cholestasis was not associated with the low percentage of bilirubin diglucuronide. The observation of reduced values for bilirubin diglucuronide could not be ascribed to duodenal diverticula or Billroth-II gastric resection. CONCLUSION: The percentage of the main bilirubinate conjugate, bilirubin diglucuronide, is decreased in the common duct bile of patients with pigmented compared with cholesterol stones. PMID- 10720121 TI - Cephalic phase of lipolysis is impaired in pancreatic insufficiency: role of gastric lipase. AB - BACKGROUND: Gastric lipase contributes significantly to overall lipolysis and is regulated by interacting neuro-hormonal mechanisms. Patients with alcoholic chronic pancreatitis (ACP) have low, or even absent, activity of pancreatic lipases. In that state the secretion of gastric lipase could be essential and compensate for the pancreatic defect. However, conflicting studies have not resolved the order of magnitude of gastric lipase secretion in these patients. This could be explained by differences in regulatory mechanisms, gastric mucosal changes, and abdominal vagal tone. METHODS: Nasogastric intubation with modified sham feeding and upper endoscopy including biopsies for histologic classification and Helicobacter pylori infection status were performed in eight ACP patients, and eight healthy volunteers were studied on separate occasions. Vagal nerve function was assessed by calculation of heart rate variability in ACP patients. Gastric lipase was measured in aspirates by means of enzyme-linked immunosorbent assay and an enzyme kinetic assay. Plasma concentrations of gastrin, secretin, cholecystokinin, and pancreatic polypeptide were measured throughout the study. RESULTS: Sham feeding rapidly and significantly increased gastric lipase secretion in healthy volunteers, whereas ACP patients did not respond to sham feeding. Two of eight patients were infected with H. pylori and had mucosal changes accordingly. The lack of lipase response could not be ascribed to dysfunction of the abdominal vagus. CONCLUSIONS: The cephalic phase of gastric lipase secretion is impaired in ACP patients. Although their fundic cells continue to secrete gastric lipase, they are not subject to normal neuro-hormonal regulation. PMID- 10720122 TI - The impact of storage time of transfused blood on postoperative infectious complications in rectal cancer surgery. Danish RANX05 Colorectal Cancer Study Group. AB - BACKGROUND: We have studied the impact of storage time of transfused allogeneic blood together with other known risk factors on postoperative infectious complications after operation for rectal cancer. METHODS: Intra-abdominal abscess, anastomotic leakage, septicaemia, wound infection, and pneumonia were prospectively recorded in 303 patients undergoing elective resection for primary rectal cancer in 18 Danish hospitals. Patient risk variables and variables related to operation technique and transfusion were recorded prospectively, whereas amount given before infectious complication and storage time of saline adenine-glucose-mannitol (SAGM) blood, administered to each patient, were recorded retrospectively. RESULTS: The overall infection rate was 24% in 78 non transfused and 40% in 225 transfused patients (P = 0.011). The proportion of SAGM blood stored for > or = 21 days administered to each transfused patient was a median of 60% in patients developing postoperative infections versus 25% (P = 0.037) in patients without infections. A multivariate analysis of significant risk variables showed weight > 75 kg (odds ratio, 2.0 versus < 65 kg) and transfusion of SAGM blood stored > or = 21 days (odds ratio, 2.5 versus no transfusion) to be independent variables predicting infectious complications. CONCLUSION: Blood storage time may, along with other risk factors, play a significant role in blood transfusion-associated development of postoperative infectious complications. PMID- 10720123 TI - Preoperative diagnosis of gallbladder torsion by magnetic resonance cholangiopancreatography. AB - The patient was a 78-year-old woman who was diagnosed as having gallbladder torsion preoperatively. This is the first reported case diagnosed by magnetic resonance cholangiopancreatography (MRCP). Signs and symptoms of this condition are often subtle. Radiologic evaluation by ultrasonography and computed tomography (CT) showed acute cholecystitis with stone. Drip-infusion cholangiography CT failed to outline the gallbladder, and distortion of the extrahepatic bile ducts and interruption of the cystic duct were observed. MRCP showed 1) a v-shaped distortion of the extrahepatic bile ducts due to traction by the cystic duct, 2) tapering and twisting interruption of the cystic duct, 3) a distended and enlarged gallbladder that was deviated to the midline of the abdomen, and 4) a difference in intensity between the gallbladder and the extrahepatic bile ducts and the cystic duct. A definitive diagnosis of gallbladder torsion (volvulus) was made by MRCP preoperatively. If treated surgically, gallbladder detorsion before cholecystectomy is a helpful technique to avoid bile duct injury. This condition should be suspected in elderly women with acute cholecystitis or acute abdominal pain of unknown origin, and MRCP may be very useful in making a definitive diagnosis. PMID- 10720124 TI - A simple method to evaluate the thickness of collagen in collagenous colitis. PMID- 10720125 TI - Antisense therapeutics in chronic myeloid leukaemia: the promise, the progress and the problems. AB - DNA sequences which are complementary or 'antisense' to a target mRNA can inhibit expression of that mRNA's protein product. Antisense therapeutics has therefore received attention for inhibiting oncogenes in haematological malignancy, in particular in chronic myeloid leukaemia. However, it is now becoming clear that antisense therapeutics is considerably more problematic than was naively initially assumed. In this article, some of these difficulties are discussed, together with the achievements in CML so far. Considerable further research is required in order to define an optimal antisense therapeutics strategy for clinical use. PMID- 10720126 TI - Benefit of intensified treatment for all children with acute lymphoblastic leukaemia: results from MRC UKALL XI and MRC ALL97 randomised trials. UK Medical Research Council's Working Party on Childhood Leukaemia. AB - Treatment of children with acute lymphoblastic leukaemia (ALL) aims to cure all patients with as little toxicity as possible and, if possible, to restrict further intensification of chemotherapy to patients with an increased risk of relapse. However in Medical Research Council (MRC) trial UKALL X two short myeloablative blocks of intensification therapy given at weeks 5 and 20 were of benefit to children in all risk groups. The successor trials, MRC UKALL XI and MRC ALL97, tested whether further intensification would continue to benefit all patients by randomising them to receive, or not, an extended third intensification block at week 35. After a median follow-up of 4 years (range 5 months to 8 years), 5 year projected event-free survival was superior at 68% for the 894 patients allocated a third intensification compared with 60% for the 887 patients who did not receive one (odds ratio 0.75, 95% CI 0.63-0.90, 2P = 0.002). This difference was almost entirely due to a reduced incidence of bone marrow relapses in the third intensification arm (140 of 891 in the third intensification arm vs. 171 of 883 in the no third intensification, 2P = 0.02). Subgroup analysis suggests benefit of the third intensification for all risk categories. Overall survival to date is no different in the two arms, indicating that a greater proportion of those not receiving a third intensification arm and subsequently relapsing can be salvaged. These results indicate that there is benefit of additional intensification for all risk subgroups of childhood ALL. PMID- 10720127 TI - Use of dual-color interphase FISH for the detection of inv(16) in acute myeloid leukemia at diagnosis, relapse and during follow-up: a study of 23 patients. AB - The value of dual-color fluorescence in situ hybridization (FISH) for the detection of inv(16), using two contigs of cosmid probes mapping on both sides of the chromosome 16p breakpoint region, was evaluated in 23 acute myeloid leukemias (AML) in different phases of the disease. At diagnosis interphase FISH detected inv(16) in 19/19 (100%) cases with conventional cytogenetics (CC) evident aberration and excluded the rearrangement in two patients with CC suspected inv(16). Moreover, it also identified an associated del(16p) in two patients. At relapse, it revealed the inv(16) in 8/8 (100%) studied cases. These results were concordant with those of reverse transcriptase-polymerase chain reaction (RT PCR). From 13 patients who obtained at least one complete remission (CR), 31 follow-up samples were analyzed using interphase FISH. Twenty-nine specimens scored negative for inv(16) and two were positive. RT-PCR detected CBFbeta/MYH11 transcripts in four of the nine CR samples analyzed, being more sensitive than interphase FISH. Eight of the 13 patients relapsed at a median time of 6.5 months (range 1-15) from the last negative FISH analysis. Of the two patients with positive FISH in CR, one relapsed soon after. At diagnosis and relapse, interphase-FISH proved to be an effective technique for detecting inv(16) appearing more sensitive than CC. Prospective studies with more frequent controls and possibly additional FISH probes are needed to assess the value of interphase FISH for minimal residual disease (MRD) and relapse prediction. PMID- 10720128 TI - Effects of cranial radiation in children with high risk T cell acute lymphoblastic leukemia: a Pediatric Oncology Group report. AB - Contemporary chemotherapy has significantly improved event-free survival among patients with T cell-lineage acute lymphoblastic leukemia (T-ALL). Unlike B precursor ALL, most investigators are still using cranial radiation (CRT) and are hesitant to rely solely on intrathecal therapy for T-ALL. In this study we assessed the effects of CRT upon event-free survival and central nervous system (CNS) relapses in a cohort of children with high risk features of T cell leukemia. In a series of six consecutive studies (1987-1995) patients were non randomly assigned their CNS prophylaxis per individual protocol. These protocols were based on POG 8704 which relied on rotating drug combinations (cytarabine/cyclophosphamide, teniposide/Ara-C, and vincristine/doxorubicin/6 MP/prednisone) postinduction. Modifications such as high-dose cytarabine, intermediate-dose methotrexate, and the addition of G-CSF, were designed to give higher CNS drug levels (decreasing the need for CRT), to eliminate epidophyllotoxin (decreasing the risk of secondary leukemia), and to reduce therapy-related neutropenia (pilot studies POG 9086, 9295, 9296, 9297, 9398). All patients included in this analysis qualified for POG high risk criteria, WBC >50000/mm3 and/or CNS leukemia. Patients without CNS involvement received 16 doses of age-adjusted triple intra-thecal therapy (TIT = hydrocortisone, MTX, and cytarabine) whereas patients with CNS disease received three more doses of TIT during induction and consolidation. Patients who received CRT were treated with 2400 cGy (POG 8704) or 1800 cGy (POG 9086 and 9295). CNS therapy included CRT in 144 patients while the remaining 78 patients received no radiation by original protocol design. There were 155 males and 57 females with a median age of 8.2 years. The median WBC for the CRT+ and CRT- patients were 186000/mm3 and 200000/mm3, respectively. CNS involvement at diagnosis was seen in 16% of the CRT+ and 23% of the CRT- groups. The complete continuous remission rate (CCR) was not significantly different for the irradiated vs. non-irradiated groups (P = 0.46). The 3-year event-free survival was 65% (s.e. 6%) and 63% (s.e. 4%) for the non-irradiated vs. the radiated group. However, the 3-year CNS relapse rate was significantly higher amongst patients who did not receive CRT; 18% (s.e. 5%) vs. 7% (s.e. 3%) in the irradiated group (P = 0.012). Our analysis in a non randomized setting, suggests that CRT did not significantly correlate with event free survival but omitting it had an adverse effect on the CNS involvement at the time of relapse. PMID- 10720129 TI - In vivo treatment of mutant FLT3-transformed murine leukemia with a tyrosine kinase inhibitor. AB - Somatic mutation of the FLT3 gene, in which the juxtamembrane domain has an internal tandem duplication, is found in 20% of human acute myeloid leukemias and causes constitutive tyrosine phosphorylation of the products. In this study, we observed that the transfection of mutant FLT3 gene into an IL3-dependent murine cell line, 32D, abrogated the IL3-dependency. Subcutaneous injection of the transformed 32D cells caused leukemia in addition to subcutaneous tumors in C3H/HeJ mice. To develop a FLT3-targeted therapy, we examined tyrosine kinase inhibitors for in vitro growth suppression of the transformed 32D cells. A tyrosine kinase inhibitor, herbimycin A, remarkably inhibited the growth of the transformed 32D cells at 0.1 microM, at which concentration it was ineffective in parental 32D cells. Herbimycin A suppressed the constitutive tyrosine phosphorylation of the mutant FLT3 but not the phosphorylation of the ligand stimulated wild-type FLT3. In mice transplanted with the transformed 32D cells, the administration of herbimycin A prolonged the latency of disease or completely prevented leukemia, depending on the number of cells inoculated and schedule of drug administration. These results suggest that mutant FLT3 is a promising target for tyrosine kinase inhibitors in the treatment of leukemia. PMID- 10720130 TI - In vitro cytotoxic effects of fludarabine (2-F-ara-A) in combination with commonly used antileukemic agents by isobologram analysis. AB - Fludarabine phosphate (2-F-ara-AMP) is an adenine nucleoside analogue that shows significant activity against chronic lymphocytic leukemia and indolent lymphoma. We assessed the cytotoxic interaction produced by the combination of the active metabolite of fludarabine phosphate, fludarabine (9-beta-D-arabinofuranosyl-2 fluoroadenine, 2-F-ara-A), and some commonly used antileukemic agents against human hairy cell leukemia cell line JOK-1, human chronic lymphocytic leukemia cell line SKW-3, and adult T cell leukemia cell lines ED-40810 (-) and SALT-3. The leukemia cells were exposed simultaneously to 2-F-ara-A and to the other agents for 4 days. Cell growth inhibition was determined using MTT reduction assay. The isobologram method of Steel and Peckham was used to evaluate the cytotoxic interaction. 2-F-ara-A and cytarabine showed synergistic effects in SKW 3 cells, additive and synergistic effects in JOK-1 and SALT-3 cells, and additive effects in ED-40810(-) cells. 2-F-ara-A and doxorubicin showed additive effects in SKW-3, ED-40810(-) and SALT-3 cell lines, and additive and synergistic effects in JOK-1 cells. 2-F-ara-A showed additive effects with etoposide, 4-hydroperoxy cyclophosphamide, and hydroxyurea in all four cell lines. 2-F-ara-A showed antagonistic effects with methotrexate and vincristine in all four cell lines. Our findings suggest that the simultaneous administration of fludarabine phosphate with cytarabine, doxorubicin, etoposide, cyclophosphamide, or hydroxyurea would be advantageous for cytotoxic effects. Among these agents, cytarabine may be the best agent for the combination with fludarabine phosphate. The simultaneous administration of fludarabine phosphate with methotrexate or vincristine would have little cytotoxic effect, and this combination may be inappropriate. These findings may be useful in clinical trials of combination chemotherapy with fludarabine phosphate and these agents. PMID- 10720131 TI - Interferon alfa as primary treatment of chronic myeloid leukemia: long-term follow-up of 71 patients observed in a single center. AB - The purpose of this study was to evaluate the long-term outcome of interferon (IFN) alfa treatment in patients with Philadelphia chromosome-positive chronic myeloid leukemia (CML). Between 1984 and 1990, a total of 71 patients with newly diagnosed CML had been enrolled into two consecutive IFN trials at our institution. Follow-up extended to December 1998, resulting in a median observation period for surviving patients of 11.4 years. The median survival time from diagnosis was 5.9 years. A plateau in the actuarial survival curve was found from 8.2 to 12.3 years following diagnosis with a projected 10-year survival rate of 32%. 'Landmark' studies showed a significant survival advantage for patients with karyotype responses. Of 68 patients accessible to calculation of the Hasford score, three were in the high risk group, 24 belonged to the medium risk group, and 41 had low risk features. The majority of cytogenetic responders including all eight assessable patients in complete cytogenetic remission were in the low risk group. Achieving a cytogenetic remission was found to provide a survival advantage also for patients with low risk disease. Of the seven patients surviving more than 11 years, six were in continuous complete cytogenetic remission. Their favorable outcome appears to translate into an out-flattening of the survival curve for the 71 single center patients presented. It will be of interest to see whether prolonged follow-ups of the large multicentric randomized trials will similarly show a subset of long-term surviving patients with ongoing IFN-induced remission. PMID- 10720133 TI - Transcription factor NF-kappaB is constitutively activated in acute lymphoblastic leukemia cells. AB - The pleiotropic transcription factor NF-kappaB controls cellular apoptotic and growth processes and increasing evidence suggests a role in tumorigenesis. We describe here that constitutively activated NF-kappaB complexes are found in the vast majority (39 out of 42 samples) of childhood acute lymphoblastic leukemia (ALL) without any subtype restriction. Electrophoretic shift analysis further demonstrates that these complexes are composed of p50-p50 and p65-p50 dimers. Proteasome inhibition in primary ALL cultures results in a hyperphosphorylated form of IkappaBalpha, indicating that activation of upstream kinases, which trigger IkappaBalpha degradation, has led to nuclear translocation of NF-kappaB. Careful inhibition of cellular proteolytic activities is of importance when analyzing extracts from primary ALL cells. Degradation of p65 and other proteins in ALL samples could be specifically suppressed by alpha-1 antitrypsin. Constitutive NF-kappaB activation is thus a common characteristic of childhood ALL and strongly suggests a critical role of this factor for leukemia cell survival. PMID- 10720132 TI - HLA class I in acute promyelocytic leukemia (APL): possible correlation with clinical outcome. AB - The majority of patients with acute promyelocytic leukemia (APL) possess either a bcr1 or a bcr3 type fusion between PML and RARalpha genes. The junction sequences may possibly be a target for immune response and influence susceptibility to the disease. In this case, HLA class I allele frequencies would be different between bcr1 and bcr3 patients. To test this hypothesis, we typed 102 APL patients for HLA-A, -B and -Cw alleles. The A*1, A*30, B*51, B*41, Cw*0602, and Cw*1701 alleles showed a different distribution between bcr1 and bcr3 patients, but in no case was this statistically significant after correction for the number of comparisons or was confirmed in an independent panel. Moreover, no difference was detected between bcr1 and bcr3 when HLA alleles were grouped according to their peptide binding specificities. Comparing HLA frequencies, clinical features at diagnosis and clinical outcome of the 64 patients homogeneously treated with all trans retinoic acid and idarubicin (AIDA protocol) we observed a statistically significant association between HLA-B*13 and risk of relapse by univariate and multivariate regression analysis. Should this finding be confirmed in larger future studies, this observation would be of outmost importance in identifying patients at high risk of relapse in which more aggressive consolidation therapies should be used. PMID- 10720134 TI - CD40 ligand upregulates expression of the IL-3 receptor and stimulates proliferation of B-lineage acute lymphoblastic leukemia cells in the presence of IL-3. AB - The proliferative response of B cell precursor acute lymphoblastic leukemia (BCP ALL) cells to IL-3 is dependent on the expression of functional IL-3 receptors (IL-3R). Here we report that CD40 ligand (CD40L) in the presence of recombinant IL-3 increased proliferation of BCP-ALL cells by upregulating expression of IL 3R. Upregulation of IL-3R in BCP-ALL cells was observed as early as 1 h after treatment with CD40L, and a 50- to 500-fold increase of IL-3R expression after 24 h was detected in all 12 cases studied. Moreover, expression of receptors for IL 7 (IL-7R) and stem cell factor (SCF-R, c-Kit) was also induced by CD40L in the majority of BCP-ALL cases examined; however, levels of induction were low compared to those for IL-3R. To test the functional activity of upregulated receptors for IL-3, SCF and IL-7, we evaluated the proliferation and growth of BCP-ALL cells cultured in serum-free media with CD40L plus these factors. When CD40L was added with either a single cytokine (IL-3, SCF and IL-7) or their combinations, cell proliferation was significantly increased as detected by DNA synthesis assay. Combinations of CD40L plus IL-3 and either SCF or IL-7 were able to support long-term growth of BCP-ALL cells for at least 8 weeks in three of the seven cases studied. Immunophenotyping and gene rearrangement studies indicated that cells in long-term cultures were monoclonal and retained their original phenotypes. The leukemic cells remained primarily dependent on the presence of IL 3 and its receptor for long-term growth, as shown by selective withdrawal of growth factors or antibody blockade of receptors. These results suggest an important role for CD40L in upregulating expression of IL-3R on BCP-ALL cells and enabling these cells to proliferate in long-term cultures in the presence of IL-3 and either SCF or IL-7. PMID- 10720135 TI - Cytokine upregulation of the antigen presenting function of acute myeloid leukemia cells. AB - Acute myeloid leukemia (AML) cells are malignant counterparts of normal myeloid pathway progenitors. Myeloid progenitors differentiate into professional antigen presenting cells (APC) under the essential influence of GM-CSF along with additional cytokines. Twelve cases of human AML were tested for ability to be differentiated toward a professional APC phenotype in short-term culture with addition of GM-CSF and the following recombinant proteins: TNFalpha, IL-4, CD40 ligand, Flt3 ligand and SCF. Significant upregulation of CD80 (B7-1) and enhancement of alloantigen presentation was seen with the addition of GM-CSF and TNFalpha alone or with additional cytokines. The combination of GM-CSF and TNFalpha, either alone or in combination with an additional cytokine, resulted in enhancing alloantigen presentation by at least two-fold over the media control group in 10/12 patients studied, and resulted in CD80 expression of greater than 15% in 11/12 patients studied. In AML cultures with GM-CSF and TNFalpha, coexpression of CD80 and either CD34 or an aberrant surface marker (CD56) was seen. In one case, sorted CD80, cells retained a characteristic cytogenetic marker and CD34 expression, proving their derivation from an AML precursor. These studies verify other reports of in vitro differentiation of human AML precursors into enhanced APC, suggesting that this phenomenon could be utilized for immunotherapy strategies aimed at enhancing presentation of leukemia antigens to T cells. PMID- 10720136 TI - Peptides spanning the junctional region of both the abl/bcr and the bcr/abl fusion proteins bind common HLA class I molecules. AB - The Philadelphia (Ph) chromosome, resulting from the t(9;22) translocation, is characteristic of chronic myeloid leukemia (CML). As a result of this translocation, two novel chimeric genes are generated and the bcr/abl and abl/bcr fusion proteins expressed. The bcr/abl fusion mRNA is present in all CML patients, whereas the reciprocal abl/bcr fusion mRNA is detectable in about 80% of the Ph+ CML patients. These fusion proteins may undergo enzymatic degradation in the cytosol and give rise to MHC class I restricted peptide epitopes originating from the junctional regions of the translocation products, which thus may serve as novel tumor specific antigens. Previously, other groups have tested peptides corresponding to the junctional region of the bcr/abl protein for their binding capacity to HLA class I molecules and have identified a few candidate epitopes. Peptides originating from the abl/bcr fusion protein have on the other hand so far been neglected, for no apparent reason. We have now extended these studies to include also the reciprocal abl/bcr translocation product by testing a large panel of synthetic peptides corresponding to the junctional regions of both the abl/bcr and the bcr/abl fusion proteins for their ability to stabilize HLA class I molecules. We find that the abl/bcr translocation product may be an even more important source of CML specific peptide antigens and together the junctional sequences of both these proteins contain peptide sequences which bind efficiently to a number of HLA molecules (HLA-A1, -A2, -A3, -A11, -B7, -B27, B35) and thus may serve as candidate CML specific tumor antigens. PMID- 10720137 TI - Frequent deletions at 11q23 and 13q14 in B cell prolymphocytic leukemia (B-PLL). AB - Deletions of the long arm of chromosomes 11 and 13 are the most frequent structural chromosome aberrations in various types of lymphoproliferative disorders. However, these regions have not been studied so far in B cell prolymphocytic leukemia (B-PLL). We have investigated the incidence of 13q deletions in 18 B-PLL cases by fluorescence in situ hybridization (FISH), using molecular probes for the RB1 and D13S25 loci. Chromosome 11q deletions were evaluated by FISH using the yeast artificial chromosome (YAC) clone 755b11 from the chromosome 11q22.3-q23.1 region, which has been previously shown to be deleted in 20% of cases of chronic lymphocytic leukemia. Chromosome 11q23 deletions were found in 7/18 (39%) cases of B-PLL. Monoallelic loss of RB1, D13S25 and BRCA2 was present in 10/18 (55%), 6/18 (33%) and 3/18 (16%) of the cases, respectively. All the cases with D13S25 and BRCA2 deletion showed RB1 loss. Deletions of 13q14 and 11q23 are frequent chromosome aberrations in B-PLL and, in contrast to CLL, there is a preferential loss of RB1 with respect to the D13S25 locus suggesting that allelic loss of the RB1 gene may play a role in the pathogenesis of B-PLL. PMID- 10720138 TI - Arsenic trioxide (As2O3)-induced apoptosis and differentiation in retinoic acid resistant acute promyelocytic leukemia model in hGM-CSF-producing transgenic SCID mice. AB - Recent clinical studies in China and USA showed that arsenic trioxide (As2O3) is an effective treatment of acute promyelocytic leukemia (APL) patients refractory to all-trans retinoic acid (RA). We here investigate the effects of As2O3 on RA resistant APL in vivo and in vitro using our RA-resistant APL model system. As2O3 can induce inhibition of cellular growth of both RA-sensitive NB4 and RA resistant UF-1 APL cells via induction of apoptosis in vitro. The expression of BCL-2 protein decreased in a dose- and time-dependent manner in NB4 cells. Interestingly, the levels of BCL-2 protein were not modulated by As2O3, but it did upregulate BAX protein in UF-1 cells. UF-1 cells (1x10(7)) were transplanted into hGM-CSF-producing transgenic SCID mice and successfully formed subcutaneous tumors. After 40 days of implantation, mice were treated with As2O3, all-trans RA and PBS for 21 days. In all-trans RA- and PBS-treated mice, tumors grew rapidly, with a 4.5-fold increase in volume at day 21 compared to the initial size. In marked contrast, tumor size was decreased to half of the initial size by the treatment of As2O3, which resulted in cells with the typical appearance of apoptosis. Interestingly, one of the As2O3-treated mice showed mature granulocytes in the diminished tumor, suggesting that As2O3 had dual effects on RA-resistant APL cells in vivo: both inducing apoptosis and differentiation of the leukemic cells. We conclude that our RA-resistant APL model will be useful for evaluating novel therapeutic approaches to patients with RA-resistant APL, and for further investigation of the metabolism of As2O3 in vivo. PMID- 10720140 TI - Lack of CD95/FAS gene somatic mutations in extranodal, nodal and splenic marginal zone B cell lymphomas. AB - Germline CD95 (also known as FAS, APT1 and APO1) gene mutations have been associated with benign lymphoproliferative diseases and autoimmune processes. Somatic mutations have been reported in human tumours, including lymphomas. Since marginal zone B cell lymphomas usually arise in a background of chronic inflammation, often of autoimmune origin, we searched for CD95 gene mutations in an unselected series of marginal zone B cell lymphomas. The CD95/FAS full coding region, comprising exon-intron junctions, was amplified from genomic DNA by polymerase chain reaction (PCR) in 10 separate reactions. PCR products were analysed by single-strand conformation polymorphism (SSCP) and visualised by silver staining. Bands exhibiting an altered electrophoretic mobility were sequenced. Twenty-seven cases of marginal zone B cell lymphomas of whom fresh or frozen tumour material was available (18 extranodal, five splenic and four nodal) were studied. Previously described silent polymorphisms in exons 7 (C836T) and 3 (T416C) were detected in 42% and in 19% of the cases, respectively. One silent T to-A substitution at bp 431, within exon 3, was found in one case. Our results did not reveal the presence of CD95 somatic mutations in unselected cases of marginal zone B cell lymphomas. On the basis of our data, we cannot rule out that other genes coding for proteins involved in the CD95-induced apoptotic pathway might be altered. However, this pathway does not seem to play an important role in the pathogenesis of these lymphoma subtypes. PMID- 10720139 TI - FLI-1 is a suppressor of erythroid differentiation in human hematopoietic cells. AB - The FLI-1 oncogene, a member of the ETS family of transcription factors, is associated with both normal and abnormal hematopoietic cell growth and lineage specific differentiation. We have previously shown that overexpression of FLI-1 in pluripotent human hematopoietic cells leads to the induction of a megakaryocytic phenotype. In this report we show that FLI-1 also acts as an inhibitor of erythroid differentiation. Following the induction of erythroid differentiation, pluripotent cells express reduced levels of FLI-1. In contrast, when FLI-1 is overexpressed in these cells, the levels of erythroid markers are reduced. The ability of FLI-1 overexpressing cells to respond to erythroid specific inducers such as hemin and Ara-C is also inhibited, and the uninduced cells show a reduced level of the erythroid-associated GATA-1 transcription factor mRNA. Furthermore, expression of a GATA-1 promoter-driven reporter construct in K562 cells is inhibited by co-transfection with a construct expressing FLI-1. Our results support the hypothesis that FLI-1 can act both positively and negatively in the regulation of hematopoietic cell differentiation, and that inhibition of GATA-1 expression may contribute to FLI-1 mediated inhibition of erythroid differentiation. PMID- 10720141 TI - MUM1/IRF4 expression as a frequent event in mature lymphoid malignancies. AB - MUM1/IRF4 is a myeloma-associated oncogene transcriptionally activated as a result of t(6;14)(p25,q32) chromosomal translocation and by virtue of its juxtaposition to the immunoglobulin heavy chain gene (IgH) locus. When this oncogene becomes non-functional, no activated B/T lymphocytes and Ig secreting plasma cells are observed, suggesting that MUM1/IRF4 is crucial for lymphoid development. Its expression was analyzed in both reactive lymphoid and lymphoma tissues by means of an immunohistochemical technique using specific goat antiserum against MUM1/IRF4. This analysis detected a 50 kDa MUM1 product whose localization was restricted to the nuclei of the lymphocytes. The MUM1+ cells in reactive lymph nodes were found to consist of plasma cells and a small fraction (approximately 7.9%) of B cells harboring CD20+CD38+, which were located in the light zone of the germinal center. MUM1 expression in peripheral blood B/T lymphocytes was upregulated by mitogenic stimuli, suggesting that MUM1 positivity represents the activated state of the B/T cells. In B cell non-Hodgkin's lymphoma (NHL), MUM1 expression was observed in 73.2% (30/41) of diffuse large B cell lymphoma (DLBCL), 20% (1/5) of marginal zone lymphoma (MZL) and 43% (3/7) of small lymphocytic lymphoma (SLL) cases, whereas it was not seen in any cases of mantle cell lymphoma (MCL) or follicle center lymphoma (FCL). Also, MUM1 was stained at high intensity in various types of T cell lymphomas including adult T cell leukemia/lymphoma (ATL/L) and anaplastic large cell lymphoma (ALCL) and in the majority of Hodgkin's diseases. Our results suggest that a major proportion of lymphomas comprise either physiologically or aberrantly activated neoplastic lymphocytes expressing the MUM1 protein. PMID- 10720142 TI - Transplantation of syngenic bone marrow contaminated with NGFr-marked WEHI-3B cells: an improved model of leukemia relapse in mice. AB - With the aim of developing a model mimicking the relapse of patients transplanted with leukemia-contaminated grafts, myelomonocytic leukemia WEHI-3B D+ cells were first transduced with a retroviral vector encoding the low-affinity human nerve growth factor receptor (NGFr). Clones with a stable and homogeneous expression of the transgene and with a similar in vitro behavior to the parental cell line were selected for further experiments. The analysis of bone marrow (BM) contaminated with WEHI-3B/NGFr cells revealed a linear correlation (r2 = 0.999) between the actual values of BM contamination and the experimental data determined by flow cytometry. Balb/c mice were myeloablated and transplanted with syngenic BM contaminated with graded numbers of leukemic cells; dose-dependent survival curves were obtained, regardless of whether parental or WEHI-3B/NGFr cells were infused. The leukemia dissemination in recipients transplanted with WEHI-3B/NGFr contaminated grafts was easily determined by means of simple flow cytometry analysis of the NGFr marker. A leukemia dose-dependent increase in the number of PB leukocytes was observed in transplanted recipients at 20 days post transplantation with no changes in myelomonocytic cells. As deduced from our observations, the transplantation of syngenic BM contaminated with WEHI-3B/NGFr cells constitutes an improved model of autograft-mediated leukemia relapse and a good tool for studies of leukemia cell purging. PMID- 10720143 TI - Novel mechanisms of drug resistance in leukemia. AB - A key issue in the treatment of acute leukemia is the development of resistance to chemotherapeutic drugs. Several mechanisms may account for this phenomenon, including failure of the cell to undergo apoptosis in response to chemotherapy, or failure of the drug to reach and/or affect its intracellular target. This review focuses on the latter mechanism, and on intracellular drug transport resistance mechanisms in particular. Expression of the ATP-binding cassette (ABC) transporter P-glycoprotein (Pgp) has generally been reported to correlate with prognosis in acute myeloid leukemia (AML). Additionally, but more controversial, expression of the ABC transporter multidrug resistance protein (MRP) and the vault-transporter lung resistance protein (LRP) have been correlated with outcome in AML. Despite these findings, functional efflux assays indicate the presence of non-Pgp, non-MRP transporters in AML. Recently, a novel ABC transporter, breast cancer resistance protein (BCRP) was cloned and sequenced in our laboratory. Transfection and overexpression of BCRP in drug-sensitive cells confers drug resistance to the cells. BCRP is a half-transporter, and may homodimerize or form heterodimers (with a yet unknown half-transporter) to produce an active transport complex. Relatively high expression of BCRP mRNA is observed in approximately 30% of AML cases, suggesting a potential role for this new transporter in drug resistance in leukemia. PMID- 10720144 TI - Monoclonal antibodies to the myeloid stem cells: therapeutic implications of CMA 676, a humanized anti-CD33 antibody calicheamicin conjugate. AB - There are several competing models of stem cell involvement in acute myeloid leukemia (AML). At issue is whether the disease origin is in the pluripotent stem cell or whether it arises later in a more mature progenitor cell. The observation that the CD33 antigen is present on AML cells, and on normal and leukemic progenitors, suggested that one might be able to target these cells while sparing the normal stem cells. Response rates of acute myelogenous leukemia patients treated with the newly developed anti-CD33 antibody-calicheamicin conjugate suggest that at least for a proportion of patients early precursors responsible for re-establishing hematopoiesis are likely to be predominantly normal in origin. PMID- 10720145 TI - Treatment of relapsed and refractory acute myelogenous leukemia. AB - Evidence suggests that the salvage therapy utilized for relapsed and refractory acute myelogenous leukemia (AML) should differ based on the duration of a patient's complete remission (CR), the principal predictor of outcome. While standard regimens have produced higher CR rates than investigational regimens, these rates have not translated into improved survival in patients with initial remission durations of <1 year. Accordingly, there is no need to give standard regimens to these patients who rather should receive investigational therapy once relapse is discovered. In contrast, in patients with initial remission durations of 1-2 years, standard regimens do increase survival compared to investigational regimens. A somewhat artificial distinction has been placed between phase I and phase II studies. The agents to be studied in phase II trials are many, but the patients are limited, so we need to be more innovative in our trial designs. One such proposal, utilizing a Bayesian selection design which calls for randomizing a small number of patients among several investigational treatments, will be discussed. PMID- 10720146 TI - Treatment of acute myelogenous leukemia in older adults. AB - The overall strategy for the treatment of older adults is summarized in Table 8. Soon after the birth of effective chemotherapy for acute leukemia, the perspective for all patients was summarized as follows: 'With all humility it may be claimed that there are, at least, grounds for hope and encouragement in this recently acquired ability occasionally to halt for a while the formerly unrelenting malignant process known as acute leukemia'. In reviewing the overall survival data for older adults one may feel that we are at a similar juncture in assessing the outcome for this particular population. It is hoped that some of the potential advances may provide greater hope and improved results over the next decade. PMID- 10720147 TI - New agents for acute myelogenous leukemia. AB - New agents for the treatment of acute myelogenous leukemia are discussed that reflect different treatment mechanisms. These include histone acetylation, angiogenesis inhibition, protein kinase inhibitors, and a novel retinoid. Efficacy and safety in phase I and phase II trials reviewed, as well as the problems involved in crossing over from treatment of solid tumors to blood disorders. PMID- 10720148 TI - Differentiation therapy in acute myelogenous leukemia (non-APL). AB - Successful treatment of acute promyelocytic leukemia (APL) has identified several novel approaches to induce leukemic cell differentiation and selective apoptosis by overcoming the site-specific transcriptional repression by dominant fusion leukemogenic proteins characteristic of APL and other forms of acute myelogenous leukemia (AML). These therapeutic approaches include the use of site-specific ligands, receptors and cytokines, disruption of dominant fusion leukemogenic proteins, chromatin remodeling and combining the above with cytotoxic chemotherapy. With the exception of cytotoxic chemotherapy, the above therapeutic strategies do not significantly affect normal hematopoiesis and their combinations have been shown to be synergistic in inducing myeloid differentiation and apoptosis in several AML cell lines and in patients with APL. These approaches are, in general, non-cross resistant and should be well tolerated particularly in elderly patients with AML. Clinical studies which include biologic end points for differentiation induction, histone acetylation and selective apoptosis are presently in development to evaluate these strategies in the treatment of AML. PMID- 10720150 TI - Transplantation of hematopoietic stem cells from alternate donors in acute myelogenous leukemia. AB - The number of allogeneic transplants from unrelated donors has grown in the past decade in part because of the expansion of the donor registry size. Patient survival has improved due to the selection of more closely matched donors and the development of effective infection prophylaxis. Relapse-free survival remains limited in patients with a large tumor burden at the time of transplantation. A higher marrow cell dose is the major factor to minimize transplant-related death. Future studies of peripheral blood stem cell transplants should be considered for patients with acute leukemia with the goal of enhancing the stem cell dose and improving survival. PMID- 10720149 TI - Is there a best transplant conditioning regimen for acute myeloid leukemia? AB - The outcome of marrow transplantation is largely determined by the effectiveness of the transplant preparative regimen. Nonetheless, there have been startlingly few randomized trials attempting to identify optimal regimens for specific conditions and, at present, no single approach has emerged as superior for the treatment of acute myeloid leukemia (AML) in the few trials that have been performed. Newer approaches that appear encouraging in phase II studies include substituting etoposide for cyclophosphamide, adding thiotepa to the traditional cyclophosphamide plus total body irradiation combination in the setting of T cell depletion, and using antibody-based targeted radiotherapy as part of the transplant regimen. The ability to obtain allogeneic engraftment with nonablative regimens may open the door to additional innovative approaches, combining very specific antileukemia therapy with relatively nontoxic measures to ensure engraftment. PMID- 10720151 TI - Use of thrombopoietic growth factors in acute leukemia. AB - Several hematopoietic growth factors have been shown to affect megakaryocyte development, and two, interleukin (IL)-11 and thrombopoietin (TPO) are presently being evaluated for use in patients with thrombocytopenia. In two studies patients who required one or more platelet transfusions during their first course of chemotherapy were found to require fewer platelet transfusions if their second cycle was augmented with IL-11. The drug was generally safe, with cardiovascular compromise the only significant complication occurring in a minority of patients. Although these reports included patients with various malignancies, studies of IL 11 in patients with myeloproliferative disorders have not been presented. In several clinical trials in cancer patients treatment with TPO was safe, and when administered early following a moderately aggressive cytotoxic insult was effective in accelerating platelet recovery. In addition, in both pre-clinical and clinical trials, TPO given to stem cell donors during mobilization lead to accelerated hematopoietic recovery. Finally, TPO appears safe when administered to patients with acute myelogenous leukemia (AML), both with respect to acute toxicity and long-term outcome of the leukemia. However, when used following a 7 day course of standard chemotherapy, the agent does not appear to accelerate platelet recovery. As such, additional clinical trials to test different growth factor regimens are ongoing. A number of studies have suggested that megakaryocytic growth factors may play a role in the biology of myeloproliferative disorders. Given the potential for adversely affecting patients with these disorders, the affects of IL-11 or TPO in patients with AML must continue to be carefully studied. PMID- 10720152 TI - Advances in the treatment of graft-versus-host disease. AB - Graft-versus-host disease (GVHD) is a complicated disease whose treatment requires an equally multifaceted approach. Recipient conditioning, donor T cell activation, and end stage effectors all may be potential targets for treatment. Many drugs used in the past are returning to the forefront for investigation. Some of the newer nucleoside analogs that are in various stages of development, such as fludarabine and pentostatin, are showing promising activity in GVHD. PMID- 10720153 TI - Molecular genetics in acute leukemia. AB - Improved techniques in identifying the chromosome changes and the affected genes that are involved in acute leukemias have led to improved treatments for these diseases. Identification of consistent chromosomal changes has allowed us to target the location of particular genes and has enabled us to focus our treatments more specifically to certain subtypes of leukemia. Translocations, in particular, are common cytogenetic abnormalities in human leukemia, and the prevalence of certain types of translocations varies with age. Cancers, lymphomas and leukemias are now known to be genetic diseases and it is recognized that genotype-specific therapies should be used that take into account the genetic alterations of the particular leukemia. PMID- 10720154 TI - Lack of p73 expression in mature B-ALL and identification of three new splicing variants restricted to pre B and C-ALL indicate a role of p73 in B cell ALL differentiation. PMID- 10720155 TI - Involvement of the MLL and RARalpha genes in a patient with acute monocytic leukemia with t(11;17)(q23;q12) PMID- 10720156 TI - Tandem duplication of the FLT3 gene in acute lymphoblastic leukemia: a marker for the monitoring of minimal residual disease. PMID- 10720157 TI - Comments on a published paper: Abe 'Infantile Leukemia and Soybeans'. PMID- 10720158 TI - Folate and carcinogenesis: an integrated scheme. AB - Collectively, the evidence from epidemiologic, animal and human studies strongly suggests that folate status modulates the risk of developing cancers in selected tissues, the most notable of which is the colorectum. Folate depletion appears to enhance carcinogenesis whereas folate supplementation above what is presently considered to be the basal requirement appears to convey a protective effect. The means by which this modulation of cancer risk is mediated is not known with certainty, but there are several plausible mechanisms which have been described. Folate plays a major role in the formation of S-adenosylmethionine, the universal methyl donor, as well as in the formation of purine and thymidine synthesis for DNA and RNA. Therefore, most mechanistic studies performed to date have focused on alterations in DNA methylation, disruption of DNA integrity and disruption of DNA repair, all of which have been observed with folate depletion. These aberrations in DNA are believed to enhance carcinogenesis by altering the expression of critical tumor suppressor genes and proto-oncogenes. Recently, the role of a common polymorphism of the methylenetetrahydrofolate reductase gene has been highlighted as well. This review presents those mechanisms which are the most likely candidates to explain folate's effects and it proposes an integrated scheme to explain how these mechanisms might interact. PMID- 10720159 TI - Magnesium deficiency modulates the insulin signaling pathway in liver but not muscle of rats. AB - Altered insulin secretion and sensitivity have been observed in Mg-deficient animals. However, the effects of Mg deficiency and supplementation on intracellular signaling events triggered by insulin are unknown. Therefore, we studied the early steps of insulin action in muscle and liver of rats fed Mg deficient (DF-6, DF-11) or control (CO-6, CO-11) diets for 6 or 11 wk, respectively, and Mg-deficient or control diets for 6 wk, followed by Mg supplementation for 5 wk (SDF and SCO groups, respectively). There were no differences in the glucose disappearance rate (K(itt)) or insulin signaling between CO-6 and DF-6 rats. Between the two groups of rats fed for 11 wk, the DF 11 group had a significantly greater K(itt). SDF and SCO rats had K(itt) that did not differ from CO-11 rats, but that were significantly lower than in DF-11 rats. In the latter rats, insulin receptor and insulin receptor substrate-1 protein and phosphorylation levels were elevated in liver and there was a greater association between the insulin receptor substrate-1 and p85 subunit of phosphatidyl-inositol 3-kinase compared with CO-11 rats. There were no differences in the early steps of insulin action in SDF and control rats. These results suggest that the normal insulin sensitivity maintained by Mg supplementation and the increased insulin sensitivity produced by a long period of Mg deprivation may result, at least in part, from alterations in or maintenance of the early molecular steps of insulin action in hepatic tissue. PMID- 10720160 TI - Orally administered leucine stimulates protein synthesis in skeletal muscle of postabsorptive rats in association with increased eIF4F formation. AB - We investigated the protein synthetic response of skeletal muscle to an orally administered dose of leucine given alone or in combination with carbohydrate. Male rats were freely fed (F) or food deprived for 18 h; food-deprived rats were then administered saline (S), carbohydrate (CHO), leucine (L) or a combination of carbohydrate plus leucine (CL). CHO and CL meals were isocaloric and provided 15% of daily energy requirements. L and CL meals each delivered 270 mg leucine. Muscle protein synthesis in S was 65% of F (P<0.01) 1 h after meal administration. Concomitant with lower rates of protein synthesis, phosphorylation of the translational repressor, eukaryotic initiation factor (eIF)4E-binding protein 1 (4E-BP1), was less in S, leading to greater association of 4E-BP1.eIF4E, and reduced formation of the active eIF4G.eIF4E complex compared with F (P<0.01). Oral administration of leucine (L or CL), but not CHO, restored protein synthesis equal to that in F and resulted in 4E-BP1 phosphorylation that was threefold greater than that of S (P<0.01). Consequently, formation of 4E BP1.eIF4E was inhibited and eIF4G.eIF4E was not different from F. The amount of eIF4E in the phosphorylated form was greater in S and CHO (P<0.01) than in all other groups. In contrast, no differences in the phosphorylation state of eIF2alpha or the activity of eIF2B were noted among treatment groups. Serum insulin was elevated 2.6- and 3.7-fold in CHO and CL, respectively, but was not different in L, compared with S (P<0.05). These results suggest that leucine stimulates protein synthesis in skeletal muscle by enhancing eIF4F formation independently of increases in serum insulin. PMID- 10720161 TI - Maternal and early dietary fatty acid intake: changes in lipid metabolism and liver enzymes in adult rats. AB - Over the last decade, much evidence has emerged to suggest that alterations in maternal nutrition during pregnancy may irreversibly affect aspects of physiological and biochemical functions in the fetus. This study was designed to determine the mechanisms involved in these alterations. Our hypothesis was that the type of maternal dietary fat received in early life could determine the level of lipoprotein lipase (LPL; EC 3.1.1.34) activity and gene expression which would be maintained into later life. A diet high in (n-3) polyunsaturated fatty acids was predicted to be associated with higher levels of lipoprotein lipase (LPL) activity and expression and lower levels of plasma triglyceride after a high fat meal challenge. Using a 2x2 factorial design, Wistar Albino rats were pair-fed either a fish oil diet (50 g/kg) or a mixed oil diet (50 g/kg) for the last 2 wk of gestation, during lactation and pups were fed these diets until 5 wk of age. After 5 wk, the rats were fed nonpurified diet. The rats were killed at 5 wk (young) or 10 wk (adult) of age after a mixed oil (50 g/kg) test meal. There were significant age effects on plasma triglyceride (P<0.02), cholesterol (P<0.001), glucose-dependent insulinotrophic polypeptide (GIP) (P<0.001) and liver glutathione reductase activity (P<0.05) which were all higher in the young rats compared to the adults. There were significant effects of diet on triglyceride (P<0.001), cholesterol (P<0.001) and LPL mRNA levels (P<0.001). GIP and triglyceride levels were significantly correlated (r = 0.66; P<0.001). Omental adipose tissue LPL activity as significantly higher in the fish-oil fed groups compared to the other groups (P<0.001), whereas Epididymal adipose tissue LPL mRNA was significantly higher in the mixed oil-fed adults compared to the other groups (P<0.001). The latter result suggested an imprinting effect of fatty acid composition in early life on LPL gene expression. Liver superoxide dismutase activity was affected by age and diet and was higher in the young than in the adults and higher in the fish oil-fed young than in those fed the mixed oil-fed (P<0.005). Catalase activity was also affected by age (P<0.001) and diet (P<0.001), and there was a significant interaction between age and diet (P<0.001). Catalase activity was higher in rats fed fish oils at both stages of development, suggesting that feeding fish oils to rats in early life raises oxidative stress throughout life. The majority of the significant differences shown were between the age groups and not between the two dietary groups, suggesting that postprandial handling of a standard fat meal is affected more by age than by early dietary fatty acid composition. However, the mechanisms of biological imprinting of fatty acids on LPL expression and on enzymes related to oxidative stress requires more investigation. PMID- 10720162 TI - Caffeine, carnitine and choline supplementation of rats decreases body fat and serum leptin concentration as does exercise. AB - The effect of a combination of caffeine, carnitine and choline with or without exercise on changes in body weight, fat pad mass, serum leptin concentration and metabolic indices was determined in 20 male, 7-wk-old Sprague-Dawley rats. They were given free access to a nonpurified diet without or with caffeine, carnitine and choline at concentrations of 0.1, 5 and 11.5 g/kg diet, respectively. In a 2x2 factorial design, one-half of each dietary group was exercised, and the other half was sedentary. Body weight and food intake of all rats were measured every day for 28 d. Rats were killed and blood and tissue samples were collected and analyzed for biochemical markers. Food intake of the groups was not different, but the body weight was significantly reduced by exercise in both dietary groups. Fat pad weights and total lipids of epididymal, inguinal and perirenal regions were significantly reduced by the supplements as well as by exercise. Regardless of exercise, supplements significantly lowered triglycerides in serum but increased levels in skeletal muscle. Serum leptin concentrations were equally lowered by supplements and exercise. Serum leptin was correlated with body weight (r = 0.55, P< or =0.01), fat pad weight (r = 0.82, P< or =0.001) and serum glucose (r = 0.51, P< or =0.05). We conclude that the indices of body fat loss due to dietary supplements were similar to those due to mild exercise, and there were no interactive effects of the two variables. PMID- 10720163 TI - Chelation of zinc amplifies induction of growth hormone mRNA levels in cultured rat pituitary tumor cells. AB - Zinc is thought to be an integral part of nuclear receptor proteins, stabilizing them in a conformation required for binding to target genes. However, we have recently shown that restriction of zinc availability with a chelator (diethylenetriaminepenta-acetic acid, DTPA) enhances, rather than inhibits, the ability of thyroid hormone to induce growth hormone mRNA expression in GH3 rat pituitary tumor cells. In this report, we have extended these observations by showing that a prolonged (48 h) exposure to DTPA is required to see these effects. The induction by DTPA can be reversed by subsequent addition of zinc, but again, this reversal is slow. A second chelator, EDTA, can also induce growth hormone gene expression in the presence of thyroid hormone, though it is less potent than DTPA. Other agents which act via the nuclear receptor pathway, all trans and 9-cis retinoic acid, also induce expression of growth hormone mRNA. Addition of DTPA amplifies these effects in a zinc-dependent manner. Thus chelation of zinc potentiates the action of ligands acting via nuclear receptors on growth hormone gene expression. The delayed nature of the response suggests an indirect effect. PMID- 10720164 TI - Rye bread decreases serum total and LDL cholesterol in men with moderately elevated serum cholesterol. AB - The objective of this study was to determine the hypocholesterolemic effects of whole meal rye and white wheat breads in healthy humans with elevated serum cholesterol concentrations, and the changes in plasma glucose and insulin concentrations during rye and wheat bread periods. The subjects were 18 men and 22 women with baseline serum cholesterol concentration of 6.4+/-0.2 mmol/L. The study design was a 2x4-wk crossover trial during which each subject randomly consumed rye and wheat breads (20% of daily energy) as part of their usual diet for 4 wk. The bread periods were separated by a 4-wk washout period. Blood samples (after fasting) were collected on two consecutive days at the beginning and end of the bread periods. Serum total cholesterol decreased by 8% (P = 0.002) in men but was not significantly altered in women during the rye bread period. The wheat bread period did not affect any of the variables studied. Analysis of the serum lipids in tertiles of rye bread consumption confirmed the reduction in total cholesterol (P = 0.048) in men and revealed the reduction in LDL cholesterol (P = 0.032); both were dependent on the amount of rye bread consumed (-2, -14 and -10% in total cholesterol and 0, -12 and -12% in LDL cholesterol). Neither rye nor wheat bread influenced the concentrations of glucose and insulin. In conclusion, rye bread is effective in reducing serum total and LDL cholesterol concentrations in men with elevated serum cholesterol. Good compliance with consuming a relatively large amount of rye bread in the usual diet indicates that rye bread offers a practical dietary means of reducing serum cholesterol in men. PMID- 10720165 TI - Walnuts lower serum cholesterol in Japanese men and women. AB - Recent studies have shown that incorporating moderate quantities of walnuts into the recommended cholesterol-lowering diet in the U.S. decreased serum concentrations of total cholesterol in normal American men. To explore whether walnut consumption would also prove effective as part of the Japanese diet, we studied the effects of walnut consumption on serum lipids and blood pressure in Japanese subjects. We randomly assigned 20 men and 20 women to two mixed natural diets, each to be consumed for 4 wk in a crossover design. Both diets conformed to the average Japanese diet (reference diet) and contained identical foods and macronutrients, except that 12.5% of the energy of the walnut diet was derived from walnuts (43-57 g/d) (offset by lesser amounts of fatty foods, meat and visible fat). Total cholesterol concentration was 0.16 mmol/L lower for men (P = 0.05) and 0.21 mmol/L lower for women (P<0.01) when they consumed the walnut diet than when they consumed the reference diet. The LDL cholesterol concentrations were 0.18 mmol/L lower for men (P = 0.13) and 0.22 mmol/L lower for women (P<0.01) when they consumed the walnut diet. The ratio of LDL cholesterol to HDL cholesterol and the apolipoprotein B concentration were also lowered by the walnut diet (P<0.05). Blood pressures did not differ between the walnut and reference diet periods. Incorporating moderate quantities of walnuts into the average Japanese diet while maintaining the intake of total dietary fat and energy decreases serum total cholesterol concentrations and favorably modifies the lipoprotein profile in Japanese, particularly in women. PMID- 10720166 TI - The pharmacokinetic responses of humans to 20 g of alanyl-glutamine dipeptide differ with the dosing protocol but not with gastric acidity or in patients with acute Dengue fever. AB - Pharmacokinetic responses to oral doses of the dipeptide, L-alanyl-glutamine (Ala Gln), were evaluated after a single, bolus load or an intermittent dosing in normal healthy subjects (n = 8) to find the optimal mode of oral administration. In a subgroup (n = 4) of the healthy subjects, the influence of a gastric acid suppressor (Omeprazole) was investigated. The influence of an acute episode of classic Dengue fever was examined in eight patients. All modes of administration to healthy subjects significantly increased free plasma Gln and alanine concentrations. Peak increments of plasma Gln concentration were 794+/-107 micromol/L (mean +/- SEM) after bolus intake of 20 g of Ala-Gln and 398+/-61 micromol/L after intermittent intake of the same cumulative dosage of the dipeptide (P<0.01). After intermittent dosing, the maximum peak increase appeared significantly later (P<0.01). Areas under the curve (AUC), expressing the integrated responses over time of plasma free Gln and alanine concentrations, did not differ after bolus and intermittent loads of Ala-Gln. Pretreatment with the acid suppressor, Omeprazole, did not influence Gln (P = 0.79) or alanine (P = 0.90) plasma increment. Dengue patients manifested the same pharmacokinetic responses to a 20 g Ala-Gln bolus as healthy controls. In general, on a micromolar concentration basis, Gln and alanine followed parallel tracks in terms of plasma appearance, clearance and elimination after the oral administration of 20 g of the Ala-Gln dipeptide through the range of conditions and dosing protocols explored here. PMID- 10720167 TI - Plasma urea appearance rate is lower when children with kwashiorkor and infection are fed egg white-tryptophan rather than milk protein. AB - In kwashiorkor, there is less endogenous proteolysis in response to acute infection than in a well-nourished state. Thus the amino acid composition of dietary protein may be more important in facilitating the acute phase response in kwashiorkor. This study tested the hypothesis that during the treatment of kwashiorkor with infection, there is a lower rate of urea appearance when the dietary intake of amino acids more closely resembles the amino acid composition of acute phase proteins. Thirty children in Malawi with kwashiorkor and acute infection were fed isoenergetic, isonitrogenous meals containing either egg white tryptophan or milk as a protein source. After 24 h, the rates of urea appearance and whole-body protein breakdown and synthesis were measured with the use of 1 13C-leucine and 15N2-urea tracers. Plasma concentrations of seven acute phase proteins, interleukin 6 and tumor necrosis factor-alpha were measured on admission, and at 24 and 48 h. The 16 children who received egg white-tryptophan had lower rates of urea appearance than those who received milk [57+/-30 vs. 87+/ 36 micromol/(kg x h), mean +/- SD, P<0.02]. No significant differences were found in the rates of whole-body protein turnover or in the concentration of any of the acute phase proteins or cytokines. The concentration of interleukin 6 was consistent with an appropriate proinflammatory response and correlated directly with the concentrations of C-reactive protein (r = 0.67, P<0.01) and alpha1 antitrypsin (r = 0.40, P<0.05). The findings suggest that egg white-tryptophan is associated with less amino acid oxidation in kwashiorkor and acute infection than is milk. PMID- 10720168 TI - Lymphocyte lycopene concentration and DNA protection from oxidative damage is increased in women after a short period of tomato consumption. AB - Several epidemiologic studies have suggested a role of tomato products in protecting against cancer and chronic diseases. In nine adult women, we evaluated whether the consumption of 25 g tomato puree (containing 7 mg lycopene and 0.3 mg beta-carotene) for 14 consecutive days increased plasma and lymphocyte carotenoid concentration and whether this was related to an improvement in lymphocyte resistance to an oxidative stress (500 micromol/L hydrogen peroxide for 5 min). Before and after the period of tomato intake, carotenoid concentrations were analyzed by HPLC and lymphocyte resistance to oxidative stress by the Comet assay, which detects DNA strand breaks. Intake of tomato puree increased plasma (P <0.001) and lymphocyte (P<0.005) lycopene concentration and reduced lymphocyte DNA damage by approximately 50% (P<0.0001). Beta-carotene concentration increased in plasma (P<0.05) but not in lymphocytes after tomato puree consumption. An inverse relationship was found between plasma lycopene concentration (r = -0.82, P<0.0001) and lymphocyte lycopene concentration (r = -0.62, P<0.01) and the oxidative DNA damage. In conclusion, small amounts of tomato puree added to the diet over a short period can increase carotenoid concentrations and the resistance of lymphocytes to oxidative stress. PMID- 10720169 TI - Current methods for estimating dietary iron bioavailability do not work in China. AB - Three current equations for estimating iron bioavailability were evaluated, and adjustments were proposed that would allow us to most effectively study iron bioavailability in China. Dietary intake data were obtained from 24-h dietary recalls taken over three consecutive days as part of the third Chinese National Nutrition Survey. Hemoglobin status was measured for 42,606 Chinese adults aged 18-60 y. The mean iron intake was 24.4 mg per capita per day, which was 177% of the Chinese RDA (209% of U.S. RDA). About 18% of the sample was classified as being anemic, indicating a large iron deficiency anemia and iron bioavailability problem in China. A number of methods proposed by World Health Organization and U.S. scholars were examined for adjusting iron bioavailability. Even the methods that consider several iron enhancers and inhibitors did not work adequately for the Chinese diet. The statistical assessment of the fit between iron bioavailability and hemoglobin status provided direction for adjusting the best of these predictive equations. We propose a new predictive approach for iron bioavailability which is more predictive of Chinese iron status. Consideration of additional dietary elements such as rice and bean consumption patterns are important. Our findings provide insight into additional factors which may influence iron bioavailability as well as possible improved methods for estimating the combined effect of multiple dietary factors on iron bioavailability, particularly in a vegetarian diet. PMID- 10720170 TI - Predictors of micronutrient status among six- to twelve-month-old breast-fed Ghanaian infants. AB - This study describes the factors associated with hemoglobin and plasma ferritin, zinc and retinol concentrations and erythrocyte riboflavin status among 208 Ghanaian infants who participated in a complementary feeding intervention trial from 6 to 12 mo of age. Anthropometric, morbidity and dietary data were collected regularly from 1 to 12 mo; blood samples were collected at 6 and 12 mo. The prevalence of low micronutrient status at 6 and 12 mo, respectively, was as follows: hemoglobin <100 g/L, 30 and 34%; plasma ferritin <12 microg/L, 17 and 43%; plasma zinc <10.7 micromol/L, 4 and 6%; plasma retinol <0.7 micromol/L, 26 and 26%; erythrocyte riboflavin <200 umol/L of packed red cells, 14 and 10%. Multiple regression was used to identify factors significantly associated with micronutrient status. From 6 to 12 mo, fever prevalence was associated with a decrease in hemoglobin, but an increase in erythrocyte riboflavin concentrations, and diarrhea prevalence was related to a decrease in plasma retinol. Seasonal differences were evident for most of the indicators of micronutrient status, and elevated C-reactive protein levels (indicative of recent infection) were related to lower hemoglobin, retinol and zinc concentrations but higher ferritin and erythrocyte riboflavin concentrations. Weight at birth or at 1 mo of age was positively related to iron, zinc and vitamin A status, but a more rapid weight gain was associated with depletion of iron stores. Socioeconomic status was related to higher hemoglobin, riboflavin and zinc concentrations. The feeding of a micronutrient-fortified food was positively associated with plasma ferritin and retinol concentrations at 12 mo. These results suggest that prenatal factors, socioeconomic status, dietary intake and morbidity all influence infant micronutrient status, and that fortification of complementary foods is one potential avenue for preventing deficiencies. PMID- 10720171 TI - Trans-vaccenic acid is desaturated to conjugated linoleic acid in mice. AB - Mice were fed pure trans11 octadecenoic acid (trans-vaccenic acid; TVA) to determine whether it is desaturated to cis9, trans11 octadecadienoic acid, a predominant isomer of conjugated linoleic acid (CLA). In a preliminary trial, 12% of the TVA consumed during a 2-wk feeding period was recovered in the carcass as CLA. As a proportion of TVA in the tissues available for bioconversion, 48.8% was desaturated. We tested whether desaturation could be modified by supplementing no modifier, 0.5% clofibric acid to stimulate desaturation, or increasing the polyunsaturated fatty acids (PUFA) (10% corn oil vs. 4% corn oil) to inhibit desaturation in diets with or without 1% TVA. These diets were fed to six groups of mice in a 3x2 factorial arrangement of treatments. Feeding 1% TVA with 10% corn oil decreased feed intake (2.70 vs. 3.73 g/d, SEM 0.23; P<0.05). Bioconversion of dietary TVA was 12.0, 7.5 and 5.1% for mice fed no modifier of desaturation, clofibrate and increased PUFA, respectively. Conversion based on TVA available for desaturation was 52.6, 55.5 and 37.0%, respectively. Thus, clofibrate did not increase bioconversion, but increasing PUFA decreased conversion by 30%. To test whether TVA decreases food intake directly or after conversion to CLA, four groups of mice were fed diets containing 1% stearic, TVA, elaidic or conjugated linoleic acid. Dietary CLA decreased food intake and body fat, but did not change body protein. CLA was found in the carcass only when TVA or CLA was fed. CLA was found in both triacylglycerol and phospholipids when CLA was fed, but only in triacylglycerol when TVA was fed, suggesting that bioconversion occurred in the adipose tissue. In three trials, conversion of dietary TVA to CLA was 11.4+/-1.25%; conversion of stored TVA was 50.8+/-1.91%. Similar bioconversion of TVA in humans would increase current estimates of CLA available for the general population by 6- to 10-fold. PMID- 10720172 TI - Dietary magnesium supplementation affects bone metabolism and dynamic strength of bone in ovariectomized rats. AB - We evaluated the effect of magnesium supplementation on apparent calcium absorption, bone metabolism and dynamic bone strength in ovariectomized (OVX) rats as a model of postmenopausal women. Two groups of OVX rats were fed a 0.05% Mg diet or a 0.15% Mg diet, and one group of sham-operated rats was fed the 0.05% Mg diet for 42 d. We collected feces and urine of all rats for 3-d periods starting from d 3, 10, 17, 24, 31 and 38 of the feeding experiment for calcium and magnesium balance studies. Urine was collected for 24 h from d 41 of the feeding experiment for measuring deoxypyridinoline. After the 42 d, the rats were killed, serum prepared and femora excised. The apparent calcium absorption in the OVX rats fed 0.15% Mg was significantly lower than both other groups. Additionally, the urinary excretion of deoxypyridinoline (a bone resorption marker) and the serum parathyroid hormone level of the OVX rats fed the 0.15% Mg diet were significantly lower than in the OVX rats fed 0.05% Mg. Serum osteocalcin (a bone formation marker) in the OVX rats fed the 0.15% Mg diet was significantly higher than in the OVX rats fed 0.05% Mg. The breaking force and breaking energy of the femur in the OVX rats fed the 0.15% Mg diet were significantly higher than in the OVX rats fed the 0.05% Mg diet. These results indicate that magnesium supplementation reduces apparent calcium absorption, but promotes bone formation and prevents bone resorption in OVX rats. Moreover, our results indicate magnesium supplementation increases the dynamic strength of bone. PMID- 10720173 TI - Serum lipid concentrations and mean life span are modulated by dietary polyunsaturated fatty acids in the senescence-accelerated mouse. AB - The senescence-accelerated mouse (SAMP8) is an animal model used in studies of aging. This study was undertaken to investigate the effects of dietary PUFA on longevity (Experiment 1) and serum lipid concentrations (Experiment 2) in SAMP8 mice. Male mice were fed either an (n-3) PUFA-rich (9 g/100 g perilla oil) or an (n-6) PUFA-rich (9 g/100 g safflower oil) diet beginning at 6 wk of age. Experiment 1: The groups did not differ in body weight gain, but those fed perilla oil had significantly lower scores of senescence relative to those fed safflower oil (P<0.05). The mean life span of mice fed perilla oil was 357+/-21 d and of those fed safflower oil, 426+/-24 d (P<0.05). Pathological studies revealed that the incidence of tumors was significantly lower in the perilla oil group than in the safflower oil group (P<0.05). Approximately half the mice fed perilla oil had died after 10 mo, and the direct causes closely connected with death could not be specified. Experiment 2: The serum total cholesterol, HDL cholesterol, triglyceride and phospholipid concentrations were significantly lower in the perilla oil group than in the safflower oil group (P<0.01). A marked decrease of serum HDL cholesterol and apolipoprotein A-II (ApoA-II)concentrations in advanced age were observed in the mice fed perilla oil (P<0.01). Ten-month-old mice fed perilla oil had a significantly greater ratio of apolipoprotein A-I (ApoA-I) to ApoA-II than those fed safflower oil. Separation of HDL subfractions revealed that the smaller HDL species were much more abundant than the larger HDL species in both dietary oil groups. These findings suggest that dietary (n-3) and (n-6) PUFA differ in their effects on serum lipid metabolism which may modulate the mean life span of SAMP8 mice fed each dietary oil. PMID- 10720174 TI - Restricted feed intake during fattening reduces intramuscular lipid deposition without modifying muscle fiber characteristics in rabbits. AB - The present study was conducted to determine the effects of feed restriction during fattening on muscle fiber characteristics and intramuscular lipid traits. From 11 wk of age onward, rabbits were given free access to feed (control group), or received 70% of the control feed intake (restricted group). At the same weight at slaughter, restricted-fed rabbits were 3 wk older than controls (18 vs. 15 wk). The longissimus lumborum (LL, white loin), biceps femoris (BF, white thigh) and semimembranosus proprius (SMP, red thigh) muscles were then removed, and biochemical and histochemical assays were performed. In the three muscles, there was no effect of feed restriction on mean fiber size or percentage of the different fiber types. Restricted vs. control feeding resulted in a significant reduction (P<0.001) in total lipid content in all three muscles. This reduction was paralleled by a decline (P<0.001) in the activities of the malic enzyme and glucose-6 phosphate dehydrogenase (G6PDH), generating NADPH for the support of fatty acid synthesis. The diet-induced variations in lipid concentration and enzyme activities were larger (P<0.05) in the pure oxidative SMP muscle than in the predominantly fast-twitch glycolytic LL and BF muscles. Whatever feeding status, the ratio of malic enzyme to G6PDH activities was sharply lower (P<0.001) in SMP than in BF and LL muscles (averaging 1.5 vs. 9 and 15, respectively). These data indicate that nutritional status regulates intramuscular lipid deposition, without changing fiber-type composition. Further studies are necessary to determine the role of G6PDH in the lipogenic process of oxidative vs. glycolytic muscles. PMID- 10720175 TI - Butyric acid is synthesized by piglets. AB - We hypothesized that there is no synthesis of butyric acid within organs or tissues not drained by the portal vein (PV). Two experiments were performed. In six piglets, the colonic vasculature was clamped (n = 4) or the entire colon resected while [1-13C]-butyric acid (99% enriched) was infused into a jejunal vein for 120 min; 13C enrichment of butyric acid was measured in the PV and carotid artery (ART) during the last 30 min of the infusion. In a second experiment, butyric acid tracer and unlabeled disaccharide were infused into the cecum for 120 min, and blood again was sampled from the PV and ART. For the four piglets studied during ligation of the colonic vasculature, the mean (+/- SD) ratio of the butyric acid enrichment in the ART to that in the PV (ART/PV) was 0.80+/-0.05 (ART vs. PV, P = 0.002) and for all six piglets in expt. 1, the ART/PV ratio was 0.74+/-0.1 (ART vs. PV, P = 0.001). The enrichment of butyric acid in the PV averaged 96.0% for the six studies, implying that splanchnic tissues other than the colon did not produce a substantial amount of butyric acid. For the second experiment, the ART/PV ratio was 0.80+/-0.15 (ART vs. PV, P = 0.03). These studies provide the first evidence for endogenous synthesis of butyric acid by piglets. PMID- 10720176 TI - Choline is required by tilapia when methionine is not in excess. AB - Choline is essential in diets fed to most young vertebrates, but previous studies did not confirm the essentiality of choline in diets fed to tilapia. Two experiments were conducted to evaluate the essentiality of dietary choline in such diets. The basal diet used in both experiments contained 32 g crude protein/100 g diet (10.1 g crude protein from casein and gelatin, and 21.9 g from a crystalline L-amino acid mixture). The total sulfur amino acid (TSAA) concentration of the basal diet was 0.28 g/100 g diet, Met:Cys 89:11. In Experiment 1, a 4x2 design was used in which crystalline L-methionine was added to the basal diet resulting in four levels of TSAA (0.28, 0.50, 0.75 or 1.0 g/100 g diet, Met:Cys 89:11, 94:6, 96:4, or 97:3, respectively). At each level of TSAA, diets also contained either 0 or 1 g choline/kg diet supplied as choline chloride. Weight gain, feed efficiency (FE) and serum methionine concentrations were significantly affected by dietary TSAA concentration, but not by dietary choline concentration or the interaction between TSAA and choline. Weight gain, feed efficiency and serum methionine concentrations indicated that the TSAA requirement was 0.5 g/100 g diet. In the second experiment, diets were formulated to contain either 0.28 or 0.5 g TSAA/100 g diet, Met:Cys 89:11 or 94:6, respectively, and graded levels of choline ranging from 1 to 4 g/kg diet in gradations of 1 g/kg. Dietary methionine significantly affected weight gain and FE, whereas dietary choline significantly affected weight gain, FE and survival, and the interaction of methionine and choline significantly affected weight gain. Fish fed diets containing 0.5 g TSAA/100 g diet and 3 g choline chloride/kg diet exhibited the highest weight gain, feed efficiency and survival. On the basis of these data, it seems clear that juvenile tilapia require choline in certain dietary formulations. PMID- 10720177 TI - Low levels of viscous hydrocolloids lower plasma cholesterol in rats primarily by impairing cholesterol absorption. AB - Hydrocolloids have been proposed as cholesterol-lowering agents, but their viscosity limits their use in human nutrition. A low level (1 %) of hydrocolloids (guar gum, (GG); xanthan gum, (XG); and konjac mannan) was investigated in rats fed 0.2 g/100 g cholesterol diets. Food intake and body weight gain were not altered by the diets. Bile flow and cholesterol bile flux were not modified by diet, whereas the bile acid flux was greater in rats fed hydrocolloid diets. The cecal pool of bile acids was greater than control rats only in rats fed the XG diet (+71%, P<0.001). The fecal excretion of neutral sterols was stimulated in rats fed the hydrocolloid diets; cholesterol apparent digestibility (60% in controls) was reduced to 30-36% in rats fed hydrocolloids. Bile acid fecal excretion was not altered by diet treatment. As a result, apparent steroid balance was about +40 micromol/d in controls and only +10 to +20 micromol/d in rats fed hydrocolloids. Both plasma cholesterol and triglycerides were significantly lower than controls in rats fed XG, but only cholesterol was lower in rats fed the GG diet. These effects were essentially found in the d <1.040 kg/L fraction. Liver cholesterol content was significantly lower than in controls in rats fed the GG or XG diets. Liver HMG CoA reductase was not affected by the hydrocolloid diets. In conclusion, a low percentage of viscous hydrocolloids lowers plasma cholesterol in cholesterol-fed rats. Inhibition of intestinal cholesterol absorption may be the primary mechanism. PMID- 10720178 TI - The rhamnogalacturonan-II dimer decreases intestinal absorption and tissue accumulation of lead in rats. AB - The rhamnogalacturonan-II dimer (dRG-II) forms strong complexes in vitro with lead (Pb) and other selected cations. We examined the in vivo bioavailability of Pb complexed with dRG-II and the effect of unleaded dRG-II on the intestinal absorption and tissue retention of Pb in rats. Forty male Wistar rats were divided into four groups. Each group consumed a purified control diet for 3 wk or the same diet supplemented with: i) 3 mg of Pb/kg, ii) 0.5 g of leaded dRG-II/kg, or iii) 0.5 g of leaded dRG-II/kg and 4.5 g of unleaded dRG-II/kg. The leaded dRG II provided approximately 3 mg of Pb/kg of diet. A chemical balance study was conducted during the last 5 d of the 3-wk study, and blood and organs were sampled for Pb and mineral analyses. The apparent intestinal absorptions of Pb were 62.3, 15.2, 11.8 and -0.1%, and Pb balances were 1.9, 9.6, 5.6 and -0.2 microg/d for the control and the three experimental groups, respectively. The Pb complexed with dRG-II was less available than Pb acetate, as reflected by significantly lower blood and tissue Pb levels. The addition of unleaded dRG-II decreased the intestinal absorption and the tissue retention of Pb significantly. We further found that the apparent absorption and status of magnesium, zinc and iron were unaffected by Pb treatment or dRG-II addition. We conclude that dRG-II may be useful in decreasing toxicity related to chronic Pb exposure. Human studies will be necessary however, to further evaluate the clinical utility of this beneficial effect. PMID- 10720180 TI - "Vitamin D mysteries"? Secretions and sloughings from skin and oral gastrointestinal mucosa contain hormone. PMID- 10720179 TI - Variations in dietary iron alter brain iron metabolism in developing rats. AB - The rat has been widely used as a model for the study of iron deficiency (ID), but the differences in the timing of development of humans and rats must be taken into account to derive appropriate conclusions from the animal model. This study was designed to evaluate the effects of dietary ID and iron excess on rat brain iron and the iron metabolism proteins, transferrin (Tf), transferrin receptor (TfR) and ferritin. The experimental design is developmentally sensitive and permits control of the timing as well as the duration of the nutritional insult. Iron-deficient and iron-supplemented (SU) rats between postnatal day (PND) 10 and 21, PND 21 and 35 and PND 10 and 35 were used to study the effects of early, late, and long-term iron deficiency and supplementation. Some ID rats were iron repleted between PND 21 and 35. These experiments demonstrated several new findings: 1) Early ID/SU (PND 10-21) altered brain iron, TfR, Tf and ferritin concentration in many regions different from those observed in the later period (PND 21-35). 2) Two weeks of iron repletion were adequate for correcting the overall Fe concentration of the brain and of individual brain regions, although larger amounts of iron were necessary to fully normalize iron and its regulatory proteins. 3) Long-term ID/SU resulted accordingly in the continued decrease or increase in brain iron concentration in some brain regions and not others. In conclusion, brain regions regulate their iron concentration in response to local needs when faced with alterations in systemic iron delivery. PMID- 10720181 TI - Epithelial cell kinetics in the inflammatory process of chicken trachea infected with infectious bronchitis virus. AB - All stages of degeneration and regeneration in chicken tracheal epithelium were studied morphologically following an intratracheal inoculation of infectious bronchitis virus (IBV). Viral antigen was detected in the cytoplasm of tracheal epithelium from 1 to 7 days post-inoculation (d.p.i.) with a peak on 3 d.p.i. At 1 d.p.i., almost all epithelial cells were involved in the degeneration. At this time, labelling index of bromodeoxyuridine (BrdU) in the basal cells showed significantly high value compared with control. At 2 and 3 d.p.i., a great number of basal cells were recognized, but the BrdU labelling index tended to decrease. At 4 and 5 d.p.i., the BrdU labelling index of basal cells significantly decreased than 1 d.p.i., and a few number of regenerated immature ciliated epithelia appeared. At 6 to 11 d.p.i., the ciliated columnar epithelia increased rapidly in number, and returned to the normal appearance except for non-ciliated patch by 13 d.p.i. These results suggested that the tracheal epithelial cells infected with IBV degenerated within 24 hours and proliferating activity of basal cells functioned immediately, and 3 to 4 days later, these basal cells were differentiated to the ciliated epithelia. PMID- 10720182 TI - Pulmonary hypoplasia induced by liquid paraffin injection into fetal thoracic cavity with special reference to renal development in rats. AB - The present study was designed to clarify lung-kidney interrelation in fetal rats. On fetal day 20, liquid paraffin (LP) was injected into fetal thoracic cavity to produce pulmonary hypoplasia. No significant difference in body and renal weights were noted between the LP injected and control fetuses. The weight of lung, however, was significantly lower in the LP injected fetuses than in the control ones. Histological examinations on the lung and kidney of the LP injected fetuses revealed that the lung was hypoplastic characterized by rich interstitium and reduced air spaces. In the kidney, mature types of glomeruli and profiles of proximal tubules near them were increased in number. Furthermore, strong expression of EGF immunoreactivity was noted in the apical cytoplasm of epithelium of the proximal tubules in the LP injected fetuses. These findings indicate that lung-kidney interrelation exists in fetal rats during late gestational days, and suggest that interruption of the lung development induces accelerated growth of the kidney in fetal rats. PMID- 10720183 TI - Effects of dizocilpine pretreatment on parvalbumin immunoreactivity and Fos expression after cerebral ischemia in the hippocampus of the Mongolian gerbil. AB - The mechanisms of ischemic neuronal death have been focused on glutamate receptor activation and subsequent elevation of intracellular Ca2+ concentration. The purpose of this study was to evaluate the effects of dizocilpine, an NMDA receptor antagonist, pretreatment on Fos expression and parvalbumin (PV, calcium binding protein) immunoreactivity in the hippocampus of the mongolian gerbil after global ischemic insults. The number of PV-immunoreactive (PV-ir) neurons in CA1 were significantly decreased from 1 day after cerebral ischemia, while dizocilpine pretreatment completely suppressed the loss of PV-ir neurons in CA1. Dizocilpine pretreatment also protected the structural loss of microtubule associated protein 2 immunoreactivity in CA1 after ischemic insults. In addition, dizocilpine pretreatment increased Fos expression in both hippocampal CA3 and CA4 after 3 hr ischemic reperfusion as compared to that of the saline pretreated group. Subsequently, the Fos-defined cellular activity of PV-ir neurons was slightly increased by dizocilpine pretreatment in the hippocampal area. This study demonstrated that NMDA receptor mediated calcium influx was associated with the loss of PV-ir neurons in CA1 hippocampal region, and that dizocilpine pretreatment increased Fos expression and the neuronal activity of PV-ir neurons in the non-vulnerable region of hippocampus after cerebral ischemia. Based on this data, we conclude that the protective effect of dizocilpine may be induced by the regulation of calcium overload, or by the upregulation of a neuroregenerative initiator such as Fos protein. PMID- 10720184 TI - A pathologic study on ocular disorders in calves in southern Kyushu, Japan. AB - Of 822 calves, ranging in age between one day and six months necropsied between 1996 and 1998 at Miyazaki University, histological examination showed that 25 (3.0%) had ocular lesions. These ocular lesions consisted of suppurative inflammation (13 cases), cataract (seven cases), and retinal atrophy (five cases). Inflammatory changes were classified as suppurative keratitis (one case), keratitis and uveitis (ten cases), and uveitis and retinitis (two cases). Cataract was subclassified into three categories; cortical (three cases), nuclear (one case), and mature (three cases). These lesions were characterized by degenerative changes in the lens fibers and the appearance of eosinophilic globules known as Morganian globules. In the most severely affected case, there was capsular rupture of the lens, resulting in severe infiltration by eosinophils and histiocytes of the whole anterior chamber. Almost all the calves with retinal atrophy had been suffering from severe hydranencephaly and three had significantly raised levels of neutralization antibodies for the Akabane and/or Aino viruses. This study indicates that congenital arbovirus infections may predispose calves to ocular diseases, especially retinal atrophy. PMID- 10720185 TI - Expression patterns of glycoconjugates in the three distinctive olfactory pathways of the clawed frog, Xenopus laevis. AB - Xenopus laevis has three distinctive olfactory neuroepithelia. We examined the axonal projection from each of these epithelia to the olfactory bulb by Di-I labeling, and confirmed that the Xenopus primary olfactory pathways involve the dorsal pathway from the olfactory epithelium to the dorsal region of the main olfactory bulb, the ventral pathway from the middle chamber epithelium to the ventral region of the main olfactory bulb, and the vomeronasal pathway from the vomeronasal epithelium to the accessory olfactory bulb. We next examined expression patterns of glycoconjugates in the three olfactory pathways by lectin histochemistry using 21 biotinylated lectins. Fourteen out of 21 lectins stained the Xenopus primary olfactory system. RCA-I stained the three olfactory pathways uniformly. PHA-E stained only the dorsal pathway. LEL, STL, PNA, ECL and UEA-I stained the dorsal pathway more intensely than the ventral pathway, and among them, only UEA-I stained the vomeronasal pathway. In contrast, s-WGA, DBA, SBA, BSL-I VVA, SJA and PHA-L showed intense stainings in the ventral pathway and moderate stainings in the vomeronasal pathway, but faint or weak stainings in the dorsal pathway. These observations suggest that the ventral pathway expresses glycoconjugates shared commonly with either the dorsal or the vomeronasal pathway. In addition, from the binding patterns of the lectins with a binding specificity for N-acetylgalactosamine, glycoconjugates containing this saccharide seem to play an important role for the organization of the olfactory pathways. PMID- 10720186 TI - Distribution and developmental change of lymphoid tissues in the chicken proventriculus. AB - In the chicken proventricular mucosa, aggregations of lymphocytes were localized in three different sites of the lamina propria, namely, underneath the surface epithelium, near the duct orifice of the deep proventricular gland, and in the gland tissue itself. In the lymphoid masses underneath the surface epithelium and in those near the duct orifice, CD4+ T lymphocytes and TCR2+ T lymphocytes occupied their central part, and B lymphocytes were localized in the periphery. CD8+ T lymphocytes and TCR1+ lymphocytes were evenly distributed in the masses. Infiltration of lymphocytes into these sites was first observed on the 20th embryonic day. At 1 week after hatching, CD3+ lymphocytes began to occupy the central area of the masses and His-C1+ B lymphocytes tended to be located in the periphery. Ultrastructurally, M cells were found neither in the epithelium of the mucosa nor in that of the excretory duct close to the lymphoid masses. In the deep proventricular gland, the lymphoid masses had a germinal center consisting of B lymphocytes, surrounded by the T lymphocyte-rich periphery. These masses were first recognized at the 3rd post-hatching week, presumably being formed against possible antigens invading into the lumen of the proventricular gland. On the other hand, the lymphoid masses beneath the surface epithelium and those near the duct orifice existing before the hatching period were considered to be prepared to establish the local mucosal immune barriers against the expectant antigenic invasion. PMID- 10720187 TI - A clinical study on velocity patterns of pulmonary venous flow in canine heartworm disease. AB - In this study, we evaluated methods of determining the velocity patterns of pulmonary venous flow (PVF) in dogs and then investigated the relationship of the patterns to cardiac functions in heartworm disease (HD) by transthoracic echocardiography (TTE). The results revealed that there was a good correlation between PVF patterns determined by transesophageal echocardiography (TEE) and TTE in animals lying on their left sides. The measurement of S and D wave velocities (PVS and PVD) by TTE was shown to allow clinical determination of the velocity patterns of PVF in dogs. The HD groups showed significant increases in PVS and PVD, and S and D wave time-velocity integrals (S-TVI and D-TVI) of the right cranial lobe PVF, when compared with the normal group, as determined by TTE (P<0.05). In contrast, the HD groups produced significant decreases in PVD and D TVI of the right caudal lobe PVF compared with the normal group (P<0.05), and a significant increase in the ratio of S-TVI to (S-TVI + D-TVI) (P<0.05). It is, therefore, suggested that measurement of the velocity patterns of the right cranial and caudal lobe PVF could be one method of assessing the stages of obstructive lesions in the pulmonary artery. PMID- 10720188 TI - Plasma concentrations of an angiotensin-converting enzyme inhibitor, benazepril, and its active metabolite, benazeprilat, after repeated administrations of benazepril in dogs with experimental kidney impairment. AB - In order to examine the safety of an angiotensin-converting enzyme (ACE) inhibitor in dogs with impaired renal excretion route, benazepril was administered orally, and plasma concentrations of benazeprilat, the active metabolite of benazepril, were determined in dogs with renal mass reduction (1/4th kidney) created by right-side nephrectomy and ligation of branches of the left renal arteries. Five dogs were administered benazepril orally at a given dose (0.5 mg/kg body weight) and 4 other dogs received 20 times that dose (10 mg/kg body weight) once daily for 15 consecutive days before (intact kidney period) and after (1/4th kidney period) creation of kidney impairment. Six control dogs received surgical treatment, but no drug. After creating a 1/4th kidney, plasma urea nitrogen and creatinine concentrations increased to approximately 30 mg/dl and 2.0 mg/dl, respectively, and renal plasma flow and glomerular filtration rate decreased to 37% and 30% of pre-treatment values, respectively. However, these parameters did not change significantly during the 1/4th kidney period both in the 0.5 mg/kg and 10 mg/kg groups. In the 0.5 mg/kg group, plasma benazeprilat concentrations increased to approximately 20 ng/ml to 340 ng/ml 2 hr after each administration, and there were no significant differences between the plasma benazeprilat concentrations during the intact and 1/4th kidney periods. In the 10 mg/kg group, plasma benazeprilat concentrations varied in the individual dog, but did not increase with the days of administration, and were not significantly different on each administration day between the intact and 1/4th kidney periods in either dose group. The AUCs(0-24) of plasma benazeprilat concentrations determined on the 15th administration day were not different between the intact and 1/4th kidney periods in dogs of either dose group. Plasma ACE activities decreased after drug administration in dogs of both groups. Benazepril seemed to have a high safety, and the adjustment of dosage regimen might not be needed in dogs with mild to moderate renal function impairment because the drug was excreted both from the kidneys and liver. PMID- 10720189 TI - Connective tissue-type mast cell leukemia in a dog. AB - An unusual case diagnosed as connective tissue-type mast cell leukemia with marked mastocyte infiltration into visceral organs in a seven-year-old female Curly-Coated retriever is presented. Acute circulatory collapse, emesis, diarrhea, abdominal enlargement, icterus, cyanosis, dyspnea, pulmonary edema, hepatomegary, ascites, and right ventricular enlargement were observed. Hematologic and biochemical examinations revealed mast cell leukemia, mature neutrophilia, monocytosis, thrombocytopenia, hemolytic hyperbilirubinemia, hyperhistaminemia, renal and hepatic injuries. Mast cells were distributed systemically, but predominantly in the diaphragm and liver with a large mass among the serosa of ileum, cecum and colon. Mast cells were stained intensely by both safranin and berberine sulfate. PMID- 10720190 TI - Wortmannin, a radiation sensitizer, enhances the radiosensitivity of WKAH rat cells but not that of LEC rat cells. AB - No significant cytotoxic effect was observed in WKAH rat cells by the treatment of wortmannin, a radiation sensitizer, at concentrations lower than 30 microM for 24 hr. The relative surviving fractions of LEC rat cells were slightly, but significantly, lower than those of WKAH rat cells at each concentration of wortmannin. When the wortmannin-treated WKAH rat cells were X-irradiated, the relative surviving fractions decreased in a wortmannin concentration-dependent manner. On the contrary, no significant difference was observed between the survival curves of untreated and wortmannin-treated LEC rat cells after X irradiation. PMID- 10720191 TI - Chronic myelomonocytic leukemia in a cat. AB - A 1-year-old spayed domestic short-haired cat was referred with anorexia and weight loss. Hematologic findings indicated nonregenerative anemia, severe neutropenia and monocytosis. The feline leukemia virus (FeLV) antigen test was positive reaction by enzyme-linked immunosorbent assay. Dysgranulopoiesis with slight increase in blast cells were observed in bone marrow smears. On the basis of blood and bone marrow findings, the cat was diagnosed as chronic myelomonocytic leukemia (CMMoL), which possibly corresponds to a kind of the subtypes in human myelodysplastic syndrome (MDS). PMID- 10720192 TI - Vimentin/cytokeratin coexpression foci in a well-differentiated canine hepatocellular carcinoma. AB - A number of pale-stained cell foci were observed within a well-differentiated hepatocellular carcinoma which developed in a 10-year-old male mongrel dog. The foci were composed of hepatocyte-like cells, but did not contain glycogen granules in their cytoplasm. Immunohistochemically, the focus cells coexpressed both bile duct type cytokeratin and vimentin. Electronmicroscopically, they were abundant in cytoplasmic organelles and contained bile pigments. Bile canaliculi were noted between the focus cells. The focus cells in the present case were considered to be neoplastic hepatocytes expressed bipotential features of hepatic stem cells. PMID- 10720193 TI - Changes in plasma testosterone and testicular transferrin concentration, testicular histology and semen quality after treatment of testosterone-depot plus PMSG to 3 dogs with asthenozoospermia. AB - Three dogs diagnosed as having asthenozoospermia were given three intramuscular injections of 50 mg testosterone(T)-depot plus 250 IU pregnant mare serum gonadotropin (PMSG) at 2-week intervals, and their plasma T and testicular transferrin (Tf) concentrations, testicular histology, and semen quality were examined during the period of hormone therapy. Plasma T concentrations temporarily increased, and there was a slight improvement in spermatogenesis. Increased Tf concentrations suggested that Sertoli cell function in all three dogs was promoted by hormone treatment. The results showed that semen quality, especially the percentages of motile sperm and abnormal sperm, were improved between 1 and 5 weeks after the start of T-depot plus PMSG treatment. PMID- 10720194 TI - Sandwich enzyme-linked immunosorbent assay of canine leptin. AB - Leptin is the ob gene product secreted from adipocytes in mammals, and thereby its plasma level reflects body fat content. To establish an assay method for leptin in the dog, rabbit anti-canine leptin antibody was obtained using canine recombinant leptin as an antigen. This antibody reacted to canine leptin much stronger than mouse, rat and human leptins. Sandwich enzyme-linked immunosorbent assay (ELISA) using this antibody was developed. The serum leptin levels of 13 healthy dogs were in a range from 1.4 to 5.6 ng/ml with the mean +/- SEM of 3.0 +/- 0.3 ng/ml. PMID- 10720195 TI - The effect of orally administered cisapride on intestinal motility in conscious horses. AB - Seven Thoroughbred horses were laparotomized and Force Transducers were fixed on the proximal jejunal and cecal serosa. After observation of the digestive tract motility in consciousness, cisapride (0, 0.5, 0.75 or 1 mg/kg) was orally administered. In horses treated with 0.75 mg/kg or 1.0 mg/kg cisapride, the migrating contraction (MC) of the jejunum was significantly increased in frequency. PMID- 10720196 TI - Pathogenicity of a new neurotropic equine herpesvirus 9 (gazelle herpesvirus 1) in horses. AB - Pathogenicity of equine herpesvirus 9 (EHV-9), a new type of equine herpesvirus isolated from Gazella thomsoni, in horses was investigated by intranasal inoculation of EHV-9 (10(7) pfu) to two conventionally reared 8-months old half bred weanling horses. Fever higher than 39 degrees C was recorded. Virus was recovered from nasal swabs and peripheral blood mononuclear cells. Both horses developed neutralizing antibody to EHV-9. Perivascular infiltration of mononuclear cells and glial reaction were found in the olfactory and limbic systems. The results suggested that EHV-9 has a pathogenicity in horses. PMID- 10720197 TI - Correlation between maternal serum antibodies and protection against bovine rotavirus diarrhea in calves. AB - The correlation between maternal serum antibodies in beef calves at 2 days old and protection against diarrhea induced by natural bovine rotavirus (BRV) infection was examined. Virus neutralizing (VN) antibody titers against BRV in sera from calves that developed diarrhea by BRV infection within 14 days of age (BRV-diarrheal calves) were significantly lower than those from calves that had no diarrhea. In the BRV-diarrheal calves, a positive correlation was found between the VN antibody titers and age of the onset of diarrhea. There were negative correlations between the VN antibody titers and duration of the diarrhea, VN antibody titers and cumulative diarrhea scores, and the VN antibody titers and duration of virus shedding. These results suggest that the VN antibody titers against BRV in newborn calf serum could be an indicator of protection against BRV-induced diarrhea. PMID- 10720198 TI - Green fluorescent protein gene insertion of Sendai Virus infection in nude mice: possibility as an infection tracer. AB - The green fluorescent protein (GFP) marker from jellyfish Aequorea victoria is considered to have potential use in the study of host-pathogen relationships, by tracing infections in living cells, organs and animals. We compared the pathogenicity of Sendai virus with an inserted GFP gene (GFP-SeV) with that of its wild-type (Wt-SeV) to determine the usefulness of the recombinant virus in long-term infection of BALB/c nude (nu/nu) mice. The results indicated that the presence of GFP in infected cells could be analyzed easily and sensitively. GFP helped in identifying and in understanding the cellular sites of viral replication in vitro and in vivo. However, the GFP insertion into the Wt-SeV genome, led to decreased pathogenicity, altering the in vivo viral kinetics. PMID- 10720199 TI - Adenovirus-mediated gene transfer of a truncated form of fibroblast growth factor receptor inhibits growth of glioma cells both in vitro and in vivo. AB - Basic fibroblast growth factor (FGF-2) and high affinity FGF receptor (FGFR) have been detected in the nucleus as well as the cytoplasm of many human gliomas, and are known to stimulate cellular proliferation and angiogenesis in the tumors. To investigate the effects of inactivation of FGFR on the growth of malignant gliomas, we constructed a replication-deficient recombinant adenovirus vector encoding a truncated form of chicken FGFR1 (AxCA delta FR). AxCA delta FR infected cells were confirmed to express truncated FGFR protein by immunoblotting and FGF-2-dependent clonogenicity of NIH3T3 cells was suppressed by infection with this virus vector. Then human malignant glioma cell lines U-251MG and T98G, both of which have been reported to express FGF-2 and FGFR, were infected with AxCA delta FR. These infected cells showed nuclear as well as cytoplasmic expression of a truncated FGFR protein. Proliferation rate and the ability to form colonies in soft agar of the cells infected with this virus vector were significantly suppressed compared with those of uninfected and lacZ-expressing adenovirus-infected cells. Moreover, intratumoral injection of AxCA delta FR significantly suppressed the subcutaneous tumor growth of the glioma cells in nude mice. We concluded that inactivation of the cytoplasmic and nuclear FGFR using this truncated FGFR-expressing adenovirus vector can inhibit the growth of malignant gliomas both in vitro and in vivo. PMID- 10720200 TI - Human astrocytomas co-expressing Fas and Fas ligand also produce TGFbeta2 and Bcl 2. AB - Human astrocytomas frequently co-express Fas (APO-1/CD95) and Fas ligand (FasL), yet do not appear to be overly susceptible to suicidal, fratricidal and immune mediated elimination. This suggests that these gliomas have acquired mechanisms to prevent Fas-mediated apoptosis from occurring. Candidates for such a role include transforming growth factor-beta2 (TGFbeta2) and B-cell lymphoma/leukemia 2 (Bcl-2). TGFbeta2 effectively functions by hiding tumor cells from the immune system. This may potentially prevent the delivery of FasL from cytolytic T cells to Fas bearing astrocytomas. Bcl-2 works by rendering gliomas resistant to Fas mediated apoptosis. Using immunohistochemistry, we analyzed seventy-six human astrocytomas (11 World Health Organization (WHO) grade I, 17 grade II, 17 grade III, and 31 grade IV) for the expression of Fas, FasL, TGFbeta2 and Bcl-2 in vivo. Positive immunoreactivity was found to significantly increase with increasing tumor grade for Fas (p < 0.0002), FasL (p < 0.0001), TGFbeta2 (p < 0.001) and Bcl-2 (p < 0.01). In addition, Fas/FasL co-expression, a counter intuitive combination of factors in regards to glioma survival, also increased with WHO grade. Forty-five of 76 (59%) astrocytomas co-expressed Fas and FasL. Of those co-expressing Fas and FasL, 44 of 45 (98%) produced TGFbeta2, and 26 of 45 (58%) expressed Bcl-2. We found a significant positive correlation between Fas/FasL co-expression and TGFbeta2 (p < 0.002) and Bcl-2 (p < 0.005) production. We conclude that Fas and FasL are frequently co-expressed in human astrocytomas and these tumors are likely to produce other immunosuppressive and antiapoptotic factors such as TGFbeta2 and Bcl-2. PMID- 10720201 TI - Proliferation of cultured human astrocytoma cells in response to an oxidant and antioxidant. AB - The role of reactive oxygen species (ROS) in initiation, promotion and progression of several (lung, skin, colon, bladder, breast) tumors is well documented. Indirect evidence for ROS involvement in tumor proliferation is provided by numerous in vivo and in vitro studies that show antioxidants inhibit tumor proliferation. However, despite strong epidemiological and experimental support for ROS involvement in brain tumor proliferation, to date little is known about the role of ROS in brain tumor promotion at a cellular level. In the present study ROS involvement in proliferation of a cultured, human astrocytoma cell line (U373-MG) was tested by studying effects of an oxidant (hydrogen peroxide, H2O2), and an antioxidant (N-acetylcysteine, NAC) on astrocytoma on proliferation of these cultured cells. Proliferation was assessed by evaluating changes in cell counts and DNA synthesis. Results from these experiments clearly indicate that NAC inhibits tumor cell proliferation and DNA synthesis induced by both serum and H2O2 (10(-5) M). NAC alone did not have any significant effects on the proliferation of serum-starved cells. Thus, ROS are capable of inducing proliferation in cultured astrocytoma cells and antioxidants block ROS- and serum induced proliferation. Further investigation using primary cultures and animal models will be needed to substantiate the therapeutic potential of antioxidants in future brain tumor therapy. PMID- 10720202 TI - Effects of radiation on a model of malignant glioma invasion. AB - We sought to characterize the effects of radiation alone and in combination with BCNU and dexamethasone on malignant glioma invasion. A model of malignant glioma invasion into a gel matrix of collagen type I was used to characterize response to radiation treatment for four malignant glioma cell lines (C6, U251, U373, A172) and nine primary human glioblastoma explants. A radiation dose dependent inhibition of invasion was noted for the C6 astrocytoma cell line but not the other cell lines or explants. Addition of BCNU and dexamethasone to radiation produced additional inhibition of invasion among the cell lines and explants but could not suppress invasion entirely. PMID- 10720203 TI - Intrathecal busulfan treatment of human neoplastic meningitis in athymic nude rats. AB - The current study was designed to evaluate the toxicity and activity of Spartaject Busulfan, a microcrystalline preparation of busulfan, following its intrathecal administration into a nude rat model of human neoplastic meningitis. Animals were treated through permanent indwelling subarachnoid catheters. Human glioma D-456 MG growing in the subarachnoid space was treated with 8.1 micromol of intrathecal Spartaject Busulfan. Single-dose therapy was also subsequently compared with 4 doses of 8.1 and 2.0 micromol busulfan, respectively, against D 456 MG neoplastic meningitis. Additional experiments evaluated a saline control versus 8.1 micromol x 1, 6.2 micromol x 4 and 4.1 micromol x 4, respectively, against D-456 MG. A single dose of 8.1 micromol of intrathecal Spartaject Busulfan resulted in an increase in median survival of 61.7% compared with the saline control. In experiment 2, all busulfan treatments showed increases in median survival of 142.8% (8.1 micromol x 1), 52.3% (2.0 micromol x 4), and 23% (8.1 micromol x 4) (p < 0.001 for all groups) compared with the saline control. These results suggest that a narrow therapeutic dose range for both toxicity and activity has been defined for intrathecal busulfan in the treatment of human neoplastic meningitis in athymic nude rats. Although busulfan has only limited activity against solid tumors, the high doses achievable in the CSF following intrathecal administration coupled with the steep dose-response relationships of alkylating agents, provide rationale for further evaluation of this agent. PMID- 10720204 TI - CNTF and its receptor subunits in human gliomas. AB - Ciliary neurotrophic factor (CNTF) promotes the survival of various neuronal cell populations. It is produced by astrocytes and influences the development and differentiation of glial cells. CNTF and related neuropoietic cytokines affect growth and differentiation of various neoplasms. Moreover, they induce the reactive transformation of astrocytes (gliosis) and influence growth and differentiation of neuroectodermal tumor cell lines in vitro. However, their role in gliomas is largely unknown. We studied the expression of CNTF and its receptor subunits in human astrocytomas and glioblastomas. In more than 95% of the tumors, CNTF transcripts were found by RNAase protection assay; in more than 80% of the cases, tumor cells were CNTF immunoreactive. CNTF receptor alpha (CNTFR alpha), the specific component of the tripartite CNTF receptor system, was detectable by Northern blot analysis in 80% of the cases. In situ hybridization revealed CNTFR alpha mRNA in the cytoplasm of neoplastic cells. Transcripts of the remaining two components of the CNTF receptor system, gp130 and LIFR beta, were found by Northern blotting in 83% and 70% of the tumors, respectively. Simultaneous expression of CNTF and all its receptor components was detected in approximately half of the tumors. These results indicate that CNTF and its receptor components are expressed by human glioma cells. The simultaneous expression of ligands and receptor subunits suggests that CNTF might act on human glioma cells via an auto- or paracrine mechanism. PMID- 10720205 TI - Analysis of proliferation and apoptosis in brain gliomas: prognostic and clinical value. AB - With the introduction of new (immuno-)histochemical techniques it is now possible to assess rates of proliferation and apoptosis in brain gliomas using archival paraffin embedded material. As proliferation and apoptosis are related to tumour growth rate quantification of these processes has prognostic value and is related to tumour grading. In this study we assessed the proliferation rate by measuring the Ki-67 labelling index using the MIB-1 antibody (MIB-LI) and the apoptotic rate using the in situ labelling of DNA strand breaks with TUNEL (TUNEL-LI) in 315 supratentorial gliomas. MIB-LI and TUNEL-LI in astrocytomas (A) where significantly lower compared to anaplastic astrocytomas (AA), glioblastomas (GBM) and oligodendroglial tumours [oligodendrogliomas (O) and anaplastic oligodendrogliomas (AO)]. MIB-LI and TUNEL-LI were significantly lower in AA compared to GBM. In astrocytic tumours MIB-LI and TUNEL-LI appeared to be correlated. As the distinction between A and AA is of clinical value but can be difficult histomorphologically we analysed the prognostic value of MIB-LI and TUNEL-LI in gliomas with particular emphasis on A and AA. MIB-LI below 10% was of prognostic value in A and AA, O and AO but not in GBM on univariate survival analysis. TUNEL-LI was of no prognostic value. With multivariate survival analysis MIB-LI lost prognostic significance in O and AO. Astrocytomas with a gemistocytic component (AG) are similar to A with respect to survival and MIB-LI and TUNEL-LI. MIB-LI is of independent prognostic value in A and AA. Assessment of MIB-LI in A and AA can be used as an aid in distinguishing A and AA. PMID- 10720206 TI - Adenoid cystic carcinoma metastatic to the dura: report of two cases. AB - Adenoid cystic carcinoma (ACC) originating in the salivary and lacrimal glands usually spreads to the intracranial space by following cranial nerves into the cavernous sinus, temporal bone and cerebellopontine angle. We present two cases in which ACC metastasized extensively to the dura, suggesting that ACC has an affinity for the dura. Case 1, a 43-year-old man, was operated on 12 years earlier for invasive ACC of the right palate. He experienced recurrence of the tumor in the left cavernous sinus and sella, and extensive involvement of the dura of both right and left temporal fossae. Case 2, a 33-year-old woman, had spread of ACC to the right convexity dura and tentorium after undergoing a resection of a left-sided ACC tumor of the lacrimal gland two years earlier. Both patients underwent multiple resections and radiation treatment. Extensive, multifocal, bilateral spread of ACC to the dura in both cases indicates that ACC has an affinity for the dura. PMID- 10720207 TI - Diagnostic accuracy of MRI compared to CCT in patients with brain metastases. AB - OBJECTIVES: In patients with extracranial neoplasms, the occurrence and number of brain metastases (BM) are critical for further diagnostic approaches and therapeutic strategies and the patient's prognosis. Although widely accepted, there is surprisingly little evidence in the literature that MRI is superior to CCT. Therefore, in patients with solitary BM according to diagnostic contrast enhanced computed tomography (CCT), we investigated, what additional information could be gained by contrast-enhanced magnetic resonance imaging (MRI). METHODS/RESULTS: Among 55 patients with solitary BM according to CCT, 17 had multiple BM on MRI (31%) and 38 had solitary BM in both. Based on a presumed binomial distribution of our data, we calculated a rate of at least 19% of patients with solitary BM on CCT, in which MRI would show multiple lesions (p = 0.05). The two main characteristics for BM missed by CCT were the smaller diameter, which averages 2 cm less than in BM identified with both modalities, and a preferential frontotemporal location. CONCLUSION: MRI is indeed superior to CCT in the diagnosis of BM the essential reasons besides detection of smaller lesions being a better soft tissue contrast, significantly stronger enhancement with paramagnetic contrast agents, the lack of bone artifacts, fewer partial volume effects, and direct imaging in three different planes. Therefore, MRI is indispensable in the diagnostic workup of patients with BM for choosing the optimum therapeutic approach, especially with regard to the decision whether to operate or to primarily irradiate the patient's metastases. PMID- 10720208 TI - Radiation-induced blood-brain barrier damage in astrocytoma: relation to elevated gelatinase B and urokinase. AB - Successful management of brain tumors prolongs life, raising the risk of delayed injury secondary to the treatment. Radiation therapy, a mainstay of brain tumor treatment, can damage the cerebral blood vessels. Acutely a breakdown of the blood-brain barrier (BBB) may be seen, but fibrosis complicates radiation injury in the chronic phase. Matrix metalloproteinases (MMPs) and plasminogen activators are two matrix-degrading proteolytic enzymes, which are induced by radiation. They disrupt the basal lamina around cerebral capillaries and open the BBB. We report a patient with an astrocytoma managed by partial resection and external beam irradiation to maximal tolerable doses. The patient later developed malignant brain edema shortly after stereotactic radiosurgery. Tissue obtained during surgical debulking to control the edema showed very high levels of gelatinase B (92 kDa type IV collagenase) and urokinase-type plasminogen activator (uPA). Tumor cells were absent from the biopsy and subsequent autopsy specimens, but necrosis with fibrosis of the blood vessels was seen. If abnormal matrix enzyme function participates in the expression of radiation injury, then inhibitors to such enzymes may provide one strategy for controlling cerebrovascular damage after therapeutic brain radiation. PMID- 10720209 TI - Suppressors of cytokine signalling (SOCS): putative modulators of cytokine bioactivity in health and disease. AB - Cytokines are important modulators of homeostatic processes such as development, haematopoiesis and host defence. A recently identified family of proteins, the supressors of cytokine signalling (SOCS) act as negative regulators of the key cytokine-activated signalling pathway, the Janus kinase/signal transducers and activators of transcription (JAK/STAT) cascade. In the current review, the discovery, structural features, regulation of expression, mechanisms of JAK/STAT inhibition and putative role in health and disease of the SOCS family are discussed. PMID- 10720210 TI - Thin glomerular basement membrane disease. AB - The term thin glomerular basement membrane disease (TBMD) refers to a condition characterised by thinning of the GBM at electron microscopy examination and, clinically, by isolated hematuria, frequently occurring in other family members, with no extra-renal manifestations. Progression towards chronic renal failure (CRF), although rare, has been reported and blood pressure is high in 30-35% of cases during follow-up. TBMD is generally considered different from Alport syndrome since immunohistological investigation does not show abnormalities of type IV collagen alpha chains in the GBM, as frequently observed in Alport patients; moreover, in familial cases, the disease is transmitted as autosomal dominant trait, rarely observed in Alport syndrome. Genetic studies suggest that TBMD is a heterogeneous disease, but some cases may be related to mutations of COL4A3/COL4A4 genes, thus belonging to the spectrum of type IV collagen diseases. TBMD may arise with other glomerular diseases, most frequently IgA nephropathy, and it remains to be established whether these cases are a casual occurrence or whether a thinner than normal GBM predisposes to immune complex deposition. PMID- 10720211 TI - Molecular biology of 5,10-methylenetetrahydrofolate reductase. AB - Methylenetetrahydrofolate reductase (MTHFR) plays a central role in the folate cycle and contributes to the metabolism of the amino acid homocysteine. It catalyzes the reduction of 5,10-methylenetetrahydrofolate to 5 methyltetrahydrofolate, thus generating the active form of folate required for remethylation of homocysteine to methionine. Deficiency of MTHFR may be associated with an increase in plasma homocysteine, which in turn is associated with an increased risk of vascular disease. This article summarizes the biochemistry, the function in the folate cycle, and the molecular genetics of this enzyme. Particular emphasis has been given to the role of two common polymorphisms (MTHFR 677C-->T, 1298A-->C) in cardiovascular disease, cerebrovascular disease, venous thrombosis, longevity, neural tube defects, pregnancy/preeclampsia, diabetes, cancer, psychiatry, renal failure and renal replacement therapy. Finally, the rare genetic defects underlying severe MTHFR deficiency are also considered. PMID- 10720212 TI - The effect of hemodiafiltration with on-line endogenous reinfusion (on-line HFR) on anemia: design of a European, open, randomised, multicentre trial. European Collaborative Study. AB - The lack of backfiltration reduces plasma levels of C-reactive protein and interleukin-6. Paired filtration dialysis is the hemodialfitration technique that abolishes backfiltration. By physically separating convection from diffusion, it allows pure ultrafiltrate to be available during the entire session, so the ultrafiltrate can be regenerated and used as infusion fluid. On these premises, we have developed a European, open, randomised, multicentre study aimed at evaluating the effect of hemodiafiltration with on-line endogenous reinfusion (on line HFR) on anemia. At least 130 chronically uremic hemodialysed (bicarbonate hemodialysis) stable patients with mild anemia (Hb between 9 and 11 g/dL) will be enrolled and normalized for iron stores by concomitantly repleting iron deposits (if ferritin <300 microg/L) and reducing the dose of erythropoietin to maintain Hb values within the range at enrollment (9-11 g/dL). Patients will be included in the study, randomized to the two treatments (test treatment: on-line HFR; control treatment: hemodiafiltration or modified forms) and followed up for nine months. Iron stores will be maintained within normal levels and the dose of erythropoietin will be kept constant. The primary question and response variable will be the mean monthly changes in hemoglobin levels over the period of nine months. As secondary questions and response variables, we will measure the nutritional status using a subjective global assessment, protein catabolic rate, urea generation rate and the dietician's assessment, serum concentrations of vitamins A, C, E and serum C-reactive protein. PMID- 10720213 TI - The cost of knowledge in nephrology and the importance of clinical databases. AB - The modern clinician uses knowledge generated on the solid ground of clinical epidemiology as a basis for clinical practice. Doctors play a leading role in continuous quality improvement of medical services, working to find and validate appropriate clinical performance measures. Large clinical databases are emerging as an important research tool for testing the performance of clinical policies in patients with chronic renal diseases. New strategies have been devised to help the clinical investigator decide when to base clinical policies on observational or randomized trial data. In the next few years the high motivation and consensus required in clinical research projects based on large data bases will certainly be created and the tantalizing managerial problems resolved. PMID- 10720214 TI - Prediction and consistency of blood pressure salt-sensitivity as assessed by a rapid volume expansion and contraction protocol. Salt-Sensitivity Study Group of the Italian Society of Hypertension. AB - BACKGROUND: This multicenter trial in essential hypertensive patients (n=94) is aimed i) to evaluate the distribution of blood pressure salt-sensitivity by a rapid volume expansion/contraction protocol over three days; ii) to investigate the within-patient reproducibility and to identify predictors of the response to the test; iii) to compare this response with the response to dietary NaCl restriction. METHODS: The study design included: 1) screening for salt sensitivity by the rapid test; 2) a controlled trial of dietary salt restriction; 3) repetition of the rapid test in a subgroup of patients. RESULTS: The mean BP response to the rapid test fitted a Gaussian curve. In multivariate regression analysis, controlling for the effect of potential confounders, the blood pressure increment during the intravenous saline infusion was the best independent predictor of the response to the test (r=0.713) with minor contributions by the 24-h urinary sodium excretion before the test and by baseline fasting serum insulin. These three variables together explained 61% of the overall variability of the response. The Spearman rank correlation coefficient between the BP response to the rapid test and the response to the dietary protocol was 0.21, p=0.05. Upon repetition of the rapid test, the correlation coefficient between the responses observed on the two occasions was 0.60 (n=19, p<0.01); there were no patients misclassified across the extreme tertiles of the distribution of salt sensitivity. CONCLUSION: We conclude that the rapid test reproducibly identified patients in the upper and lower parts of the distribution of salt sensitivity. The analysis of possible predictors of the response to the test suggested that the evaluation of the blood pressure response to saline infusion, upon careful standardization of dietary NaCl intake, may represent an alternative to the completion of the whole test for the screening of the salt-sensitivity. PMID- 10720215 TI - Plasma parathyroid hormone, phosphatemia and vitamin D receptor genotype: are they interrelated? AB - It is not clear how the rate of bone mineral loss and vitamin D receptor (VDR) Bsml polymorphism in hemodialysed patients are related. We therefore analysed the relationships between indices of calcium-phosphate metabolism in respect to VDR genotype in 180 hemodialysed patients. We measured plasma concentrations of calcium, phosphate, iPTH, 1,25(OH)2D3 and activity of the bone fraction of alkaline phosphatase on the day before dialysis. VDR genotype BB, Bb and bb were found in 39, 84 and 57 patients, respectively. The allele frequency was B 0.45 and b 0.55. Subjects with BB genotype had insignificantly higher plasma levels of phosphate, iPTH and activity of the bone fraction of alkaline phosphatase, but significantly lower (p<0.02) concentrations of 1,25(OH)2D3 [iP (mmol/l): 2.05+/ 0.09, 1.98+/-0.06, 1.93+/-0.06; iPTH (pg/ml): 257+/-50, 229+/-24, 219+/-30; AP(BF) (nmol/l/s): 515+/-45, 477+/-27, 457+/-34; 1,25(OH)2D3 (pg/ml): 23.4+/-1.5, 26.2+/-1.0, 29.3+/-1.3, for BB, Bb and bb respectively]. The strongest significant correlation between phosphatemia and iPTH was in the BB subgroup (r=0.343). Moreover, only in this subgroup did phosphatemia significantly contribute to the increase in iPTH, assessed by multiple regression analysis. In conclusion, it seems likely that BB VDR genotype in HD patients contributes to the severity of secondary hyperparathyroidism by a mechanism involving phosphatemia. PMID- 10720216 TI - Serum levels of soluble interleukin-2 receptor in patients with ANCA-associated vasculitis. AB - Serum soluble interleukin-2 receptor (sIL-2R) concentrations were determined using the ELISA method in 19 cases of ANCA-associated vasculitis. These patients were classified as 7 cases of Wegener's granulomatosis (WG) and 12 cases of microscopic polyangiitis (MPA). Elevated levels of sIL-2R were present in the sera of these patients. Levels of serum sIL-2R were not significantly different in patients with WG and MPA either in the active or inactive phase, so the results were expressed as a unified ANCA associated vasculitis group. Concentrations of serum sIL-2R were significantly higher in ANCA-associated vasculitis during the active phase than during the inactive phase (p<0.05), and serum sIL-2R levels were significantly increased in these patients, in the active or inactive stage, compared with a group of healthy subjects (p<0.05). In patients with vasculitis, serum sIL-2R levels correlated with serum levels of C reactive protein (p<0.05). In the active phase, concentrations of serum sIL-2R correlated to creatinine concentrations. No correlation was found between sIL-2R and ANCA levels in any of the stages of the disease. These findings suggest cellular immune activation in ANCA associated vasculitis. PMID- 10720217 TI - High prevalence of antithyroid antibodies in anti-glomerular basement membrane antibody-mediated disease. AB - Anti-glomerular basement membrane antibody (anti-GBM Ab)-mediated disease and autoimmune thyroiditis are characterized by the presence of organ-specific antibodies. The diagnosis of autoimmune thyroiditis is usually based on the presence of serum antithyroid antibodies. Few studies have addressed the relationship between anti GBM-Ab mediated disease and autoimmune thyroid pathology. Given that this disorder is often asymptomatic, associations of the two pathologies may be under-diagnosed. This study investigated the prevalence of serum antithyroid antibodies (antithyroglobulin (anti-TG) and anti-thyroid peroxidase (anti-TPO)) in patients with anti-GBM Ab-mediated disease. Antithyroid antibodies presence was investigated in sera from 35 patients in whom anti-GBM Ab mediated disease had been diagnosed. Anti-glomerular basement membrane antibodies and anti-thyroid antibodies (anti-TG and anti-TPO) were assayed using an enzyme linked immunosorbent assay. Forty-five percent of patients with anti-GBM Ab mediated disease (16/35) had positive antithyroid antibody titers. Eighteen percent (3/16) suffered from subclinical hypothyroidism. In conclusion, the high prevalence of antithyroid antibodies in these patients suggests a possible pathogenic link between autoimmune thyroiditis and anti GBM Ab-mediated disease. PMID- 10720218 TI - GBV-C/HGV infection in end-stage renal disease: a serological and virological survey. AB - Patients with end-stage renal disease (ESRD) appear to be at high risk for GBV C/HGV infection. This information has been obtained with virological techniques (RT-PCR) but few serological data exist. A prototype enzyme immunoassay has now been developed to detect antibodies against the putative envelope protein (E2) located on the surface of the GBV-C/HGV virion particle. We studied the prevalence of GBV-C/HGV infection, as detected by RT-PCR and anti-E2 GBV-C/HGV antibody, in a cohort of chronic dialysis patients (n=157) and renal transplant (RT) recipients (n=77); as a control group, 136 healthy blood donors were tested. The total prevalence of GBV-C/HGV in ESRD was 23% (54/234). The frequency of GBV C/HGV viremia was 7.7% (18/234) in ESRD and 4.4% (3/68) among healthy blood donors; the prevalence of anti-E2 GBV-C/HGV was 15% (36/234) and 8.8% (12/136) in ESRD and controls, respectively. No relationship was seen between anti-E2 GBV C/HGV antibody (or GBV-C/HGV viremia) and age, sex, time on renal replacement therapy, anti-HCV, HBsAg and transfusion requirement. No statistical association was observed between GBV-C/HGV and AST/ALT activity. Two of 54 GBV-C/HGV positive patients (3.7%) had raised ALT but were negative for HBV/HCV. In the majority of patients (35/36, 97%) the presence of anti-E2 GBV-C/HGV antibody was linked with the loss of GBV-C/HGV viremia from serum. In conclusion, GBV-C/HGV infection, as detected by RT-PCR and anti-E2 antibody, was common in ESRD, and the rate of infection was higher than in controls. No association was seen between GBV-C/HGV and various demographic or clinical factors. A small group of GBV-G/HGV positive patients tested negative for HBV/HCV and had raised ALT. In many patients exposed to GBV-C/HGV infection the virus was cleared. The clinical significance of GBV C/HGV in ESRD remains controversial. Prospective studies with additional serological assays are in progress. PMID- 10720219 TI - Delivery of healthy infant during hemodialysis session. AB - Acute renal failure secondary to bilateral ureteral obstruction in pregnancy is rare. We describe a case of acute renal failure secondary to bilateral ureteral obstruction. A 27 year-old woman at 35 weeks' gestation was referred to our hospital with a diagnosis of acute renal failure. The patient had been well until four days earlier, when she developed an abrupt anuria. She had been administrated excessive amounts of fluids, and unresponsive to parenteral furosemide. She had mild pitting oedema and an S3 gallop with crackles in the lungs. The uterus was enlarged to the expected size with a cervical dilatation of 2 cm in diameter. Her serum creatinine level was 7.0 mg/dl. Renal ultrasound showed bilateral hydronephrosis of severe degree. The patient was immediately hemodialyzed for advanced renal failure with hypervolemia, and a healthy infant was born at the third hour of the HD session without any complication. On the next day, her urine volume was 200 ml/day and serum creatinine level was 6.8 mg/dl. For this reason, the patient underwent cystoscopy and ureteral stents were inserted bilaterally. There was no evidence of ureteral stones or obstructive lesions. After the stenting, the urine volume increased and serum creatinine was decreased gradually to normal level at the seventh day of postpartum. Two weeks later ureteral stents were removed and both infant and patient were completely healthy. To the best of our knowledge, this is the first case of delivery of an infant during a haemodialysis session. PMID- 10720220 TI - The treatment of IgA nephropathy. PMID- 10720221 TI - Computer simulation for the convenient optimization of isocratic reversed-phase liquid chromatographic separations by varying temperature and mobile phase strength. AB - Software is described which allows the rapid development of separations by means of isocratic reversed-phase liquid chromatography (RP-LC), based on the optimization of column temperature (T) and mobile phase strength (%B). For a given sample, four initial experiments are carried out at two different temperatures, using either isocratic or (better) gradient elution. If isocratic experiments are chosen for computer simulation, it is necessary to select appropriate values of %B for these initial runs. Literature data for solute retention as a function of T are reviewed, as a basis for estimating suitable values of %B at the two values of T selected. The use of optimized values of T and %B led to acceptable separations for three representative samples. The prediction of isocratic separation on the basis of initial gradient experiments is more convenient than the use of initial isocratic experiments, but less reliable. When gradient experiments are used, one additional isocratic experiment can improve the accuracy of such predictions by a "reflection" procedure. The latter approach was confirmed for predictions of both isocratic and gradient separation from initial gradient experiments. PMID- 10720222 TI - Modelling retention in liquid chromatography as a function of solvent composition and pH of the mobile phase. AB - The aim of this work was to develop a model that accurately describes retention in liquid chromatography (LC) as a function of pH and solvent composition throughout a large parameter space. The variation of retention as a function of the solvent composition, keeping other factors constants, has been extensively studied. The linear relationship established between retention factors of solutes and the polarity parameter of the mobile phase, E(N)T, has proved to predict accurately retention in LC as a function of the organic solvent content. Moreover, correlation between retention and the mobile phase pH, measured in the hydroorganic mixture, can be established allowing prediction of the chromatographic behavior as a function of the eluent pH. The combination of these relationships could be useful for modelling retention in LC as a function of solvent composition and pH. For that purpose, the retention behavior on an octadecyl silica column of a group of diuretic compounds covering a wide range of physico-chemical properties were studied using acetonitrile as organic modifier. The suggested model accurately describes retention of ionizable solutes as concomitant effects of variables included and is applicable to all solutes studied. We also aimed to establish an experimental design that allows to reproduce to a good approximation the real retention surface from a limited number of experiments, that is from a limited number of chromatograms. Ultimately, our intention is to use the model and experimental design for the simultaneous interpretive optimization of pH and proportion of organic solvent of the mobile phase to be used in the proposed separation. PMID- 10720223 TI - Relationship between the retention characteristics on an alumina column and physico-chemical parameters of some environmental pollutants. AB - The retention behavior of 16 environmental pollutants was studied on alumina and porous graphitized carbon (PGC) columns using n-hexane as eluent. The relationship between the logarithm of the capacity factors determined on the alumina column and the physico-chemical characteristics of the solutes was elucidated by principal component analysis (PCA) followed by two-dimensional nonlinear mapping. The 12 original variables can be reduced to four with only a 10% loss of information. The logarithm of the capacity factor formed a cluster with the hydrogen donor and acceptor properties of the solutes and their Taft's constant on the two-dimensional nonlinear map of PC loadings indicating that both steric and electronic parameters play a considerable role in the retention mechanism on alumina support. Alcohols, aromatics and chlorinated alkanes formed separate clusters on the two-dimensional nonlinear map of PC variables suggesting that their retention mechanism may be different on the alumina column. Solutes were not retained on the PGC surface proving that under normal-phase conditions the retention capacity of alumina can be higher than that of PGC. PMID- 10720224 TI - Lipophilicity characterization by reversed-phase liquid chromatography of some furan derivatives. AB - Lipophilicity is one of the properties which influences the partition of a substance in biological media. The present study reports on the chromatographic behaviour of a newly synthesised series of furan derivatives by RP-HPLC and RP TLC, with methanol-water and acetonitrile-water as mobile phases, in order to establish if the linear relationships between the retention parameters (log k, R(M)) and the concentration of organic modifier in the mobile phase, phi, allows the extrapolation procedure. Good correlations between the retention parameters were obtained by RP-HPLC and RP-TLC, and the concentration of organic modifier (methanol, acetonitrile) in the mobile phase was established for the studied furan derivatives. However, for the discussed compounds, acetonitrile has a lower sensitivity to changes in the structures. A good correspondence was obtained between the extrapolated parameters for the methanol-water mobile phase when using RP-HPLC and RP-TLC. However, stronger interactions occur in RP-TLC between the compounds and the residual silanol groups than in RP-HPLC. PMID- 10720225 TI - Adsorption/partition model of liquid chromatography for chemically bonded stationary phases of the aliphatic cyano, reversed-phase C8 and reversed-phase C18 types. AB - A novel approach was introduced to modeling solute retention in the liquid chromatography systems, employing silica-based aliphatic chemically bonded stationary phases of the cyano, reversed-phase C8 and reversed-phase C18 types, and the mixed binary eluents most frequently used in the reversed-phase and normal-phase chromatography modes (i.e. using the methanol-water and the 2 propanol-n-hexane liquid mixtures, respectively). This approach takes notice of the mixed (adsorption/partition) mechanism of solute retention, in which both, the adsorptive and the dispersive forces contribute to the overall energetics of this process. Performance of our new model was compared with that of the widely recognized and on a routine basis applied Schoenmakers approach, and it was found out that both models perform with a practically equal and outstanding accuracy. PMID- 10720226 TI - Retention mechanism, isocratic and gradient-elution separation and characterization of (co)polymers in normal-phase and reversed-phase high performance liquid chromatography. AB - Synthetic (co)polymers or (co)oligomers with two (or more) repeating groups show not only molar mass distribution, but also composition and sequence distribution of the individual repeat units. To characterize such two- (or more-) dimensional distribution, liquid chromatography under "critical conditions" has been suggested, where the separation according to one type of repeating units is suppressed by balancing the adsorption and the size-exclusion effects. In present work it is shown that by combination of adequately selected separation conditions in normal-phase and in reversed-phase systems, the two-dimensional distribution mode can be adjusted to result in the separation following the distribution of any of the two repeat units in ethylene oxide-propylene oxide block (co)oligomers. Based on the retention mechanism suggested, prediction and optimization of the conditions for isocratic and gradient-elution separations of (co)oligomers is possible. HPLC-MS with atmospheric-pressure chemical ionization is a valuable tool for unambiguous identification of the individual (co)oligomers and their tracking in course of method development. Gradient elution can be used for the separation and characterization of block (co)oligomers of ethylene oxide (EO) and propylene oxide (PO) according to the number of the units in one block, while the separation according to the distribution of the units in the other block is suppressed. The effects of the arrangement of the individual EO and PO blocks in the block (co)oligomers (the sequence distribution) affects significantly the retention behavior and the selection of the optimum separation conditions. PMID- 10720227 TI - Investigations of artifact peaks in sensitive high-performance liquid chromatography methods. AB - Chromatograms of sensitive HPLC methods often show disturbing peaks, which mostly can be traced back easily. But sometimes such peaks are difficult to reproduce and therefore hard to trace. This paper describes two cases where artifact peaks arose due to contamination by the septum and by the sampling equipment, respectively, leading to misinterpretation of impurities and erroneous quantification. Furthermore, troubleshooting and ways to overcome these deficiencies are outlined. PMID- 10720228 TI - Coupling of chromatographic ion-exchange reaction with change-partner reaction. Presumed mechanism on the ion-ion interaction of inorganic ions in Sephadex G-15 columns. AB - A sample solution containing sodium or potassium dihydrogenphosphate or disodium or dipotassium hydrogenphosphate was eluted from a Sephadex G-15 column with either sodium or potassium chloride solution in various sample-eluent systems, and every one of the four kinds of ions employed was determined in the eluate. Then, the elution profiles showed the phenomenon that a negative peak of chloride ion coexisted in the fractions of a positive peak of phosphate ion, in all sample eluent systems employed. This phenomenon was assumed to occur by the coupled reaction of change-partner and ion-exchange reactions including ion exclusion. PMID- 10720229 TI - Analytical potential of fullerene as adsorbent for organic and organometallic compounds from aqueous solutions. AB - In this work, the analytical potential of C60 fullerene as a sorbent for organic and organometallic compounds from aqueous solutions was studied for the first time. Fullerene adsorbs many types of organic substances (e.g., N methylcarbamates, phenols, polycyclic aromatic hydrocarbons, amines) with efficiencies that depend on the nature of the compound concerned and never exceed 60%. Conventional sorbents such as XAD-2 or polyurethane foam are more efficient than C60 for this purpose. Organometallic compounds (viz. metalocenes and organoleads) are quantitatively adsorbed on C60 via the formation of neutral complexes or chelates; the adsorption constant is dramatically increased by the use of classical reagents such as pyrrolidinedithiocarbamate or diethyldithiocarbamate. A complementary comparative study on the adsorption of organometallic complexes on RP-C18 and silica gel 100, among others, showed C60 to be superior as sorbent. All experiments in this work were carried out by using continuous flow configurations and gas chromatography-atomic absorption spectrometry techniques. PMID- 10720230 TI - Novel cinchona alkaloid carbamate C9-dimers as chiral anion-exchange type selectors for high-performance liquid chromatography. AB - Nine new quinine (QN) carbamate C9-dimers (QN-X-QN), with different aliphatic and cyclic spacers (X), have been synthesized and immobilized onto porous silica gel for HPLC. The chiral discriminating behavior of these "dimeric" anion-exchange type chiral stationary phases (CSPs) has been investigated in detail, to elucidate the role of the presence of a second QN subunit on the chiral selector (SO), as well as the influence of the structure and length of the spacer, on the overall chiral recognition of a set of N-derivatized amino acids and other acidic drugs. The bulkiness of the intermediate spacer tuned the chiral recognition abilities of these SOs, with the 1,3-adamantylen-derived CSP being the one that led to the best separations. Shorter spacers reduced the chiral discrimination abilities of the "dimeric" selectors, with the n-hexylen bridge being the most favorable distance to allow a nearly independent interaction of the two QN subunits with the racemic analytes. The comparison to five "monomeric" CSPs showed that the "dimeric" ones usually retain the chiral analytes more strongly, though the enantioseparation is not improved. Nevertheless, the exceptional resolution abilities of dimeric SOs with a trans- 1,2-diaminocyclohexylen-bridge for the separation of DNP-derivatives of amino acids and certain acidic drugs of therapeutical interest (e.g., profens) seemed to be superior to most of the other CSPs. PMID- 10720231 TI - Stability of high-performance liquid chromatography columns packed with poly(methyloctylsiloxane) sorbed and radiation-immobilized onto porous silica and zirconized silica. AB - Reversed-phase packing materials were prepared from HPLC silica and from zirconized HPLC silica support particles having sorbed poly(methyloctylsiloxane) (PMOS) as the stationary phase. Portions of zirconized material were subjected to 80 kGy of ionizing radiation. Columns prepared from these packing materials were subjected to 5000 column volumes each of neutral and alkaline (pH 10) mobile phases, with periodic tests to evaluate chromatographic performance. It was shown that the PMOS stationary phase sorbed onto zirconized silica requires an immobilization treatment (such as gamma irradiation) for long term stability while prior surface zirconization of the silica support surface greatly improves the chromatographic stability of the stationary phase when using alkaline mobile phases. PMID- 10720232 TI - Effects in high-performance liquid chromatography of a high pH in the mobile phase on poly(methyloctylsiloxane) immobilized by gamma-radiation on titanium grafted silica. AB - Effects of high-pH environments on a stationary phase prepared by gamma-radiation immobilization of poly(methyloctylsiloxane) on titanium-grafted silica were investigated by HPLC testing with standard sample mixtures. The HPLC parameters indicate good stationary phase stability to 10000 column volumes each of mobile phases with pH of 7, 9 and 12. At pH 13, the efficiency decreases slowly, although reasonably good separations are still possible until increasing flow resistance no longer allows easy passage of the mobile phase. PMID- 10720233 TI - Development of a new cyano-bonded column for high-performance liquid chromatography. AB - A unique cyano-bonded column for high-performance liquid chromatography was developed by chemically bonding cyanopropyl groups to the silica gel support, and its chromatographic performance was described. Eight cyanopropyl-bonded silica gels were prepared under different conditions. These packing materials were packed into a stainless steel column, the chromatographic properties and durability of which were investigated in both normal- and reversed-phase partition modes. The separating selectivity and the durability of cyanopropyl bonded silica (CN) columns were dependent on the preparation conditions. Particularly, the relationship between the density of cyanopropyl groups on the surface of the silica gel and the irreversible adsorption of basic compounds was examined. Also, the experimental results indicated that endcapping resulted in the poor separating selectivity and durability of the CN columns. PMID- 10720234 TI - Characterization of an avidin-bonded column for direct injection in reversed phase high-performance liquid chromatography. AB - A denatured avidin-bonded column was suitable for use in the reversed-phase HPLC and complete recovery of serum protein over a wide range of pH. Retention property of the denatured avidin-bonded column was very nearly to a non-denatured avidin-bonded column already reported and, however, showed very high stability to organic solvent. A performance of the denatured avidin-bonded column was maintained even if 500 continuous injections of human serum (total 10 ml). PMID- 10720235 TI - Optimized stationary phases for the high-performance liquid chromatography electrospray ionization mass spectrometric analysis of basic pharmaceuticals. AB - Stationary phases were investigated for HPLC coupled with electrospray ionization mass spectrometry (ESI-MS) for the analysis of basic drugs. Tricyclic antidepressants (TCAs) and beta-blockers were used as model solutes. The functional groups, pentafluorophenyl (PFP), OH, CN or CH3 were attached to the silica via a propyl chain. The effects of these stationary phases as well as C8 and C18 phases on retention and peak shape of the basic drugs were studied. The CN and PFP phases adequately retained (tR of 2 to 6 min) the basic drugs when the mobile phase was composed of 90% acetonitrile, whereas with the C4, C8 and C18 phases, less than 40% acetonitrile had to be used to provide adequate retention of the basic drugs. Because acetonitrile provides better desolvation in ESI than an aqueous solvent, it produces an increased MS signal. As an example of the HPLC ESI-MS analysis of the beta-blocker, pindolol, on a CN phase, the use of 90% acetonitrile in the mobile phase increased the ESI-MS signal by 790% when compared to a C18 phase which could use only 5% acetonitrile in the mobile phase for retention of the solute. In addition, the CN and PFP phases provided better peak shape than the OH phase and the hydrophobic phases (C4, C8 and C18) and ion pairing or ion-suppressing agents were not required. The retention behavior of the TCAs and beta-blockers on each of the phases is described. PMID- 10720236 TI - Fast and quantitative high-performance liquid chromatography method for the determination of 9-fluorenylmethoxycarbonyl release from solid-phase synthesis resins. AB - Base catalyzed cleavage of 9-fluorenylmethoxycarbonyl (FMOC) group and subsequent analysis by UV spectrophotometry is a commonly used technique for measuring the loading of functional groups on solid supports. Recent works suggest that using 1,8-diazabicyclo[5,4,0]undec-7-ene (DBU) instead of piperidine makes it possible to use gas chromatography for quantitation, but due to the long deprotection time used, the method is not high-throughput. We observed that the dibenzofulvene released after DBU deprotection could be measured by reversed-phase (RP) HPLC. We have optimized the concentration of DBU as well as the time of the deprotection and coupled with a fast RP-HPLC separation results in a highly reproducible, high throughput method. The measured loading correspond well with the manufacturer's data on several commercial resins. Using this method we have quantitated the amine loading on several polystyrene resins and we have found that the total amount of functional group can be more than twice the amount of the available ones. We concluded that the differences were the function of the resin loading as well as the level of crosslinking. PMID- 10720237 TI - Repeatability and reproducibility of retention data and band profiles on reversed phase liquid chromatography columns. IV. Results obtained with Luna C18 (2) columns. AB - The reproducibility of retention data and band profile characteristics was investigated for a series of columns packed with Luna C18 (2), a silica-based, reversed-phase adsorbent. High precision data were obtained and statistically compared among five columns from the same batch (column-to-column reproducibility) and nine columns from as many different batches (batch-to-batch reproducibility). These data were acquired under five different sets of chromatographic conditions, for a group of 30 neutral, acidic, and basic compounds selected as probes following an experimental protocol previously described. This work is part of a study on the precision of chromatographic analyses. Its purpose is to illustrate the contribution of the stationary phase reproducibility to this precision. PMID- 10720238 TI - Control of column temperature in reversed-phase liquid chromatography. AB - When separations by reversed-phase liquid chromatography (RP-LC) are carried out at temperatures other than ambient, resulting retention times and bandwidths can depend on the equipment used. As a result, an RP-LC separation that is adequate when carried out on one LC system may prove inadequate when the separation is repeated on a second system. In the present study, various temperature-related problems which can result in a failure of method transfer for non-ambient RP-LC methods were examined. Means for correcting for such effects and thereby ensuring method transferability are described. PMID- 10720239 TI - Performance of experimental sample injectors for high-performance liquid chromatography microcolumns. AB - An experimental injector for HPLC microcolumns and a 3-nl conductivity detector connected directly to the injector outlet with a 19-nl tube were used to study injector dispersion, guide the design of improved injectors, and suggest appropriate injection techniques. With regard to the small injection volumes required when no on-column concentration technique is used, we show that in some circumstances: (i) there are two volumes to be considered, the sample volume (that which is intended to be injected) and the effective injection volume (that which contains all the sample after it has completely emerged from the injector). Due to dispersion, the latter is often many times the former. An injector performance factor is defined as the ratio of the two volumes. (ii) A smaller sample chamber volume in an injector does not necessarily produce a proportionately smaller effective injection volume, in which case there is a reduction of peak height that degrades sensitivity without a commensurate reduction in peak width that would improve resolution. (iii) Adjusting the geometry of the sample chamber and stator passage can significantly improve injector performance, as illustrated for sample volumes from 2 nl to 1 microl. (iv) In some cases, reducing the diameter of an injector passageway in an attempt to reduce dispersion actually causes performance to worsen. PMID- 10720240 TI - 2-(4-Carboxyphenyl)-6-N,N-diethylaminobenzofuran: a useful reagent for the sensitive determination of alcohols by high-performance liquid chromatography with fluorimetric detection. AB - A simple and highly sensitive method for the determination of short, medium and long-chain alcohols using high-performance liquid chromatography with fluorimetric detection is described. The alcohols were derivatized to their corresponding esters with (4-carboxyphenyl)-6-N,N-diethylaminobenzofuran. The esterification reaction proceeded rapidly and smoothly in acetonitrile at 60 degrees C with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (a coupling agent) in the presence of 4-dimethylaminopyridine (a base catalyst). The resulting esters of alcohols from methanol to eicosanol (C1-C20-ol) were separated on a reversed-phase column (Ultrasphere C8) with gradient elution (acetonitrile-water) and detected fluorometrically (excitation 387, emission 537 nm). The lower limits of detection (signal-to-noise ratio of 3) for the derivatized alcohols were in the range of 0.2-0.5 pg. PMID- 10720241 TI - 2,4-Dinitro-3,5,6-trideuterophenylhydrazones for the quantitation of aldehydes and ketones in air samples by liquid chromatography-mass spectrometry. AB - The quantification of carbonyl compounds in air samples using an internal calibration approach with stable isotope-labelled standards and HPLC-atmospheric pressure chemical ionization MS analysis is presented. 2,4-Dinitro-3,5,6 trideuterophenylhydrazine and various of its hydrazones have been synthesized and characterized for the first time. The respective stable isotope-labelled hydrazones of a series of aldehydes and ketones are applied as internal standards for the determination of the carbonyls in car exhaust samples. Various aldehydes are identified and quantified by MS detection. The results exhibit good agreement to quantification data obtained with UV detection. PMID- 10720242 TI - Micropreparative ligand fishing with a cuvette-based optical mirror resonance biosensor. AB - We have previously demonstrated the role of an optical biosensor (BIAcore 2000) as a specific detector to monitor chromatographic fractions during the purification and characterisation of ligands for orphan biomolecules. We have now extended this application to perform micropreparative ligand fishing directly on the sensor surface using an automated cuvette-based optical biosensor (Iasys Auto+) equipped with a high-capacity carboxymethyldextran surface (surface area 16 mm2). Using a F(ab)2' fragment of the A33 monoclonal antibody as bait, we have recovered microgram quantities of essentially homogeneous A33 ligand from the sensor surface in a form suitable for subsequent sensitive and specific down stream analysis (micropreparative HPLC, sodium dodecyl sulphate-polyacrylamide gel electrophoresis and Western blotting). The design of the cuvette-based system facilitates recovery of desorbed material from the constrained workspace in small volumes at high concentration. The use of on-surface detection allows the surface viability to be continuously monitored and permits direct quantitation of both bound and recovered material. PMID- 10720243 TI - Increasing the reliability of the identification by high-performance liquid chromatography by means of selective and/or sensitive detection. AB - An approach for quantitative assessment of the reliability of identification at high-performance liquid chromatography is proposed. The quantitative assessment of identification is useful for determination of selectivity at validation of the analytical methods. Chromatograms and spectra of the analytes are presented as maps in which characteristics as retention times, detector's signals, maxima and minima and another characteristics of spectra are used for identification. A formula for quantitative determination of the contribution of these characteristics on the reliability of identification is given. Using the more selective diode array detector than the convenient UV detector increases the reliability by several orders. A similar result was obtained when the UV detector was replaced with the more sensitive and selective fluorescence detector. Despite of the small contribution of the separation to the reliability its influence is very important for distinguishing of isomers because their spectra are identical. PMID- 10720244 TI - Microwave-assisted extraction by fast sample preparation for the systematic analysis of additives in polyolefins by high-performance liquid chromatography. AB - This paper presents a new approach for complete and systematic analysis of organic additives in polyolefins. The proposed procedure is a convenient combination of sample preparation, performed by microwave-assisted extraction (MAE), and direct chromatographic evaluation of extract by high-performance liquid chromatography coupled with ultraviolet and evaporative light scattering detection. In particular two microwave-assisted processes are reported and discussed: the one-step MAE, useful for additives with low-medium dipolarity (like stabilizers, flame retardant, antistatics, slip and processing agents), and the two-step MAE, useful for additives with either high dipolarity (like organic salts, antigasfading, antiacid, nucleating agent) or high molecular mass (like polymeric hindered amine light stabilizers). Both the proposed processes have been tested on representative additives in five commercially common polymeric matrices, demonstrating their satisfactory analytical results, in terms of repeatability and percentage recoveries, and their good performances, in terms of safety and time/solvent consumption, in comparison with those of traditional extraction methods. PMID- 10720245 TI - Comparison of commercial solid-phase extraction sorbents for the sample preparation of potato glycoalkaloids. AB - In this study five different commercial sorbents C18, SCX, CN, Certify and Oasis HLB were compared for the solid-phase extraction of potato glycoalkaloids. The recoveries were determined using alpha-solanine, alpha-chaconine and alpha tomatine, which contained dehydrotomatine as an impurity, as standard compounds. The samples were analysed by reversed-phase liquid chromatography under gradient elution conditions using a Zorbax Rx-C18 column and acetonitrile-25 mM triethylammonium phosphate buffer (pH 3.0) as the mobile phase. The highest recovery (approximately 100%) was achieved with Oasis HLB (60 mg) cartridges. An acetic acid extract of wild Solanum brevidens leaf material was used for the testing of a clean-up procedure. The SCX proved to be the most selective and efficient for removing the undesired components from the leaf extract. PMID- 10720246 TI - Determination of heterocyclic aromatic amines in meat extracts by liquid chromatography-ion-trap atmospheric pressure chemical ionization mass spectrometry. AB - When protein-rich foods are processed under normal cooking conditions, heterocyclic aromatic amines (HAAs) can be generated at a few parts per billion level. In this work, we have analyzed the HAAs present in a lyophilized meat extract by means of a simplified solid-phase extraction procedure. All the analytes were collected in a single extract with recoveries in the range of 45.6 75.2%, so the analysis time has been greatly reduced. Problems derived from the less exhaustive purification of the extract have been solved by using MS(ion trap) detection. The RSD for quantification ranged from 2.1% to 5.1% for run-to run precision and from 5.2% to 11% for day-to-day precision. The limits of detection for standard solutions ranged from 20 to 150 pg injected. For the meat extract analyzed limits of detection from 0.9 to 11.2 ng g(-1) were obtained. Results of the quantification are in agreement with those obtained using different clean-up procedures. PMID- 10720247 TI - Analysis of channel-geometry effects on separation efficiency in rectangular capillary electrochromatography columns. AB - A three-dimensional random walk model was developed to evaluate the impact of column geometry on separation efficiency in chromatography systems driven by electroosmotic flow. Contributions of injection plug length, cross-sectional area of channels, and aspect ratio of rectangular channels were examined in these simulation studies. Sample plug length had no impact on efficiency until it exceeded roughly 0.4% of the channel length. Plate height increased rapidly with increasing k' as expected, almost doubling in going from k'=0.25 to 0.35. Channel geometry also had a major effect on efficiency. Plate height increased sharply in rectangular channel columns until the channel aspect ratio reached 4-8. But the effect of channel depth was even more dramatic. Minimum plate height (Hmin) was roughly half that of the channel depth in ideal cases. Hmin in a 10x2 microm channel was at 1.6 mm s(-1). Rectangular channels comparable to those obtained by microfabrication are equivalent to packed column capillary electrochromatography columns in all cases. PMID- 10720248 TI - Influence of mobile phase composition on electroosmotic flow velocity, solute retention and column efficiency in open-tubular reversed-phase capillary electrochromatography. AB - The effects of some experimental parameters, such as the volume fraction and type of organic modifier in the mobile phase, and the concentration, type and pH of the buffer on the electroosmotic flow velocity, the retention behavior of test solutes, and the column efficiency have been investigated in capillary electrochromatography (CEC) using an open-tubular column of 9.60 microm I.D. with a porous silica layer chemically modified with C18 as stationary phase. The retention of a group of polycyclic aromatic hydrocarbons (PAHs) used as a test mixture varied significantly by changing the organic modifier content in the hydroorganic mobile phase according to the reversed-phase-like selectivity of the stationary phase. In addition, an increase in the percentage of organic modifier resulted in a slight increase in the linear velocity of the EOF. On the other hand, when the phosphate buffer concentration was increased over the range 1-50 mM, the electroosmotic mobility fell dramatically, the retention of the solutes decreased steadily, and the plate height showed a significant increase. The results obtained with phosphate, trishydroxymethylaminomethane or 2 morpholinoethanesulfonic acid as buffers were similar when pH remained constant. Optimization in CEC was essential to achieve further enhancement of separation performance, because the analysis time and separation resolution are essentially affected when varying operating parameters. Separations of seven PAHs with more than 100000 plates are presented within 4 min analysis time. PMID- 10720249 TI - Influence of flow patterns on chromatographic efficiency in centrifugal partition chromatography. AB - Visualization of flow patterns in centrifugal partition chromatography (CPC) was performed with an asynchronous camera and a stroboscope triggered by the CPC rotor, allowing a channel to be selected and observed regardless of rotational speed. Three main types of flow states were noted as a function of rotational speed and flow-rate: jets stuck along channel walls, broken jets and atomization. Our observations emphasize the importance of Coriolis force on flow shape. Chromatographic efficiency was related to the dispersion of the mobile phase in the stationary phase. PMID- 10720250 TI - Improvement purification of sulfated oligofucan by ion-exchange displacement centrifugal partition chromatography. AB - Centrifugal partition chromatography in ion-exchange displacement mode was used to fractionate mixtures of sulfated oligofucans obtained by partial depolymerization of brown seaweed fucoidans. Diluted (10%, v/v) protonated LA2 (a lipophilic secondary amine) is used as a weak exchanger. In an attempt to improve this method, several solvents (methyl isobutyl ketone, methyl tert.-butyl ether, BuOH) were tested to dissolve LA2H+. MtBE produced less bleeding than MiBK, whereas BuOH proved unsuitable. The sample injected needs to be highly diluted in water to ensure participation in the chromatographic process. A comparison of data (NMR, composition, molecular mass) indicated the homogeneity of the fractions obtained as well as the differences between them. PMID- 10720251 TI - Some considerations concerning the composition of the mobile phase in capillary electrochromatography. AB - In capillary electrochromatography (CEC) the propulsion of the mobile phase is effected by electroosmosis. The velocity of the electroosmotic flow is dependent on surface properties of the stationary phase and on bulk properties of the mobile phase. Therefore, in CEC the optimization of the mobile phase composition must take more factors into account than in pressure-driven LC. In this paper, the impact of the electrolyte concentration in the mobile phase and of the volume fraction of the organic mobile phase constituent on the velocity of the electroosmotic flow and on the chromatographic efficiency is investigated for CEC with capillaries packed with octadecylsilica gel. Bias of the data by an open section of the capillary has been excluded by employing completely packed capillaries and detection in a packed section. Acetonitrile as organic constituent of the mobile phase is compared to other possible organic modifiers (polar organic solvents) concerning influence on velocity of the electroosmotic flow and retention of solutes. PMID- 10720252 TI - Migration time correction for the analysis of derivatized amino acids and oligosaccharides by micellar capillary electrochromatography. AB - Migration-time reproducibility is essential in the use of capillary electrophoresis to identify components in mixtures. Two methods based on the migration time of either one or two reference markers are proposed for improving migration time reproducibility. These methods were evaluated to determine the migration time reproducibility for phenylthiohydantoin-amino acids, fluorescein thiohydantoin-amino acids, and tetramethylrhodamine labeled oligosaccharides. In the best case, the relative standard deviation of the migration time was reduced from >3% without correction to <0.04% with the two-marker correction. PMID- 10720253 TI - Separation of peptides by strong cation-exchange capillary electrochromatography. AB - Separation of small peptides on ion-exchange capillary electrochromatography (IE CEC) with strong cation-exchange packing (SCX) as stationary phase was investigated. It was observed that the number of theoretical plates for small peptides varied from 240000 to 460000/m, and the relative standard deviation for t0 and the migration time of peptides were less than 0.57% and 0.27%, respectively for ten consecutive runs. Unusually high column efficiency has been explained by the capillary electrophoretic stacking and chromatofocusing phenomena during the injection and separation of positively charged peptides. The sample buffer concentration had a marked effect on the column efficiency and peak area of the retained peptides. The influences of the buffer concentration and pH value as well as the applied voltage on the separation were investigated. It has been shown that the electrostatic interaction between the positively charged peptides and the SCX stationary phase played a very important role in IE-CEC, which provided the different separation selectivity from those in the capillary electrophoresis and reversed-phase liquid chromatography. A fast separation of ten peptides in less than 3.5 min on IE-CEC by adoption of the highly applied voltage was demonstrated. PMID- 10720254 TI - Immobilisation and evaluation of a vancomycin chiral stationary phase for capillary electrochromatography. AB - The macrocyclic antibiotic, vancomycin, is covalently bonded to LiChrospher diol silica packed columns and evaluated in capillary electrochromatography (CEC) both in the reversed-phase and polar organic mode. Initially, capillaries were packed with 5 microm LiChrospher 100 A diol silica and evaluated in CEC with a reversed phase biphenyl-pyrene achiral test resulting in resolution and efficiency values of ca. 2.5 and 100000 plates meter(-1), respectively. Repeatability for this test (resolution and efficiency) was also examined and found to be acceptable for both run-to-run (n=5, 0.74% and 1.5%) and column-to-column (n=5, 3.4% and 9.0%), respectively. Similar results were obtained when the 10 microm LiChrospher 1000 A diol silica was examined with the exception of efficiency, where a reduced plate height value of four times lower was obtained compared to the 100 A material. A simple three step in-situ vancomycin immobilisation procedure was subsequently carried out on these packed diol columns. Selectivity was obtained for thalidomide enantiomers on this vancomycin chiral stationary phase in reversed phase CEC with resolution and efficiency values of ca. 2.5 and 80000 plates meter(-1), with acceptable repeatability (n=8) 0.9% and 3.0%, respectively. Selectivity was also obtained for thalidomide enantiomers on this phase in the polar organic mode with resolution and efficiency values of ca. 2.5 and 120000 plates meter(-1), with acceptable repeatability (n=7) 0.9% and 2.0%, respectively. It was possible to deduce from a chemometric design carried out for evaluating the mobile phase component effects that organic modifier ratio, MeOH/MeCN, played a significant role in controlling both resolution and efficiency. It was also possible to separate a number of basic analytes including four beta-adrenergic blocking agents in the polar organic mode albeit with lower resolution and efficiency values, ca. 1.5 and 45000 plates meter(-1), respectively. PMID- 10720255 TI - Application of spectral libraries for high-performance liquid chromatography atmospheric pressure ionisation mass spectrometry to the analysis of pesticide and explosive residues in environmental samples. AB - The coupling of high-performance liquid chromatography (HPLC) and atmospheric pressure ionisation mass spectrometry (API-MS) seems to be the method of choice if good separation and selective detection of semi-volatile, thermolabile, and polar substances is required. Libraries of mass spectra will make the identification of unknown substances in complex environmental samples easier and more user-friendly. Unfortunately, existing GC-MS libraries are not applicable to HPLC-API-MS analysis. Thus, new and extensive mass spectral libraries were constructed. Several investigations of chromatographic (composition and salt concentration of the eluent) as well as mass spectrometric (orifice voltage) parameters and a few applications of real environmental samples are used to discuss the possibilities and limits of these libraries. PMID- 10720256 TI - Analysis of pesticide residues in matrices with high lipid contents by membrane separation coupled on-line to a high-performance liquid chromatography system. AB - Separation through membranes coupled to an HPLC system was used as a technique for the analysis of pesticide multiresidues in samples with high lipid contents. As well as the usual procedure, in the proposed system it is possible to recirculate the sample through the membrane cell, which permits the extraction system to be applied to cases in which only a very small volume of sample is available. A procedure for pesticide multiresidue analysis in egg samples was developed as a representative example of the applicability of the proposed method. To accomplish this, the analytes (dichlorvos, dimethoate, propoxur, paraoxon, pirimicarb, atrazine, ametryne, terbutryne, azinphos-methyl, folpet) were subjected to prior extraction in a Soxhlet system, after which the extract was introduced into the membrane separation device coupled to the HPLC system. This procedure afforded clean chromatograms, hence considerably facilitating determination, and at the same time was efficient in removing macromolecular compounds. For egg samples, spiked at a concentration level of 0.750 mg/kg, recoveries ranged from 60 to 98%. The detection limits varied from 0.018 mg/kg for dichlorvos to 0.002 mg/kg for atrazine. PMID- 10720257 TI - On-line ion-pair solid-phase extraction-liquid chromatography-mass spectrometry for the analysis of quaternary ammonium herbicides. AB - An ion-pair on-line solid-phase extraction procedure using C8 extraction disks, suitable for liquid chromatography-mass spectrometry analysis is developed to determine quaternary ammonium herbicides (quats) in water samples. The separation of these compounds was performed using ion-pair chromatography with heptafluorobutyric acid (15 mM, pH 3.3) and acetonitrile gradient elution. Detection was carried out using a quadrupole mass spectrometer. Water sample volumes up to 50 ml can be preconcentrated with recoveries higher than 70%. Good precision and accuracy (day-to-day and run-to-run) were obtained and the detection limits ranged from 6 to 85 ng l(-1). The proposed on-line ion-pair solid-phase method enables compliance with European Community directives for drinking waters (100 ng l(-1)). PMID- 10720258 TI - Determination of triazine herbicides in natural waters by solid-phase extraction and non-aqueous capillary zone electrophoresis. AB - Capillary zone electrophoresis (CZE) in an organic medium was used to analyse triazines at sub-ppb concentration levels in natural waters after a preconcentration step using conventional C18 cartridges and new Oasis HLB devices. With both sorbents, satisfactory results were obtained on analysing deionized water. However, on analysing natural waters, both sorbents showed very different types of behaviour. The different variables affecting the elution of both sorbents were studied, resulting in the choice of Oasis HLB as the most suitable for later separation by CZE in non-aqueous medium. Combination of a preconcentration step with electrokinetic injection revealed that capillary electrophoresis with simple UV detection can also be used satisfactorily for the quantification of micropollutants in natural waters. The detection limits obtained varied between 0.01 and 0.05 microg l(-1), depending on the type of matrix analysed. The day-to-day precision varied between 0.9% and 2.3%, expressed as the relative standard deviation. PMID- 10720259 TI - Use of solid-phase extraction and high-performance liquid chromatography for the determination of triazine residues in water: validation of the method. AB - A method for determination of some triazine residues in water has been developed. The method involves concentration with C18 solid-phase extraction cartridges followed by high-performance liquid chromatographic analysis using a C18 column with UV detection at 230 nm, a mobile phase of methanol-water (60:40, v/v) at pH 4.6 (phosphoric acid) and a flow-rate of 0.8 ml/min. After optimization of the extraction and separation conditions, the method was validated. The method can be used for determination of atrazine, simazine, cyanazine and ametryn in water, within the international limits of 0.1 microg/l. PMID- 10720260 TI - Evaluation of surface- and ground-water pollution due to herbicides in agricultural areas of Zamora and Salamanca (Spain). AB - The pollution of agricultural land due to herbicides was assessed in the Guarena and Almar river basins, situated in the provinces of Zamora and Salamanca (Spain). A set of fifteen herbicides, including triazines, ureas, amides and others, was selected owing to their frequency of use, the amounts used, their toxicity and their persistence in the environment. Solid-phase extraction with polymeric cartridges, followed by HPLC with diode-array detection, were used to monitor the herbicides. This technique was chosen owing to the wide range of functionality and polarity of the analytes under study. The detection limits obtained were in the 0.004-0.025 microg/l range (lambda=220 nm). Surface and ground waters, taken from different locations in the basins, were analyzed over a 6-month period. The presence of six out of the fifteen herbicides monitored- chlortoluron, atrazine, terbutryn, alachlor, diflufenican and fluazifop-butyl- was detected in several samples at levels ranging from the detection limit to 1.2 microg/l. The relationship of these herbicides to the agricultural activities of the zone is discussed. PMID- 10720261 TI - Study of the behaviour of azinphos-methyl in a clay mineral by batch and column leaching. AB - Research efforts dealing with the processes affecting the transport of pesticides in soils are needed in order to prevent further damage of surface and groundwater reserves. Although organic matter has been recognised as the most important contributor to the adsorption of non-ionic organic pesticides in soils, in some cases clay minerals may have an important role in the retention of these compounds. The present study was designed to improve the knowledge of the behaviour of azinphos-methyl in soils. Coefficients from adsorption isotherms and HPLC analysis of soil column leachates were used in this work for predicting pesticide mobility in soils. The studied clay mineral was a Spanish bentonite with a predominant montmorillonite fraction. The results showed that azinphos methyl was adsorbed on the clay mineral and demonstrated the catalytic effect of bentonite on the hydrolysis of the pesticide. PMID- 10720262 TI - Determination of linuron and related compounds in soil by microwave-assisted solvent extraction and reversed-phase liquid chromatography with UV detection. AB - The combination of microwave-assisted solvent extraction (MASE) and reversed phase liquid chromatography (RPLC) with UV detection has been investigated for the efficient determination of phenylurea herbicides in soils involving the single-residue method (SRM) approach (linuron) and the multi-residue method (MRM) approach (monuron, monolinuron, isoproturon, metobromuron, diuron and linuron). Critical parameters of MASE, viz, extraction temperature, water content and extraction solvent were varied in order to optimise recoveries of the analytes while simultaneously minimising co-extraction of soil interferences. The optimised extraction procedure was applied to different types of soil with an organic carbon content of 0.4-16.7%. Besides freshly spiked soil samples, method validation included the analysis of samples with aged residues. A comparative study between the applicability of RPLC-UV without and with the use of column switching for the processing of uncleaned extracts, was carried out. For some of the tested analyte/matrix combinations the one-column approach (LC mode) is feasible. In comparison to LC, coupled-column LC (LC-LC mode) provides high selectivity in single-residue analysis (linuron) and, although less pronounced in multi-residue analysis (all six phenylurea herbicides), the clean-up performance of LC-LC improves both time of analysis and sample throughput. In the MRM approach the developed procedure involving MASE and LC-LC-UV provided acceptable recoveries (range, 80-120%) and RSDs (<12%) at levels of 10 microg/kg (n=9) and 50 microg/kg (n=7), respectively, for most analyte/matrix combinations. Recoveries from aged residue samples spiked at a level of 100 microg/kg (n=7) ranged, depending of the analyte/soil type combination, from 41-113% with RSDs ranging from 1-35%. In the SRM approach the developed LC-LC procedure was applied for the determination of linuron in 28 sandy soil samples collected in a field study. Linuron could be determined in soil with a limit of quantitation of 10 microg/kg. PMID- 10720263 TI - Determination of imidacloprid and its metabolite 6-chloronicotinic acid in greenhouse air by high-performance liquid chromatography with diode-array detection. AB - A method is described for analysing and sampling imidacloprid and its metabolite 6-chloronicotinic acid in greenhouse air by high-performance liquid chromatography (HPLC) with diode-array detection (DAD). The trapping efficiency of two solid sorbents, Amberlite XAD-2 and Amberlite XAD-4 and the use of different desorption procedures have been tested. To validate the methodology, standard atmospheres containing known concentrations of these pesticides and with different relative humidities were generated. No breakthrough was observed in the range of concentrations studied. Dissipation of analytes was investigated in a 24 h period after application by using personal samplers in a field experiment. PMID- 10720264 TI - Optimization of the extraction of polycyclic aromatic hydrocarbons from wood samples by the use of microwave energy. AB - A simple and rapid microwave-assisted extraction (MAE) procedure was developed and optimized for benzo[a]anthracene, benzo[e]pyrene, benzo[b]fluoranthene, benzo[k]fluoranthene and benzo[a]pyrene in wood samples. The spiked wood used was prepared 3 months before analysis to simulate weathering processes and to allow the formation of analyte-matrix interaction. The samples, immersed in acetonitrile were irradiated with microwaves in a closed-vessel system. Optimization of the method was achieved by using a factorial design approach on parameters such as extraction time, temperature and sample amount. The analysis of extracts has been carried out by reversed-phase high-performance liquid chromatography with fluorescence detection for quantification and UV-diode-array detection for confirmation. The MAE procedure yielded extracts that could be analyzed directly without any preliminary clean-up or solvent exchange steps. PMID- 10720265 TI - Determination of polycyclic aromatic hydrocarbons in marine sediments by high performance liquid chromatography after microwave-assisted extraction with micellar media. AB - A simple and rapid method is developed for extraction and determination of polycyclic aromatic hydrocarbons (PAHs) in marine sediments. The procedure was based on the microwave-assisted extraction of PAHs in marine sediment samples using a micellar medium of Polyoxyethylene 10 lauryl ether as extractant. Two level factorial designs have been used to optimize the microwave extraction process. The analysis of extracts has been carried out by HPLC with UV detection. Fortified sediments gave an average recovery between 85.70 and 100.73%, with a relative standard deviation of 1.77-7.0% for PAHs with a ring number higher than three. PMID- 10720266 TI - Microporous membrane liquid-liquid extraction coupled on-line with normal-phase liquid chromatography for the determination of cationic surfactants in river and waste water. AB - Membrane-based continuous liquid-liquid extraction combined on-line with normal phase liquid chromatography is proposed for the determination of cationic surfactants in complex aqueous samples. The technique has the potential for complete automation. Selective enrichment of cationic surfactants from spiked river water and waste-water samples with simultaneous removal of matrix constituents, followed by a quantitative transfer of the extract onto a liquid chromatographic column and separation of the surfactant homologues yielding low detection limits, has been realised. The homologues of alkyldimethylbenzylammonium chloride (Dodigen 226) were chosen as model compounds in the method development. Dodigen homologues were ion-paired with heptanoic acid and extracted into chlorobutane by means of microporous membrane liquid-liquid extraction. It was thereby possible to attain an enrichment of over 250 times for one of the homologues, viz. the concentration in the organic liquid is 250 times higher than in the original sample. Detection limits for the three best-detected homologues of the mixture were in the range 0.7-5 microg/l in spiked river water samples. Ion-pair normal-phase liquid chromatography, again with heptanoic acid as counter-ion, gave the necessary separation of the surfactant homologues. PMID- 10720267 TI - Eye movements in parkinsonism: it's saccadic speed that counts. PMID- 10720268 TI - Biologically stressed muscle fibers in sporadic IBM: a clue for the enigmatic etiology? PMID- 10720269 TI - Choice of endpoints in antiplatelet trials: which outcomes are most relevant to stroke patients? AB - The relative efficacies of different antiplatelet agents for stroke prevention are unclear because of differences in clinical trial design, a lack of direct comparisons between individual agents, and differences in the choice of primary endpoints. Individual endpoints in a clinical trial are often combined into a single primary endpoint cluster. Theoretically, a combined endpoint may reduce the sample size required to demonstrate significant benefits of an effective therapy. However, unless all components of a composite endpoint are affected in the same direction and to a similar degree, their inclusion may not provide the anticipated increase in statistical power. In fact, the use of a combined endpoint may lead to an underestimate of therapeutic benefits when patients at high risk for one particular endpoint are studied. For patients with stroke or TIA, the single outcome of stroke is particularly important because these patients have a higher risk of recurrent stroke than any other vascular outcome during the initial years after a stroke. Because of the low incidence of myocardial infarction (MI) in stroke trials, the inclusion of MI in the primary endpoint will usually have minimal influence on trial outcome, and antiplatelet therapy has not been shown to be beneficial in preventing nonvascular death. Chance variations in the incidence of MI or death may detract from the benefit of the agent for stroke prevention when it is included in a combined endpoint. The benefit of antiplatelet therapies for patients with recent cerebrovascular events is determined most accurately if stroke alone is chosen as the primary endpoint. PMID- 10720270 TI - Longitudinal ocular motor study in corticobasal degeneration and progressive supranuclear palsy. AB - OBJECTIVE: To evaluate the usefulness of ocular motor information in the early diagnosis of corticobasal degeneration (CBD) and progressive supranuclear palsy (PSP). METHODS: Seven PSP patients, six CBD patients, and three atypical CBD patients were followed longitudinally with repeated electrooculographic (EOG) recordings, at 6-month intervals, to search for features that could confirm or modify the diagnosis. Visually guided saccades and antisaccades were studied. Data from clinical evaluations were independently collected. RESULTS: PSP patients had decreased saccade velocity throughout the disease course. Patients with probable CBD showed preserved saccade velocity but important increased saccade latency ipsilateral to the apraxia side. Similar to patients with PSP, those with atypical CBD features exhibited clinically evident abnormalities of vertical saccades and early slowing of horizontal saccade velocity, but no increase in saccade latency or early square-wave jerks. When clinical "telltale signs" appeared and the clinical diagnosis was reviewed independent of EOG recording, the three patients with atypical CBD features were diagnosed as having PSP although new or overlapping syndromes cannot be excluded. CONCLUSIONS: Consecutive EOG recordings help diagnose atypical CBD and PSP disorders earlier. PMID- 10720271 TI - AlphaB-crystallin immunolocalization yields new insights into inclusion body myositis. AB - OBJECTIVE: To study the expression of the small heat shock protein, alphaB crystallin (alphaBC), in inclusion body myositis (IBM). BACKGROUND: In humans, alphaBC is constitutively expressed in the eye lens, muscle, and heart, but not in lymphoid tissues. Induced expression of alphaBC occurs under metabolic stress, in virus-infected lymphocytes, and in degenerative brain lesions, including neurofibrillary tangles and senile plaques in AD. The previously reported pathologic similarities between AD and IBM prompted us to study alphaBC expression in IBM. METHODS: Immunolocalization of alphaBC in muscle of 11 patients with IBM, 50 patients with other muscle diseases, and 4 controls; and quantitative analysis of the frequency of fibers with 1) increased alphaBC expression in IBM and polymyositis and 2) structural abnormality (vacuolated, non necrotic and invaded by mononuclear cells, Congo red-positive, SMI-31 positive, and ubiquitin positive) in IBM. RESULTS: We detected enhanced expression of alphaBC not only in all structurally abnormal IBM fibers, but also, and with severalfold higher frequency, in IBM fibers without significant structural abnormality (X fibers) (p values in paired t-tests < 0.001). We also found enhanced alphaBC in abnormal fibers in other diseases; X fibers, however, were extremely sparse or absent, except in two atypical cases of polymyositis refractory to immunotherapy. CONCLUSION: That the X fibers are much more frequent than the structurally abnormal fibers in IBM points to a pathogenic stressor acting upstream to the development of structural abnormalities. The identification of this stressor is now of paramount importance for deciphering the enigma of IBM. PMID- 10720272 TI - Transplantation of embryonic porcine mesencephalic tissue in patients with PD. AB - OBJECTIVE: To assess the safety and the effect on standardized clinical rating measures of transplanted embryonic porcine ventral mesencephalic (VM) tissue in advanced PD. METHODS: Twelve patients with idiopathic PD underwent unilateral implantation of embryonic porcine VM tissue; six received cyclosporine immunosuppression and six received tissue treated with a monoclonal antibody directed against major histocompatibility complex class I. Patients were followed for 12 months and assessed by clinical examination, MRI, and 18F-levodopa PET. Porcine endogenous retrovirus testing was conducted by PCR-based method on peripheral blood mononuclear cells. RESULTS: Cell implantation occurred without serious adverse events in all patients. Cultures were negative for bacterial and unknown viral contamination. No porcine endogenous retrovirus DNA sequences were found. MRI demonstrated cannula tracts within the putamen and caudate, with minimal or no edema and no mass effect at the transplant sites. In the medication off state, total Unified Parkinson's Disease Rating Scale scores improved 19% (p = 0.01). Three patients improved over 30%. There were two patients with improved gait. 18F-levodopa PET failed to show changes on the transplanted side. CONCLUSIONS: Unilateral transplantation of porcine embryonic VM cells into PD patients was well tolerated with no evidence of transmission of porcine endogenous retrovirus. Changes in standardized clinical PD rating measures were variable, similar to the results of the first trials of unilateral human embryonic allografts that transplanted small amounts of tissue. PMID- 10720273 TI - Prospective validation of consensus criteria for the diagnosis of dementia with Lewy bodies. AB - OBJECTIVE: To determine the validity of a clinical diagnosis of probable or possible dementia with Lewy bodies (DLB) made using International Consensus criteria. BACKGROUND: Validation studies based on retrospective chart reviews of autopsy-confirmed cases have suggested that diagnostic specificity for DLB is acceptable but case detection rates as low as 0.22 have been suggested. METHODS: We evaluated the first 50 cases reaching neuropathologic autopsy in a cohort to which Consensus clinical diagnostic criteria for DLB, National Institute for Neurological and Communicative Disorders and Stroke-Alzheimer's Disease and Related Disorders Association criteria for AD, and National Institute of Neurological Disorders and Stroke-Association Internationale pour la Recherche et l'Enseignement en Neurosciences criteria for vascular dementia (VaD) had been prospectively applied. RESULTS: Twenty-six clinical diagnoses of DLB, 19 of AD, and 5 of VaD were made. At autopsy, 29 DLB cases, 15 AD, 5 VaD, and 1 progressive supranuclear palsy were identified. The sensitivity and specificity of a clinical diagnosis of probable DLB in this sample were 0.83 and 0.95. Of the five cases receiving a false-negative diagnosis of DLB, significant fluctuation was present in four but visual hallucinations and spontaneous motor features of parkinsonism were generally absent. Thirty-one percent of the DLB cases had additional vascular pathology and in two cases this contributed to a misdiagnosis of VaD. No correlations were found between the distribution of Lewy bodies and clinical features. CONCLUSION: The Consensus criteria for DLB performed as well in this prospective study as those for AD and VaD, with a diagnostic sensitivity substantially higher than that reported by previous retrospective studies. DLB occurs in the absence of extrapyramidal features and in the presence of comorbid cerebrovascular disease. Fluctuation is an important diagnostic indicator, reliable measures of which need to be developed further. PMID- 10720274 TI - Unilateral pallidotomy in PD: a controlled study of cognitive and behavioral effects. The Netherlands Pallidotomy Study (NEPAS) group. AB - OBJECTIVE: To investigate whether unilateral pallidotomy affects cognitive and behavioral functioning. METHODS: At baseline and after 6 months we assessed neuropsychological functioning in 35 patients with advanced PD. After baseline examination, patients were randomized to pallidotomy within 1 month (6 left sided, 13 right-sided) or to pallidotomy after follow-up assessment 6 months later (n = 16; control group). We performed neuropsychological tests of language, visuospatial function, memory, attention, and executive functions. Self ratings and proxy ratings of memory problems and dysexecutive symptoms were also collected. RESULTS: No significant differences over time were found between pallidotomy and control groups, with the exception of a decrease of verbal fluency in the left-sided pallidotomy group. CONCLUSIONS: Unilateral pallidotomy is relatively safe with respect to cognition and behavior. Left-sided pallidotomy may lead to minor deterioration in verbal fluency. The sample size of this study is too small, however, to rule out the possibility of infrequent but clinically important side effects. PMID- 10720275 TI - Prevalence and risk factors of RLS in an elderly population: the MEMO study. Memory and Morbidity in Augsburg Elderly. AB - OBJECTIVE: To evaluate prevalence, sociodemographic characteristics, and risk factors of restless legs syndrome (RLS) in a population-based survey of the elderly, using standard diagnostic criteria. BACKGROUND: Population-based studies of RLS are rare and have not yet used standard definition criteria. METHODS: The Memory and Morbidity in Augsburg Elderly (MEMO) Study is a follow-up project of the World Health Organization Monitoring Trends and Determinants in Cardiovascular Disease (MONICA) Survey-Augsburg, Germany, 1989-1990, evaluating neurologic diseases and their risk factors in a German population 65 to 83 years of age. Two RLS-trained physicians assessed the prevalence of RLS based on the four minimal standard criteria (International Restless Legs Syndrome Study Group, 1995) using standardized questions in face-to-face interviews. They also obtained information on medical history, medications, depression (Center of Epidemiologic Studies Depression Scale), and quality of life (Short Form 36) and performed a standardized neurologic examination for each participant. RESULTS: The study population included 369 participants (173 women and 196 men). The overall prevalence of RLS was 9.8% (n = 36) and higher in women (13.9% versus 6.1%; p = 0.02). In women, the prevalence did not change with age, whereas men showed a nonsignificant inverse trend with increasing age. RLS-positive individuals took more benzodiazepines and estrogen compared with non-RLS cases, but the differences were not statistically significant. Participants with RLS had higher incidence of depression (p = 0.012) and lower self-reported mental health scores (p = 0.029) than did non-RLS cases. CONCLUSIONS: RLS is a frequent syndrome in the elderly with considerable impact on self-perceived mental health, affecting women about twice as often as men. PMID- 10720276 TI - Decreased striatal dopamine transporter density in JNCL patients with parkinsonian symptoms. AB - OBJECTIVE: To explore whether striatal dopamine transporters are involved in juvenile neuronal ceroid lipofuscinosis (JNCL) with extrapyramidal signs. METHODS: Seventeen patients with JNCL entered the study (mean age, 15 years; age range, 10 to 31 years). For clinical evaluation, the authors used the motor section of the Unified Parkinson's Disease Rating Scale (UPDRS). For studying the density of dopamine transporters in the striatum, they employed iodine-123 labeled 2beta-carbomethoxy-3beta-(4-iodophenyl) tropane as a SPECT tracer. The SPECT images were evaluated visually, and tracer accumulation was semiquantified from transverse slices as striatum-to-cerebellum activity ratios. MRI (1.5-T) signal intensities of the striatum were measured and compared with those of the thalamus. RESULTS: The mean UPDRS score was 20 (range, 2 to 41). On SPECT, the mean striatum-to-cerebellum uptake ratio was lower in patients than in control subjects (3.1 +/- 0.6 versus 6.8 +/- 1.0; p < 0.001), with the decrease being more pronounced in the putamen than in the caudate nucleus. On MRI, the mean striatum-to-thalamus signal intensity ratio was higher in patients than in control subjects (1.14 +/- 0.02 versus 1.08 +/- 0.02; p < 0.001). There was a negative correlation between uptake ratios in SPECT and UPDRS scores, and a positive correlation between the MRI ratios and UPDRS. The SPECT and MRI ratios also correlated significantly, providing additional evidence for the contributions of nigrostriatal, striatal, and thalamic dysfunction to the parkinsonian symptoms. CONCLUSIONS: The observed decrease in the striatal dopamine transporter density in JNCL offers a rational basis for a trial of dopaminergic drugs in this disease. PMID- 10720277 TI - Homogeneous phenotype of the gypsy limb-girdle MD with the gamma-sarcoglycan C283Y mutation. AB - OBJECTIVE: To characterize the clinical phenotype of LGMD2C in gypsies. BACKGROUND: Limb-girdle muscular dystrophy (LGMD) in gypsies of Western Europe is caused by a homozygous C283Y mutation on the same haplotype, suggesting a founder effect. METHODS: We performed clinical, laboratory, and muscle imaging studies of 40 patients. RESULTS: Mean age at onset was 5.3 years. One half of the patients had loss of ambulation by the age of 12; 13% still could walk after age 16. Calf hypertrophy, scapular winging, macroglossia, and lumbar hyperlordosis were common. Girdle, trunk, and proximal limb flexor muscles had earlier and more severe involvement. Cardiomyopathy was not observed. Five patients in the third decade of life required mechanical ventilation. Scoliosis was common in the nonambulatory stage. CONCLUSIONS: LGMD2C in gypsy patients with C283Y mutation presents a rather homogeneous phenotype, characterized by an initial Duchenne like progressive course followed by a more prolonged survival rate possibly due to the absence of early respiratory impairment and cardiac failure. PMID- 10720278 TI - A phase II trial of nerve growth factor for sensory neuropathy associated with HIV infection. AIDS Clinical Trials Group Team 291. AB - OBJECTIVE: To evaluate the safety and efficacy of recombinant human nerve growth factor (rhNGF) in HIV-associated sensory neuropathy (SN) within a multicenter, placebo-controlled, randomized trial (ACTG 291). BACKGROUND: SN affects 30% of individuals with AIDS, is worsened by neurotoxic antiretrovirals, and its treatment is often ineffective. NGF is trophic for small sensory neurons and stimulates the regeneration of damaged nerve fibers. METHODS: A total of 270 patients with HIV-associated SN were randomized to receive placebo, 0.1 microg/kg rhNGF, or 0.3 microg/kg rhNGF by double-blinded subcutaneous injection twice weekly for 18 weeks. The primary outcome was change in self-reported neuropathic pain intensity (Gracely Pain Scale). Secondary outcomes included an assessment of global improvement in neuropathy by patients and investigators, neurologic examination, use of prescription analgesics, and quantitative sensory testing. In a subset, epidermal nerve fiber densities were determined in punch skin biopsies. RESULTS: Both doses of NGF produced significant improvements in average and maximum daily pain compared with placebo. Positive treatment effects were also observed for global pain assessments (p = 0.001) and for pin sensitivity (p = 0.019). No treatment differences were found with respect to mood, analgesic use, or epidermal nerve fiber densities. Injection site pain was the most frequent adverse event, and resulted in unblinding in 39% of subjects. Severe transient myalgic pain occurred in eight patients, usually from accidental overdosing. There were no changes in HIV RNA levels or other laboratory indices. CONCLUSIONS: We found a positive effect of recombinant human nerve growth factor on neuropathic pain and pin sensitivity in HIV-associated sensory neuropathy. rhNGF was safe and well tolerated, but injection site pain was frequent. PMID- 10720279 TI - HIV-1-related encephalopathy in infants compared with children and adults. French Pediatric HIV Infection Study and the SEROCO Group. AB - OBJECTIVE: To characterize the specificities of HIV-1-related encephalopathy in children. METHODS: Comparison of patients from the French Perinatal Cohort of children born to HIV-1-infected mothers and followed from birth with the French SEROCO Cohort of adults with a known date of infection. Our study examines 1) the characteristics of encephalopathy with onset before 1 year, after 1 year, and in adults, and 2) the maternal and birth characteristics of infants who developed AIDS before 1 year and went on to develop either encephalopathy or opportunistic infection. RESULTS: The incidence of encephalopathy was higher in children than in adults during the first year (9.9% versus 0.3%) and intermediate during the second year (4.2% versus 0%) after infection but was similar thereafter (less than 1% per year in each group). The resulting cumulative incidence at 7 years postinfection reached 16% in children and 5% in adults. Encephalopathy that developed before 1 year 1) was more frequently an isolated symptom of AIDS, 2) was associated with a reduction of intrauterine brain growth, 3) was associated with a very low level of HIV-1 RNA in CSF, 4) occurred at a higher level of immunocompetence after taking into account the decrease in CD4 lymphocytes with age, and 5) was not prevented by zidovudine treatment during gestation. CONCLUSIONS: Early encephalopathy in infants has a different pathophysiologic mechanism than that occurring in children, which in turn shows similarities with that observed in adults. Early encephalopathy is probably related to the occurrence of pathologic events during late fetal life. PMID- 10720280 TI - Diagnosis of Creutzfeldt-Jakob disease: effect of clinical criteria on incidence estimates. AB - OBJECTIVE: To assess the effect of usage of three different versions of Creutzfeldt-Jakob disease (CJD) diagnostic criteria on estimates of CJD incidence. METHODS: A total of 428 patients referred for suspected sporadic CJD between 1991 and 1997 were classified according to different criteria to be compared after analysis of medical records. Specificity, sensitivity, and positive and negative predictive values were calculated for each set of criteria in the subgroup of patients with a postmortem examination. Positive and negative predictive values of the clinical diagnosis were applied to cases without postmortem examination. Subsequently, the true number of cases of CJD among the referred cases was estimated. RESULTS: By comparison with the French and European study criteria, the Masters' criteria showed higher sensitivity but lower specificity and positive predictive value. Comparison with an estimate of the true total number of CJD cases showed that Masters' criteria overestimated the incidence by 7%, whereas the French and the European study criteria led to an underestimate of 12%. Detection of the 14-3-3 protein in CSF, considered as an additional diagnostic criterion for clinically probable CJD, resulted in a slight increase in the estimated incidence when the French or European study criteria were applied. CONCLUSIONS: Different diagnostic criteria could lead to an under- or overestimation of the true incidence of CJD. Therefore, comparisons of CJD incidence in different countries should rely on diagnostic classifications using identical criteria. Taking into account 14-3-3 protein detection as a criterion for probable CJD will result in a small increase in the estimated CJD incidence. PMID- 10720281 TI - Decreased beta-amyloid1-42 in cerebrospinal fluid of patients with Creutzfeldt Jakob disease. AB - OBJECTIVES: Decreased levels of Abeta1-42 are found in CSF of patients with AD. Because early stages of Creutzfeldt-Jakob disease (CJD) and AD share several clinical features, we investigated Abeta1-42 levels in CSF of these groups, inferring that this might give additional help in differentiating patients with CJD from AD patients. METHODS: We investigated 27 patients with CJD, 14 patients with AD, 19 patients with other dementias, and 20 nondemented controls (NDC) for Abeta1-42 in CSF. Twenty-four of the 27 CJD patients were neuropathologically verified. All the neuropathologically verified patients presented with a type 1 prion protein pattern. CJD patients were all homozygous for methionine at codon 129. Except in five CJD patients, no beta-amyloid plaques were seen. Additionally, APOE status was determined in patients with CJD. RESULTS: Levels of Abeta1-42 in CSF were decreased in patients with AD as well as in CJD. Levels of Abeta1-42 in CSF of patients with CJD and AD were significantly different from the other dementia and NDC groups. There was no substantial difference between the CJD and AD groups (p = 0.66). Decreased levels of Abeta1-42 did not correlate with the APOE epsilon4 load in patients with CJD. CONCLUSION: Low levels of Abeta1-42 in CSF do not exclude a diagnosis of CJD. Decreased levels of Abeta1-42 in CSF can occur without beta-amyloid plaque formation in the brain. However, the underlying mechanism of this phenomenon must be elucidated. PMID- 10720282 TI - Variant Alzheimer's disease with spastic paraparesis: clinical characterization. AB - OBJECTIVE: To present the clinical, neuroimaging, and electrophysiologic characteristics of a variant AD phenotype. BACKGROUND: The authors have identified a large Finnish kindred with presenile dementia and spastic paraparesis due to deletion of exon 9 of presenilin 1. Neuropathologic analysis showed unusual cortical "cotton wool" plaques, immunoreactive for the beta amyloid peptide but lacking congophilic cores. PATIENTS AND METHODS: Twenty-two affected individuals (16 men and 6 women) were identified in four successive generations. All surviving five patients were examined and subjected to molecular genetic analysis. In addition, the neurologic records of nine deceased patients were evaluated. Electrophysiologic investigations were available in eight cases. CT or MRI of the head had been performed on 11 patients and PET was performed on three patients. RESULT: The mean age at onset (+/-SD) was 50.9 +/- 5.2 years (range 40 to 61 years). Memory impairment was present in all patients. Memory impairment appeared simultaneously with or was preceded by walking difficulty due to spasticity of the lower extremities (10/14). Impaired fine coordination of hands (9/14) and dysarthria (6/14) in some patients suggested cerebellar involvement. EEG showed intermittent generalized delta-theta activity. Head MRI showed temporal and hippocampal atrophy; PET showed bilateral temporo-parietal hypometabolism. CONCLUSION: Spastic paraparesis or brisk stretch reflexes of lower extremities or clumsiness of hands combined with dementia suggests this variant of AD. PMID- 10720283 TI - Ten-year incidence of dementia in a rural elderly US community population: the MoVIES Project. AB - OBJECTIVE: To determine incidence rates by age, sex, and education of overall dementia and probable/ possible AD in a largely rural community. METHODS: Ten year prospective study of a randomly selected community sample aged 65+; biennial cognitive screening followed by standardized clinical evaluation. Incidence rates were estimated for overall dementia (Diagnostic and Statistical Manual of Mental Disorders, 3rd ed., revised, criteria and Clinical Dementia Rating [CDR]) and for probable/possible AD (National Institute of Neurological and Communicative Disorders and Stroke-Alzheimer's Disease and Related Disorders Association criteria). RESULTS: The cohort consisted of 1,298 individuals free of dementia at study entry. Among these, 199 incident (new) cases of overall (all-cause) dementia with CDR stage > or = 0.5, including 110 with CDR > or = 1, were detected during follow-up. Among the incident cases, 153 (76.9%) had probable/ possible AD. Age-specific incidence rates are reported for all dementia and for probable/possible AD, by sex and CDR stage. Among all-cause dementias with CDR = 0.5, controlling for age and education, men had a higher incidence rate than women. In the same group, those with less than high school education had significantly higher incidence rates than those with more education. Rates did not vary significantly by sex or education for probable/possible AD or for dementia with CDR > or = 1. CONCLUSIONS: Incidence rates of all dementias and of AD increased with age; men and those with lesser education had higher rates of possible/incipient dementia (CDR = 0.5) in this community. Potential explanations for these sex and education effects are discussed. PMID- 10720284 TI - The anatomy of aphasia revisited. AB - OBJECTIVE: To determine lesion locations associated with the various types of aphasic disorders in patients with stroke. BACKGROUND: The anatomy of aphasia has been challenged by several recent studies. Discrepancies are likely to be due to methodologic issues. METHODS: We examined 107 patients with a standardized aphasia battery and MRI. Three examiners blinded to the clinical data rated signal abnormalities in 69 predetermined regions of interest. The statistical procedure used classification tree testing, which selected regions associated with each aphasic disorder. RESULTS: 1) Nonfluent aphasia depended on the presence of frontal or putaminal lesions; 2) repetition disorder on insula external capsule lesions; 3) comprehension disorder on posterior lesions of the temporal gyri; 4) phonemic paraphasia on external capsule lesions extending either to the posterior part of the temporal lobe or to the internal capsule; 5) verbal paraphasia on temporal or caudate lesions; and 6) perseveration on caudate lesions. These analyses correctly classified 67% to 94% of patients. CONCLUSIONS: Lesion location is the main determinant of aphasic disorders at the acute stage. Most clinical-radiologic correlations supported the classic anatomy of aphasia. PMID- 10720285 TI - Dural arteriovenous fistula mimicking leukoencephalopathy. PMID- 10720286 TI - Frequency and clinical determinants of dementia after ischemic stroke. AB - OBJECTIVE: To investigate the frequency and clinical determinants of dementia after ischemic stroke. METHODS: The authors administered neurologic, neuropsychological, and functional assessments to 453 patients (age 72.0 +/- 8.3 years) 3 months after ischemic stroke. They diagnosed dementia using modified Diagnostic and Statistical Manual of Mental Disorders, 3rd ed., revised criteria requiring deficits in memory and two or more additional cognitive domains as well as functional impairment. RESULTS: The authors diagnosed dementia in 119 of the 453 patients (26.3%). Regarding dementia subtypes, 68 of the 119 patients (57.1%) were diagnosed with vascular dementia, 46 patients (38.7%) were diagnosed with AD with concomitant stroke, and 5 patients (4.2%) had dementia for other reasons. Logistic regression suggested that dementia was associated with a major hemispheral stroke syndrome (OR 3.0), left hemisphere (OR 2.1) and right hemisphere (OR 1.8) infarct locations versus brainstem/cerebellar locations, infarcts in the pooled anterior and posterior cerebral artery territories versus infarcts in other vascular territories (OR 1.7), diabetes mellitus (OR 1.8), prior stroke (OR 1.7), age 80 years or older (OR 12.7) and 70 to 79 years (OR 3.9) versus 60 to 69 years, 8 or fewer years of education (OR 4.1) and 9 to 12 years of education (OR 3.0) versus 13 or more years of education, black race (OR 2.6) and Hispanic ethnicity (OR 3.1) versus white race, and northern Manhattan residence (OR 1.6). CONCLUSIONS: Dementia is frequent after ischemic stroke, occurring in one-fourth of the elderly patients in the authors' cohort. The clinical determinants of dementia include the location and severity of the presenting stroke, vascular risk factors such as diabetes mellitus and prior stroke, and host characteristics such as older age, fewer years of education, and nonwhite race/ethnicity. The results also suggest that concomitant AD plays an etiologic role in approximately one-third of cases of dementia after stroke. PMID- 10720287 TI - Mild familial neurofibromatosis 2 associates with expression of merlin with altered COOH-terminus. AB - OBJECTIVE: To understand molecular details of the pathogenesis of a very mild and homogenous form of neurofibromatosis 2 (NF2). BACKGROUND: Inactivation of the NF2 tumor suppressor gene leads to the development of multiple nervous system tumors, accompanied by loss of the NF2 gene product, merlin, or schwannomin. The severity of disease varies between patients, and the biologic basis of this variation is poorly understood. METHODS: We studied the genotype-phenotype correlation in a large pedigree with extremely mild and uniform disease manifesting as slowly growing bilateral vestibular nerve schwannomas of late onset. RESULTS: The tumors demonstrated a low proliferation rate and loss of the wild-type NF2 allele. The disease is caused by a novel mutation in the NF2 gene at intron 15 splice donor site (1737 + 3 a --> t), which was identified in all carriers by the minisequencing method. The mutation resulted in splicing out of exon 15 and production of two transcripts: a novel mutant transcript with exon 16 and overexpression of isoform III, normally detected at a low level. Both transcripts encode for the COOH-terminus of isoform III. Immunoblotting of patient fibroblasts and tumor tissue demonstrated variant merlin with altered COOH terminus. CONCLUSIONS: The mutational skip of exon 15 and the expression of a protein with the COOH-terminus of isoform III correlates with the exceptionally mild NF2, and suggests tumor suppressor activity for isoform III. The detection of expressed mutant proteins may provide useful information for prediction of the clinical outcome of individual mutations. PMID- 10720288 TI - Pathologic damage in MS assessed by diffusion-weighted and magnetization transfer MRI. AB - OBJECTIVE: To compare diffusion characteristics of MS lesions, normal-appearing white matter (NAWM) from patients, and normal white matter from control subjects, and to investigate the correlations between the magnetization transfer ratio (MTR) and a directionally averaged tissue water diffusion coefficient (D) in patients. BACKGROUND: MS and other pathologic processes that modify tissue integrity can result in abnormal diffusion of water molecules detectable by diffusion-weighted imaging (DWI). METHODS: Conventional dual-echo and DWI scans were obtained from 35 patients with relapsing-remitting MS and 24 healthy control subjects. MT scans were also obtained from the patients. After coregistration of all scans, MTR and D values from MS lesions and NAWM in different regions were marked using the dual-echo scans as a reference. D values from the same brain regions in control subjects were acquired. Histograms of MTR and D were also produced. RESULTS: Patients with MS had significantly higher D values in all the areas studied. Moreover, histogram metrics (peak height, peak site, and average D) from patients were substantially different from those of control subjects. In patients, average lesion D and MTR were markedly different from those in the NAWM. There was an inverse correlation between average lesion MTR and D inside lesions, whereas no correlation was found for average MTR and D taken from the histograms. CONCLUSIONS: DWI detects severe tissue disruption inside lesions and subtle widespread abnormalities in NAWM in patients with relapsing-remitting MS. MT and DWI may provide information about different aspects of brain pathology in MS. PMID- 10720289 TI - Cladribine and progressive MS: clinical and MRI outcomes of a multicenter controlled trial. Cladribine MRI Study Group. AB - OBJECTIVE: To evaluate the safety and efficacy of two doses of cladribine in patients with progressive MS. BACKGROUND: Treatment of progressive MS patients with cladribine in a previous single-center, placebo-controlled clinical trial was associated with disease stabilization. METHODS: In the current study, 159 patients with a median baseline Kurtzke's Expanded Disability Status Scale (EDSS) score of 6.0 were randomly assigned to receive placebo or cladribine 0.07 mg/kg/day for 5 consecutive days every 4 weeks for either two or six cycles (total dose, 0.7 mg/kg or 2.1 mg/kg, respectively), followed by placebo, for a total of eight cycles. Thirty percent had primary progressive MS (PPMS) and 70% had secondary progressive MS (SPMS). EDSS and Scripps Neurologic Rating Scale (SNRS) scores were assessed bi-monthly and MRI was performed every 6 months. The primary outcome measure was disability (mean change in EDSS). RESULTS: Mean changes in disability did not differ among the groups at the end of the 12-month double-blind phase. Both cladribine treatments were superior to placebo for the proportion of patients having gadolinium-enhanced T1 lesions and for the mean volume and number of such lesions (p < or = 0.003). Differences were statistically significant at the 6-month evaluation time, with < or =90% reduction in volume and number of enhanced T1 lesions, which was maintained through final evaluation. This effect segregated largely with the SPMS group. The T2 burden of disease showed a modest improvement in cladribine-treated patients and worsened in placebo-treated patients. Most adverse events were mild or moderate in severity and not treatment limiting. CONCLUSION: No significant treatment effects were found for cladribine in terms of changes in EDSS or SNRS scores. Both doses of cladribine produced and sustained significant reductions in the presence, number, and volume of gadolinium-enhanced T1 brain lesions on MRI, and cladribine 2.1 mg/kg reduced the accumulation of T2 lesion load. Cladribine at doses up to 2.1 mg/kg was generally safe and well tolerated. PMID- 10720290 TI - A Wallerian degeneration pattern in patients at risk for MS. AB - BACKGROUND: Demyelination alone may not explain the progressive disability that frequently develops in MS. An alternative explanation for irreversible disability assumes a contribution from axonal injury or loss. In theory, axonal injury may occur in the focal areas characterized by early inflammation, or can be more distant, as in Wallerian degeneration. However, Wallerian degeneration is thought of as a rare or a late finding in MS. METHODS: Studies showing a classic Wallerian degeneration pattern in the corticospinal tract were selected from a review of MR studies from patients enrolled in a longitudinal treatment trial. Entry was based on first occurrence of an isolated neurologic syndrome consistent with MS and a positive MRI. RESULTS: This report is based on five cases followed longitudinally who showed development of a classic T2-hyperintense lesion along the ipsilateral corticospinal tract, subsequent to an initial inciting event located in the white matter located in the superior aspect of the corona radiata. Lesions were evident as T2-hyperintensity persisting throughout the 12 to 18 months of observation. CONCLUSIONS: This series suggests that Wallerian degeneration, implying axonal injury, may occur as a sequela of acute demyelinating lesions in patients presenting with their first symptoms suggestive of MS. This can produce a component of the increasing burden of T2-hyperintense lesions temporally and spatially dissociated from inflammatory or demyelinating activity. Further studies are required to determine if Wallerian degeneration is an important factor contributing to disability progression in MS. PMID- 10720291 TI - Preoperative FDG-PET temporal lobe hypometabolism and verbal memory after temporal lobectomy. AB - OBJECTIVE: To examine the relationship of preoperative fluorodeoxyglucose (FDG) PET asymmetry in temporal lobe metabolism and memory outcome after anterior temporal lobectomy (ATL). METHODS: In a university-based epilepsy surgery center, 60 ATL patients (27 left, 33 right) were divided into two groups: no/mild (n = 21) or moderate/ severe (n = 39) asymmetry in temporal lobe hypometabolism as determined by FDG-PET. All patients were nonretarded, at least 18 years of age, left-hemisphere speech dominant, without MRI abnormalities other than hippocampal atrophy, and with unilateral temporal lobe origin of intractable complex partial seizures. Neuropsychological measures of intelligence and verbal and visual memory function were assessed preoperatively and 6 months postoperatively. RESULTS: Left ATL patients with no/mild asymmetry in FDG-PET temporal lobe metabolism exhibited significantly greater verbal memory decline compared with left ATL patients with moderate/severe hypometabolism. There was no significant relationship between PET asymmetry and pre- to postsurgical IQ change. No significant relationship was observed between extent of PET hypometabolism and memory outcome for right ATL patients. CONCLUSIONS: FDG-PET asymmetry can be added to the preoperative clinical markers that appear useful in predicting verbal memory decline after left ATL. PMID- 10720292 TI - Randomized trial of modafinil as a treatment for the excessive daytime somnolence of narcolepsy: US Modafinil in Narcolepsy Multicenter Study Group. AB - OBJECTIVE: This is one of two separate clinical trials to evaluate the efficacy and safety of modafinil, a novel wake-promoting agent, in patients with excessive daytime sleepiness (EDS) associated with narcolepsy. METHODS: In this 9-week, randomized, placebo-controlled, double-blind, 21-center clinical trial, patients were randomized to receive fixed daily doses of modafinil 200 mg, modafinil 400 mg, or placebo. A placebo-controlled, 2-week treatment discontinuation phase was included to evaluate the effects of withdrawal on patients who had been receiving modafinil. A total of 271 patients who were naive to modafinil received study medication in the 9-week trial and 240 patients received study medication in the discontinuation phase. RESULTS: Treatment with modafinil resulted in significant improvement in two objective measures of EDS: the Multiple Sleep Latency Test and the Maintenance of Wakefulness Test. Additionally, patient self-assessment of sleepiness was significantly improved, as measured by the Epworth Sleepiness Scale, and level of illness was significantly reduced on the independent clinician assessment, the Clinical Global Impression of Change. Nighttime sleep, monitored by nocturnal polysomnography, was not adversely effected with modafinil treatment compared with placebo treatment. The most frequent adverse experience was headache, which was not significantly greater for modafinil than placebo. During treatment discontinuation, individuals who had been receiving modafinil experienced a return of their EDS to baseline levels. During treatment discontinuation, patients did not experience symptoms associated with amphetamine withdrawal syndrome. For up to 9 weeks of daily use there was no evidence for the development of dependence at the dose levels studied. CONCLUSION: The data indicate that modafinil has an excellent safety profile and is very well tolerated. Modafinil is an effective treatment for excessive daytime sleepiness in narcolepsy and shows continued efficacy with up to 9 weeks of daily use. PMID- 10720293 TI - Enlarged parietal foramina: association with cerebral venous and cortical anomalies. AB - OBJECTIVE: To evaluate a series of patients with enlarged parietal foramina for associated brain anomalies. BACKGROUND: Enlarged parietal foramina are usually considered a benign calvarial defect. METHODS: Ten patients with enlarged parietal foramina were identified. Seven patients were evaluated with neuroimaging: two by cranial CT and five by CT and/or MRI. Three patients who underwent MRI also underwent MR angiography or MR venography. RESULTS: Six of seven patients had cranial imaging showing a persistent falcine venous sinus. Three of six patients had variations of occipital cortical infolding. One patient had focal encephalomalacia in close proximity to the persistent falcine venous sinus and one had a previously undiagnosed atretic occipital encephalocele. CONCLUSION: This constellation of findings suggests that aberrant vascular evolution during fetal development may affect cerebrovascular, brain, or skull development. Individuals with enlarged parietal foramina (>5 mm) warrant imaging of underlying brain parenchyma and vasculature. PMID- 10720294 TI - Vagus nerve stimulation therapy for epilepsy in older adults. AB - The authors assessed the efficacy, safety, and tolerability of vagus nerve stimulation (VNS) for refractory epilepsy in 45 adults 50 years of age and older. They determined seizure frequency, adverse effects, and quality of life. At 3 months, 12 patients had a >50% decrease in seizure frequency; at 1 year, 21 of 31 studied individuals had a >50% seizure decrease. Side effects were mild and transient. Quality of life scores improved significantly with time. PMID- 10720295 TI - Benign childhood epilepsy with centrotemporal spikes: is it always benign? AB - Most children with benign childhood epilepsy with centrotemporal spikes have few seizures, and some have only one. We describe two children with interictal and ictal findings consistent with this epileptic syndrome but with severe intractable seizures and cognitive decline that resulted in consideration for epilepsy surgery. Spontaneous remission occured in one child; the other is still young. Despite the high seizure burden and cognitive decline, surgical consideration should be withheld, as these seizures are likely to remit. PMID- 10720296 TI - Topiramate therapy of epilepsy associated with Angelman's syndrome. AB - Angelman's syndrome, a genetic disorder involving a defect in the DNA coding for subunits of the gamma-aminobutyric acid (GABA) type A receptor, often is associated with intractable epilepsy. Topiramate is a novel anticonvulsant that enhances GABAergic neurotransmission. Five children with Angelman's syndrome and epilepsy were treated with topiramate for clinical indications. The drug was effective and well tolerated, possibly because of its GABAergic properties. Further studies are necessary to confirm and elucidate this observation. PMID- 10720297 TI - Phosphoglycerate kinase deficiency: an adult myopathic form with a novel mutation. AB - The authors report a 36-year-old man with exertional myoglobinuria and muscle cramp without hemolytic anemia or CNS symptoms. They found a deficiency of phosphoglycerate kinase (PGK) activity in muscle and erythrocytes and a 4-base pair deletion in exon 6 of the PGK gene. This mutation may cause a frameshift, yielding an abnormal stop codon in exon 6 by which a truncated PGK protein was produced. This phenotype is caused by a novel mutation of the PGK gene. PMID- 10720298 TI - Idiopathic trigeminal sensory neuropathy with gadolinium enhancement in the cisternal segment. AB - The authors report two patients with idiopathic trigeminal sensory neuropathy who showed gadolinium enhancement of the cisternal segment of the corresponding trigeminal nerve in cranial MRI. The resolution of these lesions in a repeat MRI suggests a similarity to Bell's palsy. PMID- 10720299 TI - Striatal dopamine in early-onset primary torsion dystonia with the DYT1 mutation. AB - Although nigrostriatal dopaminergic dysfunction has been suggested in early onset primary torsion dystonia (PTD) with the DYT1 mutation, the actual status of brain dopamine (DA) is unknown. In a DYT1 mutation-positive autopsy patient with PTD, we found that nigral cellularity was normal and that subregional striatal DA levels were within the control range, except for those in the rostral portions of the putamen and caudate nucleus (50% to 54% of control means). Our data suggest that the DYT1 mutation is not associated with significant damage to the nigrostriatal DA system, in keeping with the absence of parkinsonism and levodopa response in this disorder. PMID- 10720300 TI - Polymorphism of NACP-Rep1 in Parkinson's disease: an etiologic link with essential tremor? AB - An allele (263bp) of the nonamyloid component of plaques (NACP)-Repl polymorphism has shown association with sporadic PD in a German population. The authors studied this polymorphism in 100 American PD patients and 100 healthy controls. The authors also studied 46 essential tremor (ET) and 55 Huntington's disease (HD) patients. Allele 263bp was significantly higher in PD patients (OR = 3.86) and ET patients (OR = 6.42) but not HD patients, compared with healthy controls. The association of allele 263bp with PD and ET suggests a possible etiologic link between these two conditions. PMID- 10720301 TI - Primary hemifacial spasm and arterial hypertension: a multicenter case-control study. AB - In a case-control study, the authors found that arterial hypertension occurred more frequently among 115 patients with primary hemifacial spasm than among 115 neurologic controls matched for age (+/-5 years), sex, and referral center. The association was not confounded by education level, smoking history, diabetes, or other diseases (adjusted OR 2.64; 95% CI 1.3 to 5.33, p = 0.007). Hypertension was significantly associated with the outcome in the left-sided group (OR 4.0; 95% CI 1.4 to 11.5), but data concerning patients with right-sided spasm were inconclusive (OR 1.05; 95% CI 0.36 to 3.1). In our sample, hypertension either preceded or followed the onset of hemifacial spasm. PMID- 10720302 TI - Intravenous valproic acid for myoclonic status epilepticus. PMID- 10720303 TI - Insertion pain in needle electromyography can be reduced by simultaneous finger slapping. PMID- 10720304 TI - Equal male and female incidence of myasthenia gravis. PMID- 10720305 TI - Still's disease can cause neutrophilic meningitis. PMID- 10720306 TI - Transient tumor attacks. PMID- 10720307 TI - The use of felbamate in the treatment of patients with intractable epilepsy. PMID- 10720308 TI - Neuropathology in older people with disequilibrium of unknown cause. PMID- 10720309 TI - Relation of education to brain size in normal aging. PMID- 10720310 TI - Risk of Alzheimer's disease in relatives of Parkinson's disease patients with and without dementia. PMID- 10720311 TI - Intranasal sumatriptan for the acute treatment of migraine in children. PMID- 10720312 TI - Cerebral hemodynamic impairment: methods of measurement in association with stroke. PMID- 10720313 TI - Sodium cromoglycate on plasma protein exudation to topically-applied substance P in the tracheal airways of rats or guinea-pigs in vivo. AB - Sodium cromoglycate (SCG) was examined against substance P(SP)-induced plasma protein exudation in the trachea of anaesthetized rats and guinea-pigs in vivo to determine whether SCG is a tachykinin receptor antagonist in the airways. A segment of trachea was prepared in situ for continuous perfusion with normal saline. Plasma-derived protein in the perfusion fluid (airway lumen) was increased after topical application of SP (1 microM, 5 min contact). In rats, the SP response was not attenuated by iv SCG but was inhibited (29%) by topical SCG under certain experimental conditions and using a high concentration of SCG (500 microM, 5 min contact, 30 min before SP and with SP). The NK(1)receptor antagonist, RP 67 580 (67 microM) abolished the SP response in rats. Sodium cromoglycate did not inhibit the SP response in guinea-pigs (same protocol as in rats). Thus, SCG attenuates plasma protein exudation (and presumably microvascular leak) induced by SP in rat tracheal airways but, if SCG is a tachykinin receptor (NK(1)) antagonist, it not only lacks potency but is species selective, i.e. more effective in rats than in guinea-pigs. PMID- 10720314 TI - Drug discovery: a historical perspective. AB - Driven by chemistry but increasingly guided by pharmacology and the clinical sciences, drug research has contributed more to the progress of medicine during the past century than any other scientific factor. The advent of molecular biology and, in particular, of genomic sciences is having a deep impact on drug discovery. Recombinant proteins and monoclonal antibodies have greatly enriched our therapeutic armamentarium. Genome sciences, combined with bioinformatic tools, allow us to dissect the genetic basis of multifactorial diseases and to determine the most suitable points of attack for future medicines, thereby increasing the number of treatment options. The dramatic increase in the complexity of drug research is enforcing changes in the institutional basis of this interdisciplinary endeavor. The biotech industry is establishing itself as the discovery arm of the pharmaceutical industry. In bridging the gap between academia and large pharmaceutical companies, the biotech firms have been effective instruments of technology transfer. PMID- 10720315 TI - Target-oriented and diversity-oriented organic synthesis in drug discovery. AB - Modern drug discovery often involves screening small molecules for their ability to bind to a preselected protein target. Target-oriented syntheses of these small molecules, individually or as collections (focused libraries), can be planned effectively with retrosynthetic analysis. Drug discovery can also involve screening small molecules for their ability to modulate a biological pathway in cells or organisms, without regard for any particular protein target. This process is likely to benefit in the future from an evolving forward analysis of synthetic pathways, used in diversity-oriented synthesis, that leads to structurally complex and diverse small molecules. One goal of diversity-oriented syntheses is to synthesize efficiently a collection of small molecules capable of perturbing any disease-related biological pathway, leading eventually to the identification of therapeutic protein targets capable of being modulated by small molecules. Several synthetic planning principles for diversity-oriented synthesis and their role in the drug discovery process are presented in this review. PMID- 10720316 TI - Mechanism-based target identification and drug discovery in cancer research. AB - Cancer as a disease in the human population is becoming a larger health problem, and the medicines used as treatments have clear limitations. In the past 20 years, there has been a tremendous increase in our knowledge of the molecular mechanisms and pathophysiology of human cancer. Many of these mechanisms have been exploited as new targets for drug development in the hope that they will have greater antitumor activity with less toxicity to the patient than is seen with currently used medicines. The fruition of these efforts in the clinic is just now being realized with a few encouraging results. PMID- 10720317 TI - Harnessing the power of the genome in the search for new antibiotics. AB - Over the past 40 years, the search for new antibiotics has been largely restricted to well-known compound classes active against a standard set of drug targets. Although many effective compounds have been discovered, insufficient chemical variability has been generated to prevent a serious escalation in clinical resistance. Recent advances in genomics have provided an opportunity to expand the range of potential drug targets and have facilitated a fundamental shift from direct antimicrobial screening programs toward rational target-based strategies. The application of genome-based technologies such as expression profiling and proteomics will lead to further changes in the drug discovery paradigm by combining the strengths and advantages of both screening strategies in a single program. PMID- 10720318 TI - Monodisperse FePt nanoparticles and ferromagnetic FePt nanocrystal superlattices AB - Synthesis of monodisperse iron-platinum (FePt) nanoparticles by reduction of platinum acetylacetonate and decomposition of iron pentacarbonyl in the presence of oleic acid and oleyl amine stabilizers is reported. The FePt particle composition is readily controlled, and the size is tunable from 3- to 10 nanometer diameter with a standard deviation of less than 5%. These nanoparticles self-assemble into three-dimensional superlattices. Thermal annealing converts the internal particle structure from a chemically disordered face-centered cubic phase to the chemically ordered face-centered tetragonal phase and transforms the nanoparticle superlattices into ferromagnetic nanocrystal assemblies. These assemblies are chemically and mechanically robust and can support high-density magnetization reversal transitions. PMID- 10720319 TI - Efficient activation of aromatic C-H bonds for addition to C-C multiple bonds AB - Efficient electrophilic metalation of aromatic C-H bonds leading to new C-C bond formation through regio- and stereoselective addition to alkynes and alkenes has been realized by a catalytic amount (0.02 to 5 mole percent) of palladium(II) or platinum(II) compounds in a mixed solvent containing trifluoroacetic acid at room temperature. Various arenes undergo unexpected selective trans hydroarylation to terminal or internal C&cjs0812;C bonds inter- and intramolecularly with high efficiency (up to a turnover number of 4500 for palladium), especially for electron-rich arenes, giving thermodynamically unfavorable cis-alkenes, and the oxygen- and nitrogen-containing heterocycles. The simplicity, generality, and efficiency of this process should be very attractive to the possible industrial application for the functionalization of arenes. PMID- 10720320 TI - Thermal, catalytic, regiospecific functionalization of alkanes AB - The formation of a single product from terminal functionalization of linear alkanes from a transition metal-catalyzed reaction is reported. The rhodium complex Cp*Rh(eta(4)-C(6)Me(6)) (Cp*, C(5)Me(5); Me, methyl) catalyzes the high yield formation of linear alkylboranes from commercially available borane reagents under thermal conditions. These reactions now allow catalytic, regiospecific functionalization of alkanes under thermal conditions. The organoborane products are among the most versatile synthetic intermediates in chemistry and serve as convenient precursors to alcohols, amines, and other common classes of functionalized molecules. PMID- 10720321 TI - Detection of SO in Io's exosphere. AB - The Galileo orbiter's close pass by Io in 1995 produced evidence for extensive mass loading of the plasma torus through the ionization of SO2. On 11 October 1999, Galileo passed even closer to Io, this time across the upstream side relative to the flow of magnetospheric plasma that corotates with Jupiter. On the first flyby, ion cyclotron waves gave direct evidence for the production of SO2+ ions. On the second flyby, ion cyclotron waves associated with SO+ were stronger and more persistent. Moreover, SO+ emissions were seen closer to Io than SO2+ emissions, suggesting that the exosphere was spatially inhomogeneous. The location of the waves suggests a fan-shaped region of ion pickup extending in the anti-Jupiter direction. Because the wave spectra were different even where the 1995 and 1999 trajectories crossed, we infer that Io's exosphere is temporally variable. PMID- 10720322 TI - Secular variation of iron isotopes in north atlantic deep water AB - A high-precision iron isotope time series for a ferromanganese crust demonstrates that the iron isotope composition in North Atlantic Deep Water has changed substantially over the past 6 million years and that iron isotope variations in the crust are closely correlated to those of lead isotopes. The close correlation between the two isotope series indicates that the observed iron isotope variations predominantly reflect those of iron input from terrigenous sources and provides no evidence for biologically induced mass fractionation within North Atlantic Deep Water. PMID- 10720323 TI - Modulation of hurricane activity in the gulf of mexico by the madden-julian oscillation AB - The Madden-Julian oscillation (MJO) is a large-scale episodic modulation of tropical winds and precipitation that travels eastward from Asia to America, with a characteristic repeat time of 30 to 60 days. Here it is shown that when MJO wind anomalies in the lower troposphere of the eastern Pacific are westerly, Gulf of Mexico and western Caribbean hurricane genesis is four times more likely than when the MJO winds are easterly. Accurate predictions of the MJO may lead to improved long-range forecasts of tropical cyclone activity. PMID- 10720324 TI - Contribution of increasing CO2 and climate to carbon storage by ecosystems in the United States. AB - The effects of increasing carbon dioxide (CO2) and climate on net carbon storage in terrestrial ecosystems of the conterminous United States for the period 1895 1993 were modeled with new, detailed historical climate information. For the period 1980-1993, results from an ensemble of three models agree within 25%, simulating a land carbon sink from CO2 and climate effects of 0.08 gigaton of carbon per year. The best estimates of the total sink from inventory data are about three times larger, suggesting that processes such as regrowth on abandoned agricultural land or in forests harvested before 1980 have effects as large as or larger than the direct effects of CO2 and climate. The modeled sink varies by about 100% from year to year as a result of climate variability. PMID- 10720325 TI - Cell surface engineering by a modified Staudinger reaction. AB - Selective chemical reactions enacted within a cellular environment can be powerful tools for elucidating biological processes or engineering novel interactions. A chemical transformation that permits the selective formation of covalent adducts among richly functionalized biopolymers within a cellular context is presented. A ligation modeled after the Staudinger reaction forms an amide bond by coupling of an azide and a specifically engineered triarylphosphine. Both reactive partners are abiotic and chemically orthogonal to native cellular components. Azides installed within cell surface glycoconjugates by metabolism of a synthetic azidosugar were reacted with a biotinylated triarylphosphine to produce stable cell-surface adducts. The tremendous selectivity of the transformation should permit its execution within a cell's interior, offering new possibilities for probing intracellular interactions. PMID- 10720326 TI - A fossil snake with limbs. AB - A 95-million-year-old fossil snake from the Middle East documents the most extreme hindlimb development of any known member of that group, as it preserves the tibia, fibula, tarsals, metatarsals, and phalanges. It is more complete than Pachyrhachis, a second fossil snake with hindlimbs that was recently portrayed to be basal to all other snakes. Phylogenetic analysis of the relationships of the new taxon, as well as reanalysis of Pachyrhachis, shows both to be related to macrostomatans, a group that includes relatively advanced snakes such as pythons, boas, and colubroids to the exclusion of more primitive snakes such as blindsnakes and pipesnakes. PMID- 10720327 TI - Prostaglandin D2 as a mediator of allergic asthma. AB - Allergic asthma is caused by the aberrant expansion in the lung of T helper cells that produce type 2 (TH2) cytokines and is characterized by infiltration of eosinophils and bronchial hyperreactivity. This disease is often triggered by mast cells activated by immunoglobulin E (IgE)-mediated allergic challenge. Activated mast cells release various chemical mediators, including prostaglandin D2 (PGD2), whose role in allergic asthma has now been investigated by the generation of mice deficient in the PGD receptor (DP). Sensitization and aerosol challenge of the homozygous mutant (DP-/-) mice with ovalbumin (OVA) induced increases in the serum concentration of IgE similar to those in wild-type mice subjected to this model of asthma. However, the concentrations of TH2 cytokines and the extent of lymphocyte accumulation in the lung of OVA-challenged DP-/- mice were greatly reduced compared with those in wild-type animals. Moreover, DP /- mice showed only marginal infiltration of eosinophils and failed to develop airway hyperreactivity. Thus, PGD2 functions as a mast cell-derived mediator to trigger asthmatic responses. PMID- 10720328 TI - Facile detection of mitochondrial DNA mutations in tumors and bodily fluids. AB - Examination of human bladder, head and neck, and lung primary tumors revealed a high frequency of mitochondrial DNA (mtDNA) mutations. The majority of these somatic mutations were homoplasmic in nature, indicating that the mutant mtDNA became dominant in tumor cells. The mutated mtDNA was readily detectable in paired bodily fluids from each type of cancer and was 19 to 220 times as abundant as mutated nuclear p53 DNA. By virtue of their clonal nature and high copy number, mitochondrial mutations may provide a powerful molecular marker for noninvasive detection of cancer. PMID- 10720329 TI - Reversal of antipsychotic-induced working memory deficits by short-term dopamine D1 receptor stimulation. AB - Chronic blockade of dopamine D2 receptors, a common mechanism of action for antipsychotic drugs, down-regulates D1 receptors in the prefrontal cortex and, as shown here, produces severe impairments in working memory. These deficits were reversed in monkeys by short-term coadministration of a D1 agonist, ABT 431, and this improvement was sustained for more than a year after cessation of D1 treatment. These findings indicate that pharmacological modulation of the D1 signaling pathway can produce long-lasting changes in functional circuits underlying working memory. Resetting this pathway by brief exposure to the agonist may provide a valuable strategy for therapeutic intervention in schizophrenia and other dopamine dysfunctional states. PMID- 10720330 TI - Start sites of bidirectional DNA synthesis at the human lamin B2 origin. AB - The initiation sites of bidirectional synthesis at the DNA replication origin located at the 3' end of the human lamin B2 gene were investigated. RNA-primed nascent DNA molecules were subjected to second-strand synthesis with appropriate primers, amplified by ligation-mediated polymerase chain reaction, and size fractionated. Evidence for precise start sites was obtained. Exploration of close to 1 kilobase, coupled to inhibition of Okazaki fragment synthesis, demonstrates that the leading strands initiate at precise nucleotides on either helix, overlapping by three base pairs, within the area bound to a protein complex possibly analogous to the prereplicative complex of yeast. PMID- 10720331 TI - A role for nuclear inositol 1,4,5-trisphosphate kinase in transcriptional control. AB - Phospholipase C and two inositol polyphosphate (IP) kinases constitute a signaling pathway that regulates nuclear messenger RNA export through production of inositol hexakisphosphate (IP6). The inositol 1,4,5-trisphosphate kinase of this pathway in Saccharomyces cerevisiae, designated Ipk2, was found to be identical to Arg82, a regulator of the transcriptional complex ArgR-Mcm1. Synthesis of inositol 1,4,5,6-tetrakisphosphate, but not IP6, was required for gene regulation through ArgR-Mcm1. Thus, the phospholipase C pathway produces multiple IP messengers that modulate distinct nuclear processes. The results reveal a direct mechanism by which activation of IP signaling may control gene expression. PMID- 10720332 TI - Rapid extragranular plasticity in the absence of thalamocortical plasticity in the developing primary visual cortex. AB - Monocular deprivation during early postnatal development remodels the circuitry of the primary visual cortex so that most neurons respond poorly to stimuli presented to the deprived eye. This rapid physiological change is ultimately accompanied by a matching anatomical loss of input from the deprived eye. This remodeling is thought to be initiated at the thalamocortical synapse. Ocular dominance plasticity after brief (24 hours) monocular deprivation was analyzed by intrinsic signal optical imaging and by targeted extracellular unit recordings. Deprived-eye responsiveness was lost in the extragranular layers, whereas normal binocularity in layer IV was preserved. This finding supports the hypothesis that thalamocortical organization is guided by earlier changes at higher stages. PMID- 10720333 TI - Retinal stem cells in the adult mammalian eye. AB - The mature mammalian retina is thought to lack regenerative capacity. Here, we report the identification of a stem cell in the adult mouse eye, which represents a possible substrate for retinal regeneration. Single pigmented ciliary margin cells clonally proliferate in vitro to form sphere colonies of cells that can differentiate into retinal-specific cell types, including rod photoreceptors, bipolar neurons, and Muller glia. Adult retinal stem cells are localized to the pigmented ciliary margin and not to the central and peripheral retinal pigmented epithelium, indicating that these cells may be homologous to those found in the eye germinal zone of other nonmammalian vertebrates. PMID- 10720334 TI - Motion integration and postdiction in visual awareness. AB - In the flash-lag illusion, a flash and a moving object in the same location appear to be offset. A series of psychophysical experiments yields data inconsistent with two previously proposed explanations: motion extrapolation (a predictive model) and latency difference (an online model). We propose an alternative in which visual awareness is neither predictive nor online but is postdictive, so that the percept attributed to the time of the flash is a function of events that happen in the approximately 80 milliseconds after the flash. The results here show how interpolation of the past is the only framework of the three models that provides a unified explanation for the flash-lag phenomenon. PMID- 10720335 TI - Safe health care: are we up to it? PMID- 10720336 TI - Medical error: the second victim. The doctor who makes the mistake needs help too. PMID- 10720337 TI - Accreditation's role in reducing medical errors. PMID- 10720338 TI - Why error reporting systems should be voluntary. PMID- 10720339 TI - Let's talk about error. PMID- 10720340 TI - MPs call for safeguards for people with personality disorder. PMID- 10720341 TI - Force feeding of Ian Brady declared lawful. PMID- 10720342 TI - Type 2 diabetes is a major drain on resources. PMID- 10720343 TI - Imitating mickey mouse can be dangerous PMID- 10720344 TI - In brief PMID- 10720346 TI - Another option for Down's syndrome screening available PMID- 10720345 TI - New standards set for abortion care. PMID- 10720347 TI - Charity calls for help for people of Aral sea area. PMID- 10720348 TI - Doctors develop new treatment for cystic fibrosis PMID- 10720349 TI - UK juniors to debate pay offer. PMID- 10720351 TI - Adding fatty acids to baby milk improves development PMID- 10720350 TI - Rate of HIV transmission among Africans in UK "underestimated". PMID- 10720352 TI - US drug companies announce vaccine initiative. PMID- 10720353 TI - Transplanted pancreatic stem cells can reverse diabetes in mice PMID- 10720354 TI - Reducing errors made by emergency physicians in interpreting radiographs: longitudinal study. AB - OBJECTIVES: To reduce errors made in the interpretation of radiographs in an emergency department. DESIGN: Longitudinal study. SETTING: Hospital emergency department. INTERVENTIONS: All staff reviewed all clinically significant discrepancies at monthly meetings. A file of clinically significant errors was created; the file was used for teaching. Later a team redesigned the process. A system was developed for interpreting radiographs that would be followed regardless of the day of the week or time of day. All standard radiographs were brought directly to the emergency physician for immediate interpretation. Radiologists reviewed the films within 12 hours as a quality control measure, and if a significant misinterpretation was found patients were asked to return. MAIN OUTCOME MEASURES: Reduction in number of clinically significant errors (such as missed fractures or foreign bodies) on radiographs read in the emergency department. Data on the error rate for radiologists and the effect of the recall procedure were not available so reliability modelling was used to assess the effect of these on overall safety. RESULTS: After the initial improvements the rate of false negative errors fell from 3% (95% confidence interval 2.8% to 3.2%) to 1.2% (1.03% to 1.37%). After the processes were redesigned it fell further to 0.3% (0.26% to 0.34%). Reliability modelling showed that the number of potential adverse effects per 1000 cases fell from 19 before the improvements to 3 afterwards and unmitigated adverse effects fell from 2.2/1000 before to 0.16/1000 afterwards, assuming 95% success in calling patients back. CONCLUSION: Systems of radiograph interpretation that optimise the skills of all clinicians involved and contain reliable processes for mitigating errors can reduce error rates substantially. PMID- 10720355 TI - Incidence and types of preventable adverse events in elderly patients: population based review of medical records. AB - OBJECTIVE: To determine the incidence and types of preventable adverse events in elderly patients. DESIGN: Review of random sample of medical records in two stage process by nurses and physicians to detect adverse events. Two study investigators then judged preventability. SETTING: Hospitals in US states of Utah and Colorado, excluding psychiatric and Veterans Administration hospitals. SUBJECTS: 15 000 hospitalised patients discharged in 1992. MAIN OUTCOME MEASURES: Incidence of preventable adverse events (number of preventable events per 100 discharges) in elderly patients (>/=65 years old) and non-elderly patients (16-64 years). RESULTS: When results were extrapolated to represent all discharges in 1992 in both states, non-elderly patients had 8901 adverse events (incidence 2.80% (SE 0.18%)) compared with 7419 (5.29% (0.37%)) among elderly patients (P=0.001). Non-elderly patients had 5038 preventable adverse events (incidence 1.58% (0.14%)) compared with 4134 (2.95% (0.28%)) in elderly patients (P=0.001). Elderly patients had a higher incidence of preventable events related to medical procedures (such as thoracentesis, cardiac catheterisation) (0.69% (0.14%) v 0.13% (0.04%)), preventable adverse drug events (0.63% (0.14%) v 0.17% (0.05%)), and preventable falls (0.10% (0.06%) v 0.01% (0.02%)). In multivariate analyses, adjusted for comorbid illnesses and case mix, age was not an independent predictor of preventable adverse events. CONCLUSIONS: Preventable adverse events were more common among elderly patients, probably because of the clinical complexity of their care rather than age based discrimination. Preventable adverse drug events, events related to medical procedures, and falls were especially common in elderly patients and should be targets for efforts to prevent errors. PMID- 10720356 TI - Error, stress, and teamwork in medicine and aviation: cross sectional surveys. AB - OBJECTIVES: To survey operating theatre and intensive care unit staff about attitudes concerning error, stress, and teamwork and to compare these attitudes with those of airline cockpit crew. DESIGN: : Cross sectional surveys. SETTING: : Urban teaching and non-teaching hospitals in the United States, Israel, Germany, Switzerland, and Italy. Major airlines around the world. PARTICIPANTS: : 1033 doctors, nurses, fellows, and residents working in operating theatres and intensive care units and over 30 000 cockpit crew members (captains, first officers, and second officers). MAIN OUTCOME MEASURES: : Perceptions of error, stress, and teamwork. RESULTS: : Pilots were least likely to deny the effects of fatigue on performance (26% v 70% of consultant surgeons and 47% of consultant anaesthetists). Most pilots (97%) and intensive care staff (94%) rejected steep hierarchies (in which senior team members are not open to input from junior members), but only 55% of consultant surgeons rejected such hierarchies. High levels of teamwork with consultant surgeons were reported by 73% of surgical residents, 64% of consultant surgeons, 39% of anaesthesia consultants, 28% of surgical nurses, 25% of anaesthetic nurses, and 10% of anaesthetic residents. Only a third of staff reported that errors are handled appropriately at their hospital. A third of intensive care staff did not acknowledge that they make errors. Over half of intensive care staff reported that they find it difficult to discuss mistakes. CONCLUSIONS: Medical staff reported that error is important but difficult to discuss and not handled well in their hospital. Barriers to discussing error are more important since medical staff seem to deny the effect of stress and fatigue on performance. Further problems include differing perceptions of teamwork among team members and reluctance of senior theatre staff to accept input from junior members. PMID- 10720358 TI - The war in south africa: "Comforts" and luxuries PMID- 10720359 TI - When I use a word. nice? PMID- 10720357 TI - Implementation of rules based computerised bedside prescribing and administration: intervention study. AB - OBJECTIVES: To implement and assess a rules based computerised prescribing system with the aim of improving the safety of prescriptions and the administration of drugs. DESIGN: Analysis of performance of computerised system plus questionnaire survey of users. SETTING: 64 bed renal unit in a teaching hospital. INTERVENTION: : Introduction of the system into routine clinical use. MAIN OUTCOME MEASURES: Number of attempted prescriptions cancelled by the system; proportion of warning messages overridden; users' comparisons of the system with conventional procedures. RESULTS: Between October 1998 and August 1999 the system cancelled 58 (0.07%) out of 87 789 prescriptions on the grounds of clinical safety. In addition, 427 (57%) attempted prescriptions generating high level warnings and 1257 (8%) generating low level warnings were not completed. In a user survey 82% (31/38) of doctors and nurses considered the system to be an improvement on conventional procedures. CONCLUSIONS: The system has contributed to safety and patient care. All prescriptions are complete and legible, and transcription errors have been eliminated. The system assists clinicians when they are writing a prescription by making available information on patients. The system supports clinical decision making and has been well received by doctors, nurses, and pharmacists. PMID- 10720360 TI - Effects of preventive home visits to elderly people living in the community: systematic review. AB - OBJECTIVE: To assess the effects of preventive home visits to elderly people living in the community. DESIGN: Systematic review. SETTING: 15 trials retrieved from Medline, Embase, and the Cochrane controlled trial register. MAIN OUTCOME MEASURES: Physical function, psychosocial function, falls, admissions to institutions, and mortality. RESULTS: Considerable differences in the methodological quality of the 15 trials were found, but in general the quality was considered adequate. Favourable effects of the home visits were observed in 5 out of 12 trials measuring physical functioning, 1 out of 8 measuring psychosocial function, 2 out of 6 measuring falls, 2 out of 7 measuring admissions to institutions, and 3 of 13 measuring mortality. None of the trials reported negative effects. CONCLUSIONS: No clear evidence was found in favour of the effectiveness of preventive home visits to elderly people living in the community. It seems essential that the effectiveness of such visits is improved, but if this cannot be achieved consideration should be given to discontinuing these visits. PMID- 10720361 TI - Reporting and preventing medical mishaps: lessons from non-medical near miss reporting systems. PMID- 10720362 TI - ABC of arterial and venous disease: Acute limb ischaemia. PMID- 10720363 TI - Human error: models and management. PMID- 10720364 TI - System changes to improve patient safety. PMID- 10720365 TI - Epidemiology of medical error. PMID- 10720366 TI - How to investigate and analyse clinical incidents: clinical risk unit and association of litigation and risk management protocol. PMID- 10720367 TI - On error management: lessons from aviation. PMID- 10720368 TI - Anaesthesiology as a model for patient safety in health care. PMID- 10720369 TI - Using information technology to reduce rates of medication errors in hospitals. PMID- 10720370 TI - Gaps in the continuity of care and progress on patient safety. PMID- 10720372 TI - Realising why PMID- 10720371 TI - Detecting and reporting medical errors: why the dilemma? PMID- 10720373 TI - A cerebral massage? PMID- 10720374 TI - PFI rides again. Scepticism remains at the grassroots. PMID- 10720375 TI - Choosing between home and hospital delivery. Home birth in Britain can be safe. PMID- 10720376 TI - Helicobacter pylori and myocardial infection. Excluding group with potentially higher rates of infection with H pylori could bias estimated odds ratio. PMID- 10720377 TI - Treatment of schizophrenia. Value of diagnosis of schizophrenia remains in dispute. PMID- 10720378 TI - Preventing pressure sores. Good nursing care should prevent pressure sores. PMID- 10720379 TI - World Trade Organisation agreements should be subject to health impact assessment. PMID- 10720380 TI - Britain is ahead of US in dealing with misconduct. PMID- 10720381 TI - Chinese hypnosis can cause qigong induced mental disorders. PMID- 10720382 TI - Dental recalls are useful for detecting oral cancer. PMID- 10720383 TI - Carbon monoxide poisoning. Carboxyhaemoglobin can be measured with standard blood tests. PMID- 10720384 TI - More reluctance in accepting evidence on smoking and cancer. PMID- 10720385 TI - Student publications. Students in Birmingham have published projects for more than 10 years. PMID- 10720386 TI - Meningitis C immunisation is low among young people who are not in education. PMID- 10720387 TI - Obituaries PMID- 10720388 TI - Health secretary wants to expand NHS services PMID- 10720389 TI - Blind eye: how the medical establishment let a doctor get away with murder PMID- 10720391 TI - The arts in healthcare: learning from experience PMID- 10720390 TI - Growing up in britain: ensuring a healthy future for our children PMID- 10720392 TI - Epilepsy: A comprehensive textbook on CD-ROM PMID- 10720393 TI - Sultans of spin PMID- 10720394 TI - Error in medicine PMID- 10720395 TI - Looking back PMID- 10720396 TI - Netlines PMID- 10720398 TI - Crisis in the air PMID- 10720397 TI - Plane speaking PMID- 10720400 TI - Lower error rates in reading radiographs can be sustained by redesigning the system PMID- 10720399 TI - Facing up to medical error PMID- 10720401 TI - Older people suffer more adverse events but they have more complex care PMID- 10720402 TI - Healthcare staff deny effects of stress and tiredness PMID- 10720403 TI - Portable computerised prescribing may reduce errors PMID- 10720404 TI - Preventive home visits need improving PMID- 10720405 TI - A system approach to reducing error works better than focusing on individuals PMID- 10720406 TI - Metabolic mechanisms associated with antianginal therapy. PMID- 10720407 TI - Are Kv channels the essence of O2 sensing? PMID- 10720408 TI - LQT2 : amplitude reduction and loss of selectivity in the tail that wags the HERG channel. PMID- 10720409 TI - NAD(P)H oxidase: role in cardiovascular biology and disease. AB - Reactive oxygen species have emerged as important molecules in cardiovascular function. Recent work has shown that NAD(P)H oxidases are major sources of superoxide in vascular cells and myocytes. The biochemical characterization, activation paradigms, structure, and function of this enzyme are now partly understood. Vascular NAD(P)H oxidases share some, but not all, characteristics of the neutrophil enzyme. In response to growth factors and cytokines, they produce superoxide, which is metabolized to hydrogen peroxide, and both of these reactive oxygen species serve as second messengers to activate multiple intracellular signaling pathways. The vascular NAD(P)H oxidases have been found to be essential in the physiological response of vascular cells, including growth, migration, and modification of the extracellular matrix. They have also been linked to hypertension and to pathological states associated with uncontrolled growth and inflammation, such as atherosclerosis. PMID- 10720410 TI - beta-adrenergic receptor signaling: an acute compensatory adjustment inappropriate for the chronic stress of heart failure? Insights from Gsalpha overexpression and other genetically engineered animal models. PMID- 10720411 TI - Novel gain-of-function mechanism in K(+) channel-related long-QT syndrome: altered gating and selectivity in the HERG1 N629D mutant. AB - The N629D mutation, adjacent to the GFG signature sequence of the HERG1 A K(+) channel, causes long-QT syndrome (LQTS). Expression of N629D in Xenopus oocytes produces a rapidly activating, noninactivating current. N629D is nonselective among monovalent cations; permeation of K(+) was similar to that of Na(+) or Cs(+). During repolarization to potentials between -30 and -70 mV, N629D manifested an inward tail current, which was abolished by replacement of extracellular Na(+) (Na(+)(e)) with extracellular N-methyl-D-glucamine (NMG(e)). Because LQTS occurs in heterozygous patients, we coexpressed N629D and wild type (WT) at equimolar concentrations. Heteromultimer formation was demonstrated by analyzing the response to 0 [K(+)](e). The outward time-dependent current was nearly eliminated for WT at 0 [K(+)](e), whereas no reduction was observed for homomultimeric N629D or for the equimolar coexpressed current. To assess physiological significance, dofetilide-sensitive currents were recorded during application of simulated action potential clamps. During phase 3 repolarization, WT manifested outward currents, whereas homomultimeric N629D manifested inward depolarizing currents. During coexpression studies, variable phenotypes were observed ranging from a reduction in outward repolarizing current to net inward depolarizing current during phase 3. In summary, N629D replaces the WT outward repolarizing tail current with an inward depolarizing sodium current, which is expected to delay later stages of repolarization and contribute to arrhythmogenesis. Thus, the consequences of N629D resemble the pathophysiology seen in LQT3 Na(+) channel mutations and may be considered the first LQTS K(+) channel mutation that exhibits gain of function. PMID- 10720412 TI - NADH oxidase activation is involved in arsenite-induced oxidative DNA damage in human vascular smooth muscle cells. AB - Arsenic is atherogenic, carcinogenic, and genotoxic. Because atherosclerotic plaque has been considered a benign smooth muscle cell tumor, we have studied the effects of arsenite on DNA integrity of human vascular smooth muscle cells. By using single-cell alkaline electrophoresis, apparent DNA strand breaks were detected in a 4-hour treatment with arsenite at a concentration above 1 micromol/L. DNA strand breaks of arsenite-treated cells were increased by Escherichia coli formamidopyrimidine-DNA glycosylase and decreased by diphenylene iodinium, superoxide dismutase, catalase, pyruvate, DMSO, or D-mannitol. Extract from arsenite-treated cells showed increased capacity for producing superoxide when NADH was included in the reaction mixture; however, addition of arsenite to extract from untreated cells did not increase superoxide production. The superoxide-producing ability of arsenite-treated cells was also suppressed by diphenylene iodinium, 4,5-dihydroxy-1, 2-benzenedisulfonic acid disodium salt (Tiron), or superoxide dismutase. Superoxide production and DNA strand breaks in arsenite-treated cells were also suppressed by transfecting antisense oligonucleotides of p22phox, an essential component of NADH oxidase. Treatment with arsenite also increased the mRNA level of p22phox. These results suggest that arsenite activates NADH oxidase to produce superoxide, which then causes oxidative DNA damage. The result that arsenite at low concentrations increases oxidant levels and causes oxidative DNA damage in vascular smooth muscle cells may be important in arsenic-induced atherosclerosis. PMID- 10720413 TI - Loss of expression of the beta subunit of soluble guanylyl cyclase prevents nitric oxide-mediated inhibition of DNA synthesis in smooth muscle cells of old rats. AB - We compared the effects of NO donors and cGMP analogues on the growth of aortic smooth muscle cells (SMCs) derived from newborn, adult (aged 3 months), and old (aged 2 years) rats. We found that the NO donor S-nitroso-N-acetylpenicillamine failed to block DNA synthesis in SMCs from old rats but was effective in SMCs from newborn and adult rats. However, cGMP analogues were inhibitory in all 3 SMC types. We demonstrated that in SMCs from old rats, NO was unable to increase the concentration of intracellular cGMP, suggesting that either cGMP synthesis was defective or cGMP degradation was enhanced. Western blot analysis revealed that SMCs from old rats do not express the beta subunit of soluble guanylyl cyclase. To confirm the importance of this observation in vivo, we balloon-injured the carotid arteries of adult and old rats. Whereas soluble guanylyl cyclase was expressed at the same level in the media of injured vessels and uninjured vessels of both groups, its expression in the intimas of old rats was reduced by 70% compared with intimas from adult animals. Furthermore, N(omega)-nitro-L-arginine, an inhibitor of NO synthesis, enhanced the intimal thickening in injured vessels in adult rats but not in old rats. We conclude that the loss of NO responsiveness in aged rats is due to the lack of the beta subunit of soluble guanylyl cyclase, and we speculate that this defect contributes to the enhanced intimal thickening in response to injury in old animals. PMID- 10720414 TI - Direct observations in vivo on the role of endothelial selectins and alpha(4) integrin in cytokine-induced leukocyte-endothelium interactions in the mouse aorta. AB - The molecular mechanisms underlying leukocyte recruitment in large arteries have been extensively studied using histological techniques on fixed tissues. However, there are no reports that address the dynamics of leukocyte recruitment in large arteries in vivo. We developed an intravital microscopy technique for direct observation of leukocyte-endothelium interactions in the mouse aorta. Circulating leukocytes were labeled intravasally with rhodamine 6G and microscopically visualized within the aorta, allowing direct analysis of leukocyte rolling and adhesion. In untreated vessels, leukocyte-endothelium interactions were virtually absent. However, local pretreatment with cytokines interleukin-1beta and tumor necrosis factor-alpha induced clear-cut leukocyte rolling and adhesion, compatible with normal blood flow and wall shear rate. High shear decreased rolling leukocyte flux and increased leukocyte rolling velocity, thus decreasing the tendency for firm adhesion. Leukocyte rolling was almost abolished by an antibody blocking the function of P-selectin, whereas function-blocking antibodies against E-selectin and the alpha(4)-integrin subunit decreased rolling leukocyte flux to 51+/-34% (mean +/- SD) and 59+/-11% of the value before antibody treatment, respectively. In addition, inhibition of E-selectin function, but not of alpha(4) integrin, resulted in increased leukocyte rolling velocity (from 48+/-32 to 71+/-32 microm per second). Taken together, we introduce the first model for direct studies of leukocyte-endothelium interactions in a large artery in vivo and demonstrate cytokine-induced shear-sensitive leukocyte rolling that is critically dependent on P-selectin and modulated by E-selectin and alpha(4) integrin. PMID- 10720415 TI - Potential role for kv3.1b channels as oxygen sensors. AB - Hypoxia inhibits voltage-gated K channels in pulmonary artery smooth muscle (PASM). This is thought to contribute to hypoxic pulmonary vasoconstriction by promoting membrane depolarization, Ca(2+) influx, and contraction. Several of the K-channel subtypes identified in pulmonary artery have been implicated in the response to hypoxia, but contradictory evidence clouds the identity of the oxygen sensing channels. Using patch-clamp techniques, this study investigated the effect of hypoxia on recombinant Kv1 channels previously identified in pulmonary artery (Kv1.1, Kv1.2, and Kv1.5) and Kv3.1b, which has similar kinetic and pharmacological properties to native oxygen-sensitive currents. Hypoxia failed to inhibit any Kv1 channel, but it inhibited Kv3.1b channels expressed in L929 cells, as shown by a reduction of whole-cell current and single-channel activity, without affecting unitary conductance. Inhibition was retained in excised membrane patches, suggesting a membrane-delimited mechanism. Using reverse transcription-polymerase chain reaction and immunocytochemistry, Kv3.1b expression was demonstrated in PASM cells. Moreover, hypoxia inhibited a K(+) current in rabbit PASM cells in the presence of charybdotoxin and capsaicin, which preserve Kv3.1b while blocking most other Kv channels, but not in the presence of millimolar tetraethylammonium ions, which abolish Kv3.1b current. Kv3.1b channels may therefore contribute to oxygen sensing in pulmonary artery. PMID- 10720416 TI - Preconditioning in cardiomyocytes protects by attenuating oxidant stress at reperfusion. AB - Cardiomyocyte death after ischemia/reperfusion correlates with oxidant stress, and antioxidants confer protection in that model. Preconditioning (PC) with hypoxia or adenosine also confers protection, leading us to hypothesize that PC protects by attenuating oxidant generation during subsequent ischemia/reperfusion. Chick cardiomyocytes were preconditioned with 10 minutes of hypoxia or adenosine (100 micromol/L), followed by 1 hour of simulated ischemia and 3 hours of reperfusion. Adenosine PC decreased cell death from 50+/-3% to 18+/-4% and enhanced the return of contractions during reperfusion, as observed previously with hypoxic PC. A transient burst of dichlorofluorescein (sensitive to H2O2 oxidation that was significantly attenuated by PC initiated by hypoxia or adenosine was seen at reperfusion. The protein kinase C (PKC) inhibitor Go-6976 and the mitochondrial ATP-sensitive K(+) (K(ATP)) channel inhibitor 5 hydroxydecanoate each abolished protection and abrogated the PC-induced attenuation of reperfusion oxidant stress. By contrast, when given only at reperfusion, the K(+) channel opener pinacidil or the antioxidants 2 mercaptopropionylglycine and 1,10-phenanthroline decreased oxidant stress at reperfusion and improved survival and return of contractions. Thus, PC protection is associated with an attenuation of the oxidant burst at reperfusion, regardless of the method by which PC is triggered. Loss of PC protection associated with PKC inhibition or K(ATP) channel inhibitors is associated with a restoration of that oxidant stress. These results suggest a mechanism for PC protection and reveal a functional link between PKC activation and K(ATP) channel activation in that pathway. PMID- 10720417 TI - Redox changes of cultured endothelial cells and actin dynamics. AB - We studied the association between the production of reactive oxygen species, actin organization, and cellular motility. We have used an endothelial cell monolayer-wounding assay to demonstrate that the cells at the margin of the wound thus created produced significantly more free radicals than did cells in distant rows. The rate of incorporation of actin monomers into filaments was fastest at the wound margin, where heightened production of free radicals was detected. We have tested the effect of decreasing reactive oxygen species production on the migration of endothelial cells and on actin polymerization. The NADPH inhibitor diphenylene iodonium and the superoxide dismutase mimetic manganese (III) tetrakis(1-methyl-4-pyridyl)porphyrin (MnTMPyP) virtually abolished cytochalasin D-inhibitable actin monomer incorporation at the fast-growing barbed ends of filaments. Moreover, endothelial cell migration within the wound was significantly retarded in the presence of both diphenylene iodonium and MnTMPyP. We conclude that migration of endothelial cells in response to loss of confluence includes the intracellular production of reactive oxygen species, which contribute to the actin cytoskeleton reorganization required for the migratory behavior of endothelial cells. PMID- 10720418 TI - Changes in Ca(2+) cycling proteins underlie cardiac action potential prolongation in a pressure-overloaded guinea pig model with cardiac hypertrophy and failure. AB - Ventricular arrhythmias are common in both cardiac hypertrophy and failure; cardiac failure in particular is associated with a significant increase in the risk of sudden cardiac death. We studied the electrophysiologic changes in a guinea pig model with aortic banding resulting in cardiac hypertrophy at 4 weeks and progressing to cardiac failure at 8 weeks using whole-cell patch-clamp and biochemical techniques. Action potential durations (APDs) were significantly prolonged in banded animals at 4 and 8 weeks compared with age-matched sham operated animals. APDs at 50% and 90% repolarization (APD(50) and APD(90) in ms) were the following: 4 week, banded, 208+/-51 and 248+/-49 (n = 15); 4 week, sham, 189+/-68 and 213+/-69 (n = 16); 8 week, banded, 197+/-40 and 226+/-40 (n = 21); and 8 week, sham, 156+/-42 and 189+/-45 (n = 22), respectively; P<0.05 comparing banded versus sham-operated animals. We observed no significant differences in the K(+) currents between the 2 groups of animals at 4 and 8 weeks. However, banded animals exhibited a significant increase in Na(+) and Na(+)-Ca(2+) exchange current densities compared with controls. Furthermore, we have found a significant attenuation in the Ca(2+)-dependent inactivation of the L-type Ca(2+) current in the banded compared with sham-operated animals, likely as a result of the significant downregulation of the sarcoplasmic reticulum Ca(2+) ATPase, which has been documented previously in the heart failure animals. Our data provide an alternate mechanism for APD prolongation in cardiac hypertrophy and failure and support the notion that there is close interaction between Ca(2+) handling and action potential profile. PMID- 10720419 TI - Atrial but not ventricular fibrosis in mice expressing a mutant transforming growth factor-beta(1) transgene in the heart. AB - Increased transforming growth factor (TGF)-beta(1) activity has been observed during pathologic cardiac remodeling in a variety of animal models. In an effort to establish a causal role of TGF-beta(1) in this process, transgenic mice with elevated levels of active myocardial TGF-beta(1) were generated. The cardiac restricted alpha-myosin heavy chain promoter was used to target expression of a mutant TGF-beta(1) cDNA harboring a cysteine-to-serine substitution at amino acid residue 33. This alteration blocks covalent tethering of the TGF-beta(1) latent complex to the extracellular matrix, thereby rendering a large proportion (>60%) of the transgene-encoded TGF-beta(1) constitutively active. Although similar levels of active TGF-beta(1) were present in the transgenic atria and ventricles, overt fibrosis was observed only in the atria. Surprisingly, increased active TGF beta(1) levels inhibited ventricular fibroblast DNA synthesis in uninjured hearts and delayed wound healing after myocardial injury. These data suggest that increased TGF-beta(1) activity by itself is insufficient to promote ventricular fibrosis in the adult mouse ventricle. PMID- 10720420 TI - The antianginal drug trimetazidine shifts cardiac energy metabolism from fatty acid oxidation to glucose oxidation by inhibiting mitochondrial long-chain 3 ketoacyl coenzyme A thiolase. AB - Trimetazidine is a clinically effective antianginal agent that has no negative inotropic or vasodilator properties. Although it is thought to have direct cytoprotective actions on the myocardium, the mechanism(s) by which this occurs is as yet undefined. In this study, we determined what effects trimetazidine has on both fatty acid and glucose metabolism in isolated working rat hearts and on the activities of various enzymes involved in fatty acid oxidation. Hearts were perfused with Krebs-Henseleit solution containing 100 microU/mL insulin, 3% albumin, 5 mmol/L glucose, and fatty acids of different chain lengths. Both glucose and fatty acids were appropriately radiolabeled with either (3)H or (14)C for measurement of glycolysis, glucose oxidation, and fatty acid oxidation. Trimetazidine had no effect on myocardial oxygen consumption or cardiac work under any aerobic perfusion condition used. In hearts perfused with 5 mmol/L glucose and 0.4 mmol/L palmitate, trimetazidine decreased the rate of palmitate oxidation from 488+/-24 to 408+/-15 nmol x g dry weight(-1) x minute(-1) (P<0.05), whereas it increased rates of glucose oxidation from 1889+/-119 to 2378+/-166 nmol x g dry weight(-1) x minute(-1) (P<0.05). In hearts subjected to low-flow ischemia, trimetazidine resulted in a 210% increase in glucose oxidation rates. In both aerobic and ischemic hearts, glycolytic rates were unaltered by trimetazidine. The effects of trimetazidine on glucose oxidation were accompanied by a 37% increase in the active form of pyruvate dehydrogenase, the rate-limiting enzyme for glucose oxidation. No effect of trimetazidine was observed on glycolysis, glucose oxidation, fatty acid oxidation, or active pyruvate dehydrogenase when palmitate was substituted with 0.8 mmol/L octanoate or 1.6 mmol/L butyrate, suggesting that trimetazidine directly inhibits long-chain fatty acid oxidation. This reduction in fatty acid oxidation was accompanied by a significant decrease in the activity of the long-chain isoform of the last enzyme involved in fatty acid beta-oxidation, 3-ketoacyl coenzyme A (CoA) thiolase activity (IC(50) of 75 nmol/L). In contrast, concentrations of trimetazidine in excess of 10 and 100 micromol/L were needed to inhibit the medium- and short chain forms of 3-ketoacyl CoA thiolase, respectively. Previous studies have shown that inhibition of fatty acid oxidation and stimulation of glucose oxidation can protect the ischemic heart. Therefore, our data suggest that the antianginal effects of trimetazidine may occur because of an inhibition of long-chain 3 ketoacyl CoA thiolase activity, which results in a reduction in fatty acid oxidation and a stimulation of glucose oxidation. PMID- 10720421 TI - Role of kinins in the control of renal papillary blood flow, pressure natriuresis, and arterial pressure. AB - The present study evaluated the effects of blocking kinins with the bradykinin B(2) receptor antagonist Hoe140 on the relationship between renal perfusion pressure, papillary blood flow (PBF), and sodium excretion. To determine the relevance of renal kinins in the long-term control of arterial pressure, the effect of a chronic intrarenal infusion of Hoe140 on arterial pressure and sodium balance was also studied. PBF was not autoregulated in volume-expanded rats, and the administration of Hoe140 reduced PBF (-30%) and improved PBF autoregulation. The kinin antagonist also decreased sodium excretion (-35%) and blunted pressure natriuresis with no whole-kidney renal hemodynamic changes. These effects may be mediated through nitric oxide (NO), because in rats pretreated with N(G)-nitro-L arginine methyl ester, Hoe140 had no additional effects on PBF or pressure natriuresis. A role for NO in mediating the renal response to Hoe140 is also supported by the finding that Hoe140 reduced basal urinary NO(3)(-)/NO(2)(-) excretion (-33%), and it blunted the arterial pressure-induced increase in NO(3)( )/NO(2)(-) excretion, which is compatible with the idea that the pressure natriuresis response may be mediated through kinins and NO. The importance of kinins in long-term regulation of arterial pressure is demonstrated by the severe arterial hypertension (172+/-6 mm Hg) induced during the chronic intrarenal infusion of Hoe140 associated with sodium and volume retention. These data suggest that renal kinins and NO may be a part of the renal mechanism coupling changes in arterial pressure with modifications in PBF and sodium excretion, therefore contributing to the long-term control of arterial pressure. PMID- 10720422 TI - Sarcoplasmic reticulum Ca(2+)/Calmodulin-dependent protein kinase is altered in heart failure. AB - Although Ca(2+)/calmodulin-dependent protein kinase-II (CaMK) is known to phosphorylate different Ca(2+) cycling proteins in the cardiac sarcoplasmic reticulum (SR) and regulate its function, the status of CaMK in heart failure has not been investigated previously. In this study, we examined the hypothesis that changes in the CaMK-mediated phosphorylation of the SR Ca(2+) cycling proteins are associated with heart failure. For this purpose, heart failure in rats was induced by occluding the coronary artery for 8 weeks, and animals with >30% infarct of the left ventricle wall plus septum mass were used. Noninfarcted left ventricle was used for biochemical assessment; sham-operated animals served as control. A significant depression in SR Ca(2+) uptake and release activities was associated with a decrease in SR CaMK phosphorylation of the SR proteins, ryanodine receptor (RyR), Ca(2+) pump ATPase (SR/endoplasmic reticulum Ca(2+) ATPase [SERCA2a]), and phospholamban (PLB) in the failing heart. The SR protein contents for RyR, SERCA2a, and PLB were decreased in the failing hearts. Although the SR Ca(2+)/calmodulin-dependent CaMK activity, CaMK content, and CaMK autophosphorylation were depressed, the SR phosphatase activity was enhanced in the failing heart. On the other hand, the cAMP-dependent protein kinase-mediated phosphorylation of RyR and PLB was not affected in the failing heart. On the basis of these results, we conclude that alterations in SR CaMK-mediated phosphorylation may be partly responsible for impaired SR function in heart failure. PMID- 10720423 TI - Arterial shear stress stimulates surface expression of the endothelial glycoprotein Ib complex. AB - Exposure to shear stress has been shown to alter the expression of a number of surface components of cultured endothelial cells (EC). However, relatively few studies have examined the status of human EC surface proteins after prolonged flow, more closely corresponding to the steady state in vivo. Since the promoter region of glycoprotein (Gp) Ib alpha contains several copies of a putative shear stress response element, 5'-GAGACC-3', we investigated the response of cultured human umbilical vein EC (HUVEC) GpIb alpha to shear stress over a 72 h time period. In response to 30 dynes/cm2 of shear stress, total cell content of GpIb alpha protein was markedly increased above static levels at 7 and 24 h, as determined immunohistochemically. Western blot analysis of whole cell lysates after 24, 48, and 72 h of shear treatment demonstrated a 2.4-, 4.1-, and 3.2-fold increase in total GpIb alpha protein, respectively. Cell surface protein expression of GpIb alpha increased 2.5-fold at 7 h, as measured by quantitative immunofluorescence, and remained at that level at 24 h. After 48 h of shear stress, cell surface GpIb alpha, GpIX, and GpV, analyzed by flow cytometric analysis, were further increased over the levels observed at 24 h. The increase in cell surface membrane expression of GPIb alpha at 24, 48, and 72 h was confirmed by immunoprecipitation of biotinylated surface proteins. No upregulation of GpIb alpha was noted after exposure to shear stress of 1-3 dynes/cm2. These observations imply that under steady-state arterial shear conditions endothelial expression of the GpIb complex is significantly greater than observed in static EC cultures, and raise the possibility of a more important role for this complex under flow, rather than static conditions. PMID- 10720425 TI - A message of gratitude to the editorial board, both old and New PMID- 10720424 TI - Direct effects of prolactin on corticosterone release by zona fasciculata reticularis cells from male rats. AB - The role of prolactin (PRL) in the male is not fully defined. The aim of this study was to investigate the function and mechanism of PRL on the production of corticosterone by zona fasciculata-reticularis (ZFR) cells in vitro. The ZFR cells were obtained from male rats under normal, hyperprolactinemic, or hypoprolactinemic situation. PRL stimulated the corticosterone release in a dose dependent pattern in the ZFR cells from normal male rats. The cellular adenosine 3'-5'-cyclic monophosphate (cAMP) concentration positively correlated with PRL concentration in the presence of forskolin or 3-isobutyl-1-methylxanthine (IBMX). PRL enhanced the stimulatory effects of cAMP mimetic reagents, i.e., forskolin, 8 bromo-adenosine 3',5'-cyclic monophosphate (8-Br-cAMP), and IBMX on the release of corticosterone. The adenylate cyclase inhibitor (SQ22536) inhibited the corticosterone release in spite of presence of PRL. Nifedipine (L-type calcium channel blocker) did not inhibit corticosterone release. The hyperprolactinemic condition was actualized by transplantation of donor rat anterior pituitary glands (APs) under kidney capsule. By comparison with the cerebral cortex (CX) grafted group, AP-graft resulted in an increased release of corticosterone, 3beta hydroxysteriod dehydrogenase (HSD) activity and cAMP production by ZFR cells. Acute hypoprolactinemic status was induced by bromocriptine for 2 days. The results showed the productions of corticosterone were lower in hypoprolactinemic group than in control group, which were persistent along with different ACTH concentrations. These results suggest that PRL increase the release of corticosterone by ZFR cells via cAMP cascades and 3beta-HSD activity. PMID- 10720426 TI - Impaired bone formation in transgenic mice resulting from altered integrin function in osteoblasts. AB - To determine the role of integrins in mature osteoblasts in vivo, we expressed in transgenic mice a dominant-negative integrin subunit (beta1-DN) consisting of the beta1 subunit cytoplasmic and transmembrane domains, driven by the osteoblast specific osteocalcin promoter. Immature osteoblasts isolated from transgenic animals differentiated normally in vitro until the osteocalcin promoter became active; thereafter they detached from the substratum, suggesting that beta1-DN was impairing adhesion in mature osteoblasts. Transgenic animals had reduced bone mass, with increased cortical porosity in long bones and thinner flat bones in the skull. At 35 days, the rate of bone formation was reduced in cortical bone, and the parietal bones were 45% thinner than in wild-type animals. Active osteoblasts were less polar and had larger areas of cytoplasm with intracellular stores of matrix molecules. Osteocyte lacunae appeared normal around the cell body but did not have normal canilicular structures. At 90 days, the parietal bone of transgenic males was of normal width, suggesting that the original defect in matrix deposition had been repaired or compensated for. In contrast, transgenic females still had decreased bone mass in the parietal bone at 90 days. The decreased bone mass in TG females was accompanied by increased staining for osteoclast activity, suggesting that there was a sex-specific defect in mature animals. PMID- 10720427 TI - islet reveals segmentation in the Amphioxus hindbrain homolog. AB - The vertebrate embryonic hindbrain is segmented into rhombomeres. Gene expression studies suggest that amphioxus, the closest invertebrate relative of vertebrates, has a hindbrain homolog. However, this region is not overtly segmented in amphioxus, raising the question of how hindbrain segmentation arose in chordate evolution. Vertebrate hindbrain segmentation includes the patterning of cranial motor neurons, which can be identified by their expression of the LIM-homeodomain transcription factor islet1. To learn if the amphioxus hindbrain homolog is cryptically segmented, we cloned an amphioxus gene closely related to islet1, which we named simply islet. We report that amphioxus islet expression includes a domain of segmentally arranged cells in the ventral hindbrain homolog. We hypothesize that these cells are developing motor neurons and reveal a form of hindbrain segmentation in amphioxus. Hence, vertebrate rhombomeres may derive from a cryptically segmented brain present in the amphioxus/vertebrate ancestor. Other islet expression domains provide evidence for amphioxus homologs of the pineal gland, adenohypophysis, and endocrine pancreas. Surprisingly, homologs of vertebrate islet1-expressing spinal motor neurons and Rohon-Beard sensory neurons appear to be absent. PMID- 10720428 TI - The AML1 transcription factor functions to develop and maintain hematogenic precursor cells in the embryonic aorta-gonad-mesonephros region. AB - We examined the role of the AML1 transcription factor in the development of hematopoiesis in the paraaortic splanchnopleural (P-Sp) and the aorta-gonad mesonephros (AGM) regions of mouse embryos. The activity of colony-forming units of colonies from the P-Sp/AGM region was reduced severalfold by heterozygous disruption of the AML1 gene, indicating that AML1 functioned in a dosage dependent manner to generate hematopoietic progenitors. In addition, no hematopoietic progenitor activity was detected in the P-Sp/AGM region of embryos with an AML1 null mutation. Similar results were obtained when a dispersed culture was first prepared from the P-Sp/AGM region before assay of the activity of the colony-forming units. In a culture of cells with the AML1(+/+) genotype, both hematopoietic and endothelial-like cell types emerged, but in a culture of cells with the AML1(-/-) genotype, only endothelial-like cells emerged. Interestingly, introduction of AML1 cDNA into the P-Sp/AGM culture with the AML1( /-) genotype partially restored the production of hematopoietic cells. This restoration was observed for cultures prepared from 9.5-day postcoitum (dpc) embryos but not for cultures prepared from 11.5-dpc embryos. Therefore, the population of endothelial-like cells capable of growing in the AML1(-/-) culture would appear to contain inert but nonetheless competent hematogenic precursor cells up until at least the 9.5-dpc period. All these results support the notion that the AML1 transcription factor functions to develop and maintain hematogenic precursor cells in the embryonic P-Sp/AGM region. PMID- 10720429 TI - A novel Drosophila, mef2-regulated muscle gene isolated in a subtractive hybridization-based molecular screen using small amounts of zygotic mutant RNA. AB - It is unknown how the general patterning mechanisms that subdivide the mesoderm initiate different pathways of cell differentiation. One route to understanding these events is to isolate and analyse genes specifically expressed early in this differentiation process. I have therefore undertaken a novel molecular screen in Drosophila in a systematic search for such genes. The approach utilised subtractive hybridisation coupled to directional cDNA library construction. Libraries were made from as little as 20 microg total RNA isolated from hand picked embryos of defined stage of development and genotype. In a one-step procedure, the subtraction was 6.5- to 7.25-h wild-type embryos minus 6.5- to 7.25-h twist (twi) zygotic mutant embryos. A two-step procedure in which maternally expressed sequences were subtracted from each of these cDNA libraries, before subtracting twi from wild-type, increased the subtraction efficiency. It resulted in a cDNA population enriched more than 100-fold for mesodermal cDNAs. This was screened by determination of the embryonic expression pattern of each clone in a high throughput procedure and then DNA sequencing. The method, which is comprehensive and does not discriminate against rarer cDNAs, is generally applicable and calculations show that it would work for just 10 embryos. Analysis of one clone, Dmeso18E, that encodes a putative nuclear protein and fulfils the screen's aims is described. It is novel and its expression is mesoderm-specific, twi-dependent, and early during somatic, visceral, and heart muscle differentiation. Two pivotal regulators of mesoderm development and gene expression are Dmef2 and tinman (tin). Analysis of Dmeso18E expression revealed new aspects to their roles: there are effects of Dmef2 on developing muscle much earlier than hitherto believed, and there is tin-independent gene expression in, and invagination of, prospective midgut visceral muscle cells. Dmeso18E expression is regulated by Dmef2, although some expression is Dmef2-independent. The tin-independent and Dmef2-independent expression of Dmeso18E indicates that it either occupies a link between twi and genes like tin and Dmef2 or it lies in a parallel pathway of gene activation. PMID- 10720430 TI - Evidence that a cell-type-specific efflux pump regulates cell differentiation in Dictyostelium. AB - We have identified a cellular efflux pump, RhT, with the properties of an MDR transporter-a type of ATP-binding cassette transporter whose substrates include small hydrophobic molecules. RhT transports rhodamine 123 (Rh123) and is inhibited by low temperature, energy poisons, and several MDR transport inhibitors, such as verapamil. All vegetative cells have RhT activity, but during development prestalk cells lose RhT activity while prespore cells retain it. We also identified several RhT inhibitors. The most effective inhibitor is the stalk cell-inducing chlorinated alkyl phenone, DIF-1. The RhT inhibitors disrupted development, to varying degrees, and induced stalk cell formation in submerged culture. The inhibitors displayed the same rank order of pharmacological efficacy for stalk cell induction as they did for Rh123 transport inhibition. We also found that cerulenin, a specific inhibitor of DIF-1 biosynthesis (R. R. Kay, 1998, J. Biol. Chem. 273, 2669-2675), abolished the induction of stalk cells by each of the RhT inhibitors, and this effect could be reversed by DIF-1. Thus, DIF 1 synthesis appears to be required for the induction of stalk cells by the RhT inhibitors. Since DIF-1 is the most potent inhibitor of RhT activity, and thus a likely transport substrate itself, we propose that RhT inhibitors induce stalk cell differentiation by blocking DIF-1 export, causing DIF-1 to build up within cells. Our results provide evidence for a prespore-specific efflux pump that regulates cell fate determination, perhaps by regulating the cellular concentration of DIF-1. PMID- 10720431 TI - Signals from trunk paraxial mesoderm induce pronephros formation in chick intermediate mesoderm. AB - We used Pax-2 mRNA expression and Lim 1/2 antibody staining as markers for the conversion of chick intermediate mesoderm (IM) to pronephric tissue and Lmx-1 mRNA expression as a marker for mesonephros. Pronephric markers were strongly expressed caudal to the fifth somite by stage 9. To determine whether the pronephros was induced by adjacent tissues and, if so, to identify the inducing tissues and the timing of induction, we microsurgically dissected one side of chick embryos developing in culture and then incubated them for up to 3 days. The undisturbed contralateral side served as a control. Most embryos cut parallel to the rostrocaudal axis between the trunk paraxial mesoderm and IM before stage 8 developed a pronephros on the control side only. Embryos manipulated after stage 9 developed pronephric structures on both sides, but the caudal pronephric extension was attenuated on the cut side. These results suggest that a medial signal is required for pronephric development and show that the signal is propagated in a rostral to caudal sequence. In manipulated embryos cultured for 3 days in ovo, the mesonephros as well as the pronephros failed to develop on the experimental side. In contrast, embryos cut between the notochord and the trunk paraxial mesoderm formed pronephric structures on both sides, regardless of the stage at which the operation was performed, indicating that the signal arises from the paraxial mesoderm (PM) and not from axial mesoderm. This cut also served as a control for cuts between the PM and the IM and showed that signaling itself was blocked in the former experiments, not the migration of pronephric or mesonephric precursor cells from the primitive streak. Additional control experiments ruled out the need for signals from lateral plate mesoderm, ectoderm, or endoderm. To determine whether the trunk paraxial mesoderm caudal to the fifth somite maintains its inductive capacity in the absence of contact with more rostral tissue, embryos were transected. Those transected below the prospective level of the fifth somite expressed Pax-2 in both the rostral and the caudal isolates, whereas embryos transected rostral to this level expressed Pax-2 in the caudal isolate only. Thus, a rostral signal is not required to establish the normal pattern of Pax-2 expression and pronephros formation. To determine whether paraxial mesoderm is sufficient for pronephros induction, stage 7 or earlier chick lateral plate mesoderm was cocultured with caudal stage 8 or 9 quail somites in collagen gels. Pax-2 was expressed in chick tissues in 21 of 25 embryos. Isochronic transplantation of stage 4 or 5 quail node into caudal chick primitive streak resulted in the generation of ectopic somites. These somites induced ectopic pronephroi in lateral plate mesoderm, and the IM that received signals from both native and ectopic somites formed enlarged pronephroi with increased Pax-2 expression. We conclude that signals from a localized region of the trunk paraxial mesoderm are both required and sufficient for the induction of the pronephros from the chick IM. Studies to identify the molecular nature of the induction are in progress. PMID- 10720432 TI - Mice lacking Dad1, the defender against apoptotic death-1, express abnormal N linked glycoproteins and undergo increased embryonic apoptosis. AB - Dad1 has been shown to play a role in preventing apoptotic cell death and in regulating levels of N-linked glycosylation in Saccharomyces cerevisiae and the BHK hamster cell line. To address the in vivo role of Dad1 in these processes during multicellular development, we have analyzed mice carrying a null allele for Dad1. Embryos homozygous for this mutation express abnormal N-glycosylated proteins and are developmentally delayed by embryonic day 7.5. Such mutants exhibit aberrant morphology, impaired mesodermal development, and increased levels of apoptosis in specific tissues. These defects culminate in homozygous embryos failing to turn the posterior axis and subsequent lethality by embryonic day 10.5. Thus, Dad1 is required for proper processing of N-linked glycoproteins and for certain cell survival in the mouse. PMID- 10720433 TI - Norepinephrine transporter expression in cholinergic sympathetic neurons: differential regulation of membrane and vesicular transporters. AB - Sympathetic neurons that undergo a noradrenergic to cholinergic change in phenotype provide a useful model system to examine the developmental regulation of proteins required to synthesize, store, or remove a particular neurotransmitter. This type of change occurs in the sympathetic sweat gland innervation during development and can be induced in cultured sympathetic neurons by extracts of sweat gland-containing footpads or by leukemia inhibitory factor. Sympathetic neurons initially produce norepinephrine (NE) and contain the vesicular monoamine transporter 2 (VMAT2), which packages NE into vesicles, and the norepinephrine transporter (NET), which removes NE from the synaptic cleft to terminate signaling. We have used a variety of biochemical and molecular techniques to test whether VMAT2 and NET levels decrease in sympathetic neurons which stop producing NE and make acetylcholine. In cultured sympathetic neurons, NET protein and mRNA decreased during the switch to a cholinergic phenotype but VMAT2 mRNA and protein did not decline. NET immunoreactivity disappeared from the developing sweat gland innervation in vivo as it acquired cholinergic properties. Surprisingly, NET simultaneously appeared in sweat gland myoepithelial cells. The presence of NET in myoepithelial cells did not require sympathetic innervation. VMAT2 levels did not decrease as the sweat gland innervation became cholinergic, indicating that NE synthesis and vesicular packaging are not coupled in this system. Thus, production of NE and the transporters required for noradrenergic transmission are not coordinately regulated during cholinergic development. PMID- 10720434 TI - Translational control of cyclin B1 mRNA during meiotic maturation: coordinated repression and cytoplasmic polyadenylation. AB - Translational control is prominent during meiotic maturation and early development. In this report, we investigate a mode of translational repression in Xenopus laevis oocytes, focusing on the mRNA encoding cyclin B1. Translation of cyclin B1 mRNA is relatively inactive in the oocyte and increases dramatically during meiotic maturation. We show, by injection of synthetic mRNAs, that the cis acting sequences responsible for repression of cyclin B1 mRNA reside within its 3'UTR. Repression can be saturated by increasing the concentration of reporter mRNA injected, suggesting that the cyclin B1 3'UTR sequences provide a binding site for a trans-acting repressor. The sequences that direct repression overlap and include cytoplasmic polyadenylation elements (CPEs), sequences known to promote cytoplasmic polyadenylation. However, the presence of a CPE per se appears insufficient to cause repression, as other mRNAs that contain CPEs are not translationally repressed. We demonstrate that relief of repression and cytoplasmic polyadenylation are intimately linked. Repressing elements do not override the stimulatory effect of a long poly(A) tail, and polyadenylation of cyclin B1 mRNA is required for its translational recruitment. Our results suggest that translational recruitment of endogenous cyclin B1 mRNA is a collaborative effect of derepression and poly(A) addition. We discuss several molecular mechanisms that might underlie this collaboration. PMID- 10720435 TI - Developmental patterning genes and their conserved functions: from model organisms to humans. AB - Molecular and genetic evidence accumulated during the past 20 years in the field of developmental biology indicates that different animals possess many common genetic systems for embryonic patterning. In this review we describe the conserved functions of such developmental patterning genes and their relevance for human pathological conditions. Special attention is given to the Hox genetic system, involved in establishing cell identities along the anterior-posterior axis of all higher metazoans. We also describe other conserved genetic systems, such as the involvement of Pax6 genes in eye development and the role of Nkx2.5 type proteins in heart development. Finally, we outline some fascinating problems at the forefront of the studies of developmental patterning genes and show how knowledge obtained from model genetic organisms such as Drosophila helps to explain normal human morphogenesis and the genetic basis of some birth defects. PMID- 10720436 TI - Characterization of phenylketonuria missense substitutions, distant from the phenylalanine hydroxylase active site, illustrates a paradigm for mechanism and potential modulation of phenotype. AB - Missense mutations account for 48% of all reported human disease-causing alleles. Since few are predicted to ablate directly an enzyme's catalytic site or other functionally important amino acid residues, how do most missense mutations cause loss of function and lead to disease? The classic monogenic phenotype hyperphenylalaninemia (HPA), manifesting notably as phenylketonuria (PKU), where missense mutations in the PAH gene compose 60% of the alleles impairing phenylalanine hydroxylase (PAH) function, allows us to examine this question. Here we characterize four PKU-associated PAH mutations (F39L, K42I, L48S, I65T), each changing an amino acid distant from the enzyme active site. Using three complementary in vitro protein expression systems, and 3D-structural localization, we demonstrate a common mechanism. PAH protein folding is affected, causing altered oligomerization and accelerated proteolytic degradation, leading to reduced cellular levels of this cytosolic protein. Enzyme specific activity and kinetic properties are not adversely affected, implying that the only way these mutations reduce enzyme activity within cells in vivo is by producing structural changes which provoke the cell to destroy the aberrant protein. The F39L, L48S, and I65T PAH mutations were selected because each is associated with a spectrum of in vivo HPA among patients. Our in vitro data suggest that interindividual differences in cellular handling of the mutant, but active, PAH proteins will contribute to the observed variability of phenotypic severity. PKU thus supports a newly emerging paradigm both for mechanism whereby missense mutations cause genetic disease and for potential modulation of a disease phenotype. PMID- 10720437 TI - Amino acid homologies between human biotinidase and bacterial aliphatic amidases: putative identification of the active site of biotinidase. AB - A search of protein databases revealed amino acid homologies among human biotinidase, bacterial aliphatic amidases, and bacterial and plant nitrilases. Amino acids YRK(210-212) of biotinidase are conserved among the enzyme families. This homology and naturally occurring mutations that cause biotinidase deficiency suggest that this region is essential for enzyme activity and is conserved from bacteria. Cys(245) is likely the cysteine in the active site of biotinidase. PMID- 10720438 TI - Analysis of the expression of murine glutaryl-CoA dehydrogenase: in vitro and in vivo studies. AB - Glutaric acidemia type I (GAI) is an autosomal recessive organic acidemia caused by a mutation in the gene encoding glutaryl-CoA dehydrogenase (GCD). Clinically, GAI is characterized by progressive dystonia, resulting from degeneration of neurons in the caudate and putamen nuclei of the striatum. In an attempt to understand the basis for the specific neuropathology in GAI, we have analyzed the expression of the murine GCD gene using both in vitro and in vivo approaches. Transfection studies mapped the mouse GCD promoter to a 500-bp region of DNA 5' of the translation start site. The promoter lacks a TATA consensus sequence, but includes possible binding sites for several transcription factors with roles in the regulation of nuclear genes encoding mitochondrial proteins. Western blot and RT/PCR analyses of mouse tissues demonstrated that GCD is ubiquitously expressed, with the highest levels of expression in liver and kidney, consistent with its role in amino acid oxidation. Expression in multiple regions of the brain was also detected by Western blotting. Based on these results we conclude that the specific neuropathology associated with GCD deficiency in GAI cannot be accounted for by its expression pattern. PMID- 10720440 TI - BRCA1 suppresses insulin-like growth factor-I receptor promoter activity: potential interaction between BRCA1 and Sp1. AB - The insulin-like growth factor I receptor (IGF-I-R) has an important role in breast cancer etiology. The receptor is overexpressed by most breast cancers, where it functions as a potent antiapoptotic agent. BRCA1 is a tumor suppressor gene that is mutated in a large fraction of familial breast and ovarian cancers. Cotransfection of Saos-2, MCF7, and CHO cells with IGF-I-R promoter constructs driving luciferase reporter genes, and with a BRCA1 expression vector, suppressed promoter activity in a dose-dependent manner. Functional interactions between BRCA1 and Sp1 in the regulation of the IGF-I-R gene were studied in Schneider cells, a Drosophila cell line which lacks endogenous Sp1. In these cells BRCA1 suppressed 45% of the Sp1-induced trans-activation of the IGF-I-R promoter. These results suggest that BRCA1 is capable of suppressing the IGF-I-R promoter in a number of cell lines, thus resulting in low levels of receptor mRNA and protein. Mutant versions of BRCA1 lacking trans-activational activity can potentially derepress the IGF-I-R promoter. Activation of the overexpressed receptor by locally produced or circulating IGFs may be a crucial step in breast and ovarian cancer progression. PMID- 10720439 TI - Expression of palmitoyl protein thioesterase in neurons. AB - Infantile neuronal ceroid lipofuscinosis (INCL) is a severe neurodegenerative disorder in childhood that is caused by mutations in the gene encoding lysosomal palmitoyl protein thioesterase (PPT). INCL is characterized by massive and selective loss of cortical neurons. Here we have analyzed the intracellular processing and localization of adenovirus-mediated PPT in mouse primary neurons and NGF-induced PC-12 cells. The neuronal processing of PPT was found to be similar to that observed in peripheral cells, and a significant amount of the PPT enzyme was secreted in the primary neurons. Immunofluorescence analysis of the neuronal cells infected with wild-type PPT showed a granular staining pattern in the cell soma and neuronal shafts. Interestingly, PPT was also found in the synaptic ends of the neuronal cells and the staining pattern of the enzyme colocalized to a significant extent with the synaptic markers SV2 and synaptophysin. These in vitro data correspond with the distribution of endogeneous PPT in mouse brain and suggest that PPT may not solely be a lysosomal hydrolase. The specific targeting of PPT into the neuritic shafts and nerve terminals indicates that PPT may be associated with the maintenance of synaptic function. Furthermore, since a substantial amount of PPT is secreted by neurons, it is tempting to speculate that the enzyme could also have an extracellular substrate. PMID- 10720441 TI - Single effects of apolipoprotein B, (a), and E polymorphisms and interaction between plasminogen activator inhibitor-1 and apolipoprotein(a) genotypes and the risk of coronary artery disease in Czech male caucasians. AB - To evaluate whether polymorphisms in genes whose products are involved in lipid metabolism and fibrinolysis alter the risk of coronary artery disease (CAD), allele frequencies of four genetic polymorphisms were ascertained by PCR-based methods in 175 Czech male patients with coronary artery disease and in 222 Czech men with no symptoms of CAD. The following polymorphisms were studied: apolipoprotein B (apo B) signal peptide insertion/deletion polymorphism, 5' apolipoprotein(a) [apo(a)] TTTTA repeat polymorphism, apolipoprotein E (apo E) varepsilon2, varepsilon3, varepsilon4 polymorphism, and plasminogen activator inhibitor-1 (PAI-1) 4G/5G promoter polymorphism. Apo B and apo(a) allele frequencies differed significantly between the CAD and the control groups (P<0.01 each), with higher frequencies of apo B deletion and apo(a) shorter repeat alleles in the CAD group. We did not observe any differences in allele frequencies of either PAI-1 or apo E polymorphisms but the genotype frequencies of apo E were slightly different between the two groups (P<0.05). In addition, we observed a gene-gene interaction between the PAI-1 and apo(a) polymorphisms with respect to the risk of CAD. None of the polymorphisms studied were associated with the severity of CAD or a history of myocardial infarction. Our findings support the idea that several polymorphisms in apolipoprotein genes may by themselves and/or in interaction with other polymorphisms contribute to risk factors for CAD in men. PMID- 10720442 TI - Inositol phosphoglycans and signal transduction systems in pregnancy in preeclampsia and diabetes: evidence for a significant regulatory role in preeclampsia at placental and systemic levels. AB - Measurements have been made of the urinary content of inositol phosphoglycans IPG P-type and IPG A-type, putative insulin second messengers, in preeclampsia, in type I insulin-treated diabetic pregnant women and their matched control subjects, and nonpregnant women of child-bearing age. The content of IPG P-type and IPG A-type was also measured in the placenta from preeclamptic patients and from normal pregnancies. Pregnancy was associated with an increase, approximately twofold, in urinary output of IPG-P-type relative to nonpregnant controls (P<0.01). The 24-h output of IPG P-type in urine in preeclamptic women was significantly higher (2- to 3-fold) than in pregnant control subjects matched for age, parity, and stage of gestation (P<0.02). In contrast, insulin-dependent diabetic pregnant women did not show any significant change in urinary output of IPG P-type or IPG A-type relative to pregnant control subjects. Evidence for a possible relationship and correlation between the urinary excretion of IPG P-type and markers of preeclampsia, including proteinuria (r = 0.720, P<0.01), plasma aspartate transaminase (r = 0.658, P<0.05), and platelet counts (r = 0.613, P<0.05) is presented. A high yield of IPG P-type was extracted from human placenta, in preeclampsia some 3-fold higher (P = 0.03) than the normal value, whereas no IPG A-type (with lipogenic-stimulating activity) was found. Low concentrations of placental IPG A-type were detected relative to IPG P-type using assay systems dependent upon the effect of this mediator on cAMP-dependent protein kinase or on a proliferation assay using thymidine incorporation into DNA of EGFR T17 fibroblasts. It is postulated that the high urinary excretion IPG P type in preeclampsia reflects high placental levels and relates to the accumulation of glycogen in the placenta. The paracrine effects of placental IPG P-type (stimulation off other endocrine glands and/or endothelial cells) could contribute to the pathogenesis of the maternal syndrome. A possible theoretical link between elevated placental IPG P-type and apoptosis is proposed. PMID- 10720443 TI - Detachment of the glycolytic enzymes, phosphofructokinase and aldolase, from cytoskeleton of melanoma cells, induced by local anesthetics. AB - Cancer cells are characterized by a high rate of glycolysis, which is their primary energy source. An important mechanism that controls glycolysis is the reversible binding of glycolytic enzymes to cytoskeleton. We report here that the local anesthetics, lidocaine and bupivacaine, induced a dose-dependent detachment of the glycolytic enzymes, phosphofructokinase (EC 2.7.1.11) and aldolase (EC 4.1.2.13), from cytoskeleton of B16 melanoma cells. The detachment of glycolytic enzymes from cytoskeleton would reduce the provision of local ATP, in the vicinity of cytoskeleton-membrane and would also affect cytoskeleton structure. We show here that the local anesthetics decreased the viability of melanoma cells. The detachment of the glycolytic enzymes from cytoskeleton, induced by the drugs, preceded melanoma cell death, which indicates that this is an early effect and not a result of cell death. Bupivacaine was more potent than lidocaine both on the glycolytic enzymes and on cell viability. The present results suggest that local anesthetics, and especially bupivacaine, are promising drugs for the treatment of melanoma. PMID- 10720444 TI - Simultaneous mutations (A/G(-418) and C/T(-384)) in the apo(a) promoter of individuals with low Lp(a) levels. AB - High plasma levels of Lp(a), a low-density lipoprotein particle with an attached apo(a), are associated with the development of atherosclerosis. We present two simultaneous mutations in the apo(a) promoter (A/G(-418) and C/T(-384)) in healthy Indians with a low Lp(a) level (<5 mg/dl). No such mutations were detected in coronary artery disease positive individuals with very high Lp(a) levels (>200 mg/dl). The mutations described here might be useful in understanding the transcriptional regulation of the apo(a) gene. PMID- 10720445 TI - The arrhythmia of change: palpitations for academic medical centers. PMID- 10720446 TI - Transportation or noise is associated with tolerance to myocardial ischemia and reperfusion injury. AB - Myocardial stress can result in myocellular phenotypic changes including enhanced activity of antioxidant enzyme systems. Accordingly, endogenous tissue antioxidant enzyme activity has been associated with resistance to cardiac ischemia and reperfusion injury. The present study was designed to determine if environmental perturbations could alter myocardial antioxidant enzyme (catalase) activity and function after ischemia. Isolated perfused rat hearts (Langendorff apparatus, 37 degrees C) were subjected to 20 min global ischemia (37 degrees C) and 40 min reperfusion. Rats studied immediately following shipment had increased myocardial catalase activity (1330 +/- 3.5 U/g, P < 0.05 vs quarantined control) and increased resistance to ischemia and reperfusion injury (end reperfusion developed pressure, DP 55 +/- 4.0 mm Hg, P < 0.05 vs quarantined control). However, control rats that were quarantined for 4 weeks exhibited a progressive decrease in catalase activity (760 +/- 10 U/g) for 3 weeks of quarantine. There was a concurrent decrease in resistance to myocardial ischemia and reperfusion injury (DP 40 +/- 3.6 mm Hg). Similarly, quarantined rats subjected to construction-related noise levels in excess of 90 dB (A scale) had increased myocardial catalase activity (1140 +/- 3.3 U/g, P < 0.05) and functional tolerance to ischemia and reperfusion (DP 66 +/- 3.3 mm Hg, P < 0.05). Finally, rats experiencing 90-dB noise levels for 2 days exhibited increased myocardial catalase activity (1125 +/- 30 U/g, P < 0.05) and myocardial ischemia and reperfusion injury tolerance (DP 62 +/- 1.7 mm Hg, P < 0.05). We conclude that variations in environmental conditions can relate to changes in antioxidant defense mechanisms and tolerance to myocardial ischemia and reperfusion injury in the rat. PMID- 10720447 TI - Sustained nitric oxide production via l-arginine administration ameliorates effects of intestinal ischemia-reperfusion. AB - BACKGROUND: The role of nitric oxide in intestinal ischemia-reperfusion is unclear-some studies link it to the harmful effects of ischemia-reperfusion, while others report it to be protective. We propose that nitric oxide levels diminish in the reperfusion period in conjunction with the onset of increased capillary permeability. The aim of this study is to determine the effect of supplementing nitric oxide synthase with its substrate, l-arginine, on development of local mucosal injury and systemic capillary leak. MATERIALS AND METHODS: Rats underwent 30 min of superior mesenteric artery occlusion followed by 4 h of reperfusion. The vehicle groups received l-arginine either intravenously (4 mg/kg/min) or into the intestinal lumen. The intravenous groups received l-arginine either before the ischemic event or after 30 min of reperfusion. Capillary leak in the gut and lung were measured, as were degree of mucosal injury and number of infiltrating neutrophils. Appropriate controls were performed. RESULTS: Thirty minutes of mesenteric ischemia followed by 4 h of reperfusion significantly increased gut and lung leak, neutrophil infiltration, and the severity of mucosal injury. l-Arginine given iv prior to ischemia inhibited lung leak, mucosal injury, and neutrophil infiltration. When arginine was given during the reperfusion period, lung leak and neutrophil infiltration but not mucosal injury were reduced. Intraluminal l-arginine reduced mucosa injury, but had no effect on capillary leak. CONCLUSIONS: Supplementation with l arginine enhances NO production, resulting in reduced systemic endothelial dysfunction. This may act as a useful clinical adjunct in the management of trauma patients in preventing the development of ARDS and multiple organ failure. PMID- 10720448 TI - Severity of trauma changes expression of TNF-alpha mRNA in the brain of mice. AB - BACKGROUND: The greater nitrogen loss that occurs with increasing severity of trauma is believed to occur because activation of the hypothalamus-pituitary axis is greater with severe injury. Cytokines in the brain stimulate the hypothalamus pituitary-adrenal axis. This study was carried out to investigate whether the brain would recognize severity of trauma via TNF-alpha mRNA synthesis in the brain. METHODS: Male C57BL/6 mice (n = 70, BW: 20-28 g) were randomly assigned into four groups, (1) control (no anesthesia or incision), (2) anesthesia alone, (3) anesthesia plus laparotomy by short incision (short), and (4) anesthesia plus laparotomy by long incision (long). A laparotomy was carried out in the short and long groups by a 1.2-cm vertical incision and by a horizontal plus a vertical incision (2.4 x 2.4 cm), respectively. Exactly either 3 or 24 h after surgery, the animals were decapitated. TNF-alpha mRNA levels in the tissues were determined by semi-quantitative PCR. RESULTS: Nitrogen and catecholamine excretion were increased in the long wound group compared with the short wound group. Expression of TNF-alpha mRNA in the brain was greater in the long group after surgery than in the control, anesthesia, and short groups (brain, long: 0.150 +/- 0.005; P < 0.01 vs control, anesthesia alone, and short groups), but TNF-alpha levels in the plasma were the same in the short and long groups after surgery. CONCLUSION: Levels of TNF-alpha mRNA in the brain were enhanced according to the length of the wound probably because of greater neural stimuli from the wound site, and this elevation was involved in the greater nitrogen loss. PMID- 10720449 TI - Effect of endotoxemia on hepatic portal and sinusoidal blood flow in rats. AB - A decrease in liver blood flow leads to dysfunction of hepatocytes and Kupffer cells, with subsequent local and systemic liberation of proinflammatory mediators that may maintain systemic inflammatory response syndrome (SIRS) and may lead to multiple organ dysfunction syndrome (MODS). There is only limited knowledge on the hepatic micro- and macrocirculation during sepsis or endotoxemia. Therefore, the aim of our study was to investigate alterations in hepatic portal blood flow (PBF) and sinusoidal blood flow (SBF) during endotoxemia. In male Wistar rats endotoxemia was induced by continuous infusion of 2 mg/kg/h lipopolysaccharides from Escherichia coli 026:B6 immediately after baseline measurements (n = 8). The control group (n = 8) received an equivalent volume of Ringer's solution. Mean arterial pressure (MAP), heart rate (HR), cardiac output (CO), PBF, and SBF were measured at baseline and 60 and 120 min after induction of endotoxemia. PBF was measured using an ultrasonic flow probe that was positioned around the portal vein. SBF was detected by in vivo videomicroscopy of the left liver lobe. In the LPS group MAP decreased, but CO remained at baseline values. During endotoxemia PBF decreased significantly from 23 +/- 3 to 15 +/- 4 mL/min (60 min) and 16 +/- 3 mL/min (120 min). SBF also significantly decreased to 68.5% (60 min) and 57.1% (120 min) of baseline value. Our results demonstrate that during early endotoxemia hepatic macro- and microcirculatory perfusion is significantly decreased despite unchanged CO. This early reduction of hepatic perfusion might be caused by an increased hepatic vessel resistance as a consequence of liberation of vasoconstrictive mediators or/and by a decrease in intestinal perfusion. PMID- 10720450 TI - The important role of the gut in initiating the hyperdynamic response during early sepsis. AB - BACKGROUND: Although the initial response to sepsis includes a hyperdynamic phase and although the increased hepatic perfusion in early sepsis is due solely to the increased portal blood flow, it remains unknown whether the gut plays an important role in producing such a response. MATERIALS AND METHODS: Adult male Sprague-Dawley rats underwent a complete enterectomy (ER) before being subjected to sepsis by cecal ligation and puncture (CLP; the cecum was excised from the removed gut and stitched to the posterior peritoneum in ER groups) or sham operation. At 2 h after CLP (i.e., the early, hyperdynamic phase of sepsis), cardiac output and heart performance (+/-dP/dt(max)), as well as hepatic and renal blood flow, were measured. Systemic and regional oxygen delivery (DO(2)) and oxygen consumption (VO(2)) were also determined. RESULTS: Cardiac output, heart performance, organ blood flow, as well as DO(2) and VO(2), increased significantly 2 h after CLP. ER prior to the onset of sepsis, however, prevented the elevation of those parameters. ER in sham animals did not alter the measured parameters with the exception that portal blood flow decreased by 85% and hepatic arterial blood flow increased by 368%, resulting in no significant reduction in hepatic DO(2) and VO(2). There were no changes in circulating blood volume among groups, indicating that the effect of ER on hemodynamics after CLP was not due to alterations in blood volume. CONCLUSION: Since ER immediately before the onset of sepsis prevents the increase in cardiac output and regional hemodynamics, the gut appears to play an important role in producing the hyperdynamic response during the early stage of polymicrobial sepsis. PMID- 10720451 TI - Prolonged effect of leukocytosis on reperfusion injury of rat intestine: real time ATP change studied using (31)P MRS. AB - BACKGROUND: The intestine is one of the most sensitive tissues to ischemia and reperfusion (I/R). Polymorphonuclear neutrophils (PMN) may play an important role in ischemic injury. (31)P magnetic resonance spectroscopy (MRS) has been used to continuously monitor the energy metabolism of an animal in situ. We have applied MRS to study the effect of PMN on the I/R injury of rat intestine. MATERIAL AND METHODS: In a rat model of 30 min of intestinal ischemia and reperfusion, the number of PMNs was manipulated: group A, control; group B, leukopenia induced by cyclophosphamide; group C, leukocytosis induced by granulocyte colony-stimulating factor (G-CSF). MRS was employed to measure the level of real-time intestinal ATP and pH in vivo. RESULTS: In group A, ATP rapidly recovered on reperfusion to 61.0 +/- 11.0% of the preischemia level and maintained that level during reperfusion. The other two groups showed similar recovery of ATP at the initial phase of the reperfusion (<10 min). ATP in group B continued to recover, reaching 74.0 +/- 10.0% of the preischemia level. After the initial recovery, ATP in group C deteriorated reaching 46.0 +/- 4.4% of the preischemic level at 150 min of reperfusion. In group A and group B tissue pH decreased on ischemia and recovered on reperfusion in a similar manner. In group C, tissue pH was significantly lower than in other groups during I/R. CONCLUSION: Leukocytosis induced by G-CSF exerts a prolonged effect on ATP during I/R and leukocyte depletion helps protect against the I/R injury. PMID- 10720452 TI - Use of mathematical model to predict hemodynamics in cavopulmonary anastomosis with persistent forward flow. AB - BACKGROUND: The bidirectional cavopulmonary anastomosis with additional pulmonary blood flow is used as a staged procedure or a definitive palliation of univentricular hearts. In this paper the flow competition occurring between the caval and the pulmonary flows is investigated. The hemodynamics in the superior vena cava and the blood flow distribution into the lungs, as well as the systemic arterial oxygen availability, are correlated with the severity of the right ventricle outflow tract obstruction and the pulmonary arteriolar resistance. MATERIALS AND METHODS: Computer models of the pre- and postoperative hemodynamics of univentricular hearts were developed. The effects of increasing severity of the right ventricle outflow tract obstruction, with a pulmonary arteriolar resistance ranging from 0.8 to 7.9 nonindexed Woods units, were simulated. RESULTS: The study indicates that the presence of an additional pulmonary blood flow from the native pulmonary artery may be beneficial. Since an excessive additional blood flow may cause central venous hypertension, its optimal value should be chosen according to the value of pulmonary arteriolar resistance. The model was utilized to simulate four clinical cases. CONCLUSIONS: The simulations show that the model can predict the postoperative hemodynamics and could therefore be usefully applied to predict quantitatively the effect of the native pulmonary blood flow following bidirectional cavopulmonary anastomosis. PMID- 10720453 TI - Heparin-binding protein decreases apoptosis in human and murine neutrophils. AB - BACKGROUND: Heparin-binding protein (HBP), a serine protease without any known proteolytic activity, is found in human polymorphonuclear leukocyte (PMN) granules, but not in mice. HBP potentiates the endotoxin-induced release of tumor necrosis factor (TNF) alpha, interleukin (IL)-1, and IL-6 from isolated monocytes. HBP has also been shown to increase the survival of cultured monocytes and protect them from oxidative stress. However, whether HBP affects PMNs themselves is not known. MATERIALS AND METHODS: Based on our work with cultured monocytes and the survival benefit noted in experimental peritonitis, we hypothesized that HBP would have a beneficial effect on the survival of neutrophils. We evaluated the effect of HBP on apoptosis in murine peritoneal exudative cells elicited by intraperitoneal thioglycollate administration and in normal human neutrophils from volunteers. Leukocytes were isolated from the peritoneal cavity and blood of mice that underwent intraperitoneal thioglycollate instillation. The mouse peritoneal exudate cells and normal human neutrophils isolated from peripheral blood were used to study the effect of HBP on survival and apoptosis. RESULTS: HBP decreased percentage apoptosis of mouse cells in both serum-enriched (from 24.8 to 4.5%) and serum-deprived (from 23.1 to 8.2%) cultures. In human PMNs, the protective effect of HBP was seen only in the serum deprived group, with a decrease in percentage apoptosis from 69.1 to 43.3%. CONCLUSIONS: For the first time, we have shown that HBP, in addition to its known augmentation of the proinflammatory response of monocytes, also acts as a prosurvival protein for neutrophils themselves, and thereby enhances local host defense. PMID- 10720455 TI - Elevated interleukin-6 expression in keloid fibroblasts. AB - Keloids are characterized by a net accumulation of collagen. To date, the role of growth factors and various cytokines in the pathogenesis of these lesions has not been fully characterized. Interleukin-6 (IL-6) is an important immunoregulatory cytokine that has been implicated in a number of fibrotic autoimmune diseases such as scleroderma, interstitial nephritis, and pulmonary interstitial fibrosis. However, the role of IL-6 in the development of keloids has yet to be defined. This study demonstrates increased expression of the IL-6 gene in fibroblasts isolated from patients with keloids when compared with control fibroblasts using the ribonuclease protection assay. Subsequent detection of increased levels of IL 6 secretion by keloid fibroblasts is also demonstrated under unstimulated and stimulated conditions using serum and interferon gamma (IFN-gamma) (unstimulated: 0.3694 + 0.2499 pg/cell vs 0.0662 + 0.0786 pg/cell, P = 0.0137; serum: 1.066 + 0.513 pg/cell vs 0.233 + 0.231 pg/cell, P = 0.0027; serum and IFN-gamma: 1.286 + 0.395 pg/cell vs 0.244 + 0.199 pg/cell, P < 0.0001). These results suggest that IL-6 may play a significant role in the pathogenesis of keloids. PMID- 10720454 TI - A recombinant rat regenerating protein is mitogenic to pancreatic derived cells. AB - Pancreatic regenerating protein (reg I) is expressed in acinar cells and is mitogenic to beta- and ductal cells. Isolation of large amounts of endogenous reg I for in vivo and in vitro experiments has been difficult. The aim of this study was to develop a recombinant protein and determine its bioactivity on rat pancreatic derived cells. cDNA of the rat reg I coding region was created with unique BamHI flanking sequences using reverse transcriptase PCR. The coding region was then cloned into a bacterial expression vector in which expression is controlled by a T7 promoter. After transformation into the Escherichia coli strain B21(DE3) and induction by isopropyl-beta-d-thiogalactopyranoside, a fusion protein of 24 kDa in size, named reg-PET, was noted in the bacterial lysate. This protein contained a polyhistidine and S-peptide sequence to facilitate isolation and identification, respectively. This protein was purified using affinity chromatography, and identity was confirmed with gel electrophoresis and Western analysis. The reg-PET protein was mitogenic to both ARIP and RIN cells, rat pancreatic ductal and beta-cell lines, respectively. Antibodies raised to the protein reacted against rat reg I in pancreas. The purified recombinant reg I fusion protein, like endogenous reg I, is mitogenic to pancreatic derived cells. It is more potent than reg I isolated from pancreatic tissue. This protein can be isolated rapidly, easily, and with a high amount of purity. PMID- 10720456 TI - Carotid plaque gross morphology and clinical presentation: a prospective study of 457 carotid artery specimens. AB - BACKGROUND AND PURPOSE: In carotid artery disease, the relationship between carotid plaque morphology and the patient's neurologic symptoms is reportedly conflicting. The aim of this study was to correlate gross carotid plaque characteristics with the presenting symptoms in a relatively large series of patients who underwent carotid endarterectomy (CEA). METHODS: Four hundred and five patients who underwent 461 CEAs were divided into three groups: (1) transiently symptomatic [transient ischemic attack (TIA) or amaurosis fugax]; (2) prior stroke; and (3) asymptomatic. The degree of stenosis based on the preoperative angiograms was used in association with the presenting symptoms as the primary criterion in the decision to operate. Carotid plaque characteristics, including ulcerated plaque (UP), intraplaque hemorrhage (IH), uncomplicated plaque, and degree of stenosis, were recorded prospectively for 457 CEAs, since 4 CEAs were excluded from the study. All CEA specimens were grossly evaluated at surgery. RESULTS: There was a statistically higher incidence of UP in transiently symptomatic (P = 0.008) or prior stroke (P = 0.006) patients than in the asymptomatic group. When IH was considered independently, its incidence did not differ significantly between the three groups. Previously symptomatic patients tended to have higher-grade stenosis than asymptomatic patients, although the difference failed to reach statistical significance (P = 0.06). Although the incidences of UP and IH were higher in the higher-grade stenosis group, the difference was again not significant. CONCLUSIONS: Carotid UP correlates closely with an initial presentation of TIA, amaurosis fugax, or prior stroke, while the association between IH and presenting symptoms is less clear. Although there is an insignificant trend toward a correlation between the higher degrees of stenosis and the onset of transient symptoms, the degree of stenosis appears unaffected by the morphology of the plaque. These findings suggest that plaque morphology may play an important role in the presentation of carotid artery disease. PMID- 10720457 TI - Adaptive responses of the endothelium to stress. AB - It is now established that endothelial cells acquire several functional properties in response to a diverse array of extracellular stimuli. This expression of an altered phenotype is referred to as endothelial cell activation, and it includes several activities that promote inflammation and coagulation. While it is recognized that endothelial cell activation has a principal role in host defense, recent studies also demonstrate that endothelial cells are capable of complex molecular responses that protect the endothelium against various forms of stress including heat shock, hypoxia, oxidative stress, shock, ischemia reperfusion injury, toxins, wounds, and mechanical stress. In this review, we examine endothelial cell genotypic and phenotypic responses to stress. Also, we highlight important cellular stress responses that, although not yet demonstrated directly in endothelial cells, likely exist as part of the repertoire of stress responses in endothelium. A detailed understanding of the molecular mechanisms mediating the adaptive responses of endothelial cells to stress should facilitate the development of novel therapeutics to aid in the management of diverse surgical diseases and their complications. PMID- 10720458 TI - Editorial PMID- 10720459 TI - Applications of protein mass spectrometry in cell biology. AB - Advances in mass spectrometry combined with accelerated progress in genome sequencing projects have facilitated the rapid identification of proteins by enzymatic digestion, mass analysis, and sequence database searching. Applications for this technology range from the surveillance of protein expression in cells, tissues, and whole organisms, to the identification of proteins and posttranslational modifications. Here we consider practical aspects of the application of mass spectrometry in cell biology and illustrate these with examples from our own laboratories. PMID- 10720460 TI - Use of the two-hybrid system to identify Rab-interacting proteins. AB - The yeast two-hybrid system has been useful for identifying many partners and effectors of small GTPases of the Rab family. We describe here such a screen using Rab6, a protein involved in the regulation of intracellular transport at the level of the Golgi apparatus, as bait. PMID- 10720461 TI - Purification and identification of novel Rab effectors using affinity chromatography. AB - Rab GTPases are central regulatory elements of the intracellular transport machinery of eukaryotic cells. To regulate vesicle docking and fusion as well as organelle dynamics Rab proteins interact with effector molecules in the GTP-bound active state. The identification of Rab effectors is, therefore, of primary importance for the mechanistic understanding of intracellular transport. Here we describe the experimental system we have developed to biochemically purify and identify effectors of the small GTPase Rab5. The method, which is based on an affinity chromatography procedure, results in the large-scale purification of Rab effectors in amounts sufficient for both their identification by microsequencing techniques and their functional characterization. In the case of Rab5, the procedure allows a comprehensive analysis of the downstream effectors and regulators of this GTPase. We expect this strategy to provide fundamental insights into the molecular mechanism of membrane transport but also to be applicable to several other GTPase-dependent biological functions. PMID- 10720462 TI - Stage-specific assays to study biosynthetic cargo selection and role of SNAREs in export from the endoplasmic reticulum and delivery to the Golgi. AB - To analyze the role of coat protein type II (COPII) coat components and targeting and fusion factors in selective export from the endoplasmic reticulum (ER) and transport to the Golgi, we have developed three novel, stage-specific assays. Cargo selection can be measured using a "stage 1 cargo capture assay," in which ER microsomes are incubated in the presence of glutathione S-transferase (GST) tagged Sar1 GTPase and purified Sec23/24 components to follow recruitment of biosynthetic cargo to prebudding complexes. This cargo recruitment assay can be followed by two sequential assays that measure separately the budding of COPII coated vesicles from ER microsomes (stage 2) and, finally, delivery of cargo containing vesicles to the Golgi (stage 3). We show how these assays provide a means to identify the snap receptor (SNARE) protein rBet1 as an essential component that is not required for vesicle formation, but is required for vesicle targeting and fusion during ER-to-Golgi transport. In general, these assays provide an approach to characterize the biochemical basis for the recruitment of a wide variety of biosynthetic cargo proteins to COPII vesicles and the role of different transport components in the early secretory pathway of mammalian cells. PMID- 10720463 TI - The use of liposomes to study COPII- and COPI-coated vesicle formation and membrane protein sorting. AB - We have established systems that reconstitute the biogenesis of coated transport vesicles with liposomes made of pure lipids and purified coat proteins. Optimization of the lipid composition in the liposomes allowed the efficient binding of both coat protein I and coat protein II (COPII) coat subunits. Coated vesicles of approximately the size generated from biomembranes were detected and characterized by centrifugation analysis and electron microscopy. A variation of this budding reaction allowed us to measure the sorting of v-SNARE proteins into synthetic COPII vesicles. We developed a novel system to tether glutathione S transferase (GST)-hybrid proteins to the surface of liposomes formulated with a glutathione-derivatized phospholipid. This system allowed us to detect the positive role of cytoplasmic domains of two v-SNARE proteins that are packaged into COPII vesicles. Therefore, both generation of coated vesicles and protein sorting into the vesicles can be reproduced with liposomes and purified proteins. PMID- 10720464 TI - Fluorescent indicators of peptide cleavage in the trafficking compartments of living cells: peptides site-specifically labeled with two dyes. AB - When cells are infected with viruses, they notify the immune system by presenting fragments of the virus proteins at the cell surface for detection by T cells. These proteins are digested in the cytoplasm, bound to the major histocompatibility complex I glycoprotein (MHC-I) in the endoplasmic reticulum, and transported to the cell surface. The peptides are cleaved to the precise lengths required for MHC-I binding and detection by T cells. We have developed fluorescent indicators to study the cleavage of peptides in living cells as they are transported from the endoplasmic reticulum to the Golgi apparatus. Specific viral peptides known to be "trimmed" prior to cell surface presentation were labeled with two dyes undergoing fluorescence resonance energy transfer (FRET). When these fluorescent peptides were proteolytically processed in living cells, FRET was halted, so that each labeled fragment and the intact peptide exhibited different fluorescence spectra. Such fluorescent cleavage indicators can be used to study a wide range of biological behaviors dependent on peptide or protein cleavage. However, labeling a peptide with two dyes at precise positions can present a major obstacle to using this technique. Here, we describe two approaches for preparing doubly labeled cleavage indicator peptides. These methods are accessible to researchers using standard laboratory techniques or, for more demanding applications, through cooperation with commercial or core peptide synthesis services using minor modifications of standard synthetic procedures. PMID- 10720465 TI - In vitro generation from the trans-Golgi network of coatomer-coated vesicles containing sialylated vesicular stomatitis virus-G protein. AB - We describe an in vitro system in which post-Golgi vesicles containing metabolically labeled, sialylated, vesicular stomatitis virus (VSV) G protein molecules (VSV-G) are produced from the trans-Golgi network (TGN) of an isolated Golgi membrane fraction. This fraction is prepared from VSV-infected Madin-Darby canine kidney (MDCK) cells in which the (35)S-labeled viral envelope glycoprotein was allowed to accumulate in the trans-Golgi network during a prolonged incubation at 20 degrees C. The vesicles produced in this system are separated from the remnant Golgi membranes by differential centrifugation or by velocity sedimentation in a sucrose gradient. Vesicle production, quantified as the percentage of labeled VSV-G released from the Golgi membranes, is optimal at 37 degrees C and does not occur below 20 degrees C. It requires GTP and the small GTP-binding protein Arf (ADP-ribosylation factor), as well as coat protein type I (COPI) coat components (coatomer) and vesicle scission factors-one of which corresponds to the phosphatidylinositol transfer protein (PITP). Formation of the vesicles does not require GTP hydrolysis which, however, is necessary for their uncoating. Thus, vesicles generated in the presence of the nonhydrolyzable GTP analogs, GTPgammaS or GMP-PNP, retain a coatomer coat visible in the electron microscope, sediment more rapidly in sucrose density gradients than those generated with ATP or GTP, and can be captured with anticoatomerantibodies. The process of coatomer-coated vesicle formation from the TGN can be dissected into two distinct sequential phases, corresponding to coat assembly/bud formation and vesicle scission. The first phase is completed when Golgi fractions are incubated with cytosolic proteins and nonhydrolyzable GTP analogs at 20 degrees C. The scission phase, which leads to vesicle release, takes place when coated Golgi membranes, recovered after phase I, are incubated at higher temperatures in the presence of cytosolic proteins. The scission phase does not take place if protein kinase C inhibitors are added during the first phase, even though these inhibitors do not prevent membrane coating and bud formation. The phosphorylating activity of a protein kinase C, however, plays no role in vesicle formation, since this process does not require ATP. PMID- 10720466 TI - Site-specific photocrosslinking to probe interactions of Arf1 with proteins involved in budding of COPI vesicles. AB - ADP-ribosylation factor 1 (Arf1) plays an important role in early and intra-Golgi protein trafficking. During this process, Arf1 interacts with many different proteins and other molecules that regulate its state of activation or are involved in its intracellular function. To determine which of these proteins interact directly with Arf1 during coat protein type I (COPI) vesicle biogenesis, we probed the molecular environment of Arf1 by use of site-specific photocrosslinking. This method was first used successfully in the field of protein trafficking to study the mechanisms involved in protein translocation across the endoplasmic reticulum during protein synthesis. In such a hydrophobic environment, crosslink yields of up to 30% have been observed. We have now applied this method to study the mechanism of vesicle budding from the cytosolic face of the Golgi apparatus, an aqueous environment. Although the crosslink yield is significantly lower under these conditions, due to predominant reaction of the photolabile probes with water, a specific interaction of Arf1 with subunits of coatomer, the major coat protein of COPI vesicles, could readily be identified. PMID- 10720467 TI - Analysis of phosphoinositides in protein trafficking. AB - Phosphoinositides are key regulators of vesicle-mediated protein trafficking. Their roles include recruiting vesicle coat and effector proteins to the site of budding and promoting vesicle fusion. The intracellular levels of phosphoinositides and their localization to intracellular membranes are critical to their functions. An analytical procedure was developed that optimizes the recovery of radiolabeled cellular phosphoinositides. Quantitative analyses of yeast cellular phosphoinositides indicated that this approach is useful for examining the intracellular membrane phosphoinositide compositions related to trafficking phenomena. The approach will also enable investigators to determine whole-plant phosphoinositide compositions that have been difficult to achieve in the past. These analytical advances should be generally applicable to studies of phosphoinositide dynamics related to membrane trafficking in yeast, plant, and animal cells. PMID- 10720468 TI - Domain structure and function of dynamin probed by limited proteolysis. AB - Dynamin is a 100-kDa GTPase with multiple domains. Some of these have known functions, namely, the N-terminal GTPase domain, the PH domain that binds phosphatidylinositol lipids, and the C-terminal proline-arginine-rich domain (PRD) that binds to several SH3 domain-containing dynamin partners. Others, for example, the "middle" located between the GTPase domain and the PH domain and a predicted alpha-helical domain located between the PH domain and PRD, have unknown functions. Dynamin exists as a homotetramer in solution and self assembles into higher-order structures resembling rings and helical stacks of rings. Dynamin self-assembly stimulates its GTPase activity. We used limited proteolysis to dissect dynamin's domain structure and to gain insight into intradomain interactions that regulate dynamin self-assembly and stimulate GTPase activity. We found that the PH domain functions as a negative regulator of dynamin self-assembly and stimulates GTPase activity and that the alpha-helical domain, termed GED for GTPase effector domain, is required for stimulated GTPase activity. PMID- 10720469 TI - Development of HIV entry inhibitors targeted to the coiled-coil regions of gp41. AB - The discoveries that synthetic peptides corresponding to the N- and C-terminal heptad repeat (HR) regions of gp41 have potent anti-HIV activity opened a new avenue to identification of small molecule HIV entry inhibitors targeted to the HIV gp41 coiled-coil regions. Based on the structural information of the HIV gp41 core, three distinct approaches to develop small molecule anti-HIV agents have been reported. Each of these approaches has specific advantages, which will have complementary effects on the design of new strategies for identification of more potent HIV entry inhibitors. It is expected that novel antiviral drugs targeted to the HIV gp41 coiled-coil regions will be developed in the near future for the chemotherapy and/or prophylaxis of HIV infection and AIDS. PMID- 10720470 TI - Enhanced lipid oxidation by oxidatively modified myoglobin: role of protein-bound heme. AB - The formation of oxidized low density lipoprotein (LDL) is believed to play a significant role in the pathogenesis of atherosclerosis. Myoglobin in the presence of H(2)O(2) has been shown to catalyze LDL oxidation in vitro. It is established that an oxidatively altered form of myoglobin (Mb-H), which contains a prosthetic heme covalently crosslinked to the apoprotein, is a major product in the reaction of native myoglobin with peroxides. In the current study, we have shown for the first time that Mb-H, in the absence of exogenously added peroxides, oxidizes LDL and purified lipids, as determined by the formation of conjugated dienes, lipid peroxides, and thiobarbituric acid reactive substances. Moreover, the rate of oxidation of pure phosphatidylcholine by Mb-H was found to be at least sevenfold greater than that observed for native myoglobin. The current study strongly suggests a role for Mb-H in the lipid peroxidation observed with myoglobin. PMID- 10720471 TI - Rho-kinase inhibitor retards migration and in vivo dissemination of human prostate cancer cells. AB - The Rho-kinase inhibitor, Y-27632, inhibited in vitro chemotactic migration to bone marrow fibroblast conditioned media and metastatic growth in immune compromised mice of highly invasive human prostatic cancer (PC3) cells. Y-27632 also reduced myosin light chain phosphorylation and markedly altered the morphology of cells that developed numerous processes containing microtubules. A strikingly different, rounded phenotype was induced by an inhibitor of myosin light chain kinase, ML9. The M(110-130) subunit of the myosin phosphatase that is regulated by Rho-kinase was present in PC3 cells that contained significantly more RhoA than the less invasive, LNCaP cells. Y-27632 also inhibited angiogenesis as measured by endothelial cell tube formation on Matrigel. We conclude that invasiveness of human prostate cancer is facilitated by the Rho/Rho kinase pathway, and exploration of selective Rho-kinase inhibitors for limiting invasive progress of prostate cancer is warranted. PMID- 10720472 TI - Genetic dissection of the syndrome X in the rat. AB - In 1988, Reaven used the term syndrome X to describe a relation between several disorders including hypertension, dyslipidemia, impaired glucose tolerance, obesity, and coronary heart disease. Despite a number of studies dealing with syndrome X, its genetic basis remains poorly understood. Regarding the complexity of this syndrome, it is important to use animal models developing the traits of the disease. Here we show a genetic dissection of syndrome X in the WOKW rat, an animal model of genetically determined syndrome X. We found a major quantitative trait locus (QTL) for glucose metabolism on chromosome 3 and further QTLs influencing obesity and body weight on chromosomes 1 and 5. Genetic determinants of dyslipidemia were mapped to chromosomes 4 and 17. In addition, suggestive linkage for serum insulin was found on chromosome 1 to the region previously shown to be associated with type-1 diabetes mellitus. This is the first study demonstrating independent genetic factors influencing traits of the syndrome X in the rat as well as a possible genetic relationships between syndrome X and diabetes mellitus. Moreover, regarding the close similarities between WOKW rat and human syndrome X, the study could help in a search of genetic factors involved in this complex metabolic disorder in human. PMID- 10720473 TI - Expression of monocyte chemoattractant protein-1 by nonenzymatically glycated albumin (Amadori adducts) in vascular smooth muscle cells. AB - The biological effects of Amadori adducts that are early nonenzymatically glycated protein on vascular cells were poorly defined. We examined the effect of glycated serum albumin (GA) on the expression of monocyte chemoattractant protein 1(MCP-1) that is an important chemokine recruiting monocyte to blood vessel. GA increased MCP-1 mRNA expression with a peak after 3 h of stimulation. The induction of MCP-1 by GA was dose-dependent. The MCP-1 mRNA expression by GA was completely inhibited by PD98059 and genistein that inhibit mitogen activated protein (MAP) kinase kinase and tyrosine kinase, respectively. N-Acetylcysteine, a potent antioxidant, also suppressed the GA-induced MCP-1 expression. These results suggest that GA induces production of reactive oxygen species and activates tyrosine kinase and MAP kinase in VSMC. Activation of these signals results in MCP-1 expression. GA-induced MCP-1 expression may be one of the mechanisms by which the diabetic patients suffer from accelerated atherosclerosis. PMID- 10720474 TI - Recognition of human activated CD44 by tumor vasculature-targeted antibody. AB - TES-23 monoclonal antibody (MAb), which targets rat CD44H on tumor vascular endothelial cells (TEC), dominantly reacted to human activated CD44 rather than human inactive CD44. TES-23 MAb reacted to HT-1080 fibrosarcoma cells almost comparably to anti-human CD44 MAb and moderately to HUVEC; however, it hardly reacted to PBMC. The binding of soluble hyaluronate to HT-1080 cells and HUVEC was clearly noted, but not to PBMC. In addition, stimulation with phorbol 12 myristate 13-acetate induced soluble hyaluronate binding of MOLT-4 human T lymphoma cells and relatively increased the reactivity of TES-23 MAb. Our results suggest that TES-23 MAb can potentially recognize human activated CD44 and hence might be potentially useful for the treatment of human solid tumors containing TEC that express activated CD44. PMID- 10720475 TI - GSTM1 and mEPHX polymorphisms in Parkinson's disease and age of onset. AB - Both environmental and genetic factors are involved in the development of PD and biotransformation of exogenous and endogenous compounds and may play a role in inter-individual susceptibility. Therefore, we investigated the presence of null genotypes of GSTM1, GSTT1, and two polymorphisms of mEPHX in subjects with Parkinson's disease and in a reference population. The study included 35 male PD patients and a male control group including 283 subjects. Homozygosity of the histidine (H) 113 isoform of mEPHX was significantly increased in PD patients (odds ratio = 3.8 CI 95% 1. 2-11.8) and analysis of allele frequencies displayed an increased frequency of the H-allele among PD patients (odds ratio = 1.9 CI 95% 1.1-3.3). However, a significantly elevated median age for the onset of PD was found among GSTM1 gene carriers (median age = 68 years) compared to PD patients being GSTM1 null genotypes (median age = 57 years). Our observations suggest that (H) 113 isoform of mEPHX, which has been suggested as a low activity isoform, is overrepresented in PD patients and that inherited carriers of the GSTM1 gene postpone the onset of PD. These detoxification pathways may represent important protective mechanisms against reactive intermediates modifying the susceptibility and onset of PD. PMID- 10720476 TI - Pressure effects on tryptophan and its derivatives. AB - The high pressure effects on fluorescence of free tryptophan (Trp) and its derivatives, N-acetyl-tryptophan (AT), N-acetyl-tryptophanamide (NATA), tryptophanamide (TA), and tryptophan, containing 6-polypeptides in aqueous solution, were investigated in a pressure range from 0.1 to 650 MPa. It was found by analyzing the center of spectral mass in the wavelength range from 300 to 450 nm that high pressure shifted the fluorescence spectra of all these species to red direction: 421 cm(-1) for Trp, 305 cm(-1) for AT, 310 cm(-1) for NATA, 265 cm(-1) for TA, and 220 cm(-1) for single tryptophan containing 6-polypeptides. All the fluorescence efficiencies (i.e., quantum yield) of the compounds were reduced with pressure except free tryptophan where its fluorescence efficiency was enhanced with pressure. Glycerol, ethanol, and pH obviously influenced the pressure effects on their fluorescence characteristics. Since the tryptophan fluorescence is usually used as a probe for protein structural investigation, these findings suggested that the intrinsic pressure effect on tryptophan (or its derivatives) must be taken in consideration to explain the phenomenon observed in high pressure study on biomolecules when using the usual fluorospectroscopic approaches. In the present investigation, the mechanisms involved for pressure effects on tryptophan and its derivatives were explored and discussed. PMID- 10720477 TI - Pharmacological concentrations of arginine influence human whole blood viscosity independent of nitric oxide synthase activity in vitro. AB - l-Arginine, the natural precursor of NO, is infused in patients to restore endothelial function. Concentrations up to 7.5 mM l-arginine have been measured after parenteral administration. We investigated whether such high concentrations of amino acids influence blood viscosity in vitro. Incubation of whole blood from healthy volunteers with l-arginine, d-arginine, which has no effect on stereospecific NO synthases (NOS), the NOS substrate L-AME, the NOS inhibitor L NNA, the amino acids l-lysine and l-glutamic acid, and finally NaCl dose dependently decreased (up to 30% at 10(-2) M) low shear viscosity, which is primarily determined by erythrocyte aggregation. In contrast, the lipophilic NOS inhibitor L-NAME had no effect on low shear viscosity. All molecules failed to influence high shear viscosity, which is primarily determined by red cell deformability, and the erythrocyte shape remained unaltered. We conclude that high concentrations amino acids may decrease blood viscosity at low shear rate independent of NOS activity. This effect may contribute to the improved blood flow after intravascular administration of l-arginine. PMID- 10720478 TI - Studies on the relationships between the synonymous codon usage and protein secondary structural units. AB - The relationship between the synonymous codon usage and protein secondary structural elements (alpha helices and beta sheets) were reinvestigated by taking structural information of proteins from Protein Data Bank (PDB) and their corresponding mRNA sequences from GenBank for four different organisms E. coli, B. subtilis, S. cerevisiae, and Homo sapiens. It was observed that synonymous codon families have non-random codon usage, but there does not exist any species invariant universal correlation between the synonymous codon usage and protein secondary structural elements. The secondary structural units of proteins can be distinguished from the occurrences of bases at the second codon position. PMID- 10720479 TI - Diabetes enhances acetaldehyde-induced depression of cardiac myocyte contraction. AB - It is well established that cardiomyopathy is a consistent feature of diabetes and that alcohol consumption increases the risk of cardiovascular disease among diabetic subjects. Acetaldehyde (ACA), the main ethanol metabolite, is considered to play a role in the ethanol-induced cardiac dysfunction. It has been reported recently that the negative inotropic effect of ACA was more potent in the diabetic myocardium. To determine whether the disparate ACA-induced myocardial depression in diabetes is due to intrinsic alterations at the cellular level, mechanical properties in response to ACA were evaluated in ventricular myocytes from both normal and streptozotocin-induced diabetic rat hearts. Myocytes were electrically stimulated to contract at 0.5 Hz and contractile properties analyzed included peak shortening (PS), time-to-PS (TPS), time-to-90% relengthening (TR(90)) and maximal velocities of shortening and relengthening (+/-dL/dt). Ca(2+) transients were measured as fura-2 fluorescence intensity (DeltaFFI) changes. ACA (0. 1-30 mM) disproportionately depressed PS in a dose-dependent manner, in myocytes from diabetic hearts compared to normal hearts. Interestingly, the degree of inhibition in DeltaFFI was similar in both groups. Neither the duration nor maximal velocities of shortening and relengthening were affected by ACA in either group. These results are the first to suggest that enhanced ACA-induced myocardial depression in diabetes is due to disparate intrinsic actions on individual myocytes. The mechanism underlying the alteration of ACA-induced myocardial depression may be due, in part, to depressed Ca(2+) responsiveness in diabetic hearts. PMID- 10720480 TI - Neurotensin-mediated activation of MAPK pathways and AP-1 binding in the human pancreatic cancer cell line, MIA PaCa-2. AB - Neurotensin (NT), a gastrointestinal (GI) hormone, binds its receptor (NTR) to stimulate proliferation of normal and neoplastic GI tissues; the molecular mechanisms remain largely undefined. Mitogen-activated protein kinases (MAPKs) are a family of intracellular kinases that transmit mitogenic signals by translocating to the nucleus and activating transcription factors. The purposes of this study were: (1) to identify whether the MAPKs (ERK1/2 and JNK) are activated by NT and (2) to determine the effect of NT on downstream transcription factors using the human pancreatic adenocarcinoma cell line, MIA PaCa-2, which possesses high-affinity NTR. Both ERK and JNK activity were stimulated within 3-6 min by treatment with NT (10 nM); steady-state levels of ERK and JNK protein were unchanged. Moreover, NT treatment resulted in increased AP-1 binding activity as determined by gel shift analysis. Delineating the signal transduction mechanisms regulating the cellular effects of NT will provide important insights into the molecular pathways responsible for NT-mediated effects on both normal and neoplastic cells. PMID- 10720481 TI - Acrylamide quenching of apo- and holo-alpha-lactalbumin in guanidine hydrochloride. AB - We have examined the fluorescence properties and acrylamide quenching of calcium loaded (holo) and calcium-depleted (apo) alpha-lactalbumin (alpha-LA) as a function of guanidine hydrochloride (GDN/HCl) concentration. The spectral changes accompanying increasing GDN/HCl are consistent with protein unfolding and a release of internal fluorescence quenching, which occurs among the three tryptophan residues located in the region of the so-called "tertiary fold." Values for the intrinsic fluorescence emission, the wavelength maximum of the emission, the Stern/Volmer dynamic quench constant, and the static quench constant are consistent with a significant stabilization effect by calcium against protein unfolding. The dynamic quench constant of apo-alpha-LA increases fourfold to its maximum, in the transition from the native state to protein in 1.5 M GDN/HCl. The dynamic quench constant for holo-alpha-LA remains unchanged until exposed to 2.5 M GDN/HCl, but increases by threefold with addition denaturant to 4 M GDN/HCl. The static quench constant of the apo-protein in the native solvent, approximately 0.2 M(-1), declines to zero in 1 M denaturant, where the molten globule folding intermediate is most populated. A more protracted denaturant-dependent decline in the static quench constant occurs for the holo-protein. Sharp increase in the static quenching occurs for apo-alpha-LA and holo-alpha-LA above 1.5 M GDN/HCl and 3.5 M GDN/HCl, respectively. The results for apo-alpha-LA in dilute GDN/HCl suggest that acrylamide can penetrate the protein molecule (as judged by the collision quenching) but is unable to form a stable complex within the quenching domain for the tryptophans (as judged by the absence of the static quench constant). It seems reasonable to suggest that the protein folding intermediate which occurs in dilute denaturant represents a structure in which the tryptophans are, on average, more accessible to collisional quenching but sufficiently compact to prevent formation of a stable, dark equilibrium complex with acrylamide. PMID- 10720482 TI - Endothelin-1 induces NAD(P)H oxidase in human endothelial cells. AB - Superoxide anions (O(*-)(2)) induce oxidative stress and reduce endothelial NO availability by peroxynitrite formation. In human endothelial cells gp91(phox) was identified as the limiting subunit of the forming NAD(P)H oxidase. Because endothelin-1 (ET-1) is considered as a pro-atherosclerotic stimulus, we analyzed the effect of ET-1 on gp91(phox) expression and O(*-)(2) generation in primary cultures of human umbilical vein endothelial cells (HUVECs). The gp91(phox) mRNA expression was quantified by standard calibrated competitive reverse transcriptase-polymerase chain reaction. ET-1 induces gp91(phox) mRNA expression in HUVEC (max. after 1 h). The induction of gp91(phox) expression was dose dependent, reaching its maximum at 10 nmol/L ET-1. The increased gp91(phox) expression is mediated by endothelial receptor type B (ET(B)). Furthermore, ET-1 augments O(*-)(2) generation in human endothelial cells as measured by coelenterazine chemiluminescence. These data support a new mechanism: how ET-1 increases oxidative stress in the vessel wall leading to endothelial dysfunction and enhanced susceptibility to atherosclerosis. PMID- 10720483 TI - Expression of p16(INK4A) induces dominant suppression of glioblastoma growth in situ through necrosis and cell cycle arrest. AB - Tumor suppressor genes may represent an important new therapeutic modality in the treatment of human glioblastoma (GBM). p16(INK4A) is a tumor suppressor gene with mutation and/or deletion found in many human tumors, including glioblastomas, melanoma, and leukemias. RT-2 rat GBM cell line was used to investigate if the p16 gene induces dominant suppression of glioblastoma growth. Close to 100% of tumor cells were infected by high titer pCL retrovirus encoding the full-length human p16 cDNA at 5 m.o.i. Infected cells showed a 98% reduction in colony forming assay and a 60% reduction in growth curves in vitro compared to vector control. Exogenous overexpression of p16 induced hypophosphorylation of Rb protein by Western blot analysis. Intracranial injection of p16-infected tumor cells into syngeneic rats resulted in a 95% reduction in tumor volume compared to the controls. Intratumoral injection of p16 retrovirus resulted in tumor necrosis and prominent human p16 transgene expressions. Proliferation marker PCNA was not detected in these human p16-expressed RT-2 tumor cells, suggesting the cells were unable to enter into S phase after p16 expression. In addition, direct repeat intracranial injections of p16 retrovirus prolonged animal survival 3.2-fold compared to the controls (48.4 +/- 13.4 vs 15.0 +/- 2.1 days, p < 0.001). Two out of ten rats were found with dormant tumors at day 60 after p16 retrovirus injection. These results showed that p16 is effective in inhibiting GBM growth in situ. The mechanisms of tumor growth reduction and necrosis in vivo might be due to G1 arrest triggered by p16 expression. PMID- 10720484 TI - Copper has differential effect on prion protein with polymorphism of position 129. AB - The pathology of human prion diseases is affected by polymorphism at amino acid residue 129 of the prion protein gene. Recombinant mouse prion proteins mimicking either form of the polymorphism were prepared to examine their effect on the conformation and the level of superoxide dismutase (SOD) activity of the prion protein. Following the binding of copper atoms to prion protein, antibody mapping and CD analysis detected conformational differences between the two forms of protein. However, neither the level of copper binding nor the level of SOD activity associated with this form of prion protein altered with the identity of codon 129. These results suggest that in the holo-metal binding form of the protein, prion structure but not its SOD activity is affected by polymorphism at codon 129. PMID- 10720486 TI - Substrate binding to 15beta-hydroxylase (CYP106A2) probed by FT infrared spectroscopic studies of the iron ligand CO stretch vibration. AB - CYP106A2 has been expressed in E. coli with a high yield of up to 130 mg per litre of culture, purified to electrophoretic homogenity and found to be active in 15beta-hydroxylation of deoxycorticosterone using the adrenal redox proteins adrenodoxin and adrenodoxin reductase. Inspite of catalytic activity no substrate binding was detectable by UV-Vis spectroscopy. In contrast, an effect of substrate binding has been detected using the CO stretch mode infrared spectrum indicating that deoxycorticosterone binds in the heme pocket near the iron ligand. PMID- 10720485 TI - Equistatin, a protease inhibitor from the sea anemone actinia equina, is composed of three structural and functional domains. AB - A cDNA encoding a precursor of equistatin, a potent cysteine and aspartic proteinase inhibitor, was isolated from the sea anemone Actinia equina. The deduced amino acid sequence of a 199-amino-acid residue mature protein with 20 cysteine residues, forming three structurally similar thyroglobulin type-1 domains, is preceded by a typical eukaryotic signal peptide. The mature protein region and those coding for each of the domains were expressed in the periplasmic space of Escherichia coli, isolated, and characterized. The whole recombinant equistatin and its first domain, but not the second and third domains, inhibited the cysteine proteinase papain (K(i) 0.60 nM) comparably to natural equistatin. Preliminary results on inhibition of cathepsin D, supported by structural comparison, show that the second domain is likely to be involved in activity against aspartic proteinases. PMID- 10720487 TI - Identification and analysis of the sap genes from Vibrio fischeri belonging to the ATP-binding cassette gene family required for peptide transport and resistance to antimicrobial peptides. AB - Partial nucleotide sequences of the sapD and sapF genes of the sap operon (GenBank Accession No. AF178651) from Vibrio fischeri ATCC 7744 have been determined, and the peptide transport system of ATP-binding proteins SapD and SapF encoded by the genes have been deduced. Alignment and comparison of the Sap proteins of V. fischeri, Escherichia coli, Salmonella typhimurium, and Haemophilus influenzae Rd show that these proteins are homologous. The sap operon residing in the genome enables V. fischeri to transport peptides and resist antimicrobial peptides. Nucleotide sequence and functional analyses confirm that the specific regulatory-region-like sequence R&R* that resides inside the sapD gene and before the sapF gene functions in gene expression and regulation; also, it is regulated by the LuxR-AI complex of the V. fischeri lux regulon. The putative upstream activator binding sequences SigmaUASI, SigmaUASII, SigmaUASIII TGTCGACTTGGGCCTCGCTGTCCGTATGCACA (72nd to 103rd bp), TGTCCGTATGCACA (90th to 103rd bp), and TGTTCAAGTACCAGAAAGACA (111st to 133rd bp) in the R&R* sequence, which are similar to the two-component regulator binding sequence TGT-N(8-12)-ACA and the LuxR-AI binding sequence ACCTGTAGGATCGTACAGGT in the regulatory region of the V. fischeri lux regulon, might be the specific sequences recognized by the LuxR-AI complex for enhancement. PMID- 10720488 TI - Function of murine adenosine deaminase in the gastrointestinal tract. AB - Adenosine deaminase (ADA) deficiency in humans leads to a combined immunodeficiency characterized by severe T and B cell lymphopenia. ADA-deficient humans also display defective development of gut-associated lymphoid tissues (GALT). They lack lymphoid cells, and the Peyer's patches are without germinal centers. In mice, ADA-deficient fetuses die perinatally due to liver damage, but they also exhibit pathology in the thymus, spleen, and the small intestine. The GI phenotype associated with ADA-deficient humans prompted us to examine the effect of ADA-deficiency on mouse small intestine tissue. The work presented here focuses on understanding the physiological role of ADA in the GI tract, using ADA deficient mice rescued from perinatal lethality by restoring Ada expression to trophoblast cells. Histologically and immunologically, the GALT was compromised at all sites in ADA-/- mice, with the most dramatic changes seen in the Peyer's patches. Profound disturbances in purine metabolism were detected in all the gastrointestinal tissues. In particular, adenosine and deoxyadenosine, the ADA substrates, increased markedly while the product inosine decreased. The activity of S-adenosylhomocysteine hydrolase decreased throughout the GI tract, indicating a possible disruption of cellular transmethylation and activation of apoptotic pathways. There were also disturbances in the purine metabolic pathway with a decrease in the production of downstream nucleosides hypoxanthine and xanthine. PMID- 10720489 TI - Tip60 inhibits activation of CREB protein by protein kinase A. AB - We have previously identified a cDNA encoding a cellular protein, Tip60 (Tat interactive protein, 60 kDa), that specifically interacts with the Tat (transactivating transcriptional regulator) protein of the human immunodeficiency virus-1 (HIV-1). In this report, we have characterized cellular Tip and find that it is a 60 kDa nuclear protein expressed in a wide variety of differentiated cell lines from insects to man. To identify cellular functions of Tip, we have assayed the effects of Tip on cellular pathways that Tat has been reported to affect. Overexpression of Tip results in an almost complete block in activation of a Gal4 CREB (cAMP response element binding protein) fusion protein by cyclic AMP dependent protein kinase A (PKA). This inhibition appears to be mediated through direct interaction of Tip and CREB, since Tip directly binds to CREB protein in vitro. We show that amino acid substitutions of two conserved amino acids found in the putative acetyl coenzyme A binding motif of Tip completely abolishes the histone acetyltransferase (HAT) activity of recombinant Tip. Inhibition of CREB activation by Tip is not diminished in a HAT negative Tip mutant, indicating that Tip can negatively regulate gene expression independent of HAT activity. Recently, Tip has also been shown to be a transcriptional coactivator of nuclear hormone receptors; therefore, Tip can both activate transcription factors of one signaling pathway (nuclear hormone receptors) and bind to a different transcription factor (CREB) and inhibit activation of another signaling pathway. PMID- 10720490 TI - A genome-wide screening in Saccharomyces cerevisiae for genes that confer resistance to the anticancer agent cisplatin. AB - Cisplatin is a potent DNA-damaging agent that has demonstrated anticancer activities against several tumors. However, manifestation of cellular resistance is a major obstacle in anticancer therapy that severely limits the curative potential of cisplatin. Therefore, understanding the molecular basis of cisplatin resistance could significantly improve the clinical efficacy of this anticancer agent. Here, we employed Saccharomyces cerevisiae as a model organism to study cisplatin resistance mechanisms and describe a one-step cisplatin selection to identify and characterize novel cisplatin resistance genes. Screening a multicopy yeast genomic library enabled us to isolate several yeast clones for which we could confirm that the cisplatin resistance phenotype was linked to the introduced fragment. In a first attempt, a number of open reading frames could be identified. Among these genes, PDE2 and ZDS2 were repeatedly identified as genes whose overexpression confers cellular resistance to cisplatin. PDE2, encoding cAMP-phosphodiesterase 2, is of particular interest because the overexpression of this yeast gene is known to induce cisplatin resistance in mammalian cells as well, providing proof of the principle of our experimental approach. In addition, the identification of PDE2 shows that our yeast screening system can directly be informative for drug resistance in mammalian cells. PMID- 10720491 TI - Oxidized monocyte-derived macrophages in aortic atherosclerotic lesion from apolipoprotein E-deficient mice and from human carotid artery contain lipid peroxides and oxysterols. AB - Oxidative stress is thought to play an important role in atherogenesis. The present study demonstrated, for the first time, that macrophages (originally derived from blood monocytes) isolated from aortas of the atherosclerotic apolipoprotein E deficient (E degrees ) mice or from human carotid artery, are oxidized as they contain lipid peroxides and oxysterols. The major oxysterol in arterial macrophages was found to be 7-ketocholesterol (51% of total oxysterols). To find out whether lipid peroxidation of monocytes occurs in vivo already in the blood, we analyzed the oxidative state of monocytes derived from E degrees mice in comparison to monocytes from control mice. Cellular lipid peroxides and total oxysterols were four and sevenfold higher respectively, in monocytes derived from E degrees mice in comparison to monocytes from control mice. The results of the present study thus demonstrated the presence of lipid-peroxidized monocytes already in the blood, which are further oxidized in the arterial wall after their conversion into macrophages. The arterial oxidized macrophages could be considered key contributors to foam cell formation, the hallmark of early atherosclerosis. PMID- 10720492 TI - Viscoelastic properties of the cell nucleus. AB - Mechanical factors play an important role in the regulation of cell physiology. One pathway by which mechanical stress may influence gene expression is through a direct physical connection from the extracellular matrix across the plasma membrane and to the nucleus. However, little is known of the mechanical properties or deformation behavior of the nucleus. The goal of this study was to quantify the viscoelastic properties of mechanically and chemically isolated nuclei of articular chondrocytes using micropipet aspiration in conjunction theoretical viscoelastic model. Isolated nuclei behaved as viscoelastic solid materials similar to the cytoplasm, but were 3-4 times stiffer and nearly twice as viscous as the cytoplasm. Quantitative information of the biophysical properties and deformation behavior of the nucleus may provide further insight on the relationships between the stress-strain state of the nucleus and that of the extracellular matrix, as well as potential mechanisms of mechanical signal transduction. PMID- 10720493 TI - Expression of cold-adapted beta-tubulins confer cold-tolerance to human cellular microtubules. AB - Isolated microtubule proteins from the cold-adapted fish, Atlantic cod (Gadus morhua), assemble at temperatures between 8 and 30 degrees C, while avian and mammalian microtubules normally do not assemble at temperatures below 20 degrees C. Tubulin, the main component in microtubules, is expressed as many isotypes. Microtubules with different isotype composition have been shown to have different dynamic properties in vitro. Our hypothesis was that cold-tolerance of microtubules is caused by tubulin isotypes that differ in the primary sequence compared to mammalian tubulins. Here we show that transfection of human HepG2 cells with cod beta-tubulin induced cold-adaptation of the endogenous microtubules. Incorporation of one single tubulin isotype can induce cold tolerance to cold-intolerant microtubules. Three cod beta-tubulin isotypes were tested and two of these (beta1 and beta2) transferred cold-tolerance to HepG2 microtubules, thus not all cod beta-tubulins were able to confer cold-stability. PMID- 10720494 TI - Effects of an inhibitor of myosin light chain kinase on amylase secretion from rat pancreatic acini. AB - Ca(2+)/calmodulin-dependent protein (CaM) kinases play an important role in Ca(2+)-mediated secretory mechanisms. Previously, we demonstrated that a CaM kinase II inhibitor KN-62 had a small inhibitory effect on amylase secretion stimulated by CCK. In the present study, we investigated the effects of a myosin light chain kinase (MLCK) inhibitor on amylase secretion and Ca(2+) signaling in rat pancreatic acini. A specific inhibitor of MLCK, wortmannin, inhibited amylase secretion stimulated by CCK-8 (30 pM) in a concentration-dependent manner. Wortmannin (10 microM) had no effects on basal secretion but reduced amylase secretion stimulated by CCK-8 (30 pM) by 67 +/- 3%. Wortmannin inhibited amylase secretion stimulated by calcium ionophore (A23187) and phorbol ester (TPA). Wortmannin also inhibited amylase response to thapsigargin by 76 +/- 8% and to both thapsigargin and TPA by 52 +/- 10%. Ca(2+) oscillations evoked by CCK-8 (10 pM) were inhibited by wortmannin (10 microM). Wortmannin had a little inhibitory effect on an initial rise in [Ca(2+)](i), and abolished a subsequent sustained elevation of [Ca(2+)](i) evoked by 1 nM CCK-8. In conclusion, MLCK plays a crucial role in amylase secretion from pancreatic acini and regulates Ca(2+) entry from the extracellular space. PMID- 10720495 TI - Autocrine/Paracrine secretion of IL-6 family cytokines causes angiotensin II induced delayed STAT3 activation. AB - We recently reported that angiotensin II (AngII) biphasically activates the JAK/STAT pathway and induces delayed phosphorylation of STAT3 in the late stage (120 min) in cardiomyocytes. This study was designed to determine the mechanism of delayed phosphorylation of STAT3. Conditioned medium prepared from AngII stimulated cardiomyocytes could reproduce the tyrosine phosphorylation of STAT3 at 5 min. This delayed phosphorylation was almost completely inhibited by anti gp130 blocking antibody RX435, but not by TAK044 (ET-A/B-R antagonist), prazosin, or propranolol. AngII induced phosphorylation of gp130 in the late stage, which was temporally in parallel with the delayed phosphorylation of STAT3. AngII augmented IL-6, CT-1, and LIF mRNA expression at 30-60 min, but not CNTF expression. AngII increased IL-6 protein levels by 3-fold in the conditioned media at 2 h compared with the control. These findings indicated that AngII induced delayed activation of STAT3 is caused by autocrine/paracrine secreted IL 6 family cytokines. PMID- 10720497 TI - Induction and regulation of matrix metalloproteinase-12 by cytokines and CD40 signaling in monocyte/macrophages. AB - Matrix metalloproteinase-12 (MMP-12) has been shown to play critical roles in atherogenesis. To determine the cellular mechanisms for control of MMP-12 expression, we studied the effects of several cytokines (GM-CSF, IL-1beta, MCP-1) and CD40 ligand on MMP-12 expression in human monocyte/macrophages. Undifferentiated U937 monocytic cells and human peripheral blood monocytes did not express MMP-12. However, in the presence of GM-CSF, these monocytes showed MMP-12 expression at both the transcriptional and protein levels. The combination of treatment with GM-CSF and IL-1beta or MCP-1 resulted in a further increase of MMP-12 expression compared to treatment with GM-CSF alone. By contrast, both U937 derived macrophages and human peripheral blood monocyte-derived macrophages showed spontaneous MMP-12 expression, which was significantly increased by the addition of either GM-CSF or anti-CD40Ab. These results indicate that expression of MMP-12 is dependent upon the state of cellular differentiation and enhanced by cytokines and CD40 signaling. PMID- 10720496 TI - Trichoplusia ni lebocin, an inducible immune gene with a downstream insertion element. AB - A cDNA clone encoding a lebocin-like protein was obtained from the cabbage looper Trichoplusia ni by using differential display PCR. Northern blot analysis showed that lebocin gene expression was inducible upon bacterial challenge. Transcripts were mainly found in fat body but were also observed in hemocytes. Expression reached its highest level at 20 h and continued at least until 60 h after bacterial injection. The deduced protein is proline-rich and contains 143 amino acid residues. At position 128, a possible O-glycosylation site is observed. The whole protein shows 35% identity to Bombyx mori lebocin. The mature peptide displays an N-terminus similar to that of lebocin and a C-terminus to that of Drosophila metchnikowin. A 39-bp repetitive element is located downstream of the coding region. PMID- 10720498 TI - Metabolism and function of 3-D-phosphorylated phosphoinositides in C5a-stimulated eosinophils. AB - Eosinophils play a central role in the pathogenesis of parasitic infections, atopic diseases, and bullous dermatoses. To understand the regulative function of phosphatidylinositol 3-kinases in cell responses of eosinophils, phospholipid metabolism and production of reactive oxygen metabolites were followed after stimulation with C5a. Measurements of phosphatidylinositol lipids and analysis of deacylated products of separated lipid extracts showed fast and transient formation of phosphatidylinositol 3,4,5-trisphosphate (PIP(3)). Cell studies in the presence of the tyrosine kinase blocker genistein indicated that C5a stimulated PIP(3) formation occurred independently of tyrosine kinase activity. To analyze the function of PI4,5P(2)-3-kinase in eosinophils, the influence of wortmannin and LY294002 on production of reactive oxygen metabolites was studied. Both compounds inhibited with similar concentration dependency C5a-induced formation of PIP(3) and production of reactive oxygen metabolites. In summary, these data showed for the first time the involvement of PI4,5P(2)-3-kinase in the production of reactive oxygen metabolites in eosinophils. PMID- 10720499 TI - Mucosal and systemic immunity against poliovirus in mice transgenic for the poliovirus receptor: the poliovirus receptor is necessary for a virus-specific mucosal IgA response. AB - In view of the planned eradication of poliovirus, the suitability of transgenic mice bearing the human receptor for poliovirus (PVRtg mice) as a nonprimate animal model to study mucosal immunity against poliovirus was investigated. After intraperitoneal (ip) priming followed by ip or oral booster with live poliovirus, PVRtg mice had detectable IgA and IgG responses. The IgA response was restricted to PVRtg mice and could not be induced by oral immunization. After ip priming, PVRtg mice did shed virus in the stool, whereas control mice did not. Moreover, the amount of virus shed in the stools of PVRtg mice that had an IgA response after immunization was significantly lower than that of nonimmunized mice. A virus-specific mucosal IgA response is dependent on expression of the poliovirus receptor and is influenced by the route of immunization and the virus strain. PVRtg mice are a suitable model for the study of poliovirus-specific immunity and protection against poliovirus infection. PMID- 10720500 TI - Respiratory syncytial virus infection induces expression of the anti-apoptosis gene IEX-1L in human respiratory epithelial cells. AB - By means of differential display reverse-transcriptase polymerase chain reaction, increased expression of the mRNA encoding the anti-apoptosis gene IEX-1L was found in respiratory epithelial cells infected with respiratory syncytial virus (RSV). IEX-1L mRNA expression increased 5-7-fold in RSV-infected cells at 72 h after infection but remained unchanged in cells exposed to irradiated, replication-incompetent RSV. Because IEX-1L is reported to protect cells from apoptosis induced by tumor necrosis factor (TNF)-alpha, the effect of TNF-alpha on epithelial cell apoptosis in the context of RSV infection was determined. Epithelial cells were exposed to vehicle, RSV, or irradiated RSV for 72 h, and then TNF-alpha was added to appropriate cultures. Cytochemical staining of cellular DNA with 4,6-diamidino-2-phenylindole demonstrated TNF-alpha-induced apoptosis in 23.4% of control cells but only 5% of RSV-infected cells. These data show that RSV infection protects epithelial cells from TNF-alpha-induced apoptosis and that this effect is temporally associated with IEX-1L gene expression. PMID- 10720501 TI - The impact of influenza epidemics on hospitalizations. AB - The traditional method for assessing the severity of influenza seasons is to estimate the associated increase (i.e., excess) in pneumonia and influenza (P&I) mortality. In this study, excess P&I hospitalizations were estimated from National Hospital Discharge Survey Data from 26 influenza seasons (1970-1995). The average seasonal rate of excess P&I hospitalization was 49 (range, 8-102) /100,000 persons, but average rates were twice as high during A(H3N2) influenza seasons as during A(H1N1)/B seasons. Persons aged <65 years had 57% of all influenza-related hospitalizations; however, the average seasonal risk for influenza-related P&I hospitalizations was much higher in the elderly than in persons aged <65 years. The 26 pairs of excess P&I hospitalization and mortality rates were linearly correlated. During the A(H3N2) influenza seasons after the 1968 pandemic, excess P&I hospitalizations declined among persons aged <65 years but not among the elderly. This suggests that influenza-related hospitalizations will increase disproportionately among younger persons in future pandemics. PMID- 10720502 TI - Selective transmission of hepatitis B virus after percutaneous exposure. AB - In 3 clusters of postsurgical hepatitis B virus (HBV) infection, HBV DNA sequence mismatches were observed between the transmitting surgeons and the patients whom they infected. Sequence analysis of clones amplified from the C gene of HBV suggested that the mismatches were due to transmission of a minority variant in the circulation of each surgeon. Compared with 5 other transmitters from whom transmission of the dominant variant was demonstrated, the 3 surgeons who transmitted minority variants carried significantly more heterogeneous HBV populations. Transmission of minority variants was not correlated with the transmitters' hepatitis B antigen status, the presence of the position 1896 precore mutant, or the level of HBV viremia. In 1 cluster, a variant comprising <10% of the HBV population circulating in the transmitting surgeon established infection in all 3 patients who acquired HBV through him, which substantiates the phenomenon of true selection. PMID- 10720503 TI - Determinants of the quantity of hepatitis C virus RNA. AB - To test the hypothesis that person-to-person variability in blood levels of hepatitis C virus (HCV) RNA can be explained, the quantity of HCV RNA was assessed in 969 persons who acquired HCV infection in the context of injection drug use. Serum HCV RNA levels ranged from 200,000 to >120 million equivalents/mL (the linear range of the assay). The median log10 HCV RNA level was 0.46 higher in 468 human immunodeficiency virus (HIV)-positive persons than in 501 HIV negative persons (P<.001). In addition, among HIV-negative persons, lower HCV RNA levels were independently associated with younger age (P<.001), ongoing hepatitis B infection (P=.005), and the absence of needle sharing (P=.02). However, >90% of the person-to-person HCV RNA level variability was not explained by these sociodemographic, environmental, and virologic factors. Additional research is necessary to ascertain what determines the level of HCV RNA in blood. PMID- 10720504 TI - Longitudinal analysis of hepatitis C virus replication and liver fibrosis progression in renal transplant recipients. AB - The pathogenesis of hepatitis C virus (HCV) infection was investigated by analysis of changes in viral and histologic parameters in 36 renal transplant recipients who were infected with HCV before transplantation. Each patient was classified according to development of liver fibrosis as assessed by 2 liver biopsies done 45 and 81 months after transplantation: 13 had progressing liver fibrosis (fibrosers) and 23 did not (nonfibrosers). All developed high-titer posttransplant viremia with a significant increase of 1.2 log RNA copies/mL. There were no significant differences in the increases in serum HCV RNA or genotype distributions in fibrosers and nonfibrosers. The hypervariable region (HVR)-1 of the HCV genome was analyzed by cloning and sequencing 20 clones per sample from 5 fibrosers and 5 nonfibrosers. Comparison of samples revealed that liver fibrosis progression was significantly associated with slower HVR-1 quasispecies diversification, suggesting the selection of more aggressive variants in fibrosers. PMID- 10720505 TI - Frequencies of memory T cells specific for varicella-zoster virus, herpes simplex virus, and cytomegalovirus by intracellular detection of cytokine expression. AB - Memory T cells specific for varicella-zoster virus (VZV), herpes simplex virus (HSV), and human cytomegalovirus (HCMV) were compared in immune adults by intracellular cytokine (ICC) detection. The mean percentages of CD4+ T cells were 0.11% for VZV and 0.22% for HSV by interferon (IFN)-gamma production; the frequency for HCMV was significantly higher at 1.21%. Percentages of VZV-, HSV-, and HCMV-specific CD4+ T cells were similar by use of tumor necrosis factor (TNF) alpha. HCMV-stimulated CD8+ T cells produced IFN-gamma (1.11%) and TNF-alpha (1.71%); VZV- and HSV-specific CD8+ T cells were not detectable. VZV CD4+ T cell numbers were similar in young adults with natural or vaccine-induced immunity. VZV CD4+ T cells were significantly less frequent in older adults. Secondary varicella immunization did not increase VZV-specific CD4+ T cell frequencies by ICC assay. Numbers of memory T cells specific for herpesviruses may vary with sites of viral latency and with host age. PMID- 10720506 TI - Quantitative image analysis of simian immunodeficiency virus replication in macrophages coinfected with Mycobacterium avium complex. AB - Mycobacterium avium is the most frequent cause of disseminated bacterial infection in patients with human immunodeficiency virus type 1 infection and in rhesus macaques with simian immunodeficiency virus (SIV) infection. This animal model of AIDS was used to test the hypothesis that this frequent association is the result of reciprocal enhancement of replication of both microorganisms. The replication of M. avium and SIV was analyzed in lymphatic tissues obtained from rhesus macaques experimentally inoculated with SIVmac who developed or remained free of overt M. avium infection. In situ hybridization, quantitative image analysis, and staining of M. avium and of macrophages were used to assess the effects of coinfection on the replication of SIV and M. avium in vivo. There was no correlation between virus load and M. avium load in coinfected lymph nodes, and, with one exception, there was no evidence that M. avium coinfection of macrophages increased SIV replication. PMID- 10720507 TI - Natural history of human immunodeficiency virus type 1 viremia after seroconversion and proximal to AIDS in a large cohort of homosexual men. Multicenter AIDS Cohort Study. AB - The natural history of human immunodeficiency virus type 1 (HIV-1) viremia and its association with clinical outcomes after seroconversion was characterized in a cohort of homosexual men. HIV-1 RNA was measured by reverse-transcription polymerase chain reaction (RT-PCR) in stored longitudinal plasma samples from 269 seroconverters. Subjects were generally antiretroviral drug naive for the first 3 years after seroconversion. The decline in CD4 lymphocyte counts was strongly associated with initial HIV RNA measurements. Both initial HIV RNA levels and slopes were associated with AIDS-free times. Median slopes were +0.18, +0.09, and -0.01 log10 copies/mL, respectively, for subjects developing AIDS <3, 3-7, and>7 years after seroconversion. In contrast, HIV RNA slopes in the 3 years preceding AIDS and HIV RNA levels at AIDS diagnosis showed little variation according to total AIDS-free time. HIV RNA load at the first HIV-seropositive visit ( approximately 3 months after seroconversion) was highly predictive of AIDS, and subsequent HIV RNA measurements showed even better prognostic discrimination. PMID- 10720508 TI - Efficacy testing of recombinant human immunodeficiency virus (HIV) gp160 as a therapeutic vaccine in early-stage HIV-1-infected volunteers. rgp160 Phase II Vaccine Investigators. AB - A phase II efficacy trial was conducted with recombinant human immunodeficiency virus (HIV) type 1 envelope glycoprotein gp160 (rgp160) in 608 HIV-infected, asymptomatic volunteers with CD4+ cell counts >400 cells/mm3. During a 5-year study, volunteers received a 6-shot primary series of immunizations with either rgp160 or placebo over 6 months, followed by booster immunizations every 2 months. Repeated vaccination with rgp160 was safe and persistently immunogenic. Adequate follow-up and acquisition of endpoints allowed for definitive interpretation of the trial results. There was no evidence that rgp160 has efficacy as a therapeutic vaccine in early-stage HIV infection, as measured at primary endpoints (50% decline in CD4+ cell count or disease progression to Walter Reed stage 4, 5, or 6) or secondary endpoints. A transient improvement was seen in the secondary CD4 endpoint for the vaccination compared with the placebo arm, but this did not translate into improved clinical outcome. PMID- 10720509 TI - Lymphoproliferative responses to recombinant HIV-1 envelope antigens in neonates and infants receiving gp120 vaccines. AIDS Clinical Trial Group 230 Collaborators. AB - Children of mothers infected with human immunodeficiency virus type 1 (HIV-1) were immunized at birth and at 1, 3, and 5 months with 1 of 3 doses of recombinant gp120 vaccines prepared from SF-2 or MN strains of HIV-1. A total of 126 children were not infected; 21 received adjuvant only. Vaccine recipients developed lymphoproliferative responses on >/=2 occasions, responding more often to homologous HIV-1 antigens than did adjuvant recipients (56% vs. 14%; P<.001). Responses were appreciated after 2 immunizations and were maintained for >84 weeks after the last immunization. An accelerated immunization schedule (birth, 2 weeks, 2 months, and 5 months) with the lowest dose of the SF-2 vaccine produced responses in all 11 vaccinees by 4 weeks. Responses to heterologous envelope antigens were also detected. Immune responses to vaccination are achievable at an age when some infection (perinatal or breast milk exposure related) may be prevented. PMID- 10720510 TI - Resistance to human immunodeficiency virus type 1 in vitro as a surrogate of vaccine-induced protective immunity. AB - An in vitro assay developed as a correlate of vaccine-induced protection from human immunodeficiency virus (HIV) was validated in populations with relative resistance to HIV-1 as well as in HIV vaccine recipients. Cultures of peripheral blood mononuclear cells (PBMC) were challenged with 10 TCID50 of HIV-1MN or HIV 1BaL, titered in PBMC from normal controls (n=57). PBMC from HIV-1-infected persons with low viremia (n=17), exposed uninfected persons (n=23), and HIV-2 infected Senegalese prostitutes (n=9) were significantly resistant to HIV-1BaL and/or HIV-1MN (P<.001). Among 34 HIV vaccine recipients of live canarypox vector expressing multiple HIV-1 gene products with or without rgp120 booster, PBMC from postvaccination samples were significantly resistant to both strains (P<.001), and cytotoxic T lymphocyte precursor-positive samples were significantly more resistant than were precursor-negative samples (P<.03). This is the first evidence of the induction by vaccination of a validated correlate of protection. This assay should serve as a useful criterion for assessing experimental HIV vaccines before phase III efficacy trials. PMID- 10720511 TI - Patterns of resistance mutations selected by treatment of human immunodeficiency virus type 1 infection with zidovudine, didanosine, and nevirapine. AB - Resistance mutations selected in reverse transcriptase (RT) by incompletely suppressive therapy with combination zidovudine and didanosine with or without nevirapine were identified in 141 human immunodeficiency virus type 1 isolates from peripheral blood mononuclear cells of 57 individuals in the AIDS Clinical Trials Group protocol 241. After prolonged treatment (16-48 weeks), the most common nevirapine-selected mutations were RT 181C (15/30 isolates [50%]), 190A (15/30 [50%]), and 101E (9/30 [30%]). RT 103N and 188L, which individually confer cross-resistance to all nonnucleoside RT inhibitors, were seen in a minority of viruses (6/30 [20%] and 4/30 [13%], respectively). Didanosine-resistance mutations arose rarely. A newly recognized mutation, RT 44D, was selected by the nucleosides. Two distinct zidovudine-resistance mutational patterns were noted. Mutations selected during treatment with zidovudine, didanosine, and nevirapine differed among individuals and changed over time. Resistance testing is necessary to identify which mutations are selected by nevirapine-containing combinations. PMID- 10720512 TI - Resistance profile of the human immunodeficiency virus type 1 reverse transcriptase inhibitor abacavir (1592U89) after monotherapy and combination therapy. CNA2001 Investigative Group. AB - Abacavir (1592U89) is a nucleoside inhibitor of human immunodeficiency virus (HIV) type 1 reverse transcriptase (RT). Resistance to abacavir was studied with abacavir alone and with abacavir in combination with other nucleoside analogues in cell culture, in virus isolates from zidovudine/lamivudine clinical trials, and in the first dose-escalating 12-week clinical trial (CNA2001) to evaluate abacavir clinical potency. Abacavir alone in vitro selected for mutations at HIV RT codons K65R, L74V, Y115F, and M184V. However, abacavir combined with zidovudine selected against virus with the M184V mutation. Abacavir therapy in vivo resulted in large decreases in HIV load (>1 log), even in 1 subject who had the M184V mutation at baseline. A total of 51% of subjects showed new mutations at any of codons K65R, L74V, and M184V after abacavir monotherapy, compared with 11% who received zidovudine/abacavir. Small changes (2- to 4-fold) in abacavir susceptibility were detected. On stopping therapy, reselection of the pretherapy sequence occurred within 4 weeks. PMID- 10720513 TI - Chimeric toxins targeted to the human immunodeficiency virus type 1 envelope glycoprotein augment the in vivo activity of combination antiretroviral therapy in thy/liv-SCID-Hu mice. AB - Highly active antiretroviral therapy (HAART), which combines multiple inhibitors of essential human immunodeficiency virus type 1 (HIV-1) enzymes, induces dramatic and sustained viral load reductions in many people infected with HIV-1. However, reservoirs of infected cells capable of producing replication-competent virus persist even after years of HAART, preventing elimination of infection. CD4 PE40 and 3B3(Fv)-PE38, chimeric toxins designed to target the HIV envelope (Env), represent a complementary class of agents that selectively kill productively infected cells. To investigate whether these Env-targeted toxins might serve as adjuncts to HAART for the elimination of infected cells, we tested their ability to augment HAART efficacy in vivo by using a thy/liv SCID-hu mouse model. CD4 PE40 and 3B3(Fv)-PE38 markedly enhanced the capacity of HAART to suppress acute HIV-1 infection and improved HAART-mediated viral load reduction in mice with established HIV-1 infection. These results represent the first demonstration of in vivo anti-HIV-1 efficacy for Env-targeted toxins and support their potential therapeutic utility in combination with HAART. PMID- 10720514 TI - CD4+ T cell surface CCR5 density as a determining factor of virus load in persons infected with human immunodeficiency virus type 1. AB - The intensity of expression of the chemokine receptor CCR5 is involved in in vitro cell infectability by human immunodeficiency virus (HIV)-1 R5 isolates. Because CCR5 expression varies among individuals, the hypothesis that this expression could determine virus load in HIV-1-infected persons was tested. The mean number of CCR5 molecules per cell was measured on peripheral blood CD4+ T lymphocytes (CCR5 density) from HIV-1-infected, asymptomatic, nontreated adults. There was a strong correlation between HIV RNA plasma level and CCR5 density (P=.009) that was independent of cell activation and was not due to an HIV induced CCR5 up-regulation. These data are compatible with the hypothesis that CCR5 density is a key factor governing cell infectability and in vivo virus production and explain the protective effect of the Delta32CCR5 deletion, which results in low CCR5 expression. CCR5 density might be of critical predictive value in HIV infection. PMID- 10720515 TI - Interleukin-2 up-regulates expression of the human immunodeficiency virus fusion coreceptor CCR5 by CD4+ lymphocytes in vivo. AB - Intermittent interleukin-2 (IL-2) therapy can substantially increase CD4+ T cell counts of human immunodeficiency virus (HIV)-infected subjects. Administration of IL-2 led to transient up-regulation of CCR5 on CD4+ T cells; up to 87% of CD4+ cells expressed CCR5 after a 5-day cycle, with return to baseline levels within 2 weeks. Unlike in vitro studies, CCR5 was coexpressed with CD45RA and CXCR4 on CD4+ T cells after IL-2 therapy. The observed increase in coreceptor expression was not associated with detectable increases in viral replication. IL-2 therapy induced CCR5 expression in >90% of circulating memory CD4+ T cells, determined to be a long-term reservoir of HIV, suggesting significant activation of these cells. These studies demonstrate that levels of expression of HIV coreceptors alone do not always correlate with HIV replication in vivo and that IL-2 therapy activates a majority of memory T cells in the circulation and likely throughout the immune system. PMID- 10720516 TI - Cytokine profile in genital tract secretions from female adolescents: impact of human immunodeficiency virus, human papillomavirus, and other sexually transmitted pathogens. AB - Quantitative enzyme-linked immunosorbent assays were used to measure interleukin (IL)-2, IL-10, and IL-12 in cervical secretions from female adolescents with and without sexually transmitted infections. Compared with human immunodeficiency virus [HIV]-negative patients, HIV-positive patients had higher concentrations of IL-10 (118.2 pg/mL vs. 34.5 pg/mL; P=.002) and IL-12 (175.5 pg/mL vs. 85.1; P=.03). IL-2 concentrations were not statistically different. Furthermore, genital tract infections were predictors of IL-10 and IL-12 concentrations. Coinfection with HIV and human papillomavirus predicted the highest IL-10 concentrations; coinfection with HIV, human papillomavirus, and other sexually transmitted pathogens predicted the highest IL-12 concentrations. The data indicate that concomitant infection of the genital tract with HIV and other viral, bacterial, or protozoan pathogens influences the local concentrations of some immunoregulatory cytokines. PMID- 10720517 TI - Sustained CD4+ T cell response after virologic failure of protease inhibitor based regimens in patients with human immunodeficiency virus infection. AB - The relationship between plasma human immunodeficiency virus (HIV) RNA levels and peripheral CD4+ T cell counts was examined in 380 HIV-infected adults receiving long-term protease inhibitor therapy. Patients experiencing virologic failure (persistent HIV RNA >500 copies RNA/mL) generally had CD4+ T cell counts that remained greater than pretherapy baseline levels, at least through 96 weeks of follow-up. The CD4+ T cell response was directly and independently related to degree of viral suppression below the pretreatment baseline. For any given HIV RNA level measured 12 weeks after virologic failure, subsequent CD4+ T cell decline was slower in patients receiving a protease inhibitor-based regimen than in a historical control group of untreated patients. These observations suggest that transient or partial declines in plasma HIV RNA levels can have sustained effects on CD4+ T cell levels. PMID- 10720518 TI - Thalidomide-induced antigen-specific immune stimulation in patients with human immunodeficiency virus type 1 and tuberculosis. AB - Thalidomide, which inhibits monocyte tumor necrosis factor (TNF)-alpha production and costimulates T cells, was tested for immune modulation in patients with human immunodeficiency virus (HIV) infection and tuberculosis (TB) in a placebo controlled study. Thalidomide therapy resulted in increased levels of plasma interleukin (IL)-2 receptor, soluble CD8, interferon-gamma, and IL-12, indicating immune stimulation. TNF-alpha levels were not reduced. Thalidomide treatment increased CD4+ and CD8+ T cell counts and lymphocyte proliferation to purified protein derivative. Immune stimulation was not associated with an increase in plasma HIV levels. In vivo, a thalidomide dose-dependent costimulatory effect on T cell proliferation and HIV replication was observed after stimulation with antigens or anti-CD3, respectively. Thalidomide-induced increased viral replication in CD4+ T cells was abrogated by adding back autologous CD8+ T cells. Thus, in the presence of thalidomide, antigen-specific immune responses in vitro and in patients with HIV/TB were enhanced. PMID- 10720519 TI - Molecular typing of Streptococcus pneumoniae in northeastern Romania: unique clones of S. pneumoniae isolated from children hospitalized for infections and from healthy and human immunodeficiency virus-infected children in the community. AB - Microbiologic, serologic, and molecular typing techniques were used to characterize 272 isolates of Streptococcus pneumoniae colonizing or infecting children in Iasi, Romania, during a surveillance study conducted in 1996-1998. The 574 children in the study were from the following groups: healthy children attending 2 institutions, healthy children hospitalized for elective surgery, hospitalized children with pneumococcal infections, and human immunodeficiency virus (HIV)-infected children in an orphanage. Pneumococci colonizing healthy children from closed communities showed close similarities to pneumococci from children with pneumococcal infections; they expressed a limited number of similar serotypes, showed high frequency of penicillin and multidrug resistance, and shared several common clonal types. In contrast, isolates recovered from healthy children hospitalized for elective surgery expressed a large variety of serotypes, were less frequently resistant to antimicrobial agents, and showed great genetic diversity. Pneumococcal flora colonizing HIV-infected children showed a more complex epidemiology. These observations suggest a possible epidemiologic connection between the flora of S. pneumoniae colonizing healthy children in closed communities and the flora found in children hospitalized for infection. PMID- 10720520 TI - Genomic differences in Streptococcus pyogenes serotype M3 between recent isolates associated with toxic shock-like syndrome and past clinical isolates. AB - Genomic differences among past Streptococcus pyogenes serotype M3 strains isolated in 1973 and before from patients with streptococcal pharyngitis, recent (1990s) serotype M3 clinical isolates from patients with pharyngitis, and recent M3 isolates from patients with toxic shock-like syndrome were investigated by restriction landmark genomic scanning and by modified random-amplified polymorphic DNA-polymerase chain reaction. Similar polymorphic DNA fragments were identified between the older M3 isolates and the recent isolates; also, the recent M3 clinical isolates from patients with pharyngitis were genetically indistinguishable, by the methods used, from the M3 isolates of patients with toxic shock-like syndrome. Although nucleotide sequences of these regions showed no apparent homology with known virulence factors, the DNA fragments could distinguish the recent M3 strains from the past strains. These results suggested that the recent strains have emerged because of genetic divergence. PMID- 10720521 TI - Molecular epidemiology of community-acquired Staphylococcus aureus in families with and without cystic fibrosis patients. AB - The molecular epidemiology of Staphylococcus aureus nasal commensal strains and community-acquired infecting strains was assessed by comparison of prevalence, persistence, transmission rate, and clonal distribution of S. aureus in families with and without cystic fibrosis (CF) patients. Isolates were typed by pulsed field gel electrophoresis. CF patients without antibiotic treatment had a significantly higher nasal prevalence (66%) of S. aureus than did treated patients (29%; P<.001) or healthy controls (32%; P<.001), suggesting that persons with CF have a higher susceptibility to this organism. Strain transmission was frequent within both CF (55%) and non-CF (62%) families. After 3 and 19 months, 57% and 21%, respectively, of all persons still harbored the same S. aureus strain. Most of the isolates (78%) belonged to 8 of 38 genome types common in CF patients and in healthy persons. The predominant occurrence of a limited number of S. aureus clones within the community suggests evolutionary mechanisms for the selection of certain strains without an obvious association with disease. PMID- 10720522 TI - Molecular analysis of the accessory gene regulator (agr) locus and balance of virulence factor expression in epidemic methicillin-resistant Staphylococcus aureus. AB - Potential relationships between virulence factor expression and transmissibility were assessed in epidemic methicillin-resistant Staphylococcus aureus (MRSA) clones CMRSA-1 and CMRSA-3. A major subtype of CMRSA-1 exhibited normal transcription of RNAIII, which facilitates the induction of secreted virulence factors and repression of colonization factor expression at high cell density. However, these isolates characteristically did not express alpha-toxin or protease and displayed a limited profile of secreted proteins. CMRSA-1 also expressed a novel cell surface glycoprotein and exhibited a unique polymorphism within the accessory gene regulator (agr) locus. CMRSA-3 displayed attenuated activation of RNAIII transcription, which was consistent with its higher fibronectin-binding and coagulase activity relative to sporadic MRSA or CMRSA-1 (P=.05), low protease activity, and limited profile of secreted proteins. Thus, the balance of virulence factor expression in CMRSA-1 and CMRSA-3 favors the colonization phase of infection, and CMRSA-1 possesses unique genotypic and phenotypic traits. PMID- 10720523 TI - Universal epitopes for human CD4+ cells on tetanus and diphtheria toxins. AB - Previous studies suggested that tetanus and diphtheria toxoids (TTD and DTD, respectively) contain "universal" epitopes for human CD4+ cells (residues 632-651 and 950-969 of TTD and 271-290, 321-350, 351-370, 411-430, and 431-450 of DTD). To investigate whether CD4+ cells of 100 randomly selected subjects recognized those sequences, the proliferation of CD4+ cell-enriched blood lymphocytes to TTD and DTD and individual synthetic universal epitopes was measured. CD4+ cells of 98 subjects recognized both toxoids, those of 1 subject only TTD, and those of 1 only DTD. The TTD peptides and DTD peptides 271-290 and 331-350 were recognized by >/=80% of the toxoid-sensitized subjects. The other DTD sequences were recognized by 63%-71% of subjects. DR-homozygous subjects recognized several universal epitopes less frequently than did DR-heterozygous subjects. The intensity of responses to the epitope peptides correlated with that to TTD or DTD, consistent with recognition of the peptides by CD4+ cells specific for the cognate toxoid. PMID- 10720524 TI - Correlation between pertussis toxin IgG antibodies in postvaccination sera and subsequent protection against pertussis. AB - All acellular pertussis vaccines contain pertussis toxoid and induce protection against pertussis. This study investigated the relation between the postvaccination levels of pertussis toxin (PT) serum IgG and protection against pertussis. PT IgG was determined in sera obtained 21-77 days after the third vaccination from 813 children who received 3 doses of pertussis toxoid. The children were followed for 21-33 months after vaccination for the occurrence of pertussis. Of the children, 126 were exposed to pertussis in their households. The median PT IgG concentration was 79 U/mL in those who developed severe pertussis (>/=21 day of paroxysmal cough), 156 U/mL with mild pertussis (<21 days of paroxysmal cough), and 246 U/mL in those who did not develop pertussis (79 vs. 246, P<.0001). Corresponding values in the 687 children with no household exposure were 99, 124, and 155 U/mL, respectively (99 vs. 155, P<.0001). Thus, there is a highly significant correlation between the level of vaccine-induced serum PT IgG and protection against pertussis. PMID- 10720525 TI - Differential antibiotic-induced endotoxin release in severe melioidosis. AB - Severe melioidosis is a life-threatening, systemic bacterial infection caused by Burkholderia pseudomallei. A prospective, randomized treatment trial was conducted in northeast Thailand to compare ceftazidime (a penicillin-binding protein [PBP]-3-specific agent that causes release of large amounts of endotoxin in vitro) and imipenem (a PBP-2-specific agent that kills B. pseudomallei more rapidly but releases low amounts of endotoxin) in severe melioidosis over a 6-h time course after the first dose of antibiotic. Despite similar clinical, microbiological, endotoxin, and cytokine measures at study entry, ceftazidime treated patients (n=34) had significantly greater systemic endotoxin (P<.001) than patients treated with imipenem (n=34) after the first dose of antibiotic. No overall difference in mortality was observed (35% in both groups [95% confidence interval, 20%-50%]). Differential antibiotic-induced endotoxin release is demonstrable in severe melioidosis. These differences in endotoxin release did not appear to have a significant impact on survival in this group of patients. PMID- 10720526 TI - The oxygen- and iron-dependent sigma factor pvdS of Pseudomonas aeruginosa is an important virulence factor in experimental infective endocarditis. AB - In Pseudomonas aeruginosa, pvdS, a key oxygen (O2)-dependent locus, regulates expression of a number of virulence genes, including toxA (which encodes exotoxin A production). To define the in vivo role of differing O2 tensions on pseudomonal virulence, 2 knockout mutants, DeltapvdS and DeltatoxA, were compared with their parental strain, PA01, in rabbit aortic and tricuspid endocarditis models (representing aerobic vs. microaerobic conditions in vivo, respectively). In aortic endocarditis, DeltapvdS densities were significantly less than those of PA01 in vegetations, kidneys, and spleen (P<.01). In contrast, in tricuspid endocarditis, there were no significant differences between DeltapvdS and PA01 tissue densities in these same target tissues. The DeltatoxA mutant proliferated within target tissues to the same extent as the parental strain. Thus, pvdS (but not toxA) appears to be required for optimal virulence of P. aeruginosa, particularly in tissues preferentially exposed to high O2 tensions (e.g., aortic vegetations). PMID- 10720527 TI - Segmented filamentous bacteria prevent colonization of enteropathogenic Escherichia coli O103 in rabbits. AB - Despite their distribution in the intestines of many mammals, including man, segmented filamentous bacteria (SFB) have not been found in rabbits, nor has any function been identified for these uncultivable microbes. New Zealand White rabbits were infected with rabbit enteropathogenic Escherichia coli O103 (REPEC O103) derivatives, followed up clinically, and randomly killed 1-4 days after inoculation. Intestinal tissue samples were examined by electron and light microscopy to search for SFB and to evaluate REPEC O103 colonization. Twelve of 21 rabbits showed SFB colonization on ileal absorptive villi. The presence of SFB was correlated with lack of REPEC 0103 ileal colonization (P<.01) and disease. Rabbits without SFB were always colonized by this pathogen. SFB appear to inhibit intestinal colonization by REPEC O103 and thus protect against REPEC 0103 disease. SFB colonization in rabbits is also described for the first time. PMID- 10720528 TI - Release of gram-negative outer-membrane proteins into human serum and septic rat blood and their interactions with immunoglobulin in antiserum to Escherichia coli J5. AB - Prior studies indicate that 3 bacterial outer-membrane proteins (OMPs) are released into serum associated with lipopolysaccharide (LPS) and are bound by IgG in antiserum to Escherichia coli J5 (anti-J5 IgG). The present studies analyzed the interaction of the OMPs with anti-J5 IgG and evaluated their release in an infected burn model of gram-negative sepsis. Affinity purification studies were performed on filtrates of bacteria incubated in human serum and plasma from rats with sepsis by use of O chain-specific anti-LPS IgG and anti-J5 IgG. All 3 OMPs were captured from septic rat blood by anti-LPS IgG. Release of OMPs into serum was highest for immature bacterial cultures and was increased by antibiotics in vitro and in vivo. Anti-J5 IgG selectively captured an 18-kDa OMP released into serum and into plasma from septic rats. The results raise the possibility that anti-J5 IgG may, in part, protect via anti-OMP antibodies. PMID- 10720529 TI - Unique gonococcal phenotype associated with asymptomatic infection in men and with erroneous diagnosis of nongonococcal urethritis. AB - The percentage of gonococcal isolates in King County, Washington, requiring citrulline and uracil (CU auxotype) increased from 1.6% in 1986 to 16.5% in 1997. Among men, urethral infection with the CU auxotype (n=93), in comparison with infection by other auxotypes (n=1211), was associated with coexisting chlamydial infection, younger age, heterosexual contact, and fewer new recent partners (P<. 05). Among heterosexual men, urethral infection with the CU auxotype, compared with infection with other auxotypes, less often produced symptoms of urethral discharge (75% vs. 92%) or dysuria (47% vs. 74%) or signs of moderate or profuse urethral discharge (57% vs. 89%, P<.05 for each comparison), produced symptoms of longer duration (7. 0 vs. 4.5 days, P<.01), less often resulted in urethral smears showing gram-negative intracellular diplococci (67% vs. 95%, P<.01), and thus more often was erroneously diagnosed as nongonococcal urethritis. Several mechanisms could explain reduced inflammatory response to the CU auxotype and its recent spread. PMID- 10720530 TI - An isogenic hemoglobin receptor-deficient mutant of Haemophilus ducreyi is attenuated in the human model of experimental infection. AB - Haemophilus ducreyi expresses a conserved hemoglobin-binding outer-membrane protein (HgbA). To test the role of HgbA in pathogenesis, we infected 9 adults with isolate 35000 and its isogenic hgbA-inactivated mutant (FX504) on their upper arms in a double-blinded, escalating dose-response study. Papules developed at similar rates at sites inoculated with the mutant or parent. The pustule formation rate was 55% (95% confidence interval [CI], 30. 8%-78.5%) at parent sites and 0 (95% CI, 0-10.5%) at mutant sites (P<.0001). The recovery rate of H. ducreyi from surface cultures was 16% (n=142) from parent sites and 0 (n=213) from mutant sites (P<. 0001). H. ducreyi was recovered at biopsy from 6 of 7 parent sites and from 0 of 3 mutant sites. The results indicate that hemoglobin may be a critical source of heme or iron for the establishment of H. ducreyi infection in humans. PMID- 10720531 TI - Endotoxin down-regulates monocyte and granulocyte interleukin-6 receptors without influencing gp130 expression in humans. AB - Interleukin (IL)-6 is important for host defense against various pathogens. The IL-6 receptor (IL-6R) complex consists of a ligand-binding component (IL-6R) and a signal-transducing component (gp130). In a study designed to obtain insight into the regulation of this receptor complex during inflammation, 8 healthy subjects received an intravenous injection of lipopolysaccharide (LPS; 4 ng/kg), and receptor expression was determined on blood leukocytes by use of fluorescence activated cell cytometry. LPS induced a transient decrease in monocyte and granulocyte IL-6R expression but did not influence gp130. The plasma concentrations of soluble IL-6R and soluble gp130 did not change after LPS administration. Expression of the receptor for leukemia inhibitory factor, a member of the IL-6R family, remained unaltered after LPS injection. In whole blood in vitro, LPS and gram-positive stimuli and proinflammatory cytokines were capable of down-modulating the IL-6R. Monocytes and granulocytes may down regulate IL-6R at their surface upon their first interaction with bacterial antigens. PMID- 10720532 TI - Growth-inhibiting antibody responses of humans vaccinated with recombinant outer surface protein A or infected with Borrelia burgdorferi or both. AB - Serial serum samples from a 2-year human trial of outer surface protein (Osp) A vaccine were analyzed by Borrelia burgdorferi growth-inhibition assay (GIA) and anti-OspA ELISA to assess the antibody responses of vaccine recipients and subjects with Lyme disease. Although 74% of OspA recipients had a reciprocal GIA titer >/=64 after 3 vaccinations, none of the placebo recipients, even those with Lyme disease, had a GIA titer this high. The correlation between GIA and ELISA titers after 3 doses of vaccine was.84; however, more vaccine recipients had an elevated ELISA titer paired with low GIA titer than had a low ELISA titer with a high GIA titer. OspA-vaccine recipients who acquired Lyme disease had significantly lower serum GIA and ELISA titers after 3 immunizations than did age and sex-matched OspA recipients without Lyme disease. Thus, vaccinated subjects had antibodies to native antigen on viable cells, and antibody assays with this specificity may predict protection of vaccinees against infection. PMID- 10720533 TI - Status of Borrelia burgdorferi infection after antibiotic treatment and the effects of corticosteroids: An experimental study. AB - Sixteen specific-pathogen-free beagles were infected with Borrelia burgdorferi. Three groups of 4 dogs were treated with antibiotics for 30 consecutive days starting 120 days after tick exposure; 4 dogs were untreated controls. At day 420 after tick exposure and again before euthanasia, 2 dogs of each group were treated with prednisone for 14 days. All dogs contracted infection and 11 developed acute arthritis 50-120 days after exposure. After day 120, one of 12 antibiotic-treated dogs and 2 of 4 untreated dogs became lame. Antibiotic therapy reduced the frequency of Borrelia-positivity in subsequent skin biopsy samples. After prednisone treatment, both control dogs developed severe polyarthritis. At euthanasia, single tissues of the antibiotic-treated dogs and multiple tissues of all control dogs were Borrelia-positive by polymerase chain reaction. Viable spirochetes were not recovered from antibiotic-treated dogs. Two antibiotic treated dogs showed histologic evidence of minimal lesions, whereas all control dogs had mild polyarthritis with periarteritis. PMID- 10720534 TI - Primary isolation of Ehrlichia chaffeensis from patients with febrile illnesses: clinical and molecular characteristics. AB - Ehrlichia chaffeensis was sought among patients with a history of tick exposure and fever, and the accuracy of other diagnostic tests was compared with that of primary isolation. Among the 38 patients enrolled, E. chaffeensis was isolated from the blood of 7 (18%) and from cerebrospinal fluid specimens of 2 of these 7. All 7 patients also were positive by polymerase chain reaction (PCR) of blood, and 6 patients developed diagnostic titers of antibody to E. chaffeensis. The isolates were characterized by molecular analysis of the 16S rRNA gene, the 120 kDa protein gene, and the variable-length PCR target (VLPT) of E. chaffeensis. On the basis of the 120-kDa and VLPT genotypes, the cerebrospinal fluid and blood isolates from the same patients were identical. This study demonstrates that both PCR and culture of blood for E. chaffeensis have high diagnostic yields. More frequent isolation of E. chaffeensis from patients with infection should further our understanding of the pathogenesis of this infection. PMID- 10720535 TI - Lewis antigen expression and stability in Helicobacter pylori isolated from serial gastric biopsies. AB - The expression of Lewis antigens by the gastric pathogen Helicobacter pylori in serial biopsy isolates was investigated to assess antigen distribution and stability. A total of 26 asymptomatic subjects were given various doses of 3' sialyllactose for up to 56 days. Gastric biopsies were performed during the dosing period, as well as 30 days after dosing, which provided 127 H. pylori isolates that were examined by use of ELISA and immunoblot. A large proportion of subjects (14/26) yielded sequential H. pylori isolates, which appeared genetically identical but had variable Lewis antigen expression. The proportion of subjects with H. pylori isolates not expressing any Lewis antigens was greater than that previously reported (10/26). Thus, the expression of the Lewis antigens by H. pylori does not appear to be a requirement for colonization, whereas the antigen expression after human infection is more variable than the previously reported rate observed with in vitro cultures. PMID- 10720536 TI - Monocyte-endothelial cell coculture enhances infection of endothelial cells with Chlamydia pneumoniae. AB - To determine the mechanism(s) by which Chlamydia pneumoniae homes to and establishes persistent infection in atheromatous lesions, the effect of the interaction of monocytes/macrophages (U937 cells) with human umbilical vein endothelial cells (HUVECs) and transformed human arterial endothelial cells (HMEC 1s) on the susceptibility of endothelial cells to infection with C. pneumoniae was investigated. Infection was enhanced 4.7-fold (HUVECs) and 4.4-fold (HMEC-1s) after coculture at monocyte-to-endothelial cell ratios of 5 and 2.5, respectively. U937 cells also directly transmitted infection to the endothelial cells, and addition of U937 cell-conditioned media dose-dependently enhanced the infectivity 2.0- to 2.5-fold. The stimulation of infectivity was specific to endothelial cells, because coculturing of monocytes with epithelial cells did not enhance the susceptibility of epithelial cells to infection. The susceptibility of endothelial cells to infection with C. trachomatis and C. psittaci was not enhanced by the monocyte-derived factor(s). PMID- 10720538 TI - Attenuation of Trypanosoma brucei is associated with reduced immunosuppression and concomitant production of Th2 lymphokines. AB - Mechanisms regulating resistance to African trypanosomes were addressed by comparing the immune responses of mice infected with attenuated Trypanosoma brucei brucei lacking the phospholipase C gene (PLC-/-) and those of mice infected with wild-type (WT) parasites. Inhibition of concanavalin A (ConA) induced T cell proliferation occurred in spleen and lymph nodes of PLC-/-- and WT infected mice. Although suppressive cells were elicited in spleen and lymph nodes of WT-infected animals, such cells were not detected in lymph nodes of PLC-/- infected mice. PLC-/--infected mice had more interleukin-4 and -10 in their blood than did WT-infected mice. Correspondingly, PLC-/--infected mice had higher IgG1 antibody levels against variant surface glycoprotein than did WT-infected mice. These data indicate that attenuation of T. b. brucei correlates with the absence of cells suppressing ConA-induced T cell proliferation in the lymph nodes, with increased production of Th2 cytokines and a stronger IgG1 antibody response to trypanosome antigens. PMID- 10720537 TI - Ulinastatin, an elastase inhibitor, inhibits the increased mRNA expression of prostaglandin H2 synthase-type 2 in Kawasaki disease. AB - Kawasaki disease is an inflammatory disease of unknown cause that causes panvasculitis, including coronary arteritis. Polymorphonucleocytosis in the early stage of the illness suggests the implication of neutrophils in the pathogenesis of the disease. In the acute phase of Kawasaki disease, mRNA expression of prostaglandin H2 synthase (PHS)-2, as determined by reverse transcription polymerase chain reaction, was markedly enhanced, and thromboxane A2 (TXA2) synthesizing activity was increased in polymorphonuclear leukocytes (PMNL). This up-regulation of PHS-2 was suppressed by ulinastatin (a neutrophil-elastase inhibitor) treatment. Lipopolysaccharide-induced enhancement of PHS-2 mRNA was also inhibited by therapeutic doses of ulinastatin in vitro by use of PMNL from healthy volunteers. Thus, ulinastatin inhibits arachidonate PHS metabolism by inhibiting new induction of PHS-2 at the mRNA level, which is a novel pharmacologic action of this substance. Ulinastatin treatment is possibly an additional therapeutic approach to Kawasaki disease. PMID- 10720539 TI - Inhibition of interferon-gamma signaling by Leishmania donovani. AB - Leishmania infection causes marked down-regulation of interferon (IFN)-gamma induced gene activity in macrophages, but the mechanism of the blockade has not been fully defined. The IFN-gamma signal transduction pathway was analyzed in Leishmania donovani-infected phorbol-differentiated U937 human promonocytic cells. IFN-gamma stimulation induced marked phosphorylation of its own receptor (IFN-gammaR)-alpha chain. Phosphorylation of the receptor subunit was significantly inhibited after 24 h of infection with the parasite, apparently because of decreased amounts of the receptor subunit. Formation of the IFN-gammaR complex, as assessed by tyrosine phosphorylation and association of Jak2, was strongly inhibited in cells infected for 24 h. Inhibition of the IFN-gammaR complex formation correlated with inhibition of STAT1alpha binding to the IFN gamma response region. Pretreatment with purified parasite lipophosphoglycan before IFN-gamma stimulation had no effect on tyrosine phosphorylation. Thus, inhibition of tyrosine phosphorylation of the IFN-gammaR-alpha chain and subsequent signal transduction are most likely due to the decreased amount of IFN gammaR-alpha protein after infection. PMID- 10720540 TI - Safety and immunogenicity of intranasally administered inactivated trivalent virosome-formulated influenza vaccine containing Escherichia coli heat-labile toxin as a mucosal adjuvant. AB - A trivalent influenza virosome vaccine containing hemagglutinin and Escherichia coli heat-labile toxin (HLT) was administered intranasally to young adults and elderly subjects. Symptoms that followed immunization were mild and transient. A significant increase in serum hemagglutination inhibition (HI) antibody was noted for the 3 vaccine strains. There was no significant difference in postimmunization geometric mean titers or seroconversion rates between age groups. The percentage of subjects attaining protective HI titers (>/=40%) was comparable in both groups for the A/Bayern (P=.5) and B/Beijing (P=.3) strains but was higher among young adults (92.2%) versus elderly subjects (76.5%; P=.057) for the A/Wuhan strain. The proportion of subjects with nonprotective baseline titers who attained protective levels after immunization was similar in both age groups for the A/Bayern and B/Beijing components. For the A/Wuhan component, significantly (P=.017) more young adults achieved protective titers versus elderly subjects (85. 7% and 53.8%, respectively). Vaccination evoked a significant (P<. 005) increase in anti-HLT antibody titers. PMID- 10720542 TI - Effect of TT virus infection on hepatocellular carcinoma development: results of a Euro-Asian survey. AB - A small percentage of persons with hepatocellular carcinoma (HCC) lack identifiable causes of liver pathology. The single-stranded DNA virus, TT virus (TTV), has been found in persons with acute and chronic liver injury. Nested polymerase chain reaction was used to search for both TTV and parvoviruses in 293 HCC samples from Asia and Europe. TTV was found in >30% of Chinese and Italian samples but in only 13% of French samples. No clinicopathologic differences were found between TTV-positive and -negative populations. A significant association was found between TTV infection and hepatitis B virus (P<.01) and herpesviruses (P<.02) in HCC patients, suggesting that factors promoting these infections are associated with enhanced TTV positivity. Parvovirus B19 and adeno-associated virus were found in only 7.5% of the tumors. Taken together, these data suggest that TTV infection is unlikely to be associated with the induction or acceleration of the hepatocarcinogenic process in humans. PMID- 10720541 TI - Correlates of immune protection induced by live, attenuated, cold-adapted, trivalent, intranasal influenza virus vaccine. AB - The authors conducted a 2-year, multicenter, double-blind, placebo-controlled efficacy field trial of live, attenuated, cold-adapted, trivalent influenza vaccine administered by nasal spray to children 15-71 months old. Overall, vaccine was 92% efficacious at preventing culture-confirmed infection by influenza A/H3N2 and influenza B. Because influenza A/H1N1 did not cause disease during the years in which this study was conducted, the authors sought to determine vaccine efficacy and correlates of immune protection against experimental challenge with 107 TCID50 of attenuated H1N1 (vaccine strain) by intranasal spray. Prechallenge assessments included serum hemaglutination inhibiting (HAI) antibody and nasal wash IgA antibody to H1N1. Vaccine was 83% efficacious (95% confidence interval, 60%-93%) at preventing shedding of H1N1 virus after challenge. Any serum HAI antibody or any nasal wash IgA antibody was correlated with significant protection from H1N1 infection as indicated by vaccine-virus shedding, and high efficacy against H1N1 challenge was demonstrated. PMID- 10720543 TI - Resistance profiles in patients with viral rebound on potent antiretroviral therapy. AB - The prevalence of phenotypic drug resistance was assessed in 60 patients with a viral rebound after they received a protease inhibitor (PI)- or nonnucleoside reverse transcriptase inhibitor (NNRTI)-containing regimen (baseline). Resistance testing was done within 36 weeks of viral rebound; no resistance testing was available at baseline. All patients had previously received zidovudine; 86.0% had received lamivudine. In total, 45.1% of the patients had strains resistant to the PI that they started and 88.9% given nevirapine had strains with reduced susceptibility to that drug. Overall, 46 patients (76.7%) harbored a strain resistant to >/=1 drug of their initial PI- or NNRTI-containing regimen. Of 53 patients who remained on treatment at the time of the study (40 had switched to a different combination from that at baseline), 6 harbored isolates susceptible to all drugs they had ever received. Thus, patients with viral rebound while on potent antiretroviral therapy usually have reduced susceptibility to >/=1 drug. Viral rebound also occurs in persons in whom resistant strains could not be detected by the assay used. PMID- 10720544 TI - Granulocyte colony-stimulating factor increases CD4+ T cell counts of human immunodeficiency virus-infected patients receiving stable, highly active antiretroviral therapy: results from a randomized, placebo-controlled trial. AB - Thirty human immunodeficiency virus (HIV)-infected patients with CD4+ T cell counts <350 cells/mm3 who had received stable, highly active antiretroviral therapy (HAART) for at least 24 weeks were randomized to receive either placebo or granulocyte colony-stimulating factor (G-CSF; 0.3 mg/mL 3 times a week) for 12 weeks. Blood samples were collected at specified time points. G-CSF treatment enhanced the total lymphocyte count (P=.002) and increased CD3+ (P=.005), CD4+ (P=.03), and CD8+ (P=.004) T cell counts as well as numbers of CD3-CD16+CD56+ NK cells (P=.001). The increases in CD4+ and CD8+ cell counts resulted from increases in CD45RO+ memory T cells and cells expressing the CD38 activation marker. Lymphocyte proliferative responses to phytohemagglutinin and Candida antigen decreased, whereas NK cell activity and plasma HIV RNA did not change during G-CSF treatment. After 24 weeks, all immune parameters had returned to baseline values. This study suggests that G-CSF treatment of HIV-infected patients receiving stable HAART increases the concentration of CD4+, CD8+, and NK cells without inducing changes in the virus load. PMID- 10720545 TI - Nucleotide sequences that distinguish Oka vaccine from parental Oka and other varicella-zoster virus isolates. AB - The sequences of approximately 34 kb from the 3' end of the varicella-zoster virus (VZV) Oka vaccine strain and the previously sequenced Dumas strain were compared. Sequence differences were noted in the coding sequences of several VZV open reading frames (ORFs), including ORFs 48, 51, 52, 55, 56, 58, 59, 60, 62, 64, and 68. Tests based on differences in the ORF62 gene and in the ORF64 poly-A region successfully distinguished the Oka vaccine strain from its wild-type parent and from other Japanese and US clinical isolates. These changes remained stable after passage of the virus in humans. PMID- 10720546 TI - Novel immunogenicity of Oka varicella vaccine vector expressing hepatitis B surface antigen. AB - Recombinant Oka (ROka) varicella vaccine expressing hepatitis B surface antigen (HBsAg) and subunit HBsAg vaccine (SHV) were used as primary and booster HBsAg vaccines in 3 combinations (SHV-SHV, SHV-ROka, and ROka-SHV) in guinea pigs. Immune responses to HBsAg and varicella-zoster virus gE:gI were evaluated. The 3 combinations induced similar levels of the lymphocyte proliferation response to HBsAg. Of the 3 combinations, SHV-SHV induced the strongest antibody response to an "a" loop of HBsAg and to the whole HBsAg. Its ratio of antibody titer to this loop versus HBsAg was significantly higher than that in SHV-ROka, suggesting the supplementary recognition of the conformational epitope of HBsAg in SHV-ROka. SHV ROka induced delayed-type hypersensitivity (DTH) to the HBsAg and gE:gI produced in infected cells. Thus, ROka induced DTH to HBsAg and enhanced recognition of the conformational epitope. ROka varicella vaccine may serve as a novel vaccine vector to induce a Th1-type immune response. PMID- 10720547 TI - Comparison of an opsonophagocytic assay and IgG ELISA to assess responses to pneumococcal polysaccharide and pneumococcal conjugate vaccines in children and young adults with sickle cell disease. AB - Children with sickle cell disease were immunized with either 2 doses of 7-valent pneumococcal conjugate vaccine followed by 1 dose of 23-valent pneumococcal polysaccharide vaccine or a single dose of 23-valent vaccine. Functional antibodies to 7 vaccine serotypes were measured by a flow cytometric opsonophagocytic assay (OPA) and compared with IgG anticapsular polysaccharide antibody concentrations measured by ELISA. Moderate correlations were found between OPA and ELISA antibody titers for all 7 serotypes (r values, 0.41-0.70; P<.001 for all serotypes). After immunization with 23-valent vaccine, geometric mean antibody titers by OPA were significantly higher in the combined schedule group for 5 of 7 vaccine serotypes but were significantly higher for only 2 of 7 serotypes as measured by ELISA. The ability of OPA to show a greater differential response to the 2 immunization schedules used in this study suggests that it may be useful in the evaluation of immunization regimens involving pneumococcal conjugate vaccines. PMID- 10720548 TI - Interferon-gamma protects against biomaterial-associated Staphylococcus epidermidis infection in mice. AB - Survival of Staphylococcus epidermidis inside macrophages has been recognized as a pivotal process in the pathogenesis of biomaterial-associated infection (BAI). Interferon (IFN)-gamma is a potent activator of macrophages. This study examined whether subcutaneous injections of IFN-gamma can reverse macrophage deactivation induced by implanted biomaterials. Mice received subcutaneous implants combined with an injection of 106 S. epidermidis to induce an experimental BAI. Subsequently, 3 groups of mice received subcutaneous injections of 25,000 IU IFN gamma 3 times weekly, 10,000 IU IFN-gamma 3 times in 2 weeks, or saline 3 times weekly (saline control), respectively. A fourth group received no injections (control). Segments and tissues of the IFN-gamma-treated mice were significantly less (P<.05) culture positive than those of the control groups. Histologically, the high numbers of intracellularly persisting gram-positive cocci observed in the control mice were absent in the IFN-gamma-treated mice. These data indicate that IFN-gamma protects against experimental BAI. PMID- 10720549 TI - Lack of immunity in university students before an outbreak of serogroup C meningococcal infection. AB - Immunity to meningococci was determined in infected and uninfected students before and during an outbreak of serogroup C meningococcal infection at a university in the United Kingdom. No immunity against the outbreak strain was detected in serum taken from infected students prior to the outbreak or at the time of admission; bactericidal activity developed during convalescence. Carriage of all strains of serogroup C meningococci in asymptomatic students was low (0.9%), and no carriage of the outbreak strain could be detected. Immunity in the at-risk student population before the outbreak was low: 90% of students had no significant bactericidal activity against the outbreak strain. A low prevalence of carriage of the outbreak strain, together with a low prevalence of protective immunity within the student population, was associated with a high incidence of invasive disease in those who acquired the outbreak strain. PMID- 10720550 TI - A pilus-deficient mutant of Haemophilus ducreyi is virulent in the human model of experimental infection. AB - Haemophilus ducreyi expresses fine tangled pili, which are composed predominantly of a major subunit (FtpA). Confocal microscopy showed that an FtpA-specific monoclonal antibody bound to bacteria in biopsy samples obtained from infected human volunteers. To test the role of pili in pathogenesis, an isogenic mutant (35000HP-SMS1) was constructed by insertionally inactivating ftpA. 35000HP-SMS1 did not express FtpA and was nonpiliated but was otherwise identical to its parent, 35000HP. Seven healthy adults were challenged on the upper arm with the isogenic isolates in a double-blinded, escalating dose-response study. Sites inoculated with the mutant produced papules and pustules at rates similar to the rates observed at sites inoculated with the parent. The recovery rate of H. ducreyi from cultures and the histopathology of biopsy samples obtained from pustules inoculated with 35000HP or 35000HP-SMS1 were similar. Although pili are expressed in vivo, FtpA is not required for pustule formation in the human challenge model. PMID- 10720551 TI - Aminoglycoside-resistance mechanisms for cystic fibrosis Pseudomonas aeruginosa isolates are unchanged by long-term, intermittent, inhaled tobramycin treatment. AB - Aminoglycoside-resistance mechanisms were characterized in Pseudomonas aeruginosa isolates from cystic fibrosis (CF) patients during a recent clinical trial of inhaled tobramycin. Impermeability, in which bacteria have reduced susceptibility to all aminoglycosides, was the predominant mode of resistance in isolates obtained both before and after 6 months of cyclic treatment with tobramycin or placebo administered by aerosol. Enzymatic resistance mechanisms were found in fewer than 10% of resistant isolates. P. aeruginosa from individual patients could be grouped on the basis of genetic relatedness. When enzymatic resistance was involved, all isolates in a group had elevated tobramycin MICs. When impermeability occurred, MICs of a genotypic group varied from susceptible to resistant. These findings suggest that impermeability resistance occurs in only a fraction of the P. aeruginosa population in lungs of persons with CF and that this form of resistance arises by a process involving multiple small changes in MIC. PMID- 10720552 TI - Soluble CD14 levels in the serum, synovial fluid, and cerebrospinal fluid of patients with various stages of Lyme disease. AB - Levels of circulating soluble CD14 (sCD14) in patients with various stages of Lyme disease (LD) were examined. Patients with early or untreated late LD had significantly higher levels of sCD14 than did healthy controls (P=.0001 and .0007, respectively); levels returned to normal within 3 months after antibiotic therapy. Patients with persistent posttreatment symptoms of LD had sCD14 levels equivalent to those of healthy controls. Differences in the serum sCD14 levels in patients with various stages of LD are likely to be directly correlated with differences in bacterial burden, suggesting that posttreatment symptoms may not require continued presence of the organism. sCD14 levels in the cerebrospinal fluid (CSF) of patients with any stage of LD were no different from those of control subjects. Levels of synovial fluid sCD14 from patients with Borrelia burgdorferi in their joints were elevated, compared with levels in normal serum, and may play a role in the pathogenesis of arthritis. PMID- 10720553 TI - Interaction of Mycobacterium avium complex with human respiratory epithelial cells. AB - Adherence of Mycobacterium avium complex (MAC) to human respiratory epithelial cells (HEp-2) induced 2 distinct modes of internalization. In the first, MAC induced ruffling of HEp-2 cell membrane and formation of surface projections securing the bacilli on the surface, and concurrent membrane depressions, beneath the sites of attachment of bacilli, resulted in internalization of the organisms. The second mode involved formation of membrane folds wrapping around the bacilli, followed by internalization. Two MAC proteins of approximately 31 kD and approximately 25 kD, respectively, were identified that mediated these interactions specific for HEp-2 cells. The N-terminal amino acid sequence of the 31-kD MAC protein displayed homology with the 21-kD hypothetical protein of Mycobacterium tuberculosis, and the 25-kD MAC protein showed homology with Mn superoxide dismutase of MAC and Mycobacterium leprae. These 2 HEp-2 cell-specific MAC proteins may be involved in the interaction of MAC with epithelial cells. PMID- 10720554 TI - Increased production of interleukin 4 by CD4+ and CD8+ T cells from patients with tuberculosis is related to the presence of pulmonary cavities. AB - In tuberculosis, cellular immunity is considered to be responsible for the eradication of infection but also for damage of host tissues. In animal models, the balance between Th1-type cytokines, especially interferon (IFN)-gamma, and Th2-type cytokines, primarily interleukin (IL)-4, seems crucial for these effects. Reports on Th1-type and Th2-type cytokines in human tuberculosis are conflicting, and little is known about their role in tissue damage. Flow cytometric assessment of cytokine responses was performed in human immunodeficiency virus (HIV)-seronegative patients with active tuberculosis and in healthy controls. Patients and controls showed no significant difference in expression of IFN-gamma. However, patients showed a striking increase in production of IL-4 in CD4+ as well as CD8+ T cells. Most remarkably, the expression of IL-4 was especially elevated in patients with cavitary tuberculosis. The Th2-type response with increased production of IL-4 in patients with tuberculosis may antagonize host defense and lead to tissue necrosis. PMID- 10720555 TI - Fungal endophthalmitis diagnosis by detection of Candida albicans DNA in intraocular fluid by use of a species-specific polymerase chain reaction assay. AB - Candida endophthalmitis is a serious infection secondary to hematogenous dissemination or direct inoculation of the organisms following trauma or eye surgery. The diagnosis is based on the characteristic findings in the infected eye and on culture of vitreous samples. Unfortunately, the yield of vitreous cultures is limited. The use of a Candida albicans species-specific polymerase chain reaction (PCR) assay in the diagnosis of Candida endophthalmitis is reported herein. Four patients with suspected fungal endophthalmitis underwent vitrectomy for diagnostic and therapeutic purposes. In 2 of the 4, vitreous cultures were negative. However, characteristic PCR products were generated in all 4 patient specimens, enabling the rapid diagnosis of Candida endophthalmitis in all 4. Clinical response was observed in all cases. These results demonstrate the utility of PCR-mediated detection of C. albicans in vitreous samples. PMID- 10720556 TI - Naturally acquired antibodies to the glutamate-rich protein are associated with protection against Plasmodium falciparum malaria. AB - The development of effective malaria vaccines depends on the identification of targets of well-defined protective immune responses. Data and samples from a longitudinal study of a cohort of children from coastal Ghana were used to investigate the role of antibody responses to 3 regions of the Plasmodium falciparum glutamate-rich protein (GLURP). The data show that levels of the GLURP specific IgG that occurs in the nonrepeat region of the antigen are significantly correlated with clinical protection from P. falciparum malaria, after correction for the confounding effect of age. Furthermore, levels of cytophilic antibodies were found to be of particular importance for protection, lending support to the hypothesis that antibody-dependent cellular inhibition is the important element in GLURP-specific protective immunity. PMID- 10720557 TI - Parasite multiplication potential and the severity of Falciparum malaria. AB - The multiplication rates and invasiveness of Plasmodium falciparum parasites isolated from adult Thai patients hospitalized with uncomplicated malaria (n=34) were compared with those from persons with severe malaria (n=42). To simulate severe malaria and control for host effects, the in vitro cultures were adjusted to 1% parasitemia and used the same red blood cell donor. P. falciparum isolates from persons with severe malaria had initial cycle multiplication rates in vitro that were 3-fold higher than those from uncomplicated malaria (median [95% confidence interval], 8.3 [7. 1-10.5] vs. 2.8 [1.7-3.9]; P=.001). Parasites causing severe malaria exhibited unrestricted red blood cell invasion, whereas those from uncomplicated malaria were restricted to a geometric mean of 40 (31% 53%) of red blood cells. P. falciparum parasites causing severe malaria were less selective and multiplied more at high parasitemias than those causing uncomplicated malaria. PMID- 10720558 TI - Female genital schistosomiasis of the lower genital tract: prevalence and disease associated morbidity in northern Tanzania. AB - Female genital schistosomiasis (FGS) is a neglected disease manifestation of schistosomiasis. A cross-sectional study was carried out to assess in a schistosomiasis-endemic area the proportion of women affected by FGS of the lower reproductive tract and to compare the frequency of symptoms and signs possibly associated with FGS between women with proven FGS (n=134), endemic referents (n=225, women living in an endemic site), and referents (n=75, women living in a nonendemic site). Urinary schistosomiasis was diagnosed in 36% (239/657) and FGS in 37% (134/359) of the women. Cervical lesions occurred in 75% of the FGS cases, in 48% of endemic referents, and in 36% of nonendemic referents. The high prevalence of FGS in all age groups and the high levels of pathologic cervical alterations such as swollen and disrupted epithelium support the hypothesis that FGS might be a risk factor for the transmission of human immunodeficiency virus. PMID- 10720559 TI - Effect of plasma human immunodeficiency virus type 1 RNA levels on neutrophil oxidative burst. PMID- 10720560 TI - Reply. PMID- 10720561 TI - Clearance of cytomegalovirus viremia after initiation of highly active antiretroviral therapy. PMID- 10720562 TI - Reply. PMID- 10720563 TI - Long-term results of laser treatment for retinal angiomatosis in von Hippel Lindau disease. AB - PURPOSE: In von Hippel-Lindau disease (VHL), retinal detachment is often a real threat if the retinal angiomas are left untreated. The aim of this trial was to diagnose and treat capillary angiomas early at an asymptomatic stage. METHODS: Since 1983, we have investigated patients with key lesions and those who are at risk for VHL in a multidisciplinary study. Classical retinal angiomas and microangiomas were treated with the Argon laser. Checks during a follow-up of 15 years have provided us with long-term results. RESULTS: We detected 189 retinal angiomas in 97 eyes of 69 patients (105 classical angiomas, including 19 fibrotic angiomas in 15 eyes, 84 microangiomas). During the study, 20 patients (twenty-one symptomatic eyes) entered the study already with a visual impairment. Sixty asymptomatic eyes from 49 patients with a total of 108 retinal angiomas (55 microangiomas and 53 classical angiomas) were treated by us with photocoagulation. Nineteen asymptomatic eyes with angiomas of fibrotic type were observed only. During a mean follow-up of 5.1 years (range: 0.25 - 15; SD: 3.88) observed after treatment, no deterioration of vision occurred in 37 patients treated with photocoagulation. Regular controls showed 5 relapsing angiomas and 9 new angiomas which were successfully coagulated again. New angiomas occurred in those eyes with several angiomas as a predisposition. Patients with a tendency to multiple angiomas need to be seen more frequently because these eyes have a high risk of becoming blind, even in already coagulated angiomas. CONCLUSION: Laser treatment was successful. Early ophthalmological examination of patients with retinal angiomatosis is a great challenge, since these lesions can only be successfully treated at an early stage. Treatment is then effective and safe, but long-term follow-up is necessary. New angiomas occurred in those eyes with several angiomas as a predisposition. PMID- 10720564 TI - Therapeutic drug monitoring of saquinavir in patients during protease inhibitor therapy with saquinavir alone or in combination with ritonavir or nelfinavir. AB - Therapeutic drug monitoring is essential in HIV-patients undergoing highly active antiretroviral therapy (HAART). Saquinavir (SQV) is used alone or in combination with ritonavir (RTV) or nelfinavir (NLF), respectively, in the context of the HAART drug regimen. The achievable SQV concentration range in clinical practice remains to be elucidated. A non-randomized prospecitve clinical trial 19 patients (group I) receiving SQV (1x600 mg/d Invirase or Fortovase), 29 patients (group II) receiving SQV (2x600 mg/d Fortovase) plus RTV (2x400 mg/d Norvir), and 21 patients (group III) receiving SQV (2x600 mg/d Fortovase) plus NLF (2x750 mg/d Viracept) was conducted to determine SQV plasma concentrations. SQV levels were determined as trough levels during routine outpatient visits. Analysis was performed by HPLC with UV detection. The lowest SQV plasma levels were found in group I (95% CI 89-177 ng/ml). Significantly higher SQV levels were found in group III (combination with NLF) ranging from 242 to 398 ng/ml (95% CI) and in group II (combination with RTV) ranging from 1354 to 1747 ng/ml (95% CI). The IC 50% of 54 ng/ml was not reached in at least one sample during the study (mean duration of study 16+/-10 months) in 14/19 patients of group I, 9/29 patients in group II and 13/21 patients in group III, respectively. A positive correlation between patient compliance, defined by SQV levels in the 95% CI of the used combination, and the HIV RNA plasma level was found. The presented data confirm that therapy with SQV alone may not be effective, since trough levels are near the lower limit of antiretroviral efficacy. Although the combination of SQV with NLF results in higher SQV plasma concentrations in a bid regimen, in more than 60% of the patients SQV concentrations below IC 50 level were detected during the twelve-months study period. The combination of SQV with RTV yields the highest SQV-trough levels. SQV concentrations below the IC 50 were seen in only 31% of patients with the SQV/RTV combination. In conclusion, therapeutic drug monitoring allows an efficient surveillance of patients compliance. In addition, therapeutic drug monitoring represents a valuable tool for management of HAART in patients receiving a complex comedication or suffering from advanced liver disease. PMID- 10720565 TI - Serum leptin levels and body weight in postmenopausal women under transdermal hormone replacement therapy. AB - Leptin, the adipocyte-specific product of the ob gene, is implicated in body weight regulation and energy balance. We investigated the influence of hormone replacement therapy (HRT) on the body mass index (BMI) and serum leptin levels in 20 postmenopausal, nonobese women treated with transdermal HRT (delivery rate 50 microg 17beta-estradiol/24 h, 1 patch per week) for 6 months. Serum leptin levels were measured by ELISA and results were compared by means of the Student's paired t-test or Pearson's correlation. The mean patient age was 55+/-6.04 years. The mean body weight prior to the start of the study was 69.39+/-9.37 kg, and the BMI before HRT was 26.92+/-4.47 kg/m2. Both parameters remained unchanged under therapy. No significant change in absolute serum leptin values (18.8+/-8.4 ng/ml; 20.47+/-9.7 ng/ml; 17.92+/-8.7 ng/ml at 0, 4 and 6 months respectively) or in adiposity-corrected values (serum leptin/BMI) (0.68+/-0.24; 0.75+/-0.29; 0.67+/ 0.26 at 0, 4 and 6 months respectively) were found. Serum leptin levels correlated well with BMI (r = 0.7193, p<0.0001). There was no significant correlation of estradiol with serum leptin levels before or during therapy. In summary, low dose, transdermal HRT exhibited no influence on serum leptin levels or BMI in postmenopausal women. These data suggest that low dose HRT does not influence body weight regulation in postmenopausal women. PMID- 10720566 TI - 1st European Symposium on Liquid Ventilation - Co-organized by Unidad Neonatal, Hospital de Cruces, Basque Country University Medical School, Bilbao, Spain and Clinic of Neonatology CCM, Charite Humboldt-University Berlin, Germany - Scientific Committee: Roland R. Wauer Berlin, Germany and Adolf Valls i Soler Bilbao, Spain - November 26-27, 1999, Charite, Berlin, Germany. AB - Addressing the most recent advances and research results in the understanding and state of the art on LIQUID VENTILATION, we learned, that not only pediatricians and neonatologists, but also anesthesiologists, intensivists, biologists, chemists, pharmacologists and biophysicians were interested. Furthermore, we were able to integrate representatives of the pharmaceutic industry interested in future developments in this exciting field. - Our main goal both was and is to provide an opportunity for all European researchers in this field to meet and get to know each other. Thus we provided both a scientific and a friendly atmosphere to foster the exchange of experiences and to facilitate future joint projects. - Our long term aim is to harmonize efforts to accelerate the different research steps to close the time gap between research and its future clinical application. We hope this meeting will be a starting point for collaboration of clinical groups in Europe, to study all research aspects of the technique to carry on future trials. Therefore we are planning to initiate a WWW webside. - There are still a lot of unanswered questions, regarding the physicochemical properties, biological effects and possible indications to use Perfluorocarbons for liquid ventilation. We are aware that none of the open questions could be answered completely, but hopefully collaboration and communication will bring us closer to achieve these goals. Moreover, concerted actions should be started to search for research grant founding. PMID- 10720567 TI - Unlimited hypothesis research. PMID- 10720568 TI - Methodological function of hypotheses in science: old ideas in new cloth. PMID- 10720569 TI - A phenotype map of the mouse X chromosome: models for human X-linked disease. AB - The identification of many of the transcribed genes in man and mouse is being achieved by large scale sequencing of expressed sequence tags (ESTs). Attention is now being turned to elucidating gene function and many laboratories are looking to the mouse as a model system for this phase of the genome project. Mouse mutants have long been used as a means of investigating gene function and disease pathogenesis, and recently, several large mutagenesis programs have been initiated to fulfill the burgeoning demand of functional genomics research. Nevertheless, there is a substantial existing mouse mutant resource that can be used immediately. This review summarizes the available information about the loci encoding X-linked phenotypic mutants and variants, including 40 classical mutants and 40 that have arisen from gene targeting. PMID- 10720570 TI - Genomic organization of the dog dystroglycan gene DAG1 locus on chromosome 20q15.1-q15.2. AB - Dystroglycan is a laminin binding protein, which provides a structural link between the subsarcolemmal cytoskeleton and the extracellular matrix. It is also involved in the organization of basement membranes. So far the genomic organization of the dystroglycan gene DAG1 has not been completely investigated. Here we report the cloning and sequencing of 162 kb of dog genomic DNA containing the complete approximately 71-kb canine DAG1 gene, which consists of three exons, with the translation start codon located in exon 2. Its 2679-nucleotide ORF encodes a polypeptide of 892 amino acids, which is highly similar to human, rabbit, and bovine orthologs. To further characterize the dog DAG1 gene we determined the transcription start site and several naturally occurring polymorphisms, which partially result in amino acid substitutions of the dystroglycan protein. The dog DAG1 gene was assigned to chromosome 20q15.1-q15.2 by FISH analysis. The analysis of the entire reported sequence revealed that the genes for aminomethyltransferase (AMT), bassoon (BSN), TCTA (T-cell leukemia translocation-associated) gene, and an as yet uncharacterized protein are located very close to the DAG1 gene. Therefore, this study defines a novel syntenic region among dog chromosome 20q15, human chromosome 3p21, and murine chromosome 9F. PMID- 10720571 TI - Evolution of the neuropeptide Y receptor family: gene and chromosome duplications deduced from the cloning and mapping of the five receptor subtype genes in pig. AB - Neuropeptide Y (NPY) receptors mediate a variety of physiological responses including feeding and vasoconstriction. To investigate the evolutionary events that have generated this receptor family, we have sequenced and determined the chromosomal localizations of all five presently known mammalian NPY receptor subtype genes in the domestic pig, Sus scrofa (SSC). The orthologs of the Y(1) and Y(2) subtypes display high amino acid sequence identities between pig, human, and mouse (92%-94%), whereas the Y(4), Y(5), and y(6) subtypes display lower identities (76%-87%). The lower identity of Y(5) is due to high sequence divergence in the large third intracellular loop. The NPY1R, NPY2R, and NPY5R receptor genes were localized to SSC8, the NPY4R to SSC14, and NPY6R to SSC2. Our comparisons strongly suggest that the tight cluster of NPY1R, NPY2R, and NPY5R on human chromosome 4 (HSA4) represents the ancestral configuration, whereas the porcine cluster has been split by two inversions on SSC8. These 3 genes, along with adjacent genes from 14 other gene families, form a cluster on HSA4 with extensive similarities to a cluster on HSA5, where NPY6R and >13 other paralogs reside, as well as another large cluster on HSA10 that includes NPY4R. Thus, these gene families have expanded through large-scale duplications. The sequence comparisons show that the NPY receptor triplet NPY1R-NPY2R-NPY5R existed before these large-scale duplications. PMID- 10720572 TI - Fine mapping suggests that the goat Polled Intersex Syndrome and the human Blepharophimosis Ptosis Epicanthus Syndrome map to a 100-kb homologous region. AB - To clone the goat Polled Intersex Syndrome (PIS) gene(s), a chromosome walk was performed from six entry points at 1q43. This enabled 91 BACs to be recovered from a recently constructed goat BAC library. Six BAC contigs of goat chromosome 1q43 (ICC1-ICC6) were thus constructed covering altogether 4.5 Mb. A total of 37 microsatellite sequences were isolated from this 4.5-Mb region (16 in this study), of which 33 were genotyped and mapped. ICC3 (1500 kb) was shown by genetic analysis to encompass the PIS locus in a approximately 400-kb interval without recombinants detected in the resource families (293 informative meioses). A strong linkage disequilibrium was detected among unrelated animals with the two central markers of the region, suggesting a probable location for PIS in approximately 100 kb. High-resolution comparative mapping with human data shows that this DNA segment is the homolog of the human region associated with Blepharophimosis Ptosis Epicanthus inversus Syndrome (BPES) gene located in 3q23. This finding suggests that homologous gene(s) could be responsible for the pathologies observed in humans and goats. PMID- 10720573 TI - Structure of the highly conserved HERC2 gene and of multiple partially duplicated paralogs in human. AB - Recombination between chromosome-specific low-copy repeats (duplicons) is an underlying mechanism for several genetic disorders. Recently, a chromosome 15 duplicon was discovered in the common breakpoint regions of Prader-Willi and Angelman syndrome deletions. We identified previously the large HERC2 transcript as an ancestral gene in this duplicon, with approximately 11 HERC2-containing duplicons, and demonstrated that recessive mutations in mouse Herc2 lead to a developmental syndrome, juvenile development and fertility 2 (jdf2). We have now constructed and sequenced a genomic contig of HERC2, revealing a total of 93 exons spanning approximately 250 kb and a CpG island promoter. A processed ribosomal protein L41 pseudogene occurs in intron 2 of HERC2, and putative VNTRs occur in intron 70 (28 copies, approximately 76-bp repeat) and 3' exon 40 through intron 40 (6 copies, approximately 62-bp repeat). Sequence comparisons show that HERC2-containing duplicons have undergone several deletion, inversion, and dispersion events to form complex duplicons in 15q11, 15q13, and 16p11. To further understand the developmental role of HERC2, a highly conserved Drosophila ortholog was characterized, with 70% amino acid sequence identity to human HERC2 over the carboxy-terminal 743 residues. Combined, these studies provide significant insights into the structure of complex duplicons and into the evolutionary pathways of formation, dispersal, and genomic instability of duplicons. Our results establish that some genes not only have a protein coding function but can also play a structural role in the genome. PMID- 10720575 TI - The human adult skeletal muscle transcriptional profile reconstructed by a novel computational approach. AB - By applying a novel software tool, information on 4080 UniGene clusters was retrieved from three adult human skeletal muscle cDNA libraries, which were selected for being neither normalized nor subtracted. Reconstruction of a transcriptional profile of the corresponding tissue was attempted by a computational approach, classifying each transcript according to its level of expression. About 25% of the transcripts accounted for about 80% of the detected transcriptional activity, whereas most genes showed a low level of expression. This in silico transcriptional profile was then compared with data obtained by a SAGE study. A fairly good agreement between the two methods was observed. About 400 genes, highly expressed in skeletal muscle or putatively skeletal muscle specific, may represent the minimal set of genes needed to determine the tissue specificity. These genes could be used as a convenient reference to monitor major changes in the transcriptional profile of adult human skeletal muscle in response to different physiological or pathological conditions, thus providing a framework for designing DNA microarrays and initiating biological studies. PMID- 10720574 TI - Evaluation of single nucleotide polymorphism typing with invader on PCR amplicons and its automation. AB - Large-scale pharmacogenetics and complex disease association studies will require typing of thousands of single-nucleotide polymorphisms (SNPs) in thousands of individuals. Such projects would benefit from a genotyping system with accuracy >99% and a failure rate <5% on a simple, reliable, and flexible platform. However, such a system is not yet available for routine laboratory use. We have evaluated a modification of the previously reported Invader SNP-typing chemistry for use in a genotyping laboratory and tested its automation. The Invader technology uses a Flap Endonuclease for allele discrimination and a universal fluorescence resonance energy transfer (FRET) reporter system. Three hundred and eighty-four individuals were genotyped across a panel of 36 SNPs and one insertion/deletion polymorphism with Invader assays using PCR product as template, a total of 14,208 genotypes. An average failure rate of 2.3% was recorded, mostly associated with PCR failure, and the typing was 99.2% accurate when compared with genotypes generated with established techniques. An average signal-to-noise ratio (9:1) was obtained. The high degree of discrimination for single base changes, coupled with homogeneous format, has allowed us to deploy liquid handling robots in a 384-well microtitre plate format and an automated end point capture of fluorescent signal. Simple semiautomated data interpretation allows the generation of approximately 25,000 genotypes per person per week, which is 10-fold greater than gel-based SNP typing and microsatellite typing in our laboratory. Savings on labor costs are considerable. We conclude that Invader chemistry using PCR products as template represents a useful technology for typing large numbers of SNPs rapidly and efficiently. PMID- 10720576 TI - A fast and scalable radiation hybrid map construction and integration strategy. AB - This paper describes a fast and scalable strategy for constructing a radiation hybrid (RH) map from data on different RH panels. The maps on each panel are then integrated to produce a single RH map for the genome. Recurring problems in using maps from several sources are that the maps use different markers, the maps do not place the overlapping markers in same order, and the objective functions for map quality are incomparable. We use methods from combinatorial optimization to develop a strategy that addresses these issues. We show that by the standard objective functions of obligate chromosome breaks and maximum likelihood, software for the traveling salesman problem produces RH maps with better quality much more quickly than using software specifically tailored for RH mapping. We use known algorithms for the longest common subsequence problem as part of our map integration strategy. We demonstrate our methods by reconstructing and integrating maps for markers typed on the Genebridge 4 (GB4) and the Stanford G3 panels publicly available from the RH database. We compare map quality of our integrated map with published maps for GB4 panel and G3 panel by considering whether markers occur in the same order on a map and in DNA sequence contigs submitted to GenBank. We find that all of the maps are inconsistent with the sequence data for at least 50% of the contigs, but our integrated maps are more consistent. The map integration strategy not only scales to multiple RH maps but also to any maps that have comparable criteria for measuring map quality. Our software improves on current technology for doing RH mapping in areas of computation time and algorithms for considering a large number of markers for mapping. The essential impediments to producing dense high-quality RH maps are data quality and panel size, not computation. PMID- 10720577 TI - RHO--radiation hybrid ordering. AB - Radiation hybrid (RH) mapping is a somatic cell technique that is used for ordering markers along a chromosome and estimating the physical distances between them. With the advent of this mapping technique, analyzing the experimental data is becoming a challenging and demanding computational task. In this paper we present the software package RHO (radiation hybrid ordering). The package implements a number of heuristics that attempt to order genomic markers along a chromosome, given as input the results of an RH experiment. The heuristics are based on reducing an appropriate optimization problem to the traveling salesman problem (TSP). The reduced optimization problem is either the nonparametric obligate chromosome breaks (OCBs) or the parametric maximum likelihood estimation (MLE). We tested our package on both simulated and publicly available RH data. For synthetic RH data, the reconstructed markers' permutation is very close to the original permutation, even with fairly high error rates. For real data we used the framework markers' data from the Whitehead Institute maps. For most of the chromosomes (18 out of 23), there is a perfect agreement or nearly perfect agreement (reversal of chromosome arm or arms) between our maps and the Whitehead framework maps. For the remaining five chromosomes, our maps improve on the Whitehead framework maps with respect to both optimization criteria, having higher likelihood and fewer breakpoints. For three chromosomes, the results differ significantly (lod score >1.75), with chromosome 2 having the largest improvement (lod score 3.776). PMID- 10720579 TI - Research means "Look back" PMID- 10720578 TI - HOBACGEN: database system for comparative genomics in bacteria. AB - We present here HOBACGEN, a database system devoted to comparative genomics in bacteria. HOBACGEN contains all available protein genes from bacteria, archaea, and yeast, taken from SWISS-PROT/TrEMBL and classified into families. It also includes multiple alignments and phylogenetic trees built from these families. The database is organized under a client/server architecture with a client written in Java, which may run on any platform. This client integrates a graphical interface allowing users to select families according to various criteria and notably to select homologs common to a given set of taxa. This interface also allows users to visualize multiple alignments and trees associated to families. In tree displays, protein gene names are colored according to the taxonomy of the corresponding organisms. Users may access all information associated to sequences and multiple alignments by clicking on genes. This graphic tool thus gives a rapid and simple access to all data required to interpret homology relationships between genes and distinguish orthologs from paralogs. Instructions for installation of the client or the server are available at http://pbil.univ-lyon1. fr/databases/hobacgen.html. PMID- 10720580 TI - CYP11B2 gene polymorphisms in idiopathic hyperaldosteronism. AB - Primary aldosteronism is characterized by autonomous production of aldosterone and arterial hypertension, and it occurs in 2 principal forms: aldosterone producing adenoma (APA) and idiopathic hyperaldosteronism (IHA). APA can be cured through removal of the adenoma, whereas IHA leads to hypertension that must be treated with medication. The origin of the autonomous aldosterone production in IHA is poorly understood, but genetic factors may contribute to its cause. To test the hypothesis that variants of the aldosterone synthase gene may contribute to susceptibility to IHA, we compared genotypes at 3 polymorphic sites in the CYP11B2 gene in patients with IHA (n=90) with those found in patients with APA (n=38), in patients with essential hypertension (n=72), and in normotensive individuals (n=102). We observed significant linkage disequilibrium among the 3 polymorphisms with 2 frequent haplotypes in all groups studied. One haplotype (C2R) was found to be increased in frequency in the IHA group (47%) compared with the other groups, which had a similar haplotype frequency (36%). The 3 polymorphisms studied have been implicated in either essential hypertension or excess aldosterone production in previous studies. Because of the strong linkage disequilibrium, the observed results could be due to the action of any 1 of the 3 alleles or to another allele in linkage disequilibrium with them. Our results suggest that variations in the CYP11B2 gene may contribute to dysregulation of aldosterone synthesis and lead to susceptibility to IHA. PMID- 10720581 TI - Lys(173)Arg and -344T/C variants of CYP11B2 in Japanese patients with low-renin hypertension. AB - We analyzed the association of 2 biallelic polymorphisms of CYP11B2 (P450c11AS) gene (1 in the Lys(173)Arg of exon 3 and the other in the promoter at position 344T/C) with hypertension in 73 hypertensive patients and 134 normotensive subjects. The association between low-renin hypertension and angiotensin I converting enzyme (ACE) gene was also analyzed. An elevated ratio of plasma aldosterone concentration to plasma renin activity was used to identify low-renin hypertension. Genotypes for CYP11B2 and ACE were determined through polymerase chain reactions. The Arg(173) allele frequency did not differ between hypertensive patients considered as 1 group (34%) and normotensive control subjects (37%). However, only 22% of 58 CYP11B2 alleles studied in 29 patients with low-renin hypertension were Arg(173) alleles, whereas the frequency of this allele was 41% in patients with normal- or high-renin hypertension (P=0.033). An analysis of the distribution of -344C and Arg(173) genotypes indicated that these 2 variants were in complete linkage disequilibrium: -344C was present in a subset of chromosomes carrying the Arg(173) (P<0.001 in low-renin hypertension). Therefore, the frequency of the -344C allele was low in the patients with low renin hypertension compared with those with normal- or high-renin hypertension. Deletion (D) allele frequencies of the ACE gene were 31% in the patients with low renin hypertension, 39% in the patients with normal- or high-renin hypertension, and 29% in normotensive control subjects. We detected an association between the CYP11B2 gene polymorphisms and low-renin hypertension with inappropriate elevation of aldosterone. The decreased frequencies of the Arg(173) and -344C variants in the CYP11B2 appear to be genetically linked to low-renin hypertension in the Japanese population studied. PMID- 10720582 TI - Evaluation of the aldosterone synthase (CYP11B2) gene polymorphism in patients with myocardial infarction. AB - Left ventricular remodeling after myocardial infarction involves activation of the renin-angiotensin-aldosterone system. Recently, the biallelic -344T/C polymorphism of the aldosterone synthase gene was associated with increased aldosterone levels, arterial hypertension, diastolic dysfunction, and left ventricular dilatation. We hypothesized that this polymorphism may also affect left ventricular geometry and function after myocardial infarction. By using a standardized questionnaire, as well as anthropometric and echocardiographic measurements, we thus studied 606 patients (533 men and 73 women) who had a myocardial infarction before the age of 60 years. The aldosterone synthase gene polymorphism was analyzed after polymerase chain reaction amplification and restriction enzyme digestion. The results demonstrated that there was no association of the aldosterone synthase gene polymorphism with echocardiographically determined left ventricular dimensions, wall thicknesses, or indexes of systolic or diastolic function. Furthermore, anthropometric data, including blood pressure levels, were balanced between the different genotypes. Finally, the allele frequency was similar for patients with myocardial infarction and a sample group from the normal population (n=1675). The data indicate that the allele status of the aldosterone synthase gene polymorphism is not useful for the identification of patients with myocardial infarction who have impaired left ventricular function or unfavorable remodeling. PMID- 10720583 TI - Aldosterone stimulation by angiotensin II : influence of gender, plasma renin, and familial resemblance. AB - The aldosterone response to infused angiotensin II (Ang II) in patients receiving a low-salt diet has been described as an important phenotype for genetic studies on human hypertension. The objectives of the present study were to determine the parameters that influence this intermediate phenotype as a quantitative trait and to assess the importance of its familial resemblance in hypertensive sibling pairs. Two hundred one white hypertensive subjects (95 families: 84 pairs and 11 trios) were selected in 3 centers. The patients followed the same protocol, which included a 4-week withdrawal period of antihypertensive therapy, a 1-week period on a low-salt diet, and a 30-minute infusion of Ang II. The increase in the aldosterone level was greater in women than in men (29.1+/-16.2 versus 18.2+/-9.6 ng/dL, P<0.0001). A strong relationship was found with age (r=-0.54, P<10(-4)) and plasma renin activity (r=0.32, P<10(-4)) in women but not in men. Weak correlations of the aldosterone response to Ang II were observed for the whole set of sibling pairs (r=0.11, NS). Conversely, strong sibling correlations were found among brother-brother pairs (r=0.40, n=36) and among sister-sister pairs as soon as age or menopausal status was considered. Similar results were obtained when the Ang I-aldosterone response was analyzed as a qualitative trait (kappa=0. 35, P<0.008 in brother-brother pairs). We conclude that age, gender, and plasma renin are strong determinants of the aldosterone response to Ang II, with strong sibling correlations in men and postmenopausal women. These relationships will have to be considered in future linkage and association studies. PMID- 10720584 TI - Angiotensin II type 1 receptor A1166C gene polymorphism is associated with an increased response to angiotensin II in human arteries. AB - An adenine/cytosine (A/C) base substitution at position 1166 in the angiotensin II type 1 receptor (AT(1)R) gene is associated with the incidence of essential hypertension and increased coronary artery vasoconstriction. However, it is still unknown whether this polymorphism is associated with a difference in angiotensin II responsiveness. Therefore, we assessed whether the AT(1)R polymorphism is associated with different responses to angiotensin II in isolated human arteries. Furthermore, we evaluated whether inhibition of the renin-angiotensin system modifies the effect of the AT(1)R polymorphism. One hundred twelve patients who were undergoing coronary artery bypass graft surgery were prospectively randomized to receive an ACE inhibitor or a placebo for 1 week before surgery. Excess segments of the internal mammary artery were exposed to angiotensin II (0.1 nmol/L to 1 micromol/L) and KCl (60 mmol/L) in organ bath experiments. Patients homozygous for the C allele (n=17) had significantly greater angiotensin II responses (percentage of this maximal KCl-induced response) than did patients genotyped with AA+AC (n=95, P<0.05). Although ACE inhibition increased the response to angiotensin II, the difference in the response to angiotensin II, between CC and AA+AC patients remained intact in ACE inhibitor-treated patients. These results indicate increased responses to angiotensin II in patients with the CC genotype. The mechanism is preserved during ACE inhibition, which in itself also increased the response to angiotensin II. This reveals that the A1166C polymorphism may be in linkage disequilibrium with a functional mutation that alters angiotensin II responsiveness, which may explain the described relation between this polymorphism and cardiovascular abnormalities. PMID- 10720585 TI - Racial differences in endothelin-1 at rest and in response to acute stress in adolescent males. AB - Blacks exhibit greater vasoconstriction-mediated blood pressure (BP) increases in response to stress than do whites. Endothelin-1 (ET-1), a potent vasoconstrictive peptide, has been proposed as having a role in racial differences in stress reactivity. We evaluated the hemodynamic and plasma ET-1 levels of 41 (23 whites, 18 blacks, mean age 18.6 years) normotensive adolescent males at rest and in response to a video game challenge and forehead cold stimulation. Measurements were performed at catheter insertion and before and immediately after the 2 stressors, which were separated by 20-minute rest periods. Blacks exhibited higher absolute levels of diastolic blood pressure, total peripheral resistance index, or both in response to catheter insertion and to the video game challenge and during recovery from video game challenge and cold stimulation (P<0. 05 for all). Blacks exhibited higher absolute levels of ET-1 at every evaluation point (P<0.05 for all) and greater increases in ET-1 in response to both stressors (ps<0.05). These findings suggest that altered endothelial function may be involved in racial differences in hemodynamic reactivity to stress and possibly in the development of essential hypertension. PMID- 10720586 TI - ET(B) receptor blockade potentiates the pressor response to big endothelin-1 but not big endothelin-2 in the anesthetized rabbit. AB - The precursor of endothelin-1, big endothelin-1, is considered to be a more reliable marker of systemic production of vasoactive peptide. However, it is largely unclear whether ET(B) receptor-dependent clearance and endothelium derived relaxing factors affect the precursor in a similar manner to mature ET-1. These ET(B)-dependent modulations of big ET-1 and big ET-2 pressor properties were therefore studied in the anesthetized rabbit. When injected into the left cardiac ventricle, ET-1 and ET-2 (0.01 to 1 nmol/kg) each induced biphasic responses (a depressor followed by a pressor response), whereas big ET-1 and big ET-2 (0.1 to 3 nmol/kg) caused only protracted pressor responses. The highest dose of big ET-1 caused significantly greater responses than ET-1, ET-2, or big ET-2. A selective ET(A) receptor antagonist, BQ-123 (1 mg/kg), markedly reduced pressor responses to all 4 peptides, whereas blockade of ET(B) receptors with BQ 788 (0.25 mg/kg) sharply potentiated the responses to ET-1, ET-2, and big ET-1, but not to big ET-2. Indomethacin (10 mg/kg) sharply potentiated the pressor response to ET-1 (1 nmol/kg), but not big ET-1, at all time points. In control animals, ET-1, but not big ET-1, also triggered an indomethacin-sensitive increase in circulating prostacyclin. Finally, systemically administered big ET 1, but not big ET-2, induced a phosphoramidon-sensitive increase in plasma IR-ET. Our results suggest a significant limiting role of ET(B) receptors on pressor responses to big ET-1. In contrast, the same receptor entities do not modulate the hemodynamic properties of the ET-2 precursor, given that, unlike big ET-1, it is poorly converted in the pulmonary or systemic circulation in anesthetized rabbits. PMID- 10720587 TI - Mechanisms of big endothelin-1-induced diuresis and natriuresis : role of ET(B) receptors. AB - Endothelin-1 (ET-1) at high concentrations has marked antidiuretic and antinatriuretic activities, whereas its precursor, big endothelin-1 (big ET-1), has surprisingly potent diuretic and natriuretic actions. The mechanisms underlying the excretory effects of big ET-1 have not been fully elucidated. To explore these mechanisms, we examined the effects of a highly selective ET(B) antagonist (A-192621.1), a calcium channel blocker (verapamil), a nitric oxide synthase inhibitor (N-nitro-L-arginine methyl ester [L-NAME]), and a cyclooxygenase inhibitor (indomethacin) on the systemic and renal actions of big ET-1 in anesthetized rats. An intravenous bolus injection of incremental doses of big ET-1 (0.3, 1. 0, and 3.0 nmol/kg) produced a significant hypertensive effect that was dose dependent and prolonged (from 113+/-7 mm Hg to a maximum of 148+/-6 mm Hg). The administration of big ET-1 induced marked diuretic and natriuretic responses (urinary flow rate increased from 8.5+/-1 to 110+/-14 microL/min, and fractional excretion of sodium increased from 0.38+/-0.13% to 7.51+/-1.24%). Glomerular filtration rate and renal plasma flow significantly decreased only at the highest dose of big ET-1. Pretreatment with A-192621.1 (3 mg/kg plus 3 mg. kg(-1). h(-1)) significantly abolished the diuretic (17+/-5 microL/min to a maximum of 19+/-3 microL/min) and natriuretic (0. 29+/-0.1% to a maximum of 1.93+/-0.37%) responses induced by big ET-1. Moreover, A-192621.1 potentiated the decline in glomerular filtration rate and renal plasma flow and the increase in mean arterial blood pressure produced by the low doses of big ET-1. Similar to A 192621.1, pretreatment with a nitric oxide synthase inhibitor (L-NAME, 10 mg/kg plus 5 mg. kg(-1). h(-1)) significantly and comparably reduced the diuretic and natriuretic actions of big ET-1 and augmented the hypoperfusion/hypofiltration and systemic vasoconstriction induced by high doses of the peptide. Pretreatment with verapamil (2 mg. kg(-1). h(-1)) slightly inhibited the diuretic/natriuretic effects of the high-dose big ET-1 and completely prevented the increase in mean arterial blood pressure provoked by the peptide. Unlike verapamil and L-NAME, only indomethacin administration was associated with significant natriuretic/diuretic responses and did not influence the pressor effect and renal actions of big ET-1. Taken together, these results suggest that big ET-1-induced diuretic and natriuretic responses are mediated mainly by stimulation of nitric oxide production coupled to ET(B) receptor subtype activation. PMID- 10720588 TI - Nitric oxide in the renal medulla protects from vasopressin-induced hypertension. AB - In the present study, we assessed whether activation of the nitric oxide (NO) system within the renal medulla could serve as a buffer against the chronic hypertensive effects of arginine vasopressin (AVP). NO concentration in the renal medulla of Sprague-Dawley rats was measured with in vivo microdialysis/oxyhemoglobin NO trapping. The results showed that medullary interstitial [NO] was increased after 2 hours of AVP infusion and remained elevated even after 10 days (by 62+/-8% and 42+/-13%, respectively). Western blot analysis showed that 2 days of AVP infusion was insufficient to increase protein expression of any of the NO synthase (NOS) isoforms, but after 10 days of AVP infusion, endothelial NOS expression was significantly increased in the inner medulla with no significant changes in noninducible NOS and inducible NOS levels. When renal medullary NOS enzyme activity was blunted with a nonpressor dose of N(G)-nitro-L-arginine methyl ester (75 microg. kg(-1). h(-1)) that was chronically infused locally into the renal medulla, intravenous AVP infusion (which was shown earlier to be subpressor in chronic studies) produced a sustained elevation in arterial pressure (from 107+/-2 to 121+/-2 mm Hg). These data indicate that chronic elevations in plasma AVP enhance renal medullary endothelial NOS protein expression, which enables sustained elevations of NO concentrations in this region of the kidney to buffer the hypertensive effects of AVP. PMID- 10720589 TI - Allopurinol normalizes endothelial dysfunction in type 2 diabetics with mild hypertension. AB - Therapeutic strategies against free radicals have mostly focused on the augmentation of antioxidant defenses (eg, vitamins C and E). A novel approach is to prevent free radical generation by the enzyme system xanthine oxidase. We examined whether the inhibition of xanthine oxidase with allopurinol can improve endothelial function in subjects with type 2 diabetes and coexisting mild hypertension compared with control subjects of a similar age. We examined 23 subjects (11 patients with type 2 diabetes and 12 healthy age-matched control subjects) in 2 parallel groups. The subjects were administered 300 mg allopurinol in a randomized, placebo-controlled study in which both therapies were administered for 1 month. Endothelial function was assessed with bilateral venous occlusion plethysmography, in which the forearm blood flow responses to intra arterial infusions of endothelium-dependent and -independent vasodilators were measured. Allopurinol significantly increased the mean forearm blood flow response to acetylcholine by 30% (3.16+/-1.21 versus 2.54+/-0.76 mL. 100 mL(-1). min(-1) allopurinol versus placebo; P=0.012, 95% CI 0.14, 1.30) but did not affect the nitroprusside response in patients with type 2 diabetes. There was no significant impact on either endothelium-dependent or -independent vascular responses in age-matched control subjects. Allopurinol improved endothelial function to near-normal levels. Regarding markers of free radical activity, the level of malondialdehyde was significantly reduced (0.30+/-0.04 versus 0. 34+/ 0.05 micromol/L for allopurinol versus placebo, P=0.03) in patients with type 2 diabetes but not in control subjects. The xanthine oxidase inhibitor allopurinol improves endothelial dysfunction in patients with type 2 diabetes with mild hypertension but not in matched control subjects. In the former group, allopurinol restored endothelial function to near-normal levels. PMID- 10720590 TI - Interaction between nitric oxide and mineralocorticoids in the long-term control of blood pressure. AB - We analyzed the effects of a possible interaction between nitric oxide deficiency and mineralocorticoids on the long-term control of blood pressure and renal and endocrine variables. Six groups of uninephrectomized male Wistar rats were used: control animals and rats that received (1) N(G)-nitro-L-arginine methyl ester (L NAME) subpressor (0.5 mg/100 mL drinking fluid), (2) L-NAME pressor (35 mg/100 mL drinking fluid), (3) deoxycorticosterone acetate (DOCA; 12. 5 mg/wk per rat), (4) DOCA plus L-NAME subpressor, or (5) L-NAME pressor plus DOCA. For all groups, the drinking fluid was tap water or 1% NaCl solution. We measured the time course of tail systolic blood pressure (SBP) and body weight for 3 weeks in all rats. At the end of the experimental period, we measured mean arterial pressure (direct recording) and endocrine and renal variables. Tail SBP rose significantly in the DOCA plus L-NAME subpressor-treated group but remained at normotensive levels in the DOCA-treated group. The addition of L-NAME to the subpressor dose accelerated the blood pressure increase in DOCA-salt hypertensive rats. The simultaneous administration of DOCA and L-NAME increased blood pressure and mortality rates in rats that drank water or saline compared with the rats treated with L-NAME alone. The subpressor dose of L-NAME did not increase blood pressure in saline-drinking rats. We conclude that impaired NO synthesis results in increased sensitivity to the pressor effect of mineralocorticoids in the presence or absence of an increased saline intake. Hence, nitric oxide contributes to the adaptative response to mineralocorticoid excess, perhaps through the facilitation of natriuresis and, thus, control of blood pressure. PMID- 10720591 TI - Acute suppression of muscle sympathetic nerve activity by hydrocortisone in humans. AB - In the present study, we examined the acute influence of hydrocortisone on human sympathetic nerve activity and cardiovascular parameters. Muscle sympathetic nerve activity (MSA), heart rate, and blood pressure were monitored in 8 healthy subjects (20 to 37 years old) before and after a bolus injection of 50 mg hydrocortisone followed by a continuous infusion at 50 mg/h during a period of 3 hours in a placebo-controlled, double-blind, crossover protocol. Recordings were performed at rest and during repeated transient sympathoexcitation induced by voluntary apneas. Resting MSA and endogenous serum cortisol concentrations were also measured in a larger study group (49 experiments, 25 subjects). During the experimental period, MSA burst number increased by 56% from the control level in the placebo group. In contrast, MSA was suppressed by 25% at the end of the hydrocortisone infusion, resulting in a significant treatment effect (P<0.05). In addition, sympathoexcitation during apnea was significantly reduced with hydrocortisone after 180 minutes. In parallel with the sympathetic outflow, blood pressure decreased in the hydrocortisone-treated group, whereas it rose in the placebo group (P<0.05 between groups). No correlation was found between basal MSA and basal cortisol levels. Our results indicate that pharmacological doses of hydrocortisone acutely influence MSA responses to short- and long-lasting environmental stimuli, whereas basal native cortisol levels do not appear to be tonically involved in the regulation of resting MSA. The suppressive hydrocortisone effect is most likely induced via supraspinal autonomic centers and cannot be explained by peripheral steroid mechanisms. The effect of elevated corticosteroid levels on sympathetic nerve discharge may be an important mechanism in cardiovascular adaptations to stress. PMID- 10720592 TI - Functional importance of angiotensin-converting enzyme-dependent in situ angiotensin II generation in the human forearm. AB - To assess the importance for vasoconstriction of in situ angiotensin (Ang) II generation, as opposed to Ang II delivery via the circulation, we determined forearm vasoconstriction in response to Ang I (0.1 to 10 ng. kg(-1). min(-1)) and Ang II (0.1 to 5 ng. kg(-1). min(-1)) in 14 normotensive male volunteers (age 18 to 67 years). Changes in forearm blood flow (FBF) were registered with venous occlusion plethysmography. Arterial and venous blood samples were collected under steady-state conditions to quantify forearm fractional Ang I-to-II conversion. Ang I and II exerted the same maximal effect (mean+/-SEM 71+/-4% and 75+/-4% decrease in FBF, respectively), with similar potencies (mean EC(50) [range] 5.6 [0.30 to 12.0] nmol/L for Ang I and 3.6 [0.37 to 7.1] nmol/L for Ang II). Forearm fractional Ang I-to-II conversion was 36% (range 18% to 57%). The angiotensin converting enzyme (ACE) inhibitor enalaprilat (80 ng. kg(-1). min(-1)) inhibited the contractile effects of Ang I and reduced fractional conversion to 1% (0.1% to 8%), thereby excluding a role for Ang I-to-II converting enzymes other than ACE (eg, chymase). The Ang II type 1 receptor antagonist losartan (3 mg. kg(-1). min( 1)) inhibited the vasoconstrictor effects of Ang II. In conclusion, the similar potencies of Ang I and II in the forearm, combined with the fact that only one third of arterially delivered Ang I is converted to Ang II, suggest that in situ generated Ang II is more important for vasoconstriction than circulating Ang II. Local Ang II generation in the forearm depends on ACE exclusively and results in vasoconstriction via Ang II type 1 receptors. PMID- 10720593 TI - Cardiovascular effects of combination of perindopril, candesartan, and amlodipine in hypertensive rats. AB - The combination therapy with ACE inhibitors, angiotensin II type 1 (AT(1)) receptor antagonists, or calcium channel antagonists may exert more beneficial effects on cardiovascular diseases than monotherapy. Perindopril, candesartan cilexetil, or amlodipine alone or the combination of low doses of each agent was administered orally to stroke-prone spontaneously hypertensive rats (SHRSP) for 4 weeks to compare the hypotensive or cardiovascular effects. Although perindopril (2 mg/kg), candesartan cilexetil (2 mg/kg), or amlodipine (3 mg/kg) alone caused comparable hypotensive effects in SHRSP, monotherapy with perindopril or candesartan decreased left ventricular (LV) weight; mRNA levels for atrial natriuretic factor, skeletal alpha-actin, and collagen types I and III; and aortic weight and platelet-derived growth factor-beta receptor tyrosine phosphorylation to a greater extent than monotherapy with amlodipine. Although monotherapy with a low dose (0.2 mg/kg) of perindopril or candesartan cilexetil did not significantly reduce the LV mRNA levels and aortic platelet-derived growth factor-beta receptor phosphorylation of the SHRSP, combination therapy at such a low dose normalized these parameters more potently than the use of amlodipine (3 mg/kg) alone. Although perindopril or candesartan cilexetil alone at 0.05 mg/kg did not decrease the blood pressure of the SHRSP, such a low dose of combination therapy decreased LV weight and atrial natriuretic factor mRNA levels of the SHRSP to a greater extent than amlodipine alone or amlodipine combined with perindopril or candesartan cilexetil. Our results provide evidence that suggests the combination of an ACE inhibitor and an AT(1) receptor antagonist may be more effective in the treatment of cardiac and vascular diseases than the combination of a calcium channel blocker with an ACE inhibitor or an AT(1) receptor antagonist or monotherapy with each agent. PMID- 10720594 TI - Effect of calcium antagonists on glomerular arterioles in spontaneously hypertensive rats. AB - Through the use of microanatomic techniques, we investigated the effects of treatment with some dihydropyridine-type calcium antagonists (CAs) (ie, lercanidipine, manidipine, and nicardipine) and with the nondihydropyridine-type vasodilator hydralazine on hypertension-dependent glomerular injury and on the morphology of afferent and efferent arterioles in spontaneously hypertensive rats (SHR). Fourteen-week-old male SHR and age-matched normotensive Wistar-Kyoto rats were left untreated (control groups). Four additional groups of 14-week-old SHR were treated for 12 weeks with daily oral doses of 2.5 mg/kg lercanidipine, 5 mg/kg manidipine, 3 mg/kg nicardipine, or 10 mg/kg hydralazine. These treatments decreased systolic blood pressure values to a similar extent in SHR. Signs of glomerular injury, as characterized by glomerulosclerosis, hypertrophy, and an increased number of mesangial cells, were observed in control SHR. The treatment with CAs improved glomerular morphology and decreased the number of mesangial cells. Lercanidipine and manidipine were more effective than nicardipine in countering glomerular injury. In the SHR, both afferent and efferent arterioles revealed luminal narrowing, accompanied by increased wall thickness in efferent arterioles. The dihydropyridine-type derivatives that were tested decreased the luminal narrowing of afferent arterioles. Lercanidipine and manidipine countered the luminal narrowing of efferent arterioles. Hydralazine had no effect on hypertension-dependent glomerular injury or vascular changes. The present data indicate that lercanidipine and manidipine vasodilate afferent and efferent arterioles in SHR. A vasodilatory activity on efferent arteriole, which is not induced by the majority of CAs, may represent an useful property in the treatment of hypertension complicated by renal disease. PMID- 10720595 TI - Angiotensinogen concentrations and renin clearance : implications for blood pressure regulation. AB - Renin (REN) requires seconds to convert angiotensinogen (AGT) to angiotensin I. We tested the hypothesis that this long catalytic cycle might indicate an influence of AGT concentrations on REN clearance. We studied 2 transgenic rat (TGR) strains for human (h) AGT; one strain has hAGT values approximately 7-fold higher than the other (68+/-18 versus 10+/-4 microg angiotensin I/mL). hREN (30 000 pg) was bolus-infused into both lines and into nontransgenic controls. The terminal half-life (T1/2beta) was increased (130 versus 82 minutes) and the metabolic clearance rate (MCR) was decreased (0.83+/-0.29 versus 2.2+/-0.66 microL. min(-1). g(-1)) in the high hAGT strain compared with the low hAGT strain. The difference was not related to volume of distribution at steady state. Infused hREN blocked with remikiren resulted in T1/2beta and MCR values that were not different from control values. Infused unblocked and blocked radiolabeled hREN was distributed similarly in the hAGT TGR strains. Infused mouse REN, which cannot convert hAGT, had similar T1/2beta and MCR values in hAGT TGR. Measuring REN with direct radioimmunoassay or by enzyme kinetic assay gave similar results. We next crossed homozygous hAGT TGR from both strains with homozygous hREN TGR. Heterozygous offspring from the low hAGT TGR strain had plasma REN activity, hREN concentration, and rat AGT values that were no different from those of their parents. However, TGR offspring with high hAGT values had massively elevated plasma REN activity and hREN concentration as well as elevated blood pressure, even though both the hREN and rREN genes are downregulated. We conclude that increased AGT concentrations decrease REN MCR and increase REN T1/2beta. The REN AGT complex may stabilize plasma REN concentration and regulate plasma REN activity independent of renal REN secretion and angiotensin II-mediated feedback. These effects could augment angiotensin I generation and influence blood pressure. The notion that AGT is merely a passive substrate reservoir for REN should be revised. PMID- 10720596 TI - Changes of nocturnal blood pressure dipping status in hypertensives by nighttime dosing of alpha-adrenergic blocker, doxazosin : results from the HALT study. AB - Abnormal nocturnal blood pressure (BP) dipping status may be partly determined by nocturnal sympathetic activity. We studied the effect of nighttime dosing of an alpha(1)-adrenergic blocker, doxazosin, on the BP dipping status of 118 hypertensives, all of whom underwent 24-hour ambulatory BP monitoring before and after treatment. The mean nighttime/daytime ratio of systolic BP was increased (0.91 after therapy versus 0.89 at baseline, P<0.05). The patients were initially divided into 4 groups on the basis of their dipping status at the baseline assessment: 18 (15%) were extreme dippers, with a nighttime systolic BP fall of at least 20% of daytime BP; 46 (39%) were dippers (fall between 10% and 20%); 48 (41%) were nondippers (fall between 0% and 10%); and 6 (5%) were risers (nocturnal increase of systolic BP). A shift in dipping status toward less nocturnal BP dipping was observed after doxazosin therapy (P<0.05). Dipping status was determined by nighttime more than by daytime BP, and this was not explained by differences in the number of daytime and nighttime readings. The effects of doxazosin on the mean nocturnal systolic BP changes were an increase of 4.3 mm Hg in extreme dippers and decreases of 0.7 mm Hg in dippers, 12 mm Hg in nondippers, and 18 mm Hg in risers; the reduction was only significant in the latter 2 groups (both P<0.01). To estimate the effects of regression to the mean on the changes in dipping status, we also defined dipping status with the average of the BPs before and after doxazosin and found no difference in the degree of nighttime BP reduction among each group. The reduction of daytime BP was now significantly greater in the subgroups with less dipping: 6. 4 mm Hg for extreme dippers and 16 mm Hg for risers (P<0.05). In conclusion, nighttime dosing with doxazosin markedly affects the nocturnal BP dipping status of hypertensives, but the apparently greater reduction in nighttime pressure in nondippers and risers may be, at least partly, due to the effect of regression to the mean. The most important determinants of the effect of doxazosin were the absolute BP levels, both day and night, rather than dipping status per se. PMID- 10720597 TI - Effects of menstrual cycle and race on peripheral vascular alpha-adrenergic responsiveness. AB - Gender differences in the incidence of many cardiovascular diseases may be due to the effects of sex hormones. Both alpha(1)- and alpha(2)-adrenergic receptors produce vasoconstriction in peripheral blood vessels and have demonstrated gender effects in previous studies. In addition, race has been shown to influence the effects of some alpha-adrenergic stimuli. We therefore sought to determine the effects of the menstrual cycle and race on peripheral blood flow responses to the intra-arterial infusion of phenylephrine (alpha(1)-agonist) and clonidine (alpha(2)-agonist). Ten white and 8 black women were studied during the early luteal phase and the follicular phase; these phases were verified in each woman through measurements of plasma estradiol and progesterone. Plasma norepinephrine was measured with HPLC. During phenylephrine infusion, there was significantly greater vasoconstriction in the luteal phase versus the follicular phase (P<0.05). There were no differences (P>0.8) between white and black women. During clonidine infusion, white women showed significantly more vasoconstriction in the follicular phase than during the luteal phase (P<0.006). For black women, the responses for both phases did not differ (P>0.9). Blood pressures were significantly higher in the black women (diastolic P<0.005, systolic P<0.05). The luteal-phase elevation of alpha(1)-adrenergic responses may be due to elevated levels of estradiol, progesterone, or both. The lack of luteal-phase reduction in alpha(2)-adrenergic vasoconstriction in black women may contribute to their increased pressor responses to adrenergic stimuli. PMID- 10720598 TI - Heme oxygenase-1 is upregulated in the kidney of angiotensin II-induced hypertensive rats : possible role in renoprotection. AB - In this study, we investigated the regulation and physiological role of heme oxygenase-1 (HO-1) in the kidney of rats with hypertension. Rats were continuously administered either angiotensin II (Ang II) or norepinephrine with an osmotic minipump for up to 7 days. Ang II infusion decreased the glomerular filtration rate (GFR) as determined through creatinine clearance (3.2+/-0.2 versus 1.2+/-0.2 mL/min with Ang II infusion, P<0.01) and increased proteinuria (9. 7+/-1.3 versus 28.1+/-7.2 mg/d with Ang II infusion, P<0.01). In contrast, norepinephrine did not alter these laboratory values. Ang II infusion significantly increased HO-1 expression in mRNA (442+/-98% of control at day 5, P<0.01) and protein levels (314+/-49% of control at day 5, P<0.01). Immunohistochemistry showed that in the kidney of normotensive rats, HO-1 was expressed mainly in the basal side in the renal tubules. After Ang II infusion, HO-1 staining was more extensively dispersed in the tubular epithelial cells. The intraperitoneal administration of zinc protoporphyrin, an HO inhibitor, to Ang II infused rats further decreased GFR (0.8+/-0. 1 mL/min) and increased proteinuria (52.5+/-13.0 mg/d). In contrast, the administration of hemin, an HO inducer, ameliorated the Ang II-induced decrease in GFR (2.4+/-0.2 mL/min) and increase in proteinuria (9.3+/-4.5 mg/d). These data suggest that HO-1 upregulation in the kidney of Ang II-induced hypertensive rats may exert a renoprotective effect against Ang II-induced renal injury. PMID- 10720599 TI - Overweight and hypertension : a 2-way street? AB - Cross-sectionally, higher weight is associated with higher blood pressure levels; prospectively, baseline weight and weight gain predict higher blood pressure. The loss of weight is frequently associated with a decrease in blood pressure. These findings suggest that weight gain may pathophysiologically contribute to blood pressure elevation. In this review, we present data to indicate that the reverse is also true; persons of equal weight who had higher initial blood pressures gain more weight in the future. We also propose a plausible hypothesis to explain this reverse relationship. Both the blood pressure elevation and the gain of weight may reflect a primary increase in sympathetic tone. It is well known that in a milieu of increased sympathetic tone, the beta-adrenergic responsiveness decreases. Sympathetic overactivity and decreased cardiovascular beta-adrenergic responsiveness have been described in hypertension. beta-Adrenergic receptors mediate increases in energy expenditure. If these metabolic receptors were downregulated in hypertension, the ability of hypertensive patients to dissipate calories would decrease and they would gain more weight. The possible relationship of decreased beta-adrenergic responsiveness to weight in hypertension can be experimentally tested. Such research may contribute to an explanation of why patients with hypertension can rarely lose weight. An understanding of this pathophysiological relationship may open new avenues for therapeutic interventions. PMID- 10720600 TI - Prevalence, treatment, and control of hypertension by sociodemographic factors among the Dutch elderly. AB - The study objective was to assess the prevalence, level of treatment, and control of hypertension in a general elderly population according to age and sociodemographic factors. We conducted a cross-sectional analysis of 7983 participants of the Rotterdam Study who were >/=55 years old and living in a district of Rotterdam. The prevalence of hypertension was based on blood pressure levels (>/=160/95 mm Hg) and the use of blood pressure-lowering medication for the indication of hypertension, type of treatment, and control of hypertension. Systolic blood pressure rises with age, whereas diastolic blood pressure declines. The prevalence of hypertension increases with age and was higher among women (39%) than among men (31%). About 80% of the hypertensives were aware of having hypertension, and 82% of the 80% were treated. For 70% of them, treatment was adequate with reference to conservative criteria. Hypertension was more prevalent among persons not living in a home for the elderly, for more-educated men, and for less-educated women. Persons without a partner and men living in a home for the elderly had a higher risk of being unaware of or of not being treated for existing hypertension. Treatment was more often successful among those living in a home for the elderly. The prevalence of hypertension was higher among older women and increased with age in both genders. A large proportion of hypertensive elderly persons were aware and were successfully treated for hypertension. The degree of awareness and control appeared to be affected by sociodemographic factors. More importantly, the majority of hypertensives did not have their hypertension well controlled. This group requires more attention by medical practitioners to reduce the burden of cardiovascular diseases in elderly persons. PMID- 10720601 TI - Glomerular hyperfiltration in hypertensive African Americans. AB - The incidence of end-stage renal disease attributable to hypertension is 5-fold greater in African Americans than in whites. To determine whether glomerular hyperfiltration is an antecedent to renal failure, we compared responses of renal blood flow and glomerular filtration rate to graded infusions of norepinephrine (0. 01, 0.025, and 0.05 microg. kg(-1). min(-1) for 30 minutes each) in 29 African Americans and 33 age-matched French Canadian whites with essential hypertension. Renal blood flow and glomerular filtration rate were measured by using a constant-infusion technique of PAH and inulin, respectively. Studies were conducted on an inpatient clinical research center, and antihypertensive medications had been discontinued for at least 1 week. Based on 24-hour blood pressure monitoring, nighttime blood pressures decreased (P<0.01) in the French Canadians but not in the African Americans. Baseline renal blood flow was higher (P<0.05) in the African Americans (1310+/-127 mL. min(-1) per 1.73 m(2)) than in the French Canadians (1024+/-42 mL. min(-1) per 1.73 m(2)); baseline glomerular filtration rate was also higher (P<0.01) in the African Americans (140+/-4 versus 121+/-4 mL. min(-1) per 1.73 m(2)). In response to norepinephrine-induced blood pressure increases, renal blood flow was autoregulated and did not change in either patient group. In the African Americans, glomerular filtration rate increased (P<0.01) to 167 mL. min(-1) per 1.73 m(2) during the first norepinephrine infusion, without subsequent change. In contrast, glomerular filtration rate did not change with norepinephrine-induced increases of blood pressure in the French Canadians. In the African Americans, the elevation of baseline glomerular filtration rate, with a further increase in response to norepinephrine, may be indicative of glomerular hyperfiltration. Glomerular hyperfiltration and lack of nocturnal blood pressure decline may contribute to the higher incidence of end-stage renal disease in hypertensive African Americans. PMID- 10720602 TI - Effect of salt on insulin sensitivity differs according to gender and degree of salt sensitivity. AB - The aim of the present study was to investigate the effect of salt intake on insulin sensitivity and the relation between salt sensitivity and insulin sensitivity in genetically hypertension-prone individuals. Twenty-eight healthy subjects (13 men and 15 women) with a family history of hypertension were examined at baseline, after 1 week of salt restriction (10 mmol/d), and after 1 week of salt loading (240 mmol/d). Insulin sensitivity was measured with the hyperinsulinemic euglycemic clamp after the low- and high-salt diets. Salt sensitivity was defined as the difference in mean arterial blood pressure between the high-salt and the low-salt diets. There was no significant relationship between insulin sensitivity and salt sensitivity after either of the 2 diets. In the men, salt sensitivity was inversely related to plasma renin activity (r= 0.61, P=0.03) and plasma aldosterone (r=-0.74, P=0.004), whereas salt sensitivity in women was directly correlated with the salt-induced increase in body weight (r=0.68, P=0.005). In men, the high-salt diet induced a change in glucose disposal that was strongly correlated with the degree of salt sensitivity (r=0.83, P=0. 0004), plasma renin activity (r=-0.82, P=0.0006), and plasma aldosterone concentrations (r=-0.87, P=0.00009) (eg, the greater the salt sensitivity and the lower the activity of the renin-angiotensin-aldosterone system, the greater improvement in insulin sensitivity). No such relationships were observed in women. In conclusion, increased salt sensitivity and decreased activity of the renin-angiotensin-aldosterone system predict improved insulin sensitivity with high-salt intake compared with low-salt intake in men, suggesting an interaction among salt intake, salt sensitivity, the renin angiotensin-aldosterone system, and insulin action. PMID- 10720603 TI - High-fat diet elevates blood pressure and cerebrovascular muscle Ca(2+) current. AB - Dietary fat contributes to the elevation of blood pressure and increases the risk of stroke and coronary artery disease. Previous observations have shown that voltage-gated Ca(2+) current density is significantly increased in hypertension and can be affected by free fatty acids (FAs). We hypothesized that a diet of elevated fat level would lead to an increase in blood pressure, an elevation of L type Ca(2+) current, and an increase in saturated FA content in vascular smooth muscle cell membranes. Male Osborne-Mendel rats were fed normal rat chow or a high-fat diet (Ob/HT group) for 8 weeks. Blood pressures in the Ob/HT group increased moderately from 122.5+/-0.7 to 134.4+/-0.8 mm Hg (P<0.05, n=26). Voltage-clamp examination of cerebral arterial cells revealed significantly elevated L-type Ca(2+) current density in the Ob/HT group. Voltage-dependent inactivation of the Ob/HT L-type channels was significantly delayed. Total serum FA contents were significantly elevated in the Ob/HT group, and HPLC analyses of fractional pools of FAs from segments of abdominal aorta revealed that arachidonic acid levels were elevated in the phospholipid fraction in Ob/HT. No differences in vascular membrane cholesterol contents were noted. Plasma cholesterol was significantly elevated in portal venous and cardiac blood samples from Ob/HT rats. These findings suggest that an elevation of plasma FAs may contribute to the development of hypertension via a process involving the elevation of Ca(2+) current density and an alteration of channel kinetics in the vascular smooth muscle membrane. PMID- 10720604 TI - Progressive resistance exercise and resting blood pressure : A meta-analysis of randomized controlled trials. AB - Hypertension is a major public health problem affecting an estimated 43 million civilian, noninstitutionalized adults in the United States (24% of this population). The purpose of this study was to use the meta-analytic approach to examine the effects of progressive resistance exercise on resting systolic and diastolic blood pressure in adult humans. Studies were retrieved via (1) computerized literature searches, (2) cross-referencing from original and review articles, and (3) review of the reference list by 2 experts on exercise and blood pressure. Inclusion criteria were as follows: (1) trials that included a randomized nonexercise control group; (2) progressive resistance exercise as the only intervention; (3) adult humans; (4) journal articles, dissertations, and masters theses published in the English-language literature; (5) studies published and indexed between January 1966 and December 1998; (6) resting systolic and/or diastolic blood pressure assessed; and (7) training studies lasting a minimum of 4 weeks. Across all designs and categories, fixed-effects modeling yielded decreases of approximately 2% and 4% for resting systolic and diastolic blood pressure, respectively (mean+/-SD systolic, -3+/-3 mm Hg; 95% bootstrap CI, -4 to -1 mm Hg; mean+/-SD diastolic, -3+/-2 mm Hg; 95% bootstrap CI, -4 to -1 mm Hg). It was concluded that progressive resistance exercise is efficacious for reducing resting systolic and diastolic blood pressure in adults. However, a need exists for additional studies that limit enrollment to hypertensive subjects as well as analysis of data with an intention-to-treat approach before the effectiveness of progressive resistance exercise as a nonpharmacological intervention can be determined. PMID- 10720606 TI - Hypertension online only : march 2000 PMID- 10720605 TI - Prognostic value of ambulatory blood pressure : current evidence and clinical implications. AB - This article is a critical review of the available evidence on the prognostic value of ambulatory blood pressure (ABP). Several event-based cohort studies have shown that ABP improves cardiovascular risk stratification over and beyond traditional risk factors, including office BP. Most of these studies have been conducted in subjects with essential hypertension who were untreated at the time of execution of ABP monitoring; other studies have been conducted in subjects who were poorly controlled with treatment or in the general population. In these studies, ABP was examined as a continuous variable or with operational risk categories. Cardiovascular risk showed a direct and independent association with the observed ABP (systolic, diastolic, and pulse) and an inverse association with the degree of BP reduction from day to night. Cardiovascular risk was also directly associated with the difference between the observed value of ABP and that predicted from the office BP. White-coat hypertension versus ambulatory hypertension and dippers versus nondippers are 2 classifications based on arbitrary operational risk categories. A blunted or absent BP reduction from day to night, defined with ABP as a continuous variable or with operational thresholds, was also associated with a worse outcome regardless of the average value of ABP during the 24 hours. Overall, these studies indicate that ABP monitoring is particularly valuable to refine cardiovascular risk stratification in untreated subjects with office hypertension and in those with resistant hypertension. Intervention studies targeted at ABP are now needed. PMID- 10720607 TI - Ambulatory blood pressure: normality and comparison with other measurements. PMID- 10720608 TI - Much ado about nothing, or much to do about something? The continuing controversy over the role of uric acid in cardiovascular disease. PMID- 10720609 TI - Organization of projections from the juxtacapsular nucleus of the BST: a PHAL study in the rat. AB - The axonal projections of the juxtacapsular nucleus of the anterior division of the bed nuclei of the stria terminalis (BSTju) were examined with the Phaseolus vulgaris-leucoagglutinin (PHAL) method in the adult male rat. Our results indicate that the BSTju displays a relatively simple projection pattern. First, it densely innervates the medial central amygdalar nucleus and the subcommissural zone and caudal anterolateral area of the BST - cell groups involved in visceromotor responses. Second, it provides inputs to the ventromedial caudoputamen (CP) and anterior basolateral amygdalar nucleus - areas presumably modulating somatomotor outflow. Third, the BSTju sends dense projections to the caudal substantia innominata, a distinct caudal dorsolateral region of the compact part of the substantia nigra, and the adjacent mesencephalic reticular nucleus and retrorubral area. And fourth, the BSTju provides light inputs to the prelimbic, infralimbic, and ventral CA1 cortical areas; to the posterior basolateral, posterior basomedial, and lateral amygdalar nuclei; to the paraventricular and medial mediodorsal thalamic nuclei; to the subthalamic and parasubthalamic nuclei of the hypothalamus; and to the ventrolateral periaqueductal gray. These projections, in part, suggest a role for the BSTju in circuitry integrating autonomic responses with somatomotor activity in adaptive behaviors. PMID- 10720610 TI - Effects of standard anticonvulsant drugs on different patterns of epileptiform discharges induced by 4-aminopyridine in combined entorhinal cortex-hippocampal slices. AB - Application of 4-aminopyridine (4-AP) has previously been reported to produce different patterns of epileptiform discharges in entorhinal cortex (EC) hippocampal slices: recurrent short discharges (RSDs) in hippocampal area CA1, seizure-like events (SLEs) and negative-going potentials (NGPs) in the medial entorhinal cortex (mEC). Using recordings of field potentials, we investigated the pharmacological effects of the clinically employed standard anticonvulsant drugs phenytoin (PHT), carbamazepine (CBZ), valproic acid (VPA) and phenobarbital (PHB) and those of pentobarbital (PB) on 4-AP-induced epileptiform activity. The anticonvulsant drugs showed different effects: SLEs were completely blocked by all tested drugs. Valproic acid, which suppressed all epileptiform activities, seemed to have the most fundamental effect of all drugs on 4-AP induced activity, because under phenytoin and carbamazepine, some epileptiform activity was still observable. The RSDs in hippocampal area CA1 of the hippocampus did not respond to the different anticonvulsants. In contrast, PB decreased the frequency of the RSDs in CA1 and enhanced the frequency of the NGPs in the EC. We propose that the activities induced by 4-AP in the combined entorhinal cortex-hippocampal slices may provide an in vitro model for the development of new drugs against difficult to-treat focal epilepsy. PMID- 10720611 TI - An adenoviral vector expressing the glucose transporter protects cultured striatal neurons from 3-nitropropionic acid. AB - Considerable interest has focused on the possibility of using gene transfer techniques to introduce protective genes into neurons around the time of necrotic insults. We have previously used herpes simplex virus amplicon vectors to overexpress the rat brain glucose transporter, Glut-1 (GT), and have shown it to protect against a variety of necrotic insults both in vitro and in vivo, as well as to buffer neurons from the steps thought to mediate necrotic injury. It is critical to show the specificity of the effects of any such transgene overexpression, in order to show that protection arises from the transgene delivered, rather than from the vector delivery system itself. As such, we tested the protective potential of GT overexpression driven, in this case, by an adenoviral vector, against a novel insult, namely exposure of primary striatal cultures to the metabolic poison, 3-nitropropionic acid (3NP). We observed that GT overexpression buffered neurons from neurotoxicity induced by 3NP. PMID- 10720612 TI - Postnatal development of NR1, NR2A and NR2B immunoreactivity in the visual cortex of the rat. AB - N-Methyl-D-aspartate receptors (NMDARs) are critically involved in some types of synaptic plasticity. The NMDAR subunits NR1, NR2A and NR2B are developmentally regulated, and it has been proposed that developmental changes in their expression may underlie developmental changes in cortical plasticity. Age dependent change in cortical plasticity is most commonly measured by the monocular deprivation effect, which occurs during a critical period between P22 and P50 in the rat. Although the development of NMDAR subunits has been studied from birth through the fourth postnatal week, there is only meager information from older ages when visual plasticity ends. We hypothesized that there will be significant age-dependent change in expression of NR1, NR2A or NR2B between P22, when the cortex is plastic, and P90, when it is not. We applied specific antibodies recognizing NR1, NR2A and NR2B to the primary visual cortex at P14, P22, P30, P45 and P90. We found age-dependent changes in NR1-IR that were negatively correlated with changes in NR2A-IR; these subunits are not regulated in unison. In contrast, NR2A-IR and NR2B-IR were positively correlated. NR2A-IR and NR2B-IR both passed through a developmental minimum around P45, then recovered to approximately their P22 level. NR1-IR passed through a maximum at P45. There were no significant differences between P22 and P90. These results do not support the simple hypothesis that the loss of plasticity corresponds to a simple transition from juvenile levels of NMDAR subunit proteins to new adult levels. On the other hand, the results do confirm the hypothesis that there are significant changes in processing of NMDAR proteins during the time that plasticity is lost. How these changes of IR relate to synaptic transmission and plasticity needs to be clarified. PMID- 10720613 TI - Involvement of M3 muscarinic receptors of the spinal cord in formalin-induced nociception in mice. AB - Subcutaneous injection of formalin into a paw of mice caused two distinct phases of licking and biting, first phase (1-5 min) and the second phase (7-30 min) after the injection. The muscarinic antagonist atropine (0.1-10 ng, i.t.) and the M(3) receptor antagonist 4-diphenylacetoxy-N-methylpiperidine methiodide (4-DAMP) (0.1-20 ng, i.t.) inhibited the second phase of this response, whereas higher doses of atropine (20-100 ng, i.t.) did not cause inhibition. The M(1) muscarinic receptor antagonist pirenzepine (10-100 ng, i.t.) did not inhibit either the first or the second phase response, but a high dose of pirenzepine (1000 ng, i.t.) tended to inhibit the second phase response. On the other hand, the M(2) muscarinic receptor antagonist 11-?(2-[(diethylamino)methyl]-1 piperidinyl?acetyl)-5, 11-dihydro-6H-pyrido(2,3-b)(1,4)benzodiazepine-6-one (AF DX116; 10-1000 ng, i.t.) had no effect on either the first or the second phase of response. The opioid receptor antagonist naloxone did not affect the 4-DAMP induced anti-nociceptive response. The i.t. injection of the acetylcholinesterase inhibitor neostigmine (25 ng) significantly inhibited only the second phase. The acetylcholine (ACh) depletor hemicholinium-3 (HC-3) (1 microg, i.t.) completely abolished the 4-DAMP-induced anti-nociceptive response. The ACh content of the spinal cord was significantly increased 14 min after formalin injection. This significant increase in the ACh content was inhibited by pretreatment with 4-DAMP (10 ng, i.t.). These results suggest that endogenous ACh in the spinal cord acts as a transmitter anti-nociception, and that ACh release regulated by presynaptic M(3) muscarinic receptors in the spinal cord is involved in the second phase of nociception induced by formalin. PMID- 10720614 TI - Suppression of acute and chronic opioid withdrawal by a selective soluble guanylyl cyclase inhibitor. AB - Previous studies have shown that activation of N-methyl-D-aspartate (NMDA) receptors and formation of nitric oxide (NO) contributes to the hyperactivity of locus coeruleus (LC) noradrenergic neurons and behavioural symptoms seen during opioid withdrawal. However, the role of soluble guanylyl cyclase (sGC), the 'physiological' target of NO, in this phenomenon is unclear. In this study, the effect of 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ), a highly selective sGC inhibitor, on the naloxone-precipitated morphine withdrawal was examined using differential normal pulse voltammetry (DNPV) to measure LC activity, in vivo microdialysis to measure glutamate/aspartate release response, and behavioural assessment to evaluate withdrawal symptoms. In halothane anaesthetized rats, acute intracerebroventricular (i.c.v.) morphine (10 microg) reduced the catecholamine oxidation current (CA.OC) (54.5+/-4.9% of baseline). Naloxone (2 mg/kg, i.v.) reversed this action of morphine and produced a rebound increase in CA.OC (136.1+/-6.0% of baseline), representing acute morphine withdrawal. Administration of ODQ (200 nmol, i.c.v.) blocked this response without affecting acute morphine action. In animals chronically treated with morphine (15 microg/microl/h, i.c.v., 5 days), naloxone significantly increased both the CA.OC signal (270.0+/-19.6% of baseline) and the release of L-glu (193+/ 30.4%) and L-asp (221.5+/-28.4%) above baseline. These responses were attenuated in animals pretreated with ODQ. In unanaesthetized chronic morphine dependent rats, ODQ treatment suppressed the signs of withdrawal precipitated by naloxone (10 mg/kg). Taken together, the results of this study suggest that sGC plays an intermediary role in the genesis of LC neuronal hyperactivity and behavioural signs of morphine withdrawal. PMID- 10720615 TI - Immunohistochemical localization of glial cell line-derived neurotrophic factor family receptor alpha-1 in the rat brain: confirmation of expression in various neuronal systems. AB - The localization of glial cell line-derived neurotrophic factor (GDNF) family receptor alpha-1 (GFRalpha-1) was investigated in rat brain by immunohistochemistry using a polyclonal antibody against a specific sequence of the rat protein. For raising the antisera in rabbits, we synthesized the oligopeptide SDVFQQVEHISKGN that corresponds to residues 139 to 152 of rat GFRalpha-1. On immunospot assay, 0.5 microg/ml of an affinity-purified antibody was capable of detecting 7.8 pmol of the rat GFRalpha-1 oligopeptides. When rat brain homogenates were examined by Western blots, the antibody revealed two main bands with molecular weights of approximately 47 kDa and 53 kDa, corresponding to the known sizes of GFRalpha-1. Immunohistochemistry in rat brain demonstrated that GFRalpha-1-like immunoreactivity was present in neurons but not in glial cells. The localization of GFRalpha-1-like immunoreactivity was largely consistent with that of the corresponding GFRalpha-1 mRNA. Positive neurons were distributed widely in various brain regions, but were particularly abundant in such regions as the olfactory bulb, diagonal band, substantia innominata, zona incerta, substantia nigra, cerebellar cortex, nuclei of the cranial nerves including auditory system and spinal motoneurons. The present study showed that GFRalpha-1 in the normal central nervous system is expressed preferentially in certain multiple neuronal systems that include cholinergic system as well as dopaminergic system and motor neurons. As GFRalpha-1 protein was found in numerous brain structures, GDNF family ligands may have therapeutic application not only in degenerative diseases affecting in specific nervous systems, such as Parkinson's disease, amyotrophic lateral sclerosis and multiple system atrophy, but in diffusely damaging diseases like cerebrovascular diseases. PMID- 10720616 TI - Intrathecal administration of an NMDA or a non-NMDA receptor antagonist reduces mechanical but not thermal allodynia in a rodent model of chronic central pain after spinal cord injury. AB - Spinal cord injuries (SCI) result in a devastating loss of function and chronic central pain syndromes frequently develop in the majority of these patients. The present study uses a rodent spinal hemisection model of SCI in which mechanical and thermal allodynia develops by 24 days after injury. Post-operative paw withdrawal responses to low threshold and high threshold mechanical stimuli compared to pre-operative responses (4.78, 9.96, and 49.9 mN) were increased and were statistically significant (p<0.05) for both forelimbs and hindlimbs indicating the development of mechanical allodynia. By contrast, post operatively, the temperature at which paw withdrawal accompanied by paw lick occurred was significantly decreased (p<0.05), indicating the development of thermal allodynia. The intrathecal application of either D-AP5, a competitive NMDA receptor antagonist, or NBQX-disodium salt, a competitive non-NMDA AMPA/kainate receptor antagonist, alleviated the mechanical allodynia and lowered the threshold of response for the high threshold mechanical stimuli in a dose dependent manner, and these decreases were statistically significant (p<0.05). By contrast, neither the D-AP5 nor the NBQX produced a statistically significant change in the thermal allodynia behavior in either forelimbs or hindlimbs in the hemisected group. No significant changes in locomotion scores, and thus no sedation, were demonstrated by the hemisected group for the doses tested. These data support the potential efficacy of competitive excitatory amino acid receptor antagonists in the treatment of chronic central pain, particularly where input from low threshold mechanical afferents trigger the onset of the painful sensation. Furthermore, these data suggest a role for both NMDA and non-NMDA receptors in the development of plastic changes in the spinal cord that provide the underlying mechanisms for central neuropathic pain. PMID- 10720617 TI - Descending projections of infralimbic cortex that mediate stimulation-evoked changes in arterial pressure. AB - The infralimbic cortex (IL) of the rat can modify autonomic nervous system activity, but the critical pathway(s) that mediate this influence are unclear. To define the potential pathways, the first series of experiments characterizes the descending projections of IL and the neighboring cortical areas using Phaseolus vulgaris leucoagglutinin (PHA-L). IL has prominent projections to the central nucleus of the amygdala (Ce), the mediodorsal nucleus of the thalamus (MD), the lateral hypothalamic area (LHA), the periaqueductal gray (PAG), the parabrachial nucleus (Pb), and the nucleus of the solitary tract (NTS). The density and selectivity of these projections suggest that the LHA and the PAG mediate the ability of the IL to regulate cardiovascular function. The second series of experiments demonstrates that locally anesthetizing neurons in either the LHA or PAG with lidocaine attenuates the hypotensive effects produced by electrical stimulation of the IL. Similarly, microinjections of cobalt chloride (a neurotransmission blocker) into the anterior portion of the LHA also decrease the arterial pressure responses to IL stimulation, suggesting that the ability of lidocaine to reversibly block the evoked response is due to inactivation of neurons in the LHA. These data indicate that hypotension evoked by stimulation of IL is mediated, at least in part, by direct or indirect projections to the LHA and through the PAG. PMID- 10720618 TI - Chronic cerebral hypoperfusion: loss of pupillary reflex, visual impairment and retinal neurodegeneration. AB - Adult rats underwent permanent bilateral occlusion of the common carotid arteries (2VO) to determine the effect of chronic cerebral ischemia on vision and retina. They were monitored post-surgically for the presence of the pupillary reflex to light. Some rats were tested for 6 months post-surgically on a radial arm maze task and then tested in another water-escape task which explicitly tested visual function. Another group of rats were tested post-surgically for 3 months on a task which simultaneously assessed visual and tactile discrimination ability. The thicknesses of the retinal sub-layers were then measured for some rats. Fourteen of the 25 rats that underwent 2VO lost the pupillary reflex. This seemed to occur within 5 days. Rats that lost the pupillary reflex but not rats whose reflex was intact, were impaired on all visually guided mazes. Tactile discrimination ability was unaffected. Only rats that lost the pupillary reflex showed reduced thickness of the retinal outer nuclear and plexiform layers, reduced cell density in the retinal ganglion cell layer and astrocytosis and degeneration of the optic tract. We conclude that 2VO can eliminate the pupillary reflex. Photoreceptors and retinal ganglion cells degenerate, but it is unclear if these are the cause(s) or result(s) of the loss of the pupillary reflex. These effects are accompanied by impairment of visually guided behavior. The possibility that visual system damage may also occur in acute ischemia merits further investigation. PMID- 10720619 TI - Altered release of prostaglandins contributes to hypoxic/ischemic impairment of NOC/oFQ cerebrovasodilation. AB - This study was designed to determine if altered release of prostaglandins contributes to impaired pial artery dilation to the newly described opioid, nociceptin/orphanin FQ (NOC/oFQ), following hypoxia/ischemia in newborn pigs equipped with a closed cranial window. Global cerebral ischemia (20 min) was induced via elevation of intracranial pressure, while hypoxia (10 min) decreased P(O(2)) to 35+/-3 mmHg with unchanged P(CO(2)). NOC/oFQ (10(-8) and 10(-6) M) modestly increased cerebrospinal fluid (CSF) 6-Keto PGF(1alpha) and TXB(2), the stable breakdown products of PGI(2) and TXA(2), in sham animals (1199+/-39 to 1704+/-104 and 299+/-9 to 409+/-12 pg/ml for control and 10(-6) M NOC/oFQ 6-Keto PGF(1alpha) and TXB(2), respectively). In 1 h post ischemia/reperfusion (I+R) animals, basal levels of 6-Keto PGF(1alpha) and TXB(2) were elevated. NOC/oFQ stimulated release of 6-Keto PGF(1alpha) was blocked while such release of TXB(2) was enhanced (526+/-15 to 822+/-36 pg/ml for control and 10(-6) M NOC/oFQ CSF TXB(2)). Similar, though more pronounced, changes were observed in hypoxia/ischemia/reperfusion (H+I+R) animals. Pretreatment with indomethacin (5 mg/kg i.v.) or SQ 29,548 (10(-4) M), cyclooxygenase and PGH(2)/TXA(2) receptor antagonists, partially restored attenuated NOC/oFQ pial artery dilation 1 h after I+R (9+/-1 and 18+/-1 vs. 3+/-1 and 6+/-1 vs. 8+/-1 and 13+/-1% for 10(-8) and 10(-6) M NOC/oFQ in sham, I+R, and I+R - SQ 29,548 pretreated animals). In contrast, NOC/oFQ-induced vasodilation was reversed to vasoconstriction in H+I+R animals and indomethacin or SQ 29,548 similarly partially restored such pial vasodilation. These data indicate that altered stimulated prostaglandin release contributes to hypoxic/ischemic impairment of NOC/oFQ-mediated pial artery dilation. PMID- 10720620 TI - Brain-derived TNFalpha: involvement in neuroplastic changes implicated in the conscious perception of persistent pain. AB - The pleiotropic cytokine tumor necrosis factor-alpha (TNFalpha) is implicated in the development of persistent pain through its actions in the periphery and in the central nervous system (CNS). Activation of the alpha(2)-adrenergic receptor is associated with modulation of pain, possibly through its autoregulatory effect on norepinephrine (NE) release in the CNS. The present study employs a chronic constriction nerve injury (CCI) pain model to demonstrate the interactive role of presynaptic sensitivity to TNFalpha and the alpha(2)-adrenergic autoreceptor in the pathogenesis of neuropathic pain. Accumulation of TNFalpha is increased initially in a region of the brain containing the locus coeruleus (LC) at day 4 post-ligature placement, followed by an increase in TNFalpha in the hippocampus at day 8 post-ligature placement, coincident with hyperalgesia. Levels of TNFalpha in the thoraco-lumbar spinal cord are also increased at day 8 post ligature placement. Concurrently, alpha(2)-adrenergic receptor and TNFalpha induced inhibition of NE release are increased, and stimulated NE release is decreased in superfused hippocampal slices isolated at day 8 post-ligature placement. Stimulated NE release is also decreased in spinal cord slices (lumbar region) from animals undergoing CCI, although in contrast to that which occurs in the hippocampus, alpha(2)-adrenergic receptor inhibition of NE release is not changed. These results indicate an important role that TNFalpha plays in adrenergic neuroplastic changes in a region of the brain that, among its many functions, appears to be a crucial link in the conscious perception of pain. We predict that neuroplastic changes, involving increased functional responses of alpha(2)-adrenergic autoreceptors and increased presynaptic sensitivity to TNFalpha, culminate in decreased NE release in the CNS. These neuroplastic changes provide a mechanism for the role of CNS-derived TNFalpha in the pathogenesis of persistent pain. PMID- 10720621 TI - Phytoestrogens decrease brain calcium-binding proteins but do not alter hypothalamic androgen metabolizing enzymes in adult male rats. AB - Phytoestrogen [plant estrogenic-like molecule(s)] research has grown rapidly in recent years due to their potential health benefits. However, little is known about phytoestrogen's effects on the CNS. Androgen metabolizing enzymes are known to regulate neuroendocrine functions and reproductive behaviors, while calcium binding proteins are associated with protecting against neurodegenerative diseases. Therefore, we examined aromatase and 5alpha-reductase enzyme activities in the medial basal hypothalamic and preoptic area (mbh-poa) and characterized mbh-poa and amygdala (amy) calbindin and calretinin levels (via Western analysis) from animals fed a phytoestrogen-free (P-free) vs. a phytoestrogen-containing diet [(P-600); that had 600 microg/g of phytoestrogens]. After approximately 5 weeks on the diets, the male rats were killed at 105 days. P-600 plasma phytoestrogen levels were 78-fold higher than the P-free values and the mbh-poa phytoestrogen content was 8-fold higher than the P-free group, demonstrating the passage of phytoestrogens into brain. In general, brain aromatase or 5alpha reductase activity levels were not significantly altered by the experimental diets. However, independent of brain site (i.e., mbh-poa or amy) the abundance of calbindin from male P-600 rats was significantly lower than P-free animals. Conversely, for calretinin there were no significant alterations in the mbh-poa tissue site, while in the amy a similar pattern of expression was seen to that of the calbindin results. These data suggest that consumption of phytoestrogens via a soy diet for a relatively short interval can significantly: (1) elevate plasma and brain phytoestrogens levels and (2) decrease brain calcium-binding proteins without altering brain androgen metabolizing enzymes. PMID- 10720623 TI - Predominant activation of I1-waves from the leg motor area by transcranial magnetic stimulation. AB - We performed transcranial magnetic stimulation (TMS) to elucidate the D- and I wave components comprising the motor evoked potentials (MEPs) elicited from the leg motor area, especially at near-threshold intensity. Recordings were made from the tibialis anterior muscle using needle electrodes. A figure-of-eight coil was placed so as to induce current in the brain in eight different directions, starting from the posterior-to-anterior direction and rotating it in 45 degrees steps. The latencies were compared with those evoked by transcranial electrical stimulation (TES) and TMS using a double cone coil. Although the latencies of MEPs ranged from D to I3 waves, the most prominent component evoked by TMS at near-threshold intensity represented the I1 wave. With the double cone coil, the elicited peaks always represented I1 waves, and D waves were evoked only at very high stimulus intensities, suggesting a high effectiveness of this coil in inducing I1 waves. Using the figure-of-eight coil, current flowing anteriorly or toward the hemisphere contralateral to the recorded muscle was more effective in eliciting large responses than current flowing posteriorly or toward the ipsilateral hemisphere. The effective directions induced I1 waves with the lowest threshold, whereas the less effective directions elicited I1 and I2 waves with a similar frequency. Higher stimulus intensities resulted in concomitant activation of D through I3 waves with increasing amount of D waves, but still the predominance of I1 waves was apparent. The amount of I waves, especially of I1 waves, was greater than predicted by the hypothesis that TMS over the leg motor area activates the output cells directly, but rather suggests predominant transsynaptic activation. The results accord with those of recent human epidural recordings. PMID- 10720622 TI - Inhibition of spinal nociceptive responses after intramuscular injection of capsaicin involves activation of noradrenergic and opioid systems. AB - Extracellular recordings of wide dynamic range neurones in the dorsal horn driven by electrical stimulation of the sciatic nerve were performed in intact urethane anaesthetized Sprague-Dawley rats. The electrically evoked neuronal responses were defined as A- and C-fibres responses according to latencies, and the effect of a deep nociceptive conditioning stimulus induced by 200 microg capsaicin (8 methyl-N-vanillyl-6-noneamide) injected into the contralateral gastrocnemius soleus muscle was studied for at least 30 min. Independent of the size and location of the receptive field of the neurone under study, a clear inhibition of the neuronal responses was observed. The electrically evoked C-fibre responses were inhibited to 53% of baseline 15-30 min after injection of capsaicin. This inhibition was only slightly attenuated by 125 nmol of the alpha-adrenoceptor antagonist phentolamine or 250 nmol of the opioid receptor antagonist naloxone applied directly onto the spinal cord when the two compounds were administered separately 5 min before capsaicin. In contrast, when a mixture of the two compounds was given 5 min before capsaicin, the effect of capsaicin was completely abolished. These results indicate that activation of the capsaicin sensitive afferents in the gastrocnemius-soleus muscle inhibits the electrically evoked C-fibre responses in the dorsal horn by activating noradrenergic and opioidergic inhibitory systems. Moreover, our data indicate that the activation of these two systems following injection of capsaicin has a sub-additive inhibitory effect on the wide dynamic range neurones in the spinal cord. We conclude that only one of these systems is sufficient for the inhibition to occur. PMID- 10720624 TI - NMDA receptor antagonist-induced dopamine release in the ventral pallidum does not correlate with motor activation. AB - The ventral pallidum is the output structure of the nucleus accumbens in the ventral corticostriato-thalamocortical loop. Information processing in this loop is critically involved in motor behavior and reinforcement. The ventral pallidum receives a direct dopaminergic input from the ventral tegmental area, but also glutamatergic input from cortical and limbic areas. It has been assumed that dopamine release in the VP is indeed modulated by glutamate. The present study investigated the effects of NMDA receptor blockade on motor behavior and dopamine release in the ventral pallidum. In a first experiment, rats were implanted with microdialysis probes in the ventral pallidum and were systemically injected or locally perfused via the microdialysis probe with dizocilpine (0.32 mg/kg, 10 and 100 microM, respectively). Effects on dopamine and on locomotion were simultaneously monitored. In a second experiment, ventral pallidum was lesioned by quinolinic acid and the effects of systemic dizocilpine (0.08 and 0.16 mg/kg) on locomotion and stereotyped sniffing behavior were determined. It was found that systemic and local dizocilpine administration increased dopamine release in the ventral pallidum to a similar extent whereas only systemic treatment was accompanied by locomotor stimulation. Lesion of the ventral pallidum did not affect locomotion and stereotyped sniffing behavior induced by systemic dizocilpine treatment. Thus, DA release in the ventral pallidum that is elevated by blockade of NMDA receptors is not relevant for activation of motor behavior. PMID- 10720625 TI - Behavioral and ultrastructural changes induced by chronic neuroinflammation in young rats. AB - We investigated the ultrastructural, immunohistochemical, biochemical and behavioral effects of chronic neuroinflammation in young rats produced by injection of lipopolysaccharide (LPS) into the 4th ventricle. The 37-day infusion of LPS impaired spatial memory but not object recognition ability. Electron microscopic studies of neurons within the hippocampus identified numerous paired cisternae of the rough endoplasmic reticulum (RER) and other ultrastructural changes that suggested impaired or reduced synthesis of cellular proteins within the cytoplasm. Immunohistochemical staining found numerous highly activated microglia distributed throughout the cingulate gyrus, entorhinal cortex, hippocampus and dentate gyrus. This animal model may be useful to test potential pharmacotherapies that are directed at the prevention of the cytotoxic consequences of chronic neuroinflammation associated with normal aging or Alzheimer's disease. PMID- 10720626 TI - Angiotensin II increases intracellular Ca(2+) concentration in folliculo-stellate cells of the rat anterior pituitary in primary culture. AB - It has been reported that pituitary gonadotrophs and lactotrophs contain angiotensin II (Ang II) and suggested that Ang II modulates hormone secretion from endocrine cells of the anterior pituitary through paracrine mechanism among the endocrine cells. However, there has been no report on the effect of Ang II on the folliculo-stellate cells (FSC) which are thought to play a regulatory role in the release of hormones from pituitary endocrine cells. We, therefore, examined the effect of Ang II on FCS in primary culture by Ca(2+) imaging technique. Certain proportion (42%) of FSC responded to 100 nM Ang II by increasing [Ca(2+)](i). In addition, Ang II elicited the Ca(2+) response in about 50% of the pituitary endocrine cells. The results indicate that Ang II functions as a paracrine factor among pituitary cells including FSC. PMID- 10720627 TI - HMG-CoA reductase inhibitor induces a transient activation of high affinity nerve growth factor receptor, trk, and morphological differentiation with fatal outcome in PC12 cells. AB - The present study was aimed at investigating the possible toxicity of simvastatin on a neuronal cell line, PC12 cells. Simvastatin clearly induced a transient morphological differentiation as evidenced by the occurrence of neurite outgrowth with a transient activation of the high affinity nerve growth factor receptor, Trk, but died at 36 h after its addition. Tyrosine autophosphorylation of the Trk protein also disappeared at 36 h after addition. During the morphological differentiation, NGF mRNA expression was upregulated transiently and returned to the basal level at 36 h after addition of simvastatin. These results suggest that simvastatin is neurotoxic and PC12 cells elicited a protective response, involving a transient activation of a Trk-mediated intracellular signal transduction pathway by an autocrine secretion of NGF, although these responses did not persist against pro-apoptotic signals and resulted in an apoptosis of the PC12 cells. PMID- 10720628 TI - N-acetylcysteine elicited increase in complex I activity in synaptic mitochondria from aged mice: implications for treatment of Parkinson's disease. AB - It has been suggested that thiolic groups are essential for complex I activity and other respiratory mitochondrial enzymes. Recent experiments showed that the thiolic antioxidant N-acetylcysteine (NAC) can protect against age-related decrease in complex I activity in mice hepatic mitochondria. The present paper shows that NAC enhances complex I activity in vitro in synaptic mitochondria isolated from old mice. The optimum NAC concentration for maximum complex I activity was 10 mM in old synaptic preparations. Our data suggest that mitochondrial thiolic groups, which are essentials to oxidative phosphorylation, are impaired by aging. Based on the finding of decreased mitochondrial complex I activity in the substantia nigra of patients with Parkinson's disease, we propose that the thiol-containing antioxidant NAC could be beneficial for treatment of the disease. PMID- 10720629 TI - Inhibition of antithrombin by protein SV-IV normalizes the coagulation of hemophilic blood. AB - The aim of the study was to evaluate the effect of the protein Seminal Vesicle Protein No. 4 (SV-IV), a potent inhibitor of antithrombin III (antithrombin), on the coagulation of blood obtained from patients affected by hemophilia A. In the coagulating blood of these patients, the antithrombin/thrombin ratio was found to be markedly higher (about 44) than in normal individuals (about 4. 4). This high ratio was related to the low efficiency of thrombin-generating reactions induced by the factor VIII deficiency and to the high levels of free (not bound to serine proteases) antithrombin present in the hemophilic serum (antithrombin concentration was the same in normal and hemophilic plasma). The elevated concentration of free antithrombin in hemophiliacs was primarily a consequence of a reduced consumption caused by the scarce availability in the hemophilic serum of factors Xa and IIa, which are serine proteases possessing strong binding affinity for antithrombin. Addition of SV-IV to coagulating hemophilic blood reduced markedly the serum antithrombin and thrombin-antithrombin complexes, normalizing, as a consequence, the clotting time and other coagulation parameters. Similar results were obtained by using appropriate concentration of factor VIII. PMID- 10720630 TI - Modulation of the Ca(2+) release channel of sarcoplasmic reticulum by amiloride analogs. AB - Dichlorobenzamil, phenamil and other amiloride analogs (1-100 microM) elicit transient tension in rabbit skinned muscle fibers. Tension requires preloading of Ca(2+) into the sarcoplasmic reticulum, is facilitated by low-[Mg(2+)] solutions, abolished by ruthenium red or by functional disruption of the sarcoplasmic reticulum, and is followed by inhibition of the caffeine-evoked tension. Bilayer recording of Cs(+) currents through the sarcoplasmic reticulum Ca(2+) release channel reveals that phenamil (10-100 microM) increases the open channel probability, whereas dichlorobenzamil affects the channel activity in a complex concentration- and time-dependent manner: stimulation occurs throughout exposure to 10 microM, but is followed by channel blockade when 100 microM dichlorobenzamil is used. It is concluded that stimulation of the sarcoplasmic reticulum Ca(2+) release channel accounts for the dichlorobenzamil- or phenamil induced tension in skinned fibers, whereas depletion of sarcoplasmic reticulum Ca(2+) stores and channel block (with dichlorobenzamil) explains the inhibition of the caffeine-evoked tension by amiloride analogs. PMID- 10720631 TI - Extracellular adenosine deprivation induces epithelial differentiation of HT29 cells: evidence for a concomitant adenosine A(1)/A(2) receptor balance regulation. AB - HT29 cells display an undifferentiated phenotype in culture. However, numerous treatments are able to induce both epithelial differentiation and cell growth inhibition. We have previously demonstrated that adenosine and its analogues act through specific adenosine receptors to modulate cell proliferation in HT29 and other human colon adenocarcinoma cell lines. Among the treatments tested, the most potent inhibition of HT29 cell growth was induced by deprivation of extracellular adenosine using adenosine deaminase. Here, we investigated the capacity of adenosine deaminase to initiate epithelial differentiation. After 1 month of daily addition of 10 U/ml adenosine deaminase to the culture medium, HT29 cells were cloned by limited dilution. Among the clones obtained, we focused our attention on clone 13. Microscopic visualization and proliferation studies indicated that cells from this clone grew very slowly and in a pseudo-monolayer, in marked contrast with the situation observed in the mother HT29 cell line. In addition, clone 13 cells displayed epithelial features that mimic the enterocytic differentiation of Caco-2 cells. These modifications were accompanied by dramatic changes in the activity of adenosine receptors, as demonstrated by pharmacological studies. In contrast to the original HT29 cells, clone 13 as well as Caco-2 cells displayed (i) a very low number of adenosine A(1) receptors, and (ii) increases in intracellular cAMP levels when challenged with adenosine analogues. It is hypothesized that a loss of adenosine A(1) receptors, with no change or a concomitant increase in adenosine A(2) receptors, results in the emergence of adenosine A(2) receptor-mediated differentiation and inhibition of proliferation, through a cAMP-dependent pathway. PMID- 10720632 TI - Inhibition of inducible nitric oxide synthase improves graft function and reduces tubulointerstitial injury in renal allograft rejection. AB - Increased levels of nitric oxide (NO) are found in rejecting renal allografts. Inducible NO synthase (iNOS) in infiltrating monocytes/macrophages could lead to NO bursts. NO may modulate the inflammatory response of early rejection due to its high reactivity with superoxide to yield peroxynitrite. To define the role of iNOS in acute renal allograft, rejection effects of the specific iNOS blockers iminoethyl-lysine and 7-butylhexahydro-1H-azepin-2-imine, monohydrochloride on renal function and morphology were investigated in renal allografts. Lewis rats received Brown Norway grafts with one kidney left in situ. All recipients were treated with low dose cyclosporine-A (2.5 mg/kg BW/day s.c.) to allow moderate rejection. In addition, one group received iminoethyl-lysine (10 mg/kg BW/day gavage) and one group received butylhexahydro-azepin-imine (3.4 mg/kg BW/day i.p.). Sham operated Brown Norway donor rats served as baseline controls. Compared to controls, low dose cyclosporine-A decreased glomerular filtration rate (P<0.05) and numerically increased renal vascular resistance. Adding iminoethyl-lysine to cyclosporine-A improved renal hemodynamics. Adding butylhexahydro-azepin-imine to cyclosporine-A practically restored glomerular filtration rate and renal vascular resistance (P<0.05) to control levels. Grafts treated with cyclosporine-A alone showed vascular, glomerular and tubulointerstitial lesions. Adding iminoethyl-lysine or butylhexahydro-azepin imine to cyclosporine-A did not significantly reduce vascular and glomerular injury, but diminished tubulointerstitial injury as well as nitrotyrosine staining in tubular epithelium (P<0.05). Thus, adding the iNOS blockers iminoethyl-lysine or butylhexahydro-azepin-imine to cyclosporine-A improved graft function and reduced tubulointerstitial lesions. PMID- 10720633 TI - Hypertrophic growth of cultured neonatal rat heart cells mediated by vasopressin V(1A) receptor. AB - Primary cultures of neonatal cardiac myocytes were used to determine both the identity of second messengers that are involved in vasopressin receptor-mediated effects on cardiac hypertrophy and the type of vasopressin receptor that is involved in vasopressin-induced cell growth. Neonatal rat myocytes were plated at a density of 1x10(6) cells per 60 mm dish and were incubated with serum-free medium for 7 days. Treatment of myocytes with vasopressin significantly increased the RNA-to-DNA ratio, by 18-25%, at culture days 4-6 and the protein-to-DNA ratio by 18-20% at culture days 5-7. Rates of protein synthesis were determined to assess their contribution to protein contents during myocyte growth. Vasopressin significantly accelerated rates of protein synthesis by 25% at culture day 6. Intracellular free Ca(2+) ([Ca(2+)](i)) was transiently increased after vasopressin exposure. After the peak increase in [Ca(2+)](i) at less than 30 s, there was a sustained increase for at least 5 min. The specific activity of protein kinase C in the particulate fraction was increased rapidly after exposure to vasopressin, and its activity remained higher for 30 min, returning to its control level within 60 min. The activity of protein kinase C in the cytosol was significantly decreased at all times after exposure to vasopressin. After vasopressin treatment, the content of c-fos mRNA was increased. The stimulatory effects of vasopressin on these parameters were significantly inhibited by vasopressin V(1A) receptor antagonist, OPC-21268, but not by vasopressin V(2) receptor antagonist, OPC-31260. These results suggest that vasopressin directly induces myocyte hypertrophic growth via the V(1A) receptor in neonatal rat heart cells. PMID- 10720634 TI - The analgesic effect profile of FR122047, a selective cyclooxygenase-1 inhibitor, in chemical nociceptive models. AB - The pharmacological profile of the analgesic agent, 1-[(4, 5-bis(4-methoxyphenyl) 2-thiazoyl)carbonyl]-4-methylpiperazine hydrochloride (FR122047), was investigated. In recombinant human cyclooxygenase enzyme assays, the inhibition of prostaglandin E(2) formation by FR122047 was 2300 times more selective for cyclooxygenase-1 than cyclooxygenase-2. Oral administration of FR122047 (3.2-100 mg/kg) dose dependently reduced the phase 2 response (10-60 min) of the formalin test in rats. This effect was 3 times less potent than that of indomethacin. FR122047 (1-32 mg/kg; p. o.) showed a dose-dependent analgesic effect against the acetic acid-induced writhing response in mice. Furthermore, FR122047 (0. 01-10 mg/kg, p.o.) inhibited the increase in 6-keto prostaglandin F(1alpha) level in acetic acid-injected mouse peritoneal cavity. However, a selective cyclooxygenase 2 inhibitor, NS-398, had no effect in these cyclooxygenase-1 sensitive pain models. These results suggest that FR122047, a selective cyclooxygenase-1 inhibitor, shows an analgesic effect in chemical nociceptive models and may be a useful analgesic agent. PMID- 10720635 TI - delta-opioid receptors and nitric oxide mediate the analgesic effect of Crotalus durissus terrificus snake venom. AB - The antinociceptive effect of Crotalus durissus terrificus venom was investigated in a model of inflammatory hyperalgesia induced by carrageenin. The rat paw pressure test was applied before and 3 h after the intraplantar (i.pl.) injection of carrageenin. The venom administered per os before and 1 or 2 h after carrageenin blocked hyperalgesia. When carrageenin was injected in both hind paws and naloxone into one hind paw, antinociception was abolished only in the paw injected with naloxone. D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr amide (CTOP) and nor binaltorphimine, antagonists of micro- and kappa-opioid receptors, respectively, did not alter the effect of the venom. N,N-diallyl-Tyr-Aib-Aib-Phe-Leu (ICI 174,864), an antagonist of delta-opioid receptors, antagonised this effect. Prolonged administration of the venom did not induce tolerance to this antinociceptive effect. N(G)-methyl-L-arginine (L-NMMA) and methylene blue, inhibitors of nitric oxide synthase and soluble guanylate cyclase, respectively, injected i.pl., antagonised antinociception. These data indicate that both delta opioid receptors and nitric oxide participate in the mediation of the peripheral antinociceptive effect of C. durissus terrificus venom. PMID- 10720636 TI - Haloperidol-induced catalepsy is absent in dopamine D(2), but maintained in dopamine D(3) receptor knock-out mice. AB - We have previously found that mice homozygous for the deletion of the dopamine D(2) receptor gene (D(2)(-/-) mice) do not present spontaneous catalepsy when tested in a "bar test". In the present study, we sought to analyse the reactivity of D(2) receptor mutant mice to the cataleptogenic effects of dopamine D(2)-like or D(1)-like receptor antagonists. In parallel, we assessed the cataleptogenic effects of these antagonists in dopamine D(3) receptor mutant mice. D(2)(-/-) mice were totally unresponsive to the cataleptogenic effects of the dopamine D(2) like receptor antagonist haloperidol (0.125-2 mg/kg i.p.), while D(2)(+/-) mice, at the highest haloperidol doses tested, showed a level of catalepsy about half that of wild-type controls. The degree of haloperidol-induced catalepsy was thus proportional to the level of striatal dopamine D(2) receptor expression (0.50, 0.30 and 0.08 pmol/mg protein as measured at 0.25 nM [3H]spiperone for D(2)(+/+), D(2)(+/-) and D(2)(-/-) mice, respectively). However, D(2)(-/-) and D(2)(+/-) mice were as sensitive as their wild-type counterparts to the cataleptogenic effects of the dopamine D(1)-like receptor antagonist R-(+)-7-chloro-8-hydroxy-3 methyl-1-phenyl-2,3,4, 5-tetrahydro-1H-3-benzazepine hydrochloride (SCH 23390: 0.03-0.6 mg/kg s.c.). Striatal dopamine D(1) receptor expression (as measured using [3H]SCH 23390 binding) was not significantly affected by the genotype. The ability of SCH 23390 to induce catalepsy in D(2)(-/-) mice suggests that their resistance to haloperidol-induced catalepsy is due to the absence of dopamine D(2) receptors, and not to the abnormal striatal synaptic plasticity that has been shown by others to occur in these mice. In agreement with the observation that dopamine D(2) and dopamine D(1) receptor expression was essentially identical in D(3)(+/+), D(3)(+/-) and D(3)(-/-) mice, dopamine D(3) receptor homozygous and heterozygous mutant mice, on the whole, did not differ from their controls in the time spent in a cataleptic position following administration of either haloperidol (0.5-2 mg/kg i.p.) or SCH 23390 (0.03-0.6 mg/kg s.c.). Also, dopamine D(3) receptor mutant mice were no more responsive than wild-type controls when co-administered subthreshold doses of haloperidol (0.125 mg/kg) and SCH 23390 (0.03 mg/kg), suggesting that dopamine D(3) receptor knock-out mice are not more sensitive than wild-types to the synergistic effects of concurrent blockade of dopamine D(2) and dopamine D(1) receptors in this model. These results suggest that the dopamine D(2) receptor subtype is necessary for haloperidol to produce catalepsy, and that the dopamine D(3) receptor subtype appears to exert no observable control over the catalepsy produced by dopamine D(2)-like, D(1)-like and the combination of D(1)-like and D(2)-like receptor antagonists. PMID- 10720637 TI - Effects of GYKI 52466 and some 2,3-benzodiazepine derivatives on hippocampal in vitro basal neuronal excitability and 4-aminopyridine epileptic activity. AB - In order to determine whether the anticonvulsant effect of 2, 3-benzodiazepines is also displayed in a model of in vitro epilepsy, such as the "epileptiform" hippocampal slice, we studied the effects of 2,3-benzodiazepine 1-(4-aminophenyl) 4-methyl-7, 8-methylenedioxe-5H 2,3-benzodiazepine hydrochloride (GYKI 52466) and some new 2,3-benzodiazepine derivatives on CA1 basal neuronal excitability and on CA1 epileptiform burst activity produced by 4-aminopyridine in rat hippocampal slices. The results showed that GYKI 52466 affected basal neuronal excitability as evidenced by its influence on the magnitude of the CA1 orthodromic-evoked field potentials. 2,3-Benzodiazepines showed their antiepileptic effect also in an in vitro model of experimental epilepsy. The effects of the new 2,3 benzodiazepine derivatives suggest that the methylenedioxidation in positions 7 and 8 of the 2,3-benzodiazepine ring is the main structural modification for the antiepileptic effect of 2,3-benzodiazepines to take place. PMID- 10720638 TI - Role of histamine H(1) receptor in pain perception: a study of the receptor gene knockout mice. AB - To study the participation of histamine H(1) receptors in pain perception, histamine H(1) receptor knockout mice were examined for pain threshold by means of three different kinds of nociceptive tasks. These included assays for thermal nociception (hot-plate, tail-flick, paw-withdrawal), mechanical nociception (tail pressure), and chemical nociception (abdominal constriction, formalin test, capsaicin test) which evoked pain by the activation in nociceptive Adelta and C fibers. The mutant mice lacking histamine H(1) receptors showed significantly fewer nociceptive responses to the hot-plate, tail-flick, tail-pressure, paw withdrawal, formalin, capsaicin, and abdominal constriction tests. Sensitivity to noxious stimuli in histamine H(1) receptor knockout mice significantly decreased when compared to wild-type mice. This data indicates that histamine plays an important role in both somatic and visceral pain perceptions through histamine H(1) receptors. The difference in the effect of histamine H(1) receptors antagonist, the active (D-) and inactive (L-) isomers of chlorpheniramine on ICR mice further substantiates the evidence of the role of histamine H(1) receptors on pain threshold. PMID- 10720639 TI - The antinociceptive effects of endomorphin-1 and endomorphin-2 in diabetic mice. AB - The antinociceptive effects of endomorphin-1 and endomorphin-2, endogenous mu opioid receptor agonists, were examined using the tail-flick test in non-diabetic and diabetic mice. Endomorphin-1, at doses of 1 to 10 microg, i.c.v., and endomorphin-2, at doses of 3 to 30 microg, i.c.v., each dose dependently inhibited the tail-flick response in both non-diabetic and diabetic mice. There was no significant difference between the antinociceptive effects of endomorphin 1 in non-diabetic mice and diabetic mice. The antinociceptive effect of endomorphin-2 was greater in non-diabetic mice than in diabetic mice. In non diabetic mice, the antinociceptive effects of endomorphin-1 and endomorphin-2 were significantly reduced by beta-funaltrexamine, a mu-opioid receptor antagonist, and naloxonazine, a selective mu(1)-opioid receptor antagonist, but not by naltrindole, a delta-opioid receptor antagonist, or nor-binaltorphimine, a kappa-opioid receptor antagonist. In diabetic mice, the antinociceptive effect of endomorphin-2 was significantly reduced by beta-funaltrexamine and naloxonazine. However, these micro-opioid receptor antagonists had no significant effect on the antinociceptive effect of endomorphin-1 in diabetic mice. The antinociception induced by endomorphin-1 in diabetic mice was significantly reduced by naltrindole and 7-benzylidenenaltrexon, a selective delta(1)-opioid receptor antagonist, administered i.c.v. However, nor-binaltorphimine had no significant effect on the antinociceptive effects of endomorphin-1 and endomorphin-2 in diabetic mice. These results indicate that the antinociceptive effects of endomorphin-1 and endomorphin-2 in non-diabetic mice are mediated through the activation of mu(1)-opioid receptors, whereas in diabetic mice, endomorphin-1 and endomorphin-2 may produce antinociception through different actions at delta(1)- and mu(1)-opioid receptors, respectively. PMID- 10720640 TI - Analgesic effect of thalidomide on inflammatory pain. AB - Tumor necrosis factor alpha (TNF-alpha) may have a pivotal role in the genesis of mechanical inflammatory hyperalgesia in rats and in the nociceptive writhing response in mice. Thalidomide has been shown to selectively inhibit TNF-alpha production. We therefore investigated the effect of thalidomide on these responses as well as on the hot plate response in mice. Hyperalgesic responses to intraplantar (i.pl.) injections of carrageenin or bradykinin, which act by stimulating TNF-alpha release, but not responses to TNF-alpha or prostaglandin E(2), were inhibited in a dose-dependent manner by pretreatment of the animals with thalidomide. The nociceptive writhing responses induced by intraperitoneal (i.p.) injections of zymosan or acetic acid were also inhibited in a dose dependent manner by pretreatment of mice with thalidomide. Moreover, the thalidomide pretreatment also reduced the TNF-alpha mRNA levels in the peritoneal cells induced by injection of zymosan in mice. The analgesic effect of thalidomide is not due to a central effect, since the drug had no effect in the hot plate test. The demonstration that thalidomide is able to inhibit inflammatory hyperalgesia in rats and the writhing nociceptive response in mice suggests that these analgesic effects seem to be a consequence of the inhibition of TNF-alpha production, and indicates the need for investigations on the possibility of the use of thalidomide for the treatment of pain refractory to classical non-narcotic analgesics. PMID- 10720641 TI - Effects of scopolamine in comparison with apomorphine and phencyclidine on prepulse inhibition in rats. AB - The potential involvement of the muscarinic cholinergic system in the underlying mechanisms of prepulse inhibition of the acoustic startle reflex was evaluated in male Sprague-Dawley rats under conditions of varying dose, prepulse intensity, and interstimulus interval. The effects of scopolamine on prepulse inhibition were also directly compared with the effects observed using apomorphine and phencyclidine under the same test parameters. Scopolamine (0. 03-1.0 mg/kg) produced a significant dose-dependent decrease in prepulse inhibition, but had no effect on startle amplitude over the dose range tested. Apomorphine (0.03-1.0 mg/kg) and phencyclidine (0. 1-5.6 mg/kg) produced significant dose-dependent decreases in prepulse inhibition and changes in startle amplitude. The scopolamine-induced decrease in prepulse inhibition varied with prepulse intensity in that the changes produced by scopolamine became smaller in magnitude as the prepulse intensity was increased from 9 to 30 dB above background. On the other hand, apomorphine and phencyclidine decreased prepulse inhibition to approximately the same magnitude across all prepulse intensities tested. The observed decreases in prepulse inhibition produced by scopolamine, apomorphine, and phencyclidine were also dependent on interstimulus interval duration. Scopolamine produced marked decreases in prepulse inhibition at the 100- and 300 ms interstimulus interval durations, but had little or no effect on prepulse inhibition at the 30- and 1000-ms interstimulus interval durations. In contrast, apomorphine decreased prepulse inhibition across all interstimulus interval durations while phencyclidine decreased prepulse inhibition across the 30- to 300 ms interstimulus interval durations. The present findings support the hypothesis that the muscarinic cholinergic system, like the dopaminergic and glutamatergic systems, is directly involved in the mechanisms of prepulse inhibition. However, these three neurotransmitter systems appear to modulate different aspects of prepulse inhibition. PMID- 10720642 TI - Ondansetron modulates pharmacodynamic effects of ketamine on electrocardiographic signals in rhesus monkeys. AB - Electrocardiographic signal dynamics were examined in rhesus monkeys (Macaca mulatta) before and after treatment with ketamine and/or ondansetron. Ketamine exerts differential pharmacodynamic effects on behavior in animals stratified according to a measure of central serotonergic turnover. We hypothesized that measures of serotonergic turnover might explain some of the variance in the electrocardiographic (ECG) response to ketamine. Electrocardiographic recordings of animals were obtained at baseline, after administration of either saline or ondansetron (0.125 mg/kg), and after administration of ketamine (15 mg/kg). Electrocardiographic signal dynamics were measured using an algorithm that extracts the Hurst parameter (H) of the interbeat interval (IBI) time-series. H decreased after ketamine administration, (mean+/-S.E.M.), 0.33+/-0.04 vs. 0.12+/ 0.02, P3.5 cm long. All fistulae were first treated with the lavage of the fistulous tract with antibiotic solution until a sterile microbiological finding was obtained. This was followed by electrocoagulation of the fistulous tract with a special probe for the eradication of granulomatous tissue. Finally the fibrin glue-antibiotic mixture (Tisseel, Immuno Ltd., Vienna, Austria) was applied. RESULTS: After a follow-up of 18-36 months (median 28) 18 patients (26%) had a recurrence; among these, intersphincteric fistula recurred in 9 patients (23%) and transsphincteric also in 9 (30%). Regarding the length of the fistulous tract, a fistula with a 3.5 cm long tract in 5 (11%). CONCLUSION: The analysis showed that the success of the treatment of anal fistulae with fibrin glue-antibiotic mixture was independent of the vertical disposition of the fistula, and was dependent on the length of the fistulous tract. Surgical treatment remains a golden standard for simple fistulae with a tract 0.05). Our data for the first time confirmed that the presence of incisional hernia is accompanied by impaired collagen synthesis in the skin. The decreased tensile strength of collagen type III may play a key role in the development of incisional hernias. Furthermore, it might explain the high recurrence rates of hernia repair by simple closure, as a repetition of the primarily failing technique, and the improvement by the additional use of alloplastic material. PMID- 10720846 TI - Surgery of liver tumors: state of the art. scientific meeting, munich, may 13-15, 1999 PMID- 10720847 TI - Why is urological laparoscopy minimally invasive? AB - OBJECTIVES: Laparoscopic procedures have been developed and established with the view that a similar operative effect can be achieved with less traumatization, especially as far as systemic stress response is concerned. We report a prospective, controlled, nonrandomized animal and patient study to determine the systemic response to laparoscopic and open surgical procedures. METHODS: In the animal study, 26 female pigs underwent either a laparoscopic bilateral varix ligation followed by bilateral nephrectomy (group I), sole introduction of trocars (group II) or sole establishment of an open surgical approach (group III). In the patient study, 145 patients underwent various laparoscopic procedures (nephrectomy, renal cyst marsupialization, varix ligation), open surgical procedures (nephrectomy, inguinal orchiectomy) or extracorporeal shockwave lithotripsy (ESWL). The serum parameters interleukin (IL)-6, IL-10 and C-reactive protein (CRP) were measured before, during and after the operative procedure. RESULTS: In animals and patients, laparoscopy resulted in significantly lower serum levels of CRP during and after the operative procedure. Animals in group I showed a 5-fold elevation, in group II a 3-fold elevation and in group III a 9-fold elevation of CRP. In patients, the increase of CRP was twice as high after open unilateral nephrectomy than after laparoscopic unilateral or bilateral nephrectomy. IL-6 showed less marked elevation during laparoscopy, ESWL and minor operative procedures like laparoscopic varix ligation or inguinal orchiectomy when compared to an open unilateral nephrectomy. The parameter IL-10 showed no significant differences among the patient groups. CONCLUSIONS: The extent of the acute phase reaction to the operative trauma correlates much more convincingly to the approach than to the extent of the procedure. Only larger operations like nephrectomy trigger a systemic acute phase reaction, which can be limited by the laparoscopic access. For minor operative procedures like varix ligation or exploration of cryptorchidism, laparoscopy offers technical advantages rather than minimal invasiveness. PMID- 10720848 TI - Long-term experience with laparoscopic retroperitoneal lymph node dissection in the management of low-stage testis cancer. AB - OBJECTIVES: We describe our experience with laparoscopic retroperitoneal lymphadenectomy (LRLA) in 34 patients with low-stage germ cell tumors treated from 1992 to 1998. All patients had clinical stage-I disease with no clinical evidence (CT scan, ultrasound, tumor markers) of metastases. A laparoscopic dissection was used to assess the pathologic status of the relevant retroperitoneal lymph nodes. MATERIAL AND METHODS: 17 patients were treated by a transperitoneal laparoscopic approach, whereas in the last 17 patients retroperitoneoscopic retroperitoneal lymph node dissection was performed. The lymph node dissection was performed identically to open surgery with the modified template according to Weissbach including the paracaval, interaortocaval, upper pre-aortic, and right common iliac zonal nodes for right-sided tumors, and para aortic, upper pre-aortic zones for left-sided tumors. Retrieval of the lymph node chains was accomplished using a small organ bag. RESULTS: The procedure could be completed successfully in 30 of 34 patients with stage-I disease. In these cases the mean duration of the procedure was 248 min. In 3 patients the lymphadenectomy was abandoned, because frozen section showed metastasis. In 1 case conversion to open surgery was necessary because of bleeding from the aorta. One patient developed a delayed ureteral stenosis which required operative repair. Three patients required temporary insertion of an indwelling ureteral stent, and another patient had a pulmonary embolism with an uneventful outcome. One patient with a LRLA on the right side later developed retrograde ejaculation. In 6 of the 33 patients (18%) embryonal carcinoma or mixed carcinoma was found. The postoperative hospital stay averaged 5.3 (3-9) days for the patients without complications or conversion to open surgery. After a median follow-up of 40 months no regional relapse occurred, but 2 patients developed pulmonary metastases which were treated successfully by three cycles of platinum-based chemotherapy. All patients have no evidence of disease. CONCLUSIONS: Our experience suggests that LRLA is a safe and accurate method for low-stage germ cell tumors with minimal invasiveness, but because of its technical difficulty it should be restricted to experienced centers. PMID- 10720849 TI - Combined subcutaneous recombinant alpha-interferon and interleukin-2 in metastatic renal cell cancer: results of the Multicentre All Ireland Immunotherapy Study Group. AB - OBJECTIVE: To analyse the toxicity and efficacy of combined interferon-alpha and interleukin-2, administered subcutaneously in a general multicentre setting, as treatment for metastatic renal cell carcinoma. METHODS: Thirty-three patients with metastatic renal cell carcinoma were scheduled to receive 2 cyclical doses of subcutaneous interferon-alpha (week 1: 5 MU x 3 days) and interleukin-2 (week 2: 36 MU x 2 days, 9 MU x 3 days; weeks 3-5: 9 MU daily). Karnofsky scores ranged from 80 to 100 (median 90). Metastases occurred in multiple organs (lung 63%, retroperitoneal 39%, liver 24%). Patients were categorised according to the risk of disease progression. Treatment toxicity, therapeutic response and actuarial survival were analysed. RESULTS: All patients received recommended doses of treatment, but 6 received less than 2 cycles. Most were treated as outpatients, although hospitalisation was usual during the 1st week of a cycle. All complained of mild flu-like symptoms. Severe side effects developed in 13 patients (39%), and treatment was discontinued in 3 of these patients. No deaths occurred as a result of treatment. The overall median survival was 10 months. The overall actuarial survival rate at 3 years was 22%. On statistical analysis, actuarial survival rates were not influenced by either response to treatment or risk group category. CONCLUSION: Subcutaneously administered, combined interferon-alpha and interleukin-2 therapy achieves durable survival rates in a minority of patients with renal cell carcinoma. Toxicity is remedial, and not fatal, when subcutaneous therapy is administered by multiple medical disciplines at a variety of centres. PMID- 10720850 TI - Increase in incidental renal cell carcinoma in the northern part of the Netherlands. AB - OBJECTIVES: Evaluating in a retrospective survey the incidence of incidental and symptomatic renal cell carcinoma (RCC) between 1977 and 1994 in the northern part of the Netherlands and the mode of their detection. PATIENTS AND METHODS: Retrospectively, 173 patients surgically treated for RCC were divided into two groups according to the period of detection, 1977-1987 (n = 87) and 1987-1994 (n = 86). Because of the increase in abdominal ultrasound in 1987, this year was used as the cutoff date. In both periods the patients were grouped according to whether the tumor was found incidentally or whether the tumor was suspected. The mode of detection was recorded together with the tumor stage at presentation and survival. RESULTS: The incidental detection rate was 33% (29/87) in the 1977-1987 group and 49% (42/86) in the 1987-1994 group, showing a significant difference (p = 0.038). In the 1977-1987 group incidental tumors were detected with ultrasound in 83% and symptomatic tumors with ultrasound in 36%. Of the cases in the 1987 1994 group this percentage was 91 and 43%, respectively. Disease-free survival rates after a mean follow-up of 10 years were 63% in the incidental RCC group and 37% in the symptomatic RCC group (p = 0.0159). CONCLUSIONS: There is an increase in incidental tumors in this part of the Netherlands with ultrasound as the mode of detection. The disease-free survival is significantly better in the incidental tumor group. PMID- 10720851 TI - Proposal for changes in cystoscopic follow-up of patients with low-grade pTa bladder tumor. AB - OBJECTIVES: The cystoscopic follow-up of superficial bladder cancer accounts for a considerable workload for urologists and is also an invasive procedure with high costs. There is a potential benefit both to the urologist and the patient if unnecessary cystoscopies can be avoided. METHODS: The recurrence and progression rates of 120 patients with pTa G1 or G2 and small (<4 cm) transitional cell carcinoma were evaluated retrospectively. RESULTS: The recurrence rate was 6.5% (8/120) at 3 months. The recurrence rates at 6 and 9 months were 6.7 (8/119) and 3.6% (4/112), respectively. However, when the third month (first check) was clear, the recurrence rates at 6- and 9-month cystoscopy were 4.3 (5/116) and 2.7% (3/111), respectively. The recurrence rate at 12 months was 8% (8/99). For G1 tumors, the recurrence rates at 3, 6, 9 and 12 months were 6 (5/84), 5 (5/83), 2.5 (2/80) and 7% (5/71), respectively. The same results for G2 tumors were 8 (3/36), 8 (3/36), 6 (2/32) and 10.5% (3/28), respectively. The progression rate for the first year was lower than 1%. The difference between G1 and G2 tumors according to recurrence rate within the first year was not statistically significant (p>0. 05). CONCLUSIONS: This study supports the proposal that for patients with small and welldifferentiated pTa tumors at diagnosis, if the first control cystoscopy is clear, it is appropriate to perform the second check cystoscopy 1 year from initial resection and subsequent controls yearly. One should note that the study group included the most suitable patients for cystoscopic follow-up according to size and multiplicity of the tumor. This change in policy is further supported by the fact that progression occured in less than 1% in this group of patients. PMID- 10720852 TI - Bladder transitional cell carcinomas: a comparative study of washing and tumor bioptic samples by DNA flow cytometry and FISH analyses. AB - OBJECTIVE AND METHODS: We compared information obtained from samples of tumor biopsy and bladder washing to evaluate the representatives of the latter type of sampling. Both types of samples from 44 cases of superficial bladder TCCs and one papilloma were analyzed by FCM, to evaluate cellular DNA content, and FISH, to detect numerical aberration of chromosome 9. RESULTS: The use of both tumor and washing samples by FCM and FISH analyses evidenced alterations in 95% of cases. FCM evidenced higher aneuploid frequency in bladder washing than in bioptic specimens. In bladder washing, FISH analysis showed higher frequency of monosomy and lower frequency of trisomy than in biopsy. No correlation was found between histological grade and centromeric chromosome 9 loss while correlation was evidenced with centromeric 9 gain. CONCLUSION: Irrigation specimens, analyzed by FCM and FISH, can complement information obtained by biopsy in cytodiagnosis and follow-up of patients with bladder TCC. PMID- 10720853 TI - Role of preoperative positive apical biopsies in the prediction of specimen confined prostate cancer after radical retropubic prostatectomy: a multi institutional study. AB - OBJECTIVES: A multi-institutional study of 280 radical prostatectomy specimens obtained from three independent academic hospitals was undertaken to validate a nomogram developed for the prediction of specimenconfined protstate cancer after prostatectomy. METHODS: Three preoperative factors - the Gleason score, prostatespecific antigen (PSA) and apical location of positive biopsies - that were identified with a previous logistic regression formula were collected. Links between margin status and preoperative criteria were confirmed by univariate methods. Subsequently, the predictive indexes of positive margins were calculated and compared to the actual margin status in terms of predictive characteristics. RESULTS: This control series, independent of the initial series that was used to identify the relevant preoperative factors, confirmed that positive apical biopsies(p<0.001), PSA (p<0.005) and the Gleason score (p<0.005) were strongly linked to the occurrence of positive margins. Different cutoff values for the predictive index were compared in a receiver operating characteristic curve. A value of 0.5, similar to the one described in the original series, gave an adequate compromise between sensitivity and specificity with respective values of 68 and 73% and a test accuracy of 72%. In practical terms, it was possible to predict 85% of negative margins, and to delineate two groups with different rates of positive margins (14.5 vs. 50%). CONCLUSIONS: We demonstrated that PSA, the Gleason score and apical biopsy status are cumulative risk factors for positive margins. Risk of positive margins increases when it is not possible to obtain a wide excision of periprostatic fascia, as at the apex. This study substantiates the independent prognostic value of positive preoperative apical biopsies for predicting positive surgical margins. PMID- 10720854 TI - Determination of the percentage of free prostate-specific antigen helps to avoid unnecessary biopsies in men with normal rectal examinations and total prostate specific antigen of 4-10 ng/ml. AB - OBJECTIVE: To assess the usefulness of measuring the percentage of free prostate specific antigen (PSA) in serum to reduce the number of prostate biopsies in men with serum PSA levels between 4 and 10 ng/ml and benign prostate examinations. MATERIALS AND METHODS: The percentage of free PSA (Immulite((R))) in serum was analyzed prospectively in 180 men with benign digital rectal examinations and total PSA serum levels of between 4 and 10 ng/ml. All patients underwent ultrasound-guided sextant prostatic biopsies. Sensitivity, specificity and positive and negative predictive values were calculated as well as the percent of patients in which biopsies could have been avoided for various cutoff values of the percentage of free PSA as an indicator for biopsy. Influence of age in the determination of cut points was evaluated. RESULTS: Cancer was detected in 22.2% (40/180) of the patients. Mean percentage of free PSA was 13.4% in patients with cancer and 18.9% in patients with benign prostatic hyperplasia (p = 0.001). Using a percentage of free PSA cutoff of 22% or less as a criterion for performing prostatic biopsy would have detected 95% of cancers, avoided 25% of benign biopsies and yielded a positive predictive value of 29% in patients who underwent biopsy. Mean percent of free PSA values increased as mean subject age increased, influencing the calculation of cut points, sensitivity and specificity. Leaving the cut point constant across all age groups will oblige older patients to undergo an increased number of unnecessary biopsies, although allowing for higher sensitivity in younger men. CONCLUSIONS: Measurement of the percentage of free serum PSA improves specificity of prostate cancer detection in patients with elevated total serum PSA levels and benign prostate examinations. Subject age seemed to influence the determination of optimal cut points. PMID- 10720855 TI - Nondissection of pelvic lymph nodes does not influence the results of perineal radical prostatectomy in selected patients. AB - OBJECTIVES: Retrospective studies have shown that pelvic lymph node dissection can be dispensed with in selected men undergoing radical prostatectomy. We prospectively evaluated the influence of nondissection of pelvic lymph nodes on tumor progression in our first 100 perineal prostatectomies. METHODS: From October 1992 to February 1998, 100 patients underwent radical perineal prostatectomy for localized prostate cancer. Preoperative PSA, the Gleason score of positive biopsies and age at surgery were noted. Forty-three of the 100 patients (group 1) did not undergo pelvic lymph node dissection because their preoperative PSA level was below 10 ng/ml (Hybritech assay, normal value 4 ng/ml) and the Gleason score of their positive biopsies was below 7. These 43 patients were compared with 25 of the 114 patients operated on during the same period by the retropubic approach and who had pelvic node dissection and the same preoperative criteria (PSA <10 ng/ml and a Gleason score of positive biopsies <7; group 2). All prostatectomy specimens were processed according to the Stanford protocol: prostate weight, Gleason score, capsular, seminal vesicle, lymph node and surgical margin status, and tumor volume were studied. Postoperative followup was based on routine serum PSA assays after 1 and 3 months and then half-yearly. Biological progression was defined as PSA level which was detectable postoperatively (>/=0.2 ng/ml). Kaplan-Meier analysis was used to evaluate the likelihood of biochemical recurrence. Results were compared by using Fisher's test, the Mann-Whitney test and the log-rank test. Differences were considered significant when the p value was <0.05. RESULTS: No differences in preoperative characteristics were observed; in groups 1 and 2, mean age was 65.9 and 64.7 years, PSA was 6.7 and 5.11 ng/ml, and the Gleason biopsy score was 5.7 vs. 5.0, respectively. In groups 1 and 2, specimen weight was 44.5 and 54.3 g (p = 0.04), the Gleason score was 6.2 and 5.6, tumor volume was 0.91 and 0.8 ml, 81.4 and 84% of patients were in stage pT2, 13.9 and 12% had extracapsular disease, 4.6 and 0% had seminal vesicle invasion, and 13.9 and 16% had positive surgical margins, respectively. The mean follow-up was 2.74 years (0.27-5.59 years). The actuarial 5-year recurrence-free rate was 78% in group 1 and 80% in group 2 (p>0.05). CONCLUSION: The lack of pelvic lymph node dissection does not influence the intermediate term results of perineal radical prostatectomy in selected patients (preoperative PSA <10 ng/ml and Gleason score for positive biopsies <7). PMID- 10720856 TI - Transurethral vaporization-resection of the prostate versus standard transurethral prostatectomy: comparative changes in histopathological features of the resected specimens. AB - OBJECTIVES: Transurethral vaporization resection of the prostate (TUVRP) is a recent modification of the standard transurethral resection of the prostate (TURP). TUVRP uses a band electrode coupled to a high electrocuting energy to achieve simultaneous resection, vaporization and coagulation of the prostate. We evaluated the histopathological resection specimens of patients treated with TUVRP to see whether the higher energy used will result in thermal artifacts that will interfere with the pathological evaluation of the prostate, and compared the results to TURP specimens. MATERIAL AND METHODS: The histopathological specimens of 50 patients that underwent TUVRP or TURP were reviewed. Artifactual pathological patterns that were identified in the specimens included: abnormal cellular orientation and spindling, artifactual cellular detachment from the underlying basement membrane, atypical cytological changes or areas of stromal coagulative necrosis. Each identified pattern was awarded 1 point. The severity of cautery artifact was graded into mild, moderate or severe according to the sum of points in each specimen. RESULTS: Mild cautery artifact changes were noted in 1 patient who underwent TURP. Moderate changes were noted in 21 patients in each TURP and TUVRP groups while severe changes were noted in 4 and 3 patients undergoing TUVRP and TURP respectively. There were no statistically significant differences between the groups with regard to the severity of the cauterization- induced changes. CONCLUSIONS: The quality of histopathological specimens produced by TUVRP is similar to the standard TURP. It seems that the higher energy use in electrovaporization technique does not result in greater thermal injury to the tissues possibly because of the cooling effect of the irrigation fluids used intraoperatively. PMID- 10720857 TI - Efficacy and safety of a new prolonged release formulation of alfuzosin 10 mg once daily versus alfuzosin 2.5 mg thrice daily and placebo in patients with symptomatic benign prostatic hyperplasia. ALFORTI Study Group. AB - OBJECTIVES: To assess the efficacy and safety of a new prolonged release formulation of the uroselective alpha(1)-blocker alfuzosin for a once-daily dosing regimen in patients with lower urinary tract symptoms (LUTS) suggestive of symptomatic benign prostatic hyperplasia (BPH). METHODS: After a 1-month run-in period, 447 patients were randomly allocated in a double-blind placebo-controlled study to receive alfuzosin 10 mg once daily (n = 143), alfuzosin 2.5 mg thrice daily (n = 150) or placebo (n = 154) for 3 months. At inclusion, 46% of the randomised population had concomitant cardiovascular disease and 30% received an antihypertensive treatment. Uroflowmetry was performed close to trough plasma concentration of alfuzosin once daily to demonstrate the 24-hour coverage with this formulation. RESULTS: Both alfuzosin formulations significantly improved urinary symptoms versus placebo assessed using the International Prostate Symptom Score (alfuzosin 10 mg once daily: -6.9; alfuzosin 2.5 mg thrice daily: -6.4; placebo: -4.9, p = 0.005). Peak flow rate increased significantly with alfuzosin 10 mg once daily (+2.3 ml/s, p = 0.03 vs. placebo) and with alfuzosin 2.5 mg thrice daily (+3.2ml/s, p<0.0001 vs. placebo) compared to placebo (+1.4 ml/s). Overall both formulations of alfuzosin were well tolerated in comparison with placebo. In addition, vasodilatory adverse events appeared to be less frequent with the once daily than the thrice daily formulation (6.3 vs. 9.4%, respectively). No first-day effect was reported with alfuzosin once daily and the effect on blood pressure did not differ from those observed in placebo, both in normotensive and hypertensive patients. No specific sexual dysfunction including ejaculation disorder was reported in the alfuzosin 10 mg once-daily group. CONCLUSION: The new once-daily formulation of alfuzosin administered at a dose of 10 mg daily is an effective 24-hour treatment of LUTS associated with BPH. Alfuzosin is as effective as the immediate formulation and shows a better cardiovascular safety. The better safety profile enables the same dose to be used in all patients, providing the patients with the benefits of a once-daily administration. PMID- 10720858 TI - Detrusor contraction duration may predict response to alpha-blocker therapy for lower urinary tract symptoms. AB - OBJECTIVES: Baseline pressure/flow parameters have not correlated well with response to medical therapy for lower urinary tract symptoms (LUTS). This open label, nonrandomized retrospective study was designed to evaluate whether the urodynamic parameter duration (in seconds) of the detrusor contraction (DCD) correlates better with alpha-blocker response than previously described urodynamic parameters. METHODS: 93 men (mean age 62.6+/-8.5) with LUTS underwent urodynamic evaluation prior to initiating therapy with doxazosin titrated to 8 mg and followed for 6 months. Parameters of evaluation included the AUA symptom score (AUASx), peak urinary flow rate (Q(max)), maximal detrusor pressure (P(max)), detrusor pressure at maximal flow (P(det)) and DCD. The correlation and predictive value of therapeutic response and baseline urodynamic parameters were assessed. RESULTS: 85 patients were evaluable at 6 months. For the entire group, AUASx decreased from 15.1+/-6.9 to 9.7+/-5.1 (-36%) and Q(max) increased from 9.3+/-3.7 to 11.9+/-5.7 ml/s (+28%). Baseline P(max) was 74.8+/-19.6 cm H(2)O, P(det) was 61.6+/-18.9 cm H(2)O and DCD was 107.6+/-28.6 s. There was weak correlation between either baseline P(max) or P(det) and therapeutic response (defined as a decrease in AUASx of 40% and an increase in Q(max) of 30%). Utilizing a baseline DCD of 90 s or more, there was a significant correlation to therapeutic response (r = 0.48, p = 0.002). CONCLUSIONS: These preliminary data suggest that DCD may be a useful urodynamic parameter to predict and optimize therapy with a-blockade. The potential utility and cost-effectiveness of DCD remains to be determined. PMID- 10720859 TI - Prognostic factors for long-term maintenance of urinary continence in patients with incontinence managed by diapers or indwelling catheters. AB - OBJECTIVE: We examined the prognostic factors for longterm maintenance of continence following nonsurgical treatments in hospitalized patients with urinary incontinence. METHODS: 313 inpatients (average age: 64 years) in whom urinary incontinence had originally been managed with diapers (n = 158) or indwelling Foley catheters (n = 155) first received nonsurgical rehabilitative treatments. The patients who became continent with these treatments were then evaluated for being either continuously continent or recurrently incontinent during the 5-year follow-up period after discharge from hospital. A multivariate logistic regression analysis was then performed to determine significant risk factors for recurrent urinary incontinence. RESULTS: By initial treatments (mean duration: 144 days), 294 of the 313 patients (94%) were continent and free from diapers or catheters. After the 5-year follow-up period, urinary continence was maintained in 103 (66%) and 62 patients (45%) initially managed with diapers (n = 157) and catheters (n = 137), respectively. Multivariate logistic analysis revealed that in both patient groups, poor posttherapeutic activities of daily living or loss of home care service lowered the maintenance rate of continence. Dementia also lowered the maintenance rate in patients with catheters, but not in those with diapers. In addition, a long history of Foley catheter drainage for over 1 year prior to the initial treatment reduced the maintenance rate (highest odds ratio). CONCLUSIONS: Physical disability and poor therapeutic assistance at home are more prominent risk factors for long-term maintenance of urinary continence in elderly patients than problems within the urinary tract. PMID- 10720860 TI - Tansvaginal needle suspension operation: the way we do it. Clinical and urodynamic study: long-term results. AB - OBJECTIVES: To evaluate the long-term results of the transvaginal needle suspension operation for urinary stress incontinence. MATERIALS AND METHODS: A total of 88 women with proved genuine stress incontinence were treated with transvaginal needle suspension with fixation of suspension sutures to the rectus fascia using the technique of crossing suspension sutures. By using this method the proximal end of suspension suture from one side is tied with the distal end of suspension suture from the other side. The suspension sutures fixed in this way ensure 3-4 cm of rectus fascia which is used as a carrier of the suspension sutures. The same urologist peformed 88 consecutive operations. Clinical and urodynamic evaluations were performed at 6 months, 1 year and 5 years after surgery with the same technique and the same equipment. RESULTS: Analysis of the questionnaire showed that 81 patients (92.0%) were continent after 6 months while 78 (88.6%) patients were still continent after 1 year. After 5 years (n = 71) there were only continent 54 (76.0%) and incontinent patients (n = 17, 23.9%). Urodynamic analysis showed that 49 (69.0%) patients were continent after 5 years (n= 71). The increase in the number of incontinent patients is achieved at the cost of the previously continent patients. Of the 22 incontinent patients (after 5 years), 16 were still stress incontinent, while 6 (8.3%) patients had urge incontinence due to de novo detrusor instability. Three patients (n = 88, 3.4%) had undergone unilateral suture removal due to infection without influence on their continence status. In 2 patients (n = 88, 2.2%) the clinical pictures were highly suggestive of ilioinguinal nerve entrapment. CONCLUSIONS: Our results suggest that the transvaginal needle suspension operation is satisfactory for the management of genuine stress incontinence in women. However, we believe that the success of any suspension operation lies in adequate mobilization of the bladder neck and urethra (anterior vaginal wall) as well as in a surgeon's familiarity with the procedure. PMID- 10720861 TI - Local anesthesia for extracorporeal shock wave lithotripsy: a double-blind, prospective, randomized study. AB - OBJECTIVE: The efficacy of local anesthesia in decreasing intravenous analgesic requirements during extracorporeal shock wave lithotripsy with a second generation lithotriptor was studied. METHODS: Subcutaneous infiltration was performed before the procedure. Sixty-nine patients (ASA I-II) were randomly allocated into four groups. Lidocaine 1% plus epinephrine (5 microg/ml) were infiltrated subcutaneously in a group of patients with ureteral stones (group UL), and a group with renal stones (group RL). The same amount of saline was administered to a group of patients with ureteral stones (group UC), and a group with renal stones (group RC). RESULTS: Patients with ureteral stones needed higher doses of intravenous analgesic. Neither patients with renal stones nor patients with ureteral stones administered local anesthetic required less intravenous analgesic than patients given placebo. CONCLUSION: Local anesthesia did not decrease the requirement of intravenous doses of analgesics in patients treated with a second-generation lithotriptor (Dornier MPL 9000). PMID- 10720862 TI - Diclofenac treatment prolongs renal transit time in acute ureteral obstruction: a renographic study. AB - OBJECTIVE: Prostaglandin inhibitors, mostly diclofenac, are currently first choice therapy for ureteral colic, their main action being reduction of intrapelvic pressure and diuresis. We hypothesized that diclofenac, by increasing tubular reabsorption, can delay excretion of contrast medium and give a false impression of severe obstruction. METHODS: Gamma camera renography was performed with 50 MBq (99)Tc(m)-MAG3 before and with 150 MBq 30 min after intramuscular injection of 75 mg diclofenac in 10 patients with acute ureteral colic. The time to maximum isotope activity in each kidney, T(max), was compared with T(max) in 10 control patients, who did not receive diclofenac but underwent two identical renographies. RESULTS: T(max) was significantly delayed after diclofenac, from 353 s at baseline to >1,200 s on the stone side, and from 225 to 465 s on the healthy side. Without diclofenac there was no T(max) retardation between the two renographies. CONCLUSION: Diclofenac treatment can lead to overestimation in ureteral stone disease, by delaying renal excretion bilaterally, but predominantly on the side of calculus. PMID- 10720864 TI - Deep dorsal vein arterialization in pure cavernoocclusive dysfunction. AB - PURPOSE: We report our 4-year experience with deep dorsal vein arterialization at 3 years' follow-up in young patients with pure cavernoocclusive dysfunction as an alternative to penile prosthesis implantation. MATERIALS AND METHODS: We performed a modified Furlow-Fisher operation (circumflex collaterals are preserved and the deep dorsal venous valves are not disrupted by a stipper) in 25 patients who did not have risk factors such as general arteriosclerosis, coronary heart disease, hypertension, hyperlipidemia and age (over 40 years). Patients with arterial disease diagnosed by Doppler examination were excluded from the study. Also, patients with abnormal penile biothesiometric and electromyographic findings were not included in the study. Beside the subjective and objective evaluation the efficacy of the operation was also assessed retrospectively in 18 patients by telephone according to items 3 (ability to achieve an erection) and 4 (ability to maintain an erection) of the 15-item International Index of Erectile Function (IIEF). RESULTS: With a mean follow-up of 28 months (range 4-42) subjective and objective success rates were 80 and 72% at 1 year's 75 and 62.5% at 2 years', and 70 and 60% at 3 years' follow-up. According to items 3 and 4 of the 15 item IIEF questionnaire the mean postoperative scores reached 1.55-3.44 and 1. 33-3.27 for items 3 and 4, respectively (p<0.01). Two patients (8%) showed signs of glans hypervascularization as a major complication and minor complications such as penile skin edema, subdermal hematoma, loss of penile skin sensation and early thrombosis of the anastomosis were found in a total of 8 patients (32%). CONCLUSIONS: Deep dorsal vein arterialization is a preferable choice in highly selected young patients as an alternative to penile prosthesis. PMID- 10720863 TI - Course of calcium stone disease without treatment. What can we expect? AB - OBJECTIVES: The knowledge of the natural history (i.e. the course of the disease without metaphylaxis is the base for establishing rational guidelines for metaphylaxis in urolithiasis. METHODS: This review is based on a Medlinetrade mark Search (1966-1999) and the proceedings of the Bonn-Vienna and European symposia on urolithiasis. Only 31 references were sufficient for the purpose of this review. RESULTS: In idiopathic calcium stone disease, stone frequency without metaphylaxis is 0.10-0.15 stones per patient per year. The average recurrence rate is 30-40%. Recurrence rate increases with age and observation time. Risk for recurrence is highest during the first 4 years after the first stone episode. More than 50% of all recurrent stone formers have only one recurrence during their lives. 10% of recurrent stone formers have more than 3 recurrences. Risk factors for recurrence are: male sex, multiple and lower calyx stones, early onset, familial history, complications after stone removal. Metabolic evaluation is a poor predictor of the risk for recurrence. CONCLUSIONS: Renunciation of metaphylaxis is justified in first stone formers with idiopathic calcium oxalate and apatite stones. All patients, however, should be advised to increase their fluid intake. PMID- 10720865 TI - Genetic alterations in urinary bladder carcinosarcoma: evidence of a common clonal origin. AB - The cellular origin of carcinosarcoma of the bladder is unknown. We addressed this issue by using microsatellite analysis for loss of heterozygosity (LOH) in both the carcinomatous and sarcomatous components of 6 bladder tumors. We tested 40 microsatellite markers from 19 human chromosomes and compared the genetic alterations between the two separately isolated components. The potential relevance of the E-cadherin pathway was also evaluated by immunohistochemistry. All 6 cases revealed identical LOH on chromosomal arms 9p, 9q, 8p, and 8q, corresponding to relatively early events in bladder carcinogenesis. Discordant losses between two alleles in the remaining chromosomes, associated with progression, were seen in all tumors with a trend toward a higher incidence in the more advanced tumors (N1M1 and N1Mx). E-cadherin was strongly expressed in the carcinomatous components (5 of 6), whereas most of sarcomatous elements displayed absence of the protein product (4 of 6). These results indicate that both the carcinomatous and sarcomatous components of carcinosarcoma are derived from a common stem cell. Downregulation of E-cadherin may define one of the pathways responsible for conversion of epithelial cells to the sarcomatous phenotype. PMID- 10720866 TI - Chromosomal numerical aberrations detected by fluorescence in situ hybridization on bladder washings from patients with bladder cancer. AB - OBJECTIVE: Previous studies on touch biopsy specimens have determined numerical or structural changes involving many different chromosomes in bladder cancer. The aim of this study was to evaluate the use of fluorescence in situ hybridization (FISH) assay in bladder washings as an objective technique to detect chromosomal numerical aberrations in bladder cancer. The main advantages of bladder washings are that they can be easily collected during the clinical follow-up of patients with superficial bladder cancer and they do not contain so many degenerate cells as urine samples. METHODS: We collected specimens from 25 patients who underwent transurethral resection of bladder tumors. Double target FISH assays with centromeric labeled probes for chromosomes 7, 8, 9 and 11 were used on the bladder washings and on the touch biopsy slides. The results were compared to flow cytometry and tumor grade and stage. RESULTS: We found monosomy 9 and trisomy 7, 8, 9 and 11 in 28, 32, 36, 28 and 25% respectively of the patients. FISH analysis of bladder washing versus touch biopsy specimens were concordant in approximately 90% of the slides. Total DNA aneuploidy correlated well with numerical aberrations of chromosomes 7, 8 and 11, but not with chromosome 9. CONCLUSION: Although better hybridization efficiency was obtained on touch biopsy slides, the results in bladder washings were in high concordance. FISH analysis on bladder washing samples may become a simple tool to improve the accuracy of cytology. PMID- 10720867 TI - Chronic renal failure. Initial assessment, diagnosis and therapy PMID- 10720869 TI - Mixed lymphocyte culture of human fetal liver cells. AB - OBJECTIVE: In order to study the immunological function of the human fetus in the first and second trimesters, mixed lymphocyte culture (MLC) of fetal liver and thymic cells was performed. MLC is a functional test to determine human lymphocyte antigen-D incompatibilities. METHODS: Human fetal liver and thymic tissue was obtained from abortions in gestational weeks 7-17.5. Forty-seven fetuses were studied with one-way MLC. The cells were stimulated by adding irradiated fetal liver cells, adult bone marrow and peripheral blood lymphocytes. The activity was measured as DNA incorporation of radiolabeled thymidine. RESULTS: The results indicate that the human fetus is competent to react as early as 11-12 weeks of gestation and in some cases even earlier. In very immature fetal livers (< 8 weeks), the MLC seems to be inhibited. CONCLUSIONS: Our data suggest that the human fetus can react against foreign transplantation antigens earlier than previous papers have claimed. The onset of reactivity seems to differ considerably among fetuses. The present findings may explain some of the limited success of in utero transplantations of hematopoietic stem cells in human fetuses of normal immunological status. PMID- 10720868 TI - Transvaginal assessment of fetal anatomy at 11 to 16 weeks of gestation in relation to fetal position. AB - OBJECTIVE: To investigate the ability of the high-frequency transvaginal scanning technique to assess embryo-fetal anatomy in early pregnancy in relation to fetal position and in comparison with transabdominal scan. METHODS: A study population of 1,402 pregnant women were examined by transvaginal ultrasonography performed at 10(+1) to 16(+0) weeks of pregnancy. At 14(+1) to 16(+0) weeks of gestation, an ultrasonographic examination was performed in 247 pregnant women by transvaginal and transabdominal route. The criteria necessary for an adequate visualization of fetal organs and structures were met. RESULTS: The visualization rate of complete fetal anatomy increased with increase in menstrual age and fetal position affected this possibility. A detailed brain anatomy was more easily viewed with the fetus in the cephalic or transverse position than in the breech position, whereas the thoracic and abdominal anatomy were viewed more easily with the transverse position compared to the cephalic or breech position. Complete surveys of fetal anatomy were possible in 50% and 62% of women with transabdominal scan and in 85% and 85% with transvaginal one at 15 and 16 weeks' gestation, respectively (p < 0.001). CONCLUSION: A detailed assessment of fetal structures was possible in most cases at 13 weeks of gestation. Fetal position can influence this capability. At 14(+1) to 15(+0) weeks' gestation more anatomical surveys were completed with a transvaginal scan compared to a transabdominal one and this was influenced by fetal position. PMID- 10720870 TI - Amniotic fluid pressure in both cavities of twin-to-twin transfusion syndrome: a vote against septostomy. AB - OBJECTIVE: To report our results of the measurement of the amniotic fluid pressure in twin-to-twin transfusion syndrome before and after amniodrainage and the separate measurement in the cavity of the stuck twin and the recipient, respectively, in 2 cases. METHOD: Six patients with severe TTS underwent therapeutic amniodrainage. Amniotic fluid pressure was obtained before and after reduction of the fluid volume. In 2 of these cases we were able to calculate the pressure separately in both amniotic cavities, the donors (stuck twin) and the recipients. RESULTS: In all cases of TTS, amniotic pressure was high before and reduced after amniodrainage. The pressure was equal in the donors and in the recipients cavity. CONCLUSION: Amniotic pressure seems to be high in TTS, but equal in both cavities. Therefore, amniotic septostomy is probably of low value, especially if a high-risk monoamniotic situation is artificially created. PMID- 10720871 TI - In utero repair of myelomeningocele: a comparison of endoscopy and hysterotomy. AB - OBJECTIVE: To compare endoscopic coverage of myelomeningocele with a maternal split-thickness skin graft in utero to definitive neurosurgical closure through a hysterotomy. METHODS: Four fetuses with isolated myelomeningocele underwent endoscopic coverage of the defect with a maternal split-thickness skin graft in a CO(2) environment at 22-24 weeks' gestation. Subsequently, 4 fetuses underwent standard neurosurgical closure of their myelomeningoceles at 28-29 weeks' gestation. RESULTS: The mean operating time for the endoscopic procedures was 297 +/- 69 min. Two fetal losses occurred as a result of chorioamnionitis and placental abruption, respectively. A third baby delivered at 28 weeks' gestation after prolonged disruption of the membranes. The 2 survivors required standard closure of the myelomeningocele after delivery. The mean operating time for the hysterotomy procedures was 125 +/- 8 min. No mortality occurred, and all the infants delivered between 33 and 36 weeks with well-healed myelomeningocele scars. At present, the functional levels of all infants approximate the anatomical levels of the lesions. CONCLUSION: With current technology, in utero repair of congenital myelomeningocele through a hysterotomy appears to be technically superior to procedures performed endoscopically. PMID- 10720872 TI - Molecular and fetal tissue biopsy capabilities are needed to maximize prenatal diagnosis of junctional epidermolysis bullosa: fetal skin biopsy using a 1-mm microendoscope. AB - OBJECTIVE: To describe a minimally invasive micro-endoscopic technique for fetal skin biopsy and direct examination for a lethal skin condition. MATERIALS AND METHODS: Direct fetoscopic examination of a fetus was undertaken along with full thickness skin biopsies at 19 weeks' gestation. RESULTS: No phenotypic expressions of the lethal condition were visualized and six full thickness skin biopsies were collected. Pathological examination revealed normal skin structures not consistent with junctional epidermolysis bullosa (JEB). CONCLUSION: Minimally invasive examination with the 1 mm endoscope allows direct fetal phenotypic evaluation, full thickness skin biopsies, with risks similar to amniocentesis. PMID- 10720873 TI - Efficacy and safety of OK-432 sclerotherapy for giant cystic hygroma in a newborn. AB - BACKGROUND: OK-432, a lyophilised incubation mixture of group A Streptococcus pyogenes of human origin, was used as a sclerosant for the involution of a giant cervical cystic hygroma in a newborn. RESULTS: There were no systemic side effects. Blood tests and double immune diffusion tests showed no systemic infection or generalised inflammatory response, or antibody production. Cellular and cytokine-induced localised inflammatory reaction within the cystic hygroma, was observed on analysis of the intracystic fluid. CONCLUSIONS: The leucocytosis induced and activated by OK-432 probably increases the endothelial permeability of the lymphatics. This probably accelerated lymph drainage leading to involution of the cystic hygroma. Intralesional injection of OK-432 was safe and effective therapy for cystic hygroma in this newborn as its inflammatory reaction was localised. PMID- 10720874 TI - Sonographic, cytogenetic and DNA analysis in four 69,XXX fetuses diagnosed in the second trimester. AB - OBJECTIVE: To describe the ultrasound findings and its relationship with the cytogenetic study and the origin of the extra haploid chromosome set in four 69,XXX cases. METHODS: Four pregnant women were referred because of abnormal 2nd trimester ultrasound. Karytoypes, FISH and DNA analysis were performed. RESULTS: All cases presented asymmetrical intrauterine growth retardation, marked oligohydramnios and placental alterations and showed a 69,XXX karyotype. In three cases, DNA analysis allowed to establish the origin of the extra haploid chromosome set. CONCLUSIONS: At least three fetuses had a maternal extra haploid chromosome set. Thus, it has been possible to establish the main ultrasonographic markers and to observe the survival of the fetus until the second trimester when they have a maternal origin. PMID- 10720875 TI - High sensitivity of fetal DNA in plasma compared to serum and nucleated cells using unnested PCR in maternal blood. AB - DNA analysis of blood is conventionally performed on cells - plasma and serum are discarded. Free DNA has been demonstrated in serum in cancer and autoimmune disorders and in pregnancy. We investigated possible noninvasive prenatal diagnosis using fetal DNA from maternal plasma and serum in pregnancy. Fetal gender was determined by PCR on DNA from maternal venous blood, serum and plasma of 65 women by boiling with or without phenol/chloroform extraction. When sensitivities were compared for plasma, additional phenol/chloroform extraction proved more sensitive than boiling alone (89 vs. 50%), the observed difference was 50% (CI 19 to 81%). Extracted plasma amplified better than extracted serum (89 vs. 46%), the observed difference being 44% (CI 22 to 66%). In contrast, fetal gender could not reliably be determined from DNA extracted from maternal nucleated blood cells. The size of plasma and serum DNA at 15-17 weeks of gestation was >1,500 bp. This work confirms the presence of fetal DNA in maternal plasma and serum which may be applicable to noninvasive prenatal diagnosis of paternally derived DNA sequences. We conclude that optimal sensitivity requires two methods of DNA extraction and that the use of plasma is preferred to that of serum. PMID- 10720876 TI - Maternal serum screening for down syndrome by using alpha-fetoprotein and human chorionic gonadotropin in an asian population. a prospective study. AB - The purpose of the present study is to evaluate the efficacy of second-trimester maternal serum screening program by using alpha-fetoprotein (AFP) and total human chorionic gonadotropin (hCG) in an Asian population. During June 1994 to July 1998, we conducted a prospective study of serum screening protocol for Down syndrome. The cut-off point for a positive result in this analysis was a risk of >/=1/270. A total of 17,742 pregnant women with singleton pregnancy were screened, and 1,153 (6.5%) had positive result. Sixteen of the 17,742 pregnancies had Down syndrome, and 10 of them had positive result. The positive rate and detective rate for Down syndrome were 6.5 and 62.5%, respectively. However, the detective rate will reduce to 47.6% after being adjusted by age-specific risk. It is indicated that the double-marker test using AFP and total hCG is an effective screen strategy for second-trimester detection of Down syndrome in Asian women. PMID- 10720877 TI - Efficacy of maternal serum screening in the prenatal detection of fetal chromosome abnormalities in Japanese women. AB - OBJECTIVE: This prospective study assesses the efficacy of maternal serum screening for use in prenatal diagnosis of fetal anomaly and chromosome imbalance in Japanese women. METHODS: Maternal serum alpha-fetoprotein, human chorionic gonadotropin, and unconjugated estriol were measured in 1,055 singleton pregnant women between 14 and 20 weeks of gestation. A calculated risk for trisomy 21 of >/=1/299 or alpha-fetoprotein >/=2.5 multiples of the median was adopted as positive. RESULTS: Three hundred and seventy-eight of the 1, 055 women screened (35.8%) were identified as positive. Sensitivity, false-positive rate, and positive predictive value in women aged <35 years were 60.0, 10.6, and 6.8%, respectively, and these values were 87.5, 49.3, and 4.2%, respectively, in women aged >/=35 years. The false-positive rate in women aged <35 years was significantly lower than that for women aged >/=35 years (p < 0.001). Chromosomal abnormalities were identified in 21 cases, including 10 with trisomy 21, 5 with trisomy 18, 2 with trisomy 13, and 4 with other chromosomal disorders. Seventeen of the 21 cases (81.0%) showed screen-positive results, and among these all 10 cases with trisomy 21 were detectable. Two cases with trisomy 18, 1 with trisomy 13 and 1 with isochromosome X showed extremely low human chorionic gonadotropin levels (0.4 +/- 0.1 multiples of the median, mean +/- SE), although they were screen negative. Of the 264 women who did not undergo amniocentesis, none had any clinical findings consistent with aneuploidy after birth. CONCLUSIONS: Our results suggest that the evaluation of each serum marker, as well as of the calculated risk, was significantly important in the prenatal detection of fetal aneuploidy. PMID- 10720878 TI - Maternal contamination at fetal muscle biopsy. AB - Duchenne muscular dystrophy (DMD) can be diagnosed by fetal muscle biopsy and immunohistochemical staining showing the absence of dystrophin. We report a case of fetal muscle biopsy in which the needle gun was successfully fired within the fetal gluteal muscle but the sample was contaminated by maternal tissue. This was attributed to the design of the biopsy needle, allowing transient opening of the biopsy core as the needle penetrated the maternal rectus sheath, muscle, and myometrium. Histology showed tissue suggestive of maternal origin, confirmed by DNA analysis. Repeat sampling revealed fetal muscle with normal dystrophin expression. We recommend that care be taken during needle insertion to avoid maternal contamination, and tests be used to confirm the fetal source of the sample. PMID- 10720879 TI - Efficacy of oral iodide therapy on neonatal hyperthyroidism caused by maternal Graves' disease. AB - OBJECTIVE: To verify the efficacy of oral iodide therapy in treating a case of early neonatal hyperthyroidism due to maternal Graves' disease. METHODS: We report a case of neonatal hyperthyroidism which occurred in a 2,650-gram, female baby, born at 39 weeks' gestational age (GA) to a 30-year-old mother affected by Graves' disease and treated with thionamides (propylthiouracil) from the 20th week of gestation. A fetal goiter, due to maternal therapy, had been observed by ultrasound scan at 31 and 35 weeks of gestation, with contemporary low cord thyroid hormone levels. Two intra-amniotic injections of levothyroxine were then performed at 34 and 36 weeks of gestation, which led to a significant reduction of fetal goiter and to normalization of cord thyroid hormone levels. The neonatal clinical course was characterized by symptoms of hyperthyroidism from the 2nd to 3rd days of life (irritability, tachycardia, tachypnea, hyperphagia), mostly during feeding. Oral treatment with potassium iodide (KI, 8 mg x 3 times a day) was started at 23 days of life. RESULTS: Treatment with KI led to a significant reduction of neonatal clinical symptoms and to a normalization of hormone levels within 4 days of therapy. The treatment was discontinued in 13th week of life because of neonatal well-being and normal hormone levels. CONCLUSIONS: We believe that KI therapy is effective in treating neonatal hyperthyroidism and does not cause suppression of neonatal thyroid activity, which is possible using antithyroid drugs like thionamides. PMID- 10720880 TI - Chemokine receptors and their role in vascular biology. AB - Chemokines play an important role in the process of leukocyte recruitment and activation at sites of inflammation. Until recently, the actions of chemokines and the expression of their receptors have only been described on different leukocyte populations. However, increasing evidence has suggested that non haematopoietic cell types are capable of binding and responding to a number of chemokines. The functional expression of certain chemokine receptors has recently been described on vascular endothelial and smooth muscle cells. These findings provide new insight into the activities of chemokines and indicate that these molecules have a more widespread cellular target than first envisaged. Studies carried out to date indicate that chemokines and their respective receptors play an important role in the regulation of angiogenesis and angiostasis. They may also be involved in developmental and pathological processes such as organ vascularization, embryogenesis and arteriosclerosis. PMID- 10720881 TI - Blockade of angiotensin II type 1 receptors: effect on carotid and radial artery structure and function in hypertensive humans. AB - Converting-enzyme inhibition reduces cardiovascular hypertrophy in hypertensive subjects. Whether the blockade of angiotensin II type 1 (AT(1)) receptors reduces arterial hypertrophy has never been investigated. In a double-blind study versus placebo in subjects with essential hypertension, the effect of the AT(1) blocker irbesartan (150 mg/day for 8 weeks) on blood pressure, wall thickness, diameter and stiffness of the common carotid and radial arteries was studied, using echotracking techniques of high resolution. With irbesartan, mean blood pressure decreased significantly and proportionally to the baseline levels of active renin, and angiotensin I and II. There was a significant decrease in radial artery wall thickness. The percent change from baseline (+/- SEM) was -10.51 +/- 3.42 versus 6.18 +/- 4.77. There was no significant change in diameter or distensibility. This effect was correlated neither to blood pressure changes nor to hormonal baseline levels of the renin-angiotensin system. Carotid wall thickness and diameter were unchanged. Thus a 2-month treatment with an AT(1) antagonist significantly reduced radial but not carotid artery wall thickness. Blood pressure reduction could be explained on the basis of circulating renin angiotensin activity. On the contrary, radial artery wall thickness reduction was independent of the baseline circulating renin-angiotensin activity and was not correlated with the effects of AT(1) blockade on blood pressure, thus implying the involvement of local hemodynamic and/or cellular mechanisms. PMID- 10720882 TI - Role of voltage-dependent and Ca(2+)-activated K(+) channels on the regulation of isometric force in porcine coronary artery. AB - We investigated the role of K(+) channels in the regulation of vascular tone in de-endothelialized porcine coronary artery. Isometric force and intracellular Ca(2+) ([Ca(2+)](i)) under resting conditions were increased by treatment with 4 aminopyridine (4-AP, 1 mM), an inhibitor of voltage-dependent K(+) (K(v)) channels, but not by tetraethylammonium chloride (TEA, 1 mM) or charybdotoxin (100 nM), both inhibitors of Ca(2+)-activated K(+) (K(Ca)) channels, or glibenclamide (10 microM), an inhibitor of ATP-sensitive K(+) channels. Under stimulated conditions with 9,11-dideoxy-11alpha, 9alpha-epoxymethano prostaglandin F(2alpha) (U46619), 4-AP as well as TEA or charybdotoxin increased isometric force and [Ca(2+)](i), but not glibenclamide. 4-AP was the most potent in terms of depolarization of membrane potential compared with TEA or glibenclamide in the presence or absence of EGTA. In the presence of U46619, a high concentration of 4-AP (10 mM) caused a further contraction with oscillations. The force oscillations induced by 4-AP were inhibited by diltiazem (10 microM), an inhibitor of voltage-dependent Ca(2+) channels, or TEA (1 mM), but not by glibenclamide (10 microM). These force oscillations may be associated with the periodic activation of K(Ca) channels. These findings suggested that 4 AP-sensitive K(v) channels play an important role in the control of vascular tone in both resting and stimulated conditions. Moreover, under stimulated conditions, K(Ca) channels also have an important role in the regulation of vascular tone. Dysfunction of these channels induces abnormal vasoconstriction and may be implicated in vascular diseases such as hypertension and vasospasm. PMID- 10720883 TI - Off-line analysis of red blood cell velocity in renal arterioles. AB - Videomicroscopic methods with off-line analysis of microcirculatory parameters by multifunctional computer-assisted image analysis systems have significant advantages for in vivo microvascular research. A limitation of these methods is, however, that red blood cell velocities (V(RBC)) exceeding 2 mm/s cannot be measured using standard video framing rates. In the present study, a high-speed video camera, recording up to 600 frames per second, was incorporated in the set up, and V(RBC) was measured off-line with the line-shift-diagram method. The aim of this study was to test the reproducibility and validity of the method using a high-speed video camera and to evaluate its applicability in vivo. V(RBC) were measured in arterioles of the split hydronephrotic kidney. The intra- and interindividual variability was small for V(RBC) below 40 mm/s. The validity of the method was tested using the mass conservation principle and found to be at least as good as that of the dual-slit photometric technique. The present approach extends the application of videomicroscopy coupled to image analysis systems to the analysis of high V(RBC). PMID- 10720884 TI - Reactive oxygen species and acute modulation of albumin microvascular leakage in the microcirculation of diabetic rats in vivo. AB - Endothelial cells have been reported to generate reactive oxygen species such as the superoxide anion, hydrogen peroxide, and the hydroxyl radical. The aim of this work was to evaluate the role of reactive oxygen species in diabetes-induced changes in vascular permeability. Intravital videomicroscopy was used to study albumin microvascular leakage in the cremaster muscle. The extravasation of a fluorescent macromolecular tracer (FITC-albumin) was measured for 1 h and, after computer-aided image analysis, was expressed as variations of normalized gray levels (arbitrary units). The extravasation of the macromolecular tracer was greater in diabetic rats (5.28 +/- 1.29 vs. 1.96 +/- 0.41 AU at 1 h in diabetic and control rats, respectively). Administration of superoxide dismutase (SOD), which dismutates.O(-)(2) to H(2)O(2), and of catalase which reacts with H(2)O(2) to form H(2)O and molecular oxygen failed to inhibit the increased extravasation of the macromolecular tracer when administered separately. However, a significant inhibition of diabetic increase in albumin extravasation was found when these 2 drugs were administered simultaneously, and in this case, the extravasation of the macromolecular tracer at 1 h was similar in diabetic rats (2.11 +/- 0.61 AU) and normoglycemic rats (1.43 +/- 0. 48 AU). No difference was found in adherent leukocytes or in the leukocyte rolling flux between diabetic and normoglycemic rats. We conclude that reactive oxygen species are responsible for an increase in microvascular permeability likely via leukocyte-independent mechanisms. PMID- 10720885 TI - Neural endothelin in hypertension: increased expression in ganglia and nerves to cerebral arteries of the spontaneously hypertensive rat. AB - Endothelin has previously been localised in perivascular nerves of the rat basilar artery. Considering its potent vasoconstrictor and mitogenic properties on vascular smooth muscle, the potential role of a neural source of this peptide in hypertension has been investigated. The trigeminal, superior cervical and sphenopalatine ganglia of Wistar Kyoto (WKY) rats and spontaneously hypertensive rats (SHR) at 16 weeks of age have been examined for immunolocalisation of endothelin at the light and electron microscope level. At the light microscope level, neurones immunopositive for endothelin were detected in these ganglia of the SHR but were not seen in ganglia from WKY rats. This difference was particularly marked in the trigeminal ganglia where endothelin-positive neurones colocalised with substance P immunoreactivity. Using in situ hybridisation techniques, endothelin-1 mRNA was localised to the cytoplasm of neurones in the ganglia and was more prominent in the SHR. At the electron microscope level, endothelin-immunoreactivity was localised at the peripheral perikarya of some neuronal cell bodies of the trigeminal, superior cervical and sphenopalatine ganglia of WKY rats but was more prominent with heavy labelling throughout the cytoplasm of neurones in the SHR. Notably, in the trigeminal ganglia of the SHR only, some endothelin-immunopositive nerve fibres appeared to be damaged and contained vacuoles with granular material. Ultrastructural examination of the basilar artery revealed an increased number of endothelin-positive axons in the SHR, but these axons usually showed selective damage. In summary, in the SHR, there was a marked increase in endothelin particularly in sensory neurones projecting to the basilar artery which also appear to be undergoing degenerative changes. An increased neural source of endothelin in the SHR may contribute to the development of hypertension or may be a consequence of selective degenerative change. PMID- 10720886 TI - Effect of TGF-beta(1) antisense S-oligonucleotide on synthesis and accumulation of matrix proteoglycans in balloon catheter-injured neointima of rabbit carotid arteries. AB - Arterial matrix proteoglycans (PG) are necessary for the maintenance of viscoelastic properties of the vessel wall, but excess levels, particularly of versican and biglycan in primary and restenotic intimal thickenings, are correlated with increased tissue volume and with atherogenicity. There is good evidence that the primary stimulus to increased PG synthesis, including versican and biglycan, is transforming growth factor-beta(1) (TGF-beta(1)). The aim of this study was to determine the effects of reducing endogenous TGF-beta(1) on rates and patterns of PG synthesis and on versican, biglycan and decorin accumulation in vivo. Rabbit common carotid arteries subjected to balloon catheter injury were treated with a TGF-beta(1) antisense phosphorothioate oligonucleotide applied in a pluronic gel to the adventitia. Control animals received a nonsense oligonucleotide or gel alone. TGF-beta(1) antisense (1) significantly (p < 0.005) inhibited, at day 2, the balloon catheter-induced increase in TGF-beta(1) mRNA relative to beta-actin mRNA; (2) inhibited intimal thickening at 23 days by approximately 40% (p < 0.05); (3) inhibited (p < 0.05) PG synthesis, measured by autoradiographic detection of [(3)H]glucosamine, in the media of day 2 ballooned carotids and in the subendothelial zone of day 23 neointima, and (4) decreased immunostaining intensity for versican (p < 0.03) and TGF-beta(1) (p < 0.001) in the neointima. Biglycan was reduced to a lesser extent but not significantly and decorin was not affected. Proliferating cell nuclear antigen indices were variable and not significantly changed. These findings confirm a role for TGF-beta(1) in developing neointima and demonstrate a specific effect on the synthesis, distribution, and accumulation of matrix PG, particularly versican. PMID- 10720887 TI - Effect of venous and lymphatic congestion on lymph capillary pressure of the skin in healthy volunteers and patients with lymph edema. AB - The aim of the present study was to assess the influence of venous and lymphatic congestion on lymph capillary pressure (LCP) in the skin of the foot dorsum of healthy volunteers and of patients with lymph edema. LCP was measured at the foot dorsum of 12 patients with lymph edema and 18 healthy volunteers using the servo nulling technique. Glass micropipettes (7-9 microm) were inserted under microscopic control into lymphatic microvessels visualized by fluorescence microlymphography before and during venous congestion. Venous and lymphatic congestion was attained by cuff compression (50 mm Hg) at the thigh level. Simultaneously, the capillary filtration rate was measured using strain gauge plethysmography. The mean LCP in patients with lymph edema increased significantly (p < 0.05) during congestion (15.7 +/- 8.8 mm Hg) compared to the control value (12.2 +/- 8.9 mm Hg). The corresponding values of LCP in healthy volunteers were 4.3 +/- 2.6 mm Hg during congestion and 2.6 +/- 2.8 mm Hg during control conditions (p < 0.01). The mean increase in LCP in patients with lymph edema was 3.4 +/- 4.1 mm Hg, and 1.7 +/- 2.0 mm Hg in healthy volunteers (NS). The maximum spread of the lymph capillary network in patients increased from 13.9 +/- 6.8 mm before congestion to 18.8 +/- 8.2 mm during thigh compression (p < 0.05). No increase could be observed in healthy subjects. In summary, venous and lymphatic congestion by cuff compression at the thigh level results in a significant increase in LCP in healthy volunteers as well as in patients with lymph edema. The increased spread of the contrast medium in the superficial microlymphatics in lymph edema patients indicates a compensatory mechanism for lymphatic drainage during congestion of the veins and lymph collectors of the leg. PMID- 10720888 TI - Guanylins - are they of nephrological relevance? PMID- 10720889 TI - Plasma concentration of uroguanylin in patients on maintenance dialysis therapy. AB - BACKGROUND: Uroguanylin, originally isolated from urine, is a new natriuretic peptide. Its plasma level is increased in association with renal impairment and fluid retention in patients with renal diseases. METHODS: Uroguanylin concentrations were measured in patients on hemodialysis (HD, n = 76) and those on continuous ambulatory peritoneal dialysis (CAPD, n = 10) using a sensitive ra- dioimmunoassay for human uroguanylin. RESULTS: Plasma concentrations of immunoreactive (ir)-uroguanylin in the patients on HD and CAPD (212.0 +/- 17.4 and 245.3 +/- 39.5 fmol/ml) were significantly higher than the value for the normal controls (5.0 +/- 0.3 fmol/ml). Plasma ir-uroguanylin levels before the start of regular HD were correlated with predialysis excess weight based on their dry weights (r = 0.33, p < 0.01) and with dialysis duration (r = 0.26, p < 0.05). The plasma levels in patients with HD, for whom high-flux membranes were used, were decreased at the end of regular HD as compared with the prior levels (p < 0.05), but not in those who underwent HD with conventional membranes. CONCLUSION: These findings suggest that the plasma ir-uroguanylin level is related to the patient's volume status as well as renal impairment. Whether the accumulation of uroguanylin has a pathological effect has yet to be determined. PMID- 10720890 TI - The ferroxidase activity of caeruloplasmin is reduced in haemodialysis patients. AB - Increased free-radical production leading to oxidative stress may contribute to the development of cardiovascular complications in haemodialysis patients. The ferroxidase activity of caeruloplasmin forms an important component of antioxidant defences in body fluids. The aim of this study was to assess ferroxidase activity in haemodialysis patients. Venous blood was collected from 83 haemodialysis patients immediately prior to and after dialysis and from 52 healthy controls. Immunoreactive caeruloplasmin was measured by rate nephelometry, and ferroxidase activity determined by measuring loading of ferrous iron onto iron-free transferrin. A significant reduction in ferroxidase activity was observed in dialysis patients when compared with controls (37 +/- 1.20 and 46 +/- 1.14 mU/l, respectively; p < 0.001). Following dialysis, ferroxidase activity rose significantly to 41 +/- 1.16 mU/l, with a significant difference still remaining between control and patient ferroxidase activity (p < 0.005). Immunoreactive caeruloplasmin was found to be similar in all groups (before dialysis 0.40 +/- 0.07 g/l, after dialysis 0.39 +/- 0.07 g/l, control 0.42 +/- 0.09 g/l: p = NS). A significant difference in caeruloplasmin-specific activity was therefore observed between predialysis, postdialysis and control samples (97 +/- 2.31, 105 +/- 1.74 and 112 +/- 1.51 mU/g; p < 0.001, p < 0.01, respectively). Ferroxidase activity of caeruloplasmin is impaired in renal failure. Inhibition of caeruloplasmin ferroxidase activity in dialysis patients may contribute to increased oxidative stress in these patients. PMID- 10720891 TI - Protective role of extracellular superoxide dismutase in hemodialysis patients. AB - BACKGROUND: The superoxide anion and other oxygen radicals have been implicated in the progression of chronic renal failure, and are removed by extracellular superoxide dismutase (EC-SOD) in the extracellular space on the surface of the endothelium. A single-base substitution of the EC-SOD gene which reduces the binding capability to endothelial cells resulting in an increased serum concentration, has been identified in healthy persons and hemodialysis patients. RESULTS: The proportion of patients with this mutation among hemodialysis patients in each 20 months' duration after the initiation of hemodialysis was retrospectively studied. The percentage of substitution-positive patients declined 80 months after the start of hemodialysis in non-DM patients. In contrast, in DM patients, the rapid decrease was obvious as early as 40 months after the initiation of hemodialysis. By prospective study for 5 years, there were significant differences in the survival rate between patients with and without R213G in DM, but not in non-DM patients. Among those who died, the incidence of ischemic heart disease and cerebrovascular disease in cases with R213G was significantly higher than in cases without R213G. CONCLUSION: These results suggest that the presence of a substitution in the EC-SOD gene at the heparin-binding domain could be a prognostic marker of dialysis patients. PMID- 10720892 TI - Restoration of impaired T-cell proliferation after parathyroidectomy in hemodialysis patients. AB - BACKGROUND/AIMS: Severe secondary hyperparathyroidism is not infrequent in hemodialysis patients and recent studies suggest that parathyroid hormone (PTH) may play a role in the genesis of cell immunity abnormalities in uremia. The aim of the present study is to describe the effect of parathyroidectomy on T- and B cell functions in hemodialysis patients. METHODS: The study was performed on 6 patients with severe secondary hyperparathyroidism. iPTH, B, CD4(+), CD8(+), total number of lymphocytes, lymphoproliferative response to PHA, PWM and Candidin, and IgG, IgM, IL-2 production in vitro were determined 1 day before and 4 months after parathyroidectomy. RESULTS: The lymphoproliferative response to PHA increased significantly after parathyroidectomy. We also observed a trend to an increase in production of IgG and IgM after PWM stimulation before therapy. CONCLUSION: The present study suggests that patients with extremely high levels of PTH show a complete restoration of impaired T-cell proliferation after parathyroidectomy. PMID- 10720893 TI - Urinary N-acetyl-beta-D-glucosaminidase and neopterin aid in the diagnosis of rejection and acute tubular necrosis in initially nonfunctioning kidney grafts. AB - AIM: The study aimed at investigating urinary neopterin, a marker of cellular immune response, and urinary N-acetyl-beta-D-glucosaminidase (NAG), a marker of tubular damage, as noninvasive means to differentiate between acute tubular necrosis (ATN) and rejection in initially nonfunctioning (INF) human renal transplants. METHODS: Seventy-two renal transplant patients were studied. Forty five of them experienced an uncomplicated early posttransplant course, 27 patients suffered from INF. Twenty-two patients experienced ATN, 5 patients had a total of six biopsy-proven rejections. The NAG activity was measured by a colorimetric assay, neopterin by high-performance liquid chromatography. Receiver operating characteristics (ROC) analysis was applied to compute diagnostic performance and an optimal discriminating threshold. RESULTS: Demographic characteristics (age, gender, cold and warm ischemia periods, HLA mismatches) and posttransplant urinary NAG and neopterin excretions did not differ between ATN and rejection groups. Both urinary NAG and neopterin excretions were lower in the control group (NAG 1.8 +/- 1.0 U/mmol urinary creatinine; neopterin 270 +/- 126 nmol/mmol urinary creatinine; mean +/- SD) as compared with the ATN group (NAG 12 +/- 10 U/mmol, p < 0.001 vs. control group; neopterin 303 +/- 195 nmol/mmol, n.s.) and the rejection group (NAG 7 +/- 8 U/mmol, p < 0. 01; neopterin 508 +/- 419 nmol/mmol, p < 0.01). The ratio of urinary neopterin to NAG excretion (uNNR; dimension nmol neopterin/U NAG activity) increased during rejections as compared with ATN (139 +/- 74 vs. 50 +/- 38 nmol/U, p < 0.01). The area under the ROC curve for uNNR was 0.88 +/- 0.07 (p < 0.001). Applying a ROC-estimated optimal discriminator of uNNR (80 nmol/U), 16 patients with ATN and all six rejection episodes were classified correctly. CONCLUSION: The uNNR provides a noninvasive means to aid in the differential diagnosis of rejection and ATN in INF human renal transplants. PMID- 10720894 TI - Renal findings and glomerular pathology in diabetic subjects. AB - BACKGROUND: To describe the relationship between proteinuria, hematuria, and renal insufficiency, on one hand, and glomerular pathology, on the other hand, in a consecutive biopsy series of diabetic patients. SUBJECTS AND METHODS: All diabetic subjects (n = 200) biopsied from 1979 to 1995 at Tampere University Hospital were identified in retrospect. The clinician-based indication (any unexplained renal finding) for renal biopsy was consistent during the years and was: proteinuria alone in 68%; combined with hematuria in 10%; with renal insufficiency in 10%; with both in 9%, and with isolated hematuria or renal failure in 3%. One third of the subjects had proteinuria of >/=3 g/24 h and 16% a serum creatinine level of >/=200 microM. Glomerulopathy was found in 171 specimens and defined as nodular diabetic (group A), diffuse diabetic (group B) and primary (group C). The 24-hour urinary protein excretion rate [mean (range)] was 3.5 (1.6-6.9), 1.0 (0.5-3.5), and 3.6 (1.1-6.6) g in groups A, B and C, respectively (ANOVA p = 0.001). The corresponding serum creatinine values [mean (SD)] were 175 (115), 105 (142) and 169 (138) microM (p = 0.001). RESULTS: Nodular diabetic glomerulopathy was found in 40%, diffuse diabetic glomerulopathy in 42% and primary glomerulopathy in 18%. A primary glomerulopathy was found in any indication and in both types of diabetes (prevalence range 14-26%). The best multivariate logistic regression model obtained (chi(2) = 13.5, p = 0.008) in predicting the presence of diabetic glomerulosclerosis (group A + B) in contrast to a primary glomerulopathy (group C) included retinopathy (p = 0.04), renal insufficiency (p = 0.03), hematuria (p = 0.12) and type of diabetes (p = 0.10). CONCLUSION: In this series of diabetic subjects, biopsied due to proteinuria, hematuria and not severe renal insufficiency, 18% had evidence of a primary glomerulopathy. PMID- 10720895 TI - Effect of vitamin E and pentoxifylline on glycerol-induced acute renal failure. AB - The pathogenesis of acute renal failure may involve, among other causes, ischemia, vascular congestion, arachidonic acid pathways, and reactive oxygen metabolites. The aim of this study is to evaluate the effects of pentoxifylline and vitamin E on the prevention of experimental acute renal failure induced by glycerol. Eighty-five Sprague-Dawley rats weighing 170-230 g were included in the study. The rats were randomly divided into four groups: group 1 was given 1 ml saline; group 2, glycerol; group 3, glycerol plus vitamin E, and group 4, glycerol plus pentoxifylline. Extent of histological renal tubular necrosis and regeneration in each animal were graded. Blood urea nitrogen, serum creatinine, and creatine kinase concentrations were measured. Mean blood urea nitrogen and serum creatinine concentrations and tubular injury scores were significantly lower in group 1 than in groups 2-4 (p < 0.001), but there were no significant differences among groups 2-4. We conclude that postinsult administration of vitamin E and pentoxifylline does not have a beneficial effect on prevention and severity of acute renal failure and that controlled, multicenter studies involving a large number of patients are needed to clarify this subject. PMID- 10720896 TI - Heparin treatment reduces glomerular injury in rats with adriamycin-induced nephropathy but does not modify tubulointerstitial damage or the renal production of transforming growth factor-beta. AB - In this study we investigated the effect of heparin on renal injury and renal transforming growth factor-beta (TGF-beta) production in adriamycin (AD)-injected rats. Thirty-nine female Wistar rats were injected with AD (3.5 mg/kg body weight, i.v.) and 27 with 0.15 M NaCl solution (group C). Fifteen days later we started to inject heparin, 500 U/day, s.c., in 20 of the AD-injected animals (AD H group). Three months after beginning treatment, urine samples were collected to quantify albumin, creatinine and TGF-beta. The rats were killed and the kidneys removed for histological, immunohistochemical, ELISA and RNA studies. All AD injected animals showed structural renal changes (p < 0.05). However, the glomerular alterations were less intense in rats from group AD-H (p < 0.05). The percentage of glomerulosclerosis was 0.11 +/- 0.08 in group C, 14.7 +/- 12.8 in group AD (treated only with AD) and 3.42 +/- 2.3 in group AD-H. Renal cortex immunostaining for TGF-beta and mRNA content of this polypeptide was higher in both groups of animals injected with AD compared to controls (p < 0.05). These animals also presented a higher rate of urinary TGF-beta excretion (p < 0.05), which was 202 +/- 11 in group C, 1,103 +/- 580 in group AD and 1,564 +/- 328 pg/mg Ucreat in group AD-H. However, TGF-beta activity in the glomerular conditioned media from the rats of group AD was higher than in the glomerular conditioned media from the rats of group AD-H. In conclusion, treatment with heparin reduces glomerular damage in rats with AD-induced nephropathy but does not modify tubulointerstitial lesions or the renal production of TGF-beta. PMID- 10720897 TI - Hemolytic uremic syndrome associated with influenza A virus infection in an adult renal allograft recipient: case report and review of the literature. AB - Hemolytic uremic syndrome (HUS) is a rare but serious complication following renal transplantation. It usually develops early after transplantation, and ciclosporin treatment is the most common triggering factor. We report the case of a 35-year-old male with posttransplant HUS which developed 1 year after renal transplantation. He became febrile 4 days before the onset of HUS, and the significant rise in viral titer confirmed the diagnosis of influenza A virus infection. The association of ciclosporin treatment with HUS was unlikely, because of the late onset of HUS and the low ciclosporin trough levels. The patient was treated successfully without a dose reduction of ciclosporin. An etiologic relationship between influenza A virus and HUS was highly probable in our patient. We also review a total of 156 adult cases with HUS after renal transplantation described in the literature. The prognosis of posttransplant HUS differs according to the cause. The advent of ciclosporin has improved the graft survival rate and mortality of patients with rejection-induced HUS. PMID- 10720898 TI - Exacerbation of renal failure due to hypothyroidism in a patient with ischemic nephropathy. AB - A case of acute-on-chronic renal failure in a 70-year-old woman with ischemic nephropathy and primary hypothyroidism is presented. Her renal function became progressively worse as the level of serum creatinine increased from 283 to 628 micromol/l (3.2-7.1 mg/dl) within 8 months. Her thyroid function had been normal before the exacerbation of renal failure, but it was markedly reduced with a marked elevation of serum thyroid-stimulating hormone. Thyroid hormone replacement therapy resulted in rapid improvement of the renal function to 159 micromol/l (1.8 mg/dl) of serum creatinine. The development of primary hypothyroidism seemed to worsen the already impaired renal function in this case. We suggest the assessment of thyroid function in patients with unexplained deterioration of renal failure. PMID- 10720899 TI - Effects of potassium citrate/citric acid intake in a mouse model of polycystic kidney disease. AB - The kidney function in a model of autosomal dominant polycystic kidney disease (PKD), the Han:SPRD rat, is dramatically improved by chronic ingestion of a solution of potassium citrate and citric acid (KCitr). This study investigated whether this treatment would also be beneficial in the pcy/pcy mouse, a model of autosomal recessive PKD. Starting at 1 month of age, male CD-1 pcy/pcy and normal CD-1 mice were provided with a solution of 55 mM K(3) citrate/67 mM citric acid or tap water to drink. The pcy/pcy mice on the KCitr solution failed to grow normally and showed elevated plasma urea levels when compared to water-drinking littermates. Growth of normal CD-1 mice was not affected by KCitr intake. The pcy/pcy mice were then provided with a more dilute solution of KCitr to drink: this resulted in greater kidney wet and dry weights and a higher kidney weight/body weight ratio, but no beneficial effects. We conclude that pcy/pcy mice cannot tolerate a high level of KCitr intake and that a lower level is of no benefit. Whether KCitr therapy would be helpful in patients with PKD is still an open question. PMID- 10720900 TI - Lewis(y) expression and renal tubular atrophy in IgA nephritis. PMID- 10720903 TI - Reply: decreased plasma glutathione peroxidase activity in uraemic patients PMID- 10720901 TI - Skin biopsy in Berger's disease: is it really useful in the diagnosis? PMID- 10720902 TI - Decreased plasma glutathione peroxidase activity in uraemic patients. PMID- 10720904 TI - Low prevalence of serum HGV-RNA among patients with chronic renal failure without dialysis. PMID- 10720905 TI - Low temperature hemodialysis prevents hypote nsive episodes by reducingnitric oxide synthesis. PMID- 10720906 TI - Annual changes of dialysate interleukin-6 and peritoneal ultrafiltration capacity. PMID- 10720907 TI - Effect of low-protein diet supplemented with keto acids on progression of disease in patients with chronic renal failure. PMID- 10720908 TI - Anti-HCV antibodies in primary glomerular diseases in India. PMID- 10720909 TI - Effect of oral levamisole supplementation to hepatitis B vaccination on the rate of immune response in chronic hemodialysis patients. PMID- 10720910 TI - Influence of hepatitis C virus infection upon parenteral iron and erythropoietin responsiveness in regular haemodialysis patients. PMID- 10720911 TI - Isolated ciclosporin-associated arteriopathy does not deteriorate residual renal function in patients with kidney transplantation. PMID- 10720912 TI - Comparative study of brain lesions detected by magnetic resonance imaging between strabismus and nonstrabismus in infancy. AB - PURPOSE: To elucidate the causative factors in infantile esotropia, we evaluated morphological abnormalities in brain structures of esotropia patients showing any abnormal neurological signs in comparison to those of normal controls. METHODS: Sixty-five developmentally normal children participated in this study. Of these 65, 38 demonstrated infantile esotropia and 27 were normal controls. All underwent magnetic resonance imaging (MRI) of the brain between 2 and 30 months. RESULTS: Abnormal brain findings were noted in 3 (7. 9%) children in the strabismus group, whereas none of the children in the normal control group showed brain lesions. In these 3 cases, brain lesions involved periventricular leukomalacia, enlargement of the lateral ventricles with hypoplasia of the corpus callosum and myelination delay at the anterior horn adjacent to the lateral ventricles. CONCLUSIONS: Brain lesions that may disturb normal maturation of the visuomotor system and eventually lead to strabismus could be found in some patients without any episode that would cause birth injury. PMID- 10720913 TI - Tolerance of N-chlorotaurine, a new antimicrobial agent, in infectious conjunctivitis - a phase II pilot study. AB - N-Chlorotaurine (NCT) is an endogenous microbicidal oxidant. This open pilot study (phase IIa) with 9 patients was done to gain first knowledge on the tolerance of NCT in infectious conjunctivitis. By application of 1% NCT 5 times a day, no adverse effects could be observed. All 6 subjects with bacterial conjunctivitis were cured within 3-5 days. Two subjects with epidemic keratoconjunctivitis were treated for 7-10 days and 1 subject with herpes simplex blepharitis for 3 days with no rapid improvement but probable mitigation of inflammation. Therefore, NCT seems to be useful in the treatment of infectious conjunctivitis, and further investigation on its therapeutic efficacy is suggested. PMID- 10720914 TI - Primary open-angle glaucoma is associated with sleep apnea syndrome. AB - INTRODUCTION: The etiology of primary open-angle glaucoma remains unclear. Various risk factors, including vascular abnormalities, have been associated with this disease. Sleep-associated diseases, like sleep apnea syndrome, might also represent a risk factor. Sleep apnea syndrome is characterized by repetitive upper airway obstructions during sleep, inducing hypoxia and sleep disruption with the risk of cardiovascular and neurological sequelae. In this study, we determined the prevalence of sleep apnea syndrome in primary open-angle glaucoma patients. METHODS: Overnight transcutaneous finger oximetry was performed in 30 consecutive patients having primary open-angle glaucoma. We assessed the oximetry disturbance index during night sleep, a parameter used to diagnose sleep apnea syndrome and to grade its severity. RESULTS: Sleep apnea syndrome was more prevalent among primary open-angle glaucoma patients compared to normal historic controls of the same age and sex distribution (chi(2) = 9.35, d.f. = 3, p < 0.025). The oximetry disturbance index grade was significantly larger in the primary open-angle glaucoma group compared to normal controls (U = 3, 352, p = 0.01). According to the oximetry disturbance index, 20% (6/30) of primary open angle glaucoma patients had sleep apnea syndrome. CONCLUSION: Primary open-angle glaucoma is associated with sleep apnea syndrome. Early recognition and treatment of sleep apnea syndrome are important to avoid cardiovascular and neurological complications. PMID- 10720915 TI - Visual field defects and normal nerve fiber layer: may they coexist in primary open-angle glaucoma? AB - The retinal nerve fiber layer (RNFL) is the anatomical structure most sensitive to glaucoma injury. Before a functional loss such as a visual field defect is displayed, a large number of nerve fibers can be damaged. However, there are glaucoma patients in which an apparently normal RNFL coexists with evident visual field defects. A total of 54 eyes affected with primary open-angle glaucoma were studied. Visual field was examined with the Humphrey Field Analyzer (Zeiss) using program 30-2. The Nerve Fiber Analyzer II (Laser Diagnostic Technologies) was used to study the RNFL of these patients. Mean deviation of the visual field ranged from 6 to 31 dB in all eyes that were examined. The average thickness of the RNFL ranged from 20 to 90 microm. According to our previous experience 75 microm was fixed as the cutoff between normal and pathological values of RNFL thickness. We identified 5 eyes with a RNFL thickness over 75 microm and a visual field with a mean deviation over 6 dB; 9% of the studied eyes were found to have a visual field defect with no changes in RNFL. We conclude that not all subjects have the same number of fibers at birth and that it is therefore possible to underestimate the RNFL changes. Our study illustrates that the concept of normal and altered has to be considered as a relative one for all the aspects characterizing the glaucomatous disease. PMID- 10720916 TI - Pigment epithelium defects after submacular surgery for choroidal neovascularization: first results. AB - It was the aim of this study to compare postoperative pigment epithelium defects after submacular surgery for well-defined choroidal neovascularization (CNV) with preoperative fluorescein angiogram (FAG), indocyanine green angiogram (ICG), and intraoperative findings of the excised neovacularization. Surgical removal of the CNV was videotaped. By means of a gauge the absolute size of an anatomic structure was determined which was visible on video, FAG, and ICG as well. Postoperatively another FAG was performed to further examine the funduscopically visible pigment epithelium defect. By comparison with the above-mentioned anatomic structure it was thus possible to determine the exact size of CNV and pigment epithelium defect. The extent of the pigment epithelium defect as determined on postoperative FAG after submacular surgery surpasses the size of CNV as measured in FAG, ICG, and anatomical preparation. PMID- 10720918 TI - Risk factors in retinopathy of prematurity. a multivariate statistical analysis. AB - OBJECTIVE: To analyze risk factors in retinopathy of prematurity (ROP). PATIENTS AND METHODS: Four hundred forty-seven surviving very-low-birth-weight infants (birth weight AB > AA. PDGF AA also dose dependently stimulated PI 3 kinase activity measured in antiphosphotyrosine immunoprecipitates of treated cells. A comparison of PI 3 kinase activity in antiphosphotyrosine immunoprecipitates from mesangial cells stimulated with three different PDGF isoforms showed significant activation of this enzyme with a decreasing order of activity: BB > AB > AA. CONCLUSION: Taken together, these data demonstrate that all three isoforms of PDGF significantly stimulate PLC gamma 1 and PI 3 kinase, two enzymes necessary for both DNA synthesis and directed migration. However, activation of alpha receptor by PDGF AA with a subsequent increase in PLC and PI 3 kinase activities is not sufficient to induce these biological responses in mesangial cells. These data indicate that the extent of activation of signal transduction pathways may be a major determinant of the biological activity of different PDGF isoforms in mesangial cells. PMID- 10720946 TI - Inhibition of monocyte chemoattractant protein-1 expression in tubular epithelium attenuates tubulointerstitial alteration in rat Goodpasture syndrome. AB - BACKGROUND: To examine the role of monocyte chemoattractant protein-1 (MCP-1) expressed by tubular epithelium in tubulointerstitial alterations in situ, the level of MCP-1 mRNA in tubular epithelium was lowered selectively in the rat model of Goodpasture syndrome (GPS). METHODS: Intravenously administered antisense oligodeoxynucleotide (ODN) is taken up by renal tubular epithelium and has been found to block expression of target genes in rats. MCP-1 antisense ODN was injected into GPS rats every second day from days 27 to 35 after immunization (this represents the time when renal MCP-1 mRNA level was increased and interstitial mononuclear cell infiltration was aggravated). RESULTS: In addition to a reduction in the level of tubular MCP-1 mRNA, antisense ODN treatment attenuated monocyte infiltration significantly and preserved renal function in GPS rats. However, ODN injection did not affect glomerular MCP-1 expression and glomerular histopathology, and there were no significant changes in the urinary protein excretion rate. CONCLUSION: Our findings provide direct evidence that MCP 1, expressed by tubular epithelium, plays a pivotal role in mediating secondary tubulointerstitial alterations in the GPS model. PMID- 10720947 TI - Reciprocal balance of hepatocyte growth factor and transforming growth factor beta 1 in renal fibrosis in mice. AB - BACKGROUND: Transforming growth factor-beta 1 (TGF-beta 1) plays a central role in the pathogenesis of renal fibrosis. In contrast, hepatocyte growth factor (HGF) may be an important molecule for tissue repair. As TGF-beta 1 is a suppressor molecule for HGF expression, we asked whether a decrease in HGF expression would be accompanied by an increase in TGF-beta 1 and whether the progression of renal fibrosis would be modulated. METHODS: We used the ICR strain derived glomerulonephritis (ICGN) mice as a model of chronic renal disease and examined changes in local HGF expression during the natural course of renal fibrosis. To determine the significance of intrinsic HGF noted during progression of renal fibrosis, we administered an anti-HGF antibody to mice at the early stage of renal fibrosis. RESULTS: At an early stage of renal fibrosis, the mice showed strong peritubular HGF expression, coinciding with tubular proliferation. In the late stages, the renal HGF level was markedly decreased, coinciding with a reduction in proliferative tubular areas. Renal TGF-beta 1 levels were increased in accordance with expansion of fibrotic areas. Notably, the anti-HGF antibody treatment of early-stage mice decreased the HGF level and reduced tubular areas, whereas collagen-deposited areas were expanded in parallel with increased TGF beta 1 levels. Consequently, in HGF-neutralized mice, there was a rapid progression of renal dysfunction. CONCLUSIONS: Not only an increase in TGF-beta 1 level, but also a decrease in local HGF expression may be responsible for the manifestation of renal fibrosis, particularly tubular destruction. PMID- 10720948 TI - Induction of proliferation and apoptotic cell death via P2Y and P2X receptors, respectively, in rat glomerular mesangial cells. AB - BACKGROUND: Cell surface receptors for adenosine 5'-triphosphate (ATP; P2 receptors) have been subdivided into two families: ligand-gated ion channels (P2X1-7) and G-protein-coupled (P2Y1-8) receptors. We investigated the potential role of P2 receptors on rat glomerular mesangial cells. METHODS: To investigate cell proliferation, DNA synthesis was assayed by measuring [3H]thymidine incorporation into DNA. For detecting apoptosis, morphological features, DNA fragmentation, and exposure of phosphatidylserine on the outside surface of the cell membrane were investigated. Expression of mRNA and distribution of receptors were detected by reverse transcription-polymerase chain reaction and immunohistochemistry, respectively. RESULTS: ATP triggered a dose-dependent increase in DNA synthesis. This response was also induced by uridine triphosphate (UTP), an agonist equipotent with ATP at P2Y2 and P2Y4 receptors; both P2Y2 and P2Y4 mRNA are expressed in glomerular mesangial cells and isolated glomeruli. In contrast, the P2X7 receptor agonist 2'-83'-O-(4-benzoyl benzoyl) ATP (BzATP) caused a decrease in cell number. BzATP produced DNA cleavage and exposure of phosphatidylserine on the outside of the cell membrane. P2X7 receptors were distributed heterogeneously in unstimulated cells. The expression of P2X7 mRNA was maintained at a low level, but was induced by tumor necrosis factor-alpha. CONCLUSIONS: Stimulation of glomerular mesangial cells via P2Y2 and/or P2Y4 and via P2X7 receptors can induce proliferation and apoptotic cell death, respectively. The balance between proliferation and apoptosis will depend on the relative stimulation and expression of these P2 receptor subtypes, and could play an important role in normal and abnormal glomerular function. PMID- 10720949 TI - Overexpression of truncated I kappa B alpha potentiates TNF-alpha-induced apoptosis in mesangial cells. AB - BACKGROUND: Dysregulation of apoptosis is one of the likely underlying mechanisms of mesangial proliferative glomerulonephritis (GN), a disease in which proinflammatory cytokines exhibit a wide range of biological activities. Among them, tumor necrosis factor-alpha (TNF-alpha) induces two conflicting pathways, one leading to activation of the nuclear factor-kappa B (NF-kappa B), and the other leading to caspase-mediated apoptosis. We investigated whether or not specific inhibition of NF-kappa B affects TNF-alpha-induced apoptosis in rat mesangial cells (MCs). METHODS: To specifically inhibit NF-kappa B activation, we constructed a recombinant adenovirus vector expressing a truncated form of I kappa B alpha (AdexI kappa B delta N) that lacks the phosphorylation sites essential for the activation of NF-kappa B. Electrophoretic mobility shift assay was performed to evaluate NF-kappa B activity. Nuclear morphology was observed by staining with Hoechst-33258. DNA fragmentation was detected using an ELISA kit with an antihistone antibody. To investigate the regulation of apoptosis, we measured caspase-3 and caspase-8 activity by ELISA, and examined the Bcl-2 and Bax protein level by Western blot. RESULTS: TNF-alpha-induced NF-kappa B activation was blocked by overexpression of I kappa B delta N. Overexpression of I kappa B delta N potentiated TNF-alpha-induced apoptosis compared to mock transfection, and the potentiation was abolished by treatment with a caspase-3 inhibitor, Z-DEVD-FMK. Overexpression of I kappa B delta N augmented TNF-alpha induced caspase-3 and caspase-8 activity, but did not affect Bcl-2 or Bax protein expression. CONCLUSION: Overexpression of I kappa B delta N potentiates TNF-alpha induced apoptosis and augments caspase-8 and caspase-3 activity in rat MCs without changing Bcl-2 or Bax protein expression. These results suggest the potential usefulness of AdexI kappa B delta N to induce apoptosis in MCs under inflammatory conditions. PMID- 10720950 TI - Expression of apoptosis regulatory proteins in tubular epithelium stressed in culture or following acute renal failure. AB - BACKGROUND: While tubular cell death is a characteristic of acute renal failure (ARF), the molecular mechanisms that modulate this cell death are unclear. Cell fate in acute renal failure hinges on a balance of survival and mortality factors in a changing environment. We further explored this issue by studying selected cell death-related proteins in experimental renal failure. METHOD: The expression of genes that promote (c-myc, Bax, BclxS) or protect (Bcl2, BclxL) from cell death was studied by Northern blot, Western blot, and immunohistochemistry in murine kidneys following ARF induced by folic acid or in renal tubular epithelial cells (MCT) stressed in culture. RESULTS: Renal mRNA levels encoding for c-myc and BclxL were elevated in ARF while the Bcl2/Bax ratio was decreased (Bcl2 decreased and Bax increased; P < 0.05). Protein levels of BclxL increased and Bcl2 protein decreased. Expression of tumor necrosis factor (TNF-alpha), a mediator of ARF, was also increased. Immunohistochemistry further demonstrated that BclxL was increased in some tubuli and absent in others, while Bcl2 expression decreased diffusely. Bax staining was also patchy among tubuli and individual cells in the tubular wall and lumen. As a relative deficit of survival factors is present in ARF, MCT epithelium were deprived of serum survival factors. This resulted in apoptosis, decreased Bcl2/Bax and BclxL/Bax ratios (P < 0.05) and sensitization to TNF-alpha-induced apoptosis (P < 0.05). The latter was prevented by enforced overexpression of BclxL (P < 0.01). TNF-alpha increased the mRNA levels encoding for c-myc and decreased BclxL expression. Neither MCT cells nor the kidney expressed BclxS. CONCLUSIONS: A relative deficit of survival factors likely contributes to changes in levels of BclxL and Bax in ARF. These deficits predispose to cell death induced by persistent lethal factors such as TNF-alpha that is increased in ARF and a potential source of increased c-myc, a downstream facilitator of cell death. These findings implicate members of the Bcl2 family of proteins as regulators of tubular cell death in ARF and single them out as potential therapeutic targets. PMID- 10720951 TI - Effect of angiotensin-converting enzyme inhibition on growth factor mRNA in chronic renal allograft rejection in the rat. AB - BACKGROUND: Despite considerable progress in immunosuppression, the incidence of chronic renal allograft rejection has not decreased. Recent studies have revealed that angiotensin-converting enzyme (ACE) inhibition ameliorates graft arteriosclerosis, glomerulosclerosis, and tubular atrophy. Moreover, it decreases systemic and glomerular capillary hydrostatic pressure in a rat kidney allograft model. We evaluated the effects of the ACE inhibitor enalapril on cytokine and growth factor expression in chronically rejecting rat kidney allografts. METHODS: Kidneys of Fisher (F344) rats were orthotopically transplanted into Lewis (Lew) rats. To prevent acute rejection, cyclosporine A (1.5 mg/kg/day) was given to all recipients during the first 10 days after transplantation. Enalapril (60 mg/L) or vehicle was added to the drinking water 10 days after transplantation. Animals were harvested 20 weeks after transplantation for histologic and immunohistologic studies, as well as for evaluation of cytokine and growth factor mRNA by semiquantitative polymerase chain reaction. RESULTS: Controls developed severe signs of chronic rejection, such as glomerular and vascular lesions, associated with a large number of infiltrating leukocytes. Enalapril-treated animals developed less proteinuria and other signs of chronic rejection. The mRNA levels of transforming growth factor-beta 1 (TGF-beta 1), platelet-derived growth factor A and B chain (PDGF A and B), insulin-like growth factor-I (IGF-I), interleukin-1 (IL-1), and monocyte chemoattractant protein-1 (MCP-1) were significantly reduced in the enalapril group and were most pronounced for IL-1 and PDGF A. In addition, we found an increased level of renal angiotensinogen mRNA after treatment with enalapril. CONCLUSIONS: Treatment with enalapril attenuated the development of proteinuria, ameliorated morphological damage, decreased leukocyte infiltration, and prevented a rise in renal mRNA levels of growth factors and cytokines in kidney grafts in a rat model of chronic renal allograft rejection. PMID- 10720952 TI - Tandem antifibrotic actions of L-arginine supplementation and low protein diet during the repair phase of experimental glomerulonephritis. AB - BACKGROUND: Based upon the central role transforming growth factor-beta (TGF beta) overexpression appears to play in renal fibrotic diseases, we have recently advocated reduction of TGF-beta as a therapeutic target. As part of efforts to determine the strength of this approach, we have undertaken studies to quantitate the effects of currently used and promising therapies in terms of their potential to reduce markers of disease in anti-thymocyte-serum (ATS)-glomerulonephritis in the rat. Here we assess the therapeutic effect of L-arginine supplementation, which has been shown to reduce fibrosis in a number of hypertensive models, given alone or in combination with low protein diet and started 24 hours after disease induction. METHODS: Glomerulonephritis was induced by intravenous injection of OX 7 monoclonal antibody into 200 g Sprague-Dawley rats. Twenty-four hours later animals were placed in groups that were either untreated, treated with 1% L arginine in drinking water or 6% protein diets or both. On the fifth day of disease 24-hour urine specimens were collected and systemic blood pressure was measured. On the sixth day rats were anesthetized. Kidneys were perfused, tissue was taken for PAS staining and glomeruli were isolated. Aliquots of glomeruli were used for RNA preparation and for culture to determine 72-hour production of TGF-beta, fibronectin and plasminogen activator-type 1 (PAI-1), which were assayed by ELISA on culture supernatants. Measures of nitrate and nitrite (NOx) production included plasma NOx, urinary NOx and glomerular production of NOx in culture. RESULTS: All disease measures except proteinuria and including matrix accumulation, TGF-beta, fibronectin and PAI-1 production and mRNA expression for TGF-beta, fibronectin and PAI-1 were significantly and similarly reduced by about 50% in groups treated with L-arginine or with low protein diet. Proteinuria was reduced in low protein treated but not in L-arginine supplemented rats. Neither systemic blood pressure nor measures of NO synthesis showed differences between groups that could be attributed to L-arginine supplementation. In contrast, disease-related increases in glomerular production of NOx were markedly reduced by low protein. Combined therapy resulted in small, but statistically significant decreases in most measures of disease. CONCLUSIONS: L-arginine supplementation reduces fibrotic disease in ATS-induced glomerulonephritis if started after disease induction. The absence of evidence for increased NO production related to L-arginine supplementation suggests that L-arginine is acting here through different pathways from those demonstrated in hypertensive models of disease. The data support the ideas that TGF-beta reduction is a valid therapeutic target and that quantitation of TGF-beta reduction is a useful approach for comparing antifibrotic drug candidates. PMID- 10720954 TI - Hepatic alpha 2 mu-globulin localizes to the cytosol of rat proximal tubule cells. AB - BACKGROUND: Alpha 2 mu-Globulin (A2), an 18.6 kD protein of hepatic origin, accumulates in the proximal tubule as an abundant, 15.5 kD cleavage product termed "A2-fragment" (A2-f). A2-f facilitates proximal tubule fatty acid oxidation, presumably by binding hydrophobic ligands. This requires some A2-f to enter the cytosol of the renal epithelial cell (REC). The localization of A2/A2-f in the proximal tubule cell was evaluated in this study. METHODS: Immunoblot analysis of renal cortical homogenates separated by differential centrifugation and quantitative immunoelectron microscopy (IEM) was performed to localize A2/A2 f using an affinity-purified antibody that detects both proteins. To evaluate A2 as a physiologically relevant ligand, the accumulation of A2-f in the female rat kidney (normally devoid of A2-f) was examined after the induction of hepatic A2 synthesis. Ligand binding, uptake, and degradation assays were used to assess A2 processing by RECs in vitro. RESULTS: Although A2 and A2-f were detected in the "lysosomal" fraction, only A2-f was found in the soluble protein fraction. IEM confirmed the presence of significant signal in the vesicular and lysosomal as well as the cytosolic compartments. In contrast, both beta 2 mu globulin (B2) and cathepsin B were restricted to endosomes. In the female rat, induction of hepatic A2 production resulted in A2-f accumulation in the renal cortex. In RECs in culture, uptake of A2 and B2 demonstrated nonsaturable, nondisplacable surface binding and similar uptake rates. Compared with B2, A2 was markedly resistant to degradation. CONCLUSIONS: A fraction of A2 escapes lysosomal degradation, permitting A2-f to accumulate in the cytosol of the proximal tubule epithelial cell. A2 may represent an unusual example of a physiologic protein capable of accumulating in a distant cell type. PMID- 10720953 TI - Role of glomerular ultrafiltration of growth factors in progressive interstitial fibrosis in diabetic nephropathy. AB - BACKGROUND: The present in vivo and in vivo experiments were performed to test the hypothesis that in rats with glomerular proteinuria, the bioactive growth factors transforming growth factor-beta (TGF-beta) and hepatocyte growth factor (HGF) are ultrafiltered into tubular fluid, can interact with respective receptors in apical tubular cell membranes, increase the expression and basolateral secretion of C-C-chemokines, which interact with cells in the renal interstitium and indirectly cause myofibroblasts to increase the expression of extracellular matrix proteins. METHODS: HGF and TGF-beta were measured by Western blot and bioassay in glomerular ultrafiltrate that was collected by nephron micropuncture from rats with diabetic nephropathy and control rats. Proximal tubular and collecting duct cells were incubated with diluted proximal tubular fluid or recombinant human HGF (rhHGF) or rhTGF-beta and expression of C-C chemokines was measured by RT-PCR and ELISA. Interactions of tubular cell chemokines with macrophages and indirectly with myofibroblasts were also examined using cell culture models. RESULTS: In rats with glomerular proteinuria due to diabetic nephropathy mature, bioactive HGF as well as active and latent TGF-beta were detected in early proximal tubular fluid. Specific HGF- and TGF-beta type II receptors were expressed in apical tubular membranes more in diabetic compared to control rats. Incubation of cultured mouse proximal tubular cells (mPTC) or medullary collecting duct cells (mIMCD-3) with diabetic rat proximal tubular fluid increased MCP-1 and RANTES mRNA levels as well as secreted peptide up to threefold. In contrast, high glucose (450 mg/dL), bovine serum albumin (BSA) or rat albumin (each at 100 micrograms/mL) or 10 nmol/L insulin-like growth factor-I (IGF-I; which was also present in glomerular ultrafiltrate in rats with diabetic nephropathy) did not affect expression of these chemokines. Recombinant human TGF beta as well as rhHGF each increased MCP-1 and RANTES mRNA as well as peptide levels several-fold. In cultured macrophages MCP-1 raised the secretion of TGF beta, which in turn increased the expression of collagen type I and III as well as fibronectin in renal interstitial myofibroblasts about 2.5 to 4-fold. CONCLUSIONS: Proteinuria-induced progressive renal interstitial fibrosis may be caused by glomerular ultrafiltration of high molecular weight bioactive growth factors, HGF and TGF-beta, which "activate" tubular cells through apical membranes. These apical signals are translated into basolateral events that are recognized by cells in the interstitium, such as the basolateral secretion of the C-C-chemokines MCP-1 and RANTES, which may (via macrophages) stimulate interstitial myofibroblasts, and thus lead to accumulation of extracellular matrix proteins and progressive interstitial fibrosis. PMID- 10720955 TI - Suppressive effects of fish oil on mesangial cell proliferation in vitro and in vivo. AB - BACKGROUND: Mesangial cell proliferation is a characteristic feature of IgA nephropathy and many other forms of glomerulonephritis. Recent clinical studies have shown that dietary fish oil supplementation retards renal disease progression in patients with IgA nephropathy. The mechanism by which this effect occurs is unknown. METHODS: The anti-Thy 1.1 (ATS) model of mesangial proliferative glomerulonephritis was employed to test the hypothesis that dietary fish oil supplementation reduces mesangial cell proliferation following acute injury. Subcultured rat mesangial cells were used to determine the in vitro effects of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), the primary components of fish oil, on proliferation. RESULTS: Following antithymocyte serum (ATS) administration, proteinuria was significantly decreased in animals treated with fish oil compared with sesame oil-treated controls. In ATS rats given fish oil, there was less mesangial cell and matrix expansion, mesangiolysis, or basement membrane disruption (delta% = -40%). ATS rats receiving fish oil had less glomerular cell proliferation (PCNA-delta% = -50%) and a reduction of alpha-smooth muscle actin expression (delta% = -27%) by mesangial cells. In subcultured rat mesangial cells, DHA, but not EPA, significantly inhibited proliferation. CONCLUSIONS: Fish oil inhibits mesangial cell activation and proliferation in ATS glomerulonephritis, reduces proteinuria, and decreases histologic evidence of glomerular damage. In vitro, the antiproliferative effects of fish oil are more likely related to the action of DHA. We suggest that orally administered fish oil, or purified DHA, may have a suppressive effect in acute phases or relapses of glomerulopathies by inhibiting activation and proliferation of mesangial cells. PMID- 10720956 TI - Eicosapentaenoic acid suppresses PDGF-induced DNA synthesis in rat mesangial cells: involvement of thromboxane A2. AB - BACKGROUND: The administration of eicosapentaenoic acid (EPA) derived from marine oils has been shown to suppress vascular myocyte, lymphocyte, keratinocyte, and mesangial cell proliferation in vitro, although the effects are variable and most reports provide fragmented insight into the mechanism(s) responsible for altering cell growth, particularly the relationship of membrane lipid remodeling to changes in cell proliferation. Thus, these studies were designed to elucidate the effects of mesangial cell membrane fatty acid remodeling (induced by EPA) on cell growth, and to define the relevance of changes in the synthesis of growth modulating eicosanoids. METHODS: Mesangial cells were grown in RPMI and 17% fetal calf serum, and were subcultured and grown for 48 hours in 17% delipidated serum or delipidated serum supplemented with 0 to 50 micrograms/mL of EPA. Quiescent EPA-loaded and control mesangial cells were subjected to stimulation with 20 ng/mL of platelet-derived growth factor (PDGF) followed by measurement of 3H thymidine incorporation and cell number. RESULTS: Mesangial cells remodeled with EPA exhibited a significant decrease in PDGF-stimulated 3H-thymidine incorporation and cell number associated with a reduction in thromboxane A2 (TXA2) in the media. Importantly, the phospholipid fatty acid composition of mesangial cells grown in media enriched with EPA revealed an increase in EPA (0.5 +/- 0.02% to 17.02 +/- 0.52%) coupled with a reciprocal decrease in the precursor for TXA2, arachidonic acid (18.9 +/- 3.17% to 3.55 +/- 0.30%). Blockade of TXA2 synthesis in mesangial cells treated with indomethacin (0.1 to 100 mumol/L) or the specific TXA2 synthase inhibitor, U-63557A (0.1 to 100 mumol/L), evoked a similar reduction in PDGF-stimulated proliferation and TXA2 synthesis. Coincubation of PDGF with the TXA2 mimetic, U-46619 (1 mumol/L), reversed the growth suppression induced by cell membrane remodeling. CONCLUSIONS: These studies suggest that changes in membrane fatty acid composition induced by EPA modulates PDGF-stimulated proliferation by engendering a change in PDGF stimulated TXA2 synthesis. Furthermore, we conclude that TXA2 functions as a comitogen for PDGF-stimulated mesangial cell growth. PMID- 10720957 TI - Complement-induced phospholipase A2 activation in experimental membranous nephropathy. AB - BACKGROUND: In the passive Heymann nephritis (PHN) model of membranous nephropathy, C5b-9 induces glomerular epithelial cell (GEC) injury and proteinuria, which is partially mediated by eicosanoids. By analogy, in cultured rat GEC, sublytic C5b-9 injures plasma membranes and releases arachidonic acid (AA) and eicosanoids, due to activation of phospholipase A2 (PLA2). This study addresses the mechanisms of PLA2 activation. METHODS: PLA2 expression was assessed with the polymerase chain reaction or immunoblotting, and activity was determined using an in vitro assay or by measurement of free AA. RESULTS: Under basal conditions, GEC in culture expressed a relatively low level of cytosolic PLA2 (cPLA2) protein, while mRNAs of groups IB, IIA and V secretory PLA2s (sPLA2) were not detectable. Incubation of GEC with sublytic C5b-9 induced 1.5- to 2.0 fold increases in free [3H]AA at 40 minutes, and three and 24 hours. C5b-9 did not increase cPLA2 protein, and did not induce group IB, IIA or V sPLA2 mRNAs. Stable overexpression of cPLA2 in GEC amplified the C5b-9-induced increases in free [3H]AA, while analogous overexpression of group IIA sPLA2 had no effect. PLA2 activity was increased in glomeruli of rats with PHN, and this enhanced activity was characterized as cPLA2. There were no differences in cPLA2 protein expression between PHN and control glomeruli. CONCLUSIONS: Release of AA by C5b-9 in GEC in culture and in vivo is mediated by cPLA2, and the mechanism is consistent with post-translational regulation of cPLA2 activity. C5b-9 does not induce expression or stimulate activity of sPLA2 isoforms in GEC. PMID- 10720958 TI - Glucocorticoids enhance the expression of the basolateral Na+:HCO3- cotransporter in renal proximal tubules. AB - BACKGROUND: Studies have shown that glucocorticoids enhance HCO3- reabsorption in proximal tubules. Functional and molecular studies indicate that HCO3- reabsorption in proximal tubules is mediated via luminal H(+)-ATPase and Na+/H+ exchanger (NHE-3), and basolateral Na+:HCO3- cotransporter (NBC) acting in series. The purpose of these experiments was to examine the effect of adrenal steroids on NBC-1 and NHE-3 expression and activity in rat renal proximal tubules. METHODS: Rats were injected subcutaneously with dexamethasone (100 mu/day) or deoxycorticosterone acetate (30 mg/kg), potent glucocorticoid, or mineralocorticoid analogues, respectively. Animals were sacrificed after two or four days, and NBC-1 and NHE-3 mRNA expression and activities were measured in cortex and proximal tubules. RESULTS: Northern hybridizations indicated that cortical NBC-1 mRNA expression increased by approximately 92% in rats treated with dexamethasone for four days (N = 6, P < 0.03) but not two days. NHE-3 mRNA expression remained unchanged. NBC and NHE-3 activities were measured as the Na dependent pHi recovery from an acid load in the presence or absence of HCO3-, respectively, and appropriate inhibitors in proximal tubule suspensions loaded with BCECF. NBC activity increased by approximately 80% in rats treated with dexamethasone for four days (P < 0.01, N = 5) but not two days. NHE-3 activity increased by 34 and 42% in rats treated with dexamethasone for two and four days, respectively (P < 0.05 and P < 0.02 for each group vs. control). Treatment with deoxycorticosterone acetate did not alter NBC-1 expression. CONCLUSION: Glucocorticoids at pharmacologic concentrations enhance the mRNA expression and functional activity of renal proximal tubule NBC-1. Enhanced NBC and NHE-3 activities could result in increased HCO3- reabsorption in proximal tubule and could contribute to the maintenance of metabolic alkalosis in pathophysiologic states associated with increased glucocorticoid production. PMID- 10720959 TI - Sialic acid and crystal binding. AB - BACKGROUND: We studied the role of cell surface sialic acid in the adherence of calcium oxalate monohydrate (COM) crystals to Madin-Darby canine kidney (MDCK) cells. METHODS: Studies were performed with undifferentiated (crystal-binding) cells in subconfluent cultures and maturated (noncrystal-binding) cells in confluent cultures. Lectins were used to study the emergence and abundance of oligosaccharides at the cell surface during epithelial development. The effect of neuraminidase treatment on crystal binding was studied with [14C]COM crystals, and the enzyme-induced release of cell surface-associated sialic acid molecules was monitored by labeling the cells metabolically with [3H]glucosamine. RESULTS: Binding studies with lectins derived from Maackia Amurensis II (MALII) and Sambucus Nigra (SNA) demonstrated that the cells expressed terminal sialic acids attached to penultimate galactose through alpha 2,3 and alpha 2,6 bonds at different stages of epithelial development. Neuraminidase treatment strongly reduced the affinity of the cell surface for COM crystals in subconfluent cultures. Nevertheless, neuraminidase cleaved more sialic acids from cells in confluent cultures than from those in subconfluent cultures. Peanut agglutinin (PNA), which binds only to sialylated terminal galactose units, adhered to developing but not to maturated cells, unless the latter were pretreated with neuraminidase. Both results indicate that the surface of maturated MDCK cells is more heavily sialylated than that of undifferentiated cells. Free sialic acid molecules showed little or no affinity for COM crystals and did not affect the adherence of the crystals to undifferentiated cells. CONCLUSIONS: There are at least two models that may explain these results. First, sialic acids are presented at the surface of immature cells in an orientation that specifically matches crystal surface characteristics favoring crystal-cell interactions. Second, sialic acid molecules are not directly associated with the crystals, but may be involved in the exposure of another crystal binding molecule at the cell surface. PMID- 10720960 TI - Synergistic effect of alpha-adducin and ACE genes causes blood pressure changes with body sodium and volume expansion. AB - BACKGROUND: The genetic dissection of a polygenic, multifactorial, quantitative disease such as arterial hypertension is hampered by a large environmental variance and by genetic heterogeneity. METHODS: To reduce the environmental variance, we measured the pressor response to a saline load (PRSL) and the basal plasma renin activity (PRA) under very controlled conditions in 145 essential hypertensive patients, as they may have the most direct clinical expression of the putative genetic alteration in renal Na handling and blood pressure (BP) regulation caused by the alpha-adducin and angiotensin-converting enzyme (ACE) polymorphism. RESULTS: PRSL was smaller in patients homozygous for the wild-type (Gly460) variant of alpha-adducin compared with that of patients bearing at least one copy of the 460Trp variant (2.5 +/- 0.6 vs. 7.0 +/- 0.9 mm Hg, P = 0.0001), whereas the ACE genotype was not associated with differences in PRSL. Both alpha adducin and ACE affect PRA, with lower values correlated with the number of 460Trp or D alleles (P = 0.019 and 0.017, respectively). Most important, alpha adducin and ACE interact epistatically in determining the PRSL, doubling the variance explained when epistasis is taken into account (variance from 7.7 to 15.5%). CONCLUSION: These findings support the involvement of ACE and alpha adducin in PRSL and PRA control, which are of paramount importance in setting the BP level and its response to therapy. PMID- 10720961 TI - Postischemic vasodilation, endothelial activation, and cardiovascular remodeling in end-stage renal disease. AB - BACKGROUND: Cardiovascular complications are the major cause of death in end stage renal disease (ESRD) patients. These complications are associated with concomitant cardiac and vascular remodeling, including left ventricular (LV) hypertrophy and hypertrophy of arterial walls. The endothelium influences the process of arterial remodeling. ESRD patients are characterized by the development of both cardiovascular remodeling and endothelial dysfunction. METHODS: Common carotid artery (CCA) intima-media thickness (IMT), CCA diameter, CCA distensibility, LV mass, and function were determined in 60 stable ESRD patients on hemodialysis and 34 age-, sex-, and blood pressure (BP)-matched controls, and their relationships with endothelial alterations were estimated by forearm postischemic vasodilation [flow debt repayment (FDR)] measured by venous plethysmography. We also evaluated the relationships between FDR and several cardiovascular risk factors or markers of inflammatory response or endothelial activation, for example, duration of dialysis, BP, smoking habits, cholesterol, parathormone (PTH), serum albumin, plasma fibrinogen, C-reactive protein (CRP), plasma homocysteine, plasminogen activator inhibitor (PAI-1), and von Willebrand factor (vWF). RESULTS: ESRD patients had increased LV mass, CCA diameter and CCA IMT, and had decreased CCA distensibility (P < 0.05). While the postischemic peak flow was comparable in controls and ESRD patients (29.2 +/- 9.1 vs. 27.9 +/- 0.2 mL/100 mL/min), FDR was lower in ESRD patients (116 +/- 31 vs. 88 +/- 32%, P < 0.001) because of the shorter duration of vasodilation (127 +/- 36 vs. 96 +/- 32 s, P < 0.001). The time to complete FDR was longer in ESRD patients (110 +/- 54 vs. 162 +/- 72 s, P < 0.001). ESRD patients had lower high-density lipoprotein cholesterol and serum albumin (P < 0.01) and higher triglycerides, fibrinogen, plasma homocysteine, vWF (P < 0.01), and PAI-1 (P < 0.05). For ESRD patients, significant negative age- and pressure-independent correlations were established between FDR and CCA diameter, duration of dialysis, and PAI-1. FDR was positively correlated with serum albumin. FDR and time to FDR were negatively correlated with CCA IMT and LV mass. CCA distensibility was positively associated with FDR (P < 0.001) and negatively with time to FDR (P < 0.001). The PAI-1 concentration was positively correlated with CCA IMT (P < 0.01) and negatively with CCA distensibility (P < 0.001). CONCLUSIONS: Our data provide the first evidence that cardiac and arterial remodeling in ESRD patients are inversely related to forearm reactive hyperemia. The diminished hyperemic response is due to the shorter duration of hyperemia and is associated with higher concentrations of serum markers of endothelial activation, suggesting that, in ESRD patients, endothelial dysfunction may be a factor influencing cardiovascular changes. PMID- 10720962 TI - Endothelial dysfunction in renal transplant recipients maintained on cyclosporine. AB - BACKGROUND: Hypertension is almost universal following renal transplantation and may contribute to the already poor cardiovascular prognosis of this group. Cyclosporine-induced hypertension is a particular problem and has variously been attributed to increased sympathetic nerve activity, salt and water retention, and increased circulating endothelin levels. However, the effects of cyclosporine on the L-arginine/nitric oxide (NO) system in vivo in humans are unknown. In this present study, we examined basal and stimulated NO production from the vascular endothelium in cyclosporine-treated renal transplant recipients using the technique of forearm venous plethysmography. METHODS: In study 1, stimulated NO production was assessed in 9 cyclosporine-treated renal transplant recipients (CsA), 7 azathioprine-treated renal transplant recipients (AZA), and 12 controls, using carbachol (an endothelium-dependent vasodilator) and sodium nitroprusside (an endothelium-independent vasodilator). In study 2, basal NO production was assessed in 9 cyclosporine-treated patients and 11 controls using L-NMMA (inhibits NO synthase), with norepinephrine as a control vasoconstrictor. Drugs were infused into the nondominant forearm through a sterile 27-gauge needle, and changes in forearm blood flow (FBF) were measured using venous occlusion plethysmography. RESULTS: In study 1, sodium nitroprusside caused a similar dose dependent increase in FBF in all groups. However, the median (range) percentage increase FBF to carbachol (3 micrograms/min) was markedly reduced in the CsA patients (188.8; 72.5 to 385.1) compared with AZA patients (378.1; 124.0 to 548.9; P = 0.042) and to controls (303.8; 124.8 to 813.3; P = 0.028). In study 2, the maximum percentage reduction in FBF to L-NMMA (4 mumol/min) was less pronounced in CsA patients (-19.5; -4.7 to -63.1) compared with controls (-39.5; 15.7 to -52.8; P = 0.056), and while controls vasoconstricted to the maximum dose of norepinephrine (240 pmol/min) as expected (-26.9; -1.4 to -38.6), CsA patients as a group tended to vasodilate (7.9; -36.8 to 92.6; P = 0.02). CONCLUSION: These data demonstrate impaired stimulated and basal NO production in CsA patients, indicating endothelial dysfunction. This may predispose patients to atherosclerosis and may be involved in the etiology of post-transplant hypertension. PMID- 10720963 TI - Increased bone strontium levels in hemodialysis patients with osteomalacia. AB - BACKGROUND: In this study, we report on the association between increased bone strontium levels and the presence of osteomalacia in end-stage renal failure patients treated by hemodialysis. METHODS: We performed a histologic examination and determined the strontium content and strontium/calcium ratios in bone biopsies of 100 hemodialysis patients recruited from various centers all over the world. Aside from the bone strontium concentration, the bone aluminum content was assessed. The bone zinc concentration, a nonrelevant element for bone toxicity, was also measured. RESULTS: Bone strontium levels and bone strontium/calcium ratios were increased in subjects with osteomalacia when compared with those with the other types of renal osteodystrophy. Bone strontium and bone calcium levels correlated with each other. The slope of the linear regression curve correlating these parameters was much steeper in the osteomalacic group (Y = 2.22X - 120) as compared with the other types of renal osteodystrophy (Y = 0.52X - 5.7). Within the group of patients with osteomalacia, bone strontium levels also significantly correlated with the bone aluminum content (r = 0.72, P = 0.018). No such correlation was found for the other types of renal osteodystrophy. The bone zinc concentration of subjects with normal renal function did not differ significantly from the values noted for the various types of renal osteodystrophy taken as separate groups, nor could increased bone zinc concentrations be associated with a particular bone lesion. CONCLUSIONS: Our data demonstrate an association between osteomalacia and increased bone strontium concentrations in dialysis patients. Further studies are warranted to establish whether strontium plays either a primary, secondary, or contributive role in the development of the latter type of renal osteodystrophy. PMID- 10720964 TI - Effects of low animal protein or high-fiber diets on urine composition in calcium nephrolithiasis. AB - BACKGROUND: The purpose of this article is to evaluate the impact of low protein and high fiber intakes on risk factors of stone recurrence in idiopathic calcium stone formers (ICSFs). METHODS: Ninety-six ICSFs were randomly assigned a low animal protein diet (< 10% of total energy), a high-fiber diet (> 25 g/day), or a usual diet (control group); all patients were recommended to increase their fluid intake. Their daily urine compositions were analyzed at baseline and at four months. Compliance with dietary recommendations was checked by validated food frequency questionnaires. Compliance with total and animal protein intakes was assessed by 24-hour urea and sulfate outputs, respectively. The nutritional intervention (oral instructions, written leaflet, phoning) and food assessment were carried out by a research dietitian. RESULTS: At baseline, diets and the daily urine composition did not differ between the three groups. At four months, while diets differed significantly, the 24-hour output of calcium and oxalate did not differ significantly within and between groups after adjustment for potential confounders (age, sex, and personal and family history of calcium stones) and baseline values. However, as many as 12 out of 31 ICSFs (95% CI, 22 to 58%) assigned to a low animal protein diet achieved a reduction in the urine urea excretion rate of more than 50 mmol/day and also exhibited a significant decrease in urinary calcium excretion that averaged 1.8 mmol/day. A significant correlation between urea and calcium outputs was observed only among patients with hypercalciuria. CONCLUSIONS: These results show that only ICSFs who markedly decrease their animal protein intake, especially those with hypercalciuria, can expect to benefit from dietary recommendations. PMID- 10720965 TI - Treatment of failed native arteriovenous fistulae for hemodialysis by interventional radiology. AB - BACKGROUND: We studied the feasibility, technical problems, safety, and effectiveness of percutaneous declotting of thrombosed native arteriovenous fistulae for hemodialysis. METHODS: Between 1992 and 1998, 93 declotting procedures were performed in 73 consecutive upper limb native fistulae (forearm 56 and upper arm 17), and 162 procedures were performed in 78 prosthetic grafts using manual catheter-directed thrombo-aspiration, with or without previous urokinase infusion. Detection of restenosis by clinical surveillance led to redilation or stent placement. Rethrombosis in four forearm and six upper arm fistulae were treated by 20 further declottings by aspiration. RESULTS: The initial success was 93% in the forearm and 76% in the upper arm (99% in grafts). The complications included one pulmonary embolism, one acute pseudoaneurysm, and one blood depletion requiring transfusion. Primary patency rates at one year were 49% in the forearm and 9% in the upper arm (14% in grafts). Secondary patency rates were 81 and 50% at one year, respectively (83% in grafts). Reinterventions were necessary every 19.6 months in the forearm and every 5.7 months in the upper arm (every 6.4 months in grafts, P < 0.05). Stents were placed in 11% of forearm fistulae and in 41% of upper arm fistulae (45% of grafts) for treatment of acute rupture (5 out of 19), stenosis recoil (6 out of 19), and early (< 6 months) recurring stenosis (8 out of 19). CONCLUSIONS: The percutaneous declotting of forearm fistulae by manual catheter-directed thrombo-aspiration was effective in more than 90% of cases and yielded 50% primary and 80% secondary patency rates at one year. The results were poorer in upper arm fistulae. The need for maintenance reinterventions was three times smaller in forearm fistulae than in upper arm fistulae and grafts. PMID- 10720966 TI - Interdialytic weight gain and survival in hemodialysis patients: effects of duration of ESRD and diabetes mellitus. AB - BACKGROUND: Medical mortality determinants in end-stage renal disease (ESRD) patients treated with hemodialysis (HD) are well known. More recently, associations have been established between the dose of dialysis administered and patient survival. We showed in a prospective study that both dialyzer type and patient compliance with the dialysis prescription were independently associated with survival. Although several parameters of dialytic technique and patient compliance are associated with differential survival in patients with ESRD treated with HD, the association of interdialytic weight gain (IWG) with survival is unclear. No study has assessed the relationship between IWG and mortality in HD patients, controlled for multiple medical risk factors. The aim of our study was to determine whether IWG was associated with survival in patients with ESRD treated with HD, controlling for multiple medical and dialytic risk factors. METHODS: We prospectively conducted an observational, longitudinal, multicenter study of 283 urban HD patients to determine the relationship of IWG with several dialytic parameters and patient survival. Medical risk factors such as demographic indices and comorbid conditions were assessed. We studied Kt/V, the protein catabolic rate (PCR), serum albumin and anthropometric measurements, behavioral compliance indices, dialyzer characteristics, and serum electrolyte concentrations, and correlated these with IWG. In addition, the duration of dialysis was assessed in HD patients with and without diabetes mellitus. Cox proportional hazards models assessed the relative mortality risk of increased IWG, controlling for variations in medical comorbidity and other mortality determinants. RESULTS: The mean (+/- SD) age of our population was 54.6 +/- 14.1 years, and the mean time they were treated with HD was 30.4 +/- 46.9 months. The mean IWG was 1.54 +/- 0.71% dry wt/day. Correlations were found between increased IWG and younger age, and lower midarm circumference, and increased Kt/V, PCR, and serum potassium concentration. The mean follow-up period was 48.9 +/- 10.6 months. An increase in IWG was associated with a significantly increased relative mortality risk in diabetic ESRD patients treated with HD when variations in age, comorbidity, serum albumin concentration, and dialyzer type and site were controlled. There was, however, no association of increased mortality risk with increased IWG in the larger population of patients without diabetes. In further analyses, the increased mortality risk associated with increased IWG was found to be present only in patients with diabetes mellitus who had recently started HD therapy for ESRD. CONCLUSION: IWG is correlated with several nutritional and dialytic variables and with parameters that predict survival in HD patients. Increased IWG is independently associated with decreased survival of diabetic ESRD patients treated with HD, after adjusting for variation in other medical risk factors. The population of incident diabetic HD patients is particularly susceptible to increased risk associated with increased IWG. The mechanisms underlying these results are obscure, but IWG might be associated with poorer survival in this population if it were linked to worsened hypertension, cardiovascular stress, or poorer glycemic control. Interventions to improve compliance with IWG in incident diabetic HD patients are warranted. PMID- 10720967 TI - Accuracy of hemodialysis modeling. AB - BACKGROUND: One- and two-compartmental models of hemodialysis (HD) are well known. These models make it possible to analyze the course of treatment and to predict the effect of dialysis procedures. Mathematical modeling helps physicians to match dialysis therapy to the individual needs of the patient; however, the efficiency of the models depends on the accuracy of the coefficients. How to select coefficients in the case of one-compartmental models is known for urea and creatinine. Less information is available for two-compartmental models. Results on modeling of uric acid concentrations have not been published. METHODS: The identification of the mathematical model coefficients was based on the concentration measurements of three markers of uremic toxicity (urea, creatinine, and uric acid) in both patients' blood and dialysate. Blood samples were taken from the arterial line several times throughout the dialysis period. Simultaneously, dialysate samples were taken from a test port in the dialyzer outflow line. The mathematical model parameters were determined so as to minimize the deviations between the measured points and the calculated curves. In this way, distribution volumes, cellular clearances, and dialyzer mass transfer coefficients were estimated. RESULTS: For a one-compartmental model, the median value of distribution volume V = 0.56 DW was obtained, where DW is the patient's dry weight. For a two-compartmental model, intercompartment volume Vi = 0.36 DW and extracompartment volume Ve = 0.21 DW. The following median values for cellular clearances were established: urea 415 (mL/min), creatinine 207 (mL/min), and uric acid 257 (mL/min). CONCLUSIONS: One- and two-compartmental models describe the concentration of the urea, creatinine, and uric acid very effectively, in contrast with phosphorus, in which modeling results are not satisfactory. Although two-compartmental models are more effective, they are much more complicated than one-compartmental models, which justifies using the one compartmental model for hemodialysis modeling. A two-compartmental model must be used in the case of rebound phenomenon modeling. The total body water values we have obtained are similar to the anthropometrically based values for urea and creatinine and to a lesser degree for uric acid. Distribution volumes for one- and two-compartmental models obtained from patient weight are the simplest coefficients for mathematical models and have sufficient precision as well. The global value of both compartments is slightly greater than the corresponding value for a one-compartmental model. The effectiveness of dialyzers is in practice lower than might be expected on the basis of the data provided by their manufacturers. Urea cellular clearance is two times greater than creatinine and uric acid cellular clearances. The clearance differences are more prominent for the cellular membrane than for artificial semipermeable membranes. PMID- 10720968 TI - Pharmacokinetics of mycophenolate mofetil in patients with end-stage renal failure. AB - BACKGROUND: Mycophenolate mofetil (MMF) acts as a prodrug for the immunosuppressive drug mycophenolic acid (MPA). It is rapidly converted to MPA following oral ingestion. MPA is metabolized to MPA glucuronide (MPAG), which is renally excreted. This study examines the pharmacokinetics of MPA and MPAG in patients with end-stage renal failure who were on hemodialysis (N = 10) or peritoneal dialysis (N = 10) treatment. METHODS: After an overnight fast, a single oral dose of 1 g MMF was given. Plasma concentrations of MPA and MPAG were measured from 0 (predose) to 36 hours after administration, using high performance liquid chromatography (HPLC). The area under the concentration time curve (AUC) from 0 to 36 hours was calculated using the trapezoidal rule. RESULTS: Mean (+/- SD) AUC for MPA was 55.7 +/- 32.6 mg/L.h for hemodialysis patients and 44.7 +/- 14.7 mg/L.h for peritoneal dialysis patients, which is similar to expected values for subjects with normal renal function. The mean (+/- SD) maximum plasma concentration (Cmax) for MPA was lower than would be expected for subjects with normal renal function (16.01 +/- 10.61 mg/L for hemodialysis, 11.48 +/- 4.98 mg/L for peritoneal dialysis). MPAG clearance was prolonged with AUC approximately five times what would be expected in subjects with normal renal function (1565 +/- 596 mg/L.h for hemodialysis, 1386 +/- 410 mg/L.h for peritoneal dialysis). There was no significant difference for any of the pharmacokinetic parameters between subjects on hemodialysis and those on peritoneal dialysis. Plasma concentrations of MPA and MPAG did not fall significantly during hemodialysis. No MPA was detectable in hemodialysis or peritoneal dialysis fluid, but small amounts of MPAG were detected in hemodialysis fluid in 1 out of 10 subjects and in peritoneal dialysis fluid in 3 out of 10 subjects. CONCLUSIONS: The accumulation of MPAG may be responsible for the poor gastrointestinal tolerance of this drug in dialysis patients and probably limits the maximum dose of MMF that can be tolerated. PMID- 10720969 TI - Percutaneous treatment of thrombosed primary arteriovenous hemodialysis access fistulae. AB - BACKGROUND: We reviewed the efficacy of percutaneous intervention in acute thrombotic occlusion of native arteriovenous (AV) fistulae for hemodialysis. METHODS: Eight-one percutaneous procedures were performed in 54 patients presenting with a clotted native dialysis fistula. There were 60 cases of a long segment thrombosis of the fistula. In 20 cases, a small thrombus usually caused by an underlying severe stenosis was observed. A proximal arterial occlusion was seen in one case. Treatment depended on clot size and included balloon dilation (N = 20), mechanical thrombectomy with various devices (N = 58), as well as pharmacomechanical thrombolysis (N = 3). RESULTS: Full restoration of flow was established in 72 cases (88.9%). Early reobstruction within 14 days occurred in eight cases (11.1%). Primary patency rates after a 1-, 3-, 6-, and 12-month period were 74, 63, 52, and 27%, respectively. Overall fistula patency was 75% after 3 months, 65% after 6 months, 51% after 12 months, and 22% after 24 months. CONCLUSIONS: Acute thrombotic occlusion of native AV fistulae is a major complication of hemodialysis. The results of treatment are believed to be less successful than thrombosis treatment in synthetic grafts. Our results, however, indicate the efficacy of percutaneous treatment in native fistulae, and demonstrate comparable technical results and patency rates. PMID- 10720970 TI - Vintage, nutritional status, and survival in hemodialysis patients. AB - BACKGROUND: The link between dialysis "vintage" (length of time on dialysis in months to years) and survival has been difficult to define, largely because of selection effects. End-stage renal disease (ESRD) is thought to be a wasting illness, but there are no published reports describing the associations between vintage and body composition in hemodialysis patients. METHODS: We explored the relationships among vintage, nutritional status, and survival in a 3009 patient cohort of prevalent hemodialysis patients. Body weight, total body water, body cell mass, and phase angle by bioelectrical impedance analysis were the body composition parameters of interest. We examined vintage as an explanatory variable in multiple linear regression analyses (adjusted for age, gender, race, and diabetes) using body composition parameters and biochemical indicators of nutritional status as dependent variables. Proportional hazards regression was used to evaluate the association of vintage and survival with and without adjustment for case mix and laboratory variables. RESULTS: Dialysis vintage was 3.8 +/- 3.7 (median 2.6) years. Body composition parameters tended to be lower after dialysis year 2. Linear estimates per year of vintage beyond year 2 include -0.66 kg body wt (P < 0.0001), -0.17 kg total body water (P = 0.0003), -0.14 kg body cell mass (P < 0.0001), and -0.07 degrees phase angle (P < 0.0001). In unadjusted analyses, vintage was not associated with survival, either as a linear or higher order term. The adjustment for case mix yielded a vintage term associated with an increased relative risk (RR) of death (RR 1.04 (95% CI, 1.01 to 1.07 per year). A further adjustment for laboratory data yielded a RR of 1.06 (95% CI, 1.03 to 1.09 per year). CONCLUSION: Dialysis vintage is related to nutritional status in hemodialysis patients, with vintage of more than years associated with a significant decline in all measured nutritional parameters. Cross-sectional analyses probably underestimate these effects. A year accrued on dialysis is associated with a 6% increase in the risk of death, all else equal. Longitudinal assessments of nutritional status, including body composition, are required to better understand the natural history of wasting with ESRD and its implications for long-term survival. PMID- 10720971 TI - Dialysis solution containing hyaluronan: effect on peritoneal permeability and inflammation in rats. AB - BACKGROUND: Hyaluronan (HA), a high molecular weight mucopolysaccharide found in interstitial tissues and fluid, is lost from the peritoneal cavity during peritoneal dialysis. In order to determine the role of HA in peritoneal function, we investigated the effects of exogenous HA on peritoneal permeability, markers of intraperitoneal inflammation, and peritoneal morphology in rats exposed to peritoneal dialysis solution for four weeks. METHODS: Wistar rats were infused intraperitoneally, twice daily, with conventional, hypertonic dialysis solution (Dianeal 3.86%; control) or Dianeal solution containing 10 mg/dL of high molecular weight HA. Peritoneal permeabilities and clearances of solutes and protein were determined using a modified peritoneal permeability test (peritoneal equilibration test) at the beginning and the end of the treatment. Peritoneal volume and ultrafiltration were determined using a macromolecular marker and by gravimetric methods. Peritoneal inflammation was determined by cell counts and differential and by the measurement of cytokine concentrations in the dialysate effluent. Peritoneal thickness and HA content were determined in liver and mesentery biopsies taken at the end of the experiment. RESULTS: After four weeks of exposure to the dialysis solution, transperitoneal protein equilibration was significantly lower in HA-treated rats compared with rats treated with Dianeal alone (46% lower for albumin, P < 0.003; 33% lower for total protein, P < 0.001). The total drained volume after a four hour dwell was 29% higher in the HA group compared with the control (P < 0.001), yielding a positive net ultrafiltration in the HA group versus a negative net ultrafiltration in controls. Peritoneal clearances of urea and creatinine tended to be elevated in HA-treated rats, while clearances of total protein and albumin tended to be lower. Dialysate effluent from rats exposed to HA contained a lower percentage of neutrophils (8.8 +/- 22.8 +/- 9.5%, P < 0.01) and lower levels of the cytokines, tumor necrosis factor alpha (11.2 +/- 14.7 vs. 42.3 +/- 35.3 pg/mL, P < 0.05) and monocyte chemoattractant protein-1 MCP-1 (72.0 +/- 86.5 vs. 402.4 +/- 258.3 pg/mL, P < 0.02), compared with rats treated with Dianeal alone. The thickness of the peritoneal interstitium showed a similar increase in both groups, but mesenteric tissue from the HA group contained more HA (48%, P < 0.01) than tissue from control animals. CONCLUSIONS: The addition of HA to peritoneal dialysis solution decreases protein permeability, increases ultrafiltration, and decreases cytokine levels and the proportion of peritoneal neutrophils in dialysate from rats exposed to hypertonic dialysis solution. These results suggest that exogenous HA may help to protect the peritoneal membrane during exposure to dialysis solutions. These benefits, if sustained in the clinical setting, could lead to improvements in the therapy of peritoneal dialysis. PMID- 10720972 TI - Michelangelo: art, anatomy, and the kidney. AB - Michelangelo (1475-1564) had a life-long interest in anatomy that began with his participation in public dissections in his early teens, when he joined the court of Lorenzo de' Medici and was exposed to its physician-philosopher members. By the age of 18, he began to perform his own dissections. His early anatomic interests were revived later in life when he aspired to publish a book on anatomy for artists and to collaborate in the illustration of a medical anatomy text that was being prepared by the Paduan anatomist Realdo Colombo (1516-1559). His relationship with Colombo likely began when Colombo diagnosed and treated him for nephrolithiasis in 1549. He seems to have developed gouty arthritis in 1555, making the possibility of uric acid stones a distinct probability. Recurrent urinary stones until the end of his life are well documented in his correspondence, and available documents imply that he may have suffered from nephrolithiasis earlier in life. His terminal illness with symptoms of fluid overload suggests that he may have sustained obstructive nephropathy. That this may account for his interest in kidney function is evident in his poetry and drawings. Most impressive in this regard is the mantle of the Creator in his painting of the Separation of Land and Water in the Sistine Ceiling, which is in the shape of a bisected right kidney. His use of the renal outline in a scene representing the separation of solids (Land) from liquid (Water) suggests that Michelangelo was likely familiar with the anatomy and function of the kidney as it was understood at the time. PMID- 10720973 TI - Diabetes, genes, and kidney development. PMID- 10720974 TI - New insights into the pathogenesis of membranous nephropathy. PMID- 10720975 TI - URR, weight, and mortality. PMID- 10720976 TI - Catecholamines and transplantation. PMID- 10720978 TI - Malnutrition in dialysis: malnourishment or uremic inflammatory response? PMID- 10720977 TI - Renal handling of albumin in normal rat. PMID- 10720979 TI - Vulvar disease in children: a clinical audit of 130 cases. AB - We evaluated 130 prepubertal girls presenting with a vulvar complaint to determine the spectrum and frequency of conditions seen in this age group. Of the patients, 41 (33%) had atopic or irritant dermatitis, 23 (18%) had lichen sclerosus, 21 (17%) had psoriasis, 15 (12%) had vulvar lesions, most often hemangiomas and nevi, and 13 (10%) had streptococcal vulvovaginitis. Diagnoses less frequently seen were staphylococcal folliculitis (four patients), labial fusion (three patients), genital warts (two patients), molluscum contagiosum of the vulva only (one patient), vulvar bullous pemphigoid (two patients), scabies nodules (one patient), erythema annulare centrifugum (one patient), tinea (two patients), and vitiligo (one patient). We also encountered vulvar presentations of systemic diseases (varicella, staphylococcal scalded skin syndrome, and Henoch Schonlein purpura, all one patient each). We did not see candidal vulvovaginitis in this age group nor did we encounter bacterial infection with pathogens other than Staphylococcus aureus and S. pyogenes. PMID- 10720980 TI - Nail matrix arrest following hand-foot-mouth disease: a report of five children. AB - Hand-foot-mouth disease (HFMD) is a contagious enteroviral infection occurring primarily in children and characterized by a vesicular palmoplantar eruption and erosive stomatitis. Nail matrix arrest has been associated with a variety of drug exposures and systemic illnesses, including infections, and may result in a variety of changes, including transverse ridging (Beau's lines) and nail shedding (onychomadesis). The association of HFMD with Beau's lines and onychomadesis has not been reported previously. Five children, ages 22 months-4 years, presented with Beau's lines and/or onychomadesis following physician-diagnosed HFMD by 3-8 weeks. Three of the five patients experienced fever with HFMD, and none had a history of nail trauma, periungual dermatitis, periungual vesicular lesions, or a significant medication intake history. All patients experienced HFMD within 4 weeks of one another, and all resided in the suburbs of the Chicago metropolitan area. In all patients the nail changes were temporary with spontaneous normal regrowth. The mechanism of the nail matrix arrest is unclear, but the timing and geographic clustering of the patients suggests an epidemic caused by the same viral strain. PMID- 10720981 TI - Elastin gene deletions in Williams syndrome patients result in altered deposition of elastic fibers in skin and a subclinical dermal phenotype. AB - Williams syndrome (WS) is a complex developmental disorder with multisystem involvement known to be the result of a microdeletion in the q11.23 region of chromosome 7. This deletion involves several genes, including the elastin gene. Although elastic fibers are important constituents of skin, little is known about the skin phenotype in WS patients. We have therefore studied the skin of four WS patients in which we've shown the deletion of one copy of the elastin gene. Physical examination and indirect immunofluorescent microscopy of elastin did not detect any major phenotypic or morphologic changes in the skin. We were able, however, to show subtle textural changes in skin and, by electron microscopy, that the amorphous component of elastic fibers in WS patients was consistently reduced when compared to normal controls. These findings indicate that deletion of one copy of the elastin gene results in reduced deposition of elastin in dermal elastic fibers, an altered elastic fiber ultrastructure, and a subclinical dermal phenotype in the children and young adult patients analyzed in this study. PMID- 10720982 TI - Uncombable hair syndrome with angel-shaped phalango-epiphyseal dysplasia. AB - Uncombable hair syndrome or "cheveux incoiffables" is due to a characteristic longitudinal grooving of the hair shaft resulting in a triangular cross section (pili trianguli et canaliculi). In the majority of cases the abnormality is an isolated finding, although uncombable hair-type changes have been observed in conjunction with other features of ectodermal dysplasia. Ultrastructural studies in the latter have revealed more complex changes of the hair shaft, such as longitudinal grooving in combination with torsion, suggesting classification as a different entity. We describe a 6-year-old girl with typical "cheveux incoiffables," as confirmed by scanning electron microscopy, in combination with angel-shaped phalango-epiphyseal dysplasia. The relationship to a previously described syndrome of uncombable hair in combination with retinal dystrophy, juvenile cataract, and brachydactyly, in which both hair and skeletal abnormalities are common, remains to be further elucidated. PMID- 10720983 TI - Idiopathic localized unilateral hyperhidrosis in a child. AB - Idiopathic unilateral circumscribed hyperhidrosis is an extremely rare form of increased sweat production that occurs within a sharply demarcated area on the face or upper extremities of otherwise healthy patients. There are no associated neurovascular or metabolic abnormalities. We report idiopathic localized unilateral hyperhidrosis on the upper extremity of a healthy 4-year-old girl. PMID- 10720984 TI - A pediatric case of dermatofibrosarcoma protuberans: an immunohistochemical study. AB - Dermatofibrosarcoma protuberans (DFSP) is a fibrohistiocytic tumor of intermediate malignancy that usually occurs in early or mid-adult life as a nodular cutaneous mass. DFSP has rarely been reported during childhood or at birth. We report a case of childhood DFSP that illustrates the usefulness of immunohistochemical analysis in the differential diagnosis between DFSP and other fibrohistiocytic proliferations occurring in the skin. A prompt and correct diagnosis is very important in order to ensure appropriate treatment and to prevent local recurrences. PMID- 10720985 TI - Chemical burn caused by topical vinegar application in a newborn infant. AB - Although there is increasing interest in "alternative medicine," including nontraditional and homeopathic remedies, all around the world, they are not always safe and beneficial and may have adverse effects. We report a chemical burn caused by vinegar applied topically to lower body temperature in a febrile newborn and discuss briefly chemical skin burns caused by organic acids. PMID- 10720986 TI - Papules and pustules of the elbows and knees: an uncommon clinical sign of dermatomyositis in oriental children. AB - We report two children with dermatomyositis in whom the initial manifestation was a papular eruption on the extensor surfaces of the elbows and knees. In each there was a follicular component to the eruption and one child had pustular lesions. The extensor eruption predated the onset of muscle weakness by 1 year in the first child and by 2 years in the second. Both children had Vietnamese parents. There is some evidence in the literature that Oriental patients may be predisposed to this type of eruption. PMID- 10720987 TI - Tinea incognito due to Microsporum gypseum in three children. AB - Tinea incognito is a dermatophytosis of atypical clinical character due to the absence of classic features of ringworm. It is caused by prolonged use of topical steroids, sometimes prescribed as a result of incorrect diagnosis. The cases reported in the literature have different clinical presentations and have generally been in adults. We report three children with tinea incognito in whom the lesions were psoriasis-like, eczema-like, and lichenoid, respectively. Diagnosis was confirmed by mycologic examination, which led to the identification of Microsporum gypseum, a geophilic dermatophyte which is an infrequent agent of mycotic infection in humans. PMID- 10720989 TI - Development of diaper rash in the newborn. AB - Diaper rash is a common infant malady. This study documents the earliest stages of rash in a cohort of 31 healthy term newborns over the first 28 days of life. The diaper area was evaluated using a standardized diaper rash grading scale. The anal, buttock, genital, intertriginous, waistband, and leg areas were assessed separately. At birth the average grade was 0.1 and none of the infants had specific features of advanced rash. Nineteen percent had dryness and/or slight redness. By day 7, 71% of infants had some features of skin compromise, giving rise to an overall grade of 0.6. Both the frequency and overall grade increased during postnatal weeks 2 and 3. Overall scores for days 21 and 28 were the same (1.1). The perianal area had the highest overall regional rash grade. Gender differences were present for the genital area only. These findings indicate that epidermal barrier breakdown is an uncommon finding at birth. Clinical signs of irritated skin in the diaper area develop progressively over the first postnatal month. A better understanding of the mechanisms conferring epidermal barrier protection at birth may be important for developing skin care products and practices to extend this protection later into life. PMID- 10720990 TI - Neonatal urticaria due to prostaglandin E1. AB - Prostaglandin E1 is commonly used in the management of cyanotic congenital heart disease. While cutaneous flushing and peripheral edema are well recognized side effects of prostaglandin E1 therapy, other cutaneous effects have not been described in the dermatologic literature. We report a neonate with transposition of the great vessels who developed urticaria during treatment with prostaglandin E1. PMID- 10720991 TI - Congenital midline nasal mass in a toddler. PMID- 10720988 TI - Changes in diapered and nondiapered infant skin over the first month of life. AB - Time- and site-dependent differences in epidermal barrier properties were investigated over the first 28 days of life in healthy term newborn infants. Diapered and nondiapered skin sites were contrasted to the volar forearm of adults (mothers). Thirty-one term infants were evaluated in the hospital on postnatal day 1 and at home on days 4, 7, 14, 21, and 28 for a total of six visits. Measurements included baseline skin hydration, continuous capacitive reactance, peak water sorption, rate of water desorption, skin pH, skin temperature, and environmental conditions. Changes in epidermal barrier properties over the first 4 weeks of life included an increase in surface hydration, a decrease in transepidermal water movement under occlusion, a decrease in surface water desorption rate, and a decrease in surface pH. Diapered and nondiapered regions were indistinguishable at birth but exhibited differential behavior over the first 14 days, with the diapered region showing a higher pH and increased hydration. Maternal measurements remained constant throughout the period. We conclude that healthy newborn skin undergoes progressive changes in epidermal barrier properties over the first 28 days. Adult skin testing does not replicate newborn skin during the first month of life. PMID- 10720992 TI - What syndrome is this? Hidrotic ectodermal dysplasia (Clouston syndrome). PMID- 10720993 TI - Diagnosing tinea versicolor: don't scrape, just tape. PMID- 10720994 TI - Incontinentia pigmenti: an extensive second episode of a "first-stage" vesicobullous eruption. PMID- 10720995 TI - ATOPIC DERMATITIS (AD): EVALUATION AND THERAPY. PMID- 10720996 TI - PHOTOSENSITIVITY DISEASES IN CHILDREN AND ADOLESCENTS. PMID- 10720997 TI - CONTROVERSIES IN PEDIATRIC DERMATOLOGY: CONGENITAL NEVI. PMID- 10720998 TI - CUTANEOUS LYMPHOMAS AND PSEUDOLYMPHOMAS. PMID- 10720999 TI - PORPHYRIA UPDATE. PMID- 10721000 TI - UPDATE: DIAPERS AND DIAPER DERMATITIS. PMID- 10721001 TI - Understanding human behavior is central to improving health. PMID- 10721002 TI - The role of integrins in reproduction. AB - Fertilization, implantation, and placentation are dynamic cellular events that require not only synchrony between the maternal environment and the embryo, but also complex cell-to-cell communication. This communication involves integrins, a large family of proteins involved in the attachment, migration, invasion, and control of cellular function. Over the past decade, investigators have learned that integrins participate in multiple reproductive events including fertilization, implantation, and placentation in many species. This review will describe: (i) the expression of integrins on gametes and during the establishment and development of the placenta; (ii) regulatory pathways for controlling expression of integrins in the uterus and developing placenta; (iii) the function of integrins as determined by null-mutations; and (iv) reproductive dysfunction in women related to inappropriate integrin expression in the uterus and/or placenta. PMID- 10721003 TI - Conditional knockout of mouse insulin-like growth factor-1 gene using the Cre/loxP system. AB - Insulin-like growth factor-1 (IGF-1) is an essential growth factor for normal intrauterine development and postnatal growth. Mice with a complete deficiency of IGF-1 (IGF-1-null mice), created by homologous recombination, were found to exhibit postnatal lethality, growth retardation, infertility, and profound defects in the development of major organ systems. Furthermore, IGF-1-null mice were resistant to growth hormone (GH) treatment in peri-pubertal somatic growth. Using the Cre/loxP-induced conditional knockout system, we generated a mouse that lacks IGF-1 specifically in the liver, the primary site of IGF-1 production. Interestingly, although circulating and serum levels of IGF-1 were decreased by approximately 75% in these mice, they exhibited no defect in growth or development. When administered exogenously, GH stimulated IGF-1 production in several extra-hepatic tissues as well as body growth. The "Somatomedin hypothesis" originally proposed that circulating IGF-1 acting in various tissues mediate the effects of GH. These striking in vivo results, obtained using homologous recombination technology, call for a major modification of the Somatomedin hypothesis. These gene targeting studies confirm that IGF-1 is essential for GH-stimulated postnatal body growth. However, liver-derived (endocrine) IGF-1 is not essential for normal postnatal growth, though it does exert a negative feedback on GH secretion. Instead, local production of IGF-1, acting in a paracrine/autocrine fashion, appears to mediate GH-induced somatic growth. This review will discuss the effects of tissue-specific IGF-1 gene deficiency created by the Cre/loxP system versus the conventional IGF-1 knockout. PMID- 10721004 TI - Anatomy and blood supply of the lower four cranial and cervical nerves: relevance to surgical neck dissection. AB - This study is a continuation of previous work searching for possible anatomic reasons to explain variable and usually unpredictable postoperative pain and dysfunction after the same nerve losses with similar neck dissection operations. The study consisted of dissections of 19 deceased unpreserved elderly subjects arterially injected with dyed latex. Of the 19 subjects, 14 had brain stem and cervical spinal cord dissections, and all had neck dissections. The findings suggested two possible anatomic reasons for the pain and dysfunction: (i) The intracranial anatomy of the lower four cranial nerves, the glossopharyngeal (IX), the vagus (X), the spinal accessory (XI), and the hypoglossal (XII), was just as variable as the previously reported peripheral spinal accessory nerve plexus; and (ii) Both the intracranial and neck dissections indicated that the blood supply to the lower four cranial and cervical nerves, particularly to the brachial plexus, could be impaired by atherosclerosis and/or neuroforaminal impingement or operative loss. This loss of blood supply theoretically could result in ischemia as another possible cause of postoperative pain and dysfunction. It is concluded that because of the potential importance of each nerve and vessel, often unknown at operation, it is very important to spare as many of them as possible to avoid subsequent painful impairment. PMID- 10721005 TI - Leptin receptor transcripts are constitutively expressed in placenta and adipose tissue with advancing baboon pregnancy. AB - The baboon (Papio sp.) is an accepted nonhuman primate model for the study of the endocrinology of human pregnancy. To further characterize this model with regard to leptin function, messenger RNA transcripts for both long (Ob-RL) and short (Ob RS) leptin receptor isoforms were identified in maternal tissues at various stages of gestation. Thus, placental villous, subcutaneous and omental adipose tissues were collected upon cesarean delivery at early (Days 60-62), mid (Days 98 102) and late (Days 159-164) pregnancy (term approximately 184 days). Additionally, amniochorion, decidua, and corpus luteum were collected in late gestation. Expression of Ob-RL and Ob-RS transcripts was determined in relation to constitutively expressed glyceraldehyde-3-phosphate dehydrogenase via reverse transcriptase-polymerase chain reaction, and transcripts were localized within specific placental cell types by in situ hybridization. Ob-RL and Ob-RS transcripts were present in amniochorion, decidua, and corpus luteum at term and appeared constitutively expressed throughout gestation in placenta and adipose tissues. Ob-RS was expressed in greater (P < 0.02) abundance than Ob-RL in all tissues. Within the placenta, receptor isoforms were localized predominantly to the syncytiotrophoblast. The expression of leptin receptor transcripts in maternal adipose tissues, as well as in the syncytiotrophoblast, amniochorion, decidua, and corpus luteum, suggests the potential for autocrine/paracrine roles for the polypeptide in the endocrinology of primate pregnancy. These are the first such observations in a nonhuman primate and support the use of the baboon as a model for the study of leptin in human pregnancy. PMID- 10721006 TI - Association between elevated prolactin levels and circulating erythroid precursors in dialyzed patients. AB - The prolactin (PRL) receptor (R), a member of the cytokine hemopoietin receptor superfamily, has been shown to activate early differentiation steps along the erythroid pathway. In particular PRL, a product of bone marrow stroma, induces functional erythropoietin (EPO)-R on CD34+ hemopoietic progenitors. In this study, expression of EPO-R mRNA and responsiveness to EPO were assessed on enriched hemopoietic progenitor cells (HPC) from seven hyperprolactinemic and three normoprolactinemic patients and two normal subjects. Expression of EPO-R mRNA by semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR) was found in HPC of four out of seven hyperprolactinemic patients but not in normoprolactinemic patients or normal donors. Development of EPO-dependent Colony Forming Unit-Erythroid (CFU-E) colonies in semi-solid medium was observed only in hyperprolactinemic patients (six out of seven). A much higher number of CFU-E colonies was observed in the four patients with a positive EPO-R message. We conclude from these data that abnormally high levels of PRL may increase the number of EPO-responsive hemopoietic precursors in vivo as they do in vitro. Since hyperprolactinemia associates in these patients with depressed EPO production, it may be regarded as a compensatory mechanism for the reduced availability of the hemopoietic factor. PMID- 10721007 TI - Dietary phytoestrogens have anti-inflammatory activity in a guinea pig model of asthma. AB - Phytoestrogens are a normal constituent of soy protein and have been shown to have anti-inflammatory activity in various in vitro and in vivo models. The present study was designed to determine if a diet enriched in the phytoestrogen isoflavones, genistin and daidzin, would alter the antigen-induced cellular infiltration, particularly eosinophilia, characteristic of a guinea pig model of asthma. Throughout the duration of the study, guinea pigs were maintained on a control diet (standard guinea pig chow) or the same diet enriched in isoflavones. The animals were placed on the diet 2 weeks prior to active sensitization with ovalbumin (OA). Three weeks after sensitization, animals were challenged with OA aerosol. The cellular infiltration into the lung and protein and red blood cells (RBC) in the bronchoalveolar lavage fluid (BAL) were determined 17 hr later. In animals maintained on the control diet, OA aerosol challenge resulted in the expected increase in eosinophils in both the BAL and the lung tissue, an increase in neutrophils in the BAL, and an increase in protein and the number of RBC in the BAL. In contrast, in animals maintained on the isoflavone diet, the OA induced eosinophilia in the lung tissue was significantly attenuated. In addition, OA challenge caused a greater increase in BAL protein in animals maintained on the isoflavone diet compared with animals on the control diet. Our results indicated that a diet enriched in isoflavones results in reduced antigen induced eosinophilia in the lung in the guinea pig model of asthma. However, this beneficial anti-inflammatory effect of dietary phytoestrogens is accompanied by a potentially detrimental increase in antigen-induced leakage of protein into the airspace. PMID- 10721008 TI - Clonal heterogeneity in telomerase activity and telomere length in tumor-derived cell lines. AB - The ribonucleoprotein, telomerase, is responsible for the maintenance of telomere length in most immortal and cancer cells. Telomerase appears to be a marker of human malignancy with at least 85% of human cancers expressing its activity. In the present study, we examined a series of tumor-derived and in vitro immortalized cell lines for telomerase activity levels, telomere lengths, and expression levels of the RNA and catalytic components of telomerase. We found significant variability in both telomere lengths and telomerase activity in clones from tumor cells. In addition, the levels of telomerase components or telomerase activity were not predictive of telomere length. Data from clonally derived cells suggest that critically shortened telomeres in these tumor-derived cell lines may signal activation of telomerase activity through an increase in the expression of the catalytic subunit of telomerase. Although clones with low telomerase shorten their telomeres over time, their subclones all have high levels of telomerase activity with no telomere shortening. In addition, analysis of early clones for telomerase activity indicates substantial variability, which suggests that activity levels fluctuate in individual cells. Our data imply that cell populations exhibit a cyclic expression of telomerase activity, which may be partially regulated by telomere shortening. PMID- 10721009 TI - Evidence of endogenous mono-ADP-ribosylation of cardiac proteins via anti-ADP ribosylarginine immunoreactivity. AB - Arginine-specific mono-ADP-ribosylation of proteins and arginine-specific mono ADP-ribosyltransferase occur in heart. We developed a polyclonal antiserum, R-28, against ADP-ribosylpolyarginine that recognized mono-ADP-ribosylated proteins and identified the major mono-ADP-ribosylation products of quail heart. Treatment of Immobilon-bound ADP-ribosylated Gs protein with hydroxylamine under conditions that remove ADP-ribose from its arginines eliminated R-28 immunoreactivity to Gs. Also, R-28 immunoreactivity to quail heart proteins was removed by NaOH and phosphodiesterase I treatments. Similar treatment with mercuric chloride did not remove the immunoreactivity but did remove exogenously (via in vitro pertussis toxin treatment) added ADP-ribose from cysteine of cardiac Gi/Go proteins. The antiserum did not appear to react with ADP-ribosylasparagine of Rho (formed by C3 toxin), ADP-ribosyldiphthamide of elongation factor 2 (formed by diphtheria toxin) in quail heart preparations, or polyADP-ribosylated proteins of a neonate rat cardiac nuclear preparation. Thus, the R-28 antiserum appears to contain predominantly antibodies directed against ADP-ribosylarginine. To test the usefulness of R-28, immunoblotting of subcellular fractions of quail heart was performed. R-28 showed the greatest immunoreactivity in the sarcolemma with significant immunoreactivity in denser membrane fractions. The cytosol also contained an immunoreactive band distinct from those found in the membranes. Hydroxylamine treatment eliminated immunoreactivity in the sarcolemma and denser membrane fractions but not the cytosol, suggesting the membranous immunoreactive bands contain ADP-ribosylarginine. In conclusion, a polyclonal antiserum that recognizes ADP-ribosylarginine proteins has been raised. The usefulness of the antiserum is demonstrated by the characterization of endogenous arginine mono-ADP ribosylation products in quail heart. The quail heart has several sarcolemmal and denser membrane fraction proteins that appear to be mono-ADP-ribosylated on arginines. PMID- 10721011 TI - A nice challenge for health economics. PMID- 10721010 TI - Aluminum increases levels of beta-amyloid and ubiquitin in neuroblastoma but not in glioma cells. AB - Several epidemiological studies suggest the involvement of aluminum (Al) in the pathogenesis of Alzheimer's disease (AD). There is an increase in the levels of Abeta and ubiquitin in the pathological lesions of AD. Therefore, we have investigated whether aluminum (Al) treatment alters the levels of Abeta and ubiquitin in murine neuroblastoma (NBP2) and rat glioma (C-6) cell cultures. At a low concentration (10 microM), aluminum sulfate stimulated the level of immunoreactive Abeta and ubiquitin in NBP2 cells without changing the levels of the amyloid precursor protein (APP). However, at higher concentrations (100 and 500 microM), aluminum failed to elicit any significant effect on beta-amyloid, whereas ubiquitin levels continued to increase. No changes in the Abeta and ubiquitin content were found in the C-6 glioma cells following treatment with Al at any of the concentrations tested. Exposure of cells to aluminum salts did not alter the rate of proliferation in either of the two cell lines. These data suggest that one of the mechanisms by which Al may play a role in AD is by promoting the formation of Abeta and ubiquitin in neurons. PMID- 10721012 TI - Enhancing performance in health care: a theoretical perspective on agency and the role of information. AB - This paper examines the role of information in securing control of health care systems. The discussion focuses on the impact of the proposed 'Performance Framework', which entails a significant increase in the importance attached to formal performance indicators in the management of the UK National Health Service. The paper starts with a discussion of the role of performance data in securing organizational control within health care systems and summarizes recent research into the behavioural consequences of seeking to control health care agents using such information. A theoretical principal/agent model is then used to illustrate the incentives that exist for dysfunctional behaviour within health care when only imperfect information systems are available. The theoretical results are then examined in the context of a qualitative empirical study, which elicited the perceptions of managers and health care professionals connected with eight NHS hospitals. The study confirmed the existence and importance of serious dysfunctional consequences arising from the use of information as a means of control, and concludes that the Performance Framework will be successful only if it is used in careful conjunction with other means of control. PMID- 10721013 TI - Sensitivity and perspective in the valuation of health status: whose values count? AB - The literature was studied on the existence of differences in valuation for hypothetical and actual health states between patients and other-rater groups. It was found that nine different study designs have been used to study this question and two of these designs were applied in a study involving dialysis patients and other rater groups. In the first study, both dialysis patients and students had to value hypothetical health states with Standard Gamble (SG) and Time Trade Off (TTO). Patients assigned higher values to hypothetical health states than students did. In the second study, dialysis patients who were being treated with four different dialysis modalities were asked to value their own health state with SG, TTO and a visual analogue scale (EQ(VAS)), and to describe their health state on the EQ-5D(profile). Several EQ-5D(index) values (health index values derived from general population samples) were calculated for the four dialysis treatment groups, based on the EQ-5D(profile). These health indexes could discriminate between treatment groups, according to clinical impressions. Treatment groups could not be differentiated based on patients' valuations of own health state. The results suggest that general population samples, using EQ 5D(index) values, may be more able to discriminate between patient groups than the patients themselves are. The implications of this finding for valuation research and policy-making are discussed. PMID- 10721014 TI - Societal value, the person trade-off, and the dilemma of whose values to measure for cost-effectiveness analysis. AB - In a previous paper, it was argued that Societal Value measurement through person trade-off (PTO) elicitation offers a way to include the values of both general public and patients into cost-effectiveness analysis (CEA). It was said that patients' values could be used to estimate the effect that various health care dimensions have on health-related utility and that public values could be used to estimate the Societal Value of these changes in utility. However, this previous proposal still creates opportunities for the public to misvalue the benefit of health care interventions because of bias or misunderstanding about what the health-related utility really is of various illnesses or disabilities. A procedure that combines patient and public values into CEA to partially correct for this bias is suggested in this paper. In addition, it is pointed out that, although Societal Value measurement offers a role for distinctly public preferences in CEA, it still does not answer the question of whose utilities ought to be included in CEA. PMID- 10721015 TI - Public preferences for the allocation of donor liver grafts for transplantation. AB - To investigate the nature of public preferences in the allocation of donor liver grafts for transplantation a social conjoint analysis (CA) technique was developed for a questionnaire survey. A convenience sample of academic and non academic employees of a British University were invited to participate in the survey. Respondents were presented with eight choice situations in which they were asked to allocate 100 donor liver grafts between two groups of 100 individuals in urgent need of a transplant. The groups of individuals differed in terms of the length of time spent waiting, the life years gained following transplantation, age, personal responsibility for their illness and whether they were primary or re-transplant candidates. Only two respondents (0.7%) consistently chose to give all of the donor organs to the group of individuals with the highest expected length of survival whilst seven respondents (2%) exhibited strict egalitarian preferences, allocating equal numbers of donor organs to both groups irrespective of their characteristics. The vast majority of respondents indicated that they would be prepared to sacrifice some gain in the efficiency of the transplantation programme for an increase in equity or fairness in the allocation of donor livers. Using social CA it was possible to establish the relative weight attached to each characteristic in determining individual's allocation decisions. PMID- 10721016 TI - Establishing health state valuations for disease specific states: an example from heart disease. AB - This study considers the feasibility of defining a QALY from disease-specific data using the New York Heart Association (NYHA) classification of heart failure. The study derives health state values for the four different NYHA classifications of disease progression using the time trade-off (TTO) instrument associated with the five dimensional (EQ-5D) health state valuation method. Consistent mappings between the disease classification and the chosen QALY instrument are found. With this being the case, the assumption of constant proportionality, which is necessary to define the QALY as an acceptable measure of health related preferences, is considered. It is found that constant proportionality does not hold across the more severe health states, thus questioning the use of QALYs as representing cardinal preference structures. PMID- 10721017 TI - The impact of clean indoor-air laws and cigarette smuggling on demand for cigarettes: an empirical model. AB - This study examines the impact of clean indoor-air laws and smuggling activities on states' per capita cigarette consumption and revenues by using a static demand model. The analysis was based on data for 50 states and the District of Columbia (DC) of the United Sates over the period 1970-1995. The estimated price elasticities of demand for cigarettes ranged from -0.48 to -0.62, indicating that a 10% increase in price would reduce consumption per capita by 4.8% to 6.2%. Anti smoking laws had a significant negative impact on per capita consumption. In 1995, consumption was reduced by 4.7 packs per capita among states with anti smoking laws, or 1.1 billion fewer packs of cigarettes consumed. Both short distance smuggling between neighbouring states and long-distance smuggling from Kentucky, North Carolina and Virginia existed and were significant. Smuggling activities from military bases and Indian reservations, however, were not significant. On average, 6% of states' tax revenues were lost due to smuggling activities in 1995. Results also showed that short-distance smuggling was less important than long-distance smuggling as a source of the revenue loss. PMID- 10721018 TI - Negative and zero time preference for health. AB - The assumption of positive time preference is seldom challenged in analyses of intertemporal choices, despite considerable evidence of zero and negative discount rates. In this study, the majority of respondents have positive discount rates, but a substantial number have negative or zero discount rates. Using probit regression, the perception of the severity of the health-state, gender, education and perception of the questions in terms of difficulty are shown to influence whether individuals have positive discount rates. PMID- 10721019 TI - Are QALYs an appropriate measure for valuing morbidity in acute diseases? AB - In this paper, we examine the problems associated with using quality adjusted life years (QALYs) as the measure of effectiveness to evaluate interventions for acute conditions. We illustrate the way in which using commonly accepted benchmarks for costs per QALY, in order to adopt interventions for acute conditions, might result in decisions that are not consistent with maximizing net societal benefit. We suggest that an alternate methodology, such as willingness to pay, may be more appropriate to make allocation decisions for acute conditions. PMID- 10721020 TI - MASTOCYTOSIS: BASIC ASPECTS, CLINICAL EVALUATION, AND THERAPY. PMID- 10721021 TI - VITILIGO: IS THERE A TREATMENT THAT WORKS? PMID- 10721022 TI - INHERITED EPIDERMOLYSIS BULLOSA: SELECTED OUTCOMES AND A REVISED CLASSIFICATION SYSTEM. PMID- 10721024 TI - ERYTHEMA MULTIFORME. PMID- 10721023 TI - NEONATAL LUPUS ERYTHEMATOSUS. PMID- 10721025 TI - STEROIDS IN ERYTHEMA MULTIFORME (EM). PMID- 10721026 TI - ERYTHEMA MULTIFORME AND STEROIDS. PMID- 10721027 TI - REACTIVE NEUTROPHILIC SKIN DISEASES. PMID- 10721028 TI - PRACTICAL MAGIC: MOLECULAR DIAGNOSTICS FOR THE CLINICIAN. PMID- 10721029 TI - TRANSCRIPTION: A COMMON PATHWAY FOR UNCOMMON GENODERMATOSES. PMID- 10721030 TI - SCLERODERMA. PMID- 10721031 TI - CONTROVERSIES IN PEDIATRIC DERMATOLOGY: SHOULD ALL CONGENITAL NEVI BE EXCISED? PMID- 10721032 TI - Acute regulation and long-term modulation of presynaptic serotonin output. AB - This contribution summarizes the present knowledge about the mechanisms involved in the regulation of presynaptic serotonin release. Under conditions of a permanently altered environmental, (psychosocial, drug-induced, nutritional etc.) input, each one of these mechanisms may be adaptively adjusted to the novel use dependent requirements. Examples of such long-term alterations of the 5-HT-output pattern in distant projection fields of the raphe-neurons are the downregulation of cortical serotonin-transporters after long-term food restriction, the loss of serotonergic nerve terminals caused by substituted amphetamines, the serotonergic hyperinnervation of the frontal cortex seen after olfactory bulbectomy in rats, or the long lasting changes in the levels of tryptophanhydroxylase, of serotonin transporter depression or in the density of serotonergic nerve terminals in distant projection fields caused by long-term antidepressant treatments. All these long-lasting alterations and imbalances of a previously established serotonin-output-pattern will not only affect the impact of individual regional networks on whole brain information processing but also the established synaptic connectivity in distant projection fields of the central serotonergic system. PMID- 10721033 TI - Pet studies of serotonin synthesis in the human brain. AB - The method for measuring serotonin synthesis in human brain uses [11C]alpha methyl-L-tryptophan as a tracer and positron emission tomography. The alpha methyl-L-tryptophan is converted to alpha-methylserotonin, which is not a substrate for monoamine oxidase and therefore accumulates in the brain. In a pilot study published recently, rates of serotonin synthesis were found to be higher in men than in women. This was due to the lower plasma free tryptophan in the women under the experimental conditions used, and does not necessarily reflect the situation in all circumstances. Acute tryptophan depletion lowered brain serotonin synthesis by 90% or more. Patients with borderline personality disorder, who exhibit emotional lability and impulsivity, may have lower brain serotonin synthesis rates than healthy controls. PMID- 10721034 TI - Tryptophan hydroxylase regulation. Drug-induced modifications that alter serotonin neuronal function. AB - Tryptophan hydroxylase is the initial and rate-limiting enzyme in the biosynthesis of the neurotransmitter serotonin. A variety of drugs are known to diminish the function of this enzyme, and possibly cause damage to serotonin neurons. These include the substituted amphetamines methamphetamine and 3,4 methylenedioxy-methamphetamine, as well as L-DOPA, the most common therapy for Parkinsons Disease. In view of the important role for dopamine in the effects of these drugs on tryptophan hydroxylase and on serotonin neurons, we tested whether dopamine could alter the activity of this important enzyme. We found that dopamine-derived quinones, but not dopamine, inactivate tryptophan hydroxylase and convert the protein to a redox-cycling quinoprotein. This posttranslational modification of tryptophan hydroxylase could play a role in the drug-induced reduction in serotonin synthesis. PMID- 10721035 TI - Monoamine depletion in non-pharmacological treatments for depression. AB - Non-pharmacological treatments such as light therapy for seasonal affective disorder or sleep deprivation for non-seasonal depression have been shown to treat depression effectively. With the use of the tryptophan depletion paradigm and the catecholamine depletion paradigm we assessed the role of brain serotonergic and catecholaminergic systems respectively. We found that disturbances in brain serotonin systems play a key role in the pathogenesis of seasonal affective disorder and that light therapy may compensate for the underlying deficit. Moreover there is evidence that catecholaminergic systems may be involved in the mechanism of action of light therapy. Tryptophan depletion studies suggest that sleep deprivation does not exert its antidepressant effects by involving brain serotonin systems alone. Interestingly, tryptophan depletion prevented the relapse after the recovery night, possibly by enhancing brain serotonin transmission after the depletion procedure. PMID- 10721036 TI - The tryptophan depletion test. Impact on sleep in healthy subjects and patients with obsessive-compulsive disorder. AB - The tryptophan depletion test is a research tool to study the functional consequences of decreasing the brain serotonin metabolism. Since serotonin is involved in sleep regulation and assumed to be of high importance in the etiology of psychiatric disorders, the acute polysomnographic effects of tryptophan depletion were studied in healthy subjects and patients with obsessive compulsive disorder (OCD). According to the reciprocal interaction model of non-REM and REM sleep regulation we expected that tryptophan depletion in healthy controls should provoke alterations of sleep similar to depression, whereas we assumed that these effects would be more pronounced in patients with OCD. METHODS: 12 healthy subjects with a mean age of 34 years and 12 patients suffering from OCD with a mean age of 30 years had 4 polysomnographic investigations. After 1 adaption and 1 baseline night subjects received a low protein diet on day 3 and 4 until midday. On day 4 at 18.00 h subjects ingested an aminoacid mixture devoid of tryptophan. This procedure resulted in a decrease of 85% in healthy subjects and 80% in OCD patients at 22.00 h. RESULTS: The tryptophan depletion led to more pronounced disturbances of sleep continuity in OCD patients than in healthy subjects in terms of an increase of wake time and a decrease of total sleep time. In both groups a decrease of sleep stage 2 could be observed. Healthy subjects showed significant alterations of phasic REM parameters as REM density and total number of rapid eye movements, what was not the case for OCD patients. CONCLUSIONS: Our results indicate the important role of the serotonergic system for sleep maintainance and the phasic aspects of REM sleep. Furthermore our data demonstrate that the tryptophan depletion test is a useful tool to evaluate the hypothesis of a serotonergic involvement in sleep regulation and the etiology of psychiatric disorders. PMID- 10721037 TI - 5-HT and human anxiety. Evidence from studies using acute tryptophan depletion. AB - There is abundant evidence that serotonin (5-hydroxytryptamine, 5-HT) is involved in anxiety in both animals and humans but there is conflicting evidence for the precise role it plays. Acute tryptophan depletion provides a technique for investigating a global reduction in brain 5-HT function and we have investigated its effect on anxiety in drug-free panic disorder patients and normal volunteers. We found little effect on general levels of anxiety but it enhanced the effect of a panic challenge using 5% carbon dioxide (5%CO2) in panic disorder patients. The effect in normal volunteers was less clear with no overall effect following 5%CO2 challenge or the psychological challenge of a simulated public speaking task. These results are discussed in relation to the literature and are broadly supportive of the hypothesis that 5-HT acts to inhibit panic anxiety at the level of the periaqueductal grey but facilitates general and conditioned anxiety at the level of medial temporal lobe structures. PMID- 10721038 TI - Plasma L-tryptophan depletion and aggression. AB - There is a well-established relationship between aggression and lowered serotonin neuro-transmission. Recently developed methodologies for manipulating L tryptophan levels (and brain serotonin) have been applied to human laboratory studies of aggression. Collectively, these studies provide further evidence for the serotonin-aggression relationship. Two important findings have been made recently: (1) subsets of individuals (e.g., persons self-rating high on aggressive or hostility scales) may differ in their susceptibility to aggression produced through plasma tryptophan depletion; and (2) alcohol in combination with L-tryptophan depletion has an additive effect on aggression. All previous studies have been conducted with men. Extending these studies to women appears to be the much-needed next step given that serotonergic levels appear to vary both as a function of the menstrual cycle phase and menstrual symptomatology. PMID- 10721039 TI - A comparison of the effects of acute tryptophan depletion and acute phenylalanine/tyrosine depletion in healthy women. AB - Acute tryptophan depletion (ATD), which is thought to lower serotonin levels, can result in a lowering of mood. In the present study we compared the effect of ATD with acute phenylalanine/tyrosine depletion (APTD) in healthy women. Although considerable evidence relates catecholamines to the regulation of anxiety, there was no difference in anxiety responses in the ATD and APTD groups when the women underwent a mildly stressful psychological challenge. Both ATD and APTD caused a similar lowering of mood. Both depletions also increased heart rate. These results suggest that APTD is a useful method for studying the effect of low catecholamine levels in humans, and that catecholamines are involved in the regulation of mood. PMID- 10721040 TI - Fructose malabsorption is associated with decreased plasma tryptophan. AB - Fructose malabsorption is characterized by the inability to absorb fructose efficiently. Consequently fructose reaches the colon and is broken down by bacteria to short-fatty-acids, CO2 and H2. Recently we found that fructose malabsorption was associated with signs of depression. It was therefore of interest to find out whether fructose malabsorption is associated with abnormal tryptophan metabolism. Breath hydrogen concentrations were measured in 50 after an oral dose of 50 g fructose allowing to classify them as normals (n = 15) or fructose malabsorbers (n = 35). Blood samples were taken for tryptophan and kynurenine measurements. Fructose malabsorbers showed significantly lower plasma tryptophan concentrations and significantly higher depression scores compared to normals. Fructose malabsorption is associated with lower tryptophan levels which may play a role in the development of depressive disorders. PMID- 10721041 TI - Use of melatonin in the treatment of phase shift and sleep disorders. AB - When administered to humans the pineal hormone melatonin can phase shift a number of circadian rhythms. This property has prompted the investigation of exogenous melatonin in sleep disorders known to have an underlying chronophysiological basis (i.e. circadian rhythm sleep disorders). Both in field and simulated studies of jet lag and shift work suitably timed melatonin improved sleep and, in some cases, hastened readaptation of the circadian rhythms following the phase shift. Melatonin treatment has also been evaluated in the circadian sleep disorders: delayed sleep phase syndrome (DSPS) and non-24-hour sleep wake disorder. Compared with placebo, melatonin advanced the sleep period in subjects with DSPS. Melatonin also improved a number of sleep parameters in blind subjects suffering from non-24-hour sleep wake disorder. PMID- 10721042 TI - A placebo-controlled study of the effects of L-tryptophan in patients with premenstrual dysphoria. AB - In a randomized controlled clinical trial, 37 patients with premenstrual dysphoric disorder were treated with L-tryptophan 6 g per day and 34 were given placebo. The treatments were given under double-blind conditions for 17 days, from the time of ovulation to the third day of menstruation, during three consecutive cycles. Visual Analog Mood Scales revealed a significant (p = 0.004) therapeutic effect of L-tryptophan relative to placebo for the cluster of mood symptoms comprising the items dysphoria, mood swings, tension and irritability. These results suggest that increasing serotonin synthesis during the late luteal phase of the menstrual cycle is therapeutic in patients with premenstrual dysphoric disorder. PMID- 10721044 TI - The serotonergic appetite suppressant fenfluramine. Reappraisal and rejection. AB - Medical and social pressures have led to increased emphasis on dieting. However, there has been a concurrent world wide increase of obesity. Therefore, much attention has been paid to the development of drugs which decrease appetite. The most extensively used drug of this type over the past three decades has been the serotonergic compound fenfluramine. Recent findings have cast doubt on the previously accepted view that its action requires the release of central 5-HT. Instead, it seems likely that action on specific 5-HT receptors independently of 5-HT stores is involved. It is ironic that these new developments in understanding its mechanism of action have coincided with the recognition of its cardiovascular side-effect apparent especially in patients treated with d fenfluramine combined with phentermine. This has forced the withdrawal of fenfluramine (both as racemate and d-isomer) from clinical use. The implications of these developments are commented upon. PMID- 10721043 TI - Nocturnal melatonin secretion and its modification by treatment in patients with sleep disorders. AB - Data on the circadian melatonin secretion in sleep disordered patients and effects of sleep medication on melatonin are still missing. We studied plasma melatonin concentration, sleep, and effects of some hypnotics in 15 patients and 10 controls. Nocturnal melatonin levels were significantly decreased in patients with a more than five years history of sleep complaints compared to controls or patients with a shorter duration of illness. Independent of their sleep promoting properties drugs increased or decreased nocturnal melatonin in controls and patients. Patients with chronic sleep-wake rhythm disorders showed altered relations between their circadian melatonin secretion pattern and sleep. We conclude that nocturnal melatonin secretion is primarily independent of sleep regulation but represents a neuroendocrine feature of chronically disturbed sleep. PMID- 10721045 TI - Serotonin, dieting, and bulimia nervosa. AB - Dieting is a common behaviour which may trigger eating disorders such as bulimia nervosa in predisposed subjects. We found that in healthy women moderate dieting for 3 weeks lowered plasma concentrations of the 5-HT precursor, L-tryptophan (TRP) and impaired brain 5-HT neurotransmission as judged by 5-HT neuroendocrine tests. In recovered female subjects with a history of bulimia nervosa we found that TRP depletion produced by an amino acid mixture lacking TRP caused a temporary return of depressive symptoms together with concerns about weight and shape and fear of loss of control of eating. Taken together the data suggest that dieting-induced decreases in TRP availability may trigger the development of bulimia nervosa is susceptible individuals. PMID- 10721046 TI - Peripherally injected IL-1 induces anorexia and increases brain tryptophan concentrations. AB - Interleukin-1 is an anorexigenic cytokine, and is involved in the pathogenesis of cancer anorexia. Interleukin-1 induced anorexia is mediated by direct action within the hypothalamus, and by peripheral mechanism(s) yet to be determined. Here we present evidence showing that in an animal model the peripheral injection of interleukin-1 is followed by a significant rise in brain tryptophan concentrations. Tryptophan is the precursor of the neurotransmitter serotonin, known to mediate the onset of satiety under normal and pathological conditions. By inference, we conclude that interleukin-1 induced anorexia is mediated by at least two different mechanism: i) interleukin-1 direct action within the hypothalamus; ii) increased brain serotonergic activity, secondary to interleukin 1 induced increased brain availability of the serotonin precursor, tryptophan. PMID- 10721047 TI - Involvement of nitric oxide in the 5-HT1A autoreceptor-mediated hyperphagia in rats. AB - Effects of nitric oxide synthase(NOS) inhibitors on 8-hydroxy-2-di-n (propylamino)tetralin (8-OH-DPAT)-induced hyperphagia which is mediated by the 5 HT autoreceptor were investigated. The non-selective NOS inhibitor NG-nitro-L arginine methyl ester (L-NAME) and neuronal NOS (nNOS) inhibitor 7-nitroindazole (7-NI) clearly suppressed increases in food intake by 8-OH-DPAT. Both hypophagic effects of L-NAME and 7-NI were reversed by the nitric oxide precursor, L arginine. The findings suggest that nitric oxide formed in the brain is involved in 8-OH-DPAT-induced hyperphagia. PMID- 10721048 TI - Modulation and function of kynurenic acid in the immature rat brain. AB - Using in vivo and in vitro paradigms, the regulation and function of the brain metabolite kynurenic acid (KYNA) was examined in rats on postnatal days (PND) 7 and 14. As shown previously in adult rats, glucose removal and d-amphetamine (d Amph) administration caused decreases in KYNA formation, while exposure to pyruvate up-regulated KYNA synthesis. The effect of glucose deprivation was substantially blunted in immature animals. In PND 14 rats, d-Amph pre-treatment exacerbated the excitotoxic effects of an intrastriatal N-methyl-D-aspartate (NMDA) injection. This potentiation was prevented by m-nitrobenzoylalanine, a kynurenine 3-hydroxylase inhibitor that also antagonized the KYNA reduction caused by d-Amph. These and additional experiments with the competitive NMDA receptor antagonist CGP 40116 indicate the existence of a functionally significant, novel high-affinity receptor for KYNA in the brain. PMID- 10721049 TI - Kynurenine pathway metabolism in human astrocytes. AB - The involvement of astrocytes in Kynurenine pathway (KP) metabolism is still poorly understood. In the present study, we investigated the ability of human fetal astrocytes in vitro to produce quinolinic and picolinic acids using mass spectrometry. In parallel, we estimated the level of expression of five major KP enzymes using RT-PCR. The results demonstrated that astrocytes express most KP enzymes, except for kynurenine-hydroxylase. This in vitro study provides novel informations regarding the ability of human fetal astrocytes to degrade L tryptophan along the KP. PMID- 10721050 TI - Degradation of tryptophan in neurodegenerative disorders. AB - In patients with neurodegenerative disorders, namely Alzheimer's disease and Huntington's disease, we compared serum concentrations of tryptophan, kynurenine and the kynurenine per tryptophan ratio with concentrations of soluble immune activation markers. Significantly lower tryptophan concentrations were observed in the patients, and lower tryptophan levels as well as higher kynurenine levels and higher kynurenine per tryptophan ratios correlated with higher concentrations of neopterin, and soluble receptors for TNF and interleukin-2. In both groups of patients tryptophan concentrations correlated inversely with the degree of mental retardation. No such association existed for the duration of the disease. The data show that systemic chronic immune activation in patients with Alzheimer's disease and Huntington's disease is associated with significant degradation of tryptophan, which is most likely due to activation of indoleamine (2,3) dioxygenase by immunologic stimuli. Further studies will be necessary to investigate a potential role of tryptophan degradation in the pathogenesis of neurodegenerative disorders. PMID- 10721051 TI - Brain tryptophan/serotonin perturbations in metabolic encephalopathy and the hazards involved in the use of psychoactive drugs. AB - Several combined pathogenetic factors such as hyperammonemia, different brain tryptophan metabolic disturbances and serotonin physiological/pharmacological alterations not yet defined in all details, will often give rise to the clinical neuropsychiatric condition known as hepatic encephalopathy (HE). Indeed, to this the probable exposure to novel potent CNS-monoamine acting drugs today may put such patients at certain risk for other pharmacodynamic (PD) responses than usually are expected from these "safe" drugs. Moreover, with a compromised liver function in HE, also pharmacokinetic (PK) features for the drugs are likely changed in these patients. Thus, the ultimate clinical outcome by this probable but unknown PD/PK-deviation for such psychoactive drugs when given to HE-patients needs further clarification. Accordingly, delineation of both PD- and PK-effects in experimental HE should shed light on this issue of relevance for monoamine active drug safety as well as on some further details in the complex tryptophan/monoamine-related pathophysiology that comes into play in HE. PMID- 10721052 TI - Tryptophan metabolism and hepatic encephalopathy. Studies on the sedative properties of oxindole. AB - Oxindole administration (1-100 mg/kg i.p.) to mammals decreases locomotor activity, reduces muscular tone and blood pressure and at larger doses causes coma and death. Utilizing both HPLC and GC/MS, we showed that oxindole is present in the blood, brain and other organs of several animal species, including humans. We demonstrated that oxindole is a tryptophan metabolite able to significantly decrease neuronal excitability by modifying the function of voltage-operated sodium channels. Its synthesis requires the availability of indole, which is formed in the gut. When liver function is impaired, a sufficient amount of indole reaches systemic circulation and is oxidized into oxindole, which seems to be one of the responsible agents for the neurological symptoms found in the course of liver impairment. PMID- 10721053 TI - Sleep disorders and portal-systemic encephalopathy following transjugular intrahepatic portosystemic stent shunt in patients with liver cirrhosis. Relation to plasma tryptophan. AB - Neuropsychiatric symptoms due to any type of dysfunction and/or portal-systemic shunting are summarized as hepatic encephalopathy (HE). HE in the presence of liver cirrhosis and/or portal-systemic shunting has been termed portal-systemic encephalopathy (PSE). PSE is most frequent among the HE syndromes and is almost exclusively seen in patients with advanced cirrhosis and portal hypertension. Portal-systemic shunting either spontaneous due to portal hypertension, following surgical portocaval anastomosis, or subsequent to transjugular intrahepatic portosystemic stent-shunt (TIPSS) is regarded as the primary causative condition for PSE, not hepatic dysfunction per se. PSE may be considered as a disorder of multiple neurotransmitter systems among which derangements of the serotonergic system have been documented most consistently. Incipient PSE is frequently paralleled by the occurrence of sleep disorders, however, their relation to PSE remains unclear. We observed a transient increase of sleep disorders post-TIPSS, which were only in part correlated to other symptoms of PSE. Among the biochemical parameters studied only an association between arterial ammonia levels and sleep disorders became apparent, whereas no significant relation was observed for peripheral tryptophan. PMID- 10721054 TI - L-5-hydroxytryptophan can prevent nociceptive disorders in man. AB - Prevention of primary pain is a new topic, endowed with social and economic interest. We observed that L-5-HTP can induce a significant decrease of the cropping out of migraine, the commonest primary pain. This finding seems interesting, since it represents the first data in the field and was obtained in a prospective, long-term, placebo controlled study. The result obtained suggests that CNS abnormalities underlying the mechanism of migraine can be changed by L-5 HTP, if the amino acid is administered to subjects who are predisposed to headache. PMID- 10721055 TI - Triptans. A support to the central link between serotonin and acetylcholine in migraine. AB - Recently synthesized triptans, sumatriptan derivatives, display aborting migraine activity at doses and with plasmatic maximum peak dramatically lower (20-40 times) than sumatriptan, the 5-HT like agonist, which is the original molecule. That triptans easily cross blood-brain barrier strongly supports the central theory of migraine. We recently discovered the anti-migraine prophylactic action of centrally acting anti-cholinesterase agents, that seems a further support to the central theory of migraine. Indeed, acetylcholine is an important analgesia neurotransmitter and is intertwined with 5-HT central pathways. PMID- 10721056 TI - Plasma and urinary serotonin and 5-hydroxyindol-3-acetic acid in adults with migraine and tension-type headache. AB - Each headache can be a complex diagnostic, therapeutic, prognostic and social problem. The pain in the head can be connected with many organic and non-organic causes. In this work, the levels of plasma and urinary free 5-HT and 5-HIAA were investigated in eight migraine (aged 23-59 years) and ten tension-type headache suffers (aged 38-61 years). Based on the data obtained and their correlation with clinical features and in comparison with a control group, the following can be stated: (1) there is involvement of serotonin in migraine and tension-type headache during the attacks, although the positive 5-HT values from plasma were small; (2) urinary 5-HT values in migraine and tension-type headache were normal in comparison to the control group; (3) significantly decreased values of 5-HIAA in urine were found both in migraine and tension-type headache groups. These findings show that catabolism of 5-HT is probably decreased during headache periods; (4) visual aura was found in five out of ten subjects with tension-type headache. PMID- 10721057 TI - Neuroprotective effects of kynurenine-3-hydroxylase inhibitors in models of brain ischemia. AB - The neuroprotective effects of two kynurenine hydroxylase inhibitors, (m nitrobenzoyl)-alanine (mNBA) and 3,4-dimethoxy-[-N-4-(nitrophenyl)thiazol-2yl] benzenesulfona mide (Ro 61-8048), were studied in vitro and in vivo. In organotypic hippocampal slice cultures deprived of oxygen and glucose, these inhibitors significantly reduced neuronal damage. In gerbils subjected to bilateral carotid occlusion for 5 min, the administration of mNBA (400 mg/kg i.p., 3 times) or Ro 61-8048 (40 mg/kg i.p., 3 times) dramatically decreased the percentage of damaged pyramidal neurones in the hippocampal CA1 region. Finally, in rats with permanent occlusion of the middle cerebral artery, mNBA (200-400 mg/kg i.p.) and Ro 61-8048 (40 mg/kg i.p.) administration reduced the infarct volume. Our results demonstrate that ischemic neuronal damage may be significantly decreased by inhibiting kynurenine hydroxylase. PMID- 10721058 TI - Further evidences for neuroprotective effects of melatonin. AB - The physiological roles of the pineal hormone melatonin are still not completely clarified. Recently it has been shown that melatonin is a potent, endogenous scavenger of reactive oxygen species suggesting that it might interfere with neurodegenerative processing involving free-radical formation and excitatory aminoacid release. These neuroprotective effects of melatonin may result, at least in part, from a sparing of glutathione reductase, which is decreased following administration of the neurotoxic agent kainate (KA) in rats. Moreover, KA causes a rapid decrease in glutathione (GSH) content of cultured cerebellar granule neurons but not in astrocytes. These cell types both express functional KA receptors, but only the former is sensitive to reactive oxygen species dependent KA injury. Melatonin counteracts the changes in GSH, induced by KA, in cultured cerebellar granule neurons. PMID- 10721059 TI - Therapeutic potential of melatonin in immunodeficiency states, viral diseases, and cancer. AB - Maintenance of health depends on the ability to respond appropriately to environmental stressors via reciprocal interactions between the body and the brain. In this context, it is well recognized that the pineal hormone melatonin (MLT) plays an important role. T-helper cells bear G-protein-coupled MLT cell membrane receptors and, perhaps, MLT nuclear receptors. Activation of MLT receptors enhances the release of T-helper cell cytokines, such as gamma interferon and interleukin-2 (IL-2), as well as activation of novel opioid cytokines which crossreact immunologically with both interleukin-4 and dynorphin B. MLT has been reported also to enhance the production of interleukin-1, interleukin-6 and interleukin-12 in human monocytes. These mediators may counteract secondary immunodeficiencies, protect mice against lethal viral and bacterial diseases, synergize with IL-2 against cancer and influence hematopoiesis. Hematopoiesis is influenced by MLT-induced-opioids (MIO) acting on kappa 1-opioid receptors present on bone marrow macrophages. Clinically, MLT could amplify the anti-tumoral activity of low dose IL-2, induce objective tumor regression, and prolong progression-free time and overall survival. MLT seems to be required for the effectiveness of low dose IL-2 in those neoplasias that are generally resistant to IL-2 alone. Similar findings were obtained in a study in which MLT was combined with gamma-interferon in metastatic renal cell carcinoma. In addition, MLT in combination with low-dose IL-2 was able to neutralize the surgery-induced lymphocytopenia in cancer patients. IL-2 treatment in patients results in activation of the immune system and creates the most suitable biological background for MLT. The finding that MLT stimulates IL-12 production from human monocytes only if incubated in presence of IL-2 further supports this concept. On the other hand, high concentrations of MLT have been found in human breast cancer tissue. The MLT concentration, which was 3 orders of magnitude higher than that present in the plasma, correlated positively with good prognostic markers such as estrogen receptor status and nuclear grade. Whether this relates to the immunoneuroendocrine action of MLT remains to be established. Clinical studies are needed on the effect of MLT in combination with IL-2 or other cytokines in cancer patients and viral diseases including HIV-infected patients. PMID- 10721060 TI - Clinical experiences with the use of indole-3-pyruvic acid. AB - In the last few years, Indol-3-pyruvic acid (IPA) has been subjected to 5 pilot clinical trials in volunteers and a phase II study on patients affected by anxiety, with or without sleep problems. Overall, results indicated a very good safety profile, relief from anxiety and a better quality of sleep in patients treated with IPA. Moreover, the drug showed no withdrawal signs, but positive effects on mood, improving feelings of relaxation, calmness and happiness. The mechanism of action of IPA, depending on increased turnover of some indoles in the CNS, appears clearly distinct from that of benzodiazepines, suggesting that the drug might be used in the treatment of symptoms of mild anxiety and stress experienced by busy and anxious people. PMID- 10721061 TI - Regulation of quinolinic acid synthesis by mitochondria and o methoxybenzoylalanine. AB - o-Methoxybenzoylalanine, a selective kynureninase inhibitor, caused unexpected accumulation of 3-hydroxyanthranilic acid (3OH-ANA), the product of kynureninase activity and the precursor of quinolinic acid (QUIN) in liver homogenates incubated with 3OH-kynurenine (3OH-KYN). In order to explain this observation, we investigated the interaction(s) of o-methoxybenzoylalanine with 3 hydroxyanthranilic acid dioxygenase, the enzyme responsible of QUIN formation. When the purified enzyme, or partially purified cytosol preparations were used, oMBA did not affect 3-hydroxyanthranilic acid dioxygenase activity. The addition of purified mitochondria to 3-hydroxyanthranilic acid dioxygenase preparations reduced the enzymatic activity and the synthesis of QUIN. In the presence of mitochondria oMBA further reduced QUIN synthesis. The administration of oMBA reduced QUIN content in both blood and brain of mice. Our results suggest that mitochondrial protein(s) interact(s) with soluble 3-hydroxyanthranilic acid dioxygenase and cause(s) modifications in the enzyme resulting in a decrease in its activity. These modifications also allow the enzyme to interact with oMBA, thus leading to a further reduction in QUIN synthesis. PMID- 10721062 TI - Regulation of indoleamine 2,3-dioxygenase, the first enzyme in UV filter biosynthesis in the human lens. Relevance for senile nuclear cataract. AB - 3-Hydroxykynurenine (3OHKyn), the precursor of UV filters in human lens, is highly autooxidizable, generates H2O2, and binds to lens proteins, yielding a tanned/yellow product resembling senile nuclear cataractous materials. Thus, if 3OHkyn can be shown to be the causative agent in cataract, it may be possible to prevent the disease by lowering the level of 3OHKyn. To this end, indoleamine 2,3 dioxygenase, the first enzyme in UV filter synthesis, was studied using lens epithelial cell lines. The results indicated that the IDO expression is mediated by IFN-gamma. Immuno-suppressants which inhibit production of IFN-gamma may act as anti-cataract agents. Another way to lower the level of 3OHKyn is to use specific inhibitors for IDO. A recombinant human IDO was expressed to develop the inhibitors. PMID- 10721063 TI - Melatonin in cancer patients and in tumor-bearing animals. AB - A review of findings is given which relate to the levels of circulating melatonin as well as the urinary excretion of its main peripheral metabolite 6 sulphatoxymelatonin (aMT6s) in patients with different types of cancer as well as in tumor-bearing animals. Clinical results show that circulating melatonin tends to be depressed in patients with primary tumors of different histological types including both endocrine-dependent (mammary, endometrial, prostate cancer) and endocrine-independent tumors (lung, gastric, colorectal cancer). Reduction of melatonin is most pronounced in patients with advanced localized primary tumors, such as mammary and prostate cancer where a clear negative correlation with tumor size exists. The phenomenon of a reduction of circulating melatonin appears to be a transient one since patients with recidives show a normalization of melatonin. Surgical removal of the primary tumor does, however, not lead to normalization indicating that complex systemic changes appear to be involved in the down regulation of melatonin. It is unclear at present, whether circulating melatonin is depleted in cancer patients due to a reduced production by the pineal gland or due to certain peripheral metabolic processes, although no evidence for an enhanced hepatic degradation to aMT6s, the main peripheral metabolite of melatonin, was found. The reduction of circulating melatonin is accompanied by neuroendocrine changes affecting the circadian secretion of the adenohypophyseal hormones prolactin, somatotropin and thyroid-stimulating hormone. In contrast to the above-described types of tumors many patients with ovarian cancer show highly elevated levels of melatonin perhaps due to the production of tissue-specific growth factors that could affect pineal melatonin secretion. Experiments with tumor-bearing animals clearly demonstrate that nocturnal circulating melatonin is modulated due to malignant growth. Detailed investigations with chemically induced mammary tumors in rats and serial transplants derived thereof show that slow-growing and well-differentiated tumors containing epithelial cell elements (adenocarcinomas and carcinosarcomas) lead to an enhanced production of melatonin involving activation of the rate-limiting enzyme of pineal melatonin biosynthesis (serotonin N-acetyltransferase) probably due to elevation of the sympathetic tone in response to a stimulation of the cellular immune system by malignant growth. As opposed to that nocturnal melatonin is depleted in animals with fast-growing mammary tumor transplants when myoepithelial-mesenchymal conversion leads to pure sarcomas. The reduction of melatonin appears to be due to either a reduced availability of the precursor amino acid tryptophan because of a glucocorticoid induced activation of the hepatic enzyme tryptophan 2,3-dioxygenase or a direct peripheral degradation of melatonin via indoleamine 2,3-dioxygenase expressed in tumor and/or other tissues. The significance of these clinical and experimental findings relating to melatonin is discussed both in terms of their practical application as a possible tumor marker and from a theoretical point of view to understand better the mechanisms involved in complex host-tumor interactions involving the neuroimmunoendocrine network. PMID- 10721064 TI - Tryptophan metabolism in alcoholism. AB - Studies of tryptophan (Trp) metabolism in relation to the serotonin status in alcoholism are of 2 types: (1) those related to the pharmacological effects of ethanol; (2) those concerning the serotonin status in the absence of alcohol intake. In experimental animals, acute and chronic ethanol administration and subsequent withdrawal exert a variety of effects on brain serotonin synthesis and turnover mediated by corresponding changes in Trp availability to the brain secondarily mainly to modulation of liver Trp pyrrolase (TP) activity. Alcohol preferring mice and rats exhibit a central serotonin deficiency caused by, or in some cases associated with, a higher TP activity. Liver TP also appears to be a target of ethanol in man and evidence has recently emerged that alcoholics with positive family history are serotonin-deficient because of a lower availability of circulating Trp to the brain. Acutely, ethanol depletes brain serotonin in normal subjects, which may explain alcohol-induced aggression in susceptible individuals and also the incidence of depression in alcoholism. Trp availability to the brain is increased before the appearance of the alcohol-withdrawal syndrome in man, raising the possibility that the associated behavioural disturbances may involve the excitotoxic Trp metabolite quinolinate. Further studies of the Trp and serotonin status in relation to these important clinical features of alcohol dependence and alcoholism may therefore yield fruitful results. PMID- 10721065 TI - Effect of cisplatin and paclitaxel on plasma free tryptophan levels. An in vitro study. AB - Emesis is a common side effect of some antineoplastic drugs. Cisplatin (CP) induces a biphasic pattern of emesis referred to as acute (AE) and delayed (DE) emesis. The serotonergic system plays a major role in the pathogenesis of CP induced AE, as suggested by the therapeutic efficacy of 5HT3 receptor antagonists. The pathogenesis of CP-induced DE are not clear. To date, there are no pharmacological agents which satisfactorily control DE. We hypothesize that increased availability of tryptophan (TRP) for the synthesis of brain serotonin (5-HT) could, at least in part, contribute to CP-induced DE. In fact, within 2-4 hrs of administration, CP is largely bound to albumin (ALB) with consequent possible displacement of TRP which circulates in plasma mostly (90% of total plasma TRP) bound to its natural carrier, ALB. To test this hypothesis, we studied in vitro the effect of increasing doses of cisplatin on F-TRP in plasma obtained from healthy volunteers. We also tested the effects of therapeutic amounts of paclitaxel, an antineoplastc agent which does not cause emesis. RESULTS: F-TRP concentrations increased in a dose-dependent manner following incubation with cisplatin, in contrast to paclitaxel (PTX). CONCLUSIONS: The preliminary data obtained suggest that CP, but not PTX, at therapeutic doses, increases plasma F-TRP concentrations. This increase has likely negligible relevance in CP-induced AE, which is induced by the 5-HT released by the enterochromaffin cell system, while it might play a role in the pathogenesis of CP-induced DE. In fact, CP binding to ALB is stable for 4-5 days following administration, thus suggesting long-term TRP displacement from ALB and enhanced brain 5-HT synthesis and release. Whether increased TRP availability for 5-HT synthesis might be the pathogenic mechanism for CP-induced DE in vivo, is currently being tested. PMID- 10721066 TI - Effect of kynurenine on tryptophan-albumin binding in human plasma. AB - Increased plasma free tryptophan levels have been reported in cancer patients and causally associated to the presence of anorexia. The pathogenesis of this occurrence is yet to be completely understood. Kynurenine is a metabolite of tryptophan, and has been reported increased in plasma during tumor growth. Because of the similarities between tryptophan and kynurenine we speculate that their rise in the presence of a tumor might be causally related. To test this hypothesis, we performed a series of in-vitro studies, showing that kynurenine supplementation reduces the amount of tryptophan bound to albumin, and thus, by competition, increases free tryptophan levels. The likely clinical consequences of these findings are discussed. PMID- 10721067 TI - Vitamin effects on tryptophan-niacin metabolism in primary hepatoma patients. AB - Metabolism of 14C-labeled tryptophan and 3-hydroxyanthranilic acid were administered to early hepatoma patients to evaluate the conversion of these precursors to niacin metabolites and to assess the effect of dietary supplementation with vitamin B-6, riboflavin, thiamin and vitamin C on the extent of conversion. Expired labeled carbon dioxide and urinary excretion of picolinic acid (PA), quinolinic acid (QA), nicotinic acid (NA), N1-methylnicotinamide (N1MeNAm) and N1-methyl-2-pyridone-5-carboxamide (MPCA) were measured by carrier isolations. There were no consistent statistical differences in these conversions before and after vitamin supplementation, suggesting that the patients' nutrition was adequate and that none of the vitamins were rate-limiting under these conditions. PMID- 10721068 TI - Serum tryptophan to large neutral amino acid ratio and urinary tryptophan in three patients with phenylketonuria in a family. A clinical and biochemical study. AB - In this work clinical and biochemical findings are presented in three untreated children with phenylketonuria in a family. Their clinical pictures were not typical for classical phenylketonuria. As a result, diagnosis was missed. It has been shown that patterns of large neutral amino acids in serum and urine were somewhat different. Significantly lower serum TRP/LNAA ratio was observed in all patients with phenylketonuria, compared to the control group. These findings suggest that there was subnormal tryptophan availability in the central nervous system leading to its decreased metabolism through the serotonin and kynurenine pathways. These results may explain decreased children's growth and their mental deficiency. PMID- 10721069 TI - Plasma and urinary serotonin and 5-HIAA in children treated with lamotrigine for intractable epilepsy. AB - Alteration of monoamine levels by some antiepileptic drugs (AEDs) was elucidated in this study. Lamotrigine (LTG) is a new AED, acting the sodium-channels. LTG was given as add-on therapy to 16 patients aged 4.5-18 yrs with intractable epilepsy and comedicated with carbamazepine or valproate. An equal group of epileptics with comparable clinical characteristics and treatment served as control. Plasma and urinary (24 h-samples) serotonin and 5-HIAA were determined before onset of LTG therapy and after 2-3 months. HPLC and electrochemical detection was used for the determination of serotonin (5-HT) and 5-hydroxy indoleacetic acid (5-HIAA). No significant effect of LTG on both urinary 5-HT and 5-HIAA levels was found, whereas plasma 5-HT concentrations significantly decreased in comparison with levels before LTG starting and relevant values in controls. This findings was noted in 7/16 children with favourable response to LTG. Increased serotonin catabolism may be result of LTG action. PMID- 10721071 TI - Labeled kynurenine pharmacokinetic modeling studies in gerbils. Nonequilibrium between infused and endogenous kynurenine. AB - In order to complete pharmacokinetic studies on the central vs. peripheral origin of several tryptophan metabolites, we infused gerbils with labelled kynurenine (2H4 or 15N2). Osmotic minipumps charged with kynurenine solutions were surgically implanted subcutaneously in adult female gerbils (50-60 g). After a variable number of hours, the gerbils were sacrificed and organs taken for determination of labelled/unlabelled kynurenine ratios using mass spectrometric assay of a pentafluorobenzyl derivative as described previously. Surprisingly high ratios of 2H to 1H-kynurenine were measured in the kidney (0.25-0.40) and urine (4.0-8.0), although the ratio of deuterium labelled to endogenous kynurenine remained below detection limits (< 0.05) in serum and other tissues. Infusion of greater quantities of 2H4-kynurenine confirmed these observations in gerbils in which ratios of 2H4-to-1H kynurenine were measurable in serum and tissues. Synthesis and infusion of 15N2-kynurenine demonstrated that these effects were not due to deuterium isotope substitution. The data demonstrate a non-equilibrium between infused and endogenous kynurenine, which is related to differential rates of protein binding and the rapid clearance of free, infused kynurenine by kidney. PMID- 10721070 TI - Effect of continuous melatonin infusions on steady-state plasma melatonin levels, metabolic fate and tissue retention in rats under near physiological conditions. AB - The fate and disposition of melatonin released into the circulation is still poorly understood, and almost all current knowledge is derived from measurements made after one single, often very large dose of labeled melatonin. In continuous infusion experiments in freely moving rats, 500 ng melatonin/mL hr had to be infused in order to elevate the circulating hormone from low daytime levels to the 10-fold higher nocturnal steady state concentrations. To study the fate and tissue accumulation of the infused melatonin, tritiated melatonin was added to the infusion solution, and the retention of [3H]-melatonin and chloroform insoluble [3H]-melatonin-metabolites were measured in almost all body tissues and their subcellular compartments immediately at the end of the infusion period and six hours later. A considerable amount of the infused melatonin was found in the gut and in all tissues, some melatonin was covalently attached to proteins. PMID- 10721072 TI - Antioxidative properties of serotonin and the bactericidal function of polymorphonuclear phagocytes. AB - We investigated the antioxidative properties of platelet-released serotonin on the bactericidal function of polymorphonuclear neutrophils (PMN) since there is a surprising co-incidence of low blood serotonin and an increased rate of infections. The antioxidative properties of serotonin were demonstrated by its suppressive effects on phagocytosis-associated, luminol-enhanced chemiluminescence (CL). The bactericidal activity of PMN was determined by a microbiological assay using opsonized Staphylococcus aureus. Serotonin suppresses luminol-enhanced chemiluminescence in a dose-dependent manner indicating an interaction with reactive oxygen species, which are responsible for effective bacterial killing during the phagocytosis-associated "respiratory burst". The modulation of the bactericidal function of PMN by serotonin is complex and depends upon the amount of serotonin: at concentrations normally present at sites of tissue injury and consecutive thrombus formation (10(-6) to 10(-5) M) bacterial killing increases by about 50%. In contrast, at pharmacological concentrations (10(-3) to 10(-2) M) an adverse effect can be observed: the elimination of opsonized S. aureus is reduced by 30 to 90%. Exogenous serotonin is capable of modulating important biological functions of human PMN in vitro. At appropriate concentrations, the antibacterial defence improves significantly probably due to reduced autooxidation, whereas higher concentrations counteract an efficient bacterial killing. PMID- 10721073 TI - Preventive effect of melatonin on the progression of carbon tetrachloride-induced acute liver injury in rats. AB - We examined the preventive effect of melatonin on the progression of carbon tetrachloride (CCl4)-induced acute liver injury in rats. In rats injected with CCl4 (1.0 ml/kg body weight), liver injury appeared 6 h after the injection and progressed at 24 h. This liver injury progression was prevented by treatment with melatonin (50 or 100 mg/kg body weight), which was conducted 6 h after CCl4 injection. An increase in liver lipid peroxide content and a decrease in liver reduced glutathione content occurred with the liver injury progression. These changes were prevented by the post-treatment of melatonin (50 or 100 mg/kg body weight). These results indicate that melatonin can prevent the progression of CCl4-induced acute liver injury through its antioxidant action. PMID- 10721074 TI - Effects of peroxisome-proliferators on the TRP-NAD pathway. AB - The mechanism of the elevation of hepatic NAD level in the rats administered clofibrate and other peroxisome-proliferators were investigated. In the hepatocytes from the clofibrate-fed rats, NAD biosynthesis from Trp, but not from nicotinic acid, was specifically stimulated. The activities of peroxisomal marker enzymes, the peroxisome-proliferator activated receptors (PPAR)-dependent enzymes and key enzymes in the Trp-NAD pathway changed in parallel with the hepatic NAD increase; the activity of quinolinate phosphori-bosyltransferase (QPRT) was increased whereas that of alpha-amino-beta-carboxymuconate-epsilon-semialdehyde decarboxylase (ACMSD) was drastically reduced. The results strongly suggest that the hepatic NAD increase might be caused by transcription of genes coding the key enzymes of the Trp-NAD pathway via PPAR. PMID- 10721075 TI - Effect of albumin administration on L-tryptophan levels in the serum and tissues of rats with puromycin aminonucleoside-induced nephrosis. AB - We examined the effect of albumin administration on L-tryptophan (Trp) levels in the serum and tissues of rats with puromycin aminonucleoside (PAN)-induced nephrosis. PNA-injected rats showed increased urinary protein and serum non esterified fatty acids levels and decreased serum albumin level at 8 days after the injection. The nephrotic rats showed a decrease in total serum Trp level and increases in free serum Trp and liver and kidney Trp levels but no change in urinary Trp level. At 5 min after intravenous injection of bovine serum albumin, the nephrotic rats had serum albumin concentration similar to the level of non nephrotic rats and showed a recovery of decreased total serum Trp with the reduction of increased free serum Trp and liver Trp. PMID- 10721076 TI - The influence of kynurenine, neopterin, and norepinephrine on tubular epithelial cells and alveolar fibroblasts. AB - We have examined the effect of kynurenine, neopterin and norepinephrine on epithelial cells and fibroblasts: the total DNA synthesis (by adding 3H thymidine), the mitotic index, the amount of pathological mitosis and apoptosis. We have observed that kynurenine, neopterin and norepinephrine induce pathological mitosis and apoptosis of epithelial cells of kidney. The total DNA synthesis increases by the incubation of the epithelial cells with neopterin and norepinephrine. We have found an increase of pathological mitosis after addition of kynurenine, neopterin and norepinephrine to fibroblast cell culture. The total DNA synthesis is promoted by neopterin and norepinephrine, while kynurenine does not alter it. These data allow us to suggest that kynurenine, neopterin and norepinephrine promote epithelial cell damage leading to cell death through apoptosis, and can therefore be added to the factors, which promote the pathological mitosis of fibroblasts. PMID- 10721077 TI - Changes in membrane fluidity induced by tryptophan and its metabolites. AB - Incorporation of fatty acids into phospholipids as well as cholesterol and phospholipid concentration has been investigated using samples of rat liver homogenate, Krebs-Ringer-phosphate buffer (pH = 7.4), containing 0.3% albumin, fatty acid mixture and glycerol. The addition of kynurenine, kynurenic, xanthurenic, picolinic, and 5-hydroxyindoleacetic acids to incubation medium for phospholipid biosynthesis in vitro induced the elevation of cholesterol/phospholipid ratio, cholesterol concentration in samples, an increase of saturated and a decrease of polyunsaturated fatty acids, especially arachidonic acid incorporation into phospholipids. It allowed us to suggest that these metabolites of tryptophan can decrease the membrane fluidity, depress cell cycle, cell transformation and may stimulate cholesterol precipitation. The addition of tryptophan, 3-hydroxykynurenine, 3-hydroxyanthranilic, quinolinic, nicotinic acids, serotonin together with iproniasid, acetylserotonin, and melatonin to incubation medium for phospholipid biosynthesis in vitro induced an inverse relationship. Tryptophan and above mentioned metabolites decreased cholesterol/phospholipid ratio, cholesterol concentration in samples and incorporation of saturated fatty acids into phospholipids. The incorporation of polyunsaturated fatty acids, especially arachidonic acid increased. It allowed us to suggest that tryptophan and these metabolites, may increase membrane fluidity, stimulate cell cycle, cell transformation and can protect against cholesterol precipitation. PMID- 10721078 TI - Hormonal influences on tryptophan binding to rat hepatic nuclei. AB - We evaluated whether selected hormones, 3,5,3'-triiodothyronine (T3), hydrocortisone (HC) or insulin, would influence the binding (saturable, stereospecific, and of high affinity) of L-tryptophan to rat hepatic nuclei or nuclear envelopes. T3 (10(-14) to 10(-10) M) appreciably inhibited in vitro L-(5 3H) tryptophan binding to hepatic nuclei and T3 (10(-16) to 10(-4) M) appreciably ameliorated the inhibitory effect of unlabeled tryptophan (10(-4) M) on such binding. In vivo tryptophan administration (1 h) stimulated hepatic protein synthesis but the addition of T3 negated such stimulation. HC (10(-12) to 10(-4) M) did not affect and insulin (10(-16) to 10(-4) M) had only a small inhibitory effect on 3H-tryptophan binding to hepatic nuclei, but each (10(-12) to 10(-4) M) when added to unlabeled tryptophan (10(-4) M) diminished the inhibitory binding effect of unlabeled tryptophan alone. Thus, T3 competes with tryptophan for hepatic nuclear tryptophan binding and also negates tryptophan's stimulatory effect on hepatic protein synthesis. PMID- 10721079 TI - Melatonin and tryptophan derivatives as free radical scavengers and antioxidants. AB - Several tryptophan derivatives function as free radical scavengers and antioxidants. The molecule that has been most widely investigated in this regard is N-acetyl-5-methoxytryptamine (melatonin); however, pinoline (6-methoxy-1,2,3,4 tetrahydro-beta-carboline) and N-acetylserotonin also possess free radical scavenging activity. Experimental studies have shown that melatonin directly scavenges the hydroxy radical, peroxyl radical, peroxynitrite anion, and singlet oxygen. Furthermore, this tryptophan derivative stimulates a number of antioxidative enzymes and stabilizes cell membranes; this latter action helps membranes to resist free radical damage. While the antioxidative actions of most molecules are limited by their specific intracellular distribution, e.g., vitamin E in lipid-rich membranes, melatonin's antioxidative actions include the protection of lipids in the cell membrane, proteins in the cytosol, and DNA in the nucleus. Furthermore, melatonin crosses all morphophysiological barriers and enters equally well all cells in the organism. PMID- 10721081 TI - Radical scavenging properties of tryptophan metabolites. Estimation of their radical reactivity. AB - Radical scavenging properties of tryptophan metabolites were estimated using their radical reactivity. Metabolites of the kynurenine and the melatonin biosynthesis pathway were mainly examined by use of a kinetical model. Their radical reactivity was determined as the reaction rate constant with a stable free radical, such as galvinoxyl; that is a phenoxy radical. The rate constants of the metabolites have a widely ranged spectrum, which can be divided into three groups. The first group (3-hydroxykynurenine, 3-hydroxyanthranilic acid, and indole-3-pyruvic acid) is more reactive than alpha-tocopherol; the reactivity of the second group (xanthurenic acid, serotonin, N-acetylserotonin) is similar to that of butylated hydroxytoluene (BHT); the third group (kynurenic acid, melatonin, and other ones) is less reactive than BHT. PMID- 10721080 TI - Indole-3-pyruvic and -propionic acids, kynurenic acid, and related metabolites as luminophores and free-radical scavengers. AB - Chemiluminescence associated with oxidation by free radicals was investigated in an alkaline, hemin-catalysed hydrogen peroxide system, using the following tryptophan metabolites as radical scavengers: indole-3-pyruvic, indole-3 propionic, kynurenic, xanthurenic and quinaldic acids and 4-hydroxy-quinoline. Light emission from oxidation of the indolic compounds was only partially inhibited by the hydroxyl-radical scavenger DMSO, but strongly suppressed by the superoxide-anion scavenger Tiron, whereas chemi-luminescence generated from kynurenic acid was strongly inhibited by either of these compounds. Light emission from oxidation of kynurenic acid lasts for a surprisingly long period of time: At 0.4 mM and 20 degrees C, luminescence increased for 5 hours and continued at a high rate for more than a day. Comparison of structural analogues indicated that the 4-hydroxyl and carboxyl groups of kynurenic acid are essential for effective light emission, and that an additional 8-hydroxyl residue leading to an intramolecular hydrogen bond diminishes the reaction rate. PMID- 10721082 TI - Tryptanthrins and other tryptophan-derived agonists of the dioxin receptor. AB - 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) and related environmental pollutants exert most of their adverse effects via the aryl hydrocarbon or dioxin receptor (AhR). While most potent agonists of the AhR are of synthetic origin, an increasing number of natural compounds is now recognized as receptor agonists. Our findings demonstrate that some tryptanthrin derivatives biosynthesized in incubations of Candida lipolytica with tryptophan and anthranilic acid or its derivatives activate the AhR measured as induction of cytochrome P4501A1 mRNA and protein in rat hepatocytes in primary culture. The specificity of the inducing effect of tryptanthrins was demonstrated in gel retardation experiments in Hepa-1 mouse hepatoma cells using an oliogonucleotide comprising the sequence of the dioxin-responsive element. Furthermore, unidentified AhR agonists were formed in incubations of rat feces with a minimal medium supplemented with tryptophan. It is suggested that the receptor may be part of a defense system protecting higher organisms from secondary tryptophan-derived metabolites formed by the microflora of the host or its environment. PMID- 10721083 TI - Induction of cytochrome P4501A1 by ozone-oxidized tryptophan in Hepa lclc7 cells. AB - The present investigation was undertaken to determine whether administration of O3-oxidized amino acids to mouse hepatoma cells, Hepa lclc7 (Hepa-1), in culture would effect Cyp1a1 gene expression. The results demonstrate that, of all the amino acids tested, only O3-oxidized tryptophan caused a significant induction of CYP1A1-dependent 7-ethoxyresorufin O-deethylase (EROD) activity compared to the controls (p < 0.01). CYP1A1 mRNA and protein were markedly induced in the O3 oxidized tryptophan administered group compared to the controls. Gel mobility shift assays using nuclear extracts of Hepa-1 cells revealed that oxidized products of tryptophan can induce both aryl hydrocarbon receptor (AhR) transformation and binding of the liganded AhR complex to its specific DNA recognition site, thereby initiating transcription of the Cyp1a1 gene with concomitant increase of CYP1A1 protein and EROD activity in Hepa-1 cells. PMID- 10721084 TI - Cinnabarinic acid was formed in damaged mitochondria and its effect on mitochondrial respiration. AB - Tryptophan administration aggravates experimental mouse liver injury caused by carbon tetrachloride when 3-hydroxyanthranilic acid concentration elevates in serum. Tryptophan metabolism is changed by macrophages in injured liver. 3 Hydroxyanthranilic acid may be oxidized to cinnabarinic acid by injured mitochondria in the liver aggravating the state of injured liver. Mitochondria prepared from the liver 24 hr after CCl4 treatment have lost their ability of respiratory control. In consequence, 3-hydroxyanthranilic acid is oxidized to cinnabarinic acid by incubation with these mitochondria, whereas 3 hydroxykynurenine is not. Thus, formed cinnabarinic acid is able to inhibit completely the mitochondrial respiratory control at concentration of 10 microM. PMID- 10721085 TI - Effects of Fusarium mycotoxins on levels of serotonin, melatonin, and 5 hydroxytryptophan in pineal cell cultures. AB - Analysis of melatonin (MEL) in pineal cell cultures by enzyme linked immunosorbent assay showed its concentration was increased by fusaric acid (FA), a mycotoxin produced by Fusarium species and associated with toxic duck and ostrich feeds. Subsequent cell culture studies demonstrated the precursors of MEL, 5-hydroxytryptophan (5HTP) and serotonin (5HT), were also affected by FA as well as other Fusarium mycotoxins. Herein we describe a technique for the analysis of 5HTP and 5HT in pineal cell cultures using HPLC with electrochemical detection (EC), and report on the effects of FA alone and in combination with fumonisin B1 (FB1) and deoxynivalenol (DON) on the levels of these MEL precursors. PMID- 10721086 TI - Dioxin-induced perturbations in tryptophan homeostasis in laboratory animals. AB - Polychlorinated dioxins (PCDD) are widespread environmental contaminants. The most potent and the general model compound for dioxins is 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD). Our laboratory has developed a new model for studies of dioxin toxicity based on totally disparate sensitivity to the lethal action of TCDD between Long-Evans (L-E, Turku AB; LD50 ca. 10 micrograms/kg) and Han/Wistar (H/W, Kuopio; LD50 over 10,000 micrograms/kg) rat strains. We have shown that body weight regulation is differentially regulated by TCDD in these rat strains: body weight gain is permanently reduced in the sensitive L-E but not in the resistant H/W strain. In concert with reduced body weight, TCDD increased brain TRP concentration, 5-HT synthesis and its metabolism to 5-HIAA at lethal doses in TCDD-susceptible L-E rats, and almost not at all in resistant H/W rats in which lethal dose levels were not reached. Further studies showed that TCDD indirectly increases free TRP concentration in the circulation in TCDD susceptible L-E rats. Blood free fatty acids seem to be involved in the latter phenomenon. It is not likely that the enhanced serotonergic tone in the CNS is a causative factor in TCDD-induced anorexia. However, the present results may open up an interesting avenue to better understand physiology of TRP and the complex regulation of energy balance. PMID- 10721087 TI - From sugar beet molasses to Lyphan. Integrated quality management from the raw material to the drug. AB - L-tryptophan is produced at the AMINO GmbH (Frellstedt, FRG) via biocatalytical condensation of the amino acid L-serine with indole. As a biocatalyst, tryptophan synthetase is used which is produced in high activities by a natural mutant Escherichia coli strain. The enzyme mechanism and specificity and the individual process-parameters for the biotransformation procedure are explained as well as the purification process of educts and products. This includes a detailed description of the quality control of educts, intermediates and final product. The active ingredient L-tryptophan is subsequently used by AMINO's subsidiary company esparma GmbH to produce and distribute the pharmaceutical Lyphan. The quality management system and the production procedure for Lyphan are described and discussed. PMID- 10721088 TI - Structural characterization of case-associated contaminants peak C and FF in L tryptophan implicated in eosinophilia-myalgia syndrome. AB - We have characterized the structures of two case-associated contaminants of the Showa Denko L-tryptophan known to cause eosinophilia-myalgia syndrome (EMS). A combination of on-line accurate mass HPLC-electrospray ionization-mass spectrometry (LC-MS), HPLC-tandem mass spectrometry (LC-MS-MS) and HPLC-in source collision induced dissociation-MS-MS (LC-sCID-MS-MS) allowed the structure determination of both Peak C and FF. Peak C is identified as 3a-hydroxy 1,2,3,3a,8,8a-hexahydropyrrolo-[2-3b]-indole-2-carboxyl ic acid, whereas Peak FF is characterized as 2-(2-hydroxy-indoline)-tryptophan. Both contaminants contain indoline rings, and the significance of this finding is discussed. PMID- 10721089 TI - Eosinophilia-myalgia syndrome case-associated contaminants in commercially available 5-hydroxytryptophan. AB - Recently, 5-hydroxy-L-tryptophan (5-OHTrp) has been promoted as an alternative to banned L-tryptophan as a dietary supplement. It has been claimed to help alleviate obesity, insomnia, depression, and headaches. However, eosinophilia myalgia syndrome (EMS)-like symptoms have also been associated with ingestion or exposure to 5-OHTrp. HPLC-UV analysis of EMS-implicated 5-OHTrp revealed the presence of peak X, described as case-implicated. We show that peak X is actually a family of contaminants with the same molecular weight (234 Da) and similar HPLC retention times. We also demonstrate that all eight samples of commercially available 5-OHTrp analyzed by HPLC-MS contained three or more contaminants of the peak X family. The significance of these findings is discussed. PMID- 10721090 TI - Synthesis, formation, and occurrence of contaminants in biotechnologically manufactured L-tryptophan. AB - The pattern of contaminants in pharmaceutical and feed grade L-tryptophan (Trp) was investigated in a market survey of 22 lots of 6 different manufacturers. To date, 5 case associated contaminants in Showa Denko tryptophan (SD-Trp) known to cause the autoimmune disease eosinophilia-myalgia syndrome (EMS) have been structurally elucidated: 3a-hydroxy-1,2,3,3a,8,8a-hexahydropyrroloindole-2 carboxylic acid (PIC), an indoline compound, is one of the most abundant degradation compounds of unbound Trp during oxidative treatment. 2-(3 indolylmethyl)-L-tryptophan (IMT) and 2-(2-hydroxyindoline)-tryptophan (HIT) are both 2-substituted Trp-derivatives. IMT was synthesized by the reaction of Trp and indole-3-methanol or indole-3-acetaldehyde, respectively. From this finding it is proposed that Trp-metabolites can decompose under formation of transitional, mesomerism-stabilized cations that react with excess Trp to yield 2 substituted Trp derivatives. The decomposition of Trp-metabolites could be induced by elevated or low pH-values that occur during the downstream processing of the Trp fermentation broth. IMT was detected in pharmaceutical-grade and feed grade Trp in amounts of < 20-1,400 mg/kg. 1,1'-Ethylidenebis-(L-tryptophan) (EBT) is formed from acetaldehyde and Trp under acidic conditions and serves as a marker for EMS-suspicious Trp. 3-(Phenylamino)alanine (PAA) is the only not Trp derived case associated contaminant. Low amounts of PAA (20 mg/kg) could be detected in feed-grade Trp of one manufacturer. Non-EMS correlated 1,2,3,4 tetrahydro-beta-carboline-3-carboxylic acids of Trp and formaldehyde, acetaldehyde and indole-3-acetaldehyde could be detected in the examined Trp raw materials (< 10-13,500 mg/kg). In order to guarantee the safety of Trp containing drugs the amount of EBT (< 10 mg/kg Trp) and the sum of UV220 nm detectable contaminants (< 400 mg/kg Trp) are limited by the European authorities. PMID- 10721091 TI - Synthesis and formation of an EMS correlated contaminant in biotechnologically manufactured L-tryptophan. AB - Contaminants in biotechnologically manufactured L-Tryptophan (Trp) are suspected to be responsible for the outbreak of an unknown autoimmune disease in 1989. The contaminants, found in Trp-lots of a Japanese manufacturer, are classified in EMS correlated and non EMS-correlated substances. Up to now six EMS-correlated substances are known. One of these compounds is 2-(3'-indolylmethyl)-indole (IMT). IMT was detected as a major contaminant in two investigated EMS-associated trp-samples. In a seven step chemical synthesis IMT was obtained for use as a reference substance. A model system to investigate the formation of IMT was created using Trp and 3-indolylmethanol (IM). IMT formation was observed at acidic and alkaline pH-values and the optimal molar ratio of Trp to IM is 100:1. In addition an IMT formation was observed from indole, formaldehyde and Trp as well as from Trp and 3-indolylacetaldehyde. PMID- 10721092 TI - Is there any relationship between eosinophilia myalgia syndrome (EMS) and fibromyalgia syndrome (FMS)? An analysis of clinical and immunological data. AB - The eosinophilia-myalgia syndrome (EMS) caused by intake of contaminated L tryptophan resembles in its clinical presentation the fibromyalgia syndrome (FMS). We, therefore, analysed clinical and immunological parameters in 16 patients with chronic EMS and 100 patients with FMS in order to see whether there may be a relationship between both disorders. From 12 FMS patients and 12 controls also peripheral blood mononuclear cells (PBMC) were obtained. Myalgia and arthralgia was observed in chronic EMS in the same incidence as in patients with FMS (81%). Also antibodies to serotonin, gangliosides and phospholipids were present in both groups. In vitro stimulation of PBMC with different L-tryptophan preparations revealed in six of the 12 FMS patients but only two of the control individuals a production of type 2 cytokines (IL-5, IL-10). We, therefore, conclude that EMS may have developed in patients suffering primarily from FMS as an allergic reaction towards a more immunogenic L-tryptophan preparation. PMID- 10721093 TI - Oxidative damage in rat tissue following excessive L-tryptophan and atherogenic diets. AB - Numerous reports were published on the connection between diets containing excessive L-tryptophan and the development of Eosinophilia Myalgia Syndrome. It has been also demonstrated that some cell functions depend on fatty acid composition which can result in increased lipid peroxidation in cells such as macrophages and other inflammatory cells. The purpose of the present study was to investigate the combined effects of an atherogenic diet enriched with tryptophan on lipid peroxidation in rats. 3-week-old CD-1 female rats were fed (3 weeks) control or atherogenic diets and the same diets supplemented with 0.4% or 1.0% L tryptophan. Liver and skeletal muscle samples from all groups were taken for histology, autoradiography and for determination of lipid peroxidation. Infiltration of cells into fascia of muscle was observed following tryptophan or atherogenic diet consumption. However, no change in 3H-thymidine incorporation into DNA was observed by autoradiography. A significant increase of lipid peroxidation was detected in muscle following consumption of L-tryptophan-rich diets, with no significant difference from control in animals treated with atherogenic diets. Contrastly, a reduced lipid peroxidation was detected in liver of animals treated with excessive tryptophan as well as in animals fed on L tryptophan and atherogenic diet or atherogenic diet alone. Our results indicated that excessive dietary tryptophan, when consumed with an atherogenic diet, increased lipid peroxidation in muscle but not in liver. Consumption of these feeding diets with or without supplementation of tryptophan resulted in reduced lipid peroxidation in muscle as well as in liver. PMID- 10721094 TI - Tryptophan toxicity--time and dose response in rats. AB - During the past decade L-tryptophan (Trp) ingestion have been associated with a multisystemic syndrome, known as eosinophilia myalgia syndrome (EMS). Even though an epidemic studies indicated that a contaminant, 1,1'-ethylidene-bis-L tryptophan was involved in EMS, abnormalities in metabolism of Trp have been reported in other similar clinical syndromes such as carcinoid syndrome, scleroderma or eosinophilic fasciitis. The purpose of the study was to investigate the role of Trp or its metabolite, given in different dosing regimens in induction of tissue damage. METHOD: 3 months old female rats (Charles River CD 1) were fed for 3, 6, 12 weeks on a diet containing 20% protein diet derived from casein and supplemented with 1%, 2%, or 5% Trp. On the last week of feeding, half of the animals fed on a control diet and half of the animals fed on the Trp diet were injected with 2 injections of para-chlorophenyl alanine (p-CPA), a Trp hydroxylase inhibitor, 300 mg/kg i.p. followed by 3 injection of 100 mg/kg every alternate day. RESULTS: Body weight of rats fed higher levels of Trp increased slowly and injection of p-CPA induced loss in body weight. 2/6 of the animals treated with 1% Trp and 1/6 treated with 5% Trp for 3 weeks and 2/4 animals treated with 1% Trp and 1/4 treated with 5% Trp for 12 weeks died after injection of p-CPA. No mortality was detected in 1-5% Trp treated animals. Alopecia and skin changes were seen after p-CPA in 1-5% Trp treated animals. Increased amounts of connective tissue and induction of inflammatory cell proliferation were observed in lung, spleen and in gastrocnemia muscle of rats treated with higher dose of Trp for longer period. Induction of kynurenine pathway by injection of p CPA caused more tissue damage. It is concluded that excessive Trp or elevation of its metabolites could play a role in amplifying some of pathological features of EMS. This pathological damage is further augmented by metabolites of the kynurenine pathway. PMID- 10721095 TI - IFN-gamma activated indoleamine 2,3-dioxygenase activity in human cells is an antiparasitic and an antibacterial effector mechanism. AB - In nearly all human cells IFN-gamma stimulation leads to an activation of indoleamine 2,3-dioxygenase (IDO) activity, which is responsible for anti toxoplasma and anti-chlamydia effects. We have recently shown that IDO activation is also a defense mechanism against extracellular beta-hemolytic streptococci groups A, B, C and G in human glioblastoma cells, fibroblasts and macrophages. Similar effects were also seen with enterococci and in approximately 65% of staphylococci tested, including multiresistant strains of both species. In addition, we have found that IDO activity is differentially regulated in different cells. For example we have found that TNF-alpha enhances IFN-gamma induced IDO activity and antimicrobial effect in human glioblastoma cells whereas both IFN-gamma mediated effects were blocked by TNF-alpha as well as by IL-1 in a human uroepithelial cell line. We were able to show that the IL-1 and TNF-alpha mediated inhibition of IFN-gamma-induced IDO activity in uroepithelial cells is due to stimulation of inducible nitric oxide synthase. In human astrocytoma cells, IL-1 and TNF-alpha did not inhibit IDO activity and in concordance with this finding these cells did not show a detectable nitric oxide production. PMID- 10721096 TI - Importance of L-tryptophan metabolism in trypanosomiasis. AB - African trypanosomiasis or sleeping sickness is caused by extracellular trypanosomes. The presence of seric antibodies directed to a tryptophan-like epitope in trypanosome infected patients and animals led us to investigate the roles of tryptophan in trypanosomiasis. These antibodies are directed against a tryptophan-rich conserved sequence inside the major parasite surface glycoprotein. In vitro, a rapid uptake of tryptophan by trypanosomes is measured. Seric tryptophan levels are decreased during trypanosomiasis. This decrease may be linked with an increase in indoleamine 2,3-dioxygenase (IDO) induced by Interferon-gamma. In vivo inhibition of IDO by norharman provokes a dramatic increase in circulating parasite number. All these data show the essential role of tryptophan in parasite growth. Moreover, antibodies against tryptophan, the decreased concentration of the neurotransmitter serotonin in the brain following infection and the tryptophan metabolites (tryptophol) produced by trypanosomes may participate to the pathophysiological mechanisms provoking sleeping sickness. PMID- 10721097 TI - Cytokine mediated regulation of interferon-gamma-induced IDO activation. AB - Stimulation of human monocyte-derived-macrophages (MDM) with interferon gamma induces the L-tryptophan degrading enzyme indoleamine 2,3-dioxygenase (IDO). It has been well documented that the growth of some intra-cellular parasites such as Chlamydia and Toxoplasma in human fibroblasts and glioblastoma cells is inhibited by IDO mediated L-tryptophan depletion. We have recently shown that IDO induction in cord blood MDM is also responsible for the growth inhibition of extra-cellular group B streptococci and thus for the first time shown an anti-bacterial effect of IDO activation. In view of this immunological function we sought to investigate the regulation, and in particular the downregulation of IDO by the immune system. We describe here the effect of cytokines on IDO activation and in particular the inhibitory function of IL-10, TGF beta and IL-4. PMID- 10721098 TI - Antioxidant activities and redox regulation of interferon-gamma-induced tryptophan metabolism in human monocytes and macrophages. AB - This article summarises studies supporting the proposal that induction of L tryptophan (Trp) degradation along the kynurenine pathway in human monocytes and macrophages by interferon-gamma (IFN gamma) represents a novel extracellular antioxidant defence that acts to prevent inadvertent oxidative damage to host tissue during inflammation. The studies show that formation and release of the aminophenolic antioxidant 3-hydroxyanthranilic acid (3-HAA) is responsible for the ability of IFN gamma-primed human macrophages to inhibit the oxidation of low density lipoprotein (LDL); an event implicated as an important event in atherogenesis. 3-HAA efficiently inhibits LDL oxidation by acting as an aqueous oxidant scavenger and a synergist for LDL-associated vitamin E. Indoleamine 2,3 dioxygenase activity (IDO) is the initial and rate limiting enzyme of Trp degradation along the kynurenine pathway. Nitric oxide inhibits IDO activity in IFN gamma-primed human macrophages and this may represent a physiological regulatory mechanism of the dioxygenase during inflammation. PMID- 10721099 TI - Interferon-gamma-dependent/independent expression of indoleamine 2,3-dioxygenase. Studies with interferon-gamma-knockout mice. AB - The role of IFN-gamma in the expression of indoleamine 2,3-dioxygenase (IDO), a tryptophan oxidizing enzyme, in mouse tissues under physiological and pathological conditions was investigated using IFN-gamma-knockout mice. The results revealed that i) the expression of IDO in the large intestine or in the cecum is mediated by IFN-gamma, ii) for the systemic IDO induction under endotoxin shock, IFN-gamma is a dominant inducer but not essential, and an IFN gamma-independent mechanism is also operative, iii) the systemic induction of IDO caused by IL-12 or Pokeweed mitogen is mediated by IFN-gamma, and iv) the constitutive IDO expression in the epididymis is IFN-gamma-independent. PMID- 10721100 TI - L-tryptophan-kynurenine pathway metabolite 3-hydroxyanthranilic acid induces apoptosis in macrophage-derived cells under pathophysiological conditions. AB - Accumulation of L-kynurenine and 3-hydroxyanthranilic acid (3HAA) occurs in the monocyte-derived cells following immune stimulation, and may derive from L tryptophan following induction of indoleamine-2,3-dioxygenase. In the present study, we evaluate the possibility that 3HAA acts as an endogenous inducer of monocyte/macrophage apoptosis. Supplementation with 200 microM of 3HAA, but not other L-tryptophan metabolites tested, significantly increased the number of apoptotic cells in both THP-1 and U937 cells. Catalase, superoxide dismutase and manganese ions markedly enhanced apoptosis in the presence of 3HAA in these cells. The present results suggest that 3HAA induces the macrophage/monocyte apoptosis under certain conditions, which may be relevant to pathophysiology of inflammatory conditions. PMID- 10721101 TI - Tryptophan catabolism in synovial fluid of various arthropathies and its relationship with inflammatory cytokines. AB - Synovial fluids (SF) from patients with rheumatoid arthritis (RA), psoriatic arthritis (PsA), gout, and osteoarthritis (OA) were investigated for the levels of interleukin (IL)-1 beta, IL-6 and IL-8, tryptophan (Trp) and indoleamine 2,3 dioxygenase (IDO) activity. Significant differences exist in the levels of IL-1 beta between inflammatory arthritides RA, PsA and gout and non inflammatory arthritis, such as OA. The highest concentration of IL-1 beta was found in RA, that showed high levels also of IL-6 and IL-8. In the same disease we also found the highest IDO activity and the lowest Trp concentration. In addition, IDO activity seems to be related with the decrease in Trp, as demonstrated by the inverse correlation found between these two substances in the SF of all patients. PMID- 10721102 TI - Degradation of tryptophan in patients with systemic lupus erythematosus. AB - Systemic lupus erythematosus (SLE), a progressive autoimmune disorder, is associated with chronic stimulation of various components of the immune system. Since cell-mediated immunity is also activated, we were interested to test for abnormalities in tryptophan metabolism in SLE which may result from activation of indoleamine 2,3-dioxygenase by cytokines released during the immune response. We measured serum tryptophan and kynurenine concentrations in 52 patients with SLE as well as serum neopterin as an indicator for the degree of immune activation. Compared to controls, we found significantly decreased tryptophan and increased kynurenine concentrations in SLE. The extent of tryptophan catabolism correlates with neopterin concentrations or with the disease activity index. Tryptophan depletion may be associated with neurologic/psychiatric disturbances in patients suffering from SLE. PMID- 10721104 TI - Structure, function, and applications of tryptophan tryptophylquinone enzymes. AB - Tryptophan and tyrosine residues in proteins may be posttranslationally modified to form enzyme cofactors. Tryptophan tryptophylquinone (TTQ), the cofactor of methylamine dehydrogenase (MADH), is formed by covalent cross-linking of two tryptophan residues and incorporation of two oxygen atoms into one of the indole rings to form a quinone. MADH converts primary amines to their corresponding aldehydes plus ammonia. During the catalytic cycle, TTQ mediates electron transfer from substrate to a copper protein, amicyanin. These electrons are transferred to the respiratory chain via a c-type cytochrome. Structural, kinetic and site-directed mutagenesis studies have characterized protein-protein interactions, and mechanisms of catalysis and electron transfer by TTQ. Preliminary results obtained with MADH enzyme-electrodes demonstrate the potential for quinoprotein-based biosensors. PMID- 10721105 TI - An unusual role of tryptophan in PQQ-containing quinoproteins. AB - Methanol dehydrogenase is a bacterial quinoprotein containing PQQ at its active site, which is in the centre of a superbarrel structure made up of eight beta sheets arranged radially as the blades of a propeller. A series of novel tryptophan-docking interactions between the beta-sheets make planar, stabilizing girdles around the periphery of the protein. The tryptophan residues form stacking interactions, and hydrogen bonds through their indole ring NH groups. PMID- 10721106 TI - Specific enzymatic chlorination of tryptophan and tryptophan derivatives. AB - In the search for an alternative to chemical halogenation reactions using the free halogens, a novel type of halogenating enzymes was detected. In contrast to haloperoxidases, these NADH-dependent halogenases are specific. Tryptophan halogenase which catalyses the regioselective chlorination of tryptophan to 7 chlorotryptophan can also chlorinate tryptamine, tryptophol, indole-3 acetonitrile, and 3-methylindole. However, indole-3-acetonitrile is not chlorinated in the 7-position, but in positions two and three of the indole ring. Chlorination in the 3-position is obviously stereospecific. In addition to tryptophan and indole derivatives, aminophenylpyrrole is also accepted as a substrate for regioselective chlorination. Since the new NADH-dependent halogenases have a fairly broad substrate specificity and catalyse regioselective chlorination reactions they could be a good alternative to chemical halogenation. PMID- 10721103 TI - Kynurenine and neopterin in chronic glomerulonephritis. AB - The results of our clinical observations of 102 patients with chronic glomerulonephritis with normal renal function have shown that hyperkynureninemia in 22.5% of patients develops in cases of pyridoxal-5-phosphate deficiency (hyperkynureninemia after peroral L-tryptophan load), but in 14.8% of patients through the stimulation of the cellular immune system (hyperkynureninemia at fasting state, increase of serum neopterin concentration). In all 20 patients with chronic renal failure hyperkynureninemia develops due to decreased renal function (increased serum kynurenine, neopterin and creatinine concentrations). Therefore, L-tryptophan peroral loading test with the determination of serum concentration of kynurenine before and at 3rd hour after the load, as well as the detection of serum concentration of neopterin and creatinine are helpful for the differentiation of following pathologies in patients with chronic glomerulonephritis: pyridoxal-5-phosphate deficiency, cellular immune stimulation and chronic renal failure. PMID- 10721107 TI - Characterization and functional expression of the cDNA encoding human brain quinolinate phosphoribosyltransferase. AB - To elucidate the molecular basis underlying quinolinate metabolism, the cDNA encoding human brain QPRTase was cloned. PMID- 10721108 TI - Identification of cDNAs encoding alpha-amino-beta-carboxymuconate-epsilon semialdehyde decarboxylase (ACMSDase). AB - To elucidate the molecular basis underlying quinolinate metabolism, the cDNA encoding ACMSDase was cloned. PMID- 10721109 TI - Purification of L-kynurenine 3-monooxygenase from mitochondrial outer membrane of pig liver. AB - The aim of the present work is to obtain the homogeneous L-kynurenine 3 monooxygenase (Fpk) enzyme preparation by a simple and rapid immunoaffinity purification method. Fpk was purified by monoclonal antibody (mAb) immunoabsorbent column. The column was prepared using hydrazide-activated agarose beads (Affi-Gel Hz) to which IgG molecules were coupled via carbohydrate moieties located on the Fc region and peroxidized with periodate. Partially purified Fpk was charged on the column and after washing the column with buffer containing 0.5 M NaCl and 0.5% Triton-X-100 and then with buffer alone, the enzyme was eluted with acidic elution buffer. Despite the loss of the catalytic activity due to the acidic elution, the immunoaffinity preparation may be useful for the analysis of the chemical structure of Fpk and for the production of the polyclonal antibody toward Fpk. PMID- 10721111 TI - Liver and kidney kynurenine aminotransferase activity in different strains of rats. AB - Variations in liver and kidney kynurenine aminotransferase (KAT) activity in Pittsburg-Yoshida, Brown-Norway, albino Wistar, Sprague-Dawley, Long Evans and heterozygous Gunn rats were studied. In liver, values of KAT specific activity, expressed as mumoles of kynurenic acid formed per hour per mg of protein, were different in the groups of Brown-Norway and Pittsburg-Yoshida rats versus Long Evans and Sprague-Dawley rats. The activity expressed as mumoles of kynurenic acid per g of fresh liver showed other differences, being significantly higher in Gunn with respect to other strains of rats and lower in Pittsburg-Yoshida and Brown-Norway rats. In addition, KAT activity was significantly lower in Pittsburg Yoshida than in Brown-Norway rats. In kidney, the specific activity of kynurenine aminotransferase showed significant differences in the values of Sprague-Dawley and Long Evans rats versus the other strains. The activity expressed per g of fresh tissue was significantly higher in Wistar, Sprague-Dawley, Long Evans and Gunn than in Pittsburg-Yoshida and Brown-Norway rats. No significant differences were found in values between hyperlipidemic Pittsburg-Yoshida and their control Brown-Norway rats. These results high-light the importance of considering various rat strains when inbred animal experimental models are used. PMID- 10721110 TI - Quantification of anthranilic acid and its related enzyme activity in several different species. AB - Anthranilic acid (AA) has been attracted considerable attention as one of the L tryptophan-kynurenine pathway metabolites in the central nervous system. In this study, the concentration of L-kynurenine (L-KYN) and AA in serum and CSF, and its related enzyme activities were determined in several species. In rabbits, CSF AA concentrations were lower and serum AA concentrations were slightly higher than those in other species. However, the concentrations of L-KYN were substantially higher in rabbits in both serum and CSF compared with other species. Tissue enzyme activities varied among species. In rabbits, lung IDO activities were higher, but liver kynurenine 3-hydroxylase activities were lower than those of the other species tested. Furthermore, brain kynurenine 3-hydroxylase activities were higher in gerbils than those in other species. These results clearly demonstrated that kynurenine pathway enzyme activities and metabolite concentrations vary with species. PMID- 10721112 TI - Latest on enzymology of serotonin biosynthesis in walnut seeds. AB - Serotonin (5-HT) accumulation in walnut cotyledons is seen as a detoxification mechanism protecting the sensitive plant tissues of seeds from highly toxic ammonia concentrations following seed desiccation. Different metabolic pathways and cell compartments are involved in biosynthesis and storage of 5-HT. Ammonia fixation and incorporation into the indole moiety of tryptophan is followed by 5 HT biosynthesis via tryptamine in a two-step pathway with the adaptive tryptophan decarboxylase and the constitutive tryptamine 5-hydroxylase. Evidence is provided that tryptamine 5-hydroxylase is a member of the cytochrome P450 family which is involved in lipid hydroxylation processes in the very early period of seed development. PMID- 10721113 TI - Ommochrome genesis in an albino strain of a terrestrial isopod. AB - The contents of tryptophan (Trp) metabolites and the activities of the enzymes involved in ommochrome biosynthesis were measured in an albino strain of a terrestrial isopod Armadillidium vulgare. There was little difference between the Trp content in the albino mutant and that in the wild type, although the contents of 3-hydroxykynurenine (3-OH-Kyn), 3-hydroxyanthranilic acid (3-OH-AA) and xanthommatin in the albino were significantly lower than those in the wild type. Tryptophan 2,3-dioxygenase (TDO) activity in the albino was extremely low, while the activities of Kyn-3-hydroxylase and kynureninase did not differ significantly between the two phenotypes. The extremely low activity of TDO is probably one of main reasons why almost no ommochrome pigment is produced in the albino mutant. PMID- 10721114 TI - Involvement of tryptophan metabolism in the body color of crustacea. AB - The terrestrial isopod, Armadillidium vulgare is usually grey or black in color, however, red ones are occasionally found in the field. This is caused by the mutation of the ommochrome genesis in the integument. We focused our experiments on the mechanism of pigment genesis in which tryptophan metabolism had been expected to be different from the grey or black wild types. We obtained the result that 3-hydroxyanthranilic acid content was significantly higher in the red phenotype than in the wild type, and kynureninase activity was also higher in the red phenotype. PMID- 10721115 TI - Correlation between tryptophan and hair pigmentation in human hair. AB - The concentration of tryptophan in human hair of various colours is determined in order to study their correlation with hair pigmentation. The mean levels of this amino acid in hair samples are higher in men than in women. Therefore, sex influences the content of tryptophan in human hair. In addition, age influences the distribution, the highest levels are observed in the 1-5 year age-group and in ageing subjects in the groups up to 61-80 years in both sexes. The hair samples subdivided, according the colour, into blond, dark blond, red, light brown, brown, black, grey, and white demonstrate that in both sexes the concentrations of tryptophan are higher in brown and black hair than in blond hair. However, the tryptophan levels are highest in grey and white hair, showing that tryptophan accumulates among hair fibres with age. Therefore, there is a correlation between tryptophan content and hair pigmentation. PMID- 10721116 TI - Off-flavor compounds in wine and other food products formed by enzymatical, physical, and chemical degradation of tryptophan and its metabolites. AB - Tryptophan (TRP) and its metabolites are considered as potential precursors of 2 aminoacetophenone (AAP) in different food products causing different off-flavors. AAP is also responsible for the "untypical aging flavor (UTA)" in wine, developing a floor polish-like flavor in white wines within a few months of storage. In this study the formation of AAP was elucidated by GC-MS analysis of volatile components in model systems, grape musts and wines, spiked with TRP and different TRP metabolites like indole-3-acetic acid (IAA) and sulfite. In sulfurized wines and model solutions which were stored at different temperatures (20 degrees C, 45 degrees C) formylaminoacetophenone (FAP) and AAP were formed mainly from IAA with formation rates up to 20 mole%. Minor formation rates of AAP (< 1 mole%) were found in sulfurized solutions of TRP, indole-3-lactic acid, and indole-3-pyruvic acid. The results showed that the formation of AAP in wine can be referred to an oxidative degradation of IAA by superoxide- and hydroxyl radicals, which can be formed in wine after the sulfuration by cooxidation of sulfite to sulfate. After decarboxylation, pyrrole oxidation, and ring cleavage, FAP was the main volatile compound of the nonenzymatic degradation of IAA by sulfite which was quantitatively hydrolyzed to AAP. The formation of AAP and FAP was significantly lower in white wines than in ethanolic solutions spiked with IAA. However AAP formation rates of up to 5 mole% were still enough for an UTA. Due to the fact that the AAP- and UTA-formation by cooxidation of sulfite and IAA was completely blocked in red wines, it could be deduced that polyphenolic compounds, typical for red wines, have a scavenger effect on the radical oxidation of sulfite. Possibilities for an inhibition of the IAA degradation during winemaking to avoid the UTA in white wines by addition of radical scavengers like grape marc or ascorbic acid are discussed. PMID- 10721117 TI - Determination of tryptophan and tryptophan metabolites in grape must and wine. AB - Tryptophan (Trp) and its metabolites, especially indole-3-acetic acid (IAA), are considered as potential precursors of 2-aminoacetophenone (AAP), an aroma compound which causes the "untypical aging off-flavor" (UTA) in Vitis vinifera white wines. In this study RP-HPLC with fluorescence detection was used for the qualitative and quantitative analysis of Trp and Trp-metabolites in 39 grapes, 22 grape musts and 16 wines, to which different viticultural conditions (ripeness, pruning, strip of leaves, soil condition) have been applied. A sensitive and selective determination was achieved after solid phase extraction using an anion exchange material. Only traces of Trp-metabolites could be determined in the examined grapes and grape musts, but their amounts increased significantly during fermentation, whereas the amount of Trp decreased. Different viticultural measures, besides the time of grape harvest, showed no significant influences on the amount of Trp and Trp-metabolites. PMID- 10721118 TI - Influences on skatole formation from tryptophan in the pig colon. AB - Variable amounts of skatole (3-methyl-indole) are formed by microbes out of tryptophan in the colon of pigs. It is resorbed and accumulated in fat, leading to a fecal odor of the meat. We investigated the mechanisms by which differences in diet composition lead to variations in skatole concentrations in blood plasma and fat. The experiments were based on the hypothesis that tryptophan is derived from mucosa cell debris from the small intestine. It was found, that gut cell mitosis is stimulated by the growth factor IGF-I. This factor increases when energy in the diet is high. In addition, the mitotic rate is elevated when high amounts of purines are available in the diet, allowing a more rapid DNA- and RNA synthesis. Thus, high energy combined with high purines in the diet lead to a remarkable increase of gut cell mitosis which is accompanied by an increase of apoptosis. These apoptotic cells ultimately provide the substrate for skatole formation. In consequence, a dramatically rise of skatole in blood plasma and fat was measurable. PMID- 10721119 TI - Determination of hydroxytryptophan isomers in gamma-irradiated egg white, chicken meat, and shrimps. AB - The radiation induced products of tryptophan (TRP) were determined in gamma irradiated egg white, chicken meat and shrimps using RP-HPLC and electrochemical detection. A two-step hydrolysis with proteinase K and carboxypeptidase A was developed to release the radiation products from egg white and chicken meat and with proteinase K and pronase E from shrimps. The four hydroxytryptophan isomers (OH-Trp) were identified and quantified as radiation products in all samples. The amounts ranged between 0.02 and 1.97 mg/kg protein. A significant difference between irradiated and unirradiated samples was found for irradiation doses of more than 3 kGy for egg white and chicken meat. For shrimps no significant increase of OH-Trp isomers was measured up to a radiation dose of 5 kGy. PMID- 10721120 TI - Formation of the comutagenic beta-carboline norharman in a simple tryptophan containing model system at low temperature (40 degrees C-80 degrees C). AB - The formation of the comutagenic tryptophan derivate norharman in heat-processed food is a well known phenomenon. As a first step to investigate the possible formation of norharman during food storage particularly in regard to fortified food we developed a model system and studied the non-enzymatic formation of the comutagen at temperatures below 100 degrees C. With standard conditions (4 h, 80 degrees C) iron and copper (Fe2+/Cu2+) was required for the formation of norharman. Addition of glucose to the reaction mixture increased the norharman formation and a plateau was reached with 400 microM tryptophan. Decreasing the reaction temperature to 40 degrees C led to a significant formation of norharman after 72 hours of incubation indicating that the formation of norharman can take place at temperatures that are close to ranges occurring during food storage. PMID- 10721121 TI - The role of tryptophan in fatigue in different conditions of stress. AB - Tryptophan is the precursor for the neurotransmitter 5-hydroxytryptamine (5-HT), which is involved in fatigue and sleep. It is present in bound and free from in the blood, where the concentration is controlled by albumin binding to tryptophan. An increase in plasma free tryptophan leads to an increased rate of entry of tryptophan into the brain. This should lead to a higher level of 5-HT which may cause central fatigue. Central fatigue is implicated in clinical conditions such as chronic fatigue syndrome and post-operative fatigue. Increased plasma free tryptophan leads to an increase in the plasma concentration ratio of free tryptophan to the branched chain amino acids (BCAA) which compete with tryptophan for entry into the brain across the blood-brain barrier. The plasma concentrations of these amino acids were measured in chronic fatigue syndrome patients (CFS) before and after exercise (Castell et al., 1998), and in patients undergoing major surgery (Yamamoto et al., 1997). In the CFS patients, the pre exercise concentration of plasma free tryptophan was higher than in controls (p < 0.05) but did not change during or after exercise. This might indicate an abnormally high level of brain 5-HT in CFS patients leading to persistent fatigue. In the control group, plasma free tryptophan was increased after maximal exercise (p < 0.001), returning towards baseline levels 60 min later. The apparent failure of the CFS patients to change the plasma free tryptophan concentration or the free tryptophan/BCAA ratio during exercise may indicate increased sensitivity of brain 5-HT receptors, as has been demonstrated in other studies (Cleare et al., 1995). In post-operative recovery after major surgery plasma free tryptophan concentrations were markedly increased compared with baseline levels; the plasma free tryptophan/BCAA concentration ratio was also increased after surgery. Plasma albumin concentrations were decreased after surgery: this may account for the increase in plasma free tryptophan levels. Provision of BCAA has improved mental performance in athletes after endurance exercise (Blomstrand et al., 1995, 1997). It is suggested that BCAA supplementation may help to counteract the effects of an increase in plasma free tryptophan, and may thus improve the status of patients during or after some clinically stressful conditions. PMID- 10721122 TI - The significance of tryptophan in infant nutrition. AB - In the newborn, tryptophan (Trp) and its metabolites are essential to brain maturation and to the development of neurobehavioral regulations of food intake, satiation and sleep-wake-rhythm. Due to the high Trp concentration in human milk in relation to the total of neutral amino acids, the blood-brain transfer of tryptophan as a precursor of its metabolites serotonin and melatonin is optimal. In contrast, commercial infant formulas are lower in Trp and higher in neutral amino acid levels resulting in comparatively lower Trp serum concentrations. alpha-lactalbumin enriched, protein-reduced formulas adapted to 2.2% Trp were shown to be capable of producing Trp serum values that did not differ from those in breast-fed infants. PMID- 10721123 TI - Nutritional factors that regulate on the conversion of L-tryptophan to niacin. AB - Niacin is biosynthesized from L-tryptophan. As a lot of L-tryptophan exists in the body, the pathway is very important because niacin can be efficiently supplied even when the body emergently needs niacin. Therefore, it is very important to know factors affecting the conversion ratio of L-tryptophan to niacin. The conversion ratio is decreased with increasing dietary protein levels. In the effects of fat, feeding of diets containing unsaturated fatty acids increased the conversion ratio, while feeding of diets saturated fatty acids did not. In the effects of carbohydrate, the conversion ratio was higher in diets containing starch than in diets containing sucrose. PMID- 10721124 TI - Kynurenine concentration of serum was increased by exercise. AB - Kynurenine pathway of tryptophan makes a lot of physiological active substances, such as quinolinate, NAD and so on, suggesting that kynurenine itself may play a very important role physiologically. Therefore, we examined the influence of exercise on serum kynurenine concentration. At first, we assayed kynurenine concentration of students (n = 13) who took part in a rugby camp for three days. The mean value of kynurenine concentration of before and after training were 1.362 +/- 0.306 microM and 1.725 +/- 0.511 microM respectively. These data means that severe exercise rise the serum kynurenine concentration. Then we tried to examine the relationship between the level of exercise and serum kynurenine concentration. Serum kynurenine concentration had significantly increased immediately after the exercise from 1.869 +/- 0.285 microM to 2.138 +/- 0.248 microM of 24 hours later by loading of 65% heart rate max exercise for each subject. These results suggested that at least the severe exercise affect on the tryptophan metabolism. We will discuss the change of serum kynurenine concentration by another sports such as soccer game and 20 km run. PMID- 10721125 TI - Absorption of trigonelline from the small intestine of the specific pathogen-free (SPF) and germ-free (GF) rats in vivo. AB - After the direct injection of trigonelline (Tg) to the upper end of the duodenum in SPF and GF rats, the remaining Tg in the contents of small intestine was determined after fractionation by ion-exchange and Nucher column chromatography, and by using a high performance thin-layer chromatography (HPTLC). When Tg was injected to the upper end of the duodenum of SPF rats, the remaining Tg in the sac markedly decreased with time. The results obtained from the experiment with GF rats almost correspond to that with SPF rats. It was therefore clear that Tg is not destroyed by the small intestinal microflora and a greater part of Tg is absorbed from the small intestine. PMID- 10721126 TI - Physiological roles of tryptophan in pig nutrition. AB - Tryptophan (TRP) is shown to play original roles in the physiology of pigs. Dietary TRP deficiency induces depression of the appetite leading to reduced growth performance. Brain hydroxy-indoles, including the neurotransmitter serotonin, are closely related to dietary TRP supply. Excess protein, namely large neutral amino acids (LNAA) enhanced the appetite depression, providing some support to a role for plasma TRP:LNAA in the regulation of protein intake through serotonin. Other implications of TRP, as a precursor for serotonin, in the susceptibility of pigs to stress and in their consequences on meat quality were reported. Furthermore, we concluded to a role for TRP in the insulin response to the meal and in tissue sensitivity to insulin. Implications in pig feeding practice are briefly reviewed. PMID- 10721127 TI - Efficiency of a lysine-tryptophan blend as a tryptophan source in animal nutrition. AB - The use of L-tryptophan for balancing tryptophan deficient diets in animal feeding is currently impeded by the cost of feed-grade product. Recently ADM CORP. (Decatur, IL, USA) developed a method whereby liquid L-tryptophan and liquid L-lysine-HCl could be blended before drying. The final product contains at least 15% L-tryptophan and 70% L-lysine-HCl (Trademark Tryptosine). Experiments have been conducted with pigs and broilers to evaluate the efficiency of tryptophan from Tryptosine in comparison with feed grade L-tryptophan. Results indicated a clear tryptophan deficiency of the basal diet. Graded doses of tryptophan to the basal diet resulted in improvements in weight gain, daily feed intake and feed conversion rate. In pigs the differences in weight gain, feed intake and feed conversion rate between both tryptophan sources were not significant. In broilers feed intake with commercial L-tryptophan was higher than with Tryptosine. Weight gain, feed conversion rate and health state did not differ significantly between both sources. Protein deposition in broilers was significantly improved with dietary supplementation of 0.3 g/kg tryptophan, whereas the addition with 0.6 g/kg showed no further response. No differences were observed between both tryptophan sources. From the results of the two studies it was concluded that the biological activity of tryptophan from Tryptosine is equal to that of L-tryptophan for pigs and broilers. PMID- 10721128 TI - Approaches in novel feed--EU legislation. AB - The relevant EC-legislation in the field of novel feed, inclusive the legislation about genetically modified micro-organisms is presented and discussed. PMID- 10721130 TI - Structural characterization of contaminants in commercial preparations of melatonin by on-line HPLC-electrospray ionization-tandem mass spectrometry. AB - Three different commercially available melatonin preparations were analyzed by on line HPLC-electrospray ionization-tandem mass spectrometry. All three samples contained the same impurities at the approximately 0.1-0.5% level of parent melatonin. Based on accurate mass-HPLC-MS and tandem mass spectrometric analyses, two contaminants (both MH+ = 249) were identified as hydroxylation products of melatonin. One compound (MH+ = 265) was determined to be a C-2 oxidation product of hydroxymelatonin and a group of four regioisomers (MH+ = 477) were identified as melatonin-formaldehyde condensation products. These latter contaminants are structural analogues of the case-associated peak "E" found in L-tryptophan implicated in onset of eosinophilia-myalgia syndrome. The significance of these findings is discussed. PMID- 10721129 TI - Electrospray ionization-tandem mass spectrometry for the analysis of tryptophan derivatives in food. AB - Our knowledge about bioactive tryptophan derivatives in the diet is rather limited. Consequently, our attention was focused on the efficient profiling, i.e. the structure specific detection, of novel tetrahydro-beta-carbolines and tryptophan glycoconjugates in food samples. Applying HPLC-MS/MS for screening and structural characterization, numerous products derived from the reaction of tryptophan with alpha-oxo acids and carbohydrates could be identified by means of neutral loss scanning. Subsequently, product ion experiments followed by the synthesis of the respective reference compounds accomplished structure elucidation of tryptophan derivatives. PMID- 10721131 TI - Melanogenesis from 5-hydroxytryptamine, 5,6- and 5,7-dihydroxytryptamines. An in vitro study using MALDI-TOF. AB - The role of tyrosinase and peroxidase in melanogenesis of 5-hydroxytryptamine, 5,6- and 5,7-dihydroxytryptamines was investigated by matrix-assisted laser desorption/ionization mass spectrometry. Each enzyme was incubated with the tryptamine derivatives and samples were drawn at various times, ultrafiltered and immediately lyophilized. The results indicated that peroxidase promotes oligomerization of 5-HT with fast kinetics but with yields lower than those achieved by tyrosinase. 5,6- and 5,7-DHT formed low molecular mass oligomers in the presence of peroxidase alone. The addition of hydrogen peroxide evidences different reactivity of the two isomers: 5,6-DHT formed immediately a black precipitate while oligomers of the molecule itself and of its oxidation products were detectable for 5,7-DHT. PMID- 10721133 TI - Advances in the analysis of tryptophan and its related compounds by chromatography. AB - Advances in the analysis of tryptophan,--(both in its free form and bound, alone or together with its metabolites),--has been compiled on the basis of the relevant papers published in the last 4-5 years, including author's experiences associated with the preparation of derivatives and with any of those conditions arising from the analytical procedure itself. The special requirements of various, tryptophan containing matrices were also taken into consideration (biological tissues or fluids, food and feed stuffs, etc). For the sake of completeness in addition to the most common HPLC/UV/F1 techniques, HPLC/MS, GC/MS, CE/UV/F1 and spectrophotometry will be also discussed. PMID- 10721132 TI - Tryptophans in membrane proteins. X-ray crystallographic analyses. AB - While tryptophans are generally found in low abundance in soluble proteins, in many integral membrane proteins they comprise a significantly higher proportion of the amino acid composition. Now that crystal structures are available for a number of membrane proteins, it has been possible to examine the distribution and disposition of the tryptophans within these structures. The tryptophan locations with respect to the lipid bilayer (along the direction normal to the membrane surface) are strikingly non-uniform in nearly all of the membrane proteins examined. They tend to cluster at the interface between the polar head group region and the hydrophobic interior, in a relatively uniform layer just below the surface. In many cases, their distributions with respect to the extra- and intra cellular surfaces tend to be asymmetric. These observations provide evidence for possible structural roles for tryptophans in transmembrane sheets and helices, where they may play a part in the stabilization of the transmembrane segments and perhaps in the orientation and bilayer insertion processes. PMID- 10721134 TI - Determination of N1-methylnicotinamide by HPLC postcolumn photoirradiation. AB - A method for the determination of N1-Methylnicotinamide (NMNA) was developed by the fluorescence formed by UV irradiation after separation by HPLC with a mobile phase containing hydrogen peroxide and 18-crown-6. This method was applied to human urine and serum. PMID- 10721135 TI - Simultaneous determination of 5-hydroxyindoles and catecholamines by HPLC with fluorometric precolumn derivatization. AB - We have developed a highly sensitive and simple precolumn derivatization method for simultaneous determination of 5-hydroxyindoles (5HIs) and catecholamines (CAs) with 4-dimethylaminobenzylamine (DMBA). The method was successfully applied to the determination of 5HIs and CAs in plasma and urine. PMID- 10721136 TI - An HPLC method to determine tryptophan and kynurenine in serum simultaneously. AB - A method was established to measure tryptophan and kynurenine in serum simultaneously. Tryptophan is converted to kynurenine by the action of the enzyme indoleamine 2,3-dioxygenase induced by interferon-gamma (IFN-gamma). Since IFN gamma is a Th1-cell derived cytokine, an increased tryptophan degradation rate via the kynurenine pathway can be found when the cellular immune system is activated as it is, e.g., in viral infections or in autoimmune diseases. Thus, the ratio kynurenine per tryptophan provides a possibility to estimate IFN-gamma activity in vivo and furthermore reflects the degree of immune activation. The HPLC method requires 100 microL serum. Protein is removed by trichloroacetic acid. An external albumin-based calibrator is applied, and analysis is referred to an internal standard, 3-nitro-L-tyrosine. Kynurenine and nitrotyrosine are detected via UV absorbance at 360 nm wavelength, and tryptophan is detected via its natural fluorescence at 285 nm extinction and 365 nm emission. Representative normal values of kynurenine and tryptophan were measured in the sera of 49 healthy blood donors. PMID- 10721137 TI - Enhancement of the specificity of an enzyme-based biosensor for L-tryptophan. AB - A new selective amperometric biosensor for reagentless L-tryptophan determination has been developed using immobilized tryptophan-2-monooxygenase (TMO, EC 1.13.12.3). This enzyme-based biosensor provides a rapid-response detection system for concentrations of L-tryptophan between 25 and 1,000 microM in a batch mode system and between 100 and 50,000 microM in a flow-injection mode. The response time was 30 seconds, and the total analysis time was less than 3 minutes. The biosensor retained catalytic activity and fidelity of phenylalanine and tryptophan response for greater than 4 months with repeated usage. The biosensor selectivity to L-tryptophan was dramatically increased relative to phenylalanine when a competitive inhibitor of TMO, indole acetamide (IA), was included. The biosensor was successfully used for L-tryptophan determination in nutrition broth, giving values identical to those determined by HPLC analysis. PMID- 10721138 TI - Background lead and cadmium exposure of adult women in Xian City and two farming villages in Shaanxi Province, China. AB - The objectives of the present study are: (a) to clarify the current levels of environmental exposure to lead (Pb) and cadmium (Cd) in Shaanxi Province in China in comparison with levels in other parts of mainland China; (b) to examine if there is any urban-rural difference in Pb and Cd exposure; and (c) to quantify the role of cereals as the dietary source of environmental exposure to these metals. For this purpose, triplet surveys on lead and cadmium exposure were conducted in the provincial capital of Xian and two farming villages A and B in Shaanxi Province, China in 1997. The grand geometric mean for lead (Pb) intake via foods (Pb-F), Pb in blood (Pb-B) and Pb in urine as corrected for creatinine concentration (Pb-Ucr) were 30 micrograms/day, 33 micrograms/l and 5 micrograms/g creatinine, respectively, with significant differences among the survey sites, e.g. Pb-B being higher in Xian (43 micrograms/l) than in the two villages (38 and 22 micrograms/l). The counterpart values for cadmium (Cd) intake via foods (Cd F), Cd in blood (Cd-B) and Cd in urine (Cd-Ucr) were 6.1 micrograms/day, 0.46 microgram/l and 2.8 micrograms/g creatinine, respectively, with no substantial inter-survey site difference. Thus, it was possible to conclude that, from comparison with the values reported in 1990s literature, the exposure of Shaanxi people to Pb and Cd is no higher than, and even possibly lower than, the levels reported for people in other parts of mainland China. The exposure to Cd was almost exclusively from foods, whereas the exposure to air-borne Pb was large enough in Xian to explain higher Pb-B and Pb-Ucr than the level in Village B despite lower Pb-F in Xian than in Village B. Cereals (wheat, rice, maize and foxtail millet) contributed 26 and 84% of dietary Pb and Cd intake, respectively. PMID- 10721139 TI - The character of the suspended and dissolved phases in the water cover of the flooded mine tailings at Stekenjokk, northern Sweden. AB - Studies of the suspended and dissolved phases of the pond water, material collected from sediment traps, and surficial sediments/tailings from the flooded tailings pond at Stekenjokk have been performed. The aim was to characterise the material, to study the seasonal variations and to quantify possible resuspension of the tailings in the pond. The element concentrations in the pond at Stekenjokk seem to be largely controlled by processes controlling the precipitation and dissolution of Mn- and Fe-oxyhydroxides in both the water column and in the surficial tailings. Physiochemical processes such as weathering of silicates on the surrounding mountain slopes or dykes contributes both dissolved elements and detrital particles. The suspended phase consists of detrital silicate material as well as Fe- and Mn-oxyhydroxides. The average heavy metal concentrations are high, e.g. 0.42% Cu, 0.15% Pb and 3.1% Zn, which is probably due to sorption onto Fe- and Mn-oxyhydroxides. The suspended phase is richer in Fe, and particularly Mn, during the winter. The suspended phase resembles the material collected in sediment traps and the material in the surficial sediments. The pond water is well mixed during the ice-free season. The dissolved heavy metal concentrations are generally rather low with, e.g. maximum concentrations of 2.03 micrograms/l Cu, 0.23 microgram/l Pb and 268 micrograms/l Zn during the winter. Higher dissolved concentrations are found below the ice-cover above the sediment surface during the winter, caused by diffusion of elements from the sediment-water interface up into the pond water. Most of the metals occurring in the pond are dissolved and resuspension of tailings is negligible. PMID- 10721140 TI - Risk of respiratory exposure of dental personnel to amalgam alternatives. AB - Although the use of alternatives to dental amalgam is increasing, the possible hazard associated with their occupational exposure has received inadequate attention. The purpose of this study is to use available toxicological and environmental information in a qualitative risk assessment to address potential health hazards associated with exposure to these materials by dental personnel. The members of dental profession should be aware of risk due to long-term exposure to dental materials. PMID- 10721141 TI - Alteration of bioluminescence in Amphipholis squamata (Ophiuroidea: Echinodermata) by heavy metals contamination: a field study. AB - The ophiuroid Amphipholis squamata (Echinodermata) is a bioluminescent species whose light production varies with physico-chemical parameters of the medium. Individuals collected in the bay of Portman along a gradient of heavy metal contamination show different patterns of light production: the ones from the highest contaminated area showing a bioluminescence weaker and slower than those from the lowest contaminated area. Individuals that were transferred for 3 days from the lowest to the highest contaminated area displayed a light production that became weaker and slower. It is suggested that the decrease of the bioluminescent capability due to heavy metal pollution could indirectly affect the ophiuroid ecological success (bioluminescence is associated with defense functions in ophiuroids. PMID- 10721142 TI - Chromium levels in spices and aromatic herbs. AB - We determined the presence of chromium in a total of 72 samples of 17 different spices and aromatic herbs. Electrothermal atomization atomic absorption spectrometry (ETA-AAS) was used to determine Cr content in the samples mineralized with HNO3 and V2O5. The analytical characteristics of the proposed method were tested, and the accuracy and precision was also verified against an NBS-certified reference material. Chromium concentrations ranged from not detectable to 1.42 micrograms/g (dry wt.) and Cr presence was detected in 95% of samples. Spices and aromatic herbs are widely consumed in the Spanish diet and in the Mediterranean diet, in general. PMID- 10721143 TI - Carbon and nitrogen fluxes in a closed seawater facility. AB - A variety of empirical and calculated data from the largest tank at the New Jersey State Aquarium were used to quantify the fluxes of carbon and nitrogen before and after the installation of denitrification in this facility. Before denitrification, the stock of dissolved inorganic carbon (DIC) in Ocean Tank exhibited a decrease of 6.9 kg C/month and sodium bicarbonate had to be added to maintain DIC in steady state. We were able to explain the DIC decrease by two non conservative processes: the formation of carbonaceous precipitates (removes 4.2 kg C/month) and outgassing of carbon dioxide due to acidity from nitrification (independently determined to remove 3.2 kg C/month). Nitrogen budget in Ocean Tank before denitrification is in contrast to that of carbon, and it shows an increase of 4.8 kg N/month in the form of nitrate. Denitrification is currently removing 53.3 kg N/month (in the form of nitrogen gas), so this element should eventually reach steady state. The use of methanol for denitrification has resulted in a flux of 26.3 kg C/month into the aquarium and, as predicted, an increase in Ocean Tank DIC stock has been observed without any additions of sodium bicarbonate. Our approach can be used to model carbon and nitrogen balances in closed seawater facilities that host heterotrophic organisms and operate either with or without a biological denitrification system. PMID- 10721144 TI - Oxidation of arsenite in groundwater using ozone and oxygen. AB - Oxidation of arsenite [As(III)] with ozone and oxygen was investigated in groundwater samples containing 46-62 micrograms/l total dissolved arsenic, 100 1130 micrograms/l Fe and 9-16 micrograms/l Mn. Conversion of As(III), which constituted over 70% of dissolved arsenic in the samples, to As(V) was fast with ozone, but sluggish with pure oxygen and air. Iron and manganese in the samples were also oxidized and, by sequestering the resultant As(V), played a significant role in the rate of reaction. Sorption capacity of freshly precipitated Fe(OH)3 was estimated to be 15.3 mg As/g. The kinetics of As(III) oxidation were interpreted using modified pseudo-first-order reaction. Half-lives of As(III) in experimental solutions involving saturation with each gas were approximately 4 min for the ozone reaction and, depending on the Fe concentrations, 2-5 days for pure oxygen and 4-9 days for air. PMID- 10721145 TI - Detection of carcinogenic aromatic amines in the urine of non-smokers. AB - Smoking is thought to be one of the most important anthropogenic risk factors involved in the development of urinary bladder cancer in humans. Tobacco smoke contains a complex mixture of chemicals including potent carcinogens such as aromatic amines. In the present study the amounts of several freebase aromatic amines including the potent carcinogens 2-aminonaphthalene and 4-aminobiphenyl have been analyzed in the urine of 48 German urban living smokers and non smokers. The results indicate that (i) both groups excrete the identical set of four aromatic amines; (ii) smokers excrete approximately twice the total amount of these amines, but similar amounts of 2-aminonaphthalene and 4-aminobiphenyl are found in non-smokers; and (iii) the excreted aromatic amines are decomposed in the urine within a few hours thus, explaining why aromatic amines are difficult to detect in this matrix. Their decomposition could be prevented by adding small amounts of p-toluidine to the freshly collected urine. Unlike smokers the origin of aromatic amines detected in the urine of non-smokers is at present unknown. Based on the cotinine levels found in the urine of non-smokers environmental tobacco smoke can be excluded as a major source of aromatic amines. In addition, neither diesel exhaust-related nitroarenes nor the corresponding amino-derivatives, to which they may be metabolically converted, were found. The detected urinary levels of aromatic amines arising from sources other than tobacco smoke or diesel exhaust may play a role in the bladder cancer etiology of non-smokers. PMID- 10721146 TI - Ultrastructural features of "solid cell nest" of the human thyroid gland: a study of 8 cases. AB - The ultrastructural features of solid cell nests (SCN), made of squamous cells, and associated calcitonin cells (C cells), of the thyroid gland were studied in only a few cases in humans. A study was performed on 8 paraffin-embedded SCN, postembedded in Epon, to look for their ultrastructural features. Immunohistochemical analysis using calcitonin antibody was performed on semithin sections of SCN to explore the presence of C cells. Three cases (37.5%) of SCN were positive for calcitonin, and electron-dense secretory granules were observed in the cytoplasm. In two of these cases, an increased number of C cells in the adjacent thyroid parenchyma was observed. The presence of ciliated and lymphoid cells, in addition to intracytoplasmic microvacuolar and microfollicular (microglandular) structures, was noticed. Ciliated cells have already been reported in embryonic rests of human and animals, but ultrastructurally for the first time in human SCN. The presence of microfollicular structures, intracytoplasmic microvacuolar, secretory granules features, and ciliated cells, in addition to lymphoid cell, suggests the existence of a common ultimobranchial stem cell for C cells or for one or more cell types of the thyroid gland. PMID- 10721147 TI - Phagocytosis of hemozoin (native and synthetic malaria pigment), and Plasmodium falciparum intraerythrocyte-stage parasites by human and mouse phagocytes. AB - Hemozoin, the detoxification product of hemoglobin heme, piles up as electron dense material in the food vacuole (FV) of intraerythrocytic malaria parasites (malaria pigment). In infected individuals, pigment is internalized by both circulating and resident phagocytes, thus modulating their functions. Synthetic beta-hematin, prepared in vitro from hematin (ferriprotoporphyrin IX hydroxide) in acidic condition, is spectroscopically identical to hemozoin. In this electron microscopy study, native and synthetic hemozoin also prove to be morphologically indistinguishable (large polygonal crystals with apparent transverse banding) and to undergo the same process when internalized by phagocytes (primarily a direct uptake of crystals, similar to what is described for asbestos fibers). On the contrary, whole parasites appear to follow a classical endocytic pathway. This suggests that there may be differences between the ingestion of free particles and whole parasites in terms of modulation of phagocytes' functions. PMID- 10721148 TI - Glomerulopathies with fibrillary deposits. AB - Renal diseases involving glomerular deposits of fibrillary material are an important diagnostic challenge for the ultrastructural pathologist. Two primary disorders of this type, termed "fibrillary glomerulonephritis" (characterized by fibrils measuring approximately 20 nm in diameter) and "immunotactoid glomerulopathy" (characterized by larger, microtubular deposits), have been described. The possible relatedness of these two disorders and their potential association with other systemic illnesses are subjects of current debate. Other multisystemic diseases, including amyloidosis and various forms of cryoglobulinemia, can also present with fibrillary or microtubular deposits in the kidney. Five cases are presented in which fibrillar or microtubular structures were identified in renal biopsies by ultrastructural examination. The distinction between fibrillary glomerulonephritis, immunotactoid glomerulopathy, and other processes that have similar ultrastructural features are discussed. PMID- 10721149 TI - Cell death in retinoblastoma: electron microscopic, immunohistochemical, and DNA fragmentation studies. AB - Apparent cell loss by apoptosis occurs in carcinomatous tissue. To investigate cell death in retinoblastoma (Rb), ultrastructural examination, ApopTag staining, electrophoresis to detect apoptotic DNA fragmentation, and flow cytometric studies were performed. Immunostaining for the oncogenic products bcl-2 and p53 was also carried out. Relationships between the proliferation fraction (PF), apoptotic index (AI), and the distribution of bcl-2 and p53 were investigated according to the degree of histologic differentiation of Rb. Ultrastructurally, two patterns of cell death were seen. Necrotic cells exhibited vacuolation of cytoplasmic organelles with a marked lytic change in the karyoplasm and cytoplasm. In contrast, apoptotic cells were characterized by crescentic margination of chromatin, condensation of karyoplasm and cytoplasm, and fragmentation of the nucleus. Differentiated Rb had a low AI value (< 1%), whereas undifferentiated Rb had a high AI value (> 8%). The PF of undifferentiated RB (31%) was significantly higher than that of differentiated RB (14%). Analysis of DNA fragmentation using 3'-end labeling with terminal transferase indicated that undifferentiated Rb has increased DNA cleavage. The distribution of apoptotic bodies within Rb was inversely correlated with the expression of bcl-2. A majority of tumor cells of differentiated Rb were negative for p53, whereas 20-40% of tumor cells of undifferentiated Rb showed a positive reaction for p53. These findings suggest that the degree of susceptibility to apoptosis is closely related to PF, is inversely related to the degree of differentiation of Rb, and is protected by oncogene bcl-2. PMID- 10721150 TI - Xanthogranulomatous appendicitis--an incidental finding of localized pathology. AB - The clinical, histopathological, and electron microscopic features of an unusual case of xanthogranulomatous appendicitis are reported. The patient, a 37-year-old female, presented with typical signs of acute appendicitis and the appendix appeared slightly dilated at laparatomy. The histopathological sections showed numerous xanthoma cells mixed with inspissated fecaliths. Electron microscopy disclosed the presence of xanthoma cells filled with electron-lucent lipid droplets of variable size. The ultrastructural characteristics of these cells enabled the distinction of two types of lipid-laden histiocytes, in relationship to the size of the lipid droplets. Since the lipid droplets were seen also in cells other than histiocytes, it appears that these changes are secondary to a common mechanism, comprising factors such as obstruction, hemorrhage, inflammation, and local hypoxia. PMID- 10721151 TI - Case for the panel. Numerous small vesicles in a case of clear cell leiomyoma of deep soft tissue: an ultrastructural study. PMID- 10721152 TI - Case for the panel. An unusual, common (?) finding in skeletal muscle biopsies. PMID- 10721153 TI - [Hypertension--normotension]. PMID- 10721154 TI - [How reproducible is blood pressure measurement? Intra- and interindividual blood pressure variability in routine practice]. AB - Considering the reproducibility of blood pressure readings the epidemiologist and the physician in his clinical work take two different points of view. Studying drug effects the epidemiologist is interested in significant differences of blood pressures in a study population. The physician however, who estimates the effect of his prescription given to an individual patient has to compare single blood pressure readings in single individuals. In a pharmacological study we accept without any doubt the need of certain numbers of investigations to define a difference of 10 mm Hg as significant. In clinical routine however we behave quite differently and judge single readings intuitively as equal or different, independent of any statistical considerations. Not only the doctor but also the scientific societies with the highest reputation share the same point of view. In their recommendations they accept an arithmetic mean of two blood pressure readings as sufficient to judge blood pressures in individuals. In order to discuss this topic primarily from the doctors point, we analysed repeated blood pressure readings of 21 volunteers who tested 20 different blood pressure devices. We conclude, that the ongoing practice of judging the blood pressures of the patients meets only the needs of the epidemiologist. Reproducibility of single readings or mean values of two readings however are insufficient to draw satisfactory conclusions from the distributions of blood pressure in single individuals. Due to the short and long term blood pressure variability increasing the numbers of readings within a single day does not improve reproducibility sufficiently. It seems that the amount of blood pressure readings which are necessary to improve reproducibility of "the patients blood pressures" should be increased and taken over several days. If this is true, daily self-recordings should be the most promising approach. PMID- 10721155 TI - [Hypertension--the status in Austria]. AB - In Austria approximately 25-30% of the population have hypertension. Nearly half of them is not aware of their condition. Only one third undergoes treatment. Of those who are treated only 10% reach normotension. Among physicians these values are even worse: nearly two thirds show an elevated pressure, but only 20% of them are treated. Treatment itself is unsatisfactory: nearly 100% of patients receive a monotherapy only. PMID- 10721156 TI - [A critical analysis of the Hypertension Optimal Treatment (HOT) Study (appeared in Lancet 1998; 351: 1755-1762)]. AB - In the HOT-study 18,790 patients in 26 countries (age 50-80 years, mean age 61.5 years) with hypertension (diastolic blood pressure between 100 and 115 mm Hg- mean value 105 mm Hg) were randomised into 3 groups with different target blood pressures (90 mm Hg, 85 mm Hg, 80 mm Hg). The basic treatment was with Felodipin (5 mg q. d.): if the target pressure was not achieved, further steps were initiated: either ACE-inhibitors or -blockers were added, in a third step the Felodipindose was increased to 100 mg and in a further step the doses of ACE inhibitors or -blockers were doubled. If target pressure was still not achieved, a diuretic was added. Furthermore half of the patients in all groups received either placebo or 75 mg acetyl-salicylic acid. To prevent cardiovascular events the best benefit was shown, when a diastolic pressure of 82.6 mm Hg was reached. Acetyl-salicylic acid showed a further benefit in preventing myocardial infarction, but not in preventing strokes. PMID- 10721157 TI - [Target blood pressure values from the cardiologic viewpoint]. AB - Recent large intervention trials in hypertension have shown that diastolic blood pressure--even in the presence of ischemic heart disease--can be reduced below 90 mm Hg with further benefit for the patient. This has lead to discussions about a new definition of target blood pressure in hypertension treatment. From a cardiological point of view, the total cardiovascular risk of the patient has to be calculated according to a coronary risk profile. This risk stratification of the patient helps to make a decision about pharmacological treatment of blood pressure and target blood pressure. The higher the risk of the individual patient, the more aggressive blood pressure lowering is indicated. PMID- 10721158 TI - [Goal blood pressure values from the nephrologic viewpoint]. AB - In order to achieve optimal blood pressure control a combination drug therapy is necessary in the majority of hypertensive patients. Preferred treatment strategies comprise fixed combination drugs which enhance patients' compliance. Low dosages of each substance minimize side effects. The choice of antihypertensive drugs depends on concomitant diseases. The current blood pressure goals are 135/80 mm Hg for patients with essential hypertension and 120/70 mm Hg for patients with diabetes mellitus or renal diseases. In patients with primary renal disease or renal involvement in systemic diseases, ACE inhibitors or angiotensin II blockers with or without diuretics are preferably used. PMID- 10721159 TI - [Family practice quality circles between goals and reality--an interaction analysis]. AB - Quality circles are considered a key method for quality assurance in health care. However, there is a lack of systematic evaluation for quality circles in general practice, especially regarding the process quality of quality circle work. This article presents the results of an interaction analysis completing the systematic evaluation of quality circles in general practice in a region of south Germany. Using the so-called conference encoding method for interaction analysis we analyzed 7 out of 25 evaluated quality circles and 2348 interactions between the quality circle members. The participation rate of the moderators is high compared to the relative low contribution of the group members to the quality circle work. We could show that quality circles work topic-oriented, there is a wide exchange of experience between the group members and the group climate is positive. However, there were almost no specific activities to develop guidelines for diagnostic and therapeutic procedures. The results showed a significant discrepancy between the aims of quality circles and their practical realisation. Besides improved option for information and training programs for moderators and participants, we recommend further evaluation studies complemented with specific analysis of the process quality for example with the conference encoding method. PMID- 10721160 TI - [Indicators for ambulatory quality management in patients with bronchial asthma]. AB - The process of quality assurance and quality improvement in ambulatory care is bound to the development of quality indicators. The performance and outcomes of medical services offered by physicians have to be assessed on the basis of valid, reliable and practicable indicators. The aim of this analysis was to identify relevant process and outcome measures which could be used as quality indicators in ambulatory care of asthmatic patients in Germany. Therefore, the process of routine ambulatory care of asthmatic patients was analyzed and compared to existing management guidelines of national and international expert panels. A set of 51 indicators was derived from this analysis and a systematic review of 96 articles on quality management of asthma care selected after research in MEDLINE 1966-1997. This list of quality indicators should be discussed in quality circles for practicability in ambulatory care settings. PMID- 10721161 TI - [Diagnostic spectrum and treatment requirements of general practice clients. Results of the ADT Panel of the Central Institute of National Health Insurance Management]. AB - Results of the ADT-Panel of the Central Research Institute of ambulatory health care in Germany. The ADT-Panel of the Central Research Institute contains the remuneration data that are quarterly collected by office-based physicians and transmitted in anonymous form. These data can be classified according to the status of the insured patient, the diagnoses made or the treatment provided. The Patient Panel does therefore represent an important instrument which allows a rapid scientific analysis (3 to 4 weeks after the end of a quarter) about how patients are treated by different specialists. It might also be used to forecast trends. The Patient Panel does as well allow the demonstration of specialty related statistics on diagnoses and disease-related treatment provision. The complete publication and some exemplary tables can be found in the DGN Internet (www.dgn.de; Deutsches Gesundheitsnetz). A short summary is available on the home page of the Central Research Institute (www/zi-koeln.de). PMID- 10721162 TI - [Quality management and consumer orientation: survey of referring pediatricians of a Berlin pediatric clinic]. AB - Quality management in hospitals not only includes performance according to international medical standards but also the optimization of processes regarding internal staff as well as external customers. Total Quality Management (TQM) and the Business Excellence Model of the European Foundation of Quality Management (EFQM) require continuous evaluation of customer satisfaction. Specialists and family physician as external customers influence the patient's choice of a hospital. The aim of the present study is to evaluate the satisfaction of admitting physicians of a children's hospital with the help of a questionnaire. The results describe their needs and their level of satisfaction regarding service, information, cooperation and communication within the hospital. PMID- 10721163 TI - [Approaches to ethical decision making in correlation with medical quality management]. AB - The subject as well as the tool of quality management deal with ethical questions. It is shown what ethics can contribute to quality management. Three different concepts of assessing an act (consequentialism, deontology, virtue ethics) are presented and their consequences for the everyday decisions of physicians are described. Ethical considerations are made about the notion quality in the framework of quality management, about the function of economical aspects in quality-planning, and about the role of guidelines as instruments for quality-improvement and -assurance. PMID- 10721164 TI - [Evaluating the cost effectiveness of an ENT clinic by the physician. Development of a special cost and proceeds overview]. AB - The realization of expense and proceeds-correctional measures in the area of medicine is a current problem caused by the lack of transparency of medical costs. The responsibility and the influence on management success is increasingly assigned on the head physicians. The concerning people should be informed about the economic status of their clinic. Only on this precondition selective and efficient measures are possible. Within the scope of a project in three different ENT-clinics, a special system of expense and proceeds-survey was developed using the existing cost accounting of the hospital. This survey enables the physician to estimate the economic efficiency and weakness of his clinic. After providing four auxiliary tables, it is possible to get a global view on the expenses and proceeds which are relevant for the physician. This method will allow to find the reasons for low rentability using different options of calculation. Systematic control of medical-economic processes will be considerably simplified for the physicians. PMID- 10721165 TI - [Determining priorities in the development of medical guidelines. 1: Criteria, procedures and actors: a methodological review of international experiences]. AB - Setting priorities for the development of clinical practice guidelines has- similar to other decision-making procedures in health care--as much a political as a scientific component. Prioritizing guidelines aims to allocate resources to those health problems likely to maximize medical, social and economic outcomes associated with the use of these guidelines. This is a review and critical appraisal of international initiatives of systematically setting priorities for the development of clinical practice guidelines. Priority-setting criteria, both quantitative and qualitative methods as well as participation by relevant stakeholders will be discussed. This review provides possible decision-makers with an information basis which may assist in the development of concepts for setting priorities in a given context. PMID- 10721166 TI - [Practice guidelines--disorders of lipid metabolism. From recommendations to therapy of disorders of lipid metabolism by the Drug Committee of the German Medical Society. Drug Committee of the German Medical Society]. PMID- 10721167 TI - [Practice guidelines--legal aspects]. AB - Medical guidelines of professional institutions establish rules for good medical treatment (quality of care). They take the form of international, national, regional and local guidelines and are set up by international and national medical organisations, mainly by medical societies but also by regional (e.g. medical associations in the states) and local institutions (up to hospital departments). The rules of good medical treatment (guidelines) conform to the corresponding medical standard which is defined by scientific knowledge (evidence based medicine), practical experience and professional acceptance. The binding character of the standard for medical practice increases with the reliability of scientific evidence and practical experience. Insofar, medical guidelines which correspond to the standard fulfil a quality assurance function for medical treatment as a means of communication in the medical profession, an implementation function for the enforcement of standards and thus also a protective function for patients. Furthermore, they rationalize decisions of the courts by increasing the transparency of their medical basis and thus also contribute to the quality improvement of legal decisions on medical malpractice. Medical guidelines can also influence social law directives. On principle. Medical guidelines do not include any economic considerations (efficiency) and should be strictly separated from these (principle of separation and transparency). Medical guidelines have to be differentiated from health insurance law directives and recommendations. In addition to the quality assurance function, the function of guaranteeing the efficiency of treatment is a priority for health insurances. These objectives may be in conflict with each other. This paper is mainly concerned with the significance of medical guidelines for medical malpractice law. PMID- 10721168 TI - [Basic tenets for quality circles in North Rhine Medical Service of Public Health Insurance]. AB - The Medical Advisory Service of the Health Insurance in the area of the Northern Rhine (MDK Nordrhein) has set up an internal concept for quality management since 1998. This concept includes the installation and performance of quality circles. Staff members were internally qualified as "presenters". They worked out principles for quality circles of the MDK North Rhine which were implemented as binding basic rules by the managing conference of the MDK. The principles will be presented in detail. PMID- 10721169 TI - [Mediastinal lymphadenopathy: an extrahepatic manifestation of chronic hepatitis C?]. AB - Normal and enlarged perihepatic and mediastinal lymph nodes are detectable by ultrasonography. Aim of the present study is to determine the detection rate, size, and correlation of mediastinal and perihepatic lymphadenopathy in patients with chronic hepatitis C, healthy controls, and patients with inflammatory or neoplastic mediastinal lymphadenopathy. The mediastinum and liver hilus of 89 patients with chronic hepatitis C as well as of 34 healthy volunteers and 20 patients with mediastinal lymphadenopathy of different origin with adequate sonographic visualization were screened for the number and size of lymph nodes by high resolution ultrasonography. Lymph nodes were detectable in the mediastinum of 75/89 (84%) patients with chronic hepatitis C and 22/34 (65%) healthy volunteers (total lymph node volume [LNV]: 1.0 +/- 0.8 mL versus 0.3 +/- 0.4 mL, p < 0.001). In all patients with mediastinal lymphadenopathy, the mediastinal lymph node volume was above 15 mL. In patients with chronic hepatitis C a trend could be observed, that patients with larger perihepatic lymph nodes reveal also larger mediastinal lymph nodes. High resolution ultrasonography is able to detect enlarged mediastinal lymph nodes in patients with chronic hepatitis C. Mediastinal lymphadenopathy is considered as an extrahepatic manifestation of chronic hepatitis C. In general, the mediastinal lymph node volume differs in size to patients with lymphadenopathy related to neoplasia or sarcoidosis. The mechanism of lymphadenopathy in the liver hilus and mediastinum in patients with chronic hepatitis C is yet unknown. PMID- 10721170 TI - Weekly oxaliplatin, high-dose infusional 5-fluorouracil and folinic acid as palliative third-line therapy of advanced colorectal carcinoma. AB - The efficacy of oxaliplatin combined with high-dose 5-fluorouracil (5-FU) and folinic acid (FA) as an outpatient salvage treatment for patients with metastasized colorectal cancer was retrospectively analyzed in one center. Tumor progression had occurred for the majority of patients during two regimens (n = 11) otherwise during one (n = 1) regimen of prior 5-FU-based chemotherapy, which had been applied in a standardized sequential fashion. As third-line therapy oxaliplatin was infused intravenously over 2 h at a dose of 60 mg/m2 prior to a 2 h infusion of FA (500 mg/m2). 5-FU (2,600 mg/m2) was subsequently given over 24 h. A favorable response was observed in 9/12 (75%) of the heavily pretreated patients, including partial remissions in 3/12, minor responses in 2/12 and stable disease in 4/12 patients. The median progression free time was 23 weeks (interquartile range i. r. 0-28) for all patients, the median survival time from start of third-line therapy 55 weeks (i. r. 40-86). The median survival time from the beginning of first-line palliative chemotherapy was 34 months (i. r. 25-45 months). The highest toxicity was WHO grade III and was observed in six patients: Nausea (2), diarrhea (3), vomiting (2) and peripheral neuropathy (1). The quality of life was not adversely affected by the oxaliplatin/5-FU/FA-regimen as assessed by the EORTC QLQ-C30 questionnaire. Thus, the results show the efficiency and low toxicity of oxaliplatin/high-dose 5-FU/FA as palliative third-line therapy of patients with metastasized colorectal cancer and emphasize that sequential palliative chemotherapy may lead to extended survival of these patients. PMID- 10721171 TI - Interferon-alpha-2b + ribavirin for the treatment of chronic hepatitis C in non responders to interferon-alpha-monotherapy. AB - The efficacy of a therapy with interferon-alpha-2b and ribavirin in patients with hepatitis C not responding to the initial treatment remains controversial. Objective of the present study was to determine the efficacy of a combination therapy with 3 MU interferon-alpha thrice weekly and 1,000-1,200 mg ribavirin daily for six months in an open label trial. METHODS: 44 consecutive patients (genotype 1: 79.5%) who had been previously treated with a minimum dose of 3 MU interferon-alpha thrice weekly for at least three months were monitored. Sustained response was defined as virological response with clearance of HCV-RNA after a follow-up period of six months after the end of therapy. RESULTS: A significant decrease of the mean levels of AST (34.3 +/- 27.6 vs. 16.4 +/- 13.5 U/L) and ALT (59.6 +/- 48.8 vs. 23.2 +/- 20.7 U/L) was observed. In 15/44 patients (34.1%) a virological end-of-treatment response was evident. 8/44 patients (18.2%) achieved a sustained response. In 33.3% of the previous non responders biochemical response with normal aminotransferases, but no virological response was evident. A significant decrease of the mean hemoglobin level was observed during therapy (12.0 +/- 1.7 vs. 14.6 +/- 1.0 g/dl; p < 0.005), which returned to the pretreatment values after the end of treatment (14.4 +/- 1.6 g/dl; n.s.). In ten patients (22.7%) decrease of hemoglobin levels < 10 g/dl led to a dose reduction of ribavirin. CONCLUSION: This study--in agreement to recently published data--provides evidence for the efficacy of the combination therapy with interferon-alpha-2b and ribavirin in a relevant subset of non responders to interferon-alpha. Therapeutic nihilism should be reevaluated in non responders to interferon-alpha. PMID- 10721172 TI - [Dilatation of pancreatic duct stenoses with the ring coagulator--initial experiences]. AB - Stenoses of the main pancreatic duct in chronic pancreatitis should be treated by interventional endoscopy. If mechanical dilatation was unsuccessful, a thermodilator has been in use in our clinic since September 1998. From September 1998 to March 1999 dilation of pancreatic duct stenoses was done by the thermodilator in six patients. An asymptomatic elevation of amylase and lipase was noticed in one patient. There were no further complications. The termodilator is a promising instrument for dilatation of difficult stenoses of the main pancreatic duct. Before general use, however, evaluation of the success and complication rates in a greater number of patients is needed. PMID- 10721173 TI - [Diarrhea in a patient with Down syndrome and endemic sprue]. AB - BACKGROUND: Down syndrome is associated with disorders such as celiac disease, hypothyroidism, and insulin-dependent diabetes mellitus. In patients with mono- or oligosymptomatic celiac disease the time interval between the onset of symptoms and diagnosis often is unacceptably long. CASE REPORT: A female patient with Down syndrome is presented who had acute watery diarrhea, which spontaneously ceased but recurred after a few days. After endoscopic and histologic evaluation and measurement of gliadin, endomysium, and reticulin antibodies celiac sprue was diagnosed. Further investigation showed findings of autoimmune hypothyroidism and secondary hyperparathyreoidism. After the patient was put on a gluten-free diet her state quickly improved. CONCLUSION: Associations between Down syndrome and autoimmune diseases exist. Patients with acute gastrointestinal symptoms should be evaluated as to celiac disease. The time interval between the onset of symptoms and diagnosis of celiac disease can be shortened, if all diagnostic tools are used at the appropriate time. PMID- 10721174 TI - [Pulmonary metastasis of extranodal high malignancy B-cell non-Hodgkin lymphoma of the bulbus duodeni and pylorus of the stomach]. AB - We report a 71-year-old female patient with repeated vomitus, meteorism, epigastric pain and reflux for more than four month. She had a palpable mass in the upper abdomen and lost 7 kg of weight during the last four months. Chest X ray showed two masses, 2 cm and 3 cm in diameter, in the left and right lower lung. A stenosing polypoid mucosal swelling in the antrum and the duodenal bulb. The pulmonal masses were biopsied under CT-guidance. Biopsy proved a high malignant B-cell non-Hodgkin's lymphoma of the stomach. The masses in the lung were identified as metastases of the gastrointestinal lymphoma. In conclusion on this tumor was an extranodal non-Hodgkin's lymphoma stadium BE IV according to Musshoff. A CHOP-chemotherapy was initiated. Restaging after three cycles of CHOP revealed a complete remission. Primary gastrointestinal non-Hodgkin's lymphomas are relatively rare neoplasms of the abdomen. Unusual and interesting in this case ist the metastatic pattern involving the lung periphery without local lymph node metastases. PMID- 10721175 TI - [Signet ring cell carcinoma of the stomach--a case of diagnostic dilemma]. AB - Inguinal lymphonodal metastases of an adenocarcinoma were diagnosed in a 40-year old patient by ultrasound guided puncture. The leading symptom was elephantiasis preferentially of the right lower extremity. In addition, atypical mycobacteriosis was detected later on. The causal gastric cancer had not been identified during two years until the fourth gastroscopy assisted by endoscopic ultrasonography revealed the lesion. No regional lymph node metastases were found while distant metastases in terms of inguinal lymph nodes were already present. PMID- 10721177 TI - [Comparison of virtual with conventional colonoscopy in detection of colorectal polyps]. PMID- 10721178 TI - [Value of genetic diagnosis of C282Y mutation in patients with hereditary hemochromatosis]. PMID- 10721176 TI - [Intestinal vasculitis--a diagnostic-therapeutic challenge]. AB - Intestinal vasculitis is a rare cause of mesenteric ischemia. It results in chronic arterial insufficiency in most cases, sometimes in acute mesenteric ischemia. Abdominal symptoms like postprandial intestinal angina, diarrhea, anorexia, and perforation are nonspecific and do not allow for differentiation between vasculitic and noninflammatory causes of mesenteric ischemia. Conventional radiography and endoscopy can not prove the underlying process either. Therefore, extraintestinal symptoms of vasculitis must be observed carefully for diagnosing a systemic vasculitis with potential involvement of intestinal arteries. Extraintestinal manifestations are multifacetted including malaise, rheumatic symptoms and more specific findings like cutaneous efflorescences and organ-specific vasculitic damages due to ischemia of inner organs, nerves and sensory organs. While some vasculitic disorders are characterized by specific laboratory markers (ANCA, anti-ds-DNA antibodies), others appear with less specific signs. Prior to treatment, the diagnosis should be established by biopsy of suspect tissue and subsequent histologic analysis. Angiography can be helpful in diagnosis of syndromes involving medium-sized or larger vessels. The treatment of choice is glucocorticoids, while in patients with extensive visceral, especially renal involvement, cyclophosphamide should be added. When glucocorticoids can not be tapered or the disease can not be controlled other immunosuppressive agents should be employed. In difficult diagnostics with mere suspicion of vasculitis glucocorticoids may be given ex juvantibus and fairly often prove effective. PMID- 10721179 TI - [Diagnosis of aneuploidy with fluorescence in situ hybridization (FISH); value in pregnancies with increased risk for chromosome aberrations]. AB - BACKGROUND: In the case of abnormal ultrasound findings, abnormal serum-screening and age-risk in advanced pregnancy a rapid diagnosis or exclusion of a chromosomal aneuploidy of the fetus is of great value for the clinical management. With fluorescence in situ hybridization (FISH) on uncultured amniotic fluid cells the detection of the most common aneuploidies, which account for about 2/3 of all chromosomal aberrations [1], is possible within 24 hours. The aim was to evaluate if the FISH-technique in combination with karyotyping after cell culturing could replace other methods like diagnostics form umbilical cord blood or placental biopsies. MATERIALS AND METHODS: For the FISH assays commercially available directly with fluorochromes labelled DNA-probes (Vysis, Stuttgart) were used. FISH assays were performed on amniotic fluid samples from pregnancies at risk for fetal chromosome aberrations parallel to standard cytogenetic analysis. The method was performed on 230 samples of amniotic fluid. We tried to optimize the method concerning preparation of the cell material, the denaturation- and hybridization-steps and well as stringency of post hybridization washes. RESULTS: All trisomies 13, 18, 21 and the sex chromosome aneuploidies (n = 34) which were diagnosed by conventional cytogenetics were identified correctly by FISH analysis with the exception of one case of trisomy 21 mosaicism, in which hybridization failed. As structural chromosome aberrations and mosaicisms cannot be detected with this method, six additional chromosome aberrations were identified exclusively by cytogenetic analysis. The mean frequency of nuclei with abnormal signal pattern in the aneuploid cases was 89%. A minimum of 50 nuclei for each DNA-probe could be counted in 86% of the samples. The results of 12 cases were classified as uninformative, because only less than 15 hybridized nuclei or no hybridization signals could be scored. Maternal contamination was found in 17.4% of the samples. CONCLUSIONS: In clinical cases with a high risk for an abnormal fetal karyotype and the need of quick clinical consequences, methods which make possible a karyotyping within shortest time should be preferred to amniocentesis and FISH-analysis, because Chromosomal mosaicism and structural aberrations, which represent up to 20% of all chromosomal abnormalities in this group, cannot be detected, uninformative cases can occur in up to 15% of all investigated samples and There is a risk for false negative results through contamination of the sample with cells of maternal origin. In comparison with methods which permit rapid karyotyping from umbilical cord blood or placental biopsies, a delay in the diagnostic procedure has to be accepted, when the result of the FISH-analysis has to be confirmed by cell culturing and standard cytogenetic analysis. PMID- 10721180 TI - [Interphase FISH test as a rapid test for trisomies in amniotic fluid--results of a prospective study]. AB - BACKGROUND: Specific DNA probes allow rapid prenatal diagnosis of numerical chromosome disorders (chromosomes 13, 18, 21, X, Y) by FISH on interphase nuclei. The diagnostic reliability is presently under evaluation. STUDY GROUP: In a period of 1.5 years a total 1126 amniotic fluid samples was investigated by FISH compared to standard cytogenetic analysis. RESULTS: The success rate was 93 percent (< or = 30 nuclei) and 84% (< or = 50 nuclei). An abnormal karyotype was detected by FISH in 27 of 28 successfully hybridised samples, including trisomy 21 [16], trisomy 13 [4], trisomy 18 [4], aberrations of sex chromosomes [4]. Two cases with clinically relevant cytogenetic abnormalities were in principle not detectable by FISH. One false-negative finding was observed, possibly arising from maternal cell contamination of the sample. 6% of all samples, respectively 23% of the bloody samples were contaminated by maternal cells (more than 10%). CONCLUSION: Maternal contamination represents the most important limitation of the diagnostic reliability in routine practice. PMID- 10721181 TI - [Early auditory evoked potentials in very small premature infants]. AB - BACKGROUND: Brainstem auditory evoked response (BAER) is a sensitive test of the functional integrity of the auditory pathway. Latencies and amplitudes of measured waveforms reflect maturation of this brainstem pathway and may allow a prediction for neurological outcome. PATIENTS AND METHODS: BAER were measured prospectively in preterm infants with a birthweight less than 1500 g at 3 to 4 day following birth and during treatment on neonatal intensive care unit (NICU) every two weeks. Pre- and postnatal risk factors and frequency of apnea were evaluated. Outcome was scored after 1 year corrected age by neurological examination. RESULTS: There were no significant differences in latencies of waves between 5 children with severe asphyxia and/or IVH resulting in major neurological handicaps after 1 year. However, two of these children have bilateral abnormalities. The slope of decay of wave latencies decreased between 30 and 32 weeks gestational age, while the number of registered apnea increased. In follow up investigations during treatment on NICU there was a decrease in interpeak latencies wave V to wave III from 24/25 weeks gestational age to term infants and increase in amplitudes. CONCLUSION: Early performed BAER alone is not a good predictor for neurological outcome of preterm infants because of great interindividual variability of normal neonates and the anatomical site of cerebral lesions. The method contributes information on the brainstem maturation and may also reflect the different maturational stages of the respiratory system. PMID- 10721182 TI - [Hair cell function diagram in premature and mature newborn infants]. AB - BACKGROUND: The grade of maturation of the cochlear function must be taken into account when the cochlear function is investigated in premature infants and neonates. Quantitative analysis has not yet been performed. METHODS: Using the otodynamic analyzer ILO 88/92, we therefore determined haircell functional diagrams and the echo 65 dB levels in 35 healthy neonates as well as 30 healthy premature infants with a median gestational age (GA) of 32 weeks (28-35 weeks) 2 3 days post natum. Follow up investigations were performed in 10 neonates and 16 premature infants. RESULTS: The mean echo 65 dB levels were 4.88 +/- 4.22 dB for the right ears and 5.72 +/- 5.16 dB for the left ears in premature infants < or = 32 weeks of GA, 10.25 +/- 3.89 dB and 10.69 +/- 5.55 dB respectively in premature infants > 32 weeks of GA, and 16.01 +/- 4.99 and 14.90 +/- 4.41 dB respectively in neonates. There was no significant difference between neonates at day 2 and 4 post natum. In 16 premature infants the echo 65 dB levels increased from 4.92 dB for the right ears and 4.79 dB for the left ears at the second day of life to 11.72 dB and 11.73 dB respectively at the age of 4 weeks. CONCLUSION: We present therefore strong evidence for a maturation of the cochlear function in premature infants with increasing age. The individual grade of maturation of the cochlea is of relevance for the auditive stimulation of very premature infants. PMID- 10721183 TI - [Maternal and fetal digoxin level in fetofetal transfusion syndrome (FFTS)]. AB - BACKGROUND: Even though invasive intrauterine techniques for the treatment of TTTS such as punction of amniotic fluid and laser coagulation of placental vascular anastomoses are established methods in specialized centers, invasive methods are not always sufficiently successful. In conservative treatment of TTTS oral or intravenous maternal digoxin therapy in order to improve fetal cardiac insufficiency in combination with or after failure of invasive techniques is an useful method. PATIENTS AND METHODS: We investigated 12 TTTS pregnancies and 4 singleton pregnancies, which had been treated by maternal digoxin treatment for TTTS or arrhythmias, respectively. At birth, which was performed by means of caesarian section, venous cord blood samples of the newborns and venous maternal blood samples were collected, centrifugated and stored at minus 20 degrees C. Digoxin determinations were performed by radioimmunoassay. RESULTS: Fetal digoxin levels varied between 0.38 and 1.73 ng/ml, maternal levels ranged from 0.97 to 3.23 ng/ml. The fetomaternal digoxin gradient reached a mean of 0.56 (range 0.35 to 1.09). Donator and acceptor gradients were comparable and increased with birth weight or gestational week, respectively. CONCLUSIONS: In cases of pregnancies with TTTS a relatively high maternal digoxin level is necessary, especially during early gestational weeks, in order to reach therapeutical levels in the fetal circulation. Too low dosages might be responsible for unfavourable results in digoxin treatment of TTTS. Whether the maturation of placental villi during gestation could be the reason for increasing digoxin gradients requires further investigations. PMID- 10721184 TI - [Venous sinus thrombosis in puerperium]. AB - BACKGROUND, PATIENT RESPECTIVELY AND METHODS: Based on a case-report we demonstrate which symptoms are indicative of a cerebral sinus-venous thrombosis, how it can develop, and which diagnostic and therapeutic methods are available. RESULTS: Cerebral sinus-venous thrombosis in puerperium can be hidden behind unspecific symptoms like strong headache, nausea, vomiting. Literature reorts varying mortality between 5 and 30%. CONCLUSIONS: The obstetrician has to be attentive whenever these symptoms appear for the course and prognosis decisively depend on a quick diagnosis by means of computertomography and angiography. DISCUSSION: Blood coagulation disorders such as lack of Antithrombin III, protein C, or protein S, have been identified as a particularly important origin of the cerebral sinus-venous thrombosis. It is subject to discussion whether the patient should be advised against any further pregnancy. From a medical point of view, a subsequent pregnancy seems acceptable because the risk of another thrombosis, which is a rare event anyway, can be significantly reduced by providing anticoagulation substances. PMID- 10721186 TI - [Unfulfilled desire for a child--coping strategies of involuntarily childless women and men]. AB - OBJECTIVE: We analyzed whether coping strategies vary depending on gender and sterility diagnosis. MATERIAL AND METHODS: We investigated 110 couples using the "Freiburg Questionnaire of Coping with Illness" by Muthny [14]. The questionnaire consists of 5 analytic scales, covering one coping-strategy each: F1: depressive coping; F2: problem-faced coping; F3: diversion and building-oneself-up; F4: religion and sense-seeking; F5: trivialization and wishful thinking. RESULTS: Women with unfulfilled child-wish score lower than the group of chronically sick only on the scale "religion and sense-seeking", whereas involuntarily childless men activate all coping strategies to a lesser extent than the standardized collective. Compared to their partners, women score higher on the scales "depressive coping" and "diversion and building-oneself-up". CONCLUSIONS: Gender and sex-role expectations related to it influence the experience of infertility. PMID- 10721185 TI - [Procalcitonin as monocytic marker for early diagnosis in septic abortion]. AB - BACKGROUND: In search of sensitive and specific markers of systemic infection procalcitonin (PCT) recently was promoted to the focus of clinical research. Little is known about the biology of PCT and until now no data have been presented about clinical importance of PCT in obstetric patients. PATIENT AND METHODS: Daily PCT values in a 17 year old patient with septic abortion were compared with established markers of systemic inflammation. Cultivated monocytes were analyzed by the means of indirect immunofluorescence for intracellular distribution of PCT. Additionally, PCT release into culture medium was examined. RESULTS: PCT values in comparison with established inflammation markers was demonstrated in the patient with septic abortion. Indirect immunofluorescence studies revealed the presence of PCT within monocytes. In the supernatants of monocyte cultures PCT was detectable under control conditions. Stimulation with lipopolysaccharide resulted in the increased PCT concentrations both in the supernatants of healthy and patient monocyte cultures. CONCLUSIONS: In the given patient PCT was superior to other inflammation markers with regard to early and progression diagnosis. Peripheral blood monocytes appear to be a potential site of inflammation-induced PCT production. For the first time intracellular distribution pattern and release of PCT from human monocytes was described. DISCUSSION: Based on the presented data broad clinical studies devoted to PCT evaluation in obstetric patients seem to be promising. As till now the interpretation of increased PCT values depended on empirical knowledge, extensive studies of the potential production site as well as its biological significance should be performed, too. PMID- 10721188 TI - [Estrogen and progesterone receptors of primary breast carcinoma and their axillary lymph node metastases--immunohistochemical investigations of routine formalin-fixed paraffin-embedded tissue]. AB - OBJECTIVE: The estrogen- (ER) and progesterone receptors (PR) of 27 pre- and 33 postmenopausal primary breast carcinomas with 320 axillary lymphonodal metastases were investigated in formalin-fixed paraffin-embedded tissue. MATERIAL AND METHODS: Special interest was devoted to the comparison of the receptor equipment between the axillary lymphonodal metastases and the appertained primary carcinoma. ER-antibody D5 and PR-antibody NCL-PGR were used for investigation. RESULTS: ER-positive carcinomas of premenopausal women and ER-respectively PR positive carcinomas of postmenopausal women corresponded significantly more frequent to receptor-positive lymphonodal metastases. Carcinomas with receptor positive metastases were mostly ER-positive and PR-negative. About all receptor positive metastases had the same combination of receptors. In cases of high numbers of lymphonodal metastases these were in general more often receptor negative. Respectively, the receptor-positive metastases numbered lower than the receptor-negative. Immunohistochemical receptor-negative primary carcinomas can inherit receptor-positive lymphonodal metastases. CONCLUSIONS: Immunohistochemical investigations render it possible to determine ERs and PRs not only in primary carcinomas but in lymphonodal metastases, too. Eventually the prognostic factors "ER" and "PR" will be used more often in the determination of therapeutic decisions. PMID- 10721187 TI - Pre-operative staging of cervical cancer: comparison of magnetic resonance imaging (MRI) and computed tomography (CT) with histologic results. AB - Thirty-two patients with histologically confirmed cervical carcinoma were preoperatively investigated using MRI; in addition, a CAT-scan was performed on 15 of these patients. The diagnostic results using both modalities were compared with the histological findings (after hysterectomy according to Wertheim-Meigs, including lymph node dissection in the pelvic and, in part, in para-aortal regions). Determination of tumour volume was possible with high accuracy using MRI. Accuracy in assessing the parametria was 86%, vagina 90%, bladder and rectum 97%. The shortcoming of MRI is still the detection of infiltrated lymph nodes. The accuracy of 69% achieved for lymph nodes is equal to results with computed tomography. The general accuracy for our patients in staging was 81% for MRI versus 47% for CT. MRI-based diagnosis enables us to determine a correct tumour staging preoperatively, and is therefore very helpful in planning an adequate therapy. If MRI were used more widely it would contribute to simplification and shortening of the preoperative diagnostic procedure in patients with cervical carcinoma. PMID- 10721189 TI - [Effect of isoflavones on mammary gland and endometrium of postmenopausal macaques (Macaca fascicularis)]. AB - OBJECTIVE: Because of their beneficial effects on atherosclerosis and cancer risk, isoflavones may be useful as a dietary alternative or supplement to postmenopausal hormone replacement therapy. Isoflavones belong to the most important phytoestrogens. They may have estrogenic and antiestrogenic effects. We examined this in a well-characterised primate model of the postmenopause. MATERIAL AND METHODS: Adult, surgically postmenopausal female macaques were treated with isoflavones, estradiol or placebo for 6 months. After 6 months of therapy histopathological, morphometrical, and immunohistochemical measurements of endometrium and mammary glands were performed. RESULTS: 6 months of isoflavone therapy did not induce proliferation or any other clinical important changes in endometrium and mammary tissue. Phytoestrogentherapy did not show estradiol comparable effects. We are discussing our results with the results of other studies, which are sometimes in contrast. CONCLUSIONS: Our results indicate that isoflavones do not have estrogenic effects in the tissues studied. PMID- 10721190 TI - [Turner' syndrome (monosomy) and pregnancy]. AB - Fertility in cases of 45-XO-Turner's syndrome is extremely rare. In cases with pregnancy, spontaneous abortions, stillborns and chromosomal defects are frequently diagnosed. We report on the unusual case of a now 36-year-old woman, who is to our knowledge the 25th case of monosomy X to achieve pregnancy, but is the first one to give birth to three healthy children. PMID- 10721191 TI - [Incidental diagnosis of a vaginal foreign body during early pregnancy after an unusually long-term presence]. AB - A case of a metallic vaginal foreign body not recognized a remarkable period of time is presented. The foreign body, a screw, was incorporated at the age of 3 years. It was found and removed when the young woman had to be admitted to hospital because of an early pregnancy complication. PMID- 10721192 TI - [Treatment of vaginal atresia by a cecum transplantation after radical operation for cervical cancer and postoperative radiotherapy]. AB - Case report is given about treatment of atretic vagina after radical operation of cervical cancer and postoperative radiation. Neovagina was created with coecum. Good plastic results with uncomplicated cohabitation were reached. This method is recommended in cases with actinic lesions of vagina. PMID- 10721193 TI - [German Cohort Study of Women's Health--benefits of oral contraceptives. Study protocol]. AB - This publication is about the study protocol of the German Cohort Study on Women's Health. The main objective is to investigate medical benefits of a long term oral contraceptive use. The design is an analytical cohort study based on inquiries. Additional cases will be recruited to analyse rare events in separate case-control studies. Voluntary participants who signed to participate in a long term study are included. An annual drop-out rate of 15% is expected. Study variables encompass personal characteristics, lifetime history of diseases, but also disturbances of the state of health, and quality of life. It is anticipated to achieve 400,000 women-years of observation by 2001 (historic and concurrent follow-up). The study started April 1, 1998 and the current financial phase finishes December 31, 2001. 6000 participants were recruited until December 1998 equivalent to about 190,000 observation-years. Until the end of 1999, an additional 70,000 women-years should be included. There have been many suggestions from participants' to include additional issues of women's health into the study. PMID- 10721194 TI - [Consultation with regard to benefit-risk assessment and possible side effects of a long-term postmenopausal hormone substitution with estrogens-gestagens. New developments in hormone substitution therapy]. PMID- 10721195 TI - [25 years "Anesthesiology and Reanimation"--a historical review]. AB - The 25th anniversary of the foundation of the journal "Anaesthesiology und Reanimation" seems to be a good occasion, first of all, to look back at the special situation regarding the opportunities open to East German anaesthetists for publishing anaesthesiological papers before and after the Berlin Wall was built and then to give a review of the history of this journal. As the author's own publication list shows, East Germans could publish papers in West German journals without any problems before a major reform of the universities, bringing drastic changes, was introduced in East Germany in 1969. It became practically impossible to publish papers in West German journals because the "Directorates of International Relations", which had been installed at all universities in 1969, supervised the entire correspondence with persons and institutions in all foreign countries, in particular West Germany, the other West European countries and the countries of North and South America. Thus, East German anaesthetists were forced to publish in non-anaesthesiological East German journals because there was no journal of anaesthesiology in East Germany until "Anaesthesiologie und Reanimation" was founded as journal of the "Society of Anaesthesiology and Reanimation of the GDR" in 1976. The problems arising from the introduction of this journal under socialist conditions, including political pressure and control through the "General Secretariat of the Medical Scientific Societies of the Ministry of Health of the GDR" as well as technical problems with the publisher and the printers, are described. In spite of all these problems, which were overcome by the editor-in-chief with the aid of his colleagues on the editorial board and the scientific advisory council, this journal was initially published with a circulation of 1,200 copies in 1976 and its circulation increased to 1,600 copies in 1989. The journal proved to be of great benefit to East German anaesthetists and anaesthetists from other East European countries. It was included in an international exchange programme of anaesthesiological journals, which was particularly helpful for East German anaesthetists because they could not subscribe to West German, West European or American journals due to a lack of hard currency. The international exchange of the journal led to an increasing number of authors from West Germany and other West European countries and even from the USA and Canada who published papers in "Anaesthesiology und Reanimation". The "silent revolution" in 1989 brought new problems. The journal was primarily an organ of the "Society of Anaesthesiology and Intensive Therapy of the GDR", but with the end of the GDR, this society was dissolved on 23rd October 1990. Fortunately, "Anaesthesiologie und Reanimation" was taken over by the "German Society of Anaesthesiology and Intensive Medicine" as an organ of this society, in which the former members of the East German society were gathered. The next problem was that the publisher "VEB Verlag Volk und Gesundheit", Berlin and "Verlag Gesundheit GmbH", Berlin respectively ceased to exist in 1992 and we had to look for another publisher. We were very happy that "Selecta Verlagsgesellschaft mbH", Munich, later Wiesbaden, was interested in this journal and took it on in the same year and has now published it since that time with a circulation of 1,000 copies. The chequered history of "Anaesthesiology und Reanimation", the subtitle of which has been changed to "Journal of Anaesthesiology, Intensive Therapy, Emergency Medicine and Pain Therapy" in 1991, clearly shows, on the one hand, the difficult political circumstances under which the development of anaesthesiology took place in East Germany, and demonstrates, on the other, the special problems of the foundation of a medical journal under socialist conditions with which the editorial board in general and the editor-in-chief in particular were confronted and how they tried to overcome the PMID- 10721196 TI - [Exposure of anesthetists to sevoflurane and nitrous oxide during inhalation anesthesia induction in pediatric anesthesia]. AB - Inhalational mask induction with nitrous oxide and sevoflurane in young children is an appropriate alternative to intravenous induction and is considered safe and of rapid onset. Disadvantages of this technique are environmental pollution and occupational exposure to the inhalation agents used. Moreover, the potential health hazards are not yet completely clear. The purpose of the present study was to examine the anaesthesiologist's occupational exposure to nitrous oxide and sevoflurane in paediatric anaesthesia and mask induction. Twenty children underwent inhalational induction with nitrous oxide and sevoflurane in the operating theatre (air exchange rate 20.2/h, anaesthetic waste gas scavenger 40 l/min). Anaesthesia was maintained with the same agents. Air samples were taken from the edge of the anaesthesiologist's mouth continuously every 90 seconds, and trace concentrations of nitrous oxide and sevoflurane were analyzed with a direct reading infrared spectrometer (Bruel & Kjaer 1302, Denmark). Measurements taken during anaesthesia showed an increase in the concentrations of the anaesthetics used, but these were low. The highest mean concentrations occurred during induction (3.35 +/- 4.23 ppm for sevoflurane and 37.09 +/- 11.65 ppm for nitrous oxide). The overall peak levels measured were 6.31 +/- 4.23 ppm for sevoflurane and 68.78 +/- 40.79 ppm for nitrous oxide. Though the induction period was short compared to the whole length of anaesthesia, its impact on the overall waste gas exposure was 46.3% for sevoflurane (nitrous oxide 40.6%). Nonetheless, applicable German health law regulations were never infringed. The trace concentrations measured during inhalational mask induction and maintenance of anaesthesia were very low. With regard to modern workplace laws and health care regulations, gaseous induction in paediatric anaesthesia does not threaten the personnel's health. PMID- 10721197 TI - [Effects of sevoflurane versus propofol on oculocardiac reflex--a comparative study in 180 children]. AB - Oculocardial reflex (OCR) occurs particularly through manipulation of the medial rectus muscle and results in a bradycardic arrhythmia. In children the incidence is between 60 and 80%. After using sevoflurane in clinical practice, the absence or non-occurrence of this reflex was observed. The data of 180 healthy children aged between four and 14 years who had to undergo strabismus surgery under general anaesthesia were analysed: group I (n = 92), group II (n = 88). All children received standard premedication with midazolam, no anticholinergic drugs were administered. During narcosis, analgesia was maintained routinely with alfentanil. In group I sevoflurane was inhaled for hypnosis and in group II propofol was injected as intravenous hypnotic drug. The depth of anaesthesia was adjusted according to clinical criteria. To compare both groups, heart rate (HR) was determined before, during and after surgical intervention. OCR was defined as a heart rate declining by more than 20% from the initial HR.OCR is described in all methods of general anaesthesia. Under sevoflurane the occurrence of the reflex was significantly (p < 0.05) reduced to 14% of all patients as compared to 75% in patients who received a propofol infusion. Sufficient reflex reduction according to the depth of narcosis under sevoflurane in combination with the sympathomimetic effects of this drug could therefore be discussed as a reason for its positive effects. In our opinion, the use of sevoflurane should be considered as an option for general anaesthesia in strabismus surgery. PMID- 10721198 TI - [Analgosedation (managed anesthesia care--MAC) with propofol and piritramide for controlled cyclophotocoagulation of the eye]. AB - The new method of controlled cyclophotocoagulation of the eye is an example of a short, non-invasive procedure that is still too painful to be done under local anaesthesia alone. The risks associated with general anaesthesia, on the other hand, seem to be inappropriately high compared to the risks associated with the procedure itself. Therefore, for this procedure, we combined local anaesthesia with 0.5% proxymetacain and 10% cocaine and sedation with propofol and analgesia with piritramide. Our experiences with this method have been positive. We have applied our method to 42 patients undergoing a total of 53 procedures and we have seen no major changes in haemodynamics and only two cases of momentary slight ventilatory depression. Therefore, we conclude that our method of managed anaesthesia care is suitable for patients undergoing cyclophotocoagulation of the eye, combining patient comfort with haemodynamic stability and minimal risk for the patient. PMID- 10721199 TI - Physical control of the eutrophic response in the northern Adriatic Sea, illustrated by a nitrogen budget from ELNA data. Eutrophic Limits of the Northern Adriatic. AB - The northern Adriatic Sea would normally classify as an oligotrophic sea except for the eutrophic effect caused by its large runoff of nutrients. The coastal transports and the fresh-water distributions of the 1994 ELNA data suggests that, in the mean, a high percentage (approximately 84%) of the fresh water remains trapped in the nearshore and is exported from the northern Adriatic. The distributions of fresh water and primary production are well correlated: high values of fresh water and primary production (approximately 178 gC/m2/yr) in the nearshore, "dystrophic zone"; less fresh water and deeper production of approximately 85 gC/m2/yr in the interior, "eutrophic zone" compared to the background level of production of approximately 56 gC/m2/yr in the "oligotrophic zone". An inorganic nitrogen budget, revealed that only about 16% of the total nutrient input is directly exported; that 67% is utilized in the dystrophic zone; and that the production in the eutrophic zone was supported mostly by nutrients recycled from the organic matter produced in the dystrophic zone but subsequently sedimented out to the deeper interior. Thus, biological transport dominated over physical mixing as the more important mechanism of transferring nutrients to the interior. PMID- 10721200 TI - General features of the Adriatic Sea: the key role of carbon cycling. AB - Major inputs into the northern Adriatic Sea and critical problems affecting this basin are revised strengthening the need for a thorough knowledge of organic carbon cycling. Results gathered in the context of the PRISMA 2 project by our research group give information on the distribution and composition of organic matter in terms of major biochemical components, molecular weight sizes and biodegradability characteristics. Seasonal accumulation of dissolved organic matter in summer along with an increased role of carbohydrate and colloidal matter (being on average 15-20 and 60-65% of DOC, respectively) and restricted bacterial activity due to P deficiency are all priority subjects to be further investigated. PMID- 10721201 TI - The mucilage phenomenon in the northern Adriatic Sea. A critical review of the present scientific hypotheses. AB - In the summers of 1988, 1989, and 1991 large quantities of sticky mucilaginous masses occurred in the Adriatic Sea, particularly in its northern part. The mucilage phenomenon has been studied by scientists during past events, but the previous scientific reports back to the thirties of this century. Great efforts have been made since 1988 to understand the nature of the phenomenon. Although remarkable scientific results have been achieved, many questions related to such a complex phenomenon have remained open. In this paper results and hypotheses related to the chemical and biological composition, causes, triggering mechanisms, and responsible organisms for the mucilage phenomenon are briefly reviewed. Finally, some suggestions for future researches are proposed. PMID- 10721202 TI - Seasonal fluctuations of DIN/DIP and DON/DOP ratio in the northern Adriatic Sea. AB - Within the frame of PRISMA 1 "Biogeochemical cycles" research project (April 1995 January 1996) the quantities and the compartments of dissolved nitrogen and phosphorus have been studied in the northern Adriatic basin, considering also the organic pools. The research aimed to provide a better understanding of nutrient availability and to investigate the possible factors which promote the phenomenon of mucilage formation. For this purpose, the availability and the ratios between dissolved nitrogen and phosphorus considering both inorganic and organic fractions have been studied in relation to variations of river outflow and of biological activities. The results obtained reveal the large importance of organic nitrogen (annual average 16 microM) and phosphorus (annual average 0.13 microM) in contributing to the total nutrient availability (annual average total dissolved nitrogen: 29 microM and phosphorus: 0.18 microM) and the pronounced seasonal variability mainly ascribable to biological processes of uptake and remineralization. Furthermore, beside the well documented unbalanced ratio between inorganic nitrogen and phosphorus, the results obtained point out, for the first time, the unbalance also in the organic compartment (ratio between organic nitrogen and phosphorus ranges between 50 and 530), whose consequences might be important in relation to the phenomenon of mucilage formation. PMID- 10721203 TI - Structure/function/activity relationships in marine snow. Current understanding and suggested research thrusts. AB - Marine snow and marine snow components contribute to the mucilage phenomenon in the northern Adriatic Sea. Of special relevance is the matrix material, composed of extracellular polymeric substances, which are packaged into fibrils of colloidal dimensions. These 0.005 micron diameter fibrils are physical units of mucilage which can be visualized by transmission electron microscopy (TEM). They form polymer bridges between the various biotic and mineral components of marine snow, creating three-dimensional networks which affect floc porosity, density and settling behaviour. Recent observations of the matrix by TEM reveal complex fibril-delimited channels and capillary systems which partially traverse marine snow flocs and which are postulated to play roles in anomalous settling. Considering the marine snow floc as a microecosystem, the relationships between ultrastructure, chemistry and environmental properties are being explored. On the assumption that colloidal matrix materials, including those released into the bulk water, might provide advance information on anomalous floc behaviour, two new methods are recommended for monitoring the northern Adriatic Sea. One is a technique for chemical quantification of colloidal organic carbon. The other uses TEM, applied to water fractions derived from cascade ultracentrifugation, to estimate fibril quantities as a proportion of colloidal organic carbon. PMID- 10721204 TI - The potential role of particulate diatom exudates in forming nuisance mucilaginous scums. AB - Polysaccharide-specific staining techniques have revealed the existence and high abundance of gel-like mucilaginous particles formed from the polysaccharide exudates of diatoms. Evidence from both the field and laboratory indicates that these transparent exopolymer particles, known as TEP, are a major component of diatom aggregates and facilitate the flocculation of diatom blooms. TEP are composed primarily of sulfated polysaccharides, molecules known to be resistant to bacterial decomposition. Furthermore, the presence of TEP in diatom aggregates reduces the permeability of aggregates causing them to become neutrally buoyant and accumulate at picnoclines where they eventually develop gas bubbles and float to the surface. Diatom species, such as Cylindrotheca closterium, found abundantly in the Adriatic Sea, may produce particularly unique and copious quantities of TEP resistant to sinking and degradation. This material could accumulate as mucilaginous nuisance scums when environmental conditions favor strong stratification of the water column, high diatom biomass, and the dominance of these species. PMID- 10721205 TI - Phytoplankton extracellular production and leakage with considerations on the polysaccharide accumulation. AB - It is well known that many diatoms release and accumulate copious amounts of extracellular polysaccharides. Dramatic differences between diatom species exist. Examples from Prymnesiophyceae and Prasinophyceae show that algae from these classes as well may produce large amounts of extracellular polysaccharides. Soluble cell content may be released as extracellular components as a result of leaky membranes caused by unfavourable light conditions, pH, salinity, temperature, nutrient status and attack by bacteria or virus. Cellular soluble compounds may differ markedly from extracellular compounds produced under normal photosynthetic conditions. Production of polysaccharides in the Adriatic Sea may be promoted by heavy phytoplankton growth over long time under nutrient limitation. Sinking and flotation seem important processes in making aggregates and massive gels. PMID- 10721206 TI - Interaction between specific hydrological and microbial activity leading to extensive mucilage formation in the northern Adriatic Sea. AB - The massive formation of marine snow and the senescent stage of it, the mucilage, is a phenomenon largely restricted to the Adriatic Sea. In this contribution the major environmental factors potentially leading to the formation of this mucilage are discussed. It is proposed that the specific hydrological conditions in combination with severe phosphorus depletion lead to excessive formation of colloidal organic matter by phytoplankton. This colloidal organic matter coagulates to marine snow due to the low-turbulence regimes prevailing in the water column. Subsequently, this marine snow is colonized by bacteria which, in turn, produce and release copious amounts of capsular polymers into the matrix of marine snow. It is speculated that a significant fraction of the later stages of marine snow (mucilage) consists of bacterial-derived organic matter which has been shown to be semi-labile to refractory for further bacterial utilization. The marine snow matrix acts as efficient adsorption site and allows the bacteria to utilize scavenged molecules from the ambient water. Thus it is proposed that the matrix ages without significant biotic degradation. PMID- 10721208 TI - Human health implications associated with mucilage in the northern Adriatic Sea. AB - Mucilage in the northern Adriatic Sea is well known for its negative impact not only on the ecology of the affected area and on fishing activities but on tourism as well. The microhabitat mucilage creates in the sea can provide favourable conditions for the growth and/or survival of some environmental microorganisms that include human opportunistic pathogens. It also seems to favour the selective development of some marine toxic algae. Finally, mucilage can concentrate chemical contaminants from surrounding waters, hence increasing their bioaccumulation in seafoods. This paper examines the possible direct and indirect effects on human health of mucilage and other forms of marine aggregates. PMID- 10721207 TI - Significance of bacteria in the mucilage phenomenon in the northern Adriatic Sea. AB - Episodes of massive mucilage formation in the northern Adriatic Sea have been recorded for over a century but their cause is still a matter of conjecture and debate. It is generally thought that mucilage forms due to copious polysaccharide exudation by phosphorus limited algae. In this paper we develop the thesis that bacteria play major roles in mucilage formation. We argue that mucilage is largely produced as a consequence of bacteria-organic matter interactions and bacterial capsular polysaccharide synthesis. Ectohydrolytic enzymes of bacteria are critical in producing long-lived polysaccharides. Further, bacteria cause efficient P regeneration, particularly intensely in microscale features e.g. phycospheres, detritus and aggregates. Bacteria thus help sustain high rates of primary production despite vanishingly low levels of phosphorus in the bulk phase seawater. We integrate these roles of bacteria into a conceptual model which emphasizes microscale interactions of microbes within a seawater gel matrix as the basis for a mechanistic understanding of the accumulation of long-lived polysaccharide to form mucilage. PMID- 10721209 TI - Use of amoxicillin and amoxicillin-clavulanic acid and hospitalization for acute liver injury. AB - Increase of acute liver injury in patients taking amoxicillin-clavulanic acid (co amoxiclav) as compared to those taking amoxicillin has been suggested. To further investigate the potential hepatotoxicity of the two drugs a historical cohort study was conducted in the Italian region of Friuli-Venezia Giulia. One hundred and eighteen potential cases of acute liver injury were identified through the regional hospital information system and medical records were reviewed for all of them. Overall, 12 cases of acute liver injury were identified: 3 cases occurred in the amoxicillin exposure category, 2 among co-amoxiclav group, and 7 in the non-use category. The adjusted estimate of the rate ratio was 5.7 (CI 95% 1.5 22.1) among users of amoxicillin alone and 6.2 (CI 95% 1.3-29.7) among users of co-amoxiclav. PMID- 10721211 TI - [The comminution of fibrous and prismatic tremolites: the effect on the diffractometric response]. AB - The aim of the present work is to study the comminution of tremolite contained in an acid-washed dolomite sample and in a New York talc sample using a gentle, wet, comminution technique. Tremolite in acid-washed dolomite shows a fibrous habit of a length/diameter ratio greater than 10/1; on the contrary, tremolite particles in the New York talc have a quite acicular or prismatic habit of a length/diameter ratio in general less than 3/1. For both tremolite types, the diffractometric responses follow trends similar and are well correlated to the comminution degree. Results confirm that the comminution of amphibolic asbestos is a crucial step for the quantitative determination of this substance by electronic or light microscopy and X-ray diffractometry. PMID- 10721210 TI - Idiopathic chronic fatigue and chronic fatigue syndrome: a comparison of two case definitions. AB - The aim of the study was to compare the signs and symptoms of individuals meeting two different definitions of chronic fatigue syndrome (CFS). Ninety-four patients fitting the eligibility criteria for idiopathic fatigue were enrolled into the study. Of the 94 patients, 48 met the 1988 definition of CFS, 20 the 1994 (but not the 1988) definition of CFS, and 26 met neither definition. The 1994 defined cases were more likely than 1988 defined cases, and non-syndromal individuals to be male, married, and high school educated. The 1994 cases were less likely than 1988 cases to present acute onset, self reported sore throat, mild fever lymphadenopathy, pharyngitis. In conclusion, the 1994 criteria increased the number of patients classified as CFS; however, those who fit only the 1994 criteria were less likely to have an acute symptomatic onset and signs and symptoms suggestive of an infectious process. PMID- 10721212 TI - [The fine fraction of airborne particulate: a pollutant of increasing environmental and health relevance. Methodologies for sampling and characterizing single particles]. AB - Among the atmospheric pollutants detectable in the environment, the inhalable airborne particulate (PM10) is regarded with increasing care. Indeed a number of epidemiological studies support the correlation between both acute and chronic adverse health effects and the PM10 pollution in the urban environment. According to recent results the health effects could be related to the physico-chemical characteristics of the particulate itself. With the aim of characterising the individual particles, PM10 samples collected by multi-stage cascade impactor are examined by scanning electron microscopy (SEM) and X-ray microanalysis with energy dispersion spectroscopy (EDS). The obtained data are analysed using multivariate statistical techniques (hierarchical clustering methods, principal factor analysis) to determine the components of the particulate. By these methods a series of studies was carried out by the Istituto Superiore di Sanita to investigate the PM10 pollution in several environmental situations. PMID- 10721213 TI - [Seasonal levels of respirable quartz measured at a site in the metropolitan area of Rome in 1997-8]. AB - After the International Agency for Research on Cancer (IARC), Lyon classified crystalline silica within Class I, i.e. substances for which there is clear evidence of carcinogenicity to man, the need was felt for further studies to quantify the concentration levels and ensuing risks the general population is exposed to. Crystalline silica is a well-known pollutant which has been largely dealt with in literature as far as occupational exposure is concerned. On the contrary, the data resulting from environmental studies are scarce and usually inadequate, hence the importance of determining the levels of quartz (the most common form of crystalline silica) in the respirable fraction of the airborne particulate in urban areas in a reliable way. This paper is the continuation of a previous study which reported data on quartz concentrations in PM10 collected in an urban site of the metropolitan area of Rome in 1994 (the first year PM10 started to be collected). Measurements had been made with a new procedure that involved the use of X-ray diffractometry and silver filters, permitting the reproducible quantification of small amounts of quartz in the urban atmosphere. In the present work we report the mean daily quartz concentrations in PM10 collected for four weeks representative of the four seasons, from May 1997 to February 1998, in the same sampling site as in 1994. We then compare the two sets of data; in addition, we report preliminary estimates of the potential risk of contracting lung cancer for the general population. PMID- 10721214 TI - [The microbiological characterization of the bioaerosol and leachate from an urban solid refuse dump: preliminary data]. AB - The present paper shows the results of an experimental study aimed at arranging a microbiological characterization of bioaerosol and leachate resulting from a sanitary landfill for solid urban waste situated near Rome. Bioaerosol sampling was performed by using the active sampling method referred to as surface air system, that is extensively used during indoor environmental monitoring. The microorganisms (bacteria and fungi) believed to be of relevance on bioaerosol and leachate with a view to hygienic risks, were investigated in order to estimate the potential risks to which the population and the workers can be exposed and consequently to allow corrective measures by monitoring campaigns of the examined matrices and by models of low environmental impact landfill. PMID- 10721215 TI - [The physician-patient therapeutic alliance within the solidarity pact for health]. AB - In an attempt to address the limitations of traditional health care, the National Health Service of Italy has recently begun to evaluate so-called "unconventional" forms of health care, with the aim of better meeting individual health care needs and preferences. To realize this objective, it is necessary to evaluate unconventional forms of care in terms of demand, efficacy, and appropriateness; it is also necessary to evaluate the possibility of integrating unconventional forms of health care into the existing National Health Service. Another extremely important aspect of health care in general is the relationship between the physician and the patient. Specifically, this relationship must consist of the physician and patient working together to choose a form of health care, including unconventional forms, that is acceptable to both the physician and the patient. PMID- 10721216 TI - [Evidence-based medicine and the diverse cultures of healing]. AB - The term "unconventional medicine" refers to a remarkably heterogeneous group of theories and practices (homeopathy, herbal medicine, acupuncture, etc.) different from those peculiar to the dominant health system of a particular society. An unifying characteristic of these practices is that they have not been scientifically tested and that unconventional practitioners largely deny the need for such testing. However, established research procedures are to be considered adequate to address the majority of questions related to unconventional therapies and promising unconventional therapies should be subjected to the same level of scientific scrutiny that is required for drug therapies used within the official medicine. While many questions about the risk/benefit ratio of unconventional therapies remain unanswered, millions of people are spending millions of dollars each year in several developed countries, thus, to define strategies aimed at the careful evaluation of these practices is needed. PMID- 10721217 TI - [The characteristics of the use and the levels of diffusion of nonconventional medicine]. AB - The national trends of the utilisation of non conventional therapies suggest that an increasing number of patients employ remedies that are outside the mainstream of what has been defined as conventional western medicine. The extent to which these practices have clinical efficacy according to biomedical criteria is a matter of ongoing debate. It may be that independent of any such efficacy, the attraction of alternative medicine is related to the power of its underline shared beliefs and cultural assumptions. The fundamental premises are an advocacy of nature, vitalism, "science" and spirituality. For patients, who choose alternative medicine, the most important reason to abandon conventional therapies could be to move from the sterile "high-tech" realm of official medicine to a more intimate "high-touch" intervention offered by non-physicians. PMID- 10721218 TI - [The physician-patient relationship in the contexts of different medicines]. AB - The quality of the doctor-patient relationship depends at least in part on the way in which various non-specific factors influence disease and healing processes, which applies in particular to the subjects' beliefs (attributions) concerning what causes or prevents disease and the efficacy of various possible remedies. This favours the alternative medicines since in the exchanges with the patients, their models of etiopathogenesis and of mode and mechanism of action of proposed remedies come much closer to common sense models than those of modern scientific medicine. Such an advantage is increased by the fact that at least 80% of the encounters between physicians and patients concern situations of malaise or discomfort not identifiable as specific diseases. In such situations, official medicine often fails to exert the necessary functions of listening, explaining, counselling and reassuring, but tends to an inappropriate use of diagnostic and therapeutic tools developed for specific pathologies. Therefore, the dialogue between the different medicines, by promoting the re-establishment of a patient centered approach, can increase the efficacy of the interventions which depends both on specific technical factors and the quality of the relationship. PMID- 10721219 TI - [Phytotherapy between traditional medicine and alternative practices: how much safety and how much efficacy?]. AB - Clinical evaluation of modern phytotherapy is mainly carried out today using the method of meta-analysis. These generally reveal a marked contradiction of the results of individual clinical studies. There are various hypotheses to explain this fact: in the first place the products used are often of a different chemical and pharmacological nature. Secondly, there is the problem of the "phytocomplex". According to this concept, all the components of a medicinal plant, and not only its active ingredient, can influence its therapeutic action, but as generally only the active ingredient is standardized, the other ingredients end up by influencing the quality of the individual preparations in a way that is unknown. In the last place, there is the problem of the actual quality of the individual clinical studies and the possibility of taking the indications conveyed by traditional medicine into account when designing them. PMID- 10721220 TI - [Acupuncture and traditional Chinese medicine]. AB - Acupuncture and other therapeutic techniques of traditional Chinese medicine (moxibustion, herbal therapy, massage and physiokinesitherapy, Chinese medical exercises) were introduced in Italy and Europe after the Second World War. Especially acupuncture is utilized in private and public health services; in Italy only medical doctors can practice acupuncture because it is considered a "medical act" from 1982. Some private schools organize four years courses of acupuncture and Chinese medicine and have acquired an interesting experience in teaching, clinical practice and editing in this field. The National Institutes of Health (NIH) in USA stated, in a consensus conference held in November 1977, that "there is clear evidence that needle acupuncture is efficacious for adult post operative and chemotherapy nausea and vomiting and probably for the nausea of pregnancy". PMID- 10721221 TI - [Homeopathy in the perspective of scientific research]. AB - A significant portion of the traditional concepts of homeopathy (similia principle, experimentation on healthy humans, the cure of whole person, the use of minimum doses or high dilution/potency of medicines) are amenable to investigations conducted according to criteria accepted by biomedical science. Even though many randomized and controlled clinical studies seem to demonstrate the efficacy of some homeopathic medicines and their superiority to placebo, other trials have given negative results. A definite clear answer is not yet possible due to the scarce quality of some published reports, to lack of reproduction by independent investigators and to the uncertainty regarding the methodologies to be used for testing the claims of homeopathy. As regards the possible physiopathological, biophysical and pharmacological explanations for the action of homeopathic remedies, there are models which tend to set the similia principle as a general expression of the action-reaction principle, within the context of dynamic systems theory. The clarification of the more controversial aspects regarding dilution/potency of medicines remains tied to several promising developments in physics of condensed matter, in chaos theory and in biophysics. PMID- 10721222 TI - [Scientific languages and the science of complexity]. AB - For centuries the goal of galileian and newtonian science has been the discovery of the universal laws of nature governing all natural phenomena. But, starting with the sixties of this century, advanced research has concentrated on the study of the characteristic aspects of irregular and unrepeatable evolutionary processes. Signs of this change are the rediscovery of Poincare's results on the dynamics of non linear systems, and the extraordinary development of evolutionary thinking. Three properties characterize complex systems: the multiplicity of their levels of organization, the presence of self-referential loops of internal communication, and the irreducible nature of their past history to structural properties. The traditional cartesian gap between mind and body therefore falls down, as argued at length by the works of Gregory Bateson. His arguments are further strengthened by the neurologist Antonio Damasio in his book on reason, emotion and human brain, whose title is L'errore di Cartesio (The error of Descartes), where the negative consequences of this error on the development of science in general and on contemporary medicine in particular, are illustrated. PMID- 10721223 TI - [The evolution of the biomedical paradigm and nonconventional medicines]. AB - At the moment, it could be possible to integrate scientific, conventional and unconventional and alternative medicine to improve the efficacy and the quality of therapies. This could be feasible thanks to the following details: a) after two centuries, the biomedical paradigm is radically changed: from mechanistic to complex, holistic vision of human physiopathology and therapeutics. This emerging paradigm enables the best assessment of unconventional medical theories and complementary and alternative therapies; b) into the diversified unconventional medicines world an open-mind to scientific debate is rising, closing the ancient contrast to conventional medicine. The pathway is crucial, it could set out the scientific quality of the conventional, complementary and alternative medicine integration. PMID- 10721224 TI - [Excitement versus competence. The logic of communication in the mass media]. AB - Italian media are very attentive to health and medicine issues. They face them- nonetheless--without any critical approach. Clinically tested medicine and "alternative" practices are presented as if they were both part of the same context. Such a lack of critical sense is no longer a question connected with sole medicine. It applies also to several different fields: from economics to foreign politics. The above situation is the result of a deep and radical change in the work organisation of the Italian mass media system. This change induced experienced professionals to leave their editorial offices--in so doing media critical esprit was no longer nourished. This change was the (avoidable) consequence of the exceptional increase in "infos" reaching the editorial offices through new telematics. The new situation led to a new conception of job organization and hierarchical structure. Where once small top managers' groups lived next to numerous specialised journalists, now editorial offices are composed by over 60 percent of staff concerned only with selection of news incoming through press agencies. PMID- 10721225 TI - [The liberty of care]. AB - In our societies everybody has a right to decide in order to treatments on her/his own body, according to her/his ideas, sensibility and wills. Physicians cannot answer to the patient's world only in terms of scientific rationality. The future challenge is to find the way to improve, at the same time, the scientific standard of medical treatments and the response to single patient's needs. PMID- 10721226 TI - Macrophage inflammatory protein-1 alpha (MIP-1 alpha) and leukemia inhibitory factor (LIF) protect the repopulating ability of purified murine hematopoietic stem cells in serum-deprived cultures stimulated with SCF and IL-3. AB - The engraftment potential of murine stem cells (HSC) is greatly reduced when these cells are expanded in vitro with stem cell factor and interleukin-3. We have evaluated if the addition of MIP-1 alpha or LIF to these cultures would protect the ability of murine wild type HSC to engraft the stem cell defective W/Wv recipient. In this transplantation model red and white blood cell reconstitution is assessed by hemoglobin electrophoresis and c-kit PCR genotyping, respectively. The results obtained indicate that both MIP-1 alpha and LIF protect, at least transiently, the HSC repopulating ability in vivo in spite of the modest expansion in the number of nucleated and progenitor cells observed. PMID- 10721227 TI - Oxidative stress markers: specificity and measurement techniques. AB - With time, increasing evidence has been obtained of the involvement of oxidative stress in the pathophysiology of several diseases and ageing. Likewise, a large number of epidemiological studies have suggested that some pathologies can be prevented or delayed to some extent by dietary changes such as increased consumption of fruits, grains and vegetables. The above mentioned studies have suggested that the measurement of oxidative stress status, coupled to measurement of antioxidant status, may serve a role in diagnosis and/or treatment. The objective of this paper is to provide a review which, owing to the extent of the available literature, is obviously not exhaustive of current and most recent methods employed for the determination of the most specific markers of DNA, lipid and protein oxidative damage. PMID- 10721228 TI - Sedimentable mineral organic detritus as radioecological indicator. AB - The sedimentable mineral organic detritus (SMOD) drawn by rivers can be considered an important matrix for the monitoring of contaminants in aquatic environments. In Italy the collection and radioactivity analysis of SMOD has been introduced and standardised in the 80s. However hydrological parameters, like the flow and the amount of suspended matter close to the sampling points, must be considered to improve the methodology. This technique has been applied by the authors since 1992 to monitor the concentration of both 137Cs (following the Chernobyl accident) and 131I (because of possible waste discharge from Perugia University Hospital) along the Umbrian course of the Tiber River. In this paper the results of 137Cs water concentration are presented and discussed in the light of our interpretative working hypothesis based on hydrological parameters. This work allowed us to reduce the number of sampling points without loss of radioecological information. PMID- 10721229 TI - [An experimental quality assurance program for asbestos measurements performed with x-ray diffractometry in bulk samples]. AB - A quality assurance program for the diffractometric determinations of asbestos in bulk samples is presented. The program, developed upon request of the Italian Health Ministry, includes two tests: the qualitative determination of the asbestos forms present in three samples and their quantitative measurements. The samples to be analyzed, the asbestos standards and the criteria for results evaluation are described in detail. The program will start in the year 2000. Compliance with the tests minimum requirements will be considered mandatory for a public or private laboratory be legally allowed to work in the field of asbestos determinations by X-ray diffractometry. The description of the program is preceded by an in-depth discussion on the definition of the terms "quality control" and "quality assurance", which are often confused, and on their operational meaning when they are applied to laboratory measurements. The analytical difficulties involved with the measurement procedures currently used are also discussed. Moreover, the results of a preliminary intercalibration test among fifteen Italian laboratories of proven experience are reported since they provided important information for the arrangement of the present quality assurance program. PMID- 10721230 TI - [Computer-assisted evaluation of surface electromyograms of 8 different arm muscles]. AB - The aim of the present study was, via digitization of the EMG signals and processing them in the computer, to make possible effective evaluation of electromyographically detectable modifications of the activity of the forearm muscles developing during wrist movements under defined conditions of loading. The search for the optimal measuring parameter was of particular importance. In groups of test subjects the electric voltage potentials generated by 8 arm muscles were measured simultaneously on the skin and evaluated using suitable variables (integral, RMS [root mean square] value, mean amplitude, summed amplitude, amplitude peak count [peaks], zero crossings) with the aid of a PC and a computer program specially developed for the task. On evaluation it was found that the parameters reacting sensitively mainly to amplitude size--integral, RMS value, mean amplitude height and summed amplitude--reflected, with gradually decreasing clarity, the activities of each muscle. The correlation among the first three parameters was almost linear. The parameters sensitive primarily to frequency modulations, such as peaks and number of zero crossings, yielded little information under the load applied. They did not adequately reflect the extension and flexion of the hand. PMID- 10721231 TI - Evaluation of fast time-domain based impedance measurements on biological tissue. AB - Bio-impedance measurements are widely used for characterization of biological objects. Although the measured impedance of such objects is independent of the measurement method used, slight differences between measurements in the frequency and time domain are found. For many practical applications time domain based measurements are advantageous, but they are often rejected as not accurate. In order to show their suitability for bio-impedance measurements we used a special arrangement of time domain and frequency domain based measurements at the same biological specimen (canine liver) with the same electrodes. A reasonable coincidence in the measurement results could be shown. Moreover we used only a fraction of the time domain measurement data in order to demonstrate a significant reduction in measurement time while maintaining a reasonable accuracy. An algorithm for fast processing of the time domain data without transformation into the frequency domain is provided. PMID- 10721232 TI - [Basis for information transmission with biosignals for development of technical communication aids for handicapped patients]. AB - For the design of communication aids controlled by biosignals for the handicapped, an analysis of the aspects of the significance of information is mandatory. The definition of information in its five aspects statistics, syntax, semantics, pragmatics and apobetics, enables us to conclude that the transmission of information is possible only with voluntarily influenceable biosignal components. The voluntary influencing of the biosignal may be interpreted as a modulation of the amplitude density of the Fourier integral. By calculating the highest possible statistical information content of a biosignal, it is possible to estimate the technical complexity of a biosignal-based communication system. The construction of efficient communication aids is possible when many biosignal components that can be readily and rapidly controlled voluntarily are to be found. PMID- 10721233 TI - The effects of the use of piezoelectric motors in a 1.5-Tesla high-field magnetic resonance imaging system (MRI). AB - PURPOSE: This paper presents the results of an experimental investigation with two different rotatory piezomotors in a closed 1.5 Tesla high-field MRI. The focus of the investigation was on testing the functionality of these motors within the MRI and to determining the image interference they caused. MATERIALS AND METHODS: To obtain a differentiated estimate of the interference the motors were tested in both the passive (turned off, i.e. without current flow) and active (turned on, i.e. with current flow) state during MRI scanning. Three different types of sequences were used for the test: Spin-Echo (SE), Gradient Echo (GE) and Echo-Planar Imaging (EPI). A plastic container filled with a gadolinium-manganese solution was used for representation of the artefacts. The motors investigated were placed parallel to the container at predetermined distances during the experiment. RESULTS AND CONCLUSIONS: The results show that the motors investigated suffered no functional limitations in the magnetic field of the MRI but, depending on the type of motor, the measurement distance and the state of the motor, the motors had different effects on the sequence images. A motor in the off-state placed immediately next to the object to be measured mainly causes artefacts because of its material properties. If, on the other hand, the piezomotor is in the on-state images with strong noise result when the motor is immediately next to the object being measured. The images regain their normal quality when the motor is approximately at a distance of 1 m from the object being investigated. Driving the motor inside the MRI, therefore, is only to be recommended during the pauses in scanning: this delivers artefact-free images if minimal, motor-specific distances are kept to. With regard to the three different types of sequences it was determined that the SE sequence was the least sensitive and the EPI sequence the most sensitive to disturbance. The GE sequence showed only minimal differences to the SE sequence with regard to signal-to-noise ratios. Since it requires considerably shorter scan-times it can be considered to be the most effective type of sequence under these conditions. PMID- 10721234 TI - [Pressure-dependent flow resistance in craniospinal cerebrospinal fluid dynamics: a calculation model for diagnosis of normal pressure hydrocephalus]. AB - Computer-aided processing of the results obtained with the intrathecal infusion test using our newly developed mathematical model simplifies the investigation technique and thus the diagnosis of normal pressure hydrocephalus. Simultaneous determination of resistance and compliance in a single session markedly reduces the examination-related stress on the patient. In contrast to the classical methods, the new calculation does not require the ICP to reach a plateau. Unlike the static approach, our model describes the functional pressure-dependent course of the resistance. This means that account is taken of the non-linearity of the CSF dynamics during the processing of the biosignal. The intrathecal infusion test used to measure resistance and compliance is a reliable diagnostic method in patients with a normal pressure hydrocephalus. PMID- 10721235 TI - Multiple object tracking and attentional processing. AB - How are attentional priorities set when multiple stimuli compete for access to the limited-capacity visual attention system? According to Pylyshyn (1989) and Yantis and Johnson (1990), a small number of visual objects can be preattentively indexed or tagged and thereby accessed more rapidly by a subsequent attentional process (e.g., the traditional "spotlight of attention"). In the present study, we used the multiple object tracking methodology of Pylyshyn and Storm (1988) to investigate the relation between what we call "visual indexing" and attentional processing. Participants visually tracked a subset of a set of identical, independently randomly moving objects in a display (the targets), and made a speeded identification response when they noticed a target or a nontarget (distractor) object undergo a subtle form transformation. We found that target form changes were identified more rapidly than nontarget form changes, and that the speed of responding to target form changes was unaffected by the number of nontargets in the display when the form-changing targets were successfully tracked. We also found that this enhanced processing only applied to the targets themselves and not to nearby nontarget distractors, showing that the allocation of a broadened region of visual attention (as in the zoom-lens model of attentional allocation) could not account for these findings. These results confirm that visual indexing bestows a processing priority to a number of objects in the visual field. PMID- 10721236 TI - Strategy selection in causal reasoning: when beliefs and covariation collide. AB - The present study investigated how people combine covariation information (Cheng & Novick, 1990, 1992) with pre-existing beliefs (White, 1989) when evaluating causal hypotheses. Three experiments, using both within- and between-subjects designs, found that the use of covariation information and beliefs interacted, such that the effects of covariation were larger when people assessed hypotheses about believable than about unbelievable causal candidates. In Experiment 2, this interaction was observed when participants made judgments in stages (e.g., first evaluating covariation information about a causal candidate and then evaluating the believability of a candidate), as well as when the information was presented simultaneously. Experiment 3 demonstrated that this pattern was also reflected in participants' metacognitive judgments: Participants indicated that they weighed covariation information more heavily for believable than unbelievable candidates. Finally, Experiments 1 and 2 demonstrated the presence of individual differences in the use of covariation- and belief-based cues. That is, individuals who tended to base their causality judgments primarily on belief were less likely to make use of covariation information and vice versa. The findings were most consistent with White's (1989) causal power theory, which suggests that covariation information is more likely to be considered relevant to believable than unbelievable causes. PMID- 10721237 TI - Executive control in set switching: residual switch cost and task-set inhibition. AB - Executive processes necessary for flexibly switching between different tasks were studied using a set switching paradigm that requires participants to rapidly switch between different tasks across consecutive trials. Switch cost reflects poorer performance for task-switch trials than for consecutive same-task trials. Significant switch cost was observed even with considerable preparation time before a task-switch, an effect known as residual switch cost. This study tested the hypothesis that one process underlying residual switch cost is inhibition of the previous task-set. We used semantic categorization tasks to compare switch cost between alternating task series (ABA) and nonalternating series (ABC) in order to test the generality of a task-set inhibition effect previously observed with perceptual judgment tasks (Mayr & Keele, in press). The results yielded significant switch cost only for alternating tasks, in both response times and errors resulting from performance of the wrong task. Thus, resolving inhibition associated with previously abandoned task-sets may be the main process underlying residual switch costs, suggesting that task-set inhibition is an important executive control process. PMID- 10721238 TI - Interactions between symmetry and elongation in determining reference frames for object perception. AB - Many theories of object recognition posit that objects are encoded with respect to a perceptual frame of reference. Such theories assume that factors such as symmetry and elongation are critical for the assignment of an object's primary axis, and consequently for the extraction of an object's reference frame. The present experiments directly examined the relative roles played by symmetry and elongation in the determination of an object's primary axis, and the extent to which symmetry and elongation interact with one another. We found that observers use both symmetry and elongation in extracting an object's primary axis, that the extent to which each cue dominates depends on its relative salience, and that symmetry and elongation are processed interactively, rather than in encapsulated modules. PMID- 10721239 TI - [Indicators and standards for evaluating the program process for cervical cancer screening. Operation manual. Italian Group for Cervical Carcinoma Screening]. PMID- 10721240 TI - Stress relaxation and recovery behaviour of composite orthodontic archwires in bending. AB - The viscoelastic behaviour of prototype composite orthodontic archwires was evaluated using a bend stress relaxation test. Archwires having 10 different volume fractions of reinforcement were subjected to constant bending radii in a water bath at 37 degrees C for time periods of up to 90 days. The wires were subsequently released and left unconstrained for the same testing conditions. Creep-induced changes in the unconstrained bending radii of the wires were measured at specific times during both phases (stress relaxation and recovery) of the test. The statistical analysis showed that stress relaxation behaviour was strongly correlated with the archwire reinforcement level. The final relaxation varied, with decreasing reinforcement, from 2 to 8 per cent. Archwire recovery was not correlated with reinforcement level, and revealed a final viscous loss of only 1 per cent. The relaxed elastic moduli in bending of the composite wires were similar to the elastic moduli in bending of several conventional orthodontic archwire materials. Losses that were associated with the viscoelastic behaviour varied with decreasing reinforcement level from 1.2 to 1.7 GPa. Because these modulus losses were minimal, each archwire retained sufficient resilience to be applicable to the early and intermediate stages of orthodontic treatment. PMID- 10721241 TI - The inappropriateness of conventional orthodontic bond strength assessment protocols. AB - The purpose of this article is to examine the soundness of conventional orthodontic bonding assessment methods. A classification of bond strength studies is proposed with the testing environment (in vivo, in vitro, and ex vivo), loading mode (shear, tensile, and torsion), and bonding substrate (enamel, restorative, and prosthetic materials) serving as discriminating variables. Inconsistencies throughout the various stages of research protocols are analysed. These include the following: tooth selection, storage, and preparation; bonding; testing; and data analysis with regard to the clinical applicability of the reported information, as well as the scientific integrity of the testing procedure. Contradictory models may partially account for the considerable variability noted for reported bond strength values of different orthodontic bonding systems. Such discrepancies may also explain the conflicting evidence reported on the failure characteristics of the components of the bonding system in different trials examining the efficacy of nominally identical materials. A novel approach to study the fatigue life of materials is proposed to understand the processes occurring prior to bond failure. Mock research data manipulation is also utilized to illustrate the correct statistical treatment of findings, and recommendations for future research are made to ensure scientific soundness and clinical applicability of data. PMID- 10721242 TI - The effect of a maxillary lip bumper on tooth positions. AB - The effect of the use of a lip bumper with anterior vestibular shields on the maxilla was studied in twenty-two 9-14-year-old children with a space deficiency in the maxillary dental arch. The lip bumper was used for 1 year. The effect of the treatment was evaluated from dental casts and profile cephalograms made before and after treatment. Both the width of the maxillary dental arch at the premolars and the length of the arch increased significantly by about 2 mm. The effect of the treatment on the antero-posterior position of the first molars was small. In one subject the molar was distalized 2.8 mm. The average effect was, however, a reduction in the anterior movement of the molar within the face by about 0.5 mm, i.e. the maxilla moved anteriorly 1 mm, but the molar only 0.4 mm. No skeletal effects were found when the group of subjects treated with a lip bumper was compared with a reference sample of untreated individuals. The main effects of a maxillary lip bumper thus seem to be a widening of the dental arch across the premolars, a moderate increase in arch length due to eruption and slight proclination of the incisors, and moderate distal tipping of the first molars. PMID- 10721243 TI - The retraction of upper incisors with the PG retraction system. AB - The aim of this study was to evaluate the effect on the dentoalveolar structures of the application of PG springs for retraction of upper incisors and to compare the outcome with the effect of a closed coil spring retraction system. Thirty-six subjects with Angle Class I or Class II malocclusions were selected for the study. Each subject had the two upper first premolars extracted and presented a symmetrical extraction space of at least 3 mm distal to the lateral incisors after canine retraction. The subjects were divided into two groups, the PG group with 17 subjects and the coil group with 19 patients. One group had the incisors retracted by PG universal retraction springs, whereas in the other a closed coil spring system was used. The average chronological ages were 18 years 4 months for the PG group, and 18 years 7 months for the coil group. In both groups the springs were activated to produce an initial force of 150 g per side. To examine the type of movement of the anterior and posterior teeth, and the time and rate of space closure, 20 parameters were measured and evaluated statistically with Wilcoxon and Mann-Whitney U-tests. In both groups the incisor retraction was accompanied by mesial movement of the buccal segments. Distal movement of the root apex of the incisors was observed in both groups, although more pronounced in the PG group (P < 0.01). A significant incisor intrusion resulting in a decrease in overbite was found in the PG group, whereas the deep bite increased significantly in the coil spring group. The PG spring produced a three dimensional control in the movement of the upper incisors, so that application of additional intrusive mechanics after completion of the incisor retraction became unnecessary. PMID- 10721244 TI - A comparison of chincap and maxillary protraction appliances in the treatment of skeletal Class III malocclusions. AB - The purpose of this retrospective investigation was to compare cephalometrically the treatment effects of chincap and maxillary protraction appliances in subjects with a Class III skeletal malocclusion with a combination of an underdeveloped maxilla and prominent mandible. Twenty-four patients were divided into two groups according to the treatment type; the chincap group (mean age 11.03 years, n = 12) and the Delaire type maxillary protraction appliance group (mean age 10.72 years, n = 12). In both groups, a significant increase in ANB, molar relationship, and overjet showed the effect of the appliances in the treatment of Class III malocclusions. In comparing the two groups, the maxilla was displaced more anteriorly and the molar relationship correction was greater in the maxillary protraction appliance group (P < 0.05). Angular and dimensional parameters for lower incisor/NB and nasolabial angle showed significant differences between the groups (P < 0.05). PMID- 10721245 TI - Craniomandibular status and function in patients with habitual snoring and obstructive sleep apnoea after nocturnal treatment with a mandibular advancement splint: a 2-year follow-up. AB - The aim of the investigation was to evaluate the status and function of the temporomandibular joint (TMJ) and masticatory system in patients with habitual snoring and obstructive apnoea after 2 years nocturnal treatment with a mandibular advancement splint. Thirty-two patients participated in the study, ranging from 43.0 to 79.8 years of age (mean 54.4 years, SD 8.78) at the start of treatment. All patients had been referred from the ENT department for treatment with a mandibular advancement splint. The acrylic splint advanced the mandible 50 70 per cent of maximal protrusion, opened 5 mm vertically, and was used 6-8 hours per night and 5-7 nights per week. Overjet, overbite, and molar relationship were measured on dental casts. The patients were asked to answer a questionnaire concerning symptoms of craniomandibular dysfunction (CMD). They were also clinically examined in a standardized manner, including registration of range of mandibular movements, TMJ sounds, pain on movement, and palpatory tenderness of the TMJ and the masticatory muscles. None of the patients showed more than five symptoms of dysfunction either at the start of or after 2 years of treatment. A decrease in the frequency of headache was found for nine of those 18 patients that reported headache (P = 0.004). A minor, but significant decrease in overjet and overbite was found and the molar relationship was also changed. It was concluded that 2 years' treatment with a mandibular advancement splint had no adverse effects on the craniomandibular status and function, but the observed occlusal changes requires further evaluation. PMID- 10721246 TI - Long-term follow-up of clinical symptoms in TMD patients who underwent occlusal reconstruction by orthodontic treatment. AB - Fifty-eight patients (mean age 18.4 years) who had received splint therapy for internal derangement of the temporomandibular joint (TMJ) were examined retrospectively to investigate the efficacy of occlusal reconstruction by orthodontic treatment. The subjects were divided into three groups: 18 patients (mean age 18.6 years) who underwent orthodontic treatment combined with the use of splints (ST group); 27 patients (mean age 18.2 years) who underwent orthodontic treatment without the use of splints (NST group); and 13 patients (mean age 17.9 years) who received only splint therapy for temporomandibular joint disorders (TMD; control group). TMJ sound, pain on movement and restriction of mandibular movement were examined at the initial examination (T1), at the end of the splint therapy for TMD or beginning of orthodontic treatment (T2), at the end of orthodontic treatment (T3), and at recall or 1 year after orthodontic treatment (T4). The following results were found. (1) The percentage of patients with no joint sound at T2 was 20-30 per cent. The percentage of such patients in both the ST and NST groups increased to over 50 per cent at T3, but slightly decreased to 39-50 per cent at T4. There were no significant inter-group differences at any time point. (2) The number of patients who had no pain on movement at T2 was 60-80 per cent. The percentage of such patients in both the ST and NST groups increased to over 90 per cent at T3, but then slightly decreased to 80 per cent at T4. There were no significant inter-group differences at any time point. (3) None of the patients showed restriction of movement of the TMJ at T2 or T4. One patient in the ST group was found to have restriction at T3. There were no significant inter-group differences at any time point. (4) The most frequent type of malocclusion in both ST and NST groups was anterior open bite. These results suggest that TMD symptoms that have been eliminated by splint therapy are not likely to recur due to subsequent orthodontic treatment, but it cannot be concluded that orthodontic treatment itself had a positive effect on TMD symptoms. The results also indicate that there is a relationship between anterior open bite and TMD. PMID- 10721247 TI - A sella turcica bridge in subjects with severe craniofacial deviations. AB - In earlier studies, a sella turcica bridge was stated to occur in 1.75 to 6 per cent of the population. The occurrence of a sella turcica bridge has not previously been studied in a group of patients with craniofacial deviations treated by surgery. Profile radiographs from 177 individuals who had undergone combined orthodontic and surgical treatment at the Copenhagen School of Dentistry were studied. A sella turcica bridge was registered in those subjects where the radiograph revealed a continuous band of bony tissue from the anterior cranial fossa to the posterior cranial fossa across the sella turcica. Two types of sella turcica bridge were identified. A sella turcica bridge occurred in 18.6 per cent of the subjects. PMID- 10721248 TI - Overbite depth and anteroposterior dysplasia indicators: the relationship between occlusal and skeletal patterns using the receiver operating characteristic (ROC) analysis. AB - This study was carried out to investigate the validity of the overbite depth indicator (ODI) and the anteroposterior dysplasia indicator (APDI), based on the cephalometric analysis of 122 Caucasians selected at random for assessment of vertical and sagittal relationships. Considering the occlusion, the sample was divided into three classifications in the sagittal component: 36 cases of neutrocclusion, 54 cases of distocclusion, and 34 cases of mesiocclusion. The sample was also categorized according to the overbite relationship: 54 cases of normal overbite, 34 cases of open bite, and 34 cases of deep overbite. In the sagittal component analysis, the APDI measurement resulted in significant differences between the neutrocclusion, distocclusion, and mesiocclusion groups. In the vertical component analysis, the ODI significantly distinguished between the normal and deep overbite groups, and the open bite and deep overbite groups, but not between the normal overbite and the open bite groups. A receiver operating characteristic (ROC) analysis showed that the APDI matched the anteroposterior molar relationship in 88 per cent, and the ODI matched the amount of incisor overbite in 81 per cent. PMID- 10721249 TI - Long-term follow-up of maxillary incisors with severe apical root resorption. AB - The purpose of the study was to analyse the mobility of teeth with severe orthodontically induced root resorption, at follow-up several years after active treatment, and to evaluate mobility in relation to root length and alveolar bone support. Seventy-three maxillary incisors were examined in 20 patients, 10-15 years after active treatment in 13 patients (age 24-32 years) and 5-10 years after active treatment in seven patients (age 20-25 years). All had worn fixed or removable retainers; seven still had bonded twistflex retainers. Total root length and intra-alveolar root length were measured on intra-oral radiographs. Tooth mobility was assessed clinically according to Miller's Index (0-4) and the Periotest method. Crestal alveolar bone level, periodontal pocket depth, gingival, and plaque indices, occlusal contacts during occlusion and function, and dental wear were recorded. There was a significant correlation (P < 0.05) between tooth mobility, and total root length and intra-alveolar root length. No correlation was found between tooth mobility and retention with twistflex retainers. None of the variables for assessment of periodontal status, occlusion and function were related to total root length or tooth mobility. It is concluded that there is a risk of tooth mobility in a maxillary incisor that undergoes severe root resorption during orthodontic treatment, if the remaining total root length is < or = 9 mm. The risk is less if the remaining root length is > 9 mm. Follow-up of teeth with severe orthodontically induced root resorption is indicated. PMID- 10721250 TI - Initial cleft size does not correlate with outcome in unilateral cleft lip and palate. AB - Clinical outcomes in children born with a cleft lip and palate (CLP) have been an area of interest for orthodontists for a number of years. Whilst tools for measurement of these outcomes are available, there is no widely accepted measure of initial cleft severity and no known quantitative indices. Therefore, the potential influence of initial severity remains unmeasured and largely ignored. The aim of this investigation was to determine the importance of initial cleft severity in determining patient outcome. The longitudinal records of 49 children born with a unilateral cleft lip and palate (UCLP), and treated in a single centre were examined. An index of initial cleft severity was developed that categorizes the cleft area as a percentage of the total palate area. The dental arch relationships of the same patients at 6 years of age were also determined. The nature of the association between these was investigated for agreement and correlation by calculation of weighted Kappa and Spearman's correlation coefficient, respectively. No evidence was found in this sample that the initial cleft area had any bearing on the quality of outcome at 6 years of age. PMID- 10721251 TI - Magnetic resonance imaging in temporal lobe epilepsy: usefulness for the etiological diagnosis of temporal lobe epilepsy. AB - With improvement in magnetic resonance (MR) imaging techniques, the ability to identify lesions responsible for temporal lobe epilepsy has increased. MR imaging has also enabled the in vivo diagnosis of hippocampal sclerosis. Brain tumors are responsible for 2-4% of epilepsies in adult population and 10-20% of medically intractable epilepsy. The sensitivity of MR imaging in the diagnosis of tumors and other lesions of the temporal lobe (vascular malformations, etc.) is around 90%. Both hippocampal sclerosis and other temporal lobe lesions are amenable to surgical therapy with excellent postsurgical seizure outcome. In this article, we characterize and underline distinguishing features of the different pathological entities. We also suggest an approach to reviewing the MR images of an epileptic patient. PMID- 10721252 TI - Changes in local cerebral blood flow, glucose utilization, and mitochondrial function following traumatic brain injury in rats. AB - The pathophysiology of secondary brain damage following experimental traumatic brain injury was investigated by measuring local cerebral blood flow (lCBF), local cerebral glucose utilization (lCGU), and activity of succinate dehydrogenase (SDH), which is a mitochondrial enzyme of the tricarboxylic acid cycle, in the rat brain after moderate lateral fluid percussion injury. Measurements used autoradiography for lCBF and lCGU with [14C]iodoantipyrine and [14C]2-deoxyglucose, respectively. Regional SDH activity was determined using quantitative imaging of formazan produced from 2,3,5-triphenyl tetrazolium chloride by SDH. lCBF decreased at 1 hour after injury and was significantly lower than the preinjury level in almost all regions of both hemispheres at 6 and 24 hours, and remained low at 2 weeks. lCGU increased 1 hour after injury but was significantly decreased at 6 and 24 hours, and at 2 weeks in most regions of both hemispheres. The ipsilateral hemisphere showed a significant decrease in the activity of SDH in the cortices, hippocampus, thalamus, and caudate/putamen, most conspicuously 72 hours after injury, whereas no significant decrease was observed in the contralateral hemisphere at any time. Necrosis in the injured cortex and reduction of the number of neurons in the ipsilateral hippocampus were observed 2 weeks after injury. The present study showed that a decrease in lCBF and mitochondrial dysfunction occur with glucose hypermetabolism around 1 hour after lateral fluid percussion injury, and that lCBF, lCGU, and mitochondrial function all deteriorate after 6 hours. This suggests that lCBF and cellular metabolism may change dynamically during the several hours following traumatic brain injury, and afterwards neuronal damage may result in an irreversible change in the areas with depressed glucose hypermetabolism in the early period after injury in combination with mitochondrial dysfunction. PMID- 10721253 TI - Use of microvascular Doppler sonography in aneurysm surgery on the anterior choroidal artery. AB - Anterior choroidal artery (AChA) syndrome is still one of the most serious complications of the clipping of internal carotid artery aneurysms. No monitoring method can detect ischemia in the area of the AChA during surgery. This artery may be obstructed when a clip is applied to the neck of the aneurysm, and patency is sometimes difficult to confirm by microscopy because of the artery's small size and site of origin (usually behind the internal carotid artery as viewed surgically). However, microvascular Doppler sonography (MVDS) can detect flow instantaneously even in such a small vessel. In our series, AChA syndrome occurred in three of 19 patients treated for AChA aneurysm before the introduction of MVDS, but only one of 19 patients treated with the aid of this device. In that patient, one of the two AChA branches was intentionally sacrificed by applying a clip to the prematurely ruptured aneurysm. MVDS detected hypoperfusion of the AChA after clipping in five other patients, and so the clip was readjusted to preserve AChA flow. Use of MVDS is very effective to prevent inadvertent injury to the AChA during aneurysm surgery on this artery. PMID- 10721254 TI - Indications for shunting in patients with idiopathic normal pressure hydrocephalus presenting with dementia and brain atrophy (atypical idiopathic normal pressure hydrocephalus). AB - The indications for shunt operation in patients with idiopathic normal pressure hydrocephalus accompanied by brain atrophy (atypical idiopathic normal pressure hydrocephalus: AINPH) were investigated in 25 patients who satisfied the diagnostic criteria and underwent ventriculoperitoneal (VP) shunting. All patients had no apparent history of intra- or extracranial disease; dementia and gait disturbance as the main complaints; moderate to severe cerebral atrophy and ventricular dilatation and at least periventricular low density around the anterior horn on computed tomography; normal cerebrospinal fluid (CSF) pressure and filling of ventricles or cortical surface space with contrast medium at 24 hours on cisternography. The 15 male and 10 female patients were aged 47-83 years (mean 60.4 years). VP shunting was effective in 12 improved patients and not effective in 13 unimproved patients according to NPH grading. Pathological pressure wave on epidural pressure monitoring was observed in eight of 12 improved patients, but none of 13 unimproved patients. CSF outflow resistance was 35.33 +/- 11.16 mmHg/ml/min in improved patients and 9.12 +/- 3.51 mmHg/ml/min in unimproved patients. Preoperative serum alpha-1-antichymotrypsin value (alpha-1 ACT) was 42.02 +/- 8.64 mg/dl in improved patients and 61.72 +/- 11.03 mg/dl in unimproved patients. Alpha-1-ACT over 55 mg/dl occurred only in unimproved patients. Cerebral arteriovenous difference of oxygen content value (c-AVDO2) before and after surgery was 6.34 +/- 0.9 ml% and 5.91 +/- 0.78 ml% in improved patients and 4.75 +/- 1.85 ml% and 4.81 +/- 1.73 ml% in unimproved patients, respectively. The two cases with preoperative c-AVDO2 value over 8.5 ml% were both unimproved. Mean cerebral blood flow value before and after surgery was 23.51 +/- 4.20 ml/100 g/min and 45.22 +/- 8.11 ml/100 g/min in improved patients and 21.77 +/- 5.12 ml/100 g/min and 24.82 +/- 4.97 ml/100 g/min in unimproved patients, respectively. Cerebral atrophy in improved patients is caused by a cerebral circulation disturbance defined as a cerebral blood flow of penumbra or more due to cerebral arteriosclerosis, etc. A flow-chart of indications of shunt surgery for AINPH was prepared based on the results of the present study. PMID- 10721255 TI - Far lateral approach for foramen magnum lesions. AB - Twelve patients with lesions in the anterior or anterolateral regions of foramen magnum were treated through the far lateral approach. The patients presented with neck pain, dysesthesia, quadriparesis, numbness, respiratory distress, and spastic contractures. Most lesions were meningiomas and neurofibromas, with one patient each with a posterior inferior cerebellar artery aneurysm, neurenteric cyst, and chordoma. All mass lesions were excised totally and the aneurysm was clipped. Three patients had severe respiratory problems preoperatively and two of them died. The other patients made a satisfactory neurological recovery. It was not found necessary to resect the condyle or mobilize the vertebral artery in any of the patients. PMID- 10721256 TI - Intraventricular aneurysms--three case reports. AB - Three rare cases of purely intraventricular aneurysms are described, including a unique aneurysm in the fourth ventricle. A 30-year-old female, a 47-year-old male, and an 11-year-old girl presented with disturbance of consciousness due to massive intraventricular hemorrhage. Digital subtraction angiography revealed an idiopathic peripheral aneurysm in the fourth ventricle in the first patient, and aneurysms in the lateral ventricle associated with moyamoya disease in the latter two patients. The former two aneurysms were treated surgically and histologically confirmed to be pseudoaneurysms. The latter aneurysm disappeared spontaneously within 2 months after onset. The aneurysm in the lateral ventricle was resected via a parietal corticotomy with stereotactic insertion of an 8-Fr silicone tube to guide the approach route. This method was very useful because computerized neuronavigation was not available. The aneurysm in the fourth ventricle was resected via a midline suboccipital approach with C-1 laminectomy. Conservative treatment is usually recommended initially for patients with intraventricular aneurysm because spontaneous cure often occurs. We recommend direct surgery if the size of the aneurysm remains unchanged, because the risk of surgery has decreased recently owing to new techniques for neuronavigation. PMID- 10721257 TI - Transvenous embolization of carotid-cavernous sinus fistula associated with a primitive trigeminal artery--case report. AB - A 58-year-old female presented with right conjunctival chemosis and right abducens nerve paresis. Cerebral angiography demonstrated a right carotid cavernous sinus fistula associated with persistent primitive trigeminal artery. The fistula was treated by introducing detachable coils through the transvenous approach, as the detachable balloon was not available. Follow-up angiography performed 14 days after the embolization revealed complete disappearance of the carotid-cavernous sinus fistula due to thrombosis, which was presumably accelerated by the coils. Transvenous coil embolization should be considered as an alternative treatment for high-flow carotid-cavernous sinus fistula, but only if transarterial balloon embolization is not successful or unavailable. PMID- 10721259 TI - Hemangioblastomas with blood supply from the dural arteries--two case reports. AB - Hemangioblastomas are benign vascular tumors that often occur in the cerebellum, and are located near the pia mater. The blood supply is usually received through the pia mater, and rarely through the external carotid artery. The present cases of hemangioblastoma received blood supply from the external carotid artery (occipital artery) and a branch of the internal carotid artery (carotico-tympanic artery or artery of Bernasconi Cassinari) through the dural branches. The dural arteries were not the main feeders in either case, but preoperative embolization of the occipital artery contributed to minimum bleeding during the operation in one case. Incomplete resection of hemangioblastoma is related to multicentricity of the tumors, small mural nodules, or brain stem involvement. Angiography is valuable for demonstrating arterial supply to small or multiple mural nodules. Conventional angiography is necessary for investigation of the external carotid artery branches. PMID- 10721258 TI - Crossed cerebellar hyperperfusion in symptomatic epilepsy--two case reports. AB - A 74-year-old female with cerebral infarction and a 78-year-old female with cerebral glioblastoma suffered complex partial seizure. Ictal perfusion single photon emission computed tomography in these patients showed the interesting phenomenon of 'crossed cerebellar hyperperfusion,' a reversed crossed cerebellar diaschisis. The mechanism is probably spread of electrical seizure through efferent projections, and may be related to the cerebellar atrophy seen in patients with long-standing partial epilepsy. PMID- 10721260 TI - Epidermoid tumor within Meckel's cave--case report. AB - A rare case of an epidermoid tumor lying within Meckel's cave is reported. A 27 year-old housewife presented with complaints of right facial hypesthesia for two and a half years. On examination she had partial loss of touch sensation in the right trigeminal nerve distribution. Magnetic resonance imaging revealed a tumor located at the right petrous apex and cavernous sinus. The epidermoid tumor was excised through a lateral basal subtemporal approach. The symptoms resolved following surgery. PMID- 10721261 TI - Polypropylene mesh substitute for the fascial defect after using for the dural repair--technical note. AB - Use of the anterior sheath of the rectus abdominis muscle (anterior sheath) as a dural substitute and patching of the large defect of the anterior sheath with polypropylene mesh are described. Five patients were treated using the anterior sheath and the mesh. No postoperative complications such as cerebrospinal fluid leakage, infection, or abdominal wall hernia occurred. The mesh is useful as a patch for the sheath defect. PMID- 10721262 TI - Gap junctional intercellular communication and carcinogenesis. AB - Intercellular communications are indispensable for the maintenance of homeostasis in the multicellular organisms. Gap-type junctions are one of the most common and perhaps most interesting intercellular connections. Of various communication systems gap junctional intercellular communication is the only means for cells to directly exchange signals. Substantial progress has recently been made in the studies on the role of gap junctions both in experimental and human tumorigenesis. The present study is a review of the potential mechanisms associated with gap junctional cellular communication included in non-genotoxic tumorigenesis. PMID- 10721263 TI - Nuclear morphometry of MIB-1 positive and negative tumor cells in primary and metastatic malignant melanoma of the skin. AB - The purpose of this study was the evaluation of nuclear area, nuclear axis ratio, perimeter and roundness of nuclei of tumor cells with and without Ki-67 antigen expression (demonstrated immunohistochemically using MIB-1 antibody) in primary and metastatic malignant melanoma of the skin. The parameters were further analyzed with respect to their association with the depth of malignant invasion according to Clark [7] and tumor thickness according to Breslow [6]. 142 malignant melanomas (53 primary and 89 metastatic), were assessed employing a computerized image analyzer Quantimet 600S (Leica). The mean nuclear area of MIB 1 positive nuclei was significantly larger than that of the negative ones (p < 0.0001) both in primary and metastatic malignant melanoma. In comparison to the primary melanoma the nuclei of metastatic melanoma cells had a larger area and were more rounded, while the MIB-1 positive nuclei additionally showed a greater degree of polymorphism of their area and shape. With growing invasion thickness according to Breslow and increased Clark's level, the mean nuclear area of tumor cells increased, and their shape became more round. The MIB-1 positive tumor cell nuclei of primary melanomas with metastases were significantly out of round in comparison to primary melanomas without metastases. The results indicate an association between the area and shape of melanoma cell nuclei and the presence of metastases, and between the nuclear area of tumor cells and such factors related to poor prognosis as the depth of invasion and the tumor thickness. PMID- 10721264 TI - Cytogenetic and proliferative potentials in meningiomas. AB - Cytogenetic analysis and Ki-67 staining index (SI) were performed on the series of 51 meningiomas, to estimate the relationship between the extent of chromosomal changes and the growth potential of tumors. The tumors were classified according to histological subtype (22 meningiotheliomatous, 15 transitional, 12 fibroblastic and 2 angiomatous) and grade (40 benign, 5 atypical and 6 malignant ones). There was no significant difference in the complexity of chromosomal changes among the histologically distinct tumor subtypes. In contrast, there was a significant association between the number of chromosomal abnormalities and tumor grade. Normal karyotypes were found in 50% and complex in 20% of benign tumors. All grade II or III tumors revealed complex karotype. The tumors classified as benign revealed significantly lower mean Ki-67 SI than atypical or malignant ones (1.6%, 7.4% and 14.7%, respectively). However, within tumors classified as benign, mean Ki-67 SI of these with normal or simple karyotypic changes did not differ significantly from those with complex karyotype (1.6% and 0.9%, respectively). Thus, the extent of genome abnormalities in meningiomas, measured on the chromosomal level, does not relate directly to their proliferative potential. PMID- 10721265 TI - Studies on tissue expression of HBV in children with chronic hepatitis type B using Immunomax technique. AB - The study aimed at employing the Immunomax technique to detect the markers of HBV replication (HBcAg and HBV-DNA) in liver biopsy material, obtained from children with chronic hepatitis type B. In line with the currently modified classification of chronic hepatitis and with the increasing potential of antiviral therapy it seemed purposeful to supplement routine staining techniques with studies at the molecular level. Our studies demonstrated the effective detection of both the core antigen and HBV-DNA in liver tissue in children using immunocytochemical techniques and in situ hybridization, amplified with the Immunomax technique. HBcAg was detected in 26 out of 27 liver biopsies from patients with chronic hepatitis type B and with replication of the virus. HBV-DNA was detected in all study children with HBV infection and in 2 out of 5 cases of chronic hepatitis of a distinct etiology. No significant relationships could be found between the detection of tissue HBV markers on the one hand and the intensity of inflammatory lesions or severity of fibrosis on the other. PMID- 10721266 TI - BN52021 inhibits activation of peritoneal mast cells caused by hemorrhage and blood volume restoration. AB - An extensive blood loss activates generalized inflammatory response. Abdominal organs and especially intestines are very sensitive to the ischemia-reperfusion insults due to hemorrhagic shock (HS) and blood volume restoration. Previously obtained results suggest that studies on peritoneal lavage fluid (PLF) can contribute to elucidation of inflammatory processes in abdominal organs in HS. Histamine (H) levels, total cell, and mast cell (MC) numbers, and MC ultrastructure in the fluid lavaged from peritoneal cavity were compared in the following groups of rats: control (gr. 1), sham operation (gr. 2), untreated hemorrhagic shock (gr. 3), shock treated with blood volume restoration with lactated Ringer's solution (LR) (gr. 4), shock treated with platelet activating factor (PAF)-receptor antagonist Ginkgolide B (BN52021), and LR (gr. 5). A shock related significant increase in total cell numbers, MC numbers, MC degranulation, and histamine levels in PLF were observed. The restoration of blood volume caused further elevation of the above phenomena (gr. 4) while BN52021 seemed to inhibit peritoneal MC mobilization and degranulation as well as to attenuate increase in peritoneal H level (gr. 5). The peritoneal cavity is a place of rapid and strong reaction to hemorrhage. Evaluation of peritoneal histamine levels might be helpful in the monitoring of shock dependent intra-abdominal processes. Peritoneal MC mobilization and degranulation, and increase in histamine level is inhibited by BN52021. PMID- 10721267 TI - Ultrastructural features of immaturity in blood vessels of neonatal rat spinal cord. AB - Ultrastructural study was carried out on the lumbo-sacral segment of the spinal cord in 1, 3, 6, 9, 12, 15, 18, 21 and 25-day-old rats. Unusual features of blood vessels in postnatal life of rats were observed: capillaries with abundant endothelial cells, numerous invaginations and narrow lumen. In addition, microvilli of diverse size and number on the luminal surface of capillaries as well as larger vessels were present. They were more numerous in younger animals. An intriguing finding, not observed in older animals, was the appearance of abluminal protrusions projecting into surrounding tissue. Closed endothelial junctions were noted in all the investigated vessels. The above mentioned ultrastructural features indicate an immature character of blood vessels. The presence of this kind of vessels during the postnatal period may be connected with vascularisation of spinal cord due to the myelination process. PMID- 10721268 TI - Development of the fetal and newborn lung--morphometric studies (rat model). AB - The aim of this study was to investigate the development of the fetal lung by morphometric means. Lung specimens from 16-20 day-old fetuses, and 1, 3, 5, 14 and 21 day-old newborns were used. Tissue specimens were evaluated by light microscopy and transmission electron microscopy. The material was divided into four groups--related to four main stages of lung development (pseudoglandular, tubular, saccular and alveolar). For each developmental stage, 30 photographs of semi-thin sections were saved on computer hard disk for future morphometric measurements. OPTIMAS 4.02 software was used for evaluation of the relative area occupied by vascular, epithelial, stromal and fluid/gaseous compartments. GraphPad InStat software was used for statistical analysis. All measurements and calculations were done per observation field. The area occupied by vessels was greatest at the pseudoglandular stage (11.7%), and smallest at alveolar stage (5.7%). The stroma also occupied the largest area at the pseudoglandular stage (58.4%) and the smallest at alveolar stage (17.0%). The area occupied by epithelial cells was most extensive at the tubular stage (52.5%) and smallest at the alveolar stage (6.1%). The fluid/gas compartment was largest at the alveolar stage (70.5%) and smallest at the pseudoglandular stage (0.58%). The present results indicate that the most dynamic lung development occurs during thin air blood barrier formation (19th day of intrauterine life to 5th day after delivery- saccular and the beginning of alveolar stage). PMID- 10721270 TI - Pleomorphic adenoma (benign "mixed" tumor) of the human female breast. Case report. AB - A case of solitary pleomorphic adenoma, ("mixed" tumor of salivary gland type) of the left breast associated with the right breast fibroadenoma in 43-year-old woman is reported. The paper describes clinical, cytological, immunohistological and pathological findings in this case and indicates the importance of separating this benign entity from malignances with stromal metaplasia. PMID- 10721269 TI - Effect of Helicobacter pylori infection, smoking and dietary habits on the occurrence of antrum intestinal metaplasia. Clinico-epidemiological study in Poland. AB - The purpose of the study was to assess risk factors for intestinal metaplasia arising from H. pylori-related chronic gastritis in a subset of the population referred to endoscopic examinations due to dyspeptic complaints. We aimed specifically to establish whether H. pylori itself may be responsible for the occurrence of intestinal metaplasia and to which extent the metaplasia may be associated with life style factors such as cigarette smoking, alcohol consumption or dietary habits. The study was carried out in a sample of 1290 outpatients referred for the first time to gastroenterologic outpatient clinics in 6 university centers in Poland. The study methods covered standardized health interviews, endoscopy and histology of gastric antral specimens taken at endoscopy. The interviews performed by trained interviewers sought information on tobacco and alcohol intake, diet, socioeconomic status, and other variables. In non-ulcer dyspepsia subjects there was 54.9% H. pylori related gastritis and 25.1% of non-H. pylori-related gastritis. The corresponding rates in the group of ulcer dyspepsia were 67.5% and 20.5%. The increased risk of chronic gastritis in antrum was associated with Helicobacter pylori infection (OR = 2.28; 95% CI:1.93 2.69), and with gastric peptic ulcer (OR = 1.88; 95% CI:1.20-2.94). In the non ulcer dyspepsia the prevalence of metaplasia was 11.1% and in ulcer dyspepsia 19.7%. The risk of intestinal metaplasia within antrum depended greatly upon the presence of gastric peptic ulcer (OR = 3.85; 95% CI:2.35-6.32) and increased with age (OR = 1.05; 95% CI:1.04-1.07), smoking cigarettes currently or in the past (OR = 1.42; 95% CI:1.10-1.84), higher frequency of drinking vodka (OR = 1.32, 95% CI:1.01-1.75) and antral chronic gastritis (OR = 1.31; 95% CI:1.00-1.70), however, it was inversely related to daily consumption of fresh fruits or vegetables (OR = 0.59; 95% CI:0.38-0.93). The results of the study suggest that there is no sufficient evidence supporting the hypothesis about an association between H. pylori gastritis and intestinal metaplasia, however, the transition of gastritis to metaplasia depends greatly on life style factors such as cigarette smoking or vodka drinking and is impeded by daily consumption of fresh fruits or vegetables. PMID- 10721271 TI - Hyalinizing trabecular adenoma of the thyroid gland--a report of two cases. AB - Hyalinizing trabecular adenoma (HTA) is a specific variant of thyroid adenoma with a particular histological pattern. Immunohistochemistry is the best way of differentiating these adenomas. Of special interest is a strong positive cytoplasmic reaction for MIB-1. In FNAB and intra-operative examination HTA is usually misdiagnosed as a papillary carcinoma. Histological examination of paraffin-embedded material and immunohistochemical stainings provide a correct diagnosis. PMID- 10721272 TI - [Separation anxiety and agoraphobia in 8-year-old children]. AB - In the literature on the aetiology of panic disorder, the separation-anxiety hypothesis is discussed, in which separation anxiety disorder is conceived as a precursor of panic disorder. Using the representative sample of the Dresden Child Anxiety Study (DKAS) we examined weather agoraphobic and separation anxiety symptoms do already co-occur systematically in eight-year-olds. After N = 826 children had been screened, N = 230 took part in an individual diagnostic interview. With a total prevalence of 9.5% for all anxiety disorders, the prevalence rate for separation anxiety amounted to 2.8%. Another 2.5% of the children (almost exclusively girls) received a diagnosis of specific phobia in agoraphobic situations. Separation anxiety, social anxiety, agoraphobic anxiety and panic anxiety were assessed on a dimensional level, as well. However, no specific pattern of co-morbidity could be found in terms of an increased frequency of agoraphobic fears and separation anxiety occurring simultaneously. The symptom profiles of children with separation anxiety and those with agoraphobic anxieties differed considerably. Children currently living in a separation situation do not exhibit separation anxiety or agoraphobia more frequently than the rest of the children. PMID- 10721273 TI - [Analysis of pragmatic aspects of communication behavior of verbal and nonverbal autistic children]. AB - The analysis and intervention of communication is an important focus of autism research. The present study is a microanalysis of the communicative behaviour of 10 autistic children with their parents and a therapist. Protests, appropriate initiation and responses of the children were analysed in relation to demands and the specific feedback of the adults. After 20 months of structured therapy changes in the communicative behaviour of the participants were demonstrated. Autistic children showed different communicative pattern with their parents compared to a therapist. The non-verbal group exhibited significantly more protests and decreased responsivity with their parents compared to the therapist. The verbal group interacted with their parents predominantly by echolalia. After 20 months a significant reduction in protests, increased compliance and responsivity were obvious in the non-verbal group. The verbal group showed a reduction in echolalia as well as increased responsive and spontaneous communication. The results demonstrate that even non-verbal autistic children are sensitive towards different interaction partners. Over the observation period participants showed a reduction in behaviour problems and positive developments of communicative behaviour. PMID- 10721274 TI - [From program to metaphor--managing the needs of children in a separation or divorce process involving their parents]. AB - In German-speaking regions there are several independent intervention programs, derived from concepts originating in the USA, which are designed to assist children whose parents are separated or divorced. Two of these programs will be presented here. Consistent with the thematic implication that divorce entails trauma and stress, the childrens' need of advice and help is placed at the conceptual center of the intervention. This results in a behavioral-cognitive training program, which should enable children to overcome the stress of their situation. In contrast to these two programs, an understanding of divorce may be achieved, through phenomenological analysis, which gives due consideration to the various aspects and meanings of the divorce process. The concept of intervention derived from this approach is to assist children in further developing their own search for solutions, focused on the parents-child-relation. The primary emphasis of this approach is not the childrens' difficulty in dealing with their parents' separation and divorce, but rather their own attempt to deal with the problem, as is visible in the metaphor of their spontaneous descriptions and images of experiences and events. The central concept of the proposed course is the further development of this creative process in the form of a dynamic-communicative group happening. Furthermore it is shown how children can be assisted in a practical way, and encouraged to create their individual and personally adequate solution to the experience of divorce. PMID- 10721276 TI - [Ethics and/or evidence-based psychotherapy--a challenge]. PMID- 10721275 TI - [Children and adolescents with mental disorders as orphans of managed care?]. AB - Failure of incentive to investigation in infantile pharmaceutical trials has lead to insufficient supply with drugs within pediatrics and child- and adolescent psychiatry in Germany. In practice, drugs that are approved for adults are used to treat minors as a remedy. In many areas such "off label use" has become the medical standard and is therefore owed by the doctor under medical malpractice law. The principle of economy, that rules the law of public health insurance, forbids the prescription of drugs, that are not tested. However, as minors have the right to equal protection, the principle of economy can not prohibit off label use to the determent of minors. By virtue of the European orphan drug direction we can expect an improvement of drug supply in the field of pediatrics and child- and adolescent psychiatry in the long run. But meanwhile it would be unethical and unlawful to deny minors through restrictions on drug prescription a standard treatment, where the medical standard demands off label use of a pharmaceutical product. PMID- 10721277 TI - [Sexual and physical abuse during early childhood or adolescence and later drug addiction]. AB - Sexual or physical abuse of children are discussed as possible causes or risk factors for psychiatric disorders like posttraumatic stress disorder, alcohol and drug addiction. The aim of this study was to identify possible differences between sexually or physically abused and non-abused patients with polytoxic drug abuse. METHOD: 100 patients with polytoxic drug abuse were interviewed during their therapy about a history of sexual abuse prior to the age of sixteen. Using different questionnaires we tried to find possible differences between drug users being sexually abused or not and risk factors for later drug addiction. RESULTS: 70% of the female and 56% of the male drug users had been sexually abused as children, 40% of the male and 50% of the female participants had a history of severe sexual abuse with sexual intercourse. In over 50% friends or relatives were the perpetrators committed the crime, in no case the parents had. More than 40% showed also a history of physical abuse. Significantly more drug users than alcohol abusers had a sexual trauma. Especially severe sexual abuse was associated with abuse of hard illegal drugs. Furthermore, we could find significantly more symptoms such as autoaggressive and suicidal behaviour, social isolation, reduced emotional binding to others, tendency to be persistently victimized, prostitution and violence against others in the group of sexually abused. Many of these symptoms are not only characteristic of addiction, but can be found also in other psychiatric diseases such as borderline and eating disorder. In conclusion, we could not find a significant correlation between sexual abuse and later drug addiction. 80% of the drug users themselves did not relate the fact of being sexually abused as child to later drug abuse. However, there seems to be a positive correlation between sexual abuse and a more severe addiction to illegal drugs as well as higher rates of symptoms with a negative course of the disease. For this group of patients with an unfavourable prognosis special therapeutic concepts are needed. PMID- 10721278 TI - [When does a study of different therapies allow comparisons of their relative efficacy?]. AB - Psychotherapy research is a form of evaluation research. Among many others, two types of evaluation can be distinguished: "comparative" and "non-comparative" evaluation studies. This differentiation, though its consequences are often neglected, is fundamental, since a study designed to answer a question concerning the non-comparative or "isolated" efficacy of a therapy cannot simultaneously serve to answer the question concerning the relative efficacy of two or more therapies aiming at the same goals or objectives--and vice versa. In this article, a small part of psychotherapy research, the 22 studies which have already been used by Grawe et al. (1994) in their comparisons of behaviour therapies and short-term psychodynamic therapies, is reanalyzed with respect to the two types of evaluation and to some consequences of this differentiation. RESULTS: On the whole, it is not possible to draw conclusions about the comparative efficacy of behaviour therapies and short-term psychodynamic therapies with regard to their specific goals, even if only some consequences of the distinction between two types of evaluation are taken into account. Among other reasons, this is due to the fact that the studies have not consequently been planned and executed as comparative evaluations. Only amelioration of the 22 studies can be regarded--with certain restrictions--as comparative outcome studies with respect to amelioration of certain symptoms. A further analysis of these studies shows that there is no evidence of a "highly significant" superiority of behaviour therapies over short-term psychodynamic therapies. PMID- 10721279 TI - [Narcissistic regulation and metabolic control in type-II diabetics]. AB - The influence of narcissistic regulation on metabolic control was investigated in 130 patients with Type-II-diabetes in the course of a rehabilitation programme. Two groups were studied: a) diabetics on a non-drug therapy regimen, b) diabetics treated by insulin. At the end of the rehabilitation programme 55.6% of the non insulin patients and 60.1% of the insulin treated patients exhibited a clinically significant improvement as measured by HbA1c. On the basis of the Dusseldorf Questionnaire for Narcissistic Regulation in Chronic Disease (DNACE) predictors of metabolic control were established. Using decision analysis, a correct prediction was possible in 73.1% resp. 85.4% of the cases. In the non-drug group, low values of the "helplessness/powerlessness" scale and high values of the "mourning" scale predicted an improvement of metabolic control at the end of the rehabilitation programme. On the other hand, in the insulin treated group the more successful patients were shown to have low values in the scales "distrust", "feeling of being ashamed of disease", "feeling of inferiority" and "fighting self-image" and high values in the scales "value ideal" and "symbiotic protection of the self". The results are discussed within the theoretical framework. PMID- 10721281 TI - [Our time with the Revista Clinica Espanola]. PMID- 10721280 TI - [Specific dimensions of anxiety in surgical patients. Development of a questionnaire and empirical results]. AB - This article reports on the construction and empirical evaluation of an instrument for the measurement of different components of surgery-related state anxiety. The anxiety inventory KASA identifies three components: cognitive, autonomic, and somatic anxiety reactions. Internal consistencies of the three scales related to these components were highly satisfactory (Cronbach's alpha between 0.80 and 0.91). An analysis of scores of these three components across the perioperative period demonstrates that the KASA is a sensitive indicator of state anxiety changes. Further analyses concerning external relationships of this new instrument with indicators of coping, trait anxiety, perioperative mood states and postoperative adjustment are reported. Results of these analyses indicate that the KASA is a reliable and valid measure of different components of surgery-related anxiety. PMID- 10721282 TI - [Gastric carcinoma associated with other primary malignant neoplasms. A retrospective study of 25 cases]. AB - BACKGROUND: The risk of developing a second neoplasm in a person with gastric carcinoma (GC) is higher than among general population. OBJECTIVE: To analyze the clinical findings in patients with GC associated with other primary malignant neoplasms. PATIENTS AND METHODS: A total of 25 patients with GC associated with extragastric tumours were retrospectively studied. The following characteristics were studied: age, sex, location and staging, free interval, therapy, and survival. Survival of 13 patients with GC diagnosed as primary tumour was compared with that observed in a control group of 62 patients with GC alone. RESULTS: Twenty-five out of 792 (3.1%) patients with GC had other primary malignant neoplasms (seven synchronous and 18 metachronous). GC was associated with respiratory tumours in 7 cases. Sixty percent of patients with GC who had a second neoplasm had it diagnosed within the first year after gastric tumour was diagnosed (8 out of 13). Survival at 18 months was similar, both in the GC group with a second tumour as in the control group. CONCLUSIONS: The development of a second neoplasm among patients with GC usually occurs within the first year after diagnosis. Most commonly, the second neoplasm seats in the respiratory tract. PMID- 10721283 TI - [Nevirapine induces abstinence symptoms in patients on a methadone maintenance program with HIV infection]. AB - The objective of this study was to investigate the potential inductive effect of nevirapine (NVP) with methadone. Eight patients on the methadone maintenance programme with anti-retroviral therapy because of their infection with HIV, well maintained with methadone and without symptoms of abstinence were studied. All were included in a study of measurement of plasma levels of methadone. Anti retroviral medication was changed, including NVP, and patients began with symptoms of abstinence 5 to 10 days later. Our results indicate an inductive effect of NVP on the methadone metabolism which caused symptoms of abstinence in all patients, which prompted an increase in the dose and plasma concentration of methadone was lost; patients continued with significantly low plasma levels (p < 0.01) after a therapy mean duration of 6.5 months, with no full recovery. PMID- 10721284 TI - [Invasive aspergillosis: a study of 20 cases]. AB - OBJECTIVE: To assess the epidemiology of invasive aspergillosis (IA) and the frequency of recognition of this clinical entity. PATIENTS AND METHODS: Retrospective analysis of patients with the diagnosis of IA in the last three years. The diagnostic criteria of the American Institute of Infectious Diseases Mycoses Group were followed. RESULTS: During this period, 20 patients were diagnosed of IA: 9 (45%) had a hematologic malignancy, 14 (70%) had received corticosteroids, five (25%) had neutropenia, and three (15%) had no factors for immunosuppression. The disease was suspected in 15 cases (75%). Aspergillus spp. was recovered from sputum samples of the 16 patients who had the sample obtained. Seventeen patients (85%) died, 12 of them in spite of receiving antifungal therapy. Time relapsed since the beginning of symptoms and therapy was 14 days. CONCLUSIONS: The proportion of patients without neutropenia or severe immunosuppression is higher than usually thought. IA is a clinical entity of difficult diagnosis and occasionally it is diagnosed only at post-mortem examination. The high sensitivity of sputum culture may be due to the selection of cases with more severe infections as stringent diagnostic criteria were used. To improve the prognosis of IA it is necessary to initiate antifungal therapy early in the course of the disease and therefore, a high suspicion index is required, not only of the immunocompromised but also of the immunocompetent patient. PMID- 10721285 TI - [Breast metastases from embryonal rhabdomyosarcoma: apropos 2 cases and a review of the literature]. AB - The presence of breast metastases is uncommon in oncology. In children and young adults, the most common type of primary tumour is rhabdomyosarcoma, and primary breast rhabdomyosarcomas are exceptional. Two cases are reported of breast metastatic rhabdomyosarcoma in two adolescents with several atypical characteristics in their presentation forms: the embryonal variety and the primary location at maxillary sinus. The cases reported in the literature up to now were reviewed, with a special emphasis on epidemiologic, clinical, diagnostic, and therapeutic data of this rare entity. PMID- 10721286 TI - [The influence of histopathological diagnosis on the clinical expression of localized Castleman's disease: apropos 2 cases]. AB - The localized Castleman's disease (CD) seemingly has a different clinical behaviour depending on whether the pathologic study is consistent with the plasma cell (PC) or with the hyaline-vascular types. We report here two cases of localized CD with associated analytical changes, although only the PC modality was associated with symptoms. Surgery proved effective, and the systemic picture disappeared in both cases, as well as clinical manifestations in the PC variant. PMID- 10721287 TI - [Skin and mucosal lesions following labial herpes]. PMID- 10721288 TI - [A 65-year-old woman with a pathological fracture of the humerus]. PMID- 10721289 TI - [An intracranial mass in a patient with HIV infection]. PMID- 10721290 TI - [Prolonged fever and a patchy spleen]. PMID- 10721291 TI - [Chest pain and asthenia]. PMID- 10721292 TI - [What is healthy nutrition?]. PMID- 10721293 TI - [About the misnamed "idiopathic hirsutism"]. PMID- 10721294 TI - [Arterial calcifications detected in mammographies]. PMID- 10721295 TI - [Hyperthyroidism and hypercalcemia associated with lithium treatment]. PMID- 10721296 TI - [The ABO erythrocytic antigenic system as a possible predisposing factor for the appearance of venous thrombotic pathology]. PMID- 10721297 TI - [Community-acquired Acinetobacter baumanii sepsis with a fatal evolution]. PMID- 10721298 TI - [Meningeal carcinomatosis with a normal cell count and biochemistry in the cerebrospinal fluid]. PMID- 10721299 TI - [Postoperative infection in hip and knee surgery: does a relationship exist between hypercoagulability and immunomodulation induced by the transfusion practice?]. PMID- 10721300 TI - [Prosthetic valve endocarditis due to Enterobacter cloacae]. PMID- 10721301 TI - [The oral rehabilitation of a female bulimia patient. A case report]. PMID- 10721303 TI - [Interdisciplinary health and illness--from the philosophical viewpoint]. PMID- 10721302 TI - [The fixed denture care of the periodontally reduced dentition]. PMID- 10721304 TI - [Health, illness, healing--an interdisciplinary perspective. From the academic medicine viewpoint]. PMID- 10721305 TI - [Interdisciplinary aspects of health and illness--from the complementary medicine viewpoint]. PMID- 10721306 TI - [Intuition: the bridge from healing technique to healing arts]. PMID- 10721307 TI - [Welfare and healing from the theological viewpoint]. PMID- 10721308 TI - [Salutogenesis]. PMID- 10721310 TI - [Modern therapeutic progress: tension field between ethics and technique]. PMID- 10721309 TI - [A multitude of therapeutic possibilities--from the viewpoint of the clinician]. AB - Modern medicine offers to patients and physicians an ever increasing multitude of diagnostic and therapeutic possibilities. Depending upon the individual viewpoint of the patient and the caregivers there may be different opinions about the "right" choice. This article reviews some of the potential problems and conflicts, which may arise from multiple diagnostic and therapeutic options. We feel, that it is of utmost importance, that the informed patient is involved in the decision process. PMID- 10721311 TI - MRI safety limits: is MRI safe or not? PMID- 10721312 TI - Renal angiography using carbon dioxide. AB - The many advantages of carbon dioxide (CO2) angiography in the investigation of renal arterial disease include an absence of both nephrotoxicity and allergic reactions. An automated delivery system facilitates injection of CO2 whilst ensuring that there is no contamination of the injection with air. We report our initial experience using a prospective study of this delivery system in 47 patients referred for renal angiography, and assess diagnostic image quality and adverse reactions to CO2 angiography using the automated delivery system. The majority (37/47; 79%) of angiograms were of diagnostic quality and there were no significant adverse reactions in response to the CO2 contrast agent. PMID- 10721313 TI - Paediatric sedation for CT scanning: the safety and efficacy of quinalbarbitone in a district general hospital setting. AB - The aim of this study was to review retrospectively the safety and efficacy of a paediatric sedation protocol in a district general hospital radiology department. 256 children attended for CT scanning over a 40-month period. 40 children required sedation and were given quinalbarbitone. 34 (85%) of this group were adequately sedated. Of the children who received quinalbarbitone, 35 were under 5 years of age. 32 of this group (91.4%) were adequately sedated. Failures in children under 5 years were all caused by problems with administration whilst failures in the older children were due to paradoxical excitement. No problems with respiratory depression were encountered. Sedation can be safely performed in a district general hospital radiology department if a structured protocol is adhered to. Quinalbarbitone is a safe, effective oral agent in children under the age of 5 years. PMID- 10721314 TI - Differential bore sheath breast biopsy--a pilot study of a new technique. AB - The increasing requirement for core biopsy in the diagnosis of early breast disease puts additional demands on radiologists' time and expertise. We present a new approach to percutaneous breast biopsy of impalpable lesions with core biopsy and simultaneous hook wire localization. The technique allows accurate localization of breast lesions and is unique in that it facilitates simultaneous hook wire localization. Percutaneous biopsy by this method was accurate in 9 of 11 patients. PMID- 10721315 TI - Differential effects of menopause and metabolic disease on trabecular and cortical bone assessed by peripheral quantitative computed tomography (pQCT). AB - The usefulness of peripheral quantitative computed tomography (pQCT) was investigated in the diagnosis of metabolic bone diseases, including osteoporosis, and especially in the different diagnostic values in trabecular and cortical components. The subjects were 460 Japanese women aged 20-86 years, including 318 healthy volunteers, 58 osteoporotics with fracture and 84 patients with diseases including amenorrhoea, steroid-induced osteoporosis, renal osteodystrophy (ROD) and primary hyperparathyroidism. Bone mineral density (BMD) was measured for more than 4 years in 74 of the healthy volunteers. BMD was measured by spinal QCT, dual X-ray absorptiometry (DXA) of the spine, radius, and heel, and pQCT of the radius and tibia. High resolution images were obtained for geometry of the radius. Radial pQCT showed a higher correlation with radial DXA than with spinal QCT, and spinal QCT showed a higher correlation with spinal DXA than with radial pQCT. The annual bone loss rates at predominantly trabecular bone sites were accelerated in both the axial and appendicular skeleton. In the fracture study, radial pQCT showed a higher odds ratio (OR = 4.4) than radial DXA, and cortical area ratio seemed to be a good predictor of fracture risk (OR = 5.2). Amenorrhoea and steroid-induced osteoporosis predominantly affected trabecular bone, ROD predominantly affected cortical bone and hyperparathyroidism affected both components, especially the cortical component. pQCT is useful for assessing both trabecular and cortical bone, to provide information on individual bone changes in metabolic bone disease and to estimate the risk of fracture. PMID- 10721316 TI - Comparison of an imaging heel quantitative ultrasound device (DTU-one) with densitometric and ultrasonic measurements. AB - The purpose of this study was to evaluate a new imaging ultrasound scanner for the heel, the DTU-one (Osteometer MediTech, Denmark), by comparing quantitative ultrasound (QUS) results with bone mineral density (BMD) of the heel and femur from dual X-ray absorptiometry (DXA), and by comparing the DTU-one with another QUS device, the UBA 575+. The regions of interest in the DXA heel scan were matched with the regions evaluated by the two QUS devices. 134 healthy and 16 osteoporotic women aged 30-84 years old were enrolled in the study. In vivo short term precision of the DTU-one for broadband ultrasound attenuation (BUA) and speed of sound (SOS) was 2.9% and 0.1%, respectively, and long-term precision was 3.8% and 0.2%, respectively. Highest correlations (r) between QUS and BMD measurements were achieved when comparing DTU-one results with BMD in matched regions of the DXA heel scan. Correlation coefficients (r) were 0.81 for BUA and SOS. Highest correlations with the UBA 575+ were 0.68 and 0.72, respectively. The comparison of BMD in different femoral sites with BUA and SOS (DTU-one) varied from 0.62 to 0.69 when including the entire study population. The correlation between BMD values within different sites of the femur tended to be higher (from r = 0.81 to 0.93). When comparing BUA with BUA and SOS with SOS on the two QUS devices, the absolute QUS values differed significantly. However, correlations were relatively high, with 0.76 for BUA and 0.82 for SOS. In conclusion, the results of the new quantitative ultrasound device, the DTU-one, are highly correlated (r = 0.8) with results obtained using the UBA 575+ and with BMD in the heel. The precision of the DTU-one is comparable to other QUS devices for BUA and is high for SOS. PMID- 10721317 TI - Comparison of bone mineral density and quantitative ultrasound of the calcaneus: site-matched correlation and discrimination of axial BMD status. AB - The performance of quantitative ultrasound (QUS) and dual energy X-ray absorptiometry (DXA) bone densitometry of the calcaneus have been compared, both in terms of site-matched correlation and their discriminatory ability to identify osteoporotic and osteoporotic or osteopenic subjects. 91 female subjects (aged 56.9 +/- 9.6 years, 31-84 years) who were routinely referred for axial BMD assessment of the lumbar spine and femoral neck by DXA (Lunar DPX-L), consented to have additional measurements of QUS (McCue CubaClinical Mk II) and DXA (Lunar PIXI) of their left calcaneus. The site-matched correlation between calcaneal BMD with QUS parameters were: (a) broadband ultrasound attenuation (BUA) alone, adj R2 = 62.7%, p < 0.0001; (b) velocity (VOS) alone, adj-R2 = 48.4%, p < 0.0001; and (c) BUA and VOS combined, adj-R2 = 65.2%, p < 0.0001. The site-matched correlations are towards the higher end of data reported by other researchers, indicative of the exacting measurement protocol implemented here. 30 subjects were categorized as normal, 38 being osteopenic and 23 being osteoporotic. Optimum accuracies and odds ratios were obtained using logistic regression. The differences in accuracy between calcaneal BMD and calcaneal QUS parameters were statistically insignificant, with zero included within the confidence intervals, for the identification of both (a) osteoporotic and (b) osteoporotic or osteopenic subjects. The odds ratios for the discrimination of subject status achieved with calcaneal BMD were higher, although statistically insignificant, than achieved with the QUS parameters. Receiver operator characteristic (ROC) analysis for the identification of subjects into the categories (a) and (b) above was performed. Areas under the ROC curve (AUC) (95% confidence intervals) for the logit of the probability that a subject would be osteoporotic were: 0.814 (0.700, 0.928) for calcaneal BMD; 0.791 (0.673, 0.909) for BUA; 0.717 (0.588, 0.846) for VOS; and 0.793 (0.675, 0.911) for BUA and VOS combined. For the identification of osteoporotic or osteopenic subjects, the ROC areas were: 0.851 (0.774, 0.928) for calcaneal BMD; 0.773 (0.678, 0.868) for BUA; 0.783 (0.690, 0.877) for VOS; and 0.778 (0.685, 0.871) for BUA and VOS combined. Again, calcaneal BMD provided a higher, yet statistically insignificant, AUC than any QUS parameter. In conclusion, for the identification of subjects defined by World Health Organization criteria for axial BMD, the performance of BMD and QUS calcaneal parameters were statistically comparable. The choice of peripheral bone densitometry modality should therefore be made upon factors external to their discriminatory performance. PMID- 10721318 TI - Skin sparing in interventional radiology: the effect of copper filtration. AB - Complex and lengthy interventional radiological techniques have resulted in a number of patients developing skin reactions in recent years. To safeguard against these side effects, we have investigated the degree to which entrance skin dose can be reduced by inserting 0.18 mm and 0.35 mm copper filtration in the incident beam. The potential reduction was measured on a 22 cm water phantom for each of eight models of a fluoroscopy unit. Using the catheter laboratory fluoroscopy unit on which radiofrequency ablations are routinely performed, we assessed the relative effectiveness of adding filtration and increasing the kV:mA ratio. Image quality was subjectively assessed for diagnostic and therapeutic acceptability in two groups of 10 patients undergoing radiofrequency ablations, pacemaker insertions or electrophysiology studies. One of the groups was screened with 0.35 mm copper filtration in place and the other group acted as the control. Maximum patient skin dose proved difficult to measure directly because of the unpredictable dose pattern. This pattern was studied in four patients using a film method in conjunction with thermoluminescent dosemeters. Copper filtration 0.35 mm thick inserted in the beams of the eight fluoroscopy units produced a mean reduction in entrance dose to the phantom of 58% with a mean increase in tube loading of 29%. At 100 kV the increased loading on the X-ray tube was equivalent to increasing the anteroposterior separation of the patient by 2 cm. Measurements on the catheter laboratory unit showed that the tube voltage would need to be raised above the normal diagnostic range to obtain an equivalent entrance dose reduction without the filter. The blackening of films under the patients showed complex patterns, but the estimated skin doses were consistent with those predicted by the phantom experiments. All six cardiologists considered there to be insignificant detriment to image quality in the procedures investigated. PMID- 10721319 TI - Application of draft European Commission reference levels to a regional CT dose survey. AB - A survey of CT doses in Northern Ireland in the period between October 1995 and March 1997 was carried out. The survey included all but one of the 10 scanners in use at the time, and, additionally, two others that were replacement machines. The method used was to study standard protocols and calculate doses to the NRPB mathematical phantom, so that a direct comparison could be made with other surveys carried out in a similar fashion elsewhere. The survey addressed the patient radiation dose but not image quality or clinical outcomes. It is estimated that in Northern Ireland the contribution to collective dose to the population from CT is about 40% of that from all medical X-rays. The proposed European Commission reference quantities, weighted CT dose index and dose-length product were computed and their potential use evaluated. A full study of mean values of effective dose per examination revealed the average dose per examination was not significantly different from that found in the 1989 UK survey, although several procedures gave rise to doses that were high enough to be investigated with a view to justification or reduction. One of the scanners was found to give consistently high doses. It is likely that a revision of the mAs values used on this scanner will produce a significant reduction in patient doses without compromising image quality. When compared with the draft EC reference levels, fewer procedures were found to have excessively high dose values. The proposed EC reference levels would therefore be useful for continual monitoring of CT dose status, but do not appear to provide as comprehensive an assessment of patient exposure as that given by consideration of effective doses. PMID- 10721320 TI - Disappearance of stress-induced hyperthermia following a low dose of X irradiation: involvement of the vomeronasal system in the modulation of the radiation-induced effects. AB - When the rectal temperatures of group-housed mice are measured sequentially, the temperature of the last mouse to be measured is higher than that of the first mouse. The hyperthermia effect observed in the last animal to be measured forms the basis of an experimental paradigm for studying the neurobiology of anticipatory anxiety. Stress-induced hyperthermia (SIH) was calculated as the difference (delta T) between the basal temperature (the averages of the first three mice) and the final temperature (the averages of the last three mice) when the temperatures of the 15 mice were measured sequentially, with a 2 min interval between each temperature measurement. The hyperthermia observed in the last animals measured was abolished by prior treatment with X-irradiation at the relatively low dose of 5-15 cGy. Prevention of the SIH response could be found when the irradiation was confined to the head region only, suggesting the importance of the brain in the radiation-induced effect. Relatively higher doses of 25 or 35 cGy failed to reduce the hyperthermia stress effect. Furthermore, the effect of X-irradiation was not observed following olfactory bulbectomy or resection of the vomeronasal tract. These results indicate that the disappearance of SIH response may only be found following irradiation at low dose levels. Furthermore, the results implicate the olfactory system in the radiation-induced anti-stress effect. PMID- 10721321 TI - Dose response characteristics and basic dose distribution data for a polymerization-based dosemeter gel evaluated using MR. AB - A safe and reproducible mixing procedure for the manufacture of a polymerization based dosemeter gel evaluated using MRI (PoMRI) is presented. The dose response, obtained by irradiating gel-filled vials with absorbed doses in the interval 0-20 Gy and evaluated with respect to 1/T2, was found to be linear in the interval 0-8 Gy, with a sensitivity of 0.211 s-1Gy-1 (r2 = 0.998) at 1.5 T. Evaluation of the same set of vials with respect to 1/T1 gave a sensitivity of 0.018 s-1Gy-1 (r2 = 0.960). PoMRI and diode data were compared for standard photon and electron treatment beams. A deviation of less than 3% was found between the two methods for central depth dose curves as well as dose profiles (2 mm for electrons in the steep dose gradient regions). The importance of the method used for background correction for the reliability of the results was also evaluated. Barex (with a wall thickness of 1.5 mm) was investigated for use as phantom material and found to be favourable compared with glass. The results obtained in this study show that PoMRI has excellent potential as a 3D detector. PMID- 10721322 TI - Intrathoracic extension of aggressive fibromatoses. AB - Intrathoracic extension or primary thoracic origin of fibromatoses is distinctly uncommon. We describe the imaging features in three patients with this condition. PMID- 10721323 TI - Issues of threshold selection when determining the fractal dimension in HRCT slices of lumbar vertebrae. AB - The box counting dimension is a frequently applied tool for the classification of trabecular bone structure. The algorithm requires a binarization of the gray value data, for example that acquired by high resolution CT (HRCT). We recently proposed a method to eliminate bone mineral density (BMD) by applying a linear normalization scheme. Further consideration has shown that full BMD independence has not been achieved, and the structural parameter proposed was therefore difficult to interpret. In this study we present an alternative approach to obtain a structural parameter that is independent of BMD. HRCT volume data was acquired on 21 lumbar vertebrae from five cadavers. In the segmented spongiosa, thresholding was based on different quantiles of the gray value histogram, yielding invariance over linear and non-linear transformations. Thresholding at high gray value levels (80% quantile) shows the highest level of significance when discriminating between osteoporotic and non-osteoporotic cases. As an addition to the measurement of BMD alone, the determination of structural properties allows an improvement of the assessment of the individual fracture risk. PMID- 10721324 TI - Pseudomembranous necrotizing bronchial aspergillosis. AB - We report a case of pseudomembranous necrotizing bronchial aspergillosis in a patient with acute myelocytic leukaemia who died of massive haemoptysis. Lobar collapse was demonstrated on chest radiography. CT showed a marked necrotic thickening of the lobar bronchus with extension of the disease in to the peribronchial region. PMID- 10721325 TI - Thrombosis of mechanical heart valve prostheses: revisiting the role of fluoroscopy. AB - Prosthetic valve thrombosis is a rare but potentially fatal complication of heart valve replacement. Symptoms may be misleading, yet the condition may rapidly lead to death. A prompt diagnosis is therefore crucial. Ultrasound is the most often used technique for evaluating prosthesis dysfunction. We describe two cases of prosthesis thrombosis with negative Doppler results but with distinctly abnormal leaflet motion at fluoroscopy. A correct diagnosis would have been missed if the Doppler evaluation alone was relied on. Fluoroscopy should always form part of the diagnostic work-up in patients with artificial heart valves. PMID- 10721326 TI - Intraosseous venous drainage anomaly of the tibia treated with imaging-guided sclerotherapy. AB - A 23-year-old man presented with a pre-tibial soft tissue mass. Magnetic resonance images demonstrated the subcutaneous, intracortical and intramedullary components of an intraosseous venous drainage anomaly, which was confirmed by direct venography. Sclerotherapy using absolute alcohol was subsequently performed under imaging guidance with complete resolution of the subcutaneous component of the lesion. PMID- 10721327 TI - Pancreatic serous cystadenoma associated with islet cell tumour. AB - We report the case of a 29-year-old female patient with a diffuse type of serous cystadenoma involving the entire pancreas except for part of the head, which was replaced by islet cell tumour. Ultrasound and CT showed multiple cysts in the entire pancreas and a solid mass with calcification in the head. MRI characterized the fluid content of the cysts and the extent of disease. PMID- 10721328 TI - Struma ovarii: MRI findings. AB - We describe the MRI findings in three cases of struma ovarii. In all three cases, MRI showed a multilocular cystic mass with a variable signal intensity within loculi. Some loculi or small cysts within septations showed low signal intensity on T1 weighted images and very low signal intensity on T2 weighted images, corresponding pathologically to gelatinous colloid material in large follicles. In one case, with Gd-DTPA enhanced T1 weighted images, the thick septations and locally thickened wall showed marked enhancement, corresponding microscopically to thyroid tissue. PMID- 10721329 TI - The radiological appearances of tuberous sclerosis. AB - Tuberous sclerosis is an autosomal dominant disorder often associated with a chromosome 9 abnormality, although up to 60% of cases occur spontaneously. The incidence of the disorder is between 1/100,000 and 1/10,000, and it leads to multiple organ and skeletal abnormalities. The classical triad of epilepsy, mental retardation and adenoma sebaceum defines the syndrome clinically. Other cutaneous associations include shagreen patches, ash leaf-shaped areas of depigmentation, subungual fibromas and cafe-au-lait spots. This review describes the commoner radiological manifestations of the syndrome, and briefly mentions the rarer associations that have been reported to date. PMID- 10721330 TI - An inspiring case. PMID- 10721331 TI - Molecular dissection of Mycoplasma hominis. AB - M. hominis is commonly found as part of the normal flora in the female genital tract, but several studies have shown that it may be involved in a variety of urogenital infections. The basis for clinical manifestations in some patients has varyingly been attributed to host and M. hominis factors. The host factors involved in the infection process are largely unknown. M. hominis have no cell wall and outer membranes, and at present it seems plausible that M. hominis possesses genetic systems allowing the bacteria in vivo to alter its antigenic structure on the membrane surface and consequently circumvent the host immune system. The studies of M. hominis have shown that the antigenic variation is pronounced between surface exposed membrane proteins from different isolates. The genetic background for this variation has been investigated for three surface exposed membrane proteins: P120, Lmp, and Vaa. P120 and P120' are similar proteins in M. hominis without any homology to other known proteins. A hypervariable region in the otherwise conserved P120 protein seems to be very antigenic in patients with immunologically verified M. hominis infection. The remaining part of P120 as well as the entire P120' protein do not seem to elicit significant antibody formation. Two genes in M. hominis, lmp1 and lmp3, contain numerous highly similar 0.5 kb tandem repeats at their 3'-end. The proteins, Lmp1 and Lmp3, are synthesized from the lmp1 and lmp3 genes, respectively. Lmp1 shows size variation among M. hominis isolates. M. hominis isolates investigated in detail show that the size variation of Lmp1 corresponds to the variation in number of 0.5 kb repeats contained within the lmp1 gene. Lmp3 appears to have a lesser tendency to size variation. M. hominis isolates were found with deletions involving the lmp1 stop codon leading to translation of the downstream gene lmp2 and expression of a chimeric Lmp1-Lmp2 protein. The number of repeated elements in the lmp1 gene of a M. hominis isolate correlates with the extent of anti-Lmp antibody induced agglutination between the bacteria. Vaa is a protein involved in cell adherence. vaa is a single copy gene containing tandem repeated elements like the lmp gene family. The number of repeats in the Vaa protein differs between M. hominis isolates leading to size variation. It has been suggested that the number of repeated elements is of importance in the bacteria-host adhesion process. Beside the size variation Vaa demonstrates phase variation due to frequent frame shift mutation in a specific region near the 5'-end of the structural gene. Based on the investigations of M. hominis and other mycoplasmas several genetic mechanisms seem to be responsible for the antigenic variation of surface exposed membrane proteins in mycoplasmas: 1) variation in protein size due to insertions or deletion of repeated elements in the structural gene, 2) presence of multi-gene families, and 3) phase variation due to mutations in the promotor region or the coding region. The influence of specific antibodies on antigenic variation of membrane proteins has not been studied in greater detail in mycoplasmas. In M. hominis it was investigated whether the presence in the culture medium of monoclonal antibodies directed against the repeated elements in the M. hominis Lmp proteins would affect gene structure and consequently protein expression. The presence of anti-Lmp antibodies resulted in overgrowth of bacteria with specific deletions in the repeated elements of lmp1 leaving the lmp3 gene unchanged. The precise mechanism leading to the dominance of M. hominis isolates with fewer 0. (ABSTRACT TRUNCATED) PMID- 10721332 TI - Chemotherapy Foundation Symposium XVII: Innovative Cancer Therapy for Tomorrow. New York, New York, USA. November 3-6, 1999. Abstracts. PMID- 10721334 TI - William Osler: a model for the 21st century? PMID- 10721333 TI - Doctors and the "environment". PMID- 10721335 TI - Humour in medical teaching. PMID- 10721336 TI - The burden of disease and injury in Australia: time for action. PMID- 10721337 TI - Dead in the water: how safe are our water sports? PMID- 10721338 TI - Patterns of drowning in Australia, 1992-1997. AB - OBJECTIVE: To determine patterns of victims, circumstances and locations of drownings in Australia in 1992-1997, inclusive. METHODS: Population figures and available details of all drownings were obtained from the Australian Bureau of Statistics. Accidental non-boating drownings (ICD E910), boating incidents (E830 832), homicide (E964), suicide (E954), and other deaths without a drowning E code but "flagged" because drowning was involved (although not the primary cause of death) were included. RESULTS: The overall accidental non-boating drowning rate was 1.44/100,000 population/year. The commonest sites for non-boating drowning were ocean or estuary (22%), private swimming pools (17%), non-tidal lakes and lagoons (17%), surfing beach (10%) and bathtub (7%). 22% of victims were aged under 5 years; this group had a drowning rate of 4.6/100,000 population/year. Very few young children drowned in the ocean or in boating incidents. The rate of boating drownings was 0.29/100,000 population/year. Overseas tourists comprised 4.7% of all non-boating drownings, 18% of surf and ocean drownings, and 25% of all scuba drownings. Indigenous people had a much higher drowning rate than the general population. CONCLUSIONS: Drownings in children aged less than 5 years continue to be the greatest challenge for water safety organisations and legislators. Drownings in the Indigenous community and among tourists requires more detailed study and action. To assist in developing preventive strategies, the National Water Safety Council will need to clarify the categories described as "ocean/estuary" and "lake, lagoon, dam and waterhole". PMID- 10721339 TI - Snorkelling deaths in Australia, 1987-1996. AB - OBJECTIVE: To examine the causes and circumstances of snorkelling deaths in Australia from 1987 to 1996. DESIGN: Retrospective case extraction. CASES AND DATA SOURCES: 60 snorkelling deaths extracted from an ongoing diving fatality survey and from coroners' reports. Further details were obtained from police reports, diving industry (incorporating commercial operators, relevant government departments and instructors' organisations) inquiries and coronal inquests. MAIN OUTCOME MEASURES: Cause of death (determined by the authors from information obtained and from detailed autopsy findings) and the circumstances surrounding death. RESULTS: 15 of the 60 snorkellers who died were female. The three major causes of death were drowning (27 cases), cardiac events (18) and hypoxia with breath-holding after hyperventilation and/or during ascent producing unconsciousness then drowning (12). Overseas tourists were notable among those who drowned, while middle-aged men dominated the group who died of cardiac events (mostly on the surface). Those who died of breath-holding hypoxia were all young, Australian and male. The use of "buddy" diving was infrequent overall, and many of those who drowned or suffered cardiac events were not wearing flippers to aid propulsion. Adverse environmental conditions were implicated in 14 deaths. CONCLUSIONS: Hyperventilation to increase breath-hold time is a dangerous practice which should be discouraged. Safety measures, such as the use of flippers for propulsion and employment of the "buddy" system, should be encouraged, and made mandatory in commercial diving operations. PMID- 10721340 TI - Scuba diving medical examinations in practice: a postal survey. AB - OBJECTIVE: To assess variability of opinion regarding fitness to dive among doctors currently doing diving medical examinations. DESIGN: Anonymous, reply paid postal survey containing 15 clinical scenarios for which respondents were asked to declare the prospective scuba diver fit, unfit, fit after investigation or to offer specialist referral. PARTICIPANTS: All 81 doctors in Queensland, identified as members of the South Pacific Underwater Medical Society, who had completed approved training in underwater medicine, and who were doing diving medical examinations in June 1998. MAIN OUTCOME MEASURES: Variability in responses, and agreement with our interpretation of the action recommended by Australian Standard (AS) 4005.1-1992, the medical standard for fitness to scuba dive for recreational divers. RESULTS: 52 of the 81 questionnaires were returned (64% response rate). There was a wide variety of opinion about fitness to dive for all 15 hypothetical cases, with 70% consensus about unfitness in only four cases (one of which should have been referred according to AS guidelines) and fitness in only two cases (both of which should have been referred according to AS guidelines). No case was considered either fit or unfit by all respondents. Only 17.6% of responses recommended specialist referral, although the AS guidelines suggest that 10 of the 15 cases should be referred. One doctor failed 13 of the 15 potential divers outright and another passed seven outright and failed only four. For each case that the AS guidelines firmly indicate as unfit to dive, at least one respondent passed the hypothetical prospective diver as fit. CONCLUSIONS: There is no consensus among doctors who perform diving medical examinations as to what constitutes fitness to dive; current guidelines need to be improved. PMID- 10721341 TI - What should we do about overweight and obesity? PMID- 10721342 TI - The sources of risk factor information for general practitioners: is physical activity under-recognised? AB - OBJECTIVE: To identify and compare the amount of material on physical activity and the management of smoking, hypertension and hypercholesterolaemia in medical journals and magazines frequently read by general practitioners. METHOD: Qualitative study assessing the total number of articles and advertisements to which Australian GPs are exposed in journals and medical magazines they are likely to read. RESULTS: Only 6% of articles about cardiovascular disease (CVD) risk factors in the Medline search and 5% in the medical magazine search discussed exercise prescription or how to start and maintain an exercise program. Most CVD risk factor articles were on the pharmacological treatment of hypertension (42%), followed by hypercholesterolaemia (32%) and smoking cessation (20%). A review of medical magazines found similarly ranked results, and a count of advertisements indicated 67% related to hypertension, 26% to hypercholesterolaemia and 7% to smoking cessation. CONCLUSIONS: GPs are less well informed by the medical media about physical activity than about other traditional CVD risk factors, although the epidemiological evidence for their health benefits is similar. Strategies should be developed to inform doctors about the evidence of benefits from regular moderate physical activity, and for GPs to recommend exercise in most clinical encounters. PMID- 10721343 TI - The effectiveness of popular, non-prescription weight loss supplements. AB - OBJECTIVES: To review the evidence for the effectiveness of popular, non prescription weight loss supplements. DATA SOURCES: A detailed literature search including all relevant medical and supplementary medicine databases and evidence submitted from manufacturers. DATA SYNTHESIS: The theoretical basis and rationale for the use of each substance is considered along with available research in the published literature on effectiveness and potential risks. We classified the level of evidence represented by the main research studies on each substance. CONCLUSIONS: There is no good evidence for any weight loss benefits from most of the substances reviewed here. There is some support for mild effects of capsaicin, caffeine and fibre, but only in whole foods. In some cases (e.g., chitosan), there is a plausible theoretical basis for the product, but no supporting proof of effect in humans in the absence of a calorie-controlled diet. Possible synergistic effects of different ingredients cannot be dismissed, but cannot be assessed from current data. There is an absence of good quality research on many substances, which means that advertising claims may be misleading. PMID- 10721344 TI - Obesity and virtue. Is staying lean a matter of ethics? PMID- 10721345 TI - Why staying lean is not a matter of ethics. PMID- 10721346 TI - Accessibility to general practitioners in rural South Australia. A case study using geographic information system technology. AB - OBJECTIVE: To demonstrate the potential of GIS (geographic information system) technology and ARIA (Accessibility/Remoteness Index for Australia) as tools for medical workforce and health service planning in Australia. DESIGN: ARIA is an index of remoteness derived by measuring road distance between populated localities and service centres. A continuous variable of remoteness from 0 to 12 is generated for any location in Australia. We created a GIS, with data on location of general practitioner services in non-metropolitan South Australia derived from the database of RUMPS (Rural Undergraduate Medical Placement System), and estimated, for the 1170 populated localities in South Australia, the accessibility/inaccessibility of the 109 identified GP services. MAIN OUTCOME MEASURES: Distance from populated locality to GP services. RESULTS: Distance from populated locality to GP service ranged from 0 to 677 km (mean, 58 km). In all, 513 localities (43%) had a GP service within 20 km (for the majority this meant located within the town). However, for 173 populated localities (15%), the nearest GP service was more than 80 km away. There was a strong correlation between distance to GP service and ARIA value for each locality (0.69; P < 0.05). CONCLUSIONS: GP services are relatively inaccessible to many rural South Australian communities. There is potential for GIS and for ARIA to contribute to rational medical workforce and health service planning. Adding measures of health need and more detailed data on types and extent of GP services provided will allow more sophisticated planning. PMID- 10721347 TI - Mobile intensive care services in rural South Australia. AB - In the 12 years from 1984 to 1995, Adelaide-based mobile intensive care teams transported 4443 critically ill patients from rural areas in South Australia and adjacent States to tertiary-level hospitals in Adelaide. The SA Ambulance Service undertook communications, support staffing and deployment of transport. Average radial distances in 819 road missions were 71 km, in 808 helicopter missions 122 km, and in 2777 fixed-wing aircraft missions 398 km. The largest groups of patients were neonates (23%) and those with trauma (25%). Rural hospitals made 96% of the requests for intensive care transport; 4% came from ambulance or other emergency service crews at accident locations. Emergency surgical or operative obstetrical procedures were performed on 2.7% of patients before transport. One hundred and thirteen patients (2.5%) died during resuscitation or transport, with one death deemed to be preventable. PMID- 10721348 TI - Rural doctors--who are they? PMID- 10721349 TI - Ups and downs of rural practice: general practice. PMID- 10721350 TI - Medicine in the bush: a consultant physician's view. PMID- 10721351 TI - Ups and downs of rural practice: a surgeon's view. PMID- 10721352 TI - Retirement at last! PMID- 10721353 TI - Funding Australia's basic biomedical research of 1993 and 1994. AB - The meshing of two databases of scientific publications--the Wellcome Trust's Research Outputs Database, and the Research Evaluation and Policy Project's database of Australian publications--allows a detailed analysis of the funding agencies providing external (as opposed to intramural) support for Australia's basic biomedical research. This analysis shows the success Australian researchers are having in attracting funding from overseas, and the high citation rates achieved by publications with external funding. PMID- 10721354 TI - The Centenary Institute of Cancer Medicine and Cell Biology. PMID- 10721355 TI - Nature, nurture and my experience with smallpox eradication. PMID- 10721356 TI - The rise and rise of academic general practice in Australia. PMID- 10721357 TI - Stinging insect allergy. PMID- 10721358 TI - Funnel-web spider (Hadronyche infensa) envenomations in coastal south-east Queensland. AB - Five patients with confirmed funnel-web spider bites (Hadronyche infensa) presented to Nambour General Hospital, in south-east Queensland, between 1992 and 1998. Two patients required antivenom; low doses of antivenom were effective. Patients were bitten in spring and early summer. In areas such as this, where funnel-web spider bites are reported less frequently than in New South Wales, clinicians and the community should be aware of the risks and immediate management of these bites. PMID- 10721359 TI - Exotic myiasis with Lund's fly (Cordylobia rodhaini). AB - After a four-week holiday in East Africa, a woman was diagnosed with furuncular myiasis: a third-instar larva of the fly Cordylobia rodhaini (Lund's fly) was found in a skin lesion. This is the first report of exotic myiasis and importation of this species of fly into Australia, and reflects the increasing risk of introducing exotic flies of public health and veterinary importance to Australia. PMID- 10721360 TI - The suicide of Thomas Wentworth Wills. AB - Thomas Wentworth Wills was the most important Australian sportsman of his time. He captained the Victorian colony at cricket and was the first hero of Australian Rules football. Although his picture now adorns the conservative Melbourne Cricket Club, he died in 1880, an isolated, destitute alcoholic, after stabbing himself in the heart. Wills embodied a tradition, as prevalent today as it was over 100 years ago, that weds sport with alcohol in Australian culture. PMID- 10721361 TI - Will sex survive to 2099? PMID- 10721362 TI - What if there is a "sunset clause" on the Y chromosome? PMID- 10721363 TI - Future change in sexual behaviour? AB - Virtual sex is not a substitute for normal sexual intercourse, lacking emotional interaction between couples and the intimacy and reinforcement of a loving relationship. PMID- 10721364 TI - Sex, reproduction and impregnation: by 2099 let's not confuse them. PMID- 10721365 TI - Reproductive technology, efficiency and equality. PMID- 10721366 TI - CLINICam. PMID- 10721367 TI - Some thoughts while nymphing. PMID- 10721368 TI - A new, occasional instrument for measuring marital quality. The time required to make a cream cheese and salmon bagel following funnel-web spider bite. PMID- 10721369 TI - Breaking the rules: a thoracic impalement injury. AB - In the case of a patient with an impalement injury, the object should be removed in a controlled operating theatre environment. We report an 18-year-old man for whom this rule could not be followed. He was removed from a metal pipe transfixing his chest at the roadside. PMID- 10721370 TI - A riposte for a fencer. The residency appointment of Alfred Edmund Finckh to Sydney Hospital in 1905. Was it just a matter of male chauvinism? AB - Alfred Finckh gained a residency at Sydney Hospital in 1905 in preference to a more academically successful female medical graduate, amid some controversy over the place of female doctors in hospitals. Here, two of his descendants argue his cause: that he was an experienced scientist with qualities that merited his selection for the post. PMID- 10721371 TI - "Ute surfing": a novel cause of severe head injury. AB - Riding on the load tray of a moving utility vehicle, often after drinking alcohol, is a recognized pastime among young Australian men. The cases presented here show that associated accidents can result in serious head injuries and probable permanent neurological deficits. PMID- 10721372 TI - Letter from Yemen. PMID- 10721373 TI - On dry land. PMID- 10721374 TI - Sporotrichosis mimicking necrotising arachnidism. PMID- 10721375 TI - Browne's Law of Medicine. PMID- 10721376 TI - Summer vomiting disease? PMID- 10721377 TI - The present status of electroconvulsive therapy: a systematic review. PMID- 10721378 TI - Doctor migration--Daniel in the lions' den. PMID- 10721379 TI - Doctor migration--Daniel in the lions' den. PMID- 10721380 TI - Interpretation of HIV results for insurance purposes. PMID- 10721381 TI - Medical testing and rape. PMID- 10721382 TI - Are Bartonella emerging and re-emerging pathogens in southern Africa? PMID- 10721383 TI - Focus on AIDS. PMID- 10721384 TI - The new medical aid laws and the consumer. PMID- 10721385 TI - Appropriate laboratory monitoring of HIV. PMID- 10721386 TI - 'Is this treatment worth while?' How to read the medical journals. PMID- 10721387 TI - Unrecognized acute renal failure following the comrades marathon. PMID- 10721388 TI - Potentially serious drug interactions with grapefruit juice. PMID- 10721389 TI - The criminalization of psychiatry. PMID- 10721390 TI - An unusual cause of deep venous thrombosis of the lower limb. PMID- 10721391 TI - An experiment with problem-based learning at the University of Zambia. PMID- 10721392 TI - Factor IX deficiency and anaphylaxis. PMID- 10721393 TI - Thyroid function tests among South African children with minor illnesses. PMID- 10721394 TI - A structured record to implement the national guidelines for diabetes and hypertension care. AB - BACKGROUND: Guidelines to improve standards of care for hypertension and diabetes were disseminated by the National Department of Health in 1996 but have generally not been implemented by health professional in local primary care. A strategy for the adoption and implementation of the Guidelines was developed in collaboration with health professionals in primary care. OBJECTIVES: The development of a structured record, with prompts for the management of diabetes and hypertension according to the Guidelines. SETTING: Three community health centres (CHCs) in the Western Cape. PARTICIPANTS: Doctors and nurses managing patients with diabetes and hypertension. METHODS: A draft of the structured record was developed at a single-pilot CHC in the Western Cape. Focus group discussions established the core requirements for a structured record. Process, result and structural indicators in line with the national Guidelines were considered for inclusion in the draft record. This draft record was then piloted at two other CHCs. Comments from semi-structured interviews and pre- and post-test evaluation questionnaires were used to compile the final instrument. RESULTS: Eleven doctors and 8 nurses participated in the development of the final instrument. Important considerations in the design were a single-page, user-friendly format, tick-boxes to reduce writing, prompts, provision for sequential recording, target setting, and compatibility with the Guidelines. The final instrument was piloted and elicited a favourable overall response. CONCLUSION: The structured record simplifies the application of the Guidelines and the systematic recording of processes of care. The effectiveness of the Guidelines will be evaluated further in a randomised control trial using the structured record. PMID- 10721395 TI - A survey of hospital outpatient services for chronic diseases in Gauteng. AB - OBJECTIVES: The rapid evaluation of hospital-based services for chronic non communicable diseases, in particular aspects of the organisation of services, and indirect indicators of patient care. DESIGN: A postal survey of services for asthma, epilepsy, diabetes and hypertension at nine hospitals. Assessment over 1 week of single blood pressure (BP) and blood glucose readings at the hypertension and diabetes clinics at one regional hospital. SETTING: Nine community and secondary hospitals in Gauteng. RESULTS: Eight hospitals responded. Most did not provide specific clinics for each condition. None of the professional staff had received additional training in chronic disease management, and 7 considered their services to be understaffed. On average, nurses managed 33 patients per day (range 19-50), and doctors 53 (20-80). Mean consultation time was 9 minutes (4-20 minutes). Management guidelines were used for all conditions in 5 hospitals. Modern routine assessments were seldom employed. Estimates of regular patient attendance ranged from 25% to 75%. At the single hospital surveyed, hypertension (N = 233) was controlled in 42.5% of patients using World Health Organisation criteria (BP < 160/95), but in only 24.5% of patients by The Joint National Committee on Prevention, Detection, Evaluation and Treatment of High Blood Pressure (JNC6) standards (BP < 140/90). Random blood glucose was satisfactory (< 10 mmol/l) in 45.2% of diabetic patients (N = 157) while hypertension (N = 100) was well controlled (< 140/90) in 10% of hypertensive diabetic patients. CONCLUSIONS: Services for chronic diseases at non-academic hospitals in Gauteng were characterised by perceived inadequate staff numbers and training, short consultation times, infrequent use of management guidelines and standard assessments, little patient education with regard to self care, and perceived low rates of regular attendance (and hence compliance with medication). At one hospital there was a low rate of hypertension control, and unsatisfactory rates of acceptable glycaemic and BP control among diabetic patients. There is an urgent need for restructuring of services for chronic diseases and for more detailed outcomes research. PMID- 10721396 TI - Follow-up of patients with arrhythmogenic right ventricular cardiomyopathy dysplasia. AB - OBJECTIVE: The enlargement of data on the natural course and management of patients with arrhythmogenic right ventricular cardiomyopathy/dysplasia (ARVC/D). DESIGN: Retrospective and partly prospective observational study. SETTING: Cardiac Unit, Wentworth Hospital, Durban--the only unit in KwaZulu-Natal providing an arrhythmia and electrophysiology service. STUDY POPULATION: Those included were: (i) patients referred for palpitations, unexplained syncope, or ventricular tachycardia and in whom ARVC/D was diagnosed according to multiple criteria; and (ii) family members of patients with ARVC/D in whom the disease was documented using the same criteria. MAIN OUTCOME AND MEASUREMENTS: Diagnosis, management, morbidity and mortality were analysed. RESULTS: Twelve patients were diagnosed with ARVC/D over a period or 6 years. At the end of follow-up for 3.4 +/- 3.2 years, 7 of them were well and alive on anti-arrhythmic medication, 2 were asymptomatic, and 3 had died. One death was sudden, 1 patient died due to left ventricular failure, and 1 patient died due to a low cardiac output syndrome 3 months after right ventricular isolation, i.e. the mortality rate was 25%. ARVC/D was found in all racial groups and was familial in 5 patients (42%). In all but one patient the correct diagnosis was not suspected by the referring institution, physician or cardiologist. CONCLUSIONS: ARVC/D needs to be included into a differential diagnosis of unexplained syncope, palpitations, or ventricular tachycardia by all health service providers. Its management remains a complex challenge with varying results. PMID- 10721397 TI - Extraordinary arousal from semi-comatose state on zolpidem. A case report. AB - A young semi-comatose male patient was investigated using 99mTc hexamethyl propylene amine oxime (99mTc HMPAO) brain single photon emission computed tomography (SPECT) before and after administration of the gamma-aminobutyric acid (GABA) agonist zolpidem. It was observed that 15 minutes after application of the drug the patient awoke from his semi-comatose condition and remained awake for the next 3-4 hours. When drug action subsided he returned to his semi-comatose state. Brain SPECT before drug application showed large hypo-active areas in certain parts of the brain. Brain SPECT after drug application showed a generalised cortical activation relative to the cerebellum and a marked and amplified activation of the areas that were hypo-active before drug application. PMID- 10721398 TI - Abhorrent weapons and 'superfluous injury or unnecessary suffering': a concern for trauma surgeons? PMID- 10721400 TI - Median nerve compression associated with displaced Salter-Harris type II distal radial epiphyseal fracture. AB - Three children with grossly displaced Salter-Harris Type II fractures of the distal radial epiphysis underwent immediate manipulation under anaesthetic (MUA) because of rapidly developing median nerve compression. In each case nerve function was quickly restored with no late neurological sequelae. We believe that in children who sustain this injury with signs of median nerve compression, immediate MUA without carpal tunnel release is acceptable initial management. Late exploration of the median nerve can be considered should a neurological deficit persist. PMID- 10721399 TI - A review of post-traumatic stress disorder. Part I: Historical development and classification. AB - This paper describes the history of the development of understanding of psychological responses to traumatic life-events and their treatment. One major response, post-traumatic stress disorder (PTSD), is a recognized condition which has appeared relatively recently in diagnostic manuals. PTSD is a condition of major significance, not only to mental health professionals, but also to trauma surgeons and allied professionals. This paper focuses on the current definition of PTSD in the International Classification of Diseases (ICD-10, 1992, World Health Organization) and the Diagnostic and Statistical Manual of the American Psychiatric Association (DSM-IV, 1994). PTSD first appeared as an operational diagnosis in DSM-III (1980) and was revised in DSM-III-R (1987) and DSM-IV (1994). It made its first appearance in the ICD system later, in 1992. This paper seeks to increase awareness of PTSD across the broad spectrum of trauma management professionals, to emphasize the practical value of identifying the disorder and to encourage optimism for its treatment. This paper is in two parts. The first part deals with historical development and classification. The second part (to appear in the next issue of Injury) deals with treatment. PMID- 10721401 TI - Gunshot injuries of the lower oesophagus. AB - We present seven cases of lower oesophageal gunshot injury cared for by one surgeon. Diagnosis was made clinically, with the help of chest X-rays and with oesophagography and oesophagoscopy. Five were treated with wide debridement and resection of the distal esophagus and oesophago-gastric anastomosis with a Nissen wrap to protect the anastomosis. Two lesser injuries were treated by primary repair. The five treated with resection and oesophago-gastric anastomosis did not leak and the patients were discharged after oesophagography 10 days postoperatively. Primary repairs in two patients were complicated by oesophageal leaks, one subclinical and one with an empyema. The oesophageal blood supply is segmental in areas and variable in the distal part. Injury due to a bullet wound may cause more damage than is evident at surgery. Additional mobilization can further devascularize the distal oesophagus and lead to anastomotic leaks. We advocate wide debridement of oesophageal gunshot injuries and resection of the distal oesophagus. Continuity is restored with a primary oesophago-gastric anastomosis (double layer) with a fundoplication to protect the anastomosis. PMID- 10721402 TI - Patient preference for route of diclofenac. AB - The purpose of this study was to investigate whether patients attending an A&E department would prefer injection or suppository diclofenac as analgesia if oral medication was contraindicated. Patients presenting to the 'walking wounded' side were asked to fill in a questionnaire indicating their preference. The results showed a strong preference for intramuscular diclofenac. If we wish to prescribe rectal diclofenac, we should be prepared to take into account the patient's wishes first and, if necessary, explain our reasoning. PMID- 10721403 TI - Angulated screw placement in the lateral condylar buttress plate for supracondylar femoral fractures. AB - Certain supracondylar femoral fractures are not amenable to internal fixation with fixed angle devices. In these instances, the condylar buttress plate is the recommended alternative; however, this is a less rigid device. Because of the decreased rigidity and strength of this device, there is a tendency toward varus angulation and malunion. In six fresh-frozen human knee specimens, segmental osteotomies were created to mimic supracondylar femoral fractures. The medial cortex was completely removed to make the fracture unstable to varus deformity. The fracture was fixed with a lateral condylar buttress plate using 4.5 mm screws. Each specimen was tested once with all the screws installed perpendicular to the plate, and again with the middle screw, just proximal to the fracture, angled 45 degrees diagonally across the fracture into the subchondral bone of the medial femoral condyle. For the construct with all screws placed perpendicular to the buttress plate, the initial stiffness was 410 N/mm, and after 1000 cycles it was 230 N/mm. With a screw placed diagonally across the fracture site, stiffness increased to 833 N/mm on the first cycle, and 796 N/mm after 1000 cycles. In all specimens with the screws placed perpendicular to the plate, the distal fragment had a permanent varus deformity after 1000 cycles, under no load, of 0.91 mm. For the diagonal screw condition, the average magnitude for all six specimens was 0.42 mm. This simple means of screw angulation in the plate strengthened the overall construct to resist the tendency toward varus deformity. The attractive features include the ease of application, and the use of an existing construct. PMID- 10721404 TI - Revascularization of the limbs using vein grafts after vascular injuries. AB - We report on 32 patients with vascular injury of a limb undergoing a total of 41 revascularization procedures with interposition vein grafts. A combined arterial and venous injury was present in nine cases, an isolated venous injury in four, and an isolated arterial injury in 19 cases. Eighteen per cent of patients with arterial injuries had normal distal pulses on initial examination. Preoperative arteriography was performed in 12 cases, and intraoperative arteriography in four. All venous injuries were diagnosed at operation. In most cases, the contralateral greater saphenous vein was used for grafting. Four patients had postoperative thrombosis after arterial reconstruction resulting in below knee amputation in two cases. Two patients suffered from postoperative swelling caused by venous insufficiency, one after ligation of an injured axillary vein, and the other one following venous thrombosis of a superficial femoral vein repair. It is concluded that revascularization of arterial and venous injuries of the extremities with interposition vein grafts is successful in most cases resulting in low amputation rates, and should be attempted in all major vascular injuries in viable limbs. PMID- 10721405 TI - MR imaging of suspected acute spinal instability. AB - Twenty-two patients with suspected acute vertebral instability were imaged within 48 h of injury using a 0.5 Tesla magnet. In all patients plain radiographs, T1 weighted gradient echo (GE) and STIR sequences were performed. Two radiologists blindly evaluated the magnetic resonance imaging (MRI) scans defining injuries with bony or soft tissue disruption of both columns as unstable. Indirect signs of instability such as soft tissue haemorrhage were recorded and correlated where possible with operative findings. Sixteen patients were radiologically unstable on MRI, five more than on plain films alone. Instability was confirmed operatively in 10 patients. The six other patients with unstable injuries were treated conservatively. Two of these patients had evidence of increased deformity before fracture union. The radiologically stable patients were treated as such and at 6-month review showed no evidence of progressive instability. The presence of soft tissue haemorrhage in the interspinous gap was not associated with ligament rupture unless actual discontinuity was demonstrated at that level. We conclude that using MRI acutely, most unstable spinal injuries can be rapidly and accurately evaluated without the need for further imaging. PMID- 10721406 TI - Improvement in the therapy of multiply injured patients by introduction of clinical management guidelines. AB - A trauma algorithm representing the guidelines for the management of emergency treatment of severe blunt trauma was implemented at our institution in 1994. By comparison of two prospectively recorded cohorts of multiply injured patients, the clinical efficacy of these guidelines was analysed. The algorithm cohort comprised 74 patients over the period January 1994 to June 1996, and the Control cohort 126 patients over the period April 1988 to December 1993. To evaluate procedural quality of early clinical trauma management, nine criteria were applied. After implementation of the algorithm there was an improvement in all parameters reflected by a significant reduction of missed injuries and important time savings. Mortality rates in the cohorts were calculated after subdivision into three groups (I-III) with moderate (ISS: 18-24), high (ISS: 25-49) and very high (ISS: 50-75) injury severity. All cohort subgroups were comparable with respect to ISS values, age, initial loss of consciousness (GCS) and shock rate. In all subgroups of the algorithm cohort mortality rates were reduced: group I: 0 versus 20 per cent (p < 0.05); group II: 8 versus 24 per cent (p < 0.05); group III: 40 versus 71 per cent. Improvements in both therapeutic process and outcome were observed after implementation of the trauma algorithm. PMID- 10721407 TI - Fat embolism syndrome in isolated femoral fractures: does timing of nailing influence incidence? AB - In a consecutive series of 274 patients with isolated femoral shaft fractures 11 patients (4%) developed fat embolism syndrome. There were no cases of fat embolism syndrome in patients over the age of 35 years. Of the remaining patients, 60 operated on within 10 h of injury did not develop fat embolism. This left 109 patients who had nailing performed more than 10 h after injury of whom eleven (10%) developed fat embolism syndrome (p < 0.027). Patients under the age of 35 years with isolated femoral fractures should have nailing performed as early as possible after injury to minimize fat embolism syndrome. PMID- 10721408 TI - Kocher's painless reduction of anterior dislocation of the shoulder: a prospective randomised trial. AB - Forty-five patients with an anterior dislocation of the shoulder were randomised into one of two treatment groups and manipulation performed using Kocher's original method (without traction). A successful reduction was achieved in 80.9% of patients administered Entonox only and in 100% of patients sedated intravenously. No statistical significance was found in the pain scores between the two groups. The study shows that Kocher's original method is a reliable technique for reducing anterior dislocation of the shoulder and a successful outcome can be expected using nitrous oxide only, obviating the need for intravenous sedation and analgesia in the majority of patients. PMID- 10721409 TI - Delayed onset of retrobulbar haemorrhage following severe head injury: a case report and review. PMID- 10721410 TI - Traumatic pseudomeningocele at cranio-vertebral junction following stab injury. PMID- 10721411 TI - Fibular malreduction in AO/Weber type C ankle fractures. PMID- 10721412 TI - Risk factors for 'whiplash' in drivers: a cohort study of rear-end traffic crashes. PMID- 10721413 TI - Phenotypic plasticity and prognostic factors in extraskeletal myxoid chondrosarcoma. AB - Extraskeletal myxoid chondrosarcoma (EMC) is an intriguing entity with a broad morphologic spectrum that overlaps features found in other benign and malignant mesenchymal tumors. This phenotypic plasticity should be known to avoid erroneous diagnosis. The identification of older age, larger tumor size, proximal tumor location, and metastatic disease as independent adverse predictors of survival is useful in achieving optimal treatment and follow-up in high-risk patients. PMID- 10721414 TI - Villous adenoma of urinary tract: a common tumor in an uncommon location. AB - The presence of colonic-type epithelium in the urinary tract is not an uncommon occurrence, but tumors derived from it are rare. Tumors arising from colonic-type epithelium, including villous adenoma and adenocarcinoma, have been reported in the renal pelvis, ureter, urinary bladder, and urethra. Villous adenomas of the urinary tract are rare, being most common in the urinary bladder, followed by the urethra. Morphologic features of these tumors are similar to those of the colonic adenomas. The largest published series of villous adenomas of the urinary tract was a study of 23 patients. This study is reviewed and other reports on villous adenomas of the urinary tract are discussed. PMID- 10721415 TI - Mismatch repair proteins and microsatellites hit clinical practice. AB - Hereditary nonpolyposis colon cancer (HNPCC) is one of the most common familial cancers with characteristic molecular changes that are different from those found in familial adenomatous polyposis (FAP) coli. Genetic mutations in the germline and somatic cells lead to loss of expression of one of the two most commonly involved mismatch repair genes, hMSH2 or hMLH1, and consequently, to expansion of certain repetitive DNA sequences (microsatellite instability (MSI)). The paper describes a distinct subtype of "HNPCC-like" sporadic colonic carcinoma that can easily be identified by immunohistochemistry. Recognition of this subtype of colonic cancer is important because it occurs in the younger age group and is associated with better survival, but also a five-fold chance of developing a second colorectal carcinoma compared to "conventional" colorectal carcinomas. PMID- 10721416 TI - Atypical subcutaneous fatty tumors. AB - Atypical fatty tumors present problems of terminology and diagnosis. While atypical fatty tumors that are located in subcutaneous tissues have an extremely good prognosis, local recurrence and transformation to a higher grade tumor may occur. Allen et al document the morphologic features and behavior of 37 atypical subcutaneous fatty tumors and describe a system of grading. The commentary discusses the terminology of atypical fatty tumors, the relevance of the proposed grading system, and the significance of a spindle cell component in atypical subcutaneous fatty tumors. PMID- 10721417 TI - Detection of stromal invasion in breast cancer: the myoepithelial markers. AB - Breast cancer can only be life threatening when it becomes invasive, at which point it carries potential for spreading and metastasis. Therefore, it is critical to distinguish invasive carcinomas (IC) from noninvasive lesions, the latter including ductal carcinoma in situ (DCIS) and benign breast lesions. While this distinction is usually made based on histologic evaluation alone, in a small but significant number of cases, accurate diagnosis may be impossible, particularly in the context of core needle biopsies. To this end, a number of immunohistochemical markers have been utilized to help establish the presence (or absence) of stromal invasion. The fact that the loss of the myoepithelial cell (MEC) layer is a hallmark of IC (but not DCIS) suggests the use of antibodies to MEC to distinguish IC from DCIS. However, these markers have a wide range of specificity and sensitivity, with the potential for problems in interpretation. The use of many of these markers is limited by high rates of 'false positive' or 'false negative' immunostaining. In this review, the biology of stromal invasion in breast carcinomas will be discussed, as well as the various myoepithelial markers, with emphasis on pitfalls related to their sensitivity and specificity in the detection of MECs in the breast. Finally, the authors will discuss diagnostically challenging breast lesions, which may require the use of MEC marker studies to reach a definitive diagnosis. PMID- 10721418 TI - An update on "special stain" histochemistry with emphasis on automation. AB - For nearly 100 years, pathologists have utilized "special histochemical stains" to assist in tissue-based diagnosis. As illustrated in Figures 1 and 2, histochemical stains have been used to identify infectious microorganisms (e.g., Mycobacterium tuberculosis with acid-fast bacillus (AFB) stain), to detail inflammatory stromal or structural alterations (e.g., fibrosis in liver cirrhosis with Masson trichrome), to identify microanatomic sites of disease (e.g., basement membrane in glomerulonephritis with Jones methenamine silver), to identify abnormal chemical deposits (e.g., iron in hemochromatosis with Prussian blue stain), or abnormal immune deposits (e.g., amyloid via Congo red stain). The current surgical pathology laboratory may employ a repertoire of 20 to 25 "special stains" to ensure the full diagnostic complement. While the diagnostic repertoire and the biochemical recipes for the stains are now a well-established, codified part of surgical pathology, there is an ever-moving, leading edge of new developments including new reagents, applications, and methods. This review seeks to update the reader on some of the new applications including both new reagents and methods. Particular emphasis will be placed on the recent technologic advance of automating special stains in kinetic-mode (1-4). The authors consider in turn: 1. In brief, the "news" (recent literature review) of new staining applications; 2. In greater detail, two new applications for detection of Microsporidia and Helicobacer pylori; 3. The new technologic advancement of kinetic mode automation of special stains. PMID- 10721420 TI - Integrated chicken genetic and cytogenetic maps with FISH identification of microchromosomes. PMID- 10721419 TI - Thyroid transcription factor-1 is a marker of lung and thyroid carcinomas. AB - Thyroid transcription factor-1 (TTF-1) is a tissue specific transcription factor expressed in epithelial cells of the thyroid and lung, as well as in certain areas of the brain. Recent investigations have shown that TTF-1 is also commonly present in a wide variety of thyroid and lung tumors. Because of its restrictive expression, TTF-1 is a useful immunohistochemical marker in the diagnosis of tumors of thyroid or lung origin. PMID- 10721421 TI - Contribution to the comparative map between humans and pigs. PMID- 10721422 TI - Applications of chromosomal fish in the Bovidae with emphases on physical mapping in domestic cattle and comparative cytogenetic analyses of the tribe Bovini. PMID- 10721423 TI - Establishment of bovine chromosome region specific high resolution maps using microdissection. AB - An alternative approach for the direct analysis of chromosome regions corresponding to economical traits on the basis of chromosome microdissection is described. Large fragment clones isolated with primer pairs designed from chromosome fragment-specific DNA sequences were localized by FISH to the scraped chromosome region of interest. The chromosome fragment-specific clones are a useful tool for the generation of region specific high density marker and gene maps and represent the source material for the development of a DNA contig including the economical trait. PMID- 10721424 TI - Novel chromosomal insertional translocation in chicken uncovered by double color FISH. AB - A novel insertion 78,ZZ or 78,ZW, ins(3;1)(q25q27;undetermined) was revealed in chicken by double color fluorescent in situ hybridization (FISH). A fragment of chromosome 1 spanning either from q13-14 to q34-35, or from q14-21 to q36-41 bands, had been translocated to chromosome 3 at a site located between q25 to q27 bands. This has resulted in the generation of an interstitial deletion in chromosome 1 and an insertional translocation in chromosome 3. Chickens with this balanced insertional translocation are asymptomatic carriers and their fertility is not affected, but embryo mortality increases. Greater than 50% occurrence of unbalanced gametes are observed. However, progeny sex ratio is not affected. PMID- 10721425 TI - Molecular cytogenetic analysis of the chicken and red-legged partridge chromosome 4 repatterning. AB - Conserved genome homologies between the chicken and partridge have been demonstrated for chromosomes 1 and Z in previous studies. Morphological differences between the chicken and partridge for chromosome 4 have also been identified. The chicken chromosome 4 is submetacentric while the partridge chromosome 4 is acrocentric. We now report that in spite of this morphological difference, both species share extensive homology for chromosome 4 as determined by fluorescent in situ hybridization (FISH). Since only two chromosomes of the partridge karyotype showed FISH signals, our observation suggests that a chromosome rearrangement (peri- or paracentric inversion) has occurred in the partridge chromosome 4. PMID- 10721426 TI - Progeny testing of heterozygote carriers of the Robertsonian translocation 1/29 in Bovidae. AB - In our recent population study, 220 daughters of a heterozygote carrier of the Robertsonian translocation 1/29 were analysed by screening of metaphase spreads and typing of microsatellite markers. The segregation between markers near the centromere of chromosomes 1 and 29 and the fusion were analysed. The microsatellite markers were selected from the USDA, MARC cattle genome map. Analyses were done on AGLA17, BM6438, TGLA49, BMS 1928, BM8139, INRA117, BMS574, BMS711 and BMS4015 of chromosome 1, and on BM4602, BMC2228 and BMS1857 of chromosome 29. The two markers BMC2228 and BMS4018 in the linkage group originating from the fusion were able to either recognise or exclude 167 daughters out of 220 as carriers of the Robertsonian translocation 1/29. Fifty three daughters showed double heterozygote markers like their father, and were therefore not informative. The use of conventional cytogenetics in combination with molecular studies has allowed a more precise evaluation of the Robertsonian translocations than either approach alone might have done. PMID- 10721428 TI - Utility of chicken-specific microsatellite primers for mapping the turkey genome. AB - As part of the University of Minnesota's initiative to map the turkey genome, we are currently evaluating chicken microsatellite loci for use in mapping the turkey genome. To date, 141 primer pairs have been tested for amplification at six different combinations of temperature and MgCl2 concentration. Microsatellite primer pairs from the Chicken Comprehensive Mapping Kit #2, and additional unpublished chromosome 1 and 2 primers were screened. Analyzable PCR products were produced from 78 of the 141 (55%) primer combinations. In the majority of cases (68%), PCR fragments obtained from the turkey were similar in size to respective chicken loci. The presence of dinucleotide repeats (CA/TG repeats) was determined by Southern hybridization with a (TG)15, oligonucleotide probe. Five of 12 (41.63%) turkey fragments hybridized under low stringency conditions. The length of the dinucleotide repeats in the turkey, relative to the chicken sequences, were found to correspond directly with hybridization intensity. Amplification of homologous loci was confirmed by direct sequencing and subsequent alignment of the turkey and chicken sequences. The results of this study indicate that the use of chicken-specific microsatellite primers will rapidly and significantly enhance construction of a genetic map for the turkey. PMID- 10721427 TI - Identification and radiation hybrid mapping of members of the porcine proteasome/ubiquitin system. AB - We report the identification and radiation hybrid mapping of members of the proteasome/ubiquitin system in pigs that, so far, have only been identified in humans and cattle. Expressed sequence tags (ESTs) were constructed from ten oligo(dT)-primed individually tagged, directionally cloned and normalized cDNA libraries from peripheral blood cells (PBC), spleen (Sp), thymus (Th), lymph node (LN) and bone marrow (BM) from immunologically naive and challenged pigs as part of an implant-associated orthopedic infection model. The ESTs mapped using the 7000 rad IMpRH panel (Hawken et al., 1999) were ubiquitin fusion-degradation 1 like protein (UFD1L), ubiquitin activating enzyme E1 and ubiquitin-S27a fusion protein which mapped to porcine chromosomes 14, 7 and X, respectively. PMID- 10721429 TI - Physical mapping of porcine seasonality genes. AB - Seasonal infertility in sows is a problem in the pig industry characterized by delayed onset of puberty in summer and decreased farrowing rate resulting from silent oestrus and aborted pregnancy. Summer infertility is thought to be influenced by heat, sunburn and stress. However, the strongest contributory factor is photoperiod. The difference in seasonality between wild boar and commercial pig breeds suggests that there may be a genetic component to this trait. The maps and associated molecular tools emerging from the pig genome project have created opportunities to examine the genetic component of seasonal infertility. We are identifying and mapping genes that are likely to be involved in biological clock mechanisms and the melatonin pathways as candidate seasonality genes. PMID- 10721431 TI - Status of oocytes and embryos of X-autosome translocation carrier sows. AB - The impact of an X-autosome translocation t(Xp+; 14q-), on ovulation, fertilization and embryo survival in carrier sows, was examined and compared with these parameters of normal sows. Corpora lutea counts during week-2 and week-4 of gestation were similar in normal and carrier sows (14.4 +/- 1.36 and 15.5 +/- 2.18) although embryo recovery (11.0 +/- 1.87 and 6.0 +/- 1.47) was lower than that from normal sows (12.8 +/- 1.46 and 11.5 +/- 0.87), at these stages. Among the embryos karyotyped from the week-2 embryos of carrier sows, 42% were normal, 26.4% were carriers and 31.6% were of unbalanced chromosome make-up, and of the week-4 embryos of carriers, 33.3% were normal, 57.1% were carriers and 9.1% were chromosomally unbalanced females. The preponderance of females among the unbalanced embryos recovered at week-2 of gestation (11_ and 1_) and the total absence of males among those recovered at week-4, suggest that oocytes with unbalanced chromosome constitution are eliminated before week-2 of gestation if they are fertilized by Y bearing sperm, and that the unbalanced oocytes fertilized by X bearing sperm survive up to the peri-attachment stage even though all chromosomally unbalanced embryos are eliminated before term regardless of their sex. PMID- 10721430 TI - Analysis of allele segregation distortion in a swine resource family. AB - Segregation distortion of alleles was found in several regions of the genome in allele type analyses of genetic markers for a swine resource family that had been constructed by crossing Gottingen miniature pig and Meishan breeds. From these regions, a region on chromosome 6 presented a distortion over several consecutive markers. This chromosome 6 region was subsequently further investigated to reveal that alleles of a chromosome 6 homologue of the Gottingen miniature pig were not found in a homozygous family member. The litter size of F1-crosses which were able to produce homozygotes in this region were 24% smaller, on average, than F1 crosses which were unable to produce homozygotes. This indicated that this region of the chromosome 6 homologue contained a recessive gene or genes which could terminate fetal development. An additional 10 markers were subsequently used to investigate the region more precisely. These studies revealed that this region spans 7 cM and is located between markers Sw855 and Sw122. Since current comparative maps show that this region corresponds to the human chromosome 19 q arm, genes positioned on the human chromosome 19 q-arm were screened to select 20 candidate genes. These included the pregnancy-specific beta-1-glycoprotein gene. PMID- 10721433 TI - One size does not fit all: the case for tailoring print materials. AB - Printed health education materials frequently consist of mass-produced brochures, booklets, or pamphlets designed for a general population audience. Although this one-size-fits-all approach might be appropriate under certain circumstances and even produce small changes at relatively modest costs, it cannot address the unique needs, interests, and concerns of different individuals. With the advent and dissemination of new communication technologies, our ability to collect information from individuals and provide feedback tailored to the specific information collected is not only possible, but practical. The purpose of this article is to: (a) distinguish between tailored print communication and other common communication-based approaches to health education and behavior change; (b) present a theoretical and public health rationale for tailoring health information; and (c) describe the steps involved in creating and delivering tailored print communication programs. Studies suggest computer tailoring is a promising strategy for health education and behavior change. Practitioners and researchers should understand the approach and consider the possibilities it presents for enhancing their work in disease prevention. PMID- 10721432 TI - The integration of canine genetic maps with the canine karyotype using specific gene amplification of chromosome-specific DNA. AB - We have used a rapid approach to place markers that are already represented in current genetic maps onto individual chromosomes in species for which chromosome paints exist. PCR-based techniques are used to look for the presence of individual marker genes within each chromosome-specific DNA pool. The presence of a given marker within a DNA pool allows assignment of the complete radiation hybrid group, or linkage group from which the marker is drawn, to an individual chromosome. We have used this method with a new set of canine chromosome paints (Yang et al., 1999). In this way, we have assigned 39 of 44 published RH or syntenic RH groups to canine chromosomes, together with 33 of 40 canine linkage groups in a recently published map (Neff et al., 1999). PMID- 10721434 TI - The potential variances of tailoring in health behavior interventions. AB - The success of tailored print communications depends upon having a sufficiently diverse inventory of both content messages and delivery formats to respond to important differences among individuals. This article discusses means by which this diversity--the variances of tailoring--may be developed. One of the foundations of tailoring is the definition of a "focal point" for intervention. A focal point is characterized by a simultaneous combination of variables which specify a population group of interest, a target health behavior, and a setting in which the behavior occurs. All persons defined by the focal point may receive some intervention in common (i.e. targeted intervention). Tailored content responds to individuals within the focal point, based upon the antecedents of behavior within that focal point. This article elaborates on the focal point concept and then discusses factors that contribute to variations of tailoring. The psychosocial resources required by health behaviors are also reviewed, because tailoring must prepare the individual to make changes specific to the nature of a particular health behavior. This article does not specify what the variations of tailoring should be; the potential diversity of tailored messages is too great. Instead, the article presents basic elements that will go into the development of tailored interventions. PMID- 10721435 TI - How effective is tailored print communication? AB - This article reviews the "first generation" of tailored print communications studies in the published literature, describing the purpose, theoretical framework, sample, research design, message type and source, outcomes measured, and findings of each. Eight studies compared tailored versus similar nontailored print; one compared tailored print versus an alternate intervention, and three included tailored print as one of several intervention components. Although studies varied by behavioral topic, type of tailoring, and measurement of behavioral outcomes, several themes persist. Compared to their nontailored counterparts, tailored print communications have been consistently better remembered, read, and perceived as relevant and/or credible. There is also evidence that tailored print communications are more effective for influencing health behaviors. Six of the eight tailored/nontailored comparisons found more behavior change among tailored than nontailored recipients. Tailored print communications have also demonstrated effectiveness as an adjunct to other intervention components such as self-help smoking cessation manuals. However, studies comparing tailored print communications with tailoring via other media such as telephone counseling have shown mixed results. Additional research is needed to assess whether the behavioral topic itself may make a difference in whether tailoring is appropriate and effective. PMID- 10721436 TI - Challenges and future directions for tailored communication research. AB - As informatics technology advances, a growing number of research trials on tailored communications provide an accumulation of promising evidence to support their efficacy. These trials also reveal gaps and opportunities for future research. The scope and boundaries of tailoring must be redefined in terms of both new technology and the trade-offs between complexity, demand burden on participants, and the minimal information required for effective and efficient tailoring. Basic and methods research is needed to broaden theory, develop a common language, standardize measures, and isolate the key mediating mechanisms that facilitate tailored communications. Applied research must consider more rigorous research designs for efficacy trials and conduct more effectiveness trials to investigate the mechanisms of technology transfer to enhance large scale diffusion of tailored communications. The role of contextual variables needs to be examined, as well as their interaction with different population groups, and also the channels, modes, and methods of tailored message delivery. Research is also needed on the feasibility of tailoring across clusters of multiple risk factors to identify the commonalities, differences, and interrelations among diverse behaviors. The potential cost-effectiveness of tailored communications must also be examined. No matter how efficacious, tailored communications delivered to large populations (i.e. mass-customization) will not make a public health impact unless proven to be practical and cost efficient. PMID- 10721437 TI - Context, confidentiality, and consent in tailored health communications: a cautionary note. AB - This article highlights key contextual factors that emerge when the evolution of tailored health communications is viewed against the backdrop of dynamic changes in the nation's health care system--including the shift from fee-for-service medicine to managed care and the proliferation of direct-to-consumer and tailored marketing strategies in the pharmaceutical industry. It focuses on contextual variables with potential to significantly mediate the impact of personally tailored health advice--including those related to confidentiality, privacy, and informed consent and to the perceived aims, intents, and sources of tailored health messages. To protect the future of tailored health messages, more research attention must be given to defining these contextual factors and understanding the roles that they play and the ways in which they can be controlled to assure the best outcomes. Such research could point the way towards a set of empirical and ethical "best practices" based on a scientific understanding of how to maximize the benefits, and minimize the potential harms, of the widescale use of tailored health communications. PMID- 10721439 TI - Depression and outpatient medical utilization: a naturalistic 10-year follow-up. AB - The current investigation described the relationship between depression and outpatient medical utilization in a sample of 424 treatment-seeking individuals diagnosed with a depressive disorder and a demographically matched community sample of 424 men and women. This relationship was assessed longitudinally from baseline (intake for the patient sample) to 1-, 4-, and 10-year follow-ups. Patients and community individuals demonstrated distinct patterns of depressive symptoms and outpatient medical utilization: patients declined in symptoms and medical utilization following treatment, although they continued to have higher levels of depressive symptoms and outpatient utilization than controls at each follow-up period. Community controls demonstrated no change from baseline in symptoms or utilization at any follow-up. Higher levels of depressive symptoms was associated with increased outpatient medical utilization over the 10 years, even when age, sex, marital status, medical comorbidity, and patient status were controlled. Results add further evidence for a relationship between symptoms of depression and outpatient utilization by documenting this relationship in a posttreatment sample. Furthermore, the findings underscore the need for long-term follow-ups in investigations of the association between treatment for depression and the outpatient medical utilization of depressed individuals. PMID- 10721438 TI - Family environment, intrusive ideation, and adjustment among renal transplant candidates. AB - Waiting for an organ transplant is a stressful experience frequently associated with symptoms of depression and anxiety. Little empirical work has examined patients during the stressful period prior to transplantation, particularly among patients waiting for a renal transplant. A large body of research has demonstrated that social and family support variables are associated with psychological adjustment in a variety of medical populations. Little research has examined the mechanism by which social support exerts its effects on psychological well-being. We examined two possible models of the role of intrusive thoughts on the relationship between a supportive family environment and both depression and anxiety in a sample of 75 patients with end-stage renal disease (ESRD) waiting for a kidney transplant. Path analyses provided modest support for a mediational model, showing that intrusive thoughts partly accounted for the relationship between family expressiveness and psychological distress. A moderational model examining the interactive effects of family environment and intrusive thinking on adjustment was not supported. PMID- 10721440 TI - Age-related macular degeneration: a randomized clinical trial of a self management intervention. AB - The purpose of this study was to conduct a randomized clinical trial to assess whether a self-management group intervention can improve mood, self-efficacy, and activity in people with central vision loss due to age-related macular degeneration (AMD). Ninety-two elderly patients with AMD (average age = 79) from a university ophthalmology clinic were randomly assigned to the self-management intervention (n = 44) or to a wait-list (n = 48). All patients were legally blind in at least one eye. The intervention consisted of 6 weekly 2-hour group sessions providing education about the disease, group discussion, and behavioral and cognitive skills training to address barriers to independence. All participants eventually completed the intervention allowing pre-post comparisons for all patients. The battery of measures included the Profile of Mood States (POMS); Quality of Well-Being Scale; and assessments of self-efficacy, participation in activities, and use of vision aids. Participants' initial psychological distress was high (mean total POMS = 59.72) and similar to distress experienced by other serious chronic illness populations (e.g. cancer, bone marrow transplant). Analysis of covariance testing the primary hypothesis revealed that intervention participants experienced significantly (p = .04) reduced psychological distress (pre mean = 61.45; post mean = 51.14) in comparison with wait-list controls (pre mean = 57.72; post mean = 62.32). Intervention participants also experienced improved (p = .02) self-efficacy (pre mean = 70.16; post mean = 77.27) in comparison with controls (pre mean = 67.71; post mean = 69.07). Further, intervention participants increased their use of vision aids (p < .001; pre mean = 3.37, post mean = 6.69). This study demonstrates that a relatively brief behavioral intervention can substantially reduce psychological distress and increase self-efficacy in elderly adults experiencing vision loss due to macular degeneration. Self-management intervention appears to improve mood, self efficacy, and use of vision aids, further enhancing the lives of poorly sighted individuals with AMD. PMID- 10721441 TI - Socioeconomic status, hostility, and risk factor clustering in the Normative Aging Study: any help from the concept of allostatic load? AB - OBJECTIVE: To examine the relationships between socioeconomic status (SES), psychosocial vulnerability (hostility), and allostatic load. Allostatic load refers to the cumulative physiological cost of adaptation to stress. METHOD: We examined the relationships between SES (as measured by educational attainment), hostility, and allostatic load in the Normative Aging Study, a longitudinal study of community-dwelling men aged 21 to 80 years and free of known chronic medical conditions at entry in the 1960s. In 1986, the revised Minnesota Multiphasic Personality Inventory was administered by mail, from which a hostility measure was derived by summing the scores from three Cook-Medley subscales: Hostile Affect, Hostile Attribution, Aggressive Responding. An index of allostatic load was constructed from data collected during physical exams conducted between 1987 and 1990 (i.e. measures reflecting "wear and tear" on the cardiovascular, endocrine, and metabolic systems). Cross-sectional relationships between education, hostility, and allostatic load were examined in 818 men. RESULTS: Separate linear regression analyses indicated that lower levels of educational attainment and greater hostility were both associated with higher allostatic load scores (p < .05 and p < .01, respectively). Less education was also associated with higher hostility (p < .001). When allostatic load was regressed simultaneously on education and hostility, the effect of education was attenuated, while hostility (p < .05) maintained an independent effect. CONCLUSIONS: Our findings suggest that lower levels of education and greater hostility are associated with greater "wear and tear" on the body. The effects of education on allostatic load may be mediated by hostility. PMID- 10721442 TI - Predictors of attrition from behavioral medicine treatments. AB - Despite the efficacy of a range of behavioral medicine interventions, high rates of attrition are a persistent problem in both clinical and research settings. Appropriately, studies have begun to focus on predictors of attrition with the hope of identifying important client or treatment characteristics. This article reviews attrition predictors in outpatient behavioral medicine treatments for headache, pain, stress, and weight management. Across all areas, psychological variables and severity of symptom variables were more predictive than demographic variables. However, as 13 of the 20 studies reviewed were in the area of weight management, generalizability of the findings to other treatment areas requires further investigation. Recommendations are made for improving attrition research by (a) developing clinically valid definitions of attrition, (b) recognizing important within-group differences among those who prematurely terminate treatment, and (c) focusing on theoretically grounded psychological and treatment process variables. A working definition of attrition based on the integration of clients' and clinicians' perspectives is also provided. PMID- 10721443 TI - [The best of arterial hypertension in 1999]. AB - THE RELIEF OF THE RESULTS OF THE HOPE TRIAL WITH RESPECT TO THE INCIDENCE OF CEREBROVASCULAR EVENT UNDER ACE INHIBITOR THERAPY: In 1998, the CAPP trial had raised a serious concern about whether captopril therapy increased the risk of cerebrovascular accidents. When compared with betablocker therapy (+/- diuretics) in 11,000 hypertensives, there was a very worrying number of excess cerebrovascular accidents in the captopril group (+25%) (with no difference in the number of cerebrovascular accidents overall). There were several reasons which led to believe that the captopril was not the causal factor. But a doubt remained. In 1999, the results of the HOPE trial with ramipril, though not primarily for a hypertensive population, provided reassurance beyond the investigators' hopes concerning the value of ACE inhibitors in the prevention of vascular events, including cerebrovascular accidents. THE FRAMINGHAM EXPERIENCE OF LVH AND THE TREATMENT OF HYPERTENSION: The Framingham study reported unique data concerning the effects of antihypertensive therapy on LVH in 10,333 subjects of 45 to 74 years of age followed up for 40 years (1950-1989). As the incidence of antihypertensive therapy increased during the observation period, that of hypertension and LVH decreased in parallel. Although these data were retrospective, they are compatible with a causal relationship between the treatment and regression of LVH. This could explain up to 50% of the decrease in cardiovascular mortality observed in the United States during this period. The Framingham study so reposition, in an epidemiological context, the considerable benefits of antihypertensive therapy and of the regression of the associated LVH. THE SEVERITY OF THE WHO AND IHS RECOMMENDATIONS: Less than 130/85: this is the target value of the blood pressure in adults under antihypertensive therapy according to WHO and IHS. In patients with diabetes or renal failure with proteinuria > 1 g/j, the target is even lower. These recommendations incite physicians to beware of any laxness in the treatment of hypertension. The most recent epidemiological data from France indicates that only a minority of the hypertensive patients under treatment are well controlled and that this advice is probably not superfluous. Moreover, in the decision to treat hypertension (drug therapy or not), these recommendations underline the evaluation of the individual risk of the subject on the basis of associated risk factors, target organ complications and previous history of vascular events. PMID- 10721444 TI - [The best of cardiac failure in 1999]. AB - Cardiac failure has become a major but underestimated public health problem. The EPICAL study in France confirmed the severity of this condition. In addition to the neuro-hormones, the role played by inflammatory cytokines in the progression of the disease has been emphasized. The importance of the "genetic background" in the development and evolution of cardiac failure has been demonstrated (deletion or prospective polymorphism). From the therapeutic point of view, besides the hopes raised by multisite pacing, the betablockers and spironolactone have been shown to provide major functional improvement and prolonged survival, and they take their place with the angiotensin converting enzyme inhibitors in our pharmacological arsenal. Cellular transplantation is associated with encouraging pre-clinical results which open up a new field of interest. Finally, global management, including physical rehabilitation and patient education by plury disciplinary teams, provides medical and economic benefits. The year 1999 has been particularly rich in the field of cardiac failure from the basis of fundamental research to the organisation of health care. PMID- 10721445 TI - [The best of arrhythmias in 1999]. AB - Recent studies of atrial fibrillation, or rather "atrial fibrillations", have been based on focal atrial fibrillation which seems to be one of the commonest forms of paroxysmal atrial fibrillation. This observation led to the possibility of ablative therapy, usually near the pulmonary veins. Technological advances are awaited before a wider diffusion of this therapy becomes possible. In the field of defibrillation, the MUSTT study demonstrated the value of implantable defibrillators in the prevention of sudden death, in this trial in the case of asymptomatic high risk patients after myocardial infarction, with a decreased ejection fraction and non-sustained ventricular tachycardia. Two large scale randomised controlled trials (MERIT-HF and CIBIS II) have confirmed the value of betablockers in preventing sudden death, in patients with moderate or severe cardiac failure in association with classical treatment by diuretics, ACE inhibitors and digitalis. Similarly, for spironolactone, the RALES study showed a reduced incidence of sudden death in cardiac failure, as its physiopathological modes of action suggested. New techniques of three-dimensional mapping have confirmed their value. Finally, significant progress has been made in the field of genetics of cardiac arrhythmias, indicating that, in years to come, it will be possible to identify most of the genes responsible for conditions predisposing to arrhythmias. PMID- 10721446 TI - [The best of echocardiography in 1999]. AB - As many techniques of medical investigation, echocardiography regularly benefits from technical innovations which, with application, prove to be extremely useful and, for some of them, even widen the field of investigation. The end of this decade has seen the introduction of major improvements. In daily practice, second harmonic imaging has been the most important technical advance with such improved quality of imaging that this mode has rapidly become the routine for transthoracic investigations in adults. All modern echocardiographs are, or can be, equipped at modest cost. Stress echocardiography, the diagnostic reliability of which is closely related to the quality of the imaging, has greatly benefited from this technique, to the point of obtaining equivalent results as nuclear medicine in the detection of myocardial ischaemia and cellular viability. The results are now sufficiently convincing for the technique to have a real prognostic value in myocardial ischaemia. Doppler tissue imaging is also a major advance but the clinical value is still under evaluation: the pulsed Doppler mode is quantifiable during the investigation, contrary to the calculation of transparietal velocity gradients which requires computerisation techniques not provided by all manufacturers. The regain in interest in contrast echocardiography is due to the development of agents which, injected intravenously, cross the pulmonary capillary barrier and opacify the left heart chambers. The reinforcement of the Doppler signal and improved detection of the endocardial echoes have justified the authorization of their commercialisation, but the essential point is their use in the investigation of myocardial perfusion which is under evaluation. Three-dimensional reconstruction has made great strides but its diffusion is still limited by the limited availability of the required powerful computers. PMID- 10721447 TI - [The best of cardiac pacing in 1999]. AB - Since the first clinical application to man forty years ago, for the treatment of bradycardia, cardiac pacing has been the object of continuous technological innovation in parallel with those in electronics and computerisation. However, independently of these expected advances, there has been a surprising widening of the field of application of pacing into those of haemodynamics and rhythmology. The recent publication of the long-term results of the Pacing in Cardiomyopathy (PIC) study confirmed the sustained decrease of intraventricular pressure gradient, of NYHA functional stage and improved quality of life of patients with hypertrophic obstructive cardiomyopathy paced in the DDD mode. The investigators also underlined the placebo effect of the pacemaker. The decrease in risk of sudden death and the reduction in ventricular remodelling have not been demonstrated yet. More recently, biventricular pacing has been proposed for the treatment of dilated cardiomyopathy and a French study showed a long-term improvement in NYHA stage and effort capacity. Several prospective randomised trials are under way to validate this indication. Acute haemodynamic evaluations have confirmed the efficacy of biventricular stimulation but also underline the value of left ventricular pacing alone. The effects on mortality, the selection of patients and the optimal configuration of pacing remain to be defined. In the field of prevention of atrial arrhythmias, the results of the multicenter SYNBIAPACE study, investigating biatrial pacing in patients with interatrial conduction defects, only showed a tendency to an increase in the delay before recurrence of atrial fibrillation. The value of the memory functions of pacemakers and the algorithms of prevention of atrial arrhythmias are still under investigation. Haemodynamic transducers have been introduced in some recent pacemakers to assess myocardial contractility and have applications in the evaluation of different pacing modes and in the optimisation of the atrioventricular interval. Their value in the treatment of neurocardiogenic syncope is under evaluation. In conclusion, it is not overoptimistic to imagine that, in the near future, the cardiac pacemaker will be part of a "control and treatment system" well over the limits of treating patients with bradycardia. PMID- 10721448 TI - [The best of coronary artery disease in 1999]. AB - Despite the (modest) regression observed in the MONICA study, coronary artery disease remains the leading cause of mortality in industrialised countries and it is worryingly progressive in emerging countries. Therapeutic progress has been considerable but only a small number of coronary patients benefit from it. The techniques are costly and justify efforts to improve selection of high risk patients. The results of the VA-HIT study, which demonstrated the importance of HDL cholesterol level in prevention, now raise the question of the optimal treatment of these patients and that of the association of fibrates and statins. The value of ACE inhibitors in the HOPE study in coronary artery disease was outstanding. In parallel, the efficacy of Mediterranean diet has been confirmed in secondary prevention as that of supplements of omega-3 fatty acids in the GISSI-Prevenzione study. When reference anti-anginal agents are ineffective in the most resistant forms of coronary artery disease, other classes of drugs have been shown to be effective in association. Finally, the theory of inflammation and infection of atherosclerosis has found new indirect arguments, whereas angiogenesis and vasculogenesis have been confirmed as the main leaders in the future treatment of coronary artery disease. PMID- 10721449 TI - [The best of pediatric cardiology in 1999]. AB - Paediatric cardiology is a dynamic field of progress for results. Those which have marked the year 1999 include the introduction of new techniques of cardiovascular imaging and interventional cardiology, and the new consequences of collaboration with workers in foetal cardiology and medical and molecular genetics. The advances in imaging are the result of those of microprocessors which enable three-dimensional reconstruction of ultrasonic, radiological or magnetic resonance images. This provides intracardiac or intravascular views which are very similar to those seen by the surgeon. This is a major tool for improving the diagnosis and treatment of congenital heart disease. Similarly, the introduction of programmes of tissue recognition enables fine ultrasonic analysis of the vascular wall and of endothelial function leading to the opening of a new chapter of preventive vascular medicine from the earliest age. Paediatric interventional cardiology has also progressed rapidly and the past year has been that of a consensus on the closure of a great number of atrial septal defects by new prostheses implanted and anchored in a simpler and safer manner. Prenatal diagnosis has become a crucial factor in the treatment and prognosis of congenital heart disease which is life-threatening in the first hours of life, explaining the benefit when this is applied to transposition of the great vessels or to coarctation of the aorta. Finally, advances in genetics have led to the identification of several genes of heart malformations and the correlations between interstitial microdeletions and syndromes often associated with heart disease: chromosome 22q11 and the Di George syndrome, chromosome 7q and the Williams syndrome. They have even allowed linking of myocardial and cerebellar abnormalities of a degenerative neuropathy (Friedreich's disease) to an abnormality of the mitochondrial respiratory chain, thus giving the opportunity of a real treatment. PMID- 10721450 TI - [Coronary angioplasty at the dawn of the 21st century]. AB - In about 20 years, transluminal coronary angioplasty has become the most commonly used method world wide for revascularisation. During this short period, the technique has progressed from a problematical technique reserved for rare privileged indications to reliable, safe therapy, the primary results of which are predictable with the widespread use of coronary stents. These rapid strides were made possible by material innovations, the progression of radiological and echographic imaging and, above all, by the professionalism and experience of interventional cardiologists. Besides the immediate impact on the safety of the procedure and quality of the immediate result, the implantation of stents also reduces the risk of restenosis in the long-term. However, the risk is not totally eliminated because although the stent prevents the constrictive scarring remodelling of the dilated artery. It also exacerbates the fibroproliferative phenomena which now play a dominant part in the mechanisms of restenosis. The efforts of research are now directed towards this target: in situ drug delivery, gene therapy, brachytherapy. PMID- 10721451 TI - [The best of thrombosis and thromboembolic disease in 1999]. AB - 1999 has been a good year in the field of innovation in thrombosis. In coronary syndrome without ST elevation: low molecular weight heparin has been confirmed to be more effective than non-fractionated heparin (enoxaparin) and to improve the prognosis of non-revascularised patients (dalteparin) after the hospital phase; hirudin has been shown to be more effective in terms of incidence of myocardial infarction and recurrence of angina than non-fractionated heparin without a higher incidence of bleeding complications; the anti-GP IIb-IIIa (abciximab) has confirmed all its advantages at 6 months and 1 year after a coronary event. The association of heparin and aspirin, which has been the mainstay of antithrombotic treatment of acute coronary syndromes without ST elevation, will soon be improved upon at the beginning of the third millennium. In myocardial infarction, medical thrombolysis has probably reached a turning point in its history. The association of half doses of rt-Pa and anti-GP IIb-IIIa has been shown to be more effective in obtaining good reflow than the thrombolytic agent alone at conventional doses. These results were obtained without any increase in bleeding complication. The same anti-GP IIb-IIIa also improve mechanical revascularisation by optimising reperfusion after the 24th hour. This benefit is rapidly transformed into reduced left ventricular dysfunction. Pulmonary embolism remains a critical illness as the ICOPER registry reports a 3 year mortality of nearly 16%. This emphasises the importance of early diagnosis which is usually possible without resorting to invasive procedures and by modulating all the results of paraclinical investigations with respect to the pretest clinical probability. PMID- 10721453 TI - [The best of non-invasive cardiac imaging in 1999]. AB - The term non-invasive and non-echographic myocardial imaging is used to describe the advances in nuclear cardiology and magnetic resonance imaging underlying the most recent developments in investigating the myocardium. The value of new techniques such as rapid CT scan and cardiological applications of the synchroton are described. The main clinical applications of these techniques in the present and future are reviewed, especially with regards to ischaemic heart disease. PMID- 10721452 TI - [The best of vascular pathology in 1999]. AB - In vascular pathology, the discovery of the ABC1 receptor (ATP-binding-cassette transporter 1), the deficit of which is responsible for Tangier disease and familial hypoalphalipoproteinaemias, has opened the greatest perspectives with the possibility of new active treatments in the prevention of atherosclerosis. Other advances were more expected. A large British trial convincingly demonstrated that the follow-up of small abdominal aortic aneurysms is reliable. The MEDENOX trial showed the value of prophylaxis of thromboembolic disease in a medical setting and the reduced incidence of phlebographic events. The ICAI study, on the other hand, showed the difficulty of treatment of critical ischaemia of the lower limbs: alprostadil (PGE1) was ineffective with a 6 month follow-up in this pathology. Finally, low dose aspirin is at least as effective as high doses. PMID- 10721454 TI - [The best of valvular heart disease in 1999]. AB - The year 1999 confirmed important changes in the clinical presentation, the methods of investigation and the treatment of valvular heart disease. The near disappearance of acute rheumatic fever in the developed world, associated with the increase in life expectancy has resulted in degenerative aetiologies becoming the most common causes of valvular heart disease with a dominance of aortic stenosis and mitral incompetence. The increase in average age of the operated patients explains the increasing role of comorbidity and the higher incidence of mixed (valvular and coronary artery) surgery. Doppler echocardiography is now the reference method of investigating valvular heart disease, both pre- and post operatively (especially in mitral incompetence). The value of tri-dimensional echocardiography is beginning to be recognised. The technical advances in surgical techniques are also important, especially the extension of conservative methods both in mitral incompetence and parietal lesions of dystrophic aortic incompetence. The good long-term results of homograft aortic valves have been confirmed, especially in young patients and infectious endocarditis complicated by abscess. The Ross procedure is an interesting alternative in children and adolescents in the absence of available homografts. The persistence of good results in the long term has made percutaneous mitral commissurotomy the reference in mitral stenosis. The improvement in surgical and interventional methods has widened the operative indications which are now considered in patients who are pauci- or a-symptomatic. PMID- 10721455 TI - The non-enumerable described retrovirus integrase inhibitors are not a lure, as evidenced by ten years of clinical experience. AB - Two retrovirus integrase inhibitors have been discovered on the c-erb proto oncogene test in 1989, acriflavine and hydroxymethylellipticine. A ten-year follow-up of their applications to individual treatments of AIDS patients has provided evidence of their HIV-1 (even AZT-resistant) virostatic efficacy, as of their possible clinical use without toxicity. Hundreds of HIV-1 integrase inhibitors have recently been discovered with an in vitro test. The clinical effect of vitamin B12, which belongs to them, has already been observed in patients with one-year follow-up. A positive coordination of return to this Eden of HIV-1-AIDS complex pharmacology must be organized at the highest scientific, and not at the politico-administrative level. PMID- 10721456 TI - Update on highly active antiretroviral therapy: progress and strategies. AB - HIV patients who have detectable viral loads (1,000-5,000 copies/mL) and/or evidence of immunologic dysfunction (CD4+ T-lymphocyte count < 500/mm3) should be treated with a potent combination antiretroviral regimen. Currently, this is to consist of two nucleoside reverse transcriptase inhibitors (NRTIs) with at least one protease inhibitor (PI), or a non-nucleoside reverse transcriptase inhibitor (NNRTI), or another combination with adequate potency. The specific regimen should be designed to achieve an undetectable viral load by an ultrasensitive method (< 50 copies/mL). It should be chosen with a view towards maximizing adherence and minimizing significant drug-drug interactions and side effects. Long-term adherence with the initial highly-active regimen will minimize development of resistant viral mutants and preclude the use of higher complexity regimens. If a regimen is failing in the setting of adequate patient adherence, a new regimen should be chosen using at least two new medications to which viral strains are likely to be sensitive (by genotyping determination). If this is not possible, one may consider treatment interruption or, depending on the clinical status of the patient, continue the current suboptimal regimen until new drugs become available. The predictive value of resistance testing is currently being examined in patients who have failed their first therapy. Further developments include vaccine, cytokine-, and gene therapy-based treatment strategies. PMID- 10721457 TI - Residual HIV-1 replication. PMID- 10721458 TI - Spontaneous apoptosis and highly active antiretroviral therapy (HAART). AB - Increased programmed cell death (PCD) or apoptosis has been detected in the T cells of HIV-infected subjects; it is held partially responsible for the continuous loss of CD4+ T cells during the natural course of HIV infection. Highly active antiretroviral therapy (HAART) decreases the viral load and leads to an increase of CD4+ count in vivo. In this study we evaluated PCD in total peripheral blood mononuclear cells, CD8+ and CD4+ lymphocytes before and four weeks after initiation of HAART. Seven HIV-1-infected patients were investigated. Viral load was assessed by RT-polymerase chain reaction and PCD by flow cytometry using apoptosis by 7 amino actinomycin D (7AAD) and propidium iodide (PI). After four weeks of HAART, CD4+ T and CD8+ T cell levels were stable, and plasma HIV RNA copies were significantly decreased. In four of the patients (4/7), HIV-RNA levels were reduced to undetectable levels (fewer than 400 copies per milliliter). A statistically significant reduction of apoptosis among CD4+ cells was observed (P < 0.03), though neither in the CD8+ T cell population nor in peripheral blood mononuclear cells (PBMCS). These results demonstrate the beneficial effect of HAART on apoptosis of CD4+ cells in the early treatment stage. PMID- 10721459 TI - Analysis of telomere length and thymic output in fast and slow/non-progressors with HIV infection. AB - There are two models for CD4+ T-cell depletion leading to AIDS: a kinetic model and an immune suppression model. In the kinetic model, direct cell killing due to viral replication results in a continuous demand for CD4+ T-cells, which eventually exhausts their capacity for renewal by proliferative mechanisms. In the immune suppression model, CD4+ T-cell decline is due to an indirect global inhibitory effect of the virus on uninfected immune cell function. In order to address differences in the two models, we investigated proliferative history and thymic output in PBMC from the GRIV cohort of fast (FP) and slow/non-progressors (S/NP), and uninfected controls. Proliferative history and thymic output were assessed by measurement of mean telomeric restriction fragment (TRF) length and T cell receptor Rearrangement Excision Circles (TREC) levels, respectively. Mean TRF lengths were significantly shorter in S/NP (n = 93, 7.59 +/- 0.11 kb) and FP (n = 42, 7.25 +/- 0.15 kb) compared to controls (n = 35, 9.17 +/- 0.19 kb). Mean TRF length in PBMC (n = 9, 7.32 +/- 0.31 kb) and CD4+ enriched fractions (n = 9, 7.41 +/- 0.30 kb) from a subset of non-GRIV HIV-1 infected samples were also significantly smaller than PBMC (n = 8, 9.77 +/- 0.33 kb) and CD4+ fractions (n = 8, 9.41 +/- 0.32 kb) from uninfected controls. Rates of telomeric shortening, however, were similar among S/NP (n = 93, -45 +/- 20 bp/yr), FP (n = 42, -41 +/- 14 bp/yr) and controls (-29 +/- 17 bp/yr). Paralleling differences observed in mean TRF length, TREC levels were significantly reduced in S/NP (n = 10, 3,433 +/ 843 mol/mu and FP (n = 8, 1,193 +/- 413) compared to controls (n = 15, 22,706 +/ 5,089), indicative of a defect in thymopoiesis in HIV-1 infection. When evaluated in the context of reduced thymopoiesis, the difference in mean TRF length between S/NP and controls (1.58 +/- 0.30 kb) is similar to that observed between memory and naive T-cells (1.4 +/- 0.1 kb), and may reflect perturbations in the peripheral T-cell population due to a decline in thymic output of naive T cells rather than increased turnover. Based on the different clinical criteria used to select S/NP and FP, the sight difference in TREC between these two groups suggests the threshold for pathogenesis as a result of naive T-cell depletion may be quite low, and incremental increases in thymic output may yield substantial clinical results. Future studies regarding therapeutic vaccination, specifically with HIV-1 Tat targeted anti-immunosuppressive vaccines, should address the defect in thymic output in HIV-1 infection by using TREC analysis as a rapid method for biological evaluation. PMID- 10721460 TI - Opportunistic AIDS-associated malignancies in HIV-infected patients. AB - Three major neoplasms are associated with AIDS definition in the course of HIV infection due to their increased incidence. The relationship between the immune system and the epidemiology of these virus-induced tumors is of importance in order to identify new therapeutic approaches for treating or preventing these neoplasms. Major improvements in the understanding of the pathogenesis have already been performed. The impact of highly active antiretroviral therapy (HAART) on their incidence likely confirms the concept of 'opportunistic malignancies'. Other neoplasms are likely more prevalent in HIV-infected individuals, but their relative importance requires further prospective case control cohort studies. PMID- 10721461 TI - Procedures for preparing biologically inactive, but immunogenic HIV-1 Tat protein (Tat toxoid) for human use. AB - Extracellular Tat can exercise its deleterious effects on cells surrounding HIV-1 infected cells and allow spreading of virus. Extracellular Tat should be neutralized by anti-Tat vaccination using as an immunogen a functionally inactivated but immunogenic Tat preparation (Tat toxoid). In the present paper we show that native Tat can be inactivated without impairment of its immunogenicity by subjecting it to various chemical treatments. Since the carboxyamidation reaction can be easily monitored and the carboxyamidated Tat retained the whole immunogenicity of the native molecule, it should be the toxoid of choice for mass immunization. PMID- 10721462 TI - The effectiveness of desensitization versus rechallenge treatment in HIV-positive patients with previous hypersensitivity to TMP-SMX: a randomized multicentric study. C.I.S.A.I. Group. AB - Our study was undertaken to evaluate if desensitization treatment is more effective than rechallenge in preventing hypersensitivity reactions in HIV positive patients with previous allergic reactions to TMP-SMX; the secondary aim was to evaluate the frequency of reactions to TMP alone. This was a randomized, multicentre open study. Patients with previous documented hypersensitivity to TMP SMX who required primary or secondary PCP prophylaxis were enrolled; subjects who had previously had serious adverse reactions to TMP-SMX were excluded. All eligible patients assumed 200 mg TMP for 14 days and in case of no reactions were randomized for desensitization or rechallenge with TMP-SMX. The patients were then followed up by periodical visits for six months. Seventy-three patients were enrolled; 14 subjects (19%) presented reactions on TMP alone during the pre enrollment phase. The remaining 59 subjects were randomly assigned to the two treatment groups: 34 carried out desensitization (group 1) and 25 rechallenge (group 2) with TMP-SMX. Seven patients in group 1 (20.5%) and seven in group 2 (28%) showed hypersensitivity reactions during treatment; this difference was not statistically significant. No serious reaction occurred in either group. This study showed the comparable effectiveness of the desensitization procedure and rechallenge in patients with a previous, not serious, allergic reaction to TMP SMX. PMID- 10721463 TI - Application of monoclonal antibodies to monitor the synthesis of a glycoprotein core of envelope glycoproteins of human immunodeficiency virus (HIV-1). AB - Using monoclonal antibodies 0.5 beta or G3-42, directed against V3 and C4 domains of glycoprotein 120 (gp120), we monitored the synthesis of oligomeric and monomeric forms of HIV-1 envelope glycoprotein 120 by flow cytometry or immunoprecipitation analysis in chronically infected MoIT-4 cells, cultured in the presence of tunicamycin. We observed that the inhibition of glycosylation by high concentrations of tunicamycin results in the reduction of an oligomeric gp120 on the surface of infected MoIT 4 cells as well as the decrease in the concentration of a monomeric form in the cytoplasm. Our studies revealed that the antibody 0.5 beta (exhibited higher sensitivity in the detection of gp120 than the antibody G3-42). We also observed that both antibodies did not recognise nonglycosylated precursor core envelope protein. PMID- 10721464 TI - The cause of invasive cervical cancer could be multifactorial. AB - Cancer of the cervix is the third most common cancer among women worldwide, with incidence rates ranging from 3.8 per 100,000 women per year in Israel to 48.2 per 100,000 per year in Colombia. Epidemiologic and clinical data suggest that human papillomaviruses, especially HPV-16 and HPV-18, play the major role in the etiology of cervical cancer. However, many investigators acknowledge that HPV is neither necessary nor sufficient in the etiology of cervical cancer and that a multifactorial etiology is likely. HPV cannot be found in every patient with the disease and other factors, such as herpes simplex virus type 2 infection, cigarette smoking, vaginal douching, nutrition, and use of oral contraceptives, have been associated with cervical cancer. In two different animal models, tumors can be produced following exposure to DNA viruses and tars. Using those animal models as prototypes, we propose that the etiology of cervical cancer in humans could be an interaction between DNA viruses, specifically papillomavirus and/or HSV-2 infection, and tar exposure through cigarette smoking and/or tar-based vaginal douching. PMID- 10721465 TI - The absent patient: a meditation on a Chardin painting. PMID- 10721466 TI - A sentimental patient. PMID- 10721467 TI - The creation of partial patients. PMID- 10721469 TI - An ethical analysis of the barriers to effective pain management. PMID- 10721468 TI - Morally managing medical mistakes. PMID- 10721470 TI - Managed care: effects on the physician-patient relationship. PMID- 10721471 TI - Alternative medicine or alternatives to medicine? A physician's perspective. PMID- 10721472 TI - The culture of physician autonomy; 1900 to the present. PMID- 10721473 TI - Response to "commentary on Thomson's Violinist and Conjoined Twins" by John K. Davis (CQ vol 8, no 4). PMID- 10721475 TI - The illegal alien who needs surgery. PMID- 10721474 TI - Response to "Difference and the Delivery of Healthcare" (special section) (CQ Vol 7, No 1). PMID- 10721476 TI - Serogroups of the beer spoilage bacterium Megasphaera cerevisiae correlate with the molecular weight of the major EDTA-extractable surface protein. AB - Megasphaera cerevisiae is a Gram-negative obligate anaerobe that causes turbidity and off-flavour and aroma in beer. Seven isolates of M. cerevisiae were obtained worldwide, and their extractable surface antigens were focused upon to determine if there is more than one serogroup of this bacterium. Sodium dodecyl sulphate polyacrylamide gel electrophoresis of ethylenediaminetetraacetic acid (EDTA) bacterial extracts revealed a predominant protein with apparent molecular weights of 46,000, 45,000, and 43,000 for three, two, and two isolates, respectively. When mouse anti-serum generated against any of the EDTA extracts was reacted with denatured bacterial proteins in immunoblots, all bacterial isolates exhibited extensive cross-reactivity involving three antigens, one being the major EDTA extractable protein. In contrast, when the sera were tested for surface reactivity with intact bacteria, three cross-reactivity groups were observed, with the groups individually comprised of bacteria having the same size major EDTA-extractable surface protein. When BALB/c mice immunized with a bacterium from each of the three serogroups were used for monoclonal antibody (Mab) hybridoma production, bacterial surface-reactive Mabs were obtained whose reactivities parallel the three polyclonal antibody-defined serogroups. Through combining these surface-reactive Mabs, it will be possible to rapidly detect and identify beer contamination by M. cerevisiae belonging to any serogroup. PMID- 10721477 TI - Chemical components and their locations in the Verticillium fungicola cell wall. AB - The chemical structure of cell walls and fractions of Verticillium fungicola, a pathogen of Agaricus bisporus, as well as their corresponding ultrastructures were studied. There are at least three chemically distinct types of carbohydrate polymers: one yielding mannose with lower amounts of galactose and glucose (glucogalactomannan), another one composed mainly of glucose (glucan), and a third one containing only N-acetylglucosamine (chitin). Attempts were made to locate these materials in situ by comparing electron micrographs of shadowed and sectioned cell walls, and also by indirect immunofluorescence. It was shown that none of these polymers constituted a completely physically distinct layer, but there seem to be different solubility properties in the outer, inner, and intermediate layers. It was also shown that fibrillar material (chitin) embedded in cementing glucan constituted the residual inner fraction of the original wall material. Indirect immunofluorescence showed the location of a significant amount of glucogalactomannan on the surface of the walls in which rodlet structures were visualized by electron microscopy. PMID- 10721478 TI - Tween 80 enhanced TNT mineralization by Phanerochaete chrysosporium. AB - The effect of a nonionic surfactant (Tween 80) on 2,4,6-trinitrotoluene (TNT) mineralization by the white-rot fungus Phanerochaete chrysosporium strain BKM-F 1767, was investigated in a liquid culture at 20, 50, and 100 mg TNT.L-1. The presence of 1% (w/v) Tween 80, at 20 mg.L-1 TNT, added to a 4-d-old culture, allowed the highest TNT mineralization level, that is 29.3% after 24 d, which is two times more than the control culture, without Tween 80 (13.9%). The mineralization of TNT resumed upon additional Tween 80 supplementation, consequently, 39.0% of the TNT was respired on day 68. Orbital agitation of the fungal culture was found detrimental to TNT mineralization, with or without Tween 80 in the culture medium. The surfactant also stimulated the growth of P. chrysosporium without any notable effect on either the glycerol consumption rate or the extracellular LiP and MnP activity levels. Respirometric assays highlighted some differences between the oxygen uptake rate of the fungal culture supplemented with or without Tween 80. PMID- 10721479 TI - Origin of p-cresol in the anaerobic degradation of trinitrotoluene. AB - p-Cresol was repeatedly detected as a trace metabolite in anaerobic slurry reactors treating 2,4,6-trinitrotoluene (TNT)-contaminated soils. This study shows that p-cresol was not a metabolite of the anaerobic degradation of TNT, by using a combination of analytical techniques and 13C-labelled TNT. Instead, p cresol, an intermediate in the degradation pathway of some amino acids, was shown to be inhibited by TNT and its metabolites. The range and persistence of inhibition to p-cresol microbial degradation decreased with the level of amino substitution of the derivatives. This explains why p-cresol accumulated within the TNT-treating anaerobic bioslurry, as it could not be further biodegraded in the presence of TNT. PMID- 10721480 TI - Biological control of fusarial wilt of pigeon pea by Bacillus brevis. AB - A virulent strain of pigeon pea wilt pathogen was isolated from wilted pigeon pea plants and was identified as Fusarium oxysporum f. sp. udum. Many bacterial cultures showing antagonism to the pathogen were isolated from various ecological niches. When tested under pot and field conditions, development of fusarial wilt symptoms was prevented in pigeon pea seeds treated with one such antagonist, Bacillus brevis. A formulation of B. brevis with vermiculite as a carrier had a shelf life of at least 6 months. Bacillus brevis produced an extracellular antagonistic substance which induced swelling of the pathogen's hyphal tips, and cells were bulbous and swollen with shrunken and granulated cytoplasm. The antagonistic substance also inhibited germination of conidia, and was fungicidal to the vegetative mycelia of the pathogen. Comparison of the properties of our antagonistic substance with that of known antibiotics produced by B. brevis suggests that our antagonistic substance is a novel compound. The observations reported here indicate that this strain of B. brevis may have potential as a biocontrol agent against fusarial wilt in pigeon pea. PMID- 10721481 TI - Molecular analysis and development of 16S rRNA oligonucleotide probes to characterize a diclofop-methyl-degrading biofilm consortium. AB - Genomic DNA from nine individual bacteria, isolated from a diclofop-methyl degrading biofilm consortium, was extracted for genetic characterization. The degradation of diclofop-methyl produces metabolites that are known intermediates or substrates for bacteria that degrade a variety of chlorinated aromatic compounds. Accordingly, oligonucleotide primers were designed from specific catabolic genes for chlorinated organic degradation pathways, and tested by PCR to determine if these genes are involved in diclofop-methyl degradation. DNA homology between the PCR products and the known catabolic genes investigated by Southern hybridization analysis and by sequencing, suggested that novel catabolic genes are functioning in the isolates. Specific fluorescent oligonucleotides were designed for two of the isolates, following 16S rDNA sequencing and identification of each of the isolates. These probes were successfully used for fluorescent in situ hybridization (FISH) studies of the two isolates in the biofilm consortium. PMID- 10721483 TI - Construction of an equalized cDNA library from Colletotrichum lagenarium and its application to the isolation of differentially expressed genes. AB - To establish an efficient screening system for differentially expressed genes of a phytopathogenic fungus Colletotrichum lagenarium, we constructed an equalized (normalized) cDNA library from C. lagenarium and used this library for differential screening. For the isolation of genes involved in infection-related developments of conidia, conidia undergoing appressorium differentiation were selected as the source of materials for construction of the cDNA library. The equalization of cDNA was performed twice using a kinetic method, and the products were cloned into a plasmid vector. Colony hybridization with nine probes of different abundance showed a reduction in abundance variation from at least 276 fold in the original library to 10-fold in the equalized cDNA library, which demonstrated that the cDNA was successfully equalized. By differential hybridization of 1900 cDNA clones in the equalized cDNA library and RNA blot analysis of candidate clones, we identified 11 independent cDNA clones, designated CAD1 through CAD11, that were expressed in appressorium differentiating conidia, but not in vegetative mycelia. The transcripts of CAD1 and CAD2 hardly accumulated in preincubated conidia, whereas those of CAD3 and CAD4 accumulated highly and slightly, respectively. The amount of the four CAD transcripts increased at the early stage of the appressorium formation process. Sequence analysis of CAD1 revealed that CAD1 would encode for 101 amino acid polypeptides, which showed homology to metallothioneins. Deduced amino acid sequence of CAD2 would encode 278 amino acid polypeptides, and showed high homology to genes in aflatoxin, and sterigmatocystin gene clusters of Aspergillus parasiticus and A. nidulans, respectively. PMID- 10721482 TI - Is Fusarium culmorum isotrichodermin-15-hydroxylase different from other fungal species? AB - Fusarium spp. are ubiquitous fungi infecting cereals and grains, and therefore constitute a major problem for agriculture. Their trichothecene metabolites, and in particular deoxynivalenol and its 3-acetylated derivative, are the mycotoxins involved. The major metabolite produced by Fusarium culmorum is 3 acetyldeoxynivalenol. Studies in vivo with Fusarium culmorum have established that its tricyclic intermediate, isotrichodermin, is a major biosynthetic precursor, which is hydroxylated at position 15 to give 15-deacetylcalonectrin, prior to being converted to the product. In a preliminary in vitro investigation of the cell-free system involved in this transformation, we suggested that cytochrome P450 enzymes are not involved. In this paper, the isotrichodermin-15 hydroxylase from the microsomal fraction of Fusarium culmorum was solubilized and partially purified (60 fold). Our studies with cofactors indicate that this enzyme is a flavoprotein, and the inducers tested highly indicate that indeed the hydroxylase is not attached to cytochrome P450. This is particularly interesting, since the only other enzyme catalyzing the same reaction isolated from Fusarium sporotrichiodes is attached to cytochrome P450. PMID- 10721484 TI - Inactivation of MS-2 phage and poliovirus in groundwater. AB - Since temperature affects the inactivation rate of viruses in natural water systems, the aim of this study was to determine if a temperature shift could influence the structural integrity of model viruses. When crude lysates of MS-2 phage were seeded into groundwater microcosms and incubated at 27 degrees C, complete virus inactivation took place in eight days. The temperature was then shifted to 4 degrees C. Three days after the temperature shift, a two-log increase in virus titer (reactivation) occurred. However, when purified MS-2 lysates were added to groundwater microcosms, no reactivation was obtained. No reactivation of poliovirus took place when similar microcosm experiments were done. The sedimentation coefficients of MS-2 shifted from 80S to 58S, 48S, 37S, 32S, and 18S as inactivation proceeded in groundwater and distilled water controls. Similarly, the sedimentation coefficients of polioviruses changed from 156S to 142S, 135S, 117S, 105S, 95S, and 80 S as inactivation took place. There was no correlation between % virus inactivation and % decrease in virions with intact sedimentation coefficients, as reported earlier for poliovirus inactivated by chlorine. The results presented support our hypothesis that virus inactivation proceeds gradually, involving the rearrangement and (or) loss of capsomere components that may eventually lead to the ejection of nucleic acids. The intermediate particles generated as inactivation proceeds may be in a reversibly inactivated state, and may revert back to a fully infectious state when chemical components stabilize the virus particle. PMID- 10721485 TI - PCR cloning, heterologous expression, and characterization of isopenicillin N synthase from Streptomyces lipmanii NRRL 3584. AB - A key step which involves the cyclization of delta-(L-alpha-aminoadipyl)-L cysteinyl-D-valine to the bicyclic ring structure of isopenicillin N in the penicillin and cephalosporin biosynthetic pathway, is catalyzed by isopenicillin N synthase (IPNS). In this study, an IPNS gene from Streptomyces lipmanii NRRL 3584 (slIPNS) was cloned via PCR-based homology cloning, sequenced and expressed in Escherichia coli. Soluble slIPNS was overexpressed up to 21% of total soluble protein, and verified to be functionally active when in an IPNS enzymatic assay. Sequence comparison of the slIPNS gene obtained (excluding the consensus primer sequences) with another cloned IPNS from S. lipmanii 16884.3, revealed one three nucleotide deletion and three closely-spaced single nucleotide deletions. Furthermore, this paper also reports the first instance of the usage of PCR as an alternative and rapid strategy for IPNS cloning using consensus primers. PMID- 10721486 TI - Defining a novel domain of staphylococcal toxic shock syndrome toxin-1 critical for major histocompatibility complex class II binding, superantigenic activity, and lethality. AB - Staphylococcal toxic shock syndrome toxin-1 (TSST-1) is implicated in the pathogenesis of superantigen-mediated shock. We previously identified TSST-1 residues G31/S32 to be important for major histocompatibility complex (MHC) class II binding, as well as superantigenic and lethal activities. However, the site directed TSST-1 mutant toxin, G31R, could still induce mitogenesis and low-level TNF alpha secretion, suggesting that additional MHC class II binding sites other than G31/S32 may exist. In the current study, a TSST-1-neutralizing monoclonal antibody, MAb5, was found to inhibit TSST-1 binding to human peripheral blood mononuclear cells, neutralize TSST-1-induced mitogenesis and cytokine secretion, and protect against TSST-1-induced lethality in vivo. Epitope mapping revealed that MAb5 bound to TSST-1 residues 51-56 (T(51-56); 51YYSPAF56). Peptide T(51-56) was synthesized and found to also inhibit TSST-1 binding to human monocytes as well as TSST-1-induced mitogenesis, cytokine secretion, and lethality in vivo. This T(51-56) epitope, located within the beta 3/beta 4 loop, and the previously identified G31/S32 epitope, within the beta 1/beta 2 loop of TSST-1, are separated within the primary sequence, but spatially juxtaposed to each other. Collectively, these findings suggest that a discontinuous epitope comprising of regions within both the beta 1/beta 2 and beta 3/beta 4 loops, are critical for MHC class II binding, and the consequent superantigenic and lethal activities of TSST-1. PMID- 10721487 TI - The effects of ultraviolet radiation on the moderate halophile Halomonas elongata and the extreme halophile Halobacterium salinarum. AB - Both the moderately halophilic bacterium, Halomonas elongata, and the extremely halophilic archaea, Halobacterium salinarum, can be found in hypersaline environments (e.g., salterns). On complex media, H. elongata grows over a salt range of 0.05-5.2 M, whereas, H. salinarum multiplies over a salt range of 2.5 5.2 M. The purpose of this study was to illustrate the effect that solar (UV-A and UV-B) and germicidal radiation (UV-C) had on the growth patterns of these bacteria at varied salt concentrations. Halomonas elongata grown on a complex medium at 0.05, 1.37, and 4.3 M NaCl was found to be more sensitive to UV-A and UV-B radiation, as the salt concentration of the medium increased. Halobacterium salinarum grown on a complex medium at 3.0 and 4.3 M NaCl did not show a significant drop in viability after 39.3 kJ.m-2 of UV-A and UV-B exposure. When exposed to UV-C, H. elongata exhibited substantially more sensitivity than H. salinarum. In H. elongata, differential sensitivity to UV-C was observed. At 0.05 M NaCl, H. elongata was less sensitive to UV-C than at 1.37 and 4.3 M NaCl. Both bacteria showed some photoreactivation when incubated under visible light following both UV-A, UV-B, and UV-C exposure. Mutagenesis following UV-C exposure was demonstrated by both organisms. PMID- 10721488 TI - Melanoma cell autonomous growth: the Rb/E2F pathway. AB - Transformation of normal melanocytes to metastatic melanoma cells is characterized by loss of dependency on external growth factors required for the viability and proliferation of normal melanocytes. The molecular events that lead to melanoma cell autonomous growth are not well defined, but are likely to include sustained activity of cyclin-dependent kinases (CDK2, CDK4 and CDK6) as a result of loss of CDK inhibitors (such as p16INK4a and possibly p27KIP1), and persistent upregulation of several cyclins (cyclin D1, cyclin A and cyclin E), the positive regulators of CDKs. CDKs phosphorylate, and consequently, inactivate the retinoblastoma family of tumor suppressor proteins (pRb, p107 and p130), termed pocket proteins. The inactivation of pocket proteins liberates E2F transcription factors from suppressive complexes ('free' E2F) that, in turn, induces the continuous expression of target genes whose products promote cell cycle progression. In normal melanocytes, external growth factors suppress the activity of all three pocket proteins, allowing E2F activity to accumulate and sustain transcription of target genes required for cell proliferation. In contrast, in melanoma cells from advanced lesions, all three pocket proteins are highly phosphorylated and inactive, even in the absence of environmental mitogens, and free E2F activity is constitutively high. Manipulations of normal mouse melanocytes in vitro, and in vivo in transgenic mouse expressing ectopic genes, further support the notion that growth rate, and release from dependency on external mitogens, positively correlate with inactivation of pocket proteins. The latter has been accomplished by sustained cell surface receptor stimulation, such as constitutive high expression of a growth factor, or by sequestration with dominantly acting viral proteins. Taken together, chronic hyperphosphorlyation/inactivation of pRb, p107 and p130 is probably one of the key events in converting growth-factor dependent normal melanocytes, to autonomously growing melanoma cells. Since all pocket proteins are regulated by CDKs activity, it is likely that agents that inhibit this class of enzymes will be effective in treating melanoma patients. PMID- 10721489 TI - Cell adhesion molecules in the development and progression of malignant melanoma. AB - Cell adhesion molecules belonging to the integrin, cadherin and immunoglobulin superfamilies have been implicated in tumor progression in cutaneous melanoma. Expression of the alpha v beta 3 integrin first appears with the change from radial to vertical growth, a step which is associated with the development of metastatic potential. VLA-4 expression is characteristic of advanced primary tumors and may mediate interaction of the tumor cells with VCAM-1 on vascular endothelium. Expression of these integrins is a marker of poor prognosis in patients and can confer invasive (alpha v beta 3) and metastatic (VLA-4) properties to human melanoma cells injected into nude mice. Expression of the immunoglobulin superfamily molecules MUC18/MCAM and ICAM-1 are associated with primary tumors and metastases. MUC18/MCAM expression confers metastatic potential and increased tumorigenicity to human melanoma cells. Expression of ICAM-1 has been shown to be a marker of poor prognosis in stage I tumors and interfering with its expression inhibits experimental metastasis by melanomas in nude mice. E cadherin is used by epidermal melanocytes to interact with neighboring keratinocytes. Changes in E-cadherin expression and cellular localization is first observed in the radial growth phase, the earliest stage in melanoma development. Loss of E-cadherin function is associated with upregulation or induction of MUC18/MCAM and alpha v beta 3 in melanocytic cells in vitro and with alterations in the levels and cellular distribution of the transcriptional regulator beta-catenin in melanomas in vivo. These observations suggest that disturbances in E-cadherin function is not only important in carcinomas but may also be a critical event in melanoma tumor progression. PMID- 10721490 TI - Molecular role(s) for integrins in human melanoma invasion. AB - There are fundamental issues regarding the role of integrins in human disease which remain to be elucidated. Human cutaneous melanoma is an attractive model for studying integrin involvement in tumor progression because it generally follows a sequential series of definable stages. Furthermore, the most specific marker for the transition of cells from the more benign, non-metastatic radial growth phase stage to the more malignant, metastatically competent vertical growth phase stage is associated with the onset of alpha v beta 3 integrin expression and function. This same pattern, however, does not hold true for human ocular/uveal melanomas which do not progress through these stages, but preferentially metastasize to the liver by dissemination of the cells via a direct hematogenous pathway. It is also unclear whether the alpha v beta 3 integrin is functionally involved in uveal melanoma metastasis or not. Our results show that perturbation of the alpha v beta 3 integrin on moderately invasive A375M human cutaneous melanoma cells with either specific antibodies or ligands results in an increase in the cells' ability to invade in vitro coincident with an increase in the cells' expression and extracellular levels of matrix metalloproteinase-2 (MMP-2, gelatinase A). The highly invasive C8161 human cutaneous melanoma cells express little-to-no alpha v beta 3 integrin, but are more invasive and express higher levels of MMPs after perturbation of their alpha 5 beta 1 integrin. This augmented invasiveness could subsequently be abrogated with a function-blocking anti-MMP-2 antibody. Primary uveal melanoma cells and cells derived from uveal metastases appear to grow in either a spindle or epithelioid morphology. The less invasive uveal melanoma cells are spindle shaped and express higher levels of the alpha v beta 3 integrin, while the more invasive cell lines are epithelioid shaped and express reduced levels of the alpha v beta 3 integrin. The apparent conflict between these results and the current model for cutaneous melanoma progression may be addressed as follows: The expression and function of the alpha v beta 3 integrin plays an important role(s) during the transition of cells from the radial growth phase stage to the vertical growth phase stage. However, further progression leading to metastases may require changes in the cells' integrins that would facilitate their ability to leave the primary tumor, and aid in their ability to invade and ultimately form metastases. It is also conceivable that the alpha v beta 3 integrin is reexpressed during various stages of metastatic dissemination, and, in particular, during tumor reestablishment. PMID- 10721491 TI - Role of AP-2 in tumor growth and metastasis of human melanoma. AB - We previously demonstrated that expression of the cell surface adhesion molecule MCAM/MUC18 correlates directly with the metastatic potential of human melanoma cells. In addition, the progression of human melanoma towards the metastatic phenotype is associated with loss of expression of the tyrosine-kinase receptor c KIT. This review summarizes our recent data demonstrating that the expression of both genes is regulated by the AP-2 transcription factor. Moreover, we have observed a loss of AP-2 expression in metastatic melanoma cells. Re-expression of AP-2 in the highly metastatic A375SM cells decreased their tumorigenicity and inhibited their metastatic potential in nude mice. MCAM/MUC18 mRNA and protein expression was significantly downregulated while c-KIT expression was upregulated in the AP-2 transfected cells. Since AP-2 also regulates other genes that are involved in the progression of human melanoma such as E-cadherin, MMP-2, p21WAF 1, HER-2, BCL-2, and insulin like growth factor receptor-1, we propose that loss of AP-2 is a crucial event in the development of malignant melanoma. PMID- 10721492 TI - The biology of melanoma brain metastasis. AB - Brain metastases are clinically diagnosed in the majority of patients with metastatic melanoma. The prognosis for patients with melanoma brain metastasis is poor with a median survival time of 6 months after diagnosis. Development of better therapies requires a better understanding of the biology of melanoma brain metastasis. The development of a relevant in vivo model offers this possibility. The intracarotid injection of different murine or human melanoma cells into syngeneic or nude mice produces metastases in different regions of the brain. This site-specific metastasis is not due to patterns of initial cell arrest, motility, or invasiveness, but rather to the ability of melanoma cells to proliferate in the brain parenchyma or the meninges. The blood-brain barrier is intact in metastases that are smaller than 0.25 mm in diameter. Although in larger metastases the blood-brain barrier is leaky, the lesions are resistant to many chemotherapeutic drugs. We have also analyzed the malignant behavior of several melanoma cell lines isolated from brain or visceral metastases of patients. The cells from brain metastases showed a slower growth rate and exhibited lower metastatic potential than cells from visceral metastases, indicating that brain metastases do not necessarily represent the end stage in the metastatic cascade. Rather, brain metastases are likely to originate from a unique subpopulation of cells within the primary neoplasm. PMID- 10721493 TI - Human xenografts, human skin and skin reconstructs for studies in melanoma development and progression. AB - Melanoma develops from a series of architectural and phenotypically distinct stages and becomes progressively aggressive. Considerable progress has been made in understanding the biological, pathological, and immunological aspects of human melanoma. Genetic and cytogenetic studies have revealed broad chromosomal abnormalities and wide mutational spectra. Precise biological and molecular determinants responsible for melanoma progression are not yet known. This is in part due to lack of experimental models that mimic human melanomas. Experimental models in melanoma should not only identify cause and origin of malignancy, but also should represent the ordered progression steps that culminate in metastasis to distant organs. Currently, there are several mouse and other vertebrate melanoma models under investigation; several of them promise to shed light on mechanisms of melanomagenesis. However, many of them suffer from lack of context to human skin architecture and hence, are of basic interest. The lack of appropriate models impeded the efforts to understand origin, etiology, progression and ultimately therapeutic benefits to humans. Development of human skin-mouse chimeric models has appeal because it mimics human diseases. In addition, human artificial skin constructs in vitro promises to be a versatile and efficient model to study not only origin and mechanisms of melanoma, but also progression. This review will focus on the recent progress in establishing tumor models in melanoma in general and their relevance to human melanoma as molecular determinants of tumor progression. PMID- 10721494 TI - The co-occurrence of substance use in other mental disorders: editor's introduction. AB - The co-occurrence of substance use disorders with other mental disorders has a profound impact on theoretical and conceptual issues in the study of psychopathology and poses unique problems regarding assessment and treatment. Given the high rate of comorbidity, it is not possible to simply treat comorbid substance use as error variance that must be eliminated or controlled for in psychopathology research. Rather, it is proposed that the direct study of the co occurrence of substance use disorders with other mental disorders will enhance the generalizability of research findings and allow for a better understanding of causal processes underlying comorbidity. The contributors to this Special Issue of Clinical Psychology Review have marshaled the latest evidence concerning comorbid substance use in a range of disorders and each contributor proposes integrative perspectives that will facilitate future research. PMID- 10721495 TI - The relationship between anxiety disorders and alcohol use disorders: a review of major perspectives and findings. AB - It is generally agreed that problems related to alcohol use and anxiety tend to occur within the same individual ("comorbidity"); however, the cause of this association remains controversial. Three prominent perspectives are that anxiety disorder promotes pathological alcohol use, that pathological alcohol use promotes anxiety disorder and that a third factor promotes both conditions. We review laboratory, clinical, family, and prospective studies bearing on the validity of these explanatory models. Findings converge on the conclusion that anxiety disorder and alcohol disorder can both serve to initiate the other, especially in cases of alcohol dependence versus alcohol abuse alone. Further, evidence from clinical studies suggests that anxiety disorder can contribute to the maintenance of and relapse to pathological alcohol use. Relying heavily on pharmacological and behavioral laboratory findings, we tentatively propose that short-term anxiety reduction from alcohol use, in concert with longer-term anxiety induction from chronic drinking and withdrawal, can initiate a vicious feed-forward cycle of increasing anxiety symptoms and alcohol use that results in comorbidity. PMID- 10721496 TI - The comorbidity of depression and substance use disorders. AB - Depression and substance use disorders are highly prevalent in the general population and often co-occur within the same individual. This association is most commonly explained either by a causal relationship or a shared etiologic factor underlying both disorders. In light of these mechanisms of association, this article summarizes evidence from clinical, epidemiologic, and genetic epidemiologic studies. Details of a large family study designed to addresses key methodological and conceptual issues identified in the review are also presented. The association of alcoholism with depression is likely to be attributable to causal factors rather than a shared etiology, but the scarcity of information for other classes of substance use disorders precludes similar conclusions regarding their association with depression. The lack of unidirectional and consistent patterns of association for depression and substance use disorders indicates that multiple mechanisms of comorbidity are likely to be simultaneously active in this population. PMID- 10721497 TI - The co-occurrence of bipolar and substance use disorders. AB - Substance use disorders are exceptionally common in bipolar patients. Although the frequency of this co-occurrence is well-documented, the reasons for this association are not clear. In this review, the authors examine four potential hypotheses for why substance use and bipolar disorders co-occur: (a) substance abuse occurs as a symptom of bipolar disorder; (b) substance abuse is an attempt by bipolar patients to self-medicate symptoms; (c) substance abuse causes bipolar disorder; and (d) substance use and bipolar disorders share a common risk factor. None of these four hypotheses have unequivocal support for explaining all cases of bipolar and substance use disorder co-occurrence, and it is probable that all four mechanisms play some role in the excess of substance abuse observed in bipolar patients. Additional studies are warranted to clarify the complex relationships between these two conditions as better understanding of this co occurrence could lead to better treatment for patients afflicted with both disorders. PMID- 10721498 TI - Substance use disorders in schizophrenia: review, integration, and a proposed model. AB - Substance use disorders occur in approximately 40 to 50% of individuals with schizophrenia. Clinically, substance use disorders are associated with a variety of negative outcomes in schizophrenia, including incarceration, homelessness, violence, and suicide. An understanding of the reasons for such high rates of substance use disorders may yield insights into the treatment of this comorbidity in schizophrenia. This review summarizes methodological and conceptual issues concerning the study of substance use disorders in schizophrenia and provides a review of the prevalence of this co-occurrence. Prevailing theories regarding the co-occurrence of schizophrenia and substance use disorders are reviewed. Little empirical support is found for models suggesting that schizophrenic symptoms lead to substance use (self-medication), that substance use leads to schizophrenia, or that there is a genetic relationship between schizophrenia and substance use. An integrative affect-regulation model incorporating individual differences in traits and responses to stress is proposed for future study. PMID- 10721499 TI - Borderline personality disorder and substance use disorders: a review and integration. AB - The empirical literature on the comorbidity between borderline personality disorder (BPD) and substance use disorders (SUDs) is reviewed. BPD-SUD comorbidity data obtained from studies published from 1987 to 1997 document the frequent co-occurrence of these diagnoses. Methodological issues and theoretical models for understanding this co-occurrence are discussed. Finally, we present our conceptualization of the relations and interactions of the major factors influencing the development of BPD and contributing to the comorbidity between BPD and SUDs. PMID- 10721500 TI - Antisocial behavior and alcoholism: a behavioral genetic perspective on comorbidity. AB - Similar to many domains in the psychopathology literature, overlap and covariation between antisocial behavior (ASB) and alcohol dependence (AD) are oft documented but little understood. Although the relation between ASB and AD is reliably found and of substantial magnitude, it is not possible given the extant research to discriminate among alternative causal models that could give rise to this relation (e.g., ASB-->AD, AD-->ASB, reciprocal causation between ASB and AD, common causes of ASB and AD). In our opinion, true comorbidity among disorders can only be demonstrated and understood in the context of considerable knowledge regarding the disorders' underlying causes (viz., pathology and etiology). In this article, we present a number of behavior genetic models that may be useful for illuminating the causes of comorbidity among two or more disorders, as well as for understanding the etiology of each disorder individually. Using these behavior genetic approaches, psychopathology researchers can directly test alternative models for the comorbidity among disorders, as well as estimate the magnitude of different etiological factors (i.e., genetic and environmental influences) on comorbidity. Although not a panacea and somewhat demanding technically, behavior genetic approaches can shed new light on the comorbidity among disorders. PMID- 10721501 TI - Molden: a pre- and post-processing program for molecular and electronic structures. AB - Molden is a software package for pre- and postprocessing of computational chemistry program data. Interfacing to the ab initio programs Games-US/UK and Gaussian and to the semi-empirical package MOPAC is provided. The emphasis is on computation and visualization of electronic and molecular properties but, e.g., reaction pathways can be simulated as well. Some molecular properties of interest are processed directly from the output of the computational chemistry programs, others are calculated in MOLDEN before display. The package features different options to display MOLecular electronic DENsity, each focusing on a different structural aspect: molecular orbitals, electron density, molecular minus atomic density and the Laplacian of the electron density. To display difference density, either the spherically averaged atomic density or the oriented ground state atomic density can be used for a number of standard basis sets. The quantum mechanical electrostatic potential or a distributed multiple expansion derived electrostatic potential can be calculated and atomic charges can be fitted to these potentials calculated on Connolly surface(s). Reaction pathways and molecular vibrations can be visualized. Input structures can be generated with a Z-matrix editor. A variety of graphics languages is supported: XWindows, postscript, VRML and Povray format. PMID- 10721502 TI - Conformational preferences of the potent dopamine reuptake blocker BTCP and its analogs and their incorporation into a pharmacophore model. AB - Molecular mechanics calculations using MM3-92 and ab initio quantum mechanical calculations using SPARTAN 5.0 were performed on the structurally similar PCP and BTCP, in which only the latter has a cocaine-like pharmacological profile as a dopamine reuptake blocker. Calculations were also performed on BTCP analogs with a methyl group in various positions of the cyclohexane ring. The results for the cis-2-methyl compound, which retains good pharmacological activity, allowed us to determine that an aryl-axial conformer is the biologically active form for at least some of the compounds in this series. However, an aryl-equatorial conformer presents the identical pharmacophore, as shown by superposition of the two conformers. X-ray crystallographic structures were also obtained for BTCP and related compounds with a 2-methyl group on the cyclohexane ring, with reasonable agreement between the computational and experimental results. Superposition studies were performed with two rigid analogs of cocaine which illustrate the optimal orientations of the ammonium hydrogen for monoamine transporters. There is excellent agreement between a 'back-bridged' cocaine analog that is optimal as a dopamine reuptake blocker and the previously proposed biologically active conformer of methylphenidate. However, BTCP is found to be a better fit to the 'front-bridged' cocaine analog that is optimal for a serotonin reuptake blocker. PMID- 10721504 TI - Design of dimerization inhibitors of HIV-1 aspartic proteinase: a computer-based combinatorial approach. AB - Inhibition of dimerization to the active form of the HIV-1 aspartic proteinase (HIV-1 PR) may be a way to decrease the probability of escape mutations for this viral protein. The Multiple Copy Simultaneous Search (MCSS) methodology was used to generate functionality maps for the dimerization interface of HIV-1 PR. The positions of the MCSS minima of 19 organic fragments, once postprocessed to take into account solvation effects, are in good agreement with experimental data on peptides that bind to the interface. The MCSS minima combined with an approach for computational combinatorial ligand design yielded a set of modified HIV-1 PR C-terminal peptides that are similar to known nanomolar inhibitors of HIV-1 PR dimerization. A number of N-substituted 2,5-diketopiperazines are predicted to be potential dimerization inhibitors of HIV-1 PR. PMID- 10721503 TI - Automated docking and molecular dynamics simulations of nimesulide in the cyclooxygenase active site of human prostaglandin-endoperoxide synthase-2 (COX 2). AB - Molecular models of the complex between the selective COX-2 inhibitor nimesulide and the cyclooxygenase active site of human prostaglandin-endoperoxide synthase-2 have been built using a combination of homology modelling, conformational searching and automated docking techniques. The stability of the resulting complexes has been assessed by molecular dynamics simulations and interaction energy decomposition. It is found that nimesulide exploits the extra space made available by the replacement at position 523 of an isoleucine residue in COX-1 by a valine in COX-2 and establishes electrostatic interactions with both Arg-106 and Arg-499 (Arg-120 and Arg-513 in PGHS-1 numbering). Two alternate binding modes are proposed which are compatible with the pharmacological profile of this agent as a COX-2 selective inhibitor. PMID- 10721505 TI - Homology model directed alignment selection for comparative molecular field analysis: application to photosystem II inhibitors. AB - The use of a computational docking protocol in conjunction with a protein homology model to derive molecular alignments for Comparative Molecular Field Analysis (CoMFA) was examined. In particular, the DOCK program and a model of the herbicidal target site, photosystem II (PSII), was used to derive alignments for two PSII inhibitor training sets, a set of benzo- and napthoquinones and a set of butenanilides. The protein design software in the QUANTA molecular modeling package was used to develop a homology model of spinach PSII based on the reported amino acid sequence and the X-ray crystal structure of the purple bacterium reaction center. The model is very similar to other reported PSII protein homology models. DOCK was then used to derive alignments for CoMFA modeling by docking the inhibitors in the PSII binding pocket. The molecular alignments produced from docking yielded highly predictive CoMFA models. As a comparison, the more traditional atom-atom alignments of the same two training sets failed to produce predictive CoMFA models. The general utilities of this application for homology model refinement and as an alternative scoring method are discussed. PMID- 10721506 TI - Morphological similarity: a 3D molecular similarity method correlated with protein-ligand recognition. AB - Recognition of small molecules by proteins depends on three-dimensional molecular surface complementarity. However, the dominant techniques for analyzing the similarity of small molecules are based on two-dimensional chemical structure, with such techniques often outperforming three-dimensional techniques in side-by side comparisons of correlation to biological activity. This paper introduces a new molecular similarity method, termed morphological similarity (MS), that addresses the apparent paradox. Two sets of molecule pairs are identified from a set of ligands whose protein-bound states are known crystallographically. Pairs that bind the same protein sites form the first set, and pairs that bind different sites from the second. MS is shown to separate the two sets significantly better than a benchmark 2D similarity technique. Further, MS agrees with crystallographic observation of bound ligand states, independent of information about bound states. MS is efficient to compute and can be practically applied to large libraries of compounds. PMID- 10721507 TI - Validity of diagnostic ultrasound as a measure of delayed onset muscle soreness. AB - STUDY DESIGN: Repeated measures were taken to evaluate delayed onset muscle soreness (DOMS) following eccentric bicep contractions of the nondominant arm at 140% of 1 repetition maximum (RM) while the dominant arm served as control. OBJECTIVES: To explore the usefulness of a noninvasive method to assess delayed onset muscle soreness. BACKGROUND: Although many methods have been proposed to assess DOMS, most are somewhat subjective or require a blood sample. This study compared the assessment of DOMS following eccentric exercise using common assessment techniques with diagnostic ultrasound (US). METHODS AND MEASURES: Forty nonimpaired women (18-40 years) used a Cybex isotonic biceps curl machine to eccentrically lower, using their nondominant arm, 140% of their dominant arm 1 RM to induce muscle soreness. Four assessment methods, (1) goniometry assessing spontaneous muscle shortening (SMS); (2) subjective muscle soreness ratings (MSRs); (3) serum creatine kinase (CK); and (4) diagnostic US scans of muscle cross-sectional area (CSA), were conducted at 5 different assessment times: (1) pre-eccentric exercise; (2) postexercise; (3) 24 hours postexercise; (4) 48 hours postexercise; and (5) 72 hours postexercise. RESULTS: Significant differences existed across assessment times for 3 of the 4 assessment techniques, CK, SMS, and MSR. CONCLUSIONS: Previously published methodologies used to assess DOMS (CK, SMS, and MSR) were able to provide consistent and expected results relative to the onset and progression of soreness with a high degree of relatedness (r = 0.48 0.84). However, it appeared that the ability to assess muscle soreness by diagnostic US, as evidenced by intramuscular swelling, was limited. Thus, the technique was not sensitive enough to detect any statistically significant changes in muscle CSA. PMID- 10721509 TI - Diagnosis of patellofemoral pain after arthroscopic meniscectomy. PMID- 10721508 TI - Comparison of supervised exercise with and without manual physical therapy for patients with shoulder impingement syndrome. AB - STUDY DESIGN: A prospective randomized clinical trial. OBJECTIVE: To compare the effectiveness of 2 physical therapy treatment approaches for impingement syndrome of the shoulder. BACKGROUND: Manual physical therapy combined with exercise is a commonly applied but currently unproven clinical treatment for impingement syndrome of the shoulder. METHODS AND MEASURES: Thirty men and 22 women (age 43 years +/- 9.1) diagnosed with shoulder impingement syndrome were randomly assigned to 1 of 2 treatment groups. The exercise group performed supervised flexibility and strengthening exercises. The manual therapy group performed the same program and received manual physical therapy treatment. Both groups received the selected intervention 6 times over a 3-week period. The testers, who were blinded to group assignment, measured strength, pain, and function before treatment and after 6 physical therapy visits. Strength was a composite score of isometric strength tests for internal rotation, external rotation, and abduction. Pain was a composite score of visual analog scale measures during resisted break tests, active abduction, and functional activities. Function was measured with a functional assessment questionnaire. The visual analog scale used to measure pain with functional activities and the functional assessment questionnaire were also measured 2 months after the initiation of treatment. RESULTS: Subjects in both groups experienced significant decreases in pain and increases in function, but there was significantly more improvement in the manual therapy group compared to the exercise group. For example, pain in the manual therapy group was reduced from a pretreatment mean (+/- SD) of 575.8 (+/- 220.0) to a posttreatment mean of 174.4 (+/- 183.1). In contrast, pain in the exercise group was reduced from a pretreatment mean of 557.1 (+/- 237.2) to a posttreatment mean of 360.6 (+/- 272.3). Strength in the manual therapy group improved significantly while strength in the exercise group did not. CONCLUSION: Manual physical therapy applied by experienced physical therapists combined with supervised exercise in a brief clinical trial is better than exercise alone for increasing strength, decreasing pain, and improving function in patients with shoulder impingement syndrome. PMID- 10721510 TI - Gross measures of exercise-induced muscular hypertrophy. AB - STUDY DESIGN: Pretest-posttest, single factor design. OBJECTIVES: To compare several indices that might be used to depict muscle size. BACKGROUND: The particular strategy used during heavy-resistance training may determine the magnitude of hypertrophic adaptations. At the same time, assorted measures supposedly reflecting muscle size may provide different results. METHODS AND MEASURES: Four groups of men (n = 38, mean age = 21.1 years, SD = 2.1) were exposed to conditions designed to elicit differential hypertrophic adaptations following 21 sessions of squat training. Three of the groups performed 4 sets of multiple repetitions maximum (RM): group I, 3-5 RM; group II, 13-15 RM; and group III, 23-25 RM. A control (C) group did no formal physical training. Tests used to represent muscle size included body weight, thigh girth, net thigh girth, and quadriceps femoris and hamstring thicknesses via B-mode ultrasound. RESULTS: Changes in the groups subsequent to training were similar for body weight and hamstring thickness. Results differed for the remaining 3 dependent variables (mean +/- SD): thigh girth was greater in groups II (1.42 +/- 1.00) and III (1.35 +/- 1.16) than in group C (0.24 +/- 0.69); net thigh girth was greater in groups II (1.33 +/- 0.77) and III (1.40 +/- 1.03) than in group C (0.10 +/- 0.84); and quadriceps femoris thickness was greater in all 3 training groups (I, 0.61 +/- 0.54; II, 0.43 +/- 0.30; III, 0.55 +/- 0.39) than in group C (0.05 +/- 0.11). CONCLUSIONS: Observed muscle mass change following heavy-resistance training is dependent upon both the training intervention and tool used for measurement. PMID- 10721511 TI - Criterion-related validity of the figure-of-eight method of measuring ankle edema. AB - STUDY DESIGN: Single-session, prospective, repeated-measures design. OBJECTIVE: To determine intratester reliability and criterion-related validity of the figure of-eight method of measuring ankle edema. BACKGROUND: The measurement of edema is often necessary when physical therapists assess patients with ankle injuries. The figure-of-eight method has been shown to be a reliable method in measuring the size of ankles in subjects without edema but not in subjects with ankle swelling. The validity of this method of measurement has not been established. METHODS AND MEASURES: The subjects (mean age, 22.7 +/- 4.4 years) were 7 men and 8 women with observable ankle edema secondary to acute or chronic ankle sprains or injury to the lower extremity. Three figure-of-eight measurements were taken by one tester. These measurements were correlated to measurements taken by another tester using a foot volumeter. RESULTS: The intraclass correlation coefficient (ICC [2,1]) for the figure-of-eight measurements was 0.99. The Pearson Product Moment correlation coefficients between the figure-of-eight measurements and the first volumetric measurement were 0.89 (first figure-of-eight), 0.88 (second figure-of-eight), 0.91 (third figure-of-eight), and 0.90 (mean of all 3 figure-of-eight measurements). CONCLUSION: The figure-of-eight method was demonstrated to be a reliable and valid indirect method of measuring ankle edema in individuals with edema secondary to sprains or other lower-extremity musculoskeletal disorders. PMID- 10721512 TI - More on evidence-based practice. PMID- 10721513 TI - Society of Cardiovascular and Interventional Radiology (SCVIR) 25th annual scientific meeting. San Diego, California, USA. March 25-30, 2000. Abstracts. PMID- 10721514 TI - Cryptococcal meningitis in the immunocompromised host: intracranial hypertension and other complications. AB - Cryptococcosis as a complication of the immunocompromised host has dramatically increased in frequency since the start of the AIDS epidemic. This trend has heightened awareness of the complications of cryptococcal meningitis; of these, intracranial hypertension is common, severe, and life-threatening, as exemplified by three cases in our institutions presented here in detail. An aggressive approach to management of this complication has not been the standard of care, but neurosurgical interventional studies combined with physiologic observations suggest early intervention may reduce the devastating morbidity and mortality. PMID- 10721515 TI - Isolation of a new potent cytotoxic pigment along with indigotin from the pathogenic basidiomycetous fungus Schizophyllum commune. AB - An indole derivative, schizocommunin, was isolated along with indigotin (indigo), indirubin, isatin, and tryptanthrin, from the liquid culture medium in which a culture of Schizophyllum commune, isolated from the bronchus of a human patient with allergic bronchopulmonary mycosis, had been grown. The structure of schizocommunin was established by spectroscopic investigation. Schizocommunin showed the strong cytotoxicity against murine lymphoma cells. The assignments of the 1H- and 13C-NMR signals of indigotin were also listed. PMID- 10721516 TI - Production of proteinase and phospholipase by Paracoccidioides brasiliensis. AB - We have investigated the production of proteinase and phospholipase by 20 different isolates of Paracoccidioides brasiliensis. Isolates were grown in Bacto peptone, Dextrose, pH 5.5, agar slants, at 27 degrees C for 30 days, and cultures were transferred onto Petri dishes containing basis medium and bovine serum albumin fraction V and sterile egg yolk as substrates for enzyme production, and incubated at 27 degrees C. After 30 days net enzyme activity was visualized and quantitatively evaluated, measuring a ratio between colony diameter and diameter of the transparent (proteinase) or white (phospholipase) ring zone surrounding it. Results demonstrated that all isolates had the ability to produce proteinase and phospholipase, even though variability in enzyme production was noted among different isolates of P. brasiliensis. PMID- 10721517 TI - Aspergillus and other moulds in the air of Kuwait. AB - A one-year survey was carried out to study the aerial prevalence of Aspergillus species and other moulds in the outdoor and indoor environments of Kuwait. Petri plates containing rose-Bengal medium were exposed for 20 minutes twice a month using a six-stage Andersen air sampler at the pre-determined sites. The exposed plates were incubated at 28 degrees C +/- 1 degree C up to 5 days and colonies were enumerated and identified by colonial and microscopic morphology. The data revealed that Aspergillus species were the predominant component (27.7%) of the outdoor aerospora of Kuwait and A. fumigatus alone accounted for 21.3% of the total aspergilli. In contrast, Cladosporium species formed the major component of the indoor aerospora (22.8%), followed by Aspergillus species (20.9%), Penicillium species (14.6%), and Bipolaris species (10.6%). A comparison of the fungi recorded in the outdoor and in the indoor air revealed that Aspergillus, Alternaria and Fusarium were significantly higher in the outdoor environment, whereas Cladosporium, Penicillium, and Bipolaris were significantly higher in the indoor environment. The relative prevalence of Aspergillus species and other moulds in the outdoor and indoor air of Kuwait was as follows: A. fumigatus 5.9 and 9.8%, A. flavus 4.9 and 3.9%, other aspergilli 16.8 and 7.0%, Alternaria species 19.8 and 7.9%, Cladosporium species 13.7 and 22.8%, Penicillium species 7.6 and 14.6%, and other moulds 31.2 and 34.1%, respectively. During the study, 25 different genera were identified, indicating a wide diversity in the spectrum of local fungal aerospora. The study provides useful information on the prevalence of allergenic fungi in the outdoor and indoor environments of Kuwait. PMID- 10721518 TI - Dependence of the entomopathogenic fungus, Beauveria bassiana, on high humidity for infection of Rhodnius prolixus. AB - The impact of relative humidity (RH) on the infective potential of the isolate Bb INRA 297 of Beauveria bassiana (Bals.) Vuillemin (Deuteromycotina: Hyphomycetes) against first instar nymphs of Rhodnius prolixus Stal. (Hemiptera: Reduviidae) was determined. Fungus-treated insects were exposed to RHs ranged from 75 to 100% at 25 degrees C. Results clearly showed a threshold of humidity at ca. 96% for high and rapid mortality. After initial exposure to increasing periods of 97% (4, 8, 16, 24, 36 and 48 h) and subsequent transfer to constant lower RHs (43, 53, 75 and 86%) at a constant 25 degrees C, an incubation of at least 48 h at 97% RH was necessary to kill all insects. On changing RHs of 97/75% and different regimes of temperature (15/28 degrees C, 20/25 degrees C, 25/28 degrees C, and 25/35 degrees C), at least 72 h of initial exposure at 97% RH for the 15/28 degrees C regime, 48 h for the 20 degrees/25 degrees C and 25/28 degrees C regimes and 36 h for 25/35 degrees C were needed to kill all insects over a 6-day incubation time. Delayed exposure to favorable moisture condition (97% RH), significantly affected infection for up to a 3-day delay within the various temperature-humidity regimes tested. PMID- 10721519 TI - Environment factors influence in vitro interspecific interactions between A. ochraceus and other maize spoilage fungi, growth and ochratoxin production. AB - The effect of water availability (water activity, aw; 0.995-0.90) and temperature (18-30 degrees C) on in vitro interactions between an ochratoxin producing strain of Aspergillus ochraceus and six other spoilage fungi was assessed in dual culture experiments on a maize meal-based agar medium. In primary resource capture of nutrient substrate, A. ochraceus was dominant against many of the interacting species, being able to overgrow and replace A. candidus, and sometimes A. flavus and the Eurotium spp. regardless of aw or temperature. However, with freely available water (0.995 aw) A. alternata and A. niger were dominant, with mutual antagonism between A. ochraceus and A. flavus at 25-30 degrees C. In the driest conditions tested (0.90 aw) there was also mutual antagonism between A. ochraceus and the two Eurotium spp. Overall, under all conditions tested the Index of Dominance for A. ochraceus was much higher than for other competing species combined suggesting that A. ochraceus was a good competitive colonist able to replace a number of other species. However, the growth rate of A. ochraceus was modified and decreased by the interaction with competitors. Interaction between A. ochraceus and species such as A. alternata (18 degrees C/0.995) and Eurotium spp. (0.995-0.95 and 25-30 degrees C) resulted in a significant stimulation of ochratoxin production. The results are discussed in relation to the effect that environmental factors have on the possible competitiveness of A. ochraceus in the maize grain ecosystem and the role of ochratoxin in niche exclusion of competitors. PMID- 10721520 TI - [Surgical treatment of spinal perimedullary AVF/AVM]. PMID- 10721521 TI - [Clinical role and possibility of magnetoencephalography: functional approach to intracranial lesions]. PMID- 10721522 TI - [Usefulness of Doppler echocardiography in patients with ischemic cerebrovascular disease]. AB - We carried out transthoracic Doppler echocardiography on 213 patients with ischemic cerebrovascular disease to examine cardiac functions. The prevalence of valvular regurgitation, left ventricular diastolic function (E/A ratio), and ejection fraction (EF) of each patient were studied by transthoracic Doppler echocardiography. In patients with embolic strokes, the prevalence of valvular regurgitation of the left ventricular system such as mitral regurgitation was significantly higher than that of normal controls. In contrast, there was no significant difference in the prevalence of valvular regurgitation in the right ventricular system. Although there was no significant difference in the ejection fraction between groups, the E/A ratio in patients with ischemic cerebrovascular disease showed a significantly lower value than that in normal controls. Patients with ischemic cerebrovascular disease showed a tendency to have an atherosclerotic change in the systemic arteries including cerebral and coronary arteries, reflecting an aggravation of cardiac wall compliance and diastolic dysfunction resulting in lowering of the E/A ratio. Thus, transthoracic Doppler echocardiography was useful to evaluate cardiac diastolic dysfunction and valvular regurgitation in patients with ischemic cerebrovascular disease. PMID- 10721523 TI - [The three-dimensional CT angiography findings of ruptured aneurysms hardly detectable by repeated cerebral angiography]. AB - We reported three cases of cerebral aneurysms hardly detectable by cerebral angiography, but easily detected by three-dimensional CT angiography (3D-CTA). These cases were ruptured aneurysms with subarachnoid hemorrhage. After detection of subarachnoid hemorrhage on CT scan, cerebral angiography was performed at first, but aneurysms were not detected. Subsequently 3D-CTA was carried out, and aneurysms were detected. In all cases, cerebral angiography was repeated, after the aneurysms had been found by 3D-CTA. This time aneurysms were all detected by cerebral angiography, but each case needed photographs from special direction. The aneurysms were small by usual cerebral angiography and they were almost invisible behind the artery near which they existed. 3D-CTA was very useful for detection of small aneurysms, but small perforating arteries around the aneurysms were invisible by 3D-CTA. To find these perforating arteries, cerebral angiography was needed. PMID- 10721524 TI - [A case of the syndrome of the sinking skin flap: case report]. AB - A case of "the syndrome of the sinking skin flap" was presented. A 40-year-old man had suffered from severe SAH 9 months before. An aneurysm of the anterior communicating artery was successfully clipped and the bone flap was removed for the purpose of the external decompression. Cranioplasty and V-P shunt were performed 1 month after SAH, but both were removed because of postoperative wound infection, viz. epidural and subdural abscess 4 months after SAH. Following this, L-P shunt was performed, and the patient was discharged with mild dementia. A concave deformity of the skin flap developed about 4 months after the L-P shunt. Neurological examination showed progressive left hemiparesis and akinetic mutism. A low CSF pressure was demonstrated, but RI cisternography revealed normal CSF circulation. Intrathecal infusion of the artificial CSF was carried out via lumbar puncture and concavity of the skin flap gradually improved. This procedure resulted in improvement of the neurological deficits. Cranioplasty with artificial bone was performed under continuous intrathecal infusion of the artificial CSF. Postoperative course was satisfactory and neurological examination revealed only mild dementia. The pathological mechanism in our case was probably due to the compression of the brain by the atmospheric pressure following the external decompression. Moreover, L-P shunt exaggerated this pathology by the overdrainage of CSF. PMID- 10721525 TI - [A case of isolated fourth ventricle in the early postoperative period of subarachnoid hemorrhage]. AB - An adult case of isolated fourth ventricle which developed in the early postoperative period of SAH is reported. A 72-year-old male with Hunt & Kosnik Grade 4 subarachnoid hemorrhage (SAH) underwent emergent neck-clipping of an anterior communicating artery aneurysm along with setting of external ventricular drainage (EVD) and cisternal drainage (CD). The lamina terminalis (LT) was opened. Preoperative study had showed diffuse SAH with intraventricular hemorrhage (IVH), and mild dilatation of the whole ventricular system on CT scan (Fisher Group 4). Twelve hours after surgery, both of the drainages were opened with the pressure setting of 20 cm H2O for EVD and 10 cm H2O for CD. Although his neurological state had been improving, 2 hours after the opening of the drainages he suddenly fell into respiratory arrest and coma, when 20 ml of CFS through CD was drained. On CT scan, isolated fourth ventricle was recognized. The patient died on the ninth postoperative day. In case of severe SAH with diffuse IVH, we should be careful when setting the pressure of EVD or CD with the LT opened, because of the possibility of occlusion of the fourth ventricle outlet and aqueduct, that results in fourth ventricle isolation. PMID- 10721526 TI - [Postoperative hemorrhage due to normal pressure hyperperfusion breakthrough after a trapping of VA-PICA dissecting aneurysm]. AB - We described a case of cerebellar hemorrhage after trapping of a vertebral artery dissecting aneurysm. A forty-eight-year-old man had suffered from severe headache, vomiting and disturbance of consciousness. He was transferred to our hospital in an ambulance. Emergency CT scan showed subarachnoid hemorrhage in the posterior fossa predominantly, intraventricular hemorrhage and hydrocephalus change. Chest X-ray showed radiological evidence of pulmonary edema. The initial blood-gas determinations demonstrated a marked reduction in PaO2 and increased PaCO2. Five days after admission, the patient's condition was improving. Cerebral angiography was performed using the Seldinger method. It revealed a right vertebral artery dissecting aneurysm just distal to the posterior inferior cerebellar artery. We performed an operation to trap the VA dissecting aneurysm. Blood pressure was well controlled under 140 mmHg during the operation and he recovered from anesthesia completely. On the day after the operation, suddenly the patient's consciousness began to deteriorate. Emergency CT scan was performed and it showed SAH, cerebellar hemorrhage and diffuse swelling of the cerebellum on the same side as the operation. We suspected rebleeding of the aneurysm due to a clip's having slipped. Reoperation was performed, but the clip was not displaced and there were no definite bleeding vessels on the operative field. Consequently only external decompression and resection of the right cerebellum were performed. We discuss pathogenesis of the occurrence of hemorrhage in this particular case after trapping. We also review the relevant literature. PMID- 10721527 TI - [A case of intrasellar pure germinoma]. AB - A 19-year-old male presented with progressive loss of vision and diabetes insipidus due to an intra- and suprasellar tumor. Transsphenoidal exploration revealed a pure germinoma. Seven days after the operation, bleeding from the residual tumor caused headache and right occulomotor palsy. The residual tumor and hematoma were removed using pterional approach. The residual tumor disappeared after postoperative irradiation. After 5 months of radiation, multiple lesions due to dissemination of the germinoma were discovered in the suprasellar region. Adjuvant chemotherapy and whole supratentorial irradiation were performed. All lesions regressed completely. Mid-sagittal magnetic resonance image was useful in our patient for differential diagnosis between intrasellar germinoma and pituitary adenoma. Before initiating an operation for intrasellar germinoma, awareness is needed for the fact of postoperative bleeding. We notice that transsphenoidal surgery should be selected for treatment of postoperative bleeding from intrasellar germinoma. PMID- 10721528 TI - [An arterial dissection of the distal anterior inferior cerebellar artery treated by endovascular therapy]. AB - A rare case of a dissection of the distal anterior inferior cerebellar artery (AICA) is presented. A 68-year-old woman with sudden onset of headache was admitted. Computed tomographic (CT) scan demonstrated no subarachnoid hemorrhage. Seven days later, CT scan revealed subarachnoid and intraventricular hemorrhage. Left vertebral angiogram showed an aneurysmal dilatation on the distal AICA with a diagnosis of suspected arterial dissection. At that time, we chose delayed craniotomy to observe the lesion directly. However, rebleeding causing aggravation of the patient's systemic condition delayed radical treatment. 19 days after rebleeding, by a superselective angiogram, endovascular treatment was selected to prevent further bleeding. The lesion was diagnosed as dissection of the AICA. The parent AICA was occluded with a Guglielmi detachable coil and fibered platinum coils. Dissection of the distal portion of the cerebellar artery is rare. Only six cases have been reported in the posterior inferior cerebellar artery (PICA) and two cases in the superior cerebellar artery (SCA). However, to our knowledge, no such case has been reported in the AICA. Ruptured dissection of distal PICA or SCA is reported to require early treatment to prevent further bleeding. Ruptured dissection of the distal AICA also requires early treatment. Two cases of distal dissection of PICA and SCA successfully treated by endovascular treatment are reported. Endovascular treatment has some benefits in that it does not always require general anesthesia and in that it can follow diagnostic angiography. On the other hand, saccular aneurysms of the distal AICA do not always require early treatment, such as removal of hematoma because of low incidence of vasospasm. So, to decide the treatment, precise diagnosis of the dissection is very important. PMID- 10721529 TI - [Brain abscess following thalamic hemorrhage: a case report]. AB - Brain abscess following stroke is rare. We present a case of brain abscess which developed after hypertensive thalamic hemorrhage. A 51-year-old man had a left thalamic hemorrhage. Three months later, the patient was admitted to our hospital due to deterioration of right hemiparesis and motor aphasia. On admission, CT scan showed a ring-like high density area with peripheral edema in the left thalamus. T1-weighted magnetic resonance (MR) images revealed a large ring enhanced lesion with surrounding edema in the same region. Laboratory examination demonstrated no signs of infectious disease. The patient's neurological state rapidly deteriorated 10 days after admission. Yellowish pus was aspirated during the emergent surgery. Bacteriological study of the pus revealed Staphylococcus Aureus. His symptoms improved after surgery followed by antibiotics therapy. The correct diagnosis prior to surgery was difficult in the present case because of lack of any signs of infection. Our case indicates that possibility of brain abscess should be taken into consideration as a rare differential diagnosis following intracerebral hemorrhage. PMID- 10721531 TI - Evolutionary origins of human alcoholism in primate frugivory. AB - Evolutionary origins of alcohol consumption have rarely been considered in studies of ethanol addiction. However, the occurrence of ethanol in ripe and decaying fruit and the substantial heritability of alcoholism in humans suggest an important historical association between primate frugivory and alcohol consumption. Olfactory localization of ripe fruit via volatilized alcohols, the use of ethanol as an appetitive stimulant, and the consumption of fruits with substantial ethanol content potentially characterize all frugivorous primates, including hominoids and the lineage leading to modern humans. Patterns of alcohol use by humans in contemporary environments may thus reflect a maladaptive co option of ancestral nutritional strategies. Although diverse factors contribute to the expression of alcoholism as a clinical syndrome, historical selection for the consumption of ethanol in the course of frugivory can be viewed as a subtle yet pervasive evolutionary influence on modern humans. PMID- 10721530 TI - [Giant osteoma of the temporal bone with otitis media: a case study]. AB - A 71-year-old female presented with a giant osteoma of the right temporal bone and otitis media. Computed tomography (CT) and magnetic resonance imaging (MRI) confirmed the presence of the giant osteoma. An operation was performed to alleviate the difficult-to-treat otitis media and to address the related cosmetic problem. During the operation, three-dimensional CT (3D-CT) was very helpful in understanding the relationship between the tumor and the peripheral structures. Removal of the tumor improved the patient's otitis media. Osteoma of the temporal bone is rare. Only twenty-one cases of mastoid osteoma have been reported in Japan. In the present study, we review osteomas of the temporal bone and discuss their management. PMID- 10721532 TI - The evolution of ADHD: a disorder of communication? AB - Attention deficit hyperactivity disorder (ADHD) is the most commonly diagnosed psychiatric condition. Many believe that the central disability is impaired inhibition, which leads to reduced abilities in social skills, self-control, organization and time management. The behaviors identified by clinicians as problematic--inattention, hyperactivity and impulsivity--have been incorporated into several evolutionary models as selectively adaptive cognitive skills for surviving the challenges of a variable Pleistocene environment. We propose that the "disabilities" exhibited by individuals with ADHD are maladaptive, and we concur with Barkley that there is a central impairment in the behavioral inhibition system. The underlying neural anatomy and physiology support the possibility that neurotransmitter pathology may have an impact on other interlinked systems (including language), and may also account for the frequent comorbidity of aggression, anxiety, depression, and learning disabilities (many of which are language-related). Language skills compete with other cognitive activities for the attentional system, and thus the evolution of language could not in fact be independent of the evolution of attention. If language represents the ultimate expression of the attentional system, and some individuals with ADHD are seriously impaired in the coordination of interlinked neural systems (including language), then ADHD fits Jerome Wakefield's definition of "harmful dysfunction," and communication impairments should be investigated more thoroughly by clinicians. PMID- 10721533 TI - Forum on Respiratory Infections. February 2000. Abstracts. PMID- 10721534 TI - GPs take stock. Use it or lose it! PMID- 10721535 TI - Alcohol and other drug use as normal behaviour. PMID- 10721536 TI - Local register-reminder systems for immunisation in general practice. PMID- 10721537 TI - Liver function tests (LFTs) PMID- 10721538 TI - Acute sinusitis. Who should we be treating? AB - BACKGROUND: The diagnosis of acute sinusitis has been regarded as a serious condition that requires the use of antibiotics. However the increasing incidence of resistant organisms means antibiotics need to be used carefully. OBJECTIVE: To look at the evidence available regarding antibiotic use for sinusitis, and to discuss its application to general practice. DISCUSSION: There have been surprisingly few randomised double blind placebo controlled trials for sinusitis, and fewer still have been based in a representative population of primary care patients. This article discusses studies relevant to general practice. Several practical clinical symptoms and signs have been shown to increase the likelihood of a patient having acute bacterial sinusitis, and therefore benefit from antibiotics. When antibiotics are used, comparative data suggest that amoxycillin should be used first line. The issue of patient experience, expectations and satisfaction is also raised. PMID- 10721539 TI - The doctor's bag. What do you really need? AB - BACKGROUND: The doctor's bag should contain essential drugs to treat life threatening emergencies and other serious medical conditions. Practitioners may require these drugs for treating patients in their offices, on home visits and particularly in rural practice for emergency home and roadside calls. OBJECTIVE: To evaluate which medications and equipment need to be included in the doctor's bag. DISCUSSION: The selection of the most appropriate drugs provided by the Pharmaceutical Benefits Scheme (PBS) is discussed. Oral and inhaler preparations for certain specific conditions are also included. Storage and safe keeping of drugs is an important consideration. PMID- 10721540 TI - Safe sex messages for adolescents. Do they work? AB - BACKGROUND: Adolescence is a time of risk taking. Constructive risk taking aids the developmental task of becoming a mature, confident adult with a sense of mastery of self and the world. However, ill judged and misinformed risk taking in sexual behaviour can have serious and life long consequences. OBJECTIVE: To assess strategies used for promoting safe sex messages to adolescents and to delineate the role of the general practitioner in promoting these messages effectively. DISCUSSION: The most successful sexual health promotion strategies are those that acknowledge the social and media influences on young people and use these to help strengthen group norms around safe sexual behaviour. GPs have an important role to play in adopting a non judgmental approach to accepting young people as sexual beings and engaging in one to one opportunistic health promotion. PMID- 10721541 TI - Calf and ankle swelling. AB - BACKGROUND: Swelling of the calf and ankle region is a common presenting symptom and historical features such as speed of onset, trauma and mechanism of injury are important in aiding diagnosis. OBJECTIVE: To discuss diagnosis and management of musculoskeletal causes of calf and ankle swelling. DISCUSSION: Calf muscle injuries and injuries around the ankle including Achilles tendon injuries, ankle ligament injuries and overuse injuries are discussed. PMID- 10721542 TI - New insights into the genetic basis of intellectual disabilities. AB - BACKGROUND: Within general practice patients with intellectual disabilities are common. OBJECTIVE: This article sheds some new light on the underlying genetic mechanisms of intellectual disabilities. It outlines the genetic basis of known inherited and sporadic causes that have recently been understood through new molecular advances. DISCUSSION: General practice has an important role to play in pursuing a precise biomedical diagnosis by improving information provided to the patient and family regarding recurrence risks and prognoses as well as management issues specific to the underlying condition. New molecular techniques are providing us with specific diagnostic tools as well as improving our understanding of the complex mechanisms that contribute to what is known as human intelligence. PMID- 10721543 TI - Difficult doctor-patient relationships. AB - BACKGROUND: Although difficult consultations constitute about 15% of general practice work, patients considered 'difficult' by one doctor, may not be thought 'difficult' by another. Rather than labelling and therefore dismissing patients, it is more helpful to consider that the relationships may be difficult rather than the patient. OBJECTIVE: To change the perspective from labelling a patient as difficult to considering the difficulties in the relationship between doctor and patient. This empowers the doctor to use communication skills to develop appropriate strategies for change. DISCUSSION: This article describes a method for managing difficult doctor-patient relationships that involves acknowledging the problem, settling boundaries, using additional communication skills and, when necessary, bringing in external resources to assist both doctor and patient. PMID- 10721544 TI - Coxsackie B virus. The great pretender. PMID- 10721545 TI - Conditions responding to lasers. Hair removal & resurfacing lasers. PMID- 10721546 TI - Can I fly Doc? Eustachian tube dysfunction. PMID- 10721548 TI - Medicine. What has love got to do with it? PMID- 10721547 TI - Evidence based hypolipidaemic drug treatment. PMID- 10721549 TI - Chronic fatigue syndrome. PMID- 10721550 TI - General practitioners' beliefs, attitudes and reported actions towards chronic fatigue syndrome. AB - OBJECTIVE: To undertake a survey of Australian general practitioners (GPs) to explore their beliefs, attitudes and reported actions with respect to chronic fatigue syndrome (CFS). METHOD: A random sample of 2090 Australian GPs, stratified by state, was surveyed in May-August 1995. RESULTS: A 77% response rate was obtained. For the majority of practitioners who pursue a diagnosis of CFS, six symptoms were considered to be of significance: chronic unremitting fatigue for over 6 months; failure to recover energy after rest; reduced exercise tolerance; prostration for several days after exercise; generalised myalgia and poor concentration. Individual counselling was the most frequently used treatment. Thirty-one percent of practitioners reported that they did not believe that CFS is a distinct syndrome. Of these, 70% reported that the most likely cause of chronic fatigue was depression. CONCLUSION: There is considerable diversity of opinion between practitioners about CFS. The diversity extends from questioning whether the syndrome even exists to different strategies for diagnosis and management. PMID- 10721551 TI - General practitioners' views on patient care research. AB - BACKGROUND: Little research has been undertaken into the factors affecting recruitment by Australian general practitioners of patients for clinical trials. Understanding the differences between recruiters and non-recruiters will assist researchers in better supporting general practitioners involved in such research. METHOD: A survey of general practitioners involved in recruiting patients for clinical trials for the RACGP Research Program was undertaken. RESULTS: Recruiters were more likely to be interested in learning more about research, to perceive involvement as worthwhile, to desire a good relationship with Research Program staff and to feel the doctor-patient relationship assists recruitment. DISCUSSION: Recruiters in general are average general practitioners, male, middle aged and work in group practices. Most felt some discomfort in recruiting patients, but believed the strong doctor-patient relationship assisted in the process. CONCLUSION: The Research Program needs to recruit general practitioners interested in research, choose topics of interest, keep recruitment protocols simple and stay in contact. PMID- 10721552 TI - Update on the treatment of Wegener's granulomatosis. AB - Mortality due to WG has been significantly decreased by cytotoxic therapy with cyclophosphamide and glucocorticoids. Several studies have addressed different treatment regimens, particularly different maintenance regimens, in order to reduce the potential for cyclophosphamide-induced toxicity. Relapse may be precipitated by the chronic carrier-state of S aureus in the nasopharynx, and is sometimes heralded by rising c-ANCA titers. The treatment of the relapse is determined by the severity of its manifestations. Options for maintenance therapy include methotrexate and azathioprine. The value of therapy with TMSx for maintenance of remission still is uncertain, although its use 3 times a week is recommended for Pneumocystis prophylaxis. The ACR guidelines for monitoring drug toxicity should be followed when treating WG. PMID- 10721553 TI - [Out-sourcing clinical microbiology?]. PMID- 10721554 TI - [Intravenous drug users serologic control: what may be prevented?]. AB - OBJECTIVE: To report of IDU serologic control done in 1997 in comparison with another of 1990, in a Health-Care Area of Madrid, Spain, focussing to hepatitis B virus prevention. PATIENTS AND METHODS: Cross-sectional study. LOCATION: Fuenlabrada-Leganes, 400,000 inhabitants Health-Care Area in the Southeast of Madrid. PATIENTS: 233 IDU submitted for serological study during 1997 (mean age, 29.5; standard deviation, 6.8; 85.8% males). INTERVENTIONS: HIV, HBV, HCV and Treponema pallidum; serological markers were done at Severo Ochoa Hospital and a search for previous reports since 1991. Study of HBV previous vaccination was done with HBsAb marker in HBV seronegative patients. RESULTS: Prevalence: HBV 39.5% (IC = 33.2-46.1), HCV 49.3% (IC = 42.8-55.9), HIV 11.1% (IC = 7.5-16.1), syphilis 3.4% (IC = 1.5-7). Statistically meaning falls in rates were detected comparing them with results from 1990 (HBV: 67.5%; HCV: 75.1%; HIV: 50.1%; n = 261; p < 0.001). Only 5 (3.55%) out of 141 HBV seronegative patients showed HBsAb titres above 10 mU/I, although 55 (39%) of them had been tested for HBV previously. Nineteen (20.6%) out of 92 patients with HBV markers had previous seronegative reports and 3 of them suffered an acute viral infection. CONCLUSIONS: We have seen a statistically meaning fall in seroprevalence of the studied agents although a low HBV vaccination rate was achieved. PMID- 10721555 TI - [Comparative analysis of viral load by bDNA HCV RNA-2.0 and amplicor HCV monitor in patients with hepatitis C infection]. AB - BACKGROUND: Two standardized techniques, Quantiplex (bDNA-2.0) and Amplicor Monitor have been evaluated for the quantification of virus load of HCV with these objectives: a) determinate the relationship between virus load and genotype, and b) evaluate the virus load in serial serum samples and in patients with normal or slightly increased liver enzymes in an area with a high prevalence of genotype 1. RESULTS: A significant correlation of 0.7 (p < 0.0001) in virus load has been observed by both methods, but the virus load is smaller by Monitor than by Quantiplex and does not depend on genotype. The relationship Monitor/Quantiplex is smaller in patients with non-1 genotype than in patients with genotype 1a (p = 0.01) and 1b (p = 0.005). Virus characteristics are similar in patients with normal or slightly increased enzymes than in patients with high enzymes. Virus load by both methods is not related to the age, sex, know duration of the infection, transmission manner of the infection neither to the histologic activity index. CONCLUSION: The virus load not depends on genotype. The determination of virus load in a single serum sample adequately reflects the virus load are in several serum samples in patients with chronic HCV infection. The genotype and the virus load are similar in patients with normal enzymes than in patients with high enzymes. PMID- 10721556 TI - [Confirmation by molecular typing of cross-contamination in a mycobacteria laboratory]. AB - BACKGROUND: Detection of cross-contamination in the laboratory by restriction fragments length polymorphism (RFLP) analysis of Mycobacterium tuberculosis strains. MATERIAL AND METHODS: 224 strains isolated during a five years period were characterized by IS6110 fingerprinting performed (RFLP) by standardized protocols. RESULTS: Four groups of isolates with smear-negative specimens and low number of colony form its units were detected. They were processed in the same batch and day than other smear-positive specimens with identical RFLP patterns. Fifteen patients were involved and the review of four patients' charts showed that they did not have the typical manifestations of tuberculosis. CONCLUSIONS: When M. tuberculosis isolates were obtained from smear-negative specimens, the results of specimens processed in the same batch and the patients' charts should be reviewed. If with these data the possibility of cross-contamination is suspected, the isolates must be analyzed by molecular typing methods. PMID- 10721557 TI - [Fulminant streptococcus infection of soft tissue]. AB - BACKGROUND: In recent years there has been an apparent increase in severe infections produced by group A beta-hemolytic Streptococci in developed countries. Necrotizing fascitis and myositis are two rare but fearsome complications caused by this microorganism. METHODS: Two cases of fulminant soft tissue infection recently observed in the authors' center are presented and the clinical presentation and differential diagnosis commented upon. RESULTS: The first case was a necrotizing fascitis at a surgical wound which appeared following a gynecological surgery. The second case reports gluteal myositis following intramuscular injection. In both cases the evolution was disastrous. CONCLUSIONS: Streptococcus pyogenes may produce fulminant infections in patients without underlying disease either spontaneously or following minimum traumas. The most frequent involvement is of the soft tissues. This virulence at a local tissue and systemic level has been associated with the production of exotoxin. PMID- 10721558 TI - [Cutaneous myiasis by Sarcophaga sp]. AB - BACKGROUND: Myasis are due to the invasion of tissue or cavities of animal organism by dipterous larvae. PATIENTS AND METHODS: Two cases of semi-specific myasis caused by Sarcophaga larvae are described. RESULTS: Case 1: A 77-year-old woman with Kaposi sarcoma in the left leg developed cutaneous radionecrosis secondary to radiotherapy. In June, 1998 five fly larvae were observed moving freeing within the wound. These were removed with forceps and local dressing of the wound was performed with povidone-iodine. Case 2: A 87-year-old man with moderate dementia, progressive immobilization syndrome prostasic neoplasm and gastric ulcer reported. In the posterior part of the right outer ear the presence of three fly larvae were observed with some dermic orifices made. The larvae were removed with forceps and local dressing was carried out with povidone-iodine. In the laboratory an adult form was obtained from one of the larva. CONCLUSIONS: Myasis in infrequent in Spain and appears particularly in people with some predisposing factor. Treatment consists in the elimination of the larvae in the infected tissue and disinfection of the wound. PMID- 10721559 TI - [The activity of eight fluoroquinolones versus biolayers of Escherichia coli and Pseudomonas aeruginosa in siliconized latex urinary catheters]. AB - OBJECTIVE: To evaluate the in vitro activity of eight fluoroquinolones against Escherichia coli and Pseudomonas aeruginosa biofilms on siliconized latex urinary catheters. MATERIAL AND METHODS: The MICs and MBCs of norfloxacin, ciprofloxacin, ofloxacin, levofloxacin, clinafloxacin, sparfloxacin, trovafloxacin, and moxifloxacin for two strains of E. coli (CBR-3 and CBR-4) and two strains of P. aeruginosa (HUS-3 and PBR-2) were determined according to the NCCLS guidelines. The susceptibility of bacteria attached to siliconized latex catheters to fluoroquinolones was also evaluated. Catheter segments containing 6 or 24 hours old biofilms were used as inocula for the studies of antimicrobial activity against bacterial biofilms. RESULTS: MICs of ciprofloxacin for planktonic and attached bacteria were equal. MICs values for the others fluoroquinolones increased two or more times when bacterial biofilms were used as inocula, except for ofloxacin and E. coli CBR-4, trovafloxacin and E. coli CBR-3, and levofloxacin and trovafloxacin and P. aeruginosa PBR-4; in these cases the MIC values for planktonic and attached bacteria were similar. When bacteria attached to siliconized latex were used as inocula, MBCs values increased 8-> 4,096-fold for all the fluoroquinolones evaluated. CONCLUSIONS: E. coli and P. aeruginosa biofilms on siliconized latex were more resistant to the bactericidal activity of fluoroquinolones than planktonic bacteria. PMID- 10721560 TI - [The recommendations of GESIDA/SEFH/PNS for improving adherence to antiretroviral treatment. AIDS Study Group of the Spanish Society of Hospital Pharmacy and the National Plan on AIDS of the Minister of Health and Consumers]. AB - The main objective of HAART is to achieve a complete suppression of the viral replication for long time. However, when the therapeutic drug levels are low, HIV can replicate and it can develop resistances. This fact can be the reason of treatment failure, HIV transmission of resistant strains and therefore an inappropriate use of the economical resources. In order to get the adequate therapeutic drug levels it is necessary to have a good adherence to the treatment. We review the factors that influence the adherence, the evaluation methods and we recommend the possible intervention strategies which should be given by a multidisciplinary team, integrated by physicians, pharmacists, nurses, psychologists and other personal support. To start HAART is not an emergency. For this reason is very important to prepare to the patient and to identify the non adherence factors in order to correct it. Once the HAART is indicated it is very important to offer information during the medical prescription and when the drugs are dispensed. During the therapy is necessary to follow actively all patients on HAART. In order to make therapeutical decisions we need to know the patient drug adherence rate. We recommend to use several methods to calculate the drug adherence rate, being the most commonly used the patient interview, the patient questionnaire, the refill count, the pharmacy visits rate together with the viral load evolution of the patient. In order to get all this information it is necessary to have a very good communication between all the people involved in HIV infected patients care. If non-adherence is detected it is necessary to start the intervention strategies to correct it and if they fail it might be necessary in some cases to stop HAART. The potential benefits of the adherence programs can justify the economical spend in human and hospital facilities resources. PMID- 10721561 TI - [Chronic unilateral canaliculitis]. PMID- 10721562 TI - [Invasive otitis in an immunosuppressed patient]. PMID- 10721563 TI - [Clinical microbiology: is it a specialty in crisis?]. PMID- 10721564 TI - [Laryngeal tumor of laryngeal tuberculosis?]. PMID- 10721566 TI - [Bacteremia by Kluyvera ascorbata in a patient with neutropenia and fever]. PMID- 10721567 TI - [Bacteremia by Vibrio cholera no 01, two cases]. PMID- 10721565 TI - [Diagnosis of cerebral toxoplasmosis in an immunocompetent patient]. PMID- 10721568 TI - [Cutaneous sporotricoid infection by Mycobacterium marinum]. PMID- 10721569 TI - [Pneumonia and empyema caused by Moraxella osloensis]. PMID- 10721570 TI - [Rhabdomyolysis associated with pneumonia by Streptococcus pneumoniae and Legionella pneumophila]. PMID- 10721571 TI - [Bacterial vaginosis and vaginitis by Candida spp.: contradictory or compatible diagnosis?]. PMID- 10721572 TI - [Mycobacteriosis as an emerging disease]. PMID- 10721573 TI - [Diagnosis of pneumonia in intubated patients: the unsuccessful search for "El Dorado"]. PMID- 10721574 TI - [The microbiology laboratory in the diagnosis of bacteremia associated with catheters]. AB - Catheter related sepsis is an outstanding problem in patients in every age group. The microbiological diagnosis should consider the main pathways of infection (catheter-skin interface, endoluminal). With this aim we analysed 1496 central and peripheral short term catheters and 119 episodes of catheter related bacteremia. Catheters were cultured according to the quantitative technique of Brun Buisson (QT), the semiquantitative technique of Maki (SQ) and qualitative broth culture (QL). The following results of sensitivity, specificity, positive predictive value, negative predictive value and Youden index were obtained: SQ = 87%, 88%, 40%, 99%, 0.75; QT (> or = 10(2) CFU/ml) = 88%, 89%, 43%, 99%, 0.77; QT (> or = 10(3) CFU/ml) = 77%, 92%, 48%, 97%, 0.69; QL = 94%, 68%, 20%, 99%, 0.62. Results of SQ and QT > or = 10(2) were comparable, nevertheless, the addition of QT to SQ increased the detection of bacteremia by 12.8%, while in the opposite situation the increase was 10%. According to this, it is advisable to combine routinely SQ and QT. Finally, in 42 episodes of bacteremia related to implanted catheters processed by quantitative differential culture of blood drawn through the catheter and blood drawn through the peripheral vein the relationships were: > 1000 in 79% of cases, between 100 and 1000 in 9% of cases and between 5 and 10 in just 5% of cases. PMID- 10721575 TI - [Microbiological study of post mortem lung biopsy in pediatric patients]. AB - AIM: The aim of this study was to correlate the morphologic and microbiologic findings of the post mortem biopsies of a series of pediatric patients. MATERIAL AND METHODS: A complete morphologic study of all the organs of the patients included in the study was performed. In the pulmonary samples provided by the pathologist, the presence of aerobic and anaerobic microorganisms and fungi were determined. RESULTS: Ninety-three pulmonary biopsies corresponding to 77 dead patients (47 infants and 30 fetuses) were undertaken. Forty-five patients showed pulmonary histology of infectious processes. The concordance between the morphology and the microbiologic culture was of 66.2%. The microorganisms most frequently isolated were gram positive cocci (46.4%) followed by gram negative bacillus (30.4%). CONCLUSIONS: Although post mortem pulmonary tissue cultures should be interpreted with caution without first relating these with the morphologic findings and the clinical characteristics of the patient, a statistically significant agreement was observed between the anatomo-pathological study and the microbiologic findings. PMID- 10721576 TI - [Prevalence of Enterococcus faecalis and Enterococcus faecium with high resistance to aminoglycosides in the cities of Resistencia and Corrientes, Republic of Argentina]. AB - BACKGROUND: The objective of this study was characterize the prevalence of high level aminoglycosides resistance (HLRA) in Enterococcus faecalis and E. faecium, determine the relationship between high-level gentamicin resistance (HLGR) and other aminoglycosides, and their distribution according clinical samples (blood, urine and others). MATERIALS AND METHOD: A total of 177 strain (157 E. faecalis and 20 E. faecium) isolated from 1996 to 1998 were studied. They were identified by using classic methods. Their susceptibility to gentamicin, streptomycin, and kanamycin was tested by the disk diffusion technique using high-level disks in agar Muller Hinton. RESULTS: E. faecalis showed HLRG of 28.7%, streptomycin 28.7% and kanamycin 37.6%, E. faecium showed 50%, 40%, and 60% respectively. The strains with HLRA have a tendency to high-level resistances to streptomycin and kanamycin (p < 0.0005). Statistical analysis demonstrated significative differences among strains with HLRA isolated from blood, urine and other clinical samples (p < 0.0005 to gentamicin and streptomycin and 0.004 < p < 0.007 to kanamycin). CONCLUSIONS: The prevalence of HLRA enterococci found in the area os this study, justify its detection, particularity in cases of serious infections. PMID- 10721578 TI - [Urinary infection caused by non typhi Salmonella]. AB - BACKGROUND: The aim of this study was to know the frequency and the clinical characteristics of urinary infection by non typhi Salmonella in our area in Spain. PATIENTS AND METHODS: The clinical histories of patients with urinary infection by non typhi Salmonella diagnosed in the Hospital General del Guadalajara from January 1990 to July 1999 were reviewed. RESULTS: During the period studied nine patients with urinary infection by non typhi Salmonella were diagnosed, representing 0.056% of the urinary infection diagnosed in our hospital over the same period. All the patients presented underlying disease and five were undergoing immunosuppressor treatment. Four patients presented urological disease. The most frequent serogroup was Salmonella enteritidis (7 cases). All the episodes were symptomatic. The same microorganism was isolated in stools in four patients. The evolution was favorable in five of the nine cases. Recurrence was observed in two patients and secondary bacteremia in one. Six patients required antibiotic treatment over two or more weeks. The mean length of treatment was of 2.5 weeks. CONCLUSIONS: Urinary infection by non typhi Salmonella is predominantly observed in patients undergoing immunosuppression or with urological disease. Prolonged antibiotic treatment is recommended due to its bad evolution. PMID- 10721577 TI - [Seroepidemiological study of brucellosis in a rural endemic area]. AB - BACKGROUND: This study investigated the prevalence of Brucella spp. antibodies in the general population in the Health Area of Tremp (Region of Pallars Jussa, Lleida). It also identified the risk factors with the presence of these. PATIENTS AND METHODS: A total of 346 (191 men and 155 women) were studied. Information about the sex, age, location, the personal and familiar antecedents of brucellosis, occupational risk, contact with the animals and the consumption of non-hygienic dairy products was recorded. The estimation of the seroprevalence was carried out by the ELISA IgG test. The association of independent variables with the presence of antibodies was assessed by the Coombs to Brucella and the ELISA IgG tests. It was assessed by using the calculation of the analysis variance. RESULTS: The personal antecedents, the contact with the animals and the occupational risk all showed a statistically significant relation (p < 0.05) with the Coombs and ELISA IgG tests. The familiar antecedents showed a significant relation with the ELISA IgG. The consumption of dairy products and the location showed no statistically significant relation. A seroprevalence was obtained among the researched population of 11.9%, the maximum occurred in Isona surgery (25.6%) and the minimum in Tremp (9.8%). CONCLUSIONS: The seroprevalence is high and the epidemiological profile associated with the fact of being seropositive is associated with the profession of the study subject and it coincides with de infection mechanisms present in the area. PMID- 10721579 TI - [Infection and lipoproteins]. PMID- 10721580 TI - [Eye foreign object]. PMID- 10721581 TI - [Micronodular pulmonary infiltration in a patient with acquired immunodeficiency syndrome]. PMID- 10721582 TI - [Lumbosacral spondylodiscitis as the first manifestation of subacute bacterial endocarditis caused by Streptococcus sanguis type II]. PMID- 10721583 TI - [Sepsis caused by Streptococcus pneumoniae. Post mortem atypical diagnosis]. PMID- 10721584 TI - [Bilateral tubal abscess without peritonitis caused by Streptococcus pneumoniae]. PMID- 10721585 TI - [Asymptomatic pneumoperitoneum as the first manifestation of complicated diverticulitis in a renal transplant receptor]. PMID- 10721586 TI - [Aseptic meningitis caused by Mycoplasma pneumoniae]. PMID- 10721587 TI - [Hemophagocytic syndrome associated with cytomegalovirus and Mycobacterium xenopi disseminated disease in a patient infected by the human immunodeficiency virus who had a fatal outcome]. PMID- 10721588 TI - [Fibrinous peritonitis caused by Brucella sp]. PMID- 10721589 TI - [Bilateral tuberculous mastitis in a woman infected by the human immunodeficiency virus]. PMID- 10721590 TI - [Chronic otorrhea caused by Mycobacterium tuberculosis]. PMID- 10721591 TI - [Questions and answers on cryptococcal meningitis associated with human immunodeficiency virus infection]. PMID- 10721592 TI - [Medical practice]. PMID- 10721593 TI - [Valvulopathy in primary antiphospholipid syndrome. Prospective echocardiography study]. AB - Echocardiographic studies have demonstrated a high prevalence of valvular disease in patients with primary antiphospholipid syndrome (PAPS). However, there are no studies assessing changes over time in valvular abnormalities. We conducted a study to determine whether there are changes over time in valvular lesions as detected by serial transesophageal echocardiography (TEE). Twelve patients with a first TEE had a second evaluation after a mean period of 13.5 months. There were 10 women and two men with a mean age of 38 years. Two patients had normal TEE on both initial and follow-up studies. Ten patients (83%) had valvular abnormalities, predominantly of the mitral and aortic valves in both studies. Abnormalities consisted of thickening, nodules, regurgitation, regurgitation and stenosis, and calcification. The type and frequency of lesions changed over time. As an example, one mitral valve nodule disappeared on follow up but three new aortic nodules developed even though all patients were receiving anticoagulant therapy. Two patients with mitral and aortic nodules presented cerebral ischemia. Mitral valvuloplasty was performed in one case. These results show that valvular abnormalities in patients with PAPS resolve, appear, or persist irrespective of anticoagulant therapy. Regurgitation is often mild or moderate, but stenosis may appear. PMID- 10721594 TI - [Orbit decompression surgery in patients with exophthalmos caused by Graves Basedow disease]. AB - OBJECTIVE: To evaluate in a prospective trial two surgical technique routinely used for orbital decompression in Graves' disease ophthalmopathy. PATIENTS AND METHODS: Patients with Graves' disease exophthalmus (greater than 22 mm) that was euthyroid for at least six months after treatment were admitted to the study and randomly assigned to two groups, twenty six orbits in 17 patients were surgically decompressed with the Walsh and Ogura technique (group I) and 18 orbits in 18 patients were decompressed with Kennedy's surgical approach (group II). RESULTS: Both surgical techniques were equally effective in reducing the exophthalmus (p < 0.05). Morbility was significantly greater in terms of diplopia and infraorbital nerve lesion with Walsh and Ogura surgical approach. CONCLUSIONS: Due to reduced morbility, orbital surgical decompression with Kennedy's technique is recommended in patients with Graves' disease exofthalmus. PMID- 10721595 TI - [Management of hypertension emergencies in elderly patients with isosorbide dinitrate aerosol]. AB - In this clinical trial, we assessed the effectiveness and safety of isosorbide dinitrate spray administered through the oral mucosa in 20 elderly patients (> 60 years old) with a hypertensive emergency (mean arterial pressure > 140 mmHg and target-organ damage). The patents were given a first dose of 1.25 mg of spray when they were admitted; a second dose was administered 15 min. later if the mean arterial pressure had not decreased by > 15%. An electrocardiogram (ECG) was done on every patient immediately prior and 30 min. after administering the medication. Three patients (15%) had a good response with one dose while 17 patients (85%) required a second dose. Thirty patients had a significant reduction in arterial blood pressure (193 +/- 91,123 +/- 5.4 to 154 + 7.1/92.5 + 6.2 mmHg p < 0.005) as well as of the mean arterial pressure (146.8 +/- B to 113 +/- 5 mmHg 23%, p < 0.005 > in a period of 30 min. No adverse effects, rebound hypertension nor severe hypotension were observed. These figures remained under control for 3 h. Both ECG, were normal. A reduction of 13.5% heart rate was obtained (p < 0.005). Our observations suggest that isosorbide dinitrate aerosol is an effective and safe alternative for the treatment of elderly patients with hypertensive emergencies. PMID- 10721596 TI - [Plasma malondialdehyde in patients with type 2 diabetes mellitus and in patients with coronary disease]. AB - After intracoronary platelet aggregation, malondialdehyde (MDA), a lipid peroxide product is released. MDA renders some lipoproteins more atherogenic. OBJECTIVE: The objective of this study was to determine the sanguineous concentration of MDA in patients with type 2 diabetes mellitus (DM2) and patients with coronary disease. We measured methods and material MDA in plasma of 131 consecutive normal subjects, 44 hyperlipidemic, hyperglycemic patients with type 2-diabetes mellitus (DM2), 60 normolipidemic patients with angina, and 62 normolipidemic patients with acute myocardial infarction with and without DM2. STATISTICAL ANALYSIS: The concentration of MDA was lowest in normal subjects (42.5 +/- 7.2 micrograms per deciliter), intermediate in those with DM2 (62.7 +/- 10.1 micrograms per deciliter, p < 0.002), and highest in those with myocardial infarction (101.6 +/- 31.7 micrograms per deciliter, p < 0.001). The mean MDA concentration of patients with infarction was similar to that of patients with angina (121.8 +/- 51.9 micrograms per deciliter, p < 0.07). Stepwise logistic regression analysis showed that MDA was a possible predictor of myocardial infarction. CONCLUSIONS: The increase of plasma MDA might be a biochemical marker of coronary artery disease. We suggest that MDA levels greater than 62.7 micrograms per deciliter could indicate a high risk for myocardial infarction. PMID- 10721598 TI - [In Process Citation] PMID- 10721599 TI - [In Process Citation] PMID- 10721597 TI - [The right to free choice. 25 years of family planning in Mexico]. PMID- 10721601 TI - [In Process Citation] PMID- 10721600 TI - [In Process Citation] PMID- 10721602 TI - [Hyperbaric oxygenation therapy, basic concepts]. AB - Hyperbaric oxygen therapy (HBO) is defined as a treatment in which a patient breathes 100% oxygen in a pressurized environment of at least 1.4 absolute atmospheres. The first written reports data from the 15th century, when it was used to treat respiratory diseases. For some time its applications lacked scientific support until the second half of this century when scientific publications were carried out using current methodology. This type of therapy is grounded basically in three gas laws: Henry's Law, Dalton's Law and Boyle's Law. The beneficial effects are: wound healing enhancement; increased neutrophil bactericidal capacity; direct toxic effect against some microorganisms; arteriolar vasoconstriction with subsequent edema reduction and decreased ischemia/reperfusion injury, among others. These are the result of increased environmental pressure and high oxygen tension in body tissues. Currently there are 13 accepted conditions to be treated with HBO and others are still under investigation. Following UHMS-accepted treatment protocols, complications and/or adverse effects are limited. PMID- 10721603 TI - [A 57-year-old man with diabetes mellitus, liver cirrhosis, ascites and fever]. PMID- 10721604 TI - [Hemophilic pseudotumor in pediatric patient]. PMID- 10721605 TI - [Thyroiditis induced by radioactive iodine (I-131). Report of a case and review of the literature]. AB - A 23 years old female with Graves' disease initiated a symptomatic treatment with propanolol and then received radioactive iodine (I-311) 370 MBq (10 mCi) as definitive treatment. Three weeks later she was hospitalized because she presented acute radiation thyroiditis. She was discharged after 48 h of treatment, but clinical symptoms and signs persisted for over 30 days, but in less degree. One month later she developed hypothyroidism. There are three therapeutics options for the treatment of Grave's disease (antithyroid drugs, radioiodine and surgery). There are some hospitals in which the use of radioactive iodine is preferred as the first option, without a pretreatment with antithyroid drugs, but this could lead to acute thyroiditis or thyroid storm. PMID- 10721606 TI - [Contributions of the medical community to the Mexican revolution]. AB - Mexican physicians, faithful to their tradition of honor and patriotism, were present in the military and political events of the great Revolution, the began in 1910 and ended triumphantly in 1917. In the first phase, a Madero supporter and opposed to presidential reelection was doctor Francisco Vazquez Gomez, a specialist in otorhinolaryngology, Professor at the National Medical School and past President of our Academy of Medicine. The second phase of this Revolution, characterized by the struggle against the Huerta dictatorship and then by combats among revolutionary factions, also saw the intervention of many physicians and surgeons, such as senator Belisario Dominguez of Chiapas, a victim of dictatorial oppression. Among them were distinguished academicians such as doctors Rafael Silva of Mexico and Francisco Castillo Najera of Durango. Likewise devoted nurses were in Carranza's group, while medical students enlisted in Zapata's forces. The last phase of the Revolution was dominated by the activities of the Constituent Congress in Queretaro, which promulgated the New Mexican Constitution. Among 223 elected representatives, 20 were physicians and two pharmacists (10%), who had an excellent participation in the different sessions. The new Constitution, sworn and signed on February 5, 1917, added social guarantees to individual guarantees already established by the Constitution of 1857. PMID- 10721607 TI - [Finding and description of a new syndrome]. PMID- 10721608 TI - [Obstructive symmetric hypertrophic cardiomyopathy]. PMID- 10721609 TI - [The first sequenced human chromosome]. PMID- 10721610 TI - [Osteoporosis in males is a frequently overlooked risk]. AB - For many years, osteoporosis has been considered as a women's disease. Data show that the incidence of this bone thinning disease is high in men. Aging is accompanied with changes in circulating hormone levels. Thus, low testosterone levels is associated with osteoporosis in men. Like osteoporosis in females, in men, might be treated with calcium supplements, vitamin D and bisphosphonates. Recent report of treatment for osteoporotic men with bisphosphonate alendronate was successful in improving bone density at several sites in the male skeleton, in reducing the incidence of fractures and in preventing loss of height. The benefits and risks of hormone androgen replacement therapy in men are not fully defined, but the androgens supplementation is also expected to have favorable consequences on bone. PMID- 10721611 TI - Anthropological genetics in the 21st century: introduction. PMID- 10721612 TI - Spatial and temporal stability of mtDNA haplogroup frequencies in native North America. AB - Mitochondrial DNA lineage frequencies in prehistoric Aleut, eastern Utah Fremont, Southwestern Anasazi, Pyramid Lake, and Stillwater Marsh skeletal samples from northwest Nevada and the Oneota of western Illinois are compared with those in 41 contemporary aboriginal populations of North America. The ancient samples range in age from 300 years to over 6,000 years. The results indicate that the prehistoric inhabitants of North America exhibit the same level of mtDNA variability as contemporary populations of the continent. Variation in modern mtDNA haplogroup frequencies is highly geographically structured, and the prehistoric samples exhibit the same geographic pattern of variation. This indicates that differentiation of regional patterns of mtDNA lineage variation occurred early in North American prehistory (much more than 2,000 years B.P.), has remained relatively stable since its origin, and was little influenced by the disruptions hypothesized for other genetic systems as a result of population declines and relocations at contact. PMID- 10721613 TI - Quantitative trait locus mapping using human pedigrees. AB - In the past decade phenomenal progress has been made in molecular and statistical genetic methods for localizing quantitative trait loci. Because of these advances, we can anticipate a long period of active genetic research in which the genes influencing human quantitative variability will be mapped and their effects accurately evaluated. Here, we review the current state of the science in statistical genetic methods for quantitative trait linkage analysis. In particular, we detail a variance component-based framework for localizing quantitative trait loci and for accurately estimating their relative effect sizes. Attention is paid to the optimal design of human family studies for localizing genes of small to moderate effect. In addition, methods and strategies are described for dealing with the most important complications of quantitative variation, including the assessment of genotype x environment interaction and epistasis. PMID- 10721614 TI - Gene mapping in the 20th and 21st centuries: statistical methods, data analysis, and experimental design. AB - In the 20th century geneticists began to unravel some of the simpler aspects of the etiology of inherited diseases in humans. The theory of linkage analysis was developed and applied long before the advent of molecular biology, but only the technological advances of the second half of the 20th century made large-scale gene mapping with a dense genome-spanning set of markers a reality. More recently, the primary topic of interest has shifted from simple Mendelian diseases, for which genotypes of some gene are the cause of disease, to more complex diseases, for which genotypes of some set of genes together with environmental factors merely alter the probability that an individual gets the disease, although individual factors are typically insufficient to cause the disease outright. To this end, a great deal of dogma has evolved about the best way to skin this cat, although to date success has been minimal with any approach. We postulate that the main reason for this is a lack of attention to experimental design. Once the data have been ascertained, the most powerful statistical methods will not be able to salvage an inappropriately designed study (Andersen 1990). Each phenotype and/or population mandates its own individually tailored study design to maximize the chances of successful gene mapping. We suggest that careful consideration of the available data from real genotype phenotype correlation studies (as opposed to oversimplified theoretically tractable models), and the practical feasibility of different ascertainment schemes dictate how one should proceed. In this review we review the theory and practice of gene mapping at the close of the 20th century, showing that most methods of linkage and linkage disequilibrium analysis are similar in a fundamental sense, with the differences being related more to study design and ascertainment than to technical details of the underlying statistical analysis. To this end, we propose a new focus in the field of statistical genetics that more explicitly highlights the primacy of study design as the means to increase power for gene mapping. PMID- 10721615 TI - Geographic patterns: how to identify them and why. AB - Geographic patterns of genetic diversity allow us to make inferences about population histories and the evolution of inherited disease. The statistical methods describing genetic variation in space, such as estimation of genetic variances, mapping of allele frequencies, and principal components analysis, have opened up the possibility to reconstruct demographic processes whose effects have been tested by a variety of approaches, including spatial autocorrelation, cladistic analyses, and simulations. These studies have significantly contributed to our understanding of human genetic variation; however, the molecular data that have accumulated since the mid-1980s have also created new complications. Reasons include the generally limited sample sizes, but, more generally, it is the nature of molecular variation itself that makes it necessary to develop and apply specific models and methods for the treatment of DNA data. The foreseeable diffusion of laboratory techniques for the rapid typing of many DNA markers will force us to change our approach to the study of human variation anyway, moving from the gene level toward the genome level. Because extensive variation among loci is the rule rather than the exception, an important practical tip is to be skeptical of inferences based on single-locus diversity. PMID- 10721616 TI - Admixture studies in Latin America: from the 20th to the 21st century. AB - The present study is a review of admixture studies in Latin America, an interesting subject because of the unique history of the area, in which populations from 3 different origins had contact and intercrossed. The most often used methods of analysis of admixture in Latin America and some problems related to them, such as the determination of the parental populations and selection of genetic markers, are briefly reviewed. Several sources of data for admixture studies (surnames, quantitative traits, proteins, and molecular information) are summarized. The results obtained using protein systems and blood groups, the most often used markers in Latin America, are considered. They are classified according to their application in 3 groups of populations: urban centers, native Americans, and African-descended subjects. The data show that almost every population is dihybrid or trihybrid, and when African influence is not detected, it is probably due more to the method than to an absence of that contribution. A special section is dedicated to the direction of gene flow, and results about directional mating based on mtDNA, Y-chromosome, and nuclear DNA or proteins are also given. From these studies it is possible to conclude that Amerindian admixture came mainly from female lineages, but it is difficult to establish what happened with the African contribution. A last subject considered is the relation between interethnic crosses and diseases; it is easy to analyze that relation when the pathological condition is related to a unique allele, but when complex diseases are considered, the results are not as clear because of the influence of nongenetic factors. Finally, the perspectives for admixture studies in the 21st century are considered, and some attempts to predict their future in Latin America are made. PMID- 10721617 TI - Human adaptability research into the beginning of the third millennium. AB - Human adaptability, as a field of inquiry within human biology, became defined during the research activities of the International Biological Program (IBP) (1964-1974). During this period, research was focused on ecological, physiological, and genetic studies of human populations within the theoretical frameworks of adaptation and evolution. Other defining characteristics of the IBP human adaptability research were standardization of methods, multidisciplinary projects, international cooperation, and a concern with human health issues. Some observers suggest that this research contributed to the ongoing transformation of physical anthropology and related fields from a largely descriptive to an analytical science. During the 25 years between the end of the IBP and the present, a number of research trends have continued: Several new multidisciplinary projects were initiated and completed; a subfield of demography within human biology has matured; nutrition, infant and child growth, and health studies have proliferated; and molecular genetics and DNA analysis have superseded the earlier population genetics. International programs today are geared toward more practical and applied studies with less emphasis on basic science. Continuation of human adaptability research into the 21st century is likely to make contributions in 3 broad areas: population, environment, and health. Productive research is likely to contribute to these 3 areas in the following categories: reproduction, psychosocial stress, life span approaches to health, effects of losses in biodiversity on health, a human biology of poverty, emerging infectious diseases, epidemiology of modernization, evolutionary medicine, and aging. The success of much of this research in its contribution to knowledge will come from the integrated perspectives of a biobehavioral framework of inquiry. PMID- 10721618 TI - Tibetan and Andean patterns of adaptation to high-altitude hypoxia. AB - Understanding the workings of the evolutionary process in contemporary humans requires linking the evolutionary history of traits with their current genetics and biology. Unusual environments provide natural experimental settings to investigate evolution and adaptation. The example of high-altitude hypoxia illustrates some of the progress and many of the remaining challenges for studies of evolution in contemporary populations. Current studies exemplify the frequently encountered problem of determining whether large, consistent population differences in mean values of a trait reflect genetic differences. In this review I describe 4 quantitative traits that provide evidence that indigenous populations of the Tibetan and Andean plateaus differ in their phenotypic adaptive responses to high-altitude hypoxia. These 4 traits are resting ventilation, hypoxic ventilatory response, oxygen saturation, and hemoglobin concentration. The Tibetan means of the first 2 traits were more than 0.5 standard deviation higher than the Aymara means, whereas the Tibetan means were more than 1 standard deviation lower than the Aymara means for the last 2 traits. Quantitative genetic analyses of within-population variance revealed significant genetic variance in all 4 traits in the Tibetan population but only in hypoxic ventilatory response and hemoglobin concentration in the Aymara population. A major gene for oxygen saturation was detected among the Tibetans. These findings are interpreted as indirect evidence of population genetic differences. It appears that the biological characteristics of sea-level humans did not constrain high-altitude colonists of the 2 plateaus to a single adaptive response. Instead, microevolutionary processes may have operated differently in the geographically separated Tibetan and Andean populations exposed to the same environmental stress. Knowledge of the genetic bases of these traits will be necessary to evaluate these inferences. Future research will likely be directed toward determining whether the population means reflect differences identified at the chromosomal level. Future research will also likely consider the biological pathways and environmental influences linking genotypes to phenotypes, the costs and benefits of the Tibetan and Andean patterns of adaptation, and the question of whether the observed phenotypes are indeed adaptations that enhance Darwinian fitness. PMID- 10721620 TI - Coil design for real and sham transcranial magnetic stimulation. AB - Transcranial magnetic stimulation (TMS) can be used to excite the human cortex noninvasively. TMS also activates scalp muscles and sensory receptors; additionally, the loud sound from the stimulating coil activates auditory pathways. These side effects complicate the interpretation of the results of TMS studies. For control experiments, we have designed a coil that can produce both real and sham stimulation without moving the coil. The sham TMS is similar to the real TMS, except for the different relative direction of the currents in the two loops of the figure-of-eight coil. While the real TMS elicited activation of hand muscles, sham TMS had no such effect; however, the auditory-evoked potentials were similar. PMID- 10721619 TI - Contributions and promise of human behavioral genetics. AB - Human behavioral genetics has contributed greatly to our understanding of human behavioral development. Twin, family, and adoption studies have shown that genetic effects are ubiquitous and that both genes and environments contribute to individual differences in behavior. The unique ability of behavioral genetic methods to separate genetic from environmental effects has also led to important discoveries about how the environment works in development and to the elucidation of the complex ways environments and genes interact across the life span. Although quantitative methods have been the mainstay of the field of human behavioral genetics since Galton's time, the Human Genome Project and advances in molecular genetics are providing new tools and promise as we enter the 21st century. Thus the future of human behavioral genetics lies in the cross disciplinary exchanges and collaborations that will increasingly occur in the years to come among quantitative and molecular scientists who work with both animal and human systems. This research may someday culminate in an understanding of the biological basis of behavior that spans from how the brain develops and functions to a grasp of how genes influence thought at the molecular level. PMID- 10721621 TI - Applications of SPICE for modeling miniaturized biomedical sensor systems. AB - This paper proposes a model for a miniaturized signal conditioning system for biopotential and ion-selective electrode arrays. The system consists of three main components: sensors, interconnections, and signal conditioning chip. The model for this system is based on SPICE. Transmission-line based equivalent circuits are used to represent the sensors, lumped resistance-capacitance circuits describe the interconnections, and a model for the signal conditioning chip is extracted from its layout. A system for measurements of biopotentials and ionic activities can be miniaturized and optimized for cardiovascular applications based on the development of an integrated SPICE system model of its electrochemical, interconnection, and electronic components. PMID- 10721622 TI - A micropower dry-electrode ECG preamplifier. AB - This paper describes the development of a very low-power preamplifier intended for use in pasteless-electrode recording of the human electrocardiogram. The expected input signal range is 100 microV-10 mV from a lead-II electrode configuration. The amplifier provides a gain of 43 dB in a 3-dB bandwidth of 0.05 Hz-2 kHz with a defined high input impedance of 75 M omega. It uses a driven common electrode to enhance rejection of common-mode interfering signals, including low-frequency motion artifact, achieving a common-mode rejection ratio (CMRR) of better than 80 dB over its entire bandwidth. The gain and phase characteristics meet the recommendations of the American Heart Association, ensuring low distortion of the output ECG signal and making it suitable for clinical monitoring. The amplifier has a power consumption of 30 microW operating from a 3.3-V battery and is intended for use in small, lightweight, portable electrocardiographic equipment and heart-rate monitoring instrumentation. PMID- 10721623 TI - Effects of sample geometry and electrode configuration on measured electrical resistivity of skeletal muscle. AB - Over the past 40 years, researchers from a variety of scientific backgrounds have been using Rush's equations to analyze results of their electrophysiological studies. A lack of understanding of the constraints and the domain in which these equations are valid, often results in situations in which it is challenging to evaluate and compare results obtained by different investigators. In this paper, we reanalyzed the conditions for which Rush's equations were derived, and using mathematical modeling, computer simulation and in vitro measurements, we delineated areas of their appropriate application. Our studies showed that both sample geometry and test electrode configuration affect the measured tissue electrical resistivities: 1) The sample can be considered semi-infinite only if its dimensions are > 50 inter-electrode separation distances (IESD), and thickness > 2.5 IESD, 2) smaller sample sizes increase the transversally measured resistivity, 3) semi-infinite samples thinner than 2.5 IESD, and samples tested with needle electrodes demonstrate reduced anisotropy, and 4) when surface-spot electrodes are longitudinally aligned, as the IESD/tissue thickness ratio decreases, the measured resistivity increases. Our conclusion is that in most experimental situations, it is necessary to use modeling techniques to decouple the electrode configuration/sample geometry influence from the measured tissue resistivity. PMID- 10721624 TI - A viscoelastic model of phagosome motion within cells based on cytomagnetometric measurements. AB - Cytomagnetometry is a noninvasive method to investigate intracellular movements of organelles such as phagosomes by introducing magnetic particles into cells by phagocytosis, magnetizing them and measuring the field from the cells. To analyze the results of the cell-field measurement, we introduce a model for intracellular phagosome motion and investigate their behavior in terms of the cell field. The model includes an elastic body and two viscosity components which are ascribed to the filamentous structures surrounding the phagosomes. The magnetic relaxation phenomenon is assumed to derive from the rotationary Brownian motion as in our previous model. Although the model is simple, its behavior is not trivial because it contains a nonlinear term and the Brownian motion term. This model is the simplest one possible having a viscoelastic body and its behavior hence should be investigated thoroughly. PMID- 10721625 TI - Magnetostatic image current and its application to an analytic identification of a current dipole inside a conducting sphere. AB - The image solution for the static magnetic field outside a conducting sphere with an internal current dipole is considered. The image current, which is a linear distribution of magnetic dipoles on the line segment between the dipole point and the center of the sphere, is derived by using the fact that the induced current does not have any contribution to the radial component of the magnetic field outside the sphere. The image is then used to obtain some explicit formulas for identifying the location and tangential moment of the primary current dipole. This explicit identification method is also tested with a real model for a patient's brain. PMID- 10721626 TI - A model of EMG generation. AB - Simulation models are unavoidable in experimental research when the point is to develop new processing algorithms to be applied on real signals in order to extract specific parameter values. Such algorithms have generally to be optimized by comparing true parameter values to those deduced from the algorithm. Only a simulation model can allow the user to access and control the actual process parameter values. This constraint is especially true when dealing with biomedical signals like surface electromyogram (SEMG). This work is an attempt to produce an efficient SEMG simulation model as a help for assessing algorithms related to SEMG features description. It takes into account the most important parameters which could influence these characteristics. This model includes all transformations from intracellular potential to surface recordings as well as a fast implementation of the extracellular potential computation. In addition, this model allows multiple graphically-programmable electrode-set configurations and SEMG simulation in both voluntary and elicited contractions. PMID- 10721627 TI - Experimental and numerical determination of SAR distributions within culture flasks in a dielectric loaded radial transmission line. AB - The effect of dielectric loading on the cell layer specific absorption rate (SAR) within a T-75 culture flask being irradiated within a transverse electromagnetic (TEM) cell was studied both experimentally and numerically. Direct thermal measurements of a T-75 containing 40 mL of culture medium and resting upon a 3-mm thick slab of alumina ceramic (epsilon r = 9.6) revealed that, compared to the same flask resting upon a foam slab (epsilon r = 1.0) of the same thickness, the average SAR at the cell layer was increased roughly fourfold. This fourfold increase is significant experimentally because it allows biologists to perform experiments over a larger range of SAR values needed to determine possible dose response curves without the costs and difficulties of a fourfold increase in amplifier power. Finite-difference time-domain (FDTD) simulations of the SAR distribution were in good quantitative agreement with the experimental measurements. It is concluded that FDTD modeling can be a cost effective and scientifically acceptable means of obviating the thermal measurement of SAR. PMID- 10721628 TI - Quality driven gold washing adaptive vector quantization and its application to ECG data compression. AB - The gold washing (GW) adaptive vector quantization (AVQ) (GW-AVQ) is a relatively new scheme for data compression. The adaptive nature of the algorithm provides the robustness for wide variety of the signals. However, the performance of GW AVQ is highly dependent on a preset parameter called distortion threshold (dth) which must be determined by experience or trial-and-error. We propose an algorithm that allows us to assign an initial dth arbitrarily and then automatically progress toward a desired dth according to a specified quality criterion, such as the percent of root mean square difference (PRD) for electrocardiogram (ECG) signals. A theoretical foundation of the algorithm is also presented. This algorithm is particularly useful when multiple GW-AVQ codebooks and, thus, multiple dth's are required in a subband coding framework. Four sets of ECG data with entirely different characteristics are selected from the MIT/BIH database to verify the proposed algorithm. Both the direct GW-AVQ and a wavelet-based GW-AVQ are tested. The results show that a user specified PRD can always be reached regardless of the ECG waveforms, the initial selection of dth or whether a wavelet transform is used in conjunction with the GW-AVQ. An average result of 6% in PRD and 410 bits/s in compressed data rate is obtained with excellent visual quality. PMID- 10721629 TI - Vibration arthrometry in the patients with failed total knee replacement. AB - This is a preliminary research on the vibration arthrometry of artificial knee joint in vivo. Analyzing the vibration signals measured from the accelerometer on patella, there are two speed protocols in knee kinematics: 1) 2 degrees/s, the signal is called "physiological patellofemoral crepitus (PPC)", and 2) 67 degrees/s, the signal is called "vibration signal in rapid knee motion". The study has collected 14 patients who had revision total knee arthroplasty due to prosthetic wear or malalignment represent the failed total knee replacement (FTKR), and 12 patients who had just undergone the primary total knee arthroplasty in the past two to six months and have currently no knee pain represent the normal total knee replacement (NTKR). FTKR is clinically divided into three categories: metal wear, polyethylene wear of the patellar component, and no wear but with prosthesis malalignment. In PPC, the value of root mean square (rms) is used as a parameter; in vibration signals in rapid knee motion, autoregressive modeling is used for adaptive segmentation and extracting the dominant pole of each signal segment to calculate the spectral power ratios in f < 100 Hz and f > 500 Hz. It was found that in the case of metal wear, the rms value of PPC signal is far greater than a knee joint with polyethylene wear and without wear, i.e., PPC signal appears only in metal wear. As for vibration signals in rapid knee motion, prominent time-domain vibration signals could be found in the FTKR patients with either polyethylene or metal wear of the patellar component. We also found that for normal knee joint, the spectral power ratio of dominant poles has nearly 80% distribution in f < 100 Hz, is between 50% and 70% for knee with polyethylene wear and below 30% for metal wear, whereas in f > 500 Hz, spectral power ratio of dominant poles has over 30% distribution in metal wear but only nonsignificant distribution in polyethylene wear, no wear, and normal knee. The results show that vibration signals in rapid knee motion can be used for effectively detecting polyethylene wear of the patellar component in the early stage, while PPC signals can only be used to detect prosthetic metal wear in the late stage. PMID- 10721630 TI - Time-frequency analysis of myoelectric signals during dynamic contractions: a comparative study. AB - In this paper, we introduce the nonstationary signal analysis methods to analyze the myoelectric (ME) signals during dynamic contractions by estimating the time dependent spectral moments. The time-frequency analysis methods including the short-time Fourier transform, the Wigner-Ville distribution, the Choi-Williams distribution, and the continuous wavelet transform were compared for estimation accuracy and precision on synthesized and real ME signals. It is found that the estimates provided by the continuous wavelet transform have better accuracy and precision than those obtained with the other time-frequency analysis methods on simulated data sets. In addition, ME signals from four subjects during three different tests (maximum static voluntary contraction, ramp contraction, and repeated isokinetic contractions) were also examined. PMID- 10721631 TI - Nonlinear adaptive noise compensation in electrogastrograms recorded from healthy dogs. AB - Adaptive noise compensation is a popular method for improving signal-to-noise ratio in a variety of biomedical applications with its major disadvantage being the requirement for a reference channel containing noise strongly correlated to the noise in the primary channel. In many biomedical applications the utilization of a channel containing such noise without any representation of the information signal is difficult if not impossible. In this study we investigated the possibility of applying adaptive compensation in nonideal noise environments containing substantial presence of information signal in the reference channel. The signal in the reference channel was subjected to nonlinear manipulations for reducing the signal-to-noise ratio, thus diminishing the representation of information signal. The methodology was tested on canine electrogastrographic (EGG) signals of four unconscious dogs which underwent laparotomy and implantation of six pairs internal stainless steel electrodes in addition to the eight-channel abdominal EGG. Fourteen-channel (six internal and eight cutaneous) were obtained from each dog for 1/2 h. The signals were digitized and processed by computer. All internal signals showed regular and coupled gastric electrical activity with frequency of repetition in the normogastric range [3-9 cycles-per minute (cpm)]. A single pair of primary and reference channels was selected from each cutaneous recording and exponential manipulators in the reference channels were introduced. The manipulators were tuned to maximize the percent distribution of spectral components in the canine normogastric range of each frequency spectrum calculated from the signal at the output of the adaptive compensator. Significant increment in the percent distributions in the normogastric range (p < 0.01) was noted after tuning the exponential manipulator, and in many frequency spectra the recovery of the genuine dominant frequency peak of gastric electrical activity as determined by the internal recordings was noted. This study indicated that low percent distributions registered by some EGG channels are related to external nonlinear factors, the impact of which can be partially compensated. PMID- 10721632 TI - A spectral-discrimination method for tear-film lipid-layer thickness estimation from fringe pattern images. AB - Examination of the tear-film lipid layer is often helpful in the prognosis of prospective contact lens patients and contact lens related problems, and in the analysis of symptomatic noncontact lens-wearing patients. In particular, the thickness of the lipid layer is considered to be an informative cue in studying the tear-film stability and uncovering of certain disorders. We propose a method for the accurate estimation of the lipid-layer thickness, exploiting the intensity and color information in Fizeau fringe images. The technique is based on a quantitative measure for discriminating among the spectra associated with different thicknesses. We propose an optical system for imaging the interference patterns, develop a mathematical model based on the physics of the fringe formation and sensing, and describe the calibration of the optical system using this model. The thickness extraction is readily carried out utilizing a lookup table. The proposed method would enable objective evaluation of the lipid layer characteristics, and provide a means for examining the dynamic changes in its thickness and spatial distribution during inter-blink periods. PMID- 10721633 TI - Determination of intracranial tumor volumes in a rodent brain using magnetic resonance imaging, Evans blue, and histology: a comparative study. AB - The measurement of tumor volumes is a practical and objective method of assessing the efficacy of a therapeutic agent. However, the relative accuracy of different methods of assessing tumor volume has been unclear. Using T1-weighted, gadolinium enhanced magnetic resonance Imaging (T1-MRI), Evans Blue infusion and histology we measured intracranial tumor volumes in a rodent brain tumor model (RT2) at days 10, 16 and 18 after implantation of cells in the caudate putamen. There is a good correlation between tumor volumes comparing T1-MRI and Evans Blue (r2 = 0.99), T1-MRI and Histology (r2 = 0.98) and histology and Evans Blue (r2 = 0.93). Each of these methods is reliable in estimating tumor volumes in laboratory animals. There was significant uptake of gadolinium and Evans Blue in the tumor suggesting a wide disruption of the blood-brain barrier. PMID- 10721634 TI - In vivo quantification of a homogeneous brain deformation model for updating preoperative images during surgery. AB - Clinicians using image-guidance for neurosurgical procedures have recently recognized that intraoperative deformation from surgical loading can compromise the accuracy of patient registration in the operating room. While whole brain intraoperative imaging is conceptually appealing it presents significant practical limitations. Alternatively, a promising approach may be to combine incomplete intraoperatively acquired data with a computational model of brain deformation to update high resolution preoperative images during surgery. The success of such an approach is critically dependent on identifying a valid model of brain deformation physics. Towards this end, we evaluate a three-dimensional finite element consolidation theory model for predicting brain deformation in vivo through a series of controlled repeat-experiments. This database is used to construct an interstitial pressure boundary condition calibration curve which is prospectively tested in a fourth validation experiment. The computational model is found to recover 75%-85% of brain motion occurring under loads comparable to clinical conditions. Additionally, the updating of preoperative images using the model calculations is presented and demonstrates that model-updated image-guided neurosurgery may be a viable option for addressing registration errors related to intraoperative tissue motion. PMID- 10721635 TI - Three-dimensional blind deconvolution of SPECT images. AB - Thanks to its ability to yield functionally rather than anatomically-based information, the three-dimensional (3-D) SPECT imagery technique has become a great help in the diagnostic of cerebrovascular diseases. Nevertheless, due to the imaging process, the 3-D single photon emission computed tomography (SPECT) images are very blurred and, consequently, their interpretation by the clinician is often difficult and subjective. In order to improve the resolution of these 3 D images and then to facilitate their interpretation, we propose herein to extend a recent image blind deconvolution technique (called the nonnegativity support constraint-recursive inverse filtering deconvolution method) in order to improve both the spatial and the interslice resolution of SPECT volumes. This technique requires a preliminary step in order to find the support of the object to be restored. In this paper, we propose to solve this problem with an unsupervised 3 D Markovian segmentation technique. This method has been successfully tested on numerous real and simulated brain SPECT volumes, yielding very promising restoration results. PMID- 10721636 TI - Facilitation of thrombolysis in acute myocardial infarction. PMID- 10721637 TI - Lipid lowering strategies. PMID- 10721638 TI - Supporting a failing heart: a review. PMID- 10721639 TI - Fibrinogen and homocysteine levels in coronary artery disease. AB - Conventional risk factors like high serum cholesterol, smoking and hypertension do not explain all the mortality and morbidity due to coronary artery disease in Indian population. Novel factors like plasma fibrinogen and homocysteine have been currently recognised as independent risk factors for coronary artery disease. A case-control study was carried out to examine the role of plasma fibrinogen, homocysteine, lipid profile and anthropometric parameters in angiographically established coronary artery disease patients. The relationship between the biochemical and anthropometric parameters was also examined. Fifty eight male patients in the age range of 35-60 years with angiographically established coronary artery disease and equal number of matched-controls were the subjects of this study. Cases with coronary artery disease had significantly higher waist-to-hip ratio, waist-to-thigh ratio, plasma fibrinogen and total cholesterol. Mean plasma total homocysteine levels were not significantly different between cases and controls. In Indian population, elevated plasma fibrinogen and abdominal obesity appear to be significantly associated with coronary artery disease. PMID- 10721640 TI - Clinical and angiographic profile and follow-up of myocardial bridges: a study of 21 cases. AB - Myocardial bridging describes an angiographic entity, which is any degree of systolic narrowing of a coronary artery observed in at least one angiographic projection. Among the cineangiograms of 3200 patients reviewed, there were 21 cases (19 males) of myocardial bridges--incidence of 0.6 percent. Of these, seven had hypertrophic cardiomyopathy, six had atherosclerotic coronary artery disease and remaining eight had no evidence of either. All 21 patients had myocardial bridges in proximal or mid left anterior descending coronary artery. In addition, one case of hypertrophic cardiomyopathy had whole posterior descending coronary artery under a myocardial bridge. Another case of hypertrophic cardiomyopathy had a short normal segment of 5 mm inside a long myocardial bridge of 35 mm (tandem myocardial bridges). The length of the bridges varied from 10 to 35 mm (mean 24.5 +/- 4.5 mm) and diameter stenosis during systole varied from 40-90 percent (mean 70 +/- 8%). Two patients had large saccular coronary aneurysms proximal to the muscle bridge. Four of the eight patients who had neither hypertrophic cardiomyopathy nor coronary artery disease presented with acute anterior wall myocardial infarction and three of them had regional wall motion abnormality of left descending territory. Of the six patients who had coronary artery disease, one had 60 percent left descending artery lesion and two had recanalized segments proximal to the bridge. Five of the above six patients had significant stenosis of other coronary vessels. Four patients were lost to follow-up (mean period 3.4 +/- 2 years). In the coronary artery disease group, one patient underwent coronary artery bypass graft surgery for 3-vessel disease including graft to left descending artery and one developed inferior wall myocardial infarction. The patients in the hypertrophic cardiomyopathy group and "no hypertrophic cardiomyopathy-no coronary artery disease" group were free of events at last follow-up. Long-term prognosis of isolated myocardial bridges appears to be excellent. Degree of systolic narrowing or length of myocardial bridge does not correlate with event rates on follow-up. PMID- 10721641 TI - Post-infarction ischaemic mitral regurgitation: what determines the outcome. AB - Ischaemic mitral regurgitation is an important determinant of survival in patients with coronary artery disease. A retrospective analysis was performed to evaluate the overall outcome and its determinants in patients with ischaemic mitral regurgitation. Over a period of 10 years, 72 patients underwent operations for mitral regurgitation of ischaemic origin. Age ranged from 37 to 68 years (mean 54.6 +/- 10.4 years), and 62 (86.1%) were male. Thirteen (18%) patients had acute and 59 (82%) had chronic ischaemic mitral regurgitation. Twenty-one patients were in New York Heart Association class II, 32 in class III and 19 in class IV. Moderate to severe left ventricular dysfunction was present in 42 patients. Valve prolapse was present in 35 (48.6%) patients and restricted leaflet motion secondary to myocardial dysfunction was present in 37 (51.4%) patients. All the patients were operated using standard cardiopulmonary bypass technique. Mitral valve was replaced in 33 patients and repaired in 39. Repair included a combination of techniques: chordal transposition (n = 2), chordal shortening (n = 18), leaflet resection (n = 2), posterior collar annuloplasty (n = 35) and annuloplasty with flexible Duran's ring (n = 3). Operative mortality was 18.1 percent (13/72). Low cardiac output was the cause of death in the majority (n = 10). Acute presentation and presence of restricted leaflet motion were the significant predictors of early mortality. Follow-up ranged from 3 to 84 months (mean 41.6 +/- 10.2 months). Late mortality was 46.2 percent. Actuarial survival in operative survivors at five years was 44.4 +/- 8.8 percent. To conclude, ischaemic mitral regurgitation carries a poor early and late outcome, with left ventricular dysfunction and presence of restricted leaflet motion being important contributors to it. In addition, acute presentation also reflects greater early mortality. PMID- 10721643 TI - Captopril for prevention of contrast-induced nephropathy in diabetic patients: a randomised study. AB - Contrast-induced nephrotoxicity is an important cause of hospital-acquired acute renal insufficiency. Different modalities have been used to prevent contrast induced-nephrotoxicity namely saline infusion, mannitol, furosemide, calcium channel blockers, atrial natriuretic factor and dopamine infusion with variable success. The possible role of medullary ischaemia mediated by renin angiotensin system in genesis of contrast-induced nephrotoxicity prompted us to investigate the role of captopril (a sulfhydryl group containing angiotensin-converting enzyme inhibitor) in its prevention. Seventy-one patients of diabetes mellitus undergoing coronary angiography were included in the study. Patients randomised to receive captopril, received the drug in a dose of 25 mg thrice a day for three days, starting one hour prior to angiography while the patients in the control group underwent angiography in a routine manner without receiving captopril. Following angiography, patients in the control group developed a significant increase in serum creatinine and blood urea nitrogen levels, as compared to those who received captopril. Contrast-induced nephrotoxicity (i.e. a rise of 0.5 mg/dL in serum creatinine) developed in 29 percent of the control group. Administration of captopril reduced the risk of development of contrast-induced nephrotoxicity by 79 percent. Glomerular filtration rate as measured by Tc DTPA renal scanning prior to and 24-72 hours following angiography demonstrated a mean fall of 9.6 ml/min in the control group while those in the captopril group had a mean increase of 13 ml/min in glomerular filtration rate. We conclude that abnormalities of renal perfusion possibly mediated by renin angiotensin system are responsible for development of contrast-induced nephrotoxicity. Administration of the angiotensin-converting enzyme inhibitor, captopril, offers protection against development of contrast-induced nephrotoxicity. PMID- 10721642 TI - Immune response in acute coronary syndromes. AB - Inflammatory response in the atherosclerotic lesions of coronary artery disease, mediated by cellular immune mechanisms is well appreciated. The significance of the immuno-inflammatory processes for the development of acute ischaemic sequelae of these lesions remains unsettled. Fifty patients of acute coronary syndromes were studied for complement components and immunoglobin levels by single radial immunodiffusion method. Twenty-eight patients of acute myocardial infarction showed significantly lower levels of complement components C3 and C4 at admission (C3--69.19 +/- 12.91 mg% compared to 82.40 +/- 9.26 mg% in controls, p < 0.01; C4 -14.56 +/- 2.46 mg% compared to 18.53 +/- 2.69 mg% in controls, p < 0.01). Twenty two patients of unstable angina did not show any significant change (C3--83.14 +/ 8.01 mg% and C4--19.07 +/- 4.47 mg%). Sixteen patients of acute myocardial infarction who were thrombolysed with streptokinase showed a steep rise in the levels of complement components immediately after thrombolysis (C3--69.19 +/- 12.91 mg% before and 100.56 +/- 17.09 mg% after thrombolysis, p < 0.001; C4- 14.56 +/- 2.46 mg% before and 21.48 +/- 4.78 mg% after thrombolysis, p < 0.001). Plasma C3 and C4 levels in acute myocardial infarction showed no relationship with peak CPK levels. Plasma immunoglobulins remained unchanged in patients of acute coronary syndromes. PMID- 10721644 TI - Dilated cardiomyopathy in non-specific aortoarteritis. AB - Out of 195 cases of Takayasu's arteritis who presented in our institute between January 1988 and December 1997, 12 (5.58%) had dilated cardiomyopathy. Age of these patients ranged from 10 to 30 years (17.25 +/- 5.30 years) and male-female ratio was 1:11. All the cases had cardiovascular system features (dyspnoea, oedema, palpitation, angina, etc. but without hypertension), three had central nervous system features (headache, vomiting, convulsion etc.) and all had general systemic features like weight loss, malaise, fever, arthralgia etc. Electrocardiography, chest X-ray and echocardiographic findings were consistent with dilated cardiomyopathy. Haemodynamic findings showed raised left ventricular end-diastolic pressure and pulmonary capillary wedge pressure in all; raised pulmonary artery pressure, pulmonary vascular resistance, right ventricular pressure and right atrial pressure in 6, 6, 4 and 2 cases, respectively; reduced left ventricular peak systolic pressure in 10 cases but central aortic pressure and systemic vascular resistance in all the cases were within normal limits. Angiography showed type I, II and III involvement in 7 (majority), 3 and 2 cases, respectively. Coronary and pulmonary angiography were normal and left ventricular angiography showed poor left ventricular systolic function in all the cases. Histopathological study (on 3 autopsy cases) showed non-specific inflammation of myocardium with lymphocyte/mononuclear cell infiltration and normal coronary vessels. So, dilated cardiomyopathy in Takayasu's arteritis is not rare, though not much reported, and can influence the prognosis of aortoarteritis cases. PMID- 10721645 TI - Importance of venting the left ventricle in aortic valve surgery. AB - Routine use of left ventricular vent is controversial in patients undergoing open heart surgery. However, surgeons use it during valvular surgery to maintain a dry field to make the operation easier. In addition it helps to prevent left ventricular distention during the critical period of rewarming and reperfusion, if ventricular function does not return immediately following the release of aortic cross clamp. In our country, patients present for valvular surgery at a very late stage and they often have severe left ventricular hypertrophy. This may affect the return of cardiac rhythm after the release of aortic cross clamp with progressive left ventricular distention. In the authors' experience, insertion of left ventricular vent through the apex is occasionally necessary to decompress the left ventricle as the left atrial vent usually fails to do so. This paper deals with retrospective analysis of the seven patients (out of a total of 395 patients who underwent valve surgery) who required insertion of left ventricular vent through the apex and reviews the beneficial effects of an apical left ventricular vent under refractory circumstances. It is recommended that insertion of left ventricular vent through apex should be strongly considered in patients having severe aortic valve disease with hypertrophied hearts, if cardiac rhythm in not restored with conventional management with left atrial vent and 3 to 5 DC shocks following the release of aortic cross clamp. PMID- 10721646 TI - Surgical repair of pulmonary artery aneurysm following infective endocarditis in a patient with persistent ductus arteriosus. PMID- 10721647 TI - De novo coronary artery stenting in dextrocardia with acute coronary syndrome. PMID- 10721648 TI - Submitral pseudoaneurysm and anticardiolipin antibodies. PMID- 10721649 TI - Allograft vasculopathy in cardiac transplant recipients: an early experience. PMID- 10721650 TI - Stenting of left common femoral artery using the VascuCoil stent. PMID- 10721651 TI - Traumatic rupture of the mitral valve apparatus. PMID- 10721652 TI - Exceptional survival of a patient with ventricular septal defect and Eisenmenger syndrome. PMID- 10721653 TI - ECG differentiation between RCA and LCx occlusion. PMID- 10721654 TI - Molecular mechanism of carcinogenesis by human papillomavirus-16. AB - Human papillomaviruses (HPVs) are common DNA viruses in humans. Recently, epithelial cancers associated with HPV infection have been used as models of virus-induced carcinogenesis. HPVs can be divided into two groups, mucosal and cutaneous. HPV-16 is the most frequent mucosal type associated with cervical cancer. Although the molecular mechanisms of carcinogenesis by HPV-16 have not been completely elucidated, it is apparent that HPV infection is the major risk factor in cervical carcinogenesis. Two viral early genes, E6 and E7, and an upstream regulatory region (URR) are preserved in cervical carcinoma cell lines as well as in clinical samples of cervical cancer, indicating that these regions are important in cancer development. E6 and E7 function as transforming genes. E6 protein binds to and promotes degradation of the tumor suppressor protein, p53, while E7 protein complexes and inactivates the Rb protein; together, they disrupt cell cycle regulation. E6 and E7 are transcribed from a promoter, P97. P97 is regulated by complex interactions between multiple, positive and negative, cellular factors and the viral E2 product. E2 disruption caused by the integration into the cellular genome may induce overexpression of E6 and E7. The E6 and E7 proteins are thought to act as critical factors in cervical carcinogenesis by inactivating the two tumor suppressor proteins, p53 and Rb, which are commonly mutated in other human cancers. PMID- 10721655 TI - Acute infectious urticaria: clinical and laboratory analysis in nineteen patients. AB - We treated 19 Japanese patients with acute urticaria presumably caused by infection during the five years from 1994 to 1998. The patients' ages ranged from 2 to 66 years (8 males and 11 females). Most of them had urticaria, angioedema, high fever, neutrophilia, and high serum levels of C reactive protein (CRP). The skin rash lasted more than 24 hours. In four patients, a flow cytometric analysis revealed that the percentage of circulating T cells bearing T-cell receptor V beta 3 was decreased during the active stage and that this decrease was sustained for at least 2 to 3 weeks. This suggests that certain T-cell populations were numerically altered in association with the occurrence of the disease. A retrospective review indicated that the combination therapy with corticosteroid and antibiotics was more effective than the single use of either agent. PMID- 10721656 TI - Lichen planus with involvement of all twenty nails and the oral mucous membrane. AB - A 57-year-old man had had deformities of all ten fingernails for one and a half years before presentation and deformities of all ten toenails for the previous six months. The surfaces of the nails were rough, with excessive longitudinal striations. The bases of the nails were slightly hypertrophic, and the tips were atrophic and itchy. A longitudinal nail biopsy including the nail matrix revealed the typical histology of lichen planus. Reticulated pigmentation, maceration, and erosion on the buccal mucous membrane were also discovered. Histological analysis of the buccal mucous membrane revealed lichen planus intermingled with eosinophils. Immunological blood analysis revealed elevated CD4+ T cells and CD4/CD8 ratio. He worked as a tinsmith and had dental metal. The metal series patch test revealed positive reactions to chromate and tin. Treatment with systemic steroids was quite effective in treating the nail lesions. PMID- 10721657 TI - Perianal cytomegalovirus ulcer in an HIV infected patient: case report and review of literature. AB - We report the case of a 25-year-old man with acquired immunodeficiency syndrome, presenting with perianal ulcer and diarrhea. He had positive immunocytochemical tests for Cytomegalovirus (CMV) in circulating polymorphonuclear cells (PMN). The biopsy specimen was suggestive of CMV infection, and specific immunoperoxidase for CMV antigens positively stained endothelial cells and fibroblasts. In this report we review cutaneous CMV infection in immunocompromised patients. PMID- 10721658 TI - Primary pachydermoperiostosis: a case report. AB - Pachydermoperiostosis (PDP), a rare genodermatosis, occurred in a 38-year-old Indian male. He presented with progressive thickening of the skin on the face and scalp of 15 years duration. Widening of his wrists and ankles and broadening of the fingers and toes had also developed since then. He was born of a consanguineous marriage and had no family history of a similar disorder. He had the typical findings of complete form of PDP including cutis verticis gyrata, coarse facial features, clubbing of the digits in the skin, and periostosis and cortical thickening at the distal ends of long bones of the extremities and small bones of the hands and feet. PDP has two different forms--primary and secondary. These two entities are differentiated by family history and presence or absence of a primary lesion, usually in the lungs. Clinically, in secondary PDP, the cutaneous findings (pachydermia, seborrhoea, oiliness) are less severe than primary PDP; osteoarthropathy is more severe and painful in secondary PDP, especially with congenital heart disease. The present case was suffering from primary PDP that had expressed itself in its complete form. PMID- 10721659 TI - Complete remission of hypereosinophilic syndrome after interferon-alpha therapy: report of a case and literature review. AB - Hypereosinophilic syndrome (HES) is a multisystem disease with a significant mortality rate. It is characterized by peripheral blood eosinophilia and infiltration of eosinophils into many organs including skin. We describe a patient with cutaneous, pulmonary, and gastric symptoms of HES. He was first treated with systemic and topical corticosteroids, antihistamines, and photochemotherapy. Therapy was switched to interferon-alpha, because we had to consider side effects of corticosteroids and no significant improvement had been achieved. During the period of usage of interferon-alpha for 50 weeks, his symptoms of HES nearly cleared. There has been no recurrence in the eight months since the discontinuation of interferon-alpha therapy. PMID- 10721660 TI - Conventional cold excision combined with dermabrasion for rhinophyma. AB - A 65-year-old man, farmer by occupation, presented with redness and gradual enlargement of the nose. Examination revealed marked nodular enlargement of the nose and loss of normal nasal contours. Sebaceous material could be expressed from widened pores. The patient was diagnosed as rhinophyma of moderate degree. He was treated with cold knife excision combined with dermabrasion. A literature scan revealed that currently there is no evidence of superiority of much popular laser surgery over conventional cold knife surgery combined with dermabrasion for rhinophyma. Conventional surgery is time-tested, and it does not require expensive equipment or special training. PMID- 10721661 TI - Idiopathic calcinosis of the areola of the nipple. AB - Idiopathic calcinosis cutis involving the breast is a rare condition. Previously reported cases were detected by mammography without specific cutaneous findings. We report a case of idiopathic calcinosis of the areola of the nipple in a 32 year-old Korean woman that has unique clinical features resembling scrotal calcinosis. PMID- 10721662 TI - Keratosis lichenoides chronica: marked response to calcipotriol ointment. AB - Keratosis lichenoides chronica (KCL) is a rare dermatosis characterized by a distinctive seborrheic dermatitis-like facial eruption, together with violaceous, papular, and nodular lesions on the extremities and trunk, typically arranged in a linear and reticulate pattern. KLC is resistant to therapy, although spontaneous remission has been reported. We describe a 35-year-old woman with KLC who had the typical features of widespread violaceous, reticulate, and striae like eruptions with a prominent keratotic component over a nine-year period and who responded well to treatment with calcipotriol ointment. The immunohistochemical profiles are presented in addition to typical histopathologic features. PMID- 10721663 TI - Acantholytic acanthoma. PMID- 10721664 TI - Koebner phenomenon due to sacred thread in lichen planus. PMID- 10721665 TI - Angiokeratoma of the oral cavity and scrotum. PMID- 10721666 TI - Mitochondrial DNA mutations in Leigh syndrome and their phylogenetic implications. AB - Of 100 patients with the clinical diagnosis of Leigh syndrome, 21 were found to have specific enzyme defects: 15 involving cytochrome c oxidase (COX); 4, pyruvate dehydrogenase complex (PDHC); one, complex I (reduced nicotinamide adenine dinucleotide [NADH]-coenzyme Q reductase) and one, complex II (succinate ubiquinone reductase) deficiencies. In addition to the most common form of COX deficiency, mtDNA mutations in the adenosine triphosphatase (ATPase) 6 coding region were also commonly seen. Eighteen patients (18%) had mtDNA mutations at nucleotide position (np) 8993 or 9176. The mutated DNAs were present in a heteroplasmic state, comprising more than 90% of the DNA in muscle and/or blood samples from all patients. Patients with the T-to-G mutation at np 8993 usually had early onset of the disease with rapid progression, showing the typical clinical features of Leigh syndrome. On the other hand, those with the T-to-C 8993 mutation showed a milder and more chronic course. Patients with the mutation at np 9176 showed variable courses. Phylogenetic analysis of mtDNA D-loop sequences for the patients with the ATPase 6 mutations and normal Japanese subjects revealed that a T-to-G/C mutation at np 8993 and a T-to-C mutation at np 9176 occurred many times independently in the Japanese population. PMID- 10721667 TI - Y chromosomal DNA variation in east Asian populations and its potential for inferring the peopling of Korea. AB - We have examined variations of five polymorphic loci (DYS287, DXYS5Y, SRY465, DYS19, and DXYS156Y) on the Y chromosome in samples from a total of 1260 males in eight ethnic groups of East Asia. We found four unique haplotypes constructed from three biallelic markers in these samples of East Asians. The Japanese population was characterized by a relatively high frequency of either the haplotype I-2b (-/Y2/T) or II-1 (+/Y1/C). These dual patterns of the distribution of Y chromosomes (I-2b/II-1) were also found in Korea, although they were present at relatively low frequencies. The haplotype II-1 was present in Northeast Asian populations (Chinese, Japanese, Koreans, and Mongolians) only, except for one male from the Thai population among the Southeast Asian populations (Indonesians, Philippines, Thais, and Vietnamese). The Japanese were revealed to have the highest frequency of this haplotype (27.5%), followed by Koreans (2.9%), Mongolians (2.6%), and mainland Chinese (2.2%). In contrast, the frequency of the haplotype I-2b was found to be 17.1% in the Japanese, 9.5% in Indonesian, 6.3% in Korean, 3.8% in Vietnamese, and 2.7% in Thai samples. These findings suggested that the chromosomes of haplotype I-2b were likely derived from certain areas of Northeast Asia, the region closest to Southeast Asia. Phylogenetic analysis using the neighbor-joining tree also reflected a general distinction between Southeast and Northeast Asian populations. The phylogeny revealed a closer genetic relationship between Japanese and Koreans than to the other surveyed Asian populations. Based on the result of the dual patterns of the haplotype distribution, it is more likely that the population structure of Koreans may not have evolved from a single ancient population derived from Northeast Asians, but through dual infusions of Y chromosomes entering Korea from two different waves of East Asians. PMID- 10721668 TI - A nonsense mutation at Arg-1947 in the NF1 gene in a case of neurofibromatosis type 1 in a Korean patient. AB - We report a case of neurofibromatosis (NF) 1 presenting as a C-to-T transition changing an Arg-1947 codon to a stop codon. Because this mutation has been described in multiple Caucasian and Japanese families, the codon CGA for Arg-1947 in the NF1 gene is considered to be a hotspot for mutation in neurofibromatosis type 1 in all ethnic groups. PMID- 10721669 TI - Novel mutations of the ATP7B gene in Japanese patients with Wilson disease. AB - Wilson disease (WD) is an autosomal recessive disorder characterized by copper accumulation in the liver, brain, kidneys, and corneas, and culminating in copper toxication in these organs. In this study, we analyzed mutations of the responsible gene, ATP7B, in four Japanese patients with WD. By direct sequencing, we identified five mutations, of which two were novel, and 16 polymorphisms, of which 6 were novel. The mutations 2871delC and 2513delA shift the reading frame so that truncated abnormal protein is expected. In contrast to these mutations found in patients with hepatic-type of early onset, the mutations A874V, R778L, and 3892delGTC were either missense mutations or in frame 1-amino acid deletion, and occurred in the patients with hepato-neurologic type of late onset. The mutations 2871delC and R778L have been previously reported in a relatively large number of Japanese patients. In particular, R778L is known to be more prevalent in Asian countries than in other countries of the world. Our data are compatible with the hypothesis that the mutations tend to occur in a population-specific manner. Therefore, the accumulation of the types of mutations in Japanese patients with WD will facilitate the fast and effective genetic diagnosis of WD in Japanese patients. PMID- 10721670 TI - Human aryl hydrocarbon receptor nuclear translocator gene (ARNT) D/N511 polymorphism. AB - We found a novel A-->C change in codon 511 of the ARNT gene, which predicted the substitution of Asn (AAC) for Asp (GAC) at this position. Amplification using mismatched primers allowed the ARNT D/N511 polymorphism to be detected by digestion with endonuclease Tth111I. The frequency of the ARNT N511 allele was 0.019 in Caucasians and 0.026 in Africans. Because of the importance of the ARNT gene product in the metabolism of xenobiotics, this polymorphism may be useful in the study of associations with metabolic phenotypes and in pharmacogenetic studies. PMID- 10721671 TI - Human cathepsin S gene (CTSS) promoter -25G/A polymorphism. AB - We found a novel G-->A change at nucleotide -25 within the promoter of the CTSS gene encoding the elastase cathepsin S. The CTSS -25G/A polymorphism could be detected by digestion with endonuclease BfmI. The frequency of the CTSS -25A allele was 0.457 in Caucasians and 0.431 in Canadian Inuit. Because of the importance of the CTSS gene product in vascular matrix remodeling, this polymorphism may be useful in the study of associations with atherosclerosis and related phenotypes. PMID- 10721672 TI - An NsiI RFLP in the human long QT intronic transcript 1 (LIT1). AB - An NsiI polymorphic site has been found in the human long QT intronic transcript 1 (LIT1). In this transcript, we found a C-to-T transition, which was located between exons 10 and 11 of KVLQT1, and was confirmed by sequencing analysis. The allelic frequency of this polymorphism, was 0.82: 0.18 in Japanese individuals. Our novel polymorphism, combined with other polymorphisms, could be very useful in helping to determine whether the imprinting of LIT1 is disrupted in Beckwith Wiedemann syndrome (BWS) or in human cancers. PMID- 10721673 TI - Human hepatocyte nuclear factor-1 beta (HNF1B) 1968A/G polymorphism. AB - We found a novel A-->G change at nucleotide 1968 within the 3'-untranslated region of the HNF1B gene encoding the hepatocyte nuclear factor-1 beta. The HNF1B 1968A/G polymorphism could be detected by digestion with endonuclease MspI. The frequency of the HNF1B 1968G allele was 0.060 in Caucasians and 0.129 in Canadian Oji-Cree. Because of the importance of the HNF1B gene product in the regulation of transcription of several hepatic proteins, this polymorphism may be useful in the study of associations with metabolic phenotypes such as diabetes. PMID- 10721674 TI - Human C-reactive protein (CRP) 1059G/C polymorphism. AB - We found a novel G-->C change at nucleotide 1059 within exon 2 of the CRP gene encoding the C-reactive protein. The CRP 1059G/C polymorphism could be detected by digestion with endonuclease MaeIII. The frequency of the CRP 1059C allele was 0.109 in Caucasians, but it was absent from Canadian Oji-Cree. Because of the importance of the CRP gene product in inflammation and its recent association with ischemic heart disease syndromes, this polymorphism may be useful in the association studies of atherosclerosis and its related phenotypes. PMID- 10721675 TI - A novel nonsense mutation of the PEPD gene in a Japanese patient with prolidase deficiency. AB - A nonsense mutation at amino acid residue 184 in the human peptidase D (PEPD) gene caused the production of a truncated polypeptide. Characterizing molecular defects in patients provides clues to elucidate the relationship between the phenotype and the genotype. PMID- 10721676 TI - A novel type X collagen gene mutation (G595R) associated with Schmid-type metaphyseal chondrodysplasia. AB - Metaphyseal chondrodysplasia of the Schmid type (MCDS) is a skeletal dysplasia affecting the long bone metaphyses; it is characterized by short stature, bowlegs, and coxa vara. The spine is generally not involved. Here we report a novel missense mutation of the type X collagen gene in a sporadic case of MCDS. The mutation was a heterozygous single base-pair transition of G-to-A at nucleotide 1783, which predicted a substitution of glycine by arginine at codon 595 (G595R) in the carboxyl-terminal noncollagenous domain. Interestingly, another mutation of the same codon, in which glycine is substituted by glutamic acid (G595E), was previously reported to cause spondylometaphyseal dysplasia (SMD), another group of skeletal dysplasias with involvement of the spine in addition to the long tubular bones. The novel G595R mutation identified in the present study supports the concept of type X collagenopathy. PMID- 10721677 TI - Combination of mtDNA mutations in a patient with a mitochondrial multisystem syndrome. AB - We report a patient who manifested a heterogeneous clinical presentation, including hypertrophic cardiomyopathy and hypothyroidism, with initially limited central nervous system involvement, and who harbored the mitochondrial (mt)DNA A3243G mutation. MtDNA analysis also revealed deleted genomes in muscle and blood. This atypical molecular combination may have influenced the clinical phenotype. PMID- 10721678 TI - A novel missense mutation in the HMG box region of the SRY gene in a Japanese patient with an XY sex reversal. AB - The sex-determining region of the Y chromosome, the SRY gene, located on the short arm of the Y chromosome, is appreciated as one of the genes that is responsible for directing the process of sex differentiation. To date, 34 different mutations, including 29 missense and nonsense mutations in the SRY gene, have been described in XY female patients. We investigated the molecular basis of the sex reversal in one Japanese XY female patient by determining the nucleotide sequence of the SRY gene, using polymerase chain reaction and direct sequencing. We identified a novel mutation, of the substitution of Tyr for Asn at nucleotide position 87 (N87Y). This Asn residue is located within the DNA-binding high-mobility-group (HMG) motif, which is considered to be the main functional domain of the SRY protein. Further, this amino acid, Asn, is a conserved residue among mammalian SRY genes. These findings indicate that this amino acid substitution may be responsible for the sex reversal in this patient. PMID- 10721679 TI - Fibrillin gene (FBN1) mutations in Japanese patients with Marfan syndrome. AB - Marfan syndrome (MFS; MIM #154700) is a connective tissue disorder characterized by cardiovascular, skeletal, and ocular abnormalities. The fibrillin-1 gene (FBN1; MIM no. 134797) on chromosome 15 was revealed to be the cause of Marfan syndrome. To date over 137 types of FBN1 mutations have been reported. In this study, two novel mutations and a recurrent de-novo mutation were identified in patients with MFS by means of single-strand conformational polymorphism (SSCP) analysis. The two novel mutations are a 4-bp deletion at nucleotide 2820-2823 and a G-to-T transversion at nucleotide 1421 (C474F), located on exon 23 and exon 11, respectively. A previously reported mutation at the splicing donor site of intron 2 (IVS2 G + 1A), which is predicted to cause exon skipping, was identified in a sporadic patient with classical MFS. PMID- 10721680 TI - Functional activities of the amygdala: an overview. AB - Research to date into the amygdala shows that it has an integrative role in behavioural, vegetative and endocrine activities of animals in their relation with their environment. Animal studies show that amygdala has a role in emotional response, integrating input signals and initiating activities related to them. Different nuclei seem to have different effects. A complete picture of the functional roles of the amygdala is unavailable, and it has been suggested that the amygdala is functionally and anatomically heterogeneous. Amygdaloid subnuclei appear to have a role in the modulation of fear, in memory and attention, and in some sexual and sex-related behaviour of rats. In humans, functional magnetic resonance imaging shows that the amygdala responds preferentially to emotionally charged stimuli. Bilateral amygdala damage in humans can compromise the recognition of fear in facial expressions, an important ability in social judgement. Future study of the amygdala promises to shed light on emotional disorders in humans. PMID- 10721681 TI - Citalopram controls phobic symptoms in patients with panic disorder: randomized controlled trial. AB - OBJECTIVE: To examine the effects of long-term treatment with citalopram or clomipramine on subjective phobic symptoms in patients with panic disorder. DESIGN: Double-blind, parallel-group, five-arm study. PATIENTS: Patients aged 18 to 65 years with panic disorder (DMS-III-R diagnosis) and with no major depressive symptoms. INTERVENTIONS: Four hundred and seventy-five patients were randomized to 8 weeks of treatment with either citalopram (10 to 15 mg per day; 20 to 30 mg per day; or 40 to 60 mg per day), clomipramine (60 to 90 mg per day) or placebo. Two hundred and seventy-nine patients continued treatment after the 8 week acute phase. OUTCOME MEASURES: Phobic symptoms were assessed using the Phobia Scale and the Symptom Checklist's (SCL-90) phobia-related factors. RESULTS: At all dosages, citalopram was more efficacious than placebo, with 20 to 30 mg generally being the most effective dosage. Citalopram (20 to 30 mg) generally decreased phobic symptoms significantly more than placebo after Month 3. Interpersonal sensitivity decreased when measured on the respective SCL-90 sub scale. Alleviation of phobic symptoms generally continued to increase towards the end of the treatment. The effect of clomipramine was not as consistent. CONCLUSIONS: All active treatment groups, especially the group receiving 20 to 30 mg per day of citalopram, effectively controlled phobic symptoms in patients with panic disorder. Long-term treatment with citalopram further decreased phobic symptoms. PMID- 10721682 TI - Cholecystokinin-induced anxiety in rats: relevance of pre-experimental stress and seasonal variations. AB - OBJECTIVE: To examine the influence of pre-experimental stress on the anxiogenic like action of caerulein, an agonist of cholecystokinin (CCK) receptors. Differences in the anxiety levels of rats in summer and winter, and the role of CCK in these behavioural alterations, were also examined. DESIGN: Prospective animal study. INTERVENTIONS: Male Wistar rats were injected with the CCK agonist caerulein, or the CCK antagonists L-365,260 or devazepide, after being exposed to pre-experimental stress (handling and isolation). OUTCOME MEASURES: Performance in the plus-maze model of anxiety; serum levels of prolactin, thyrotropin and growth hormone; brain density and affinity of dopamine D2, serotonin 5-HT2 and CCK receptors. RESULTS: Caerulein (5 micrograms/kg, subcutaneous injection) caused the strongest action in animals brought to the experimental room immediately before the experiment and kept in isolation after the administration of caerulein. Caerulein did not cause any reduction of exploratory activity in rats made familiar with the experimental room and kept in the home-cage after the injection of the CCK agonist. The anti-exploratory action of caerulein in stressed rats was reversed by the CCK antagonist L-365,260 (100 micrograms/kg, intraperitoneal injection), demonstrating the involvement of the CCKB receptor subtype. In addition, seasonal fluctuations occur in the exploratory activity of rats; such activity was much lower in July than in November. The rats displaying the reduced exploratory activity had an increased number of CCK receptors in the frontal cortex and hippocampus. Simultaneously, the density of serotonin 5-HT2 receptors in the frontal cortex, but not that of dopamine D2 receptors in the striatum, was elevated. The blood level of growth hormone was also higher in July. CONCLUSIONS: The anti-exploratory action of caerulein appears to be dependent on the pre-experimental stress of rats. Moreover, the seasonal variations of exploratory behaviour of rats are evident in the plus-maze model of anxiety. The reduced exploratory activity in summer appears to be related to the elevated density of CCK and 5-HT2 receptors in the brain. PMID- 10721683 TI - Antidepressants upregulate messenger RNA levels of the neuroprotective enzyme superoxide dismutase (SOD1). AB - OBJECTIVE: To investigate the effect of amitriptyline, bupropion, doxepin or venlafaxine on the gene expression of the neuroprotective enzyme superoxide dismutase (SOD1) in a catecholamine cell in vitro model. DESIGN: Molecular study of a cultured cell line. INTERVENTIONS: Rat pheochromocytoma (PC12) cells were incubated in 1 and 10 mumol/L of various antidepressant medications for 24 or 48 hours. OUTCOME MEASURES: Northern blot analysis. RESULTS: Amitriptyline up regulated SOD1 messenger RNA in a time- and dose-dependent manner. The greatest up-regulation was following incubation with 10 mumol/L amitriptyline for 48 hours. The addition of bupropion, doxepin or venlafaxine to PC12 cell cultures also up-regulated SOD1 mRNA. CONCLUSIONS: These findings suggest that some antidepressants have the ability to positively regulate neuroprotective genes. PMID- 10721684 TI - Use of slow-release melatonin in treatment-resistant depression. AB - OBJECTIVE: To examine antidepressant augmentation with and hypnotic effects of slow-release melatonin (SR-melatonin) in patients with treatment-resistant depression. DESIGN: Open-label trial. SETTING: Tertiary care outpatient depression clinic. PATIENTS: Nine outpatients who had failed to respond to 2 or more 8-week trials of antidepressant medication. INTERVENTIONS: Patients received SR-melatonin 5 mg per day for the first 2 weeks and 10 mg per day for the final 2 weeks, in addition to their antidepressant medication. OUTCOME MEASURES: Structured Clinical Interview for DSM-IV, Axis 1 Disorders, Hamilton Rating Scale for Depression (HRSD), Beck Depression Inventory, Response Style Questionnaire, sleep and fatigue measures. RESULTS: One patient was excluded after 1 week because of the development of a mixed affective state. In the remaining 8 patients there was a 20% mean decrease in HRSD scores after 4 weeks of treatment, with no individual achieving an improvement of 50% or more. There was a 36% decrease on the 3-item HRSD related to insomnia, with 4 of 8 patients showing at least a 50% improvement on this measure. The greatest decrease in insomnia occurred during the last 2 weeks of the study, following the increase in dosage to 10 mg per day of SR-melatonin. Patients also reported significantly lower levels of fatigue post-treatment. CONCLUSIONS: SR-melatonin may be a useful adjunct for sleep, but does not substantially augment existing antidepressant therapies in some patients with treatment-resistant depression. PMID- 10721685 TI - Early- versus late-onset bipolar II disorder. AB - OBJECTIVE: To compare the clinical features and the outcome between patients with early- and late-onset bipolar II disorder. DESIGN: Case series. SETTING: Outpatient private practice. PATIENTS: One hundred and seventy-nine consecutive outpatients with bipolar II disorder presenting for treatment of a major depressive episode. OUTCOME MEASURES: Duration of illness, severity of depression, recurrences, psychosis, chronicity, atypical features and comorbidity. RESULTS: Patients with early-onset (before 20, 25 or 30 years of age) bipolar II disorder had a significantly longer duration of illness and more recurrences compared with patients with late-onset (after 20, 25 or 30 years of age) bipolar II disorder. All other variables were not significantly different between the 2 groups. CONCLUSIONS: Indicators of worse outcome (severity of depression, psychosis, chronicity, comorbidity) were not significantly different between patients with early- and late-onset bipolar II disorder. PMID- 10721686 TI - Use of home and community-based services by elderly black and white females. AB - The study purpose was to determine the impact of demographic, social, environmental, and health indicators on utilization of community-based services among black and white female elders. Existing data from a regional Area Agency on Aging was used and the sample (N = 1816) included low income and rural females. Races differed in use and services most frequently used were case management, outreach, congregate meal, and home delivered meals. Multiple linear and logistic regression indicated that age, payment source, income adequacy, residence, health conditions, sensory impairment, and function were associated with the number and types of services used, but these differed by race. Study findings have implications for health care providers, educators, policy makers, and planners. PMID- 10721688 TI - Geographic variation in preventable hospitalization of older women and men: implications for access to primary health care. AB - This study demonstrates how readily available data and small area analysis can be used to identify potential problems of access to primary care services for older women and men. Gender and socioeconomic differences in rates of preventable hospitalization are examined. Using hospital discharge data, five county and twenty-four intra-county areas in Upstate New York are studied. There is significant variation in preventable hospitalization within counties. Areas having significantly higher rates of these hospitalizations tend to have higher rates for both women and men. Problems of access are associated with lower income areas for women and men. PMID- 10721687 TI - Gender differences in positive and negative self-assessments of health status in a national epidemiological study of Israeli aged. AB - The literature in subjective health appraisals frequently notes that elderly women, more so than men, generally experience a lower quality of life in all major indicators (physical health status, functional ability, perceived income adequacy, social contacts, psychological distress, and cognitive ability). The current epidemiological study, of 1,352 reporting Israeli subjects between the ages of 75-94, was undertaken in order to obtain reliable estimates of "poor" and "excellent/good" self assessments of health in a national sample of aged; to identify the most significant correlates of "poor" and "excellent/good" assessments; and to ascertain whether the models of "poor" and "good/excellent" subjective health are different for elderly men and women. While it was found that women indeed rate their health as being poorer than men, of greater theoretical interest was the finding that the pattern of variables predicting to "poor" and "good/excellent" health are different for men and women. The findings point to the fact that the simple health self-evaluation question is not a unitary construct, but rather a complex attitudinal measure which yields different structural and conceptual results when controlling for the subjective health outcome ("poor" or "good/excellent") and when analyzing gender dichotomized models. PMID- 10721689 TI - How women experience menopause: the importance of social context. AB - This study analyzes personal accounts of women's menopausal experiences to understand why most women view menopause as an insignificant event, despite negative cultural and medical constructions of menopause as a time of "loss." We analyze 16 in-depth interviews with a diverse sample of women to examine how social contexts affect women's experiences with menopause and the meaning of those experiences. We find that most women view menopause as inconsequential because other events of midlife are more important or stressful to them. However, when cultural and medical contexts are examined, we find that some women do not avoid others' negative constructions of menopause as a time of "loss." PMID- 10721690 TI - Sarcopenia and decreased muscle strength in the elderly woman: resistance training as a safe and effective intervention. AB - A principle component of age-related weakness and frailty in women is sarcopenia. This decrease in skeletal muscle mass is a progressive syndrome that will affect the quality of life for elderly women by decreasing the ability to perform many activities of daily living. Strength training is known to be an effective means of increasing muscular strength and size in many populations, and can be utilized successfully to significantly improve muscle strength, muscle mass and functional mobility in elderly women up to the age of 96 years. Such exercise can minimize the syndrome of physical frailty due to decreased muscle mass and strength. Any rehabilitation or exercise program for the elderly woman would benefit from the inclusion of such a training regime. PMID- 10721691 TI - Caring together for elderly mothers: a qualitative study of relations between adult daughters and supportive home care workers. AB - This qualitative study examines relationships between adult daughters caring for elderly disabled mothers and the mothers' personal care workers (PCWs) paid directly by the Wisconsin Community Options Program (COP). A subset of a larger study, in these five cases PCWs provide substantial hands on care without substituting for the heavy care also provided by the daughters. Direct payment offers the daughters and workers freedom to schedule around their family obligations and other limits and tailor care to the abilities of all three participants. It also allows the daughter to be the paid provider when she chooses. Expansion of such supportive services could benefit many more low and middle income families. PMID- 10721692 TI - TAFs revisited: more data reveal new twists and confirm old ideas. AB - Synthesis of messenger RNA by RNA polymerase II requires the combined activities of more than 70 polypeptides. Coordinating the interaction of these proteins is the basal transcription factor TFIID, which recognizes the core promoter and supplies a scaffolding upon which the rest of the transcriptional machinery can assemble. A multisubunit complex, TFIID consists of the TATA-binding protein (TBP) and several TBP-associated factors (TAFs), whose primary sequences are well conserved from yeast to humans. Data from reconstituted cell-free transcription systems and binary interaction assays suggest that the TAF subunits can function as promoter-recognition factors, as coactivators capable of transducing signals from enhancer-bound activators to the basal machinery, and even as enzymatic modifiers of other proteins. Whether TAFs function similarly in vivo, however, has been an open question. Initial characterization of yeast bearing mutations in particular TAFs seemingly indicated that, unlike the situation in vitro, TAFs played only a minor role in transcriptional regulation in vivo. However, reconsideration of this data in light of more recent results from yeast and other organisms reveals considerable convergence between the models derived from in vitro experiments and those derived from in vivo studies. In particular, there is an emerging consensus that TAFs represent one of several classes of coactivators that participate in transcriptional activation in vivo. PMID- 10721693 TI - MDM2--master regulator of the p53 tumor suppressor protein. AB - MDM2 is an oncogene that mainly functions to modulate p53 tumor suppressor activity. In normal cells the MDM2 protein binds to the p53 protein and maintains p53 at low levels by increasing its susceptibility to proteolysis by the 26S proteosome. Immediately after the application of cellular stress, the ability of MDM2 to bind to p53 is blocked or altered in a fashion that prevents MDM2 mediated degradation. As a result, p53 levels rise, causing cell cycle arrest or apoptosis. In this review, we present evidence for the existence of three highly conserved regions (CRs) shared by MDM2 proteins and MDMX proteins of different species. These highly conserved regions encompass residues 42-94 (CR1), 301-329 (CR2), and 444-483 (CR3) on human MDM2. These three domains are respectively important for binding p53, for binding the retinoblastoma protein, and for transferring ubiquitin to p53. This review discusses the major milestones uncovered in MDM2 research during the past 12 years and potential uses of this knowledge in the fight against cancer. PMID- 10721694 TI - Identification and analysis of a third mouse Polycomb gene, MPc3. AB - A new mouse Polycomb (Pc) gene, MPc3, has been identified. The MPc3 protein contains the highly conserved chromodomain and carboxy-terminal COOH box of other known Pc proteins from diverse species. Like other Pc proteins, MPc3 physically interacts with the RING finger proteins RING1A and dinG/RING1B. MPc3 maps to the distal arm of mouse chromosome 11 (11E2), a region that contains other known Pc genes in addition to several disease loci that may be linked to abnormal Pc gene function. PMID- 10721695 TI - Characterization of the mouse JAB1 cDNA and protein. AB - JAB1 was originally described as a transcriptional coactivator of c-Jun and Jun D. Recent data suggests that JAB1 is a component of a large protein complex, the JAB1 signalosome in mammals and the COP9 complex in plants. The JAB1 signalosome is implicated in the phosphorylation of selected transcription factors, while the COP9 complex is involved in repression of photomorphogenesis in Arabidopsis. In this study, we describe the partial characterization of mouse JAB1 (mJAB1). The murine JAB1 protein is encoded by a gene located on mouse chromosome 1. mJAB1 mRNA is abundantly expressed in a variety of adult tissues as well as in mouse embryos. The JAB1 protein was readily detectable in many cell types and localized to both the nucleus and cytoplasm. Endogenous JAB1 protein is relatively stable and its degradation is not perturbed by blocking 26S proteasome activity, suggesting that this protein is not degraded by the ubiquitin-mediated proteolytic pathway. PMID- 10721696 TI - Cloning, genomic organization and chromosomal localization of the gene encoding the murine sodium-dependent, purine-selective, concentrative nucleoside transporter (CNT2). AB - A PCR-based strategy was used to isolate a 2653 bp cDNA encoding the mouse sodium dependent, purine nucleoside selective, concentrative nucleoside transporter (designated mCNT2). The deduced protein sequence exhibits 93 and 80% identity to the previously cloned rat and human sodium-dependent, purine nucleoside selective, nucleoside transporters, respectively. Characterization of 3H nucleoside uptake by COS-1 cells transiently transfected with the cDNA demonstrated that it encoded a functional nucleoside transport activity with selectivity for purine nucleosides. The cDNA was used to screen a murine (strain 129SvJ/6) genomic library in pBeloBAC11 to identify a clone containing the mCNT2 gene. A PCR strategy was used to identify and sequence the intron-exon boundaries and to determine the approximate sizes of the introns. The mCNT2 gene spans approximately 13.7 kb and is encoded by 15 exons. The gene was mapped to mouse chromosome 2e3 by fluorescence in situ hybridization. PMID- 10721697 TI - Structural analysis of the gene encoding RP58, a sequence-specific transrepressor associated with heterochromatin. AB - RP58, a sequence-specific transcriptional repressor sharing homology with the POZ domain of a number of zinc-finger proteins, is highly synthesized in brain and localized in condensed chromatin regions, suggesting a role in transcriptional repression in the central nervous system. In the present study, genomic clones of the human rp58 gene were isolated to determine the complete genomic organization. Sequence analyses indicated that the human rp58 gene encoding the functional protein is uninterrupted over its entire 4.2 kb length. Comparison of the human and mouse rp58 genes revealed that they share not only a high homology in the amino acid sequences of their encoded proteins, but also a high degree of structural similarity at the genomic level. RT-PCR analysis also demonstrated the existence of an alternatively spliced form of rp58 similar to the previously reported zinc-finger cDNA, C2H2-171. Chromosomal mapping by fluorescence in situ hybridization analysis allowed localization of the rp58 gene to human chromosome 1q44 ter, a genetic region associated with a number of human malignancies and neurological disorders. PMID- 10721698 TI - Characterization of the cystatin B gene promoter harboring the dodecamer repeat expanded in progressive myoclonus epilepsy, EPM1. AB - Mutations in the gene encoding cystatin B (CSTB) are responsible for the primary defect in progressive myoclonus epilepsy of Unverricht-Lundborg type (EPM1). A novel and unique type of disease-causing mutation, an unstable dodecamer repeat expansion, accounts for the majority of EPM1 patients world-wide. This minisatellite repeat expansion, located in the putative promoter of CSTB 175 bp upstream from the translation initiation codon, appears to downregulate CSTB gene expression in vivo. We report here the characterization of the CSTB promoter using different promoter-luciferase gene constructs. Transient transfections of cultured mammalian cells suggest that the region from -670 to -1 bp from the translation initiation codon functions as the CSTB promoter. Active binding to five Sp1 and four AP1 sites as well as weak binding to an androgen response element (ARE) half site was demonstrated by electrophoretic mobility shift assays. The effect of the minisatellite expansion on the promoter activity was evaluated by comparing the activity of constructs containing wild-type and expanded alleles. An increase in the number of dodecamer units from three to 19 repeats lowered transcription in vitro by 10-fold. Northern analysis of lymphoblastoid RNA from individuals with 'premutation' length dodecamer repeat (12-17 copies) expansions showed decreased levels of CSTB mRNA expression. These data indicate that expansion of the dodecamer repeat located in the proximal promoter of CSTB severely disrupts the function of the promoter and thereby reduces transcription of CSTB. PMID- 10721699 TI - Structural analysis and promoter characterization of the human membrane-type matrix metalloproteinase-1 (MT1-MMP) gene. AB - Membrane type-1 matrix metalloproteinase (MT1-MMP) degrades extracellular matrix components directly and indirectly by activation of other matrix metalloproteinases (MMPs). In the present study, we have isolated and characterized the human MT1-MMP gene and its promoter. The gene consists of 10 exons and nine introns spanning more than 10 kilobases (kb). The locations of two exon-intron splicing sites are distinct from the preserved positions among other known MMP genes. Primer extension and RNAse and S1 nuclease protection analyses indicated that there are four major and several minor transcription start sites. The 5'-flanking sequence of the gene contains putative regulatory elements, including one Sp-1 site and four CCAAT-boxes, whereas there is no TATA-box. The Sp-1 binding site was functional, as shown by gel shift and supershift analyses. Transfection studies with promoter constructs containing 0.1 to 7.2 kb of 5' flanking sequence coupled to a luciferase reporter gene indicated that the promoter contains additional positive and negative regulatory sequences. Deletion of the Sp-1 binding site by site-directed mutagenesis reduced luciferase activity by about 90%, demonstrating the crucial role of this element in maintaining MT1 MMP transcription. Our findings indicate that the human MT1-MMP promoter has distinctive structural and functional features compared with other MMP genes, which may lead to a unique expression pattern and regulation during physiological and pathological processes. PMID- 10721700 TI - Cryptic polyadenylation sites within the coding sequence of three yeast genes expressed in tobacco. AB - Three yeast genes, MIP (mitochondrial DNA polymerase) and two genes, YCF1 (yeast cadmium factor 1) and PDR5 (pleiotropic drug resistance 5), conferring multidrug resistance, were provided with the cauliflower mosaic virus 35S transcription promoter and introduced into tobacco using an Agrobacterium tumefaciens T-DNA derived vector. Transcripts of each gene much shorter than those expected were found in the transgenic plants. RT-PCR and S1 nuclease mapping of the PDR5 and MIP transcripts demonstrated the presence of one (PDR5), or several close (MIP), cryptic polyadenylation site(s) within the coding sequence of these yeast genes. Possible sequences involved in polyadenylation are discussed. PMID- 10721701 TI - Cloning and characterization of complementary DNA encoding human N acetylglucosamine-phosphate mutase protein. AB - Endothelial cells express erythropoietin receptor (EpoR) and are responsive to erythropoietin (Epo). Upon ligand binding, EpoR activates multiple signaling cascades. Identification of genes expressed in response to Epo is important for understanding the molecular nature of the signals. Applying the differential display approach, an effective method for analysis of gene expression, we identified five differentially expressed mRNAs. In this study, we cloned human N acetylglucosamine-phosphate mutase from a human microvascular endothelial cell (HMVEC) cDNA library using one of the differentially expressed fragments as a probe. The nucleotide (nt) sequence analysis of the longest clone displayed a 2 kb cDNA fragment and encodes a protein of approximately 542 amino acids with a predicted MW of approximately 60 kDa. Northern blotting and reverse transcriptase polymerase chain reaction analysis revealed an upregulation of the N acetylglucosamine-phosphate mutase mRNA after 2 h of stimulation of cells with Epo. This gene was shown to be variably expressed in human tissues and is located on chromosome 6. These studies demonstrate that the expression of N acetylglucosamine-phosphate mutase mRNA responds to cytokines, and the presence of a 10 aa motif similar to the putative active site of several hexose-phosphate mutases provides a basis for future studies of the role of this gene in the regulation of Epo-stimulated endothelial cell proliferation. PMID- 10721702 TI - Construction of an efficient expression system for Agrobacterium tumefaciens based on the coliphage T5 promoter. AB - A versatile expression vector utilizing a promoter of coliphage T5, P(N25) (Gentz and Bujard, 1985. J. Bacteriol. 164, 70-77) and a derivative of the IncW broad host-range plasmid pJB20 (Beaupre et al., 1997. J. Bacteriol. 179, 78-89) has been developed. This vector successfully expresses virulence proteins of Agrobacterium tumefaciens encoded by virG and a mutant allele of virA, virA (delta1-284, G665D) in Escherichia coli as well as in A. tumefaciens. The signal transduction proteins VirA (delta1-284, G665D) and VirG are fully functional when expressed in Agrobacterium, and the P(N25) driven expression overrides the complex transcriptional regulation present with the native promoters. This expression system will enable a more detailed analysis of the activation events in signal transduction in A. tumefaciens, and we expect it to be useful in other prokaryotes. PMID- 10721703 TI - Genomic structure and chromosomal localization of the rat protein kinase Cdelta gene. AB - Protein kinase Cdelta (PKCdelta) is a widely expressed calcium-independent PKC isozyme that is induced at mRNA and protein levels upon stimulation of different cellular pathways. We found the rat PKCdelta gene to consist of 19 exons and to span approximately 29 kb. The exon-intron junctions follow the GT/AG rule. The 5' untranslated region is nearly 12 kb in length, and the transcription initiation site is surrounded by CG-rich sequences. The 5' flanking region contains putative binding sites for activator protein 1 (AP-1), nuclear factor kappa B (NFkappaB), stimulatory protein-1 (Sp-1) and nerve growth factor induced-C (NGFI-C) transcription factors. The PKCdelta gene is localized at the rat chromosome 19p14. The cloned gene will help to elucidate the role of PKCdelta in growth, differentiation and death of mammalian cells. PMID- 10721704 TI - Analysis of the 5' end of the mouse Elavl1 (mHuA) gene reveals a transcriptional regulatory element and evidence for conserved genomic organization. AB - mHuA (Elavl1) belongs to a highly conserved family of genes encoding RNA-binding proteins and has been linked to cell growth and proliferation through its regulation of mRNA stability. Here, we use an RNase protection assay to demonstrate that the mHuA transcript is relatively abundant in a range of mouse tissues, with the highest levels being found in lung and embryonic stem cells. We then cloned and mapped an 18 kb DNA fragment which encompasses the 5' end of the mHuA gene. The genomic organization in this region is similar to the neural restricted family members, Hel-N1 (ELAVL2) and mHuD (Elavl4). The first exon is lengthy and untranslated, and the second exon, which includes the methionine start site, ends between the ribonucleoprotein motifs of the first RNA binding domain. Mapping of the mHuA transcript by primer extension demonstrated three potential transcription-initiation sites which were detected consistently among different tissues and cell lines. Analysis of the sequence flanking these sites revealed the presence of transcriptional elements including TATA, CREB, c-ets, and AP1 sites. Transfection analysis of this promoter region using a luciferase reporter-gene assay indicated strong transcriptional activity both in HeLa and in mouse macrophage (RAW) cells which is consistent with the ubiquitous expression pattern of mHuA. Thus, while the genomic organization of mHuA is similar to the neural-restricted members of the Elav family, the promoter element differs substantially both by sequence analysis and transcriptional activity in non neural cell types. PMID- 10721705 TI - The growth of mdp1/rsp5 mutants of Saccharomyces cerevisiae is affected by mutations in the ATP-binding domain of the plasma membrane H+ -ATPase. AB - Mutations in the PMA1 gene, encoding plasma membrane H+ -ATPase, were isolated that are able to suppress the temperature sensitivity (ts) phenotype of mdp1 mutations located in RSP5, the ubiquitin-protein ligase gene. The mdp1 mutants were previously found to change the mitochondrial/cytosolic distribution of Mod5p I, the tRNA modifying enzyme, and to affect fluid phase endocytosis. The data presented reveal that mdp1 mutants are also pH sensitive, and hypersensitive to hygromycin B and paromomycin. The ts phenotype, hygromycin B and paromomycin sensitivity are suppressed by pmal-t, but the pH sensitivity, the effect of mdp1 on Mod5p-I cytoplasmic/mitochondrial localization and endocytosis are not. Characterization of pmal-t revealed the substitution of amino acid G(653)V in the ATP-binding domain of the H+ -ATPase. Our results indicate that Rsp5 ubiquitin protein ligase may also influence, in addition to protein distribution, the functioning of plasma membrane H+ -ATPase and the response of cells to stress. PMID- 10721706 TI - Molecular characterization of KEX1, a kexin-like protease in mouse Pneumocystis carinii. AB - Expression screening of a Pneumocystis carinii-infected mouse lung cDNA library with specific monoclonal antibodies (mAbs) led to the identification of a P. carinii cDNA with extensive homology to subtilisin-like proteases, particularly fungal kexins and mammalian prohormone convertases. The 3.1 kb cDNA contains a single open reading frame encoding 1011 amino acids. Structural similarities to fungal kexins in the deduced primary amino acid sequence include a putative proenzyme domain delineated by a consensus autocatalytic cleavage site (Arg-Glu Lys-Arg), conserved Asp, His, Asn and Ser residues in the putative catalytic domain, a hydrophobic transmembrane spanning domain, and a carboxy-terminal cytoplasmic domain with a conserved tyrosine motif thought to be important for localization of the protease in the endoplasmic reticulum and/or Golgi apparatus. Based on these structural similarities and the classification of P. carinii as a fungus, the protease was named KEX1. Southern blotting of mouse P. carinii chromosomes localized kex1 to a single chromosome of approximately 610 kb. Southern blotting of restriction enzyme digests of genomic DNA from P. carinii infected mouse lung demonstrated that kex1 is a single copy gene. The function of kexins in other fungi suggests that KEX1 may be involved in the post translational processing and maturation of other P. carinii proteins. PMID- 10721707 TI - cDNA cloning of pig testicular lactate dehydrogenase-C, thermal stability of the expressed enzyme, and polymorphism among strains. AB - Pig testicular lactate dehydrogenase-C (LDHC) cDNA was cloned and sequenced. The deduced sequence of 332 amino acids from pig LDHC shows 73% and 67% identity with that of pig LDHA (muscle) and LDHB (heart) respectively, whereas pig LDHA and LDHB isozymes shows 74% sequence identity. Pig and mouse LDHC cDNAs were subcloned into bacterial expression vector, and the expressed pig LDHC isozyme was shown to be as thermally stable as mouse LDHC isozyme. Pig genomic DNAs from Chinese Meishan, English Yorkshire, Danish Landrace and American Duroc were shown to exhibit polymorphic sites for restriction enzymes EcoRI, BamHI and PstI. PMID- 10721708 TI - The genetic switch for the regulatory pathway of Lactobacillus plantarum phage (phi)g1e: characterization of the promoter P(L), the repressor gene cpg, and the cpg-encoded protein Cpg in Escherichia coli. AB - The structural and functional features of the approximately 530 bp P(L)/Gb5-Gb6 cpg-Gb7 region (P(L) overlaps Gb5) for the lysogenic pathway of L. plantarum phage (phi)gle were investigated using the cat gene of E. coli plasmid pKK232-8 as a reporter. In E. coli XL1-Blue, a recombinant plasmid pKPL2 (cat under P(L)/Gb5-Gb6) exhibited distinct CAT activity, whereas the activity of pKPLCP1 (cat under P(L)/Gb5-Gb6-cpg) was only marginal. When pKPL2 was coexistent with a compatible derivative of plasmid pACYC177 carrying P(L)/Gb5-Gb6-cpg, the CAT activity was declined to the level of pKPLCP1. On the other hand, the cpg-encoded protein Cpg was overproduced in E. coli under P(T7). The molecular mass of the purified Cpg (14.5 kDa on a SDS gel) corresponded well with that (15.1 kDa) predicted from the DNA sequence. Gel-shift and footprinting assays demonstrated that Cpg selectively binds to about 25 bp bases centered around the GATAC-box (from 1 to 7). Moreover, protein crosslinking experiments using glutaraldehyde showed that Cpg most likely functions as a dimeric form. Thus, the present results indicate that Cpg probably represses P(L) through binding to the operator GATAC-box(es), and the P(L)/cpg region might participate in the lysogenic pathway. PMID- 10721709 TI - Detailed structural analysis on both human MRP5 and mouse mrp5 transcripts. AB - The multidrug-resistant phenotype in tumor cells is attributed in part to anti cancer drug efflux transporters such as the MRP family. The amino-terminal structure of MRP5 has not been refined. To determine the amino-terminal structure of a major transcript of the MRP5 gene, we performed primer extension analysis to determine a major transcriptional start site of this gene and compared the structure of human MRP5 and that of mouse mrp5. We successfully determined the structures of human MRP5 and mouse mrp5. Estimated amino acid sequences are 1437 and 1436 amino acids for human MRP5 and mouse mrp5 respectively, and were highly conserved (94.1%). We further showed that our previously identified SMRP mRNA was a splicing variant of the MRP5 gene, which was expressed in various human tissues, suggesting that a short form of MRP5 protein encoded by the SMRP mRNA may have a physiological role. PMID- 10721710 TI - Collapsin response mediator protein-3/unc-33-like protein-4 gene: organization, chromosomal mapping and expression in the developing mouse brain. AB - CRMPs (collapsin response mediator proteins)/ULIPs (unc-33-like proteins) are a family of intracytoplasmic proteins that are expressed mainly in the brain. The involvement of CRMP/ULIP members in neuronal differentiation, growth cone motility and axonal collapse has been suggested. We recently found that a member of this family, CRMP3/ULIP4, corresponds to POP66 (paraneoplastic oligodendrocyte protein of 66 kDa), a protein which may be associated with auto-immune induced neuronal degeneration in paraneoplastic neurological syndromes. However, the physiological functions of these proteins remain to be elucidated. Further studies, including the generation of cell lines and of animals with modified/disrupted CRMP/ULIP gene expression, are necessary to explore the functions of this protein. We have cloned and determined the organization and chromosomal localization of the mouse gene encoding CRMP3/ULIP4. The gene is composed of 14 exons and spans more than 20 kb. We assigned the mouse CRMP3/ULIP4 gene to the distal end of chromosome 7. In mouse brain, in situ hybridization showed that CRMP3/ULIP4 mRNA is expressed mainly in the dentate gyrus of hippocampus, in the granular layers of cerebellum and in the inferior olive of the pons, the nucleus which controls movement and posture, and adjusts the major output of descending motor system. PMID- 10721711 TI - Fission yeast contains an rDNA binding activity that interacts specifically with regulatory sequences for ribosomal RNA synthesis. AB - Basal level transcriptional initiation of fission yeast ribosomal RNA genes is dependent on the core ribosomal RNA gene promoter and is stimulated by an upstream rDNA promoter element and by regulatory sequences located in its approximately 3.5 kb intergenic rDNA spacer. A Schizosaccharomyces pombe sequence specific rDNA binding activity was characterized that interacted with the upstream rDNA promoter region and that associated with required RNA polymerase I transcription components in initial fractionation steps. The rDNA binding activity was further purified and found to specifically associate with a region of the rDNA promoter between -80 and -56. The promoter region required for stable binding correlates with that mediating activated levels of transcriptional initiation. This rDNA binding activity stimulates in vitro rRNA synthesis supported by templates bearing this upstream promoter domain but not by templates lacking it. PMID- 10721712 TI - Cloning and characterization of a novel histone acetyltransferase homologue from the protozoan parasite Toxoplasma gondii reveals a distinct GCN5 family member. AB - In an effort to identify gene products involved in transcriptional regulation in apicomplexan parasites, the Toxoplasma gondii expressed sequence tag (EST) database was examined for sequences containing similarity to known transcriptional components. One EST (dbEST ID #466792) exhibited strong similarity to yeast GCN5 and other histone acetyltransferases (HATs). Primers were designed based on the EST sequence and used to amplify an 850 bp fragment (containing an intron) from T. gondii genomic DNA which was used to identify four cDNA clones from a tachyzoite cDNA library. The complete open reading frame (ORF) of 3.5 kb was elucidated using 5' RACE and genomic sequence. The deduced amino acid sequence of the coding region shows that the C-terminal domain possesses unequivocal similarity to GCN5 family members. However, unlike other lower eukaryotes, T. gondii GCN5 has an extended N-terminal domain similar in length, but not in composition, to metazoan HAT proteins. These features distinguish T. gondii GCN5 as a novel member of the GCN5 family. A portion of the cDNA sequence was used as a probe to isolate three overlapping clones from a T. gondii genomic library, generating a approximately 7.5 kb map of the GCN5 locus which contains seven exons separated by six introns. Southern analysis verifies the predicted map and suggests that a similar locus may be present elsewhere in the genome. PMID- 10721713 TI - Green fluorescent protein as a second selectable marker for selection of high producing clones from transfected CHO cells. AB - Mammalian cells are often used for the expression of recombinant proteins. The process of screening transfected cells randomly for high producing clones is tedious and time consuming. We evaluated using green fluorescent protein (GFP) for selection of high producing clones by fluorescence-activated cell sorter (FACS) to reduce screening effort. We expressed neurotrophin-3 (NT3), deoxyribonuclease (DNase), or vascular endothelial growth factor (VEGF) with GFP in Chinese hamster ovary cells. The vector expressed the desired secreted protein and the selectable marker, dihydrofolate reductase, in one expression unit and the intracellular GFP in a second expression unit. Transfected cells were grown in selection medium and sorted by FACS. High fluorescence clones were obtained and found to produce high amounts of the desired protein; VEGF productivity correlated well with GFP fluorescence in 48 clones. Further studies demonstrated that productivity correlated very well with RNA of the desired protein. For comparison, we randomly picked and screened 144 VEGF clones, and the highest producing VEGF clone obtained produced 0.7 pg/cell/day. In contrast, the highest producing VEGF clone obtained by FACS sorting produced 4.4 pg/cell/day. FACS sorting therefore selected high producing clones efficiently. Since an assay for the desired protein is not required, high producing clones for a protein of unknown function can be obtained by FACS sorting followed by measuring the RNA level of the desired protein in the highly fluorescent clones. PMID- 10721714 TI - The rat Mist1 gene: structure and promoter characterization. AB - Transcription factors of the basic Helix-Loop-Helix (bHLH) protein family play key roles in several developmental processes. Mist1 belongs to this group of proteins and shares several properties with the other family members. For example, Mist1 is capable of dimerization with the ubiquitously expressed E2A bHLH proteins and exhibits a strong DNA-binding activity to the core E-box sequence. Using in-situ hybridization and Northern blot hybridization, Mist1 mRNA has been detected in a variety of embryonic and adult rodent tissues. To understand the molecular mechanisms involved in the expression of the gene, we have cloned the rat Mist1 gene and analyzed 2.5 kb of its 5' flanking region. The Mist1 gene spans over 5 kilobases and is composed of two exons separated by a unique intron. The entire coding region is localized in the second exon. Sequence analysis of the promoter region indicated an absence of TATA-box or CAAT-box sequence, but several consensus Sp1-binding sites were present near the transcription start site. Deletion analysis of the promoter region identified a 272 bp proximal fragment to be sufficient to drive expression of a reporter gene in NIH3T3 fibroblasts. Subsequent deletion of potential Sp1 sites results in a marked decrease in promoter activity. Electrophoretic mobility shift assays revealed that Sp1 binds to two different regions in the proximal promoter, a typical Sp1 site located at (-38; -33) and a G/C-rich region between (-67; -62). These data suggest that the basal expression of this TATA-less gene might be driven by general transcription factors, such as Sp1. PMID- 10721715 TI - Activation of human gamma-globin gene expression via triplex-forming oligonucleotide (TFO)-directed mutations in the gamma-globin gene 5' flanking region. AB - Human beta-globin disorders, such as sickle cell anemia and beta-thalassemia, are relatively common genetic diseases cause by mutations in the beta-globin gene. Increasing gamma-globin gene expression has been found to greatly reduce the disease symptom. However, the gamma-globin gene is developmentally regulated and normally expressed at high levels only during the fetal stage of human development. We have explored the possibility of activating the gamma-globin gene expression by triplex-forming oligonucleotide (TFO)-directed targeted mutagenesis. Using a psoralen-conjugated TFO designed to bind to a site overlapping with an Oct-1 binding site at the -280 region of the gamma-globin gene, targeted mutagenesis of the Oct-1 binding site has been achieved by transfecting the in-vitro-formed plasmid-oligo complex into human normal fibroblast (NF) cells. The mutation frequency at the target site was estimated to be 20% by direct DNA sequencing analysis. In-vitro protein binding assays indicated that these mutations reduced Oct-1 binding to the target site. In-vivo gene expression assays demonstrated activation of gamma-globin gene expression from these mutations in mouse erythroleukemia (MEL) cells. The levels of the gamma-globin gene expression increased by as much as fourfold in mutants with single base changes. These results suggest that the -280 region of the Agamma globin gene negatively regulates the gamma-globin gene expression, and mutations at the Oct-1 binding site can lead to activation of the gamma-globin gene and generate the hereditary persistence of fetal hemoglobin (HPFH) condition. This study may provide a novel approach for gene therapy of sickle cell disease. The data may also have implications in gene therapy for other diseases including genetic diseases and cancers by introducing mutations into transcription factor binding sites to alter the levels of target gene expression. PMID- 10721716 TI - Structure and chromosomal localization of the human glycogenin-2 gene GYG2. AB - Glycogenin-2 is one of two self-glucosylating proteins involved in the initiation phase of the synthesis of the storage polysaccharide glycogen. Cloning of the human glycogenin-2 gene, GYG2, has revealed the presence of 11 exons and a gene of more than 46 kb in size. The structure of the gene explains much of the observed diversity in glycogenin-2 cDNA sequences as being due to alternate exon usage. In some cases, there is variation in the splice junctions used. Over regions of protein sequence similarity, the GYG2 gene structure is similar to that of the other glycogenin gene, GYG. A genomic GYG2 clone was used to localize the gene to Xp22.3 by fluorescence in-situ hybridization. Localization close to the telomere of the short arm of the X chromosome is consistent with mapping information obtained from glycogenin-2 STS sequences. Glycogenin-2 maps between the microsatellite anchor markers AFM319te9 (DXS7100) and AFM205tf2 (DXS1060), and its 3' end is 34.5 kb from the 3' end of the arylsulphatase gene ARSD. GYG2 is outside the pseudoautosomal region PAR1 but still in a region of X-Y shared genes. As is true for several other genes in this location, an inactive remnant of GYG2, consisting of exons 1-3, may be present on the Y chromosome. PMID- 10721717 TI - Isolation, characterization, and mapping of the mouse Fgd3 gene, a new Faciogenital Dysplasia (FGD1; Aarskog Syndrome) gene homologue. AB - FGD1 gene mutations result in faciogenital dysplasia (FGDY, Aarskog syndrome), an X-linked developmental disorder that adversely affects the formation of multiple skeletal structures. FGD1 encodes a guanine nucleotide exchange factor (GEF) that specifically activates the Rho GTPase Cdc42. By way of Cdc42, FGD1 regulates the actin cytoskeleton and activates the c-Jun N-terminal kinase signaling cascade to regulate cell growth and differentiation. Previous work shows that FGD1 is the founding member of a family of related genes including the mouse Fgd2 gene and the rat Frabin gene. Here, we report on the isolation, characterization, and mapping of the mouse Fgd3 gene, a new and novel member of the FGD1 gene family. Fgd3 cDNA encodes a 733-amino-acid protein with a predicted mass of 81 kDa. Fgd3 and FGD1 share a high degree of sequence identity that spans >560 contiguous amino acid residues. Like FGD1, Fgd3 contains adjacent RhoGEF and pleckstrin homology (PH) domains, a second carboxy-terminal PH domain, and a distinctive FYVE domain. Together, these domains appear to form a canonical core structure for FGD1 family members. In addition, compared to other FGD1 family members, Fgd3 contains different structural regions that may be involved in distinct signaling interactions. Microinjection studies show that Fgd3 stimulates fibroblasts to form filopodia, actin microspikes formed upon the stimulation of Cdc42. Fgd3 transcripts are present in several diverse tissues and during mouse embryogenesis, suggesting a developmentally regulated pattern of expression and a potential role in embryonic development. Genetic linkage and radiation hybrid mapping data show that Fgd3 and the human FGD3 ortholog map to syntenic regions of murine chromosome 13 and human chromosome 9q22, respectively. We conclude that Fgd3 is a new and novel member of the FGD1 family of RhoGEF proteins. PMID- 10721718 TI - Positive tetracycline control of expression of p15INK4B from an Epstein-Barr autonomous plasmid in a human melanoma cell line. AB - Homozygous deletions in the region of chromosome 9p21 are frequent in human melanoma. Mutations in the p16INK4A cyclin-dependent kinase inhibitor (CDI) gene at this locus have implicated the product of this gene as a tumor suppressor. Less attention has been focused on the homologous, closely linked p15INK4B gene. To facilitate study of the phenotypic effects of restoring expression of the latter in aggressive melanoma cells lacking INK4 expression, we inserted the cDNA encoding p15INK4B into an autonomously maintained plasmid under positive tetracycline control ('TET ON' system). Similarly regulated luciferase and herpes thymidine kinase sequences were used as controls. We demonstrate that this system enabled efficient, and reasonably uniform, induction of p15INK4B expression in a human melanoma cell line exposed to the tetracycline derivative, doxycycline. Flow cytometry showed that this induction resulted in substantial accumulation of cells in the G0/G1 phase of the cell cycle. This system will facilitate detailed analysis of the cell cycle inhibitory mechanisms of this CDI in human melanoma cells. PMID- 10721719 TI - Two promoters regulate transcription of the mouse folylpolyglutamate synthetase gene three tightly clustered Sp1 sites within the first intron markedly enhance activity of promoter B. AB - The process of polyglutamylation mediated by folylpolyglutamate synthetase (FPGS) in mammalian cells has nutritional and pharmacological importance. In murine cells, FPGS expression is controlled by two promoters that, as we show here, vary substantially in their efficiency, at least in the context of a reporter gene assay. Characteristics of the most efficient promoter (promoter B) were examined in the present studies. Insertion in pGL3 of a 1635 bp segment of upstream sequence including the most upstream exon (B1c), intron B1c and only 26 bp of the more downstream exon Bla resulted in a 15-20-fold increase in transcription in NIH3T3 and Hep1-6 cells compared with the promoterless vector. Deletion analysis of DNA sequence upstream of exon B1c showed that transcription was regulated by putative cis active elements only within two distally located upstream segments which when deleted cumulatively increased transcription three- to four-fold. However, deletion of the 56 bp intron B1c immediately downstream of the most upstream exon (Blc) resulted in 1/10 the rate of transcription. Primer extension analysis with NIH3T3 cells revealed start sites for transcription appreciably upstream of and within exon B1c as well as downstream in exon B1a. This result is consistent with the frequent occurrence in murine cells of an FPGS variant (variant III) incorporating exon B1c [Roy et al., J. Biol. Chem. 271 (1996) 23820; 272 (1997) 5587]. Site-directed mutagenesis and DNAse I footprinting revealed that three canonical GC boxes, either overlapping or tightly clustered within intron B1c, bound Sp1 and markedly enhanced transcription, accounting for the maximal promoter B activity. Moreover, in a cellular background devoid of Sp1 activity, we demonstrate that Spl can induce high levels of promoter B activity in pGL3 transfectants, but only when intron B1c is included within the reporter gene construct used. These results suggest that the unusually tight cluster of active Sp1 sites within intron B1c are essential and sufficient for maximal activity of this promoter. These tightly clustered sites appear to act as an enhancer element in promoting transcription and efficiently stabilize transcription initiation complexes at both distal and proximal start sites. PMID- 10721720 TI - Lens epithelium-derived growth factor (LEDGF/p75) and p52 are derived from a single gene by alternative splicing. AB - A human gene that encodes lens epithelium-derived growth factor (LEDGF) was isolated, and the DNA sequence and the exon/intron organization was determined. The gene contains at least 15 exons and 14 introns and encodes LEDGF mRNA and p52 mRNA. Exons 1-15 encode LEDGF mRNA, and exons 1-9, and a part of the ninth intron encode a splice variant (p52). Sequences of the exon/intron junctions of the gene have the highly conserved GT/AG rule. Most intron/exon junctions correspond to junctions of individual protein motifs. Almost equal amounts of LEDGF and p52 are expressed in lens epithelial cells in culture. The LEDGF gene is assigned to chromosome 9p22.2, which is adjacent to the major cell malignancy locus. PMID- 10721721 TI - Polypurine.polypyrimidine sequences in complete bacterial genomes: preference for polypurines in protein-coding regions. AB - The genomes of Methanococcus jannaschii, Mycoplasma genitalium, Haemophilus influenzae, Archaeoglobus fulgidus, Helicobacter pylori, Treponema pallidum, Borrelia burgdorferri, Rickettsia prowazekeii, Mycobacterium tuberculosis, Methanobacterium thermoautotrophicum, Synechocystis sp. PCC6803, Bacillus subtilis, Chlamydia trachomatis, Pyrococcus horikoshii, Aquifex aeolicus, Mycoplasma pneumoniae and Escherichia coli have been analysed for the presence of polypurine.polypyrimidine tracts, in order to understand their distribution in these genomes. We observed a variation in abundance of such sequences in these bacteria, with the archaeal genomes forming a high-abundance group and the canonical eubacteria forming a low-abundance group. The genomes of M. tuberculosis and A. aeolicus are unique among the organisms analysed here in the abnormal underrepresentation and overrepresentation of polypurine.polypyrimidine, respectively. We also observe a strand bias, i.e., a preferential occurrence of polypurines in coding strands. It varies widely among the bacteria, from the very high bias in M. jannaschii to the slightly inverse bias in the parasitic genomes of T. pallidum and C. trachomatis. The extent of strand bias, however, cannot be explained on the basis of the GC-content of the genome, use of all-purine codons or an excess in the amino acids that are encoded by such codons. The probable causes and effects of this phenomenon are discussed. PMID- 10721722 TI - Cloning of three novel neuronal Cdk5 activator binding proteins. AB - Neuronal Cdc2-like kinase (Nclk) is involved in the regulation of neuronal differentiation and neuro-cytoskeleton dynamics. The active kinase consists of a catalytic subunit, Cdk5, and a 25 kDa activator protein (p25nck5a) derived from a 35 kDa neuronal-specific protein (p35nck5a). As an extension of our previous study (Qi, Z., Tang, D., Zhu, X., Fujita, D.J., Wang, J.H., 1998. Association of neurofilament proteins with neuronal Cdk5 activator. J. Biol. Chem. 270, 2329 2335), which showed that neurofilament is one of the p35nck5a-associated proteins, we now report the isolation of three other novel p35nck5a-associated proteins using the yeast two-hybrid screen. The full-length forms of these three novel proteins, designated C42, C48 and C53, have a molecular mass of 66, 24, and 57 kDa, respectively. Northern analysis indicates that these novel proteins are widely expressed in human tissues, including the heart, brain, skeletal muscle, placenta, lung, liver, kidney and pancreas. The bacterially expressed glutathione S-transferase (GST)-fusion forms of these three proteins were able to co precipitate p35nck5a complexed with Cdk5 from insect cell lysate. Among these three proteins, only C48 and C53 can be phosphorylated by Nclk, suggesting that they may be the substrates of Nclk. Sequence homology searches have suggested that the C48 protein is marginally related to restin protein, whereas the C42 protein has homologues of unknown function in Caenorhabditis elegans and Arabidopsis thaliana. PMID- 10721724 TI - Compositional pressure and translational selection determine codon usage in the extremely GC-poor unicellular eukaryote Entamoeba histolytica. AB - It is widely accepted that the compositional pressure is the only factor shaping codon usage in unicellular species displaying extremely biased genomic compositions. This seems to be the case in the prokaryotes Mycoplasma capricolum, Rickettsia prowasekii and Borrelia burgdorferi (GC-poor), and in Micrococcus luteus (GC-rich). However, in the GC-poor unicellular eukaryotes Dictyostelium discoideum and Plasmodium falciparum, there is evidence that selection, acting at the level of translation, influences codon choices. This is a twofold intriguing finding, since (1) the genomic GC levels of the above mentioned eukaryotes are lower than the GC% of any studied bacteria, and (2) bacteria usually have larger effective population sizes than eukaryotes, and hence natural selection is expected to overcome more efficiently the randomizing effects of genetic drift among prokaryotes than among eukaryotes. In order to gain a new insight about this problem, we analysed the patterns of codon preferences of the nuclear genes of Entamoeba histolytica, a unicellular eukaryote characterised by an extremely AT-rich genome (GC = 25%). The overall codon usage is strongly biased towards A and T in the third codon positions, and among the presumed highly expressed sequences, there is an increased relative usage of a subset of codons, many of which are C-ending. Since an increase in C in third codon positions is 'against' the compositional bias, we conclude that codon usage in E. histolytica, as happens in D. discoideum and P. falciparum, is the result of an equilibrium between compositional pressure and selection. These findings raise the question of why strongly compositionally biased eukaryotic cells may be more sensitive to the (presumed) slight differences among synonymous codons than compositionally biased bacteria. PMID- 10721723 TI - Woodchuck lymphotoxin-alpha, -beta and tumor necrosis factor genes: structure, characterization and biological activity. AB - We cloned and characterized the woodchuck tumor necrosis factor (TNF) and lymphotoxin-alpha, -beta (LT-alpha, -beta) cDNAs, genes and proteins to facilitate study of the functions of these cytokines during the course of woodchuck hepatitis virus (WHV) infection. Woodchuck cDNA and genomic DNA libraries were screened with woodchuck-specific DNA probes to isolate the cDNA and gene clones for TNF, LT-alpha and LT-beta. The cDNAs for woodchuck TNF, LT alpha and LT-beta code for proteins of 233, 205 and 310 amino acids respectively. The polypeptide encoded by each gene among woodchucks, humans and mice can differ: the human TNF, LT-alpha and LT-beta genes encode polypeptides of 233, 205 and 244 amino acids respectively, whereas the mouse TNF, LT-alpha and LT-beta genes encode polypeptides of 235, 202 and 306 amino acids respectively. In the woodchuck, there are four exons for TNF, four exons for LT-alpha and three exons for LT-beta. The RNA splicing patterns for TNF, LT-alpha and LT-beta genes are identical among woodchucks, humans and mice, except that the human LT-beta gene contains four exons. The woodchuck TNF gene promoter contains consensus sequences for binding of AP-1, AP-2, C/EBPbeta, CRE, Egr-1, Ets, NF-AT, NF-kappaB and SP-1 transcription factors. LT-alpha has AP-2, Ets, NF-kappaB, SP-1 and STAT binding sites, and LT-beta has Egr-1/SP-1, Ets and NF-kappaB binding sites. The bacterially expressed woodchuck TNF and LT-alpha proteins exhibited cytotoxic activities on both mouse L929B and woodchuck A2 cells in the presence of actinomycin D. The specific activities of TNF and LT-alpha were 2.62x10(8) units/mg and 2.22x10(3) units/mg respectively for L929B cells, and 1.05x10(9) units/mg and 3.56x10(4) units/mg respectively for A2 cells. However, only woodchuck TNF showed cytotoxic activity on human HepG2 cells, with a specific activity of 6.55x10(7) units/mg in the presence of actinomycin D. The data obtained from this study will be useful to future investigations of the TNF and LT antitumor and anti-viral activities, and their therapeutic potential in the woodchuck model for human hepatitis B virus (HBV). PMID- 10721725 TI - Structural characterization of the human fast skeletal muscle troponin I gene (TNNI2). AB - Three troponin I genes have been identified in vertebrates that encode the isoforms expressed in adult cardiac muscle (TNNI3), slow skeletal muscle (TNNI1) and fast skeletal muscle (TNNI2), respectively. While the organization and regulation of human cardiac and slow skeletal muscle genes have been investigated in detail, the fast skeletal troponin I gene has to date only been examined in birds. Here, we describe the structure and complete sequence of the human fast skeletal muscle troponin I gene (TNNI2) and identify putative regulatory elements within both the 5' flanking region and the first intron. In particular, a region containing MEF-2, E-box, CCAC and CAGG elements was identified in intron 1 that closely resembles the fast internal regulatory element (FIRE) of the quail intronic enhancer. We have previously shown that the fast skeletal muscle troponin I gene is located at 11p15.5 and noted potential close linkage with the fast skeletal muscle troponin T gene (TNNT3). Here, we have isolated two independent human PAC genomic clones that contain either TNNI2 or TNNT3 and demonstrate by interphase FISH mapping that they are less than 100 kb apart in the genome. The results demonstrate that the human TNNI2 gene is closely related to its avian counterparts with conserved elements within both the putative promoter and first intron. Our data further confirm close physical linkage of TNNI2 and TNNI3 on 11p15.5. PMID- 10721726 TI - Molecular cloning of mouse thioredoxin reductases. AB - We screened clones for thioredoxin reductase genes with a degenerate PCR-based strategy and have isolated two novel cDNA clones from a mouse thymocyte cDNA library. These encode two distinct thioredoxin reductases (TrxR1 and TrxR2) with 499 and 527 amino acid (aa) residues and calculated molecular masses of 54.5 kDa and 56.8 kDa respectively. These proteins share 90% and 50% aa sequence identity with those of previously cloned human TrxR, containing the redox-active cysteines, FAD binding domain, and the selenocysteine (SeCys) insertion sequence, which is composed of a putative stem-loop sequence located in the 3'-untranslated region (UTR). TrxR2 showing less homology to human TrxR has a mitochondrial translocation signal and a mitochondrial prepeptide protease cleavage site in the N-terminal domain. Transient expression experiments of each gene as fusion proteins with Xpress-tagged protein in NIH 3T3 cells indicated that TrxR1 was localized in the nucleus and cytoplasm and TrxR2 in the mitochondria. Furthermore, we mapped the TrxR1 gene to chromosome 10 (placed 1.71 cR from D10Mit42, lod>3.0) and the TrxR2 gene to chromosome 16 (placed 22.56 cR from D16Mit34, lod>3.0). Thus, the mouse has at least two distinct nuclear genes for TrxR that have different translocation sites in the cell. PMID- 10721727 TI - Complete nucleotide sequence of a plant tumor-inducing Ti plasmid. AB - Crown gall tumor disease in dicot plants is caused by Agrobacterium tumefaciens harboring a giant tumor-inducing (Ti) plasmid. Here, for the first time among agrobacterial plasmids, the nucleotide sequence of a typical nopaline-type Ti plasmid (pTi-SAKURA) was determined completely. In total, 195 open reading frames (ORFs) were estimated in the 206479 bp long sequence. 20 genes for conjugation, three for replication, 22 for pathogenesis and 37 for genetic colonization of host plants were found within two-thirds of the plasmid. These genes formed seven functional gene clusters with narrow inter-cluster spaces. In the remaining one third of the plasmid, novel genes including homologs of mutT, Rhizobium nodQ and Sphingomonas ligE genes were found, which are likely to be responsible for the broad host range. Restriction fragment length variation indicates extreme plasticity of the part required for conjugational gene transfer and the above mentioned one-third of the plasmid, even among closely related Ti plasmids. PMID- 10721729 TI - The development of TnNuc and its use for the isolation of novel secretion signals in Lactococcus lactis. AB - We have previously used Tn917 for the identification and characterization of regulated promoters from Lactococcus lactis [Israelsen et al., Appl. Environ. Microbiol. 61 (1995) 2540-2547]. We describe here the construction of a new Tn917 transposon derivative, termed TnNuc, which includes the Staphylococcus aureus nuclease gene (nuc) as a reporter for secretion. Transposition of TnNuc into the L. lactis chromosome allows the generation of fusions in-frame with the nuc gene. TnNuc includes also lacZ, a reporter used for identification of relevant clones from the library, i.e. clones with Lac+ phenotype result from transposition of TnNuc into a functional gene on the L. lactis chromosome. The presence of a functional signal sequence at the upstream flanking region of the left repeat of the transposed element results in the detection of nuclease activity using a sensitive plate assay. TnNuc was used for the identification of novel secretion signals from L. lactis. The sequences identified included known and unknown lactococcal-secreted proteins containing either a signal peptidase-I or -II recognition sequence. In one case, the gene identified codes for a transmembrane protein. The sequences identified were used to study functionality when located in a plasmid under the control of the pH and growth phase-dependent promoter P170 [Madsen et al., Mol. Microbiol. 32 (1999) 75-87]. In all cases, concurrent secretion of nuclease was observed during induction of P170 in a fermentor. PMID- 10721728 TI - Ermap, a gene coding for a novel erythroid specific adhesion/receptor membrane protein. AB - Ermap (erythroid membrane-associated protein), a gene coding for a novel transmembrane protein produced exclusively in erythroid cells, is described. It is mapped to murine Chromosome 4, 57 cM distal to the centromere. The initial cDNA clone was isolated from a day 9 murine embryonic erythroid cell cDNA library. The predicted peptide sequence suggests that ERMAP is a transmembrane protein with two extracellular immunoglobulin folds, as well as a highly conserved B30.2 domain and several phosphorylation consensus sequences in the cytoplasmic region. ERMAP shares a high homology throughout the entire peptide with butyrophilin, a glycoprotein essential for milk lipid droplet formation and release. A GFP-ERMAP fusion protein was localized to the plasma membrane and cytoplasmic vesicles in transiently transfected 293T cells. Northern blot analysis and in-situ hybridization demonstrated that Ermap expression was restricted to fetal and adult erythroid tissues. ERMAP is likely a novel adhesion/receptor molecule specific for erythroid cells. PMID- 10721730 TI - The ftsZ gene of Haloferax mediterranei: sequence, conserved gene order, and visualization of the FtsZ ring. AB - We sequenced the ftsZ gene region of the halophilic archaeon Haloferax mediterranei and mapped the transcription start sites for the ftsZ gene. The gene encoded a 363-amino-acid long FtsZ protein with a predicted molecular mass of 38 kDa and an isoelectric point of 4.2. A high level of similarity to the FtsZ protein of Haloferax volcanii was apparent, with 97 and 90% identity at the amino acid and nucleotide levels, respectively. Structural conservation at the protein level was shown by visualization of the FtsZ ring structure in H. mediterranei cells using an antiserum raised against FtsZ of H. volcanii. FtsZ rings were observed in cells in different stages of division, including cells with pleomorphic shapes and cells that appeared to be undergoing asymmetric division. Cells were also observed that displayed constriction-like invaginations in the absence of an FtsZ ring, indicating that morphological data are not sufficient to determine whether pleomorphic Haloferax cells are undergoing cell division. Both the upstream and downstream gene order in the ftsZ region was found to be conserved within the genus Haloferax. Furthermore, the downstream gene order, which includes the secE and nusG genes, is conserved in almost all euryarchaea sequenced to date. The secE and nusG genes are likely to be transcriptionally and translationally coupled in Haloferax, and this co-expression may have been a selective force that has contributed to keeping the gene cluster intact. PMID- 10721731 TI - A genomic approach of the hepatitis C virus generates a protein interaction map. AB - The hepatitis C virus (HCV) causes severe liver disease, including liver cancer. A vaccine preventing HCV infection has not yet been developed, and, given the increasing number of infected people, this virus is now considered a major public health problem. The HCV genome is a plus-stranded RNA that encodes a single polyprotein processed into at least 10 mature polypeptides. So far, only the interaction between the protease NS3 and its cofactor, NS4A, which is involved in the processing of the non-structural region, has been extensively studied. Our work was aimed at constructing a protein interaction map of HCV. A classical two hybrid system failed to detect any interactions between mature HCV polypeptides, suggesting incorrect folding, expression or targetting of these proteins. We therefore developed a two-hybrid strategy, based on exhaustive screens of a random genomic HCV library. Using this method, we found known interactions, such as the capsid homodimer and the protease dimer, NS3-NS4A, as well as several novel interactions such as NS4A-NS2. Thus, our results are consistent with the idea that the use of a random genomic HCV library allows the selection of correctly folded viral protein fragments. Interacting domains of the viral polyprotein are identified, opening the possibility of developing specific anti viral agents, based on their ability to modulate these interactions. PMID- 10721732 TI - PCR-based gene targeting in the filamentous fungus Ashbya gossypii. AB - We have investigated a PCR-based approach for one-step gene targeting in the filamentous fungus Ashbya gossypii. Short guide sequences with 40-46 bp of homology to two sequences of a targeted gene, provided by PCR, were sufficient to mediate homologous recombination. The PCR products used for transformation were generated from the newly constructed chimeric selection marker GEN3. This consists of the open reading frame of the Escherichia coli kanR gene under the control of promoter and terminator sequences of the Saccharomyces cerevisiae TEF2 gene and allows selection of G418/geneticin-resistant transformants. Verification of gene targeting was performed either by PCR or by DNA hybridization analyses, and in all 18 cases tested, correct targeting was confirmed. This approach was used for the complete deletion of the open reading frame of the A. gossypii RHO4 gene for which a double-strand sequence was available as information source for the design of PCR primers. We also demonstrated successful partial deletion of four other ORFs using single-read sequences (SRS) as sole information for the design of targeting primers. A gossypii is the first filamentous fungus in which a PCR-based gene disruption technique has been established. Since short target guide sequences are sufficient to direct homologous integration into the A. gossypii genome it is not necessary to obtain and sequence large DNA fragments from a target locus to provide the long flanking homology regions usually required for efficient targeting of cloned disruption cassettes in filamentous fungi. Thus functional analysis of A. gossypii genes is already possible, based on single-pass sequence information. PMID- 10721733 TI - A molecular genetic system for the pathogenic yeast Candida dubliniensis. AB - Candida dubliniensis is a recently described pathogenic yeast of the genus Candida that is closely related to Candida albicans but differs from it in several phenotypic and genotypic characteristics, including putative virulence traits, which may explain differences in the spectrum of diseases caused by the two species. In contrast to C. albicans, a molecular genetic system to study virulence of C. dubliniensis is lacking. We have developed a system for the genetic transformation of C. dubliniensis that is based on the use of the dominant selection marker MPA(R) from C. albicans that confers resistance to mycophenolic acid (MPA). Using this transformation system, a GFP (green fluorescent protein) reporter gene that was genetically engineered for functional expression in C. albicans and placed under control of the inducible C. albicans SAP2 (secreted aspartic proteinase) promoter was integrated into the C. dubliniensis genome. MPA-resistant transformants containing the SAP2P-GFP fusion fluoresced under SAP2-inducing conditions but not under SAP2-repressing conditions. These results demonstrate that the MPA(R) selection marker is useful for transformation of C. dubliniensis wild-type strains, that the GFP reporter gene is functionally expressed in C. dubliniensis, and that the C. albicans SAP2 promoter can be used for controlled gene expression in C. dubliniensis. These genetic tools will allow the dissection of the differences in virulence characteristics between the two pathogenic yeast species at the molecular level. PMID- 10721734 TI - Molecular cloning and expression analysis of a cDNA encoding a glutamate transporter in the honeybee brain. AB - We have cloned and characterized a cDNA encoding a putative glutamate transporter, Am-EAAT, from the brain of the honeybee, Apis mellifera. The 543 amino-acid AmEAAT gene product shares the highest sequence identity (54%) with the human EAAT2 subtype. Am-EAAT is expressed predominantly in the brain, and its transcripts are abundant in the optic lobes and inner compact Kenyon cells of the mushroom bodies (MBs), with most other regions of the brain showing lower levels of Am-EAAT expression. High levels of Am-EAAT message are found in pupal stages, possibly indicating a role for glutamate in the developing brain. PMID- 10721735 TI - Transcriptional regulation of the human DNA Methyltransferase (dnmt1) gene. AB - DNA methylation is an important component of the epigenetic control of genome functions. Understanding the regulation of the DNA Methyltransferase (dnmt1) gene expression is critical for comprehending how DNA methylation is coordinated with other critical biological processes. In this paper, we investigate the transcriptional regulatory region of the human dnmt1 gene using a combination of RACE, RNase protection analysis and CAT assays. We identified one major and three minor transcription initiation sites in vivo (P1-P4), which are regulated by independent enhancers and promoter sequences. The minimal promoter elements of P1, P2 and P4 are mapped within 256 bp upstream of their respective transcription initiation sites. P1 is nested within a CG-rich area, similar to other housekeeping genes, whereas P2-P4 are found in CG-poor areas. Three c-Jun dependent enhancers are located downstream to P1 and upstream to P2-P4, thus providing a molecular explanation for the responsiveness of dnmt1 to oncogenic signals that are mediated by the Ras-c-Jun oncogenic signaling pathway. PMID- 10721736 TI - Molecular cloning and functional characterization of the mouse mafB gene. AB - The Maf family of the transcription factors plays a pivotal role in controlling development and cellular differentiation. To clarify the molecular mechanisms controlling mafB expression, a genomic clone of the mouse mafB gene was isolated and analyzed. RNase protection analysis determined the transcription initiation site at 389 bp upstream from the translation initiation site. The 3' end of the gene is located at 946 bp downstream from the termination codon. The gene lacks intron structure. Sequence analysis showed a TATA-like sequence (5'-GATAAAA-3') and an inverted CCAAT-box (5'-ATTGG-3') in the promoter region. Upstream of these sequences, there are several potential regulatory elements, including two GC boxes (5'-GGGCGG-3'), and a palindromic sequence (5'-GTCAGCTGAC-3') which contains two Maf recognition elements (MARE, 5'-GCTGAC-3') and an E-box (5' CAGCTG-3'). Transient transfection analysis with the 5'-flanking region of the mafB gene demonstrated that these elements are important for mafB gene expression. In addition, cotransfection analysis indicated that the MyoD activates the mouse mafB promoter and the gene is positively auto-regulated by its own product. PMID- 10721737 TI - Characterization of a ubiquitous expressed gene family encoding polygalacturonase in Arabidopsis thaliana. AB - Pectin, as one of the major components of plant cell wall, has been implicated in many developmental processes occurring during plant growth. Among the different enzymes known to participate in the pectin structure modifications, polygalacturonase (PG) activity has been shown to be associated with fruit ripening, organ abscission and pollen grain development. Until now, sequence analyses of the deduced polypeptides of the plant PG genes allowed their grouping into three clades corresponding to genes involved in one of these three activities. In this study, we report the sequence of three genomic clones encoding PG in Arabidopsis thaliana. These genes, together with 16 other genes present in the databases form a large gene family, ubiquitously expressed, present on the five chromosomes with at least two gene clusters on chromosomes II and V, respectively. Phylogenetic analyses suggest that the A. thaliana gene family contains five classes of genes, with three of them corresponding to the previously defined clades. Comparison of positions and numbers of introns among the A. thaliana genes reveals structural conservation between genes belonging to the same class. The pattern of intron losses that could have given rise to the PG gene family is consistent with a mechanism of intron loss by replacement of an ancestral intron-containing gene with a reverse-transcribed DNA copy of a spliced mRNA. Following this event of intron loss, the acquisition of introns in novel positions is consistent with a mechanism of intron gain at proto-splice sites. PMID- 10721738 TI - Sequence, expression and functional analysis of the Coccidioides immitis ODC (ornithine decarboxylase) gene. AB - The ornithine decarboxylase (ODC) gene of the human respiratory fungal pathogen, Coccidioides immitis (Ci) was cloned, sequenced, chromosome-mapped, and expressed in Escherichia coli (Ec). The genomic, cDNA and translated sequences are presented. Transformation of an ODC null mutant strain of Ec (EWH 319) with the Ci ODC gene was conducted to confirm function of the protein encoded by the fungal gene. Activity of the enzyme by the bacterial transformant was inhibited by 1, 4-diamino-2-butanone (DAB), a known inhibitor of eukaryotic ODC. Temporal expression of the Ci ODC gene during the parasitic cell cycle is constitutive, based on results of RT PCR. However, results of enzyme activity assays of cell homogenates obtained at different stages of parasitic cell development in vitro showed that the functional protein is present only during periods of isotropic growth and segmentation, and these morphogenetic events can be arrested by the addition of DAB. The observed absence of a difference in steady-state mRNA transcript amounts, and the developmentally correlated variation in levels of enzyme activity, suggest a translational or post-translational mechanism of ODC regulation. Since no PEST sequence was detected in the Ci ODC, enzyme regulation by programmed protein degradation as reported for many other eukaryotic ODCs may not occur in this case. ODC activity appears to play a key role in the morphogenesis of Ci, and the enzyme could be a rational target for therapy of disseminated coccidioidomycosis. PMID- 10721739 TI - The life stages of weight: setting achievable goals appropriate to each woman. AB - Weight is a preoccupation in America, especially among women. Using body mass index (BMI) as the measure, many women are found to be overweight (BMI 25-30 kg/m2) or obese (BMI >30 kg/m2). A BMI that is less than 18.5 kg/m2 reflects underweight, which is best considered separately. This paper is concerned with the classification of body weight, the distribution of body fat, and the nutritional, medical, and stage-of-life factors that affect fat deposition and weight. The main stages of life in regard to adiposity are the prenatal period, infancy and the period of adiposity rebound (childhood), adolescence, and finally adulthood, with some special considerations at and after menopause. PMID- 10721740 TI - The central nervous system and HRT. AB - In postmenopausal women, both the aging process and the hypoestrogenism due to the loss of ovarian function seem to be related to the progressive impairment of cognitive functions and to a higher risk of developing Alzheimer's disease (AD). This paper reviews the potentially beneficial effects of hormonal replacement therapy (HRT) on cognition and on the risk of developing AD. Articles relevant to the topic were selected by reviewing MEDLINE data and references of previous published reviews on this subject. Epidemiological studies on the effects of HRT on cognitive functioning have yielded disparate results, perhaps because of varying methodology and designs. However, the available data suggest that the use of HRT could be associated with a lower risk for AD. This conclusion should be interpreted with caution, since most of the studies were case-control studies, and thus subjected to several sources of bias. Further well-designed and conducted clinical trials and longitudinal studies would be required to clarify the effects of estrogens on cognition and AD. PMID- 10721742 TI - Fertility and uterine size among Asian women undergoing hysterectomy for leiomyomas. AB - OBJECTIVE: To ascertain whether an epidemiological relationship exists between fertility and uterine leiomyomas, and whether uterine size is associated with fertility among Asian women undergoing hysterectomy for leiomyomas. METHODS: The study was conducted in Sapporo, Japan. 91 women undergoing hysterectomy for myomas were compared with age-matched controls with respect to reproductive factors. Further comparisons were made of these factors in women with large myomas and those with small ones. RESULTS: Women with leiomyomas were more likely to be nulliparous than controls (P < .01), and the risk of leiomyoma increased as the number of births decreased (P < .01). Time since last birth was associated with increased risk in women with large myomas (P < .05). Women with fewer births undergoing hysterectomy for leiomyomas tended to have greater uterine weight. CONCLUSION: Fewer births may be a cause of larger leiomyomas. PMID- 10721741 TI - Treatment of hyperlipidemia in women. AB - Coronary artery disease (CAD) is the most common cause of death in the United States. It is responsible for the deaths of 480,000 people annually. Half of these fatalities are in women. More women die of CAD than due to all cancers combined. The clinical presentation of women with CAD can be very subtle, and atypical as compared to men. Furthermore, women also face a worse prognosis than men following surgical therapy for CAD. Hyperlipidemia is a well-known risk factor for CAD in women, particularly elevated triglycerides and low HDL cholesterol levels. Although estrogen replacement therapy has been considered a primary modality to alleviate some cardiovascular risk in post menopausal women, the results of the recently published HERS trial highlight the need for more research in this field. PMID- 10721743 TI - Cryopreservation of human spermatozoa: comparison of TEST-yolk buffer and glycerol. AB - OBJECTIVE: Comparison of human sperm motility, morphology, and sperm membrane integrity using two cryoprotective media, Test-yolk buffer with glycerol and glycerol alone. METHODS: Semen samples from 10 healthy donors were divided and frozen with either glycerol or a combination of glycerol and TEST-yolk buffer. Semen characteristics were evaluated before freezing and after thawing. Motility was measured at 0, 60, 120, and 180 minutes post-thaw following removal of the cryoprotectant (post-wash). RESULTS: Percentage of motile sperm decreased significantly compared to prefreeze values in both groups. Post-thaw motility following removal of the freezing media was higher in specimens that were frozen in TEST-yolk buffer compared to those frozen in glycerol at 0 minutes (P = 0.004). Similarly, specimens cryopreserved in TEST-yolk buffer had higher sperm motility compared to aliquots that were frozen in glycerol at 180 minutes (P = 0.013). The percentage of normal sperm forms was significantly higher post-wash and post-thaw in specimens that were cryopreserved in TEST-yolk buffer (P = 0.04). CONCLUSIONS: Sperm cryopreserved in TEST-yolk buffer had significantly better motility, morphology, and sperm membrane integrity than sperm preserved in glycerol alone. PMID- 10721744 TI - Bioresorbable bone graft substitutes of different osteoconductivities: a histologic evaluation of osteointegration of poly(propylene glycol-co-fumaric acid)-based cement implants in rats. AB - Bioresorbable bone graft substitutes may significantly reduce the disadvantages associated with autografts, allografts and other synthetic materials currently used in bone graft procedures. We investigated the biocompatibility and osteointegration of a bioresorbable bone graft substitute made from the unsaturated polyester poly(propylene-glycol-co-fumaric acid), or simply poly(propylene fumarate), PPF, which is crosslinked in the presence of soluble and insoluble calcium filler salts. Four sets of animals each having three groups of 8 were evaluated by grouting bone graft substitutes of varying compositions into 3-mm holes that were made into the anteromedial tibial metaphysis of rats. Four different formulations varying as to the type of soluble salt filler employed were used: set 1--calcium acetate, set 2--calcium gluconate, set 3- calcium propionate, and set 4--control with hydroxapatite, HA, only. Animals of each of the three sets were sacrificed in groups of 8 at postoperative week 1, 3, and 7. Histologic analysis revealed that in vivo biocompatibility and osteointegration of bone graft substitutes was optimal when calcium acetate was employed as a soluble salt filler. Other formulations demonstrated implant surface erosion and disintegration which was ultimately accompanied by an inflammatory response. This study suggested that PPF-based bone graft substitutes can be designed to provide an osteoconductive pathway by which bone will grow in faster because of its capacity to develop controlled porosities in vivo. Immediate applicability of this bone graft substitute, the porosity of which can be tailored for the reconstruction of defects of varying size and quality of the recipient bed, is to defects caused by surgical debridement of infections, previous surgery, tumor removal, trauma, implant revisions and joint fusion. Clinical implications of the relation between developing porosity, resulting osteoconduction, and bone repair in vivo are discussed. PMID- 10721745 TI - Coatings on zirconia for medical applications. AB - In order to combine the mechanical properties of a high-strength inert ceramic (yttria-stabilised zirconia, ZrO2-3%Y2O3, defined as zirconia in the text) with the specific properties of bioactive materials, some zirconia samples were coated by two bioactive phosphosilicate glasses and glass-ceramics: RKKP and AP40. Coatings of about 200-300 microm thickness were prepared by a simple and low-cost firing method. They were characterised by optical and scanning electron microscopy (SEM) and compositional analysis (EDS). The adhesion of the coatings on zirconia was tested by shear tests. Vickers indentations at the coating/zirconia interface were performed in order to observe the crack propagation path. The reactivity of glasses and glass-ceramics coatings towards a simulated body fluid (SBF), having the same ion concentration as that of human plasma, was evaluated and compared to that of the bulk glass and glass-ceramics, by examining the morphology of the reaction layer formed on the surface of the coated zirconia after one month of soaking in the SBF at 37 degrees C. PMID- 10721746 TI - Correlation between impact strength and fracture toughness of PMMA-based bone cements. AB - The thrust of the work involved determining the impact strength, IS (in kJ m(-2)) [using non-ASTM-sized Charpy-type specimens and an in-house impact tester] and fracture toughness, K1c (in MPa square root(m)) [using ASTM-sized rectangular compact tension specimens] of Surgical Simplex P and three variants of Palacos R acrylic bone cements. The attractions and drawbacks of this method for determining IS are detailed. The best fit to the K1c and IS results yielded a power relationship K1c = 0.795(IS)(0.59). The usefulness and limitations of this relationship are detailed. PMID- 10721747 TI - Control of hepatocyte function on collagen foams: sizing matrix pores toward selective induction of 2-D and 3-D cellular morphogenesis. AB - While microporous biopolymer matrices are being widely tested as cell culture substrates in hepatic tissue engineering, the microstructural basis for their control of cell differentiation is not well understood. In this paper, we studied the role of void size of collagen foams in directing the induction of liver specific differentiated morphology and secretory activities of cultured rat hepatocytes. Hepatocytes cultured on collagen foams with subcellular sized pore diameters of 10 microm assumed a compact, cuboidal cell morphology, rapidly achieving monolayer coverage, and secreted albumin at the rate of 40 +/- 8 pg/cell/d. Increasing the pore size to 18 microm elicited a distinctly spread cellular phenotype, with discontinuous surface coverage, and albumin secretion rates declined precipitiously to 3.6 +/- 0.8 pg/cell/d. However, when collagen foams with an even higher average void size of 82 microm were used, hepatocytes exhibited high degree of spreading within an extensive three-dimensional cellular network, and exhibited high albumin secretory activity (26 +/- 0.6 pg/cell/d). The effect of void geometry on cellular ultrastructral polarity was further analyzed for the three void size configurations employed. The distribution of the cell-cell adhesion protein, E-cadherin, was primarily restricted to cell-cell contacts on the 10 microm foams, but was found to be depolarized to all membrane regions in cells cultured on the 18 and 82 microm foams. Vinculin-enriched focal adhesions were found to be peripherally clustered on cells cultured on 10 microm foams, but were found to redistribute to the entire ventral surface of cells cultured on the 18 and 82 microm foams. Overall, we demonstrate the significance of designing pore sizes of highly adhesive substrates like collagen toward selective cell morphogenesis in 2-D or 3-D. Subcellular and supercellular ranges of pore size promote hepatocellular differentiation by limiting 2-D cell spreading or effecting 3-D intercellular contacts, while intermediate range of pore sizes repress differentiation by promoting 2-D cell spreading. PMID- 10721748 TI - Articular cartilage repair in rabbits by using suspensions of allogenic chondrocytes in alginate. AB - The feasibility of allogenic implants of chondrocytes in alginate gels was tested for the reconstruction in vivo of artificially full-thickness-damaged articular rabbit cartilage. The suspensions of chondrocytes in alginate were gelled by the addition of calcium chloride solution directly into the defects giving in situ a construct perfectly inserted and adherent to the subchondral bone and to the walls of intact cartilage. The tissue repair was controlled at 1, 2, 4 and 6 months after the implant by NMR microscopy, synchrotron radiation induced X-ray emission to map the sulfur of glycosaminoglycans and by histochemistry. Practically a complete repair of the defect was observed 4-6 months from the implant of the chondrocytes with the recovery of a normal tissue structure. Controls in which Ca-alginate alone was implanted developed only a fibrous cartilage. PMID- 10721749 TI - In vitro assessment of new embolic liquids prepared from preformed polymers and water-miscible solvents for aneurysm treatment. AB - Treatment of cerebral aneurysms by embolic liquids has been proposed as an alternative strategy to coil or balloon techniques. In order to assess the feasibility of this approach, a general screening of preformed polymers dissolved in biocompatible, water-miscible solvents has been carried out. The solubilizing capacity of the solvents has been evaluated by the solubility parameters approach. The viscosity of the solutions has been determined and the precipitation characteristics of the embolic liquids have been investigated in phosphate buffer solution pH 7.4 at 37 degrees C to mimic physiological conditions. The radiopaque agent bismuth (III) oxide was added to solutions having appropriate precipitation characteristics and the angiographic assessment, in an in vitro aneurysm model, were consistent with the precipitation properties and confirmed that only hard and coherent masses allowed satisfactory embolization. However, the solubilizing prediction using the calculation of the solubility parameters was only partially successful owing to the highly hydrophilic functional groups of the chosen solvents. This failing justifies the experimental screening that was carried out. This study pointed out that the frequently used solvent dimethyl sulfoxide could be replaced by more biocompatible solvents offering the possibility of using other preformed polymers. In conclusion, nine solutions of the selected polymer-solvent combinations could be used as embolic liquids for the treatment of cerebral aneurysms with respect to their satisfactory precipitation properties and viscosity. PMID- 10721750 TI - Flow cytometry analysis of the effects of pre-immersion on the biocompatibility of glass-reinforced hydroxyapatite plasma-sprayed coatings. AB - Multilayered coatings composed of mixtures of HA and P2O5-based bioactive glasses are of potential clinical benefit in orthopaedic and dental surgery. Pre immersion of these materials has been reported to further enhance their efficacy in vivo, although the precise biological effects of this treatment are not yet known. In this study we have therefore prepared double-layer plasma-sprayed coatings and evaluated the effects of pre-immersion on the growth and function of human osteosarcoma cells in vitro, using the MTT assay and flow cytometry analysis, respectively. The results showed that the increase in numbers of viable cells was the same or elevated following incubation on the pre-immersed HA and glass-reinforced HA coatings compared with the non-immersed materials. In addition, the expression of bone sialoprotein and fibronectin, two key connective tissue antigens, was up-regulated in cultures grown on the pre-immersed surfaces compared with the non-treated materials. Moreover, cell numbers and antigen expression both improved as the proportion of glass increased, particularly in the pre-immersed samples. Our findings thus suggest that the immersion treatment of these materials appeared to improve the response of these bone-like cells. PMID- 10721751 TI - Influence of different parameters on drug release from hydrogel systems to a biomembrane model. Evaluation by differential scanning calorimetry technique. AB - A comparative study on the drug release capacity of four water swellable polymeric systems was carried out by differential scanning calorimetry (DSC). The polymeric systems chosen were alpha,beta-polyaspartahydrazide (PAHy) crosslinked by glutaraldehyde (GLU) (PAHy-GLU) or by ethyleneglycoldiglycidylether (EGDGE), (PAHy-EGDGE), polyvinylalcohol (PVA) crosslinked by glutaraldehyde (PVA-GLU) and alpha,beta-poly(N-hydroxyethyl)-DL-aspartamide (PHEA) by gamma irradiation (PHEA gamma matrices). The degree of crosslinking for PAHy-GLU, PAHy-EGDGE and PVA-GLU samples was about 0.4 and 0.8. These hydrogels were characterized as free of drugs and were loaded with diflunisal (DFN) (approximately 2.5% w/w). Diflunisal, a non-steroidal anti-inflammatory drug, has been chosen as a model drug to be incorporated into polymeric matrices to follow the release processes of a drug from these hydrogels to a model membrane made by unilamellar vesicles of dipalmitoylphosphatidylcholine (DPPC). Differential scanning calorimetry appears to be a suitable technique to follow the transfer kinetics of the drug from the controlled release system to the biomembrane model. The drug releases from all the considered polymeric hydrogels, were compared with the release observed from the drug solid form by examining the effects on the thermotropic behaviour of DPPC unilamellar vesicles. The release kinetics of the drug from hydrogels were followed at 25, 37 and 50 degrees C to evidence the influence of temperature on the drug release and on the successive transfer to biological membrane model. Particularly, it appears evident that the total amount of drug transferred and the release rate are affected by the polymer crosslinking degree (it increases with crosslinking decrease) as well as by the nature of crosslinking agent. In fact, the drug release profiles from PAHy-GLU samples are more differentiated than those from PAHy-EGDGE. The effect of parameters correlating with the properties of starting polymer, such as water-affinity, crystallinity, glass-to rubber transition temperature and affinity towards drug molecules, has been also evaluated. PMID- 10721752 TI - Cultured differentiated human urothelial cells in the biomaterials field. AB - To review the use of normal cultured differentiated human urothelial cells in the biomaterials field, we checked the literature for human urothelial cells in culture (HUC) both for their use in biocompatibility assessment and as bioartificial devices. The in vitro culture of differentiated human urothelium is now a simple and reliable procedure. These techniques provide new tools for biocompatibility assessment of urinary biomaterials, because for the rational design of a testing procedure, it is preferable that the particular cell culture models selected should be closely related to the end-use application. The emerging use of HUC culture should lead to the development of bioartificial tissue for urinary tract reconstruction. Tissue engineering techniques require urothelial cells and cell delivery matrices. The cytocompatibility of novel artificial delivery matrices should be assessed in vitro before implantation using cultured HUC to find the best material available. PMID- 10721753 TI - Bond strength of plasma-sprayed hydroxyapatite/Ti composite coatings. AB - One of the most important clinical applications of hydroxyapatite (HA) is as a coating on metal implants, especially plasma-sprayed HA coating applied on Ti alloy substrate. However, the poor bonding strength between HA and Ti alloy has been of concern to orthopedists. In this paper, an attempt has been made to enhance the bonding strength of HA coating by forming a composite coating with Ti. The bioactivity of the coating has also been studied. HA/Ti composite coatings were prepared via atmospheric plasma spraying on Ti-6Al-4V alloy substrates. The bond strength evaluation of HA/Ti composite coatings was performed according to ASTM C-633 test method. X-ray diffractometer and scanning electron microscopy were applied to identify the phases and the morphologies of the coatings. The bioactivity of HA/Ti composite coating was qualified by immersion of coating in simulated body fluid (SBF). The obtained results revealed that the addition of Ti to HA improved the bonding strength of coating significantly. In the SBF test, the coating surface was covered by carbonate apatite, which was testified by X-ray photoelectron spectroscope, indicating good bioactivity for HA/Ti composite coating. The bioactivity of the coating has not been reduced by the addition of Ti. PMID- 10721754 TI - Analysis of released products from oxidized ultra-high molecular weight polyethylene incubated with hydrogen peroxide and salt solutions. AB - The wear of ultra-high molecular weight polyethylene (UHMWPE) implants generates polymeric and metallic particulate, which can be phagocytosed by human macrophages. The generation of these UHMWPE particles has been attributed to wear mechanisms and oxidation of the material. Many cell/particle studies have focused specifically on investigating particles of virgin materials themselves (i.e. virgin UHMWPE), while in fact, there is a strong likelihood that the oxidation processes encountered by the materials will yield particles with very different surface chemistries. Therefore, it is conceivable that chemical changes in the material would lead to altered cellular responses, as measured in the various cell study models. This paper has focused on the characterization of UHMWPE particulates that have been exposed to various conditions simulating processing steps and some of the oxidative and hydrolytic agents related to inflammatory responses. These include gamma-irradiation, thermal treatment and chemical oxidation by H2O2 and saline solutions. Oxidation of the particles was measured using Fourier transform infrared spectroscopy (FTIR). Degradation products were isolated from the incubation solutions using high-performance liquid chromatography (HPLC). UHMWPE particulates underwent extensive oxidation after gamma-irradiation and thermal treatments. There were marked differences following treatments of film samples taken from bar stock and the virgin particle samples. Polymer-related products, containing alkenes, alkanes and hydroxyl groups, were found in the incubation solutions. The study concluded that future work must consider both the particulates' surface chemistry and the possibility of soluble degradation products when assessing UHMWPE/cellular interactions. PMID- 10721755 TI - Neuromuscular rehabilitation and electrodiagnosis. 1. Central neurologic disorders. AB - This self-directed learning module highlights the medical treatment and rehabilitation intervention of certain central neurologic disorders encountered in physiatric practice. It is part of the chapter on neuromuscular rehabilitation and electrodiagnosis in the Self-Directed Physiatric Education Program for practitioners and trainees in physical medicine and rehabilitation. This article contains sections on multiple sclerosis, Parkinson's disease, and amyotrophic lateral sclerosis. Information covered in these sections includes discussions of the current medical management and the benefits of comprehensive rehabilitation and interventions for specific impairments seen in these conditions. PMID- 10721756 TI - Neuromuscular rehabilitation and electrodiagnosis. 2. Localized peripheral neuropathy. AB - This self-directed learning module highlights advances in diagnosis and treatment of focal injuries to peripheral and cranial nerves. It is part of the chapter on neuromuscular rehabilitation and electrodiagnosis in the Self-Directed Physiatric Education Program for practitioners and trainees in physical medicine and rehabilitation. Nerve conduction and electromyographic techniques are employed as extensions of the physician's senses in clinical examination and diagnosis. The findings are used to plan treatment, and to predict and measure outcomes. Electrodiagnosis and medical and surgical treatments of nerve injuries are discussed in light of the managed-care utilization review of services. PMID- 10721757 TI - Neuromuscular rehabilitation and electrodiagnosis. 3. Generalized peripheral neuropathy. AB - This self-directed learning module provides an updated tool for establishing the differential diagnosis and subsequently designing a cost-effective workup for patients with peripheral neuropathy. It is part of the chapter on neuromuscular rehabilitation and electrodiagnosis in the Self-Directed Physiatric Education Program for practitioners and trainees in physical medicine and rehabilitation. Medication intervention for neuropathic pain is reviewed by medication class with recommendations for starting doses and review of side-effect profiles. This article also reviews the current treatment standards for a patient with juvenile onset diabetes, including recommendations for glucose control, management of nephropathy and retinopathy, and care of foot complications. PMID- 10721758 TI - Neuromuscular rehabilitation and electrodiagnosis. 4. Specialized neuropathy. AB - This self-directed learning module briefly highlights the differential diagnosis for acute weakness in patients with acute respiratory failure requiring prolonged mechanical ventilation. It is part of the chapter on neuromuscular rehabilitation and electrodiagnosis in the Self-Directed Physiatric Education Program for practitioners and trainees in physical medicine and rehabilitation. This article includes a discussion on the role of exercise in the treatment of patients with the late effects of poliomyelitis or with acute inflammatory demyelinating polyradiculoneuropathy. PMID- 10721759 TI - Neuromuscular rehabilitation and electrodiagnosis. 5. Myopathy. AB - This self-directed learning module highlights hypotonia, facioscapulohumeral dystrophy, and herbal supplements causing muscle weakness. It is part of the chapter on neuromuscular rehabilitation and electrodiagnosis in the Self-Directed Physiatric Education Program for practitioners and trainees in physical medicine and rehabilitation. This section presents advances in the diagnosis of myotubular dystrophy, myoblast transfer, and problems associated with the increased use of herbal supplements. PMID- 10721760 TI - Acute weakness syndromes in the critically ill patient. AB - Over the past three decades there has been increasing interest in acute weakness syndromes in critically ill mechanically ventilated patients. Many of these patients require rehabilitation, and some understanding of potential etiologies and functional outcomes for these syndromes is useful to rehabilitation practitioners. A clearer understanding of these syndromes has evolved over time, as has the terminology to describe these conditions. This article will review commonly encountered causes of acute weakness in critically ill patients, including disorders of the peripheral nerves, the neuromuscular junction, and muscle. PMID- 10721762 TI - Rehabilitation of orthopedic and rheumatologic disorders. 2. Connective tissue diseases. AB - This self-directed learning module highlights assessment and therapeutic options in the rehabilitation of patients with connective tissue diseases. It is part of the chapter on rehabilitation of orthopedic and rheumatologic disorders in the Self-Directed Physiatric Education Program for practitioners and trainees in physical medicine and rehabilitation. New advances covered in this article include the management of patients who have polymyalgia rheumatica, juvenile rheumatoid arthritis, ankylosing spondylitis, rheumatoid cervical spine, and rheumatoid hand. PMID- 10721761 TI - Rehabilitation of orthopedic and rheumatologic disorders. 1. Osteoporosis. AB - This self-directed learning module reviews and summarizes recent literature on osteoporosis. It is part of the chapter on rehabilitation of orthopedic and rheumatologic disorders in the Self-Directed Physiatric Education Program for practitioners and trainees in physical medicine and rehabilitation. The areas covered include pathophysiology of primary and secondary osteoporosis, effects of various pharmacologic treatments on bone metabolism, and the utility of available diagnostic tests. Management strategies for perimenopausal women as compared with postmenopausal women with established osteoporosis are discussed. This is followed by an evaluation and management plan for the older man with acute osteoporotic fracture. PMID- 10721763 TI - Rehabilitation of orthopedic and rheumatologic disorders. 3. Degenerative joint disease. AB - This self-directed learning module highlights assessment and therapeutic options in the rehabilitation of patients with osteoarthritis. It is part of the chapter on rehabilitation of orthopedic and rheumatologic disorders in the Self-Directed Physiatric Education Program for practitioners and trainees in physical medicine and rehabilitation. New advances covered in this article include updates on conservative and operative treatment of lumbar spinal stenosis and pediatric hip diseases, prophylactic therapy for thromboembolic disease after lower limb joint replacement, new therapies for osteoarthritis, and the impact of exercise on outcome following hip replacement in active persons. PMID- 10721764 TI - Rehabilitation of orthopedic and rheumatologic disorders. 4. Musculoskeletal disorders. AB - This self-directed learning module highlights assessment and therapeutic options in the rehabilitation of patients with orthopedic and musculoskeletal disorders. It is part of the chapter on rehabilitation of orthopedic and rheumatologic disorders in the Self-Directed Physiatric Education Program for practitioners and trainees in physical medicine and rehabilitation. This article discusses new advances in such topics as idiopathic scoliosis, nontraumatic shoulder pain, rotator cuff tendinitis, and Dupuytren's disease. PMID- 10721765 TI - Lumbosacral epidural steroid injections. AB - Epidural steroid injections for the treatment of low back pain and associated leg pain have been used for half a century. Initial use was empirical and not without controversy. An abundant literature has accumulated describing their use and potential benefit. Previous controlled studies are limited by methodology and technique. A growing body of literature suggests that they exert some of their clinical effect by reducing epidural and perineural inflammation. More current studies using fluoroscopy with radiographic contrast and precise epidural steroid placement suggests they may obviate surgery in some patients with true radicular pain. PMID- 10721766 TI - CD164--a novel sialomucin on CD34+ cells. AB - Hematopoiesis in adult bone marrow is a tightly regulated process involving interactions between cytokine and adhesion receptors on hematopoietic progenitor cells and their cognate ligands in the immediate microenvironment. These interactions control hematopoietic stem cell self-renewal, quiescence, commitment and migration. Recently, sialomucins have assumed some importance in hematopoiesis, with six of these receptors, CD34, PSGL-1, CD43, PCLP, CD45RA and CD164, having been identified on primitive hematopoietic precursor cells and/or their associated stromal/endothelial elements. This article reviews the cloning, expression and function of the recently identified sialomucin, CD164, which is highly expressed by primitive hematopoietic progenitor cells. The CD164 receptor is implicated in mediating or regulating hematopoietic precursor cell adhesion to stroma, and may serve as a potent negative regulator of hematopoietic progenitor cell proliferation. PMID- 10721767 TI - Effect of thrombopoietin on acute myelogenous leukemia blasts. AB - It has been shown that the expression of c-mp1, which is a specific receptor for thrombopoietin (TPO), is restricted to the surface of megakaryocytes, platelets, human CD34+ progenitor cells and human erythroid/megakaryocytic leukemic cell lines. Recently, however, it has been reported that some acute myelogenous leukemia (AML) blasts expressed c-mp1 on their cell surface and proliferated in response to TPO. We therefore investigated the effect of thrombopoietin on the growth of leukemic blasts from patients with CD7-positive acute myelogenous leukemia (AML), which is a distinct biological and clinical subtype of AML. Significant growth responses of leukemic blasts to TPO were seen in 10/10 CD7+ and 7/20 CD7- AML cases using 3H-thymidine incorporation, while synergistic stimulatory effects of TPO with stem cell factor (SCF), interleukin-3 (IL-3), granulocyte colony-stimulating factor, and granulocyte-macrophage colony stimulating factor were observed in both groups. In a leukemic blast colony assay, significant growth response to TPO was observed in 5/6 CD7+ and 4/17 CD7- AML cases examined. Furthermore, the expression of c-mp1 seemed to be higher in CD7+ AML cases than in CD7- cases, suggesting a relationship between the expression of c-mp1 and the proliferative response to TPO. These findings imply that CD7+ leukemic blasts express functional TPO receptors and proliferate in response to TPO. Thus CD7 expression on AML blasts may indicate the involvement of leukemic progenitors at an early stage of multipotent hemopoietic stem cells. In this review, we discuss the effect of TPO on AML blasts, especially in CD7+ AML cases. PMID- 10721768 TI - The cAMP signaling pathway as a therapeutic target in lymphoid malignancies. AB - Certain subsets of lymphoid cells, such as thymocytes or peripheral B cells, undergo apoptosis after treatment with agents which elevate intracellular 3',5' cyclic adenosine monophosphate (cAMP). Investigators have also noted induction of apoptosis of chronic lymphocytic leukemia (CLL) cells following treatment with methylxanthines, a phenomenon that may, at least in part, be due to the activity of these drugs as non-specific phosphodiesterase (PDE) inhibitors. We discuss three general strategies for altering cAMP-mediated signal transduction in lymphoid cells. After a review of what is known about the expression and regulation of PDE families in human lymphoid cells, we focus on the use of isoform-specific PDE inhibitors as potential therapeutic agents in CLL. Our work has suggested that despite the presence of PDE1, PDE3B, PDE4 and PDE7 enzymes in CLL, inhibition of PDE4 results in uniquely potent induction of apoptosis in CLL cells. This effect is relatively specific as comparable treatment of human peripheral blood T cells does not induce apoptosis. Clinical trials utilizing PDE4 inhibitors are indicated in the therapy of CLL patients resistant to standard therapy. PMID- 10721769 TI - Arsenic-induced apoptosis in malignant cells in vitro. AB - Arsenic trioxide-induced apoptosis was identified by morphological change and nucleosomal DNA fragmentation in hematopoietic malignant cells and neuroblastoma cells. Arsenic trioxide directly induced apoptosis in the acute promyelocytic cell line NB4 cells at a low dose of 1 microM, whereas all-trans-retinoic acid caused the cells to differentiate and finally induced apoptosis. In addition to the involvement of caspase 3 in arsenic trioxide-induced apoptosis of NB4 cells, the activation of caspase 8 was also shown to be involved by Western blot analysis or by apoptosis inhibition assay using caspase 8 inhibitor Ac-IETD-CHO. The down-regulation of Bcl-2 protein was shown in arsenic trioxide-treated pre apoptotic and early apoptotic mouse B-cell line LyH7 cells, which overexpress Bcl 2 protein, by the studies of Western blot and immunoelectron microscopy. Arsenic trioxide also induced apoptosis in the majority of neuroblastomas cell lines. The arsenic-induced apoptosis in neuroblastoma cell lines was mediated by the activation of caspase 3 in all cases tested. In regard to the intracellular content of reduced glutathione in various neuroblastoma cell lines, the level in the cells sensitive to arsenic trioxide was under 40 nmol/mg protein, but the cells having more than 40 nmol/mg protein did not undergo apoptosis. N acetylcysteine protected neuroblastoma cells from arsenic-induced apoptosis. Therefore, the intracellular glutathione content may be a good indicator of application of arsenic trioxide for various kinds of cancer cells. Our results raise the possibility that arsenic trioxide will be effective even against a solid tumor such as neuroblastoma and warrants clinical trials for patients with other kinds of tumors not only by systemic therapy but also using local therapy. PMID- 10721770 TI - Cost analyses of adjunct colony stimulating factors for acute leukemia: can they improve clinical decision making. AB - Colony stimulating factors reduce the duration of neutropenia following intensive chemotherapy in a variety of settings, but the advantages in the management of leukemia are inconclusive. The variations in clinical results and the high costs of granulocyte colony-stimulating factor (G-CSF) and granulocyte macrophage colony-stimulating factor (GM-CSF) have led to confusion over appropriate use for leukemia patients. In this paper, we reviewed published information on costs and cost-effectiveness of growth factors for childhood and adult leukemia patients. Medline and Healthstar databases were searched for original research articles that contain cost or cost-effectiveness analyses of G-CSF (filgrastim) and GM-SCF (sargramostim) in oncology cooperative group trials. Published manuscripts and abstracts presented at national or international oncology conferences were included. The cost of adjunct treatment was evaluated in two studies of pediatric ALL, one study of adult AML, and two studies of AML in older adults (>55 years). The use of G-CSF for children with ALL was associated with reductions in days to ANC recovery, fewer documented infections, a shorter duration of hospitalization, and small (but not significant) additional costs. In adult AML patients, benefits included a shortening of the duration of neutropenia and hospital stays, a lower incidence of infection and febrile episodes, less use of antibiotics, and cost savings of $2,230 and $2,310 in two studies and an increase if $120 in the third study. This summary suggests that economic analyses can provide useful information to assist clinical decision-making. For pediatric ALL patients, this information indicates that G-CSF use is unlikely to have significant cost implications, and its use should be based on clinical considerations. In studies of adult and older adult AML patients, both GM-CSF and G-CSF have clinical benefits and can be expected to lead to a decrease in overall costs. PMID- 10721771 TI - Prediction of prognosis following fludarabine used as secondary therapy for chronic lymphocytic leukemia. AB - A prognostic factor analysis for survival and response was conducted on 374 previously treated patients who were treated with one of four fludarabine regimens. As several prognostic factors were associated with response and survival, multivariate analyses were conducted to identify the combination of factors which best describe the prognosis as regards response to fludarabine and survival. Statistical models to identify the hazard for survival and probability of response were developed. The characteristics which were included in the model predicting for survival were sex, age, number of prior treatments, performance status, hemoglobin level, serum albumin, and alkaline phosphatase levels. The characteristics most strongly associated with response were the hemoglobin levels, serum albumin, and the number of prior treatments. These models can be used to identify risk groups to guide physicians in decision-making and to assist in the design and analysis of clinical trials. PMID- 10721772 TI - Oral idarubicin in patients with late chronic phase chronic myelogenous leukemia or chronic myelomonocytic leukemia. AB - Patients with chronic myelogenous leukemia (CML) who have failed or cannot receive interferon alpha (IFN-alpha) based regimens or patients with advanced chronic myelomonocytic leukemia (CMML) have very limited current therapeutic options. Hence, there is a need to develop new strategies for these patients. This study was undertaken to determine the efficacy and toxicity of a chronic low dose oral idarubicin regimen in these patients as positive data has been generated on this agent in shorter schedules given to patients with other hematological malignancies. Eighteen patients were treated on study. The starting dose of oral idarubicin was 2 to 5 mg/m2 daily depending in initial WBC count. This dose was escalated in the absence of Grade 4 myelosuppression or Grade 3 or 4 extramedullary toxicity. Oral idarubicin was given daily for 28 days followed by a 21 day break off treatment in repeated cycles until there was evidence of disease progression or intolerable toxicity. The dose of idarubicin was adjusted, at 2-week intervals, by 25% to maintain a white blood cell (WBC) count between 2 and 4x10(9)/L and a platelet count of >75x10(9)/L. The dose was reduced by 25% for grade 2 extramedullary toxicity and held until toxicity resolved to grade 2 or better for grade 3 toxicity. Oral idarubicin was then restarted at 75% of the initial dose. Five out of 14 CML patients achieved a complete hematologic remission. No CMML patient responded (median survival 3 months). The overall median survival was 24 months. CML patients had a median survival of 28 months. Major toxicities (myelosuppression, gastrointestinal, cardiac) were infrequent with a median cumulative dose of 1110 mg/m2 (range 54-9750). Five patients have received oral idarubicin for > 1 year with no overt cardiotoxicity, reaching median cumulative dose of 2756 mg/m2 (range 2550-9750) which is higher than those documented in prior studies. We conclude that oral idarubicin is sufficiently safe and active to warrant phase II studies investigating it as part of interferon-based regimens in patients with advanced CML. PMID- 10721773 TI - Development of lymphoproliferative disorder of granular lymphocytes in association with hairy cell leukemia. AB - Lymphoproliferative disorder of granular lymphocytes (LDGL) is a low grade T-cell disease characterized by clonal expansion of large granular lymphocytes of either T cell or natural killer (NK) cell lineage that express the cytotoxic T-cell/NK cell antigens CD16, CD56 and/or CD57. LDGL has been described in association with other malignancies, leading to theories of a common abnormal stem cell as well as development of the LDGL as an immune response to a primary tumor. We have studied 32 patients with hairy cell leukemia (HCL). In 15 patients (47%) we detected an increase in cells expressing cytotoxic T-cell/NK cell antigens. In 10(31%) patients these cells were of T cell lineage, while 5 patients (16%) had increased NK-cells. T cell clonality was detected by PCR in all cases with increased cytotoxic T-cells in which adequate DNA was obtained from peripheral blood. Since in 2 patients the LDGL was not present at diagnosis but developed during follow up, our data suggests that clonal LDGL may develop in response to the HCL. The significance of LDGL in the setting of HCL and flow cytometric evaluation of HCL versus LDGL will be discussed. PMID- 10721774 TI - Minimally differentiated acute myeloid leukemia (AML M0): clinico-biological findings of 29 cases. AB - Twenty-nine cases of minimally differentiated acute myeloid leukemia or AML M0 identified among 441 AML diagnosed in the last 12 years are reported. In all cases, flow cytometric analysis using a large panel of monoclonal antibodies and cytogenetic and molecular studies (IgH, TcRbeta, BCR/ABL, AML1/ETO and CBFB-MYH11 rearrangements) were performed. Of the 29 patients, 27 were treated with intensive chemotherapy based on GIMEMA protocols. We noted a greater incidence of older (over 60 years) and male patients (52% and 65%, respectively). CD33, CD13, CD7 and TdT were expressed in 79.3%, 82.7%, 58.6% and 42.8% of cases, respectively. Antigenic MPO was present in 17 of 22 cases (77.3%). Most cases expressed CD34 (93.1%), HLA-DR (93.1%), CD117 (80%) and CD45RA (87%). CD45RO and CD90 were consistently negative. In all cases, we observed an up-expression of bcl-2 and a down-expression of CD95 with an inverse trend between the two markers (r -5253; p 0.03). Karyotypic abnormalities were demonstrated in 53.6% of cases. Of these, 6 involved chromosomes 5, 7 and 8, t(9;22), confirmed by the BCR/ABL transcript, was detected in one case. Rearrangements of the TcRbeta and IgH chains were observed in 3 and 2 cases, respectively. No AML1/ETO and CBFB-MYH11 transcripts were found. Twelve out of 27 patients (44%) achieved a complete remission (CR) (in 2 cases after rescue therapy). Seven early (range 1-9 months) and one late (32 months) relapses were observed. Five patients are alive, but only the 4 who underwent bone marrow transplantation are in persistent first CR. In conclusion, AML M0 is a subtype of AML antigenically well detectable, endowed with many adverse parameters (older age, TdT and CD34 expression, resistance to apoptosis, unfavorable cytogenetic abnormalities) and poor prognosis. A very aggressive consolidation treatment can be useful to improve the outcome. PMID- 10721775 TI - Dexamethasone, carmustine, etoposide, cytarabine, and melphalan (dexa-BEAM) followed by high-dose chemotherapy and stem cell rescue--a highly effective regimen for patients with refractory or relapsed indolent lymphoma. AB - We performed a phase II study to determine the efficacy of maximal cytoreductive therapy with up to five cycles of Dexa-BEAM (dexamethasone, carmustine [BCNU], etoposide, cytarabine, and melphalan) followed by high-dose chemotherapy (HDCT) and autologous stem cell transplantation (ASCT) for patients with advanced relapsed or refractory indolent lymphoma. Thirty-two patients with primary refractory or relapsed indolent lymphoma were treated with the Dexa-BEAM regimen. Thirteen patients had primary refractory disease, 4 patients partial remission, and 15 patients first or subsequent relapse. Patients achieving PR or CR received HDCT with ASCT. The conditioning regimen used was BEAM (carmustine [BCNU], etoposide, cytarabine, and melphalan). Twenty-two patients responded to Dexa-BEAM resulting in a response rate of 78%. Maximum response was observed after 3.2 (range 2-5) courses. One patient with progressive disease died in septic shock during neutropenia. Nineteen patients with partial or complete remission after Dexa-BEAM received HDCT. Hematopoietic stem cells (HSC) were collected after two cycles of Dexa-BEAM. The median number of CD34+ HSC reinfused was 3.1 x 10(6)/kg (range 1.6-8.2 x 10(6)/kg). There was no transplantation-related death. All patients receiving HDCT achieved complete remission. Overall survival (OS) and freedom from treatment failure (FFTF) for all patients are estimated to be 68% and 65% at two years, respectively. With a mean follow-up of 20 months (range 8 42 months), 16/19 patients receiving HDCT are in continuous complete remission. The Dexa-BEAM regimen is effective in overcoming drug resistance in patients with indolent lymphoma who failed to respond to conventional treatment or who relapsed. The CR rate of 100% of those patients receiving HDCT and ASCT after maximal cytoreductive treatment with Dexa-BEAM suggests the use of HDCT at the time of maximal response. PMID- 10721776 TI - Delayed onset of autoimmune hemolytic anemia complicating cladribine therapy for Waldenstrom macroglobulinemia. AB - Therapeutic options for patients with Waldenstrom macroglobulinemia (WM) now include the purine nucleoside analogues, fludarabine and cladribine. Both these agents have been associated with the onset of severe life-threatening autoimmune hemolytic anemia (AIHA) in patients with chronic lymphocytic leukemia (CLL). In these reports, AIHA developed within 6 weeks of drug administration and was generally resistant to conventional therapy. AIHA following purine analogues has not been reported in other hematologic malignancies. We report here on 4 patients with WM who developed AIHA following cladribine therapy. Cladribine was administered as a 2-hour infusion at a dose of 0.12 mg/kg per day for 5 consecutive days, at 28-day intervals. The median number of cycles was four (range, 4 to 6). AIHA occurred at a median of 40 months (range, 24 to 60) from cladribine administration. Only 1 patient responded to oral steroids while the other 3 were resistant to multiple therapeutic interventions. Two patients, 1 in complete remission and 1 requiring transfusional support, remain alive, and 2 have died. In contrast to published reports of the early onset of AIHA occurring after purine analogues in CLL patients, we observed AIHA in WM patients as a delayed event. PMID- 10721777 TI - EBVD combination chemotherapy plus low dose involved field radiation is a highly effective treatment modality for early stage Hodgkin's disease. AB - To evaluate the efficacy of EBVD combination chemotherapy followed by low dose (LD) involved field (IF) radiation therapy (RT) in patients with clinical stage (CS) I-IIA Hodgkin's disease (HD), we analyzed 148 patients treated in our Unit from March 1988 to November 1995. EBVD consisted of Epirubicine 40 mg/m2, Bleomycin 10 mg/m2, Vinblastine 6 mg/m2 and Dacarbazine 300 mg. All drugs were administered i.v. at days 1 and 15, every 4 weeks, for a total of 4-6 cycles. LDIF RT (24-32 Gy) was scheduled for patients with complete response (CR) or >90% reduction of tumor load, after EBVD. Patients with stable or progressive disease (SD, PD) after EBVDx3 or poor compliance to the regimen received mantle or inverted Y RT at standard dose. The median follow-up of patients currently alive was 71.5 months. 129 patients achieved a CR after EBVD and 10 a >90% reduction of tumor load, for a post-CT response rate of 94%. Eight patients had SD after EBVDx3 and one had a partial response with poor compliance. All 9 patients received mantle or inverted Y RT and 8/9 achieved a CR. Nine patients relapsed at a median of 7 months from the end of treatment. At 10 years, FFS was 90% and overall survival 95%. Six patients have died so far; 5 of HD and one of stroke. One patient developed a diffuse large cell lymphoma 48 months after the diagnosis of HD. We conclude that EBVD followed by LDIF RT is a highly effective regimen for patients with CS I-IIA HD. Longer follow up is required to assess the risk of secondary malignancies, especially solid tumors. PMID- 10721778 TI - Expression of the CD11c antigen in B-cell chronic lymphoproliferative disorders. AB - This study analyzed the expression of the beta2 integrin CD11c in 155 patients with well-characterized B-cell chronic lymphoproliferative disorders: 106 B-cell chronic lymphocytic leukemias (B-CLL), 21 hairy cell leukemias (HCL), 9 B-cell prolymphocytic leukemias (PLL) and 19 low grade non-Hodgkin's lymphomas (NHL) in leukemic phase. CD11c was expressed in 100% of patients with HCL and B-PLL, while in B-CLL and NHL it was expressed in only 49 and 57%, respectively. Furthermore, in B-CLL the expression of CD11c was found mainly in patients with early stage of disease. In addition, when the fluorescence intensity of CD11c, calculated by MFI, was evaluated, it proved significantly higher in HCL and B-PLL compared to the values recorded in B-CLL and NHL (325 and 387 vs 34 and 56, respectively) (p < 0.05). Our results demonstrate that the evaluation of CD11c, both in terms of overall positivity and of fluorescence intensity, represents an additional useful parameter for a more precise differential diagnosis within the spectrum of B-cell chronic lymphoproliferative disorders. PMID- 10721779 TI - Prognostic significance of serum soluble interleukin-2 receptor level in non Hodgkin's lymphoma: a single center study in Japan. AB - Interleukin 2 receptor is expressed not only on the surface of activated T or B lymphocytes, but also on certain lymphoid malignancies. The receptor is released from the cell membrane as soluble form (sIL-2R). Serum sIL-2R level is a sensitive and quantitative marker of circulating peripheral blood mononuclear cell activation or specific tumor cell growth including non-Hodgkin's lymphoma (NHL). However, the relevance of serum sIL-2R levels relating to clinical outcome in adult patients with NHL remains uncertain. Therefore, we investigated the serial serum sIL-2R levels in 28 untreated patients with NHL to evaluate its correlation with clinical characteristics. High serum sIL-2R level (>1000 U/ml) at diagnosis was associated with a high incidence of treatment failure (p=0.03) and poor overall survival (p=0.057). The serum sIL-2R levels decreased significantly after achieving complete remission (p=0.003). Further larger studies are required to evaluate whether serum sIL-2R level is an independent prognostic factor or not. However, adding this parameter to those already employed in the International Prognostic Index would perhaps provide a better prognostic index for adult patients with NHL. PMID- 10721780 TI - Fludarabine containing-regimens may adversely affect peripheral blood stem cell collection in low-grade non Hodgkin lymphoma patients. AB - Fludarabine (FLUDA) based chemotherapy has shown promise in both initial and salvage treatment of low-grade non Hodgkin's lymphomas (LG-NHL). Recently, more aggressive therapies followed by autologous hemopoietic progenitor cell rescue, have also been successfully employed in these patients. However, this procedure, due to several factors including previous therapeutic regimens, is often limited by an inadequate collection of peripheral blood stem cell (PBSC). At present, very little data is available on the effect of FLUDA containing regimens in PBSC collection. We report our preliminary experience showing a possible correlation between FLUDA based chemotherapy regimens employed before mobilization and inability to collect an adequate number of blood derived hematopoietic progenitors for autologous PBSC transplantation in LG-NHL patients. PMID- 10721781 TI - Follicular lymphoma. A series of 11 patients with minimal or no treatment and long survival. AB - Follicular lymphoma is the commonest low-grade lymphoma. Its indolent nature even in advanced stages and the failure of conservative or aggressive treatments to achieve a cure have questioned the need for immediate treatment. Eleven patients with follicular lymphoma who had minimal or no treatment were retrospectively reviewed. Median age was 44 years. Staging was: I (4), III (6) and IV (1). Eight were confirmed to have follicular lymphoma of whom six did not receive treatment at presentation. Four of these patients remain in remission after 14 to 30 years of follow-up and the other two have relapsed after 10 and 13 years of follow-up, respectively. Two patients who were treated at diagnosis remained disease free for 18 years. Three patients had diffuse large cell lymphoma on review. They received no treatment, radiotherapy or chemotherapy and have been in remission for 36, 14 and 23 years respectively. The overall survival is 58% at 30 years, and median survival has not been reached for the whole group. Observation seems to be a valid alternative to treatment in patients with stages I to III until signs of progression. PMID- 10721782 TI - Multicenter phase II study of oral idarubicin in treated and untreated patients with B-chronic lymphocytic leukemia. AB - Idarubicin is the first anthracycline that can be administered orally facilitating antineoplastic chemotherapy at an improved quality of life. In different studies idarubicin has proved clinical effectiveness in patients with advanced low grade non Hodgkin's lymphoma. We performed a phase II study in 19 patients with untreated and pretreated B-CLL of Binet stage A-C. Idarubucin was administered orally at a dose of 15 mg/m2 over 3 days every 4 weeks. Of 19 evaluable patients (m:f, 16:3, median age 64 years, range 41-80 years) 7 were previously untreated while 12 patients had received prior therapy with fludarabine, chlorambucil or similar non-anthracycline containing regimens. 12 pts had Binet stage C, 5 Binet stage B and 2 Binet stage A. Five patients achieved a partial remission (26%), 5 patients had stable disease (26%) and 9 patients showed progressive disease (47%), resulting in an overall response of 26% (5/19). There was no correlation of response rate with Binet stages or previous treatment regimens. Treatment associated side effects consisted predominantly of mild nausea and vomiting (26%) as well as minor infections (21%) and diarrhoea (16%). These data demonstrate that oral idarubicin as a single agent is well tolerated but of limited effectiveness in B-CLL. Further studies are needed to assess different doses and schedules of oral idarubucin and to test it in combination with other chemotherapeutic agents. PMID- 10721783 TI - Incidence and pathology of primary brain lymphoma in Hong Kong Chinese patients. AB - Primary brain lymphoma (PBL) is an uncommon extranodal lymphoma. Its incidence is rapidly increasing in both immunocompromised and immunocompetent patients in Western countries. Eighteen cases of PBL were identified during a 16-year period among HIV negative patients in Queen Mary Hospital, Hong Kong. One case of post transplantation lymphoproliferative disease (PTLD) was positive for Epstein Barr virus (EBV) encoded RNA (EBER) by in situ hybridization. All the remaining 17 immunocompetent cases were classified as diffuse large B-cell lymphoma, except for one case of Burkitt's lymphoma. EBER expression was negative in all 13 cases tested. Immunostaining for bcl-2 and bcl-6 was positive in 8/11 and 6/11 cases tested, with heterogeneous combination of expression and intensity. The incidence rate of PBL in immunocompetent patients was stable at 1.03 per million per year. The incidence of PBL in post transplantation (0.16%) and HIV related setting (0.29%) is also low in Chinese. PBL in Chinese patients is almost uniformly represented by EBV negative, diffuse large B-cell lymphoma, confined to the brain. However, the molecular pathogenesis may be heterogeneous. PMID- 10721784 TI - Is mantle cell lymphoma a sex-related disease? AB - Mantle cell lymphomas are characterized by a male predominance with a range between 55 and 65 years (sex ratio M/F of 6.5). When the sex ratio of patients having mantle cell lymphoma was compared to that of each of the subtypes of non Hodgkin's lymphomas, it was significantly higher in all cases except Burkitt's and lymphoblastic T-cell lymphomas. These observations suggest a possible relation between the chromosome X and mantle cell lymphomas which has to be explored. PMID- 10721785 TI - Dexa-BEAM: an effective regimen for cytoreduction prior to high-dose chemotherapy with autologous stem cell support for patients with relapsed/refractory mantle cell lymphoma. AB - Mantle-cell lymphoma (MCL) is not a curable disease using conventional chemotherapy. Patients with MCL have the shortest median time to progression and the shortest median survival of all lymphoma subtypes after first-line treatment. In the present study we determined the efficacy of maximal cytoreductive therapy with up to four cycles of Dexa-BEAM (dexamethasone, carmustine [BCNU], etoposide, cytarabine, and melphalan) followed by high-dose chemotherapy (HDCT) and autologous hematopoietic stem cell support (ASCT) for patients with advanced relapsed or refractory MCL. Nine consecutive patients with relapsed or refractory MCL were included. Three patients had partial remission (PR), three patients progressive disease (PD) upon first line tretment, and three patients first or subsequent relapse. After 2 to four cycles of Dexa-BEAM eight patients achieved complete remission (CR), resulting in a response rate of 88%. Six of 8 patients responding to Dexa-BEAM received high-dose chemotherapy HDCT (BEAM) and autologous hematopoietic stem cell transplantation (ASCT). With a median follow up of 24 months six patients are alive. Five of those six patients are still in contiuous CR (range 13-54 months). PMID- 10721787 TI - Serologic prevalence of antibody to human herpesvirus type 8 in patients with various monoclonal gammopathies. AB - Both viral and serologic studies have consistently shown an association of human herpesvirus type 8 (HHV-8) with Kaposi's sarcoma, primary effusion lymphoma, and Castleman's disease. The presence of HHV-8 DNA in patients with myeloma has been reported by some investigators but not substantiated by others. In addition, variable results have been obtained with serologic studies for HHV-8 in patients with myeloma and certain other monoclonal gammopathies (MG). We tested 238 coded serum or plasma samples from 96 patients with various MG for antibodies to lytic and latent HHV-8 antigens by indirect immunofluorescence. Thirty-four of 96 (35%) patients were positive for the lytic antibody, but none were positive for the latent antibody. Patients with kappa or lambda light chain myeloma were often positive for the lytic antibody when compared to patients with IgG or IgA myeloma (8 of 11 [73%] vs. 12 of 38 [32%], P = 0.033). The patients with light chain myeloma also were more likely to be positive when compared to patients with Waldenstrom's macroglobulinemia (WM) (4 of 15 [27%], P = 0.045) or AL amyloidosis (4 of 13 [31%], P = 0.047). Four of 9 (44%) patients with monoclonal gammopathy of undetermined significance (MGUS) were positive. However, 4 other patients who progressed from MGUS to myeloma were negative. Subgroup analysis of MG may help clarify the role of HHV-8 in these disorders. PMID- 10721786 TI - Plasma erythropoietin concentrations in polycythaemia vera with special reference to myelosuppressive therapy. AB - In 80 patients with polycythaemia vera (PV) a total of 108 venous blood samples were obtained and analysed for EDTA-plasma erythropoietin (EPO) concentration. At the time of study 21 of the PV patients were newly diagnosed and had prior to blood sampling neither received phlebotomy treatment nor therapy with myelosuppressive agents; these subjects had a mean plasma EPO concentration of 0.5+/-0.9 IU/L. Thirty-seven patients treated with phlebotomy only had a mean plasma EPO concentration of 2.5+/-2.9 IU/L. The mean plasma EPO concentrations for 26 patients treated with hydroxyurea, 13 patients treated with radiophosphorous and 11 patients given a combination of myelosuppressive agents were 8.9+/-8.0, 10.9+/-12.6 and 7.2+/-7.4 IU/L, respectively. Untreated patients and patients on phlebotomy only had significantly lower values for plasma EPO than patients on therapy with myelosuppressive drugs. This finding persisted also after a correction for differences in haemoglobin levels had been introduced. Thereby, the present results would suggest a difference in the EPO feedback system in untreated and phlebotomised PV patients compared to PV patients treated with myelosuppressive agents. PMID- 10721788 TI - Detection of BCR-ABL transcripts by multiplex and nested PCR in different haematological disorders. AB - Single-step Multiplex RT-PCR was used as a rapid and highly sensitive method for screening patients with myeloproliferative conditions and ALL for the presence of underlying BCR-ABL gene fusions. Positive and negative results obtained with the multiplex assay were subsequently confirmed by nested PCR. We studied 21 patients for detecting the presence of b3a2, b2a2 and e1a2 BCR-ABL transcripts at diagnosis and following treatment with different therapeutical procedures. These studies allowed the molecular characterisation of patients with different haematological disorders and for demonstrating BCR-ABL transcripts in Ph-CML. In a Ph+ CML patient, a switch of isoforms was detected after bone marrow transplantation and infusion with donor lymphocytes, implying substitution of e1a2 for b3a2 coexisting with a myeloid/lymphoid biphenotypic profile. In ALL, one Ph+ patient showed coexpression of e1a2 and b2a2 at diagnosis followed by persistence of e1a2 after bone marrow transplantation. Our results were compared to previous findings in the literature on molecular diagnosis of leukaemias. PMID- 10721789 TI - Trisomy of the long arm of chromosome 1 resulting in a dicentric derivative (6)t(1;6) chromosome in a child with myelodysplastic syndrome following treatment for a primitive neuroectodermal tumor. AB - We report the clinical, hematologic, and cytogenetic findings for a child with secondary myelodysplastic syndrome (MDS) after treatment for a primitive neuroectodermal tumor. At the time of conversion to MDS, conventional cytogenetics revealed an unbalanced der(6)t(1;6) that resulted in trisomy of the long arm of chromosome 1 and partial monosomy and duplication of 6p. Using alpha satellite probes, fluorescence in situ hybridization of bone marrow cells showed that the rearranged chromosome contained the centromeres of both chromosomes 1 and 6, thus forming a dic(1;6) resulting in trisomy 1q. This report is the first to describe a case of childhood secondary myelodysplastic syndrome associated with a trisomy 1q involving chromosome 6. PMID- 10721790 TI - Intrasinusoidal bone marrow infiltration revealing intravascular lymphomatosis. AB - Intravascular lymphoma is a rare, often fatal disease characterized by a widespread intravascular proliferation of neoplastic lymphoid cells. Dermatological and bizarre neurological manifestations usually predominate. We report a case of intravascular lymphomatosis with an exceptional clinical presentation showing splenomegaly combined with early bone marrow involvement. The diagnosis was made on bone marrow biopsy examination using both immunohistochemistry and molecular biology analysis. We stress the histopathological features of bone marrow involvement by intravascular lymphoma which allow the prompt recognition of this disease. Early systemic chemotherapy, which represents the only chance of remission in such an aggressive disease, can then be initiated. PMID- 10721791 TI - The first report of familial adult T-cell leukemia/lymphoma in Argentina. AB - Here we describe two Caucasian brothers who developed adult T-cell leukemia/lymphoma (ATLL), within a short period of time. These two patients have never left Argentina. Their parents are dead and according to the family history it is possible that the mother may have been affected by spastic paraparesis. The daughters reported that their mother had suffered from increasing difficulty in walking for many years which finally made it impossible for to her walk. There are no other data to support the presumptive diagnosis. One of the patients presented with acute disease while the other had a lymphoma type disorder. Both were positive for HTLV 1. The first patient died with disease progression ten months after diagnosis and the second is in partial remission 13 months after diagnosis. Immunophenotyping showed CD4+, CD5+, CD3+, CD2+, CD8 (-). Two asymptomatic brothers with positive HTLV 1 serology were detected. This is the first family case that has been reported in Argentina. PMID- 10721792 TI - Lymphomatous skin infiltration at the site of previous varicella zoster virus infection in a patient with T cell lymphoma. AB - Cutaneous infiltrations in hemopoietic neoplasias are not uncommon. They are generally localized on the legs, arms, back, anterior chest, scalp and face. In rare cases specific infiltration of neoplastic cells is localized in the site of herpes zoster and herpes simplex scars. In this report a case with T cell lymphoma in leukemic phase with skin infiltration in the previous Varicella Zoster Virus (VZV) site of infection is reported and literature is reviewed. PMID- 10721793 TI - Effects of mutant c-kit in early myeloid cells. AB - Activating mutations in c-Kit, the receptor for Stem Cell Factor (SCF), have been identified in dysplasias and leukaemias of the mast cell lineage and have been shown to contribute to transformation in model systems. Early myeloid cells also normally express c-Kit and their survival, proliferation and differentiation is promoted by SCF. It might therefore be expected that c-Kit mutations could also be involved in some acute and/or chronic myeloid leukaemias. We have found that mutant c-Kit (and normal c-Kit in the presence of SCF) provides a strong differentiation stimulus in normal and immortalised murine early myeloid cells. Since maturation of haemopoietic cells, with the exception of mast cells, results in down-regulation of c-Kit expression, the transforming effects of mutant receptor may be self-limiting in most lineages. This is consistent with the observation that multipotential progenitor cells from some patients with systemic mastocytosis express mutant c-Kit. However, c-Kit mutations have been observed in a few cases of myelodysplastic syndromes or AML without mast cell features. Oncogenesis involves multiple genetic changes and the phenotype of malignant haemopoietic cells expressing mutant c-Kit may be influenced by co-oncogenic events. For example mutations blocking the differentiative effect of mutant c-Kit might result in AML rather than mastocytosis. Thus the extent to which c-Kit mutations contribute to malignancies of early myeloid phenotype remains unknown, and resolution of this issue is complicated by the heterogeneity of this family of diseases. PMID- 10721794 TI - Lectins for gastrointestinal targeting--15 years on. AB - In the mid-1980s, the concept of bioadhesion using synthetic polymers emerged, and brought with it the promise of improved efficiency for the delivery of drugs via mucosal surfaces. Studies in the author's laboratory concentrated on 'biological' bioadhesion using the naturally-occurring proteins, lectins, which recognise and bind sugars in glycoconjugates, such as those found on the surfaces of cells. Tomato Lectin (TL) was extensively studied as a putative non-toxic lectin with potential for drug targeting/delivery to the gastrointestinal (GI) tract. In vitro, the TL displayed impressive binding to the intestinal mucosa, but in vivo failed to significantly modify intestinal transit. A number of research groups have coupled the TL to microparticles, and significant systemic uptake of these has been observed in animal studies. Polymers with pendant sugars have also been shown to be bioadhesive, by interacting with endogenous lectins present on the cells of the GI tract. The use of lectins to target to Peyer's patches and diseased tissues in the colon is an interesting development, but much work remains to be done. Lectins also have potential in mucosal vaccines. Before advanced drug delivery systems using lectins can be realised, rigorous evaluation of their toxicity and immunogenicity will be required, but they clearly offer a number of possibilities for GI drug targeting systems in the future. PMID- 10721795 TI - Rates of protein transport across rat alveolar epithelial cell monolayers. AB - The transport of model proteins, ranging from 12,300 to 150,000 Da, across tight rat alveolar epithelial cell monolayers (> 2000omegacm2) grown on polycarbonate filters, was studied. Model proteins were 14C-cytochrome c, 14C-ovalbumin, granulocyte-colony stimulating factor (G-CSF), 14C-bovine serum albumin (BSA), 125I-transferrin, and 14C-immunoglobulin G. Cytochrome c was extensively metabolized, as indicated by < 10% of the dose being translocated in intact form. This contrasts with 20-80% for the other model proteins studied. The flux of cytochrome c and G-CSF was symmetric in the apical-to-basolateral (ab) and basolateral-to-apical (ba) directions. By contrast, the flux of intact ovalbumin, BSA, transferrin and immunoglobulin G showed asymmetry, with the ab flux being higher by 2-5 times. There was no relationship between ab or ba fluxes and the molecular weights of these four model proteins. Since some of the proteins were translocated at much greater rates than are consistent with restricted diffusion or pinocytosis, receptor-mediated or adsorptive transcytosis may be involved. PMID- 10721796 TI - PLGA microspheres phagocytosis by pig alveolar macrophages: influence of poly(vinyl alcohol) concentration, nature of loaded-protein and copolymer nature. AB - The study aimed to investigate on a pig alveolar macrophage culture model the influence of: (1) poly(D,L-lactide-co-glycolide) characteristics, (2) the residual poly(vinyl alcohol) (PVA) and (3) the nature of encapsulated proteins, immunoglobulin Y (IgY) or bovine serum albumin, on the microspheres phagocytosis efficiency. The phagocytosis evaluation was performed by flow cytometry and has allowed a screening of microspheres formulations. The hydrophilicity of microspheres resulting from the nature of the polymer and/ or from the residual hydrophilic surface agent (PVA) led to a decrease of phagocytosis intensity. The phagocytosis results of IgY-loaded microspheres strongly suggested that the phagocytosis was increased when the phagocytic cell possessed a receptor for this protein on its surface. PMID- 10721797 TI - Improved nasal bioavailability of FITC-dextran (Mw 4300) from mucoadhesive microspheres in rabbits. AB - The nasal bioavailability of fluorescein isothiocyanate-dextran (FITC-dextran) (Mw = 4300) encapsulated in non-mucoadhesive and mucoadhesive microspheres in New Zealand White rabbits was investigated. FITC-dextran was administered nasally encapsulated in carbopol 934P, chitosan and lactose microspheres and the bioavailability compared to intravenous administration of FITC-dextran solution. Administration of FITC-dextran as carbopol microspheres produced a significantly greater bioavailability (33%) than after administration as chitosan (13%) and non mucoadhesive rapidly dissolving control lactose microspheres (9%). The FITC dextran terminal plasma half-lives after carbopol 934P and chitosan microsphere administration were significantly longer than after intravenous administration of FITC-dextran. The FITC-dextran terminal plasma half-life after carbopol 934P microspheres administration was significantly longer than after lactose microsphere administration. This data suggested that the increase in FITC-dextran bioavailability after carbopol 934P microspheres administration was due to increased residence at the absorptive site via mucoadhesion and reduced mucociliary clearance. A change in mucosal permeability could not however be discounted especially for the chitosan microspheres. PMID- 10721798 TI - Effect of cationic liposomes on intracellular trafficking and efficacy of antisense oligonucleotides in mouse peritoneal macrophages. AB - We have investigated the intracellular fate and antisense effect of oligonucleotide/cationic liposome complexes using phosphorothioate oligonucleotides (S-Oligo) targeted to inducible nitric oxide synthase in mouse peritoneal macrophages. Confocal laser microscopic analysis revealed that, after application of fluorescein isothiocyanate (FITC)-labeled S-Oligo alone, the intracellular localization of fluorescence exhibited a punctate pattern in the cytoplasm, suggesting that the oligonucleotides were mainly confined to the endosomal and/or lysosomal compartments. In the case of complexation with Lipofectin and DMRIE-C liposomes, cellular uptake of FITC-S-Oligo was not greatly enhanced and the fluorescence localization in the cells was similar to that of FITC-S-Oligo alone. LipofectAMINE slightly enhanced cellular uptake of FITC-S Oligo; however, the intracellular localization profile of FITC-S-Oligo remained largely unchanged. The antisense effect was slightly enhanced by LipofectAMINE under only very limited experimental conditions. It was concluded that cationic liposomes are not a potential carrier for S-Oligo in peritoneal macrophages because of their inability to promote the release of S-Oligo from the endosomal compartments to the cytosol over a non-toxic concentration range. PMID- 10721799 TI - Studies on uptake, sub-cellular trafficking and efflux of antisense oligodeoxynucleotides in glioma cells using self-assembling cationic lipoplexes as delivery systems. AB - The cellular uptake of antisense oligodeoxynucleotides (ODNs) may be enhanced by the use of carriers such as cationic liposomes or lipoplexes, but little is known about the intracellular fate and subcellular trafficking of these systems in target cells. In this study, we report on the cellular uptake and biodistribution of ODNs in the presence and absence of optimised self-assembled cationic lipoplexes using the C6 glioma cell line as an in vitro model. Biotin or radiolabelled 15-mer phosphorothioate (PS) ODNs were synthesised and their cellular uptake and subcellular biodistribution characterised in the presence and absence of an optimised cationic lipoplex delivery system using studies ranging from cellular association, cellular efflux and transmission electron microscopy (TEM). Ultrastructural studies clearly showed PS ODNs in the absence of liposomal delivery to be sequestered within endosomal and lysosomal vesicular bodies indicative of endocytic uptake. ODNs were also visible, to a lesser extent, in the nucleus and cytoplasm. By employing DOSPA (2'-(1",2" dioleoyloxypropyldimethyl-ammonium bromide)-N-ethyl-6-amidospermine tetra trifluoroacetic acid) and DOPE (dioleoylphosphatidylethanolamine) complex in a 3 : 1 ratio, as a delivery system for ODNs at a optimal lipid/DNA charge ratio of 1 : 1, the level of ODN cellular association was significantly increased by approximately 10-12 fold with a concomitant change in subcellular distribution of PS ODN. TEM studies indicated enhanced penetration of ODN within the cytosol and the cell nucleus with reduced presence in vesicular compartments. Efflux studies confirmed that cationic lipoplexes promoted entry of ODNs into 'deeper' cellular compartments, consistent with endosomal release. Optimised cationic lipoplexes improved cellular delivery of ODNs by enhancing cell association, uptake and by favourably modulating the intracellular trafficking and distribution of ODNs into non-vesicular compartments including the cytosol and nucleus. PMID- 10721800 TI - Influence of coenzyme Q10 on tissue distribution of archaeosomes, and pegylated archaeosomes, administered to mice by oral and intravenous routes. AB - The biodistribution of orally and intravenously administered archaeosomes in mice was compared to that of archaeosomes containing either coenzyme Q10 (archaeosome CoQ10), polyethylene glycol (archaeosome-PEG), or PEG plus CoQ10 (archaeosome-PEG CoQ10). The archaeosome formulations were prepared by a reverse-phase evaporation method using the total polar lipids from the archaeobacterium Methanosarcina mazei. In the case of oral gavage, the most striking observation was that a significantly (p < 0.05) higher concentration (42.28+/-4.17%) of administered dose was found in the stomach content 3 h after administration of unmodified archaeosomes, as compared to that of archaeosome-CoQ10 (16.98+/-2.48%) and archaeosome-PEG-CoQ10 (5.8 +/-4.05"/ vesicles. This correlated with an increased uptake, notably of the archaeosome-PEG-CoQ 0 vesicles.,into liver and spleen; however, no more than 7% of the administered dose was found in liver, spleen and blood at any time point studied. In the case of intravenous administration, a significantly higher percentage of injected dose of unmodified archaeosomes was found in the liver (66.4 +/-.92%) and spleen (11.445+/-.68%) at 48 h, compared to archaeosome-CoQ10, archaeosome-PEG, and archaeosome-PEG-CoQ10 vesicles. The combination of PEG and CoQ10 significantly prolonged the circulation of archaeosomes in the blood, but after 48 h the amount of the vesicle marker in blood had declined to only about 0.5% of administered dose. These data indicate that the biodistribution of archaeosome formulations given orally or intravenously can be altered significantly by incorporating PEG or CoQ 10, alone or in combination, and these vesicles have the potential to act as a carrier for therapeutics and vaccines. PMID- 10721801 TI - Synthesis, characterisation and in vivo behaviour of a norfloxacin-poly(L-lysine citramide imide) conjugate bearing mannosyl residues. AB - With the aim of promoting the targeting of macrophage mannose receptors and the internalisation of the norfloxacin antibiotic, which is active against some intracellular bacteria, a macromolecular prodrug was synthesised where the antibiotic and mannosyl moieties were coupled to a polymeric carrier, namely poly(L-lysine citramide imide). This carrier, which derived from two metabolites, citric acid and L-lysine, is known to be biocompatible and slowly degradable under slight acidic conditions. Norfloxacin was coupled onto the acid groups present along the polymer chains, and conjugates were characterised by UV, TLC and SEC. The mannosyl groups selected to promote the targeting of the mannose specific lectin present on the outer membrane of macrophages were incorporated through a biodegradable glycolic spacer arm. Two different strategies were considered to synthesise the full conjugates, namely coupling norfloxacin onto mannosylated conjugates, and coupling mannose onto PLCAI/Nflx conjugates. The second pathway led to better results regarding mannosylation. The presence of norfloxacin and mannose caused chain aggregation, especially for conjugates with a high content of mannosyl residues. The targeting ability of the prodrug was investigated using a method based on the competition between the mannosylated macromolecules and glucose oxidase, a mannosyl-bearing non-human protein. This method showed that prodrug macromolecules competed effectively with glucose oxidase and thus should be able to bring the drug up to the mannosyl receptor bearing membranes of macrophages infected by intracellular bacteria. PMID- 10721802 TI - Cystic lesions of the pancreas. Introduction. PMID- 10721803 TI - Pseudocysts and other non-neoplastic cysts of the pancreas. AB - Among the cystic lesions of the pancreas, pseudocysts are most common. This article summarizes the current state of knowledge on the morphology and pathogenesis of pseudocysts. Specifically, the pathogenetic relationship between pancreatic pseudocysts and acute and chronic pancreatitis and the natural history of pseudocysts will be discussed. In addition, this article gives a brief description of a few non-neoplastic pancreatic cysts, including retention cysts, congenital cysts, cystic acinar transformation, para-ampullary duodenal wall cysts, enterogenous cysts, endometrial cysts, and parasitic cysts. PMID- 10721804 TI - Pancreatic tumors with cystic dilatation of the ducts: intraductal papillary mucinous neoplasms and intraductal oncocytic papillary neoplasms. AB - Intraductal papillary mucinous neoplasms (IPMNs) and intraductal oncocytic papillary neoplasms (IOPNs) are the 2 types of intraductal neoplasms of the pancreas that may appear cystic because of dilatation of the ducts. Both are characterized by intraductal proliferation of mucinous cells usually arranged in papillary patterns. This proliferation is often associated with intraluminal mucin accumulation, which produces cystic dilatation of the ducts, mimicking mucinous cystic neoplasms. Endoscopic and radiologic studies and careful macroscopic examination are crucial for the correct diagnosis of IPMNs and IOPNs by showing the origin within the native ducts. Microscopically, these tumors display a spectrum of cytoarchitectural atypia that ranges from adenoma to borderline and to carcinoma-in-situ. Although they are defined as "intraductal tumors," IPMNs and IOPNs are associated with invasive carcinoma in about a third of the cases. It, therefore, appears that, like mucinous cystic neoplasms or pancreatic intraepithelial neoplasia involving the smaller ducts associated with ordinary ductal adenocarcinomas, these tumors are precursors of invasive carcinoma. Invasive carcinomas associated with IPMNs are of either tubular or colloid (mucinous noncystic) types, whereas those associated with IOPNs may be oncocytic. Even in the presence of invasive carcinoma, these tumors may follow a more protracted clinical course than ordinary ductal adenocarcinoma. On the other hand, rare examples of IPMNs after an aggressive clinical course despite the lack of any identifiable invasive carcinoma are on record. Therefore, IPMNs and IOPNs should be examined carefully and sampled extensively, first, to confirm that the main pathology is an intraductal process and, more importantly, to rule out the presence of an invasive carcinoma. PMID- 10721806 TI - Serous cystic tumors of the pancreas. AB - Serous cystic tumors of the pancreas are unique among pancreatic cystic neoplasms in that they are almost always cytologically monomorphous and biologically benign. The diagnostic challenge for the surgical pathologist is to recognize the spectrum of architectural variation that may lead to misdiagnosis, either preoperatively or postoperatively, and the additional lesions and conditions with which serous tumors may be associated. PMID- 10721805 TI - Mucinous cystic neoplasms of the pancreas. AB - Since their initial description, mucinous cystic neoplasms have been difficult to classify. This article attempts to clarify histological, clinical, and genetic criteria so that the pathologist can categorize each mucinous cystic neoplasm into 1 of 4 possible categories. Mucinous cystadenomas contain a single layer of mucin-producing, columnar epithelium lacking significant atypia. Borderline mucinous cystic neoplasms contain cells with moderate atypia. Mucinous cystic neoplasms with in situ carcinoma show significant architectural and cytological atypia. When invasive carcinoma is present in association with a mucinous cystic neoplasm, then the diagnosis of invasive mucinous cystadenocarcinoma should be made. The categorization of mucinous cystic neoplasms into these groups is essential because it accurately predicts outcome, provided that the tumor has been sampled and examined thoroughly. Completely removed mucinous cystadenomas, borderline mucinous cystic neoplasms, and mucinous cystic neoplasms with in situ carcinoma follow benign courses. Partial resection should be avoided as evidence suggests that mucinous cystic neoplasms can progress from adenomas to borderline lesions to carcinomas in situ to invasive carcinomas over time; partial resection should be avoided if possible. Modern molecular genetic techniques are helping to unravel the origins of rare variants of mucinous cystic tumors, such as the mucinous cystic tumor with an associated osteoclast-like giant cell tumor and the mucinous cystic tumor with sarcomatous stroma. PMID- 10721807 TI - Squamous-lined cysts of the pancreas: lymphoepithelial cysts, dermoid cysts (teratomas), and accessory-splenic epidermoid cysts. AB - In the pancreas, 3 types of morphologically similar lesions may present as "squamous cysts": Lymphoepithelial cysts, dermoid cysts (monodermal teratomas), and epidermoid cysts in intrapancreatic accessory spleen. Lymphoepithelial cysts (LECs) are seen predominantly in men (M/F: 4/1) and in adulthood (mean age, 56, and range, 35 to 74 years). They may occur at any site of the organ (head, body, or tail). LECs are well-delineated cysts that may be multilocular (60%) or unilocular (40%), and they are characterized microscopically by stratified squamous epithelium surrounded by a band of mature lymphoid tissue with intervening well-formed germinal centers. Solid lymphoepithelial clusters are seldom seen. The pathogenesis of LECs is unclear; clinical diseases that are known to be associated with their counterparts in the salivary glands such as Sjogren disease or human immunodeficiency virus have not been documented for the LECs of the pancreas. The second type of squamous-lined cyst in the pancreas is the epidermoid cyst arising in intrapancreatic accessory spleen. These are located almost exclusively in the tail of the pancreas, in the fourth decade of life (mean age = 38). Their mean size is 4.5 cm (range, 2.3 to 6.5). In some cases, the cyst lining may be partly mucinous. Dermoid cysts of the pancreas are also rare. The cases that appear to be true dermoid cysts occur in a younger age group (mean age, 23, range, 2 to 53 years), and in contrast with LEC, there is no gender predominance. Mucinous epithelium, respiratory-type mucosa and sebaceous units are more readily identifiable in dermoid cysts, and they may contain hair. Subepithelial lymphoid tissue is not a feature. They are sometimes complicated by suppurative infections. The importance of these lesions is in their distinction from other cystic neoplasms, especially mucinous cystic tumors. PMID- 10721808 TI - Solid-pseudopapillary tumor of the pancreas: a typically cystic carcinoma of low malignant potential. AB - Solid-pseudopapillary tumors are uncommon neoplasms of low malignant potential generally occurring in young women. They often cause few symptoms and may reach a large size by the time they are detected. Degenerative cystic changes are common, and the clinical presentation is often that of a cystic pancreatic tumor. Pathological features include solid, cellular, hypervascular regions without gland formation, and degenerative pseudopapillae. The cells contain eosinophilic granules rich in alpha-1-antitrypsin and the nuclei are typically grooved. Despite its characteristic microscopic appearance, the immunophenotype (positive for vimentin, alpha-1-antitrypsin, and neuron specific enolase) is not specific and does not define a line of differentiation corresponding to any normal pancreatic cell type. Ultrastructural studies have also failed to identify specific differentiated features. Nonetheless, the biological behavior of solid pseudopapillary tumor is well established. The tumor is indolent, with infrequent metastases to liver or peritoneum and usually long survival, even in the presence of disseminated disease. PMID- 10721809 TI - Cystic forms of typically solid pancreatic tumors. AB - In addition to the well-known cystic lesions, the differential diagnosis of pancreatic cysts also includes cystic counterparts (or cystic change in) otherwise typically solid tumors of this organ. Pancreatic endocrine neoplasms (islet cell tumors) and ductal adenocarcinomas sometimes undergo cystic degeneration, the latter usually due to necrosis. Also, although acinar cell carcinoma is typically a solid tumor, its rare cystic counterpart, "acinar cell cystadenocarcinoma," has been reported. A rare variant of pancreatic ductal adenocarcinoma referred to as "large-duct-type" is characterized by microcystic ectasia of the invasive glands and may mimic other cystic tumors at the microscopic level. Secondary tumors, although exceedingly rare, may also potentially fall into the differential diagnosis of pancreatic cysts. Often, the morphological characteristics of these lesions are identical to those of their solid counterparts, and their diagnosis is not difficult if one is aware of their existence. This review focuses on these cystic counterparts of otherwise characteristically solid pancreatic neoplasms. Solid pseudopapillary tumor, in which cystic degeneration is so common that "cystic" has been a part of many alternate terms assigned to this neoplasm, is discussed in another article of this issue of Seminars in Diagnostic Pathology. PMID- 10721811 TI - Considerations on pharmacodynamics and pharmacokinetics: can everything be explained by the extent of drug binding to its receptor? AB - It is frequently assumed that pharmacological responses depend solely on the extent of drug binding to its receptor according to the occupational theory. It is therefore presumed that the intensity of the effect is determined by drug concentration at its receptor site, yielding a unique concentration-effect relationship. However, when dependence, abstinence, and tolerance phenomena occur, as well as for pharmacological responses in vivo that are modulated by homeostatic mechanisms, the rate of drug input shifts the concentration-effect relationship. Hence, such responses cannot be explained on the sole basis of the extent of drug binding to its receptor. Information on the cellular and molecular processes involved in the generation of abstinence, dependence, and tolerance will undoubtedly result in the development of pharmacodynamic models allowing a satisfactory explanation of drug effects modulated by these phenomena. Notwithstanding, integrative physiology concepts are required to develop pharmacokinetic-pharmacodynamic models allowing the description of drug effects in an intact organism. It is therefore important to emphasize that integrative physiology cannot be neglected in pharmacology teaching and research, but should be considered as an equally valuable tool as molecular biology and other biomedical disciplines for the understanding of pharmacological effects. PMID- 10721810 TI - Heterotrimeric G proteins in heart disease. AB - Guanine nucleotide binding proteins (G proteins) are largely grouped into three classes: heterotrimeric G proteins, ras-like or small molecular weight GTP binding proteins, and others like Gh. In the heart G proteins transduce signals from a variety of membrane receptors to generate diverse effects on contractility, heart rate, and myocyte growth. This central position of G proteins forming a switchboard between extracellular signals and intracellular effectors makes them candidates possibly involved in the pathogenesis of cardiac hypertrophy, heart failure, and arrhythmia. This review focuses primarily on discoveries of heterotrimeric G protein alterations in heart diseases that help us to understand the pathogenesis and pathophysiology. We also discuss the underlying molecular mechanisms of heterotrimeric G protein signalling. PMID- 10721812 TI - Opposite change with ischaemia in the antifibrillatory effects of class I and class IV antiarrhythmic drugs resulting from the alteration in ion transmembrane exchanges related to depolarization. AB - It is known that class I antiarrhythmic drugs lose their antifibrillatory activity with severe ischaemia, whereas class IV antiarrhythmic drugs acquire such activity. Tachycardia, which is also a depolarizing factor, has recently been shown to give rise to an alteration of ion transmembrane exchanges which is particularly marked in the case of calcium. This leads one to wonder if the change in antifibrillatory activity of antiarrhythmic drugs caused by ischaemia depends on the same process. The change in antifibrillatory activity was studied in normal conditions ranging to those of severe ischaemia with a class I antiarrhythmic drug, flecainide (1.00 mg x kg(-1) plus 0.04 mg x kg(-1)x min(-1), a sodium channel blocker, and a class IV antiarrhythmic drug, verapamil (50 microg x kg(-1) plus 2 microg x kg(-1) x min(-1)), a calcium channel blocker. The experiments were performed in anaesthetized, open-chest pigs. The resulting blockade of each of these channels was assessed at the end of ischaemic periods of increasing duration (30, 60, 120, 180, 300, and 420 s) by determining the ventricular fibrillation threshold (VFT). VFT was determined by means of trains of diastolic stimuli of 100 ms duration delivered by a subepicardial electrode introduced into the myocardium (heart rate 180 beats per min). Ischaemia was induced by completely occluding the left anterior descending coronary artery. The monophasic action potential was recorded concurrently for the measurement of ventricular conduction time (VCT). The monophasic action potential duration (MAPD) varied with membrane polarization of the fibres. The blockade of sodium channels by flecainide, which normally raises VFT (7.0 +/- 0.4 to 13.8 +/- 0.8 mA, p < 0.001) and lengthens VCT (28 +/- 3 to 44 +/- 5 ms, p < 0.001), lost its effects in the course of ischaemia. This resulted in decreased counteraction of the ischaemia-induced fall of VFT and decreased aggravation of the ischaemia induced lengthening of VCT. The blockade of calcium channels, which normally does not alter VFT (between 7.2 +/- 0.6 and 8.4 +/- 0.7 mA, n.s.) or VCT (between 30 +/- 2 and 34 +/- 3 ms, n.s.), slowed the ischaemia-induced fall of VFT. VFT required more time to reach 0 mA, thus delaying the onset of fibrillation. Membrane depolarization itself was opposed as the shortening of MAPD and the lengthening of VCT were also delayed. Consequently there is a progressive decrease in the role played by sodium channels during ischaemia in the rhythmic systolic depolarization of the ventricular fibres. This reduces or suppresses the ability of sodium channel blockers to act on excitability or conduction, and increases the role of calcium channel blockers in attenuating ischaemia-induced disorders. PMID- 10721813 TI - Nitric oxide, prostaglandins, and impaired cerebral blood flow autoregulation in group B streptococcal neonatal meningitis. AB - Impaired autoregulation of cerebral blood flow (CBF) contributes to CNS damage during neonatal meningitis. We tested (i) the hypothesis that cerebrovascular autoregulation is impaired during early onset group B streptococcal (GBS) meningitis, (ii) whether this impairment is regulated by vasoactive mediators such as prostaglandins and (or) nitric oxide (NO), and (iii) whether this impairment is preventable by specific and (or) nonspecific inhibitors: dexamethasone, ibuprofen, and Nomega-nitro-L-arginine, a NO inhibitor. Sterile saline or 10(9) colony-forming units (cfu) of heat-killed GBS was injected into the cerebral ventricle of newborn piglets. CBF autoregulation was determined by altering cerebral perfusion pressure (CPP) with balloon-tipped catheters placed in the aorta. GBS produced a narrow range of CBF autoregulation due to an impairment at the upper limit of CPP. We report that in vivo in the early stages (first 2 h) of induced GBS inflammation (i) GBS impairs the upper limit of cerebrovascular autoregulation; (ii) ibuprofen, dexamethasone, and Nomega-nitro-L arginine not only prevent this GBS-induced autoregulatory impairment but improve the range of cerebrovascular autoregulation; (iii) these autoregulatory changes do not involve circulating cerebral prostanoids; and (iv) the observed changes correlate with the induction of NO synthase gene expression. Thus, acute early onset GBS-induced impairment of the upper limit of CBF autoregulation can be correlated with increases of NO synthase production, suggesting that NO is a vasoactive mediator of CBF. PMID- 10721814 TI - Levels of specifically bound [3H]ketanserin compared with levels of serotonin (5HT) in the brain regions of juvenile and sexually recrudescing female rainbow trout, Oncorhynchus mykiss. AB - This study compared the distribution of specifically bound [3H]ketanserin (Bsp) with serotonin (5HT) in brain regions of juvenile and sexually recrudescing female trout. Amounts of Bsp varied widely among brain regions and consistently differed between juvenile and sexually recrudescing females. Levels of Bsp were significantly greater in the hypothalamus than the olfactory lobe, which were at least threefold greater than in all other tissues examined (Kruskal-Wallis test, p < 0.05). Bsp densities in the hypothalamus, preoptic area, and optic lobe were significantly greater in juveniles compared with corresponding tissues from sexually recrudescing females (Mann-Whitney U test, p < 0.05); in contrast, Bsp in olfactory lobe and spinal cord did not differ significantly between the two classes of fish. 5HT concentration was determined by high performance liquid chromatography - electrochemical detection (HPLC-EC) analysis. Biogenic amine standards eluted in a stereotypic pattern, with peaks consistently separable in time. 5HT concentration was significantly greater in hypothalamus than in olfactory lobe and undetectable in the pituitary (Kruskal-Wallis test, p < 0.05). Trends in distribution of Bsp and 5HT were comparable in the hypothalamus and preoptic area in juvenile and sexually recrudescing females. In general, density of specific [3H]ketanserin binding sites was directly related to 5HT content of brain regions in juvenile and sexually recrudescing females. 5HT concentrations (pmol/g tissue) were approximately 900-fold greater than Bsp (fmol/g tissue) in all brain regions, and approximately 300-fold greater than Bsp in the olfactory lobe. These results suggest important regulatory role(s) for 5HT in the trout preoptic-hypothalamo-hypophysial axis, which may differ from 5HT role(s) in trout olfactory lobe. PMID- 10721815 TI - Distribution of alpha1-adrenoceptor subtype proteins in different tissues of neonatal and adult rats. AB - Distribution of alpha(1)-adrenoceptor (alpha(1)AR) subtype (alpha(1A), alpha(1B), alpha(1D)) proteins in brain, heart, kidney, and liver of 1-week-old rats and in brain, heart, aorta, kidney, liver, vas deferens, prostate, and adrenal glands of adult rats was investigated by Western analysis, using receptor subtype specific polyclonal antibodies. High levels of immunoreactive alpha(1A)AR and alpha(1D)AR in brain and heart and of alpha(1B)AR in liver and heart of neonatal rats were detected. In adult rat tissues, the abundance of alpha(1A)AR protein was most marked in the brain, intermediate in heart, aorta, liver, vas deferens, and adrenals, and minimal in the kidney and prostate; relative to other tissues, the expression of alpha(1B)AR was higher in brain and heart and that of alpha(1D)AR in brain. All the three receptor subtypes increased with age in the brain cortex, whereas the abundance of alpha(1B)AR increased in the heart but decreased in the liver; alpha(1A)AR and alpha(1D)AR in liver, kidney, and heart were not affected by age. It is concluded that alpha(1)AR subtypes are widely expressed in different neonatal and adult rat tissues. PMID- 10721816 TI - Acute plasma volume expansion alters cardiovascular but not thermal function during moderate intensity prolonged exercise. AB - To investigate the hypothesis that the increase in plasma volume (PV) that typically occurs with training results in improved cardiovascular and thermal regulation during prolonged exercise, eight untrained males (V(O2)peak = 3.52 +/- 0.12 L x min(-1)) performed 90 min of cycle ergometry at 62% V(O2)peak before and after acute PV expansion. Subjects were infused with a PV-expanding solution (dextran (6%) or Pentaspan (10%)) equivalent to 6.7 mL x kg(-1) body mass (PVX) or acted as their own control (CON) in a randomized order. PVX resulted in a calculated 15.8% increase in resting PV, which relative to CON, was maintained throughout the exercise (P < 0.05). During PVX, heart rate was lower (P < 0.05) and stroke volume and cardiac output were higher (P < 0.05) during the exercise. Mean arterial pressure and total peripheral resistance, although altered by exercise (P < 0.05), were not different between the two conditions. Core temperature, which was progressively increased by the exercise (P < 0.01), was not affected by PVX. A similar decrease in body weight was observed between the conditions as a result of the exercise (P < 0.01). These results indicate that acute PVX alters cardiovascular performance without affecting the thermoregulatory response to prolonged cycle exercise. PMID- 10721817 TI - Vasodilation in human subcutaneous arteries induced by neuropeptide Y is mediated by neuropeptide Y Y1 receptors and is nitric oxide dependent. AB - Neuropeptide Y (NPY) is known as a potent vasoconstrictor of peripheral blood vessels both in vivo and in vitro. There have been reports suggesting that NPY also has a dilatory effect. The aim of the present study was to elucidate whether NPY dilates small human subcutaneous arteries. Subcutaneous arteries, obtained from patients undergoing abdominal surgery, were mounted in in vitro tissue baths, and the vascular responses to NPY were investigated. The presence of mRNA encoding the human NPY Y1 receptor in endothelial cells from human umbilical veins was studied by the use of reverse transcriptase - polymerase chain reaction (RT-PCR). In arteries precontracted with the prostaglandin analogue U46619, NPY induced a concentration-dependent vasodilation (Emax 30 +/- 10% of the U46619 induced contraction), which was significantly inhibited by the NPY Y1 receptor antagonist BIBP3226 (1 microM), causing a rightward shift of the concentration response curve, pEC50 7.1 +/- 0.3 vs. 7.7 +/- 0.3 for NPY alone. After pretreatment with the nitric oxide synthetase inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME) (10 microM), the dilation was abolished (Emax 6 +/- 5% of the U46619-induced contraction). mRNA encoding the human NPY Y1 receptor was detected in endothelial cells from human umbilical veins. It was concluded that NPY induces vasodilation in human subcutaneous arteries. The dilation is mediated via the NPY Y1 receptor and is dependent on nitric oxide. PMID- 10721818 TI - Neuropeptide Y stimulates DNA synthesis in human vascular smooth muscle cells through neuropeptide Y Y1 receptors. AB - We investigated the mitogenic effect, measured as [3H]thymidine incorporation, of neuropeptide Y (NPY) on smooth muscle cells (SMCs) from human subcutaneous arteries (diameter: 0.4 mm). NPY stimulated DNA synthesis in a concentration dependent manner, Emax 32 +/- 5% relative to control. The effect was potently antagonised by the NPY Y1 receptor antagonist BIBP3226 ((R)-N2-(diphenylacetyl)-N [(4-hydroxy-phenyl)methyl]-D-arginine-a mide), indicating the effect to be mediated via the NPY Y1 receptor. Noradrenaline (NA) also induced mitogenesis, Emax 35 +/- 10% relative to control. When added together, NPY and NA potentiated the [3H]thymidine incorporation, Emax 109 +/- 38% relative to control. Also, this effect seems to be mediated by the NPY Y1 receptor, since BIBP3226 blocked the effect (44 +/- 9% relative to control). The mitogenic effect of NPY and NA, two important transmitters of the sympathetic nervous system, might have clinical consequences on conditions with elevated sympathetic nerve activity. PMID- 10721819 TI - Effects of long-term pretreatment with isoproterenol on bromocriptine-induced tachycardia in conscious rats. AB - It has been shown that bromocriptine-induced tachycardia, which persisted after adrenalectomy, is (i) mediated by central dopamine D2 receptor activation and (ii) reduced by 5-day isoproterenol pretreatment, supporting therefore the hypothesis that this effect is dependent on sympathetic outflow to the heart. This study was conducted to examine whether prolonged pretreatment with isoproterenol could abolish bromocriptine-induced tachycardia in conscious rats. Isoproterenol pretreatment for 15 days caused cardiac hypertrophy without affecting baseline blood pressure and heart rate. In control rats, intravenous bromocriptine (150 microg/kg) induced significant hypotension and tachycardia. Bromocriptine-induced hypotension was unaffected by isoproterenol pretreatment, while tachycardia was reversed to significant bradycardia, an effect that was partly reduced by i.v. domperidone (0.5 mg/kg). Neither cardiac vagal nor sympathetic tone was altered by isoproterenol pretreatment. In isolated perfused heart preparations from isoproterenol-pretreated rats, the isoproterenol-induced maximal increase in left ventricular systolic pressure was significantly reduced, compared with saline-pretreated rats (the EC50 of the isoproterenol-induced increase in left ventricular systolic pressure was enhanced approximately 22 fold). These results show that 15-day isoproterenol pretreatment not only abolished but reversed bromocriptine-induced tachycardia to bradycardia, an effect that is mainly related to further cardiac beta-adrenoceptor desensitization rather than to impairment of autonomic regulation of the heart. They suggest that, in normal conscious rats, the central tachycardia of bromocriptine appears to predominate and to mask the bradycardia of this agonist at peripheral dopamine D2 receptors. PMID- 10721820 TI - Quantification: the soft underbelly of molecular biology. PMID- 10721821 TI - Variation in epidermal housekeeping gene expression in different pathological states. AB - Using non-radioactive in situ hybridization we studied the expression of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and PolyA+ RNA in psoriasis and normal skin, and GAPDH in epidermis following application of a range of noxious stimuli, including ultraviolet radiation, Sellotape stripping and various irritants. In keeping with what might have been expected from previous results on cell culture of various non-epidermal cell types, expression of these putative control gene products is not constant. We suggest that the use of GAPDH, and possibly other control genes, may lead to error in interpreting experiments using Northern blotting or even RT-PCR of epidermal samples. PMID- 10721822 TI - Decreased mRNA levels of retinoic acid receptor alpha, retinoid X receptor alpha and thyroid hormone receptor alpha in lesional psoriatic skin. AB - Retinoic acid, vitamin D3 and triiodothyronine regulate keratinocyte proliferation and differentiation--processes that are disturbed in psoriatic skin -via binding to nuclear receptors for retinoic acid (RAR-alpha,-gamma), vitamin D3 (VDR), thyroid hormone (TR-alpha,-beta) plus the common heterodimer partners, the 9-cis-retinoic acid receptors (RXR-alpha,-beta). By using a new quantitative real-time polymerase chain reaction assay, the expression of these receptors and three housekeeping genes (cyclophilin, GAPDH and beta-actin) was studied in psoriatic skin. The expression of housekeeping genes was consistently 2.7-4.3 times higher in lesional than in non-lesional skin. When the beta-actin expression was used to normalize the receptor mRNA values, the RARalpha, RXRalpha and TRalpha transcripts were found to be 58-75% lower in lesional vs. non lesional skin and the RXRalpha:RARgamma ratio was reduced from 3.2 to 1.5. Topical treatment for 4 days with 0.025% all-trans-retinoic acid or calcipotriol under occlusion did not normalize the altered mRNA expression of RARs, RXR and VDR in lesional skin. The results suggest that retinoid and thyroid hormone signalling is abnormal in lesional psoriatic skin, but how this relates to the pathogenesis of the disease is still unclear. PMID- 10721823 TI - In vivo gene transfer method in keratinocyte gene therapy: intradermal injection of DNA complexed with high mobility group-1 protein in rats. AB - In order to develop a more efficient method of introducing genes into keratinocytes in vivo, we intradermally injected DNA bound to high mobility group 1 protein, thereby taking advantages of the naked DNA and hemagglutinating virus of the Japan-liposome method reported previously. First we performed a gel mobility shift assay, which confirmed DNA binding to high mobility group 1. Then we injected beta-galactosidase expression vector complexed with high mobility group 1 into the rat skin and the activity of sample with the protein was 2-3 times higher than that without the protein as control. Semiquantification of transferred-DNA content using polymerase chain reaction and a time course of transgene expression in keratinocytes suggested that high mobility group 1 protein increased transfer of the DNA from the cytoplasm to the nucleus. Direct injection of the DNA-high-mobility-group-1 complex is a highly efficient method for introducing genes into keratinocytes in connection with gene therapy. PMID- 10721824 TI - Expression of PGP9.5 on Langerhans' cells and their precursors. AB - Langerhans' cells are epidermal dendritic cells, derived from blood precursors. Their main function is antigen presentation to T-cells. They are able to express neuronal proteins, such as neuron-specific enolase or substance P-receptor. They are closely associated with nerve fibres. PGP9.5 is the most specific neuronal protein in the epidermis. Epidermal Langerhans' cells can express PGP9.5 if denervated. Using flow cytometry, we found that cultured CD34+ precursors did not express PGP9.5, whereas suspensions of fresh or cultured Langerhans' cells could express this neuronal protein. Precursors of Langerhans' cells are not able to express PGP9.5, suggesting that they are not mature enough or that the capacity to express PGP9.5 may be acquired only in the epidermis. The function of PGP9.5 on Langerhans' cells and mature dendritic cells remains unknown. PGP9.5 might be related to dendritic cell maturation or to the lack of contacts with nerve endings. PMID- 10721825 TI - Superantigen Staphylococcal enterotoxin B induces release of IL-1beta in human epidermis. AB - Lesional skin in patients with inflammatory skin diseases is often colonized with Staphylococcus aureus, which is capable of releasing superantigens. We therefore studied whether application of superantigen on the skin led to release of cytokines, especially IL-1beta. Suction blisters were raised on vehicle- and superantigen-treated skin and IL-1beta protein levels measured in suction blister fluid and supernatant from blister roofs. In all volunteers studied, application of the superantigen Staphylococcal enterotoxin B led to increased release of IL 1beta protein from suction blister roofs (n=7). In contrast, we did not detect any difference in IL-1beta in the blister fluid (n=5). IL-1beta is known as a mediator of inflammation, and the increase in IL-1beta may be involved in the aggravation of inflammatory skin diseases seen following Staphylococcus aureus colonization. PMID- 10721826 TI - Blood perfusion studies on basal cell carcinomas in conjunction with photodynamic therapy and cryotherapy employing laser-Doppler perfusion imaging. AB - Superficial blood perfusion was monitored using laser-Doppler perfusion imaging in connection with a phase III clinical trial comparing photodynamic therapy, utilizing topically applied delta-aminolevulinic acid, with cryotherapy of basal cell carcinomas. A total of 526 images were recorded before and immediately after the treatment and during the follow-up period. Before treatment, the lesions exhibited a blood perfusion 3+/-2 times that in normal tissue. Both treatment modalities induced an increased blood perfusion inside the lesions, which slowly approached normal values in conjunction with successful treatments. The blood perfusion in successfully treated lesions approached normal values 2 months after photodynamic therapy, and about 1 year after cryotherapy. The tissue perfusion in recurrent lesions did not decrease to normal values after the treatment, suggesting that the laser-Doppler perfusion imaging technique can be used to follow the healing process and discover possible persistent tumour growth. PMID- 10721827 TI - Antipruritic action of thalidomide. AB - The effect of thalidomide on itch was studied in 11 patients with chronic pruritus from psoriasis, eczema, nodular prurigo, senile pruritus and primary biliary cirrhosis. Itch, assessed subjectively by the patients on a 10 cm line and measured objectively as nocturnal scratch movement, was decreased by thalidomide 200 mg on the 2 nights it was given. There was no improvement in the underlying disorders and it is concluded that thalidomide is a primary antipruritic agent. The patients all became drowsy and it seems likely that, as with most other antipruritic agents, the antipruritic action of thalidomide results from a central depressant effect. The primary antipruritic effect of thalidomide should now be assessed therapeutically. PMID- 10721828 TI - Topical photodynamic therapy for localized scleroderma. AB - Therapy of localized scleroderma is unsatisfactory, with numerous treatments being used that have only limited success or considerable side-effects. The aim of this trial was to determine whether topical photodynamic therapy would be effective in patients with localized scleroderma. Five patients with progressive disease, in whom conventional therapies had failed, were treated by application of a gel containing 3% 5-aminolevulinic acid followed by irradiation with an incoherent lamp (40 mW/cm2, 10 J/cm2). The treatment was performed once or twice weekly for 3-6 months. In all patients the therapy was highly effective for sclerotic plaques, as measured by a quantitative durometer score and a clinical skin score. The only side-effect was a transient hyperpigmentation of the treated lesions. These cases document the beneficial effect of topical photodynamic therapy in localized scleroderma. Controlled trials are now necessary to confirm these preliminary results. PMID- 10721829 TI - Excision of lipodermatosclerotic tissue: an effective treatment for non-healing venous ulcers. AB - In longstanding venous ulcers, the development of lipodermatosclerosis of the skin surrounding the ulcer is common. According to our clinical experience lipodermatosclerosis impairs the opportunities for the ulcer to heal. In this combined retrospective and prospective study the lipodermatosclerotic skin area was excised in 7 non-healing venous ulcers and then covered with split skin graft. All 7 legs had previously been treated with superficial venous surgery. Laser Doppler scanning of the ulcer area was performed pre- and postoperatively. Five of the 7 ulcers healed within 4 months and 1 healed within 9 months. Laser Doppler scanning showed increased blood flow in the lipodermatosclerotic skin area, which was decreased after the operation. This study indicates that excision of the lipodermatosclerotic skin area followed by split skin grafting can accomplish healing in non-healing venous leg ulcers that have failed to respond to previous superficial venous surgery. PMID- 10721830 TI - Azathioprine as a corticosteroid sparing agent for the treatment of dermatitis caused by the weed Parthenium. AB - Air-borne contact dermatitis caused by Parthenium hysterophorus is a serious problem in India. Patients with this condition have to use corticosteroids regularly in order to maintain clinical remissions, but prolonged usage causes serious side-effects. The weed cannot be eradicated. We have used 3 therapeutic regimens with azathioprine, which led to an effective control with minimal side effects even when used for long periods. A total of 22 patients (group I) were given 50 mg azathioprine twice a day; 11 patients (group II) received 50 mg azathioprine per day and 300 mg azathioprine every 28 days, and 10 patients (group III) were given 50 mg azathioprine twice a day along with 300 mg azathioprine every 28 days. The duration of treatment varied from 6 months to 3 years. Twenty patients in group I and 9 patients each in groups II and III had complete remission. Nine, 7 and 6 patients in the respective groups needed additional oral betamethasone 1-2 mg per day for brief periods only during the peak season in order to maintain complete remission. One patient in each group had only partial relief. The need for oral betamethasone during the second and the third year was further reduced. One patient each in group I and group II could not continue azathioprine due to the side-effects of the drug. PMID- 10721831 TI - Patch-testing with serial dilutions of tixocortol pivalate and potential cross reactive substances. AB - Of patch-tested patients with dermatitis, 4-5% are allergic to corticosteroids. Four groups of corticosteroids are recognized (A-D), where substances from the same group may cross-react. We investigated the potential cross-reactivity pattern and dose-response relationship for several corticosteroids from group A. We also included the corresponding aldehyde to hydrocortisone, as this degradation product has been proposed to be immunogenic. Eleven patients shown to be allergic to tixocortol pivalate were patch-tested with several corticosteroids from group A, as well as with the aldehyde, all in serial dilutions. All 11 reacted to both tixocortol pivalate and hydrocortisone. The dose-response relationship for the corticosteroids tended to be similar to sensitizers lacking anti-inflammatory potential. Patients with simultaneous reactions to many substances had high patch-test reactivity to tixocortol pivalate and hydrocortisone, while patients with few such reactions showed low reactivity (p=0.001 and 0.003, respectively). Several patients reacted to the aldehyde, supporting the theory that it is an intermediate in sensitization. PMID- 10721832 TI - Dorfman-Chanarin syndrome in a Turkish kindred: conductor diagnosis requires analysis of multiple eosinophils. AB - Dorfman-Chanarin syndrome is a rare, autosomal recessive inherited lipid storage disease with congenital ichthyotic erythroderma due to an acylglycerol recycling defect. Demonstration of lipid vacuoles in neutrophils from peripheral blood smears (Jordans' anomaly) in patients with ichthyotic erythroderma leads to the diagnosis. In spite of frequent liver, muscle, ear, eye and central nervous system involvement, Dorfman-Chanarin syndrome may present clinically as monosymptomatic ichthyosis. Here, we report clinical and laboratory investigations in a consanguineous family from Turkey with 3 affected family members, and demonstrate the lipid vacuoles in epidermal Langerhans' cells for the first time. Langerhans' cell phenotyping suggests that the skin inflammation is due to the gene defect and not to underlying atopic dermatitis. Microscopic examination of eosinophils for lipid vacuoles to identify conductors revealed variable percentages of normal and vacuolized eosinophils in conductors, suggesting the microscopic analysis of at least 10 eosinophils for conductor identification. PMID- 10721833 TI - Behavioural and social characteristics of subjects with repeated sexually transmitted diseases. AB - A case-control study was performed in order to assess risk factors for repeated sexually transmitted diseases. The study comprised 101 patients who had had sexually transmitted diseases 3 or more times during their lives and 182 controls who had no history of sexually transmitted disease. The subjects all attended the City Department for Skin and Venereal Diseases in Belgrade, Yugoslavia, from June 1997 to April 1998. According to multivariate logistic regression analysis, sexually transmitted diseases repeaters, in comparison with the controls, were older, more frequently divorced and widowed and without a regular partner, had more sexual partners and more sexual intercourse, and had more frequent sexual contact with people on the same day as meeting them. They also consumed alcohol, used sedatives and were prosecuted for criminal offences more frequently than the controls. The results of this study support the hypothesis that sexually transmitted diseases repeaters are different from their controls in terms of their behavioural and social characteristics. PMID- 10721834 TI - Frequent presence of Helicobacter pylori infection in chronic urticaria. PMID- 10721835 TI - Perianal ulcer as a leading symptom of paediatric Langerhans' cell histiocytosis. PMID- 10721836 TI - Cross-allergy to latex and spinach. PMID- 10721837 TI - Disseminated superficial porokeratosis induced by furosemide. PMID- 10721838 TI - Successful treatment of Hailey-Hailey disease with a scanned carbon dioxide laser. PMID- 10721839 TI - Bullous herpes zoster. PMID- 10721840 TI - Ehlers-Danlos syndrome type VIII with severe periodontitis and apical root resorption after orthodontic treatment. PMID- 10721841 TI - Clinical features of Ehlers-Danlos syndrome type VII in chromosome 6q deletion. PMID- 10721842 TI - Ultrasonic measurement of skin thickness in patients with systemic sclerosis. PMID- 10721843 TI - Elevated serum xylosyltransferase activity correlates with a high level of hyaluronate in patients with systemic sclerosis. PMID- 10721844 TI - Linear scleroderma associated with hypertrichosis in the absence of melorheostosis. PMID- 10721845 TI - Eosinophil activation in Wells' syndrome demonstrated immunohistochemically with antibodies against eosinophilic cationic protein. PMID- 10721846 TI - Pseudoaneurysm of the superficial temporal artery. PMID- 10721848 TI - Leukocytoclastic vasculitis in a patient with HHV-6 infection. PMID- 10721847 TI - Topical all-trans retinoic acid does not influence minimal erythema doses for UVB light in normal skin. PMID- 10721849 TI - Monoterpenoids and tetralin as pediculocides. PMID- 10721850 TI - Treatment of eosinophilic annular erythema with chloroquine. PMID- 10721851 TI - Itraconazole-induced drug eruption confirmed by challenge test. PMID- 10721852 TI - Interferon-gamma production in the peripheral lymphocytes of a patient with carbamazepine hypersensitivity syndrome. PMID- 10721853 TI - Primary cutaneous aspergillosis in an immunocompetent host. PMID- 10721854 TI - Asymptomatic bilateral optic perineuritis in secondary syphilis. PMID- 10721856 TI - Ashy dermatosis in an HIV antibody-positive patient. PMID- 10721855 TI - Multiple pilomatricomas, sternal cleft and mild coagulative defect. PMID- 10721857 TI - Accumulated p53 protein and UVA protection level of sunscreens. AB - Nuclear p53 expression is a sensitive parameter for the detection of ultraviolet (UV)-induced skin damage, and it has been used as an endpoint to evaluate the effectiveness of sunscreens. In this study, we compared the protection provided by two sunscreens having identical sun protection factors (SPF) but different UVA protection factors (UVA-PF) measured by the persistent pigment darkening method (PPD). The SPF of the sunscreens was 7 and the UVA-PF were respectively 7 and 3. Nuclear p53 protein was quantified in human skin biopsies treated with sunscreens and exposed 8 times to 5 MED of solar simulated radiation (SSR). The results showed that both sunscreens offered only partial protection against the increased expression of nuclear p53 protein induced by repetitive SSR exposures. However, a significantly lower level of p53-positive cells was found in areas protected with the sunscreen having the higher UVA-PF compared to the other sunscreen protected areas. In order to verify whether the difference in efficacy of these products was due to the difference in UVA absorption capacity, we quantified epidermal p53 protein accumulation after 8 exposures to either UVA (320-400 nm) or UVA1 (340 400 nm). We showed that as with SSR, repetitive exposures to 12.5 and 25 J/cm2 of UVA or UVA1 induced a significant increase in p53-positive cells in the human epidermis. These results confirmed that SPF determined on the basis of an acute erythemal reaction does not predict the level of protection against cumulative damage. They also showed that the protection provided by two sunscreens with different UVA protection factors is different (based on nuclear p53 protein accumulation), and that the PPD method can distinguish varying levels of sunscreen efficacy against UVA-induced cell damage. PMID- 10721858 TI - The kinetics of the tanning response to tanning bed exposures. AB - No clinical studies have been published documenting the development of pigmentation following the use of the Food and Drug Administration (FDA) recommended exposures from a tanning bed. Panelists were exposed three times weekly for eight weeks (24 exposures) using the FDA recommended exposure schedule. The initial tan was noted after only six exposures and quantitatively increased through the remainder of the study. PMID- 10721859 TI - What do young people think about the dangers of sunbathing, skin cancer and sunbeds? A questionnaire survey among Italians. AB - Previous experience in Australia and Sweden showed that public education programs produced substantial changes in people's opinions, attitudes and perceptions about melanoma, non-melanoma skin cancer, sunlight, sunbeds and suntanning. In order to organize effective prevention campaigns, more must be known about the sunbathing habits of children and adolescents. The aim of our study was to assess the knowledge that young people in a southern European country have about sun exposure. A total of 764 young people ranging from 16 to 22 years old (mean age: 19.3+/-1.2 years) responded to a questionnaire. Our study indicates that young people are very aware of the risks associated with sunbathing but that they continue to expose themselves without taking precautions. This suggests that: a) the majority of young Italians are reasonably well-informed but they do not take preventative measures; b) one effective measure could be promotion of the idea that an untanned body is more esthetically pleasing than a tanned one; c) a crucial point in the programming of future safety measures in suntan centers involves rigorous and regular controls. PMID- 10721860 TI - Effect of childhood and adolescent ultraviolet exposures on cumulative exposure in South East Queensland schools. AB - Quantitative estimates of the childhood and adolescent erythemal ultraviolet (UV) exposure received in South East Queensland schools are provided in this paper for age groups 0 to 6, 7 to 12 and 13 to 19 years. For the neck, hand and lower arm, sites of high UV exposure that are generally not covered by clothing, 13 to 19 year olds received the highest exposure of the three age groups, followed by 7 to 12 year olds. Exposure for 13 to 19 year olds contributed up to 44% of cumulative exposure to 20 years of age, and exposures for the 7 to 12 year olds contributed up to 31%. If the annual UV exposure for these two age groups were reduced to the average of all the age groups, cumulative erythemal UV exposure from 0 to 20 years would be reduced by up to 16%. On the other hand, if mothers can protect their babies by reducing the level of annual exposure to 30% of the annual UV exposure of the 7 to 12 year olds for the first four years then cumulative exposure to UV to age 20 would be reduced by up to 19%. These data confirm the importance of targeting young age groups in public campaigns for sun protection. PMID- 10721861 TI - Does smoking influence the efficacy of bath-PUVA therapy in chronic palmoplantar eczema? AB - Bath-PUVA therapy has been described as successful treatment for palmoplantar eczema. However, our own observations showed that patients with palmoplantar eczema of the dyshidrotic or hyperkeratotic type responded only partially to bath PUVA therapy. In order to evaluate environmental influences possibly having an impact on the efficacy of this therapy, smokers and non-smokers suffering from palmoplantar eczema treated with bath-PUVA therapy were compared. A retrospective study was conducted involving 62 patients, 39 non-smokers and 23 smokers, with palmar and/or plantar eczema resistant to local corticosteroids. Bath-PUVA therapy was performed according to the European standard regimen for oral PUVA therapy. The total number of treatments and the cumulative UVA-dose were similar in smokers and non-smokers (smokers 24+/-17.7 (mean+/-SD) and 67.6+/-51.3 J/cm2 vs. non-smokers 25.7+/-16.3 and 68.5+/-49.3 J/cm2). In the group of non-smokers, 31% showed complete remission (CR; 100% clearance), 33% partial remission (PR; more than 50% clearance) and 36% no change after treatment (NC; less than 50% clearance). In contrast, the group of smokers showed only 13% CR and 22% PR, whereas 65% exhibited NC. The differences regarding complete or partial remission between the groups were statistically significant (Student t-test for paired samples; P<0.05). Regarding the different type of eczema, bath-PUVA proved to be more successful in the dyshidrotic type of eczema as compared to the hyperkeratotic type in non-smokers (P<0.05). In the group of smokers no CR was achieved in patients suffering from the dyshidrotic form of eczema. Smoking is likely to be a reason for the failure of bath-PUVA therapy in the treatment of chronic palmoplantar eczema, in particular regarding smokers with eczema of the dyshidrotic type where no complete remission was achieved. PMID- 10721862 TI - Solar urticaria: report of two cases with augmentation spectrum. AB - Two adult Japanese patients with severe solar urticaria are reported. In both patients, an action spectrum existed in the visible light range, and an augmentation spectrum was demonstrated in the visible light range longer than the action spectrum. The augmentation phenomenon has rarely been documented, and in the previous reports, it was induced by preirradiation with the augmentation spectrum. In our cases, however, only postirradiation with the augmentation spectrum enhanced urticarial reactions. PMID- 10721863 TI - Low intensity laser irradiation in the treatment of recalcitrant radiation ulcers in patients with breast cancer--long-term results of 3 cases. AB - Radiotherapy can be followed by recalcitrant skin ulcers. As low intensity laser irradiation has been demonstrated to have a beneficial effect on impaired wound healing, we investigated its efficacy and safety in three patients with chronic radiation ulcers. The three patients, previously mastectomized due to breast cancer, with recalcitrant radiation ulcers of the skin were treated with a 30 mW helium-neon laser (wavelength: 632.8 nm, intensity: 3 mW/cm2, dose: 30 J/ cm2) three times weekly. In all patients, complete wound closure was achieved within a period of 7, 5, and 8 weeks. One patient died 6 weeks after laser treatment due to tumor cachexia. Neither of the other patients showed recurrence of radiation ulcers or neoplasm during a follow-up of 36 months. Low intensity helium-neon laser irradiation has been shown to be effective in the induction of wound healing in radiotherapy-induced ulcers in three patients with breast cancer. PMID- 10721864 TI - A case of actinic prurigo showing hypersensitivity of skin fibroblasts to ultraviolet A (UVA). AB - We here report a patient with actinic prurigo. He had had erythematous papulovesicular eruptions on the sun-exposed sites from fall to early summer for 4 years. The lesions healed leaving atrophic scars. The histology showed epidermal necrosis and dermal dense perivascular lymphohistiocytic infiltration and edema. His minimal erythema doses to ultraviolet B (UVB) and UVA were normal and lowered, respectively. Skin lesions were produced by repeated irradiation with UVA plus UVB, but not with UVA alone. Then he was diagnosed as having actinic prurigo. Skin fibroblasts from the patient were hypersensitive to UVA. We believe that the hypersensitivity relates to the pathomechanisms of the photosensitivity in the case. UVA sensitivity of fibroblasts may be useful for differentiating actinic prurigo, hydroa vacciniforme, and other similar photosensitive disorders. PMID- 10721865 TI - Placebo revisited. PMID- 10721866 TI - An overview of epidemiological studies on seasonal affective disorder. AB - OBJECTIVE: To review and systematize all epidemiological studies of seasonal affective disorder (SAD). METHOD: The relevant papers were identified by searches in Medline, Excerpta Medica, PsychLIT and other databases. The primary reports were reviewed for additional citations. The studies were classified into retrospective and prospective population surveys, surveys of patient populations and studies of seasonal variations in psychiatric illnesses other than mood disorders. RESULTS: The prevalence estimates of SAD across 20 retrospective studies varied from 0% to 9.7%. All prospective population studies, except one, find seasonal variations in mood, depressive symptoms usually peaking in winter. SAD was more prevalent at higher northern latitudes, but the prevalence varied across ethnic groups. SAD has also been identified in children and adolescents. Seasonal exacerbations and remissions are not limited to mood disorders, it has also been found in bulimia nervosa, anxiety disorders and other psychiatric illnesses. CONCLUSION: The actuality of seasonal variation in mood has been documented thoroughly by both retrospective and prospective studies. In the general population, depressive symptoms peak in winter, and the most extreme form of this disposition, SAD, appears to be a relatively common disorder. PMID- 10721867 TI - A population study on the influence of depression on neuropsychological functioning in 85-year-olds. AB - OBJECTIVE: To examine cognitive function in very old depressed individuals. METHOD: Individuals with major depression (MDS) or dysthymia according to the DSM III-R were compared to mentally healthy regarding tests of verbal ability, inductive logical reasoning, spatial ability, perceptual speed, basic arithmetics, primary memory and secondary memory in a population-based sample of 85-year-olds. RESULTS: Individuals with MDS performed worse than mentally healthy individuals in tests of verbal ability, inductive logical reasoning, spatial ability, perceptual speed and secondary memory. There were no differences between the groups regarding basic arithmetics and primary memory. The poor test performance was mainly associated with psychomotor retardation and decreased concentration in depressed individuals. Memory complaints were not correlated to poor test performance, neither in the mentally healthy nor in the depressed. CONCLUSION: MDS in elderly individuals is associated with reduced cognitive test performance, especially regarding more complex and time-demanding tests and in tests of secondary memory. PMID- 10721868 TI - Somatoform syndromes and disorders in a representative population sample of adolescents and young adults: prevalence, comorbidity and impairments. AB - OBJECTIVE: To present prevalence findings of DSM-IV somatoform symptoms, syndromes and disorders in a representative sample of adolescents and young adults. METHOD: Data come from the Early Developmental Stages of Psychopathology (EDSP) study, in which a total of 3021 respondents aged 14-24 years were assessed by the Munich-Composite International Diagnostic Interview. RESULTS: Although specific DSM-IV somatoform disorders were relatively rare with a lifetime rate of 2.7%, a considerably higher proportion of respondents met criteria for clinically significant somatoform syndromes as defined by the Somatic Symptom Index SSI4, 6 (lifetime: 1.7%), as well as the undifferentiated somatoform/dissociative syndrome USDS (lifetime: 9.1%), resulting in an overall prevalence rate of 12.6%. Somatoform conditions are often comorbid with other mental disorders and were found to be associated with remarkable impairments and disabilities. CONCLUSION: Somatoform disorders and syndromes in young adults are frequent, impairing and often associated with the development of other mental disorders. PMID- 10721869 TI - Maternity blues and attachment to children in mothers of full-term normal infants. AB - OBJECTIVE: This study was conducted for evaluating incidence of maternity blues in Japan, in addition to clarifying the relationship between maternity blues and maternal attachment, and the factors involved. METHOD: A questionnaire survey was conducted on 417 mothers having given birth at the Nagoya Daini Red Cross Hospital. The questionnaire consisted of Zung's self-rating depression scale, and a 'postpartum maternal attachment' scale, consisting of subscales on 'core maternal attachment' and 'anxiety regarding children'. The survey was conducted 5.2 days +/-1.46 postpartum. RESULTS: ZSDS scores over 40 amounted to 66.8% of the responses. Analysis of the two scales revealed significant correlation/inverse correlation between 'maternity blues' and 'anxiety regarding children'/ 'core maternal attachment'. Path analysis revealed 'maternity blues' to be influencing 'core maternal attachment' and 'anxiety regarding children'. CONCLUSION: It was found that the incidence of maternity blues may be higher in Japan than was believed previously, and that an intimate association exists between 'maternity blues' and 'postpartum maternal attachment'. PMID- 10721870 TI - A placebo-controlled comparison of zotepine versus chlorpromazine in patients with acute exacerbation of schizophrenia. AB - OBJECTIVE: The aim of this study was to evaluate the efficacy of zotepine in the treatment of acute episodes of schizophrenia. METHOD: Patients with acute exacerbation of schizophrenia (DSM-III-R criteria; n = 158) were allocated on a random, double-blind basis to receive zotepine (150 or 300 mg/day), chlorpromazine (300 or 600 mg/day) or placebo for 8 weeks. Symptoms were assessed on the BPRS, SANS and CGI scales at baseline and weeks 1, 2, 4, 6 and 8 and patients were assessed at these times for adverse effects. Analysis was by analysis of variance on the intent-to-treat population, with last observation carried forward. RESULTS: Mean BPRS scores improved statistically significantly more with zotepine than chlorpromazine (point estimate of difference -12.4, 95% CI -18.3 to -6.5) or placebo (point estimate of difference -12.7, 95% CI -18.6 to -6.8). Zotepine produced significantly fewer extrapyramidal symptoms (EPS) than chlorpromazine. CONCLUSION: Zotepine is an effective antipsychotic with low propensity for EPS. PMID- 10721871 TI - Decreased platelet monoamine oxidase activity in female anorexia nervosa. AB - OBJECTIVE: To study if platelet MAO activity, previously described as a serotonergic index, is modified in a sample of pure restrictive anorectic patients. METHOD: Twenty-five female patients with DSM-IV anorexia nervosa restricting type were studied and compared with 30 healthy female controls. Platelet MAO activity was measured by isotopic methods using C-14 benzylamine. Impulsive personality features were measured with specific rating scales and temperament studied with Cloninger's TCI. RESULTS: Platelet MAO activity was significantly lower (4.3+2.7 nmol/h/ 108 platelets) in the anorectic patients than in the control group (6.7+2.8) (P<0.01). Platelet MAO was inversely correlated with scores on impulsivity scales and positively correlated with the dimension 'persistence' of Cloninger's TCI. CONCLUSION: Platelet MAO activity is lowered in a group of patients with anorexia nervosa and might involve some dysfunction in the regulation of impulse control. PMID- 10721872 TI - Low serum cholesterol levels and depressive state in human dock visitors. AB - OBJECTIVE: The purpose of the present study was to investigate an association between serum cholesterol levels and depressive state. METHOD: Physical examinations and mental assessments were performed for 13702 human dock visitors. We obtained 13571 sets of complete data for serum total cholesterol, the scores for depressive state and possible confounders such as sex, age, body weight, body mass index (BMI), recent weight loss, total protein and concomitant medical diagnoses. RESULTS: Their depressive state varied significantly across the serum cholesterol levels after adjusting age and gender. After controlling all the above confounding variables the significance still remained, not only in the categorical analysis but also in the continuous analysis. CONCLUSION: The present findings suggest that there is an association between cholesterol and depressive state. PMID- 10721873 TI - Psychometric measures of individual change: an empirical comparison with the Brief Psychiatric Rating Scale (BPRS). AB - OBJECTIVE: An empirical comparison of treatment efficacy estimates as based on psychometric measures of intra-individual change (Reliable Change methods). METHOD: All seven different methods of assessing Reliable Change that have been advocated in the past two decades are compared empirically in a large in-patient sample (n = 107). Estimates of treatment efficacy by each of these seven Reliable Change methods are computed, using pre-/post-score changes on the Brief Psychiatric Rating Scale (BPRS). RESULTS: It is demonstrated that Reliable Change methods may yield very different estimates of treatment efficacy. The Reliable Change method with the fewest statistical assumptions is one of the least sensitive Reliable Change methods. CONCLUSION: Disagreement on the proper definition of Reliable Change is not merely of theoretical importance, but also has major practical implications. PMID- 10721874 TI - Smoking and the risk of suicide. AB - OBJECTIVE: To estimate the relationship between cigarette smoking and the risk of suicide. METHOD: The mortality of 36527 adult men and women was monitored for the mean 14.4 years. Information on deaths caused by suicide was obtained from the National Mortality Register. Suicides were subclassified by the level of violence used. Current smokers of 1-20 cigarettes per day were considered light/moderate smokers and heavy smokers were defined as those smoking > or =21 cigarettes per day. RESULTS: There were 134 suicides among 17798 men and 31 suicides among 18729 women. The most common suicide methods were hanging, firearms and drug overdose. According to the Cox model the adjusted relative risk of both violent and non violent suicide was significantly and linearly increased among light/moderate and heavy smokers compared with non-smokers. CONCLUSION: Smoking was associated with an increased risk of suicide irrespective of the level of violence used. PMID- 10721875 TI - Transcranial magnetic stimulation induces 'pseudoabsence seizure'. AB - OBJECTIVE: Several studies support the hypothesis of an antidepressive or mood enhancing effect of repetitive transcranial magnetic stimulation (rTMS) on depressive patients. The most acute concern regarding rTMS is possible seizure induction; therefore, reports on seizure during rTMS are of special significance. METHOD: We describe a case in which high frequency rTMS over the left dorsolatero prefrontal cortex (DLPC) applied as an add-on antidepressive strategy may have induced a frontal lobe complex partial seizure in a female patient affected by drug-resistant depression. RESULTS: The epileptic seizure was self-limited, and the patient did not report any physical sequelae. The psychopathological improvement, observed immediately after the incident in question, did not last. CONCLUSION: In this case train duration in rTMS, combined with drugs modulating the norepinephrine turnover, may have contributed to the occurrence of this complex partial seizure, which neuroanatomically seems to be localized in the DLPC. PMID- 10721876 TI - Nocturnal sweating and temperature in depression'. PMID- 10721877 TI - Delineating the longitudinal structure of depressive illness: beyond clinical subtypes and duration thresholds. AB - Through the use of polysomnographic, epidemiologic, and prospective clinical follow-up studies, the authors document that the course of major depressive disorder (MDD) is expressed by fluctuating symptoms in which depressive subtypes included in official diagnostic systems do not represent discrete disorders, but are stages along a dimensional continuum of symptomatic severity. Depressive symptoms at the major, minor, dysthymic or otherwise sub-threshold levels are all integral components of the longitudinal clinical structure of MDD with each symptom level representing a different phase of illness intensity, activity and severity. Detailed analyses indicate that patients are symptomatic 60 % of the time, much of it at the minor, dysthymic or subthreshold level. The symptomatic phases of illness activity are interspersed sporadically with inactive phases, when patients are asymptomatic. These findings are pertinent to both clinical cohorts and community-based epidemiologic samples. Each level of depressive symptom severity is associated with significant psychosocial impairment; such impairment increases progressively with each stepwise increment in symptom severity. When patients are asymptomatic their psychosocial functioning returns to good or very good levels. Residual subthreshold symptoms in the course of MDD are associated with high risk for early episode relapse and a significantly more chronic course of illness. Asymptomatic recovery from MDD is associated with significant delays in episode relapse and recurrence and a more benign course of illness. We submit that, as in the case of chronic medical conditions, the goal of treating unipolar depressive illness should optimally be to return the patient to as asymptomatic a level as is feasible by all available therapeutic means. PMID- 10721878 TI - Mortality in 497 patients with affective disorders attending a lithium clinic or after having left it. AB - The impact of lithium prophylaxis on mortality has been studied in 497 patients, 405 bipolars and 92 unipolars, who attended the same out-patient lithium clinic for up to 30 years. In order to avoid preselection, no minimum period of lithium treatment was required in our study. Of a total of 6014 patient-years, 4330 were spent in regular contact with the study clinic. General mortality due to natural causes was not significantly increased; among cardiovascular diseases, only pulmonary embolism showed an excess mortality. No patients died of lithium intoxication or chronic renal insufficiency. Patients were divided into three groups: Group A, 277 patients, attended the study clinic until death or the end of the study, Group B, 86 patients, left the clinic but continued to take lithium, and Group C, 134 patients, both left the clinic and stopped taking lithium. Among bipolars, the suicide rate compared to the general population was in excess in all three groups. Among unipolars, suicides occurred only after the patients had left the study clinic and stopped taking lithium. A special analytical method was used for intergroup comparisons of suicide rates. Bipolars in Group A attending the study clinic regularly had a suicide rate of 3.5 per 1000 patient-years. The rate increased to 6.3 or by 80 % if patients had left the clinic and did not take lithium any longer as in Group C. The suicide rate in Group C increased by 45% compared to Group B, patients who left the clinic but continued to take lithium. Our results support the hypothesis that lithium has a significant antisuicidal effect in bipolars as well as in unipolars. The suicide mortality can be further reduced by regular attendance in a specialised mood disorder clinic. PMID- 10721879 TI - Effect of acute administration of hypericum perforatum-CO2 extract on dopamine and serotonin release in the rat central nervous system. AB - The hydromethanolic extract of Hypericum perforatum has been shown to be an effective antidepressant, although its mechanism of action is still unclear. In this study, in vivo microdialysis was used to investigate the effects of Hypericum perforatum-CO2 extract on dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC), serotonin (5-HT), and 5-hydroxyindoleacetic acid (5-HIAA) release in various areas of brain. Administration of Hypericum perforatum extract (1 mg/kg, p.o.) caused a slight, but significant increase of DA outflow both in the nucleus accumbens and the striatum. The maximal increase of DA efflux (+19.22+/-1.93%, relative to the control group) in the nucleus accumbens occurred 100 min after administration of Hypericum perforatum. In the striatum, the extract maximally enhanced DA outflow (+24.83+/-7.49 %, relative to the control group) 80 min after administration. Extraneuronal DOPAC levels were not significantly affected by Hypericum perforatum treatment. Moreover, Hypericum perforatum (1 mg/kg, p.o.) did not produce any significant effect on either 5-HT or 5-HIAA efflux in the ventral hippocampus. This study shows for the first time that Hypericum perforatum extract is capable of increasing in vivo DA release. PMID- 10721880 TI - The benzamide tiapride: treatment of extrapyramidal motor and other clinical syndromes. AB - The benzamide derivative tiapride (Tiapridex, Synthelabo) has a highly selective antagonistic effect on striatal adenylate cyclase-independent dopamine-2 receptors. Its in vitro binding affinity is especially high for dopamine receptors which have been sensitized by pre-incubation with dopamine. The involvement of altered dopamine receptor sensitivity in several extrapyramidal dys- and hyperkinesia has been hypothesized. By its high affinity for these receptors, without any affinity for other neurotransmitter receptors of the brain, tiapride is especially well suited for the treatment of movement disorders related to functional dopamine hyperactivity. Even at higher doses, tiapride does not exceed a D2-receptor occupancy of 80%, which is in accordance with the finding that tiapride rarely causes acute extrapyramidal syndromes and has, up to now, never implicated in inducing tardive dyskinesias. On the contrary, clinical studies demonstrate its excellent efficacy in neuroleptic-induced tardive dyskinesia, L-Dopa-induced dyskinesias, psychomotor agitation in geriatric patients and choreatic movement disorders. Since tiapride is not available in the USA as yet, most of the studies concerning tiapride have been carried out in Europe. In a recent study, based on objective measurements, tiapride effectively controlled choreatic movements in patients suffering from Huntington's disease (HD). Tiapride is well tolerated in daily doses between 300 and 1200 mg. Adverse events are generally rare and mild. PMID- 10721881 TI - Amantadine influences cognitive processing in patients with multiple sclerosis. AB - We investigated the effect of amantadine on cognitive processing in patients with multiple sclerosis (MS) and fatigue with objective electrophysiological measures. Behavioral methods (Reaction Time, RT) and two different Event Related Potential (ERP) components measuring i) stimulus selection (Selection Negativity, SN) and ii) response selection (Lateralized Readiness Potential, LRP) were employed. Twenty-four patients with clinical definite MS (10 relapsing remitting and 14 secondary progressive) and confirmed fatigue in the past three months (Fatigue Severity Scale (FSS) > 4) were included. Patients were randomized in a double blind, placebo-controlled cross-over design. We found a difference between the two treatments for ERP measures to stimuli with relevant colour starting at about 200 ms. This negativity had a higher amplitude during amantadine treatment regardless of treatment order. The RT did not differ significantly between the treated and untreated groups. Additional analysis indicated that patients with a disease duration of less than 7 years had a significant test position (practice effect), but no treatment effect, while patients with a longer MS duration showed no practice effect, but rather an improved reaction speed and increased ERP amplitude effects when treated with amantadine. The present findings suggest that amantadine exerts beneficial effects on early cognitive processes in patients with MS, but appears to be limited to subjects with a longer duration of the disease. PMID- 10721883 TI - Clozapine withdrawal symptoms after change to sertindole in a schizophrenic patient. AB - A 30-year-old male patient with paranoid schizophrenia was on clozapine therapy for more than five years. Discontinuation of clozapine and an attempt to change his medication to sertindole has led to serious psychotic and somatic symptoms. After readministration of clozapine the psychotic symptoms rapidly disappeared. The patient was monitored by BPRS and PANSS positive and negative scale. Also clinical and labor parameters of the patient were monitored. The change of his medication from clozapine to sertindole was unsuccessful. This case report suggests that although atypical antipsychotics may be generally different from the classical neuroleptic drugs, there are also significant differences among the atypical antipsychotic drugs in their effects on the receptors of the central nervous system. Therefore the change of clozapine to another atypical antipsychotic medication in the clinical practice should be cross-tapered and the symptoms of withdrawal closely monitored. PMID- 10721882 TI - Posthallucinogen-like visual illusions (palinopsia) with risperidone in a patient without previous hallucinogen exposure: possible relation to serotonin 5HT2a receptor blockade. AB - BACKGROUND: Previous reports document visual illusions resembling hallucinogen persisting perception disorder (HPPD) after risperidone treatment in patients with histories of previous LSD exposure. METHODS: We report a case with visual disturbances resembling HPPD after each of three consecutive risperidone dose increases. RESULTS: Contrasting with previous reports, our patient lacked any history of substance abuse, particularly hallucinogen exposure. She lacked neurologic or other contributory illnesses. Illusions generally remitted within 48 hours each time. Coadministration of trazodone and clonazepam may have contributed to these phenomena, although clonazepam has been used to treat this condition. She had been unusually sensitive to the side-effects of many psychotropics. CONCLUSIONS: This case is unique due to the absence of substance abuse. This and another report note heightened sensitivity to medication side effects. Visual phenomena resembling HPPD evidently can occur with risperidone and, possibly, other atypical antipsychotics and certain antidepressants regardless of previous hallucinogen use. Several lines of evidence implicate reduced 5HT2a serotonin receptor stimulation rather than increased 5HT2c stimulation. PMID- 10721884 TI - Dietary composition and obesity: do we need to look beyond dietary fat? PMID- 10721885 TI - The role of energy density in the overconsumption of fat. AB - In recent years, research has focused on why fat is so readily overconsumed. Although the palatability of many high fat foods can encourage overconsumption, another possibility is that fat is not very satiating. A number of studies have compared the effects of fat and carbohydrate on both satiation (the amount eaten in a meal) and satiety (the effect on subsequent intake), but have found little difference between these macronutrients when the palatability and energy density were similar. On the other hand, the energy density of foods has been demonstrated to have a robust and significant effect on both satiety and satiation, independently of palatability and macronutrient content. It is likely that the high energy density of many high fat foods facilitates the overconsumption of fat. An understanding of the role that the energy density of foods plays in the regulation of food intake should lead to better dietary management of hunger and satiety in conditions associated with both over- and underconsumption of energy, such as obesity and anorexia. PMID- 10721886 TI - Dietary fiber and energy regulation. AB - Dietary fiber has many functions in diet, one of which may be to aid in energy intake control and reduced risk for development of obesity. The role of dietary fiber in energy intake regulation and obesity development is related to its unique physical and chemical properties that aid in early signals of satiation and enhanced or prolonged signals of satiety. Early signals of satiation may be induced through cephalic- and gastric-phase responses related to the bulking effects of dietary fiber on energy density and palatability, whereas the viscosity-producing effects of certain fibers may enhance satiety through intestinal-phase events related to modified gastrointestinal function and subsequent delay in fat absorption. The goal of this paper is to provide a brief overview of the role of dietary fiber in energy intake regulation, highlighting the relationship between fiber properties and physiologic action. PMID- 10721887 TI - Dietary determinants of energy intake and weight regulation in healthy adults. AB - Until recently, the percentage of energy from dietary fat has been considered a primary determinant of body fatness. This review covers recent studies from our laboratory that challenge this notion. High and low fat diets matched for energy density, palatability and fiber resulted in similar mean voluntary energy intakes over 9 d; analysis of the individual foods in these diets showed that energy density and palatability were significant determinants of energy intake, independent of fat content. Path analysis further revealed that the influence of energy density on energy intake was in part direct, and in part indirect and mediated by palatability. In another study, dietary variety within food groups was shown to be an important predictor of body fatness, and the direction of the association depended on which food groups provided the variety, i.e., the variety of sweets, snacks, condiments, entrees and carbohydrates consumed was positively associated with body fatness, whereas the variety of vegetables was negatively associated. Last, a study of restaurant food and body fatness showed that the frequency of consumption of restaurant food was positively associated with body fatness, independent of education level, smoking status, alcohol intake and physical activity. Restaurant meals tend to be high in fat and low in fiber, and thus energy dense. Restaurants also typically serve a variety of palatable foods in large portions. The increasing variety of high energy foods available and the increasing proportion of household income spent on foods consumed away from home may help explain the U.S. national rising prevalence of obesity. PMID- 10721888 TI - Dietary glycemic index and obesity. AB - Obesity is among the most important medical problems in America today. Currently, approximately 1 in 4 children and 1 in 2 adults are overweight, prevalence rates that have increased by 50% since the 1960s. In an attempt to combat this problem, the Federal government and various official medical agencies have advocated decreasing intake of total fat and sugar, while increasing consumption of "complex carbohydrate." Despite a recent reduction in fat consumption to near the recommended 30% of total energy, rates of obesity have continued to rise, suggesting that other dietary factors may play a critical role in body weight regulation. One such factor may be glycemic index. This review examines the physiologic effects of glycemic index and argues for the need for controlled clinical trials of a low glycemic index diet in the treatment of obesity. PMID- 10721889 TI - Dietary fat intake and regulation of energy balance: implications for obesity. AB - Obesity represents a major threat to health and quality of life. Although obesity has strong genetic determinants, the increasing prevalence of obesity in populations around the world suggests that environmental factors are promoting or exacerbating the problem. Experts are calling for public health efforts to deal with the global epidemic of obesity. Such a campaign would require that we identify and modify environmental factors that promote obesity. Our current food supply is high in fat, and high fat diets have been suggested to promote obesity by increasing energy intake, thus increasing the probability of positive energy balance and weight gain. However, others argue that high fat diets are not promoting obesity. In this paper, we review evidence from animal studies, carefully controlled laboratory studies, cross-sectional studies, clinical trials and studies in individuals at high risk to develop obesity. Although there are many environmental factors promoting excess energy intake and discouraging energy expenditure, it is clear that consumption of a high fat diet increases the likelihood of obesity and that the risk of obesity is low in individuals consuming low fat diets. On the basis of the available data, the current public health recommendations to lower dietary fat intake appear to be appropriate. PMID- 10721890 TI - Long-chain acyl-CoA as a multi-effector ligand in cellular metabolism. AB - Fatty acyl-CoA esters have the ability to bind at specific sites on certain proteins through their CoA moiety, thereby acting as modulators of cellular metabolism. In some cases at least, the acyl-CoA competes with cofactors (nucleotides) for binding to the proteins and results in either their activation or inhibition of catalytic activity. Photolabeling derivatives of acyl-CoA permit covalent binding of the esters to the proteins, which should lead to determination of amino acid residues required for ligand binding, if a common binding motif exists. On the basis of the accumulation of published results, there is now evidence to implicate acyl-CoA esters in the regulation of a variety of biological processes, ranging from mitochondrial metabolism to gene transcription to insulin secretion and signaling. PMID- 10721891 TI - Role of acyl-CoA binding protein in acyl-CoA metabolism and acyl-CoA-mediated cell signaling. AB - Long-chain acyl-CoA esters (LCA) act both as substrates and intermediates in metabolism and as regulators of various intracellular functions. Acyl-CoA binding protein (ACBP) binds LCA with high affinity and is believed to play an important role in intracellular acyl-CoA transport and pool formation and therefore also for the function of LCA as metabolites and regulators of cellular functions . The free concentration of cytosolic LCA is efficiently buffered to low nanomole concentration by ACBP and fatty acid binding protein (FABP). An additional important factor is the activity of acyl-CoA hydrolases. The estimated cellular free LCA concentration is two to four orders of magnitude lower than the concentrations reported to be necessary to regulate most LCA-affected cellular functions. Preliminary evidence indicates that the regulatory effect of LCA might be mediated by the LCA/ACBP complex. PMID- 10721892 TI - The role of long-chain fatty acyl-CoA esters in beta-cell signal transduction. AB - Glucose-induced insulin secretion is associated with inhibition of free fatty acid (FFA) oxidation, increased esterification and complex lipid formation by pancreatic beta-cells. Abundant evidence favors a role for cytosolic long-chain acyl-CoA (LC-CoA), including the rapid rise in malonyl CoA, the inhibitory effect of hydroxycitrate or acetyl CoA carboxylase knockout, both of which prevent malonyl CoA formation, and the stimulatory effect of exogenous FFA. On the other hand, some evidence opposes the concept, including the fall in total LC-CoA levels in response to glucose, the stimulatory effect of LC-CoA on K(ATP) channels and the lack of inhibition of glucose-stimulated secretion either by overexpression of malonyl CoA decarboxylase, which markedly lowers malonyl CoA levels, or by triacsin C, which blocks FFA conversion to LC-CoA. Alternative explanations for these data are presented. A revised model of nutrient-stimulated secretion involving two arms of signal transduction that occur simultaneously is proposed. One arm depends on modulation of the K(ATP) channel evoked by changes in the ATP/ADP ratio. The other arm depends upon anaplerotic input into the tricarboxylic acid cycle, generation of excess citrate, and increases in cytosolic malonyl-CoA. Input from this arm is increased LC-CoA. Signaling through both arms would be required for normal secretion. LC-CoA esters and products formed from them are potent regulators of enzymes and channels. It is hypothesized that their elevations directly modulate the activity of enzymes, genes and various beta-cell functions or modify the acylation state of key proteins involved in regulation of ion channels and exocytosis. PMID- 10721893 TI - Long-chain acyl-CoA-dependent regulation of gene expression in bacteria, yeast and mammals. AB - Fatty acyl-CoA thioesters are essential intermediates in lipid metabolism. For many years there have been numerous conflicting reports concerning the possibility that these compounds also serve regulatory functions. In this review, we examine the evidence that long-chain acyl-CoA is a regulatory signal that modulates gene expression. In the bacteria Escherichia coli, long-chain fatty acyl-CoA bind directly to the transcription factor FadR. Acyl-CoA binding renders the protein incapable of binding DNA, thus preventing transcription activation and repression of many genes and operons. In the yeast Saccharomyces cerevisiae, genes encoding peroxisomal proteins are activated in response to exogenously supplied fatty acids. In contrast, growth of yeast cells in media containing exogenous fatty acids results in repression of a number of genes, including that encoding the delta9-fatty acid desaturase (OLE1). Both repression and activation are dependent upon the function of either of the acyl-CoA synthetases Faa1p or Faa4p. In mammals, purified hepatocyte nuclear transcription factor 4alpha (HNF 4alpha) like E. coli FadR, binds long chain acyl-CoA directly. Coexpression of HNF-4alpha and acyl-CoA synthetase increases the activation of transcription of a fatty acid-responsive promoter, whereas coexpression with thioesterase decreases the fatty acid-mediated response. Conflicting data exist in support of the notion that fatty acyl-CoA are natural ligands for peroxisomal proliferator-activated receptor alpha (PPARalpha). PMID- 10721894 TI - Functional characterization of mammalian mitochondrial carnitine palmitoyltransferases I and II expressed in the yeast Pichia pastoris. AB - Mitochondrial carnitine palmitoyltransferases I and II (CPTI and CPTII), together with the carnitine carrier, transport long-chain fatty acyl-CoA from the cytosol to the mitochondrial matrix for beta-oxidation. Recent progress in the expression of CPTI and CPTII cDNA clones in Pichia pastoris, a yeast with no endogenous CPT activity, has greatly facilitated the characterization of these important enzymes in fatty acid oxidation. It is now well established that yeast-expressed CPTI is a catalytically active, malonyl CoA-sensitive, distinct enzyme that is reversibly inactivated by detergents. CPTII is a catalytically active, malonyl CoA insensitive, distinct enzyme that is detergent stable. Reconstitution studies with yeast-expressed CPTI have established for the first time that detergent inactivation of CPTI is reversible, suggesting that CPTI is active only in a membrane environment. By constructing a series of deletion mutants of the N terminus of liver CPTI, we have mapped the residues essential for malonyl CoA inhibition and binding to the conserved first six N-terminal amino acid residues. Mutation of glutamic acid 3 to alanine abolished malonyl CoA inhibition and high affinity malonyl CoA binding, but not catalytic activity, whereas mutation of histidine 5 to alanine caused partial loss in malonyl CoA inhibition. Our mutagenesis studies demonstrate that glutamic acid 3 and histidine 5 are necessary for malonyl CoA inhibition and binding to liver CPTI, but not catalytic activity. PMID- 10721895 TI - Regulation of the fatty acid synthase promoter by insulin. AB - Expression of critical enzymes in fatty acid and fat biosynthesis is tightly controlled by nutritional and hormonal stimuli. The expression of fatty acid synthase, which catalyzes all reactions for synthesis of palmitate from acetyl CoA and malonyl-CoA, and of mitochondrial glycerol-3-phosphate acyltransferase, which catalyzes the first acylation step in glycerophospholipid synthesis, is decreased to an undetectable level during fasting. Food intake, especially a high carbohydrate, fat-free diet after fasting, causes a dramatic increase in the transcription of these genes. Insulin secretion is increased during feeding and has a positive effect on expression. By using adipocytes in culture and transgenic mice that express the reporter gene driven by the fatty acid synthase promoter, the cis-acting sequence that mediates insulin regulation of the fatty acid synthase promoter was defined. Upstream stimulatory factors (USF) that bind to the -65 E-box are required for insulin-mediated transcriptional activation of the fatty acid symthase gene. Sterol regulatory element binding protein (SREBP)-1 may be also involved in induction of these genes during feeding. Using specific inhibitors and expressing various signaling molecules, we found that insulin regulation of the fatty acid synthase promoter is mediated by the phosphatidylinositol (PI)3-kinase signaling pathway and that protein kinase B/akt is a downstream effector. PMID- 10721896 TI - Mechanistic aspects of vitamin and coenzyme utilization and function: a symposium in recognition of the distinguished career of Donald B. McCormick. PMID- 10721897 TI - A trail of research on cofactors: an odyssey with friends. AB - Over the span of 40 y and with the participation of over 60 students and postdoctoral colleagues, my laboratory has been able to elucidate numerous aspects of cofactor metabolism and function. Findings have been on the absorption, transport, utilization and excretion of vitamin B-6, riboflavin, biotin, lipoate and ascorbate. Specificity studies on those trace but essential enzymes that catalyze conversion of such vitamins as B-6 and riboflavin to their functional coenzymes led to our development of "biochemically specific absorbents" that prototypically exemplified what later was called "affinity chromatography." Characterization of the purified kinases for B-6 and riboflavin revealed preference for Zn2+ with the eucaryotic enzymes and delimited effects of inhibitors that relate to drug action. Flavin adenine dinucleotide synthetase, separable from flavokinase in mammals, prefers Mg2+. Specifics for binding and function of flavocoenzymes were delineated for several flavoproteins. The flavin mononucleotide-dependent oxidase that converts the 5'-phosphates of pyridoxine and of pyridoxamine to pyridoxal phosphate is a connection between riboflavin and B-6 that we characterized in mechanistic detail and found to be the primary control point for conversion of B-6 to its coenzyme. Sequencing and cloning of a side-chain oxidase for riboflavin was achieved. Isolation and identification of metabolites of biotin and of lipoic acid, first from bacteria obtained by enrichment culture and then from mammals, provided seminal information on catabolic pathways involved, as have our other studies with flavin catabolites isolated from milk and urine. PMID- 10721898 TI - Probing the mechanisms of the biological intermolecular transfer of reduced flavin. AB - NAD(P)H-flavin oxidoreductases [flavin reductases (FR)] are a class of enzymes capable of producing reduced flavin for bacterial bioluminescence and other biological processes. Bacterial luciferase utilizes oxygen, reduced FMN (FMNH2) and a long-chain aliphatic aldehyde as substrates for light emission. The Vibrio harveyi luciferase and FRP (for which we have cloned the gene and determined the crystal structure) is a model for the elucidation of the reduced flavin transfer mechanism using both a flavin reductase single-enzyme assay monitoring the NADPH oxidation and a flavin reductase-luciferase coupled assay measuring bioluminescence intensity or quantum output. The FRP exhibits a ping-pong kinetic pattern in the single-enzyme assay but changes to a sequential pattern in the coupled assay. Furthermore, FMN at >2x10(-6) mol/L reduced both the light intensity and quantum yield of the coupled reaction by noncompetitively inhibiting NADPH and competitively inhibiting luciferase. These results support a scheme in which the luciferase forms specific complex(es) with FRP. Indeed, such complexes were shown by fluorescence anisotropy to exist between luciferase and monomeric FRP either in the holo- or apoenzyme form. Furthermore, the reduced flavin cofactor of FRP is transferred directly to luciferase for bioluminescence, whereas the reduced flavin product of FRP is inefficient in supporting the luminescence reaction. The mechanism of reduced flavin transfer is apparently flavin and flavin reductase specific. PMID- 10721899 TI - Effects of vitamin B-6 on (n-3) polyunsaturated fatty acid metabolism. AB - To investigate interactions between vitamin B-6 and fatty acid metabolism, male Wistar rats were fed a vitamin B-6 (B-6)-deficient diet consisting of 70% vitamin free casein and 10% perilla oil [approximately 63% alpha-linolenic acid, (n-3)] for 5 wk. The amounts of linoleic acid (n-6) and arachidonic acid (n-6) in the B 6-deficient group changed only slightly compared with those in a pair-fed control group. The amount of linoleic acid increased and arachidonic acid decreased in the plasma total lipid fraction, and the ratios of both eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) in the B-6-deficient group were significantly lower than for the controls. The ratios of alpha-linolenic acid and EPA were higher, and DHA lower, in the B-6-deficient group than in the pair-fed control group in the total lipid as well as phospholipid fractions in liver microsomes. The activity of delta6-desaturase was significantly lower in the B-6 deficient group than in the pair-fed control group (approximately 64%), and acyl CoA oxidase activity, an initial enzyme of the peroxisomal beta-oxidation pathway, was reduced by approximately 80% in the B-6-deficient group. These data suggest that B-6 deficiencies impair the metabolism of (n-3) PUFA from alpha linolenic acid to EPA and DHA with the most pronounced reduction in the production of DHA. PMID- 10721900 TI - Utilization of biotin in proliferating human lymphocytes. AB - Lymphocytes are part of the immune system and respond to antigenic stimulation with proliferation. We sought to determine whether mitogen-stimulated, proliferating lymphocytes increase the cellular uptake of biotin and, if so, to identify mechanisms that mediate the increase. Lymphocytes were isolated from human peripheral blood; proliferation of lymphocytes was induced by incubation with pokeweed lectin, concanavalin A or phytohemagglutinin. Biotin uptake was quantitated by determination of [3H] uptake into the lymphocytes during incubation with [3H]biotin after establishing that [3H]biotin is not metabolized within the lymphocytes during the incubation period (<5%). Biotin uptake into proliferating lymphocytes increased to 278-722% of the control values for nonproliferating lymphocytes. Kinetic analysis of biotin transport provided evidence that the increase is mediated by an increased number of transporters on the cell surface rather than by an increase in transporter affinity. Cycloheximide, an inhibitor of protein synthesis, completely suppressed the mitogen-stimulated increase in biotin transport. This observation is consistent with the hypothesis that proliferating lymphocytes increase biotin uptake by increasing the synthesis of new transporters. Biotin affinity and structural specificity were similar in proliferating and nonproliferating lymphocytes, suggesting that mitogens induced an increase in the number of the same transporter molecule that mediates transport in unstimulated lymphocytes. Mitogen stimulated lymphocytes exhibited 2.5 times greater activities of biotin-dependent beta-methylcrotonyl-CoA carboxylase compared with time 0 (at 72 h after addition of mitogen). This observation is consistent with the hypothesis that proliferating lymphocytes increase biotin uptake at least in part to provide adequate coenzyme for biotin-dependent carboxylases. PMID- 10721901 TI - Vitamin nutrition and gastroesophageal cancer. AB - Nitrosamines have been suspected in the etiology of esophageal/gastric cardia cancer in the high incidence area of Linxian of the Henan Province in northern China, but marginal deficiencies in riboflavin, vitamins A and C, and other micronutrients may also be involved. A joint U.S.-China nutritional intervention study with investigators from the Cancer Institute of the Chinese Academy of Medical Sciences and the U.S. National Cancer Institute tested the effects of the following four combinations of nutrients on 29,584 subjects in an eight-group design: 1) retinol and zinc; 2) riboflavin and niacin; 3) vitamin C and molybdenum; and 4) vitamin E, beta-carotene and selenium. Supplementation with Group 4 nutrients significantly decreased mortality rate from stomach cancer, primarily due to the decrease in deaths resulting from adenocarcinomas of the gastric cardia; it lowered the total mortality rate and showed signs of other beneficial effects. Another study of nutrition and gastric cancer in a high incidence area of Linqu of the Shangdong province in northern China (in collaboration with the Beijing Institute for Cancer Research and the U. S. National Institutes of Health) found significantly lower serum concentrations of vitamin C and beta-carotene among individuals with intestinal metaplasia; an intervention trial with vitamins C and E and selenium (combined) is ongoing in Linqu. Other studies are also elucidating the mechanisms for the pathogenesis of adenocarcinoma at the gastroesophageal junction with the use of a rat model. Such studies are expected to shed light on the etiology and prevention of gastroesophageal cancers in humans. PMID- 10721902 TI - Lipases and carboxylesterases: possible roles in the hepatic utilization of vitamin A. AB - The formation and hydrolysis of retinyl esters are key processes in the metabolism of the fat-soluble micronutrient vitamin A. Long-chain acyl esters of retinol are the major chemical form of vitamin A (retinoid) stored in the body. Although retinyl esters are found in a variety of tissues and cell types, most of the total body retinoid is accounted for by the retinyl esters stored in the liver. Thus, these esters represent the major endogenous source of retinoid that can be delivered to peripheral tissues for conversion to biologically active forms. This paper summarizes the current state of our knowledge about the identity, function and regulation of the hepatic enzymes that are potentially involved in catalyzing the hydrolysis of retinyl esters. These enzymes include several known and characterized lipases and carboxylesterases. PMID- 10721903 TI - Causes of iron and zinc deficiencies and their effects on brain. AB - Low consumption of foods rich in bioavailable iron (Fe) and zinc (Zn) such as meat, particularly red meat, and high consumption of foods rich in inhibitors of Fe and Zn absorption, such as phytate, certain dietary fibers and calcium, cause Fe and Zn deficiencies. Neuropsychologic impairment is one of several potential outcomes of these deficiencies. PMID- 10721904 TI - Developmental sequel from early nutritional deficiencies: conclusive and probability judgements. AB - Results from quasi-experimental longitudinal studies of children and from experimental research with animal models have led several investigators to state that early iron deficiency anemia leaves a permanent cognitive deficit. However, neither source of information provides a basis for such a claim. Some key confounders were not controlled by the quasi-experimental studies, and the external validity of the animal data is questionable. Further, three decades of research on the functional consequences of protein-energy malnutrition have shown that the social environment moderates the effects of an early nutritional insult; it can keep such effect unchanged, or increase or decrease its severity. The prediction of later life on the basis of a particular nutritional event carries a large error factor, which suggests that the search would be more fruitful if we tracked probability statements. PMID- 10721905 TI - Moderate zinc-iron deprivation influences behavior but not growth in adolescent rhesus monkeys. AB - Primate species demonstrate a prolonged period of development before reproductive maturity that includes distinctive periods of rapid growth in the late fetal, late infancy and early adolescent stages. Rhesus monkeys resemble humans in this discontinuous pattern of growth and also in its relationship to brain development. Studies of zinc deprivation in rhesus monkeys have suggested an important relationship among growth rate, nutrient status and behavioral performance in infancy as well as adolescence. Recently, moderate combined zinc and iron deprivation (intake 0.2 mg Zn and 0.8 mg Fe/d, compared with control intake of 2.9 mg Zn and 1.7 mg Fe/d) during the adolescent growth spurt (29-32 mo. of age) of female rhesus monkeys (n = 8/group) was shown to influence behavior without affecting growth. Behavioral assessments included the Continuous Performance Test, the Delayed Nonmatch to Sample Test and activity (measured with an actimeter). The behavioral syndrome was characterized by reduced activity, reduced participation in behavioral testing and slower response. These changes could be reversed or prevented to some extent by altering the diet to include tablets of powdered beef (adding approximately 1.7 mg Zn and 0.7 mg Fe to daily intake). The study suggests that behavior may be sensitive to the quality of the diet available during the period of rapid adolescent growth and development. PMID- 10721906 TI - Dietary zinc and iron sources, physical growth and cognitive development of breastfed infants. AB - Iron and zinc are trace minerals that are of critical importance to the young infant for normal growth and development. Exclusive feeding of human milk provides adequate amounts of both of these nutrients for normal term infants for approximately the first 6 mo. of life. Current recommendations for introduction of complementary foods at this age do not emphasize the order of introduction of specific foods because the infant's gastrointestinal tract is considered mature at this time. Consideration of nutritional needs at 6 mo. has generally focused on the increasing risk of iron deficiency the longer the diet is without an additional source of iron. Recently, there has been more recognition of the risk of zinc deficiency in the second half of the first year of life in breastfed infants. Review of common feeding practices indicates that early complementary foods are typically iron fortified but low in zinc. Several studies have now investigated the effects of meat as an earlier complementary food on iron and zinc status. Results of these studies, although requiring further verification, suggest that increased meat intake by breastfed infants >6 mo. old would adequately support both iron and zinc requirements. PMID- 10721907 TI - Behavioral data and methodology issues in studies of zinc nutrition in humans. AB - Despite the widespread incidence of childhood zinc (Zn) deficiency and strong evidence that Zn deprivation during periods of rapid growth affect brain development and behavior in animals, there is little research on the behavioral effects of Zn deficiency in children or adults. A brief review of previous human studies is followed by more detailed discussion of recent studies of Chinese and Mexican-American children, which showed beneficial effects of Zn repletion on neuropsychologic function. Methodology issues are reviewed and recommendations are made to assess the following: 1) a broad range of cognitive, psychomotor, emotional and social factors; 2) performance in the presence of secondary stressors to approximate real-world conditions more accurately; 3) continuous activity and rest in older children by the use of electronic activity monitors; and 4) electrophysiologic measures of brain function. It is concluded that research on cognition, behavioral activity and brain electrophysiology as outcomes of Zn deficiency and response to improved Zn nutrition is critical, given that Zn deficiency is common in both developing and developed countries. PMID- 10721908 TI - Mechanisms of homocysteine toxicity on connective tissues: implications for the morbidity of aging. AB - It is proposed that chronic moderate hyperhomocysteinemia has a causal role in a number of common diseases of late life, including occlusive vascular disease, cognitive decline, senile osteoporosis and presbyopia. These diseases are seen as clinical counterparts of the main manifestations of homocystinuria (vascular occlusions of arteries and veins, mental retardation, osteoporosis and ectopia lentis, respectively) that develop only after many years of exposure to moderately elevated homocysteine (Hcy) levels. The multisystem toxicity of Hcy is attributed to its spontaneous chemical reaction with many biologically important molecules, primarily proteins. The formation of these Hcy-adducts is dependent on time and Hcy concentration and leads to loss or diminution of function of the derivatized molecules. Irreversible homocysteinylation of long-lived proteins should lead to cumulative damage and progressive clinical manifestations. Fibrillin 1 is seen as the paradigm of extracellular connective tissue proteins that are specially susceptible to Hcy (and presumably Hcy thiolactone) attack. The prominent presence of epidermal growth factor (EGF)-like domains in fibrillin and in many other extracellular proteins of the coagulation, anticoagulation, and lipoprotein transport pathways, all of which malfunction in hyperhomocysteinemia, suggests that EGF-like domains may be preferential sites of homocysteinylation. PMID- 10721909 TI - Homocysteine and endothelial dysfunction: a link with cardiovascular disease. AB - The nature of the link between homocysteine and cardiovascular disease has not yet been clearly established. Impaired endothelium-independent vasodilatation is an early feature of vascular disease. In human studies, methionine loading, which acutely elevates plasma homocysteine, induces endothelial dysfunction. Folate therapy, which lowers homocysteine, enhances endothelial function. This is consistent with, but not proof of, homocysteine toxicity to endothelium in vivo. Homocysteine, in high concentration, can induce endothelial dysfunction in vitro. This is accompanied by increased superoxide production, which when inhibited, restores normal endothelial function. These observations suggest that homocysteine may induce vascular endothelial dysfunction by a mechanism involving reactive oxygen species. PMID- 10721910 TI - Inhibition of endothelial cell thromboresistance by homocysteine. AB - Homocysteine (HC) is a highly reactive thiol intermediate in amino acid metabolism, which can modify the function of endothelial cells in a myriad of ways. In vitro, homocysteine can inhibit the thromboresistance properties of the endothelial cell by induction of procoagulant factors, inactivation of natural anticoagulant systems, and suppression of vasodilatory and platelet-modulating factors. HC also inhibits the fibrinolytic system by impairing the ability of the endothelial cell to bind tissue plasminogen activator (t-PA), by interacting directly with the t-PA binding "tail" domain of its endothelial cell receptor, annexin II. Moreover, HC influences endothelial cell gene expression as exemplified by induction of the elongation factor-1 family of polypeptides, which promote polypeptide chain elongation during mRNA translation. Induction of EF-1 subunits alpha, beta, gamma and delta by homocysteine is associated with increased turnover of at least one free thiol-containing protein, suggesting that up-regulation of these subunits may represent a mechanism for replacement of damaged or modified proteins. A more complete understanding of the diverse effects of homocysteine on endothelial cell function may provide important clues to the precise role homocysteine may play in the initiation and progression of vascular disease. PMID- 10721911 TI - Homocysteine thiolactone: metabolic origin and protein homocysteinylation in humans. AB - Homocysteine thiolactone, an intramolecular thioester of homocysteine, is synthesized by methionyl-tRNA synthetase in an error-editing reaction that prevents translational incorporation of homocysteine into proteins. The synthesis of thiolactone occurs in all human cell types investigated. An increase in homocysteine levels leads to elevation of thiolactone levels in human cells. In cultured human cells and in human serum, homocysteine thiolactone reacts with proteins by a mechanism involving homocysteinylation of protein lysine residues. The homocysteinylation leads to protein damage. A calcium-dependent homocysteine thiolactonase, tightly associated with HDL in human serum, may prevent protein damage by detoxifying thiolactone. PMID- 10721912 TI - Considerations for use of probiotic bacteria to modulate human health. AB - Oral consumption of probiotic bacteria has the potential to support the health of American consumers. This paper will discuss the rationale of the probiotic theory, several health targets for probiotic bacteria, probiotic products in the U.S. and, finally, issues pertaining to communication about probiotic products to the consumer. PMID- 10721913 TI - Aspects of in vitro and in vivo research approaches directed toward identifying probiotics and prebiotics for human use. AB - The microbiota of the human gastrointestinal tract plays a key role in nutrition and health. Through the process of fermentation, gut bacteria metabolize various substrates (principally dietary components) to end products such as short-chain fatty acids and gases. This anaerobic metabolism is thought to contribute positively toward host daily energy requirements. However, under certain circumstances, the fermentative process may produce undesirable metabolites. This may cause the onset of gut disorders that can be manifest through both acute and chronic conditions. Moreover, the gut flora may become contaminated by transient pathogens that serve further to upset the normal community structure. There has been a recent increase in the use of dietary components that help to maintain, or even improve, the gut microflora "balance." Probiotics are live microbial feed supplements added to appropriate food vehicles (usually fermented milks), whereas prebiotics are dietary carbohydrates that have a selective metabolism in the colon and serve to increase numbers of bacteria seen as desirable. Because of their purported health-promoting properties, lactic acid-producing bacteria, including bifidobacteria, are the usual target organisms. The market value and biological potential of both approaches are enormous. This article will summarize how efficacious types can be identified. PMID- 10721914 TI - The role of probiotic cultures in the control of gastrointestinal health. AB - The use of probiotics to enhance intestinal health has been proposed for many years. Probiotics are traditionally defined as viable microorganisms that have a beneficial effect in the prevention and treatment of specific pathologic conditions when they are ingested. There is a relatively large volume of literature that supports the use of probiotics to prevent or treat intestinal disorders. However, the scientific basis of probiotic use has been firmly established only recently, and sound clinical studies have begun to be published. Currently, the best-studied probiotics are the lactic acid bacteria, particularly Lactobacillus sp. and Bifidobacterium sp. However, other organisms used as probiotics in humans include Escherichia coli, Streptococcus sp., Enterococcus sp., Bacteroides sp., Bacillus sp., Propionibacterium sp. and various fungi. Some probiotic preparations contain mixtures of more than one bacterial strain. Probiotics have been examined for their effectiveness in the prevention and treatment of a diverse spectrum of gastrointestinal disorders such as antibiotic associated diarrhea (including Clostridium difficile-associated intestinal disease), infectious bacterial and viral diarrhea (including diarrhea caused by rotavirus, Shigella, Salmonella, enterotoxigenic E. coli, Vibrio cholerae and human immunodeficiency virus/acquired immunodeficiency disorder, enteral feeding diarrhea, Helicobacter pylori gastroenteritis, sucrase maltase deficiency, inflammatory bowel disease, irritable bowel syndrome, small bowel bacterial overgrowth and lactose intolerance. Probiotics have been found to inhibit intestinal bacterial enzymes involved in the synthesis of colonic carcinogens. There are many mechanisms by which probiotics enhance intestinal health, including stimulation of immunity, competition for limited nutrients, inhibition of epithelial and mucosal adherence, inhibition of epithelial invasion and production of antimicrobial substances. Probiotics represent an exciting prophylactic and therapeutic advance, although additional investigations must be undertaken before their role in intestinal health can be delineated clearly. PMID- 10721915 TI - Probiotic immunomodulation in health and disease. AB - Probiotics, microorganisms that have a favorable influence on physiologic and pathological processes of the host by their effect on the intestinal flora, may play a role in improving human health. One of the putative effects is the modulation of immune function. Thus, the mucosal immune system and methods to assess its function are reviewed briefly. Probiotic modulation of humoral, cellular and nonspecific immunity is reviewed, with emphasis placed on immune response in disease models. There are very few reports of human intervention studies with probiotics. However, some of the possible future directions for research with respect to probiotics, immunity, and human health are discussed. Although the application of probiotics has demonstrated trends with respect to altered aspects of immune response, the underlying mechanisms by which that occurs are unclear. PMID- 10721916 TI - The role of probiotic cultures in the prevention of colon cancer. AB - Risk factors for colon cancer include both hereditary and environmental factors. Dietary patterns represent controllable risk factors for the development of colon cancer. Much attention has focused on decreasing colon cancer risk through increasing intake of dietary fiber; recently, this has included interest in the consumption of prebiotics and probiotics. Because factors involved in the initiation and promotion of colon cancer might be separated in time from actual tumor development, it is difficult to choose "outcomes" or "end points" that are definitive indicators of efficacy of probiotics or prebiotics. Studies that have explored the cause-effect relationship directly have used animal models. In this review, we have confined our discussion to animal studies from the last 10 years that have examined most directly the relationship between prebiotic and probiotic consumption and colon cancer development. To present the consensus of these studies first, it appears that probiotics with or without prebiotics have an inhibitory effect on the development of aberrant crypts (precancerous lesions) and tumors in animal models. The effect is not completely consistent and is small in some studies, but this may represent a dose or time effect. PMID- 10721917 TI - Probiotic bacteria: today and tomorrow. AB - This paper provides an overview of the key issues raised during this symposium. Probiotic cultures have been associated historically with cultured milks and dairy products, from which there is substantial evidence for positive effects on human health and general well-being. PMID- 10721918 TI - Overview: the clinical perspective. PMID- 10721919 TI - Adverse host responses to bacterial toxins in human infants. AB - Bacterial toxin interaction with the intestinal epithelium is regulated developmentally as well as by nutritional factors. It is the binding of bacterial toxins to the epithelium followed by several events that forms the basis of infantile diarrhea, a leading cause of morbidity and mortality world-wide. There has been increasing interest in bacterial toxin interaction with the enterocyte, postreceptor events that follow and the effect of developmental regulation on necrotizing enterocolitis. Diet and environmental factors can provide a major influence on bacterial-enterocyte interaction. Particularly important is the role of breast milk and its constituents, as well as probiotics, in this regard. The purpose of this review is to provide a brief overview on this complex interaction. PMID- 10721920 TI - Modulation of the gastrointestinal tract of infants by human milk. Interfaces and interactions. An evolutionary perspective. AB - Human milk contains agents that affect the growth, development and functions of the epithelium, immune system or nervous system of the gastrointestinal tract. Some human and animal studies indicate that human milk affects the growth of intestinal villi, the development of intestinal disaccharidases, the permeability of the gastrointestinal tract and resistance to certain inflammatory/immune mediated diseases. Moreover, one cytokine in human milk, interleukin (IL)-10, protects infant mice genetically deficient in IL-10 against an enterocolitis that resembles necrotizing enterocolitis (NEC) in human premature infants. There are seven overlapping evolutionary strategies regarding the relationships between the functions of the mammary gland and the infant's gastrointestinal tract as follows: 1) certain immunologic agents in human milk compensate directly for developmental delays in those same agents in the recipient infant; 2) other agents in human milk do not compensate directly for developmental delays in the production of those same agents, but nevertheless protect the recipient; 3) agents in human milk enhance functions that are poorly expressed in the recipient; 4) agents in human milk change the physiologic state of the intestines from one adapted to intrauterine life to one suited to extrauterine life; 5) some agents in human milk prevent inflammation in the recipient's gastrointestinal tract; 6) survival of human milk agents in the gastrointestinal tract is enhanced because of delayed production of pancreatic proteases and gastric acid by newborn infants, antiproteases and inhibitors of gastric acid production in human milk, inherent resistance of some human milk agents to proteolysis, and protective binding of other factors in human milk; and 7) growth factors in human milk aid in establishing a commensal enteric microflora. PMID- 10721921 TI - Interactions mediating bacterial translocation in the immature intestine. AB - Systemic disease caused by transmucosal passage of enterovirulent bacteria and toxins from the gut lumen into the mesenteric lymph nodes (MLN) is reviewed, with particular concern for bacterial interactions in the developing gut of premature newborns. Anaerobic bacteria are rarely observed to translocate to the MLN. Bifidobacterial strains have been tested for their abilities to adhere to enterocyte-like Caco-2 cells in culture. We have investigated the inhibitory effect of adherent human bifidobacterial strains against colonization by a number of diarrheagenic bacteria (Escherichia coli O157; Salmonella typhimurium) and viruses (murine and rhesus rotavirus), in various in vitro and in vivo models. The phagocytic cell (macrophage) may be a key factor in bacterial translocation (BT). Human breast milk contains abundant bioactive substances (immunologic, nutritional) that provide protective effects through inhibition of bacterial overgrowth and BT. New biotherapeutic therapies that stimulate beneficial anaerobic microflora (Lactobacillus, Bifidobacterium) are promising avenues of research to combat BT in disease treatment. PMID- 10721922 TI - Improving adolescent iron status before childbearing. PMID- 10721923 TI - Iron requirements in adolescent females. AB - Adolescence is characterized by a large growth spurt and the acquisition of adult phenotypes and biologic rhythms. During this period, iron requirements increase dramatically in both boys and girls as a result of the expansion of the total blood volume, the increase in lean body mass and the onset of menses in young females. The overall iron requirements increase from a preadolescent level of approximately 0.7-0.9 mg Fe/d to as much as 2.2 mg Fe/d or perhaps more in heavily menstruating young women. These increased requirements are associated with the timing and size of the growth spurt as well as sexual maturation and the onset of menses. The available data on iron intakes in adolescents suggest that adolescent girls are unlikely to acquire substantial iron stores during this time period because intakes may average as little as 10-11 mg Fe/d. The bioavailability from diets in developing and industrialized countries indicates a negative iron balance is likely in many female populations. The low iron stores in these young women of reproductive age will make them susceptible to iron deficiency anemia during pregnancy because dietary intakes alone are insufficient, in most cases, to meet the requirements of pregnancy. PMID- 10721924 TI - Anemia, iron and pregnancy outcome. AB - When maternal anemia is diagnosed before midpregnancy, it has been associated with an increased risk of preterm delivery. Maternal anemia detected during the later stages of pregnancy, especially the third trimester, often reflects the expected (and necessary) expansion of maternal plasma volume. Third-trimester anemia usually is not associated with increased risk of preterm delivery. High hemoglobin concentration, elevated hematocrit and increased levels of serum ferritin late in pregnancy, however, all have been associated with increased preterm delivery. This increased risk may reflect in part the failure to expand maternal plasma volume adequately, thus diminishing appropriate placental perfusion. Although controlled trials of iron supplementation during pregnancy have consistently demonstrated positive effects on maternal iron status at delivery, they have not demonstrated reductions in factors that are associated with maternal anemia, i.e., increased risk of preterm delivery and infant low birth weight. One reason for discordant findings may be the exclusion of many gravidas with iron deficiency from these trials or the data concerning gravidas with pregnancy outcomes such as preterm delivery from the analysis. Finally, recent concerns have been voiced about harmful effects of iron supplementation during pregnancy. No adverse effects of iron supplementation on pregnancy outcome have been demonstrated to date. Questions about the efficacy of iron supplementation during pregnancy for reducing adverse outcomes such as preterm delivery and side effects from iron supplementation, including the potential for oxidation of lipids and DNA, require further research in iron-deficient women. PMID- 10721925 TI - The potential impact of iron supplementation during adolescence on iron status in pregnancy. AB - Iron deficiency anemia (IDA) during pregnancy is associated with significant morbidity for mothers and infants. Over 50% of pregnant women in developing countries suffer from IDA. It is also prevalent among adolescent girls because the growth spurt and onset of menstruation increase iron requirements. Women who conceive during or shortly after adolescence are likely to enter pregnancy with low or absent iron stores or IDA. Iron supplementation during adolescence is one of the new strategies advocated to improve iron balance in pregnancy. However, iron requirements are highest in the second and third trimesters and the model described here indicates that iron balance at this stage depends more on adequate intakes of bioavailable iron than on the size of the iron stores at conception. Furthermore, although supplementation will correct anemia and increase iron stores in girls, the positive effect on iron status will be temporary if their diets do not contain adequate bioavailable iron. Although iron status in early pregnancy may be improved if the period of supplementation continues up to the time of conception, supplementation before pregnancy should be viewed as an additional strategy to supplementation during the second and third trimesters. PMID- 10721926 TI - Supplementation with iron and folic acid enhances growth in adolescent Indian girls. AB - The prevalence of anemia is high in adolescent girls in India, with over 70% anemic. Iron-folic acid (IFA) supplements have been shown to enhance adolescent growth elsewhere in the world. To confirm these results in India, a study was conducted in urban areas of Vadodora, India to investigate the effect of IFA supplements on hemoglobin, hunger and growth in adolescent girls 10-18 y of age. Results show that there was a high demand for IFA supplements and >90% of the girls consumed 85 out of 90 tablets provided. There was an increment of 17.3 g/L hemoglobin in the group of girls receiving IFA supplements, whereas hemoglobin decreased slightly in girls in the control group. Girls and parents reported that girls increased their food intake. A significant weight gain of 0.83 kg was seen in the intervention group, whereas girls in the control group showed little weight gain. The growth increment was greater in the 10- to 14-y-old age group than in the 15- to 18-y-old group, as expected, due to rapid growth during the adolescent spurt. IFA supplementation is recommended for growth promotion among adolescents who are underweight. PMID- 10721927 TI - Reaching young Indonesian women through marriage registries: an innovative approach for anemia control. AB - In an effort to build iron stores before pregnancy and reduce the high prevalence of anemia in Indonesia, the Ministry of Health/Indonesia and the MotherCare project implemented an anemia control program for newly wed women. As part of an existing program to counsel couples about marriage and require them to obtain tetanus toxoid immunization before obtaining a marriage certificate, women also were counseled to buy and take 30-60 iron-folate (IFA) tablets. Women (n = 344) were enrolled from one of three participating districts in South Kalimantan, Indonesia. At first monitoring, at least 30 d after baseline, 261 women were tested for hemoglobin and asked about their IFA tablet consumption and knowledge of information, education, and communications (IEC) materials promoted through the program. Results showed that there was a decrease in the prevalence of anemia from 23.8 to 14.0% over the course of the program, 98% of women had taken at least some IFA tablets and 56% had taken >30 tablets. PMID- 10721928 TI - Improving dietary intake to prevent anemia in adolescent girls through community kitchens in a periurban population of Lima, Peru. AB - Peru has high rates of iron deficiency anemia. The prevalence is 35% in nonpregnant women of fertile age and 24.7% in adolescent girls in slums of periurban Lima. The major cause of anemia is low intake of dietary iron. A community-based, randomized behavioral and dietary intervention trial was conducted to improve dietary iron intake and iron bioavailability of adolescent girls living in periurban areas of Lima, Peru. Results show that there was a change in knowledge about anemia and improved dietary iron intake in the 71 girls who completed the study compared with the 66 girls in the control group. Although the 9-mo. intervention was not sufficient to improve hemoglobin levels significantly, there appeared to be a protective effect in maintaining the iron status of girls in comparison with the control group. PMID- 10721929 TI - Efficacy and acceptability of two iron supplementation schedules in adolescent school girls in Lima, Peru. AB - To assess the efficacy and acceptability of a daily and intermittent iron supplementation, a double-blind, placebo-controlled trial was conducted in a public school located in periurban Lima, Peru. Adolescent girls (n = 312), 12-18 y old, were randomly assigned to one of the following three groups: 1) 60 mg iron as ferrous sulfate daily from Monday to Friday; 2) 60 mg iron as ferrous sulfate 2 d/wk and 3 d placebo (intermittent); 3) placebo, from Monday to Friday. Field workers gave the girls supplements during school hours for 17 wk; 296 girls completed the trial. Girls took 94% of the expected dose of 85 pills. Few side effects were reported. Postintervention, hemoglobin (Hb), serum ferritin (SF) and free erythrocyte protoporphyrin (FEP) were improved significantly in the iron supplemented groups compared with placebo (P<0.05). Daily supplements led to higher Hb increases than intermittent supplements (P<0.05), but SF and FEP were similar between the two groups. Thus, both iron supplementation schedules were efficacious in preventing iron deficiency in adolescent girls through the school system, and the daily schedule was better than the intermittent schedule at increasing Hb values and reducing anemia. PMID- 10721930 TI - Diet, natural products and cancer chemoprevention. AB - There is considerable scientific evidence to suggest that nutritive and nonnutritive plant-based dietary factors can inhibit the process of carcinogenesis effectively. Cancer chemoprevention involves pharmacologic intervention with synthetic or naturally occurring chemicals to prevent, inhibit or reverse carcinogenesis or prevent the development of invasive cancer. In light of the considerable effort that has been expended by scientists from the academic, governmental and private sectors in identifying, characterizing and utilizing potential cancer chemopreventive agents, it is reasonable to inquire about the progress that has been made to date and the promise that this field holds in the fight against cancer. The symposium entitled Diet, Natural Products and Cancer Chemoprevention: Progress and Promise was therefore organized at Experimental Biology 99 by the American Society for Nutritional Sciences to address in part these two issues. Progress in the development of cancer chemopreventive agents, examples of current clinical and experimental research of particular relevance to cancer prevention and the promise of chemoprevention in effectively contributing to the conquest of cancer were highlighted. PMID- 10721931 TI - Progress in cancer chemoprevention: development of diet-derived chemopreventive agents. AB - Because of their safety and the fact that they are not perceived as "medicine," food-derived products are highly interesting for development as chemopreventive agents that may find widespread, long-term use in populations at normal risk. Numerous diet-derived agents are included among the >40 promising agents and agent combinations that are being evaluated clinically as chemopreventive agents for major cancer targets including breast, prostate, colon and lung. Examples include green and black tea polyphenols, soy isoflavones, Bowman-Birk soy protease inhibitor, curcumin, phenethyl isothiocyanate, sulforaphane, lycopene, indole-3-carbinol, perillyl alcohol, vitamin D, vitamin E, selenium and calcium. Many food-derived agents are extracts, containing multiple compounds or classes of compounds. For developing such agents, the National Cancer Institute (NCI) has advocated codevelopment of a single or a few putative active compounds that are contained in the food-derived agent. The active compounds provide mechanistic and pharmacologic data that may be used to characterize the chemopreventive potential of the extract, and these compounds may find use as chemopreventives in higher risk subjects (patients with precancers or previous cancers). Other critical aspects to developing the food-derived products are careful analysis and definition of the extract to ensure reproducibility (e.g., growth conditions, chromatographic characteristics or composition), and basic science studies to confirm epidemiologic findings associating the food product with cancer prevention. PMID- 10721932 TI - Tea and tea polyphenols in cancer prevention. AB - The inhibitory action of tea (Camellia sinensis) and tea components against cancer formation has been demonstrated in different animal models involving different organ sites in many laboratories. The possible preventive activity of tea against cancer in humans, however, is not clear. A critical question is whether the information obtained from animal studies is applicable to humans because of possible species differences or the difference in the quantity of tea used in animal studies and that consumed by humans. This article will discuss the results from animal studies and possible cancer inhibitory mechanisms that might be applicable to human cancer prevention. To provide a basis for more quantitative analyses of the effect of tea on carcinogenesis, the levels of tea polyphenols in blood, urine and tissue samples have been analyzed, and the pharmacokinetic properties of tea polyphenols studied. Studies with cell lines have demonstrated that tea polyphenols affect signal transduction pathways, inhibit cell proliferation and induce apoptosis, but the effective concentrations are usually much higher than those observed in blood and tissues. More mechanistic studies in these areas will help us to understand the inhibitory action of tea against carcinogenesis and provide background for evaluating the effects of tea consumption on human carcinogenesis. PMID- 10721933 TI - Advances in the development of retinoids as chemopreventive agents. AB - With the inclusion of brief discussions of retinoid drug development in animal carcinogenesis models (e.g., skin, breast, oral cavity, lung, prostate or bladder) and clinical trials (e.g., head and neck or cervix), this review will focus on recent advances in retinoid molecular targeting studies designed primarily to develop retinoids with reduced toxicity, while maintaining or enhancing activity in the context of chemoprevention. Major current retinoid molecular targets include the six known nuclear retinoid receptors (RAR and RXR). Receptor numbers, distinct functions, tissue-expression patterns, ligand specificities, functional redundancy and regulation of multiple pathways make retinoid signaling highly complex. Development of receptor-selective synthetic retinoids is a major focus of molecular retinoid development. RAR heterodimerize with RXR and mediate classic retinoid activity/toxicity. RXR are more promiscuous, heterodimerizing with several other members of the steroid receptor superfamily [e.g., peroxisome proliferator-activated receptors (PPAR) or vitamin D receptors]. RXR-selective ligands are less toxic and more active in animal breast cancer prevention studies and less toxic than RAR ligands in clinical trials. Other new avenues of retinoid molecular drug development include newly identified retinoid-regulated genes, orphan-receptor ligands/functions, novel retinoid mechanisms involving potent receptor-independent apoptosis-inducing activity (e.g., 4-HPR or anhydroretinol), synergistic combinations [e.g., RXR agonists plus selective estrogen receptor modulators (SERM)], activity in other diseases and novel delivery systems. PMID- 10721934 TI - History of nutrition symposium: trace element nutrition and human health. PMID- 10721935 TI - The selenium-coxsackievirus connection: chronicle of a collaboration. AB - This review provides a historical account of a collaboration established between a nutritionist and a virologist to investigate the interrelationship of host nutritional status and viral virulence. The parties to this collaboration consider themselves specialists in the fields of antioxidant nutrition and viral immunology, respectively. The advantages of such talent pooling are discussed (rapid startup, well-focused experimentation, ability to visualize the "big picture"), as are some of the disadvantages (limited common scientific vocabulary, proper apportioning of credit, lack of institutional infrastructure to house such efforts). The common perception that some of the most exciting science occurs when the advancing edges of two disparate disciplines intersect is borne out by this project because host nutriture was shown for the first time to influence the genetic make-up of an invading viral pathogen. Encouragement of joint cooperative ventures should have a high priority as demanded by increasingly difficult scientific problems and as desired by scientists themselves who wish to see their research progress more quickly. PMID- 10721936 TI - Cardiovascular disease from copper deficiency--a history. AB - Although the nutritional essentiality of copper was established in 1928, a preoccupation with hematology delayed the discovery of cardiovascular disease from copper deficiency for more than a decade. Anatomical studies of several species of deficient animals revealed, interalia, aortic fissures and rupture, arterial foam cells and smooth muscle migration, cardiac enlargement and rupture, coronary artery thrombosis and myocardial infarction. Abnormal biochemistry in deficiency probably contributes to these lesions, e.g., decreased activities of lysyl oxidase and superoxide dismutase which result in failure of collagen and elastin crosslinking and impaired defense against free radicals. Copper deficiency also decreases copper in hearts and other organs and cells and increases cholesterol in plasma. Abnormal physiology from deficiency includes abnormal electrocardiograms, glucose intolerance and hypertension. People with ischemic heart disease have decreased cardiac and leucocyte copper and decreased activities of some copper-dependent enzymes. Copper depletion experiments with men and women have revealed abnormalities of lipid metabolism, blood pressure control, and electrocardiograms plus impaired glucose tolerance. The Western diet often is as low in copper as that proved insufficient for these people. Knowledge of nutritional history can be useful in addressing contemporary nutritional problems. PMID- 10721937 TI - Iodine and neuropsychological development. AB - The establishment of the essential link among iodine deficiency, thyroid function and brain development has emerged from a fascinating combination of clinical, epidemiologic and experimental studies. The central human phenomenon that focuses this relationship is the condition of endemic cretinism, described from the Middle Ages and characterized in its fully developed form by severe brain damage, deaf mutism and a spastic state of the hands and feet. The demonstration of the prevention of cretinism in a double-blind controlled trial with injections of iodized oil in Papua New Guinea (1966-1970) established the causal role of iodine deficiency in cretinism by an effect on the developing fetal brain. Cretinism could not be prevented unless the iodized oil was given before pregnancy. Iodine deficiency is now regarded by the WHO as the most common preventable cause of brain damage in the world today, with at least 30 million suffering from this preventable condition. Since 1986 the international NGO, the International Council for Control of Iodine Deficiency Disorders, has worked closely as an expert group with WHO and UNICEF in assisting countries with a program of universal salt iodization for the elimination of iodine deficiency as a cause of brain damage by the year 2000. In 1996, WHO reported that 56% of the population of 83 developing countries now had adequate access to iodized salt. This represents an increase of 750 million since 1990 with protection of 12 million children. PMID- 10721938 TI - History of zinc as related to brain function. AB - Zinc (Zn) is essential for synthesis of coenzymes that mediate biogenic-amine synthesis and metabolism. Zn from vesicles in presynaptic terminals of certain glutaminergic neurons modulates postsynaptic N-methyl-D-aspartate (NMDA) receptors for glutamate. Large amounts of Zn released from vesicles by seizures or ischemia can kill postsynaptic neurons. Acute Zn deficiency impairs brain function of experimental animals and humans. Zn deficiency in experimental animals during early brain development causes malformations, whereas deficiency later in brain development causes microscopic abnormalities and impairs subsequent function. A limited number of studies suggest that similar phenomena can occur in humans. PMID- 10721939 TI - What is the optimal method to predict axillary lymph node metastases in breast cancer patients? Is sentinel lymph node biopsy ready to be an alternative to complete axillary dissection? PMID- 10721940 TI - Establishment and analysis of biological characteristics of a human duodenal carcinoma cell line, WDC-1. AB - BACKGROUND: Duodenal carcinoma is very rare and its culture cell lines have rarely been established. METHODS: Tumor cells separated from a surgically resected primary tumor of duodenal carcinoma were put into culture. The patient was an 81-year-old female and had metastatic lymph nodes. We investigated the biological characteristics of the culture cells including in vitro cell kinetics, karyotype, expression of tumor markers and integrins and tumorigenicity and histology in nude mice. RESULTS: A new cell line, designated WDC-1, was established. This duodenal carcinoma cell line proliferated in a monolayered sheet with a doubling time of 50 h. The histological findings of the xenograft in nude mice were similar to those of the primary tumor. WDC-1 cells produced carcinoembryonic antigen and expressed 1 integrin and very late antigen (VLA)-4d in vitro. CONCLUSIONS: A duodenal carcinoma cell line was established, which is rare and may contribute to progress in understanding the biological features of duodenal cancer. PMID- 10721941 TI - Effect of nebulized morphine in cancer patients with dyspnea: a pilot study. AB - BACKGROUND: It is known that opioids may decrease subjective dyspnea. The recent finding that opioid binding sites are present in the peripheral bronchus supports the possibility of a local action of opioids. However, the clinical benefit of nebulized morphine is controversial. The purpose of this study was to confirm the feasibility of nebulized morphine and to evaluate its clinical benefits. PATIENTS AND METHODS: Fifteen cancer patients with dyspnea in the Thoracic Oncology Division and Palliative Care Unit in the National Cancer Center Hospital East were given 20 mg of morphine hydrochloride dissolved in 5 ml of normal saline through an ultranebulizer. The subjective effects were evaluated using a visual analog scale (VAS) immediately before and 60 min after inhalation. Respiratory rate (RR), hemoglobin oxygen saturation (SpO2) and blood pressure also were measured twice at these two time points. A questionnaire about adverse reactions was included. RESULTS: No major adverse reactions such as respiratory depression, sleepiness, nausea or vomiting were observed. VAS was significantly decreased after nebulization (p = 0.005) without any significant change in RR or SpO2. In eight of 15 patients, dyspnea was improved as measured by a decrease in VAS of more than 10%. This correlated with the desire of the patients to continue its use. CONCLUSION: Our preliminary data confirmed the feasibility of nebulized morphine and suggested its possible clinical benefit for dyspneic patients. A randomized controlled study is warranted to exclude a placebo effect and to compare the clinical benefits of nebulized morphine with those of other methods of treatment. PMID- 10721942 TI - Sentinel node biopsy guided by indocyanine green dye in breast cancer patients. AB - BACKGROUND: We aimed to evaluate whether dye-guided sentinel node biopsy is a useful indicator of axillary node involvement in breast cancer patients and whether clinicopathological features affect its success in identifying sentinel nodes. METHODS: Sentinel node biopsy was performed in patients with stage I or II breast cancer using an indocyanin green dye-guided method. RESULTS: We could identify sentinel nodes in 127 (73.8%) of 172 patients. The mean number of sentinel nodes per patient was 1.7 (range, 1-8) and the mean node size was 9.3 mm (range, 3.0-28.0 mm). Of the 127 patients, 40 (31.5%) also had axillary node involvement. In 16 (40.0%) of these, the sentinel node was the only node involved. There was concordance between sentinel node and axillary node status in 122 (96.1%) of the 127 patients. Success in identifying sentinel nodes was not affected by tumor size, operative procedure, histological type of tumor or tumor location; however, the success rate was significantly lower in patients with axillary node involvement (65.7 vs 79.0% in axillary node-negative patients, p = 0.039) and the presence or absence of lymphatic or vascular invasion in the tumor (63.8 vs 78.9% in patients without lymphatic or vascular invasion, p = 0.043). Sentinel nodes could also be identified significantly more frequently in patients under 50 years old (83.3%) than in those over 50 years old (64.8%, p = 0.009). CONCLUSIONS: Sentinel node biopsy guided by indocyanin green dye is an easy technique with an acceptable success rate in detecting sentinel nodes and predicting axillary nodal status. Axillary node status, the presence or absence of lymphatic or vascular invasion in the tumor and patient age affect its success in identifying sentinel nodes. PMID- 10721943 TI - Primary and adjuvant therapy, prognostic factors and survival in 1053 breast cancers diagnosed in a trial of mammography screening. AB - BACKGROUND: As mammographic screening becomes more widespread, larger numbers of tumours are diagnosed while small and node negative. METHODS: We examined detection mode, tumour size, node status, histological type, therapy and outcome in 1053 breast cancers diagnosed in one county of the Swedish Two-County Trial of mammographic screening for breast cancer. RESULTS: Of patients undergoing total mastectomy with axillary dissection, 65% were found to be node negative. For tumours of size 1-9 mm, 95% were node negative. The major effects on survival were tumour size, node status and histological type. Primary adjuvant therapy had no significant association with survival. CONCLUSIONS: The advent of mammography has substantially enhanced the possibilities for less radical treatment. There is an urgent need for therapeutic trials utilizing mammographic-pathological correlations to ascertain in advance which tumours can and which cannot benefit from more radical therapy. PMID- 10721944 TI - Determination of reference values for total PSA, F/T and PSAD according to prostatic volume in japanese prostate cancer patients with slightly elevated serum PSA levels. AB - BACKGROUND: For screening prostate cancer (CAP) using prostate-specific antigen (PSA), the indications of biopsies for patients showing slightly elevated PSA values are still controversial. Furthermore, the dependence of total PSA, free-to total PSA ratio (F/T) and PSA density (PSAD) on prostatic volume in gray zone cases is unclear. METHODS: By analyzing 1913 patients with a serum total PSA ranging from 2.0 to 20 ng/ml, we evaluated the correlation between total PSA and age or prostatic volume and positive predictive value (PPV) in each range for total PSA, age and prostatic volume. Then suitable reference values for total PSA, F/T and PSAD were decided according to prostatic volume. RESULTS: There was no close correlation between PSA and age or volume. The PPV was high in the group with a prostatic volume of 10-30 ml. Prostatic volume was categorized into three groups, <20, 20-40 and > or =40 ml, and reference values for obtaining a sensitivity of 90% were proposed. The reference values of total PSA and PSAD were lowered and that of F/T was raised with increase in prostatic volume. The specificity was very low for the > or =40 ml group. The highest specificity of 36% in PSAD was obtained for the <20 ml group. CONCLUSION: The reference values for total PSA, F/T and PSAD must be changed according to prostatic volume in order to maintain a sufficient diagnostic sensitivity of CAP. Of these parameters, PSAD showed a high specificity in the group with a prostatic volume of <40 ml. PMID- 10721945 TI - The increased accumulation of [18F]fluorodeoxyglucose in untreated prostate cancer. AB - BACKGROUND: To evaluate the clinical usefulness of [18F]fluorodeoxyglucose positron emission tomography (FDG-PET) compared with histopathological grading, clinical stage and serum prostatic specific antigen (PSA) level in the detection and characterization of prostate cancer. METHODS: Forty-four patients with histologically proven prostate cancer and five control subjects with benign prostatic hyperplasia (BPH) were prospectively investigated with FDG-PET prior to treatment. RESULTS: By visual inspection, FDG accumulation was positive in 28 patients with prostate cancer (sensitivity 64%), whereas all were negative in the control group. FDG-PET in three patients with lymph node metastases did not show any high intrapelvic accumulations corresponding to metastatic sites. Among 12 patients with multiple bone metastases which were detected with 99m-HMDP bone scintigraphy, nine (75%) showed moderate to high FDG accumulation at the sites of bone metastases. Quantitatively, FDG accumulation in prostate cancer was significantly higher than in BPH and there was a tendency for FDG uptake of tumors to be higher with higher histological Gleason grades. Furthermore, FDG uptake in tumors with lymph node and/or bone metastasis was significantly higher than that of localized stages. However, the correlation between PSA and FDG uptake in the prostate cancer was very weak for clinical relevance. CONCLUSIONS: Although FDG-PET was not sensitive enough to detect prostate cancer in clinical use, it is suggested that glucose metabolism in prostate cancer tended to be higher in patients with tumors of advanced stages. PMID- 10721946 TI - Minimally invasive thymoma with extensive intravascular growth. AB - A 70-year-old male with grossly non-invasive thymic tumor associated with myasthenia gravis was subjected to thymothymectomy. Microscopic examination showed extensive intravascular tumor extensions into veins of thymic tissue and surrounding muscles and a minute direct invasion of the thymic tissue. Histologically, the tumor showed mixed-type thymoma with polygonal epithelial cells. These pathological findings indicated that the tumor cells extended mainly into vessels beyond the tumor capsule via tumor drainage veins rather than invading neighboring structures. After chemotherapy and mediastinal irradiation, the patient is now in complete remission of myasthenia gravis and is recurrence free 15 months after surgery. PMID- 10721947 TI - Pain-relieving posterior rod fixation with segmental sublaminar wiring for Pancoast tumor invading the vertebrae. AB - We describe the case of a 44-year-old male patient with Pancoast lung cancer invading the vertebrae. Because irradiation did not relieve his symptoms, we conducted tumor resection with posterior rod fixation with segmental sublaminar wiring of the vertebrae. This enabled the patient to walk and to discontinue morphine immediately after surgery. Although the tumor recurred within the region of the fixation 4 months after surgery, the patient complained of no pain until his death. Although Pancoast lung cancer with extensive vertebral invasion cannot be cured surgically, posterior rod fixation with segmental sublaminar wiring with tumor resection can improve a patient's quality of life by providing immediate, long-term pain relief. PMID- 10721948 TI - Primary low-grade lymphoma of mucosa-associated lymphoid tissue of the urinary bladder: a case report with special reference to the use of ancillary diagnostic studies. AB - We report a case of primary low-grade B-cell lymphoma of the mucosa-associated lymphoid tissue (MALT) type of the urinary bladder. The patient, a 77-year-old woman, presented with a sense of urinary retention. An intravenous pyelogram and cystoscopy revealed a wide-based submucosal mass measuring 3 cm in the left wall of the urinary bladder. Histological findings of the tissue obtained by transurethral resection (TUR) showed a dense, monomorphic atypical lymphoid (centrocyte-like) infiltrate with reactive lymph follicles in the subepithelial tissue. Monocytoid and plasmacytoid features were readily evident in a population of these cells. Lymphoepithelial lesions involving the urothelium were also noticed in some areas. These features were strongly suggestive of primary low grade lymphoma of the MALT type. The diagnosis was confirmed by immunohistochemical and flow cytometric studies, both of which showed a clear immunoglobulin restriction to lambda light chain and also by polymerase chain reaction-based assay using a formalin-fixed paraffin-embedded TUR tissue sample, which showed a clonal Ig heavy-chain gene rearrangement. Clinical staging procedures revealed that the tumor was localized in the urinary bladder. The patient has not received chemotherapy and is alive and well with no evidence of recurrence, 3 years after TUR. This case demonstrates that these ancillary tests are worth performing for confirmation of B-cell clonality in TUR tissue samples showing dense B-lymphocytic infiltration. PMID- 10721949 TI - Uterine cervix metastasis from rectal carcinoma: a case report and a review of the literature. AB - A 59-year-old woman underwent a radical hysterectomy for a metastatic uterine cervix tumor caused by rectal carcinoma, which had been previously resected. Metastatic carcinoma from the large bowel to the uterus is rare. A total of 48 patients (including nine Japanese patients) with metastasis from the large bowel to the uterus were reviewed. The metastatic site of the uterus was the cervix in 27 cases and the corpus in 18. The interval between primary carcinoma and the secondary diagnosis was 17 months. The mean survival after the diagnosis of the secondary deposit was 11 months. Our patient died of lymph node, lung, local and bone metastases 7 months after the diagnosis of the secondary deposit. PMID- 10721950 TI - Disconnection of a venous Port-A-Cath followed by embolization after saline flush: rare case report. AB - A 77-year-old man presented with painful swelling of his Port-A-Cath insertion site soon after flushing with normal saline. No discomfort or abnormality was found during the saline flush. A chest roentgenogram showed that the disconnected catheter had separated from the disc and was absent from its original location. The disconnected catheter was found embolized, by chest roentgenogram and CT scan, to the right atrium and hepatic vein. The patient was treated successfully with an X-ray guided extraction of the catheter. The possibility of catheter disconnection with embolization should be considered and a chest roentgenogram performed immediately in cases of rapid swelling of subcutaneous tissue around the port chamber after fluid infusion. PMID- 10721951 TI - Desire or early death in cancer patients and clinical oncology. PMID- 10721952 TI - A strategy for the isolation of DNA fragments containing methylated CpG islands in human adenocarcinomas of the lung. PMID- 10721953 TI - Cancer statistics digest. Comparison of crude and age-standarized death rates in Japan. PMID- 10721954 TI - A preliminary study of photodynamic therapy using verteporfin for choroidal neovascularization in pathologic myopia, ocular histoplasmosis syndrome, angioid streaks, and idiopathic causes. AB - OBJECTIVE: To evaluate short-term safety and the effects on visual acuity and fluorescein angiography of single or multiple sessions of photodynamic therapy with verteporfin for choroidal neovascularization (CNV) not related to age related macular degeneration (AMD), including pathologic myopia, the ocular histoplasmosis syndrome, angioid streaks, and idiopathic causes. DESIGN: A nonrandomized, multicenter, open-label, dose-escalation phase 1 and 2 clinical trial. SETTING: Four ophthalmic centers in Europe and North America providing retinal care. PARTICIPANTS: Thirteen patients with subfoveal CNV due to pathologic myopia, the ocular histoplasmosis syndrome, angioid streaks, or idiopathic causes. METHODS: Standardized protocol refraction, visual acuity testing, ophthalmic examinations, color photographs, and fluorescein angiograms were used to evaluate the results of photodynamic therapy treatments with verteporfin. Follow-up ranged from 12 weeks for patients who were treated once to 43 weeks for patients who were treated up to 4 times. RESULTS: Verteporfin therapy was well tolerated in patients with CNV not related to AMD. No deterioration in visual acuity was observed; most patients gained at least 1 line of vision. Reduction in the size of leakage area from classic CNV was noted in all patients as early as 1 week after verteporfin therapy, with complete absence of leakage from classic CNV in almost half of the patients. Improvement in visual acuity after verteporfin therapy was greatest (+6, +8, and +9 lines) in 3 patients with relatively poor initial visual acuity (between 20/200 and 20/800). Up to 4 treatments were found to have short-term safety even with retreatment intervals as short as 4 weeks. CONCLUSIONS: Treatment of CNV not related to AMD with verteporfin therapy achieves short-term cessation of fluorescein leakage from CNV in a small number of patients without loss of vision. Further randomized clinical trials including a larger number of patients are under way to confirm whether verteporfin therapy is beneficial for subfoveal CNV not related to AMD. PMID- 10721955 TI - Risk factors for glaucoma filtering bleb infections. AB - OBJECTIVE: To determine risk factors for bleb-related ocular infection after glaucoma filtering surgery. METHODS: A case-control study comparing all consecutive cases of glaucoma filtering bleb-related infections (55 eyes of 55 patients) with matched control eyes between January 1, 1990, and June 30, 1998, was performed. Bleb-related infection was classified as blebitis when a mucopurulent infiltrate was identified within the bleb and associated with mild to moderate anterior segment inflammation. Eyes with endophthalmitis had hypopyon, cells in the anterior vitreous cavity, or a positive vitreous biopsy sampling result. A control was selected for each case based on matching of the surgeon, date and type of glaucoma surgery, and type of antifibrotic agent used. Multivariate, matched, case-control logistic regression analysis was performed using age, race, sex, diagnosis, number of previous incisional operations, filtering bleb location, and presence of bleb leak to determine which variables were associated with bleb-related infection. RESULTS: The odds of an eye with a bleb-related infection being seen with a concomitant late-onset bleb leak are 25.8 times the odds of a noninfected eye having a late-onset bleb leak at any time in the postoperative period (P<.001; 95% confidence interval, 2.3-294.1). Other risk factors for bleb-related infection included younger age (P = .05), black race (P = .03), diagnosis of primary open-angle glaucoma (P = .03), and inferior location of the filtering bleb (P = .04). CONCLUSIONS: Late-onset bleb leakage is a significant risk factor for bleb-related infection. The risk of infection may warrant closure of late-onset bleb leaks in selected eyes. PMID- 10721956 TI - Subretinal surgery for choroidal neovascularization in patients with high myopia. AB - OBJECTIVE: To analyze the visual outcome in patients undergoing surgical removal of subfoveal choroidal neovascularization (CNV) in eyes with high myopia. METHODS: We retrospectively reviewed the medical records of 48 consecutive patients with high myopia (> or =6 diopters [D]) who underwent vitrectomy with surgical removal of subfoveal CNV. The patient population consisted of 2 groups. Group 1 included 23 patients with findings only of myopic degeneration, and group 2 included 25 patients with presumed ocular histoplasmosis syndrome and myopia of 6 D or more. RESULTS: In group 1, the visual acuity improved by 2 or more Snellen lines in 9 eyes (39%), decreased in 8 eyes (35%), and remained unchanged in 6 (26%), with a mean follow-up of 24 months (range, 8-60 months). The preoperative visual acuity was 20/40 or better in only 1 eye (4%), but 8 (35%) achieved a final visual acuity of 20/40 or better. In group 2, the visual acuity improved in 16 eyes (64%), was stable in 4 (16%), and deteriorated in 5 (20%), with a mean follow-up of 18 months (range, 6-44 months). Only 3 eyes (12%) had a preoperative visual acuity of 20/40 or better, but 11 (44%) achieved a final visual acuity of 20/40 or better. Recurrence occurred in 13 (57%) of the 23 eyes in group 1 and in 9 (36%) of the 25 eyes in group 2. Univariate analysis demonstrated a significant relation between younger patient age (group 1) and absence of postoperative CNV recurrence (group 2) and an improvement of visual acuity (P<.01). CONCLUSIONS: Surgical removal of CNV may provide visual benefit in selected cases of subfoveal CNV associated with high myopia. The determination of whether surgical intervention is appropriate in these cases requires a prospective, randomized, clinical trial. PMID- 10721957 TI - Hypertension, cardiovascular disease, and age-related macular degeneration. Age Related Macular Degeneration Risk Factors Study Group. AB - OBJECTIVES: To describe a case-control study of risk factors for neovascular and non-neovascular age-related macular degeneration (AMD) and to present findings on associations with systemic hypertension and cardiovascular disease. METHODS: Participants with and without neovascular and non-neovascular AMD were recruited from 11 ophthalmology practices in the New York, NY, metropolitan area. Comprehensive data collection included (1) a standardized interview, (2) blood pressure measurements, and (3) blood samples. Cases and controls were classified from fundus photograph gradings. Polychotomous logistic regression analyses were used to evaluate associations. RESULTS: Classification of 1222 sets of available photographs resulted in the inclusion of a neovascular case group (n = 182), a non-neovascular case group (n = 227), and a control group (n = 235). Neovascular AMD was positively associated with diastolic blood pressure greater than 95 mm Hg (odds ratio [OR] = 4.4), self-reported use of potent antihypertensive medication (OR = 2.1), physician-reported history of hypertension (OR = 1.8), use of antihypertensive medication (OR = 2.5), combinations of self-reported and physician-reported data on hypertension and its treatment (OR = 1. 7), high density lipoprotein level (OR = 2.3), and dietary cholesterol level (OR = 2.2). Non-neovascular AMD was unrelated to hypertension or cholesterol level. No associations were found between either AMD type and other definitions of hypertension or other cardiovascular disease. CONCLUSIONS: These findings suggest that neovascular AMD is associated with moderate to severe hypertension, particularly among patients receiving antihypertensive treatment. They also support the hypotheses that neovascular and non-neovascular AMD may have a different pathogenesis and that neovascular AMD and hypertensive disease may have a similar underlying systemic process. PMID- 10721959 TI - Diode laser ablation for threshold retinopathy of prematurity: short-term structural outcome. AB - OBJECTIVE: To describe short-term structural outcomes and associated ocular complications in premature infants treated with diode laser ablation for retinopathy of prematurity. METHODS: The records of all infants who were diagnosed as having threshold retinopathy of prematurity and treated with diode laser therapy at our hospital from January 1, 1992, through December 31, 1996, were retrospectively reviewed. Sixty-four eyes reached threshold during this period. Three eyes received cryotherapy in addition to laser treatment and were excluded, leaving 61 eyes eligible for review. RESULTS: Of the 61 eyes with threshold disease treated exclusively with diode laser, 4 (7%) had zone I disease and 57 (93%) had zone II disease at the time of initial laser treatment. Three (5%) of the 61 eyes progressed to stage 4 disease (2 eyes, stage 4A; 1 eye, stage 4B). There were no cataracts or other ocular complications noted secondary to laser treatment based on short-term follow-up (mean follow-up, 120 days). CONCLUSION: In this population of infants, diode laser ablation appears to be a safe and effective treatment for threshold retinopathy of prematurity. PMID- 10721958 TI - Combination of clinical factors predictive of growth of small choroidal melanocytic tumors. AB - OBJECTIVE: To better define the effect of individual risk factors and combinations thereof on the growth of small choroidal melanocytic tumors. DESIGN: Retrospective analysis. SETTING: Clinical practice of ocular oncology. PATIENTS: The study included 1287 patients with small suspicious choroidal melanocytic tumors, measuring 3 mm or less in thickness, managed with observation. RESULTS: On multivariate analysis, the clinical risk factors predictive of growth of small choroidal melanocytic tumors include tumor thickness greater than 2.0 mm, posterior tumor margin touching the disc, visual symptoms, orange pigment, and subretinal fluid. Tumor growth was detected in 4% of those patients with no risk factors. Growth was detected in approximately 36% of patients with 1 risk factor, 45% of patients with 2 risk factors, 50% of patients with 3 risk factors, 51% of patients with 4 risk factors, and 56% of patients with all 5 risk factors. The combination of risk factors offering the greatest risk for growth was tumor thickness greater than 2.0 mm, tumor margin touching disc, and subretinal fluid that was associated with tumor growth in 63% of the affected patients. The relative risk for growth was 1.9 for 1 factor, 3.8 for 2 factors, 7.4 for 3 factors, 14.1 for 4 factors, and 27.1 for all 5 risk factors combined. CONCLUSIONS: Five risk factors for growth of small choroidal melanocytic tumors have been identified. The combinations of various factors increase the risk for tumor growth from 4% if no factors are present to more than 50% if 3 or more risk factors are present. These factors may be important when counseling patients with small suspicious choroidal melanocytic tumors. PMID- 10721960 TI - Prognostic indicators for vision and mortality in shaken baby syndrome. AB - OBJECTIVE: To study ocular and nonocular signs of patients diagnosed as having "shaken baby syndrome" and determine prognostic indicators for vision and mortality. METHODS: Medical records of child abuse cases involving bilateral retinal hemorrhages were reviewed. Particular attention was paid to visual function and pupillary light reaction at the time of admission as well as the location of retinal hemorrhages, neuroimaging findings, ventilatory requirement, and associated skeletal injuries. These findings were correlated with visual prognosis and mortality. RESULTS: Thirty consecutive cases met the criteria for review. At the initial visit, mean age of the children was 9.3 months (range, 1 39 months) and 12 children (40%) had at least fix-and-follow vision. Preretinal and intraretinal hemorrhages (93% [n = 28] and 100% [n = 30]) were more common than vitreous hemorrhage (10% [n = 3]). Subdural hematomas were detected in 21 patients (70%). Twenty children (67%) had seizures and 16 (53%) required ventilatory support; bruises and long bone fractures were seen in 14 (47%) and 4 (13%) children, respectively. Eight patients died. All patients with nonreactive pupils on presentation died, while all patients with a pupillary light reaction lived (P<.001). Six (86%) of 7 patients with midline shift died, whereas 21 (91%) of 23 with no midline shift lived (P<.001). At follow-up, retinal hemorrhages had resolved in nearly all children by 4 months, and 16 children (73%) had at least fix-and-follow vision. Ventilatory requirement was associated with poorer vision (P<.01). CONCLUSIONS: Nonreactive pupils and midline shift of the brain structures correlate highly with mortality. Ventilatory requirement, but not visual acuity on presentation, predicts visual outcome. PMID- 10721961 TI - Loss of neurons in magnocellular and parvocellular layers of the lateral geniculate nucleus in glaucoma. AB - OBJECTIVES: To determine whether there is loss of lateral geniculate nucleus relay neurons, which convey visual information to the visual cortex, in experimental glaucoma in monkeys. METHOD: Four cynomolgus monkeys with experimentally induced glaucoma in the right eye (referred to as the glaucoma group) and 5 control monkeys were studied. In both groups, the same conditions of fixation, tissue processing, staining, and measurement were used. In each monkey, the left lateral geniculate nucleus target neurons in magnocellular layer 1 and parvocellular layers 4 and 6, connected to the right glaucomatous eye, were studied. Immunocytochemistry with antibody to parvalbumin was used to specifically label relay neurons connecting to the visual cortex. The number of parvalbumin-immunoreactive neurons was estimated using an unbiased 3-dimensional counting method. The t test was used to compare the experimental and control groups. RESULTS: The mean ( SD) number of neurons in mavnocellular layer 1 was significantly decreased in the glaucoma group compared with the control group (20 692 9567 vs 37 687 8017; P = .02). The mean ( SD) number of neurons in parvocellular layers 4 and 6 was significantly decreased in the glaucoma group compared with the control group (100 141 44 906 vs 174 090 39 136; P = .03). Data are given as the mean SD. CONCLUSION: Significant loss of lateral geniculate nucleus relay neurons terminating in the primary visual cortex occurs in the magnocellular and parvocellular layers in an experimental monkey model of glaucoma. CLINICAL RELEVANCE: Knowledge of the fate of neurons in the central visual system may lead to a better understanding of the nature and progression of visual loss in glaucomatous optic neuropathy. PMID- 10721962 TI - Light exposure and the risk of cortical, nuclear, and posterior subcapsular cataracts: the Pathologies Oculaires Liees a l'Age (POLA) study. AB - BACKGROUND: Exposure to light may be an important risk factor for the development of cataracts. OBJECTIVE: To present the relation of ambient solar radiation and professional and leisure exposures to light with the different types of cataracts. METHODS: Pathologies Oculaires Liees a l'Age (POLA) is a population based study on cataract and age-related macular degeneration and their risk factors in 2584 residents of Sete (southern France). Cataract classification was based on lens examination at slitlamp according to Lens Opacities Classification System III. A questionnaire about light exposure was administered. RESULTS: After multivariate adjustment, participants who had higher ambient solar radiation had a 2.5-fold (95% confidence interval [CI], 1.2-5.0), 4.0-fold (95% CI, 2.0-8.0), and 2.9-fold (95% CI, 1.5-5.3) increased risk of cortical and mixed cataract and cataract surgery, respectively. Solar ambient radiation was not significantly associated with posterior subcapsular and nuclear cataracts. By contrast, posterior subcapsular cataracts were significantly associated with professional exposure to sunlight (odds ratio [OR], 1.63; 95% CI, 1.01-2.63) and frequent use of sunglasses (OR, 0.62; 95% CI, 0.43-0.90). Mixed cataract was also associated with professional exposure to artificial light (OR, 3.02; 95% CI, 1.03-8.82). CONCLUSION: Our study further confirms the role of sunlight exposure in the pathogenesis of cataract, in particular in its cortical localization. PMID- 10721963 TI - Mixed lens opacities and subsequent mortality. AB - BACKGROUND: Previous studies have found an association between cataract or lens opacity and increased risk of mortality. Further work on determining explanatory factors for this association is needed. OBJECTIVES: To determine, in a population based cohort of older persons, the 2-year risk of death associated with different types of lens opacities; whether an association of mortality and lens opacity is explained by confounding risk factors such as smoking, diabetes, age, race, and sex, which are known to be related to opacity and mortality; whether lens opacity is a marker for health status; and whether there are differences in cause specific mortality for persons with and without lens opacity. MAIN OUTCOME MEASURE: Two-year mortality rate. METHODS: The Salisbury Eye Evaluation Project consists of a random sample of 2520 residents of Salisbury, Md, aged 65 to 84 years. At baseline, lens photographs were taken to document nuclear, cortical, posterior subcapsular cataract, and mixed opacities. Data on education, smoking, alcohol use, hypertension, diabetes and other comorbid conditions, handgrip strength, and body mass index were also collected. Two-year follow-up was conducted for mortality and cause of death. RESULTS: Nuclear opacity, particularly severe nuclear opacity, and mixed opacities with nuclear were significant predictors of mortality independent of body mass index, comorbid conditions, smoking, age, race, and sex (mixed nuclear: odds ratio, 2.23; 95% confidence interval, 1.26-3.95). CONCLUSION: Lens opacity status is an independent predictor of 2-year mortality, an association that could not be explained by potential confounders. PMID- 10721964 TI - Dietary fat and fish intake and age-related maculopathy. AB - OBJECTIVE: To assess whether dietary intake of fat or fish is associated with age related maculopathy (ARM) prevalence. DESIGN: Cross-sectional, urban population based study. PARTICIPANTS: People (N = 3654) aged 49 years or older. MAIN OUTCOME MEASURES: Subjects with ARM were identified from masked grading of retinal photographs. A 145-itemself-administered, semiquantitative food frequency questionnaire was completed adequately by 88.8% of participants and was used to assess intakes of dietary fat and fish. RESULTS: A higher frequency of fish consumption was associated with decreased odds of late ARM (odds ratio for frequency of consumption more than once per week compared with less than once per month, 0.5). Subjects with higher energy-adjusted intakes of cholesterol were significantly more likely to have late ARM, with an increased risk for late ARM for the highest compared with the lowest quintile of intake (odds ratio, 2.7). CONCLUSION: The amount and type of dietary fat intake may be associated with ARM. PMID- 10721965 TI - Perish, then publish: Thomas Harriot and the sine law of refraction. AB - A talented young scientist, Thomas Harriot, wrote the first English account of the New World, "A Briefe and True Report of the New Found Land of Virginia," distinguished by its serious effort to describe and understand the American Indian. Harriot went on to make innovations in mathematics and was one of the first astronomers to use the telescope. His largely unappreciated contribution to the history of ophthalmology was the first formulation of the sine law of refraction of light, found in his unpublished papers long after his death in 1621. Willebrord Snell discovered the sine law in Holland in 1621 but also died without formally publishing it. Rene Descartes first published the sine law in 1637. The sine law of refraction became not only the prime law of all lens systems but ushered in a new world of physical laws. PMID- 10721966 TI - A simple maneuver to reposit a subluxed globe. AB - In patients with thyroid orbitopathy, severe lid retraction and proptosis may produce spontaneous axial globe subluxation. This acute event is characterized by anterior displacement of the globe beyond the orbital rim, retraction of both upper and lower eyelids behind the globe, and tethering of the optic nerve. This frightening occurrence causes severe pain because of exposure keratopathy and forward displacement of the globe, as the retracted eyelids are squeezing the retrobulbar tissues. Spontaneous globe subluxation frequently occurs at home or at work, and patients experiencing this for the first time often panic because they have never been forewarned of this possibility nor received any instruction on how to attend to such an emergency. The lack of concise patient instruction for a quick and safe method of self-administered globe reposition is the main impetus for the design of current technique. PMID- 10721967 TI - A call for innovative operations for glaucoma. PMID- 10721968 TI - The triple procedure--is it the best approach for the patient? The triple procedure may be superior to sequential surgery. PMID- 10721969 TI - Sequential surgery may be the best approach for the patient. PMID- 10721970 TI - Traumatic Acremonium atrogriseum keratitis following laser-assisted in situ keratomileusis. AB - A 52-year-old man underwent bilateral laser-assisted in situ keratomileusis. Eight months later, he sustained a penetrating corneal injury to the left eye. A dense white infiltrate, unresponsive to antimicrobial therapy, developed in the corneal stroma. Corneal biopsy and eventual penetrating keratoplasty were performed, and both specimens demonstrated fungal elements with branching, septate hyphae. Culture identified the organism as Acremonium atrogriseum. Histopathologic features of this organism and its differentiation from other, more common fungal organisms are discussed herein. PMID- 10721971 TI - Well-differentiated squamous cell carcinoma of the eyelid arising during a 20 year period. AB - An 81-year-old man had a keratotic eyelid lesions for 20 years. He eventually sought treatment by ophthalmic plastic surgery. Clinically, the lesion resembled a keratoacanthoma. Findings from histologic examination of the excision biopsy specimen showed a squamous cell carcinoma. The lesion was completely excised. This case demonstrates the difficulty in making a correct clinical diagnosis of a keratotic eyelid lesion. Performing a histologic examination of nonregressed keratotic lesions is essential to exclude a squamous cell carcinoma. PMID- 10721973 TI - Retinal toxic effects associated with intravitreal fomivirsen. PMID- 10721974 TI - Reversible atorvastatin-associated external ophthalmoplegia, anti-acetylcholine receptor antibodies, and ataxia. PMID- 10721972 TI - Lipemia retinalis in acquired immunodeficiency syndrome treated with protease inhibitors. PMID- 10721975 TI - Familial occurrence of ablepharon macrostomia syndrome: eyelid structure and surgical considerations. PMID- 10721976 TI - Use of apically based periosteal flaps as globe tethers in severe paretic strabismus. AB - OBJECTIVE: To evaluate the technique of using an intact autogenous periosteal flap for tethering of the globe in patients with severe paretic strabismus. METHODS: We performed a periosteal flap procedure on 5 patients and followed their postoperative course. The flap was created from the medial, lateral, or superior orbital walls. A description of the harvesting and manipulation of the flap and the initial postoperative findings are presented. RESULTS: All patients showed marked reduction in their postoperative strabismic deviation compared with preoperative measurements. Greater early postoperative swelling was noted after this procedure than with the standard strabismus surgery. No complications were experienced during or after surgery. Two patients required a second operation for adjustment of the periosteal flap for adequate alignment. CONCLUSIONS: The vascularized periosteal flap technique provides an excellent tether for the globe. Early and late stability has been favorable. PMID- 10721977 TI - A macular lesion simulating an aberrant cryotherapy lesion in retinopathy of prematurity. PMID- 10721978 TI - Cholesterosis following chemoreduction for advanced retinoblastoma. PMID- 10721979 TI - Diffuse infiltrating retinoblastoma simulating uveitis in a 7-year-old boy. PMID- 10721980 TI - Analysis of costs. PMID- 10721981 TI - Coinjection of hyaluronic acid and hyaluronidase. PMID- 10721982 TI - Efficacy of hyaluronidase. PMID- 10721983 TI - Improving the diagnostic yield of vitrectomy for intraocular lymphoma. PMID- 10721984 TI - Eponym error. PMID- 10721985 TI - DC-SIGN: a guide to some mysteries of dendritic cells. PMID- 10721986 TI - Wnts as retrograde signals for axon and growth cone differentiation. PMID- 10721987 TI - Looping, linking, and chromatin activity: new insights into beta-globin locus regulation. PMID- 10721988 TI - Secondary structure of vertebrate telomerase RNA. AB - Telomerase is a ribonucleoprotein enzyme that maintains telomere length by adding telomeric sequence repeats onto chromosome ends. The essential RNA component of telomerase provides the template for telomeric repeat synthesis. To determine the secondary structure of vertebrate telomerase RNA, 32 new telomerase RNA genes were cloned and sequenced from a variety of vertebrate species including 18 mammals, 2 birds, 1 reptile, 7 amphibians, and 4 fishes. Using phylogenetic comparative analysis, we propose a secondary structure that contains four structural domains conserved in all vertebrates. Ten helical regions of the RNA are universally conserved while other regions vary significantly in length and sequence between different classes of vertebrates. The proposed vertebrate telomerase RNA structure displays a strikingly similar topology to the previously determined ciliate telomerase RNA structure, implying an evolutionary conservation of the global architecture of telomerase RNA. PMID- 10721989 TI - Coupled leading- and lagging-strand synthesis of mammalian mitochondrial DNA. AB - Analysis of mammalian mtDNA by two-dimensional agarose gel electrophoresis revealed two classes of replication intermediate. One was resistant to single strand nuclease digestion and displayed the mobility properties of coupled leading- and lagging- strand replication products. Intermediates of coupled, unidirectional mtDNA replication were found in mouse liver and human placenta and were the predominant species in cultured cells recovering from transient mtDNA replication. Replication intermediates sensitive to single-strand nuclease were most abundant in untreated cultured cells. These are presumed to derive from the orthodox, strand-asynchronous mode of mtDNA replication. These findings indicate that two modes of mtDNA replication operate in mammalian cells and that changes in mtDNA copy number involve an alteration in the mode of mtDNA replication. PMID- 10721990 TI - Axonal remodeling and synaptic differentiation in the cerebellum is regulated by WNT-7a signaling. AB - Synapse formation requires changes in cell morphology and the upregulation and localization of synaptic proteins. In the cerebellum, mossy fibers undergo extensive remodeling as they contact several granule cells and form complex, multisynaptic glomerular rosettes. Here we show that granule cells secrete factors that induce axon and growth cone remodeling in mossy fibers. This effect is blocked by the WNT antagonist, sFRP-1, and mimicked by WNT-7a, which is expressed by granule cells. WNT-7a also induces synapsin I clustering at remodeled areas of mossy fibers, a preliminary step in synaptogenesis. Wnt-7a mutant mice show a delay in the morphological maturation of glomerular rosettes and in the accumulation of synapsin I. We propose that WNT-7a can function as a synaptogenic factor. PMID- 10721991 TI - Solution structure of the E. coli 70S ribosome at 11.5 A resolution. AB - Over 73,000 projections of the E. coli ribosome bound with formyl-methionyl initiator tRNAf(Met) were used to obtain an 11.5 A cryo-electron microscopy map of the complex. This map allows identification of RNA helices, peripheral proteins, and intersubunit bridges. Comparison of double-stranded RNA regions and positions of proteins identified in both cryo-EM and X-ray maps indicates good overall agreement but points to rearrangements of ribosomal components required for the subunit association. Fitting of known components of the 50S stalk base region into the map defines the architecture of the GTPase-associated center and reveals a major change in the orientation of the alpha-sarcin-ricin loop. Analysis of the bridging connections between the subunits provides insight into the dynamic signaling mechanism between the ribosomal subunits. PMID- 10721992 TI - Structural basis of presequence recognition by the mitochondrial protein import receptor Tom20. AB - Most mitochondrial proteins are synthesized in the cytosol as precursor proteins with a cleavable N-terminal presequence and are imported into mitochondria. We report here the NMR structure of a general import receptor, rat Tom20, in a complex with a presequence peptide derived from rat aldehyde dehydrogenase. The cytosolic domain of Tom20 forms an all alpha-helical structure with a groove to accommodate the presequence peptide. The bound presequence forms an amphiphilic helical structure with hydrophobic leucines aligned on one side to interact with a hydrophobic patch in the Tom20 groove. Although the positive charges of the presequence are essential for import ability, presequence binding to Tom20 is mediated mainly by hydrophobic rather than ionic interactions. PMID- 10721993 TI - Multivalent binding of nonnative substrate proteins by the chaperonin GroEL. AB - The chaperonin GroEL binds nonnative substrate protein in the central cavity of an open ring through exposed hydrophobic residues at the inside aspect of the apical domains and then mediates productive folding upon binding ATP and the cochaperonin GroES. Whether nonnative proteins bind to more than one of the seven apical domains of a GroEL ring is unknown. We have addressed this using rings with various combinations of wild-type and binding-defective mutant apical domains, enabled by their production as single polypeptides. A wild-type extent of binary complex formation with two stringent substrate proteins, malate dehydrogenase or Rubisco, required a minimum of three consecutive binding proficient apical domains. Rhodanese, a less-stringent substrate, required only two wild-type domains and was insensitive to their arrangement. As a physical correlate, multivalent binding of Rubisco was directly observed in an oxidative cross-linking experiment. PMID- 10721996 TI - The evolving role of combined modality therapy in head and neck cancer. PMID- 10721994 TI - Identification of DC-SIGN, a novel dendritic cell-specific ICAM-3 receptor that supports primary immune responses. AB - Contact between dendritic cells (DC) and resting T cells is essential to initiate a primary immune response. Here, we demonstrate that ICAM-3 expressed by resting T cells is important in this first contact with DC. We discovered that instead of the common ICAM-3 receptors LFA-1 and alphaDbeta2, a novel DC-specific C-type lectin, DC-SIGN, binds ICAM-3 with high affinity. DC-SIGN, which is abundantly expressed by DC both in vitro and in vivo, mediates transient adhesion with T cells. Since antibodies against DC-SIGN inhibit DC-induced proliferation of resting T cells, our findings predict that DC-SIGN enables T cell receptor engagement by stabilization of the DC-T cell contact zone. PMID- 10721995 TI - DC-SIGN, a dendritic cell-specific HIV-1-binding protein that enhances trans infection of T cells. AB - Dendritic cells (DC) capture microorganisms that enter peripheral mucosal tissues and then migrate to secondary lymphoid organs, where they present these in antigenic form to resting T cells and thus initiate adaptive immune responses. Here, we describe the properties of a DC-specific C-type lectin, DC-SIGN, that is highly expressed on DC present in mucosal tissues and binds to the HIV-1 envelope glycoprotein gp120. DC-SIGN does not function as a receptor for viral entry into DC but instead promotes efficient infection in trans of cells that express CD4 and chemokine receptors. We propose that DC-SIGN efficiently captures HIV-1 in the periphery and facilitates its transport to secondary lymphoid organs rich in T cells, to enhance infection in trans of these target cells. PMID- 10721997 TI - Birth of a new society: American Head and Neck Society challenges for the future. PMID- 10721998 TI - What is the role of primary surgery in the treatment of laryngeal and hypopharyngeal cancer? Hayes Martin Lecture. PMID- 10721999 TI - Who will care for the uninsured in a managed care world? John Conley Lecture. PMID- 10722000 TI - Reconstruction of the pediatric maxilla and mandible. AB - BACKGROUND: The creation of osseous defects in the upper and lower jaws in children is an uncommon occurrence. It is therefore likely that a head and neck reconstructive surgeon will accumulate only limited experience in restoring such defects. We have reviewed 7 pediatric bone-containing microvascular free flap reconstructions in 6 patients for reconstruction of the upper or lower jaws. Three patients were available for long-term follow-up to evaluate the effect of osseous free flap reconstruction on function and growth and development of the donor site. DESIGN: Retrospective review. SETTING: Academic tertiary referral center for otolaryngology. PATIENTS AND METHODS: Six pediatric patients ranging in age from 8 to 16 years underwent 2 fibular, 4 scapular, and 1 iliac free flap procedure for restoration of 2 maxillary and 5 mandibular defects from 1992 to 1997. Three of the 6 patients were available for long-term follow-up to assess the postoperative donor site function in an effort to determine the effect of this surgery on long-term donor site morbidity and development. RESULTS: Two patients were lost to follow-up, and 1 died secondary to complications related to distant metastatic disease. Three of 6 patients were observed for 2 years 6 months, 4 years, and 4 years 2 months, respectively. Two of the 3 patients who were observed long term have undergone full dental rehabilitation and currently maintain a regular diet and deny pain with mastication or deglutition. One patient did not require dental rehabilitation. All 3 patients demonstrate gross facial symmetry and normal dental occlusion. Assessment of the fibular donor site demonstrated normal limb length and circumference. The patients denied pain or restriction to recreational activity. Scapular donor sites demonstrated normal range of motion, strength, and shoulder stability. CONCLUSIONS: Free flap reconstruction of the pediatric maxilla and mandible requires harvesting bone from actively growing donor sites. We have found no evidence of functional deficit after bone harvest from the fibular or scapular donor sites. Patients demonstrate normal growth at the donor sites, and symmetry of the mandible and maxilla is preserved. PMID- 10722001 TI - Posterior scalping flap revisited. AB - OBJECTIVE: To report our experience in a case series of 5 posterior scalping flaps. DESIGN: Retrospective review of a case series. SETTING: A tertiary academic care otolaryngology-head and neck surgery referral center. PATIENTS: Five patients having undergone posterior scalping flap reconstruction of cutaneous midface defects. METHODS: Reconstruction was performed for 4 cheek defects, 1 of which included the lateral third of the upper and lower lips, and 1 combined midfacial and lateral nasal wall defect. RESULTS: All 5 patients had excellent cosmetic and functional results. The only complication was a single case of partial-thickness distal flap necrosis. CONCLUSION: The posterior scalping flap offers a reliable source of skin with appropriate color and texture and minimal donor-site morbidity. PMID- 10722002 TI - Prognostic importance of vascular invasion in papillary thyroid carcinoma. AB - BACKGROUND: The prognostic importance of vascular invasion has not been extensively studied in patients with papillary thyroid cancer. OBJECTIVE: To determine whether the presence of vascular invasion in papillary thyroid carcinoma, even within the thyroid gland, is associated with more aggressive disease at diagnosis and a higher incidence of tumor recurrence. PATIENTS AND METHODS: We identified 410 patients who had been diagnosed with papillary thyroid cancer since 1986 who had a follow-up period of longer than 1 year (median follow up, 5.5 years). Pathology reports were reviewed and patients were separated into 3 groups: no vascular invasion, intrathyroidal vascular invasion, and extrathyroidal vascular invasion. MAIN OUTCOME MEASURES: Statistical comparison was performed by univariate and multivariate analysis. RESULTS: Patients with intrathyroidal vascular invasion were more likely to have distant metastasis at the time of diagnosis (26.1% vs 2.2%, P = .001). Similarly, patients with extrathyroidal vascular invasion had a higher incidence of distant metastases at diagnosis (40% vs 4.4%, P = .02). Patients with tumors identified to have intrathyroidal vascular invasion were more likely to develop distant recurrence (20% vs 3%, P = .002). CONCLUSIONS: These associations were found to be independent by multiple regression analysis. Patient age, sex, palpable or fixed lymph nodes, radiation exposure, and race did not differ between the patient group with and those without vascular invasion. Preliminary analysis of our data suggests that the presence of vascular invasion in papillary, thyroid carcinoma, even within the thyroid gland, is associated with more aggressive disease at diagnosis and with a higher incidence of tumor recurrence. PMID- 10722003 TI - Cervical sentinel lymph node biopsy for melanomas of the head and neck and upper thorax. AB - OBJECTIVE: To describe a clinical experience with sentinel lymph node biopsy (SLNB) of head and neck nodal basins for clinical stage I melanomas draining to these areas. DESIGN: Consecutive clinical case series with a mean follow-up of 10.7 months. SETTING: University tertiary care referral medical center. PATIENTS: Seventy patients with clinical stage I cutaneous melanoma who underwent SLNB of cervical and/or parotid lymph node basins. INTERVENTIONS: Patients underwent same day preoperative technetium Tc 99m lymphoscintigraphy followed by SLNB using gamma probe and blue dye (66 patients) and blue dye alone (4 patients). Patients with histological evidence of tumor (here in after "positive") according to SLNB results underwent modified cervical completion lymph node dissection, including parotidectomy, as appropriate. Patients without histological evidence of tumor (hereinafter "negative") according to SLNB results were followed up clinically without undergoing completion lymph node dissection. MAIN OUTCOME MEASURES: The rates of SLNB success, SLNB positivity, completion lymph node dissection positivity, complications, and SLNB false-negative results were determined by clinical follow-up. RESULTS: Locations of melanomas in the 70 patients were the face (n = 20), neck (n = 14), ear (n = 9), scalp (n = 9), and upper thorax (n = 18). Locations of basins that underwent biopsy (n = 104) were in the cervical (n = 68), parotid (n = 19), and axillary (n = 17) regions. The mean Breslow thickness was 2.1 mm (range, 0.4-12.0 mm). Sentinel lymph node biopsy was successful in 103 basins (99%). The mean number of sentinel lymph nodes per basin was 2.5 (range, 1.0-8.0). Positive sentinel lymph nodes were found in 12 patients (17%) and 15 basins (14%). Sentinel lymph node biopsy results correlated with the American Joint Committee on Cancer tumor stage (P = .05) and a Breslow thickness of 1.23 mm or greater (P = .03). Additional tumor-containing nodes were noted in 5 (42%) of the 12 patients who underwent completion lymph node dissection, and these results correlated with the presence of multiple positive sentinel lymph nodes (P = .01). There were complications in 3 patients (4%) (seromas in 2 patients and temporary spinal accessory nerve paresis in 1 patient). One nodal recurrence in a basin that was negative according to SLNB results (SLNB with blue dye only) was noted (false-negative rate, 2%). The results of SLNB were accurate in 69 patients (99%). CONCLUSIONS: Sentinel lymph node biopsy using lymphoscintigraphy and blue dye to manage cutaneous melanomas draining to the head and neck nodal areas is reliable and safe. Sentinel lymph node biopsy results correlated with a Breslow thickness of 1.23 mm or greater and the American Joint Committee on Cancer tumor stage. Completion lymph node dissection is recommended after determining positive SLNB results. PMID- 10722004 TI - The impact of clinical pathways on the practice of head and neck oncologic surgery: the University of Texas M. D. Anderson Cancer Center Experience. AB - OBJECTIVE: To assess the impact of clinical pathways on the practice of head and neck oncologic surgery in an academic center. DESIGN: Cross-sectional study. SETTING: Cancer treatment center. PATIENTS: The study population consisted of 3 groups of patients who underwent unilateral neck dissection and were treated in the Department of Head and Neck Surgery of the University of Texas M. D. Anderson Cancer Center, Houston. Additional procedures which may have been performed were direct laryngoscopy, rigid esophagoscopy, and/or dental extractions. Ninety-six patients treated during 1993-1994 prior to the implementation of the clinical pathway (historical control group) were compared with 94 patients treated during 1996-1998, 64 who were not (contemporaneous nonpathway group) and 30 who were managed on the clinical pathway (pathway group). Patients from 1995 were excluded since the pathway was in the planning stages then. MAIN OUTCOME MEASURES: Median length of stay; median total costs of care. RESULTS: The median length of hospital stay of the historical control, contemporaneous nonpathway, and pathway groups decreased from 4.0 to 2.0 days (P<.001). The total median costs of care were less in the pathway group as compared with the historical control group ($6,227 and $8,459, respectively, P<.001) and also less in the contemporaneous nonpathway group compared with the historical control group (S6885 and $8,459, respectively, P<.001). Mean and median length of hospital stay and costs were lower in the pathway group as compared with the nonpathway group but not significantly (P = .11 and P = .07, respectively) The contemporaneous nonpathway and pathway groups did not differ in complications or readmissions. CONCLUSIONS: Development and implementation of this clinical pathway played a statistically significant role in decreasing length of hospital stay and total costs of care associated with neck dissection between nonpathway and pathway patients. Thus, a more cost-effective practice environment has resulted for all of our patients. PMID- 10722005 TI - Quality of life in patients with head and neck cancer: lessons learned from 549 prospectively evaluated patients. AB - OBJECTIVES: To summarize our quality-of-life (QOL) research findings for patients with head and neck cancer, to suggest areas for future productive QOL research, and to discuss how to undertake QOL studies in a cost-effective manner. DESIGN: Review of previously published analyses of advanced larynx cancer, advanced oropharynx cancer, and neck-dissection cases and current data from the complete set of patients. PATIENTS: From January 1, 1993, through December 31, 1998, data on 549 patients were entered in our head and neck database. Of these patients, 364 met additional criteria for histologic findings (squamous cell carcinoma) and the restriction of their cancer to 4 major anatomical sites (oral, oropharynx, hypopharynx, or larynx). Of these, 339 patients were more than 1 year beyond initial treatment. Complete baseline TNM staging and QOL data were obtained for 260 of these patients, of whom 210 presented with an untreated first primary tumor (index cases) to the University of Washington, Seattle. INTERVENTION: Pretreatment QOL was assessed with an interviewer-supervised self-administered questionnaire. Subsequent self-administered tests were completed at 3, 6, 12, 24, and 36 months. Other data collected on each patient included cancer site, stage, treatment, histologic findings, type of surgical reconstruction, and current disease and vital status. RESULTS/CONCLUSIONS: It is difficult to achieve "statistically significant" results in a single-institution setting. The "composite" QOL score may not be a sufficiently sensitive tool. Analysis of separate domains may be more effective. PMID- 10722006 TI - Preoperative chemotherapy-sensitized radiation therapy for cervical metastases in head and neck cancer. AB - OBJECTIVE: To determine the efficacy of concurrent preoperative cisplatin chemotherapy and radiotherapy (CT/RT) for patients with advanced head and neck cancer and cervical metastatic disease. DESIGN: Retrospective analysis. SETTING: University hospitals. PATIENTS: Eighty-eight patients with operable stage III and IV squamous cell carcinoma of the head and neck and palpable cervical lymphogenous metastases received preoperative concurrent CT/RT followed by planned neck dissection. INTERVENTIONS: All patients undergoing CT/RT received concomitant continuous infusions of cisplatin (20 mg/m2) on days 1 to 4 and 22 to 25 of CT/RT. Thirty-nine patients underwent single-fraction (1.8-Gy) radiotherapy to 45.0 Gy, and 49 patients received 10 single-fraction (1.8-Gy) treatments, which were hyperfractionated (1.2-Gy twice a day) to 46.8 Gy. MAIN OUTCOME MEASURES: The 71 patients for whom complete post-CT/RT data were available were evaluated for clinical response in addition to survival. Histologic complete response (HCR) was confirmed from planned neck dissection specimens (n = 48) after clinical complete response (CCR) from initial CT/RT. Kaplan-Meier statistical analysis for disease-specific survival and overall survival was performed on all 88 patients who received CT/RT. RESULTS: A CCR and an HCR were noted in 78% (18/23) and 59% (10/17) of patients with N1 lesions, respectively, and in 60% (29/48) and 45% (14/31) of patients with N2-3 lesions, respectively. The percentage of patients with CCR who also had HCR was 67% (10/15) for patients with N1 lesions and 54% (14/26) for patients with N2-3 lesions. With a median follow-up of 18.5 months, the Kaplan-Meier disease-specific survival rate at 54 months (n = 88) was 70% (21/30) for patients with N1 lesions, 60% (24/40) for patients with N2 lesions, and 39% (7/18) for patients with N3 lesions. The overall survival and disease-specific survival rates at 5 years for all nodal groups combined were 36% (32/88) and 59% (52/88), respectively. CONCLUSIONS: A CCR to CT/RT was achieved in nearly two thirds of patients with head and neck cervical lymphogenous metastases, independent of nodal tumor load. Most patients (59% [24/41]) with CCR were pathologically tumor free before neck dissection. PMID- 10722007 TI - Combination immunotherapy of squamous cell carcinoma of the head and neck: a phase 2 trial. AB - OBJECTIVES: To test the efficacy of a natural cytokine mixture (IRX-2), cyclophosphamide, indomethacin, and zinc to induce immune regression of squamous cell carcinoma (SCC) of the head and neck (H&N) prior to conventional therapy and to characterize the responses. PATIENTS AND DESIGN: A phase 2 trial was performed in 15 adults with recently diagnosed, biopsy-confirmed H&N SCC (3 with stage II disease, 6 with stage III disease, and 6 with stage IV disease). The patients were treated with 20 days of perilymphatic injections of IRX-2 (administered subcutaneously at the base of the skull) in combination with contrasuppression consisting of a low-dose infusion of cyclophosphamide (300 mg/m2), and daily oral indomethacin and zinc (StressTabs) in a 21-day cycle before surgery and/or radiotherapy. Tumor dimensions, toxic effects, and disease-free survival were monitored. The tumor sections were histologically examined after surgery, and tumor reduction, fragmentation, and lymphoid infiltration were assessed. RESULTS: All 15 patients responded clinically to the 21-day IRX-2 protocol: 1 with a complete response, 7 with a partial response, and 7 with a minor response. All 15 patients responded pathologically with tumor reduction (mean, 42%) and fragmentation (mean, 50%) in the histological section and increased lymphoid infiltration. The adverse effects of the IRX-2 protocol were negligible except for an allergic skin rash (n = 1) and parotiditis (n = 1). Indomethacin caused gastritis in 1 patient. Reduction of pain and ulceration and bleeding were observed in 8 and 4 patients, respectively. Four of 5 patients with lymphopenia showed increased CD3, CD4, and CD8 cell counts. After surgery (n = 13) and/or radiotherapy (n = 10) and with a mean follow-up of 17 months, 3 patients have had recurrences, 1 patient has died of disease, 1 patient has been re-treated with immunotherapy and has no evidence of disease, and 1 patient is alive with disease. Two patients died of other causes with no evidence of disease. CONCLUSIONS: The IRX-2 immunotherapy induced lymphocyte mobilization and infiltration in H&N SCC associated with clinical and histological tumor responses indicative of immune regression in all 15 patients. Minimal toxic effects were observed, and overall survival may have been improved. A phase 3 trial seems warranted. PMID- 10722008 TI - The cervical wound of General James Longstreet. AB - BACKGROUND: Lieutenant General James Longstreet was arguably the finest corps commander on either side during the Civil War. He was severely wounded at the Battle of the Wilderness in Virginia on May 6, 1864, after a successful flank attack that nearly routed the Union army. DESIGN: A thorough review of the firsthand accounts of the events leading up to and following Longstreet's wounding was made. In addition, all articles listed in the medical literature describing Longstreet's care and numerous recent texts and articles about Longstreet have been researched. RESULTS: After being wounded on May 6, Longstreet received appropriate care by John Syng Dorsey Cullen, MD. Cullen controlled the hemorrhage from Longstreet's wound, helped evacuate him from the battlefield, and diligently cared for him during his convalescence. CONCLUSIONS: Longstreet was wounded by "friendly fire." The bullet's trajectory and the location of the gunshot wound suggest a posterior wound of entry rather than an anterior one as has been previously assumed. PMID- 10722010 TI - Computed tomography--guided needle biopsy of head and neck lesions. AB - OBJECTIVE: To evaluate the diagnostic efficacy of computed tomography (CT)-guided needle biopsies of head and neck lesions. DESIGN: All CT-guided needle biopsies of head and neck lesions performed between September 1994 and February 1999 were included. Cytopathologic and histologic records, along with patient clinical records, were reviewed. SETTING: A tertiary care medical center. PATIENTS: Patients referred for evaluation of lesions inaccessible to routine methods of needle biopsy. RESULTS: Thirty-seven patients underwent 42 CT-guided biopsies. There were included 12 lesions in or adjacent to the skull base and 9 lesions around the pharyngoesophageal or laryngotracheal complex; the other lesions were located in the deep lobe of the parotid gland (n = 7), deep neck area (n = 12), and thyroid gland (n = 2). Diagnostic cytologic biopsy specimens were obtained in 38 (91%) of 42 needle biopsy procedures. The results were supported histologically and/or clinically in 36 cases (95%). Eighteen patients underwent open surgical procedures. Histologic confirmation was found in 86% of cases. Nineteen patients (51%) avoided an open surgical procedure: 11 with benign disease and 8 with recurrent malignancy. There were no false-positive or false negative results, and no complications were identified. CONCLUSIONS: Computed tomography-guided needle biopsy is a safe and reliable minimally invasive technique for the diagnosis of poorly accessible or deep-seated lesions of the head and neck. Diagnostic needle biopsies allow improved preoperative planning and patient counseling in surgical patients and avoidance of open surgical procedures in patients with benign disease or recurrent malignant neoplasms. PMID- 10722009 TI - The use of clinical criteria alone in the management of the clinically negative neck among patients with squamous cell carcinoma of the oral cavity and oropharynx. AB - BACKGROUND: Management of the clinically negative neck among patients with oral and oropharyngeal squamous cell carcinoma at the Royal Prince Alfred Hospital, Sydney, Australia has been based on the site and stage of the primary cancer, the likely incidence of microscopic nodal involvement, the treatment modality used for the primary cancer, and whether the neck will be entered during resection or reconstruction. This report analyzes the results of treatment when patients are allocated to either treatment or observation of the neck based on these clinical factors. METHODS: This is a prospectively documented series of 162 consecutively treated patients with squamous cell carcinoma of the oral cavity and oropharynx and clinically negative necks, treated by 1 surgeon (C.J.O.). There were 128 oral cavity and 34 oropharyngeal cancers clinically staged at T1 for 62 patients, T2 for 61, T3 for 16, and T4 for 23 patients. Management of the neck consisted of elective neck dissection (END) in 96 patients (12 bilateral), elective radiotherapy in 8, and observation in 58. Neck treatment correlated with the T stage in a statistically significant way. Forty-six patients underwent postoperative radiotherapy, which was directed to the neck in 22 patients because of pathological findings following neck dissection. Free-flap reconstruction was used in 90 patients. RESULTS: Metastatic squamous cell carcinoma was identified in 32 of 108 neck dissections (30%). There was 1 positive node in 15 necks, 2 positive nodes in 11 necks, and 3 or more positive nodes in 6 necks. Extracapsular spread was present in 8 of 32 positive END specimens (25%). Regional control rates in the neck at 3 years were 94% for END, 100% for elective radiotherapy, and 98% for patients initially observed and then treated by therapeutic neck dissection. Death with uncontrolled disease in the neck occurred in 4 of 96 patients (4%) after END and 1 of 58 patients (2%) after neck observation. Overall disease-specific survival was 83%, comprising an 86% rate for patients with pathologically negative necks and 68% if pathologically positive. Disease-specific survival was 86% at 3 years for patients having END, 67% following radiotherapy, and 94% for the observation group. CONCLUSIONS: Elective neck dissection was performed in most patients, and occult metastatic disease was found in nearly 30% of neck dissections. Observation was most frequently used for patients with early stage disease, and subsequent development of neck metastases was uncommon (9%) in this group. Selective treatment of the clinically negative neck based on the primary tumor site and stage led to a high rate of regional disease control in this series. PMID- 10722011 TI - Swallowing function in patients with head and neck cancer prior to treatment. AB - OBJECTIVE: To define the site-specific swallowing dysfunctions of patients with head and neck cancer with respect to tumor site and stage by, videofluoroscopic oropharyngeal motility (OPM) study prior to initiation of treatment. DESIGN: Retrospective survey. SETTING: Academic university institution. PATIENTS: A consecutive sample of 79 patients with stage III or IV head and neck cancer without prior treatment or tracheotomy. Patients were divided into groups according to tumor site: oral cavity (n = 7), oropharynx (n = 27), larynx (n = 24), and hypopharynx (n = 10). Patients with sinonasal, nasopharyngeal, and unknown primary carcinomas served as the comparison group (n = 11). INTERVENTION: All patients underwent OPM study prior to treatment. MAIN OUTCOME MEASURES: Parameters of swallowing function, including oral impairment, pharyngeal impairment, cervical esophageal impairment, aspiration, and Swallowing Performance Status Scale (SPSS) score (a global measure of swallowing function) were extracted from the pretreatment OPM study and analyzed with reference to tumor site, T stage, and overall stage. The relations between tumor site and area or degree of dysfunction, and between stage of disease and area or degree of dysfunction were analyzed using chi2 and Fisher exact tests. RESULTS: Aspiration status, cervical esophageal impairment, and pharyngeal impairment examined as a function of disease site showed statistically significant differences between groups, with laryngeal and hypopharyngeal sites revealing the most severe dysfunctions. The SPSS score did not correlate with tumor site, T stage, or overall stage. Other OPM parameters analyzed as a function of T stage and overall stage revealed no consistent patterns. CONCLUSIONS: Hypopharyngeal and laryngeal disease sites have a high degree of pretreatment functional impairment. The SPSS score is a good global measure of swallowing dysfunction. In addition, significant site-specific dysfunctions are found when the OPM study is analyzed via its separate parameters. It is therefore critical that posttreatment function is compared with baseline pretreatment dysfunction. PMID- 10722012 TI - Video fluoroscopic evaluation after glossectomy. AB - BACKGROUND: The swallowing deficits that result from oral or oropharyngeal resections vary considerably depending on the site, extension of the resection, and type of reconstruction. Most patients will experience some degree of dysphagia despite the reconstructive effort. Furthermore, a glossectomy is frequently associated with voice and speech difficulties. OBJECTIVES: To characterize swallowing in patients who underwent a glossectomy and to define the limits and the compensatory movements using video fluoroscopic analysis. DESIGN AND SETTING: Video fluoroscopic evaluation of 15 patients who underwent glossectomies at the Centro de Tratamento e Pesquisa Hospital do Cancer A. C. Camargo, S*ao Paulo, Brazil. PATIENTS: We examined 15 patients: 5 who underwent a partial glossectomy, 2 who underwent a subtotal glossectomy, and 8 who underwent a total glossectomy with laryngeal preservation and reconstruction with myocutaneous flaps (9 pectoralis major flaps and 1 latissimus dorsi flap). The 15 patients were enrolled in a program that included voice, speech, and swallowing rehabilitation. RESULTS: All patients who underwent a partial glossectomy had difficulties with formation and anteroposterior propulsion of the bolus in the oral cavity and an increase in oral transit time, which was more evident with materials of thicker consistencies. All patients who underwent a total or subtotal glossectomy with laryngeal preservation had an increase in oral transit time and stasis of food in the oral cavity, the pharynx, and the superior esophageal sphincter. Of the 15 patients, 2 had moderate and asymptomatic aspiration. These 2 patients had swallowing compensations, such as increased buccal, mandibular, pharyngeal, and laryngeal activity and voluntary protection of the larynx during swallowing. CONCLUSIONS: This study demonstrates the effectiveness of swallowing in patients who were enrolled in voice, speech, and swallowing rehabilitation after undergoing a partial or total glossectomy. An increase in oral transit time was detected in all patients. Only 2 of the 10 patients who underwent a total glossectomy had persistent asymptomatic aspiration. PMID- 10722013 TI - Long-term swallowing problems after organ preservation therapy with concomitant radiation therapy and intravenous hydroxyurea: initial results. AB - OBJECTIVE: To evaluate the long-term effects on swallowing function of concomitant continuous infusion hydroxyurea and hyperfractionated radiation therapy used to treat advanced head and neck carcinoma. DESIGN: A prospective evaluation of swallowing function was performed on an inception cohort by analyzing posttreatment videoflouroscopic swallow function studies using radiological descriptors for pharyngeal transport abnormalities and temporal measures of structural movements, as well as by conducting patient interviews to assess alimentation, more than 1 year after tumor treatment (range, 52-124 weeks; median, 70 weeks). SETTING: Academic tertiary care referral medical center. PATIENTS: Ten patients, aged 44 to 71 years, with stage III and IV squamous cell carcinoma of the oral cavity, oropharynx, or hypopharynx. MAIN OUTCOME MEASURE: Radiographic and temporal swallow abnormalities, as well as functional status, were documented and compared with published norms and results of earlier swallowing studies when possible. RESULTS: Pharyngeal transport dysfunction and anterior segment abnormalities, manifested by epiglottic dysmotility, vallecular residue, laryngeal penetration, or aspiration, were evident in all 10 patients. Posterior segment abnormalities, such as pharyngeal stasis, constrictor dysmotility and piriform residue were documented in 8 patients. Three patients developed late aspiration, and the majority of patients showed persistent or worsened delay in laryngeal movement compared with their earlier posttreatment evaluations. Also, 3 patients developed a hypopharyngeal stricture, and 6 patients continued to require gastrostomy tube supplementation beyond 1 year. There was no association between site of primary, duration to swallowing evaluation, and severity of dysfunction. CONCLUSION: Prolonged and debilitating functional swallowing abnormalities may occur after this aggressive concomitant chemotherapy and radiotherapy regimen. PMID- 10722014 TI - Experience with various 3-dimensional navigation systems in head and neck surgery. AB - OBJECTIVE: To evaluate the benefits and difficulties encountered when using various 3-dimensional (3-D) navigation systems in head and neck procedures. DESIGN: Five different navigation systems were used for preoperative planning and intraoperative 3-D navigation in procedures at the paranasal sinuses, the frontal and lateral skull bases, and the petrous bone. INTERVENTION: Intraoperative 3-D localizing systems (position-sensitive mechanical arms, infrared cameras, etc) demand reliable patient fixation on the operating table. We achieved this by developing a noninvasive head holder. Other systems allow patient movements by using magnetic digitizing technology (ARTMA System) and sophisticated programming. RESULT: Having surpassed an initial learning curve, we now achieve an accuracy of 1 to 2 mm regularly. Especially in paranasal and frontal basal surgery, all navigation systems used provide valuable positioning information during surgery. In particular for revision or tumor surgery, decisive benefits resulted from use of these systems: shorter overall operation time; safer manipulation near delicate structures; and reliable identification of the skull base even in patients with bleeding, scarring, or missing anatomical landmarks. CONCLUSIONS: We performed approximately 250 operations with different systems and introduced navigation at the lateral skull base and the petrous bone with mechanical, optic, and magnetic digitizers. In these anatomical areas, navigation was used successfully; the technical challenge is greatest at the lateral skull base, however. PMID- 10722015 TI - New reconstructive technologies in skull base surgery: role of titanium mesh and porous polyethylene. AB - OBJECTIVE: To report on 8 years of experience with 156 titanium mesh and porous polyethylene implants used for craniofacial reconstruction after skull base surgery in 100 patients. DESIGN: Cohort study with a mean follow-up of 5 years. SETTING: Population based. PATIENTS: A consecutive sample of 100 patients treated for skull base tumors or craniofacial trauma who underwent reconstruction with 156 3-dimensional titanium mesh and/or porous polyethylene implants. A retrospective review of the Skull Base Program database, along with photographic and imaging documentation, was undertaken. MAIN OUTCOME MEASURES: Rate of complications as well as the degree of functional and esthetic reconstruction. INTERVENTION: The reconstructive technique focused primarily on the substitution of removed craniofacial skeleton for oncologic reasons or soft tissue defects. RESULTS: After completion of follow-up (mean, 5 years), all 100 patients remained healed except for 7 patients (7%) with 8 implants (5%). Overall, excellent craniofacial symmetry and stability were achieved with both types of implants. CONCLUSIONS: Immediate craniofacial skeletal reconstruction and soft tissue augmentation is feasible with 3-dimensional titanium mesh and porous polyethylene implants. The reviewed 8-year evolution in the use of these technologies (156 implants in 100 patients) highlights the excellent tolerance of these implants (5% implant complication rate) in 100 patients (7% complication rate). The few encountered complications were judged to be primarily related to the quality of the overlying soft tissue and not to the implants themselves. The advantages of using these implants for immediate 3-dimensional skeletal and soft tissue substitution, including availability, easy contouring, stability, primary healing, and tolerance of adjuvant therapy, translate to an improved function and esthetic appearance, with a better quality of life for patients. PMID- 10722016 TI - PH20: a novel tumor marker for laryngeal cancer. AB - OBJECTIVE: To determine whether levels of PH-20, a hyaluronidase similar to that found in human sperm, are elevated in laryngeal cancer tissue. DESIGN: In this case-control study. reverse transcription polymerase chain reaction was used to measure levels of PH-20 messenger RNA in tissue taken from laryngectomy specimens. SETTING: A university medical center. PATIENTS: We compared tissue samples taken from 11 patients with laryngeal cancer, and from 2 metastatic lymph nodes, with samples of normal, healthy laryngeal tissue and prostate cancer tissue (positive control). MAIN OUTCOME MEASURE: PH-20 complementary DNA expression as quantified by densitometric analysis. RESULTS: Expression of PH-20 was significantly higher in nonirradiated laryngeal cancer specimens than in normal laryngeal tissue (P<.01). Metastatic lymph nodes also had higher levels of PH-20 expression than did primary laryngeal cancer tissue (P = .11) and normal laryngeal tissue (P<.01). Irradiated laryngeal cancer specimens had PH-20 levels comparable to normal. CONCLUSIONS: We report the first data on PH-20 expression in laryngeal cancer tissue. PH-20 expression is significantly elevated in primary laryngeal cancer tissue and seems to be even higher in metastatic lesions compared with normal laryngeal tissue. PH-20 may be a useful tumor marker and prognostic tool for laryngeal cancer. PMID- 10722017 TI - Acid/pepsin promotion of carcinogenesis in the hamster cheek pouch. AB - OBJECTIVE: While clinical observation has suggested an association between gastroesophageal reflux and laryngeal carcinoma, the nature of this relationship has yet to be defined. The purpose of this study is to determine the carcinogenic potential of acid and pepsin mixtures in the hamster cheek pouch animal model. DESIGN: A blinded intervention study. SUBJECTS: One hundred male Syrian hamsters aged approximately 5 weeks. INTERVENTIONS: A control group of 20 hamsters received application of the carcinogen 9,10-dimethyl-1,2-henzanthracene (DMBA) to their cheek pouch mucosa. One experimental group (n = 20) received applications of DMBA plus hydrochloric acid, and another (n = 20) received DMBA plus an acid and pepsin solution. Latency to squamous cell tumor production, size of tumors, and numbers of tumors were compared among groups. RESULTS: Latency to tumor production and size of tumor were similar among groups, with both experimental and control groups developing tumors of comparable size after 12 weeks of chemical application. However, the number of tumors produced was significantly higher in the DMBA/acid and DMBA/acid/ pepsin groups than in the DMBA only group at 18 weeks, with 23, 27, and 10 tumors in these groups, respectively (P<.02). Likewise, a cumulative dysplasia score was different among groups at 18 weeks with the DMBA/acid and DMBA/acid/pepsin groups scoring higher degrees of dysplasia than the DMBA only group. CONCLUSION: These results suggest that application of acid and acid/pepsin mixtures may promote experimental carcinogenesis in the hamster cheek pouch. PMID- 10722018 TI - Ipsilateral neck cancer recurrences after elective supraomohyoid neck dissection. AB - DESIGN: Retrospective analysis of a case series. SETTING: Referral center, private or institutional practice, hospitalized care. OBJECTIVE: To analyze the level (site) of ipsilateral neck recurrences after supraomohyoid (SOH) dissection in patients with lip, oral, and oropharyngeal cancer treated in a single institution. INTERVENTION: Supraomohyoid neck dissection. PATIENTS AND METHODS: From 1979 to 1997, 154 patients with oral and oropharyngeal carcinoma and no palpable lymph nodes at the neck underwent ipsilateral elective SOH dissection. RESULTS: Tumor sites were the lip, 5 cases (3.3%); oral cavity, 128 cases (83.1%); and oropharynx, 21 cases (13.6%). Tumor stages were T1, 13 cases (8.4%); T2, 77 cases (50.0%); T3, 40 cases (27.0%); and T4, 22 cases (14.3%). There were 7 cases (4.5%) of ipsilateral neck recurrences. Three were beyond the limits of the SOH dissection, and 4 were inside these limits. There was no association of neck recurrences with the pathological status of the lymph nodes. Six of the 7 recurrences were in patients who underwent postoperative radiotherapy. CONCLUSIONS: The incidence of neck recurrence after selective neck dissection was 4.5%, and it occurred either inside (57.1%) or beyond (42.9%) the limits of the selective neck dissection. PMID- 10722019 TI - The role of supraomohyoid neck dissection in patients with positive nodes. AB - BACKGROUND: Supraomohyoid neck dissection (SOHND) is currently used as a staging procedure for patients with clinically negative nodes in the neck who are at increased risk (>20%) for metastatic disease. OBJECTIVE: To assess the potential role of SOHND in patients with clinically positive nodes at levels I, II, or III. We evaluated, in particular, whether selective neck dissection in patients with clinically positive nodes results in decreased regional control and/or diminished survival. PATIENTS AND METHODS: We retrospectively reviewed the charts of all patients who underwent SOHND from January 1, 1971, to December 31, 1997. The oral cavity and oropharynx represented the primary sites in the majority of the patients. Two-year follow-up information was available on all patients. RESULTS: During the study period, 69 patients underwent 84 SOHNDs. Of the 69 patients, there were 30 patients with clinically negative nodes and 39 patients with clinically positive nodes in the neck. The overall regional control rates were 88% vs 71% for pathologically negative vs positive nodes, respectively, with or without adjuvant radiation therapy. Adjuvant radiation therapy significantly improved regional control in patients with pathologically positive nodes but not in patients with NO disease (P = .005). Similar results were noted in patients with both clinically and pathologically positive nodes. CONCLUSIONS: Supraomohyoid neck dissection in patients with pathologically positive nodes in the neck is inadequate therapy for regional control without postoperative radiation therapy. However, in patients with pathologically positive nodes in the neck, SOHND with postoperative radiation therapy can achieve regional control comparable to that of comprehensive neck dissection and postoperative radiation therapy. PMID- 10722020 TI - Endoscopic neck dissection in an animal model: comparison of nodal yield with open-neck dissection. AB - OBJECTIVE: To evaluate the possibility, complications, and efficacy of endoscopic neck dissection (END) in a porcine model. DESIGN: Experimental self-controlled study. SUBJECTS: Minipigs. INTERVENTION: Endoscopic neck dissection was performed using general anesthesia with techniques adapted from laparoscopic surgery. The tissue specimens removed were divided according to porcine equivalents of human neck groups. After the completion of END, open-neck dissection was performed using standard surgical techniques, and the remaining tissue within each neck group was retrieved. A pathologist evaluated each specimen without knowing its exact origin in terms of neck group or side and the type of surgical technique used. For each specimen, the number of retrieved lymph nodes and their anatomical integrity were analyzed. RESULTS: Ten neck dissections were performed in 8 minipigs without any major complications. The number of retrieved lymph nodes by END was 18.4 +/- 7.4 (mean +/- SD). Completed open-neck dissection retrieved an additional 3.3 +/- 1.8 lymph nodes. The efficacy rate of END was 88% +/- 10% (+/ SD). The majority of retrieved lymph nodes were intact, with less than 5% of nodes exhibiting crushing artifacts. CONCLUSIONS: Endoscopic neck dissection in a porcine model seems to be free of major complications and able to retrieve the majority of neck lymph nodes. A larger number of animals and their survival need to be studied before human studies can begin. PMID- 10722021 TI - A pilot study evaluating the treatment of postparotidectomy sialoceles with botulinum toxin type A. AB - OBJECTIVE: To report our experience in treating 4 cases of recurrent siacloceles with botulinum toxin type A after partoid surgery. DESIGN: This is a prospective, nonrandomized, nonblinded pilot study describing a new use for botulinum toxin type A. SETTING: Tertiary academic medical center. PATIENTS: Four patients (2 men and 2 women) with persistent postparotidectomy sialoceles who had undergone various treatment failures were included. The diagnosis was made by fine-needle aspiration of the mass based on well-recognized cytologic features of the entity, as well as an elevated amylase level and no evidence of tumor or infection. INTERVENTIONS: Sialoceles were aspirated before local injection of botulinum toxin type A (30-50 U) subcutaneously. MAIN OUTCOME MEASURES: The patients were followed up 1 week after receiving botulinum toxin type A injection and then at monthly intervals. They were extensively questioned and examined for any evidence of side effects or recurrence. RESULTS: All patients had total resolution of sialocele or external salivary fistula within 1 month of treatment. None of the patients to date have demonstrated recurrences at 7 through 13 months, and there were no complications, particularly facial nerve weakness. CONCLUSION: Our findings suggested that botulinum toxin type A offers a highly effective, safe, and noninvasive method of treatment in postparotidectomy sialocele. PMID- 10722022 TI - Sensory changes associated with selective neck dissection. AB - OBJECTIVE: To evaluate sensory changes in the head and neck region associated with selective neck dissection with or without preservation of cervical root branches. DESIGN: Retrospective cohort study. SETTING: University tertiary referral hospital and a Veterans Affairs hospital. PATIENTS: Fifty-seven patients who had undergone 84 neck dissections with or without preservation of the sensory cervical root branches 3 or more months before evaluation. INTERVENTIONS: Questionnaire combined with head and neck sensory examination. MAIN OUTCOME MEASURES: Neck and facial sensory function. RESULTS: Neck dissections with preservation of the cervical rootlets were most likely to be associated with a small area of anesthesia in the upper neck below the body of the mandible and anterior to the mid-body of the mandible (P=.03). Neck dissections without rootlet-preserving technique increased the area of anesthesia to include all other areas of the neck (P= .02). CONCLUSIONS: Preservation of the cervical root branches resulted in a small, limited, and uniform area of the neck rendered permanently anesthetic. Conversely, sacrifice of the nerve branches led to a pattern of anesthesia involving the entire neck. PMID- 10722023 TI - Pulmonary complications in patients with head and neck and lung neoplasms. AB - BACKGROUND: The occurrence of second primary neoplasms in patients with head and neck carcinoma assumes greater importance as our ability to control local disease improves. Both the primary lesions and the therapeutic interventions can predispose patients to pulmonary complications. OBJECTIVE: To explore the incidence of pulmonary complications in patients with head and neck cancer who also undergo lung surgery. DESIGN: Survey; case series. SETTING: A tertiary care university hospital. RESULTS: The clinic and hospital charts of 32 patients with multiple interventions of the head and neck and lung were retrospectively reviewed, and data were recorded on the clinical and pathologic specifics of primary and secondary neoplasms, pulmonary complications, and treatment outcomes. Twenty-eight (88%) of these patients underwent a diagnostic or therapeutic surgical procedure for a head and neck primary neoplasm. All patients (100%) underwent a pulmonary resection for malignant or nonmalignant pulmonary disease. Overall, 31 patients (97%) experienced either major or minor pulmonary complications after surgery, 51 (79%) of which occurred during the immediate postoperative course. Major complications occurred in 11 patients (34%), which were fatal in one. CONCLUSIONS: Our data suggest that patients with head and neck cancer who also experience a second pulmonary disease requiring lung resection are at high risk of developing serious pulmonary complications. These risks should be considered in planning optimal therapy. PMID- 10722024 TI - Management of malignant melanoma of the head and neck using dynamic lymphoscintigraphy and gamma probe-guided sentinel lymph node biopsy. AB - BACKGROUND: The sentinel lymph node (SLN) biopsy is revolutionizing the surgical management of primary malignant melanoma. It allows accurate nodal staging, and targets patients who may benefit from regional lymphadenectomy and systemic therapy; however, its use in the management of head and neck melanoma has not been widely accepted. METHODS: A retrospective review of patients treated for clinical stages I and II malignant melanoma of the head and neck with dynamic lymphoscintigraphy and gamma probe-guided SLN biopsy. RESULTS: Fifty-eight patients (47 male and 11 female) were identified. Primary melanoma sites included the scalp (21), ear (8), face (13), neck (15), and eyelid (1). Primary tumor staging was T2 (11), T3 (24), and T4 (23). Dynamic lymphoscintigraphy visualized SLNs in 57 patients (98.3%). In 43 cases (75%) a single draining nodal basin was identified, and in 14 cases there were multiple draining nodal basins. Sentinel lymph nodes were successfully identified in 72 (96%) of 75 nodal basins. Positive SLNs were identified in 10 patients (17.5%). Sentinal lymph node positivity by tumor staging was T3, 16.7% and T4, 27.3%. Completion lymphadenectomy revealed residual disease in 3 patients (30%). Relapse occurred in 10 (21.3%) of the 47 patients with negative SLN biopsy results and 7 (70%) of those with positive results. CONCLUSIONS: Gamma probe-guided SLN localization in the head and neck region was successful in 96% of draining nodal basins. It can target regional lymphadenectomy in patients who may benefit from regional nodal dissection. PMID- 10722025 TI - The impact of laparoscopy and laparoscopic ultrasound on the management of pancreatic cystic lesions. AB - HYPOTHESIS: Laparoscopic ultrasound examination combined with biopsy of the cystic wall and aspiration of cystic fluid improves differential diagnosis of pancreatic cystic lesions contributing to surgical decision making. STUDY DESIGN: A prospective evaluation of the impact of laparoscopic ultrasound on surgical decision making in patients with pancreatic cysts. SETTING: A general community hospital; the department of surgery serves as referral for pancreatic surgery. PATIENTS: During a 36-month period, 15 patients with pancreatic cystic lesions were prospectively evaluated by laparoscopy and laparoscopic ultrasound with ultrasound-guided biopsy of the cystic wall and aspiration of cystic fluid for cytologic study, viscosity, and determination of levels of amylase and tumor markers (carcinoembryonic antigen, cancer antigen 19.9). RESULTS: Laparoscopic ultrasound contributed new, additional data in 8 patients (53%) when compared with compiled imaging data obtained by conventional ultrasound, computed tomography, magnetic resonance imaging, and endoscopic ultrasound. A solid cystic component was detected in 6 patients and additional small (<1 cm) cysts in 3 patients. Amylase and tumor marker levels, biopsy of the cystic wall, and cytologic examination had significant impact on surgical decision making in 6 patients. Nine patients underwent resection of the cystic lesion. Three patients diagnosed as having benign cysts had laparoscopy with laparoscopic ultrasound only. Three patients with suspicious lesions refused surgery. Laparoscopic ultrasound predicted correctly the nature of the cyst in 7 of 9 surgically treated patients (sensitivity, 78%). Two patients with serous cystadenoma had high levels of tumor markers (false-positive). CONCLUSION: Although a rather invasive procedure that requires general anesthesia and hospitalization, laparoscopy with laparoscopic ultrasonography was found to significantly contribute to the differential diagnosis of pancreatic cystic lesions. PMID- 10722026 TI - Effect of posture on popliteal artery hemodynamics. AB - HYPOTHESIS: Marked peripheral vasodilation and rubor characterize critically ischemic limbs on dependency. We believe that intermittent claudication is also associated with peripheral hemodynamic changes on postural alteration, which differ distinctly from normal. Evaluation of such differences and understanding of the underlying physiological derangements may be essential in the development of treatments for intermittent claudication. We comparatively assess the effect of posture on lower limb arterial hemodynamics in normal subjects and in patients with intermittent claudication (or Fontaine II) due to peripheral vascular disease, determined in the popliteal artery. DESIGN: A cohort study. SETTING: A university-associated tertiary care hospital. PATIENTS: Thirty-seven legs of 29 normal subjects (group A) and 50 legs of 36 patients with intermittent claudication (ankle-brachial index range, 0.39-0.76; median, 0.57) (group B). INTERVENTIONS: Popliteal artery volume flow (vFl), mean velocity, and luminal diameter were measured on (1) recumbency, (2) sitting, and (3) return to recumbency in groups A and B using color duplex imaging. MAIN OUTCOME MEASURES: The pulsatility index, peak systolic velocity, and end diastolic velocity (EDV) were measured on (1) recumbency, (2) sitting, and (3) return to recumbency. RESULTS: Popliteal artery vFl in normal subjects decreased from 110 +/- 43 mL/min on recumbency to 57 +/- 27 mL/min on sitting (P<.001) and returned to 111 +/- 46 mL/ min on resumption of recumbency (P<.001). Similarly, in patients with intermittent claudication, vFl decreased from 113 +/- 52 mL/min on recumbency to 76 +/- 41 mL/min on sitting (P<.001) and increased on resumption of recumbency to 114 +/- 53 mL/min (P<.001). There was no difference (P = .97) in the vFl between the study groups on recumbency, but sitting vFl in normal subjects was significantly lower than in patients with intermittent claudication (P = .04). The mean velocity, peak systolic velocity, and EDV displayed a similar pattern of change as vFl. The pulsatility index in both groups increased significantly on sitting (P<.001) and decreased on return to recumbency (P<.001). All data are given as mean +/- SD. CONCLUSIONS: Lower limb arterial vFl, mean velocity, peak systolic velocity, and EDV decrease significantly (P<.001) when posture is altered from recumbency to sitting, in normal subjects and in patients with intermittent claudication. A decrease in the EDV and an increase in the pulsatility index on sitting indicate enhancement of arterial resistance to flow secondary to peripheral vasoconstriction. Quantitative differences between the groups in vFl (P<.04), EDV (P<.01), and pulsatility index (P<.001) on dependency indicate that the orthostatic vasoactive response in patients with intermittent claudication is significantly subdued, reflecting a marked derangement in venoarteriolar response. PMID- 10722027 TI - Papillary thyroid carcinoma: prognostic index for survival including the histological variety. AB - BACKGROUND: Numerous prognostic factors have been studied for survival in patients with papillary thyroid carcinoma (PTC), although there are few multivariate studies that include the histological variety of PTC. HYPOTHESIS: There are prognostic factors that influence survival in a series of patients with PTC, including the histological variety, and a new prognostic index (PI) for survival can be formulated by accounting for these factors. DESIGN: A retrospective study. SETTING: A university hospital department of surgery. PATIENTS: Between January 1970 and December 1995, 200 patients undergoing surgery for PTC were observed (mean follow-up, 8 years). MAIN OUTCOME MEASURES: A univariate analysis was done for survival rates using the Kaplan-Meier estimation method. The possible prognostic factors were evaluated using a multivariate analysis according to the Cox model. We formulated a PI and defined 3 risk groups (low, medium, and high) for mortality. RESULTS: Of the 200 patients, 175 (87.5%) are still alive. Of the 25 deaths, 19 (9.5%) were due to the tumor. The survival was 97.5% at 1 year, 92.8% at 5, 89.5% at 10, and 83.9% at 15 and 20 years. The prognostic factors obtained after the multivariate analysis were age, tumor size, extrathyroid spread, and histological variant of the PTC. The PI is calculated as follows: PI = (2 x size) + (6 x spread) + (2 x variant) + (3 x age). As for the risk groups, the low-risk group showed a mortality of 0%; the medium-risk group, 17.1%; and the high-risk group, 76.5%. CONCLUSIONS: The histological variety of PTC has prognostic value for survival in patients with PTC. As risk factors for PTC mortality, we consider an age of 50 years or older, a tumor larger than 4 cm, the existence of extrathyroid spread, and a certain histological subtype of PTC. With these risk factors, it is possible to formulate a PI and classify patients into low-, medium-, and high-risk groups for mortality. PMID- 10722028 TI - Neurological damage and duodenopancreatic reflux in the pathogenesis of alcoholic pancreatitis. AB - OBJECTIVE: To present a new theory on the pathogenesis of acute alcoholic pancreatitis based on experimental data, the significance of which has not been recognized, and on evidence from the current literature. HYPOTHESIS: That chronic alcoholism damages muscarinic receptors in the pancreas, duodenum, and Oddi sphincter, producing heightened sensitivity to acetylcholine, stimulation of protein-rich pancreatic juice, hypertonicity of the duodenum and esophagus, relaxation of the Oddi sphincter, and intraduodenal pressures exceeding those shown to cause duodenopancreatic reflux and acute pancreatitis in humans and experimental animals. OUTCOME: The duodenopancreatic reflux mechanism can explain all of the clinical features of acute alcohol pancreatitis, including the intraductal site and rapid activation of zymogens by enterokinase, the recurrent episodes of pancreatitis, the precipitation of protein plugs by partial proteolytic hydrolysis, the severe vascular changes, the relation to infection by the most direct route, and the progression to chronic pancreatitis via the necrosis-fibrosis sequence. CONCLUSIONS: Damage to the nervous system, with a time lag of 5 to 15 years between the onset of heavy drinking and the development of neurological disorders (peripheral neuropathy and cerebellar degeneration), is a characteristic complication of chronic alcoholism. The similarity to events in alcoholic pancreatitis is striking. PMID- 10722029 TI - Contrast-enhanced dynamic computed tomography does not aggravate the clinical severity of patients with severe acute pancreatitis: reevaluation of the effect of intravenous contrast medium on the severity of acute pancreatitis. AB - BACKGROUND: Contrast-enhanced abdominal computed tomography (CT) is useful in demonstrating pancreatitis necrosis, but the administration of contrast medium in animal models with acute pancreatitis may worsen the severity. HYPOTHESIS: The use of contrast-enhanced CT in clinical patients with acute pancreatitis may actually aggravate the severity of the disease. DESIGN: A randomized prospective study. SETTING: Chang Gung Memorial Hospital, Taipei, Taiwan. PATIENTS: Twenty patients with severe acute pancreatitis were randomly divided into 2 groups. Those in group A (n = 10) underwent a CT examination with a contrast-enhanced medium, and those in group B (n = 10) underwent a CT examination without a contrast-enhanced medium. MAIN OUTCOME MEASURES: The patients' serum amylase, lipase, C-reactive protein, leukocyte, glutamicoxaloacetic transaminase, creatinine, calcium, and phosphate levels were serially checked before the CT examination and at 2, 4, 6, 8, 12, and 24 hours after the examination was performed. The biochemical data between the 2 groups were compared. The morbidity, length of stay, and mortality were also compared. RESULTS: There were no significant changes in the level of pancreatic enzymes, C-reactive proteins, and leukocytes and in the biochemical data of either group before or after the CT examination. The difference in the previously examined values between the 2 groups was also not significant. There was also no difference in the morbidity, length of hospital stay, and mortality between the 2 groups. CONCLUSION: Contrast enhanced abdominal CT does not aggravate the severity of clinical patients with severe acute pancreatitis. PMID- 10722030 TI - Is interleukin 6 an early marker of injury severity following major trauma in humans? AB - HYPOTHESIS: Interleukin 6 (IL-6), a multifunctional cytokine, is expressed by various cells after many stimuli and underlies complex regulatory control mechanisms. Following major trauma, IL-6 release correlates with injury severity, complications, and mortality. The IL-6 response to injury is supposed to be uniquely consistent and related to injury severity. Therefore, we designed a prospective study starting as early as at the scene of the unintentional injury, to determine the trauma-related release of plasma IL-6 in multiple injured patients. PATIENTS AND METHODS: On approval of the local ethics committee, 94 patients were enrolled with different injuries following trauma (Injury Severity Score [ISS] median, 19; range, 3-75). The patients were rescued by a medical helicopter. Subsets were performed according to the severity of trauma--4 groups (ISS, <9, 9-17, 18-30, and >32)-and survival vs nonsurvival. The first blood sample was collected at the scene of the unintentional injury before cardiopulmonary resuscitation, when appropriate. Then, blood samples were collected in hourly to daily intervals. Interleukin 6 plasma levels were determined using a commercial enzyme-linked immunosorbent assay test. The short term phase protein, C-reactive protein, was measured to characterize the extent of trauma and to relate these results to IL-6 release. RESULTS: As early as immediately after trauma, elevated IL-6 plasma levels occurred. This phenomenon was pronounced in patients with major trauma (ISS, >32). Patients with minor injury had elevated concentrations as well but to a far lesser extent. In surviving patients, IL-6 release correlated with the ISS values best during the first 6 hours after hospital admission. All patients revealed increased C reactive protein levels within 12 hours following trauma, reflecting the individual injury severity. This was most pronounced in patients with the most severe (ISS, >32) trauma. CONCLUSIONS: To our knowledge, this is the first study that elucidates the changes in the IL-6 concentrations following major trauma in humans as early as at the scene of the unintentional injury. The results reveal an early increase of IL-6 immediately after trauma. Moreover, patients with the most severe injuries had the highest IL-6 plasma levels. There is strong evidence that systemic IL-6 plasma concentrations correlate with ISS values at hospital admission. Therefore, IL-6 release can be used to evaluate the impact of injury early regardless of the injury pattern. PMID- 10722031 TI - Minimally invasive cardiac surgery defined. AB - There is no operation as complex, yet as fundamentally unchanged over time, as conventional coronary artery bypass grafting (CABG). This remarkable achievement is attributed to the operation's adaptability to a wide variety of clinical settings; its reproducibility, although performed by surgeons all across the world; and its proved track record for safety and effectiveness. A monumental effort, however, is currently under way to redefine CABG. This paradigm shift has received a groundswell of support as advances in minimally invasive surgery in other areas, such as arthroscopy, laparoscopic cholecystectomy, and thoracoscopy, combined with an increasing focus on cost containment, have forever changed the milieu of the cardiac surgeon. This review examines the clinical and research issues surrounding minimally invasive CABG from the vantage point of a surgeon scientist working in the field. PMID- 10722032 TI - Major liver resection for carcinoma in jaundiced patients without preoperative biliary drainage. AB - BACKGROUND: The role of preoperative biliary drainage (PBD) before liver resection in the presence of obstructive jaundice remains controversial. Our patients with proximal duct carcinoma undergo noninvasive assessment followed by rapid laparotomy without PBD if the lesion is deemed resectable. HYPOTHESIS: Our aim was to report operative outcome of these patients and to analyze their specific features by comparison with patients without biliary obstruction who underwent major liver resection. DESIGN: A case-comparison study. SETTING: A tertiary care university hospital in a metropolitan area. PATIENTS: Twenty consecutive jaundiced patients underwent major liver resection without PBD. The jaundiced patients were matched with 27 nonjaundiced patients with normal underlying liver selected from a computer bank of 261 patients undergoing liver resections and identical for age, tumor size, type of liver resection, and vascular occlusion. MAIN OUTCOME MEASURE: Postoperative course including mortality, morbidity, transfusion rates, and results of liver function tests. RESULTS: Seventeen jaundiced patients (85%) and 13 nonjaundiced patients (48%) received blood transfusions (P = .03). Morbidity was 50% in jaundiced and 15% in nonjaundiced patients (P = .006), mainly resulting from subphrenic collections and bile leaks occurring only in jaundiced patients. In contrast, there were no significant differences for mortality (5% vs 0%) and liver failure (5% vs 0%). Postoperative changes in liver function test results were comparable between groups. CONCLUSIONS: Major liver resections without PBD are safe in most patients with obstructive jaundice. Recovery of hepatic synthetic function is identical to that of nonjaundiced patients. Transfusion requirements and incidence of postoperative complications, especially bile leaks and subphrenic collections, are higher in jaundiced patients. Whether PBD could improve these results remains to be determined. PMID- 10722033 TI - Chlorhexidine lavage in the treatment of experimental intra-abdominal infection. AB - HYPOTHESIS: Closed postoperative peritoneal lavage (CPPL) with chlorhexidine gluconate reduces the number of intraperitoneal bacteria and improves the outcome of intra-abdominal infection. DESIGN: Laboratory animal trial. INTERVENTIONS: Intra-abdominal infection was produced in mice by the cecal ligation and puncture technique. After 16 to 18 hours, the animals underwent relaparotomy and placement of an intra-abdominal catheter for CPPL. In the first experiment animals were randomly divided into 4 groups: no lavage (served as a control), CPPL with chlorhexidine. CPPL with cefoxitin, and CPPL with lactated Ringer solution (LR). Lavage was continued intermittently every 8 hours for 24 hours. All animals received systemic cefoxitin every 8 hours for 7 days. Mortality was recorded every 8 hours for 10 days. In the second experiment, animals were divided into 3 groups: no lavage (served as a control), CPPL with chlorhexidine, and CPPL with LR. Lavage was continued intermittently every 8 hours for 24 hours. The animals were killed 48 hours after reoperation. Bacterial counts from peritoneal fluid and biopsy specimens, as well as peritoneal white blood cell counts, were measured before and after lavage. RESULTS: Closed postoperative peritoncal lavage with chlorhexidine reduced mortality from 71% in a control group to 37% (P = .003). There was no survival benefit in either the CPPL with cefoxitin (91% mortality) (P = .14) or CPPL with LR groups (90% mortality) (P = .17). The statistically significant findings of analysis of variance evaluation demonstrated a decrease in bacterial counts after cecal excision in all 3 groups. There was a greater reduction in bacterial counts in the chlorhexidine group compared with the control group (P<.05). Bacterial counts decreased in peritoneal fluid, as well as in tissue biopsy specimens, after cecal excision. White blood cell counts significantly decreased after cecal excision in all 3 groups. There was no difference in white blood cell counts between the groups. Correlation analyses demonstrated weak interaction between bacterial and white blood cell counts before or after treatment in all the groups. Pearson r ranged from -0.37 to +0.35, none of which were statistically significant. CONCLUSIONS: In our experiments chlorhexidine lavage resulted in a 50% reduction in mortality and a significant reduction in bacterial counts compared with the control group. There was no survival benefit from lavage with either cefoxitin or LR. There was no reduction in bacterial counts in the LR group relative to the control group. Thus, the survival benefit and the reduction in bacterial numbers are attributed to the antibacterial properties of chlorhexidine rather than to the mechanical washing of the abdominal cavity. Closed postoperative peritoneal lavage with 0.05% chlorhexidine gluconate might be useful in the multimodal treatment of intra-abdominal infection. PMID- 10722034 TI - Emergency medical services (EMS) vs non-EMS transport of critically injured patients: a prospective evaluation. AB - BACKGROUND: A previous report of 5,782 trauma patients demonstrated higher mortality among those transported by emergency medical services (EMS) than among their non-EMS-transported counterparts. HYPOTHESIS: Trauma patients who are transported by EMS and those who are not differ in the injury-to-hospital arrival time interval. DESIGN: Prospective cohort-matched observation study. SETTING: Level I trauma center, multidisciplinary study group. PATIENTS: All non-EMS patients were matched with the next appropriate EMS patient by an investigator who was unaware of the outcome and mode of transport. Every 10th EMS patient with an Injury Severity Score (ISS) of 13 or greater was also randomly enrolled. Matching characteristics included age, ISS, mechanism of injury, head Abbreviated Injury Score, and presence of hypotension. An interview protocol was developed to determine the time of injury. Interview responses from patients, witnesses, and friends were combined with data obtained from police, sheriff, and medical examiner reports. MAIN OUTCOME MEASURES: Time to the hospital, mortality, morbidity, and length of stay. RESULTS: A total of 103 patients were enrolled (38 non-EMS, 38 EMS matched, 27 random EMS). Injury time was estimated using all available data made on 100 patients (97%). Independent raters agreed in 81% of cases. Deaths, complications, and length of hospital stay were similar between the EMS- and non-EMS-transported groups. Although time intervals were similar among the groups overall, more critically injured non-EMS patients (ISS > or = 13) got themselves to the trauma center in less time than their EMS counterparts (15 minutes vs 28 minutes; P<.05). CONCLUSIONS: A multidisciplinary approach can be utilized, and an interview protocol created to determine actual time of injury. Critically injured non-EMS-transported patients (ISS > or =13) arrived at the hospital earlier after their injuries. PMID- 10722035 TI - Outcome for older burn patients. AB - BACKGROUND: Physicians will be increasingly responsible for an aging society whose members demonstrate a notable striving for independence. HYPOTHESIS: With standard treatment of burns, older patients will have a survival rate of more than 70%, with at least 60% of patients becoming fully functional 6 months after hospital discharge. METHODS: A 7-year retrospective medical review of burn unit patients was performed, and 221 ( 11%) of 1957 patients who were at least 59 years old were identified. RESULTS: Of 97 women (44%) and 124 men (56%), 64 (29%) had an associated smoke inhalation injury; 146 (66%), flame injury; and 44 (20%), scald injury. The bedroom and/or living room were the most common areas of injury (90 [41%]), followed by outdoors and the workplace (62 [28%]), the kitchen (40 [18%]), the bathroom ( 18 [8%]), and the garage or basement (11 [5%]) (P<.005). One hundred twenty-six injuries (57%) were associated with impaired judgment, mobility, or both. On hospital admission, 74 patients (36%) were intubated, 60 (30%) required intubation postoperatively, and 34 (18%) required both. The survival rate was 159 patients (72%) overall. Findings from an ethanol screening and a drug toxicology screening were positive in 22 and 32 patients (10% and 29%) on admission, respectively. Of the survivors, most were discharged to home with (87 [64%) or without visiting nurse supervision, and at 6 months after discharge, 16 patients (50%) in transitional care facilities were able to return to an independent level of functioning. Of the 59- to 69-year-old age group, 83 (86%) survived compared with 59 (69%) in the 70- to 79-year-old age group and 18 (47%) in the 80 years and older age group. CONCLUSIONS: In contrast to the usual male preponderance in patients with thermal injury, older women, many of whom are widowed, constituted almost half of the older patients admitted to the hospital. Modalities for injury prevention are necessary to provide optimal and safe household environments for a growing population of older persons. PMID- 10722036 TI - Reduction of resuscitation fluid volumes in severely burned patients using ascorbic acid administration: a randomized, prospective study. AB - HYPOTHESIS: High-dose ascorbic acid (vitamin C) therapy (66 mg/kg per hour) attenuates postburn lipid peroxidation, resuscitation fluid volume requirements, and edema generation in severely burned patients. STUDY DESIGN AND SETTING: A prospective, randomized study at a university trauma and critical care center in Japan. SUBJECTS AND METHODS: Thirty-seven patients with burns over more than 30% of their total body surface area (TBSA) hospitalized within 2 hours after injury were randomly divided into ascorbic acid and control groups. Fluid resuscitation was performed using Ringer lactate solution to maintain stable hemodynamic measurements and adequate urine output (0.5-1.0 ml/kg per hour). In the ascorbic acid group (n = 19; mean burn size, 63% +/- 26% TBSA; mean burn index, 57 +/- 26; inhalation injury, 15/ 19), ascorbic acid was infused during the initial 24-hour study period. In the control group (n = 18; mean burn size, 53% +/- 17% TBSA; mean burn index, 47 +/- 13; inhalation injury, 12/18), no ascorbic acid was infused. We compared hemodynamic and respiratory measurements, lipid peroxidation, and fluid balance for 96 hours after injury. Two-way analysis of variance and Tukey test were used to analyze the data. RESULTS: Heart rate, mean arterial pressure, central venous pressure, arterial pH, base deficit, and urine outputs were equivalent in both groups. The 24-hour total fluid infusion volumes in the control and ascorbic acid groups were 5.5 +/- 3.1 and 3.0 +/- 1.7 mL/kg per percentage of burn area, respectively (P<.01). In the first 24 hours, the ascorbic acid group gained 9.2% +/- 8.2% of pretreatment weight; controls, 17.8% +/- 6.9%. Burned tissue water content was 6.1 +/- 1.8 vs 2.6 +/- 1.7 mL/g of dry weight in the control and ascorbic acid groups, respectively (P<.01). Fluid retention in the second 24 hours was also significantly reduced in the ascorbic acid group. In the control group, the ratio of PaO2 to fraction of inspired oxygen at 18, 24, 36, 48, and 72 hours after injury was less than that of the ascorbic acid group (P<.01). The length of mechanical ventilation in the control and ascorbic acid groups was 21.3 +/- 15.6 and 12.1 +/- 8.8 days, respectively (P<.05). Serum malondialdehyde levels were lower in the ascorbic acid group at 18, 24, and 36 hours after injury (P<.05). CONCLUSIONS: Adjuvant administration of high-dose ascorbic acid during the first 24 hours after thermal injury significantly reduces resuscitation fluid volume requirements, body weight gain, and wound edema. A reduction in the severity of respiratory dysfunction was also apparent in these patients. PMID- 10722037 TI - Thoracoscopic splanchnicectomy for pain relief in unresectable pancreatic cancer. AB - HYPOTHESIS: Unilateral truncal thoracoscopic splanchnicectomy (TS) provides safe and effective treatment for pain relief in patients with unresectable pancreatic cancer. DESIGN: Before-and-after trial of 24 patients undergoing 25 TS procedures. SETTING: Surgical unit at a university teaching hospital. PATIENTS: A consecutive sample of 24 patients with severe pain due to unresectable (primary or recurrent) pancreatic cancer refractory to drug therapy and with a life expectancy of less than 6 months. INTERVENTION: The key point of the reported operation is intrathoracic carbon dioxide insufflation, which allows a more distal division of the greater splanchnic nerve and a 2-port technique. MAIN OUTCOME MEASURES: Pain and the effect of this symptom on quality of life were assessed before and after TS using a 10-point visual analog pain scale (VAS) and the Nottingham Health Profile questionnaire, respectively. RESULTS: Four TS procedures were technical failures because of pleural adhesions. One patient required a contralateral procedure 12 weeks after TS. Mean (+/- SD) preoperative VAS basal score was 7.4 +/- 1.7. Twenty-four hours after TS, it was reduced to 0.6 +/- 1.0. Significant reduction of VAS scores persisted over the first 3 months after TS (P<.001). Recurrence of pain of low intensity (mean VAS basal score, 4.2) was observed in 8 patients. Significant improvement (P<.001) in each area covered by the Nottingham Health Profile questionnaire was reported at 1 month after TS. CONCLUSION: Thoracoscopic splanchnicectomy offered substantial short-term relief of pain in patients with unresectable pancreatic cancer, and significantly ameliorated the quality of their residual life. PMID- 10722038 TI - Safety of donors in live donor liver transplantation using right lobe grafts. AB - HYPOTHESIS: Right lobe donation was advocated for adult-to-adult live donor liver transplantation but the safety of the donor is still a major concern. We hypothesize that right lobe donation is safe if the lowest limit of volume of liver remnant that can support donor survival is known. DESIGN: Retrospective analysis of data collected prospectively. SETTING: Tertiary hepatobiliary surgery referral center. PATIENTS: Twenty-two live donors involved in adult-to-adult right lobe liver transplantation from May 1996 to June 1999. INTERVENTIONS: The right lobe grafts were obtained by transecting the liver on the left side of the middle hepatic vein. Liver transection was performed by using an ultrasonic dissector, without using the Pringle maneuver. The left lobe volume was measured by computed tomographic volumetry and the ratio of left lobe volume to the total liver volume was calculated. MAIN OUTCOME MEASURES: Hospital mortality rate and complication rate. RESULTS: The median blood loss was 719 mL (range, 200-1,600 mL). Only one donor, who had thalassemia, received 1 U of homologous blood transfusion. Postoperative complications included wound infection, incision hernia, and cholestasis in 1 donor whose liver showed 20% fatty change and who had a left lobe-total liver volume of 0.34. Another donor with 15% fatty change in the liver and a left lobe-total liver volume ratio of 0.27 developed prolonged cholestasis. Two other donors with left lobe-total liver volume ratios of 0.27 but with mild steatosis (<5%) did not develop postoperative cholestasis. Postoperative complications also included 1 case of biliary stricture and 1 case of small bowel obstruction. Both complications were adequately treated. There was no donor mortality. All donors are well and have returned to their previous occupations. CONCLUSION: Live donation of right lobe graft for adult-to-adult liver transplantation is safe provided that the residual liver volume exceeds 30% of the total liver volume and the liver itself is normal or only mildly affected by steatosis. PMID- 10722039 TI - Selective use of tube cholecystostomy with interval laparoscopic cholecystectomy in acute cholecystitis. AB - HYPOTHESIS: Tube cholecystostomy followed by interval laparoscopic cholecystectomy is a sale and efficacious treatment option in critically ill patients with acute cholecystitis. DESIGN: Retrospective cohort study within a 4 1/2%-year period. SETTING: University hospital. PATIENTS: Of 324 patients who underwent laparoscopic cholecystectomy, 65 (20%) had acute cholecystitis; 15 of these 65 patients (mean age, 75 years) underwent tube cholecystostomy. INTERVENTION: Thirteen patients at high risk for general anesthesia because of underlying medical conditions underwent percutaneous tube cholecystostomy with local anesthesia. Laparoscopic tube cholecystostomy was performed on 2 patients during attempted laparoscopic cholecystectomy because of severe inflammation. Interval laparoscopic cholecystectomy was attempted after an average of 12 weeks. MAIN OUTCOME MEASURES: Technical details and clinical outcome. RESULTS: Prompt clinical response was observed in 13 (87%) of the patients after tube cholecystostomy. Twelve patients (80%) underwent interval cholecystectomy. Laparoscopic cholecystectomy was attempted in 11 patients and was successful in 10 (91%), with 1 conversion to open cholecystectomy. One patient had interval open cholecystectomy during definitive operation for esophageal cancer and another had emergency open cholecystectomy due to tube dislodgment. Two patients (13%) had complications related to tube cholecystostomy and 2 patients died from sepsis before interval operation. One patient died from sepsis after combined esophagectomy and cholecystectomy. Postoperative minor complications developed in 2 patients. At a mean follow-up of 16.7 months (range, 0.5-53 months), all patients were free of biliary symptoms. CONCLUSIONS: Tube cholecystostomy allowed for resolution of sepsis and delay of definitive surgery in selected patients. Interval laparoscopic cholecystectomy was safely performed once sepsis and acute infection had resolved in this patient group at high risk for general anesthesia and conversion to open cholecystectomy. Just as catheter drainage of acute infection with interval appendectomy is accepted in patients with periappendiceal abscess, tube cholecystostomy with interval laparoscopic cholecystectomy should have a role in the management of selected patients with acute cholecystitis. PMID- 10722040 TI - Restorative proctocolectomy with J-pouch ileoanal anastomosis. AB - Restorative proctocolectomy with ileoanal anastomosis, complemented by a pouch formed with the last foot of terminal ileum, is the procedure of choice for patients in need of surgical treatment for ulcerative colitis and familial polyposis. The procedure has undergone many technical modifications that have ensured a very high degree of continence and an acceptable number of daily bowel movements. Herein we describe the operative technique we use in the majority of our patients, a restorative proctocolectomy with hand-sewn J-pouch ileoanal anastomosis with protecting ileostomy. We also comment on the immediate postoperative care and on the long-term functional results. PMID- 10722041 TI - Surgery in Saudi Arabia. PMID- 10722042 TI - The professor touch. PMID- 10722043 TI - On scalpels and bistouries. PMID- 10722044 TI - Enzymatic halogenation catalyzed via a catalytic triad and by oxidoreductases. AB - During the search for haloperoxidases in bacteria we detected a type of enzymes that catalyzed the peroxide-dependent halogenation of organic substrates. However, in contrast to already known haloperoxidases, these enzymes do not contain a prosthetic group or metal ions nor any other cofactor. Biochemical and molecular genetic studies revealed that they contain a catalytic triad consisting of a serine, a histidine, and an aspartate. The reaction they catalyze is actually the perhydrolysis of an acetic acid serine ester leading to the formation of peracetic acid. As a strong oxidizing agent the enzymatically formed peracetic acid can oxidize halide ions, resulting in the formation of hypohalous acid which then acts as the actual halogenating agent. Since hypohalous acid is also formed by the heme- and vanadium-containing haloperoxidases, enzymatic halogenation catalyzed by haloperoxidases and perhydrolases in general lacks substrate specificity and regioselectivity. However, detailed studies on the biosynthesis of several halometabolites led to the detection of a novel type of halogenases. These enzymes consist of a two-component system and require NADH and FAD for activity. Whereas the gene for one of the components is part of the biosynthetic cluster of the halometabolite, the second component is an enzyme which is also present in bacteria from which no halometabolites have ever been isolated, like Escherichia coli. In contrast to haloperoxidases and perhydrolases the newly detected NADH/FAD-dependent halogenases are substrate-specific and regioselective and might provide ideal tools for specific halogenation reactions. PMID- 10722045 TI - Structural analysis of modified forms of recombinant IFN-beta produced under stress-simulating conditions. AB - The present study focused on the investigation of the chemical stability of recombinant human interferon-beta (rhIFN-beta) tested in vitro by chemical treatments that simulate stress-induced conditions that may occur during handling, storage or ageing of protein samples. Mild oxidation and/or alkylation of the recombinant protein showed that the four methionines occurring in the interferon displayed different chemical susceptibility in that Met36 and Met117 were fully modified, whereas Met1 showed only little modification and Met62 was completely resistant. Moreover, incubation of rhIFN-beta under alkaline conditions resulted in the formation of a covalent dimeric species stabilised by an intermolecular disulphide bridge involving the free SH group of Cys17 from each polypeptide chain. Analysis of biological activity of the different IFN-beta derivatives showed that rhIFN-beta fully retains its specific activity following mild oxidation treatments whereas reaction with a high concentration of alkylating agents or incubation under alkaline conditions strongly reduce its specific antiviral activity. PMID- 10722046 TI - A synthetic peptide derived from the non-structural protein 3 of hepatitis C virus serves as a specific substrate for PKC. AB - A synthetic peptide corresponding to the amino acid sequence Arg1487-Arg-Gly-Arg Thr-Gly-Arg-Gly-Arg-Arg-Gly-Ile-Tyr-Arg1500 of the hepatitis C virus (HCV) polyprotein was found to be a selective substrate for protein kinase C (PKC). In the presence of Ca2+, TPA and phospholipid, PKC phosphorylates the peptide [termed HCV(1487-1500)] with a Km of 11 microM and Vmax of 24 micromol x min(-1) x mg(-1). HCV(1487-1500) acts as a competitive inhibitor of PKC towards other peptide or protein substrates and inhibits the kinase activity with an IC50 corresponding to the Km values measured for the substrates. N- or C-terminally deleted analogs of HCV(1487-1500) did not show inhibitory effects and were only marginally or not phosphorylatable. We designed an additional peptide in which the tyrosine residue was replaced by phenylalanine ([Phe1499]HCV(1487-1500)). This peptide was neither phosphorylated by other serine/threonine kinases tested nor by whole cell extracts prepared from PKC-depleted cells. [Phe1499]HCV(1487 1500) was used to monitor the TPA-induced translocation of PKC activity to the particulate fraction in JB6 cells. The use of SDS/PAGE to separate the peptide from ATP and Pi allowed to monitor simultaneously PKC autophosphorylation and phosphorylation of the peptide. The data presented here show that[Phe1499]HCV(1487-1500) can serve as a convenient tool for investigations of PKC activity also in the presence of other kinases in tissues or in crude cell extracts. PMID- 10722047 TI - Neoglycolipids derived from phosphatidylethanolamine serve as probes in cell culture studies on glycolipid metabolism. AB - The neoglycolipid (NeoGL) N-acetyl-1-deoxy-1-phosphatidylethanolamino lacto-N tetraositol [Lc4Ose-PtdEtn(NAc)] and the radioactivly labeled analog [Lc4Ose PtdEtn(N[14C]Ac)] were synthesized by coupling the corresponding oligosaccharide to phosphatidylethanolamine (dihexadecyl) via reductive amination and subsequent N-acetylation with unlabeled and [14C]acetic acid anhydride, respectively. Lc4Ose PtdEtn(N[14C]Ac) was then incubated with homogenates of rat small intestine epithelial cells (IEC-6) at pH 4. The reaction products were shown to be the degradation products formed by glycosidases by fast atom bombardment mass spectrometry (FAB MS). On the other hand, incubation of Lc4Ose-PtdEtn(NAc) with IEC-6 cell homogenates in sialyltransferase assays yielded the corresponding sialylated product. When Lc4Ose-PtdEtn(N[14C]Ac) was fed to IEC-6 cells as BSA complex, up to 5% of the NeoGL administered were taken up by the cells. After extraction of the NeoGL and separation by thin layer chromatography (TLC) the catabolic products Lc3Ose-PtdEtn(N[14C]Ac), Lac-PtdEtn(N[14C]Ac), and Glc PtdEtn(N[14C]Ac), as well as the main anabolic product NeuGc-Lc4Ose PtdEtn(N[14C]Ac) could be identified by FAB MS. These results demonstrate that PtdEtn-derived NeoGL can be used as probes for studies on the metabolism of specific oligosaccharide structures in cell culture. PMID- 10722048 TI - Anomalous binding of MgADP to myosin of skeletal muscle. AB - Binding of adenosine diphosphate to skeletal muscle myosin was studied using a range of concentrations from 0 to 2 mM. Up to 0.2 mM adenosine diphosphate two equivalent and independent nucleotide binding sites were detected, characterized by the single association constant of 5 x 10(4)M(-1). At greater adenosine diphosphate concentrations a decreasing binding capacity was noticed, bound nucleotide being essentially approximately 0.1 mol/mol at a 1-2mM adenosine diphosphate concentration. We tentatively propose that nucleotides act indirectly on myosin by promoting the perturbation of the solvent, which is supported by the fact that polyphosphates are known powerful kosmotropes. PMID- 10722050 TI - Activation of progelatinase B by membranes of human polymorphonuclear granulocytes. AB - Isolated human granulocyte plasma membranes contain progelatinase B. The binding of progelatinase B to the membrane, however, is relatively weak, and a considerable part of progelatinase B can be removed by simply washing the membrane with buffer. This detachment does not depend on the ionic strength of the buffer, indicating that electrostatic forces do not play an important role in the binding of progelatinase B to the membrane. A complete removal of progelatinase B is achieved by chromatography of neutrophil membranes on gelatin agarose. The plasma membrane of human granulocytes activates added progelatinase B. This activation is inhibited by soybean trypsin inhibitor and is thus performed by membrane bound serine proteinases. In contrast to other reports that claimed an important role of elastase in activating progelatinase B, we found that this activation is mostly inhibited by chymostatin and not by elastatinal and is thus primarily due to cathepsin G. Proteinase 3 was shown to activate progelatinase B as efficient as neutrophil elastase, i. e. much weaker than cathepsin G. Binding of cathepsin G and elastase to the neutrophil membrane does not change their ability to activate progelatinase B. However, cathepsin G, the most potent activator of the three neutrophil serine proteinases, is only a weak activator, when compared to stromelysin-1. This, as well as only a weak binding of progelatinase B, make it doubtful that activation of membrane-bound progelatinase B by membrane-bound serine proteinases is of significant physiological importance. PMID- 10722049 TI - Engineering, purification and applications of His-tagged recombinant antibody fragments with specificity for the major birch pollen allergen, bet v1. AB - Type I allergy, an immunodisorder affecting almost 20% of the population worldwide, is based on the production of IgE antibodies against per se harmless allergens. We report the expression of hexahistidine-tagged antibody fragments (Fabs) with specificity for Bet v1, the major birch pollen allergen, in Escherichia coli. The cDNA coding for the heavy chain fragment of a mouse monoclonal anti-Bet v1 antibody, Bip 1, was engineered by PCR to contain a hexahistidine-encoding 3' end. The modified Bip1 heavy chain cDNA was co expressed in E. coli XL-1 Blue with the Bip 1 light chain cDNA using the combinatorial plasmid pComb3H. His-tagged recombinant (r) Bip 1 Fabs were isolated by nickel affinity chromatography and rBip 1 Fabs without His-tag were purified via affinity to rBet v1. rBip 1 Fabs with and without His-tag bound specifically to rBet v1 and, like Bet v1 -specific human serum IgE and rabbit anti rBet v1 antibodies, cross-reacted with Bet v1-related allergens in other plant-species (alder, oak, hazelnut). We demonstrate the usefulness of His-tagged rBip 1 Fabs (1) for the identification of pollen samples containing Bet v 1 by particle blotting, (2) forthe detection of Bet v1-specific IgE antibodies in human serum samples by sandwich ELISA and (3) for the quantification of Bet v1 in solution. Based on these examples we suggest to use rBip 1 Fabs for the detection of Bet v1 and Bet v1-related allergens in natural allergen sources for allergy prevention, as well as for the standardization of natural allergen extracts produced for diagnosis and immunotherapy of birch pollen allergy. PMID- 10722051 TI - Bait region cleavage and complex formation of human alpha2M with a Porphyromonas gingivalis W50 protease is not accompanied by enzyme inhibition. AB - Three isoforms of extracellular Arg-specific proteases of P. gingivalis, W50, HRgpA, RgpAcat and mt-RgpAcat, which are all products of the same gene, show identical enzymatic properties toward small chromogenic substrates but have different subunit organisation and molecular size. In order to examine the potential inhibition of these proteases in vivo by host protease inhibitors, the interaction of HRgpA (approximately 110 kDa) and RgpAcat (approximately 55 kDa) with human (alpha2M and their cytotoxicity toward cultured fibroblasts were investigated. Both enzymes formed complexes with (alpha2M as shown by gel filtration chromatography and both cleaved the 'bait' region at Arg696-Leu697. However, whereas (alpha2M-RgpAcat) complex was unable to hydrolyse large substrates such as hide powder azure, (alpha2M-HRgpA) complex hydrolysed both small and large substrates. HRgpA was able to bind to alpha2M saturated with trypsin and also to methylamine-treated alpha2M. This suggested that HRgpA is able to bind to both 'slow' and 'fast' forms of alpha2M and formation of (alpha2M:HRgpA) complex does not trap HRgpA and cause inhibition of activity toward hide powder azure. However, the (alpha2M-HRgpA) complex is not able to cleave other alpha2M molecules, which suggests that the active site of HRgpA in the complex is constrained probably due to steric reasons. The (alpha2M-HRgpA) complex was cytotoxic to 3T3 cells, causing them to round up and detach from the surface with a reduction in metabolic activity. PMID- 10722052 TI - Adjuvant and haemolytic activities of 47 saponins derived from medicinal and food plants. AB - Adjuvant and haemolytic activities of 47 saponins purified from medicinal and food plants were examined. The compounds showed various levels of both adjuvant and haemolytic activities. Soyasaponins and lablabosides showed strong adjuvant activity but little haemolytic activity. Jujubosides showed strong adjuvant and haemolytic activities. Escins showed weaker adjuvant activity than the adjuvant control, but strong haemolytic activity. Comparison of the functional groups of each saponin revealed that the acyl residue in saponin, the aldehyde group at carbon 4 in aglycone, and branched sugar chains attached to aglycone, were not essential for adjuvant activity. Furthermore, saponins with an acyl residue or oxide-ring moiety tended to show haemolytic activity. These results suggest that the adjuvant activity of saponins does not relate with haemolytic activity. It is considered that not only the functional groups themselves, but the overall conformation harmoniously consisting of such functional groups, affects adjuvant activity of saponins. PMID- 10722054 TI - Investigation of the effect of freezing on protease-catalyzed peptide synthesis using cryoprotectants and frozen organic solvent. AB - In order to investigate the effect of freezing on aqueous protease-catalyzed peptide synthesis systems, the influence of polyethylene glycols as cryoprotecting substances on alpha-chymotrypsin-catalyzed coupling of a N protected acyl donor ester and various nucleophilic amino components was studied. Changes in S'-specificity of alpha-chymotrypsin in frozen aqueous systems were suppressed by polyethylene glycols even at concentrations below 1% (w/v). Furthermore, the influence of freeze-concentration in organic solvents on protease-catalyzed peptide synthesis was investigated for the first time. In frozen tert-butanol, alpha-chymotrypsin-catalyzed peptide synthesis took advantage from freeze-concentration, but in contrast to frozen aqueous systems, no changes in S'-specificity of the biocatalyst were observed. The results suggest that freeze-concentration is not the only cause of freezing-induced yield improvement in aqueous peptide synthesis systems, but interactions between enzyme and ice structures strongly contribute to the observed effects. PMID- 10722053 TI - Tissue-specific regulation of IRS-2/PI 3-kinase association in aged rats. AB - We have examined the insulin-stimulated IRS-2 association with PI 3-kinase and the phosphorylation of AKT/PKB, which is functionally located downstream of the PI 3-kinase, in aged (obese) rats. The IRS-2 protein levels were similar in 2 and 20 month-old rats in both tissues, liver and muscle. There were reductions in insulin-induced IRS-2 tyrosine phosphorylation in liver and muscle, accompanied by a decrease in IRS-2/PI 3-kinase association and in AKT/PKB phosphorylation only in muscle tissue of aged rats. This regulation may be important in the altered glucose metabolism observed in aged (obese) rats. PMID- 10722055 TI - Cloning and expression of functional equistatin. AB - Equistatin is a 199-residue protein composed of three thyroglobulin type-1 domains. It strongly inhibits cysteine proteinases as well as the aspartic proteinase cathepsin D. In order to initiate structure-function studies by protein engineering, a cDNA library from sea anemone, Actinia equina, was screened. A positive clone of 888 nucleotides was shown to encode a protein of 231 amino acids, including the signal sequence. The mature protein region was amplified by PCR, cloned into the pET22b(+)cas expression vector and expressed in Escherichia coli. Isolation of active recombinant equistatin required only one purification step, the His-tag affinity column. The protein displays physical and inhibitory properties closely similar to the native inhibitor. PMID- 10722056 TI - Micellar liquid chromatography for prediction of drug transport. AB - The vast majority of well absorbed drugs are transported passively across the cell membranes. Physicochemical descriptors of drug molecules that are believed to influence transcellular transport are routinely used to predict drug absorption by means of complex mathematical models. In this paper, a new in vitro method, based on the retention data in micellar liquid chromatography (MLC), is validated for the prediction of passive drug absorption. The retention of a heterogeneous drugs set in MLC using Brij 35 as surfactant in the mobile phase is compared with the retention data reported in literature obtained in red cell membrane lipid liposomes, human red cell membranes vesicles (vesicles), native membranes of adsorbed red cells (ghosts) and egg phospholipids liposomes [Beigi et al., Int. J. Pharm., 164 (1998) 129-137]. Finally, the correlation between the logarithm of retention factors in MLC and reported oral drug absorption values for barbiturates and beta-blockers is studied. Predictive regression models for estimating oral drug absorption using the logarithm of the retention values as independent variable are proposed. PMID- 10722057 TI - Determination of N-acetylcysteine in human plasma by liquid chromatography coupled to tandem mass spectrometry. AB - An analytical method for the determination of total N-acetylcysteine in human plasma has been developed, validated and applied to the analysis of samples from a phase I clinical trial. The analytical method consists of plasma digestion with dithiothreitol in order to reduce all the oxidized forms of N-acetylcysteine, and extraction with ethyl acetate followed by determination of levels by an LC-MS-MS method. The intra- and inter-assay precision and accuracy of this technique were good and the limit of quantitation was 50 ng/ml of plasma. The concentration working range was established between 50 ng/ml and 1000 ng/ml. This method has been used in the analysis of approximately 800 human plasma samples from a clinical study with 24 volunteers; the precision of the quality controls was in the range 8.7 to 13.4% and the accuracy was in the range -5.9 to 8.5%, expressed as the RSD and the relative error, respectively. PMID- 10722058 TI - Comparison of various reversed-phase columns for the simultaneous determination of ephedrines in urine by high-performance liquid chromatography. AB - Different reversed-phase columns for basic analytes were compared for the simultaneous determination of ephedrines in urine, such as LiChrospher 60 RP Select B, LiChrospher 100 RP18, Hypersil BDS-C18, Inertsil ODS-2, Spherisorb ODS B and Symmetry Shield RP8. Symmetry Shield was the only column which did not require the use of high concentrations of buffer and triethylamine. With this column, a good separation of the six ephedrines and the internal standard was achieved using 50 mM phosphate buffer-25 mM triethylamine as a mobile phase. Linearity, precision and accuracy were satisfactory for the levels usually found in urine. Due to these all parameters the developed analytical method was found to be suitable for the application in the doping field. PMID- 10722060 TI - Analysis of barbiturates by micro-high-performance liquid chromatography with post-column photochemical derivatization. AB - This study attempts to combine the advantages of microcolumn high-performance liquid chromatography (micro-HPLC) with those of post-column photochemical derivatization in barbiturate analysis. Some barbiturates are photochemically unstable, leading to photoproducts that show maximum absorption at 270 nm and not the typical one at approximately 220 nm. For this purpose, a laboratory-built photoreactor has been developed to work with micro-HPLC instruments. Its performance is satisfactory in the forensic analysis of barbiturates. PMID- 10722059 TI - Application of liquid separation techniques to the determination of the main urinary nicotine metabolites. AB - A rapid procedure for the analysis of the main nicotine metabolites (cotinine, trans-3'-hydroxycotinine) in urine has been worked out. The procedure includes isolation of nicotine and its metabolites from urine by means solid-liquid extraction technique using resin Amberlite XAD-2 and then quantitation by the use of thin-layer chromatography with densitometry (in reflection mode). GC-MS was applied to confirm the results obtained by TLC. The procedure was applied to the analysis of cotinine concentrations in urine samples taken from children living in Upper Silesia region (Poland). Among 444 investigated children we did not find cotinine almost in 60% but in 15% of this population, there were children who could have been exposed to cigarette smoke. PMID- 10722061 TI - Full and fractionated experimental designs for robustness testing in the high performance liquid chromatographic analysis of codeine phosphate, pseudoephedrine hydrochloride and chlorpheniramine maleate in a pharmaceutical preparation. AB - This paper describes the testing of a saturated factorial design using a full factorial design. Saturated factorial designs are often used to test the robustness of high-performance liquid chromatography (HPLC) methods, however they are based on several assumptions. A full factorial design relies on fewer assumptions and hence could be used to evaluate the effectiveness of the saturated design. Both designs were used to test a gradient HPLC method for the assay of codeine phosphate, pseudoephedrine hydrochloride and chlorpheniramine maleate. Six HPLC conditions, including wavelength, mobile phase pH and ion pairing reagent concentration were tested using the saturated design. Three of these factors were selected for full evaluation using a full factorial design. The results showed that the main effects calculated by each design were comparable. However, the saturated design showed higher standard errors, probably due to the effects of changing several more factors. One interaction effect was indicated as a confounding effect by the saturated design and this was confirmed by the calculation of the same interaction effect using the full design. Overall the method was shown to be robust under the variety of HPLC conditions tested. PMID- 10722062 TI - Profiling of impurities in illicit methamphetamine by high-performance liquid chromatography and capillary electrochromatography. AB - High performance liquid chromatography (HPLC) with photodiode array (PDA) UV and fluorescence (FL) detection, and capillary electrochromatography (CEC) with laser induced fluorescence (LIF) detection were investigated for the analysis of acidic extracts derived from illicit methamphetamine. These compounds include major impurities from the hydriodic acid/red phosphorous reduction method, i.e., 1,3 dimethyl-2-phenylnaphthalene and 1-benzyl-3-methylnaphthalene, and other trace level, structurally related impurities. For certain of these solutes, HPLC with conventional FL detection gave at least a 60x increase in sensitivity over UV detection. In addition, other highly fluorescent impurities were detected in methamphetamine produced via four other synthetic routes. The use of a rapid scanning FL detector (with acquisition of "on the fly" excitation or emission) provided structural information and gave "optimum" excitation and emission detection wavelengths. CEC with LIF detection using UV laser excitation provided greatly improved chromatography over HPLC, with good detection limits in the low ng/ml range. Both methodologies provide good run-to-run repeatability, and have the capability to distinguish between samples. PMID- 10722063 TI - Validation of the removal of acetylsalicylic acid. Recovery and determination of residues on various surfaces by high performance liquid chromatographic. AB - The validation of a procedure to clean glass, vinyl and stainless steel surfaces that have been exposed to acetylsalicylic acid during its manufacture is described. The cleaning procedure using two cotton swabs moistened with the mobile phase was validated using a wipe-test and a high-performance liquid chromatography (HPLC) method developed to determine low quantities of the acid. The HPLC method involves an octadecylsilane column at 55 degrees C, a mixture of water-acetonitrile-orthophosphoric acid (779:220:1, v/v) as mobile phase and detection at 226 nm. Recoveries of 86%, 90% and 94% were obtained from vinyl, glass and stainless steel plates respectively. The validation gave acceptable levels of sensitivity, recovery, precision and linearity. PMID- 10722064 TI - Simultaneous determination of paracetamol and chlorpheniramine in human plasma by liquid chromatography-tandem mass spectrometry. AB - An analytical method for the determination of paracetamol and chlorpheniramine in human plasma has been developed, validated and applied to the analysis of samples from a phase I clinical trial. The analytical method consists in the extraction of paracetamol and chlorpheniramine with diethyl ether, followed by the determination of both drugs by an LC-MS-MS method, using 2-acetamidophenol as internal standard. The intra-assay and inter-assay precision and accuracy of this technique were good and the limit of quantitation was 0.5 microg/ml of plasma for paracetamol and 0.2 ng/ml for chlorpheniramine. The concentration working range was established between 0.5 microg/ml and 25 microg/ml for paracetamol and between 0.2 ng/ml and 50 ng/ml for chlorpheniramine. This method has been used for analyzing more than 1200 human plasma samples from a clinical study with 24 volunteers. PMID- 10722065 TI - Determination of theophylline and its metabolites in biological samples by liquid chromatography-mass spectrometry. AB - Liquid chromatography-mass spectrometry (LC-MS) is a powerful tool for analysis of drugs and their metabolites. We used a column-switching system in combination with atmospheric pressure chemical ionization LC-MS (LC-APCI-MS) for the determination of theophylline and its metabolites in biological samples. The separation was carried out on a reversed-phase column using methanol-20 mM ammonium acetate as a mobile phase at a flow-rate of 1 ml/min in 30 min. In the mass spectrum, the molecular ions of these drugs and metabolites were clearly observed as base peaks. This method is sufficiently sensitive and accurate for the pharmacokinetic studies of these drugs. PMID- 10722066 TI - Optimization and validation of a method for the determination of caffeine, 8 chlorotheophylline and diphenhydramine by isocratic high-performance liquid chromatography. Stress test for stability evaluation. AB - The optimization of a HPLC method for caffeine, 8-chlorotheophylline and diphenhydramine separation with UV detection at 229 nm is described. The conditions studied included: stationary phase, compositions of mobile phases with pH modulators. Optimal conditions were: SymmetryShield RP8 column and acetonitrile-(0.01 M H3PO4-triethylamine, pH 2.8) (22:78, v/v). Validation was performed using standards and a pharmaceutical preparation containing the compounds described above. Results from both standards and samples show suitable validation parameters. The pharmaceutical grade substances were tested by factors that could influence the chemical stability. These reaction mixtures were analyzed to evaluate the capability of the method to separate degradation products. Degradation products did not interfere with the determination of the substances tested by the assay. PMID- 10722067 TI - High-performance liquid chromatography with amperometric detection applied to the screening of 1,4-dihydropyridines in human plasma. AB - A high-performance liquid chromatographic method with electrochemical detection has been developed for the determination of six 1,4-dihydropyridines: nifedipine, nimodipine, nisoldipine, nicardipine, felodipine and lacidipine. The chromatographic separation was performed using a Supelcosil LC-ABZ+Plus C18 column. A mobile phase of methanol-water (70:30), containing 2 mM CH3COOH CH3COONa at a flow-rate of 1 ml/min and a pH of 5.0, was used. The temperature was optimized at 30+/-0.2 degrees C. The amperometric detector, equipped with a glassy carbon electrode, was operated at 1000 mV versus Ag/AgCl in the direct current mode. The method was applied to the determination of these compounds at ng/ml concentrations, obtaining intra-day reproducibilities of lower than 5.0% in terms of relative standard deviations and detection limits ranging from 16 to 44 ng/ml. The method was applied to the screening of 1,4-dihydropyridines in spiked plasma samples, with a total elution time of lower than 18 min, obtaining the best recoveries for nimodipine and felodipine (91 and 88%, respectively). These recoveries together with the low detection limits achieved allow its application to the analysis of these drugs in human plasma. PMID- 10722068 TI - Quantitation of nifedipine in human plasma by on-line solid-phase extraction and high-performance liquid chromatography. AB - An analytical methodology for nifedipine quantitation in plasma by on-line solid phase extraction (SPE) and high-performance liquid chromatography (HPLC) is described. The SPE cartridges contain C2 and the analytes nifedipine and nitrendipine (internal standard) are separated on a C18 column with a mobile phase consisting of acetonitrile-13 mM phosphate buffer pH 7 (65:35, v/v) followed by UV detection at 338 nm. Validation of the method demonstrated good recoveries (>90%), sensitivity (limit of quantification, 2 ng/ml), based on a 500 microl sample volume, accuracy and precision (<5.5% in concentrations greater than the limit of quantitation). This methodology has been used for bioequivalence studies. PMID- 10722070 TI - Direct identification and quantitation of prednisone in the presence of overlapping hydrocortisone by liquid chromatography with electrospray and atmospheric-pressure chemical-ionisation mass spectrometry. AB - The paper describes the application of liquid chromatography interfaced to a triple quadrupole mass spectrometer utilising the multiple reaction monitoring (MRM) mode. The technique was shown to provide detection limits lower than 0.01% for the analysis of prednisone in the presence of hydrocortisone. Prednisone was mixed in concentrations from 0.500 to 0.0005 ppm (corresponding to 1% to 0.001% of the hydrocortisone concentration). These solutions were assayed using MRM observing the product ion transitions of 359.2-->147.1 and 359.2-->171.2 and was shown to be capable of detecting co-eluting impurities at concentrations of less than 0.001% of the major component. The assay of prednisone was shown to be linear over the range 0.500-0.0005 ppm with a correlation coefficient of 0.999 and a precision of 6.9% at the concentration of 0.005 ppm. The analysis was carried out using both atmospheric pressure chemical ionisation (APCI) and electrospray ionisation (ESI) as an interface. However, for these compounds APCI provided significantly more sensitive data compared to ESI. PMID- 10722069 TI - Determination of albendazole and its main metabolites in ovine plasma by liquid chromatography with dialysis as an integrated sample preparation technique. AB - Albendazole is a benzimidazole derivative with a broad-spectrum activity against human and animal helminth parasites. In order to determine the main pharmacokinetic parameters in sheep after oral and intravenous administration of a new formulation of albendazole (an aqueous solution), a fully automated method was developed for the determination of this drug and its main metabolites, albendazole sulfoxide (active metabolite) and sulfone in ovine plasma. This method involves dialysis as purification step, followed by enrichment of the dialysate on a precolumn and liquid chromatography (LC). All sample handling operations were executed automatically by means of an ASTED XL system. After conditioning of the trace enrichment column (TEC) packed with octadecyl silica with pH 6.0 phosphate buffer containing sodium azide, the plasma sample, in which a protein releasing reagent (1 M HCl) containing Triton X-100 was automatically added, was loaded in the donor channel and dialysed on a cellulose acetate membrane in the static-pulsed mode. The dialysis liquid consisted of pH 2.5 phosphate buffer. By rotation of a switching valve, the analytes were eluted from the TEC in the back-flush mode by the LC mobile phase and transferred to the analytical column, packed with octyl silica. The chromatographic separation was performed at 35 degrees C and the analytes were monitored photometrically at 295 nm. Due to the differences in hydrophobic character between albendazole and its metabolites, a gradient elution was applied. The mobile phase consisted of a mixture of acetonitrile and pH 6.0 phosphate buffer. The proportion of organic modifier was increased from 10.0 to 50.1% in 12.30 min, then from 50.1 to 66.9% in 1.70 min. First, the gradient conditions and the temperature were optimised for the LC separation using the DryLab software. Then, the influence of some parameters of the dialysis process on analyte recovery was investigated. Finally, the method developed was validated. The mean recoveries for albendazole and its metabolites were about 70 and 65%, respectively. The limits of quantification for albendazole and its metabolites were 10 and 7.5 ng/ml, respectively. PMID- 10722071 TI - Determination by high-performance liquid chromatography of ketoprofen in vitro in rat skin permeation samples. AB - A direct, simple and rapid high-performance liquid chromatographic method has been developed for the determination of ketoprofen with ibuprofen as internal standard. Samples were chromatographed on a 5 microm Kromasil 100 C18 column. The mobile phase was a mixture of acetonitrile-0.01 M KH2PO4 adjusted to pH 1.5 with orthophosphoric acid 85% (60:40, v/v). Detection was at 260 nm and the run time was 10 min. The detector response was found to be linear in the concentration range 0.02 to 40 microg/ml. This HPLC assay has been applied to measure the "in vitro" percutaneous penetration of ketoprofen through rat skin. PMID- 10722072 TI - Automated liquid membrane extraction for high-performance liquid chromatography of Ropivacaine metabolites in urine. AB - An automatic method for the determination of metabolites of Ropivacaine in urine was set up. It utilizes supported liquid membrane extraction for sample clean-up and enrichment, followed by ion-pair chromatography determination using UV detection. The extraction was very selective with no observed interfering compounds from the urine matrix, permitting simple isocratic chromatographic analysis. The detection limits for spiked urine samples were 2-18 nM for the different compounds. The repeatability was 1-3% (RSD) with an internal standard that was also extracted, and about twice without this standard. A throughput of 3.3 samples per hour was achieved and the liquid membrane was stable for more than a week. PMID- 10722073 TI - Development of a sensitive method for the determination of ganciclovir by reversed-phase high-performance liquid chromatography. AB - Ganciclovir is a nucleoside analogue widely used in the treatment of cytomegalovirus infections, which affects mainly immunocompromised patients. Recently, new pharmaceutical dosage forms based on the use of albumin nanoparticles have been developed for improving the efficacy of this drug. The aim of this study was to develop an analytical HPLC method for the determination of ganciclovir in both pharmaceuticals (i.e. albumin nanoparticles) and biological medium samples. The chromatography was performed on a reversed-phase encapped column (LiChrospher Select B C8) with a mobile phase consisting of acetonitrile in 0.05 M ammonium acetate (pH 6.5; 2: 98, v/v). Acyclovir was used as internal standard and the detection wavelength was 254 nm. The limit of quantitation of ganciclovir was 50 ng/ml and the average recoveries over a concentration range of 0.05-10 microg/ml ranged from 98 to 102%. Precision did not exceed 5%. In summary, this assay is a selective, sensitive and reproducible method for the determination of the ganciclovir in albumin nanoparticles. It can be successfully applied to the estimation of the ganciclovir uptake by cultured human corneal fibroblasts. PMID- 10722074 TI - Simultaneous determination of sulfaquinoxaline, sulfamethazine and pyrimethamine by liquid chromatography. AB - A liquid chromatography method is described to determine sulfaquinoxaline (SQX), sulfamethazine (SMT), and pyrimethamine (PMT), by using a Kromasil C18 column and a 40 mM NaH2PO4 buffer solution, containing 10 mM NaClO4 (pH 3.0)-acetonitrile (65:35) as mobile phase. The mobile phase flow-rate and sample volume injected were 1.5 ml/min and 20 microl, respectively and the samples were dissolved in the mobile phase. The limits of quantification were found to be about 180 microg/l (3.6 ng) for each compound. The method was applied in veterinary commercial formulations. Analyses were made by means of the standard addition method, whose results were compared with those obtained by preparing "tests" (from the stock solutions) and with those obtained by a capillary electrophoresis method. Both methods showed similar results, and then it was proved that some commercial claimed levels were not in agreement with the obtained results by using our analytical method, as they were in other cases. PMID- 10722075 TI - Stability-indicating high-performance liquid chromatographic assay and analysis of decomposition products of 2-[4-[(7-chloro-2-quinoxalinyl)oxy]phenoxy]propanoic acid. AB - A stability-indicating HPLC assay has been developed for 2-[4-[(7-chloro-2 quinoxalinyl)oxy]phenoxy]propanoic acid (XK469). XK-469 is the 7-chloro analog of herbicide Quizalofop and is currently under development as an antineoplastic agent. HPLC separation of XK469 is achieved with an ODS column using isocratic elution of an aqueous MeOH mobile phase. The assay is reproducible (RSD=0.9%), linear (r2=0.999), accurate (error=1.2%) and sensitive (LDL=1.2 ng). The HPLC separates XK469 from its forced decomposition products. Identities of the decomposition products have been elucidated. PMID- 10722076 TI - Determination of the N-methyl-D-aspartate receptor NR2B subunit blocker Ro 63 1908 in rat, cynomolgus monkey and human plasma by high-performance liquid chromatography with column switching and fluorescence detection. AB - A HPLC method with automated column switching was developed and validated for the determination of Ro 63-1908 in rat and cynomolgus monkey plasma. Human plasma was used for calibration and was also included in the validation process. Ro 63-1908 belongs to a class of neuroprotective N-methyl-D-aspartate (NMDA) receptor blockers which were in development for the treatment of stroke and traumatic brain injury. The method involves deproteinisation of plasma samples with ethanol and direct injection of the supernatant (1.4 ml) into the HPLC column-switching system. To prevent a breakthrough of the analyte and the internal standard on the precolumn (Purospher RP-18, 75x4 mm) due to the high ethanol content, the injection solution was diluted, on-line, using an additional pump and a T-piece. 1% ammonium acetate-ethanol (100:2, v/v) was used as mobile phase for injection, as well as for on-line dilution, resulting in pre-concentration of the analyte and the internal standard on the precolumn. As Purospher RP-18 is a non-endcapped stationary phase with a special selectivity for amines, the analyte and the internal standard could then be selectively eluted with 30% acetonitrile (without any buffer in the mobile phase) and transferred to the analytical column [consisting of two coupled columns (125+250x4 mm) packed with Superspher 60 RP select B], where they were separated by gradient elution and detected by fluorescence detection. Compared to the use of a 125 mm long precolumn and dilution of the supernatant with ammonium acetate prior to injection, the 75 mm precolumn and the on-line dilution procedure allowed about one third shorter run times (21 min) and, therefore, a higher sample throughput. The limit of quantification was 1 ng/ml using 0.4 ml plasma. The method was applied to more than 670 plasma samples from pharmacokinetic and toxicokinetic studies and is also suitable for other matrices and NMDA receptor blockers. PMID- 10722077 TI - Determination of sunscreen agents in cosmetic products by supercritical fluid extraction and high-performance liquid chromatography. AB - A rapid supercritical fluid extraction (SFE) procedure for the isolation of five of the most common sunscreen agents (2-ethylhexyl-p-dimethylaminobenzoate, 2 hydroxy-4-methoxybenzophenone, 2-ethylhexyl-p-methoxycinnamate, 4 methylbenzylidene camphor and 4-tert.-butyl-4'-methoxydibenzoylmethane) from cosmetic products is described. Investigation of the factors affecting the extraction efficiency in SFE indicated that sunscreen recoveries were affected mainly by the supercritical CO2 pressure and by the trapping method. The sunscreens were analyzed by reversed-phase high-performance liquid chromatography after a 10-min extraction of the cosmetic product with CO2 at 250 bar and 40 degrees C, using sequential glass surface and C18 sorbent as collection system. A quantitative comparison of SFE with a liquid extraction procedure was performed on commercial cosmetics. The SFE method yielded recoveries higher than 94.8% compared with conventional liquid extraction and exhibited a precision better than 5.3% relative standard deviation. Moreover, SFE minimized sample handling, reduced the consumption of harmful solvents and afforded a more effective purification of the cosmetic matrices. PMID- 10722078 TI - Determination of eight water- and fat-soluble vitamins in multi-vitamin pharmaceutical formulations by high-performance liquid chromatography. AB - In the present work, a reversed-phase high-performance liquid chromatographic procedure has been developed for the determination of water-soluble vitamins (thiamine hydrochloride, pyridoxine hydrochloride, nicotinamide, riboflavin phosphoric ester and cyanocobalamine) and fat-soluble vitamins (retinol palmitate, cholecalciferol, alpha-tocopherol acetate) in multi-vitamin pharmaceutical formulations. The sample treatment proposed consists of a solid phase extraction with C18 AR cartridges that allow the separation of fat-soluble vitamins, which were retained on the sorbent, from water-soluble vitamins. Afterwards, the water-soluble vitamins were analysed by HPLC on a Nova-Pack C18 (150x3.9 mm, 4 microm) analytical column, using CH3OH-0.05 M CH3COONH4 as mobile phase. The chromatographic analysis of the fat-soluble vitamins was carried out after their sequential elution with methanol and chloroform from C18 sorbent, on the above column. The mobile phase employed was MeOH-CH2CN (95:5, v/v) working at a flow-rate of 2 ml min(-1) in isocratic mode. The solid-phase extraction for these vitamins had been previously optimised. The experimental variables studied were: application volume, elution solvents and cleaning solutions. The UV-Vis detection of vitamins was made at 270 nm for all the water-soluble vitamins (362 nm for B12) and 285 nm for the water-soluble and fat-soluble vitamins present in real samples at different concentration levels. The accuracy of the method was tested obtaining an average recovery ranging between 78 and 116%. PMID- 10722079 TI - Determination of vitamin B12 in multivitamin tablets by multimode high performance liquid chromatography. AB - Quantitative determination of vitamin B12 in B-complex tablets was performed by using multimode high-performance liquid chromatography. The multivitamin tablets (B1, B6 and B12) were sonicated for 30 min in methanol-water (50:50, v/v) and diluted to appropriated volume with the same solvent. The resulting solution was filtered and the filtrate was analysed on a phenylpropanolamine bonded silica column (15 cm x 4.6 mm I.D., 5 microm). The optimized mobile phase was 30 mM phosphate buffer (pH 3.00) containing 6% (v/v) acetonitrile at a flow-rate of 1 ml min(-1) and the detection was measured at 361 nm. The calibration graph prepared using standards was linear from 0.05 to 0.25 microg. The determination limit was 25 ng, the relative standard deviation was 0.47% and recovery from tablet solution was 100%. An analysis was completed in 5 min. The new method is simple, rapid and precise. PMID- 10722080 TI - Measurement and pharmacokinetics of unbound 20(S)-camptothecin in rat blood and brain by microdialysis coupled to microbore liquid chromatography with fluorescence detection. AB - To characterize the pharmacokinetics of protein-free camptothecin in blood and brain we implanted microdialysis probes into the jugular vein and striatum of rats for unbound drug sampling and determination. Camptothecin (2 or 5 mg/kg, i.v., n=6) was then administered from the femoral vein, and microdialysates were collected from blood and brain of both sites and assayed by a validated microbore scale high-performance liquid chromatographic method. The mobile phase consisted of methanol-100 mM monosodium phosphoric acid (35:65, v/v, pH 2.5) with a flow rate 0.05 ml/min. The fluorescence response for camptothecin was observed at excitation and emission wavelengths of 360 and 440 nm, respectively. Pharmacokinetic parameters were calculated from the corrected data for dialysate concentrations of camptothecin versus time. The results suggest that the pharmacokinetics of unbound camptothecin in blood and brain can be fitted best to a two- and one-compartment model, respectively. Camptothecin rapidly entered the extracellular fluid of brain striatum at 10 min following camptothecin administration. PMID- 10722081 TI - Separation of erythromycin and related substances on base-deactivated reversed phase silica gel columns. AB - An official liquid chromatographic method for the analysis of erythromycin and related substances, which is based on a polymer reversed-phase, is described in the European Pharmacopoeia and in the United States Pharmacopeia. The pH of the mobile phase used in this system is 9.0. Recent advanced technology has led to the introduction of a new generation of silica-based reversed-phase column packings, which are claimed to be much more stable towards bases. They are useful for the analysis of basic compounds. Studies to verify the separation of erythromycin and related substances on Hypersil BDS C18, Luna C18(2), Inertsil ODS-2 and Supelcosil ABZ+ have been performed and the results are presented. It is shown that these base-deactivated phases give a better sensitivity and selectivity towards erythromycins than the polymer phase, provided that an adapted mobile phase is used. This is the first liquid chromatographic method described for the separation of erythromycin D from erythromycin A. PMID- 10722082 TI - Liquid chromatography of polymyxin B sulphate. AB - A reversed-phase liquid chromatography method for analysis of polymyxin B sulphate is described. The method uses a YMC-Pack Pro, C18, 5 microm, 250x4.6 mm I.D. column maintained at 30 degrees C. The mobile phase comprises acetonitrile sodium sulphate (0.7%, m/v)-phosphoric acid (6.8%, v/v dilution of 85%, m/m phosphoric acid)-water (22.25:50:5:22.75) at a flow-rate of 1.0 ml/min. Detection was by UV at 215 nm. The method is able to resolve polymyxin B1, the major component, from more than thirty other components present in the complex. Robustness was evaluated by performing a full-factorial design experiment. The method showed good selectivity, repeatability, linearity and sensitivity. PMID- 10722083 TI - Ion-pair chromatography on a porous graphitic carbon stationary phase for the analysis of twenty underivatized protein amino acids. AB - The analysis of twenty underivatized protein amino acids has been achieved on porous graphitic carbon packing material (Hypercarb). Five perfluoroalkyl carboxylic acids (trifluoroacetic, heptafluorobutyric, nonafluoropentanoic, tridecafluoroheptanoic and pentadecafluorooctanoic acid) have been studied as ion pairing reagent. Several parameters (equilibration time, quantities adsorbed onto the chromatographic support, concentration and nature of the ion-pairing reagent, as well as temperature effect) have been studied leading to the complete separation of these compounds in gradient elution mode. Evaporative light scattering detector has allowed the detection of these non UV-visible absorbent molecules. The chromatographic methodology developed can also be easily coupled with pneumatically assisted electrospray mass spectrometry. PMID- 10722084 TI - Identification of unusual (modified and non-encoded) amino acid residues in peptides by combinations of high-performance liquid chromatography and high performance capillary electrophoresis with matrix-assisted laser desorption ionization time-of-flight mass spectrometry. AB - The techniques for micro-level analysis of some widespread unusual amino acids (phosphorylated and hydroxylated ones) as well as of some genetically non-encoded amino acids were developed for their subsequent identification in the peptide and protein amino acid sequence by narrow-bore column high-performance liquid chromatography (approximately 10 pmol of the sample), high-performance capillary electrophoresis (approximately 1-10 pmol), matrix-assisted laser desorption ionization time-of-flight mass spectrometry (approximately 1-10 pmol), and automatic protein gas phase sequencing (approximately 1-50 pmol). PMID- 10722085 TI - Retention/quantitation properties of the o-phthaldialdehyde-3-mercaptopropionic acid and the o-phthaldialdehyde-N-acetyl-L-cysteine amino acid derivatives in reversed-phase high-performance liquid chromatography. AB - The separation/identification of 25 amino acids as their o-phthaldialdehyde-3 mercaptopropionic acid (OPA/MPA) and o-phthaldialdehyde-N-acetyl-L-cysteine (OPA/NAC) derivatives have been optimized [paying particular attention to those amino acids which elute with more than one derivative (glycine, histidine, gamma aminobutyric acid, beta-alanine, ornithine, lysine) and that are expected to be present in apples in their free form]. Optimum separation conditions are reported on six reversed-phase columns: Nucleosil 3 and 5 microm, 150(+20 guard)x4.0 mm; Gromsil 3 microm, 150(+10 guard)x4.0 mm; Hypersil 5 microm, 150(+20 guard)x4.0 mm and 200(+20 guard)x4.0 mm; and Hypersil 3 microm, 150(+20 guard)x4.0 mm. Elutions were followed, simultaneously, with photodiode array and fluorescence detectors connected in line. Optimization studies carried out in model solutions as a function of temperature (30-55 degrees C) and eluent flow-rate (0.8-2.5 mL/min) demonstrated that optimum resolutions are obtained with the highest flow-rate applicable (remaining on the safe side with a column pressure of << 3500 p.s.i.; 1 p.s.i.=6894.76 Pa) in the temperature range 30-50 degrees C. Twenty-five amino acids, eluting in 31 separate, characteristic derivatives, were determined on all six columns (the main component, asparagine, present in overwhelming excess, together with the minor constituents glutamine, beta-alanine, gamma-aminobutyric acid, homoserine, and homoarginine). Optimum conditions in the case of both derivatives were obtained on the same type of column (Hypersil, 5 microm), as follows: for the OPA/MPA amino acids with programmed flow-rate [1.3-2.3 ml/min; column, 200(+20 guard)x4 mm], at 50 degrees C, while, for the OPA/NAC amino acids at 2.1 ml/min flow rate, at 30 degrees C [column, 150(+20 guard)x4 mm], with 40 and 37 min run times, including equilibration. Responses of the corresponding amino acids proved to be independent of the column used; reproducibility in the concentration range 6-12,000 pmol, related to the injected amount of amino acids, was <3.4% RSD (average relative standard deviation percentage). The utility of the protocol was demonstrated in the quantitation of the free amino acid content of five apple varieties (Jonagored, Idared, Jonica, Florina, Freedom) on various harvesting dates and after different storage times. Derivatization of the apple pulp was performed with filtered samples, applying any special isolation processes. PMID- 10722086 TI - Screening for defects in tryptophan metabolism. AB - We introduced a two-step procedure for the detection of defects in metabolism of tryptophan: (1) HPTLC (described previously) is suitable when starting the investigation, (2) two HPLC methods with isocratic elution and spectrophotometric detection are used at the next step, when pathological findings are to be confirmed and the individual metabolites quantified. The first method enables the assessment of tryptophan, 5-hydroxyindolylacetic acid, indolylacetic acid, indolylacryloylglycine, indolylacrylic acid and its possible precursors, namely indolyllactic and indolylpropionic acids. The second procedure is intended for the monitoring of anthranilic, 3-hydroxyanthranilic, kynurenic and xanthurenic acids, kynurenine, 3-hydroxykynurenine and indoxyl-sulfate. The same pre-treated sample is used for all methods. PMID- 10722087 TI - Signature-peptide approach to detecting proteins in complex mixtures. AB - The objective of the work presented in this paper was to test the concept that tryptic peptides may be used as analytical surrogates of the protein from which they were derived. Proteins in complex mixtures were digested with trypsin and classes of peptide fragments selected by affinity chromatography, lectin columns were used in this case. Affinity selected peptide mixtures were directly transferred to a high-resolution reversed-phase chromatography column and further resolved into fractions that were collected and subjected to matrix-assisted laser desorption ionization (MALDI) mass spectrometry. The presence of specific proteins was determined by identification of signature peptides in the mass spectra. Data are also presented that suggest proteins may be quantified as their signature peptides by using isotopically labeled internal standards. Isotope ratios of peptides were determined by MALDI mass spectrometry and used to determine the concentration of a peptide relative to that of the labeled internal standard. Peptides in tryptic digests were labeled by acetylation with acetyl N hydroxysuccinimide while internal standard peptides were labeled with the trideuteroacetylated analogue. Advantages of this approach are that (i) it is easier to separate peptides than proteins, (ii) native structure of the protein does not have to be maintained during the analysis, (iii) structural variants do not interfere and (iv) putative proteins suggested from DNA databases can be recognized by using a signature peptide probe. PMID- 10722088 TI - Separation and characterization of multicomponent peptide mixtures by liquid chromatography-electrospray ionization mass spectrometry. Application to crude products of the synthesis of leuprolide. AB - Leuprolide is a synthetic structural analogue of luteinizing hormone-releasing hormone used for the treatment of a large number of diseases related with the regulation of sexual hormones. Solid-phase peptide synthesis is used to obtain leuprolide peptidic hormone, but this synthetic procedure results in complex mixtures that need separation and characterization. Here, liquid chromatography coupled with mass spectrometry using electrospray ionization, (LC-ES-MS), was used for the separation and characterization of multicomponent peptide mixtures of crudes of synthesis of leuprolide. To optimize the LC separation process, the method of linear solvation energy relationships was applied and the powerful coupling LC-ES-MS permitted rapid and reliable characterization of the reaction product. PMID- 10722089 TI - Separation of potentially therapeutic peptide hormones by liquid chromatography. Optimisation of the composition and pH of the mobile phase. AB - The aim of this work is to optimise the proportion of the organic modifier and the pH of the mobile phase, in order to separate a series of peptide hormones with therapeutic interest in the molecular mass range from 500 to 6000. The composition of the mobile phase was optimised by establishing relationships between retention parameters and either the scale of solvent polarity, or the Kamlet-Taft multiparameter solvent scale of the eluent, using linear solvation energy relationships. Likewise, linear correlations between the chromatographic retention and Reichardt's E(T)N parameter were obtained. These relationships allowed an important reduction of the experimental retention data needed for developing a given separation. In addition, a model describing the effect of the correctly measured pH of the mobile phase on retention in LC was established and tested for the series of selected peptides using an octadecylsilica column. The proposed equations permit the prediction of the optimum pH and also permit the determination of the acidity constants of the peptides in the hydro-organic mixtures using a minimum number of measurements. PMID- 10722090 TI - Simultaneous separation of angiotensin and endothelin peptide families by high performance liquid chromatography: application to the specific radioimmunoassay measurement of angiotensin II or endothelin-1 from tissue. AB - Currently available measurements of endogenous angiotensin II (ANG II) and endothelin-1 (ET-1) concentrations by radioimmunoassay (RIA) lack specificity to ANG II or ET-1. ANG II and ET-1 antibodies cross-react with immuno-reactive angiotensin and endothelin family members, respectively. We have therefore developed an ion-pair reversed-phase high-performance liquid chromatography (HPLC) for simultaneously separating angiotensin and endothelin peptides and enhancing RIA specificity in the measurement of ANG II and ET-1. The developed HPLC separation was applied to canine myocardium extracts; ANG II or ET-1 fractions were collected and quantified by RIA. Elution times for both peptide families, ANG I, ANG II, ANG III, ANG IV, ANG II(4-8), bET-1, ET-1, ET-2 and ET-3 were within 25 min. In normal canine myocardium from the right atrium, right ventricle, left atrium and left ventricle, ANG II concentrations were 39+/-11, 28+/-21, 31+/-11 and 21+/-8 fmol/g and ET-1 concentrations were 43+/-16, 42+/-19, 55+/-21 and 57+/-34 fmol/g (mean+/-SD, N=7), respectively. The combination of HPLC with RIA renders the measurement of ANG II or ET-1 specific and convenient, and saves time. This HPLC separation may be applied to the specific measurement of other immuno-reactive angiotensin and endothelin peptides. PMID- 10722091 TI - Quantitative analysis of hydrophobic pulmonary surfactant proteins by high performance liquid chromatography with light-scattering detection. AB - A new method for the separation and quantification of two hydrophobic lung surfactant proteins (SPs) is described. It is based on size-exclusion chromatography using the apolar stationary phase butyl silicagel with a pore size of 30 nm and isocratic elution with chloroform, methanol and trifluoroacetic acid. The samples were prepared from sheep lung lavage fluid by centrifugation and fractional extraction with butanol and chloroform-methanol. The chromatograms show three peaks in the elution order SP-B, SP-C and lipids. A small peak ahead of SP-B, which disappeared after reduction with 2-mercaptoethanol, was oligomeric SP-B. The response of the evaporative light-scattering detector was non-linear. For preparative high-performance liquid chromatography ultraviolet detection at 279 nm is recommended. PMID- 10722092 TI - Analysis of major ovine milk proteins by reversed-phase high-performance liquid chromatography and flow injection analysis with electrospray ionization mass spectrometry. AB - Ovine milk proteins were analyzed both by coupling HPLC and electrospray ionization mass spectrometry (ESI-MS) and by flow injection analysis and ESI-MS detection after separation and collection of fractions from gel permeation chromatography. These methods resolved the four ovine caseins and whey proteins and made it possible to study the complexity of these proteins associated with genetic polymorphism, post-translational changes (phosphorylation and glycosylation) and the presence of multiple forms of proteins. The experimental molecular masses of ewe milk proteins were: 19,373 for kappa-casein 3P; 25,616 for alpha(s2)-casein 10P; 23,411 for alpha(s1)-casein C-8P; 23,750 for beta casein 5P; 18,170 and 18,148 for beta-lactoglobulins A and B; 14,152 for alpha lactalbumin A and 66,322 for serum albumin. PMID- 10722093 TI - Determination of lipoic acid and dihydrolipoic acid in human plasma and urine by high-performance liquid chromatography with fluorimetric detection. AB - A highly sensitive method for the determination of alpha-lipoic acid (LA) and dihydrolipoic acid (DHLA) in human plasma and urine has been developed. Samples were acidified and extracted with organic solvent, and the free sulfhydryls of DHLA protected as the dicarboxyethylate by treatment with ethylchloroformate. The free carboxylic function of LA and the SH-protected DHLA were converted into their amide derivatives with the strong fluorophore 2-(4-aminophenyl)-6 methylbenzothiazole in the presence of a coupling agent and a base catalyst. The resulting fluorescent amides of both LA and DHLA were separated on a reversed phase column (Ultrasphere C8) using simple isocratic elution with acetonitrile water (80:20) and detected fluorimetrically (excitation 343, emission 423 nm). The method is highly sensitive, reproducible, and is easily applied for the simultaneous determination of LA and DHLA in biological samples. PMID- 10722094 TI - Monitoring of extracellular pyruvate, lactate, and ascorbic acid during cerebral ischemia: a microdialysis study in awake gerbils. AB - In vivo microdialysis coupled with liquid chromatography was developed for the continuous monitoring of brain neurochemicals during cerebral ischemia in awake, free moving gerbils. The dead volume of the microdialysis system was estimated to be less than 30 microl. The detection limits of the present assay were 0.2 to 2.0 microM for analytes at a signal to noise ratio of five. To validate this assay, a focal cerebral ischemia was produced by occlusion of one common carotid artery for 60 min and then reperfusion for additional 3 h in awake gerbils. A microdialysis probe was inserted into the striatum of the gerbil. Dialysates were autoinjected and analyzed extracellular pyruvate, lactate, and ascorbic acid by liquid chromatography with a UV detector during cerebral ischemia. Significant changes of pyruvate and the lactate/pyruvate ratio were observed. Biphasic and dynamic changes in ascorbic acid and lactate were proposed to correlate a secondary damage. This assay can be used as a tool to study dynamic changes of brain neurochemicals in awake animals. PMID- 10722095 TI - Analysis of biogenic amines by solid-phase microextraction and high-performance liquid chromatography with electrochemical detection. AB - We investigated the new solid-phase microextraction method by high-performance liquid chromatography with electrochemical detection for the analysis of biogenic amines. The Carbowax-Templated Resin 50 microm (purple) fibre coating offers good performances for dopamine and serotonin separation, i.e., good selectivity and high sensibility (0.1 microg l(-1)). We also tested this fibre for biogenic amines quantification of rat striatum. The coating seems to be selective towards the amines and has low affinity for the metabolites, allowing a good separation and preventing drawbacks from the biological matrix. These first results obtained using this original separation method offer large perspectives of application to many biological samples. PMID- 10722096 TI - Determination of catecholamines in pheochromocytoma cell (PC-12) culture medium by microdialysis-microbore liquid chromatography. AB - An in vitro microdialysis system was constructed for the measurement of catecholamines in pheochromocytoma cell culture medium. The novel microdialysis device is composed of a petri dish, a dialysis membrane and two transmission tubes. The dialysis membrane is located in the space of a petri dish such that it is immersed in the culture medium. Catecholamines contained in the culture medium diffused into a designed dialysis membrane with sufficient recovery (about 60%). Dialysates were collected by a sampling loop and introduced by an on-line injector to a microbore liquid chromatographic system for analysis of catecholamines. This assay yielded a detection limit of 0.2-0.5 pg/injection with acceptable intra- and inter-assay reproducibilities in 5 microl of dialysates. To evaluate the on-line microdialysis system, PC-12 cells were cultured in a petri dish within an incubator. The baseline concentration of dopamine in PC-12 cell culture medium was about 0.29 ng/ml which was elevated to 2.43 ng/ml after treatment with 0.5 mM potassium cyanide. In conclusion, the present microassay provides for the sensitive, direct measurement of catecholamines in culture medium while minimizing pretreatment procedures for sample preparation. PMID- 10722097 TI - Sheath liquid effects in capillary high-performance liquid chromatography electrospray mass spectrometry of oligonucleotides. AB - Fused-silica capillary columns of 200 microm inner diameter were packed with micropellicular, octadecylated, 2.3 microm poly(styrene-divinylbenzene) particles and applied to the separation of oligonucleotides by ion-pair reversed-phase high performance liquid chromatography. Oligonucleotides were eluted at 50 degrees C with gradients of 3-13% acetonitrile in 50 mM triethylammonium bicarbonate. Addition of sheath liquid to the column effluent allowed the detection of oligonucleotides by electrospray ionization mass spectrometry using full-scan data acquisition with a detectability comparable to that obtained with UV detection. The signal-to-noise ratios with different sheath liquids increased in the order isopropanolA) and initiation codon (ATG-->ACG), were found in the homozygous condition in patients belonging to Balochi and Sindhi ethnic groups of Pakistan, together with heterozygous and homozygous alpha(-3.7) deletions, respectively. A frameshift mutation at codon 44 (-C) was identified in a patient belonging to the Gujrati ethnic group together with IVS-I-1 (G-->T) and a normal complement of four a globin genes. Haplotype analysis was performed to identify the chromosomal background associated with these mutations, and for tracing the origin and spread of these mutations. PMID- 10722110 TI - Molecular characterization of beta-thalassemia in Syria. AB - This study concerns the determination of beta-thalassemia alleles and other hemoglobin variants in 82 patients from Syria. We have characterized 146 chromosomes and found 17 different beta-thalassemia mutations, and one beta globin chain variant that gives rise to the abnormal Hb S. The eight most common beta-thalassemia mutations were the IVS-I-110 (G-->A), IVS-I-1 (G-->A), codon 5 ( CT), -30 (T-->A), codon 39 (C-->T), IVS-I-6 (T-->C), IVS-II-1 (G-->A), and codon 15 (TGG-->TAG). These mutations accounted for almost 75% of the total beta thalassemia chromosomes. We identified 34 different genotypes with a high level of homozygosity. The various beta-thalassemia mutations were characterized using gene amplification with specific oligonucleotide primers, restriction enzyme analysis, denaturing gradient gel electrophoresis and direct sequencing. By combining these three approaches we were able to detect mutations in almost 90% of the chromosomes studied. Our findings provide a sound foundation on which to base a preventive program for thalassemia and we believe that the data that we present will facilitate the improvement of medical services such as carrier screening, genetic counseling, and prenatal diagnosis. Furthermore a detailed knowledge of the molecular pathology of beta-thalassemia will strongly improve the prenatal diagnosis services in Syria. PMID- 10722112 TI - Beta-thalassemia intermedia associated with homozygosity for the -87 (C-->T) mutation in a Turkish family. AB - We report on two siblings with beta+-thalassemia intermedia. Molecular studies of the beta-globin gene indicated that the patients are homozygous for the -87 (C- >T) mutation. This genotype has not been previously described. Homozygosity for the -87 (C-->T) mutation produces a mild form of beta+-thalassemia associated with moderate Hb F elevation (26-38%) and highly elevated Hb A2 (10-8.6%) levels, respectively. Hematological parameters of homozygous -87 (C-->G) and -87 (C-->A) mutations, and compound heterozygous patients with either C-->T, C-->G, or C-->A at -87 and one of the severe beta+- or beta0-thalassemia mutations, are given for comparison. PMID- 10722113 TI - Hb Sallanches [alpha104(G11)Cys-->Tyr]: a rare alpha2-globin chain variant found in the homozygous state in three members of a Pakistani family. AB - We have identified a rare alpha2-globin chain variant, Hb Sallanches [alpha104(G11) Cys-->Tyr], in a Pakistani family having three homozygous patients with transfusion-dependent Hb H disease. This variant, previously reported in a French patient and a West Indian homozygous patient with Hb H disease, is due to a mutation at codon 104 (TGC-->TAC). This is the third case of Hb Sallanches and the first case with three homozygous patients reported in Pakistan. Due to the different ethnic origins of the patients, it is very likely an independent mutation. PMID- 10722114 TI - A new, electrophoretically silent, fetal hemoglobin variant: Hb F-Calabria [Ggamma118(GH1)Phe-->Leu]. AB - Hb F-Calabria [Ggamma118(GH1)Phe-->Leu] is a new fetal hemoglobin variant that was found during routine screening for abnormal hemoglobins in a newborn of Calabrian (Southern Italy) ancestry. The variant chain was identified (acid urea gel electrophoresis of dissociated globin chains in the presence of Triton X-100, and by reversed phase high performance liquid chromatography) as a slightly hydrophilic Ggamma chain. Sequencing of the polymerase chain reaction-amplified exon 3 of the Ggamma-globin gene demonstrated the TTC-->CTC mutation at codon 118 leading to the Phe-->Leu conservative substitution at position GH1. A molecular modeling study supports that the variant might not have clinical implications. This is the 40th example of a Ggamma chain variant. PMID- 10722115 TI - Two new Ggamma chain variants: Hb F-clamart [gamma17(A14)Lys-->Asn] and Hb F Ouled Rabah [gamma19(B1)Asn-->Lys]. AB - Two new fetal hemoglobin variants affecting the Ggamma chain are reported. Hb F Clamart was found during investigation of a French newborn who presented with a mild microcytemia. The second variant was found during neonatal screening for hemoglobinopathies of 30,000 babies from a population-at-risk living in the Paris region. It was named Hb F-Ouled Rabah because its structural modification and ethnic distribution is similar to that of Hb D-Ouled Rabah [beta19(B1)Asn-->Lys]. Hb F-Ouled Rabah is clinically silent and occurs at a frequency of ca. 0.1% in newborns originating from Maghreb. Structural characterization of both variants was done by protein chemistry methods, including amino acids analysis and mass spectrometry. PMID- 10722116 TI - Characterization by DNA sequencing of Hb F-Columbus-GA [Ggamma94(FG1)Asp-->Asn] observed in Sardinian newborn. PMID- 10722117 TI - Hb Vila Real [beta36(C2)Pro-->His]: a newly discovered high oxygen affinity variant. PMID- 10722118 TI - Hb Ube-2 [alpha68(E17)Asn-Asp] and Hb Hafnia [beta116(G18)His-->Gln] observed during neonatal screening in Brussels. PMID- 10722119 TI - Hb Siam [alpha15(A13)Gly-->Arg] is a GGT-->CGT mutation in the alpha1-globin gene. PMID- 10722120 TI - The role of receptor structure in determining adenosine receptor activity. AB - Adenosine produces a wide variety of physiological effects through the activation of cell surface adenosine receptors (ARs). ARs are members of the G-protein coupled receptor family, and currently, four subtypes, the A1AR, A2AAR, A2BAR, and A3AR, are recognized. This review focuses on the role of receptor structure in governing various facets of AR activity. Ligand-binding properties of ARs are primarily dictated by amino acids in the transmembrane domains of the receptors, although a role for extracellular domains of certain ARs has been suggested. Studies have identified certain amino acids conserved amongst AR subtypes that are critical for ligand recognition, as well as additional residues that may differentiate between agonist and antagonist ligands. Receptor regions responsible for activation of Gs have been identified for the A2AAR. The location of these intracellular sites is consistent with findings described for other G protein-coupled receptors. Site-directed mutagenesis has been employed to analyze the structural basis for the differences in the kinetics of the desensitization response displayed by various AR subtypes. For the A2AAR and A3AR, agonist stimulated phosphorylation of the AR, presumably via a G-protein receptor kinase, has been shown to occur. For these AR subtypes, intracellular regions or individual amino acids that may be targets for this phosphorylation have been identified. Finally, the role of A1AR gene structure in regulating the expression of this AR subtype is reviewed. PMID- 10722121 TI - Pharmacokinetic profile of levetiracetam: toward ideal characteristics. AB - Levetiracetam is a novel orally active antiepileptic drug with a unique preclinical profile. It has a high therapeutic index and potential antiepileptogenic effects. Results of clinical trials indicate activity in partial-onset and generalized seizures. The pharmacokinetic profile of levetiracetam closely approximates the ideal characteristics expected of an antiepileptic drug, with good bioavailability, rapid achievement of steady-state concentrations, linear and time-invariant kinetics, minimal protein binding, and minimal metabolism. The major metabolic pathway of levetiracetam is not dependent on the hepatic cytochrome P450 system, and levetiracetam does not inhibit or induce hepatic enzymes to produce clinically relevant interactions. Sixty-six percent of an administered levetiracetam dose is eliminated unchanged in urine; 24% is metabolized to an inactive metabolite that is detectable in blood and is also excreted in urine. Total body clearance of levetiracetam is decreased in patients with renal impairment, and doses should be modified according to creatinine clearance values. Levetiracetam is not appreciably protein-bound, nor does it affect the protein binding of other drugs. Thus, because of its minimal protein binding and lack of hepatic metabolism, the risk of drug interactions is very low. Levetiracetam has a wide margin of safety and patient-friendly pharmacokinetics that distinguish it from other currently available antiepileptic drugs. This profile may facilitate the clinical management of patients with epilepsy by providing a safer and less-complicated therapeutic strategy. PMID- 10722122 TI - Muscarinic toxins from the green mamba. AB - Muscarinic acetylcholine receptors are involved in many important physiological processes. Discovery of different subtypes of muscarinic receptors that are responsible for modulating specific physiological events was a key development in muscarinic receptor research. However, the lack of highly selective muscarinic agonists and antagonists has made the classification of a muscarinic receptor subtype responsible for the mediation or modulation of a particular response very difficult. Toxins have previously proved to be highly useful pharmacological tools, due to their high potency and selectivity. This review looks at a new class of muscarinic ligand isolated from the venom of the Eastern green mamba (Dendroaspis angusticeps). Just over a decade ago, it was found that two toxins from the green mamba venom appeared to distinguish between different muscarinic receptor subtypes. Since then, at least 10 more muscarinic toxins (MTs) have been isolated from mamba venom. In recent years, some of the MTs have been used as pharmacological tools; for example, to determine the muscarinic receptor subtype involved in inhibition of adenylyl cyclase in rat striatum. This review looks at the progress that has been made over the past 10 years in the area of MT research and examines whether or not these new peptides are a new way forward in the field of muscarinic receptor research. PMID- 10722123 TI - A community--wide outbreak of Salmonella enterica serotype Typhimurium infection associated with eating a raw milk soft cheese in France. AB - In 1997, a community-wide outbreak of Salmonella enterica serotype Typhimurium (S. typhimurium) infection occurred in France. The investigation included case searching and a case-control study. A case was defined as a resident of the Jura district with fever or diarrhoea between 12 May and 8 July 1997, from whom S. typhimurium was isolated in stool or blood. One hundred and thirteen cases were identified. Thirty-three (83 %) of 40 cases but only 23 (55 %) of 42 community controls, matched for age and area of residence, reported eating Morbier cheese (Odds ratio: 6.5; 95 % Confidence Interval: 1.4-28.8). Morbier cheese samples taken from the refrigerators of two case-patients and one symptom-free neighbour cultured positive for S. typhimurium of the same phage type as the human isolates. The analysis of distribution channels incriminated one batch from a single processing plant. These findings show that an unpasteurized soft cheese is an effective vehicle of S. typhimurium transmission. PMID- 10722124 TI - Characterization of multi-drug resistant Salmonella typhi isolated from Pakistan. AB - Thirty-nine strains of Salmonella typhi, isolated in 1995 from four Districts in Pakistan, Rawalpindi, Islamabad, Kharian and Jehlem, were catalogued and examined. Chromosomal DNA from each isolate was digested with XbaI restriction endonuclease and subjected to pulsed-field gel electrophoresis. Three clonal variants comprising of 17-19 DNA fragments were identified. Antibiotic susceptibility testing identified that 37 of the S. typhi were resistant to chloramphenicol, trimethoprim and ampicillin. These antibiotic resistance genes were found to be located on one of four plasmids belonging to incompatibility group IncHI1 and ranging in size from 150-175 Kb. The genes responsible for this resistance in each case were the chloramphenicol acetyltransferase (CAT) type I, the dihydrofolate reductase (DHFR) type VII and the beta-lactamase TEM-1 respectively. PMID- 10722125 TI - The antibiotic resistance patterns of Salmonella Typhi isolates in Italy, 1980 96. The Italian SALM-NET Working Group. Salmonella Network. AB - In this paper we report the distribution of Salmonella Typhi isolates in Italy and their resistance patterns to antibiotics. The data were collected by the Italian SALM-NET surveillance system in a pilot retrospective study of the period 1980-96. Data on drug-resistance were available for 82 isolates out of 176 S. Typhi isolated in Italy. Of these 82 isolates, 32 (39%) were resistant or intermediate to 1 or more antibiotics. Eight isolates were resistant and 7 intermediate to streptomycin; 4 isolates were resistant to ampicillin alone or in association with other antibiotics; only 2 strains (1 isolated in Lombardia in 1993 and the other 1 in Lazio in 1994) were resistant to chloramphenicol, and 2 (isolated in Sardegna and Piemonte in 1995 and 1996, respectively) showed intermediate resistance to chloramphenicol. The strains showing resistance to 3 or more antibiotics were very scarce: 1 (with 5 complete resistances) was isolated in Lazio in 1994, and another 1 (with complete resistance to 10 antibiotics and intermediate resistance to 2 antibiotics) was isolated in Molise in 1988. In conclusion, besides the routine activities to control typhoid fever, an accurate and continuous surveillance is necessary in order to quickly identify multidrug-resistant (MDR) S. Typhi strains and prevent their spread, even though their level, in our country, is still quite low. PMID- 10722126 TI - Molecular evidence of clonality amongst Vibrio cholerae O1 biotype El Tor during an outbreak in Malaysia. AB - Forty-three clinical strains of V. cholerae O1 biotype E1 Tor were isolated between 3 May and 10 June 1998 during an outbreak in the metropolitan area of Kuala Lumpur and its suburbs. With the exception of three Inaba strains that were restricted to three members of a family, all the others belonged to the Ogawa serotype. The strains were analysed for clonality using ribotyping and pulsed field gel electrophoresis (PFGE). Two ribotypes, V/B21a and B27, were identified among 40 Ogawa isolates using BglI restriction endonuclease. Ribotype V/B21a has been described previously from Taiwan and Colombia and several Asian countries while B27 has been reported among isolates from Senegal. The three Inaba strains belonged to one ribotype, designated type A, not previously reported. PFGE analysis using NotI revealed that all isolates within a ribotype had identical profiles demonstrating clonality amongst the strains. Dice coefficient analysis of the two Ogawa genotypes revealed 89% similarity on ribotype patterns and 91.3% on PFGE profiles. Ribotype V/B21a isolates were associated with cases from dispersed areas of Kuala Lumpur and its suburbs while ribotype B27 was restricted to cases from one particular area suggesting a common-source outbreak. PMID- 10722127 TI - An outbreak of Escherichia coli O157:H7 infections and haemolytic uraemic syndrome associated with consumption of unpasteurized apple cider. AB - During October 1996, an outbreak of Escherichia coli O157:H7 infections among Connecticut residents occurred. An epidemiologic investigation included enhanced surveillance and a case-control study. Clinical isolates of Escherichia coli O157:H7 were typed by pulsed-field gel electrophoresis (PFGE). Implicated cider samples were analysed by culture and polymerase chain reaction (PCR). Consumption of implicated cider was associated with illness; (matched odds ratio = undefined, 95 % confidence interval = 3.5-infinity). Ultimately, a total of 14 outbreak associated patients were identified. All isolates analysed by PFGE yielded the outbreak-associated subtype. Escherichia coli O157:H7 was not cultured from three cider samples; PCR analysis detected DNA fragments consistent with Escherichia coli O157:H7 in one. This outbreak was associated with drinking one brand of unpasteurized apple cider. PFGE subtyping supported the epidemiologic association. PCR analysis detected microbial contaminants in the absence of live organisms. Washing and brushing apples did not prevent cider contamination. PMID- 10722128 TI - A five year outbreak of methicillin-susceptible Staphylococcus aureus phage type 53,85 in a regional neonatal unit. AB - We identified a 5-year outbreak of a methicillin-susceptible Staphylococcus aureus (MSSA) strain, affecting 202 babies on a neonatal unit, by routine weekly phage typing all S. aureus isolates. Multiple staged control measures including strict emphasis on hand hygiene, environmental and staff surveillance sampling, and application of topical hexachlorophane powder failed to end the outbreak. S. aureus PT 53,85 (SA5385) was found on opened packs of Stomahesive, used as a neonatal skin protectant. Only following the implementation of aseptic handling of Stomahesive, and the use of topical mupirocin for staff nasal carriers of SA5385, and for babies colonized or infected with S. aureus, did the isolation rate of SA5385 decline. DNA fingerprinting indicated that > or =, 95% of SA5385 isolates were clonal. In vitro death rates of SA5385 on Stomahesive with human serum were significantly lower than on Stomahesive alone (P = 0.04), and on cotton sheet with serum (P = 0.04), highlighting the potential of this material as a survival niche. Phage typing remains a valuable, inexpensive and simple method for monitoring nosocomial MSSA infection. PMID- 10722129 TI - Isolation rates of Streptococcus pyogenes in patients with acute pharyngotonsillitis and among healthy school children in Iran. AB - We examined three populations from the Tehran region and the North part of Iran (Gilan), in all more than 5000 individuals, for carriage of Streptococcus pyogenes (group A streptococci; GAS). Children or adults with acute pharyngotonsillitis and healthy school children harboured GAS in 34-1, 20.0 and 21.0%, respectively. Typing of 421 randomly selected isolates showed a predominance of M-types M4, M5, M11, M12, as well as the provisional type 4245; however, many of the isolates were T and M non-typable. Forty-three percent of all strains were opacity factor (OF) negative. The type distribution differed markedly from that reported in 1973-4, when M types 1 and 12 were predominant. PMID- 10722130 TI - Eradication of vancomycin resistant Enterococcus faecium from a paediatric oncology unit and prevalence of colonization in hospitalized and community-based children. AB - We previously reported an outbreak of vancomycin resistant enterococci (VRE) in a paediatric oncology unit in December 1995 which was associated with widespread environmental contamination of the unit with VRE. We undertook this study to evaluate the effectiveness of the infection control policy instituted subsequent to the outbreak and to investigate the underlying prevalence of VRE colonization in hospitalized, outpatient and community-based children. We sought to establish the molecular similarity of VRE isolates from the study. Stool specimens were obtained from outpatients at risk of VRE, hospital inpatients and from healthy community-based children. VRE colonization was eradicated from the inpatient unit within 11 months, but in outpatients, 16 months after the outbreak, 4 of 137 (2.9 %) attending oncology outpatients, 5 of 65 (7.7%) with cystic fibrosis and 1 of 12 (8.3 %) with liver disease were found to be colonized with VRE. The isolates were all Enterococcus faecium, Van A phenotype except one E. casseliflavus of the Van C phenotype. All were unique in SmaI DNA macrorestriction patterns with the exception of two isolates, which were similar to the original outbreak strain and three further isolates of a single strain but which differed from the outbreak strain. Of 315 hospital inpatients, 2.5 % were colonized with VRE of the Van C resistance phenotype but VRE was not detected in 116 healthy, community-based children. We conclude that effective strategies can successfully control spread of VRE but despite a low prevalence of VRE colonization in hospital patients and in community-based children, outbreaks can occur when infection control practices are not optimal. Continued vigilance to detect VRE and limit spread within hospitals is therefore necessary. PMID- 10722131 TI - Occurrence of Clostridium perfringens beta2-toxin amongst animals, determined using genotyping and subtyping PCR assays. AB - Clostridium perfringens isolates are currently classified into one of five biotypes on the basis of the differential production of alpha-, beta-, epsilon- and iota-toxins. Different biotypes are associated with different diseases of man and animals. In this study a multiple PCR assay was developed to detect the genes encoding these toxins. In addition, detection of the genes encoding the C. perfringens enterotoxin and beta2-toxin allowed subtyping of the bacteria. C. perfringens isolates taken from a variety of animals, including foals, piglets or lambs, were genotyped using this assay. Most of the isolates were found to be genotype A and the gene encoding beta2-toxin [corrected] was present in 50% of the isolates genotyped. A significant association between C. perfringens possessing the beta2-toxin gene and diarrhoea in piglets was identified, suggesting that beta2-toxin may play a key role in the pathogenesis of the disease. PMID- 10722132 TI - Serogroup C meningococcal disease outbreak associated with discotheque attendance during carnival. AB - In the week following a carnival during 19-24 February 1998, an outbreak of meningococcal disease occurred in a rural German county. The available isolates belonged to phenotype C:2a:P1.2,5 and were clonally related by pulsed-field gel electrophoresis. A case-control study was done to identify risk factors for the outbreak and to define possible vaccination target groups. Five persons aged 13 16 years who fell ill during 24-27 February were included in the study. Four of 5 cases and 10 of 32 controls visited local discotheques (OR = 8.8; P = 0.06). Cases also visited discotheques more frequently than controls (chi2 for trend, P = 0.0002). Multiple discotheques during the carnival may have been predominant locations of transmission in this outbreak. Because this risk factor was limited in time, a mass community vaccination campaign was not initiated. PMID- 10722133 TI - Management of an outbreak of meningococcal meningitis in a Sudanese refugee camp in Northern Uganda. AB - We describe an outbreak of meningitis at a Sudanese refugee camp in Northern Uganda that lasted over a year from February 1994. Some 291 cases occurred in a refugee population of 96860 (averaged over the year), an attack rate of 0.30%. The case fatality rate was 13.3%. From a small number of samples taken for culture N. meningitidis serogroup A, serotype 21:P1.9, clone III-1 was identified as the causative organism. The outbreak started in the camp's reception centre which had the highest attack rate. Spread from the reception centre was rapid and the epidemic reached its peak within 3 weeks. All of the cases amongst residents of the reception centre reported having had meningococcal vaccine before arriving at the camp and so were not immunized on arrival as would normally have been the case. Some 37 547 doses of meningococcal vaccine were used in a mass immunization campaign in February and March 1994. Following this the outbreak was declared over in August 1994 when no cases were registered for 2 consecutive weeks. However, following a massive and sudden influx of refugees a new epidemic peak occurred during February 1995. Many of these new refugees were also not immunized on arrival due to pressures of numbers. A follow-up immunization campaign then brought an end to the outbreak. Our experience confirms the effectiveness of timely and high-coverage immunization campaigns in controlling group A meningitis outbreaks amongst refugees in Africa. PMID- 10722134 TI - Restriction enzyme analysis and ribotyping distinguish Bordetella avium and Bordetella hinzii isolates. AB - Fifty-seven bacterial isolates previously identified as Bordetella avium or B. hinzii were characterized by restriction enzyme analysis (REA) and/or ribotyping. Twenty restriction endonucleases were evaluated for REA. Digestion of chromosomal DNA from the 42 B. avium and 15 B. hinzii isolates with HinfI produced 8 and 7 distinct fingerprint profiles, respectively. Digestion with DdeI further discriminated these Bordetella species and produced 12 fingerprint profiles for B. avium and 4 profiles of B. hinzii. In addition, B. avium isolates were clearly distinguishable from B. hinzii isolates by ribotyping with the restriction endonuclease PvuII. The ribotype patterns of these two species of Bordetella were unique when compared to previously reported ribotype patterns for B. bronchiseptica isolates. Since it was possible to discern differences among isolates within each Bordetella species by REA analysis, we suggest that REA could be used in developing a typing system based on the fingerprint profiles generated. PMID- 10722135 TI - Molecular epidemiology of Helicobacter pylori: separation of H. pylori from East Asian and non-Asian countries. AB - The predominant H. pylori strain circulating among geographic locations differs with regard to the genomic structure. This study determined whether structural subtypes of the cagA 3' repeat region could be used to identify the population of origin of H. pylori isolates. We examined 600 cagA-positive H. pylori (Colombia, 100; USA, 100; France, 100; Canada, 20; Italy, 20; Korea, 100; Japan, 100; Hong Kong, 20; Taiwan, 20; Vietnam, 20). The cagA 3' region was amplified by PCR using primers specific to Japanese and Western 3' cagA gene sequences. PCR using Japanese cagA primers resulted in PCR products in 99-6 % of strains from East Asia but no non-Asian strains. Conversely, PCR using Western cagA primers resulted in amplicons in 100% of non-Asian strains, and only one from East Asia. cagA genotyping is useful for molecular epidemiological studies as strains can be completely separated by differences in the cagA 3' region. PMID- 10722136 TI - Attentional function in secondary school students receiving isoniazid prophylaxis for tuberculosis infection. AB - Reports have suggested that isoniazid treatment may be associated with poor concentration and subtle reduction in memory. This study examines attentional function and processing speed in a group of 25 adolescents who received isoniazid prophylaxis for at least 6 months. As adolescents often face major educational assessment milestones, such cognitive side effects may have important implications. Participants were assessed before treatment, 1 month into treatment and at least 1 week after treatment cessation. Measures included the Paced Auditory Serial Addition Test and subtests of the appropriate Wechsler scale sensitive to attention and speed of information processing. Isoniazid does not appear to cause significant adverse effects on attentional function in adolescents. PMID- 10722137 TI - Molecular typing of Mycoplasma pneumoniae strains by PCR-based methods and pulsed field gel electrophoresis. Application to French and Danish isolates. AB - Restriction fragment length polymorphism (RFLP) analysis of the amplified P1 gene was used to type 153 strains of Mycoplasma pneumoniae isolated in France between 1977 and 1994, and in Denmark between 1962 and 1994, and an additional group of 28 strains isolated from Belgium and Germany between 1990 and 1993. Random amplified polymorphic DNA (RAPD) analysis was tested on French, Belgian and German strains. Both methods separated the strains into two groups corresponding to the two reference strains M129 (group I) and FH (group II), and gave concordant results. When 75 selected strains of different geographical origin were analysed by pulsed-field gel electrophoresis (PFGE), strains of group II fell into two closely related subgroups, subgroup IIa corresponding to the reference strain FH, and subgroup IIb. Most of the strains isolated in Denmark in the period 1962-86 belonged to group I. Almost all strains isolated in France and Denmark between 1987 and 1988 were from group II, the two subgroups being present. In 1991-3, almost all strains from France as well as Denmark, Germany and Belgium belonged to group I. PMID- 10722138 TI - The logic of causation and the risk of paralytic poliomyelitis for an American child. AB - Beginning in January 1997, American immunization policy allowed parents and physicians to elect one of three approved infant vaccination strategies for preventing poliomyelitis. Although the three strategies likely have different outcomes with respect to prevention of paralytic poliomyelitis, the extreme rarity of the disease in the USA prevents any controlled comparison. In this paper, a formal inferential logic, originally described by Donald Rubin, is applied to the vaccination problem. Assumptions and indirect evidence are used to overcome the inability to observe the same subjects under varying conditions to allow the inference of causality from non-randomized observations. Using available epidemiologic information and explicit assumptions, it is possible to project the risk of paralytic polio for infants immunized with oral polio vaccine (1.3 cases per million vaccinees), inactivated polio vaccine (0.54 cases per million vaccinees), or a sequential schedule (0.54-0.92 cases per million vaccinees). PMID- 10722139 TI - Hepatitis A in New South Wales, Australia from consumption of oysters: the first reported outbreak. AB - Between 22 January and 4 April 1997, 467 hepatitis A cases were reported to the New South Wales Health Department, Australia. To identify the cause of the outbreak, we conducted a matched case-control study, and an environmental investigation. Among 66 cases and 66 postcode-matched controls, there was a strong association between illness and consumption of oysters (adjusted odds ratio 42; 95 % confidence interval 5-379). More than two-thirds of cases reported eating oysters, including one third of cases and no controls who reported eating oysters in the Wallis Lake area. A public warning was issued on 14 February, and Wallis Lake oysters were withdrawn from sale. Hepatitis A virus was subsequently identified in oyster samples taken from the lake. Hepatitis A virus poses a special risk to consumers who eat raw oysters because it can survive for long periods in estuaries and cause severe disease. PMID- 10722140 TI - A secondary school outbreak of mumps following the childhood immunization programme in England and Wales. AB - Since the introduction of routine measles, mumps and rubella immunization for children in England and Wales in 1988, the incidence of mumps has declined steadily. We describe an outbreak of mumps in 1996 attacking 34 of a cohort of 98 schoolchildren born in 1982 and 1983. This is the largest outbreak in the UK since the introduction of the vaccine into the childhood immunization schedule. Salivary IgM assay was used as a simple, minimally invasive test to confirm the diagnosis. The occurrence of the outbreak demonstrates that British children who were just too old to receive mumps immunization in 1988 continue to be at risk of this disease as a result of diminished natural exposure. Further cases and outbreaks in this cohort are to be expected. Cohorts born before 1982 appear to be at less risk, presumably because of naturally acquired infection before the introduction of immunization. PMID- 10722141 TI - Prevalence of serum antibodies against bloodborne and sexually transmitted agents in selected groups in Somalia. AB - Somalia has suffered from a civil war during the last 10 years. In this period the use of whole blood has increased at least twofold in Mogadishu, Somalia compared with pre-war. Screening possibilities are limited. Recent data concerning the prevalence of infections with blood-borne and sexually transmitted agents are not available from this country. To investigate the spread of human immunodeficiency virus (HIV-1/2) and other blood-borne or sexually transmitted agents we tested a total of 256 serum samples collected in the summer of 1995 from blood donors, hospitalized children and adults in Mogadishu. The hepatitis B surface antigen (HbsAg) carrier rate was 191%, 5.6% and 21.3 % among blood donors, hospitalized children and hospitalized adults, respectively. However, no children under 2 years of age were HbsAg positive. The overall presence of antibodies against hepatitis C virus (HCV) was 2.4% (6/256). In blood donors this was 0.6% (1/157). In none of the samples tested, antibodies against HIV 1 and 2 or human T-cell lymphotropic viruses (HTLV I and II) were detected. Our results indicate that, during the civil war in Somalia, no evidence of an increase of HIV infections was found. Our findings indicate that preventive measures in Somalia should focus mainly on prevention of HBV-infections. HBV-vaccine could be administered within the framework of the expanded programme on immunization, as none of the children less than 2 years of age were HbsAg positive. PMID- 10722142 TI - Cross-sectional study on risk factors of HIV among female commercial sex workers in Cambodia. AB - To describe epidemiological features on HIV prevalence among female commercial sex workers (CSWs), a cross-sectional study on sexual behaviour and serological prevalence was carried out in Cambodia. The CSWs were interviewed on their demographic characters and behaviour and their blood samples were taken for testing on sexually transmitted diseases, including HIV, Chlamydia trachomatis, syphilis, and hepatitis B. Associations between risk factors and HIV seropositivity were analysed. High seroprevalence of HIV and Chlamydia trachomatis IgG antibody (CT-IgG-Ab) was shown among the CSWs (54 and 81.7%, respectively). Univariate logistic regression analyses showed an association between HIV seropositivity and age, duration of prostitution, the number of clients per day and CT-IgG-Ab. Especially, high-titre chlamydial seropositivity showed a strong significant association with HIV prevalence. In multiple logistic regression analyses, CT-IgG-Ab with higher titre was significantly independently related to HIV infection. These suggest that existence of Chlamydia trachomatis is highly related to HIV prevalence. PMID- 10722143 TI - Genetic analysis of IgG subclass responses against RESA and MSP2 of Plasmodium falciparum in adults in Papua New Guinea. AB - Contributions of environmental and genetic factors to IgG subclass responses against Plasmodium falciparum antigens RESA and MSP2 were investigated among adults in a highly endemic area of Papua New Guinea. Heritabilities were estimated using variance component analysis. Familial aggregation of several responses was found, including IgG1, IgG2 and IgG3 responses against RESA, IgG1 and IgG3 responses against the 3D7 form of MSP2 and IgG1, IgG2 responses against the FC27 form of MSP2. Allowance for sharing of houses explained some of the non genetic variance but not the familial aggregation. The variance of IgG3 responses against RESA and IgG1, IgG2 against MSP2 (FC27) was partly explained by sharing of HLA class II genotypes, although heritability was low. Segregation analyses indicated that any genetic regulation was more complex than governed by a single major gene. Such host genetic variation in responses to specific malaria antigens has implications for immuno-epidemiology and vaccine development. PMID- 10722144 TI - The occurrence of Trypanosoma evansi in buffaloes in Indonesia, estimated using various diagnostic tests. AB - The prevalence and incidence of Trypanosoma evansi infections in village buffaloes in Central Java were estimated using parasitological tests, two antigen detection ELISAs (2G6 Ag-ELISA and Tr7 Ag-ELISA), an antibody-detection ELISA (IgG ELISA) and a card agglutination test (CATT). Of 2387 village buffaloes tested in five districts, 4 % (95 % confidence interval [CI]: 3 %, 5 %) were positive with the microhaematocrit test (MHCT), 58 % (95 % CI: 56 %, 60 %) were positive with the 2G6 Ag-ELISA and 70 % (95 % CI: 68 %, 72 %) were positive with the Tr7 Ag-ELISA. An increasing prevalence with age was found and the proportion of positive buffaloes was highest in the over 84 months-old age-group (68 %) with the 2G6 Ag-ELISA and in the 37-60 months-old age-group (78 %) with the Tr7 Ag ELISA. Parasitaemic buffaloes were found in more than half of the villages visited. Corrected village-specific prevalence values obtained with the two Ag ELISAs ranged from 0% to over 100%, and prevalence differed significantly (P < or = 0.0001) between villages in four of the five districts. Overall, 10% of buffaloes tested in markets were found to be parasitaemic and 39, 56 and 47 % were found positive with the 2G6 Ag-ELISA, IgG ELISA and CATT, respectively. Incidence rates varied according to the test used and ranged from 0.22 (95 % CI: 0.09, 0.44) to 0.44 (95 % CI: 0.24, 0.76), per animal-year at risk, in two villages. The results highlight the importance of using validated diagnostic tests to obtain accurate estimates of prevalence and incidence. These parameters are needed, for example in mathematical models, for the development and evaluation of different control strategies for T. evansi infections in buffaloes. PMID- 10722145 TI - Introduction, persistence and fade-out of porcine reproductive and respiratory syndrome virus in a Dutch breeding herd: a mathematical analysis. AB - The objective of this study was to investigate the dynamics of PRRSV infection and to quantify transmission within a breeding herd, and its impact on herd performance. For this purpose a longitudinal study was performed in a closed breeding herd of 115 sows. Statistical methods and Monte Carlo simulations based on stochastic SIR models were used to analyse the observational data. Moreover, a case-control study was performed to determine whether seroconversion of sows during gestation was associated with aberrant litters. The transmission parameter R was estimated to be 3.0 (95% confidence interval 1.5-6.0) for the model version based on the most plausible assumptions that the infectious period lasts 56 days and no lifelong immunity exists after infection. Based on simulations using a breeding herd of equal size the average time-to-extinction was estimated to be 6 years; using a herd of twice the size, it was 80 years. Furthermore, in contrast to the epidemic phase of the disease, the endemic phase was not detrimental to herd performance. PMID- 10722146 TI - Immobilised activated sludge based biosensor for biochemical oxygen demand measurement. AB - A biochemical oxygen demand (BOD) sensor, based on an immobilised mixed culture of microorganisms in combination with a dissolved oxygen electrode, has been developed for the purpose of on-line monitoring of the biological treatment process for waste and wastewater. The sensor was designed for easy replacement of the biomembrane, thereby making it suitable for short-term use. The drawbacks of activated sludge based sensor, such as short sensor lifetime, were thereby circumvented. The sensor BOD measurements were carried out in the kinetic mode using a flow injection system, resulting in 25 s for one measurement followed by 4-8 min recovery time. Based on the results of normalised sensor responses, the OECD synthetic wastewater was considered to be a more suitable calibration solution in comparison with the GGA solution. Good agreement was achieved between the results of the sensor BOD measurement and those obtained from BOD5 analysis of a wastewater sample from a food-processing factory. Reproducibility of responses using one sensor was below +/- 5.6%, standard deviation. Reproducibility of responses using different sensors was within acceptable bias limits, viz. +/- 15% standard deviation. PMID- 10722147 TI - Fiber optic monitoring of carbamate pesticides using porous glass with covalently bound chlorophenol red. AB - An optical fiber biosensor for the determination of the pesticides propoxur (Baygon) and carbaryl, two of the most commonly used carbamate insecticides in vegetable crops, is described. A pH indicator, chlorophenol red, is used as optical transducer of the inhibition of the enzyme acetylcholinesterase by the analytes. The biorecognition element is covalently immobilized onto controlled pore glass beads (CPG) and packed in a thermostatized bioreactor connected to a flow-through cell that contains CPG-immobilized chlorophenol red placed at the common end of a bifurcated fiber optic bundle. In the presence of a constant acetylcholine concentration, the colour of the pH sensitive layer changes and the measured reflectance signal can be related to the carbamate concentration in the sample solution. The performance of the biosensor has been optimized using a flow injection system. The linear dynamic range for the determination of carbaryl and propoxur spans from 0.8 to 3.0 mg l(-1) and from 0.03 to 0.50 mg l(-1), respectively. The detection limit (3 s) of the biosensor for propoxur (0.4 ng) is lower than that measured for carbaryl (25 ng). Reproducibility, stability and interference studies of the optical device are reported. The biosensor has been applied to the determination of propoxur in spiked vegetables (onion and lettuce) using ultrasound extraction, achieving recovery values between 93 and 95% for onion samples at the different concentration levels assayed. PMID- 10722148 TI - Conductimetric membrane strip immunosensor with polyaniline-bound gold colloids as signal generator. AB - For point-of-care examination, an immuno-chromatographic assay system based on conductimetric detection was investigated by utilizing, as signal generator, colloidal gold with polyaniline bound on the metal surface. Although the gold is a widely used label for antibodies to produce colorimetric signals, the tracer does not lend itself for a suitable electric conduction along the gold particles due to the presence of protein barriers (e.g. immunoglobulin and blocking agent) against electron transfer. To overcome this problem, we introduced a conducting polymer, for instance, polyaniline, as a conductivity-modulating agent on the gold surface after immobilizing an antibody specific to human albumin used as model analyte. This novel signal generator amplified the conductimetric signal 4.7 times compared with the plain gold, and the signal was also maximum 2.3-fold higher than that from the photometric system under the same analytical conditions. The latter effect resulted from an exponential pattern in the dose response curve of the electric signal that was different from the conventional sigmoidal shape. PMID- 10722150 TI - Current awareness in biosensors & bioelectronics. PMID- 10722149 TI - AC voltammetric carbon paste-based enzyme immunosensors. AB - Carbon paste electrodes, previously anodised in a basic media, are the basis for the development of a new voltammetric immunosensor device. Passive adsorption of the appropriate immunochemical reagent was performed onto the electrode surface. Alkaline Phosphatase labelled immunoglobulin was the tracer used in this work, 3 indoxyl phosphate being a very suitable enzymatic substrate for the electrochemical detection of the corresponding affinity reaction. The hydrolysis of this molecule generates indigo dimmer. This product was detected by alternating current voltammetry taking advantage of the adsorptive and inherent electrodic properties that it exhibits. The same electrochemical anodisation was used at the end of one assay to remove the entire protein layer attached to the carbon paste surface, allowing the formation of a new sensing phase and the use of the same support in several consecutive experiments. The methodology was applied to the design of two different immunoassays for the determination of human IgG. Good reproducibility of the electrodic signal and a limit of detection around 10(-10) M were achieved. PMID- 10722151 TI - Synthesis and hybridization properties of DNA-PNA chimeras carrying 5-bromouracil and 5-methylcytosine. AB - The preparation of 5-bromouracil and 5-methylcytosine peptide nucleic acid (PNA) monomers is described. These PNA monomers were used for the preparation of several DNA-PNA chimeras and their hybridization properties are described. The substitution of cytosine by 5-methylcytosine in DNA-PNA chimeras increased duplex stability while substitution of thymine by 5-bromouracil maintained it. Moreover, binding of DNA-PNA chimeras to double-stranded DNA to form triple helices was studied. In contrast to DNA, the presence of 5-methylcytosine and 5-bromouracil in DNA-PNA chimeras destabilized triple helix. PMID- 10722152 TI - Steroidal derived acids as inhibitors of human Cdc25A protein phosphatase. AB - A group of steroidal derived acids were synthesized and found to be human Cdc25A inhibitors. Their potency ranged from 1.1 to > 100 microM; the best ones compare very favorably with that of the novel cyano-containing 5,6-seco-cholesteryl acid 1 (IC50=2.2microM) reported by us recently (Peng, H.; Zalkow, L. H.; Abraham, R. T.; Powis, G. J. Med. Chem. 1998, 41, 4677). Structure-activity relationships of these compounds revealed that a hydrophobic cholesteryl side chain and a free carboxyl group are crucial for activity. The distance between these two pharmacophores is also important for the potency of these compounds. Several of the compounds showed selective growth inhibition effects in the NCI in vitro cancer cell line panel. PMID- 10722153 TI - Synthesis of C2-symmetric guanidino-sugars as potent inhibitors of glycosidases. AB - A series of enantiomerically pure C2-Symmetric guanidino-sugars was synthesized from D-mannitol. The first method described involves direct opening of a bis epoxide by guanidine, whereas the second one deals with a mercury-catalyzed transformation of a cyclic thiourea into a N,N',N"-trisubstituted guanidine as a key step. The biological activity of these compounds towards several glycosidases has been evaluated. One of them (5) was found to selectively inhibit alpha-L fucosidase of bovin kidney (2.8 microM). PMID- 10722154 TI - Benzamides derived from 1,2-diaminocyclopropane as novel ligands for human D2 and D3 dopamine receptors. AB - Benzamides (3a-f) derived from 4-amino-5-chloro-2-methoxybenzoic acid and either cis or trans 1,2-diaminocyclopropane were synthesised and were evaluated in binding assays employing, bovine striatal D2 receptors, recombinant human hD2 and hD3 receptors expressed in CHO cells and rat, cortical 5-HT3 and striatal 5-HT4 receptors. The cis and trans isomers of the derivatives were isolated and characterised. The results demonstrated the superiority of the cis conformers over the trans conformers in dopamine receptor binding assays (Ki hD2 = 13.4 and 6.9 nM and Ki hD3 = 17.7 and 4.5 nM for the cis-3b and cis-3f compounds, respectively; Ki hD2 = 816 and >l000 nM and Ki hD3 = 469 and >1000 nM for the corresponding trans-3b and trans-3f compounds respectively). The cis compounds are folded: the benzamide group and the basic nitrogen atom were in a syn relationship. Compound 3f can be superimposed with a conformation of the tropane derivative, BRL 25594, having the benzyl group in an axial position to give a suitable fit, indicating that both compounds may have a common binding site in the dopamine receptor. PMID- 10722155 TI - Density functional and Ab initio studies on N-acetylduocarmycin SA: insight into its DNA interaction properties. AB - Density functional (DF) and Moller-Plesset second order perturbation (MP2) calculations were carried out on N-acetylduocarmycin SA (N-Ac-DSA), an analogue of a series of potent antitumor antibiotics that include the duocarmycins. These computational methods were used to investigate the degree of ground state destabilization of duocarmycins that would result upon binding to DNA. Ground state destabilization has been proposed as the origin of the ligand's enhanced rate of alkylation by more than a millionfold. The conformations of the 'Unbound' and 'DNA-Bound' N-Ac-DSA were generated using available geometric data for duocarmycin SA. Specifically, the dihedral angles chi1/chi2 were locked at 6.9 degrees/4.5 degrees for the Unbound and 22.0 degrees/11.0 degrees for the Bound form. The structures were optimized using DF theory, with subsequent MP2 calculations to improve the electronic energies. All of the calculations were performed on the unprotonated (1) as well as the C6-carbonyl protonated form (2). The results showed that the ground state destabilization energies of the Unbound and Bound forms, for the unprotonated and protonated series, were fairly small (< 0.8 kcal/mol). Similarly, the difference in the electronic nature of the Unbound and Bound forms, as indicated by changes in bond lengths and charge density, were also small. In summary, it appears that twisting of two key torsional angles, the concomitant ground state destabilization, and C6-carbonyl protonation may not fully account for the significant rate increase of adenine-N3 alkylation upon binding to DNA. PMID- 10722156 TI - Large scale, liquid phase oligonucleotide synthesis by alkyl H-phosphonate approach. AB - A new approach to the liquid phase synthesis of oligonucleotide is described, it is based on oxidative coupling using alkyl H-phosphonate synthon and polyethylene glycol (PEG5000) as a soluble support. Nucleoside alkyl H-phosphonate undergoes oxidative coupling in presence of NBS. The use of polyethylene glycol as a soluble polymeric support preserves some convenient features of the solid phase synthesis with new interesting advantages. This liquid phase method appears effective in terms of speed and coupling yield and can be evaluated for the production of large amount of oligonucleotide (100 microM). PMID- 10722157 TI - gem-diamine 1-N-iminosugars of L-fucose-type, the extremely potent L-fucosidase inhibitors. AB - An efficient route from D-ribono-gamma-lactone to gem-diamine 1-N-iminosugars of L-fucose-type, a new family of glycosidase inhibitor, has been developed in a formation of a gem-diamine 1-N-iminopyranose ring by the Mitsunobu reaction of an aminal as a key step. The analogues were proved to be the extremely potent inhibitors against alpha-L-fucosidase (IC50 approximately 3 ng mL(-1), Ki approximately 5 x 10(-9) M). The present study has shown that a cyclic methanediamine generated in media affects glycosidases as a real active-form of the gem-diamine 1-N-iminosugars of L-fucose-type. PMID- 10722158 TI - Synthesis and structure-activity relationships of a new class of selective EP3 receptor agonist, 13,14-didehydro-16-phenoxy analogues of prostaglandin E1. AB - A series of 13,14-didehydro-16-phenoxy analogues of prostaglandin E1 was synthesized and their agonistic activity on EP receptor subtypes was evaluated. 13,14-Didehydro-16-phenoxy-1-decarboxy analogues, 7e and 7f, display highly selective activity on the EP3 receptor subtype, thus, their utility as a selective anti-ulcer agent can be expected. PMID- 10722159 TI - Syntheses and cytotoxicity evaluation of bis(indolyl)thiazole, bis(indolyl)pyrazinone and bis(indolyl)pyrazine: analogues of cytotoxic marine bis(indole) alkaloid. AB - 2,4-Bis(3'-indolyl)thiazoles, 3,5-bis(3'-indolyl)-2(1H)pyrazinone and 3,6-bis(3' indolyl)pyrazine were synthesized and evaluated for cytotoxic activity against diverse human cancer cell lines by the National Cancer Institute. These compounds demonstrated significant inhibitory effects in the growth of a range of cancer cell lines. 2,4-Bis(3'-indolyl)thiazole displayed selective cytotoxicity against certain leukemia cell lines with GI50 values in the low micromolar range while the substituted derivatives showed a broad spectrum of cytotoxic activity. 3,5 Bis(3'-indolyl)-2(1H)pyrazinone and 3,6-bis[3'-(N-methyl-indolyl)]pyrazine possessed strong inhibitory activity against a wide range of human tumor cell lines. The mechanism of action remained unknown. The results suggested that 2,4 bis(3'-indolyl)thiazoles, 3,5-bis(3'-indolyl)-2(1H)pyrazinone and 3,6-bis[3'-(N methyl-indolyl)] pyrazine offer potential as lead compounds for the discovery of anticancer agents. PMID- 10722161 TI - Synthesis and antitumor activity of duocarmycin derivatives: modification at C-8 position of A-ring pyrrole compounds bearing the simplified DNA-binding groups. AB - A series of the 8-O-substituted A-ring pyrrole derivatives of duocarmycin bearing the simplified DNA-binding moieties such as cinnamoyl or heteroarylacryloyl groups were synthesized, and evaluated for in vitro anticellular activity against HeLa S3 cells and in vivo antitumor activity against murine sarcoma 180 in mice. In addition, the stability of the 8-O-substituted analogues in aqueous solution and the conversion to their active form (cyclopropane compound) from the 8-O substituted analogues in mice or human serum were examined. The 8-O-substituted A ring pyrrole derivatives bearing the simplified DNA-binding moieties showed remarkably potent in vivo antitumor activity and low peripheral blood toxicity compared with the 8-O-substituted A-ring pyrrole derivatives having the trimethoxyindole skeleton in segment-B (Seg-B), which were equal to 8-O-[(N methylpiperazinyl)carbonyl] derivatives of 4'-methoxycinnamates and 4'-methoxy beta-heteroarylacrylates. Moreover, among 8-O-substituted analogues, several compounds can be chemically or enzymatically converted to their active form in human serum. This result indicated that new 8-O-substituted derivatives were different prodrugs from KW-2189 and 8-O-substituted analogues being the same type of prodrug as KW-2189. PMID- 10722160 TI - Synthesis and biological activities of novel antiallergic agents with 5 lipoxygenase inhibiting action. AB - Novel benzimidazole derivatives were synthesized and their pharmacological activities were examined. These compounds showed a good suppressive action on histamine release from rat peritoneal mast cells produced by antigen-antibody reaction, an antagonistic action on guinea pig ileum contraction caused by histamine, an inhibitory action on 5-lipoxygenase in rat basophilic leukemia-1 (RBL-1) cells, and a preventive action on NADPH dependent lipid peroxidation induced by Fe3+-ADP in rat liver microsomes. In addition, 1-[2-[2-(4-Hydroxy 2,3,5-trimethylphenoxy)ethoxy]-ethyl]-2-(4-meth yl-1-homopiperazino)-1H benzimidazole difumarate (BOM1006) exhibited a dose dependent suppressive action on 48 h homologous passive cutaneous anaphylaxis (PCA) reaction in rats orally administered the drug. PMID- 10722162 TI - Novel potassium channel openers. Part 4: transformation of the 1,4-benzoxazine skeleton into 1,4-benzothiazine, 1,2,3,4-tetrahydroquinoline, 1,2,3,4 tetrahydroquinoxaline, indoline, and 1,5-benzoxazepine. AB - As part of a search for a new potassium channel opener, the 1,4-benzoxazine skeleton derived from the benzopyran skeleton of cromakalim, was transformed into other fused rings such as 1,4-benzothiazine, 1,2,3,4-tetrahydroquinoline, 1,2,3,4 tetrahydroquinoxaline, indoline, and 1,5-benzoxazepine. The 1,4-benzothiazine derivative displayed approximately 20 times more potent vasorelaxant activity than cromakalim. PMID- 10722163 TI - para-Substituted N-nitroso-N-oxybenzenamine ammonium salts: a new class of redox sensitive nitric oxide releasing compounds. AB - N-Nitroso-N-oxybenzenamine ammonium salts with -OMe, -Me, -H, -F, -Cl, -CF3, and SO2Me substituents at the para position of the phenyl ring constitute a new class of-redox sensitive nitric oxide (NO) releasing compounds. These compounds yield nitric oxide and the corresponding nitrosobenzene derivatives by a spontaneous dissociation mechanism after undergoing a one electron oxidation. Oxidation of these compounds can be achieved through chemical, electrochemical and enzymatic methods. It was observed electrochemically that the amount of NO generated was dependent on the substituent effect and the applied oxidation potential. Electron withdrawing substituents increase the oxidation potential of the compound. A linear correlation was observed when the peak potentials for the oxidation were graphed versus the Hammett substituent constant. Density functional theory calculations were also performed on this series of compounds. The theoretical oxidation energies of the corresponding anions show a strong linear correlation with the experimental potentials. Furthermore, enzymatic oxidation using horseradish peroxidase showed a similar substituent effect. These results indicate that substitution at the para position of the phenyl ring has a profound effect on the stability, oxidation potential and enzymatic kinetic properties of the compounds. Thus para-substituted N-nitroso-N-oxybenzenamine salts comprise a new class of redox-sensitive nitric oxide releasing agents. PMID- 10722164 TI - An investigation of antibody acyl hydrolysis catalysis using a large set of related haptens. AB - An aspect of catalytic antibody research that receives little attention in the literature involves hapten systems that fail to elicit antibody catalysts despite a high affinity immune response and hapten designs that resemble those known to elicit catalysts. We have investigated a series of 12 phosphate and phosphonate haptens in a total of three animal systems. Dramatic and reproducible differences were observed in the catalytic activities of polyclonal antibodies elicited by the different haptens. A phosphate hapten with a phenyl ring on the side of the hapten opposite the linker elicited reproducibly high levels of polyclonal antibody catalytic activity. The other 11 haptens, most with benzyl groups on the side of the hapten opposite the linker, elicited immune responses in which catalytic activity was significantly weaker in terms of the level of observed catalytic activity, as well as frequency of elicited catalysts. Our results indicate that subtle features of transition state analogue hapten structure can have a dramatic and reproducible influence over the catalytic activity of elicited antibodies in related haptens. Whatever the explanation, subtle changes in mechanistic features due to altered leaving group ability/location or overall hapten flexibility, the comprehensive data presented here indicate that phenyl or 4-nitrophenyl leaving groups located opposite the hapten linker are to be preferred in order to elicit highly active antibody catalysts for acyl hydrolysis reactions. PMID- 10722165 TI - Paclitaxel esters of malic acid as prodrugs with improved water solubility. AB - The synthesis of paclitaxel esters of malic acid is described. These compounds were found to have improved water solubility and are stable in solution at neutral pH. The C2' modified compounds behave as prodrugs, that is, paclitaxel is generated upon exposure to human plasma, whereas the C7 modified derivatives do not. 2'-Malyl paclitaxel sodium salt demonstrated enhanced antitumour activity and less toxicity in a P388 murine leukaemia in vivo model when compared to paclitaxel. PMID- 10722166 TI - The reactivity of phenolic and non-phenolic residual kraft lignin model compounds with Mn(II)-peroxidase from Lentinula edodes. AB - Three phenolic model compounds representing bonding patterns of residual kraft lignin were incubated with manganese peroxidase from Lentinula edodes. Extensive degradation of all the phenolic models, mainly occurring via side-chain benzylic oxidation, was observed. Among the tested model compounds the diphenylmethane alpha-5 phenolic model was found to be the most reactive, yielding several products showing oxidation and fragmentation at the bridging position. The non phenolic 5-5' biphenyl and 5-5' diphenylmethane models were found unreactive. PMID- 10722167 TI - Synthesis and pharmacology of a hybrid cannabinoid. AB - A pentacyclic hybrid cannabinoid (4) has been synthesized, which combines structural elements of traditional cannabinoids and cannabmimetic indoles. Cannabinoid 4 contains a 1-pentylindole structure fused to the 2,3-positions of the partially reduced hydroxydibenzopyran system of THC. The successful approach to 4 employed 9-benzoyl-5,7-dimethoxy-1,2,3,4-tetrahydrocarbazole (17) as the starting material. Dehydrogenation to carbazole 18, followed by demethylation and condensation with trans-p-menthadienol gave N-benzoyl hybrid cannabinoid 22, N alkylation of which afforded target cannabinoid 4. The hybrid cannabinoid had affinity for the CB1 receptor approximately equal to that of delta8-THC (Ki = 19.3+/-3 nM), and shows comparable potency in vivo. PMID- 10722168 TI - Synthesis and in vitro muscarinic activities of a series of 3-(pyrazol-3-yl)-1 azabicyclo[2.2.2]octanes. AB - A series of 3-(pyrazol-3-yl)-1-azabicyclo[2.2.2]octane derivatives C (Fig. 1) was synthesized and tested for muscarinic activity in receptor binding assays using [3H]-oxotremorine-M (OXO-M) and [3H]-pirenzepine (PZ) as ligands. Potential muscarinic agonistic or antagonistic properties of the compounds were determined using binding studies measuring their potencies to inhibit the binding of OXO-M and PZ. Preferential inhibition of OXO-M binding was used as an indicator for potential muscarinic agonistic properties; this potential was confirmed in functional studies on isolated organs. PMID- 10722169 TI - Elucidation of strict structural requirements of brefeldin A as an inducer of differentiation and apoptosis. AB - Brefeldin A (BFA) can induce a wide variety of human cancer cells to differentiation and apoptosis and is in development as an anticancer agent. To elucidate structural requirements for cytotoxicity and induction of differentiation and apoptosis, BFA was structurally modified into various derivatives including 4-epi-BFA in this study. Their inducing activities of apoptosis were evaluated with their cytotoxicities, DNA fragmentation and morphological changes in human colon cancer cell HCT 116. The cytotoxicity of 4 epi-BFA (TX-1923) (IC50 = 60 microM) was 300 times lower than that of BFA (IC50 = 0.2 microM). Furthermore, 4-epi-BFA induced DNA fragmentation and apoptotic morphological changes at much higher concentrations (70 and 50 microM, respectively) than BFA (0.11 and 0.36 microM, respectively). These results indicated that the configuration of 4-hydroxyl group of brefeldin A plays a key role in the cytotoxicity and induction of apoptosis. On the other hand, 7-O acetyl-BFA, 4-O-acetyl-BFA, and 4,7-di-O-acetyl-BFA exhibited potential activities in cytotoxicity and inducibility of apoptosis. We suggested that the structural determinants for BFA include the moiety of the Michael acceptor, the conformational rigidity of the 13-membered ring, and the configuration of 4 hydroxyl group. PMID- 10722170 TI - DNA-binding peptides searched from the solid-phase combinatorial library with the use of the magnetic beads attaching the target duplex DNA. AB - We have exhibited successful and rapid screening of DNA-binding peptide ligands from solid-phase library beads with the use of the target DNA-conjugated magnetic beads. The target duplex DNA (3) has a polyether linker between two complementary sequences (T4A3G-ether linker-CT3A4) and is stable in the duplex form during the selection procedure. Finally, 71 pentapeptide sequences were identified from the solid-phase pentapeptide library. From an analysis of the peptide sequences identified in this study, it has been revealed that peptide ligands contain hydrophobic amino acids as the major component. The synthetic peptides with identified sequences and a combination of the major components have exhibited moderate to high binding affinity to the duplex DNA in competition experiments with ethidium-DNA complexes. PMID- 10722171 TI - Personality and personality disorder: current issues and directions. PMID- 10722172 TI - Does old age reduce the risk of anxiety and depression? A review of epidemiological studies across the adult life span. AB - BACKGROUND: There is considerable disagreement about what happens to the risk of anxiety and depression disorders and symptoms as people get older. METHODS: A search was made for studies that examine the occurrence of anxiety, depression or general distress across the adult life span. To be included, a study had to involve a general population sample ranging in age from at least the 30s to 65 and over and use the same assessment method at each age. RESULTS: There was no consistent pattern across studies for age differences in the occurrence of anxiety, depression or distress. The most common trend found was for an initial rise across age groups, followed by a drop. Two major factors producing this variability in results were age biases in assessment of anxiety and depression and the masking effect of other risk factors that vary with age. When other risk factors were statistically controlled, a more consistent pattern emerged, with most studies finding a decrease in anxiety, depression and distress across age groups. This decrease cannot be accounted for by exclusion of elderly people in institutional care from epidemiological surveys or by selective mortality of people with anxiety or depression. CONCLUSION: There is some evidence that ageing is associated with an intrinsic reduction in susceptibility to anxiety and depression. However, longitudinal studies covering the adult life span are needed to distinguish ageing from cohort effects. More attention needs to be given to understanding the mechanism behind any ageing-related reduction in risk for anxiety and depression with age. Possible factors are decreased emotional responsiveness with age, increased emotional control and psychological immunization to stressful experiences. PMID- 10722173 TI - Risk factors and life processes associated with the onset of suicidal behaviour during adolescence and early adulthood. AB - BACKGROUND: This study examined associations between childhood circumstances, adolescent mental health and life events, and the development of suicidal behaviour in young people aged between 15 and 21 years. METHOD: Data were gathered over the course of a 21-year longitudinal study of a birth cohort of 1265 children born in New Zealand. The measures collected included: (1) patterns of suicidal behaviour (ideation, attempt) (15-21 years); (2) social background, family functioning, parental and individual adjustment during childhood (0-16 years); and (3) time dynamics of mental health and stressful life events during adolescence and early adulthood (15-21 years). RESULTS: By the age of 21 years, 28.8% of the sample reported having thought about killing themselves and 7.5% reported having made a suicide attempt. The childhood profile of those at greatest risk of suicidal behaviour was that of a young person reared in a family environment characterized by socio-economic adversity, marital disruption, poor parent-child attachment and exposure to sexual abuse, and who as a young adolescent showed high rates of neuroticism and novelty seeking. With the exception of the socio-economic and personality measures, the effects of childhood factors were largely mediated by mental health problems and exposure to stressful life events during adolescence and early adulthood. Mental health problems including depression, anxiety disorders, substance use disorder, and to some extent conduct disorder, in addition to exposure to adverse life events, were significantly associated with the onset of suicidal behaviours. CONCLUSIONS: Findings support a life course model of the aetiology of suicidal behaviour in which risk of developing suicidal behaviour depends on accumulative exposure to a series of social, family, personality and mental health factors. PMID- 10722174 TI - Early sexual abuse and lifetime psychopathology: a co-twin-control study. AB - BACKGROUND: This study was designed to determine lifetime prevalence of psychiatric disorders among twins who reported childhood sexual abuse (CSA), and to compare these rates with those among non-abused co-twins. The contribution of familial and individual-specific factors to reported sexual abuse was also examined. METHOD: Information about lifetime psychopathology and substance use was obtained by structured telephone interviews with 5995 Australian twins. Twins who reported a history of childhood sexual abuse (CSA) were contrasted on lifetime psychopathology with subjects without such a history; in addition, comparisons were made between same-sex twin pairs discordant for CSA. RESULTS: A history of CSA was reported by 5.9% of the women and 2.5% of the men. In the sample as a whole, those reporting CSA were more likely to receive lifetime diagnoses of major depression, conduct disorder, panic disorder and alcoholism, and were more likely to report suicidal ideation and a history of suicide attempt. Abused women, but not men, were also more likely to report social phobia. When comparisons were restricted to non-abused co-twins, no differences in psychopathology were seen. However, rates of major depression, conduct disorder and suicidal ideation were higher if both co-twins were abused than if the respondent alone reported CSA. Model-fitting indicated that shared environmental factors influenced risk for reported CSA in women, but not in men. CONCLUSION: The association between CSA and psychopathology arises at least in part through the influence of shared familial factors on both risk of victimization and risk of psychopathology. PMID- 10722175 TI - The childhood and family background of women with clinical eating disorders: a comparison with women with major depression and women without psychiatric disorder. AB - BACKGROUND: Childhood antecedents are often put forward as being of possible aetiological significance for both anorexia nervosa and bulimia nervosa. METHOD: Comparisons were made of groups of women with eating disorders with groups of women with major depression or without current psychiatric disorder, using the Childhood Experience of Care and Abuse interview (CECA). RESULTS: Women with bulimia nervosa (or mixed bulimia and anorexia nervosa) tended to report more troubled childhood experiences than did women from the non-morbid comparison group. In this respect, they resembled those with major depression. In contrast, those with anorexia nervosa resembled the non-morbid women rather than the other psychiatric groups. CONCLUSIONS: Adversity in childhood as measured by the CECA may play a part in the causation of bulimia nervosa but not of anorexia nervosa. It remains possible that more specific or subtle family influences may be relevant. PMID- 10722176 TI - Quality of rearing practices as predictor of short-term outcome in adolescent anorexia nervosa. AB - BACKGROUND: Studies of family relationships in anorexia nervosa have produced conflicting results. Some authors claim that family factors are related to short term outcomes. METHODS: Perceived rearing practices, as measured by the EMBU (Egna Minnen Betraffande Uppfostran: 'My memories of Upbringing') were examined in a sample (N = 158) of adolescents with anorexia nervosa and compared with the perceptions of adolescents (N = 159) from the general population. A further comparison was made between the groups of patients with good and bad short-term outcomes. Logistic regression analysis was performed to evaluate the predictive value of different variables on short-term outcome. RESULTS: Overall, small differences were observed in the perceptions of rearing practices as expressed by the controls and the anorexic patients. Patients with bad short-term outcome perceived more rejection and control-overprotection from both parents than those with good outcome. In the logistic regression analysis only Rejection from father and the EAT (Eating Attitudes Test) total score gave independent prediction of treatment response. CONCLUSIONS: Taken as a whole, these results do not support the idea of altered rearing practices in anorexic patients, at least in young patients with a short evolution of the disease. Perceived rearing practices, especially 'rejection', appear to have an appreciable effect on the short-term outcome. PMID- 10722177 TI - The association between childhood feeding problems and maternal eating disorder: a community study. AB - BACKGROUND: A possible association between childhood feeding problems and maternal eating disorder has been suggested by a clinic-based self-report questionnaire study. A community study was conducted, using standardized psychiatric interviews, to investigate the strength and specificity of this putative association. METHODS: Four-year-old children were screened using a self report version of the Behaviour Screening Questionnaire, completed by mothers, and the Pre-School Behaviour Checklist, completed by teachers. Three groups of children were identified for follow-up: children with feeding problems (N = 42), children with a non-feeding form of disturbance (i.e. shyness, fearfulness or behavioural disturbance; N = 79), and a random sample of children with no disturbance (N = 29). The presence of feeding problems was confirmed by assessment of a filmed family meal, with ratings made blind to child group and maternal mental state. Maternal current and past affective disorder and current and past eating disorder were systematically assessed, blind to child status, using the Anxiety Disorders Interview Schedule and the Eating Disorder Examination respectively. RESULTS: Compared with the mothers of the two comparison groups of children, the mothers of the children with feeding problems had no raised rate of any affective disorder, either current or past, but they did have a markedly raised rate of both current and past DSM-IV eating disorder. The odds ratio of maternal eating disorder for the children with feeding problems was significantly raised at 11.1 (CI 1.4-91.8). CONCLUSION: There is a strong and specific association between childhood feeding problems and maternal eating disorder. PMID- 10722178 TI - Effects of rapid tryptophan depletion in patients with seasonal affective disorder in natural summer remission. AB - BACKGROUND: Serotonergic mechanisms have been proposed for the pathophysiology of seasonal affective disorder (SAD) and the therapeutic effect of bright-light treatment. Previously, we showed that SAD patients, in clinical remission with light therapy during the winter, experienced transient depressive relapses after a rapid tryptophan depletion (RTD) technique, which results in decreased brain serotonin levels. The objective of this study was to investigate the effect of RTD in SAD patients who were in natural summer remission. METHODS: Twelve drug free patients with SAD by DSM-IV criteria and 10 normal subjects participated in this double-blind, placebo-controlled, crossover study. SAD patients were in natural summer remission for at least 8 weeks. Behavioural ratings and plasma tryptophan levels were obtained before, and 5 h after, ingesting an amino acid (AA) mixture +/- tryptophan. Experimental RTD and control sessions were scheduled 1 week apart. RESULTS: The RTD session resulted in significant reduction in total and free plasma tryptophan levels compared to the control session. The behavioural data were analysed using repeated measures analysis of variance. This analysis found significant main effects of time (higher scores after AA ingestion) and diagnosis (higher scores in SAD patients), but no main effect of session or significant interaction effects between the three factors. Thus, there were no significant behavioural effects of RTD compared to the sham depletion control session. CONCLUSIONS: The summer remission experienced by SAD patients is not dependent on plasma tryptophan levels (and presumably brain serotonin function) in the same manner as that of remission after light therapy. These results conflict with those of other laboratories, perhaps because of differences in study samples. PMID- 10722179 TI - Safety of fluoxetine during the first trimester of pregnancy: a meta-analytical review of epidemiological studies. AB - BACKGROUND: This study was designed to examine whether there is an increased risk for major malformations following the use of fluoxetine during the first trimester of pregnancy. METHODS: Published and unpublished reports were identified through computerized and manual searches of bibliographical databases, reference lists from primary articles, and letters to editors, agencies, foundations and content experts. Meta-analysis was undertaken of prospective controlled and uncontrolled studies on the use of fluoxetine during first trimester of pregnancy. RESULTS: The pooled relative risk and 95% confidence interval for major malformations does not suggest an association between the use of fluoxetine during the first trimester and an increased risk of major malformations. Combination of controlled and uncontrolled studies shows a weighted risk of 26% (95% CI 1-4.2%). The summary odds ratio from the two controlled studies (OR = 1.33, 95% CI 0.49-3.58) was not significant. Homogeneity testing shows that the effect sizes are similar throughout all studies. Power analysis indicates that 26 controlled studies of similar size, would be required, to reverse this finding. CONCLUSIONS: The use of fluoxetine during the first trimester of pregnancy is not associated with measurable teratogenic effects in human. PMID- 10722180 TI - A multicentre, double-blind, randomized comparison of quetiapine (ICI 204,636, 'Seroquel') and haloperidol in schizophrenia. AB - BACKGROUND: Quetiapine (ICI 204,636, 'Seroquel') is a new atypical antipsychotic agent with a similar binding profile to the original atypical antipsychotic, clozapine. Its clinical efficacy has already been demonstrated at multiple fixed doses (150-750 mg/day) and has been suggested to be comparable with haloperidol (12 mg/day). METHODS: This international, 6-week, multicentre, double-blind, randomized, parallel-group trial compared quetiapine with haloperidol (455 mg and 8 mg mean total daily doses, respectively) in 448 hospitalized patients with acute exacerbation of chronic or subchronic schizophrenia (DSM-III-R), in order to establish their equivalence in terms of efficacy, and the nature of their tolerability profiles, especially in terms of extrapyramidal symptoms (EPS) and serum prolactin levels. RESULTS: Both quetiapine and haloperidol produced a clear reduction in the Positive and Negative Syndrome Scale (PANSS) scores and Clinical Global Impression (CGI) Severity of Illness and Global Improvement scores. At day 42, the PANSS total score was reduced by -18.7+/-1.63 in the quetiapine group, and -22.1+/-1.63 in the haloperidol group (P = 0.13, between-treatment). Quetiapine was better tolerated than haloperidol in terms of EPS as demonstrated by the significant differences in the Simpson Scale and Abnormal Involuntary Movement Scale scores (P < 0.05). Although patients in both groups had elevated serum prolactin concentrations at baseline, mean serum prolactin concentration decreased (by 16.5 microg/l) in quetiapine-treated patients, yet increased (by 5.9 microg/l) in patients treated with haloperidol. CONCLUSION: Quetiapine is an effective and well tolerated antipsychotic of comparable efficacy to haloperidol and lacks the latter compound's effect on prolactin and EPS. PMID- 10722181 TI - Altered brain energy metabolism in lithium-resistant bipolar disorder detected by photic stimulated 31P-MR spectroscopy. AB - BACKGROUND: Previous 31P-MRS (magnetic resonance spectroscopy) studies suggested altered brain energy metabolism in bipolar disorder. This study characterized brain energy metabolism in lithium-resistant bipolar disorder using the photic stimulation paradigm. METHODS: Subjects were 19 patients with DSM-IV bipolar disorder (nine responders and 10 nonresponders, 13 with bipolar I and six with bipolar II) in the euthymic state and 25 healthy volunteers. Energy metabolism in the occipital region was examined by 31P-MRS during photic stimulation (PS). Six 31P-MR spectra were obtained, one was before PS (Pre), two during 12 min of PS (PS1, PS2), and three after the PS (Post 1, Post 2, Post 3). RESULTS: Significant effect of diagnosis (lithium-responsive bipolar disorder, lithium-resistant bipolar disorder, and control) was found for the phosphocreatine peak area ratio during the course of the photic stimulation (P < 0.05 by repeated measures ANOVA). The phosphocreatine peak area ratio was significantly decreased at Post 1 and Post 2 compared with Pre in lithium-resistant bipolar patients (P = 0.01 and P = 0.01 by Dunnett's multiple comparison). CONCLUSIONS: The finding that phosphocreatine decreased after photic stimulation may be compatible with mitochondrial dysfunction. It is possible that mitochondrial function is impaired in lithium-resistant bipolar disorder. PMID- 10722182 TI - Quantitative MRI of the hippocampus and amygdala in severe depression. AB - BACKGROUND: There is little evidence to support possible structural changes in the amygdala and hippocampus of patients with severe depression. METHODS: Quantitative MRI of the amygdala and hippocampus, as well as proton spectroscopy (MRS) of mesial temporal structures were studied in 34 drug-resistant in-patients with major depression and compared with 17 age-matched controls. Volumetric MRI data were normalized for brain size. RESULTS: The volume of the left hippocampus was significantly smaller in the patients compared with the controls. Both groups exhibited similar significant hippocampal asymmetry (left smaller than right). The patients, but not the controls, had significant asymmetry of the amygdalar volumes (right smaller than left). No differences were observed between the patients and controls in the T2 relaxation times for the hippocampus and amygdala. Mesial temporal lobe MRS revealed a significantly elevated choline/creatine ratio in the patients compared with the controls. CONCLUSIONS: This quantitative MRI study provides support for a possible association between structural and biochemical substrates and severe drug-resistant major depression. PMID- 10722183 TI - Thought disorder index of Finnish adoptees and communication deviance of their adoptive parents. AB - BACKGROUND: Diverse forms of thought disorder, as measured by the Thought Disorder Index (TDI), are found in many conditions other than schizophrenia. Certain thought disorder categories are primarily manifest during psychotic schizophrenic episodes. The present study examined whether forms of thought disorder qualify as trait indicators of vulnerability to schizophrenia in persons who are not clinically ill, and whether these features could be linked to genetic or environmental risk or to genotype-environment interactions. The Finnish Adoptive Study of Schizophrenia provided an opportunity to disentangle these issues. METHODS: Rorschach records of Finnish adoptees at genetic high risk but without schizophrenia-related clinical diagnoses (N = 56) and control adoptees at low genetic risk (N = 95) were blindly and reliably scored for the Thought Disorder Index (TDI). Communication deviance (CD), a measure of the rearing environment, was independently obtained from the adoptive parents. RESULTS: The differences in total TDI between high-risk and control adoptees were not statistically significant. However, TDI subscales for Fluid Thinking and Idiosyncratic Verbalization were more frequent in high-risk adoptees. When Rorschach CD of the adoptive rearing parents was introduced as a continuous predictor variable, the odds ratio for the Idiosyncratic Verbalization component of the TDI of the high-risk adoptees was significantly higher than for the control adoptees. CONCLUSIONS: Specific categories of subsyndromal thought disorder appear to qualify as vulnerability indicators for schizophrenia. Genetic risk and rearing-parent communication patterns significantly interact as a joint effect that differentiates adopted-away offspring of schizophrenic mothers from control adopted-away offspring. PMID- 10722184 TI - Is auditory imagery defective in patients with auditory hallucinations? AB - BACKGROUND: A variant of the 'inner speech' theory of auditory verbal hallucinations in schizophrenia suggests that there is an abnormality of the relationship between the 'inner voice' and 'inner ear', such that hallucinators are unable to distinguish inner 'imagined' speech from real external speech, and so misrecognize inner speech as alien. METHODS: Five experiments were carried out comparing 12 schizophrenic patients who were highly prone to hallucinate, with seven patients who were not, on a series of auditory imagery tasks that are differentially dependent on inner voice/inner ear partnership for successful performance: parsing meaningful letter/number strings; the verbal transformation effect; phoneme judgements; pitch judgements, and homophony and rhyme judgements. RESULTS: Contrary to our hypothesis, there was no evidence that the group with the propensity to hallucinate were impaired on tasks requiring normal inner ear/inner voice partnership. CONCLUSIONS: Together with previous work indicating no impairment of the phonological loop in patients who hallucinate, these results suggest that inner speech and auditory verbal hallucinations are not connected in a simplistic or direct way. Indeed, a reappraisal of psychological models of hallucinations in general may be warranted. PMID- 10722185 TI - Schizophrenics know more than they can tell: probabilistic classification learning in schizophrenia. AB - BACKGROUND: Previous studies have demonstrated impaired explicit and preserved implicit memory functions in schizophrenia. However, it is less clear whether schizophrenics can learn complex information (e.g. probabilistic stimulus response associations) with or without access for conscious recollection. In this study we applied a classification learning task to assess explicit and implicit processes concurrently. METHODS: Two test procedures were administered to 40 schizophrenic subjects and 20 healthy volunteers: a probabilistic classification learning (PCL) task to evaluate implicit memory functions; and a category cue recognition test to investigate the explicit memory system. The PCL task included feedback guided category learning of geometrical shapes. These shapes were called category cues, predicting class membership with certain probabilities. The gradual increase of categorization performance during the feedback learning was a potentially implicit process, whereas the subsequent recognition of category cues required explicit memory functions. RESULTS: The schizophrenic patients improved their categorization performance to a similar extent to the controls, but they failed to recognize the category cues. Memory performances were independent of the positive and negative symptoms. CONCLUSIONS: Patients with schizophrenia were able to establish representations of complex categories, but these remained unconscious. This is consistent with earlier reports, suggesting damaged explicit and spared implicit memory in schizophrenia. PMID- 10722186 TI - Perception of threat in schizophrenics with persecutory delusions: an investigation using visual scan paths. AB - BACKGROUND: Cognitive theories of persecutory delusions in schizophrenia include increased attention to threat and reduced re-appraisal of information during decision-making. METHODS: We employed visual scan path measurements, an 'on-line' marker of attention, in schizophrenic patients with persecutory delusions (N = 19), negative symptom- and medication-matched patients with non-persecutory delusions (N = 8), and normal controls (N = 18). Stimuli comprised black-and white photographs of social scenes rated as depicting either neutral, ambiguous or overtly threatening activity. Foreground areas containing salient information with regard to the overall scene were rated independently as either threatening or non-threatening in both the overtly threatening and ambiguous scenes; all foreground areas were rated as non-threatening in the neutral scene. RESULTS: For the ambiguous scene only, schizophrenics with persecutory delusions directed gaze to less threatening areas, and, for all three scenes, demonstrated reduced re appraisal of information compared with both control groups. All subjects showed similar viewing strategies for the overtly threatening and neutral scenes. CONCLUSIONS: These findings suggest abnormal information gathering and evaluation in schizophrenics, specifically related to the presence of persecutory delusions. In particular, the results point to biased processing of contextual information in an ambiguous setting in these patients, and perhaps perception of threat in inappropriate places. PMID- 10722187 TI - Attentional bias for drug cues in opiate dependence. AB - BACKGROUND: In a number of theories of compulsive drug use conditioned responses to stimuli associated with drug taking play a pivotal role. For example, according to incentive-sensitization theory (Robinson & Berridge, 1993), drug related stimuli selectively capture attention, and the neural mechanisms underlying this attentional bias play a key role in the development and maintenance of drug dependence, and in relapse. However, there has been little work that assesses attentional biases in addiction. METHODS: We used a pictorial probe detection task to investigate whether there is an attentional bias to stimuli associated with drug use in opiate dependence. Stimuli presented included pairs of drug-related and matched neutral pictures. Methadone-maintained opiate addicts (N = 16) were compared with age-matched controls (N = 16). RESULTS: A mixed design analysis of variance of response times to probes revealed a significant three-way interaction of group x drug picture location x probe location. Opiate addicts had relatively faster reaction times to probes that replaced drug pictures rather than neutral pictures, consistent with the predicted attentional bias to drug-related stimuli. CONCLUSIONS: These results support the idea that an attentional bias for drug-related stimuli occurs in opiate dependence. This is consistent with the concept of a central role for such salient stimuli in compulsive drug use. PMID- 10722188 TI - Non-linear relationship between an index of social deprivation, psychiatric admission prevalence and the incidence of psychosis. AB - BACKGROUND: Indicators of population socio-economic disadvantage expressed as weighted deprivation indices show strong relationships with mental health and underpin national funding of psychiatric services. A new index of social deprivation, the Mental Illness Needs Index, has been devised specifically to predict need for psychiatric services. Its validity has not been established outside the area in which it was developed. METHODS: We explored the relationship between the Mental Illness Needs Index and two alternative indicators of need for mental health services: the prevalence of psychiatric admission for electoral wards in Nottingham (calculated from Hospital Episode Statistics for the years 1992 and 1993) and ward-based incidence rates for psychosis (ICD-10 F1X-F33). Relationships were explored graphically using local regression models, and estimated using Generalized Linear and Additive Models, and Poisson regression. RESULTS: Social deprivation was strongly related to admission prevalence and psychosis incidence (Spearman's rho 0.63 and 0.44 respectively). Neither admission prevalence, nor the incidence of psychosis were linearly related to social deprivation. Areas with above average social deprivation had both more new cases of psychoses and a higher proportion of the population admitted than expected from a linear function. CONCLUSIONS: Application of a linear function to funding gradients may underfund high and low need areas and overfund median need areas. Improving the precision of estimates of the relationship between social deprivation and need for services is crucial to more equitable resource allocation. PMID- 10722189 TI - Homeless youth in London: II. Accommodation, employment and health outcomes at 1 year. AB - BACKGROUND: While there is considerable evidence of a high prevalence of psychiatric disorder among homeless youth, much less is known about its long-term course or the impact it may have on accommodation outcomes. METHOD: A random sample of 161 homeless people 16-21 years of age were recruited from consecutive attendees at two of London's largest facilities for homeless young people. These young people were traced and re-interviewed a year later to examine accommodation, occupation and health outcomes. RESULTS: A total of 107 (67%) people were successfully re-interviewed. Psychiatric disorder was identified in 55% at follow up. Two thirds of those with a psychiatric disorder at index interview remained symptomatic at follow-up. Persistence of psychiatric disorder was associated with adverse childhood experiences and rough sleeping. Satisfactory accommodation outcomes were achieved by 45 subjects (42%). Better accommodation outcomes were associated with three variables measured at the index assessment: ethnic minority status; educational achievement; and, the presence of accommodation plans negotiated through a resettlement agency. While psychiatric disorder at index interview was not associated with accommodation outcome, persistent substance use in the follow-up year was associated with poor accommodation outcome. Over half of the young people had been involved in petty crime and just under a third had been convicted for more serious criminal activity. Offending and antisocial behaviour in the follow-up year were related to a history of conduct disorder, persistent substance abuse and poor accommodation outcomes. CONCLUSIONS: Young homeless people are characterized by multiple social and medical needs. Successful resettlement of this population may depend upon integrated services that address problems of persisting substance use and mental illness as well as the immediate housing need. PMID- 10722190 TI - The expressed emotion of case managers of the seriously mentally ill: the influence of expressed emotion on clinical outcomes. AB - BACKGROUND: Expressed emotion (EE) measured from relatives and informal carers has been consistently demonstrated to be associated with clinical outcome in schizophrenic patients. There have also been published studies that have investigated EE in professional carers that have suggested that the quality of the relationship between staff and patient may also be associated with patient outcomes. A large controlled trial of the effectiveness of different intensities of case management provided the opportunity to assess the association between the EE of case managers, including the quality of the relationship they had with patients under their care, and later clinical outcomes. METHOD: This was a prospective naturalistic study of EE present in a case manager patient dyad and subsequent patient outcomes. EE was assessed from the Five Minute Speech Sample (FMSS) at least 3 months after the case manager became responsible for the patient's care and a range of clinical outcomes were assessed 6 to 9 months later. Assessment of clinical outcomes was made independent and blind of the EE ratings. RESULTS: High EE ratings were significantly associated with individual case managers and not to symptom or illness factors. High EE was not associated with later clinical outcome, however, the positive relationship between case manager and patient was. The absence of a positive relationship was significantly associated with poorer outcomes. CONCLUSIONS: In spite of very low face-to-face contact between case managers and patients, compared with the amount of contact patients have with their informal carers and family, aspects of staff attitudes and behaviour did influence clinical outcome. There are potential implications of these results for staff training and clinical practice. PMID- 10722191 TI - Factors associated with neuropsychological performance in HIV-seropositive subjects without AIDS. AB - BACKGROUND: Previous research has suggested that several factors may influence the presence of cognitive impairment in human immunodeficiency virus (HIV) infection. The objective of this study was to assess the impact of cognitive reserve capacity and other variables on neuropsychological performance in early HIV infection. METHODS: The neuropsychological performance of 100 HIV seropositive subjects without AIDS (71 men and 29 women) was compared with that of 63 seronegative controls (51 men and 12 women). Measures included a neuropsychological battery, a medical examination and a psychiatric assessment. Cognitive reserve scores were based on a combination of years in school, a measure of educational achievement, and an estimate of pre-morbid intelligence. RESULTS: HIV-positive subjects had longer reaction time latencies than HIV negative subjects. Those in the HIV-positive group with low cerebral reserve scores showed the poorest performance on the neuropsychological tests. The prevalence of cognitive impairment was significantly higher in the HIV-positive group (27%) than in the controls (32%). Multiple regression analysis and logistic regression analysis were used to identify factors associated with global neuropsychological performance and cognitive impairment. Older age, lower cerebral reserve scores and not being on zidovudine treatment were associated with lower global neuropsychological scores and with the presence of cognitive impairment. CONCLUSIONS: Our results suggest that although cognitive impairment is not characteristic of early HIV infection, there is a subgroup of subjects who perform more poorly than expected. A lower reserve capacity, older age and not being on zidovudine treatment are factors that lower the threshold for neuropsychological abnormalities in cases of early HIV infection. PMID- 10722192 TI - Modelling a loss event: effect of imagined bereavement on the hypothalamic pituitary-adrenal axis. AB - BACKGROUND: Loss events are the stressors most closely associated with the onset of depressive illnesses. The acute cortisol response to loss has been little studied although it could be an important mediator of the effects of environmental stress on psychological state. METHODS: The salivary cortisol response to an established negative mood induction procedure involving music and an imagined bereavement was measured in 30 healthy volunteers. RESULTS: Considerable but transient mood lowering in response to the negative mood induction was associated with a small increase in cortisol output over 30 min. CONCLUSIONS: This procedure has some potential as a tool to investigate individual differences in the neuroendocrine response to loss events, but this is limited. There remains a need for laboratory models of relevant psychosocial stressors in mood disorders research. PMID- 10722193 TI - Quality of life in first-admitted schizophrenia patients: a follow-up study. AB - BACKGROUND: While most studies of quality of life (QoL) in schizophrenia have investigated long-term patients, relatively little is known about QoL early in the illness and how it changes over time. This study was conducted to investigate objective and subjective quality of life in first-admitted schizophrenia patients as compared to patients with long-term schizophrenia, changes between first admission and 9-month follow-up and predictors of changes. METHOD: Eighty-six patients were examined after first admission and 51 were re-interviewed at follow up. Results were compared with samples of in-patients and out-patients with long term schizophrenia. QoL was assessed using a German version of the Lancashire Quality of Life Profile. RESULTS: Although some objective QoL data were more favourable in first-admitted patients, subjective QoL was lower than in each of the other two groups, even when psychopathology and age were controlled for. On a group level, patients showed a slight improvement in subjective QoL, which was not statistically significant. Individual changes over time were not predicted by initial data, but were correlated with changes in anxiety/depression. CONCLUSION: Subjective QoL appears to be lower in first-admitted schizophrenics than in groups with long-term illness and, on a group level, it changes little within 9 months. On an individual level, changes in depressive symptoms need to be considered when interpreting changes in satisfaction with life. PMID- 10722194 TI - Alteration of platelet serotonin transporter in romantic love. PMID- 10722195 TI - Facial reconstruction using 3-D computer graphics. AB - Facial reconstruction using 3-D computer graphics is being used in our institute as a routine procedure in forensic cases as well as for skulls of historical and archaeological interest. Skull and facial data from living subjects is acquired using an optical laser scanning system. For the production of the reconstructed image, we employ facial reconstruction software which is constructed using the TCL/Tk scripting language, the latter making use of the C3D system. The computer image may then be exported to enable the production of a solid model, employing, for example, stereolithography. The image can also be modified within an identikit system which allows the addition of facial features as appropriate. PMID- 10722196 TI - Fatal suffocation by rubber balloons in children: mechanism and prevention. AB - This paper presents five autopsy cases of children who died as a result of an accidental choking on latex (rubber) balloons. All of the investigated cases suffocated by an intact (non-broken), pear-shaped and medium-sized balloon of less than 9 cm in length and 4 cm in width. The mouthpieces of all the involved balloons were upward and above the vocal cords, while the nosepieces were downward in the larynx and the trachea. The application of continuous suction forces to uninflated or partially air-filled balloon is considered the most crucial factor in causing asphyxiation by rubber balloon. An uninflated balloon is usually placed outside the mouth cavity where the child sucks the balloon into the mouth, either during a repeated trial of inverting the balloon inside-out or during the application of a direct suction to their bulb. A partially air-filled balloon is usually placed inside the mouth cavity then it is pushed further inside by movements like sucking or chewing. Other important factors are a sudden slippage of the mouthpiece from the fingers or teeth of the child, as well as the panic of the suffocation. The process of suffocation is also facilitated by the counterforce, which results from a sudden pressure of the mouth cavity over the bulb of the balloon, and the consistency of the balloons as pliable objects. Once the balloon enters the airway, it is less likely to be expelled. Supervision of children by adults and proper education programs are needed to decrease this problem. PMID- 10722197 TI - The forensic analysis of soil organic by FTIR. AB - In order to elucidate the discriminating power of various soil analytical techniques, over 100 soils samples were analyzed using conventional analysis (i.e., color, percent organic and density gradient) and a novel FTIR technique. The FTIR technique involves collecting a spectrum of a soil sample that has been oxidatively pyrolysed, and therefore all organic have been degraded. This spectrum is subtracted from the spectrum of the same sample that contained the organic prior to pyrolysis. This resultant IR spectrum represents the organic portion of the sample. The use of organic components increases the discrimination in soils that are otherwise similar. We have illustrated the usefulness of this technique by selecting four soil samples, which have identical Munsel color values but can be discriminated by subtractive ETIR. We propose that the ETIR spectra of the organic portion of soil serves a useful purpose in forensic investigations. PMID- 10722198 TI - Swiss Caucasian population data for the STR loci D2S1338 and D19S433 using the AmpFISTR SGM plus PCR amplification kit. AB - Allele and genotype frequencies for the ten STR loci D3S1358, VWA, D16S539, D2S1338, D8S1179, D21S11, D18S51, D19S433, TH01, FGA were determined in a Swiss Caucasian population sample (n=206) using the AmpFISTR SGM Plus Amplification kit. Electrophoresis was carried out on an ABI PRISM CE 310 Genetic Analyzer instrument. Previously, allele frequencies were published for the 13 STR loci D3S1358, VWA, FGA, D8S1179, D21S11, D18S51, D5S818, D13S317, D7S820, THO1, TPOX, CSF1PO and D16S539 for the same samples (n=206) amplified with the AmpFISTR Profiler Plus and Cofiler PCR Amplification kits. Since the results for the eight loci D3S1358, VWA, FGA, D8S1179, D21S11, D18S51, THO1, D16S539 shared between the AmpFISTR SGM Plus, Profiler Plus and Cofiler PCR Amplification kits already are published, only the allele frequencies for the two STR loci D2S1338 and D19S433 are reported in this paper. The two loci meet Hardy-Weinberg expectations. In addition, there is little evidence for association of alleles among the 15 loci (amplified with the Profiler, Cofiler, and SGM Plus amplification kits). The allelic frequency data can be used in forensic analyses to estimate the frequency of a multiple STR locus DNA profile in the Swiss population. PMID- 10722199 TI - DNA evidence, probabilistic evaluation and collaborative tests. AB - Forensic scientists working in 12 state or private laboratories participated in collaborative tests to improve the reliability of the presentation of DNA data at trial. These tests were motivated in response to the growing criticism of the power of DNA evidence. The experts' conclusions in the tests are presented and discussed in the context of the Bayesian approach to interpretation. The use of a Bayesian approach and subjective probabilities in trace evaluation permits, in an easy and intuitive manner, the integration into the decision procedure of any revision of the measure of uncertainty in the light of new information. Such an integration is especially useful with forensic evidence. Furthermore, we believe that this probabilistic model is a useful tool (a) to assist scientists in the assessment of the value of scientific evidence, (b) to help jurists in the interpretation of judicial facts and (c) to clarify the respective roles of scientists and of members of the court. Respondents to the survey were reluctant to apply this methodology in the assessment of DNA evidence. PMID- 10722200 TI - Distribution of the HLA-DQA1 and polymarker alleles in the Basque population of Spain. AB - HLA-DQA1 and polymarker (LDLR, GYPA, HBGG, D7S8, and GC) genotypic and allelic frequencies are determined for a population sample of 102 unrelated Basque individuals using PCR-based methodology. All six loci met Hardy-Weinberg expectations in at least two of the three analyses performed (HLA-DQA1 failed to meet Hardy-Weinberg requirements in the heterozygote deficiency test). Three linkage analysis programs (GDA, GENEPOP and LINKDOS) detected possible linkage disequilibrium between LDLR and HBGG and results from one (GDA) indicated a possible non-random association between HBGG and HLA-DQA1 as well. Allelic data for the six loci are compared to that previously established for other populations (18 for polymarker alone, 16 for polymarker plus HLA-DQA1) to determine homogeneity between the Basque sample and these groups. According to the results of G-tests based on these loci, the Tadjik, a nomadic Caucasian group from western Asia, and the Basque residents are the only sample populations surveyed that are homogenous with the Basque sample. Phylogenetic analysis places the Basque sample correctly within the Caucasian cluster. PMID- 10722201 TI - In vivo and in vitro immunomodulatory activities of Trichilia glabra aqueous leaf extracts. AB - The effects of Trichilia glabra (Meliaceae) aqueous leaf extract on mouse lymphocytes were studied. The in vitro proliferation of T and B lymphocytes was completely impaired. Besides, the extract significantly diminished both antibody and delayed hypersensitivity responses in treated mice. These results suggest that the extract exerts a marked immunomodulatory effect on the murine immune system. PMID- 10722202 TI - Hypoglycemic activity of root water decoction, sesquiterpenoids, and one polysaccharide fraction from Psacalium decompositum in mice. AB - The hypoglycemic activity of Psacalium decompositum (Asteraceae) was investigated in fasting healthy mice and alloxan-diabetic mice. The freeze-dried water decoction significantly reduced the blood glucose in normal mice (from 50.9 +/- 4.7 to 32.5 +/- 3.1 mg/dl) and in mild diabetic mice (from 208.5 +/- 13.0 to 52.3 +/- 7.0 mg/dl), 240 min after intraperitoneal administration (P < 0.005). This preparation also diminished fasting glycemia in severe diabetic mice, but the effects were minor (from 394.4 +/- 9.4 to 289.3 +/- 39.5 mg/dl). The main sesquiterpenoid constituents from P. decompositum roots, cacalol, cacalone and maturin, as well as the transformation product cacalol acetate, did not show a hypoglycemic effect on healthy mice. Nevertheless, two polysaccharide fractions (F1 and F3) obtained from the freeze-dried water extract significantly reduced the fasting glycemia in healthy mice. The best results were obtained with the F1 fraction. PMID- 10722203 TI - Inhibitory effect of Carum copticum on histamine (H1) receptors of isolated guinea-pig tracheal chains. AB - In a previous study, the relaxant and anti-cholinergic (functional antagonism) effects of Carum copticum were demonstrated on guinea-pig tracheal chains. To elucidate the other mechanisms responsible for this relaxant effect, the inhibitory effect of this plant on histamine H1 receptors was examined in this study. The anti-histaminic effects of extracts, essential oil, 5 nM chlorpheniramine, and saline were tested by performing the cumulative log concentration-response curves of histamine induced contraction of isolated guinea pig tracheal chains incubated with three different conditions: 1.4 microM indomethacin (group 1, n = 9); indomethacin, 1 microM propranolol, and 10 nM atropine (group 2, n = 8); and indomethacin and propranolol (group 3, n = 7). The results showed clear rightward shifts in histamine response curves obtained in the presence of extracts, essential oil, and chlorpheniramine in all three sets of experiments compared with the curves obtained in the presence of saline. The effective concentrations of histamine causing 50% of maximum response (EC50) obtained in the presence of extracts, essential oil, and chlorpheniramine in all three sets of experiments were significantly higher than those of saline (P < 0.05- < 0.001) except EC50 of macerated extract in group 1. However, maximum response to histamine obtained in the presence of extracts and essential oil were lower (P = 0.005- < 0.001) except maximum response of essential oil in group 2. In addition, the maximum responses obtained in the presence of extracts in group 2 experiments compared to the other two sets of experiments were improved. Comparison of the slope of histamine-response curves showed parallel shifts of the curves obtained in the presence of all extracts and essential oil in group 2 experiments. The results of this study indicated a competitive antagonism effect of C. copticum at histamine H1 receptors. A beta-adrenergic stimulatory effect of essential oil and an anti-cholinergic property of the plant were also suggested. PMID- 10722204 TI - Gastroprotective effect of essential oil from Croton cajucara Benth. (Euphorbiaceae). AB - The gastroprotective activity of the essential oil from the bark of Croton cajucara Benth (Euphorbiaceae) was assessed in three different models of experimentally induced gastric ulcer in mice. At oral dose of 100 mg/kg the essential oil reduced gastric lesions induced by hypothermic restraint stress and HCl/ethanol significantly. In the HCl/ethanol model a dose-dependent gastroprotective effect was found. Moreover, significant changes in gastric parameters such as pH, secretion rate and total gastric acid were found after intraduodenal administration of essential oil under ligated pylorus (Shay) conditions. The acute toxicity of essential oil was assessed in mice. The LD50 values were 9.3 and 680 mg/kg for oral and intraperitoneal administrations, respectively. The cytotoxicity of essential oil was studied also. A dose dependent cell viability inhibition was found in V79 fibroblast cell cultures with an IC50 of 22.9 microg/ml. Our results support the pharmacological study of this essential oil. PMID- 10722205 TI - Inhibiting and disaggregating effect of gel-filtered Galega officinalis L. herbal extract on platelet aggregation. AB - The in vitro inhibiting and disaggregating effect on platelet aggregation of a gel-fractionated herbal extract from Galega officinalis L. is examined. The obtained Sephadex G-25 filtered fraction was 35-36 times more active than the crude extract. The threshold concentration at which this fraction inhibits platelet aggregation (5-10% inhibition) by 50 microM adenosine 5'-diphosphate (ADP) is 4.5-5 microg per 1 ml platelet-rich plasma (PRP). At a concentration of 35 microg/ml PRP the fraction inhibits 50% of aggregation by ADP and at a concentration of 125 microg/ml PRP fully inhibits the aggregation of PRP by ADP. At a concentration of 40 microg/ml PRP the fraction inhibits initiation of platelet aggregation by 0.18 mg/ml collagen and at 50 microg/ml PRP inhibits the initiation of aggregation by 0.7 units/ml thrombin. The G-25 filtered fraction shows a strong disaggregating effect on aggregated PRP. At a concentration of 65 75 microg/ml PRP, the fraction is able to disaggregate the 50-53% of aggregated platelet-rich plasma by 50 microM ADP, and 25% of aggregated PRP by 0.18 mg/ml collagen. PMID- 10722206 TI - Zulu medicinal plants with antibacterial activity. AB - Aqueous, methanolic and ethyl acetate extracts of 14 plants used in traditional Zulu medicine for treatment of ailments of an infectious nature were screened for antibacterial activity. Most of the activity detected was against gram-positive bacteria. Tuber bark extracts of Dioscorea sylvatica had activity against gram negative Escherichia coli and extracts of Dioscorea dregeana, Cheilanthes viridis and Vernonia colorata were active against Pseudomonas aeruginosa. The highest antibacterial activity was found in extracts of C. viridis, D. dregeana, D. silvatica, Melianthus comosus and V. colorata. In general, methanolic extracts exhibited higher activity than aqueous and ethyl acetate extracts. PMID- 10722207 TI - Effects of Opuntia megacantha on blood glucose and kidney function in streptozotocin diabetic rats. AB - The purpose of the study was to investigate the effects of Opuntia megacantha leaf extracts on blood glucose concentrations and kidney function in normal and streptozotocin (STZ)-diabetic rats. STZ-diabetic and non-diabetic rats were orally administered extracts of O. megacantha leaves (20 mg/100 g body weight) daily for 5 weeks and respective control rats were administered normal saline (0.1 ml/100 mg body weight). Urine volume, urinary outputs of Na+, K+ and creatinine were monitored daily over the 5-week period. Plasma concentrations of Na+, K+, urea and creatinine and the glomerular filtration rate (GFR) as assessed by creatinine clearance were determined after 5 weeks. Plasma glucose concentrations in STZ-diabetic and non-diabetic rats were reduced by the administration of leaf extracts of O. megacantha. However, leaf extracts increased urinary Na+ output in STZ-diabetic and non-diabetic rats, concomitantly with a reduction in plasma concentration of the ion. O. megacantha leaf extracts significantly increased plasma creatinine and urea concentrations in non-diabetic and STZ-diabetic rats. Administration of the leaf extract was also associated with an increased GFR in STZ-diabetic rats (from 1.8 +/- 0.3 ml/min to 2.8 +/- 0.3 ml/min, n = 8) although the rate was unaltered in non-diabetic rats. The results suggest that leaf extracts of O. megacantha not only reduce blood glucose levels, but may be toxic to the kidney as shown by the elevation in plasma urea and creatinine concentrations and the reduction of plasma Na+ concentration. PMID- 10722208 TI - In vitro snake venom detoxifying action of the leaf extract of Guiera senegalensis. AB - The extract of the leaves of Guiera senegalensis was found to detoxify (in vitro) venom from two common northern Nigerian snake species, Echis carinatus and Naja nigricollis, in separate experiments. There was a remarkable reduction in the mortality of albino mice after intra-peritoneal (i.p.) administration of reconstituted venom incubated with the extract, when compared to those challenged with the venom only. The survival of the animals exposed to the venom incubated with the different concentrations of the extract was used as the in vitro detoxification parameter. PMID- 10722209 TI - In vitro antioxidant activity of Anthriscus cerefolium L. (Hoffm.) extracts. AB - Standardised aqueous extracts of chervil (Anthriscus cerefolium L. Hoffm.) (Apiacae) were investigated for antioxidant effect. Numerous in vitro test methods were used to determine whether the extracts, from different vegetative parts (root, herb) had H-donor, metal binding, reductive, free radical scavenging and membrane protective activity. Apiin was used as a reference material. The herb extract showed better activity in all experiments than the root extract. The present results underline that the wateric chervil extracts have antioxidant and anti-lipoperoxidant activity. PMID- 10722210 TI - The mating consequences of sexual segregation within inflorescences of flowering plants. AB - Many co-sexual plants segregate female and male function among flowers on an inflorescence through dichogamy or the production of unisexual flowers. Sexual segregation may reduce self-pollination among flowers within inflorescences (geitonogamy), thereby increasing the pollen available for export to other plants. To assess these complementary roles we manipulated the simultaneously hermaphroditic (adichogamous) flowers of Eichhornia paniculata to produce ten flowered inflorescences with either female above male flowers (female/male inflorescences) or male/female inflorescences, which competed for mating opportunities with five-flowered adichogamous inflorescences. Because of the upward movement of bumble-bees, selfing increased upward in adichogamous inflorescences (overall female selfing rate s+/-s.e.=0.320+/-0.026). Female flowers of male/female inflorescences selfed less than flowers in corresponding positions in adichogamous inflorescences so s fell to 0.135+/-0.027. In contrast, all-female flowers of female/male inflorescences selfed similarly to upper flowers on adichogamous inflorescences, elevating s (0.437+/-0.043). During 1997, male/female inflorescences sired more outcrossed seeds than female/male or adichogamous inflorescences, whereas during 1994 flowers on male/female inflorescences received fewer visits than those of adichogamous inflorescences, reducing their outcross siring success. Hence, sexual segregation limits geitonogamy and enhances outcross siring success when it does not affect pollinator behaviour, illustrating the importance of both female and male function in inflorescence design. PMID- 10722211 TI - Song as an indicator of male parental effort in the sedge warbler. AB - Repertoire size has been found to be a sexually selected trait in a number of bird species, although the advantages of mating with a male who possesses a complex song remain unclear. We studied the potential role of song as an indicator of male parental effort in the sedge warbler Acrocephalus schoenobaenus. The male provisioning rate was used as a measure of male parental effort and was found to increase with nestling age and brood size. When controlling for chick age, brood size and other variables, we found a highly significant positive correlation between a measure of song complexity (repertoire size) and male parental effort. Both male parental effort and repertoire size were found to be positively correlated with chick weight when controlling for chick age. We found no correlation between a measure of song output (amount of song flighting) or territory size and parental effort. Repertoire size is known to be the most important cue in female choice amongst sedge warblers and we discuss the possible reasons for this. We suggest that, in choosing a male with a large repertoire, a female obtains not only indirect benefits but also direct benefits in the form of increased parental effort. PMID- 10722212 TI - The inheritance of female preference functions in a mate recognition system. AB - Mate recognition systems (MRSs) play a major role in sexual selection and speciation, yet few studies have analysed both male and female components in detail. Here, female preference functions have been characterized for the tettigoniid bushcricket Ephippiger ephippiger, and the inheritance of male song and female preference functions followed in crosses between subspecies. Songs are disproportionately determined by sex-linked genes. However, there is no evidence for a role of maternally derived sex-linked genes in female preference or of maternal effects. At the genetic level, there is a mismatch between peak preferences and male song, consistent with an evolutionary history of persistent directional preferences. Such a pattern of inheritance could contribute to the process of speciation via the evolution of new MRSs. PMID- 10722213 TI - Risk-taking restraints in a bird with reduced egg-hatching success. AB - Risk taking, as is any other phenotypic and/or behavioural trait, is determined by proximate constraints related to time or resource availability and by evolutionary adaptive restraints related to the differences in the costs of risk taking and its benefits in terms of fitness. Because risk taking is influenced by many confounding variables related to experimental design, environment, parents and offspring, few field studies have been reported which unambiguously separate the effects of restraints from those of constraints. We compared parental risk taking in blue tits (Parus caeruleus) during brood defence towards a nest predator in broods with experimentally reduced and natural egg-hatching success leaving the original number of eggs in the nest. The experimentally reduced broods had more time or resources available and lower risk-taking benefits compared to the control broods. 'Constraint' would predict more risk taking in broods having experimentally reduced egg-hatching success, whereas 'restraint' would predict the opposite effect with more risk taking in broods with natural egg-hatching success. We report, to our knowledge, the first field study experimentally demonstrating a brood defence restraint in response to reduced egg hatching success. This demonstration was only possible after controlling for more than 20 potential confounding variables showing once more how complicated it is to separate proximate from evolutionary levels of analyses in natural populations. PMID- 10722214 TI - Sensory exploitation as an evolutionary origin to nuptial food gifts in insects. AB - Nuptial food gifts given by males to females at mating are widespread in insects, but their evolutionary origin remains obscure. Such gifts may arise as a form of sensory trap that exploits the normal gustatory responses of females, favouring the selective retention of sperm of gift-giving males. I tested this hypothesis by offering foreign food gifts, synthesized by males of one cricket species, to females of three non-gift-giving species. Females provisioned with novel food gifts were 'fooled' into accepting more sperm than they otherwise would in the absence of a gift. These results support the hypothesis that nuptial food gifts and post-copulatory female mating preferences coevolve through a unique form of sensory exploitation. PMID- 10722215 TI - The allometry of patch selection in ruminants. AB - An axiomatic feature of food consumption by animals is that intake rate and prey abundance are positively related. While this has been demonstrated rigorously for large herbivores, it is apparent from patch selection trials that grazers paradoxically tend to prefer short, sparse swards to tall, dense swards. Indeed, migratory herbivores often shift from areas of high to low sward biomass during the growing season. As nutritional quality is an inverse function of grass abundance, herbivores appear to sacrifice short-term intake for nutritional gains obtainable by eating sparse forage of higher quality. Explicit models of this trade-off suggest that individual ruminants maximize daily rates of energy gain by choosing immature swards of intermediate biomass. As body mass is related positively to both ruminant cropping rates and digestibility, there should be an allometric link between grass abundance and energy maximization, providing a tool for predicting patterns of herbivore habitat selection. We used previously published studies to develop a synthetic model of trade-offs between forage abundance and quality predicting that optimal sward biomass should scale allometrically with body size. The model predicts size-related variation in habitat selection observed in a guild of grazing ungulates in the Serengeti ecosystem. PMID- 10722216 TI - Experimental tests of predation and food hypotheses for population cycles of voles. AB - Pronounced population cycles are characteristic of many herbivorous small mammals in northern latitudes. Although delayed density-dependent effects of predation and food shortage are often proposed as factors driving population cycles, firm evidence for causality is rare because sufficiently replicated, large-scale field experiments are lacking. We conducted two experiments on Microtus voles in four large predator-proof enclosures and four unfenced control areas in western Finland. Predator exclusion induced rapid population growth and increased the peak abundance of voles over 20-fold until the enclosed populations crashed during the second winter due to food shortage. Thereafter, voles introduced to enclosures which had suffered heavy grazing increased to higher densities than voles in previously ungrazed control areas which were exposed to predators. We concluded that predation inhibits an increase in vole populations until predation pressure declines, thus maintaining the low phase of the cycle, but also that population cycles in voles are not primarily driven by plant-herbivore interactions. PMID- 10722217 TI - A new demographic function maximized by life-history evolution. AB - A goal of life-history theory has been to understand what combination of demographic traits is maximized by natural selection. In practice, researchers usually choose either density-independent population growth rate, lambda, or lifetime reproductive success, R0 (expected number of offspring produced in a lifetime). Others have shown that the maxima of density-independent lambda and R0 are evolutionarily stable strategies under specific density-dependent conditions: population regulation by equal density dependence among all age classes for lambda and by density dependence on a single age class for R0. Here I extend these connections between density-independent optimization models and density dependent invasion function models in two ways. First, I derive a new demographic function for which a maximum corresponds to attainability of the equilibrium strategy or stability of the mean rather than stability of the variance of the strategy distribution. Second, I show explicitly a continuous range of cases with maxima between those for the lambda and R0. Graphical and biological interpretations are given for an example model. Finally, exceptions to a putative life-history generality (from lambda and R0 models), that high early-life mortality selects for high iteroparity, are shown. PMID- 10722218 TI - A phylogenetic approach to community assembly from a local species pool. AB - Ecological theory provides two contrasting predictions about the characteristics of the species combining to form communities. Classical competition theory states that they will be less similar than expected by chance, whilst the environmental structuring hypothesis states that they will be more similar. We investigated these predictions by applying phylogenetic methods of analysis (PICs) to a grassland community, examining species on the basis of their traits. At the scale of investigation most useful in making predictions about the presence and abundance of species (the community level, the species forming the community were more similar than would be expected by chance. The use of PICs resulted in a more sensitive test than if phylogeny had been ignored, allowing the detection of effects that would otherwise have been overlooked or underestimated. Selected traits from the PICs analysis were used to develop a predictive model of community membership using discriminant analysis. This correctly identified species in the pool which were present in the community but failed to predict absences accurately, implying that dispersal limitation may operate in the community. PMID- 10722219 TI - Extreme environmental change and evolution: stress-induced morphological variation is strongly concordant with patterns of evolutionary divergence in shrew mandibles. AB - Morphological structures often consist of simpler traits which can be viewed as either integrated (e.g. correlated due to functional interdependency) or non integrated (e.g. functionally independent) traits. The combination of a long-term stabilizing selection on the entire structure with a short-term directional selection on an adaptively important subset of traits should result in long historical persistence of integrated functional complexes, with environmentally induced variation and macroevolutionary change confined mostly to non-integrated traits. We experimentally subjected populations of three closely related species of Sorex shrews to environmental stress. As predicted, we found that most of the variation in shrew mandibular shape was localized between rather than within the functional complexes; the patterns of integration did not change between the species. The stress-induced variation was confined to nonintegrated traits and was highly concordant with the patterns of evolutionary change--species differed in the same set of non-integrated traits which were most sensitive to stress within each species. We suggest that low environmental and genetic canalization of non-integrated traits may have caused these traits to be most sensitive not only to the environmental but also to genetic perturbations associated with stress. The congruence of stress-induced and between-species patterns of variation in non-integrated traits suggests that stress-induced variation in these traits may play an important role in species divergence. PMID- 10722220 TI - Environmental heterogeneity and balancing selection in the acorn barnacle Semibalanus balanoides. AB - The northern acorn barnacle Semibalans banlanoides occupies several intertidal microhabitats which vary greatly in their degree of physical stress. This environmental heterogeneity creates distinct selection regimes which can maintain genetic variation in natural populations. Despite considerable attention placed on the link between spatial variation in fitness and balancing selection at specific loci, experimental manipulations and fitness estimates for molecular polymorphisms have rarely been conducted in the wild. The aim of this transplant experiment was to manipulate the level of physical stress experienced by a cohort of barnacles in the field and then investigate the spatial variation in fitness for genotypes at three loci: two candidate allozymes and the mitochondrial DNA control region. The viability of mannose-6-phosphate isomerase (Mpi) genotypes was dependent on the level of physical stress experienced in the various treatments; alternative homozygotes were favoured in alternative high stress-low stress environments. In contrast, the fitness of genotypes at other loci was equivalent among treatments and unaffected by the manipulation. Evaluated in the light of balancing selection models, these data indicate that the presence of multiple environmental niches is sufficient to promote a stable Mpi polymorphism in barnacle populations and that allelic variation at this locus reflects the process of adaptation to the heterogeneous intertidal landscape. PMID- 10722221 TI - Evolutionary trade-offs at two time-scales: competition versus persistence. AB - The evolution of many natural systems is complicated due to dynamics at a mixture of time-scales. This is especially true when there is a trade-off between large reproductive rates and long-term persistence; such behaviour is frequently observed in disease models. In this paper, a simple partial differential equation model is formulated which describes the evolutionary dynamics of two disease strains in a metapopulation: one strain is a better short-term competitor; the other has greater persistence. By considering the behaviour of means and higher order moments, analytical expressions for the evolutionary behaviour are produced in the case when the two strains are phenotypically close. PMID- 10722223 TI - Human-like, population-level specialization in the manufacture of pandanus tools by New Caledonian crows Corvus moneduloides. AB - The main way of gaining insight into the behaviour and neurological faculties of our early ancestors is to study artefactual evidence for the making and use of tools, but this places severe constraints on what knowledge can be obtained. New Caledonian crows, however, offer a potential analogous model system for learning about these difficult-to-establish aspects of prehistoric humans. I found new evidence of human-like specialization in crows' manufacture of hook tools from pandanus leaves: functional lateralization or 'handedness' and the shaping of these tools to a rule system. These population-level features are unprecedented in the tool behaviour of free-living non-humans and provide the first demonstration that a population bias for handedness in tool-making and the shaping of tools to rule systems are not concomitant with symbolic thought and language. It is unknown how crows obtain their tool behaviour. Nevertheless, at the least they can be studied in order to learn about the neuropsychology associated with early specialized and/or advanced population features in tool making such as hook use, handedness and the shaping of tools to rule systems. PMID- 10722222 TI - Variance of molecular datings, evolution of rodents and the phylogenetic affinities between Ctenodactylidae and Hystricognathi. AB - The von Willebrand factor (vWF) gene has been used to understand the origin and timing of Rodentia evolution in the context of placental phylogeny vWF exon 28 sequences of 15 rodent families and eight non-rodent eutherian clades are analysed with two different molecular dating methods (uniform clock on a linearized tree; quartet dating). Three main conclusions are drawn from the study of this nuclear exon. First, Ctenodactylidae (gundis) and Hystricognathi (e.g. porcupines, guinea-pigs, chinchillas) robustly cluster together in a newly recognized clade, named 'Ctenohystrica'. The Sciurognathi monophyly is subsequently rejected. Pedetidae (springhares) is an independent and early diverging rodent lineage, suggesting a convergent evolution of the multiserial enamel of rodent incisors. Second, molecular date estimates are here more influenced by accuracy and choice of the palaeontological temporal references used to calibrate the molecular clock than by either characters analysed (nucleotides versus amino acids) or species sampling. The caviomorph radiation at 31 million years (Myr) and the pig porpoise split at 63 Myr appear to be reciprocally compatible dates. Third, during the radiation of Rodentia, at least three lineages (Gliridae, Sciuroidea and Ctenohystrica) emerged close to the Cretaceous-Tertiary boundary, and their common ancestor separated from other placental orders in the Late Cretaceous. PMID- 10722224 TI - Myosin monomer density and exchange in synthetic thick filaments investigated using fluorescence microscopy with single molecule sensitivity. AB - The number of myosin molecules in synthetic thick filaments, prepared by dialysis at 0.12 M NaCl and pH 7.0, was estimated to be between 400 and 800 molecules per micrometre under conditions appropriate for in vitro motility assays. This estimate was based on a number count of Cy3-labelled myosin molecules incorporated into filaments at a nominal ratio of 1:1000. At this dilution, single fluorescent spots were resolved corresponding to individual labelled myosins. The spots usually bleached with a one- or two-step profile but, in around 30% of the cases, fluctuations were observed indicating that additional photophysical or photochemical events had occurred. Myosin molecules were shown to exchange between filaments in suspension on a time-scale of several hours at 4 degrees C, but the reaction was only 75% complete after 48 h, suggesting a non exchangeable core. However, myosin exchange does not appear to be the predominant source of the fluctuations in fluorescence intensity in the single molecule assays. PMID- 10722226 TI - Molecular aspects in the pathogenesis of human systemic lupus erythematosus. AB - Systemic lupus erythematosus is a common and often devastating systemic autoimmune disease of unknown etiology. In this communication we review the latest developments of the molecular pathogenesis of human lupus. Novel genetic studies of multiplex lupus families have revealed potential disease-associated genome intervals, put special emphasis on genetic loci mapping in the long arm of chromosome 1 and have underscored the complexity of the underlying genetic background. New data have emerged on the role of estrogens in the function of lymphocytes and a number of studies have recently emphasized the relative Th-1/Th 2 cytokine imbalance in favor of a Th-2 type cytokine immune response. Finally, novel experiments have revealed an abnormal antigen receptor-mediated signaling process in lupus T and B cells, which may influence the aberrant expression and function of costimulatory molecules as well as of other aspects of immune cell function. It is important to decipher the underlying molecular mechanisms that govern the expression of human lupus, because we may design novel, rational approaches in the treatment of a human lupus, a disease that has high morbidity and mortality. PMID- 10722225 TI - The immunopathology of primary biliary cirrhosis: thoughts for the millennium. AB - Primary biliary cirrhosis is an organ specific autoimmune disease that produces progressive cholestatic liver failure. It is predominantly a disease of women characterized by chronic progressive destruction of small intrahepatic bile ducts with portal inflammation and ultimately fibrosis. The serologic hallmark of primary biliary cirrhosis (PBC) is the presence of antibodies to mitochondria. The mechanisms by which and if which such antibodies produce liver tissue injury is unknown. However, the presence of these antibodies have allowed detailed immunological definition of the antigenic epitopes, the nature of reacting autoantibodies and the characterization of T cell responses. Several mechanisms may now be proposed regarding the immune mediated bile duct damage in PBC, including the possible role of T cell-mediated cytotoxicity and intracellular interaction between the IgA class of antimitochondrial antibodies (AMA) and mitochondrial autoantigens. The advent of molecular biology, the ability to clone and define epitopes, and the use of in situ nucleic acid hybridization, have all led to advances in understanding the natural history of immunopathology in PBC. There are major questions which remain unanswered, including, of course, etiology, but also including the questions of why there is female predominance, the absence of PBC in children, the relative ineffectiveness of immunosuppressive drugs, and the specific role of mitochondrial antigens. In this review, we focus on these issues and particularly on the immunobiology of patients with this disease. PMID- 10722227 TI - Preventing staphylococcal disease by disarming the immune responses to infection. AB - Use of experimental models of staphylococcal infections clarified several bacterial virulence factors as well as many hematopoetic cell types and their products that are involved in the pathogenesis of infection. For many decades it has been believed that antibody mediated response to staphylococci and their products was the major, if not the only one, hallmark of immune reactivity during infection. Recent studies have documented that T cell mediated responses to superantigens produced by staphylococci are not only prominent but also decisive with respect to sequels. Also the nonantigen specific immune responsiveness to staphylococcal infection is reviewed including roles of neutrophils, complement system and nitric oxide. The knowledge gained regarding staphylococcal virulence factors and the host immune responses has prompted researchers to develop new strategies how to interact in vivo witl the infectious process. Some of these approaches are commented in this review regarding e.g. vaccination procedures in order to prevent severe infections as well as therapeutic procedures to minimize organ damage during an ongoing infectious process. PMID- 10722229 TI - Effective phage therapy is associated with normalization of cytokine production by blood cell cultures. AB - The aim of this study was to investigate the effect of phagotherapy on tumor necrosis factor alpha (TNF-alpha) and interleukin 6 (IL-6) serum levels and the ability of blood cells to produce these cytokines in culture. Fifty one patients with long-term, suppurative infections of various tissues and organs were enrolled. The ability of cells to secrete cytokines was tested using whole blood cell cultures, unstimulated or stimulated with lipopolysaccharide (LPS) from E. coli. In addition, cytokine serum levels were determined. Measurement of cytokine activity was performed using bioassays. We showed that TNF-alpha, but not IL-6 serum levels, were regulated upon division of patients into categories exhibiting initial: low, moderate and high cytokine levels. The low spontaneous production of IL-6 by blood cell cultures was elevated significantly on day 21 of phage therapy, whereas high release of this cytokine was inhibited. No such correlation was observed with LPS-induced IL-6 production in cell cultures when cells from low-, moderately- or highly-reactive patients were studied. Phage therapy modified TNF release according to the initial ability to produce that cytokine: it reduced TNF production in high responders and increased it in low responders. Patients infected only with Gram-positive bacteria demonstrated analogous changes in the spontaneous and LPS-induced TNF-alpha production as in the whole studied group. A similar kind of regulation was observed in TNF-alpha and LPS-induced production, i.e. low production was significantly elevated, high strongly inhibited, and moderate only slightly affected. In summary, we demonstrated for the first time that effective phage therapy can normalize TNF-alpha serum levels and the production of TNF-alpha and IL-6 by blood cell cultures. PMID- 10722228 TI - Crosstalk between G protein-coupled receptors and tyrosine kinase signaling: Src take centre stage. AB - Although the role of G protein-coupled receptors in the regulation of metabolic, secretory and contractile responses is well established, they have only recently been recognized as important mediators of cellular growth and differentiation. G protein-coupled signaling pathways had been previously thought to be totally independent of the tyrosine kinase receptor pathway. It was previously believed that molecular switches responsible for growth factor tyrosine kinase receptor signaling and G protein-coupled signaling were divided into a distinct sets of protein families. Recent evidence has demonstrated, however, that G protein coupled receptors can crosstalk to tyrosine kinase signaling. In the past few years several groups have found that G protein-coupled receptors utilize non receptor tyrosine kinases, mostly that of Src family, and some adapter proteins, to regulate tyrosine kinase cascades in cells. PMID- 10722230 TI - Abnormal distribution of gammadelta T lymphocytes in Graves' disease and insulin dependent diabetes type 1. AB - There is an increasing evidence that CD3+ cells, bearing gammadelta T cell receptors representing a minor subpopulation of T cells in the peripheral blood of humans are involved in the development of autoimmunity. The aim of the present study was determination of the gammadelta T cell subpopulation levels in the peripheral blood of subjects with Graves' disease and newly diagnosed type 1 diabetes in comparison to age-matched healthy controls. The percentages of CD3+, CD8+, gammadelta TCR+CD8+, gammadelta TCR+CD8- lymphocyte subsets were measured by flow cytometry. In the peripheral blood of newly diagnosed Graves' disease patients we showed a significant decrease of gammadelta TCR+ cells and gammadelta TCR+CD8- subset content in comparison to the percentages observed in subjects after methimazole treatment and in healthy controls. We also found a significant increase of gammadelta TCR+CD8+ cells in the peripheral blood of subjects with insulin-dependent diabetes, treated with insulin for 3-6 months. The present findings confirm our previous hypothesis that gammadelta TCR+CD8+ lymphocyte subset could play a role in the pathogenesis of diabetes type 1, probably as regulatory T cells and could be induced by delivery of exogenous insulin. Our results suggest that gammadelta T cells (gammadelta TCR+CD8- subset) could also play an important role in the development of Graves' disease and that their levels are modulated by thyreostatic treatment. PMID- 10722231 TI - Changes in glucocorticoid-induced apoptosis and in expression of Bcl-2 protein during long-term culture of thymic lymphoma. AB - Lymphomagenesis is a multistep process progressively freeing transformed thymocytes from external regulatory signals, i.e. thymic developmental program controlling growth, differentiation or apoptosis. Here we report that cells of thymic lymphoma overexpressing Ras/Raf proteins, initially resistant to TCR dependent apoptosis but sensitive to dexamethasone- and etoposide-induced cell death, became insensitive to dexamethasone after long-time culture. That transition correlated with a strong increase in the expression of the anti apoptotic Bcl-2 protein. Interestingly, lymphoma cells were still sensitive to p53-mediated apoptosis induced by etoposide. It suggests that the anti-apoptotic activity of Bcl-2 is correlated with a resistance to glucocorticoid-induced apoptosis but not to p53-mediated apoptosis. The sequence of mutations in the process of lymphomagenesis seems to be composed of at least 3 main hits which equip the cells with independence from external mitogenic signals (activation of Ras/Raf), resistance to inducers of apoptosis (activation of Bcl-2) and generation of cellular heterogeneity (deletion of p53) important in tumor progression. PMID- 10722232 TI - HLA-DR antigens in patients with pulmonary tuberculosis in northern Poland. Preliminary report. AB - The aim of the present study was the analysis of the association between particular class II HLA antigens and the incidence of tuberculosis in northern Poland. HLA-DR antigens in a group of 26 patients with pulmonary tuberculosis (PTB) and 58 healthy volunteers were determined. Histocompatibility typing was performed by the PCR-SSP method using primers from the Dynal company. For statistical analysis, the chi2 test was used with Yates' correction. The probability values were weighted for the number of antigens tested (pc). The relative risk (RR) was calculated by Woolf's method. We found that HLA-DR16(2) antigen expression was significantly higher in patients with tuberculosis than in the tested group of healthy controls (p < 0.001, pc < < 0.01); the highest relative risk (RR = 12.4) of tuberculosis incidence was connected with DR16(2) antigen, the prevalence of HLA-DR13(6) antigen was significantly lower in pulmonary tuberculosis (with RR = 0.09) than the control (p < 0.001, pc < 0.01). The results obtained suggest that the presence of HLA-DR16(2) antigen can extend the risk of developing tuberculosis whereas HLA-DR13(6) antigen occurrence was significantly more rare in pulmonary tuberculosis than in healthy individuals and that the relative risk (RR = 0.09) can be connected to their relation with the genes of insusceptibility to tuberculosis. PMID- 10722233 TI - Novel Ru(III), Rh(III), Pd(II) and Pt(II) complexes with ligands incorporating azole and pyrimidine rings. I. Antiproliferative activity in vitro. AB - A number of co-ordination compounds of Ru(III), Rh(III), Pd(II) and Pt(II) with ligands incorporating azole and pyrimidine rings has been synthesized. The in vitro cell proliferation-inhibitory activity of these compounds was examined against human cancer cell lines: A 549 (lung carcinoma), LS-180 (colon cancer) and MCF-7 (breast cancer), using SRB technique. Six out of 13 compounds studied revealed cytotoxic activity in vitro. Inhibitory dose 50% (ID50) was lower than 4 microg/ml, which is an activity criterion accepted in conventional in vitro cytotoxic screening tests. Two compounds revealed weak cytotoxic activity with ID50 higher than 4 microg/ml and five compounds were inactive. PMID- 10722234 TI - Thrombus organization plays no major role in late neointimal formation after angioplasty in porcine coronary arteries. AB - Thrombus organization has been suggested to play a major role in late neointimal formation after coronary angioplasty. We sought to describe the time sequence of lesion formation after angioplasty in porcine coronary arteries and to quantify the relation between early thrombosis and late neointimal formation. Deep vessel wall injury was induced by conventional balloon angioplasty in the circumflex (CX) and right coronary (RCA) arteries and by retraction of a chain-encircled balloon in the left anterior descendent artery (LAD). Lesions were assessed by histomorphometry at days 0, 1, 4, 7, 14, 28, and 56 after angioplasty. A response to-injury index (lesion area/injury length) was determined for each artery. Angioplasty led to rupture/removal of media. Thrombus was present at the exposed adventitia at days 0, 1, and 4. From day 7, neointima was observed on the luminal side of the arterial wall. All thrombus had disappeared at day 28, at which only neointima was observed. Histomorphometry revealed that lesion formation after angioplasty was a gradually increasing process from day 0 to day 28 with no further growth from day 28 to day 56. Maximal thrombus size (day 4, RCA: 0.07+/ 0.04 mm, CX: 0.23+/-0.16 mm, LAD: 0.15+/-0.11 mm) was significantly smaller than late neointimal formation (day 28, RCA: 0.68+/-0.18 mm, CX: 0.63+/-0.23 mm, LAD: 0.71+/-0.18 mm) in all three arteries (p < .03). Lesion formation after angioplasty is a gradually increasing process for 4 weeks. Maximal thrombus size is about four times smaller than late neointimal formation. Thus, thrombus organization plays no major role in late neointimal formation. PMID- 10722235 TI - Histological analysis of coronary artery lesions in fatal postoperative myocardial infarction. AB - We sought to evaluate the underlying coronary pathology of fatal postoperative myocardial infarction (MI). It has been hypothesized that most MIs following noncardiac surgery occur in the setting of increased oxygen demand that exceeds coronary blood supply. However, most MIs not associated with surgery are caused by plaque rupture and intracoronary thrombosis. In a retrospective cohort study, we reviewed 1841 consecutive autopsy records from 1981 to 1995 at two institutions and identified 26 cases of postoperative MI with coronary arteries available. Plaque rupture was present in 12 cases (46%, 95% confidence interval [CI] 27%-67%). Of the 9 (35%) patients with intracoronary thrombus, 5 (56%; 19% of entire group) had total occlusion. Thrombus occurred on a >50% stenosis (by cross-sectional area) in a total of 33% (95% CI 16%-55%) of patients. The only statistically significant difference in clinical variables between patients with and without plaque rupture was longer interval from surgery to death in patients with plaque rupture (7.8+/-4.4 days versus 4.4+/-4.8 days; p = 0.047). In this autopsy series, coronary plaque rupture was associated with almost half of fatal postoperative MI cases. Strategies aimed at reducing triggers of plaque rupture with coronary occlusion might reduce postoperative MI fatality. PMID- 10722236 TI - Coronary artery erosion and dissection: an unusual complication of mitral annular calcification. AB - This article describes a 42-year-old male patient with a longstanding history of insulin-dependent diabetes mellitus, systemic arterial hypertension, and chronic renal failure. The patient had severe mitral annular calcification (MAC) identified at autopsy. This MAC was of an amorphous, caseous-appearing type; it displaced the posterior mitral valve leaflet and extruded into the myocardium of the lateral and posterolateral left ventricle to involve the epicardial surface. The MAC produced extramural erosion of the wall, and dissection into the media, of the first left/obtuse marginal coronary artery. This coronary artery involvement by, and other complications of, MAC are discussed. PMID- 10722237 TI - Increased expression of tumor necrosis factor-alpha in diabetic macrovasculopathy. AB - Large vessel disease, a common feature of diabetes mellitus, appears to run an aggressive course, but its cellular and molecular aspects remain partially elucidated. Although in common atherosclerosis and especially in other forms of accelerated vasculopathy, immunoinflammatory mechanisms participate in the disease process, it is unclear whether this is present in diabetic vasculopathy. We hypothesized that diabetic macrovasculopathy, compared with classical atherosclerosis, is associated with increased immunoinflammatory features and matrix accumulation. In this study, vessel segments obtained after lower-limb amputation for advanced atherosclerotic disease, from type 2 diabetic patients (n = 20; 68.9+/-10.9 years) and nondiabetic patients (n = 16; 67.1+/-14.6 years) were analyzed. Histological characteristics (extent of intimal proliferation, cellularity, and fibrosis) were semiquantitatively graded in the two lesion types. Using immunohistochemistry, the presence of T cells and macrophages, accumulation of fibronectin, and expression of tumor necrosis factor-alpha was also assessed. Histological features of these advanced atherosclerotic lesions were similar in the two lesions examined. By immunohistochemistry, a similar pattern of T-cell and macrophage infiltration and fibronectin accumulation was observed. Nevertheless, increased expression of tumor necrosis factor-alpha was observed in diabetic lesions (13/19 patients had positive staining), whereas only 2 of 16 lesions from nondiabetic patients had positive staining (p < 0.003), with an odds ratio of 15.17 (confidence interval 2.12-139.5). These data suggest that increased expression of tumor necrosis factor-alpha observed in the diabetic lesions may reflect an enhanced inflammatory activity associated with the development of vascular lesions in type 2 diabetic patients. PMID- 10722238 TI - In vivo assessment of a novel dacron surface with covalently bound recombinant hirudin. AB - Prosthetic arterial graft surfaces are relatively thrombogenic and fail to heal with a cellular neointima. The goal of this study was to characterize the in vivo antithrombin properties of a novel Dacron surface with covalently linked recombinant hirudin (rHir) implanted in a canine thoracic aorta with high flow and shear rates. rHir was bound to a knitted Dacron patch using crosslinker modified bovine serum albumin (BSA) as a basecoat protein. BSA was first reacted with the heterobifunctional crosslinker, sulfo-SMCC. This BSA-SMCC complex was then bound to the carboxylic acid groups of hydrolyzed Dacron patches using the carbodiimide crosslinker, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride. Iodinated, Traut's-modified rHir (125I-rHir-SH) was then reacted with the Dacron-BSA-SMCC surface, thereby covalently binding 125I-rHir. Graft segments were washed and sonicated to remove any nonspecifically bound 125I-rHir. Dacron-BSA-SMCC-S-125I-rHir patches (n = 5) and control Dacron-BSA patches (n = 5) were implanted in series in the thoracic aortas of canines. These patches were exposed to nonheparinized, arterial blood flow for 2 hours. Patches were explanted and assessed for 125I-rHir loss. Antithrombin activity of explanted 1 cm2 patch segments was evaluated using a chromogenic assay with 1, 5, 10, 15 units of added thrombin. Light microscopy was performed to qualitatively examine the pseudointima. Two animals were excluded from the study owing to excessive bleeding through the knitted 125I-rHir patch. Comparison of preoperative and postoperative 125I-rHir gamma counts revealed an overall decrease of 20+/-5.4% over the period studied. Explanted 125I-rHir patch segments were able to inhibit 1, 5, and 7 NIHU of thrombin, demonstrating retained antithrombin activity. Gross and microscopic examination of the control and test Dacron surfaces showed marked differences. Dacron surfaces with covalently bound 125I-rHir had no gross thrombus and a thin pseudointima of platelets and plasma proteins. In contrast, the control patches had a thick pseudointima composed of fibrin-rich thrombus. rHir, covalently bound to Dacron patches, maintains its biologic activity as well as prevents thrombus formation on the graft surface. This novel antithrombin coating, by modifying the blood/ graft interface, may improve both short- and long-term patency in small-diameter prosthetic arterial grafts and has applications with respect to other implantable or indwelling biomaterials. PMID- 10722239 TI - Time course of the regression of intimal hyperplasia in experimental vein grafts. AB - A previous study in which vein grafts were removed from the arterial circulation and reimplanted into the venous circulation of the same animal demonstrated regression of vein graft intimal hyperplasia and medial thickening within 14 days. The present study was designed to characterize the kinetics of the morphological and ultrastructural changes over this 14-day period. Twenty-one male New Zealand White rabbits received a reversed vein interposition bypass graft of the right common carotid artery. Fourteen days after the procedure, 21 vein grafts were isolated, removed, and reimplanted into the contralateral external jugular venous system as veno-venous interposition bypass grafts (reversal grafts). The grafts were harvested at 60 minutes, 1 day, 3 days, 5 days, 7 days, and 14 days after reversal. Before insertion into the venous circulation, the vein graft had a confluent endothelial cell surface with multiple layers of smooth muscle cells representing intimal hyperplasia. After 1 hour, the reversal graft retained an intact endothelial cell layer with no evidence of tissue edema or cellular disruption. By 24 hours, there were a few blood cells on the endothelial cell surface. There was no inflammatory infiltrate seen in the subendothelium, and the smooth muscle cells were unaltered. At 3 days, the endothelial cell lining remained intact with no polymorphonucleocytes in the subendothelium or within the graft wall. Underlying smooth muscle cells at this time were noted to contain cytoplasmic vacuoles. At 5 days, there were no inflammatory cells seen on the surface or within the vein graft wall, but many of the underlying smooth muscle cells within the intimal hyperplasia were noted to be fragmented and to have clumping of chromatin. After 7 days, the endothelial cells remained intact and there was widespread evidence of apoptosis beneath the subendothelium with highly fragmented smooth muscle cells, some of which were histologically in the process of breaking up. At 14 days, the grafts retained uniform endothelial cell surfaces. Most of the smooth muscle cells that composed the intimal hyperplasia seen before implantation as a reversal graft were gone. Areas of newly laid down collagen could be observed. There were no acute inflammatory cells but for some mast cells seen in the graft wall. This study demonstrates that in this model, regression of intimal hyperplasia was associated with apoptosis of the smooth muscle cells and the deposition of collagen. There was no evidence that this process is mediated by an acute inflammatory response. Regression therefore appears to be due to induction of smooth muscle cell apoptosis by either a reduction in pressure or flow or a combination of both factors. The findings will enable a systematic cellular and molecular analysis of the biology of regression, which may afford clues to better understand the biology of the developing intimal hyperplasia. PMID- 10722240 TI - Characterization and quantitation of extracellular collagen matrix in myocardium of pigs with spontaneously occurring hypertrophic cardiomyopathy. AB - The extracellular matrix between cardiocytes has been suggested to play an important role in maintaining the structure and function of the heart. The purpose of this study was to elucidate the morphological changes in the collagen of the extracellular matrix (ECM) in the hearts of pigs with hypertrophic cardiomyopathy. Sixty pigs diagnosed with hypertrophic cardiomyopathy from 605 purebred Landrace pigs ages 6 to 9 months were used in this study. Morphologically, these pigs with hypertrophic cardiomyopathy had increased heart weight and heart-to-body weight ratio, thickening of the left ventricular (LV) and right ventricular (RV) free walls and septum, disorientation of cardiocytes, myocardial fibrosis, and intramural coronary arteriosclerosis. Similar observations have been described in our preliminary report (Cardiovasc Pathol 3:261, 1994). In the present study, we have modified the silver impregnation technique to stain paraffin-embedded sections to demonstrate three types of ECM. There were endomysial struts, perimysial weaves, and epimysial coils in the myocardium. The light microscopic findings of the struts, weaves, and coils were also confirmed by scanning electromicroscopic examination. The numbers of these fine structures were increased significantly in the pigs with hypertrophic cardiomyopathy. In addition, the amounts of collagen in the LVs, RVs, and septum (Sep) in pigs with hypertrophic cardiomyopathy (LV = 19.37+/-0.79, RV = 23.72+/ 0.72, Sep = 20.38+/-0.94 microg/mg, n = 60) were significantly higher (p < 0.01) than that in similar areas of normal pigs (LV = 14.56+/-1.11, RV = 18.90+/-1.02, Sep = 14.99+/-1.33 microg/mg, n = 30, respectively). Our findings of an overall increase of collagen content suggested that the accumulation of collagen matrix might be another factor responsible for the diastolic dysfunction of hypertrophic cardiomyopathy. These results might also infer that the increased collagen matrix could contribute to the stiffness of the cardiac chambers, thereby markedly affecting systolic and diastolic function of the heart. These observations provide further support that the pig may be an animal model for human cardiovascular disease. PMID- 10722241 TI - Anitschkow and the cholesterol over-fed rabbit. PMID- 10722242 TI - Myocardial connexin43 expression in left ventricular hypertrophy resulting from aortic regurgitation. AB - Intercellular conduction in the working myocardium of the mammalian heart is mediated by gap junctions composed of connexin43 or 45. Recently, it has been shown that myocardial connexin expression is malleable and may be altered with disease. To better understand myocardial conduction in left ventricular hypertrophy resulting from volume overload, we used indirect immunofluorescence microscopy to examine cardiac connexin43 expression in 10 New Zealand white rabbits with surgically induced aortic regurgitation (AR) and in 10 age-matched sham-operated controls. Animals were sacrificed at approximately 1 month or > or =2.5 years after operation. All AR animals developed eccentric hypertrophy; none evidenced heart failure. The heart-to-body weight ratios for the 1 month AR and control groups were 2.9+/-0.8 vs 1.8+/-0.2 g/kg (p < or = 0.01) while ratios for the > or =2.5 year AR and control groups were 2.4+/-0.3 vs 1.9+/-0.3 (p < or = 0.05). No significant differences in posterior wall thickness were found among any of the groups. Although the overall pattern of connexin43-like immunoreactivity was similar for all four groups, staining in the I month AR animals tended to be less than that of age-matched controls; staining was increased in the > or =2.5 year AR animals and was greater than control (p < 0.05), in which staining did not change with animal age. This disease duration related increase differs from the long-term decrease in connexin43 expression associated with other forms of heart disease and suggests that alterations in connexin expression may play a role in the rhythm abnormalities commonly seen in AR. PMID- 10722243 TI - Activation of calpain in myocardial infarction: an immunohistochemical study using a calpain antibody raised against active site histidine-containing peptide. AB - Tissue damage resulting from ischemia due to myocardial infarction is thought to be intensified by the proteolytic action of endogenous enzymes. Calpain (calcium dependent cysteine protease) is considered to be a highly likely candidate, since it is activated by calcium ion which increases in concentration under conditions of ischemia. We prepared a mono-specific antibody against the active site histidine stretch, Lys-Leu-Val-Lys-Gly-His-Ala-Tyr-Ser-Val, in the calpain 80 kDa large subunit. The specificity of the antibody was verified by its inhibitory effect on the caseinolytic activity of both mu- and m-calpains, western blotting analysis, and by absorption with the antigen peptide. The antibody was used to localize the intracellular distribution of activated calpains in infarcted regions of the human heart. The results showed that myocardial cells affected by ischemia were stained by the antibody, allowing damaged cells to be distinguished from cells of unaffected regions and that the immunostained regions were essentially the same regions as those identified by dense eosinophilic staining with hematoxylin and eosin. However, the staining pattern obtained with the antibody, was characteristic in denser staining at the cell periphery, whereas the damaged cells were stained homogeneously by hematoxylin and eosin. By the former method, results of staining indicated that the activation site of the calpain proenzyme was in the peri-plasma membrane, whereas by the latter method, diffusely distributed plasma proteins such as albumin and immunoglobulins were visualized as demonstrated in earlier reports. PMID- 10722244 TI - Discordance between pre and post cardiac transplant diagnosis: implications for pre- and postoperative decision making. AB - A correct clinical diagnosis in end-stage patients undergoing cardiac transplantation may have important prognostic and therapeutic implications. A retrospective clinico-pathologic study was carried out in 257 patients who had undergone cardiac transplantation at the University of Padua. A discrepancy between clinical and pathological diagnosis was found in 20 cases (8%). Among 126 patients with the clinical diagnosis of dilated cardiomyopathy, seven were found eventually to have ischemic heart disease (IHD), five myocarditis, one arrhythmogenic right ventricular cardiomyopathy (ARVC), and one non-compacted myocardium. Among the 87 patients with clinical diagnosis of IHD, three turned out to be dilated cardiomyopathy and one granulomatous myocarditis. Among the 10 patients with the clinical diagnosis of hypertrophic-restrictive cardiomyopathy, one had ARVC and one had cardiac fibroma. Altogether, only 24.5% underwent endomyocardial biopsy (EMB) and 75% coronary angiography before transplantation. Missed diagnosis of myocarditis occurred in patients in whom EMB was not carried out. EMB and coronary angiography might be indicated routinely in patients with apparent dilated cardiomyopathy, before proceeding to cardiectomy. PMID- 10722245 TI - Unusual myocardial (myostromal) "repair" in cardiac transplant patient. AB - A 57-year-old man received a cardiac allograft for severe ischemic heart disease. His endomyocardial biopsy at eight weeks postoperatively showed a focus of unusual myocardial morphology characterized by small diameter myocytes associated with loose, myxoid appearing stroma and a myocytic mitotic figure. We feel this may represent a unique type of myocardial repair. PMID- 10722246 TI - Histological and ultrastructural features of atherosclerosis in progeria. AB - This histological and ultrastructural study of a limited amount of vascular tissue from a progeric woman of 20 years who died of traumatic subdural hemorrhage supports the belief that the vascular changes are atherosclerotic. The unusual features observed were collagen fibrils with a relatively small diameter in the atherosclerotic intima and media, extensive loss of mural smooth muscle cells particularly in the aorta, and widespread contraction bands in smooth muscle cells in vascular and nonvascular tissues. Smooth muscle cells appear to be unusually susceptible to hemodynamic and ischemic stress. Further autopsy studies are required to elucidate the etiology and pathogenesis of this unique disease. PMID- 10722247 TI - Distribution of collagen deposition in cardiomyopathic hamster hearts determined by infrared microscopy. AB - Fibrillar collagens are major proteins of the cardiac extracellular matrix and play a significant role in the structural organization of the healthy heart. The aim of this study was (i) to investigate and compare the patterns of cardiac collagen deposition in different layers taken from both cardiomyopathic and normal myocardium using infrared (IR) microspectroscopy and (ii) to evaluate IR microspectroscopy as an alternative means for in vitro detection of collagen deposition in heart. Frozen sections from UM-X7.1 strain hamsters expressing the cardiomyopathic phenotype associated with ventricular remodeling and age-matched control (F1-beta) strain hamsters were examined using IR microspectroscopy. The presence of collagen was identified by the appearance of a typical collagen band at 1204 cm(-1), and the results were compared with identical tissue sections stained with trichrome, a routine discriminator for interstitial matrix proteins in cardiac myocytes. Spatial information addressing collagen deposition was obtained and viewed using contour mapping and three-dimensional band intensity maps at 1204 cm(-1). Perivascular and interstitial collagen deposition was detected in control samples taken from both ventricles as indicated with relative low intensities of the band of 1204 cm(-1). When compared with these control levels, the concentration of collagen was increased in cardiomyopathic left ventricular samples with some focal depositions, and these results were confirmed with the trichrome references. Our study suggests that collagen deposition from normal and diseased hearts may be successfully analyzed directly in the absence of any chemihistological or immunological staining, by infrared microscopy. PMID- 10722248 TI - Hypoplastic left heart syndrome with restrictive atrial septal defect and congenital pulmonary lymphangiectasis. AB - The presence of a restrictive atrial septal defect in hypoplastic left heart syndrome represents a surgical emergency and may negatively affect survival after operation. A neonate with such a disease association, requiring septectomy upon birth developed intractable respiratory failure due to congenital pulmonary lymphangiectasis. The therapeutic implications of this rare pathologic condition are discussed. PMID- 10722249 TI - Bilateral Ebstein's malformation. PMID- 10722250 TI - Collaboration between the Society for Cardiovascular Pathology (SCVP) and the International Society for Applied Cardiovascular Biology (ISACB) is justified. PMID- 10722251 TI - Pain in the neck for a rheumatologist. AB - Rheumatologists are frequently asked to see whiplash patients for an independent medical examination. There are many diagnostic and other assessment issues which arise in the setting of an independent assessment, and which do not usually arise in the usual physician-patient relationship. This article thus reviews aspects of the history, physical examination, radiological evaluation, and causation by way of the accident that are of importance to the rheumatologist as independent examiner. We address questions such as Was a physical injury likely to have occurred in the accident? What are the pertinent psychosocial aspects in the history? What are useful diagnostic signs on examination? and What is the significance of radiological findings? These are important and sometimes troublesome questions to answer in the setting of the independent medical examination, but rheumatologists can look to recent literature and research findings to assist in answering these questions. PMID- 10722252 TI - Rheumatologists and neck pain. AB - Many authors have suggested that chronic pain syndromes are psychosocial in origin; maladaptive behaviours favoured by psychosocial and political factors. Sometimes this may be true, but neither the individual patients nor the accumulated scientific evidence deserve such a routine dismissal. In this editorial I will review issues of responsibility, the nature of referred pain and referred tenderness, evidence for the value of tender point examination as an objective measure, techniques of assessment of the cervical spine, techniques of assessment of pain behaviour, and the determinants of the specific symptom patterns associated with cervical injury. PMID- 10722253 TI - Isotype distribution of anti-Ro/SS-A and anti-La/SS-B antibodies in plasma and saliva of patients with Sjogren's syndrome. AB - In this study an ELISA assay was used to quantify the levels of antibodies against the recombinant Ro 52 kD, Ro 60 kD, and La 48 kD proteins in plasma and saliva of 17 Sjogren's syndrome patients. The levels of total IgG, IgA, and IgM were also quantified. About one third of the patients had salivary enrichment of IgA and IgM against the Ro 52 kD, Ro 60 kD, and La 48 kD antigens and IgG against La 48 kD, while no enrichment of IgG against Ro 52 kD and Ro 60 kD was found. Most correlations between plasma and saliva levels of antigen specific antibodies were highly significant, as were most correlations between focus score and levels of antigen specific antibodies in plasma and saliva. In total, the results support the hypothesis that autoantibodies are produced in the salivary glands. The strong correlation between focus score and plasma and saliva levels of autoantibodies, indicates that the local autoantibody production is a consequence of local inflammation. PMID- 10722254 TI - Health-related quality of life in primary Sjogren's syndrome, rheumatoid arthritis and fibromyalgia compared to normal population data using SF-36. AB - OBJECTIVE: To investigate the health-related quality of life in women with primary Sjogren's syndrome (prim SS) and compare with normative data and the health-related quality of life in women with rheumatoid arthritis (RA) and women with fibromyalgia. METHODS: A questionnaire including the MOS Short-Form 36 (SF 36) was completed by 42 prim SS women, 59 RA women, and 44 women with fibromyalgia. RESULTS: All three patient groups experienced a decreased quality of life level ranging from 5 to 65 % in all SF-36 scales compared to normative data. Differences between groups were seen in 7 of the 8 scales (p< or = 0.004). The prim SS patients experienced a higher quality of life level with regard to physical function than the women with RA and fibromyalgia, whereas in the psychological dimensions the quality of life level was comparable to that of the two other groups. CONCLUSION: The health-related quality of life was significantly decreased as compared to norms in prim SS women and comparable to the levels of women with RA and fibromyalgia. PMID- 10722255 TI - Mortality in a cohort of Norwegian patients with rheumatoid arthritis followed from 1977 to 1992. AB - This study aimed to describe the mortality pattern of Norwegian patients with rheumatoid arthritis (RA). The subjects were 149 patients (52 males(M)) who were discharged from a Norwegian rheumatology hospital in 1977 after their first admission for RA. 126 patients (85%) met the 1958 criteria for definite or classical RA. By the end of 1992, 2 patients, both with definite/classical RA, were lost to follow-up. Of the remaining 147 patients (51M), 68 (25M) had died. The overall standardised mortality ratio (SMR) was 149 (95% CI: 115-188). The mortality was significantly raised for females with SMR= 168 (120-223). The moderate increase in the male SMR of 126 (81-181) was restricted to the early years of follow up. Patients with definite/classical RA had a somewhat higher SMR (159 (120-202)). Excess deaths were due to malignant disease in males and cardiovascular disease in females. RA was mentioned on one third of the death certificates. This study supports previous findings that patients with RA have a reduced long term survival, most prominent in females. PMID- 10722256 TI - Social network size of female patients with rheumatoid arthritis compared to healthy controls. AB - To estimate how rheumatoid arthritis (RA), the disease duration, and level of physical disability, influence the total size of patients' social network and the size of different subsets. Two hundred sixty four female patients (mean age 57 yrs) with RA of more than 6 yrs duration (mean 20 yrs) were compared to 61 healthy controls matched for sex, age, and residential area. Network size was measured by Social Network Delineation Questionnaire (SNDQ), physical disability by Health Assessment Questionnaire (HAQ). RA patients had a significantly smaller total network compared to the healthy controls (RA: 15.8 persons; CONTROLS: 18.1), mostly due to a significant difference in the subset of important others in favour of the controls (RA: 1.1; CONTROLS: 2.3). There were no significant differences regarding the network size of family, friends, and neighbours. The same results remained after statistical control for sociodemographic variables. Neither disease duration nor physical disability had any significant association with network size. The interaction analysis did, however, show that non-working patients with long disease duration (> 15 yrs) had fewer important others than occupationally active patients. Furthermore, a high degree of physical disability was related to a smaller number of friends for patients > 57 yrs than for equally disabled patients below this age. Most patients with RA seem to maintain contact with the family network-members, despite the challenges connected with chronic disease. PMID- 10722257 TI - Co-existence of chronic fatigue syndrome with fibromyalgia syndrome in the general population. A controlled study. AB - OBJECTIVE: To determine the proportion of adults with fibromyalgia syndrome (FMS) in the general population who also meet the 1988 Centre for Disease Control (CDC) criteria for chronic fatigue syndrome (CFS). METHODS: Seventy-four FMS cases were compared with 32 non-FMS controls with widespread pain and 23 with localized pain, all recruited in a general population survey. RESULTS: Among females, 58.0% of fibromyalgia cases met the full criteria for CFS, compared to 26.1% and 12.5% of controls with widespread and localized pain, respectively (p=0.0006). Male percentages were 80.0, 22.2, and zero, respectively (p=0.003). Compared to those with FMS alone, those meeting the case definitions for both FMS and CFS reported a worse course, worse overall health, more dissatisfaction with health, more non CFS symptoms, and greater disease impact. The number of total symptoms and non CFS symptoms were the best predictors of co-morbid CFS. CONCLUSIONS: There is significant clinical overlap between CFS and FMS. PMID- 10722258 TI - Sonographic analysis of the ankle in patients with psoriatic arthritis. AB - Foot involvement is very frequent in patients affected by psoriatic arthritis (PsA). However, evaluation of the painful foot can be problematic, because it is often difficult to distinguish between arthritis, tenosynovitis, and enthesopathy. Plain radiographs can show bone erosion or other features of joint involvement, but give little information about the soft tissues. We therefore studied foot involvement in 31 PsA patients using high resolution sonography, and compared the results with the findings on x-ray and clinical examination. Ultrasound revealed pathological findings in a large proportion of the patients, most of whom exhibited no clinical (pain or swelling) or radiological signs of foot involvement at the time of the study. Our data suggest that involvement of the tendons and entheses may be more frequent in PsA patients than has thus far been supposed, even in cases of not particularly aggressive disease, and that clinical evaluation tends to underestimate these manifestations. PMID- 10722260 TI - The time of day that etidronate is ingested does not influence its therapeutic effect in osteoporosis. AB - To investigate whether the therapeutic effect of etidronate is affected by the time of ingestion we have retrospectively studied 110 osteoporotic patients. They had been taking etidronate according to the manufacturer's instructions for a mean duration of 2.6 years either on waking (n=47), on retiring (n=47), or during the day (n= 16). No significant differences were found between the three groups with respect to percentage change in bone mineral density at all sites either for the first year of treatment or for the mean yearly percent change throughout the total course of treatment. These findings suggest that the time of ingestion of etidronate does not influence the therapeutic effect if a two hour fast before and after ingestion is adhered to. PMID- 10722259 TI - Ultrasonography in the quantification of arterial involvement in Takayasu's arteritis. AB - OBJECTIVE: To evaluate the accuracy of ultrasonography (US) in the detection and quantification of arterial involvement in Takayasu's arteritis (TA). METHODS: The common carotid and subclavian arteries, and the abdominal aorta of 15 patients with TA were studied by Color Doppler (CD) US and Doppler spectral analysis and compared with angiography. RESULTS: The mean difference (+/-SD) between the percent luminal stenoses measured at angiography and by CD US was 2.0+/-10.3% for the common carotid artery, 4.0+/-23.6% for the subclavian artery and -1.3+/-16.8% for the abdominal aorta. The differences were not statistically significant. However, the agreement of the methods was less than satisfactory as shown by the wide standard deviations. CONCLUSIONS: More efforts are needed to improve the less than optimal agreement of US with angiography regarding the severity of individual stenoses. The technique performs best in the study of carotid arteries. PMID- 10722261 TI - Mesangial nephropathy in Sjogren's syndrome. AB - We describe a 36-year-old woman with Primary Sjogren's Syndrome (PSS). Purpura, corneal perforation, metabolic acidosis, decreased glomerular filtration, hypokalemia, hyposthenuria, and polyuria were present. Chronic renal insufficiency and renal tubular acidosis type I were diagnosed. Kidney biopsy revealed mesangial glomerulonephritis, interstitial nephritis, and tubular atrophy. Replacement treatment with saliva, tears, and potassium citrate was started. She was given prednisone and cyclophosphamide. This would be the first description of PSS, mesangial glomerulonephritis, and chronic renal insufficiency. PMID- 10722263 TI - Avian Behavioural Neurosciences. Proceedings of a meeting. Tihany, Hungary, August 1996. PMID- 10722264 TI - Paper alert. Cell biology. PMID- 10722262 TI - Simultaneous presence of C-ANCA and P-ANCA in a patient with concurrent Churg Strauss syndrome and giant cell temporal arteritis. AB - We herein describe the case of a 77-year-old woman, who presented clinical and histopathological evidence of giant cell arteritis (GCA) involving the temporal artery, together with a Churg-Strauss syndrome (CSS). Our patient presented positive anti-neutrophil cytoplasmic antibodies (ANCA), with cytoplasmic staining pattern (C-ANCA) that was specific against proteinase 3 (PR3), and also a perinuclear pattern (P-ANCA) with specificity against myeloperoxidase (MPO). To our knowledge, the simultaneous presence in the same patient of both types of antibodies has not been previously reported. PMID- 10722265 TI - Web alert. Cytoskeleton. PMID- 10722266 TI - Research discredits rebel dentists. PMID- 10722267 TI - Increased serum neopterin levels in mycosis fungoides and Sezary syndrome. PMID- 10722268 TI - Successful treatment of dissecting cellulitis and acne conglobata with oral zinc. PMID- 10722269 TI - Finasteride as a therapy for hidradenitis suppurativa. PMID- 10722270 TI - A case of serratiopeptidase-induced subepidermal bullous dermatosis. PMID- 10722272 TI - Severity of disease in inflammatory cutaneous disease. PMID- 10722271 TI - Successful treatment of orogenital ulceration with transdermal nicotine patches. PMID- 10722273 TI - Acquired tufted angioma in association with a complex cutaneous vascular malformation. PMID- 10722274 TI - Porokeratotic eccrine ostial and dermal duct naevus: treatment with carbon dioxide laser. PMID- 10722275 TI - Kawasaki disease associated with varicella: a rare association. PMID- 10722276 TI - Childhood cutaneous T-cell lymphoma in association with pityriasis lichenoides chronica. PMID- 10722277 TI - Cidofovir for the treatment of Kaposi's sarcoma in an HIV-negative homosexual man. PMID- 10722278 TI - Confluent and reticulated papillomatosis: treatment with topical calcipotriol. PMID- 10722279 TI - Undeclared constituents in antiscabetic treatments. PMID- 10722280 TI - Mees' lines. PMID- 10722281 TI - MAP kinase localizes to the platelet-yielding demarcation membrane system in megakaryocytes. PMID- 10722282 TI - The casual relation between benzene exposure and multiple myeloma. PMID- 10722283 TI - Paper alert. Biotechnology. PMID- 10722284 TI - Web alert. Analytical biotechnology. PMID- 10722285 TI - Analytical biotechnology: sorting needles and haystacks. PMID- 10722286 TI - True symbol of medicine. PMID- 10722287 TI - First myocardial infarction in patients of Indian subcontinent and European origin: comparison of risk factors, management, and long term outcome. PMID- 10722288 TI - Home treatment--engimas and fantasies. PMID- 10722289 TI - Debate on cot death. Standard system for postmortem examination and certification needs to be agreed. PMID- 10722290 TI - Debate on cot death. In 1994, 29% of suspicious deaths were officially recorded as due to sudden infant death syndrome. PMID- 10722291 TI - Stable chronic obstructive pulmonary disease. Published correction for one of studies must be borne in mind. PMID- 10722292 TI - Moving the research agenda. Academic GPs can get research ideas from their experiences in practice. PMID- 10722294 TI - Commentary: why do doctors overestimate? PMID- 10722295 TI - Commentary: prognoses should be based on proved indices not intuition. PMID- 10722293 TI - Moving the research agenda. RCGP is encouraging improvements in primary care research. PMID- 10722297 TI - Notice of inadvertent duplicate publication. PMID- 10722296 TI - Commentary: much still to learn about relations between tumour biology, prognosis, and treatment outcome in early breast cancer. PMID- 10722298 TI - Doctors write on patients' eye view of quality. Demand is too great for GPs to provide patients with longer appointments. PMID- 10722299 TI - Doctors write on patients' eye view of quality. Patients must "get real". PMID- 10722300 TI - Doctors write on patients' eye view of quality. Longer consultation time that patients wish for is not available in NHS. PMID- 10722301 TI - Accuracy of perceptions of hepatitis B and C status. Results require further clarification. PMID- 10722302 TI - Accuracy of perceptions of hepatitis B and C status. Value of screening for hepatitis C is still debatable. PMID- 10722303 TI - Melanocyte and melanogenesis. Proceedings of the 8th meeting of the European Society for Pigment Cell Research. Prague, Czech Republic, September 23-26, 1998. PMID- 10722304 TI - Genetic relationship of 32 cell lines of Euplotes octocarinatus species complex revealed by random amplified polymorphic DNA (RAPD) fingerprinting. AB - Random amplified polymorphic DNA (RAPD) fingerprinting was used in this study to determine the genetic relationship of different cell lines of the hypotrichous ciliate Euplotes octocarinatus. Stocks isolated from different habitats in the USA, and from a group of genetically recombined laboratory strains, were characterized. Band-sharing indices (D) for all possible pairwise comparisons revealed a remarkable genetic diversity between the different cell lines. Investigation of the genetic structure in natural populations found diversity- although to a different extent--in all populations investigated. No clonal structure could be observed, as proposed for several protozoa and recently shown for E. daidaleos. These findings suggest frequent conjugation in the population of E. octocarinatus. No correlation between the genetic relationship of cell lines from different habitats and the distance between the corresponding sampling locations was found. Once separated geographically, the exchange of genetic material between populations appears to be nearly impossible. Therefore, these groups tend to separate into sibling species. The data generally support the occurrence of different syngens in the E. octocarinatus species complex. This finding is in accordance with our observation that the morphological 'species' of E. octocarinatus consists of several syngens or sibling species, similar to findings for the Paramecium aurelia-, Tetrahymena pyriformis- and E. vannus- species complexes. PMID- 10722305 TI - Seeds of conflict. AB - Recent studies have shown that the Arabidopsis MEDEA gene is imprinted, so that paternally and maternally inherited alleles are differentially expressed during seed development. Futhermore, a chromatin remodelling factor has been implicated as a novel trans-acting regulator of imprinting. PMID- 10722306 TI - Hormone resistance and new aspects of GH therapy. Proceedings of the 2nd Workshop on Clinical Paediatric Endocrinology. Salzburg, Austria, September 7-8, 1998. PMID- 10722307 TI - Canadian Cardiovascular Society 1998 Consensus Conference on the Prevention of Cardiovascular Diseases: The Role of the Cardiovascular Specialist. PMID- 10722308 TI - Preventing cardiovascular diseases in people with diabetes. PMID- 10722310 TI - International symposium on cardiac rhythms in animals.Muroran, Japan, March 1998. PMID- 10722309 TI - 6th Annual Congress and 1st Advanced Course of the European Society of Magnetic Resonance in Neuropediatrics (ESMRN). Marseille, France, 19-22 January 2000. Abstracts. PMID- 10722311 TI - Urinary tract infections. PMID- 10722313 TI - Head tilt during driving. AB - In order to distinguish between the use of visual and gravito-inertial force reference frames, the head tilt of drivers and passengers were measured as they went around corners at various speeds. The visual curvature of the corners were thus dissociated from the magnitude of the centripetal forces (0.30-0.77 g). Drivers' head tilts were highly correlated with the visually-available estimate of the curvature of the road (r2=0.86) but not with the centripetal force (r2<0.1). Passengers' head tilts were inversely correlated with the lateral forces (r2=0.3-0.7) and seem to reflect a passive sway. The strong correlation of the tilt of drivers' heads with a visual aspect of the road ahead, supports the use of a predominantly visual reference frame for the driving task. PMID- 10722314 TI - Yra1p, a conserved nuclear RNA-binding protein, interacts directly with Mex67p and is required for mRNA export. AB - Mex67p and Mtr2p constitute an essential mRNA export complex that interacts with poly(A)+ RNA and nuclear pore proteins. We have identified Yra1p, an intranuclear protein with in vitro RNA-RNA annealing activity, which directly binds to Mex67p. The complex between Yra1p and Mex67p was reconstituted in vitro and shown by UV crosslinking to bind directly to RNA. Mutants of YRA1 are impaired in nuclear poly(A)+ RNA export at restrictive growth conditions. ALY, the mouse homologue of Yra1p and a transcriptional coactivator, can bind in vitro to yeast and human Mex67p and partly complements the otherwise non-viable yra1 null mutant. Thus, Yra1p is the first RNA-binding protein characterized, which bridges the shuttling Mex67p/Mtr2p exporter to intranuclear mRNA transport cargoes. PMID- 10722315 TI - Cost-effectiveness of interferon alfa 2b and ribavirin in the treatment of chronic hepatitis C. PMID- 10722316 TI - Hepatitis C virus infection and lymphoproliferative disorders after liver transplantation. PMID- 10722317 TI - Carvedilol--A new nonselective beta blocker. PMID- 10722318 TI - Hepatitis C and autoimmune hepatitis. PMID- 10722319 TI - [Promotion of complete, follow-up tumor documentation]. PMID- 10722320 TI - ICTR 2000. 1st International Conference on Translational Research and Pre Clinical Strategies in Clinical Radio-Oncology. Lugano, Switzerland, March 5-8, 2000. Abstracts. PMID- 10722321 TI - Interference of Pseudomonas strains in the identification of Helicobacter pylori. PMID- 10722322 TI - Horizontal transmission of a hepatitis B virus surface antigen mutant. PMID- 10722323 TI - Criteria for performing PCR in cases of suspected herpes encephalitis. PMID- 10722324 TI - Molecular basis for distinction of the ET-15 clone within the ET-37 complex of Neisseria meningitidis. PMID- 10722325 TI - PCR for detection of bacteremia. PMID- 10722326 TI - Variation in microfilariae and infective stages of two types of Wuchereria bancrofti from the Thai-Myanmar border. AB - Comparative morphometric and morphological studies of microfilariae and infective stages were undertaken in nocturnally periodic and subperiodic Wuchereria bancrofti. For microfilariae, the body dimensions of nocturnally periodic (NP) were significantly smaller than nocturnally subperiodic (NSP), i.e., body length 268.03+/-14.75 microm (NP), 307.61+/-11.52 microm (NSP); cephalic space length 4.21+/-0.62 microm (NP), 5.32+/-0.79 microm (NSP); head to nerve ring 49.39+/ 5.43 microm (NP), 57.40+/-4.46 microm (NSP); innenkorper length 33.05+/-5.89 microm (NP), 44.02+/-8.71 microm (NSP); cephalic space width 4.28+/-0.59 microm (NP), 6.04+/-0.68 microm (NSP); body width at nerve ring 5.01+/-0.57 microm (NP), 7.45+/-0.75 microm (NSP). The number of nuclei between the cephalic space and nerve ring of NP (66.67+/-5.19) was also significantly less than in NSP (94.74+/ 6.95). For infective stages, the body dimensions of NP were significantly smaller than NSP, i.e., body length 1632.50+/-131.48 microm (NP), 2002.63+/-222.60 microm (NSP); head to nerve ring 103.09+/-7.47 microm (NP), 122.44+/-9.62 microm (NSP); head to oesophago-intestinal junction 567.69+/-94.84 microm (NP), 666.75+/-110.08 microm (NSP); body width at oesophago-intestinal junction 23.15+/-1.55 microm (NP), 26.78+/-1.62 microm (NSP). It is too early to infer the NP type as an additional sibling species of W. bancrofti but it is reasonable to treat it as a new variety and additional work is needed to clarify its status. PMID- 10722327 TI - Elevated MIA serum levels are of relevance for management of metastasized malignant melanomas: results of a German multicenter study. PMID- 10722328 TI - Barrier function in K-10 heterozygote knockout mice. PMID- 10722329 TI - Chronic liver disease. PMID- 10722330 TI - Proceedings of the 5th International Symposium on Reproduction in Domestic Ruminants. Colorado Springs, Colorado, USA. 1-5 August 1998. PMID- 10722331 TI - Phytoestrogens. PMID- 10722332 TI - Aspirin for primary prevention of cardiovascular disease. PMID- 10722333 TI - Osteopathic treatment of low back pain. PMID- 10722334 TI - Osteopathic treatment of low back pain. PMID- 10722335 TI - Osteopathic treatment of low back pain. PMID- 10722336 TI - Osteopathic treatment of low back pain. PMID- 10722337 TI - Osteopathic treatment of low back pain. PMID- 10722338 TI - Osteopathic treatment of low back pain. PMID- 10722339 TI - Osteopathic treatment of low back pain. PMID- 10722340 TI - The economic implications of HLA matching in cadaveric renal transplantation. PMID- 10722341 TI - Treatment of Alzheimer's disease. PMID- 10722342 TI - Treatment of Alzheimer's disease. PMID- 10722343 TI - Administration of epinephrine by emergency medical technicians. PMID- 10722344 TI - Antiapoptotic agents in brain ischemia. PMID- 10722345 TI - Long-term care for the frail elderly. PMID- 10722346 TI - Necrotizing fasciitis due to Photobacterium damsela in a man lashed by a stingray. PMID- 10722347 TI - Proposal for an addendum to the recommendations made by the European College of Radiological Education and the Education Committee Working Group. PMID- 10722348 TI - Absence of evidence is not evidence of absence (again) response to Dr. Fernando Cervero and Dr. Jennifer M.A Laird. PMID- 10722349 TI - R3 component of the blink reflex is not a suitable model to investigatetrigeminal nociception. Comment on Jaaskelainen et al., PAIN 80 (1999) 191-200. PMID- 10722350 TI - When the whole is more than the sum of its parts, comments on Treede et al; PAIN 79 (1999) 105-111. PMID- 10722351 TI - Comments on Treede et al; PAIN 79 (1999) 105-111. PMID- 10722352 TI - Comments on Moore et al; PAIN 78 (1998) 209-216. PMID- 10722353 TI - The bigger the better or small but perfectly formed? PMID- 10722354 TI - Comments on France and Suchowiecki, PAIN 81 (1999) 77-84. PMID- 10722355 TI - Index of suspicion. Case 2. Diagnosis: Henoch-Schoenlein purpura. PMID- 10722356 TI - Index of suspicion. Case 3. Diagnosis: septic arthritis. PMID- 10722357 TI - Image interpretation session: 1998. Bilateral epididymal cystadenomas, pancreatic and renal cysts, and cerebellar hemanigioblastoma in the setting of VHL disease. PMID- 10722358 TI - Image interpretation session: 1998. Malignant melanoma metastatic to the liver and spine. PMID- 10722359 TI - Image interpretation session: 1998. Langerhans cell histiocytosis (LCH) with pulmonary and bone involvement. PMID- 10722360 TI - Image interpretation session: 1998. Thyroglossal duct cyst with a nodule of thyroid tissue containing papillary adenocarcinoma and cervical lymph node metastases. PMID- 10722361 TI - Image interpretation session: 1998. Pulmonary arteriovenous malformation (AVM) in the setting of hereditary hemorrhagic telangiectasia (HHT). PMID- 10722362 TI - Image interpretation session: 1998. Plexiform neurofibroma of the genitourinary tract in the setting of neurofibromatosis type 1. PMID- 10722363 TI - Image interpretation session: 1998. Cecal and pulmonary sarcoidosis. PMID- 10722365 TI - Image interpretation session: 1998. Papillary endolymphatic sac tumor. PMID- 10722364 TI - Image interpretation session: 1998. Tuberous sclerosis with multiple bilateral renal cysts and a subependymal giant cell astrocytoma. PMID- 10722367 TI - Proceedings of the 9th International Meeting of the Leksell Gamma Knife Society. Hong Kong, November 1998. PMID- 10722368 TI - [In-Hospital meeting of the AMUB (Association des Medecins anciens etudiants de l'Universite libre de Bruxelles): endocrine digestive tumors]. PMID- 10722366 TI - Image interpretation session: 1998. Swyer-James (Macleod) syndrome. PMID- 10722369 TI - Genes, receptors, signals and responses to lipopolysaccharide endotoxin. AB - C3H/HeJ inbred mice have been very useful for identifying genetic elements responsible for endotoxin mediated responses. Depending on the type of assays employed, Tlr-2, Tlr-4 and Lps/Ran have been shown to be important in lipopolysaccharide (LPS)-mediated responses. The concept of a single LPS gene being responsible for the genetic defect found in C3H/HeJ mice should therefore be re-examined more closely. Given the most recent discoveries, it is probable that more than one signal transduction pathway is involved. One is a CD14 dependent pathway, the other a CD14-independent pathway. Identification of the genetic elements involved in these pathways will be beneficial in designing therapeutic strategies for treating patients with endotoxic or septic shock. PMID- 10722370 TI - Possible role of the plasminogen receptor as a site of interaction of the human immunodeficiency virus p24 immunosuppressive heptapeptide Ch7 with the host immune system. AB - Previous work from our laboratory demonstrated that a synthetic heptapeptide (Ch7: RGSDIAG), corresponding to a conserved sequence of human immunodeficiency virus (HIV) core protein p24 (amino acids 232- 238), was able to specifically abrogate antigen-induced responses in cultures of normal human peripheral blood mononuclear cells (PBMC), probably acting at the level of monocytes. The Ch7 peptide displays sequence homology to human plasminogen. In the present report we show that a compound (6-aminoexanoic acid), known to prevent plasminogen binding to monocyte-like cells, greatly reduced the immunosuppressive capacity of Ch7. We suggest that the plasminogen receptor may represent a target structure on human monocytes for the immunosuppressive p24 sequence. PMID- 10722371 TI - Effects of subcutaneous interleukin-2 therapy on phenotype and function of peripheral blood mononuclear cells in human immunodeficiency virus infected patients. AB - In the context of clinical therapy with recombinant human interleukin-2 (IL-2), we monitored immunological alteration in 10 human immunodeficiency virus type-I (HIV-1)-infected individuals, on stable antiretroviral therapy, who had a CD4+ cell count between 200 and 500 cells/mm3. Subcutaneous IL-2 was prescribed thrice weekly (at a dose of 3 x 10(6) IU) for 24 weeks and the patients were followed-up for 32 weeks. IL-2 treatment induced an increase in the CD4+ percentage (P<0.001) and CD4+ cell count (P<0.009). Furthermore. natural killer (NK) cell activity was increased (P<0.001) at week 8 of treatment, whereas lymphokine-activated killer (LAK) cell activity showed a transient, nonsignificant increase at week 8 and was reduced (P<0.001) at 32 weeks. However, the cytotoxic T-lymphocyte (CTL) activity decreased against HIV antigens, and the proliferative response to Candida, IL-2 and phytohaemagglutinin (PHA) declined during the first 8 weeks (P<0.05) and returned to baseline levels after 32 weeks. The HIV RNA level did not change during IL-2 therapy; however, after 8 weeks of follow-up a significant increase (P<0.001) in viral load was observed. In PMID- 10722372 TI - Human T-cell response to meningococcal immunoglobulin A1 protease associated alpha-proteins. AB - A unique feature of the immunoglobulin A1 (IgA1) protease from pathogenic Neisseriae, i.e. N. meningitidis and N. gonorrhoeae, is its co-secretion with an amphipathic a-protein. Polymerase chain reaction (PCR) analysis of the respective iga(alpha). gene region in 48 meningococcal strains revealed that this protein domain is conserved throughout all isolates in four different principal variants. Despite strain-dependent size and sequence variations, sequence analysis showed common structural characteristics. More than 80% of the amino acid sequence of all a-proteins is dependent on the five amino acids Q, E, A, K and R, resulting in a pI> 10. The sequences are highly conserved at the N-terminus and the C terminus and contain long amphipathic alpha-helical stretches. These stretches have a strong probability of forming coiled coil conformations and comprise short repetitive sequence modules with pronounced similarities to T-cell epitopes. We therefore analyzed the T-cell response of 20 volunteer blood donors to four peptides, representing such predicted epitopes, and a recombinant meningococcal alpha-protein. Sixteen donors reacted against at least one peptide after culture of peripheral blood mononuclear cells in interleukin (IL)-2-rich medium, while two individuals showed a positive reaction only against an IgA1 protease-derived control peptide. From one donor, we established and maintained T-cell clones specific for purified alpha-protein. Characterization of the T-cell clones revealed a CD3- and a CD4-positive phenotype and the secretion of IL-2 and interferon-gamma (IFN-gamma), PMID- 10722373 TI - Genomics. Fruit fly genome yields data and validation. PMID- 10722374 TI - Intellectual property. HHS probes genesis of gene sequencer. PMID- 10722375 TI - Gene patents. Patent on HIV receptor provokes an outcry. PMID- 10722376 TI - Molecular genetics. Spider genes reveal flexible design. PMID- 10722377 TI - Neuroscience. New stroke treatment strategy explored. PMID- 10722378 TI - Neuroscience. Death triggers regrowth of zebra finch neurons. PMID- 10722380 TI - Ecology. The unbearable capriciousness of Bering. PMID- 10722379 TI - Meltdown on Long Island. PMID- 10722381 TI - Ecology. New corn plant draws fire from GM food opponents. PMID- 10722382 TI - In search of stem cell policy. PMID- 10722383 TI - Limits to our knowledge. PMID- 10722384 TI - Transgenic maize in Mexico: no need for concern. PMID- 10722385 TI - Firearms the target. PMID- 10722386 TI - Firearms the target. PMID- 10722387 TI - Policy forum: European policy. Interesting times--biology, European science, and EMBL. PMID- 10722388 TI - Perspectives: paleoecology. The refugial debate. PMID- 10722389 TI - Perspectives: microbiology. Mice are not furry petri dishes. PMID- 10722390 TI - Can old cells learn new tricks? PMID- 10722391 TI - The business of stem cells. PMID- 10722392 TI - Fetal neuron grafts pave the way for stem cell therapies. PMID- 10722393 TI - Proceedings of the 5th Congress of the Mediterranean Society of Clinical Pharmacology. Marrakech, Morocco, 28-30 October 1998. PMID- 10722394 TI - Developments in pretransfusion testing and compatibility testing. PMID- 10722396 TI - Satellite Symposium of the 9th Congress of the European Society for Organ Transplantation, Oslo, Norway. From Immunosuppression to Tolerance: Visions of the Future. Foreword. PMID- 10722395 TI - European neuroscience meets the snow. PMID- 10722397 TI - Apricot orchards: doctors and patients in Japan. PMID- 10722398 TI - Current awareness on yeast. PMID- 10722399 TI - Proceedings of the International Congress on Melioidosis. Bangkok, Thailand, November 22-25, 1998. PMID- 10722400 TI - American Thoracic Society develops guidelines on diagnosis of venous thromboembolism. PMID- 10722402 TI - Centers for Disease Control and Prevention. 4th Decennial International Conference on Nosocomial and Healthcare-Associated Infections. Atlanta,Georgia, USA. March 5-9, 2000. Abstracts. PMID- 10722403 TI - Molecular and clinical immunology and immunopathology. Association of Clinical Scientists meeting. La Jolla, California, USA. 4-7 November 1999. Abstracts. PMID- 10722401 TI - Mosaicism in Prader-Willi syndrome. PMID- 10722404 TI - [Hermann Niemeyer Fernandez (1918-1991) and science in Chile]. PMID- 10722405 TI - Adolescent inpatient units. PMID- 10722406 TI - Dipstick examination for urinary tract infections. PMID- 10722407 TI - Emergency management of meningococcal disease. PMID- 10722408 TI - Estimating the genetic potential in stature. PMID- 10722409 TI - Examination of children who may have been sexually abused. PMID- 10722410 TI - Iron fortified follow on formula from 9 to 18 months improves iron status but not development or growth. PMID- 10722411 TI - IGFBP-3 as a predictor of growth hormone deficiency. PMID- 10722412 TI - New developments in the treatment of cardiac failure. PMID- 10722413 TI - Pacifier use and SIDS. PMID- 10722414 TI - Planning for major incidents involving children by implementing a Delphi study. PMID- 10722415 TI - Prevalence of bruising in babies. PMID- 10722416 TI - Raised serum transaminases: not always liver disease. PMID- 10722417 TI - Recommendations for the management of galactosaemia. PMID- 10722418 TI - Transition from paediatric to adult care. Bridging the gaps or passing the buck? PMID- 10722419 TI - Use of duvets and SIDS. PMID- 10722420 TI - Microchip mess. PMID- 10722421 TI - The West Nile Virus outbreak of 1999 in New York: the Flushing Hospital experience. AB - West Nile Virus (WNV) is a mosquito-borne flavivirus, which has been known to cause human infection in Africa, the Middle East, and southwestern Asia. It has also been isolated in Australia and sporadically in Europe but never in the Americas. Clinical features include acute fever, severe myalgias, headache, conjunctivitis, lymphadenopathy, and a roseolar rash. Rarely is encephalitis or meningitis seen. During the month of August 1999, a cluster of 5 patients with fever, confusion, and weakness were admitted to the intensive care unit of the same hospital in New York City. Ultimately 4 of the 5 developed flaccid paralysis and required ventilatory support. Three patients with less-severe cases presented shortly thereafter. With the assistance of the New York City and New York State health departments and the Centers for Disease Control and Prevention, these were documented as the first cases of WNV infection on this continent. PMID- 10722422 TI - Giardia lamblia carriage in Israeli Bedouin infants: risk factors and consequences. AB - Giardiasis is a common protozoan infection, with varying clinical manifestations. We investigated the associations between Giardia lamblia carriage and environmental, family, illness, and growth characteristics. Bedouin infants (n=234) were followed from birth to age 18-23 months. At monthly home visits, stool samples were obtained, history of illness was determined, and an environmental assessment was done. The comparisons presented are between 4 groups defined by length of carriage of G. lamblia. Study children had a mean+/-SD of 4.1+/-2.9 diarrhea episodes. No illness, environmental, or family characteristics were associated with length of carriage. Significant differences were found in weight-for-age and weight-for-height z scores between the never-positive-for-G. lamblia group and all other carriage groups combined. Faltering growth was shown to be subsequent to G. lamblia infection rather than preceding it. Our findings confirm that G. lamblia carriage is not associated with diarrhea. However, the effect on growth deserves further investigation. PMID- 10722423 TI - Epidemiology of nosocomial infection and resistant organisms in patients admitted for the first time to an acute rehabilitation unit. AB - The objectives of this study were to define the epidemiology of nosocomial bacterial colonization and infection and to define predictors of nosocomial infection among a cohort (n=423) of admissions to an acute rehabilitation unit. Overall, methicillin-resistant Staphylococcus aureus (MRSA) and enterococci were the most commonly identified colonizing organisms. Escherichia coli and Pseudomonas aeruginosa were the most commonly identified colonizing gram-negative bacilli. During 70 (16.5%) of the 423 hospitalizations in the unit, 94 nosocomial infections occurred. The most common infections were those of the urinary tract (30% of 94 infections) or a surgical site (17%), Clostridium difficile diarrhea (15%), and bloodstream infection (12.8%). Antibiotic-resistant bacteria most commonly caused bloodstream infection (41.7%) and surgical site infection (56.3%). Independent predictors of nosocomial infection at the time of admission were functional status (measured with the functional independence measure), APACHE III score, and spinal cord injury. In conclusion, gram-positive organisms were the predominant strains causing nosocomial colonization and infection. The logistic model, if verified, may be useful in defining patients who should be targeted for measures to prevent nosocomial infection. PMID- 10722424 TI - National survey of extended-interval aminoglycoside dosing. AB - A random sample survey of 500 acute care hospitals in the United States was conducted to evaluate the adoption of extended-interval aminoglycoside dosing (EIAD). The survey revealed that EIAD has been adopted in 3 of every 4 acute care hospitals, a 4-fold increase since 1993. Of the 74.7% of hospitals reporting EIAD, 64% had written guidelines. Equal or less toxicity (87.1%), equal efficacy (76.9%), and cost-savings (65.6%) were common rationales. There has been a trend toward higher adult dosages of gentamicin (e.g., >5 mg/kg/dose) and an increase in the adoption of EIAD across all age groups (neonatal, 11%, and pediatric, 23%). Monitoring of aminoglycoside concentrations has shifted to a single determination of concentration, at 6-18 h after drug administration. The most common methods of dosage adjustment for declining renal function were an interval extension with the same dose (47%) or use of the Hartford nomogram (32%). PMID- 10722425 TI - Single daily dosing of aminoglycosides--A community standard? PMID- 10722426 TI - Mycobacterium terrae: case reports, literature review, and in vitro antibiotic susceptibility testing. AB - Mycobacterium terrae infection can cause debilitating disease that is relatively resistant to antibiotic therapy. Two cases are presented, and data from an additional 52 reports from the literature are reviewed. Tenosynovitis of the upper extremity, often following trauma, was the most commonly reported presentation (59% of cases), with pulmonary disease occurring in an additional 26% of cases. Underlying medical problems were absent (44%) or not reported (28%) in 72% of the cases. One-half of the patients with upper extremity tenosynovitis were treated with local or systemic corticosteroids, before microbiological identification. Only one-half of the patients with tenosynovitis who were followed up for 6 months had clinical improvement or were cured. The other one half of the patients required repeated debridement, tendon extirpation, or amputation. The best antimicrobial therapy for M. terrae infection is unknown but might include a macrolide antibiotic plus ethambutol and one other effective drug for at least 12 months after clinical response. Parenteral treatment with an aminoglycoside and surgery may be useful in selected cases. PMID- 10722427 TI - Antimicrobial susceptibility and frequency of occurrence of clinical blood isolates in Europe from the SENTRY antimicrobial surveillance program, 1997 and 1998. AB - As part of the European arm of the SENTRY Antimicrobial Surveillance Program, 25 European university hospitals referred 9613 blood isolates for in vitro testing against >20 antimicrobial agents. Escherichia coli, Staphylococcus aureus, coagulase-negative Staphylococcus, Pseudomonas aeruginosa, and Klebsiella pneumoniae were the 5 most frequent isolates and accounted for two-thirds of all referrals, with minor regional variation. Of these, approximately 0.36% of E. coli and 16.7% of K. pneumoniae isolates proved to be potential extended-spectrum beta-lactamase producers, and their incidence clearly varied regionally. Quinolone resistance was detected among gram-negative species; in particular, P. aeruginosa and Acinetobacter species. Considerable regional variation was observed in the incidences of methicillin resistance in S. aureus and penicillin resistance in Streptococcus pneumoniae. The incidence of vancomycin resistance in enterococci was relatively low overall and primarily associated with Enterococcus faecium. However, extrapolation of these data to smaller and nonteaching hospitals should be undertaken with caution, since resistance rates may be lower in these facilities. PMID- 10722428 TI - Haemophilus influenzae pneumonia in human immunodeficiency virus-infected patients. The Grupo Andaluz para el Estudio de las Enfermedades Infecciosas. AB - Although Haemophilus influenzae is a common etiologic agent of pneumonia in patients infected with human immunodeficiency virus (HIV), the characteristics of this pneumonia have not been adequately assessed. We have prospectively studied features of H. influenzae pneumonia in 26 consecutive HIV-infected inpatients. Most of these patients were severely immunosuppressed; 73.1% had a CD4+ cell count <100/microL. A subacute clinical presentation was observed in 27% of the patients and was associated with a higher degree of immunosuppression (P=.04). Bilateral lung infiltrates were noted radiographically in 57.7% of the cases. The mortality attributable to H. influenzae pneumonia was 11.5%. Thus, pneumonia caused by H. influenzae affects mainly patients with advanced HIV disease, and since its clinical and radiological features may be diverse, this etiology should be considered when pneumonia occurs in patients with advanced HIV infection. The mortality rate associated with H. influenzae pneumonia is not higher than that occurring in the general population. PMID- 10722429 TI - Association between resistance to vancomycin and death in cases of Enterococcus faecium bacteremia. AB - We conducted a retrospective cohort study to determine the association between resistance to vancomycin and mortality among hospitalized patients with Enterococcus faecium bacteremia. We compared outcomes for patients infected with vancomycin-resistant versus vancomycin-susceptible E. faecium among 69 patients with bacteremia defined according to the National Nosocomial Infections Surveillance system. The univariate odds ratio (OR) for death associated with vancomycin resistance was 2.1 (P=.172). After controlling for severity of illness, we found that vancomycin resistance was not associated with mortality (OR, 1.74; 95% confidence interval, 0.5-6.12; P=.39). Vancomycin resistance does not independently increase mortality among patients with E. faecium bacteremia. PMID- 10722430 TI - Epidemiology of ciprofloxacin resistance and its relationship to extended spectrum beta-lactamase production in Klebsiella pneumoniae isolates causing bacteremia. AB - A prospective study of Klebsiella pneumoniae bacteremia was performed in 12 hospitals in 7 countries. Of 452 episodes of bacteremia, 25 (5.5%) were caused by K. pneumoniae that was resistant in vitro to ciprofloxacin. Extended-spectrum beta-lactamase (ESBL) production was detected in 15 (60%) of 25 ciprofloxacin resistant isolates, compared with 68 (16%) of 427 ciprofloxacin-susceptible strains (P=.0001). Multivariate analysis revealed that risk factors for ciprofloxacin resistance in K. pneumoniae included prior receipt of a quinolone (P=.0065) and an ESBL-producing strain (P=.012). In all, 18% of ESBL-producing isolates were also ciprofloxacin-resistant. Pulsed-field gel electrophoresis showed that 11 of the 15 ciprofloxacin-resistant ESBL-producing strains belonged to just 4 genotypes, suggesting that patient-to-patient transmission of such strains occurred. The close relationship between ESBL production and ciprofloxacin resistance is particularly worrisome because the first reported instance of plasmid-mediated ciprofloxacin resistance has been in an isolate of K. pneumoniae also possessing an ESBL. PMID- 10722431 TI - Posterior uveitis in patients with positive serology for syphilis. AB - The clinical features and ophthalmologic findings of 20 patients with syphilitic posterior uveitis seen at the Detroit Medical Center from November 1993 through February 1996 were reviewed. The mean age was 58 years; 8 patients were male and 12 were female; and all patients were black. Three of 9 patients tested were HIV positive. Patients were divided into 2 groups: those with acute (8) and those with chronic (12) syphilitic posterior uveitis. Chorioretinitis was the predominant uveitic pattern (15/20). Eighteen patients presented with blurred vision. All patients had reactive serum fluorescent treponemal antibody, absorbed (FTA-ABS); 3 had nonreactive rapid plasma reagin (RPR). Mean RPR titer in the chronic uveitis group and in the acute uveitis group was 1:27.3 and 1:209.8, respectively. Seven patients had abnormal cerebrospinal fluid (CSF); CSF VDRL was reactive in 2 patients. All patients were treated with intravenous penicillin G. Eight of 14 patients seen at follow-up showed improvement of ophthalmologic findings. Syphilis should be considered in the differential diagnosis of posterior uveitis. PMID- 10722433 TI - Stereotactic biopsy of cerebral lesions in AIDS. AB - Stereotactic brain biopsy was used to establish diagnoses of conditions in patients with AIDS. Two hundred fifty stereotactic biopsies and one open resection were performed for 243 patients. Pathologically abnormal tissue was obtained in 246 (98%) of the procedures, and 16 patients (6%) had >1 diagnosis. Diagnoses included lymphoma in 82 (33%), progressive multifocal leukoencephalopathy in 73 (30%), and tumors not ordinarily associated with AIDS in 7 (3%). In one-third of the cases, the tissue diagnosis differed from the predicted diagnosis. Four of the first 32 patients (12%) developed intracranial bleeding hours after surgery, which was fatal in 3 (9%). Subsequently, all patients were treated with a coagulopathy protocol that included preoperative and postoperative administration of coagulation factors, and there were no further instances of delayed bleeding in the 218 subsequent patients. Among those later patients, there were 7 complications (3%), leading to 4 deaths (2%), a complication rate that compares favorably with that among patients without AIDS. PMID- 10722432 TI - Primary human immunodeficiency virus type 1 infection: clinical manifestations among women in Mombasa, Kenya. AB - The occurrence of clinical manifestations associated with primary human immunodeficiency virus type 1 (HIV-1) infection was evaluated in a prospective cohort study of female sex workers in Mombasa, Kenya. Among 103 women who seroconverted to HIV-1, fever, vomiting, diarrhea, headache, arthralgia, myalgia, skin rash, swollen lymph nodes, extrainguinal lymphadenopathy, inguinal lymphadenopathy, and vaginal candidiasis were noted significantly more frequently at visits in which seroconversion first became evident. Eighty-one percent of seroconverting women had >/=1 of these 11 symptoms or signs. Among 44% of the women, the acute illness was severe enough to prevent them from working. Having >/=2 of 6 selected symptoms and signs yielded a sensitivity of 51%, specificity of 83%, positive likelihood ratio of 3.2, and negative likelihood ratio of 0.5 for acute HIV-1 infection. The recognition of primary HIV-1-infection illness in high-risk populations and subsequent risk-reduction counseling could potentially reduce secondary HIV-1 transmission during this highly infectious period. PMID- 10722434 TI - Raynaud's phenomenon as a manifestation of parvovirus B19 infection: case reports and review of parvovirus B19 rheumatic and vasculitic syndromes. AB - Infection with human parvovirus B19 is manifested as erythema infectiosum, transient aplastic crisis, or hydrops fetalis. Rheumatic manifestations include arthropathy and various vasculitic syndromes. Isolated Raynaud's phenomenon due to parvovirus B19 has never been described. We report on 2 previously healthy sisters with new-onset Raynaud's phenomenon accompanied by severe generalized polyarthralgia. A full workup was negative, except serology for parvovirus B19, which was positive. All symptoms gradually subsided within 3-5 months, and no recurrence has been noted during the 3 years since onset. We review all the studies in the English-language literature on parvovirus B19-induced rheumatic and vasculitic syndromes. We hypothesize that the pathogenesis of Raynaud's phenomenon in our patients involved immune-mediated endothelial damage leading to platelet activation and vasoconstriction. We recommend that in cases of unexplained Raynaud's phenomenon, serology for parvovirus B19 be included in the evaluation. PMID- 10722436 TI - Recurrence of Mycobacterium avium infection in patients receiving highly active antiretroviral therapy and antimycobacterial agents. AB - The known effects of highly active antiretroviral therapy (HAART) on opportunistic infections (OIs) range from immune restoration disease to remission of specific OIs. In the present study, Mycobacterium avium complex infection recurred in 3 patients receiving antimycobacterial therapy and HAART. At the time of the initial M. avium infection, the mean CD4 cell count was 22.3 cells/mm3, and the HIV viral load was 181,133 copies/mL. Relapse occurred a mean of 14. 3 months after the first episode; the mean follow-up CD4 cell count was 89/mm3 (mean elevation of 66 cells/mm3), and the HIV viral load was <400 copies/mL in each patient. M. avium was isolated from blood (1 patient), blood and lymph node (1), and small-bowel tissue (1). M. avium infection may recur as a generalized or focal disease in those who are receiving antimycobacterial agents but whose HAART associated CD4 cell recovery, although significant, is not optimal. PMID- 10722435 TI - Reversal of human immunodeficiency virus type 1-associated hematosuppression by effective antiretroviral therapy. AB - The immunodeficiency of human immunodeficiency virus type 1 (HIV-1) disease may be due to accelerated destruction of mature CD4+ T cells and/or impaired differentiation of progenitors of CD4+ T cells. HIV-1 infection may also inhibit the production of other hematopoietic lineages, by directly or indirectly suppressing the maturation of multilineage and/or lineage-restricted hematopoietic progenitor cells. To test this hypothesis, the effects of durable viral suppression on multilineage hematopoiesis in 66 HIV-1-seropositive patients were evaluated. Administration of effective antiretroviral therapy resulted in an increase in circulating CD4+ T cell counts and statistically significant increases in circulating levels of other hematopoietic lineages, including total white blood cells, lymphocytes, polymorphonuclear leukocytes, and platelets. These results suggest that a significant lesion in untreated HIV-1 disease may lie at the level of cell production from hematopoietic progenitors. PMID- 10722437 TI - Emergence of antimicrobial-resistant shigellosis in Oregon. AB - Ampicillin and trimethoprim-sulfamethoxazole (TMP-SMZ) are currently considered acceptable empirical therapy for shigellosis in developed countries. However, there are few recently reported studies on antimicrobial resistance among shigellae isolated in the United States. We examined the epidemiology of shigellosis and the antimicrobial susceptibility of Shigella species isolated in Oregon from July 1995 through June 1998. Of 430 isolates, 410 were identified to the species level: Shigella sonnei accounted for 55% of isolates, and Shigella flexneri, for 40%. The overall annual incidence of shigellosis was 4.4 cases per 100,000 population. Children aged <5 years (annual incidence, 19.6 cases per 100,000 population) and Hispanics (annual incidence, 28.4 cases per 100,000 population) were at highest risk. Of 369 isolates tested, 59% were resistant to TMP-SMZ, 63% were resistant to ampicillin, 1% were resistant to cefixime, and 0.3% were resistant to nalidixic acid; none of the isolates were resistant to ciprofloxacin. Thirteen percent of the isolates had multidrug resistance to ampicillin, chloramphenicol, streptomycin, sulfisoxazole, and tetracycline. Infections due to multidrug-resistant shigellae are endemic in Oregon. Neither ampicillin nor TMP-SMZ should be considered appropriate empirical therapy for shigellosis any longer; when antibiotics are indicated, a quinolone or cefixime should be used. PMID- 10722439 TI - High prevalence of varicella-zoster virus reactivation in herpes simplex virus seronegative patients with acute peripheral facial palsy. AB - Varicella-zoster virus (VZV) and herpes simplex virus (HSV) are considered to be the major causes of acute peripheral facial palsy (APFP). One hundred and forty two patients with APFP were analyzed by serological assays and polymerase chain reaction analysis. Ramsay Hunt syndrome was diagnosed in 21 patients. Of the remaining 121 patients clinically diagnosed with Bell's palsy, VZV reactivation without zoster (zoster sine herpete) was detected in 35 patients (29%). The prevalence of antibodies to HSV among patients with Bell's palsy was significantly higher than the prevalence among those with VZV reactivation (Ramsay Hunt syndrome or zoster sine herpete). In contrast, a high incidence (88%) of VZV reactivation among HSV-seronegative patients with APFP was observed. Our data indicate that VZV is one of the major etiologic agents of clinically diagnosed Bell's palsy and that VZV reactivation causes APFP in most patients who lack antibodies to HSV. PMID- 10722438 TI - Impact of penicillin susceptibility on medical outcomes for adult patients with bacteremic pneumococcal pneumonia. AB - The impact of penicillin susceptibility on medical outcomes for adult patients with bacteremic pneumococcal pneumonia was evaluated in a retrospective cohort study conducted during population-based surveillance for invasive pneumococcal disease in the greater Atlanta region during 1994. Of the 192 study patients, 44 (23%) were infected with pneumococcal strains that demonstrated some degree of penicillin nonsusceptibility. Compared with patients infected with penicillin susceptible pneumococcal strains, patients whose isolates were nonsusceptible had a significantly greater risk of in-hospital death due to pneumonia (relative risk [RR], 2.1; 95% confidence interval [CI], 1-4.3) and suppurative complications of infection (RR, 4.5; 95% CI, 1-19.3), although only risk of suppurative complications remained statistically significant after adjustment for baseline differences in severity of illness. Among adults with bacteremic pneumococcal pneumonia, infection with penicillin-nonsusceptible pneumococci is associated with an increased risk of adverse outcome. PMID- 10722440 TI - Prolonged afebrile nonproductive cough illnesses in American soldiers in Korea: a serological search for causation. AB - A serological study was undertaken to investigate infections in active-duty United States soldiers with illnesses characterized by prolonged, afebrile, nonproductive coughs. Fifty-four soldiers were enrolled with such illness of >/=2 weeks' duration (case patients) along with 55 well soldiers (control subjects). Serum samples were tested for IgG and IgA antibody to 3 Bordetella pertussis antigens, pertussis agglutinins, IgM antibodies to Mycoplasma pneumoniae, IgM and IgG antibodies to Chlamydia pneumoniae, and IgM antibody to adenoviruses. Forty six case patients (85%) had evidence of recent infection with Bordetella species, M. pneumoniae, or C. pneumoniae, and many had evidence of mixed infections; there were 27 Bordetella species, 20 C. pneumoniae, and 33 M. pneumoniae recent infections. Fifteen case patients had high titers of IgG or IgA to B. pertussis filamentous hemagglutinin without high titers of antibodies to other B. pertussis antigens, which suggested the presence of cross-reacting antibodies to M. pneumoniae and perhaps C. pneumoniae or unidentified infectious agent or agents. Since illnesses due to Bordetella species, M. pneumoniae, and C. pneumoniae can all be treated with macrolide antibiotics and B. pertussis illness can be prevented by immunization, and since military readiness was affected in 63% of the cases, it seems important to conduct further studies in military populations. PMID- 10722441 TI - A pilot study evaluating ceftriaxone and penicillin G as treatment agents for neurosyphilis in human immunodeficiency virus-infected individuals. AB - To compare intravenous (iv) ceftriaxone and penicillin G as therapy for neurosyphilis, blood and CSF were collected before and 14-26 weeks after therapy from 30 subjects infected with human immunodeficiency virus (HIV)-1 who had (1) rapid plasma reagin (RPR) test titers >/=1&rcolon;16, (2) reactive serum treponemal tests, and (3) either reactive CSF-Venereal Disease Research Laboratory (VDRL) tests or CSF abnormalities: (a) CSF WBC values >/=20/microL or (b) CSF protein values >/=50 mg/dL. At baseline, more ceftriaxone recipients had skin symptoms and signs (6 [43%] of 14 vs. 1 [6%] of 16; P=.03), and more penicillin recipients had a history of neurosyphilis (7 [44%] of 16 vs. 1 [7%] of 14; P=.04). There was no difference in the proportion of subjects in each group whose CSF measures improved. Significantly more ceftriaxone recipients had a decline in serum RPR titers (8 [80%] of 10 vs. 2 [13%] of 15; P=. 003), even after controlling for baseline RPR titer, skin symptoms and signs, or prior neurosyphilis were controlled for. Differences in the 2 groups limit comparisons between them. However, iv ceftriaxone may be an alternative to penicillin for treatment of HIV-infected patients with neurosyphilis and concomitant early syphilis. PMID- 10722442 TI - A limitation of 2-stage serological testing for Lyme disease: enzyme immunoassay and immunoblot assay are not independent tests. AB - To improve the accuracy of testing for antibody to Borrelia burgdorferi, 2-stage conditional testing has been recommended, in which sera that yield positive or equivocal results in a first-stage test (e.g., an ELISA) are then tested by immunoblot assay. The increased specificity anticipated with sequential testing, however, depends on immunoblot assays and ELISAs being independent tests. To examine whether they are independent, control serum samples were tested with 2 different commercially available IgM ELISAs and with an IgM immunoblot assay kit. The frequency of false-positive IgM immunoblot assays was significantly higher with ELISA-reactive than with ELISA-negative serum samples (P17 and <100 nmol/liter in 25% of the children, 100 to 499 nmol/liter in 14% of the children, and > or =500 nmol/liter in 13% of the children. Young age, attendance at health posts, and absence of parasitemia were factors independently associated with CQ in blood. With increasing concentrations of CQ, the prevalence of P. falciparum infection and parasite densities decreased. However, at concentrations corresponding to those usually attained during regular prophylaxis (> or =500 nmol/liter), 62% of children were still harboring P. falciparum parasites. In contrast, no infection with P. malariae and only one infection with P. ovale were observed in children with CQ concentrations of > or =100 nmol/liter. These data show the high prevalence of subcurative CQ concentrations in Nigerian children and confirm the considerable degree of CQ resistance in that country. Subtherapeutic drug levels are likely to further promote CQ resistance and may impair the development and maintenance of premunition in areas where malaria is endemic. PMID- 10722479 TI - Horizontal transfer of parC and gyrA in fluoroquinolone-resistant clinical isolates of Streptococcus pneumoniae. AB - We have analyzed genetically three clinical isolates (3180, 3870, and 1244) of Streptococcus pneumoniae with high-level ciprofloxacin resistance. Isolates 3180 and 3870 were atypical because of their insolubility in deoxycholate. However, they hybridized specifically with pneumococcal autolysin and pneumolysin gene probes and have typical pneumococcal atpC and atpA gene sequences. Analysis of the complete sequences of the parC and gyrA genes revealed total variations of 8 and 8.7% (isolate 3180) and 7.4 and 3.6% (isolate 3870), respectively, compared to the wild-type strain R6 sequence. The variations observed between the sequences of R6 and isolate 1244 were less than 0.9%. The structure of the gyrA and parC genes from isolates 3180 and 3870 was organized in sequence blocks that show different levels of divergence, suggesting a pattern of recombination. These results are evidence for recombination at the fluoroquinolone target genes in clinical isolates of S. pneumoniae. The genetically related viridans group streptococci could act as a reservoir for fluoroquinolone resistance genes. PMID- 10722480 TI - Antimicrobial activities of mefloquine and a series of related compounds. AB - Mefloquine was found to have bactericidal activity against methicillin- and fluoroquinolone-susceptible and -resistant strains of Staphylococcus aureus and Staphylococcus epidermidis and gentamicin- and vancomycin-resistant strains of Enterococcus faecalis and Enterococcus faecium. The MICs were 16 microg/ml, and the minimal bactericidal concentrations (MBCs) were 16 to 32 microg/ml. These concentrations cannot be achieved in serum. Mefloquine was active at a more achievable concentration against penicillin-susceptible and -resistant Streptococcus pneumoniae, with MICs of 0.2 to 1.5 microg/ml. Mefloquine was not active against gram-negative bacteria and yeasts. In an attempt to find more active derivatives, 400 mefloquine-related compounds were selected from the chemical inventory of The Walter Reed Army Institute of Research. We identified a series of compounds containing a piperidine methanol group attached to pyridine, quinoline, and benzylquinoline ring systems. These had activities similar to that of mefloquine against S. pneumoniae but were far more active against other gram positive bacteria (MICs for staphylococci, 0.8 to 6.3 microg/ml). They had activities similar to that of amphotericin B against Candida spp. and Cryptococcus neoformans. Combinations of the compounds with gentamicin and vancomycin were additive against staphylococci and pneumococci. The MIC and MBC of gentamicin were decreased by four- to eightfold when this drug was combined with limiting dilutions of the compounds. There was no antagonism with other antimicrobial drugs. The compounds were rapidly bactericidal. They appear to act by disrupting cell membranes. Combinations of the compounds with aminoglycoside antibiotics may have potential for therapeutic use. PMID- 10722481 TI - Intracellular metabolism of beta-L-2',3'-dideoxyadenosine: relevance to its limited antiviral activity. AB - The intracellular metabolism of the beta-L- enantiomer of 2', 3'-dideoxyadenosine (beta-L-ddA) was investigated in HepG2 cells, human peripheral blood mononuclear cells (PBMC), and primary cultured human hepatocytes in an effort to understand the metabolic basis of its limited activity on the replication of human immunodeficiency virus and hepatitis B virus. Incubation of cells with 10 microM [2',3',8-(3)H]-beta-L-ddA resulted in an increased intracellular concentration of beta-L-ddA with time, demonstrating that these cells were able to transport beta L-ddA. However, it did not result in the phosphorylation of beta-L-ddA to its pharmacologically active 5'-triphosphate (beta-L-ddATP). Five other intracellular metabolites were detected and identified as beta-L-2', 3'-dideoxyribonolactone, hypoxanthine, inosine, ADP, and ATP, with the last being the predominant metabolite, reaching levels as high as 5.14 +/- 0.95, 8.15 +/- 2.64, and 15.60 +/ 1.74 pmol/10(6) cells at 8, 4, and 2 h in HepG2 cells, PBMC, and hepatocytes, respectively. In addition, a beta-glucuronic derivative of beta-L-ddA was detected in cultured hepatocytes, accounting for 12.5% of the total metabolite pool. Coincubation of hepatocytes in primary culture with beta-L-ddA in the presence of increasing concentrations of 5'-methylthioadenosine resulted in decreased phosphorolysis of beta-L-ddA and formation of associated metabolites. These results indicate that the limited antiviral activity of beta-L-ddA is the result of its inadequate phosphorylation to the nucleotide level due to phosphorolysis and catabolism of beta-L-ddA by methylthioadenosine phosphorylase (EC 2.4.2.28). PMID- 10722482 TI - Broad-spectrum antiviral activity of the IMP dehydrogenase inhibitor VX-497: a comparison with ribavirin and demonstration of antiviral additivity with alpha interferon. AB - The enzyme IMP dehydrogenase (IMPDH) catalyzes an essential step in the de novo biosynthesis of guanine nucleotides, namely, the conversion of IMP to XMP. The major event occurring in cells exposed to competitive IMPDH inhibitors such as ribavirin or uncompetitive inhibitors such as mycophenolic acid (MPA) is a depletion of the intracellular GTP and dGTP pools. Ribavirin is approved as an inhaled antiviral agent for treatment of respiratory syncytial virus (RSV) infection and orally, in combination with alpha interferon (IFN-alpha), for the treatment of chronic hepatitis C virus (HCV) infection. VX-497 is a potent, reversible uncompetitive IMPDH inhibitor which is structurally unrelated to other known IMPDH inhibitors. Studies were performed to compare VX-497 and ribavirin in terms of their cytotoxicities and their efficacies against a variety of viruses. They included DNA viruses (hepatitis B virus [HBV], human cytomegalovirus [HCMV], and herpes simplex virus type 1 [HSV-1]) and RNA viruses (respiratory syncytial virus [RSV], parainfluenza-3 virus, bovine viral diarrhea virus, Venezuelan equine encephalomyelitis virus [VEEV], dengue virus, yellow fever virus, coxsackie B3 virus, encephalomyocarditis virus [EMCV], and influenza A virus). VX 497 was 17- to 186-fold more potent than ribavirin against HBV, HCMV, RSV, HSV-1, parainfluenza-3 virus, EMCV, and VEEV infections in cultured cells. The therapeutic index of VX-497 was significantly better than that of ribavirin for HBV and HCMV (14- and 39-fold, respectively). Finally, the antiviral effect of VX 497 in combination with IFN-alpha was compared to that of ribavirin with IFN alpha in the EMCV replication system. Both VX-497 and ribavirin demonstrated additivity when coapplied with IFN-alpha, with VX-497 again being the more potent in this combination. These data are supportive of the hypothesis that VX-497, like ribavirin, is a broad-spectrum antiviral agent. PMID- 10722483 TI - Pefloxacin-induced achilles tendon toxicity in rodents: biochemical changes in proteoglycan synthesis and oxidative damage to collagen. AB - Despite a relatively low incidence of serious side effects, fluoroquinolones and the fluoroquinolone pefloxacin have been reported to occasionally promote tendinopathy that might result in the complication of spontaneous rupture of tendons. In the present study, we investigated in rodents the intrinsic deleterious effect of pefloxacin (400 mg/kg of body weight) on Achilles tendon proteoglycans and collagen. Proteoglycan synthesis was determined by measurement of in vivo and ex vivo radiosulfate incorporation in mice. Collagen oxidative modifications were measured by carbonyl derivative detection by Western blotting. An experimental model of tendinous ischemia (2 h) and reperfusion (3 days) was achieved in rats. Biphasic changes in proteoglycan synthesis were observed after a single administration of pefloxacin, consisting of an early inhibition followed by a repair-like phase. The depletion phase was accompanied by a marked decrease in the endogenous serum sulfate level and a concomitant increase in the level of sulfate excretion in urine. Studies of ex vivo proteoglycan synthesis confirmed the in vivo results that were obtained. The decrease in proteoglycan anabolism seemed to be a direct effect of pefloxacin on tissue metabolism rather than a consequence of the low concentration of sulfate. Pefloxacin treatment for several days induced oxidative damage of type I collagen, with the alterations being identical to those observed in the experimental tendinous ischemia and reperfusion model. Oxidative damage was prevented by coadministration of N acetylcysteine (150 mg/kg) to the mice. These results provide the first experimental evidence of a pefloxacin-induced oxidative stress in the Achilles tendon that altered proteoglycan anabolism and oxidized collagen. PMID- 10722484 TI - A rapid phenotypic assay for detection of acyclovir-resistant varicella-zoster virus with mutations in the thymidine kinase open reading frame. AB - Susceptibility assays by cell culture methods are time-consuming and are particularly difficult to perform with varicella-zoster virus (VZV). To overcome this limitation, we have adapted a functional test of the viral thymidine kinase (TK) in TK-deficient (tdk mutant) bacteria to detect ACV-resistant VZV in clinical samples. After PCR amplification, the complete viral TK open reading frame (ORF) is purified from PCR primers, digested with two restriction enzymes, and ligated in an oriented fashion into a bacterial expression vector. The ligation products are then used to transform tdk mutant bacteria. After transformation, an aliquot of the bacteria is plated onto a plate with minimal medium containing (i) ampicillin to select for plasmids carrying the viral TK ORF and (ii) isopropyl beta-D-thiogalactopyranoside (IPTG) to induce its expression. An identical aliquot of bacteria is also plated onto a medium containing, in addition to the components described above, 5-fluorodeoxyuridine (FUdR). Compared to the number of transformants on FUdR-free medium, the number of colonies carrying TK derived from susceptible strains was reduced by 86%, on average, in the presence of FUdR. In contrast, the number of transformants carrying TK from resistant strains with a mutant TK were reduced by only 4%, on average, on FUdR containing plates. We have assessed the validity of this assay with cell culture isolates and several clinical samples including two cerebrospinal fluid samples from which no virus could be isolated. This colony reduction assay allowed the correct identification of the TK phenotype of each VZV isolate tested and can be completed within 3 days of receipt of the sample. PMID- 10722485 TI - Comparative pharmacodynamics of gatifloxacin and ciprofloxacin in an in vitro dynamic model: prediction of equiefficient doses and the breakpoints of the area under the curve/MIC ratio. AB - To demonstrate the impact of the pharmacokinetics of gatifloxacin (GA) relative to those of ciprofloxacin (CI) on the antimicrobial effect (AME), the killing and regrowth kinetics of two differentially susceptible clinical isolates each of Staphylococcus aureus, Escherichia coli, and Klebsiella pneumoniae were studied. With each organism, a series of monoexponential pharmacokinetic profiles of GA (half-life [t(1/2)], 7 h) and CI (t(1/2) = 4 h) were simulated to mimic different single doses of GA and two 12-h doses of CI. The respective eightfold ranges of the ratios of the area under the concentration-time curve (AUC) to the MIC were 58 to 466 and 116 to 932 (microg. h/ml)/(microg/ml). The species- and strain independent linear relationships observed between the intensity of AME (I(E)) and log AUC/MIC were not superimposed for GA and CI (r(2) = 0.99 in both cases). The predicted AUC/MIC ratio for GA that might be equivalent to a clinically relevant AUC/MIC breakpoint for CI was estimated to be 102 rather than 125 (microg. h/ml)/(microg/ml). The respective MIC breakpoints were 0.32 microg/ml (for a 400 mg dose of GA) and 0.18 microg/ml (for two 500-mg doses of CI). On the basis of the I(E)-log AUC/MIC relationships, equiefficient 24-h doses (D(24h)s) of GA and CI were calculated for hypothetical strains of S. aureus, E. coli, and K. pneumoniae for which the MICs were equal to the MICs at which 50% of isolates are inhibited. To provide an "acceptable" I(E) equal to 200 (log CFU/ml). h, i.e., the I(E) provided by AUC/MIC of 125 (microg. h/ml)/(microg/ml) for ciprofloxacin, the D(24h)s of GA for all three organisms were much lower (115, 30, and 60 mg) than the clinically proposed 400-mg dose. Although the usual dose of CI (two doses of 500 mg) would be in excess for E. coli and K. pneumoniae (D(24h) = two doses of 40 mg and two doses of 115 mg, respectively), even the highest clinical dose of CI (two doses of 750 mg) might be insufficient for S. aureus (D(24h), > two doses of 1,000 mg). The method of generalization of data obtained with specific organisms to other representatives of the same species described in the present report might be useful for prediction of the AMEs of new quinolones. PMID- 10722486 TI - Efficacies of imipenem, meropenem, cefepime, and ceftazidime in rats with experimental pneumonia due to a carbapenem-hydrolyzing beta-lactamase-producing strain of Enterobacter cloacae. AB - The antibacterial activities of imipenem-cilastatin, meropenem-cilastatin, cefepime and ceftazidime against Enterobacter cloacae NOR-1, which produces the carbapenem-hydrolyzing beta-lactamase NmcA and a cephalosporinase, and against one of its in vitro-obtained ceftazidime-resistant mutant were compared by using an experimental model of pneumonia with immunocompetent rats. The MICs of the beta-lactams with an inoculum of 5 log(10) CFU/ml were as follows for E. cloacae NOR-1 and its ceftazidime-resistant mutant, respectively: imipenem, 16 and 128 microg/ml, meropenem, 4 and 32 microg/ml, cefepime, <0.03 and 1 microg/ml, and ceftazidime, 1 and 512 microg/ml. The chromosomally located cephalosporinase and carbapenem-hydrolyzing beta-lactamase NmcA were inducible by cefoxitin and meropenem in E. cloacae NOR-1, and both were stably overproduced in the ceftazidime-resistant mutant. Renal impairment was induced (uranyl nitrate, 1 mg/kg of body weight) in rats to simulate the human pharmacokinetic parameters for the beta-lactams studied. Animals were intratracheally inoculated with 8.5 log(10) CFU of E. cloacae, and therapy was initiated 3 h later. At that time, animal lungs showed bilateral pneumonia containing more than 6 log(10) CFU of E. cloacae per g of tissue. Despite the relative low MIC of meropenem for E. cloacae NOR-1, the carbapenem-treated rats had no decrease in bacterial counts in their lungs 60 h after therapy onset compared to the counts for the controls, regardless of whether E. cloacae NOR-1 or its ceftazidime-resistant mutant was inoculated. A significant decrease in bacterial titers was observed for the ceftazidime-treated rats infected with E. cloacae NOR-1 only. Cefepime was the only beta-lactam tested effective as treatment against infections due to E. cloacae NOR-1 or its ceftazidime-resistant mutant. PMID- 10722487 TI - Characterization of VIM-2, a carbapenem-hydrolyzing metallo-beta-lactamase and its plasmid- and integron-borne gene from a Pseudomonas aeruginosa clinical isolate in France. AB - Pseudomonas aeruginosa COL-1 was identified in a blood culture of a 39-year-old woman treated with imipenem in Marseilles, France, in 1996. This strain was resistant to beta-lactams, including ureidopenicillins, ticarcillin-clavulanic acid, cefepime, ceftazidime, imipenem, and meropenem, but remained susceptible to the monobactam aztreonam. The carbapenem-hydrolyzing beta-lactamase gene of P. aeruginosa COL-1 was cloned, sequenced, and expressed in Escherichia coli DH10B. The deduced 266-amino-acid protein was an Ambler class B beta-lactamase, with amino acid identities of 32% with B-II from Bacillus cereus; 31% with IMP-1 from several gram-negative rods in Japan, including P. aeruginosa; 27% with CcrA from Bacteroides fragilis; 24% with BlaB from Chryseobacterium meningosepticum; 24% with IND-1 from Chryseobacterium indologenes; 21% with CphA-1 from Aeromonas hydrophila; and 11% with L-1 from Stenotrophomonas maltophilia. It was most closely related to VIM-1 beta-lactamase recently reported from Italian P. aeruginosa clinical isolates (90% amino acid identity). Purified VIM-2 beta lactamase had a pI of 5.6, a relative molecular mass of 29.7 kDa, and a broad substrate hydrolysis range, including penicillins, cephalosporins, cephamycins, oxacephamycins, and carbapenems, but not monobactams. As a metallo-beta lactamase, its activity was zinc dependent and inhibited by EDTA (50% inhibitory concentration, 50 microM). VIM-2 conferred a resistance pattern to beta-lactams in E. coli DH10B that paralleled its in vitro hydrolytic properties, except for susceptibility to ureidopenicillins, carbapenems, and cefepime. bla(VIM-2) was located on a ca. 45-kb plasmid that in addition conferred resistance to sulfamides and that was not self-transmissible either from P. aeruginosa to E. coli or from E. coli to E. coli. bla(VIM-2) was the only gene cassette located within the variable region of a novel class 1 integron, In56, that was weakly related to the bla(VIM-1)-containing integron. VIM-2 is the second carbapenem hydrolyzing metalloenzyme characterized from a P. aeruginosa isolate outside Japan. PMID- 10722489 TI - Identification of a streptogramin A acetyltransferase gene in the chromosome of Yersinia enterocolitica. AB - Streptogramins are polypeptide antibiotics inhibiting protein synthesis by the prokaryotic ribosome. Gram-positive organisms are susceptible to streptogramins, while most gram-negative bacteria are intrinsically resistant. We have found a genomic fragment from a Yersinia enterocolitica isolate with an open reading frame coding for a polypeptide similar to the virginiamycin acetyltransferases found in various plasmids from gram-positive bacteria. The susceptible Escherichia coli strain DB10 was transformed to resistance to the type A streptogramins and to mixed (A + B) streptogramins upon introduction of a plasmid containing that gene. In addition, we showed streptogramin acetylating activity in vitro dependent on the presence of the Y. enterocolitica sat gene. Southern blot hybridization experiments showed that the sat gene was present in all the Y. enterocolitica isolates examined. These data together show that the gene in the Y. enterocolitica chromosome encoded an active streptogramin acetyltransferase. The deduced sequence of the Y. enterocolitica Sat protein was close to those of sat gene products found in gram-positive bacteria and cyanobacteria, suggesting a common evolutionary origin. PMID- 10722490 TI - Interspecies comparison of pharmacokinetics of the novel triazole antifungal agent SYN-2869 and its derivatives. AB - The pharmacokinetics and distribution in tissue of several novel triazole antifungal agents were studied in different animal species in order to select an appropriate lead compound. The purpose of the study was also to determine species differences in pharmacokinetics for SYN azoles to select the most appropriate species for secondary efficacy and toxicological evaluation of the selected compound. SYN-2836, SYN-2869, SYN-2903, and SYN-2921 were rapidly absorbed into the systemic circulation and reached maximum concentrations (C(max)s) of 7.31 +/- 2.53, 6.29 +/- 0.85, 6.16 +/- 0.39, and 3.41 +/- 0.34 microg/ml, respectively, in BALB/c mice after administration of an oral dose of 50 mg/kg of body weight, with bioavailability being greater than 45% in all mice. The areas under the concentration-time curve from time zero to infinity (AUC(0-infinity)s) after administration of a single intravenous dose of 20 mg/kg to mice varied between 25.0 and 63.6 microg. h/ml. The half-life was in the range of 4.5 to 6 h. In Sprague-Dawley rats there was no significant difference in AUC(0-infinity) after administration of a single intravenous dose of 20 mg/kg, but on oral administration, the bioavailability of SYN-2836 was extremely low, while that of SYN-2869 was only 14.7%. In New Zealand White rabbits the C(max) and the time to reach C(max) for SYN-2836 and SYN-2869 after administration of a single oral dose of 50 mg/kg were similar. There were significant differences in AUC(0-infinity) and half-life between SYN-2836 and SYN-2869. On the other hand, in beagle dogs the C(max) and AUC(0-infinity) of SYN-2836 after administration of a single oral dose of 30 mg/kg were 4.82 +/- 1.54 microg/ml and 41.8 +/- 15.7 microg. h/ml, respectively, which were threefold higher than those of SYN-2869. The concentrations of the SYN compounds in tissue indicated that the AUC(0-infinity)s of SYN-2836, SYN-2869, SYN-2903, and SYN-2921 in mouse lungs were significantly different from each other. The ratios of the concentrations of the SYN azoles in lungs to those in plasma were also significantly different from those for itraconazole. Among the SYN azoles the highest concentration in the lungs was found for SYN-2869. The higher level of distribution of SYN-2869 into lung tissue was considered to contribute to the potent efficacy in respiratory tract infection models compared with the potency of itraconazole. Significant differences in the pharmacokinetics of these compounds were observed in different animal species, and selection of an animal model for further evaluation was based on results obtained from these studies. PMID- 10722488 TI - Comparative pharmacokinetics, tissue distributions, and effects on renal function of novel polymeric formulations of amphotericin B and amphotericin B-deoxycholate in rats. AB - The pharmacokinetic profiles of a traditional formulation of amphotericin B (Fungizone) and novel nanosphere and mixed micelle delivery systems developed for amphotericin B were compared and described. Six groups of male Wistar rats received intravenous injections of the different formulations. Plasma and tissue samples were obtained at 11 different times after dosing, with three animals used each time. The amphotericin B concentrations in plasma and tissues were analyzed by high-performance liquid chromatography. The plasma drug concentration-time profiles were best described by a two-compartment model. Models that described the observed single or double peak disposition kinetics in kidney, liver, and spleen were also developed. Parameter estimates from those models show that components of the formulation such as poloxamer 188, which is present in all new formulations, seem to play an important role in the rate of drug uptake by the tissues; in general, the levels of amphotericin B in tissues were increased after the administration of the new formulations compared with those after the administration of Fungizone. The increment in the baseline plasma creatinine level was used as an index of renal function. All formulations increased this baseline value, but the novel formulations exhibited fewer renal effects than Fungizone did. However, a direct relationship between drug exposure in the kidneys and development of renal damage could not be found. PMID- 10722491 TI - Pharmacokinetics of an everninomicin (SCH 27899) in mice, rats, rabbits, and cynomolgus monkeys following intravenous administration. AB - The pharmacokinetics of SCH 27899, a novel oligosaccharide compound of the everninomicin class with excellent activity against gram-positive strains, was studied with mice, rats, rabbits, and cynomolgus monkeys following intravenous administration as SCH 27899-N-methylglucamine-hydroxypropyl beta-cyclodextrin. Concentrations of SCH 27899 in mouse serum, rat plasma, and rabbit serum were determined by a high-pressure liquid chromatography method on a poly(styrene divinyl benzene) column, and those in monkey plasma were determined by a paired ion chromatographic method. Plasma and serum concentrations of SCH 27899 exhibited a biexponential decline in all species following intravenous administration. The half-lives at beta phase were 3.0 to 7.9 h in mice, rats, and rabbits and 24 h in cynomolgus monkeys. There was a linear relationship between the area under the curve extrapolated to infinity [AUC(I)] in mice and dose. Rabbits also exhibited dose proportionality in AUC(I). However, in rats, increasing the dose from 3 to 60 mg/kg of body weight resulted in a 49-fold increase in AUC(I). When the species was changed from mouse to rat, rabbit, or cynomolgus monkey, AUC(I) increased, whereas clearance (CL) decreased. It was concluded that the pharmacokinetics of SCH 27899 in animals varied with species; CL was the highest in mice and rats, followed by rabbits and cynomolgus monkeys. PMID- 10722492 TI - A novel phenotypic drug susceptibility assay for human immunodeficiency virus type 1. AB - Although combination antiretroviral therapy has resulted in a considerable improvement in the treatment of human immunodeficiency virus (HIV) type 1 (HIV-1) infection, the emergence of resistant virus is a significant obstacle to the effective management of HIV infection and AIDS. We have developed a novel phenotypic drug susceptibility assay that may be useful in guiding therapy and improving long-term suppression of HIV replication. Susceptibility to protease (PR) and reverse transcriptase (RT) inhibitors is measured by using resistance test vectors (RTVs) that contain a luciferase indicator gene and PR and RT sequences derived from HIV-1 in patient plasma. Cells are transfected with RTV DNA, resulting in the production of virus particles that are used to infect target cells. Since RTVs are replication defective, luciferase activity is measured following a single round of replication. The assay has been automated to increase throughput and is completed in 8 to 10 days. Test results may be useful in facilitating the selection of optimal treatment regimens for patients who have failed prior therapy or drug-naive patients infected with drug-resistant virus. In addition, the assay can be used to evaluate candidate drugs and assist in the development of new drugs that are active against resistant strains of HIV-1. PMID- 10722493 TI - Antiviral properties of a series of 1,6-naphthyridine and 7, 8 dihydroisoquinoline derivatives exhibiting potent activity against human cytomegalovirus. AB - A series of 1,6-naphthyridine (L. Chan, H. Jin, T. Stefanac, J. F. Lavallee, G. Falardeau, W. Wang, J. Bedard, S. May, and L. Yuen, J. Med. Chem. 42:3023-3025, 1999) and isoquinoline (L. Chan, H. Jin, T. Stefanac, W. Wang, J. F. Lavallee, J. Bedard, and S. May, Bioorg. Med. Chem. Lett. 9:2583-2586, 1999) analogues exhibiting a high level of anti-human cytomegalovirus (HCMV) activity were investigated in a series of studies aimed at better understanding the mechanism of action of some representatives of this class of compounds. In vitro antiviral profiling revealed that these compounds were active against a narrow spectrum of viruses, essentially the human herpesviruses and type 2 rhinovirus. In HCMV assays, a 39- to 223-fold lower 50% inhibitory concentration was obtained for compound A1 than for ganciclovir against strains AD 169 and Towne. In addition, ganciclovir, foscarnet, cidofovir, and BDCRB (2-bromo-5,6-dichloro-1-beta-D ribofuranosylbenzimidazole)-resistant HCMV strains remained susceptible to 1,6 naphthyridines and 7, 8-dihydroisoquinolines tested in this study, supporting the view that a novel mechanism of action could be involved. Drug combination studies showed a small but significant synergistic antiviral effect between compound B2 and ganciclovir. Cytotoxicity profiling of representative compounds under various cell growth conditions indicated a generally similar cytotoxic effect, relative to ganciclovir, in log-phase growing cells. However, in stationary cells, a relatively higher level of toxicity was observed than that for control compound. Effect of time of drug addition showed that the anti-HCMV activity of compound A1, ganciclovir, and cidofovir was lost at approximately the same time (72 h postinfection), indicating that the compound was affecting events at the early and late stage of virus replication. This interpretation is also supported by reduction of de novo synthesis of pp65 tegument protein and lack of any effect of the compound on viral adsorption. A reduction of the HCMV enhancer-promoter directed luciferase expression was also observed in a stably transfected cell line when compound A1 was present at relatively high concentrations. PMID- 10722494 TI - In vivo characterization of the pharmacodynamics of flucytosine in a neutropenic murine disseminated candidiasis model. AB - In vivo pharmacodynamic parameters have been characterized for a variety of antibacterial agents. These parameters have been studied in correlation with in vivo outcomes in order to determine (i) which dosing parameter is predictive of outcome and (ii) the magnitude of that parameter associated with efficacy. Very little is known of the pharmacodynamics of antifungal agents. We used a neutropenic murine model of disseminated candidiasis to correlate the pharmacodynamic parameters (percentage of time above the MIC, area under the concentration-time curve [AUC]/MIC and peak level/MIC) for flucytosine (5-FC) in vivo with efficacy as measured by organism number in homogenized kidney cultures after 24 h of therapy. The pharmacokinetics of 5-FC in infected mice were linear. Serum half-lives ranged from 0.36 to 0.43 h. Infection was achieved by intravenous inoculation of 10(6) CFU of yeast cells per ml via the lateral tail vein of neutropenic mice. Groups of mice were treated with fourfold escalating total doses of 5-FC ranging from 1.56 to 400 mg/kg of body weight/day divided into one, two, four, or eight doses over 24 h. Increasing doses produced minimal concentration-dependent killing ranging from 0 to 0.9 log(10) CFU/kidneys. 5-FC did, however, produce a dose-dependent suppression of growth after levels in serum had fallen below the MIC. The fungistatic dose increased from 6 to 8 mg/kg with dosing every 3 and 6 h to 70 mg/kg at with dosing every 24 h. Nonlinear regression analysis was used to determine which pharmacodynamic parameter best correlated with efficacy. Time above the MIC was the parameter best predictive of outcome, while AUC/MIC was only slightly less predictive (time above MIC, R(2) = 85%; AUC/MIC, R(2) = 77%; peak level/MIC, R(2) = 53%). Maximal efficacy was observed when levels exceeded the MIC for only 20 to 25% of the dosing interval. If one considers drug kinetics in humans, these results suggest reevaluation of current dosing regimens. PMID- 10722495 TI - In vivo pharmacodynamic activities of two glycylcyclines (GAR-936 and WAY 152,288) against various gram-positive and gram-negative bacteria. AB - The in vivo pharmacodynamic activities of two glycylcyclines (GAR-936 and WAY 152,288) were assessed in an experimental murine thigh infection model in neutropenic mice. Mice were infected with one of several strains of Streptococcus pneumoniae, Staphylococcus aureus, Escherichia coli, or Klebsiella pneumoniae. Most infections were treated with a twice-daily dosing schedule, with administration of 0.75 to 192 mg of GAR-936 or WAY 152,288 per kg of body weight. A maximum-effect dose-response model was used to calculate the dose that produced a net bacteriostatic effect over 24 h of therapy. This dose was called the bacteriostatic dose. More extensive dosing studies were performed with S. pneumoniae 1199, E. coli ATCC 25922, and K. pneumoniae ATCC 43816, with doses being given as one, two, four, or eight equal doses over a period of 24 h. The dosing schedules were designed in order to minimize the interrelationship between the various pharmacokinetic and pharmacodynamic parameters studied. These parameters were time above 0.03 to 32 times the MIC, area under the concentration time curve (AUC), and maximum concentration of drug in serum (C(max)). The bacteriostatic dose remained essentially the same, irrespective of the dosing frequency, for S. pneumoniae 1199 (0.3 to 0.9 mg/kg/day). For E. coli ATCC 25922 and K. pneumoniae ATCC 43816, however, more frequent dosing led to lower bacteriostatic doses. Pharmacokinetic studies demonstrated dose-dependent elimination half-lives of 1.05 to 2.34 and 1.65 to 3.36 h and serum protein bindings of 59 and 71% for GAR-936 and WAY 152,288, respectively. GAR-936 and WAY 152,288 were similarly effective against the microorganisms studied, with small differences in maximum effect and 50% effective dose. The glycylcyclines were also similarly effective against tetracycline-sensitive and tetracycline resistant bacteria. Time above a certain factor (range, 0.5 to 4 times) of the MIC was a better predictor of in vivo efficacy than C(max) or AUC for most organism-drug combinations. The results demonstrate that in order to achieve 80% maximum efficacy, the concentration of unbound drug in serum should be maintained above the MIC for at least 50% of the time for GAR-936 and for at least 75% of the time for WAY 152,288. The results of these experiments will aid in the rational design of dose-finding studies for these glycylcyclines in humans. PMID- 10722496 TI - Compartmental pharmacokinetics and tissue distribution of multilamellar liposomal nystatin in rabbits. AB - The plasma pharmacokinetics of multilamellar liposomal nystatin were studied in normal, catheterized rabbits after single and multiple daily intravenous administration of dosages of 2, 4, and 6 mg/kg of body weight, and drug levels in tissues were assessed after multiple dosing. Concentrations of liposomal nystatin were measured as those of nystatin by a validated high-performance liquid chromatography method, and plasma concentration data were fitted into a two compartment open model. Across the investigated dosage range, liposomal nystatin demonstrated nonlinear kinetics with more than proportional increases in the AUC(0-24) and decreasing clearance, consistent with dose-dependent tissue distribution and/or a dose-dependent elimination process. After single-dose administration, the mean C(max) increased from 13.07 microg/ml at 2 mg/kg to 41.91 microg/ml at 6 mg/kg (P < 0.001); the AUC(0-24) changed from 11.65 to 67.44 microg. h/ml (P < 0.001), the V(d) changed from 0.205 to 0. 184 liters/kg (not significant), the CL(t) from 0.173 to 0.101 liters/kg. h (P < 0.05), and terminal half-life from 0.96 to 1.51 h (P < 0.05). There were no significant changes in pharmacokinetic parameters after multiple dosing over 14 days. Assessment of tissue concentrations of nystatin near peak plasma levels after multiple dosing over 15 days revealed preferential distribution to the lungs, liver, and spleen at that time point. Substantial levels were also found in the urine, raising the possibility that renal excretion may play a significant role in drug elimination. Liposomal nystatin administered to rabbits was well tolerated and displayed nonlinear pharmacokinetics, potentially therapeutic peak plasma concentrations, and substantial penetration into tissues. Pharmacokinetic parameters were very similar to those observed in patients, thus validating results derived from infection models in the rabbit and allowing inferences to be made about the treatment of invasive fungal infections in humans. PMID- 10722497 TI - Pentamidine inhibition of group I intron splicing in Candida albicans correlates with growth inhibition. AB - We previously demonstrated that pentamidine, which has been clinically used against Pneumocystis carinii, inhibits in vitro a group I intron ribozyme from that organism. Another fungal pathogen, Candida albicans, also harbors a group I intron ribozyme (Ca.LSU) in the essential rRNA genes in almost half of the clinical isolates analyzed. To determine whether pentamidine inhibits Ca.LSU in vitro and in cells, phylogenetically closely related intron-containing (4-1) and intronless (62-1) strains were studied. Splicing in vitro of the Ca.LSU group I intron ribozyme was completely inhibited by pentamidine at 200 microM. On rich glucose medium, the intron-containing strain was more sensitive to growth inhibition by pentamidine than was the intronless strain, as measured by disk or broth microdilution assays. On rich glycerol medium, they were equally susceptible to pentamidine. At pentamidine levels selectively inhibiting the intron-containing strain (1 microM) in glucose liquid cultures, inhibition of splicing and rRNA maturation was detected by quantitative reverse transcription PCR within 1 min with a 10- to 15-fold accumulation of precursor rRNA. No comparable effect was seen in the intronless strain. These results correlate the cellular splicing inhibition of Ca.LSU with the growth inhibition of strain 4-1 harboring Ca.LSU. Broth microdilution assays of 13 Candida strains showed that intron-containing strains were generally more susceptible to pentamidine than the intronless strains. Our data suggest that ribozymes found in pathogenic microorganisms but absent in mammals may be targets for antimicrobial therapy. PMID- 10722498 TI - Macrolide resistance genes in Enterococcus spp. AB - Seventy-eight isolates of different Enterococcus species (E. faecalis, n = 27; E. faecium, n = 23; E. durans, n = 8; E. avium, n = 6; E. hirae, n = 9; E. gallinarum, n = 3; and E. casseliflavus, n = 2) with a variety of erythromycin resistance phenotypes were examined for the presence of macrolide resistance genes (ermA, ermB, ermC, ermTR, mefA/E, and msrA). Positive PCR amplifications of ermB were obtained for 39 of 40 highly erythromycin-resistant Enterococcus isolates (MICs, >128 microg/ml) of different species; the remaining highly resistant E. faecium isolate was positive for PCR amplification of ermA but was negative for PCR amplification of the ermB and ermC genes. For all enterococcal strains for which erythromycin MICs were < or =32 microg/ml PCRs were negative for erm methylase genes. For all E. faecium isolates PCR amplified products of the expected size of 400 bp were obtained when msrA primers were used, with the results being independent of the erythromycin resistance phenotype. All the other enterococcal species gave negative results by msrA PCRs. Sequencing of the msrA PCR products from either erythromycin-susceptible, low-level-resistant, or highly resistant E. faecium strains showed that the amplicons did not correspond to the msrA gene described for Staphylococcus epidermidis but corresponded to a new putative efflux determinant, which showed 62% identity with the msrA gene at the DNA level and 72% similarity at the amino acid level. This new gene was named msrC. PMID- 10722499 TI - Antimalarial bioavailability and disposition of artesunate in acute falciparum malaria. AB - The pharmacokinetic properties of oral and intravenous artesunate (2 mg/kg of body weight) were studied in 19 adult patients with acute uncomplicated Plasmodium falciparum malaria by using a randomized crossover design. A sensitive bioassay was used to measure the antimalarial activity in plasma which results from artesunate and its principal metabolite, dihydroartemisinin. The oral study was repeated with 15 patients during convalescence. The mean absolute oral bioavailability of the antimalarial agent in patients with acute malaria was 61% (95% confidence interval [CI], 52 to 70%). The absorption and elimination of oral artesunate were rapid, with a mean elimination half-life of antimalarial activity of 43 min (95% CI, 33 to 53 min). Following oral administration to patients with acute falciparum malaria, peak antimalarial activity in plasma and the area under the plasma concentration-time curve were approximately double those during convalescence and the apparent volume of distribution and clearance were approximately half those during convalescence (P < or = 0.005). Acute malaria is associated with a significant reduction in the clearance of artesunate-associated antimalarial activity. PMID- 10722500 TI - Pharmacokinetic interaction between amprenavir and clarithromycin in healthy male volunteers. AB - The P450 enzyme, CYP3A4, extensively metabolizes both amprenavir and clarithromycin. To determine if an interaction exists when these two drugs are coadministered, the pharmacokinetics of amprenavir and clarithromycin were investigated in healthy adult male volunteers. This was a Phase I, open-label, randomized, balanced, multiple-dose, three-period crossover study. Fourteen subjects received the following three regimens: amprenavir, 1,200 mg twice daily over 4 days (seven doses); clarithromycin, 500 mg twice daily over 4 days (seven doses); and the combination of the above regimens over 4 days (seven doses of each drug). Twelve subjects completed all treatments and the follow-up period. The erythromycin breath test (ERMBT) was administered at baseline, 2 h after the final dose of each of the three regimens and at the first follow-up visit. Coadministration of clarithromycin and amprenavir significantly increased the mean amprenavir AUC(ss), C(max,ss), and C(min,ss) by 18, 15, and 39%, respectively. Amprenavir had no significant effect on the AUC(ss) of clarithromycin, but the median T(max,ss)for clarithromycin increased by 2.0 h, renal clearance increased by 34%, and the AUC(ss) for 14-(R) hydroxyclarithromycin decreased by 35% when it was given with amprenavir. Amprenavir and clarithromycin reduced the ERMBT result by 85 and 67%, respectively, and by 87% when the two drugs were coadministered. The baseline ERMBT value did not correlate with clearance of amprenavir or clarithromycin. A pharmacokinetic interaction occurs when amprenavir and clarithromycin are coadministered, but the effects are not likely to be clinically important, and coadministration does not require a dosage adjustment for either drug. PMID- 10722501 TI - Single-dose intrapulmonary pharmacokinetics of rifapentine in normal subjects. AB - The intrapulmonary pharmacokinetics of rifapentine were studied in 30 volunteers who received a single, oral dose of rifapentine (600 mg). Subgroups of five subjects each underwent bronchoscopy and bronchoalveolar lavage (BAL) at timed intervals following drug administration. Drug concentrations, including the concentration of the primary metabolite 25-desacetyl rifapentine, were determined in plasma, BAL fluid, and alveolar cells (AC) by high-pressure liquid chromatography. The concentrations in epithelial lining fluid (ELF) were calculated by the urea diffusion method. The concentration-time data were fit to two-compartment (plasma) or one-compartment (AC and ELF) models. The peak concentrations in plasma, ELF, and AC, 26.2, 3. 7, and 5.3 microg/ml, respectively, occurred at 5, 5, and 7 h after drug administration, respectively. The half-lives and areas under the curve for plasma, ELF, and AC were 18.3 h and 520 microg. h/ml, 20.8 h and 111 microg. h/ml, and 13.0 h and 133 microg. h/ml, respectively. Although the intrapulmonary rifapentine concentrations were less than the plasma rifapentine concentrations at all time periods, they remained above the proposed breakpoint for M. tuberculosis (0.5 microg/ml) for the 48-h observation period. These data provide a pharmacokinetic rationale for extended interval dosing. The optimum dosing regimen for rifapentine will have to be determined by controlled clinical trials. PMID- 10722502 TI - Towards an understanding of the mechanism of pyrimethamine-sulfadoxine resistance in Plasmodium falciparum: genotyping of dihydrofolate reductase and dihydropteroate synthase of Kenyan parasites. AB - The antifolate combination of pyrimethamine (PM) and sulfadoxine (SD) is the last affordable drug combination available for wide-scale treatment of falciparum malaria in Africa. Wherever this combination has been used, drug-resistant parasites have been selected rapidly. A study of PM-SD effectiveness carried out between 1997 and 1999 at Kilifi on the Kenyan coast has shown the emergence of RI and RII resistance to PM-SD (residual parasitemia 7 days after treatment) in 39 out of 240 (16.25%) patients. To understand the mechanism that underlies resistance to PM-SD, we have analyzed the dihydrofolate reductase (DHFR) and dihydropteroate synthase (DHPS) genotypes of 81 patients. Fifty-one samples were obtained, before treatment, from patients who remained parasite free for at least 7 days after treatment. For a further 20 patients, samples were obtained before treatment and again when they returned to the clinic with parasites 7 days after PM-SD treatment. Ten additional isolates were obtained from patients who were parasitemic 7 days after treatment but who were not sampled before treatment. More than 65% of the isolates (30 of 46) in the initial group had wild-type or double mutant DHFR alleles, and all but 7 of the 47 (85%) had wild-type DHPS alleles. In the paired (before and after treatment) samples, the predominant combinations of DHFR and DHPS alleles before treatment were of triple mutant DHFR and double mutant DHPS (41% [7 of 17]) and of double mutant DHFR and double mutant DHPS (29% [5 of 17]). All except one of the posttreatment isolates had triple mutations in DHFR, and most of these were "pure" triple mutants. In these isolates, the combination of a triple mutant DHFR and wild-type DHPS was detected in 6 of 29 cases (20.7%), the combination of a triple mutant DHFR and a single mutant (A437G) DHPS was detected in 4 of 29 cases (13.8%), and the combination of a triple mutant DHFR and a double mutant (A437G, L540E) DHPS was detected in 16 of 29 cases (55.2%). These results demonstrate that the triply mutated allele of DHFR with or without mutant DHPS alleles is associated with RI and RII resistance to PM-SD. The prevalence of the triple mutant DHFR-double mutant DHPS combination may be an operationally useful marker for predicting the effectiveness of PM-SD as a new malaria treatment. PMID- 10722503 TI - TLA-1: a new plasmid-mediated extended-spectrum beta-lactamase from Escherichia coli. AB - Escherichia coli R170, isolated from the urine of an infected patient, was resistant to expanded-spectrum cephalosporins, aztreonam, ciprofloxacin, and ofloxacin but was susceptible to amikacin, cefotetan, and imipenem. This particular strain contained three different plasmids that encoded two beta lactamases with pIs of 7.0 and 9.0. Resistance to cefotaxime, ceftazidime, aztreonam, trimethoprim, and sulfamethoxazole was transferred by conjugation from E. coli R170 to E. coli J53-2. The transferred plasmid, RZA92, which encoded a single beta-lactamase, was 150 kb in length. The cefotaxime resistance gene that encodes the TLA-1 beta-lactamase (pI 9.0) was cloned from the transconjugant by transformation to E. coli DH5alpha. Sequencing of the bla(TLA-1) gene revealed an open reading frame of 906 bp, which corresponded to 301 amino acid residues, including motifs common to class A beta-lactamases: (70)SXXK, (130)SDN, and (234)KTG. The amino acid sequence of TLA-1 shared 50% identity with the CME-1 chromosomal class A beta-lactamase from Chryseobacterium (Flavobacterium) meningosepticum; 48.8% identity with the VEB-1 class A beta-lactamase from E. coli; 40 to 42% identity with CblA of Bacteroides uniformis, PER-1 of Pseudomonas aeruginosa, and PER-2 of Salmonella typhimurium; and 39% identity with CepA of Bacteroides fragilis. The partially purified TLA-1 beta-lactamase had a molecular mass of 31.4 kDa and a pI of 9.0 and preferentially hydrolyzed cephaloridine, cefotaxime, cephalothin, benzylpenicillin, and ceftazidime. The enzyme was markedly inhibited by sulbactam, tazobactam, and clavulanic acid. TLA-1 is a new extended-spectrum beta-lactamase of Ambler class A. PMID- 10722504 TI - Risk factors for recovery of ampicillin-sulbactam-resistant Escherichia coli in hospitalized patients. AB - Ampicillin-sulbactam resistance in Escherichia coli is an emerging problem. This study determined risk factors for the recovery of ampicillin-sulbactam-resistant E. coli in hospitalized patients. A case-control design was used to compare two groups of case patients with control patients. The first group of case patients consisted of patients from whom nosocomially acquired ampicillin-sulbactam resistant E. coli strains were isolated, and the second group of case patients consisted of patients from whom ampicillin-sulbactam-susceptible E. coli strains were isolated. Control patients were a random selection among 5% of all patients admitted during the same time period. Risk factors analyzed included antimicrobial drug exposure, comorbid conditions, and demographics. Univariate and multivariate analyses were performed. Ampicillin-sulbactam-resistant E. coli strains were isolated from 175 patients, and ampicillin-sulbactam-susceptible E. coli strains were isolated from 577 patients. Nine hundred thirty-four control patients were selected. Exposure to penicillin antibiotics as a class and to ampicillin and ampicillin-sulbactam individually were the only significant, independent risk factors associated with the isolation of ampicillin-sulbactam resistant E. coli (odds ratio [OR] = 2.32 [P < 0.001], OR = 3.04 [P = 0.02], and OR = 1.72 [P = 0.04], respectively), but they were not associated with the isolation of ampicillin-sulbactam-susceptible E. coli. Interestingly, exposure to piperacillin-tazobactam tended to protect against the isolation of E. coli strains resistant to ampicillin-sulbactam, but this did not reach statistical significance (OR = 0.13; P = 0.11). PMID- 10722505 TI - In vivo activity and pharmacokinetics of ziracin (SCH27899), a new long-acting everninomicin antibiotic, in a murine model of penicillin-susceptible or penicillin-resistant pneumococcal pneumonia. AB - The effectiveness of ziracin (SCH27899), a novel everninomicin, was at first investigated against lethal pneumonia caused by a penicillin-susceptible Streptococcus pneumoniae strain. A single intravenous injection of ziracin at a dose of 60 mg/kg of body weight given at 18 h postinfection protected 100% mice and led to the complete clearance of bacteria from their lungs. The activity of ziracin was observed to be the same as that of ceftriaxone: the 50% protective doses (PD(50)s) of ziracin and ceftriaxone were 24.8 and 24.6 mg/kg, respectively. Evaluation of this therapy with leukopenic mice showed that a single injection of ziracin protected 75% of these mice. A delay in therapy with ziracin, which was initiated at 48 h postinfection with 30 mg/kg given once daily for 3 days, resulted in an 83% survival rate of immunocompetent mice. The efficacy of ziracin was further compared to that of vancomycin against lethal pneumonia caused by a penicillin-resistant S. pneumoniae strain in leukopenic mice. The PD(50)s of ziracin and vancomycin were 40.5 and 44.2 mg/kg, respectively. Treatment with ziracin at 30 mg/kg once daily for 2 days (initiated 18 h postinfection) yielded an 83% survival rate and achieved complete eradication of the bacteria. The results were the same as those obtained with vancomycin administered at 15 mg/kg twice daily for 2 days. It is notable that the high survival rates for mice treated with ziracin were associated with effective eradication of the bacteria and rapid recovery of pulmonary tissues from pneumonia. The pharmacokinetic properties of ziracin, ceftriaxone, and vancomycin were estimated following intravenous administration of a single dose of 30 mg/kg to immunocompetent mice. The half-life of ziracin was observed to be longer than those of ceftriaxone and vancomycin (2.3 h versus 1.0 and 0.36 h in the bloodstream and 3 h versus 1.9 and 0. 45 h in lung tissues). The areas under the concentration-time curves (AUCs) in lung tissue for ziracin versus those for ceftriaxone and vancomycin were 36 microg. h/g versus 20 and 9.5 microg. h/g. The prolonged half-life and high AUC for ziracin in tissue contributed to its excellent in vivo activities. PMID- 10722506 TI - Identification of Cryptosporidium parvum dihydrofolate reductase inhibitors by complementation in Saccharomyces cerevisiae. AB - There is a pressing need for drugs effective against the opportunistic protozoan pathogen Cryptosporidium parvum. Folate metabolic enzymes and enzymes of the thymidylate cycle, particularly dihydrofolate reductase (DHFR), have been widely exploited as chemotherapeutic targets. Although many DHFR inhibitors have been synthesized, only a few have been tested against C. parvum. To expedite and facilitate the discovery of effective anti-Cryptosporidium antifolates, we have developed a rapid and facile method to screen potential inhibitors of C. parvum DHFR using the model eukaryote, Saccharomyces cerevisiae. We expressed the DHFR genes of C. parvum, Plasmodium falciparum, Toxoplasma gondii, Pneumocystis carinii, and humans in the same DHFR-deficient yeast strain and observed that each heterologous enzyme complemented the yeast DHFR deficiency. In this work we describe our use of the complementation system to screen known DHFR inhibitors and our discovery of several compounds that inhibited the growth of yeast reliant on the C. parvum enzyme. These same compounds were also potent or selective inhibitors of the purified recombinant C. parvum DHFR enzyme. Six novel lipophilic DHFR inhibitors potently inhibited the growth of yeast expressing C. parvum DHFR. However, the inhibition was nonselective, as these compounds also strongly inhibited the growth of yeast dependent on the human enzyme. Conversely, the antibacterial DHFR inhibitor trimethoprim and two close structural analogs were highly selective, but weak, inhibitors of yeast complemented by the C. parvum enzyme. Future chemical refinement of the potent and selective lead compounds identified in this study may allow the design of an efficacious antifolate drug for the treatment of cryptosporidiosis. PMID- 10722508 TI - In vitro activities of nontraditional antimicrobials against multiresistant Acinetobacter baumannii strains isolated in an intensive care unit outbreak. AB - Fifteen multiresistant Acinetobacter baumannii isolates from patients in intensive care units and 14 nonoutbreak strains were tested to determine in vitro activities of nontraditional antimicrobials, including cefepime, meropenem, netilmicin, azithromycin, doxycycline, rifampin, sulbactam, and trovafloxacin. The latter five drugs were further tested against four of the strains for bactericidal or bacteriostatic activity by performing kill-curve studies at 0.5, 1, 2, and 4 times their MICs. In addition, novel combinations of drugs with sulbactam were examined for synergistic interactions by using a checkerboard configuration. MICs at which 90% of the isolates tested were inhibited for antimicrobials showing activity against the multiresistant A. baumannii strains were as follows (in parentheses): doxycycline (1 microg/ml), azithromycin (4 microg/ml), netilmicin (1 microg/ml), rifampin (8 microg/ml), polymyxin (0.8 U/ml), meropenem (4 microg/ml), trovafloxacin (4 microg/ml), and sulbactam (8 microg/ml). In the kill-curve studies, azithromycin and rifampin were rapidly bactericidal while sulbactam was more slowly bactericidal. Trovafloxacin and doxycycline were bacteriostatic. None of the antimicrobials tested were bactericidal against all strains tested. The synergy studies demonstrated that the combinations of sulbactam with azithromycin, rifampin, doxycycline, or trovafloxacin were generally additive or indifferent. PMID- 10722507 TI - Pharmacologic characteristics of indinavir, didanosine, and stavudine in human immunodeficiency virus-infected children receiving combination therapy. AB - The use of human immunodeficiency virus (HIV) protease inhibitors in children has lagged behind that in adults because of the lack of suitable pediatric formulations and information on safe and effective dosing regimens. This study was designed to obtain pharmacokinetic information on indinavir, administered to HIV-infected children also receiving therapy with two nucleoside agents, and to explore relationships between pharmacokinetic parameters and anti-HIV effect. Indinavir was initiated at a dose of 500 mg/m(2) every 8 h. Plasma indinavir concentrations were measured every 4 weeks; the dose or dosing interval was adjusted to maintain trough concentrations of > or =0.1 mg/liter. All children were evaluated clinically at baseline and every 4 weeks. Plasma HIV RNA was quantitated at baseline and at weeks 4, 12, and 24. Eighteen children participated in this study. The average daily dose of indinavir was 2,043 mg/m(2); nine children received indinavir at 6-h intervals. Pharmacokinetic characteristics of indinavir (mean +/- standard deviation) were the following: oral clearance, 1.4 +/- 0.5 liters/h/kg; half-life, 1.1 +/- 0.43 h; and trough concentration, 0. 29 +/- 0.32 mg/liter. In nine children that completed 24 weeks of therapy, the baseline-to-week-24 change in HIV RNA level was related to indinavir trough concentration and didanosine area under the curve. This study illustrates the ability to obtain pharmacokinetic information from children during routine clinic visits and to use this information to provide a safeguard against underdosing. The incorporation of pharmacologic knowledge with virologic, immunologic, and behavioral considerations should result in improved clinical outcomes for children infected with HIV. PMID- 10722509 TI - Single-dose pharmacokinetics and safety of the oral antiviral compound adefovir dipivoxil in children infected with human immunodeficiency virus type 1. The Pediatrics AIDS Clinical Trials Group. AB - The acyclic phosphonate analog adefovir is a potent inhibitor of retroviruses, including human immunodeficiency virus (HIV) type 1, and, unlike some antiviral nucleosides, does not require the initial phosphorylation step for its activity. Two oral dosages of the adefovir prodrug adefovir dipivoxil were evaluated in a phase I study with children with HIV infection. A total of 14 patients were stratified into age groups ranging from 6 months to 18 years of age. Eight patients received 1.5 mg of adefovir dipivoxil per kg of body weight, and six patients received 3.0 mg of adefovir dipivoxil per kg. Serum samples were obtained at intervals during the 8 h postdosing and were analyzed for adefovir concentrations. Patients were monitored for adverse effects. All samples collected resulted in quantifiable levels of adefovir (lower limit of quantitation, 25 ng/ml) from each patient. The areas under the concentration versus-time curves (AUCs) were similar (P = 0.85) for the 1.5- and 3.0-mg/kg doses, while the apparent oral clearance (CL/F) was significantly higher (P = 0.05) for the 3-mg/kg dose. Pharmacokinetic parameters differed by patient age. In comparing those children older and younger than the median age of 5.1 years, AUC (P = 0.03), maximum concentration of drug in serum (P = 0.004), and the concentration at 8 h postdosing (P = 0.02) were significantly lower for the younger children. There were no significant differences for apparent volume of distribution and CL/F normalized to body surface area, but there was a suggestive difference in half-life (P = 0.07) among the subjects in the older and younger age groups. No significant adverse events were encountered. These data provide the basis for a multidose phase II study of adefovir dipivoxil in HIV-infected infants and children. PMID- 10722510 TI - Potent and selective activity of a combination of thymidine and 1843U89, a folate based thymidylate synthase inhibitor, against Plasmodium falciparum. AB - Unlike mammalian cells, malarial parasites are completely dependent on the de novo pyrimidine pathway and lack the enzymes to salvage preformed pyrimidines. In the present study, first, it is shown that 1843U89, even without polyglutamylation, is a potent folate-based inhibitor of purified malarial parasite thymidylate synthase. The binding was noncompetitive with respect to methylenetetrahydrofolate, and 1843U89 had a K(i) of 1 nM. The compound also had potent antimalarial activity in vitro. Plasmodium falciparum cells in culture were inhibited by 1843U89, with a 50% inhibitory concentration of about 70 nM. The compound was effective against drug-sensitive as well as drug-resistant clones of P. falciparum. As predicted by the biochemistry of the parasite, the potent inhibition of parasite proliferation by 1843U89 could not be reversed with 10 microM thymidine. In contrast, in the presence of 10 microM thymidine, mammalian cells were unaffected by 1843U89 even at concentrations as high as 0.1 mM, thus offering a selectivity window of more than 1,000-fold. On this basis, folate-based thymidylate synthase inhibitors may represent a powerful additional tool that can be used to combat drug-resistant malaria. PMID- 10722511 TI - The triple combination indinavir-zidovudine-lamivudine is highly synergistic. AB - Administration of the combination of indinavir-zidovudine-lamivudine has been demonstrated to cause a large fraction of treated patients to have a decline in human immunodeficiency virus type 1 (HIV-1) copy number to below the detectability of sensitive assays. A recent investigation (G. L. Drusano, J. A. Bilello, D. S. Stein, M. Nessly, A. Meibohm, E. A. Emini, P. Deutsch, J. Condra, J. Chodakewitz, and D. J. Holder, J. Infect. Dis. 178:360-367, 1998) demonstrated that the durability of the antiviral effect was affected by combination chemotherapy. Zidovudine-lamivudine-indinavir differed significantly from the combination of zidovudine plus indinavir. We hypothesized that the addition of lamivudine might alter the regimen, producing a synergistic anti-HIV effect. In vitro analysis of drug interaction demonstrated that zidovudine-indinavir interacted additively. The addition of lamivudine in concentrations which suppressed viral replication by 20% or less by itself demonstrated marked increases in the synergy volume, increasing the synergy volume 20-fold with the addition of 320 nM lamivudine (which does not suppress HIV by itself) and 40-fold with the addition of 1,000 nM lamivudine (20% viral inhibition as a single agent). A fully parametric analysis with a newly developed model for three-drug interaction confirmed and extended these observations. The interaction term (alpha(IND,AZT, 3TC)) for all three drugs showed the greatest degree of synergy. This marked synergistic interaction among the three agents may explain some of the clinical results which differentiate this regimen from the double-drug regimen of zidovudine plus indinavir. PMID- 10722512 TI - Pronounced postantibiotic effect of quinupristin-dalfopristin in static cultures of Staphylococcus aureus: an effect not seen with other antibiotics. AB - Quinupristin-dalfopristin produced postantibiotic effects (PAEs) on exponentially growing (log phase) and nongrowing (lag phase) cultures of 0.4 to 6.9 h. PAEs of the other antibiotics tested were 0.5 to 1.9 h, determined with exponentially growing cultures. None of these antibiotics show a PAE on lag-phase bacteria. PMID- 10722513 TI - In vitro activities of daptomycin, vancomycin, linezolid, and quinupristin dalfopristin against Staphylococci and Enterococci, including vancomycin- intermediate and -resistant strains. AB - The in vitro activity of daptomycin was compared with those of vancomycin, linezolid, and quinupristin-dalfopristin against a variety (n = 203) of gram positive bacteria, including methicillin-resistant Staphylococcus aureus and S. epidermidis (MRSA and MRSE, respectively), vancomycin-resistant enterococci (VRE), and vancomycin-intermediate S. aureus (VISA). Overall, daptomycin was more active against all organisms tested, except Enterococcus faecium and VISA, against which its activity was similar to that of quinupristin-dalfopristin. In time-kill studies with MRSA, MRSE, VRE, and VISA, daptomycin demonstrated greater bactericidal activity than all other drugs tested, killing > or =3 log CFU/ml by 8 h. Daptomycin may be a potential alternative drug therapy for multidrug resistant gram-positive organisms and warrants further investigation. PMID- 10722514 TI - Coordinate suppression of superantigen-induced cytokine production and T-cell proliferation by a small nonpeptidic inhibitor of class II major histocompatibility complex and CD4 interaction. AB - Proinflammatory cytokines mediate the toxic effect of superantigenic staphylococcal exotoxins (SE). TJU103, a small nonpeptidic molecule that blocks the interaction between major histocompatibility complex class II and CD4 molecules inhibited SE-stimulated T-cell proliferation (by 92%) and production of tumor necrosis factor, interleukin 1beta, interleukin 6, and gamma interferon (by 66, 56, 76, and 72%, respectively) by human peripheral blood mononuclear cells. These data suggest that TJU103 may be useful for mitigating the pathogenic effects of SE. PMID- 10722515 TI - Nucleotide sequence of the bla(RTG-2) (CARB-5) gene and phylogeny of a new group of carbenicillinases. AB - We determined the nucleotide sequence of the bla gene for the Acinetobacter calcoaceticus beta-lactamase previously described as CARB-5. Alignment of the deduced amino acid sequence with those of known beta-lactamases revealed that CARB-5 possesses an RTG triad in box VII, as described for the Proteus mirabilis GN79 enzyme, instead of the RSG consensus characteristic of the other carbenicillinases. Phylogenetic studies showed that these RTG enzymes constitute a new, separate group, possibly ancestors of the carbenicillinase family. PMID- 10722516 TI - Mutations in the beginning of the rpoB gene can induce resistance to rifamycins in both Helicobacter pylori and Mycobacterium tuberculosis. AB - A clinical isolate of Helicobacter pylori that developed resistance to rifabutin during therapy carried an rpoB gene that retained a wild-type cluster region sequence but had acquired a novel codon 149 (V149F) mutation. In transformation experiments, the mutation was shown to confer high-level rifabutin resistance. The equivalent mutation (V176F) was present in several resistant isolates of Mycobacterium tuberculosis. PMID- 10722517 TI - Susceptibilities of oral and nasal isolates of Streptococcus mitis and Streptococcus oralis to macrolides and PCR detection of resistance genes. AB - The susceptibility of viridans group streptococci to macrolides was determined. Thirteen isolates (17%) were resistant to erythromycin. Five strains carried an erm gene that was highly homologous to that in Tn917. Four strains had mefE genes that coded erythromycin efflux ability. PMID- 10722518 TI - SHV-13, a novel extended-spectrum beta-lactamase, in Klebsiella pneumoniae isolates from patients in an intensive care unit in Amsterdam. AB - Eleven clonally related Klebsiella pneumoniae isolates were examined. These had been isolated at an intensive care unit in Amsterdam in 1994. Their resistance was associated with a conjugative 170-kb plasmid which encoded a novel SHV beta lactamase designated SHV-13. The SHV-13 enzyme had two substitutions compared with SHV-1: Leu35Gln and Gly238Ala. It hydrolyzed cefotaxime much more rapidly than ceftazidime or aztreonam. PMID- 10722519 TI - Antipneumococcal activities of GAR-936 (a new glycylcycline) compared to those of nine other agents against penicillin-susceptible and -resistant pneumococci. AB - GAR-936, a new glycylcycline, had lower MICs (< or =0.016 to 0.125 microg/ml) for 201 penicillin- and tetracycline-susceptible and -resistant pneumococcal strains than tetracycline (< or = 0.06 to 128 microg/ml), minocycline (< or =0.06 to 16.0 microg/ml), or doxycycline (< or =0.06 to 32.0 microg/ml). GAR-936 was also bactericidal against 11 of 12 strains tested at the MIC after 24 h, with significant kill rates at earlier time points. PMID- 10722520 TI - A Canadian national surveillance study of urinary tract isolates from outpatients: comparison of the activities of trimethoprim-sulfamethoxazole, ampicillin, mecillinam, nitrofurantoin, and ciprofloxacin. The Canadian Urinary Isolate Study Group. AB - Ampicillin, trimethoprim-sulfamethoxazole, mecillinam, nitrofurantoin, and ciprofloxacin mean resistance rates for 2,000 urinary tract isolates collected from outpatients across Canada in 1998 were 41.1, 19.2, 14.7, 5.0, and 1.8%, respectively. For Escherichia coli isolates alone (n = 1,681) comparable rates were 41. 0, 18.9, 7.4, 0.1, and 1.2%, respectively. The majority of E. coli isolates resistant to ampicillin, trimethoprim-sulfamethoxazole, or ciprofloxacin were susceptible (MIC, <16 microg/ml) to mecillinam. PMID- 10722522 TI - Toward antiviral strategies that resist viral escape. AB - We studied the effect on viral growth of drugs targeting different virus functions using a computer simulation for the intracellular growth of bacteriophage T7. We found that drugs targeting components of negative-feedback loops gain effectiveness against mutant viruses that attenuate the drug-target interaction. The greater inhibition of such mutants than of the wild type suggests a drug design strategy that would hinder the development of drug resistance. PMID- 10722521 TI - Control of staphylococcal adhesion to polymethylmethacrylate and enhancement of susceptibility to antibiotics by poloxamer 407. AB - We studied the antiadhesive effect of Poloxamer 407 (P407), together with modifications in the antimicrobial susceptibility of residual adherent staphylococci. Bacterial adherence was markedly inhibited (77% to more than 99.9%) whether polymethylmethacrylate was exposed to P407 before or during the adherence assay. Furthermore, residual adherent staphylococci appeared to be more susceptible to antibiotic activity, suggesting that combination of P407 with antibiotics could be a promising approach to the prevention of infection of foreign material. PMID- 10722523 TI - Effect of fluconazole on the pharmacokinetics of doxorubicin in nonhuman primates. AB - Antifungal prophylaxis in cancer patients who are undergoing chemotherapy is associated with prolonged neutropenia. We measured the effect of fluconazole on doxorubicin pharmacokinetics in nonhuman primates to determine if neutropenia is related to a pharmacokinetic interaction that delays the clearance of the chemotherapeutic agent. Fluconazole pretreatment had no effect on doxorubicin pharmacokinetics. PMID- 10722524 TI - In vitro activities of sitafloxacin (DU-6859a) and six other fluoroquinolones against 8,796 clinical bacterial isolates. AB - The in vitro activities of sitafloxacin, ciprofloxacin, trovafloxacin, levofloxacin, clinafloxacin, gatifloxacin, and moxifloxacin against 5,046 gram negative bacteria, 3,344 gram-positive cocci, and 406 anaerobes were determined. Sitafloxacin was the most active agent against gram-positive cocci and anaerobes. Against Enterobacteriaceae and nonfermenters, its activity was either equivalent to or better than that of clinafloxacin. PMID- 10722525 TI - Postantifungal effects of echinocandin, azole, and polyene antifungal agents against Candida albicans and Cryptococcus neoformans. AB - The postantifungal effect (PAFE) of fluconazole, MK-0991, LY303366, and amphotericin B was determined against isolates of Candida albicans and Cryptococcus neoformans. Concentrations ranging from 0. 125 to 4 times the MIC were tested following exposure to the antifungal for 0.25 to 1 h. Combinations of azole and echinocandin antifungals (MK-0991 and LY303366) were tested against C. neoformans. Fluconazole displayed no measurable PAFE against Candida albicans or Cryptococcus neoformans, either alone or in combination with either echinocandin antifungal. MK-0991, LY303366, and amphotericin B displayed a prolonged PAFE of greater than 12 h against Candida spp. when tested at concentrations above the MIC for the organism and 0 to 2 h when tested at concentrations below the MIC for the organism. PMID- 10722527 TI - New antimicrobial agents approved by the U.S. Food and Drug Administration in 1999 and new indications for previously approved agents. PMID- 10722526 TI - In vitro activity of ABT 773, a new ketolide antibiotic, against Chlamydia pneumoniae. AB - The in vitro activities of ABT 773, telithromycin (HMR 3647), azithromycin, clarithromycin, erythromycin, and levofloxacin were tested against 20 strains of Chlamydia pneumoniae. The MIC at which 90% of the isolates were inhibited and the minimal bactericidal concentration at which 90% of the isolates were killed by ABT 773 were 0.015 microg/ml (range, 0.008 to 0.015 microg/ml). ABT 773 was the most active antibiotic tested in this study. PMID- 10722528 TI - Troponin T or troponin I as cardiac markers in ischaemic heart disease. PMID- 10722530 TI - Balloon mitral valvuloplasty in the elderly. PMID- 10722531 TI - The aortic root: structure, function, and surgical reconstruction. PMID- 10722532 TI - Protecting the ischaemic and reperfused myocardium in acute myocardial infarction: distant dream or near reality? PMID- 10722533 TI - Images in cardiology. Candle flame appearance in dissection of aorta. PMID- 10722534 TI - Patients' interpretation of symptoms as a cause of delay in reaching hospital during acute myocardial infarction. AB - OBJECTIVE: To examine whether the association between expected symptoms of acute myocardial infarction and actual symptoms predicted delay in reaching hospital and help seeking behaviour. DESIGN: During hospital convalescence, participants completed a structured interview designed to measure symptom experience and help seeking behaviour following the onset of symptoms of acute myocardial infarction. PATIENTS: 88 patients admitted to hospital with their first myocardial infarction MAIN OUTCOME MEASURES: Delay in reaching hospital from onset of worst symptoms, obtained from ambulance and hospital records. RESULTS: The most common symptoms expected by patients with myocardial infarction were central chest pain (76%), radiating arm or shoulder pain (34%), and collapse (26%). The most common symptoms experienced were sweats or feeling feverish (78%), chest pain (64%), and arm, shoulder, or radiating pain (66%). A mismatch between symptoms experienced and those expected occurred in 58% of patients, and was associated with delay. Patients who experienced a mismatch between expectation and actual symptoms also were more likely to have a third party decide to call for help. CONCLUSIONS: The experience and interpretation of symptoms is an important source of delay and help seeking following onset of myocardial infarction symptoms. PMID- 10722535 TI - Images in cardiology. Intermittent mitral valve occlusion by a mobile left atrial thrombus. PMID- 10722536 TI - Alcohol intake and mortality in middle aged men with diagnosed coronary heart disease. AB - OBJECTIVE: To examine the effects of alcohol on risk of mortality from coronary heart disease (CHD), cardiovascular disease, and all causes in men with established CHD. METHODS AND RESULTS: In a population based prospective study of 7169 men aged 45-64 years followed for a mean of 12.8 years, 655 men (9.1%) had a physician diagnosis of CHD (myocardial infarction 455, angina only 200). In these 655 men, there were 294 deaths from all causes including 175 CHD deaths. Ex drinkers had the highest risk of CHD, cardiovascular mortality, and all cause mortality even after adjustment for lifestyle characteristics and pre-existing disease. Using occasional drinkers as the reference group, lifelong teetotallers, occasional drinkers, and light drinkers all showed similar risks of mortality from CHD, cardiovascular disease, and all causes. Moderate/heavy drinkers showed increased risk of mortality from CHD, cardiovascular disease, and all causes compared to occasional drinkers. The adverse effect of moderate/heavy drinking was confined to the 455 men with previous myocardial infarction (adjusted relative risk for all cause mortality 1.50, 95% confidence interval 1.01 to 2.23). In contrast to lighter drinking, giving up smoking within five years of the start of follow up was associated with a considerable reduction in risk of all cause and cardiovascular mortality compared to those who continued to smoke. CONCLUSION: Compared to occasional drinking, regular light alcohol consumption (1 14 units per week) in men with established coronary heart disease is not associated with any significant benefit or deleterious effect for CHD, cardiovascular disease or all cause mortality. Higher levels of intake (>/= 3 drinks per day) are associated with increased mortality in men with previous myocardial infarction. In contrast, smoking cessation in men with established CHD substantially reduces the risk of mortality. PMID- 10722537 TI - Transthoracic ultrasonic visualisation of coronary aneurysm, stenosis, and occlusion in Kawasaki disease. AB - OBJECTIVE: To determine the sensitivity and specificity of our transthoracic echocardiographic technique using high frequency (7.5 MHz) transducers for identification of the presence and type of coronary artery disease in patients with Kawasaki disease. DESIGN: The results of the prospective echocardiographic study in each of seven segments of the four major coronary arteries were compared with the selective coronary angiograms. SETTING: Kitasato University Hospital. SUBJECTS: 60 patients with Kawasaki disease, ranging in age from 8.0 months to 22 years (median, 6.0 years). RESULTS: Adequate echocardiographic images were obtained in 397 (95%) of 420 coronary segments. Coronary angiography showed the presence of coronary aneurysms in 87 segments and stenosis or occlusion in 28. The overall sensitivity and specificity of cross sectional echocardiography for correctly identifying coronary aneurysms were 95% and 99%, respectively; for correctly identifying coronary stenosis or occlusion the values were 85% and 98% for the right coronary artery, and 80% and 97% for the left anterior descending coronary artery. Agreement on the presence or absence of coronary aneurysms and obstructive lesions on echocardiograms between the two observers was 1.0 and 0.98, respectively. CONCLUSIONS: Echocardiography may provide a non-invasive means of identifying the presence and type of coronary artery disease in patients with Kawasaki disease. PMID- 10722538 TI - Treatment with epoprostenol reverts nitric oxide non-responsiveness in patients with primary pulmonary hypertension. AB - OBJECTIVE: To assess whether long term treatment with epoprostenol might restore primary non-responsiveness to nitric oxide (NO) in patients with primary pulmonary hypertension. METHODS: Seven patients with primary pulmonary hypertension receiving intravenous epoprostenol continuously because of failure of NO to influence pulmonary haemodynamics during initial testing were followed over a period of 13-29 months. Afterwards, acute vascular reactivity towards NO was tested again during right heart catheterisation. RESULTS: Administration of NO after continuous epoprostenol treatment for a mean period of 18 months improved arterial oxygen saturation (p < 0.01) and cardiac index (p < 0.05), and decreased mean pulmonary artery pressure (p < 0.01) and total pulmonary vascular resistance (p < 0.01) in patients previously unresponsive to NO. CONCLUSIONS: Long term treatment with epoprostenol reverts initial refractoriness to NO in patients with primary pulmonary hypertension. Thus the addition of NO to epoprostenol treatment might cause further improvement in the course of the disease. PMID- 10722539 TI - Sudden death in children and adolescents. AB - OBJECTIVE: To identify the incidence, causes, and characteristics of sudden death at age 1-20 years. DESIGN: A review of all deaths at age 1-20 years. Death certificates were obtained from the Office for National Statistics, and further information, where appropriate, from coroners, paediatricians, physicians, and pathologists. SETTING: The resident population of one English health region in 1985-1994. RESULTS: In a population of 806 500 children and adolescents aged 1-20 years there were 2523 deaths in 10 years. Medical causes accounted for 1017 deaths (40%); 1236 (49%) were unnatural, and 270 (11%) were sudden. These sudden deaths comprised 142 with a previous diagnosis, the commonest being epilepsy 49 (34%), cardiovascular disease 33 (23%), and asthma 30 (21%); 87 attributed to a cause discovered at necropsy, which was respiratory infection in 32 (37%), other infections in 17 (20%), and unsuspected cardiovascular abnormalities in 26 (30%); 41 remained unexplained. CONCLUSIONS: Half of all sudden deaths in children or adolescents were attributed to an already diagnosed condition. Abnormalities identified at necropsy accounted for one third of sudden deaths. Undiagnosed hypertrophic cardiomyopathy caused less than one death per million person years in the population aged 1-20 years. Unexplained sudden death, which may be caused by primary cardiac arrhythmia, is probably about 10 times more common. PMID- 10722540 TI - Temporal variability in birth prevalence of cardiovascular malformations. AB - OBJECTIVE: To investigate changes over time in the prevalence at live birth of cardiovascular malformations and to compare "anatomical" and "physiological" diagnostic hierarchies within a population. DESIGN: Retrospective and prospective ascertainment of all congenital cardiovascular malformations diagnosed in infancy. SETTING: The resident population of one health region. PATIENTS: All infants live born from 1985 to 1997 with cardiovascular malformations confirmed by echocardiography, cardiac catheterisation, surgery or autopsy. MAIN OUTCOME MEASURES: Year to year variation in prevalence of individual malformations and of "complex", "significant", and "minor" groups. RESULTS: 2671 babies with cardiovascular malformations were confirmed in a denominator population of 477 960 live births (5.6 per 1000). There was no change over 13 years in the birth prevalence of "complex" or "significant" defects, but a highly significant increase in "minor" defects (p < 0.0001), mainly small ventricular septal defects. Termination of pregnancy increased from no cases in 1985 to 16 in 1997 with no demonstrable effect on live born babies with heart defects. A one dimensional "anatomical" diagnostic hierarchy led to under ascertainment of pulmonary atresia by 27%, coarctation of the aorta by 39%, and interruption of the aorta by 100%. CONCLUSIONS: The apparent increase in live born cardiovascular malformations results mainly from improved diagnosis of minor defects. There has been no change over time in birth prevalence of more serious defects. Spontaneous year to year variation in numbers will make it difficult to ascribe any short term changes to any particular intervention. A two dimensional diagnostic hierarchy is offered as a standard. PMID- 10722541 TI - Serial assessment of left ventricular diastolic function after Fontan procedure. AB - OBJECTIVE: To assess longitudinal changes in systemic ventricular diastolic function late after the Fontan procedure. DESIGN AND PATIENTS: Prospective study of 13 patients at 2.8 (2.0) years (early) and again at 11.4 (2.0) years (late) after the Fontan procedure by Doppler echocardiography with simultaneous ECG, phonocardiogram, and respirometer. SETTING: Tertiary paediatric cardiac centre. RESULTS: The isovolumic relaxation time (IVRT) was significantly longer, and E wave deceleration time, E and A wave velocities, and E:A velocity ratio were reduced compared to normal both early and late after the procedure. The mean (SD) z score of IVRT decreased significantly from +2.50 (1.00) to +1.24 (0.80) (p = 0.002), and the z score of the E wave deceleration time decreased from -1.69 (1.31) to -2.40 (1.47) (p = 0.03) during follow up. The A wave deceleration time also tended to decrease (early 80 (12) ms v late 73 (11) ms, p = 0.13) with increased follow up. There were no changes of the E and A wave velocities and E:A velocity ratio. The E wave velocity was inversely related to IVRT both early (r = -0.82, p = 0.001) and late (r = -0.59, p = 0.034) after the operation. The prevalence of diastolic flow during isovolumic relaxation decreased from 85% (11/13) to 38% (5/13) (p = 0.04), while that of mid diastolic flow increased from 23% (3/13) to 77% (10/13) (p = 0.02) between the two assessments. CONCLUSIONS: Left ventricular diastolic function remains highly abnormal late after the Fontan procedure. The longitudinal changes demonstrated on follow up are compatible with reduction of left ventricular compliance in addition to persisting abnormalities of relaxation. PMID- 10722542 TI - Images in cardiology. Ventricular fibrillation provoked by cardioversion and asynchronous pacing. PMID- 10722544 TI - Parsonnet score is a good predictor of the duration of intensive care unit stay following cardiac surgery. AB - OBJECTIVE: To investigate the value of the Parsonnet score (PS) in identifying preoperatively patients that are likely to spend < 24 hours on the intensive care unit (ICU) following cardiac surgery. METHOD: Prospectively collected data on 5591 patients were analysed. PS, mortality, the length of stay on the ICU (ICU LOS), number of patients with clinical evidence of stroke, need for haemofiltration, resternotomy for bleeding, tracheostomy, and use of intra-aortic balloon pump were documented as outcomes. A receiver operating characteristic (ROC) curve constructed using PS as a predictor of ICU stay < 24 hours identified a PS of 10 as the best cut off point that would predict ICU-LOS < 24 hours. The patients were therefore stratified by PS into two groups, those with a PS of 0 to 9 (PS 0-9) and those with a PS of 10 and above (PS 10+). RESULTS: The ROC curve constructed using PS as a predictor of ICU stay < 24 hours had an area under the curve of 0.70 (0.01). The maximum efficiency of the test was at a sensitivity of 0.68. This corresponded to PS 10. The positive predictive value of the test at this score was 90.5%. Patients with PS 0-9 had a mean ICU stay of 1.49 days, while patients with PS 10+ had a mean ICU stay of 2.89 days (p = 0.01). The risk of stroke, use of intra-aortic balloon pump, requirement for haemofiltration, need for tracheostomy, and risk of resternotomy for bleeding were each significantly less in patients with PS 0-9 versus those with a score of PS 10+ (p < 0.01 in all cases). The risk of a single complication was 4.7% (PS 0-9) v 15.2% (PS 10+) (p < 0.01). CONCLUSION: PS is an impartial and objective method of predicting postoperative complications and ICU stay < 24 hours. This is of value in selecting a cohort of patients likely to maintain a smooth flow of patients through the cardiothoracic unit when resources are limited to a few free ICU beds. PMID- 10722543 TI - Intrapulmonary arteriovenous shunting may be a universal phenomenon in patients with the superior cavopulmonary anastomosis: a radionuclide study. AB - OBJECTIVE: To evaluate the extent of intrapulmonary right to left shunting in children after bidirectional cavopulmonary anastomosis (BCPA). DESIGN: Prospective study of patients who underwent BCPA in a single centre. PATIENTS: 17 patients with complex cyanotic congenital cardiac malformations who underwent BCPA at 1-45 months of age (median 21 months) were evaluated 15-64 months postoperatively (median 32 months). Five children between 1 and 10 years (median 5 years) with normal or surgically corrected intracardiac anatomy and peripheral pulmonary circulation who required V/Q scanning for other reasons were used as controls. INTERVENTIONS: All patients underwent cardiac catheterisation to exclude angiographically demonstrable venovenous collaterals followed by pulmonary perfusion scanning using (99m)technetium ((99m)Tc) labelled albumen microspheres to quantify the intrapulmonary right to left shunt. MAIN OUTCOME MEASURE: Percentage of intrapulmonary right to left shunt. RESULTS: The mean (SD) level of physiological right to left shunting found in the control group was 5.4 (2.3)%. All patients with BCPA showed the presence of a significantly higher level of intrapulmonary shunting (26.8 (16.9)%, p < 0.001). The degree of shunting was significantly increased in the subgroup of 11 patients with BCPA as the only source of pulmonary blood flow (34.9 (15.8)%), when compared to the six remaining patients with an additional source of pulmonary blood supply (12.0 (2.6)%, p < 0.001). There was a negative correlation between age at BCPA and the shunt percentage found in the patients with a competitive source of pulmonary blood flow (r = -0.63, p < 0. 01). CONCLUSIONS: Intrapulmonary right to left shunting develops in all patients following BCPA. This may be caused by a sustained and inappropriate vasodilatation resulting from absence or decreased levels of a substance that inhibits pulmonary vasodilatation. Augmenting BCPA with an additional source of blood flow containing hepatic factor limits the degree of intrapulmonary arteriovenous shunting and may help provide successful longer term palliation. PMID- 10722545 TI - Images in cardiology. Postcoarctation giant aneurysm of aorta. PMID- 10722547 TI - Images in cardiology. Exclusion of a pulmonary artery aneurysm using a covered stent. PMID- 10722546 TI - Long term outcome of percutaneous mitral balloon valvotomy in patients aged 70 and over. AB - OBJECTIVE: To assess the immediate haemodynamic improvement and long term symptomatic benefit of percutaneous mitral balloon valvotomy in patients aged over 70 years. DESIGN: Pre- and postprocedure haemodynamic data and follow up for 1 to 10 years by clinic visit or telephone contact. SETTING: Tertiary referral centre in Scotland. SUBJECTS: 80 patients age 70 and over who had mitral balloon dilatation: 55 were considered unsuitable for surgical treatment because of frailty or associated disease. In an additional four patients mitral dilatation was not achieved. MAIN OUTCOME MEASURES: Increase in valve area after balloon dilatation and survival, freedom from valve replacement, and symptom class at follow up. RESULTS: Mean (SD) valve area increased by 89% from 0.84 (0.28) to 1. 59 (0.67) cm(2). There was a low rate of serious complications, with only two patients having long term major sequelae. Of 55 patients unsuitable for surgical treatment, 28 (51%) were alive without valve replacement and with improvement by at least one symptom class at one year, and 14 (25%) at five years. In the 25 patients considered suitable for surgical treatment, 16 (64%) achieved this outcome at one year and nine (36%) at five years. CONCLUSIONS: Percutaneous mitral balloon valvotomy is a safe and useful palliative procedure in elderly patients who are unsuitable for surgery. Balloon dilatation should also be used for elderly patients whose valve appears suitable for improvement by commissurotomy, but echo score is an imperfect predictor of haemodynamic improvement. PMID- 10722548 TI - Neurohormonal activation late after cavopulmonary connection. AB - OBJECTIVE: To determine whether patients with cavopulmonary connection have higher levels of vasoactive/water-salt regulating hormones and if so, whether hormone levels are related to postoperative haemodynamics and postoperative follow up. DESIGN: Cross sectional study. SETTING: University hospital. PATIENTS: 20 patients (New York Heart Association functional class I-II), mean age 11 years (range 4 to 22), were studied at a mean of 2 years (0.5 to 6) after a total cavopulmonary connection (TCPC, n = 12) or a bidirectional Glenn anastomosis (BDG, n = 8). INTERVENTIONS: Cardiac catheterisation was performed and blood samples were drawn. Control blood samples were drawn from 33 healthy children, mean age 12 years (6 to 16). MAIN OUTCOME MEASURES: Plasma levels of angiotensin II, renin, aldosterone, arginine, vasopressin, atrial natriuretic factor (ANF), brain natriuretic peptide (BNP). RESULTS: All neurohormones were significantly increased in both TCPC and BDG patients (p < 0. 05), with a fourfold increase in angiotensin II, renin, and aldosterone, and a twofold increase in vasopressin, ANF, and BNP (compared with healthy controls). There was no correlation between haemodynamic variables and hormone levels. Angiotensin II and renin were inversely correlated with time to follow up. All subjects over 15 years (n = 5) had normal neurohormonal levels. CONCLUSIONS: Neurohormones were raised for years after successful cavopulmonary operations but lower levels were observed with time on follow up. This supports the hypothesis that neurohormonal activation is primarily related to altered postoperative physiology and that adaptation takes place over time. PMID- 10722549 TI - The atrioventricular junctions in Ebstein malformation. AB - OBJECTIVE: To review the anatomical structure of the right atrioventricular junction, including the specialised atrioventricular conduction system, in hearts with Ebstein's malformation, to identify potential substrates for the abnormalities in conduction. METHODS: Five heart specimens representing the morphological spectrum of Ebstein malformation were examined grossly and histologically. RESULTS: On the endocardial surface, the atrioventricular junction was marked by a faint line in two hearts, and by a small ridge in the other three. Analysis of the right parietal junction in four hearts revealed only two accessory muscular atrioventricular connections. A plane of fibrofatty tissue separated atrial from ventricular myocardium in the right parietal junction in all hearts. The compact atrioventricular node was closer to the coronary sinus than usual. Accessory nodoventricular connections were present in four hearts, while accessory fasciculo-ventricular connections were found in one. The right bundle branch was hypoplastic or absent in four hearts. CONCLUSIONS: In this small series, the parietal atrioventricular junction was better developed than previously thought. Structural abnormalities of the atrioventricular conduction system, however, were present. These may account for some of the conduction abnormalities frequently observed with the Ebstein malformation. PMID- 10722550 TI - Production of hepatocyte growth factor during acute myocardial infarction. AB - OBJECTIVE: To investigate the clinical significance of circulating hepatocyte growth factor (HGF) and the role of peripheral blood mononuclear cells (monocytes), which are a possible source of HGF, in patients with acute myocardial infarction. DESIGN AND PATIENTS: 37 patients with acute myocardial infarction and 13 normal control subjects were recruited. Peripheral venous blood samples were drawn from the infarct patients 1, 7, 14, and 21 days after onset. Monocytes were isolated from peripheral blood at those times. HGF concentrations in serum and in a culture medium of monocytes after incubation for 24 hours (monocyte HGF levels) were measured by enzyme linked immunosorbent assay. RESULTS: Serum HGF and monocyte HGF values within seven days after onset of myocardial infarction were significantly higher than those of control subjects and decreased by day 14. There were significant positive correlations between serum HGF and monocyte HGF levels on day 7; between maximum plasma creatine phosphokinase levels and serum HGF levels on day 1; between maximum plasma C reactive protein and serum HGF levels; and between maximum C reactive protein and monocyte HGF levels. Monocyte HGF levels were raised in the patients with progression of ventricular enlargement in the course of acute myocardial infarction. CONCLUSIONS: Early serum HGF concentrations reflect the extent of myocardial damage in acute myocardial infarction patients. Inflammation after acute myocardial infarction is supposed to be involved in enhanced HGF production. Monocytes may play an important role in ventricular remodelling after acute myocardial infarction by releasing the cardiovascular protective mitogen HGF. PMID- 10722552 TI - Coronary stent symmetry and vascular injury determine experimental restenosis. AB - OBJECTIVE: To assess the impact of stent symmetry on restenosis using the coronary overstretch sheep model. METHODS: Neointimal thickness, injury index, and percentage diameter and area stenosis were calculated by digital morphometry. The standard deviation of the angular burden was used to assess stent symmetry for each section. MATERIALS: 15 healthy Merino sheep (63-75 kg) underwent implantation of 30 slotted tube stents (7 mm). Restenosis was induced by calculated overstretch of the coronary artery. Twenty eight days after implantation, stents were excised and underwent histological examination using quantitative digital morphometry. RESULTS: The severity of vessel injury was positively correlated with neointimal thickness and with percentage diameter and area stenosis (p < 0.001). Mean neointimal thickness and mean vascular injury per cross section were strongly related to the standard deviation of angular burden, with correlation coefficients of 0.6 and 0.8, respectively (p < 0.001). CONCLUSIONS: The well known relation between vascular injury and restenosis was confirmed, and a new relation was discovered between stent asymmetry and restenosis. If these results apply to human coronary arteries, restenosis may also be dependent on the degree of asymmetric stent expansion. These results should influence the development of new stent designs to reduce asymmetric stent expansion, leading to a more homogeneous strain distribution in stented coronary segments. PMID- 10722554 TI - Positron emission tomography and myocardial imaging. PMID- 10722553 TI - The cardiomyopathies: an overview. PMID- 10722551 TI - Hibernating myocardium: morphological correlates of inotropic stimulation and glucose uptake. AB - BACKGROUND: In patients with postischaemic left ventricular dysfunction, segments recovering function after revascularisation (hibernating myocardium) may not respond during dobutamine echocardiography, despite preserved [(18)F] 2-fluoro-2 deoxy-D-glucose (FDG) uptake at positron emission tomography. OBJECTIVE: To investigate whether this lack of response might reflect the degree of ultrastructural change in hibernating myocardium. METHODS: Transmural biopsies were obtained from 22 dysfunctional segments in 22 patients during coronary artery bypass grafting and examined by light and electron microscopy. Wall motion scores and coronary vasodilator reserve were assessed before and after coronary artery bypass grafting (CABG). RESULTS: Mean (SD) wall motion score improved in all segments following CABG (from 2.24 (0.4) to 1.55 (0.4); p < 0.0001), confirming hibernating myocardium. In these segments myocardial blood flow (positron emission tomography with H(2)(15)O) before CABG was similar to that in normal volunteers (1.02 (0.24) v 1.02 (0.23) ml/min/g), while the coronary vasodilator reserve was blunted (1.26 (0.7) v 3.2 (1.6); p < 0.0001). Myocardial blood flow was unchanged after CABG, whereas coronary vasodilator reserve increased to 2.10 (0.90) (p < 0.0007). In hibernating myocardium myofibrillar loss, interstitial fibrosis, and glycogen-rich myocytes were more marked than in control donor hearts. On the basis of the response to dobutamine before CABG, two functional groups were identified: group A, segments with inotropic reserve (n = 15); group B, segments without inotropic reserve (n = 7). FDG uptake was similar in group A and group B (0.40 (0.1) v 0.44 (0.1) micromol/min/g). In group B there was more myofibrillar loss (26 (8)% v 11 (5)%; p = 0.0009) and glycogen-rich myocytes (28 (11)% v 17 (10)%; p = 0.02), whereas interstitial fibrosis, myocardial blood flow, and coronary vasodilator reserve were similar in the two groups. Myofibrillar loss was the only independent predictor of inotropic reserve (p = 0.01). CONCLUSIONS: Hibernating myocardium is characterised by a reduced coronary vasodilator reserve which improves on revascularisation and shows a spectrum of ultrastructural changes that influence the response to dobutamine, while FDG uptake is invariably preserved. PMID- 10722555 TI - Intervention in coronary artery disease. PMID- 10722556 TI - Angiographic documented coronary arterial spasm in absence of critical coronary artery stenoses in a patient with variant angina episodes during exercise and dobutamine stress echocardiography. AB - Dobutamine stress echocardiography is widely performed as a useful diagnostic tool in patients with known or suspected coronary artery disease. Dobutamine induced myocardial ischaemia is frequently associated with ST segment depression. ST segment elevation is uncommon and is almost always associated with prior myocardial infarction or transient total coronary occlusion. Dobutamine induced ST segment elevation in absence of significant coronary artery disease is a rare condition and is supposed to be a consequence of severe coronary artery spasm. The case of a 58 year old man with variant angina episodes at rest, during exercise test, and dobutamine stress echocardiography is reported, in whom coronary spasm without significant coronary artery stenoses was documented angiographically. PMID- 10722557 TI - Acute severe thrombocytopenia after treatment with ReoPro (abciximab). AB - ReoPro (abciximab) is an extremely potent inhibitor of the glycoprotein IIb/IIIa receptor, the final common pathway of platelet activation and aggregation. Its main role is the maintenance of coronary patency after suboptimal results with coronary intervention. However, one of the complications of this treatment is excessive bleeding, a problem which may be compounded by a rare idiosyncratic thrombocytopenic reaction. A severe episode of thrombocytopenia in a 64 year old man is described; he was treated with ReoPro for a right coronary stenosis which had not been resolved by angioplasty. His platelet level dropped quickly and only improved after 20 units of platelets were given. PMID- 10722558 TI - Cystic tumour of the atrioventricular nodal region: report of a case successfully treated with surgery. AB - A case is reported of a 59 year old woman who presented with palpitations. Electrocardiographic studies revealed atrial fibrillation and atrioventricular block. Echocardiography and magnetic resonance imaging showed a right atrial cystic mass attached to the interatrial septum. The patient underwent surgical excision of the mass. Histopathological findings were of a cystic tumour of the atrioventricular nodal region. This is the second report of this condition diagnosed antemortem and treated successfully with surgical excision. PMID- 10722559 TI - Left ventricular opacification during selective intracoronary injection of echocardiographic contrast in patients with hypertrophic cardiomyopathy. AB - Percutaneous alcohol ablation of the interventricular septum via the first septal perforator branch of the left anterior descending artery can successfully treat dynamic left ventricular outflow tract obstruction in patients with hypertrophic cardiomyopathy. Increasingly, echocardiographic contrast agents are used before alcohol injection to identify the perfusion bed of the septal perforator vessels. This study describes the unexpected opacification of the left ventricular cavity in three of five consecutive patients following selective injection of the first septal perforator with Optison. This case study demonstrates that direct communication between the first septal perforator vessel and the left ventricle is common, an observation that may have considerable relevance to the technique of alcohol septal reduction. PMID- 10722560 TI - Serum amyloid P component bound to gram-negative bacteria prevents lipopolysaccharide-mediated classical pathway complement activation. AB - Although serum amyloid P component (SAP) is known to bind many ligands, its biological function is not yet clear. Recently, it was demonstrated that SAP binds to lipopolysaccharide (LPS). In the present study, SAP was shown to bind to gram-negative bacteria expressing short types of LPS or lipo-oligosaccharide (LOS), such as Salmonella enterica serovar Copenhagen Re and Escherichia coli J5, and also to clinical isolates of Haemophilus influenzae. It was hypothesized that SAP binds to the bacteria via the lipid A part of LPS or LOS, since the htrB mutant of the nontypeable H. influenzae strain NTHi 2019-B29-3, which expresses a nonacetylated lipid A, did not bind SAP. This was in contrast to the parental strain NTHi 2019. The binding of SAP resulted in a clear inhibition of the deposition of complement component C3 on the bacteria. SAP inhibited only the activation of the classical complement pathway; the alternative route remained unaffected. In the classical route, SAP prevented the deposition of the first complement component, Clq, probably by interfering with the binding of Clq to LPS. Since antibody-mediated Clq activation was not inhibited by SAP, SAP seems to inhibit only the LPS-induced classical complement pathway activation. The SAP induced inhibition of C3 deposition strongly diminished the complement-mediated lysis as well as the phagocytosis of the bacteria. The binding of SAP to gram negative bacteria, therefore, might influence the pathophysiology of an infection with such bacteria. PMID- 10722561 TI - Comparison of the immune profile of nonhealing cutaneous Leishmaniasis patients with those with active lesions and those who have recovered from infection. AB - Th1-type cellular immune responses play a critical role in protection against infection with Leishmania parasites, whereas activation of Th2-type cells results in progressive disease. Cutaneous leishmaniasis caused by Leishmania major is often a self-healing disease; however, persistent nonhealing forms are also known. In the present study, we have described cell-mediated immune responses in nonhealing patients by measuring T-cell proliferation, cytokine production, and phenotypic characterization of these cells. The responses were compared with those of patients with active lesions, patients who had recovered from infection, and healthy controls. Peripheral blood mononuclear cells from patients with active lesions and recovered donors proliferated vigorously and produced Th1-type cytokine when stimulated with L. major antigens, whereas in nonhealing patients the proliferative responses were significantly lower and showed a Th2-type response to Leishmania antigens. Interleukin-10 (IL-10) production was not a feature of L. major stimulation. Flow cytometric analysis revealed that L. major antigen induced proliferation of the CD4-positive population and that these cells were the major source of gamma interferon and IL-4. These results show a distinct dichotomy in the cytokine response to L. major infection. PMID- 10722562 TI - Altered gene expression in Staphylococcus aureus upon interaction with human endothelial cells. AB - Staphylococcus aureus is isolated from a substantial number of patients with infective endocarditis who are not known to have predisposing heart abnormalities. It has been suggested that the infection is initiated by the direct binding of S. aureus to human vascular endothelium. To determine the mutual response of the endothelial cells and the bacteria, we studied the interaction between S. aureus and human vascular endothelium. Scanning electron microscopic analyses showed that binding of S. aureus to human umbilical vein endothelial cells (HUVEC) mainly occurred via thread-like protrusions extending from the cell surface. Bound bacteria appeared to be internalized via retraction of the protrusions into newly formed invaginations of the endothelial cell surface. The growth phase of S. aureus had a major impact on the interaction with HUVEC. Logarithmically growing bacteria showed increased binding to, and were more readily internalized by, HUVEC compared to stationary-phase bacteria. To assess the bacterial response to the cellular environment, an expression library of S. aureus was used to identify genes whose expression was induced after 4 h of exposure to HUVEC. The identified genes could be divided into different categories based on the functions of the encoded proteins (transport, catabolism, biosynthesis, and DNA repair). Further analyses of five of the S. aureus transposon clones showed that HUVEC as well as human serum are stimuli for triggering gene expression in S. aureus. PMID- 10722563 TI - Interleukin-4 receptor alpha-deficient BALB/c mice show an unimpaired T helper 2 polarization in response to Leishmania major infection. AB - We recently generated interleukin-4 (IL-4) receptor alpha-deficient (IL-4Ralpha( /-)) BALB/c mice and showed evidence for a protective role of IL-13-mediated functions in leishmaniasis. In this study, we investigated the IL-4 expression and T helper 2 (Th2) development in Leishmania major-infected IL-4Ralpha(-/-) mice. Here we show that the early burst of IL-4 expression observed in L. major infected BALB/c mice is independent of IL-4Ralpha-mediated functions. Subsequently, we confirmed an impaired Th2 development in vitro. Unexpectedly, during L. major infection, isolated CD4(+) IL-4Ralpha(-/-) T cells expressed high IL-4- but low gamma interferon (IFN-gamma)-specific mRNA, comparable to Th2 polarized BALB/c CD4(+) cells and in contrast to Th1-polarized C57BL/6 CD4(+) cells. Since antigen-specific restimulated popliteal lymph node cells (PLN) of IL 4Ralpha(-/-) mice also responded with high IL-4 but low IFN-gamma production, comparable to Th2-polarized cells from wild-type BALB/c mice and in contrast to Th1-polarized C57BL/6 cells, these results suggested an unimpaired Th2 polarization during an established infection with L. major. To further define the observed IL-4 receptor-independent Th2 cell phenotype, we determined an independent Th2 marker, the IL-12 receptor beta-2 (IL-12Rbeta2)-specific transcript levels of CD4(+) T cells. Confirming Th2 polarization in L. major infected IL-4Ralpha(-/-) mice, comparable IL-12Rbeta2 message levels between CD4(+) T cells from infected IL-4Ralpha(-/-) mice and Th2 cells from BALB/c mice were found, whereas Th1-polarized C57BL/6 cells showed strikingly increased IL 12Rbeta2 expression levels. These results indicate that signals mediated by the IL-4Ralpha are not necessary to induce and sustain an efficient IL-4 expression and Th2 polarization in L. major-infected BALB/c mice and suggest that IL-4Ralpha independent mechanisms underlie the default Th2 development in L. major-infected BALB/c mice. PMID- 10722564 TI - Role of toxin functional domains in anthrax pathogenesis. AB - We investigated the role of the functional domains of anthrax toxins during infection. Three proteins produced by Bacillus anthracis, the protective antigen (PA), the lethal factor (LF), and the edema factor (EF), combine in pairs to produce the lethal (PA+LF) and edema (PA+EF) toxins. A genetic strategy was developed to introduce by allelic exchange specific point mutations or in-frame deletions into B. anthracis toxin genes, thereby impairing either LF metalloprotease or EF adenylate cyclase activity or PA functional domains. In vivo effects of toxin mutations were analyzed in an experimental infection of mice. A tight correlation was observed between the properties of anthrax toxins delivered in vivo and their in vitro activities. The synergic effects of the lethal and edema toxins resulted purely from their enzymatic activities, suggesting that in vivo these toxins may act together. The PA-dependent antibody response to LF induced by immunization with live B. anthracis was used to follow the in vivo interaction of LF and PA. We found that the binding of LF to PA in vivo was necessary and sufficient for a strong antibody response against LF, whereas neither LF activity nor binding of lethal toxin complex to the cell surface was required. Mutant PA proteins were cleaved in mice sera. Thus, our data provide evidence that, during anthrax infection, PA may interact with LF before binding to the cell receptor. Immunoprotection studies indicated that the strain producing detoxified LF and EF, isogenic to the current live vaccine Sterne strain, is a safe candidate for use as a vaccine against anthrax. PMID- 10722565 TI - Cable-piliated Burkholderia cepacia binds to cytokeratin 13 of epithelial cells. AB - Although the Burkholderia cepacia complex consists of several genomovars, one highly transmissible strain of B. cepacia has been isolated from the sputa of cystic fibrosis (CF) patients throughout the United Kingdom and Canada. This strain expresses surface cable (Cbl) pili and is thought to be the major strain associated with the fatal "cepacia syndrome." In the present report we characterize the specific 55-kDa buccal epithelial cell (BEC) protein that binds cable pilus-positive B. cepacia. N-terminal sequences of CNBr-generated internal peptides identified the protein as cytokeratin 13 (CK13). Western blots of BEC extracts probed with a specific monoclonal antibody to CK13 confirmed the identification. Mixed epidermal cytokeratins (which contain CK13), cytokeratin extract from BEC (which consists essentially of CK13 and CK4), and a polyclonal antibody to mixed cytokeratins inhibited B. cepacia binding to CK13 blots and to normal human bronchial epithelial (NHBE) cells. Preabsorption of the antikeratin antibody with the BEC cytokeratin fraction reversed the inhibitory effect of the antibody. A cytokeratin mixture lacking CK13 was ineffective as an inhibitor of binding. Colocalization of CK13 and B. cepacia by confocal microscopy demonstrated that intact nonpermeabilized NHBE cells express small amounts of surface CK13 and bind Cbl-positive B. cepacia in the same location. Binding to intact NHBE cells was dependent on bacterial concentration and was saturable, whereas a Cbl-negative isolate exhibited negligible binding. These findings raise the possibility that surface-accessible CK13 in respiratory epithelia may be a biologically relevant target for the binding of cable piliated B. cepacia. PMID- 10722566 TI - Monoclonal antibody-mediated modulation of the humoral immune response against mucosally applied Streptococcus mutans. AB - Systemic immunization with antigen coupled to monoclonal antibody (MAb) has been used by several investigators to increase the number of MAb-producing hybridomas against an antigen and to elicit antibodies specific for poorly immunogenic epitopes. This strategy has implications for vaccine design in that protective immunity is not necessarily directed at immunodominant epitopes of pathogens and may be improved by deliberately shifting the immune response toward subdominant epitopes. To our knowledge, no studies to date have addressed the potential for immunomodulatory activity mediated by MAbs bound to mucosally applied antigen. To test whether administration of an exogenous MAb directed against a streptococcal surface protein could influence the humoral immune response, BALB/c mice were immunized orally by gastric intubation or intranasally with Streptococcus mutans alone or S. mutans complexed with a MAb directed against the major surface protein P1. Significant changes in the subclass distribution, as well as the specificity, of anti-P1 serum immunoglobulin G antibodies were demonstrated in groups of mice which received S. mutans coated with the anti-P1 MAb versus those which received S. mutans alone. Alterations in the humoral immune response were dependent on the amount of anti-P1 MAb used to coat the bacteria. In addition, differences in the anti-P1 immune responses were observed between groups of mice immunized via oral versus intranasal routes. In summary, an exogenous MAb complexed with a streptococcal antigen prior to mucosal immunization can influence the immunoglobulin isotype and specificity of the host humoral immune response against the antigen. PMID- 10722567 TI - Activation of intercellular adhesion molecule 1 expression by Helicobacter pylori is regulated by NF-kappaB in gastric epithelial cancer cells. AB - Interactions between leukocytes and epithelial cells may play a key role in Helicobacter pylori-associated gastric mucosal inflammation. This process is mediated by various cell adhesion molecules. The present study examined the molecular mechanisms leading to H. pylori-induced epithelial cell intercellular adhesion molecule-1 (ICAM-1; also called CD54) expression. Coculture of epithelial cells with cytotoxin-associated gene pathogenicity island-positive (cag PAI(+)) H. pylori strains, but not with a cag PAI(-) strain or H. pylori culture supernatants, resulted in upregulation of steady-state mRNA levels and cell surface expression of ICAM-1. Coculture with H. pylori induced an increase in luciferase activity in cells which were transfected with a luciferase reporter gene linked to the 5'-flanking region of the ICAM-1 gene. H. pylori activated the ICAM-1 promoter via the NF-kappaB binding site. An inducible nuclear protein complex bound to the ICAM-1 NF-kappaB site and was identified as the NF-kappaB p50-p65 heterodimer. H. pylori induced the degradation of IkappaB-alpha, a major cytoplasmic inhibitor of NF-kappaB, and stimulated the expression of IkappaB alpha mRNA. Pretreatment of epithelial cells with pyrrolidine dithiocarbamate, which blocks NF-kappaB activation, inhibited H. pylori-induced ICAM-1 expression. THP-1 macrophagic cells, peripheral blood mononuclear cells, and purified neutrophils adhered to H. pylori-infected epithelial cells to a greater extent than to uninfected cells. These results show that H. pylori directly induces expression of ICAM-1 on gastric epithelial cells in an NF-kappaB-dependent manner that may support leukocyte attachment during inflammation. PMID- 10722568 TI - Peripheral cytokine responses to Trichuris muris reflect those occurring locally at the site of infection. AB - The study of human cellular immune responses to parasite infection under field conditions is very complex. Often, the only practical site from which to sample the cellular responses is the peripheral blood. Sampling peripheral blood lymphocytes (PBL) relies on the assumption that these peripheral responses accurately reflect the immune responses acting locally at the site of infection. This is a particularly important point for the human intestinal helminth Trichuris trichiura, which solely inhabits the cecum and large intestine and so will stimulate a localized immune response. Using the well-defined model of T. trichiura, T. muris in the mouse, we have demonstrated that the dominant cytokine responses of the mesenteric lymph nodes (MLN) can be detected by sampling PBL. Resistant mice which mount a type 2 cytokine response in their MLN had PBL producing interleukin-4 (IL-4), IL-5, and IL-9, with negligible levels of gamma interferon (IFN-gamma). Conversely, susceptible mice which mount a type 1 cytokine response in their MLN had PBL producing IFN-gamma and negligible levels of type 2 cytokines. We have also shown that the PBL are capable of mounting a functional immune response against T. muris. PBL from immune mice were capable of transferring immunity to T. muris-infected severe combined immunodeficient (C.B 17 scid/scid) mice. Sampling PBL responses is therefore a viable option for monitoring human intestinal immune responses during T. trichiura infection in the field. PMID- 10722569 TI - Production of protective human antipneumococcal antibodies by transgenic mice with human immunoglobulin loci. AB - Infections with Streptococcus pneumoniae remain a significant cause of morbidity and mortality. To gain insight into structure-function relationships for human antibodies to pneumococcal capsular polysaccharide (PPS), we studied the response of transgenic mice reconstituted with human immunoglobulin loci, XenoMouse, to PPS antigens in a pneumococcal vaccine. Enzyme-linked immunosorbent assays of sera from mice vaccinated with a 23-valent pneumococcal vaccine revealed that they produced serotype-specific human antibodies, with the greatest response being to the PPS of serotype 3 (PPS 3). Molecular sequence analysis of three monoclonal antibodies (MAbs) to PPS 3 generated from lymphoid cells from mice vaccinated with a 23-valent pneumococcal vaccine or a PPS 3-bovine serum albumin conjugate revealed that they all used heavy-chain immunoglobulin genes from the V(H)3 family, two expressed light chain genes from the human Vkappa1 family, and one expressed a mouse lambda light chain. The protective efficacy of the two MAbs was examined in mice. A 10-microgram dose of both, and a 1-microgram dose of one, significantly prolonged survival from a lethal serotype 3 infection in CBA/N mice. Our data show that XenoMouse mice produced protective, serotype-specific human antibodies to PPS 3, and they lend support to the proposal that these animals represent a useful model to study the human antibody response to PPS antigens. PMID- 10722570 TI - Gamma interferon dominates the murine cytokine response to the agent of human granulocytic ehrlichiosis and helps to control the degree of early rickettsemia. AB - The cytokine response to the agent of human granulocytic ehrlichiosis (HGE) was assessed in a murine infection model and the role of gamma interferon (IFN gamma), a cytokine that is crucial for host defenses against intracellular pathogens, was investigated by using IFN-gamma-deficient mice. The agent of HGE (aoHGE) is an obligate intracellular bacterium that survives within neutrophils: morulae (vacuoles containing HGE organisms) are evident in polymorphonuclear leukocytes of experimentally infected immunocompetent mice for 1 to 2 weeks. We now show that IFN-gamma levels increase during early infection of C3H/HeN or C57BL/6 mice with HGE bacteria. Moreover, in response to aoHGE extracts or concanavalin A, splenocytes from ehrlichia-infected mice produced more IFN-gamma and less interleukin-4 than controls, suggesting that aoHGE partially skewed the immune response towards a Th1 phenotype. Absolute concentration of morulae containing neutrophils in blood was 122 +/- 22 cells/microliter on day 8. The bacterial DNA burden was also highest on day 8 and then declined after IFN-gamma levels peaked. In contrast, IFN-gamma-deficient mice had a markedly elevated HGE bacteria burden with morulae concentration of 282 +/- 48 cells/microliter on day 5 (P = 0.004) and 242 +/- 63 cells/microliter on day 8 (P = 0.005). Rickettsemia resolved in immunocompetent and IFN-gamma deficient mice after 2 weeks, while both the immunocompetent and the IFN-gamma-deficient mice had increased serum antibodies against aoHGE antigens at this time point. These data demonstrate that the HGE agent elicits a prominent IFN-gamma response in mice and that IFN-gamma is important in controlling the degree of rickettsemia during the early phase of infection, while IFN-gamma independent mechanisms play a role at later time points. PMID- 10722571 TI - Impact of siderophore production on Pseudomonas aeruginosa infections in immunosuppressed mice. AB - Pseudomonas aeruginosa produces siderophores, pyoverdin and pyochelin, for high affinity iron uptake. To investigate their contribution to P. aeruginosa infections, we constructed allelic exchange mutants from strain PAO1 which were deficient in producing one or both of the siderophores. When inoculated into the calf muscles of immunosuppressed mice, pyochelin-deficient and pyoverdin deficient mutants grew and killed the animals as efficiently as PAO1. In contrast, the pyochelin- and pyoverdin-deficient (double) mutant did not show lethal virulence, although it did infect the muscles. On the other hand, when inoculated intranasally, all mutants grew in the lungs and killed immunosuppressed mice. Compared with PAO1, however, the pyoverdin-deficient mutant and the double mutant grew poorly in the lungs, and the latter was significantly attenuated for virulence. Irrespective of the inoculation route, the pyoverdin-deficient and doubly deficient mutants detected in the blood were significantly less numerous than PAO1. Additionally, in vitro examination demonstrated that the growth of the double mutant was extremely reduced under a free-iron-restricted condition with apotransferrin but that the growth reduction was completely canceled by supplementation with hemoglobin as a heme source. These results suggest that both pyoverdin and pyochelin are required for efficient bacterial growth and full expression of virulence in P. aeruginosa infection, although pyoverdin may be comparatively more important for bacterial growth and dissemination. However, the siderophores were not always required for infection. It is possible that non-siderophore-mediated iron acquisition, such as via heme uptake, might also play an important role in P. aeruginosa infections. PMID- 10722572 TI - Enhanced hematopoietic activity accompanies parasite expansion in the spleen and bone marrow of mice infected with Leishmania donovani. AB - In this study, we have analyzed hematopoietic activity in the spleen, bone marrow, and blood of BALB/c and scid mice infected with Leishmania donovani. Our analysis demonstrates that infection induces a rapid but transient mobilization of progenitor cells into the circulation, associated with elevated levels of granulocyte/macrophage colony-stimulating factor (GM-CSF) and MIP-1alpha. From 14 to 28 days postinfection, when parasite expansion begins in the spleen and bone marrow, both the frequency and cell cycle activity of hematopoietic progenitors, particulary CFU-granulocyte, monocyte, are dramatically increased in these organs. This is associated with increased accumulation of mRNA for GM-CSF, M-CSF, and G-CSF, but not interleukin-3. Our data also illustrate that hematopoietic activity, as assessed by changes in the frequency of progenitor cell populations and their levels of cell cycle activity, can be regulated in both a T-cell independent and T-cell-dependent, as well as in an organ-specific, manner. Collectively, these data add to our knowledge of the long-term changes which occur in organs in which L. donovani is able to persist. PMID- 10722573 TI - Cloning, sequencing, and role in serum susceptibility of porin II from mesophilic Aeromonas hydrophila. AB - We cloned and sequenced the structural gene for Aeromonas hydrophila porin II from strain AH-3 (serogroup O:34). The genetic position of this gene, like that of ompF in Escherichia coli, is adjacent to aspC and transcribed in the same direction. However, upstream of the porin II gene no similarities with E. coli were found. We obtained defined insertion mutants in porin II gene either in A. hydrophila (O:34) or A. veronii sobria (serogroup O:11) serum-resistant or sensitive strains. Furthermore, we complemented these mutants with a plasmid harboring only the porin II gene, which allowed us to define the role of porin II as an important surface molecule involved in serum susceptibility and C1q binding in these strains. PMID- 10722574 TI - Secreted enzymatic activities of wild-type and pilD-deficient Legionella pneumophila. AB - Legionella pneumophila, the agent of Legionnaires' disease, is an intracellular pathogen of protozoa and macrophages. Previously, we had determined that the Legionella pilD gene is involved in type IV pilus biogenesis, type II protein secretion, intracellular infection, and virulence. Since the loss of pili and a protease do not account for the infection defect exhibited by a pilD-deficient strain, we sought to define other secreted proteins absent in the mutant. Based upon the release of p-nitrophenol (pNP) from p-nitrophenyl phosphate, acid phosphatase activity was detected in wild-type but not in pilD mutant supernatants. Mutant supernatants also did not release either pNP from p nitrophenyl caprylate and palmitate or free fatty acid from 1 monopalmitoylglycerol, suggesting that they lack a lipase-like activity. However, since wild-type samples failed to release free fatty acids from 1,2 dipalmitoylglycerol or to cleave a triglyceride derivative, this secreted activity should be viewed as an esterase-monoacylglycerol lipase. The mutant supernatants were defective for both release of free fatty acids from phosphatidylcholine and degradation of RNA, indicating that PilD-negative bacteria lack a secreted phospholipase A (PLA) and nuclease. Finally, wild-type but not mutant supernatants liberated pNP from p-nitrophenylphosphorylcholine (pNPPC). Characterization of a new set of mutants defective for pNPPC-hydrolysis indicated that this wild-type activity is due to a novel enzyme, as opposed to a PLC or another known enzyme. Some, but not all, of these mutants were greatly impaired for intracellular infection, suggesting that a second regulator or processor of the pNPPC hydrolase is critical for L. pneumophila virulence. PMID- 10722575 TI - Identification of a fibronectin binding protein from Streptococcus mutans. AB - The interaction of viridans streptococci with components of the extracellular matrix (ECM) plays an important role in the pathogenesis of infective endocarditis. We have identified a surface protein of Streptococcus mutans which binds the ECM constituent fibronectin (Fn). Initially, we found that S. mutans could adsorb soluble Fn in plasma, but with lower efficiency than Streptococcus pyogenes. In addition, S. mutans could bind immobilized Fn in a dose-dependent manner when tested using an enzyme-linked immunosorbent assay. Crude extracts of cell wall-associated proteins or extracellular proteins from S. mutans MT8148 specifically bound Fn through a protein with the molecular mass of ca. 130 kDa, as detected by far-Western immunoblotting. The candidate Fn binding protein (FBP 130) was purified to near homogeneity by using Fn coupled Sepharose 4B affinity column chromatography. A rabbit polyclonal antibody against FBP-130 reacted specifically with a protein of molecular mass of ca. 130 kDa in both cell wall and extracellular fractions, and the abundance of FBP was higher in the former than in the latter fractions. The purified FBP bound specifically to immobilized Fn, whereas the binding of soluble Fn to coated FBP could only be detected in the presence of high concentrations of Fn. The purified FBP, as well as anti-FBP immunoglobulin G, inhibited the adherence of S. mutans to immobilized Fn and endothelial cells (ECV304) in a dose-dependent manner. These results demonstrated that FBP-130 mediated the adherence of S. mutans specifically to Fn and endothelial cells in vitro. The characteristics of S. mutans and FBP-130 in binding Fn confirmed that viridans streptococci adopt different strategies in their interaction with ECM. PMID- 10722577 TI - Human dendritic cells are superior to B cells at presenting a major histocompatibility complex class II-restricted heterologous antigen expressed on recombinant Streptococcus gordonii. AB - Bacteria are being actively investigated as vaccine carriers for inducing or boosting protective immune responses. In this study, human monocyte-derived dendritic cells (DCs) and normal B cells were compared for their capacity to present the C fragment of tetanus toxin (TTFC), expressed on the surface of recombinant Streptococcus gordonii, to specific CD4(+) T lymphocytes. DCs were more efficient than B cells at presenting soluble TTFC and remarkably more capable of presenting bacterium-associated TTFC both in terms of the amount of antigen required to obtain a given T-cell response and on a per-cell basis. This difference was associated with a much lower capacity of B cells to endocytose soluble TTFC and phagocytose recombinant S. gordonii. In addition, S. gordonii induced the phenotypic maturation of DCs but not of B cells. The results thus indicate that DCs but not B cells play a crucial role in the amplification of class II-restricted immune responses induced by immunization with recombinant gram-positive bacteria. PMID- 10722576 TI - Functional activities and immunoglobulin variable regions of human and murine monoclonal antibodies specific for the P1.7 PorA protein loop of Neisseria meningitidis. AB - The meningococcal PorA protein is considered a promising vaccine candidate. Although much is understood regarding the structure of PorA proteins, little is known about the structure-function relationships of PorA antibodies. The aim of this study was to compare the functional and molecular characteristics of a human monoclonal antibody (MAb) and three murine MAbs specific for the PorA P1.7 serosubtype. Murine MAbs 207,B-4 (immunoglobulin G2a [IgG2a]) and MN14C11.6 (IgG2a) were both bactericidal and opsonophagocytic for P1.7-expressing meningococci, whereas human MAb SS269 (IgG3) and murine MAb 208,D-5 (IgA) initiated neither effector function. Epitope mapping with synthetic peptides revealed that MAbs 207,B-4 and 208,D-5 recognized the sequence ASGQ, which is the same specificity motif that a previous study had established for SS269 and MN14C11.6. Nucleotide and amino acid sequence analyses of the variable regions of the four MAbs showed that the SS269 V(H) region belonged to the VH3 family and was approximately 70% homologous to those of the murine MAbs which were all from the 7183 family, whereas the SS269 V(L) region belonged to the Vlambda1-b family and was less than 40% homologous to those of the murine MAbs which were all members of the Vkappa1 family. The Fab fragment of SS269 was cloned and expressed in Escherichia coli and was shown by enzyme-linked immunosorbent assay analyses to bind as well as intact SS269 MAb to P1.7,16 serosubtype group B strain 44/76. We conclude that distinct differences exist in the effector function activities and variable region gene sequences of human and murine P1.7-specific MAbs despite their recognition of similar epitopes. PMID- 10722578 TI - Identification of a Treponema denticola OppA homologue that binds host proteins present in the subgingival environment. AB - Proteins secreted or exported by Treponema denticola have been implicated as mediators of specific interactions between the spirochete and subgingival tissues in periodontal diseases. However, limited information is available on the ability of this peptidolytic organism to bind or transport soluble peptides present in the subgingival environment. A prominent 70-kDa protein was isolated from surface extracts of T. denticola ATCC 35405. A clone expressing a portion of the protein was identified in an Escherichia coli expression library of T. denticola DNA. DNA sequence analysis showed that the cloned gene encoded a peptide homologous to OppA, the solute binding protein of an ATP-binding cassette-type peptide transporter involved in peptide uptake and environmental signaling in a wide range of bacteria. Genes encoding OppB, -C, -D, and -F were identified directly downstream of oppA in T. denticola. OppA was present in representative strains of T. denticola and in Treponema vincentii but was not detected in Treponema pectinovorum or Treponema socranskii. Immunogold electron microscopy suggested that OppA was accessible to proteins at the surface of the spirochete. Native OppA bound soluble plasminogen and fibronectin but did not bind to immobilized substrates or epithelial cells. A T. denticola oppA mutant bound reduced amounts of soluble plasminogen, and plasminogen binding to the parent strain was inhibited by the lysine analog epsilon-aminocaproic acid. Binding of soluble host proteins by OppA may be important both for spirochete-host interactions in the subgingival environment and for uptake of peptide nutrients. PMID- 10722579 TI - Induction of apoptotic cell death in peripheral blood mononuclear and polymorphonuclear cells by an oral bacterium, Fusobacterium nucleatum. AB - It is largely unknown why a variety of bacteria present in the oral cavity are capable of establishing themselves in the periodontal pockets of nonimmunocompromised individuals in the presence of competent immune effector cells. In this paper we present evidence for the immunosuppressive role of Fusobacterium nucleatum, a gram-negative oral bacterium which plays an important role in the generation of periodontal disease. Our studies indicate that the immunosuppressive role of F. nucleatum is largely due to the ability of this organism to induce apoptotic cell death in peripheral blood mononuclear cells (PBMCs) and in polymorphonuclear cells (PMNs). F. nucleatum treatment induced apoptosis of PBMCs and PMNs as assessed by an increase in subdiploid DNA content determined by DNA fragmentation and terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end-labeling assays. The ability of F. nucleatum to induce apoptosis was abolished by either heat treatment or proteinase digestion but was retained after formaldehyde treatment, suggesting that a heat-labile surface protein component is responsible for bacterium-mediated cell apoptosis. The data also indicated that F. nucleatum-induced cell apoptosis requires activation of caspases and is protected by NF-kappaB. Possible mechanisms of F. nucleatum's role in the pathogenesis of periodontal disease are discussed. PMID- 10722580 TI - Comparable endotoxic properties of lipopolysaccharides are manifest in diverse clinical isolates of gram-negative bacteria. AB - In general there is a poor correlation between serum lipopolysaccharide (LPS; the biologically active constituent of endotoxin) levels and mortality in septic patients. The objective of this study was to determine if chemical, structural, or biological differences among LPS from different clinical isolates of gram negative bacteria might explain this discrepancy. LPS preparations were made using the hot phenol-water extraction method from eight clinical isolates of gram negative bacteria. As a percentage of the total weight of the LPS, the phosphate content ranged from 3.0 to 13.8% (average, 6.7 +/- 3.6%), and the 2-keto-3 deoxyoctonate content ranged from 1.9 to 27.4% (average, 8.9 +/- 8.5%). These values were not dissimilar to those obtained for a reference endotoxin. In a standard measure of LPS activity, the Limulus amoebocyte lysate assay, there was approximately a twofold difference between the least and most active preparations. The two preparations with the greatest difference in their ability to elicit the secretion of tumor necrosis factor alpha from a mouse peritoneal macrophage cell line were similar in lethality when administered to mice sensitized to the effects of LPS by D(+)-galactosamine. These relatively minor differences in LPS activity seem unlikely to explain the generally observed discrepancy between serum endotoxin levels and mortality in patients with gram negative sepsis. PMID- 10722581 TI - Immunity to onchocerciasis: cells from putatively immune individuals produce enhanced levels of interleukin-5, gamma interferon, and granulocyte-macrophage colony-stimulating factor in response to Onchocerca volvulus larval and male worm antigens. AB - Antigen-specific interleukin-5 (IL-5), gamma interferon (IFN-gamma), and granulocyte-macrophage colony-stimulating factor (GM-CSF) responses in individuals living in an area of hyperendemicity for onchocerciasis in Cameroon were examined. The responses against antigens prepared from Onchocerca volvulus third-stage larvae (L3), molting L3 (mL3), and crude extract from adult males (M OvAg) were compared to the responses against antigens from adult female worms and skin microfilariae. Cytokine responses for the putatively immune individuals (PI) and the infected individuals (INF) were compared. A differential cytokine profile of IL-5 (Th2 phenotype) and IFN-gamma (Th1 phenotype) was found in these individuals in response to the antigens. In both the PI and the INF, Th2 responses against all the antigens tested were dominant. However, in the PI group as a whole, there was an enhanced Th2 response against the larval antigens and the adult male and adult female antigens, and a Th1 response in a subgroup of the PI (27 to 54.5%) against L3, mL3, and M-OvAg antigens was present. While the PI produced significantly higher levels of GM-CSF against L3, mL3, and M-OvAg antigens than the INF, there was no difference in the GM-CSF responses of the groups against the other antigens. The present study indicated that, in comparison to the INF, the PI have distinct larva-specific and adult male specific cytokine responses, thus supporting the premise that immunological studies of the PI would lead to the identification of immune mechanisms and the target genes that play a role in protective immunity. PMID- 10722582 TI - Antibodies to tyvelose exhibit multiple modes of interference with the epithelial niche of Trichinella spiralis. AB - Infection with the parasitic nematode Trichinella spiralis is initiated when the L1 larva invades host intestinal epithelial cells. Monoclonal antibodies specific for glycans on the larval surface and secreted glycoproteins protect the intestine against infection. Protective antibodies recognize tyvelose which caps the target glycan. In this study, we used an in vitro model of invasion to further examine the mechanism(s) by which tyvelose-specific antibodies protect epithelial cells against T. spiralis. Using cell lines that vary in susceptibility to invasion, we confirmed and clarified the results of our in vivo studies by documenting three modes of interference: exclusion of larvae from cells, encumbrance of larvae as they migrated within epithelial monolayers, and inhibition of parasite development. Excluded larvae bear cephalic caps (C. S. McVay et al., Infect. Immun. 66:1941-1945, 1998) of immune complexes that may physically block invasion or may interfere with sensory reception. Monovalent Fab fragments prepared from a tyvelose-specific antibody also excluded larvae from cells, demonstrating that antibody binding can inhibit the parasite in the absence of antigen aggregation and cap formation. In contrast, encumbered larvae caused extensive damage to the monolayer yet were not successful in establishing a niche, as reflected by their failure to molt. These results show that antibodies to tyvelose exhibit multiple modes of inhibitory activity, further implicating tyvelose-bearing glycoproteins as mediators of invasion and niche establishment by T. spiralis. PMID- 10722583 TI - Bordetella pertussis TonB, a Bvg-independent virulence determinant. AB - In gram-negative bacteria, high-affinity iron uptake requires the TonB/ExbB/ExbD envelope complex to release iron chelates from their specific outer membrane receptors into the periplasm. Based on sequence similarities, the Bordetella pertussis tonB exbB exbD locus was identified on a cloned DNA fragment. The tight organization of the three genes suggests that they are cotranscribed. A putative Fur-binding sequence located upstream from tonB was detected in a Fur titration assay, indicating that the tonB exbB exbD operon may be Fur-repressed in high iron growth conditions. Putative structural genes of the beta-subunit of the histone-like protein HU and of a new two-component regulatory system were identified upstream from tonB and downstream from exbD, respectively. A B. pertussis DeltatonB exbB::Km(r) mutant was constructed by allelic exchange and characterized. The mutant was impaired for growth in low-iron medium in vitro and could not use ferrichrome, desferal, or hemin as iron sources. Levels of production of the major bacterial toxins and adhesins were similar in the TonB(+)/TonB(-) pair. The DeltatonB exbB mutant was still responsive to chemical modulators of virulence; thus, the BvgA/BvgS two-component system is not TonB dependent. Nevertheless, in vivo in the mouse respiratory infection model, the colonization ability of the mutant was reduced compared to the parental strain. PMID- 10722584 TI - Cell vacuolation, a manifestation of the El tor hemolysin of Vibrio cholerae. AB - Culture supernatants of nontoxigenic nonepidemic clinical strains of Vibrio cholerae belonging to diverse serogroups were found to induce vacuolation of nonconfluent HeLa cells. The vacuoles became prominent 18 h after introduction of culture supernatant, and vacuolated cells survived for 48 h and then died. Only a fraction of the vacuolated cells took up neutral red dye, implying that there were differences in the vacuolar microenvironment. Further tests showed that the factor responsible for vacuolation was heat labile and proteinaceous. Vacuolating activity was completely neutralized by antibody to hemolysin of V. cholerae but not by antibody to vacuolating cytotoxin of Helicobacter pylori. Partial purification of the vacuolating factor led to elution of fractions, which showed both hemolytic and vacuolating activity. PCR amplification and cloning of the hemolysin structural gene (hlyA) into Escherichia coli DH5alpha led to isolation of clones producing cell vacuolating factor in a cell-associated form. Further, a null insertion mutation in the hlyA gene of a high-vacuolating-factor-producing strain led to complete abolition of both cell vacuolating and hemolytic activities. These analyses establish vacuolation as a potentially important but previously unrecognized property of V. cholerae El Tor hemolysin. PMID- 10722585 TI - Role of adhesins and toxins in invasion of human tracheal epithelial cells by Bordetella pertussis. AB - Bordetella pertussis, the agent of whooping cough, can invade and survive in several types of eukaryotic cell, including CHO, HeLa 229, and HEp-2 cells and macrophages. In this study, we analyzed bacterial invasiveness in nonrespiratory human HeLa epithelial cells and human HTE and HAE0 tracheal epithelial cells. Invasion assays and transmission electron microscopy analysis showed that B. pertussis strains invaded and survived, without multiplying, in HTE or HAE0 cells. This phenomenon was bvg regulated, but invasive properties differed between B. pertussis strains and isolates and the B. pertussis reference strain. Studies with B. pertussis mutant strains demonstrated that filamentous hemagglutinin, the major adhesin, was involved in the invasion of human tracheal epithelial cells by bacteria but not in that of HeLa cells. Fimbriae and pertussis toxin were not found to be involved. However, we found that the production of adenylate cyclase-hemolysin prevents the invasion of HeLa and HTE cells by B. pertussis because an adenylate cyclase-hemolysin-deficient mutant was found to be more invasive than the parental strain. The effect of adenylate cyclase-hemolysin was mediated by an increase in the cyclic AMP concentration in the cells. Pertactin (PRN), an adhesin, significantly inhibited the invasion of HTE cells by bacteria, probably via its interaction with adenylate cyclase hemolysin. Isolates producing different PRNs were taken up similarly, indicating that the differences in the sequences of the PRNs produced by these isolates do not affect invasion. We concluded that filamentous hemagglutinin production favored invasion of human tracheal cells but that adenylate cyclase-hemolysin and PRN production significantly inhibited this process. PMID- 10722586 TI - Predictive value of nuclear factor kappaB activity and plasma cytokine levels in patients with sepsis. AB - The relationship between fluctuating cytokine concentrations in plasma and the outcome of sepsis is complex. We postulated that early measurement of the activation of nuclear factor kappaB (NF-kappaB), a transcriptional regulatory protein involved in proinflammatory cytokine expression, may help to predict the outcome of sepsis. We determined NF-kappaB activation in peripheral blood mononuclear cells of 34 patients with severe sepsis (23 survivors and 11 nonsurvivors) and serial concentrations of inflammatory cytokines (interleukin-6, interleukin-1, and tumor necrosis factor) and various endogenous antagonists in plasma. NF-kappaB activity was significantly higher in nonsurvivors and correlated strongly with the severity of illness (APACHE II score), although neither was related to the cytokine levels. Apart from NF-kappaB activity, the interleukin-1 receptor antagonist was the only cytokine tested whose level in plasma was of value in predicting mortality by logistic regression analysis. These results underscore the prognostic value of early measurement of NF-kappaB activity in patients with severe sepsis. PMID- 10722587 TI - Expression of polymorphic msp1beta genes during acute anaplasma Marginale rickettsemia. AB - Immunization of cattle with native MSP1 induces protection against Anaplasma marginale. The native immunogen is composed of a single MSP1a protein and multiple, undefined MSP1b polypeptides. In addition to the originally sequenced gene, designated msp1beta(F1), we identified three complete msp1beta genes in the Florida strain: msp1beta(F2), msp1beta(F3), and msp1beta(F4). Each of these polymorphic genes encodes a structurally unique MSP1b protein, and unique transcripts can be identified during acute A. marginale rickettsemia. The structural polymorphism is clustered in discrete variable regions, and each MSP1b protein results from a unique mosaic of five variable regions. Although each of the MSP1b proteins in the Florida strain contains epitopes recognized by serum antibody induced by protective immunization with the native MSP1 complex, the variable regions also include epitopes expressed by some but not all of the MSP1b proteins. These data support testing recombinant vaccines composed of the multiple antigenically and structurally unique MSP1b proteins combined with MSP1a in order to mimic the efficacy of native MSP1 immunization. PMID- 10722588 TI - Macrophage class A scavenger receptor-mediated phagocytosis of Escherichia coli: role of cell heterogeneity, microbial strain, and culture conditions in vitro. AB - Macrophage class A scavenger receptors (SR-AI and SR-AII) contribute to host defense by binding polyanionic ligands such as lipopolysaccharide and lipoteichoic acid. SR-A knockout (SR-A(-/-)) mice are more susceptible to endotoxic shock and Listeria monocytogenes infection in vivo, possibly due to decreased clearance of lipopolysaccharide and microorganisms, respectively. We have used flow cytometry to analyze the role of SR-A and other scavenger-like receptors in phagocytosis of bacteria in vitro. Chinese hamster ovary cells stably transfected with human SR-A bound Escherichia coli and Staphylococcus aureus but ingested few organisms. Primary human monocyte-derived macrophages (Mphi) bound and ingested E. coli more efficiently, and this was partially but selectively blocked by the general SR inhibitor, poly(I). A specific and selective role for SR-A was shown, since bone marrow culture-derived Mphi from SR A(-/-) mice ingested fewer E. coli organisms than did wild-type cells, while uptake of antibody-opsonized E. coli was unaffected. SR-A-dependent uptake of E. coli varied with the bacterial strain; ingestion of DH5alpha and K1 by SR-A(-/-) Mphi was reduced by 30 to 60% and 70 to 75%, respectively. Phagocytosis and endocytosis via SR-A were markedly down-modulated when Mphi were plated on serum coated tissue culture plastic compared to bacteriologic plastic, where cell adhesion is mediated by SR-A and CR3, respectively. This paper demonstrates that SR-A can bind and ingest bacteria directly, consistent with a role in host defense in vivo, and highlights the importance of the source of the Mphi, bacterial strain, and culture conditions on receptor function in vitro. PMID- 10722589 TI - Antibodies against the Plasmodium falciparum receptor binding domain of EBA-175 block invasion pathways that do not involve sialic acids. AB - The 175-kDa Plasmodium falciparum erythrocyte binding protein (EBA-175) binds to its receptor, sialic acids on glycophorin A. The binding region within EBA-175 is a cysteine-rich region identified as region II. Antibodies against region II block the binding of native EBA-175 to erythrocytes. We identified a P. falciparum strain, FVO, that could not invade erythrocytes devoid of sialic acids due to prior neuraminidase treatment, and in addition, we used a strain, 3D7, that could invade such sialic acid-depleted erythrocytes. We used these two strains to study the capacity of anti-region II antibodies to inhibit FVO and 3D7 parasite development in vitro. Analysis of growth-inhibitory effects of purified FVO anti-region II immunoglobulin G (IgG) with the FVO and 3D7 strains resulted in similar levels of growth inhibition. FVO and 3D7 strains were inhibited between 28 and 56% compared to control IgG. There appeared to be no intracellular growth retardation or killing of either isolate, suggesting that invasion was indeed inhibited. Incubation of recombinant region II with anti-region II IgG reversed the growth inhibition. These results suggest that antibodies against region II can also interfere with merozoite invasion pathways that do not involve sialic acids. The fact that EBA-175 has such a universal and yet susceptible role in erythrocyte invasion clearly supports its inclusion in a multivalent malaria vaccine. PMID- 10722591 TI - Effects of interleukin-4 deprivation and treatment on resistance to Trypanosoma cruzi. AB - Trypanosoma cruzi (Y strain)-infected interleukin-4(-/-) (IL-4(-/-)) mice of strains 129/J, BALB/c, and C57BL/6 showed no significant difference in parasitemia levels or end point mortality rates compared to wild-type (WT) mice. Higher production of gamma interferon (IFN-gamma) by parasite antigen (Ag) stimulated splenocytes was observed only for C57BL/6 IL-4(-/-) mice. Treatment of 129/J WT mice with recombinant IL-4 (rIL-4), rIL-10, anti-IL-4, and/or anti-IL-10 monoclonal antibodies (MAbs) did not modify parasitism. However, WT mice treated with rIL-4 and rIL-10 had markedly increased parasitism and suppressed IFN-gamma synthesis by spleen cells stimulated with parasite Ag, concanavalin A, or anti CD3. Addition of anti-IL-4 MAbs to splenocyte cultures from infected WT 129/J, BALB/c, or C57BL/6 mice failed to modify IFN-gamma synthesis levels; in contrast, IL-10 neutralization increased IFN-gamma production and addition of rIL-4 and/or rIL-10 diminished IFN-gamma synthesis. We conclude that endogenous IL-4 is not a major determinant of susceptibility to Y strain T. cruzi infection but that IL-4 can, in association with IL-10, modulate IFN-gamma production and resistance. PMID- 10722590 TI - Mutations in the extracellular protein secretion pathway genes (eps) interfere with rugose polysaccharide production in and motility of Vibrio cholerae. AB - Vibrio cholerae is the causal organism of the diarrheal disease cholera. The rugose variant of V. cholerae is associated with the secretion of an exopolysaccharide. The rugose polysaccharide has been shown to confer increased resistance to a variety of agents, such as chlorine, bioacids, and oxidative and osmotic stresses. It also promotes biofilm formation, thereby increasing the survival of the bacteria in the aquatic environments. Here we show that the extracellular protein secretion system (gene designated eps) is involved directly or indirectly in the production of rugose polysaccharide. A TnphoA insertion in epsD gene of the eps operon abolished the production of rugose polysaccharide, reduced the secretion of cholera toxin and hemolysin, and resulted in a nonmotile phenotype. We have constructed defined mutations of the epsD and epsE genes that affected these phenotypes and complemented these defects by plasmid clones of the respective wild-type genes. These results suggest a major role for the eps system in pathogenesis and environmental survival of V. cholerae. PMID- 10722592 TI - Characterization of heat-inducible expression and cloning of HtpG (Hsp90 homologue) of Porphyromonas gingivalis. AB - Porphyromonas gingivalis is implicated in the etiology of periodontal disease. Associations between microbial virulence and stress protein expression have been identified in other infections. For example, Hsp90 homologues in several microbial species have been shown to contribute to virulence. We previously reported that P. gingivalis possessed an Hsp90 homologue (HtpG) which cross reacts with human Hsp90. In addition, we found that elevated levels of serum antibody to Hsp90 stress protein in individuals colonized with this microorganism were associated with periodontal health. However, the role of HtpG in P. gingivalis has not been explored. Therefore, we cloned the htpG gene and investigated the characteristics of HtpG localization and expression in P. gingivalis. htpG exists as a single gene of 2,052 bp from which a single message encoding a mature protein of approximately 68 kDa is transcribed. Western blot analysis revealed that the 68-kDa polypeptide was stress inducible and that a major band at 44 kDa and a minor band at 40 kDa were present at constitutive levels. Cellular localization studies revealed that the 44- and 40-kDa species were associated with membrane and vesicle fractions, while the 68-kDa polypeptide was localized to the cytosolic fractions. PMID- 10722593 TI - Purified lipopolysaccharide from Francisella tularensis live vaccine strain (LVS) induces protective immunity against LVS infection that requires B cells and gamma interferon. AB - Previous results have demonstrated that nonspecific protective immunity against lethal Francisella tularensis live vaccine strain (LVS) or Listeria monocytogenes infection can be stimulated either by sublethal infection with bacteria or by treatment with bacterial DNA given 3 days before lethal challenge. Here we characterize the ability of purified lipopolysaccharide (LPS) from F. tularensis LVS to stimulate similar early protective immunity. Treatment of mice with surprisingly small amounts of LVS LPS resulted in very strong and long-lived protection against lethal LVS challenge within 2 to 3 days. Despite this strong protective response, LPS purified from F. tularensis LVS did not activate murine B cells for proliferation or polyclonal immunoglobulin secretion, nor did it activate murine splenocytes for secretion of interleukin-4 (IL-4), IL-6, IL-12, or gamma interferon (IFN-gamma). Immunization of mice with purified LVS LPS induced a weak specific anti-LPS immunoglobulin M (IgM) response and very little IgG; however, infection of mice with LVS bacteria resulted in vigorous IgM and IgG, particularly IgG2a, anti-LPS antibody responses. Studies using various immunodeficient mouse strains, including LPS-hyporesponsive C3H/HeJ mice, muMT(-) (B-cell-deficient) knockout mice, and IFN-gamma-deficient mice, demonstrated that the mechanism of protection does not involve recognition through the Lps(n) gene product; nonetheless, protection was dependent on B cells as well as IFN-gamma. PMID- 10722594 TI - Reduced virulence of HWP1-deficient mutants of Candida albicans and their interactions with host cells. AB - The Candida albicans gene HWP1 encodes a surface protein that is required for normal hyphal development in vitro. We used mutants lacking one or both alleles of HWP1 to investigate the role of this gene in virulence. Mice infected intravenously with the homozygous hwp1 null mutant, CAL3, survived a median of >14 days, whereas mice infected with a control strain containing two functional alleles of HWP1 survived only 3.5 days. After 1 day of infection, all strains produced similar levels of infection in the kidneys, spleen, and blood. However, after 2 and 3 days, there was a significant decrease in the number of organisms in the kidneys of the mice infected with CAL3. This finding suggests that the hwp1 homozygous null mutant is normal in its ability to initiate infection but deficient in its capacity to maintain infection. CAL3 also germinated minimally in the kidneys. The ability of the heterozygous null mutant to germinate and cause mortality in mice was intermediate to CAL3, suggesting a gene dosage effect. To investigate potential mechanisms for the diminished virulence of CAL3, we examined its interactions with endothelial cells and neutrophils in vitro. CAL3 caused less endothelial cell injury than the heterozygous hwp1 mutant. We conclude that the HWP1 gene product is important for both in vivo hyphal development and pathogenicity of C. albicans. Also, the ability to form filaments may be critical for candidal virulence by enabling the fungus to induce cellular injury and maintain a deep-seated infection. PMID- 10722595 TI - Staphylococcal enterotoxin A acts through nitric oxide synthase mechanisms in human peripheral blood mononuclear cells to stimulate synthesis of pyrogenic cytokines. AB - The pyrogenic response to supernatant fluids obtained from human peripheral blood mononuclear cells (PBMC) stimulated with staphylococcal enterotoxin A (SEA) was characteristic of a response to an endogenous pyrogen in that it was brief and monophasic and was destroyed by heating supernatant fluids at 70 degrees C for 30 min. The febrile responses were in parallel with the levels of interleukin-1 (IL 1), tumor necrosis factor (TNF), interferon-gamma (IFN-gamma), IL-2, and IL-6 in supernatant fluids obtained from PBMC treated with SEA. Both the pyrogenicity and the levels of IL-1, TNF, IFN-gamma, IL-2, and IL-6 in supernatant fluids started to rise at 6 to 18 h and reached their peak levels at 24 to 96 h after SEA incubation. Both the fever and the increased levels of IL-1, TNF, IFN-gamma, IL 2, and IL-6 in supernatant fluids obtained from the SEA-stimulated PBMC were decreased by incubating SEA-PBMC with anisomycin (a protein synthesis inhibitor), aminoguanidine (an inhibitor of inducible nitric oxide synthase [NOS]), or dexamethasone (an inhibitor of NOS). The febrile response to supernatant fluids obtained from the SEA-stimulated PBMC was attenuated by adding either anti-IL 1beta, anti-TNF-alpha, or anti-IFN-gamma monoclonal antibody (MAb) to supernatant fluids. The antipyretic effects exerted by anti-IL-1beta MAb were greater than those exerted by anti-TNF-alpha or anti-IFN-gamma MAb. The data suggest that SEA acts through the NOS mechanisms in PBMC to stimulate synthesis of pyrogenic cytokines (in particular, the IL-1beta). PMID- 10722596 TI - Molecular epidemiologic approaches to urinary tract infection gene discovery in uropathogenic Escherichia coli. AB - Urinary tract infection (UTI) is one of the most frequently acquired bacterial infections. The vast majority of UTIs are caused by a large, genetically heterogeneous group of Escherichia coli. This genetic diversity has hampered identification of UTI-related genes. A three-step experimental strategy was used to identify genes potentially involved in E. coli UTI transmission or virulence: epidemiologic pairing of a UTI-specific strain with a fecal control, differential cloning to isolated UTI strain-specific DNA, and epidemiologic screening to identify sequences among isolated DNAs that are associated with UTI. The 37 DNA sequences initially isolated were physically located all over the tester strain genome. Only two hybridized to the total DNA of the sequenced E. coli K-12 strain; eight sequences were present significantly more frequently in UTI isolates than in fecal isolates. Three of the eight sequences matched to genes for multidrug efflux proteins, usher proteins, and pathogenicity island insertion sites, respectively. Using population characteristics to direct gene discovery and evaluation is a productive strategy applicable to any system. PMID- 10722597 TI - Helicobacter pylori possesses two CheY response regulators and a histidine kinase sensor, CheA, which are essential for chemotaxis and colonization of the gastric mucosa. AB - Infection of the mucous layer of the human stomach by Helicobacter pylori requires the bacterium to be motile and presumably chemotactic. Previous studies have shown that fully functional flagella are essential for motility and colonization, but the role of chemotaxis remains unclear. The two-component regulatory system CheA/CheY has been shown to play a major role in chemotaxis in other enteric bacteria. Scrutiny of the 26695 genome sequence suggests that H. pylori has two CheY response regulators: one a separate protein (CheY1) and the other (CheY2) fused to the histidine kinase sensor CheA. Defined deletion mutations were introduced into cheY1, cheY2, and cheA in H. pylori strains N6 and SS1. Video tracking revealed that the wild-type H. pylori strain moves in short runs with frequent direction changes, in contrast to movement of cheY2, cheAY2, and cheAY2 cheY1 mutants, whose motion was more linear. The cheY1 mutant demonstrated a different motility phenotype of rapid tumbling. All mutants had impaired swarming and greatly reduced chemotactic responses to hog gastric mucin. Neither cheY1 nor cheAY2 mutants were able to colonize mice, but they generated a significant antibody response, suggesting that despite impaired chemotaxis, these mutants were able to survive in the stomach long enough to induce an immune response before being removed by gastric flow. Additionally, we demonstrated that cheY1 failed to colonize gnotobiotic piglets. This study demonstrates the importance of the roles of cheY1, cheY2, and cheA in motility and virulence of H. pylori. PMID- 10722598 TI - Role of Bordetella bronchiseptica fimbriae in tracheal colonization and development of a humoral immune response. AB - Fimbriae are filamentous, cell surface structures which have been proposed to mediate attachment of Bordetella species to respiratory epithelium. Bordetella bronchiseptica has four known fimbrial genes: fim2, fim3, fimX, and fimA. While these genes are unlinked on the chromosome, their protein products are assembled and secreted by a single apparatus encoded by the fimBCD locus. The fimBCD locus is embedded within the fha operon, whose genes encode another putative adhesin, filamentous hemagglutinin (FHA). We have constructed a Fim(-) B. bronchiseptica strain, RB63, by introducing an in-frame deletion extending from fimB through fimD. Western blot analysis showed that RB63 is unable to synthesize fimbriae but is unaffected for FHA expression. Using this mutant, we assessed the role of fimbriae in pathogenesis in vitro and in vivo in natural animal hosts. Although RB63 was not significantly defective in its ability to adhere to various tissue culture cell lines, including human laryngeal HEp-2 cells, it was considerably altered in its ability to cause respiratory tract infections in rats. The number of DeltafimBCD bacteria recovered from the rat trachea at 10 days postinoculation was significantly decreased compared to that of wild-type B. bronchiseptica and was below the limit of detection at 30 and 60 days postinoculation. The number of bacteria recovered from the nasal cavity and larynx was not significantly different between RB63 and the wild-type strain at any time point. The ability of fimbriae to mediate initial attachment to tracheal tissue was tested in an intratracheal inoculation assay. Significantly fewer RB63 than wild-type bacteria were recovered from the tracheas at 24 h after intratracheal inoculation. These results demonstrate that fimbriae are involved in enhancing the ability of B. bronchiseptica to establish tracheal colonization and are essential for persistent colonization at this site. Interestingly, anti-Bordetella serum immunoglobulin M (IgM) levels were significantly lower in animals infected with RB63 than in animals infected with wild-type B. bronchiseptica at 10 days postinoculation. Even at 30 days postinoculation, RB63-infected animals had lower serum anti-Bordetella antibody titers in general. This disparity in antibody profiles suggests that fimbriae are also important for the induction of a humoral immune response. PMID- 10722599 TI - Cytokine gene expression in innately susceptible BALB/c mice and relatively resistant C57BL/6 mice during infection with virulent Burkholderia pseudomallei. AB - Production of cytokines including gamma interferon (IFN-gamma) and tumor necrosis factor alpha (TNF-alpha) is an important early-stage host response following infection with intracellular pathogens. Development of immunity to these pathogens is determined to a large extent by the timing and relative level of expression of the cytokines. Numerous studies have shown that early cytokine responses involving interleukin-12 (IL-12) and IFN-gamma are important for resistance to intracellular pathogens, whereas responses involving IL-4 and IL-10 increase host susceptibility. These often-indistinct early cytokine responses influence the differentiation of naive CD4(+) T helper cells, which later develop into what have commonly been termed Th1- and Th2-type cells. The characterization of CD4(+) T-helper-cell responses as Th1 or Th2 type is based largely on the cytokine profiles during the specific phase and has been used in recent years to account for the innate resistance and susceptibility of different inbred strains of mice to several intracellular pathogens. Studies investigating cytokine production in terms of CD4(+) T-helper-cell polarization in Burkholderia pseudomallei infection have not been undertaken. In this study, we used semiquantitative reverse transcription-PCR to assess induction of cytokine mRNA in liver and spleen of B. pseudomallei-susceptible BALB/c and relatively resistant C57BL/6 mice following infection with virulent B. pseudomallei. The levels of mRNA for IFN-gamma, TNF-alpha, IL-1beta, IL-6, IL-10, and IL-12 increased in both BALB/c and C57BL/6 mice 24 to 36 h after infection. A comparison of BALB/c and C57BL/6 responses revealed the relative levels of expression of mRNA for several of these cytokines, including IFN-gamma, were greater in BALB/c mice, suggesting a role for endotoxic shock and cytokine mediated immunopathology in the development of acute melioidosis. Early induction of mRNA for the cytokines classically associated with development of Th1- and Th2 type responses was absent or minimal, and induction levels in both strains of mice were similar. During the specific phase, cytokine mRNA profiles occurred as a combination of Th1- and Th2-type patterns. Collectively, these results demonstrate that cytokine mRNA responses in BALB/c and C57BL/6 mice following infection with virulent B. pseudomallei do not develop as polarized Th1- or Th2 type profiles. Considering the role of TNF-alpha and IFN-gamma in the processes of endotoxic shock, these results also indicate that selected cytokines, while important for resistance to B. pseudomallei infection, are also potential contributors to immunopathology and the development of acute fulminating disease. PMID- 10722600 TI - Trehalose 6,6'-dimycolate (Cord factor) enhances neovascularization through vascular endothelial growth factor production by neutrophils and macrophages. AB - Trehalose 6,6'-dimycolate (TDM) plays important roles in the development of granulomatous inflammation during infection with Mycobacterium spp., Rhodococcus spp., etc. To reveal the augmenting effect of TDM on vascular endothelial growth factor (VEGF) production and neovascularization, we investigated murine granulomatous tissue air pouches induced by Rhodococcus sp. strain 4306 TDM dissolved in Freund's incomplete adjuvant (FIA), comparing them to pouches treated with FIA alone. Histologically, granulomatous tissue and new vessel formation, which reached a maximum at day 7, was greatly enhanced by treatment with TDM. At day 1, VEGF-positive neutrophils accumulated in the pouch wall with frequency of 95% of total infiltrating cells, adhering to TDM-containing micelles. By day 3, granulomatous tissue and new vessels started to develop, and VEGF-positive macrophages appeared in a small number and gradually increased in number thereafter. The pouch contents of VEGF, interleukin-1beta, tumor necrosis factor alpha, and transforming growth factor beta were significantly elevated in TDM-treated pouches, with peaks at days 1, 0.5, 1, and 3, respectively, compared to those of control pouches, while that of basic fibroblast growth factor showed no significant increase. Treatment with anti-VEGF antibody inhibited TDM-induced granulomatous tissue formation and neovascularization, and administration of recombinant VEGF into pouches treated with FIA alone induced neovascularization comparable to that in the TDM-treated pouches. Incubation of neutrophils and macrophages on TDM-coated plastic dishes increased the VEGF release. The present results indicate that TDM augments VEGF production by neutrophils and macrophages and induces neovascularization in the granulomatous tissue. PMID- 10722601 TI - Nonopsonic binding of type III Group B Streptococci to human neutrophils induces interleukin-8 release mediated by the p38 mitogen-activated protein kinase pathway. AB - Nonopsonic interaction of host immune cells with pathogens is an important first line of defense. We hypothesized that nonopsonic recognition between type III group B streptococcus and human neutrophils would occur and that the interaction would be sufficient to trigger neutrophil activation. By using a serum-free system, it was found that heat-killed type III group B streptococci bound to neutrophils in a rapid, stable, and inoculum-dependent manner that did not result in ingestion. Transposon-derived type III strain COH1-13, which lacks capsular polysaccharide, and strain COH1-11 with capsular polysaccharide lacking terminal sialic acid demonstrated increased neutrophil binding, suggesting that capsular polysaccharide masks an underlying binding site. Experiments using monoclonal antibodies to complement receptor 1 and to the I domain or lectin site of complement receptor 3 did not inhibit binding, indicating that the complement receptors used for ingestion of opsonized group B streptococci were not required for nonopsonic binding. Nonopsonic binding resulted in rapid activation of cellular p38 and p44/42 mitogen-activated protein kinases. This interaction was not an effective trigger for superoxide production but did promote release of the proinflammatory cytokine interleukin-8. The release of interleukin-8 was markedly suppressed by the p38 mitogen-activated protein kinase inhibitor SB203580 but was only minimally suppressed by the mitogen-activated protein/extracellular signal regulated kinase inhibitor PD98059. Thus, nonopsonic binding of type III group B streptococci to neutrophils is sufficient to initiate intracellular signaling pathways and could serve as an arm of innate immunity of particular importance to the immature host. PMID- 10722602 TI - Staphylococcus aureus protein A recognizes platelet gC1qR/p33: a novel mechanism for staphylococcal interactions with platelets. AB - The adhesion of Staphylococcus aureus to platelets is a major determinant of virulence in the pathogenesis of endocarditis. Molecular mechanisms mediating S. aureus interactions with platelets, however, are incompletely understood. The present study describes the interaction between S. aureus protein A and gC1qR/p33, a multifunctional, ubiquitously distributed cellular protein, initially described as a binding site for the globular heads of C1q. Suspensions of fixed S. aureus or purified protein A, chemically cross-linked to agarose support beads, were found to capture native gC1qR from whole platelets. Moreover, biotinylated protein A bound specifically to fixed, adherent, human platelets. This interaction was inhibited by unlabeled protein A, soluble recombinant gC1qR (rgC1qR), or anti-gC1qR antibody F(ab')(2) fragments. The interaction between protein A and platelet gC1qR was underscored by studies illustrating preferential recognition of the protein A-bearing S. aureus Cowan I strain by gC1qR compared to recognition of the protein A-deficient Wood 46 strain, as well as inhibition of S. aureus Cowan I strain adhesion to immobilized platelets by soluble protein A. Further characterization of the protein A-gC1qR interaction by solid-phase enzyme-linked immunosorbent assay techniques measuring biotinylated gC1qR binding to immobilized protein A revealed specific binding that was inhibited by soluble protein A with a 50% inhibitory concentration of (3.3 +/- 0.7) x 10(-7) M (mean +/- standard deviation; n = 3). Rabbit immunoglobulin G (IgG) also prevented gC1qR-protein A interactions, and inactivation of protein A tyrosil residues by hyperiodination, previously reported to prevent the binding of IgG Fc, but not Fab, domains to protein A, abrogated gC1qR binding. These results suggest similar protein A structural requirements for gC1qR and IgG Fc binding. Further studies of structure and function using a truncated gC1qR mutant lacking amino acids 74 to 95 demonstrated that the protein A binding domain lies outside of the gC1qR amino-terminal alpha helix, which contains binding sites for the globular heads of C1q. In conclusion, the data implicate the platelet gC1qR as a novel cellular binding site for staphylococcal protein A and suggest an additional mechanism for bacterial cell adhesion to sites of vascular injury and thrombosis. PMID- 10722603 TI - Interleukin-12 neutralization alters lung inflammation and leukocyte expression of CD80, CD86, and major histocompatibility complex class II in mice infected with Histoplasma capsulatum. AB - Histoplasma capsulatum induces a cell-mediated immune response in lungs and lymphoid organs of mammals. Resolution of primary infection in mice depends on interleukin-12 (IL-12), since neutralization of this monokine increases susceptibility to infection. The present study was designed to determine if blockade of IL-12 disrupts the protective immune response by altering the influx of lineage-specific cells into infected lungs and the numbers of cells expressing CD80, CD86, CD119, and major histocompatibility complex class II (MHC II) molecules. In mice given anti-IL-12, there was a 2.5-fold decrease in total numbers of T cells on days 3 to 10 of infection and a 4-fold increase in Mac-1/Gr 1(+) cells on days 7 and 10 compared to infected controls. CD80(+) lung cells from anti-IL-12-treated mice were 2- to 3-fold greater than those from controls on days 7 and 10, whereas the total numbers of CD86(+) cells were 2- to 3-fold less and MHC II(+) cells were 1.5- to 2-fold less on days 3 and 5. Cells expressing CD119 were reduced 1.5-fold on day 5. Treatment with monoclonal antibodies (MAb) to CD80, CD86, or both reduced the fungal burden slightly compared to that in rat immunoglobulin G-treated controls, whereas after IL-12 neutralization, blocking of CD80 reduced the tissue burden by 2. 5-fold and this correlated with a decrease in IL-4. Regardless, mortality was not altered by treatment with MAb to CD80 or CD86. We conclude that (i) IL-12 neutralization alters the nature of the inflammatory response in lungs and the expression of CD80 and CD86 on lineage-specific cells, (ii) the immune response during infection with H. capsulatum is controlled via mechanisms independent of the CD80 and CD86 costimulatory pathways, and (iii) decreased expression of CD86 and MHC II may modulate generation of optimal protective immunity. PMID- 10722604 TI - Dual role for transforming growth factor beta-dependent signaling in Trypanosoma cruzi infection of mammalian cells. AB - Expression of functional transforming growth factor beta (TGF-beta) receptors (TbetaR) is required for the invasion of mammalian cells by the protozoan parasite Trypanosoma cruzi. However, the precise role of this host cell signaling complex in T. cruzi infection is unknown. To investigate the role of the TGF-beta signaling pathway, infection levels were studied in the mink lung epithelial cell lines JD1, JM2, and JM3. These cells express inducible mutant TbetaR1 proteins that cannot induce growth arrest in response to TGF-beta but still transmit the signal for TGF-beta-dependent gene expression. In the absence of mutant receptor expression, trypomastigotes invaded the cells at a low level. Induction of the mutant receptors caused an increase in infection in all three cell lines, showing that the requirement for TGF-beta signaling at invasion can be divorced from TGF beta-induced growth arrest. TGF-beta pretreatment of mink lung cells expressing wild-type TbetaR1 caused a marked enhancement of infection, but no enhancement was seen in JD1, JM2, and JM3 cells, showing that the ability of TGF-beta to stimulate infection is associated with growth arrest. Likewise, expression of SMAD7 or SMAD2SA, inhibitors of TGF-beta signaling, did not block infection by T. cruzi but did block the enhancement of infection by TGF-beta. Taken together, these results show that there is a dual role for TGF-beta signaling in T. cruzi infection. The initial invasion of the host cell is independent of both TGF-beta dependent gene expression and growth arrest, but TGF-beta stimulation of infection requires a fully functional TGF-beta signaling pathway. PMID- 10722605 TI - Molecular and biological analysis of eight genetic islands that distinguish Neisseria meningitidis from the closely related pathogen Neisseria gonorrhoeae. AB - The pathogenic species Neisseria meningitidis and Neisseria gonorrhoeae cause dramatically different diseases despite strong relatedness at the genetic and biochemical levels. N. meningitidis can cross the blood-brain barrier to cause meningitis and has a propensity for toxic septicemia unlike N. gonorrhoeae. We previously used subtractive hybridization to identify DNA sequences which might encode functions specific to bacteremia and invasion of the meninges because they are specific to N. meningitidis and absent from N. gonorrhoeae. In this report we show that these sequences mark eight genetic islands that range in size from 1.8 to 40 kb and whose chromosomal location is constant. Five of these genetic islands were conserved within a representative set of strains and/or carried genes with homologies to known virulence factors in other species. These were deleted, and the mutants were tested for correlates of virulence in vitro and in vivo. This strategy identified one island, region 8, which is needed to induce bacteremia in an infant rat model of meningococcal infection. Region 8 encodes a putative siderophore receptor and a disulfide oxidoreductase. None of the deleted mutants was modified in its resistance to the bactericidal effect of serum. Neither were the mutant strains altered in their ability to interact with endothelial cells, suggesting that such interactions are not encoded by large genetic islands in N. meningitidis. PMID- 10722606 TI - Identification of regions of the Escherichia coli chromosome specific for neonatal meningitis-associated strains. AB - Specific virulence factors associated with the pathogenesis of Escherichia coli strains causing neonatal meningitis (ECNM), such as the K1 capsular polysaccharide, the S fimbriae, and the Ibe10 protein, have been previously identified. However, some other yet unidentified factors are likely to be involved in the pathogenesis of ECNM. To identify specialized unique DNA regions associated with ECNM virulence, we used the representational difference analysis technique. The genomes of two strains belonging to nonpathogenic phylogenetic group A of the ECOR reference collection were subtracted from E. coli strain C5, isolated from a case of neonatal meningitis. Strain C5 belongs to the phylogenetic group B2 as do the majority of ECNM. We have isolated and mapped 64 DNA fragments which are specific for strain C5 and not found in nonpathogenic strains. Of these clones, 44 were clustered in six distinct regions on the chromosome. The sfa and ibe10 genes were located in regions 2 and 6, respectively. A group of genes (cnf1, hra, hly, and prs) known to be present in a pathogenicity island of the uropathogenic strain E. coli J96 colocalized with region 6. The occurrence of these DNA regions was tested in a set of meningitis associated strains and in a control group composed of non-meningitis-associated strains belonging to the same B2 group. Regions 1, 3, and 4 were present in 91, 82, and 81%, respectively, of the meningitis strains and in 40, 13, and 47% of the control strains. Together, these data suggest that regions 1, 3, and 4 code for factors associated with the ability of E. coli to invade the meninges of neonates. PMID- 10722607 TI - Linkage of exogenous T-cell epitopes to the 19-kilodalton region of Plasmodium yoelii merozoite surface protein 1 (MSP1(19)) can enhance protective immunity against malaria and modulate the immunoglobulin subclass response to MSP1(19). AB - The degree of protection against Plasmodium yoelii asexual blood stages induced by immunization of mice with the 19-kDa region of merozoite surface protein 1 (MSP1(19)) is H-2 dependent. As a strategy to improve the protection, mouse strains with disparate H-2 haplotypes were immunized with glutathione S transferase (GST)-MSP1(19) proteins including either a universal T-cell epitope from tetanus toxin (P2) or an I-A(k)-restricted T-cell epitope (P8) from Plasmodium falciparum Pf332. In H-2(k) mice which are poorly protected following immunization with GST-MSP1(19), GST-P2-MSP1(19) significantly improved the protection. In mice partially (H-2(k/b)) or well protected by GST-MSP1(19) (H 2(d) and H-2(b)), P2 did not further increase the protection. However, the protection of H-2(k/b) mice and to some extent H-2(k) mice was improved by immunization with GST-P8-MSP1(19). The magnitudes of immunoglobulin G1 (IgG1) and IgG2a responses in mice immunized with the GST-MSP1(19) variants correlated with low peak parasitemia, indicating a protective capacity of these IgG subclasses. In H-2(k) mice immunized with GST-P2-MSP1(19), both IgG1 and IgG2a responses were significantly enhanced. The epitope P2 appeared to have a general ability to modulate the IgG subclass response since all four mouse strains displayed elevated IgG2a and/or IgG2b levels after immunization with GST-P2-MSP1(19). In contrast, GST-P8-MSP1(19) induced a slight enhancement of IgG responses in H 2(k/b) and H-2(k) mice without any major shift in IgG subclass patterns. The ability to improve the protective immunity elicited by P. yoelii MSP1(19) may have implications for improvement of human vaccines based on P. falciparum MSP1(19). PMID- 10722608 TI - Chronic atrophic gastritis in SCID mice experimentally infected with Campylobacter fetus. AB - Campylobacter fetus is a cause of enteritis and invasive extraintestinal disease in humans. In order to develop an animal model of C. fetus infection, outbred ICR SCID mice were orally challenged with a clinical isolate of C. fetus. The stomachs of SCID mice were heavily colonized with C. fetus, and colonization was associated with the development of chronic atrophic gastritis. This lesion was characterized by an inflammatory infiltrate of granulocytes and macrophages that over time resulted in a loss of specialized parietal and chief cells in the corpus and the appearance of a metaplastic mucous epithelium. This lesion bears similarity to that encountered during experimental murine infection with Helicobacter pylori or Helicobacter felis. Despite colonization of the cecum and colon tissues by C. fetus in SCID mice, no lesions were noted in these tissues. A follow-up study confirmed these findings for SCID mice and also demonstrated that C. fetus could also infect the gastric mucosa of wild-type, outbred ICR mice. However, in ICR mice, the anatomic extent of colonization was more limited and the severity of inflammation and epithelial alterations was significantly less than that observed in infected SCID mice. The stomach may represent an unrecognized environmental niche for Campylobacter species. PMID- 10722610 TI - Group B streptococci and other gram-positive cocci bind to cytokeratin 8. AB - Group B streptococci (GBS) adhere to surface receptors present on epithelial cells; these receptors include fibronectin and laminin. To identify other possible receptors, plasma membranes from A549 cells, a respiratory tract epithelial cell line, were prepared. These plasma membranes were tested in a protein blot analysis using radiolabeled GBS as a probe. GBS adhered to two species, with molecular masses of 50 kDa (p50) and 57 kDa (p57). We concluded that p50 and p57 correspond to two forms of cytokeratin 8 (CK8) on the basis of the following results: (i) protein blot results demonstrated that p50 and p57 exactly comigrated with two forms of CK8 after separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (PAGE); (ii) p50 and p57 exactly comigrated with CK8 after separation by two-dimensional PAGE; (iii) CK8 in solution bound to GBS, as demonstrated by immunoblot analysis of proteins from A549 lysates that bound to GBS in a liquid-phase assay; and (iv) radiolabeled GBS bound to A549 lysate-derived CK8 that had been captured in anti-CK8-coated microtiter wells. CK8 bound to COH1-13, an acapsular mutant of COH1, demonstrating that adherence is not mediated by capsular polysaccharide. Trypsin treated GBS did not bind to CK8, indicating that adherence is mediated via a protein on the surface of GBS. Soluble CK8 bound to six of six GBS strains tested. Soluble CK8 also bound to Staphylococcus aureus, Lactococcus lactis, Enterococcus faecalis, and Streptococcus pyogenes. We hypothesize that adherence of GBS to cytokeratin may be important for maintenance of colonization at sites of keratinized epithelium, such as the vagina, or for adherence of these bacteria to damaged epithelial cells at other sites. PMID- 10722609 TI - Epitope mapping of the outer membrane protein P5-homologous fimbrin adhesin of nontypeable Haemophilus influenzae. AB - To identify potential immunodominant and/or adhesin binding domains of the outer membrane protein P5-homologous fimbrin adhesin of nontypeable Haemophilus influenzae (NTHI), three sets of synthetic peptides were synthesized and assayed in an adherence inhibition assay, by Western blotting, and in a biomolecular interaction analysis (BIA) system. The first series of 34 8- to 10-mer peptides represented the entire mature protein sequentially. The second set of four peptides (each 19 to 28 residues) represented the four predicted major surface exposed regions (or loops) of this adhesin. The third series of seven peptides (each 27 to 34 residues) were specifically designed to map the third surface exposed region. Data obtained by BIA indicated limited reactivity of a panel of high-titered immune chinchilla sera to the 8- to 10-mer peptides representing the mature protein, likely because these linear peptides did not represent continuous epitopes. However, several of these short peptides did inhibit adherence of multiple NTHI strains to a human respiratory epithelial cell. Overall, greatest relative reactivity in both BIA and adherence inhibition assays was demonstrated against, or shown by, peptides mapping to the third and fourth predicted surface exposed regions of this adhesin, thereby indicating the presence of immunodominant and adhesin binding domains at these sites. Middle ear fluids sequentially recovered from a chinchilla with an ongoing NTHI-induced otitis media (OM) as well as sera from children with OM due to NTHI also reacted exclusively with peptides representing the third and fourth surface-exposed regions of the P5-fimbrin adhesin, indicating a similarity in immune recognition of this bacterial protein by these two hosts. Collectively, these data together with the previously demonstrated protective efficacy of immunogens derived from this adhesin in chinchilla models support the continued development of P5-fimbrin based vaccine components. PMID- 10722611 TI - Pilot study of phoP/phoQ-deleted Salmonella enterica serovar typhimurium expressing Helicobacter pylori urease in adult volunteers. AB - Attenuated Salmonella enterica serovar Typhi has been studied as an oral vaccine vector. Despite success with attenuated S. enterica serovar Typhimurium vectors in animals, early clinical trials of S. enterica serovar Typhi expressing heterologous antigens have shown that few subjects have detectable immune responses to vectored antigens. A previous clinical study of phoP/phoQ-deleted S. enterica serovar Typhi expressing Helicobacter pylori urease from a multicopy plasmid showed that none of eight subjects had detectable immune responses to the vectored antigen. In an attempt to further define the variables important for engendering immune responses to vectored antigens in humans, six volunteers were inoculated with 5 x 10(7) to 8 x 10(7) CFU of phoP/phoQ-deleted S. enterica serovar Typhimurium expressing the same antigen. Two of the six volunteers had fever; none had diarrhea, bacteremia, or other serious side effects. The volunteers were more durably colonized than in previous studies of phoP/phoQ deleted S. enterica serovar Typhi. Five of the six volunteers seroconverted to S. enterica serovar Typhimurium antigens and had strong evidence of anti-Salmonella mucosal immune responses by enzyme-linked immunospot studies. Three of six (three of five who seroconverted to Salmonella) had immune responses in the most sensitive assay of urease-specific immunoglobulin production by blood mononuclear cells in vitro. One of these had a fourfold or greater increase in end-point immunoglobulin titer in serum versus urease. Attenuated S. enterica serovar Typhimurium appears to be more effective than S. enterica serovar Typhi for engendering immune responses to urease. Data suggest that this may be related to a greater stability of antigen-expressing plasmid in S. enterica serovar Typhimurium and/or prolonged intestinal colonization. Specific factors unique to nontyphoidal salmonellae may also be important for stimulation of the gastrointestinal immune system. PMID- 10722612 TI - Innate lung defenses and compromised Pseudomonas aeruginosa clearance in the malnourished mouse model of respiratory infections in cystic fibrosis. AB - Cystic fibrosis (CF) is characterized by dysfunction of the digestive and respiratory tracts resulting in generalized malnutrition and chronic respiratory infections. Chronic lung infections with Pseudomonas aeruginosa, intense neutrophil-dominated airway inflammation, and progressive lung disease are the major cause of high morbidity and mortality in CF. Here we investigated the effects of malnutrition in CF on innate lung defenses, susceptibility to P. aeruginosa colonization, and associated inflammation, using aerosol models of acute and chronic infections in normal, malnourished, and transgenic mice. CFTR(m1Unc-/-) knockout mice displayed body weight variations and showed variable pulmonary clearance of P. aeruginosa. This variability was not detected in bitransgenic CFTR(m1Unc-/-)(FABP-hCFTR) mice in which the intestinal defect had been corrected. Diet-induced protein calorie malnutrition in C57BL/6J mice resulted in impaired pulmonary clearance of P. aeruginosa. Tumor necrosis factor alpha (TNF-alpha) and nitrite levels detected upon exposure to P. aeruginosa aerosols were lower in the lungs of the malnourished C57BL/6J mice relative than in lungs of mice fed a normal diet. The role of TNF-alpha and reactive nitrogen intermediates in P. aeruginosa clearance was tested in TNF-alpha and inducible nitric oxide synthase (iNOS) knockout mice. P. aeruginosa clearance was diminished in transgenic TNF-alpha- and iNOS-deficient mice. In contrast to the effects of TNF-alpha and iNOS, gamma interferon knockout mice retained a full capacity to eliminate P. aeruginosa from the lung. Malnutrition also contributed to excessive inflammation in C57BL/6J mice upon chronic challenge with P. aeruginosa. The repeatedly infected malnourished host did not produce interleukin 10, a major anti-inflammatory cytokine absent or diminished in the bronchoalveolar fluids of CF patients. These results are consistent with a model in which defective CFTR in the intestinal tract leads to nutritional deficiency which in turn contributes to compromised innate lung defenses, bacterial colonization, and excessive inflammation in the CF respiratory tract. PMID- 10722614 TI - Upregulation of monocyte urokinase plasminogen activator receptor during human endotoxemia. AB - The receptor for urokinase-type plasminogen activator (uPAR) (CD87) plays an important role in leukocyte adhesion and migration. To assess the effect of endotoxin on cellular uPAR, uPAR expression was determined on leukocytes by fluorescence-activated cell sorter analysis in seven healthy subjects following intravenous injection of endotoxin (lot G; 4 ng/kg). Endotoxin induced a transient increase in uPAR expression on monocytes, reaching a 92% +/- 46% increase over baseline expression after 6 h (P < 0.05). Endotoxin did not influence uPAR expression on granulocytes, while uPAR remained undetectable on lymphocytes. Endotoxin also increased soluble uPAR levels in plasma (P < 0.05). Stimulation of human whole blood with endotoxin or gram-positive stimuli in vitro also resulted in an upregulation of monocyte uPAR expression. Although tumor necrosis factor alpha (TNF) upregulated monocyte uPAR expression, anti-TNF did not influence the endotoxin-induced increase in monocyte uPAR expression. These data suggest that infectious stimuli may influence monocyte function in vivo by enhancing the expression of uPAR. PMID- 10722613 TI - A large toxin from pathogenic Escherichia coli strains that inhibits lymphocyte activation. AB - The mechanisms by which bacteria resist cell-mediated immune responses to cause chronic infections are largely unknown. We report the identification of a large gene present in enteropathogenic strains of Escherichia coli (EPEC) that encodes a toxin that specifically inhibits lymphocyte proliferation and interleukin-2 (IL 2), IL-4, and gamma interferon production in response to a variety of stimuli. Lymphostatin, the product of this gene, is predicted to be 366 kDa and shares significant homology with the catalytic domains of the large clostridial cytotoxins. A mutant EPEC strain that has a disruption in this gene lacks the ability to inhibit lymphokine production and lymphocyte proliferation. Enterohemorrhagic E. coli strains of serotype O157:H7 possess a similar gene located on a large plasmid. Loss of the plasmid is associated with loss of the ability to inhibit IL-2 expression while transfer of the plasmid to a nonpathogenic strain of E. coli is associated with gain of this activity. Among 89 strains of E. coli and related bacteria tested, lifA sequences were detected exclusively in strains capable of attaching and effacing activity. Lymphostatin represents a new class of large bacterial toxins that blocks lymphocyte activation. PMID- 10722615 TI - Clostridium difficile recombinant toxin A repeating units as a carrier protein for conjugate vaccines: studies of pneumococcal type 14, Escherichia coli K1, and Shigella flexneri type 2a polysaccharides in mice. AB - Unlike the native protein, a nontoxic peptide (repeating unit of the native toxin designated rARU) from Clostridium difficile toxin A (CDTA) afforded an antigen that could be bound covalently to the surface polysaccharides of pneumococcus type 14, Shigella flexneri type 2a, and Escherichia coli K1. The yields of these polysaccharide-protein conjugates were significantly increased by prior treatment of rARU with succinic anhydride. Conjugates, prepared with rARU or succinylated (rARUsucc), were administered to mice by a clinically relevant dosage and immunization scheme. All conjugates elicited high levels of serum immunoglobulin G both to the polysaccharides and to CDTA. Conjugate-induced anti-CDTA had neutralizing activity in vitro and protected mice challenged with CDTA, similar to the rARU alone. Conjugates prepared with succinylated rARU, therefore, have potential for serving both as effective carrier proteins for polysaccharides and for preventing enteric disease caused by C. difficile. PMID- 10722616 TI - Survival of group B streptococcus type III in mononuclear phagocytes: differential regulation of bacterial killing in cord macrophages by human recombinant gamma interferon and granulocyte-macrophage colony-stimulating factor. AB - Phagocytic and killing capacities of resident and cytokine-activated human macrophages against group B Streptococcus (GBS) type III were studied. Evidence is presented that monocyte-derived macrophages from cord and adults ingest serum opsonized GBS but that killing of bacteria was negligible in resident cells. Treatment of adult macrophages with recombinant human gamma interferon (rhIFN gamma; 100 U/ml) or recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF; 200 U/ml) resulted in significant increases of killing of GBS (P < 0.01 for each). The killing capacity of cord macrophages treated with rhGM CSF was also enhanced compared to that of untreated cells (P < 0.01). However, treatment with rhIFN-gamma resulted in only a moderate increase in the capacity of cord macrophages to kill GBS (P > 0.1). These results mirrored the effect of rhIFN-gamma on candidacidal capacities of cord and adult macrophages, reported earlier from our laboratory. These data indicate differential modulation of neonatal macrophages by rhGM-CSF and rhIFN-gamma. We suggest that administration of rhGM-CSF to neonates with invasive GBS disease may enhance host resistance to these bacteria. PMID- 10722618 TI - Acquisition of coinfection and simultaneous transmission of Borrelia burgdorferi and Ehrlichia phagocytophila by Ixodes scapularis ticks. AB - The agents of Lyme disease (Borrelia burgdorferi) and human granulocytic ehrlichiosis (Ehrlichia phagocytophila) are both transmitted by the tick Ixodes scapularis. In nature, ticks are often infected with both agents simultaneously. We studied whether previous infection with either Borrelia or Ehrlichia in ticks would affect acquisition and transmission of a second pathogen. Ehrlichia infected I. scapularis nymphs were fed upon Borrelia-infected mice, and Borrelia infected I. scapularis nymphs were fed upon Ehrlichia-infected mice. The efficiency with which previously infected nymphal ticks acquired a second pathogen from infected hosts was compared to that of uninfected ticks. An average of 51% +/- 15% of ticks acquired Ehrlichia from infected mice regardless of their prior infection status with Borrelia. An average of 85% +/- 10% of ticks acquired Borrelia from infected mice regardless of their prior infection status with Ehrlichia. Also, we assessed the efficiency with which individual nymphs could transmit either agent alone, or both agents simultaneously, to individual susceptible hosts. An average of 76% +/- 9% of Borrelia-infected ticks and 84% +/ 10% of Ehrlichia-infected ticks transmitted these agents to mice regardless of the presence of the other pathogen. There was no evidence of interaction between the agents of Lyme disease and human granulocytic ehrlichiosis in I. scapularis ticks. The presence of either agent in the ticks did not affect acquisition of the other agent from an infected host. Transmission of the agents of Lyme disease and human granulocytic ehrlichiosis by individual ticks was equally efficient and independent. Dually infected ticks transmitted each pathogen to susceptible hosts as efficiently as ticks infected with only one pathogen. PMID- 10722617 TI - Role of tir and intimin in the virulence of rabbit enteropathogenic Escherichia coli serotype O103:H2. AB - Attaching and effacing (A/E) rabbit enteropathogenic Escherichia coli (REPEC) strains belonging to serogroup O103 are an important cause of diarrhea in weaned rabbits. Like human EPEC strains, they possess the locus of enterocyte effacement clustering the genes involved in the formation of the A/E lesions. In addition, pathogenic REPEC O103 strains produce an Esp-dependent but Eae (intimin) independent alteration of the host cell cytoskeleton characterized by the formation of focal adhesion complexes and the reorganization of the actin cytoskeleton into bundles of stress fibers. To investigate the role of intimin and its translocated coreceptor (Tir) in the pathogenicity of REPEC, we have used a newly constructed isogenic tir null mutant together with a previously described eae null mutant. When human HeLa epithelial cells were infected, the tir mutant was still able to induce the formation of stress fibers as previously reported for the eae null mutant. When the rabbit epithelial cell line RK13 was used, REPEC O103 produced a classical fluorescent actin staining (FAS) effect, whereas both the eae and tir mutants were FAS negative. In a rabbit ligated ileal loop model, neither mutant was able to induce A/E lesions. In contrast to the parental strain, which intimately adhered to the enterocytes and destroyed the brush border microvilli, bacteria of both mutants were clustered in the mucus without reaching and damaging the microvilli. The role of intimin and Tir was then analyzed in vivo by oral inoculation of weaned rabbits. Although both mutants were still present in the intestinal flora of the rabbits 3 weeks after oral inoculation, neither mutant strain induced any clinical signs or significant weight loss in the inoculated rabbits whereas the parental strain caused the death of 90% of the inoculated rabbits. Nevertheless, an inflammatory infiltrate was present in the lamina propria of the rabbits infected with both mutants, with an inflammatory response greater for the eae null mutant. In conclusion, we have confirmed the role of intimin in virulence, and we have shown, for the first time, that Tir is also a key factor in vivo for pathogenicity. PMID- 10722619 TI - Antibody-mediated elimination of the obligate intracellular bacterial pathogen Ehrlichia chaffeensis during active infection. AB - It is generally accepted that cellular, but not humoral immunity, plays an important role in host defense against intracellular bacteria. However, studies of some of these pathogens have provided evidence that antibodies can provide immunity if present during the initiation of infection. Here, we examined immunity against infection by Ehrlichia chaffeensis, an obligate intracellular bacterium that causes human monocytic ehrlichiosis. Studies with mice have demonstrated that immunocompetent strains are resistant to persistent infection but that SCID mice become persistently and fatally infected. Transfer of immune serum or antibodies obtained from immunocompetent C57BL/6 mice to C57BL/6 scid mice provided significant although transient protection from infection. Bacterial clearance was observed when administration occurred at the time of inoculation or well after infection was established. The effect was dose dependent, occurred within 2 days, and persisted for as long as 2 weeks. Weekly serum administration prolonged the survival of susceptible mice. Although cellular immunity is required for complete bacterial clearance, the data show that antibodies can play a significant role in the elimination of this obligate intracellular bacterium during active infection and thus challenge the paradigm that humoral responses are unimportant for immunity to such organisms. PMID- 10722620 TI - CD8(+) T-cell priming against a nonsecreted Listeria monocytogenes antigen is independent of the antimicrobial activities of gamma interferon. AB - Sublethal infection of mice with recombinant Listeria monocytogenes expressing a model epitope in either secreted or nonsecreted form results in similar CD8(+) T cell priming. Since nonsecreted bacterial proteins have no obvious access to the endogenous major histocompatibility complex (MHC) class I presentation pathway, presentation of these antigens requires destruction of the bacterium to reveal the nonsecreted molecules to an exogenous MHC class I presentation pathway. Gamma interferon (IFN-gamma), a cytokine made by multiple cell types in response to L. monocytogenes infection, could be required for exogenous presentation of nonsecreted bacterial antigens via its capacity to upregulate the expression of molecules involved in antigen presentation, its capacity to activate macrophages to kill bacteria to expose nonsecreted molecules or both. IFN-gamma knockout (KO) mice were used to address the requirement for IFN-gamma in CD8(+) T-cell priming against (i) a model exogenous antigen and (ii) secreted and nonsecreted L. monocytogenes antigens. We demonstrate that IFN-gamma KO mice are capable of cross-presenting the model exogenous antigen ovalbumin to prime CD8(+) T-cell responses that are only slightly weaker than that in wild-type (WT) mice. Despite their extreme susceptibility to primary L. monocytogenes infection, previously immunized and naive IFN-gamma KO mice were able to generate CD8(+) T-cell responses against both secreted and nonsecreted L. monocytogenes antigens which were similar to responses of WT mice. Interestingly, IFN-gamma KO mice were as capable as WT mice in mediating the characteristic drop in bacterial load in the liver at 4 h postinfection, although the IFN-gamma KO mice have exacerbated bacterial loads as early as 24 h postinfection. These results demonstrate that the regulatory functions of IFN-gamma are not required for priming of CD8(+) T cells by cross-presentation of a model exogenous antigen or in response to a nonsecreted L. monocytogenes antigen. In addition, the capacity of IFN-gamma to activate the microbicidal activities of macrophages is not required for the very early innate immune response to L. monocytogenes or priming of CD8(+) T cells against a nonsecreted bacterial antigen. PMID- 10722622 TI - Immunogenicity and efficacy in aotus monkeys of four recombinant Plasmodium falciparum vaccines in multiple adjuvant formulations based on the 19-kilodalton C terminus of merozoite surface protein 1. AB - The immunogenicity and protective efficacy of four versions of recombinant C terminal 19-kDa epidermal growth factor-like region of the major surface protein 1 (rMSP1(19)) of Plasmodium falciparum was studied in Aotus monkeys. Vaccination with each of the four rMSP1(19) constructs elicited high levels of antibodies to MSP1(19) but only one construct, the 19-kDa fragment expressed as a secreted fusion protein from Saccharomyces cerevisiae (yP30P2MSP1(19)), induced a high degree of protective immunity in Aotus nancymai against lethal P. falciparum challenge. Protective formulation required Freund's adjuvant; vaccination with yP30P2MSP1(19) in six other adjuvants that are suitable for human use induced lower levels of antibody response and no protection. These results emphasize the need to continue the search for an adjuvant that is comparable to Freund's adjuvant in potency and is safe for use in humans. PMID- 10722621 TI - Four different genes responsible for nonimmune immunoglobulin-binding activities within a single strain of Escherichia coli. AB - Certain Escherichia coli strains bind the Fc fragment of immunoglobulin G (IgG) at the bacterial cell surface. Previous work established that this nonimmune Ig binding depends on several large proteins with apparent molecular masses that can exceed 200 kDa. For E. coli strain ECOR-9, four distinct genes (designated eibA, eibC, eibD, and eibE) are responsible for Ig binding. Two eib genes are linked to eaa genes, which are homologous to genes for the autotransporter family of secreted proteins. With reference to the E. coli K-12 chromosome, the eibA-eaaA cluster is adjacent to trpA (min 28.3) while the eibC-eaaC cluster is adjacent to aspS (min 42. 0). Sequence adjacent to the eibA-eaaA cluster converges with that of strain K-12 precisely as observed for the Atlas family of prophages, suggesting that eibA is part of one of these. All four eib genes, when cloned into plasmid vectors, impart IgG binding to E. coli K-12 strains, and three impart IgA binding also. The IgG binding occurs at the bacterial cell surface, and its expression increases survival in serum by up to 3 orders of magnitude. The eib sequences predict a C-terminal peptide motif that is characteristic of outer membrane proteins, and the protein sequences show significant similarity near the C terminus to both the YadA virulence factor of Yersinia species and the universal surface protein A II of Moraxella catarrhalis. The sizes predicted for Eib proteins from DNA sequence are much smaller than their apparent sizes on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, possibly reflecting stable oligomerization. PMID- 10722623 TI - gammadelta T cells are a component of early immunity against preerythrocytic malaria parasites. AB - We tested the hypothesis that gammadelta T cells are a component of an early immune response directed against preerythrocytic malaria parasites that are required for the induction of an effector alphabeta T-cell immune response generated by irradiated-sporozoite (irr-spz) immunization. gammadelta T-cell deficient (TCRdelta(-/-)) mice on a C57BL/6 background were challenged with Plasmodium yoelii (17XNL strain) sporozoites, and then liver parasite burden was measured at 42 h postchallenge. Liver parasite burden was measured by quantification of parasite-specific 18S rRNA in total liver RNA by quantitative competitive reverse transcription-PCR and by an automated 5' exonuclease PCR. Sporozoite-challenged TCRdelta(-/-) mice showed a significant (P < 0.01) increase in liver parasite burden compared to similarly challenged immunocompetent mice. In support of this result, TCRdelta(-/-) mice were also found to be more susceptible than immunocompetent mice to a sporozoite challenge when blood-stage parasitemia was used as a readout. A greater percentage of TCRdelta(-/-) mice than of immunocompetent mice progressed to a blood-stage infection when challenged with five or fewer sporozoites (odds ratio = 2.35, P = 0.06). TCRdelta(-/-) mice receiving a single irr-spz immunization showed percent inhibition of liver parasites comparable to that of immunized immunocompetent mice following a sporozoite challenge. These data support the hypothesis that gammadelta T cells are a component of early immunity directed against malaria preerythrocytic parasites and suggest that gammadelta T cells are not required for the induction of an effector alphabeta T-cell immune response generated by irr-spz immunization. PMID- 10722624 TI - Interaction of HLA and age on levels of antibody to Plasmodium falciparum rhoptry associated proteins 1 and 2. AB - The Plasmodium falciparum rhoptry-associated proteins 1 and 2 (RAP1 and RAP2) are candidate antigens for a subunit malaria vaccine. The design of the study, which looks at the acquisition of immunity to malaria from childhood to old age, has allowed us to document the interaction of HLA and age on levels of antibody to specific malarial antigens. Antibodies reach maximum levels to RAP1 after the age of 15 but to RAP2 only after the age of 30. The effect of HLA-DRB1 and -DQB1 and age on levels of antibody to rRAP1 and rRAP2 was analyzed with a multiple regression model in which all HLA alleles and age were independent variables. DQB1*0301 and -*03032 showed an age-dependent association with levels of antibody to rRAP1, being significant in children 5 to 15 years (P < 0.001) but not in individuals over 15 years of age. DRB1*03011 showed an age-dependent association with antibody levels to rRAP2; however, this association was in adults over the age of 30 years (P < 0.01) but not in individuals under the age of 30 years. No associations were detected between DRB1 alleles and RAP1 antibody levels or between DQB1 alleles and RAP2 antibody levels. Thus, not only the HLA allele but also the age at which an interaction is manifested varies for different malarial antigens. The interaction may influence either the rate of acquisition of antibody or the final level of antibody acquired by adults. PMID- 10722627 TI - Effects of Shiga toxin 2 on lethality, fetuses, delivery, and puerperal behavior in pregnant mice. AB - Shiga toxin 2 (Stx2) is produced by enterohemorrhagic Escherichia coli (EHEC) and is known as the major virulence factor of EHEC. The aim of this study was to evaluate the effects of Stx2 on (i) maternal lethality, (ii) fetuses, (iii) delivery period, and (iv) maternal behavior after delivery. Timed pregnant ICR mice were injected intravenously with Stx2 on day 5 of pregnancy (early stage) or on day 15 (late stage). In early-stage experiments, the number of normal fetuses of mice injected with Stx2 was significantly lower than that of control mice. In late-stage experiments, mothers injected with Stx2 delivered normal numbers of neonates, but could not take care of them. The lethal doses of Stx2 were not different for pregnant and nonpregnant female mice at either stage. We conclude that Stx2 is toxic to the fetus in early pregnancy and affects maternal puerperal behavior in late pregnancy. PMID- 10722626 TI - Roles of interleukin-12 and gamma interferon in murine Chlamydia pneumoniae infection. AB - BALB/c and strain 129 mice infected intranasally with Chlamydia pneumoniae displayed a moderate-to-severe inflammation in the lungs and produced interleukin 12 (IL-12), gamma interferon (IFN-gamma), tumor necrosis factor alpha (TNF alpha), and IL-10, with peak levels on days 1 to 3 postinfection (p.i.), returning to basal levels by day 16 p.i. Anti-IL-12 treatment resulted in less severe pathological changes but higher bacterial titers on days 3 and 7 p.i. By day 16 p.i., the inflammatory responses of control antibody-treated mice subsided. The bacterial titers of both anti-IL-12- and control antibody-treated mice decreased within 3 weeks to marginally detectable levels. Anti-IL-12 treatment significantly reduced lung IFN-gamma production and in vitro spleen cell IFN-gamma production in response to either C. pneumoniae or concanavalin A. In gamma-irradiated infected mice, cytokine production was delayed, and this delay correlated with high bacterial titers in the lungs. Following C. pneumoniae infection, 129 mice lacking the IFN-gamma receptor alpha chain gene (G129 mice) produced similar IL-12 levels and exhibited similarly severe pathological changes but had higher bacterial titers than 129 mice. However, by day 45 p.i., bacterial titers became undetectable in both wild-type 129 and G129 mice. Thus, during C. pneumoniae lung infection, IL-12, more than IFN-gamma, plays a role in pulmonary cell infiltration. IFN-gamma and IL-12, acting mostly through its induction of IFN-gamma and Th1 responses, play an important role in controlling acute C. pneumoniae infection in the lungs, but eventually all mice control the infection to undetectable levels by IL-12- and IFN-gamma-independent mechanisms. PMID- 10722625 TI - Effect of Chlamydia trachomatis infection and subsequent tumor necrosis factor alpha secretion on apoptosis in the murine genital tract. AB - The pathology observed during Chlamydia infection is due initially to localized tissue damage caused by the infection itself, followed by deleterious host inflammatory responses that lead to permanent scarring. We have recently reported that the infection by Chlamydia in vitro results in apoptosis of epithelial cells and macrophages and that infected monocytes secrete the proinflammatory cytokine interleukin-1beta. At the same time, proinflammatory cytokines such as tumor necrosis factor alpha (TNF-alpha) can also trigger apoptosis of susceptible cells. To study the possible relationship between Chlamydia trachomatis infection and apoptosis in vivo, we used the terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling technique to determine whether infection may cause apoptosis in the genital tract of mice and, conversely, whether cytokines produced during the inflammatory response may modulate the level of apoptosis. Our results demonstrate that infected cells in the endocervix at day 2 or 7 after infection are sometimes apoptotic, although there was not a statistically significant change in the number of apoptotic cells in the endocervix. However, large clumps of apoptotic infected cells were observed in the lumen, suggesting that apoptotic cells may be shed from the endocervix. Moreover, there was a large increase in the number of apoptotic cells in the uterine horns and oviducts after 2 or 7 days of infection, which was accompanied by obvious signs of upper tract pathology. Interestingly, depletion of TNF-alpha led to a decrease in the level of apoptosis in the uterine horns and oviducts of animals infected for 7 days, suggesting that the inflammatory cytokines may exert part of their pathological effect via apoptosis in infected tissues. PMID- 10722628 TI - Macrophage migration inhibitory factor release by macrophages after ingestion of Plasmodium chabaudi-infected erythrocytes: possible role in the pathogenesis of malarial anemia. AB - Human falciparum malaria, caused by Plasmodium falciparum infection, results in 1 to 2 million deaths per year, mostly children under the age of 5 years. The two main causes of death are severe anemia and cerebral malaria. Malarial anemia is characterized by parasite red blood cell (RBC) destruction and suppression of erythropoiesis (the mechanism of which is unknown) in the presence of a robust host erythropoietin response. The production of a host-derived erythropoiesis inhibitor in response to parasite products has been implicated in the pathogenesis of malarial anemia. The identity of this putative host factor is unknown, but antibody neutralization studies have ruled out interleukin-1beta, tumor necrosis factor alpha, and gamma interferon while injection of interleukin 12 protects susceptible mice against lethal P. chabaudi infection. In this study, we report that ingestion of P. chabaudi-infected erythrocytes or malarial pigment (hemozoin) induces the release of macrophage migration inhibitory factor (MIF) from macrophages. MIF, a proinflammatory mediator and counter-regulator of glucocorticoid action, inhibits erythroid (BFU-E), multipotential (CFU-GEMM), and granulocyte-macrophage (CFU-GM) progenitor-derived colony formation. MIF was detected in the sera of P. chabaudi-infected BALB/c mice, and circulating levels correlated with disease severity. Liver MIF immunoreactivity increased concomitant with extensive pigment and parasitized RBC deposition. Finally, MIF was elevated three- to fourfold in the spleen and bone marrow of P. chabaudi infected mice with active disease, as compared to early disease, or of uninfected controls. In summary, the present results suggest that MIF may be a host-derived factor involved in the pathophysiology of malaria anemia. PMID- 10722629 TI - Developmental expression of a tandemly repeated, glycine- and serine-rich spore wall protein in the microsporidian pathogen Encephalitozoon cuniculi. AB - Microsporidia are intracellular organisms of increasing importance as opportunistic pathogens in immunocompromised patients. Host cells are infected by the extrusion and injection of polar tubes located within spores. The spore is surrounded by a rigid spore wall which, in addition to providing mechanical resistance, might be involved in host cell recognition and initiation of the infection process. A 51-kDa outer spore wall protein was identified in Encephalitozoon cuniculi with the aid of a monoclonal antibody, and the corresponding gene, SWP1, was cloned by immunoscreening of a cDNA expression library. The cDNA encodes a protein of 450 amino acids which displays no significant similarities to known proteins in databases. The carboxy-terminal region consists of five tandemly arranged glycine- and serine-rich repetitive elements. SWP1 is a single-copy gene that is also present in the genomes of Encephalitozoon intestinalis and Encephalitozoon hellem as demonstrated by Southern analysis. Indirect immunofluorescence and immunoelectron microscopy revealed that SWP1 is differentially expressed during the infection cycle. The protein is absent in replicative meronts until 24 h postinfection, and its expression is first induced in early sporonts at a time when organisms translocate from the periphery to the center of the parasitophorous vacuole. Expression of SWP1 appears to be regulated at the mRNA level, as was shown by reverse transcriptase PCR analysis. Further identification and characterization of stage-specific genes might help to unravel the complex intracellular differentiation process of microsporidia. PMID- 10722630 TI - The leptospiral major outer membrane protein LipL32 is a lipoprotein expressed during mammalian infection. AB - We report the cloning of the gene encoding the 32-kDa lipoprotein, designated LipL32, the most prominent protein in the leptospiral protein profile. We obtained the N-terminal amino acid sequence of a staphylococcal V8 proteolytic digest fragment to design an oligonucleotide probe. A Lambda-Zap II library containing EcoRI fragments of Leptospira kirschneri DNA was screened, and a 5.0 kb DNA fragment which contained the entire structural lipL32 gene was identified. Several lines of evidence indicate that LipL32 is lipid modified in a manner similar to that of other procaryotic lipoproteins. The deduced amino acid sequence of LipL32 would encode a 272-amino-acid polypeptide with a 19-amino-acid signal peptide, followed by a lipoprotein signal peptidase cleavage site. LipL32 is intrinsically labeled during incubation of L. kirschneri in media containing [(3)H]palmitate. The linkage of palmitate and the amino-terminal cysteine of LipL32 is acid labile. LipL32 is completely solubilized by Triton X-114 extraction of L. kirschneri; phase separation results in partitioning of LipL32 exclusively into the hydrophobic, detergent phase, indicating that it is a component of the leptospiral outer membrane. CaCl(2) (20 mM) must be present during phase separation for recovery of LipL32. LipL32 is expressed not only during cultivation but also during mammalian infection. Immunohistochemistry demonstrated intense LipL32 reactivity with L. kirschneri infecting proximal tubules of hamster kidneys. LipL32 is also a prominent immunogen during human leptospirosis. The sequence and expression of LipL32 is highly conserved among pathogenic Leptospira species. These findings indicate that LipL32 may be important in the pathogenesis, diagnosis, and prevention of leptospirosis. PMID- 10722631 TI - Intracellular trafficking and killing of Streptococcus pneumoniae by human alveolar macrophages are influenced by opsonins. AB - Human alveolar macrophages (HAM) are the major resident phagocytic cells of the gas-exchanging areas of the lung. Following contact with macrophages, bacteria enter phagosomes, which gradually acquire the characteristics of terminal phagolysosomes, with incorporation of lysosome-associated membrane protein (LAMP). We measured the binding of type 1 Streptococcus pneumoniae to the surface of HAM and then measured subsequent internalization and phagosomal incorporation of LAMP-1 under various opsonic conditions. Opsonization with serum containing immunoglobulin resulted in significantly greater binding of pneumococci to HAM compared with opsonization with immunoglobulin G (IgG)-depleted serum containing complement, which in turn resulted in marginally increased binding over that observed in the absence of opsonization. Internalization of opsonized S. pneumoniae gradually increased to a maximum of 20% of bound bacteria by 120 min of warm incubation, with 20% of internalized pneumococci being localized within LAMP-containing compartments by 80 min. Internalization of opsonized S. pneumoniae by HAM correlated with a reduction of bacterial viability. When inocula were adjusted so that pneumococcal binding under different conditions was equalized, subsequent internalization, trafficking to LAMP-containing compartments, and reduction of bacterial viability were less efficient in the absence of opsonization than that observed following opsonization with adsorbed or IgG-replete adsorbed serum. Once bound to the surface of HAM, pneumococci opsonized with adsorbed serum with or without IgG were internalized, processed, and killed equally well. In conclusion, binding, intracellular trafficking, and killing of S. pneumoniae by HAM are each significantly increased by opsonization with serum containing immunogloblin and/or complement. PMID- 10722632 TI - Induction of specific immunoglobulin A and Th2 immune responses to P6 outer membrane protein of nontypeable Haemophilus influenzae in middle ear mucosa by intranasal immunization. AB - Nontypeable Haemophilus influenzae (NTHI) is a major pathogen of otitis media. One of the outer membrane proteins of NTHI, P6, is an antigen common to all strains and is considered as a candidate for mucosal vaccine. To elucidate the possibility of developing a nasal vaccine against nontypeable Haemophilus influenzae (NTHI) and to investigate mucosal immune responses in the middle ear, mice were immunized intranasally with the P6 outer membrane protein of NTHI, and P6-specific immune responses in the middle ear mucosa were examined. Mice were given with P6 and cholera toxin intranasally as an adjuvant on days 0, 7, and 14 and were killed on day 21. The P6-specific immunoglobulin A (IgA) antibody titer in ear wash was significantly elevated. Mononuclear cells were isolated from middle ear mucosa, and an increase in P6-specific IgA-producing cells was shown with an enzyme-linked immunospot assay. In addition, an increase in memory T cells in middle ear mucosa was detected with flow cytometric analysis after intranasal immunization. Moreover, in vitro stimulation with P6 resulted in proliferation of purified CD4(+) T cells from immunized mice, and these T cells expressed Th2 cytokine mRNA. These results indicate that P6-specific IgA-B-cell immune responses and selected Th2 cytokine expressing Th cells were induced in middle ear mucosa by intranasal immunization. These findings suggest that a nasal vaccine is useful for preventing otitis media with effusion. PMID- 10722633 TI - Differential host inflammatory responses to viable versus antibiotic-killed bacteria in experimental microbial sepsis. AB - Staphylococcus aureus killed during imipenem or ceftazidime chemotherapy in mice elicited an early release of tumor necrosis factor alpha (TNF-alpha) into the systemic circulation. This response was coincident in time with an increase in leukocyte-endothelium adhesive interactions in the microvasculature. Equivalent responses were not observed without the antibiotic treatment (imipenem or ceftazidime). Protective efficacy of the same antibiotic treatment was markedly diminished in D-galactosamine-treated mice compared to controls; e.g., it dropped from 2,000-fold to 70-fold with 4 mg of imipenem per kg given at the time of challenge. Nevertheless, protection was quantitatively restored upon concurrent administration of neutralizing anti-TNF-alpha antibody or 4 mg of dexamethasone per kg to these TNF-alpha-hypersensitive mice. Importantly, protection afforded by dexamethasone was not seen when the animals were challenged with viable organisms but without the concurrent administration of antibiotic. An early TNF alpha response could also be demonstrated upon challenge with Escherichia coli, but in this instance, neither the timing nor the magnitude of that response was influenced by treatment with these antibiotics. We conclude from these studies that the inflammatory response to viable versus killed bacteria may differ markedly depending on the particular bacterium, host sensitivity to TNF-alpha, and possibly the Gram stain classification. PMID- 10722634 TI - Localization of Haemophilus ducreyi at the pustular stage of disease in the human model of infection. AB - To localize Haemophilus ducreyi in vivo, human subjects were experimentally infected with H. ducreyi until they developed a painful pustule or for 14 days. Lesions were biopsied, and biopsy samples were fixed in 4% paraformaldehyde, and cryosectioned. Sections were stained with polyclonal anti-H. ducreyi antiserum or H. ducreyi-specific monoclonal antibodies (MAbs) and fluorescently tagged secondary antibodies and examined by confocal microscopy. We identified H. ducreyi in 16 of 18 pustules but did not detect bacteria in the one papule examined. H. ducreyi was observed as individual cells and in clumps or chains. Staining with MAbs 2D8, 5C9, 3B9, 2C7, and 9D12 demonstrated that H. ducreyi expresses the major pilus subunit, FtpA, the 28-kDa outer membrane protein Hlp, the 18-kDa outer membrane protein PAL, and the major outer membrane protein (MOMP) or OmpA2 in vivo. By dual staining with polyclonal anti-H. ducreyi antiserum and MAbs that recognize human skin components, we observed bacteria within the neutrophilic infiltrates of all positively staining pustules and in the dermis of 10 of 16 pustules. We were unable to detect bacteria associated with keratinocytes in the samples examined. The data suggest that H. ducreyi is found primarily in association with neutrophils and in the dermis at the pustular stage of disease in the human model of infection. PMID- 10722635 TI - Cryptosporidium parvum induces host cell actin accumulation at the host-parasite interface. AB - Cryptosporidium parvum is an intracellular protozoan parasite that causes a severe diarrheal illness in humans and animals. Previous ultrastructural studies have shown that Cryptosporidium resides in a unique intracellular compartment in the apical region of the host cell. The mechanisms by which Cryptosporidium invades host intestinal epithelial cells and establishes this compartment are poorly understood. The parasite is separated from the host cell by a unique electron-dense structure of unknown composition. We have used indirect immunofluorescence microscopy and confocal laser scanning microscopy to characterize this structure. These studies indicate that host filamentous actin is assembled into a plaque-like structure at the host-parasite interface during parasite invasion and persists during parasite development. The actin-binding protein alpha-actinin is also present in this plaque early in parasite development but is lost as the parasite matures. Other actin-associated proteins, including vinculin, talin, and ezrin, are not present. We have found no evidence of tyrosine phosphorylation within this structure. Molecules known to link actin filaments to membrane were also examined, including alpha-catenin, beta-catenin, plakoglobin, and zyxin, but none was identified at the host-parasite junction. Thus, Cryptosporidium induces rearrangement of the host cell cytoskeleton and incorporates host cell actin and alpha-actinin into a host-parasite junctional complex. PMID- 10722636 TI - Identification of secreted proteins of Mycobacterium tuberculosis by a bioinformatic approach. AB - Proteins secreted by Mycobacterium tuberculosis are usually targets of immune responses in the infected host. Here we describe a search for secreted proteins that combined the use of bioinformatics and phoA' fusion technology. The 3,924 proteins deduced from the M. tuberculosis genome were analyzed with several computer programs. We identified 52 proteins carrying an NH(2)-terminal secretory signal peptide but lacking additional membrane-anchoring moieties. Of these 52 proteins-the TM1 subgroup-only 7 had been previously reported to be secreted proteins. Our predictions were confirmed in 9 of 10 TM1 genes that were fused to Escherichia coli phoA', a marker of subcellular localization. These findings demonstrate that the systematic computer search described in this work identified secreted proteins of M. tuberculosis with high efficiency and 90% accuracy. PMID- 10722637 TI - Stable expression of a new chimeric fluorescent reporter in the human malaria parasite Plasmodium falciparum. AB - Stable transfection of a new, chimeric reporter in the human malaria parasite Plasmodium falciparum confers green fluorescence and methotrexate resistance that can be quantitated by Western blotting and flow cytometry. This provides a sensitive, live reporter for exploitation of genomic and high-throughput assays for the identification of new pathogenic determinants. PMID- 10722638 TI - Specificity of human antibodies reactive with pneumococcal C polysaccharide. AB - Antibodies reactive with C polysaccharide (PS) were found in healthy adults, pneumococcal patients, and vaccinees. These antibodies were not directed to the phosphocholine determinant of the C PS, as they appear to be in mice, since the human antibodies were inhibitable only with C PS. We found another population of phosphocholine-specific antibodies inhibitable only by phosphocholine and related compounds. PMID- 10722639 TI - Hydroxamate siderophores of Histoplasma capsulatum. AB - The zoopathogenic fungus Histoplasma capsulatum, like other eukaryotic aerobic microorganisms, requires iron for growth. Under conditions of low iron availability, the fungus secretes hydroxamates that function as siderophores (iron chelators). The experiments to be reported were designed to gather further information on the hydroxamate siderophores of H. capsulatum. The fungus was grown in a synthetic medium deferrated with the cationic exchange resin Chelex 100. Siderophores were detected after 4 days of incubation at 37 degrees C in media containing 0.3 to 1.0 microM iron. The secretion was suppressed by 10 microM iron. The hydroxamates were purified by reverse-phase and size-exclusion chromatography. On the basis of ions observed during electrospray mass spectroscopy, five hydroxamate siderophores were tentatively identified: dimerum acid, acetyl dimerum acid, coprogen B, methyl coprogen B, and fusarinine (monomeric). A polyclonal antibody to dimerum acid was generated. This reagent cross-reacted with coprogen B and fusarinine. Thus, the antibody detects hydroxamates in all three families of siderophores excreted by H. capsulatum. PMID- 10722640 TI - Identification of a new repetitive element in Staphylococcus aureus. AB - The Staphylococcus aureus repeat (STAR) element is a sequence identified in two intergenic regions in S. aureus. The element is found in 13 to 21 copies in individual S. aureus strains, and elements in the homologous intergenic location are variable in length. The element sequence consists of several small and unusually GC-rich direct repeats with recurring intervening sequences. In addition, STAR-like elements may be present in related staphylococcal species. PMID- 10722641 TI - Epitope mapping of the immunodominant invariable region of Borrelia burgdorferi VlsE in three host species. AB - VlsE, the variable surface antigen of Borrelia burgdorferi, contains a 26-amino acid-long immunodominant invariable region, IR(6). In the present study, three overlapping 14-mer peptides reproducing the sequence of IR(6) were used as peptide-based enzyme-linked immunosorbent assay antigens to map this invariable region in infected monkeys, mice, and human Lyme disease patients. Antibodies of the two primate species appeared to recognize IR(6) as a single antigenic determinant, while mouse antibodies recognized multiple epitopes within this region. PMID- 10722642 TI - UV-B irradiation increases susceptibility of mice to malarial infection. AB - We here examined whether exposure of mice to UV-B affected their susceptibility to the murine malaria parasite Plasmodium chabaudi. When BALB/c mice with depilated skin were irradiated with UV-B and subsequently infected with the parasite, 80 to 100% of the UV-B-irradiated mice died within 12 days of infection with a sublethal dose. In addition, UV-B irradiation of C57BL/10 (B-10) mice, which are otherwise naturally resistant to the parasites, rendered them susceptible, and 100% of irradiated B-10 mice died within 11 days postinfection. The level of plasma gamma interferon (IFN-gamma) in unirradiated B-10 mice at 5 days after infection increased to 566 pg/ml, whereas the UV-B exposure of mice impaired the production of IFN-gamma, which showed a maximum level of 65 pg/ml in response to the parasite infection. The maximum level of plasma interleukin-10 in UV-B-irradiated mice in response to the parasite infection was approximately 1,100 pg/ml, which was approximately fourfold higher than the maximum level in unirradiated control mice. When UV-B-irradiated B-10 mice were administered murine recombinant IFN-gamma after infection, the mice regained parasite resistance. These results demonstrated that the UV-B exposure of mice enhances the susceptibility to the malaria parasites and suggested that the enhanced susceptibility following UV-B exposure was mediated by impairment of IFN-gamma production in response to the parasite infection. PMID- 10722643 TI - Escherichia coli O157:H7 causes more-severe systemic disease in suckling piglets than in colostrum-deprived neonatal piglets. AB - Our objective was to determine if suckling neonatal piglets are susceptible to enterohemorrhagic Escherichia coli (EHEC) O157:H7 disease. Surprisingly, EHEC O157:H7 caused more-rapid and more-severe neurological disease in suckling neonates than in those fed an artificial diet. Shiga toxin-negative O157:H7 did not cause neurological disease but colonized and caused attaching-and-effacing intestinal lesions. PMID- 10722644 TI - In vivo-induced genes in Pseudomonas aeruginosa. AB - In vivo expression technology was used for testing Pseudomonas aeruginosa in the rat lung model of chronic infection and in a mouse model of systemic infection. Three of the eight ivi proteins found showed sequence identity to known virulence factors involved in iron acquisition via an open reading frame (called pvdI) implicated in pyoverdine biosynthesis, membrane biogenesis (FtsY), and adhesion (Hag2). PMID- 10722645 TI - Disparate requirement for T cells in resistance to mucosal and acute systemic candidiasis. AB - Although highly susceptible to orogastric candidiasis, T-cell receptor delta- and alpha-chain knockout mice, deficient in gammadelta and alphabeta T cells, respectively, were found to be resistant to disseminated candidiasis of endogenous origin and to acute systemic candidiasis (resulting from intravenous injection). PMID- 10722646 TI - Staphylococcal exfoliative toxins cleave alpha- and beta-melanocyte-stimulating hormones. AB - The staphylococcal exfoliative toxins (ETs) A and B (ETA and ETB) are 27-kDa exotoxins produced by certain strains of Staphylococcus aureus and are the causative agents of staphylococcal scalded-skin syndrome. The crystal structures of the ETs strongly indicate that the proteins are members of the serine protease family of enzymes, although protease activity until now has not yet been conclusively demonstrated. Here, we show that the peptide beta-melanocyte stimulating hormone (beta-MSH) is cleaved by ETA and that both ETA and ETB are capable of cleaving alpha-MSH. Both toxins exhibit cleavage at specific glutamic acid residues in MSH peptides. Moreover, biologically inactive mutants of ETA were incapable of cleaving beta-MSH. PMID- 10722647 TI - Molecular and immunological analyses of the Borrelia turicatae Bdr protein family. AB - Here, we describe the molecular and immunological characterization of the bdr gene family of Borrelia turicatae, a relapsing-fever spirochete. Nine bdr alleles belonging to two different subfamilies were sequenced and localized to linear plasmids. Anti-Bdr antiserum was generated and used to analyze Bdr expression in pre- and postinfection isogenic populations. The analyses presented here provide a detailed characterization of the Bdr proteins in a relapsing-fever spirochete species, enhancing our understanding of these proteins at the genus-wide level. PMID- 10722648 TI - Xanthine oxidase contributes to host defense against Burkholderia cepacia in the p47(phox-/-) mouse model of chronic granulomatous disease. AB - Chronic granulomatous disease (CGD) is an inherited disorder of the NADPH oxidase in which phagocytes are defective in generating superoxide and downstream microbicidal reactive oxidants, leading to recurrent life-threatening bacterial and fungal infections. Xanthine oxidase (XO) is another enzyme known to produce superoxide in many tissues. Using the p47(phox-/-) mouse model of CGD, we evaluated the residual antibacterial activity of XO. Clearance of Burkholderia cepacia, a major pathogen in CGD, was reduced in p47(phox-/-) mice compared to that in wild-type mice and was further inhibited in p47(phox-/-) mice by pretreatment with the specific XO inhibitor allopurinol. Hepatic B. cepacia burden was similar in the two genotypes, but allopurinol significantly reduced net hepatic killing and killing efficiency only in p47(phox-/-) mice. Clearance and killing of intravenous Escherichia coli was intact in p47(phox-/-) mice and was unaffected by pretreatment with allopurinol. In CGD, XO may contribute to host defense against a subset of reactive oxidant-sensitive pathogens. PMID- 10722649 TI - Ultrastructural analysis of developmental events in Chlamydia pneumoniae-infected cells. AB - Chlamydia pneumoniae is an obligate intracellular parasite with a developmental cycle believed to be common to all members of the genus Chlamydia. We present a detailed description based on transmission and scanning electron microscopy of temporal events and inclusion structures throughout the C. pneumoniae AR-39 developmental cycle. PMID- 10722650 TI - Induction of emetic, pyrexic, and behavioral effects of Staphylococcus aureus enterotoxin B in the ferret. AB - Ferrets which had been orally dosed with 5 mg of Staphylococcal enterotoxin B (SEB) responded with an increase in subcutaneous temperature. At 75 min, the subcutaneous temperature was significantly higher (+ 0.9 degrees C +/- 0.38 degrees C, P < 0.007) than in control animals. Animals dosed with 1 or 2 mg of SEB responded with a small, but not significant, increase in subcutaneous temperature. All of the animals dosed with 5 mg of SEB retched and vomited. The mean latency for the onset of retching was 105 +/- 36 min, and the mean latency for the onset of vomiting was 106 +/- 34 min. The mean number of retches was 17.8 +/- 19.6, and the mean number of vomits was 2.0 +/- 1.5. These findings indicate that ferrets can be used as alternatives to primates for the study of the biological activities of SEB. PMID- 10722651 TI - Stimulus-specific interaction between activator-coactivator cognates revealed with a novel complex-specific antiserum. AB - A number of second messenger pathways propagate inductive signals via protein protein interactions that are phosphorylation-dependent. The second messenger, cAMP, for example, promotes cellular gene expression via the protein kinase A mediated phosphorylation of cAMP-response element-binding protein (CREB) at Ser(133), and this modification in turn stimulates the association of CREB with the co-activator, CREB-binding protein (CBP). The solution structure of the CREB.CBP complex, using relevant interaction domains, kinase inducible domain and kinase-induced domain interacting domain, referred to as KID and KIX, respectively, shows that KID undergoes a coil to helix transition, upon binding to KIX, that stabilizes complex formation. Whether such changes occur in the context of the full-length CREB and CBP proteins, however, is unclear. Here we characterize a novel antiserum that specifically binds to the CREB. CBP complex but to neither protein individually. Epitope mapping experiments demonstrate that the CREB.CBP antiserum detects residues in KID that undergo a conformational change upon binding to KIX. The ability of this antiserum to recognize full length CREB.CBP complexes in a phospho-(Ser(133))-dependent manner demonstrates that the structural transition observed with the isolated KID domain also occurs in the context of the full-length CREB protein. To our knowledge, this is the first report documenting formation of endogenous cellular protein-protein complexes in situ. PMID- 10722652 TI - Smad6 as a transcriptional corepressor. AB - Smad6 and Smad7, a subgroup of Smad proteins, antagonize the signals elicited by transforming growth factor-beta. These two Smads, induced by transforming growth factor-beta or bone morphogenetic protein (BMP) stimulation, form stable associations with their activated type I receptors, blocking phosphorylation of receptor-regulated Smads in the cytoplasm. Here we show that Smad6 interacts with homeobox (Hox) c-8 as a transcriptional corepressor, inhibiting BMP signaling in the nucleus. The interaction between Smad6 and Hoxc-8 was identified by a yeast two-hybrid approach and further demonstrated by co-immunoprecipitation assays in cells. Gel shift assays show that Smad6, but not Smad7, interacts with both Hoxc 8 and Hoxa-9 as a heterodimer when binding to DNA. More importantly, the Smad6 Hoxc-8 complex inhibits interaction of Smad1 with Hoxc-8- and Smad1-induced transcription activity. These data indicate that Smad6 interacts with Hox transcription factors as part of the negative feedback circuit in the BMP signaling pathway. PMID- 10722653 TI - Akt/protein kinase B is regulated by autophosphorylation at the hypothetical PDK 2 site. AB - The function of Akt (protein kinase B) is regulated by phosphorylation on two sites conserved within the AGC kinase family: the activation loop (Thr-308) in the kinase core and a hydrophobic phosphorylation site on the carboxyl terminus (Ser-473). Thr-308 is phosphorylated by the phosphoinositide-dependent kinase-1, (PDK-1), whereas the mechanism of phosphorylation of the hydrophobic site, tentatively referred to as the PDK-2 site, is unknown. Here we report that phosphorylation of the hydrophobic motif requires catalytically competent Akt. First we show that a kinase-inactive construct of Akt fails to incorporate phosphate at Ser-473 following IGF-1 stimulation in vivo but does incorporate phosphate at Thr-308 and a second carboxyl-terminal site, Thr-450; this ligand triggers the phosphorylation of both sites in wild-type enzyme. Neither does a catalytically inactive construct in which phosphorylation at the activation loop is blocked, T308A, become phosphorylated on the hydrophobic site in response to stimulation. Second, we show that Akt autophosphorylates on the hydrophobic site in vitro: phosphorylation of the activation loop by PDK-1 triggers the phosphorylation of the hydrophobic site in kinase-active, but not thermally inactivated, Akt alpha. Thus, Akt is regulated by autophosphorylation at the Ser 473 hydrophobic site. PMID- 10722654 TI - Disulfide bond formation between RNA binding domains is used to regulate mRNA binding activity of the chloroplast poly(A)-binding protein. AB - Binding of the chloroplast poly(A)-binding protein, RB47, to the psbA mRNA is regulated in response to light and is required for translation of this mRNA in chloroplasts. The RNA binding activity of RB47 can be modulated in vitro by oxidation and reduction. Site-directed mutations to individual cysteine residues in each of the four RNA binding domains of RB47 showed that changing single cysteines to serines in domains 2 or 3 reduced, but did not eliminate, the ability of RB47 to be redox-regulated. Simultaneously changing cysteines to serines in both domains 2 and 3 resulted in the production of RB47 protein that was insensitive to redox regulation but retained the ability to bind the psbA mRNA at high affinity. The poly(A)-binding protein from Saccharomyces cerevisiae lacks cysteine residues in RNA binding domains 2 and 3, and this poly(A)-binding protein lacks the ability to be regulated by oxidation or reduction. These data show that disulfide bond formation between RNA binding domains in a poly(A) binding protein can be used to regulate the ability of this protein to bind mRNA and suggest that redox regulation of RNA binding activity may be used to regulate translation in organisms whose poly(A)-binding proteins contain these critical cysteine residues. PMID- 10722655 TI - The cytoplasmic tail of CD1d contains two overlapping basolateral sorting signals. AB - CD1d is a member of the CD1 polypeptide family that represents a new arm of host defense against invading pathogens. In our previous work (Rodionov, D. G., Nordeng, T. W., Pedersen, K., Balk, S. P., and Bakke, O. (1999) J. Immunol. 162, 1488-1495) we have shown that CD1d contained a classic tyrosine-based internalization signal (YQGV) in its short cytoplasmic tail. CD1d is expressed in polarized epithelial cells, and we found that the cytoplasmic tail of CD1d also contained information for basolateral sorting. Interestingly, a mutation of the critical tyrosine residue of the endosomal sorting signal did not result in the loss of basolateral targeting of the mutant CD1d. To search for a basolateral sorting signal we have constructed a full set of alanine mutants, but no single alanine substitution inactivated the signal. However, deletions or mutations of either the C-terminal valine/leucine pair or the critical tyrosine residue from the internalization signal and either residue from the C-terminal valine/leucine pair inactivated basolateral sorting. Our data thus suggest that the cytoplasmic tail contains two overlapping basolateral signals, one tyrosine- and the other leucine-based, each being sufficient to direct CD1d to the basolateral membrane of polarized Madin-Darby canine kidney cells. PMID- 10722656 TI - Evidence that ThiI, an enzyme shared between thiamin and 4-thiouridine biosynthesis, may be a sulfurtransferase that proceeds through a persulfide intermediate. AB - ThiI is an enzyme common to the biosynthetic pathways leading to both thiamin and 4-thiouridine in tRNA. Comparison of the ThiI sequence with protein sequences in the data bases revealed that the Escherichia coli enzyme contains a C-terminal extension displaying sequence similarity to the sulfurtransferase rhodanese. Cys 456 of ThiI aligns with the active site cysteine residue of rhodanese that transiently forms a persulfide during catalysis. We investigated the functional importance of this sequence similarity and discovered that, like rhodanese, ThiI catalyzes the transfer of sulfur from thiosulfate to cyanide. Mutation of Cys-456 to alanine impairs this sulfurtransferase activity, and the C456A ThiI is incapable of supporting generation of 4-thiouridine in tRNA both in vitro and in vivo. We therefore conclude that Cys-456 of ThiI is critical for activity and propose that Cys-456 transiently forms a persulfide during catalysis. To accommodate this hypothesis, we propose a general mechanism for sulfur transfer in which the terminal sulfur of the persulfide first acts as a nucleophile and is then transferred as an equivalent of S(2-) rather than S(0). PMID- 10722657 TI - The underlying mechanism for the diversity of disulfide folding pathways. AB - The disulfide folding pathway of bovine pancreatic trypsin inhibitor (BPTI) is characterized by the predominance of folding intermediates with native-like structures. Our laboratory has recently analyzed the folding pathway(s) of four 3 disulfide-containing proteins, including hirudin, potato carboxypeptidase inhibitor, epidermal growth factor, and tick anticoagulant peptide. Their folding mechanism(s) differ from that of BPTI by 1) a higher degree of heterogeneity of 1 and 2-disulfide intermediates and 2) the presence of 3-disulfide scrambled isomers as folding intermediates. To search for the underlying causes of these diversities, we conducted kinetic analyses of the reductive unfolding of these five proteins. The experiment of reductive unfolding was designed to evaluate the relative stability and interdependence of disulfide bonds in the native protein. It is demonstrated here that among these five proteins, there exists a striking correlation between the mechanism(s) of reductive unfolding and that of oxidative folding. Those proteins with their native disulfide bonds reduced in a collective and simultaneous manner exhibit both a high degree of heterogeneity of folding intermediates and the accumulation of scrambled isomers along the folding pathway. A sequential reduction of the native disulfide bonds is associated with the presence of predominant intermediates with native- like structures. PMID- 10722658 TI - The transcriptional response of Saccharomyces cerevisiae to osmotic shock. Hot1p and Msn2p/Msn4p are required for the induction of subsets of high osmolarity glycerol pathway-dependent genes. AB - We have analyzed the transcriptional response to osmotic shock in the yeast Saccharomyces cerevisiae. The mRNA level of 186 genes increased at least 3-fold after a shift to NaCl or sorbitol, whereas that of more than 100 genes was at least 1.5-fold diminished. Many induced genes encode proteins that presumably contribute to protection against different types of damage or encode enzymes in glycerol, trehalose, and glycogen metabolism. Several genes, which encode poorly expressed isoforms of enzymes in carbohydrate metabolism, were induced. The high osmolarity glycerol (HOG) pathway is required for full induction of many but not all genes. The recently characterized Hot1p transcription factor is required for normal expression of a subset of the HOG pathway-dependent responses. Stimulated expression of the genes that required the general stress-response transcription factors Msn2p and Msn4p was also reduced in a hog1 mutant, suggesting that Msn2p/Msn4p might be regulated by the HOG pathway. The expression of genes that are known to be controlled by the mating pheromone response pathway was stimulated by osmotic shock specifically in a hog1 mutant. Inappropriate activation of the mating response may contribute to the growth defect of a hog1 mutant in high osmolarity medium. PMID- 10722659 TI - Nicotinic acid adenine dinucleotide phosphate-induced Ca(2+) release. Interactions among distinct Ca(2+) mobilizing mechanisms in starfish oocytes. AB - An intracellular mechanism activated by nicotinic acid adenine dinucleotide phosphate (NAADP(+)) contributes to intracellular Ca(2+) release alongside inositol 1,4,5-trisphosphate (Ins-P(3)) and ryanodine receptors. The NAADP(+) sensitive mechanism has been shown to be operative in sea urchin eggs, ascidian eggs, and pancreatic acinar cells. Furthermore, most mammalian cell types can synthesize NAADP(+), with nicotinic acid and NADP(+) as precursors. In this contribution, NAADP(+)-induced Ca(2+) release has been investigated in starfish oocytes. Uncaging of injected NAADP(+) induced Ca(2+) mobilization in both immature oocytes and in oocytes matured by the hormone 1-methyladenine (1-MA). The role of extracellular Ca(2+) in NAADP(+)-induced Ca(2+) mobilization, which was minor in immature oocytes, was instead essential in mature oocytes. Thus, the NAADP(+)-sensitive Ca(2+) pool, which is known to be distinct from those sensitive to inositol 1,4,5-trisphosphate or cyclic ADPribose, apparently migrated closer to (or became part of) the plasma membrane during the maturation process. Inhibition of both Ins-P(3) and ryanodine receptors, but not of either alone, substantially inhibited NAADP(+)-induced Ca(2+) mobilization in both immature and mature oocytes. The data also suggest that NAADP(+)-induced Ca(2+) mobilization acted as a trigger for Ca(2+) release via Ins-P(3) and ryanodine receptors. PMID- 10722660 TI - Influenza virus-induced NF-kappaB-dependent gene expression is mediated by overexpression of viral proteins and involves oxidative radicals and activation of IkappaB kinase. AB - Influenza A viruses are capable of inducing the expression of a variety of cytokine and proapoptotic genes in infected cells. The promoter regions of most of these genes harbor binding sites for the transcription factor NF-kappaB which is an important mediator of immune and inflammatory responses. Our present study is based on an observation that influenza A virus infection of cells stimulates transcriptional activation of the HIV-1 long terminal repeat (LTR) which harbors two regulatory NF-kappaB elements, and is aimed at identifying the molecular mechanisms involved in this process. We found that the expression of influenza virus hemagglutinin (HA), matrix protein (M), and nucleoprotein (NP), as single factors is sufficient to transcriptionally activate the HIV-1 LTR. This process is mediated by oxidative radicals because treatment of cells with pyrrolidine dithiocarbamate, a scavenger of such radicals, abolished the transactivating ability. Expression of different influenza proteins induces activation of NF kappaB-dependent gene expression but not transcriptional activation of an AP 1/Ets-dependent promoter, indicating a selectivity for NF-kappaB transactivation. Furthermore, influenza protein expression induces activation of IkappaB kinase (IKK). Accordingly coexpression of a catalytically inactive mutant of IKK abolishes influenza protein induced activation of NF-kappaB as well as HIV-1 LTR dependent reporter gene expression, suggesting that IKK is an important intermediate within this signaling process. Taken together, our results show that various influenza virus proteins act as viral transactivators to modulate transcriptional activity of kappaB-element harboring promoters such as the HIV LTR. PMID- 10722662 TI - The roles of carbohydrate chains of the beta-subunit on the functional expression of gastric H(+),K(+)-ATPase. AB - Gastric H(+),K(+)-ATPase consists of alpha and beta-subunits. The alpha-subunit is the catalytic subunit, and the beta-subunit is a glycoprotein stabilizing the alpha/beta complex in the membrane as a functional enzyme. There are seven putative N-glycosylation sites on the beta-subunit. In this study, we examined the roles of the carbohydrate chains of the beta-subunit by expressing the alpha subunit together with the beta-subunit in which one, several, or all of the asparagine residues in the N-glycosylation sites were replaced by glutamine. Removing any one of seven carbohydrate chains from the beta-subunit retained the H(+),K(+)-ATPase activity. The effects of a series of progressive removals of carbohydrate chains on the H(+),K(+)-ATPase activity were cumulative, and removal of all carbohydrate chains resulted in the complete loss of H(+), K(+)-ATPase activity. Removal of any single carbohydrate chain did not affect the alpha/beta assembly; however, little alpha/beta assembly was observed after removal of all the carbohydrate chains from the beta-subunit. In contrast, removal of three carbohydrate chains inhibited the surface delivery of the beta-subunit and the alpha-subunit assembled with the beta-subunit, indicating that the surface delivery mechanism is more dependent on the carbohydrate chains than the expression of the H(+),K(+)-ATPase activity and alpha/beta assembly. PMID- 10722661 TI - Isolation of Trypanosoma brucei CYC2 and CYC3 cyclin genes by rescue of a yeast G(1) cyclin mutant. Functional characterization of CYC2. AB - Two Trypanosoma brucei cyclin genes, CYC2 and CYC3, have been isolated by rescue of the Saccharomyces cerevisiae mutant DL1, which is deficient in CLN G(1) cyclin function. CYC2 encodes a 24-kDa protein that has sequence identity to the Neurospora crassa PREG1 and the S. cerevisiae PHO80 cyclin. CYC3 has the most sequence identity to mitotic B-type cyclins from a variety of organisms. Both CYC2 and CYC3 are single-copy genes and expressed in all life cycle stages of the parasite. To determine if CYC2 is found in a complex with previously identified trypanosome cdc2-related kinases (CRKs), the CYC2 gene was fused to the TY epitope tag, integrated into the trypanosome genome, and expressed under inducible control. CYC2ty was found to associate with an active trypanosome CRK complex since CYC2ty bound to leishmanial p12(cks1), and histone H1 kinase activity was detected in CYC2ty immune-precipitated fractions. Gene knockout experiments provide evidence that CYC2 is an essential gene, and co-immune precipitations together with a two-hybrid interaction assay demonstrated that CYC2 interacts with CRK3. The CRK3 x CYC2ty complex, the first cyclin-dependent kinase complex identified in trypanosomes, was localized by immune fluorescence to the cytoplasm throughout the cell cycle. PMID- 10722663 TI - Sp1/Sp3 and PU.1 differentially regulate beta(5) integrin gene expression in macrophages and osteoblasts. AB - Murine osteoclast precursors and osteoblasts express the integrin alpha(v)beta(5), the appearance of which on the cell surface is controlled by the beta(5), and not the alpha(v), subunit. Here, we show that a 173-base pair proximal region of the beta(5) promoter mediates beta(5) basal transcription in macrophage (osteoclast precursor)-like and osteoblast-like cells. DNase I footprinting reveal four regions (FP1-FP4) within the 173-base pair region, protected by macrophage nuclear extracts. In contrast, osteoblast nuclear extracts protect only FP1, FP2, and FP3. FP1, FP2, and FP3 bind Sp1 and Sp3 from both macrophage and osteoblast nuclear extracts. FP4 does not bind osteoblast proteins but binds PU.1 from macrophages. Transfection studies show that FP1 and FP2 Sp1/Sp3 sites act as enhancers in both MC3T3-E1 (osteoblast-like) and J774 (macrophage-like) cell lines, whereas the FP3 Sp1/Sp3 site serves as a silencer. Mutation of the FP2 Sp1/Sp3 site totally abolishes promoter activity in J774 cells, with only partial reduction in MC3T3-E1 cells. Finally, we demonstrate that PU.1 acts as a beta(5) silencer in J774 cells but plays no role in MC3T3-E1 cells. Thus, three Sp1/Sp3 sites regulate beta(5) gene expression in macrophages and osteoblast-like cells, with each element exhibiting cell-type and/or activation-suppression specificity. PMID- 10722664 TI - Intermolecular V(D)J recombination. AB - V(D)J recombination plays a prominent role in the generation of the antigen receptor repertoires of B and T lymphocytes. It is also likely to be involved in the formation of chromosomal translocations, some of which may result from interchromosomal recombination. We have investigated the potential of the V(D)J recombination machinery to perform intermolecular recombination between two plasmids, either unlinked or linked by catenation. In either case, recombination occurs in trans to yield signal and coding joints, and the results do not support the existence of a mechanistic block to the formation of coding joints in trans. Instead, we observe that linearization of the substrate, which does not alter the cis or trans status of the recombination signals, causes a specific and dramatic reduction in coding joint formation. This unexpected result leads us to propose a "release and recapture" model for V(D)J recombination in which coding ends are frequently released from the postcleavage complex and the efficiency of coding joint formation is influenced by the efficiency with which such ends are recaptured by the complex. This implies the existence of mechanisms, operative during recombination of chromosomal substrates, that act to prevent coding end release or to facilitate coding end recapture. PMID- 10722665 TI - Identification and cDNA cloning of alveolin, an extracellular metalloproteinase, which induces chorion hardening of medaka (Oryzias latipes) eggs upon fertilization. AB - Chorion hardening is triggered by the contents of cortical alveoli that are released upon fertilization of medaka (Oryzias latipes) eggs. We purified the chorion hardening-inducing activity as a single protein from the exudate of cortical alveoli of medaka eggs. This activity was co-purified with proteolytic activity of the chorion protein ZI-1,2. Based on the amino acid sequence of purified protein, we cloned the cDNA of this protein from a medaka ovarian cDNA library. Sequence analyses revealed typical sequence features, a zinc-binding motif and a methionine turn motif, of the astacin metalloproteinase family. We termed this protein "alveolin." Alveolin has a molecular mass of 21.5 kDa deduced by the amino acid sequence and neutral optimal pH range. Alveolin hydrolyzes ZI 1,2. Alveolin activity was strongly inhibited by metal-chelating agents but not by various proteinase inhibitors. To our knowledge, this is the first description of the isolation and identification of the chorion hardening-inducing factor from cortical alveoli exudate of teleost eggs. PMID- 10722666 TI - Requirement of divalent galactoside-binding activity of ecalectin/galectin-9 for eosinophil chemoattraction. AB - We have previously isolated and cloned a novel eosinophil chemoattractant (ECA) from a human T-cell-derived expression library. This ECA, termed ecalectin, is a variant of human galectin-9, a member of a beta-galactoside binding animal lectin family, which contains two conserved carbohydrate recognition domains (CRDs). In the present study, we addressed whether carbohydrate binding activity is required for the ECA activity of ecalectin and whether both CRDs are essential for this activity. Recombinant full-length wild-type ecalectin (ecalectin-WT) and N terminal and C-terminal CRD (ecalectin-NT and -CT, respectively) were generated. All of these recombinant proteins exhibited affinity for lactose, a property shared by galectins, but ecalectin-WT exhibited substantially higher hemagglutination activities than ecalectin-NT and -CT. Furthermore, ecalectin-WT showed over 100-fold higher ECA activity than ecalectin-NT and -CT; combination of recombinant domain fragments did not reconstitute the ECA and hemagglutination activities of the full-length protein. ECA activity of ecalectin-WT was inhibited by lactose in a dose-dependent manner. Site-directed mutation of positions Arg(65) of ecalectin-NT and Arg(239) of ecalectin-CT to an aspartic acid residue resulted in the loss of both lactose-binding and ECA activities. We conclude that divalent galactoside-binding activity is required for eosinophil chemoattraction by ecalectin. PMID- 10722667 TI - Mex67p mediates nuclear export of a variety of RNA polymerase II transcripts. AB - Mex67p is essential for nuclear poly(A)(+) RNA export in yeast, but which specific transcripts are transported by Mex67p is not known. We observed that thermosensitive mex67-5 cells do not produce a heat shock response at 37 degrees C but will induce heat shock proteins (Hsp) (e.g. Hsp104p and Hsp70p) when shifted back from the restrictive to permissive temperature (30 degrees C). This memory of a previous heat stress in mex67-5 cells could be explained if HSP mRNAs accumulated inside the nucleus during heat shock and were exported and translated in the cytoplasm on return to the permissive temperature. To test this hypothesis, nuclear export of heat shock mRNAs was directly analyzed by in situ hybridization using fluorescent-labeled oligonucleotide probes specific for SSA transcripts. This revealed that Mex67p is required for nuclear export of heat shock mRNAs. Furthermore, other polymerase II transcripts encoding the transcriptional repressor ASH1 and the glycolytic enzyme PGK1 are shown to require Mex67p for their export into the cytoplasm. Thus, Mex67p is an mRNA export factor for a broad range of polymerase II transcripts. PMID- 10722668 TI - Down-regulation of beta-catenin by the colorectal tumor suppressor APC requires association with Axin and beta-catenin. AB - The tumor suppressor adenomatous polyposis coli (APC) is mutated in familial adenomatous polyposis and in sporadic colorectal tumors. APC forms a complex with beta-catenin, Axin, and glycogen synthase kinase-3beta and induces the degradation of beta-catenin. In the present study, we examined whether APC association with Axin is required for degradation of beta-catenin. We found that a fragment of APC that induces beta-catenin degradation was rendered inactive by disruption of its Axin-binding sites. Also, overexpression of an Axin fragment spanning the regulator of the G-protein signaling domain inhibited APC-mediated beta-catenin degradation. An APC fragment with mutated beta-catenin-binding sites but intact Axin-binding sites also failed to induce degradation of beta-catenin. These results suggest that APC requires interaction with Axin and beta-catenin to down-regulate beta-catenin. PMID- 10722669 TI - Kinetic and pharmacological properties of cloned human equilibrative nucleoside transporters, ENT1 and ENT2, stably expressed in nucleoside transporter-deficient PK15 cells. Ent2 exhibits a low affinity for guanosine and cytidine but a high affinity for inosine. AB - We stably transfected the cloned human equilibrative nucleoside transporters 1 and 2 (hENT1 and hENT2) into nucleoside transporter-deficient PK15NTD cells. Although hENT1 and hENT2 are predicted to be 50-kDa proteins, hENT1 runs as 40 kDa and hENT2 migrates as 50 and 47 kDa on SDS-polyacrylamide gel electrophoresis. Peptide N-glycosidase F and endoglycosidase H deglycosylate hENT1 to 37 kDa and hENT2 to 45 kDa. With hENT1 being more sensitive, there is a 7000-fold and 71-fold difference in sensitivity to nitrobenzylthioinosine (NBMPR) (IC(50), 0.4 +/- 0.1 nM versus 2.8 +/- 0.3 microM) and dipyridamole (IC(50), 5.0 +/- 0.9 nM versus 356 +/- 13 nM), respectively. [(3)H]NBMPR binds to ENT1 cells with a high affinity K(d) of 0.377 +/- 0.098 nM, and each ENT1 cell has 34,000 transporters with a turnover number of 46 molecules/s for uridine. Although both transporters are broadly selective, hENT2 is a generally low affinity nucleoside transporter with 2.6-, 2.8-, 7. 7-, and 19.3-fold lower affinity than hENT1 for thymidine, adenosine, cytidine, and guanosine, respectively. In contrast, the affinity of hENT2 for inosine is 4-fold higher than hENT1. The nucleobase hypoxanthine inhibits [(3)H]uridine uptake by hENT2 but has minimal effect on hENT1. Taken together, these results suggest that hENT2 might be important in transporting adenosine and its metabolites (inosine and hypoxanthine) in tissues such as skeletal muscle where ENT2 is predominantly expressed. PMID- 10722670 TI - Identification of a lumenal sequence specifying the assembly of Emp24p into p24 complexes in the yeast secretory pathway. AB - The p24 proteins are transmembrane proteins of the endomembrane system that play a poorly defined role in vesicle traffic between the endoplasmic reticulum and the Golgi apparatus. Various lines of evidence indicate that p24 proteins fall into four subfamilies (alpha, beta, gamma, and delta) and that tetramers are assembled containing one representative from each subfamily; however, the nature of the protein-protein interactions within these hetero-oligomers is unknown. We have identified a lumenal segment of yeast p24beta (Emp24p) that is necessary for its assembly into p24 complexes. Replacement of 52 C-terminal residues of Emp24p with the corresponding sequence from Erv25p (p24delta) generates a chimeric protein able to replace Emp24p in p24 complexes that retain partial function in vivo, ruling out a role for the transmembrane and cytosolic domains in specifying p24 interactions. Substitution of a further 50 residues, encompassing a heptad repeat region, abolishes the ability of the chimera to replace Emp24p but instead creates a protein that resembles its Erv25p parent in its requirement for stabilization by Emp24p. These data point to a role for coiled-coil interactions in directing subfamily-specific assembly of p24 oligomers that project into the lumen of transport vesicles, where they may act to exclude secretory cargo from coat protein complex type I-coated retrograde transport vesicles. PMID- 10722671 TI - Regulation of calcium-sensitive tyrosine kinase Pyk2 by angiotensin II in endothelial cells. Roles of Yes tyrosine kinase and tyrosine phosphatase SHP-2. AB - Calcium-sensitive tyrosine kinase Pyk2 has been implicated in the regulation of ion channels, cellular adhesion, and mitogenic and hypertrophic reactions. In this study, we have investigated the regulation of Pyk2 by angiotensin II (Ang II) in pulmonary vein endothelial cells. We found that the Ang II-induced tyrosine phosphorylation of Pyk2, which requires the activity of Src family kinase, was specifically regulated by the Src family kinase member, Yes kinase. Moreover, we identified for the first time the constitutive association of Pyk2 with an Src homology 2 (SH2) domain-containing tyrosine phosphatase SHP-2. SHP-2 interacts with Pyk2 through a region other than its SH2 domains. Pyk2 can be dephosphorylated in vitro in SHP-2 immunoprecipitates and in intact cells expressing an NH(2) terminus-truncated form of SHP-2, which lacks the two SH2 domains but has an enhanced phosphatase activity. Ang II activates the endogenous SHP-2. Finally, the SHP-2-mediated dephosphorylation of Pyk2 correlates with the negative effect of SHP-2 on the Ang II-induced activation of extracellular signal regulated kinase and c-Jun NH(2)-terminal kinase. Thus, the balance of Pyk2 tyrosine phosphorylation in response to Ang II is controlled by Yes kinase and by a tyrosine phosphatase SHP-2 in endothelial cells. PMID- 10722672 TI - Hrs1/Med3 is a Cyc8-Tup1 corepressor target in the RNA polymerase II holoenzyme. AB - The Srb/Mediator, a multisubunit subcomplex of the RNA polymerase II (RNA pol II) holoenzyme has been proposed to function as a control panel regulating transcription in response to gene-specific activator proteins. In this report, we identify the Mediator subunit Hrs1/Med3 as a physical target for Cyc8-Tup1, a yeast transcriptional corepressor. Two-hybrid and glutathione S-transferase interaction assays show that Hrs1 can associate directly with Cyc8-Tup1. Moreover, affinity chromatography experiments, using yeast protein extracts, reveal that Cyc8-Tup1 co-purifies with Hrs1 and with additional Mediator subunits in a Hrs1-dependent manner. These observations suggest that Cyc8-Tup1 contacts the Mediator complex via its interaction with the Hrs1 subunit. Further on, genetic analysis indicates that increased Hrs1 dosage can alleviate Cyc8-Tup1 mediated repression, suggesting that Hrs1/Mediator's function is inhibited upon its interaction with Cyc8-Tup1. Finally, artificial holoenzyme recruitment assays support a model by which the contact between the corepressor and the Hrs1/Mediator may prevent pol II holoenzyme recruitment to the core promoter. These data, together with previous genetic evidence, establish a functional and physical interaction between the Cyc8-Tup1 corepressor and the RNA pol II holoenzyme and support a central role of the Mediator complex in transcriptional repression. PMID- 10722673 TI - A Xestospongin C-sensitive Ca(2+) store is required for cAMP-induced Ca(2+) influx and cAMP oscillations in Dictyostelium. AB - Xestospongin C (XeC) is known to bind to the inositol 1,4, 5-trisphosphate (IP(3))-sensitive store in mammalian cells and to inhibit IP(3)- and thapsigargin induced Ca(2+) release. In this study we show that this is also true for Dictyostelium. In addition, XeC inhibited Ca(2+) uptake into purified vesicle fractions and induced Ca(2+) release. This suggests that, in the case of Dictyostelium, XeC opens rather than plugs the IP(3) receptor channel as was proposed for mammalian cells (Gafni, J., Munsch, J. A. , Lam, T. H., Catlin, M. C., Costa, L. G., Molinski, T. F., and Pessah, I. N. (1997) Neuron 19, 723-733). In order to elucidate the function of the XeC-sensitive Ca(2+) store in Dictyostelium during differentiation, we applied XeC to the cells and found that it caused a time-dependent increase of basal [Ca(2+)](i) and inhibited cAMP induced Ca(2+) influx in single cells as well as in cell suspensions. Moreover, XeC blocked light scattering spikes and pulsatile cAMP signaling. PMID- 10722674 TI - Purification of the serine palmitoyltransferase complex responsible for sphingoid base synthesis by using affinity peptide chromatography techniques. AB - Serine palmitoyltransferase (SPT), a membrane-bound enzyme of the endoplasmic reticulum, catalyzes the condensation of palmitoyl coenzyme A (CoA) and L-serine to produce 3-ketodihydrosphingosine. This enzyme contains at least two different subunits, named the LCB1 and LCB2 proteins. In the present study, we expressed a FLAG- and His(6) peptide-tagged version of the hamster LCB1 protein in a Chinese hamster ovary cell mutant strain lacking the endogenous LCB1 subunit and purified SPT from the cells near to homogeneity by affinity peptide chromatography. The endogenous LCB2 protein was co-purified with the tagged LCB1 protein in purification of SPT. In various aspects, including optimum pH, acyl-CoA specificity, and sphingofungin sensitivity, the activity of purified SPT was consistent with the activity detected in lysates of wild-type Chinese hamster ovary cells. The optimum concentration of palmitoyl-CoA for 3 ketodihydrosphingosine formation by purified SPT was approximately 25 microM, and the apparent K(m) of L-serine was 0.28 mM. Competition analysis of the SPT reaction with various serine analogs showed that all of the amino, carboxyl, and hydroxyl groups of L-serine were responsible for the substrate recognition of the enzyme. SDS-polyacrylamide gel electrophoretic analysis of purified SPT, together with immunoprecipitation analysis of metabolically labeled LCB proteins, strongly suggested that the SPT enzyme consisted of the LCB1 and LCB2 proteins with a stoichiometry of 1:1. PMID- 10722675 TI - Mechanisms of hepatic very low density lipoprotein overproduction in insulin resistance. Evidence for enhanced lipoprotein assembly, reduced intracellular ApoB degradation, and increased microsomal triglyceride transfer protein in a fructose-fed hamster model. AB - A novel animal model of insulin resistance, the fructose-fed Syrian golden hamster, was employed to investigate the mechanisms mediating the overproduction of very low density lipoprotein (VLDL) in the insulin resistant state. Fructose feeding for a 2-week period induced significant hypertriglyceridemia and hyperinsulinemia, and the development of whole body insulin resistance was documented using the euglycemic-hyperinsulinemic clamp technique. In vivo Triton WR-1339 studies showed evidence of VLDL-apoB overproduction in the fructose-fed hamster. Fructose feeding induced a significant increase in cellular synthesis and secretion of total triglyceride (TG) as well as VLDL-TG by primary hamster hepatocytes. Increased TG secretion was accompanied by a 4.6-fold increase in VLDL-apoB secretion. Enhanced stability of nascent apoB in fructose-fed hepatocytes was evident in intact cells as well as in a permeabilized cell system. Analysis of newly formed lipoprotein particles in hepatic microsomes revealed significant differences in the pattern and density of lipoproteins, with hepatocytes derived from fructose-fed hamsters having higher levels of luminal lipoproteins at a density of VLDL versus controls. Immunoblot analysis of the intracellular mass of microsomal triglyceride transfer protein, a key enzyme involved in VLDL assembly, showed a striking 2.1-fold elevation in hepatocytes derived from fructose-fed versus control hamsters. Direct incubation of hamster hepatocytes with various concentrations of fructose failed to show any direct stimulation of its intracellular stability or extracellular secretion, further supporting the notion that the apoB overproduction in the fructose-fed hamster may be related to the fructose-induced insulin resistance in this animal model. In summary, hepatic VLDL-apoB overproduction in fructose-fed hamsters appears to result from increased intracellular stability of nascent apoB and an enhanced expression of MTP, which act to facilitate the assembly and secretion of apoB containing lipoprotein particles. PMID- 10722676 TI - Stepwise regulated chromatin assembly of MCM2-7 proteins. AB - Acquisition of the competence to replicate requires the assembly of the MCM2-7 (minichromosome maintenance) protein complex onto pre-replicative chromatin, a step of the licensing reaction. This step is thought to occur through binding of a heterohexameric MCM complex containing the six related MCM subunits. Here we show that assembly of the MCM complex onto pre-replicative chromatin occurs through sequential stabilization of specific MCM subunits. Inhibition of licensing with 6-dimethylaminopurine results in chromatin containing specifically bound MCM4 and MCM6. A similar result was obtained by interference of the assembly reaction with an MCM3 antibody. The presence of chromatin-bound MCM intermediates was confirmed by reconstitution experiments in vitro with purified proteins and by the observation of an ordered association of MCM subunits with chromatin. These results indicate that the assembly of the MCM complex onto pre replicative chromatin is regulated at the level of distinct subunits, suggesting an additional regulatory step in the formation of pre-replication complexes. PMID- 10722677 TI - 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced degradation of aryl hydrocarbon receptor (AhR) by the ubiquitin-proteasome pathway. Role of the transcription activaton and DNA binding of AhR. AB - Activation of the aryl hydrocarbon receptor (AhR) by 2,3,7, 8-tetrachlorodibenzo p-dioxin (TCDD), a potent agonist of AhR, induces a marked reduction in steady state AhR. To analyze the mechanism of regulation of ligand-activated AhR, we examined the biochemical pathway and function of the down-regulation of the receptor by TCDD. Pulse-chase experiments reveal that TCDD shortens the half-life (t1/2) of AhR from 28 to 3 h in mouse hepatoma cells. Inhibitors of the 26 S proteasome, lactacystin and MG132, block the TCDD-induced turnover of AhR. The TCDD-induced degradation of AhR involves ubiquitination of the AhR protein, because (a) TCDD induces formation of high molecular weight, ubiquitinated AhR and (b) degradation of AhR is inhibited in ts20 cells, which bear a temperature sensitive mutation in the ubiquitin-activating enzyme E1, at a nonpermissive temperature. Inhibition of proteasomal degradation of AhR increases the amount of the nuclear AhR.Arnt complex and "superinduces" the expression of endogenous CYP1A1 gene by TCDD, indicating that the proteasomal degradation of AhR serves as a mechanism for controlling the activity of the activated receptor. We also show that deletion of the transcription activation domain of AhR abolishes the degradation, whereas a mutation in the DNA-binding region of AhR or Arnt reduces the degradation; these data implicate the transcription activation domain and DNA binding in AhR degradation. Our findings provide new insights into the regulation of TCDD-activated AhR through ubiquitin-mediated protein degradation. PMID- 10722678 TI - Hsc70 chaperones clathrin and primes it to interact with vesicle membranes. AB - When Hsc70 uncoats clathrin-coated vesicles in an auxilin- and ATP-dependent reaction, a single round of rapid uncoating occurs followed by very slow steady state uncoating. We now show that this biphasic time course occurs because Hsc70 sequentially forms two types of complex with the dissociated clathrin triskelions. The first round of clathrin uncoating is driven by formation of a pre-steady-state assembly protein (AP)-clathrin-Hsc70-ADP complex. Then, following exchange of ADP with ATP, a steady-state AP-clathrin-Hsc70-ATP complex forms that ties up Hsc70, preventing further uncoating. This steady-state complex forms only during uncoating in the presence of APs; in the absence of APs, Hsc70 rapidly dissociates from the uncoated clathrin and continues to carry out uncoating. Whether it is complexed with ATP or ADP, the steady-state complex has very different properties from the pre-steady-state complex in that it cannot be immunoprecipitated by anti-clathrin antibodies and is readily dissociated by fast protein liquid chromatography. Remarkably, when the steady-state complex is incubated with uncoated vesicle membranes in ATP, the pre-steady-state complex reforms, suggesting that the clathrin triskelions in the steady-state complex rebind to the membranes and are again uncoated by Hsc70. We propose that Hsc70 not only uncoats clathrin but also chaperones it to prevent it from inappropriately polymerizing in the cell cytosol and primes it to reform clathrin coated pits. PMID- 10722679 TI - The C-terminal domain of MutY glycosylase determines the 7,8-dihydro-8-oxo guanine specificity and is crucial for mutation avoidance. AB - Escherichia coli MutY is an adenine DNA glycosylase active on DNA substrates containing A/G, A/8-oxoG, or A/C mismatches and also has a weak guanine glycosylase activity on G/8-oxoG-containing DNA. The N-terminal domain of MutY, residues 1-226, has been shown to retain catalytic activity. Substrate binding, glycosylase, and Schiff base intermediate formation activities of the truncated and intact MutY were compared. MutY has high binding affinity with 8-oxoG when mispaired with A, G, T, C, or inosine. The truncated protein has more than 18 fold lower affinities for binding various 8-oxoG-containing mismatches when compared with intact MutY. MutY catalytic activity toward A/8-oxoG-containing DNA is much faster than that on A/G-containing DNA whereas deletion of the C-terminal domain reduces its catalytic preference for A/8-oxoG-DNA over A/G-DNA. MutY exerts more inhibition on the catalytic activity of MutM (Fpg) protein than does truncated MutY. The tight binding of MutY with GO mispaired with T, G, and apurinic/apyrimidinic sites may be involved in the regulation of MutM activity. An E. coli mutY strain that produces an N-terminal 249-residue truncated MutY confers a mutator phenotype. These findings strongly suggest that the C-terminal domain of MutY determines the 8-oxoG specificity and is crucial for mutation avoidance by oxidative damage. PMID- 10722680 TI - Mechanism of insulin resistance in A-ZIP/F-1 fatless mice. AB - Insulin resistance is a major factor in the pathogenesis of type 2 diabetes and may be related to alterations in fat metabolism. Fatless mice have been created using dominant-negative protein (A-ZIP/F-1) targeted gene expression in the adipocyte and shown to develop diabetes. To understand the mechanism responsible for the insulin resistance in these mice, we conducted hyperinsulinemic euglycemic clamps in awake fatless and wild type littermates before the development of diabetes and examined insulin action and signaling in muscle and liver. We found the fatless mice to be severely insulin-resistant, which could be attributed to defects in insulin action in muscle and liver. Both of these abnormalities were associated with defects in insulin activation of insulin receptor substrate-1 and -2-associated phosphatidylinositol 3-kinase activity and a 2-fold increase in muscle and liver triglyceride content. We also show that upon transplantation of fat tissue into these mice, triglyceride content in muscle and liver returned to normal as does insulin signaling and action. In conclusion, these results suggest that the development of insulin resistance in type 2 diabetes may be due to alterations in the partitioning of fat between the adipocyte and muscle/liver leading to accumulation of triglyceride in the latter tissues with subsequent impairment of insulin signaling and action. PMID- 10722681 TI - Expression and functional analysis of Apaf-1 isoforms. Extra Wd-40 repeat is required for cytochrome c binding and regulated activation of procaspase-9. AB - Apaf-1 is an important apoptotic signaling molecule that can activate procaspase 9 in a cytochrome c/dATP-dependent fashion. Alternative splicing can create an NH(2)-terminal 11-amino acid insert between the caspase recruitment domain and ATPase domains or an additional COOH-terminal WD-40 repeat. Recently, several Apaf-1 isoforms have been identified in tumor cell lines, but their expression in tissues and ability to activate procaspase-9 remain poorly characterized. We performed analysis of normal tissue mRNAs to examine the relative expression of the Apaf-1 forms and identified Apaf-1XL, containing both the NH(2)-terminal and COOH-terminal inserts, as the major RNA form expressed in all tissues tested. We also identified another expressed isoform, Apaf-1LN, containing the NH(2) terminal insert, but lacking the additional WD-40 repeat. Functional analysis of all identified Apaf-1 isoforms demonstrated that only those with the additional WD-40 repeat activated procaspase 9 in vitro in response to cytochrome c and dATP, while the NH(2)-terminal insert was not required for this activity. Consistent with this result, in vitro binding assays demonstrated that the additional WD-40 repeat was also required for binding of cytochrome c, subsequent Apaf-1 self-association, binding to procaspase-9, and formation of active Apaf-1 oligomers. These experiments demonstrate the expression of multiple Apaf-1 isoforms and show that only those containing the additional WD-40 repeat bind and activate procaspase-9 in response to cytochrome c and dATP. PMID- 10722683 TI - The Lurcher mutation of an alpha-amino-3-hydroxy-5-methyl- 4-isoxazolepropionic acid receptor subunit enhances potency of glutamate and converts an antagonist to an agonist. AB - A point mutation of the GluRdelta2 (A654T) glutamate receptor subunit converts it into a functional channel, and a spontaneous mutation at this site is thought to be responsible for the neurodegeneration of neurons in the Lurcher mouse. This mutation is located in a hydrophobic region of the M3 domain of this subunit, and this alanine is conserved throughout many of the glutamate receptors. We show here that site-directed mutagenesis of the homologous alanine (A636T; GluR1-L(c)) in the GluR1 AMPA receptor subunit alters its channel properties. The apparent potencies of both kainate and glutamate were increased 85- and 2000-fold, respectively. Furthermore, 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX)was converted from a competitive antagonist into a potent agonist. Our results demonstrate that a single amino acid within or near the putative second transmembrane region of the GluR1 subunit is critical for the binding/gating properties of this AMPA receptor. PMID- 10722682 TI - Interaction between the conserved region in the C-terminal domain of GRK2 and rhodopsin is necessary for GRK2 to catalyze receptor phosphorylation. AB - The C-terminal domain of G protein-coupled receptor kinases (GRKs) consists of a conserved region and a variable region, and the variable region has been shown to direct the membrane translocation of cytosolic enzymes. The present work has revealed that the C-terminal domain may also be involved in kinase-receptor interaction that is primarily mediated by the conserved region. Truncation of the C-terminal domain or deletion of the conserved region in this domain of GRK2 resulted in a complete loss of its ability to phosphorylate rhodopsin and in an obvious decrease in its sensitivity to receptor-mediated phosphorylation of a peptide substrate. On the contrary, deletion of the betagamma subunit binding region in the C-terminal domain of GRK2 did not significantly alter the ability of the enzyme to phosphorylate rhodopsin. In addition, the recombinant proteins that represent the C-terminal domain and the conserved region of GRK2 could inhibit GRK2-mediated phosphorylation of rhodopsin and receptor-mediated activation of GRK2 but not GRK2-mediated phosphorylation of the peptide substrate. Furthermore, the conserved region as well as the C-terminal domain could directly bind rhodopsin in vitro. These results indicate that the C terminal domain, or more precisely, the conserved region of this domain, is important for enzyme-receptor interaction and that this interaction is required for GRK2 to catalyze receptor phosphorylation. PMID- 10722684 TI - Kinetic studies of cAMP-induced allosteric changes in cyclic AMP receptor protein from Escherichia coli. AB - Cyclic AMP receptor protein (CRP) regulates the expression of several genes in Escherichia coli. The ability of CRP to bind specific DNA sequences and stimulate transcription is achieved as result of binding of an allosteric ligand: cAMP. Stopped-flow fluorimetry was employed to study the kinetics of the conformational changes in CRP induced by cAMP binding to high and low affinity receptor sites. Results of experiments using CRP labeled at Cys-178 with 1,5-I-AENS indicate change in conformation of the helix-turn-helix, occurring after the formation of CRP-cAMP(2) complex, i.e. after saturation of the high affinity sites. The observed conformational change occurs according to sequential model of allostery and is described by rate constants: k(c) = 9.7 +/- 0.1 s(-1) and k(-c) = 0.31 +/- 0.05 s(-1), for the forward and backward reaction, respectively. Results of experiments monitored using CRP intrinsic fluorescence suggest that conformational change precedes the formation of CRP-cAMP(4) complex and results from displacement of equilibrium between two forms of CRP-cAMP(2), caused by binding of cAMP to low affinity sites of one of these forms only. The observed conformational change occurs according to concerted model of allostery and is described by rate constants: k(on) = 28 +/- 1.5 s(-1) and k(off) = 75.5 +/- 3 s( 1). Results of experiments using single-tryptophan-containing CRP mutants indicate that Trp-85 is mainly responsible for the observed total change in intrinsic fluorescence of wild-type CRP. PMID- 10722685 TI - Selenite negatively regulates caspase-3 through a redox mechanism. AB - Selenium, an essential biological trace element, exerts its modulatory effects in a variety of cellular events including cell survival and death. In our study we observed that selenite protects HEK293 cells from cell death induced by ultraviolet B radiation (UVB). Exposure of HEK293 cells to UVB radiation resulted in the activation of caspase-3-like protease activity, and pretreatment of the cells with z-DEVD-fmk (N-benzyloxycarbonyl-Asp-Glu-Val-Asp-fluoromethylketone), a caspase-3 inhibitor, prevented UVB-induced cell death. Interestingly, enzymatic activity of caspase-3-like protease in cell lysates of UVB-exposed cells was repressed in vitro by the presence of selenite. Selenite also inhibited the in vitro activity of purified recombinant caspase-3 in cleaving Ac-DEVD-pNA (N acetyl-Asp-Glu-Asp-p-nitroanilide) or ICAD(L) (inhibitor of a caspase-activated deoxyribonuclease) and in the induction of DNA fragmentation. The inhibitory action of selenite on a recombinant active caspase-3 could be reversed by sulfhydryl reducing agents, such as dithiothreitol and beta-mercaptoethanol. Furthermore, pretreatment of cells with selenite suppressed the stimulation of the caspase-3-like protease activity in UVB-exposed cells, whereas dithiothreitol and beta-mercaptoethanol reversed this suppression of the enzymatic activity. Taken together, our data suggest that selenite inhibits caspase-3-like protease activity through a redox mechanism and that inhibition of caspase-3-like protease activity may be the mechanism by which selenite exerts its protective effect against UVB-induced cell death. PMID- 10722686 TI - The mechanism of GTP hydrolysis by Ras probed by Fourier transform infrared spectroscopy. AB - Time-resolved Fourier transform infrared spectroscopy (FTIR) in combination with photo-induced release of (18)O-labeled caged nucleotide has been employed to address mechanistic issues of GTP hydrolysis by Ras protein. Infrared spectroscopy of Ras complexes with nitrophenylethyl (NPE)-[alpha-(18)O(2)]GTP, NPE-[beta-(18)O(4)]GTP, or NPE-[gamma-(18)O(3)]GTP upon photolysis or during hydrolysis afforded a substantially improved mode assignment of phosphoryl group absorptions. Photolysis spectra of hydroxyphenylacyl-GTP and hydroxyphenylacyl GDP bound to Ras and several mutants, Ras(Gly(12))-Mn(2+), Ras(Pro(12)), Ras(Ala(12)), and Ras(Val(12)), were obtained and yielded valuable information about structures of GTP or GDP bound to Ras mutants. IR spectra revealed stronger binding of GDP beta-PO(3)(2-) moiety by Ras mutants with higher activity, suggesting that the transition state is largely GDP-like. Analysis of the photolysis and hydrolysis FTIR spectra of the [beta-nonbridge-(18)O(2), alphabeta bridge-(18)O]GTP isotopomer allowed us to probe for positional isotope exchange. Such a reaction might signal the existence of metaphosphate as a discrete intermediate, a key species for a dissociative mechanism. No positional isotope exchange was observed. Overall, our results support a concerted mechanism, but the transition state seems to have a considerable amount of dissociative character. This work demonstrates that time-resolved FTIR is highly suitable for monitoring positional isotope exchange and advantageous in many aspects over previously used methods, such as (31)P NMR and mass spectrometry. PMID- 10722687 TI - Different catalytically competent arrangements of arachidonic acid within the cyclooxygenase active site of prostaglandin endoperoxide H synthase-1 lead to the formation of different oxygenated products. AB - Arachidonic acid is converted to prostaglandin G(2) (PGG(2)) by the cyclooxygenase activities of prostaglandin endoperoxide H synthases (PGHSs) 1 and 2. The initial, rate-limiting step is abstraction of the 13-proS hydrogen from arachidonate which, for PGG(2) formation, is followed by insertion of O(2) at C 11, cyclization, and a second O( 2) insertion at C-15. As an accompaniment to ongoing structural studies designed to determine the orientation of arachidonate in the cyclooxygenase site, we analyzed the products formed from arachidonate by (a) solubilized, partially purified ovine (o) PGHS-1; (b) membrane-associated, recombinant oPGHS-1; and (c) a membrane-associated, recombinant active site mutant (V349L oPGHS-1) and determined kinetic values for formation of each product. Native forms of oPGHS-1 produced primarily PGG(2) but also several monohydroxy acids, which, in order of abundance, were 11R-hydroxy-5Z, 8Z,12E,14Z eicosatetraenoic acid (11R-HETE), 15S-hydroxy-5Z,8Z,11Z, 13E-eicosatetraenoic acid (15S-HETE), and 15R-HETE. V349L oPGHS-1 formed primarily PGG(2), 15S-HETE, and 15R-HETE but only trace amounts of 11R-HETE. With native enzyme, the K(m) values for PGG(2), 11-HETE, and 15-HETE formation were each different (5.5, 12.1, and 19.4 microM, respectively); similarly, the K(m) values for PGG(2) and 15-HETE formation by V349L oPGHS-1 were different (11 and 5 microM, respectively). These results establish that arachidonate can assume at least three catalytically productive arrangements within the cyclooxygenase site of oPGHS-1 leading to PGG(2), 11R-HETE, and 15S-HETE and/or 15R-HETE, respectively. IC(50) values for inhibition of formation of the individual products by the competitive inhibitor, ibuprofen, were determined and found to be the same for a given enzyme form (i.e. 175 microM for oPGHS-1 and 15 microM for V349L oPGHS-1). These latter results are most simply rationalized by a kinetic model in which arachidonate forms various catalytically competent arrangements only after entering the cyclooxygenase active site. PMID- 10722688 TI - Urocortin protects against ischemic and reperfusion injury via a MAPK-dependent pathway. AB - Urocortin (UCN) is a peptide related to hypothalamic corticotrophin-releasing hormone and binds with high affinity to corticotrophin-releasing hormone receptor 2beta, which is expressed in the heart. In this study, we report that UCN prevented cell death when administered to primary cardiac myocyte cultures both prior to simulated hypoxia/ischemia and at the point of reoxygenation after simulated hypoxia/ischemia. UCN-mediated cell survival was measured by trypan blue exclusion, 3'-OH end labeling of DNA (TUNEL), annexin V, and fluorescence activated cell sorting. To explore the mechanisms that could be responsible for this effect, we investigated the involvement of MAPK-dependent pathways. UCN caused rapid phosphorylation of ERK1/2-p42/44, and PD98059, which blocks the MEK1 ERK1/2-p42/44 cascade, also inhibited the survival-promoting effect of UCN. Most important, UCN reduced damage in isolated rat hearts ex vivo subjected to regional ischemia/reperfusion, with the protective effect being observed when UCN was given either prior to ischemia or at the time of reperfusion after ischemia. This suggests a novel function of UCN as a cardioprotective agent that could act when given after ischemia, at reperfusion. PMID- 10722689 TI - Purification and magneto-optical spectroscopic characterization of cytoplasmic membrane and outer membrane multiheme c-type cytochromes from Shewanella frigidimarina NCIMB400. AB - Two membranous c-type cytochromes from the Fe(III)-respiring bacterium Shewanella frigidimarina NCIMB400, CymA and OmcA, have been purified and characterized by UV visible, magnetic circular dichroism, and electron paramagnetic resonance spectroscopies. The 20-kDa CymA is a member of the NapC/NirT family of multiheme cytochromes, which are invariably anchored to the cytoplasmic membrane of Gram negative bacteria, and are postulated to mediate electron flow between quinols and periplasmic redox proteins. CymA was found to contain four low-spin c-hemes, each with bis-His axial ligation, and midpoint reduction potentials of +10, -108, -136, and -229 mV. The 85-kDa OmcA is located at the outer membrane of S. frigidimarina NCIMB400, and as such might function as a terminal reductase via interaction with insoluble Fe(III) substrates. This putative role is supported by the finding that the protein was released into solution upon incubation of harvested intact cells at 25 degrees C, suggesting an attachment to the exterior face of the outer membrane. OmcA was revealed by magneto-optical spectrocopies to contain 10 low-spin bis-His ligated c-hemes, with the redox titer indicating two sets of near iso-potential components centered at -243 and -324 mV. PMID- 10722690 TI - Recruitment of a foreign quinone into the A(1) site of photosystem I. I. Genetic and physiological characterization of phylloquinone biosynthetic pathway mutants in Synechocystis sp. pcc 6803. AB - Genes encoding enzymes of the biosynthetic pathway leading to phylloquinone, the secondary electron acceptor of photosystem (PS) I, were identified in Synechocystis sp. PCC 6803 by comparison with genes encoding enzymes of the menaquinone biosynthetic pathway in Escherichia coli. Targeted inactivation of the menA and menB genes, which code for phytyl transferase and 1,4-dihydroxy-2 naphthoate synthase, respectively, prevented the synthesis of phylloquinone, thereby confirming the participation of these two gene products in the biosynthetic pathway. The menA and menB mutants grow photoautotrophically under low light conditions (20 microE m(-2) s(-1)), with doubling times twice that of the wild type, but they are unable to grow under high light conditions (120 microE m(-2) s(-1)). The menA and menB mutants grow photoheterotrophically on media supplemented with glucose under low light conditions, with doubling times similar to that of the wild type, but they are unable to grow under high light conditions unless atrazine is present to inhibit PS II activity. The level of active PS II per cell in the menA and menB mutant strains is identical to that of the wild type, but the level of active PS I is about 50-60% that of the wild type as assayed by low temperature fluorescence, P700 photoactivity, and electron transfer rates. PS I complexes isolated from the menA and menB mutant strains contain the full complement of polypeptides, show photoreduction of F(A) and F(B) at 15 K, and support 82-84% of the wild type rate of electron transfer from cytochrome c(6) to flavodoxin. HPLC analyses show high levels of plastoquinone-9 in PS I complexes from the menA and menB mutants but not from the wild type. We propose that in the absence of phylloquinone, PS I recruits plastoquinone-9 into the A(1) site, where it functions as an efficient cofactor in electron transfer from A(0) to the iron-sulfur clusters. PMID- 10722691 TI - Recruitment of a foreign quinone into the A(1) site of photosystem I. II. Structural and functional characterization of phylloquinone biosynthetic pathway mutants by electron paramagnetic resonance and electron-nuclear double resonance spectroscopy. AB - Electron paramagnetic resonance (EPR) and electron-nuclear double resonance studies of the photosystem (PS) I quinone acceptor, A(1), in phylloquinone biosynthetic pathway mutants are described. Room temperature continuous wave EPR measurements at X-band of whole cells of menA and menB interruption mutants show a transient reduction and oxidation of an organic radical with a g-value and anisotropy characteristic of a quinone. In PS I complexes, the continuous wave EPR spectrum of the photoaccumulated Q(-) radical, measured at Q-band, and the electron spin-polarized transient EPR spectra of the radical pair P700(+) Q(-), measured at X-, Q-, and W-bands, show three prominent features: (i) Q(-) has a larger g-anisotropy than native phylloquinone, (ii) Q(-) does not display the prominent methyl hyperfine couplings attributed to the 2-methyl group of phylloquinone, and (iii) the orientation of Q(-) in the A(1) site as derived from the spin polarization is that of native phylloquinone in the wild type. Electron spin echo modulation experiments on P700(+) Q(-) show that the dipolar coupling in the radical pair is the same as in native PS I, i.e. the distance between P700(+) and Q(-) (25.3 +/- 0.3 A) is the same as between P700(+) and A(1)(-) in the wild type. Pulsed electron-nuclear double resonance studies show two sets of resolved spectral features with nearly axially symmetric hyperfine couplings. They are tentatively assigned to the two methyl groups of the recruited plastoquinone-9, and their difference indicates a strong inequivalence among the two groups when in the A(1) site. These results show that Q (i) functions in accepting an electron from A(0)(-) and in passing the electron forward to the iron-sulfur clusters, (ii) occupies the A(1) site with an orientation similar to that of phylloquinone in the wild type, and (iii) has spectroscopic properties consistent with its identity as plastoquinone-9. PMID- 10722692 TI - JAB1 interacts with both the progesterone receptor and SRC-1. AB - JAB1 (Jun activation domain-binding protein-1) has previously been described as a coactivator of AP1 transcription factor. We show here, by yeast and mammalian two hybrid analyses and by pull-down experiments, that JAB1 also interacts with both the progesterone receptor (PR) and the steroid receptor coactivator 1 (SRC-1) and that it stabilizes PR-SRC-1 complexes. We also show that JAB1 potentiates the activity of a variety of transcription factors known to associate with SRC-1 (nuclear receptors, activator protein-1, and nuclear factor kappaB). This occurs without any modification of PR or SRC-1 concentration. JAB1 is a subunit of a large multiprotein complex that has been called the COP9 signalosome. The latter is present in plant and animal cells and has been shown to be involved in a variety of cellular mechanisms including transcription regulation, cell cycle control, and phosphorylation cascades. We now show that it is also involved in the mechanisms of action of nuclear receptors and of their coactivators. PMID- 10722693 TI - Pervanadate-induced nuclear factor-kappaB activation requires tyrosine phosphorylation and degradation of IkappaBalpha. Comparison with tumor necrosis factor-alpha. AB - Tumor necrosis factor activates nuclear transcription factor kappaB (NF-kappaB) by inducing serine phosphorylation of the inhibitory subunit of NF-kappaB (IkappaBalpha), which leads to its ubiquitination and degradation. In contrast, pervanadate (PV) activates NF-kappaB and induces tyrosine phosphorylation of IkappaBalpha (Singh, S., Darney, B. G., and Aggarwal, B. B. (1996) J. Biol. Chem. 271, 31049-31054; Imbert, V., Rupec, R. A., Antonia, L., Pahl, H. L., Traenckner, E. B.-M., Mueller-Dieckmann, C., Farahifar, D., Rossi, B., Auderger, P., Baeuerle, P. A., and Peyron, J.-F. (1996) Cell 86, 787-798). Whether PV also induces IkappaBalpha degradation and whether degradation is required for NF kappaB activation are not understood. We investigated the effect of PV-induced tyrosine phosphorylation on IkappaBalpha degradation and NF-kappaB activation. PV activated NF-kappaB, as determined by DNA binding, NF-kappaB-dependent reporter gene expression, and phosphorylation and degradation of IkappaBalpha. Maximum degradation of IkappaBalpha occurred at 180 min, followed by NF-kappaB-dependent IkappaBalpha resynthesis. N-Acetylleucylleucylnorlucinal, a proteasome inhibitor, blocked both IkappaBalpha degradation and NF-kappaB activation, suggesting that the IkappaBalpha degradation is required for NF-kappaB activation. PV did not induce serine phosphorylation of IkappaBalpha but induced phosphorylation at tyrosine residue 42. Unlike tumor necrosis factor (TNF), PV did not induce ubiquitination of IkappaBalpha. Like TNF, however, PV induced phosphorylation and degradation of IkappaBalpha, and subsequent NF-kappaB activation, which could be blocked by N-tosyl-L-phenylalanine chloromethyl ketone, calpeptin, and pyrrolidine dithiocarbomate, suggesting a close link between PV-induced NF-kappaB activation and IkappaBalpha degradation. Overall, our studies demonstrate that PV activates NF-kappaB, which, unlike TNF, requires tyrosine phosphorylation of IkappaBalpha and its degradation. PMID- 10722694 TI - Proteolytic processing of the C terminus of the alpha(1C) subunit of L-type calcium channels and the role of a proline-rich domain in membrane tethering of proteolytic fragments. AB - Although most L-type calcium channel alpha(1C) subunits isolated from heart or brain are approximately 190-kDa proteins that lack approximately 50 kDa of the C terminus, the C-terminal domain is present in intact cells. To test the hypothesis that the C terminus is processed but remains functionally associated with the channels, expressed, full-length alpha(1C) subunits were cleaved in vitro by chymotrypsin to generate a 190-kDa C-terminal truncated protein and C terminal fragments of 30-56 kDa. These hydrophilic C-terminal fragments remained membrane-associated. A C-terminal proline-rich domain (PRD) was identified as the mediator of membrane association. The alpha(1C) PRD bound to SH3 domains in Src, Lyn, Hck, and the channel beta(2) subunit. Mutant alpha(1C) subunits lacking either approximately 50 kDa of the C terminus or the PRD produced increased barium currents through the channels, demonstrating that these domains participate in the previously described (Wei, X., Neely, a., Lacerda, A. E. Olcese, r., Stefani, E., Perez-Reyes, E., and Birnbaumer, L. (1994) J. Biol. Chem. 269, 1635-1640) inhibition of channel function by the C terminus. PMID- 10722695 TI - Apolipoprotein E is resistant to intracellular degradation in vitro and in vivo. Evidence for retroendocytosis. AB - Apolipoprotein E (apoE) is an important determinant for the uptake of triglyceride-rich lipoproteins and emulsions by the liver, but the intracellular pathway of apoE following particle internalization is poorly defined. In the present study, we investigated whether retroendocytosis is a unique feature of apoE as compared with apoB by studying the intracellular fate of very low density lipoprotein-sized apoE-containing triglyceride-rich emulsion particles and LDL after LDLr-mediated uptake. Incubation of HepG2 cells with [(3)H]cholesteryl oleate-labeled particles at 37 degrees C led to a rapid release of [(3)H]cholesterol within 30 min for both LDL and emulsion particles. In contrast, emulsion-derived (125)I-apoE was more resistant to degradation (>/=120 min) than LDL-derived (125)I-apoB (30 min). Incubation at 18 degrees C, which allows endosomal uptake but prevents lysosomal degradation, with subsequent incubation at 37 degrees C resulted in a time-dependent release of intact apoE from the cells (up to 14% of the endocytosed apoE at 4 h). The release of apoE was accelerated by the presence of protein-free emulsion (20%) or high density lipoprotein (26%). Retroendocytosis of intact particles could be excluded since little intact [(3)H]cholesteryl oleate was released (<3%). In contrast, the degradation of LDL was complete with virtually no secretion of intact apoB into the medium. The intracellular stability of apoE was also demonstrated after hepatic uptake in C57Bl/6 mice. Intravenous injection of (125)I-apoE and [(3)H]cholesteryl oleate-labeled emulsions resulted in efficient LDLr-mediated uptake of both components by the liver (45-50% of the injected dose after 20 min). At 1 h after injection, only 15-20% of the hepatic (125)I-apoE was degraded, whereas 75% of the [(3)H]cholesteryl oleate was hydrolyzed. From these data we conclude that following LDLr-mediated internalization by liver cells, apoE can escape degradation and can be resecreted. This sequence of events may allow apoE to participate in its hypothesized intracellular functions such as mediator of the post-lysosomal trafficking of lipids and very low density lipoprotein assembly. PMID- 10722696 TI - Characterization of the selectivity filter of the epithelial sodium channel. AB - The epithelial sodium channel (ENaC) is composed of three homologous subunits termed alpha, beta, and gamma. Previous studies suggest that selected residues within a hydrophobic region immediately preceding the second membrane-spanning domain of each subunit contribute to the conducting pore of ENaC. We probed the pore of mouse ENaC by systematically mutating all 24 amino acids within this putative pore region of the alpha-subunit to cysteine and co-expressing these mutants with wild type beta- and gamma-subunits of mouse ENaC in Xenopus laevis oocytes. Functional characteristics of these mutants were examined by two electrode voltage clamp and single channel recording techniques. Two distinct domains were identified based on the functional changes associated with point mutations. An amino-terminal domain (alpha-Val(569)-alpha-Gly(579)) showed minimal changes in cation selectivity or amiloride sensitivity following cysteine substitution. In contrast, cysteine substitutions within the carboxyl-terminal domain (alpha-Ser(580)-alpha-Ser(592)) resulted in significant changes in cation selectivity and moderately altered amiloride sensitivity. The mutant channels containing alphaG587C or alphaS589C were permeable to K(+), and mutation of a GSS tract (positions alpha587-alpha589) to GYG resulted in a moderately K(+) selective channel. Our results suggest that the C-terminal portion of the pore region within the alpha-subunit contributes to the selectivity filter of ENaC. PMID- 10722697 TI - Unusual oxidative chemistry of N(omega)-hydroxyarginine and N-hydroxyguanidine catalyzed at an engineered cavity in a heme peroxidase. AB - Heme enzymes are capable of catalyzing a range of oxidative chemistry with high specificity, depending on the surrounding protein environment. We describe here a reaction catalyzed by a mutant of cytochrome c peroxidase, which is similar but distinct from those catalyzed by nitric-oxide synthase. In the R48A mutant, an expanded water-filled cavity was created above the distal heme face. N hydroxyguanidine (NHG) but not guanidine was shown to bind in the cavity with K(d) = 8.5 mM, and coordinate to the heme to give a low spin state. Reaction of R48A with peroxide produced a Fe(IV)=O/Trp(.+) center capable of oxidizing either NHG or N(omega)-hydroxyarginine (NHA), but not arginine or guanidine, by a multi turnover catalytic process. Oxidation of either NHG or NHA by R48A did not result in the accumulation of NO, NO(2)(-), NO(3)(-), urea, or citrulline, but instead afforded a yellow product with absorption maxima of 257 and 400 nm. Mass spectrometry of the derivatized NHA products identified the yellow species as N nitrosoarginine. We suggest that a nitrosylating agent, possibly derived from HNO, is produced by the oxidation of one molecule of substrate. This then reacts with a second substrate molecule to form the observed N-nitroso products. This complex chemistry illustrates how the active sites of enzymes such as nitric oxide synthase may serve to prevent alternative reactions from occurring, in addition to enabling those desired. PMID- 10722698 TI - Mechanism for proton conduction of the M(2) ion channel of influenza A virus. AB - The M(2) integral membrane protein of influenza A virus forms a proton-selective ion channel. We investigated the mechanism for proton transport of the M(2) protein in Xenopus oocytes using a two-electrode voltage clamp and in CV-1 cells using the whole cell patch clamp technique. Membrane currents were recorded while manipulating the external solution to alter either the total or free proton concentration or the solvent itself. Membrane conductance decreased by approximately 50% when D(2)O replaced H(2)O as the solvent. From this, we conclude that hydrogen ions do not pass through M(2) as hydronium ions, but instead must interact with titratable groups that line the pore of the channel. M(2) currents measured in solutions of low buffer concentration (<15 mM in oocytes and <0.15 mM in CV-1 cells) were smaller than those studied in solutions of high buffer concentration. Furthermore, the reversal voltage measured in low buffer was shifted to a more negative voltage than in high buffer. Also, at a given pH, M(2) current amplitude in 15 mM buffer decreased when pH-pK(a) was increased by changing the buffer pK(a). Collectively, these results demonstrate that M(2) currents can be limited by external buffer capacity. The data presented in this study were also used to estimate the maximum single channel current of the M(2) ion channel, which was calculated to be on the order of 1-10 fA. PMID- 10722699 TI - Oxidative stress disrupts glucocorticoid hormone-dependent transcription of the amiloride-sensitive epithelial sodium channel alpha-subunit in lung epithelial cells through ERK-dependent and thioredoxin-sensitive pathways. AB - The amiloride-sensitive epithelial Na(+) channel (ENaC) plays a critical role in the maintenance of alveolar fluid balance. It is generally accepted that reactive oxygen and nitrogen species can inhibit ENaC activity and aggravate acute lung injury; however, the molecular mechanism for free radical-mediated ENaC inhibition is unclear. Previously, we showed that the expression of the alpha subunit of ENaC, alpha-ENaC, which is indispensable for ENaC activity, is repressed by Ras activation in salivary epithelial cells. Here, we investigated whether exogenous H(2)O(2) modulates alpha-ENaC gene expression in lung epithelial cells through a similar molecular mechanism. Utilizing transient transfection reporter assays and site-directed mutagenesis analyses, we found that the glucocorticoid response element (GRE), located at -1334 to -1306 base pairs of the alpha-ENaC 5'-flanking region, is the major enhancer for the stimulated alpha-ENaC expression in A549 lung epithelial cells. We further demonstrate that the presence of an intact GRE is necessary and sufficient for oxidants to repress alpha-ENaC expression. Consistent with our hypothesis, exogenous H(2)O(2)-mediated repression of alpha-ENaC GRE activity is partially blocked by either a specific inhibitor for extracellular signal-regulated kinase (ERK) pathway activation, U0126, or dominant negative ERK, suggesting that, in part, activated ERK may mediate the repressive effects of H(2)O(2) on alpha-ENaC expression. In addition, overexpression of thioredoxin restored glucocorticoid receptor action on the alpha-ENaC GRE in the presence of exogenous H(2)O(2). Taken together, we hypothesize that oxidative stress impairs Na(+) transport activity by inhibiting dexamethasone-dependent alpha-ENaC GRE activation via both ERK-dependent and thioredoxin-sensitive pathways. These results suggest a putative mechanism whereby cellular redox potentials modulate the glucocorticoid receptor/dexamethasone effect on alpha-ENaC expression in lung and other tight epithelia. PMID- 10722700 TI - Tumor necrosis factor-alpha and Fas activate complementary Fas-associated death domain-dependent pathways that enhance apoptosis induced by gamma-irradiation. AB - Activation of either tumor necrosis factor receptor 1 or Fas induces a low level of programmed cell death in LNCaP human prostate cancer cells. We have shown that LNCaP cells are entirely resistant to gamma-radiation-induced apoptosis, but can be sensitized to irradiation by TNF-alpha. Fas activation also sensitized LNCaP cells to irradiation, causing nearly 40% cell death 72 h after irradiation. Caspase-8 was cleaved and activated after exposure to tumor necrosis factor (TNF) alpha. However, after exposure to anti-Fas antibody caspase-8 cleavage occurred only between the 26-kDa N-terminal prodomain and the 28-kDa C-terminal region that contains the protease components. Although anti-Fas antibody plus irradiation induced apoptosis that could be blocked by the pancaspase inhibitor zVAD, there was no measurable caspase-8 activity after exposure to anti-Fas antibody. The effector caspases-6 and -7, and to a lesser extent caspase-3, were activated by TNF-alpha, but not by anti-Fas antibody. Anti-Fas antibody, like TNF alpha also activated serine proteases that contributed to cell death. Exposure of LNCaP cells simultaneously to TNF-alpha and anti-Fas antibody CH-11 resulted in marked enhancement of apoptosis that occurred very rapidly and was still further augmented by irradiation. Rapid apoptosis that ensued from combined treatment with TNF-alpha, anti-Fas antibody, and irradiation was completely blocked either by zVAD or expression of dominant negative Fas-associated death domain. Our data shows that there are qualitative differences in caspase activation resulting from either TNF receptor 1 or Fas. Simultaneous activation of these receptors was synergistic and caused rapid epithelial cell apoptosis mediated by the caspase cascade. PMID- 10722701 TI - Active site serine promotes stabilization of the reactive glutathione thiolate in rat glutathione transferase T2-2. Evidence against proposed sulfatase activity of the corresponding human enzyme. AB - Ser(11) in rat glutathione transferase T2-2 is important for stabilization of the reactive enzyme-bound glutathione thiolate in the reaction with 1-menaphthyl sulfate. The S11A mutation increased the pK(a) value for the pH dependence of the rate constant for pre-steady-state product formation, from 5.7 to 7.9. This pH dependence is proposed to reflect titration of enzyme-bound glutathione thiol. Further, the mutation lowered the k(cat) value but not because of the impaired stabilization of the glutathione thiolate. In fact, several steps on the reaction pathway were affected by the S11A mutation, and the cause of the decreased k(cat) for the mutant was found to be a slower product release. The data presented here contradict the hypothesis that glutathione transferase T2-2 could act as a sulfatase that is not dependent on Ser(11) for the catalytic activity, as proposed for the corresponding human enzyme (Tan, K.-L., Chelvanayagam, G., Parker, M. W., and Board, P. G. (1996) Biochem. J. 319, 315-321; Rossjohn, J., McKinstry, W. J., Oakley, A. J., Verger, D., Flanagan, J., Chelvanayagam, G., Tan, K.-L., Board, P. G., and Parker, M. W. (1998) Structure 6, 309-322). On the contrary, Ser(11) governs both chemical and physical steps of the catalyzed reaction. PMID- 10722702 TI - New aspects of siglec binding specificities, including the significance of fucosylation and of the sialyl-Tn epitope. Sialic acid-binding immunoglobulin superfamily lectins. AB - The siglecs (sialic acid-binding immunoglobulin superfamily lectins) are immunoglobulin superfamily members recognizing sialylated ligands. Most prior studies of siglec specificities focused on alpha2-3- and alpha2-6 sialyllactos(amin)es and on one or two of the siglecs at a time. Here, we explore several new aspects of specificities of the first six reported siglecs, using sialylated glycans presented in multivalent form, on synthetic polyacrylamide backbones, or on mucin polypeptides. First, we report that binding of siglec-1 (sialoadhesin), siglec-3 (CD33), siglec-4a (myelin-associated glycoprotein), and siglec-5 to alpha2-3 sialyllactosamine is affected markedly by the presence of an alpha1-3-linked fucose. Thus, while siglecs may not interfere with selectin mediated recognition, fucosylation could negatively regulate siglec binding. Second, in contrast to earlier studies, we find that siglec-3 prefers alpha2-6 sialyllactose. Third, siglec-5 binds alpha2-8-linked sialic acid, making it the siglec least specific for linkage recognition. Fourth, siglecs-2 (CD22), -3, -5, and -6 (obesity-binding protein 1) showed significant binding to sialyl-Tn (Neu5Acalpha2-6-GalNAc), a tumor marker associated with poor prognosis. Fifth, siglec-6 is an exception among siglecs in not requiring the glycerol side chain of sialic acid for recognition. Sixth, all siglecs require the carboxyl group of sialic acid for binding. Finally, the presentation of the sialyl-Tn epitope and/or more extended structures that include this motif may be important for optimal recognition by the siglecs. This was concluded from studies using ovine, bovine, and porcine submaxillary mucins and Chinese hamster ovary cells transfected with ST6GalNAc-I and/or the mucin polypeptide MUC1. PMID- 10722703 TI - Loss of N-glycolylneuraminic acid in human evolution. Implications for sialic acid recognition by siglecs. AB - The common sialic acids of mammalian cells are N-acetylneuraminic acid (Neu5Ac) and N-glycolylneuraminic acid (Neu5Gc). Humans are an exception, because of a mutation in CMP-sialic acid hydroxylase, which occurred after our common ancestor with great apes. We asked if the resulting loss of Neu5Gc and increase in Neu5Ac in humans alters the biology of the siglecs, which are Ig superfamily members that recognize sialic acids. Human siglec-1 (sialoadhesin) strongly prefers Neu5Ac over Neu5Gc. Thus, humans have a higher density of siglec-1 ligands than great apes. Siglec-1-positive macrophages in humans are found primarily in the perifollicular zone, whereas in chimpanzees they also occur in the marginal zone and surrounding the periarteriolar lymphocyte sheaths. Although only a subset of chimpanzee macrophages express siglec-1, most human macrophages are positive. A known evolutionary difference is the strong preference of mouse siglec-2 (CD22) for Neu5Gc, contrasting with human siglec-2, which binds Neu5Ac equally well. To ask when the preference for Neu5Gc was adjusted in the human lineage, we cloned the first three extracellular domains of siglec-2 from all of the great apes and examined their preference. In fact, siglec-2 had evolved a higher degree of recognition flexibility before Neu5Gc was lost in humans. Human siglec-3 (CD33) and siglec-6 (obesity-binding protein 1) also recognize both Neu5Ac and Neu5Gc, and siglec-5 may have some preference for Neu5Gc. Others showed that siglec-4a (myelin-associated glycoprotein) prefers Neu5Ac over Neu5Gc. Thus, the human loss of Neu5Gc may alter biological processes involving siglec-1, and possibly, siglec 4a or -5. PMID- 10722704 TI - A natural ErbB4 isoform that does not activate phosphoinositide 3-kinase mediates proliferation but not survival or chemotaxis. AB - ErbB4 is a member of the epidermal growth factor receptor (ErbB) family that mediates cellular responses activated by neuregulins (NRG) and other epidermal growth factor-like growth factors. Two naturally occurring ErbB4 isoforms, ErbB4 CYT-1 and ErbB4 CYT-2, have previously been identified. Unlike ErbB4 CYT-1, ErbB4 CYT-2 lacks a phosphoinositide 3-kinase (PI3-K)-binding site and is incapable of activating PI3-K. We have now examined the consequences of the inability of this isoform to activate PI3-K on cell proliferation, survival, and chemotaxis in response to NRG-1beta: (i) NRG-1beta stimulated proliferation of cells expressing either ErbB4 CYT-1 or ErbB4 CYT-2. Consistent with the mitogenic responsiveness, analysis of downstream signaling showed that Shc and MAPK were phosphorylated after stimulating either isoform with NRG-1beta. (ii) NRG-1beta protected cells expressing ErbB4 CYT-1 but not cells expressing ErbB4 CYT-2 from starvation induced apoptosis as measured by effects on cell number and 4', 6-diamidino-2 phenylindole staining. Furthermore, in cells expressing ErbB4 CYT-2, Akt, a protein kinase that mediates cell survival, was not phosphorylated. (iii) NRG 1beta stimulated chemotaxis and membrane ruffling in cells expressing ErbB4 CYT-1 but not in cells expressing ErbB4 CYT-2. In summary, ErbB4 CYT-2 can mediate proliferation but not chemotaxis or survival. These results suggest a novel mechanism by which cellular responses such as chemotaxis and survival may be regulated by the expression of alternative receptor-tyrosine kinase isoforms that differ in their coupling to PI3-K signaling. PMID- 10722705 TI - CD95(Fas/APO-1) signals ceramide generation independent of the effector stage of apoptosis. AB - Although numerous studies document caspase-independent ceramide generation preceding apoptosis upon environmental stress, the molecular ordering of ceramide generation during cytokine-induced apoptosis remains uncertain. Here, we show that CD95-induced ceramide elevation occurs during the initiation phase of apoptosis. We titrated down the amount of FADD transfected into HeLa and 293T cells until it was insufficient for apoptosis, although cycloheximide (CHX) still triggered the effector phase. Even in the absence of CHX, ceramide levels increased rapidly, peaking at 2.7 +/- 0.2-fold of control 8 h post-transfection. Dominant negative FADD failed to confer ceramide generation or CHX-mediated apoptosis. Ceramide generation induced by FADD was initiator caspase-dependent, being blocked by crmA. Limited pro-caspase 8 overexpression also increased ceramide levels 2.7 +/- 0.2-fold, yet failed, without CHX, to initiate apoptosis. Expression of membrane-targeted oligomerized CD-8 caspase 8 induced apoptosis without CHX, yet elevated ceramide only to a level equivalent to limited pro caspase 8 transfection. Ceramide elevations were detected concurrently by diacylglycerol kinase and electrospray tandem mass spectrometry. These investigations provide evidence that ceramide generation is initiator caspase dependent and occurs prior to commitment to the effector phase of apoptosis, definitively ordering ceramide as proximal in CD95 signaling. PMID- 10722706 TI - Role of acidic sphingomyelinase in Fas/CD95-mediated cell death. AB - Engagement of the Fas receptor has been reported to induce ceramide generation via activation of acidic sphingomyelinase (aSMase). However, the role of aSMase in Fas-mediated cell death is controversial. Using genetically engineered mice deficient in the aSMase gene (aSMase(-/-)), we found that thymocytes, concanavalin A-activated T cells, and lipopolysaccharide-activated B cells derived from both aSMase(-/-) and aSMase(+/+) mice were equally sensitive to Fas mediated cell death, triggered by either anti-Fas antibody or Fas ligand in vitro. Similarly, activation-induced apoptosis of T lymphocytes was unaffected by the status of aSMase, and aSMase(-/-) mice failed to show immunological symptoms seen in animals with defects in Fas function. In vivo, intravenous injection of 3 microg/25 g mouse body weight of anti-Fas Jo2 antibody into aSMase(-/-) mice failed to affect hepatocyte apoptosis or mortality, whereas massive hepatocyte apoptosis and animal death occurred in wild type littermates. Animals heterozygous for aSMase deficiency were also significantly protected. Susceptibility of aSMase(-/-) mice to anti-Fas antibody was demonstrated with higher antibody doses (>/=4 microg/25 g mouse). These data indicate a role for aSMase in Fas-mediated cell death in some but not all tissues. PMID- 10722707 TI - Subtype-specific trafficking of endothelin receptors. AB - We investigated the subcellular localization of two endothelin receptors (ET(A)R and ET(B)R). To visualize these receptors directly, the C terminus of each receptor was fused to the N terminus of enhanced green fluorescent protein (designated as ETR-EGFP). When transiently expressed in various mammalian cell lines, ET(A)R-EGFP was predominantly localized on the plasma membrane. By contrast, ET(B)R-EGFP was, independent of ligand stimulation, predominantly localized on the intracellular vesicular structures containing Lamp-1. Immunoblot analyses revealed that at steady state ET(B)R-EGFP was highly degraded, and its degradation was inhibited by bafilomycin A(1). Antibody uptake experiments suggested that the ET(B)R-EGFP molecules were internalized from the plasma membrane. It is therefore likely that ET(B)R is first transported to the plasma membrane and then internalized, irrespective of ligand stimulation, to lysosomes where it undergoes proteolytic degradation. Exchanging the C-terminal cytoplasmic tails of the two ETRs revealed that the cytoplasmic tail is responsible for both the intracellular localization and the degradation of the receptors. Deletion of the extreme C-terminal 35 amino acids from both receptors allowed the receptor proteins to localize predominantly in the intracellular vesicles and to degrade. These observations indicate that the cytoplasmic tail of ET(A)R determines its plasma membrane localization. Stimulation with endothelin-1 increased the amount of intact ETR-EGFP fusion proteins without increasing their de novo synthesis, suggesting that binding of endothelin-1 stabilizes the ETRs. PMID- 10722708 TI - Intracellular localization of processing events in human surfactant protein B biosynthesis. AB - Surfactant protein B (SP-B) is essential to the function of pulmonary surfactant and to alveolar type 2 cell phenotype. Human SP-B is the 79-amino acid product of extensive post-translational processing of a 381-amino acid preproprotein. Processing involves modification of the primary translation product from 39 to 42 kDa and at least 3 subsequent proteolytic cleavages to produce the mature 8-kDa SP-B. To examine the intracellular sites of SP-B processing, we carried out immunofluorescence cytochemistry and inhibitor studies on human fetal lung in explant culture and isolated type 2 cells in monolayer culture using polyclonal antibodies to human SP-B(8) (Phe(201)-Met(279)) and specific epitopes within the N- (NFProx, Ser(145)-Leu(160); NFlank Gln(186)-Gln(200)) and C-terminal (CFlank, Gly(284)-Ser(304)) propeptides of pro-SP-B. Fluorescence immunocytochemistry using epitope-specific antisera showed colocalization of pro-SP-B with the endoplasmic reticulum resident protein BiP. The 25-kDa intermediate was partially endo H-sensitive, colocalized with the medial Golgi resident protein MG160, and shifted into the endoplasmic reticulum in the presence of brefeldin A, which interferes with anterograde transport from endoplasmic reticulum to Golgi. The 9 kDa intermediate colocalized in part with MG160 but not with Lamp-1, a transmembrane protein resident in late endosomes and lamellar bodies. Brefeldin A induced a loss of colocalization between MG160 and NFlank, shifting NFlank immunostaining to a juxtanuclear tubular array. In pulse-chase studies, brefeldin A blocked all processing of 42-kDa pro-SP-B whereas similar studies using monensin blocked the final N-terminal processing event of 9 to 8 kDa SP-B. We conclude that: 1) the first enzymatic cleavage of pro-SP-B to the 25-kDa intermediate is in the brefeldin A-sensitive, medial Golgi; 2) cleavage of the 25 kDa intermediate to a 9-kDa form is a trans-Golgi event that is slowed but not blocked by monensin; 3) the final cleavage of 9 to 8 kDa SP-B is a monensin sensitive, post-Golgi event occurring prior to transfer of SP-B to lamellar bodies. PMID- 10722709 TI - Nuclear magnetic resonance solution conformation of alpha-conotoxin AuIB, an alpha(3)beta(4) subtype-selective neuronal nicotinic acetylcholine receptor antagonist. AB - The neuronal nicotinic acetylcholine receptors constitute a highly diverse group, with subtypes consisting of pentameric combinations of alpha and beta subunits. alpha-Conotoxins are a homologous series of small peptides that antagonize these receptors. We present the three-dimensional solution structure of alpha-conotoxin AuIB, the first 15-residue alpha-conotoxin known to selectively block the alpha(3)beta(4) nicotinic acetylcholine receptor subtype. The pairwise backbone and heavy-atom root mean square deviation for an ensemble of 20 structures are 0.269 and 0.720 A, respectively. The overall fold of alpha-conotoxin AuIB closely resembles that of the alpha4/7 subfamily alpha-conotoxins. However, the absence of Tyr(15), normally present in other alpha4/7 members, results in tight bending of the backbone at the C terminus and effectively renders Asp(14) to assume the spatial location of Tyr(15) present in other neuronal alpha4/7 alpha-conotoxins. Structural comparison of alpha-conotoxin AuIB with the alpha(3)beta(2) subtype specific alpha-conotoxin MII shows different electrostatic surface charge distributions, which may be important in differential receptor subtype recognition. PMID- 10722710 TI - S100A13. Biochemical characterization and subcellular localization in different cell lines. AB - S100 proteins became of major interest because of their divergent cell- and tissue-specific expression, their close association with a number of human diseases, and their importance for clinical diagnostics. Here, we report for the first time the purification and characterization of human recombinant S100A13. Flow dialysis revealed that the homodimeric S100A13 binds four Ca(2+) in two sets of binding sites, both displaying positive cooperativity but of very different affinity. Fluorescence and difference spectrophotometry indicate that the Trp/Tyr signal changes are almost complete upon binding of Ca(2+) to the two high affinity sites, which probably correspond to the C-terminal EF-hands in each subunit. The far-UV circular dichroic signal also changes upon binding of the first two Ca(2+). So far, the tissue distribution of S100A13 has not been well characterized. Here, we show that S100A13 is widely expressed in various types of tissues with a high expression level in thyroid gland. Using specific antisera against S100A13, high protein expression was detected in follicle cells of thyroid, Leydig cells of testis, and specific cells of brain. In human smooth muscle cells, which co-express S100A2 in the nucleus and S100A1 in stress fibers, S100A13 shows a unique subcellular localization in the perinuclear area. These data suggest diverse functions for this protein in signal transduction. PMID- 10722711 TI - Cbfa1 is a positive regulatory factor in chondrocyte maturation. AB - Cbfa1 is a transcription factor that belongs to the runt domain gene family. Cbfa1-deficient mice showed a complete lack of bone formation due to the maturational arrest of osteoblasts, demonstrating that Cbfa1 is an essential factor for osteoblast differentiation. Further, chondrocyte maturation was severely disturbed in Cbfa1-deficient mice. In this study, we examined the possibility that Cbfa1 is also involved in the regulation of chondrocyte differentiation. mRNAs for both Cbfa1 isotypes, type I Cbfa1 (Pebp2alphaA/Cbfa1) and type II Cbfa1 (Osf2/Cbfa1 or til-1), which are different in N-terminal domain, were expressed in terminal hypertrophic chondrocytes as well as osteoblasts. In addition, mRNA for type I Cbfa1 was expressed in other hypertrophic chondrocytes and prehypertrophic chondropcytes. In a chondrogenic cell line, ATDC5, the expression of type I Cbfa1 was elevated prior to differentiation to the hypertrophic phenotype, which is characterized by type X collagen expression. Treatment with antisense oligonucleotides for type I Cbfa1 severely reduced type X collagen expression in ATDC5 cells. Retrovirally forced expression of either type I or type II Cbfa1 in chick immature chondrocytes induced type X collagen and MMP13 expression, alkaline phosphatase activity, and extensive cartilage-matrix mineralization. These results indicate that Cbfa1 is an important regulatory factor in chondrocyte maturation. PMID- 10722712 TI - Sporadic inclusion body myositis correlates with increased expression and cross linking by transglutaminases 1 and 2. AB - Sporadic inclusion body myositis (SIBM) is characterized by vacuolar degeneration of muscle fibers and intrafiber clusters of paired helical filaments with abnormal amyloid deposition. Because of their potential involvement in other degenerative disorders, we have examined the expression of transglutaminases (TGases) in normal and SIBM tissues. We report that at least two different enzymes, the ubiquitous TGase 2 as well as the TGase 1 enzyme, are present in muscle tissues. However, in comparison with normal tissue, the expression of TGases 1 and 2 was increased 2.5- and 4-fold in SIBM, accompanied by about a 20 fold higher total TGase activity. By immunohistochemical staining, in normal muscle, TGase 2 expression was restricted to some endomysial connective tissue elements, whereas TGase 1 and beta-amyloid proteins were not detectable. In SIBM muscle, both TGases 1 and 2 as well as amyloid proteins were brightly expressed and co-localized in the vacuolated muscle fibers, but none of these proteins colocalized with inflammatory cell markers. Next, we isolated high molecular weight insoluble proteins from SIBM muscle tissue and showed that they were cross linked by about 6 residues/1000 residues of the isopeptide bond. Furthermore, by amino acid sequencing of solubilized tryptic peptides, they contain amyloid and skeletal muscle proteins. Together, these findings suggest that elevated expression of TGases 1 and 2 participate in the formation of insoluble amyloid deposits in SIBM tissue and in this way may contribute to progressive and debilitating muscle disease. PMID- 10722713 TI - Ribozyme ablation demonstrates that the cardiac subtype of the voltage-sensitive calcium channel is the molecular transducer of 1, 25-dihydroxyvitamin D(3) stimulated calcium influx in osteoblastic cells. AB - 1,25-Dihydroxyvitamin D(3) (1,25(OH)(2)D(3)) stimulates transmembrane influx of Ca(2+) through L-type voltage-sensitive Ca(2+) channels (VSCCs) in ROS 17/2.8 osteoblastic cells. Ca(2+) influx modulates osteoblastic activities including matrix deposition, hormone responsiveness, and Ca(2+)-dependent signaling. 1, 25(OH)(2)D(3) also regulates transcript levels encoding VSCCs. L-type VSCCs are multisubunit complexes composed of a central pore-forming alpha(1) subunit and four additional subunits. The alpha(1) subunit is encoded by one gene in a multimember family, defining tissue-specific subtypes. Osteoblasts synthesize two splice variants of the alpha(1C) cardiac VSCC subtype; however, the molecular identity of the 1,25(OH)(2)D(3)-regulated VSCC remained unknown. We created a ribozyme specifically cleaving alpha(1C) mRNA. To increase target ablation efficiency, the ribozyme was inserted into U1 small nuclear RNA (snRNA) by engineering the U1 snRNA expression cassette, conferring the ribozyme transcript with stabilizing stem-loops at both sides and the Sm binding site that facilitates localization into nucleoplasm. After transfection of ROS 17/2.8 cells with U1 ribozyme-encoding vector, stable clonal cells were selected in which the expression of alpha(1C) transcript and protein were strikingly reduced. Ca(2+) influx assays in ribozyme transfectants showed selective attenuation of depolarization and 1, 25(OH)(2)D(3)-regulated Ca(2+) responses. We conclude that the cardiac subtype of the L-type VSCC is the transducer of stimulated Ca(2+) influx in ROS 17/2.8 osteoblastic cells. PMID- 10722714 TI - A protein encoded within the Down syndrome critical region is enriched in striated muscles and inhibits calcineurin signaling. AB - Here we describe a small family of proteins, termed MCIP1 and MCIP2 (for myocyte enriched calcineurin interacting protein), that are expressed most abundantly in striated muscles and that form a physical complex with calcineurin A. MCIP1 is encoded by DSCR1, a gene located in the Down syndrome critical region. Expression of the MCIP family of proteins is up-regulated during muscle differentiation, and their forced overexpression inhibits calcineurin signaling to a muscle-specific target gene in a myocyte cell background. Binding of MCIP1 to calcineurin A requires sequence motifs that resemble calcineurin interacting domains found in NFAT proteins. The inhibitory action of MCIP1 involves a direct association with the catalytic domain of calcineurin, rather than interference with the function of downstream components of the calcineurin signaling pathway. The interaction between MCIP proteins and calcineurin may modulate calcineurin-dependent pathways that control hypertrophic growth and selective programs of gene expression in striated muscles. PMID- 10722716 TI - The effect of a reducing-end extension on pentasaccharide binding by antithrombin. AB - Antithrombin requires heparin for efficient inhibition of the final two proteinases of the blood coagulation cascade, factor Xa and thrombin. Antithrombin binds heparin via a specific pentasaccharide domain in a two-step mechanism whereby initial weak binding is followed by a conformational change and subsequent tight binding. The goal of this study is to investigate the role of a reducing-end extension in the binding of the longer oligosaccharides that contain the cognate pentasaccharide sequence. We determined the antithrombin binding properties of a synthetic heptasaccharide containing the natural pentasaccharide sequence (DEFGH) and an additional reducing-end disaccharide (DEFGHG'H'). Binding at low ionic strength is unaffected by the disaccharide addition, but at ionic strengths >/=0.2 the mode of heptasaccharide binding changes resulting in a 2 fold increase in affinity due to a decrease in the off-rate caused by a greater nonionic contribution to binding. Molecular modeling of possible binding modes for the heptasaccharide at high ionic strength indicates a possible shift in position of the pentasaccharide domain to occupy the extended heparin-binding site. This conclusion supports the likely presence of a range of sequences that can bind to and activate antithrombin in the natural heparan sulfates that line the vascular endothelium. PMID- 10722715 TI - The CafA protein required for the 5'-maturation of 16 S rRNA is a 5'-end dependent ribonuclease that has context-dependent broad sequence specificity. AB - The CafA protein, which was initially described as having a role in either Escherichia coli cell division or chromosomal segregation, has recently been shown to be required for the maturation of the 5'-end of 16 S rRNA. The sequence of CafA is similar to that of the N-terminal ribonucleolytic half of RNase E, an essential E. coli enzyme that has a central role in the processing of rRNA and the decay of mRNA and RNAI, the antisense regulator of ColE1-type plasmids. We show here that a highly purified preparation of CafA is sufficient in vitro for RNA cutting. We detected CafA cleavage of RNAI and a structured region from the 5'-untranslated region of ompA mRNA within segments cleavable by RNaseE, but not CafA cleavage of 9 S RNA at its "a" RNase E site. The latter is consistent with the finding that the generation of 5 S rRNA from its 9 S precursor can be blocked by inactivation of RNase E in cells that are wild type for CafA. Interestingly, however, a decanucleotide corresponding in sequence to the a site of 9 S RNA was cut efficiently indicating that cleavage by CafA is regulated by the context of sites within structured RNAs. Consistent with this notion is our finding that although 23 S rRNA is stable in vivo, a segment from this RNA is cut efficient by CafA at multiple sites in vitro. We also show that, like RNase E cleavage, the efficiency of cleavage by CafA is dependent on the presence of a monophosphate group on the 5'-end of the RNA. This finding raises the possibility that the context dependence of cleavage by CafA may be due at least in part to the separation of a cleavable sequence from the 5'-end of an RNA. Comparison of the sites surrounding points of CafA cleavage suggests that this enzyme has broad sequence specificity. Together with the knowledge that CafA can cut RNAI and ompA mRNA in vitro within segments whose cleavage in vivo initiates the decay of these RNAs, this finding suggests that CafA may contribute at some point during the decay of many RNAs in E. coli. PMID- 10722717 TI - Zinc finger proteins as designer transcription factors. AB - Recent progress in the design and selection of novel zinc finger proteins with desired DNA binding specificities now allows construction of tailor-made DNA binding proteins that specifically recognize almost any predetermined DNA sequence. Such novel or "designer" zinc finger proteins with desired DNA binding specificities can serve as efficient transcription factors in various mammalian cell lines. In addition, they may be broadly useful in the regulation of endogenous genes in transgenic organisms and eventually in gene therapy applications. In this report, we use a series of transient and stable transfection experiments to demonstrate that the expression of a target gene can be controlled by changing the in vivo concentration of designer zinc finger proteins in a dose-dependent manner. We also report that designer zinc finger proteins can access their binding sites integrated into the genome and function as potent transcription factors. Our results suggest that designer zinc finger transcription factors that specifically recognize appropriate sites in the promoter of a target gene may have broad applications in the post-genomic era. PMID- 10722718 TI - The pro-alpha3(V) collagen chain. Complete primary structure, expression domains in adult and developing tissues, and comparison to the structures and expression domains of the other types V and XI procollagen chains. AB - The low abundance fibrillar collagen type V is widely distributed in tissues as an alpha1(V)(2)alpha2(V) heterotrimer that helps regulate the diameters of fibrils of the abundant collagen type I. Mutations in the alpha1(V) and alpha2(V) chain genes have been identified in some cases of classical Ehlers-Danlos syndrome (EDS), in which aberrant collagen fibrils are associated with connective tissue fragility, particularly in skin and joints. Type V collagen also exists as an alpha1(V)alpha2(V)alpha3(V) heterotrimer that has remained poorly characterized chiefly due to inability to obtain the complete primary structure or nucleic acid probes for the alpha3(V) chain or its biosynthetic precursor, pro alpha3(V). Here we provide human and mouse full-length pro-alpha3(V) sequences. Pro-alpha3(V) is shown to be closely related to the alpha1(V) precursor, pro alpha1(V), but with marked differences in N-propeptide sequences, and collagenous domain features that provide insights into the low melting temperature of alpha1(V)alpha2(V)alpha3(V) heterotrimers, lack of heparin binding by alpha3(V) chains and the possibility that alpha1(V)alpha2(V)alpha3(V) heterotrimers are incorporated into heterotypic fibrils. In situ hybridization of mouse embryos detects alpha3(V) expression primarily in the epimysial sheaths of developing muscles and within nascent ligaments adjacent to forming bones and in joints. This distribution, and the association of alpha1(V), alpha2(V), and alpha3(V) chains in heterotrimers, suggests the human alpha3(V) gene COL5A3 as a candidate locus for at least some cases of classical EDS in which the alpha1(V) and alpha2(V) genes have been excluded, and for at least some cases of the hypermobility type of EDS, a condition marked by gross joint laxity and chronic musculoskeletal pain. COL5A3 is mapped to 19p13.2 near a polymorphic marker that should be useful in analyzing linkage with EDS and other disease phenotypes. PMID- 10722719 TI - Three subunit a isoforms of mouse vacuolar H(+)-ATPase. Preferential expression of the a3 isoform during osteoclast differentiation. AB - Vacuolar H(+)-ATPase (V-ATPase) is a multi-subunit enzyme with a membrane peripheral catalytic (V(1)) and an intrinsic (V(o)) sector. We have identified three cDNA clones coding for isoforms of mouse V(o) subunit a (a1, a2, and a3). They exhibit 48-52% identity with each other and high similarity to subunit a of other species. The a1 isoform was mainly expressed in brain and liver. The a2 isoform was observed in heart and kidney in addition to brain and liver. Transcripts for the a3 isoform were strongly expressed in heart and liver. The a3 isoform was induced during osteoclast differentiation, and localized in the plasma membrane and cytoplasmic filamentous structures. In contrast to a3, the a1 isoform was constitutively expressed and localized in the cytoplasmic endomembrane compartments of the same cells. These findings suggest that the a3 isoform is a component of the plasma membrane V-ATPase essential for bone resorption. PMID- 10722720 TI - Activation of MST/Krs and c-Jun N-terminal kinases by different signaling pathways during cytotrienin A-induced apoptosis. AB - We found that antitumor drugs such as cytotrienin A, camptothecin, taxol, and 5 fluorouracil induced the activation of a 36-kDa protein kinase (p36 myelin basic protein (MBP) kinase) during apoptosis in human promyelocytic leukemia HL-60 cells. This p36 MBP kinase, which phosphorylates MBP in an in-gel kinase assay, results from the caspase-3-mediated proteolytic cleavage of MST/Krs protein, a mammalian Ste20-like serine/threonine kinase. Herein the correlation between cytotrienin A-induced apoptosis and the activation of MST/Krs proteins was examined in human tumor cell lines, including leukemia-, lung-, epidermoid-, cervix-, stomach-, and brain-derived cell lines. In cytotrienin A-sensitive cell lines, we observed a strong activation of p36 MBP kinase by cleavage of the C terminal regulatory domain of full-length MST/Krs proteins by caspase-3. When the kinase-inactive mutant form of MST/Krs protein was overexpressed in cytotrienin A sensitive HL-60 cells, the cytotrienin A-induced apoptosis was partially inhibited. Because cytotrienin A also activated c-Jun N-terminal kinase, we examined the effect of the expression of dominant negative c-Jun on cytotrienin A induced apoptosis. The expression of dominant negative c-Jun also partially inhibited cytotrienin A-induced apoptosis. Furthermore, coexpression of kinase inactive MST/Krs protein and dominant negative c-Jun completely suppressed cytotrienin A-induced apoptosis. These findings suggest that the proteolytic activation of MST/Krs and c-Jun N-terminal kinase activation are involved in cytotrienin A-induced apoptosis in human tumor cell lines. PMID- 10722721 TI - Chaperones Hsp70 and Hsp40 suppress aggregate formation and apoptosis in cultured neuronal cells expressing truncated androgen receptor protein with expanded polyglutamine tract. AB - Spinal and bulbar muscular atrophy (SBMA) is one of a group of human inherited neurodegenerative diseases caused by polyglutamine expansion. We have previously demonstrated that the SBMA gene product, the androgen receptor protein, is toxic and aggregates when truncated. Heat shock proteins function as molecular chaperones, which recognize and renaturate misfolded protein (aggregate). We thus assessed the effect of a variety of chaperones in a cultured neuronal cell model of SBMA. Overexpression of chaperones reduces aggregate formation and suppresses apoptosis in a cultured neuronal cell model of SBMA to differing degrees depending on the chaperones and their combinations. Combination of Hsp70 and Hsp40 was the most effective among the chaperones in reducing aggregate formation and providing cellular protection, reflecting that Hsp70 and Hsp40 act together in chaperoning mutant and disabled proteins. Although Hdj2/Hsdj chaperone has been previously reported to suppress expanded polyglutamine tract-formed aggregate, Hsdj/Hdj2 showed little effect in our system. These findings indicate that chaperones may be one of the key factors in the developing of CAG repeat disease and suggested that increasing expression level or enhancing the function of chaperones will provide an avenue for the treatment of CAG repeat disease. PMID- 10722722 TI - Phragmoplastin polymerizes into spiral coiled structures via intermolecular interaction of two self-assembly domains. AB - Phragmoplastin, a high molecular weight GTPase belonging to the dynamin superfamily of proteins, becomes associated with the cell plate during cytokinesis in plants. Growth of the cell plate requires continuous fusion of vesicles, and phragmoplastin appears to play a role in the formation of vesicle tubule-vesicle structures at the cell plate. In this study, we have demonstrated that two self-assembly domains (SA1 and SA2) are involved in polymerization of phragmoplastin. SA1 is about 42 amino acids long and is located near the N terminus overlapping with the GTP-binding region. SA2, containing at least 24 amino acids, is located in the middle of the molecule outside the GTP-binding domain. Peptides containing either SA1 or SA2 interact efficiently with the full length phragmoplastin. The SA1 domain of one phragmoplastin molecule also binds to SA2 of another as confirmed in vitro by using radiolabeled peptides. This interaction leads to the formation of polymers with a staggered contoured spiral structure. Electron microscopy studies revealed that helical arrays of phragmoplastin can be induced by reducing salt concentration. Our results suggest that phragmoplastin may assemble into helical arrays that wrap around and squeeze vesicles into vesicle-tubule-vesicle structures observed on the forming cell plate. PMID- 10722723 TI - Biosynthesis, post-translation modification, and functional characterization of Drm/Gremlin. AB - Down-regulated by mos (Drm)/Gremlin is a highly conserved protein whose properties and expression pattern suggest a role in early development, tissue specific differentiation, and cell transformation. We have investigated the biosynthesis and processing of Drm expressed endogenously in rat fibroblasts or overexpressed following transient or stable transfection. Analysis of metabolically labeled cells revealed that Drm exists in secreted and cell associated forms that exhibit similar mobilities in SDS-polyacrylamide gel electrophoresis. Protein analysis indicated that Drm is present in two major species: a slow migrating glycosylated form and a nonglycosylated form. Both forms of Drm are able to undergo phosphorylation. Drm is released into the media within 30 min of synthesis and is detectable for up to 4-5 h, whereas the cell associated form has a half-life of about 1 h. Confocal immunofluorescent microscopy indicates that Drm is present both on the external surface of expressing cells, as well as within the endoplasmic reticulum and the Golgi. Both glycosylated and nonglycosylated forms of Drm exhibit identical distributions and are able to antagonize bone morphogenetic protein signaling. Like the soluble form, the cell-associated forms are capable of binding (125)I-bone morphogenetic protein-4. These properties are consistent with a role for Drm in interfering with signaling and indicate that Drm may act at the cell surface during tissue development and transformation. PMID- 10722724 TI - Substrate specificity and inhibition studies of human serotonin N acetyltransferase. AB - Arylalkylamine N-acetyltransferase (AANAT) catalyzes the reaction of serotonin with acetyl-CoA to form N-acetylserotonin and plays a major role in the regulation of the melatonin circadian rhythm in vertebrates. In the present study, the human cloned enzyme has been expressed in bacteria, purified, cleaved, and characterized. The specificity of the human enzyme toward substrates (natural as well as synthetic arylethylamines) and cosubstrates (essentially acyl homologs of acetyl-CoA) has been investigated. Peptide combinatorial libraries of tri-, tetra-, and pentapeptides with various amino acid compositions were also screened as potential sources of inhibitors. We report the findings of several peptides with low micromolar inhibitory potency. For activity measurement as well as for specificity studies, an original and rapid method of analysis was developed. The assay was based on the separation and detection of N-[(3)H]acetylarylethylamine formed from various arylethylamines and tritiated acetyl-CoA, by means of high performance liquid chromatography with radiochemical detection. The assay proved to be robust and flexible, could accommodate the use of numerous synthetic substrates, and was successfully used throughout this study. We also screened a large number of pharmacological bioamines among which only one, tranylcypromine, behaved as a substrate. The synthesis and survey of simple arylethylamines also showed that AANAT has a large recognition pattern, including compounds as different as phenyl-, naphthyl-, benzothienyl-, or benzofuranyl-ethylamine derivatives. An extensive enzymatic study allowed us to pinpoint the amino acid residue of the pentapeptide inhibitor, S 34461, which interacts with the cosubstrate-binding site area, in agreement with an in silico study based on the available coordinates of the hAANAT crystal. PMID- 10722725 TI - Cross-talk between epidermal growth factor receptor and c-Met signal pathways in transformed cells. AB - In rat liver epithelial cells constitutively expressing transforming growth factor alpha (TGFalpha), c-Met is constitutively phosphorylated in the absence of its ligand, hepatocyte growth factor. We proposed that TGFalpha and the autocrine activation of its receptor, epidermal growth factor receptor (EGFR), leads to phosphorylation and activation of c-Met. We found that there is constitutive c Met phosphorylation in human hepatoma cell lines and the human epidermoid carcinoma cell line, A431 which express TGFalpha, but not in normal human hepatocytes. Constitutive c-Met phosphorylation in A431, HepG2, AKN-1, and HuH6 cells was inhibited by neutralizing antibodies against TGFalpha and/or EGFR. Exposure to exogenous TGFalpha or EGF increased the phosphorylation of c-Met in the human epidermoid carcinoma cell line, A431. The increase of c-Met phosphorylation by TGFalpha in A431 cells was inhibited by neutralizing antibodies against TGFalpha and/or EGFR and by the EGFR-specific inhibitor tyrphostin AG1478. These results indicate that constitutive c-Met phosphorylation, and the increase of c-Met phosphorylation by TGFalpha or EGF, in tumor cell lines is the result of the activation via EGFR. We found that c-Met in tumor cells co-immunoprecipitates with EGFR regardless of the existence of their ligands in tumor cells, but not in normal human hepatocytes. We conclude that c Met associates with EGFR in tumor cells, and this association facilitates the phosphorylation of c-Met in the absence of hepatocyte growth factor. This cross talk between c-Met and EGFR may have significant implications for altered growth control in tumorigenesis. PMID- 10722726 TI - Membrane association and protein conformation of alpha-synuclein in intact neurons. Effect of Parkinson's disease-linked mutations. AB - Two missense mutations (Ala-30 --> Pro and Ala-53 --> Thr) in the gene encoding alpha-synuclein are associated with rare autosomal dominant forms of familial Parkinson's disease. In addition, alpha-synuclein is an abundant component of Lewy bodies in sporadic Parkinson's disease and diffuse Lewy body disease. However, the normal conformation of alpha-synuclein, its cellular localization in neurons, and the effects of the mutations remain to be determined. In the present study, we examine these questions using sensitive fluorescence resonance energy transfer techniques. Transient transfection of alpha-synuclein expression constructs into primary cortical neurons and counterstaining with the lipophilic fluorescent marker, DiI, demonstrates a close association between alpha-synuclein and cellular membranes. Both the N- and C-terminal regions of alpha-synuclein are tightly associated with membranes. A weak interaction also occurs between the N and C termini themselves. The Parkinson's disease-associated mutations have no effect on membrane interaction; however, the Ala-30 --> Pro mutation alters the three-dimensional conformation of alpha-synuclein, as measured by significantly increased fluorescence resonance energy transfer between the N and C termini. PMID- 10722727 TI - The Ras/phosphatidylinositol 3-kinase and Ras/ERK pathways function as independent survival modules each of which inhibits a distinct apoptotic signaling pathway in sympathetic neurons. AB - Ras promotes robust survival of many cell systems by activating the phosphatidylinositol 3-kinase (PI3-kinase)/Akt pathway, but little is understood about the survival functions of the Ras/ERK pathway. We have used three different effector-loop mutant forms of Ras, each of which activates a single downstream effector pathway, to dissect their individual contributions to survival of nerve growth factor (NGF)-dependent sympathetic neurons. The PI3-kinase pathway selective protein Ras(Val-12)Y40C was as powerful as oncogenic Ras(Val-12) in preventing apoptosis induced by NGF deprivation but conferred no protection against apoptosis induced by cytosine arabinoside. Identical results were obtained with transfected Akt. In contrast, the ERK pathway-selective protein Ras(Val-12)T35S had no protective effects on NGF-deprived neurons but was almost as strongly protective as Ras(Val-12) against cytosine arabinoside-induced apoptosis. The protective effects of Ras(Val-12)T35S against cytosine arabinoside were completely abolished by the ERK pathway inhibitor PD98059. Ras(Val-12)E37G, an activator of RalGDS, had no survival effect on either death pathway, similar to RasS17N, the full survival antagonist. Thus, Ras provides two independent survival pathways each of which inhibits a distinct apoptotic mechanism. Our study presents one of the few clear-cut cases where only the Ras/ERK, but not the Ras/PI3K/Akt pathway, plays a dominant survival signaling role. PMID- 10722728 TI - The MSG1 non-DNA-binding transactivator binds to the p300/CBP coactivators, enhancing their functional link to the Smad transcription factors. AB - The MSG1 nuclear protein has a strong transcriptional activating activity but does not bind directly to DNA. When cotransfected, MSG1 enhances transcription mediated by the Smad transcription factors in mammalian cells in a manner dependent on ligand-induced Smad hetero-oligomerization. However, the mechanism of this MSG1 effect has been unknown. We now show that MSG1 directly binds to the p300/cAMP-response element-binding protein-binding protein (CBP) transcriptional coactivators, which in turn bind to the Smads, and enhances Smad-mediated transcription in a manner dependent on p300/CBP. The C-terminal transactivating domain of MSG1 is required for binding to p300/CBP and enhancement of Smad mediated transcription; the viral VP16 transactivating domain could not substitute for it. In the N-terminal region of MSG1, we identified a domain that is necessary and sufficient to direct the specific interaction of MSG1 with Smads. We also found that the Hsc70 heat-shock cognate protein also forms complex with MSG1 in vivo, suppressing both binding of MSG1 to p300/CBP and enhancement of Smad-mediated transcription by MSG1. These results indicate that MSG1 interacts with both the DNA-binding Smad proteins and the p300/CBP coactivators through its N- and C-terminal regions, respectively, and enhances the functional link between Smads and p300/CBP. PMID- 10722729 TI - Interleukin-1beta suppresses retinoid transactivation of two hepatic transporter genes involved in bile formation. AB - Cytokines have been implicated in the pathogenesis of inflammatory cholestasis. This is due to transcriptional down-regulation of hepatic transporters including the Na(+)/bile acid cotransporter, ntcp, and the multispecific organic anion exporter, mrp2. We have recently shown that ntcp suppression by lipopolysaccharide in vivo is caused by down-regulation of transactivators including the previously uncharacterized Footprint B-binding protein. Both the ntcp FpB element and the mrp2 promoter contain potential retinoid-response elements. We hypothesized that retinoic acid receptor (RAR) and retinoid X receptor (RXR) heterodimers would activate these two genes and that cytokines that reduce bile flow might do so by suppressing nuclear levels of these transactivators. Retinoid transactivation and interleukin-1beta down-regulation of the ntcp and mrp2 promoters were mapped to RXRalpha:RARalpha-response elements. Gel mobility shift assays demonstrated specific binding of RXRalpha:RARalpha heterodimers to the ntcp and mrp2 retinoid-response elements. The RXRalpha:RARalpha complex was down-regulated by IL-1beta in HepG2 cells. An unexpected finding was that an adjacent CAAT-enhancer-binding protein element was required for maximal transactivation of the ntcp promoter by RXRalpha:RARalpha. Taken together, these studies demonstrate regulation of two hepatobiliary transporter genes by RXRalpha:RARalpha and describe a mechanism which likely contributes to their down-regulation during inflammation. PMID- 10722731 TI - The dominant negative Ras mutant, N17Ras, can inhibit signaling independently of blocking Ras activation. AB - Ras plays an important role in a variety of cellular functions, including growth, differentiation, and oncogenic transformation. For instance, Ras participates in the activation of Raf, which phosphorylates and activates mitogen-activated protein kinase kinase (MEK), which then phosphorylates and activates extracellular signal-regulated kinase (ERK), a mitogen-activated protein (MAP) kinase. Activation of MAP kinase appears to be essential for propagating a wide variety of extracellular signals from the plasma membrane to the nucleus. N17Ras, a GDP-bound dominant negative mutant, is used widely as an interfering mutant to assess Ras function in vivo. Surprisingly, we observed that expression of N17Ras inhibited the activity and phosphorylation of Elk-1, a physiological substrate of MAP kinases, in response to phorbol myristate acetate. The activity and phosphorylation of the MAP kinase hemagglutinin epitope (HA)-ERK1 were not affected by N17Ras in response to the same stimulus. Additionally, expression of N17Ras, but not L61S186Ras, a GTP-bound interfering mutant, inhibited MEK-induced Elk-1 phosphorylation, suggesting that inhibition of Elk-1 may be unique to GDP bound Ras mutants. Finally, we observed that V12Ras-induced focus formation in NIH3T3 cells is inhibited by coexpression of GDP-bound Ras mutants, such as N17, A15, and N17N69. Therefore, N17Ras and V12 Ras may be codominant with respect to Elk-1 activation and cellular transformation. These results indicate that N17Ras appears to have at least two distinguishable functions: interference with endogenous Ras activation and inhibition of Elk-1 and transfomation. Furthermore, our data imply the possibility that GDP-bound Ras, like N17Ras, may have a direct role in signal transduction. PMID- 10722730 TI - Cloning and characterization of a new member of the Nudix hydrolases from human and mouse. AB - Proteins containing the Nudix box "GX(5)EX(7)REUXEEXGU" (where U is usually Leu, Val, or Ile) are Nudix hydrolases, which catalyze the hydrolysis of a variety of nucleoside diphosphate derivatives. Here we report cloning and characterization of a human cDNA encoding a novel nudix hydrolase NUDT5 for the hydrolysis of ADP sugars. The deduced amino acid sequence of NUDT5 contains 219 amino acids, including a conserved Nudix box sequence. The recombinant NUDT5 was expressed in Escherichia coli and purified to near homogeneity. At the optimal pH of 7, the purified recombinant NUDT5 catalyzed hydrolysis of two major substrates ADP ribose and ADP-mannose with K(m) values of 32 and 83 microM, respectively; the V(max) for ADP-mannose was about 1.5 times that with ADP-ribose. The murine NUDT5 homolog was also cloned and characterized. mNudT5 has 81% amino acid identity to NUDT5 with catalytic activities similar to NUDT5 under the optimal pH of 9. Both NUDT5 and mNudT5 transcripts were ubiquitously expressed in tissues analyzed with preferential abundance in liver. The genomic structures of both NUDT5 and mNudT5 were determined and located on human chromosome 10 and mouse chromosome 2, respectively. The role of NUDT5 in maintaining levels of free ADP-ribose in cells is discussed. PMID- 10722732 TI - Purification, cloning, and characterization of an acidic ectoprotein phosphatase differentially expressed in the infectious bloodstream form of Trypanosoma brucei. AB - We purified an ecto-phosphatase of 115 kDa (TryAcP115) specifically expressed by bloodstream forms of Trypanosoma brucei. The corresponding gene coded for a 45 kDa protein potentially including a signal peptide, a membrane-spanning domain and an N-terminal domain containing 8 N-glycosylation sites. There was no significant sequence homology with other phosphatases. Antiserum to the Escherichia coli recombinant N-terminal domain, Petase7, recognized a protein of 55 kDa in Western blots after deglycosylation of the TryAcP115 protein by N glycosidase F. Immunofluorescence and trypsin treatment of living parasites showed that TryAcP115 was localized to the surface of the parasite and that its N terminal domain was oriented extracellularly. The recombinant N-terminal domains, expressed in E. coli and Leishmania amazonensis, harbored phosphatase activity against Tyr(P)-Raytide, Ser(P)-neurogranin, and ATP. The enzymatic properties of native TryAcP115 and the recombinant proteins for the substrate Tyr(P)-Raytide were virtually identical and included: (i) K(m) and V(max) values of 15 nM and 200 pmol/min/mg, (ii) no requirement for divalent cations, and (iii) sensitivity to vanadate, sodium fluoride, and tartrate, but insensitivity to okadaic acid and tetramisole. Although the function of TryAcP115 remains unknown, a differentially expressed, unique ecto-phosphatase could regulate growth or influence parasite host interactions and might provide a useful target for chemotherapy. PMID- 10722733 TI - Suppression of heat shock protein-70 by ceramide in heat shock-induced HL-60 cell apoptosis. AB - Ceramide has emerged as a mediator of cell growth, differentiation, and apoptosis in many biological systems. Many kinds of stresses are reported to induce apoptosis with an increase of ceramide generation. Here we showed that the intracellular ceramide levels increased in parallel with heat shock (HS)-induced apoptosis in an intensity- and time-dependent manner, and synthetic N acetylsphingosine (C(2)-ceramide) synergistically enhanced HS-induced apoptosis in HL-60 cells. In order to know the role of ceramide generation in HS-induced apoptosis, we examined the effects of C(2)-ceramide on the levels of mRNA and protein of heat shock proteins (HSPs). The increase of HSP-70 mRNA levels 1-2 h after HS at 42 degrees C for 30 min was suppressed by C(2)-ceramide in a dose dependent manner. In comparison with HSP-70, the levels of HSP-60 and -90 mRNAs were faintly suppressed by C(2)-ceramide. Similarly, the increase in the protein levels of HSP-70 was significantly suppressed 4-8 h after HS by C(2)-ceramide in a dose-dependent manner. Additionally, in 293 cells, which are constitutively overexpressing HSP-70 gene, the levels of HSP-70 mRNA were suppressed by C(2) ceramide in parallel with the increase of apoptotic cells. We next examined the mechanisms by which C(2)-ceramide suppressed HS-increased HSP-70 expression. The treatment with C(2)-ceramide did not affect both an activation of a nuclear transcription factor for HSP-70, heat shock factor-1, and an increased transcriptional rate of HSP-70 by HS, but increased the rates of HSP-70 mRNA degradation. In summary, ceramide may efficiently induce HS-induced apoptosis by suppressing anti-apoptotic HSP-70 through a post-transcriptional regulation. PMID- 10722734 TI - RNase-L-dependent destabilization of interferon-induced mRNAs. A role for the 2 5A system in attenuation of the interferon response. AB - The 2-5A system is an interferon-regulated RNA degradation pathway with antiviral, growth-inhibitory, and pro-apoptotic activities. RNase-L mediates the antiviral activity through the degradation of viral RNAs, and the anticellular effects of the 2-5A system are thought to be similarly mediated through the degradation of cellular transcripts. However, specific RNase-L-regulated cellular RNAs have not been identified. To isolate candidate RNase-L substrates, differential display was used to identify mRNAs that exhibited increased expression in RNase-L-deficient N1E-115 cells as compared with RNase-L transfected cells. A novel interferon-stimulated gene encoding a 43-kDa ubiquitin specific protease, designated ISG43, was identified in this screen. ISG43 expression is induced by interferon and negatively regulated by RNase-L. ISG43 induction is a primary response to interferon treatment and requires a functional JAK/STAT signaling pathway. The kinetics of ISG43 induction were identical in wild type and RNase-L knock-out fibroblasts; however, the decline in ISG43 mRNA following interferon treatment was markedly attenuated in RNase-L knock-out fibroblasts. The delayed shut-off kinetics of ISG43 mRNA corresponded to an increase in its half-life in RNase-L-deficient cells. ISG15 mRNA also displayed RNase-L-dependent regulation. These findings identify a novel role for the 2-5A system in the attenuation of the interferon response. PMID- 10722735 TI - The structures of the H(C) fragment of tetanus toxin with carbohydrate subunit complexes provide insight into ganglioside binding. AB - The entry of tetanus neurotoxin into neuronal cells proceeds through the initial binding of the toxin to gangliosides on the cell surface. The carboxyl-terminal fragment of the heavy chain of tetanus neurotoxin contains the ganglioside binding site, which has not yet been fully characterized. The crystal structures of native H(C) and of H(C) soaked with carbohydrates reveal a number of binding sites and provide insight into the possible mode of ganglioside binding. PMID- 10722736 TI - Identification and functional characterization of a conserved, nuclear factor 1 like element in the proximal promoter region of CYP1A2 gene specifically expressed in the liver and olfactory mucosa. AB - CYP1A2 is a major cytochrome P-450 isoform in the liver and the olfactory mucosa but is essentially not expressed in other tissues. A nuclear factor 1 (NF-1) like element was identified in the proximal promoter region of rat, mouse, rabbit, and human CYP1A2 genes through data base analysis. In vitro DNase I footprinting with a -211 to +81 probe from the rat CYP1A2 gene and nuclear extracts from rat liver and olfactory mucosa revealed a single protected region corresponding to the NF-1-like element at -129 to -111. Protein binding to this NF-1-like element was tissue-selective and was confirmed by in vivo footprinting in native chromatin from rat liver. Multiple DNA-binding complexes were detected in gel-shift assays using the CYP1A2 NF-1-like element and nuclear extracts from liver and olfactory mucosa, all of which were supershifted in the presence of an anti-NF1 antibody. The NF-1-like element was essential for transcriptional activity of the CYP1A2 gene in an in vitro transcription assay using nuclear extracts from the two tissues. Thus, members of the NF-1 family of transcription factors may play an important role in the tissue-selective expression of the CYP1A2 gene in the liver and olfactory mucosa. PMID- 10722737 TI - Cell-specific transcription of leukotriene C(4) synthase involves a Kruppel-like transcription factor and Sp1. AB - Leukotriene C(4) synthase (LTC(4)S) is responsible for the biosynthesis of cysteinyl leukotrienes that participate in allergic and asthmatic inflammation. We analyzed 2.1 kilobases of the 5'-flanking region of the human LTC(4)S gene, which contains three DNase I hypersensitivity sites, for its transcriptional activity when fused to a promoterless and enhancerless luciferase gene. Deletion analysis revealed a nonspecific basal promoter region between nucleotides -122 and -56 upstream of the translation start site which contains a consensus Sp1 binding site and a putative initiator element (Inr) and cell-specific enhancer regions further upstream. A single mutation of either the Sp1 binding site between nucleotides -120 and -115 or the Inr (CAGAC) between nucleotides -66 and 62 reduced the expression of the reporter gene by approximately 60%, whereas double mutations decreased the expression by approximately 80%. The incubation of nuclear extracts from THP-1 and K562 cells with a (32)P-labeled oligonucleotide containing the Sp1 site or the Inr sequence gave gel-shifted complexes that were blocked by their respective cold oligonucleotides, and antisera specific for Sp1 and Sp3 provided supershifts for the former. Linker-scanning mutations of a cell specific regulatory region revealed that mutations from nucleotides -165 to -125 reduced reporter activity. This region contains a tandem CACCC repeat (at nucleotides -149 to -145 and -139 to -135). An oligonucleotide containing the distal CACCC motif was gel shifted by THP-1 cell nuclear extract and was supershifted by antisera to Sp1 and Sp3. Cotransfection of an Sp1 expression plasmid into Drosophila SL2 cells with a -228 to -3 LTC(4)S reporter construct transactivated the reporter gene, whereas mutations at the CACCC repeat region reduced Sp1 transactivation by approximately 66%. Similarly, the Kruppel-like factor Zf9/CPBP (core promoter-binding protein) transactivated the -228 construct in COS cells but not its CACCC mutant construct. These findings indicate the involvement of Sp1 and an Inr in non-cell-specific regulation and a Kruppel-like transcription factor and Sp1 in the cell-specific regulation of the LTC(4)S gene. These are the first such analyses of a member of a newly recognized superfamily of membrane-associated proteins involved in eicosanoid and glutathione metabolism, which contains key proteins involved in the generation of both prostanoids and cysteinyl leukotrienes. PMID- 10722738 TI - cAMP-independent activation of the adenovirus type 12 E2 promoter correlates with the recruitment of CREB-1/ATF-1, E1A(12S), and CBP to the E2-CRE. AB - Expression of the transcription unit early region 2 (E2) is of crucial importance for adenoviruses because this region encodes proteins essential for viral replication. Here, we demonstrate that the E1A(12S) protein of the oncogenic adenovirus serotype 12 activates the E2 promoter in dependence of the N terminus and the conserved region 1. Activation is mediated through a cAMP-response element that is bound by CREB-1 and ATF-1. Moreover, the Ad12 E2 promoter is inducible by protein kinase A and repressed by either a dominant-negative cAMP response element-binding protein (CREB) mutant or the highly specific protein kinase A inhibitor protein underscoring the participation of CREB-1/ATF-1 in promoter activation. E1A(12S) binds to CREB-1 and ATF-1 in dependence of the N terminus and CR1 and is recruited to the E2 cAMP-response element through both cellular transcription factors. Most interestingly, point mutations revealed that E1A(12S) domains essential for binding to CREB-1/ATF-1 and for activation of the Ad12 E2 promoter are also essential for binding to the CREB-binding protein. Due to these data and results obtained in DNA-dependent protein-protein interaction assays, we propose a model in which the cAMP-independent activation of the Ad12 E2 promoter is mediated through a ternary complex consisting of CREB-1/ATF-1, E1A(12S), and CREB-binding protein, which assembles on the E2 cAMP-response element. PMID- 10722739 TI - GABA(A) receptor assembly. Identification and structure of gamma(2) sequences forming the intersubunit contacts with alpha(1) and beta(3) subunits. AB - GABA(A) receptors are ligand-gated chloride channels composed of five homologous subunits that specifically recognize one another and assemble around an aqueous pore. To identify domains responsible for the specificity of subunit association, we constructed C-terminal truncated gamma(2) subunits, as well as mutated and chimeric fragments. From their ability to interfere with alpha(1)beta(3)gamma(2) receptor assembly and to associate with full-length subunits, we concluded that amino acid sequences gamma(2)-(91-104) and gamma(2)-(83-90) form the sites mediating assembly with alpha(1) and beta(3) subunits, respectively. Neural network-based secondary structure prediction, Monte Carlo optimization, and hydrophobicity analysis led to the conclusion that these sites also form the intersubunit contacts in the completely assembled receptor and provided important information on the benzodiazepine-binding site and structure of GABA(A) receptors. PMID- 10722740 TI - The amyloid precursor protein-binding protein APP-BP1 drives the cell cycle through the S-M checkpoint and causes apoptosis in neurons. AB - APP-BP1 binds to the amyloid precursor protein (APP) carboxyl-terminal domain. Recent work suggests that APP-BP1 participates in a novel ubiquitinylation related pathway involving the ubiquitin-like molecule NEDD8. We show here that, in vivo in mammalian cells, APP-BP1 interacts with hUba3, its presumptive partner in the NEDD8 activation pathway, and that the APP-BP1 binding site for hUba3 is within amino acids 443-479. We also provide evidence that the human APP-BP1 molecule can rescue the ts41 mutation in Chinese hamster cells. This mutation previously has been shown to lead to successive S phases of the cell cycle without intervening G(2), M, and G(1), suggesting that the product of this gene negatively regulates entry into the S phase and positively regulates entry into mitosis. We show that expression of APP-BP1 in ts41 cells drives the cell cycle through the S-M checkpoint and that this function requires both hUba3 and hUbc12. Overexpression of APP-BP1 in primary neurons causes apoptosis via the same pathway. A specific caspase-6 inhibitor blocks this apoptosis. These findings are discussed in the context of abnormalities in the cell cycle that have been observed in Alzheimer's disease. PMID- 10722741 TI - Type XIII collagen forms homotrimers with three triple helical collagenous domains and its association into disulfide-bonded trimers is enhanced by prolyl 4 hydroxylase. AB - Type XIII collagen is a type II transmembrane protein predicted to consist of a short cytosolic domain, a single transmembrane domain, and three collagenous domains flanked by noncollagenous sequences. Previous studies on mRNAs indicate that the structures of the collagenous domain closest to the cell membrane, COL1, the adjacent noncollagenous domain, NC2, and the C-terminal domains COL3 and NC4 are subject to alternative splicing. In order to extend studies of type XIII collagen from cDNAs to the protein level we have produced it in insect cells by means of baculoviruses. Type XIII collagen alpha chains were found to associate into disulfide-bonded trimers, and hydroxylation of proline residues dramatically enhanced this association. This protein contains altogether eight cysteine residues, and interchain disulfide bonds could be located in the NC1 domain and possibly at the junction of COL1 and NC2, while the two cysteine residues in NC4 are likely to form intrachain bonds. Pepsin and trypsin/chymotrypsin digestions indicated that the type XIII collagen alpha chains form homotrimers whose three collagenous domains are in triple helical conformation. The thermal stabilities (T(m)) of the COL1, COL2, and COL3 domains were 38, 49 and 40 degrees C, respectively. The T(m) of the central collagenous domain is unusually high, which in the light of this domain being invariant in terms of alternative splicing suggests that the central portion of the molecule may have an important role in the stability of the molecule. All in all, most of the type XIII collagen ectodomain appears to be present in triple helical conformation, which is in clear contrast to the short or highly interrupted triple helical domains of the other known collagenous transmembrane proteins. PMID- 10722742 TI - Mdm2 is a RING finger-dependent ubiquitin protein ligase for itself and p53. AB - Mdm2 has been shown to regulate p53 stability by targeting the p53 protein for proteasomal degradation. We now report that Mdm2 is a ubiquitin protein ligase (E3) for p53 and that its activity is dependent on its RING finger. Furthermore, we show that Mdm2 mediates its own ubiquitination in a RING finger-dependent manner, which requires no eukaryotic proteins other than ubiquitin-activating enzyme (E1) and an ubiquitin-conjugating enzyme (E2). It is apparent, therefore, that Mdm2 manifests an intrinsic capacity to mediate ubiquitination. Mutation of putative zinc coordination residues abrogated this activity, as did chelation of divalent cations. After cation chelation, the full activity could be restored by addition of zinc. We further demonstrate that the degradation of p53 and Mdm2 in cells requires additional potential zinc-coordinating residues beyond those required for the intrinsic activity of Mdm2 in vitro. Replacement of the Mdm2 RING with that of another protein (Praja1) reconstituted ubiquitination and proteasomal degradation of Mdm2. However, this RING was ineffective in ubiquitination and proteasomal targeting of p53, suggesting that there may be specificity at the level of the RING in the recognition of heterologous substrates. PMID- 10722743 TI - Activation of a Plasmodium falciparum cdc2-related kinase by heterologous p25 and cyclin H. Functional characterization of a P. falciparum cyclin homologue. AB - Several Plasmodium falciparum genes encoding cdc2-related protein kinases have been identified, but the modalities of their regulation remains largely unexplored. In the present study, we investigated the regulation in vitro of PfPK5, a putative homologue of Cdk1 (cdc2) in P. falciparum. We show that (i) PfPK5 is efficiently activated by heterologous (human) cyclin H and p25, a cyclin like molecule that specifically activates human Cdk5; (ii) the activated enzyme can be inhibited by chemical Cdk inhibitors; (iii) Pfmrk, a putative P. falciparum homologue of the Cdk-activating kinase, does neither activate nor phosphorylate PfPK5; and (iv) PfPK5 is able to autophosphorylate in the presence of a cyclin. Taken together, these results suggest that the regulation of Plasmodium Cdks may differ in important aspects from that of their human counterparts. Furthermore, we cloned an open reading frame encoding a novel P. falciparum protein possessing maximal homology to cyclin H from various organisms, and we show that this protein, called Pfcyc-1, is able to activate recombinant PfPK5 in vitro with an efficiency similar to that of human cyclin H and p25. This work opens the way to the development of screening procedures aimed at identifying compounds that specifically target the parasite Cdks. PMID- 10722744 TI - Structural and functional characterization of the leukocyte integrin gene CD11d. Essential role of Sp1 and Sp3. AB - CD11d encodes the latest alpha-subunit of the leukocyte integrin family to be discovered, and it is expressed predominantly in myelomonocytic cells. We have isolated a genomic clone that contains CD11d and showed this gene to be 11,461 bp downstream and oriented in the same direction as the related CD11c gene. CD11d transcription begins 69-79 nucleotides upstream of the ATG codon. Transfection analysis of CD11d-luc reporter constructs revealed that the -173 to +74 region is sufficient to confer leukocyte-specific expression of luciferase in myelomonocytic cells (THP1 and HL60), B-cells (IM9), and T-cells (Jurkat). Transfection analysis showed that down-regulation of CD11d expression by phorbol ester was myelomonocyte-specific and is mediated by one or more cis-elements within the -173 to +74 region. In vitro DNase I footprint analysis and electrophoretic mobility shift analysis showed that Sp1 and Sp3 bind at -63 to 40. Deletion of the Sp-binding site significantly reduced CD11d promoter activity. Overexpression of either Sp1 or Sp3 in THP1 cells led to activation of the CD11d promoter even in the presence of phorbol ester, whereas down-regulation of either factor by antisense oligonucleotides decreased CD11d promoter activity. In contrast, overexpression of Sp3 in IM9 and Jurkat cells down-regulated CD11d promoter expression. In vivo genomic footprinting revealed that the -63 to -40 region is bound by a Sp protein in unstimulated HL60 cells but not in phorbol ester-stimulated HL60 cells. In contrast, this site is bound in both unstimulated and phorbol ester-stimulated IM9 and Jurkat cells. Together, these results show that myelomonocyte-specific phorbol ester down-regulation of CD11d is mediated through both Sp1 and Sp3. PMID- 10722745 TI - Salivary film expresses a complex, macromolecular binding site for Streptococcus sanguis. AB - Teeth in the oral cavity are coated with a salivary film or pellicle, which lacks apparent intermolecular organization. This heterogeneous film facilitates binding of early commensal colonizing bacteria, including Streptococcus sanguis. To test the hypothesis that sufficient intermolecular organization exists in salivary films to form binding sites for S. sanguis, an in vitro model of saliva-coated teeth was probed with murine anti-idiotypical monoclonal antibodies (mAb2, anti ids). The anti-ids were harvested from hybridomas that were developed in response to first generation murine hybridomas that produced anti-S. sanguis adhesin monoclonal antibodies (mAb1). The anti-ids (i) reacted with experimental salivary films and inhibited S. sanguis adhesion in a dose-dependent fashion. In Western blots, the anti-ids (ii) recognized a high molecular weight salivary antigen and (iii) secretory IgA (sIgA) light chain and alpha-amylase. After isolation by gel filtration from whole saliva or mixed secretory IgA and alpha-amylase, the high molecular weight component, containing amylase activity and sIgA, bound to hydroxyapatite to promote adhesion of S. sanguis. Therefore, a complex enriched in secretory immunoglobulin A and alpha-amylase forms a S. sanguis-binding site. PMID- 10722746 TI - Molecular cloning and expression of chondroitin 4-sulfotransferase. AB - Chondroitin 4-sulfotransferase (C4ST) catalyzes the transfer of sulfate from 3' phosphoadenosine 5'-phosphosulfate to position 4 of N-acetylgalactosamine residue of chondroitin. The enzyme has been previously purified to apparent homogeneity from the serum-free culture medium of rat chondrosarcoma cells (Yamauchi, A., Hirahara, Y., Usui, H., Takeda, Y., Hoshino, M., Fukuta, M., Kimura, J. H., and Habuchi, O. (1999) J. Biol. Chem. 274, 2456-2463). The purified enzyme also catalyzed the sulfation of partially desulfated dermatan sulfate. We have now cloned the cDNA of the mouse C4ST on the basis of the amino acid sequences of peptides obtained from the purified enzyme by protease digestion. This cDNA contains a single open reading frame that predicts a protein composed of 352 amino acid residues. The protein predicts a Type II transmembrane topology. The predicted sequence of the protein contains all of the known amino acid sequence and four potential sites for N-glycosylation, which corresponds to the observation that the purified C4ST is an N-linked glycoprotein. The amino acid sequence of mouse C4ST showed significant sequence homology to HNK-1 sulfotransferase. Comparison of the sequence of mouse C4ST with human HNK-1 sulfotransferase revealed approximately 29% identity and approximately 48% similarity at the amino acid level. When the cDNA was introduced in a eukaryotic expression vector and transfected in COS-7 cells, the sulfotransferase activity that catalyzes the transfer of sulfate to position 4 of GalNAc residue of both chondroitin and desulfated dermatan sulfate was overexpressed. Northern blot analysis showed that, among various mouse adult tissues, 5.7-kilobase message of C4ST was mainly expressed in the brain and kidney. PMID- 10722747 TI - Down-regulation of the neurotrophin receptor TrkB following ligand binding. Evidence for an involvement of the proteasome and differential regulation of TrkA and TrkB. AB - This study examines the mechanisms by which the tyrosine kinase receptor TrkB is down-regulated following binding of brain-derived neurotrophic factor (BDNF). In primary cultures of cerebellar granule neurons, BDNF-induced reduction of TrkB receptors was largely prevented by the addition of specific proteasome inhibitors. HN10 cells, a neuronal cell line that can be readily transfected, also showed a marked down-regulation of cell surface TrkB following BDNF exposure. In addition, we observed that prolonged exposure to nerve growth factor of TrkA-transfected cells did not lead to the down-regulation seen with BDNF and TrkB. TrkA and TrkB chimeric molecules were therefore expressed in HN10 cells and tested for ligand-induced regulation. These experiments led to the conclusion that the motives responsible for down-regulation are contained in the cytoplasmic domain of TrkB, and a short sequence in the juxtamembrane domain of TrkB was identified that confers nerve growth factor-induced down-regulation when inserted into TrkA. PMID- 10722748 TI - Ran-mediated nuclear export of the von Hippel-Lindau tumor suppressor protein occurs independently of its assembly with cullin-2. AB - Inactivating mutations of the von Hippel-Lindau (VHL) tumor suppressor gene cause the VHL cancer syndrome and sporadic renal clear cell carcinoma. VHL engages in a nucleocytoplasmic shuttle, which is required for its function. Here, we pursue our investigation to identify mechanisms by which VHL-green fluorescent protein (VHL-GFP) is exported from the nucleus. We show that nuclear export of VHL-GFP in living cells requires ongoing RNA polymerase II activity, and is mediated by mechanisms that are temperature-sensitive and energy-dependent. In vitro nuclear export of VHL-GFP is inhibited by nuclear pore-specific lectins, requires ATP hydrolysis and polyadenylated mRNAs, and occurs with kinetics that are similar to those of proteins containing a nuclear export signal. Biochemical fractionation has revealed that nuclear export of VHL-GFP occurs by way of a Ran-dependent pathway. Size exclusion column chromatography and deletion mutant analysis suggest that VHL-GFP does not require assembly with one of its associated proteins, cullin-2, to engage in nuclear export. These results demonstrate that nuclear export of VHL-GFP is Ran-mediated and ATP hydrolysis-dependent. They also suggest that sequences outside the elongin C binding box may function as a nuclear export domain, potentially providing a novel role for this region of VHL frequently mutated in renal cell carcinoma. PMID- 10722749 TI - Identification of endoglycan, a member of the CD34/podocalyxin family of sialomucins. AB - CD34 and podocalyxin are structurally related sialomucins, which are expressed in multiple tissues including vascular endothelium and hematopoietic progenitors. These glycoproteins have been proposed to be involved in processes as diverse as glomerular filtration, inhibition of stem cell differentiation, and leukocyte endothelial adhesion. Using homologies present in the cytoplasmic tails of these proteins, we have identified a novel member of this family, which we designate endoglycan. This protein shares a similar overall domain structure with the other family members including a sialomucin domain, but also possesses an extremely acidic amino-terminal region. In addition, endoglycan contains several potential glycosaminoglycan attachment sites and is modified with chondroitin sulfate. Endoglycan mRNA and protein were detected in both endothelial cells and CD34(+) bone marrow cells. Thus, CD34, podocalyxin, and endoglycan comprise a family of sialomucins sharing both structural similarity and sequence homology, which are expressed by both endothelium and multipotent hematopoietic progenitors. While the members of this family may perform overlapping functions at these sites, the unique structural features of endoglycan suggest distinct functions for this molecule. PMID- 10722750 TI - Characterization of the N-terminal domain of the yeast transcriptional repressor Tup1. Proposal for an association model of the repressor complex Tup1 x Ssn6. AB - The yeast Tup1 and Ssn6 proteins form a transcriptional repression complex that represses transcription of a broad array of genes. It has been shown that the N terminal domain of the Tup1 protein interacts with a region of the Ssn6 protein that consists of 10 tandem copies of a tetratricopeptide motif. In this work, we use a surface plasmon resonance assay to measure the affinity of the N-terminal domain of Tup1 for a minimal 3-TPR domain of Saccharomyces cerevisiae Ssn6 that is sufficient for binding to Tup1. This domain of Ssn6 binds with comparable affinity to S. cerevisiae and Candida albicans Tup1, but with 100-fold lower affinity to Tup1 protein containing a point mutation that gives rise to a defect in repression in vivo. Results from studies using analytical ultracentrifugation, CD spectroscopy, limited proteolysis, and (1)H NMR show that this domain of Tup1 is primarily alpha-helical and forms a stable tetramer that is highly nonglobular in shape. X-ray diffraction recorded from poorly ordered crystals of the Tup1 tetramerization domain contains fiber diffraction typical of a coiled coil. Our results are used to propose a model for the structure of the N-terminal domain of Tup1 and its interaction with the Ssn6 protein. PMID- 10722751 TI - Formation of spherical, reconstituted high density lipoproteins containing both apolipoproteins A-I and A-II is mediated by lecithin:cholesterol acyltransferase. AB - Previous studies have provided detailed information on the formation of spherical high density lipoproteins (HDL) containing apolipoprotein (apo) A-I but no apoA II (A-I HDL) by an lecithin:cholesterol acyltransferase (LCAT)-mediated process. In this study we have investigated the formation of spherical HDL containing both apoA-I and apoA-II (A-I/A-II HDL). Incubations were carried out containing discoidal A-I reconstituted HDL (rHDL), discoidal A-II rHDL, and low density lipoproteins in the absence or presence of LCAT. After the incubation, the rHDL were reisolated and subjected to immunoaffinity chromatography to determine whether A-I/A-II rHDL were formed. In the absence of LCAT, the majority of the rHDL remained as either A-I rHDL or A-II rHDL, with only a small amount of A-I/A II rHDL present. By contrast, when LCAT was present, a substantial proportion of the reisolated rHDL were A-I/A-II rHDL. The identity of the particles was confirmed using apoA-I rocket electrophoresis. The formation of the A-I/A-II rHDL was influenced by the relative concentrations of the precursor discoidal A-I and A-II rHDL. The A-I/A-II rHDL included several populations of HDL-sized particles; the predominant population having a Stokes' diameter of 9.9 nm. The particles were spherical in shape and had an electrophoretic mobility slightly slower than that of the alpha-migrating HDL in human plasma. The apoA-I:apoA-II molar ratio of the A-I/A-II rHDL was 0.7:1. Their major lipid constituents were phospholipids, unesterified cholesterol, and cholesteryl esters. The results presented are consistent with LCAT promoting fusion of the A-I rHDL and A-II rHDL to form spherical A-I/A-II rHDL. We suggest that this process may be an important source of A-I/A-II HDL in human plasma. PMID- 10722752 TI - Identification of Gbetagamma binding sites in the third intracellular loop of the M(3)-muscarinic receptor and their role in receptor regulation. AB - Gbetagamma binds directly to the third intracellular (i3) loop subdomain of the M(3)-muscarinic receptor (MR). In this report, we identified the Gbetagamma binding motif and G-protein-coupled receptor kinase (GRK2) phosphorylation sites in the M(3)-MR i3 loop via a strategy of deletional and site-directed mutagenesis. The Gbetagamma binding domain was localized to Cys(289)-His(330) within the M(3)-MR-Arg(252)-Gln(490) i3 loop, and the binding properties (affinity, influence of ionic strength) of the M(3)-MR-Cys(289)-His(330) i3 loop subdomain were similar to those observed for the entire i3 loop. Site-directed mutagenesis of the M(3)-MR-Cys(289)-His(330) i3 loop subdomain indicated that Phe(312), Phe(314), and a negatively charged region (Glu(324)-Asp(329)) were required for interaction with Gbetagamma. Generation of the full-length M(3)-MR Arg(252)-Gln(490) i3 peptides containing the F312A mutation were also deficient in Gbetagamma binding and exhibited a reduced capacity for phosphorylation by GRK2. A similar, parallel strategy resulted in identification of major residues ((331)SSS(333) and (348)SASS(351)) phosphorylated by GRK2, which were just downstream of the Gbetagamma binding motif. Full-length M(3)-MR constructs lacking the 42-amino acid Gbetagamma binding domain (Cys(289)-His(330)) or containing the F312A mutation exhibited ligand recognition properties similar to wild type receptor and also effectively mediated agonist-induced increases in intracellular calcium following receptor expression in Chinese hamster ovary and/or COS 7 cells. However, the M(3)-MRDeltaCys(289)-His(330) and M(3)-MR(F312A) constructs were deficient in agonist-induced sequestration, indicating a key role for the Gbetagamma-M(3)-MR i3 loop interaction in receptor regulation and signal processing. PMID- 10722753 TI - Comparison of nucleosome remodeling by the yeast transcription factor Pho4 and the glucocorticoid receptor. AB - Chromatin reorganization of the PHO5 and murine mammary tumor virus (MMTV) promoters is triggered by binding of either Pho4 or the glucocorticoid receptor (GR), respectively. In order to compare the ability of Pho4 and GR to remodel chromatin and activate transcription, hybrid promoter constructs were created by insertion of the MMTV B nucleosome sequence into the PHO5 promoter and then transformed into a yeast strain expressing GR. Activation of either Pho4 (by phosphate depletion) or GR (by hormone addition) resulted in only slight induction of hybrid promoter activity. However, simultaneous activation of both Pho4 and GR resulted in synergistic activation to levels exceeding that of the wild type PHO5 promoter. Under these conditions, Pho4 completely disrupted the nucleosome containing its binding site. In contrast, GR had little effect on the stability of the MMTV B nucleosome. A minimal transactivation domain of the GR fused to the Pho4 DNA-binding domain is capable of efficiently disrupting the nucleosome with a Pho4-binding site, whereas the complementary hybrid protein (Pho4 activation domain, GR DNA-binding domain) does not labilize the B nucleosome. Therefore, we conclude that significant activation by Pho4 requires nucleosome disruption, whereas equivalent transcriptional activation by GR is not accompanied by overt perturbation of nucleosome structure. Our results show that the DNA-binding domains of the two factors play critical roles in determining how chromatin structure is modified during promoter activation. PMID- 10722754 TI - Inhibition of caspase-3-mediated poly(ADP-ribose) polymerase (PARP) apoptotic cleavage by human PARP autoantibodies and effect on cells undergoing apoptosis. AB - Autoantibodies directed to nuclear antigens are serological hallmarks of autoimmune rheumatic diseases such as systemic lupus erythematosus. Although much more is known about the molecular identity and functions of targeted self antigens, with few exceptions, evidence that autoantibodies to these targets have a particular function and contribute directly to the pathological process is lacking. Here we show that human autoantibodies reacting with the zinc fingers of poly(ADP-ribose) polymerase involved in the recognition of damaged DNA totally prevent the cleavage of poly(ADP-ribose) polymerase by caspase-3, a process that normally occurs during early apoptosis. Furthermore, these antibodies, which are frequent in certain autoimmune rheumatic and bowel diseases, affect the characteristic features of apoptosis and increase cell survival ex vivo. This new observation is important, because failure to remove autoimmune or abnormal cells can give rise to prolonged autoimmune stimulation and tumor formation. PMID- 10722755 TI - The c-Jun NH(2)-terminal kinase promotes insulin resistance during association with insulin receptor substrate-1 and phosphorylation of Ser(307). AB - Tumor necrosis factor alpha (TNFalpha) inhibits insulin action, in part, through serine phosphorylation of IRS proteins; however, the phosphorylation sites that mediate the inhibition are unknown. TNFalpha promotes multipotential signal transduction cascades, including the activation of the Jun NH(2)-terminal kinase (JNK). Endogenous JNK associates with IRS-1 in Chinese hamster ovary cells. Anisomycin, a strong activator of JNK in these cells, stimulates the activity of JNK bound to IRS-1 and inhibits the insulin-stimulated tyrosine phosphorylation of IRS-1. Serine 307 is a major site of JNK phosphorylation in IRS-1. Mutation of serine 307 to alanine eliminates phosphorylation of IRS-1 by JNK and abrogates the inhibitory effect of TNFalpha on insulin-stimulated tyrosine phosphorylation of IRS-1. These results suggest that phosphorylation of serine 307 might mediate, at least partially, the inhibitory effect of proinflammatory cytokines like TNFalpha on IRS-1 function. PMID- 10722756 TI - A critical role for the proteasome activator PA28 in the Hsp90-dependent protein refolding. AB - The 90-kDa heat shock protein, Hsp90, was previously shown to capture firefly luciferase during thermal inactivation and prevent it from undergoing an irreversible off-pathway aggregation, thereby maintaining it in a folding competent state. While Hsp90 by itself was not sufficient to refold the denatured luciferase, addition of rabbit reticulocyte lysate remarkably restored the luciferase activity. Here we demonstrate that Hsc70, Hsp40, and the 20 S proteasome activator PA28 are the effective components in reticulocyte lysate. Purified Hsc70, Hsp40, and PA28 were necessary and sufficient to fully reconstitute Hsp90-initiated refolding. Kinetics of substrate binding support the idea that PA28 acts as the molecular link between the Hsp90-dependent capture of unfolded proteins and the Hsc70- and ATP-dependent refolding process. PMID- 10722757 TI - c-Myb-binding sites mediate G(1)/S-associated repression of the plasma membrane Ca(2+)-ATPase-1 promoter. AB - We demonstrate that two Myb-binding sites of the mouse plasma membrane Ca(2+) ATPase-1 (PMCA1) promoter are required for G(1)/S cell cycle stage-associated repression of PMCA1 promoter activity. Nuclear run-on experiments revealed G(1)/S associated repression of PMCA1 transcription. Ribonuclease protection assays revealed two transcription initiation sites between two Myb-binding sites in the PMCA1 promoter. Gel shift assays showed that c-Myb can bind to wild-type but not point mutated Myb binding sequences of the PMCA1 promoter. Transient transfection assays using cell cycle-synchronized vascular smooth muscle cells (VSMC) and PMCA1 promoter-luciferase constructs showed a 2-fold decrease in reporter activity at G(1)/S as compared with G(0). Overexpression of wild-type c-Myb severely repressed PMCA1 promoter activity at both G(0) and G(1)/S while co transfection of a dominant negative c-Myb, or a construct encoding an anti-c-Myb neutralizing antibody, completely abolished the repression seen at G(1)/S. Single nucleotide substitutions in the first, second, or both Myb-binding sites alleviated the G(1)/S-associated repression of PMCA1 promoter activity in transformed rat VSMC and primary mouse VSMC cultures. We conclude that c-Myb mediates G(1)/S-associated transcriptional repression of the PMCA1 Ca(2+) pump in rodent VSMC by direct binding to the PMCA1 promoter. PMID- 10722758 TI - Hypertonic induction of aquaporin-5 expression through an ERK-dependent pathway. AB - Aquaporin-5 (AQP5) is a water channel protein expressed in lung, salivary gland, and lacrimal gland epithelia. Each of these sites may experience fluctuations in surface liquid osmolarity; however, osmotic regulation of AQP5 expression has not been reported. This study demonstrates that AQP5 is induced by hypertonic stress and that induction requires activation of extracellular signal-regulated kinase (ERK). Incubation of mouse lung epithelial cells (MLE-15) in hypertonic medium produced a dose-dependent increase in AQP5 expression; AQP5 protein peaked by 24 h and returned to baseline levels within hours of returning cells to isotonic medium. AQP5 induction was observed only with relatively impermeable solutes, suggesting an osmotic pressure gradient is required for induction. ERK was selectively activated in MLE-15 cells by hypertonic stress, and inhibition of ERK activation with two distinct mitogen-activated extracellular regulated kinase kinase (MEK) inhibitors, U0126 and PD98059, blocked AQP5 induction. AQP5 induction was also observed in the lung, salivary, and lacrimal glands of hyperosmolar rats, suggesting potential physiologic relevance for osmotic regulation of AQP5 expression. This report provides the first example of hypertonic induction of an extrarenal aquaporin, as well as the first association between mitogen-activated protein kinase signaling and aquaporin expression. PMID- 10722759 TI - Serine palmitoyltransferase regulates de novo ceramide generation during etoposide-induced apoptosis. AB - The de novo pathway of sphingolipid synthesis has been identified recently as a novel means of generating ceramide during apoptosis. Furthermore, it has been suggested that the activation of dihydroceramide synthase is responsible for increased ceramide production through this pathway. In this study, accumulation of ceramide mass in Molt-4 human leukemia cells by the chemotherapy agent etoposide was found to occur primarily due to activation of the de novo pathway. However, when the cells were labeled with a substrate for dihydroceramide synthase in the presence of etoposide, there was no corresponding increase in labeled ceramide. Further investigation using a labeled substrate for serine palmitoyltransferase, the rate-limiting enzyme in the pathway, resulted in an accumulation of label in ceramide upon etoposide treatment. This result suggests that the activation of serine palmitoyltransferase is the event responsible for increased ceramide generation during de novo synthesis initiated by etoposide. Importantly, the ceramide generated from de novo synthesis appears to have a distinct function from that induced by sphingomyelinase action in that it is not involved in caspase-induced poly (ADP-ribose)polymerase proteolysis but does play a role in disrupting membrane integrity in this model system. These results implicate serine palmitoyltransferase as the enzyme controlling de novo ceramide synthesis during apoptosis and begin to define a unique function of ceramide generated from this pathway. PMID- 10722760 TI - Leukotriene receptor antagonists: useful in acute asthma? PMID- 10722761 TI - Stopping smoking: the importance of nicotine addiction. PMID- 10722762 TI - Orthostatic increase of respiratory gas exchange in hyperventilation syndrome. PMID- 10722763 TI - Comparison of the effects of intravenous and oral montelukast on airway function: a double blind, placebo controlled, three period, crossover study in asthmatic patients. AB - BACKGROUND: Montelukast, a leukotriene receptor antagonist, improves parameters of asthma control including forced expiratory volume in one second (FEV(1)) when given orally to patients aged six years or older. This study was undertaken to compare the effect on FEV(1) of intravenous and oral montelukast and placebo during the 24 hour period following administration. METHODS: Fifty one asthmatic patients (FEV(1) 40-80% predicted and > or =15% improvement after inhaled beta agonist) were enrolled in a double blind, single dose, three period, crossover study to receive intravenous montelukast (7 mg), oral montelukast (10 mg), or placebo in a randomised fashion. The primary end point was area under the curve (AUC)(0-24 h) of the percentage change from baseline in FEV(1). Additional end points were maximum percentage change in FEV(1) and percentage change at different time points. RESULTS: Compared with placebo, intravenous and oral montelukast significantly increased the AUC(0-24 h) (means of 20.70%, 15.72%, and 7.75% for intravenous, oral and placebo, respectively; no statistical difference between intravenous and oral). The difference in least square means from placebo for intravenous montelukast was 13.27% (95% CI 7.07 to 19.46), p<0.001 and for oral montelukast was 7.44% (95% CI 1.20 to 13.68), p = 0.020. The maximum percentage change in FEV(1) was not significantly different for intravenous and oral montelukast (difference in least square means 6.78% (95% CI -0.59 to 14.15), p = 0.071). The mean percentage change in FEV(1) for intravenous montelukast was greater than for oral montelukast within the first hour (15.02% vs 4.67% at 15 min, p< or =0.001; 18.43% vs 12.90% at one hour, p<0.001 for intravenous and oral montelukast, respectively (placebo 3.05% at 15 minutes, 7.33% at one hour). Intravenous and oral montelukast were similar to placebo in the frequency of adverse events. CONCLUSIONS: The onset of action for intravenous montelukast was faster than for oral montelukast and the improvement in airway function lasted over the 24 hour observation period for both treatments. Although not well understood, there was a trend toward a greater improvement in FEV(1) with intravenous than with oral montelukast. These findings suggest that leukotriene receptor antagonists should be investigated as a treatment for acute severe asthma. PMID- 10722764 TI - Frequent paracetamol use and asthma in adults. AB - BACKGROUND: The pulmonary antioxidant glutathione may limit airway inflammation in asthma. Since paracetamol (acetaminophen) depletes the lung of glutathione in animals, a study was undertaken to investigate whether frequent use in humans was associated with asthma. METHODS: Information was collected on the use of analgesics as part of a population based case-control study of dietary antioxidants and asthma in adults aged 16-49 years registered with 40 general practices in Greenwich, South London. The frequency of use of paracetamol and aspirin was compared in 664 individuals with asthma and in 910 without asthma. Asthma was defined by positive responses to questions about asthma attacks, asthma medication, or waking at night with shortness of breath. The association between analgesic use and severity of disease amongst asthma cases, as measured by a quality of life score, was also examined. RESULTS: Paracetamol use was positively associated with asthma. After controlling for potential confounding factors the odds ratio for asthma, compared with never users, was 1.06 (95% CI 0.77 to 1.45) in infrequent users (/= 150 degrees ) consisted of 8 digits treated with dermatofasciectomy and full-thickness skin graft, and group C (TAM >/= 150 degrees ) consisted of 13 digits treated with fasciectomy and local flaps. Total active range of motion reflecting the preoperative, immediately postoperative, and final follow-up values revealed that group C (fasciectomy and local flap) was the only group to maintain a statistically significant TAM improvement from preoperative (205 degrees ) to final follow-up (230 degrees ) analysis. Dermatofasciectomy and full-thickness skin grafting did not prevent recurrent contracture (preoperative TAM = 175 degrees; final follow-up TAM = 150 degrees ). Thirteen patients had abnormal Semmes-Weinstein monofilament testing and 8 had abnormal 2-point discrimination. There were 3 anesthetic digits. Despite these findings, 18 of the 19 patients were unconditionally satisfied with their experience and would undergo the procedure again. PMID- 10722822 TI - Long-term results of volar ligament reconstruction for symptomatic basal joint laxity. AB - It has been hypothesized that instability of the thumb trapeziometacarpal joint is a major factor in the etiology of degenerative disease. Theoretically, surgically stabilized joints should be subject to less shear force and, hence, will be less likely to develop degenerative changes. The long-term results of volar ligament reconstruction were assessed in 19 patients (24 thumbs). The average age at surgery was 33 years (range, 18-55 years). Twenty-three thumbs were radiographic stage I; a preoperative x-ray was not available in 1. The follow-up period averaged 15 years (range, 10-23 years). At the final follow-up visit 15 thumbs were stage I, 7 were stage II, and 2 were stage III. Fifteen patients were at least 90% satisfied with the results of the surgery. Only 8% of thumbs advanced to radiographic arthritic disease, which compares favorably with the 17% to 33% reported incidence of stage III/IV basal joint arthritis in the general population. PMID- 10722823 TI - Kinematics of the proximal interphalangeal joint of the finger after surface replacement. AB - Nine fresh-frozen normal human cadaveric long fingers were used to compare the kinematics of the proximal interphalangeal joint (PIP) before and after a resurfacing metal-polyethylene prosthetic replacement (Avanta prosthesis, San Diego, CA) using the magnetic Isotrak system (Polhemus Navigational Systems, Colchester, VT). The kinematics of the PIP joint after replacement were similar to that of the normal joint. The maximum angular displacement was 5 degrees for lateral deviation and 9 degrees for rotation during the passive flexion and extension motion. The center of rotation after implant insertion was nearly identical to the center of rotation of the normal joint. This anatomically designed PIP prosthesis has potential to restore normal motion to the finger PIP joint while resisting physiologic out-of-plane forces such as pinch and grasp. PMID- 10722824 TI - The effects of transection and reconstruction of the ulnar collateral ligament complex on the position of the proximal phalanx of the thumb during simulated tip pinch. AB - Injuries to the ulnar collateral ligament (UCL) of the metacarpophalangeal joint of the thumb are common and may result in functional instability of the joint. Eight cadaveric hands were studied. Physiologic levels of muscle loads were applied to the extrinsic flexor tendon of the thumb to simulate tip pinch of the thumb. We investigated the effects of transection of the UCL and accessory UCL (UCL complex) with and without transection of the dorsal capsule and volar plate and of reconstruction of the UCL, for 2 surgical techniques, on the position of the proximal phalanx with respect to the thumb metacarpal. The spatial positions of the metacarpal and proximal phalanx were measured with a 6 degrees of freedom digitizing system for flexion angles from 0 degrees to 60 degrees in 15 degrees increments. Transection of the UCL complex, dorsal capsule, and volar plate (ulnar capsuloligamentous structures) of the metacarpophalangeal joint did not affect radioulnar deviation or radioulnar shift, but did produce significant increases in supination by 8 degrees and volar translation by 2 mm at 45 degrees and 60 degrees compared with those found for the intact joint. The UCL was reconstructed with a tendon graft using the autogenous extensor digiti quinti. The first surgical technique, a traditional technique, and the second surgical technique, a technique based on anatomy, returned the position of the proximal phalanx on the metacarpal head to normal, with the exceptions of volar translation of the proximal phalanx at 60 degrees and trends toward abnormal supination of the proximal phalanx for flexion angels of 45 degrees and 60 degrees. PMID- 10722825 TI - Correction of lunate malalignment when bone grafting scaphoid nonunion with humpback deformity: rationale and results of a technique revisited. AB - Patients with scaphoid nonunion with humpback deformity and collapse of the wrist were treated with palmar wedge bone grafting combined with reduction of the lunate to correct the dorsal intercalated segment instability deformity. Union was obtained at an average of 3 months, and patient satisfaction with functional outcome and pain relief was high. Palmar wedge bone grafting combined with correction of lunate malalignment successfully achieved scaphoid union, restored scaphoid length, and avoided the potential complication of scaphoid malunion. This report revisits a technique that facilitates accurate correction of lunate malalignment (dorsal intercalated segment instability) by initial reduction and pin fixation to the radius before insertion of scaphoid bone graft and internal fixation. PMID- 10722826 TI - Responsiveness of the short form-36, disability of the arm, shoulder, and hand questionnaire, patient-rated wrist evaluation, and physical impairment measurements in evaluating recovery after a distal radius fracture. AB - We evaluated the responsiveness of patient questionnaires and physical testing in the assessment of recovery after distal radius fracture. Patients (n = 59) were assessed at their baseline clinic visit and again 3 and 6 months after injury. At each visit patients completed a short form-36, Disability of the Arm, Shoulder, and Hand questionnaire, and patient-rated wrist evaluation (PRWE). At 3 and 6 months grip strength, range of motion, and dexterity were analyzed. Standardized response means (SRM) and effects sizes were calculated to indicate responsiveness. The PRWE was the most responsive. Both the PRWE (SRM = 2.27) and the Disability of the Arm, Shoulder, and Hand (SRM = 2.01) questionnaire were more responsive than the short form-36 (SRM = 0.92). The physical component summary score of the short form-36 was similar to that of the physical component subscales. Questionnaires were highly responsive during the 0- to 3-month time period when physical testing could not be performed. Of the physical tests, grip strength was most responsive, followed by range of motion. Responsive patient rating scales and physical performance evaluations can assist with outcome evaluation of patients with distal radius fracture. PMID- 10722827 TI - The role of the distal radioulnar ligaments, interosseous membrane, and joint capsule in distal radioulnar joint stability. AB - The individual contribution of the distal radioulnar ligaments to dorsal and palmar translational stability during forearm rotation remains controversial. Furthermore, the role of the distal radioulnar joint capsule as a restraint and contributor to stability has not been investigated. A biomechanical study was performed in 11 fresh cadaver specimens to simultaneously measure dorsal and palmar radioulnar ligament tension. Joint rotation and radial translation were measured after sequential excision of the disk, interosseous membrane, joint capsule, and radioulnar ligaments. Results confirmed that the dorsal ligament tightens during pronation while the palmar ligament becomes progressively lax; the converse occurred during supination. Translational stability remained intact at all positions throughout the sectioning sequence until one of the radioulnar ligaments was sectioned. The most significant increases in translation occurred after sectioning the dorsal radioulnar ligament in pronation and after sectioning the palmar radioulnar ligament in supination. Forearm rotation increased significantly after excising either hemicapsule. PMID- 10722828 TI - The importance of the pronated grip x-ray view in evaluating ulnar variance. AB - Although dynamic increases in ulnar variance may accompany functional activity, radiographic assessment of ulnar variance traditionally has used a neutral rotation x-ray of the wrist that provides an image of the radioulnar length with the wrist unloaded. Such a view may underestimate variance in wrists in which power grip and pronation result in significant proximal migration of the radius. This study investigates the effect of a maximum grip effort in combination with forearm pronation on ulnar variance in 22 patients who presented with ulnar wrist pain. The pronated grip x-ray view resulted in statistically significant increases in ulnar variance. Preoperative ulnar variance should be measured using both neutral rotation and pronated grip x-rays before selecting treatment for causes of ulnar wrist pain that are affected by radioulnar length so that dynamic increases in ulnar variance are considered when operative treatment is necessary. PMID- 10722829 TI - Acute compartment syndrome of the forearm in association with ulnar shortening osteotomy: a case report. AB - A 41-year-old man experienced severe pain in the forearm after undergoing ulnar shortening osteotomy to treat positive ulnar variance, a complication of a fracture of the distal end of the radius. The patient had compartment syndrome with compartment pressure of 55 mm Hg. A decompressive fasciotomy of the volar compartment provided total relief of pain and, subsequently, full recovery of all functions. We report the case and discuss the serious nature of compartment syndrome, its associated complications, and methods of diagnosis and management. PMID- 10722830 TI - Ulnar nerve function following total elbow arthroplasty: a prospective study comparing preoperative and postoperative clinical and electrophysiologic evaluation in patients with rheumatoid arthritis. AB - A study was conducted to determine the incidence of ulnar and peripheral neuropathy in patients with rheumatoid arthritis undergoing total elbow arthroplasty and the effect it has on ulnar nerve function after surgery. Preoperative and postoperative clinical and electrodiagnostic examinations were completed in 10 patients. Before surgery 4 patients had clinical and electrophysiologic evidence of a neuropathy (2 each with a peripheral neuropathy and an ulnar neuropathy). One patient had subclinical evidence of a chronic T-1 radiculopathy. After surgery 2 patients showed neurologic improvement (1 had ulnar neuropathy and 1 had diabetic neuropathy). One patient who had normal test results before surgery developed transient ulnar sensory symptoms after surgery. An electrodiagnostic study confirmed an ulnar neuropathy that was not detected on physical examination; the electrodiagnostic findings improved 4 months later. We found that a large percentage of patients (40%) with rheumatoid arthritis had evidence of ulnar or peripheral neuropathy before surgery. The presence of an ulnar or peripheral neuropathy did not predispose patients to develop postoperative ulnar nerve dysfunction either clinically or electrophysiologically. Preoperative and postoperative physical and electrodiagnostic examination results correlated in 9 of the 10 patients. PMID- 10722831 TI - Intraosseous neurilemmoma of the radius: a case report. AB - Neurilemmomas are benign tumors that arise from Schwann cells. They are rarely found on bone. We describe a neurilemmoma in a 45-year-old patient that affected the distal metaphysis of the radius. Only 1 previous case has been described in the literature. We discuss the clinical presentation, the radiographic aspect, particularly the magnetic resonance imaging characteristics, and the microscopic findings. PMID- 10722832 TI - Dynamic axial carpal instability. PMID- 10722833 TI - Dynamic axial carpal instability PMID- 10722834 TI - Development of a novel biomarker of free radical damage in reperfusion injury after cardiac arrest. AB - In a porcine model of cardiopulmonary resuscitation (CPR), we investigated changes in the plasma levels of 8-iso-PGF(2alpha), a marker for oxidative injury, and 15-keto-dihydro-PGF(2alpha), an inflammatory response indicator during the post-resuscitation period after cardiac arrest. Twelve piglets were subjected to either 2 or 5 min (VF2 and VF5 group) of ventricular fibrillation (VF) followed by 5 min of closed-chest CPR. Six piglets without cardiac arrest were used as controls. In VF5 group, 8-iso-PGF(2alpha) in the jugular bulb plasma (draining the brain) increased four-fold. Jugular bulb 8-iso-PGF(2alpha) in the control group remained unchanged. The 15-keto-dihydro-PGF(2alpha) also increased four fold in the VF5 group. Thus, 8-iso-PGF(2alpha) and 15-keto-dihydro-PGF(2alpha) measurements in jugular bulb plasma may be used as biomarkers for quantification of free radical catalyzed oxidative brain injury and inflammatory response in reperfusion injury. PMID- 10722835 TI - The symbiotically essential cbb(3)-type oxidase of Bradyrhizobium japonicum is a proton pump. AB - Purified cbb(3)-type oxidase of Bradyrhizobium japonicum was reconstituted into phospholipid vesicles. Tight vesicles were obtained as shown by the disturbance of deltapH with CCCP and the membrane potential with valinomycin, which led to a six-fold increase in cytochrome c oxidase activity. The vesicles were thus suitable for proton translocation experiments. In the presence of valinomycin, a pulse with reduced cytochrome c caused an acidification with a subsequent alkalinization, whereas the same pulse caused only an alkalinization in the presence of valinomycin plus CCCP. We conclude that the cbb(3)-type oxidase of B. japonicum is a proton pump. PMID- 10722836 TI - Rapid internal dynamics of BPTI is insensitive to pressure. (15)N spin relaxation at 2 kbar. AB - Pressure effects on the backbone dynamics of a native basic pancreatic trypsin inhibitor (BPTI) have been measured by (15)N spin relaxation and chemical shifts at 30 and 2000 bar. The experiments utilized the on-line variable pressure cell nuclear magnetic resonance system on (15)N-uniformly labeled BPTI at a proton frequency of 750.13 MHz at 36 degrees C. Longitudinal (R(1)) and transverse (R(2)) (15)N relaxation times and ((1)H)-(15)N nuclear Overhauser effects were measured for 41 protonated backbone nitrogens at both pressures. The model free analysis of the internal dynamics gave order parameters for individual H-N vectors at both pressures. The results indicate that rapid internal dynamics in the ps-ns range for the polypeptide backbone is not significantly affected by pressure in the range between 30 bar and 2 kbar. The result is consistent with the linear pressure dependence of (1)H and (15)N chemical shifts of BPTI, which suggests that local compressibilities and amplitudes of associated conformational fluctuation are nearly invariant in the same pressure range. Overall, we conclude that at 2 kbar BPTI remains within the same native ensemble as at 1 bar, with a small shift of population from that at 1 bar. PMID- 10722837 TI - Termination of IL-6-induced STAT activation is independent of receptor internalization but requires de novo protein synthesis. AB - The interleukin-6 (IL-6) receptor complex comprises the IL-6 receptor (IL-6R, gp80) and the signal transducer gp130. Binding of IL-6 to its receptor results in dimerization of gp130, activation of the Jak/STAT pathway, and in a down regulation of IL-6 binding sites by endocytosis. The STAT activation after stimulation is transient, being maximal after 15-30 min and disappearing after 60 90 min. The mechanism which leads to the termination of the signal is still unknown. In this paper we have studied whether the down-modulation of the STAT signal requires the endocytosis of the receptor complex. Our results suggest that the desensitization of the IL-6 signal is not due to internalization of the receptor complex but requires de novo protein synthesis. PMID- 10722838 TI - Homocysteine accelerates endothelial cell senescence. AB - In this study we demonstrate that exposure of cultured endothelial cells to homocysteine significantly accelerates the rate of endothelial senescence. Examination of telomere length demonstrates that homocysteine increases the amount of telomere length lost per population doubling. The effects of homocysteine on both senescence and telomere length are inhibited by treatment with the peroxide scavenger catalase. Chronic exposure of endothelial cells to homocysteine also increases the expression of two surface molecules linked to vascular disease, intracellular adhesion molecule-1 (ICAM-1) and plasminogen activator inhibitor-1 (PAI-1). Interestingly, the level of expression of both ICAM-1 and PAI-1 correlates with the degree of endothelial senescence. Taken together, these results suggest that homocysteine accelerates the rate of cellular senescence through a redox-dependent pathway. In addition, it suggests that chronic oxidative stress in the vessel wall may hasten the rate of senescence and that the senescent endothelial cell may in turn be pro atherogenic. PMID- 10722839 TI - Regulation of the golgi structure by the alpha subunits of heterotrimeric G proteins. AB - Disassembly of the Golgi apparatus is elicited by the action of nordihydroguaiaretic acid (NDGA) and this disassembly is prevented by the activation of heterotrimeric G proteins. In the present study we showed that overexpression of Galpha(z) or Galpha(i2) significantly suppresses the disassembly of the Golgi apparatus induced by NDGA. Overexpression of Gbeta(1)gamma(2), on the other hand, had no effect on NDGA-induced Golgi disassembly. Galpha(z) neither blocked Golgi disassembly induced by brefeldin A or nocodazole, nor interfered with protein transport, suggesting its specificity on the action of NDGA. Our results suggest that the alpha subunits of heterotrimeric G proteins are responsible for the maintenance of the Golgi structure. PMID- 10722840 TI - MFH-1 is required for bone morphogenetic protein-2-induced osteoblastic differentiation of C2C12 myoblasts. AB - Mesenchyme forkhead-1 (MFH-1), a winged helix/forkhead transcription factor, is expressed in developing cartilaginous tissues, kidney and arch arteries, and is essential for the normal development of the axial skeleton and aortic arch formation of mice. To investigate the possible role of MFH-1 in osteogenesis and osteoblast differentiation, we examined expression of MFH-1 induced by bone morphogenetic protein-2 (BMP-2) in C2C12 myoblasts, and found that MFH-1 protein and also MFH-1 mRNA increased markedly in C2C12 cells after treatment with BMP-2. To confirm the hypothesis that BMP-2 induced osteoblastic differentiation of C2C12 cells by increasing MFH-1 expression, we lowered the endogenous MFH-1 level by stably transfecting C2C12 cells with antisense MFH-1 sequence, and found that in antisense MFH-1 cell lines, both alkaline phosphatase (ALP) activity and production of osteocalcin induced by BMP-2 decreased markedly in comparison with control cell lines. Our results suggest that the BMP-2-induced MFH-1 protein may play a key role in regulating the commitment to osteoblastic differentiation of C2C12 myoblasts and production of osteoblast markers including ALP and osteocalcin. PMID- 10722841 TI - Induction of cell death by the lysosomotropic detergent MSDH. AB - Controlled lysosomal rupture was initiated in lysosome-rich, macrophage-like cells by the synthetic lysosomotropic detergent, O-methyl-serine dodecylamide hydrochloride (MSDH). When MSDH was applied at low concentrations, resulting in partial lysosomal rupture, activation of pro-caspase-3-like proteases and apoptosis followed after some hours. Early during apoptosis, but clearly secondary to lysosomal destabilization, the mitochondrial transmembrane potential declined. At high concentrations, MSDH caused extensive lysosomal rupture and necrosis. It is suggested that lysosomal proteases, if released to the cytosol, may cause apoptosis directly by pro-caspase activation and/or indirectly by mitochondrial attack with ensuing discharge of pro-apoptotic factors. PMID- 10722842 TI - Domain 1 of the urokinase receptor (uPAR) is required for uPAR-mediated cell binding to vitronectin. AB - In the present paper we have analyzed uPAR-mediated cellular binding to vitronectin using the murine erythroid progenitor cell line 32D. We show that expression of uPAR in 32D cells promotes cellular binding to vitronectin, but fails to support cell spreading. The strength of binding is correlated to the expression level of uPAR and is strongly stimulated by the presence of uPAR ligands. Using a truncated variant of uPAR lacking domain 1 and by antibody inhibition experiments, we demonstrate that domain 1 plays a crucial role in uPAR mediated cellular binding. The failure of the mutant uPAR to promote cellular binding is paralleled by a strong reduction in the affinity for vitronectin in vitro. PMID- 10722843 TI - The salicylate metabolite gentisic acid, but not the parent drug, inhibits glucose autoxidation-mediated atherogenic modification of low density lipoprotein. AB - Oxidation of low density lipoprotein (LDL) by glucose-derived radicals may play a role in the aetiology of atherosclerosis in diabetes. Salicylate was shown to scavenge certain radicals. In the present study, aspirin, salicylate and its metabolites 2,5- and 2, 3-dihydroxybenzoic acid (DHBA) were tested for their ability to impair LDL oxidation by glucose. Only the DHBA derivatives, when present during LDL modification, inhibited LDL oxidation and the increase in endothelial tissue factor synthesis induced by glucose oxidised LDL. The LDL glycation reaction was not affected by DHBA. The antioxidative action of DHBA may be attributed to free radical scavenging and/or chelation of transition metal ions catalysing glucose autoxidation. PMID- 10722845 TI - Preparation of synthetic human islet amyloid polypeptide (IAPP) in a stable conformation to enable study of conversion to amyloid-like fibrils. AB - Human synthetic islet amyloid polypeptide (hIAPP) is rapidly converted to beta sheet conformation and fibrils in aqueous media. Optimal solubility conditions for hIAPP were determined by circular dichroism spectroscopy and transmission electron microscopy. hIAPP in trifluoroethanol or hexafluoro-2-isopropanol (HFIP) diluted in water or phosphate buffer (PB) exhibited random structure which was converted to beta-sheet and fibrils with time. hIAPP, solubilised in HFIP, filtered and lyophilised remained in stable random structure for up to 7 days in water; in PB, insoluble aggregates precipitated from which protofilaments and fibrils formed with time. This suggests that amorphous aggregates of hIAPP could initiate islet amyloidosis in vivo. PMID- 10722844 TI - Characterization of the interactions between the glycine transporters GLYT1 and GLYT2 and the SNARE protein syntaxin 1A. AB - In this study we have examined the effect of the SNARE protein syntaxin 1A on the glycine transporters GLYT1 and GLYT2. Our results demonstrate a functional and physical interaction between both glycine transporters and syntaxin 1A. Co transfection of syntaxin 1A with GLYT1 or GLYT2 in COS cells resulted in approximately 40% inhibition in glycine transport. This inhibition was reversed by the syntaxin 1A-binding protein, Munc18. Furthermore, immunoprecipitation studies showed a physical interaction between syntaxin 1A and both transporters in COS cells and in rat brain tissue. Finally, we conclude that this physical interaction resulted in a partial removal of the glycine transporters from the plasma membrane as demonstrated by biotinylation studies. PMID- 10722846 TI - Rab37 is a novel mast cell specific GTPase localized to secretory granules. AB - GTPases regulate a myriad of cellular functions including signal transduction, cytoskeletal organization and membrane trafficking. Rab GTPases act to coordinate the membrane dynamics of cells by organizing and regulating the activity of effector proteins important in vesicle trafficking. Rab37 is a novel Rab GTPase specifically expressed in the MC-9 mast cell line and bone marrow mast cells. Rab37 is 74% identical to Rab26 and 47% identical to Rab8, a GTPase important in Golgi to plasma membrane vesicle trafficking in mammalian cells. When green fluorescent protein tagged Rab37 is expressed in bone marrow mast cells, the secretory granules are labeled. These data suggest that Rab37 may play an important role in mast cell degranulation making this protein a potentially important target for therapeutic intervention in the treatment of allergy. PMID- 10722847 TI - Elucidation of determinants of protein stability through genome sequence analysis. AB - Sequences of putative soluble proteins from complete genomes of eight thermophiles and 12 mesophiles were analyzed to gain insight into determinants of protein thermostability. The predator algorithm was used to assign secondary structures to each protein sequence. Based on simple statistical tests, a set of stabilizing factors was identified. These include reduced protein size, increases in number of residues involved in hydrogen bonding, beta-strand content and helix stabilization through ion pairs. There are also significant increases in the relative amounts of charged and hydrophobic beta-branched amino acids and decreases in uncharged polar amino acids in proteins from thermophiles relative to mesophilic organisms. Factors such as the relative proportion of residues in loops, proline and glycine content and helix capping do not appear to be important. PMID- 10722849 TI - Profile of gene expression regulated by induced p53: connection to the TGF-beta family. AB - The transcription regulatory function of p53 was analyzed by using two inducible p53 systems in the human lung cancer cell line H1299. cDNA probes derived from RNA harvested 12 h after p53 induction were used to probe filters containing cDNA arrays. Over 20 genes were found to be significantly induced or suppressed by p53. The induced genes can be classified mainly as cell cycle inhibitors like p21waf, GADD45, apoptosis-related genes like Fas/APO1 and PIG3 or DNA repair genes like DDB2, DNA ligase and G/T mismatch DNA glycosylase. The suppressed genes include mainly cell cycle regulators like cyclin B1, cyclin H and kinases like c-abl, CLK1 and others. The most notable induced gene was MIC-1, encoding a TGF-beta-related secretory protein, suggesting a potential paracrine component for p53 growth suppression. PMID- 10722848 TI - A novel C-terminal kinesin subfamily may be involved in chromosomal movement in caenorhabditis elegans. AB - C-terminal kinesin motor proteins, such as the Drosophila NCD and yeast KAR3, are involved in chromosomal segregation. Previously we have described two orthologs of NCD in Caenorhabditis elegans, KLP-3 and KLP-17, which also participate in chromosome movement. Here we report cDNA cloning of klp-15 and klp-16, and the expression pattern of the genes encoding C-terminal motor kinesins including klp 15 and klp-16. Interestingly KLP-15 and KLP-16 form a unique class of C-terminal kinesins, distinct from the previously known C-terminal motors in other organisms. Using in situ hybridization and RNA interference assay, we show that although all of these motors mediate chromosome segregation, they do so in a combination of unique and overlapping manners, suggesting a complex hierarchy of kinesin motor function in metazoans. PMID- 10722850 TI - Biochemical characterization and subcellular localization of the sterol C-24(28) reductase, erg4p, from the yeast saccharomyces cerevisiae. AB - The yeast ERG4 gene encodes sterol C-24(28) reductase which catalyzes the final step in the biosynthesis of ergosterol. Deletion of ERG4 resulted in a complete lack of ergosterol and accumulation of the precursor ergosta-5,7,22,24(28) tetraen-3beta-ol. An erg4 mutant strain exhibited pleiotropic defects such as hypersensitivity to divalent cations and a number of drugs such as cycloheximide, miconazole, 4-nitroquinoline, fluconazole, and sodium dodecyl sulfate. Similar to erg6 mutants, erg4 mutants are sensitive to the Golgi-destabilizing drug brefeldin A. Enzyme activity measurements with isolated subcellular fractions revealed that Erg4p is localized to the endoplasmic reticulum. This view was confirmed in vivo by fluorescence microscopy of a strain expressing a functional fusion of Erg4p to enhanced green fluorescent protein. We conclude that ergosterol biosynthesis is completed in the endoplasmic reticulum, and the final product is supplied from there to its membranous destinations. PMID- 10722851 TI - Effects of fatty acids, nucleotides and reactive oxygen species on durum wheat mitochondria. AB - Linoleic acid (LA) and other fatty acids added to respiring durum wheat mitochondria (DWM) were found to cause a remarkable membrane potential (deltaPsi) decrease, as monitored by measuring safranin fluorescence. The rate of deltaPsi decrease showed (i) saturation dependence on LA concentration; (ii) fatty acid specificity; (iii) inhibition by externally added ATP, GDP, GTP and Mg(2+) and (iv) sigmoid dependence upon initial DeltaPsi, thus suggesting the existence of an active plant mitochondrial uncoupling protein (PUMP) in mitochondria from monocotyledonous species (durum wheat, Triticum durum Desf.). Surprisingly, the rate of the linoleate dependent DeltaPsi decrease was found to be activated by reactive oxygen species (ROS) (hydrogen peroxide and superoxide anion) and, moreover, linoleate proved to lower the mitochondrial generation of superoxide anion. These results suggest that ROS can activate PUMP, thus protecting the cell against mitochondrial ROS production. PMID- 10722852 TI - Tubulin folding cofactor D is a microtubule destabilizing protein. AB - A rapid switch between growth and shrinkage at microtubule ends is fundamental for many cellular processes. The main structural components of microtubules, the alphabeta-tubulin heterodimers, are generated through a complex folding process where GTP hydrolysis [Fontalba et al. (1993) J. Cell Sci. 106, 627-632] and a series of molecular chaperones are required [Sternlicht et al. (1993) Proc. Natl. Acad. Sci. USA 90, 9422-9426; Campo et al. (1994) FEBS Lett. 353, 162-166; Lewis et al. (1996) J. Cell Biol. 132, 1-4; Lewis et al. (1997) Trends Cell Biol. 7, 479-484; Tian et al. (1997) J. Cell Biol. 138, 821-823]. Although the participation of the cofactor proteins along the tubulin folding route has been well established in vitro, there is also evidence that these protein cofactors might contribute to diverse microtubule processes in vivo [Schwahn et al. (1998) Nature Genet. 19, 327-332; Hirata et al. (1998) EMBO J. 17, 658-666; Fanarraga et al. (1999) Cell Motil. Cytoskel. 43, 243-254]. Microtubule dynamics, crucial during mitosis, cellular motility and intracellular transport processes, are known to be regulated by at least four known microtubule-destabilizing proteins. OP18/Stathmin and XKCM1 are microtubule catastrophe-inducing factors operating through different mechanisms [Waters and Salmon (1996) Curr. Biol. 6, 361-363; McNally (1999) Curr. Biol. 9, R274-R276]. Here we show that the tubulin folding cofactor D, although it does not co-polymerize with microtubules either in vivo or in vitro, modulates microtubule dynamics by sequestering beta-tubulin from GTP bound alphabeta-heterodimers. PMID- 10722853 TI - Similarities in reflex control of laryngeal and cardiac vagal motor neurones. AB - We sought to test the hypothesis that laryngeal adductor and cardiac vagal motor neurones respond similarly to the activation of certain afferent inputs. Experiments were performed on a working heart-brainstem preparation of rat devoid of pulmonary stretch receptor feedback. Upper airway negative pressure receptors (UANPR), peripheral arterial chemoreceptors and receptors at the junction of the pharynx and oesophagus were stimulated selectively while recording heart rate, recurrent laryngeal, phrenic and hypoglossal motor outflows, subglottic pressure during constant translaryngeal airflow (as an index of laryngeal resistance), and single unit respiratory neurone activity. Stimulation of all three receptor types produced bradycardia, evoked discharges in the recurrent laryngeal and hypoglossal motor outflows during the post-inspiratory period and caused swallowing. Stimulation of pharyngoesophageal receptors and peripheral chemoreceptors evoked an increase in laryngeal resistance during the post inspiratory phase indicative of laryngeal adductor motoneurone activation. Although this reflex response cannot be evaluated during UANPR stimulation, some post-inspiratory neurones were powerfully activated suggesting that UANPR probably drive laryngeal adductor muscles. Our data show that motor outflows controlling cardiac rate and laryngeal patency are concurrently activated by these sensory inputs. This may constitute the basis for a stereotyped defensive reflex response which maintains end expiratory lung volume, thus conserving oxygen in conditions of upper airway obstruction. Our observations lend further support to models of cardiorespiratory control which propose close coupling and shared central mechanisms for the regulation of the cardiovascular and respiratory systems. PMID- 10722855 TI - Sleep influences on homeostatic functions: implications for sudden infant death syndrome. AB - The mechanisms underlying the sudden infant death syndrome (SIDS) appear to have origins in the fetal environment resulting in neural damage which later compromises responses to breathing or blood pressure challenges during sleep. The deficits appear to involve alterations in neurotransmitter receptors within regions involved in chemoreception and cardiovascular control. SIDS risk is enhanced by pre- and postnatal nicotine exposure, and possibly by hypoxic experiences. The prone sleeping position plays a significant role in risk, as do head positions that minimize facial escape from enclosed spaces; elevated body temperature may also be a factor. Compensatory mechanisms, including diminished gasping ability, relative failure to arouse to a safer state, or a failure to recruit respiratory efforts to overcome a blood pressure loss have been the object of recent research efforts. The findings suggest that the fatal event involves a neurally-compromised infant, circumstances that challenge vital physiology, most likely during sleep, at a particular developmental period. PMID- 10722854 TI - Impact of brainstem sleep mechanisms on pharyngeal motor control. AB - Suppression of respiratory muscle activity in sleep, particularly evident in the pharyngeal muscles, is pivotal to the pathogenesis of common sleep-related breathing disorders such as obstructive sleep apnea. Obstructive apneas are caused by sleep-related decrements in pharyngeal muscle activity that leads to snoring and airway obstruction in individuals with underlying structural narrowing of the upper airway. Since obstructive apneas occur exclusively during sleep, this disorder by definition is state-dependent and ultimately caused by the influences of brainstem sleep mechanisms on pharyngeal motoneurons in individuals with compromised upper airway anatomy. This paper reviews the central neuronal mechanisms by which sleep reduces the output to the pharyngeal muscles and the neurotransmitters implicated in this alteration. The experimental approaches used to address this problem are also mentioned and their relative advantages and disadvantages discussed. In particular, the information derived from reduced animal preparations is reviewed and the need for studies in natural sleep is emphasised. Identifying the central neuronal mechanisms and neurotransmitters involved in sleep-related suppression of pharyngeal muscle activity not only has important basic relevance to understanding state-dependent respiratory control, it also has immediate clinical relevance to understanding common sleep-related breathing disorders at the central neuronal level. Determining these basic mechanisms also has immediate clinical relevance to understanding the pathogenesis of airway occlusions, and guiding neuro pharmacological approaches aimed at preventing the sleep-related decrements in pharyngeal muscle tone that are ultimately the root cause of obstructive sleep apnea. PMID- 10722856 TI - Significance of vagal innervation in perinatal breathing and gas exchange. AB - The mechanisms responsible for the establishment of continuous breathing at birth remain unknown. Several studies have shown that postnatal vagal denervation produces deleterious effects on ventilation as well as breathing patterns during the newborn period. However, the validity of these studies was compromised by anesthesia, tracheostomy or possible secondary laryngeal obstruction. We have recently developed an unanesthetized lamb model in which vagal denervation was performed antenatally and below the recurrent laryngeal nerves thereby avoiding anesthesia, tracheostomy and laryngeal paralysis. The denervated animals developed life-threatening respiratory failure shortly after birth, implying that vagal innervation of the lungs plays an essential role in establishing adequate gas exchange in the first hours after birth. We have subsequently investigated various mechanisms of respiratory failure in denervated animals. Our results show that the surfactant system dysfunction and loss of vagally mediated volume feedback likely contributed to the respiratory failure observed in the intrathoracically denervated animal model. PMID- 10722857 TI - Pathophysiology of childhood obstructive sleep apnea: current concepts. AB - The obstructive sleep apnea syndrome (OSAS) is a common and serious condition during childhood. Its pathophysiology remains poorly understood. Although OSAS is related to adenotonsillar hypertrophy in children, adenotonsillar hypertrophy is not likely the sole cause of sleep-disordered breathing in this age group. Rather, large tonsils and adenoids appear to precipitate OSAS in children with underlying abnormalities of upper airway function. Normal children have a relatively narrow upper airway, but maintain airway patency during sleep because of increased upper airway neuromotor tone and an increased central ventilatory drive. We speculate that OSAS occurs in those children lacking the compensatory upper airway neuromotor responses. PMID- 10722858 TI - Respiratory-related activation and mechanical effects of the pharyngeal constrictor muscles. AB - We have examined the respiratory-related activation of pharyngeal constrictor (PC) muscles in decerebrate cats, normal adult humans and patients with obstructive sleep apnea. In decerebrate cats and awake normal adult humans, phasic expiratory PC activity is uniformly present under hypercapnic and hypoxic conditions. While the PC muscles are electrically silent during quiet breathing in normal adult humans in NREM sleep, an activation pattern very similar to that of other upper airway dilators, such as the genioglossus muscle, is present during spontaneous and induced apneas in patients with obstructive sleep apnea. Experiments using an isolated, sealed upper airway preparation in decerebrate cat show that selective activation of the PC muscles stiffens the pharyngeal airway. The results also show that activation of the PC muscles constricts the airway at relatively high airway volumes but dilates the airway at relatively low airway volumes. These results suggest that PC muscle activation at the end of an apneic episode, when airway volume is relatively low, may help restore airway patency in patients with obstructive sleep apnea. PMID- 10722859 TI - Leptin, obesity, and respiratory function. AB - Leptin is a protein produced by adipose tissue that circulates to the brain and interacts with receptors in the hypothalamus to inhibit eating. The importance of this single peptide is vividly demonstrated by the profound obesity exhibited by the ob/ob mouse (C57BL/6J-Lep(ob)) which is unable to produce functional leptin. The measurement of respiratory function in the ob/ob mouse shows that the profound obesity is associated with impaired respiratory mechanics and depressed respiratory control, particularly during sleep. Longitudinal studies and leptin replacement studies in the ob/ob mouse indicate that leptin may act as both as a growth factor in the lung and as a neurohumoral modulator of central respiratory control mechanisms. Moreover, wildtype mice with diet-induced obesity have normal respiratory function associated with markedly elevated leptin levels. Human obesity, similar to obesity in wildtype mice, also causes an elevation in circulating leptin. However, unlike the tight relationship between obesity and elevated leptin present in an inbred strain of wildtype mice, human obesity is associated with more variable leptin levels for a given degree of adiposity. Thus, the possibility exists that a relative deficiency in leptin, or a leptin resistance, may play a role in obesity-related breathing disorders such as obesity hypoventilation syndrome (OHS) or obstructive sleep apnea (OSA). PMID- 10722860 TI - Effects of positive intrathoracic pressure on pulmonary and systemic hemodynamics. AB - The Frank-Starling Law accounts for many changes in cardiac performance previously attributed to changes in contractility in that changes in contractility might have been incorrectly inferred from changing ventricular function curves (i.e. systolic performance plotted against filling pressure) if diastolic compliance also changed. To apply the Frank-Starling Law in the presence of changing diastolic compliance, it is necessary to measure end diastolic volume directly or to calculate end-diastolic transmural pressure, which requires that pericardial pressure be known. Under most normal circumstances, increased intrathoracic pressure (and other interventions, such as vasodilators or lower-body negative pressure, that decrease central blood volume) decreases the transmural end-diastolic pressures of both ventricles, their end diastolic volumes and stroke work. However, when ventricular interaction is significant, the effects of these interventions might be quite different; this may be important in patients with heart-failure. Although these interventions decrease RV transmural pressure, they may increase LV transmural pressure, end diastolic volume, and thus stroke work by the Frank-Starling mechanism. PMID- 10722861 TI - Neural and humoral mechanisms mediating cardiovascular responses to obstructive sleep apnea. AB - Patients with obstructive sleep apnea are at increased risk for hypertension. The mechanisms underlying this increased risk are not known. During sleep, repetitive apneic episodes result in hypoxemia and carbon dioxide retention, which cause increases in sympathetic nerve activity and elicit humoral vasoconstrictor responses. While these mechanisms explain nocturnal elevations in blood pressure, it is unclear why hypertension and elevated sympathetic nerve activity prevail even during the daytime. This review will examine briefly some of the neural and humoral mechanisms that are activated by nocturnal apneas and which may contribute to persistent increases in blood pressure even during daytime normoxia. Disruption of the autonomic and hemodynamic profile of normal sleep by apneic events manifests as raised blood pressure and heightened sympathetic nerve traffic during sleep. During awake daytime normoxia, baroreflex and chemoreflex dysfunction may contribute to maintenance of higher blood pressure and sympathetic activity. Sustained vasoconstrictor effects of nocturnal endothelin release may also be implicated in the elevated daytime blood pressures. PMID- 10722862 TI - Effect of episodic hypoxia on sympathetic activity and blood pressure. AB - One of the major manifestations of obstructive sleep apnea (OSA) is profound and repeated (episodic) hypoxia during sleep. Acute hypoxia leads to stimulation of the peripheral chemoreceptors, which in turn directly increase sympathetic outflow. It is believed that this increase in sympathetic outflow is directly responsible, at least in part, for the acute blood pressure (BP) changes seen in OSA. It is difficult however, to study the chronic effects of repeated episodic hypoxia (EH) in humans since the chronic cardiovascular changes may take many years to manifest. For this reason, we developed a method of providing recurrent short periods of hypoxia (resembling the episodic desaturation in humans with OSA) to rats for 35 days, stimulating the chemoreceptors and the sympathetic nervous system, allowing examination of the chronic cardiovascular response to EH. The result of EH in rats is a 10-14 mmHg increase in resting (unstimulated) mean BP that lasts for several weeks after cessation of the daily EH. This BP increase is blocked by carotid body denervation, sympathetic nerve ablation, renal sympathectomy, adrenal medullectomy, and the angiotensin-1 receptor blocker losartan. Thus, it appears that adrenergic and renin-angiotensin system over activity contribute to the early chronic elevated BP in EH and perhaps in human hypertension associated with OSA. PMID- 10722863 TI - Important role of carotid chemoreceptor afferents in control of breathing of adult and neonatal mammals. AB - This review provides a summary and prospective on the importance of carotid/peripheral chemoreceptors to the control of breathing during physiologic conditions. For several days after carotid body denervation (CBD), adult mammals hypoventilate (+10 mmHg increase in Pa(CO(2))) at rest and during exercise and CO(2) sensitivity is attenuated by about 60%. In addition, if the rostral ventrolateral medulla is cooled during NREM sleep after CBD, a sustained apnea is observed. Eventually, days or weeks after CBD, a peripheral ventilatory chemoreflex redevelops and there is a normalization of breathing (rest and exercise) and CO(2) sensitivity. The site (s) of the regained chemosensitivity has not been established. This plasticity/redundancy after CBD appears greater in neonates than in adult mammals. These data suggest the carotid and other peripheral chemoreceptors provide an important excitatory input to medullary respiratory neurons that is essential for breathing when wakeful stimuli and central chemoreceptors are absent. PMID- 10722864 TI - Sleep of the great. AB - Both Lewis Carroll and William Shakespeare appear to have made clinical observations of sleep apnea syndromes long before they were discovered by medical science, and to have understood something about their physiological mechanisms. The somnolent dormouse in Alice in Wonderland indicates that his problem is one of sleep and breathing and is subject to modern treatment for obstructive apnea. Shakespeare in Henry IV presents a case of obstructive apnea along with a case of Cheyne-Stokes breathing and uses the plot of these history plays to explain by analogy the theoretical basis for periodic breathing. PMID- 10722865 TI - Soluble interleukin-6 receptor (sIL-6R) makes IL-6R negative T cell line respond to IL-6; it inhibits TNF production. AB - The receptor for interleukin-6 (IL-6) consists of two subunits: a ligand specific IL-6Ralpha and gp130 that is responsible for signal-transduction. A soluble form of the ligand specific chain was described that when complexed to IL-6 is capable of binding to the membrane-bound gp130 subunit and thus can elicit signal transduction. This soluble receptor can act on cells that express only the gp130 but not the ligand-specific subunit of the IL-6R. This phenomenon, called trans signaling, introduced a novel aspect of cytokine action. In this study we examined the response of Jurkat cells, that are known not to express IL-6Ralpha, to IL-6, the soluble IL-6 receptor (sIL-6R) and a covalent complex of IL-6 and sIL-6R termed Hyper-IL-6. We studied the expression of tumour necrosis factor (TNF) and interferon-gamma (IFN-gamma). The complex of IL-6+sIL-6R and Hyper-IL-6 inhibited significantly the production of TNF in a gp130-dependent manner, whereas no differences in IFN-gamma expression were found. IL-6 and sIL-6R alone were not effective. Because we did not detect major differences in the TNF mRNA levels upon treatments, we conclude that the inhibition of TNF production should occur at the post-transcriptional level. These results provide another example of trans-signaling and underline the physiological importance of sIL-6R, and in the case of Hyper-IL-6 its possible therapeutic application can also be considered. PMID- 10722866 TI - Differential regulation of nitric oxide production by increase of intracellular cAMP in murine primary fibroblasts and L929 fibrosarcoma cell line. AB - The effect of intracellular cAMP rise on nitric oxide (NO) production was compared in murine primary fibroblasts isolated from the spleens of CBA mice, and L929 fibrosarcoma cell line. Treatment of confluent L929 cells with cAMP analogues -dibutyryl-cAMP (db-cAMP) or 8-Cl-cAMP caused dose-dependent augmentation of inducible NO synthase (iNOS)-mediated NO production, which has been abrogated by inhibition of protein synthesis with cycloheximide or addition of selective iNOS inhibitor aminoguanidine. In contrast, under the same cultivating conditions, cAMP analogues were not able to upregulate NO synthesis in primary fibroblasts. Treatment with cAMP analogues or non-selective phosphodiesterase (PDE) inhibitor pentoxifylline affected IFNgamma-induced NO synthesis in both cell types, but in the opposite manner-enhancing in L929 cells and suppressive in primary fibroblasts. The induction of iNOS, but not its catalytic activity, was impaired in cAMP-treated primary fibroblasts. Finally, PDE type IV inhibitor rolipram enhanced IFN-gamma-triggered NO synthesis in L929 cells, but was unable to mimic cAMP analogue or PTX-mediated suppression of NO synthesis in spleen fibroblasts. These results suggest that, in contrast to L929 fibrosarcoma cell line, intracellular cAMP rise might have a role in downregulation of NO production in murine primary fibroblasts. PMID- 10722867 TI - Influences of amino acid sequences in FR1 region on binding activity of the scFv and Fab of an antibody to human gastric cancer cells. AB - Murine anti-gastric cancer mAb 3H11 has promising clinical applications. In this work, engineering of 3H11 scFv and Fab was conducted to increase its usefulness. 3H11 scFv and Fab were constructed by PCR amplification of the V-domains with degenerate primers for FR1. The bacterially expressed 3H11 Ab fragments showed no antigen binding activity. Then the N-terminal sequences of V regions were mutated to the 3H11 original sequence. The expressed scFv and Fab in bacterial culture supernatant showed binding activity to gastric cancer cells. Comparing the expression of unmutated and mutated 3H11 Fab, we found that the sequence changes of the V region N terminus introduced by PCR may seriously affect antigen binding but not the expression of antibody. Correction of either VL or VH N-terminal sequences can partially restore the antigen binding activity (61% for VL and 73% for VH). PMID- 10722868 TI - Expression and function of sialoadhesin in rat alveolar macrophages. AB - Alveolar macrophages (Amφ) represent an immunologically distinct sub population within the reticuloendothelial system. Phagocytosis and possibly antigen presentation by Amφ are essential components of specific and innate primary immune defence processes against inhaled material. The mφ-restricted sheep erythrocyte receptor sialoadhesin (Sn) is a member of the immunglobulin superfamily and binds specifically to sialic acid-containing structures such as selectins and was originally identified as the sheep erythrocyte receptor (SER) responsible for sialic acid-dependent binding of native sheep erythrocytes (SE) to resident murine bone marrow macrophages in rosetting assays. Sn expression has been demonstrated on murine and rat mφ in lymphatic organs and is recognised by the monoclonal antibody (mAb) ED3 in the rat. In addition, sialic acid dependent receptor (SAR) activities that mediate rosette formation of alveolar, peritoneal, splenic and bone marrow-resident rat mφ with SE pretreated with gangliosides and SER-like activities between native SE and trypsinised Amφ, have been described. The binding activities of both SAR and Sn show similar characteristics suggesting that these molecules are closely structurally related or identical. To clarify the relationship between Sn, SAR and SER-like activities, the binding of mAb ED3 to isolated rat Amφ was investigated by flow cytometry and rosetting assays. It is demonstrated that rat Amφ express Sn and evidence is provided that SAR and SER-like activities are mediated by Sn. PMID- 10722869 TI - Nitric oxide induced expression of stress proteins in virulent and avirulent promastigotes of Leishmania donovani. AB - Intracellular survival and replication of Leishmania donovani inside macrophage is essential for establishment of the disease. Cytokines play an important role in this process through activation or inhibition of macrophage antimicrobial activity. Nitric oxide (NO) has been demonstrated to be the principal effector molecule mediating intracellular killing of Leishmania. We have examined the effect of NO and various other cytokines on stress protein synthesis by promastigotes of L. donovani virulent and avirulent strains. Virulent promastigotes exposed to NO showed appreciable increase in relative synthesis of HSPs 83, 70 and 65. The overexpression of HSPs on exposure of parasite to NO was observed to be more pronounced at 37 degrees C than at 24 degrees C. In contrast, the avirulent promastigotes responded by an increase in relative synthesis of HSP70 alone at 37 degrees C. Furthermore, treatment of promastigotes of L. donovani with gammaIFN, TGF-beta or IL-4 did not significantly alter the stress proteins expression. The overexpression of HSPs in promastigotes of L. donovani in response to sublethal doses of NO suggests that HSPs may be playing a protective role for parasite survival in the mammalian host. This is further supported by the observation that a significantly higher induction of HSPs is seen in the virulent as compared to the avirulent strain of L. donovani. PMID- 10722870 TI - Limitation of nitric oxide production: cells from lymph node and spleen exhibit distinct difference in nitric oxide production. AB - Many types of cells in the immune system have been found to produce nitric oxide (NO), which performs multiplex functions. However, in myelin basic protein peptide 68-86 (MBP 68-86)-induced experimental autoimmune encephalomyelitis (EAE) in Lewis rats, we found that elevated NO production was generated from spleen cells (SC), not from lymph node cells (LNC). LNC expressed lower NO synthase 2 (NOS2) and produced lower levels of NO than SC upon MBP 68-86 stimulation. Expression of B7-1(CD80) and B7-2(CD86) molecules was much lower on LNC than on SC. Blocking of B7-1 or B7-2 ligation resulted in reduced NO production by SC. Unlike SC, LNC were resistant to interferon-gamma- or lipopolysaccharide-induced NO production. NO derived from SC suppressed cell proliferation and induced apoptosis in vitro. Addition of N(omega)-nitrol-L-arginine methylester (L-NAME) into cell cultures promoted cell expansion and reduced apoptosis. These results indicate that NO production originates from SC, but not from LNC. Low expression of co-stimulatory molecules and NOS2 of LNC limits NO induction. The high levels of NO derived from SC are involved in the self-limiting mechanisms of autoimmune responses by inhibiting cell expansion and promoting cell apoptosis. PMID- 10722871 TI - MRP8/MRP14, CD11b and HLA-DR expression of alveolar macrophages in pneumonia. AB - Activation of alveolar macrophages is characterised by specific alterations to the expression pattern of surface markers under certain pathological conditions. MRP8/MRP14 and CD11b are involved in the regulation of macrophage migration and adhesion. HLA-DR regulates the antigen presentation by alveolar macrophages. The aim of this study was to investigate the phenotype of alveolar macrophages in pneumonia particularly in relationship to the changes in concentrations of TGF beta1 and IL-8. Using cytofluorimetry, we analysed the surface expression of MRP8/MRP14, CD11b, and HLA-DR on alveolar macrophages of 42 pneumonia (PN) patients, 14 patients with interstitial lung diseases (ILD), five patients with chronic obstructive lung disease (COPD), and 58 patients without lung disease. Phenotypic characteristics were correlated to the concentration of TGF-beta1 and IL-8 in the bronchoalveolar lavage fluid (BALF) of the same patients. The direct influence of TGF-beta1 and IL-8 on expression of MRP8/MRP14, CD11b and HLA-DR of cultured monocytes and MonoMac cells was analysed. Significantly more MRP8/MRP14 and CD11b positive macrophages and less HLA-DR-positive macrophages were found in PN but not in ILD or COPD. The percentage of CD11b-positive macrophages correlated with the TGF-beta1 as well as the IL-8 concentrations. The amount of HLA-DR-positive macrophages correlated negatively to the concentration of TGF beta1 and IL-8. These findings document a significant activation of alveolar macrophages during pneumonia. TGF-beta1 led to a modulation of HLA-DR and MRP8/MRP14-antigen expression in vitro. In conclusion, it was shown that in pneumonia but not in ILD or COPD alveolar macrophages were characterised by an increased MRP8/MRP14 and CD11b expression and a diminished HLA-DR expression. The characterisation of subpopulations within the alveolar macrophages may be a useful tool for the monitoring of disease progression. PMID- 10722872 TI - State-of-the-art of clinical immunology in Europe. PMID- 10722873 TI - Myosins: a diverse superfamily. AB - Myosins constitute a large superfamily of actin-dependent molecular motors. Phylogenetic analysis currently places myosins into 15 classes. The conventional myosins which form filaments in muscle and non-muscle cells form class II. There has been extensive characterization of these myosins and much is known about their function. With the exception of class I and class V myosins, little is known about the structure, enzymatic properties, intracellular localization and physiology of most unconventional myosin classes. This review will focus on myosins from class IV, VI, VII, VIII, X, XI, XII, XIII, XIV and XV. In addition, the function of myosin II in non-muscle cells will also be discussed. PMID- 10722874 TI - Regulation of the enzymatic and motor activities of myosin I. AB - Myosins I were the first unconventional myosins to be purified and they remain the best characterized. They have been implicated in various motile processes, including organelle translocation, ion channel gating and cytoskeletal reorganization but their exact cellular functions are still unclear. All members of the myosin I family, from yeast to man, have three structural domains: a catalytic head domain that binds ATP and actin; a tail domain believed to be involved in targeting the myosins to specific subcellular locations and a junction or neck domain that connects them and interacts with light chains. In this review we discuss how each of these three domains contributes to the regulation of myosin I enzymatic activity, motor activity and subcellular localization. PMID- 10722875 TI - Class V myosins. PMID- 10722876 TI - Are class III and class IX myosins motorized signalling molecules? AB - Myosins exist that are fused to domains that harbour signalling activities. Class III myosins (NINAC) are protein kinases that play important roles in phototransduction. Class IX myosins inactivate the small G-protein Rho that acts as molecular switch. Because these myosins interact via their myosin head domain with actin filaments, they link signal transduction to the actin cytoskeleton. The exact motor properties of these myosins, however, remain to be determined. PMID- 10722878 TI - Distinct cytoplasmic dynein complexes are transported by different mechanisms in axons. AB - In neurons, cytoplasmic dynein is synthesized in the cell body, but its function is to move cargo from the axon back to the cell body. Dynein must therefore be delivered to the axon and its motor activity must be regulated during axonal transport. Cytoplasmic dynein is a large protein complex composed of a number of different subunits. The dynein heavy chains contain the motor domains and the intermediate chains are involved in binding the complex to cargo. Five different intermediate chain polypeptides, which are the result of the alternative splicing of the two intermediate chain genes, have been identified. We have characterized two distinct pools of dynein that are transported from the cell body along the axon by different mechanisms. One pool, which contains the ubiquitous intermediate chain, is associated with the membranous organelles transported by kinesin in the fast transport component. The other pool, which contains the other developmentally regulated intermediate chains, is transported in slow component b. The mechanism of dynein regulation will therefore depend on which pool of dynein is recruited to function as the retrograde motor. In addition, the properties of the large pool of dynein associated with actin in slow component b are consistent with the hypothesis that this dynein may be the motor for microtubule transport in the axon. PMID- 10722877 TI - The dynein microtubule motor. AB - Dyneins are large multi-component microtubule-based molecular motors involved in many fundamental cellular processes including vesicular transport, mitosis and ciliary/flagellar beating. In order to achieve useful work, these enzymes must contain motor, cargo-binding and regulatory components. The ATPase and microtubule motor domains are located within the very large dynein heavy chains that form the globular heads and stems of the complex. Cargo-binding activity involves the intermediate chains and several classes of light chain that associate in a subcomplex at the base of the soluble dynein particle. Regulatory control of dynein motor function is thought to involve the phosphorylation of various components as well as a series of light chain proteins that are directly associated with the heavy chains. These latter polypeptides have a variety of intriguing attributes, including redox-sensitive vicinal dithiols and Ca(2+) binding, suggesting that the activity of individual dyneins may be subject to multiple regulatory inputs. Recent molecular, genetic and structural studies have revealed insight into the roles played by these various components and the mechanisms of dynein-based motility. PMID- 10722879 TI - The role of cytoplasmic dynein in the human brain developmental disease lissencephaly. AB - Lissencephaly is a brain developmental disorder characterized by disorganization of the cortical regions resulting from defects in neuronal migration. Recent evidence has implicated the human LIS-1 gene in Miller-Dieker lissencephaly and isolated lissencephaly sequence. LIS-1 is homologous to the fungal genes NudF and PAC1, which are involved in cytoplasmic dynein mediated nuclear transport, but it is also almost identical to a subunit of PAF acetylhydrolase, an enzyme which inactivates the lipid mediator platelet activating factor. Recent evidence from our laboratory has revealed that cytoplasmic dynein coimmunoprecipitates with LIS 1 in bovine brain cytosol, supporting a role in the dynein pathway in vertebrates. Overexpression of LIS-1 interferes with cell division, with noteworthy effects on chromosome attachment to the mitotic spindle and on the interaction of astral microtubules with the cell cortex. Other aspects of dynein function, such as the organization of the Golgi apparatus, are not affected. Together, these results suggest a role for LIS-1 in cytoplasmic dynein functions involving microtubule plus-ends. Furthermore, they suggest that mutations in LIS 1 may produce a lissencephalic phenotype either by interfering with the movement of neuronal nuclei within extending processes, or by interference with the division cycle of neuronal progenitor cells in the ventricular and subventricular zones of the developing nervous system. PMID- 10722880 TI - Mitotic motors in Saccharomyces cerevisiae. AB - The budding yeast Saccharomyces cerevisiae provides a unique opportunity for study of the microtubule-based motor proteins that participate in mitotic spindle function. The genome of Saccharomyces encodes a relatively small and genetically tractable set of microtubule-based motor proteins. The single cytoplasmic dynein and five of the six kinesin-related proteins encoded have been implicated in mitotic spindle function. Each motor protein is unique in amino acid sequence. On account of functional overlap, no single motor is uniquely required for cell viability, however. The ability to create and analyze multiple mutants has allowed experimental dissection of the roles performed by each mitotic motor. Some of the motors operate within the nucleus to assemble and elongate the bipolar spindle (kinesin-related Cin8p, Kip1p, Kip3p and Kar3p). Others operate on the cytoplasmic microtubules to effect spindle and nuclear positioning within the cell (dynein and kinesin-related Kip2p, Kip3p and Kar3p). The six motors apparently contribute three fundamental activities to spindle function: motility, microtubule cross-linking and regulation of microtubule dynamics. PMID- 10722881 TI - Directional motility of kinesin motor proteins. AB - Kinesin motor proteins are molecules capable of moving along microtubules. They share homology in the so-called core motor domain which acts as a microtubule dependent ATPase. The surprising finding that different members of the superfamily move in opposite directions along microtubules despite their close similarity has stimulated intensive research on the determinants of motor directionality. This article reviews recent biophysical, biochemical, structural and mutagenic studies that contributed to the elucidation of the mechanisms that cause directional motion of kinesin motor proteins. PMID- 10722882 TI - Roles of motor proteins in building microtubule-based structures: a basic principle of cellular design. AB - Eukaryotic cells must build a complex infrastructure of microtubules (MTs) and associated proteins to carry out a variety of functions. A growing body of evidence indicates that a major function of MT-associated motor proteins is to assemble and maintain this infrastructure. In this context, we examine the mechanisms utilized by motors to construct the arrays of MTs and associated proteins contained within the mitotic spindle, neuronal processes, and ciliary axonemes. We focus on the capacity of motors to drive the 'sliding filament mechanism' that is involved in the construction and maintenance of spindles, axons and dendrites, and on a type of particle transport called 'intraflagellar transport' which contributes to the assembly and maintenance of axonemes. PMID- 10722883 TI - Understanding the functions of kinesin-II. AB - Species ranging from Chlamydomonas to humans possess the heterotrimeric kinesin II holoenzyme composed of two different motor subunits and one non-motor accessory subunit. An important function of kinesin-II is that it transports the components needed for the construction and maintenance of cilia and flagella from the site of synthesis in the cell body to the site of growth at the distal tip. Recent work suggests that kinesin-II does not directly interact with these components, but rather via a large protein complex, which has been termed a raft (intraflagellar transport (IFT)). While ciliary transport is the best-established function for kinesin-II, evidence has been reported for possible roles in neuronal transport, melanosome transport, the secretory pathway and during mitosis. PMID- 10722884 TI - Lack of carcinogenicity of gardenia blue colour given chronically in the diet to F344 rats. AB - The carcinogenicity of gardenia blue colour was examined in Fischer 344 (F344) rats. Groups of 50 males and 50 females were given the material at dietary doses of 0 (control), 2.5 or 5% for 104 weeks and then sacrificed. The doses were selected on the basis of results from a 13-week subchronic toxicity study. A slight increase in relative organ weights of the left lung was observed in male rats of the 5% group. However, no significant differences between the control and treated groups were noted with regard to clinical signs, mortality and haematological findings. A variety of tumours developed in all groups, including the controls, but all were histologically similar to those known to occur spontaneously in F344 rats, and no statistically significant increase in the incidence of any type of neoplastic lesion was found for either sex in the treated groups. Thus, it was concluded that, under the present experimental conditions, gardenia blue colour is not carcinogenic in F344 rats. PMID- 10722885 TI - Lack of oestrogenic effects of food preservatives (parabens) in uterotrophic assays. AB - The oestrogenic activity of the parabens, methyl-, ethyl- and propyl p hydroxybenzoate, widely used as antimicrobials in food, and butyl p hydroxybenzoate, which is used in cosmetic products, and their shared main metabolite p-hydroxybenzoic acid was investigated in a mouse uterotrophic assay. Immature B6D2F1 mice were treated with oral or subcutaneous doses of the test compounds for three consecutive days. p-Hydroxybenzoic acid and butyl p hydroxybenzoate were also tested by the subcutaneous route in a rat uterotrophic assay. A significant increase in the uterus weight at day 4 was considered an oestrogenic effect. In the mouse assay, none of the compounds tested produced any oestrogenic response at dose levels up to 100mg/kg body weight per day, for ethyl p-hydroxybenzoate even at a dose level of 1000mg/kg body weight per day. In immature Wistar rats, subcutaneous administration of butyl p-hydroxybenzoate produced a weak oestrogenic response at 600mg/kg body weight per day. PMID- 10722886 TI - The effect of maternal exposure to flaxseed on spermatogenesis in F(1) generation rats. AB - Pregnant Sprague-Dawley rats were exposed to a flaxseed (20 or 40%), flaxmeal (13 or 26%) or standard NIH AIN-93 (0% flaxseed control) diet throughout gestation and until their offspring were weaned. After weaning, F(1) generation males were placed in the same diet treatment groups as their mothers for 70 days. Statistically significant differences were not observed between either low-dose or high-dose flaxseed and flaxmeal-treated animals and the 0% flaxseed control animals for testis weights, homogenization resistant spermatid counts, daily sperm production rates, epididymal weights, seminal vesicle weights, seminiferous tubule fluid testosterone concentrations and the percentage of sperm abnormalities. The following statistically significant differences were observed when treated groups and the 0% flaxseed control groups were compared: (1) increases in serum LH in the 20% and 40% flaxseed treatment groups and in serum LH and testosterone in the 26% flaxmeal treatment group; (2) increases in the cauda epididymal weight from the 20% and 40% flaxseed groups; (3) increases in cauda epididymal sperm numbers/g epididymis from the 20% and 40% flaxseed and the 13% and 26% flaxmeal treatment groups; (4) a decrease in prostatic weight from the 20% flaxseed and 13% and 26% flaxmeal treatment groups. Prostate weight in the 40% flaxseed treatment group was lower but not statistically significantly different than the 0% flaxseed control group. Histological effects on spermatogenesis were not observed in either the control group, flaxmeal or the flaxseed treated groups. PMID- 10722887 TI - Lack of DNA-damaging activity of five non-nutritive sweeteners in the rat hepatocyte/DNA repair assay. AB - The non-nutritive sweeteners acesulfame-K, aspartame, cyclamate, saccharin and sucralose were tested for DNA damaging activity in the rat hepatocyte/DNA repair assay. Using hepatocytes from F344 and Sprague-Dawley male rats, all were inactive despite strong responses for the positive control, 2-aminofluorene. PMID- 10722888 TI - Promotion of morphological transformation by Di-n-butyltin dichloride in C3H/10T1/2 cells: prediction by prior expression of tumour promoter-responsive genes. AB - Previous studies in our laboratory have shown that chemical treatments may induce increases in proliferin gene family mRNA accumulation in cultured murine embryonic cells. Proliferin inductions are highly correlated with subsequent promotional outcomes during two-stage focus-formation assays in C3H/10T1/2 cell cultures. In work reported here, the strong affiliation between these two responses was further validated after treating cells with di-n-butyltin dichloride which is a polyvinyl chloride (PVC) plastic additive that often contaminates food and water. Increased proliferin expression and promotion of morphological transformation occurred at similar concentrations. Promotion of transformation was detected at di-n-butyltin dichloride concentrations of 80 nM (24 ng/ml) and above, if added to initiated cultures before confluent monolayers had formed. Proliferin induction and morphological transformation were both reduced in confluent cultures treated with di-n-butyltin dichloride, as compared to subconfluent cultures. Proliferin expression measured in near-confluent cultures was induced up to 10-fold during the 36-hr period following di-n butyltin dichloride exposure and was accompanied by increased accumulation of transcripts from many genes regulated by oxidative stresses, growth-inducing agents, and/or other promoting agents (asbestos, superoxide radicals ). Di-n butyltin dichloride-induced mRNA species included members of the fos and jun proto-oncogene families, c-myc, egr1, ribonucleotide reductase (R2 subunit), odc, macrophage chemotactic protein/je, hsp70, metallothionine IIA, c-sod and mn-sod. The observed patterns of RNA accumulation suggested that a small subset of mRNA species, including proliferin, exhibit regulatory behaviour as a response to dissimilar agents or conditions that promote focus-formation in C3H/10T1/2 cultures. Plausible predictions of promotional effects in two-stage morphological transformation assays can be made from gene-expression responses to test agents. PMID- 10722889 TI - Divergent antibody isotype responses induced in mice by systemic exposure to proteins: a comparison of ovalbumin with bovine serum albumin. AB - Whereas many foreign proteins are immunogenic, only a proportion is associated commonly with allergy, having the potential to induce the quality of immune response necessary for IgE antibody production and the development of immediate type hypersensitivity reactions in the gastrointestinal and/or respiratory tracts. In the context of toxicological evaluations there is a need to identify those properties that confer on proteins the ability to provoke allergic reactions. The characteristics of antibody responses induced in BALB/c strain mice following administration of ovalbumin (OVA), a significant human allergen, have been compared with those provoked by bovine serum albumin (BSA), a protein considered to have more limited allergenic potential. Intranasal or intraperitoneal (ip) administration of BSA or OVA elicited vigorous IgG and IgG1 antibody responses. Differential IgE antibody production was observed, however, with OVA stimulating relatively high IgE antibody titres at all doses tested whereas no or low titre IgE antibody was detected following exposure to BSA. Furthermore, a differential capacity for IgG2a antibody responses was observed, with only BSA provoking high titres of this IgG subclass. The relative quality of induced responses was equivalent following administration of these proteins via mucosal (in) tissue or via a non-mucosal (ip) route of exposure. IgG2a antibody production is promoted by the type 1 cytokine interferon gamma (IFN-gamma), whereas IFN-gamma and the type 2 cell product interleukin 4 exert reciprocal antagonistic effects on IgE antibody responses. Although cytokine expression patterns were not analysed in this series of experiments, the differential IgE and IgG subclass antibody responses induced by BSA and OVA are consistent with the preferential activation of T helper (Th) 1- and Th2-type cells, respectively. These data indicate that proteins can provoke in mice characteristic antibody (IgE and IgG) isotype profiles suggestive of discrete T lymphocyte responses and that such differences may be associated with variable allergenic activity. PMID- 10722890 TI - Percutaneous penetration and dermal metabolism of triclosan (2,4, 4'-trichloro-2' hydroxydiphenyl ether). AB - triclosan is widely used in many products that contact the skin of consumers. This study compares in vivo and in vitro skin absorption of triclosan and determines the potential of skin to metobolize it prior to entering the blood stream. After in vivo topical application of a 64.5mM alcoholic solution of [(3)H]triclosan to rat skin, 12% radioactivity was recovered in the faeces, 8% in the carcass 1% in the urine, 30% in the stratum corneum and 26% was rinsed from the skin surface at 24 hours after application. Free triclosan and the glucuronide and sulfate conjugates of triclosan were found in urine and faeces. triclosan penetrated rat skin more rapidly and extensively than human skin in vitro. 23% of the dose had penetrated completely through rat skin into the receptor fluid by 24 hours, whereas penetration through human skin was only 6.3% of the dose. Chromatographic analysis of the receptor solutions showed that triclosan was metabolized to the glucuronide, and to a lesser extent to the sulfate, during passage through the skin. triclosan glucuronide appeared rapidly in the receptor fluid whereas triclosan sulfate remained in the skin. Although the major site of metabolism was the liver, conjugation of triclosan in skin was also demonstrated in vitro and in vivo, particularly to the glucuronide conjugate which was more readily removed from the skin. The in vitro system provides a reasonable estimate of dermal absorption in vivo for the rat. Therefore by extrapolation of the comparative in vitro data for human and rat skin it is reasonable to deduce that dermal absorption in human of triclosan applied at the same dose is about one-third of that in the rat in vivo. PMID- 10722891 TI - "IARC group 2A Carcinogens" reported in cigarette mainstream smoke. AB - As a follow-up to an earlier study on IARC Group I compounds, further efforts have been made to evaluate the international literature on cigarette mainstream smoke for reports on constituents classified as IARC "Group 2A: probably carcinogenic to humans" and IARC "Group 2B: possibly carcinogenic to humans." IARC classifies 59 agents, mixtures and exposures as Group 2A. Of the overall list of 59, 50 represent chemical entities or complex mixtures ( [IARC,] ). When only chemical entities which have their origin from cigarette components (tobacco and paper) are considered, further searching of the international literature has revealed that nine chemical compounds of the 50 Group 2A listings have been reported in cigarette mainstream smoke ( Table 1 ). In micrograms/cigarette (mug/cig), the ranges reported for each of the nine compounds are as follows: formaldehyde (3.4-283); benzo[a]pyrene (B[a]P) (0.004-0. 108); dibenz[a,h]anthracene (DB[a,h]A) (0.004-0.076); N-nitrosodiethylamine (DEN) (non detectable-0.0076); benz[a]anthracene (B[a]A) (trace-0.08); N nitrosodimethylamine (DMN) (non-detectable-0.7-1.62); acrylamide (1.1-2.34); 1,3 butadiene (16-77); and 2-amino-3-methyl-3H-imidazo[4,5-f]quinoline (IQ) (0. 00026 0.00049). PMID- 10722892 TI - Allergenicity of refined vegetable oils. AB - Several commercially important refined vegetable oils are derived from plants which are recognized as potent food allergens (e.g. peanut, soy). Full refining of oils results in the almost complete removal from oils of protein, which is responsible for allergic reactions. However, it is uncertain whether the minute amounts remaining could provoke allergic reactions in highly susceptible individuals. This has led to a vigorous debate about the safety of refined oils and specifically whether to label each oil individually because of the potential risk of allergenicity. Peanut oil has been the most thoroughly studied. It has been shown, in well-designed studies, that refined peanut oil can be safely consumed by the vast majority of peanut-allergic individuals, whereas unrefined oil can provoke reactions in some of the same individuals. However, some other studies report cases of allergic individuals reacting to oils, which are presumed to be refined. While it is likely that the discrepancy between these observations is due to differences in the processing of the oils, and possibly the protein content, this has not been formally demonstrated. Few data exist on the potential allergenicity of other edible vegetable oils; what data there are suggest that the major oils (soy, maize, sunflower, palm) do not provoke allergic reactions in susceptible individuals. Determining the content and immunoreactivity of the residual protein of refined oils is crucial to assessing the allergenic risk they present. Current methodology is inadequate and has not been validated for use with oils and aqueous extracts from oils. Little is known about the importance of different processing steps on allergenicity, although this information is crucial to risk assessment, particularly when considering process modifications. Available data suggest that the protein content of crude oils is of the order of 100-300 microg and that refining results in levels up to about 100-fold lower. The review concludes that peanut oil, and by extrapolation other edible vegetable oils, presents no risk of provoking allergic reactions in the overwhelming majority of susceptible people. However, there is a need to standardize and validate methodology for measuring the protein content and immunoreactivity of such so that they can be used to maintain process specifications. Thresholds of reactivity to allergens in man also need to be established in order to assess fully the risk from very small amounts. PMID- 10722893 TI - A systematic review of single-tooth restorations supported by implants. AB - OBJECTIVES: To make an inventory of clinical studies on single-tooth restorations supported by implants using a systematic review procedure and to aggregate overall survival results. DATA SOURCES: Papers referring to single-tooth implants were located by a MEDLINE search 1990 to April 1998. Three hundred and twenty references were found, and they were subjected to a systematic review procedure. STUDY SELECTION: A three-step inclusion/exclusion procedure was applied to identify papers that represented: good scientific practice (GSP), reported results of all patients, implants and crowns for more than 2years, and had sufficient data to generate life-table analyses. The outcomes were 'implant failure' and 'crown completion'. Nine studies survived. These data showed an overall mean GSP of 0.37 with a predicted 4year implant survival of 97% (n=459), and an uncomplicated crown maintenance of 83% (n=240). CONCLUSION: Single-tooth implants show an acceptable short-term survival of 4years, but crown complications are common. PMID- 10722894 TI - The usefulness of radiographs in diagnosis and management of periodontal diseases: a review. AB - OBJECTIVES: To review the periodontally significant diagnostic information obtainable from radiographs and the stages during periodontal therapy when the information may influence patient management and treatment outcomes. DATA: Confined to studies involving conventional radiography, as this remains the commonest imaging method in clinical dental practice and primary dental care setting. SOURCES: Literature was reviewed using Medline and manual tracing of references cited in key papers not otherwise elicited. STUDY SELECTION: Studies were selected in order to (i) define the role of radiographs in periodontal diagnosis and management at the initial, corrective and supportive (maintenance) phases of periodontal therapy and (ii) critically review the evidence for the value added by radiographs. CONCLUSIONS: Radiographs provide diagnostic information on alveolar bone levels, plaque retention factors, caries, furcation defects, subgingival calculus and additional pathology. Features visualised are dependent on the radiographic view. A relationship exists between probing attachment loss and radiographic bone height, with a range in level of correlation; clinical attachment may correspond more closely to surgical measurements of bone height. Radiographs can be used in planning initial, corrective and supportive phases of therapy, though some decisions may be made on clinical assessments alone. Evidence in the literature on benefit gained from radiographs taken for periodontal patients is sparse; the extent to which they influence the treatment provided and treatment outcomes is poorly addressed. Further research is indicated to define the role of radiographs when managing the periodontal patient to maximise the potential gain for the patient. PMID- 10722895 TI - Quality evaluation of process of root canal treatments performed on young adults in Finnish public oral health service. AB - OBJECTIVES: To evaluate the quality of root canal treatments performed in every day practice on young adults. METHODS: A quality index related to the treatment process was developed based on the guidelines of the European Society of Endodontology. A random computerised selection of 134 young adults (born 1966 1971) produced 125 (93%) eligible oral health documents, including information on 148 teeth that were root canal treated by 47 dentists. The process of root canal treatment was evaluated according to the original oral health documents. The technical quality of root canal fillings was assessed on postoperative periapical radiographs. RESULTS: The overall Kappa statistics for the inter-examiner reliability of assessments was 0.63; the proportional agreement being 87%. The mean quality index was 6.2, s.d. 2.0, with none of the treatments scoring the maximum 13. On the 56 available postoperative radiographs, 52% of the fillings were optimal, showing a length within 0-3mm from the radiological apex, no voids in the apical part nor lumen apical to the filling. Of the completed 144 root canal treatments, only 11 included some recorded follow-up information within 15months from the termination of the treatment. CONCLUSIONS: The technical quality of the root fillings was similar to that found in previous studies. However, the quality index, paucity of pre- and post-operative radiographs, and the lack of follow-up all indicated a discrepancy between consensus guidelines and every-day practice. PMID- 10722896 TI - Restorative treatment provided over five years for adults regularly attending general dental practice. AB - OBJECTIVES: To investigate the distribution and type of restorative treatment, including re-treatment, provided for adults who attend annually. METHODS: In 1991 a selected group of 24 general dental practitioners in the North West of England recruited 4211 of their regularly attending adult patients. Dentists recorded the reason for and type of treatment provided during the following 5 years. RESULTS: Approximately 40% of the participants received treatment (restorations and/ or extractions) at each annual examination. Of the 2293 patients who attended every examination 1959 (85%) had received a restoration and/ or extraction during the 5 years. A total of 8187 teeth, 15% of those present at baseline, received treatment, 3030 (37%) for caries and 5157 (63%) for other reasons. The proportion of adults who received treatment by age group differed significantly with those aged 25-34 years (80%) being least likely and those 35-44 years of age (89%) most likely. Of the 1744 teeth restored in the first year of the study, 170 (10%) were retreated within 1 year and 402 (23%) during the subsequent 4 years. The 4 year survival of amalgam and tooth coloured fillings was 84% and that of crowns 92%. CONCLUSIONS: This study documented the extent and type of restorative care provided for regularly attending adults during a 5-year period. The majority of treatment was provided for reasons other than caries. Of the teeth restored over the first year, 23% were retreated in the subsequent 4 years. PMID- 10722897 TI - A new electrical method for detecting marginal leakage of in vitro resin restorations. AB - OBJECTIVES: Ingress of bacteria at sites of marginal leakage has been suggested to cause pulpal inflammation. The purpose of this in vitro study was to evaluate the validity of a new electrical method to detect marginal leakage of restoratives by comparing the results obtained with those of a dye penetration test. METHODS: After cavities were prepared on the buccal coronal surfaces and root surfaces of 16 extracted non-carious human molar teeth, eight specimens were treated with a dentin bonding system (bonding group) and the other eight specimens were not treated (non-bonding group). Resin composites were filled in the cavities of all specimens, and physiological saline was applied to the margin of the restorative. Any excess saline was wiped off, leaving only the electrolyte, which had penetrated into the marginal gap. The change in conductance was measured continuously across the margin of each specimen during this process. The marginal leakage of specimens was confirmed using the dye penetration test, and the results were evaluated by the microleakage score. RESULTS: In both coronal and root surface cavities, the changes in conductance in the non-bonding group after filling were significantly larger than those of the bonding group (p<0.05). The change in conductance of each specimen correlated with the microleakage score (p<0.05). CONCLUSIONS: It was concluded that the relative electrical method could detect marginal leakage in both coronal and root surface cavities. PMID- 10722898 TI - In vitro caries inhibition at the enamel margins of glass ionomer restoratives developed for the ART approach. AB - OBJECTIVES: To assess the ability of conventional glass ionomer cements manufactured specifically for the atraumatic restorative treatment (ART) approach to inhibit the in vitro demineralization of enamel. METHODS: Twenty-four sound permanent premolar teeth, extracted for orthodontic reasons, had cervical cavities (4x2x1. 5mm(3)) prepared in enamel. These were restored with Fuji IX, Fuji IX GP, Ketac-Molar and Compoglass, and then thermocycled 300 times between 5 55 degrees C before being placed in a demineralizing solution (0.1M lactic acid with 1g/l dissolved hydroxyapatite at pH 4.7) for four weeks. Buccolingual planoparallel sections were cut axially through the restorations, and subsequently lapped to approximately 100 microm thickness. The sections were examined with a polarized light microscope, and lesion measurements made using image analysis software. ANOVA and coefficients of variance were used to compare the findings. RESULTS: Compoglass and Ketac-Molar showed significantly less surface erosion than did the other two cements (p<0.0001). Inhibition of enamel demineralization immediately adjacent to the restoration margins was more frequent with the glass ionomer cements (20.5-25.0%) than with Compoglass (13.0%). However, the widths of the inhibition zones varied between materials and sites. CONCLUSIONS: Fluoride ion release from the restorative materials afforded some degree of protection to the adjacent enamel against in vitro demineralization. PMID- 10722899 TI - Erosion of deciduous and permanent dental hard tissue in the oral environment. AB - OBJECTIVES: The objectives of this study were two-fold: (1) to determine (by surfometry) loss of deciduous and permanent enamel and dentine following consumption of a single low pH orange drink for 15days; and (2) to determine (by surfometry) loss of deciduous and permanent enamel and dentine following consumption of the product 2 versus 4 times per day for 15days. METHODS: Sixteen healthy volunteers participated in a single centre, single blind, 2-phase crossover study, conducted according to Good Clinical Practice, and employing the validated model described by West and co-workers (Journal of Dentistry 1998; 26:329-335). RESULTS: In all tissues, erosion was progressive over time, the pattern being more linear in enamel than in dentine. In general, erosion of deciduous enamel was greater than that of permanent enamel, though this difference was significant only for those specimens exposed to 4 drinks per day. Conversely, erosion of dentine was generally greater in the permanent tissue, though differences rarely reached conventional levels of statistical significance. Increasing frequency of consumption resulted in increased loss of tissue, but this difference was neither proportional nor consistently statistically significant. CONCLUSIONS: It is concluded that statistically significant differences in susceptibility of deciduous and permanent enamel to erosion appear to emerge over time and with increasing frequency of consumption. This is of importance clinically given the reduced dimensions of the deciduous dentition and the element of 'abuse' of soft drinks by the child population. Further development of soft drinks with low erosive potential is recommended. PMID- 10722900 TI - Relative susceptibility of deciduous and permanent dental hard tissues to erosion by a low pH fruit drink in vitro. AB - OBJECTIVES: The objectives of this study were two-fold: (1) to determine (by surfometry) loss of deciduous and permanent enamel and dentine following 15days' exposure to a single low pH orange drink; and (2) to determine (by surfometry) loss of deciduous and permanent enamel and dentine following exposure to the product 2 versus 4 times per day for 15days. METHODS: This in vitro study employed the validated methodology described by West and co-workers [Journal of Dentistry, 1998;26:329-335.] RESULTS: In all four tissues, erosion was progressive over time, though this pattern was more linear in enamel than in dentine. In general, erosion of enamel was greater in the deciduous tissue, while erosion of dentine was greater in the permanent tissue. However, these differences were rarely of statistical significance. Increasing frequency of exposure resulted in a non-proportional increase in tissue loss. CONCLUSIONS: Differences in susceptibility of deciduous and permanent tissues to erosion by a low pH drink in vitro appear to exist, though these may not be of statistical significance. Care may be indicated in the delivery of dietary advice, since reduced frequency of exposure to a low pH drink does not appear to result in a proportional reduction in tissue loss. PMID- 10722901 TI - In vitro approach to evaluate potential harmful effects of beer on teeth. AB - OBJECTIVE: The purpose of the present work was to examine some properties of different brands of beer manufactured in Brazil that may be important to oral health. METHODS: Samples from seven different beer brands were analyzed for pH, titratable acidity, calcium and phosphate concentrations. Demineralization experiments were carried out by incubating samples with crown tooth particles (40 80 mesh) at 37 degrees C under agitation (100 strokes/min). RESULTS: The pH was lower than 4.0 for three of the seven samples and higher than 4.0 for the others. The amount of titratable acidity, expressed as the volume of 0.1N NaOH solution consumed to raise the initial pH to 7.0, and the concentrations of calcium and phosphate varied. Calcium concentration ranged from 0.21 to 1.59 micromol/ml, while phosphate concentration varied from 0.048 to 0.094 micromol/ml. Calcium released to the incubation medium was proportional to the time of incubation up to 5min. Maltose, a disaccharide, was detected in all samples studied. CONCLUSION: Differences in the properties examined indicated that some brands of beer studied may have potential dental effects. PMID- 10722902 TI - Human odontoblast cell numbers after dental injury. AB - OBJECTIVES: The purpose of this study was to measure the changes in odontoblast cell numbers in response to cavity restoration variables and patient factors, and the effect these factors have on dental repair by tertiary dentinogenesis. The number of vital odontoblasts is a critical factor for pulpal repair following restorative surgery, and yet little information is available on these cell numbers. METHODS: Class V non-exposed cavities were prepared in the buccal surface of intact first or second premolar teeth of 27 patients, between 9 and 17 years of age. Following tooth extraction (28-163 days) the area of reactionary dentine and the area of the odontoblasts were measured histomorphometrically. RESULTS: Patient factors, as well as cavity preparation and restoration variables, had little effect on the numbers of odontoblasts per pulpal unit area. However, the age of the patient did appear to have an effect on the reactionary dentine secretory capacity of odontoblasts per unit area, and on the relative number of odontoblasts beneath cut dentinal tubules. CONCLUSIONS: Odontoblast cell numbers were maintained following the preparation of cavities cut into dentine with a 0.5mm residual dentine thickness. The repair capacity of the pulp dentine complex would appear to be age dependent, this may explain differences in the success of various restorative treatments between patients. PMID- 10722903 TI - Effect of thione primers on bonding of noble metal alloys with an adhesive resin. AB - OBJECTIVES: The purpose of the current study was to evaluate the effects of two metal conditioners on the bond durability of an adhesive resin joined to noble metal alloys by comparing pre- and post-thermocycling bond strengths. METHODS: Two different sizes of disk specimens (10 and 8mm in diameter by 2.5mm thickness) were prepared from silver-indium (Ag-In-Zn, Salivan), silver-palladium-copper gold (Ag-Pd-Cu, Castwell M.C.12), metal-ceramic gold (Au-Pt-Pd, Degudent Universal), metal-ceramic palladium (Pd-Ga-Co, PTM 88), type IV gold (type IV, Casting Gold) alloys, and pure silver (pure Ag). The specimens were air-abraded with 50-microm grain sized alumina, conditioned either with a thiouracil primer (Metaltite) or with a thione-phosphate primer (Alloy Primer), then bonded with an adhesive resin (Super-Bond Opaque). Shear bond strengths were determined both before and after repeated thermocycling (4 degrees C and 60 degrees C, 1min each, 100, 000cycles). The results were compared by analysis of variance and post-hoc multiple comparison intervals. RESULTS: The average post-thermocycling bond strengths in MPa (n=8) generated with the thiouracil primed and thione-phosphate primed groups, respectively, were: 3.4 and 5.8 for the Ag-In-Zn alloy, 40.4 and 37.7 for the Ag-Pd-Cu alloy, 26.4 and 33.5 for the Au-Pt-Pd alloy, 27.4 and 36.6 for the Pd-Ga-Co alloy, 40.2 and 40.3 for the type IV alloy, and 37.3 and 32.4 for the pure Ag. The Ag-In-Zn alloy exhibited significantly lower bond strength than the other alloys, whereas the Ag-Pd-Cu and type IV alloys exhibited the greatest magnitude of bond strength for both primers (p<0.05). CONCLUSIONS: It can be concluded that the combined use of either of the two thione primers and the adhesive resin is effective for bonding the metal/alloys examined, with the exception of the Ag-In-Zn alloy. PMID- 10722904 TI - Reasons for tooth extraction in Scotland. AB - OBJECTIVES: A 1984 study investigated the reasons underlying the extraction of teeth in Scotland. The survey described in this paper, used a similar methodology and aimed to determine the reasons for the extraction of permanent teeth by general dental practitioners and investigate changes in the influences on tooth extraction over a 10 year period. METHODS: During a 1 week period in November 1994, 139 general dental practitioners working throughout Scotland, recorded the reasons for all permanent tooth extractions. RESULTS: A total of 917 permanent teeth were extracted from 613 patients, the reason for extraction being stated as dental caries (51%), periodontal disease (21%), orthodontics (11%) and failed endodontics (4%). Trauma, pericoronitis and other reasons accounted for 5.5% of extractions whilst, in 7.5% of cases, patients requested extraction in preference to other treatments. The proportion of extractions attributed to periodontal disease increased from age 31-60 years, but declined thereafter. CONCLUSIONS: Comparing the results with those obtained in the 1984 study, whilst the mean number of teeth extracted by each practitioner had reduced, the overall relative contribution of different reasons for extraction was similar. PMID- 10722905 TI - Three-dimensional ultrasound in the assessment of fetal cerebellar transverse and antero-posterior diameters. AB - Fetal cerebellum scanning by prenatal ultrasound (US) is very important for early detection of fetal central-nervous-system anomaly, as well as for the determination of gestational age (GA). Due to the small organ size and the unique shape of the fetal cerebellum (CL), accurate measurement of the dimensions of CL by two-dimensional (2-D) US is not easy if the appropriate plane cannot be reached. With the advent of three-dimensional (3-D) US, the disadvantages of 2-D US in assessing the fetal CL dimensions can be avoided. The purpose of this study was to assess the fetal cerebellar transverse diameter (CTD) and cerebellar antero-posterior diameter (CAD) using 3-D US. First, we compared the reproducibility of 2-D and 3-D US on the assessment of fetal cerebellar dimensions. Second, we prospectively measured CTD and CAD in 223 healthy fetuses using a cross-sectional design with an attempt to establish the normal growth charts of fetal CL. Our results showed 3-D US is superior to 2-D US in the reproducibility test of fetal cerebellar dimensions. In addition, with GA as the dependent variable, polynomial regression analysis showed that the best-fit equations for both CTD vs. GA and CAD vs. GA were the first-order. The best-fit predictive equation of GA by CTD was GA (week) = 9.0281 + 0. 58533 x CTD (mm) (r = 0.95, n = 223, SE = 1.82 weeks, p < 0.0001), and the best-fit predictive equation of GA by CAD was GA (week) = 10. 855 + 1.1672 x CAD (mm) (r = 0.82, n = 223, SE = 3.41 weeks, p < 0. 0001). Furthermore, all the correlation coefficients of CTD or CAD vs. the common fetal growth indexes were also highly significant (all p < 0.0001). In conclusion, our data of fetal CL dimensions assessed by 3-D US may serve as a useful reference in assessing fetal CL growth, dating GA or detecting fetal CL anomalies. PMID- 10722906 TI - Evaluation of tissue harmonic imaging for the diagnosis of focal liver lesions. AB - This was a prospective study to evaluate tissue harmonic imaging (THI) for the diagnosis of focal liver lesions. A total of 15 reviewers read 100 randomly arranged liver images, a fundamental grey-scale image (FGI) and a THI (transmitted: 2 MHz, received: 4 MHz) of each of 50 patients (29 with liver cirrhosis, 42 with focal lesions) taken from the same section. The mean value of overall accuracy for detecting lesions (presence or absence) was significantly higher with THI (82.3%) than with FGI (79.6%) (t = 1. 96, p< 0.05). When only the 29 cirrhosis patients were analyzed, the difference was more significant (t = 2.48, p < 0.02). The correct count rate of the number of focal lesions was higher with THI (78. 0%) than with FGI (67.0%) (t = 3.61, p< 0.005) in 23 cirrhosis patients with focal lesions. The correct diagnosis of HCC was achieved at a higher rate with THI (42.5%) than with FGI (36.8%). THI was statistically effective for detecting focal lesions, particularly in cirrhotic livers. PMID- 10722907 TI - Transcranial sonography of the brain parenchyma: comparison of B-mode imaging and tissue harmonic imaging. AB - Transcranial color-coded Duplex sonography (TCCS) has been used for the identification of cerebrovascular disorders. Recently, its value in the diagnosis of disorders of the brain parenchyma has been proposed. The object of this study was to determine systematically the echo pattern of the brain parenchyma and to compare conventional B-mode imaging with tissue harmonic imaging (THI). Transcranial sonography (TCS) was performed in 54 healthy individuals through the temporal bone window using conventional B-mode imaging and THI by two experienced investigators. Identification rates for several brain structures were assessed, and the quality of depiction of each method was graded semiquantitatively. In addition, several parts of the ventricular system and the basal cerebral cisterns were measured. Four subjects did not have an adequate bone window for transcranial examination. In the remaining people, the bone window was assessed to be adequate (59%) or excellent (33%). In the majority (> 80%), TCS allowed an unequivocal identification of various brain structures. Inter-rater variability of the assessments of tissue echogenicity and measurements of the ventricular width were found to be low for several structures (e.g., brainstem, thalamus, or 3rd ventricle). The echo pattern of brain tissue in THI is identical to that described for B-mode imaging. Using THI, contours of brain structures were typically visualized more clearly and the reproducibility of measurements was more consistent. In our experience, insonation of the contralateral lobes was limited when depths were higher than 12 cm using THI. In conclusion, TCS allowed the sonographic examination of the brain parenchyma in the majority of our subjects. THI substantially improves the identification of parenchymal structures when the depth is below 12 cm. PMID- 10722908 TI - Evaluation of posterior cerebral artery flow velocity by transcranial color-coded real-time sonography. AB - Using transcranial color-coded real-time sonography (TCCS), we measured peak systolic flow velocities (PSVs) in segment P2 of 102 posterior cerebral arteries (PCAs) in 61 patients, with angiography. We divided 102 PCAs into four groups: control group (n = 70) with no significant stenotic lesions; PCS group (n = 7) with stenosis >/= 50% of P2 segment; Col (+) group (n = 13) and Col (-) group (n = 12) had occlusive lesions in the carotid system with or without collateral flow from PCA to the middle cerebral artery through the leptomeningeal anastomosis. In the PCS group, PSV (255.7 +/- 67.2 cm/s) was higher than in the other three groups (p < 0.0001). PSV was higher in the Col (+) group (127.6 +/- 31.2) than in the Col (-) (86.6 +/- 20.1) and control (83.8 +/- 24.8) groups (p < 0.001). The measurement of PSV in the P2 segment of PCA using TCCS may help to identify a significant stenosis in PCA. PMID- 10722909 TI - Real-time 3-D ultrasound acquisition and display for cardiac volume and ejection fraction evaluation. AB - The objective of this study was to test if three-dimensional (3-D) ultrasound (US) provides accurate determination of the cardiac volumes and ejection fraction. The 3-D device (Model 1-Volumetrics, ) is a 3-D acquisition system using a 2-Mhz matrix probe that insonates the whole cardiac volume in a 4-chamber view and collects the entire backscattered US echoes from this volume within one cardiac cycle. The complete 3-D US information stored in the memory can then be cut into 2-D views of any arbitrary orientation. For volume determination, the best 4-chamber view was selected into the memory, then 6 transverse views were displayed at different depths along the ventricle long axis, and the contour of the ventricle was drawn on each of these views. The left ventricle volume in diastole (LVDV) and the ejection fraction (EF) obtained by 3-D US were compared with those from x-ray and isotopic angiographies, and 2-D echo-time motion (2-D Echo-TM). The variations in stroke volume (SV) during a stand test, measured by 3 D US, and aortic Doppler were compared. The correlation between EF evaluated from 3-D US and x-ray or isotopic angiographies was found to be good (r = 0.80 p < 0.001; r = 0.86 p < 0.001), but lower with 2-D Echo-TM (r = 0.59 p < 0.001). For LVDV, the correlation was acceptable with x-ray angiography (r = 0.75 p < 0.001), but much lower with isotopic angiography and 2-D Echo-TM (r = 0.47 p < 0.001; r = 0.55 p < 0.001). A good correlation was also found between the SV changes measured by 3-D US and aortic Doppler (r = 0.79 p < 0.001). PMID- 10722910 TI - Calculating blood flow from Doppler measurements in the systemic-to-pulmonary artery shunt after the Norwood operation: a method based on computational fluid dynamics. AB - Hypoplastic left heart syndrome is currently the most lethal cardiac malformation of the newborn infant. Survival following a Norwood operation depends on the balance between systemic and pulmonary blood flow, which is highly dependent on the fluid dynamics through the interposition shunt between the two circulations. We used computational fluid dynamic (CFD) models to determine the velocity profile in a systemic-to-pulmonary artery shunt and suggested a simplified method of calculating the blood flow in the shunt based on Doppler measurements. CFD models of systemic-to-pulmonary shunts based on the finite element method were studied. The size of the shunt has been varied from 3 to 5 mm. Velocity profiles at proximal and distal positions were evaluated and correlations between maximum and mean spatial velocity were found. Twenty-one Doppler measurements in the proximal and distal part of the shunt were obtained from six patients with hypoplastic left heart syndrome. Combining Doppler velocities and CFD velocity profiles, blood flow rate in the shunt was calculated. Flow rate evaluated from aortic Doppler and oxygen saturation measurements were performed for comparison. Results showed that proximal shunt Doppler velocities were always greater than the correspondent distal ones (ratio equal to 1.15 +/- 0.11). CFD models showed a similar behaviour (ratio equal to 1.21 +/- 0.03). CFD models gave a V(mean)/V(max) ratio of 0. 480 at the proximal junction and of 0.579 at the distal one. The agreement between the flow evaluated in the proximal and distal areas of the shunt was good (0.576 +/- 0.150 vs. 0.610 +/- 0.166 l/min). Comparison of these data with saturation data and aortic Doppler measurements correlate less well (0.593 +/- 0.156 vs. 1.023 +/- 0.493 l/min). A formula easily to quantify shunt flow rate is proposed. This could be used to evaluate the effects of different therapeutic and pharmacological manoeuvres in this unique circulation. PMID- 10722911 TI - Accuracy of coronary flow measurements performed by means of Doppler wires. AB - To estimate in patients the accuracy of coronary flow measurements performed by means of 0.014" Doppler wires, the time-averaged maximal blood velocity (APV) was recorded in the 3 branches of 36 angiographically normal coronary artery bifurcations selected in 21 patients undergoing cardiac catheterization for various diseases. Contrast medium injections filmed under two incidences allowed identification of the 3 sample volume locations and computing of the 3 corresponding vessel cross-sectional areas (CSA) at subsequent data analysis. Multiplication of the velocities APV/2 (range: 3 to 20.5 cm/s) by the CSA (obtained by averaging the two calibrated vessel diameters; range: 1.6 to 5.4 mm) yielded 108 flow rates (range: 5.4 to 169 mL/min). The average relative flow error was then estimated using the continuity equation (Q(in) = Q(out,1) + Q(out, 2)) and the central limit theorem. The result was that the relative flow error decreased from 30% at Q = 30 mL/min to 13% at Q = 160 mL/min. We conclude that coronary flow measurements are reasonably accurate, except perhaps for very low flows. PMID- 10722912 TI - Assessment of myocardial velocities in healthy children using tissue Doppler imaging. AB - The objective was to determine the normal range of tissue velocities in paediatric hearts as measured by tissue Doppler imaging. A prospective study was carried out involving 160 healthy children (mean age 10.8 y, range 4.0-17.9 y). Using tissue Doppler imaging (TDI) from parasternal long axis and apical views, peak velocities and peak myocardial velocity differences across the right ventricular anterior wall, interventricular septum and left ventricular posterior wall were assessed during systole, early and late diastole. The existence of transmyocardial velocity differences between the left and right side of the interventricular septum, as well as between the endocardium and epicardium of the left ventricular posterior wall was observed throughout the heart cycle. With range-gated TDI from apical four-chamber view, peak velocities were measured within the basal, mid and apical parts of the interventricular septum, and the left and right free ventricular walls. The highest peak systolic, early and late diastolic velocities were measured within the basal parts of all myocardial walls. The ranges of the calculated velocity ratios (early-to-late diastolic velocity and early diastolic-to-systolic velocity) for the various wall parts appeared to be overlapping. The correlations of peak myocardial tissue velocities and their ratios with age and weight were weak and practically irrelevant. These normal values of peak myocardial velocities, transmyocardial velocity differences and the ratios of peak wall velocities can be used as reference values in future investigations of ventricular dysfunction in this age group. PMID- 10722913 TI - The effects of temporal bone on transcranial Doppler ultrasound beam shape. AB - A knowledge of beam shape is desirable for many Doppler ultrasound applications, and is especially important for transcranial Doppler recordings where the beam may undergo significant distortion by the skull. Although it may not be possible to determine the precise beam shape for individual cases due to variations in the physical characteristics of the media in the beam path, information about the range of beam shapes that are likely to arise for in vivo recordings from the middle cerebral artery may in future allow limits of uncertainty to be derived, and may even allow partial correction in some cases. In order to assess the potential intersubject variation in beam sensitivity across the middle cerebral artery, the beam shapes generated by four commercial Transcranial Doppler transducers and the effects on beam shape of four different samples of temporal bone were investigated. The results demonstrate that there seems to be relatively little difference in the beam shapes of commercial transducers, but that the distortion effects of temporal bone are variable and unpredictable. PMID- 10722914 TI - The effects of beam shape on the ability to predict changes in vessel size from Doppler signal power. AB - Theory suggests that, under certain ideal circumstances, the power of a Doppler signal is proportional to the size of the vessel from which it is recorded and can, therefore, be used to assess the scale of any changes in vessel size that occur during clinical recordings of cerebral blood flow. However, the relationship between signal power and vessel size depends on the intensity of the incident beam being uniform across the vessel. This is unlikely to be the case for cerebral vessels insonated by commercial transducers. A model was used to estimate the signal power received from vessels insonated by a beam passing through a homogeneous medium, and also by beams passing through each of five samples of temporal bone. In each case, the effects of initial vessel size and various changes in cross-sectional area were investigated for different vessel positions in the beam. The results for the beam paths through bone predict that the power change arising from a change in vessel cross-sectional area is between 5% and 75% smaller than that occurring in a uniform beam. If these results are representative of those arising for an in vivo change in middle cerebral artery size, then the potential magnitude of the error illustrates the caution that may need to be applied when interpreting changes in Doppler signal power. PMID- 10722915 TI - Doppler flow measurement using surface integration of velocity vectors (SIVV): in vitro validation. AB - Blood flow measurement using an improved surface integration of velocity vectors (SIVV) technique was tested in in vitro phantoms. SIVV was compared with true flow (12-116 mL/s) in a steady-state model using two angles of insonation (45 degrees and 60 degrees ) and two vessel sizes (internal diameter = 11 and 19 mm). Repeatability of the method was tested at various flow rates for each angle of insonation and vessel. In a univentricular pulsatile model, SIVV flow measured at the mitral inlet was compared to true flow (29-61 mL/s). Correlation was excellent for the 19-mm vessel (r(2)= 0.99). There was a systematic bias but close limits of agreement (mean +/- 2 SD = -24.1% +/- 7.6% at 45 degrees; +16.4% +/- 11.0% at 60 degrees ). Using the 11-mm vessel, a quadratic relationship was demonstrated between between SIVV and true flow (r(2) = 0.98-0.99), regardless of the angle of insonation. In the pulsatile system, good agreement and correlation were shown (r(2) = 0.94, mean +/- 2 SD = -4.7 +/- 10.1%). The coefficients of variation for repeated SIVV measurements ranged from 0.9% to 10.3%. This method demonstrates precision and repeatability, and is potentially useful for clinical measurements. PMID- 10722917 TI - An early vision-based snake model for ultrasound image segmentation. AB - Due to the speckles and the ill-defined edges of the object of interest, the classic image-segmentation techniques are usually ineffective in segmenting ultrasound (US) images. In this paper, we present a new algorithm for segmenting general US images that is composed of two major techniques; namely, the early vision model and the discrete-snake model. By simulating human early vision, the early-vision model can capture both grey-scale and textural edges while the speckle noise is suppressed. By performing deformation only on the peaks of the distance map, the discrete-snake model promises better noise immunity and more accurate convergence. Moreover, the constraint for most conventional snake models that the initial contour needs to be located very close to the actual boundary has been relaxed substantially. The performance of the proposed snake model has been shown to be comparable to manual delineation and superior to that of the gradient vector flow (GVF) snake model. PMID- 10722916 TI - Reconstruction and visualization of irregularly sampled three- and four dimensional ultrasound data for cerebrovascular applications. AB - Although recent studies have demonstrated the potential value of compounded data for improvement in signal-to-noise ratio and speckle contrast for three dimensional (3-D) ultrasonography, clinical applications are lacking. We investigated the potential of six degrees-of-freedom (6-DOF) scanhead position and orientation measurement (POM) devices for registration of in vivo multiplanar, irregularly sampled ultrasound (US) images to a regular 3-D volume space. The results demonstrate that accurate spatial and temporal registration of four-dimensional (4-D) US data can be achieved using a 6-DOF scanhead tracking system. For reconstruction of arbitrary, irregularly sampled US data, we introduce a technique based upon a weighted, ellipsoid Gaussian convolution kernel. Volume renderings of 3-D and 4-D compounded in vivo US data are presented. The results, although restricted to the field of cerebrovascular disease, will be of value to other applications of 3-D sonography, particularly those in which compounding of data through irregular sampling may provide superior information on tissue or vessel structure. PMID- 10722918 TI - Semiautomatic contour detection in ultrasound M-mode images. AB - We have developed a method for semiautomatic contour detection in M-mode images. The method combines tissue Doppler and grey-scale data. It was used to detect: 1. the left endocardium of the septum, the endocardium and epicardium of the posterior wall in 16 left ventricular short-axis M-modes, and 2. the mitral ring in 38 anatomical M-modes extracted pair-wise in 19 apical four-chamber cine-loops (healthy subjects). We validated the results by comparing the computer-generated contours with contours manually outlined by four echocardiographers. For all boundaries, the average distance between the computer-generated contours and the manual outlines was smaller than the average distance between the manual outlines. We also calculated left ventricular wall thickness and diameter at end diastole and end-systole and lateral and septal mitral ring excursions, and found, on average, clinically negligible differences between the computer generated indices and the same indices based on manual outlines (0.8-1.8 mm). The results were also within published normal values. In conclusion, this initial study showed that it was feasible in a robust and efficient manner to detect continuous wall boundaries in M-mode images so that tracings of left ventricular wall thickness, diameter and long axis could be derived. PMID- 10722919 TI - Segmentation of 3D intravascular ultrasonic images based on a random field model. AB - Segmentation of intravascular ultrasound images provides important information about the degree of vessel obstruction as well as about the shape and size of plaques. To address the problems of inter- and intra-observer variances associated with conventional manual tracing, a fully automated segmentation was developed. The algorithm is based on the optimisation of a maximum a posteriori estimator, implementing the Rayleigh distribution of speckle and a priori information about the contours. Within 3D image sets, additional information by the blood flow resulting in a decorrelation of the pixels within the luminal boundary is used to initialise the segmentation. To accelerate the estimation, dynamic programming was used. The segmentation algorithm was realised as a Windows 95 application on a Pentium II/233 MHz and delivered reliable and reproducible results independent of the catheter position and the total image brightness (except overflow). In contrast, contours drawn by two physicians for an evaluation of 29 clinical cases showed large intra- and inter-observer variances. In vivo images were acquired with a 20 MHz transducer array (EndoSonics InVision). Comparison with the contours drawn by the physicians and histology demonstrates the potential of the segmentation algorithm. PMID- 10722920 TI - The influence of ultrasound frequency and gas-body composition on the contrast agent-mediated enhancement of vascular bioeffects in mouse intestine. AB - The induction by ultrasound (US) of petechiae and hemorrhages in mouse intestine was examined with injection of gas body-based contrast agents. Production of petechiae in the intestinal wall was enhanced by contrast agents for both continuous and pulsed (10 micros pulses repeated at 1 kHz) exposure relative to a gas body-free blank. For pulsed exposure with 10 mL/kg of Albunex, apparent thresholds for peak negative pressure amplitude were 0.42 MPa at 0.4 MHz, 0.85 MPa at 1.09 MHz and 2.3 MPa at 2.4 MHz. Results at these frequencies were the same for 10-11 cycle pulses with fixed duty cycle (0.01). Thresholds for hemorrhage into the intestinal lumen were not appreciably enhanced by added Albunex, and appear to be compatible with previously reported lithotripsy data when duty factor differences are considered. The agents PESDA, Optison and Levovist had lower thresholds (for example, 1.8 MPa for Levovist) than Albunex at 2.3 MHz, and yielded more petechiae. The thresholds for petechiae induction by US with contrast agents encroach upon the exposure range relevant to diagnostic US practice. PMID- 10722921 TI - Direct ultrasound application had no effect on cardiac hemodynamic performance in a baseline isolated rat heart model. AB - Therapeutic ultrasound (US) has been used for more than 3 decades to promote tissue healing in cases of tissue injury and muscle soreness. It was previously suggested that US may have vasorelaxatory and inotropic properties. However, the direct effect of therapeutic US in a whole heart model has not yet been investigated. Our hypothesis was that application of US might enhance cardiac function. The Langendorf model was modified in a special manner to allow application of US to the heart. Using this model, 20 male rats were equally divided into two groups. Group 1: the hearts were perfused for 15 min, to obtain baseline measurements, and then they were perfused for another 15 min in a special bath full of perfusate. Group 2: after 15 min of baseline measurements, continuous US of 1 MHz 2 W/cm(2) was applied for another 15 min. The parameters that were measured at 5-min intervals were: left ventricular pressure P(max), first derivative of the rise and fall in left ventricular pressure (dP/dt(max), dP/dt(min)), and pressure-time integral. There was no significant difference between the two groups in all parameters at baseline and during US application. P(max) and dP/dt(max) remained constant. After 15 min of US propagation, P(max) was 98% +/- 3 from baseline level vs. 98% +/- 7 in the control group, and dP/dt(max) was 98% +/- 3 vs. 99% +/- 9 in the control. In dP/dt(min), a gradual decline after 15 min of perfusion was measured. In the US- treated group, it declined to 80% +/- 10 vs. 83% +/- 5 in the controls. In conclusion, US radiation at the dose specified does not improve healthy isolated heart hemodynamic performance. We established a model that may be used for further investigation. PMID- 10722922 TI - Measurement of acoustic streaming using magnetic resonance. AB - Magnetic resonance imaging (MRI) has been used to explore acoustic streaming caused in water under ultrasonic exposure conditions similar to those used for diagnostic applications. Streaming was established in an enclosed tube with acoustically transparent end windows, using a pulsed, weakly-focused transducer of acoustic frequency 3.5 MHz. Phase-detection MRI was used to image and quantify streaming profiles in the region of the acoustic focus. Acoustic powers in the range 0.4 mW to 100 mW were used. The sensitivity of the technique enabled streaming velocities down to 0. 1 mm s(-1) to be measured, generated by acoustic power less than 1 mW. In addition, acoustic streaming generated within open meshes with minimum pore dimensions of 3.0 mm and 2.0 mm was measured. The flow velocity in the coarser mesh reached 0.9 mm s(-1) at 95 mW total acoustic power. These observations demonstrate that acoustic streaming is probably a much more general phenomenon in diagnostic ultrasound (ultrasound) than previously recognised. The combination of magnetic resonance and ultrasound shows promise as a diagnostic method for the differentiation of cystic lesions in vivo, and for their characterisation, with sensitivity significantly greater than using ultrasound alone. PMID- 10722923 TI - Schlieren photography study of energy absorption by uric acid nuclei. AB - Previous studies using microfocus x-ray radiography on concentric laminated uric acid calculi following in vitro extracorporeal shock-wave lithotripsy (ESWL) has demonstrated the enlargement of the matrix layers. The Schlieren technique was used to verify the hypothesis that the incident energy would be concentrated in the matrix layers by total internal reflection. Because of the rough surface of the stone, the ultrasonic beam (4 MHz) was incident at various angles. At the critical Rayleigh angle (45.9 degrees +/- 0.5 degrees ), the reflected beam consisted of a spectacularly reflected lobe and a reradiated leaky Rayleigh wave. Different critical Lamb angles, theta(L), of 30.8 degrees, 38.1 degrees and 50.8 degrees (experimental uncertainty 0.5 degrees ) were also determined. Depending on the angle of incidence on the stone surface, a Rayleigh wave can be produced and/or one or more layers, as a whole, are set into vibration (Lamb excitation), with the result that shell-like fragments consisting of one or more layers, break off the stones at sites of weaker bonding, as has been noted in previous in vitro lithotripsy experiments by our group. PMID- 10722924 TI - Power Doppler-derived speckle tracking image of intraventricular flow in patients with anterior myocardial infarction: correlation with left ventricular thrombosis. AB - The abnormal spatial distribution of intraventricular flow is superior to clinical and two-dimensional (2-D) echocardiographic variables in predicting left ventricular thrombosis after myocardial infarction. Echocardiography was prospectively performed in 79 patients within 72 h after anterior wall myocardial infarction onset and repeated before discharge. The apical rotating flow pattern in color flow map was recognized as abnormal. By power Doppler echocardiography, the moving blood could generate speckle tracking images to delineate the intraventricular flow. A swirling flow pattern indicating the compartmentalization of left ventricular blood flow with some blood stagnant in the apical dyssynergic area was identified. The flow pattern shown by the speckle tracking image was superior to the color-flow map in correlating with left ventricular thrombosis. It implicated that the more the detail in which we can describe the blood flow pathway, the more information we can realize. PMID- 10722925 TI - The experimental investigation of ultrasonic properties for a sonicated contrast agent and its application in biomedicine. AB - The ultrasonic properties of a promising ultrasound (US) contrast agent, named SDA (sonicated dextrose albumin) are reported in this paper. SDA is a suspension of stable microencapsulated gas bubbles with average diameter 2.0 microm prepared from sonicated dextrose albumin. The ultrasonic linear and nonlinear parameters, such as acoustic velocity, sound attenuation and acoustic nonlinearity parameter B/A of SDA, as a function of its bubble concentration from 1.0 x 10(7) to 2.05 x 10(8) microbubbles/mL in the frequency range of 2-6 MHz are measured in vitro. The sound attenuation coefficients over 2-6 MHz are linearly proportional to the bubble concentration and frequency. It is important to point out that the acoustic nonlinearity parameter B/A for SDA has a very large value that nonlinearly increases with the increase of bubble concentration. PMID- 10722926 TI - The PCAF acetylase complex as a potential tumor suppressor. PMID- 10722927 TI - The transcription factor NF-kappaB: control of oncogenesis and cancer therapy resistance. AB - Discovered in 1986 as a DNA binding activity that recognized the immunoglobulin light chain intronic enhancer, NF-kappaB has been studied intensively for its role in controlling expression of genes involved in immune and inflammatory function. However, more recently, NF-kappaB has been implicated in controlling cell growth and oncogenesis. The link between NF-kappaB and cancer stems, in part, from the fact that this transcription factor is capable of inducing gene products that control proliferative responses and that suppress apoptotic cascades, such as those induced by tumor necrosis factor (TNF), expression of oncoproteins, and genotoxic stress. This latter observation is likely to be important in developing new approaches aimed at improving the efficacy of cancer chemotherapy. PMID- 10722928 TI - From apoptosis to angiogenesis: new insights into the roles of nuclear orphan receptors, chicken ovalbumin upstream promoter-transcription factors, during development. PMID- 10722929 TI - Cyclooxygenase-2 and carcinogenesis. AB - Numerous investigations have shown that COX-2 is a participant in the pathway of colon carcinogenesis, especially when mutation of the APC tumor suppressor is the initiating event. Moreover, it seems that the amount of COX-2 is important, since there is a correlation between its level of expression and the size of the tumors and their propensity to invade underlying tissue [40]. Inhibiting COX-2 at an early stage blocks the development of malignant tumors, causes pre-malignant tumors to regress and may improve the outcome once the cancer is completely established. This set of findings seems to link very strongly with the traditional observation that chronic inflammation is a precursor to a variety of types of cancer. By this formulation, inflammatory stimuli increase COX-2 and the downstream events that it induces promote tumor formation. All of these finding suggest that existing NSAIDs will be useful for the prophylaxis of colon cancer and polyps and we eagerly await clinical investigations that will generate guidelines that suggest those individuals that are the most appropriate recipients for such therapy. Although this field has progressed rapidly in the last few years, many important questions remain. PMID- 10722930 TI - Intercellular invasion and the organizational stability of tissues: a role for fibronectin. AB - Intracellular invasion is the movement of cells of one type into the fabric of other, contiguous tissues. Invasion is a signature behavior of the malignant tumor and also is found as part of the normal behavior of inflammatory blood cells and tissues engaged in the morphogenetic movements of normal embryogenesis and in a number of instances of normal and pathological tissue remodeling in the adult. Informed by the view that the underlying mechanisms of invasion will be similar for tumor cells and invasive blood and embryonic cells, this review adopts a comparative approach to the analysis of invasion. Invasion results in the development of a diffuse interface between contiguous tissues. Its alternative is the maintenance of stable, planar tissue boundaries. This is the more usual condition for contiguous tissues in the animal. This review will focus on the processes that, on the one hand, stabilize planar contact interfaces between tissues, and, on the other, promote the destabilization of tissue integrity by fostering intercellular invasion. Particular attention is devoted to a role for adhesive interactions mediated by the matrix adhesion molecule, fibronectin. In certain instances, fibronectin in the matrix promotes invasion whereas in others, the presence of fibronectin prevents invasion. The distinction appears to depend on whether the invasive tissue is migrating into an acellular extracellular matrix or whether invasion involves densely cellular tissues. In the first instance, fibronectin promotes invasion, whereas in the second, it stabilizes the interface of the contacting tissues and prevents invasion. PMID- 10722931 TI - ISREC conference 'Cancer and the Cell Cycle'. 26-30 January 1999, Lausanne, Switzerland. PMID- 10722932 TI - EMBL Mouse Molecular Genetics Meeting, Heidelberg, 1-5 September, 1999. PMID- 10722933 TI - The integrative nature of biochemistry: challenges of biochemical education in the USA. AB - The intricate interplay of Biochemistry with well-established disciplines often blurs the identity of the subject. The issues are many. What is biochemical education? Who should be educated? What should be taught? What should the requirements be for Biochemistry major? What is a career in Biochemistry? The curriculum, course syllabus and application of Biochemistry are ever-evolving concerns. The challenges are particularly keen in the USA due to the diversity in its teaching modes, and in the composition of the student body. The constant changes in technologies also shift the needs of skill and knowledge of the graduates. This presentation is to examine the Biochemistry degree programs in the USA, particularly the curricula of private and public research universities, contrasting them with those of the liberal arts colleges. The goal is to sense the trends of changes, probing how the challenges are met, and to solicit and formulate recommendations as we approach a new millennium. PMID- 10722934 TI - Journal club as a supplement to the undergraduate biochemistry laboratory. AB - After purification of lysozyme, our biochemistry students write a research proposal that outlines a strategy for studying this enzyme after alteration by site-directed mutagenesis. Despite a literature search that yielded a wealth of background information, students were often overwhelmed by the assignment because they were not familiar with advanced techniques of protein analysis. We therefore developed a series of journal clubs in which teams of students present methods and data found in papers dealing with lysozyme. The five topics for journal clubs include; substrate binding and mechanism; spectroscopic techniques; stability analysis; two-dimensional NMR; and X-ray crystallography. After the adoption of the group talks, the quality of the research proposals improved immensely and students found the assignment to be an educationally rewarding exercise. PMID- 10722935 TI - Strategies for building criticism skills in undergraduate biochemists. AB - This paper describes an easy method for empowering biochemistry students to think critically about research data. The technique encourages students to formulate their own opinions by removing the usually dominant 'expert' commentary. The exercise can be done individually or in very large groups, requires no specialist materials and confirmation that articulation and criticism skills have been learnt can be assessed under standard examination conditions. The skills learnt are not discipline specific and, as well as learning how best to read research papers, students also learn something about the research process. PMID- 10722936 TI - How do proteins fold? AB - This article emphasizes the importance of getting students to understand the ways in which polypeptides fold to form protein molecules with complex higher-ordered structures. Modern views on how this folding occurs in vitro and in the cell are summarized and set within an appropriate biological context. PMID- 10722937 TI - Does the transport of oxaloacetate across the inner mitochondrial membrane during gluconeogenesis require carrier proteins other than those used in the malate aspartate shuttle? AB - When authors of general biochemistry textbooks mention carrier proteins involved in the transport of oxaloacetate across the inner mitochondrial membrane for gluconeogenesis, they only make use of the two transporters involved in the malate-aspartate shuttle. As a result of only using the malate-2-oxoglutarate and the glutamate-aspartate carrier proteins, I show that the reaction describing the overall process is unsatisfactory since, in addition to oxaloacetate being transported from the mitochondrial matrix to the cytosol, 2-oxoglutarate is also transported in the reverse direction. I therefore point out that, if only oxaloacetate is to be transported from the mitochondrial matrix to the cytosol, then it is necessary to also make use of other carrier proteins in the inner mitochondrial membrane, namely, the dicarboxylate transporter and the phosphate transporter. PMID- 10722938 TI - Integration of post-graduate studies with corporately funded biotechnology programs. AB - Post graduate research students need to undertake programs that will develop their potential as a scientist. They need to acquire skills to develop scientific argument, design an experimental approach to test a hypothesis, obtain and analyse data, and effectively communicate their ideas and findings to the scientific community. Many biotechnology-based programs are now funded by industry partnerships and integrating post graduate students into these programs require special considerations. PMID- 10722939 TI - A suggested module detailing biopharmaceuticals suitable for inclusion in undergraduate applied biochemistry/biochemistry programs. AB - A 26 lecture module is outlined which details the biochemistry and biotechnology of biopharmaceutical substances. It is designed to equip students undertaking programs in applied biochemistry/biochemistry with an understanding of concepts, both academic and applied, directly relevant to working in the biopharmaceutical sector. In addition to the syllabus, a bank of relevant resource material is provided. PMID- 10722940 TI - Interrelationship between oxidative damage and antioxidant enzyme activities: an easy and rapid experimental approach. AB - This article describes a method for determining some antioxidant enzyme activities (catalase and/or glutathione peroxidase) and the oxidative status (protein oxidative damage and/or lipid peroxidation) of human blood. However, the main objective of the work is to illustrate the relationship between antioxidant defences and oxidative damage, showing to students their correlation and the general importance of the biochemical regulation in health and diseases. PMID- 10722942 TI - A guided experimental approach to practical molecular pharmacology teaching: demonstration of sequence selectivity of DNA-binding drugs by arrested polymerase chain reaction. AB - Here, we report a simple experimental approach for demonstrating sequence selectivity of DNA-binding drugs by using the polymerase-chain reaction (PCR). The experiments described could be easily and reproducibly performed by medical and science students and biochemistry teachers, do not require complex or expensive instruments and 32P labelled probes or primers. The procedure described could represent a novel approach to practical biochemistry teaching in the field of biochemical pharma-cology. PMID- 10722941 TI - Experimental treatment of the laws of heterogeneous catalysis with immobilized yeast cells (Saccharomyces cerevisiae). AB - A series of simple, low-cost experiments is described in this paper that allows students to be introduced to some basic kinetic laws relating to heterogeneous catalysis. Immobilized yeast cells are used as the example and therefore simultaneously offer the opportunity to acquaint the students with the theoretical and practical background of an important branch of biotechnology. PMID- 10722943 TI - The excellence of turnip mitochondrial fraction. AB - Several mitochondrial fractions were screened for suitability in practical experiments designed for students, with the following issues in mind: avoiding the use of animals; minimal expenditure of labour and time; high enzyme activities; accessible instruments and low-cost materials. Turnips and potato tubers were identified as the best materials from which to extract purified mitochondria according to these criteria, with high respiratory activities and integrity maintained during five consecutive days. Mitochondrial respiration was assayed for succinate, exogenous NADH, malate/pyruvate and alpha-ketoglutarate oxidations with excellent results. At the fifth day, the respiratory control was still about 3, the integrity of the outer mitochondrial membrane maintained at values higher than 80%, and the enzymes retained more than 55% of the initial activities. PMID- 10722944 TI - A single protein research integrated advanced biochemistry laboratory course; spectroscopic determination of tyrosyl side chain pKa. AB - The unique bio-analytical properties of the amino acid tyrosine (Tyr) are the focus of this experiment from the research oriented biochemistry laboratory course at our university. In the present study pK(a(1)), pK(a(2)), and pK(a(3)) values for free Tyr were estimated to be 2.30, 9.40, and 9.97, respectively, when free Tyr was titrated with 1mM NaOH and 1mM HCl using a pH meter. Spectrophotometric analysis of the phenolic side chain pK(a(3)) revealed a value of 10.14, which was consistent with the pK(a)s estimated from the pH meter. The results from this experiment will allow students to compare the free Tyr properties with those present in a protein. PMID- 10722945 TI - An experiment on apoptosis induced by polyamine adducts produced in the presence of serum amine oxidase. AB - A two-session experiment is proposed to train students with an easy, economic and educative procedure to detect the typical DNA ladder produced in many apoptotic events. The procedure is accurate enough to provide an easy way to compare degrees of damage in DNA caused by different treatments. PMID- 10722946 TI - Stability assessment of ketoconazole in aqueous formulations. AB - Ketoconazole is an imidazole antifungal agent. It has a wide antifungal spectrum and possesses some antibacterial activity. In inappropriate formulations, especially in aqueous media, ketoconazole molecules may be unsteady. The stability of ketoconazole in aqueous media was assessed as a function of pH, antioxidant and ketoconazole concentrations. It was found that ketoconazole was least stable at pH 1 among the pH values studied (pH 1-9). Since the major degradation pathway was specific acid catalysis, based upon the transition-state theory, the entropy (DeltaS) of the activation was calculated and found to be negative indicating that the activated complex was more constrained than the individual species. The free energy of activation (DeltaG) was estimated to be 30 kcal mol(-1). The viscosity of the formulation was found to be more stable at high pH because carbopol is stable at basic pH and protected ketoconazole. It appears that the amount of ketoconazole in the formulation has a low influence on the degradation mechanisms. The increase of the butylated hydroxytoluene antioxidant levels from 0.05 to 0.4%, adversely affected the stability of ketoconazole. In conclusion, the expected shelf life of the final ketoconazole formulation (pH 7, 0.1% butylated hydroxytoluene) was 15 months. PMID- 10722947 TI - Nonionic oil-in-water microemulsions: the effect of oil type on phase behaviour. AB - The formation of oil-in-water (o/w) microemulsions stabilized by the nonionic surfactants, polyoxyethylene-10-dodecyl ether, polyoxyethylene-10-oleyl ether, N,N-dimethyldodecylamine-N-oxide and N,N-dimethyloleylamine-N-oxide and containing a variety of pharmaceutically acceptable oils, namely ethyl butyrate, ethyl caprylate, ethyl oleate and the triglycerides, soybean oil, Miglyol 812 and tributyrin, has been examined at 298 K. The effect on microemulsion formation of replacing water with phosphate buffered saline (PBS) and complete PBS has been established. In addition, the effect of changing temperature (from 298 to 310 K) on the phase behaviour of microemulsions formulated using PBS as continuous phase has been determined. Although some small differences in phase behaviour were noted when altering the continuous phase, the greatest difference in phase behaviour was observed when changing the experimental temperature, particularly for microemulsions stabilized by polyoxyethylene-10-oleyl ether. Regardless of the temperature and aqueous phase used, however the larger molecular volume oils (soybean oil, Miglyol 812 and ethyl oleate) were solubilized to a lower extent than the smaller molecular volume oils (namely, ethyl butyrate and ethyl caprylate). The only exception to this rule was when polyoxyethylene-10-oleyl ether was used as surfactant, particularly at 298 K, where it was the larger molecular volume oils that were solubilized to the greatest extent. Cloud point/phase inversion temperature experiments suggested that the higher molecular volume oils were incorporated into the microemulsions prepared using the polyoxyethylene-based surfactants in a different way than the smaller molecular volume oils and suggest that the smaller molecular volume oils are acting in much the same way as a cosurfactant in that they interchelate with their hydrophilic group interspersed in the surfactant head group region. As N,N dimethyldodecylamine-N-oxide does not exhibit a cloud point it was not possible to determine the mode of oil incorporation in microemulsions prepared with this surfactant. PMID- 10722948 TI - Prolonged effect of liposomes encapsulating pilocarpine HCl in normal and glaucomatous rabbits. AB - The possibility of using liposomes as an ophthalmic drug delivery carrier for the lipophilic drug, pilocarpine HCl, was investigated on the eyes of normal and glaucomatous pigmented rabbits. The intraocular pressure (IOP) of rabbits was measured, using a Shi∅tz tonometer, as a function of time after topical administration with free drug, neutral and negatively charged multilamellar vesicles (MLVs) encapsulating pilocarpine HCl. The results showed that administration with neutral MLVs displayed the most prolonged effect with respect to negatively charged MLVs and free drug. The efficiency of MLVs encapsulating pilocarpine HCl, measured using spectrophotometric technique, was found to be 96% in our modified preparations. The storage stability of MLVs encapsulating pilocarpine HCl was investigated by measuring phase transition and size distribution using light scattering technique. The results show that liposomes encapsulating pilocarpine HCl have kept their integrity and physicochemical properties for at least 15 months, which makes them suitable for commercial use. PMID- 10722949 TI - Improved kinetic parameter estimation in pH-profile data treatment. AB - Statistical problems in temperature stability parameter estimation have been the subject of many papers whereas statistics in, pH-profile parameter estimation have focused little attention. However, the conventional two step method used in data treatment in both cases leads to identical statistical problems. The aim of this study is then to introduce a method that improves statistics in pH-profile parameter estimation. A one step non-linear method that takes into account the errors in drug content determination is proposed. A mathematical relationship between drug content C, pH and time t is tested. The proposed method allows the estimation of the specific kinetic constants and the dissociation constant (pK(a)) in a single run. The most likely experimental initial drug contents C(0j),. where j is the index of a given experiment, are also determined. This approach that takes into account all relevant experimental information for the estimation of kinetic parameters is more rigorous from a statistical viewpoint than the classical two step methods. Kinetic data from acetylsalicylic acid (ASA) hydrolysis was used for the tests. PMID- 10722950 TI - Effect of process parameters on compressibility of granulation manufactured in a high-shear mixer. AB - Various processing variables that can influence granulation characteristics of a lactose-based formulation were evaluated using a Plackett-Burman experimental design. These parameters were impeller speed, granulating solution addition rate, total amount of water added in the granulation step, wet massing time, moisture content of the granulation after drying, and screen size used for the dry milling. Results showed that granulation growth was enhanced by the increase in the amount of added water, high impeller speed, and short wet massing time. On the other hand, moisture content had the largest impact on granulation compressibility, followed by the wet massing time and impeller speed. Increasing moisture content of the granulation and decreasing wet massing time or impeller speed increased granulation compressibility. Increasing impeller speed and/or wet massing time decreased granule porosity and fragmentation propensity, which led to decreased granulation compressibility. Granulation compressibility was extremely sensitive to processing conditions. Tablets from all runs showed acceptable weight variation and friability, suggesting that the parameters evaluated had little effect on these responses in the ranges tested. PMID- 10722951 TI - Comparison of in vitro and in vivo efficiencies of a novel unit-dose liquid aerosol generator and a pressurized metered dose inhaler. AB - Gamma scintigraphic imaging was employed in 10 healthy volunteers to compare the total and regional lung deposition of aerosols generated by two delivery platforms that permitted microprocessor-controlled actuation at an optimal point during inhalation. An aqueous solution containing 99mTc-DTPA was used to assess the deposition of aerosols delivered by inhalation from two successive unit dosage forms (44 microl volume) using a prototype of a novel liquid aerosol system (AERx Pulmonary Delivery System). This was compared with aerosol deposition after inhalation of two 50 microl puffs of a 99mTc-HMPAO-labeled solution formulation from a pressurized metered dose inhaler (MDI). The in vitro size characteristics of the radiolabeled aerosols were determined by cascade impaction. For the AERx system, the predicted lung delivery efficiency based on the product of emitted dose (60.8%, coefficient of variation (CV)=12%) and fine particle fraction (% by mass of aerosol particles <5.7 microm in diameter) was 53.3% (CV=13%). For the solution MDI, the emitted dose was 62.9% (CV=13%) and the predicted lung dose was 44. 9% (CV=15%). The AERx system demonstrated efficient and reproducible dosing characteristics in vivo. Of the dose loaded into the device, the mean percent reaching the lungs was 53.3% (CV=10%), with only 6. 9% located in the oropharynx/stomach. In contrast, the lung deposition from the solution MDI was significantly less (21.7%) and more variable (CV=31%), with 42.0% of the radiolabel detected in the oropharynx/stomach. Analysis of the regional deposition of the radioaerosol indicated a homogeneous pattern of deposition after delivery from the AERx system. A predominantly central pattern of distribution occurred after MDI delivery, where the pattern of deposition was biased towards a central zone depicting the conducting airways. The AERx system, in contrast to MDIs, seems highly suited to the delivery of systemically active agents via pulmonary administration. PMID- 10722952 TI - Synthesis and bacterial degradation of an azopolymer. AB - Azopolymers were synthesised with differing degrees of hydrophobicity, from 2 hydroxyethylmethacrylate (HEMA), styrene and 2,2'-dimethylacryloyloxyazobenzene as azo crosslinker. Bacterial degradation of the series of polymers was assessed using a pure culture of the colonic organism Enterococcus faecalis and rat caecal contents. Polymer degradation was determined in terms of weight loss on polymer coated glass beads and using scanning electron microscopy after incubation. Similar weight loss occurred on incubation of polymers in both bacterial cultures and non-bacterial control. The presence of styrene was found to decrease the amount of weight loss. The polymer surfaces showed microscopic cracks and holes after incubation, again, this phenomenon was less pronounced with increasing styrene content. As there was no increase in polymer degradation in the presence of azo reducing microorganisms, the results of this study suggest that these polymers are degraded by mechanisms other than azo reduction. PMID- 10722953 TI - Modulating pharmacokinetics of an anti-interleukin-8 F(ab')(2) by amine-specific PEGylation with preserved bioactivity. AB - By covalently attaching biocompatible polyethylene-glycol (PEG) groups to epsilon amino groups of the F(ab')(2) form of a humanized anti-interleukin-8 (anti-IL-8) antibody, we sought to decrease the in vivo clearance rate to give a potentially more clinically acceptable therapeutic. The in vivo clearance was modulated by changing the hydrodynamic size of the PEGylated antibody fragments. To achieve significant increases in the hydrodynamic size with minimal loss in bioactivity, high molecular weight linear or branched PEG molecules were used. Modification involved N-hydroxy-succinamide reaction of the PEGs with primary amines (lysines and/or the N-terminus) of the anti-IL-8 F(ab')(2). The process of adding up to four linear 20 kDa PEG, or up to two branched 40 kDa PEG, gave reproducible distribution of products. The components with uniform size (as assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis) were purified by a single step ion-exchange high-performance liquid chromatography and showed no significant loss of biological activity in ligand binding and cell-based assays. Addition of a single branched 40 kDa PEG to a F(ab')(2) (molecular weight (MW)=1.6 million Da) or up to two 40 kDa branched PEG (MW=1.9 million Da) increased the serum half-life to 48 h as compared with the unPEGylated F(ab')(2) with a half-life of 8.5 h. This study shows that by attaching high molecular weight PEGs at a one or two sites, bioactive antibody fragments can be made reproducibly with sizes tailored to achieve the desired pharmacokinetics. PMID- 10722954 TI - Chitosan citrate as film former: compatibility with water-soluble anionic dyes and drug dissolution from coated tablet. AB - Chitosan citrate solution containing 25% w/w propylene glycol was prepared and tested for its compatibility with some water soluble anionic dyes. The immiscibility between erythrosine, ponceau 4R, sunset yellow or tartrazine solutions and chitosan citrate solution was evident. The Fourier transform infrared spectra revealed charged interaction between anionic dye and chitosan. Brilliant blue and green FS at concentration of 0.02-1.00% w/w polymer could be miscible with chitosan citrate solution due to the decrease in charge interaction by the positive charge on molecule of brilliant blue, which was also the composition in green FS. Propranolol HCl tablets coated with these colored film coating solutions exhibited good appearance and no color migration. Drug dissolution from coated tablets was pH dependent, corresponding to the ability of chitosan to protonate in the medium. Color incorporation slightly retarded drug dissolution in acidic medium. Drug dissolved from coated tablet colored with brilliant blue was faster than from that colored with green FS. This was because brilliant blue had positive charge and more SO(3)H groups on its molecular structure, and exhibited higher water solubility. Accelerated condition could alter dissolution characteristics, and the Td+t(0) value from curve fitting between the dissolution profiles and Weibull equation was increased. However, drug dissolution from freshly prepared coated tablets, coated tablets after exposure to accelerated condition and after storage at room temperature for 12 months conformed to the monograph in USP XXIII. PMID- 10722955 TI - Thermodynamic analysis of compact formation; compaction, unloading, and ejection. I. Design and development of a compaction calorimeter and mechanical and thermal energy determinations of powder compaction. AB - The aim of this investigation was to determine and evaluate the thermodynamic properties, i.e. heat, work, and internal energy change, of the compaction process by developing a 'Compaction Calorimeter'. Compaction of common excipients and acetaminophen was performed by a double-ended, constant-strain tableting waveform utilizing an instrumented 'Compaction Simulator.' A constant-strain waveform provides a specific quantity of applied compaction work. A calorimeter, built around the dies, used a metal oxide thermistor to measure the temperature of the system. A resolution of 0.0001 degrees C with a sampling time of 5 s was used to monitor the temperature. An aluminum die within a plastic insulating die, in conjunction with fiberglass punches, comprised the calorimeter. Mechanical (work) and thermal (heat) calibrations of the elastic punch deformation were performed. An energy correction method was outlined to account for system heat effects and mechanical work of the punches. Compaction simulator transducers measured upper and lower punch forces and displacements. Measurements of the effective heat capacity of the samples were performed utilizing an electrical resistance heater. Specific heat capacities of the samples were determined by differential scanning calorimetry. The calibration techniques were utilized to determine heat, work, and the change in internal energies of powder compaction. Future publications will address the thermodynamic evaluation of the tablet sub processes of unloading and ejection. PMID- 10722956 TI - When antiepileptic drugs aggravate epilepsy. AB - Paradoxically, an antiepileptic drug (AED) may aggravate epilepsy. The number of AEDs is steadily increasing, and the occurrence of paradoxical aggravation will probably become a frequent problem. The overall status of the patient treated for epilepsy can be altered due to maladjustment to the diagnosis of epilepsy, to unwanted side-effects, to overdosage and to the occurrence of tolerance. However, the main mechanism of aggravation is the occurrence of an inverse pharmacodynamic effect. The specific effect of the AED is such that it controls epilepsy in most cases and increases seizures in other cases. Idiopathic generalised epilepsies (IGE) are particularly prone to pharmacodynamic aggravation: typical absences are constantly increased by carbamazepine (CBZ), vigabatrin, tiagabine, gabapentin, while phenytoin (PHT) is less aggravating. Juvenile myoclonic epilepsy is often aggravated by CBZ, less constantly by PHT and other AEDs. Generalised tonic clonic seizures found in IGEs may respond to AEDs that aggravate the other seizure types. In symptomatic generalised epilepsies, patients have often several seizure types that respond differently to AEDs: myoclonias are generally aggravated by the same drugs that aggravated IGEs; tonic seizures in the Lennox Gastaut syndrome respond to CBZ, which may however aggravate atypical absences. In severe myoclonic epilepsy of infancy, there is a nearly constant aggravating effect of lamotrigine. In some patients with benign rolandic epilepsy, a clear aggravation may be produced by CBZ, with occurrence of negative myoclonias, atypical absences, drop attacks, and at the maximum evolution into a state of electrical status epilepticus during sleep. It is much more difficult to pinpoint specific pharmacological sensitivity in other focal epilepsies, but aggravation clearly occurs. When treating epilepsy, the clinician should act according to seizure type, or, better, to epilepsy type. Patients are usually aware of aggravation before their doctors: we should listen carefully whenever they express a 'dislike' for an AED. PMID- 10722957 TI - Medical education and postgraduate training in the USA: special reference to children's hospital. AB - The American medical education from undergraduate to postgraduate education is introduced. All physicians and surgeons require residency training before becoming eligible to be certified by the American Board of Medical Specialty. The residency program is tightly overseen and regulated by the Accreditation Council for Graduate Medical Education (ACGME) and the Residency Review Committee in order to maintain its quality. The American healthcare system with its current trend towards managed care is also presented. Managed care has changed the scope of American medical education. Medicare and Medicaid have been the primary source of funding for graduate medical education (GME). With these governmental health insurance programs now involving managed care, GME funding has been reduced. Uprising of managed care is followed by a strong emphasis to train more primary care physicians than specialists. The current trend in child health care is moving towards decreased hospitalization, increased ambulatory care and increased community-based service. PMID- 10722958 TI - Sleep in subjects with autistic disorder: a neurophysiological and psychological study. AB - Polysomnography (EOG, EEG, EMG) was carried out in 17 male children and adolescents with autistic disorder, in seven patients with mental retardation and fragile X syndrome, and in five age- and sex-matched normal male subjects. Density of rapid eye movements was not significantly different in the three groups of subjects; however, some sleep parameters such as time in bed, sleep period time, and total sleep time were significantly lower in subjects with autistic disorder than in normal controls; moreover, patients with autistic disorder showed values of sleep period time, first REM latency and percent (%) sleep stage 1 lower than those of patients with fragile X syndrome with mental retardation. Density of muscle twitches was significantly higher in patients with autistic disorder than in normal controls. In contrast only minor differences were observed between patients with autistic disorder and those with fragile X syndrome with mental retardation. Furthermore, some psychoeducational profile revised items such as perception and eye-hand coordination, showed significant correlation with some sleep parameters (time in bed, sleep latency, stage shifts, first REM latency and wakefulness after sleep onset). Childhood Autism Rating Scale (CARS) scores to visual response and non-verbal communication showed significant correlation with some tonic sleep parameters, such as sleep period time, wakefulness after sleep onset, and total sleep time. Relating to people and activity level items were found to be significantly correlated with rapid eye movement density. Our results suggest the existence of a sleep pattern in autistic patients different from that observed in subjects with mental retardation and from that of normal controls. In addition, these findings indicate that sleep parameters in these patients are correlated with some psychological indices generally used for the diagnosis of autistic disorder; for this reason, polysomnographies might be useful in the comprehension of the neurophysiological mechanisms underlying this condition. PMID- 10722959 TI - Benign partial epilepsy in infancy and early childhood with vertex spikes and waves during sleep: a new epileptic form. AB - International epilepsy classification includes different epileptic syndromes with favourable outcomes in paediatric age, both partial and generalised. This is true in childhood while no partial benign forms are accepted in infancy. In 1987, Watanabe first described a new entity and he defined it as 'benign complex partial epilepsies in infancy'. In 1992, Vigevano referred similar but familial cases whose seizures had secondary generalisation. Both these forms had no interictal EEG abnormalities neither awake nor during sleep. This article presents a survey of 12 cases of partial epilepsy with favourable outcome differing from Watanabe and Vigevano's cases, both for the presence of interictal EEG abnormalities only during sleep and for seizure picture. All our patients are neurologically and neuroradiologically normal. Psychomotor development is unremarkable. Age onset range is 13-30 months. All cases present characteristic spikes and waves during slow-sleep in vertex cerebral areas. Awake EEG is always normal, at follow-up too. Our cases have such homogeneous electroclinical features as to hypothesise a new partial idiopathic epileptic syndrome with favourable outcome in infancy and early childhood. We propose to define it as 'benign partial epilepsy in infancy and early childhood with vertex spikes and waves' (BVSE). PMID- 10722960 TI - Head stability during whole body movements in spastic diplegia. AB - Head angular stability is essential for postural control in whole body movement. Using the opto-electronic ELITE system, we have studied head orientation during the movements of squatting from the standing position and straightening-up from the squatting position in 12 children with spastic diplegia and 12 age-matched controls. Although no instruction was given regarding the head, diplegic children consistently performed excessive neck flexion in the squatting movement and excessive hyperextension in the straightening-up movement, whereas normal children maintained the initial orientation throughout both movements. We discuss pathophysiological implications. PMID- 10722961 TI - Epilepsy in adolescents and young adults with autistic disorder. AB - Since the first description by Kanner (1943) the association between autistic disorder (AD) and epilepsy has been observed in 4-42% of patients. Some authors reported that seizures prevailed in adolescence but a systematic investigation has never been undertaken. We examined retrospectively 60 patients divided into two groups (with and without epilepsy and EEG paroxysmal abnormalities) with AD unrelated to a congenital or acquired encephalopathy (mean age 17 years 2 months). The aim was to investigate epilepsy, EEG paroxysmal abnormalities and possible etiological factors. The prevalence of epilepsy was 38.3%, much higher than that in a normal population of a similar age (6.6 per thousand). The prevalence of EEG paroxysmal abnormalities without epilepsy was 6.7%, higher than that in a population of adolescents and adults with psychiatric pathologies (2. 6%). Seizure onset was after age 12 years in 66.7% of cases. The most common type of epilepsy was partial in 65.2% and four patients (17.4%) had a benign childhood epilepsy with centro-temporal spikes. At the last observation 44.4% of patients had been seizure-free for 2 years or more. There were no organic factors influencing the development of epilepsy but familial and personal antecedents, mental retardation and CT scan/MRI data may suggest an early brain dysfunction responsible for AD and epilepsy. PMID- 10722962 TI - Complete skipping of exon 66 due to novel mutations of the dystrophin gene was identified in two Japanese families of Duchenne muscular dystrophy with severe mental retardation. AB - Severe mental retardation is a rare complication of Duchenne muscular dystrophy (DMD). Here we report that two DMD cases showing severe mental retardation exhibit the same exon skipping event induced by different intron mutations. In the two Japanese DMD patients studied, the complete sequence of exon 66 of the dystrophin gene was found to be absent from the dystrophin mRNA, creating a premature stop codon in exon 67. Novel point mutations at the consensus sequence of the splice donor site of intron 66 (T9857(+2) to C in one case and G9857(+5) to T in the other case) were found to be the cause of complete exon skipping. Remarkably, severe mental retardation cosegregated with an exon 66-skipping event in their families. Furthermore, pachygyria was disclosed by magnetic resonance imaging (MRI) examination of the brain of one case. Our results suggested that exon 66 skipping should be examined in DMD cases with a severe form of mental retardation. PMID- 10722963 TI - Ethical attitudes of Japanese physicians regarding life-sustaining treatment for children with severe neurological disabilities. AB - Ethical attitudes of Japanese physicians regarding life-sustaining treatment for children with severe neurological disabilities (SND) were investigated by mailing a translated questionnaire which the Child Neurology Society (CNS) of the United States used for their survey. The questionnaire was sent to 202 council members of the Japanese Society of Child Neurology (JSCN), and the answers of 147 respondents (72.8%) were analyzed. It was found that the majority (85. 0%) of respondents believed that the same level of care should be provided to children with SND as those without it. However, fewer respondents (15.6%) believed that cardiopulmonary resuscitation was indicated for children with progressive or degenerative brain disorders. With respect to the authoritative role of medical indications and family/guardian's wishes in clinical decision-making for children with SND, about 30% of respondents believed that medical indications should override family/guardian's wishes. However, almost as many respondents (29.9%) chose an ambivalent answer. If compared with the results of the preceding CNS survey, considerably more respondents gave ambivalent answers (average 26. 6%) than in the CNS survey (5.8%). About half of the respondents (49. 0%) acknowledged the need for ethical guidelines to help physicians make ethically difficult decisions. Although statistical comparison was not possible, there were considerable differences between the results of the current study and those of the CNS survey. PMID- 10722964 TI - Age-related changes in the posterior limb of the internal capsule revealed by magnetic resonance imaging. AB - In this study, we examined the age-related differences in the corticospinal tract (CST) of the posterior limb (PL) of the internal capsule (IC) by analyzing 53 magnetic resonance (MR) images from 45 subjects. Regions in the posterior part of the PL of the IC were observed as hypointense areas in T1-weighted images of axial sections at 4 years of age. These regions became clear, spotty hyperintense areas in T2-weighted images of subjects older than 9 years. These regions persisted in all cases beyond 9 years of age, level on both T1- and T2-weighted images. The region is consistent with the CST from MR images of coronal and sagittal sections in our study. These results suggest that differences among the region may occur, such as an increase in fiber size and an increase in the thickness of the myelin sheaths, depending on age after the completion of general myelination in the CST. PMID- 10722965 TI - Plasmapheresis in a child affected by acute disseminated encephalomyelitis. AB - We describe an eight year old girl with acute relapsing disseminated encephalomyelitis (ADEM) who began to improve concomitantly with plasmapheresis therapy. The patient had previously undergone high-dose intravenous methylprednisolone, intravenous immunoglobulins and Interferon beta-1b treatment which did not control the clinical course of the disease. The long term follow-up suggests that plasmapheresis is effective in this disorder and may give better results than steroids or IVIG. PMID- 10722966 TI - Clinical, fluorine-18 labeled 2-fluoro-2-deoxyglucose positron emission tomography (FDG PET), MRI of the brain and biochemical observations in a patient with 4-hydroxybutyric aciduria; a progressive neurometabolic disease. AB - We report a five-year-old boy with 4-hydroxybutyric aciduria. The child presented with global developmental delay, severe hypotonia and myoclonic seizures. The urine 4-hydroxybutyric acid was 1038 times that of normal, and other organic acids related to its further metabolism were also increased. Electroencephalography showed findings indicative of cerebral dysfunction. However, other neurophysiological studies were normal. Clinical improvement was observed after the administration of vigabatrin and dextromethorphan. Magnetic resonance imaging of the brain revealed cerebellar vermin atrophy and subtle white matter changes in the cerebral hemispheres. Fluorine-18 labeled 2-fluoro-2 deoxyglucose positron emission tomographic (FDG PET) scan of the brain showed a marked decrease in the cerebellar metabolism, probably related to atrophy of cerebellar vermis and secondary cerebellar deafferentation. FDG PET scan is found to be of value in the understanding and assessment of brain functional alterations. It may be useful in monitoring and optimizing treatment strategies of this rare disease. PMID- 10722967 TI - Congenital myotonic dystrophy: report of paternal transmission. AB - A female neonate born to a healthy mother was hospitalized because of enlargement of the lateral ventricles, muscle weakness, irregular respiration, and poor sucking. Characteristic facial appearance such as high forehead and carp mouth were noted. The father had mild manifestations of adult type myotonic dystrophy, including muscle weakness of the extremities, percussion myotonia and atrophy of the facial muscles. PCR analysis and southern blot analysis revealed that CTG repeats in the myotonic dystrophy gene of the infant and the father were about 1000 and 400, respectively. This is a rare case showing paternally transmitted congenital myotonic dystrophy and seems to be the first report describing a neonate. PMID- 10722968 TI - Suppression-burst patterns in intractable epilepsy with focal cortical dysplasia. AB - We report on a patient with early-onset spasms in series and partial seizures associated with focal cortical dysplasia whose EEGs showed suppression-burst patterns during early infancy. These electroclinical characteristics suggested a diagnosis of Ohtahara syndrome, but the EEG findings were atypical because of the lack of suppression-burst patterns during wakefulness. In addition, the patient did not have severe psychomotor retardation. With high-dose pyridoxal phosphate therapy, seizures were suppressed and suppression-burst patterns disappeared at 2 months of age. Focal motor seizures recurred later and they often evolved into epilepsia partialis continua. Patients with early-onset intractable seizures associated with suppression-burst patterns on EEGs have several different etiologies, and these patients should be categorized according to their etiology in addition to their syndromic diagnosis. PMID- 10722969 TI - Applications of (7)Li NMR in biomedicine. AB - The applications of (7)Li NMR spectroscopy and imaging in biology and experimental medicine have been progressing steadily. The interest derives primarily from the clinical use of Li salts to treat mania and manic-depressive illness. One area of investigation is ionic transport across the cellular membrane and compartmentation, so as to elucidate the mechanism(s) of therapeutic action and toxicity in clinical practice. The second is the development of a noninvasive, in vivo analytical tool to measure brain Li concentrations in humans, both as an adjunct to treatment and as a mechanistic probe. Here we review progress to date in this area. PMID- 10722970 TI - Dual-echo breathhold T(2)-weighted fast spin echo MR imaging of liver lesions. AB - The purpose of this study was to develop a multi-shot dual-echo breathhold fast spin echo technique (DFSE) and compare it with conventional spin echo (T2SE) for T(2)-weighted MR imaging of liver lesions. The DFSE acquisition (EffTE1/EffTE2/TR = 66/143/2100 ms) imaged 5 sections per 17 s breathhold. T2SE imaging (TE1/TE2/TR = 60/120/2500 ms) required 16:55 (min:s) for 14 sections. Both techniques used a receive-only phased-array abdominal multicoil and provided 192 x 256 effective resolution. The results showed first and second echo relative DFSE/T2SE contrast values for 27 representative lesions (15 consecutive patients) were 1.08 +/- 0.05 and 1.16 +/- 0.09 (mean +/- STD mean), respectively. Corresponding CNR values were 1.12 +/- 0.09 and 0.97 +/- 0.12. Overall DFSE was comparable-to-superior to T2SE for lesion sizing and image artifact. DFSE lesion detection was inferior to T2SE's in several patient studies because of decreased conspicuity of lesions located near multicoil edges and because of poor breathhold-to-breathhold reproducibility and lack of breathholding. However both DFSE (and T2SE) provided lesion detection rated to be of diagnostic quality for all patient studies. In conclusion, we found that DFSE provides diagnostically useful dual-echo T(2) weighted MR liver images in a greatly decreased acquisition time. PMID- 10722971 TI - MR imaging of radiation osteitis in the sacroiliac joints. AB - The purpose of this study was to analyze magnetic resonance (MR) images of radiation osteitis of sacroiliac joints, retrospectively. Seven patients with radiation osteitis, which was diagnosed by pelvic plain radiographs and CT images, underwent MRI. T(1)-weighted spin echo images and T(2)-weighted fast spin echo images were obtained in all patients. Four patients were examined after gadolinium injection. Major signal changes of radiation osteitis were distributed on the iliac side. T(1)-weighted images showed diffuse low intensity both in sacral and iliac sides. T(2)-weighted images showed very low intensity adjacent to sacroiliac joints, but mixed intensity was illustrated apart from joints, and high intensity in the peripheral areas. Radiation osteitis showed slight to mild, but irregular enhancement in four patients after gadolinium administration. MRI can illustrate abnormal bone change distribution and is useful for diagnosing this entity by characteristic intensity patterns on T(1)-weighted images with and without gadolinium and T(2)-weighted image. However, the diagnosis of accompanied insufficiency fractures in the area of radiation osteitis is occasionally difficult with conventional MRI. PMID- 10722972 TI - T(1) fast acquisition relaxation mapping (T(1)-FARM): optimized data acquisition. AB - A theoretical procedure for estimating the precision of the T(1) Fast Acquisition Relaxation Mapping sequence as a function of a number of acquisition parameters has been validated by both simulations and experimental results. These results have clarified the selection of sequence parameters to give optimal accuracy and precision in the R(1)* measurements. There is excellent agreement between theory, simulation, and experiment except for flip angles greater than 9 degrees, at which point slice profile imperfections significantly degrade the precision of the technique. The experimental results indicate that over a range of T(1)s that would be seen in a bolus tracking experiment (25-1200 ms), T(1) Fast Acquisition Relaxation Mapping can be used to obtain 64 x 128 R(1)* maps at a rate of 1 map/s, with a precision of 10% or better. PMID- 10722973 TI - Activity revealed in MRI of multiple sclerosis without contrast agent. A preliminary report. AB - Disease activity in multiple sclerosis is usually accompanied by blood-brain barrier disruption, which can be assessed with contrast-enhanced magnetic resonance imaging (MRI). This paper describes a technique that gives information about disease activity using magnetization transfer MRI. Image combination methods using follow-up scans, like the one presented here, have the potential to show MS lesions that correlate with enhancement. PMID- 10722974 TI - Peliosis hepatis and neoplastic/dysplastic lesions in aged male Long-Evans Cinnamon rats: MR imaging with pathologic correlation. AB - The Long-Evans Cinnamon (LEC) rat, an animal model of Wilson's disease, abnormally accumulates copper in the liver. There have been a lot of reports on preneoplastic and neoplastic hepatic tumors in LEC rats, but few studies have been focused on other lesions. The aim of this study was to describe the MR findings of the liver of LEC rats with pathologic correlation to characterize the hepatic lesions developed in them. We measured MR images of the liver of six aged (over the age of 70 weeks old) male LEC rats. Measurements of T(1), T(2)-weighted images, and the dynamic and delayed studies after i.v. gadolinium injection were performed. The rats were sacrificed immediately after the measurements, and the diagnosis was histologically made. We identified seven lesions of peliosis hepatis, three neoplastic/dysplastic lesions, three cysts and one cholangiofibrosis. Peliosis hepatis was characterized as showing a significantly long T(2) relaxation time of 57.9 +/- 13.3 ms (mean +/- standard deviation) compared with 41.3 +/- 1.7 ms in normal liver, and prolonged enhancement after a gadolinium injection. Neoplastic/dysplastic lesions tended to show prolonged T(2), and they showed isointensity on T(1)-weighted images. They were best characterized by early enhancement followed by a rapid wash-out after a gadolinium injection. In conclusions, the frequent occurrence of peliosis hepatis observed in the present study suggests this can be a characteristic lesion in aged LEC rats. The characteristic MR findings enable us to distinguish between peliosis hepatis and neoplastic/dysplastic lesions. PMID- 10722975 TI - Ex vivo measurement of tissue distribution of a nitroxide radical after intravenous injection and its in vivo imaging using a rapid scan ESR-CT system. AB - To establish the usefulness of ESR-CT imaging with 3-carbamoyl-2,2,5, 5 tetramethylpyrrolidine-1-oxyl (carbamoyl-PROXYL) in living animals, we investigated the tissue distribution of carbamoyl-PROXYL after i. v. injection. Ten minutes after injection of carbamoyl-PROXYL, its concentrations in the liver, spleen, kidney, and plasma were higher than those in the small intestine and stomach. However, the inter-organ differences in concentrations were not striking. We selected the liver as a representative organ and attempted to measure the concentration of carbamoyl-PROXYL in it after washing out all of the blood by in situ perfusion with saline. The ESR spectrum of the liver homogenate after complete blood washout revealed that the concentration of carbamoyl-PROXYL was significantly reduced. Thus, at this time, carbamoyl-PROXYL was distributed predominantly in the plasma and/or loosely attached to the surfaces of cells. We obtained high-quality ESR-CT images of the murine abdomen at a measurement time of 40 s and found that a high-intensity area of carbamoyl-PROXYL appeared in the liver and kidneys, indicating an abundant blood circulation. Although the organ specificity of carbamoyl-PROXYL was weak, we consider that ESR-CT imaging with carbamoyl-PROXYL will be a powerful new tool for non-invasive anatomic analysis of the liver and the kidneys. PMID- 10722976 TI - Improving image quality and T(1) measurements using saturation recovery turboFLASH with an approximate K-space normalisation filter. AB - We present a method for reducing the image point-spread function and measuring T(1) using saturation recovery turboFLASH (SRTF) with centric-ordered k-space and a k-space correction filter designed to compensate for longitudinal magnetisation evolution during image acquisition. The method provides a two point T(1) measurement that reduces inaccuracies and image artefacts caused by longitudinal magnetisation evolution in conventional turboFLASH methods. The method is designed for use in rapid, quantitative measurements of contrast agent uptake in vivo. PMID- 10722977 TI - A wavelet-based method for improving signal-to-noise ratio and contrast in MR images. AB - Magnetic resonance (MR) images acquired with fast measurement often display poor signal-to-noise ratio (SNR) and contrast. With the advent of high temporal resolution imaging, there is a growing need to remove these noise artifacts. The noise in magnitude MR images is signal-dependent (Rician), whereas most de noising algorithms assume additive Gaussian (white) noise. However, the Rician distribution only looks Gaussian at high SNR. Some recent work by Nowak employs a wavelet-based method for de-noising the square magnitude images, and explicitly takes into account the Rician nature of the noise distribution. In this article, we apply a wavelet de-noising algorithm directly to the complex image obtained as the Fourier transform of the raw k-space two-channel (real and imaginary) data. By retaining the complex image, we are able to de-noise not only magnitude images but also phase images. A multiscale (complex) wavelet-domain Wiener-type filter is derived. The algorithm preserves edges better when the Haar wavelet rather than smoother wavelets, such as those of Daubechies, are used. The algorithm was tested on a simulated image to which various levels of noise were added, on several EPI image sequences, each of different SNR, and on a pair of low SNR MR micro-images acquired using gradient echo and spin echo sequences. For the simulated data, the original image could be well recovered even for high values of noise (SNR approximately 0 dB), suggesting that the present algorithm may provide better recovery of the contrast than Nowak's method. The mean-square error, bias, and variance are computed for the simulated images. Over a range of amounts of added noise, the present method is shown to give smaller bias than when using a soft threshold, and smaller variance than a hard threshold; in general, it provides a better bias-variance balance than either hard or soft threshold methods. For the EPI (MR) images, contrast improvements of up to 8% (for SNR = 33 dB) were found. In general, the improvement in contrast was greater the lower the original SNR, for example, up to 50% contrast improvement for SNR of about 20 dB in micro-imaging. Applications of the algorithm to the segmentation of medical images, to micro-imaging and angiography (where the correct preservation of phase is important for flow encoding to be possible), as well as to de-noising time series of functional MR images, are discussed. PMID- 10722978 TI - A quantitative assessment of liver metabolites during jaundice using three dimensional phosphorus chemical shift imaging. AB - Phosphorus metabolites in the jaundiced rat liver were studied by three dimensional phosphorus chemical shift imaging (CSI). Animals were studied at 1, 2, and 3 weeks post-ligation of the common bile duct. Quantitation of metabolites was performed using an external standard. Metabolite T(1) values were assessed in CSI experiments on normal untreated animals. High-performance liquid chromatography (HPLC) was used to measure adenine nucleotides in a separate group of jaundiced rats. 3D-CSI did not detect significant changes in NTP in jaundiced animals relative to baseline controls. At two and three weeks post bile duct ligation, pH was significantly elevated. HPLC data comparing ATP levels to baseline controls also detected no change except for elevated ATP detected on Day 21. (31)P NMR chemical shift imaging may be used to assess liver metabolites under conditions of stress such as jaundice. However, absolute quantitation requires careful attention to many factors including point spread function, correct T(1) values, and adequate signal-to-noise ratio. PMID- 10722979 TI - An interleaved sampling strategy for MR spectroscopy in vivo: applications on human calf musculature. AB - Assessment of relaxation times, magnetization transfer rates, or apparent diffusion coefficients by volume selective (1)H MR spectroscopy requires data from several single spectra with variable sequence parameters. Unintentional movements during the examination lead to inaccuracies, especially if the spatial distribution of concentrations is inhomogeneous. Improved comparability of the single spectra in a series recorded in vivo were obtained using a modified spectroscopic technique with INTerleaved ACquisiTion of multiple SPECtra (INTACTSPEC). INTACTSPEC series of spectra from the tibialis anterior muscle (m. tib. ant.), soleus muscle (m. soleus), and tibial bone marrow of 20 healthy volunteers were analyzed. Transverse relaxation times T(2) of methylene signals in muscular lipid stores ranged from 77 ms (intramyocellular methylene component in m. tib. ant.) to 88 ms (intramyocellular methylene component in m. soleus) and were similar to those from yellow tibial bone marrow (T(2) = 84 ms). Echo time dependent signal intensities of choline and creatine deviated markedly from a monoexponential behavior in m. tib. ant., but were nearly exponential in m. soleus. Results from water diffusion measurements parallel and perpendicular to the axis of the lower leg showed significant differences between m. tib. ant. and m. soleus, probably due to the spatial orientation of the muscle fibers. Apparent diffusion coefficients along the leg axis were found to be higher in m. tib. ant. (2.10 +/- 0.08 x 10(-3) mm(2)/s) compared to m. soleus (1.78 +/- 0.11 x 10(-3) mm(2)/s), but m. soleus showed less restricted diffusion in perpendicular orientation (1.59 +/- 0.19 x 10(-3) mm(2)/s versus 1.20 +/- 0.08 x 10(-3) mm(2)/s in m. tib. ant.). Magnetization transfer experiments with various RF preparation pulse amplitudes led to very similar results for m. tib. ant. and m. soleus. PMID- 10722980 TI - Visualization of Taylor-Couette and spiral Poiseuille flows using a snapshot FLASH spatial tagging sequence. AB - A new magnetic resonance imaging technique was applied to the Taylor-Couette and spiral Poiseuille (Taylor-Couette with superposed mean axial flux) flows for the first time. The experimental technique is a combination of spatial tagging methods and a snapshot FLASH imaging sequence, which allows the full-field visualization of 2-D slices of the flow field, with image acquisition times approximately half a second. By acquiring images every few seconds, direct visualization of flow patterns can be obtained in the form of cinematography. Tagged images of the Taylor-Couette flow were acquired in both the axial and transverse planes and confirmed previously reported numerical predictions of Taylor cell size. Tagged images of the spiral Poiseuille flows verified that the cells in this flow propagate at a higher velocity than the mean axial flow. In addition, intermittent cell formation was observed as the axial flow was increased. PMID- 10722981 TI - Simultaneous measurement of temperature and velocity maps by inversion recovery tagging method. AB - A new method, called the inversion recovery (IR) tagging method, for simultaneous measurement of temperature and velocity maps of flowing fluid has been developed. The present method employs a set of tagging pulses which acts as an inversion pulse of the conventional IR method, based on the temperature dependence of the spin-lattice relaxation of water proton in a fluid, and has the advantage of being able to compensate the reduction of the NMR signal intensity due to flow motion and to reduce the total time to measure these maps. First, the accuracy of the temperature measurement of stagnant doped water in a differentially heated cell using the conventional IR method, as the basic sequence of the IR tagging method, has been evaluated. The accuracy was within 10% of the temperature difference DeltaT = 17.2 degrees C and the measurable temperature resolution was within +/-0.5 degrees C. Then temperature and velocity maps of the flowing doped water through a cooled pipe were measured simultaneously by the IR tagging method, and the accuracy of temperature measurement was evaluated. The accuracy obtained using the present method was within 15% of the temperature difference DeltaT = 15 degrees C. PMID- 10722983 TI - High-pressure magnetic resonance imaging up to 40 MPa. AB - A high-pressure vessel designed for use with commercial magnetic resonance imaging equipment at up to 40 MPa of pressure was used and tested. Special features of the vessel are the following: 1) 12.6 mm sample chamber i.d.; 2) only non-magnetic parts; 3) visible sample from the outside; 4) resistant to corrosive chemicals, and; 5) sample could be manually translated and rotated in situ. This apparatus was demonstrated through observation of CO(2) clathrate-hydrate growth in a water droplet injected into liquid CO(2) at 20 MPa. PMID- 10722982 TI - Raysum reconstruction algorithm in MR cholangiopancreatography. AB - Magnetic resonance cholangiopancreatography (MRCP) is a new, non-invasive imaging technique for the visualization of the biliary ducts. The presence of stones within the choledocus is easily detectable in source images. However, three dimensional reconstructions using the maximum intensity pixel (or projection) algorithm (MIP) fail to reproduce accurately the eventual presence of filling defects or parietal irregularities due to biliary stones. We used the Raysum algorithm in addition to the MIP in evaluating MRCPs of twelve patients with known choledocolithiasis. A visualization of the stones was obtained in nine (75%) patients by using the Raysum while visualization was obtained in one patient by using MIP. No additional sequences are required, and the post processing time takes only a few seconds. The Raysum reconstruction can be successfully associated to the MIP in the three-dimensional evaluation of biliary stones in MRCP. PMID- 10722984 TI - Adoption of a health education intervention for family members of breast cancer patients. AB - BACKGROUND: Relatives of breast cancer patients often face substantial uncertainty and psychological stress regarding their own health risks and optimal strategies for prevention and early detection. Efficacious educational and counseling interventions are rarely evaluated for their potential adoption and use in medical practice settings. This study evaluates a health education program for first-degree relatives of breast cancer patients based on the program's potential for being adopted and used by medical practices affiliated with cancer centers. METHODS: A randomized, controlled trial was implemented in four community hospital-based medical practices. After 9 months, clinical and administrative staff at each practice were given self-administered surveys. Of 90 staff members recruited to respond, useable responses were received from 60 (67%), including 13 physicians (31%), 43 nurses (98%), and four program managers (100%). Participants made self-reports of program awareness, program support, perceived program performance, likelihood of program adoption and use, and barriers to adoption. RESULTS: A strong majority of respondents (80%) reported that all or most staff agreed with the need for the program. Perceived program performance in meeting goals was generally favorable but varied across sites and across staff types. Overall, 56% of respondents indicated that their practices were likely or highly likely to adopt the program in full. The likelihood of adoption varied substantially across sites and across program components. CONCLUSIONS: Evaluating the potential for program adoption offers insight for tailoring preventive health interventions and their implementation strategies to improve diffusion in the field of practice. PMID- 10722985 TI - The effectiveness of mammography promotion by volunteers in rural communities. AB - INTRODUCTION: The Community Trial of Mammography Promotion assessed the effectiveness of mammography promotion by community volunteer groups in rural areas. Three interventions were tested. One used an individual counseling strategy, one used a community activities strategy, and a third combined the two strategies. METHODS: The effects of the interventions were tested by randomizing 40 communities either to the study interventions or to a control group. A cohort of 352 women from each community was randomly selected and used to evaluate the interventions' effectiveness. Of these, 6592 women were eligible for screening mammography at baseline and follow-up and were successfully interviewed prior to and after study intervention activities. RESULTS: Although the interventions did not significantly increase women's overall use of mammography, the community activities intervention increased use at follow-up by regular users over baseline by 2.9% (p = 0.01). Intervention appears to have increased the use of mammography among certain groups of women who were not regular users at baseline, including those in communities without female physicians (10% to 16%; p < 0.05), and among women with no health insurance (10% to 23%; p 21. Sensitivity and specificity were calculated for three chronic conditions and use of six preventive services. RESULTS: Sensitivity was highest for hypertension (83%), moderate for diabetes (73%), and lowest for hypercholesterolemia (59%); specificity was >80% for all three conditions. Sensitivity ranged from 86% to 99% for influenza immunization, clinical breast examination, blood cholesterol screening, mammography, Pap test, and blood pressure screening; specificity was <75% for all preventive services. CONCLUSIONS: Self-reports are reasonably accurate for certain chronic conditions and for routine screening exams and can provide a useful estimate for broad measures of population prevalence. PMID- 10722988 TI - Clinician satisfaction with a preventive services implementation trial. The IMPROVE project. AB - OBJECT: To discover how attempts to increase the delivery of preventive services affect clinician satisfaction. METHODS: The IMPROVE project was a randomized clinical trial conducted in 44 clinics in and around Minneapolis-St. Paul, Minnesota. Personnel were trained in continuous quality improvement techniques to organize preventive services delivery systems. Satisfaction with delivery of these services and with the sponsoring organizations was measured before the intervention (Time 1), at the end of the intervention (Time 2), and 1 year post intervention (Time 3). RESULTS: At no time was the intervention associated with a change in the respondents satisfaction with their places of work or with their job roles. Satisfaction with preventive services delivery increased from Time 1 to Time 3 among intervention-clinic respondents. Satisfaction with the IMPROVE project and the efforts of the two managed care organizations to help the clinics deliver preventive services peaked at Time 2 and declined toward baseline at Time 3. Satisfaction with preventive services delivery tended to increase more in the 13 intervention clinics that implemented a preventive services delivery system than in the nine intervention clinics that did not implement a preventive services delivery system (p = 0.15). CONCLUSIONS: Planned organizational change to create systems for preventive services delivery can be associated with increased clinician satisfaction with the way these services are delivered. However, increased satisfaction with preventive services does not necessarily indicate that service delivery rates have increased. PMID- 10722989 TI - Incorporating physical activity advice into primary care: physician-delivered advice within the activity counseling trial. AB - INTRODUCTION: The Activity Counseling Trial (ACT) was designed to compare the effectiveness of physician advice alone with physician advice plus behavioral counseling, provided by ACT-trained health educators, to increase levels of physical activity in healthy, sedentary patients. The objective was to determine health care providers' adherence to the ACT protocol for delivering initial "physician" advice on physical activity and to determine providers' satisfaction with the protocol. METHODS: Fifty-four physicians or physician assistants from 11 primary care practices located in California, Texas, and Tennessee volunteered to participate as ACT-trained physicians. Providers were trained to integrate 3 to 4 minutes of initial physical activity advice into the routine office visits of sedentary patients, aged 35 to 75 years, with no acute or serious chronic conditions. This advice included assessment of current physical activities, advising the patient about an appropriate physical activity goal, and referring the patient to the health educator. Providers initialed forms to document delivery of advice, and ACT health educators recorded their advice on a computerized tracking system. A provider survey measured length of time spent advising patients about physical activity and provider satisfaction with the program. RESULTS: Ninety-nine percent of patients received the initial physician advice about physical activity. Eighty-three percent of the providers spent less than 5 to 6 minutes, and 46% spent the recommended 3 to 4 minutes providing advice. Sixty-three percent said the advice resulted in little or no increase in the length of an office visit, and 83% said participation was an asset to their clinics. CONCLUSIONS: Providers incorporated brief physical activity advice into routine primary care visits with little disruption. Their response to the ACT advice protocol was positive and participation in the study was viewed as beneficial. PMID- 10722990 TI - Promoting physical activity in rural communities: walking trail access, use, and effects. AB - INTRODUCTION: Environmental and policy approaches to promote physical activity, such as walking trail construction and promotion, are being widely recommended, yet sparse data exist on their effectiveness. In conjunction with ongoing community-intervention projects in Missouri, walking trails are being built, promoted, and evaluated. Objectives include determining: (1) patterns and correlates of walking, (2) the availability of places to walk and perform other forms of physical activity, (3) the extent of walking trail use and possible effects on rates of physical activity, and (4) attitudes toward the trails and their uses. METHODS: In 12 rural counties in Missouri we used a cross-s ectional telephone survey to ask a population-based sample of residents aged >18 years (n=1269) some standard and specially developed questions about walking behaviors, knowledge, and attitudes. RESULTS: Only 19.5% of respondents were classified as regular walkers. About one third of respondents (36.5%) reported having access to walking trails in their area, and 50.3% reported having access to indoor facilities for exercise. Among persons with access to walking trails, 38.8% had used the trails. Groups who were more likely to have used the walking trails included women, persons with more education, those making $35,000 or more per year, and regular walkers. Among persons who had used the trails, 55.2% reported they had increased their amount of walking since they began using the trail. Women and persons with a high school education or less were more than twice as likely to have increased the amount of walking since they began using the walking trails. CONCLUSIONS: Walking trails may be beneficial in promoting physical activity among segments of the population at highest risk for inactivity, in particular women and persons in lower socioeconomic groups. PMID- 10722991 TI - What students bring to medical school: attitudes toward health promotion and prevention. AB - INTRODUCTION: Health care providers' positive attitudes toward prevention and health promotion are important in achieving national health care goals. Limited studies of incoming medical students have been conducted that measure predictors of positive attitudes toward health promotion and prevention. METHODS: Data were obtained from a 1993 curriculum evaluation survey of first-year students at five different medical schools in California. Attitudes toward health promotion and prevention were measured using a nine-question Prevention Attitude Scale (PAS). We developed 2 multivariate linear regression models using demographics, education choices, and personal social values and beliefs to predict PAS scores. We also performed bivariate analysis. RESULTS: Five hundred ninety-nine completed surveys were analyzed, with a response rate of 95%. Mean PAS score was 36.47 +/- 3.7 on a 0 to 44 scale. Female gender, Democratic party preference, and a planned specialty choice in preventive medicine or primary care predicted the highest PAS scores on bivariate analysis (p < 0.002). Significant correlation ( p < 0. 001) was shown between PAS scores and 2 additional scales regarding beliefs in associations of social factors and illness and in the importance of caring for the poor. Linear regression model using personal social values and beliefs explained 34% of the variance, as opposed to the demographic model that explained only 9% of the variance. CONCLUSIONS: In the schools studied, participating first year medical students had moderately positive attitudes toward health promotion and prevention, as measured by PAS scores. In designing curriculum to improve medical students' attitudes toward health promotion and prevention, medical educators may need to consider other personal and social values held by medical students and to address the "political" aspects of health promotion and prevention. Future studies are needed to longitudinally follow medical student attitudes. PMID- 10722992 TI - Interest in genetic testing among first-degree relatives of colorectal cancer patients. AB - PURPOSE: The present study examined colorectal cancer screening behaviors, risk perceptions, and willingness to receive genetic testing to determine colorectal cancer susceptibility. METHODS: We recruited 95 first-degree relatives of colorectal cancer patients, then conducted a brief telephone interview using a structured questionnaire that elicited information on sociodemographics, cancer screening behaviors, risk perceptions, and interest in genetic testing. RESULTS: Among these high-risk individuals who were aged 40 years or older, only 31% reported fecal occult blood testing within the past year and 59% reported undergoing sigmoidoscopy or colonoscopy within the past 5 years. The majority of participants believed their relative risk of colorectal cancer was increased (68%). Eighty-four percent of the participants indicated that they would have a genetic test if one were available. Participants who believed that <50% of colorectal cancers were caused by heredity were more likely to be interested in genetic testing than were participants who believed that 50% or more of colorectal cancers were caused by heredity. Referral source, sociodemographic factors, clinical factors, and perceived personal risk were not significantly associated with interest in genetic testing. CONCLUSION: Our results suggest that the demand for colorectal cancer susceptibility testing may be high among individuals with a family history of colorectal cancer. We also observed that a substantial number of first-degree relatives were not adhering to colorectal cancer screening guidelines. Accurate information on the genetic aspects of colorectal cancer and the benefits and limitations of genetic testing may help relatives of colorectal cancer patients make informed decisions about whether to undergo enhanced screening and genetic testing. PMID- 10722993 TI - Can compliance with nonpharmacologic treatments for cardiovascular disease be improved? AB - OBJECTIVE: To critically review the literature regarding the effectiveness of interventions aimed at improving cardiovascular patient compliance with nonpharmacologic treatments. METHODS: We searched Medline, Healthplan, and Psychlit from 1985 to 1996; searched the bibliographies of located studies; contacted Australian government departments and nongovernment organizations; and two experts examined the resulting study list. We selected 27 studies, which randomly allocated patients to groups and were published in English, and we evaluated interventions aimed at increasing compliance with nonpharmacologic treatments for cardiovascular disease. These trials were critically appraised against eight methodologic criteria and, subsequently, classified as of good, fair, or poor quality. Information about target groups, samples, trial intervention strategies and their effectiveness were extracted from the 18 good- and fair-quality trials. Interrater reliability was high on the 20% of references that were double-coded. The 18 studies reviewed described the effectiveness of 27 intervention strategies at improving compliance with dietary, smoking-cessation, exercise, weight-loss, stress-reduction, general lifestyle, relaxation, and blood pressure screening programs. RESULTS: Tentative recommendations were made for or against most trial strategies: partner-focused and structural strategies showed the most consistent benefits, physician-focused strategies were unanimously unsuccessful, and patient-focused strategies were of mixed benefit. CONCLUSIONS: The methodologic quality of many of the located trials was less than optimal. Therefore, further good-quality, randomized trials are necessary to clarify the effectiveness of those strategies identified as potentially useful in this review. PMID- 10722994 TI - Measuring immunization registry costs: promises and pitfalls. AB - INTRODUCTION: The medical and public health communities advocate the use of immunization registries as one tool to achieve national goals for immunization. Despite the considerable investment of resources into registry development, little information is available about the costs of developing or maintaining a registry. METHODS: The objective of this study was to measure the direct costs of maintaining one immunization registry. Cost and resource-use data were collected by interviewing registry personnel and staff at participating pediatric practices, collecting available financial records, and direct observation. RESULTS: The estimated direct cost for maintaining the registry during the 3 calendar years 1995 through 1997 was $439,232. In 1997, this represented an annual cost of $5.26 per child immunized whose record was entered into the registry. In all years, personnel expenses represented at least three fourths of the total costs, with the majority of administrative effort donated. Yearly costs increased over time largely because of growing administrative personnel requirements as the registry became fully operational. CONCLUSION: Considerable resources are required to establish and maintain immunization registries. Because personnel costs, particularly nontechnical personnel, represent a large portion of total registry costs, it is important to accurately account for donated effort. Recommendations for future registry cost studies include prospective data collection and focusing upon the costs of providing specific outreach or surveillance functions rather than overall registry costs. In addition, registry effectiveness evaluations are needed to translate registry costs into cost effectiveness ratios. PMID- 10722995 TI - Honeycomb-like structure of the intermediate layers of the rat superior colliculus, with additional observations in several other mammals: AChE patterning. AB - The aim of the present study was to reinvestigate the stereometric pattern of acetylcholinesterase (AChE) activity staining in the intermediate layers of the superior colliculus in several mammalian species. A pioneering study in the cat and the monkey by Graybiel (1978) stressed the regular arrangement of AChE staining in the deep collicular layers. According to her description, made in the frontal plane, the enzyme was arranged in a mediolateral series of patches, the cores of which tended to line up in the longitudinal axis of the structure, so they formed roughly parallel bands. As exhaustive a description as possible of the AChE distribution was undertaken in the rat by compiling observations in the frontal, sagittal, and tangential planes. It emerged that AChE-positive elements are organized in the form of a conspicuous honeycomb-like network that is divided into about 100 rounded compartments, over virtually the full extent of the intermediate layers. The generality of the rat model was then tested in other rodents such as mouse and hamster and also in cat and monkey. For these species we resorted to a single tangential cutting plane, which proved to be more appropriate for disclosing such a modular arrangement. The data revealed that in all species AChE staining followed the same architectural plan and identified the striking similarity in the number of compartments that compose the various honeycomb-like lattices. In conclusion, the present findings support a unified model of the AChE arrangement within the intermediate layers of the mammalian colliculus; the model comprehensively incorporates the classical description of the patchy and stripy features of the enzyme distribution. We hypothesize here that the modular AChE arrangement might be the anatomical basis for collicular vectorial encoding of orienting movements. PMID- 10722996 TI - Remodeling of the leg sensory system during metamorphosis of the hawkmoth, Manduca sexta. AB - The adult legs of the hawkmoth Manduca sexta are supplied by a diverse array of sensory organs and associated neurons (Kent and Griffin [1990] Cell Tissue Res. 259:209-223) that differ from those in the larval legs. In the present study, a combination of nerve-tracing techniques [biocytin, 1,1;-dioctadecyl-3,3,3;, 3; tetramethyl-indocarbocyanine perchlorate (DiI)], birth date labeling (5 bromodeoxyuridine), confocal microscopy, and electrophysiology were used to describe the remodeling of the prothoracic leg sensory system. Four primary sensory branches carry the axons of all of the sensory neurons in the larval leg. At the onset of metamorphosis, the imaginal leg epidermis develops underneath the larval cuticle and encircles the sensory neurons, thus separating them from their target-organs. Most of the larval neurons degenerate during the larval-to-pupal transition and are replaced by new-adult sensory neurons that are born and differentiate in the pupa. Six sensory neurons that supply hair sensilla in the larval leg, together with 13 femoral and tibial chordotonal organ neurons, persist into the developing adult leg to serve similar functions. Early in the pupal stage, electrical activity can be recorded from these neurons despite the absence of target sensory structures. During the differentiation of the adult sensory system, the axons of the new-adult sensory neurons contact and fasciculate with the axons of the persistent neurons. Thus, five of the primary sensory branches of the adult leg are built on the preexisting larval sensory trajectories. Two sensory branches, however, are established de novo by the axons of specific adult sensory neurons. PMID- 10722997 TI - Calpastatin immunoreactivity in the monkey and human brain of control subjects and patients with Parkinson's disease. AB - Parkinson's disease is characterized by a selective loss of dopaminergic neurons in the nigrostriatal pathway. However, not all dopaminergic neurons degenerate in this disease, and calcium has been suspected of playing a role in this differential vulnerability. An overexpression of the calcium-dependent protease calpain II has recently been reported in the parkinsonian substantia nigra, suggesting that a rise in intracellular calcium concentrations may be involved in the mechanism leading to cell death. The proteasic activity of calpain is regulated by an endogenous inhibitory protein called calpastatin. Because little is known about the distribution of calpastatin in the primate brain, we first analyzed immunohistochemically the calpastatin expression in normal human and monkey brain. A ubiquitous distribution of calpastatin immunostaining was observed in both cases, but its expression was variable from one region to another. In the basal ganglia, staining was intense in the striatum, in the pallidal complex, and in some nuclei of the thalamus. The cerebellum was stained intensely, particularly in the granular and Purkinje cell layers. A dense, heterogeneous staining was observed in the hippocampal formation, mostly in the pyramidal and granular layers. The distribution of staining was similar in the different cerebral cortices studied, and it was most intense in layer V. In the brainstem, staining was particularly prominent in the substantia nigra pars reticulata and compacta, the central gray substance, the superior colliculus, and the cuneiform nucleus, and staining was moderate in the tegmenti pedonculopontinus nucleus and the griseum pontis. In the second part of the study, the authors compared calpastatin expression in the mesencephalon between patients with Parkinson's disease and control subjects. Sequential double staining revealed that some dopaminergic neurons coexpress calpastatin, the proportion of double-stained neurons ranging between 52% and 76% among the different dopaminergic cell groups. Quantitative analysis of the number of calpastatin-stained neurons evidenced a loss of both calpastatin-positive and calpastatin-negative neurons in the substantia nigra of patients with Parkinson's disease. These data suggest that calpain II overexpression in Parkinson's disease is not compensated for by a concomitant increase in calpastatin expression. PMID- 10722998 TI - Rostrocaudal branching within the climbing fibre projection to forelimb-receiving areas of the cerebellar cortical C1 zone. AB - The inferior olive climbing fibre projection to two somatotopically corresponding regions of the paravermal C1 zone in lobule V of the anterior lobe and the rostral folia of the paramedian lobule (PML) in the posterior lobe were investigated in cats by using a combined electrophysiological and retrograde double-labelling technique. In each experiment (n = 7), a small injection of rhodamine-tagged beads was made into the forelimb-receiving part of the C1 zone in one region of the cortex, and an injection of fluorescein-tagged beads was made into the other region. The two regions were found to receive olivary input from cells located in a partially overlapping territory within the rostromedial dorsal accessory olive (DAO). At progressively more rostral levels of DAO, the territory providing climbing fibres to the anterior lobe was centred more laterally than the territory projecting to the rostral PML. Where overlaps occurred, cells that provided climbing fibres to one or the other region were intermingled with a smaller population of double-labelled cells that had axons that branched to supply climbing fibres to both regions of the zone. The double labelled cells represented on average 26% of the smaller total population of labelled cells within the area of overlap and 7% of the total population of neurones projecting to both regions of the zone. Overall, the findings suggested that the C1 zone within spatially separate but somatotopically corresponding regions of the paravermal cerebellar cortex receives at least partially independent olivary inputs, consistent with the presence of distinct microzones at different rostrocaudal levels of the zone. PMID- 10722999 TI - Topographical organization of projections from the subiculum to the hypothalamus in the rat. AB - The projections from the subiculum to the hypothalamus were comprehensively examined in the rat by using the anterograde Phaseolus vulgaris leucoagglutinin (PHA-L) and retrograde cholera toxin B subunit (CTb) methods. Tracing of efferents with PHA-L indicated that the medial preoptic region received projection fibers from the temporal two-thirds of the subiculum, whereas the anterior, tuberal, and mammillary regions received those from the full longitudinal extent of the subiculum. The subicular projections to the anterior and tuberal hypothalamic regions were also found to be organized in a topographical manner such that the temporal-to-septal axis of origin in the subiculum determined a ventromedial-to-dorsolateral axis of termination in the medial zone of the hypothalamus: Massive labeled fibers from the temporalmost part of the subiculum terminated in the subparaventricular zone and its caudal continuum around the dorsal and medial aspects of the ventromedial nucleus, and those from progressively more septal parts terminated in progressively more dorsolateral parts of the medial zone. In addition, the temporal-to-septal axis of origin in the subiculum tended to determine a medial-to-lateral axis of termination in the preoptic region as well as a ventral-to-dorsal axis of termination in the mammillary region. Furthermore, the temporal-to-septal axis of origin in the septal two-thirds of the subiculum corresponded to a ventrolateral to-dorsomedial axis of termination in the medial mammillary nucleus. The topographical projections from the subiculum to the medial zone of the hypothalamus were confirmed by CTb experiments, representatively in the subicular projections to the anterior hypothalamic region. These results suggest that different populations of neurons existing along the longitudinal axis of the subiculum may exert their influences on the execution of different hypothalamic functions. PMID- 10723000 TI - Distribution of GAD-immunoreactive neurons in the diencephalon of the african lungfish Protopterus annectens: colocalization of GAD and NPY in the preoptic area. AB - The distribution of GABAergic neurons was investigated in the diencephalon of the African lungfish, Protopterus annectens, by using specific antibodies directed against glutamic acid decarboxylase (GAD). A dense population of immunoreactive perikarya was observed in the periventricular preoptic nucleus, whereas the caudal hypothalamus and the dorsal thalamus contained only scattered positive cell bodies. Clusters of GAD-positive cells were found in the intermediate lobe of the pituitary. The diencephalon was richly innervated by GAD-immunoreactive fibers that were particularly abundant in the hypothalamus. In the periventricular nucleus, GAD-positive fibers exhibited a radial orientation, and a few neurons extended processes toward the third ventricle. More caudally, a dense bundle of GAD-immunoreactive fibers coursing along the ventral wall of the hypothalamus terminated into the median eminence and the neural lobe of the pituitary. Double-labeling immunocytochemistry revealed that GAD and neuropeptide tyrosine (NPY)-like immunoreactivity was colocalized in a subpopulation of perikarya in the periventricular preoptic nucleus. The proportion of neurons that coexpressed GAD and NPY was higher in the caudal region of the preoptic nucleus. The distribution of GAD-immunoreactive elements in the diencephalon and pituitary of the African lungfish indicates that GABA may act as a hypophysiotropic neurohormone in Dipnoans. The coexistence of GAD and NPY in a subset of neurons of the periventricular preoptic nucleus suggests that GABA and NPY may interact at the synaptic level. PMID- 10723001 TI - Functional organization of crayfish abdominal ganglia. III. Swimmeret motor neurons. AB - Swimmerets are limbs on several segments of the crayfish abdomen that are used for forward swimming and other behaviors. We present evidence that the functional modules demonstrated previously in physiological experiments are reflected in the morphological disposition of swimmeret motor neurons. The single nerve that innervates each swimmeret divides into two branches that separately contain the axons of power-stroke and return-stroke motor neurons. We used Co(++) or biocytin to backfill the entire pool of neurons that innervated a swimmeret, or functional subsets whose axons occurred in particular branches. Each filled cell body extended a single neurite that projected first to the Lateral Neuropil (LN), and there branched to form dendritic structures and its axon. All the motor neurons that innervated one swimmeret had cell bodies located in the ganglion from which their axons emerged, and the cell bodies of all but two of these neurons were located ipsilateral to their swimmeret. Counts of cell bodies filled from selected peripheral branches revealed about 35 power-stroke motor neurons and 35 return-stroke motor neurons. The cell bodies of these two types were segregated into different clusters within the ganglion, but both types sent their neurites into the ipsilateral LN and had their principle branches in this neuropil. We saw no significant differences in the numbers or distributions of these motor neurons in ganglia A2 through A5. These anatomical features are consistent with the physiological evidence that each swimmeret is controlled by its own neural module, which drives the alternating bursts of impulses in power-stroke and return-stroke motor neurons. We propose that the LN is the site of the synaptic circuit that generates this pattern. PMID- 10723002 TI - Differential distribution in rat brain of mu opioid receptor carboxy terminal splice variants MOR-1C-like and MOR-1-like immunoreactivity: evidence for region specific processing. AB - The present study examined immunohistochemically the regional distribution of the mu opioid receptor splice variant MOR-1C by using a rabbit antisera generated against the C-terminal peptide sequences and compared it with MOR-1. Overall, the distribution of MOR-1C-like immunoreactivity (-LI) differed from MOR-1-LI. Both MOR-1C-LI and MOR-1-LI were prominent in a few central nervous system regions, including the lateral parabrachial nucleus, the periaqueductal gray, and laminae I-II of the spinal trigeminal nuclei and the spinal cord. In the striatum, hippocampal formation, presubiculum and parasubiculum, amygdaloid nuclei, thalamic nuclei, locus coeruleus, and nucleus ambiguous MOR-1-LI predominated, whereas MOR-1C-LI was absent or sparse. Conversely, MOR-1C-LI exceeded MOR-1-LI in the lateral septum, the deep laminae of the spinal cord, and most hypothalamic nuclei such as the median eminence, periventricular, suprachiasmatic, supraoptic, arcuate, paraventricular, ventromedial, and dorsomedial nuclei. Double-labeling studies showed colocalization of the two receptors in neurons of the lateral septum, but not in the median eminence or in the arcuate nucleus, even though both MOR-1 isoforms were expressed. Because both MOR-1 and MOR-1C are derived from the same gene, these differences in regional distribution represent region specific mRNA processing. The regional distributions reported in this study involve the epitope seen by the combinations of exons 7, 8, and 9. However, if other MOR-1 variants containing exons 7, 8, and 9 exist, the antisera would not distinguish between them and MOR-1C. PMID- 10723003 TI - Ultrastructural organization of transmitters in the cat lateralis medialis suprageniculate nucleus of the thalamus: an immunohistochemical study. AB - The lateralis medialis-suprageniculate nuclear (LM-Sg) complex of the cat's posterior thalamus receives a rather wide variety of inputs from diverse cortical and subcortical areas. Previous ultrastructural studies of this nucleus demonstrated the presence of four types of vesicle-containing profiles and characterized some of these as gamma-aminobutyric acid (GABA)-containing terminals (Norita and Katoh [1987] J. Comp. Neurol. 263:54-67; Norita and Katoh [1988] Prog. Brain Res. 75:109-118). The present study has extended these observations by examining the immunoreactivity (ir) of LM-Sg, with antibodies raised against aspartate (Asp), glutamate (Glu), GABA, the acetylcholine (ACh) marker, choline acetyltransferase (ChAT), and substance P (SP), by using light and electron microscopy. Neuronal somata immunopositive for the excitatory amino acids (EAAs) Asp and Glu, were of medium size. EAA-ir terminals also were of medium size and contained round synaptic vesicles; they made asymmetrical synaptic contacts with dendritic profiles. Neuronal somata immunopositive for GABA were small. GABA-positive terminals also were small and contained pleomorphic synaptic vesicles; they formed symmetrical synaptic contacts with dendritic profiles. No neurons immunolabeled for ChAT were found. Terminals immunopositive for ChAT were small and contained round synaptic vesicles; these made symmetrical synaptic contacts, asymmetrical synaptic contacts, or both, of the en passant type with dendritic profiles. SP-immunolabeled neuronal somata were not found. Immunolabeled terminals were small, contained round synaptic vesicles, and made asymmetrical synaptic contacts with dendritic profiles. ChAT ir and SP-ir axon terminals were not expressed evenly within LM-Sg. This difference in distribution suggests that within the LM-Sg, there may be a difference in specific sensory processing functions which correlate with transmitter type. PMID- 10723004 TI - Trigeminal ganglion innervates the auditory brainstem. AB - A neural connection between the trigeminal ganglion and the auditory brainstem was investigated by using retrograde and anterograde tract tracing methods: iontophoretic injections of biocytin or biotinylated dextran-amine (BDA) were made into the guinea pig trigeminal ganglion, and anterograde labeling was examined in the cochlear nucleus and superior olivary complex. Terminal labeling after biocytin and BDA injections into the ganglion was found to be most dense in the marginal cell area and secondarily in the magnocellular area of the ventral cochlear nucleus (VCN). Anterograde and retrograde labeling was also seen in the shell regions of the lateral superior olivary complex and in periolivary regions. The labeling was seen in the neuropil, on neuronal somata, and in regions surrounding blood vessels. Retrograde labeling was investigated using either wheatgerm agglutinin-horseradish peroxidase (WGA-HRP), BDA, or a fluorescent tracer, iontophoretically injected into the VCN. Cells filled by retrograde labeling were found in the ophthalmic and mandibular divisions of the trigeminal ganglion. We have previously shown that these divisions project to the cochlea and middle ear, respectively. This study provides the first evidence that the trigeminal ganglion innervates the cochlear nucleus and superior olivary complex. This projection from a predominantly somatosensory ganglion may be related to integration mechanisms involving the auditory end organ and its central targets. PMID- 10723005 TI - Cervical motoneuron topography reflects the proximodistal organization of muscles and movements of the rat forelimb: a retrograde carbocyanine dye analysis. AB - Behavioral evidence reveals that the laboratory rat and other rodent species display skilled paw and digit use in handling food during eating and skilled limb use in reaching for food in formal laboratory skilled reaching tests that is comparable to that described in carnivores and primates. Because less is known about the central control of skilled movements in rodents than in carnivores or primates, the purpose of the current study was to examine the relation between the rat's spinal motoneurons and the individual forelimb muscles that they innervate. In two experiments, 14 forelimb muscles (in the shoulder and the upper and lower arm segments) were injected with carbocyanine dye tracers. The topography of spinal motoneurons was reconstructed by using fluorescence microscopy. Motor neurons were found to be organized in columns throughout the length of the cervical and upper thoracic area, with 1) extensor motoneurons located more laterally than flexor motoneurons, 2) rostral motoneurons innervating more proximal muscles than caudal motoneurons, and 3) more dorsally located motoneurons innervating more distal muscles. These results reveal that the topography of rodent cervical spinal cord motoneurons is very similar to that of carnivores and of primates, which also are characterized by well-developed, skilled movements. In addition, the proximal-distal organization of motoneuron columns parallels the proximal-to-distal pattern of forelimb movement used by the rat when reaching. The data from this study enable the development of predictions about the specific movements that would be compromised by experimental transections or other injuries at different levels of the spinal cord in rat models of spinal injury. PMID- 10723006 TI - Functional differentiation of ganglion cells from multipotent progenitor cells in sliced retina of adult goldfish. AB - Multipotent progenitor cells at the retinal margin of adult goldfish give rise to all cell types in the rest of the retina. We took advantage of this spatial arrangement of progenitor and mature cells in slices of peripheral retina, to investigate the appearance and maturation of voltage-activated Na(+) current. We divided the peripheral retina into three broad regions (marginal, intermediate, and mature) on the basis of their morphological development. Whole-cell patch clamp recordings were performed in ruptured-patch mode, so that cells from which currents were recorded could be identified by Lucifer Yellow fills. No voltage activated Na(+) current was detected in the slender, peripherally located marginal cells. Voltage-activated Na(+) currents were detected in rounded cells found alongside or near marginal cells, facing the vitreal side of the retina. Some of these "intermediate cells" had a long axon-like process which ran along the vitreal surface. Intermediate cells adjacent to the marginal region tended to have smaller Na(+) currents than intermediate cells closer to the mature region. On average, the maximum Na(+) current amplitude recorded from intermediate cells was roughly 6-fold smaller than that of mature ganglion cells. In addition, the activation threshold of the Na(+) current in intermediate cells was nearly 14 mV more positive than that of mature ganglion cells. The results indicate that voltage-activated Na(+) current, as a possible marker of retinal ganglion cells, begins to develop well before these cells migrate to their adult position within the retina. PMID- 10723007 TI - Termination of the geniculocortical projection in the striate cortex of macaque monkey: a quantitative immunoelectron microscopic study. AB - The goal of this present study was to derive a new estimate of the synaptic contribution of the dorsal lateral geniculate nucleus (dLGN) to the subdivisions of its main recipient layer, layer 4C, of striate cortex of macaque monkey. The projection from the dLGN and its terminal boutons within layer 4C were visualized by immunodetection of the calcium binding protein, parvalbumin (PV), which is expressed in relay cells of the dLGN. The proportion of asymmetric synapses formed by PV-positive boutons within the alpha and beta sublayers of 4C was estimated by using a nonbiased stereological counting method. The proportion of asymmetric synapses contributed by the PV-positive boutons to layer 4Calpha is 8.7%; to 4Cbeta is 6.9%. Assuming all the PV-positive asymmetric synapses derive from the dLGN relay cells, this gives a ratio of dLGN synapses per neuron of 192 in layer 4Calpha and 128 in layer 4Cbeta. Thus, the recurrent excitatory input from neighboring cortical neurons must play an important part in responses of the neurons lying at the input stage of the cortical circuit. PMID- 10723008 TI - Distribution of the mRNA encoding the four dopamine D(1) receptor subtypes in the brain of the european eel (Anguilla anguilla): comparative approach to the function of D(1) receptors in vertebrates. AB - Four subtypes of D(1) dopamine receptors are expressed in the brain of the European eel (Anguilla anguilla), an elopomorph teleost. To correlate this molecular multiplicity with specific localisation and functions, the distribution of the D(1) receptor transcripts was analysed by in situ hybridisation. The four D(1) receptor transcripts exhibit largely overlapping expression territories. In telencephalon, they are found in the olfactory bulb and the dorsal telencephalon (except its lateral part) but are most abundant in the subpallial areas. More caudally, the entopeduncular nucleus, preoptic nuclei, preglomerular nuclear complex, ventral thalamus, periventricular hypothalamus, optic tectum and cerebellum, all contain various amounts of D(1) receptor transcripts. Finally, D(1) receptor mRNAs are present in nuclei associated with the cranial nerves. The two D(1A) receptor subtypes are generally the most abundant and present a different distribution in several areas. The D(1B) mRNA, although present in fewer areas than D(1A) transcripts, is the most abundant in ventrolateral telencephalon and torus semicircularis. The D(1C) receptor transcript, which has not been found in mammals, is restricted to diencephalon and cerebellum. In view of the expression territories of D(1) receptor transcripts and previous data, some areas of the everted telencephalon of teleost may be homologous to regions of the tetrapod brain. In particular, D(1) expression territories of the ventral telencephalon are likely to be equivalent to striatal areas. These observations suggest an evolutionary scenario in which the D(1A) receptor subtype was highly conserved after the first gene duplication during the evolution of craniates, whereas D(1B) and D(1C), and their associated specific characteristics, appeared later, probably in the gnathostome lineage. PMID- 10723009 TI - GABA(A) receptor subunit expression within hypophysiotropic CRH neurons: a dual hybridization histochemical study. AB - Dual hybridization histochemical studies were conducted to investigate the extent of colocalization of mRNA transcripts encoding the alpha1-2 and beta1-3 subunits of the gamma aminobutyric acid (GABA)(A) receptor with those for corticotropin releasing hormone (CRH) within the rat hypothalamic paraventricular nucleus (PVN). A vast majority of CRH neurons (>94.5%) were found to express transcripts specific for the the alpha2, beta1 and beta3 subunits; mRNAs for the alpha1 and beta2 subunits of the GABA(A) receptor were detected within 53.3% and 65.7% of PVN CRH neurons, respectively. The results may have important implications for studies aimed at understanding GABAergic influences upon the hypothalamic pituitary-adrenocortical (HPA) axis. Hypophysiotropic CRH neurons serve as the origin of the final common pathway for glucocorticoid secretion in response to stressful stimuli, and GABAergic afferents have been implicated in afferent control of these neurons. The subunit composition of GABA(A) receptors at this key regulatory locus may affect the efficacy of a major inhibitory input, and thus the magnitude and/or duration of stress-induced glucocorticoid secretion. The present findings reveal basal expression patterns of transcripts encoding several subunits of the GABA(A) receptor within stress-integrative CRH neurons, data which may be used to guide regulatory studies of GABAergic influences on the HPA axis under a variety of conditions. PMID- 10723010 TI - Tagma-specific distribution of FXPRLamides in the nervous system of the American cockroach. AB - FXPRLamide (pyrokinin) distribution in the central nervous system and major neurohaemal organs of the American cockroach and related cockroach species was investigated using immunocytochemistry and MALDI-TOF mass spectrometry. Six isoforms (Pea-PK-1 through -6) were found in different neurohaemal release sites. Pea-PK-1-4 and Pea-PK-6 are all stored in the retrocerebral complex and are all produced in cells located in both the suboesophageal ganglion (SOG) and the tritocerebrum. These pyrokinins were found to be concentrated in and around the corpora allata. No other known peptides were detectable in such high concentrations in this neurohaemal organ. They reach the corpora cardiaca/allata via the nervi corporis cardiaci-1 (NCC-1), NCC-3, and nervi corporis allati-2 (NCA-2). Abdominal perisympathetic organs contained only Pea-PK-5 and low quantities of the sequence-related Pea-PK-6. Neither Pea-PK-5 nor -PK-6 was detected in thoracic perisympathetic organs. It is likely that the expression of pyrokinins in the central nervous system is tagma (body region)-specific. Pea-PK 6 was identified during this study as follows: Ser-Glu-Ser-Glu-Val-Pro-Gly-Met Trp-Phe-Gly-Pro-Arg-Leu-NH(2). PMID- 10723011 TI - Effects of aging on myelinated nerve fibers in monkey primary visual cortex. AB - In monkeys, myelin sheaths of the axons in the vertical bundles of nerve fibers passing through the deeper layers of primary visual cortex show age-related alterations in their structure. These alterations have been examined by comparing the myelin sheaths in young monkeys, 5-10 years old, with those in old monkeys, between 25 and 33 years of age. The age-related alterations are of four basic types. In some sheaths, there is local splitting of the major dense line to accommodate dense cytoplasm derived from the oligodendrocytes. Other sheaths balloon out, and in these locations, the intraperiod line in that part of the sheath opens up to surround a fluid-filled space. Other alterations are the formation of redundant myelin so that a sheath is too large for the enclosed axon and the formation of double sheaths in which one layer of compact myelin is surrounded by another one. These alterations in myelin increase in frequency with the ages of the monkeys, and there is a significant correlation between the breakdown of the myelin and the impairments in cognition exhibited by individual monkeys. This correlation also holds even when the old monkeys, 25 to 33 years of age, are considered as a group. It is suggested that the correlation between the breakdown of myelin in the old monkeys and their impairments in cognition has not to do specifically with visual function but to the role of myelin in axonal conduction throughout the brain. The breakdown of myelin could impair cognition by leading to a change in the conduction rates along axons, resulting in a loss of synchrony in cortical neuronal circuits. PMID- 10723012 TI - Visual cortical projections and chemoarchitecture of macaque monkey pulvinar. AB - We investigated the patterns of projections from the pulvinar to visual areas V1, V2, V4, and MT, and their relationships to pulvinar subdivisions based on patterns of calbindin (CB) immunostaining and estimates of visual field maps (P(1), P(2) and P(3)). Multiple retrograde tracers were placed into V1, V2, V4, and/or MT in 11 adult macaque monkeys. The inferior pulvinar (PI) was subdivided into medial (PI(M)), posterior (PI(P)), central medial (PI(CM)), and central lateral (PI(CL)) regions, confirming earlier CB studies. The P(1) map includes PI(CL) and the ventromedial portion of the lateral pulvinar (PL), P(2) is found in ventrolateral PL, and P(3) includes PI(P), PI(M), and PI(CM). Projections to areas V1 and V2 were found to be overlapping in P(1) and P(2), but those from P(2) to V2 were denser than those to V1. V2 also received light projections from PI(CM) and, less reliably, from PI(M). Neurons projecting to V4 and MT were more abundant than those projecting to V1 and V2. Those projecting to V4 were observed in P(1), densely in P(2), and also in PI(CM) and PI(P) of P(3). Those projecting to MT were found in P(1)- P(3), with the heaviest projection from P(3). Projections from P(3) to MT and V4 were mainly interdigitated, with the densest to MT arising from PI(M) and the densest to V4 arising from PI(P) and PI(CM). Because the calbindin-rich and -poor regions of P(3) corresponded to differential patterns of cortical connectivity, the results suggest that CB may further delineate functional subdivisions in the pulvinar. PMID- 10723013 TI - Lazarillo expression reveals a subset of neurons contributing to the primary axon scaffold of the embryonic brain of the grasshopper Schistocerca gregaria. AB - The authors studied the contribution of seven clusters of Lazarillo-expressing cells to the primary axon scaffold of the brain in the grasshopper Schistocerca gregaria from 26% to 43% of embryogenesis. Each cluster, which was numbered according to when Lazarillo expression first appeared, was uniquely identifiable on the basis of its stereotypic position in the brain and the number of Lazarillo expressing cells it contained. At no time during embryogenesis was Lazarillo expression found in brain neuroblasts: It was found only in progeny. For ease of analysis, axogenesis was followed in a cell cluster that contained only a single Lazarillo-expressing cell (the lateral cell) in the dorsal median domain of the brain midline. Bromodeoxyuridine incorporation revealed the presence of only a single midline precursor cell in this region during embryogenesis. Intracellular injection of Lucifer yellow into the lateral cell at various ages showed that there was no dye coupling to the midline precursor or to the nearby term-1 expressing primary commissure pioneers. The lateral cell is not related lineally to these cells and most likely differentiates directly from the neuroectoderm of the brain midline. Lazarillo expression appears at the onset of axogenesis as the lateral cell projects an axon laterally toward the next Lazarillo-expressing cell cluster. The cells of this target cluster direct axons into separate brain regions, thereby establishing an orthogonally organized scaffold that the lateral cell axon follows as it navigates away from the brain midline. The primary axon scaffold of the brain results from a stepwise interlinking of discrete brain regions, as exemplified by axons from neighboring Lazarillo-expressing cell clusters. PMID- 10723014 TI - Low-level exposure to pulsed 900 MHz microwave radiation does not cause deficits in the performance of a spatial learning task in mice. AB - There is some concern that short-term memory loss or other cognitive effects may be associated with the use of mobile cellular telephones. In this experiment, the effect of repeated, acute exposure to a low intensity 900 MHz radiofrequency (RF) field pulsed at 217 Hz was explored using an appetitively-motivated spatial learning and working memory task. Adult male C57BL/6J mice were exposed under far field conditions in a GTEM cell for 45 min each day for 10 days at an average whole-body specific energy absorption rate (SAR) of 0.05 W/kg. Their performance in an 8-arm radial maze was compared to that of sham-exposed control animals. All behavioral assessments were performed without handlers having knowledge of the exposure status of the animals. Animals were tested in the maze immediately following exposure or after a delay of 15 or 30 min. No significant field dependent effects on performance were observed in choice accuracy or in total times to complete the task across the experiment. These results suggest that exposure to RF radiation simulating a digital wireless telephone (GSM) signal under the conditions of this experiment does not affect the acquisition of the learned response. Further studies are planned to explore the effects of other SARs on learned behavior. Bioelectromagnetics 21:151-158, 2000. Published 2000 Wiley-Liss, Inc. PMID- 10723016 TI - Inter-laboratory comparison of numerical dosimetry for human exposure to 60 Hz electric and magnetic fields. AB - In recent years, with the availability of high resolution models of the human body, numerical computations of induced electric fields and currents have been made in more than one laboratory for various exposure conditions. Despite the verification of computational methods, questions are often asked about the reliability of these data. In this paper, computational results from two laboratories that presented data in compatible formats are compared, supplemented with additional data from the third laboratory. Two exposures to uniform fields at 60 Hz are evaluated. The human body models used in the computations are different and so are the computation al methods and codes. There are some differences in the conductivity values used for some of the tissues, as well. The results of the comparison confirm that these data are reliable, as the overall agreement is reasonably good and the differences can be rationally explained. This comparison also underscores the importance of accurate data on the dielectric properties of tissues. PMID- 10723015 TI - Cardiovascular and thermal effects of microwave irradiation at 1 and/or 10 GHz in anesthetized rats. AB - Relatively large thermal gradients may exist during exposure of an animal to microwaves (MWs), particularly at high frequencies. Differences in thermal gradients within the body may lead to noticeable differences in the magnitude of cardiovascular changes resulting from MW exposure. This study compares the thermal distribution and cardiovascular effects of exposure to a single MW frequency with effects of simultaneous exposure to two frequencies. Ketamine anesthetized male Sprague-Dawley rats (n = 58) were exposed individually to one of three conditions: 1-GHz, 10-GHz, or combined 1- and 10-GHz MWs at an equivalent whole-body specific absorption rate of 12 W/kg. The continuous-wave irradiation was conducted under far-field conditions with animals in E orientation (left lateral exposure, long axis parallel to the electric field) or in H orientation (left lateral exposure, long axis perpendicular to the electric field). Irradiation was started when colonic temperature was 37.5 degrees C and was continued until lethal temperatures were attained. Colonic, tympanic, left and right subcutaneous, and tail temperatures, and arterial blood pressure, heart rate, and respiratory rate were continuously recorded. In both E and H orientations, survival time (i.e., time from colonic temperature of 37.5 degrees C until death) was lowest in animals exposed at 1-GHz, intermediate in those exposed at 1- and 10-GHz combined, and greatest in the 10-GHz group (most differences statistically significant). At all sites (with the exception of right subcutaneous), temperature values in the 1- and 10-GHz combined group were between those of the single-frequency exposure groups in both E and H orientations. During irradiation, arterial blood pressure initially increased and then decreased until death. Heart rate increased throughout the exposure period. The general, overall patterns of these changes were similar in all groups. The results indicate that no unusual physiological responses occur during multi frequency MW exposure, when compared with results of single-frequency exposure. Bioelectromagnetics 21:159-166, 2000. Published 2000 Wiley-Liss, Inc. PMID- 10723017 TI - Dantrolene modulates the influence of steady magnetic fields on hippocampal evoked potentials in vitro. AB - Direct current-generated magnetic fields (2-3 mT, 20-min exposure) exerted biphasic effects on the population spike recorded from hippocampal slices. The initial decrease in the potential, observed during exposure of the slices to magnetic fields was followed by a recovery/amplification phase, which began after terminating the magnetic field action. During that phase the population spike exceeded the amplitude observed before application of the magnetic fields. The pattern of magnetic fields influence was not affected either by (+)-5-methyl 10,11-dihydro-5H-dibenzo (a,d) cyclohepten-5, 10-imine maleate (MK801), or by D,L,-2amino-5phosphonovalerate (APV), a noncompetitive and competitive NMDA receptor antagonist, respectively. The rising phase of the potential, however, was eliminated by dantrolene, an inhibitor of intracellular Ca(2 +) channels. This suggests that intracellular calcium channels participate in the mechanism of the influence of the direct current magnetic fields on the function of the hippocampal tissue. PMID- 10723018 TI - High-intensity static magnetic fields modulate skin microcirculation and temperature in vivo. AB - We investigated the acute effect of static magnetic fields of up to 8 T on skin blood flow and body temperature in anesthetized rats. These variables were measured prior to, during, and following exposure to a magnetic field in a superconducting magnet with a horizontal bore. The dorsal skin was transversely incised for 1 cm to make a subcutaneous pocket. Probes of a laser Doppler flowmeter and a thermistor were inserted into the pocket and positioned at mid dorsum to measure skin blood flow and temperature. Another thermistor probe was put into the rectum to monitor rectal temperature. After baseline measurement outside the magnet, the rat was inserted into the bore for 20 min so that mid dorsum was exactly positioned at the center, where the magnetic field was nearly homogeneous. Post-exposure changes were then recorded for 20 min outside the bore. Sham-exposed animals were submitted to exactly the same conditions, except that the superconducting magnet was not energized. Skin blood flow and temperature decreased significantly during magnetic field exposure and recovered after removal of the animal from the magnet. The rectal temperature showed a tendency to decrease while the animal was in the magnet. The microcirculatory and thermal reactions in the present study were consistent and agreed with some of the predictions based on mathematical simulations and model experiments. PMID- 10723019 TI - The effect of static magnetic fields on the rate of calcium/calmodulin-dependent phosphorylation of myosin light chain. AB - This study reports an attempt to confirm a published and well-defined biological effect of magnetic fields. The biological model investigated was the phosphorylation of myosin light chain in a cell free system. The rate of phosphorylation has been reported to be affected in an approximately linear manner by static magnetic field strengths in the range 0-200 microT. We performed three series of experiments, two to test the general hypothesis and a third that was a direct replication of published work. We found no effect of static magnetic field strength on the rate of phosphorylation. Hence, we were unable to confirm that weak static magnetic fields affect the binding of calcium to calmodulin. In view of the difficulty we and other authors have had making independent verifications of claimed biological effects of magnetic fields, we would urge caution in the interpretation of published data until they have been independently confirmed. There are still few well defined biological effects of low level magnetic fields that have been successfully transferred to an independent laboratory. PMID- 10723020 TI - Rate of occurrence of transient magnetic field events in U.S. residences. AB - Recent interest in the transient magnetic field events produced by electrical switching events in residential and occupational environments has been kindled by the possibility that these fields may explain observed associations between childhood cancer and wire codes. This paper reports the results of a study in which the rate of occurrence of magnetic field events with 2-200 kHz frequency content were measured over 24 h or longer periods in 156 U.S. residences. A dual channel meter was developed for the study that, during 20 s contiguous intervals of time, counted the number of events with peak 2-200 kHz magnetic fields exceeding thresholds of 3. 3 nT and 33 nT. Transient activity exhibited a distinct diurnal rhythm similar to that followed by power frequency magnetic fields in residences. Homes that were electrically grounded to a conductive water system that extended into the street and beyond, had higher levels of 33 nT channel transient activity. Homes located in rural surroundings had less 33 nT transient activity than homes in suburban/urban areas. Finally, while transient activity was perhaps somewhat elevated in homes with OLCC, OHCC, and VHCC wire codes relative to homes with underground (UG) and VLCC codes, the elevation was the smallest in VHCC and the largest in OLCC homes. This result does not provide much support for the hypothesis that transient magnetic fields are the underlying exposure that explains the associations, observed in several epidemiologic studies, between childhood cancer and residence in homes with VHCC, but not OLCC and OHCC, wire codes. PMID- 10723021 TI - Children's exposure to magnetic fields produced by U.S. television sets used for viewing programs and playing video games. AB - Two epidemiologic studies have reported increased risk of childhood leukemia associated with the length of time children watched television (TV) programs or played video games connected to TV sets. To evaluate magnetic field exposures resulting from these activities, the static, ELF, and VLF magnetic fields produced by 72 TV sets used by children to watch TV programs and 34 TV sets used to play video games were characterized in a field study conducted in Washington DC and its Maryland suburbs. The resulting TV-specific magnetic field data were combined with information collected through questionnaires to estimate the magnetic field exposure levels associated with TV watching and video game playing. The geometric means of the ELF and VLF exposure levels so calculated were 0.0091 and 0.0016 microT, respectively, for children watching TV programs and 0.023 and 0.0038 microT, respectively, for children playing video games. Geometric means of ambient ELF and VLF levels with TV sets turned off were 0.10 and 0.0027 microT, respectively. Summed over the ELF frequency range (6-3066 Hz), the exposure levels were small compared to ambient levels. However, in restricted ELF frequency ranges (120 Hz and 606-3066 Hz) and in the VLF band, TV exposure levels were comparable to or larger than normal ambient levels. Even so, the strengths of the 120 Hz or 606-3066 Hz components of TV fields were small relative to the overall ambient levels. Consequently, our results provide little support for a linkage between childhood leukemia and exposure to the ELF magnetic fields produced by TV sets. Our results do suggest that any future research on possible health effects of magnetic fields from television sets might focus on the VLF electric and magnetic fields produced by TV sets because of their enhanced ability relative to ELF fields to induce electric currents. PMID- 10723022 TI - Effects on Rb(+)(K+) uptake of HeLa cells in a high K(+) medium of exposure to a switched 1.7 Tesla magnetic field. AB - Effects of a switched, time-varying 1.7 T magnetic field on Rb(+)(K+) uptake by HeLa S3 cells incubated in an isosmotic high K(+) medium were examined. The magnetic flux density was varied intermittently from 0.07-1.7 T at an interval of 3 s. K(+) uptake was activated by replacement of normal medium by high K(+) medium. A membrane-permeable Ca(2+) chelating agent (BAPTA-AM) and Ca(2+) dependent K(+) channel inhibitors (quinine, charibdotoxin, and iberiotoxin) were found to reduce the Rb(+)(K+) uptake by about 30-40%. Uptake of K(+) that is sensitive to these drugs is possibly mediated by Ca(2+)-dependent K(+) channels. The intermittent magnetic field partly suppress ed the drug-sensitive K(+) uptake by about 30-40% (P < 0.05). To test the mechanism of inhibition by the magnetic fields, intracellular Ca(2+) concentration ([Ca(2+)]c) was measured using Fura 2 AM. When cells were placed in the high K(+) medium, [Ca(2+)]c increased to about 1.4 times the original level, but exposure to the magnetic fields completely suppressed the increase (P < 0.01). Addition of a Ca(2+) ionophore (ionomycin) to the high K(+) medium increased [Ca(2+)]c to the level of control cells, regardless of exposure to the magnetic field. But the inhibition of K(+) uptake by the magnetic fields was not restored by addition of ionomycin. Based on our previous results on magnetic field-induced changes in properties of the cell membrane, these results indicate that exposure to the magnetic fields partly suppresses K(+) influx, which may be mediated by Ca(2+)-dependent K(+) channels. The suppress ion of K(+) fluxes could relate to a change in electric properties of cell surface and an inhibition of Ca(2+) influx mediated by Ca(2+) channels of either the cell plasma membrane or the inner vesicular membrane of intracellular Ca(2+) stores. PMID- 10723023 TI - Modulation of cytokine production by interferential current in differentiated HL 60 cells. AB - The influence of interferential current (IFC) on the release of four cytokines was investigated. IFC is an amplitude-modulated 4 kHz current used in therapeutic applications. Human promyelocytes (HL-60) were differentiated to monocytes/macrophages by treatment with calcitriol. Release of tumor necrosis factor alpha (TNFalpha) and interleukines 1beta, 6, and 8 (IL-1beta, IL-6, and IL 8) into the supernatant was measured after exposure to IFC at different modulation frequencies. TNFalpha release was stimulated about twofold by 4 kHz sine waves alone. The influences of exposure time (5-30 min) and current density (2.5-2500 microA/c m(2)) were tested. A maximum field effect was found at an exposure time of 15 min and a current density of 250 microA/cm(2). With these exposure conditions (15 min and 250 microA/cm(2) ), cells were treated at different modulation frequencies and reacted for TNFalpha, IL-1beta, and IL-8 release in a complex manner. Within the frequencies studied (0-125 Hz), we found stimulation as well as depression of the release. In a second run the cells were activated by pretreatment with 10 microg/ml lipopolysaccharide (LPS) and exposed in the same way as the nonactivated cells. Again the modulation frequency influenced, in a complex way, the induction of TNFalpha, IL-1beta, and IL-8, resulting in a pattern of stimulation and depression of release different from that found in nonactivated cells. For IL-6 production no significant changes were detected in activated or non-activated cells. PMID- 10723024 TI - A(3) adenosine receptor ligands: history and perspectives. AB - Adenosine regulates many physiological functions through specific cell membrane receptors. On the basis of pharmacological studies and molecular cloning, four different adenosine receptors have been identified and classified as A(1), A(2A), A(2B), and A(3). These adenosine receptors are members of the G-protein-coupled receptor family. While adenosine A(1) and A(2A) receptor subtypes have been pharmacologically characterized through the use of selective ligands, the A(3) adenosine receptor subtype is presently under study in order to better understand its physio-pathological functions. Activation of adenosine A(3) receptors has been shown to stimulate phospholipase C and D and to inhibit adenylate cyclase. Activation of A(3) adenosine receptors also causes the release of inflammatory mediators such as histamine from mast cells. These mediators are responsible for processes such as inflammation and hypotension. It has also been suggested that the A(3) receptor plays an important role in brain ischemia, immunosuppression, and bronchospasm in several animal models. Based on these results, highly selective A(3) adenosine receptor agonists and/or antagonists have been indicated as potential drugs for the treatment of asthma and inflammation, while highly selective agonists have been shown to possess cardioprotective effects. The updated material related to this field of research has been rationalized and arranged in order to offer an overview of the topic. PMID- 10723025 TI - The HIV-1 reverse transcription (RT) process as target for RT inhibitors. AB - Since the Human Immunodeficiency Virus Type 1 (HIV-1) was identified as the etiologic agent of the Acquired Immune Deficiency Syndrome (AIDS), the HIV-1 reverse transcriptase (RT) has been the subject of intensive study. The reverse transcription entails the transition of the single-stranded viral RNA into double stranded proviral DNA, which is then integrated into the host chromosome. Therefore, the HIV-1 reverse transcriptase plays a pivotal role in the life cycle of the virus and is consequently an interesting target for anti-HIV drug therapy. In the first section, we describe the complex process of reverse transcription and the different activities involved in this process. We then highlight the structure-function relationship of the HIV-1 reverse transcriptase, which is of great importance for a better understanding of resistance development, a major problem in anti-AIDS therapies. Finally, we summarize the mechanisms of HIV resistance toward various RT inhibitors and the implications thereof for the current anti-HIV drug therapies. PMID- 10723026 TI - Chemokine receptor antagonists. AB - Chemokines belong to a large family of chemoattractant molecules involved in the directed migration of immune cells. They achieve their cellular effects by direct interaction with cell surface receptors. Chemokines appear to be involved in a variety of proinflammatory and autoimmune diseases and this makes them and their receptors very attractive therapeutic targets. This review highlights current efforts toward obtaining highly specific chemokine receptor antagonists and discusses their chemistry and potential role as disease modifiers. PMID- 10723027 TI - The development of delivery agents that facilitate the oral absorption of macromolecular drugs. AB - Macromolecules comprise a growing group of new drugs with great clinical promise. To date, the therapeutic application of these drugs has been limited, because they are effective only when administered parenterally. Unfortunately, macromolecular drugs are not absorbed following nonparenteral dosing, because the mechanisms of the human body are designed to degrade and/or exclude them. To overcome the numerous obstacles to the noninvasive delivery of these drugs, various approaches are under investigation including the use of delivery agents to promote drug absorption. This review provides a summary of the novel approaches currently in progress in the areas of transdermal, transmucosal, and oral delivery of macromolecular drugs facilitated by delivery agents. We review our own novel work in this area in some detail, including the methods developed for the synthesis of the delivery agents, in vitro screening techniques developed to select compounds for in vivo testing, and the results of in vivo screening in both rats and primates, including preliminary safety and efficacy studies. Finally, the results of Phase I clinical studies showing the oral delivery of heparin are presented. PMID- 10723028 TI - Spiny legs and prickled bodies: new insights and complexities in planar polarity establishment. AB - Epithelial cells can be polarized along two axes, namely in the apical basolateral axis and in the horizontal plane of the epithelium. Vertebrate examples of planar polarization include aspects of skin development or features in internal organs, such as the inner ear epithelium. In insects like Drosophila, adult cuticular structures show planar polarization. Studies on planar polarity in Drosophila have identified several genes that regulate this process. Notably, the Frizzled receptor and its signaling cascade provide an entry point to the molecular aspects of planar polarization. A recent study by Gubb et al.((1)) of the prickle locus, which encodes a cytoplasmic protein with three LIM domains, provides new insights and raises several interesting questions that can now be addressed. Pk might serve a scaffolding function involved in assembling a protein complex required for planar polarity establishment. PMID- 10723029 TI - Peeling by binding or twisting by cranking: models for promoter opening and transcription initiation by RNA polymerase II. AB - The precise, sequence-specific regulation of RNA synthesis is the primary mechanism underlying differential gene expression. This general statement applies to both prokaryotic and eukaryotic organisms, as well as to their viral pathogens. Thus, it is not surprising that genomes use a substantial portion of their protein-coding content to regulate the process of RNA synthesis. Transcriptional regulation in bacterial systems is particularly well understood. In this essay, we build on this knowledge and propose two opposing models to describe promoter opening and transcription initiation in the eukaryotic RNA polymerase II system. Promoter opening in the "twisting by cranking" model is based on changes in the trajectory of DNA. In contrast, invasion of single stranded DNA-binding proteins between the DNA strands drives the reaction in the "peeling by binding" model. PMID- 10723031 TI - Ca(2+)-binding proteins in the retina: structure, function, and the etiology of human visual diseases. AB - The complex sensation of vision begins with the relatively simple photoisomerization of the visual pigment chromophore 11-cis-retinal to its all trans configuration. This event initiates a series of biochemical reactions that are collectively referred to as phototransduction, which ultimately lead to a change in the electrochemical signaling of the photoreceptor cell. To operate in a wide range of light intensities, however, the phototransduction pathway must allow for adjustments to background light. These take place through physiological adaptation processes that rely primarily on Ca(2+) ions. While Ca(2+) may modulate some activities directly, it is more often the case that Ca(2+)-binding proteins mediate between transient changes in the concentration of Ca(2+) and the adaptation processes that are associated with phototransduction. Recently, combined genetic, physiological, and biochemical analyses have yielded new insights about the properties and functions of many phototransduction-specific components, including some novel Ca(2+)-binding proteins. Understanding these Ca(2+)-binding proteins will provide a more complete picture of visual transduction, including the mechanisms associated with adaptation, and of related degenerative diseases. PMID- 10723030 TI - Transcription elongation factor SII. AB - RNA chain elongation by RNA polymerase II (pol II) is a complex and regulated process which is coordinated with capping, splicing, and polyadenylation of the primary transcript. Numerous elongation factors that enable pol II to transcribe faster and/or more efficiently have been purified. SII is one such factor. It helps pol II bypass specific blocks to elongation that are encountered during transcript elongation. SII was first identified biochemically on the basis of its ability to enable pol II to synthesize long transcripts. ((1)) Both the high resolution structure of SII and the details of its novel mechanism of action have been refined through mutagenesis and sophisticated in vitro assays. SII engages transcribing pol II and assists it in bypassing blocks to elongation by stimulating a cryptic, nascent RNA cleavage activity intrinsic to RNA polymerase. The nuclease activity can also result in removal of misincorporated bases from RNA. Molecular genetic experiments in yeast suggest that SII is generally involved in mRNA synthesis in vivo and that it is one type of a growing collection of elongation factors that regulate pol II. In vertebrates, a family of related SII genes has been identified; some of its members are expressed in a tissue-specific manner. The principal challenge now is to understand the isoform specific functional differences and the biology of regulation exerted by the SII family of proteins on target genes, particularly in multicellular organisms. PMID- 10723032 TI - Checkpoints controlling mitosis. AB - Each year many reviews deal with checkpoint control.((1-5)) Here we discuss checkpoint pathways that control mitosis. We address four checkpoint systems in depth: budding yeast DNA damage, the DNA replication checkpoint, the spindle assembly checkpoint and the mammalian G2 topoisomerase II-dependent checkpoint. A main focus of the review is the organization of these checkpoint pathways. Recent work has elucidated the order-of-function of several checkpoint components, and has revealed that the S phase, DNA damage and spindle assembly checkpoints each have at least two parallel branches. These steps forward have largely come from kinetic studies of checkpoint-defective mutants. PMID- 10723033 TI - Targeting and insertion of nuclear-encoded preproteins into the mitochondrial outer membrane. AB - Most mitochondrial proteins are synthesized in the cytosol as preproteins with a cleavable presequence and are delivered to the import receptors on the mitochondria by cytoplasmic import factors. The proteins are then imported to the intramitochondrial compartments by the import systems of the outer and inner membranes, TOM and TIM. Mitochondrial outer membrane proteins are synthesized without a cleavable presequence and most of them contain hydrophobic transmembrane domains, which, in conjunction with the flanking segments, function as the mitochondria import signals. Some of the proteins are inserted into the outer membrane by the TOM machinery; the import signal probably arrests further translocation and is released from the translocation channel to the lipid bilayer. The other proteins are inserted into the membrane by a novel pathway independent of the TOM machinery. This article reviews recent developments in the biogenesis of mitochondrial outer membrane proteins. PMID- 10723034 TI - Canalization in evolutionary genetics: a stabilizing theory? AB - Canalization is an elusive concept. The notion that biological systems ought to evolve to a state of higher stability against mutational and environmental perturbations seems simple enough, but has been exceedingly difficult to prove. Part of the problem has been the lack of a definition of canalization that incorporates an evolutionary genetic perspective and provides a framework for both mathematical and empirical study. After briefly reviewing the importance of canalization in studies of evolution and development, we aim, with this essay, to outline a research program that builds upon the definition of canalization as the reduction in variability of a trait, and uses molecular genetic approaches to shed light on the problems of canalization. PMID- 10723035 TI - To be or not to be active: the stochastic nature of enhancer action. AB - Transcriptional enhancers are traditionally considered to regulate the rate at which a linked promoter transcribes mRNA, but recent experiments suggest a reevaluation of this model is necessary. Single-cell assays of transgenes reveal that enhancers increase the probability that a reporter gene will be active, but have little or no effect on the transcription rate once a gene has been activated. These results raise the question of how enhancers affect gene expression in their native contexts. A simple interpretation is that enhancers act in a stochastic fashion to increase the probability that a regulated gene will be transcribed; such a model is compatible with programs of cell differentiation in which multiple similar cells subject to similar environmental stimuli do not respond uniformly. PMID- 10723036 TI - Mutation is modulated: implications for evolution. AB - Evolution occurs through genome variation followed by selection. Because DNA sequence context affects the activity of enzymes that copy, move and repair DNA, there are intrinsic variations in the probability of genetic variation along a genome. These intrinsic variations can be affected by selective pressure. Codon changes that do not alter the encoded amino acids may still have effects on the local rate of sequence change. Large gene families could encode a successful genetic framework by which to evolve new, functional members. The speed of adaptation to environmental challenges may be improved when the distinct mechanisms of genetic change come under regulatory control. Natural selection operates on mechanisms that generate and modulate diversity as it does on all biological functions. PMID- 10723037 TI - Illusory defects and mismatches: why must DNA repair always be (slightly) error prone? AB - There is growing evidence that recombination is mu;tagenic and that some forms of DNA repair synthesis are error prone. DNA synthesis in mismatch repair might also be error prone. DNA-repair systems detect structural defects in DNA with high efficiency but they occasionally also strike at normal sections of DNA. Considering the diversity of local DNA structure, some DNA sections with complementary sequences are bound to act as preferential false targets for a repair system (i.e. as "illusory defects"). However, if the repair system never changes the sequence upon repair, it will be solicited again and again by the illusory defect, a potentially harmful situation. It is therefore advantageous for a repair system to be, to some extent, error prone. Strong illusory defects may arise at the decanucleotide level and could be the cause of local increases in mutation levels. They might be used to initiate somatic hypermutation pathways. PMID- 10723038 TI - Set down in bone PMID- 10723039 TI - Incidence of occupational asthma by occupation and industry in Finland. AB - BACKGROUND: Systematic research on occupation or industry-specific incidence of occupational asthma (OA) is sparse. We calculated the incidence of notified OA by occupation, industry and causative agent in Finland for the years 1989-95. METHODS: The numbers of cases of reported OA were retrieved from the Finnish Registry of Occupational Diseases for the population between 20 and 64 years of age. The numbers of employed workers were retrieved from Statistics Finland. Incidence rates were calculated for each occupation, industry and the total workforce. RESULTS: Altogether 2602 cases of OA were notified and the mean annual incidence rate was 17.4 cases/100,000 employed workers. The incidence rate was the highest in bakers, other painters and lacquerers, veterinary surgeons, chemical workers, farmers, animal husbandry workers, other food manufacturing workers, welders, plastic product workers, butchers and sausage makers, and floor layers. Cases caused by animal epithelia, hairs and secretions or flours, grains, and fodders accounted for 60% of the total. CONCLUSIONS: Estimation of occupation and industry-specific incidence rates forms the basis for successful prevention of OA, but necessitates collection of data over several years from well established surveillance systems. PMID- 10723040 TI - Upper airway inflammation assessed by nasal lavage in compost workers: A relation with bio-aerosol exposure. AB - BACKGROUND: Exposure to microbial agents in the composting industry may cause work related airway inflammation. Nasal lavage (NAL) has been proposed as a noninvasive method to assess such effects in population studies. METHODS: Pre- and post-shift NAL were performed in the workers of a compost plant visited in 1995 (n = 14) and 1996 (n=15), of whom only four participated in both surveys. Total cells, cytokines and other inflammation markers were measured in NAL fluid, and pre-shift levels and post/pre concentration ratios were compared with NAL results obtained in the same periods in 10 and 9 controls, respectively, and with levels of airborne exposure to microbial agents endotoxin and beta(1,3)-glucan as measured in personal air samples. RESULTS: Job-title specific exposure levels in the first survey ranged from 75 to 527 EU/m(3) for endotoxin and from 0.54 to 4.85 microg/m(3) for beta(1,3)-glucan. In the second survey these values were lower, 29-285 EU/m(3) and 0.36-4.44 microg/m(3), respectively. In the first survey pre-shift NAL concentrations of total cells, MPO, IL-8, NO and albumin were significantly (1.1-4.8 fold) higher in compost workers than in controls. Post/pre ratios for various markers were significantly (1.2-3.2 fold) higher in compost workers in both surveys. NAL cells were mainly neutrophils, while eosinophils were only incidentally observed. A weak relation with exposure was found for pre-shift levels of MPO, uric acid and urea in the first survey. CONCLUSIONS: Occupational exposure of compost workers may cause acute and possibly (sub-)chronic inflammatory reactions in the upper airways, presumably induced by non-allergenic pro-inflammatory agents like endotoxins and beta(1, 3) glucans. PMID- 10723041 TI - Lung cancer and exposure to man-made vitreous fibers: results from a pooled case control study in Germany. AB - BACKGROUND: To investigate the association between lung cancer and occupational exposure to man-made vitreous fibers (MMVF), a pooled analysis of two case control studies was conducted in the years 1988-1994. METHODS: The case series consisted of 3498 males who were histologically or cytologically verified primary lung cancer cases. 3541 male population controls were drawn at random from the general population and matched to cases by sex, age, and place of residence. To examine the relationship between MMVF and lung cancer we asked all study subjects who worked for at least 6 months as construction and installation workers whether they ever installed or removed insulations and what kind of insulation material they used. RESULTS: Some 304 (8.7%) cases and 170 (4.8%) controls reported to have insulated with glass wool or mineral wool mats. Coded as ever/never exposed, the odds ratio was 1.48 (95% CI: 1.17-1.88), adjusted for smoking and asbestos. To be sure to exclude any confounding effect of asbestos, we tried to identify those cases and controls who insulated with glass wool or mineral wool mats only and never reported any asbestos exposure. For this group we calculated an odds ratio of 1.56 (95% CI: 0.92-2.65), after adjustment for smoking. An elevated risk was also estimated on the basis of an expert rating which was done for a subgroup of cases and controls. Ever exposure to MMVF (but not to asbestos) in this subgroup yielded an odds ratio of 1.30 (95% CI: 0.82-2.07). CONCLUSIONS: Our study provides some indication for an excess risk of man-made vitreous fibers. This result also persists after adjustment for smoking and asbestos. PMID- 10723043 TI - Survival of infectious Pseudomonas aeruginosa genotypes in occupational saturation diving environment and the significance of these genotypes for recurrent skin infections. AB - BACKGROUND: Occupational saturation divers suffer from various skin disorders, of which skin infections are the most serious and frequent. Pseudomonas aeruginosa is the microbe most often isolated. METHODS: P. aeruginosa isolates from 292 skin infections in operational saturation divers and about 800 isolates from occupational saturation diving systems have been collected during the period 1986 to 1998. Genotyping of the isolates has been performed by using restriction enzyme fragmentation and pulsed field gel electrophoresis. RESULTS: Four hundred and seventy-two P. aeruginosa isolates have been analyzed, of which 181 originate from skin infections in divers. Ninety-seven significantly different P. aeruginosa genotypes have been defined. Some of these genotypes are solely found from skin infections, some solely from the saturation environment and about 25% were found both from infections and from the saturation environment. Eight frequent infectious genotypes have been identified, and these are shown to be present over several years, both in infections and in the saturation environment. CONCLUSIONS: The study suggests that skin infections in occupational saturation divers are commonly caused by environmental strains. PMID- 10723044 TI - Wood dust exposure and cancer incidence: a retrospective cohort study of furniture workers in Estonia. AB - BACKGROUND: Occupational wood dust exposure is associated with increased risk of sinonasal cancer in men. However, little is known whether it is associated with sinonasal cancer in women or with malignancies of other sites. METHODS: In a retrospective cohort study of furniture workers, cancer incidence in 3723 men and 3063 women between 1968 and 1995 was compared to the incidence in the general population of Estonia. Cancer risks were analyzed by employment duration and occupation. RESULTS: The standardized incidence ratio (SIR) for all cancers did not differ significantly from one. Two men and one woman had sinonasal cancer (expected 1.07 and 0.53, respectively). Significantly increased risk of colon cancer was seen in the cohort (SIR 1.65, 95% confidence interval (CI) 1.22-2.17). Subjects employed for 10 years and over had significant excess of colon cancer (SIR 2.29, 95% CI 1.28-3.77) and rectal cancer (SIR 2.10, 95% CI 1.05-3.76) in the analysis by employment duration using exposure with a latency of 20 years. The nonsignificant excess of pharyngeal cancer in men (SIR 1.82) and lung cancer in women (SIR 1.43) was restricted to short-term workers. CONCLUSIONS: This study found an excess of colon and rectal cancer in furniture workers. There was no increase in total cancer risk. PMID- 10723042 TI - Proportionate mortality among unionized roofers and waterproofers. AB - BACKGROUND: The United Union of Roofers, Waterproofers, and Allied Workers (UURWAW) is one of the 15 building and construction trades departments in the AFL CIO. The U.S. roofing industry, including both roofing and waterproofing applications, both unionized and nonunionized, comprises about 25,000 firms, employing approximately 300,000 people, about 200,000 of whom are involved in the application of roofs. The specific toxins to which roofers may be exposed at the job site include, among others, bitumens (asphalt and/or coal tar pitch) as well as asbestos and fiberglass from roof removal operations. Excess deaths from occupational injuries are also of concern. METHODS: This study evaluated causes of mortality among 11,144 members of the UURWAW. Age-adjusted proportionate mortality ratios (PMRs) were computed with 95% confidence intervals (CI) using U.S. age-, gender-, and race-specific proportional mortality rates for the years of the study, 1950-1996. RESULTS: Statistically significant increased PMRs were found for all injuries (PMR = 142, CI = 134-150), especially falls (PMR = 464, CI = 419-513) and other injuries (PMR = 121, CI = 107-137), cancers of the lung (PMR = 139, CI = 131-148), bladder (PMR = 138, CI = 111-170), esophagus (PMR = 134, CI = 107-166), larynx (PMR = 145, CI = 106-193), and cancers of other and unspecified sites (PMR = 130, CI = 112-149), pneumoconioses and other nonmalignant respiratory diseases (PMR = 115, CI = 103-128), and homicides (PMR = 153, CI = 135-172). The occupational exposures which may have contributed to the excess risks of malignant and nonmalignant respiratory diseases include, among others, asphalt fumes, coal tar pitch volatiles and asbestos; however, cigarette smoking must also be considered a contributing factor. CONCLUSIONS: The present study underscores the need to control airborne exposures to hazardous substances and especially to examine fall prevention efforts within the roofing industry. Am. J. Ind. Med. 37:478-492, 2000. Published 2000 Wiley-Liss, Inc. PMID- 10723045 TI - Recent geographic patterns of lung cancer and mesothelioma mortality rates in 49 shipyard counties in the United States, 1970-94. AB - BACKGROUND: Lung cancer mortality rates among white males in the United States were observed to be elevated during 1950-69 in counties with shipbuilding industries during World War II; risk was found to be associated with asbestos exposure. We evaluated the geographic patterns in more recent years, 1970-94, for whites and compared them with the 1950-69 patterns. METHODS: We calculated age adjusted rates and estimated rate ratios between comparison groups. RESULTS: Rates generally were higher in shipyard counties than in all nonshipyard counties and in coastal nonshipyard counties for both sexes and time periods. Rates increased markedly from 1950-69 to 1970-94 in all groups, with the changes more pronounced in females than males. Pleural mesothelioma mortality rates were also significantly higher in shipyard counties than coastal nonshipyard counties in all regions among males but not among females. CONCLUSIONS: The more pronounced changes in lung cancer mortality rates among females in shipyard counties may be attributed to the combined effects of low asbestos exposures and changes in smoking behavior. Am. J. Ind. Med. 37:512-521, 2000. Published 2000 Wiley-Liss, Inc. PMID- 10723046 TI - Validation of biomarkers in humans exposed to benzene: urine metabolites. AB - BACKGROUND: The present study was conducted among Chinese workers employed in glue- and shoe-making factories who had an average daily personal benzene exposure of 31+/-26 ppm (mean+/-SD). The metabolites monitored were S phenylmercapturic acid (S-PMA), trans, trans-muconic acid (t,t-MA), hydroquinone (HQ), catechol (CAT), 1,2, 4-trihydroxybenzene (benzene triol, BT), and phenol. METHODS: S-PMA, t,t-MA, HQ, CAT, and BT were quantified by HPLC-tandem mass spectrometry. Phenol was measured by GC-MS. RESULTS: Levels of benzene metabolites (except BT) measured in urine samples collected from exposed workers at the end of workshift were significantly higher than those measured in unexposed subjects (P < 0.0001). The large increases in urinary metabolites from before to after work strongly correlated with benzene exposure. Concentrations of these metabolites in urine samples collected from exposed workers before work were also significantly higher than those from unexposed subjects. The half-lives of S-PMA, t,t-MA, HQ, CAT, and phenol were estimated from a time course study to be 12.8, 13.7, 12.7, 15.0, and 16.3 h, respectively. CONCLUSIONS: All metabolites, except BT, are good markers for benzene exposure at the observed levels; however, due to their high background, HQ, CAT, and phenol may not distinguish unexposed subjects from workers exposed to benzene at low ambient levels. S-PMA and t,t-MA are the most sensitive markers for low level benzene exposure. PMID- 10723047 TI - Occupational exposure to chlorophenol and the risk of nasal and nasopharyngeal cancers among U.S. men aged 30 to 60. AB - BACKGROUND: Elevated rates of nasal and nasopharyngeal cancers have been associated with wood-related occupational exposures, including chlorophenols, formaldehyde, and wood dust. METHODS: Occupational information was obtained from 43 nasal carcinoma cases, 92 nasopharyngeal carcinoma cases, and 1909 controls, by interview. Exact conditional logistic regression was used to evaluate the association of these cancers with chlorophenol exposure, estimated from a review of verbatim responses. RESULTS: Both nasal and nasopharyngeal cancers were significantly associated with estimated duration of chlorophenol exposure. For nasopharyngeal cancer, elevated risk was observed among those who held jobs assigned medium or high intensity chlorophenol exposure (n(exposed)=18, OR=1.94, 95% CI=1.03-3.50) and among those with 10+ years in jobs assigned high intensity with high certainty (n(exposed)=3, OR=9.07, 95% CI=1.41-42. 9). Controlling for estimated formaldehyde and wood dust exposure did not alter these findings, as much of the estimated chlorophenol exposure was among machinists. CONCLUSIONS: These findings support the hypothesis that occupational exposure to chlorophenol is a risk factor for nasal and nasopharyngeal cancer, although the role of machining-related exposures warrants further assessment. PMID- 10723048 TI - Nature, incidence, and cause of work-related amputations in Minnesota. AB - BACKGROUND: The Minnesota Sentinel Event Notification System for Occupational Risks (SENSOR) has collected data on the nature, incidence, and cause of work related amputation injuries that have taken place since 1992. METHODS: SENSOR defined an amputation as any finger amputation or the loss of any other body part; 832 workers were identified as having amputation injuries between 1994 and 1995 and 72% of these workers completed telephone interviews. RESULTS: The amputation injury rate for Minnesota workers was 39 per 100,000 workers, with agriculture and manufacturing having the highest rates. Sixty-six percent of the injuries involved one finger; 14% involved two or more fingers. Persons working with machinery reported 73% of the injuries. CONCLUSIONS: A closer examination of the incidence and causes for amputation injuries shows that these were not random events. Reliance on human reactions to prevent injury is inadequate; therefore, additional research needs to be conducted. PMID- 10723049 TI - Medical, personal, and occupational outcomes for work-related amputations in Minnesota. AB - BACKGROUND: The Minnesota Sentinel Event Notification System for Occupational Risks (SENSOR) surveillance system has collected data on the medical, personal, and occupational outcomes associated with work-related amputations since 1992. METHODS: SENSOR defined amputations as any finger amputation or the loss of any other body part; 832 workers were identified as having amputation injuries between 1994 and 1995 and 72% of these workers completed a telephone interview. RESULTS: Twenty percent of those injured required overnight hospitalization. Ninety-one percent of the cases reported having missed work, with 56% reporting missing ten or more days. Individuals working on their usual jobs at the time of injury were more likely to report less serious medical and occupational outcomes. CONCLUSIONS: Severe injuries were significantly associated with worse medical, personal, and occupational outcomes. Two groups of machines, material handling, and powered handtools were associated with a higher proportion of severe injuries. PMID- 10723050 TI - Low back pain disability: relative costs by antecedent and industry group. AB - BACKGROUND: Previous studies of workers' compensation claims for low back pain (LBP) have revealed that the preponderance of disability is borne by a fraction of cases. However, less is known regarding the influence of occupational factors on these extreme conditions. METHODS: Workers' compensation claims (n=107,867) for LBP reported to a large, national insurer in 1992 were examined by antecedent event and industry class. In addition to summaries of the frequency and cost distribution, each factor was examined at two points on its cost distribution: one more representative of the typical case and one more representative of the case with long disability. These alternative disability indicators were introduced to explore a different perspective of LBP disability. RESULTS: The information provided by the alternative indicators was distinct from the information provided by the traditional aggregate indicators (claim frequency and claim cost frequency). In particular, this method identified increased severity for claims in the construction and services sectors, as well as for claims arising from falls and motor vehicle crashes. CONCLUSIONS: The results suggest that the construction and service sectors confront unique challenges to prevention and management of LBP disability. LBP related to discrete antecedents such as falls and motor vehicle crashes merits consideration on the basis of exceptionally severe disability. PMID- 10723051 TI - Regional localization of dopamine and ionotropic glutamate receptor subtypes in striatolimbic brain regions. AB - Localization of dopamine (D(1)-, D(2)-like, and D(4)) and ionotropic glutamate (NMDA, AMPA, and KA) receptor subtypes within the striatolimbic forebrain remains incomplete, but basic to understanding the functional organization of this important brain region. We found that frontal cortical ablation supported colocalization of D(4) and NMDA receptors on corticostriatal afferents to caudate putamen and nucleus accumbens in rat forebrain. Local injection of kainic acid into caudate-putamen, nucleus accumbens, or hippocampus produced massive local postsynaptic losses of D(1)- and D(2)-like, as well as NMDA, AMPA, and KA receptors, and kainic acid ablation of hippocampal-striatal projections indicated the selective expression of presynaptic NMDA and KA autoreceptors. Degeneration of nigrostriatal dopamine projections with 6-hydroxydopamine showed that all three glutamatergic subtypes exist as heteroceptors on nigrostriatal dopaminergic terminals. Our findings suggest common interactions between excitatory glutamatergic and inhibitory dopaminergic receptors in rat forebrain. Further localization of these receptor subtypes in striatolimbic forebrain should help to clarify their contributions to the pathophysiology of neuropsychiatric disorders and their treatment. PMID- 10723052 TI - Interleukin-6 (IL-6) and its soluble receptor support survival of sensory neurons. AB - The cytokine interleukin-6 (IL-6) has multiple functions in the immune and hematopoietic systems. IL-6 is related to ciliary neurotrophic factor (CNTF), a trophic factor for motoneurons, sensory dorsal root ganglion (DRG) neurons, and other neuronal subpopulations. Both act via related receptor complexes, consisting of one ligand-specific alpha-receptor subunit (IL-6R and CNTFR, respectively) and two signal-transducing receptor components. Even though IL-6 is expressed by neurons and glia, the functions of IL-6 in the nervous system are poorly understood. Here, we report that exogenous human IL-6 promotes the survival of dissociated newborn rat DRG neurons in vitro if supplemented with soluble human IL-6-alpha-receptor. The dosages of human IL-6 and soluble human IL 6R necessary to achieve neurotrophic effects could be reduced markedly by linking ligand and alpha-receptor component in a designer cytokine. Furthermore, we show that newborn rat DRG neurons express and secrete bioactive IL-6. Endogenously secreted IL-6 does not enhance survival of these neurons in vitro, suggesting that DRG neurons do not sufficiently express cell surface IL-6R. Exogenously added soluble rat IL-6R rendered DRG neurons responsive to secreted IL-6. Our results indicate an autocrine function of IL-6 in DRG neuron survival which depends on membrane-bound or soluble IL-6R as a neurotrophic cofactor. PMID- 10723053 TI - Role of nitric oxide in photoreceptor survival in embryonic chick retinal cell culture. AB - The presence of nitric oxide synthase (NOS) in chick retina during development has allowed us to study the role of nitric oxide (NO) during retinal differentiation in dissociated chick retinal cell culture from embryonic day 6. We have demonstrated the presence of nicotinamide adenine dinucleotide phosphate diaphorase staining in these cultures after 3 days in vitro (Div), with a maximal intensity after 8 Div, corresponding to embryonic day 14. Immunohistochemistry studies confirmed the presence of the two isoforms of NOS, NOS-I and -III, in dissociated retinal cell cultures at 8 Div. Addition of NG-monomethyl-L-arginine, a NOS inhibitor, to retinal cell cultures prevented NO production but did not modify the appearance and the survival of ganglion and amacrine cells. However, immunohistochemical analysis with distinct markers for photoreceptor cells (rods and cones) showed that inhibition of endogenous NOS in retinal cell cultures prevented the developmental decrease of rod number between 5 and 8 Div, thus supporting the hypothesis that NO may be involved in the cell death of rods during the development of the retina. PMID- 10723054 TI - Gender differences in protection from EAE induced by oral tolerance with a peptide analogue of MBP-Ac1-11. AB - Mechanisms that contribute to increased female susceptibility to multiple sclerosis can be studied in the murine model of experimental autoimmune encephalomyelitis (EAE). In this report, we compared oral tolerance induction in male and female B10.PL mice using fed myelin basic protein (MBP) Ac1-11 peptide or a high-affinity analogue, Ac1-11[4Y]. We found that fed Ac1-11[4Y] peptide, but not native Ac1-11, could limit cellular infiltration into the central nervous system (CNS) and protect male mice from EAE, an effect that was completely obviated by castration. In contrast, female mice could not be orally tolerized or protected from EAE with either peptide. Tolerance induction in males was reflected by the appearance of Ac1-11[4Y]-reactive splenocytes that produced a sharply increased ratio of transforming growth factor (TGF)-beta:interleukin (IL) 2 and induced bystander suppression. These data directly demonstrate gender differences in regulatory T cells, and support the concept that androgens are involved in governing oral tolerance to EAE. PMID- 10723055 TI - Calcitonin gene-related peptide immunoreactivity in the hippocampus and its relationship to cellular changes following exposure to trimethyltin. AB - Calcitonin gene-related peptide (CGRP) is a neuropeptide that is regionally regulated following peripheral insult and in central nervous system (CNS) damage models targeting limbic structures. Functional studies have shown this neuropeptide to be involved in neuronal protection and remodeling, vasodilation, immunomodulation, and apoptosis, thus making it an important constituent of the acute phase response. In the present study, we characterized the anatomic expression and distribution of CGRP immunoreactivity (CGRP-IR) after exposure to the toxin, trimethyltin (TMT). We chose this model because TMT causes dramatic changes in the endocrine system, the limbic system, particularly the hippocampus, as well as in the immune response. We have specifically focused on comparing the changes in CGRP-IR with the pattern of apoptosis (via TUNEL staining), cell-cycle activation (Ki67-IR), and in alteration in microglia (OX-42-IR) and astrocyte (gGFAP-IR) immunocytochemistry in TMT-treated hippocampus. Our results show a marked change in CGRP-IR in regions of the hippocampus that are temporally and anatomically correlated with the induction of apoptosis and activation of microglia, astrocyte, and the cell-cycle marker. Given the known effects of CGRP on these cell types and on programmed cell death elsewhere, these findings are consistent with a regional immunoregulatory/injury response role for CGRP following organotin poisoning. PMID- 10723056 TI - Organization of point contacts in neuronal growth cones. AB - Growth cones from rat dorsal root ganglia plated on laminin contain integrin clusters over the entire growth cone surface, and growth cones make transient adhesions at sites called point contacts. We examined, by immunocytochemistry and confocal microscopy, the composition and distribution of point contacts in neuronal growth cones. Vinculin was concentrated in the central domain of growth cones and at the tips of filopodia. Vinculin was specifically associated with integrin clusters at the membrane-substrate interface and thus marked point contacts. The cytoskeletal proteins paxillin and talin colocalized with beta1 integrin in a subpopulation of clusters restricted to the central domain of the growth cone and to the tips of filopodia. The neuron-specific kinase, FAK+ also distributed with the vinculin-positive clusters. The Rho family proteins RhoA, RhoB, and Cdc42 were present in growth cones, and a few Rho clusters were colocalized with vinculin. Examination of proteins resistant to detergent extraction in PC12 cells confirmed the retention of beta1 integrin, paxillin, talin, and vinculin with the cytoskeleton. Moreover, we detected FAK+ and RhoA in the detergent-resistant cytoskeleton, supporting their distribution to point contacts. Our observations indicate that two types of integrin clusters are present in growth cones: those associated with vinculin at the cell substratum interface, and those not associated with vinculin. Point contacts are mature adhesion sites defined by the presence of both beta1 integrin and vinculin, and they are associated with signaling proteins. PMID- 10723057 TI - Metabotropic glutamate receptor subtypes independently modulate neuronal intracellular calcium. AB - Metabotropic glutamate receptors (mGluRs) modulate several G-protein-related signal transduction pathways including intracellular calcium (iCa(2+)) that control both neuronal development and demise. As an initial investigation, we characterized the ability of specific mGluR subtypes to modulate iCa(2+) by using Fura-2 microfluorometry in primary hippocampal neurons. Activation rather than inhibition of the metabotropic system with the group I and group II mGluR agonist 1S, 3R-1-aminocyclopentane-1,3-dicarboxylic acid (ACPD), the specific group I agonist (S)-3,5-dihydroxyphenylglycine (DHPG), and the specific group II agonist (2S,1'S,2'S)-2-(carboxycyclopropyl)glycine (LCCG-I) increased iCa(2+) with increasing concentrations. In contrast, the group III mGluR agonist, L(+)-2-amino 4-phosphonobutyric acid (L-AP4) produced no significant increase in iCa(2+). Through the pharmacological modulation of individual mGluR subtypes, we further examined the role of iCa(2+) release by the mGluR system. Release of iCa(2+) by both 1S,3R-ACPD and LCCG-I was prevented only through the administration of the antagonists (2S)-alpha-ethylglutamic acid (EGlu; mGluR2 and mGluR3) and (2S,1'S,2'S,3'R)-2-(2'-carboxy-3'-phenylcyclopropyl)glycine (PCCG-IV; mGluR2), suggesting that the mGluR2 subtype was responsible for the release of iCa(2+). As a control, the group I antagonists, L(+)-2-amino-3-phosphonopropionic acid (L AP3) and (RS)-1-aminoindan-1,5-dicarboxylic acid (AIDA), prevented DHPG release of iCa(2+) but were ineffective against iCa(2+) release by 1S,3R-ACPD. Although extracellular calcium influx did not significantly contribute to the release of iCa(2+) by the mGluR system, pharmacological inhibition of calcium-induced calcium-release-sensitive calcium pools played a critical role in the release of iCa(2+). Further characterization of the cellular calcium pools modulated by the mGluR subtypes may provide greater insight into the mechanisms that mediate neuronal function. PMID- 10723058 TI - Vascularization of human glioma spheroids implanted into rat cortex is conferred by two distinct mechanisms. AB - Aim of this study was to develop and characterize an applicable in vivo model to investigate angiogenesis of human gliomas. An established glioblastoma spheroid model was used to investigate the neovascularization of a standardized avascular solid tumor mass. Spheroids of two human glioma cell lines were labeled with an in vivo fluorescent dye. Single spheroids were implanted into the cortex of athymic rats. After 1, 3, 7, 14, and 21 days, brain sections containing the spheroid were immunostained for endothelial cells or vascular endothelial growth factor (VEGF). The dye-stained glioma spheroid and the endothelial cells were visualized by confocal microscopy. Two distinct mechanisms of tumor vascularization could be observed. (1) "Classical" angiogenesis with new vessels sprouting from existing host vessels into the spheroid was seen. (2) Individual endothelial cells were found to migrate towards and into the center of the spheroid where they coalesced to form new vessels. This process occurred as early as 24 hr after spheroid implantation. Spheroid vascularization was accompanied by an increase of VEGF expression, which peaked 7 days after implantation and returned to normal patterns by 14-21 days. Besides the "classical" angiogenesis by angiogenic blood vessels, the recruitment of individual endothelial cells seems to be an additional mechanism in early glioma vascularization. Our model proves to be a reliable, reproducible system to study in vivo angiogenesis of human gliomas. PMID- 10723059 TI - Glycosaminoglycans treatment increases IGF-I muscle levels and counteracts motor neuron death: A novel nonanticoagulant action. AB - The present study shows that sciatic nerve crush in 2-day-old rats causes extensor digitorum longus (EDL) muscle atrophy and motor neuron loss and that treatment with glycosaminoglycans (GAGs) promotes muscle reinnervation, motor neuron survival, and markedly increases insulin-like growth factor-I (IGF-I) content in the denervated muscles. EDL muscle denervation-induced atrophy in saline-treated rats is progressive and reaches the greatest extent at 42 days after birth, which correlates with reduced EDL weight growth. There is also a partial reinnervation as shown by the number of reinnervated EDL muscle fibers (65.4% of control) and by the poor restoration of the indirect isometric twitch tension (62% of control) that is further reduced under tetanic stimulation (34% of control). The number of surviving motor neurons that innervate EDL muscle drops from 55 +/- 3 to 29 +/- 8. In GAGs-treated 42-day-old rats, the effects of neonatal nerve lesioning on EDL muscle atrophy and denervation are successfully reversed, and the isometric twitch tension and the capacity to hold tetanic stimulation are restored to almost control levels. The number of surviving EDL motor neurons is also increased to 43 +/- 4. Treatment with GAGs selectively affects IGF-I content in denervated hindlimb muscles, which is augmented from 7.02 +/- 0.71 ng/mg tissue to 25.72 +/- 0.7 in the EDL and from 3.2 +/- 0.18 to a robust 211 +/- 9.6 in the soleus. PMID- 10723060 TI - Oligodendrocyte-specific gene expression in mouse brain: use of a myelin-forming cell type-specific promoter in an adeno-associated virus. AB - To explore the feasibility of cell type-specific gene expression in oligodendrocytes as a possible therapeutic approach for demyelinating diseases, the cell specificity, tissue specificity, and duration of gene expression were investigated using recombinant adeno-associated viral vectors (rAAV) carrying a green fluorescence protein (GFP) gene. Recombinant AAV vectors carrying either the myelin basic protein (MBP) promoter (rAAV-MBP-GFP) or the cytomegalovirus (CMV) immediate early promoter (rAAV-CMV-GFP) were semistereotactically injected into the brain of C57BL/6J mice. Injection of the rAAV-MBP-GFP vector into or near the corpus callosum resulted in high levels of GFP expression in white matter regions. Double immunostaining with cell- specific markers proved that these GFP-expressing cells were oligodendrocytes. Injection of the rAAV- MBP-GFP vector into gray matter rarely produced GFP expression. In contrast, injection of the rAAV-CMV-GFP vector resulted in few GFP-expressing cells in the white matter, with most of the GFP-expressing cells being neurons located in the cerebral cortex along the needle track. The expression of the GFP driven by the MBP promoter persisted for at least 3 months. PMID- 10723061 TI - Differential responses of oligodendrocytes to tumor necrosis factor and other pro apoptotic agents: role of ceramide in apoptosis. AB - Staurosporine induced apoptosis in a human oligodendroglioma cell line (HOG), neonatal rat oligodendrocyte (O2A(+)) precursors, and mature rat oligodendrocytes. In all three cell culture systems, the activation of caspase-3 like activity (CPP32) coincided with the increased formation of ceramide from sphingomyelin and the onset of DNA fragmentation. Further, the addition of exogenous C(2)-ceramide induced CPP32 activation and DNA fragmentation in all three culture systems. Raising endogenous ceramide levels by the addition of the ceramidase inhibitor, oleoylethanolamine, enhanced apoptosis in both a time- and concentration-dependent manner. Inhibitors of phosphatidylinositol 3-kinase (wortmannin and LY294002) also induced caspase-3 (CPP32) activation, increased ceramide formation, induced DNA fragmentation, and reduced cell viability. In contrast, cytokines such as tumor necrosis factor-alpha (TNF-alpha) had a differential effect on the three cell cultures. Thus, TNF-alpha (160 ng/ml) induced 70% apoptosis in 24 hr in freshly isolated rat brain O2A(+) precursor cells, 60% apoptosis in 24 hr in a human oligodendroglioma (HOG) cell line, but no apoptosis in mature neonatal rat oligodendrocytes. Interferon-gamma augmented the activation of CPP32 by TNF-alpha in HOG cells and O2A(+) oligodendrocyte precursor cells but had no effect on mature oligodendrocytes. Thus, the death pathway appears to be similar in the three cell lines but the lack of coupling between TNF-alpha receptors and the apoptotic pathway leads to a lack of response to cytokines in mature oligodendrocytes. PMID- 10723062 TI - Effect of oxidant systems on the ubiquitylation of proteins in the central nervous system. AB - Ubiquitin (Ub) modification of different proteins plays an important role in many cellular processes. However, the best studied function of Ub is the labeling of proteins committed to rapid degradation, by an ATP-dependent pathway. We previously found that this pathway is operative in the central nervous system (CNS) of adult rats (Adamo et al. [1994] J. Neurosci. Res. 38:358-364). In the present study, we examined the changes in the capacity to form high-molecular weight Ub protein conjugates (UbPC) and the changes in the production of 2 thiobarbituric acid-reactive substances (TBARS), in the content of protein associated carbonyl groups (PAC), and in the activity of glutamine synthetase produced by in vitro peroxidation of the cell cytosolic proteins and of the mitochondrial fraction isolated from rat brain. Under these experimental conditions, there was an increase in PAC and TBARS in the cytosol, indicating that damage to certain cellular components had occurred. Simultaneously there was a marked increase in UbPC in comparison with the nonoxidized controls. These conjugates showed an active turnover and accumulated when Ub-mediated proteolysis was inhibited. In vitro peroxidation of the mitochondrial fractions resulted in an increase in the production of PAC and in an enhancement in the formation of UbPC. These results demonstrate that the oxidized proteins can be recognized by the ubiquitylating system and that in the CNS the Ub-dependent proteolytic pathway is one of the possible mechanisms involved in the removal of cytosolic and mitochondrial fraction damaged proteins. PMID- 10723063 TI - FGF9: a motoneuron survival factor expressed by medial thoracic and sacral motoneurons. AB - In the nervous system, fibroblast growth factor-9 (FGF9) is produced mainly by neurons. By whole-mount in situ hybridization, on embryonic rat spinal cord, we observed Fgf9 expression in a subpopulation of motoneurons located in the thoracic and sacral regions of the median motor column that innervate the axial muscles. Furthermore, FGF9 prevented death of purified rat and chicken motoneurons in culture in the same concentration range as FGF2. The targets of FGF9 are more restricted than that of the other FGFs, however, because conversely to FGF1 or FGF2, FGF9 had only weak or inexistent survival effects on chicken ciliary neurons or rat DRG. FGF9 may therefore play a role as an autocrine/paracrine survival factor for motoneurons. PMID- 10723064 TI - Down-regulation of mu-opioid receptor expression in rat oligodendrocytes during their development in vitro. AB - In the central nervous system, opioid receptors are found in neurons and also in glial cells. To gain more information on their presence and possibly on their function, we investigated the expression of mu-opioid receptors (MOR) during oligodendroglial cell development in two culture systems. In these models, during the first days, the cells are O-2A bipotential progenitor cells (also called OPCs; oligodendrocyte precursor cells), and then they differentiate into oligodendrocytes, which mature. In the first system, oligodendroglial cells, derived from newborn rat brain hemispheres, are grown in primary culture in the presence of a confluent layer of astrocytes, and they differentiate slowly. In the second, cells are specifically detached from the mixed cultures of the first system and are grown thereafter alone in secondary culture, a condition allowing a rapid cell differentiation. Under both conditions OPCs and immature oligodendrocytes were found to express a high level of MOR mRNA, whereas mature oligodendrocytes did not express it at all. The decrease of MOR expression during oligodendrocyte maturation was progressive, suggesting that it was not a primary effect of differentiation but an indirect secondary effect. Our study also shows that basic fibroblast growth factor (bFGF), which has been claimed by some authors to induce a dedifferentiation of the mature oligodendrocytes, and retinoic acid (RA), which had not been tested before, were not able to restore MOR expression in mature oligodendrocytes. These results indicate that bFGF and RA neither reverse the maturation process nor dedifferentiate the cells. However, RA was found to inhibit almost completely the expression of the myelin basic protein. The main result of this study is that MOR is expressed in progenitors and in immature oligodendrocytes, but not in mature oligodendrocytes. This suggests that MOR could be involved in some developmental process of the cells of the oligodendroglial lineage. PMID- 10723065 TI - Chondroitin sulfates expressed on oligodendrocyte-derived tenascin-R are involved in neural cell recognition. Functional implications during CNS development and regeneration. AB - Tenascin-R (TN-R), an extracellular matrix constituent of the central nervous system (CNS), has been implicated in a variety of cell-matrix interactions underlying axon growth inhibition/guidance, myelination and neural cell migration during development and regeneration. Although most of the functional analyses have concentrated exclusively on the role of the core protein, the contribution of TN-R glycoconjugates present on many potential sites for N- and O glycosylation is presently unknown. Here we provide first evidence that TN-R derived from whole rat brain or cultured oligodendrocytes expresses chondroitin sulfate (CS) glycosaminoglycans (GAGs), i.e., C-4S and C-6S, that are recognized by CS-56, a CS/dermatan sulfate-specific monoclonal antibody. Based on different in vitro approaches utilizing substrate-bound glycoprotein, we found that TN-R linked CS GAGs (1) promote oligodendrocyte migration from white matter microexplants and increase the motility of oligodendrocyte lineage cells; (2) similar to soluble CS GAGs, induce the formation of glial scar-like structures by cultured cerebral astrocytes; and (3) contribute to the antiadhesive properties of TN-R for neuronal cell adhesion in an F3/F11-independent manner, but not to neurite outgrowth inhibition, by mechanism(s) sensitive to chondroitinase or CS 56 treatments. Furthermore, after transection of the postcommissural fornix in adult rat, CS-bearing TN-R was found to be stably upregulated at the lesion site. Our findings suggest the functional impact of TN-R-linked CS on neural cell adhesion and migration during brain morphogenesis and the contribution of TN-R to astroglial scar formation (CS-dependent) and axon growth inhibition (CS independent), i.e., suppression of axon regeneration after CNS injury. PMID- 10723066 TI - Differential distribution of the tetrodotoxin-sensitive rPN4/NaCh6/Scn8a sodium channel in the nervous system. AB - Voltage-gated sodium channels underlie the generation of action potentials in excitable cells. Various sodium channel isoforms have been cloned, functionally expressed and distinguished on the basis of their biophysical properties or differential sensitivity to tetrodotoxin (TTX). In the present study, we have investigated the immunolocalization of the TTX-sensitive sodium channel, rPN4/NaCh6/Scn8a, in discrete areas of the rat nervous system. Thus, in naive animals, PN4 was abundantly expressed in brain, spinal cord, dorsal root ganglia (DRG) and peripheral nerve. The presence of PN4 at the nodes of Ranvier in the sciatic nerve suggests the importance of this sodium channel in peripheral nerve conduction. In addition, the pattern of PN4 immunolabeling was determined in DRG, spinal cord and sciatic nerve in rats subjected to chronic constriction nerve injury (CCI). PMID- 10723067 TI - Expression pattern of galectin-3 in neural tumor cell lines. AB - Galectin-3 is a member of the galectin family of beta-galactoside-specific animal lectins. Here we show that galectin-3 is constitutively expressed in 15 out of 16 glioma cell lines tested, but not by normal or reactive astrocytes, oligodendrocytes, glial O-2A progenitor cells and the oligodendrocyte precursor cell line Oli-neu. Galectin-3 is also expressed by one oligodendroglioma cell line, but not by primitive neuroectodermal tumor and 4 neuroblastoma cell lines tested so far. In all galectin-3 expressing cell lines, the lectin is predominantly, if not exclusively, localized intracellularly and carries an active carbohydrate recognition domain (shown for C6 rat glioma cells). Moreover, in contrast to primary astrocytes, glioma cells do not or only weakly adhere to substratum-bound galectin-3, probably reflecting an unusual glycosylation pattern. Our findings indicate that the expression of galectin-3 selectively correlates with glial cell transformation in the central nervous system and could thus serve as a marker for glial tumor cell lines and glial tumors. PMID- 10723068 TI - RGS7 complex formation and colocalization with the Gbeta5 subunit in the adult rat brain and influence on Gbeta5gamma2-mediated PLCbeta signaling. AB - This study describes the colocalized distribution and dimeric complex formation between RGS7, a GTPase-activating protein for several heterotrimeric Galpha protein families, and the Gbeta5 subunit in the adult rat brain. Confocal dual immunofluorescence labeling studies indicated a broad regional specificity in the cellular coexpression between RGS7 and Gbeta5 within the cerebral cortical layers I and V-VI, hippocampal formation, caudate-putamen, medial habenula, most thalamic nuclei, and cerebellar molecular and granular layers. In all instances, Gbeta1-beta4 immunoreactivities exhibited no observable colocalization with RGS7, despite their widespread codistribution throughout similar neuronal networks. Coimmunoprecipitation studies confirmed the selective protein-protein interaction between RGS7 and Gbeta5 within brain regions that displayed immunohistochemical colocalization. The influence of RGS7 to modulate Gbeta5gamma2-mediated phosphatidyl inositol (PI) production was examined in COS-7-cotransfected cells. In the presence of Gbeta5gamma2 only, intracellular PI accumulation was increased by 25% above basal levels; addition of RGS7 produced no significant alteration in Gbeta5gamma2-mediated PI accumulation. A similar trend was exhibited when full length RGS7 was substituted with an RGS7 construct lacking the Gbeta5-interacting region (G protein gamma-like domain; GGL domain) or with RGS4. In conclusion, RGS7/Gbeta5 dimers occurred within most brain regions in which both proteins were cellularly coexpressed. However, an influence of RGS7 on Gbeta5gamma2-mediated PLCbeta signaling activity was not apparent, athough this was in COS-7 cell transfection studies. PMID- 10723069 TI - Impairment of hippocampal long-term potentiation by Alzheimer amyloid beta peptides. AB - Although it is generally believed that amyloid beta (Abeta) peptides contribute to the pathogenesis of Alzheimer's disease, the precise role of these peptides in the development of memory loss of Alzheimer's disease, has not been fully understood. The present study examined the effect of several synthetic Abeta peptides on long-term potentiation (LTP), a cellular model of learning and memory, in rat hippocampal slices. Brief perfusion of slices with low concentrations (200 nM or 1 microM) of Abeta(1-42), Abeta(1-40) or their active fragment Abeta(25--35) significantly inhibited LTP induction without affecting the basal synaptic transmission and posttetanic potentiation in the dentate medial perforant path. A similar effect of Abeta(25-35) was also observed in the Schaffer collateral-CA1 pathway. When comparing actions of several Abeta variants derived from Abeta(25-35), the N-terminal sequence of Abeta(25-35) was found necessary for inhibiting LTP. In addition, Abeta variants lacking neurotoxic action and aggregating property were also able to block LTP, suggesting that this effect was neurotoxicity independent. Our findings demonstrated that subneurotoxic concentrations of Abeta peptides could strongly suppress long-term synaptic plasticity in the hippocampus. Such an effect might underlie the memory deficits seen in Alzheimer's disease before neuronal cell loss. PMID- 10723070 TI - Broadly altered expression of the mRNA isoforms of FE65, a facilitator of beta amyloidogenesis, in Alzheimer cerebellum and other brain regions. AB - FE65 is a key "adapter" protein that links a multiprotein complex to an intracellular domain of beta-amyloid precursor protein (betaPP). Its overexpression modulates the trafficking of betaPP and facilitates the generation of beta-amyloid (Abeta). FE65 is predominantly expressed in brain tissues. An exon 9-inclusive isoform is exclusively expressed in neurons, and an exon 9 exclusive isoform is only expressed in non-neuronal cells. We quantitated the two isoforms in middle temporal cortex, middle frontal cortex, cerebellar cortex and caudate nucleus of 17 Alzheimer disease (AD) patients, 12 normal controls and 9 non-AD neurodegenerative disease controls by reverse transcription-competitive polymerase chain reaction (RT-cPCR). Expression of the two isoforms was significantly and differentially altered, with a 30-57% decrease in levels of the neuronal form (P < 0.05-0.002) and a 73-135% increase in levels of non-neuronal form (P < 0.02-0.001), in the temporal and frontal cortex of AD brains. These alterations presumably reflect advanced neurodegenerative processes of these regions. Surprisingly, expression of both isoforms was significantly up-regulated by 42-66% in the cerebellar cortex and caudate nucleus of AD brains when compared to normal brains (P < 0.05-0.005). Diffuse Abeta-positive plaques were observed in the cerebellum of these AD subjects but not in the normal controls. Selective up-regulation of only the FE65 neuronal isoform was seen in the cerebellar cortex in association with other neurodegenerative diseases (largely Parkinson's disease). Because FE65 modulates trafficking of betaPP toward the production of Abeta, the up-regulation of FE65 in AD cerebellum may be relevant to the genesis of diffuse plaques. Thus, early biochemical alterations in AD, not complicated by advanced pathology, may be beneficially investigated in the less-affected regions of the brain, such as the cerebellum. PMID- 10723071 TI - Increasing neurite outgrowth capacity of beta-amyloid precursor protein proteoglycan in Alzheimer's disease. AB - Progressive cerebral deposition of beta-amyloid peptide either in blood vessels or around neurites is one of the most important features of Alzheimer's disease (AD). The beta-peptide, known as Abeta or A4, is produced by proteolytic cleavage of the amyloid precursor protein (APP). Two APP processing pathways have been proposed as physiological alternatives; only one of which leads to the production of Abeta or amyloidogenic peptides. However, we have little information regarding these processing pathways in the brain, or on whether posttranslational modifications such as glycosylation affect APP processing in vivo. Furthermore, the physiological function(s) of this protein in nervous tissue remains unclear, although modulatory roles in cell adhesion and neuritic extension have been suggested. It has been reported that APP may be glycosylated as a proteoglycan. We purified this APP population from human brain, and our data indicate that PG APP supports neurite extension of hippocampal neurons. Neurons grown on this substratum showed an increased capacity to elongate neurites and increased neuritic "branching" compared to culture on laminin. These effects were enhanced with PG-APP samples obtained from AD brains. Our results suggest that this APP population may act as a neurite outgrowth and branching promoter and may thus play a role in some pathological conditions. These findings may have significant implications in understanding normal brain development and pathological situations (such as AD). PMID- 10723072 TI - Ultrastructural evidence of calcium involvement in experimental autoimmune gray matter disease. AB - Experimental studies have suggested that increased calcium and inappropriate calcium handling by motoneurons might have a significant role in motoneuron degeneration. To further define the involvement of calcium in motoneuron loss we used the oxalate-pyroantimonate technique for calcium fixation and monitored the ultrastructural distribution of calcium in spinal motoneurons in experimental autoimmune gray matter disease (EAGMD). In cervical and hypoglossal motoneurons from animals with relatively preserved upper extremity and bulbar function, increased calcium precipitates were present in the cytoplasm as well as in mitochondria, endoplasmic reticulum and Golgi complex without significant morphologic alterations. In surviving lumbar motoneurons of animals with hindlimb paralysis, however, there was massive morphological destruction of intracellular organelles but no significant accumulation of calcium precipitates. These findings suggest that altered calcium homeostasis is involved in motoneuron immune-mediated injury with increased calcium precipitates early in the disease process and decreased to absent calcium precipitates later in the pathogenesis of motoneuron injury. PMID- 10723073 TI - Different testosterone metabolism by immortalized embryonic and postnatal hippocampal neurons from C57BL/6 mice: a crucial role for androstenedione. AB - Steroid hormones influence the development of undifferentiated brain during ontogenesis. In the present study we investigated the metabolic pathway of testosterone in immortalized embryonic and postnatal hippocampal neurons from C57BL/6 mice. Both cell lines are capable of metabolizing testosterone to 6alpha hydroxytestosterone, 6beta-hydroxytestosterone and androstenedione. The formation was found to correlate with protein concentration and time of incubation. These linearities were significant for all metabolites except androstenedione that was the main metabolite in embryonic hippocampal neurons and nearly absent in postnatal neurons. Moreover, only embryonic cells react to testosterone with a decrease of beta-tubulin expression, that was a typical effect indicating induced neuronal maturation. Application of androstenedione caused the same decrease of beta-tubulin expression as testosterone did before. Our results of hippocampal testosterone metabolism in vitro confirm that not only estradiol and 5alpha dihydrotestosterone could impact neural tissue but also androstenedione is a powerful metabolite involved in prenatal neuronal differentiation. PMID- 10723074 TI - Nordidemnin potently inhibits inositol uptake in cultured astrocytes and dose dependently augments lithium's proconvulsant effect in vivo. AB - It has been suggested that inositol uptake across the cell membrane is of importance for maintenance of the inositol pool involved in lithium's therapeutic effect in bipolar disease and in the lithium-pilocarpine seizure test in freely moving rats (measuring the latency of a normally subconvulsive concentration of pilocarpine to seizure induction in the additional presence of lithium). We have tested this hypothesis by: 1) demonstrating an extremely high potency of nordidemnin as an inhibitor of myo-inositol uptake in primary cultures of mouse astrocytes; and 2) determining the dose-response correlation of a nordidemnin induced decrease in the latency before appearance of seizures in the lithium pilocarpine test after intracerebroventricular injection of minute samples (10 microl) of virtually isotonic saline solution. PMID- 10723075 TI - Distinct differences in the cannabinoid receptor binding in the brain of C57BL/6 and DBA/2 mice, selected for their differences in voluntary ethanol consumption. AB - The two inbred strains of mice C57BL/6 and DBA/2 mice have been shown to differ significantly in their preference for alcohol (EtOH). These strains of mice have been employed to study various aspects of pharmacological and behavioral effects of EtOH. We have previously demonstrated that chronic EtOH exposure down regulated cannabinoid receptors (CB1) in mouse synaptic plasma membranes and enhanced the synthesis of endogenous cannabimimetic compound anandamide (AnNH) in human neuroblastoma cells. The purpose of the present study was to investigate whether there were differences in the density and the affinity of CB1 receptors in the brains of the two inbred C57BL/6 (alcohol-preferring) and DBA/2 (alcohol avoiding) mice. The results indicate the presence of specific CB1 receptors in the brain membranes of both the strains. It was also found that the CB1 receptor densities (B(max)) were 25% lower in C57BL/6 (0.66 +/- 0.15 pmol/mg protein) compared with that of DBA/2 (0.88 +/- 0.08 pmol/mg protein) mice. Significant differences in the affinity were also observed between the two lines (K(d), 0.68 +/- 0.15 nM for C57BL/6 and 2.21 +/- 0.56 nM for DBA/2). The competition studies with SR141716A, a CB1 receptor antagonist, and 2-arachidonylglycerol (2-AG) and anandamide (AnNH), known CB1 receptor agonists, all showed a substantial decrease in [(3)H]CP-55,940 binding in both strains of mice with a higher K(i) values in the DBA/2 mice. These results suggest that CB1 receptor signal transduction may play an important role in controlling the voluntary EtOH consumption by these strains of mice. PMID- 10723076 TI - Head-capsule design and mandible control in beetle larvae: a three-dimensional approach. AB - This work provides the morphological basis of mandible kinematics of beetle larvae with different types of head capsules, orientation of mouthparts, mandible design, and feeding behavior. Short descriptions are provided for the hypognathous head of Liocola marmorata (Scarabaeidae), the prognathous head of Cybister lateralimarginalis (Dytiscidae), and the hyperprognathous head of Hydrophilus piceus (Hydrophilidae). Using 3D measurements of a set of points selected on the head, 3D models of head capsules and mandible articulations were made for the larvae of these three species. Further calculations of the models showed differences in spatial positions of mandible axes and insertion points of mandible muscles. Functional consequences of these differences and evolutionary aspects of changes in head design and mandible articulations are discussed. It is suggested that the approach used in this study may be useful for further comparative studies of mandible mechanics of a broad variety of insect taxa. PMID- 10723077 TI - Developmental study of the shortened spinal cord in the adult tiger puffer fish, Takifugu rubripes (Teleostei). AB - ABSTRACT The spinal cords of vertebrates are generally divided into the cord proper and the minute filum terminale. While the spinal cord extends the entire length of the vertebral canal in the adult tiger puffer, Takifugu rubripes, the cord proper is greatly reduced in length and almost all of the canal is occupied by the filum terminale, which is tape-like rather than thread-like. The dorsal and ventral roots of the spinal nerves extend, respectively, above and below the filum terminale; as a whole, these form a massive cauda equina. Supramedullary cells are found in the rostral half of the medulla oblongata caudal to the cerebellum. In 4-mm long tiger puffers, the spinal cord is cylindrical and supramedullary cells are found in the rostral half of the cord. In 7-mm puffers, the longitudinally arranged ventral roots appear ventrally in the middle portion of the spinal cord. In 15-mm puffers, the dorsal and ventral roots run longitudinally along the spinal cord and have noticeably increased in number. Supramedullary cells are located in the rostral 15% of the cord. In 21-mm puffers, the spinal cord in large part becomes dorsoventrally flattened. In 30-mm puffers, the spinal cord becomes much flatter, and supramedullary cells now are located mainly in the medulla oblongata. These observations indicate that formation of the shortened spinal cord proper is due to at least two developmental processes. First, the elongation of the spinal cord proper is remarkably less than that of the vertebral canal. Second, the bulk of the spinal cord proper is translocated to the cranial cavity, where it is transformed into part of the medulla oblongata. PMID- 10723078 TI - Peptidergic and aminergic neurotransmitters of the exocrine pancreas of the Houbara bustard (Chlamydotis undulata). AB - The immunochemical distribution of peptidergic and aminergic neurotransmitters in the exocrine pancreas of the Houbara bustard, Chlamydotis undulata, was determined. Immunoreactivity to choline acetyltransferase (ChAT), vasoactive intestinal polypeptide (VIP), and galanin (Gal) occurred mainly as varicose terminals in the walls of capillaries around the acini and arterioles within the connective tissue. Neuronal cell bodies immunoreactive to ChAT were infrequently observed. Neuropeptide Y (NPY), pancreatic polypeptide (PP), and somatostatin (Som) were observed mainly in intra-acinar cell bodies but nerve fibers immunoreactive to these neuropeptides were also seen along the basal surfaces of the acini. Immunoreactivity to NPY and PP was also discernible in cells of the pancreatic ducts. In addition, NPY occurred as varicose terminals in vessels around the ducts. SP occurred rarely in interacinar ganglia. The distribution of tyrosine hydroxylase (TH) was similar to that of ChAT and, in addition, the occasional TH immunoreactive intra-acinar neuronal cell body was observed. Neuronal nitric oxide synthase (nNOS) occurred in neuronal cell bodies among the acinar cells as well as nerve fibers along the bases of the acini. The potential roles of these peptidergic and aminergic neurotransmitters in the neurohormonal control of pancreatic secretion are discussed. PMID- 10723079 TI - Developmental implications of differential effects of calcium in tail and body skin of Anuran tadpoles. AB - The effects of external Ca(++) on metamorphosis of Rana catesbeiana tadpoles were assessed. Treatment of tadpoles with Ca(++) (0.05 mM) during early prometamorphic stages induced precocious metamorphic events such as tail regression, shortening of the intestine, forelimb emergence, and keratinization of body epidermis within 23 days of treatment compared to control tadpoles still in mid-prometamorphic stages. These effects of Ca(++) are probably mediated by the thyroid gland, as indicated by histological features of the gland at the light and electron microscopic levels. Calcium levels of tail and body skin were measured at various stages of development by atomic absorption spectrophotometry. In control and experimental groups, body skin had significantly higher Ca(++) concentrations than tail skin. There were no statistically significant effects of developmental stage on Ca(++) levels of tail or body skin. Experimental Ca(++) treatment significantly increased Ca(++) concentration in tail but not body skin. Ultrastructure studies and gel electrophoresis indicated that calcium induced keratinization of body skin, but not tail epidermis. Ca(++)-treated tail epidermis showed various autolysing figures in apoptotic cells. In summary, calcium treatment accelerated metamorphosis and induced the following region dependent cellular events: keratinization of body skin-a characteristic of adult epidermis-and programmed cell death in the tail. Whatever signal elicited by calcium in this experimentally induced accelerated metamorphosis is probably mediated via the thyroid gland. PMID- 10723080 TI - Ectal mandibular gland in Polistes dominulus (Christ) (Hymenoptera, vespidae): ultrastructural modifications over the secretory cycle. AB - An ultrastructural study was carried out on the secretory activity of the ectal mandibular gland in the wasp Polistes dominulus (foundress and worker females as well as males). Secretory activity in foundresses proceeds slowly during hibernation and early spring, becoming prominent in late spring and then falling sharply during the summer. This sequential pattern of ultrastructural modifications follows a functional, annual cycle. However, by comparing the subcellular changes in the gland with colonial development, it appears that secretory activity fits in with the specie's social cycle rather than merely following the seasons. The highest levels of secretory activity correspond to the early, critical breeding phases, while activity slows down with an increase in colony protection, based on both primary (passive) and secondary (active) defenses, with the emergence of the workers. These correlations suggest that the ectal mandibular gland secretory product in P. dominulus is involved in chemical nest defense. PMID- 10723081 TI - Evolutionary mechanisms of rib loss in anurans: a comparative developmental approach. AB - ABSTRACT The presence of free ribs is presumed to be a primitive morphological character observed only in a few families of Recent anurans, whereas the absence of ribs has been considered to be a derived condition that is widespread within this order. A comparative study of rib development based on representatives of several anuran lineages (Alytes, Bombina, Bufo, Discoglossus, Hyla, Pelobates, Pelodytes, Rana, and Xenopus) reveals a previously undetected diversity of developmental features in the formation and interaction between neural arches and ribs. The absence of free ribs at premetamorphic or later stages is verified in some groups, but we present for the first time evidence of the existence of larval rib rudiments in others, both in the anterior (Rana, Hyla) and posterior (Bufo, Discoglossus, Pelobates) presacral regions. Heterochrony seems to have played a major role in the processes underlying rib reduction. The intracolumnar differences between anterior (V(2)-V(4)) and posterior (V(5)-V(8)) regions are based on perturbations in the timing of early differentiation. Furthermore, a clear shift in the relative timing of ossification among evolutionary lineages was detected. In this respect Xenopus has a highly derived condition. The use of the morphological character of "rib loss" in phylogenetic analyses must be reconsidered due to the different convergent developmental paths described here. The phylogenetic analysis of a "sequence units" matrix of rib development is compared with current anuran phylogenies. Some evolutionary information appears to be clearly present in the ontogenetic data of this "missing morphology," but its value for evolutionary inferences is rather limited. PMID- 10723082 TI - Anatomy of the baculum-corpus cavernosum interface in the norway rat (Rattus norvegicus), and implications for force transfer during copulation. AB - The baculum is a nonappendicular bone found in the glans tissue of members of five orders of mammals. Its function during copulation is unknown. Anatomical examination of the baculum and corpus cavernosum in the Norway rat (Rattus norvegicus) shows that the two structures are connected by a layer of fibrocartilage, and that the distal tip of the corpus cavernosum swells during erection to surround the proximal end of the baculum. Microradiographs of bacula from sexually experienced males show that regions of the bone may be remodeling; these data suggest that the baculum is load-bearing. On the basis of this anatomy, I propose that the baculum increases the overall flexural stiffness of the penis during copulation by transferring bending and compressive forces from the distal end of the glans to the tensile wall of the corpus cavernosum. Forces on the distal end of the penis during copulation press the baculum against the corpus cavernosum, reducing its internal volume and increasing intracavernosal pressure and corpus cavernosum wall strains. Because the wall of the erect corpus cavernosum is reinforced with inextensible collagen fibers, an increase in wall strain will also increase wall tissue stiffness, and thereby increase the flexural stiffness of the corpus cavernosum. PMID- 10723084 TI - A TGF-beta-inducible cell adhesion molecule, betaig-h3, is downregulated in melorheostosis and involved in osteogenesis. AB - Melorheostosis is a rare bone disease characterized by linear hyperostosis and associated soft tissue abnormalities. The skin overlying the involved bone lesion is often tense, shiny, erythematous, and scleodermatous. In order to look for genes differentially expressed between the normal and involved skin, we cultured skin fibroblasts from the skin lesions of several afflicted patients, and identified differentially expressed genes by reverse dot-blot hybridization. We found that the genes human TGF-beta-induced gene product (betaig-h3), osteoblast specific factor 2, osteonectin, fibronectin, and type I collagen were all downregulated in the affected skin fibroblasts, with betaig-h3 the most significantly affected. The expression of betaig-h3 was induced by TGF-beta in both affected and normal fibroblasts. In an effort to determine the mechanism of bone and skin abnormalities in melorheostosis, we made recombinant betaig-h3. Both immobilized and soluble recombinant betaig-h3 proteins with or without an RGD motif inhibited bone nodule formation of osteoblasts in vitro. Taken together, our results suggest that altered expression of several adhesion proteins may contribute to the development of hyperostosis and concomitant soft tissue abnormalities of melorheostosis, with betaig-h3 in particular playing an important role in osteogenesis. PMID- 10723085 TI - Ordered structure acquisition by the N- and C-terminal domains of the small proline-rich 3 protein. AB - The cell envelope (CE) is a vital structure for barrier function in terminally differentiated dead stratified squamous epithelia. It is assembled by transglutaminase (TGase) cross-linking of several proteins, including hSPR3 in certain specialized epithelia normally subjected to mechanical trauma. Biochemical studies show that hSPR3 serves as a complete substrate for TGase1, TGase2, and TGase3. Multiple adjacent glutamines and lysines of only head-and tail domain sequences are used by each enzyme for cross-linking. Structural data suggest that the hSPR3 central repeats, as well as hSPR1 and hSPR 2, are highly flexible and mobile; thus, the TGases might not be able to recognize the residues localized on the repeats as adequate substrate. To investigate this hypothesis further and to complete the structural investigation of hSPR3, we performed circular dichroism (CD) studies on peptides corresponding to the N- and C terminal domain. CD spectra have also been carried out in the presence of different concentrations of the structure-promoting agent cosolvent trifluoroethanol (TFE), which mimics a partial hydrophobic environment found in vivo in or next to the membrane. In fact, this agent increases the dielectric constant of water proportionally, depending on its concentration, and confers structuring properties to the solution, to peptides and proteins that have a structuring propensity. The results indicate that in both the N-terminal and C terminal, peptides acquire a more ordered structure as a function of the TFE concentration in water. This ability of both N- and C-terminal domain to acquire a more stable ordered conformation might be relevant for SPR3 to act as substrate of TGases. Indeed, only the N- and C-terminus is cross-linked by TGase1 and 3. PMID- 10723086 TI - Transforming growth factor-beta1 expression in cultured corneal fibroblasts in response to injury. AB - The mechanisms underlying TGF-beta regulation in response to injury are not fully understood. We have developed an in vitro wound model to evaluate the expression and localization of transforming growth factor-beta1 in rabbit corneal fibroblasts in response to injury. Experiments were conducted in the presence or absence of serum so that the effect of the injury could be distinguished from exogenous wound mediators. Cultures were wounded and evaluations conducted over a number of time points. Expression of TGF-beta1 RNA was determined using Northern blot analysis and in situ hybridization, while the TGF-beta receptors were identified by affinity cross-linking. Injury increased the expression of TGF beta1 mRNA in cells at the wound edge after 30 min; this response was amplified by the addition of serum. TGF-beta1 mRNA expression was observed in a number of cells distal from the wound. After wound closure, TGF-beta1 mRNA was negligible and resembled unwounded cultures. The half-life of TGF-beta1 mRNA was two times greater in the wounded cultures, indicating that the injury itself maintained the expression, while cell migration was present. Analogous to these findings, we found that binding of TGF-beta to its receptors was maximal at the wound edge, decreasing with time and distance from the wound. These results indicate that injury increases the level of expression of TGF-beta1 mRNA and maintains a higher level of receptor binding during events in wound repair and that these might facilitate the migratory and synthetic response of stromal fibroblasts. PMID- 10723088 TI - Inhibition of neointima formation by a nonpeptide alpha(v)beta(3) integrin receptor antagonist in a rabbit cuff model. AB - This study was performed to determine whether a highly selective nonpeptide alpha(v)beta(3) antagonist (SH306) would prove effective in inhibiting neointima formation in a rabbit cuff model. The animals were dosed with SH306, 5 mg/kg i.v., followed by 10 mg/kg s. c., 3 times daily for 3 days, or with vehicle (10% DMAC). Rabbits were sacrificed and perfused on days 1, 3, and 21; the vessels were paraffin embedded. A reduction in the intima/media (I/M) of the SH306 treated rabbits, as compared with the vehicle-treated control group, was noted (0.20 vs 0.36 [n = 4]). A significant increase in the area of the media was observed in the SH306-treated group versus the control group (0.20 vs 0.13). No difference was observed in cell proliferation between SH306 and vehicle after 1 day and 3-day dosing. Thrombi were found in 43% of the control vessels and in only 14% of the drug-treated vessels. No anticoagulant was used during the surgical procedure. No increase in inhibition of GPIIb/IIIa was observed in SH306 treated animals, as compared with the vehicle control group. We conclude that selective inhibition of alpha(v)beta(3) reduced neointima formation in a rabbit model at 3 weeks. PMID- 10723087 TI - Role of protein kinase C in 1,25(OH)(2)-vitamin D(3) modulation of intracellular calcium during development of skeletal muscle cells in culture. AB - Regulation of muscle cell Ca(2+) metabolism by 1, 25-dihydroxy-vitamin D(3) [1,25(OH)(2)D(3)] is mediated by the classic nuclear mechanism and a fast, nongenomic mode of action that activates signal transduction pathways. The role of individual protein kinase C (PKC) isoforms in the regulation of intracellular Ca(2+) levels ([Ca(2+)](i)) by the hormone was investigated in cultured proliferating (myoblasts) and differentiated (myotubes) chick skeletal muscle cells. 1,25(OH)(2)D(3) (10(-9) M) induced a rapid (30- to 60-s) and sustained (>5 min) increase in [Ca(2+)](i) which was markedly higher in myotubes than in myoblasts. The effect was suppressed by the PKC inhibitor calphostin C. In differentiated cells, PKC activity increased in the particulate fraction and decreased in cytosol to a greater extent than in proliferating cells after 5-min treatment with 1,25(OH)(2)D(3). By Western blot analysis, these changes were correlated to translocation of the PKC alpha isoform from cytosol to the particulate fraction, which was more pronounced in myotubes than in myoblasts. Specific inhibition of PKC alpha activity using antibodies against this isoform decreased the 1, 25(OH)(2)D(3)-induced [Ca(2+)](i) sustained response associated with Ca(2+) influx through voltage-dependent calcium channels. Neomycin, a phospholipase C (PLC) inhibitor, blocked its effects on [Ca(2+)](i), PKC activity, and translocation of PKC alpha. Exposure of myotubes to 1,2-dioleyl-rac glycerol (1,2-diolein), also increased [Ca(2+)](i), PKC activity, and the amount of PKC alpha associated with the particulate fraction. Changes in [Ca(2+)](i) induced by diolein were inhibited by calphostin C and nifedipine. The results indicate that PKC alpha activation via PLC-catalyzed phosphoinositide hydrolysis is part of the mechanism by which 1, 25(OH)(2)D(3) regulates muscle intracellular Ca(2+) through modulation of the Ca(2+) influx pathway of the Ca(2+) response to the sterol. PMID- 10723089 TI - Activation of a rel-A/CEBP-beta-related transcription factor heteromer by PGG glucan in a murine monocytic cell line. AB - PGG-Glucan is a soluble beta-glucan immunomodulator that enhances a variety of leukocyte microbicidal activities without activating inflammatory cytokines. Although several different cell surface receptors for soluble (and particulate) beta-glucans have been described, the signal transduction pathway(s) used by these soluble ligands have not been elucidated. Previously we reported that PGG Glucan treatment of mouse BMC2.3 macrophage cells activates a nuclear factor kappa-B-like (NF-kappaB) transcription factor complex containing subunit p65 (rel A) attached to an unidentified cohort. In this study, we identify the cohort to be a non-rel family member: a CCAAT enhancer-binding protein-beta (C/EBP-beta) related molecule with an apparent size of 48 kDa, which is a different protein than the previously identified C/EBP-beta p34 also present in these cells. C/EBP beta is a member of the bZIP family whose members have previously been shown to interact with rel family members. This rel/bZIP heteromer complex activated by PGG-Glucan is different from the p65/p50 rel/rel complex induced in these cells by lipopolysaccharide (LPS). Thus, our data demonstrate that PGG-Glucan uses signal transduction pathways different from those used by LPS, which activates leukocyte microbicidal activities and inflammatory cytokines. We further show that heteromer activation appears to use protein kinase C (PKC) and protein tyrosine kinase (PTK) pathways, but not mitogen-activated protein kinase p38. Inhibitor kappa-B-alpha (IkappaB-alpha) is associated with the heteromer; this association decreases after PGG-Glucan treatment. These data are consistent with a model whereby treatment of BMC2.3 cells with PGG-Glucan activates IkappaB-alpha via PKC and/or PTK pathways, permitting translocation of the rel-A/CEBP-beta heteromer complex to the nucleus and increases its DNA-binding affinity. PMID- 10723090 TI - Regulation of amino acid-sensitive TOR signaling by leucine analogues in adipocytes. AB - In adipocytes, amino acids stimulate the target of rapamycin (TOR) signaling pathway leading to phosphorylation of the translational repressor, eIF-4E binding protein-I (4E-BP1), and ribosomal protein S6. L-leucine is the primary mediator of these effects. The structure-activity relationships of a putative L-leucine recognition site in adipocytes (LeuR(A)) that regulates TOR activity were analyzed by examining the effects of leucine analogues on the rapamycin-sensitive phosphorylation of the translational repressor, eIF-4E binding protein-I (4E BP1), an index of TOR activity. Several amino acids that are structurally related to leucine strongly stimulated 4E-BP1 phosphorylation at concentrations greater than the EC(50) value for leucine. The order of potency was leucine > norleucine > threo-L-beta-hydroxyleucine approximately Ile > Met approximately Val. Other structural analogues of leucine, such as H-alpha-methyl-D/L-leucine, S-(-)-2 amino-4-pentenoic acid, and 3-amino-4-methylpentanoic acid, possessed only weak agonist activity. However, other leucine-related compounds that are known agonists, antagonists, or ligands of other leucine binding/recognition sites did not affect 4E-BP1 phosphorylation. We conclude from the data that small lipophilic modifications of the leucine R group and alpha-hydrogen may be tolerated for agonist activity; however, leucine analogues with a modified amino group, a modified carboxylic group, charged R groups, or bulkier aliphatic R groups do not seem to possess significant agonist activity. Furthermore, the leucine recognition site that regulates TOR signaling in adipocytes appears to be different from the following: (1) a leucine receptor that regulates macroautophagy in liver, (2) a leucine recognition site that regulates TOR signaling in H4IIE hepatocytes, (3) leucyl tRNA or leucyl tRNA synthetase, (4) the gabapentin-sensitive leucine transaminase, or (5) the system L-amino acid transporter. PMID- 10723091 TI - Upregulation of myogenin by N-cadherin adhesion in three-dimensional cultures of skeletal myogenic BHK cells. AB - Cells of the baby hamster kidney (BHK) line express the skeletal muscle determining transcription factor MyoD but fail to differentiate. Unlike most skeletal myogenic cells, which express multiple members of the cadherin family of cell-cell adhesion proteins, the BHK cells lack a robust cadherin adhesion system. We previously published that forced expression of N- (or E)-cadherin in BHK cells increases the level of endogenous catenins, mediates strong cell-cell adhesion, and enhances differentiation of BHK cells induced to differentiate by placing them in three-dimensional (3-D) culture (Redfield et al. [1997] J. Cell. Biol. 138:1323-1331). This report demonstrates that N-cadherin adhesion upregulates the protein level of nuclear myogenin in cells induced to differentiate by 3-D culture. Myogenin is a transcription factor required for differentiation of skeletal muscle. It was not detected in monolayer culture, whether the cells expressed N-cadherin or not, nor was it upregulated in 3-D cultures of cells lacking N-cadherin. The activity of two myogenin chloramphenicol acetyltransferase (CAT) reporter constructs containing 3.7 or 1.1 kb upstream regulatory region of the mouse myogenin gene was increased significantly in N-cadherin-expressing cells induced to differentiate by 3-D culture. Our observations indicate that N-cadherin adhesion stimulates skeletal myogenesis by upregulating myogenin. PMID- 10723092 TI - Regulation of alphaVbeta3 and alphaVbeta5 integrins by dexamethasone in normal human osteoblastic cells. AB - Long-term administration of pharmacological doses of glucocorticoids inhibits bone formation and results in osteoporosis. Since integrin-mediated cell-matrix interactions are essential for osteoblast function, we hypothesized that the detrimental effect of glucocorticoids on bone derived, at least in part, from decreased integrin-matrix interactions. Because alphavbeta3 and alphavbeta5 integrins can interact with several bone matrix proteins, we analyzed the effects of dexamethasone (Dex) on the expression of these integrins in normal human osteoblastic cells. We found adhesion of these cells to osteopontin and vitronectin to be dependent on alphavbeta3 and alphavbeta5, respectively; this ligand specificity was not altered by Dex. The effects of Dex on the adhesion of human osteoblastic cells to osteopontin and vitronectin were biphasic with an increase after 2 days, followed by a decrease after 8 days of treatment. Consistently, surface alphavbeta3 and alphavbeta5 integrins, which were increased after 2 days of Dex treatment, were decreased after 8 days. Similarly, total cellular alphav, beta3, and beta5 proteins, which were increased by Dex early in the culture, were diminished after 8 days. Metabolic labeling studies indicated that Dex exhibited biphasic regulation on the biosynthesis of alphavbeta5, with stimulation observed during the second day of treatment, followed by inhibition during the 8th day of exposure. By contrast, the biosynthesis of alphavbeta3 was inhibited by Dex on day 1 and remained inhibited on day 8. Analysis of the mRNA indicated that alphav and beta5 levels were increased by Dex during early exposure (1-3 days), followed by inhibition after prolonged exposure (>/=7 days). By contrast, Dex decreased beta3 mRNA level at all the time points analyzed. Consistently, Dex decreased beta3 promoter activity after 1 day and persisted over 8-day period. By contrast, Dex stimulated beta5 promoter activity after 1 or 2 days but had no effect after 8 days. To further evaluate mechanism(s) leading to the decreased integrin expression after prolonged Dex treatment, mRNA stability was analyzed. Dex was found to accelerate the degradation of alphav, beta3 and beta5 mRNA after an 8-day treatment. Thus, the regulation of alphavbeta3 was dependent on transcription and posttranscriptional events whereas the expression of alphavbeta5 was dependent mainly on posttranscriptional events after prolonged Dex treatment. In conclusion, Dex exhibited time-dependent regulation on the expression of alphavbeta3 and alphavbeta5 integrins in normal human osteoblastic cells. Short-term exposure to Dex increased the levels of alphavbeta3 and alphavbeta5 on the surface and cell adhesion to osteopontin and vitronectin whereas long-term exposure to Dex decreased the expression of both integrins and inhibited the cell adhesion to matrix proteins. PMID- 10723093 TI - C-terminal variations in beta-thymosin family members specify functional differences in actin-binding properties. AB - Mammalian cells express several isoforms of beta-thymosin, a major actin monomer sequestering factor, including thymosins beta4, beta10, and beta15. Differences in actin-binding properties of different beta-thymosin family members have not been investigated. We find that thymosin beta15 binds actin with a 2.4-fold higher affinity than does thymosin beta4. Mutational analysis was performed to determine the amino acid differences in thymosin beta15 that specify its increased actin-affinity. Previous work with thymosin beta4 identified an alpha helical domain, as well as a conserved central motif, as crucial for actin binding. Mutational analysis confirms that these domains are also vital for actin binding in thymosin beta15, but that differences in these domains are not responsible for the variation in actin-binding properties between thymosins beta4 and beta15. Truncation of the unique C-terminal residues in thymosin beta15 inhibits actin binding, suggesting that this domain also has an important role in mediating actin-binding affinity. Replacement of the 10 C-terminal amino acids of thymosin beta15 with those of thymosin beta4 did, however, reduce the actin binding affinity of the hybrid relative to thymosin beta15. Similarly, replacement of the thymosin beta4 C-terminal amino acids with those of thymosin beta15 led to increased actin binding. We conclude that functional differences between closely related beta-thymosin family members are, in part, specified by the C-terminal variability between these isoforms. Such differences may have consequences for situations where beta-thymosins are differentially expressed as in embryonic development and in cancer. PMID- 10723094 TI - Characterization of the inhibition of DNA synthesis in proliferating mink lung epithelial cells by insulin-like growth factor binding protein-3. AB - Insulin-like growth factor binding protein-3 (IGFBP-3) can inhibit cell growth by directly interacting with cells, as well as by forming complexes with IGF-I and IGF-II that prevent their growth-promoting activity. The present study examines the mechanism of inhibition of DNA synthesis by IGFBP-3 in CCL64 mink lung epithelial cells. DNA synthesis was measured by the incorporation of 5-bromo-2' deoxyuridine, using an immunocolorimetric assay. Recombinant human IGFBP-3 (rh[N109D,N172D]IGFBP-3) inhibited DNA synthesis in proliferating and quiescent CCL64 cells. Inhibition was abolished by co-incubation of IGFBP-3 with a 20% molar excess of Leu(60)-IGF-I, a biologically inactive IGF-I analogue that binds to IGFBP-3 but not to IGF-I receptors. DNA synthesis was not inhibited by incubation with a preformed 1:1 molar complex of Leu(60)-IGF-I and IGFBP-3, indicating that only free IGFBP-3 inhibits CCL64 DNA synthesis. Inhibition by IGFBP-3 is not due to the formation of biologically inactive complexes with free IGF, since endogenous IGFs could not be detected in CCL64 conditioned media; any IGFs that might have been present could only have existed in inactive complexes, since endogenous IGFBPs were present in excess; and biologically active IGFs were not displaced from endogenous IGFBP complexes by Leu(60)-IGF-I. After incubation with CCL64 cells, (125)I-IGFBP-3 was covalently cross-linked to a major thick similar400-kDa complex. This complex co-migrated with a complex formed after incubation with (125)I-labeled transforming growth factor-beta (TGF-beta) that has been designated the type V TGF-beta receptor. (125)I-IGFBP-3 binding to the thick similar400-kDa receptor was inhibited by co-incubation with unlabeled IGF-I or Leu(60)-IGF-I. The ability of Leu(60)-IGF-I to decrease both the inhibition of DNA synthesis by IGFBP-3 and IGFBP-3 binding to the thick similar400-kDa receptor is consistent with the hypothesis that the thick similar400-kDa IGFBP-3 receptor mediates the inhibition of CCL64 DNA synthesis by IGFBP-3. PMID- 10723095 TI - Alteration of proteoglycan synthesis in human lung fibroblasts induced by interleukin-1beta and tumor necrosis factor-alpha. AB - Important constituents of extracellular matrix are collagen, fibronectin, hyaluronan, and various types of proteoglycans, such as versican, perlecan, decorin, and biglycan. Remodeling of extracellular matrix occurs continuously and is affected by various cytokines. The aim of this study was to investigate how interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha), separately and in combination, alter fibroblast proliferation, as well as the production of extracellular matrix molecules produced by human fibroblasts from lung. Fibroblast proliferation was inhibited significantly by all treatments, by 12% with IL-1beta and by 16% with TNF-alpha. The combination of IL-1beta and TNF alpha increased the inhibition further, by 27%. Hyaluronan production was increased by all treatments: 1.7-fold by IL-1beta and 1.5-fold by TNF-alpha. The combination of the two gave a further increase (2.5-fold). Similarly, the production of total proteoglycans was increased. The small proteoglycans, decorin, and biglycan, were regulated differently. Decorin production was inhibited by about 34% by all treatments, while biglycan was upregulated 1.3-fold by TNF-alpha. Versican was upregulated by IL-1beta (1.7-fold), whereas TNF-alpha was without effect. Perlecan was mostly unaffected. The changes in protein production of the various proteoglycans were due to increased transcription, as mRNA levels were also changed to the same extent. Synthesis of mRNA for collagen type I was inhibited by up to 75% with the IL-1beta/TNF-alpha combination. The separate cytokines also decreased the level of collagen type I mRNA, but to a lesser extent: 60% with IL-1beta and 40% with TNF-alpha. In summary, our study indicates that these proinflammatory cytokines affect the regulation of extracellular matrix production, which is of importance for the inflammatory process. PMID- 10723096 TI - Insulin-induced redistribution of the insulin-like growth factor II/mannose 6 phosphate receptor in intact rat liver. AB - The ability of acute insulin treatment to elicit a redistribution of the liver insulin-like growth factor-II/ mannose 6-phosphate (IGF-II/M6P) receptor has been studied in rats, using cell fractionation. Injection of insulin (0.4-50 microg) led to a time- and dose-dependent decrease in IGF-II binding activity in Golgi endosomal (GE) fractions, along with an increase in activity in the plasma membrane (PM) fraction; only receptor number was affected. Quantitative subfractionation of the microsomal fraction on sucrose density gradients showed that IGF-II binding activity distributed similarly to galactosyltransferase (a Golgi marker), at slightly higher densities than in vivo internalized (125)I insulin, and at lower densities than 5' nucleotidase and alkaline phosphodiesterase (two plasma membrane markers). Insulin treatment led to a slight time-dependent and reversible shift of IGF-II binding activity toward higher densities. Subfractionation of the GE fraction on Percoll gradients showed that IGF-II binding activity was broadly distributed, with about 60% at low densities coinciding with galactosyltransferase and early internalized (125)I insulin and with 40% at high densities in the region of late internalized (125)I insulin. Insulin treatment caused a time-dependent and reversible shift of the distribution of IGF-II binding activity toward low densities. On SDS-PAGE, the size of the affinity-labeled IGF-II/M6P receptor was comparable in GE and PM fractions (about 255 kDa), but on Western blots receptor size was slightly lower in the latter (245 kDa) than in the former (255 kDa). Insulin treatment did not affect the size, but modified the abundance of the IGF-II/M6P receptor in a manner similar to that of IGF-II binding. In vivo chloroquine treatment fully suppressed the changes in IGF-II binding activity in liver GE and PM fractions observed in insulin-treated rats. We conclude that insulin elicits a time dependent and reversible redistribution of liver IGF-II receptors from Golgi elements and endosomes to the plasma membrane, presumably via early endosomes. PMID- 10723097 TI - ATP-stimulated c-fos and zif268 mRNA expression is inhibited by chemical hypoxia in a rat brain-derived type 2 astrocyte cell line, RBA-2. AB - The stimulus-transcriptional coupling during ischemia/hypoxia was examined for ATP-stimulated expression of immediate early genes (IEGs; c-fos, zif268, c-myc and nur77) in a rat brain-derived type 2 astrocyte cell line, RBA-2. Incubation of cells with 1 mM of extracellular ATP stimulated time-dependent expression of c fos and zif268. ATP induced the largest increases in zif268 mRNA and a lesser one in c-fos mRNA. ATP also induced a slight increase in nur77 mRNA but was ineffective in inducing c-myc expression in these cells. Brief exposure of cells to potassium cyanide to simulate chemical hypoxia induced 9-fold and 7-fold transient increases in c-fos and zif268 expression, respectively, but did not affect c-myc or nur77 expression. When cyanide and ATP were added together, the expression of c-fos and zif268 expression was inhibited, and the effect was mimicked by simulating chemical hypoxia with sodium azide. To elucidate the mechanism involved, the effect of cyanide on ATP-stimulated increases in intracellular Ca(2+) concentrations, [Ca(2+)](i), and phospholipase D (PLD) activities were measured. Cyanide induced an increase in [Ca(2&plus);](i) and further enhanced the ATP-stimulated increases in [Ca(2+)](i) and PLD activities. Nevertheless, metabolic inhibitor, iodoacetate, blocked the ATP-induced c-fos and partially inhibited zif268 expression, and deprivation of cells with glucose also inhibited the ATP-induced c-fos expression. Taken together, these results demonstrate that both extracellular ATP and chemical hypoxia induce c-fos and zif268 expression in RBA-2 type 2 astrocytes. The chemical hypoxia inhibited ATP stimulated c-fos and zif268 expression is not due to alterations in Ca(2+) and PLD signaling, and is at least partially related to metabolic disturbance in these cells. PMID- 10723099 TI - Introduction to international perspectives on therapeutic jurisprudence AB - Therapeutic Jurisprudence (TJ), a concept first conceived by law professors David Wexler (University of Puerto Rico and University of Arizona) and Bruce Winick (University of Miami) only a little more than a decade ago, has emerged as the leading conceptual perspective in the mental health law field. Indeed, a LEXIS search reveals well over 150 articles on, or citing to, therapeutic jurisprudence in American law review publications in the past decade (terms used were "therapeutic w/5 jurisprudence w/25 Wexler or Winick"). Copyright 1999 John Wiley & Sons, Ltd. PMID- 10723098 TI - Methylglyoxal induces apoptosis in Jurkat leukemia T cells by activating c-Jun N terminal kinase. AB - Methylglyoxal (MG) is a physiological metabolite, but it is known to be toxic, inducing stress in cells and causing apoptosis. This study examines molecular mechanisms in the MG-induced signal transduction leading to apoptosis, focusing particularly on the role of JNK activation. We first confirmed that MG caused apoptosis in Jurkat cells and that it was cell type dependent because it failed to induce apoptosis in MOLT-4, HeLa, or COS-7 cells. A caspase inhibitor, Z-DEVD fmk, completely blocked MG-induced poly(ADP-ribose)polymerase (PARP) cleavage and apoptosis, showing the critical role of caspase activation. Inhibition of JNK activity by a JNK inhibitor, curcumin, remarkably reduced MG-induced caspase-3 activation, PARP cleavage, and apoptosis. Stable expression of the dominant negative mutant of JNK also protected cells against apoptosis notably, although not completely. Correspondingly, loss of the mitochondrial membrane potential induced by MG was decreased by the dominant negative JNK. These results confirmed a crucial role of JNK working upstream of caspases, as well as an involvement of JNK in affecting the mitochondrial membrane potential. PMID- 10723100 TI - Therapeutic jurisprudence and the resolution of value conflicts: what we can realistically expect, in practice, from theory. AB - This article assesses the criticisms of therapeutic jurisprudence that it cannot resolve value conflicts, especially between autonomy rights and therapeutic values, or, less radically, that it has not provided a general method for resolving conflicts. Grounded in general jurisprudential principles about conflict resolution, including novel developments respecting the meaning of weighing and balancing, the article rejects the criticisms as unfounded. The article also develops and critiques arguments maintaining that therapeutic jurisprudence cannot resolve certain value conflicts because the values are incommensurable. The argument is illustrated by examples concerning the right to refuse treatment, and jurisprudential analyses of that right. PMID- 10723101 TI - Therapeutic jurisprudence: integrated inquiry and instrumental prescriptions. AB - This paper considers the manner in which one could apply a therapeutic jurisprudence (TJ) analysis to a controversial approach to the treatment of sex offenders. Rather than attempting to resolve the difficult questions regarding this form of intervention, however, this paper uses this issue as an opportunity to examine some theoretical concerns about the nature and limits of the TJ research program. As interpreted here, TJ consists of a program of scholarship and law reform intended to promote integration of conceptual, empirical, and normative inquiry. TJ does not in itself resolve normative issues, but it promotes integrated analysis that can sharpen our ability to articulate and examine those normative concerns. TJ remains normatively neutral at the levels of value and theoretical neutrality. Prescriptively, it cannot generate principled normative prescriptions, but it can generate instrumental prescriptions. The long term measure of the TJ program rests upon its success in promoting innovative and integrated interdisciplinary scholarship and law reform. PMID- 10723102 TI - State of the nation: therapeutic jurisprudence and the evolution of the right of self-determination in international law. AB - This article expands the scope of the therapeutic jurisprudence enterprise and applies the concept at a collective global level. The right of self determination, arguably the most important and certainly the most controversial part of international law, is examined through the lens of therapeutic jurisprudence. By detailing the manner in which nations move towards their goal of statehood, this article opens up dialogue about collective healing, shared memory and alternative approaches to autonomy. The article poses the question of whether groups of people can share in common delusions, forms of folie a gens. PMID- 10723103 TI - From status to contract: a future for mental health law. AB - Given the many complex issues and interests involved it is hardly surprising that mental health is a controversial topic. The law has a critical role in protecting interests and balancing claims. But the law is a major part of the problem. Indeed analysis of its role in producing anti-therapeutic outcomes led to the development of therapeutic jurisprudence. And we keep changing the law; for example on 16 October, 1999 the British Government announced proposals for radical reform of the mental health laws of England and Wales which, in the current form, were heavily influenced by developments in the United States of America. A major problem, it is submitted, is that we do not have an agreed model of the law, upon which to develop an appropriate structure for the delivery of mental health services. Currently we emphasis 'status', which puts a premium upon drawing distinctions which it is difficult, if not unrealistic, to achieve in practice. Another approach would be to emphasize the centrality of the relationship between the patient, or client, and the inter-disciplinary team of people providing him or her with services. This approach, outlined and argued for in this article, would involve developing the law of contract. PMID- 10723104 TI - Should victim impact influence sentences? Understanding the community's justice reasoning. AB - Victim impact statements have been introduced in response to growing community concern about apparent neglect of victims in the criminal justice system. Their use in sentencing is a contentious issue, because victim characteristics such as resilience or fragility can contribute to impacts. Is it appropriate for sentences to be influenced by consequences arising from chance victim circumstances unforeseeable by the offender? In the interest of achieving an optimal fit between the justice system and community expectations, this research examined a neglected question: how does the public reason about the issue? Using offense vignettes presented to 260 people in Western Australia, sentencing decisions were found to vary according to consequences arising from victim characteristics. There was little to indicate participants fully appreciated the issue; thus, further research is needed to clarify how justice reasoning principles are used, and to ascertain whether different decisions are taken when people are informed about the problem. PMID- 10723105 TI - Use of conditional and marginal odds-ratios for analysing familial aggregation of binary data. AB - We propose a method for measuring familial aggregation of binary outcomes that incorporates unconditional, i.e., marginal, odds-ratios, for measuring association between outcomes within generations while using conditional odds ratios to measure association between generations. This method may be used together with marginal odds-ratio models to explore the strength of association between individuals within a generation while accounting for the outcomes of predecessors. Interpretation of resulting odds-ratios is straightforward and a number of features of the method are discussed. An example, using smoking status data from Busselton Health Study families, is used to illustrate our approach. PMID- 10723106 TI - Frequency and allelic association of common variants in the lipoprotein lipase gene in different ethnic groups: the Wandsworth Heart and Stroke Study. AB - The lower serum triglyceride (Tg), higher high density cholesterol (HDL-C) levels and low coronary heart disease (CHD) mortality in black populations, contrast with that in whites. By comparison, South Asian populations display a higher mortality from CHD associated with increased Tg and low HDL-C levels. Lipoprotein lipase (LPL) plays a major role in Tg metabolism. To determine if variation in the LPL gene contributes to the differences in lipid levels, we studied the frequencies and allelic associations of five common variants in the lipoprotein lipase (LPL) gene (-93T/G, D9N, N291S, S447X, and the HinddIII RFLP in intron 8) with serum Tg and HDL-cholesterol concentrations in population samples of middle aged men and women of whites, South Asians, and blacks of African origin co resident in South London. Significantly higher frequencies of the H(-) (P < 0.00001), N9 (P < 0.001), and -93G (P < 10(-10)) alleles were seen in blacks compared to the other two groups. Allelic association between -93G and N9, and H(+) and X447 was strong in all three groups. However, no association was observed between serum Tg and HDL-cholesterol concentrations and these variants in the three ethnic groups. A single common polymorphism in the LPL gene is unlikely to account for the differences in fasting serum Tg in populations of different ethnic background. The importance of the differences in frequencies and the mechanism(s) whereby these may contribute towards a beneficial LPL genotype in black populations remain to be determined. PMID- 10723108 TI - Family risk score of coronary heart disease (CHD) as a predictor of CHD: the Atherosclerosis Risk in Communities (ARIC) study and the NHLBI family heart study. AB - Family history of coronary heart disease (CHD) has been found to be a risk factor for CHD in numerous studies. Few studies have addressed whether a quantitative measure of family history of CHD (family risk score, FRS) predicts CHD in African Americans. This study assessed the association between FRS and incident CHD of participants, and the variation of the association by gender and race. Participants in the study were a biracial population-based cohort with 3,958 African Americans and 10,580 Whites aged 45-64 years old in the ARIC baseline survey (1987-1989). They were randomly selected from four U. S. communities. During follow-up (1987-1993), 352 participants experienced the onset of CHD. Incidence density of CHD (per 1,000 person-years) was 7.8 and 3.6 among African American men (AAM) and women (AAW), and 7.2 and 2.2 among White men (WM) and women (WW). The hazard rate ratio (HRR) of CHD associated with one standard deviation increase of FRS was 1.52 in AAW, 1.46 in AAM, 1.41 in WW, and 1.68 in WM. The HRRs decreased 4.6% in AAW, 1.4% in WW, 5.7% in AAM, and 3.0% in WM, but increased 2.1% in AAM after adjustment for selected covariates. FRS predicts incident CHD in African Americans and Whites, men and women. The relation of FRS to incident CHD can be only partially explained by the selected risk factors in the biological causal pathways: IMT, T-G, LDL, HDL, Lp(a), fibrinogen and hypertension. No significant difference by race has been found in this study. PMID- 10723107 TI - Ascertainment bias in the estimation of sibling genetic risk parameters. AB - The sibling recurrence risk, sibling relative risk, and locus-specific sibling relative risk are fundamental quantities in genetic epidemiologic research and are often estimated without accounting for the sampling scheme. For data generated under some genetic models, bias of estimates may be large if the sampling method is incorrectly modeled. In this paper, we explore the relationship between ascertainment of sibships and estimation and interpretation of genetic risk parameters. In particular, we observe that, although traditional definitions of these population parameters are consistent with each other, implied assumptions about ascertainment and the nature of ascertainment correction differ. In the absence of ascertainment correction, unbiased estimation of sibling recurrence risk and overall sibling relative risk requires single ascertainment, while unbiased estimation of locus-specific sibling relative risk requires complete ascertainment. PMID- 10723110 TI - Effect of physical activity on lipid levels in a population-based sample of men with and without the Arg192 variant of the human paraoxonase gene. AB - The prevalence of cardiovascular risk factors in Gerona, Spain, is high for the low myocardial infarction incidence and mortality rates in the province. Physical activity is a protective factor against coronary heart disease. We investigated whether the genetic variants Q and R of the paraoxonase Gln-Arg 192 polymorphism were involved in different responses of lipids to physical activity. Serum triglycerides, HDL-cholesterol concentrations, and the paraoxonase Gln-Arg 192 polymorphism were determined in 262 men randomly selected from a representative population sample in a cross-sectional study conducted in Gerona, Spain. The Minnesota Leisure Time Physical Activity Questionnaire was used to assess energy expenditure in leisure time physical activity. No differences were found in lipid levels among tertiles of physical activity distribution in subjects with the QQ genotype. However, R carriers showed a significant decreasing trend in triglyceride levels and in log-triglyceride-to-HDL-cholesterol ratio and a significant increasing trend in HDL-cholesterol concentration with the amount of physical activity. R carriers included in the low tertile of physical activity distribution had HDL-cholesterol levels significantly lower than those of QQ homozygous men in the same physical activity category (1.04 mmol/L vs. 1.22 mmol/L, P = 0.024). R carriers of the higher tertile of physical activity distribution showed the most favorable lipid profile in this genetic group. A statistically-significant interaction between paraoxonase genotypes and physical activity was observed for log triglycerides (P = 0.018), HDL-cholesterol concentration (P = 0.017), and log triglyceride-to-HDL-cholesterol ratio (P = 0.008). The beneficial association of the amount of physical activity and lipid traits found in men with the R allele suggests that this population subgroup needs to be physically active to achieve a favorable lipoprotein phenotype similar to that observed in QQ homozygous men. PMID- 10723109 TI - Linkage analysis of 150 high-risk prostate cancer families at 1q24-25. AB - Confirmation of linkage and estimation of the proportion of families who are linked in large independent datasets is essential to understanding the significance of cancer susceptibility genes. We report here on an analysis of 150 high-risk prostate cancer families (2,176 individuals) for potential linkage to the HPC1 prostate cancer susceptibility locus at 1q24-25. This dataset includes 640 affected men with an average age at prostate cancer diagnosis of 66. 8 years (range, 39-94), representing the largest collection of high-risk families analyzed for linkage in this region to date. Linkage to multiple 1q24-25 markers was strongly rejected for the sample as a whole (lod scores at theta = 0 ranged from -30.83 to -18. 42). Assuming heterogeneity, the estimated proportion of families linked (alpha) at HPC1 in the entire dataset was 2.6%, using multipoint analysis. Because locus heterogeneity may lead to false rejection of linkage, data were stratified based on homogeneous subsets. When restricted to 21 Caucasian families with five or more affected family members and mean age at diagnosis < = 65 years, the lod scores at theta = 0 remained less than -4.0. These results indicate that the overall portion of hereditary prostate cancer families whose disease is due to inherited variation in HPC1 may be less than originally estimated. PMID- 10723111 TI - When will the ACE-inhibitors be declared obsolete? PMID- 10723112 TI - Fifty years of Framingham Study contributions to understanding hypertension. AB - The Framingham Study established hypertension as a major cardiovascular risk factor and quantified its atherogenic cardiovascular disease potential. An historical perspective is presented on the epidemiological insights about hypertension derived from 50 years of Framingham Study research into the prevalence, incidence, determinants and hazards of hypertension. Existing misconceptions about the presence of critical levels of blood pressure, the impact of the systolic and diastolic components of blood pressure, the hazard 'mild' hypertension, the impact in advanced age and the hazard of left ventricular hypertrophy. The importance of isolated systolic hypertension and the pulse pressure were demonstrated. It has been demonstrated that hypertension seldom occurs in isolation of other atherogenic risk factors, with which it tends to cluster. This clustering with other metabolically linked risk factors has been shown to reflect insulin resistance promoted by weight gain and abdominal obesity. Obesity was shown to be one of the major determinants of hypertension in the general population. Left ventricular hypertrophy was shown to be an ominous harbinger of cardiovascular disease rather than an incidental compensatory phenomenon. Multivariate risk profiles for coronary disease, stroke, peripheral artery disease and heart failure have been devised to facilitate incorporation of elevated blood pressure in a global, multivariate cardiovascular risk assessment. PMID- 10723113 TI - Alpha-1-antitrypsin phenotypes in patients with renal arterial fibromuscular dysplasia. AB - Fibromuscular dysplasia (FMD) is a significant cause of renal artery stenosis, especially in young females. A rare association between FMD and alpha 1 antitrypsin (alpha 1-AT) deficiency has been reported. We compared the alpha 1-AT phenotype distribution in 83 patients with renal arterial FMD with those published for Australian populations. alpha 1-AT phenotyping was performed by isoelectric focusing between pH 4.2 and pH 4.9 on polyacrylamide gels with PiM1M2, PiFM (non-deficiency alleles), PiMS and PiMZ (deficiency alleles) markers. Following phenotyping, alpha 1-AT genotyping was performed in 10 patients to confirm the presence of S and/or Z alleles. The phenotype distribution and allele frequencies were similar to those reported for normal subjects from two Australian populations (72 (86.7%) PiMM phenotype, one (1.2%) PiFM, seven (8.4%) PiMS, two (2.4%) PiMZ and one (1.2%) PiSZ), suggesting that alpha 1-AT deficiency is not a common aetiological factor in renal arterial FMD. However, despite FMD being three times less common in males than females, and carotid artery dissection being a rare occurrence, a male with PiMS deficiency phenotype presented with internal carotid artery dissection and had bilateral renal artery FMD. Further, a patient with PiSZ deficiency phenotype was one of two sisters with FMD and was more severely affected than her PiMM normal phenotype sibling. These two patients from the present series together with nine culled from the literature with alpha 1-AT deficiency phenotype and FMD suggest that the chance combination of alpha 1-AT deficiency and FMD may predispose to severe manifestations of FMD. PMID- 10723114 TI - Association of the alpha-adducin polymorphism with blood pressure and risk of myocardial infarction. AB - Genetic variation in adducin, a protein associated with the inner leaflet of the plasma membrane, may be in part responsible for salt-sensitive hypertension. In the Netherlands, 560 men who survived a myocardial infarction and 646 men who had undergone an orthopaedic intervention participated in a case-control study. In men in this study, the alpha-adducin polymorphism was not associated with the risk of myocardial infarction either among those with or among those without a clinical history of hypertension. In a cross-sectional analysis of blood pressure data from the controls, the alpha-adducin polymorphism was associated neither with self-reported hypertension (OR = 0.78, 95% CI = 0.51-1.19) nor with mean levels of systolic or diastolic blood pressure. Additional studies in other populations are needed to assess the contribution of alpha-adducin to high blood pressure and cardiovascular risk. PMID- 10723115 TI - A randomised placebo controlled trial of the effects of tibolone on blood pressure and lipids in hypertensive women. AB - The effects of hormone replacement therapy in hypertensive women are controversial. This randomised placebo controlled trial assessed the effect of tibolone 2.5 mg on blood pressure and fasting plasma lipids in 29 hypertensive postmenopausal women over 6 months using a 2:1 randomisation to tibolone. The primary clinical end-point was mean office blood pressure. At 6 months systolic blood pressure declined by 5.30 +/- 2.87% vs 4.94 +/- 3.37% whilst diastolic blood pressure declined 5.38 +/- 2.65% vs 0.85 +/- 3.69% on tibolone and placebo respectively. These differences were not statistically significant. Triglycerides decreased by 33.3 +/- 6.1% vs 7.6 +/- 7.9% (P < 0.01) and high-density lipoprotein (HDL)-cholesterol by 21.7 +/- 3.8% vs 2.4 +/- 2.6% (P < 0.01) with tibolone as opposed to placebo. No significant differences were observed in total cholesterol, low-density lipoprotein (LDL)-cholesterol and lipoprotein (a). Fibrinogen levels were reduced by 13.6 +/- 6.8% on tibolone compared to a 19.3 +/ 15.4% rise (P < 0.05) on placebo. This study suggests that tibolone has no deleterious effect on blood pressure in women with hypertension but has contrasting effects on biochemical risk factors. Large-scale studies are required to determine the overall effect of tibolone on cardiovascular morbidity and mortality. PMID- 10723116 TI - Thromboviscometry as a tool for evaluation of thrombotic risk in systemic hypertension. AB - In the present study, thromboviscometry was used to analyse the dynamic coagulation of blood in patients with severe systemic hypertension. Fibrinogen levels and whole blood viscosity, corrected for 45% haematocrit, were also monitored. The efficacy of thromboviscometry as an adjunct diagnostic tool, for determination of thrombogenic potential, was compared with that of detection of fibrinogen levels in the blood. Twenty-five cases of severe systemic hypertension (HT) in the 40 to 50-year age group were compared with 50 age and sex-matched normal controls (NC). The changes in whole blood viscosity were monitored with time at a constant shear rate, in a concentric cylinder viscometer, during the clotting process. The total thrombus formation time was significantly less in the HT group when compared with NC (238.9 +/- 38.72 s vs 315.1 +/- 32.93 s, P < 0.0005). The time required for a sudden increase in viscosity during clotting was also significantly lower in the HT group (205.9 +/- 34.37 s vs 272.9 +/- 28.83 s, P < 0.0005) and the overall rate of increase of thrombus viscosity was significantly higher in HT (245.2 +/- 36.44 centiPoise/s vs 183.6 +/- 16.32 centiPoise/s, P < 0.0005). There was, however, no significant change in the fibrinogen levels of the two groups. Thus, thromboviscometry was a more sensitive indicator of the thrombogenic potential of blood in HT than fibrinogen levels. The increased thrombogenic potential of hypertensive blood could be due to acceleration of the initial part of the coagulation process during the activation of factor Xa and the formation of thrombin. PMID- 10723117 TI - Blood pressure control in subjects with type 2 diabetes. AB - Blood pressure (BP) control of type 2 diabetic subjects aged under 65 years was assessed in a primary care setting. In addition, the usefulness of 24-h ambulatory BP measurement (ABPM) in the treatment of hypertension was assessed in subjects with diastolic BP (DBP) > or = 90 mm Hg. Of the total 381 diabetic subjects, 260 (68%) participated in the first phase, and 48 of the 110 subjects with DBP > or = 90 mm Hg were equipped with a Meditech ABPM-02 monitor in the second phase. The mean BP of the 260 participants was 156/91 (s.d. 22/11) mm Hg. According to the WHO criteria, 58% had hypertension, and 42% had a diagnosis of hypertension. Albuminuria > or = 20 micrograms/min was detected in 32% of the subjects. Ten percent of the subjects with diagnosed hypertension had a mean BP < 140/90 mm Hg and 50% had a mean BP > or = 160/95 mm Hg, as many as 38% of those not having a diagnosis of hypertension. Only long-term poor BP control in casual measurements was associated with albuminuria (42% vs 27%, P = 0.018). It is concluded that BP control was unsatisfactory and diagnosis of hypertension was delayed in most subjects with type 2 diabetes. Occurrence of microalbuminuria was associated with poor BP control and urinary albumin excretion rate may be useful in assessing the BP control. Further studies are needed to assess the position of 24-h ABPM in the treatment of hypertension of subjects with type 2 diabetes. PMID- 10723118 TI - Determinants of adolescent blood pressure: findings from the Glasgow University student cohort. AB - OBJECTIVE: To establish whether the blood pressure (BP) of young adults is related to their physical or behavioural attributes. DESIGN: Cross-sectional study. SUBJECTS: A total of 11,284 males and 3491 females who attended the University of Glasgow between 1948 and 1968. OUTCOME MEASURES: Systolic and diastolic BP. RESULTS: Body mass index (BMI) and weight were positively associated with BP in males and females. Height had a small positive association with male systolic BP and female diastolic BP. Haemoglobin levels were associated with raised BP in females. Albuminuria was associated with raised BP in males but not females. Smoking and alcohol consumption were associated with small negative effects on BP, although the measurement of alcohol consumption in the study was crude, and these results may not be free of confounding. Insufficient outdoor exercise was associated with higher BP in females but not males. No social class gradients in BP levels were seen, although having a higher number of siblings was associated with slightly lower BP in males. CONCLUSION: The results from the current study indicate that modifiable risk factors, such as weight and exercise affect the BP of young adults. The impact of these factors, coupled with the known tracking of BP from adolescence to adulthood, emphasise the importance of healthy behaviour patterns in young people. PMID- 10723119 TI - The effects of exercise duration on post-exercise hypotension. AB - STUDY 1: Thirteen normotensive participants with average baseline blood pressure of 126/71 mm Hg participated in the study. Participants performed bouts of cycle ergometry for 15, 30 and 45 min at 70% VO2 Peak. Blood pressure was monitored by the Finapres method with 2 min windows recorded at rest, 5, 10, 15, 30, 45 and 60 min post-exercise. Following exercise, systolic blood pressure (SBP) was similar between the three trials and was reduced from pre-exercise values at 5 through 60 min of measurement. Diastolic blood pressure (DBP) was also unaffected by the duration of exercise and was lower than before exercise at 30 through 45 min post exercise. STUDY 2: Eight borderline hypertensive participants with average baseline blood pressure of 133/79 mm Hg participated in the study. Subjects performed bouts of cycle ergometry for 10 and 30 min at 70% VO2 Peak. Following exercise, blood pressure was monitored as in study 1. SBP was similar between both trials and was reduced from baseline at 5 through 60 min post-exercise. The largest decrement of SBP was 14 mm Hg and occurred 15 min post-exercise. DBP was also unaffected by the duration of exercise and was lower than pre-exercise levels at 5 min and again at 15 through 45 min post-exercise. Mean arterial pressure (MAP) also showed significant decrements throughout the entire 1 h post exercise period by a maximum of 9 mm Hg at 15 min post-exercise, irrespective of exercise duration. We conclude that moderately intense exercise may be as brief as 10 min in duration in order to elicit a decrease in resting blood pressure and may have potential benefits as a non-pharmacological aid to hypertension. PMID- 10723120 TI - Angiotensin-converting enzyme (ACE) gene polymorphisms, serum ACE activity and blood pressure in a Spanish-Mediterranean population. AB - Angiotensin-converting enzyme (ACE) levels and ACE gene polymorphisms have been related with hypertension but with contradictory results between populations. We have investigated the association among the allelic distribution of the insertion deletion (I/D) polymorphism of the ACE gene, identified by polymerase chain reaction (PCR), serum ACE activity determined by spectrophotometry, and the blood pressure (BP), in a Mediterranean population in the southwest of Europe. A total of 1322 randomised individuals were analysed, and a comparative study was conducted analysing 205 individuals from the group with highest BP (fifth quintyl) and 196 from the group with lowest BP (first quintyl). In addition we have studied the frequencies of alleles in separated groups of women and men. We conclude that in this population there is no association between I/D polymorphism and hypertension. However, we have found a statistically significant association between the presence of the D allele in the genotypes and an elevation of serum ACE activity. PMID- 10723121 TI - Association between 24-hour ambulatory heart rate and arterial stiffness. AB - Clinical and experimental studies point to a positive association between carotid femoral pulse wave velocity (PWVcf) and casual heart rate. However, an association with 24-h ambulatory heart rate has never been investigated. Twenty four hour ambulatory heart rate and PWVcf (automatic computerised technique) were simultaneously measured in 213 subjects with untreated mild-to-moderate essential hypertension. It was found that mean ambulatory heart rate was higher in women than in men but the difference reached statistical significance only in those below 50 years of age during night-time measurements. As well as age and blood pressure, 24-h ambulatory heart rate was also an independent factor influencing PWVcf. Independent of gender, the relationship between PWVcf and ambulatory heart rate was stronger in patients over 50 years of age. Casual heart rate was not a significant determinant of PWVcf in this population. In conclusion, ambulatory heart rate contribution to explain pulse wave velocity is more important than casual heart rate. The relationship between 24-h heart rate and pulse wave velocity is stronger for subjects older than 50 years of age independent of gender. PMID- 10723122 TI - Blood pressure levels and hypertension status among ethnic groups in England. AB - The prevalence of cardiovascular disease and hypertension show wide variability among different ethnic groups in the UK. We combined data collected annually between 1991-1996 in the Health Surveys for England--nationwide surveys that provide information on the health status in a representative sample of the population living in England, to compare blood pressure (BP) levels, hypertension rates (systolic BP > or = 160 mm Hg or diastolic BP > or = 95 mm Hg, or those on antihypertensive medication), hypertension treatment and control rates in people of white, black (combining black-Caribbean, black-African and black-other), and South Asian origin (combining Indians, Pakistanis and Bangladeshis). Analyses were stratified into two age groups, 16-39 (younger) and > or = 40 years (older), but were focused on older adults (30,619 whites, 295 blacks and 529 South Asians). Age-adjusted mean BP levels and hypertension rates of older adults were highest among blacks, while South Asian men showed BP levels and hypertension rates similar to black men and South Asian women had mean BP levels and hypertension rates similar to white women. After controlling for age, BMI, smoking, alcohol consumption, and social class the odds ratio (OR) of being hypertensive among older adults was higher in black men (OR 2.0; 95% CI 1.4, 2.9; P < 0.001); black women (OR 1.7; 95% CI 1.2, 2.5; P < 0.01); and South Asian men (1.9; CI 1.4, 2.4; P < 0.001), than in their white counterparts. Among those studied with hypertension, treatment rates were highest among black men and women. Among those on antihypertensive medication, the odds of having BP controlled (SBP < 160 mm Hg and DBP < 95 mm Hg) did not differ among the three groups of older men but was reduced in older South Asian women, compared with white women. PMID- 10723123 TI - Page's syndrome: a case of pseudophaeochromocytoma. PMID- 10723124 TI - Coexistence of atherosclerotic renal artery stenosis with primary hyperaldosteronism. AB - The discovery of two forms of secondary hypertension in the same patient is unusual and suggests similar pathophysiological mechanisms, a predisposition to one type in the presence of the other or a chance occurrence. We describe two patients with renal artery stenosis who after successful correction of the stenotic lesions were discovered to have primary hyperaldosteronism associated with bilateral adrenal hyperplasia. Initially prior to revascularisation of the renal artery stenosis, the diagnosis of primary hyperaldosteronism was not evident. Both patients were subjected to further diagnostic evaluation after the appearance of hypokalaemia in one patient and continued resistant hypertension in both patients. The addition of spironolactone therapy reduced blood pressure impressively in both patients. Clinicians should be aware of the possibility that these two forms of secondary hypertension may be present in the same patient and that optimal blood pressure control requires diagnostic assessment and intervention for both disorders. PMID- 10723125 TI - Gingival hyperplasia caused by calcium channel blockers. PMID- 10723126 TI - Screening for the GRA mutation in Jamaica. PMID- 10723127 TI - Activation of NF-kappa B by the dsRNA-dependent protein kinase, PKR involves the I kappa B kinase complex. AB - Besides its known role as a translational controlling factor, the double stranded RNA-dependent protein kinase (PKR) is a key transcriptional regulator exerting antiviral and antitumoural activities. We have recently described that induction of NF-kappa B by PKR is involved in apoptosis commitment. To define how PKR mediates NF-kappa B activation by dsRNA, we have used two different approaches, one based on expression of PKR by a vaccinia virus (VV) recombinant and the other based on induction of endogenous PKR by poly I:C (pIC) treatment. We found that NF-kappa B complexes induced by PKR are composed primarily of p50-p65 heterodimers and also of c-rel-p50 heterodimers. As described for other stimuli, following pIC treatment, PKR phosphorylates the NF-kappa B inhibitor I kappa B alpha at serine 32 before degradation. Expression by VV recombinants of IKK1 or IKK2 dominant negative mutants together with PKR showed inhibition of PKR-induced NF-kappa B activation, as measured both by gel shift and luciferase reporter assays. Immunoprecipitation analysis revealed that PKR interacts with the IKK complex. Our findings demonstrate that physiological function(s) of PKR involve activation of the I kappa B kinase complex. Oncogene (2000) 19,1369 - 1378. PMID- 10723129 TI - Ionizing radiation activates the ATM kinase throughout the cell cycle. AB - The ATM protein kinase is a critical intermediate in a number of cellular responses to ionizing irradiation (IR) and possibly other stresses. ATM dysfunction results in abnormal checkpoint responses in multiple phases of the cell cycle, including G1, S and G2. Though downstream targets of the ATM kinase are still being elucidated, it has been demonstrated that ATM acts upstream of p53 in a signal transduction pathway initiated by IR and can phosphorylate p53 at serine 15. The cell cycle stage-specificity of ATM activation and p53Ser15 phosphorylation was investigated in normal lymphoblastoid cell line (GM536). Ionizing radiation was found to enhance the kinase activity of ATM in all phases of the cell cycle. This enhanced activity was apparent immediately after treatment of cells with IR, but was not accompanied by a change in the abundance of the ATM protein. Since IR activates the ATM kinase in all phases of the cell cycle, DNA replication-dependent strand breaks are not required for this activation. Further, since p53 protein is not directly required for IR-induced S and G2-phase checkpoints, the ATM kinase likely has different functional targets in different phases of the cell cycle. These observations indicate that the ATM kinase is necessary primarily for the immediate response to DNA damage incurred in all phases of the cell cycle. PMID- 10723128 TI - Activation of the c-fos enhancer by the erk MAP kinase pathway through two sequence elements: the c-fos AP-1 and p62TCF sites. AB - The c-fos enhancer can be activated by many signaling pathways through distinct elements of the enhancer. The enhancer contains at its core the serum response element (SRE) that binds serum response factor (SRF). On the 5' side of the SRE is a site for p62TCF which binds only when SRF is bound as well. p62TCF is encoded by three ets-related genes, Elk-1, SAP1 and SAP2. Each of these factors contain a transcriptional activation domain that is activated by phosphorylation by MAP kinases. On the 3' side of the SRE is the 'c-fos AP1 site' (FAP1) whose role has been less clear. We find here that the FAP1 site contributes strongly to phorbol ester (TPA) and Erk MAP kinase activation of the c-fos enhancer and that both the p62TCF and FAP1 sites are required for effective activation of the enhancer. We further find that the FAP1 site binds ATF1 and CREB from HeLa nuclear extracts and that the phosphorylation of these factors is induced by TPA. ATF1 and CREB can be phosphorylated by Rsk2 which is a protein kinase directly activated by Erk MAP kinases. These results suggest a signaling pathway in which Erk MAP kinase activates the c-fos enhancer by direct phosphorylation of p62TCF and by activation of Rsk related kinases that phosphorylate ATF1 and CREB. PMID- 10723130 TI - Combined LOH/CGH analysis proves the existence of interstitial 3p deletions in renal cell carcinoma. AB - We have recently developed an allele titration assay (ATA) to assess the sensitivity and influence of normal cell admixture in loss of heterozygosity (LOH) studies based on CA-repeat. The assay showed that these studies are biased by the size-dependent differential sensitivity of allele detection. Based on these data, we have set up new criteria for evaluation of LOH. By combining these new rules with comparative genome hybridization (CGH) we have shown the presence of interstitial deletions in renal cell carcinoma (RCC) biopsies and cell lines. At least three out of 11 analysed RCC cell lines and three out of 37 biopsies contain interstitial deletions on chromosome 3. Our study suggests the presence of several regions on human chromosome 3 that might contribute to tumor development by their loss: (i) 3p25-p26, around the VHL gene (D3S1317); (ii) 3p21. 3-p22 (between D3S1260 and D3S1611); (iii) 3p21.2 (around D3S1235 and D3S1289); (iv) 3p13-p14 (around D3S1312 and D3S1285). For the first time, AP20 region (3p21.3-p22) was carefully tested for LOH in RCC. It was found that the AP20 region is the most frequently affected area. Our data also suggest that another tumor suppressor gene is located near the VHL gene in 3p25-p26. PMID- 10723131 TI - The amino-terminus and membrane-spanning domains of LMP-1 inhibit cell proliferation. AB - The LMP-1 oncoprotein of EBV is required to maintain proliferation of infected B cells and shares several features with CD40, TNF-R1, and related receptors. Members of this family can bind TRAF and TRADD molecules and activate NF-kappaB and AP-1, as can LMP-1. While CD40 and TNF-R1 are dependent on binding their ligands for their signaling, LMP-1 apparently is not. We have found that LMP-1 can act as a governor of cell proliferation and thereby limit its own activities. Its inhibition of proliferation is not mediated by apoptosis but results in cytostasis in four cell lines tested. The structural moiety of LMP-1 that distinguishes it from CD40 and TNF-R1, its amino-terminus and multiple membrane spanning segments, alone can mediate its cytostatic activity. PMID- 10723132 TI - An IFN regulatory factor-2 DNA-binding domain dominant negative mutant exhibits altered cell growth and gene expression. AB - In order to study interferon regulatory factor (IRF) family mediation of cell growth regulation, we established U937 cell lines stably transfected with a truncated form of IRF-2 lacking the transcriptional repressor domain. The truncated IRF-2 contained the DNA binding domain (DBD) and bound the ISRE. Phenotypically, the IRF-2 DBD transfectants exhibited reduced cell growth, altered morphology and increased cell death. Consistent with alterations in growth characteristics, the IRF-2 DBD transfectants constitutively expressed higher levels of the cyclin dependent kinase inhibitor p21WAF1/Cip1 than did control clones. The level of p21WAF1/Cip1 expression was positively correlated with the level of DBD expressed, as well as with the level of growth inhibition in these clones. DBD expression also correlated with expression of other members of the growth regulatory complex, cyclin dependent kinase 2 and cyclin A, but not proliferating cell nuclear antigen. These results imply active repression by IRF 2 to keep p21WAF1/Cip1 transcriptionally silent. PMID- 10723133 TI - The p44S10 locus, encoding a subunit of the proteasome regulatory particle, is amplified during progression of cutaneous malignant melanoma. AB - Gene amplification is frequently present in human tumors, although specific target genes relevant to many amplified loci remain unidentified. An expression cloning assay enabled identification of a candidate oncogene derived from human chromosome 3p14.1. The cDNA retrieved from morphologically transformed cells contained the full-length protein coding region and detected an abundant transcript in the same cells. Sequence analysis revealed identity with the wild type sequence of p44S10, a highly conserved subunit of the 26S proteasome that exhibits similarity to the Arabidopsis fus6/cop11 family of signaling molecules. p44S10 gene copy number and mRNA expression were increased in association with segmental 1.8 - 11-fold chromosomal gains in cutaneous malignant melanoma cell lines (5/13; 40%) and tumors (2/40; 5%), and in breast cancer MCF-7 cells. Likewise, malignant progression of human radial growth phase WM35 melanoma cells was associated with amplification and increased expression of endogenous p44S10, and increased expression of p44S10 was sufficient to induce proliferation of WM35 cells in vivo. The results demonstrate segmental copy number gains within chromosome 3p in cutaneous malignant melanoma and suggest that deregulation of a proteasome regulatory particle subunit may contribute to the malignant phenotype. PMID- 10723134 TI - Distinct involvement of cdc42 and RhoA GTPases in actin organization and cell shape in untransformed and Dbl oncogene transformed NIH3T3 cells. AB - The Dbl oncogene is a putative exchange factor for the small GTPases RhoA and Cdc42, which are involved in actin polymerization into stress fibers and filopodia, respectively. We report here that, upon adhesion to fibronectin, Dbl transformed NIH3T3 cells display a contracted, polygonal shape with a high number of short stress fibers. In contrast, untransformed NIH3T3 cells acquire the characteristic fibroblast morphology and organize a regular mesh of long stress fibers. We show that in Dbl-transformed and in untransformed NIH3T3 cells the different shape and actin cytoskeleton organization observed in the early steps of adhesion involves activation of distinct GTPases. Upon adhesion to fibronectin, cell morphology of Dbl-transformed NIH3T3 cells depends on activation of RhoA and not of Cdc42. In contrast Cdc42 activation is necessary to untransfected NIH3T3 cells to acquire their fibroblast shape. In both Dbl transformed and in untransformed NIH3T3 cells a basal Rac activation is necessary to support stress fiber organization, while constitutive Rac activation promotes ruffles and lamellipodia formation. As a consequence of RhoA activation, Dbl transformed cells show high activity of ROCK-alpha and CRIK kinases, two known RhoA effectors. In addition Dbl-transformed and NIH3T3 cells expressing the constitutive active form of RhoA are less motile on fibronectin than cells expressing constitutive active Cdc42. We conclude that in NIH3T3 cells in response to fibronectin the expression of the Dbl oncogene leads to a predominant activation of RhoA which both supports the peculiar cell shape and actin cytoskeleton organization in stress fibers and regulates cell motility. PMID- 10723135 TI - Inhibition of Myc-dependent apoptosis by eukaryotic translation initiation factor 4E requires cyclin D1. AB - Ectopically expressed eukaryotic translation initiation factor 4E (eIF4E) stimulates cell proliferation, suppresses apoptosis in growth factor restricted cells, and induces malignant transformation in primary rodent fibroblasts when coexpressed with protooncogene myc. We report here that eIF4E rescued rat embryo fibroblasts ectopically expressing c-Myc (REF/Myc) from genotoxic and non genotoxic cytostatic drugs and identify cyclin D1 as a downstream effector in the antiapoptotic mechanism. In clones of REF/Myc ectopically expressing eIF4E, resistance to apoptosis paralleled steady state levels of cyclin D1. Stable expression of cyclin D1 in REF/Myc inhibited apoptosis in response to a broad range of cell cycle specific cytostatic agents. Partial loss-of-cyclin D1 function in REF/Myc ectopically expressing eIF4E (REF/Myc/4E) significantly increased chemosensitivity; either soluble antisense cyclin D1 oligomers or transfection with a dominant negative cyclin D1 mutant that prevents translocation of cyclin D-dependent kinases to the nucleus, significantly blunted the antiapoptotic effect of eIF4E. These data directly link eIF4E rescue from cytostatic drugs to cyclin D1. Since overexpression of eIF4E and cyclin D1 is observed in many aggressive forms of chemoresistant cancers, these findings provide insight into possible mechanisms responsible for this biological behavior. PMID- 10723137 TI - Modulation of retinoic acid receptor function alters the growth inhibitory response of oral SCC cells to retinoids. AB - Retinoids have been shown to inhibit the growth of many human tumor cells including breast, ovarian and squamous cell carcinoma (SCC). While the exact mechanism of retinoid mediated growth suppression is not known, a role for the retinoic acid receptors (RARs) and retinoid X receptors (RXRs) has been established in both the breast and ovarian tumor cell models. We set out to determine if modulation of RAR/RXR function would alter the retinoid sensitivity of oral SCC cells. We found that the growth of SCC cells was significantly inhibited by treatment with either all-trans-retinoic acid (trans-RA) or the synthetic, conformationally restricted RARgamma selective retinoids MM11254 and MM11389. In order to demonstrate a role for RAR/RXR function in this process, stable oral SCC cell clones constitutively overexpressing the dominant negative mutant RARbeta2 (R269Q) were prepared and shown to exhibit reduced RAR/RXR transcriptional transactivation activity. We found that oral SCC cells exhibiting reduced RAR/RXR function became resistant to growth inhibition by all-trans-RA, MM11254 and MM11389. Likewise, treatment of oral SCC cells with the RARgamma antagonist MM11253 was found to block the ability of MM11254 and MM11389 to inhibit SCC cell growth. Thus, modulation of RAR function through the use of RAR gamma selective agonists, an RAR-gamma selective antagonist or a pan-RAR dominant negative mutant significantly alters the growth inhibitory response of oral SCC cells to retinoids. PMID- 10723136 TI - Somatic mutagenesis studies of NF-kappa B signaling in human T cells: evidence for an essential role of IKK gamma in NF-kappa B activation by T-cell costimulatory signals and HTLV-I Tax protein. AB - NF-kappa B plays a pivotal role in normal T-cell activation and may also mediate human T-cell leukemia virus (HTLV)-induced T-cell transformation. Activation of NF-kappa B by both T-cell costimulatory signals and the HTLV Tax protein involves stimulation of I kappa B kinase (IKK). As a genetic approach to dissect the intermediate steps involved in NF-kappa B activation in human T cells, we performed somatic cell mutagenesis to isolate signaling-defective mutant Jurkat T cell lines. One of the mutant cell lines was shown to have a specific blockade in the IKK signaling pathway but remained competent in the c-Jun N-terminal kinase and MAP kinase pathways. Interestingly, this mutant cell line lacks expression of IKK gamma, a non-catalytic component of the IKK complex. Expression of exogenous IKK gamma in the mutant cells restored NF-kappa B activation by both the T-cell costimulation agents and Tax. These findings provide genetic evidence for the requirement of IKK gamma in NF-kappa B signaling triggered by both T-cell costimulatory signals and HTLV-I Tax protein. PMID- 10723138 TI - Allelotyping defines minimal imbalance at chromosomal region 17q25 in non-serous epithelial ovarian cancers. AB - Allelic deletions of multiple chromosome 17q loci in sporadic ovarian cancer of epithelial origin suggest that inactivation of tumor suppressor gene(s) in these regions may be important for ovarian tumorigenesis. To further define the pattern of allelic imbalance in epithelial ovarian tumors of different histologies, a PCR based assay was used to assess loss of heterozygosity (LOH) of polymorphic markers representative of TP53, BRCA1, NME1 and GH1, and region 17q23-25. LOH was observed for at least one marker in 68% of malignant tumors (n=60) and in 18% tumors of borderline malignancy (n=11), but not in benign tumors (n=5). The highest frequency of LOH in malignant tumors (64%) was observed with D17S801 on 17q25. Ten of 39 malignant ovarian tumors displaying LOH of at least one 17q marker, displayed a LOH pattern enabling the determination of a minimal region of overlapping deletion defined by D17S795 and D17S801. One borderline tumor also displayed an interstitial LOH pattern that overlapped this 17q25 minimal region of deletion. The histologies of malignant tumors displaying a pattern indicative of interstitial 17q deletions were of the endometrioid, clear cell and mucinous epithelial types. As the minimal region of overlap defined by these tumors overlap regions deleted in malignant tumors of all histologic types, and in a tumor of borderline malignancy, the 17q25-tumor suppressor may be implicated in the development of all types of epithelial ovarian tumors. PMID- 10723139 TI - Activity of MDM2, a ubiquitin ligase, toward p53 or itself is dependent on the RING finger domain of the ligase. AB - We previously showed that oncoprotein MDM2 has ubiquitin ligase activity toward tumor suppressor p53. In that paper, we showed very weak homology in the carboxyl terminal portion between MDM2 and E6AP (HECT domain). We mutated the cysteine residue (C464) corresponding to the residue essential for the ubiquitin ligase activity of E6AP and this mutation diminished the ligase activity of MDM2. The cysteine residue described above is also one of the cysteine residues that form the RING finger domain of MDM2. We tried to find out whether the diminishing of the activity by the mutation is attributable to the disruption of the RING finger domain or not. When the ring finger domain of MDM2 was deleted, the truncation mutant did not have the ubiquitin ligase activity. When we mutated the seven cysteine residues of RING finger domain of MDM2 in the carboxyl terminus, the disruption of each residue in the RING finger completely diminished the ubiquitin ligase activity of MDM2 toward MDM2 itself and toward tumor suppressor p53. These data indicate that the RING finger domain in MDM2 is essential for its ubiquitin ligase activity toward p53 and itself. PMID- 10723140 TI - p53-independent association between SV40 large T antigen and the major cytosolic heat shock protein, HSP90. AB - The simian double strand DNA tumor virus SV40 encodes the 90-kDa multi-functional protein, large T antigen (LT). LT functions by binding to DNA, as well as to many cellular target proteins such as p53 and retinoblastoma protein (pRB). We report here the identification of a cellular heat shock protein, HSP90, as a previously undescribed LT-associated protein. Immunoprecipitates by anti-HSP90 antibodies from LT-expressing cell lysates contained LT protein, as revealed by Western blotting. Conversely, anti-LT antibody co-immunoprecipitated HSP90. Co immunoprecipitation of HSP90 and LT was observed even after complete immuno depletion of p53, indicating that the association of LT with HSP90 is p53 independent. LT-HSP90 complexes can be reconstituted from purified HSP90 and unfolded-LT in vitro in an ATP-independent manner but not from HSP90 and native LT, suggesting that non-mature conformation of LT is required for the efficient association with HSP90. Moreover, geldanamycin, an anti-tumor drug that specifically binds and inhibits HSP90, reduced the intracellular concentration of LT by destabilizing newly synthesized LT. The above results suggest that HSP90 associates with immature forms of LT both in vivo and in vitro, and thus might assist LT in the formation of a functional, mature structure. PMID- 10723141 TI - Identification of Mad as a repressor of the human telomerase (hTERT) gene. AB - Activation of telomerase, which has been frequently associated with cellular immortality, may constitute a key step in the development of human cancer. De repression in the expression of its catalytic subunit hTERT gene has been proposed to directly link to the telomerase activation in tumor cells. Little is known about the mechanism how the hTERT gene is repressed in telomerase-negative mortal cells. This study was conducted, using an expression cloning approach, with the aim of identifying the gene(s) responsible for repressing the hTERT gene expression. Using this genetic screen, we isolated the transcription factor Mad as a repressor. Mutation of its DNA binding sites caused significant de repression of hTERT promoter activity in mortal cells. This Mad-mediated repression of the hTERT promoter in mortal cells was counteracted by ectopic expression of Myc. The antagonism between Mad and Myc was also observed with an endogenous hTERT promoter. Their potential roles in differential hTERT promoter activities were further supported by the relative amounts of Mad and Myc proteins detected in immortal and mortal cells. Thus, Mad may be a direct negative regulator of hTERT in mortal cells and this repression mechanism can be inhibited by induction of Myc in immortal cells. PMID- 10723142 TI - [Cardiac arrhythmia and conduction disorders in older patients]. PMID- 10723143 TI - [Assessment of patient's quality of life in medicine]. PMID- 10723144 TI - [Early ventricular repolarization as a probable consequence of acute viral and idiopathic myopericarditis]. AB - 64 patients who had acute viral or idiopathic myopericarditis (40 males and 24 females aged 16-48 years) were followed up for 3 to 24 years. Echocardiography has found thickening of the pericardium from 5 to 8 mm which corresponded to fibrosis in 55 patients (85.9%). Early repolarization was detected in 17 patients, in 14 of them it was associated with pericardial thickening (82.4%). Of 30 healthy controls, early repolarization in combination with pericardial fibrosis occurred in 6.7% of cases. In 7 patients with early repolarization ECG registered residual changes of the acute period, in 6 new changes arose--His block and left ventricular hypertrophy. It is suggested that in many cases early repolarization is not independent, it is sequelae of previous myopericarditis. PMID- 10723146 TI - [Variants of hemodynamic response in transesophageal pacing in patients with chronic circulatory insufficiency]. AB - Compared were hemodynamic responses to transesophageal pacing (TEP) in 86 patients with symptoms of chronic circulatory insufficiency (CCI) and 46 healthy individuals. Cardiac output was assessed by tetrapolar chest rheogram under stimulation and its discontinuation, and end diastolic pressure (EDP) in the left ventricle. Stroke volume was diminished in all the examinees. Cardiac output changed in different directions. Poststimulation recovery of cardiac output was different. Contribution of ejection period is discussed the prolongation of which indirectly indicates shorter time of diastole and, respectively, diastolic filling of cardiac chambers. EDP rose both in healthy controls and patients, being higher in the patients. The role of systolic and diastolic compensatory mechanisms in CCI development is considered. PMID- 10723145 TI - [Clinical and prognostic significance of disturbed global and regional contractility of left ventricle in diphtheria myocarditis]. AB - Examination of 159 diphtheria patients diagnosed myocarditis in 64 of them. The latter were divided into 3 groups: with mild, moderate and severe myocarditis. The patients with and without diphtheria have undergone two-dimensional echocardiography with estimation of the asinergia index (AI) and left ventricular (LV) ejection fraction. Pronounced systolic dysfunction was revealed only in severe diphtheric myocarditis (DM). AI rose in moderate and severe DM. By the degree of AI elevation it can be judged about myocarditis severity and the disease prognosis. Regional LV contractility and LV systolic dysfunction were correlated. A group of patients with severe DM was identified who had a high risk of death in akinesia of LV segments (IA > 2) and lowering of LV ejection fraction below 35%. PMID- 10723147 TI - [Disbiotic conditions in patients operated and reoperated for heart defects and ischemic heart disease]. AB - Bacteriological examination of angiocardiac system, biocenosis of the intestine and upper respiratory tracts was made in 3473 patients who were to be operated or reoperated for congenital or acquired valvular defects in the presence of chroniosepsis and chronic septic endocarditis, complicated IHD. 375 patients with other diseases served control. 74 of them have undergone surgery for varicose veins of the legs. Preoperative contamination with opportunistic microflora was found in the heart, major vessels, veins of the majority of the patients. There was also dysbiosis of natural biotops. Patients with valvular defects to be reoperated 2-20 years after the initial operation on the heart had infection in the angiocardiac structures, severe dysbiosis of natural biotops, i.e. advanced dysbiosis--severe persistent condition with permanent source of endogenic infection complicated the underlying disease and bringing postoperative septic complications. PMID- 10723148 TI - [Therapeutic complex in therapy of patients with complicated course of pylorobulbar ulcers]. AB - Combined treatment of pylorobulbar ulcers associated with duodenogastric reflux incorporated surgery, antiinflammatory and antisecretory therapy. The treatment provides preventing ulcer recurrence, gastric mucosa affection caused by duodenogastric reflux and infection Helicobacter pylori. Finally, this led to improved quality of life. PMID- 10723149 TI - [Clinical significance of cellular and humoral immunity in tropical malaria]. AB - Humoral and cellular immunity were studied in 34, 59 and 21 patients with mild, moderate and severe tropical malaria in the course of the disease. It was found that immunological indices in tropical malaria change with the disease severity. Some immunological indices may be indicative of the infection severity, outcome and prognosis of the disease and contribute to more effective chemotherapy and immunocorrection. PMID- 10723150 TI - [Losartan in therapy of chronic heart failure]. AB - To evaluate effectiveness of nonpeptide angiotensin-2 subtype-1 receptor antagonist losartan in therapy of symptomatic congestive heart failure in patients with ischemic heart disease, 116 patients were examined at the age of 36 62 (mean age 50.6 +/- 4.22). They had angina pectoris of functional class II-III (according to CCS) and congestive heart failure of functional class II-III (according to NYHA). All the patients were randomized into two groups. 60 patients of group 1 received basic medication with nitrates, diuretic (on demand), digoxin and aspirin. 56 patients of group 2 received basic medication and losartan (cozaar, MSD, USA) in the dose 25 mg/day for 48 weeks. Echocardiographic monitoring of the treatment efficacy was made. The outcomes of the treatment evidence that losartan improves the patients' clinical status and heart failure functional class. For twelve weeks losartan reduced left ventricular and atrial dilation positively influencing the isometric inotropic indices. In 48 weeks losartan arrested progression of pathologic remodelling of the left ventricle and prevents depression of total myocardial contractility. Losartan's positive effect in restriction of negative evolution of cardiac failure manifests on the treatment week 3. PMID- 10723151 TI - [Enalapril treatment of residual pulmonary hypertension in patients operated rheumatic mitral valve defects]. AB - A pilot trial of efficiency of enalapril maleate in the treatment of residual pulmonary hypertension was made in 22 patients operated for rheumatic mitral valve defects. Degree I, II and III of pulmonary hypertension was registered in 5, 13 and 4 patients, respectively. Thus, the patients had NYHA functional classes III and IV (22.7 and 77.3%, respectively. Enalapril given for 6 months in a mean daily dose 12.3 +/- 1.57 mg/m2 (5-30 mg a day) normalized pressure in the pulmonary artery in 18.2% of patients. 50% of patients showed hypertension degree I, only one female retained hypertension degree III. To the end of the treatment the functional classes were the following: II--in 68.2%, III--in 27.3% and IV--in 4.5%. PMID- 10723152 TI - [Prospects of cytomedines application in clinical medicine and gerontology]. AB - Available are the results of experimental and clinical study of a new class of peptide bioregulators--cytomedines. Mechanism of action of these substances is considered. Cytomedines-based drugs can be used for prevention and treatment of age pathology and premature aging. A new trend in clinical medicine- bioregulation therapy--is grounded. PMID- 10723153 TI - [Prescription of drugs in arterial hypertension in outpatient setting]. PMID- 10723154 TI - [Prognosis of ischemic heart disease in patients with obliterating atherosclerosis of lower limb arteries]. PMID- 10723155 TI - [Pathogenetic validation of immunotherapy in chronic gastritis and ulcer]. PMID- 10723156 TI - [Two cases of purulent pericarditis as complication of myocardial infarction]. PMID- 10723157 TI - [Variants of mitral valve prolapse clinical course]. PMID- 10723158 TI - [In Process Citation] PMID- 10723159 TI - [Medical professionals surrounding A.S. Pushkin (part II)]. PMID- 10723160 TI - The Attorney General's health care agenda. PMID- 10723161 TI - Fit to be tied. PMID- 10723162 TI - Managed care finds itself in dire straits. AB - Managed health care, in its various guises from HMOs to PPOs, has been in place in Minnesota for at least 20 years and in the rest of the country for most of the last decade. Ten years should be enough time for any idea to prove its worth. Yet, every consultant, economist, stockbroker, medical supply company executive, and pharmaceutical CEO I know believes that corporate medicine has failed to deliver its promised quality care at reduced cost. A victim not only of its own unmet promises but also of its own excesses, managed care is now facing its own demise. PMID- 10723163 TI - Faultfinding: no cure for health care woes. PMID- 10723164 TI - Late payment of physician claims. Results of an MMA survey. PMID- 10723165 TI - Rural Minnesota family physicians. Practice characteristics, gender, income, and job satisfaction. AB - This study investigated rural physicians' job satisfaction, income, and working conditions through a survey of a random sample of rural family physicians in Minnesota. Of the 300 surveys sent, 210 (70%) were returned and usable. Male physicians reported more years of experience, work hours, and call hours per week than female physicians (p < 0.0005). On a scale of 1 to 5, average overall job satisfaction among the physicians was 4.04, with men averaging 4.0 and women, 4.3. Overall job satisfaction was positively correlated with the presence of a hospital in town (p < 0.05) and negatively correlated with the number of work hours, including hours on call, per week (p < 0.05). Overall job satisfaction was lowest for those in practice 20-34 years. Average reported annual income was $127,213 (men, $131,400; women, $97,800; p < 0.0001). In a comparison of physicians working the same number of hours including call, men earned more than women. Rural Minnesota family physicians reported high job satisfaction. Income was positively correlated with size of practice, years of experience, and number of hours worked. In addition, men earned more than women, and physicians whose pay was based on production earned more than physicians on straight salary. PMID- 10723166 TI - Minnesota bound. Stability of practice location among UMD family physicians in Minnesota. AB - Previous studies indicate that physicians often move from one practice setting to another, particularly early in their career. However, data on practice relocation for a group of University of Minnesota, Duluth School of Medicine graduates show a different trend. Minnesota family physicians from the UMD School of Medicine have been remarkably stable in their practices over the past 20 years. More than 80% of these physicians have continued to practice in the same community that they selected after their training. In addition, physicians in this group who are practicing in smaller communities have not relocated to urban practices. These findings suggest that the UMD School of Medicine's emphasis on family medicine and rural practice may have influenced the practice retention rate for these physicians. PMID- 10723167 TI - Whiplash, chronic neck pain, and zygapophyseal joint disorders. A selective review. AB - Recent research on whiplash injury has challenged the long-held view of what causes chronic neck pain in car accident victims. Although it was previously thought to result from residual scarring of muscles and ligaments, such pain is now understood to be caused by zygapophyseal-joint damage, which produces no objective findings on radiologic testing and often none on physical examination. Chronic pain from a whiplash injury can be reliably diagnosed and effectively treated in most patients by z-joint testing and radiofrequency blocks. PMID- 10723168 TI - Practicing in the age of EMTALA. PMID- 10723169 TI - [Neuropathology of frontotemporal dementia]. AB - The syndrome of progressive fronto-temporal dementia represents the clinical expression of several pathological processes, mostly degenerative, preferentially involving the frontal and temporal lobes. These processes include dementia with Pick bodies, cortico-basal degeneration, dementia with ITSNU (Inclusions Tau and Synuclein Negative, Ubiquitinated), also known as dementia of motorneuron disease type, dementia with basophilic inclusion bodies, and dementia lacking specific histopathology, and in addition all variants linked to mutations in the tau gene, located in chromosome 17. The term "Pick complex" encompasses these processes and their clinical manifestations, which in addition to fronto-temporal dementia include primary progressive aphasia and apraxic-motor syndromes. The pathologic processes are better discriminated by histopathology than distribution of atrophy, but the latter is the main determinant of the clinical presentation. PMID- 10723170 TI - [Clinical manifestation of frontotemporal dementia]. AB - Frontotemporal dementia is a degenerative disease, usually of presenile onset, frequently with positive family history, whose main clinical features are early social and personal behavioral disturbances, cognitive defects mainly in attention, language and executive functions, personality disorders and blunt, unconcerned and subdepressive affect. Anatomic and functional neuroimaging shows atrophy and functional defects in anterior brain regions, and the EEG is persistently normal. Although frontotemporal dementia has been confounded with Alzheimer's disease for long time, it can be identified on clinical grounds in order to perform new therapeutic trials. PMID- 10723171 TI - [Neuropsychological exploration in frontotemporal degeneration]. AB - The present paper discusses the neuropsychological assessment in fronto-temporal lobe degeneration. Having established the neuroanatomical and functional basis for the discussion the major syndromes included in the concept of frontotemporal degeneration are reviewed from a neuropsychological standpoint. With reference to fronto-temporal dementia the different frontal or executive function tests and their limitations are discussed. With reference to progressive aphasia and semantic dementia we differentiate the distinct language profiles as observed in aphasia batteries and general neuropsychological tests. Reference is made to especially useful tests for the differentiation of the two syndromes from each other, as well as from other primary progressive disorders. Concluding remarks postulate a series of axis of cognitive function in fronto-temporal lobe degenerations, which exist at the functional as well as the anatomical level and along which the different syndromes evolve. PMID- 10723172 TI - [Neuroimaging of frontotemporal dementia]. AB - With the development of neuroimaging, frontal lobe atrophy has been demonstrated with increased frequency, first with structural studies (computed tomography and magnetic resonance imaging), then with functional images (Single photon emission computed tomography and Positron emission tomography). PMID- 10723174 TI - [Frontotemporal dementia: therapeutic possibilities]. AB - In order to treat frontotemporal dementia (FTD) we must first evaluate the patient's medical condition, as well as his or her social setting (caregiver, financial resources, home characteristics). Primary health-care team must receive information about the patient's disease, and the family should be informed about the disease itself and the social resources they can ask for. It is advisable to formulate a therapeutic scheme including some counsels to improve the suitability of environment, social help measures, behaviour therapy, cognitive stimulation and pharmacological treatment. Atypical antipsychotics have improved "positive symptoms" as logorrhoea, wandering, agitation and aggression, without impairing cognitive function. Selective serotonin reuptake inhibitors improve depressive symptoms, compulsions, food craving and disinhibition. A few reports suggest that idazoxan (alpha 2-noradrenergic antagonist) can improve attention, verbal fluency and planning efficiency. In some cases with "FTD and parkinsonism linked to chromosome 17" it could be justified to perform a genetic analysis to the offspring, in order to know if genetic counseling is necessary. An inflammatory reaction has been observed in brain damaged areas, and therefore antiinflammatory treatment efficacy should be investigated. It would also be interesting to look for neuroprotective agents that lessen the tau protein abnormality. All types of receptors which are involved in FTD should be identified, and then their selective agonists or antagonists could be administered in synergic combinations. We hope that all genetic alterations producing or facilitating FTD are eventually known, and harmless curative means are developed. PMID- 10723173 TI - [Genetics of frontotemporal dementia and alterations of the tau gene]. AB - The prevalence of fronto-temporal dementia may be estimated in 10-20% of all dementia's. In 40 to 50% of the cases it is possible to detect familial antecedents and in some of the families it is possible to identify multiple affected individuals. The study of these familial cases has allowed to narrow the responsible genomic region to chromosome 17 and to identify mutations in the tau gene as responsible for the disease. At present more than 100 cases of the disease due to mutations in the tau gene have been described in which it has been possible to detect 24 different mutations. In order to search for mutations in the tau gene in a patient all it is needed is a sample of venous blood with which it is possible to extract the genomic DNA, amplify the tau gene through PCR and to sequence it. The information obtained provides a very precise diagnosis in patients with fronto-temporal dementia, may be useful in the differential diagnosis with other types of dementia, and may allow a pre-symptomatic diagnosis in relatives from patients. At a more basic level, the detection of mutations in the tau gene allows to identify the pathogenic mechanisms involved opening up the possibility to develop new therapeutics strategies. PMID- 10723175 TI - [Characteristics of incidentally detected renal cell carcinoma by ultrasonography at health check-up]. AB - BACKGROUND: The prognosis of patients with incidentally detected renal cell carcinoma is better than that of patients with symptomatic renal cell carcinoma. These incidentalomas include those discovered by ultrasonography at health check up and those found during examinations for unrelated disease. In this study, we investigated the prognosis of the patients of the health check-up group and the unrelated disease group. METHODS: From April 1987 to March 1997, 263 patients with renal cell carcinoma were treated in our department including 166 incidentalomas (63.1%). The occasion of incidental detection was divided into 2 groups; 90 cases as health check-up group and 76 cases as unrelated disease group. RESULTS: The mean age was 52.9 +/- 9.7 years for health check-up group and 65.1 +/- 11.4 years for unrelated disease group (p < 0.01). The mean evaluated tumor size was 3.6 +/- 1.6 cm for health check-up group and 4.4 +/- 2.6 cm for unrelated disease group (p < 0.05). The survival rates were significantly different in the two groups (p < 0.01); the 5- and 10-year survival rate for health check-up group was 91.5% and 55.9%, respectively and for unrelated disease group 79.4% and 66.1%, respectively. CONCLUSION: These results suggested that examination by ultrasonography at health check-up lead to detection of smaller renal cell carcinoma and improve the prognosis further. PMID- 10723176 TI - [Immuno-histochemical identification of initial lymphatics: the contour and distribution pattern of initial lymphatics in the human foreskin of the penis]. AB - PURPOSE: There has not been an established method to distinguish initial lymphatics from blood capillaries under the light microscopy. In this study, we examined the usefulness of the immuno-histochemical staining method using a monoclonal anti-desmoplakin antibody in identifying initial lymphatics under the light microscopy. The specificity of this reaction was confirmed by the immuno electron microscopy. MATERIAL AND METHODS: The cryostat sections of the human foreskin were observed under light microscopy by indirect immunoperoxidase method with the anti-desmoplakin mouse monoclonal antibody, and compared with the hematoxylin and eosin sections. These cryostat sections were also observed under electron microscopy by pre-embedding immunoperoxidase method with the same antibody. RESULTS: Under the light microscopy, the initial lymphatics of the human foreskin were visualized by the method with anti-desmoplakin antibody. These lymphatics were mainly distributed in the dermal layer, on the other hand, rarely seen in dermal papillae. Being usually found in closed shape, the lumens of initial lymphatics were hardly recognized as initial lymphatics by the ordinary hematoxylin and eosin staining. Under the immuno-transmission electron microscopy, the peroxidase-desmoplakin antibody precipitations were located on the surface of the endothelial cells of the vasculature which lacked pericytes and basal lamina, and was composed of endothelial cells alone. By these features of the vascular structures, the vessel reacting with anti-desmoplakin antibody was identified as initial lymphatics. CONCLUSION: This study shows the reliability and specificity of the immuno-histochemical method by anti desmoplakin antibody in identifying initial lymphatics under light microscopy, and this method will be useful in studying the fine distribution of lymphatic vessels in normal human tissue. PMID- 10723177 TI - [Treatment results of VIP (etoposide, ifosfamide and cisplatin) chemotherapy as a first-line therapy in metastatic germ cell tumors]. AB - PURPOSE: We investigated the effectiveness and toxicity of VIP therapy as a first line chemotherapy for patients with metastatic germ cell tumor. PATIENTS AND METHODS: From March 1994 to October 1997, we treated 16 patients with VIP therapy consisting of etoposide (100 mg/m2), ifosfamide, (1.2 g/m2) and cisplatin (20 mg/m2), all of which were generally given daily for 5 consecutive days every 3 weeks. Of the 16 patients, 6 were classified into a good, 5 into an intermediate, and 5 into a poor prognostic group according to the International Germ Cell Consensus Classification. RESULTS: Thirteen patients (81%) achieved complete response with VIP alone or VIP plus surgery. Three-year survival rate was 100% in good and intermediate prognostic group, while 40% in poor prognostic group. Although all patients had Grade 3 or higher myelosuppression, the treatment was well tolerated and no patient died of treatment-related complications. CONCLUSIONS: VIP appears to be an effective and safe regimen as an induction chemotherapy for good and intermediate risk patients with germ cell tumor. However, more intensive regimen may be necessary for poor-risk patients. PMID- 10723178 TI - [Microsurgical penile revascularization in patients with pure arteriogenic erectile dysfunction]. AB - BACKGROUND: We report the results of microscopic penile revascularization in patients with arteriogenic erectile dysfunction. METHODS: One patient with localized obstruction of the common penile artery underwent the Michal II penile revascularization, 13 patients underwent the Furlow-Fisher procedure, and 5 patients underwent the Hauri procedure. The mean age was 33.0 years and the mean follow-up period was 32 months (4-80 months). Eight patients were tobacco smokers, one patient was over 50 years old. Surgery was considered successful when the patients had a permeable anastomosis and were able to achieve satisfactory erection resulting in normal sexual intercourse. RESULTS: All surgery was successful except for one patient who had undergone the Furlow-Fisher procedure. In Spite of antithrombotic therapy, graft occlusion occurred in two patients. Post operative glans hypervascularity occurred in two patients. CONCLUSION: Penile revascularization surgery is a highly effective treatment for selected patients. There is a need for further study of graft occlusion and glans hypervascularity. PMID- 10723179 TI - [mRNA expression of chemokines in rat kidneys with ureteral obstruction]. AB - BACKGROUND: It has been demonstrated that leukocyte infiltration, mainly of macrophages and lymphocytes, into obstructed kidneys (OBK) of rats during unilateral ureteral obstruction (UUO). Chemokines (C-C subfamily) may be involved in this mechanisms. Thus, we accessed the gene expression of chemokines in renal cortex of rats with UUO. MATERIALS AND METHODS: Female SD rats were sacrificed at various time points after UUO. mRNA expression of MCP-1, RANTES and MIP-1 alpha was determined by semi-quantitative RT-PCR. RESULTS: Control kidneys (CNK) showed a weak mRNA expression of MCP-1, RANTES and MIP-1 alpha. OBKs showed an increase in MCP-1 at 2 hours of UUO and a significant increase at 4 hours of UUO as compared with CNKs or contralateral unobstructed kidneys (CLK). The mRNA levels of RANTES and MIP-1 alpha were not increased until 72 hours of UUO in CLKs or OBKs. There were slight, but significant, differences of RANTES and MIP-1 alpha expression between OBKs and CNKs at 120 hours of UUO. CONCLUSIONS: We suggest that the early increase in MCP-1 contributes to the leukocyte infiltration and that RANTES and MIP-1 alpha plays a partial role in a late increases. PMID- 10723180 TI - [A case of ureterocalicostomy for refractory renal calculi]. AB - We have sometimes encountered intractable cases of nephrolithiasis, even though ESWL and endourology have dramatically developed at the present time. We could obtain satisfactory result in the treatment of intractable right nephrolithiasis with ureterocalicostomy. CASE: The patient was a 39-year-old man having undergone PNL, ESWL, pyelolithotomy for right nephrolithiasis. Ten-odd stones, measuring 5 20 mm in diameter, were detected, and his IVP revealed mild hydronephrosis with the ureteropelvic junction stenosis. Pyeloplasty was thought to be difficult to perform. Thus the stones were removed through an incision made on the lowermost portion of the kidney followed by ureterocalicostomy on September 9, 1993. After clamping of the renal artery, the lowermost portion of the renal parenchyma was resected, and the lower calyx was sufficiently exposed. Adequate hemostasis of the cut surface was made, and the renal artery was then declamped. Many stones, measuring 20 mm or less, were removed through the lower calyx, and the lower calyx and the ureter were anastomosed. After operation, ESWL was additionally performed for residue stones. The IVP in July 1997 demonstrated the sufficiently patent anastomosed site without hydronephrosis or recurrence of nephrolithiasis. DISCUSSION: Anastomosis between the lower calyx and the ureter is an effective therapeutic method because it dose not interfere urine stream or small stone passage. However, it is not easy to make anastomosis in this site because the calyceal wall is very fragile. Several cases have been reported to have stenosis in the anastomosed site. We supposed that it is an excellent method for the treatment of refractory nephrolithiasis if the procedure is selected only for appropriate candidates. PMID- 10723181 TI - [Segmental infarction of testicle presenting as right acute scrotum: a case report]. AB - We report a case of segmental infarction of a testicle seen in a 28-year-old man with right lower abdominal pain who visited our hospital. On physical examination, the right testicle was slightly elevated and Prehn's sign was positive. As torsion of the right spermatic cord was suspected, ultrasonography and color Doppler sonography were performed, revealing sufficient blood supply in the right spermatic cord and in the most part of the right testicle. However, a low echogenic area without blood flow was noted in the upper pole of testicle. Since CT and MRI findings couldn't rule out a testicular tumor, right high orchiectomy was performed. The specimen revealed a hemorrhagic area, histologically proved to be segmental hemorrhagic infarction of the testicle. Segmental infarction of a testicle is rare and our case is the 31st case in the world literature (the 7th case in Japan). PMID- 10723182 TI - [Traumatic brain injury in childhood: introductory remarks]. AB - Age-dependent diversity of pediatric head injury was overviewed and significance of early implementation of rehabilitation in head-injured children was emphasized. Survivors from severe traumatic brain injury (TBI) often sustains chronic physical and cognitive dysfunction. It is important for physicians and therapists to provide them with early rehabilitation treatment in a coordinated way. These children often require a long-term support from medical as well as educational specialists after finishing the hospital phase of rehabilitation. PMID- 10723183 TI - [Independent compartment phenomena: characteristics of dynamics in cerebral tissue oxygen metabolism under increased intracranial pressure in immature brain: an experimental study]. AB - The aim of this study is to clarify the specific pathophysiology of increased intracranial pressure in an immature brain in relation to its unique cerebral blood flow dynamics and brain tissue oxygen metabolism. Thirteen puppies were used for an experimental model of brain herniation due to a massive intracerebral hematoma. Along with increasing size of the hematoma, the intracranial pressure (ICP), carotid blood flow (CBF) and cerebral tissue oxygen (CTPO2) were measured simultaneously and continuously. The tolerance capacity for an acutely expanding mass lesion, or intracranial compliance, was studied. The ratio of hematoma volume/body weight was obviously higher by more than 200% in a group of younger puppies with open cranial sutures. Dynamic changes of CTPO2 were noted to be independent in the cerebral subcortical region and the medulla oblongata, when Doppler detection of arterial pulsations showed no flow in the anterior circulation in association with increased intracranial pressure caused by a supratentorial expanding mass lesion. A case with open cranial sutures (1,500 g of body weight) clearly demonstrated this and survived over 24 hours. With acutely increasing ICP CTPO2 was elevated modestly in the cerebral subcortical region (p < 0.1) and prominently in the medulla oblongata (p < 0.005). In conclusion, the posterior fossa structure (brain stem and cerebellum) in the immature age group is protected from an acutely expanding mass lesion in the supratentorial compartment. The posterior fossa behaves as an independent compartment with more prominent CBF and CTPO2 reactivity in the dynamic changes. The author proposed to name these findings as "independent compartment phenomena". PMID- 10723184 TI - [Traumatic brain injury in children]. AB - We investigated the prognosis of 42 children with traumatic brain injuries. The main etiology was a traffic accident in 46 cases, especially during walking and bicycling, and child abuse in 7 cases. Eighteen cases of acute subdural hematoma 18 cases distributed at all ages, 9 cases of diffuse axonal injury mainly during school age, 4 cases of chronic subdural hematoma under 2 years. These were all caused by child abuse. Fifteen cases showed a good prognosis with independent activities of daily living (ADL). The main type of injury was diffuse axonal injury in this group. Twelve cases showed a bad prognosis with completely dependent ADL. The bad prognostic factors were chronic subdural hematoma caused by child abuse, consciousness loss with Glasgow Coma Scale less than 8 or lasting more than 2 weeks. After rehabilitation in our hospital, 37 cases returned to school: an ordinary class in 20 cases, a special class in an ordinary school in 5 cases and a special class in 12 cases. About half of the cases returned to an ordinary class, although with problems such as learning difficulty, danger and bullying. PMID- 10723185 TI - [Rehabilitation and outcome of the adult traumatic brain injury]. AB - One-hundred and seventy traumatic brain injured persons were admitted to Kanagawa Rehabilitation Hospital for a rehabilitation program during a five-years period. The average age was 30.7 years and the male to female ratio was 5:1. The most common cause of injury was an traffic accident. They exhibited various physical and cognitive dysfunctions. Patients with severe physical and/or cognitive dysfunction had difficulty in community re-integration after finishing the hospital rehabilitation. At present, social resources for traumatic brain injury patients and family to fulfill their various needs are so few that they tend to be isolated from the community. PMID- 10723186 TI - [Brain hypothermia treatment for the management of severe pediatric brain injury]. AB - In the management of severe pediatric brain injury, attention has previously been paid to brain edema, ICP elevation and low cerebral perfusion pressure (CPP). However, in the acute stage within 3-6 hours after trauma, brain hypoxia and hyperglycemia associated with diffuse brain injury are often observed. We have pointed out brain thermo-pooling (elevation of brain tissue temperature) and brain hypoxia caused by defective release of oxygen from hemoglobin (due to decrease in red blood cell enzyme (DPG)) as a new mechanism of brain injury. To treat these pathologic changes, we have developed a brain hypothermia treatment, the major purpose of which is to prevent brain hypoxia, brain thermo-pooling, neurohormonal changes causing cytokine encephalopathy, and a selective, radical mediated damage of the dopamine A10 nervous system. The brain tissue temperature is initially adjusted to 35 degrees C with adequate cerebral oxygenation, followed by brain hypothermia at 34 degrees C for 1 weeks to prevent brain hypoxia, free radical reactions, brain edema and ICP elevation. What is most difficult in the pediatric brain hypothermia treatment is to maintain metabolic balance in the injured brain tissue and pulmonary infections associated with an immune crisis. When a rapid elevation of serum glucose is noted it is critical to lower the value because glucose quickly penetrates the blood-brain barrier and increases pyruvate and lactate by inhibiting the TCA cycle metabolism. Thus, hyperglycemia during brain hypothermia treatment is one of the major target of management. Another problem is immune crisis associated with secondary pulmonary infections. To prevent them, early enteral nutrition and replacement of L arginine were most useful, as well as preconditioning for rewarming as follows: serum albumin > 3.0 g/dl; lymphocyte > 1500/mm3; T-H (CD4) lymphocytes > 55%; serum glucose, 120-140 mg/dl; vitamin A > 50 mg/dl; Hb > 12 g/dl and 2,3 DPG, 10 15 mumol/gHb; O2 ER, 23-25% and AT-III, > 100%. The clinical benefit of this therapy is still controversial. PMID- 10723187 TI - [Management of acute-stage head trauma in childhood]. AB - Trauma victims are directly transferred to a level I trauma center bypassing local hospitals. First, airways and cervical stability are secured. Intracranial hematoma should be promptly evacuated. Endotracheal intubation and mechanical ventilation are initiated for children with a Glasgow Coma Score of 10 or less, anisocoria, apnea, and/or hypercarbia. Isotonic crystalloid is used for intravenous fluid maintenance. The goal of intracranial pressure (ICP) management is to maintain the ICP at less than 15 mmHg and to maintain minimum cerebral perfusion pressure at 45-55 mmHg. External ventricular drainage provides direct control of the ICP by allowing intermittent drainage of the CSF (5-10 ml/hour). Mannitol is effective but hyperventilation is not recommended. PMID- 10723188 TI - [Current topics of acute encephalitis and encephalopathy: introductory remarks]. AB - Encephalitis/encephalopathy is a neurological syndrome characterized by acute onset, symptoms of intracranial hypertension accompanying severe sequels or death. Encephalitis is caused by microbial infection of central nervous system, such as neurotrophic or conventional viruses. Infectious encephalopathy shows similar clinical symptoms to acute encephalitis, without any evidence of inflammation and microbial infection in brain tissues. The national epidemiological surveillance of the diseases is carried out to study the frequency and prognosis of patients with both diseases. The principal treatment is quite different in the both, in the former the eradication of microbial from the brain and in the latter the reduction of pressure of brain edema. Furthermore, the improvement of the brain with severe destruction requires such new step to reduce the activities of enzymes or cytokines to destroy brain tissues, as a mild hypothermia to lower body and brain temperature to 33-34 degrees C. PMID- 10723189 TI - [Epidemiology--surveillance information]. AB - Recent data of the National Epidemiological Surveillance of Infectious Diseases (NESID) showed that neurological complications associated with influenza virus infection, such as acute encephalitis/encephalopathy (excluding Reye's syndrome) have increased in number especially among young children. Further investigation is necessary to find the pathogenesis of these serious complications and clarify whether this phenomenon is due to unclarified factor(s) unique in Japan or it has reminded unrecognized in other countries. PMID- 10723190 TI - [Influenza encephalopathy and encephalitis]. AB - Cleavage of the hemagglutinin (HA) molecule by proteases is a prerequisite for the pathogenicity and even for the neurovirulence of influenza A viruses. WSN, a neurovirulent virus, adapted to mouse brain, grew in vitro in several types of cells including neuroblastoma cells in the absence of trypsin. When mice were intracerebrally inoculated with WSN, the viral antigen was found in the substantia nigra zona compacta and hippocampus. The mice inoculated with viruses isolated from children with acute encephalopathy associated with an influenza virus infection, on the other hand, showed no neurological symptoms. Furthermore, these viruses did not grow in the human neuroblastoma and glioblastoma cells. Since 1991, most of the human influenza A viruses have not agglutinated chicken erythrocytes. Whether this altered receptor binding specificity is related to the occurrence of influenza encephalitis and encephalopathy is now under investigation. PMID- 10723191 TI - [Infection-related acute encephalopathy: CT scan/MRI finding, laboratory examination and prognosis]. AB - CT/MRI findings, laboratory examinations and prognoses of 42 patients with acute encephalopathy (AE) (Japan Coma Scale > or = 200) were reported. 1. Findings on CT/MRI were divided into the following 7 categories: Group 1 (normal), Group 2 (CT/MRI looked normal in acute phase, but brain atrophy developed and progressed slowly by weeks or months), Group 3 (CT/MRI looked normal within a few days after the onset of AE, but cortical laminar necrosis developed at 4-5 days after the onset), Group 4 (marked brain edema developed within 2 days after the onset of AE), Group 5 (AE with symmetric thalamic lesions), Group 6 (symmetric pallidum, lesions on MRI which appeared after brain edema disappeared), and Group 7 (the brain shrinked during acute phase, which normalized on the follow up CT/MRI). 2. Serum AST elevated in approximately 50% of the patients with AE. Sixty percent of them exhibited DIC, whose prognoses were poor. Cerebrospinal fluids (CSF) neopterin (NP) and/or interleukin (IL)-6 were elevated in all the 8 patients examined. In the two cases whose serum NP and IL-6 were measured at the same time, their values in the CSF were higher than those in the serum in one case, and almost the same in the other. In a patient with a condition mimicking hemorrhagic shock and encephalopathy, serum IL-6 concentration was very high (94,000 pg/ml). 3. Mild hypothermia (around 34 degrees C) combined with methylprednisolone pulse therapy was excellently effective on AE. A 6-year-old boy exhibited tonsillar herniation at admission recovered well to be able to run. 4. Differentiation between Reye syndrome and HSE, and the pathogenesis of AE were also discussed. PMID- 10723194 TI - [A case of report of idiopathic epilepsy with combined attacks of typical absence and sylvian seizure]. PMID- 10723192 TI - [Brain thermo-pooling is the major problem in pediatric influenza encephalitis]. AB - The prognosis of pediatric encephalitides, such as infantile influenza encephalitis, is still poor because of the rapid progression, severe brain edema, selective bilateral basal ganglia necrosis, and a poor immune function, the mechanism of which is still unknown. Especially, little is known about virus es in CSF and brain tissue with influenza encephalitis, which hampers successful treatment of this condition. Recently, hypothermia treatment has attracted attention as the management of infantile influenza encephalitis to prevent severe brain edema. Recent clinical studies have revealed brain thermo-pooling (elevation of brain tissue temperature) with damage of blood-brain barrier (BBB). We then studied brain injury mechanism after severe brain injuries, cerebral strokes, reperfusion after shock, and high fever with lower cerebral perfusion pressure in our ICU. The brain thermo-pooling phenomenon results from body temperature higher than 38 degrees C, systolic blood temperature lower than 90 100 mmHg, and cerebral perfusion pressure (CPP) lower than 70 mmHg that hinders washout of brain tissue temperature by cerebral blood flow. We have recorded of brain tissue temperature of 40-44 degrees C in various brain injured patients. Some pathophysiological changes in infantile influenza encephalitis may be explained on the basis of this brain thermo-pooling phenomenon. In systemic infection, it causes severe brain edema by activation of cytokines and destruction of BBB, bilateral basal ganglia necrosis by acute severe brain hypoxia, resulting in poor prognosis without control of brain temperature. In other words, brain thermo-pooling, is the major target of treatment for infantile influenza encephalitis. In this paper, new concepts of the brain injury mechanism and methods of brain hypothermia treatment of pediatric influenza encephalitis are presented. PMID- 10723193 TI - [A case of riboflavin-responsive multiple acyl-CoA dehydrogenase deficiency (glutaric aciduria type II)]. AB - We reported a male infant with multiple acyl CoA dehydrogenase deficiency, probably due to electron transfer flavoprotein dehydrogenase deficiency. He was noted to have severe muscle weakness, a high serum creatine kinase (CK) level up to 6920 IU/L, lipid storage myopathy and fatty liver at 6 months of age. A GC/MS analysis of urinary organic acids showed excess excretion of dicarboxylic acids, including glutaric, 2-hydroxyglutaric, adipic, suberic, sebacic, malonic, ethylmalonic and methylsuccinic acids. On a urinary acylglycine analysis, hexanoylglycine and suberylglycine were increased, but not isovalerylglycine, in amount. No ketosis was noted. The muscle pathology showed increased oil-red O positive lipid droplets of various sizes indicative of lipid storage myopathy. There was diffuse decrease in the activity of cytochrome c oxidase. No ragged-red fibers were noted. His clinical symptoms improved remarkably after the administration of riboflavin (100 mg/day) and L-carnitine (1000 mg/day). He was then diagnosed as having probable riboflavin-responsive multiple acyl-CoA dehydrogenase deficiency. The glutaryl CoA dehydrogenase activity in lymphocytes was normal, as were the alpha- and beta-subunits of electron transfer flavoprotein. These findings led us to suspect electron transfer flavoprotein dehydrogenation deficiency. Although he had several episodes of short-term deterioration in clinical and laboratory findings, he developed normally with normal intelligent till 10 years of age. PMID- 10723195 TI - [Kanagawa care program for physically and mentally severe-handicapped children attending to a school for handicapped children]. PMID- 10723196 TI - False iliac artery aneurysm following renal transplantation. AB - We report a very rare case of a false iliac artery aneurysm following renal transplantation. The patient was a 51-year-old women who presented with a painful 10 x 10 cm pulsating mass in her left iliac fossa. The patient had received a second cadaveric renal transplantation 5 years previously. The graft never functioned and transplant nephrectomy was performed 2 weeks later. A CT-scanning showed a 10 x 10 cm large aneurysm arising from the left external iliac artery. At operation a large false aneurysm was identified arising from the original transplant anastomotic site. Due to the extent of the aneurysms, a Gortex graft was inserted between the external iliac artery and the common femoral artery. The patient made an uneventful post-operative recovery. PMID- 10723197 TI - Successful treatment using peroral itraconazole in a patient with acute promyelocytic leukemia complicated with splenic candidiasis. AB - A 57-year-old female with acute promyelocytic leukemia was admitted to our hospital. The PML-RAR alpha fusion transcript was reverse transcription polymerase chain reaction. Complete remission was achieved with intensive induction chemotherapy. Then a high fever unresponsive to antibiotics with increased C-reactive protein continued. Abdominal computed tomography revealed multiple low-density lesions in the spleen. Splenic candidiasis was suspected and peroral treatment with itraconazole (200 mg/day) was begun. After the fungal infection was confirmed to be inactive, splenectomy was performed. The splenic tissue showed multiple white or yellow nodules and methenamine silver stain revealed fungal hyphae characteristic to Candida. There was no evidence of relapse of Candida infection. She has been in complete remission for these two years and free of fungal infection. It is indicated from our case that splenic candidiasis in patients with acute myeloid leukemia can be successfully treated with oral administration of itraconazole and subsequent splenectomy, when it is confined in the spleen. PMID- 10723198 TI - Effect of saliva on the growth of Helicobacter pylori. AB - The effect of saliva mixed in culture media from young healthy volunteers on the growth of Helicobacter pylori (H. pylori) was investigated. Saliva was centrifuged at 15,000 rpm for 10 min and the supernatant was mixed with the reference strain of standard H. pylori which was suspended in sterilized physiological saline. The mixture was inoculated onto agar plate and incubated for 3 days. The numbers of colonies were counted. Saliva from a young male volunteer stimulated the growth of H. pylori significantly compared with the control. This result indicates that there may be such individuals who have saliva increasing the growth of H. pylori. But this is the preliminary report and more studies are needed to know what factors in saliva may affect the growth of H. pylori. PMID- 10723199 TI - Muscle cell apoptosis is responsible for the body weight loss in tumor-bearing rabbits. AB - In order to elucidate the cause of body weight loss in the early stage of tumor progression, morphological changes of striated muscle were investigated in rabbits every 10 days after VX2 carcinoma implantation. The lean body mass started to decrease in the tumor-bearing rabbits 10 days after implantation, and body fat ratio showed a significant decrease from 30 days, different from the starved rabbits, whose lean body mass and body fat ratio started to decrease from 10 days. Morphological hallmarks of apoptosis in muscle cells were detected in tumor-bearing animals prior to the tumor growth but not in starved animals. These findings suggest that muscle cell apoptosis may be responsible for the body weight loss in the early tumor-bearing. PMID- 10723200 TI - An autopsy report of graft-versus-host disease (GVHD) following blood transfusion. AB - A case of graft-versus-host disease (GVHD) associated with blood transfusion in a 71-year-old man is reported. The patient received transfusions (irradiation red cells M.A.P 14 units, fresh frozen plasma 11 units) and developed features of GVHD including fever, a skin rash, diarrhea, liver dysfunction, hypoplastic bone marrow, and pancytopenia. He died on day 32 posttransfusion. Pathologically, lymphocyte infiltration of skin and liver lesions, degeneration of small bile ducts, crypt cell necrosis in the gastrointestinal tract, and bone marrow hypoplasia were observed. Immunohistochemically, infiltrating lymphocytes were CD8+ T type, the result suggesting that they role a part in posttransfusion GVHD. PMID- 10723201 TI - Prognostic factors for brain metastasis from lung cancer after gamma knife radiosurgery. AB - To determine the factors related to optimal treatment of brain metastasis from lung cancer by gamma knife radiosurgery, we investigated those influencing survival and local tumor control. We observed 156 patients with 315 such lesions treated by gamma knife radiosurgery (GKS). The factor that most affected survival was whether extracranial disease was controlled at the time of treatment. Patients with a high Karnofsky Performance Score (KPS) tended to survive longer. The local tumor control rate was higher with tumors less than 20 mm in diameter than larger lesions. Tumors treated with a high marginal dose (23 Gy < or =) were better controlled than those treated with a lower marginal dose. Control of extranial lesions, high KPS, small (< 20 mm) lesions, and high marginal dose (23 Gy < or =) were important factors in obtaining good results with GKS for brain metastasis from lung cancer. PMID- 10723202 TI - Significant role of apoptosis in type-1 autoimmune hepatitis. AB - We used the terminal deoxyribonucleotidyl transferase-mediated deoxyuridine triphosphate-digoxigenin nick-end labeling (TUNEL) method to detect apoptosis, and immunohistochemical staining for molecules related to apoptosis, a marker of cell proliferation, and surface markers of lymphocytes to examine 20 patients with autoimmune hepatitis (AIH). Confluent hepatic necrosis was frequently found, in which rosette formation of hepatocytes and ductular proliferation were common. TUNEL staining and staining for Lewis Y antigen, Bax protein, and Fas antigen were found in biliary epithelial cells in bile ducts and proliferating atypical bile ductules in regions of confluent necrosis with severe lymphocytic infiltration. TUNEL staining and staining for Lewis Y antigen and Bax protein were found in rosette-forming hepatocytes. Many hepatocytes in lobules without injury were stained for proliferating cell nuclear antigen (PCNA). bcl-2 oncoprotein was found in many lymphocytes surrounding proliferating atypical bile ductules and rosette-forming hepatocytes in regions of confluent necrosis, in which CD20 and OPD 4 were found. Apoptosis of both hepatocytes in rosette arrangement and biliary epithelial cells in bile ducts and proliferating atypical bile ductules may play a role in progression of AIH as well as confluent hepatic necrosis. PMID- 10723203 TI - Expression of cartilage specific genes by human capsular cartilaginous cells. AB - The amount of proteoglycan production in tissue that was considered to have cartilaginously differentiated due to mechanical stress and cartilage specific gene expression were investigated. The inner layer of the joint capsule which forms a sliding surface with the femoral head in the case of dislocated hip arthoropathy showed higher proteoglycan production compared to that in the surrounding non-cartilaginous tissue. On examining gene expression, although large cartilaginous proteoglycan is originally absent in this region, gene expression of aggrecan and versican which are able to bind to hyaluronan was observed. Further, gene expression of decorin and link protein was also examined. These findings in the inner layer of joint capsule forming sliding surface with the femoral head suggested the importance of mechanical stress in cartilaginous differentiatiog of mesenchymal tissue. PMID- 10723204 TI - Phase-contrast magnetic resonance imaging study on cord motion in patients with spinal dysraphism: comparison with healthy subjects. AB - Cord motion at the cervical cord (C-3) level was measured with phase-contrast magnetic resonance imaging (MRI) in 39 healthy young subjects and 34 age-matched patients with spinal dysraphism. Cord velocity curves (composed of waves I, II, and III) were made in both sagittal images and transverse images and classified into 3 types. The measurement in the sagittal images were more reproducible than that in the transverse images. In healthy subjects, the mean cord velocity of wave I was 0.49 +/- 0.11 cm/s in the sagittal image. In the subgroup of patients with spinal dysraphism showing stable clinical courses, cord velocity was comparable to that in the healthy subjects. Patients of the subgroup with symptomatic aggravation showed a significant decrease in cord velocity in wave I, and a characteristic pattern in the cord velocity curve: the velocity change (wave I) was small. This decrease in cord velocity was thought to be due mainly to cord tethering. The measurement of cord motion with phase-contrast MRI could give objective and quantitative information about cord motion and tethering associated with spinal dysraphism. PMID- 10723205 TI - Speech perception without hearing. AB - In this study of visual phonetic speech perception without accompanying auditory speech stimuli, adults with normal hearing (NH; n = 96) and with severely to profoundly impaired hearing (IH; n = 72) identified consonant-vowel (CV) nonsense syllables and words in isolation and in sentences. The measures of phonetic perception were the proportion of phonemes correct and the proportion of transmitted feature information for CVs, the proportion of phonemes correct for words, and the proportion of phonemes correct and the amount of phoneme substitution entropy for sentences. The results demonstrated greater sensitivity to phonetic information in the IH group. Transmitted feature information was related to isolated word scores for the IH group, but not for the NH group. Phoneme errors in sentences were more systematic in the IH than in the NH group. Individual differences in phonetic perception for CVs were more highly associated with word and sentence performance for the IH than for the NH group. The results suggest that the necessity to perceive speech without hearing can be associated with enhanced visual phonetic perception in some individuals. PMID- 10723206 TI - The perception of primary and secondary stress in English. AB - Most models of word recognition concerned with prosody are based on a distinction between strong syllables (containing a full vowel) and weak syllables (containing a schwa). In these models, the possibility that listeners take advantage of finer grained prosodic distinctions, such as primary versus secondary stress, is usually rejected on the grounds that these two categories are not discriminable from each other without lexical information or normalization of the speaker's voice. In the present experiment, subjects were presented with word fragments that differed only by their degree of stress--namely, primary or secondary stress (e.g.,/'prasI/vs./"prasI/). The task was to guess the origin of the fragment (e.g., "prosecutor" vs. "prosecution"). The results showed that guessing performance significantly exceeds the chance level, which indicates that making fine stress distinctions is possible without lexical information and with minimal speech normalization. This finding is discussed in the framework of prosody-based word recognition theories. PMID- 10723207 TI - The effect of segmental order on fricative labeling by children and adults. AB - We examined whether children modify their perceptual weighting strategies for speech on the basis of the order of segments within a syllable, as adults do. To this end, fricative-vowel (FV) and vowel-fricative (VF) syllables were constructed with synthetic noises from an/[symbol: see text]/-to-/s/continuum combined with natural/a/and/u/portions with transitions appropriate for a preceding or a following /[symbol: see text]/or/s/. Stimuli were played in their original order to adults and children (ages of 7 and 5 years) in Experiment 1 and in reversed order in Experiment 2. The results for adults and, to a lesser extent, those for 7-year-olds replicated earlier results showing that adults assign different perceptual weights to acoustic properties, depending on segmental order. In contrast, results for 5-year-olds suggested that these listeners applied the same strategies during fricative labeling, regardless of segmental order. Thus, the flexibility to modify perceptual weighting strategies for speech according to segmental order apparently emerges with experience. PMID- 10723208 TI - Perceptual normalization for speaking rate. II: Effects of signal discontinuities. AB - In a series of experiments, we examined how rate normalization in speech perception is influenced by segments that occur after the target. Perception of the syllable-initial target was influenced by the durations of both the adjacent vowel and the segment after the vowel, even when the identity of the talker was changed during the syllable. These results, together with earlier findings of a temporal window that follows a target phoneme within which segment duration influences perception of the target, help to resolve apparently conflicting results that have been reported previously. Overall, the results fit within a theoretical framework in which the rate at which events take place is extracted early in processing, prior to segregating voices, and the use of this information is obligatory in subsequent processing. PMID- 10723209 TI - Tactile perception in blind Braille readers: a psychophysical study of acuity and hyperacuity using gratings and dot patterns. AB - It is not clear whether the blind are generally superior to the sighted on measures of tactile sensitivity or whether they excel only on certain tests owing to the specifics of their tactile experience. We compared the discrimination performance of blind Braille readers and age-matched sighted subjects on three tactile tasks using precisely specified stimuli. Initially, the blind significantly outperformed the sighted at a hyperacuity task using Braille-like dot patterns, although, with practice, both groups performed equally well. On two other tasks, hyperacute discrimination of gratings that differed in ridge width and spatial-acuity-dependent discrimination of grating orientation, the performance of the blind did not differ significantly from that of sighted subjects. These results probably reflect the specificity of perceptual learning due to Braille-reading experience. PMID- 10723210 TI - Temporal pattern and spectral complexity as stimulus parameters for eliciting a cardiac orienting reflex in human fetuses. AB - The purpose of this study was to determine whether temporal pattern and/or spectral complexity were important stimulus parameters for eliciting a cardiac orienting reflex (OR) in low-risk human fetuses. Each of 28 term fetuses was exposed to four sounds formed from the four different combinations of temporal pattern (pulsed, continuous) and spectral complexity (sine wave, /a/). The fetal cardiac electrical signal was captured transabdominally at a rate of 1024 Hz, and fetal R-waves were extracted by using adaptive signal-processing techniques. We found that pulsed sounds elicited a significantly greater decrease in heart rate (HR) than did continuous sounds. However, the HR response was relatively unaffected by spectral complexity. For the pure tone and the phoneme used in this study, our results indicate that temporal characteristics were more effective at eliciting a cardiac OR in human fetuses than was spectral complexity. PMID- 10723211 TI - The ventriloquist effect does not depend on the direction of deliberate visual attention. AB - It is well known that discrepancies in the location of synchronized auditory and visual events can lead to mislocalizations of the auditory source, so-called ventriloquism. In two experiments, we tested whether such cross-modal influences on auditory localization depend on deliberate visual attention to the biasing visual event. In Experiment 1, subjects pointed to the apparent source of sounds in the presence or absence of a synchronous peripheral flash. They also monitored for target visual events, either at the location of the peripheral flash or in a central location. Auditory localization was attracted toward the synchronous peripheral flash, but this was unaffected by where deliberate visual attention was directed in the monitoring task. In Experiment 2, bilateral flashes were presented in synchrony with each sound, to provide competing visual attractors. When these visual events were equally salient on the two sides, auditory localization was unaffected by which side subjects monitored for visual targets. When one flash was larger than the other, auditory localization was slightly but reliably attracted toward it, but again regardless of where visual monitoring was required. We conclude that ventriloquism largely reflects automatic sensory interactions, with little or no role for deliberate spatial attention. PMID- 10723212 TI - Infants' and adults' use of duration and intensity cues in the segmentation of tone patterns. AB - Adults and 8-month-olds were presented with sequences in which every third complex tone was either longer or more intense. Segmentation was measured by comparing the detection of silent gaps inserted into three possible locations in each pattern: Silent gaps inserted at perceived segmentation boundaries are harder to detect than gaps within perceived phrases or groups. A go/no-go conditioned head-turn (hand-raising for adults) procedure was used. In Experiment 1, detection was worse for the gaps following the longer complex tones than for the gaps at the other locations, suggesting that the longer tones marked the ends of perceived groups for both infants and adults. Experiment 2 showed that an increase in intensity did not result in any systematic grouping at either age. PMID- 10723213 TI - The effects of figure/ground, perceived area, and target saliency on the luminosity threshold. AB - Observers adjusted the luminance of a target region until it began to appear self luminous, or glowing. In Experiment 1, the target was either a face-shaped region (figure) or a non-face-shaped region (ground) of identical area that appeared to be the face's background. In Experiment 2, the target was a square or a trapezoid of identical area that appeared as a tilted rectangle. In Experiment 3, the target was a square surrounded by square, circular, or diamond-shaped elements. Targets that (1) were perceived as figures, (2) were phenomenally small in area, or (3) did not group well with other elements in the array because of shape appeared self-luminous at significantly lower luminance levels. These results indicate that like lightness perception, the luminosity threshold is influenced by perceptual organization and is not based on low-level retinal processes alone. PMID- 10723214 TI - Do McCollough effects provide evidence for global pattern processing? AB - Contingent color aftereffects (CAEs, or McCollough effects) were induced using two pairs of orthogonally related patterns (horizontal/vertical and concentric/radial) to determine whether the CAEs of the four patterns are independent. Tests using composite test patterns (like those employed by Emerson, Humphrey, & Dodwell, 1985) suggested independent aftereffects. However, tests using unitary patterns indicated additive or competing effects of the four patterns in regions where line orientations were similar, and tests isolating such regions showed clear interactions between the pattern aftereffects. The results fail to support the claim that global (rather than local) features of the patterns control these CAEs. PMID- 10723215 TI - The perceived position of the hand in space. AB - This paper reports a series of experiments of the perceived position of the hand in egocentric space. The experiments focused on the bias in the proprioceptively perceived position of the hand at a series of locations spanning the midline from left to right. Perceived position was tested in a matching paradigm, in which subjects indicated the perceived position of a target, which could have been either a visual stimulus or their own fingertip, by placing the index finger of the other hand in the corresponding location on the other side of a fixed surface. Both the constant error, or bias, and the variable error, or consistency of matching attempts, were measured. Experiment 1 showed that (1) there is a far left advantage in matching tasks, such that errors in perceived position are significantly lower in extreme-left positions than in extreme-right positions, and (2) there is a strong hand-bias effect in the absence of vision, such that the perceived positions of the left and right index fingertips held in the same actual target position in fact differ significantly. Experiments 2 and 3 demonstrated that this hand-bias effect is genuinely due to errors in the perceived position of the matched hand, and not to the attempt at matching it with the other hand. These results suggest that there is no unifying representation of egocentric, proprioceptive space. Rather, separate representations appear to be maintained for each effector. The bias of these representations may reflect the motor function of that effector. PMID- 10723216 TI - Changes in motion perception following oculomotor smooth pursuit adaptation. AB - The hypothesis that oculomotor smooth pursuit (SP) adaptation is accompanied by alterations in velocity perception was tested by assessing coherence thresholds, using random-dot kinematograms before and after the adaptation paradigm. The results showed that the sensitivity to coherent motion at 10 deg/sec (the initial target velocity during adaptation) was reduced after the SP adaptation, ending up at a level that was between those normally observed for velocities of 10 and 20 deg/sec. This is consistent with an overestimation of the velocity of the coherent motion and suggests that SP adaptation alters not only the oculomotor output, but also the perception of target velocity. PMID- 10723217 TI - Categorical perception occurs in newly learned faces, other-race faces, and inverted faces. AB - On the basis of findings that categorical perception (CP) is possible in complex visual stimuli such as faces, the present study tested for CP on continua between unfamiliar face pairs. Results indicate that CP can be observed for unfamiliar faces, in both familiar (same-race) and unfamiliar (other-race) groups. In addition, significant CP effects were observed in inverted faces. Finally, half continua were tested where midpoint stimuli became endpoints. This was done to ensure that stimulus artifacts did not account for the observed CP effects. Consistent with the perceptual rescaling associated with CP, half-continua showed a rescaled CP effect. We argue that these CP effects are based on the rapid acquisition of perceptual equivalence classes. PMID- 10723218 TI - Spelling-sound regularity effects on eye fixations in reading. AB - An interaction of word frequency and word regularity has typically been observed in naming and lexical decision experiments in which, in addition to an overall effect of word frequency, responses to low-frequency exception words are slower than those to low-frequency regular words, while no such difference occurs with high-frequency words. The only eye movement study to examine this effect in reading (Inhoff & Topolski, 1994) reported only transient effects of regularity. In the present experiment, we examined the frequency x regularity interaction using different stimuli than those of Inhoff and Topolski and also varied the parafoveal preview of the target word prior to fixation. When the preview was valid, the frequency x regularity interaction appeared. However, with an invalid preview, the effect of regularity disappeared. The results suggest that the activation of phonological codes is a very early component of reading. PMID- 10723220 TI - [41st Congress of the German Society of Pneumology. Hamburg, 1-4 March 2000. Abstracts]. PMID- 10723219 TI - Cross-modal selective attention: on the difficulty of ignoring sounds at the locus of visual attention. AB - In three experiments, we investigated whether the ease with which distracting sounds can be ignored depends on their distance from fixation and from attended visual events. In the first experiment, participants shadowed an auditory stream of words presented behind their heads, while simultaneously fixating visual lip read information consistent with the relevant auditory stream, or meaningless "chewing" lip movements. An irrelevant auditory stream of words, which participants had to ignore, was presented either from the same side as the fixated visual stream or from the opposite side. Selective shadowing was less accurate in the former condition, implying that distracting sounds are harder to ignore when fixated. Furthermore, the impairment when fixating toward distractor sounds was greater when speaking lips were fixated than when chewing lips were fixated, suggesting that people find it particularly difficult to ignore sounds at locations that are actively attended for visual lipreading rather than merely passively fixated. Experiments 2 and 3 tested whether these results are specific to cross-modal links in speech perception by replacing the visual lip movements with a rapidly changing stream of meaningless visual shapes. The auditory task was again shadowing, but the active visual task was now monitoring for a specific visual shape at one location. A decrement in shadowing was again observed when participants passively fixated toward the irrelevant auditory stream. This decrement was larger when participants performed a difficult active visual task there versus fixating, but not for a less demanding visual task versus fixation. The implications for cross-modal links in spatial attention are discussed. PMID- 10723221 TI - [Women's health--a separate medical problem?]. PMID- 10723223 TI - [Can the exercise test with nitroglycerin be useful for differentiation of patients with and without changes in coronary arteries--syndrome X?]. AB - The aim of this study was to compare the results of ECG exercise test performed before and after oral administration of nitroglycerine (NTG) in patients with coronary artery disease (CAD), and patients with typical chest pain without any changes in coronary arteriography--syndrome X. We examined 98 patients with typical chest pain, positive result of ECG exercise test, then accordingly to results of coronary arteries assessed with coronary arteriography, patients (pts.) were divided into two groups: group 1--35 pts. without any changes in coronary arteriography--syndrome X, and group 2--48 pts. with significant stenosis present in one or more coronary vessels. Each patient underwent two ECG exercise tests: first without any medication and second performed average 30 minutes after first test, and 5 min after oral administration of 1 table of nitroglycerine. During both tests the following parameters were evaluated: test duration, presence of chest pain, max. ST-T changes, heart rate (HR), and systolic blood pressure (SBP). RESULTS: In group 1 after NTG time of test duration had shortened from 5.9 +/- 0.4 min to 5.7 +/- 0.6 min. We also observed an increase in max. ST-T complex depression (2.2 +/- 0.5 mm vs 2.4 +/- 0.4 mm) but these differences were not statistically significant. In CAD group, duration of test after NTG was longer (6.2 +/- 1 vs 7.4 +/- 1.2), and normalization of max. ST-T complex depressions (2.7 +/- 0.5 vs 2.0 +/- 0.3 mm) was observed p < 0.01. CONCLUSION: Our study suggests that ECG exercise test with NTG may be useful in differentiation of patients with syndrome X and patients with typical coronary artery disease. PMID- 10723222 TI - [Usefulness of measuring serum IGG antibodies against A60 mycobacterial antigen for diagnosis of tuberculosis]. AB - Measurement of antimycobacterial antibody may be used as potential diagnostic tool in tuberculosis. The aim of the study was to evaluate the diagnostic value of serum IgG level against A60 mycobacterial antigen measured by ELISA method. Material consisted of 144 persons divided into 5 groups (76 tuberculosis patients, 20 sarcoidosis patients, 17 lung cancer patients, 8 patients with mycobacterial infections other than tuberculosis and 23 healthy controls). In the tuberculosis group there were 50 culture positive cases and 26 culture negative ones, 43 new cases and 32 chronic cases. Positive results were obtained in 51% of tuberculosis patients. Sensitivity increased to 62% in culture positive group and 63% in chronic cases. Specificity of the test was 96%. The results indicate that Immunozyme Mycobacterium test is a valuable tool in tuberculosis diagnosis. PMID- 10723224 TI - [The level of erythropoietin in serum of patients with anemia during infective endocarditis]. AB - The infective endocarditis is a septic syndrome caused by an infection in endocardium or in heart valves. The majority of patients with infective endocarditis develop normocytic anemia. The metabolic studies in septic shock syndromes documented an intensive proteolysis of muscles, visceral organs and blood proteins, and probably of erythropoietin as a glycoprotein as well. The aim of the study was to assess the erythropoietin level in patients with infective endocarditis severe anemia and preserved renal function. Erythropoietin concentration was measured in blood serum in 12 patients (11 men and 1 woman), mean age 48 +/- 8 years, with infective endocarditis. The patients had clinical symptoms of endocarditis, positive blood bacteriological cultures and echocardiography features. All patients had serious normocytic anemia with mean hemoglobin concentration 5.40 +/- 0.48 mmol/L. The control group consisted of 7 healthy persons (5 men and 2 women), mean age 50 +/- 7 years, with hemoglobin concentration 8.70 +/- 0.60 mmol/L. The concentration of erythropoietin at the patients with bacterial endocarditis was 144.04 +/- 17.80 mIU/mL versus 67.28 +/- 6.29 mIU/mL in the control group (p = 0.0002). We conclude that in patients with infective endocarditis and serious normocytic anemia without renal insufficiency the concentration of erythropoietin is increased. PMID- 10723225 TI - [Quality of sleep and periodic breathing in healthy individuals working at an altitude of 3700 meters]. AB - We performed full polysomnography (PSG) in 7 healthy miners of Kyrghyz origin (mean age 25 +/- 6 years) working in 2 weeks shifts at Kumtor gold mines at the elevation of 4200 m. They slept in comfortable dormitories situated at 3700 m. To avoid acute mountain sickness all subjects received acetazolamide 3 x 0.25 daily during 2 days preceding ascent and during 2 days at altitude: PSG was performed three times: at 760 m (1) and on the 1st (2) and 7th night (3) after rapid ascent (aircraft) to high altitude using SomnoTrac 4250 sleep laboratory. We found that sleep efficiency was good at lowland and in the mountains averaging 81.79% and 84% respectively. Although there were no significant differences in percentage of sleep stages and of total sleep time between lowland and both nights at high altitude, arousals and awakenings were more frequent in the mountains. Episodes of periodic breathing (PB) appeared at high altitude. There was a large individual variability in PB on both nights at altitude. The time spent in PB ranged from 4 to 30 minutes during the first night at altitude and from 3 to 17 minutes during the second one. PB appeared mainly during non-REM sleep and aggravated arterial blood desaturation. PMID- 10723226 TI - [Detection of pituitary autoantibody in patients with pituitary tumors and hypopituitarism]. AB - In 80 patients, 73 cases of pituitary tumours and 7 cases of hypopituitarism, we performed pituitary autoantibodies assays in serum samples because in our previous studies we had found a high prevalence of pituitary autoantibodies in several autoimmune endocrine disorders. To detect the presence of pituitary autoantibodies we applied 2 methods, radioimmunoassay (RIA) and immunoblotting. The RIA was performed by solid phase technique in human pituitary microsome coated polyethylene tubes. Following incubation with diluted sera of the patients labelled 125I-protein A was added to the tubes to detect the retained antibodies. In the sera of 33 patients we detected the presence of antibodies; in the other 47 patients no antibodies were found. The majority of the patients with positive antibody results were previously treated by pituitary irradiation. To evaluate the molecular weights of pituitary autoantigens the microsomal proteins were separated on SDS PAGE, then electrophoretically transferred to nitrocellulose membranes and reacted with diluted sera of 30 antibody-positive patients. The nitrocellulose strips were incubated with labelled 125I-protein A and autoradiographed. Using immunoblotting, 13 out of these 30 patients we found autoantibodies reacting with pituitary microsomal antigens of different molecular weights, most frequently reacting with a 68 kDa autoantigen. CONCLUSIONS: The prevalence of pituitary autoantibodies in patients with pituitary diseases is 41% lower than in autoimmune endocrine diseases. Pituitary autoantibodies usually appear in patients after pituitary irradiation or after neurosurgery followed by irradiation, but occur rarely in untreated patients with pituitary adenomas. PMID- 10723227 TI - [Bone mineral density in primary hyperparathyroidism]. AB - Skeletal complications of advanced hyperparathyroidism include clinically bone pains, muscle weakness, bone deformities and fractures. X-ray studies reveal subperiosteal bone resorption, atrophy of the cortex of long bones, cysts, brown tumours and calcifications of soft tissues; these changes appear in the late period of the disease. In recent onset of hyperparathyroidism bone changes may be detected by X-ray absorptiometry. Thus the aim of our study was to evaluate bone mineral density with the use of dual energy X-ray absorptiometry (DEXA) at two sites: in lumbar vertebral bodies consisting mainly of the trabecular bone and in 1/3 distal part of the radius composed predominantly of the cortical bone. Twenty three patients with primary hyperparathyroidism were included in our study. Hypercalcemia (ionized calcium above 5.4 mg/100 ml, total calcium above 10.6 mg/100 ml) and increased serum PTH, above 100 pg/ml, were detected in all patients. Isotope scintigraphy using 99mTc-MIBI revealed the presence of a parathyroid adenoma; this was confirmed at surgery and histopathologically. In bone densitometry we found greatly reduced bone mineral density (BMD) in 1/3 distal part of the radius amounting to 66.8 +/- 12.0% of the age-matched range and markedly smaller bone loss in lumbar spine, BMD was 91.7 +/- 14.6%. In 10 patients control densitometry, performed 6-24 months after parathyroid adenomectomy, revealed a marked 10 to 22% increase in bone density of lumbar vertebral bodies in the first year. BMD of the 1/3 distal part of the radius increased to a smaller degree 6.3% per year. CONCLUSIONS: 1. Bone densitometry in primary hyperparathyroidism reveals pronounced decrease in bone mineral density in the 1/3 distal part of the radius and much smaller decrease of the lumbar spine density. 2. Parathyroid adenomectomy leads to a rapid increase in density of the trabecular bone L1-L4 vertebral bodies and much smaller increase in the cortical bone of the radius. 3. Pronounced differences in bone mineral density of cortical bone and trabecular bone surpassing 20% are characteristic of hyperparathyroidism as they do not occur in other types of osteoporosis. PMID- 10723228 TI - [Treatment of severe uremic hyperparathyroidism using a method for percutaneous injection of the parathyroid glands with ethanol]. AB - The results of recent studies suggest that uremic patients with large parathyroid hyperplasia are often resistant to active vitamin D3 therapy. Percutaneous ethanol injection has become an interesting option in such cases, although there are only a few publications on that subject. In this work we would like to present our experience with this method. 20 patients with serum iPTH > 400 pg/ml and 1-4 hyperplastic parathyroids (mean volume 1.07) underwent 56 percutaneous ethanol injection sessions under ultrasonographic guidance. In 9 patients a marked (> 75%), long-term (12-24 months) decrease in serum iPTH was achieved; lesser (> 50%) reduction in parathyroid activity persisted for 36-42 months in 5 out of 9 patients observed in this period. In almost every patient a significant reduction of alphacalcidol dose was possible. Our data confirm that percutaneous ethanol injection therapy is a useful and safe adjunct in severe uremic hyperparathyroidism treatment strategy which allows to restore the responsiveness to active vitamin D3 metabolites. PMID- 10723229 TI - [The level of triglycerides, total cholesterol and HDL cholesterol in various stages of human immunodeficiency virus (HIV) infection]. AB - In 63 HIV-infected individuals the levels of concentration plasma triglicerides, total cholesterol and HDL cholesterol were determined by the level of immunological deficiency defined according to the CD4 lymphocytes count. The analyzed markers of lipid disorder allow to draw the following conclusions: 1. Along with the increase of immunological deficiency and clinical development of the HIV infection the disorders, which are indicated by the increase in trigliceride levels and decreased plasma concentrations for HDL cholesterol, intensify as well. 2. HIV-infected persons, particularly those treated with protease inhibitors, should have carefully monitored lipid metabolism. PMID- 10723230 TI - [Diagnostic difficulties in cases with spurious thrombocytopenia]. AB - Pseudothrombocytopenia (PTP) is a rare laboratory phenomenon of falsely low platelet count in the presence of anticoagulant, most often EDTA. It may be confused with true thrombocytopenia, leading to the refusal of further invasive procedures and inappropriate diagnosis and treatment. PTP is due to presence of antiplatelet antibodies--agglutinins--usually temperature dependent. Here we present two cases of such spurious thrombocytopenia and discuss the differential diagnosis. PMID- 10723231 TI - [Neurologic complications in Wegener's granulomatosis]. AB - Nervous system involvement is relatively frequent in Wegener's granulomatosis (WG). It may be difficult to differentiate between the primary central nervous system involvement and complications secondary to concomitant arterial hypertension, renal insufficiency and iatrogenic effects of immunosuppressive therapy. The crucial role in the assessment of intracranial pathology may be ascribed to the diagnostic imaging techniques: magnetic resonance imaging (MRI), computed tomography (CT) and cerebral blood flow imaging utilising the single photon emission computed tomography (SPECT). SPECT may prove superior sensitivity to MRI. It may be especially useful in differentiating central nervous system involvement in WG with secondary changes of other origin. PMID- 10723232 TI - [Somatostatin analogues in diagnosis and management of neuroendocrine tumors]. PMID- 10723233 TI - [Morphology, diagnosis and treatment of Zollinger-Ellison syndrome]. PMID- 10723234 TI - [Professor Ivan Duris, awarded the medal of the Polish Society of Internal Medicine]. PMID- 10723235 TI - [Mechanisms of pathogenesis and differentiation-induction in acute promyelocytic leukemia]. PMID- 10723236 TI - [Characteristics of blood cells during peripheral blood stem cell mobilization following chemotherapy in patients with non-Hodgkin's lymphoma]. AB - The objective of our study was to obtain a predictor of the timing for peripheral blood stem cell (PBSC) collection using routine blood cell counts. For that purpose, we compared recovery kinetics of peripheral blood cells and CD34+ cells in non-Hodgkin's lymphoma patients who were given granulocyte-colony stimulating factor for PBSC collection after CHOP therapy. Peaks in the monocyte/WBC ratio and reticulocyte high fluorescence ratio (HFR) were observed 2 to 3 days prior to the peak CD34+ cell count. In addition, the appearance of immature granulocytes in peripheral blood correlated well with increases in CD34+ PBSC number. These findings suggested the peak monocyte/WBC ratio, HFR, and immature granulocyte count can be used as predictors of the timing for PBSC harvests. PMID- 10723237 TI - [Total hip and knee arthroplasty for arthropathy in a hemophiliac]. AB - We evaluated the medium-term follow-up results, effectiveness, and suitability of arthroplasty for hemophilic arthropathy. We performed 26 total knee and 9 total hip arthroplasty operations on hemophilic patients between 1988 and 1998 under general anesthesia and appropriate hemostatic management. Postoperative treatment for hemophilic arthropathy is the same as that for osteoarthritis and rheumatoid arthritis. After their operations, patients experienced relief from pain and intra-articular bleeding in affected joints but only marginal improvement in the range of motion. In general, total joint arthroplasty is not indicated for young patients. However, arthropathy can have a severe impact on the active life of patients during their youth. For severe hemophilic arthritis, total knee and hip arthroplasty can lead to a pain-free and improved quality of life. We believe total knee and hip arthroplasty is a good solutions for hemophiliac arthropathy before severe deformity occurs. Although treated relatively young hemophilic patients, age was not considered to be a contraindication. PMID- 10723238 TI - [Treatment of hepatic veno-occlusive disease after bone marrow transplantation with recombinant human tissue plasminogen activator (rh-tPA)]. AB - Ten children were treated with recombinant human tissue plasminogen activator (rh tPA) for severe hepatic veno-occlusive disease (VOD) that developed after bone marrow transplantation. Treatment with rh-tPA was begun a median of 22 days (range; 13-127 days) after transplantation. Seven of 9 (78%) evaluable patients had complete resolution of their VOD. Four patients had hemorrhagic complications, and 2 of them died because of pulmonary hemorrhage and subdural hemorrhage, respectively. Although rh-tPA seems to be an effective therapy for established VOD, further studies will be necessary to determine its safety as well as the optimal dosing regimen. PMID- 10723239 TI - [A randomized trial of conventional dose vs reduced dose in BHAC-DM therapy for acute myelogenous leukemia in elderly patients]. AB - Twenty-nine patients aged 60-75 years with newly diagnosed acute myelogenous leukemia (AML) were randomized to receive BHAC-DM either at a reduced dose (S-1 group, n = 13; BHAC 150 mg/m2 1-7 day, DNR 30 mg/m2 1-3 day, 6MP 70 mg/m2 1-7 day) or the conventional dose (S-2 group, n = 16; BHAC 200 mg/m2 1-7 day, DNR 40 mg/m2 1-3 day, 6MP 70 mg/m2 1-7 day). On day 7, patients were given therapy for 2 more days if the ratio of blasts in their bone marrow was more than 15%. Granulocyte-colony stimulating factor was injected when the leukocyte count decreased below 1,000/microliter. The rates of complete remission were 46.2% in the S-1 group and 43.8% in the S-2 group. No significant differences in response distinguished the 2 groups. The mortality rate during myelosuppression was 1/13 in the S-1 group and 1/16 in the S-2 group. The rate of treatment-related death was 10.1% for all patients. Grade-4 adverse effects were not seen in any of the patients. We concluded that the conventional dose of BHAC-DM was as acceptable as the reduced dose in elderly patients with AML. PMID- 10723240 TI - [Clinical evaluation of rhG-CSF in patients with neutropenia induced by chemotherapy for multiple myeloma]. AB - A randomized controlled study of patients with multiple myeloma was performed to evaluate the efficacy and safety of recombinant human granulocyte colony stimulating factor (rhG-CSF:KW-2228) in treating neutropenia induced by chemotherapy, and its influence on the dose intensity of, and response rate to, chemotherapy. As a rule, 3 courses of chemotherapy at intervals of 4 weeks were administered both to the untreated and KW-2228-treated groups. Among 98 eligible patients evaluated for neutrophil recovery, a markedly reduced duration of neutropenia was observed during each course in the KW-2228 treated group. No significant difference distinguished the two groups in terms of incidence or duration of infection. However, febrile neutropenia appeared only in the untreated group. There was no significant difference in terms of response rate or dose intensity. However, only patients in the untreated group withdrew from the study due to protracted neutropenia. These results demonstrated that KW-2228 is effective and safe, and has a significant effect on the acceleration of neutrophil recovery in patients with neutropenia induced by chemotherapy for multiple myeloma, and is useful for the completion of chemotherapy regimens. PMID- 10723241 TI - [Werner's syndrome associated with acute myelofibrosis]. AB - A 47-year-old man was admitted to our hospital in June 1997 because of nasal bleeding. He presented with anemia in addition to physical characteristics of Werner's syndrome (WS). Peripheral blood examination disclosed pancytopenia with 4% blasts. Bone marrow aspiration was a dry tap; biopsy specimens revealed myelofibrosis. Chromosomal analysis of peripheral blood revealed hypodiploidy with complex abnormalities including -5 and del(7)(q21). Serum levels of PDGF, FGF, and TGF beta 1 were normal. A diagnosis of acute myelofibrosis was made. The patient's condition became quickly deteriorated and he died of pneumonia in October 1997. In the literature, we found 6 reported cases of myelofibrosis associated with WS. Considering that only approximately 1,100 cases of WS have been reported so far, the incidence of myelofibrosis in WS seems relatively high. This case suggested a link between WS and myelofibrosis, and the mechanism of myelofibrosis in WS was discussed. PMID- 10723242 TI - [Development of arterial thrombus of Mucorales hyphae during deferoxamine therapy in a patient with aplastic anemia in transformation to myelodysplastic syndrome]. AB - A 58-year-old woman with a diagnosis of aplastic anemia had been treated with anabolic steroid for mild anemia in 1984. In May 1995, pancytopenia progressed and the patient became dependent on red blood cell transfusions. Chromosome analysis of bone marrow cells revealed trisomy 8 and the patient was thought to have aplastic anemia in transformation to myelodysplastic syndrome. In July 1996, deferoxamine was administered for iron overload. The patient was admitted because of pneumonia on July 31, 1998. Chest computed tomograms showed arteriothrombus of the right pulmonary artery and pulmonary mycosis. Although an antifungal agent was administered, the patient experienced respiratory failure and eventually died of hypovolemic shock due to gastric bleeding from a Dieulafoy ulcer. Autopsy revealed arteriothrombus of hyphae of Mucorales in the right pulmonary artery and right renal artery branch. Cases of mucormycosis occurring in dialysis patients receiving deferoxamine have recently appeared in the literature. Deferoxamine may be a risk factor for mucormycosis. Deferoxamine has been also used in the treatment of iron overload patients with aplastic anemia. Four cases of mucormycosis developing in such patients have been reported in Japan including this case. There may be a relationship between mucormycosis and deferoxamine in patients with aplastic anemia. PMID- 10723243 TI - [Rapidly progressive, refractory eosinophilia with a 250,000/microliter eosinophil count]. AB - A 31-year-old man had received corticosteroids for 20 months for treatment of a brain tumor, and his blood eosinophil count ranged from 100/microliter to 1,000/microliter. On June 24th, 1998, he was re-admitted because of dyspnea secondary to left massive pleural effusion. Peripheral blood examination revealed an eosinophil count of 48,000/microliter. The eosinophils were hypersegmented, with abnormal distribution of eosinophilic granules and formation of cytoplasmic vacuoles. Blasts and basophils were not increased, hemoglobin was 13.4 g/dl, and the platelet count was 79,000/microliter. Bone marrow was slightly hypercellular with 55% eosinophils and 0.2% blasts. The patient's karyotype was normal, and Wilms' tumor gene was not detected. Serum IgE was normal and serum vitamin B12 and soluble IL-2 receptor were elevated. Serum levels of eosinophilopoietic cytokines, IL-3, IL-5, and GM-CSF, were low. Specimens of pleural fluid contained many eosinophils. Because the eosinophil count increased to 110,000/microliter on July 2nd, hydroxyurea was started without effect. On July 16th, the eosinophil count reached 167,000/microliter, and vincristine was added. The eosinophil count rose to 253,000/microliter the next day, and cytarabine and daunorubicin were administered, but the patient died of septic shock. Although the clinical course suggested eosinophilic leukemia, monoclonal proliferation of eosinophils was not demonstrated. To our knowledge, this is the highest peripheral blood eosinophil count reported in the literature to date. PMID- 10723244 TI - [Syndrome of inappropriate antidiuretic hormone secretion associated with meningeal infiltration of tumor cells and elevated interleukin-1 beta and interleukin-6 in cerebrospinal fluid of a patient with adult T-cell leukemia]. AB - A 73-year-old man with acute adult T-cell leukemia (ATL) in remission was re admitted to our hospital due to drowsiness, headache, and bilateral knee joint pain on May 17, 1998. On admission, examinations revealed decreased serum sodium concentration (112 mEq/l), low plasma osmotic pressure (259 mOsm/l), and elevated antidiuretic hormone(5.6 pg/ml). Cerebrospinal fluid examination showed an increased number of abnormal flower-like lymphocyte (951/microliter). Brain computed tomography and magnetic resonance imaging found no abnormality in the hypothalamus or pituitary gland. These findings yielded a diagnosis of syndrome of inappropriate antidiuretic hormone secretion (SIADH). Though ATL patients typically exhibit a variety of clinical symptoms, SIADH is rarely one of the complications. Further investigation showed that IL-1 beta and IL-6 concentrations were increased in spinal fluid but not in serum. Recently, it has been reported that exogeneous IL-6 is an inducer of ADH secretion, and that primary ATL cells and HTLV-I infected cell lines can produce IL-6. In this case, we speculated that IL-6 produced by ATL cells that infiltrated a cerebral lesion may have played an important role in the development of SIADH. PMID- 10723245 TI - [Perforation of small intestinal during hematologic recovery in an elderly man after induction therapy for acute lymphoblastic leukemia L3]. AB - A 67-year-old man with a 7-month history of dilated cardiomyopathy was admitted to our hospital because of general fatigue, shortness of breath, and anemia on laboratory examination. Increased blasts were observed in the bone marrow. The blasts were characterized by large cells with abundant, intensely basophilic, vacuolated cytoplasm, round nuclei, and prominent nucleoli. Chromosome analysis revealed a nonrandom t(8;22)(q24;q11) chromosomal abnormality, and surface-marker analysis disclosed a positive immunophenotype for CD10, CD19, CD20, CD38, HLA-DR, FMC7, and IgM-lambda. These findings yielded a diagnosis of L3 acute lymphoblastic leukemia. The patient was treated with chemotherapeutic agents. On the 39th hospital day, during hematologic recovery after induction therapy, abdominal pain developed. Abdominal X-ray films disclosed ileus with dilatation of the small bowel and Kerckring's folds. Conservative treatment was begun but the patient died. At autopsy, intestinal perforations were observed at a site 55 cm proximal to the ileocecal junction. A specimen of perforated tissue revealed a diffuse infiltration of leukemic cells through the small bowel wall. However, bone marrow specimens showed no signs of aggravation of leukemia. PMID- 10723246 TI - [Chromosome 1 abnormalities at band 1p32 in two atomic bomb survivors with myelodysplastic syndrome]. AB - We reported on 2 atomic bomb survivors(a 60-year-old man and 63-year-old woman)suffering myelodysplastic syndrome(MDS) associated with 1p32 chromosomal abnormalities. They were exposed to atomic bomb radiation at distances of 1.2 and 1.1 km, respectively, and were given a diagnosis of MDS 44 and 46 years after the bombing, respectively. The male patient had refractory anemia(RA) and a bone marrow cell karyotype of 46, XY, del(1)(p22p32), t(8;11)(p11;p15). The female patient had RA with excess of blasts (RAEB) and a karyotype of 45, X, -X, t(1;11)(p32;q23), +del(1)(p32), inv(3) (p21q27), del(5)(q15), -6, -9, -19, +mar 1, +mar 2. Multi-separated nuclear megakaryocytes were observed in both patients. These findings suggested that they had been exposed to radiation near the atomic explosion despite the fact that their symptoms of MDS developed more than 40 years after the bombing. 1p32 is known to be the locus of the TAL1 gene. However, Southern blot analysis did not reveal rearrangement of the TAL1 gene in the male patient. PMID- 10723247 TI - Sensible RAHC plan needed. PMID- 10723248 TI - Environmental symposium criticized. PMID- 10723249 TI - "To err is human.". PMID- 10723250 TI - MedBytes. PMID- 10723251 TI - 48 hours. PMID- 10723252 TI - Patient privacy. PMID- 10723253 TI - Ethics of authorship. PMID- 10723254 TI - Scientific and legal issues in fenfluramine/dexfenfluramine litigation. AB - Fenfluramine and dexfenfluramine were popular and widely used antiobesity agents until they were withdrawn from the market in 1997. Even though early research appeared to demonstrate their safety, serious concerns were raised about these medications. Primary pulmonary hypertension (PPH) was a known side effect, but it was believed that the health benefits of weight loss compensated for the risk of PPH. With widespread use of these agents, 2 other conditions--valvular heart disease and neurotoxicity--were reported as potential side effects. In this paper, we review the evidence for these adverse events and whether the current data meet federal and Texas legal standards for admissibility. We discuss also the basis for health claims against physicians, and the scientific and legal challenges faced by both plaintiffs and defendants. PMID- 10723255 TI - [The South Urals Research Center of RAMN]. PMID- 10723256 TI - [Molecular and cellular mechanisms of erythropoiesis regulation]. AB - The paper presents the results of recent studies of the molecular and cell cellular mechanisms of physiological regulation of the differentiation and proliferation of erythroid cell precursors and the reproduction of erythropoietin in the kidneys, hormone-sensitive cell reception, the mechanisms of transduction of an erythropoietic signal from the receptor of a cell to its genome, the mechanisms of erythropoietic cell-cellular interactions in the bone marrow. PMID- 10723259 TI - [Tissue regeneration and growth during exposure to graded directed mechanical loads]. AB - The paper gives the results of experimental morphological and biochemical studies made under the author's supervision in the subdivisions of the Kurgan Branch of the South Urals Research Center, Russian Academy of Medical Sciences, which death with distraction osteogenesis in long tubular, flat, and spongy bones, with the mechanism of osteogenesis and the modes of its management by mechanical and biochemical factors, with the use of methods for computer aided image analysis, three-dimensional reconstruction, and stereological analysis for quantitative characteristics of osteogenesis, with the definition of indications for its correction and with evaluation of its efficiency, with the clinical use of experimental developments in vertebrology and craniosurgery. PMID- 10723258 TI - [Role of neutrophils in regulation of immune responsiveness and reparative reactions of damaged tissue]. AB - After both thermal and mechanical injuries, mice are shown to have depressed functions of phagocytes (primarily neutrophils), suppressed ability of an immune response to antigens and decreased skin inflammatory and reparative activities. It is suggested that a neutrophilic secretory defect and a decrease in the production of stimulating factors might be a cause of the above effects. After thermal and mechanical injuries, injection of neutrophilokin extracted from latex activated neutrophilic supernatant into the mice promoted recovered phagocytic functional activity, higher humoral and cellular immune response and increased the rate of standard burn wound healing. Neutrophilokin also accelerated clavus formation in mice with mechanical injury. Thus, it can be concluded that neutrophils and their low-molecular secretory products play an important role in regulating immune and inflammatory reparative homeostasis in thermal and mechanical injuries. PMID- 10723257 TI - [Erythropoiesis and functional characteristics in bone marrow erythroblastic islets during stimulated adn inhibited erythropoiesis]. AB - When erythropiesis is stimulated (acute blood loss) or inhibited (posttransfusion polycythemia), there are early changes in the cytochemical values of erythroblastic islets (EI): in the levels of acid and neutral glucoconjugates and in the activity of nonspecific esterase. A close correlation has been found between the erythropoiesis in EI and its functional characteristics. It is concluded that central macrophages play the key role in the modulation of EI erythropoiesis. It is suggested that EI macrophages are involved in the provision of bioenergetic and reparative processes in EI. PMID- 10723260 TI - [Growth and differentiation of epi- and perineural structures under graded stretching]. AB - Light and electron microscopic and ultrastructural studies of the epi- and perineurium of sciatic and tibial nerves were performed in 63 adult mongrel dogs during lengthening the femur and leg at rates of 0.5 and 1 mm a day (no more than 0.25 mm per dose) by the Il-izarov procedure. Hypertrophy of the biosynthetic apparatus, complication of intercellular and cytostromal contacts were observed in the fibroblasts of the epineurium, in the cell elements of vascular walls and perineurium during distraction. There was growth and associated changes in the volume ratios of the wall components of epineural structures, as well as new formation of structures of the intrinsic innervation apparatus of epi- and perineural sheaths. At all levels (organ, tissue, cellular, and subcellular) the new formed structures incorporated into the preexisting ones without disintegrating them, which is a characteristic feature for intercalar growth. PMID- 10723261 TI - [Regulation of distractional osteogenesis: biochemical aspects]. AB - The samples of intact compact bone, bone tissue of new-born puppies (BP), distractional regenerate bone (DRB), new cancellous bone (NCB) filling in the transported fragment were fractionated by the degree of maturity while substituting canine tibial shaft defect using bifocal distraction-compression osteosynthesis. Comparative analysis of the composition of the mineral phase and the organic matrix of different derivatives of bone tissue revealed the identity of low-mineralized fractions of BP and NCB. It was suggested that the fractions were produced by DRB cells which were phenotypically different from osteogenic ones. There is evidence that the high proliferating potential of the least mature DRB fraction is maintained by relative hypoxia in the "growth zone" of the latter and by the changing concentrations of local factors depending on the rate of distraction. PMID- 10723262 TI - [Responsiveness and resistance of spinal cord structures in patients with closed thoracic and lumbar spinal injuries: neurophysiological and clinical aspects]. AB - The study was undertaken to explore the time course of parameters of neuromotor dysfunction in patients with thoracic and lumbar spinal fractures characterized by the varying degrees of neurological symptoms. The study was based on the results of complex neurophysiological testing (global and stimulation electroneuromyography (EMG) in 45 patients with thoracic and lumbar spinal fractures who had been admitted to the "VTO" Russian Research Center without complications. The patients were divided into 2 groups: 1) 17 patients without neurological disorders and 2) 28 with mild neurological ones. There was evidence that there were no complicated vertebral injuries. Group 1 patients were found to have steady-state changes in the EMG structure, lower voluntary and involuntary activities (M responses) of the muscles of the hip, leg, and foot, enhanced reflex excitability of leg muscles, EMG signs of spasticity and irritation of segmental radicular structures, long-term asymmetry virtually in all EMG parameters. It was also ascertained that the group of patients with uncomplicated vertebral fractures was represented by individuals having more fitness or those belonging to the so-called "muscular" somato-type. The findings lead to the conclusion that the VTO treatment of vertebrospinal injuries, that is based on the use of a refinement of an external spinal fixation apparatus, creates necessary prerequisites for prevention of further development of neurological deficit and for the optimal course of compensatory-reparative processes in the damaged spinal cord structures. PMID- 10723263 TI - [Mechanism of laser radiation action at tissue and cellular levels]. AB - The paper describes the action of low-intensity laser radiation (LILR) and high intensity one (HILR) on biological tissues. LILR produces its action on cells by changing the membranous formations-receptors and ion channels. At cardiac surgery, HILR stimulates uncontractile myocardial elements, by inducing the secretion of biologically active substances and the activation of enzyme systems that ensure the implementation of neoangiogenesis in the ischemic myocardium. PMID- 10723264 TI - [Mechanisms of persistence in bacterial pathogens]. AB - This paper gives an original classification of persistent bacterial mechanisms, that is based on the protection (isolation) of peptidoglycan, by recognizing the host immune system. The classification includes a bacterial cell wall screening and production of secreted agents, inactivating host defense, as well as antigenic mimicry, and bacterial wall-free formations (L forms). A group of novel bacterial secreted agents by suppressing host defense is described. PMID- 10723265 TI - [Role of hypothalamic nonapeptides in interaction of prokaryotic and eukaryotic cells]. AB - The role of hypothalamic nonapeptides in the interaction of prokaryotic and eukaryotic cells was studied in the experimental setting. Nonapeptides were found to stimulate the adaptive and regenerative properties of eukaryotic cells and they are likely to have an antimicrobic effect on prokaryotic ones. The paper discusses the modulating role of nonapeptides in the establishment of symbiotic relations in the bacterial agent-host system. PMID- 10723266 TI - [Rapid reduction of a variety of organic functionalities including a new selective reduction with samarium diiodide]. AB - This review deals with the rapid reduction of a variety of organic functionalities, including a new selective reduction of carboxylic acid, with samarium diiodide in the presence of additives under mild conditions. The single electron donor ability of samarium diiodide can be enhanced with ligands around Sm2+ when sufficient electrons are supplied from the ligands. Mechanistic consideration of this enhancement led to our finding that carboxylic acid, ester, amide, nitrile, pyridine and other functionalities were rapidly reduced with samarium diiodide in the presence of a base, acid, water and methanol as ligands at room temperature in good yields. Particularly, these systems directly reduced carboxylic acid into alcohol and a samarium diiodide--85% phosphoric acid system immediately reduced an aromatic primary amide to aldehyde in good yield. Pyridines were reduced to piperidines with a samarium diiodide-base or water system in excellent yields; also, the aromatic nucleus of phenol derivatives was rapidly reduced with a samarium diiodide-base system. Furthermore, these organic functionalities were rapidly reduced with an Sm or Yb metal-hydrochloric acid system. Aliphatic and aromatic carboxylic acids with a coexisting aldehyde or that bearing a formyl group were selectively reduced to the corresponding alcohols with a new samarium diiodide-samarium triflate-methanol-base or water system within a few minutes at room temperature in good to almost quantitative yield. PMID- 10723267 TI - [Targeting therapy using a monoclonal antibody against tumor vascular endothelium]. AB - Recent studies have revealed that the targeting therapy using monoclonal antibody against tumor associated antigens did not have a clinically satisfactory effect due to various physiological characters of tumor. We propose a novel approach targeting tumor vascular endothelium to solve the inefficiency of common tumor missile therapy. In this study, the tissue distribution of anti-tumor vascular endothelium monoclonal antibody (TES-23) produced by immunizing with plasma membrane vesicles obtained from isolated rat tumor-derived endothelial cells (TECs) was assessed in various tumor-bearing animals. Radiolabeled TES-23 dramatically accumulated in KMT-17 fibrosarcoma, a source of isolated TECs after intravenous injection. In Meth-A fibrosarcoma, Colon-26 adenocarcinoma in BALB/c mice and HT-1080 human tumor tissue in nude mice, radioactivities of 125I-TES-23 were also up to fifty times higher than those of control antibody with little distribution to normal tissues. Furthermore, immunostaining of human tissue sections showed specific binding of TES-23 on endothelium in esophagus and colon cancers. These results indicate that tumor vascular endothelial cells express a common antigen in different tumor types of various animal species. In order to clarify the efficacy of TES-23 as a drug carrier, an immunoconjugate, composed of TES-23 and neocarzinostatin, was tested for its antitumor effect in vivo. The immunoconjugate (TES-23-NCS) caused a marked regression of the tumor, KMT-17 in rats and Meth-A in mice. Thus, from a clinical view, TES-23 would be a novel drug carrier because of its high specificity to tumor vascular endothelium and its application to many types of cancer. PMID- 10723268 TI - [Analysis of the mechanism for the anti-inflammatory effect of the anti-rheumatic drug auranofin]. AB - Effects of auranofin, an orally active chrysotherapeutic agent, were examined on the production of prostaglandin E2 (PGE2) and nitric oxide (NO) in rat peritoneal macrophages and in RAW 264.7 cells, a murine macrophage-like cell line. Auranofin (1-10 microM) inhibited PGE2 production in rat peritoneal macrophages stimulated with 12-O-tetra-decanoylphorbol 13-acetate (TPA, 16.2 nM) at 8-20 h, but did not affect PGE2 production at 4 h. However, in non-stimulated rat peritoneal macrophages, auranofin increased PGE2 production at 4 h and had no effect on PGE2 production at 8-20 h. It was proved that auranofin (1-10 microM) increased COX (cyclooxygenase)-1-dependent PGE2 production and inhibited COX-2-dependent PGE2 production in rat peritoneal macrophages. Auranofin showed no effect on the enzyme activities of the purified COX-1 and COX-2 proteins. Furthermore, auranofin did not affect the COX-1 protein level, but inhibited the TPA-induced expression of COX-2 protein. Therefore, it was suggested that auranofin inhibited PGE2 production by inhibiting the COX-2 protein induction in TPA-stimulated macrophages. In RAW 264.7 cells, auranofin (0.3-3 microM) inhibited lipopolysaccharide-induced NO synthesis by inhibiting the induction of NO synthase (NOS) protein expression. Auranofin did not affect the enzyme activity of iNOS (inducible NOS). Finally, using rat peritoneal macrophages, the effects of auranofin on PGE2 production and NO production were determined. Auranofin (10 microM) strongly inhibited the production of PGE2 and NO, and the induction of COX-2 protein and NOS protein by TPA. Indomethacin, a COX inhibitor, partially inhibited NO production at the concentration at which PGE2 production was completely inhibited. On the other hand, L-NG-monomethyl-L-arginine acetate (L NMMA), a NOS inhibitor, partially inhibited PGE2 production. NO production was completely inhibited at the same concentration as shown above. These findings suggest that PGE2 production and NO production partially affect each other. Therefore, the inhibition of PGE2 production by auranofin might be partly due to the inhibition of NO production, and the inhibition of NO production by auranofin be partly due to the inhibition of PGE2 production. In conclusion, auranofin inhibits both PGE2 production and NO production by inhibiting the upregulation of mRNA levels of COX-2 and NOS. PMID- 10723269 TI - [Tubular localization and drug recognition of kidney-specific organic anion transporters, OAT-K1 and OAT-K2]. AB - The renal proximal tubular cells play a principal role in limiting or preventing toxicity by actively secreting organic anions from the circulation into the urine. We isolated a cDNA coding a novel rat kidney specific organic anion transporter, OAT-K1, mediating transport of methotrexate (MTX). Moreover, we have isolated a new cDNA coding a rat OAT-K2, showing the 91% amino acid identity with OAT-K1. In this study, the mRNA distribution along the nephron segments and the transport characteristics of OAT-K1 and OAT-K2 have been analyzed. By the use of a reverse transcription-coupled PCR, OAT-K1 and OAT-K2 mRNAs were detected predominantly in the proximal straight tubules. When expressed in Xenopus oocytes, OAT-K1 mediated the uptake of MTX and folate, but not of taurocholate (TCA) and prostaglandin E2 (PGE2), although OAT-K2 stimulated the uptake of MTX, folate, TCA and PGE2. In stable transfectants (MDCK OAT-K1 and MDCK-OAT-K2), each transporter was localized functionally to the apical membrane and showed transport activity similar to those in the oocytes. The efflux of preloaded MTX was enhanced in both MDCK-OAT-K1 and MDCK-OAT-K2 cells. These results suggest that both OAT-K1 and OAT-K2 are apical membrane bidirectional organic anion transporters, and participate in epithelial transport of lipophilic organic anions in the kidney. PMID- 10723270 TI - [Studies of metals and metallothionein in tissue]. AB - This paper is a review of three topics related to bio-trace metals. First, the transfer of metals into tissues of patients with chronic diseases treated with hemodialysis is examined. Such diseases include chronic hepatitis, diabetes, and chronic renal failure. In these diseases, metal contents from fingernails were flexible but non-specific. Toxicity may appear as the amount of heavy metals in tissues of patients with chronic renal failure treated with hemodialysis. For example, cadmium and lead were not excreted from the blood of patients during the hemodialysis treatment, and, therefore, their amounts gradually increased in the blood of patients. The level of zinc increased and was excreted in the urine of diabetic patients and experimental animals. Calcium accumulated in the kidney of streptozotocin (STZ)-induced diabetic rats that were fed low zinc diets; and, as a result, severe renal failure occurred. From these results, complication syndromes of either metal deficiency or excesses may occur in tissues of patients with chronic diseases. Second, the role of metallothionein (MT), an inducible protein, and the properties of MT isoforms have been studied on experimental animals. In the exocrine cells of the pancreas, MT was induced by various stresses such as zinc, STZ, alloxan and 4-aminopyrazolo-(3,4-d) pyrimidine, but the effects of those stresses were not clear in the endocrine cells. Therefore, MT may have a role in the exocrine cells of the pancreas. In addition, we were able to separate completely MT-1 and MT-2 isoforms in cytosol fractions of tissues using a capillary zone electrophoresis system at neutral pH without any detergents. Each role of the MT isoforms in the tissues soon started to become clear. Third, cisplatin, a platinum-containing anti-tumor drug, did not penetrate into the brain tissue under physiological conditions, as there is a blood-brain barrier to cerebral tissues; however, it did penetrate with either short-term hypoxia or in the case of lipopolysaccharide-treated experimental animals. Nitric oxide, prostaglandin, and free radicals are related to the penetration. Older rats had a higher sensitivity to cisplatin than younger rats. PMID- 10723271 TI - [Crystallographic characteristics of crystal habits and their peculiarity to the substance (study of crystalline drugs by mesns of a polarizing microscope. XVIII)]. AB - Crystallographic and optical characteristics of crystal habits have been investigated comparing the both results of goniometric and newly developed polarizing microscopic methods. It has been found that the predominant faces of crystal habits are mostly formed by (001) or (100) of orthorhombic or monoclinic system and (010) of monoclinic system, and a little part by triclinic (001). Not so many numbers of predominant faces were found to be formed by orthorhombic or monoclinic (110) making prismatic habits. Key refractive indices have been found to be uniquely measured from those habits, and it was also found that one or two of them were coincided with the principal refractive indexes, which will become important facts for the analytical use of crystal habits. PMID- 10723272 TI - [Absorption of a vaginal contraceptive, nonoxynol (polyoxyethylene nonylphenyl ether) and its metabolism to nonylphenol in female rabbits]. AB - After intravaginal administration of a spermicide, nonoxynol (polyoxyethylene nonylphenyl ether, NPE) in female rabbits, the pharmacokinetics of NPE were examined by the HPLC method. The plasma levels after administration of NPE revealed a considerable amount of absorption of NPE into the circulation and the bioavailability after intravaginal administration was calculated to be 66% by comparison with that after intravenous administration. Unchanged NPE was not excreted into the urine in significant amounts even on intravenous administration. Nonylphenol (NP), a presumed metabolite of NPE, was simultaneously analyzed using GC-MS to assess the risk by its endocrine disrupting effects. Although the NP concentrations in the plasma were all below the lower limit of quantitation (10 ng/ml), small amounts of NP and its glucuronide conjugate were detected in the urine after intravaginal administration of NPE. Thus it was suggested that at least part of NPE absorbed in the circulation was metabolized to give NP. However, the sum of NP and its conjugate excreted in the urine was very small amounts (0.22% of dose). Therefore, it was assumed that the production of NP was not in the major pathway of the NPE metabolism. PMID- 10723273 TI - [Hepatotoxicity induction in mice after acute DL-lactic acid intake]. AB - DL-Lactic acid and its salts are added to food as acidulants, pH control agents, leavening agents, nutrient supplements and seasonings. However, the basic data concerning the safety and toxicity of these compounds are insufficient. In this article, we examined induction of hepatotoxicity and nephrotoxicity in mice after acute intake of DL-lactic acid. Body weight change, serum glutamic pyruvic transaminase (SGPT) activity, serum urea nitrogen (SUN) concentration, liver and kidney weights, and renal lipid peroxide level could not be affected significantly in mice at 4 h after intraperitoneal administration of DL-lactic acid at 1.2 mmol/kg, indicating no induction of toxicity in the liver and kidney. In contrast, at 20 h after the treatment, SGPT activity, liver weight and lipid peroxide level were enhanced significantly, suggesting induction of hepatotoxicity. However, SUN concentration, kidney weight and lipid peroxide level could not be affected significantly at 20 h after the treatment, indicating no induction of nephrotoxicity. PMID- 10723274 TI - [In vivo toxicity, lipid peroxide lowering, and glutathione, ascorbic acid and copper elevation induced in mouse liver by low dose of oxine-copper, a fungicide]. AB - While oxine-copper (OxCu) is generally used as an agricultural fungicide and induces a harmful effect on ecosystems, little information is available regarding a toxic effect of OxCu on mammals. In this article, we examined in vivo induction of toxicity and change of levels of glutathione and ascorbic acid, major biological antioxidants, lipid peroxide and copper (Cu) in liver and kidney 4 h and 24 h after intraperitoneal administration of OxCu at a low dose (0.05 mmol/kg) to mice. Increased hepatic ascorbic acid and Cu levels were found at 4 h after the treatment. In addition, body weight change was lowered and serum glutamic pyruvic transaminase activity was elevated significantly compared to control at 24 h after the treatment, suggesting induction of systemic and hepatic toxicity respectively. These were accompanied by lowered lipid peroxide level and enhanced glutathione, ascorbic acid and Cu levels in the mouse liver. On the other hand, OxCu induced no elevation in serum urea nitrogen concentration 4 h and 24 h after the treatment, suggesting no induction of nephrotoxicity, accompanied by no change in renal lipid peroxide, glutathione, ascorbic acid and Cu levels. These results suggest that hepatic Cu elevation may induce hepatotoxicity and no renal Cu elevation may lead to no induction of nephrotoxicity after the treatment with OxCu. PMID- 10723276 TI - Collaboration in nursing PMID- 10723275 TI - [In vivo toxicity, and glutathione, ascorbic acid and copper level changes induced in mouse liver and kidney by copper(II) gluconate, a nutrient supplement]. AB - While copper(II) gluconate (CuGL) is generally used as a nutrient supplement for infant foods and as an oral deodorant, little information is available regarding a toxic effect of CuGL on mammals. In this article, we examined in vivo induction of toxicity and change of level of glutathione and ascorbic acid, major biological antioxidants, lipid peroxide and copper (Cu) in liver and kidney 4 h after single intraperitoneal administration of CuGL at 0.05 and 0.10 mmol/kg to mice. Serum glutamic pyruvic transaminase (SGPT) activity, an indicator of hepatotoxicity, significantly increased compared to control in proportion to doses of CuGL. Hepatic level of glutathione measured as nonprotein sulfhydryl was not proportional to CuGL doses, but enhanced after dosing of 0.05 mmol/kg and lowered by 0.10 mmol/kg. Like SGPT activity, serum urea nitrogen (SUN) concentration, an indicator of nephrotoxicity, significantly increased in proportion to doses of CuGL. Renal glutathione level was not different from control after dosing of 0.05 mmol/kg and lowered by 0.10 mmol/kg. In both organs, relative organ weight and lipid peroxide level were not affected by the treatment with CuGL; ascorbic acid level was elevated after dosing of 0.05 mmol/kg and was not different from control after treatment with 0.10 mmol/kg; like SGPT activity and SUN concentration, Cu level significantly increased in proportion to doses of CuGL. These results suggest that in the liver and kidney after the treatment with CuGL Cu accumulated may induce toxicity, leading to level changes of glutathione and ascorbic acid and to no induction of oxidative damage. PMID- 10723277 TI - Developing new nursing roles. PMID- 10723278 TI - Screening the vision of school-aged children: an interdisciplinary research approach. AB - The authors are registered nurses (RNs) and members of an Interdisciplinary Vision Screening Research Group investigating how best to screen children's vision, incorporating the assessment of binocular visual function. The aims of this pilot study were: to detect any visual skill problems of children in a Reception class (n = 28) using the Oyarzun Vision Screening Kit; to describe the behaviour of the children during the screening; to calculate the time taken to screen each child and to establish the interrater reliability (IRR) between the RNs and an optometrist. Thirteen of the children (46%) had at least one visual problem, necessitating follow-up assessment. Other results and research implications are also discussed. PMID- 10723279 TI - A descriptive study of nurses employed by general practitioners in south-east Queensland. AB - The aim of this study was to describe the demographic and occupational characteristics of a sample of nurses employed by general medical practitioners and the factors perceived to be influential in their role development. Telephone and mail surveys were undertaken within one general practice division in South East Queensland. Thirty-seven of the 67 (55%) practice nurses responded to the mailed questionnaire. Of these respondents, ten were collectively interviewed to elaborate on the survey results. Findings indicated that the primary work of these nurses is one of assistant to the doctor. Autonomous nursing initiatives are largely opportunistic. Perceived barriers to role expansion included Medicare restrictions, inadequate basic and ongoing education programs, financial and space limitations of the practice, reluctance of general practitioners, and a lack of professional support. PMID- 10723280 TI - Midwives and women in partnership: the ideal and the real. AB - This qualitative study sought to identify the strategies used by hospital based midwives in assisting women to manage pain in labour, as well as the support they offer in all aspects of the birth process. Data were collected via participant observation of five birthing mothers and their midwives, and pre and post birth interviews. Analysis was by means of the software package QSR NUD.IST (Non numerical, Unstructured, Data, Indexing, Search and Theorizing) Version 3. Analysis of observation of labour and pre and post birth interviews revealed that all women wanted a birth with minimal intervention, however safety was an overriding consideration. During labour women relied on the midwife to oversee and direct care, and trust in care by a midwife was established as a result of prior experience, or during antenatal care. Factors inhibiting full partnership between women and midwives, and also the use of alternative pain strategies, included staffing levels, lack of emphasis on evidence based care and environmental constraints such as the room set up. PMID- 10723281 TI - Patient education Web sites. PMID- 10723282 TI - Twenty first century patient education. PMID- 10723283 TI - Identifying sources of stress and job satisfaction in the nursing environment. AB - As nursing has previously been identified as a stressful occupation (Tyler and Cushway 1992; 1995), sources of job stress and levels of job satisfaction were investigated in a study of occupational stress with 129 Victorian nurses. The study aimed to identify what the nurses perceived as the major causes of stress in the workplace using a standardized questionnaire, Gray-Toft and Anderson's Nursing Stress Scale (NSS), and by way of written reports. Level of job satisfaction was measured using the Nursing Stress Index (Harris Hingley and Cooper 1988). Results showed that the nurses rated their workload as highly stressful in terms of both frequency of its occurrence and its perceived effect upon themselves. As expected, higher levels of reported nursing stress were associated with lower levels of job satisfaction. Analyses of the written descriptions of a recent stressful work episode provided by 66 of the nurses included examples of relevant nursing stressors that were not covered by the NSS. Findings from the qualitative data are also discussed in terms of the implications for nursing practice arising from the impact of workload, and professional interpersonal conflicts. PMID- 10723284 TI - Perceptions of nursing students on men entering nursing as a career. AB - The purpose of this study was to examine the perceptions of nursing students who were enrolled in an under-graduate nursing programme in a rural university, on men entering nursing as a career. Questionnaires were returned by 38 males (63%) and 117 females (50%). The major findings were that male nursing students intend to work in specialty areas such as emergency nursing, operating room nursing and intensive care. Both male and female nursing students perceived that the public displayed an increasing respect and acceptance of male nurses. Differences between male and female nursing students were evident in demographic characteristics, course interests, career aspirations, motivating factors and barriers associated with males choosing a nursing career. PMID- 10723285 TI - Surgery, pain, and immune function. AB - Research findings have demonstrated that surgery negatively affects immune function. Pain also has deleterious effects on immune function. Most research in this area has been conducted on animals. Psychoneuroimmunology provides the framework for this article. Psychoneurological phenomena, such as stress, depression, and pain, can influence immune system functioning through neuroendocrine pathways. Biological and behavioral processes appear to be closely related and need to be jointly considered when planning and providing anesthesia care. The anesthesia planning process increasingly involves aggressive assessment and treatment of postoperative pain, which may benefit immune function. PMID- 10723286 TI - Pain in human immunodeficiency virus/acquired immune deficiency syndrome patients: a review for nurse anesthetists. AB - Pain is the most common reason for the hospitalization of people with human immunodeficiency virus/acquired immune deficiency syndrome (HIV/AIDS). The epidemiology of this disease and limitations of available therapies suggest that HIV/AIDS pain will be a treatment challenge within and beyond the perioperative realm for some time to come. This paper examines HIV/AIDS pain by major body systems. Suggestions are provided for pain assessment in HIV/AIDS patients. Both pharmacological and nonpharmacological treatment of pain in this complex population are described. PMID- 10723287 TI - Human immunodeficiency virus/acquired immune deficiency syndrome-related drug therapy: anesthetic implications. AB - This article reviews advances in the diagnosis and treatment of human immunodeficiency virus/acquired immune deficiency syndrome (HIV/AIDS). Newer drugs used to treat HIV/AIDS, such as protease inhibitors and non-nucleoside reverse transcriptase inhibitors, are described, including their potential interactions with anesthetic drugs. Common areas of concern for CRNAs related to these drugs are reviewed, such as end-organ toxicity, liver and kidney effects, and bone marrow suppression. PMID- 10723288 TI - Current perspectives on the perioperative management of the latex-allergic patient. AB - The increasing incidence of latex allergy necessitates thorough preanesthetic screening for risk factors, which will be delineated in this article, that are associated with latex allergy. The pathophysiology, epidemiology, and testing procedures for latex allergy will be reviewed. This case report will illustrate the management of a patient who was found to be latex-sensitive during surgery and the management of intraoperative anaphylaxis is provided. Safe perioperative care can be provided for latex-sensitive patients if latex avoidance techniques are used consistently. PMID- 10723289 TI - Alterations in immune response associated with anxiety in surgical patients. AB - Anesthesia practitioners have begun to focus on the immune function of their patients as more research is done on the interface between anesthesia, surgery, and immune alterations. The anxiety associated with anesthesia and surgery produces alterations in immune function through several mechanisms which affect recovery from surgery and wound healing. Immune status may be assessed by traditional measures such as complete blood count and differential as well as using newer technologies such as flow cytometry, lymphocyte proliferation assays and natural killer cell cytotoxicity. Anxiety produces immune changes through sympathetic adrenal medullary and hypothalamic pituitary adrenal cortical mechanisms, as well as through the neurotransmitter substance P. Anxiety also produces changes in immune function through alteration in health behaviors such as increased smoking, increased alcohol consumption, drug use, and changes in diet and sleep. Recommendations for psychological and pharmacological measures that are effective in reducing stress-induced immunosuppression among this group of patients are provided. The future use of substance P antagonists, which are currently under investigation, as well as cytokine manipulation, hold promise for further measures to reduce immune alterations associated with anxiety. PMID- 10723290 TI - Anesthetic drug interactions. Quarterly update. PMID- 10723291 TI - Regulation of health care professionals, Part 2: Validation of continued competence. AB - Today's demand for professional accountability regarding patient outcomes of care stems from two principle concerns: (1) the risks associated with health care services and their delivery, and (2) health care cost-containment and wanting to get the best care with the best patient outcomes at the best cost. Whereas patient outcomes are a function of multiple factors, health care professionals and their practice are considered key to those outcomes. Today's society and particularly the payers for health care services, have grown skeptical of professions and their willingness to do a good job of self-policing their own members, incompetent performers. In the 1970s, this early skepticism and the concurrent malpractice crisis led professionals and state regulators over time to move beyond the one-time testing for a lifelong credential so long as you didn't get into major problems, to considering alternatives for assuring continued competency in practice. The first efforts focused on voluntary or mandatory requirements for continuing education as a basis of attesting to continuing or recredentialing (licensure or certification). Unfortunately, this continuing education process has never been validated, demonstrating its efficacy in assuring continued professional competence and currency of knowledge as it plays out in practice and patient outcomes. Reasonable assumptions are no longer publicly acceptable; evidence is being demanded. Also, a problem that has plagued private credentialers has been the legal concept of a license or certification as a lifelong property right based on the such conditions when it was awarded. Credentialing bodies have been at legal risk in removing a license or certification for failure to comply with new criteria, particularly if the criteria has not been validated as making a difference. Rather than looking solely at currency of knowledge and competence in its utilization (including the associated technology), it is now being recommended that a multifaceted methodology be used in assessing those professional attributes deemed essential for achieving quality patient outcomes. PMID- 10723293 TI - Combined spinal-epidural anesthesia/analgesia. AB - Combined spinal epidural anesthesia offers the advantages of each method while minimizing their respective disadvantages. First described in 1937, this technique has risen in popularity over the last 15 years and is being used successfully in orthopedic, urologic, and gynecologic surgeries and for anesthesia/analgesia for labor and delivery as well as cesarean section. The history and development of combined spinal epidural anesthesia/analgesia, the different techniques, and controversies and problems associated with its use are discussed. The use of the technique of obstetric anesthesia/analgesia is also examined. PMID- 10723292 TI - Differential diagnosis of postdural puncture headache in the parturient. AB - In the obstetric setting, spinal and epidural analgesia/anesthesia are the 2 most frequently used forms of analgesia and anesthesia. One of the potential complications of these procedures is the postdural puncture headache (PDPH), and there is a high probability that the anesthetist will have occasion to evaluate the headache complaints of the parturient. The author reviews the differential process and discusses some of the causes and treatments of headaches in the parturient. PMID- 10723294 TI - Placenta percreta: report of a case. AB - Placenta accreta, increta, or percreta are rare but potentially lethal obstetric emergencies. Removal of abnormal growth of the placenta into the uterine wall is difficult or impossible and results in massive blood loss. Hysterectomy may be necessary to save the mother's life. The common predisposing factors in development of placenta percreta are repeat cesarean and placenta previa. The diagnosis of placenta percreta may remain undiagnosed until delivery. The case presented describes a scenario involving placenta percreta with bladder involvement in which the diagnosis was known in advance. The article describes the preoperative preparation, intraoperative events, and postoperative status of this particular case. PMID- 10723295 TI - Accuracy of blood loss determination by health care professionals. AB - Patients undergoing surgery will likely experience some degree of blood loss. There is much literature examining effects of blood loss, but little was found that examined accuracy of estimation of blood loss. The research question for this study was: How accurate are surgical health care professionals in their estimations of blood loss? This study was a pre-experimental between-subject design that used a convenience sample of 85 volunteers who worked in the surgical and postsurgical units of a rural southern 450-bed hospital. The participants viewed 1 of 3 randomly chosen samples of laparotomy pads with variable amounts of blood and saline. Only the researchers knew the exact amount contained on the pads. The variables that were examined and were compared included the professional group, years of experience in surgery or the postanesthesia care unit (PACU), and their estimation of blood loss. Their estimation of blood loss was compared with the actual amount of blood to determine whether one group was more accurate than another statistically and whether increasing years of experience improved accuracy. The statistical tests used were simple and multiple regressions. PMID- 10723297 TI - Extrauterine abdominal pregnancy: report of a case. AB - A healthy, 34-year-old, gravida 3, para 1,011, patient presented for cesarean delivery in her 35th week of gestation with a diagnosis of complete placenta previa. During her 26th week of gestation, the patient was admitted to a high risk obstetric unit with the diagnosis of premature rupture of membranes. Numerous ultrasonographic studies were conducted throughout her 10-week hospital stay, confirming the admitting diagnosis. A routine cesarean section was planned, and preparations were made for a potential increase in blood loss related to the placenta previa. The procedure began under spinal anesthesia and, upon incision of the abdomen, an extrauterine pregnancy was identified. The patient was immediately anesthetized and intubated at the request of the surgeon. During the 3-hour surgical procedure, the patient sustained massive blood loss, transfusions, central line placement, and aggressive pharmacological therapy. The patient was extubated the day after surgery, and was discharged approximately 1 week later. The only major complication was compartment syndrome of the left upper extremity related to the infiltration of vasopressors requiring fasciotomy and closure 2 days later. The incidence, morbidity/mortality, and anesthetic implications of abdominal pregnancy are reviewed. PMID- 10723296 TI - Meperidine utilization and compliance with Agency for Health Care Policy and Research guidelines in a tertiary care hospital. AB - The Agency for Health Care Policy and Research (AHCPR) established guidelines for the use of meperidine (demerol), a common inpatient analgesic. These guidelines define standards of care for acute and chronic cancer pain management and address many of the problems with meperidine and its metabolite, normeperidine. The purpose of this study was to determine whether meperidine was prescribed in compliance with AHCPR guidelines, whether patients exhibited any adverse reactions to meperidine, and to determine the analgesic efficacy of meperidine. Three hundred inpatient charts were reviewed and identified meperidine as the primary analgesic in 157 nonobstetric inpatients. Age, sex, weight, dosing interval, route of administration, duration of meperidine use, serum chemistry values, primary diagnosis, associated medical conditions, and medications concurrently being taken with meperidine were the parameters analyzed. An interview was conducted to ascertain medical and drug history, chronicity of pain syndromes, analgesic drug history, and analgesic efficacy. A visual analog scale for pain (range = 0 to 10) and an analgesic satisfaction survey (range = 1 to 5) were used. Of 157 patients, 124 (79.8%) were in conflict with AHCPR guidelines. The most frequent conflict was found to be suboptimal dosing regimen and treatment of chronic pain. Often concurrent analgesics were given with the meperidine to achieve adequate analgesia. Higher analgesic satisfaction scores were noted when meperidine was given with concurrent analgesics. Meperidine also was administered to patients in renal failure or with medications contraindicated with meperidine use. No significant adverse effects were noted with meperidine use in this sample population other than an increased incidence of confusion in the elderly population. PMID- 10723298 TI - Anesthetic drug interactions. Quarterly update. PMID- 10723299 TI - Health care delivery and change: thoughts on Lema's "... of dinosaurs, dodos and anesthesia personnel". AB - Problems in health care delivery relative to access, costs, and quality have been debated for more than a quarter of a century. Health care costs have significantly increased since the implementation of the Medicare/Medicaid legislation. Cost containment has been high on the agendas of government officials, legislators, health policy decision makers, business leaders, and economists since the 1980s. There has been a shift toward market medicine and managed care as a means for cost containment. Although some costs were contained for a short period, they are once again rising significantly, and there is growing dissatisfaction with this shift. The United States is not alone in this dilemma. Mark Lema, MD, PHD, editor of the ASA Newsletter, wrote a thought provoking editorial in the July 1999 issue, raising concerns about change, relationships, reimbursement, and demise relative to anesthesia personnel. In response, this article primarily raises the issue of health manpower mix as a major factor in the cost of health care delivery regarding these systems. Whereas change is inevitable, it is difficult for state and federal governments in the United States to force change because of the number of special interests involved in campaign financing involving elected government officials. It is nevertheless important for health professionals to be involved in the changes that come about, or change will be made for them. It is essential to renew society, institutions, and individuals in order to prevent decay and obsolescence. If we don't make the future, the future will make us. PMID- 10723300 TI - Blood flow displayed by dialysis machines: is it accurate? AB - Blood flow is a major determinant of dialysis efficiency. We reported in 1980, at the EDTNA conference in Prague (1), that the actual blood flow may be much lower than displayed by the dialysis machines. We demonstrated that the Negative Inflow Pressure (NIP), induced by the arterial needle and therefore generally referred to as arterial pressure, partly collapses the proximal blood pump segment whose inner volume is decreased. Thus the actually pumped volume per revolution is lower than the displayed blood flow. PMID- 10723301 TI - IV 3000 dressing on Permcath exit sites. AB - With an increasing number of elderly clients now being accepted onto our haemodialysis programme, Permcath (Quinton Instrument Co., Seattle, WA, USA) catheters have become the chosen form of long term vascular access in many of these people. With this has come the need not only to adapt, but where necessary change the previous format of covering dressings at the exit site of these catheters. PMID- 10723302 TI - New location for polytetrafluorethylene graft. AB - Cimino-Brescia performed in 1966 an internal by-pass between an artery and a peripheral vein resulting in an internal arteriovenous fistula (FAVI). Since then, it has been the most used and first chosen vascular access for haemodialysis. Due to the regular use of the access, as well as the patients' chronic condition and the associated pathologies, vascular access can become exhausted, necessitating continuous research and new alternatives to be resorted to. PMID- 10723303 TI - Infection control: do we all have a role to play? AB - Infection control became a recognised discipline in 1970. Its principles, however, had been in existence for some time. In the middle of the nineteenth century, Semelweiss showed the importance of hand washing in coping with the transmission of infections. Better acceptance of Semelweiss's idea came after Pasteur, Lister and Koch related asepsis to the prevention of the spread of infection. Florence Nightingale, on the other hand, at about the same time, made a remarkable contribution to sanitation and isolation practices. PMID- 10723304 TI - HIV and dialysis: is it a real risk? AB - The need to expound this work was born of the necessity to confront all the organisational difficulties that present themselves with the arrival of a HIV+ patient for dialysis in our centre. On such occasions, in fact, we were unable to find a protocol, guidelines or a text that could help us in a complete and adequate way, to resolve the doubts, the fears and the practical, ethical and legal problems that we suddenly had to face. PMID- 10723305 TI - Hygienic plan for a haemodialysis unit: structure and responsibilities. AB - Prevention of therapeutic environment-related contamination and infection of patients and staff has had a high priority in renal care delivery since the beginning. Instructions, recommendations and advice are provided in various rules and guidelines deemed to protect patients and staff. References can be found often in working policies and quality standards of care. PMID- 10723306 TI - Ethical considerations: educational programme for assisting staff. AB - With the advancement of dialysis technique, it has become possible to treat older patients with more complicated conditions. More and more often, staff have to face difficult treatment decisions. This has led to many questions, concerning for example the indications for life support and the risk of "over-treatment". Patients, relatives, doctors, nurses and social workers all have different ways of looking at these problems. PMID- 10723307 TI - Non-compliance to treatment: can professionals ethically refuse to treat? AB - Professionals who work in health care have long been aware of patients who continually fail to adhere to treatment regimes recommended to them by their nurses, doctors and dieticians. This is no less a problem within the field of renal medicine. In the current climate where who to treat often causes more of a dilemma than how to treat, professionals in renal care frequently deliberate the fairness of treating those who persistently "non-comply". PMID- 10723308 TI - CE marking & the MDD: what difference will the new regulations make? AB - CE marks are symbols used to show that products comply with regulations, or 'Directives', issued by the Council of Ministers of the European Commission. The EC Directives cover all kinds of products, from toys to construction materials. They are intended to make it easier for manufacturers to sell their products throughout the European Community. There will be three directives for medical devices. PMID- 10723309 TI - Use of erythropoietin in emergencies: massive intoxication by chloramines. AB - Recombinant human Erythropoietin (rHuEPO) is normally used to correct anaemia in patients with End Stage Renal Disease (ESRD), that are in Regular Dialysis Treatment (RDT). This anaemia is usually due to the existence of two factors: A decrease in the erythropoiesis of the bone marrow and an increase in peripheral haemolysis and, consequently, a decrease in the life span of the red cells. PMID- 10723310 TI - Erythropoietin and its use in renal failure. AB - Causative treatment of anaemia associated with renal failure with human recombinant erythropoietin (rHuEPO) represents undoubtedly one of the most exciting benefits in the complex therapeutic care of patients on maintenance dialysis. Ten years have passed since the first clinical experience with rHuEPO. At present, the number of patients on rHuEPO therapy has increased to more than 300,000 worldwide. All of us being involved in renal and dialysis care should have knowledge on how to deal with this drug, what its benefits are as well as its potential untoward effects and limits. PMID- 10723311 TI - Psychological needs of adult patients following renal transplantation and implications for care. AB - Previous work undertaken by the author has detailed factors that are known to contribute to psychological reactions experienced post renal transplantation by the recipient (Dowsett, 1991). Such information has thus prompted a more detailed investigation into identifying the emotional and psychological needs of adult renal transplant recipients and to establish whether the care received from health professionals and significant others is meeting the needs of these patients following renal transplantation. PMID- 10723312 TI - Public opinion and renal transplantation: results of a sample poll. AB - Organ transplantation is one of the most urgent and deeply felt problems in this day and age. It is a very delicate topic with its ethical, scientific, legal and political implications. Transplant surgery can be defined as a sure and irreplaceable opportunity capable of solving situations of life-threatening seriousness which could not otherwise be so effectively treated. PMID- 10723313 TI - The difficulties for transplant co-ordinators in finding cadaveric grafts. AB - The increase in patients who happen to be candidates for solid organ transplantation without the proportional increase in organ donation, is a phenomenon that is observed in every centre although at different rates (1). In Greece this fact tends to take dramatic dimensions in spite of the effort that is undertaken for reducing its intensity. PMID- 10723314 TI - Organ donation, information and the mass media. AB - Many transplantation specialists have criticised negative broadcasts from the mass media and their effect on public opinion as being one of the main causes for the degraded image of transplantation and the increase in refusals they generate, both among citizens and even among health workers not specifically involved in transplantation. PMID- 10723315 TI - Public attitudes towards organ transplantation in the United Kingdom. AB - Transplant activity is being increasingly constrained by the shortage of organs available. It has to be recognised that relatives giving their consent is the key to donation actually taking place. In the 1990's transplantation is the treatment of choice for end stage renal disease. Transplant activity, however, is being increasingly constrained by the shortage of organs available. PMID- 10723316 TI - The role of education in developing specialist practice. AB - It really is a great pleasure for me to write this paper. Before EDTNA was born, when it was a only a glimmer in its Mother's eye, I fell in love with renal nursing. And it was through renal nursing that I became involved with the Royal College of Nursing of the United Kingdom, which has played such an important role in advancing the art and science of nursing, and in encouraging the development of clinical nursing expertise, and the role of specialist practice. PMID- 10723317 TI - Bridging the gap, spreading the load and mixing the skills: a training for junior staff nurses. AB - This inner London renal unit, like others, was facing an increase in patient numbers together with a shortage of renal trained staff nurses. For some time the patient population had been shifting towards a predominance of elderly dependent, although stable, clientele with various physical, psychological and social needs. PMID- 10723318 TI - Individualisation of active participation of dialysis patients: are patients satisfied? AB - In the Netherlands there are 52 dialysis units, where approximately 3,600 patients with end stage renal disease are being treated with renal replacement therapy. The dialysis department in Dordrecht is a medium to large-sized department, where about 85 patients have their renal replacement therapy. PMID- 10723319 TI - Nurse, haemodialysis, patient: is our caring based within a helping relationship? AB - Due to our experience in haemodialysis (HD) we became interested in evaluating the quality of care given during HD treatment. In May 1994, we conducted a cross sectional survey comparing the assessment of the total interaction between nurses and haemodialysis patients in a Lisbon dialysis facility. Our purpose was to identify nurses' and patients' opinion on the total interaction established during the dialysis treatment among both groups. PMID- 10723320 TI - A nursing approach: towards the integral health care of renal patients. AB - In February 1993, after several multicentre meetings held together with nephrologists, the decision was taken to create the Peritoneal Dialysis Nursing Group for the Central Area; having as its primary purpose the provision of integrated medical care to renal patients. PMID- 10723321 TI - Nursing models: a useful strategy for nephrology nursing practice? AB - The use and misuse of nursing models has been a subject which has provoked intense debate amongst nurses both in clinical areas as well as within the realms of nursing academia. The impetus for such analysis has been increased by the many changes that are evolving in modern nursing. The Oxford Renal Unit, along with many of the UK units, has begun to question available nursing models, and whether they actually represent the reality of renal patient experience. PMID- 10723322 TI - Peritoneal dialysis: year 2010. PMID- 10723323 TI - Patient and technique survival after treatment shifts between CAPD and haemodialysis in a single centre. AB - In the last decade, there has been a renewed interest in peritoneal dialysis and this modality has been proposed as a possible alternative to haemodialysis in the care of patients with end-stage renal disease. Attempts have been made to compare various aspects of these two modes of dialysis (4,6). Clinical trials have been performed particularly comparing CAPD with haemodialysis in the treatment of certain subgroups of patients, such as children or patients with diabetic nephropathy (1,5). PMID- 10723324 TI - Hydro-therapy in CAPD training. AB - Exercise has been identified as being one of the five most important principles in renal rehabilitation (1). It is well recognised that physical exercise enhances the patients' ability to carry out normal daily activities such as walking, climbing stairs, pursuing gardening or other hobbies and holding employment (1). PMID- 10723325 TI - Functional status of CAPD patients and their mood state, dialysis dose, comorbidity and quality of life. AB - Whilst Continuous Ambulatory Peritoneal Dialysis (CAPD) has sustained the lives of thousands of patients with End Stage Renal Disease (ESRD), renal team members are increasingly confronted with concerns about quality of life. Uraemic symptoms in patients with ESRD are reported to be a source of functional and psychosocial problems, thus diminishing quality of life (1). PMID- 10723326 TI - Quality of life: but in whose judgement? AB - Today, many renal units scientifically measure the clinical quality of care, monitor morbidity and mortality and provide care to patients based upon clinical and social norms for this client group. Implicit within these measurements is the assumption that the absence of clinical complications of End Stage Renal Failure (ESRF) will directly result in an increased level of satisfaction and quality of life for patients (1). PMID- 10723327 TI - Looking into the factors affecting renal patients' quality of life. AB - Quality of life, according to Horquist is "the extent to which one's needs are satisfied, in the context of physical, psychological, social and environmental conditions" (I). Self-esteem is the basic element of a good quality of life. Health-related quality of life consists of a number of components; including family relationships, friendships, finances, physical and psychological status, adjustment to therapy and feeling of security during the treatment. PMID- 10723328 TI - Continuous quality improvement (CQI) in rhu-Epo management: the outcome. AB - The establishment of an adequate recombinant erythropoietin (epo) treatment for haemodialysis patients has challenged nephrology workers for the last 5 years. Starting with epo, it was quickly clear that its management was complex and a lot of conditions were involved in its effective application. PMID- 10723329 TI - Patient categories: a tool for quality management. AB - Throughout recent years the daily work in haemodialysis centres has changed in Germany as well as in other European countries. The reason is that the percentage of older and multi-morbidity patients on chronic renal replacement therapy has progressively increased. PMID- 10723330 TI - Computer-assisted production of dietary advice: a useful tool for the future? AB - Adequate nutrition is vital for the health and well-being of all patients. Of all aspects of treatment the dietary guidelines are often the most difficult for the patient to accept (1,2). Compliance may be poor, particularly if the dietary prescription differs markedly from the patient's diet of choice (3-6). Reports of non-compliance range from 20-78%, depending on the criteria used. Equally as worrying are the patients whose nutritional intake is compromised by over compliance (7). PMID- 10723331 TI - Trauma counselling: what is it and does it exist within the caring professions? PMID- 10723332 TI - Psychosocial needs in renal failure: development of a renal support team. AB - The purpose of this paper is to illustrate how social, emotional and educational support is provided by the Renal Support Team at Dulwich Hospital to clients approaching or already receiving renal replacement therapy. We aim to illustrate how psychosocial needs were met before the creation of the team, how support has developed since this time and how clients are benefiting from this. We will conclude by discussing some future developments within the team. PMID- 10723333 TI - Haemodialysis patients' knowledge and beliefs about medication. AB - Patient adherence to treatment continues to concern health professionals. Non adherence is costly to: a) the patient in terms of health, psychological well being and quality of life, b) the health care providers in terms of individual professionalism, job satisfaction and the provision of optimum care packages, and c) the managers in terms of finance and service planning. Several factors are thought to be influential in treatment adherence. PMID- 10723334 TI - Occupational therapy in the dialysis unit. AB - Occupational therapy is defined by the American Occupational Therapy Association as: the art and science of directing man's participation in selected tasks, to restore, reinforce and enhance performance, facilitate learning of those skills and functions essential for adaptation and productivity, diminish or correct pathology and maintain health. PMID- 10723335 TI - A patient's guide to kidney failure. AB - Imagine how bewildered you would feel if you had just been told by a doctor or a nurse that your kidneys had failed and that you would shortly require dialysis. This news is broken to 80 per million of the population or approximately 4000 people in the United Kingdom each year, and must come as a dreadful blow to each and every patient, their family, carers and friends. PMID- 10723336 TI - Introduction to the research forum. PMID- 10723337 TI - Encouraging environmental awareness in renal units. AB - Humans have inhabited Earth for centuries, but unfortunately during the past two centuries we have caused untold damage and destruction to our planet, through pollution and waste of natural resources. Baxter Healthcare Ltd. is committed to a programme of environmental protection. The company adopted an Environmental policy in 1990 which applied to all Baxter operations world-wide. This policy stated that each area of the company would develop and implement its own environmental management programme. PMID- 10723338 TI - Automated on-line bicarbonate concentrate production: microbiological, chemical and economic benefits. AB - We all know that "bicarbonate dialysate" became the world wide used fluid for use in haemodialysis and related techniques. To prepare bicarbonate dialysate we actually have to use two separate concentrates: one containing sodium bicarbonate, solely or in combination with other electrolytes other than calcium, and a second concentrate containing mostly all the other electrolytes (sodium, magnesium, potassium, chloride and/or some acetate). PMID- 10723340 TI - Water treatment for haemodialysis: not as safe as anticipated. AB - "It is not exaggerated to state that inadequate water treatment is one of the gravest risks posed to the health of the patient on dialysis". PR Keshaviah. Water purification for haemodialysis relies on a combination of filtration, carbon adsorption, ion exchange and Reverse Osmosis (RO). This procedure is commonly considered safe and adequate (1). PMID- 10723339 TI - Clinical use of continuous blood volume monitoring. AB - Our department has been using continuous blood volume monitoring (Critline instrument, In-line Diagnostics, Riverdale, USA) for over two years now. First during research on sodium concentration control in haemodialysis (HD), later in the project on assessment and control of hydration in HD patients. PMID- 10723341 TI - Haemodialysis adequacy in children: urea kinetic modelling. AB - The delivered dialysis dose is quantified by KT/V, the normalized whole body urea clearance. Calculations of KT/V are most often performed in single pool urea kinetic modelling, from the post and predialysis serum values. Although the exact levels of KT/V required is a matter of some debate, a minimum KT/V level of 1.2 1.4 is now thought to be necessary. PMID- 10723342 TI - Nephrocamp: the holistic approach to care. AB - The care process for a child suffering from renal impairment and its family is very complex and can cause an enormous strain However the technical aspects of the therapy is often dominant leaving the psychological aspects aside. This paper describes a possibility of care intervention to address the situation in a holistic way. PMID- 10723344 TI - Haemodialysis in infants. AB - Chronic ambulatory peritoneal dialysis (CAPD) is the treatment most often used in neonates and infants with chronic kidney failure. However, in some cases it is impossible to use CAPD, e.g. due to major abdominal surgery. Then chronic intermittent haemodialysis is the only remaining treatment, until kidney transplantation can be carried out. PMID- 10723345 TI - New approach to vascular access revascularisation: the Hydrolyser Thrombectomy Catheter. AB - The appearance of thrombosis in the vascular access (VA) for haemodialysis (HD) is one of the most important complications of HD, with a great impact on the survival of definitive VA-both autologous internal arteriovenous fistulae (IAVFs) and prostheses (Gore-tex). PMID- 10723343 TI - Chronic single needle predilutional haemofiltration in a pre-school child. AB - Peritoneal dialysis is the preferred dialysis mode for children with end stage renal disease (1). It avoids problems with vascular access and enables near normal life style. Haemodialysis is the only mode of treatment for a child waiting for renal transplantation when peritoneal dialysis is not possible. Haemofiltration, as a mode of renal replacement therapy, was introduced in 1967 by Lee Henderson (2). PMID- 10723346 TI - Self-expandable endovascular stent for treatment of venous stenoses. AB - The increase in survival among patients undergoing haemodialysis, the non limitation of patient age regarding the start of renal substitution treatment, and the high incidence of vascular problems--particularly in diabetics--have caused the positioning of percutaneous catheters in central vessels to become a common practice in haemodialysis. PMID- 10723347 TI - Integrated and specialised care of arteriovenous fistulae improves quality of life. AB - The purpose of this paper is to enable you to become acquainted with integrated care of vascular access as a substantial part of the overall treatment of patients who depend on renal replacement therapy. PMID- 10723348 TI - Quality in practice: setting a standard for the insertion of fistula needles. AB - Nursing research, involving haemodialysis patients completing mood diaries, conducted at St. James' Hospital in Leeds, has indicated that consecutive problematic haemodialysis sessions can significantly affect a patient psychologically (1,2). In response to these findings a nursing standard was set at the Lincoln Haemodialysis unit, a satellite unit of Leicester General Hospital Department of Nephrology, to identify pain on needle insertion. PMID- 10723349 TI - Contribution of an effective venopuncture to recirculation. AB - Patency and function of permanent vascular access appears to be one of the most important elements of an adequate haemodialysis session. Since, it has been established that adequate haemodialysis is closely related to the mortality and morbidity of these patients, we all realise the importance of the strategies involved in handling the av fistula (1). PMID- 10723350 TI - Nursing protocol for manipulation of haemodialysis catheters. AB - This study looked at the incidence of infection complications, in relation to central vein catheterisation as a provisional HD access, by means of the establishment of a nursing protocol for the handling of these catheters. Central vein catheterisation is a classical technique in Nephrology. PMID- 10723351 TI - Organ transplantation: the role of critical care nursing. PMID- 10723352 TI - Despite the pressures of busy lives, oncology nurses make time for professional involvement. PMID- 10723353 TI - Splitting the difference. PMID- 10723354 TI - An asthma policy for schools in Cornwall. PMID- 10723355 TI - Value added decisions. PMID- 10723356 TI - Enteral nutrition in critical illness: Part One. PMID- 10723357 TI - The Lifetime Service: a model for children with life-threatening illnesses and their families. PMID- 10723358 TI - Young children in A&E: a local review. AB - Between two and three million children attend accident and emergency (A&E) departments every year in the United Kingdom, making up one quarter of all A&E attendances (Bentley 1996). Despite government, professional and consumer guidance (Audit Commission 1996, Action for Sick Children 1997), the majority of children are not seen in a children's A&E or cared for by appropriately-trained staff. This study explored the pattern of attendance of 375 children aged five years and under who attended the A&E department in a district general hospital. Issues of service provision and quality of care were also explored using secondary data from A&E records. Key findings were that one third of the children were triaged with non-urgent illness, suggesting a need for improved access to GP services and that a number of children were referred by GPs and admitted via A&E rather than directly to the paediatric ward. Although waiting times were generally short, many children did not appear to receive analgesia, suggesting the need for improved pain assessment and management. PMID- 10723359 TI - Play and culture. PMID- 10723361 TI - How to ... prepare an abstract. PMID- 10723360 TI - The ethics of caring: reflection on the death of a child. PMID- 10723362 TI - Principles and practice of child protection. AB - This article aims to update the registered nurse's knowledge and understanding of the signs of child abuse, the legislation that exists to protect children and the action available to healthcare staff in cases where abuse of a child is suspected. PMID- 10723363 TI - Widening ripples from Bristol. PMID- 10723364 TI - Preparing an information leaflet. PMID- 10723365 TI - Tomorrow's nurse for tomorrow's child: a student perspective. PMID- 10723367 TI - The anaesthetic nurse specialist in the pre-admission clinic. PMID- 10723366 TI - Catherine Wood: children's nursing pioneer. PMID- 10723368 TI - Promoting children's rights: the role of the children's nurse. PMID- 10723370 TI - An evaluation of five years' work in Romania. PMID- 10723371 TI - Nurse-led care for children with asthma. PMID- 10723369 TI - Enteral nutrition in critical illness: Part Two. PMID- 10723372 TI - Consent in practice. PMID- 10723373 TI - Positive moves. PMID- 10723374 TI - Still much to be done about pain. PMID- 10723375 TI - Neonatal sensitisation to latex: a hypothesis. AB - The exposure of babies to latex gloves at birth may be behind the dramatic increase in childhood allergies in the past 40 years. Jennifer Worth explores this theory and calls for a ban on the use of pre-powdered latex gloves in maternity units. PMID- 10723376 TI - Comparison of costing tools in paediatric intensive care. AB - The cost of service provision is an issue of concern to all professionals working in health care. Department of Health reports have highlighted the need for more effective costing mechanisms for provision of paediatric intensive care (PIC). This study aimed to determine whether nurse-patient dependency scoring would be as effective as the Therapeutic Intervention Scoring System (TISS) at providing a differential costing tool for PIC. Data were collected on 251 patient admissions over 1,741 nursing shifts and analysis of variance undertaken. Findings suggest that a modified nurse-patient dependency score would be as good as TISS as a potential costing tool in PICU. PMID- 10723377 TI - Are Newcastle urine collection pads suitable as a means of collecting specimens from infants? AB - The collection of uncontaminated urine specimens from infants is not straightforward. This quasi-experimental study was designed to investigate whether 'Newcastle urine collection pads' are suitable for this purpose. A minimum of two urine specimens were collected from 50 children aged one day to five years, one using a pad and the other by using a sterile adhesive bag. Since pad specimens successfully demonstrated the presence of urinary tract infection and a lower contamination rate than bag specimens; it is concluded that they are suitable. Pad specimens contained fewer cells than bag specimens, which must be acknowledged by clinicians. PMID- 10723378 TI - Improving staffing and quality: a nursing support team. PMID- 10723379 TI - Children in mind--new CAMHS audit report. PMID- 10723380 TI - Holistic pain management. PMID- 10723381 TI - Epilepsy education in schools. AB - The objective of this study was to assess the effect of an epilepsy education initiative on the management of epilepsy in Southampton schools. A self completion questionnaire survey was undertaken of staff in 19 Southampton primary schools and ten secondary schools. Changes in knowledge and management of seizures in children with epilepsy were measured following an education session and a recommended implementation of an individual seizure protocol for children with epilepsy. There were significant changes in some areas of seizure management and knowledge. Only a small number (three) of primary schools had a protocol in place. Two primary and four secondary schools were in the process of putting the protocol in place. An improvement in knowledge and seizure management has been demonstrated. Further work is needed to implement an individual seizure protocol for all children with epilepsy. PMID- 10723382 TI - Advancing practice roles: reflections from the US. PMID- 10723383 TI - Mental health assessment for children and young people. AB - To provide appropriate help to children with mental health problems, a thorough assessment of the nature and extent of the disorder is essential. This article identifies some common disorders encountered among children and describes how to tailor assessments to particular problems. PMID- 10723384 TI - Time to be proud of children's nursing. PMID- 10723385 TI - Global millennium targets: UNICEF Child-Friendly Hospital Initiative. AB - Twelve standards for children's hospitals have been drafted by the UK Committee for UNICEF and are to be piloted in the UK. Here is a summary from UNICEF of the targets that will help to change attitudes to child care. PMID- 10723386 TI - Child branch: criteria for selection. PMID- 10723387 TI - Nurses' attitudes to disability. PMID- 10723388 TI - Piloting a clinical educator role. PMID- 10723389 TI - Mouth care in PICU. PMID- 10723390 TI - 'A child's eye view': the development and evaluation of a teaching video. PMID- 10723391 TI - Three electives. AB - Electives are an important part of students' education and often provide long lasting memories and friendships, as well as knowledge. The following three accounts are of students who spent their electives in Sweden, Finland and Romania. They report on their experiences abroad, what they learnt from them and draw comparisons with the healthcare system in the United Kingdom. PMID- 10723392 TI - Infant nutrition: Part One. PMID- 10723393 TI - Women behaving badly. PMID- 10723394 TI - Caregiver support for women during childbirth. PMID- 10723395 TI - The Wessex Maternity Centre. A midwifery service outside the NHS. PMID- 10723396 TI - Asian women's maternity language course. PMID- 10723397 TI - Midwife teachers and clinical practice. Part 3: Rebuilding the relationship. PMID- 10723398 TI - Is the partogram a help or a hindrance? An exploratory study of midwives' views. PMID- 10723399 TI - Biochemical and blood tests in midwifery practice (1). Pre-eclampsia. PMID- 10723400 TI - Getting what you want from antenatal education.... PMID- 10723401 TI - Pregnant women and testing for HIV. PMID- 10723402 TI - Pleased to meet you. Nicky Leap. PMID- 10723403 TI - Which course? Leading antenatal classes. PMID- 10723404 TI - Knee deep at a home birth. PMID- 10723405 TI - The force of law. PMID- 10723406 TI - Support for breastfeeding mothers. PMID- 10723407 TI - 'Now just pop up here, dear...' Revisiting the art of antenatal abdominal palpation. PMID- 10723408 TI - Intrauterine pollution and human milk pollution. PMID- 10723409 TI - Confidential Enquiry into Stillbirths and Deaths in Infancy (CESDI). Highlights of the 6th annual report. PMID- 10723410 TI - Baby Friendly. Where has it got to? PMID- 10723411 TI - Skin-to-skin contact immediately after birth. PMID- 10723412 TI - Informing women about pre-eclampsia. PMID- 10723413 TI - HIV screening and pregnancy. Testing is accurate. PMID- 10723414 TI - More doubt required. PMID- 10723415 TI - Pleased to meet you. Patricia Purton. PMID- 10723416 TI - Real women. PMID- 10723417 TI - Desperately seeking continuity. PMID- 10723418 TI - [The nurse and the accessibility of medical records]. PMID- 10723420 TI - [School nurses: extended duties but insufficient means]. PMID- 10723419 TI - [Postcoital contraception]. PMID- 10723421 TI - [School nurses: a profession under constant change]. PMID- 10723422 TI - [Evaluation of the nutritional status of adults]. PMID- 10723423 TI - [Different methods of enteral nutrition]. PMID- 10723424 TI - [Feeding solutions]. PMID- 10723425 TI - [Indications for enteral nutrition]. PMID- 10723426 TI - [Surveillance of enteral nutrition]. PMID- 10723427 TI - [Complications of enteral nutrition]. PMID- 10723429 TI - [An individual program for the management of premature infants]. PMID- 10723428 TI - [The value of nursing diagnosis in the management of alcohol dependence]. PMID- 10723430 TI - [Artificial blood in the future]. PMID- 10723431 TI - [Education of hypertensive patients assisted by computers]. PMID- 10723433 TI - [Therapeutic observation and the role of the nurse]. PMID- 10723432 TI - [Nutritional evaluation and re-equilibration in oncology]. PMID- 10723434 TI - ["Europa Donna", association to fight breast cancer]. PMID- 10723435 TI - [Postcoital contraception: two new medications]. PMID- 10723436 TI - [Kinesitherapy and gerontology]. PMID- 10723437 TI - [Treatment refusal and energy mobilization]. PMID- 10723438 TI - [Psychomotor performance in institutionalized aged persons]. PMID- 10723439 TI - [Muscular atrophy, nutrition and aging]. PMID- 10723440 TI - [Low vision, equilibrium and accidental falls]. PMID- 10723441 TI - [Changes in physical mobility]. PMID- 10723442 TI - [Neuro-psychomotor rehabilitation]. PMID- 10723444 TI - [Tuberculosis in the aged]. PMID- 10723443 TI - [On the menu, a whole variety of food: fresh, frozen, preserved...]. PMID- 10723445 TI - [Foot care in patients under long-term care]. PMID- 10723446 TI - [The humanizing function of clothing: value of self concept]. PMID- 10723447 TI - [Oxygen therapy in the home]. PMID- 10723448 TI - [Installation of arm chairs for rest]. PMID- 10723449 TI - [Legal protection of the aged]. PMID- 10723450 TI - [Intergenerational cinema festival at the Marine Hospital]. PMID- 10723451 TI - [Rate fixing reform soon to be published]. PMID- 10723453 TI - [Theme issue pneumology]. PMID- 10723452 TI - Pulmonary endarterectomy. PMID- 10723454 TI - [Interventional catheter treatment of bypass graft stenosis: comparison of intracoronary stent implantation and balloon angioplasty]. AB - BACKGROUND AND OBJECTIVE: Balloon angioplasty of a stenosed bypass graft has a much higher risk of recurrent stenosis than dilatation of a stenosed native coronary artery. Intracoronary stent implantation has established itself as the better treatment of native coronary artery stenosis than conventional balloon angioplasty. However, there is still uncertainty whether intracoronary stent implantation in stenosed bypass vessels gives better long-term results than conventional balloon angioplasty. PATIENTS AND METHODS: Results were retrospectively analyzed of unrandomized 224 primarily successful interventions- 122 balloon dilatations and 102 stent implantations--performed between January 1996 and June 1998 on stenosed coronary bypass grafts, re-examined by coronary angiography an average of 6 months later. All but 11 patients were on combined aspirin and ticlopidine antiplatelet aggregation treatment. RESULTS: There was a significantly lower 6-month restenosis rate (30.4%) after stent implantation than after balloon dilatation (51.6%). The re-intervention rate was also significantly lower after stent implantation. CONCLUSION: These data suggest that stent implantation of stenosed coronary bypass grafts under cover of platelet aggregation inhibition with aspirin and ticlopidine provides a lower restenosis and thus higher revascularization rate than conventional balloon dilatation. PMID- 10723455 TI - [Hepatitis after blood transfusion in inpatients. Post-transfusion hepatitis, or infection of other etiology?]. AB - BACKGROUND AND OBJECTIVE: Hepatitis after a transfusion of blood products is often uncritically called post-transfusion hepatitis (PTH). It was the aim of this study to ascertain in how many reported cases of suspected PTH a causal relationship between transfusion and the infection in the recipient can be proven. PATIENTS AND METHODS: Full investigations (look-back method) were made of 23 cases of PTH reported in the 10-year period of 1987-1997 (11 cases of PTH B and 12 of PTH non-A-non-B or C). The recipients had been given a mean of 7.7 blood components (range 1-57). The clinical diagnosis had been made a mean of 5.2 (range 1-27) months after transfusion. RESULTS: One case each of hepatitis B and C had been caused by transfusion of blood products. In 5 of 12 suspected cases of PTH non-A-non-B hepatitis C, PTH could not be definitively excluded (despite absent seroconversion for anti-hepatitis C virus [HCV]), because HCV-RNA findings were not available. In 16 of 23 investigated cases serological and molecular biological tests firmly excluded PTH B or C. CONCLUSIONS: The small number of confirmed cases of PTH indicates that hepatitis that has occurred post transfusion is frequently due to a cause not related to the transfusion. The number of transfusion-associated cases will be further reduced with the introduction (standard since 1.4.1999) of VCH-RNA screening of donors. This will raise the importance of hepatitis cases not associated with transfusion. PMID- 10723456 TI - [Macro AST]. PMID- 10723457 TI - [Corticosteroids and chronic obstructive lung disease (COPD)]. PMID- 10723458 TI - [Cortisone therapy in chronic obstructive lung disease]. PMID- 10723459 TI - [Established new drugs in the chemotherapy of non-small-cell lung carcinoma (NSCLC)]. PMID- 10723460 TI - [Therapy of fungal infections of the lung]. PMID- 10723461 TI - [Diagnosis of acute pulmonary artery embolism]. PMID- 10723462 TI - [Interventional bronchoscopy]. PMID- 10723463 TI - [New aspects in the epidemiology of ambulatory-acquired pneumonia]. PMID- 10723464 TI - [Therapeutic options in the management of stable chronic obstructive lung disease (COPD)]. PMID- 10723465 TI - [Care of the mind in scientific medicine? First writings on psychoanalysis at the beginning of the 20th century]. PMID- 10723466 TI - [First HIV triple therapy with one tablet]. PMID- 10723467 TI - [Detection of asymptomatic venous thrombosis after lower limb prosthetic surgery. Retrospective evaluation of a systematic approach using Doppler ultrasonography: 400 cases]. AB - OBJECTIVES: The purpose of this study was to evaluate a pragmatic approach using duplex ultrasonography (US) for detecting deep vein thrombosis (DVT) after total hip (THA) and total knee (TKA) replacement. METHODS: Venous B-mode and color duplex US examination of both legs including a systematic evaluation of calf veins was performed twice during hospital stay (Between day 1 and day 4 for the first exam and between day 7 and day 11 for the second) in 400 consecutive patients. RESULTS: Deep vein thrombosis was diagnosed in 53 patients (13.5%) including 7 patients with proximal DVT. Thrombosis was asymptomatic in 46 patients (85%), and was bilateral or concerned the non-operated leg in 8 patients (14.5%). No clinical pulmonary embolism (PE) occurred during hospital stay (mean hospital stay: 12.3). Prior phlebitis and age over 70 were identified as a statistically significant risk-factor for post-operative DVT (p = 0.001 and p < 0.01 respectively) concerning the whole series and the THA series (p < 0.02 and p < 0.04 respectively). No statistically significant risk factor was founded for the TKA series (p < 0.2 and p < 0.2 respectively). All patients were seen at three months. Four patients (1.16%) developed DVT between hospital discharge and the 3-month follow-up visit. One patient with coronary disease died suddenly on post-operative day 24, without clinical signs or symptoms of PE or DVT. CONCLUSION: Venous US performed twice after total hip replacement detected asymptomatic DVT in 85% of patients. This approach might explain the absence of PE in our series and thus justify systematic ultrasonographic evaluation of lower limb veins after prosthetic replacement. PMID- 10723468 TI - [Prescriptions for antidepressants in a population of psychiatrists and non psychiatrists]. AB - OBJECTIVE: Much proof has been accumulated over the last decade demonstrating that depression is a major public health issue. Use of psychotropics and more precisely antidepressants is considered to be excessive. It is however paradoxical that prescribing antidepressants has become commonplace. The aim of this study was to better assess the process of prescribing antidepressants in the hospital setting. METHODS: An epidemiological study was carried out to examine prescribing practices used by psychiatrists and non psychiatrists working in the Rennes University Hospital. The psychiatrist population was used as the reference population for univariate and multivariate analysis designed to ascertain differences concerning prescription practices. RESULTS: Duration of the clinical examination (shortest for non-psychiatric physicians, p = 0.0001), use of a diagnostic scale (more frequently for psychiatrists, p = 0.0008), reasons for choosing an antidepressant (pharmacological considerations more frequent among psychiatrists, p = 0.0009), and co-therapies (neuroleptic association more frequent among psychiatrists, p = 0.0001) were found to be different between the two prescribing populations. CONCLUSION: All patients with signs of depression are not necessarily given optimal care. Errors in assessing antidepressants is probably a common problem. PMID- 10723469 TI - [Disseminated Penicillium marneffei infection suggesting visceral leishmaniasis in an HIV infected patient]. AB - BACKGROUND: Penicilliosis is a fungal infection caused by Penicillium marneffei. In Southeast Asia, this infection is common in HIV-infected patients but few cases have been reported in Europe. CASE REPORT: The patient lived in southern France. He had HIV infection and was immunocompromised. The clinical signs suggested visceral leishmaniasis. Iterative travels to Southeast Asia were reported leading to the diagnosis of P. marneffei infection. DISCUSSION: In southern France, imported penicilliosis marneffei may be misdiagnosed as visceral leishmaniasis. PMID- 10723470 TI - [Horton's disease in the course of chronic hepatitis C]. AB - BACKGROUND: Vascularitis is a well-known extrahepatic manifestation of chronic hepatitis C. Mixed cryoglobulinemia is the most common form. To our knowledge, the present case is the first report associating chronic hepatitis C and temporal arteritis. CASE REPORT: A 56-year-old man with chronic hepatitis C in the precirrhogenic phase presented with fever and weight loss. The patient complained of pain of the scapular and pelvic girdles and headache and physical examination revealed claudication of the jaw and abolition of the upper limb pulses. Biopsy of the temporal artery confirmed the diagnosis of Horton's disease. The patient also had bilateral stenosis of the sub-clavian arteries. DISCUSSION: This observation of Horton's disease involving large vessels in a patient with chronic hepatitis C suggests that an infectious factor might trigger vascularitis. PMID- 10723471 TI - [Breast cancer in two male dizygotic twins]. PMID- 10723472 TI - [Familial sarcoidosis]. PMID- 10723473 TI - [Do patients understand their usual treatment? Inquiry upon admission to a medical service]. PMID- 10723474 TI - [Polyp screening: is virtual colonoscopy a valid alternative to conventional colonoscopy?]. PMID- 10723475 TI - [Guidelines for the use of intravenous thrombolytic therapy cerebrovascular ischemic accident. French Society for Neurovascular Disease]. PMID- 10723476 TI - [With respect to the guidelines for thrombolytic therapy for cerebral ischemia. Interviews with President D. Leys and Dr. C. Masson. Interview by C. Mammand]. PMID- 10723477 TI - [Good use of medications in outpatient clinics. Seminar organized by the AMC]. PMID- 10723478 TI - [Skin and osteoarticular bacterial infections in the diabetic foot. The need for multidisciplinary management]. PMID- 10723480 TI - [Skin and osteoarticular infections of the diabetic foot. Role of infection]. AB - A MAJOR PROBLEM: Two-thirds of all amputations involve infection. Infection is favored by dysfunction of the antibacterial defense systems due to high blood glucose and vascular disorders. DIAGNOSIS: General signs of infection are usually not found. A careful exploration is required to rule out or confirm osteitis in order to guide surgery and plan the antibiotic regimen. A history of chronic and/or recurrent ulceration or direct signs at inspection may be suggestive of osteitis. Radiographic signs are late and nonspecific. Scintigraphy scans are difficult to interpret. Magnetic resonance imaging can be quite helpful in difficult cases. BACTERIOLOGICAL PROOF: Staphylococcus aureus and to a lesser extent streptococci account for almost all of the superficial infections in the diabetic foot. In case of deep ulceration, it is important to obtain deep specimens at surgical cleansing as more superficial samples are easily contaminated. Nevertheless, if Staphylococcus aureus is isolated from pus coming from a deep zone fistulizing to the skin, it is likely the causal agent since 80% of all bone infections involve S. aureus. Other germs besides staphylococci and streptococci include enterobacteria (40%), enterococci (26%) and pseudomonas (7%). Several germs are involved in about 70% of cases with a probable synergetic effect between the different bacterial colonies within the infected tissues. PMID- 10723479 TI - [Skin and osteoarticular bacterial infections of the diabetic foot. Ulcers of the diabetic foot: epidemiology and physiopathology]. AB - EPIDEMIOLOGY: There are more than 2 million diabetics in France. Fifteen percent have suffered at one time or another from a foot ulcer. This condition accounts for 20% of all admissions of diabetic patients and for 50% of corresponding hospitalization stays. Fifteen to 25% of diabetic foot ulcers lead to an amputation, the patients being in the 45-65 year age range. Fifty percent of the amputated patients will have a contralateral amputation within the next 5 years. PATHOPHYSIOLOGY: Diabetic foot ulcers result from damage caused by diabetic neuropathy and micro- or macroangiopathy. Ulceration is favored by usually minimal trauma and secondary infection. The neuropathy causes deformations and sensorial disorders. Repair is hindered by the often precarious vascular supply. Reduced antibacterial defense related to high serum glucose levels and impaired diapedesis favor superinfection. MANAGEMENT: Careful physical examination and appropriate explorations are required for proper care giving the patient the best chances for cure. PMID- 10723481 TI - [Skin and osteoarticular bacterial infections of the diabetic foot. Treatment]. AB - MULTIDISCIPLINARY CARE: A multidisciplinary approach is essential. General measures include immobilization of the focus, controlling blood glucose, anticoagulation, and anti-tetanus vaccination. Topical application of growth factors is currently under evaluation. ANTIBIOTIC THERAPY: The antibiotics chosen should diffuse well into bone tissue. Combinations with synergetic or additive effects against Staphylococcus aureus are best. Treatment duration depends on the depth of the ulceration. Two weeks is generally advised for superficial ulcers. For deep ulcers, treatment duration depends on the presence or not of osteitis and the quality of surgical debridement. In case of osteitis, after amputation with a healthy margin, antibiotics can generally be discontinued 2 weeks after surgery. Six weeks are required if the amputation margins do not lie in healthy zones. Finally, if no surgery is attempted, the antibiotic regimen should be continued for 3 months, or even longer, with a risk of failure greater than 50%. The best criterion for successful treatment is the absence of late recurrence. SURGERY: Surgery is an indispensable element in the overall treatment of deep infections and/or osteitis. The operation should be performed as early as possible to improve prognosis. Well-conducted early surgical debridement can prevent the infection from spreading and avoid the need for much more mutilating "salvage" procedures. Vascular surgery can help maintain sufficient blood supply for wound healing and antibacterial defense. Plastic surgery can be very helpful. PREVENTION: A certain number of simple measures help reduce the risk of diabetic foot ulcers. However, many patients, and practitioners, are insufficiently aware of their effectiveness. Prevention and treatment can best be accomplished by a multidisciplinary approach calling upon the endocrinologist and the vascular and orthopedic surgery teams. A carefully planned rehabilitation program using adapted soles, orthesis, orthopedic shoes or prostheses as needed can considerably reduce the frequency of recurrence. The risk of recurrence in a patient wearing adapted footwear is only 26% at 5 years compared with 83% in other cases. PMID- 10723482 TI - [Interventional magnetic resonance imaging--non-invasive imaging for interventions]. AB - As a prerequisite for MR-guidance of interventional procedures, instruments have to be well depicted in the MR image without obscuring or distorting the underlying anatomy. For non-vascular interventions the imaging speed has to be in the range of seconds while control of vascular interventions requires real time imaging speed. The imaging contrast has to be maintained as well as a high spatial resolution. Furthermore, sufficient patient access has to be provided by the MR scanner. Neither an ideal magnet nor the optimal single sequence are available to fulfill the above-mentioned criteria. The type of sequence--gradient echo versus spin echo--together with changing of the echo time and phase encoding direction will ensure an appropriate size of the artifact and thereby of the appearance of the instrument in the MR image. The feasibility of non-vascular MR guided interventions has been proved at field strengths ranging from 0.064 T to 1.5 T. Bone biopsies, soft tissue biopsies, drainages, and control of interstitial thermo- and cryotherapy have been reported. For vascular interventions, different real time MR strategies are currently under investigation. The development of dedicated catheters and guide wires has enabled MR-guided dilatations, stenting, placement of vena cava filters, and TIPS procedures. Considering the fast progress being made in this field, there can be no question that interventional MRI will become a well-accepted clinical tool offering potential advantages such as excellent soft tissue contrast, multiplanar imaging, flow measurements, high resolution imaging of vessel walls, and lack of ionizing radiation. PMID- 10723483 TI - [Diagnosis of tracheal instability: spiral CT in inspiratory and expiratory and respiratory cine CT]. AB - PURPOSE: In tracheo- and bronchomalacia, localization and determination of collapse is necessary for planning a surgical procedure. We compared inspiratory and spiral CT, cine CT, and bronchoscopy and evaluated the relevance of each method. METHODS: Seventeen patients with suspected or verified tracheal stenosis or collapse underwent paired inspiratory and expiratory spiral CT and cine CT during continuous respiration (temporal increment 100 ms). The tracheal cross sectional area was calculated and compared. RESULTS: In addition to bronchoscopy, further information concerning localization, extent, collapse, stability of the tracheal wall, distal portions of the stenosis, and extraluminal compressions was obtained. A significantly higher degree of tracheal collapse was seen using cine CT compared to paired spiral CT (p < 0.002). The findings changed the further surgical procedure in 3/17 patients. Further distal stenoses were excluded and bronchoscopy was verified in another 10/17. CONCLUSION: Paired inspiratory and expiratory spiral CT localizes tracheal stenoses and demonstrates clinically relevant extraluminal compression. Improved evaluation of expiratory collapse and further information of localized tracheal instability is provided by cine CT. PMID- 10723484 TI - [Contrast media enhanced three dimensional MR angiography of the pulmonary arteries in patients with chronic recurrent pulmonary embolism--comparison with selective intra-arterial DSA]. AB - PURPOSE: This study compares contrast-enhanced 3D-MR angiography (MRA) of the pulmonary arteries with selective intra-arterial DSA in patients with chronic thromboembolic pulmonary hypertension. MATERIALS AND METHODS: 20 patients preoperatively underwent a contrast-enhanced 3D-MRA of the pulmonary arteries at 1.5 T using the phased-array body coil. For MRA, we used a 3D-Flash-sequence after bolus timing. 2 radiologists analyzed the acquired image material in consensus with respect to the detection of central thromboembolic material and the visualization of the pulmonary arterial tree. Finally, the MR angiograms were compared with selective DSA images using surgical findings as the definitive standard. RESULTS: MRA demonstrated central thromboembolic material, vessel cut offs and abnormal proximal-to-distal tapering in all patients. Compared to DSA, MRA depicted the pulmonary vessels up to the segmental level in all cases, it was inferior to DSA in delineation of the subsegmental arteries (sensitivity 87%, specificity 100%). The central beginning of the thromboembolic occlusions seen at MRA corresponded to the beginning of the deobliteration procedure during pulmonary thromboendarterectomy in every case. CONCLUSIONS: Contrast-enhanced 3D MRA of the pulmonary arteries enables the reliable detection of central thromboembolic material in patients with CTPEH. It also allows identification of those patients who may be treated surgically. PMID- 10723486 TI - [Contrast-enhanced MR angiography of abdominal vessels using a 1.0 T system]. AB - PURPOSE: To evaluate the efficacy of breath-hold, three-dimensional, contrast enhanced magnetic resonance angiography with a 1.0 T system for imaging the abdominal vessels in comparison to conventional arteriography (CA). METHODS: The abdominal aorta and visceral arteries were studied in 54 patients (60 examinations) on a 1.0 T scanner using an ultrafast gadolinium-enhanced gradient echo sequence with the following parameters: TR/TE = 3.8/1.4 ms, flip angel 25 degrees, matrix 198 x 256, field 380-420 mm, pixel size 1.9 x 1.48 mm2, slice thickness 1.5-2.5 mm, acquisition time 22-26 sec. Individual circulation times were determined by a test bolus before each MR angiography. Conventional arteriography was performed in 23 of the 60 cases. RESULTS: 172 vessel segments of 23 MR angiographies were compared with CA, sensitivity and specificity were 96.4% and 97.2%. Over-estimations of stenoses or occlusions (n = 4) were caused by the limited resolution of small vessel branches and one stent artifact. CONCLUSION: Contrast-enhanced MR angiography of the abdominal vessels may replace invasive digital subtraction angiography in certain cases like perioperative or peri-interventional diagnostics. Imaging of small peripheral vessels remains a problem and limits use of the method. PMID- 10723485 TI - [Ultrafast MRI of lung ventilation using hyperpolarized helium-3]. AB - OBJECTIVE: Assessment of the temporal and spatial dynamics of hyperpolarized Helium-3 (3He) distribution in the lung with ultrafast gradient-echo magnetic resonance imaging. MATERIAL AND METHODS: Coronal images of the lung were acquired using ultrafast gradient-echo pulse sequences with TR/TE = 3.3 ms/1.3 ms (slice thickness, 40 mm) and TR/TE = 2.0 ms/0.7 ms (without slice selection). A series of 80 or 160 projection images was obtained with 210 ms or 130 ms temporal resolution, respectively. Imaging was performed during several respiratory cycles after application of a single bolus of 300 mL hyperpolarized 3He. Measurements were performed in six healthy volunteers (spontaneous breathing). RESULTS: Different phases of in- and expiration could be visualized. During the course of consecutive respiratory cycles the 3He signal decreased due to dilution of 3He in residual alveolar gas and by inspired air, relaxation due to oxygen and the RF pulses, and due to Helium-3 washout. The signal of a single bolus of 3He was detected in the lung for up to four respiratory cycles. Anatomical structures were better visualized on slice selective images than on images without slice selection. CONCLUSION: Distribution of inspired 3He within the tracheobronchial tree and alveolar space and its washout can be visualized by ultrafast imaging of a single bolus of hyperpolarized 3He gas. This method may allow for regional analysis of lung function with temporal and spatial resolution superior to conventional methods. PMID- 10723487 TI - [Automated detection of spleen volume by spiral CT scans using neural networks and "fuzzy logic"]. AB - PURPOSE: To assess spleen segmentation and volumentry in spiral CT scans with and without pathological changes of splenic tissue. METHODS: The image analysis software HYBRIKON is based on region growing, self-organized neural nets, and fuzzy-anatomic rules. The neural nets were trained with spiral CT data from 10 patients, not used in the following evaluation on spiral CT scans from 19 patients. An experienced radiologist verified the results. The true positive and false positive areas were compared in terms to the areas marked by the radiologist. The results were compared with a standard thresholding method. RESULTS: The neural nets achieved a higher accuracy than the thresholding method. Correlation coefficient of the fuzzy-neural nets: 0.99 (thresholding: 0.63). Mean true positive rate: 90% (thresholding: 75%), mean false positive rate: 5% (thresholding > 100%). Pitfalls were caused by accessory spleens, extreme changes in the morphology (tumors, metastases, cysts), and parasplenic masses. CONCLUSIONS: Self-organizing neural nets combined with fuzzy rules are ready for use in the automatic detection and volumetry of the spleen in spiral CT scans. PMID- 10723488 TI - [Segmental anatomy of the liver in computed tomography: do we localize the lesion accurately?]. AB - PURPOSE: To evaluate if Couinaud's model using the planes of the major veins is an adequate tool for the presurgical localization of focal liver lesions. METHODS: Biphasic helical CT scans were performed on patients evaluated for liver resection using an increased IV bolus of contrast medium (180 ml lopamidol) and 2 mm image reconstruction increments. During the first evaluation, all liver lesions were localized in the conventional way using the planes of the 3 major hepatic veins and the portal trunks as segmental boundaries. In a second review, all lesions were attributed to the nearest peripheral portal branches. The path and the segmental attribution of the portal branches were analysed. Evaluations were performed using an interactive cine mode as well as three-dimensional reconstructions. RESULTS: 20 of 126 (16%) liver lesions had a different segmental location if the individual anatomy of the peripheral portal branch was used instead of the conventional technique. These different locations were due to the path of the portal trunks or the path of the peripheral portal branches crossing the planes of the major hepatic veins. CONCLUSION: The segmental anatomy of the liver using the planes of hepatic veins and portal trunks according to Couinaud is not an accurate tool for the presurgical localization of liver lesions in many cases. PMID- 10723490 TI - [CT angiography hemodynamically relevant to renal artery stenosis. Evaluation of AXIAL, MPR, MIP and SSD reconstruction procedures under standard investigation conditions]. AB - PURPOSE: To evaluate the various reconstruction methods of helical-CT angiography for the assessment of hemodynamically relevant renal artery stenoses in comparison to i.a. DSA. METHODS: In 76 renal arteries the reconstruction modalities AXIAL, MRP, MIP and SSD of helical-CT angiography were compared with the results of i.a. DSA for the determination of the grade and location of the stenosis. RESULTS: The highest accuracy of stenosis grading was 76% with AXIAL reconstruction. In 8% of the cases grading of the stenosis was not evaluable by the AXIAL reconstruction. In these cases, a higher sensitivity in the detection of hemodynamically relevant stenoses (> grade II, > 50%) was achieved with the reconstruction mode MPR (96%) than with MIP (92%). In 51% of the cases the reconstruction mode SSD was not suitable for any diagnosis of renal artery stenosis because of overlying calcified plaques. CONCLUSIONS: The evidence of hemodynamically relevant stenosis in helical-CT angiography in comparison to i.a. DSA succeeds most reliable by using the reconstruction modality AXIAL in combination with MPR. The MIP reconstruction provides information about the anatomy of the renal arteries within one image. PMID- 10723489 TI - [Assessment of inflammatory activity in Crohn disease with hydro-MRI]. AB - PURPOSE: To assess the value of hydro-MRI in the assessment of the activity of Crohn's disease. MATERIALS AND METHODS: After an oral bowel opacification using 1000 ml of a 2.5% mannitol solution, axial and coronal breath-hold sequences (T2W HASTE +/- FS, contrast-enhanced T1W FLASH FS) were acquired in 63 patients with Crohn's disease at 1.0 T. The enhancement of the bowel wall was correlated with other MRI findings, with the Crohn's disease activity index (CDAI), and the C reactive protein (CRP). RESULTS: In Crohn's disease, contrast enhancement of the affected bowel wall is markedly increased in comparison with the normal bowel wall (+80 +/- 23% vs. +43 +/- 11%; p = 8 x 10(-11)). Positive correlations could be established between the increase of bowel wall enhancement and other MRI findings. Between the increase of bowel wall enhancement and the CDAI a poor correlation was found (r = 0.25; p = 0.046). There was no statistical correlation between the increase of bowel wall enhancement and the CRP (r = 0.09; p = 0.24). CONCLUSION: Hydro-MRI allows an assessment of the activity of Crohn's disease. PMID- 10723491 TI - [Magnetic resonance imaging of the wrist--comparison of high resolution pulse sequences and different fat signal suppression techniques in cadavers]. AB - PURPOSE: To evaluate high resolution sequences with and without fat-suppression techniques for MR imaging of the wrist. MATERIALS AND METHODS: 10 cadaver wrist specimens were imaged with 12 MR sequences (SE: 400 ms/20 ms, TSE: 3000 ms/119 ms/17 ms, fatsat (FS) TSE: 3000 ms/17 ms and 3000 ms/45 ms, STIR: 2619 ms/29 ms/160 ms, DESS 3D: 43.7 ms/9 ms/35 degrees FS and 25.4 ms/9 ms/35 degrees water excitation (WE), CISS 3D: 12.2 ms/5.9 ms/40 degrees and FLASH-sequences: 53 ms/11 ms/40 degrees FS, 23 ms/11 ms/40 degrees WE and 45 ms/11 ms/30 degrees FS) at 1.5 T. Slice thickness was 3 mm, FOV 80 x 70 mm (pixel size 0.31 x 0.31 mm). Signal intensity was measured by an ROI in bone marrow, fluid, hyaline cartilage, scapholunate (SL) ligament and triangular fibrocartilage and S/N- and C/N-ratios were calculated. Additionally, a visual evaluation was performed. RESULTS: The highest homogeneity and the least artifacts were achieved by the T1-w SE sequence. For the STIR and PD-FS TSE sequence high rankings were found for the detection of free water. The PD FS sequence had high ranking also for visualization of the SL ligament and the triangular fibrocartilage. The best sequence for the assessment of hyaline cartilage was the FLASH-FS sequence. For detailed analysis of bony structures the CISS sequence performed best. CONCLUSION: The isolated use of a PD-FS-TSE sequence enables for evaluation of all clinically relevant structures at the wrist. Dedicated questions for hyaline cartilage are answered best by the use of a FLASH 3D-FS sequence. Selective water excitation reduces acquisition time to 60%, nevertheless FS sequences are still diagnostically superior to WE sequences. PMID- 10723492 TI - [Laser-induced thermotherapy (LITT) for liver metastasis in an open 0.2T MRI]. AB - OBJECTIVE: To test the feasibility and safety of the laser-induced thermotherapy (LITT) for liver metastases in open MR imaging system operating at 0.2 Tesla. METHOD: Laser therapy using the Nd:YAG laser was performed on 25 patients with a total of 41 liver metastases. An open low-field MRI scanner was used for puncture, positioning of the laser applicator, and monitoring the therapy. A true FISP sequence was used to track the puncture in close to real-time. Localization diagnostics and temperature monitoring were aided by T1-weighted gradient echo sequences in the breath-holding technique. In the first follow up after 24-48 hours, a contrast-enhanced T1-weighted gradient-echo sequence was performed in an MRI scanner at 1.5T. The pre-, intra- and postinterventional volumes of the liver metastases as well as the thermolesions and the thermonecroses were determined. RESULTS: LITT in an open MRI system was technically feasible in all patients with no clinically relevant complications. The mean volumes of the thermolesions measured during intervention in low-field MRI were lower than the volumes of the thermonecroses measured after intervention in high-field MRI. CONCLUSION: The technique presented here of laser-induced thermotherapy for liver metastases in an open MRI system is technically feasible and safe. PMID- 10723493 TI - [Catheter tract embolization after percutaneous transhepatic biliary intervention]. AB - PURPOSE: Evaluation of the feasibility and efficacy of an embolization technique of the transhepatic tract after intervention, in order to prevent peritoneal bile leakage. PATIENTS AND METHOD: Twenty patients (mean age 62 yr) with malignant (17 cases) or benign (three cases) biliary obstruction were treated by percutaneous transhepatic biliary intervention (stent implantation in 17 cases). Mean diameter of the transhepatic tract was 3.2 +/- 0.6 mm. Tract embolization was performed 3.0 +/- 4.0 days after intervention by injecting Histoacryl-Lipiodol via a coaxial catheter-sheath system. RESULTS: Tract embolization was feasible in all cases and resulted in a continuous cast of the catheter tract. There were no signs or symptoms of peritonitis in any of the patients. One patient with stent occlusion developed a biliocutaneous fistula via the former tract after 60 days. CONCLUSION: After embolization of fresh transhepatic tracts there were no signs of leakage during follow-up of the patients. One case with biliocutaneous fistula shows that prevention of leakage is not permanent if biliary reobstruction occurs. PMID- 10723494 TI - [In vitro investigation of biological and technical prosthetic heart valves using MRI: evaluation of possible deflection and heating of the implants]. AB - PURPOSE: In vitro evaluation of possible deflection and heating of present-day prosthetic heart valves during MR imaging at 1.5 T. METHODS: 17 prosthetic heart valves, 12 technical and 5 biological, were investigated using a 1.5 Tesla Siemens Vision system. Deflection was measured at the edge of a 1.5 Tesla superconducting magnet. Each valve was then submerged in a vial of a 1/1 electrolyte solution and temperature was measured before and after imaging with a turbo-spin-echo sequence (TR 5200 ms, TE 138 ms, Flip angle 180 degrees, acquisition time 10.5 minutes, length of echo train 29). MR imaging was performed with phase encoding parallel and perpendicular to the plane of the valves. RESULTS: None of the investigated prosthetic heart valves were deflected. The maximal observed temperature rise was 0.5 degree C. During MR investigation of the prostheses, artifacts caused by metallic parts were less evident using a spin echo sequence than a gradient-echo sequence. CONCLUSIONS: Patients with the tested present-day prosthetic heart valves can be safely imaged by MRI. PMID- 10723495 TI - [Pressure load of the aneurysm sac after endovascular treatment of aortic aneurysm]. AB - PURPOSE: Stent grafting of aortic aneurysms should result in relief of pressure within the excluded aneurysmal sac, however confirming data are not available. This study evaluates the intra-extraluminal pressure translation and translation of maximum pressure increase (dp/dtmax) into the excluded aneurysmal sac after endovascular treatment of experimental aortic aneurysms. MATERIALS AND METHODS: Experimental autologous aneurysms were created surgically using a patch from the sheath of the rectus abdominis muscle in 12 mongrel dogs. After 12 weeks reconvalescence, endovascular treatment was performed viafemoral access using dacron-covered nitinol stents. Spiral CT and angiography were performed at one week and six weeks follow-up and 6 animals each. Laparotomy was performed for hemodynamic measurements. A manometer-tipped catheter was introduced into the excluded aneurysmal sac. A second manometer-tipped catheter was placed intraluminally within the stent graft. Pressure curves and their first derivative dp/dt were simultaneously recorded to calculate the intra-extraluminal transmembraneous pressure transmission. RESULTS: At one week follow-up systolic blood pressure and mean blood pressure were significantly reduced by factors of 0.60 +/- 0.17 (p < 0.01) and 0.78 +/- 0.3 (p < 0.05), respectively in the excluded aneurysm sac. The maximal pressure increase, dp/dt max, was considerably reduced by a factor of 0.06 +/- 0.05 (p < 0.01). However, the diastolic blood pressure was not significantly changed. There were no hemodynamic differences between one and six weeks follow-up. CONCLUSION: In experimental aortic aneurysms, endovascular treatment with stent grafts significantly reduces the systolic peak pressure and dp/dt max in the excluded aneurysmal sac, and thereby significantly reduces the wall stress in the diseased aneurysm wall. Despite complete exclusion of the aneurysm, a considerable pressure load remains in the excluded aneurysmal sac. PMID- 10723496 TI - [Implementation of the quality management system ISO/CD2 9001-2000 in a radiology institute]. AB - Quality assurance in health care, a relatively new discipline, has developed rapidly over the last years and is now required by law. The Quality Management System (QMS) ISO 9001 aims at the definition of requirements necessary in order to achieve perfect products and continuous quality improvement. Implementation of this QMS necessitates the analysis and written documentation of all working processes and modes of operation. Furthermore, potential improvement possibilities are defined to guarantee highly qualified, generally applicable and standardised procedures. Time-consuming data assessment is an instrument to enable disclosure and analysis of existing errors as well as to show possibilities of optimization, thus forming the basis for continuous improvement. The new ISO NORM 9001-2000 is process-orientated with an organisational structure strongly recommendable for service institutions and therefore also for a department of radiology. PMID- 10723497 TI - [Radial k-scanning for real-time MR imaging of central and peripheral pulmonary vasculature]. AB - PURPOSE: To depict the central and peripheral pulmonary vessel anatomy with real time radial MR scanning without respiratory control. METHODS: Three healthy volunteers and one patient with pulmonary embolism proven by spiral CT angiography were studied with a 1.5 T MR imaging system. First, a breath-hold, contrast-enhanced MR angiography was performed for comparison of accuracy. Gradient echo images (TR 16 ms, TE 4 ms, flip angle 18 degrees) were obtained applying a combination of radial scanning and the sliding window reconstruction technique. A dedicated back-projector allowed data reconstruction in real-time with a frame rate of 20 images per second. Scanning was performed during free breathing. RESULTS: The described technique depicted the central and peripheral portions of the normal pulmonary anatomy with comparable image quality as the 3D contrast-enhanced angiography. Visualization of the pulmonary emboli as demonstrated by spiral CT was possible. CONCLUSION: Real-time radial scanning allows promising image acquisition of the pulmonary vasculature without respiratory control. Further technical improvements and clinical trials are required to evaluate its role in the diagnosis of vascular pathologies. PMID- 10723498 TI - Comparison of once and twice daily dosing of didanosine in combination with stavudine for the treatment of HIV-1 infection. AI 454-146 Team. AB - OBJECTIVE: To compare the antiviral activity, safety and tolerability of didanosine dosed once and twice daily when administered in combination with stavudine dosed twice daily in human immunodeficiency virus type 1 (HIV-1) infected individuals with little or no previous exposure to antiretroviral drugs. DESIGN: Comparative, multicentre, randomized, open-label, short-term study. PATIENTS AND METHODS: Eighty-four HIV-1-infected adults with qualifying baseline CD4 cell counts of 200 to 500 cells/mm3 were included in the study. Of these, 43 patients received once daily didanosine plus twice daily stavudine (group A) and 41 subjects received twice daily didanosine plus twice daily stavudine (group B). The primary efficacy analysis used was the time-averaged difference (TAD) between treatment regimens of variations in plasma HIV-1 RNA levels from baseline over the first 12 weeks of therapy. Plasma HIV-1 RNA levels, CD4 cell counts and adverse events were monitored. RESULTS: At week 12, median HIV-1 RNA variations were -1.18 log10 copies/ml in group A and -0.88 log10 copies/ml in group B. For patients who were followed up to week 24, median variations of HIV-1 RNA levels from baseline were -1.21 log10 copies/ml in group A and -0.78 log10 copies/ml in group B. The TAD between the two treatment groups for variations from baseline plasma HIV-1 RNA levels over the first 12 weeks was 0.10 log10 copies/ml (95% confidence interval, -0.19 to 0.40), indicating equivalence. CONCLUSION: Once daily didanosine plus twice daily stavudine and twice daily didanosine plus twice daily stavudine are equally effective in reducing plasma HIV-1 RNA levels and increasing CD4 cell counts. Both regimens are safe and well tolerated. PMID- 10723499 TI - Sensitivity and resistance to (+)-calanolide A of wild-type and mutated forms of HIV-1 reverse transcriptase. AB - We have tested both wild-type and drug-resistant mutated, recombinant human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) molecules for sensitivity to each of two non-nucleoside RT inhibitors (NNRTI), (+)-calanolide A and nevirapine, in primer extension assays. We found that RT containing either the V106A or Y181C substitutions, associated with NNRTI resistance, displayed approximately 90-fold resistance to nevirapine but remained fully sensitive to (+)-calanolide A and that the Y181C mutation marginally enhanced susceptibility to the latter drug. In contrast, the Y188H substitution in RT resulted in about 30-fold resistance to (+)-calanolide A in these assays but did not result in diminished sensitivity to nevirapine. Tissue culture results indicated that the combination of (+)-calanolide A and nevirapine possessed an additive to weakly synergistic effect in blocking replication of HIV-1 in tissue culture. These results suggest that (+)-calanolide A and nevirapine might have rationale as a combination therapy for HIV disease. PMID- 10723501 TI - Second International Workshop on Salvage Therapy for HIV Infection. 19-21 May 1999, Toronto, Canada. PMID- 10723500 TI - Intravenous ribavirin for hantavirus pulmonary syndrome: safety and tolerance during 1 year of open-label experience. Ribavirin Study Group. AB - Intravenous ribavirin was provided non-selectively for investigational open-label use among persons with suspected hantavirus pulmonary syndrome (HPS) in the United States between 4 June 1993 and 1 September 1994. Therapy was initiated prior to laboratory confirmation of hantavirus infection because most deaths from HPS occur within 48 h of hospitalization. Thirty patients with confirmed HPS, 105 patients without HPS and 5 patients without adequate diagnostic testing for HPS were enrolled. This observational study arguably provides the most complete information available on ribavirin-associated adverse effects. Although ribavirin was generally well tolerated, 71% of recipients became anaemic and 19% underwent transfusion. An apparent excess of hyperamylasaemia/pancreatitis was either therapy-associated or due to enrollment bias. The 30 enrolled HPS patients had a case-fatality rate of 47% (14/30). It is not possible to assess efficacy with this study design. However, comparison of survival curves for the 30 enrolled HPS patients and 34 patients who developed HPS during the same time period but were not enrolled did not suggest an appreciable drug effect. A randomized, placebo controlled trial that enrolls patients during the prodrome phase would be necessary to assess the efficacy and further define the safety of intravenous ribavirin for HPS. PMID- 10723502 TI - Resistance to antiretroviral drugs. AB - There are many questions regarding testing for viral resistance to antiretroviral drugs and interpretation of test results. The most appropriate time for testing and the compartment from which to derive virus are still uncertain. Both phenotypic and genotypic tests are used to measure drug susceptibility; however, interpretation of results of these assessments is not clear-cut. Furthermore, correlation of these test results with clinical outcome or pharmacological data is limited by lack of studies and other confounding factors. PMID- 10723503 TI - Pharmacological considerations in antiretroviral therapy. AB - Suboptimum drug exposure arising from pharmacological and pharmacokinetic factors is an important reason that combination antiretroviral therapy fails in patients with human immunodeficiency virus type 1 infection. With all three available drug classes, the major causes are drug-drug interactions and interpatient pharmacokinetic variability. Inadequate concentrations of protease inhibitors and non-nucleoside reverse transcriptase inhibitors occur when concurrently administered drugs interfere with their metabolism by the hepatic cytochrome P450 enzyme system, the major metabolic pathway for these drugs. Combined therapy with certain nucleoside reverse transcriptase inhibitors can reduce systemic drug exposure by interfering with the intracellular conversion of administered drugs to their active metabolites. Important causes of interpatient variability in pharmacokinetic parameters include low oral drug bioavailability and individual differences in drug disposition and metabolism. Careful selection and monitoring of combination drug therapy along with individualized rather than standard dosage regimens may minimize these pharmacological problems and help ensure optimum antiviral activity. PMID- 10723504 TI - Sanctuary sites in HIV-1 infection. AB - The existence of sanctuary sites for human immunodeficiency virus type 1 (HIV-1) may potentially endanger the efficacy of antiretroviral therapy in the long term and may even make eradication of HIV-1 from the infected body impossible. Potential 'classic' sanctuary sites for HIV-1 are the central nervous system and the testes, but long-lived cell populations (such as macrophages) or latently infected (resting) CD4 cells may also be considered a sanctuary for HIV-1. These potential sanctuary sites, and putative underlying biochemical mechanisms such as the divergent phosphorylation properties of nucleoside reverse transcriptase inhibitors in different cell populations and the affinity of drugs for the multidrug transporter P-glycoprotein, are discussed. PMID- 10723505 TI - Immunological effects of antiretroviral therapy. AB - The striking decline in HIV load with highly active antiretroviral combination therapy (HAART) is accompanied by substantial improvements in immune function, even in patients with more advanced disease. These include a general reduction in immune activation with increases in total CD4 and CD8 cell counts, memory and naive T cell subsets and antigen responses to certain opportunistic pathogens. At this time, it appears that HAART-induced improvements in function are limited to those T cells that have not yet been completely depleted by HIV. Long-term studies are needed to determine whether complete functional restoration of the repertoire is possible. Clinically, HAART improves survival and reduces progression of HIV disease. Some patients with active opportunistic disorders demonstrate complete or partial resolution of the infection or malignancy. However, persons with subclinical Mycobacterium avium complex or cytomegalovirus retinitis and those with chronic hepatitis B virus infection may sometimes experience acute flares if prophylactic therapy against the underlying disorder is not included in the regimen. PMID- 10723506 TI - Hydroxyurea: mechanisms of HIV-1 inhibition. AB - Hydroxyurea inhibits cellular ribonucleotide reductase, resulting in decreased pools of dNTPs and thus inhibition of DNA synthesis. Studies in vitro have shown that hydroxyurea reduces dNTP pools in cells infected with human immunodeficiency virus type 1 (HIV-1), inhibiting HIV-1 DNA synthesis in infected quiescent and activated primary human lymphocytes and macrophages. Hydroxyurea also potentiates the activity of nucleoside reverse transcriptase inhibitors (NRTIs): the activated triphosphate forms of NRTIs compete with naturally occurring dNTPs for incorporation into nascent viral DNA during reverse transcription. A synergistic effect is observed between hydroxyurea and didanosine (2',3'-dideoxyinosine; DDI). This combination exerts persistent suppression of HIV-1 replication without evidence of viral rebound for over 1 year in HIV-1-infected patients. Didanosine resistant HIV-1 mutants retain sensitivity to didanosine in the presence of hydroxyurea. The incorporation of didanosine triphosphate by resistant reverse transcriptase is increased in the context of the hydroxyurea-induced depletion of dATP. Although hydroxyurea has a reduced effect on dNTPs competing with the triphosphate forms of pyrimidine NRTIs, it appears to augment the anti-HIV-1 activity of these agents by increasing their intracellular phosphorylation; this may be of particular interest for salvage strategies given recent data indicating disruption of NRTI phosphorylation with specific NRTI treatment regimens. Finally, by exerting a cytostatic effect on CD4 and CD8 T lymphocytes, hydroxyurea may (i) reduce HIV-1 replication by decreasing CD4 T cell proliferation; and (ii) prevent the exhaustion of CD8 T cell populations that may occur as a result of excessive activation in the context of HIV-1 infection. PMID- 10723507 TI - Didanosine once daily: an overview. AB - The current focus on simplifying treatment regimens for patients with human immunodeficiency virus (HIV) infection has contributed to the interest in once daily therapy. The triphosphate of didanosine (2',3'-dideoxyinosine or DDI) has a long intracellular half-life, which supports the use of didanosine in a once daily dosing schedule. Early clinical studies found that changes in surrogate markers were similar whether didanosine was dosed once or twice daily, while toxic effects occurred less frequently with once-daily dosing. Didanosine once-a day used in combination with other drugs has also been studied, and results of some of these trials are summarized in this paper. PMID- 10723508 TI - Stavudine plus a non-thymidine nucleoside reverse transcriptase inhibitor as a backbone for highly active antiretroviral therapy. AB - Current guidelines for treatment of human immunodeficiency virus (HIV) disease favour the use of triple-drug combinations consisting of two nucleoside analogue reverse transcriptase inhibitors (NRTIs) plus a protease inhibitor to achieve maximum suppression of HIV replication. There is considerable evidence for including one thymidine NRTI to target activated HIV host cells and one non thymidine NRTI to target quiescent host cells as the backbone of such regimens. A number of recent studies have shown that stavudine in combination with didanosine or lamivudine is at least as effective as zidovudine-based combinations with regard to virological outcomes, with available data suggesting an enhanced effect on immunological outcome with stavudine-based combinations. When considered along with such advantageous characteristics of stavudine as infrequent and low-level resistance and good cerebrospinal fluid penetration, these findings indicate that stavudine in combination with didanosine or lamivudine should be considered for use as the backbone of multiple-agent highly active antiretroviral therapy (HAART). PMID- 10723509 TI - Post-exposure prevention of HIV-1 infection. AB - Post-exposure prevention of human immunodeficiency virus type 1 (HIV-1) infection involves a combined modality approach to subjects exposed to the HIV-1 virus through sexual or occupational contact or intravenous drug use. Identification of those at highest risk of transmission, which involves patient factors as well as the infectivity of the partner and the local prevalence of HIV-1, will help to determine appropriate candidates for post-exposure treatment. Early institution of therapy (within 72 h) should be followed with education and counselling to help maximize therapeutic adherence while reinforcing safer behaviour for the future. PMID- 10723510 TI - Early antiretroviral therapy: rationale, protease inhibitor-sparing regimens and once daily dosing. AB - In 1998 it seems reasonable and widely accepted that all human immunodeficiency virus type 1 (HIV-1)-infected patients willing to be treated may benefit from receiving antiretroviral therapy. Only those with undetectable plasma HIV-1 RNA, normal CD4 lymphocyte counts and lack of markers of immunological system activation may be possible exceptions. The rationale supporting the early initiation of antiretroviral therapy are (i) data on viral dynamics; (ii) preliminary data pointing toward a better and a quicker restoration of immune function when treatment is initiated in very early stages (during or within a few weeks or months of acute symptomatic or asymptomatic HIV-1 infection); (iii) the lack of a stable viral load set-point even in patients in the early stages (CD4 > 500 cells/mm3) who have a very low viral load (< 5000 copies/ml); (iv) the relatively high likelihood of clinical progression at mid-term of the approximately 50-75% of patients in very early disease stages (CD4 > 500 cells/mm3) who have a plasma viral load above 5000 to 10,000 HIV-1 RNA copies/ml; (v) data from the Spanish Earth-1 study, which used a composite endpoint (virological, immunological or clinical progression), demonstrating that even in these very early stages of HIV-1 disease any antiretroviral therapy (double or triple combination) was better than no treatment. Even in early disease stages, a triple combination is needed to achieve a durable and profound virological and immunological response. In addition, the combination of stavudine plus didanosine has several advantages and can be considered one of the best double nucleoside combinations to combine with a protease inhibitor or with a non-nucleoside reverse transcriptase inhibitor. The INCAS study and the preliminary results of the ongoing Spanish SCAN study have demonstrated the possibility of protease inhibitor-sparing combinations for initial antiretroviral treatment, at least in selected patient subsets, such as those with a relatively low baseline viral load. Furthermore, components of these protease inhibitor-sparing combinations such as didanosine or nevirapine are suitable for once daily administration. PMID- 10723511 TI - Nevirapine/didanosine/lamivudine once daily in HIV-1-infected intravenous drug users. AB - Human immunodeficiency virus (HIV)-infected, active intravenous drug users received once-daily therapy consisting of a combination of didanosine (2',3' dideoxyinosine or DDI), lamivudine [(-)-beta-L-2',3'-dideoxy-3'-thiacytidine or 3TC] and nevirapine. Preliminary results for the first 24 weeks show that the regimen provides potent immunological and antiviral effects. PMID- 10723512 TI - Stavudine plus didanosine and nevirapine in antiretroviral-naive HIV-infected adults: preliminary safety and efficacy results. VIRGO Study Team. AB - The objective of this open-label trial is to evaluate the virological and immunological effects of triple therapy with stavudine (40 mg twice daily if > or = 60 kg, 30 mg twice daily if < 60 kg)/didanosine (400 mg once daily if > or = 60 kg, 300 mg once daily if < 60 kg)/nevirapine (200 mg daily from day 1 to 14, then 200 mg twice daily) in 60 antiretroviral-naive HIV-infected adults with CD4 cell counts > or = 200 cells/mm3 and plasma HIV RNA > or = 5000 copies/ml. At present, 59 patients have begun receiving the trial regimen. Characteristics of patients at baseline were as follows: 46 men/13 women, CDC stage A, 75%; mean CD4 cell count, 429 cells/mm3; mean HIV RNA, 4.6 log10 copies/ml). Mean decrease of viral load was -1.9 log10 at week 4 (n = 39), -1.9 log10 at week 16 (n = 20), with HIV RNA below the detectable level (< 500 copies/ml) in 62% of patients at week 4 and 85% at week 16. Mean CD4 cell count increase was +118 cells/mm3 at week 4. Cutaneous intolerance occurred within the first 4 weeks in 11/59 (19%) patients after a mean of 14 days (range, 3-24 days) and led to nevirapine discontinuation in 3/11 patients. Preliminary results of this ongoing trial show that combination therapy with stavudine/didanosine/nevirapine is a convenient (seven pills in two daily intakes) triple-therapy regimen with rapid immunological and antiviral effects. Rash, frequent in the first weeks of therapy, usually can be managed without stopping nevirapine. Long-term suppression of plasma HIV RNA with this combination needs to be confirmed but may support use of nevirapine as a component of first-line anti-HIV therapy along with two nucleosides. PMID- 10723513 TI - dNN study: stavudine, nelfinavir and nevirapine. Preliminary safety, activity and pharmacokinetic interactions. AB - The dNN study evaluated the safety, efficacy and pharmacokinetic interactions of the combination of stavudine (2',3'-didehydro-2',3'-dideoxythymidine; D4T), nelfinavir and nevirapine in 25 HIV-infected subjects who received treatment for up to 29 weeks. This brief report presents the study design and preliminary data. PMID- 10723514 TI - Ongoing open-label trials of triple therapy with stavudine and lamivudine or stavudine and didanosine plus nelfinavir. AB - The VZN and ZEN studies are non-comparative trials assessing triple combination therapy comprising thymidine and non-thymidine analogue nucleoside reverse transcriptase inhibitors plus a protease inhibitor. Preliminary data are available for the VZN study and recruitment for the ZEN study is underway. PMID- 10723515 TI - Didanosine plus stavudine with or without hydroxyurea in HIV-1-infected patients: 1 year follow-up. Swiss HIV Cohort Study. AB - A total of 144 human immunodeficiency virus (HIV)-infected patients (mean CD4 cell count, 367 cells/mm3) were included in a double-blind placebo-controlled trial testing the efficacy on surrogate markers of HIV progression of the combination didanosine (2',3'-dideoxyinosine or DDI) plus stavudine (2',3' didehydro-2',3'-dideoxythymidine or D4T) with or without hydroxyurea. The primary end point was a reduction of HIV RNA levels to below 200 copies/ml after 12 weeks of treatment. The results showed that the triple combination was associated with a more profound decrease in HIV RNA with an increased proportion of patients with viraemia < 200 copies/ml. This effect persisted for the majority of the patients after a 48 week follow-up. In contrast, the increase in CD4 cell counts was less in patients treated with hydroxyurea because of lymphopenia, and adverse events were more frequent in hydroxyurea-treated patients. In conclusion, the addition of hydroxyurea consistently improved the antiviral activity of the didanosine/stavudine combination over a 48 week follow-up. Increased toxicity and decreased effect on CD4 cell counts might inspire caution. PMID- 10723516 TI - Stavudine-based multiple agent combinations: initial studies and ongoing comparative trials. AB - Initial studies of multiple-agent antiretroviral combinations including the thymidine nucleoside analogue reverse transcriptase inhibitor (RTI) stavudine (2',3'-didehydro-2',3'-dideoxythymidine; D4T) have shown potent anti-HIV effects in both treatment-naive and -experienced patients. A number of ongoing randomized comparative trials are assessing stavudine-based multiple agent combinations, including protease inhibitor-containing and -sparing regimens, triple nucleoside RTI regimens and regimens including non-nucleoside RTIs and/or hydroxyurea as initial or subsequent therapy. Results of these studies should help to define additional first-line treatment options and strategies for sequencing antiretroviral treatment regimens. PMID- 10723518 TI - Cellular factors: targets for the treatment of HIV infection. AB - On 19-20 April 1998, researchers and clinical investigators from around the world gathered in Pavia, Italy, for Cellular Factors: Targets for the Treatment of HIV Infection, a comprehensive 2 day meeting designed to share the most recent findings in human immunodeficiency virus (HIV) and AIDS treatment research. Because viral replication is dependent on the host cell machinery, many researchers are seeking new treatment strategies that control HIV by limiting the availability of cellular proteins and metabolic functions. Other approaches, such as gene therapy, seek to confer cellular resistance to the effects of viral gene products. Conference participants discussed a range of new therapeutic approaches, as well as new research into the workings of the cells that are targets of HIV infection. The meeting was sponsored by the Research Institute for Genetic and Human Therapy (RIGHT), with the support of Bristol-Myers Squibb Immunology and Policlinico San Matteo. Established in 1994 and codirected by Drs Julianna Lisziewicz and Franco Lori, RIGHT is a non-profit organization dedicated to translating basic research into clinical trials. With laboratories at Georgetown University in Washington, DC, and at the Policlinico San Matteo in Pavia, Italy, RIGHT works with an international research network of molecular biologists, virologists and immunologists to understand how diseases might be attacked at the molecular level. Current research focuses on prevention and treatment of HIV infection. Several clinical studies are now underway. Hydroxyurea, which targets a cellular enzyme, is being tested in combination with antiretroviral drugs to inhibit HIV-1 replication and control the onset of resistance. In gene therapy pilot studies, a novel antiviral gene is being tested for its ability to confer cellular resistance to HIV. PMID- 10723517 TI - Strategies for rescue therapy. AB - Human immunodeficiency virus (HIV) RNA load will be detectable in approximately 40% of non-naive HIV patients who are taking triple therapy after 1 year of follow-up. Treatment failure may involve factors related to the physician (poor prescribing), the patient (poor compliance due to complicated regimens or side effects), or the drug (pharmacological aspects such as poor bioavailability). The primary mechanism for drug failure is viral resistance to available HIV therapies. Whereas initial therapy should be aggressive, second- or third-line therapy demands a more measured approach, as treatment options become more limited. Rescue or salvage therapy refers to therapy after everything else has failed. Issues related to rescue therapy for HIV-positive subjects include drug holidays, drug recycling and the potential for new drugs in this setting. PMID- 10723519 TI - Low birthweight associated with maternal anaemia and Plasmodium falciparum infection during pregnancy, in a peri-urban/urban area of low endemicity in Uganda. AB - A cross-sectional study of pregnant women was conducted at Nsambya Hospital in Kampala, to investigate the prevalence and effect of Plasmodium falciparum infections during pregnancy, in a peri-urban/urban location. Overall, 544 pregnant women were recruited when they presented at the labour ward for delivery. After giving informed consent, each subject answered a questionnaire and underwent a physical examination, and peripheral-blood samples were obtained. After each uncomplicated delivery, samples of placental and cord blood were obtained from the placenta and infant, respectively, and infant birthweights were recorded. Smears were prepared from the blood samples and checked for parasites. Only 46 and 36 of the 537 women investigated were positive for P. falciparum infection in their peripheral and placental blood, respectively. Plasmodium falciparum was the only parasite encountered. The prevalences of low birthweight and maternal parasitaemia and the intensities of maternal infection were each greater in primigravidae than secundi- or multi-gravidae. Despite the low prevalence of parasitaemia in this population, P. falciparum infection in the primigravidae was a significant contributor to their ill health, leading to low birthweights in their infants. PMID- 10723520 TI - Malariometric update for the rainforest and savanna of Ashanti region, Ghana. AB - The epidemiological features of human infection with Plasmodium were studied in a community-based survey of 35 villages in the Ashanti region of Ghana. The overall prevalences of malarial parasitaemia in subjects aged > or = 2 years were 50.72% in forest areas and 49.72% in savanna. Plasmodium falciparum was the predominant species everywhere, followed by P. malariae in the savanna and P. ovale in the forest. The highest prevalence of asexual parasitaemia (of any species) occurred in the youngest age-group (2-9 years). The geometric mean intensities of parasitaemia among the parasitaemic (i.e. the parasite density indices) were 557, 640 and 452 parasites/microliter for P. falciparum, P. ovale and P. malariae, respectively. For each Plasmodium species encountered, the mean intensity of parasitaemia decreased with age. Mixed infections were observed in 24% and 30% of the parasitaemic subjects from the forest and savanna, respectively. Those infected with P. falciparum were more likely to carry P. ovale (odds ratio = 2.02) or P. malariae (odds ratio = 2.63) than those who were not infected with P. falciparum. Mean intensities of the parasitaemias in mixed infections were substantially higher than the sums of those in the corresponding single infections. When comparing villages, parasite density indices were found to be correlated with the prevalences of parasitaemia (r = 0.56). PMID- 10723521 TI - The chemotherapy of rodent malaria. LVIII. Drug combinations to impede the selection of drug resistance, Part. 2: The new generation--artemisinin or artesunate with long-acting blood schizontocides. AB - The search for combinations of antimalarial drugs that will impede the selection of drug resistance, especially in Plasmodium falciparum, is currently focused on the use of a member of the artemisinin family, with a short half-life, in association with a relatively long-acting blood schizontocide. Experiments with such 'third-generation' combinations, in mice infected either with chloroquine sensitive P. berghei or P. chabaudi, or chloroquine-resistant P. yoelii ssp. NS, have produced interesting results. The data collected, using the '2% relapse technique' (2%RT), indicate that a combination of artemisinin with mefloquine can impede to a significant degree, although by no means completely, the selection of resistance to both compounds in P. berghei and in P. yoelii ssp. NS. Similarly, a combination of artesunate with pyronaridine impedes the selection of resistance to these compounds in P. berghei. Parallels are drawn between observations with such combinations in man and in the rodent models which, it is argued, once again demonstrate their value in predicting the protective value of using different types of antimalarials together. Evidence is presented that resistance to single compounds may emerge more rapidly when a high dose is employed in the 2%RT than a lower dose. It is noted also that the rate at which resistance to pyronaridine is selected by a given dose varies with the species of rodent Plasmodium, and the relevance of this to the malarial parasites of human is discussed. PMID- 10723522 TI - Clinical manifestations of visceral leishmaniasis associated with HIV infection: a retrospective study of 91 French cases. AB - A retrospective study was conducted in France to determine the clinical features of visceral leishmaniasis (VL) seen, between 1986 and 1997, in 91 patients co infected with human immunodeficiency virus (HIV). Fever (87% of patients), splenomegaly (74%) and hepatomegaly (49%) were common, 43% of the patients having all of these signs and only 9% having none of them. Amastigotes were reported in atypical locations in 31 (34%) of the patients, and 15 patients had only had their VL diagnosed following accidental discovery of amastigotes in samples from their digestive tract and lungs (one), digestive tracts only (11 patients), lungs only (two), or skin (one). Some of the digestive symptoms observed are probably attributable to the intestinal infections with Leishmania. Overall, VL diagnosis was fortuitous in 27% of the subjects. Even in endemic areas, therefore, VL is not considered routinely by physicians attempting diagnoses. PMID- 10723523 TI - Altered calcium homeostasis and membrane destabilization in erythrocytes of hamsters infected with Leishmania donovani. AB - Homeostatic mechanisms regulating intracellular concentrations of Ca2+ at a low level are prerequisites for maintaining the integral and cytoskeletal structure of erythrocytes under normal physiological conditions. The present study was undertaken to assess the contribution of Ca2+ homeostasis in modifying red-cell stability in hamsters, during the anaemia caused by Leishmania donovani. Erythrocytes from the infected animals became increasingly fragile as infection progressed. This fragility may be the result of a gradual change in membrane permeability, as indicated by enhanced uptake of 45Ca2+. The increase in cytosolic Ca2+ and decrease in membrane-bound Ca2+ observed indicate the release of Ca2+ from the membrane store, leading to [Ca2+]i accumulation in the later stages of the post-infection period. Decline in the efficacy of Ca(2+)-effluxing enzyme may also contribute to the enhanced [Ca2+]i level, with subsequent degradation of membrane proteins in the erythrocytes of the infected animals. Marked inhibition of proteolytic degradation by the Ca(2+)-dependent thiol protease inhibitor leupeptin, with concomitant thiol depletion, indicates the involvement of Ca(2+)-induced thiol protease in the observed degradation of membrane proteins. The results indicate that an altered Ca2+ homeostasis in erythrocytes following leishmanial infection causes enhanced cellular accumulation of Ca2+, which in turn may lead to haemolysis in experimental visceral leishmaniasis. PMID- 10723524 TI - Patterns of infection of haemoparasites in the fat sand rat, Psammomys obesus, in Tunisia, and effect on the host. AB - Two bacterial and one protozoan blood parasite, belonging to the genera Bartonella, Borrelia and Babesia, were studied in a Tunisian population of Psammomys obesus. Seasonal changes in the abundance of the parasites and host were monitored in a longitudinal field survey lasting 17 months. Blood samples collected during eight rodent-trapping sessions, between September 1995 and January 1997, were examined microscopically. Bartonella sp. showed a seasonal pattern, with most transmission occurring in summer and autumn; most rodents (90%) were infected in August-September, when they were at low density and adult. Borrelia sp. showed low prevalences, with few seasonal fluctuations, and Babesia sp. showed an intermediate pattern, differing from one year to another. In the cohort of adult rats, infections with Bartonella sp. and Babesia sp. were less prevalent in winter than in the previous summer. Single and mixed infections were equally prevalent in females and males, and in sexually active and inactive adults. In addition, infection had no apparent effect on the weight of adult P. obesus. The observation that the proportion of erythrocytes infected with Bartonella sp. decreased with increasing host age is probably indicative of some acquired immunity to this micro-organism. The absence of detectable infections with Borrelia sp. in old rats indicates that the prevalence and/or intensity of infection declines with host age or that infected animals die selectively. However, there was no indication that any of these parasites combined sufficient pathogenicity and abundance to have any measurable effect on the rodent population. PMID- 10723525 TI - Risk factors associated with human cystic echinococcosis in Jordan: results of a case-control study. AB - The parasites causing cystic echinococcosis (CE) are transmitted to man and domestic animals either directly or indirectly from dogs. High levels of human infection have been frequently described in sheep-rearing areas of the world, where the infection cycles between dogs and sheep through the use of working dogs and the feeding of sheep offal to dogs. A case-control study was undertaken in northern Jordan to examine the epidemiological characteristics of echinococcosis in a Muslim community. Forty-four indigenous Jordanians who had been treated for CE between 1990 and 1996 were contacted and three controls for each case, matched for sex and age, were selected from the Jordanian population. The most important single factor associated with treatment for CE was the main source of domestic water; 42 (95%) of the cases but only 81 (61%) of the controls reported that they used a public, piped, water supply as their principal source of household water [odds ratio (OR) = 13.22; 95% confidence interval (CI) = 2.91-83.7]. The keeping of dogs or close association with dogs or farm animals was not associated with any increased risk of CE. However, individuals who grew their own vegetables had a significantly decreased risk of acquiring CE (OR = 0.30; CI = 0.08-0.98). There was evidence of widespread ignorance of the disease and how it is transmitted. PMID- 10723526 TI - Apparent failure of ultrasonography to detect adult worms of Brugia malayi. AB - Adult worms of Wuchereria bancrofti, or rather their characteristic movements (the 'filarial dance'), can now be detected in the scrotal lymphatics of microfilaraemic males, using ultrasonography. This ability has been used to delineate the lymphatic pathology of bancroftian filariasis, guide the surgical removal of the adult worms and, most importantly, assess the macrofilaricidal effects of antifilarial drugs. In the present study, the first report of the use of ultrasonography in brugian filariasis, 22 men (aged 18-62 years) with 60-2972 (median = 370) Brugia malayi microfilariae/ml blood were subjected to ultrasonography using a linear, 7.5-MHz probe. In addition, four other men (aged 19-35 years), with W. bancrofti microfilaraemia [28-524 (median = 234) microfilariae/ml], were similarly examined. Adult worms were not detectable in any of the patients with B. malayi parasitaemia but were detected in the scrotal lymphatics of two of the four individuals with W. bancrofti infection. The reasons for the failure to detect adult B. malayi and the limitations of ultrasound as a screening tool are examined. The results highlight the differences between the two species that cause most lymphatic filariasis and the need for rapid development of tools that can be used for the control of brugian lymphatic filariasis. PMID- 10723527 TI - Recovery of a species of Brugia, probably B. ceylonensis, from the conjunctiva of a patient in Sri Lanka. AB - A species of Brugia, probably B. ceylonensis, was recovered from the conjunctiva of a patient in Sri Lanka for the first time. This infection represents only the second record of Brugia in the human conjunctiva, and is clearly zoonotic, acquired from a dog. Brugia ceylonensis has a distinct head bulb like that of Wuchereria bancrofti and B. malayi. However, the parasite recovered was not W. bancrofti, as specific IFAT and DNA probes gave negative results, and B. malayi is believed to have been eradicated from Sri Lanka several years ago. The presence of a distinct head bulb excludes the possibility that the parasite was B. buckleyi. PMID- 10723528 TI - The transmission status of Bulinus on Zanzibar Island (Unguja), with implications for control of urinary schistosomiasis. PMID- 10723529 TI - [Allergies, an increasing public health problem: causes and consequences]. AB - Allergies due to IgE immunoglobulins and belonging to the atopic syndrome, such as asthma, allergic rhinitis and atopic eczema have increased in prevalence three times during the past 20-30 years in most industrialized countries. The causes of that increase are still much debated but seem to be related to multiple changes in the environment. Our recent studies on dog atopy indicate that the interaction of a dominant gene responsible for high IgE production but with variable expression, according to several environmental factors acting during infancy, could explain observations made in man. Allergic diseases have become an important portion of public health costs, amounting to approximately 200 billions French Francs in the European Community. Early and adequate care for the about 20% of allergic patients which are severely affected, as well setting up a systematic prevention policy would have a moderating influence on the increasing costs, and would achieve improvements in the quality of life of allergic patients. Appropriate measures include fostering medical and political awareness about the problem's urgency, the formation of an appropriate body of specialists and an Allergological education at all levels, as well as the definition of a screening and care providing policy taking in account existing medical structures. PMID- 10723530 TI - [The multifood allergy syndrome]. AB - Multiple food intolerance in infants and young children is increasingly diagnosed. More than 40% of infants less than 1 y.o. could be affected. The syndrome is characterized by the seriousness of atopic dermatitis (SCORAD > 50), by enterocolitis or failure to thrive or various associations of symptoms that may change over time. The evolution is long-lasting. Common food allergens are milk, egg, soy, wheat, but other ones can be implicated. The diagnosis is established by standardized oral challenges. Multiple etiopathogenic factors are involved: atopy, gastro-enteritis induced intestinal hyperpermeability, precocity of food diversification, breast-feeding continued after the onset of symptoms. Amino-acid based formulas have changed the evolution. PMID- 10723531 TI - [The wasp-mosquito syndrome: extension of cross-allergenicity to the horsefly]. AB - The crossed allergenicity between wasp venom and mosquito extract was shown during recent work based on clinical observations and correlation studies between different biological parameters, indicating an IgE-dependent biological mechanism. A common protein was identified by Immunoblot. From observation of one of our patients involved in this work, we examined the possibility of the extension of crossed reactivity between wasp, mosquito and horsefly. In effect, our patient presented an anaphylactic reaction with neurological complications from attack on the central grey nucleus, shown by IRM; the immunological study showed a common protein between wasp venom and the total extracts of mosquito and horsefly. PMID- 10723532 TI - [Value of specific IgE and IgG4 measurements in the follow-up of a hymenoptera venom immunotherapy treatment: apropos of 82 cases]. AB - The work is devoted to the study of 82 dossiers of patients who were treated by immunotherapy (IT) to hymenoptera venoms, following various manifestations. The study has permitted analysis of the value of measurement of specific immunoglobulins, both IgE and IgG4 (1), in following IT in patients who were treated for at least three years. It especially emphasized the usefulness of adopting management that is appropriate to the clinical and biological characteristics of each patient. PMID- 10723533 TI - Resilience among urban African American male adolescents: a study of the protective effects of sociopolitical control on their mental health. AB - Resilience refers to the notion that some people succeed in the face of adversity. In a risk-protective model of resilience, a protective factor interacts with a risk factor to mitigate the occurrence of a negative outcome. This study tested longitudinally the protective effects of sociopolitical control on the link between helplessness and mental health. The study included 172 urban, male, African American adolescents, who were interviewed twice, 6 months apart. Sociopolitical control was defined as the beliefs about one's capabilities and efficacy in social and political systems. Two mental health outcomes were examined--psychological symptoms and self-esteem. Regression analysis to predict psychological symptoms and self-esteem over time were conducted. High levels of sociopolitical control were found to limit the negative consequences of helplessness on mental health. The results suggest that sociopolitical control may help to protect youths from the negative consequences of feelings of helplessness. Implications for prevention strategies are discussed. PMID- 10723534 TI - If it's offered, will they come? Influences on parents' participation in a community-based conduct problems prevention program. AB - This study examined influences on the rate and quality of parent participation in the Fast Track Program, a multi-system, longitudinal preventive intervention for children who are at risk for conduct problems. A theoretical model of the relations among family coordinator characteristics, parent characteristics, the therapeutic engagement between family coordinator and parent, and rate and quality of parent participation was the basis for this study. "Family coordinators" are the Fast Track program personnel who conduct group-based parent training sessions and home visits. Participants in this study included 12 family coordinators (42% were African American, 58% European American) and 87 parents (55% were African American, 45% European American). The level of therapeutic engagement between the parent and the family coordinator was positively associated with the rate of parent attendance at group training sessions. The extent of family coordinator-parent racial and socioeconomic similarity and the extent of the family coordinator's relevant life experiences were highly associated with the level of therapeutic engagement. The quality, but not the rate, of participation was lower for African American parents. Implications of these findings for preventive intervention with this population are discussed. PMID- 10723535 TI - Ecological factors impacting provider attitudes towards human service delivery reform. AB - Although reform efforts are substantially altering the structural operations and guiding ideological framework of the human service delivery system, little empirical work has been done to systematically examine these transformations. This study examines providers' attitudes regarding two reform elements that are being widely implemented: an increased emphasis on interagency collaboration and a shift from a medical model service delivery philosophy, that focuses on client deficits, to one that emphasizes consumer strengths. Through survey data collected from 186 providers from 32 human service agencies in one county, the relationship between providers' perceptions of contextual support for human service delivery reform and providers' attitudes towards these initiatives is explored. The findings from this study support the importance of attending to the ecology in which we initiate system reform efforts. For both reform elements, working within contexts that are perceived as providing ideological and functional support for change was associated with positive provider attitudes towards those changes. Staffs' perceptions of the external environment played the most critical role in shaping staff attitudes. Interestingly, unique aspects of providers' work environments were related to positive attitudes towards the two different reforms. The implications of these findings for the success of human service delivery reform are discussed. PMID- 10723536 TI - A longitudinal assessment of teacher perceptions of parent involvement in children's education and school performance. AB - This study examines the ways in which parental involvement in children's education changes over time and how it relates to children's social and academic functioning in school. Teachers provided information on parent involvement and school performance for 1,205 urban, kindergarten through third-grade children for 3 consecutive years. They rated the following four dimensions of parent involvement: frequency of parent-teacher contact, quality of the parent-teacher interactions, participation in educational activities at home, and participation in school activities. As predicted, the frequency of parent-teacher contacts, quality of parent-teacher interactions, and parent participation at school declined from Years 1 to 3. Every parent involvement variable correlated moderately with school performance and parent involvement in Years 1 and 2, and accounted for a small, but significant amount of variance in Year 3 performance after controlling for initial performance level. Participation in educational activities at home predicted the widest range of performance variables. Results suggest that enhancing parental involvement in children's schooling relates to improvements in school functioning. PMID- 10723537 TI - Giving and receiving help: interpersonal transactions in mutual-help meetings and psychosocial adjustment of members. AB - The helping transactions that occur in group meetings have been theorized to be important therapeutic mechanisms within mutual-help (or self-help) groups. Hypothesized links between giving and receiving help and psychosocial adjustment were examined in a mutual-help group for individuals with serious mental illness (GROW). Participants' adjustment was assessed at two time points and helping behaviors were measured with observational coding of weekly group interactions during the period between assessments. Frequencies of helping behaviors were used to predict Time 2 adjustment after controlling for initial adjustment. Consistent with the helper therapy principle, giving help to others predicted improvements in psychosocial adjustment; giving advice was a unique predictor. Total amount of help received was not associated with adjustment, but receiving help that provided cognitive reframing was associated with better social adjustment. A predicted interaction suggested that receiving help was related to better functioning when members experienced high levels of group integration. PMID- 10723538 TI - Ethnicity and social support during pregnancy. AB - Data from two multi-ethnic prospective studies of African American, Latina, and non-Hispanic White pregnant women were used to examine the influence of contextual factors on social support processes during pregnancy. Multiple types of support (perceived support, received support, support satisfaction, network support) and sources of support (baby's father, family, friends) were assessed. The role of ethnicity in social support was examined after controlling for the contribution of related contextual factors (SES, marital status, age, parity, employment) to these processes. The impact of ethnicity and related contextual factors differed across sources of social support. Ethnic differences in support from family and friends, but not from the baby's father, emerged. However, marital status was a consistent predictor of support from the baby's father, and SES was a consistent predictor of support from friends. Overall, the findings of two studies suggest that although ethnicity is associated with support from friends and family, other contextual factors, such as marital status and SES, influence support processes during pregnancy. PMID- 10723539 TI - Profiles of self-esteem in early adolescence: identification and investigation of adaptive correlates. AB - Investigated profiles of self-esteem during early adolescence and their adaptive correlates in two separate longitudinal studies. Using multidimensional ratings of self-esteem within a developmental-ecological framework, cluster analysis revealed five distinct profiles for each sample. The profiles found were characterized by differing patterns of self-evaluation across major contexts of development, including consistently positive or negative ratings for all domains as well as more variable patterns in which ratings for one or more domains (e.g., school) were elevated or diminished relative to those for other areas. Profiles, in turn, were found to be related to measures of youth adjustment both concurrently and longitudinally, independent of their associations with ratings of global self-esteem. Prospective analyses in each study further revealed that profile type predicted differential change in measures of youth adjustment over time, whereas this type of relationship was not found for global ratings of self esteem. Implications for esteem-enhancement interventions with youth are discussed. PMID- 10723540 TI - The reactions of myoglobin, normal adult hemoglobin, sickle cell hemoglobin and hemin with hydroxyurea. AB - The kinetics of the reaction of hydroxyurea (HU) with myoglobin (Mb), hemin, sickle cell hemoglobin (HbS), and normal adult hemoglobin (HbA) were determined using optical absorption spectroscopy as a function of time, wavelength, and temperature. Each reaction appeared to follow pseudo-first order kinetics. Electron paramagnetic resonance spectroscopy (EPR) experiments indicated that each reaction produced an FeNO product. Reactions of hemin and the ferric forms of HbA, HbS, and myoglobin with HU also formed the NO adduct. The formation of methemoglobin and nitric oxide-hemoglobin from these reactions may provide further insight into the mechanism of how HU benefits sickle cell patients. PMID- 10723541 TI - Backbone cluster identification in proteins by a graph theoretical method. AB - A graph theoretical algorithm has been developed to identify backbone clusters of residues in proteins. The identified clusters show protein sites with the highest degree of interactions. An adjacency matrix is constructed from the non-bonded connectivity information in proteins. The diagonalization of such a matrix yields eigenvalues and eigenvectors, which contain the information on clusters. In graph theory, distinct clusters can be obtained from the second lowest eigenvector components of the matrix. However, in an interconnected graph, all the points appear as one single cluster. We have developed a method of identifying highly interacting centers (clusters) in proteins by truncating the vector components of high eigenvalues. This paper presents in detail the method adopted for identifying backbone clusters and the application of the algorithm to families of proteins like RNase-A and globin. The objective of this study was to show the efficiency of the algorithm as well as to detect conserved or similar backbone packing regions in a particular protein family. Three clusters in topologically similar regions in the case of the RNase-A family and three clusters around the porphyrin ring in the globin family were observed. The predicted clusters are consistent with the features of the family of proteins such as the topology and packing density. The method can be applied to problems such as identification of domains and recognition of structural similarities in proteins. PMID- 10723542 TI - A calorimetric study of the influence of calcium on the stability of bovine alpha lactalbumin. AB - Bovine alpha-lactalbumin has been studied by differential scanning calorimetry with various concentrations of calcium to elucidate the effect of this ligand on its thermal properties. In the presence of excess calcium, alpha-lactalbumin unfolds upon heating with a single heat-absorption peak and a significant increase of heat capacity. Analysis of the observed heat effect shows that this temperature-induced process closely approximates a two-state transition. The transition temperature increases in proportion with the logarithm of the calcium concentration, which results in an increase in the transition enthalpy as expected from the observed heat capacity increment of denaturation. As the total concentration of free calcium in solution is decreased below that of the proteins, there are two temperature-induced heat absorption peaks whose relative area depends on the calcium concentration, such that further decrease of calcium concentration results in a increase of the low-temperature peak and a decrease of the high-temperature one. The high-temperature peak occurs at the same temperature as the unfolding of the holo-protein, while the low-temperature peak is within the temperature range associated with the unfolding of the apo-protein. Statistical thermodynamic modeling of this process shows that the bimodal character of the thermal denaturation of bovine alpha-lactalbumin at non saturated calcium concentrations is due to a high affinity of Ca2+ for alpha lactalbumin and a low rate of calcium exchange between the holo- and apo-forms of this protein. Using calorimetric data, the calcium-binding constant for alpha lactalbumin has been determined to be 2.9 x 10(8) M-1. PMID- 10723543 TI - Ultrasonic surfactant nebulization with different exciting frequencies. AB - Intratracheal bolus instillation of natural lung surfactant is the treatment of choice in neonatal respiratory distress syndrome and an increasing part in adults' therapy. For reasons of hemodynamics, surfactant distribution and efficiency the application mode should be improved. Nebulization seems to have some advantages but its technical realization is difficult. The aim of the present study was to investigate if ultrasonic nebulization with exciting frequencies higher than 2.8 MHz can improve the efficiency of surfactant nebulization without changing the surface-active properties of the material. Exciting frequencies of 1.7, 3.3 and 4.0 MHz were used to produce a surfactant aerosol. The phospholipid content in the liquefied aerosol and particle size distinctly dropped with higher frequencies. The surface activity was not altered in the produced aerosol and neither in the surfactant remaining in the nebulizer. Although possible, ultrasonic nebulization of surfactant suspensions is ineffective because of a striking decrease in phospholipid content. PMID- 10723544 TI - Thermodynamics of reactions catalyzed by anthranilate synthase. AB - Microcalorimetry and high performance liquid chromatography have been used to conduct a thermodynamic investigation of reactions catalyzed by anthranilate synthase, the enzyme located at the first step in the biosynthetic pathway leading from chorismate to tryptophan. One of the overall biochemical reactions catalyzed by anthranilate synthase is: chorismate(aq) + ammonia(aq) = anthranilate(aq) + pyruvate(aq) + H2O(l). This reaction can be divided into two partial reactions involving the intermediate 2-amino-4-deoxyisochorismate (ADIC): chorismate(aq) + ammonia(aq) = ADIC(aq) + H2O(l) and ADIC(aq) = anthranilate(aq) + pyruvate(aq). The native anthranilate synthase and a mutant form of it that is deficient in ADIC lyase activity but has ADIC synthase activity were used to study the overall ammonia-dependent reaction and the first of the above two partial reactions, respectively. Microcalorimetric measurements were performed on the overall reaction at a temperature of 298.15 K and pH 7.79. Equilibrium measurements were performed on the first partial (ADIC synthase) reaction at temperatures ranging from 288.15 to 302.65 K, and at pH values from 7.76 to 8.08. The results of the equilibrium and calorimetric measurements were analyzed in terms of a chemical equilibrium model that accounts for the multiplicity of ionic states of the reactants and products. These calculations gave thermodynamic quantities at the temperature 298.15 K and an ionic strength of zero for chemical reference reactions involving specific ionic forms. For the reaction: chorismate2 (aq) + NH4+(aq) = anthranilate-(aq) + pyruvate-(aq) + H+(aq) + H2O(l), delta rHmo = -(116.3 +/- 5.4) kJ mol-1. For the reaction: chorismate2-(aq) + NH4+(aq) = ADIC (aq) + H2O(l), K = (20.3 +/- 4.5) and delta rHmo = (7.5 +/- 0.6) kJ mol-1. Thermodynamic cycle calculations were used to calculate thermodynamic quantities for three additional reactions that are pertinent to this branch point of the chorismate pathway. The quantities obtained in this study permit the calculation of the position of equilibrium of these reactions as a function of temperature, pH, and ionic strength. Values of the apparent equilibrium constants and the standard transformed Gibbs energy changes delta rG'mo under approximately physiological conditions are given. PMID- 10723545 TI - Kinetic study of the oxidation of 3-hydroxyanisole catalysed by tyrosinase. AB - Tyrosinase hydroxylates 3-hydroxyanisole in the 4-position. The reaction product accumulates in the reaction medium with a lag time (tau) which diminishes with increasing concentrations of enzyme and lengthens with increasing concentrations of substrate, thus fulfilling all the predictions of the mechanism proposed by us for 4-hydroxyphenols. The kinetic constants obtained, kcatM = (46.87 +/- 2.06) s 1 and KmM = (5.40 +/- 0.60) mM, are different from those obtained with 4 hydroxyanisole, kcatM = (184.20 +/- 6.1) s-1 and KmM = (0.08 +/- 0.004) mM. The catalytic efficiency, kcatM/KmM is, therefore, 265.3 times greater with 4 hydroxyanisole. The possible rate-determining steps for the reaction mechanism of tyrosinase on 3- and 4-hydroxyanisole, based on the NMR spectra of both monophenols, are discussed. These possible rate-determining steps are the nucleophilic attack of hydroxyl's oxygen on the copper and the electrophilic attack of the peroxide on the aromatic ring. Both steps may be of similar magnitude, i.e. take place in the same time scale. PMID- 10723546 TI - Thermodynamical model of mixed aggregation of ligands with caffeine in aqueous solution. Part II. AB - A statistical-thermodynamical model of mixed association in which one component's self-association is unlimited while the second component does not self-aggregate is described. The model was tested with 4',6-diamidino-2-phenyl-indole dihydrochloride (DAPI) and ethidium bromide (EB) using light absorption spectroscopy and calorimetry. The system is controlled by two parameters, which represent self-aggregation 'neighborhood' association constant KCC and mixed 'neighborhood' association constant KAC. Calculated, using this model, KAC = 58.2 +/- 1 M-1, KAC = 64.6 +/- 2 M-1 for DAPI and EB, respectively, are in good agreement with known values of stacking interactions. The titration microcalorimetric measurement of DAPI-CAF interaction delta H = -11.1 +/- 0.4 kcal/mol is also consistent with this type of reaction. The structures of the stacking complexes were also confirmed by semi-empirical molecular modeling in the presence of water. The data indicate that CAF forms stacking complexes with DAPI and EB, thus effectively lowering the concentration of the free ligands in the solution, and therefore, CAF can be used to modulate aromatic compound activity. PMID- 10723547 TI - Correcting for different amplification rates of internal standard and template when measuring cDNA by competitive PCR. PMID- 10723548 TI - High copy number plasmids compatible with commonly used cloning vectors. PMID- 10723550 TI - Size-fractionation of RNA by hot agarose electrophoresis. PMID- 10723551 TI - Improved transfection efficiency of 293 cells by radio frequency electroporation. PMID- 10723552 TI - Making colony PCR easier by adding gel-loading buffer to the amplification reaction. PMID- 10723549 TI - Quantifying adenoviral titers by spectrophotometry. PMID- 10723553 TI - Heating greatly speeds Coomassie blue staining and destaining. PMID- 10723554 TI - Modified CTAB protocol using a silica matrix for isolation of plant genomic DNA. PMID- 10723555 TI - BLAST on the Web. PMID- 10723556 TI - Optimized rapid amplification of cDNA ends (RACE) for mapping bacterial mRNA transcripts. AB - A simple, efficient and sensitive RACE-based procedure was developed for the determination of unknown 5' regions from bacterial cDNA. A number of critical modifications were made to the standard RACE method, including the optimization of the RNA extraction, reverse transcription and PCR conditions. This procedure was used to accurately determine the site of transcript initiation and structure of the promoter region of the Helicobacter pylori aspartate carbamoyltransferase gene (pyrB). The technique avoids many of the difficulties associated with established bacterial transcript mapping protocols and can be performed in two days starting with less than 1 microgram of total RNA. The modifications described here have significant potential for the identification of transcript start sites of bacterial genes and non-polyadenylated eukaryotic RNA. PMID- 10723557 TI - Monitoring and purification of proteins using bovine papillomavirus E2 epitope tags. AB - We describe here the use of two newly mapped bovine papillomavirus type 1 (BPV-1) E2 protein epitopes as tags. We constructed several vector plasmids for overexpression as well as for moderate expression of single- or double-tagged proteins in either Escherichia coli or eukaryotic cells. The new tags were fused to several proteins, and the activity of the tagged proteins was tested in different assays. The tags were shown not to interfere with the function of these proteins in vivo and in vitro. Interaction of the monoclonal antibodies 3F12 and 1E2 with their respective epitopes was specific and had high affinity in a variety of conditions. We have demonstrated that the 3F12 antibody-epitope interaction tolerates high salt concentrations up to 2 M. This permits immunoprecipitation and immunopurification of the tagged proteins in high-salt buffers and reduction of the nonspecific binding of the contaminating proteins. We also provide a protocol for DNA binding and DNase I footprinting assays using the tagged, resin-bound DNA-binding proteins. The BPV-1 E2-derived tags can be recommended as useful tools for detection and purification of proteins. PMID- 10723558 TI - Determining SNP allele frequencies in DNA pools. AB - Single nucleotide polymorphisms (SNPs) are among the most common types of polymorphism used for genetic association studies. A method to allow the accurate quantitation of their allele frequencies from DNA pools would both increase throughput and decrease costs for large-scale genotyping. However, to date, most DNA pooling studies have concentrated on the use of microsatellite polymorphisms. In the case of SNPs that are restriction fragment length polymorphisms (RFLPs), studies have tended to use methods for the quantitation of allele frequency from pools that rely on densitometric evaluation of bands on an autoradiograph. Radiation-based methods have well-known drawbacks, and we present two alternative methods for the determination of SNP allele frequencies. For RFLPs, we used agarose gel electrophoresis of digested PCR products with ethidium bromide staining combined with densitometric analysis of gel images on a PC. For all types of SNP, we used allele-specific fluorescent probes in the Taqman assay to determine the relative frequencies of two different alleles. Both methods gave accurate and reproducible results, suggesting they are suitable for use in DNA pooling experiments. PMID- 10723559 TI - Use of fluorescent microspheres to localize in vivo gene transfer injection sites. AB - The potential for using gene therapy to treat a variety of disease states is growing rapidly. Many vector types and delivery systems have been developed that allow the optimization of protein production levels and kinetics for a given therapeutic gene product. In cases in which a transient, localized delivery of gene product is desired, any determination of the locale of transfected tissue by non-marker genes is problematic. We describe a technique by which the use of fluorescent microspheres can help in identifying potentially transfected tissue. Adenovirus containing the gene for beta-galactosidase (beta-gal) was mixed with fluorescent microspheres and injected into rat skeletal muscle and porcine myocardium. The injection sites could be visualized under ultraviolet light and correlated with beta-gal enzyme expression. This method is simple, inexpensive and generally useful for in vivo gene transfer experiments. PMID- 10723560 TI - Detection and molecular mass determination of an HIV replication-enhancing female genital tract factor using a blot bioassay. AB - We previously reported that cervicovaginal lavages (CVL) contain a factor that enhances the replication of human immunodeficiency virus (HIV) by increasing virus transcription in T cells and monocytoid cells. This factor was named the HIV-inducing factor (HIF). To determine the molecular mass of HIF, we adapted a blot technique that involves nonreducing SDS-PAGE of CVL samples and electrophoretic transfer onto nitrocellulose paper followed by incubation of paper slices with HIV-infected monocytoid U1 cells. The slices with HIF bioactivity were detected by increased HIV production and measured by an HIV core protein (p24) ELISA. We refer to this technique as the "BioBlot" assay. The BioBlot assay successfully detected bioactivity of HIF anchored onto nitrocellulose and determined that HIF has a molecular mass of about 14 kDa. Paper slices with HIF-negative CVL samples as well as nitrocellulose paper samples without CVL did not enhance HIV production. This finding suggested that SDS-PAGE and nitrocellulose binding do not functionally alter the bioactive domain(s) of HIF structure. In addition to the detection of HIF bioactivity anchored to nitrocellulose and HIF molecular mass determination, the BioBlot technique offers an alternative, rapid method for other applications. These include the study of receptor-ligand interactions of mucosal proteins, direct bioactivity testing and molecular mass determination of secretory substances. PMID- 10723561 TI - Preparation of sensitive and specific oligonucleotide probes tailed using terminal transferase and dITP. AB - An oligonucleotide probe tailed with deoxyadenosine-5'-triphosphate or deoxythymine-5'-triphosphate is detectable with high sensitivity, but has a major drawback--the tail co-hybridizes specifically to complementary sequences. This can be a problem when screening cDNA clones that contain poly(dA) sequences. While it is possible to mask the cDNA tail with unlabeled poly(dA) or poly(A) oligonucleotides, false-positive clones are still produced because complete masking of extremely long (dA) tails is difficult. As a result, only cDNA clones that have extremely long poly(dA) sequences are often obtained by hybridization screening using tailed probes. In this report, we describe an oligonucleotide probe tailed with DIG-labeled nucleotide in combination with deoxyinosine-5' triphosphate that was highly specific and sensitive to cDNAs. Terminal deoxynucleotidyl transferase efficiently adds dI nucleotides to the 3'-end. The dI of the tails did not pair with any nucleotides under stringent hybridization so that the specificity of hybridization assays remained high without affecting the sensitivity of the test. Colony hybridization experiments demonstrated that there were very few (1 of 80 tested) false positives using this technique. Its use may increase the accuracy of cDNA screening. PMID- 10723562 TI - Protein microdeposition using a conventional ink-jet printer. AB - Many recent bioanalytical systems based on immunologic and hybridization reactions in a mono- or bidimensional microarray format require technology that can produce arrays of spots containing biospecific molecules. Some microarray deposition instruments are commercially available, and other devices have been described in recent papers. We describe a system obtained by adapting a commercial ink-jet printer and used to produce mono- and bidimensional arrays of spots containing horseradish peroxidase on cellulose paper. In a few minutes, it was possible to obtain bidimensional arrays containing several thousands of spots with a diameter as low as 0.2 mm, with each of which requiring only a few nanoliters of the enzyme deposition solution. The quantity of enzyme in each spot was evaluated with a chemiluminescent reaction and a charge-coupled device-based, low-light imaging luminograph. The chemiluminescence measurements revealed that the reproducibility of the enzyme deposition was satisfactory for analytical purposes, with the variation coefficients being lower than 10% in almost all cases. PMID- 10723563 TI - Restriction site-free insertion of PCR products directionally into vectors. AB - A restriction site-free cloning method has been developed for inserting a PCR product into a vector flexibly and precisely at any desired location with high efficiency. The method uses a pair of DNA integration primers with two portions. The 3' portion isolates the inserts by PCR, and the 5' portion integrates the PCR products into the homologous region of the vector. For mutagenesis, a third portion of mutation-generating sequences can be placed in between the 3' and 5' portions. This method has been used to clone the E. coli gene that codes for peptidyl-tRNA hydrolase, expressing it as a native protein and as a glutathione S transferase fusion protein. It was also applied to convert a construct of the E. coli fatty acid biosynthesis protein with an N-terminal hexa-histidine tag into a construct with a C-terminal hexa-histidine tag. PMID- 10723564 TI - Cryptic enhancer elements in luciferase reporter vectors respond to the osteoblast-specific transcription factor Osf2/Cbfa1. AB - Luciferase reporter vectors are commonly used for the functional analysis of basal promoter and enhancer elements of eukaryotic genes. Randomly occurring cisacting elements in the vector sequences that can spuriously respond to various transcription factors, combined with the high sensitivity of the luciferase assay system, could make these vectors unsuitable for functional studies with certain transcription factors. Here, we provide evidence that pGL2-Basic and pGL3-Basic are transactivated by the osteoblast-specific transcription factor Cbfa1 and estrogen receptor alpha probably through randomly occurring cisacting elements in the vector sequences. Our results highlight the limitations of pGL2-Basic and pGL3-Basic vectors in promoter transactivation/repression studies. The results also emphasize the need to perform appropriate controls and test the expression levels with a particular transcription factor and promoterless luciferase reporter vector combination. PMID- 10723565 TI - Polycationic lipids translocate lipopolysaccharide into HeLa cells. AB - We have investigated the ability of LIPOFECTAMINE, a polycationic lipid reagent used in DNA transfection, to translocate E. coli lipopolysaccharide (LPS) into HeLa cells. Although HeLa cells did not spontaneously take up fluorescein isothiocyanate-labelled LPS (FITC-LPS) from the culture medium, the cells that were co-incubated with greater than 1 g/mL FITC-LPS and LIPOFECTAMINE showed punctate fluorescence. Virtually all cells were loaded on incubation with 100 micrograms/mL FITC-LPS. Confocal scanning laser microscopy showed extensive FITC LPS loading in the cytoplasm of HeLa cells, but no label was evident in the nuclear regions of these cells. Loading with LPS for up to six hours had no effect on the viability of HeLa cells, beyond the 30% reduction in live cells that is attributable to the toxic effect of LIPOFECTAMINE itself. In contrast to cells treated with etoposide for six hours, LPS-loaded cells did not display apoptotic bodies. Exposure of cells to 4 beta-phorbol 12-myristate 13-acetate led to the induction of the immediate early gene c-fos and resulted in an enhanced c Fos signal, detected by Western blot analysis. In contrast, LPS loading did not alter the c-fos expression in HeLa cells. The loading of LPS into HeLa cells by means of polycationic lipids results in relatively low acute toxicity, as judged from cell viability, morphology and c-fos expression. Therefore, our method appears well suited to the study of acute actions of LPS in the intracellular compartment of mammalian cells. PMID- 10723566 TI - Simple method for preparation of fluor/hapten-labeled dUTP. AB - Many projects, such as multiplex-fluorescence in situ hybridization (M-FISH) karyotyping, require the use of relatively large amounts of multiple fluor- or hapten-labeled nucleotides for the preparation of DNA probes. Such a requirement makes these experimental approaches prohibitively expensive for many researchers. The cost of such nucleotides can be reduced approximately 99% by purchasing the chemical precursors, fluor or hapten succinimidyl esters and 5-(3-aminoallyl)-2' deoxyuridine 5' triphosphate (AA-dUTP), and performing the simple coupling/purification described here. It is possible to finish four to ten different fluor/hapten dUTP preparations of 2.5 microM scale within a 24 h period. The reagent cost for each preparation ranges from $33-$237 per microM, depending on the fluor/hapten. This laboratory uses such nucleotide preparations to prepare FISH probes by nick translation or PCR amplification. PMID- 10723567 TI - Assessment of RNA quality by semi-quantitative RT-PCR of multiple regions of a long ubiquitous mRNA. AB - A simple method to assess the degree of degradation present in a total RNA preparation from cells or tissues is based on the increasing probability of RNA cleavage with increasing length of an RNA molecule. Under ideal conditions, reverse transcription of a particular mRNA species with oligo-dT as the primer generates a population of cDNAs, terminating at the 5' end of the mRNA if all template RNA molecules are intact, or at the first cleavage site 5' to the polyA if some template RNAs are partially degraded. Consequently, for cellular RNA preparations with some degradation, the 5' end of an mRNA is represented in the cDNA population to a lesser extent than the 3' end of the mRNA. We describe a sensitive assay of mRNA quality that compares the relative PCR amplification of 5' and 3' regions of a long and ubiquitous mRNA following oligo-dT-primed reverse transcription. PMID- 10723568 TI - Mutagenesis and gene cloning in Schizosaccharomyces pombe using nonhomologous plasmid integration and rescue. AB - Genes are commonly cloned in yeasts and bacteria by plasmid complementation, where the introduction of the gene of interest into a host strain carrying a recessive mutation in that gene suppresses the host's mutant phenotype. However, a lack of low copy cloning vectors in the fission yeast Schizosaccharomyces pombe can complicate this approach especially when overexpression of one gene may suppress a defect in another gene or when overexpression of the desired gene is detrimental, if not lethal, to the cell. We describe here a method of identifying mutations in S. pombe that allows for the rapid and direct cloning of the defective gene. This involves the nonhomologous integration of a marked plasmid into the yeast genome and its subsequent rescue into Escherichia coli, so that DNA at the site of insertion is incorporated into the recovered plasmid. As two of three insertions obtained in this study occurred outside of the affected gene's open reading frame, this method should be applicable to cloning both essential genes and nonessential genes. PMID- 10723569 TI - Optical aberrations and objective choice in multicolor confocal microscopy. AB - Refinements in design have simplified confocal microscopy to the extent that it has become a standard research tool in cell biology. However, as confocal microscopes have become more powerful, they have also become more demanding of their optical components. In fact, optical aberrations that cause subtle defects in image quality in wide-field microscopy can have devastating effects in confocal microscopy. Unfortunately, the exacting optical requirements of confocal microscopy are often hidden by the optical system that guarantees a sharp image, even when the microscope is performing poorly. Optics manufacturers provide a wide range of microscope objectives, each designed for specific applications. This report demonstrates how the trade-offs involved in objective design can affect confocal microscopy. PMID- 10723570 TI - GCN4-based expression system (pGES): translationally regulated yeast expression vectors. AB - The expression of foreign proteins in Saccharomyces cerevisiae is a powerful tool for basic research and the biotechnological industry. In spite of the potential of S. cerevisiae, only a few useful expression vectors have been developed for this yeast. These vectors are based on an increasing transcription rate in combination with an increase in gene dosage. Most vectors are maintained as plasmids, which forces growth of cultures on poor selective media. Expression of the yeast Gcn4 protein is regulated at the translational level and increases strongly under amino acid starvation. Because under these conditions protein synthesis in general ceases, it is conceivable that regulatory elements that control Gcn4 expression could support selective expression of foreign genes. We cloned DNA fragments residing upstream from the GCN4 coding sequence (including the 5' UTR) and ligated them to a cDNA that encodes the human serum albumin (HSA) gene. These GCN4 regulatory elements induced efficient HSA expression at the translational level under amino acid starvation. The GCN4/HSA cassette promoted efficient, inducible expression on either a multicopy or integrative plasmid. The integrated cassette induced a high level of HSA in dense cultures grown on rich media. Thus, the GCN4-based expression system (pGES) provides high protein quantities. pGES is the first expression vector to be induced at the translational level. PMID- 10723571 TI - Restriction mapping of retroviral vector episomal DNA. PMID- 10723572 TI - Routine detection of the replication error phenotype in clinical tumor specimens using fluorescence-SSCP. AB - Almost all tumors from patients with hereditary non-polyposis colon carcinoma and approximately 10%-15% of sporadic colon and gastric carcinomas contain widespread deletions within mono- and dinucleotide repeat sequences in their DNA. This is referred to as the replication error (RER+) phenotype and, in the case of colon carcinoma, is often associated with an improved tumor prognosis and possibly also with response to chemotherapy. The RER+ status of tumors is usually determined by examining several dinucleotide and mononucleotide repeats for size variations when compared with the matching normal DNA. Alternately, the identification of deletions within BAT-26, a quasi-monomorphic polyadenine tract within the hMSH2 gene, has been shown to establish the RER+ status of tumors with greater than 99% accuracy and without the need for normal DNA. Here, we use fluorescent PCR in combination with single strand conformation polymorphism analysis to detect deletions in BAT-26. This technique provides a sensitive, rapid, reproducible and inexpensive assay suitable for the routine identification of RER+ status in clinical tumor specimens. PMID- 10723573 TI - Cloning and characterization of a retroviral plasmid, pCC1, for combination suicide gene therapy. AB - Introduction of the herpes simplex virus thymidine kinase (HSV-TK) gene into mammalian cells confers specific sensitivity to killing by the anti-herpes drug ganciclovir (GCV). This gene has therefore been used in a number of cancer gene therapy protocols as a therapeutic gene. However, the therapeutic efficacy of HSV TK/GCV in cancer gene therapy experiments can be augmented by additional therapeutic genes. We have cloned a retroviral plasmid, pCC1, containing a fusion gene of HSV-TK and Sh-ble driven by an internal simian virus 40 early promoter. This gene encodes a fusion protein that confers GCV sensitivity and Zeocin resistance when introduced into mammalian cells. A multiple cloning site (MCS) allows the introduction of a second therapeutic gene under the transcriptional control of the Moloney murine leukemia virus 5' long terminal repeat. We have generated packaging cell lines electroporated with pCC1 or pCC1 rtIL-2 S (rat interleukin-2 gene cloned in the sense direction in the MCS), the supernatants of which transfer GCV sensitivity only, or both GCV sensitivity and rtIL-2 production, respectively to rat ovarian cancer cells. This plasmid may be useful for the study of combination suicide gene therapy strategies. PMID- 10723574 TI - IPTG-inducible episomal expression system for exogenous genes in primate cells. AB - An isopropyl beta-D-thiogalactopyranoside (IPTG)-inducible episomal expression system has been established for the human breast carcinoma cell line MCF7. This two-component system includes: (i) a primate cell-specific episomal vector, pEpiLac, that contains an IPTG-inducible promoter and (ii) a cell line derived from MCF7, MCF7/LAP5, which expresses the IPTG-dependent transactivator LAP267. Treatment of MCF7/LAP5 cells with IPTG results in efficient inducible expression of exogenous genes from the inducible promoter in pEpiLac. Up to 300-fold induction can be observed when luciferase is used as a reporter. Inducible expression of the p27KIP1 cyclin-dependent kinase (CDK) inhibitor, the orphan nuclear receptor Nur77 and the angiogenic inducer Cyr61 has also been demonstrated. Expression of the exogenous gene is promptly halted on removal of IPTG. Moreover, the episomal vector can be stably maintained in and easily recovered from MCF7/LAP5 cells. Taken together, this inducible expression system should be applicable for the regulated expression of exogenous genes, especially growth inhibitory or cytotoxic genes, in cells of primate origin. PMID- 10723575 TI - Mini-allografts: ongoing trials in humans. AB - Conventional allogeneic stem cell transplantation is a valuable approach to therapy for many hematologic malignancies. However, high-dose conditioning regimens designed both to control the malignancy and to prevent graft rejection are associated with a high incidence of acute and long-term side-effects. This has largely precluded the use of allografting for patients older than 55 years or for younger patients with certain pre-existing organ damage. In order to manage the side-effects, transplants have traditionally been delivered in highly specialized hospital wards or intensive care settings. Thus, an important goal is to develop safer allografting procedures that can be extended to older patients or patients with pre-existing organ dysfunction who are currently excluded from consideration for transplant. Recent observations have shown that donor lymphocyte infusions (DLI) can eradicate some malignancies that relapse after conventional allografting. These observations confirmed earlier evidence in favor of a graft-versus-leukemia effect based on the association of graft-versus-host disease (GVHD) with a lower likelihood of relapse of malignancy after allografting. Given the potential efficacy of DLI as the sole modality for eradication of malignancy, new strategies for allografting can incorporate the concept of less intensive conditioning therapy which is given with the sole aim of facilitating allogeneic engraftment. Recent pre-clinical studies in a canine model have shown that conditioning regimens for allografting can be markedly reduced in intensity yet still achieve the goal of engraftment. This review briefly summarizes the initial translational clinical studies, using a minimally myelosuppressive-conditioning regimen based on low dose total body irradiation (TBI) or fludarabine alone or in combination with other drugs followed by a short course of immunosuppression with post-grafting cyclosporine and methotrexate or mycophenolate mofetil. PMID- 10723576 TI - Stem cell transplantation for treatment of severe autoimmune diseases: current status and future perspectives. AB - Autoimmune diseases include a heterogeneous group of disorders with variable presentation and severity. Immunosuppressive and immunomodulatory therapies are often used for treatment with considerable success in some cases. These diseases may also be severe and refractory to conventional treatment. Thus more aggressive intervention might be indicated in a subset of patients. Animal studies suggest that high-dose therapy supported by stem cell transplantation may lead to remissions in experimental autoimmune disease models. Anecdotal case reports suggest that the same may be the case in some human autoimmune diseases as well. This review attempts to summarise some current concepts and future perspectives on stem cell transplantation in the treatment of severe autoimmune diseases. PMID- 10723577 TI - Conditioning regimens for the treatment of experimental arthritis with autologous bone marrow transplantation. AB - Adjuvant induced arthritis (AA) in Buffalo rats is a chronic progressive disease that responds very well to treatment with myeloablation and rescue with autologous BM. These previous results were obtained by conditioning with a lethal single dose of TBI. In the present study various other conditioning regimens were compared with single dose TBI. Fractionated TBI using adjusted total dose was equally effective. CY and BU when used as single agents at the highest tolerated dose were less effective. Combinations of CY (2 x 60 mg/kg) with lower dose (4 Gy) TBI and of BU with CY were as beneficial as high-dose TBI. These results indicate that a very intense reduction of T lymphocytes, in the order of 3-4 log, is required for obtaining the highest rate of long-lasting complete remissions. A similar conclusion was reached from our studies of various conditioning regimens in rats suffering from experimental allergic encephalitis (EAE). If extrapolated to the clinic, such a degree of T lymphocyte eradication poses upper limits to the number of T lymphocytes that can be safely reintroduced with the autograft. The exact limits cannot be derived from these experiments because the addition of autologous T lymphocytes to the graft yielded different results in the two models of autoimmune disease (AID). PMID- 10723578 TI - Melphalan/TBI is not more carcinogeneic than cyclophosphamide/TBI for transplant conditioning: follow-up of 725 patients from a single centre over a period of 26 years. AB - As there is concern regarding the high carcinogenic potential of melphalan (Mel), 725 patients with haematological malignancies who received allogeneic (n = 714) or syngeneic (n = 11) transplants over the last 26 years were followed-up to evaluate if melphalan was more likely to result in secondary malignant neoplasms (SMNs) than cyclophosphamide (Cy). Three hundred and ninety-five were treated with Cy/TBI and 330 with Mel/TBI. Twelve patients developed non-haematological SMN. Median time to develop a SMN was 7 years (range 2-17 years). Age-adjusted rate was significantly higher than in the general population (observed 12 expected 1.2, risk 10; P < 0.0001). The cumulative overall risk of developing a SMN at 2, 5, 10 and 15 years post transplant was 0.4% (95% CI 0.1-2.6%), 1.7% (95% CI 0.6-4.4%), 6.4% (95% CI 2.8-10.8%) and 6.6% (95% CI 3.4-12.4%), respectively. Even though age-adjusted rates were higher than the general population melphalan/TBI was not associated with higher age-adjusted risk than Cy/TBI (increased risk 7.9 vs 11.4; P = NS). The cumulative overall risk of SMNs was not different with CY/TBI or Mel/TBI (8/393 vs 4/363; 10 year risk 4.4%, 95% CI 1.8-10.6 vs 8.4%, 95% CI 2.9-22.9; P = NS). The risk was significantly higher with use of additional cranial or cranio-spinal irradiation (17.5% vs 2.7% at 10 years; P = 0.0241). Transplants for acute lymphatic leukaemia resulted in a higher incidence of SMNs than did transplants for other diseases (ALL: 17.4%, 95% CI 6.3-42.6%; other diseases: 3.4% (95% 1.3-8.5%, P = 0.0469). The risk of SMN for patients with chronic GVHD was 8.4% (95% CI 3.7-18.7%) as compared to 3.5% (95% CI 1-11.1%) for patients without chronic GVHD (P = NS). No factor was associated with independently increased risk in multivariate analysis. Use of melphalan and TBI for transplant conditioning does not appear to be associated with higher risk of second malignant neoplasms than cyclophosphamide and TBI. PMID- 10723579 TI - Lenograstim-mobilized peripheral blood progenitor cells in volunteer donors: an open label randomized split dose escalating study. AB - Mobilization of peripheral blood cell progenitor cells was investigated in 36 healthy sibling donors using three different split doses of glycosylated rhG-CSF (lenograstim). The donors were randomized into three groups: group 1 was given lenograstim at 8, group 2 at 11 and group 3 at 15 micrograms/kg/day in two split doses, subcutaneously for 4 and 5 days, respectively. Leukapheresis was performed on day 4 or 5 depending on the WBC and CD34+ cell count. We were able to demonstrate that there was a significant correlation between circulating CD34+ cells on the day of harvest and CD34+ cells in the apheresis products in all three groups. The number of CD34+ cells pre-apheresis was inversely correlated with age in group 1 and group 2. However, in group 3, the number of CD34+ cells pre-apheresis did not correlate with age. There was also a difference between the number of progenitor cells mobilized in the three dose groups regarding the time of harvest. Apheresis was performed in groups 1 and 2 on day 5 of mobilization in order to obtain a sufficient number of stem cells for allogeneic transplantation. In contrast, with the split dose of 15 micrograms/kg/day, harvest could be routinely performed on day 4 of stimulation. We conclude that lenograstim given twice a day at doses of 8, 11 and 15 micrograms/kg/day provided different CD34+ cell yields in normal donors, in particular, with regard to the time of harvest. The number of CD34+ cells pre-apheresis was not correlated with age in the group of donors mobilized with a split dose of 15 micrograms/kg/day, indicating that this dosage might also be suitable for older donors. PMID- 10723580 TI - Characterization of CD34+ subsets derived from bone marrow, umbilical cord blood and mobilized peripheral blood after stem cell factor and interleukin 3 stimulation. AB - We characterized CD34+ cells purified from bone marrow (BM), mobilized peripheral blood (PB) and cord blood (CB) and we tried to establish correlations between the cell cycle kinetics of the CD34+CD38- and CD34+CD38+ subpopulations, their sensitivity to SCF and IL-3 and their expression of receptors for these two CSFs. At day 0, significantly fewer immature CD34+CD38- cells from CB and mobilized PB are in S + G2M phases of the cell cycle (respectively 2.0 +/- 0.4 and 0.9 +/- 0.3%) than their BM counterpart (5.6 +/- 1.2%). A 48-h incubation with SCF + IL-3 allows a significant increase in the percentage of cycling CD34+CD38- cells in CB (19.2 +/- 2.2%, P < 0.0002) and PB (14.1 +/- 5.5%, P < 0.05) while the proliferative potential of BM CD34+CD38- progenitors remains constant (8.6 +/- 1.0%, NS). CD123 (IL-3 receptor) expression is similar in the three sources of hematopoietic cells at day 0 and after 48-h culture. CD117 (SCF receptor) expression, although very heterogeneous according to the subpopulations and the sources of progenitors evaluated, seems not to correlate with the difference of progenitor cell sensitivity to SCF nor with their proliferative capacity. Considering the importance of the c-kit/SCF complex in the adhesion of stem cells to the microenvironment, several observations are relevant. The density of CD117 antigen expression (expressed in terms of mean equivalent soluble fluorescence, MESF) is significantly lower on fresh PB cells than on their BM (P < 0.017) and CB (P < 0.004) counterparts, particularly in the immature CD34+CD38- population (560 +/- 131, 2121 +/- 416 and 1192 +/- 129 MESF respectively); moreover, when PB and BM CD34+CD38- cells are stimulated for 48 h with SCF + IL-3, the CD117 expression decreases by 1.5- and 1.66-fold, respectively. This reduction could modify the functional capacities of ex vivo PB and BM manipulated immature progenitor cells. PMID- 10723581 TI - The extent of HLA class II allele level disparity in unrelated bone marrow transplantation: analysis of 1259 National Marrow Donor Program donor-recipient pairs. AB - A comprehensive analysis of the HLA-D region loci, DRB1, DRB3, DRB5, DQA1, DQB1, DPA1 and DPB1, was performed to determine allelic diversity and underlying HLA disparity in 1259 bone marrow recipients and their unrelated donors transplanted through the National Marrow Donor Program. Although 43.0% of DRB1 alleles known to exist at the beginning of the study were found in this predominantly Caucasian transplant population, a few alleles predominated at each locus. In recipients, 67.1% of DRB1 alleles identified were one or two of six common DRB1 alleles. Only 118 (9.4%) donor-recipient pairs were matched for all alleles of DRB1, DQA1, DQB1, DPA1 and DPB1. While 79.4% of the pairs were matched for DRB1, only 13.2% were matched for DPB1 alleles. Almost 66% of pairs differed by more than one allele mismatch and 59.0% differed at more than one HLA-D locus. DQB1 was matched in 85.9% of DRB1-matched pairs. In contrast, only 13.9% of the pairs matched for DRB1, DQA1 and DQB1 were also matched for DPA1 and DPB1. This database, highlighting the underlying HLA disparity within the pairs, forms the foundation of an ongoing study to establish the relationship between HLA matching and successful outcome in unrelated allogeneic stem cell transplant. PMID- 10723582 TI - Pediatric marrow transplantation for acute leukemia using unrelated donors and T replete or -depleted grafts: a case-matched analysis. AB - A retrospective, case-matched analysis of the short-term toxicity, risk of GVHD and relapse as well as outcome in pediatric unrelated marrow transplantation was conducted by comparing recipients of T-replete and -depleted grafts in a two center setting. Both groups contained 30 patients with acute leukemia matched by age at transplant, gender, primary diagnosis and disease status. Acute (90% vs 53%) and chronic (48% vs 0%) GVHD were more common among recipients of T-replete grafts. No significant differences in graft rejection/failure or viral infections were encountered between the two groups. Relapses were more prevalent (37% vs 15%) among recipients of T-depleted grafts. Outcome (EFS) was similar in the two groups. Consequently, in the analysis of transplant outcome, the higher risk of procedure-related, toxic complications among pediatric recipients of T-replete marrow grafts appears to be balanced by an increased risk of relapse among the recipients of T-depleted grafts. PMID- 10723583 TI - Long-term survival of ex-thalassemic patients with persistent mixed chimerism after bone marrow transplantation. AB - Twenty-six transplanted thalassemic patients out of 295 analyzed, showed the presence of persistent mixed chimerism, over a period of time varying between 2 and 11 years after BMT. Despite the presence of large numbers of residual host cells, these transplanted thalassemic patients no longer require red blood cell transfusions and have a functional graft, producing sufficient levels of hemoglobin A ranging from 8.3-14.7 g/dl. These ex-thalassemic patients with persistent mixed chimerism, although they did not achieve complete donor engraftment are no longer exposed to the risk of graft rejection. The mechanisms underlying this apparent state of tolerance or education in these patients are at the present time unknown. However, these observations may be useful for physicians involved in defining optimal strategies for clinical gene therapy, in utero hematopoietic stem cell transplantation and adoption of less toxic conditioning regimens in mini-transplantation. PMID- 10723584 TI - An engraftment syndrome in autologous stem cell transplantation related to mononuclear cell dose. AB - Engraftment syndrome (ES) is a toxicity of autologous stem cell transplantation that occurs unexpectedly and is occasionally fatal. This syndrome, manifested as fever, rash and pulmonary deterioration which becomes evident at marrow engraftment, has been described by several centers but as yet remains enigmatic. We describe this syndrome at a single institution and note that it has accompanied the transition from the use of autologous marrow rescue to peripheral blood stem cell rescue. In this study, the occurrence of ES is related to the mononuclear cell dose at reinfusion. We found, in agreement with other reports, that patients developing ES are predominantly women undergoing therapy for solid tumors who demonstrate neutrophil engraftment at a significantly greater rate than do those patients not expressing the syndrome. We did not note a significant relationship between growth factor use (G-CSF) or amphotericin B exposure and the syndrome, as has been previously reported. The progenitor cell populations obtained with autologous marrow and peripheral blood stem cells are different. We hypothesize that the interaction of committed myeloid precursors from the stem cell product with the pulmonary vascular endothelium can be deleterious, especially under the influence of the inflammatory cytokines present at the time of engraftment. PMID- 10723585 TI - The effect of the serotherapy regimen used and the marrow cell dose received on rejection, graft-versus-host disease and outcome following unrelated donor bone marrow transplantation for leukaemia. AB - Unrelated donor (UD) transplantation is the only potentially curative therapy for many leukaemia patients but is associated with a high mortality and morbidity. We sought to identify factors that could be optimised to improve outcome following UD transplantation in adults. Data was retrospectively analysed on 55 patients sequentially receiving UD transplants for CML or acute leukaemia (AL), all of whom received serotherapy for the prevention of GVHD and rejection. All patients received standard conditioning regimens. The first 28 patients transplanted also received combined pre- and post-transplant serotherapy with Campath 1G (days -5 to +5) and standard dose CsA plus MTX as GVHD prophylaxis (protocol 1). The subsequent 27 patients received a 5-day course of pre-transplant serotherapy alone either with ATG (CML patients) or Campath 1G (AL patients) on days -5 to -1 inclusive, with high-dose CSA plus MTX (protocol 2). The incidence of acute GVHD was low with no patient receiving either protocol developing > grade 2 disease. The use of protocol 2 and the administration of a bone marrow cell dose above the median (2.17 x 10(8)/kg) were the most important factors predicting engraftment (P = 0.03 and P = 0.001, respectively) but this only remained significant for cell dose in multivariate analysis (P = 0.03). Overall survival for the group was 45% at 3 years and was influenced by both age (P = 0.02) and disease status at transplantation (P = 0.001). Receiving a cell dose above the median was also associated with a trend towards better survival (P = 0.08), due primarily to a reduction in the TRM to 8.2% compared with 54.5% in those receiving a lower cell dose (P = 0.002). We conclude that pre-transplant serotherapy alone is highly effective at preventing acute GVHD following UD BMT and that additional post transplant serotherapy does not confer any benefit. Furthermore, a high marrow cell dose infused has a major effect in reducing transplant related mortality following UD BMT. PMID- 10723586 TI - Quantitative detection of t(14;18)-positive cells in patients with follicular lymphoma before and after autologous bone marrow transplantation. AB - The aim of this study was to evaluate whether a quantitative analysis of circulating t(14;18)-positive cells is of prognostic significance in patients with follicular lymphoma (FL) after myelo-ablative therapy supported by ABMT. We tested DNA from primary lymphoma tissue as well as PBMC before and after ABMT from 15 patients for the presence of the t(14;18) translocation. Nine patients showed a t(14;18) translocation, six patients were t(14;18)-negative. Circulating t(14;18)-positive cells of seven patients were quantitatively determined by limiting dilution assays combined with a two-step PCR and by real-time quantitative PCR. The results of both methods correlate very well. The number of circulating t(14;18)-positive cells decreased significantly in all patients after myeloablative therapy and ABMT, t(14;18)-negative blood samples were found in five of seven patients. In all patients circulating t(14;18)-positive cells reappeared within 2 years after ABMT showing two different patterns. During continuous CR the numbers of circulating t(14;18)-positive cells were found to be stable within one order of magnitude. In contrast, in one patient the relapse was accompanied by a logarithmic increase of t(14;18)-positive cells. In a second patient the enlargement of lymph nodes developing over a period of 12 months was accompanied by very slowly increasing numbers of t(14;18)-positive cells. In all cases where diagnostic lymph node tissue was available, the same t(14;18) translocation was found at first diagnosis and after ABMT as shown by nucleotide sequence analysis. We conclude that the quantitative detection of circulating t(14;18)-positive cells during follow-up of patients with FL after ABMT reflects the clinical course of the disease. Relapses are associated with increasing numbers of circulating t(14;18)-positive cells and continuous complete remissions with stable cell counts. PMID- 10723587 TI - Hematopoietic reconstitution after lethal irradiation and bone marrow transplantation: effects of different hematopoietic cytokines on the recovery of thymus, spleen and blood cells. AB - Lethally irradiated mice were grafted with syngeneic bone marrow cells or left ungrafted. Mice of each group were injected with different hematopoietic cytokines for 5 consecutive days starting immediately after irradiation or left uninjected. The recovery of lymphoid tissues induced by hematopoietic cytokines 7 days after irradiation and bone marrow cell transplantation was comparable to that observed at days 21-28 in irradiated, bone marrow-grafted, but cytokine uninjected mice. IL-11 or IL-6, in combination with IL-3, was able to hasten thymus, spleen and blood cell numbers and functions. SCF also displayed a detectable effect when used with IL-3. Conversely, the IL-6 superagonist K-7/D-6 was able, when injected alone, to induce significant recovery of thymus, spleen and blood cells. Thus, K-7/D-6 appears to be a most efficient cytokine for fast reconstitution of lymphoid tissues after irradiation and bone marrow transplantation. PMID- 10723588 TI - The CD34+90+ cell dose does not predict early engraftment of autologous blood stem cells as well as the total CD34+ cell dose. AB - CD90 or Thy-1 is an antigen co-expressed with CD34+ on putative immature hematopoietic stem cells. Peak mobilization of CD34+90+ cells into the blood occurs a few days earlier than peak mobilization of total CD34+ cells. Because it is not known which cell type best correlates with engraftment, the optimal timing of apheresis remains unclear. The purpose of the study was to determine if the CD34+90+ cell dose predicts engraftment of autologous blood stem cells independent of the total CD34+ cell dose/kg, the dose of other CD34+ cell subsets (CD34+33-, CD34+38-, CD34+41+), or various clinical factors. Data were analyzed on 125 consecutive patients ranging in age from 19 to 66 years (median 46) who underwent autologous blood stem cell transplantation (ABSCT) for breast cancer (54), lymphoma (59), or other malignancies (12). By univariate analysis, neutrophil (> or = 0.5 x 10(9)/l) and platelet (> or = 20 x 10(9)/l or > or = 100 x 10(9)/l) engraftment correlated better with the total CD34+ cell dose than with the CD34+90+ cell subset. Using Cox proportional hazards models, factors independently associated with both neutrophil engraftment (> or = 0.5 x 10(9)/l) and platelet engraftment (> or = 20 x 10(9)/l and > or = 100 x 10(9)/l) were higher total CD34+ dose/kg and high-dose regimen (melphalan-containing slower than other regimens). In conclusion, the total CD34+ dose/kg was a better predictor of hematopoietic engraftment following ABSCT than the dose of any CD34+ subset, including CD34+90+ cells. Apheresis should continue to be timed according to peak CD34+ levels. PMID- 10723589 TI - Shortening of telomeres in recipients of both autologous and allogeneic hematopoietic stem cell transplantation. AB - Telomere length of peripheral blood mononuclear cells (PBMCs) from 23 autologous HSCT patients ranging from 4 to 61 years old, and 46 allogeneic HSCT recipients from 6 to 52 years old were studied to confirm whether excessive shortening of telomeres is associated with HSCT. After autologous HSCT, telomere length of PBMCs ranged from 6.8 to 12.0 kb. The comparison between transplanted PBMCs and PBMCs after autologous HSCT showed shortening by up to 1.9 kb (mean +/- s.d.: 0.64 +/- 0.50 kb). There was a difference between autologous HSCT patients and normal volunteers in the slopes of regression lines. After allogeneic HSCT, telomere length of PBMCs ranged from 6.8 to 12.0 kb. Telomeres of recipients were up to 2.1 kb (0.60 +/- 0.468 kb) shorter than those of donors. The slope of regression lines for allogeneic HSCT patients and normal volunteers were parallel. Although all patients were transplanted with more than 2.0 x 10(8) cells/kg, telomere length did not correlate with the number of transplanted cells. There was no significant correlation between telomere length and recovery of hematological parameters. However, three patients with an average telomere length of 6.8 kb after HSCT took a longer period to reach the normal hematological state. Taken together, these data suggest that most HSCTs are performed within the biological safety range of telomeres, while the patients who have telomeres shorter than 7.0 kb after HSCT should be observed carefully for long-term hematopoiesis and the occurrence of hematopoietic disorders. PMID- 10723590 TI - Blood tacrolimus concentrations in bone marrow transplant patients undergoing plasmapheresis. AB - Microangiopathic hemolytic anemia (MAHA) is a well-described complication of stem cell transplantation. Plasmapheresis is one modality utilized as therapy for patients who develop this complication. However, plasmapheresis may alter whole blood levels of certain medications and its effect on tacrolimus in bone marrow transplant patients is unknown. Because tacrolimus has a narrow therapeutic range, the effect of plasmapheresis on whole blood concentrations would be important to know. We report three allogeneic BMT patients who were receiving tacrolimus as acute GVHD therapy while undergoing plasmapheresis for MAHA. Tacrolimus levels seemed unaffected by plasmapheresis in these patients. PMID- 10723591 TI - Detailed monitoring of hematopoietic chimerism in a child treated by adoptive immunotherapy for high risk of relapse after BMT for acute myeloid leukemia. AB - We report a case of a 13-year-old boy who was transplanted for relapse of acute myeloid leukemia (AML). A detailed study of hematopoietic chimerism was performed using polymerase chain reaction (PCR) of variable number of tandem repeats (VNTR) at very short time intervals. We used discontinuation of post-transplant immunosuppression and donor lymphocyte infusions (DLI) in order to prevent leukemia relapse that was indicated by a progressive increase in autologous hematopoiesis. Despite the fact that the boy relapsed 10 months after BMT, we could see a significant influence of adoptive immunotherapy on the mixed chimerism status during the post-transplant period. PMID- 10723592 TI - Detection of an anti-RhD antibody 2 years after sensitization in a patient who had undergone an allogeneic BMT. AB - We describe an HLA matched bone marrow transplantation with minor ABO incompatibility and RhD mismatch (donor RhD negative and recipient RhD positive). GVHD appeared on day +96 and therapy with steroid and cyclosporin was started. When GVHD disappeared and immunosuppressive therapy was stopped (2 years after BMT), an anti-RhD antibody was detected in the patient's serum. The delayed appearance of this antibody may have been associated with the prolonged immunosuppression that was required for treatment of the patient's GVHD. PMID- 10723593 TI - Diagnosis of Klinefelter syndrome in the donor after peripheral blood stem cell transplantation. PMID- 10723594 TI - The journal jigsaw--fitting the pieces together. PMID- 10723595 TI - An ethicist's commentary on the case of the clients who believe a vaccine created problems for their animal. PMID- 10723596 TI - Bacteria and fungi of marine mammals: a review. AB - A list of the different bacterial and fungal agents isolated from marine mammals in different parts of the world is presented. Importance is given to some of the most recently identified bacterial agents, including Actinobacillus delphinicola, A. scotiae, and Brucella spp. A list, in alphabetical order, of bacteria recovered from different tissues or organs from marine mammals is presented for the integumentary, respiratory, digestive, genitourinary, and reticuloendothelial systems. Infectious bacterial agents associated with abscesses and with cases of septicemia are also listed. Information about the different fungal agents recovered from marine mammals is summarized. A section covering some of the zoonotic infectious agents recovered from marine mammals is included. PMID- 10723597 TI - Arthrodesis of the proximal interphalangeal joint affected with septic arthritis in 8 horses. AB - Arthrodesis was performed to treat septic arthritis of the proximal interphalangeal joint of 8 horses. Records of the horses were reviewed to determine outcome and possible factors that influenced success or failure. All horses were female. Seven horses had 1 joint treated and 1 horse was treated for bilateral pelvic limb involvement. The duration of sepsis before surgery ranged from 1 to 66 days. Bone lysis and production was radiographically apparent in 7 horses before surgery. Six horses had multiple bacterial organisms cultured from bone or synovial tissues; 2 horses had single isolates identified. After aggressive curettage, arthrodesis was accomplished with 3 parallel screws in 1 horse, 2 divergent narrow dynamic compression plates in 3 horses, and a single broad dynamic compression plate in 4 horses. Casts were applied to all horses for 1 to 6 weeks. Four horses survived to successful brood mare status. Four horses were euthanized during hospitalization because of continued discomfort or complications of sepsis. Arthrodesis of the proximal interphalangeal joint affected with septic arthritis appears to be an acceptable alternative to euthanasia for some horses. PMID- 10723598 TI - The use of computer imaging technology to facilitate the capture of feedlot necropsy information. AB - The collection of necropsy information is an integral component of veterinary feedlot consulting. Computer imaging technology can be employed to facilitate the capture of feedlot necropsy data. A digital camera is used to capture necropsy images. Subsequently, the images are electronically transferred to a central site for veterinary interpretation and diagnosis. PMID- 10723599 TI - Caseous lymphadenitis caused by Corynebacterium ulcerans in the dromedary camel. AB - Caseous lymphadenitis that affected the dorsal and ventral superficial lymph nodes in the left cervicothoracic region of a young dromedary camel is described. The agent isolated was Corynebacterium ulcerans. To our knowledge, this is the first description of purulent lymphadenitis caused by C. ulcerans in a species belonging to the Camelidae. PMID- 10723600 TI - Shivering in a thoroughbred mare. AB - An 11-year-old mare presented with neuromuscular deficits and what resembled shivering in the left hind limb. On necropsy, there was no evidence of denervation atrophy of the left hind gastrocnemius muscle. The spinal cord had a small, right-sided lesion at C3-C4 and C4-C5. Tests for equine herpesvirus-1 and Sarcocystis spp. were negative. PMID- 10723601 TI - Eosinophilic cystitis in 3 dogs. PMID- 10723602 TI - Headroom requirements for horses in transit. AB - Horses intended for slaughter in Western Canada are frequently transported in double-deck trailers, where headroom may be restricted. Poll and withers height was estimated from type photographs of various horse breeds. The headroom required by Canadian legislation and codes of practice may not be sufficiently restrictive to protect the welfare of sport type horses when transported. PMID- 10723603 TI - Intestinal spirochetosis in a guinea pig with colorectal prolapse. PMID- 10723604 TI - Hydromorphone: a cost-effective alternative to the use of oxymorphone. PMID- 10723605 TI - Hidden messages. PMID- 10723606 TI - Synthesis and 1H NMR studies of 3-(D-erythro-glycerol-1-yl)-1H-pyrazolo[3,4 b]quinoxaline and its 7-chloro and 7-methyl analogues. AB - 3-(D-erythro-Glycerol-1-yl)-1H-pyrazolo[3,4-b]quinoxaline and its 7-chloro and 7 methyl analogues (11 and 12) were prepared from the corresponding quinoxalines. The 7-substituted analogues 11 and 12 were obtained as the preponderant isomers, and the 6-substituted analogues as the minor isomers. The structure and position of the substituent were determined by 1H NMR studies. The effect of substitution on the chemical shift of other protons is discussed. PMID- 10723607 TI - A one-step phosphorylation of D-aldohexoses and D-aldopentoses with inorganic cyclo-triphosphate. AB - The phosphorylation reaction by inorganic cyclo-triphosphate (P3m) having a six membered ring was examined for D-aldohexoses and D-aldopentoses in aqueous solution. Similar to the process for D-glucose, D-galactose, D-xylose or D-allose were phosphorylated with P3m to give stereoselectively beta-D-galactopyranosyl 1 triphosphate, beta-D-xylopyranosyl 1-triphosphate or beta-D-allopyranosyl 1 triphosphate with maximum yields of 31.3, 32.5 or 32.1%, respectively. On the other hand, in the reaction of D-ribose, D-lyxose, D-mannose or D-arabinose with P3m, the yields of beta-D-ribopyranosyl 1-triphosphate, alpha-D-lyxopyranosyl 1 triphosphate, alpha-D-mannopyranosyl 1-triphosphate or alpha-D-arabinopyranosyl 1 triphosphate were 8.0, 16.5, 9.6 or 14.1%, respectively. The phosphorylation mechanism of D-aldopyranoses with P3m was also discussed. PMID- 10723608 TI - Bivalency and epitope specificity of a high-affinity IgG3 monoclonal antibody to the Streptococcus group A carbohydrate antigen. Molecular modeling of a Fv fragment. AB - The binding of Strep 9, a mouse monoclonal antibody (mAb) of the IgG3 subclass directed against the cell-wall polysaccharide of Group A Streptococcus (GAS), has been characterized. The intact antibody and proteolytic fragments of Strep 9 bind differently to GAS: the intact mAb and F(ab)2' have greater affinity for the carbohydrate epitope than the monomeric Fab or F(ab)'. A mode of binding in which Strep 9 binds bivalently to portions of the polysaccharide on adjacent chains on GAS is proposed. A competitive ELISA protocol using a panel of carbohydrate inhibitors shows that the branched trisaccharide, beta-D-GlcpNAc-(1-->3)-[alpha-L Rhap-(1-->2)]-alpha-L-Rhap, and an extended surface are key components of the epitope recognized by Strep 9. Microcalorimetry measurements with the mAb and two synthetic haptens, a tetrasaccharide and a hexasaccharide, show enthalpy-entropy compensation as seen in other oligosaccharide-protein interactions. Molecular modeling of the antibody variable region by homology modeling techniques indicates a groove-shaped combining site that can readily accommodate extended surfaces. Visual docking of an oligosaccharide corresponding to the cell-wall polysaccharide into the site provides a putative model for the complex, in which a heptasaccharide unit occupies the site and the GlcpNAc residues of two adjacent branched trisaccharide units occupy binding pockets within the groove-shaped binding site. PMID- 10723609 TI - Lanthanide-saccharide chemistry: synthesis and characterisation of Ce(III) saccharide complexes. AB - A series of nine Ce(III) complexes has been synthesised with seven different monosaccharides (D-glucose, D-fructose, D-galactose, D-mannose, L-sorbose, D ribose and D-xylose) and two different disaccharides (D-maltose and L-lactose), and these have been characterised with various analytical, spectral, magnetic and electrochemical techniques. The NMR studies have highlighted some interesting features about the metal-ion-binding pattern of the saccharides. Some additional coordination has been proposed along with the chelating groups in the saccharide molecules, based on the shifts in 13C NMR spectra. On the other hand, solution absorption studies and solid-state magnetic susceptibilities have indicated the contribution from the d-character to the spectral features to some extent. PMID- 10723610 TI - Isolation and identification of poly-alpha-(1-->4)-linked 3-O-methyl-D mannopyranose from a hot-water extract of Mycobacterium vaccae. AB - A polysaccharide around 3.6 kDa has been identified as the major carbohydrate moiety of a antineoplastic protein-polysaccharide complex (PS4A) obtained by boiling intact cells of Mycobacterium vaccae in water. 1H and 13C NMR spectra of this polysaccharide suggested it was a highly homogeneous polymer composed substantially of one monomer, probably an alpha-linked O-methylated mannose. Comparison of the COSY spectra of the original and acetylated polymer indicated that the glycosidic linkage and the methyl ether were interchangeable, at O-3 and O-4. Further study demonstrated that the benzyolated hydrolysate of the polymer was 1,2,4,6-tetra-O-benzoyl-3-O-methyl-beta-mannopyranose. The hydrolysate was 3 O-methyl-alpha, beta-mannopyranose and the polymer was therefore poly-alpha-(1- >4)-linked 3-O-methyl-D-mannopyranose. This conclusion was further confirmed with an authentic sample of the monomer, which had spectral data identical to those of the hydrolyzate and co-eluted from an ion-exchange HPLC with the major sugar in the hydrolysate. PMID- 10723611 TI - Sugar interaction with metal ions. FT-IR study on the structure of crystalline galactaric acid and its K+, NH4+, Ca2+, Ba2+, and La3+ complexes. AB - The FT-IR spectra of galactaric acid and its K+, NH4+, Ca2+, Ba2+, and La3+ salts have been recorded and interpreted. Spectroscopic evidence shows that the dimeric carboxylic groups of the free acid are dissociated upon formation of the salt, and the asymmetric and symmetric stretching vibrations of the anionic COO- group in these salts are observed at about 1600 and 1400 cm-1, respectively. The two carboxylic groups of the galactarate coordinate with Ca2+ ions in a monodentate form. One of the carboxylic groups in the Ba2+ salt coordinates in a monodentate state; another group interacts with three cations in a tetradentate form. In the K+, NH4+, and La3+ salts, the COO- groups coordinate in a polydentate manner with the cations. By comparison of the spectra of the salts with that of the free acid, it is concluded that the hydroxyl groups of the galactarate skeleton take part in metal-oxygen interaction, and the hydrogen-bonding network is rearranged upon sugar metalation. The degree of participation of the sugar OH groups in metal-galactarate interaction is varied from the K+ and NH4+ salts to the Ca2+, Ba2+, and La3+ salts. PMID- 10723612 TI - NMR spectroscopy, molecular dynamics, and conformation of a synthetic octasaccharide fragment of the O-specific polysaccharide of Shigella dysenteriae type 1. AB - A synthetic octasaccharide fragment (2) of the O-specific polysaccharide (1) of Shigella dysenteriae type 1 has been studied as its methyl glycoside by one- and two-dimensional homo- and heteronuclear NMR spectroscopy. Complete 1H and 13C NMR assignments have been generated, and the 13C spin-lattice relaxation times have been measured for the octasaccharide 2. A congener (6) of this octasaccharide containing one D-galactose residue with a specific 13C label at C-1 has been synthesized and used to measure interglycosidic 13C-1H coupling by the 2D J resolved 1H NMR method. From the NMR data, three types of conformational restraints were developed: (a) 29 inter-residue, distance restraints; (b) 48 intra-residue, ring atom dihedral angle restraints, and (c) one heteronuclear, inter-residue dihedral angle restraint. The use of these restraints in a restrained molecular dynamics computation with simulated annealing yielded a conformation resembling a short, irregular spiral, with methyl substituents on the exterior. PMID- 10723613 TI - A new 6-C-alkylation from an alkyl mannofuranoside 5,6-cyclic sulfate. AB - Methyl 6-C-alkyl-6-deoxy-alpha-D-mannofuranoside derivatives have been synthesized from methyl 2,3-O-isopropylidene-5,6-O-sulfuryl-alpha-D mannofuranoside (1). In a Path A, reaction of the 5,6-cyclic sulfate 1 with 2 lithio-1,3-dithiane afforded 2-(methyl 6-deoxy-2,3-O-isopropylidene-alpha-D mannofuranosid-6-yl)-1,3-dith iane (2). Treatment of 2 with n-butyllithium then alkyl iodide gave the corresponding 2-(methyl 5-O-alkyl-6-deoxy-2,3-O isopropylidene-alpha-D-mannofuranosid-6-yl )-1,3- dithiane. Reaction of 2 with n butyllithium and 5,6-cyclic sulfate 1 furnished 2-[methyl 6-deoxy-2,3-O isopropylidene-5-O-(methyl 6-deoxy-2,3-O-isopropylidene-alpha-D-manno-furanosid-6 yl)-alpha-D - mannofuranosid-6-yl]-1,3-dithiane. 2-(Methyl 6-deoxy-2,3-O isopropylidene-5-O-methyl-alpha-D-mannofuranosid- 6-yl)-1,3-dithiane was converted into the lithiated anion, which after treatment with alkyl halide afforded the corresponding 2-alkyl-C-(methyl 6-deoxy-2,3-O-isopropylidene-5-O methyl-alpha-D-mannofuranosid-6-y l)-1,3- dithiane. In a Path B, 5,6-cyclic sulfate 1 reacted with 2-alkyl-2-lithio-1,3-dithiane derivatives, which led after acidic hydrolysis to 2-alkyl-2-(methyl 6-deoxy-2,3-O-isopropylidene-alpha-D mannofuranosid-6-yl)-1,3-dith iane accompanied by methyl 6-deoxy-2,3-O isopropylidene-alpha-D-lyxo-hexofuranos-5-u loside as the by-product. This methodology was applied to synthesize 2-(methyl 6-deoxy-2,3-O-isopropylidene-5-O methyl-alpha-D-mannofuranosid-6-y l)-2- (methyl 6-deoxy-2,3-O-isopropylidene alpha-D-mannofuranosid-6-yl)-1,3-dith iane. PMID- 10723614 TI - Communication between older patients and their physicians. AB - This article provides an overview of communication between older patients and their physicians. The authors discuss distinctive features of geriatric medical visits and empirical investigations of communication between physicians and older patients in real life clinical encounters highlighting the content, interactional processes, and outcomes of care. They also discuss strategies for improving communication between physicians and older patients using new and innovative technologies. The authors conclude that healing in its broadest sense can occur only through a humanistic approach to geriatric care. PMID- 10723615 TI - The influence of older patient-physician communication on health and health related outcomes. AB - Effective patient-physician communication significantly influences health outcomes of older patients. For example, concordance between patient and physician expectations and patient participation in the decision-making process affects older patients. Communication is also linked to patient recall, adherence, and satisfaction. Furthermore, communication impacts emotional and physical outcomes of older patients, although evidence of improved physical outcomes remains under-investigated in this population. Dimensions of communication, such as continuity of relationship, seem to be important in decreasing hospitalization of older patients. This article explores the link between communication and health care outcomes in the older population. PMID- 10723616 TI - The seasons of the patient-physician relationship. AB - There are seasons in the geriatric patient-physician relationship. As they age together, doctors and patients see one another through crises, recoveries, uncertainties, and losses, slowly weaving between them at times tapestries of complex narrative richness. What is known about the patient-physician relationship from scientific research can be contemplated by what is known about intimate human relationships. This article turns to three poems, written by Charles Simic, Wallace Stevens, and T. S. Eliot, to understand how the passage of time might lead to human intersubjective understanding. A method of reflecting on individual clinical relationships, first through a close reading of the medical records of patients and then through narrative writing about the individual patient-physician relationship as illuminated by retrospection, is then introduced. Three patients in the practice of the author are described in detail within the frameworks suggested by the poems, and directions for future research are outlined. PMID- 10723617 TI - Caregiving issues in the geriatric medical encounter. AB - Families provide much-needed care for dependent older persons, which can be both burdensome and stressful. In addition to providing personal care, families often are essential for optimal chronic disease management. Thus, two critical functions of the medical encounter are to provide empathic support to family caregivers and to provide education about chronic diseases and their management. Concomitantly, a conscious effort must be made to not compromise the doctor-older patient relationship, insofar as possible. Managing the doctor-patient-family caregiver relationship is challenging, especially in the settings of cognitive impairment and end-of-life care. In these circumstances in particular, both older patients and their families need the care of their physicians. PMID- 10723619 TI - Clinical pragmatism, ethics consultation, and the elderly patient. AB - This article describes a process of moral problem solving for medicine termed clinical pragmatism. A structured and disciplined method of addressing ethical problems in medical practice that clinicians will find useful in their daily routines is provided. After outlining the method of clinical pragmatism, the authors illustrate its use in a case involving an older patient refusing medical treatment and conclude with comments on the importance of the process in ethics case consultation. PMID- 10723618 TI - Empowering older patients to communicate more effectively in the medical encounter. AB - Active involvement of patients in medical encounters has been associated with several desirable outcomes, including greater satisfaction, increased adherence to treatment, and positive treatment outcomes. Older patients, particularly the very old and less well educated, are more likely to place physicians in a dominant role and themselves in a submissive role. Intervention trials to increase patient involvement have shown positive results. Activation interventions with older patients to increase a sense of control and self efficacy are promising. Most of the attention to improving doctor-patient interaction has been directed toward physicians. The results of these few intervention trials support increased attention to patient behaviors. PMID- 10723620 TI - The doctor-patient relationship in the home. AB - The rapid growth of the geriatric sector of the population has, in part, led to the explosion of the home care industry and the rebirth of the house call. Understanding the population being served is always the first step in providing excellent care. Older people who are frail and homebound have physical, social, and emotional needs that require careful assessment and team intervention in the home. To provide care, the practitioner must establish trust with the patient and the caregivers. Often, the agenda of the patient, family, and physician differ. A multidisciplinary team can offer help to meet the needs of each party. Allowing the patient to be an active participant in the care plan is paramount. Changes should be made slowly with careful explanation and education at each step, and the right of an individual to refuse treatment should be respected. Support for the caregiver and accessibility for consultation and advice can make a difference in the quality of care an older individual receives. PMID- 10723621 TI - The outpatient medical encounter and elderly patients. AB - There is little evidence of systematic negative bias against older patients in medical visits. The nature of the current narrative review, largely based on studies conducted after 1985, is consistent with the author's previous metaanalysis of over 40 studies published between 1965 and 1985. In that review, based on videotapes or audiotapes of medical visits, consistent relationships between patient age and physicians' interviewing skills were found. Older patients received more information, more total communication and questions concerning drugs, more courtesy, and perhaps more formality reflected in less laughter and joking than younger patients. Ultimately, the subtle ageism that may be present in medical visits with older patients is probably balanced somewhat by communication advantages afforded them challenging the negative views of older patients' care prevalent in the literature. This balance may help explain the ubiquitous finding that older patients are more satisfied with their health care, despite poorer health status, than younger patients. Nevertheless, other patients, especially those in the oldest cohorts, are at high risk for passive relationships and communication complications related to low literacy and poor health status and deserve the attention and special consideration of providers and health service researchers. PMID- 10723622 TI - Communication between the hospitalized older patient and physician. AB - Much research has shown that how physicians communicate with patients can have profound influence on behavioral, psychosocial, and clinical outcomes of the encounter. Communication with older patients, however, is often compromised by some attributes of the aging process. Communication can also be affected by the setting in which it takes place, and the hospital presents some barriers not found in ambulatory sites. These concerns are often compounded in end-of-life decisions for older patients when discussed in hospital settings. PMID- 10723624 TI - Managed care and the geriatric patient-physician relationship. AB - As managed care proliferates in the United States and other countries, its structure has patterned changes in patient-doctor relationships, including those between older patients and their physicians. The physician as gatekeeper now limits the access of the patient to information and services. Patient trust in the physician, essential to an effective patient-doctor relationship, will be damaged under this system of care. Additionally, examples from medical encounters demonstrate that many of the problems in the doctor-older patient relationship found under fee for service will remain, including the lack of attention to the contextual issues of health care of older adults. PMID- 10723623 TI - What nursing home residents value in their doctors. AB - Patient satisfaction is influenced by multiple factors, and different populations are expected to define satisfaction in terms of their novel perspectives. Despite growing interest in patient satisfaction, an extensive literature search reveals no studies of nursing home residents' satisfaction with respect to medical care. In an initial qualitative study using transcripts of interviews conducted as part of a state quality control mandate, categories are identified that make up this population's construct of satisfaction and dissatisfaction. These categories serve as building blocks for designing future studies investigating these issues and allowing for comparison of nursing home residents' ideas of satisfaction and dissatisfaction to other older patients, including those in an outpatient geriatric setting. PMID- 10723625 TI - Alzheimer's disease and other dementias. Implications for physician communication. AB - This article addresses the importance of language, speech, and communication accommodations between the physician and the patient with dementia. Following a brief summary of common profiles of speech, language, and communication in several different types of dementia, the authors discuss key elements based on a comprehensive model of communication enhancement for individuals with dementia. The primary emphasis of this article is that physicians must select individually tailored communication strategies drawn from a sound knowledge of patients' skills. More importantly, the article stresses that communication is not just the exchange of information, but communication includes components that help establish mutual respect and maintain patients' self-identity and autonomy. PMID- 10723626 TI - Physician-older patient communication at the end of life. AB - Communication with dying patients and their families requires special skills to assist them in this extremely stressful period. This article begins with a case that illustrates many of the challenges of communicating with the dying. It then reviews the literature about communication with older patients at the end of life, focusing on physician-patient discussions, decision-making, advance directives, and cultural factors. The article concludes with a practical discussion of problems that physicians may encounter when working with older patients at the end of life and their families and recommendations to improve communication. PMID- 10723627 TI - Archaeobacterial ether lipid liposomes (archaeosomes) as novel vaccine and drug delivery systems. AB - Liposomes are artificial, spherical, closed vesicles consisting of one or more lipid bilayer(s). Liposomes made from ester phospholipids have been studied extensively over the last 3 decades as artificial membrane models. Considerable interest has been generated for applications of liposomes in medicine, including their use as diagnostic reagents, as carrier vehicles in vaccine formulations, or as delivery systems for drugs, genes, or cancer imaging agents. The objective of this article is to review the properties and potential applications of novel liposomes made from the membrane lipids of Archaeobacteria (Archaea). These lipids are unique and distinct from those encountered in Eukarya and Bacteria. Polar glycerolipids make up the bulk of the membrane lipids, with the remaining neutral lipids being primarily squalenes and other hydrocarbons. The polar lipids consist of regularly branched, and usually fully saturated, phytanyl chains of 20, 25, or 40 carbon length, with the 20 and 40 being most common. The phytanyl chains are attached via ether bonds to the sn-2,3 carbons of the glycerol backbone(s). It has been shown only recently that total polar lipids of archaeobacteria, and purified lipid fractions therefrom, can form liposomes. We refer to liposomes made with any lipid composition that includes ether lipids characteristic of Archaeobacteria as archaeosomes to distinguish them from vesicles made from the conventional lipids obtained from eukaryotic or eubacterial sources or their synthetic analogs. In general, archaeosomes demonstrate relatively higher stabilities to oxidative stress, high temperature, alkaline pH, action of phospholipases, bile salts, and serum proteins. Some archaeosome formulations can be sterilized by autoclaving, without problems such as fusion or aggregation of the vesicles. The uptake of archaeosomes by phagocytic cells can be up to 50-fold greater than that of conventional liposome formulations. Studies in mice have indicated that systemic administration of several test antigens entrapped within certain archaeosome compositions give humoral immune responses that are comparable to those obtained with the potent but toxic Freund's adjuvant. Archaeosome compositions can be selected to give a prolonged, sustained immune response, and the generation of a memory response. Tissue distribution studies of archaeosomes administered via various systemic and peroral routes indicate potential for targeting to specific organs. All in vitro and in vivo studies performed to date indicate that archaeosomes are safe and do not invoke any noticeable toxicity in mice. The stability, tissue distribution profiles, and adjuvant activity of archaeosome formulations indicate that they may offer a superior alternative to the use of conventional liposomes, at least for some biotechnology applications. PMID- 10723628 TI - Echocardiographic assessment of prosthetic heart valves. PMID- 10723629 TI - Relationship of cognitions to fear of somatic symptoms: a test of the cognitive theory of panic. AB - The relationship between fear of physical anxiety symptoms and cognitive misinterpretation of those symptoms, as measured by responses to the Body Sensations Questionnaire and the Agoraphobic Cognitions Questionnaire, respectively, was examined for two samples of outpatients with panic disorder. Factor analytic and correlational analyses demonstrated that the patients' self rated fear of specific physical and psychological symptoms was related to the frequency of specific logically related catastrophic thoughts (e.g., fears of heart palpitations or chest pressure with thoughts of a heart attack). This specific relationship between the somatic sensations and the catastrophic thoughts experienced by agoraphobic individuals provides further support for the cognitive theory of panic disorder. When the responses to the two questionnaires were factor-analyzed together, four factors were identified: symptoms and thoughts relevant to cardiovascular, neurological, gastrointestinal, and behavioral control systems, respectively. These findings suggest that the nature of panic-related fears varies across patients, and that the use of specific treatment interventions designed to modify the specific variations in their expression may be advisable. PMID- 10723630 TI - Convergent, discriminant, and criterion-related validity of the Social Phobia and Anxiety Inventory. AB - The psychometric properties of the Social Phobia Anxiety Inventory (SPAI) were assessed in a sample of participants who either had a primary diagnosis of social phobia or were normal volunteers. Positive evidence was obtained on the SPAI's concurrent and predictive convergent validity: it was significantly correlated with other measures of social anxiety obtained from self-report questionnaires of social impairment and with behavioral assessment measures administered in conjunction with a conversation role-play, including measures of negative thinking, subjective anxiety, and self-perceived skill and apparent nervousness. Discriminant validity was demonstrated by the SPAI's significantly stronger relationship with public than private self-consciousness. There was no overlap in SPAI scores in the normal volunteer and socially phobic groups, demonstrating criterion-related validity. PMID- 10723631 TI - Autonomic reactivity of panic patients during a CO2 inhalation procedure. AB - The objectives of this study were to compare continuous subjective and physiological responses of panic disorder patients and normal controls during 5% CO2 inhalation. Psychophysiological responses of panic disorder patients (n = 42) and controls (n = 25) were monitored during baseline (20 min), 5% CO2 inhalation (20 min), and recovery (20 min). The data were compared at baseline and over periods of the experiment using analysis of variance. A subgroup of patients who experienced panic attacks during the CO2 inhalation (n = 12) were significantly different from the other subjects on baseline heart rate and on variability of systolic and diastolic blood pressure, skin conductance, and breathing variability (length and number of breathing pauses and length of breathing cycle variability). Inspection of the data showed that elevation in blood pressure and breathholding were present during some of the panic attacks, suggesting that some attacks may represent a complex psychophysiological response with elements of a "freezing" reaction, well described in animal experiments, which can quickly shift to a "fight/flight" reaction that is usually characterized by an increase in heart and breathing rate. However, some patients had only minimal changes in breathing and others had minimal psychophysiological changes during the time they indicated that they had a panic attack. Panic attacks are not homogeneous and may be characterized by a variety of physiological and cognitive responses. This may indicate that biological mechanisms of panic include abnormality in many functionally connected areas of the brain responsible for complex psychophysiological reactions to multiple threatening situations. PMID- 10723632 TI - Flumazenil challenge in social phobia. AB - The benzodiazepine antagonist, flumazenil, can provoke panic attacks in some panic disorder patients. It has been predicted that panic responses to flumazenil may be associated with situational fear. Patients with social phobia frequently experience situational anxiety and panic attacks. The current study tested whether flumazenil induces panic in patients with social phobia. Fourteen patients with social phobia (DSM-III-R) and 14 age- and sex-matched controls were tested in a single session, double blind crossover challenge design, using intravenous flumazenil 2 mg/20 ml or matched placebo infusions 1 hour apart. Panic attacks occurred during flumazenil challenge in 2/14 subjects with social phobia. The rate of panic attacks and the severity of panic symptoms following flumazenil were not significantly greater in patients than in controls. Situational fears that are provoked by social cues therefore do not predict panic responses to flumazenil. PMID- 10723633 TI - Cell-surface expression of L-selectin (CD62L) by blood lymphocytes: correlates with affective parameters and severity of panic disorder. AB - The surface immune phenotype of peripheral blood lymphocytes (PBL) was examined in 30 patients meeting DSM-III-R criteria for panic disorder and in 10 normal controls by immunostaining and cytofluorimetry. Patients with panic disorder and controls showed comparable numbers of PBL and no differences in the percentages of blood T-cells, B-cells, or NK-cells. The PBL in panic disorder patients showed a trend toward enrichment for "naive" CD45RA+ T-lymphocytes (35.0 +/- 7.6 vs. 28.7 +/- 9.8, P = 0.09) and significant enrichment for cells expressing CD62L (L selectin, 22.9 +/- 5.9 vs. 14.6 +/- 6.3, P = 0.002), a lymphocyte homing receptor that mediates binding to lymph node endothelium. Increased expression of CD62L correlated directly with the global severity of illness, Hamilton Anxiety (HAM-A) and Hamilton Depression (HAM-D) scores. Although in the normal range, plasma cortisol levels were significantly increased in patients with panic disorder (P = 0.003) with respect to controls and correlated with the expression of CD62L by PBL. We conclude that the peripheral blood in panic disorder shows phenotypic changes that may reflect diminished cell activation in vivo. PMID- 10723634 TI - Predictors of anxiety in patients with Alzheimer's disease. PMID- 10723635 TI - Acute responses of anxiety disorder patients after a natural disaster. PMID- 10723637 TI - Variation of psychiatric emergencies across seasons in San Diego county. PMID- 10723638 TI - Moving from innovation to implementation: trends in foods for particular nutritional uses. Introduction. PMID- 10723636 TI - Treatment of social phobia with the dopamine agonist pergolide. PMID- 10723639 TI - The role of research--how much is enough? AB - A great deal of research is involved in bringing PARNUTs to the marketplace. Research often begins with identifying an unfilled nutritional need, determining if that need is great enough to warrant development of a new product, and then evaluating the efficacy and safety of the potential product in animal models and clinical trials. This approach tends to emphasize short-term outcomes, however, while neglecting the issues of whether a product offers long term benefit or holds long term risks. This article presents discussion centered around the need for selecting appropriate outcomes for nutritional intervention trials, designing trials with a follow-up time sufficient to allow for outcome measurement, and enrolling a patient population large enough to accurately gauge efficacy and tolerability. PMID- 10723640 TI - Nutrition throughout the life cycle. PMID- 10723641 TI - The role of industry in bringing foods for particular nutritional uses (PARNUTS) to the market. AB - The process of bringing new food products from innovation to implementation requires a high level of interaction between researchers, marketers, and consumers. Researchers from industry and academia have the task of developing products that are not only efficacious, but also have a high probability of consumer acceptance. For most foods, industry must provide the most leadership in finding new product concepts, determining which products will have the widest markets, and in funding research and development. To accomplish these tasks, industry has forged partnerships with academic centers and scientists who excel in research and development, and continues to search for the best ways to communicate with consumers. For some FSMPs, other considerations, such as medical and nutritional needs (e.g., products for inborn errors of metabolism), might change the pattern of industry leadership. The following article explores the ways in which industry can facilitate the development and acceptance of beneficial and marketable food products. PMID- 10723642 TI - The role of scientific and expert advisory committees in food product research and approval. AB - Scientific and expert advisory committees responsible for food products often have the advantage of being relatively unhindered by rigid regulations and the simultaneous disadvantage of having few guidelines to clarify their role in directing the research and approval process. Committees can thus miss opportunities to function in a proactive advisory capacity, and to assist in predetermining what research and documentation are necessary for regulatory approval of a particular food product. This paper examines the ways scientific and expert committees for nutritional products can contribute to formulation of procedures for effective hypothesis and study design development, preparation of well-structured, complete dossiers for product approval, and transparent interactions with petitioners. PMID- 10723643 TI - Regulating food products without impeding innovation. AB - Industry and regulatory bodies share a common goal of making beneficial products available to consumers, but the relationship between industry and regulators can become adversarial if it is not handled properly. When industry views the regulatory process as an obstacle to product development and marketing, and regulators view petitioners as having only a profit motive, the opportunity to work together efficiently to get products to market is lost. While it is difficult to codify how industry and regulators should interact, it is worthwhile to look at some of the most provocative issues and see how they might be addressed. The discussion that follows will examine the functioning of regulatory bodies in the European Union, consider how more flexibility might be added to the regulatory process, and raise the issue of how regulatory bodies can function to serve the consumer while promoting innovation. PMID- 10723644 TI - Beyond implementation: the role of physicians, health care professionals and consumers in the development and use of food products. AB - Physicians and consumers play an essential role in determining the ultimate success of food products. Effective communication of product efficacy and safety data to physicians, health care professionals and consumers can be a deciding factor in whether a food product is recommended or consumed. Food products deemed safe and beneficial will be further judged for cost and cost effectiveness. Physician and consumer reaction to newly launched products can be a valuable source of feedback if used to create communication plans and to reevaluate products. Long term monitoring of safety, efficacy, and consumer use patterns in the context of post-marketing surveillance studies is another potential feedback mechanism. The present review focuses on how physicians and consumers are involved in food product development and refinement, how they use the information available to make choices, and what kinds of information must be supplied to allow the public to be fully informed. PMID- 10723645 TI - Trends in sexual activity among adolescent American women: 1982-1995. AB - CONTEXT: The formulation of policies and development of programs regarding adolescent sexual and reproductive health requires up-to-date information on levels of and trends in teenage sexual activity. METHODS: Analysis of three NSFG surveys, carried out in 1982, 1988 and 1995, allows examination of the sexual behavior of teenage women over a 13-year time period, using comparable data for the entire time period. RESULTS: The proportion of adolescent women who ever had sexual intercourse increased somewhat during the 1980s, but this upward trend stabilized between the late 1980s and the mid-1990s. Throughout the period, there has been little change in the proportion currently sexually active: In each of the surveys, about 40% of all 15-19-year-olds had had sexual intercourse in the last three months. The average number of months in the past year in which sexually experienced teenagers had had intercourse declined during the 1980s, with no change in the continuity of sexual intercourse taking place between 1988 and 1995, when the mean remained at 8.6 months. Differences in teenage sexual behavior across poverty and racial and ethnic subgroups were large in the early 1980s, but narrowed over the 13-year period. CONCLUSIONS: Only continued monitoring will tell whether the patterns observed during 1988-1995 signify a temporary leveling off in the trend toward increasing adolescent sexual activity, stability in behavior or the beginnings of a decline. Nevertheless, the sustained level of initiation of sexual activity during adolescence is by now a recognized pattern of behavior, and is an important characteristic of the transition to adulthood in the United States. PMID- 10723646 TI - The correspondence between intention to avoid childbearing and subsequent fertility: a prospective analysis. AB - CONTEXT: Retrospective studies of pregnancy intendedness have revealed some characteristics that can help identify which women are more likely than others to experience an unintended birth. A comparison of these findings with those from a prospective analysis may shed greater light on the characteristics associated with unintended pregnancy. METHODS: Data were taken from the 1988 National Survey of Fertility Growth and a telephone reinterview of respondents conducted in 1990. Separate analyses were conducted of women intending to postpone childbearing for at least three years and of women intending to forgo all future childbearing. Logistic regression models were used to identify the effects of social and demographic characteristics, as well as change in marital status and certainty of intentions, on the odds of experiencing a birth in the interval between interviews. RESULTS: Only 10% of women intending to postpone pregnancy for more than three years and 8% of respondents seeking to forgo future childbearing had a birth in the interval between interviews. (These births, referred to as unpredicted births in this article, are roughly analogous to those labeled unintended in retrospective analyses.) Women with incomes below the poverty level were 2-3 times as likely as women with incomes between 100% and 199% of poverty to experience an unpredicted birth. Race was not a significant factor among women intending to avoid future childbearing, and became nonsignificant among those intending to postpone when change in marital status and contraceptive status were taken into account. Women aged 35 and older who wanted no more children were significantly less likely than women aged 20-29 to have an unpredicted birth. Women aged 30-34 who wanted to postpone childbearing were roughly 70% less likely than women aged 20-29 to experience an unpredicted birth. Overall, women who were at risk for a pregnancy but not practicing contraception were 2-3 times more likely than women using an effective method to have an unpredicted birth. CONCLUSIONS: There are at least two potential explanations for instances where the correlates of unintended births in the prospective analysis differ from those identified in retrospective studies. Certain subgroups of women may be more likely to classify births as wanted when they are asked retrospectively; alternatively, they may be more likely to experience changes in their living conditions that alter their fertility intentions. PMID- 10723647 TI - Sexual partnership patterns as a behavioral risk factor for sexually transmitted diseases. AB - CONTEXT: Women's and men's number of sexual partners and protective practices such as condom use can have a direct effect on their risk of contracting sexually transmitted diseases (STDs), including HIV. METHODS: The 1988 and 1995 cycles of the National Survey of Family Growth and five rounds of the General Social Survey conducted from 1988 to 1996 are used to examine women's and men's numbers of recent sexual partners. Levels of direct risk for STDs (two or more partners in the past year) and the social and demographic correlates of multiple partnership are analyzed among women and men. In addition, women's indirect risk for STDs (their partners' involvement with other partners in the past year) is used to estimate their overall risk of STDs through multiple partnerships. RESULTS: At least three-quarters of sexually active U.S. women and men in the late 1980s and mid-1990s had had only one sexual partner in the preceding 12 months. Moreover, there is no indication that the proportion with more than one partner in the past year changed substantially over that period. Nevertheless, combining women's and men's partnership reports suggests that about 17 million women aged 15-44--34% of those sexually active in the past year--were at risk for STDs because of direct exposure to multiple partners (5.4 million), indirect exposure (6.3 million) or both direct and indirect exposure (5.5 million). In all, 21% of women were at direct risk and 23% were at indirect risk. In comparison, among men aged 18-44, 24% were at direct risk for STDs and an unknown proportion were at indirect risk. Multivariate analyses indicated that unmarried individuals, women younger than 40 and men aged 20-29, blacks and women in the South were all at elevated risk for STDs because of multiple partnership. Overall, in 1995, 19% of sexually active women aged 15-44 had used condoms to protect against STDs over the preceding year, and 19% of those sexually active in the three months before the survey were current condom users. Condom use specifically for STD prevention was more common among women reporting both direct and indirect risk for STDs (58%) and among those at direct risk (46%) than among other women; women whose partners put them at indirect risk only were less likely to be current or recent condom users than women who were not at risk or were only at direct risk. CONCLUSIONS: There is a continuing need to educate people regarding their risk for STDs, to increase the use of existing barrier methods and to develop new methods that protect against STD infection. In addition, if we are to develop a better understanding of the extent of STD risk through multiple partnership, the collection of information on number of partners and relationships between partners must be expanded and improved. PMID- 10723648 TI - Women's experience and satisfaction with emergency contraception. AB - CONTEXT: If any new contraceptive technology is to become a viable option for decreasing unintended pregnancies, women must be willing to use the method and find it acceptable. However, because emergency contraceptive pills have not been widely used, very little is known about this method's acceptability. METHODS: Telephone interviews were conducted with 235 women who had received emergency contraceptive pills through a demonstration project at 13 Kaiser Permanente medical offices in San Diego to assess women's experience and satisfaction with the pills. RESULTS: More than two-thirds of the women (70%) were using a contraceptive method prior to their need for emergency contraception, and 73% of these users were relying on condoms. When asked about the situation that led to unprotected intercourse, 45% reported that their condom broke or slipped, while 23% said they had had unplanned sex. More than three-quarters of the sample (81%) experienced at least one side effect. The overwhelming majority were satisfied with emergency contraceptive pills (91%) and would recommend them to friends and family members (97%). Just one-quarter of the sample (28%) believed that emergency contraceptive pills should be dispensed over the counter, and an even lower proportion agreed that they should be available from vending machines (6%). CONCLUSIONS: Because women were overwhelmingly accepting of emergency contraceptive pills, found them easy to use and did not intend to substitute them for regular contraceptive use, this new method is an important addition to the contraceptive options available to women, providing a way to prevent pregnancy after unprotected intercourse or method failure. PMID- 10723649 TI - Abortion services in rural Washington State, 1983-1984 to 1993-1994: availability and outcomes. AB - CONTEXT: Fewer rural health providers offer abortion services than a decade ago. It is unknown how the reduction in service availability has affected women's pregnancy outcomes, the extent to which they must travel to obtain an abortion or whether abortions are delayed as a result. METHODS: Population, birth and fetal death data, as well as pregnancy termination reports, obtained from Washington State were used to calculate abortion rates and ratios and birthrates for Washington residents in 1983-1984 and in 1993-1994. Residence of abortion patients was classified by county only, and location of providers was recorded as large urban county, small urban county, large rural county or small rural county. Distances that women traveled to obtain an abortion were calculated. Chi-square tests were used to compare urban and rural rates and ratios within time periods, and to compare changes that occurred between time periods. RESULTS: Birthrates and abortion rates decreased for both rural and urban Washington women between 1983-1984 and 1993-1994, but the magnitude of the decrease was greater for rural women. The rural abortion rate fell 27%, from 14.9 abortions per 1,000 women to 10.9 per 1,000, while the urban rate dropped 17%, from 21.8 to 18.2 per 1,000. The decline in the abortion rate was larger for adolescents than it was for other age-groups. In rural areas, the abortion rate decreased from 16.5 per 1,000 adolescents aged 10-19 in 1983-1984 to 10.8 per 1,000 in 1993-1994, while it declined from 23.3 per 1,000 to 16.9 per 1,000 in urban areas. From the earlier to the later time period, rural women traveled on average 12 miles farther each way to obtain an abortion, and the proportion who obtained the procedure in a rural county decreased from 25% to 3%. In the earlier time period, 62% of rural women traveled 50 miles or more to obtain an abortion, compared with 73% in 1993 1994. From 1983-1984 to 1993-1994, the proportion of rural women who traveled out of state for an abortion increased from 8% to 14%. The proportion of rural women terminating their pregnancy after the first trimester increased from 8% in 1983 1984 to 15% in 1993-1994. CONCLUSION: Rural Washington women are traveling farther and more often to urban and out-of-state locations for abortion services, and are obtaining their abortions at a later gestational age, which is associated with a decade-long decline in the number of abortion providers. PMID- 10723650 TI - Are all contraceptive failures unintended pregnancies? Evidence from the 1995 National Survey of Family Growth. AB - CONTEXT: The incidence of unintended pregnancy has long been used as a primary indicator of the state of reproductive health. However, the definition--and therefore the measurement--of this indicator has been elusive. METHODS: Data from the 1995 National Survey of Family Growth (NSFG) were used to compare levels of unintended pregnancy among contraceptive users based on two definitions--the standard definition based on women's reports of contraceptive failure, and the NSFG definition based on pregnancy timing (wanted then, wanted later, or not wanted then or in the future). An attitudinal scale was used to examine women's feelings about their unintended pregnancy. RESULTS: Of pregnancies classified as contraceptive failures under the standard definition, only 68% were unintended pregnancies--94% of those ending in abortion and 60% of those ending in birth. Just 59% of women with a contraceptive failure classified as an unintended pregnancy reported feeling unhappy or very unhappy about their pregnancy, while 90% of those with a failure classified as an intended pregnancy reported being happy or very happy. CONCLUSIONS: Measures of wantedness based on women's feelings about their pregnancy may correlate more closely with important pregnancy outcomes than do traditional measures of intendedness. PMID- 10723651 TI - A reminder that human behavior frequently refuses to conform to models created by researchers. PMID- 10723652 TI - Pregnancy intentions may not be a useful measure for research on maternal and child health outcomes. PMID- 10723653 TI - Ambivalent feelings about parenthood may lead to inconsistent contraceptive use- and pregnancy. PMID- 10723654 TI - Intended pregnancies and unintended pregnancies: distinct categories or opposite ends of a continuum? PMID- 10723655 TI - Options for measuring unintended pregnancy in cycle 6 of the National Survey of Family Growth. PMID- 10723656 TI - [Preventive effect of chlormadinone acetate on flare-up phenomenon in advanced prostate cancer administered with a luteinizing hormone-releasing hormone analogue]. AB - To investigate whether chlormadinone acetate (CMA) could prevent the flare-up phenomenon induced by a luteinizing hormone-releasing hormone analogue (LH-RHa), we treated 4 cases of stage C and 17 cases of stage D prostate cancer with CMA for 4 weeks and CMA plus monthly injection of LH-RHa for following 24 weeks. Serum LH, testosterone, and prostate-specific antigen (PSA) levels were closely monitored before and 3 days, 1-, 2-, and 4-weeks after LH-RHa injection. Subjective and objective responses were also investigated. Serum LH and testosterone levels significantly elevated 3 days after the initial injection of LH-RHa. However, they resumed 1 week after LH-RHa injection with fluctuation under the normal range. Out of 21 cases, 3 cases (14%) consisting of 2 poorly and 1 moderately differentiated adenocarcinoma showed increased serum PSA levels 1 week after LH-RHa injection in spite of suppressed serum testosterone levels. The objective response of these 2 poorly differentiated cases was progressive disease at 24 weeks. No cases indicated worsening of clinical symptoms concerning flare up phenomenon. CMA seemed to be capable of preventing flare-up phenomenon in advanced prostate cancer. PMID- 10723658 TI - [Renal cell carcinoma in a horseshoe kidney with abdominal aortic aneurysm: a case report]. AB - We herein report a case of renal cell carcinoma in a horseshoe kidney with an abdominal aortic aneurysm in a 69-year-old man. Radiological examinations showed a left renal tumor, horseshoe kidney and abdominal aortic aneurysm. We performed a left radical nephrectomy with the division of the isthmus and artificial graft through an abdominal transperitoneal approach. Histological findings revealed clear cell type renal cell carcinoma without invasion of the capsule or renal pelvis. Only 31 cases of renal cell carcinoma in a horseshoe kidney have been reported in Japan, and our case is the 32nd. No case with abdominal aortic aneurysm has been reported previously. We assume that abdominal aortic aneurysm was associated with renal cell carcinoma by chance in the horseshoe kidney in this case. The arterial and venous supplies vary from case to case. We emphasize that arteriography and venography are very important preoperative procedures. PMID- 10723657 TI - [Endocrine therapy of stage D2 prostate cancer--comparison of drugs used for total androgen blockade]. AB - Patients with Stage D2 prostate cancer were treated with surgical or medical (LHRH analog) castration combined with either estrogen, chlormadinone acetate or flutamide as initial therapy. The effect of each medication was compared. The overall survival, cause-specific survival and relapse-free survival were not different among the three medications. Patients given each medication were divided into two groups each according to grade, extent of diseases on bone metastases, and levels of tumor marker. Survivals of the corresponding two groups were compared with each other among different medications. No differences were revealed with any medication. There were no serious side effects in whole patients, except that grade 2 liver dysfunction was accompanied in 12% of flutamide-treated group. It is concluded that the three drugs used with castration did not make any difference in the survival of stage D2 patients, and differences between medications were seen in the frequency of side effects. PMID- 10723659 TI - [Chromophobe cell renal carcinoma: a case report]. AB - Chromophobe cell renal carcinoma is an uncommon subtype of renal cell carcinoma and the number of cases studied is still limited in Japan. We here report a case of chromophobe cell renal carcinoma in a 41-year-old Mexican male. He visited our branch hospital with the symptom of upper abdominal pain. Ultrasound examination showed a left renal mass. He was admitted to our hospital for treatment of a left renal mass. Radiological examinations revealed a hypervascular tumor in the left kidney. Under the clinical diagnosis of possible renal cell carcinoma, left radical nephrectomy was performed. This tumor was diagnosed as chromophobe cell renal carcinoma with a microscopic examination of H & E stained specimens, histochemical staining using Hale's colloidal iron and an ultrastructural study. PMID- 10723660 TI - [Renal inflammatory pseudotumor after embolization for arteriovenous malformation: a case report]. AB - A 61-year-old man was admitted to our hospital with the chief complaint of asymptomatic macrohematuria. Because he was diagnosed with arteriovenous malformation of the right kidney by angiography, embolization was performed. However, 2 years and 3 months later, bleeding from the right kidney was detected. Computed tomography (CT) revealed a low density area (1.8 cm) in the right kidney protruding to the renal pelvic adjacent to the old embolized lesion. Since we could not make a conclusive diagnosis as malignant disease by ureteroscopy and angiography, we have decided to follow the case carefully. Nine months later, macroscopic hematuria worsened, and the low density area by CT in the right kidney expanded. Therefore, we finally decided to perform right total nephroureterectomy. The tumor was histologically diagnosed as renal inflammatory pseudotumor. This is the 14th case reported in Japan. We here report the renal inflammatory pseudotumor with a review of the Japanese literature. PMID- 10723661 TI - Mesonephric adenocarcinoma of the urinary bladder: a case report. AB - We report a very rare case of mesonephric adenocarcinoma of the urinary bladder, the origin of which is still uncertain. A non-papillary and broad-based tumor was located in the trigone and bladder neck on cystoscopic examination. Pelvic magnetic resonance imaging in T2-weighted images revealed a mass invading into the muscular layer of the bladder wall. Histologic examination of bladder cup biopsy specimens showed adenocarcinoma. She underwent total cystectomy and pelvic lymph node dissection. Histologically, the tumor was chiefly composed of cells with eosinophilic cytoplasm and partly of cells with clear cytoplasm or hobnail shaped cells, arranged in tubular or papillary structures, and infiltrated peri vesical fat tissues. She died of metastatic disease 22 months after surgery. To the best of our knowledge, the present case is the 19th reported in the literature. PMID- 10723662 TI - [Adenocarcinoma of the ileal segment with transitional cell carcinoma of the bladder following ileocytoplasty: a case report]. AB - A 67 year-old woman visited our hospital complaining of pollakisuria. She had undergone left nephrectomy and augmentation ileocystoplasty for tuberculous bladder atrophy 40 years previously. She underwent a total cystectomy and tubeless ureterocutaneostomy with a preoperative diagnosis of muscle-invading transitional cell carcinoma of the bladder. The pathological diagnosis was adenocarcinoma of the ileal segment and transitional cell carcinoma of the original bladder. This is the first case report of adenocarcinoma of the ileal segment and transitional cell carcinoma of the original bladder among 22 patients suffering from bladder cancer after ileocystoplasty. PMID- 10723663 TI - [Giant bladder stone: a case report]. AB - An 88-year-old woman was hospitalized with the chief complaints of lower abdominal pain, miction pain and pollakisuria. Radiographs showed a giant bladder stone shadow, 12.0 x 9.0 cm in size. Cystolithotomy was performed under the diagnosis of bladder stone. The extirpated stone weighed 510 g, and measured 10.0 x 7.5 x 6.0 cm in size. The stone had four compartments of stratified lamellae composed of calcium phosphate and magnesium ammonium phosphate. The postoperative course was uneventful and the bilateral hydronephrosis improved markedly on DIP. PMID- 10723664 TI - [A case of epithelioid leiomyoma (leiomyoblastoma) of the urethra]. AB - A 25-year-old woman was admitted to our hospital because of a painless, slow growing mass arising from the external genitalia. The mass had been developing for a few years. She did not have difficulty voiding nor was there hematuria. Magnetic resonance imaging revealed a well-circumscribed, 4 cm, solid tumor between the urethra and the anterior vaginal wall. After needle biopsy confirmed a benign tumor arising from the smooth muscle, the tumor was removed by a transvaginal approach. The histopathological diagnosis was epithelioid leiomyoma (leiomyoblastoma), which often occurs in the stomach or uterus, but seldom around the urethra. The patient has remained well without recurrence for 20 months after surgery. PMID- 10723665 TI - [CA19-9-producing testicular tumor: a case report]. AB - A rare case of CA19-9-producing testicular tumor is reported. A 37-year-old male who had complained of high fever and right scrotal swelling was referred to our department. Ultrasonography and computed tomography demonstrated a right testicular tumor with right lung metastasis and aortocaval lymph node metastasis. Right high orchiectomy was performed. The histopathological diagnosis was mixed type of teratoma, yolk sac tumor, embryonal carcinoma and seminoma. Immuno histochemical analysis showed CA19-9 to be expressed in the cancer cells. After 5 courses of combination chemotherapy, the operation for right lung metastasis was performed. The CA19-9 level was lowered to the normal range within four weeks after the first operation. He has been free of recurrence for about 18 months after the lung operation. PMID- 10723666 TI - [Efficacy of single-dose therapy with levofloxacin for acute cystitis: comparison to three-day therapy]. AB - To investigate the safety, efficacy and recurrence-inhibiting effect of a single dose of levofloxacin (LVFX) for the treatment of acute uncomplicated cystitis in women, 56 females with acute uncomplicated cystitis between 17 and 73 years old were studied, based on the urinary tract infection (UTI) drug efficacy evaluation. The patients were divided into two groups; in Group A a single 200 mg dose of LVFX was administered, while in Group B, 100 mg of LVFX was administered twice a day for three days. The patients were re-examined three days later. The presence or absence of recurrence was surveyed by a questionnaire 3 months after the treatment. The efficacy rate on the third day after the administration in Group A and Group B was 96.9% (32 cases) and 95.8% (24 cases), respectively, and the recurrence rate within three months after administration 17.4% (4 in 23 cases) and 5.6% (1 in 18 cases), respectively. As for adverse drug reaction, abdominal pain was seen in one case, without a clear cause-effect relationship. Although the number of cases studied was small, no significant difference was seen between the single dose group and 3-day dose group in the safety, efficacy and recurrence-inhibiting effect of the new quinolone antibacterial treatment for acute uncomplicated cystitis in women, confirming the usefulness of the single dose treatment. PMID- 10723667 TI - Anesthetic considerations for patients with severe emphysematous lung disease. AB - The pathophysiology, medical and surgical management of emphysema have been reviewed as a foundation to the physiological goals and principles of anesthetic management of patients with emphysema. An understanding of the cardiovascular and respiratory consequences of emphysema combined with anesthesia, PPV, and thoracic surgery is essential to achieving the challenging physiological goals of providing anesthesia, positive pressure and one-lung ventilation, and postoperative analgesia in a manner consistent with rapid postoperative extubation, hemodynamic stability, adequate gas exchange, and minimal barotrauma for this population of patients. PMID- 10723668 TI - Physiology of the lateral decubitus position and one-lung ventilation. AB - OLV is most frequently utilized to provide a quiet field for the performance of many different surgical procedures. In some patients, severe hypoxemia may result, mandating the implementation of other therapies to provide adequate oxygenation. This paper has reviewed the physiological consequences of the lateral position that may contribute to the hypoxemia and the techniques we utilize at our institution for establishing OLV, maintaining OLV, and treating hypoxemia during OLV. Our technique is performed with the goal of maintaining adequate gas exchange and protecting the ventilated lung from potential overdistension and injury. It remains for future study to determine if the use of a lung protective strategy during intraoperative OLV offers any benefit to patients at risk for postoperative lung injury, such as those undergoing major lung resections. PMID- 10723669 TI - Anesthesia and airway management for tracheal resection and reconstruction. PMID- 10723670 TI - The bronchospastic patient. PMID- 10723671 TI - Basic mechanisms of hyperbaric oxygen in the treatment of ischemia-reperfusion injury. AB - HBO treatment affects many of the components involved in I/R injury, including PMNL function, endothelial CAM expression, NO production, NOS expression, cellular energetics, lipid peroxidation, and microvascular blood flow. Given the variety of models used to study the individual components involved in I/R injury, it is difficult to determine which is the predominant factor affected by HBO and which generates the observed beneficial outcomes in most systems. Experimental differences in the types of I/R injury, the timing of HBO treatment relative to the I/R injury (before, during, after, or delayed), the duration of HBO treatment pressure and duration, and the time of outcome measurements confound our ability to compare studies and determine the key beneficial factor. Upon review, it is likely that the sum of many of these effects is responsible for the final outcome. We have been presented with many of the pieces of the puzzle with respect to the beneficial effect of HBO in ischemia-reperfusion injury states. Hopefully, future studies will unite them into a clear picture of the basic mechanism(s) responsible for the benefits of hyperbaric oxygen therapy. PMID- 10723672 TI - Decompression illness, iatrogenic gas embolism, and carbon monoxide poisoning: the role of hyperbaric oxygen therapy. PMID- 10723673 TI - Hyperbaric oxygen therapy for trauma: crush injury, compartment syndrome, and other acute traumatic peripheral ischemias. AB - In the future, the indications for HBO therapy in acute peripheral ischemic injuries will likely be based on objective criteria rather than, as at present, on clinical diagnoses alone. This chapter offers objective criteria for using HBO in crush injuries and compartment syndromes. The pathophysiology of ATPI are well defined. Hyperbaric oxygen mediates the effects of ATPI through four mechanisms: hyperoxygenation, vasoconstriction, reperfusion, and host factors. The cost benefits of using HBO will be substantial, since complications from ATPI are very expensive. As objective criteria replace the presently used subjective criteria, hyperbaric oxygen therapy will become an integral part of trauma management of these injuries. PMID- 10723674 TI - Management of the critically ill patient in the hyperbaric chamber. PMID- 10723675 TI - Pathways of T cell pathology in models of chronic intestinal inflammation. PMID- 10723676 TI - Colitis in HLA-B27/beta 2 microglobulin transgenic rats. AB - Rats of susceptible genetic backgrounds expressing high copy numbers of the transgene encoding HLA-B27 and human beta 2 mu develop chronic colitis complicated in the advanced stage by adenomatous polyps progressing to adenocarcinoma. Unique features of this model include a spectrum of extraintestinal manifestations resembling to some extent human spondyloarthropathy, with peripheral and axial joint, dermatologic and male genital inflammation. Inflammation is T lymphocyte mediated, although surprisingly CD4+ cells are more active in transferring disease than CD8+ cells, which would be expected to be preferentially activated by Class I MHC peptides. Inflammation is dependent on a nonlymphoid bone marrow-derived cell, expressing high copy numbers of B27, probably APCs. In vitro function of transgenic dendritic cells is deficient, and in vivo competition for peptide binding in the antigen binding site of B27 attenuates arthritis. Normal bacteria are required for disease expression, with B. vulgatus preferentially able to induce colitis, whereas other bacteria such as E. coli stimulate no inflammatory response. Inflammation and resulted complications are modulated by non-MHC genes and are amenable to treatment by bone marrow transplant from normal donors. These results support the hypothesis that gastrointestinal and systemic inflammation in B27 transgenic rats is the result of loss of tolerance to enteric bacteria, as a consequence of defective APC (? dendritic cells) function. Whether disease is the result of selective MHC binding of enteric antigens uniquely capable of inducing disease, lack of appropriate induction of a CD8+ suppressor cell population, or skewed cytokine (IL-12, IL-18) secretion by APCs remains to be determined. PMID- 10723677 TI - TNBS-colitis. AB - Disease states, such as the occurrence of gastrointestinal inflammation (Crohn's disease and ulcerative colitis), can be secondary to a host of determinants that act in conjunction to bring about pathologic change. The underlying factors that mediate the development of such mucosal inflammation has recently been brought to the forefront with the advent of animal models. The examination of these animal models have given researchers a better understanding of the mechanisms involved in the pathogenesis of inflammatory bowel disease. This review discusses one such model, TNBS-colitis, and the insights that it provides into the occurrence of IBD and its future treatment. PMID- 10723678 TI - The C3H/HeJBir mouse model: a high susceptibility phenotype for colitis. AB - C3H/HeJBir is a substrain of C3H/HeJ mice that was generated by selective breeding for the phenotype of spontaneous colitis. These mice show increased B cell and T cell reactivity to antigens of the enteric bacterial flora. CD4+ T cells from this strain cause colitis, when activated by enteric bacterial antigens and transferred to histocompatible severe combined immunodeficient recipients. The expression of the disease phenotype of spontaneous colitis is greatly influenced by housing conditions and probably requires an immunostimulatory enteric flora. This strain seems to carry multiple susceptibility genes for colitis as does the parental C3H/HeJ strain; the genes involved are being mapped. This strain represents a high susceptibility phenotype for colitis that is providing insight into the interactions among immune, environmental and genetic factors that can result in IBD. PMID- 10723679 TI - The development of animal models of inflammatory bowel disease. PMID- 10723680 TI - Chronic colitis in IL-10-/- mice: insufficient counter regulation of a Th1 response. AB - IL-10-deficient (IL-10-/-) mice, generated by a gene-targeted mutation, develop abnormal immune responses as a result of uncontrolled interactions between antigen presenting cells and lymphocytes. The studies reviewed herein have focused on the enterocolitis that spontaneously develops in IL-10-/- mice. Not unexpectedly, heightened production of proinflammatory mediators accompanied pathologic changes in the gastrointestinal tract of young mutants. In a series of studies, the proinflammatory mediators responsible for initiating the pathogenic response were distinguished from those that were elicited as a consequence of persistent inflammation. We have also investigated the possibility that different mediators are involved in the inductive versus the maintenance phase of disease. The findings of these mechanistic studies as they relate to our understanding of progressive inflammatory disease and the role of IL-10 in controlling the acute and chronic stages are discussed. PMID- 10723681 TI - Spontaneous chronic colitis in TCR alpha-mutant mice; an experimental model of human ulcerative colitis. AB - Mice with targeted disruption of the T cell receptor alpha gene (TCR alpha-/-) spontaneously develop chronic colitis. Colonic inflammation begins at 6-8 weeks of age and chronic colitis is established in about 60% of mice by 16-20 weeks of age. The disease is also associated with autoantibodies (anti-tropomyosin antibodies, anti-neutrophil cytoplasmic antibodies) and an oligoclonal immune response to luminal bacterial antigens. Although T cells, but not B cells or autoantibodies, are essential for the development of colitis, B cells and/or autoantibodies may have a regulatory role in the pathogenesis of this colitis because the colitis is more severe in B cell deficient TCR alpha-/- mice. Cytokines, specifically IL-4 and IL-1, also play an important role in the development of colitis in TCR alpha-/- mice. Enteric bacteria located in the large intestine are an important factor in the pathogenesis of this colitis because germ-free TCR alpha-/- mice do not develop colitis and appendectomy at an early age delays the onset of this colitis. The colitis in TCR alpha-/- mice resembles human ulcerative colitis and provides a useful model to study the pathogenesis of human inflammatory bowel disease. PMID- 10723682 TI - Blunt abdominal trauma in children. AB - Blunt abdominal trauma is the commonest cause of intra-abdominal injuries in children. The use of computerized axial tomography and non-operative management of haemoperitoneum are two significant developments in the last two decades in the management of blunt abdominal trauma in children. The concept of non operative management was introduced in late 1979 and wherever possible remains the optimum treatment. Computerized tomography scan for paediatric abdominal trauma was first described in 1980 and remains the investigation of choice. There is no substitute, however, for a good history, astute physical examination, and strict adherence to the principles of primary and secondary survey, prompt resuscitation, vigilant monitoring and repeated evaluation. PMID- 10723683 TI - Survival, cranial ultrasound and cerebral palsy in very low birthweight infants: 1980s versus 1990s. AB - OBJECTIVE: To determine the changes in the rates of survival, cranial ultrasound abnormalities and cerebral palsy in very low birthweight (VLBW) (birthweight 500 1499 g) infants between the early 1980s and the early 1990s. METHODOLOGY: A cohort study of consecutive VLBW live births in one tertiary perinatal hospital during two distinct eras was performed at The Royal Women's Hospital, Melbourne, a level-III perinatal centre. Consecutive VLBW infants born over the 18-month period from 1 October 1980 (n = 222), and over the 12-month period from 1 January 1992 (n = 202) were identified. The main outcome measures were the proportions of live births surviving to 5 years of age, rates of cranial ultrasound abnormalities, and rates of cerebral palsy at 5 years of age. RESULTS: Over the 18 months from 1 October 1980, 68% (150/222) VLBW live births survived to 5 years of age. The survival rate rose substantially to 82% (165/202) during 1992 (odds ratio 2.1, 95% confidence interval 1.4-3.2). The survival rate increased over time more for those of 500-999 g birthweight than for those of 1000-1499 g birthweight. The rates of cerebroventricular haemorrhage (CVH) were similar inlive births and survivors from both eras, as were the rates of cerebral palsy (7.5% in 1980-82; 7.8% in 1992) in survivors seen at 5 years of age. The positive predictive value of CVH for cerebral palsy was low, but cystic periventricular leucomalacia was followed by cerebral palsy in seven of eight survivors from the 1992 cohort. CONCLUSIONS: Despite the increasing survival rate with improvements in perinatal care, including more antenatal steroid therapy and the introduction of exogenous surfactant, the rates of CVH and of cerebral palsy in survivors have not diminished. PMID- 10723684 TI - Factors associated with recurrent hospitalization in chronically ill children and adolescents. AB - OBJECTIVES: To determine factors associated with recurrent hospitalization in children with chronic illnesses in the Barwon Region. METHODOLOGY: Patients with four or more admissions to the Geelong Hospital children's ward over a 12-month period were identified. Their records were reviewed and the opinions of involved staff (medical, nursing, psychiatry, psychology, and social work) were sought. Multidisciplinary discussions were held to identify factors precipitating or maintaining the need for hospitalization. The numbers, illnesses and profiles of those admitted recurrently were compared with the data from the Barwon Paediatric Consultation Profile from the same period, and with those patients seen by the local counselling service for young people with chronic illnesses. RESULTS: Twenty-seven children had four or more admissions over the 12 months; these represent 0.05% of the child population regionally, or 2% of those with chronic illness. They account for 8.7% of hospital admissions and 16% of inpatient days. Two-thirds (18/27) had major psychosocial issues largely responsible for their admissions. A checklist was formulated of important medical, family, social, psychological, developmental, and institutional considerations. The most frequently identified psychosocial issues were medical dependency, psychological or medical problems affecting other family members, family and medical disparity regarding the treatment agenda, the lack of more intensive community supports, and medical controversy regarding best management. CONCLUSIONS: Ongoing medicalization and medical dependency, driven both by staff and families, can perpetuate recurrent hospitalization. Further awareness and training in these issues and development of community resources will be necessary if this process is to be changed. PMID- 10723685 TI - Favourable neurological outcomes following delivery room cardiopulmonary resuscitation of infants < or = 750 g at birth. AB - OBJECTIVE: To study short- and long-term outcomes of infants < or = 750 g birthweight who received cardiopulmonary resuscitation (CPR) in the delivery room. METHODOLOGY: A retrospective analysis of all inborn live births < or = 750 g birthweight from 1990 to 1996. Cardiopulmonary resuscitation was defined as positive pressure ventilation via an endotracheal tube and chest compressions. Univeriate analysis were conducted comparing patients according to the use of CPR or positive pressure ventilation alone. RESULTS: Cardiopulmonary resuscitation was administered to 16 infants: four received chest compressions only and 12 also received adrenaline. Cardiopulmonary resuscitation recipients had significantly lower Apgar scores at both 1 and 5 min, and had delayed onset of spontaneous respiration (P < 0.01). Seven patients died, and eight of nine survivors were free of major neurodevelopmental abnormalities at follow up. All CPR recipients with a 5 min Apgar score of < or = 5 and delayed onset of spontaneous respiration beyond 5 min had poor outcomes. CONCLUSION: Contrary to the majority of published evidence, delivery room CPR in our extremely small infants was not associated with a high risk of severe neurodevelopmental disability. PMID- 10723686 TI - A quality assurance review of outpatient care of children with life-threatening asthma exacerbations. AB - OBJECTIVES: A hospital admission for asthma represents an opportunity to address and improve asthma control. The aims of this study were to compare the ambulatory care of children admitted to the intensive care unit (ICU) following a life threatening asthma exacerbation with published guidelines of asthma management and to identify areas that could be targeted for change. METHODS: A retrospective review of case notes of children admitted to the ICU with asthma over a 6-month period. Variables recorded were: demographic; asthma history (including prior pattern of asthma, hospital admissions, interval treatment and managing doctor); admission details (consultation of respiratory team and asthma educator); and discharge management. RESULTS: There were 40 admissions of 38 children (24 males) with mean age 5.7 years (range 1.1-14 years). The majority (58%) had previous admissions for asthma (55 admissions in 22 children), with 23% of these to ICU. Sixty three per cent of those with previous admissions had persistent asthma, but only 29% were on inhaled corticosteroid (ICS). Most (60%) were managed by their local medical officer (LMO). Use of ICS was more likely if managed by a paediatrician. A respiratory subspecialist was consulted in 42% and the asthma educator in 70% of ICU admissions. Discharge medication included ICS in 74%, with no interval treatment in 18% of admissions. Follow up was by a respiratory subspecialist in 25% of cases. CONCLUSION: Asthma management before and after admission with life-threatening asthma did not conform to available guidelines. Persistent asthma was under-treated. Paediatricians were more likely to use interval treatment than LMO. We identified areas in which quality of care and outcome could be improved in this vulnerable group of asthmatics. PMID- 10723687 TI - Disclosure of developmental disability: a study of paediatricians' practices. AB - OBJECTIVE: To investigate paediatricians' practices in disclosure of disability and the influences on their practices, including attitude to people with disabilities. METHODOLOGY: Interviews were conducted with 26 paediatricians regarding their disclosure practices and their experience, training, contact with children with significant disabilities and influences on practices. Anonymous self-report questionnaires to the same group of practitioners relating to attitude to disability were also employed. RESULTS: Paediatricians' practices in the disclosure process scored relatively low on an index based upon recommended practices. No significant relationships were found between index scores and the experience or training of the paediatrician or the amount of contact of the paediatrician with children with disabilities. However, more experienced paediatricians were found to be more likely to mention the practice of informing both parents together and the presence of a support person at the time of disclosure. Paediatricians having more contact with children with disabilities were more likely to mention that they would disclose disability in a child as soon as possible. The major modifying influences on disclosure practices were reported to be the intelligence of the parents and their emotional state of at the time of disclosure. Time was the most frequently reported constraint upon disclosure practices. CONCLUSIONS: The low 'disclosure practice index' scores in this study are not necessarily an indication that practices are poor, as there are challenges to the validity of the advocated practices. There were few significant associations found between the practices of paediatricians in disclosure and their experience, training, contact with children with disabilities and attitude to people with disabilities. PMID- 10723688 TI - Disclosure of developmental disability: a study of parent satisfaction and the determinants of satisfaction. AB - OBJECTIVE: To investigate the level of parent satisfaction with the first communication of a diagnosis of developmental disability in their child ('disclosure') and the determinants of this satisfaction. METHODOLOGY: Interviews with parents of children with developmental disabilities regarding their experiences at the time of disclosure and their level of satisfaction with the process were carried out. RESULTS: Parent satisfaction with disclosure overall was found to be high (82.6%). Parents were more likely to be satisfied if they received a large amount of information. Parent satisfaction was found to be higher when the disclosing professional communicates well with the parents, has an understanding of parental concerns, and is direct in manner. Having both parents, the child or support people present were not found to have any significant relationship to parent satisfaction. CONCLUSIONS: The high level of satisfaction with disclosure in this study supports the claim made by earlier researchers that parental dissatisfaction with the disclosure process is not inevitable. The major determinants of parental satisfaction with disclosure are directness, understanding of parental concerns and good communication on the part of the disclosing professional, and receiving a large amount of information. PMID- 10723689 TI - Parent reported allergy and anaphylaxis in 4173 South Australian children. AB - OBJECTIVES: To determine the prevalence of parent reported allergy and anaphylaxis in a sample of children and to investigate the first aid management of anaphylaxis in the schools and preschools that these children attend. METHODS: The study sample comprised 4173 South Australian children aged 3-17 years. Information was collected regarding parent-reported allergy and anaphylaxis. All children with known anaphylaxis were contacted and an attempt was made to contact those with reports of allergy to ascertain if these children had anaphylaxis. A telephone questionnaire was used to verify reports of anaphylaxis and determine the cause, severity and school first aid management of anaphylaxis. RESULTS: The rate of parent reported allergy and anaphylaxis was 7.3 and 0.59 per 100 children, respectively. Two-thirds of children with anaphylaxis did not have emergency medication available at school, an emergency action plan, or a teacher on site able to administer adrenaline for first aid use. CONCLUSIONS: Children attending preschool or school may have had a past history of anaphylaxis. Arrangements for first aid management of these children while in this environment are often inadequate. PMID- 10723690 TI - Reference ranges for blood pressure in preschool Australians, obtained by oscillometry. AB - OBJECTIVE: To derive reference centiles for blood pressure in children aged 1-6 years which seek to address shortcomings in available reference ranges. METHODS: Prospective cohort study of 2876 children in Perth, Western Australia, commenced in 1989 with serial blood pressure measurements through early childhood obtained by oscillometry under standardized conditions. RESULTS: Gender-specific reference centile charts for systolic and diastolic blood pressure, (i) across ages 1-6 years and (ii) across the range of corrected Body Mass Index values at ages 1, 3 and 6 years, were generated by fitting linear models with both fixed and random effects. CONCLUSIONS: Reference values for blood pressure for young children are of clinical use and may be of long-term predictive value. PMID- 10723691 TI - Methylxanthines and sensorineural outcome at 14 years in children < 1501 g birthweight. AB - OBJECTIVES: Methylxanthines, including theophylline, have been used extensively and successfully to treat apnoea in preterm infants. However, long-term consequences of such therapy are largely unknown. The aim of this study was to determine the relationship between theophylline therapy and outcome at 14 years of age in surviving preterm children of birthweight < 1501 g. METHODOLOGY: The subjects of this study were 154 consecutive survivors with birthweights < 1501 g born from 1 October 1980 to 31 March 1982; 130 (84.4%) were assessed at 14 years of age. Outcomes included motor function, psychological test scores, and growth. RESULTS: Of the 130 children assessed, 69 (53.1%) had been exposed to theophylline; 13.0% had cerebral palsy, significantly higher than 1.6% in the 61 children not exposed to theophylline (P < 0.02). This difference remained statistically significant after adjusting for potential confounding variables including the presence of cerebroventricular haemorrhage. In contrast, after adjusting for known confounding variables, children who had received theophylline achieved higher psychological test scores. There was no association between theophylline therapy and growth. CONCLUSIONS: Theophylline therapy in the newborn period is associated with some evidence of harmful, but also helpful sensorineural effects at 14 years of age. PMID- 10723692 TI - Difference in susceptibilities of different cell lines to bilirubin damage. AB - OBJECTIVE: To investigate if there are differences in susceptibilities to bilirubin toxicity of different cell lines. METHODOLOGY: A modified 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) method was adopted to study the cytotoxic effect of bilirubin on several commercially available cell lines including human glioblastoma (ATCC CRL 1690, T98G), human neuroblastoma (ATCC HTB-10, SK-N-MC), human liver (ATCC CCL 13, Chang Liver, HeLa markers) and a mouse fibroblast (ATCC CCL-1, NCTC Colon 929). RESULTS: Cytotoxicity was observed when certain bilirubin:albumin molar ratios were exceeded in the medium of a cell line in culture. Different cells exhibited different susceptibilities to the cytotoxic effects of bilirubin; neuroblastoma and glioblastoma were most susceptible, fibroblasts were the least vulnerable. CONCLUSIONS: Our findings have confirmed the clinical impression that different cells sustain different degrees of cytotoxicities caused by bilirubin. PMID- 10723693 TI - Growth and nutrition of Hong Kong children aged 0-7 years. AB - OBJECTIVE: To study the early dietary practices in relation to growth of Hong Kong children from birth to 7 years. METHODOLOGY: One hundred and seventy-three full-term Hong Kong Chinese babies were recruited at birth and were followed up for anthropometric measurements using standardized methods and dietary assessment using a combination of dietary history, 24 h recall and food frequency. At 7 years, 125 children remained in the study. RESULTS: Mean (SD) birthweight was 3.3 (0.38) kg for boys and 3.1 (0.38) kg for girls. Mean (SD) weight at 7 years was 22.4 (4.2) kg for boys and 21.1 (3.7) kg for girls, and mean (SD) height was 120.3 (4.8) cm for boys and 119.8 (5.1) cm for girls. Hong Kong children were lighter and shorter than Australian children and the National Centre for Health Statistics (NCHS) references, but the magnitude is less than one standard deviation score. Mean weight and height of Hong Kong children were lower compared to Caucasian and Beijing children, with more obvious differences between 1 and 5 years. At 1 year, mean (SD) daily energy intake was 98 (24) kcal/kg/day for boys and 100 (26) kcal/kg/day for girls. By 7 years, it decreased to 82 (18) kcal/kg/day for boys and 73 (22) kcal/kg/day for girls. Between 2 to 4 years of age the energy intake of studied children were slightly lower than the Australian and Finnish children, but the protein intake was higher. Percentage of fat contributing to total daily energy intake was lower throughout at a level of 30%. Such differences in diet reflect a lower consumption of milk fat, higher consumption of meat and lower level of physical activity in Hong Kong children. Intakes of calcium, iron and vitamin C all reached 60% or above of US recommended daily allowance. CONCLUSIONS: The smaller body build of Chinese compared to Caucasians cannot be explained by dietary differences. The diet of Hong Kong children is changing to one which is more Westernized with a higher consumption of animal products. PMID- 10723694 TI - Accidental asphyxia in bed in severely disabled children. AB - OBJECTIVE: To determine whether there are specific situations which may increase the risk of accidental asphyxia during sleep in children with physical and mental disabilities. METHODOLOGY: Review of all cases where death was attributed to accidental asphyxia caused by unsafe sleeping situations in children listed in the Department of Histopathology database over a 10-year period from March 1989 to February 1999. RESULTS: A total of 26 cases were found (M:F, 19:7; age range, 1-48 months; average age, 7.4 months). Of those cases, two involved children with significant mental and physical impairment. Case 1: A 4-year-old boy with Klippel Trenaunay-Weber syndrome, macrocephaly and severe developmental delay, was found dead with his head hanging over a wooden board attached to the side of his bed. Case 2: A 4-year-old boy with lissencephaly and severe developmental delay was found dead wedged between a retractable mesh cot side and the side of his bed. In both cases the devices resulting in death had been put in place to prevent the boys from falling out of bed. CONCLUSIONS: Accidental asphyxia in physically and mentally impaired children may be caused by devices that have been used to prevent injury from falling out of bed. Careful assessment of the specific developmental problems that children suffer should be undertaken before their beds are modified. It may be safer for these children either to have no barrier, or to have drop-sided cots/beds that meet recognized safety standards. PMID- 10723695 TI - True hermaphroditism. AB - OBJECTIVE: True hermaphroditism is a rare cause of atypical genitalia which presents significant diagnostic and management challenges. We present the clinical and laboratory findings and management of four patients with true hermaphroditism. METHODOLOGY: Case studies from a teaching hospital and literature review. RESULTS: All four patients had atypical genitalia identified at birth. All had a palpable gonad, only one of which was palpable at birth. Three patients were 46XX (SRY -ve) and one 46XY (SRY +ve). Three patients were raised as females (two 46XX and one 46XY) and one as a male. All four patients were found to have an ovotestis present. CONCLUSIONS: The management of true hermaphroditism is controversial and requires a multidisciplinary approach. It has many implications for both the parent and child. We discuss the issues involved for the patients and their parents. PMID- 10723696 TI - Statistics for clinicians. 1: Introduction. PMID- 10723697 TI - Neonatal chickenpox. PMID- 10723698 TI - Nocturnal enuresis: what is happening? AB - Primary nocturnal enuresis is common and has considerable psychological ramifications for children as they get older. It is a familial condition with complex inheritance patterns. The pathophysiology of the condition appears to be related to poor arousal from sleep, nocturia due to deficient vasopressin release in sleep and possibly a decrease in functional bladder capacity especially at night. The mainstay of treatment is the bed-wetting alarm. In recent years, desmopressin nasal spray has found a clinical niche as a short-term solution for children attending school camps or sleeping over at friends' houses and as treatment in the medium term for those unresponsive to treatment with a bed wetting alarm. It may also be used as an adjunct to the use of the alarm. Treatment with imipramine is increasingly in disfavour because the relapse rate is unacceptably high and fatal overdose is a real possibility. PMID- 10723699 TI - Tetracycline-induced benign intracranial hypertension. AB - We report on a young adolescent with benign intracranial hypertension which we attribute to the use of minocycline for acne. PMID- 10723700 TI - Pre-excitation syndrome secondary to cardiac rhabdomyomas in tuberous sclerosis. AB - Rhabdomyomas are not uncommon in infants with tuberous sclerosis. We describe a neonate who presented with hydrops fetalis arising from a tachyarrhythmia during fetal life related to rhabdomyomas. After reversion of the arrhythmia, pre excitation was noted on an interval electrocardiogram. Following regression of the tumours, the delta wave disappeared with no further arrhythmias noted. PMID- 10723701 TI - Kingella kingae infections in children. AB - OBJECTIVE: To increase awareness of Kingella kingae infections in children by presenting four cases seen at the Gold Coast Hospital, Southport, Queensland, and reviewing the literature. METHODOLOGY: Records of the four cases were reviewed and relevant information described. A MEDLINE search of the English literature from 1983 to 1998 was conducted. RESULTS: Osteoarticular infections are the commonest type of invasive paediatric infection but bacteraemia and endocarditis also occur. Isolation of the organism is difficult but inoculation of the specimen into enriched blood culture systems improves the recovery rate. The majority of isolates are sensitive to beta-lactam antibiotics but resistance has been described. CONCLUSIONS: Kingella kingae infections in children are more common than previously recognized. The organism should be actively sought in any child with suspected osteoarticular infections. Recommended empiric therapy is a third generation cephalosporin until susceptibility to penicillin is confirmed. PMID- 10723702 TI - Elevated plasma bile acid concentrations in two sisters with tyrosinaemia type I. AB - A 21-month-old girl suffering from tyrosinaemia type I and undergoing treatment with 2-(2-nitro-4-trifluoro-methylbenzoyl)-1,3-cyclohexanedione (NTBC) presented with pruritus which rapidly ceased with administration of high doses of ursodeoxycholic acid. Determination of plasma bile acids revealed clearly elevated levels both in samples taken before and after the onset of NTBC therapy, thus indicating, that the increase was not related to the administration of this drug. This result is corroborated by data from the first patient's newborn sister, diagnosed with the same disease, who showed elevated plasma bile acid concentrations in all samples examined, except for the cord plasma. This is the first report on altered bile acid concentrations in tyrosinaemia type I, and underlines the need for thorough investigation of bile acid metabolism in this disease. PMID- 10723703 TI - Gastrografin aspiration in a neonate with tracheoesophageal fistula. PMID- 10723704 TI - Sydenham's chorea: a resurgence in the 1990s? PMID- 10723705 TI - High incidence of galactosaemia among the Irish travelling community. PMID- 10723706 TI - A pilot study of teachers' perspective on attention deficit hyperactive disorder. PMID- 10723707 TI - The syntax of features. AB - The syntax of features is a complicated and densely researched domain, from a variety of theoretical perspectives. The present paper takes one influential syntactic framework [the minimalist program of Chomsky, 1995] as its vantage point, and discusses the major issues that arise within this framework in the domain of the syntax of features, with particular emphasis on Case and agreement. PMID- 10723708 TI - Syntactic features in reanalysis: positive and negative symptoms. AB - Meng and Bader have presented evidence that a Case conflict is a more effective cue for garden-path reanalysis than a number conflict is, for German wh-sentences with subject-object ambiguities. The preferred first-pass analysis has the wh trace in subject position, although object position is correct. In a speeded grammaticality judgment task, perceivers accepted Case-disambiguated examples more often and more rapidly than number-disambiguated examples, although comprehension questions indicated that both were eventually understood correctly. For ungrammatical sentences, a Case mismatch error resulted in more false positive grammaticality judgments than a number mismatch error. We offer an explanation for why Case and number features differ in these two ways in their effects on sentence processing. We propose, within the Diagnosis Model of garden path processing, that reanalysis triggered by a Case mismatch guides the parser more effectively toward the correct structure. Case is a positive symptom, which carries information about the new structure that must be built. By contrast, a number mismatch is a negative symptom; it invalidates the incorrect structure without showing how to rebuild it. This difference in the transparency of garden path repair can also account for the greater overacceptance of Case-disambiguated ungrammatical sentences. The speeded grammaticality judgment task is designed to encourage hasty responses. Usually, these are hasty rejections of garden path sentences that, on calmer reflection, the parser would find acceptable. Conversely, over-hasty acceptance could occur if some initial progress is made in resolving a grammatical problem. Thus, a higher rate of false positives on ungrammaticals is to be expected where reanalysis proceeds successfully for a while before blocking. PMID- 10723709 TI - Case and reanalysis. AB - In this paper we discuss an asymmetry in the Case system of German and its implications for human sentence processing: the asymmetry between nominative/accusative and dative case. Starting from the assumption that dative case has a distinct grammatical representation--dative DPs are embedded into an extra structural layer KP--the results of two experiments will be presented, which show that dative assignment during reanalysis is accompanied by additional processing operations that are not needed when accusative or nominative are assigned. In particular, we show that dative assignment during reanalysis triggers reaccess to the mental lexicon, giving rise to greater processing difficulty. We conclude with a discussion of empirical and theoretical consequences of our findings. PMID- 10723710 TI - Discourse before gender: an event-related brain potential study on the interplay of semantic and syntactic information during spoken language understanding. AB - A study is presented on the effects of discourse-semantic and lexical-syntactic information during spoken sentence processing. Event-related brain potentials (ERPs) were registered while subjects listened to discourses that ended in a sentence with a temporary syntactic ambiguity. The prior discourse-semantic information biased toward one analysis of the temporary ambiguity, whereas the lexical-syntactic information allowed only for the alternative analysis. The ERP results show that discourse-semantic information can momentarily take precedence over syntactic information, even if this violates grammatical gender agreement rules. PMID- 10723711 TI - Agreement checking in comprehension: evidence from relative clauses. AB - Work on agreement processing in comprehension in English has provided evidence that the plural feature is "specified" or "marked" and that the syntactic structure of the subject plays a role in checking features. It is argued here that the structure intervening between the subject and verb also plays a role in checking and that there is pressure to check features immediately. A self-paced reading study tested this proposal in a previously unstudied configuration: agreement between the verb in a subject relative clause (RC) and the relative clause head (e.g., the niece of the actors who was/were...). According to the Construal Hypothesis (Frazier and Clifton, 1995), the structural relation between the RC and head is not immediately fully specified (unlike, for example, the relation between the subject and verb of a main clause). Thus, at the point when an agreeing verb is processed, the structure underlying the check may not yet be fully specified. This assumption about structural processing, together with the proposal for agreement checking in comprehension predicts (1) that there will be a large asymmetry between the processing of singular and plural agreeing verbs in the RC (larger than those seen in direct subject-verb predication) and (2) that there could be a role for agreement in determining RC attachment. Clear evidence for the first prediction was found: evidence for the second was inconclusive. PMID- 10723712 TI - Case matching and relative clause attachment. AB - Two accounts of relative clause attachment will be discussed, the case-matching hypothesis proposed by Sauerland and Gibson (1998) and the attachment-binding dualism (Hemforth et al., in press a, b). While the case-matching hypothesis predicts that relative clauses are preferentially attached to NPs whose case matches that of the relative pronoun, attachment binding predicts that NPs are preferentially attached to the most salient host, that is NP1 in constructions with two NPs. We conducted two off-line studies, one sentence completion task and one magnitude estimation experiment using subject (nominative pronoun) and object (accusative pronoun) relative clauses that can be attached to either of the two nouns in a complex subject (NP1 = nominative, NP2 = genitive) or object NP (NP1 = accusative, NP2 = genitive). While attachment binding predicts an across-the board NP1 preference, the case-matching hypothesis predicts an NP1 prefence only in the case of subject (object) NPs followed by subject (object) relative clauses. The results of both experiments provide evidence for attachment binding and against case matching. PMID- 10723713 TI - Linear versus hierarchical agreement feature processing in comprehension. AB - Two experiments examined whether syntactic number features are tracked during comprehension with a linear slot-based memory system or with a hierarchical feature-passing system. In a construction such as The pond near the trail(s) for the horse(s) was ..., a linear account of subject-number tracking predicts greater interference from horses (N3), whereas a hierarchical account predicts greater interference from trails (N2). Experiment 1 used singular-head subject noun phrases (e.g., pond) and showed equal interference from N2 and N3, failing to differentiate between linear and hierarchical accounts. Experiment 2 used plural-head subjects and revealed more interference from N2 than N3. The pattern across the experiments accords with the ideas that (1) feature-tracking is hierarchical (e.g., Vigliocco & Nicol, 1997), (2) plurals are marked (e.g., Eberhard, 1997), and (3) subject-number information decays across intervening number-marked elements. PMID- 10723714 TI - Word frequency effects on the processing of subject-verb number agreement. AB - This paper examines the role of word frequency in the computation of subject-verb number agreement. Previous research in both production and comprehension has demonstrated that processing difficulties can arise in sentence structures containing a singular subject noun followed by a plural "distractor" noun, as in The key to the cabinets.... A whole sentence reading task was employed to determine whether the relative frequency with which the distractor noun appears in its singular or plural form would affect the degree of processing difficulty experienced by readers. Results suggest that the agreement process is, indeed, sensitive to this factor and this finding is compatible with activation-based accounts of the implementation of number agreement. PMID- 10723715 TI - Pleural tissue hyaluronan produced by postmortem ventilation in rabbits. AB - We developed a method that used Alcian blue bound to hyaluronan to measure pleural hyaluronan in rabbits postmortem. Rabbits were killed, then ventilated with 21% O2--5% CO2--74% N2 for 3 h. The pleural liquid was removed by suction and 5 ml Alcian blue stock solution (0.33 mg/ml, 3.3 pH) was injected into each chest cavity. After 10 min, the Alcian blue solution was removed and the unbound Alcian blue solution (supernatant) separated by centrifugation and filtration. The supernatant transmissibility (T) was measured spectrophotometrically at 613 nm. Supernatant Alcian blue concentration (Cab) was obtained from a calibration curve of T versus dilutions of stock solution Cab. Alcian blue bound to pleural tissue hyaluronan was obtained by subtracting supernatant Cab from stock solution Cab. Pleural tissue hyaluronan was obtained from a calibration curve of hyaluronan versus Alcian blue bound to hyaluronan. Compared with control rabbits, pleural tissue hyaluronan (0.21 +/- 0.04 mg/kg) increased twofold, whereas pleural liquid volume decreased by 30% after 3 h of ventilation. Pleural effusions present 3 h postmortem without ventilation did not change pleural tissue hyaluronan from control values. Thus ventilation-induced pleural liquid shear stress, not increased filtration, was the stimulus for the increased hyaluronan produced from pleural mesothelial cells. PMID- 10723716 TI - The in vitro efficacy of varidase versus streptokinase or urokinase for liquefying thick purulent exudative material from loculated empyema. AB - Patients with loculated parapneumonic effusion or empyema are sometimes treated with streptokinase or urokinase in an attempt to facilitate pleural fluid drainage by liquefying the pleural exudate and destroying the fibrin membranes producing the loculation. This study evaluated the effectiveness of streptokinase, urokinase, and Varidase (the combination of streptokinase and streptodornase) in liquefying gummy, purulent, exudative material from loculated empyemas. An empyema was created by injecting 10(8) Pasteurella multocida bacteria into the pleural space of New Zealand white rabbits. Twenty specimens, each containing 0.5 g of purulent material obtained 5 days after empyema induction, were placed in test tubes. Streptokinase (15,000 IU), urokinase (10,000 IU), Varidase (4,000-15,000 IU streptodornase + 15,000 IU streptokinase) or saline was added to five sets of four test tubes each. The amount of nonliquefied material that remained after incubation with the fibrinolytic agents was quantitated. Over the 6-h incubation period, the amount of nonliquefied material decreased from 0.5 g to 0.02 g in the Varidase group but never decreased to less than 0.4 g in any of the other three treatment groups. Liquefaction of thick pleural exudates from rabbits with empyema can be achieved with Varidase but not with streptokinase or urokinase. PMID- 10723717 TI - The effects of intrapleural polyclonal anti-tumor necrosis factor alpha (TNF alpha) Fab fragments on pleurodesis in rabbits. AB - The mechanism by which various agents produce a pleurodesis is unknown. The purpose of this project was to determine whether the pleurodesis that results from the intrapleural administration of talc or doxycycline depends on tumor necrosis factor alpha (TNF alpha). In a randomized, blinded, placebo-controlled study, 34 New Zealand white rabbits were given 400 mg talc or 10 mg/kg doxycycline intrapleurally as a sclerosant through a chest tube. Half the rabbits in each group were also given 2,000 units of ovine, polyclonal affinity purified anti-TNF alpha Fab, or saline as placebo immediately after and 12 h after the injection of the sclerosant. Chest tubes were aspirated at 12-h intervals until their removal at 4 days. Rabbits were killed at 28 days. The pleural fluid volume, cell counts, lactate dehydrogenase (LDH) and pleurodesis scores were compared among groups. Both talc and doxycycline produced an exudative pleural effusion. The pleural fluid volume and the pleural fluid LDH levels were significantly (p < 0.05) greater in the group given doxycycline. The administration of anti-TNF alpha Fab had no significant effect on pleural fluid volume or leukocyte count in either group. However, the administration of anti TNF alpha Fab resulted in a significant decrease (p = 0.004) in the pleurodesis score for the animals given talc (3.2 +/- 0.8 without Fab and 1.8 +/- 0.9 with Fab). In contrast, the pleurodesis score was virtually identical in the doxycycline group with (3.5 +/- 0.5) and without (3.4 +/- 0.7) Fab. The administration of anti-TNF alpha Fab diminishes the pleurodesis induced by talc but not that resulting from doxycycline. These findings suggest that different mechanisms are involved with the two different sclerosants. PMID- 10723718 TI - The effect on sound generation of varying both gas flow rate and the viscosity of sputum-like gel in a simple tubular model. AB - Gas flows of 2, 3, and 4 L/min were directed through a sputum-like gel with viscosities of 100, 150, and 200 P and placed in a tube similar in diameter to a human segmental bronchus (4 mm), which was immersed in a bath of water. The sound produced by gas flow through the gel was recorded with a hydrophone. Sound data were subjected to time-expanded waveforms and fast Fourier transform (FFT) analysis. This study demonstrated that the number of crackles generated was directly related to the flow rate and inversely related to gel viscosity. The initial deflection width (IDW), two-cycle duration (2 CD), and peak-to-peak amplitude of crackles were significantly affected by the gas flow rate but not the viscosity of the gel. A lower gas flow rate generated crackles with longer IDW and 2 CD, but higher gas flow rates generated crackles with higher amplitude. Peak sound intensity measured from FFT increased as flow rate increased but decreased as the viscosity of the gel increased. At low gas flows, no gel-induced crackle sound was generated within the data capture window when the most viscous gel was examined. A digital video image of gas flow through the gel was captured, and this confirmed the absence of bubbles or slug formation at low flows through 200 P gel during the 3 seconds of data acquisition. This study describes some characteristics of crackles generated from different combinations of gas flow and gel viscosity and suggests that "coarse crackles" results from the explosion of gas bubbles in pulmonary secretions. Health care practitioners should consider the combined effect of rate of inspiratory gas flow and sputum viscosity during auscultation of patients' lungs. PMID- 10723719 TI - Interaction of vagal lung afferents with inhalation of histamine aerosol in anesthetized dogs. AB - In seven alpha-chloralose anesthetized dogs we examined the contribution of lung afferents to the rapid, shallow breathing induced by inhalation of 10 breaths of histamine aerosol. In four spontaneously breathing dogs, the inhalation of histamine caused an increased respiratory frequency, decreased tidal volume, and decreased dynamic lung compliance. Selective blockade of pulmonary C-fibers abolished a reflex-induced increase in respiratory frequency but did not significantly affect the reductions in tidal volume or lung compliance. Terbutaline treatment in combination with C-fiber blockade abolished the reductions in tidal volume and lung compliance induced by histamine. In three separate alpha-chloralose anesthetized, open-chest, mechanically ventilated dogs, we recorded an increase in the inspiratory activity of rapidly adapting pulmonary stretch receptors (RARs) induced by the inhalation of histamine aerosol. Selective C-fiber blockade abolished histamine-induced increases in RAR activity while only partially attenuating reductions in lung compliance. We conclude that the increase in RAR activity induced by histamine depends on intact C-fibers and not on a direct effect of histamine on RARs or an indirect effect of histamine reducing lung compliance. In addition, our data illustrate the multiple interactions that occur between the various vagal afferents and their roles in the reflexes induced by histamine inhalation. PMID- 10723720 TI - Hyperoxia causes an increase in antioxidant enzyme activity in adult and fetal rat type II pneumocytes. AB - It is well known that exposure to hyperoxia results in lung inflammation and damage, which leads to the development of chronic lung disease. Previous studies have shown increased activities of antioxidant enzymes (AOE) in lung tissue from animals exposed to hyperoxia. We propose the hypothesis that the fetal type II pneumocytes (TIIP) would be resistant to oxygen toxicity by virtue of increasing AOE activity on exposure to hyperoxia. The aim of this study was to measure the activities of catalase, glutathione reductase, glutathione peroxidase (GPX), and cytosolic superoxide dismutase (SOD) in cultures of adult and fetal rat TIIP exposed to 95% oxygen for 24 h. Control cells were incubated in room air. Hyperoxia exposure resulted in 53.4 +/- 1.2% of control viability (mean +/- S.E.M.; p = 0.001) in the adult TIIP with a significant threefold increase in the activities of all the AOE. The fetal TIIP were more resistant to hyperoxia (99.4 +/- 6.1% of control viability). However, in the fetal TIIP, only SOD and GPX levels were significantly increased (fourfold and 2.3-fold, respectively) compared with fetal controls. We conclude that fetal TIIP are more resistant to hyperoxia than adult TIIP in terms of viability; other protective antioxidant factors might account for the better survival of fetal TIIP in hyperoxia. PMID- 10723721 TI - The mechanism by which IGFBP-5 exerts anabolic effects in bone. PMID- 10723722 TI - Comparative sequence analysis of the MECP2-locus in human and mouse reveals new transcribed regions. AB - Comparative sequence analysis facilitates the identification of evolutionarily conserved regions, that is, gene-regulatory elements, which can not be detected by analyzing one species only. Sequencing of a 152-kb region on human Chromosome (Chr) Xq28 and of the synthenic 123 kb on mouse Chr XC identified the MECP2/Mecp2 locus, which is flanked by the gene coding for Interleukin-1 receptor associated kinase (IRAK/Il1rak) and the red opsin gene (RCP/Rsvp). By comparative sequence analysis, we identified a previously unknown, non-coding 5' exon embedded in a CpG island associated with MECP2/Mecp2. Thus, the MECP2/Mecp2 gene is comprised of four exons instead of three. Furthermore, sequence comparison 3' to the previously reported polyadenylation signal revealed a highly conserved region of 8.5 kb terminating in an alternative polyadenylation signal. Northern blot analysis verified the existence of two main transcripts of 1.9 kb and approximately 10 kb, respectively. Both transcripts exhibit tissue-specific expression patterns and have almost identical short half-lifes. The approximately 10-kb transcript corresponds to a giant 3' UTR contained in the fourth exon of MECP2. The long 3' UTR and the newly identified first intron of MECP2/Mecp2 are highly conserved in human and mouse. Furthermore, the human MECP2 locus is heterogeneous with respect to its DNA composition. We postulate that it represents a boundary between two H3 isochores that has not been observed previously. PMID- 10723723 TI - Organization of the mouse microfibril-associated glycoprotein-2 (MAGP-2) gene. AB - A 1.4-kb EST clone encoding mouse microfibril-associated glycoprotein-2 (MAGP-2), identified by its similarity with the reported human cDNA, was used to screen a mouse 129 genomic bacterial artificial chromosome (BAC) library. The mouse gene contains 10 exons spanning 16 kb, located on the distal region of Chromosome (Chr) 6. The exons range in size from 24 to 963 bp, with the ATG located in exon 2. The tenth and largest exon contains 817 bp of 3' untranslated sequence, including a B2 repetitive element. Northern analysis demonstrates abundant expression of MAGP-2 mRNA in skeletal muscle, lung, and heart. Sequence analysis of additional cDNA clones suggests that the two mRNA forms of MAGP-2 in the mouse arise from alternative polyadenylation site usage. The promoter does not contain an obvious TATA box, and the sequence surrounding the start site does not conform to the consensus for an initiator promoter element. Additionally, the mouse promoter contains 22 copies of a CT dinucleotide repeat sequence located approximately 155 bp 5' to exon 1. MAGP-2 gene and compared it with that of the human gene (Hatzinikolas and Gibson 1998). While the mouse and human MAGP-2 proteins are similar in sequence, the promoters for the two genes share little in common. The presence of two mRNA species for MAGP-2 in the mouse raised the possibility that more than one isoform of the protein might be synthesized. We have characterized both mRNA species and determined that they do not code for different variants of the protein. PMID- 10723724 TI - Conditional mutagenesis in mice with heat shock promoter-driven cre transgenes. AB - To explore the potential of a simple and rapid approach for ubiquitous conditional gene disruption, we have generated Cre-producer mouse transgenic lines (Hs-cre1, 6 and 7) expressing a recombinase transgene (cre) from a heat shock gene promoter and tested their performance in Cre-mediated excision of target DNA in crosses with Cre-responder strains carrying loxP-modified alleles of the genes encoding the Huntington's disease gene homolog (Hdh), the epidermal growth factor receptor (Egfr), and the type 1 insulin-like growth factor receptor (Igf1r). Analyses of progeny possessing various transgene/reporter combinations showed that cre expression can occur without heat shock in early embryos, but this constitutive transcription is stochastic and transgene dependent. Thus, Hs cre1 behaves predominantly as a "deleter" strain, since the majority of progeny (approximately 70-85%) exhibit complete recombination, regardless of reporter locus. Lines Hs-cre6 and Hs-cre7, however, function successfully as "mosaicking" strains because, in addition to two extreme classes of progeny with 0% or 100% recombination, they generate an intermediate class of mosaics exhibiting various degrees of partial DNA excision. Notably, the frequency of offspring in each class varies between reporters, but mosaic embryos are consistently obtained in adequate numbers (approximately 30-60%). The Hs-cre6 transgene is also inducible and can be used to introduce mosaicism into adult tissues at preselected developmental times by heat shock treatment of mice with 0% recombination in tail DNA. By bypassing the lethality resulting from some gene knockouts, mosaic embryos and mice make particular mutational analyses possible and are also very useful for the identification of cell lineage-specific gene functions. PMID- 10723725 TI - The bovine alpha-glucosidase gene: coding region, genomic structure, and mutations that cause bovine generalized glycogenosis. AB - We report here cDNA and genomic sequence of the bovine acidic alpha-glucosidase gene, from the initiation codon to the most 3' polyadenylation signal. The 2814 bp coding sequence predicts a 937-amino acid protein, which is highly conserved compared with the human alpha-glucosidase gene (86% and 83% identity respectively). The intron/exon boundaries are also conserved between the two species. Two mutations have been identified in Brahmans, and one in Shorthorns, that lead to generalized glycogenosis. All three mutations result in premature termination of translation. Evidence is also presented for a missense mutation segregating with the Brahman population, which is responsible for a 70-80% reduction in alpha-glucosidase activity. PMID- 10723726 TI - Comparative analysis of the PCOLCE region in Fugu rubripes using a new automated annotation tool. AB - The Japanese pufferfish Fugu rubripes with a genome of about 400 Mb is becoming increasingly recognized as a vertebrate model organism for comparative gene analysis (see Elgar 1996 for review). We have isolated and sequenced two Fugu cosmids spanning a genomic region of 66 kb containing the Fugu homolog to the human PCOLCE-I (Glockner et al. 1998). We then examined if RUMMAGE-DP, a newly developed analysis tool for gene discovery which was designed for human and mouse genomic DNA, can be used for automatic annotation of Fugu genomic sequence. The exon prediction programs contained in RUMMAGE-DP performed better overall for the human sequence than for the Fugu contig. The GENSCAN program was the only exon prediction programme that performed equally well for both organisms. We show that RUMMAGE-DP is very useful in automatic analysis of Fugu sequences. Comparative analysis of the genomic structure of the PCOLCE-I genes in Fugu and human reveals that the exon/intron structure throughout the protein coding region is almost identical. We defined an additional domain based on the high degree of similarity of 26 aa between mammals and Fugu. The PCOLCE-I protein in both organisms contains two highly conserved CUB domains. Exons 6 and 7 are the only coding exons that differ in length between the two species. We assume that these exons do not code for any catalytic domain of the protein. Analysis of the remaining five Fugu genes within the 66 kb interval revealed no conserved synteny with the corresponding human 7q22 region. PMID- 10723727 TI - A revision of the human XIST gene organization and structural comparison with mouse Xist. AB - The XIST gene plays an essential role in X Chromosome (Chr) inactivation during the early development of female humans. It is believed that the XIST gene, not encoding a protein, functions as an RNA. The XIST cDNA is unusually long, as its full length is reported to be 16.5 kilobase pairs (kb). Here, comparison of sequences from the genomic interval downstream to the 3' end of the human XIST gene against the human EST database brought to light a number of human EST sequences that are mapped to the region. Furthermore, PCR amplification of human cDNA libraries and RNA fluorescence in situ hybridization (RNA-FISH) demonstrate that the human XIST gene has additional 2.8 kb downstream sequences which have not been documented as a part of the gene. These data show that the full-length XIST cDNA is, in fact, 19.3 kb, not 16.5 kb as previously reported. The newly defined region contains an intron that may be alternatively spliced and seven polyadenylation signal sequences. Sequences in the newly defined region show overall sequence similarity with the 3' terminal region of mouse Xist, and three subregions exhibit quite high sequence conservation. Interestingly, the new intron spans the first two sub-regions that are absent in one of the two isoforms of mouse Xist. Taken together, we revise the structure of human XIST cDNA and compare cDNA structures between human and mouse XIST/Xist. al. 1992). This gene, called XIST/Xist (X inactive specific transcript), shows several interesting features. First, both human and mouse XIST/Xist cDNA are unusually long, reportedly 16.5 kb and 17.8 kb, respectively (Brown et al. 1992; Hong et al. 1999). Second, the transcript does not seem to encode a protein, on the basis of the lack of a significant open reading frame, absence of the Xist RNA from polysomes, and localization of the transcript in the nucleus (Brockdorff et al. 1992; Brown et al. 1992). Third, the XIST/Xist RNA physically associates with, or 'coats,' the inactive X Chr (Brown et al. 1992; Clemson et al. 1996). Fourth, XIST/Xist transcripts can be observed as early as the four-cell stage, and upon the initiation of X-inactivation, the steady-state level of the transcript rises dramatically, apparently by stabilization of the RNA (Panning et al. 1997; Sheardown et al. 1997). Although the function of XIST/Xist is not known, deletion of the gene leads to failure of X-inactivation, and knock-out mice die around the gastrulation stage (Marahrens et al. 1997; Penny et al. 1996). In this report, we revise the structure of the human XIST cDNA and discuss structural features of the newly defined region. PMID- 10723728 TI - Gene flow of unique sequences between Mus musculus domesticus and Mus spretus. AB - Allelic diversity has been examined from a variety of Mus musculus subspecies and Mus spretus strains by sequencing at a 453-bp unique sequence locus. One M. m. domesticus classic inbred strain, C57BL/KsJ, contained a sequence identical to that in the M. spretus wild-derived inbred strain SEG, and other wild M. spretus isolates. Such a result should have been precluded by the expected divergence between the species unless there has been interspecies gene flow. Examination of C57BL/KsJ for M. spretus-specific repetitive sequences shows that it is neither a mis-identified spretus strain nor a domesticus/spretus hybrid. Thus, in addition to the previously reported presence of small amounts of Mus spretus-specific repetitive DNA in M. m. domesticus, there is a detectable flow of unique sequence between the two species. There was also ancestral polymorphism observed among the spretus alleles. The difficulty of distinguishing ancestral polymorphism from horizontal transfer is discussed. PMID- 10723729 TI - Sox14 maps to mouse chromosome 9 and shows no mutations in the neurological mouse mutants ducky and tippy. PMID- 10723730 TI - The characterization of a mouse mutant that displays abnormal mammary gland development. PMID- 10723731 TI - Molecular and embryological characterization of a new transgene-induced null allele of mouse Brachyury locus. PMID- 10723732 TI - Chromosomal localization of four HSA2 type I loci in river buffalo (Bubalus bubalis, 2n = 50) chromosomes 2q and 12. PMID- 10723733 TI - An integrated physical and genetic map of the PLS locus interval on chromosome 11q14. PMID- 10723734 TI - Cloning, structure and assignment to chromosome 19q13 of the human Kir2.4 inwardly rectifying potassium channel gene (KCNJ14). PMID- 10723735 TI - Evolution of alpha 2-fucosyltransferase genes in primates: relation between an intronic Alu-Y element and red cell expression of ABH antigens. AB - Coding sequences of the paralogous FUT1 (H), FUT2 (Se), and Sec1 alpha 2 fucosyltransferase genes were obtained from different primate species. Analysis of the primate FUT1-like and FUT2-like sequences revealed the absence of the known human inactivating mutations giving rise to the h null alleles of FUT1 and the se null alleles of FUT2. Therefore, most primate FUT1-like and FUT2-like genes potentially code for functional enzymes. The Sec1-like gene encodes for a potentially functional alpha 2-fucosyltransferase enzyme in nonprimate mammals, New World monkeys, and Old World monkeys, but it has been inactivated by a nonsense mutation at codon 325 in the ancestor of humans and African apes (gorillas, chimpanzees). Human and gorilla Sec1's have, in addition, two deletions and one insertion, respectively, 5' of the nonsense mutation leading to proteins shorter than chimpanzee Sec1. Phylogenetic analysis of the available H, Se, and Sec1 mammalian protein sequences demonstrates the existence of three clusters which correspond to the three genes. This suggests that the differentiation of the three genes is rather old and predates the great mammalian radiation. The phylogenetic analysis also suggests that Sec1 has a higher evolutionary rate than FUT2 and FUT1. Finally, we show that an Alu-Y element was inserted in intron 1 of the FUT1 ancestor of humans and apes (chimpanzees, gorillas, orangutans, and gibbons); this Alu-Y element has not been found in monkeys or nonprimate mammals, which lack ABH antigens on red cells. A potential mechanism leading to the red cell expression of the H enzyme in primates, related to the insertion of this Alu-Y sequence, is proposed. PMID- 10723736 TI - Horizontal gene transfer of glycosyl hydrolases of the rumen fungi. AB - By combining analyses of G + C content and patterns of codon usage and constructing phylogenetic trees, we describe the gene transfer of an endoglucanase (celA) from the rumen bacteria Fibrobacter succinogenes to the rumen fungi Orpinomyces joyonii. The strong similarity between different glycosyl hydrolases of rumen fungi and bacteria suggests that most, if not all, of the glycosyl hydrolases of rumen fungi that play an important role in the degradation of cellulose and other plant polysaccharides were acquired by horizontal gene transfer events. This acquisition allows fungi to establish a habitat within a new environmental niche: the rumen of the herbivorous mammals for which cellulose and plant hemicellulose constitute the main raw nutritive substrate. PMID- 10723737 TI - The phytochrome gene family in tomato and the rapid differential evolution of this family in angiosperms. AB - A reexamination of the genome of the tomato (renamed Solanum lycopersicum L.) indicates that it contains five, or at most perhaps six, phytochrome genes (PHY), each encoding a different apoprotein (PHY). Five previously identified tomato PHY genes have been designated PHYA, PHYB1, PHYB2, PHYE, and PHYF. A molecular phylogenetic analysis is consistent with the hypothesis that the angiosperm PHY family is composed of four subfamilies (A, B, C/F, and E). Southern analyses indicate that the tomato genome does not contain both a PHYC and a PHYF. Molecular phylogenetic analyses presented here, which utilize for the first time full-length PHY sequences from two completely characterized angiosperm gene families, indicate that tomato PHYF is probably an ortholog of Arabidopsis PHYC. They also confirm that the angiosperm PHY family is undergoing relatively rapid differential evolution. Assuming PHYF is an ortholog of PHYC, PHY genes in eudicots are evolving (Ka/site) at 1.52-2.79 times the rate calculated as average for other plant nuclear genes. Again assuming PHYF is an ortholog of PHYC, the rate of evolution of the C and E subfamilies is at least 1.33 times the rate of the A and B subfamilies. PHYA and PHYB in eudicots are evolving at least 1.45 times as fast as their counterparts in the Poaceae. PHY functional domains also exhibit different evolutionary rates. The C-terminal region of angiosperm PHY (codons 800-1105) is evolving at least 2.11 times as fast as the photosensory domain (codons 200-500). The central region of a domain essential for phytochrome signal transduction (codons 652-712) is also evolving rapidly. Nonsynonymous substitutions occur in this region at 2.03-3.75 times the average rate for plant nuclear genes. It is not known if this rapid evolution results from selective pressure or from the absence of evolutionary constraint. PMID- 10723738 TI - Tempo and mode of human and simian T-lymphotropic virus (HTLV/STLV) evolution revealed by analyses of full-genome sequences. AB - We investigated the tempo and mode of evolution of the primate T-lymphotropic viruses (PTLVs). Several different models of nucleotide substitution were tested on a general phylogenetic tree obtained using the 20 full-genome HTLV/STLV sequences available. The likelihood ratio test showed that the Tamura and Nei model with discrete gamma-distributed rates among sites is the best-fitting substitution model. The heterogeneity of nucleotide substitution rates along the PTLV genome was further investigated for different genes and at different codon positions (cdp's). Tests of rate constancy showed that different PTLV lineages evolve at different rates when first and second cdp's are considered, but the molecular-clock hypothesis holds for some PTLV lineages when the third cdp is used. Negative selection was evident throughout the genome. However, in the gp46 region, a small fragment subjected to positive selection was identified using a Monte Carlo simulation based on a likelihood method. Employing correlations of the virus divergence times with anthropologically documented migrations of their host, a possible timescale was estimated for each important node of the PTLV tree. The obtained results on these slow-evolving viruses could be used to fill gaps in the historical records of some of the host species. In particular, the HTLV-I/STLV-I history might suggest a simian migration from Asia to Africa not much earlier than 19,500-60,000 years ago. PMID- 10723739 TI - Molecular adaptation of alanine:glyoxylate aminotransferase targeting in primates. AB - The intermediary metabolic enzyme alanine:glyoxylate aminotransferase (AGT) is targeted to different organelles (mitochondria and/or peroxisomes) in different species. Possibly under the influence of dietary selection pressure, the subcellular distribution of AGT has changed on at least eight occasions during the evolution of mammals. AGT targeting is dependent on the variable use of two alternative transcription and translation initiation sites which determine whether or not the region encoding the N-terminal mitochondrial targeting sequence is contained within the open reading frame. In the present study, we sequenced the 5' region of the AGT gene, including both ancestral translation start sites, for 11 anthropoid primates and compared the results with data already available for two others. We show that while the more 3' of the two translation start sites is maintained in all species, the more 5' site has been lost in six species (five of seven catarrhines and one of six platyrrhines). In addition, the remaining two catarrhines, which have maintained the 5' translation start site, are predicted to have lost mitochondrial targeting by a different mechanism, possibly loss of the more 5' transcription start site. Analysis of the relative frequencies of nonsynonymous and synonymous mutations in the region encoding the extant or ancestral mitochondrial targeting sequences led us to suggest that there has been recent strong positive selection pressure to lose, or decrease the efficiency of, mitochondrial AGT targeting in several anthropoid lineages, and that the loss of mitochondrial targeting in this group of mammals is likely to have occurred on at least four, and possibly five, separate occasions. PMID- 10723740 TI - On the optimization principle in phylogenetic analysis and the minimum-evolution criterion. AB - This paper discusses the optimization principle in phylogenetic analysis, in the case of distance data. We argue that the use of this principle cannot be called into question, except for computing time reasons. We show that the minimum evolution criterion is not perfectly suited for distance data estimated from sequences, and we present another approach, implemented in the BIONJ algorithm, which allows the data features to be taken into account, while being less demanding in computing time. Simulations show that BIONJ significantly outperforms NJ. PMID- 10723741 TI - Frequent assimilation of mitochondrial DNA by grasshopper nuclear genomes. AB - Multiple copies of mitochondrial-like DNA were found in the brown mountain grasshopper, Podisma pedestris (Orthoptera: Acrididae), paralogous to COI and ND5 regions. The same was discovered using the ND5 regions of nine other grasshopper species from four separate subfamilies (Podisminae, Calliptaminae, Cyrtacanthacridinae, and Gomphocerinae). The extra ND5-like sequences were shown to be nuclear in the desert locust, Schistocerca gregaria (Cyrtacanthacridinae), and probably so in P. pedestris and an Italopodisma sp. (Podisminae). Eighty seven different ND5-like nuclear mitochondrial pseudogenes (Numts) were sequenced from 12 grasshopper individuals. Different nuclear mitochondrial pseudogenes, if descended from the same mitochondrial immigrant, will have diverged from each other under no selective constraints because of their loss of functionality. Evidence of selective constraints in the differences between any two Numt sequences (e.g., if most differences are at third positions of codons) implies that they have separate mitochondrial origins. Through pairwise comparisons of pseudogene sequences, it was established that there have been at least 12 separate mtDNA integrations into P. pedestris nuclear genomes. This is the highest reported rate of horizontal transfer between organellar and nuclear genomes within a single animal species. The occurrence of numerous mitochondrial pseudogenes in nuclear genomes derived from separate integration events appears to be a common phenomenon among grasshoppers. More than one type of mechanism appears to have been involved in generating the observed grasshopper Numts. PMID- 10723742 TI - Strand symmetry around the beta-globin origin of replication in primates. AB - Certain mutations are known to occur with differing frequencies on the leading and lagging strands of DNA. The extent to which these mutational biases affect the sequences of higher eukaryotes has been difficult to ascertain because the positions of most replication origins are not known, making it impossible to distinguish between the leading and lagging strands. To resolve whether strand biases influence the evolution of primate sequences, we compared the substitution patterns in noncoding regions adjacent to an origin of replication identified within the beta-globin complex. Although there was limited asymmetry around the beta-globin origin of replication, patterns of substitutions do not support the existence of a mutational bias between the leading and lagging strands of chromosomal DNA replication in primates. PMID- 10723743 TI - Population genetics of the porB gene of Neisseria gonorrhoeae: different dynamics in different homology groups. AB - The porB locus codes for the major outer membrane protein of Neisseria gonorrhoeae. Alleles of this locus have been assigned to two homology groups based on close sequence and immunological relationships and are designated as either PIA or PIB. Several population parameters were estimated and compared among these two groups using a data set of 22 PIA sequences and 91 PIB sequences obtained from diverse geographic localities and from time periods spanning approximately 50 years. Recombination appears to be extensive in the porB gene. While the recombination rates are similar for the PIA and PIB sequences, the relative contribution of recombination to genetic diversity is higher for the PIA sequences. Alleles belonging to the PIB group show greater genetic diversity than do those in the PIA group. Although phylogenetic analysis did not reveal temporal or geographic clustering of sequences, estimates of gene flow and the fixation index suggested that PIB sequences exhibit population substructure based on geographic locality. Selection acts in these homology groups in a different way. While positive Darwinian selection is the dominant force driving the evolution of the PIA sequences, purifying selection operates also on the PIB sequences. These differences may be attributable to the greater propensity of PIA strains, as compared with PIB strains, to cause disseminated gonococcal infection, which would expose the former to intense selection pressure from the host immune system. The molecular evolution of Neisseria gonorrhoeae seems to be driven by the simultaneous action of selection and recombination, but under different rates and selection pressures for the PIA and PIB homology groups. PMID- 10723744 TI - Relative patterns and rates of evolution in heron nuclear and mitochondrial DNA. AB - Mitochondrial cytochrome b sequence data from 15 species of herons (Aves: Ardeidae), representing 13 genera, were compared with DNA hybridization data of single-copy nuclear DNA (scnDNA) from the same species in a taxonomic congruence assessment of heron phylogeny. The two data sets produced a partially resolved, completely congruent estimate of phylogeny with the following basic structure: (Tigrisoma, Cochlearius, (((Zebrilus, (Ixobrychus, Botaurus)), (((Ardea, Casmerodius), Bubulcus), ((Egretta thula, Egretta caerulea, Egretta tricolor), Syrigma), Butorides, Nycticorax, Nyctanassa)))). Because congruence indicated similar phylogenetic information in the two data sets, we used the relatively unsaturated DNA hybridization distances as surrogates of time to examine graphically the patterns and rates of change in cytochrome b distances. Cytochrome b distances were computed either from whole sequences or from partitioned sequences consisting of transitions, transversions, specific codon site positions, or specific protein-coding regions. These graphical comparisons indicated that unpartitioned cytochrome b has evolved at 5-10 times the rate of scnDNA. Third-position transversions appeared to offer the most useful sequence partition for phylogenetic analysis because of their relatively fast rate of substitution (two times that of scnDNA) and negligible saturation. We also examined lineage-based rates of evolution by comparing branch length patterns between the nuclear and cytochrome b trees. The degree of correlation in corresponding branch lengths between cytochrome b and DNA hybridization trees depended on DNA sequence partitioning. When cytochrome b sequences were not partitioned, branch lengths in the cytochrome b and DNA hybridization trees were not correlated. However, when cytochrome b sequences were reduced to third position transversions (i.e., unsaturated, relatively fast changing data), branch lengths were correlated. This finding suggests that lineage-based rates of DNA evolution in nuclear and mitochondrial genomes are influenced by common causes. PMID- 10723745 TI - The early history of modern birds inferred from DNA sequences of nuclear and mitochondrial ribosomal genes. AB - The traditional view of avian evolution places ratites and tinamous at the base of the phylogenetic tree of modern birds (Neornithes). In contrast, most recent molecular studies suggest that neognathous perching birds (Passeriformes) compose the oldest lineage of modern birds. Here, we report significant molecular support for the traditional view of neognath monophyly based on sequence analyses of nuclear and mitochondrial DNA (4.4 kb) from every modern avian order. Phylogenetic analyses further show that the ducks and gallinaceous birds are each other's closest relatives and together form the basal lineage of neognathous birds. To investigate why other molecular studies sampling fewer orders have reached different conclusions regarding neognath monophyly, we performed jackknife analyses on our mitochondrial data. Those analyses indicated taxon sampling effects when basal galloanserine birds were included in combination with sparse taxon sampling. Our phylogenetic results suggest that the earliest neornithines were heavy-bodied, ground-dwelling, nonmarine birds. This inference, coupled with a fossil bias toward marine environments, provides a possible explanation for the large gap in the early fossil record of birds. PMID- 10723746 TI - Positive selection and propeptide repeats promote rapid interspecific divergence of a gastropod sperm protein. AB - Male-specific proteins have increasingly been reported as targets of positive selection and are of special interest because of the role they may play in the evolution of reproductive isolation. We report the rapid interspecific divergence of cDNA encoding a major acrosomal protein of unknown function (TMAP) of sperm from five species of teguline gastropods. A mitochondrial DNA clock (calibrated by congeneric species divided by the Isthmus of Panama) estimates that these five species diverged 2-10 MYA. Inferred amino acid sequences reveal a propeptide that has diverged rapidly between species. The mature protein has diverged faster still due to high nonsynonymous substitution rates (> 25 nonsynonymous substitutions per site per 10(9) years). cDNA encoding the mature protein (89-100 residues) shows evidence of positive selection (Dn/Ds > 1) for 4 of 10 pairwise species comparisons. cDNA and predicted secondary-structure comparisons suggest that TMAP is neither orthologous nor paralogous to abalone lysin, and thus marks a second, phylogenetically independent, protein subject to strong positive selection in free-spawning marine gastropods. In addition, an internal repeat in one species (Tegula aureotincta) produces a duplicated cleavage site which results in two alternatively processed mature proteins differing by nine amino acid residues. Such alternative processing may provide a mechanism for introducing novel amino acid sequence variation at the amino-termini of proteins. Highly divergent TMAP N-termini from two other tegulines (Tegula regina and Norrisia norrisii) may have originated by such a mechanism. PMID- 10723747 TI - Codon usage and the origin of P elements. PMID- 10723748 TI - [Clinical subtypes and differential diagnosis of fronto-temporal dementia]. PMID- 10723749 TI - [Pick's disease and its related disorders]. PMID- 10723750 TI - [Motor neuron disease with dementia]. PMID- 10723751 TI - [Frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17)]. PMID- 10723752 TI - [Molecular genetics on affective disorder: a review]. PMID- 10723753 TI - [Clinical study of cerebral blood flow in unilateral chronic subdural hematoma measured by 99mTc-HMPAO SPECT]. AB - Cerebral blood flow (CBF) measured by 99mTc-HMPAO SPECT before operation was studied in 60 patients with unilateral chronic subdural hematoma. The regional CBF was measured in 26 regions of the fronto-occipital 10 cortices, putamen, thalamus and cerebellar hemisphere on both sides. Sixty cases with unilateral chronic subdural hematoma were classified into four groups on the basis of clinical symptoms: 17 cases with headache (headache group), 34 cases with hemiparesis (hemiparesis group) and 9 cases with consciousness disturbance or dementia (consciousness disturbance group), and into three groups on the basis of the degree of midline brain shift on MRI: 7 cases of mild shift group, 24 cases of moderate shift group and 29 cases of severe shift group. The average CBF in 60 patients in each region indicated that the regional CBF was reduced in frontal, occipital cortices and cerebellum on the non-hematoma side, and in putamen and thalamus on the hematoma side. In the headache group, the regional CBF reduction on the non-hematoma side was found in only frontal and occipital cortices compared with the corresponding regions on the hematoma side. In the hemiparesis group, the regional CBF was reduced in frontal and occipital cortices on the non hematoma side and in putamen and thalamus on the hematoma side. The part of CBF reduction in both hemispheres was also noted in the hemiparesis group. In the consciousness disturbance group, the CBF reduction was markedly noted in whole brain. The CBF reductions in frontal and occipital cortices on the non-hematoma side and in putamen, thalamus and cerebellum on the hematoma side was not mutually related with the degree of midline brain shift. We concluded that the disturbance of CBF in chronic subdural hematoma was started from frontal and occipital cortices on the non-hematoma side observed in the headache group, and which was extended to putamen and thalamus on the hematoma side and a part of both hemispheres observed in the hemiparesis group. And such disturbance was finally observed as the CBF reductions in whole brain in the consciousness disturbance group. PMID- 10723754 TI - [Evaluation of hemodynamics in cerebral infarction using MRI with FAIR sequence]. AB - In comparison with 99mTc-ECD SPECT, the usefulness for evaluation of cerebral blood flow by the perfusion images using MRI with FAIR sequence was studied in ischemic stroke patients. Among 27 cases, 15 patients showed lacunar infarctions and 12 patients showed cortical infarctions determined by T2 weighted MR images. FAIR images were obtained as single images at the slice position running through the basal ganglia or corona radiata. The inversion times(TI) were varied, ranging from 800 to 1,400 msec. In 15 patients without definite low perfusions in the SPECT study, FAIR images showed sequentially proxymal arterial branches in early phase and distal arterial branches or capillary beds in the cortical tissues in a late phase as the TI was elongated. Nine of the 12 patients with low cerebral perfusions in the SPECT study showed perfusion defects in FAIR imaging. Five of the 12 patients with a small low cerebral perfusion area in the SPECT study showed a delay of the depiction of cortical arteries along with TI elongation. In 3 patients, ischemic lesions demonstrated by the SPECT study was not shown in the MRI study because of motion artifacts. In conclusion, FAIR imagings are considered to be useful in the evaluation of cerebral flow dynamics in the ischemic stroke patients. PMID- 10723755 TI - [Clinical usefulness of diffusion-weighted magnetic resonance imaging in patients with ischemic cerebrovascular disease]. AB - Diffusion-weighted magnetic resonance imaging was performed to determine the detectability of ischemic changes in patients with ischemic cerebrovascular disease. We retrospectively reviewed 103 patients with symptoms suggestive of ischemic cerebrovascular disease. All patients underwent computed tomography, routine magnetic resonance imaging, and diffusion-weighted imaging. Of 103 patients, 18 were imaged within 3 hours after onset, 57 were imaged between 3 and 24 hours, and 29 were imaged between 24 and 144 hours. Eighty-eight patients were diagnosed as ischemic cerebrovascular disease. Magnetic resonance imaging was performed at a 1.0 Tesla clinical machine using single-shot spin-echo/echo-planar imaging sequence. In each case, three sets of DWI with motion-probing gradient pulses in the x, y, and z directions were taken. The detectability of ischemic changes of each imaging modality was compared. DWI detected ischemic changes in 83 of 88 cases with clinical diagnoses of cerebral ischemia(sensitivity; 94.3%). In contrast, DWI showed negative findings in 15 of the 15 patients with diagnoses other than cerebral ischemia(selectivity; 100.0%). DWI detected ischemic changes in 16 out of 18 patients(88.6%) within 3 hours after the onset. In contrast, T 2 weighted image did not detect any ischemic changes in the same period. These results suggest that DWI is considered to be highly useful for the early diagnosis of cerebral ischemia. PMID- 10723756 TI - [The Japanese version of the Calgary Depression Scale for Schizophrenics (JCDSS)]. AB - Depressive features are clearly recognized within the context of schizophrenia, and clarification of the comorbidity has important clinical implications particularly when choosing the best neuroleptics for treatment. However, in some cases, it is difficult to clearly distinguish between depressive symptoms, negative symptoms and extrapyramidal side-effects. Conventionally, the Zung's Self-Rating Depression Scale or Hamilton's Depression Rating Scale has been used as an evaluation scale of depressive symptoms. However, as these scales are designed for assessment of depression in non-psychotic populations, they do not necessarily distinguish depressed from non-depressed psychotic subjects. Recently, Addington et al. developed a new depression scale, the Calgary Depression Scale for Schizophrenics(CDSS), which was designed for the assessment of depression in schizophrenia. CDSS is a nine item structured interview scale, in which each item has a four point measure, each point anchored by descriptors. It has been tested, and it has been shown that there is no overlap with negative or extrapyramidal symptoms. Therefore, for its clinical application, we prepared the Japanese-language version(JCDSS). PMID- 10723757 TI - [An autopsy of parkinsonism after solitary living in Guam Island for 28 years]. AB - The autopsy findings of an 82-year-old man with history of solitary living in the jungle of Guam, the endemic area of parkinsonism-dementia complex(PDC), for 28 years was reported in this paper. When he was 75 years old, about 20 years later to have come back to Japan, he developed parkinsonism. He noticed bradykinesia and was pointed out masked face, rigidity and tremor in his right hand. After 2 years, he was diagnosed as Parkinson's disease under the third degree of Hoehn Yahr criteria. He also showed mild cognitive dysfunction, but no pyramidal signs, muscle atrophy of fasciculation at all. Anti-parkinsonian drugs were effective for his motor symptoms. He admitted at age 82 because of anorexia, and died after 3 months. Neuropathological study disclosed neuronal loss and gliosis with Lewy bodies in the substantia nigra, locus coeruleus and dorsal vagal nucleus. There were cortical type Lewy bodies in the limbic system and scanty amount in the neocortex. A few neurofibrillary tangles(NFT) were found in the hippocampus and parahippocampal gyrus, but no dominancy in the second or third layers of the cerebral cortex as reported in PDC. Senile plaques were not observed at all. Although the exact cause of PDC has not been clarified, environmental factors such as water or food seem to influence on the outcome of PDC. However, the pathological findings of the present case were compatible to those of idiopathic Parkinson's disease. Thus it is a very important fact that the present case was not suffered from PDC in spite of his long residence in the endemic area of Guam. PMID- 10723758 TI - [Malignant lymphoma at the cavernous sinus]. AB - A 71-year-old man developed general fatigue, appetite loss, and headache. Two months later, he noticed diplopia. Examination demonstrated reduced visual acuity and complete ophthalmoplegia of the left eye. Brain MRI disclosed a mass that extended from bilateral cavernous sinus to the clivus. There were left cervical lymphadenopathy and a right abdominal mass. A needle biopsy of the abdominal mass revealed non-Hodgkin lymphoma. Although malignant lymphoma at the cavernous sinus is not common, it should be an important consideration in the differential diagnosis of mass at the cavernous sinus. PMID- 10723759 TI - [A case of cerebral postresuscitation encephalopathy detected by diffusion weighted MR imaging]. PMID- 10723760 TI - [MRI features following status epilepticus due to delayed radiation necrosis]. PMID- 10723761 TI - Dyschromatosis universalis hereditaria: a unique disorder. PMID- 10723762 TI - Factors that may be influencing the prevalence of head lice in British school children. PMID- 10723763 TI - Substituted N-phenylcarbamates as histamine H3 receptor antagonists with improved in vivo potency. AB - Novel substituted N-phenylcarbamates as derivatives of 3-(1 H-imidazol-4 yl)propanol were prepared and tested for their antagonist potency in vitro and in vivo at histamine H3 receptors. Structural modifications with different alkyl and acetyl moieties were performed in an attempt to optimize pharmacodynamic and pharmacokinetic effects. Most compounds are active in a functional test for histamine H3 receptors on rat cerebral cortex synaptosomes as well as in a peripheral model on guinea pig ileum. But only carbamates without too bulky lipophilic residues showed pronounced to high antagonist potency on the enhancement of endogenous histamine in brain after p.o. administration to mice (ED50 values of 5.5 to 0.86 mg.kg-1). The tested compounds presented weak activities at histamine H1, H2, and muscarinic M3 receptors thus demonstrating their H3-receptor selectivity. PMID- 10723764 TI - Synthesis and antimicrobial activity of acyclo C-nucleosides: 3-(alditol-1-yl)-7 oxo-5-phenyl-1,2,4-triazolo[4,3-a]pyrimidines. AB - Condensation of 2-hydrazino-4-oxo-6-phenylpyrimidine (1) with aldopentoses 2a-d or aldohexoses 2e-g gave the corresponding aldehydo-sugar (4-oxo-6 phenylpyrimidin-2-yl)hydrazones 3a-g which were acetylated to the corresponding poly-O-acetyl-aldehydo-sugar (3-acetyl-4-oxo-6-phenylpyrimidin-2-yl)hydrazones 4a g. The latter compounds underwent oxidative cyclization with bromine in acetic acid and in the presence of sodium acetate to the corresponding 8-acetyl-3- (poly O-acetyl-alditol-1-yl)-7-oxo-5-phenyl-1,2,4-triazolo[4,3-a]pyrimid ines 6a-g. Compounds 6a-g were also obtained by consecutive one-pot oxidative cyclization/acetylation in which the parent hydrazones 3a-g were treated with bromine/acetic acid/sodium acetate followed by acetic anhydride. Deacetylation of 6a-g with ammonium hydroxide in methanol gave the title compounds 7a-g. The antibacterial and antifungal activities of compounds 3c, 3f, 7c and 7f are reported. PMID- 10723765 TI - A new method of capillary electrophoresis for metabolites of coumarin. AB - The metabolism of coumarin has been very widely investigated compared with other natural compounds. We know today that genetic variations lead to human groups with a high or low coumarin-metabolism. An effective capillary-electrophoresis method has been developed to study these effects on metabolic patterns. The seven most important metabolites can be analysed in one step. Therefore also small volume samples can be examined for phase-I-reactions (hydroxylation of coumarin) as well as for secondary reactions (e.g. glucuronidation or decomposition of the lactone-structure). PMID- 10723766 TI - Performance of GC-MS and MOSES II, a hybrid modular sensor system, for the quantitative detection of the evaporation of the insect repellent N,N-diethyl-m toluamide from two different matrices. AB - The purpose of this work was to determine the evaporated amount of N,N-diethyl-m toluamide (DEET), a commonly used insect repellent, from two different matrices medium chain triglycerides (MCT) and Polyethylene glycol 400 (PEG), with GC with mass sensitive detection (GC-MS) and a hybrid modular sensor system (MOSES II). Thus, the binary mixtures are equilibrated at 80 degrees C for 45 min and then an aliquot of this headspace is analyzed by GC-MS and a sensory system consisting of an array of eight metal oxide sensors and eight quartz crystal microbalances. Both analytical methods allow a sensitive detection of even small amounts of DEET in the headspace from binary mixtures with the MCT or PEG, respectively. The two additives are found to be very different concerning their behavior to the vapor phase; i.e., the lipophilic (MCT) delivers an almost constant headspace of DEET, while the hydrophilic matrix PEG releases smaller amounts of DEET, which continuously decrease when a multiple headspace extraction is performed. Furthermore, the results reveal that both analytical methods lead to comparable results with the MCT/DEET mixtures whereas differences were seen with mixtures containing PEG. This can be attributed to the interaction of volatile portions of PEG with the sensors of the MOSES II. PMID- 10723767 TI - Fluorescent-labeled ligands for the benzodiazepine receptor. Part 2: The choice of an optimal fluorescent-labeled ligand for benzodiazepine receptor assays. AB - In this article, the binding affinities of the fluorescent-labeled benzodiazepines described in Part 1 are compared to assess the influence of the labeling position and the choice of fluorophore on the binding affinity. This comparison was extended by taking into account the data of other fluorescent labeled benzodiazepines in the literature. The differences in the binding affinities observed could partly be explained by structure-activity relationships (SAR). On the basis of this comparison, fluorescent-labeled desethylflumazenil (Ro15-3890, 19) derivatives were selected as the most suitable labeled ligands in fluorescent receptor assays. A methyl-methoxycoumarin derivative (Mmc-O-CO-(CH2)3 Ro15-3890) (20b) had a Ki-value of 6.5 nM, and a 7-nitrobenz-2-oxa-1,3diazole derivative (NBD-NH-(CH2)3-Ro15-3890, 21) had a Ki-value of 5.7 nM. In order to yield sufficient sensitivity in the final receptor assay, a suitable fluorescent labeled ligand should have a Ki < 10 nM. A further advantage of the above two ligands is that the benzodiazepine moiety has no receptor affinity of its own. Thus, if some hydrolysis of the labeled ligand were to occur, the resulting Ro15 3890 (18) would hardly affect the outcome of the assay. In the second part of this paper the prerequisites of the fluorophore are being examined. In this regard, 20b is preferred, because the coumarin derivative has higher fluorescence intensities in aqueous media than the NBD-derivative. Therefore, 20b was selected as a fluorescent-labeled ligand in the development of a non-radioactive receptor assay for benzodiazepines. PMID- 10723768 TI - Polysaccharide coated niosomes for oral drug delivery: formulation and in vitro stability studies. AB - Non-ionic surfactant vesicles (niosomes) were prepared and appended with a polysaccharide cap using hydrophobic anchors. Hydrophobized polysaccharides, O palmitoyl pullulan (OPPu) and cholesteroyl pullulan (CHPu) were anchored onto propranolol.HCL containing preformed niosomes. The coated niosomes were characterized for average vesicle size, size distribution, shape, encapsulation efficiency and in vitro release profile and were compared with their uncoated counterparts. No significant difference was observed in % encapsulation (P > 0.05 in a rank sum test) of polysaccharide coated and uncoated vesicles. In vitro release studies however, revealed a significant lowering (P < 0.01) of drug release for the coated systems in simulated gastric and intestinal fluids with a biphasic release profile. The influence of the hydrophobized polysaccharide cap on niosomal membrane integrity and stabilization against harsh bio-environment conditions was also investigated. The parameters investigated include detergent and bile (bile salts and fresh-pooled rat bile) challenge, freeze-thaw cycling, osmotic stress, and long term and shelf stability studies. It was seen that at higher bile salt concentrations and detergent content, uncoated niosomes underwent bilayer solubilization into intermediate micellar structures, whereas coated niosomes were able to maintain their structural integrity as reflected from their higher % latency for the entrapped water soluble agent. Similarly, freeze-thaw cycling could not bring any fusion or collapse of the niosomal membrane (unlike uncoated ones). Furthermore, the exceptional shelf stability of the coated vesicles both at 37 +/- 1 degrees and at 4 +/- 1 degrees C establishes the potential of polysaccharide coated niosomes as an oral delivery system for water-soluble agents. Results from OPPu and CHPu coated niosomal systems for their oral stability potential are compared. PMID- 10723769 TI - ["Non-solvent shock agglomeration"--technology of a new alternative method for sustained release of ibuprofen. 5. Production and evaluation of retard formulations]. AB - Conventional and prepared racemic and optically pure substances were assessed with regard to their suitability to be processed into retarded formulations by direct tabletting. In this respect conventional ibuprofen and specially granulated substances usually show only insufficient properties. Fluid bed granulated S(+)ibuprofen, however, can be suitably transformed into slow release forms by using traditional agents (above all cellulose ether); the low bulk and tapped volume of these granulates, however, is a limiting factor. Shock agglomerated ibuprofen (in particular racemic substance) offers all preconditions of dissolution in acid environment correspondingly prepared substances can cause significant release retardation in artificial intestinal juice. Moreover acceptable tablet qualities (e.g. hardness) can be guaranteed. PMID- 10723770 TI - Effect of cyclodextrin complexation on aqueous solubility and photostability of phenothiazine. AB - The effect of cyclodextrin (beta-CD, gamma-CD and substituted beta-CD derivatives) complexation on the solubility and photostability of phenothiazine (Ph) was compared. The phase solubility method was applied to calculate the stability constants of soluble 1:1 or 1:2 inclusion compounds formed between Ph and CDs. Photochemical decomposition in solution of phenothiazine alone and in the presence of beta-CD or beta-CD derivatives, was found to proceed according to the two stage first-order reaction and in the case of gamma-CD, in a single stage reaction. Formation of solid inclusion complexes of Ph with CDs was evaluated using IR, 13C NMR and DSC studies. The influence of the complexation technique in the solid state (kneading, heating and freeze-drying) on the solubility of Ph was compared. It was establish that the improvement in solubility and stability of Ph was dependent on the kind of CD. When the complexation proceeded in solution it was more effective. PMID- 10723771 TI - Drying of steamed Asian ginseng (Panax ginseng) roots by microwave-hot air combination. AB - Steamed Asian ginseng (Panax ginseng) roots were dried by a combined microwave hot air method in a modified experimental microwave oven. Hot air drying was used as a reference method. The drying time to achieve the desired moisture level (10%) as well as the ginsenoside contents and the color of the final product were determined. The ginsenosides Rb1, Rb2, Rc, Rd, Re, Rf, Rg1 and Ro were analyzed by HPLC. Compared with hot air drying, the combined microwave-hot air drying method resulted in a substantial decrease (approximately 30-40%) in drying time and had little influence on the ginsenoside contents and the color of the final product. PMID- 10723772 TI - Inhibition of metallopeptidases by flavonoids and related compounds. AB - To elucidate possible mechanisms of activity in medicinal plants containing flavonoids, the inhibitory potency of twenty flavones, flavonols, flavanones, phenylacrylic acids and various hydroxylated phenylacetic acids on the activity of neutral endopeptidase (NEP; EC 3.4.24.11), angiotensin-converting enzyme (ACE; EC 3.4.15.1) and aminopeptidase N (APN; EC 3.4.11.2) was investigated in vitro. The screening generally resulted that inhibition of these enzymes requires free hydroxyl groups at the flavone molecule. Flavone and methoxylated compounds (sinensetin) were without effects. Flavonoids with free hydroxyl functions in position 3',4' and 5,7 inhibited the activity of NEP (quercetin, luteolin, fisetin), with myricetin (IC50 = 42 microM) as strongest inhibitor. Inhibition of ACE and APN did not depend on this class of compounds and substitution pattern. E.g. 3,4-dihydroxyphenylacetic acid and 4-methylcatechol (urinary metabolites of flavonoids) also inhibited both APN and ACE activity, but not NEP activity. The results demonstrate that some of the pharmacological activities of flavonoids might be related to the inhibition of metallopeptidases responsible for the splitting of regulatory neuropeptides. PMID- 10723773 TI - Free-Wilson analysis of the antibacterial activity of fluoronaphthyridines against various microbes. A new application of indicator variables. AB - The quantitative structure-activity relationship of the antibacterial activity of eighteen fluoronaphthyridines against nine species of bacteria was analyzed by the Free-Wilson method. A new indicator variable, which designates the species of bacteria, was useful in predicting the antibacterial activity against the various microbes. PMID- 10723774 TI - beta-Adrenergic receptor manipulation and acid phosphatase and zinc levels in the ventral prostate of the adult rat. AB - The influence of 15-day treatments with the beta-adrenergic receptor agonist isoproterenol (120 micrograms/kg/d) or the antagonist propranolol (1.00 mg/kg/d) on acid phosphatase and zinc levels in the ventral prostate was examined in intact rats, rats simultaneously injected with dexamethasone (0.25 mg/kg/d) and animals chemically castrated with a single dose of ethane dimethanesulphonate (75 mg/kg). Isoproterenol-treatment significantly increased acid phosphatase concentration in the ventral prostate of intact rats, whereas propranolol prevented a glandular zinc loss induced by dexamethasone administration. These results demonstrate that the levels of both biochemical parameters in the prostate can be altered by beta-adrenergic receptor manipulation. The responsiveness of the two secretory processes is different and depends on the functional status of the ventral prostate. PMID- 10723776 TI - Hirsutinolides from Vernonia cinerascens. AB - The aerial parts of Vernonia cinerascens, collected in Saudi Arabia, yielded a new hirsutinolide, together with three known lactones. The structure of the new compound was elucidated using, 1H NMR, 13C NMR, 1H-1H COSY, 1H-13C HETCOR and HMBC. PMID- 10723775 TI - [Blood titanium levels before and after oral administration titanium dioxide]. AB - The normal titanium levels in the blood of males between 24 and 66 years of age were found to be 11.2 micrograms/l (rsd 4.1). After oral administration of titanium dioxide containing capsules or as powder (anatas) it could be observed that the material can be absorbed from the gastro-intestinal tract. If two titanium dioxide qualities, having different mean particle sizes (0.16 micron and 0.38 micron), are administered orally, the latter shows less absorption, most likely due to agglomeration phenomena. The blood concentration/time correlation shows the type of curves which are characteristic for a persorption mechanism of absorption and reveal a high individual fluctuation. An increase of the administered dose by twice the amount shows only a tendentious response in the corresponding blood levels. The method of analysis was ICP-AES. A pretreatment of the samples in order to eliminate the organic matrix is necessary. PMID- 10723777 TI - [Synthesis of cyclohexanone and 2-cyclohexenone from sodium cyclamate]. PMID- 10723778 TI - Determination of testosterone, testosterone propionate, testosterone enanthate, testosterone undecanoate in pharmaceutical preparations by HPLC. PMID- 10723779 TI - Development of biodegradable poly(alpha-methylmalate) microspheres. PMID- 10723780 TI - Antiinflammatory activity of diaquabis(cresoxyacetato)zinc(II) complexes. PMID- 10723782 TI - 1,2,4-Trihydroxy menthane, a contact allergen from oxidized Australian tea tree oil. PMID- 10723781 TI - Benzodihydrocarbazoles activity on triazole susceptible and resistant Candida sp. PMID- 10723783 TI - Purification of polyphenol oxidase from opium poppy latex. PMID- 10723784 TI - Composition of the essential oil of Micromeria thymifolia (Scop.) Fritsch and its chemical variation. PMID- 10723785 TI - Determination of the burden of care in families of cardiac surgery patients. AB - "Burden of care" is a term that describes the effects of the multifaceted stressors associated with providing care to an ill family member. Descriptions of burden of care in acute care populations, such as families of patients who have had coronary artery bypass grafting, are very limited. The three purposes of this study were to describe the burden of care in families of coronary artery bypass grafting surgery patients, to compare the burden of care in families grouped by length of stay, and to provide evidence for the validity of the Caregiving Burden Scale in acute care populations. A survey was done using a longitudinal design over the first six weeks following coronary artery bypass grafting surgery. The 124 spouses of coronary artery bypass grafting surgery patients who participated reported a moderate degree of burden in caring for post cardiac surgery family members. Providing emotional support, taking over household tasks, and monitoring patients' conditions created the greatest burden for the participants. Length of stay in hospital did not have an impact on burden of care. The analysis of the data supports the validity of the Care-giving Burden Scale when used in the cardiac surgery population. (Prog Cardiovasc). PMID- 10723786 TI - Nursing care of the intraaortic balloon catheter's inner lumen. AB - An inner lumen exists within the intra-aortic balloon for the purpose of introducing a guidewire to facilitate balloon insertion. This inner cannula also can be used for continuous arterial pressure monitoring. However, special precautions are necessary when utilizing this port for arterial monitoring as compared to a conventional radial arterial line. The proximal tip of this arterial pressure monitoring source rests within the proximal aorta in close proximity to the trifurcated vessels coming off the aorta. The possibility of a clot dislodging and entering the trifurcated vessels. Because of the inner lumen's small size and location, special guidelines need to be followed for its care and maintenance. This paper reviews physiological specific to the intraaortic balloon's inner cannula and recommends practice guidelines regarding heparinized or nonheparinized flush solutions, flushing of the inner lumen, and troubleshooting a damped inner cannula arterial waveform. PMID- 10723787 TI - Intraaortic balloon pumping through the common iliac artery: management of the ambulatory intraaortic balloon pump patient. AB - Indications for use of the intraaortic balloon pump have expanded as advances in the treatment of heart disease have continued. The intraaortic balloon pump is the most widely used circulatory assist device inserted as short term or long term therapy. Because the percutaneous femoral artery approach requires bedrest, new techniques for intraaortic balloon pump insertion that allow greater mobility are being developed for patients who require long term ventricular support. The goal of ambulation in these patients is to prevent potential complications associated with prolonged immobility. This paper reviews the use of the common iliac artery as an alternate site for intraaortic balloon pump insertion that allows the patient to ambulate and exercise. Pre and post procedure management as well as potential complications of intraaortic balloon pump insertion are discussed. PMID- 10723788 TI - The effects of hormone replacement therapy and oral contraceptive pills on blood pressure. PMID- 10723789 TI - Root cause analysis: a method of addressing errors and patient risk. PMID- 10723790 TI - Investing in the future of cardiovascular nursing as a research based profession: an international perspective. AB - While we all face problems with promoting research even at a local level we must not lose sight of how much there is to gain from learning from each other. Together, on an international basis, we can make more progress in alleviating the suffering of our patients. The "head in the sand" approach (even on a nice warm beach in Australia or California) will be of little benefit to our patients or ourselves in the future. We must do more to take the good, but rare, examples of international collaboration in cardiovascular nursing and make them commonplace! PMID- 10723791 TI - Why should nurses closely monitor the ECG during insertion or exchange of a central venous catheter? PMID- 10723792 TI - Dementia with stroke and its association with low density lipoprotein levels. PMID- 10723793 TI - Intracellular targeting of signalling proteins. Introduction. PMID- 10723794 TI - Cytoskeletal polarization and redistribution of cell-surface molecules during T cell antigen recognition. AB - T cell antigen recognition is accompanied by cytoskeletal polarization towards the APC and large-scale redistribution of cell surface molecules into 'supramolecular activation clusters' (SMACs), forming an organized contact interface termed the 'immunological synapse' (IS). Molecules are arranged in the IS in a micrometer scale bull's eye pattern with a central accumulation of TCR/peptide-MHC (the cSMAC) surrounded by a peripheral ring of adhesion molecules (the pSMAC). We propose that segregation of cell surface molecules on a much smaller scale initiates TCR triggering, which drives the formation of the IS by active transport processes. IS formation may function as a checkpoint for full T cell activation, integrating information on the presence and quality of TCR ligands and the nature and activation state of the APC. PMID- 10723795 TI - The role of lipid rafts in T cell antigen receptor (TCR) signalling. AB - Plasma membranes of many cell types contain domains enriched in specific lipids and cholesterol, called lipid rafts. In T lymphocytes, key T cell antigen receptor (TCR) signalling molecules associate with rafts, and disrupting raft association of certain of these abrogates TCR signalling. The TCR itself associates with lipid rafts, and TCR cross-linking causes aggregation of raft associated proteins. Furthermore, raft aggregation promotes tyrosine phosphorylation and recruitment of signalling proteins, but excludes the tyrosine phosphatase CD45. Together the data suggest that lipid rafts are important in controlling appropriate protein interactions in resting and activated T cells, and that aggregation of rafts following receptor ligation may be a general mechanism for promoting immune cell signalling. PMID- 10723796 TI - The role of membrane-associated adaptors in T cell receptor signalling. AB - Engagement of the T cell receptor leads to activation of several tyrosine kinases and phosphorylation of many intracellular proteins. This is followed by Ca2+ mobilization and activation of multiple biochemical pathways, including the Ras/MAPK cascade, and several downstream serine/threonine kinases. Membrane associated adaptor proteins play an important role in T cell activation by coupling TCR ligation at the membrane to distal signalling cascades. Several new membrane associated adaptors have been identified in recent years. LAT (linker for activation of T cells) is an adaptor molecule, which following its phosphorylation associates with Grb2, Gads, PLC-gamma 1, and other signalling molecules. The functional importance of this molecule has been demonstrated by the study of LAT-deficient cell lines and LAT-deficient mice. Two other recently identified adaptor proteins, TRIM (T cell receptor interacting molecule) and SIT (SHP2-interacting transmembrane adaptor protein), which constitutively associate with several surface molecules, bind to PI3K and SHP2, respectively, after T cell activation and might also function in the TCR signalling pathway. PMID- 10723797 TI - Intracellular adapter molecules. AB - Lymphocyte antigen receptor engagement leads to the initiation of numerous signal transduction pathways that direct ultimate cellular responses. In recent years, it has become apparent that adapter molecules regulate the coupling of receptor proximal events, such as protein tyrosine kinase activation, with end results such as inducible gene expression and cytoskeletal rearrangements. While adapter molecules possess no intrinsic enzymatic activity, their ability to mediate protein-protein interactions is vital for the integration and propagation of signal transduction cascades in lymphocytes. Recent studies demonstrate that intracellular adapter molecules function as both positive and negative regulators of lymphocyte activation. PMID- 10723799 TI - Ras protein signalling. AB - Ras proteins were identified through their association with cell transformation. Since then they have been shown to regulate cell growth, differentiation and apoptosis, as well as influencing processes such as cell migration and neuronal activity. Ras regulates a number of signalling molecules by translocating them to the plasma membrane for activation. An emerging concept is that Ras acts as a branchpoint in signal transduction because it orchestrates the activity of multiple signalling pathways to regulate diverse cellular functions. This implies a degree of selectivity in the ability of Ras to activate particular arms of each pathway, but the mechanisms by which this is achieved are not known. Ras is also an important regulator of immune function and in this review, we summarise current understanding of Ras regulation and function and discuss some new aspects of Ras signalling where understanding is less clear. PMID- 10723798 TI - Pharmacological regulation of network kinetics by protein kinase C localization. AB - Protein kinase C (PKC) is a conserved family of 11 serine/threonine kinases. Most cell types express multiple members of the family. Because the catalytic sites are homologous, and able to accommodate a broad range of substrates in vitro, specificity in function is dependent on subcellular localization of each isozyme in each cell type. Physiological stimulation can result in major changes in localization of individual PKC isozymes, mediated through binding to specific anchoring proteins. We describe data demonstrating that disruption of such translocations of PKC isozymes by pharmacological agents, peptides, or antibodies, causes profound effects on T cell functions. The pharmacological opportunity provided by distinct kinetic properties of complex assembly is also discussed. PMID- 10723800 TI - Form, function, and regulation of protein tyrosine phosphatases and their involvement in human diseases. AB - Protein tyrosine phosphatases (PTPs) are a family of enzymes that modulate the cellular level of tyrosine phosphorylation. Based on cellular location, they are classified as receptor like or intracellular PTPs. Structure and function studies have led to the understanding of the enzymatic mechanism of this class of enzymes. Proper targeting of PTPs is essential for many cellular signalling events including antigen induced proliferative responses of B and T cells. The physiological significance of PTPs is further unveiled through mice gene knockout studies and human genome sequencing and mapping projects. Several PTPs are shown to be critical in the pathogenesis of human diseases. PMID- 10723801 TI - The I kappa B kinase (IKK) and NF-kappa B: key elements of proinflammatory signalling. AB - NF-kappa B is a heterodimeric transcription factor that plays a key role in inflammatory and immune responses. In nonstimulated cells, NF-kappa B dimers are maintained in the cytoplasm through interaction with inhibitory proteins, the I kappa Bs. In response to cell stimulation, mainly by proinflammatory cytokines, a multisubunit protein kinase, the I kappa B kinase (IKK), is rapidly activated and phosphorylates two critical serines in the N-terminal regulatory domain of the I kappa Bs. Phosphorylated I kappa Bs are recognized by a specific E3 ubiquitin ligase complex and undergo polyubiquitination which targets them for rapid degradation by the 26S proteasome. NF-kappa B dimers, which are spared from degradation, translocate to the nucleus to activate gene transcription. There is strong biochemical and genetic evidence that the IKK complex, which consists of two catalytic subunits, IKK alpha and IKK beta, and a regulatory subunit, IKK gamma, is the master regulator of NF-kappa B-mediated innate immune and inflammatory responses. In the absence of IKK gamma, which normally connects IKK to upstream activators, no IKK or NF-kappa B activation can occur. Surprisingly, however, of the two catalytic subunits, only IKK beta is essential for NF-kappa B activation in response to proinflammatory stimuli. The second catalytic subunit, IKK alpha, plays a critical role in developmental processes, in particular formation and differentiation of the epidermis. PMID- 10723802 TI - Chorion type as a possible influence on the results and interpretation of twin study data. AB - The estimation of genetic effects from twin studies usually relies upon the equal environment assumption--that monozygous (MZ) and dizygous (DZ) twin pairs experience equal similarity of their environments from prenatal experiences through adulthood. However, the sharing of a chorion may make a subset of identical twins more similar, or in some cases, more different, than twins that do not share a chorion. Recent studies suggest monochorionic MZ twins resemble one another more than dichorionic MZ twins in cognitive abilities, personality, and risk for psychiatric disorder. To the extent that prenatal environment affects these characteristics, the traditional twin method will yield biased estimates of genetic and environmental influences. We develop models for quantifying this bias and estimating the influence of chorion type on estimates of heritability. PMID- 10723803 TI - Genetic and environmental causes of variation in basal levels of blood cells. AB - The genetic and environmental determinants of variation in blood cell size and number were investigated in 392 pairs of 12-year-old twins. The following blood cell indices were measured: haemoglobin, red blood cell count, haematocrit, mean corpuscular volume, platelet number, total white cell count, level of neutrophils, monocytes, eosinophils, total lymphocytes, CD3+ lymphocytes, CD4+ lymphocytes, CD8+ lymphocytes, CD19+ lymphocytes, CD56+ lymphocytes and CD4+/CD8+ ratio. Genetic factors contributed significantly to all blood cell measures accounting for between 61 and 96% of variance. Heritability estimates did not differ significantly between males and females, although the sample size of the present study was not large enough to exclude the possibility of sex-limited gene expression. Common environmental factors were important in determining red blood cell count and haematocrit, but were not important in determining basal levels of any white blood cell type. PMID- 10723804 TI - Premature singleton versus a twin or triplet infant death: parental adjustment studied through a personal interview. AB - Parental adjustment following the death of a premature singleton or multiple birth infant has hitherto been studied by mailed questionnaires or telephone survey. In the present study, using an in-depth personal interview, grief reactions and adjustment patterns of nine families who lost a singleton premature infant ('Single Group') were compared with those of nine families who lost one of a premature multiple birth cohort ('Multiple Group'). The interview was conducted 1-4 years after the death of the infant and evaluated specific areas or 'scales' of life adjustment, including individual feelings, relationship between husband and wife, and functioning at home and at work. There was no significant difference between the paternal and maternal level of adjustment of the two groups in any of the studied scales. A positive correlation was found between maternal and paternal grief reaction of the same family in the scales of individual feelings (r = 0.65), relationships between husband and wife (r = 0.70), and functioning at home (r = 0.57). Comparing the father's scale with the mother's scale revealed a significant difference only in the area of 'individual feelings'. The gestational age, maternal bonding during hospitalisation of the infant and the parental attendance at the event of death were significantly associated with the process of parental adjustment. The results of this study support previous reports of similar parental reactions following the demise of a premature singleton or multiple birth infant. Since societal environment may not recognise the need for consolation of these families, care, compassion, and sensitivity should be encouraged in dealing with these parents at the time of their infant' death, and for a long time thereafter. PMID- 10723805 TI - Differences and similarities in the intra-uterine behaviour of monozygotic and dizygotic twins. AB - Diagnostic advances have made it possible to use ultrasonograph to assess placentation and therefore zygosity in utero in the case of monochorionic monozygotic twins. Foetal behaviour of 15 monozygotic and 15 unlike-sexed dizygotic twin pairs was studied serially with ultrasounds from 10 to 22 weeks gestational age. Each twin, regardless of its zygosity, showed individualised behavioural styles. One twin was found to be 'dominant' in the sense of being more active, but less reactive, possibly due to the fewer stimuli being generated by its co-twin. Monozygotic twins, as opposed to dizygotic twins, showed greater similarities in activity and reactivity levels, but were never behaviourally identical and decreased in likeness with increasing age. Our data suggest that so called identical twins are very similar, but not behaviourally identical, from very early in pregnancy. The unequally shared intrauterine environment contributes to putting each monozygotic twin on a progressively distinct behavioural path. PMID- 10723806 TI - Peers, teachers and parents as assessors of the behavioural and emotional problems of twins and their adjustment: the Multidimensional Peer Nomination Inventory. AB - A multidimensional peer nomination inventory (MPNI; 30 items) and parallel versions--MPNI-Teacher and MPNI-Parental Rating Forms (37 items each)--were developed during a major new Finnish study of families with twins. The twins (477 girls and 467 boys) were 12 years old, representing subsets of three nationwide Finnish twin cohorts (b. 1983-1985). They were enrolled in 503 school classes, and the total number of children participating in peer nominations was 12,937. Three main factors were extracted from peer nominations and teacher and parental assessments. Intercorrelating sub-components were found, especially in parental assessments. Scales were formed, accordingly, for Behavioral Problems (including Hyperactivity-Impulsivity, Aggression, and Inattention), Emotional Problems (including Depression and Social Anxiety), and Adjustment (including Constructiveness, Compliance, and Social Activity). A framework for the development of the multidimensional inventory was a model of emotional and behavioural regulation. Peer nominations were most reliable, while parental assessments, although mostly satisfactory, were least reliable. Results provided evidence of concurrent validity of peer-referenced assessment, using teacher assessments as criteria; correlations between assessments of peers and parents were lower. The inventory has discriminative validity. Intra-pair correlations of monozygotic co-twins were higher than correlations of same-sex (SS) and opposite sex (OS) dizygotic (DZ) co-twins for all scales across all three assessors, and peer nominations of both SS and OSDZ co-twins yielded correlations significantly greater than zero for all scales. All scales, except Depression and Social Anxiety, differentiated boys from girls. PMID- 10723807 TI - Twin-to-twin transfusion syndrome: a case report. Antepartum prediction of underlying placental vascular pattern in monochorionic twin pregnancies may be possible. AB - A case of twin-to-twin transfusion syndrome is described. Comparing data of serial antepartum ultrasonography with a haemodynamic model suggests the possibility of predicting the underlying placental vascular anatomy. It is suggested that serial ultrasonography, including full biometry, pulsatility indices of the umbilical arteries, foetal echocardiography, assessment of amniotic fluid indices and foetal bladder filling could serve as ultrasound parameters for pattern recognition of the underlying placental vascular anomaly. Biometry should be plotted serially in a difference/average plot. Future application of such intensive ultrasound monitoring in monochorionic twins, as soon as monochorionicity is established, may distinguish those monochorionic twins who may benefit from treatment from those whom it would be better only to observe. PMID- 10723809 TI - Medical reports and more PMID- 10723808 TI - Kin recognition by olfactory cues: what can twins tell us? PMID- 10723811 TI - Aging and memory decline PMID- 10723810 TI - Twins born apart and together. PMID- 10723812 TI - Foetal origins of pathology risk factors PMID- 10723813 TI - Why do people differ in what and how much they eat? PMID- 10723814 TI - Loss of bone because of immobilisation PMID- 10723815 TI - Reduced smoking: an introduction and review of the evidence. AB - The major questions about reductions in the number of cigarettes-day as a treatment goal are (1) how many smokers can reduce and maintain such reduction, (2) how much compensation will occur, (3) will reduced smoking significantly decrease the risk of smoking and (4) will reduction promote or undermine cessation. Naturalistic studies of smokers who are not trying to stop smoking indicate that a substantial minority of smokers spontaneously reduce their number of cigarettes-day and can maintain significant reductions (-7% to -43%) over long periods of time. Six experimental trials of smokers not interested in quitting were able to induce large reductions in cigarettes-day (-15% to -63%) using behavioral therapy and or nicotine replacement. Reductions in toxin exposure (carbon monoxide) were not as large but still substantial (-21% to -35%). The three studies with long-term follow-ups found little loss of effects over 6-30 months. Although face-valid, there is no direct test of whether reduced smoking will decrease smoking risks and such a study would need to be very large and last for a long time. None of the above-cited studies indicate that reduction undermines the probability of future cessation attempts and several found reduction promotes future cessation. PMID- 10723816 TI - Policy interventions to reduce the harm from smoking. AB - The other papers in this series on reduced smoking discuss interventions focused on individuals. This paper illustrates possible smoking reduction interventions focused on policies rather than individuals. Target 12 of the new WHO Health For All Policy aims to significantly reduce the harm from addictive substances, including tobacco, in all member states by 2015, and the WHO Third Action Plan for Tobacco-Free Europe focuses on reducing the harm from tobacco. These documents recommend five key policy strategies: market regulation, product liability, smoke-free environments, support for smoking cessation and education, public information and public opinion. Interventions such as price increases, restricting availability, advertising bans and product control could all be used to achieve harm reduction. Research on reducing the harm of smoking needs to include policy as well as treatment research. PMID- 10723817 TI - Maximizing help for dissonant smokers. AB - 'Consonant' smokers know and accept the risks associated with tobacco consumption, and do not wish to change their smoking, whereas 'dissonant' smokers are tobacco consumers whose attitudes differ from their behaviour. Dissonant smokers have several options: to quit smoking (the optimal solution), reduce their smoking, switch products or brands, or do nothing. To date, nicotine replacement therapy (NRT) is the best-established medical aid to smoking cessation, but several important factors impact on NRT use. As smokers constitute a diverse group there is a need for various different formulations, some of which will suit certain smokers better than others. Smokers should be allowed to select their preferred products in order to increase compliance, and should also be permitted to combine various products if desired. Adequate dosage regimens should be stressed in order to avoid under-dosing, which is common with NRT. It is also essential that the medical system focuses increasingly on the diagnosis and treatment of those smokers who are unable or unwilling to quit smoking. High nicotine dependence correlates with a high risk of pulmonary and cardiovascular disease; because these smokers cannot quit, cessation efforts have little impact on the incidence of tobacco-related diseases in this population. Additional smoking control interventions, such as smoking reduction therapy, are therefore required to treat this group. Our experience in Vienna shows that these smokers can be targeted through approaches that utilize new messages offering alternatives to cessation. PMID- 10723818 TI - Practical experiences in smoking reduction and cessation. AB - Tobacco use is a major cause of cardiovascular, respiratory and oncological disease. Quitting smoking significantly benefits health, but for highly dependent smokers, unable to overcome their nicotine dependence, the concept of smoking reduction as a method of harm reduction is gaining ground. The University Hospital of Basle, Switzerland, has run double-blind, placebo-controlled smoking cessation and smoking reduction studies: the CEASE trial evaluated the efficacy of the nicotine patch in achieving abstinence, and the Rosette study is evaluating the efficacy and tolerability of the nicotine oral inhaler in smokers who do not wish to quit. Smokers were instructed either to quit smoking (CEASE) or to reduce cigarette consumption by > 50% (Rosette). In both studies, success rates for active treatment versus placebo at 4 months demonstrated that nicotine replacement therapy (NRT) is effective in achieving both smoking cessation and reduction. Current approaches to smoking cessation and reduction at our clinic are discussed. Combination NRT rather than monotherapy is commonly used to achieve both smoking reduction and cessation. Treatment appears to be most effective if subjects are allowed to select their preferred NRT product. There are clear differences in patient populations aiming to quit or reduce, the cessation population being more motivated. Smoking cessation remains the ultimate aim but, if unfeasible, a significant reduction in cigarette consumption is a valid goal. PMID- 10723819 TI - Regulatory imbalance between medicinal and non-medicinal nicotine. AB - Cigarettes are very efficient, but exceedingly 'dirty', nicotine delivery systems. Although nicotine creates dependency, it is the contaminated delivery system that causes tobacco-related harm. With an annual global tobacco market > USD$300 billion, and a large proportion of the > 1 billion tobacco users seeking to avoid the 50% risk of death, there should be a huge market for alternative nicotine delivery systems. A move towards risk reduction could significantly benefit public health, provide consumer choice and allow free market forces to combat the leading cause of preventable death. However, market forces are currently prevented from providing consumers with the risk-reducing products they want because of existing regulatory systems. Tobacco products have been exempted from consumer protection laws, but there are no such exemptions for other nicotine delivery products, e.g. NRT. This has resulted in an exceedingly uneven playing field for nicotine products, with the most harmful products subject to little regulation while the least hazardous products are stringently regulated. In effect the world is upside-down, and nicotine regulatory systems should be reformed in order to maximize the reduction in risk. In addition, regulatory bodies need to: develop nicotine- and tobacco-specific expertise, rapidly evaluate which products should be permitted and decide how these products should be marketed. Appropriate regulatory structures could harness the power of free enterprise in global efforts to control the tobacco epidemic. This can be done through the development of regulatory processes designed to ensure that all nicotine delivery products are considered in relative terms (regardless of source), and ensuring that all regulatory action strives for the greatest practical reductions in risk. PMID- 10723820 TI - Addressing regulatory barriers to licensing nicotine products for smoking reduction. AB - This paper examines the current regulatory obstacles to extending the permitted use of NRT, and to include smoking reduction ways of addressing these. There are major differences between different countries in regulations concerning NRT. These differences appear to be due to different attitudes to cigarette smoking as an issue and different preconceptions about nicotine as an addictive and potentially toxic drug. The paper considers how existing WHO and American Psychiatric Association definitions of mental disorders may be used in submitting proposals for use of NRT as an aid to smoking reduction and how concerns over safety and abuse and dependence liability can be addressed. It also examines what new evidence may be needed. Coordinating the efforts of the pharmaceutical industry, clinicians and researchers will probably be important in moving regulatory authorities further in the direction of accepting NRT for widespread use in smoking reduction. PMID- 10723821 TI - Past as prologue: the future of addiction studies. PMID- 10723822 TI - What works? A review of evaluated alcohol misuse interventions among aboriginal Australians. AB - AIMS: To identify which intervention strategies have been effective in reducing excessive consumption of alcohol, and related harm, among some segments of Australia's Aboriginal population. DESIGN: Items dealing with 'alcohol' and 'evaluation' (27) were identified from the comprehensive electronic data base on Aboriginal alcohol and other drug issues, maintained by Australia's National Centre for Research into the Prevention of Drug Abuse. From these were selected all reports (14) dealing specifically with evaluation of particular intervention projects. These were grouped and systematically reviewed under the broad categories of treatment, health promotion education, acute interventions and supply reduction. FINDINGS: A broad range of intervention strategies has been employed. However, few systematic evaluations have been undertaken and the methodologies employed have been generally insufficient to allow robust generalization. The impact of most interventions appears limited but, in part, this may be a function of inadequate resourcing and programme support. CONCLUSIONS: Despite the limitations of the evaluation reports, several conclusions can be tentatively drawn. It appears there is a need to employ a broader range of treatment models and complementary intervention strategies. Interventions are generally inadequately resourced. There is a suggestion that supply reduction interventions may be effective. Most importantly, there is a pressing need for more rigorous evaluation studies in cooperation with Aboriginal community organizations. PMID- 10723823 TI - Opium in 20th-century Britain: pharmacists, regulation and the people. AB - AIMS: To indicate how the system of pharmaceutical regulation of the sale and use of opium in Great Britain continued throughout the first half of the 20th century. DESIGN: An oral history investigation of community pharmacy in Great Britain (n = 50), together with an analysis of standard pharmaceutical texts. SETTING: Community pharmacies in Great Britain during the 20th century. PARTICIPANTS: Retired community pharmacists with experience of the sale and use of opium during the period. MEASUREMENTS: Oral testimony of retired community pharmacists about the use and sale of opium, and quantitative analysis of numbers of official preparations of opium available during the period. FINDINGS: The popular use of opium continued well after the First World War, and its use as an ingredient of prescribed medicines continued well beyond the introduction of the National Health Service in 1948. CONCLUSIONS: Although the role of pharmacy in the regulation of opiates and other drugs was displaced by medicine following the passage of dangerous drugs legislation in the 1920s, pharmacists continued to play an important part in this regulation, exercising considerable discretion in the process. PMID- 10723824 TI - The impact of programs for high-risk drinkers on population levels of alcohol problems. AB - AIMS: Historically, treatment programs and related activities for alcoholics or high-risk drinkers have been viewed as not relevant to efforts to prevent alcohol problems, and in particular population-based prevention efforts. In this review we consider evidence that high-risk programs may have an impact on population or aggregate levels of these problems. DESIGN: We first summarize recent reviews of the clinical impact of programs for high-risk drinkers, since some level of effectiveness at the individual level is necessary for these programs to have an aggregate level impact. Following that, correlational evidence on the impact of high-risk programs on aggregate problem levels is examined. Estimates of the potential impact of high-risk programs on aggregate problem levels, based on available information on the impact of these programs and the numbers of individuals affected, are then considered, as are estimations of the comparative aggregate level impact of high-risk and consumption reduction strategies. FINDINGS: There is increasing evidence that high-risk programs have beneficial effects for individuals. Available correlational evidence supports the proposal that increases in treatment and AA have contributed to the declines in alcohol related morbidity and mortality observed in some countries in recent years. Studies estimating the recent impact of increases in levels of treatment and AA membership support that interpretation, and studies comparing estimated effects of high-risk and population strategies find similar potential for aggregate effects. CONCLUSIONS: Programs for high-risk drinkers can have beneficial aggregate-level effects and are thus a valuable component of population-based efforts to reduce alcohol problems. PMID- 10723825 TI - Ecological validity or ecological fallacy? Toward an eco-epidemiology for alcohol studies. PMID- 10723826 TI - Can treatment really reduce alcohol problems in the population? PMID- 10723827 TI - Can drug treatment availability affect population health? PMID- 10723828 TI - A bridge to cross the treatment-prevention divide? PMID- 10723829 TI - Putting the past in perspective: formulating questions for the future. PMID- 10723830 TI - Investigating the impact of high-risk programs on aggregate problem indicators: towards a research agenda. A reply to the commentaries. PMID- 10723831 TI - "I already stopped": abstinence prior to treatment. AB - AIMS: To determine pre-treatment abstinence rates among treatment seekers and identify factors associated with pre-treatment abstinence. To evaluate the association between pre-treatment abstinence and subsequent outcome. DESIGN: An observational study using data collected for a randomized, experimental design. SETTING: Conducted with participants immediately after assessment for publicly funded substance abuse treatment at the King County Assessment Center (KCAC) in Seattle. PARTICIPANTS: People referred for outpatient or inpatient treatment by KCAC who had illicit drug use in the previous 90 days (N = 565). Participants waited a median of 12 days (range = 0-108 days) until either treatment entry or waiting-list dropout. MEASUREMENTS: A modified Drug History Questionnaire quantified drug use at baseline, treatment entry or waiting-list dropout and 3 months later. Other measurement methods: Stages of Change Readiness and Treatment Eagerness Scale, participant confidence ratings and KCAC chart review. FINDINGS: Sample-wide, 45% of participants reported abstinence from initial assessment to when they entered or failed to enter treatment. Higher rates of abstinence were associated with shorter waiting periods, less substance use prior to initial assessment and higher scores on change readiness. Pre-treatment abstinence was not associated with either treatment entry or completion. There was a non significant trend towards less improvement in substance use with pre-treatment abstinence, with the greatest effect observed for short waits. CONCLUSIONS: Participants can become abstinent prior to treatment, but this is not a good predictor of treatment entry, completion or outcome. A decisional balance strategy may be a more productive use of client and treatment program energy. PMID- 10723832 TI - Deaths in methadone maintenance treatment in New South Wales, Australia 1990 1995. AB - AIMS: To determine the number and causes of deaths in methadone maintenance treatment (MMT). DESIGN: Cross-sectional survey. SETTING: New South Wales (NSW), Australia. PARTICIPANTS: Two hundred and thirty-eight patients who died while registered in MMT from 1990 to 1995. MEASUREMENTS: Data on number and causes of death in MMT were obtained from data on file at the NSW Health Department, NSW Registry of Births, Deaths and Marriages, and NSW Department of Courts Administration. FINDINGS: The most common cause of death was drug-related (44%), followed by medical illness (24%). Fifty deaths (21%) occurred in the first week of MMT, 88% of which were drug-related. In 92% of these drug-related deaths, there was evidence of polydrug use. In all, 42% of all drug-related deaths occurred during the first week of MMT. Nearly half the cases of drug-related death (46%) in the first week were noted by the medical practitioner at assessment to have a history of polydrug abuse or dependence. Four (9%) drug related cases were prescribed doses of methadone in excess of the-then current national methadone clinical guidelines. CONCLUSION: The first 7 days of MMT is a high-risk period. Inadequate clinical review of subjects' tolerance to methadone and/or subjects' use of other central nervous system (CNS) depressant drugs probably contributed to most of these cases' deaths during induction. The findings from this study reinforce the importance of a thorough drug and alcohol assessment of people seeking MMT, cautious prescribing of methadone, frequent clinical review of patients' tolerance to methadone during induction and education about the dangers of additional drug use during this period. PMID- 10723833 TI - Suicidal intent in non-fatal illicit drug overdose. AB - AIM: To explore suicidal intent among drug users experiencing non-fatal overdose. DESIGN: Semi-structured interviews. SETTING AND PARTICIPANTS: Seventy-seven drug users experiencing non-fatal overdose and attending six hospital accident and emergency departments in two Scottish cities during 1997 and 1998. MEASUREMENTS: The extent of suicidal intent and motivations for intentional overdosing were examined. FINDINGS: The incidence of suicidal intent was high, with 38 respondents (49%) reporting suicidal thoughts or feelings before overdosing. Suicidal actions were significantly associated with a self-reported history of life-time mental health problems and with not using heroin prior to overdosing, but not with other demographic or drug history data. Qualitative data indicated that intentional overdosing was frequently not driven by a clear and unambiguous desire to die. Furthermore, suicidal actions were motivated by a range of psychosocial factors, including: (i) predisposing personal circumstances; (ii) precipitating events; and (iii) poor individual coping strategies. CONCLUSIONS: The issue of suicidal intent needs to be addressed routinely in hospital wards and accident and emergency departments so that the need for support can be assessed. PMID- 10723834 TI - The process of relapse in severely dependent male problem drinkers. AB - AIMS: The aim of the study was to investigate factors hypothesized to influence the relapse process, with a focus on the role of self-efficacy, alcohol dependence and cognitive functioning. DESIGN: The study was conducted in the context of a controlled trial of a relapse prevention programme. Subjects were assessed prior to treatment, at immediate conclusion of treatment and at 6- and 12-month follow-up. SETTING: The study was conducted in an Alcohol Treatment Unit (ATU) in Scotland. PARTICIPANTS: Subjects were 60 male problem drinkers who were patients at the ATU. They were heavy drinkers, with corresponding high levels of alcohol dependence and alcohol-related harm. MEASUREMENTS: The independent variables were post-treatment self-efficacy, alcohol dependence, cognitive functioning, level of depression and alcohol consumption prior to admission to treatment. The dependent variables were post-treatment drinking behaviour and functioning and time to lapse and relapse. FINDINGS: Although the methodology does not allow identification of causality, support was found for the hypothesis that post-treatment self-efficacy was an intervening variable between treatment and outcome. Higher post-treatment self-efficacy predicted better outcome at 6 month follow-up and was associated with a reduced risk of lapse and relapse over the 12-month follow-up. Poorer cognitive functioning was significantly associated with being categorized as a problem drinker at 6-month follow-up and with higher risk of a lapse over the 12-month follow-up. Level of alcohol dependence did not predict outcome. CONCLUSIONS: It was concluded that post-treatment self-efficacy rating is a predictor of treatment outcome and time to lapse and relapse and that cognitive functioning is a predictor of treatment outcome and time to lapse. PMID- 10723835 TI - Prevalence of alcohol problems among elderly patients in a university hospital. AB - AIMS: To assess the prevalence of alcohol abuse and the prevalence of alcohol related discharge diagnosis in an elderly general hospital population. DESIGN: On a randomly selected day, all patients aged 65 years and over admitted to a university hospital were screened. SETTING: University Hospital of Amiens, France. PARTICIPANTS: All patients aged 65 years and over were approached and requested to take part in the study. They were interviewed using the CAGE questionnaire and with a structured questionnaire regarding life-style, and asked about their usual daily alcohol consumption. The medical history of each patient was taken. In total, 612 patients fulfilled the age criteria, but 205 patients (33.6%) had to be excluded owing to predefined exclusion criteria (e.g. dementia, aphasia, terminal illness ...) and 37 patients (6%) refused to participate. FINDINGS: The data were derived from 370 patients. The median age was 79 years; 54% reported no alcohol consumption; 9% of patients scored positive on the CAGE questionnaire. The prevalence of patients with a CAGE questionnaire positive was significantly higher among male patients (17%) than female patients (2.5%). The prevalence of patients with alcohol-related discharge diagnosis was 7%. The frequency of higher socio-economic status or divorced status increased significantly with alcohol consumption. CONCLUSIONS: There may be a substantial prevalence of alcohol problems in elderly hospital patients. Research is needed to examine how generalized this problem is. PMID- 10723836 TI - Effects of using recommended coping strategies on drinking outcome following a brief intervention. AB - AIMS: To explore the unique contribution to outcome drinking of clients' use of six strategies for moderating drinking, after statistically accounting for variance explained by some client and intervention variables. DESIGN: An exploratory hierarchical regression analysis assessed the contributions to variance in drinking outcome of pre-intervention (client characteristics, baseline drinking), assignment (level of assessment, therapist experience) and early follow-up variables. Data came from an experimental trial which evaluated the effect of adding assessment to provision of a self-help book to heavy drinkers. SETTING: Diverse Ontario communities. PARTICIPANTS: Heavy drinkers (99 males, 56 females) were selected from 185 media-recruited applicants who were screened by telephone to exclude cases with severe alcohol-related problems. Their mean (+/- SD) pre-admission weekly quantity of alcohol consumed was 22 +/- 15 drinks. Follow-up rates at 3 and 12 months were 92% and 88%. MEASUREMENTS: Regressed onto weekly quantity at follow-up were: client characteristics, previous measures of weekly quantity, experimental condition and use of the menu of strategies (setting goals for drinking, keeping track of drinking, pacing drinking, planning ahead to avoid heavy drinking, developing free-time activities and coping with problems without drinking). FINDINGS: At 3 months the variables predicting lower weekly quantity were: pre-intervention weekly quantity, developing free-time activities, setting goals for drinking and condition. Lower weekly quantity at 12 months was predicted by lower 3-month and pre-intervention weekly quantity, and regular use of: coping with problems without drinking, setting goals for drinking and keeping track of drinking. CONCLUSIONS: This descriptive study revealed a positive association between level of use of recommended coping strategies at follow-up and drinking outcome. Controlled studies of the effects of strategy use on drinking outcome are therefore warranted. PMID- 10723837 TI - The effect of mode of data collection and of non-response on reported alcohol consumption: a split-sample study in Switzerland. AB - AIMS: To examine (a) effects of different modes of data collection on the reporting of alcohol consumption and non-response rate, and (b) differences in reported consumption between respondents and non-respondents. DESIGN: Two versions of a health questionnaire survey were assigned to two random samples, one version to each sample. Version 1 consisted of a telephone interview without alcohol questions, followed by a mailed questionnaire with alcohol questions. Version 2 consisted of a telephone interview with alcohol questions, followed by a mailed questionnaire without alcohol questions. SETTING: Participants were recruited randomly in eight Swiss cantons. PARTICIPANTS: Five hundred and thirty seven (404) respondents to the telephone interview (and subsequent mailed questionnaire) with version 1, and 451 (360) with version 2. MEASUREMENTS: Alcohol-use variables derived from a quantity-frequency measure. RESULTS: Respondents to the mailed questionnaire (version 2) did not differ significantly in alcohol consumption from non-respondents. Response rate was not affected by inclusion of alcohol questions, but respondents asked by telephone about their alcohol use were more often abstainers and less often hazardous drinkers than respondents to the mailed question. CONCLUSION: The study gives no indication that interviews are refused because alcohol consumption is a questionnaire topic, but suggests that postal questionnaires give slightly greater disclosure of alcohol consumption. PMID- 10723838 TI - Is alcohol really good for the heart? PMID- 10723839 TI - Drug misuse and suicide: a tale of two services. PMID- 10723840 TI - ICAP and the perils of partnership. PMID- 10723841 TI - Critical independence and personal integrity. PMID- 10723842 TI - The new manichaeism in alcohol science. PMID- 10723843 TI - Permission for profits. PMID- 10723844 TI - Partnership and Nordic naivety. PMID- 10723845 TI - Partnership, profits and public health. PMID- 10723846 TI - Real partnerships need trust. PMID- 10723847 TI - Perilous partnerships: a reply. PMID- 10723848 TI - Does exercise aid smoking cessation? A systematic review. AB - AIMS: To assess the effectiveness of exercise-based interventions in smoking cessation. DESIGN: A systematic review was conducted of articles published between 1980 and 1999. The review focused on randomized controlled trials (RCTs) in which the specific effects of exercise on smoking abstinence were examined. The primary dependent variable was smoking abstinence. Other studies which had both exercise programming as an independent variable and smoking behaviour as a dependent variable are briefly discussed. PARTICIPANTS: The review included interventions targeting both healthy individuals and those with specific medical conditions. SETTINGS: The interventions were delivered in both community and inpatient settings. MEASUREMENTS: Information extracted from each article included details of the participants, exercise and smoking cessation programmes, control conditions, exercise adherence rates, length of follow-up and outcomes. FINDINGS: Of the eight trials satisfying our inclusion criteria, only two trials found a positive effect for exercise on smoking abstinence. The others showed no effect. CONCLUSIONS: There is some evidence for exercise aiding smoking cessation. Of the two trials finding a positive effect one was rigorously designed, the other was found to have numerous methodological limitations. Trials showing no effect lacked sensitivity. This was principally because of small sample sizes and inadequate measurement and control of exercise adherence. There is a need for more rigorously designed studies in this area. PMID- 10723849 TI - Communities mobilizing for change on alcohol (CMCA): effects of a randomized trial on arrests and traffic crashes. AB - AIMS: We previously reported effects of the CMCA intervention in reducing social and commercial access to alcohol by youth, and reducing alcohol use by 18-20-year olds. This paper reports on effects of CMCA on arrests and car crashes. DESIGN: CMCA was a group-randomized trial that implemented and evaluated a community organizing effort to change community policies and practices to reduce youth access to alcohol. Seven Midwestern communities were randomly assigned to the intervention condition and eight communities were assigned to the control condition. INTERVENTION: For 2.5 years, a part-time community organizer worked in each of the seven intervention communities with local public officials, enforcement agencies, alcohol merchants, the media, schools and other community groups to reduce youth access to alcohol. MEASUREMENT: We collected annual arrest and quarterly traffic crash data for the years 1987-1995, providing a 6-year baseline and 3 years of data during the intervention. Data were stratified into two target age groups (15-17 and 18-20) and a control group (age 21 and over). Analyses used random-coefficient models because we had repeated observations for each unit of assignment in a group-randomized trial with heterogeneous trends across communities. FINDINGS: We observed net declines in the intervention communities for all arrest and traffic crash indicators. The decline was statistically significant for DUI arrests among 18-20-year-olds and approached significance for DUI arrests and disorderly conduct violations among 15-17-year olds. CONCLUSIONS: Together with previously published results from this study, the results reported here suggest that a community-organizing approach to limit youth access to alcohol may be effective, at least for selected end-points and subgroups. We conclude that this approach may be useful, but that a longer intervention period is required to increase effectiveness. PMID- 10723850 TI - Disulfiram treatment for cocaine dependence in methadone-maintained opioid addicts. AB - AIMS: Cocaine use by patients on methadone maintenance treatment is a widespread problem and is associated with a poorer prognosis. Recent studies have evaluated disulfiram as a treatment for individuals with comorbid alcohol and cocaine abuse. We evaluated the efficacy of disulfiram for cocaine dependence, both with and without co-morbid alcohol abuse, in a group of methadone-maintained opioid addicts. DESIGN: Randomized double-blind, placebo-controlled trial. SETTING: Urban methadone maintenance clinic. PARTICIPANTS: Sixty-seven cocaine-dependent, methadone-maintained, opioid-dependent subjects (52% female; 51% Caucasian). INTERVENTION: Study medication, either disulfiram or placebo, was placed directly in the methadone to ensure compliance for 12 weeks. MEASUREMENTS: Primary outcome measures included weekly assessments of the frequency and quantity of drug and alcohol use, weekly urine toxicology screens and breathalyzer readings. FINDINGS: Disulfiram treated subjects decreased the quantity and frequency of cocaine use significantly more than those treated with placebo. Alcohol use was minimal for all subjects regardless of the medication. CONCLUSIONS: Disulfiram may be an effective pharmacotherapy for cocaine abuse among methadone-maintained opioid addicts, even in those individuals without co-morbid alcohol abuse. Disulfiram inhibits dopamine beta-hydroxylase resulting in an excess of dopamine and decreased synthesis of norepinephrine. Since cocaine is a potent catecholamine re uptake inhibitor, disulfiram may blunt cocaine craving or alter the "high", resulting in a decreased desire to use cocaine. PMID- 10723851 TI - Dexamphetamine substitution in the treatment of amphetamine abuse: an initial investigation. AB - AIMS: Dexamphetamine substitution is a widely practised, yet under-researched and controversial treatment for amphetamine abusers. This study aimed to evaluate the usefulness of substitute prescribing, to both oral and intravenous users, and to find out which factors predict doing well in treatment. In the absence of more rigorous controlled trials, it was hoped that this study would help make some inroads into what is a hitherto unexplored area. DESIGN: The standardized records of 220 users receiving dexamphetamine prescriptions were examined retrospectively. Cross-sectional socio-demographic data, and longitudinal outcome data were obtained for 148 of them. SETTING: The amphetamine users had all attended and received treatment by Cornwall Community Drug Team, in the far South West of England, during the period 1992-96. FINDINGS: Oral and intravenous users had remarkably similar outcomes, with intravenous users making more overall gains in treatment. Over half the injectors stopped injecting, and more than a third within 2 months of coming into treatment. Variables predicting a good outcome differed between oral and intravenous users; although for both groups being female was associated with a slower change in drug-use behaviours, but a longer period in treatment. CONCLUSIONS: Dexamphetamine prescribing appears to be reasonably safe, and is associated with improvements in drug-use. Randomized trials are warranted to determine the specific efficacy of the treatment. PMID- 10723852 TI - Treatment of opioid-dependent pregnant women with buprenorphine. AB - AIMS: To assess the maternal and fetal acceptability of buprenorphine and neonatal abstinence syndrome (NAS) in children born to buprenorphine-maintained mothers. DESIGN AND SETTING: Open-label, flexible dosing, inpatient induction with outpatient maintenance, conducted at the University of Vienna within the existing pregnancy and drug addiction program. PARTICIPANTS: Fifteen opioid dependent pregnant women. INTERVENTION: Sublingual buprenorphine tablets (1-10 mg/day). MEASUREMENTS: Mothers: withdrawal symptoms (Wang Scale), nicotine dependence (Fagerstrom Scale: FTQ) and urinalysis. Neonates: birth outcome and NAS (Finnegan Scale). FINDINGS: All subjects were opioid-, nicotine- and cannabis dependent. Buprenorphine was well tolerated during induction (Wang Score < or = 4) and illicit opioid use was negligible (91% opioid-negative). All maternal, fetal and neonatal safety laboratory measures were within normal limits or not of clinical significance. Mean birth outcome measures including gestational age at delivery (39.6 +/- 1.5 weeks), Apgar scores (1 min = 8.9; 5 min = 9.9; and 10 min = 10), birth weight (3049 +/- 346 g), length (49.8 +/- 1.9 cm) and head circumference (34.1 +/- 1.8 cm) were within normal limits. The NAS was absent, mild (without treatment) and moderate (with treatment) in eight, four and three neonates, respectively. The mean duration of NAS was 1.1 days. CONCLUSIONS: Buprenorphine appears to be well accepted by mother and fetus, and associated with a low incidence of NAS. Further investigation of buprenorphine as a maintenance agent for opioid-dependent pregnant women is needed. PMID- 10723853 TI - The Severity of Dependence Scale (SDS) as screening test for benzodiazepine dependence: SDS validation study. AB - AIMS: To assess the validity of the Severity of Dependence Scale (SDS) as a screening test to detect benzodiazepine dependence in regular benzodiazepine users. METHOD: One hundred regular benzodiazepine users, recruited from neurotic benzodiazepine users attending the Mental Health Outpatient Services of the Canary Islands Health Service, were administered the SDS and responses were compared with the Composite International Diagnostic Interview (CIDI) diagnosis of benzodiazepine dependence. Receiver Operating Characteristic (ROC) analysis was used to determine which cut-off score on SDS allowed the best trade-off between sensitivity and specificity. RESULTS: SDS was shown to have high diagnostic utility, and a score higher than six on the scale appears to be an appropriate threshold for problematic benzodiazepine use. The SDS had a specificity of 94.2% and a sensitivity of 97.9%, and the area under the curve was of 0.991. CONCLUSION: The SDS was found to be a valid brief self-report questionnaire for the assessment of benzodiazepine dependence in patients using benzodiazepines. PMID- 10723854 TI - Gender differences in alcohol consumption and adverse drinking consequences: cross-cultural patterns. AB - AIMS: To examine the consistency and/or variability of gender differences in drinking behavior cross-culturally. DESIGN, SETTING, PARTICIPANTS: Women's and men's responses in 16 general population surveys from 10 countries, analyzed by members of the International Research Group on Gender and Alcohol. MEASUREMENTS: Comparable measures of drinking, versus abstention, typical drinking frequencies and quantities, heavy episodic drinking, intoxication, morning drinking, and alcohol-related family and occupational problems. FINDINGS: Women and men differed little in the probability of currently drinking versus abstaining, but men consistently exceeded women in typical drinking frequencies and quantities and in rates of heavy drinking episodes and adverse drinking consequences, while women were consistently more likely than men to be life-time abstainers. In older age groups, both men and women drank smaller quantities of alcohol and were more likely to stop drinking altogether, but drinking frequencies did not change consistently with age. CONCLUSIONS: A theoretical synthesis proposes that gender roles may amplify biological differences in reactions to alcohol, and that gender differences in drinking behavior may be modified by macrosocial factors that modify gender role contrasts. PMID- 10723855 TI - Attachment in adult daughters of alcoholic fathers. AB - AIM: This study was designed to explore the utility of attachment theory for explaining socio-emotional outcomes in adult daughters of alcoholic fathers (ADAF). It was hypothesized that ADAF would have more insecure attachment organizations than daughters of non-alcoholic parents (non-ADAF), and that ADAF would describe themselves as more disposed towards compulsive care-giving than non-ADAF. DESIGN: ADAF and a matched group of non-ADAF were compared on measures of attachment security and compulsive care-giving. PARTICIPANTS: From a larger sample of 251 female college students, 26 ADAF and a matched group of non-ADAF were identified to participate in the study. SETTING: A large, urban university in the northeastern US. MEASUREMENTS: Participants completed the Adult Attachment Interview and a questionnaire assessing characteristics of compulsive care giving. FINDINGS: As predicted, ADAF had less secure attachment organizations then did non-ADAF. Although no group differences were observed for compulsive care-giving scores, compulsive care-giving was negatively correlated with attachment security for ADAF. CONCLUSIONS: Findings indicate that the concept of attachment may be useful for understanding the developmental consequences of parenting in alcoholic families. PMID- 10723856 TI - Effects of telephone versus face-to-face interview modes on reports of alcohol consumption. AB - AIMS AND DESIGN: In order to assess the effects of survey modality on alcohol consumption estimates, data from two surveys using different interview modes (face-to-face and telephone) were compared on several alcohol measures. SETTING AND PARTICIPANTS: Face-to-face survey data were drawn from the 1990 National Alcohol Survey, while the telephone data came from the 1990 Warning Labels Survey. Both surveys used a probability sampling of the US adult general population in the 48 contiguous states. MEASUREMENTS: Measures of alcohol use derived from an identical "graduated frequencies" series included estimates of any drinking in the past 12 months, overall volume, and heavy (5+) drinking days. FINDINGS: Abstention rates did not differ by survey mode, nor did distributions of alcohol consumption by volume and reported frequency of drinking five or more drinks in a day. Multiple regression models including demographic-mode interaction terms were used to examine how mode effects might differ across demographic subgroups. Lower income respondents were under-represented in the telephone sample, and were associated with lower reports of volume and 5+ days, compared to respondents in the face-to-face mode. CONCLUSIONS: The results suggest that although there are few differences in alcohol consumption estimates by interview mode, telephone samples may need to be supplemented or estimates adjusted by income level in order to attain equivalent results. PMID- 10723857 TI - Opioid-associated effects on oxygen saturation. PMID- 10723858 TI - Impact of genomics on healthcare. Overview. PMID- 10723859 TI - New technologies and DNA resources for high throughput biology. AB - The rapid increase in DNA sequencing information is opening up new opportunities in genetics. The current methods for processing and analysing genetic data are, however, slow and labour intensive. The next wave of genetic analysis will rely on the analysis of DNA variation from large population based cohorts. These studies will provide important new data on population and disease genetics and have the potential to make a significant impact on our current healthcare practices. In order for these studies to deliver, we need to develop a new generation of ultra-rapid DNA technologies which will allow us to generate, capture and efficiently exploit these new data. This chapter describes the recent advances in DNA sequencing and genotyping technologies that will lead to 100-1000 fold increases in our ability to produce the DNA data we need to explore and exploit the new genetic opportunities to the full. PMID- 10723860 TI - DNA diagnostics: goals and challenges. AB - The exponential increase in the discovery of human disease genes over the past 10 years has transformed DNA diagnostics from a minor research-based activity to a major professional operation. Mutation testing and linkage analysis are now used to provide prenatal or postnatal diagnosis for a wide range of monogenic disorders, but robust automated procedures for scanning disease genes for mutations are not yet available. The discovery of genes which confer susceptibility to common disorders is likely to create demand for high throughput testing for specific mutations or clinically relevant polymorphisms. Widespread genetic testing must be supported by adequate genetic counselling and by education of healthcare professionals in order to ensure the appropriate application of this information for the benefit of patients and their families. PMID- 10723861 TI - Disease taxonomy--monogenic muscular dystrophy. AB - The field of the autosomal recessive progressive muscular dystrophies has clarified significantly following the recent elucidation of the genetic and molecular etiology of a number of these entities. These studies illustrate how genetics provides a rationale and objective basis for a new, refined nosology. Furthermore, whereas most of these studies point towards the pivotal role played by a number of structural proteins--all directly or indirectly associated with dystrophin--a calpain protease was shown to be involved in the Reunion-type limb girdle muscular dystrophy. This discovery raises the issue of whether these mechanisms are all part of one and the same pathway or of distinct pathophysiological pathways (structuropathy versus enzymopathy) leading to similar phenotypes. Finally, all of these diseases are considered as classical monogenic traits. Some findings suggest, however, that epistatic interactions have been overlooked and that the inheritance models could be slightly more complex. These results are discussed in light of the coming challenges of the identification of genes underlying complex multifactorial traits. PMID- 10723862 TI - Disease taxonomy--polygenic. AB - The practice of medicine depends on the recognition and classification of disease. Correct diagnosis is the cornerstone of correct treatment. The past century has seen the classification of disease move from a reliance on symptoms and signs to the use of more and more sophisticated measurements of human structure and function. However, although most diseases have now have names and schemes of classification, these names still may hide a fundamental lack of understanding of the causes of the disease. The extraordinary progress in molecular genetics in the last 20 years now means that a complete understanding of the constitutional predisposition to disease is possible. All disease results from the interaction between adverse environmental events and constitutional (genetic) resistance or susceptibility. Genetic resistance is modified by ageing. The study of genetics is the process of linking polymorphism in the genetic material to polymorphism or variation in the function or appearance of an organism. The extent to which this becomes clinically useful will be determined by the strength of the genetic effects influencing the disease. Oligogenic disorders, in which just a few genes are impacting on the disease, are more likely to be classifiable by genetic polymorphism than true polygenic disorders, in which a multiplicity of small effects give incremental risks of developing disease. Nevertheless, an improved understanding of the aetiology of disease will in all probability identify previously unrecognised yet distinct subsets of disease. PMID- 10723863 TI - Pharmacogenetics. AB - Inter-individual variability in drug response is a major clinical problem. Adverse drug reactions (ADRs) are common, are responsible for a number of debilitating side effects following drug therapy and are a significant cause of death. It is now clear that much of the observed variability in drug response has a genetic basis, arising as a result of genetically-determined differences in drug absorption, disposition, metabolism or excretion. The best characterised pharmacogenetic polymorphisms are those within the phase I cytochrome P450 family of drug metabolising enzymes. One of these enzymes, CYP2D6 (debrisoquine hydroxylase), metabolizes one-quarter of all prescribed drugs and is inactive in 6% of the Caucasian population. Individuals at risk of developing ADRs as a result of genetically-determined variation in genes such as CYP2D6 can now be identified using DNA-based tests. A detailed knowledge of the genetic basis of individual drug response is potentially of major clinical and economic importance and could provide the basis for a rational approach to drug prescription. This would have significant benefits for human health. PMID- 10723864 TI - Pathogen virulence genes--implications for vaccines and drug therapy. AB - The emergence and spread of bacteria resistant to antimicrobial drugs is a major public health problem with a growing number of infections becoming virtually untreatable. There is a need to develop interventions both to prevent and to treat diseases caused by multi-resistant microbes. We review some recently developed methods (including whole genome nucleotide sequencing projects) to study bacterial pathogenesis, and discuss how knowledge gained from understanding the molecular mechanisms of disease can be applied to combat the threat of infectious diseases. PMID- 10723865 TI - Genetics and genomics of infectious disease susceptibility. AB - Human genetic variation is a major determinant of susceptibility to many common infectious diseases. Malaria was the first disease to be studied extensively and many susceptibility and resistance loci have been identified. However, genes for other diseases such as HIV/AIDS and mycobacterial infections are now being identified using a variety of approaches. A large number of genes appear to influence susceptibility to infectious pathogens and defining these can provide insights into pathogenic and protective mechanisms and identify new molecular targets for prophylactic and therapeutic interventions. Immunogenetic associations with infectious diseases have considerable potential to guide immunomodulatory interventions and vaccine design. PMID- 10723866 TI - Communicating genetic risk information. AB - It is envisaged that genetic information will be used, together with other types of information, to assess individuals' risks of developing a variety of common conditions. Such risk assessments will involve providing probabilistic information partly based upon results of genetic tests in order to facilitate behaviour change without causing excessive anxiety. The behaviours targeted for change are likely to include adherence to prescribed medication, alteration to diet, increasing levels of exercise and quitting smoking. This paper reviews research already conducted on perceptions of risk and genes, methods of facilitating behaviour change and reducing anxiety following various types of risk assessment. Although risk assessment and interventions to reduce risks have been conducted for over 20 years, very little rigorous research exists. For the investments in the new genetics to be realised, research is now needed both in how individuals respond to risk information that involves the use of genetic information and in how to facilitate and maintain behaviour change to reduce such risks. PMID- 10723867 TI - Genomics: the implications for ethics and education. AB - Over the past 10 years, the identification of disease genes has been expanding rapidly. Those identified in the earlier part of the decade were largely achieved through positional cloning and the majority are for relatively rare disorders which involve single genes. As the Human Genome Mapping Project has progressed, the rate of gene discovery has increased substantially through the development of new DNA sequencing techniques and in silico approaches. The human genome will have been largely sequenced by ther Spring 2000. We can expect the identification of large numbers of susceptibility genes for common multifactorial polygenic diseases as well as genes which are associated with human behavioural traits. Some of these advances hold out the prospects of real progress in the diagnosis and treatment of a wide range of disorders. However, for many individuals, increased knowledge about their genes will present ethical dilemmas which are difficult to resolve. There are also wider ethical issues which concern the use of genetic information by insurers and employers and yet others which concern ownership and access. In this chapter, the main ethical issues raised by the impact of genomics on healthcare are discussed. The role of education in enabling individuals and health professionals to meet these challenges is also considered. PMID- 10723868 TI - Xenotransplantation. AB - The success of organ transplantation has led to an ever-increasing shortfall between the demand for organs and the supply. This has led to extensive investigation of the possible use of animals, especially the pig, as organ donors. However, a number of major barriers to successful xenotransplantation exist. These include immunological, physiological, anatomical, infectious and ethical problems. Of the immunological problems, the most immediate is hyperacute rejection of the organ caused by natural cytotoxic antibodies in man directed at the galactose alpha-1,3-galactose antigen present in all mammals except man, old world monkeys and the great apes. It is in this area in which genetic engineering has been applied most assiduously and essentially this problem is solved, at least in theory. However, many other problems remain to be resolved before xenotransplantation is likely to become a clinical reality. PMID- 10723869 TI - The impact of genetics on medical education and training. AB - This paper explores, mainly from the UK perspective, some of the issues relating to the current, and potential, impact of advances in genetics and molecular biology on the education and research training of healthcare professionals. We start by describing some of the expectations for progress in the use of genomic technologies and genetic data in healthcare delivery and the need for policy development to ensure timely translation of advances in science and technology into improved patient care. We review briefly the likely evolution of clinical genetics service provision to build the requisite scientific basis in primary care and explore how user needs could be addressed. Strategic issues for the future medical curriculum are introduced and linked with the concerns about the current status of clinical academic research. The issues for research training, career progression, nurturing of research 'at the bedside', definition of the research agenda and weaknesses in both academic infrastructure and support costs are reviewed in the context of the urgent imperative for medicine to harness the accelerating pace of progress in genomics. PMID- 10723870 TI - Genetics in drug discovery and development: challenge and promise of individualizing treatment in common complex diseases. PMID- 10723871 TI - Colonoscopy aided by magnetic 3D imaging: is the technique sufficiently sensitive to detect differences between men and women? AB - Colonoscopy tends to be more difficult to perform in women. Women also experience more pain during flexible sigmoidoscopy, and the mean insertion distance of the instrument is less than in men. The 'Bladen system', first described in 1993, is a non-radiological method of continuously visualising the path of the endoscope using magnetic drive coils under the patient and a chain of sensors up the biopsy channel of the instrument. In 1998, results were published that used a novel computer graphics system (the 'RMR system'), in which a much more realistic endoscope could be produced using the stored positional data from the Bladen system. The RMR computer graphics system has been further refined to enable measurement of the anatomical lengths of different parts of the large intestine to an accuracy of greater than 5 mm. The system is used to analyse the results obtained in 232 patients undergoing a total colonoscopy. In women, the colonoscope tends to form loops in the sigmoid colon more readily than in men (p < 0.05). When the first 50 cm of the endoscope are inserted for the first time, the tip passes either up to or beyond the splenic flexure in 40/116, or 34.5%, of males, compared with 24/117, or 20.5%, of females (p = 0.0137). It is demonstrated that women have longer transverse colons than men, and the differences are especially apparent when a stiffening tube is used to splint the left side of the colon (p < 0.0001). The possible relevance of these observations to biomedical engineers and those manufacturing and assessing prototype endoscopes is discussed. PMID- 10723872 TI - Adaptive fuzzy control of electrically stimulated muscles for arm movements. AB - A modified adaptive Takagi-Sugeno (TS) fuzzy logic controller (FLC) is proposed that allows a simulated elbow-like biomechanical system to accurately track sigmoidal and sinusoidal trajectories in the sagittal plane. The work is a first effort towards the implementation of a system to restore elbow movements in quadriplegics using functional neuromuscular stimulation. The single-joint musculo-skeletal system is composed of a co-contractable pair of electrically stimulated muscles; the muscle model accounts for the increase in fatigue during the tracking exercise. In the proposed controller structure, a reinforcement learning scheme is used to accomplish the parameter tuning, and the parameter projection algorithm guarantees the system stability during the adaptation process. The controller performance is evaluated using computer simulation experiments and compared with the performance achievable when a standard proportional-integrative-derivative (PID) controller is employed for the same application. The modified adaptive TSFLC outperforms the PID controller in all tested situations, with a clear-cut advantage in the case of high-frequency sinusoidal trajectories (angular frequencies spanning the interval 8-12 rad s-1). The standard controller suffers from a dramatic increase in root mean square (RMS) tracking error above the value at 8 rad s-1, e.g. ERMS > or = 0.013, whereas the correlation coefficient between the actual and desired trajectory falls almost to zero, starting from the value rho approximately equal to 0.97 at 8 rad s-1. On the other hand, the adaptive TSFLC yields ERMS < or = 0.015, with rho > or = 0.78, over the whole range of tested angular frequencies. PMID- 10723873 TI - Temporal feature estimation during walking using miniature accelerometers: an analysis of gait improvement after hip arthroplasty. AB - A new method for the detection of gait cycle phases using only two miniature accelerometers together with a light, portable digital recorder is proposed. Each subject is asked to walk on a walkway at his/her own preferred speed. Gait analysis was performed using an original method of computing the values of temporal parameters from accelerometer signals. First, to validate the accelerometric method, measurements are taken on a group of healthy subjects. No significant differences are observed between the results thus obtained and those from pressure sensors attached under the foot. Then, measurements using only accelerometers are performed on a group of 12 patients with unilateral hip osteo arthritis. The gait analysis is carried out just before hip arthroplasty and again, three, six and nine months afterwards. A mean decrease of 88% of asymmetry of stance time and especially a mean decrease of 250% of asymmetry of double support time are observed, nine months after the operation. These results confirm the validity of the proposed method for healthy subjects and its efficiency for functional evaluation of gait improvement after arthroplasty. PMID- 10723874 TI - Measuring arterial strain induced by endovascular stents. AB - Endovascular stents are expandable, fenestrated tubes that are threaded in their collapsed state through an artery to a site of occlusion, plastically enlarged and left as permanent implants to scaffold the artery open. The stent induces large-scale vascular strains that are difficult to measure in vivo and yet can be critical determinants of stent-vessel biology. A method is developed to measure the strain tensor developed on the surface of an artery as a stent is expanded in vivo. Arterial sections are marked with reference points and imaged as the stent is expanded. An axially symmetric parametric model of the artery is determined for each expansion time-point, and these reference points are back-projected onto this surface. The back-projected reference points are grouped and analysed to determine the circumferential, axial and torsional strain tensor components in each arterial subsection. The method is characterised in vitro using bovine artery segments and a latex phantom, and is then tested on rabbits to demonstrate its feasibility in vivo. In vitro experiments on stented bovine arteries show typical post-stenting strains of 0.60, -0.26, and 0.08 mm mm-1 in the circumferential, axial and torsional directions, respectively, sampled every 1 mm along the length of the stented region. Phantom experiments characterise the RMS error of system measurements as 0.1 mm mm-1. The system is shown capable of measuring strains of straight, accessible vessels in the presence of respiratory/cardiac motion and visual glare in vivo. PMID- 10723875 TI - Selection of measurement frequencies for optimal extraction of tissue impedance model parameters. AB - The results are presented of a study performed to determine the measurement frequencies that provide optimal extraction of tissue impedance model parameters from in vivo measured electrical impedance spectra. Measurement frequency sets that are logarithmic and quasi-linear, and frequency sets that produce angularly equidistant points on the Nyquist loci are used to test the parametric fitting algorithm that calculates R0, R infinity, alpha and tau tissue parameters from complex impedance spectra. Simulated data, calculated in the presence of < or = 5% measurement noise, and in vivo experiments indicate that the quality of the fitted parameters depends upon the selection of measurement frequencies. The results show that, if measurements are performed with a system that has a realistic measurement bandwidth, then, for the best estimation of: R0, the measurement frequencies should include the decade from 100 Hz-1 kHz; R infinity, the algorithm should not include frequencies under 1 kHz; alpha and tau, the measurement frequencies should be equidistantly spaced on the Nyquist locus. PMID- 10723876 TI - Magnetic and electrical stimulation of undulating nerve fibres: a simulation study. AB - Mathematical models of myelinated nerve fibres are highly stylized abstractions of real nerve fibres. For example, nerve fibres are usually assumed to be perfectly straight. Such idealizations can cause discrepancies between theoretical predictions and experimental results. One well-known discrepancy is that the currently used models predict (contradictory to experimental findings) that an activation of nerve fibres is not possible with a pure transverse electric field. This situation occurs when a magnetic coil is placed symmetrically above a straight nerve fibre for magnetic nerve stimulation, or when an anode and a cathode are placed equidistantly on a line perpendicular to the fibre in the case of electrical stimulation. It is shown that this discrepancy does not occur if the physiological undulation of peripheral nerve fibres is included in the models. Even for small undulation amplitudes (e.g. 0.02 mm), it is possible to activate the fibre in these positions. For physiological undulations, as found in the literature, and favourable (off-centre) positions, the typical reduction of the thresholds is in a range between one and five, compared with perfectly straight fibres. PMID- 10723877 TI - Three-dimensional electromagnetic model of the human eye: advances towards the optimisation of electroretinographic signal detection. AB - Classical electromagnetic theory is used to examine the topographical variation in electrical potentials over the corneal surface resulting from specific retinal stimuli. Results from a three-dimensional mathematical model show that over 97% of calculated electromagnetic field potentials lie within 3% of previous analytical model data for an axially symmetric case. Maps of corneal potentials are produced that are shown to be characteristic of specific retinal stimuli and location. The maximum variation in corneal potential for a full field global stimulus is found to be approximately 1%. This is considered encouraging, as current electrophysiology techniques measure ocular potentials from a single corneal or scleral site, the position of which is often difficult to localise and reproduce. The model is used to simulate both central and peripheral stimuli and scotoma conditions. A 20 degrees central scotoma simulation shows an overall reduction in central corneal potential of only 3%, whereas peripheral stimuli are found to cause up to 10% variations in this potential. There is therefore a possibility that a single recording site for multifocal retinal stimulation is not ideal. These data may be used to suggest more appropriate electrode recording positions for maximum signal recovery, not least in optimising signal detection for multi-focal electroretinography stimulation. PMID- 10723878 TI - Depth and intensity of equivalent current dipoles estimated through an inverse analysis of surface electromyograms using the image method. AB - The depth and intensity of equivalent current dipoles that can create the surface potentials of active motor units in human skeletal muscles are estimated through an inverse analysis of surface electromyographic (EMG) potentials in an attempt to measure detailed muscular activity non-invasively. The inverse analysis is conducted by repetition of forward analyses. In the study, the image method is used for forward analysis, because it is the simplest potential calculation method for electric currents in a semi-infinite volume conductor. Using this method, surface EMG potentials are calculated for current sources assumed to be located in a muscle. An inverse analysis is then carried out by searching for the depth and intensity of such current sources that would minimise the sum of squares difference between measured and calculated surface EMG potentials. The inverse analysis is applied to surface EMG potentials measured from the biceps brachii of three healthy subjects. As a result, the individual current sources are estimated to be 2.7 +/- 1.6 mm deep and 0.5 +/- 0.9 nAm in intensity, whereas the total current intensity for individual motor units is 2.4 +/- 2.9 nAm. PMID- 10723879 TI - Experimental analysis of the human perception threshold of a DC electric field. AB - To study the biological effects of extremely low frequency (ELF) electric fields, a fundamental study is conducted of the human perception threshold of an electric field. The perception threshold is measured with human subjects, and the results are analysed. It is clear that field perception is based on the movement of hair and not on other sensations. Variance in the perception threshold and its causes are investigated. The perception threshold decreases by almost 30% as the relative humidity increases from 50 to 90%. The perception threshold is also dependent on the physical condition (length and density) of the hair and the psychological condition (degree of awareness) of the subject. The dependence on these is much smaller than that on relative humidity. The cause of the gender difference in the threshold is ascribed to the difference in the physical condition of the hair. Through this study, some factors to be taken into account for the safety standard are made clear. PMID- 10723880 TI - Hybrid finite elements and spectral method for computation of the electric potential generated by a nerve cuff electrode. AB - An original numerical method is developed to compute the 3D electric potential generated by a dot-contact cuff electrode implanted around an axisymmetrical, inhomogeneous, anisotropic nerve. The technique is based on a 2D finite-element approach coupled with a semi-analytical Fourier spectral decomposition to approximate the solution behaviour in the azymuthal direction. The method only requires a 2D FEM mesh and allows an accurate electrode description, with any number of contacts at different angular positions. Results show that the convergence of the Fourier series is very fast: typically, the relative error due to series truncation (estimated by the norm of the difference between the solution computed with M modes and the one computed with M-1 modes, normalised by the norm of the solution computed with M modes) reaches the order of 10(-3) with six spectral modes (M = 6). As a consequence, the whole algorithm has the complexity of a 2D approach. PMID- 10723881 TI - A comparison between in vitro studies of protein lesions generated by brain electrodes and finite element model simulations. AB - The aim of this study was to develop a finite element model for simulation of the thermal characteristics of brain electrodes and to compare its performances with an in vitro experimental albumin model. Ten lesions were created in albumin using a monopolar electrode connected to a Leksell Neuro Generator and a computer assisted video system was used to determine the size of the generated lesions. A finite element model was set up of the in vitro experiments using the same thermal properties. With a very simple heat source applied to the finite element model in the proximity of the upper part of the tip, a good agreement (no deviations in width and distance from tip but a deviation in length of -1.6 mm) with the in vitro experiments (width 4.6 +/- 0.1 mm and length 7.4 +/- 0.1 mm) was achieved when comparing the outline of the lesion. In addition, a gelatinous albumin-model was set up and compared to computer simulations resulting in deviations in width of -0.4 mm, length of -2.2 mm and distance from the tip of 0.1 mm. Hence, the utilisation of finite element model simulations may be a useful complement to in-vitro experiments. PMID- 10723882 TI - Interaction between Gregg's phenomenon and coronary flow control: a model study. AB - Coronary perfusion pressure, Pp, affects coronary arterial resistance, Ra, (autoregulation) as well as myocardial oxygen consumption, MVO2 (Gregg's phenomenon). The interaction between the effects of Pp and MVO2 on coronary flow control was investigated using a coronary flow control model. Model analysis predicts that response of the pressure-flow ratio, p/q(t), following a change in Pp depends on the sensitivity of Ra to a change in tissue oxygen concentration (tone sensitivity) and on the sensitivity of MVO2 to a change in capillary pressure (Gregg's sensitivity). At high tone sensitivity Gregg's effect is small, whereas at high Gregg's sensitivity autoregulation is attenuated. In experiments glibenclamide decelerated the p/q(t) in response to a pressure step by a factor of four. However, the proposed model demonstrates that this is compatible with a reduction in rate of change of Ra by a factor of ten. This is due to the interaction of negative and positive feedback gains in the model. Model analysis demonstrates that autoregulation and Gregg's phenomenon compete with each other in controlling coronary flow. PMID- 10723883 TI - Wavelet-based enhancement of signal-averaged electrocardiograms for late potential detection. AB - An optimal wavelet filter to improve the signal-to-noise ratio (SNR) of the signal-averaged electrocardiogram is described. As the averaging technique leads to the best unbiased estimator, the challenge is to attenuate the noise while preserving the low amplitude signals that are usually embedded in it. An optimal, in the mean-square sense, wavelet-based filter has been derived from the model of the signal. However, such a filter needs exact knowledge of the noise statistic and the noise-free signal. Hence, to implement such a filter, a method based on successive sub-averaging and wavelet filtering is proposed. Its performance was evaluated using simulated and real ECGs. An improvement in SNR of between 6 and 10 dB can be achieved compared to a classical averaging technique which uses an ensemble of 64 simulated ECG beats. Tests on real ECGs demonstrate the utility of the method as it has been shown that by using fewer beats in the filtered ensemble average, one can achieve the same noise reduction. Clinical use of this technique would reduce the ensemble needed for averaging while obtaining the same diagnostic result. PMID- 10723884 TI - Evaluation of frequency and time-frequency spectral analysis of heart rate variability as a diagnostic marker of the sleep apnoea syndrome. AB - The sleep apnoea/hypopnoea syndrome (SAHS) elicits a unique heart rate rhythm that may provide the basis for an effective screening tool. The study uses the receiver operator characteristic (ROC) to assess the diagnostic potential of spectral analysis of heart rate variability (HRV) using two methods, the discrete Fourier transform (DFT) and the discrete harmonic wavelet transform (DHWT). These two methods are compared over different sleep stages and spectral frequency bands. The HRV results are subsequently compared with those of the current screening method of oximetry. For both the DFT and the DHWT, the most diagnostically accurate frequency range for HRV spectral power calculations is found to be 0.019-0.036 Hz (denoted by AB2). Using AB2, 15 min sections of non REM sleep data in 40 subjects produce ROC areas, for the DFT, DHWT and oximetry, of 0.94, 0.97 and 0.67, respectively. In REM sleep, ROC areas are 0.78, 0.79 and 0.71, respectively. In non-REM sleep, spectral analysis of HRV appears to be a significantly better indicator of the SAHS than the current screening method of oximetry, and, in REM sleep, it is comparable with oximetry. The advantage of the DHWT over the DFT is that it produces a greater time resolution and is computationally more efficient. The DHWT does not require the precondition of stationarity or interpolation of raw HRV data. PMID- 10723885 TI - Computer-controlled flow resistance. AB - A computer-controlled flow resistance (CCR), to be used in a computer-controlled lung model, is presented. Flow is forced through a slit between a cylinder and a sleeve around the cylinder. The resulting flow resistance depends on the width, circumferences and the variable length of the slit. The variation in the length is computer-controlled by the position of the sleeve with respect to the cylinder. The total flow resistance also depends on inlet and outlet resistance at both sides of the slit and on flow. The dependence on flow is primarily due to the shape of the inlet of the slit. The resistance of the slit itself is almost independent of flow. The resistance is calculated during a calibration phase at different positions of the sleeve, for flow values from 0.05 to 1.0 litre.s-1 (inflow) and from -0.05 to -1.0 litre.s-1 (outflow). To simulate a required resistance pattern, as, for instance, will occur during breathing, at each moment the set position of the sleeve is calculated by means of an interpolation from the relationship between flow resistance and position of the sleeve. The internal diameter of the sleeve is fixed. To tune the resistance range for a specific simulation, the cylinder is changed for one with different diameter, changing the width of the slit. PMID- 10723886 TI - Removal of cardiac beat artifact in oesophageal pressure measurement by frequency analysis. AB - Oesophageal pressure (Pes) measurements are important in medical research and useful in clinical diagnosis. Measurements, however, are contaminated heavily by cardiac artifacts. The spectrum and waveform of the Pes signal is obtained from the oesophageal balloon. Adaptive finite impulse response (AFIR) filter and modified adaptive noise cancellation (MANC) methods are adopted to filter out cardiac beat interference. These results are compared. In the frequency domain, frequency variations and spectral overlap between the Pes components and cardiac beat signal components impact on the performance of the filter. From our experimental results on power strength, the fourth or higher harmonics did not have any significant effect on the filter performance. However, the second harmonics of these signals had a significant effect on the filtering result. Thus, in the design of AFIR filters, attention is needed to minimise these effects. In frequency analysis, these harmonics or overlapping frequencies do not affect MANC. MANC was the better method for eliminating cardiac beat artifact in Pes measurement. The dynamic compliance (Cdyn) was also used to evaluate the performance of MANC and AFIR. The standard deviation of Cdyn was less than 0.15 using MANC, compared with standard deviations as high as 0.57 for AFIR. We conclude that MANC performs better than AFIR. PMID- 10723887 TI - Mathematical modelling of non-invasive oscillometric finger mean blood pressure measurement by maximum oscillation criterion. AB - A mathematical study is performed to assess how the arterial pressure-volume (P V) relationship, blood pressure pulse amplitude and shape affect the results of non-invasive oscillometric finger mean blood pressure estimation by the maximum oscillation criterion (MOC). The exponential models for a relaxed finger artery and for a partly contracted artery are studied. A new modification of the error equation is suggested. This equation and the results of simulation demonstrate that the value of pressure estimated by the MOC does not exactly agree with the value of the true mean blood pressure (the latter being defined as pressure corresponding to maximum arterial compliance). The error depends on the arterial pressure pulse amplitude, as well as on the difference between the arterial pressure pulse shape index and the arterial P-V curve shape index. In the case of contracted finger arteries, the MOC can give an overestimation of up to 19 mmHg, the pressure pulse shape index being 0.21 and the pulse amplitude 60 mmHg. In the case of relaxed arteries, the error is less evident. PMID- 10723888 TI - Automatic on-line analyser of microbial growth using simultaneous measurements of impedance and turbidity. AB - An apparatus for the measurement of bacterial growth is described. The instrument applies alternate adequate sequential currents of two different frequencies through a pair of electrodes immersed in a cultured medium. It monitors, detects and quantifies the growth of micro-organisms based on the measurement of the impedance across the two electrodes and, simultaneously, it measures the variation in the medium turbidity. The medium conductivity and the interface electrode impedance changes can be extracted from the measured impedance. The variations in turbidity can be calibrated in absorbance or optical density units. Moreover, all these parameters are also proportional to bacterial proliferation. The computer-controlled apparatus processes and displays the parameters on a monitor showing bulk resistance, electrode impedance and turbidity changes as time course events. The equipment can detect aerobic or anaerobic micro-organisms and permits the operator simultaneously to assess impedance and turbidity, or it can produce each parameter as a separate event. Time growth curves of different micro-organisms are presented in the results. PMID- 10723889 TI - Sensitivity of foetal magnetocardiograms versus gestation week. AB - Foetal magnetocardiograms (FMCGs) were measured using a nine-channel SQUID system equipped with first-order gradiometers (60 mm baseline, 20 mm diameter). The system was installed in a magnetically shielded room in a hospital. The white noise level was less than 10 fT/square root of Hz, and FMCGs above 1 pT were detected. These results and the depth to the foetal heart were used to estimate the current dipole. The relationship between the current dipole (Q) and the gestation week (G) was calculated and the average performance was determined as Q = 18G - 295. By using the estimated foetal current dipole, the measurement limit (average value) between depth and gestation weeks was determined. When the depth from the pickup coil to the foetal heart is 50, 60, 70 and 80 mm, the first detectable gestation weeks of FMCGs above 1 pT measured by a first-order gradiometer with 60 mm baseline were determined at 21, 23, 26, and 30 weeks respectively, and the detectable gestation weeks in the case of a 30 mm baseline were determined at 22, 26, 30 and 36 weeks respectively. PMID- 10723890 TI - Spatial and temporal variations in the magnetic fields produced by human gastrointestinal activity. AB - Magnetoenterography (MENG) is a new, non-invasive technique that measures gastrointestinal magnetic signals near the body surface. This study was undertaken to evaluate the temporal and spatial characteristics of the magnetic signals generated by gastric and duodenal slow wave activity. The gastrointestinal magnetic fields of eight normal subjects were measured for 60 minutes in both the fasting and fed state using 36 magnetic sensors simultaneously. The results were displayed as a succession of maps over time showing the temporal evolution of the spatial distribution of the signal over the upper abdomen. In all subjects, slow wave activity of the stomach centred at 3.0 +/- 0.5 cycles min-1 in both the fasting and fed state was observed. The duodenal signal at 11.0 +/- 1.0 cycles min-1 was observed in four subjects. The spatial distribution of these two signals is distinctly different. The observed spatial and temporal variations are described in terms of a model used previously to explain the potentials observed in electrogastrography (EGG). PMID- 10723891 TI - Effects of tissue resistivities on electroencephalogram sensitivity distribution. AB - The effects of tissue resistivities on EEG amplitudes were studied using an anatomically accurate computer model based on the finite difference method (FDM) and lead field analysis covering the whole brain area with 180,000 nodes. Five tissue types and three lead fields were considered for analysis. The changes in sensitivity distribution are directly comparable to changes in the potential distribution on the scalp. The results indicate that a 10% decrease in any tissue resistivity caused 3.0-4.1% differences in the sensitivity distributions of the selected EEG leads. The applied 10% decrease in the resistivity values covers only a fraction of the range of variation of 50% to 100% reported in the literature. The use of a 55% decreased skull resistivity value or a commonly applied three-compartment model increased the differences to 28% and 33%, respectively. In conclusion, both a realistic anatomy and accurate resistivity data are important in EEG head models. PMID- 10723892 TI - Classification of premature ventricular complexes using filter bank features, induction of decision trees and a fuzzy rule-based system. AB - The classification of heart beats is important for automated arrhythmia monitoring devices. The study describes two different classifiers for the identification of premature ventricular complexes (PVCs) in surface ECGs. A decision-tree algorithm based on inductive learning from a training set and a fuzzy rule-based classifier are explained in detail. Traditional features for the classification task are extracted by analysing the heart rate and morphology of the heart beats from a single lead. In addition, a novel set of features based on the use of a filter bank is presented. Filter banks allow for time-frequency dependent signal processing with low computational effort. The performance of the classifiers is evaluated on the MIT-BIH database following the AAMI recommendations. The decision-tree algorithm has a gross sensitivity of 85.3% and a positive predictivity of 85.2%, whereas the gross sensitivity of the fuzzy rule based system is 81.3%, and the positive predictivity is 80.6%. PMID- 10723893 TI - Combined wavelet transformation and radial basis neural networks for classifying life-threatening cardiac arrhythmias. AB - Automatic detection and classification of arrhythmias based on ECG signals are important to cardiac-disease diagnostics. The ability of the ECG classifier to identify arrhythmias accurately is based on the development of robust techniques for both feature extraction and classification. A classifier is developed based on using wavelet transforms for extracting features and then using a radial basis function neural network (RBFNN) to classify the arrhythmia. Six energy descriptors are derived from the wavelet coefficients over a single-beat interval from the ECG signal. Nine different continuous and discrete wavelet transforms are considered for obtaining the feature vector. An RBFNN adapted to detect and classify life-threatening arrhythmias is then used to classify the feature vector. Classification results are based on 159 arrhythmia files obtained from three different sources. Classification results indicate the potential for wavelet based energy descriptors to distinguish the main features of the signal and thereby enhance the classification scheme. The RBFNN classifier appears to be well suited to classifying the arrhythmia, owing to the feature vectors' linear inseparability and tendency to cluster. Utilising the Daubechies wavelet transform, an overall correct classification of 97.5% is obtained, with 100% correct classification for both ventricular fibrillation and ventricular tachycardia. PMID- 10723894 TI - Spatial, temporal and wavefront direction characteristics of 12-lead T-wave morphology. AB - Three new approaches for the analysis of ventricular repolarisation in 12-lead electrocardiograms (ECGs) are presented: the spatial and temporal variations in T wave morphology and the wavefront direction difference between the ventricular depolarisation and repolarisation waves. The spatial variation characterises the morphology differences between standard leads. The temporal variation measures the change in interlead relationships. A minimum dimensional space, constructed by ECG singular value decomposition, is used. All descriptors are measured using the ECG vector in the constructed space and the singular vectors that define this space. None of the descriptors requires time domain measurements (e.g. the precise detection of the T-wave offset), and so the inaccuracies associated with conventional QT interval related parameters are avoided. The new descriptors are compared with the conventional measurements provided by a commercial system for an automatic evaluation of QT interval and QT dispersion in digitally recorded 12 lead ECGs. The basic comparison uses a set of 1100 normal ECGs. The short-term intrasubject reproducibility of the new descriptors is compared with that of the conventional measurements in a set of 760 ECGs recorded in 76 normal subjects and a set of 630 ECGs recorded in 63 patients with hypertrophic cardiomyopathy (ten serial recordings in each subject of both these sets). The discriminative power of the new and conventional parameters to distinguish normal and abnormal repolarisation patterns is compared using the same set. The results show that the new parameters do not correlate with the conventional QT interval-related descriptors (i.e. they assess different ECG qualities), are generally more reproducible than the conventional parameters, and lead to a more significant separation between normal and abnormal ECGs, both univariately and in multivariate regression models. PMID- 10723895 TI - Automatic measurement of long-term heart rate variability by implanted single chamber devices. AB - Heart rate variability (HRV) measurement is an established technology for the assessment of cardiac autonomic status. Recently 24 h HRV has been shown to correlate with disease severity in heart failure. This potentially makes continuous 24 h HRV measurement suitable for monitoring of heart-failure patients. Day-to-day 24 h measurement of HRV is, in principle, feasible when implemented using implanted devices (pacemakers and defibrillators) used in patients who are predominantly in the sinus rhythm. However, a number of such devices used in heart-failure patients are single-chamber devices, in which the distinction between sinus rhythm beats and ectopic beats is problematic. The study investigates whether a reasonably accurate 24 h HRV measurement can be achieved by automatic algorithms, suitable for implementation using implanted devices, without the need for identification of ectopic beats. A set of 5321 nominal 24 h Holter recordings of cardiac patients are used. Each of the recordings contains at least one ectopic beat; approximately 30% of the recordings have more than 1% of ectopic beats. Conventional 24 h measures of HRV, that is the SDNN, HRV index, and SDANN indices, are obtained from each recording after elimination of the ectopic beats and are approximated by HRV measures computed by the same formulas without exclusion of the ectopic beats. The SDANN values are also approximated by the standard deviation of 5 min medians of all RR intervals (SDMRR measure). The errors introduced by including the ectopic beats in the HRV computation were evaluated using the Bland-Altman statistics and by Cohen's kappa statistics investigating the precision of identifying patients with depressed and preserved 24 h HRV. The SDNN measure is very sensitive to the quality of the RR interval sequence and cannot be reasonably used without distinction between sinus rhythm and ectopic beats. The HRV index measure is marginally more acceptable when used without ectopic elimination. The SDANN is rather insensitive, and its replacement by SDMRR values leads to relative errors in the region of 2-5% that are almost independent of the number of ectopic beats included. Even in recordings with a substantial proportion of ectopic beats, a practically acceptable (kappa > 0.9) identification of depressed and preserved SDANN values is possible without ectopic elimination. Thus, continuous monitoring of 24 h HRV is technically feasible within implanted devices, provided the SDANN measure is monitored and either computed from the sequence of all RR intervals or, potentially preferably, replaced by the SDMRR measure. PMID- 10723896 TI - Effects of multiple stenoses and post-stenotic dilatation on non-Newtonian blood flow in small arteries. AB - Fully-developed one-dimensional Casson flow through a single vessel of varying radius is proposed as a model of low Reynolds number blood flow in small stenosed coronary arteries. A formula for the resistance-to-flow ratio is derived, and results for yield stresses of tau 0 = 0, 0.005 and 0.01 Nm-2, viscosities of mu = 3.45 x 10(-3), 4.00 x 10(-3) and 4.55 x 10(-3) Pa.s and fluxes of 2.73 x 10(-6), x 10(-5) and x 10(-4) m3 s-1 are determined for segment of 0.45 mm radius and 45 mm length, with 15 mm abnormalities at each end where the radius varies by up to +/- 0.225 mm. When tau 0 = 0.005 N m-2, mu = 4 x 10(-3) Pa.s and Q = 1, the numerical values of the resistance-to-flow ratio vary from lambda = 0.525, when the maximum radii of the two abnormal segments are both 0.675 mm, to lambda = 3.06, when the minimum radii are both 0.225 mm. The resistance-to-flow ratio moves closer to unity as yield stress increases or as blood viscosity or flux decreases, and the magnitude of these alterations is greatest for yield stress and least for flux. PMID- 10723897 TI - Real-time automatic extraction of lumen region and boundary from endoscopic images. AB - A new approach to the automatic extraction of the lumen region and its boundary for gastrointestinal (GI) endoscopic images is presented. First, a quasi region of interest, the darker regions of the image, is segmented using a region splitting scheme termed progressive thresholding. The centre of mass of this segmented region acts as a seed for further processing. Then the lumen region is obtained using a region growing technique called the integrated neighbourhood search (INS). A new quad structure based technique is introduced to enhance the INS speed significantly. A back projection algorithm is suggested to optimise the search for pixels belonging to the lumen region and boundary. A boundary-thinning algorithm is also proposed to remove the redundant pixels from the lumen boundary and to generate a connected single pixel width boundary. The proposed approach does not need a priori knowledge about the image characteristics. The experimental results indicate that the proposed technique enhances the speed of conventional INS by 45.5% to 28.6% based on the lumen size varying from 22,709 pixels to 4947 pixels. The main advantage of the proposed technique is its high speed response that facilitates real-time analysis of endoscopic images. PMID- 10723899 TI - Design of a microwave array hyperthermia applicator with a semicircular reflector. AB - The design of a hyperthermia applicator for heating biological tissues is presented in which the applicator consists of an array antenna surrounded by a perfect electrically conducting reflector. The heat hazard to superficial tissues is reduced by the introduction of a dielectric protecting layer over them. A method of moments formulation is applied to approximate the electric field within the biological medium and a closed form expression is presented for the electromagnetic coupling problem, which enables an optimisation procedure to be performed. The applicator enhances both penetration and focusing: deep tumours, close to the bone region, are heated and the percentage of biologically healthy tissue exposed to a specific absorption rate (SAR) hazard level diminishes by 53.8%. PMID- 10723898 TI - Colonoscopy aided by magnetic 3D imaging: assessing the routine use of a stiffening sigmoid overtube to speed up the procedure. AB - There are not enough trained colonoscopists to cope with the present recommended number of examinations required for diagnostic and surveillance purposes. If colorectal cancer screening is to be introduced, endoscopic examination of the large bowel needs to be easier to learn and significantly quicker to carry out. The 'Bladen system', first described in 1993, is a non-radiological method of visualising the path of the endoscope, using magnetic drive coils under the patient and a chain of sensors along the biopsy channel of the instrument. In 1998, results were published using a novel computer graphics system (the RMR system), in which a much more realistic image of the endoscope could be produced using the stored positional data from the Bladen system. The RMR system has been further refined to allow, for the first time ever, accurate measurement of the effect of the passage of a colonoscope along the bowel on the lengths of different segments of the large intestine. The results obtained in 232 patients undergoing colonoscopy are analysed. In 77 of the patients, a stiffening overtube is used to splint the sigmoid colon once the endoscope is at or beyond the splenic flexure. The mean time taken to pass the colonoscope across the transverse colon is significantly shorter (p < 0.001) when an overtube is used, despite it resulting in significant lengthening of the transverse colon. The routine use of a stiffening overtube can be expected to reduce the total procedure time by between 10 and 20%. PMID- 10723900 TI - A catheter tactile sensor for measuring hardness of soft tissue: measurement in a silicone model and in an in vitro human prostate model. AB - Tissue hardness is related to tissue composition, and this is often changed by disease. It is therefore of interest to measure the hardness in an objective and non-invasive way. A tactile sensor based on a vibrating piezoelectric ceramic element in a feedback loop is described. When the sensor touches an object it produces a frequency shift related to the hardness of the object. The aim of this study was to develop an in vitro hardness measurement method using a catheter type version of the sensor. The method was evaluated in an established silicone tissue model and on human prostate tissue in vitro. A linear relationship was found with a high degree of explanation (R2 = 0.98) between a cone penetration hardness standard (DIN ISO 2137) applied to the silicone model and the corresponding frequency shift. The results from measurements on a human prostate tissue sample, fixed with formalin, showed that the relative hardness measured with the tactile sensor correlated (R = -0.96, p < 0.001, N = 60) with the proposed hardness related to the histological composition of the prostate tissue. The results indicated that hardness of prostate tissue, and maybe hardness of human tissue in general, can be expressed according to the cone penetration standard and that the hardness can be measured with this tactile sensory system. These findings hold the promise of further development of a non-invasive tool for hardness measurement in a clinical situation. PMID- 10723901 TI - Stresses in the normal and diabetic human penis following implantation of an inflatable prosthesis. AB - The prevalence of impotence in diabetes mellitus ranges as high as 75%. The implantation of an inflatable penile prosthesis (IPP) is frequently carried out to restore erectile function. However, clinical studies have demonstrated that severe post-implantation penile pain during erection is a common complication in diabetic men. A biomechanical model of the penis/prosthesis complex is developed, based on cross-sectional anatomy, to simulate the internal stress distribution due to interaction of the prosthesis with both normal and diabetic penile tissues. The material properties of the model components are adopted from experimental data. The model is solved by using commercial finite-element software for a characteristic inflation loading of the penile prosthesis. Elevated structural stresses during erection are found in the dorsal aspect of the tunica albuginea (normal 5.1-31.5 kPa, diabetic 5.1-70 kPa post implantation). Following IPP implantation, stresses in the diabetic penis are almost as twice as high as those in the normal one and can cause a painful sensation owing to nerve stimulation or to ischaemia in regions of compressed vascular tissue. PMID- 10723902 TI - Reliability of neural-network functional electrical stimulation gait-control system. AB - Functional electrical stimulation (FES) has been used for restoring walking in spinal-cord injured (SCI) persons. Using artificial intelligence (AI), FES controllers have been developed that allow the automatic phasing of stimulation, to replace the function of hand or heel switches. However, there has been no study to evaluate the reliability of these AI systems. Neural networks were used to construct FES controllers to control the timing of stimulation. Different numbers of sensors in the sensor set and different numbers of data points from each sensor were used. Two incomplete-SCI subjects were recruited, and each was tested on three separate occasions. The results show the neural-network controllers can maintain a high accuracy (around 90% for the two- and three sensor groups and 80% for the one-sensor group) over a period of six months. Two or three sensors were sufficient to provide enough information to construct a reliable FES control system, and the number of data points did not have any effect on the reliability of the system. PMID- 10723903 TI - A system for investigating oesophageal photoplethysmographic signals in anaesthetised patients. AB - The monitoring of arterial blood oxygen saturation in patients with compromised peripheral perfusion is often difficult, because conventional non-invasive techniques such as pulse oximetry (SpO2) can fail. Poor peripheral circulation commonly occurs after major surgery including cardiopulmonary bypass. The difficulties in these clinical situations might be overcome if the sensor were to monitor a better perfused central part of the body such as the oesophagus. A new oesophageal photoplethysmographic (PPG) probe and an isolated processing system have been developed to investigate the pulsatile signals of anaesthetised adult patients undergoing routine surgery. Measurements were made in the middle third of the oesophagus, 25 cm to 30 cm from the upper incisors. The AC PPG signals are sampled by a data acquisition system connected to a laptop computer. The signals recorded correspond to infrared and red AC PPGs from the middle third oesophagus and the finger. Preliminary results from 20 patients show that good quality AC PPG signals can be measured in the human oesophagus. The ratio of the oesophageal to finger AC PPG amplitudes was calculated for the infrared and red wavelengths for each patient. The mean (+/- standard deviation) of this ratio was 2.9 +/- 2.1 (n = 19) for the infrared wavelength and 3.1 +/- 2.4 (n = 16) for the red wavelength. The red and infrared wavelengths used are appropriate for pulse oximetry and this investigation indicates that the mid-oesophagus may be a suitable site for the reliable monitoring of SpO2 in patients with poor peripheral perfusion. PMID- 10723904 TI - Pressure measurements during injection of corticosteroids: in vivo studies. AB - Intralesional injection of corticosteroids is an effective treatment for capillary hemangiomas. Complications include embolisation of corticosteroid particles into the ocular circulation resulting in permanent loss of vision. This research is aimed at developing an injection cannula and monitoring system to prevent such inadvertent embolisation. A cannula has been designed to simultaneously estimate the pressure at its tip and the flow rate during injection. The estimation technique has previously been validated using an in vitro model. In this study, the cannula was tested in vivo with canine liver at injection flow rates of 2.5 to 21 ml min-1. The pressure generated in the tissue during injection was calculated using the technique developed. This was compared with direct in situ pressure measurements made with a coaxial outer cannula. The mean calculated pressure was seen to be linearly related to the mean measured pressure with a slope of 0.97, correlation coefficient of 0.99 and standard error of 2.74 mmHg. Similar trends were observed between the maximum calculated and maximum measured injection pressure: slope = 1.0, r = 0.99 and standard error = 5.54 mmHg. The estimation of the mean and maximum pressure from the cannula and monitoring system was accurate in canine liver. High pressures close to 250 mmHg were generated in tissues during injection. PMID- 10723905 TI - Acoustical recognition of laryngeal pathology: a comparison of two strategies based on sets of features. AB - The efficiency of sets of acoustical features discriminating pathological voices from control voices is reported. Two strategies were compared. The first (called the 'distance strategy') was built upon a statistical distance of voice features to reference values obtained for a set of healthy (reference) voices. The second strategy (called the 'range strategy') is based on the position inside or outside normal ranges established from a reference population; results based on this strategy were presented in a previous paper. Reference values were calculated from a database of 200 healthy voices distributed into 10-year age groups ranging from 20 to 70. Comparisons were made using a second database of 220 voices, including 65 control, 51 functional dysphonia, 50 with nodules on the vocal folds and 54 recurrent nerve palsy. The phonetic material was compared of 17 French vowels: 11 vowels in a sentence, three isolated vowels and three segments (beginning, middle and end) of the sustained vowel /a/. Four acoustical features were considered for each vowel: the voice fundamental (f0) and the first three formant frequencies. Acoustical features were calculated on an ILS (Interactive Laboratory System) analysis system (workstation). The separation of each pathological group from the control group, using sets of acoustical features, was statistically assessed. From the strategy point of view, results indicated that (i) the fundamental frequency f0 was the best measure to separate normal from pathological voices with the distance strategy; (ii) when the formants were taken, the range strategy performed better in separating the voices. For classification of pathologies, the best separation coefficients were obtained with nodules and the worst with recurrent nerve palsy. Overall, it was seen that the separation between control and pathological voices was most efficient when measured using the distance strategy for f0. The range strategy was useful with formant frequencies. PMID- 10723906 TI - Qualitative modelling of the response of cytoskeletal actin filaments in endothelial cells subjected to shear stress. AB - An approach to modelling cytoskeletal function based on qualitative process theory and qualitative algebra is presented. A computer environment for qualitative modelling and simulation of physical systems, specifically oriented to the construction of biomedical knowledge bases, is used. The system that is being set up for the cytoskeleton is intended to serve as a tool for testing assumptions about cytoskeletal function, for the evaluation of experimental data and for planning experimental work. The aspects of cytoskeletal function that have been modelled so far are focal adhesion and the reorganisation of the vascular endothelial cell cytoskeleton in response to fluid-imposed shear stress. Starting from a natural language description, the subsequent levels of analysis leading to the formal structural description and the resulting implementation for these aspects within the development environment are presented. PMID- 10723907 TI - A new perfusion cell chamber system for determination of heat shock effects by means of video-enhanced microscopy. AB - A user friendly microscope perfusion chamber which allows real-time observation of individual cells at high magnification has been designed. An integrated multisensor was used to monitor the cell culture conditions. To prove the potential of the system heat shock experiments were performed. By means of video enhanced contrast microscopy (VECM) the mitochondria morphology of cultured astrocytes was demonstrated to change from a rod-like to an annular shape after heat shock. For further analyses mitochondria were stained on the microscope stage. PMID- 10723908 TI - Analysis of the mechanical properties of in vitro reconstructed epidermis: preliminary results. AB - Human epidermis can be reconstructed in vitro and is currently used in autografts for the treatment of severe, extensive burns and pigmentation disorders. However, there are neither international standards nor a common nomenclature for engineered tissues. The paper discusses the results of a preliminary study on human cultured epidermis to assess its mechanical tensile strength, and to eventually establish mechanical evaluation criteria that will enable test and comparison of the behaviour of different engineered tissue products. To perform uniaxial tension tests a traditional testing machine was adapted, and dedicated sample holding frame and grips designed. PMID- 10723909 TI - [Clinical aspect of gastroesophageal reflux disease]. PMID- 10723910 TI - [Concept of gastroesophageal reflux disease]. PMID- 10723911 TI - [Epidemiology of gastroesophageal reflux disease]. PMID- 10723912 TI - [Mechanism for the development of gastroesophageal reflux disease]. PMID- 10723913 TI - [Symptoms of gastrointestinal reflux disease (including atypical symptoms)]. PMID- 10723914 TI - [Diagnosis of gastroesophageal reflux disease. 1. Endoscopic diagnosis]. PMID- 10723915 TI - [Diagnosis of gastroesophageal reflux disease. 2. Diagnosis by 24-hour esophageal pH monitoring]. PMID- 10723916 TI - [Diagnosis of gastroesophageal reflux disease. 3. Tests for esophageal motility (including esophageal manometry)]. PMID- 10723917 TI - [Diagnosis of gastroesophageal reflux disease. 4. Pathology of gastroesophageal reflux: a morphological viewpoint]. PMID- 10723918 TI - [Treatment of gastroesophageal reflux diseases. 1. General therapeutic approaches]. PMID- 10723919 TI - [Gastroesophageal reflux disease. 2. Drug therapy]. PMID- 10723920 TI - [Treatment of gastroesophageal reflux disease. 3. Surgical treatment]. PMID- 10723921 TI - [Complications of gastrointestinal reflux diseases. 1. Mechanism for the etiology of Barrett esophagus]. PMID- 10723922 TI - [Complications of gastroesophageal reflux disease. 2. Endoscopic diagnosis of Barrett esophagus--can Barrett esophagus be diagnosed by endoscopic observation alone?]. PMID- 10723923 TI - [Complications of gastroesophageal reflux disease. 3. Cancer arising from Barrett esophagus]. PMID- 10723925 TI - [Clinical aspect of gastroesophageal reflux disease: discussion]. PMID- 10723924 TI - [Helicobacter pylori elimination and gastroesophageal reflux disease]. PMID- 10723926 TI - [Donor leukocyte transfusion in early relapsed acute lymphoblastic leukemia after allogenic bone marrow transplantation]. PMID- 10723927 TI - [Dermatomyositis associated with multiple primary cancers of the stomach, colon and lung]. PMID- 10723928 TI - [Secondary amyloidosis in an autopsied case of adult onset Still disease]. PMID- 10723929 TI - [Remission for a patient with ulcerative colitis after agranulocytosis by the side effect of salazosulfapyridine]. PMID- 10723930 TI - [Acromegaly with a remarkable high growth hormone response by thyrotropin releasing hormone and symptoms occurred after total thyroidectomy]. PMID- 10723931 TI - [Progress in nuclear medicine in cardiology]. PMID- 10723932 TI - [Genetic abnormalities and diabetes mellitus--with special reference to MODY]. PMID- 10723933 TI - [Obesity and diseases in the field of internal medicine]. PMID- 10723934 TI - [Liver transplantation among adults in Japan]. PMID- 10723935 TI - Immunohistochemical analysis of connexin26 and 43 expression in the mouse alimentary tract. AB - The spatial pattern of connexins26 (Cx26) and 43 (Cx43) expressions were investigated in the mouse digestive tract by immunocytochemistry. High levels of connexin43 in the epithelium of the oesophagus, non-glandular part of the stomach, and the circular layer of duodenal and ilea muscularis externa were detected. Cx26 was expressed in stratum granulosum of oesophagal folds and in the non-glandular part of the stomach. A low level of immunoreactivity of Cx43 was observed in the circular, and very low in the longitudinal layer of the muscularis externa in the stomach and colon. No immunoreactivity for Cx26 and Cx43 was found in the entire muscularis externa of the oesophagus or in the longitudinal muscle layers of the duodenum and ileum. PMID- 10723936 TI - Depletion of head kidney neutrophils and cells with basophilic granules during peritoneal inflammation in the goldfish, Carassius auratus. AB - The head kidney morphology of the goldfish with the experimental peritoneal inflammation (on day 2 after i.p. injection with sterile 3% Thioglycollate) is compared with that in the control fish (i.e. on day 2 after i.p. injection with a strile physiological saline PBS) with special emphasis on identification of granulocytes on semithin and consecutive ultrathin sections. The most striking feature of head kidneys of goldfish in the course of peritoneal inflammation is a severe depletion of mature neutrophils and cells with basophilic granules (basophils/mast cells). These observations suggest the involvement of the head kidney, the main hematopoietic organ of teleosts, in the inflammatory process. PMID- 10723937 TI - Cytochemical identification of mucocysts in Balantidium coli trophozoites. AB - The mucocyst ultrastructure in B. coli has not been described so far. As demonstrated in this work, cytoenzymatic assays on B. coli with the use of a reaction-detecting membrane-coupled hydrolase, i.e., ATP-ase, permitted identification of the mucocysts in the ciliate studied. The shape, size, and location of mucocysts in B. coli trophozoites were found to correspond to descriptions of these structures in other ciliates. The mucocysts were more numerous in B. coli trophozoites isolated from the symptomatic balantidiosis affected pigs (Group I), and the product of reaction to ATP-ase was more copious than in Group II trophozoites. However, not all the bubble-like structures with similar morphological features reacted positively to the enzyme. The discrepancy was explained by the cytoenzymatic reaction to Beta-GR. The reaction product was visible in the vesicular structures, situated above the plasmolemma, although some of them contained no reaction product. Thus the presence of two types of secretory structure can be inferred: the mucocysts, with ATP-ase in their membranes, and other extrusomes containing active Beta-GR. PMID- 10723938 TI - [The virulence for mice of newly described mycobacterial species]. AB - The virulence for mice of some newly described slowly growing mycobacterial species, M. intermedium DSM 44049 (Group I), M. lentiflavum ATCC 51985 and M. interjectum ATCC 51457 (Group II), M. genavense ATCC 51233, M. celatum ATCC 51131, M. branderi ATCC 51788 and M. conspicuum DSM 6333 (Group III) was compared with the virulence of M. intracellulare N-260 (virulent strain). 10(6) CFU cultured in Middlebrook 7H9 medium were given i.v. to the BALB/c mice. Infected animals were sacrificed after one day and 56 days, and observed for the incidence and/or degree of gross lesions of visceral organs. The bacterial load (CFU) in tissue homogenates of the lungs and spleen was counted using 7H11 agar plates. Macroscopically, marked enlargement of the spleen and numerous small whitish nodules in the lungs of mice infected with M. intracellulare were noted, whereas with other test strains no gross disease in the lungs and slight enlargement of the spleen of mice infected with M. genavense and M. branderi were noted. The average log CFU in the lungs and spleen 56 days after the infection was in the order as follows: M. intracellulare (5.90 and 7.57, respectively), M. genavense (4.35, 7.15), M. celatum (5.01, 6.00), M. branderi (4.09, 6.33), M. conspicuum (3.11, 5.10), M. intermedium (2.26, 4.19), M. lentiflavum (1.70, 3.91), and M. interjectum (1.43, 2.93). Microscopically, granulomatous lesions mainly consisted of macrophages with acid-fast bacilli were found in the alveolar walls and splenic red pulp of mice infected with M. intracellulare. No histopathological changes were produced in the lungs of any of mice infected with other test strains, and some milder features were revealed in the spleen of mice infected with M. genavense, M. branderi and M. celatum. Thus, we concluded that all of the test strains of the newly described nontuberculous slowly growing mycobacteria were less virulent for mice than were M. intracellulare. PMID- 10723939 TI - [A study on the risk factors for tuberculosis epidemics observed from the results of extraordinary health examinations]. AB - In recent years, tuberculosis (tbc) epidemics have been increasing, and have become a social problem in Japan. This study was carried out to clarify the current risk factors on tbc epidemics. The original reports of a total of 254 tbc patients whose contacts were examined for possible tbc infection (extraordinary health examinations) during 1992-97, were investigated and compared with all new pulmonary tbc cases registered during the same period in Aichi prefecture. In addition, from registration cards in each public health center, the risk factors for 19 cases of both tbc epidemics (infecting more than 20 contacts) and microepidemics infecting 10-19 contacts were further examined. The results obtained were as follows; 1. Extraordinary health examinations were carried out in 3.2% of total pulmonary tbc cases, and were carried out at higher rates among contacts with younger patients and with those having severer tbc by radiographical (p < 0.01) and bacteriological findings (p < 0.01). 2. 11 tbc epidemics (3.4%) and 34 microepidemics (14.3%) were identified among 254 examinations over 6 years. They were frequently observed in groups with abundant bacilli discharge and long lasting cough, and also in young age groups. 3. All of the 19 cases causing epidemics or microepidemics had multiple risk factors; the main factors being late diagnosis, inadequate health managements in schools and offices and poor quality of patients' living environments. Therefore, it is necessary for public health centers to work more closely with schools and industrial circles for the prevention of tbc epidemics. PMID- 10723940 TI - [A study on tuberculosis cases among over-staying foreigners]. AB - An analysis was made on eighty-four cases of tuberculosis (TB) among over-staying foreigners during the past 9 years at Minatomachi Medical Center. All of them did not enroll in any health insurance system because they were illegal immigrants. Most of them were in their twenties and thirties. The ratio of male to female was four to one. By ethnic origin, the proportion was highest among Filipinos (30%), followed by Koreans (23%), Pakistanis (13%) and Indians (7%). Half of them had been in Japan for more than two years. Most patients sought care due to symptoms rather than as a result of the efforts of public health centers to screen foreigners. The proportion of extrapulmonary TB among all types of TB was 35% overall. Compared with pulmonary TB cases, patient's as well as doctor's delay was more marked among extrapulmonary TB cases. Forty cases were treated at Minatomachi Medical Center and forty-four cases were referred to another hospitals. The default rate including repatriation was as high as 41%, and the reasons for this high defaulting rate were as follows: 1. Language barrier, 2. Worry for loosing work during treatment, 3. Worry for high medical costs because of no coverage by a health insurance scheme, 4. Lack of information on medical systems and TB, 5. Worry for forced repatriation by the immigration office. In addition, some cases were not followed up due to unkind behaviour of attending physicians. The following measures are needed to prevent the epidemic of tuberculosis among overstaying foreigners and our societies. a) To provide free medical check-ups b) To provide easy access to medical facilities c) To utilize the tuberculosis prevention law d) To follow up patient thoroughly and strictly e) To explain TB in their mother languages f) To change the immigration law and its system g) To make liaison with other organizations such as medical facilities and NGOs, including foreign ones, and exchange informations h) To build good human relations with patients. PMID- 10723941 TI - [Evaluation of Tl-20 lung uptake and impairment of pulmonary perfusion on scintigraphies in pulmonary tuberculosis]. AB - Tl-201 lung uptake in 74 patients (85 lesions) and pulmonary perfusion in 105 patients were studied to evaluate clinical usefulness of Tl-201 lung uptake and perfusion lung scintigraphy in pulmonary tuberculosis, using a scintillation camera with a mini-computer system. As indices of Tl-201 lung uptake, lung (lesion) to upper mediastinum uptake ratio (L/M) and visual grading were used. L/M in pulmonary tuberculosis was 1.96 +/- 0.66, which was significantly larger than 1.04 +/- 0.24 in healthy controls and lower than that in heart diseases with left heart failure and idiopathic interstitial pneumonia, and showed no significant differences with that in acute pneumonia, pyothorax, primary lung cancer and malignant mediastinal tumor. L/M in pulmonary tuberculosis did not correlate with CRP, erythrocyte sedimentation rate, Gaffky number of sputum and body temperature. It correlated with the type of pulmonary tuberculosis according to the Gakken Classification reflecting the disease activity. It was larger in the exudative type, caseo-infiltrative one, disseminated one, one with cavity in infiltrative lesion than the fibro-caseous one. On perfusion lung scintigram, impairment of pulmonary perfusion larger than area of the entire unilateral lung was observed in 68 cases (64.8%). Area of hypoperfused lung field, which correlated with % vital capacity (r = 0.60, p = 0.0002) and PaO2 (r = 0.39, p = 0.0024), was significantly larger in patients with silicosis and those with bilateral pleural involvements such as pleural callosity than in those with type III according to the Gakkai Classification. Most of the patients showed decreased pulmonary perfusion and Tl-201 accumulation of which grade reflects the disease activity in active tuberculous lesion. Patients with miliary tuberculosis and those with silicotuberculosis showed diffuse Tl-201, accumulation in the both lungs. Tl-201 lung scintigraphy seems to be useful for visualizing active tuberculous lesions, particularly the ones that could not be detected by the chest radiograph in patients with destroyed lung and with pleural callosity. Joint use of Tl-201 and perfusion lung scintigraphies provides useful informations about the pathophysiology and disease process in pulmonary tuberculosis. PMID- 10723942 TI - Tuberculosis: recent progress in basic immunity and vaccine development. AB - Tuberculosis continues to be the most prevalent cause of death from an infectious agent globally, and its interaction with HIV is having devastating effects, particularly in Sub-Saharan Africa. Over the past decade, my laboratory has developed small animal models of pulmonary infection, which have revealed new information regarding the nature of acquired immunity, and subsequent immunopathology, in the lungs. We propose that cell mediated immunity comprises two separate elements; protective immunity, driven by IL-12 and IFN; and DTH, mediated by TNF and driven by chemokines. The generation of a CD4 response is critical to both processes, but other cells are also involved in the overall control of the infection. These include gamma delta T cells, which we believe control the inflammatory influx of cells; CD4+ NK cells, which may play a role in focussing lymphocytes into lung granulomas; and CD8 T cells, which play a currently undefined role after initial expression of immunity and establishment of chronic disease in the lungs has ensued. Complex interactions between these populations of cells appear to control the influx of mediator cells into the lungs and then focus them at sites of infection. Prior to adequate expression of protective immunity the correct expression of chemokine and adhesion molecules is critical. A better understanding of these processes will hopefully in turn lead to better vaccine design, a topic which is also addressed in this paper. PMID- 10723943 TI - [Current status of tuberculosis education in universities and future tasks]. AB - Symposium Topics and Presenters: 1. Education of tuberculosis in medial school: Kaoru SHIMOKATA (Department of Clinical Preventive Medicine, Nagoya University Daiko Medical Center) 2. From Medical University holding Tuberculous ward: Shosaku ABE (Third Department of Internal Medicine, Sapporo Medical University School of Medicine), et al. 3. Necessity and Significance of Sanatorium ward associated with University Hospital: Saburo SONE, et al. (Third Department of Internal Medicine, The University of Tokushima School of Medicine) 4. A proposal on education and training for tuberculosis in medical school from aspect of sanitariums: Takeshi OGURA (Toneyama National Hospital) 5. How to avoid infecting TB and to prevent contracting TB while medical and nursing practices: Keiichi NAGAO (Health Sciences, Center, Chiba University), et al. 6. Special speech: A review of the year since TB patient rooms were included in a common ward as a part of ministry of welfare's model project: Shuji KURANE (Fourth Department of Internal Medicine, Nippon Medical School) Tuberculosis began to rank first in mortality rate in Japan in the Meiji Era, and especially since it did not conform to the "national wealth and military strength" that was the national policy of the time due to the high mortality rates in the early decades of life, it was referred to as the "disease that was destroying the country" and the "pandemic disease." Even after entering the Showa Era, tuberculosis long occupied first place as the cause of death in Japan, and it raged unchecked for a period after World War II. However, the prognosis of tuberculosis as a whole improved considerably as a result of the development of antituberculosis agents, such as streptomycin, and the advent of rifampicin made it a curable disease. Its rank as a cause of death subsequently fell precipitously, and many of the TB wards that had been established in university hospitals were closed as the numbers of patients rapidly declined. At the present time, only 22 of the 80 university hospitals in the country have TB wards or TB beds, and 18 of the hospitals that had TB wards have closed them. Two of them closed them prior to 1964, 6 between 1965 and 1974, 4 between 1975 and 1984, 4 between 1985 and 1994, and 2 since 1995. Thus, it would be no exaggeration to say that there has been a steady decline in the TB wards of university hospitals. It is fairly easy to surmise that the result has been a decrease in the opportunities and time available for educating medical students about tuberculosis. Today, university hospitals not only accept medical students, but 80% of clinical residents as well, and they too have lost opportunities for education concerning tuberculosis. We would hope that the concern of Japanese physicians regarding tuberculosis has not diminished in proportion to the decrease in TB wards in our university hospitals. However, as is truly revealed by the expression "doctor's delay" in the diagnosis and treatment of tuberculosis, it is a fact that physicians no longer bear pulmonary tuberculosis in mind when diagnosing and treating patients with respiratory disease, and that as a result diagnosis is delayed, treatment is drawn out, and in the worst-case scenario, we see scattered instances of a tragic outcome. When we consider the recent conditions in society, as reported in recent newspapers, tuberculosis is not a disease that is on the decline at all in Japan today. However, as stated above, but if the concern of ordinary physicians has drifted away from tuberculosis, it is fair to say that it is not only a major problem medically, but socially as well. Consideration of the tuberculosis education in university medical schools seems to be opportune at this time. Professor Shimokata of Nagoya University, who is chairperson of the Japan Tuberculosis Education Committee, outlined the problems of tuberculosis education in his speech at this symposium. (ABSTRACT TRUNCATED) PMID- 10723944 TI - [Basic and clinical aspects of bronchus-associated lymphoid tissue]. AB - Human mucosal membrane surfaces have a distinct protective mechanism that is specifically designed to produce secretory IgA. In the airway, bronchus associated lymphoid tissue (BALT), is known to be an inductive site of secretory IgA. BALT is a lymphoid aggregate located in the submucosal area of bronchioles, and plays a central role in airway mucosal immunity by inducing the accumulation of secretory IgA-producing cells. Although previously it had been unclear whether BALT is present in the human lung, we demonstrated the expression of BALT in patients with diffuse panbronchiolitis, chronic hypersensitivity pneumonitis, and collagen disease-associated lung diseases. BALT was shown to elevate serum IgA levels and in other ways to have an influence on the symptoms and signs in patients with these pathologic conditions. We also demonstrated that continuous inhaled antigenic stimulation as well as the local production of interleukin-4 and other cytokines promote BALT development, which, in turn, may give rise to BALToma, a type of primary pulmonary mucosa-associated lymphoma. PMID- 10723945 TI - [A simple personal computer-based system for lung sound analysis]. AB - Although lung sounds provide important information about the respiratory system, the analysis of lung sounds has not been widely used in clinical practice because of the complicated procedure involved. However, personal computer technology has made impressive strides in recent years. Today, practically all personal computer models on the market are equipped with the capacity for audio signal input and output. We developed a new computer system for lung sounds acquisition and analysis. The system hardware comprises only a personal computer and a microphone, and the software was developed for a widely used operating system (Windows 95). Our system can record, save, and replay lung sounds and analyze their time and frequency domains. To verify the accuracy of sound acquisition, we examined the frequency characteristics of the system as installed and utilized on 4 different machines. The characteristics were essentially flat throughout the 200-2,000 Hz spectrum within which almost all lung sounds were contained. We feel our system can serve as a simple and useful tool for lung sound analysis. PMID- 10723946 TI - [Nocturnal oxygen desaturation during home oxygen therapy in patients with chronic respiratory disease]. AB - We investigated nocturnal oxygen desaturation (NOD) in 36 patients with stable chronic respiratory disease who were receiving home oxygen, therapy (HOT). Study data included medical history, chest roentgenograms, measurement of daytime arterial blood gases while awake, and spirometry. Each subject underwent full overnight oximetry monitoring. Three patients were excluded from further investigation because of periodic desaturation suggestive of sleep apnea. The remaining 33 subjects were divided into two groups: 21 patients with sequelae of pulmonary tuberculosis (TB-sequela) and 12 patients with chronic obstructive pulmonary disease (COPD). The COPD group was divided into two subgroups according to the Burrows classification (Am Rev Resp Dis. 90: 14-27, 1964): 5 patients with type A (Type A) and 7 patients with type B (Type B) COPD. The percentages of total sleep time with SaO2 < or = 85% (DST 85) and SaO2 < or = 90% (DST 90) were calculated for each subject. NOD was defined as DST 85 > or = 1%. Arterial oxygen partial pressure (PaO2) while awake was > or = 60 Torr in all subjects. No difference was observed in mean awake PaO2 values between the TB-sequela and COPD groups. NOD was detected in 8 TB-sequela patients but in none of the COPD patients. Mean DST 85 and DST 90 values were significantly (p < 0.05) higher for the TB-sequela group than for the COPD group. Of 15 TB-sequela patients who were able to complete spirometry tests, 6 had NOD. All 6 of these patients had hypercapnia while awake (PaCO2 > or = 50 Torr) and reduced vital capacity (< or = 50% predicted). No difference was observed in mean DST 90 or DST 85 values between the TypeA and TypeB COPD subgroups. We conclude that NOD is common in patients with chronic stable respiratory disease treated with HOT despite daytime euoxia. TB-sequela patients with hypercapnia and restrictive ventilatory impairment are at high risk for NOD. PMID- 10723947 TI - [Successful treatment of steroid-resistant bronchiolitis obliterans-organizing pneumonia with orally administered cyclosporin and pirfenidone]. AB - A 27-year-old man with fever and dyspnea was admitted to our hospital. Chest computed tomography and a lung biopsy were performed, and bronchiolitis obliterans-organizing pneumonia (BOOP) was diagnosed. The patient was treated with corticosteroid, and a marked improvement was noted. However, when the dosage was tapered, BOOP recurred. Although the dosage was increased again, the corticosteroid alone was no longer effective against BOOP. While continuing with corticosteroid therapy, we also put the patient on a daily regimen of cyclosporin and pirfenidone, a recently developed anti-fibrotic agent. Both drugs were administered orally, and were so effective that we gradually decreased the dosage of corticosteroid. Several journals have reported that cyclosporin may be effective in the treatment of interstitial pneumonitis associated with collagen disease. We concluded that cyclosporin may also be useful in the treatment of refractory BOOP. PMID- 10723948 TI - [Right tracheal bronchus with anomalous ramification of the bronchial artery disclosed during an episode of hemoptysis]. AB - A 63-year-old woman was referred to our hospital on June 18th, 1998 during an episode of hemoptysis that had lasted for 6 days. She had no hemorrhagic diathesis and no history of pulmonary disease. Chest X-ray films disclosed a ground-glass opacity in the right upper lung field. Bronchoscopic examination revealed bleeding from an anomalous ectopic orifice on the right lateral trachea, about 1 cm above the carina. Chest computed tomographic examinations by conventional and spiral methods readily disclosed an ectopic bronchus. Bronchial arteriography showed that the tracheal bronchus was fed by a branched vessel of the thyrocervical artery arising from the brachiocephalic artery. Atypical mycobacterium was detected in bronchoalveolar lavage fluid from the ectopic bronchus. A shunt had formed with the pulmonary artery and peripheral parts of the bronchial artery that fed the tracheal bronchus. It was speculated that the hemoptysis in this case might be due to the combined phenomena of infection and abnormal vessel formation in the tracheal bronchus. In our patient, the system of blood supply to the tracheal bronchus may have been a manifestation of atavism because it closely resembled the circulatory structure of the tracheal bronchi normally observed in sheep and giraffes. The tracheal bronchus should be taken into consideration as a potential cause of hemoptysis, inflammatory changes, and atelectasis during intubation. PMID- 10723949 TI - [Two cases of intravascular lymphomatosis diagnosed antemortem by transbronchial lung biopsy]. AB - Patient 1 was a 71-year-old man who had been admitted to our hospital with high grade fever. Chest computed tomographic (CT) images revealed clear peripheral and high-density central areas. Abnormal laboratory findings included elevated LDH and hypoxemia. Interstitial pneumonia was suspected, and transbronchial lung biopsy (TBLB) confirmed the diagnosis of intravascular lymphomatosis (IVL). The patient succumbed before completion of chemotherapy. Patient 2 was a 65-year-old man admitted with high-grade fever. Abnormal laboratory findings included pancytopenia, hypoxemia, and elevated levels of LDH and soluble interleukin-2 receptor. Chest CT images revealed diffuse, mildly dense areas in the upper fields of both lungs. TBLB specimens yielded a diagnosis of IVL. Complete clinical remission was obtained with CHOP multiagent chemotherapy. Although IVL is usually diagnosed at autopsy, in these 2 cases an antemortem diagnosis was made on the basis of TBLB findings. Also, multiagent chemotherapy achieved a complete clinical remission in Patient 2. PMID- 10723950 TI - [Metastatic squamous cell carcinoma of hilar lymph node with unknown primary site]. AB - An abnormal shadow was observed on chest X-ray films of a 63-year-old man presenting with cough and sputum. Chest computed tomographic scans disclosed enlargement of the right hilar lymph nodes, but no obvious primary lesion was found in the lung field. Bronchoscopic examination revealed a slightly widened second carina, but no malignant cells were detected by transbronchial aspiration cytology. At surgery, a tumor was found between the truncus superior and the truncus intermedius. The pathologic diagnosis was a metastatic lymph node of poorly differentiated squamous cell carcinoma. Because the tumor severely adhered to the bronchus and pulmonary arteries, we performed a right pneumonectomy with mediastinal node dissection. Pre- and postoperative examinations did not detect the primary lesion, and no recurrence had been observed 76 months after surgery. This was thought to be a very rare case of T0 N1 M0 lung cancer. In general, the prognosis is poor for patients with metastatic carcinoma of unknown primary site. However, patients with T 0 lung cancer, as in this case, might enjoy a better prognosis if complete resection and dissection of metastatic lymph nodes are performed. PMID- 10723951 TI - [Anaerobic bacillus pyothorax with the production of gas and severe mediastinal shift]. AB - A 62-year-old man was admitted with the complaints of chronic sputum, dyspnea, and general weakness. Chest X-ray and computed tomographic films disclosed severe mediastinal shift and left lung collapse due to the accumulation of fluid and gas in the left pleural space. A puncture of the thoracic cavity yielded a milk coffee-like purulent pleural effusion with stool odor, suggesting pyothorax with pneumothorax or broncho-pleural fistula. Chest tube drainage was performed. The elimination of gas was transient; subsequently, no air leaks were observed during deep breathing, suggesting the absence of pneumothorax and broncho-pleural fistula. An anaerobic culture of pleural effusion was prepared and a Bacteroides species was isolated. These clinical findings indicated that the intrathoracic gas could have been produced by anaerobic bacilli. Systemic antibiotic chemotherapy with chest tube drainage achieved recovery. The production of gas in focal lesions is one noted symptom of anaerobic bacillus infection. However, to our knowledge, cases of anaerobic bacillus pyothorax generating large volumes of intrathoracic gas are rare. PMID- 10723952 TI - [Multiple Aspergillus brain abscesses complicated by bronchial asthma]. AB - A 35-year-old man was hospitalized for the treatment of severe asthma attack. His condition improved with intensive steroid chemotherapy under artificial ventilation. On the 12th hospital day, he was taken off respirator support but lost consciousness afterward. Computed tomography of the brain disclosed multiple hypodense lesions with bleeding. T1-weighted magnetic resonance imaging disclosed low-intensity lesions containing high-intensity areas. T2-weighted images showed heterogeneous high-intensity lesions. The autopsy specimen demonstrated multiple brain abscesses. Histologic examination revealed branching fungal hyphae in abscess walls and also extending through arterial walls with emboli. These findings yielded a diagnosis of multiple Aspergillus infarct abscesses of the brain. PMID- 10723953 TI - [Intrathoracic neurofibroma originating in the left vagus nerve]. AB - A 20-year-old man was admitted because of an abnormal mass shadow on chest X-ray film. Computed tomography (CT) and magnetic resonance imaging (MRI) disclosed a mass lesion in the superior portion of the left mediastinum. CT scans showed a well-defined mass with low density. Axial MRI rendered the mass lesion with intermediate signal intensity on T1-weighted images and high signal intensity on T2-weighted images. The preoperative diagnosis was bronchogenic cyst. Video assisted thoracic surgery revealed that the tumor originated in the truncus of the left vagus nerve. The resected tumor was 90 x 24 x 18 mm in size. The postoperative course was uneventful and hoarseness did not develop. The pathologic diagnosis was benign mediastinal neurofibroma without von Recklinghausen's disease. Such cases are extremely rare in the Japanese literature. PMID- 10723954 TI - [Multiple nodular pulmonary amyloidosis with multiple bullae]. AB - We report a case of multiple nodular amyloidosis accompanied by multiple emphysematous bullae. The patient was a 72-year-old nonsmoking woman who presented with chronic cough and had a history of pneumothorax. Chest computed tomography disclosed multiple nodules up to 1 cm in diameter, and multiple small emphysematous bullae predominantly distributed in the middle and lower lung fields. Left lung biopsy was performed under thoracoscopy. The nodules were histopathologically diagnosed as amyloidoma. The mechanism of the accompanying bullae formation was also considered. PMID- 10723955 TI - [Testicular sarcoidosis]. AB - We report a rare case of testicular sarcoidosis. A 68-year-old man was admitted for detailed examination of uveitis and swelling of the testes. A chest X-ray film and computed tomographic scans disclosed ground-glass shadows in the lower fields of both lungs with mediastinal lymphadenopathy. Ga scintigram showed pronounced accumulations in the testes, hilum, and mediastinum. Transbronchial lung and testicular biopsy specimens demonstrated noncaseating epithelioid granulomas, thus confirming the diagnosis of sarcoidosis with testicular involvement. The patient was followed up without systemic steroids. A review of the world literature found only 12 reported cases of clinically evident testicular sarcoidosis. PMID- 10723956 TI - [Nontuberculous mycobacterial infection followed for 12 years]. AB - A 67-year-old woman presented in September 1985 with productive cough, bloody sputum, and dyspnea on exertion. Productive cough and bloody sputum had developed when the patient was 55 years old. Sputum culture and radiologic findings yielded a diagnosis of nontuberculous mycobacteriosis (NTM). Antituberculous therapy with INH, RFP, and EB was initiated in November 1987 because of the development of a cavity in the right upper lobe, and led to resolution of the lesion and clinical symptoms. Despite progression of bronchiectatic changes in both lungs and a relapse of her clinical symptoms during the following 10 years, the patient retained enough pulmonary function to be able to maintain an active daily life until she died of advanced gastric cancer at the age of 79. Autopsy revealed cystic bronchiectasis accompanied by bronchial wall thickening in both lungs, with some granuloma and acid-fast-bacteria observed in lung tissue. In this report, we concluded that patients with NTM usually experience a gradual progression of symptoms and radiographic changes during their clinical course, and that their pulmonary function may be conserved well enough to maintain an active daily life. PMID- 10723957 TI - [Pulmonary hypertrophic osteoarthropathy associated with primary lung cancer]. AB - The patient was a 61-year-old man admitted with the complaints of cough, arthralgia, and swelling of the legs. A chest roentgenogram and chest computed tomographic scan revealed a giant mass in the right upper lobe. Transperitoneal lung biopsy was performed, and a diagnosis of poorly differentiated adenocarcinoma was made. Physical examination confirmed swelling of the legs and clubbing of fingers on both hands. Bone scintigrams showed marked accumulation of 99 m-Tc-MDP in the long bones, bones of the hands, and patellae. These findings yielded a diagnosis of pulmonary hypertrophic osteoarthropathy associated with primary lung cancer. Although a high serum level of growth hormone was also detected, immunohistochemical analysis did not find growth hormone in the tumor itself. Chemotherapy and radiotherapy were performed but did not stop progression of the disease. The patient subsequently experienced worsening arthralgia and swelling of the legs. Steroid therapy rapidly alleviated the arthralgia and swelling, but not the clubbing of the fingers. Thereafter, the patient's serum CRP and ICTP dropped to normal levels, and the abnormal findings of bone scintigrams subsequently disappeared. The pulmonary hypertrophic osteoarthropathy was not clearly attributable to growth hormone. Steroid therapy was effective in this case. Bone scintigrams and serum CRP and ICTP may be useful indicators in the therapeutic follow-up and monitoring of patients with pulmonary hypertrophic osteoarthropathy. PMID- 10723958 TI - [Searching for a new risk factor of sudden cardiac death]. AB - Sudden death of cardiovascular origin has been recognized as a major cause of sudden natural death. Advanced coronary atherosclerosis often co-existent, despite this, little or no coronary atherosclerosis was seen in approximately half of the sudden coronary death cases. Some components in the blood seem to induce not only the coronary atherosclerosis but also coronary spasm and/or thrombogenesis under certain conditions, which does not necessarily relate to the severity of atherosclerosis. Postmortem plasma lipids and lipoprotein levels, especially triglyceride-rich lipoprotein remnants (abbreviated as remnants) were measured as remnant-like particles-cholesterol (RLP-C) and triglyceride (RLP-TG) in two groups of Japanese subjects who died suddenly and unexpectedly due to coronary or non-coronary causes. Our study on the postmortem plasma analysis of lipids and lipoproteins indicated that RLP-C was most strongly correlated with the severity of the coronary atherosclerosis and to be the best predictor among various parameters. Furthermore, RLP-TG appeared to be highly associated with the risk of sudden coronary event or death, especially in cases without advanced coronary atherosclerosis, which might be associated with clinical events as coronary spasm or thrombosis. Biological activities of RLP from postmortem plasmas of sudden coronary death cases showed strong effects on platelet aggregation and impaired vasorelaxation in vitro. RLP determined as RLP-TG might affect on the process of spasm or on the formation of thrombus in the coronary artery at the time of sudden coronary death, independent of the severity of coronary atherosclerosis. These data might be clinically important to predict or to prevent sudden coronary death. The studies were conducted as followed: 1) Separation, purification and quantitative analysis of remnants from postmortem plasma. 2) Isolation and characterization of remnants from postmortem plasma. 2 1) Assessment of the methods of TG measurement. 2-2) Characterization of remnants by HPLC methods. 2-3) Characterization of remnants by electrophoresis. 2-4) Electron microscopical observation of remnants. 3) Lipid and lipoprotein analysis of sudden coronary death and control cases. 3-1) Lipid and lipoprotein levels of sudden coronary death and control cases. 3-2) Plasma remnant levels in the postprandial state. 3-3) Severity of coronary atherosclerosis and plasma lipid and lipoprotein levels, especially remnant levels. 3-4) Sudden coronary death cases without advanced coronary atherosclerosis and their plasma lipid, lipoprotein levels, especially remnant levels. 4) Biological activities of the remnants. 4-1) Endothelium-dependent vasodilator function in the coronary artery. 4-2) Aggregation of human blood platelets by remnants. 5) Fatty acids analysis of remnants from sudden coronary death cases. 6) Immuno-histochemical examination of various apo-lipoproteins on the coronary artery. 7) Gene analysis of plasma lipid metabolites. PMID- 10723959 TI - [Forensic DNA analysis--past and future]. AB - Since the introduction of DNA polymorphism analysis techniques to forensic science, forensic identification research has made radical, astonishing progress at a rate that has already rendered the initial methodologies introduced fifteen years ago obsolete. DNA extraction now can be quickly and efficiently performed by various kinds of commercially available kits. The advent of PCR has enabled the use of relatively crude and minute DNA as amplification templates while many kinds of new detection methods for analyzing the amplified products have also been developed. Although many minisatellites such as MCT118, YNZ22, COL2A1, and ApoB were highlighted at the beginning of 1980s, none of these loci, with the exception of MCT118, have proved useful for forensic DNA application due to their low amplification efficiency. On the other hand, STR loci containing four base pair repeat sequences have been used routinely for human identification since the mid-1990s. In the near future, the highly efficient STR should be selected as a consensus core marker in Japan. STR systems located on the Y chromosome are widely used in forensic science for the identification of male individuals. These systems have a special significance in forensic science cases where mixtures of male and female DNA are analyzed, as happens in cases of rape or other sexual crimes. The characteristics of high copy number, maternal inheritance, and high degree of sequence variability make mtDNA a powerful tool for forensic identification. Most of the variations in mtDNA among individuals are found within the displacement loop (D-loop). In all population groups, mtDNA sequences can be useful for discriminating among unrelated individuals. Now it is necessary to get as much as possible individual genetic information as quickly as possible in order to enable individual identification. We will create a new era in which forensic identification can be performed using microarray technology. PMID- 10723960 TI - [Biological actions of acetaldehyde]. AB - Acetaldehyde (AcH), the first metabolite of ethanol (EtOH), is a chemically reactive and pharmacologically active compound. The author has been engaged in the study of AcH in cooperation with many researchers for three decades. We have found many biological actions of AcH which cause cardiovascular symptoms after drinking and also inhibited EtOH absorption via the canine and rat intestinal tract. This report covers the following five points. 1. The subjects were classified into a non-flushing group and a flushing group, according to the degree of facial flushing after drinking 200 ml of Sake (Japanese rice wire) at a rate of 100 ml per 5 min. Blood EtOH profile was much the same in both groups, yet peak blood AcH concentration in the flushing group was significantly higher than that in the non-flushing group. All subjects in the flushing group showed marked flushing and an increase in pulse rate after drinking, but these symptoms were not apparent in the non-flushing group. These results suggested that cardiovascular symptoms were caused by AcH itself. 2. Urinary excretions of both norepinephrine and epinephrine increased in the flushing cases after drinking Sake in comparison with those who drank the same volume of water. However, these catecholamines did not change in the non-flushing group. These results suggested that it is catecholamines released from the sympathetic nerve end or the adrenal medulla by AcH which caused an increase in pulse rate. 3. Bradykinin is released from high molecular kininogen by activated kallikrein and acts to dilate distal blood vessels and raise permeability in tissues. On the other hand, kallidin is released from low molecular kininogen by activated glandular kallikrein and its action is weaker than that of bradykinin. Blood low molecular kininogen levels in the flushing group decreased gradually after drinking and were mutually related to the blood AcH concentrations. But levels in the non-flushing group showed no difference before and after drinking. The decrease in low molecular kininogen levels indicates that kallidin released from glandular kallikrein exists in the glandular tissues such as the kidneys, sweat glands, saliva glands, etc. We hypothesize that kallikrein activated by AcH in the sweat glands produces kallidin which cause vessels around the glands to dilate, and flushing of the face and the whole body occurs due to escalation of the sphere of dilatation of blood vessels. 4. A isolated 30 cm length of the canine jejunum segment with intact vascular supply was performed. After pretreatment with cyanamide (CY), a potent inhibitor of aldehyde dehydrogenase, or pyrazole (PY), a potent inhibitor of alcohol dehydrogenase, a 17% EtOH solution (0.4 g/kg) was administered into the jejunum segment, and 150 min after the administration of EtOH, the fluid from the segment was collected to determine its volume and EtOH concentration. The CY pretreatment group, in which an extremely high AcH concentration developed, in comparison with the control and PY-pretreatment groups, showed a gradual increase of portal blood EtOH, a 25% reduction in the amount of absorbed EtOH, and an 85% smaller absorption rate constant value (Ka value). These facts indicate that the presence of a high AcH concentration in the blood results in a reduction of EtOH absorption and retardation of EtOH reaching the systemic circulation. The rapid reduction of portal blood flow and lower EtOH level in the portal vein observed in the CY group, in comparison with the other groups, also indicate that the reduction of EtOH permeability through the absorption site to the blood is an important retarding factor induced by AcH. 5. After segmenting a 20 cm length of rat intestine, cannulae for EtOH perfusion were inserted into each end of the intestine segment. Perfusion of EtOH solution (4%) was performed for 30 min at steady rate, beginning 60 min after pretreatment with CY and/or PY. The blood EtOH and AcH concentrations in the f PMID- 10723961 TI - [Forensic autopsy of "hardship" without a well-equipped, adequately staffed toxicological laboratory]. AB - With a limited budget, it is difficult for our department to have a well-equipped toxicological laboratory with sufficient members of trained personnel. Alcohol levels in the body fluids and carbon monoxide levels in the blood are routinely measured using gas chromatography and UV spectrophotometer, respectively. Some drugs can be detected by the drug screening test on the market, such as Triage test. When poisoning is certain in a criminal case, I entrust another forensic laboratory with analyzing. In some non-criminal cases, a clinical laboratory of a private company may be chosen. In other cases, however, the samples are only kept in a freezer. In case of outside order, a guideline to provide an adequate chain of custody will be necessary. PMID- 10723962 TI - [Strategy for investigation of forensic autopsy cases where drugs or poisonous substances are involved]. AB - In recent years, the number of drug- or toxin-related criminal cases has gradually increased, and accordingly the establishment of precise forensic autopsy examinations and appropriate chemical analysis is of urgent importance so that such cases are not overlooked. To this end, the author has set up a general instructional objective comprising "preparation of the most suitable analytical procedures for elucidation of drug- and/or poison-related cases in a forensic medicine laboratory". Specific objectives within the general instructional objective consist of the following: 1) securing adequate manpower, 2) swift acquisition and accumulation of the latest scientific information, 3) sufficient examinations during forensic autopsy, 4) appropriate collection and storage of human tissues, 5) enforcement of preliminary tests and routine examinations, 6) selection of proper analytical procedures and instruments for qualitative and quantitative analysis, 7) establishment of reliable quantitative analysis, 8) signification of quantitative data, 9) preparation of an accurate analytical report. In our laboratory, we have been able to analyze many kinds of acidic and basic substances as well as volatile hydrocarbons, alcohol and carboxyhemoglobin through the use of absorptimetry, gas chromatography, gas chromatography/mass spectrometry and high performance liquid chromatography, however it is thought that more skillful techniques should be developed as soon as possible with regard to the analysis of inorganic and water-soluble agents. PMID- 10723963 TI - [Isn't there a gap between ideal and real actions in legal toxicology?]. AB - I showed a view for some problems, which need to be solved, in the process of post mortem examinations and chemical analyses of deaths from poisoning. Basic education program of analytical chemistry has some problems for medical students, post graduated persons and analysts. Analytical procedures have also problems in quality assurance system. I expressed the necessity of "an analytical center of poisoning", following I showed a plan as a starting point for "the analytical center". PMID- 10723964 TI - [The cooperation system for drug analysis by computer network]. AB - Generally, the drugs are analyzed by the forensic medicine department in the most of autopsy cases. The forensic medicine department cannot always perform toxicological analysis because a shortage of a verified personnel, appropriate instruments, and financial resources. In order to overcome these difficulties and lack of resources, we developed a computer network called ml-poison. This network consists of the staff of forensic medicine, hospitals, governmental agencies and research facilities of police departments, etc. This network offers immediate and valuable information and expertise to these inexperienced in dealing with cases of poisoning. In cases in which the toxicological analysis cannot be performed in a facility, the network will recommend a facility where the analysis can be performed. Up this point, the network order system has assisted in many cases of poisoning. Although the effectiveness of the network for toxicological analysis has been proven, we still must deal with severed difficult problems: 1. The number of facilities assisting network orders is limited. 2. Who will pay the expenses involved in the analysis. 3. How to maintain security of the system. 4. What agency will assume responsibility for the management of the system. PMID- 10723965 TI - [Present status of forensic analyses and possible approach for a rapid identification of toxins]. AB - Frequency of public harm that associated with hazardous chemicals has been increasing in the last several years. These incidents involve nerve gas attacks on Matsumoto-city and on the subways in central Tokyo, doping in sports, amphetamines abuse problem among adolescent boys and girls, potential health risks due to a contamination of environment by endocrine disrupters, and scheduled contamination of foods and drinks by several toxic compounds. US government has learnt from unexpected Sarin-affairs in Japan, and fast-track defensive action has been taken in the USA for a better safe of general public. The Japanese system for forensic analyses consists of a number of in organizational laboratories that founded in each individual universities, hospitals and police offices etc. Such system allows laboratories to fit regional requirements and to achieve highly professional outcome on a specific area depending on their scientific interests. On the other hand, such situation sometime can cause difficulty in timely identification of certain compounds, which are not frequently analyzed in their laboratories. This paper refers to the possible bottlenecks, critical points and key success factors for a rapid identification of toxins. Two stage testing systems, namely, a) simple rapid examination such as one-step specific immunoassays for on-site emergency testing, and b) following conclusive identification of the compounds in the nearest expert laboratories seemed to be effective and practical corrective measures for the problems that were focused in the recent crises. Documentation of the course of action to be taken in case of emergency, improved supply system of reference standards, circulation of information through the Internet mailing list or other communication infrastructures, use of quality control program, and financial supporting of other educational programs for analyses, may improve the situation. PMID- 10723966 TI - [Isolation of type-A blood group active glycoproteins from salivas and their reaction with monoclonal antibodies]. AB - We have previously examined several ABO blood grouping antibodies with whole saliva and observed that there were two types of antibodies. One group of antibodies reacted with both secretory and non-secretory saliva, and the other with only secretory saliva. In order to clarify the differences in reaction of the antibodies with saliva, we needed to analyze of antigens in secretory and non secretory saliva. Therefore, we prepared blood group active glycoproteins from secretory and non-secretory saliva by affinity chromatography and gel filtration. The secretory saliva gave three active peaks, one large peak in the void volume and two small peaks in fractions of smaller molecular weights on Sepharose-CL6B after the affinity chromatography. From the non-secretory saliva, a single active peak in the void volume, which corresponded to the major peak in the secretory saliva, was found. Most blood group activities were found in those void volume fractions. Moreover, capillary electrophoresis showed that these blood group active glycoproteins were identical, and that there were immunological differences between secretory and non-secretory salivary blood group substances. PMID- 10723967 TI - [Blood-group genotyping by constant denaturing gel electrophoresis (CDGE)]. AB - Constant Denaturing Gel Electrophoresis (CDGE) was used as an analytical method of the genetic markers. Even a single base change in a fragment amplified by PCR was detected exactly by CDGE. The computational simulation of CDGE gave the calculation whether a single base change in a fragment amplified by PCR could be detected by CDGE or not. In this report, genotyping of three blood groups, MN, Duffy and Kidd, and Gc system is described. The regions reflecting allelic differences of each system were amplified from genomic DNAs. The concentration of denaturants (urea and formamide) in CDGE gels was decided with the computational simulation as follows: 15% for MN, 27% for Duffy, 24% for Kidd, 30% for Gc. CDGE was run in TBE buffer at 60 degrees C, 100 V constant voltage. PCR amplified fragments with 1-3 base changes were separated clearly in each gel. By staining the gels with ethidium bromide, the genotype of each system was determined. The genotyping of system by CDGE can avoid mistakes in the conventional method, which requires complicated and multiple troublesome operations. Analysis of PCR amplified fragments by CDGE will make a beneficial contribution to medico-legal practice. PMID- 10723968 TI - [Improvement of absorption-elution test using commercially available anti-A, anti B monoclonal antibodies--ABO blood typing from hair samples]. AB - An improved procedure for ABO blood typing by the absorption-elution test using commercially available monoclonal antibodies was established. The optimized elution temperature for anti-A monoclonal antibodies to be liberated from bloodstains was 54 degrees C and the temperature to deactivate the liberated antibodies was over 56 degrees C. The test condition was established using a monoclonal antibody manufactured by Biotest. For the anti-A monoclonal antibody the most effective elution time was 5 min and the long incubation time decreased the activity of the liberated antibodies. On the other hand, the conditions formerly used for absorption-elution tests were suitable for anti-B monoclonal antibody. A Type-A test and a Type-B test have to be performed separately for ABO blood grouping from forensic samples. The conditions established in this report were applied to the absorption-elution test to achieve an improved result for ABO blood grouping from hair samples. PMID- 10723969 TI - [An autopsy case of pulmonary thromboembolism associated with chlorine gas poisoning]. AB - We report a rare case of sudden death of a patient with acute pulmonary thromboembolism associated with chlorine gas poisoning. A 21-year-old man in a water-filtration plant accidentally inhaled highly concentrated chlorine gas. He was immediately brought to a hospital after exposure. On admission, the patient had clouding of consciousness, dyspnea, and deep cyanosis. Arterial blood gas values indicated severe hypoxemia; PaO2 was 35.9 mmHg and PaCO2 was 42.4 mmHg. The clinical course was uneventful and he was satisfactorily recovering. However, ten days after admission he became sick and markedly cyanotic. He lost consciousness and then he went into cardiopulmonary arrest. Despite efforts at resuscitation, he died. An autopsy revealed bilateral pulmonary thromboembolism, although he apparently did not have any risk factor for embolism. The toxicity of chlorine gas may be related to the pulmonary thromboembolism, but the mechanisms leading to his death are unclear. PMID- 10723970 TI - [An autopsy case of a bicycle accident with ring fracture at the base of the skull]. AB - We report the autopsy case of a 41-year old passenger who suffered a significant head injury with a typical ring fracture at the base of the skull as a result of a violent fall from a bicycle. Several reports about ring fractures of the base of the skull revealed that they were due to crashing a car at high speed, a collision and/or a fall while riding a motorcycle and a fall in piloting a gyrocopter and so on resulting in severe injury to another part of the body. In this case, the ring fracture occurred when his spine was pushed up by high impact of the parieto-occipital region against the ground. PMID- 10723971 TI - Effect of age on the outcome of angioplasty for acute myocardial infarction among patients treated at the Mayo Clinic. AB - PURPOSE: Elderly patients, especially those 80 years of age and older, have been excluded from most studies of thrombolysis or primary coronary angioplasty in patients with acute myocardial infarction. We compared the outcomes of elderly patients who underwent coronary angioplasty with the outcomes of younger patients and determined whether there were any temporal trends in survival. PATIENTS AND METHODS: We reviewed the outcomes of 1,597 consecutive patients who underwent primary coronary angioplasty between 1979 and 1997, including 127 patients who were 80 years of age or older (mean [+/-SD] age, 83 +/- 3 years, 47% male). Their in-hospital and long-term outcomes were compared with those of 524 patients who were 70 to 79 years old, 527 patients who were 60 to 69 years old, and 419 patients who were 50 to 59 years old. The oldest group of patients was divided into two groups, based on whether they had intervention through the end of 1993 (n = 56) or between 1994 and 1997 (n = 71). The survival rate of the patients who had no complications and left the hospital was compared with expected survival based on age- and sex-adjusted data. RESULTS: Patients 80 years of age or older had more adverse baseline characteristics, including risk factors and comorbid conditions, than the younger patients. The clinical success rate of primary angioplasty in this group was lower than those in the other three groups (61% versus 74% in those aged 70 to 79 years, 73% in those aged 60 to 69 years, and 81% in those aged 50 to 59 years, P < 0.001). The in-hospital mortality rate among patients 80 years of age or older was significantly greater than among patients in the other three groups (21% in those aged 80 years or older, 13% in those aged 70 to 79 years, 9% in those aged 60 to 69 years, and 4% in those aged 50 to 59 years, P < 0.001 ). The clinical success rate of the angioplasty improved significantly in the more recent period (75% versus 45%, P = 0.0006) and in-hospital mortality declined (16% versus 29%, P = 0.07). During follow-up, mortality in the oldest age group in whom angioplasty was successful was significantly greater than in the three younger groups, but was similar to the expected survival in the general US population. CONCLUSIONS: The mortality associated with primary angioplasty for acute myocardial infarction in octogenarians remains high, although there has been significant improvement in the clinical success rate. The long-term prognosis following a successful angioplasty is not different from that in an age- and sex-adjusted U.S. white population. PMID- 10723972 TI - Magnesium sulfate as a vehicle for nebulized salbutamol in acute asthma. AB - PURPOSE: Magnesium sulfate is thought to be an effective bronchodilator when administered intravenously to patients with acute severe asthma, and it can be safely administered via inhalation to patients with stable asthma. Our goal was to determine if isotonic magnesium sulfate could be used as a vehicle for nebulized salbutamol for patients with acute asthma. METHODS: We enrolled 35 patients with acute asthma in a randomized, double-blind, controlled trial. After measurement of peak expiratory flow, patients received 2.5 mg salbutamol plus either 3 mL normal saline solution (n = 16) or isotonic magnesium sulfate (n = 19) through a jet nebulizer. Peak flow was reassessed 10 and 20 minutes after treatment. RESULTS: Peak flow at baseline was similar in the two groups. Ten minutes after baseline, the mean (+/- SD) percentage increase in peak flow was greater in the magnesium sulfate-salbutamol group (61% +/- 45%) than in the normal saline-salbutamol group (31% +/- 28%; difference = 30%; 95% confidence interval [CI] for the difference: 3% to 56%; P = 0.03). At 20 minutes, the percentage increase in peak flow was 57% greater in the magnesium sulfate group (95% CI: 4% to 110%, P = 0.04). There was a significant inverse correlation between baseline peak flow (percent of predicted) and the percentage increase in peak flow at 20 minutes in the magnesium sulfate group (r = -0.82, P <0.0001), but not in the saline group (r = -0.12, P = 0.67). CONCLUSION: In patients with acute asthma, isotonic magnesium sulfate, as a vehicle for nebulized salbutamol, increased the peak flow response to treatment in comparison with salbutamol plus normal saline. PMID- 10723973 TI - Autoimmune hemolytic anemia in patients with systemic lupus erythematosus. AB - PURPOSE: We sought to evaluate the clinical and serologic associations with, and outcomes of, autoimmune hemolytic anemia, as compared with other types of anemia, in patients with systemic lupus erythematosus (SLE). SUBJECTS AND METHODS: We studied 41 consecutive patients with SLE with clinically manifest autoimmune hemolytic anemia, including 27 (66%) in whom hemolysis was the initial disease manifestation. We matched each patient for age and disease duration with a patient with SLE with anemia resulting from a different cause. RESULTS: The 41 patients had a total of 50 episodes of autoimmune hemolytic anemia. The recurrence rate was 4 per 100 person-years. Cases and controls had similar mean (+/- SD) lupus activity indexes (2.1 +/- 1.5 vs 2.4 +/- 1.3, P = 0.5). Patients with autoimmune hemolytic anemia at any time could be distinguished from patients with other causes of anemia, because they were more likely to have elevated titers of IgG anticardiolipin antibodies [odds ratio (OR) = 5.8; 95% confidence interval (CI), 1.4 to 24] and thrombosis (OR = 4.6; 95% CI, 1.0 to 21). Autoimmune hemolytic anemia at the onset of SLE was independently associated with renal involvement (OR = 5.4; 95% CI, 1.0 to 28), thrombocytopenia (OR = 7.3; 95% CI, 1.1 to 48), and possibly thrombotic episodes during follow-up (OR = 11; 95% CI, 0.8 to 160) when compared with controls with other types of anemia at the onset of SLE. CONCLUSIONS: Autoimmune hemolytic anemia usually occurs at the onset of SLE, and its recurrence rate is low among treated patients. The association with IgG anticardiolipin antibodies and thrombosis suggests that the occurrence of autoimmune hemolytic anemia may define a subgroup of patients with SLE who have characteristic serologic and clinical manifestations. PMID- 10723974 TI - Does prior use of aspirin affect outcome in ischemic stroke? AB - BACKGROUND: Large intervention studies suggest that aspirin may reduce mortality when given to patients who present with strokes or transient ischemic attacks. We sought to determine whether patients who were already using aspirin at the time of an ischemic stroke had a lower mortality than those who were not. METHODS: A prospective cohort study was undertaken in patients (mean age 76 +/- 15 years) with acute ischemic stroke. Detailed information on demography, stroke characteristics, and aspirin use prior to the stroke was collected from patients, medical records, and other sources. Patients were classified by cause and subtype of stroke using standard criteria. Mortality was measured 4 weeks after the initial episode. RESULTS: Of the 1,457 patients, 650 (45%) were using aspirin (median dose 75 mg; range 75 to 300 mg) prior to the stroke. Prior use of aspirin was associated with lower 4-week mortality (14% versus 20%, P <0.01 ). Beneficial effects of prior aspirin use on mortality were seen in patients with atherosclerotic strokes (15% versus 21%, P <0.05) and with cardioembolic strokes (21% versus 34%, P <0.05), but not among patients with strokes due to small vessel occlusion (10% versus 11%, P = 0.8). Prior aspirin use was also associated with lower mortality in patients in whom the cause of ischemic stroke could not be determined (15% versus 22%, P <0.01). The effect of prior aspirin use on mortality was independent of age, gender, other risk factors, and use of other medication. CONCLUSIONS: Prior use of low-dose aspirin may be associated with a small but significant reduction in stroke mortality. PMID- 10723976 TI - A systematic review of the effects of physician specialty on the treatment of coronary disease and heart failure in the United States. AB - PURPOSE: To assess the effects of physician specialty on the knowledge, management, and outcomes of patients with coronary disease or heart failure. MATERIALS AND METHODS: We performed a systematic search of MEDLINE from 1980 to 1997, as well as bibliographic references to articles about the effects of physician specialty on the knowledge, treatment, and outcomes of patients with coronary disease or heart failure in the United States. RESULTS: Twenty-four articles met our criteria for inclusion (including eight that involved knowledge or self-reported practices, 14 that described actual practice patterns, and six that measured clinical outcomes). Cardiologists were more knowledgeable than generalist physicians about the optimal evaluation and management of coronary disease but not about the use of angiotensin-converting enzyme (ACE) inhibitors for heart failure. Patients with unstable angina or myocardial infarction were more likely to receive proven medical therapies, and possibly had improved outcomes, if they were treated by cardiologists. The use of lipid-lowering drugs after myocardial infarction was also more common among patients of cardiologists. ACE inhibitor use for heart failure was probably greater, and short-term readmission rates were lower, with cardiology care. CONCLUSIONS: Patients with coronary disease or heart failure in the United States who are treated by cardiologists appear more likely to receive evidence-based care and probably have better outcomes. Investigation of collaborative models of care and innovative efforts to improve the use of proven therapies by physicians are needed. PMID- 10723975 TI - Calcium channel blockers and cancer. AB - PURPOSE: We sought to explore the relation that has been previously reported between calcium channel blockers and an increased risk of cancer. SUBJECTS AND METHODS: We followed 3,511 participants, age 65 years or older, in the Duke Established Populations for Epidemiologic Studies of the Elderly for up to 10 years. Information about use of medications was obtained at baseline and 3 and 6 years later. Information about hospitalization for cancer, or death from cancer, was obtained from Health Care Financing Administration data and death certificates. RESULTS: Of the 133 users of calcium channel blockers, 16 (12%) developed cancer, compared with 548 (16%) of 3,378 nonusers (hazard ratio = 0.9; 95% confidence interval, 0.5 to 1.5). Adjusting for baseline and time-dependent covariates, such as race, diabetes, or blood pressure, for dose or class of calcium channel blockers, or for length of follow-up, had no effect. CONCLUSIONS: Use of calcium channel blockers does not appear to be related to cancer risk. Earlier reports showing such a relation may have been the result of chance. PMID- 10723977 TI - New therapies for alcohol problems: application to primary care. AB - Primary care physicians are frequently involved in the longitudinal care of patients with alcohol problems and in helping patients to decrease their alcohol consumption. Recent clinical trials provide evidence in support of new treatment strategies for these patients. Brief interventions have been used successfully to reduce alcohol consumption in patients with hazardous and harmful drinking. Twelve-step facilitation, cognitive behavioral, and motivational enhancement therapies have produced sustained drinking reductions in patients with alcohol dependence. Pharmacologic therapies, such as naltrexone and acamprosate, have been effective in decreasing alcohol consumption when provided along with psychosocial counseling in patients with alcohol dependence. The current review highlights the application of these new therapies to primary care physicians' efforts on behalf of their patients with alcohol problems. PMID- 10723978 TI - Management of patients with hypertension and diabetes mellitus: advances in the evidence for intensive treatment. AB - Diabetes mellitus is common in patients with hypertension, and greatly increases the risk of cardiovascular disease, including myocardial infarction, stroke, and peripheral vascular disease. "The pharmacologic therapy of patients with hypertension and diabetes has been surrounded by controversy because of concerns about the metabolic effects of several antihypertensive agents, as well as concerns about the safety of calcium antagonists. The objective of this review is to re-evaluate the management of diabetic patients with hypertension in light of the latest clinical trials. We also review the importance of intensive blood pressure control in these patients. PMID- 10723979 TI - Positron emission tomography in giant cell arteritis and polymyalgia rheumatica: evidence for inflammation of the aortic arch. PMID- 10723980 TI - Reactivation of hepatitis B in patients with human immunodeficiency virus infection treated with combination antiretroviral therapy. PMID- 10723981 TI - Intraclot recombinant tissue plasminogen activator in the treatment of deep venous thrombosis of the lower and upper extremities. PMID- 10723982 TI - Diagnostic dilemma. Cardiogenic shock with third degree A-V block and junctional escape rhythm. PMID- 10723983 TI - Primary percutaneous intervention in octogenarians with acute myocardial infarction: the treatment of choice. PMID- 10723984 TI - Generalists and specialists caring for patients with heart disease: united we stand, divided we fall. PMID- 10723985 TI - Case of the month misdiagnosis. PMID- 10723986 TI - Physicians and drug company representatives. PMID- 10723987 TI - Do residents need unions? PMID- 10723988 TI - Commentary. Alliance for Academic Internal Medicine. PMID- 10723989 TI - Unraveling the symmetry ambiguity in a hexamer: calculation of the R6 human insulin structure. AB - Crystallographic and NMR studies of insulin have revealed a highly flexible molecule with a range of different aggregation and structural states; the importance of these states for the function of the hormone is still unclear. To address this question, we have studied the solution structure of the insulin R6 symmetric hexamer using NMR spectroscopy. Structure determination of symmetric oligomers by NMR is complicated due to 'symmetry ambiguity' between intra- and intermonomer NOEs, and between different classes of intermonomer NOEs. Hence, to date, only two symmetric tetramers and one symmetric pentamer (VTB, B subunit of verotoxin) have been solved by NMR: there has been no other symmetric hexamer or higher-order oligomer. Recently, we reported a solution structure for R6 insulin hexamer. However, in that study, a crystal structure was used as a reference to resolve ambiguities caused by the threefold symmetry; the same method was used in solving VTB. Here, we have successfully recalculated R6 insulin using the symmetry-ADR method, a computational strategy in which ambiguities are resolved using the NMR data alone. Thus the obtained structure is a refinement of the previous R6 solution structure. Correlated motions in the final structural ensemble were analysed using a recently developed principal component method; this suggests the presence of two major conformational substates. The study demonstrates that the solution structure of higher-order symmetric oligomers can be determined unambiguously from NMR data alone, using the symmetry-ADR method. This success bodes well for future NMR studies of higher-order symmetric oligomers. The correlated motions observed in the structural ensemble suggest a new insight into the mechanism of phenol exchange and the T6 <--> R6 transition of insulin in solution. PMID- 10723990 TI - Peptide self-association in aqueous trifluoroethanol monitored by pulsed field gradient NMR diffusion measurements. AB - Defining the self-association state of a molecule in solution can be an important step in NMR-based structure determination. This is particularly true of peptides, where there can be a relatively small number of long-range interactions and misinterpretation of an intermolecular NOE as an intramolecular contact can have a dramatic influence on the final calculated structure. In this paper, we have investigated the use of translational self-diffusion coefficient measurements to detect self-association in aqueous trifluoroethanol of three peptides which are analogues of the C-terminal region of human neuropeptide Y. Experimentally measured diffusion coefficients were extrapolated to D0, the limiting value as the peptide concentration approaches zero, and then converted to D(20,w), the diffusion coefficient after correction for temperature and the viscosity of the solvent. A decrease in D(20,w) of about 16% was found for all three peptides in aqueous TFE (30% by volume) compared with water, which is in reasonable agreement with the expected decrease upon dimerisation, the presence of which was indicated by sedimentation equilibrium measurements. Apparent molecular masses of these peptides in both solutions were also calculated from their diffusion coefficients and similar results were obtained. Several potential internal standards, including acetone, acetonitrile, dimethylsulfoxide and dioxane, were assessed as monitors of solution viscosity over a range of trifluoroethanol concentrations. Compared with independent measurements of viscosity, acetonitrile was the most accurate standard among these four. The practical limitations of a quantitative assessment of peptide self-association from translational diffusion coefficients measured by PFGNMR, including the calculation of apparent molecular mass, are also discussed. PMID- 10723991 TI - Measurement of dipolar couplings in a transducin peptide fragment weakly bound to oriented photo-activated rhodopsin. AB - Rhodopsin-containing disks, isolated from rod outer segments of bovine retina, align at high magnetic fields with their membrane normal parallel to the magnetic field. After light-activation of rhodopsin, transient binding of the C-terminal transducin undecapeptide, selectively labeled with 15N at Leu5 and Gly9, results in residual dipolar contributions to the 1J(NH) splittings for these two residues. Both residues show 1J(NH) splittings which are smaller than in the dark adapted or rhodopsin-free sample, and return to their isotropic values at a rate determined by the decay of the meta II state of rhodopsin. The dipolar couplings indicate that in the bound state, N-H vectors of Leu5 and Gly9 make angles of 48+/-4 degrees and 40+/-8 degrees, respectively, with the disk normal. These 'transferred' dipolar couplings potentially offer a useful method for studying the conformation and orientation of flexible, low affinity ligands when bound to oriented integral membrane receptors. PMID- 10723992 TI - NMR experiments for resonance assignments of 13C, 15N doubly-labeled flexible polypeptides: application to the human prion protein hPrP(23-230). AB - A combination of three heteronuclear three-dimensional NMR experiments tailored for sequential resonance assignments in uniformly 15N, 13C-labeled flexible polypeptide chains is described. The 3D (H)N(CO-TOCSY)NH, 3D (H)CA(CO-TOCSY)NH and 3D (H)CBCA(CO-TOCSY)NH schemes make use of the favorable 15N chemical shift dispersion in unfolded polypeptides, exploit the slow transverse 15N relaxation rates of unfolded polypeptides in high resolution constant-time [1H, 15N] correlation experiments, and use carbonyl carbon homonuclear isotropic mixing to transfer magnetization sequentially along the amino acid sequence. Practical applications are demonstrated with the 100-residue flexible tail of the recombinant human prion protein, making use of spectral resolution up to 0.6 Hz in the 15N dimension, simultaneous correlation with the two adjacent amino acid residues to overcome problems associated with spectral overlap, and the potential of the presently described experiments to establish nearest-neighbor correlations across proline residues in the amino acid sequence. PMID- 10723993 TI - Automation of NMR measurements and data evaluation for systematically screening interactions of small molecules with target proteins. AB - In this technical note we describe the setup and application of automated sample preparation and usage of flow-through NMR equipment for the characterization of ligand binding on proteins. In addition, we focus on the perspectives of automated analysis of 2D HSQC spectra to identify changes in patterns indicative for ligand binding or changes of sample conditions. In this context we discuss a combination of statistical and non-statistical data analysis. PMID- 10723994 TI - Determination of the populations and structures of multiple conformers in an ensemble from NMR data: multiple-copy refinement of nucleic acid structures using floating weights. AB - A new algorithm is presented for determination of structural conformers and their populations based on NMR data. Restrained Metropolis Monte Carlo simulations or restrained energy minimizations are performed for several copies of a molecule simultaneously. The calculations are restrained with dipolar relaxation rates derived from measured NOE intensities via complete relaxation matrix analysis. The novel feature of the algorithm is that the weights of individual conformers are determined in every refinement step, by the quadratic programming algorithm, in such a way that the restraint energy is minimized. Its design ensures that the calculated populations of the individual conformers are based only on experimental restraints. Presence of internally inconsistent restraints is the driving force for determination of distinct multiple conformers. The method is applied to various simulated test systems. Conformational calculations on nucleic acids are carried out using generalized helical parameters with the program DNAminiCarlo. From different mixtures of A- and B-DNA, minor fractions as low as 10% could be determined with restrained energy minimization. For B-DNA with three local conformers (C2'-endo, O4'-exo, C3'-endo), the minor O4'-exo conformer could not be reliably determined using NOE data typically measured for DNA. The other two conformers, C2'-endo and C3'-endo, could be reproduced by Metropolis Monte Carlo simulated annealing. The behavior of the algorithm in various situations is analyzed, and a number of refinement protocols are discussed. Prior to application of this algorithm to each experimental system, it is suggested that the presence of internal inconsistencies in experimental data be ascertained. In addition, because the performance of the algorithm depends on the type of conformers involved and experimental data available, it is advisable to carry out test calculations with simulated data modeling each experimental system studied. PMID- 10723995 TI - Resolution of the 1H-1H NOE spectrum of RNA into three dimensions using 15N-1H two-bond couplings. AB - The feasibility of using two-bond 15N-1H couplings to resolve the 1H-1H nuclear Overhauser effect spectrum of RNA into a third dimension was investigated, using the 36-nucleotide gene 32 messenger RNA pseudoknot of bacteriophage T2 as an example. The two-bond 15N-1H couplings present in adenosine and guanosine were found to be suitable for generating a three-dimensional 1H-1H-15N NOESY-HSQC spectrum with reasonably good sensitivity, as well as favorable chemical shift dispersion in the nitrogen dimension. The described NMR experiment provides a tool that can be used to complement other heteronuclear methods in the analysis of RNA structure. PMID- 10723996 TI - A novel experiment for the quantitative measurement of CSA(1H(N))/CSA(15N) cross correlated relaxation in 15N-labeled proteins. AB - An experiment is presented which allows for the quantitative measurement of the relaxation interference between the 1H(N) CSA and 15N CSA interactions in 15N labeled proteins. A constant-time buildup scheme is used to measure the differential relaxation rate, eta, between double-quantum (DQ) and zero-quantum (ZQ) 1H(N)-15N coherences. The CSA/CSA experiment was recorded at three different Bo field strengths. The CSA(1H(N))/CSA(15N) cross-correlation rate was obtained from the linear fit of the measured rate, eta, versus Bo2 for 77 residues of the EH2 domain from mouse Eps15. PMID- 10723997 TI - Long-range imino proton-13C J-couplings and the through-bond correlation of imino and non-exchangeable protons in unlabeled DNA. AB - Thanks to rather large (5-9 Hz) long-range imino proton-13C J-couplings, heteronuclear correlation experiments in H2O provide unambiguous assignment of imino protons by intranucleotide through-bond connectivities to guanosine H8 and thymidine CH3 protons, or sequence-specific assignment of non-exchangeable protons when the imino protons are identified independently. This method is demonstrated in the Dickerson dodecamer [d(CGCGAATTCGCG)]2 and in a human telomeric fragment of 22 nucleotides. PMID- 10723998 TI - Sequence-specific 1H and 15N assignment and secondary structure of transforming growth factor beta3. PMID- 10723999 TI - Sequence-specific resonance assignments and partial unfolding of extracellular domains II and III of E-cadherin. PMID- 10724000 TI - Cerebral aneurysms and medical defects, continued. PMID- 10724001 TI - Novel streptopyrroles from Streptomyces rimosus with bacterial protein histidine kinase inhibitory and antimicrobial activities. AB - A series of halogenated pyrrolo [2,1-b] [1,3] benzoxazines (1 approximately 9) was isolated from fermentations of an actinomycete strain X10/78/978 (NCIMB40808), identified as Streptomyces rimosus, during a microbial extract screening programme to identify inhibitors of bacterial histidine kinase. The structures of these compounds were elucidated by spectroscopic methods including the HMQC, HMBC and INADEQUATE NMR experiments. The structure of 1 was confirmed by X-ray crystallographic studies. Compounds 5 and 6 were produced in fermentations in the presence of NaBr and NaI respectively. The most abundant member of the series, streptopyrrole, 1, inhibited the nitrogen regulator II (NRII) histidine kinase from Escherichia coli with an IC50 of 20 microM and exhibited antimicrobial activity against a range of bacteria and fungi. PMID- 10724002 TI - Adxanthromycins A and B, new inhibitors of ICAM-1/LFA-1 mediated cell adhesion molecule from Streptomyces sp. NA-148. I. Taxonomy, production, isolation and biological activities. AB - Two new inhibitors of ICAM-1/LFA-1 mediated cell adhesion molecule, adxanthromycins A and B were isolated from the cultured broth of a streptomycete strain. The strain was identified as Streptomyces sp. NA-148 from its morphological and physiological characteristics. Adxanthromycins A and B inhibited the formation of the JY cell aggregates from 1.5 microg/ml, respectively, in a dose dependent manner. Components A and B also inhibited a human T cell leukemia cell line SKW-3 adhesion to soluble ICAM-1 in a dose dependent manner with an IC50 of 18.8 microg/ml and 25.0 microg/ml, respectively. PMID- 10724003 TI - Effect of fungal metabolites cytochalasans on lipid droplet formation in mouse macrophages. AB - Effects of seven cytochalasans including cytochalasins B, D and E and novel phenochalasins A and B were tested on cytosolic lipid droplet formation and neutral lipid synthesis in mouse peritoneal macrophages. Phenochalasin A inhibited lipid droplet formation in a dose-dependent manner at least up to 20 microM without any morphological changes in macrophages. Cytochalasins D and E also inhibited lipid droplet formation only in a narrow range of concentrations, around 1 and 0.1 microM, respectively. At higher concentrations they gave morphological changes in macrophages. The other four cytochalasans only showed severe morphological changes in macrophages. Phenochalasin A and cytochalasins D and E inhibited cholesteryl ester (CE) synthesis specifically with IC50 values of 0.61, 2.4 and 0.20 microM, respectively, while the other cytochalasans inhibited both CE and triacylglycerol syntheses. Thus, among the cytochalasans tested, phenochalasin A showed very specific inhibition of CE synthesis and gave the lowest morphological changes in macrophages, resulting in the best inhibitor of lipid droplet formation in macrophages. PMID- 10724004 TI - New cytotoxic agents, BE-54238A and B, produced by a streptomycete. AB - New cytotoxic substances, designated BE-54238A and B, were isolated from the culture broth of Streptomyces sp. A54238. The active principles were extracted from the mycelium by methanol and purified by Diaion HP-20 and Sephadex LH-20 column chromatographies. BE-54238A and B exhibited cytotoxic activity against murine and human tumor cell lines. PMID- 10724005 TI - PF1163A and B, new antifungal antibiotics produced by Penicillium sp. I. Taxonomy of producing strain, fermentation, isolation and biological activities. AB - Two novel antifungal antibiotics, PF1163A and B, were isolated from the fermentation broth of Penicillium sp. They were purified from the solid cultures of rice media using ethyl acetate extraction, silica gel and Sephadex LH-20 column chromatographies. PF1163A and B showed potent growth inhibitory activity against pathogenic fungal strain Candida albicans but did not show cytotoxic activity against mammalian cells. These compounds inhibited the ergosterol biosynthesis in Candida albicans. PMID- 10724006 TI - PF1163A and B, new antifungal antibiotics produced by Penicillium sp. II. Physico chemical properties and structure elucidation. AB - The structures of new antifungal antibiotics, PF1163A and B, were elucidated by spectroscopic analyses of the degradation products and by X-ray crystallography of the de-2-hydroxyethyl derivative of PF1163B. Both antibiotics consist of a 13 membered macrocyclic structure containing a derivative of N-methyl tyrosine and a hydroxy fatty acid. PF1163A differs from PF 1163B by having an additional hydroxyl group on the side chain. PMID- 10724007 TI - Haematocin, a new antifungal diketopiperazine produced by Nectria haematococca Berk. et Br. (880701a-1) causing nectria blight disease on ornamental plants. AB - A new antifungal diketopiperazine named haematocin was isolated from the culture broth of Nectria haematococca Berk. et Br. (880701a-1) causing blight disease on ornamental plants, Phalaenopsis spp. and Doritanopsis spp. Its structure was established by spectroscopic methods. Haematocin inhibited the germ-tube elongation and spore-germination of Pyricularia oryzae at the ED50 values of 30 microg/ml and 160 microg/ml, respectively. PMID- 10724008 TI - Arisugacins C and D, novel acetylcholinesterase inhibitors and their related novel metabolites produced by Penicilium sp. FO-4259-11. AB - The mutant of Penicillium sp. FO-4259, an arisugacins A and B producing strain, was found to produce a series of metabolites, designated arisugacins C, D, E, F, G and H, which were structurally related to arisugacins A and B. These compounds were isolated from the culture broth and the physico-chemical and biological properties were examined. The IC50 values of arisugacins C and D against acetylcholinesterase (AChE) were 2.5 microM and 3.5 microM, respectively. However arisugacins E, F, G and H did not inhibit AChE at 100 microM. Though they showed only weak or no activity against AChE compared with arisugacins A and B, they may be useful for the study of the structure-activity relationship. PMID- 10724009 TI - Effective production of dehydro cyclic dipeptide albonoursin exhibiting pronuclear fusion inhibitory activity. II. Biosynthetic and bioconversion studies. AB - Albonoursin production was greatly enhanced when cyclo (L-Leu-L-Phe) (CFL), a tetrahydro derivative of albonoursin, was added to the 2-day culture of an albonoursin-producing actinomycete, Streptomyces albulus KO-23. The increase in albonoursin production paralleled the amount of CFL added. Furthermore, the resting cells of the strain catalyzed the bioconversion of CFL to albonoursin. The optimum pH and temperature for the conversion were found to be pH 10.0 and 50 degrees C. The feeding experiments and the resting-cell reactions revealed that albonoursin is biosynthesized by dehydrogenation of CFL in the actinomycete. This is the first report for a dehydrogenation of amino acid residues at the alpha,beta-positions in cyclic dipeptides. PMID- 10724010 TI - TMC-86A, B and TMC-96, new proteasome inhibitors from Streptomyces sp. TC 1084 and Saccharothrix sp. TC 1094. II. Physico-chemical properties and structure determination. PMID- 10724011 TI - Pyrronamycin A and B, novel antitumor antibiotics containing pyrrole-amide repeating unit, produced by Streptomyces sp. PMID- 10724012 TI - A new antifungal macrolide component, brasilinolide B, produced by Nocardia brasiliensis. PMID- 10724013 TI - Antitumor activity of brasilicardin A, a novel terpenoid antibiotic from Nocardia brasiliensis. PMID- 10724014 TI - Biosynthesis of dykellic acid: origin of the carbon skeleton. PMID- 10724015 TI - Belactosin A, a novel antitumor antibiotic acting on cyclin/CDK mediated cell cycle regulation, produced by Streptomyces sp. PMID- 10724016 TI - A new actinomycin-type chromopeptide from Streptomyces sp. HKI-0155. PMID- 10724017 TI - The first total synthesis of pyralomicin 1c. PMID- 10724018 TI - Molecular epidemiology of antibiotic resistance. AB - Molecular typing methods based on the analysis of the genetic structure of bacteria, are used to address many different problems such as the study of genomic organisation and evolution, the identification of patterns of infection, the identification of sources of transmission, the epidemiological surveillance of infectious diseases and for investigations into outbreaks. Of particular interest is the application of these techniques for acquiring information on the spread of micro-organisms that have become resistant to many clinically important antibiotics. The emergence of antibiotic resistance is one of the most dangerous phenomena of the last 20 years and knowledge of the mechanisms of resistant-gene exchange means fully understanding their spread into all environments. Studies on the molecular epidemiology of antibiotic-resistance in micro-organisms should make it easier to distinguish clonality with respect to horizontal transfer of the determinants of resistance. PMID- 10724019 TI - Targeted delivery of antibiotics using liposomes and nanoparticles: research and applications. AB - This review examines current technologies for increasing the bioavailability of antibiotics by means of liposomes or nanoparticles. The main focus is on liposomes. These carriers were preferentially developed because their composition is compatible with biological constituents. Biodegradable polymers in the form of colloidal particles have also been used and show promise for future applications in antimicrobial chemotherapy. The in vivo behaviour of both types of carriers and consequently their therapeutic potential, are determined by their route of administration. Conventional carrier strategies permit the mononuclear phagocyte system to be targeted by intravenous injection of antibiotics. Stealthy strategies avoid major uptake by these cells and extend the systemic presence of these carriers. The purpose of this review is to provide background information in antibiotic targeting gathered from papers published over the last twenty years. It seems clear that such drug carriers (liposomes, nanoparticles) allow increased drug concentration at infected sites but reduce drug toxicity. PMID- 10724020 TI - Direct confocal microscopy studies of the bacterial colonization in vitro of a silver-coated heart valve sewing cuff. AB - The antimicrobial coating of prosthetic heart valve sewing cuffs has been considered a potentially effective method for preventing prosthetic valve endocarditis. Although traditional in vitro bacterial adherence studies are often useful as screening tools, they can be inadequate in examining the antiinfective efficacy of antimicrobial-coated devices. We conducted a pilot in vitro study to directly assess the antimicrobial activity of a silver-coated sewing cuff versus uncoated cuff using confocal scanning laser microscopy. Staphylococcus epidermidis adhered more to the surfaces of the silver-coated sewing cuff compared with the uncoated cuff. These pilot in vitro results cast a doubt on the antiinfective efficacy of silver-coated prosthetic heart valve sewing cuffs and suggest further assessment should be carried out using animal studies. PMID- 10724021 TI - Outer-membrane proteins pattern and detection of beta-lactamases in clinical isolates of imipenem-resistant Acinetobacter baumannii from Brazil. AB - In order to compare imipenem-sensitive and -resistant Acinetobacter baumannii strains isolated from three patients, ribotyping, plasmid, beta-lactamase detection and outer-membrane analysis were performed. Ribotyping and the use of a beta-lactam during the period when the strains were isolated suggested that they had a common origin and that resistance occurred in vivo. Outer membrane analysis showed no difference between susceptible and resistant strains with the exception of an A2 imipenem-resistant strain that lost a protein band of 31-36 kDa. Beta lactamases were detected using isoelectric focusing in all strains (pI of 7.4). In addition, two beta-lactamases (pI of 5.9 and 6.7) were found in imipenem resistant isolates. The double-disc technique demonstrated the presence of a beta lactamase capable of imipenem inactivation in resistant strains. Plasmid analysis showed that all susceptible strains had the same pattern, one resistant strain did not have any plasmid, one had the same plasmid pattern of its susceptible pair and only one had a different pattern when compared with its susceptible pair. PMID- 10724022 TI - Comparison of pivmecillinam and cephalexin in acute uncomplicated urinary tract infection. AB - The clinical and bacteriological efficacy of a 3-day course of pivmecillinam, 200 mg three times daily, was compared with that of a 7-day course of cephalexin, 250 mg four times daily, in 216 patients with a bacteriologically confirmed, acute, uncomplicated, urinary tract infection. Both treatments were similarly effective. Clinical cure or improvement was obtained in 95.3% of patients given pivmecillinam and in 93.6% of patients given cephalexin. Bacteriological success was achieved in 89.7 and 81.7% patients taking pivmecillinam or cephalexin, respectively. Eradication rates for Escherichia coli were 90.1% for pivmecillinam and 80.6% for cephalexin. Both treatments were well tolerated. This study has confirmed that a 3-day course of pivmecillinam is effective and well tolerated in uncomplicated cystitis. PMID- 10724023 TI - Efficacy of the bisbenzylisoquinoline alkaloids in acute and chronic Trypanosoma cruzi murine model. AB - We have shown previously that daphnoline and cepharanthine are active against Trypanosoma cruzi and inhibited trypanothione reductase. The effects of oral treatments with daphnoline, cepharanthine and benznidazole were examined in Balb/c mice infected with T. cruzi acutely and chronically. In acute infections, parasitaemia was significantly reduced in the daphnoline-treated mice compared with controls and benznidazole-treated mice. The parasitological cure rate was increased in mice treated with daphnoline. Fifty days after infection, the negative serological response in both models was significantly different for the three tested drugs. Daphnoline showed the highest negative serological rate (48%). In chronically infected mice treated with daphnoline, we were unable to detect parasites in 70% of mice. The results obtained of oral treatment of daphnoline suggest that this bisbenzylisoquinoline may be useful in the treatment of acute and chronic Chagas' disease. This was not seen with cepharanthine, an excellent trypanothione reductase inhibitor. PMID- 10724024 TI - Antimicrobial and antioxidant activities of Feijoa sellowiana fruit. AB - The present study was designed to evaluate the antibacterial and antioxidant activities of an aqueous extract from the tropical Feijoa sellowiana Berg. fruit which is widely used for human food. The extract was tested against gram-positive and gram-negative bacteria by a broth dilution test and on human whole blood leukocytes, as well as isolated neutrophils using a chemiluminescence (CL) assay. The extract inhibited bacterial growth; Pseudomonas aeruginosa, Enterobacter aerogenes and Enterobacter cloacae were the most sensitive. The fruit extract significantly decreased CL emission from human whole blood phagocytes and isolated polymorphonuclear leukocytes whether they were activated or not by soluble or phagocytic stimuli. F. sellowiana showed both antibacterial and antioxidant properties and therefore its extract might be used as a new multifaceted drug. PMID- 10724026 TI - Susceptibility pattern of Staphylococcus aureus isolated in Malaysian hospitals. AB - Isolates of 390 Staphylococcus aureus were tested against 13 different antibiotics by a disc diffusion method as recommended by the National Committee for Clinical Laboratory Standards (NCCLS). Strains were isolated from blood (5.7%), cerebrospinal fluid (0.5%), respiratory tract (11.8%), pus and wound (73.3%), urine (1.8%), genital specimens (1.0%) and other specimens (4.3%). Only 4.6% of the isolates were fully susceptible to all the drugs tested. Resistance to penicillin was 94.1%, methicillin, 39.7%, chloramphenicol, 8.5%, ciprofloxacin, 29.2%, clindamycin, 2.1%, erythromycin, 45.9% gentamicin, 40.5%; rifampicin, 3.3% tetracycline, 47.2%, co-trimoxazole, 38.5%, mupirocin, 2.8%, fusidic acid, 3.6%. None of the isolates was resistant to vancomycin. The susceptibility of methicillin-resistant strains to erythromycin, gentamicin, tetracycline and ciprofloxacin was low, while clindamycin, fusidic acid, mupirocin, and rifampicin remained active. PMID- 10724025 TI - Antimicrobial activity of synthetic all-D mastoparan M. AB - Mastoparan M, a tetradecapeptide toxin (INKAIAALAKKLL-NH2) from hornet venom and its D-form mastoparan M were synthesized chemically. All D- and L-mastoparan M forms were found to adopt 28% alpha-helical structures in a 30% trifluroethanol solution as shown by the circular dichroism spectrum. All-D mastoparan M caused 3H-thymidine release from labeled bacterial cells after incubation for 1 h and complete cell lysis by 4 h. Both L- and D-mastoparan M showed strong activity against gram-positive and gram-negative bacteria. All-D mastoparan M showed 2 fold higher antibacterial activity than L-mastoparan M. The effects of all-D mastoparan M on the surface morphology of Staphylococcus aureus (ATCC29213) and Escherichia coli (ATCC25922) were studied by scanning-beam electron microscopy. Blast-like bleb extrusions on the surface of some S. aureus and swellings on the end of E. coli were seen after culture with all-D mastoparan M. These findings indicated the all-D mastoparan M could kill bacteria by disrupting cells. PMID- 10724027 TI - Sulphur-containing derivatives structurally related to fenoxycarb are potent growth inhibitors against the intracellular form of Trypanosoma cruzi. AB - Sulphur-containing derivatives structurally related to the insect growth regulator fenoxycarb were shown to be extremely active antiproliferative agents against the amastigote form of Trypanosoma cruzi in in vitro assays. All of these drugs had previously been proved to be remarkably potent growth inhibitors against the epimastigote form of the parasite. PMID- 10724028 TI - Therapeutic effect of clarithromycin for respiratory-tract infections in children caused by Chlamydia pneumoniae. Research Group of Sapporo for Pediatric Chlamydial Infections. AB - Children infected with Chlamydia pneumoniae sometimes experience lower respiratory tract infections such as pneumonia and bronchitis. Although numerous anti-microbial compounds have been reported to be active against the organism, most of them have not been in a clinical trial in infants and children with C. pneumoniae infection. Clarithromycin has been shown to express anti-chlamydial effects in vitro. In this study, we evaluated the clinical anti-C. pneumoniae properties of clarithromycin in children with mainly lower respiratory tract infection. We administered clarithromycin orally to 21 infants and children at a dose of 10-15 mg/kg/day divided into two or three doses for 4-21 days. Clinical symptoms, roentgenographic and laboratory abnormal findings improved. The overall clinical efficacy rate was 85.7% (18 of 21 cases). Administration of clarithromycin was considered to be a suitable treatment for improving lower respiratory infections in infants and children caused by C. pneumoniae. PMID- 10724029 TI - Bactericidal activity of levofloxacin and ciprofloxacin on clinical isolates of different phenotypes of Pseudomonas aeruginosa. AB - Levofloxacin has been reported to have in vitro activity against both gram positive and gram-negative bacteria. A recent survey carried out at our Institution showed clinical isolates of Pseudomonas aeruginosa to be more susceptible to levofloxacin than to ciprofloxacin. The in vitro activity of the two fluoroquinolones was evaluated further by looking at their bactericidal activity against two strains of each of the following antibio-phenotypes of P. aeruginosa: levofloxacin- and ciprofloxacin-susceptible, levofloxacin susceptible/ciprofloxacin-resistant, levofloxacin-susceptible/ciprofloxacin susceptible and ceftazidime-resistant, (National Committee for Clinical Laboratory Standards susceptibility breakpoints were used). MIC and MBC values were measured and time-kill experiments were carried out. Drugs were used at susceptibility or resistance breakpoint concentrations in the time-kill experiments and results were recorded over 12 h in an attempt to link in vitro results with the clinical situation The polypeptide profiles of outer membrane preparations of the six strains were examined by gel electrophoresis. Levofloxacin was shown to be more bactericidal than ciprofloxacin in the time kill experiments. No differences were observed between the outer membrane proteins of the six strains. Levofloxacin showed greater bactericidal activity against P. aeruginosa clinical isolates than ciprofloxacin. PMID- 10724030 TI - Recombinant adeno-associated virus-mediated correction of lysosomal storage within the central nervous system of the adult mucopolysaccharidosis type VII mouse. AB - The central nervous system (CNS) is a predominant site of involvement in several lysosomal storage diseases (LSDs); and for many patients, these diseases are diagnosed only after the onset of symptoms related to the progressive accumulation of macromolecules within lysosomes. The mucopolysaccharidosis type VII (MPS VII) mice are deficient for the lysosomal enzyme beta-glucuronidase and, by early adulthood, develop a significant degree of glycosaminoglycan storage within neuronal, glial, and leptomeningeal cells. Using this animal model, we investigated whether gene transfer mediated by a recombinant adeno-associated virus (rAAV) vector is capable of reversing the progression of storage lesions within the CNS. Adult MPS VII mice received intracerebral injections of 4 X 10(7) infectious units of a rAAV vector carrying the murine beta-glucuronidase (gus s(a)) cDNA under the transcriptional direction of the cytomegalovirus immediate early promoter and enhancer. By 1 month after vector administration, transgene derived beta-glucuronidase was present surrounding the injection site. Enzyme levels were between 50 and 240% of that found in wild-type mice. This level of beta-glucuronidase activity was sufficient to reduce the degree of lysosomal storage. Moreover, the reduction in storage was maintained for at least 3 months post-rAAV administration. These data demonstrate that rAAV vectors can transduce the diseased CNS of MPS VII mice and mediate levels of transgene expression necessary for a therapeutic response. Thus, rAAV vectors are potential tools in the treatment of the mucopolysaccharidoses and other lysosomal storage diseases. PMID- 10724031 TI - The use of adeno-associated virus to circumvent the maturation-dependent viral transduction of muscle fibers. AB - Muscle-based gene therapy using adenovirus, retrovirus, and herpes simplex virus has been hindered by viral cytotoxicity, host immune response, and the maturation dependent viral transduction of muscle fibers. The development of new mutant vectors has greatly reduced the toxicity and the immune rejection problems, but the inability of viral vectors to penetrate and transduce mature myofibers remains an important issue. Research has been focused on the characterization of barriers to viral transduction in mature myofibers to develop strategies to circumvent the maturation-dependent viral transduction of myofibers. Here, we report that adeno-associated virus (AAV) can be used to overcome the maturation dependent viral transduction of myofibers. We have investigated by which mechanism AAV can penetrate and efficiently transduce mature muscle fibers, and have shown that this viral vector is not blocked by the basal lamina and that AAV transduction of myofibers is independent of myoblast mediation. Although AAV can efficiently transduce mature myofibers, a differential transduction is still observed among the different types of myofibers that correlates with the expression of the heparan sulfate proteoglycan receptors, the muscle maturity, the number of viral particles used, and the time postinjection. The identification of the mechanisms by which AAV transduces mature myofibers will help in the development of strategies to achieve an efficient muscle-based gene therapy for inherited and acquired diseases. PMID- 10724032 TI - Packageable antiviral therapeutics against human immunodeficiency virus type 1: virion-targeted virus inactivation by incorporation of a single-chain antibody against viral integrase into progeny virions. AB - To determine their activities as an antiviral agent packageable within virions and suitable for continued expression in cells, we tested a single-chain antibody (scAb) against human immunodeficiency virus type 1 (HIV-1) integrase and its three fusion proteins: fused to viral protein R (scab-Vpr), a double-cassette of the WXXF motif binding to Vpr (scAb-WXXF), and viral major capsid protein (scAb CA), respectively. Cotransfection of human 293T cells with expression plasmid for scAb-Vpr or -WXXF along with HIV-1 clone pLAI resulted in the production of a normal amount of progeny virions with infectivity decreased by more than 10(3) fold. Immunoblot analyses showed that scAb-Vpr or -WXXF was associated with virions, whereas scAb or scAb-CA was not, suggesting that scAb-Vpr or -WXXF was incorporated into virions. The incorporation of scAb-WXXF appeared to be Vpr dependent, because the fusion protein was associated with the wild-type but not with Vpr-truncated HIV-1 virions. Since G418-selected HeLa clones carrying expression plasmid for scAb-WXXF were obtained much more frequently than those for scAb-Vpr, scAb-WXXF was inferred to be less toxic to cells than scAb-Vpr. These results suggest that scAb-WXXF may serve as a novel class of antiviral therapeutic that inactivates progeny HIV virions from within. PMID- 10724033 TI - Intravenous injection of naked DNA encoding secreted flt3 ligand dramatically increases the number of dendritic cells and natural killer cells in vivo. AB - The trace number of dendritic cells (DCs) present in tissues has limited the study of DC biology and development of clinical applications utilizing DCs. Here we show that hydrodynamics-based gene delivery of naked DNA encoding secreted human flt3 ligand (hFLex) can dramatically increase the number of functional DCs and natural killer (NK) cells. After a single injection of the hFLex gene, hFLex levels in mouse serum reached approximately 40 microg/ml and remained above 1 microg/ml for 5-6 days. Sustained levels of serum hFLex correlated with significant increases in the size of the lymphoid organs and in the proportion of dendritic cells and NK cells in both lymph nodes and spleen. The increase in DC and NK cell numbers started from day 5, and reached peak levels between day 8 and day 12. The levels then returned to normal on day 20. These DCs and NK cells were functional as evidenced by mixed leukocyte reactions and lysis of YAC-1 cells, respectively. These results suggest that delivery of the hFLex gene provides a simple, efficient, and inexpensive way of increasing DC and NK cell populations in vivo, and may have broad applications in the further study of DC and NK cell biology and in the development of immunotherapy strategies. PMID- 10724034 TI - Chemoprotection of hematopoietic cells by a mutant P-glycoprotein resistant to a potent chemosensitizer of multidrug-resistant cancers. AB - Cancers are frequently chemoresistant because of overexpression of P glycoprotein. Two different approaches to improve cancer treatment are currently being investigated in clinical trials: inhibition of P-glycoprotein function by reversing agents, and alleviation of leukocytopenia by MDR1 gene transfer to normal bone marrow of patients. We report here that retroviral vectors encoding a mutant P-glycoprotein (MDR1-F983A) protect hematopoietic cells from anticancer drugs even in the presence of trans-(E)-flupentixol, an inhibitor of P glycoprotein. Transfer of either mutant or wild-type MDR1 to K562 erythroleukemia cells or primary murine bone marrow resulted in reduced accumulation of daunomycin and vinblastine because of increased drug efflux.trans-(E)-Flupentixol at concentrations up to 10 microM failed to reverse drug efflux mediated by the product of the mutant MDR1 while wild-type P-glycoprotein was inhibited. In the presence of 2 microM trans-(E)-flupentixol chemoresistance to daunomycin was circumvented only in K562 cells transduced with wild-type, but not with mutant, MDR1. Moreover, drug resistance of KB-8-5 epidermoid cancer cells, which express the wild-type MDR1 gene at levels comparable to clinical specimens from multidrug resistant cancers, was fully overcome in the presence of trans-(E)-flupentixol. Vectors expressing mutant P-glycoprotein may help improve chemotherapy by allowing safe dose intensification under conditions in which multidrug-resistant cancers are rendered drug sensitive by reversing agents. PMID- 10724035 TI - Partial correction of the urinary concentrating defect in aquaporin-1 null mice by adenovirus-mediated gene delivery. AB - The feasibility of water channel gene delivery to kidney tubules and microvessels was evaluated by delivery of an adenovirus encoding aquaporin 1 (AQP1-Ad5) to transgenic AQP1 null mice. In wild-type mice, AQP1 is expressed in kidney proximal tubule, thin descending limb of Henle, and descending vasa recta, where urine osmolality (Uosm) increases from 1000-1500 mOsm (before) to 2500-3500 mOsm after 36 hr of water deprivation. Uosm in AQP1 null mice remains nearly fixed at 650-750 mOsm. AQP1-Ad5 (with a CMV promoter) was generated and purified. Infection of CHO cells gave strong uniform AQP1 expression with plasma membrane localization and eightfold increased water permeability over noninfected cells. AQP1-Ad5 was delivered to 20 to 25-g AQP1 null mice by tail vein infusion (0 10(10) PFU). At 3-7 days, AQP1 protein expression was strongest in liver (approximately 20 microg of AQP1 protein per liver) and next strongest in kidney, with expression in proximal tubule apical and basolateral membranes, and renal microvessels. Functional analysis showed increased water permeability in apical membrane vesicles from proximal tubule. AQP1 expression was not detected in glomerulus, limb of Henle, or collecting duct. In water-deprived null mice receiving 5 x 10(9) PFU of AQP1-Ad5, Uosm increased by up to 510 mOsm (mean increase, 225 +/- 24 mOsm; n = 33 mice). Whereas the control null mice became lethargic and lost 34.2 +/- 0.6% body weight, the virus-treated mice remained relatively active and lost 32.3 +/- 0.7% body weight. Viral DNA and AQP1 transcript were detected in kidney and liver of null mice up to 17 weeks after virus infusion; partial correction of the urinary concentrating defect persisted for 3-5 weeks. These results demonstrate partial functional correction of a urinary concentrating defect by adenoviral delivery of the AQP1 gene. PMID- 10724036 TI - Control of the functional activity of an antisense RNA by a tetracycline responsive derivative of the human U6 snRNA promoter. AB - In an effort to develop a regulatable derivative of the promoter of the human gene for U6 snRNA, we generated several constructs composed of the human U6 snRNA promoter and sequences derived from the gene for the tetracycline operator of a prokaryotic tetracycline resistance transposon. One of the constructs had strong transcriptional activity in the presence of tetracycline that was equivalent to 80% of the activity of the wild-type promoter. Furthermore, the transcriptional activity was almost completely repressed in the absence of tetracycline. Transcriptional activity became detectable within 4 hr after the addition of tetracycline to the culture medium. We used this system to control the functional activity of an antisense RNA for a chimeric gene derived from genes for the epidermal growth factor receptor (EGFR) and green fluorescent protein (GFP). A plasmid that expressed the chimeric gene and a plasmid that expressed the antisense RNA under the control of the inducible U6 promoter were used to cotransfect HeLa cells that were producing the tetracycline repressor protein (Tet R). Addition of tetracycline to the culture medium 12 hr after transfection resulted in the almost complete disappearance of the fluorescent signal due to the chimeric protein within 24 hr. Our results suggest that this expression system might be a useful tool for controlling the expression of functional RNAs, such as aptamers and antisense RNAs, both in basic research and in gene therapy. PMID- 10724037 TI - Effect of changes in expression of the amphotropic retroviral receptor PiT-2 on transduction efficiency and viral titer: implications for gene therapy. AB - The aim of this study was to quantify the impact of amphotropic retroviral receptor (PiT-2) levels on susceptibility to transduction and to determine whether the low level of PiT-2 found on CD34+ hematopoietic cells is within the range likely to compromise gene transfer. Receptor-deficient Chinese hamster ovary (CHO) cells were transfected with a PiT-2 construct that could be induced by the removal of tetracycline. The level of PiT-2 expression measured by virus binding in uninduced and in fully and partially induced transfectants correlated with the efficiency of transduction by an amphotropic retroviral reporter vector. Fully induced CHO-PiT-2 cells gave apparent viral titers similar to NIH 3T3 fibroblasts while addition of tetracycline reduced titers by up to 140-fold. Binding of the same vector preparation to purified CD34+ peripheral blood stem cells (PBSCs) was always less than to uninduced CHO-PiT-2 transfectants even after preincubation in 10-ng/ml concentrations of IL-3, IL-6, and stem cell factor, which increased retroviral binding by an average of 35%. The level of expression of the amphotropic retroviral receptor PiT-2 thus significantly limits transduction efficiency within the range observed in target cells of importance in human gene therapy. PMID- 10724038 TI - Highly efficient transduction and expression of cytokine genes in human tumor cells by means of autonomous parvovirus vectors; generation of antitumor responses in recipient mice. AB - The possible use of recombinant autonomous parvoviruses as vectors to efficiently express therapeutic cytokines in human tumor cells was evaluated in vitro and in vivo. The parvovirus H1 was used to generate recombinant viruses (rH1) that carried transgenes encoding either human interleukin 2 (IL-2) or monocyte chemotactic protein 1 (MCP-1), in replacement of part of the capsid genes. Such rH11 viruses have been shown to retain in vitro the intrinsic oncotropic properties of the parental virus. On infection with the recombinant viruses at an input multiplicity of 1 replication unit (RU) per cell, HeLa cultures were induced to release 4-10 microg of cytokine per 10(6) cells over a period of 5 days. The expression of the rH1-transduced human cytokine/chemokine could also be detected in tumor material recovered from nude mice that had been subcutaneously engrafted with in vitro-infected HeLa cells. The formation of tumors from HeLa xenografts was reduced by 90% compared with wild-type or mock-infected cells as a result of cells preinfected with IL2-expressing virus at an input multiplicity as low as 1 RU per cell. Tumors arising from HeLa cells infected with transgene-free or MCP1-expressing vectors or with wild-type H1 virus were not rejected at this virus dose. Tumors infected with rH1/IL-2 virus displayed markers indicative of their infiltration with NK cells in which the cytocidal program was activated, whereas little NK activity was detected in wild-type virus or mock-infected tumors. Altogether, these data show that the IL-2 expressing H1 vector was a more potent antineoplastic agent than the parental virus, and point to the possible application of recombinant autonomous parvoviruses toward therapy of some human tumors. PMID- 10724039 TI - Genetic modification of human T cells with CD20: a strategy to purify and lyse transduced cells with anti-CD20 antibodies. AB - A retroviral vector has been constructed that contains the human CD20 cDNA under the control of the Moloney murine leukemia virus (Mo-MuLV) LTR. Freshly isolated mononuclear cells are infected for three consecutive days in the presence of PHA and hrlL-2, and a mean 15.9% of the cells (range, 6.5 to 31.7%) acquire a CD3+CD20+ phenotype. Transduced T lymphocytes grow and expand in vitro for up to 3 weeks like mock-infected cells and, as observed for the T lymphoblastoid CEM cell line, CD20 expression is maintained for several months with no change in the growth curve of the cells. CD20-expressing CEM and fresh T lymphocytes can be positively immunoselected on columns using different anti-CD20 antibodies. Exposure to monoclonal chimeric anti-CD20 IgG1(kappa) Rituximab antibody (Roche), in the presence of complement, results in effective and rapid killing of the transduced CD3+CD20+ human T cells in vitro. This approach represents a new and alternative method to gene manipulation with "suicide" genes for the production of drug-responsive T cell populations, a crucial step for the future management of graft-versus-host disease in bone marrow transplant patients. PMID- 10724041 TI - Adeno-associated virus vectors show variable dependence on divalent cations for thermostability: implications for purification and handling. AB - Recombinant adeno-associated virus (rAAV) shows significant promise as a vector for gene transfer in pre-clinical models of human disease, and is currently being evaluated in human clinical trials. As a consequence, increasing attention is being turned to the important tasks of optimizing rAAV titer, purity, and stability. We have observed dramatic variation in divalent cation dependence for thermostability of different rAAV vectors. To further investigate this observation, the thermostability of eight different vector constructs ranging in size from 73 to 107% of wild-type genome size (4.68 kilobases) was determined in the presence and absence of divalent cations. Virions containing smaller genomes (i.e., <85% wild type) were relatively divalent cation independent for thermostability. In contrast, virions containing recombinant genomes close to, or exceeding, wild-type size (i.e., >95% wild type) were dependent on divalent cations for thermostability. Genome sequence also appeared to be a factor in the thermostability of the larger rAAV vectors. These observations are of both practical and theoretical significance. Divalent cations should be included in all buffer solutions used during rAAV purification and storage, and unnecessary heat exposure avoided. These data also demonstrate that different recombinants of a particular virus should not be assumed to possess the same thermostability profile. PMID- 10724040 TI - Liver bypass significantly increases the transduction efficiency of recombinant adenoviral vectors in the lung, intestine, and kidney. AB - Recombinant adenoviruses have great potential as gene delivery systems because of their ability to infect a wide range of target cells. However, systemic delivery of viral vectors to tissues other than liver and spleen has been inefficient because of the rapid clearance of the circulating virus by the liver. In the present study we tested the hypothesis that a systemic administration of E1 deleted recombinant adenovirus vectors that bypasses the hepatic circulation will lead to enhanced expression of these vectors in extrahepatic tissues. The portal vein and hepatic artery in B6/129 F1 mice were clamped and an E1-deleted recombinant adenovirus carrying the beta-galactosidase gene (Ad.CBlacZ) was then administered through the retroorbital venous plexus. The clamp was released 30 min after viral injection with no major chronic ischemic consequences noted. High levels of LacZ expression were detected predominantly in the vessels and capillaries of the lung, intestinal wall, and renal glomeruli 7 days after viral infusion. The transgene expression persisted for at least 21 days. Intense LacZ staining was also observed in the liver, suggesting that liver infection occurred after the portal clamp was released. A retroorbital infusion of anti-adenovirus neutralizing antibodies 5 min before the release of the portal clamp significantly reduced postclamp viral infection to the liver, while LacZ expression in lung and intestine persisted after the antibody treatment. Taken together, these results suggest that liver bypass can significantly improve the transduction efficiency in the other target organs. This method could be used to develop animal models of human diseases that predominantly affect the vessels of the lung, intestine, and kidney. PMID- 10724042 TI - Gene therapy of malignant gliomas: a phase I study of IL-4-HSV-TK gene-modified autologous tumor to elicit an immune response. PMID- 10724043 TI - Resistance to antituberculosis medications: hard lessons to learn. PMID- 10724044 TI - Unraveling the Tuskegee Study of Untreated Syphilis. PMID- 10724045 TI - Cardiac manifestations of acquired immunodeficiency syndrome. AB - Acquired immunodeficiency syndrome is a serious problem worldwide. Recent advances in the knowledge about human immunodeficiency virus (HIV) replication and the treatment of HIV infection have improved survival in HIV patients. Because of the longer survival in HIV patients, the more manifestations of late stage HIV infection will be seen, including HIV-related cardiac diseases. The common cardiac manifestations in patients with the acquired immunodeficiency virus are pericardial effusion, myocarditis, dilated cardiomyopathy, endocarditis, pulmonary hypertension, malignant neoplasms, and drug-related cardiotoxicity. This review focuses on these cardiac manifestations in patients with the acquired immunodeficiency syndrome. PMID- 10724046 TI - The effects of immunosuppressive and anti-inflammatory medications on fertility, pregnancy, and lactation. AB - Many rheumatic diseases affect women of childbearing age, and the medications used to treat these diseases may affect conception, pregnancy, fetal development, and lactation. Physicians who care for these women need to be aware of the potential adverse effects of these medications, and which medications can be used safely prior to conception and during pregnancy and lactation. Although reviews of individual classes of medications are available, there is no practical and comprehensive review that summarizes all of this information, and includes anticoagulant drugs and 2 recently approved drugs for rheumatoid arthritis. Women who take cytotoxic drugs should be informed of the risks of impaired fertility and congenital malformations, and must use effective methods of contraception. During pregnancy, nonsteroidal anti-inflammatory agents may be used until the last 6 weeks, and low to moderate doses of corticosteroids are safe throughout pregnancy. Among the disease-modifying agents, sulfasalazine and hydroxychloroquine treatment may be maintained. Cytotoxic drugs may be used after the first trimester to treat life-threatening disease. During lactation, prednisone, sulfasalazine, and hydroxychloroquine may be used cautiously. Women using heparin for treatment of antiphospholipid antibody syndrome should take measures to prevent bone loss. Men taking methotrexate, sulfasalazine, cyclosporine, azathioprine, or leflunomide should be apprised of the possibilities of infertility and teratogenicity. PMID- 10724047 TI - Beta-adrenergic blocking agents in heart failure: benefits of vasodilating and non-vasodilating agents according to patients' characteristics: a meta-analysis of clinical trials. AB - BACKGROUND: In patients with heart failure, beta-adrenergic blocking agents reduce overall and cardiovascular mortality. This meta-analysis aimed at clarifying their effect on sudden death, the magnitude of their benefit according to the cause of heart failure, and whether there is any difference between vasodilating and nonvasodilating agents. METHODS: Randomized, clinical trials were included if they evaluated a beta-adrenergic blocking agent without intrinsic sympathomimetic activity, included a control group receiving placebo or standard treatment, evaluated mortality on an intention-to-treat basis, and lasted at least 8 weeks. RESULTS: Twenty-one trials with 5,849 patients (3,130 receiving beta-blockers) were included. Median length of treatment was 6 months. Most patients had mild or moderate heart failure and were treated with angiotensin-converting enzyme inhibitors, diuretics, and digitalis. The beta blockers significantly reduced overall mortality, cardiovascular mortality, and mortality due to pump failure and sudden death by 34% to 39%. The decrease in overall mortality in patients with ischemic heart disease (IHD) (30%) was no different from that among patients with non-IHD (26%) (P = .08). The reduction in overall mortality was greater with vasodilating than with nonvasodilating agents (45% vs 27%; P = .007), particularly in patients without IHD (62%), compared with those with IHD (22%; P =.03). CONCLUSIONS: In patients with heart failure, beta blockers reduce total and cardiovascular mortality at the expense of a decrease in mortality due to pump failure and sudden death. The magnitude of the benefit is similar in patients with IHD and in those with non-IHD. Vasodilating beta blockers have a greater effect on overall mortality than nonvasodilating agents, particularly in patients with non-IHD. PMID- 10724048 TI - Clinical consequences and transmissibility of drug-resistant tuberculosis in southern Mexico. AB - BACKGROUND: Consequences of drug-resistant tuberculosis (TB) in developing countries using directly observed treatment, short-course (DOTS), are not well defined. OBJECTIVE: To determine the impact of drug resistance on clinical outcome and transmission of TB under programmatic conditions. PATIENTS AND METHODS: A prospective cohort and molecular epidemiologic study was conducted in southern Mexico. Between March 1995 and February 1998 all patients with persistent cough whose sputa had acid-fast bacilli (AFB) underwent clinical and mycobacteriologic evaluation (species identification, drug susceptibility testing, and IS6110-based genotyping). Treatment was provided in accordance with Mexico's National Tuberculosis Program. Clinical and microbiologic outcomes and molecular epidemiologically defined transmission were measured. RESULTS: Mycobacterium tuberculosis was isolated from 238 of the 284 AFB smear-positive persons. The overall rate of resistance was 28.4% (new, 20.7%; retreated, 54.7%), and 10.8% (new, 3.3%; retreated, 35.8%) had multi-drug-resistant TB (ie, resistance to isoniazid and rifampin). After treatment, 75% (new, 81.0%; retreated, 52.8%) were cured, 8% (new, 7.8%; retreated, 7.5%) abandoned therapy, 9% (new, 3.9%; retreated, 28.3%) had treatment failure, and 4% (new, 3.3%; retreated, 7.5%) died. Another 2% of patients relapsed, and 9% died during a median of 24.4 months of follow-up. Drug-resistance was a strong independent risk factor for treatment failure. Being infected with multi-drug-resistant TB was the only factor associated with a decreased likelihood of being in a restriction fragment length polymorphism cluster. CONCLUSIONS: Despite the use of DOTS, patients with drug-resistant TB had a dramatically increased probability of treatment failure and death. Although multi-drug-resistant TB may have a decreased propensity to spread and cause disease, it has a profoundly negative impact on TB control. PMID- 10724049 TI - Survey of drug resistance of Mycobacterium tuberculosis in 3 Mexican states, 1997. AB - BACKGROUND: Drug resistance threatens global tuberculosis (TB) control efforts. Population-based estimates of drug resistance are needed to develop strategies for controlling drug-resistant TB in Mexico. OBJECTIVE: To obtain population based data on Mycobacterium tuberculosis drug resistance in Mexico. METHODS: To obtain drug resistance data, we conducted a population-based study of TB cases in the states of Baja California, Sinaloa, and Oaxaca, Mexico. We performed cultures and drug susceptibility testing on M tuberculosis isolates from patients with newly diagnosed, smear-positive TB from April 1 to October 31, 1997. RESULTS: Mycobacterium tuberculosis was isolated from 460 (75%) of the 614 patients. Levels of resistance in new and retreatment TB cases to 1 or more of the 3 current first-line drugs used in Mexico (isoniazid, rifampin, and pyrazinamide) were 12.9% and 50.5%, respectively; the corresponding levels of multi-drug resistant TB were 2.4% and 22.4%. Retreatment cases were significantly more likely than new cases to have isolates resistant to 1 or more of the 3 first-line drugs (relative risk [RR], 3.9; 95% confidence interval [CI], 2.8-5.5), to have isoniazid resistance (RR, 3.6; 95% CI, 2.5-5.2), and to have multi-drug-resistant TB (RR, 9.4; 95% CI, 4.3-20.2). CONCLUSIONS: This population-based study of M tuberculosis demonstrates moderately high levels of drug resistance. Important issues to consider in the national strategy to prevent M tuberculosis resistance in Mexico include consideration of the most appropriate initial therapy in patients with TB, the treatment of patients with multiple drug resistance, and surveillance or periodic surveys of resistance among new TB patients to monitor drug resistance trends. PMID- 10724050 TI - Physicians' ethical beliefs about cost-control arrangements. AB - BACKGROUND: Although much has been written about the ethics of new methods of health care financing, little is known about the extent to which physicians experience these cost-control arrangements as ethical problems. METHOD: A cross sectional telephone survey of 1,549 physicians, 8 to 17 years after residency, randomly selected from 75 US metropolitan service areas (response rate, 74.0%). RESULTS: Only 17.0% believed that financial incentives to limit services are ethically acceptable. Although 52.9% thought that physicians should try to abide by guidelines discouraging the use of interventions with possible but unproven benefit, only 14.5% thought such guidelines should be enforced by payers. Only 5.7% thought that it was morally acceptable for payers to discourage physicians from telling patients about their personal financial incentives, and only 9.1% found compliance with such restrictions morally acceptable. Changes in the health care system in the past 5 years were believed to have had a negative impact on their own patients' trust in them by 50.6%, and 80.8% believed that changes in the health care system in the past decade have diminished physicians' commitment to an ethic of undivided loyalty to patients. In multiple regression analysis, physicians who reported that the overall personal financial incentives in their practices encouraged them to reduce services were significantly more likely to have ethical objections to such incentives, to believe their own patients' trust in them had diminished, and to believe that the ethic of undivided loyalty to patients had diminished. CONCLUSIONS: Many of the methods now commonly used to influence medical decision making are considered ethically objectionable by most midcareer physicians. Whether their ethical disquiet about these arrangements is justified cannot be answered from these data. PMID- 10724051 TI - Cigarette smoking and risk of clinically overt thyroid disease: a population based twin case-control study. AB - BACKGROUND: The effects of cigarette smoking on the thyroid gland have been studied for years. However, the effect of smoking on thyroid function and size is still controversial. OBJECTIVE: To determine the impact of cigarette smoking on the development of clinically overt thyroid disease. METHODS: Matched case control study of 132 same-sex twin pairs (264 individuals) discordant for clinically overt thyroid disease, ascertained from a population-based nationwide twin register. Information on thyroid disease and smoking habits was gathered by questionnaire, and the patients' endocrinologist or general practitioner verified the diagnosis. RESULTS: Overall, smoking was associated with an increased risk of developing clinically overt thyroid disease (odds ratio, 3.0; 95% confidence interval, 1.4-6.6; P = .003). This association remained statistically significant in monozygotic and dizygotic disease-discordant pairs. The effect of smoking was more pronounced in monozygotic vs dizygotic pairs (odds ratio, 5.0 vs 2.5; P= .04 for both). Essentially similar results were obtained after subdividing the twin pairs into groups discordant for clinically overt autoimmune (49 pairs) and nonautoimmune (83 pairs) thyroid disease. Among twin pairs concordant for smoking, probands with clinically overt autoimmune thyroid disease smoked significantly more than did their healthy co-twins (17 pairs; P= .03), whereas no difference was found between probands with nonautoimmune thyroid disease and their healthy co-twins (34 pairs; P= .20). CONCLUSIONS: Smoking is associated with an increased risk of developing clinically overt thyroid disease. Furthermore, our data suggest that cumulative cigarette consumption is a risk factor, most pronounced in autoimmune thyroid disease. PMID- 10724052 TI - Postthrombotic syndrome after hip or knee arthroplasty: a cross-sectional study. AB - BACKGROUND: Although the incidence of the postthrombotic syndrome (PTS) has been addressed in patients with symptomatic deep vein thrombosis (DVT), less information is available on the incidence in patients who develop asymptomatic DVT after major hip or knee arthroplasty. OBJECTIVES: To determine whether symptomatic PTS occurs more frequently in patients who develop DVT after hip or knee arthroplasty than those who are free of DVT and to provide an estimate of the incidence of PTS in patients who had undergone major hip or knee arthroplasty and had proximal DVT, distal (calf) DVT, or no DVT. DESIGN AND SETTING: A cross sectional study conducted at the Hamilton Health Sciences Corporation, Hamilton, Ontario, and the Academic Medical Centre, Amsterdam, the Netherlands. SUBJECTS AND METHODS: Two hundred fifty-five subjects who had undergone major hip or knee arthroplasty 2 to 7 years previously and had routine predischarge venography showing proximal DVT (n = 25), distal DVT (n = 66), or no DVT (n = 164) were enrolled from March 1993 through December 1998. The presence of symptomatic PTS confirmed by the presence of objectively confirmed venous valvular incompetence was ascertained. RESULTS: The rates of PTS were low and not significantly different among the 3 subgroups: 1 (4.0%, 95% confidence interval [CI] = 0.1% 20.4%) of 25 patients with proximal DVT, 4 (6.1%, 95% CI = 1.7%-14.8%) of 66 patients with distal DVT, and 7 (4.3%, 95% CI = 1.7%-8.6%) of 164 patients with no DVT. CONCLUSIONS: Symptomatic PTS is an uncommon complaint after major hip or knee arthroplasty. Patients who develop postoperative proximal or distal DVT and who receive 6 to 12 weeks of anticoagulant therapy are not predisposed to PTS. PMID- 10724053 TI - The relationship between pyuria and infection in patients with indwelling urinary catheters: a prospective study of 761 patients. AB - BACKGROUND: Pyuria is universally considered as essential for identifying urinary tract infections in noncatheterized patients. The utility of pyuria in the catheterized patient, to identify catheter-associated urinary tract infection (CAUTI), has not been adequately defined. METHODS: We prospectively studied 761 newly catheterized patients in a university hospital; 82 (10.8%) developed nosocomial CAUTI (> 10(3) colony-forming units per milliliter). While catheterized, each patient was seen daily, a quantitative urine culture was obtained, and the urine white blood cell concentration was measured quantitatively using a hemocytometer. RESULTS: The mean urine leukocyte count in patients with CAUTI was significantly higher than in patients without infections (71 vs 4 per microliter; P= .006). Pyuria was most strongly associated with CAUTI caused by gram-negative bacilli (white blood cell count, 121 vs 4 per microliter; P = .03); infection with coagulase-negative staphylococci and enterococci (white blood cell count, 39 vs 4 per microliter; P = .25) or yeasts (white blood cell count, 25 vs 4 per microliter; P = .15) produced much less pyuria. Pyuria with a white blood cell count greater than 10 per microliter (>5 per high-power field in a conventional urinalysis) had a specificity of 90% for predicting CAUTI with greater than 10(5) colony-forming units per milliliter but a sensitivity of only 37%. CONCLUSIONS: In patients with short-term indwelling urinary catheters, pyuria is less strongly correlated with CAUTI than in noncatheterized patients with urinary tract infection. The strongest association is with CAUTI caused by gram-negative bacilli; the association is far weaker for infections caused by gram-positive cocci or yeasts. Most patients with CAUTI are asymptomatic and do not have associated fever. Pyuria should not be used as the sole criterion to obtain a urine culture in a patient with a catheter. PMID- 10724054 TI - Catheter-associated urinary tract infection is rarely symptomatic: a prospective study of 1,497 catheterized patients. AB - BACKGROUND: Catheter-associated urinary tract infection (CAUTI) is the most common nosocomial infection, accounting for more than 1 million cases each year in US hospitals and nursing homes. OBJECTIVE: To define the clinical features of CAUTI. SETTING AND PATIENTS: A university hospital; 1,497 newly catheterized patients. DESIGN: Every day that the catheter was in place, a quantitative urine culture and urine leukocyte count were obtained, and the patient was queried by a research worker regarding symptoms. To more precisely define the role of CAUTI in patients' symptoms, a subset of 1,034 patients, 89 of whom developed CAUTI with more than 10(3) colony-forming units per milliliter, who did not have another potentially confounding site of infection besides the urinary tract, was analyzed. OUTCOME MEASURES: Presence of fever, symptoms commonly associated with community-acquired urinary tract infection, and peripheral leukocytosis. RESULTS: There were 235 new cases of nosocomial CAUTI during the study period. More than 90% of the infected patients were asymptomatic; only 123 infections (52%) were detected by patients' physicians using the hospital laboratory. In the subset analysis, there were no significant differences between patients with and without CAUTI in signs or symptoms commonly associated with urinary tract infection fever, dysuria, urgency, or flank pain-or in leukocytosis. Only 1 of the 235 episodes of CAUTI that were prospectively studied was unequivocally associated with secondary bloodstream infection. CONCLUSIONS: Whereas CAUTIs are a major reservoir of antibiotic-resistant organisms in the hospital, they are rarely symptomatic and infrequently cause bloodstream infection. Symptoms referable to the urinary tract, fever, or peripheral leukocytosis have little predictive value for the diagnosis of CAUTI. PMID- 10724055 TI - Angiotensin-converting enzyme inhibitor-associated elevations in serum creatinine: is this a cause for concern? AB - BACKGROUND: Reducing the actions of the renin-angiotensin-aldosterone system with angiotensin-converting enzyme inhibitors (ACEIs) slows nephropathy progression in patients with or without diabetes. Post hoc analyses of many ACEI-based clinical trials demonstrate the greatest slowing of renal disease progression in patients with the greatest degree of renal insufficiency at study initiation. However, many physicians fail to use ACEIs or angiotensin receptor blockers in patients with renal insufficiency for fear that either serum creatinine or potassium levels will rise. OBJECTIVE: To determine if limited initial reduction in either glomerular filtration rate (GFR) or elevation in serum creatinine levels, associated with ACEI or angiotensin receptor blocker use, results in long-term protection against decline in renal function in patients with renal insufficiency. METHODS: We reviewed 12 randomized clinical trials that evaluated renal disease progression among patients with preexisting renal insufficiency. Six of these studies were multicenter, double-blinded, and placebo controlled, with the remainder being smaller randomized studies with a minimum 2-year follow up on renal function. These investigations evaluated patients with and without diabetes or systolic heart failure. Average duration of follow-up for all studies was 3 years. Trials were examined in the context of changes in either serum creatinine levels or GFR in the group randomized to an ACEI (N = 1,102). Sixty four percent of these individuals (705/1,102) had renal function data at both less than 6 months and at the end of the study. RESULTS: Most trials demonstrated that patients with preexisting renal insufficiency manifested an acute fall in GFR, a rise in serum creatinine, or both. Those randomized to an ACEI with a serum creatinine level of 124 pmol/L or greater (> or =1.4 mg/dL) demonstrated a 55% to 75% risk reduction in renal disease progression compared with those with normal renal function randomized to an ACEI. An inverse correlation was observed between the amount of renal function loss at baseline and the subsequent rate of annual decline in renal function following randomization to an antihypertensive regimen that contained an ACEI. CONCLUSIONS: A strong association exists between acute increases in serum creatinine of up to 30% that stabilize within the first 2 months of ACEI therapy and long-term preservation of renal function. This relationship holds for persons with creatinine values of greater than 124 pmol/L (>1.4 mg/dL). Thus, withdrawal of an ACEI in such patients should occur only when the rise in creatinine exceeds 30% above baseline within the first 2 months of ACEI initiation, or hyperkalemia develops, ie, serum potassium level of 5.6 mmol/L or greater. PMID- 10724056 TI - Adherence to isoniazid prophylaxis in the homeless: a randomized controlled trial. AB - OBJECTIVES: To test 2 interventions to improve adherence to isoniazid preventive therapy for tuberculosis in homeless adults. We compared (1) biweekly directly observed preventive therapy using a $5 monetary incentive and (2) biweekly directly observed preventive therapy using a peer health adviser, with (3) usual care at the tuberculosis clinic. METHODS: Randomized controlled trial in tuberculosis-infected homeless adults. Outcomes were completion of 6 months of isoniazid treatment and number of months of isoniazid dispensed. RESULTS: A total of 118 subjects were randomized to the 3 arms of the study. Completion in the monetary incentive arm was significantly better than in the peer health adviser arm (P = .01) and the usual care arm (P = .04), by log-rank test. Overall, 19 subjects (44%) in the monetary incentive arm completed preventive therapy compared with 7 (19%) in the peer health adviser arm (P = .02) and 10 (26%) in the usual care arm (P = .11). The median number of months of isoniazid dispensed was 5 in the monetary incentive arm vs 2 months in the peer health adviser arm (P = .005) and 2 months in the usual care arm (P = .04). In multivariate analysis, independent predictors of completion were being in the monetary incentive arm (odds ratio, 2.57; 95% CI, 1.11-5.94) and residence in a hotel or other stable housing at entry into the study vs residence on the street or in a shelter at entry (odds ratio, 2.33; 95% CI, 1.00-5.47). CONCLUSIONS: A $5 biweekly cash incentive improved adherence to tuberculosis preventive therapy compared with a peer intervention or usual care. Living in a hotel or apartment at the start of treatment also predicted the completion of therapy. PMID- 10724057 TI - Physical activity and osteoporotic hip fracture risk in men. AB - BACKGROUND: Physical activity has been related to reduced risk of osteoporotic hip fractures, but the evidence among men is weak. OBJECTIVE: To determine the association between baseline leisure physical activity and future risk of osteoporotic hip fracture in men. METHODS: At baseline in 1975 our prospective study cohort included 3,262 men who were 44 years or older and did not have chronic disease restricting their ability to exercise. At baseline, physical activity was assessed by a questionnaire. Hip fractures were followed for 21 years, or from the age of 50 years for subjects who were initially younger than 50 years. RESULTS: The hazard ratio of osteoporotic hip fracture, adjusted for other possible predictors (height, body mass index, baseline diseases, smoking, use of alcohol, work-related physical activity, and occupational group), in men participating in vigorous physical activity compared with men not participating was 0.38 (95% confidence interval, 0.16-0.91) (P = .03). CONCLUSION: These results provide further evidence that there is an inverse association between baseline physical activity and future hip fracture risk among men. PMID- 10724058 TI - Should elderly individuals who frequently nap take beta-blockers and/or aspirin? PMID- 10724059 TI - Is the siesta associated with sleep apnea syndrome in the elderly? PMID- 10724060 TI - Afternoon nap is good for the elderly. PMID- 10724061 TI - Is taking a siesta really a health hazard? PMID- 10724062 TI - Selective serotonin receptor uptake inhibitors can reduce restless legs symptoms. PMID- 10724063 TI - The effect of acute severe illness on CD4+ lymphocyte counts in nonimmunocompromised patients. PMID- 10724064 TI - Physical medicine and rehabilitation organizations and the future. PMID- 10724065 TI - WWPM&R@MM. World Wide Physical Medicine and Rehabilitation. PMID- 10724066 TI - Halo vest effect on balance. AB - OBJECTIVE: To determine the effect of a halo vest, a cervical orthosis, on clinically relevant balance parameters. DESIGN: Subjects performed unipedal stance (with eyes open and closed, on both firm and soft surfaces) and functional reach, with and without the application of a halo vest. SUBJECTS: A convenience sample of 12 healthy young subjects, with an equal number of men and women. OUTCOME MEASURES: Seconds for unipedal stance (maximum 45); inches for functional reach. RESULTS: Both unipedal stance times and functional reach (mean +/- standard deviation) were significantly decreased with the halo vest as compared to without it (29.1+/-5.8 vs. 32.8+/-6.4 seconds, p = .002; 12.9+/-1.4 vs. 15.1+/ 2.1 inches, p<.01). CONCLUSION: A halo vest causes an acute impairment in balance in the healthy young. It is likely that the impairment would be greater in older or injured patients, thus increasing their risk for a fall, which could have devastating consequences. PMID- 10724067 TI - Interexaminer reliability of the palpation of trigger points in the trunk and lower limb muscles. AB - OBJECTIVES: To determine the interexaminer reliability of palpation of three characteristics of trigger points (taut band, local twitch response, and referred pain) in patients with subacute low back pain, to determine whether training in palpation would improve reliability, and whether there was a difference between the physiatric and chiropractic physicians. DESIGN: Reliability study. SETTING: Whittier Health Campus, Los Angeles College of Chiropractic. PARTICIPANTS: Twenty six nonsymptomatic individuals and 26 individuals with subacute low back pain. INTERVENTION: Twenty muscles per individual were first palpated by an expert and then randomly by four physician examiners. MAIN OUTCOME MEASURES: Palpation findings. RESULTS: Kappa scores for palpation of taut bands, local twitch responses, and referred pain were .215, .123, and .342, respectively, between the expert and the trained examiners, and .050, .118, and .326, respectively, between the expert and the untrained examiners. Kappa scores for agreement for palpation of taut bands, twitch responses, and referred pain were .108, -.001, and .435, respectively, among the nonexpert, trained examiners, and -.019, .022, and .320, respectively, among the nonexpert, untrained examiners. CONCLUSIONS: Among nonexpert physicians, physiatric or chiropractic, trigger point palpation is not reliable for detecting taut band and local twitch response, and only marginally reliable for referred pain after training. PMID- 10724068 TI - Unilateral lower-limb musculoskeletal injury: its long-term effect on balance. AB - OBJECTIVE: To assess if any long-term decrements in balance occur after unilateral musculoskeletal injury. The relation between the size of decrement and the dominance, the type, and the time since injury were also considered. DESIGN: With eyes open and closed, postural sway in one-legged standing was recorded for 10 seconds in 48 subjects who sustained a unilateral musculoskeletal injury 6 months to 42 years earlier. Comparative data were also collected in 108 healthy subjects with no previous injury. SETTING: A university physiologic laboratory. PATIENTS: Injured subjects were recruited locally via the district general hospital, sports injury clinic, and the university, and had not received any treatment within the past 6 months. MAIN OUTCOME MEASURE: Postural sway of the injured and uninjured limb (or dominant and nondominant limb in the uninjured subjects). RESULTS: Postural sway was significantly greater in the injured limb compared with the uninjured limb (p = .0118). The ratio of the postural sway of the injured limb compared with the uninjured limb (I/UI%) was significantly lower in the group with nondominant injuries (p = .0085). Subjects with nondominant injuries performed significantly better than those with dominant injuries (p = .0085). No relation was found between the decrements in balance performance and the type of injury and time since injury. CONCLUSIONS: Full recovery is frequently not achieved and perhaps recovery does not continue to improve once the formal rehabilitation period is over. PMID- 10724069 TI - Stretch reflex adaptation in elbow flexors during repeated passive movements in unilateral brain-injured patients. AB - OBJECTIVE: To evaluate the effects of repeated, externally imposed, flexion extension movements of the elbow on the resulting stretch reflex response in hemiparetic spastic brain-injured patients. These effects were compared within a recording session and across sessions for the same subject to determine the impact of movement history on the quantification of spastic hypertonia using the stretch reflex response. DESIGN: Twenty to 30 sequential, constant velocity flexion-extension movements were applied to the impaired elbow of our cohort, with a 10-second hold interposed between flexion and extension. Movements were applied regularly at 1-minute intervals. Changes in stretch reflex responses were monitored during the applied movements. PARTICIPANTS: We examined a convenience sample of seven hemiparetic brain-injured subjects between the ages of 26 and 60 yrs, with moderate-to-severe spastic hypertonia of elbow muscles (Ashworth score 2-4/4). Subjects participated in 2 to 9 sessions. MEASURES: Elbow torque, position, velocity, and electromyograms of the biceps, brachioradialis, and triceps muscles were recorded for each flexion and extension movement. Stretch reflex torque was calculated by subtracting passive torque from total elbow torque, recorded over large amplitude movements. A linear regression analysis quantified both the initial torque response of the stretch reflex and the ensuing adaptation of the stretch reflex during sequential movements. Intersession variability was characterized both for spastic hypertonia measures and for stretch reflex adaptation. RESULTS: Repeated, externally imposed, sequential flexion-extension movements of the elbow decreased the elbow flexor stretch reflex in six of seven subjects. The mean reduction in reflex torque after 30 movements was 50% of the initial torque values (p = .001, t test vs. 0% change). Intersession stretch reflex responses for each subject were found to vary greatly (SDs of reflex torque ranged from 0.1 to 4.0 Nm), and there were also significant variations in the degree of adaptation between subjects. CONCLUSIONS: Stretch reflex adaptation must be taken into consideration when spastic hypertonia is quantified using repeated joint motion, as is often the case. The magnitude of intersession variation in spastic hypertonia measures suggests that ideally, such measurements should be made across multiple sessions before conclusions are made regarding the efficacy of spastic hypertonia interventions. This study provides quantitative evidence that repeated joint movements may have a significant short term beneficial effect on spastic hypertonia. PMID- 10724070 TI - Quantitative assessment of intrathecally administered baclofen in spasticity. AB - OBJECTIVE: To quantitatively assess the antispastic effect of intrathecally administered baclofen on muscle stiffness in spastic patients. DESIGN: Case control study. SETTING: Clinical laboratory in a university hospital of a city of more than 1,000,000 inhabitants. PARTICIPANTS: Eighteen healthy adult volunteers (9 men, 9 women) were recruited for establishing the normal values. Eleven spastic patients (8 men, 3 women) comprised the study group. MAIN OUTCOME MEASURES: The resistance to passive sinusoidal displacement of 5 degrees imposed to the ankle joint was measured at frequencies from 3 to 12 Hz. Torque and displacement signals were subjected to a Fourier analysis to isolate the elastic and viscous components of the total muscle stiffness. RESULTS: In comparison with the period before intrathecal injection, and with the control group, it was shown that at 4 hours after injection, stretch reflex activity was abolished and elastic and viscous muscle stiffness approached control values. The abnormal residual stiffness concerned only the elastic component due to chronic transformations of the spastic muscle and/or due to changes in joints and periarticular connective tissue. This antispastic effect was completely reversed 36 hours after injection. CONCLUSION: The present study shows that the antispastic effect of intrathecally administered baclofen in spastic patients can be quantitatively assessed by a sensitive method allowing measurement of elastic and viscous components of muscle stiffness. PMID- 10724071 TI - Anatomic motor point localization for partial quadriceps block in spasticity. AB - OBJECTIVE: To identify the location of the vastus intermedius nerve and its motor point (point M) and to precisely identify its coordinates in relation to anatomic surface landmarks. DESIGN: Descriptive study. SETTING: Anatomy institute of a university school of medicine. PARTICIPANTS: Twenty-nine adult cadaver limbs immobilized in anatomic position. INTERVENTION: Anatomic dissection to identify point M. Anatomic surface landmarks were point F, the issuing point of femoral nerve under the inguinal ligament; point R, the middle of superior edge of the patella; segment FR, which corresponds to thigh length; point M', point M orthogonal projection on segment FR. MEAN OUTCOME MEASURE: Absolute vertical coordinate, distance FM, relative vertical coordinate compared to the thigh length, FM'/FR ratio; absolute horizontal coordinate, distance MM'. RESULTS: The absolute vertical coordinate was 11.7+/-2 cm. The relative vertical coordinate was at .29+/-.04 of thigh length. The horizontal coordinate was at 2+/-.5 cm lateral to the FR line. CONCLUSION: Point M can be defined with relative precision by two coordinates. Application and clinical interest of nerve blocking using these coordinates in quadriceps spasticity should be studied. PMID- 10724072 TI - Performance of persons with juvenile-onset amputation in driving motor vehicles. AB - OBJECTIVE: To study the driving of motor vehicles by persons with juvenile-onset amputation and to compare the percentage of drivers among them with that found in the general population. DESIGN: A follow-up study of subjects who were younger than 18 years of age at amputation and who underwent one-sided amputation, covering the period 1976 to 1996. SETTING: The Prosthesis Service of the Asturias Central Hospital, Spain. SUBJECTS: A total of 236 juvenile amputee patients. RESULTS: The percentage of women with amputations who drive is lower than that of their male counterparts (p<.05). The percentage of drivers with upper limb amputations is greater than that of drivers with amputation of the lower limb (p<.05). Motor vehicle adaptations were used more frequently by people with upper limb amputations (p<.05). The ability to drive was not affected by the etiology or the side of amputation, or by the use of a prosthesis. The level of amputation affected driving ability in cases of amputation of the lower limb, but not in those of amputation of the upper limb. CONCLUSION: The percentage of persons with juvenile-onset amputation who drive (47.4%) is similar to that found in the general population (40.8%), and the use of a prosthesis does not have any influence on the capacity to drive a car--89.2% of drivers and 93.5% of nondrivers used a prosthesis. PMID- 10724073 TI - Rehabilitation and the long-term outcomes of persons with trauma-related amputations. AB - OBJECTIVE: To examine the long-term outcomes of persons undergoing trauma-related amputations, and to explore factors affecting their physical, social, and mental health and the role of inpatient rehabilitation in improving such outcomes. DESIGN: Abstracted medical records and interview data sought for a retrospective cohort of persons who had undergone a lower-limb trauma-related amputation. PARTICIPANTS: Patients identified with a principal or secondary diagnosis of a trauma-related amputation to the lower extremity at the University of Maryland Shock Trauma Center between 1984 and 1994. Patients with spinal cord injury or traumatic brain injury were excluded. RESULTS: Of 146 patients who had trauma related amputations to the lower limb at the University of Maryland Shock Trauma Center during the study period, nearly 9% died during the acute admission and 3.5% died after discharge. About 87% of all trauma-related amputations involved males, and roughly three quarters involved white persons. About 80% of all amputations occurred before age 40. The health profile of traumatic amputee subjects interviewed in the study (n = 78, 68% response rate) was systematically lower than that of the general US population for all SF-36 scores. The differences in profiles were largest among SF-36 scales sensitive to differences in physical health status, particularly physical functioning, role limitations due to physical health, and bodily pain. About one fourth of persons with a trauma-related amputation reported ongoing severe problems with the residual limb, including phantom pain, wounds, and sores. The number of inpatient rehabilitation nights significantly improved the ability of patients with amputation to function in their physical roles, increased vitality, and reduced bodily pain. Inpatient rehabilitation was also significantly correlated with improved vocational outcomes. CONCLUSIONS: These findings suggest a substantial effect of inpatient rehabilitation in improving long-term outcomes of persons with trauma-related amputations. PMID- 10724074 TI - Treadmill training with partial body weight support in nonambulatory patients with cerebral palsy. AB - OBJECTIVE: To examine the potential role of treadmill training with partial body weight support in nonambulatory children with cerebral palsy. STUDY DESIGN: Open, nonrandomized, baseline-treatment study. SETTING: An outpatient rehabilitation clinic. SUBJECTS: Ten children with cerebral palsy. Six children (group A) were nonambulatory, and four children (group B) either required continuous physical help (two cases) or were able to walk short distances with a stand-by or independently (one case each). INTERVENTION: Three months of additional treadmill training, three times a week, 25 minutes a session. MAIN OUTCOME MEASURES: Functional Ambulation Categories, standing and walking section of the Gross Motor Function Measure, assessed at two baseline measurements 6 and 3 weeks before the study onset, at the beginning, and at the end of therapy. RESULTS: Measurements during baseline and at the study onset did not differ. During therapy, the mean Functional Ambulation Category improved significantly from 1.1 to 1.9 (p<.05). The sum score of the standing section of the Gross Motor Function Measure increased by 47% (p<.05). The walking section score increased by 50% (p<.01). Of the six nonambulant children in group A, transfer abilities improved in four, one child could walk short distances independently, and two children could walk with continuous physical support after therapy. Of group B, one child could climb stairs independently, three children only needed verbal support while walking, and all subjects could then stand up arm-free after therapy. CONCLUSIONS: Treadmill training with partial body weight support is a promising treatment technique in nonambulatory children with cerebral palsy. PMID- 10724075 TI - Measuring functional status and family support in older school-aged children with cerebral palsy: comparison of three instruments. AB - OBJECTIVE: To compare a pediatric and an adult version of a functional status measure and a family support measure for assessing school-age children with spastic cerebral palsy. DESIGN: A prospective study involved functional status measurements using the Pediatric Functional Independence Measure (WeeFIM), the Adult Functional Independence Measure (FIM), and a family support measure, the Amount of Assistance Questionnaire (AAQ). PARTICIPANTS: The feasibility sample consisted of 47 children aged 2 to 12 yrs with cerebral palsy. The study sample consisted of 20 children aged 7 to 16 yrs with spastic cerebral palsy (50% diplegia, 50% quadriplegia). INTERVENTIONS: Initial assessment interview included the WeeFIM, developmental milestones, educational achievement information, and the AAQ. Within 1 month, a follow-up phone interview using the FIM was completed. MAIN OUTCOME MEASURE: The WeeFIM and FIM measure independence in self-care, sphincter control, mobility, locomotion, communication, and social cognition. The AAQ measures the time and assistance required by a child in essential daily tasks. RESULTS: Pearson's correlation coefficient exceeded .97 for WeeFIM and FIM total score in the total sample as well as in two subgroups of children: those with diplegia and quadriplegia. Total scores in WeeFIM and FIM as well as domain scores were significantly different between children with diplegia and quadriplegia. Parental amount of assistance on the AAQ was significantly correlated with WeeFIM and FIM scores. CONCLUSION: Either the WeeFIM or FIM can be used for monitoring functional status through adolescence in children with spastic cerebral palsy. Functional limitations are highly related to requirements for parental assistance. PMID- 10724076 TI - Exercise training in the debilitated aged: strength and functional outcomes. AB - OBJECTIVE: Resistance and endurance training result in gains in fitness in the aged. It is unclear whether the debilitated elderly can perform moderate intensity training and whether such training results in short-term improvements in strength, endurance, and function in this population. DESIGN: Randomized, controlled trial. SETTINGS AND PATIENTS: Subjects were from a Veterans Affairs nursing home and rehabilitation unit and a community nursing home. They were older than 60 yrs with impairment in at least one physical activity of daily living. Seventy-eight subjects volunteered and 58 (mean age, 75 yrs; 9 women, 49 men) completed the intervention and initial posttest. Only one subject withdrew because of injury or disinterest. INTERVENTION: Thrice-weekly resistance training (using an isokinetic dynamometer) and twice-weekly endurance training for 4 to 8 weeks. MAIN OUTCOMES: Isometric strength in dominant arm and leg, heart rate response to timed endurance test, and activities of daily living score. RESULTS: The mean change in isometric strength across the muscle movements tested was 32.8% in the training group and 10.2% in the control group (difference, 22.6%; 95% confidence interval, 6.2% to 39.0%). No change in heart rate during exercise was seen in the training group. Trained subjects tended to have a greater improvement in functional activity than control subjects, which was statistically significant (p = .04) for those subjects who at enrollment were most dysfunctional (i.e., activities of daily living score less than 13 [maximum score 26]). CONCLUSION: This group of debilitated elderly patients effectively performed resistance training and increased their strength, with the most impaired gaining the most function. Few in the group could effectively perform endurance training. PMID- 10724077 TI - Long-term home exercise program: effect in women at high risk of fracture. AB - OBJECTIVE: To determine whether a better outcome in terms of physical frailty could be achieved with a regular home exercise program in women at high risk of fracture. DESIGN: Prospective long-term observational study. SETTING: Outpatient clinic of physical medicine and rehabilitation. PARTICIPANTS: Women with a history of postmenopausal fractures and an age-adjusted low bone mass, as determined 7 to 12 years earlier. INTERVENTION: Home exercise program. OUTCOME MEASURES: Thirty-three women were followed. The exercise group and control group were compared with regard to fracture rates, episodes of falling, neuromuscular performance (one-leg stance, chair rise, body sway, tandem walk, tapping test), and bone mineral density (BMD). RESULTS: Twenty-five women with a mean age of 73.8+/-5.7 yrs appeared for the investigation. An exercise program had been prescribed in 19 women, and six served as controls. Seven women of the exercise group (36.8%) regularly performed the exercises. No differences between participants of the groups in terms of fracture rates, falling episodes, neuromuscular performance, and BMD were observed. CONCLUSION: It appears that a home exercise program does not affect the outcome of postmenopausal women at high risk of fracture. PMID- 10724078 TI - A controlled study on the effects of transcutaneous electrical nerve stimulation and interferential therapy upon the RIII nociceptive and H-reflexes in humans. AB - OBJECTIVE: To study the effect of transcutaneous electrical nerve stimulation (TENS) and interferential therapy (IFT) upon the RIII nociceptive reflex and H reflex. DESIGN: Double-blind conditions. PARTICIPANTS: Seventy healthy subjects were randomly allocated to one of seven groups (n = 10 per group): Control, TENS 1 (5 Hz), TENS 2 (100 Hz), TENS 3 (200 Hz), IFT 1 (5 Hz), IFT 2 (100 Hz), IFT 3 (200 Hz). INTERVENTION: In the treatment groups, stimulation was applied over the right sural nerve for 15 minutes. MAIN OUTCOME MEASURES: Ipsilateral RIII and H reflexes were recorded before treatment, immediately after treatment, and subsequently at 25, 35, and 45 minutes. Subjects rated the pain associated with the RIII reflex using a computerized visual analogue scale (VAS). RESULTS: Statistical analysis using ANOVA showed no significant differences between baseline and posttreatment measurement for RIII reflex, H-reflex, or VAS data. CONCLUSION: These results suggest that neither type of electrical stimulation (TENS or IFT) affects the RIII or H-reflexes, at least using the parameters and application time in this study. PMID- 10724079 TI - Sacroiliac joint pain referral zones. AB - OBJECTIVE: To determine the patterns of pain referral from the sacroiliac joint. STUDY DESIGN: Retrospective. PARTICIPANTS/METHODS: Fifty consecutive patients who satisfied clinical criteria and demonstrated a positive diagnostic response to a fluoroscopically guided sacroiliac joint injection were included. Each patient's preinjection pain description was used to determine areas of pain referral, and 18 potential pain-referral zones were established. OUTCOME MEASURES: Observed areas of pain referral. RESULTS: Eighteen men (36.0%) and 32 women (64.0%) were included with a mean age of 42.5 years (range, 20 to 75 yrs) and a mean symptom duration of 18.2 months (range, 1 to 72 mo). Forty-seven patients (94.0%) described buttock pain, and 36 patients (72.0%) described lower lumbar pain. Groin pain was described in 7 patients (14.0%). Twenty-five patients (50.0%) described associated lower-extremity pain. Fourteen patients (28.0%) described leg pain distal to the knee, and 6 patients (14.0%) reported foot pain. Eighteen patterns of pain referral were observed. A statistically significant relationship was identified between pain location and age, with younger patients more likely to describe pain distal to the knee. CONCLUSIONS: Pain referral from the sacroiliac joint does not appear to be limited to the lumbar region and buttock. The variable patterns of pain referral observed may arise for several reasons, including the joint's complex innervation, sclerotomal pain referral, irritation of adjacent structures, and varying locations of injury with the sacroiliac joint. PMID- 10724080 TI - Predicting elementary school participation in children with disabilities. AB - OBJECTIVE: To identify predictors of participation in school activities from two sets of functional variables using classification and regression tree analysis. DESIGN: Relational study. PARTICIPANTS: A nationwide sample of 341 children with various disabling conditions, including physical and cognitive/behavioral types of impairment and various severity levels. Children attended public elementary school in 40 states in the United States. MAIN OUTCOME MEASURE: Overall participation in elementary school, combining children's participation in six different environments (transportation, transitions, classroom, cafeteria, bathroom, and playground), as measured by the newly developed School Function Assessment. The children were dichotomized into full (n = 117) and limited (n = 224) participation categories. RESULTS: Two classification trees were developed identifying a small set of predictors from variables measuring performance of functional tasks and discrete activities. Final predictive models included physical and cognitive-behavioral variables, suggested important interactions among predictors, and identified meaningful cut-off points that classified the sample into the outcome categories with about 85% accuracy. CONCLUSIONS: Limited participation was predicted by information about children's physical capabilities. Full participation was predicted by a combination of physical and cognitive-behavioral variables. Findings underscore the relative utility of functional performance compared with impairment information to predict the outcome, and suggest pathways of influence to consider in future research and intervention efforts. PMID- 10724081 TI - Lack of predictive power of the "tethered" median stress test in suspected carpal tunnel syndrome. AB - OBJECTIVE: To investigate the value of the "tethered" median nerve stress test (TMST) in predicting electrodiagnostically confirmed carpal tunnel syndrome (CTS) in patients with symptoms suggestive of CTS. STUDY DESIGN: Blinded comparison of a clinical diagnostic test with neurophysiologic testing. SETTING: Portland (OR) Veterans Administration Medical Center Electrodiagnostic Laboratory. PATIENTS: One hundred two consecutive patients referred for symptoms suggestive of CTS. Study inclusion criteria were referral for evaluation of symptoms of paresthesia (with or without pain) inclusive of the median nerve distribution distal to the wrist. RESULTS: Fifty-seven percent of referred patients had electrodiagnostically confirmed CTS. The sensitivity of the TMST was 50%. The specificity was 59.1%. The positive predictive value was 61.7%. The negative predictive value was 47.3%. CONCLUSION: The TMST does not have utility in predicting electrodiagnostic consult results in veteran patients with symptoms suggestive of CTS. PMID- 10724082 TI - Contribution of passive stiffness to ankle plantarflexor moment during gait after stroke. AB - OBJECTIVE: To measure the contribution of passive stiffness to the ankle plantarflexor moment during gait in subjects with hemiparesis early after stroke. The relationship of passive stiffness with gait speed was also examined. DESIGN: Cross-sectional, descriptive. PATIENTS AND OTHER PARTICIPANTS: A sample of convenience of 14 patients (54.7+/-10.9 yrs) with a hemiparesis for less than 5 months and 11 healthy controls (50.6+/-11.6 yrs). MAIN OUTCOME MEASURES: The contribution of passive stiffness to the plantarflexor moment during gait was obtained using moment-angle slope (stiffness) values. Total plantarflexor stiffness was measured during gait, and passive stiffness was measured during passive dorsiflexion imposed by an isokinetic dynamometer at velocities and ranges of movement matched with values recorded during the plantarflexor lengthening period of the stance phase. The contribution of passive stiffness was obtained by dividing the passive stiffness (dynamometer) by the total plantarflexor stiffness (gait). RESULTS: On the paretic side, passive stiffness contributed more (16.8%; range 2.9% to 49.6%) to total plantarflexor stiffness during gait compared (p<.01) with both the nonparetic side (7.3%) and control values (5.9%). This increased contribution on the paretic side resulted from a large muscle-tendon passive stiffness, a decreased active muscle contribution, or both. Although in some patients the increased passive component led to the development of a total plantarflexor stiffness that was within normal values, it did not in others either because the active component was very small or because limited dorsiflexion during the stance phase prevented the passive component tension to develop. The contribution of passive stiffness was not significantly (p>.05) related to gait speed in both the patients and the controls. CONCLUSIONS: The increased contribution of passive stiffness to total plantarflexor moment during gait likely acts as an adaptation for a defective muscle active component, helping ankle push-off at the end of the stance phase. Although this mechanism is effective in most of the patients, it cannot come into action if the dorsiflexion movement during the stance phase is prevented, for instance, by enhanced stretch reflexes. PMID- 10724083 TI - Predictors for return to work after spinal cord injury: a 3-year multicenter analysis. AB - OBJECTIVE: To examine the ability of the Motor Index Score (MIS), in combination with demographic variables, to predict return to work during a 3-year period for individuals with spinal cord injury (SCI). METHODS: Prospectively collected data, between 1986 and 1995, submitted to the National Spinal Cord Injury Statistical Center were analyzed to determine the prediction of return to work utilizing variables of education, ethnicity, age, marital status, gender, and MIS. Individuals, aged 18 to 65 yrs, employed at the time of their injury, were evaluated at discharge from rehabilitation and at 1 (YR1), 2 (YR2), and 3 (YR3) years postinjury (sample sizes of 1,857, 1,486, and 1,177, respectively). RESULTS: The most important predictors of return to work were education, MIS, ethnicity, and age at onset of SCI. These variables resulted in a high rate of accuracy for predicting across all 3 yrs (YR1, 81%; YR2, 82%; YR3, 77%). CONCLUSIONS: The ability to predict return to work after SCI was shown utilizing MIS and demographic variables, with nearly 80% accuracy. This suggests that return to work after SCI is a dynamic process, with the level of importance of each variable changing with time postinjury. PMID- 10724084 TI - Postural sway of the affected and nonaffected pelvis and leg in stance of hemiparetic patients. AB - OBJECTIVES: To study potential differences between sway of the paretic and of the nonparetic pelvis and leg in standing hemiparetic patients by comparing measurements of corresponding bilateral waist and leg sites, and by comparing the results to those of healthy control subjects. DESIGN AND SETTING: Anterior posterior and mediolateral sway of 15 hemiparetic patients and 13 healthy control subjects was measured with the eyes open and closed during quiet stance. Data were collected via an ultrasonic-based system that continuously monitored the position of four transducers mounted bilaterally on the anterior aspect of the pelvis and on each tibial tuberosity. Sway of each transducer marker was calculated by the standard deviation around its mean position and by its mean speed. Descriptive statistics, analysis of variance, and cross-correlation analysis were used for comparing hemiparetic patients with healthy subjects, as well as for determining the effects of body level, body side, and vision on postural sway. RESULTS: For all four measurement sites and in both the anterior posterior and mediolateral axes, the hemiparetic patients had larger sways than the control subjects. Patients' sway on the paretic side was larger than on the nonparetic side, whereas no side differences were detected in the control subjects. For both groups, waist sway was larger than legs' sway. Cross correlation values between sway of the ipsilateral waist and leg on each body side, as well as between the two legs, were substantially lower in the patients than in healthy subjects. CONCLUSIONS: Postural sway of standing hemiparetic patients is characterized by an asymmetrical profile that is expressed both in larger sway values of the paretic than of the nonparetic side, and in low temporal synchronization between sway of the legs and of the pelvis as well as between the two legs. Impairment in the ability to stabilize the distal segments of the lower extremity on the paretic side, rather than in stabilization of the pelvis, appears to underlie the enhanced postural sway of hemiparetic patients during stance. PMID- 10724085 TI - Soft tissue injuries to USA paralympians at the 1996 summer games. AB - OBJECTIVE: To report the soft tissue injuries sustained by the members of four disabled sports organizations (DSOs) who competed as the USA Team at the 1996 Paralympic Games. SETTING: 1996 Paralympic Games, Atlanta, Georgia. METHODS: Soft tissue (strain, sprain, tendonitis, bursitis, or contusion) injury frequencies sustained by Disabled Sports USA (DSUSA, n = 66), the United States Association for Blind Athletes (USABA, n = 53), the United States Cerebral Palsy Athletic Association (USCPAA, n = 56), and Wheelchair Sports USA (WSUSA, n = 129) athletes were compared by body region with chi-square tests (p<.05) and standardized residual assessment. RESULTS: A total of 254 soft tissue injuries (67% acute onset, 170/254) were sustained by the participant DSO members. Statistical design limitations and poor USCPAA athlete homogeneity prompted their exclusion from group comparisons (descriptive results are reported). The most common injury regions for specific DSOs were shoulder (26%), hip-thigh (14%), and ankle (12%) for DSUSA; hip-thigh (21%), cervicothoracic region (19%), and shoulder (17%) for USABA; lumbar region (14%), foot-toe (13%), and ankle (9%) for USCPAA; and shoulder (18%), arm-elbow (12%), forearm-wrist (12%), and lumbar region (9%) for USUSA. Chi-square residual analysis showed that the USABA athletes contributed more to cervicothoracic and lower leg region injury frequencies than DSUSA or WSUSA athletes. The WSUSA athletes contributed more to elbow-arm and forearm wrist region injury frequencies than DSUSA or USABA athletes. The DSUSA athletes contributed more to ankle region injury frequencies than USABA or WSUSA athletes. CONCLUSIONS: Differences in soft tissue injury frequency among athletes of differing DSOs suggest that the competitive use of adaptive or assistive devices, in combination with sport-specific stressors and the athletes' disabilities, is related to the development of predictable soft tissue injury patterns. The decreased incidence of shoulder injury among WSUSA athletes suggests that the injury prevention advice provided by previous studies is being implemented among athletes at this competitive level. The increased incidence of ankle injuries among DSUSA athletes suggests lower extremity load imbalances (prosthetic vs. uninvolved) during running. The increased incidence of lower leg injuries among USABA athletes suggests "overuse" injury patterns typical of nondisabled runners, or inadvertent contacts (contused shins), whereas the increased incidence of cervicothoracic injuries suggests injuries related to falls, "near falls," or sudden directional changes prompted by guidance aids. PMID- 10724086 TI - BCG vaccine prevents insulitis in low dose streptozotocin-induced diabetic mice. AB - Autoimmune type 1 diabetes mellitus is caused by the immunologic destruction of pancreatic beta-cells; therefore, there have been many attempts at immunologic modulation as a block or prevention of the underlying process. The aim of this study is to investigate the effect of BCG vaccination on low dose streptozotocin induced diabetic (LDSD) mice. The mice were pretreated with BCG 7 days before starting low dose streptozotocin (STZ), observed body weight and blood glucose for 2 months, then analyzed the severity of the STZ-induced insulitis after the animals were sacrificed. In this experiment, the mean body weights in the BCG-STZ group on days 1, 19, 33, 47, and 61 of the experiment were 37.5 +/- 3.6, 37.3 +/- 3.6, 37.5 +/- 3.5, 39.4 +/- 3.9, and 39.3 +/- 4.5 (g), respectively. Those in the STZ group were 37.7 +/- 3.5, 38.3 +/- 4.5, 38.4 +/- 3.9, 36.2 +/- 4.5, and 36.3 +/- 4.0 (g), respectively (P < 0.05). The mean blood glucose levels in the BCG STZ group on days 1, 19, 33, 47, and 61 were 106.5 +/- 8, 150 +/- 37, 147 +/- 54, 143 +/- 60, and 142 +/- 66 (mg/dl), respectively. Those in the STZ group were 103 +/- 12, 196 +/- 90, 261 +/- 236, 236 +/- 91, and 224 +/- 89 (mg/dl), respectively (P < 0.05). The numbers developing grade 0, I, II, III, and IV insulitis in the BCG-treated group were 63, 48, 33, 4, and 2, respectively, and in the control group were 16, 23, 31, 45, and 35, respectively. This study indicates that BCG vaccination reduces the development of insulitis and overt diabetes in LDSD mice. PMID- 10724087 TI - The effects of tolbutamide on lipoproteins, lipoprotein lipase and hormone sensitive lipase. AB - Type 2 diabetic patients are at increased risk to develop atherosclerotic vascular disease. These patients are often treated with sulphonylurea derivatives, and it has been suggested that this treatment might contribute to the increased atherosclerotic process. The aim of the present study was therefore to investigate whether tolbutamide influences lipid metabolism in such a way that the atherosclerotic process may be promoted. Addition of tolbutamide (5-500 mg/l) to isolated rat fat adipocytes inhibited the lipoprotein lipase (LPL) activity in a dose-dependent manner to levels about 50% of those registered in the absence of tolbutamide. This effect was due to inhibition of the activation of the enzyme in the tissue and not to interference with the interaction of enzyme with its substrate. Addition of tolbutamide (500 mg/l) also inhibited noradrenaline (100 nM) and isoprenaline (40 nM)-induced lipolysis by 48.1 +/- 7.4% (mean +/- S.E.M.) and 47.3 +/- 5.5%, respectively. The decreased lipolysis in tolbutamide preincubated adipocytes was shown to be the result of an inhibition of the phosphorylation of hormone sensitive lipase (HSL). Three months of tolbutamide treatment (0.5 g t.i.d.) in diet treated type 2 diabetic patients did not influence the plasma concentrations of cholesterol, triglycerides, LDL cholesterol, HDL cholesterol as well as HDL triglycerides and HDL phospholipids, and there were no differences compared to placebo treated patients. There was a tendency towards a decrement in the elimination rate of exogenous triglycerides in the tolbutamide group (P = 0.0801). No differences between the groups and no treatment effects were seen on LPL and hepatic lipase activities. In conclusion, our in vitro data show that tolbutamide has dual effects on lipid transport, with impairment of the LPL system, which would tend to decrease plasma lipoproteins by reducing hepatic production of lipoproteins. In vivo, these two effects seem to balance each other and plasma lipoprotein levels remain unaffected. PMID- 10724089 TI - Serum fatty acid composition as a marker of eating habits in normal and diabetic subjects. AB - It is known that the Japanese people have had healthy eating habits, which may explain their low incidence of non-insulin-dependent diabetes mellitus (NIDDM). In the present study, in order to examine dietary habits such as fish consumption, the serum fatty acids from 190 normal people in their 30s and 50s living in Tokyo were surveyed. Furthermore, the fatty acid composition of 44 patients with DM was studied to clarify the difference in eating habits between normal and diabetic patients. The n-3:n-6 ratio of polyunsaturated fatty acids was 0.19 +/- 0.05 for the group in their 30s and 0.27 +/- 0.08 for the group in their 50s (P < 0.01). In patients with DM, the n-3:n-6 ratio was 0.23 +/- 0.05 and glycosylated haemoglobin (HbA1c) was 6.52 +/- 0.55%. After 4 months of dietary education, the n-3:n-6 ratio had increased to 0.27 +/- 0.06 (P < 0.01) and HbA1c had decreased significantly to 6.16 +/- 0.35% (P < 0.01). These findings indicated that people in their 50s tend to eat a traditional diet, which is abundant in fish oil, whereas people in their 30s, as well as patients with NIDDM in their 50s, appear to eat a diet that includes less fish than the traditional diet. Benefits of fish oil have been reported to be limited due to adverse effects in the glycemic controls in patients with NIDDM. Early instruction of newly diagnosed patients to encourage them to change their eating habits to a more traditional diet, which is abundant in fish, proved to be one of the safe methods for interventions in patients with DM. PMID- 10724088 TI - Human insulin B chain but not A chain decreases the rate of diabetes in BB rats. AB - The autoimmune response seen in insulin-dependent diabetes mellitus (IDDM) includes a humoral immune response against human insulin. Early insulin treatment has been used to prevent IDDM in the rodent models of IDDM, and a prevention trial is underway in humans. The metabolic effects of insulin may not be involved in this prevention since, in NOD mice, the use of metabolically inert human insulin B chain was effective. We wished to ascertain whether immunization of diabetes-prone BB/W rats with insulin B chain, A chain, or both could alter the incidence of diabetes. Immunizations began by 30 days of age and the rats were followed until 120 days of age. Only immunization with insulin B chain plus adjuvant was effective in reducing the rate of diabetes. All immunization frequencies were effective, but a significantly lower rate of diabetes was achieved with injections every week. All of the doses tested resulted in significantly lower rates of diabetes. These data confirm in the BB rat model that immunization with insulin B chain in the presence of adjuvant can reduce diabetes incidence. The absence of any metabolic effect of B chain and the requirement for adjuvant suggest that this effect is mediated via modulation of the autoimmune response. PMID- 10724090 TI - Type 1 diabetes mellitus in a routine diabetes clinic: the association of psycho social factors, diabetes knowledge and glycaemic control to insulin regime. AB - In controlled trials intensified diabetes therapy including multiple insulin injection regimes has been shown to improve glycaemic and microvascular disease outcomes in insulin dependent diabetes but this is not clear in routine outpatient practice. We undertook a pragmatic cross sectional study of 200 patients with Type 1 diabetes aged 18-50 years. There were 108 on two insulin injections/day (conventional) and 92 on four injections/day (multiple) with no significant difference for age, sex, social class, body mass index, diabetes duration, hypoglycaemia rate or complications prevalence. The relationship of insulin injection regime used with diabetes knowledge, psychological factors and glycaemic control outcomes was evaluated. Percent glycated HbA1c concentrations (normal range < 5.5%) were worse in the multiple injection group (7.5 +/- 1.7 vs. 6.8 +/- 1.4%, P < 0.001) (mean +/- SD). Their scores for diabetes knowledge (72.5 +/- 8.2 vs. 69.0 +/- 9.8, P < 0.01) were better but treatment satisfaction (29.9 +/- 5.1 vs. 28.5 +/- 6.1, ns) and well-being (49.1 +/- 10.7 vs. 46.5 +/- 12.7, ns) scores were not. Parameters of perceived locus of control were (multiple v conventional): personal (self), 24.5 +/- 5.0 vs. 22.3 +/- 5.9, P < 0.05; medical (doctor), 11.8 +/- 5.1 vs. 10.8 +/- 5.8, ns; situational (chance), 7.9 +/- 5.1 vs. 8.9 +/- 5.9, ns. In multiple regression of HbA1c versus multiple variables only insulin regime (P < 0.001) was significant. We conclude that in routine clinical practice the use of intensive insulin regimes are associated with worse glycaemic control despite patients being marginally more knowledgeable and self directed. PMID- 10724091 TI - Purine metabolites and malondialdehyde in platelets of diabetic patients. AB - The concentration of some of the purine nucleotides and their metabolites together with that of malondialdehyde (MDA) have been measured in resting and stimulated platelets of type 1 and type 2 diabetic patients. While control platelets show a net decrease of guanosine triphosphate (GTP) (3.1 vs. 2.3 nmol per 10(9) platelets) and guanosine diphosphate (GDP) (3.0 vs. 2.0 nmol per 10(9) platelets) and a significant increase of adenosine (0.04 vs. 0.55 nmol per 10(9) platelets) with platelet stimulation, platelets of type 1 and type 2 diabetic patients have a lesser change of these metabolites (GTP, 2.6 vs. 2.4; GDP, 2.3 vs. 2.4; adenosine, 0.04 vs. 0.30 (P < 0.05 vs. control) nmol per 10(9) platelets in type 1 diabetics; GTP, 2.4 vs. 2.7; GDP, 2.4 vs. 2.1; adenosine, 0.08 vs. 0.32 (P < 0.05 vs. control) nmol per 10(9) platelets in type 2 diabetics). These results indicate that the change (stimulated minus resting) of GTP, GDP and adenosine in diabetic platelets is significantly different from that of controls (P < 0.001). Moreover, the amount of MDA produced during platelet activation seems to be lower than controls only in type 2 diabetes (1.81 vs. 2.86 nmol per 10(9) platelets, P < 0.05). These results seem to indicate that a difference in the pattern of platelet nucleotides could be an important feature even in well controlled diabetes, while MDA is probably modified only in association with the late vascular complications of diabetes. PMID- 10724092 TI - Long term effect of a structured inpatient diabetes teaching and treatment programme in type 2 diabetic patients: influence of mode of follow-up. AB - Structured diabetes teaching and treatment programmes (STTP) are increasingly offered for patients with diabetes to improve metabolic control. We prospectively studied the long term-effect of STTP on metabolic control and knowledge of diabetes in patients with type 2 diabetes. In addition, differences in the mode of follow-up by a university diabetes centre (UDC) versus general practitioner (GP) were assessed. Of the 64 patients with type 2 diabetes (61 +/- 10 years old, diabetes duration 11 +/- 7 years) included in the study 52 could be reevaluated after 2 years. Of those, 31 were followed up by the UDC and 21 by their GPs who received detailed follow-up instructions from the UDC. In all patients, HbA1c decreased from 9.1 +/- 0.3% before the programme to 8.3 +/- 0.3% 2 years after the programme (P = 0.004), whereas body mass index increased from 28.8 +/- 0.8 to 30.3 +/- 0.9 kg/m2 (P < 0.001). Patients had a significantly better knowledge of diabetes and diet 2 years after the programme. For all parameters tested, none of the changes differed between patients managed by the UDC versus those managed by their GP. However, patients who chose follow-up by the UDC were more obese and had a better knowledge of diabetes. In conclusion, the STTP for patients with type 2 diabetes was effective in improving the long-term glycaemic control and knowledge of diabetes. Moreover, with precise therapeutic goals and follow-up instructions given to patient and GP this improvement was independent of the mode of outpatient follow-up. PMID- 10724093 TI - Circulating intercellular adhesion molecule-1, vascular cell adhesion molecule-1, and E-selectin in patients with type 2 diabetes mellitus. AB - Serum levels of intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) and E-selectin in patients with type 2 diabetes mellitus (n = 64) and control subjects (n = 40) were studied. Serum ICAM-1 concentrations in diabetic patients were significantly higher than those of control subjects (378.2 +/- 70.0 versus 220.4 +/- 31.8 ng/ml, P < 0.01). By multiple regression analysis, hemoglobin A1c was independently associated with serum ICAM-1 concentration in patients with diabetes. The serum VCAM-1 concentration of diabetic patients with macroangiopathy was higher than those of patients without macroangiopathy and of control subjects (806.9 + 276.5 versus 639.0 +/- 146.0 (P < 0.01), and 652.1 +/- 146.9 ng/ml (P < 0.01), respectively). There was no difference in serum E-selectin concentration between diabetic patients with or without macroangiopathy and normal control subjects. These results suggest that adhesion molecules may contribute to the development of atherosclerosis in the diabetic state. PMID- 10724094 TI - Peripheral blood mitochondrial DNA content correlates with lipid oxidation rate during euglycemic clamps in healthy young men. AB - Both qualitative and quantitative changes in mitochondrial DNA (mtDNA) have been implicated in the pathogenesis of diabetes mellitus. It was previously found that decreased mtDNA content preceded the development of diabetes and mtDNA content correlated with the clinical parameters of insulin resistance syndrome, including diastolic blood pressure and waist-hip ratio. These results prompted one to look whether there are correlations between mtDNA content and the biochemical parameters of insulin resistance in non-diabetic subjects. MtDNA content of peripheral blood leukocytes was measured in Korean healthy young men, and this was correlated with various parameters of fuel metabolism at baseline and during euglycemic hyperinsulinemic clamps with indirect calorimetry. MtDNA content in peripheral blood leukocytes did not correlate with insulin sensitivity index or other metabolic variables such as body mass index (BMI), waist-to-hip ratio (WHR) and blood pressure. However, mtDNA content showed a positive significant correlation with fat oxidation rate during euglycemic clamps (r = 0.61, P < 0.05). Changes in fat oxidation rate and carbohydrate oxidation rate during the clamps were significantly correlated with mtDNA content (r = 0.65, P < 0.05, r = 0.65, P < 0.05, respectively). These results suggest that mtDNA content in peripheral blood may not correlate with insulin resistance per se but with some aspect of insulin resistance in healthy young men. PMID- 10724095 TI - An aldose reductase intragenic polymorphism associated with diabetic retinopathy. AB - The polyol pathway has been considered important in the development of diabetes long-term complications. Diabetic patients with microvascular disease have increased gene expression and enzyme activity, which may be due to variants in the aldose reductase gene. An association of an intragenic BamHI polymorphic site with diabetic retinopathy and nephropathy has been suggested, but the BamHI site has not been confirmed. In the current study, long template PCR-RFLP and DNA sequencing were used to ascertain its existence. A single substitution of A to C at 95th nucleotide of intron 8 was identified. This polymorphism was investigated in 164 adolescents with type 1 diabetes in whom diabetic retinopathy was assessed by stereoscopic retinal photography. Both the wild haplotype B and homozygote BB were significantly more common in the adolescents with retinopathy than in those without retinopathy (P = 0.018 and 0.002, respectively). We also confirmed an association between a previously described (CA)n repeat sequence and retinopathy in these adolescents (P < 0.0005). However, there was no association between the two polymorphisms. PMID- 10724096 TI - Does glucose tolerance affect quality of life in an elderly population? AB - The aim of the present study was to describe the associations between glucose tolerance and quality of life in an unselected non-institutionalised elderly population aged 73 years or over (n = 259, of whom 93 were men). Diabetes was assessed on the basis of self-reports and 2-h oral glucose tolerance tests (1985 WHO criteria). Quality of life was evaluated with the Nottingham Health Profile instrument (NHP). A greater proportion of the previously diagnosed diabetic patients reported to have problems on all the three energy items, on nearly all the physical mobility items and on half of the pain items compared to the subjects with undiagnosed diabetes, impaired glucose tolerance or normal glucose tolerance. The results of the second part of the NHP were in line with those of the first part, showing that more of the persons with previously diagnosed diabetes had problems on the following items: jobs around the house, hobbies and holidays compared to the other study groups. As for the six quality of life dimensions in the first part of the NHP, the previously diagnosed diabetic persons scored clearly higher on the energy, pain and physical mobility dimensions of the NHP compared to all the other subjects. To conclude, elderly subjects with previously diagnosed diabetes had a poorer quality of life compared to those with undiagnosed diabetes, impaired glucose tolerance or normal glucose tolerance. PMID- 10724097 TI - CTLA4 gene polymorphism correlates with the mode of onset and presence of ICA512 Ab in Japanese type 1 diabetes. AB - Recently, the association of CTLA4 gene polymorphism with type 1 diabetes and AITD has been reported in several populations. CTLA4 was originally reported to regulate T-cell activity and T-B cognate interaction. To investigate the role of CTLA4 in autoimmune diseases, we examined the correlation between CTLA4 gene polymorphism and the clinical characteristics of Japanese patients with type 1 diabetes, including the mode of onset of diabetes and presence of islet-specific autoantibodies (GAD, ICA 512 Ab) in the serum. We studied 111 patients with type 1 diabetes and 445 normal subjects. CTLA4 exon 1 position 49 (A/G: codon 17: Thr/Ala) polymorphism was defined, employing PCR-RFLP. Sixty-three (57%) patients had AITD. The allele frequencies of G and A in both 111 patients (G: 65%; A: 35%) and 63 patients (G: 62%; A: 38%) were not significantly different from the control subjects (G: 63%; A: 37%). Serum samples of 69 patients were obtained within a year after onset and used for pancreas specific autoantibodies analysis. These samples were also used for further analysis between CTLA4 gene polymorphism and clinical characteristics. The allele frequencies of G and A in patients who presented with diabetic ketoacidosis (DK+) (G: 75%; A: 25%) were significantly different from those in DK- patients (G: 50%, A: 50%, P = 0.003). Allele and genotype analyses showed significant differences between DK+ patients and control subjects (P = 0.014, P = 0.046, respectively). Allele frequencies of G and A were not significant between patients who were positive and negative for GAD Ab, but significant for ICA 512 Ab (G: 83%, A:17% versus G: 59%, A: 41%: positive patients versus negative patients, P = 0.004). Our results showed a significant correlation between CTLA4 gene polymorphism and ICA 512 Ab. Our results also indicated that CTLA4 gene polymorphism is associated with the onset mode of Japanese type 1 diabetes and the presence of ICA512 Ab. Further analysis of this polymorphism is necessary to fully understand the pathogenesis and progression of type 1 diabetes. PMID- 10724098 TI - Assessment of the function and effect of diabetes education programs in Taiwan. AB - A multi-center prospective study was conducted to assess the function and impact of diabetic education programs on diabetic control. A total of 208 subjects with type 2 diabetes were recruited. Diabetes self-care, assessed by questionnaire, was evaluated before, and 4 months after attending a diabetes education course. A total of 121 subjects who received advanced diabetes education courses were designated as the experimental group. A second group of 87 cases receiving a basic course served as controls. In addition to basic knowledge, the advanced education programs included dietary control, blood glucose monitoring, management of hypoglycemia, medication compliance, foot care and exercise. Diabetes self care techniques were significantly improved in the experimental group. The overall score for diabetes self-care techniques improved in both groups at the 4th month over baseline values. The change was significant with the controls' (P < 0.001). Multiple regression analysis confirmed the intensity of diabetic education was the only significant variable correlated with the decrease of fasting blood glucose and systolic blood pressure. In conclusion, integrated and intensive diabetes education program in diabetes education centers provides an effective method for improving diabetes self-care techniques and metabolic outcome. PMID- 10724099 TI - Molecular basis of Canavan disease. PMID- 10724100 TI - Merosin-deficient congenital muscular dystrophy and cortical dysplasia. AB - Congenital muscular dystrophy (CMD) encompasses a heterogenous group of muscle disorders with autosomal recessive inheritance, characterized by muscular weakness and hypotonia at birth or within the first few months of life and developmental delay. Merosin-deficient CMD is a clinically distinct form which may be associated with significant abnormalities of the brain detectable by neuroimaging. We report two siblings of consanguineous parents with merosin deficient CMD in an Irish family who in addition to the characteristic white matter abnormalities on neuroimaging, had occipital dysplasia. Clinical, electrophysiological muscle biopsy findings and neuroimaging were very similar in both cases. Although merosin-deficient CMD with white matter abnormalities on neuroimaging is well documented in the literature, the association with occipital dysplasia has only rarely been reported. The appearance of an identical cortical defect in these siblings suggests an underlying genetic mechanism. PMID- 10724101 TI - Evaluation of magnetic resonance angiography with selective maximum intensity projection in patients with childhood moyamoya disease. AB - To determine whether magnetic resonance angiography with selective maximum intensity projection can facilitate the detection of cerebral moyamoya vessels in patients with childhood moyamoya disease, six patients with moyamoya disease (6 to 9 years old) and ten controls (4 to 16 years old) were evaluated by means of high resolution magnetic resonance angiography (matrix 512x384) with/without selective maximum intensity projection, and conventional angiography. In the patients with moyamoya disease, moderate or marked moyamoya vessels were detected but underestimated in 2/12 hemispheres on magnetic resonance angiography without selective maximum intensity projection. On magnetic resonance angiography with selective maximum intensity projection, moyamoya vessels were correctly assessed in 11/12 hemispheres (92%). In the controls, bilateral mild moyamoya vessels were detected in eight of 20 hemispheres (four of ten patients, under 7 years old), which were compatible with normal lenticulostriate arteries. Magnetic resonance angiography with selective maximum intensity projection is an accurate modality for assessing moyamoya vessels in moyamoya disease. PMID- 10724102 TI - Lower degree of fever at the initial febrile convulsion is associated with increased risk of subsequent convulsions. AB - We studied 132 children admitted consecutively with their first febrile convulsion to assess whether the degree of fever at the onset of the convulsion can predict the risk of subsequent convulsions. The children studied were reviewed at least 2 years after the initial febrile convulsion to determine the number of children who had recurrences of febrile convulsions and/or afebrile convulsions. Children with body temperatures below 39 degrees C at the onset of their initial febrile convulsion (Group 1) were two and half times more likely to experience multiple convulsions within the same illness than those with body temperatures above 39 degrees C (Group 2). This occurred when the body temperature rose above that which had triggered the initial febrile convulsion. Children in Group 1 were also over three times more likely to experience recurrent febrile convulsion in subsequent illnesses than those in Group 2. As for subsequent development of afebrile convulsion or epilepsy, although the risk was low, it only occurred in Group 1. It is suggested that the known association between multiple convulsions, recurrent febrile convulsions and epilepsy may be due to the single predisposing factor of a low degree of fever at the onset of febrile convulsion. Each child with febrile convulsion may have his own threshold for eliciting a convulsion with fever; the lower this threshold is, the more likely are subsequent convulsions. PMID- 10724103 TI - Congenital myasthenic syndromes. PMID- 10724104 TI - Mitochondrial gene mutations. PMID- 10724105 TI - How primitive is the Moro reflex? PMID- 10724106 TI - Workshop on the genetic and molecular basis of the neuronal ceroid lipofuscinoses. London, UK, 13-16 November 1997. Report and abstracts. PMID- 10724107 TI - Imaging synaptic neurotransmission with in vivo binding competition techniques: a critical review. AB - Several groups have provided evidence that positron emission tomography (PET) and single-photon emission computed tomography (SPECT) neuroreceptor imaging techniques might be applied to measure acute fluctuations in dopamine (DA) synaptic concentration in the living human brain. Competition between DA and radioligands for binding to D2 receptor is the principle underlying this approach. This new application of neuroreceptor imaging provides a dynamic measurement of neurotransmission that is likely to be informative to our understanding of neuropsychiatric conditions. This article reviews and discusses the body of data supporting the feasibility and potential of this imaging paradigm. Endogenous competition studies performed in rodents, nonhuman primates, and humans are first summarized. After this overview, the validity of the model underlying the interpretation of these imaging data is critically assessed. The current reference model is defined as the occupancy model, since changes in radiotracer binding potential (BP) are assumed to be directly caused by changes in occupancy of D2 receptors by DA. Experimental data supporting this model are presented. The evidence that manipulation of DA synaptic levels induces change in the BP of several D2 radiotracers (catecholamines and benzamides) is unequivocal. The fact that these changes in BP are mediated by changes in DA synaptic concentration is well documented. The relationship between the magnitude of BP changes measured with PET or SPECT and the magnitude of changes in DA concentration measured by microdialysis supports the use of these noninvasive techniques to measure changes in neurotransmission. On the other hand, several observations remain unexplained. First, the amphetamine-induced changes in the BP of D2 receptor antagonists [123I]IBZM and [11C]raclopride last longer than amphetamine-induced changes in DA extracellular concentration. Second, nonbenzamide D2 receptor antagonists, such as spiperone and pimozide, are not affected by changes in DA release, or are affected in a direction opposite to that predicted by the occupancy model. Similar observations are reported with D1 radiotracers. These results suggest that the changes in BP following changes in DA concentration might not be fully accounted by a simple occupancy model. Specifically, the data are reviewed supporting that agonist-mediated receptor internalization might play an important role in characterizing receptor-ligand interactions. Finally, it is proposed that a better understanding of the mechanism underlying the effects observed with benzamides is essential to develop this imaging technique to other receptor systems. PMID- 10724108 TI - Reduction of tissue plasminogen activator-induced hemorrhage and brain injury by free radical spin trapping after embolic focal cerebral ischemia in rats. AB - Thrombolytic stroke therapy with tissue plasminogen activator (tPA) remains complicated by serious risks of cerebral hemorrhage and brain injury. In this study, a novel model of tPA-induced hemorrhage was used in spontaneously hypertensive rats to examine the correlates of hemorrhage, and test methods of reducing hemorrhage and brain injury. Homologous blood clot emboli were used to occlude the middle cerebral artery in spontaneously hypertensive rats, and delayed administration of tPA (6 hours postischemia) resulted in high rates of cerebral hemorrhage 24 hours later. Compared with untreated rats, tPA significantly increased hemorrhage volumes by almost 85%. Concomitantly, infarction and neurological deficits were worsened by tPA. A parallel experiment in normotensive Wistar-Kyoto rats showed markedly reduced rates of hemorrhage, and tPA did not significantly increase hemorrhage volumes. To examine whether tPA induced hemorrhage was caused by the delayed onset of reperfusion per se, another group of spontaneously hypertensive rats was subjected to focal ischemia using a mechanical method of arterial occlusion. Delayed (6 hours) reperfusion via mechanical means did not induce hemorrhage. However, administration of tPA plus delayed mechanical reperfusion significantly increased hemorrhage volumes. Since reperfusion injury was implicated, a final experiment compared outcomes in spontaneously hypertensive rats treated with tPA plus the free radical spin trap alpha-phenyl tert butyl nitrone (alpha-PBN) versus tPA alone. tPA-induced hemorrhage volumes were reduced by 40% with alpha-PBN, and infarction and neurological deficits were also decreased. These results indicate that (1) blood pressure is an important correlate of tPA-induced hemorrhage, (2) tPA interacts negatively with reperfusion injury to promote hemorrhage, and (3) combination therapies with anti-free radical treatments may reduce the severity of tPA induced hemorrhage and brain injury after cerebral ischemia. PMID- 10724109 TI - Intraventricular infusion of apolipoprotein E ameliorates acute neuronal damage after global cerebral ischemia in mice. AB - The ability of intraventricular infusion of apolipoprotein E (apoE) to reduce neuronal damage after global cerebral ischemia was investigated in apoE-deficient and wild-type mice. ApoE (5 microg/mL lipid-conjugated derived from human plasma; 1 microL/h, continuous infusion) significantly reduced neuronal damage in the caudate nucleus and CA2 pyramidal cell layer by approximately 50% in apoE deficient mice after global ischemia compared to vehicle infusion. In wild-type mice infused with apoE, there was a trend for ischemic neuronal damage to be reduced. ApoE-infused mice had a marked reduction in 4-hydroxynonenal immunoreactivity, as a marker of lipid peroxidation. The results show that the presence of apoE at or after the time of injury can be neuroprotective, possibly via an anti-oxidant mechanism. PMID- 10724110 TI - ALS-linked Cu/Zn-SOD mutation impairs cerebral synaptic glucose and glutamate transport and exacerbates ischemic brain injury. AB - Although degeneration of lower motor neurons is the most striking abnormality in amyotrophic lateral sclerosis (ALS), more subtle alterations may occur in the brain. Mutations in copper/zinc superoxide dismutase (Cu/Zn-SOD) are responsible for some cases of inherited ALS, and expression of mutant Cu/Zn-SOD in transgenic mice results in progressive motor neuron loss and a clinical phenotype similar to that of ALS patients. It is now reported that Cu/Zn-SOD mutant mice exhibit increased vulnerability to focal ischemic brain injury after transient occlusion of the middle cerebral artery. Levels of glucose and glutamate transport in cerebral cortex synaptic terminals were markedly decreased, and levels of membrane lipid peroxidation were increased in Cu/Zn-SOD mutant mice compared to nontransgenic mice. These findings demonstrate that mutant Cu/Zn-SOD may endanger brain neurons by a mechanism involving impairment of glucose and glutamate transporters. Moreover, our data demonstrate a direct adverse effect of the mutant enzyme on synaptic function. PMID- 10724111 TI - Noninvasive functional imaging of human brain using light. AB - Analysis of photon transit time for low-power light passing into the head, and through both skull and brain, of human subjects allowed for tomographic imaging of cerebral hemoglobin oxygenation based on photon diffusion theory. In healthy adults, imaging of changes in hemoglobin saturation during hand movement revealed focal, contralateral increases in motor cortex oxygenation with spatial agreement to activation maps determined by functional magnetic resonance imaging; in ill neonates, imaging of hemoglobin saturation revealed focal regions of low oxygenation after acute stroke, with spatial overlap to injury location determined by computed tomography scan. Because such slow optical changes occur over seconds and co-localize with magnetic resonance imaging vascular signals whereas fast activation-related optical changes occur over milliseconds and co localize with EEG electrical signals, optical methods offer a single modality for exploring the spatio-temporal relationship between electrical and vascular responses in the brain in vivo, as well as for mapping cortical activation and oxygenation at the bedside in real-time for clinical monitoring. PMID- 10724112 TI - Neural substrate for the effects of passive training on sensorimotor cortical representation: a study with functional magnetic resonance imaging in healthy subjects. AB - Repetitive passive movements are part of most rehabilitation procedures, especially in patients with stroke and motor deficit. However, little is known about the consequences of repeated proprioceptive stimulations on the intracerebral sensorimotor network in humans. Twelve healthy subjects were enrolled, and all underwent two functional magnetic resonance imaging (fMRI) sessions separated by a 1-month interval. Passive daily movement training was performed in six subjects during the time between the two fMRI sessions. The other six subjects had no training and were considered as the control group. The task used during fMRI was calibrated repetitive passive flexion-extension of the wrist similar to those performed during training. The control task was rest. The data were analyzed with SPM96 software. Images were realigned, smoothed, and put into Talairach's neuroanatomical space. The time effect from the repetition of the task was assessed in the control group by comparing activation versus rest in the second session with activation versus rest in the first session. This time effect then was used as null hypothesis to assess the training effect alone in our trained group. Passive movements compared with rest showed activation of most of the cortical areas involved in motor control (i.e., contralateral primary sensorimotor cortex, supplementary motor area [SMA], cingulum, Brodmann area 40, ipsilateral cerebellum). Time effect comparison showed a decreased activity of the primary sensorimotor cortex and SMA and an increased activity of ipsilateral cerebellar hemisphere, compatible with a habituation effect. Training brought about an increased activity of contralateral primary sensorimotor cortex and SMA. A redistribution of SMA activity was observed. The authors demonstrated that passive training with repeated proprioceptive stimulation induces a reorganization of sensorimotor representation in healthy subjects. These changes take place in cortical areas involved in motor preparation and motor execution and represent the neural basis of proprioceptive training, which might benefit patients undergoing rehabilitative procedures. PMID- 10724113 TI - Dependence of oxygen delivery on blood flow in rat brain: a 7 tesla nuclear magnetic resonance study. AB - Magnetic resonance imaging (MRI) and spectroscopy (MRS) were used at a magnetic field strength of 7 T to measure CBF and CMRO2 in the sensorimotor cortex of mature rats at different levels of cortical activity. In rats maintained on morphine anesthesia, transitions to lower activity and higher activity states were produced by administration of pentobarbital and nicotine, respectively. Under basal conditions of morphine sulfate anesthesia, CBF was 0.75 +/- 0.09 mL x g(-1) x min(-1) and CMRO2 was 3.15 +/- 0.18 micromol x g(-1) x min(-1). Administration of sodium pentobarbital reduced CBF and CMRO2 by 66% +/- 16% and 61% +/- 6%, respectively (i.e., "deactivation"). In contrast, administration of nicotine hydrogen tartrate increased CBF and CMRO2 by 41% +/- 5% and 30% +/- 3%, respectively (i.e., "activation"). The resting values of CBF and CMRO2 for alpha chloralose anesthetized rats were 0.40 +/- 0.09 mL x g(-1) x min(-1) and 1.51 +/- 0.06 micromol x g(-1) x min(-1), respectively. Upon forepaw stimulation, CBF and CMRO2 were focally increased by 34% +/- 10% and 26% +/- 12%, respectively, above the resting nonanesthetized values (i.e., "activation"). Incremental changes in CBF and CMRO2, when expressed as a percentage change for "deactivation" and "activation" from the respective control conditions, were linear (R2 = 0.997) over the entire range examined with the global and local perturbations. This tight correlation for cerebral oxygen delivery in vivo is supported by a recent model where the consequence of a changing effective diffusivity of the capillary bed for oxygen, D, has been hypothetically shown to be linked to alterations in CMRO2 and CBF. This assumed functional characteristic of the capillary bed can be theoretically assessed by the ratio of fractional changes in D with respect to changes in CBF, signified by omega. A value 0.81 +/- 0.23 was calculated for omega with the in vivo data presented here, which in turn corresponds to a supposition that the effective oxygen diffusivity of the capillary bed is not constant but presumably varies to meet local requirements in oxygen demand in a similar manner with both "deactivation" and "activation." PMID- 10724114 TI - Administration of selective endothelin receptor type A antagonist Ro 61-1790 does not improve outcome in focal cerebral ischemia in cat. AB - The authors examined the effect of selective endothelin (ET) receptor type A (ET(A)) antagonism on histological and functional recovery in cat at 24 hours after reversible middle cerebral artery occlusion (MCAO). A novel and specific ET(A) antagonist, Ro 61-1790 [5-methylpyridine-2-sulfonic acid-6-(2 hydroxyethoxy)-5-(2-methoxyphenoxy)-2-(2-1H-tetrazol-5-y l-pyridin-4-yl) pyrimidin-4-ylamide sodium salt (1:2)] (Roche, Basel, Switzerland), was used at doses that produced steady-state plasma concentrations and abolished ET-induced pial arteriolar vasoconstriction. In a cranial window preparation, 8 nmol/L ET constricted pial arterioles by 33 +/- 18% (mean +/- SD), but this response was ablated by intravenous Ro 61-1790 treatment (10-mg/kg bolus, 4-mg/kg/h infusion). In additional animal cohorts, halothane-anesthetized cats were treated with 90 minutes of MCAO and 24 hours of reperfusion. Animals received Ro 61-1790 infusion beginning at the onset of reperfusion and continuing for 6 or 24 hours (n = 41). Control cats were treated with 0.9% saline by intravenous infusion throughout reperfusion. There was no difference in injury volume or neurologic evaluation score in saline-treated cats (n = 11; caudate 24 +/- 28%, cortical injury 7.5 +/- 5% of ipsilateral structure; score 52 +/- 8) versus the results in cats treated with Ro 61-1790 for either 24 hours (n = 6; caudate 22 +/- 23%, cortex 6 +/- 5%, injury volume of ipsilateral structure; score 55 +/- 3) or 6 hours (n = 11; caudate 33 +/- 30%, cortex 12 +/- 14%, injury volume of ipsilateral structure; score 50 +/- 10). Mortality was greatest in the 24-hour drug treatment group. These data suggest that blockade of ET(A) receptor activity is not beneficial to tissue or functional outcomes from experimental stroke in cat. PMID- 10724115 TI - Altered association of protein tyrosine kinases with postsynaptic densities after transient cerebral ischemia in the rat brain. AB - Transient cerebral ischemia results in an increase in the tyrosine phosphorylation of proteins associated with postsynaptic densities (PSDs). The authors investigated the possible mechanisms behind this increase by analyzing isolated PSDs for protein tyrosine kinase activity and for the presence of specific tyrosine kinases. Transient (15 minutes) global ischemia was produced in adult rats by four-vessel occlusion, and PSDs were isolated immediately after ischemia or after 20 minutes or 6 hours of reperfusion. Tyrosine phosphorylation of several PSD proteins, including the N-methyl-D-aspartate (NMDA) receptor subunits NR2A and NR2B, was enhanced relative to shams after 20 minutes of reperfusion and underwent a further increase between 20 minutes and 6 hours. The ability of intrinsic PSD tyrosine kinase to phosphorylate PSD proteins, including the NMDA receptor, increased threefold after ischemia. Whereas PSD-associated proline-rich tyrosine kinase 2 (PYK2) and gp145TrkB were elevated immediately after the ischemic event, increases in Src and Fyn were not apparent until 6 hours of reperfusion. The level of PSD-associated pp125FAK decreased after ischemia. The results demonstrate that ischemia results in selective changes in the association of protein tyrosine kinases with the PSD which may account for ischemia-induced increases in the tyrosine phosphorylation of PSD proteins. PMID- 10724116 TI - The window of opportunity for neuronal rescue with insulin-like growth factor-1 after hypoxia-ischemia in rats is critically modulated by cerebral temperature during recovery. AB - Insulin-like growth factor (IGF-1) is induced in damaged brain tissue after hypoxia-ischemia, and exogenous administration of IGF-1 shortly after injury has been shown to be neuroprotective. However, it is unknown whether treatment with IGF-1 delayed by more than a few hours after injury may be protective. Hypothermia after brain injury has been reported to delay the development of ischemic neuronal death. The authors therefore hypothesize that a reduction in the environmental temperature during recovery from hypoxia-ischemia could prolong the window of opportunity for IGF-1 treatment. Unilateral brain damage was induced in adult rats using a modified Levine model of right carotid artery ligation followed by brief hypoxia (6% O2 for 10 minutes). The rats were maintained in either a warm (31 degrees C) or cool (23 degrees C) environment for the first 2 hours after hypoxia. All rats were subsequently transferred to the 23 degrees C environment until the end of the experiment. A single dose of IGF-1 (50 microg) or its vehicle was given intracerebroventricularly at either 2 or 6 hours after hypoxia. Histologic outcome in the lateral cortex was quantified 5 days after hypoxia. Finally, cortical temperature was recorded from 1 hour before and 2 hours after hypoxia in separate groups of rats exposed to the "warm" and "cool" protocols. In rats exposed to the warm recovery environment, IGF-1 reduced cortical damage (P < 0.05) when given 2 hours but not 6 hours after insult. In contrast, with early recovery in the cool environment, a significant protective effect of IGF-1 in the lateral cortex (P < 0.05) was found with administration 6 hours after insult. In conclusion, a reduction in cerebral temperature during the early recovery phase after severe hypoxia-ischemia did not significantly reduce the severity of injury after 5 days' recovery; however, it markedly shifted and extended the window of opportunity for delayed treatment with IGF-1. PMID- 10724117 TI - Hypothermia during reperfusion after asphyxial cardiac arrest improves functional recovery and selectively alters stress-induced protein expression. AB - This study examined whether prolonged hypothermia induced 1 hour after resuscitation from asphyxial cardiac arrest would improve neurologic outcome and alter levels of stress-related proteins in rats. Rats were resuscitated from 8 minutes of asphyxia resulting in cardiac arrest. Brain temperature was regulated after resuscitation in three groups: normothermia (36.8 degrees C x 24 hours), immediate hypothermia (33 degrees C x 24 hours, beginning immediately after resuscitation), and delayed hypothermia (33 degrees C x 24 hours, beginning 60 minutes after resuscitation). Mortality and neurobehavioral deficits were improved in immediate and delayed hypothermia rats relative to normothermia rats. Furthermore, both immediate and delayed hypothermia improved neuronal survival in the CA1 region of the hippocampus assessed at 14 days. In normothermia rats, the 70-kDa heat shock protein (Hsp70) and 40-kDa heat shock protein (Hsp40) were increased within 12 hours after resuscitation in the hippocampus. Delayed hypothermia attenuated the increase in Hsp70 levels in the hippocampus but did not affect Hsp70 induction in the cerebellum. Hippocampal expression of Hsp40 was not affected by hypothermia. These data indicate that prolonged hypothermia during later reperfusion improves neurologic outcome after experimental global ischemia and is associated with selective changes in the pattern of stress induced protein expression. PMID- 10724118 TI - Importance of posttraumatic hypothermia and hyperthermia on the inflammatory response after fluid percussion brain injury: biochemical and immunocytochemical studies. AB - The purpose of this study was to investigate: 1) the temporal and regional profile of polymorphonuclear leukocyte (PMNL) infiltration after moderate traumatic brain injury using the parasagittal fluid percussion model and 2) the effects of posttraumatic hypothermia (30 degrees C) and hyperthermia (39 degrees C) on the acute and subacute inflammatory response. We hypothesized that posttraumatic hypothermia would reduce the degree of PMNL accumulation whereas hyperthermia would exacerbate this response to injury. In the first series of experiments we quantitated the temporal profile of altered myeloperoxidase activity under normothermic (37 degrees C) conditions (n = 20). The rats were allowed to survive for 3 hours, 24 hours, 3 days, or 7 days after trauma, and brains were dissected into cortical and subcortical regions ipsilateral and contralateral to injury. Additional animals were perfused and fixed for the immunocytochemical visualization of myeloperoxidase (n = 15). In the second series of experiments, rats (n = 25) were killed 3 hours or 3 days after the 3 hour monitoring period of normothermia (36.5 degrees C), hypothermia (30 degrees C), or hyperthermia (39 degrees C) (n = 4 to 5 per group), and myeloperoxidase activity was again quantitated. In normothermic rats, the enzymatic activity of myeloperoxidase was significantly increased (P < 0.05) at 3 hours within the anterior cortical segment (213.97 +/- 56.2 versus control 65.5 +/- 52.3 U/g of wet tissue; mean +/- SD) and posterior (injured) cortical and subcortical segments compared to sham-operated rats (305.76 +/- 27.8 and 258.67 +/- 101.4 U/g of wet tissue versus control 62.8 +/- 24.8 and 37.28 +/- 35.6 U/g of wet tissue; P < 0.0001, P < 0.05, respectively). At 24 hours and 7-days after trauma only the posterior cortical region (P < 0.005, P < 0.05, respectively) exhibited increased myeloperoxidase activity. However, 3 days after trauma, myeloperoxidase activity was also significantly increased within the anterior cortical segment (P < 0.05) and in posterior cortical and subcortical regions compared to sham-operated cortex (P < 0.0001, P < 0.05, respectively). Immunocytochemical analysis of myeloperoxidase reactivity at 3 hours, 24 hours, 3- and 7-days demonstrated large numbers of immunoreactive leukocytes within and associated with blood vessels, damaged tissues, and subarachnoid spaces. Posttraumatic hypothermia and hyperthermia had significant effects on myeloperoxidase activity at both 3 hours and 3 days after traumatic brain injury. Posttraumatic hypothermia reduced myeloperoxidase activity in the injured and noninjured cortical and subcortical segments compared to normothermic values (P < 0.05). In contrast, posttraumatic hyperthermia significantly elevated myeloperoxidase activity in the posterior cortical region compared to normothermic values at both 3 hours and 3 days (473.5 +/- 258.4 and 100.11 +/- 27.58 U/g of wet tissue, respectively, P < 0.05 versus controls). These results indicate that posttraumatic hypothermia decreases early and more prolonged myeloperoxidase activation whereas hyperthermia increases myeloperoxidase activity. Temperature-dependent alterations in PMNL accumulation appear to be a potential mechanism by which posttraumatic temperature manipulations may influence traumatic outcome. PMID- 10724119 TI - Rapid tau protein dephosphorylation and differential rephosphorylation during cardiac arrest-induced cerebral ischemia and reperfusion. AB - The effects of cerebral ischemia/reperfusion on phosphorylation of microtubule associated tau proteins were assessed in a canine model of cardiac arrest. As tau proteins are phosphorylated by kinases involved in different transduction signal pathways, their phosphorylation state is an excellent marker of neuronal homeostasis and microtubule dynamics. Canine brain tau proteins were characterized by immunoblotting using phosphorylation-dependent antibodies and antisera raised against different amino- and carboxy-terminal tau sequences. The present study reports a complete dephosphorylation of tau proteins during ischemia, which is shown by a higher electrophoretic mobility and the almost (if not total) disappearance of phosphorylation-dependent monoclonal antibody labeling. After 2-hour restoration of spontaneous circulation, a decrease in the electrophoretic mobility was observed, and after 24 hours of reperfusion, a full restoration of the phosphorylation was visualized using phosphorylation-dependent monoclonal antibodies directed against Ser/Thr-Pro sites. However, one particular phosphorylation site involved in tau binding to microtubules, located on Ser262/356, was never fully significantly rephosphorylated, suggesting that microtubule metabolism was still affected after 24 hours of reperfusion. Thus, the sequential and differential recovery of tau phosphorylation after ischemia followed by reperfusion is a useful marker with which to monitor neuronal integrity after brain ischemia. PMID- 10724120 TI - Novel characteristics of glutamate-induced cell death in primary septohippocampal cultures: relationship to calpain and caspase-3 protease activation. AB - Studies examined the phenotypic characteristics of glutamate-induced cell death and their relationship to calpain and caspase-3 activation. Cell viability was assessed by fluorescein diacetate and propidium iodide staining and lactate dehydrogenase release. Calpain and caspase-3 activity was inferred from signature proteolytic fragmentation of alpha-spectrin. Characterization of cell death phenotypes was assessed by Hoechst 33258 and DNA fragmentation assays. Exposure of septohippocampal cultures to 1.0, 2.0, and 4.0 mmol/L glutamate induced a dose dependent cell death with an LD50 of 2.0 mmol/L glutamate after 24 hours of incubation. Glutamate treatment induced cell death in neurons and astroglia and produced morphological alterations that differed from necrotic or apoptotic changes observed after maitotoxin or staurosporine exposure, respectively. After glutamate treatment, cell nuclei were enlarged and eccentrically shaped, and aggregated chromatin appeared in a diffusely speckled pattern. Furthermore, no dose of glutamate produced evidence of internucleosomal DNA fragmentation. Incubation with varying doses of glutamate produced calpain and caspase-3 activation. Calpain inhibitor II (N-acetyl-Leu-Leu-methionyl) provided protection only with a narrow dose range, whereas carbobenzoxy-Asp-CH2-OC(O)-2,6 dichlorobenzene (Z-D-DCB; pan-caspase inhibitor) and MK-801 (N-methyl-D-aspartate receptor antagonist) were potently effective across a wider dose range. Cycloheximide did not reduce cell death or protease activation. PMID- 10724121 TI - A temporal MRI assessment of neuropathology after transient middle cerebral artery occlusion in the rat: correlations with behavior. AB - The purpose of this study was to evaluate the temporal and spatial pathological alterations within ischemic tissue using serial magnetic resonance imaging (MRI) and to determine the extent and duration of functional impairment using objective behavioral tests after transient middle cerebral artery occlusion (tMCAO) in the rat. MRI signatures derived from specific anatomical regions of interest (ROI) were then appropriately correlated to the behavioral measures over the time course of the study (up to 28 days post-tMCAO). Sprague-Dawley rats (n = 12) were initially trained on the following behavioral tasks before surgery: bilateral sticky label test (for contralateral neglect); beam walking (for hindlimb coordination); staircase test (for skilled forelimb paw-reaching). Rats were then randomly assigned to receive either tMCAO (90 minutes, n = 6), by means of the intraluminal thread technique, or sham-control surgery (n = 6). Proton density, T2- and T2-diffusion-weighted MR images were acquired at 1, 7, 14, and 28 days post-tMCAO that were then smoothed into respective proton density, T2 relaxation, and apparent diffusion coefficient (ADC) maps. Apparent percent total lesion volume was assessed using T2W imaging. MR signatures were evaluated using the tissue maps by defining ROI for MCAO and sham-control groups, which corresponded to the caudate-putamen, forelimb, hindlimb, and lower parietal cortices both ipsilateral and contralateral to the occlusion site. Behavioral tests were undertaken daily from 1 to 28 days post-tMCAO. Results demonstrate that apparent percent lesion volume reduced from 1 to 7 days (P < 0.05) but then remained constant up to 28 days for the MCAO group. Pathological changes in the temporal profile of T2 and ADC tissue signatures were significantly altered in specific ROI across the time course of the study (P < 0.05 to <0.001), reflecting the progression of edema to necrosis and cavitation. Both T2 and ADC measures of ischemic pathology correlated with parameters defined by each of the functional tests (r > or =0.5, P < 0.05) across the time course. The staircase test revealed bilateral impairments for the MCAO group (P <0.001), which were best predicted by damage to the ipsilateral lower parietal cortex by means of hierarchical multiple regression analyses (R2 changes > or =0.21, P < or =0.03). Behavioral recovery was apparent on the beam walking test at 14 to 28 days post-MCAO, which was mirrored by MRI signatures within the hindlimb cortex returning to sham-control levels. This long-term study is the first of its kind in tracing the dynamic pathologic and functional consequences of tMCAO in the rat. Both serial MRI and objective behavioral assessment provide highly suitable outcome measures that can be effectively used to evaluate promising new antiischemic agents targeted for the clinic. PMID- 10724122 TI - Probability of metabolic tissue recovery after thrombolytic treatment of experimental stroke: a magnetic resonance spectroscopic imaging study in rat brain. AB - The effect of thrombolytic therapy on metabolic changes was studied in rats submitted to thromboembolic stroke. Reperfusion was initiated at three different time points, 1.5, 3, and 4.5 hours after embolism (n = 3 each), by injection of recombinant tissue-type plasminogen activator (rt-PA). Recovery was observed during 5 hours of reperfusion using perfusion-weighted images and a two dimensional 1H magnetic resonance spectroscopic imaging (MRSI) technique. Temporal evolution of the cerebral metabolites lactate and N-acetyl-aspartate (NAA) was determined. To analyze the chances of metabolic tissue recovery, the outcome of treatment, defined by a reversal of lactate concentration, was compared with the lactate intensity before treatment. In untreated animals (n = 4), clot embolism resulted in a drop of perfusion signal intensity in the occluded hemisphere followed by an increase of lactate concentration and a decrease of NAA that persisted throughout the observation period. Thrombolysis partially restored blood flow, but the mean lactate concentration decreased only slightly after successful lysis in animals treated 1.5 hours after embolism. If treatment was initiated later, no decline of lactate level was observed. Five hours after initiation of thrombolysis, the average tissue lactate amounted to 95 +/- 6, 111 +/- 17, and 139 +/- 60% of the early ischemic value (40 minutes after embolization) if treatment began 1.5, 3, and 4.5 hours after embolism, respectively. The NAA level declined slightly but never showed a recovery after rt-PA treatment. In individual pixels, the probability of metabolic tissue recovery clearly declined with increasing lactate concentration before thrombolysis. Interestingly, this probability was independent of treatment delay, but the number of pixels with low lactate declined with increasing ischemia time. Potential clinical applications of MRSI include monitoring of therapeutic intervention as well as support for prognosis of outcome after rt-PA treatment. PMID- 10724123 TI - Transcription factor nuclear factor-kappa B is activated in neurons after focal cerebral ischemia. AB - Nuclear factor-kappa B (NF-kappaB) is a multisubunit transcription factor that when activated induces the expression of genes encoding acute-phase proteins, cell adhesion molecules, cell surface receptors, and cytokines. NF-kappaB is composed of a variety of protein subunits of which p50-and p65-kDa (RelA) are the most widely studied. Under resting conditions, these subunits reside in the cytoplasm as an inactive complex bound by inhibitor proteins, IkappaB alpha and IkappaB beta. On activation, IkappaB is phosphorylated by IkappaB kinase and ubiquitinated and degraded by the proteasome; simultaneously, the active heterodimer translocates to the nucleus where it can initiate gene transcription. In the periphery, NF-kappaB is involved in inflammation through stimulation of the production of inflammatory mediators. The role of NF-kappaB in the brain is unclear. In vitro, NF-kappaB activation can be either protective or deleterious. The role of NF-kappaB in ischemic neuronal cell death in vivo was investigated. Adult male rats were subjected to 2 hours of focal ischemia induced by middle cerebral artery occlusion (MCAO). At 2, 6, and 12 hours after reperfusion, the expression and transactivation of NF-kappaB in ischemic versus nonischemic cortex and striatum were determined by immunocytochemistry and by electrophoretic mobility gel-shift analysis. At all time points studied, p50 and p65 immunoreactivity was found exclusively in the nuclei of cortical and striatal neurons in the ischemic hemisphere. The contralateral nonischemic hemisphere showed no evidence of nuclear NF-kappaB immunoreactivity. Double immunofluorescence confirmed expression of p50 in nuclei of neurons. Increased NF kappaB DNA-binding activity in nuclear extracts prepared from the ischemic hemisphere was further substantiated by electrophoretic mobility gel-shift analysis. Because the activation of NF-kappaB by many stimuli can be blocked by antioxidants in vitro, the effect of the antioxidant, LY341122, previously shown to be neuroprotective, on NF-kappaB activation in the MCAO model was evaluated. No significant activation of NF-kappaB was found by electrophoretic mobility gel shift analysis in animals treated with LY341122. These results demonstrate that transient focal cerebral ischemia results in activation of NF-kappaB in neurons and supports previous observations that neuroprotective antioxidants may inhibit neuronal death by preventing the activation of NF-kappaB. PMID- 10724124 TI - Acute decrease in cerebral nitric oxide levels after subarachnoid hemorrhage. AB - Disturbances in the nitric oxide (NO) vasodilatory pathway have been implicated in acute vasoconstriction and ischemia after subarachnoid hemorrhage (SAH). The authors hypothesize that blood released during SAH leads to vasoconstriction by scavenging NO and limiting its availability. This was tested by measuring the major NO metabolites nitrite and nitrate in five different brain regions before and after experimental SAH. The basal NO metabolites levels were as follows (mean +/- SD, micromol/mg wet weight): brain stem, 0.14 +/- 0.07; cerebellum, 0.12 +/- 0.08; ventral convexity cortex, 0.22 +/- 0.15; dorsal convexity cortex, 0.16 +/- 0.11; and hippocampus, 0.26 +/- 0.17. In sham-operated animals, no effect of the nitric oxide synthase (NOS) inhibitor L(G)-nitro-L-arginine-methyl-ester (30 mg/kg) was found on NO metabolites 40 minutes after administration, but a significant decrease was seen after 120 minutes. The NO metabolites decreased significantly 10 minutes after SAH in all brain regions except for hippocampus, and recovered to control levels in cerebellum at 60 minutes and in brain stem and dorsal cerebral cortex 180 minutes after SAH, while remaining low in ventral convexity cortex. Nitrite recovered completely in all brain regions at 180 minutes after SAH, whereas nitrate remained decreased in brain stem and ventral convexity cortex. Our results indicate that SAH causes acute decreases in cerebral NO levels by a mechanism other than NOS inhibition and provide further support for the hypothesis that alterations in the NO vasodilatory pathway contribute directly to the ischemic insult after SAH. PMID- 10724125 TI - Hypercarbia and mild hypothermia, only when not combined, improve postischemic bioenergetic recovery in neonatal rat brain slices. AB - In the immature brain, postischemic metabolism may be influenced beneficially by the effect of inducing hypercarbia or hypothermia. With use of 31P nuclear magnetic resonance spectroscopy, intracellular pH (pHi) and cellular energy metabolites in ex vivo neonatal rat cerebral cortex were measured before, during, and after substrate and oxygen deprivation in in vitro ischemia. Early postischemic hypothermia (fall in temperature -3.2 +/- 1.0 degrees C) delayed the normalization of pHi after ischemia by inducing an acid shift in pHi (P < 0.01). Postischemic hypercarbia (Krebs-Henseleit bicarbonate buffer equilibrated with 10% carbon dioxide in oxygen) and hypothermia induced separate, but potentially additive, reversible decreases in pHi, each of approximately -0.16 pH unit (P < 0.05). When these postischemic perturbations were applied in isolation, there was significant improvement of approximately 20% in the recovery of beta-ATP (P < 0.05). In combination, however, hypercarbia and hypothermia worsened recovery in ATP by approximately 20% (P < 0.05). In control tissue, which had not been exposed to ischemia, ATP content was also significantly reduced by co administration of the two treatments (P < 0.05), an effect that persisted even after discontinuing the perturbing conditions. Therefore, in this vascular independent neonatal preparation, early postischemic modulation of metabolism by hypercarbia or hypothermia appears to confer improved bioenergetic recovery, but only if they are not administered together. PMID- 10724126 TI - Overexpression of the cell death suppressor Bcl-w in ischemic brain: implications for a neuroprotective role via the mitochondrial pathway. AB - Bcl-w is a newly described cell death suppressor member of the Bcl-2 gene family. As these genes may have a role in the outcome of ischemic brain injury, the regional expression of Bcl-w protein in rat brain was examined at 6 to 72 hours after 90 minutes of transient middle cerebral artery occlusion. Bcl-w protein, although constitutively expressed at low levels in nonischemic brain, was found to be overexpressed in ischemic brain at all time points studied. Up-regulation of Bcl-w protein was particularly abundant in the penumbral region of the cortex and mainly in cells lacking DNA fragmentation. In the cortical penumbra, Bcl-w protein was detected predominantly in neurons and showed mitochondrial localization, as determined using double-label immunohistochemistry. Bcl-w expression was also detectable, to a lesser extent, in reactive astrocytes in the infarct border zone and in microvessel walls in the infarct regions. At the mechanistic level, incubation of isolated brain mitochondria with the addition of recombinant Bax or high concentration of calcium resulted in release of cytochrome c from the mitochondria. In the presence of recombinant Bcl-w protein, however, the release of cytochrome c induced by Bax or calcium was largely inhibited. Further, recombinant Bcl-w protein inhibited calcium-induced loss of mitochondrial transmembrane potential, indicative of permeability transition, in a dose-dependent manner. These results suggest that Bcl-w may be an endogenous neuroprotectant against ischemic neuronal death and that, like its analogues such as Bcl-2 and Bcl-x-long, Bcl-w may achieve this protection via the mitochondrial death-regulatory pathway. PMID- 10724127 TI - The efficacy of atypical antipsychotics in the treatment of depressive symptoms, hostility, and suicidality in patients with schizophrenia. AB - Depressive symptoms and syndromal depression commonly occur in patients with schizophrenia. Schizophrenia is also associated with aggression directed at self and others. For this article, the available literature regarding the efficacy of clozapine, risperidone, olanzapine, quetiapine, and ziprasidone in the treatment of depression, hostility, and suicidality in patients with schizophrenia was reviewed. These studies suggest that atypical antipsychotics may exert therapeutic effects on depression and hostility as well as psychosis and that clozapine and olanzapine may reduce suicidality in patients with schizophrenia. These therapeutic actions appear to represent additional advantages of atypical antipsychotics compared with standard agents. PMID- 10724128 TI - Hormonal side effects in women: typical versus atypical antipsychotic treatment. AB - Neuroleptic-induced hyperprolactinemia can cause menstrual disorders, impaired fertility, galactorrhea, and sexual dysfunction, as well as hypoestrogenism secondary to disruption of the hypothalamic-pituitary-ovarian axis. The development of the prolactin-sparing atypical antipsychotic drugs offers prevention and resolution of these adverse reactions. Thus far, this property of the new medications has received insufficient clinical attention. The authors use case vignettes to discuss assessment and management of clinical situations that arise as a result of antipsychotic-induced endocrine changes. PMID- 10724129 TI - Extrapyramidal side effects, tardive dyskinesia, and the concept of atypicality. AB - The most frequent problems associated with the older generation of antipsychotic agents are extrapyramidal side effects (EPS) and tardive dyskinesia. Neuroleptic induced EPS are thought to be caused by blockade of nigrostriatal dopamine tracts resulting in a relative increase in cholinergic activity; tardive dyskinesia is less well understood but is thought to be a supersensitivity response to chronic dopamine blockade. The leading hypothesis for the mechanism of action of the newer generation of atypical antipsychotics is the presence of a high serotonin to-dopamine receptor blockade ratio in the brain. When serotonergic activity is blocked-as is the case with atypical antipsychotics-dopamine release increases and balances out the dopamine blockade effect at postsynaptic receptor sites, which results in few or no EPS. Prospective data indicate that the risk of tardive dyskinesia in patients taking atypical antipsychotics is less than that for those taking typical antipsychotics. This article reviews the mechanisms of neuroleptic-induced EPS and tardive dyskinesia and discusses the relationship between these movement disorders and atypical antipsychotic agents. PMID- 10724130 TI - Recovery-oriented psychopharmacology: redefining the goals of antipsychotic treatment. AB - The traditional goals of psychopharmacology stem from the medical model. Rehabilitation interventions attempt to improve aspects of functioning in patients with chronic illnesses that are not responsive to biological intervention. Recovery is a concept emanating from the consumer self-help movement. It describes a move away from the patient role defined by a diagnostic label toward community membership defined by relationships and responsibilities in the community. Comprehensive care for people with psychotic disorders can include attention to each realm. This article provides an overview of the 3 models of care and describes a role for the psychopharmacologist in each as well as his or her unique potential to incorporate all 3. We outline potential synergistic benefits of integrating recovery-, rehabilitation-, and medical-model thinking into the practice of psychopharmacology and explore implications for the goals and outcomes of treatment for people with psychotic disorders. PMID- 10724132 TI - Re-reforming Unesco PMID- 10724131 TI - International panel for GM food? PMID- 10724133 TI - Genome leaders told to keep their eyes on the main prize. PMID- 10724134 TI - GM debate must go global, says meeting... PMID- 10724135 TI - Unesco 'worse than I imagined,' says new director PMID- 10724136 TI - Expensive space crystal programme has produced little of scientific value, says panel. PMID- 10724137 TI - Geneticists oppose consent ruling. PMID- 10724138 TI - Panel will seek 'appropriate' AIDS goals for South Africa. PMID- 10724139 TI - $350m gift boosts MIT brain power. PMID- 10724140 TI - Indian research budget favours defence. PMID- 10724141 TI - Rival demands sink genome alliance plans. PMID- 10724142 TI - Why private institutions alone will not do enough to protect biodiversity. PMID- 10724143 TI - Alzheimer's research is vital in work on ageing. PMID- 10724144 TI - Opportunism knocks? PMID- 10724145 TI - A century of cognitive decline. PMID- 10724146 TI - The song of the Neanderthal PMID- 10724147 TI - Life's downs and ups. PMID- 10724148 TI - Quantum cloning. Schrodinger's sheep. PMID- 10724149 TI - Neurobiology. A chorus line. PMID- 10724150 TI - Too hot to melt PMID- 10724151 TI - Cell biology. Moving in mysterious ways. PMID- 10724152 TI - Pushing electrons around PMID- 10724153 TI - Plate tectonics. The Antarctic connection PMID- 10724154 TI - Urban benzene and population exposure. PMID- 10724155 TI - Climate variability and North Sea cod. PMID- 10724156 TI - Bleaching patterns in reef corals. PMID- 10724157 TI - Myopia and ambient night-time lighting. CLEERE Study Group. Collaborative Longitudinal Evaluation of Ethnicity and Refractive Error. PMID- 10724158 TI - Myopia and ambient night-time lighting. PMID- 10724159 TI - Cenozoic motion between East and West Antarctica AB - The West Antarctic rift system is the result of late Mesozoic and Cenozoic extension between East and West Antarctica, and represents one of the largest active continental rift systems on Earth. But the timing and magnitude of the plate motions leading to the development of this rift system remain poorly known, because of a lack of magnetic anomaly and fracture zone constraints on seafloor spreading. Here we report on magnetic data, gravity data and swath bathymetry collected in several areas of the south Tasman Sea and northern Ross Sea. These results enable us to calculate mid-Cenozoic rotation parameters for East and West Antarctica. These rotations show that there was roughly 180 km of separation in the western Ross Sea embayment in Eocene and Oligocene time. This episode of extension provides a tectonic setting for several significant Cenozoic tectonic events in the Ross Sea embayment including the uplift of the Transantarctic Mountains and the deposition of large thicknesses of Oligocene sediments. Inclusion of this East-West Antarctic motion in the plate circuit linking the Australia, Antarctic and Pacific plates removes a puzzling gap between the Lord Howe rise and Campbell plateau found in previous early Tertiary reconstructions of the New Zealand region. Determination of this East-West Antarctic motion also resolves a long standing controversy regarding the contribution of deformation in this region to the global plate circuit linking the Pacific to the rest of the world. PMID- 10724160 TI - Autoinhibition and activation mechanisms of the Wiskott-Aldrich syndrome protein. AB - The Rho-family GTPase, Cdc42, can regulate the actin cytoskeleton through activation of Wiskott-Aldrich syndrome protein (WASP) family members. Activation relieves an autoinhibitory contact between the GTPase-binding domain and the carboxy-terminal region of WASP proteins. Here we report the autoinhibited structure of the GTPase-binding domain of WASP, which can be induced by the C terminal region or by organic co-solvents. In the autoinhibited complex, intramolecular interactions with the GTPase-binding domain occlude residues of the C terminus that regulate the Arp2/3 actin-nucleating complex. Binding of Cdc42 to the GTPase-binding domain causes a dramatic conformational change, resulting in disruption of the hydrophobic core and release of the C terminus, enabling its interaction with the actin regulatory machinery. These data show that 'intrinsically unstructured' peptides such as the GTPase-binding domain of WASP can be induced into distinct structural and functional states depending on context. PMID- 10724161 TI - Discovery of calcium in Mercury's atmosphere. AB - The composition and evolutionary history of Mercury's crust are not well determined. The planet as a whole has been predicted to have a refractory, anhydrous composition: rich in Ca, Al, Mg and Fe, but poor in Na, K, OH, and S. Its atmosphere is believed to be derived in large part from the surface materials. A combination of effects that include impact vaporization (from infalling material), volatile evaporation, photon-stimulated desorption and sputtering releases material from the surface to form the atmosphere. Sodium and potassium have already been observed in Mercury's atmosphere, with abundances that require a volatile-rich crust. The sodium probably results from photon stimulated desorption, and has a temperature of 1,500 K (ref. 10). Here we report the discovery of calcium in the atmosphere near Mercury's poles. The column density is very low and the temperature is apparently very high (12,000 K). The localized distribution and high temperature, if confirmed, suggest that the atmospheric calcium may arise from surface sputtering by ions, which enter Mercury's auroral zone. The low abundance of atmospheric Ca may indicate that the regolith is rarefied in calcium. PMID- 10724163 TI - Impossibility of deleting an unknown quantum state AB - A photon in an arbitrary polarization state cannot be cloned perfectly. But suppose that at our disposal we have several copies of a photon in an unknown state. Is it possible to delete the information content of one or more of these photons by a physical process? Specifically, if two photons are in the same initial polarization state, is there a mechanism that produces one photon in the same initial state and the other in some standard polarization state? If this could be done, then one would create a standard blank state onto which one could copy an unknown state approximately, by deterministic cloning or exactly, by probabilistic cloning. This could in principle be useful in quantum computation, where one could store new information in an already computed state by deleting the old information. Here we show, however, that the linearity of quantum theory does not allow us to delete a copy of an arbitrary quantum state perfectly. Though in a classical computer information can be deleted (reversibly) against a copy, the analogous task cannot be accomplished, even irreversibly, with quantum information. PMID- 10724162 TI - North-south geological differences between the residual polar caps on Mars. AB - Polar processes can be sensitive indicators of global climate, and the geological features associated with polar ice caps can therefore indicate evolution of climate with time. The polar regions on Mars have distinctive morphologic and climatologic features: thick layered deposits, seasonal CO2 frost caps extending to mid latitudes, and near-polar residual frost deposits that survive the summer. The relationship of the seasonal and residual frost caps to the layered deposits has been poorly constrained, mainly by the limited spatial resolution of the available data. In particular, it has not been known if the residual caps represent simple thin frost cover or substantial geologic features. Here we show that the residual cap on the south pole is a distinct geologic unit with striking collapse and erosional topography; this is very different from the residual cap on the north pole, which grades into the underlying layered materials. These findings indicate that the differences between the caps are substantial (rather than reflecting short-lived differences in frost cover), and so support the idea of long-term asymmetry in the polar climates of Mars. PMID- 10724164 TI - Molecular control over Au/GaAs diodes AB - The use of molecules to control electron transport is an interesting possibility, not least because of the anticipated role of molecules in future electronic devices. But physical implementations using discrete molecules are neither conceptually simple nor technically straightforward (difficulties arise in connecting the molecules to the macroscopic environment). But the use of molecules in electronic devices is not limited to single molecules, molecular wires or bulk material. Here we demonstrate that molecules can control the electrical characteristics of conventional metal-semiconductor junctions, apparently without the need for electrons to be transferred onto and through the molecules. We modify diodes by adsorbing small molecules onto single crystals of n-type GaAs semiconductor. Gold contacts were deposited onto the modified surface, using a 'soft' method to avoid damaging the molecules. By using a series of multifunctional molecules whose dipole is varied systematically, we produce diodes with an effective barrier height that is tuned by the molecule's dipole moment. These barrier heights correlate well with the change in work function of the GaAs surface after molecular modification. This behaviour is consistent with that of unmodified metal-semiconductor diodes, in which the barrier height can depend on the metal's work function. PMID- 10724165 TI - Sintering dense nanocrystalline ceramics without final-stage grain growth AB - Sintering is the process whereby interparticle pores in a granular material are eliminated by atomic diffusion driven by capillary forces. It is the preferred manufacturing method for industrial ceramics. The observation of Burke and Coble that certain crystalline granular solids could gain full density and translucency by solid-state sintering was an important milestone for modern technical ceramics. But these final-stage sintering processes are always accompanied by rapid grain growth, because the capillary driving forces for sintering (involving surfaces) and grain growth (involving grain boundaries) are comparable in magnitude, both being proportional to the reciprocal grain size. This has greatly hampered efforts to produce dense materials with nanometre-scale structure (grain size less than 100 nm), leading many researchers to resort to the 'brute force' approach of high-pressure consolidation at elevated temperatures. Here we show that fully dense cubic Y2O3 (melting point, 2,439 degrees C) with a grain size of 60 nm can be prepared by a simple two-step sintering method, at temperatures of about 1,000 degrees C without applied pressure. The suppression of the final stage grain growth is achieved by exploiting the difference in kinetics between grain-boundary diffusion and grain-boundary migration. Such a process should facilitate the cost-effective preparation of other nanocrystalline materials for practical applications. PMID- 10724166 TI - The influence of Antarctic sea ice on glacial-interglacial CO2 variations AB - Ice-core measurements indicate that atmospheric CO2 concentrations during glacial periods were consistently about 80 parts per million lower than during interglacial periods. Previous explanations for this observation have typically had difficulty accounting for either the estimated glacial O2 concentrations in the deep sea, 13C/12C ratios in Antarctic surface waters, or the depth of calcite saturation; also lacking is an explanation for the strong link between atmospheric CO2 and Antarctic air temperature. There is growing evidence that the amount of deep water upwelling at low latitudes is significantly overestimated in most ocean general circulation models and simpler box models previously used to investigate this problem. Here we use a box model with deep-water upwelling confined to south of 55 degrees S to investigate the glacial-interglacial linkages between Antarctic air temperature and atmospheric CO2 variations. We suggest that low glacial atmospheric CO2 levels might result from reduced deep water ventilation associated with either year-round Antarctic sea-ice coverage, or wintertime coverage combined with ice-induced stratification during the summer. The model presented here reproduces 67 parts per million of the observed glacial-interglacial CO2 difference, as a result of reduced air-sea gas exchange in the Antarctic region, and is generally consistent with the additional observational constraints. PMID- 10724167 TI - Interferometric radar measurements of water level changes on the Amazon flood plain AB - Measurements of water levels in the main channels of rivers, upland tributaries and floodplain lakes are necessary for understanding flooding hazards, methane production, sediment transport and nutrient exchange. But most remote river basins have only a few gauging stations and these tend to be restricted to large river channels. Although radar remote sensing techniques using interferometric phase measurements have the potential to greatly improve spatial sampling, the phase is temporally incoherent over open water and has therefore not been used to determine water levels. Here we use interferometric synthetic aperture radar (SAR) data, acquired over the central Amazon by the Space Shuttle imaging radar mission, to measure subtle water level changes in an area of flooded vegetation on the Amazon flood plain. The technique makes use of the fact that flooded forests and floodplain lakes with emergent shrubs permit radar double-bounce returns from water and vegetation surfaces, thus allowing coherence to be maintained. Our interferometric phase observations show decreases in water levels of 7-11 cm per day for tributaries and lakes within approximately 20 km of a main channel and 2-5 cm per day at distances of approximately 80 km. Proximal floodplain observations are in close agreement with main-channel gauge records, indicating a rapid response of the flood plain to decreases in river stage. With additional data from future satellite missions, the technique described here should provide direct observations important for understanding flood dynamics and hydrologic exchange between rivers and flood plains. PMID- 10724168 TI - Delayed biological recovery from extinctions throughout the fossil record. AB - How quickly does biodiversity rebound after extinctions? Palaeobiologists have examined the temporal, taxonomic and geographic patterns of recovery following individual mass extinctions in detail, but have not analysed recoveries from extinctions throughout the fossil record as a whole. Here, we measure how fast biodiversity rebounds after extinctions in general, rather than after individual mass extinctions, by calculating the cross-correlation between extinction and origination rates across the entire Phanerozoic marine fossil record. Our results show that extinction rates are not significantly correlated with contemporaneous origination rates, but instead are correlated with origination rates roughly 10 million years later. This lagged correlation persists when we remove the 'Big Five' major mass extinctions, indicating that recovery times following mass extinctions and background extinctions are similar. Our results suggest that there are intrinsic limits to how quickly global biodiversity can recover after extinction events, regardless of their magnitude. They also imply that today's anthropogenic extinctions will diminish biodiversity for millions of years to come. PMID- 10724169 TI - Simple rules yield complex food webs. AB - Several of the most ambitious theories in ecology describe food webs that document the structure of strong and weak trophic links that is responsible for ecological dynamics among diverse assemblages of species. Early mechanism-based theory asserted that food webs have little omnivory and several properties that are independent of species richness. This theory was overturned by empirical studies that found food webs to be much more complex, but these studies did not provide mechanistic explanations for the complexity. Here we show that a remarkably simple model fills this scientific void by successfully predicting key structural properties of the most complex and comprehensive food webs in the primary literature. These properties include the fractions of species at top, intermediate and basal trophic levels, the means and variabilities of generality, vulnerability and food-chain length, and the degrees of cannibalism, omnivory, looping and trophic similarity. Using only two empirical parameters, species number and connectance, our 'niche model' extends the existing 'cascade model and improves its fit ten-fold by constraining species to consume a contiguous sequence of prey in a one-dimensional trophic niche. PMID- 10724170 TI - Inhibitory threshold for critical-period activation in primary visual cortex. AB - Neuronal circuits across several systems display remarkable plasticity to sensory input during postnatal development. Experience-dependent refinements are often restricted to well-defined critical periods in early life, but how these are established remains mostly unknown. A representative example is the loss of responsiveness in neocortex to an eye deprived of vision. Here we show that the potential for plasticity is retained throughout life until an inhibitory threshold is attained. In mice of all ages lacking an isoform of GABA (gamma aminobutyric acid) synthetic enzyme (GAD65), as well as in immature wild-type animals before the onset of their natural critical period, benzodiazepines selectively reduced a prolonged discharge phenotype to unmask plasticity. Enhancing GABA-mediated transmission early in life rendered mutant animals insensitive to monocular deprivation as adults, similar to normal wild-type mice. Short-term presynaptic dynamics reflected a synaptic reorganization in GAD65 knockout mice after chronic diazepam treatment. A threshold level of inhibition within the visual cortex may thus trigger, once in life, an experience-dependent critical period for circuit consolidation, which may otherwise lie dormant. PMID- 10724171 TI - Attention modulates synchronized neuronal firing in primate somatosensory cortex. AB - A potentially powerful information processing strategy in the brain is to take advantage of the temporal structure of neuronal spike trains. An increase in synchrony within the neural representation of an object or location increases the efficacy of that neural representation at the next synaptic stage in the brain; thus, increasing synchrony is a candidate for the neural correlate of attentional selection. We investigated the synchronous firing of pairs of neurons in the secondary somatosensory cortex (SII) of three monkeys trained to switch attention between a visual task and a tactile discrimination task. We found that most neuron pairs in SII cortex fired synchronously and, furthermore, that the degree of synchrony was affected by the monkey's attentional state. In the monkey performing the most difficult task, 35% of neuron pairs that fired synchronously changed their degree of synchrony when the monkey switched attention between the tactile and visual tasks. Synchrony increased in 80% and decreased in 20% of neuron pairs affected by attention. PMID- 10724172 TI - Growth patterns in the developing brain detected by using continuum mechanical tensor maps. AB - The dynamic nature of growth and degenerative disease processes requires the design of sensitive strategies to detect, track and quantify structural change in the brain in its full spatial and temporal complexity. Although volumes of brain substructures are known to change during development, detailed maps of these dynamic growth processes have been unavailable. Here we report the creation of spatially complex, four-dimensional quantitative maps of growth patterns in the developing human brain, detected using a tensor mapping strategy with greater spatial detail and sensitivity than previously obtainable. By repeatedly scanning children (aged 3-15 years) across time spans of up to four years, a rostro-caudal wave of growth was detected at the corpus callosum, a fibre system that relays information between brain hemispheres. Peak growth rates, in fibres innervating association and language cortices, were attenuated after puberty, and contrasted sharply with a severe, spatially localized loss of subcortical grey matter. Conversely, at ages 3-6 years, the fastest growth rates occurred in frontal networks that regulate the planning of new actions. Local rates, profiles, and principal directions of growth were visualized in each individual child. PMID- 10724173 TI - A clonogenic common myeloid progenitor that gives rise to all myeloid lineages. AB - Haematopoietic stem cells give rise to progeny that progressively lose self renewal capacity and become restricted to one lineage. The points at which haematopoietic stem cell-derived progenitors commit to each of the various lineages remain mostly unknown. We have identified a clonogenic common lymphoid progenitor that can differentiate into T, B and natural killer cells but not myeloid cells. Here we report the prospective identification, purification and characterization, using cell-surface markers and flow cytometry, of a complementary clonogenic common myeloid progenitor that gives rise to all myeloid lineages. Common myeloid progenitors give rise to either megakaryocyte/erythrocyte or granulocyte/macrophage progenitors. Purified progenitors were used to provide a first-pass expression profile of various haematopoiesis-related genes. We propose that the common lymphoid progenitor and common myeloid progenitor populations reflect the earliest branch points between the lymphoid and myeloid lineages, and that the commitment of common myeloid progenitors to either the megakaryocyte/erythrocyte or the granulocyte/macrophage lineages are mutually exclusive events. PMID- 10724174 TI - Regulation of intracellular calcium by a signalling complex of IRAG, IP3 receptor and cGMP kinase Ibeta. AB - Calcium release from the endoplasmic reticulum controls a number of cellular processes, including proliferation and contraction of smooth muscle and other cells. Calcium release from inositol 1,4,5-trisphosphate (IP3)-sensitive stores is negatively regulated by binding of calmodulin to the IP3 receptor (IP3R) and the NO/cGMP/cGMP kinase I (cGKI) signalling pathway. Activation of cGKI decreases IP3-stimulated elevations in intracellular calcium, induces smooth muscle relaxation and contributes to the antiproliferative and pro-apoptotic effects of NO/cGMP. Here we show that, in microsomal smooth muscle membranes, cGKIbeta phosphorylated the IP3R and cGKIbeta, and a protein of relative molecular mass 125,000 which we now identify as the IP3R-associated cGMP kinase substrate (IRAG). These proteins were co-immunoprecipitated by antibodies directed against cGKI, IP3R or IRAG. IRAG was found in many tissues including aorta, trachea and uterus, and was localized perinuclearly after heterologous expression in COS-7 cells. Bradykinin-stimulated calcium release was not affected by the expression of either IRAG or cGKIbeta, which we tested in the absence and presence of cGMP. However, calcium release was inhibited after co-expression of IRAG and cGKIbeta in the presence of cGMP. These results identify IRAG as an essential NO/cGKI dependent regulator of IP3-induced calcium release. PMID- 10724175 TI - hCds1-mediated phosphorylation of BRCA1 regulates the DNA damage response. AB - Mutations in the BRCA1 (ref. 1) tumour suppressor gene are found in almost all of the families with inherited breast and ovarian cancers and about half of the families with only breast cancer. Although the biochemical function of BRCA1 is not well understood, it is important for DNA damage repair and cell-cycle checkpoint. BRCA1 exists in nuclear foci but is hyperphosphorylated and disperses after DNA damage. It is not known whether BRCA1 phosphorylation and dispersion and its function in DNA damage response are related. In yeast the DNA damage response and the replication-block checkpoint are mediated partly through the Cds1 kinase family. Here we report that the human Cds1 kinase (hCds1/Chk2) regulates BRCA1 function after DNA damage by phosphorylating serine 988 of BRCA1. We show that hCds1 and BRCA1 interact and co-localize within discrete nuclear foci but separate after gamma irradiation. Phosphorylation of BRCA1 at serine 988 is required for the release of BRCA1 from hCds1. This phosphorylation is also important for the ability of BRCA1 to restore survival after DNA damage in the BRCA1-mutated cell line HCC1937. PMID- 10724176 TI - Ligand binding and conformational motions in myoglobin. AB - Myoglobin, a small globular haem protein that binds gaseous ligands such as O2, CO and NO reversibly at the haem iron, serves as a model for studying structural and dynamic aspects of protein reactions. Time-resolved spectroscopic measurements after photodissociation of the ligand revealed a complex ligand binding reaction with multiple kinetic intermediates, resulting from protein relaxation and movements of the ligand within the protein. To observe the structural changes induced by ligand dissociation, we have carried out X-ray crystallographic investigations of carbon monoxy-myoglobin (MbCO mutant L29W) crystals illuminated below and above 180 K, complemented by time-resolved infrared spectroscopy of CO rebinding. Here we show that below 180 K photodissociated ligands migrate to specific sites within an internal cavity--the distal haem pocket--of an essentially immobilized, frozen protein, from where they subsequently rebind by thermally activated barrier crossing. Upon photodissociation above 180 K, ligands escape from the distal pocket, aided by protein fluctuations that transiently open exit channels. We recover most of the ligands in a cavity on the opposite side of the haem group. PMID- 10724177 TI - Purification and characterization of UDP-glucuronate: baicalein 7-O glucuronosyltransferase from Scutellaria baicalensis Georgi. cell suspension cultures. AB - UDP-glucuronate: baicalein 7-O-glucuronosyltransferase (UBGAT) catalyzes the transfer of glucuronic acid from UDP-glucuronic acid to the 7-OH of baicalein. UBGAT was purified from cultured cells of Scutellaria baicalensis Georgi (Lamiaceae). It was purified 95-fold using various chromatography and chromatofocusing procedures to apparent homogeneity. The Mr was estimated to be 110 kDa by gel filtration chromatography with a 52 kDa subunit by SDS-PAGE. The isoelectric point was pH 4.8. UBGAT was specific to UDP-glucuronic acid as a sugar donor and flavones with substitution ortho- to the 7-OH group such its baicalein (6-OH), scutellarein (6-OH) and wogonin (8-OMe). PMID- 10724178 TI - Ontogenic changes in enzymes of carbon metabolism in relation to carbohydrate status in developing mungbean reproductive structures. AB - The content of free sugars and the activities of enzymes involved in carbon metabolism-sucrose synthase, acid and alkaline invertase, phosphoenol pyruvate carboxylase, malic enzyme and isocitrate dehydrogenase were determined during seed development in mungbean pods. A decrease in carbohydrate content of pod wall from 10 to 25 days after flowering (DAF) and a concomitant increase in the seed till 20 DAF was observed. Sucrose remained the dominant soluble sugar in the pod wall and seed. In the branch of inflorescence and pod wall, the activities of sucrose metabolizing enzymes, viz. acid and alkaline invertase, sucrose synthase (synthesis and cleavage) and sucrose phosphate synthase were higher at 5-10 DAF, whereas in seed the maximum activities of these enzymes were observed at the time of maximum seed filling stage (10-20 DAF). High activities of sucrose synthase at the time of rapid seed filling can be correlated to its sink strength. Higher activities of phosphoenol pyruvate carboxylase in the branch of inflorescence and pod wall than in seed may indicate the involvement of the fruiting structure for recapturing respired CO2. High activities of isocitrate dehydrogenase and malic enzyme in the seed at the time of rapid seed filling could provide NADPH and carbon skeletons required for the synthesis of various seed reserves. PMID- 10724179 TI - Biosynthesis of brassinosteroids in cultured cells of Catharanthus roseus. AB - Precursor administration experiments with 2H-labeled 6-oxocampestanol, 6 deoxocastasterone and 6alpha-hydroxycastasterone in cultured cells of Catharanthus roseus were performed and the metabolites were analyzed by GC-MS. [2H6]Cathasterone was identified as a metabolite of [2H6]6-oxocampestanol, whereas [2H6]6alpha-hydroxycastasterone and [2H6]castasterone were identified as metabolites of [2H6]6-deoxocastasterone, and [2H6]castasterone was identified as a metabolite of [2H6]6alpha-hydroxycastasterone, indicating that 6 deoxocastasterone is converted to castasterone via 6alpha-hydroxycastasterone. In addition, 6-deoxocathasterone, a putative biosynthetic intermediate in the late C6-oxidation pathway, was identified as an endogenous brassinosteroid. These studies provide further evidence supporting our proposed biosynthetic pathways for brassinolide. PMID- 10724180 TI - Distribution of morphinan and benzo[c]phenanthridine alkaloid gene transcript accumulation in Papaver somninferum. AB - The opium poppy Papaver somniferum L. produces the antimicrobial benzo[c]phenanthridine alkaloid sanguinarine and the narcotic analgesic morphinan alkaloid morphine. Transcripts of three genes of alkaloid biosynthesis in P. somniferum in developing seedlings, mature plants and plant cell suspension culture were monitored for temporal/spatial or for methyl jasmonate-induced accumulation by RNA gel blot analysis. These genes encoded (S)-N-methylcoclaurine 3'-hydroxylase (CYP80B1) that is common to morphine and sanguinarine biosynthesis, the berberine bridge enzyme (BBE) that lies on the pathway to sanguinarine, and codeinone reductase (COR) the penultimate enzyme of morphine biosynthesis. In developing P. somniferum seedlings, the morphine precursor thebaine was present throughout the first twenty days of germination. In contrast, sanguinarine was present in detectable quantities only after day five after germination and continued to increase at least until day twenty. Accumulation of cyp80b1, bbe1 and cor1 gene transcripts paralleled these differences. In the mature poppy plant, cyp80b1, bbe1 and cor1 gene transcripts were detected in the root, the stem, the leaf lamina and the leaf mid rib. Only cyp80b1 and cor1, however, were found in the flower bud and the capsule. Consistent with the fact that sanguinarine accumulation, but not that of morphine, can be induced in opium poppy cell suspension culture by addition of methyl jasmonate to the culture medium, cyp80b1 and bbe1, but not cor1 transcript accumulated in response to elicitor treatment. PMID- 10724181 TI - Scission of polysaccharides by peroxidase-generated hydroxyl radicals. AB - Cell-wall polysaccharides can be broken down non-enzymatically in vitro by scission of backbone bonds in a Fenton reaction system producing hydroxyl radicals (OH*) (Fry, S.C. (1998). Biochemical Journal, 332, 507-515). OH* can also be generated enzymatically from O2 by horseradish peroxidase (HRP) in a complex reaction cycle involving NADH or dihydroxyfumarate (DHF) as reducing substrate (Chen, S.-X., & Schopfer, P. (1999). European Journal of Biochemistry, 260, 726-735). Based on these recent findings the possibility that HRP can be used to degrade cell-wall polysaccharides in vitro was investigated. The production of OH* from O2 by HRP in the presence of NADH or DHF was confirmed by EPR spectroscopy using 5,5-dimethyl-1-pyrroline-N-oxide as a spin trap. Chemical scission of polysaccharides (dextran, pectin, xyloglucan) by HRP-generated OH* was demonstrated using a viscometric assay. The reaction could be inhibited by an array of OH* scavengers, confirming the involvement OH* as the causative agent for macromolecule cleavage. The significance of these findings for the biochemical function of peroxidase in cell-wall loosening processes underlying cell expansion and related physiological processes is discussed. PMID- 10724182 TI - Acetylenic aromatic compounds from Stereum hirsutum. AB - Stereum hirsutum is a one of several fungi involved in a grapevine disease called esca. From the culture medium of this fungus, four new acetylenic compounds 1-3 and 6 have been isolated and identified. Structural elucidation and biological activity are reported. PMID- 10724183 TI - Covalently linked anthocyanin-flavonol pigments from blue Agapanthus flowers. AB - The structures of the two major anthocyanins in blue Agapanthus flowers have been determined to be a p-coumaroylated delphinidin diglycoside attached to a flavonol triglycoside via a succinic acid diester link. The structure has been determined unambiguously through degradation studies, glycosidic analysis and NMR experiments. These compounds represent unique examples of anthocyanin pigments where both types of co-pigment, an aromatic acyl group and a flavonoid co pigment, are attached covalently to the anthocyanin. PMID- 10724184 TI - An amendment of Aspergillus section Candidi based on chemotaxonomical evidence. AB - A novel 2,2'-epoxy-terphenyllin, candidusin C, in addition to the well known secondary metabolites terphenyllin, 3-hydroxyterpenyllin and chlorflavonin, has been isolated from the chemically unexplored fungus Aspergillus campestris. The latter three are known secondary metabolites from Aspergillus candidus and therefore a large number of Aspergilli were screened for production of these compounds to see whether they could be regarded as chemotaxonomical indicators of section membership in the monotypic Aspergillus section Candidi. The results indicated that A. campestris and A. taichungensis should be placed in Candidi and this was further confirmed by morphological and physiological similarities. Three species outside the section Candidi produced candidusin related secondary metabolites: Aspergillus arenarius, A. ellipticus and Penicillium raistrickii. Chlorflavonin, however, was only found in section Candidi. PMID- 10724185 TI - Bioactive alkaloids from Brunsvigia radulosa. AB - A phytochemical investigation of the bulbs of Brunsvigia radulosa yielded the new alkaloid 1-O-acetylnorpluviine, together with the known structures 1 epideacetylbowdensine, crinamine, crinine, hamayne, lycorine, anhydrolycorin-6 one and sternbergine. All structures were established by spectroscopic evidence. Some of the 13C assignments which were reported for crinamine and hamayne were corrected by means of 2D NMR techniques. In order to provide a further structure for biological testing, crinamine was converted to apohaemanthamine. The alkaloids were tested for activity against two strains of cultured Plasmodium falciparum and for cytotoxicity with BL6 mouse melanoma cells. PMID- 10724186 TI - Pinguisane and dimeric pinguisane-type sesquiterpenoids from the Japanese liverwort Porella acutifolia subsp. tosana. AB - Two new pinguisane-type, three new Diels-Alder reaction-type dimeric pinguisane sesquiterpenoids and known sesqui and diterpenoids were isolated from the ether extract of the Japanese liverwort Porella acutifolia subsp. tosano. Their absolute stereostructures were established by a combination of extensive 2D-NMR, CD spectra, X-ray crystallographic analysis, modified Mosher's method and chemical correlation. PMID- 10724187 TI - An isoflavone from Pterocarpus santalinus. AB - A new isoflavone together with liquiritigenin and isoliquiritigenin has been isolated from the heartwood of Pterocarpus santalinus. Based on spectral methods, the structure of the new compound was elucidated as 6-hydroxy,7,2',4',5' tetramethoxyisoflavone. PMID- 10724188 TI - Configurations of polyunsaturated sesterterpenoids from the diatom, Haslea ostrearia. AB - The partial configurations of C25 isoprenoid alkenes isolated from the diatom Haslea ostrearia Gaillon (Simonsen) have been established. A combination of NMR spectroscopy studies of the alkenes with chiral shift reagents in conjunction with soluble silver beta-diketonate complexes and enantioselective gas chromatography of oxidation products of the alkenes was used. Unexpected differences in highly branched isoprenoid isomer configurations were observed between different laboratory cultures of the alga. PMID- 10724189 TI - Antifungal and larvicidal cordiaquinones from the roots of Cordia curassavica. AB - In addition to the known cordiaquinones A and B, two novel meroterpenoid naphthoquinones, named cordiaquinones J and K, have been isolated from the roots of Cordia curassavica. Their structures were elucidated by spectrometric methods including EI, D/CI mass spectrometry, 1H, 13C and 2D-NMR experiments. The four naphthoquinones demonstrated antifungal activities against Cladosporium cucumerinum, Candida albicans and toxic properties against larvae of the yellow fever-transmitting mosquito Aedes aegypti. PMID- 10724190 TI - The propriety of sponsorship of dermatologic conferences. PMID- 10724191 TI - Shadow of a doubt. PMID- 10724192 TI - Frequency and characteristics of enlarging common melanocytic nevi. AB - OBJECTIVE: To analyze the frequency and characteristics of enlarging common melanocytic nevi. DESIGN: Cohort study using digital epiluminescence microscopy (ELM) for documentation and follow-up, with a median follow-up interval of 11.4 months. SETTING: A dermatology department at a university hospital in Vienna, Austria. PATIENTS: One thousand six hundred twelve melanocytic nevi appearing clinically as common nevi, obtained from 385 patients (mean [+/-SD] age, 34.2 +/- 14.8 y; 55.6% female). INTERVENTIONS: Follow-up examination and documentation by digital ELM. MAIN OUTCOME MEASURES: Frequency of enlarging nevi according to age and comparison of ELM features observed in enlarging and nonenlarging nevi. RESULTS: Enlargement was found in 5.3% (n = 86) of nevi. The frequency of enlarging nevi was inversely related to age (P<.001), in that enlarging nevi were common in patients younger than 20 years and relatively rare in older age groups. Epiluminescence microscopy revealed a peripheral rim of brown globules in 48.8% (n = 42) of enlarging nevi. In contrast, a peripheral rim of brown globules was found in only 0.7% (n = 11) of nevi without enlargement (P<.001). Enlarging nevi that were excised in children and adolescents showed no histological signs of atypia. In older age groups, 48.1% of excised enlarging nevi that were clinically diagnosed as common nevi showed some histological signs of atypia. None of the excised enlarging lesions was histologically diagnosed as melanoma. CONCLUSIONS: The frequency of enlarging common nevi is inversely related to age. In the absence of clinical signs of atypia, enlargement alone does not indicate malignancy. A peripheral rim of brown globules is a characteristic ELM feature of symmetrically enlarging melanocytic nevi. PMID- 10724193 TI - Toxic epidermal necrolysis and Stevens-Johnson syndrome: does early withdrawal of causative drugs decrease the risk of death? AB - BACKGROUND: Withdrawal of the drug(s) that cause severe cutaneous adverse reactions is usually recommended without proof that it alters the course of those reactions. OBJECTIVE: To determine whether the timing of causative drug withdrawal is related to the prognosis of patients with toxic epidermal necrolysis (TEN) or Stevens-Johnson syndrome (SJS). DESIGN: A 10-year observational study (January 1, 1987, through October 30, 1997) of patients admitted to a dermatological intensive care unit, using binary logistic regression analysis. SETTING: A single referral unit in a university hospital. PATIENTS: Consecutive sample of 203 patients with TEN or SJS. Exclusion criteria included causative drug undetermined, lack of information on disease evolution, the date of causative drug(s) withdrawal, or the date when the first definite sign of TEN or SJS appeared. MAIN OUTCOME MEASURE: Death before hospital discharge. RESULTS: One hundred thirteen patients were included; 74 had TEN and 39 had SJS; 20 died. The drug causing TEN or SJS was withdrawn early in 64 patients and late (after the first definite sign of TEN or SJS) in 49 patients. After adjustment for confounding variables (age, maximum extent of detachment, admission year, human immunodeficiency virus status), our model showed that the earlier the causative drug was withdrawn, the better the prognosis (odds ratio, 0.69 for each day; 95% confidence interval, 0.53-0.89). Patients exposed to causative drugs with long half-lives had an increased risk of dying (odds ratio, 4.9; 95% confidence interval, 1.3-18.9). The variables did not interact. CONCLUSIONS: Prompt withdrawal of drug(s) that are suspected to cause SJS or TEN may decrease mortality. Prompt withdrawal of causative drugs should be a priority when blisters or erosions appear in the course of a drug eruption. PMID- 10724194 TI - Natural history of erythromelalgia: presentation and outcome in 168 patients. AB - OBJECTIVE: To describe the demographics, presentation, and outcome in patients with erythromelalgia--a rare and poorly understood clinical syndrome defined by the triad of red, hot, painful extremities. DESIGN: Retrospective medical record review with follow-up by survey questionnaire. SETTING: Large tertiary care medical center. SUBJECTS: Patients with erythromelalgia examined at the Mayo Clinic, Rochester, Minn, between 1970 and 1994. INTERVENTION: The medical records of 168 patients were analyzed. Follow-up data, which consisted of answers to 2 survey questionnaires or the most recent information in the medical record from patients still alive and death certificates or reports of death for those deceased patients, were obtained for all but 13 patients. MAIN OUTCOME MEASURES: Survival, morbidity, and quality of life. RESULTS: All patients were white; 122 (72.6%) were female, and 46 (27.4%) were male. At presentation, the patients' mean age was 55.8 years (age range, 5-91 years). Symptoms had been present since childhood in 7 patients (4.2%). Six patients (3.6%) had a first-degree relative with erythromelalgia. Symptoms were intermittent in 163 patients (97.0%) and constant in 5 (3.0%). Symptoms predominantly involved feet (148 patients [88.1%]) and hands (43 patients [25.6%]). Kaplan-Meier survival curves revealed a significant decrease in survival compared with that expected in persons of similar age and of the same sex (P<.001). After a mean follow-up of 8.7 years (range, 1.3-20 years), 30 patients (31.9%) reported worsening of, 25 (26.6%) no change in, 29 (30.9%) improvement in, and 10 (10.6%) complete resolution of the symptoms. On a standard health status questionnaire, scores for all but one of the health domains were significantly diminished in comparison with those in the US general population. CONCLUSION: Erythromelalgia is a syndrome with significantly increased mortality and morbidity compared with the US general population. PMID- 10724195 TI - Expression of SS-A/Ro and SS-B/La antigens in skin biopsy specimens of patients with photosensitive forms of lupus erythematosus. AB - CONTEXT: The reason that only some patients with lupus erythematosus (LE) develop autoantibodies to SS-A/Ro and SS-B/La antigens and photosensitivity is unknown. One hypothesis is that both events are related to the level of expression of these antigens in the skin. OBJECTIVE AND DESIGN: To test this hypothesis, we measured the expression of the 52-kd SS-A/Ro, 60-kd SS-A/Ro, and 48-kd SS-B/La antigens in normal sun-protected and sun-exposed skin in 14 patients with LE with photosensitivity, 12 patients with LE without photosensitivity, and 4 normal individuals. The presence of circulating antibodies to these antigens was measured in all patients. SETTING: Outpatient clinic in an academic medical center. RESULTS: We found that the expression of the 52-kd SS-A/Ro, 60-kd SS A/Ro, and 48-kd SS-B/La antigens in skin biopsy specimens obtained from the same site was 4- to 10-fold higher in patients with LE with photosensitivity than in those patients with LE without photosensitivity (P<.001). Antigen expression was highly correlated with the presence and titer of circulating anti-SS-A/Ro and anti-SS-B/La antibodies (P<.001). CONCLUSIONS: These findings indicate that photosensitivity and the presence and titer of circulating anti-SS-A/Ro and anti SS-B/La antibodies are both directly correlated with the expression of accessible and immunoreactive SS-A/Ro and SS-B/La antigens in the skin specimens of patients with LE. Thus, the expression of these antigens in keratinocytes may be an important determinant of the development of both SS-A/Ro and SS-B/La autoantibodies and of photosensitive forms of LE. PMID- 10724197 TI - Top-cited dermatology authors publishing in 5 "high-impact" general medical journals. AB - BACKGROUND: In addition to publishing in the dermatologic literature, some dermatologists also publish articles in the general medical journals, which enjoy wide circulation and whose articles are often cited. OBJECTIVE: To identify articles and citations to these articles that the most frequently cited authors in the dermatologic literature published in highly cited general medical journals. DESIGN: We obtained a citation database from the Institute of Scientific Information, Philadelphia, Pa, that identified all articles published by the top-cited authors in the dermatologic literature in 5 "high-impact" general medical journals. SETTING: The 5 high-impact general medical journals with the historically highest impact factors. SUBJECTS: Two hundred top-cited authors in dermatology journals and their coauthors. MAIN OUTCOME MEASURE: Number of citations to articles published in 5 high-impact general medical journals. RESULTS: From 1981 to 1998, 120 of the 200 top-cited dermatology authors published a total of 674 papers in the 5 most highly cited general medical journals. Original articles published in these high-impact general medical journals were cited an average 7.5 times more often than articles published in dermatology journals. CONCLUSIONS: Top-cited authors in dermatology journals also frequently publish in the leading 5 high-impact general medical journals. Publications in these journals by dermatologists are often highly cited. PMID- 10724196 TI - Circumcision and genital dermatoses. AB - CONTEXT: It is well recognized that the presence of a foreskin predisposes to penile carcinoma and sexually transmitted infections. We have investigated the relationship between the presence or absence of the foreskin and penile dermatoses. OBJECTIVE: To determine whether there is an association between circumcision and penile dermatoses. DESIGN: A retrospective case control study of patients attending the department of dermatology with genital skin conditions. SUBJECTS: The study population consisted of 357 male patients referred for diagnosis and management of genital skin disease. The control population consisted of 305 male patients without genital skin disease attending the general dermatology clinics over a 4-month period. MAIN OUTCOME MEASURES: The relationship between circumcision and the presence or absence of skin disease involving the penis was investigated. The rate of circumcision in the general male dermatology population was determined. RESULTS: The most common diagnoses were psoriasis (n = 94), penile infections (n = 58), lichen sclerosus (n = 52), lichen planus (n = 39), seborrheic dermatitis (n = 29), and Zoon balanitis (n = 27). Less common diagnoses included squamous cell carcinoma (n = 4), bowenoid papulosis (n = 3), and Bowen disease (n = 3). The age-adjusted odds ratio for all penile skin diseases associated with presence of the foreskin was 3.24 (95% confidence interval, 2.26-4.64). All patients with Zoon balanitis, bowenoid papulosis, and nonspecific balanoposthitis were uncircumcised. Lichen sclerosus was diagnosed in only 1 circumcised patient. Most patients with psoriasis, lichen planus, and seborrheic eczema (72%, 69%, and 72%, respectively) were uncircumcised at presentation. The majority of men with penile infections (84%) were uncircumcised. CONCLUSIONS: Most cases of inflammatory dermatoses were diagnosed in uncircumcised men, suggesting that circumcision protects against inflammatory dermatoses. The presence of the foreskin may promote inflammation by a koebnerization phenomenon, or the presence of infectious agents, as yet unidentified, may induce inflammation. The data suggest that circumcision prevents or protects against common infective penile dermatoses. PMID- 10724198 TI - Cutaneous aspergillosis and acquired immunodeficiency syndrome. AB - BACKGROUND: Primary cutaneous aspergillosis is an uncommon finding in patients with acquired immunodeficiency syndrome (AIDS); only 13 cases have been reported in the literature. OBSERVATIONS: We describe 11 patients with primary cutaneous aspergillosis and AIDS. There does not seem to be an age, sex, race, or human immunodeficiency virus risk factor predisposition. This is a late manifestation of AIDS; patients typically have low CD4 counts (<0.050 x 10(9)/L [<50/microL]) and other AIDS-defining illnesses. The most frequent presentation is in patients with cytomegalovirus disease and neutropenia caused by ganciclovir therapy. Lesions developed at the site of occlusive dressings for an indwelling intravenous catheter site in 10 patients. Neutrophil counts may be normal at the time of diagnosis. A minor presentation is in the patient without neutropenia as a result of traumatic inoculation. Histological findings and/or culture results are required for diagnosis. Patients develop cutaneous lesions despite prophylactic therapy with fluconazole. Lesions can be treated with excision and lifelong therapy with itraconazole. CONCLUSION: Because of the potential morbidity and mortality of cutaneous aspergillosis, a high level of suspicion and prompt institution of therapy is required. PMID- 10724199 TI - Basal cell carcinoma in children: report of 3 cases. AB - BACKGROUND: The peak incidence of basal cell carcinoma occurs in the seventh decade of life and is rare in children. When found in the pediatric age group, basal cell carcinoma is usually associated with a genetic defect, such as basal cell nevus syndrome, xeroderma pigmentosum, or nevus sebaceus. In areas of intense UV radiation exposure, such as the southwestern United States, children may be at increased risk of developing this malignancy de novo. OBSERVATIONS: Three children (2 boys, aged 8 and 16 years, and an 11-year-old girl) from Tucson, Ariz, with isolated basal cell carcinoma unassociated with any other disease or syndrome are described. CONCLUSIONS: Basal cell carcinoma in children is probably the result of a combination of UV radiation exposure and genetic background. Early recognition in children can prevent extensive tissue destruction and excess scarring after excision. A higher index of suspicion for basal cell carcinoma may also aid in prompt diagnosis of a possible genetic disorder, such as basal cell nevus syndrome. PMID- 10724200 TI - Intralesional therapy with anti-CD20 monoclonal antibody rituximab in primary cutaneous B-cell lymphoma. AB - BACKGROUND: We report the use of a new treatment modality in 2 patients with primary cutaneous B-cell lymphoma. In a 58-year-old woman with progressive nodular lesions on the scalp and face, several treatment attempts either failed or could not be used because of severe adverse effects and underlying epilepsy. The patient declined radiotherapy. A 30-year-old man presented with recurrence of tumor nodules occipitally, thoracically, on the arm, and on the right thigh after several excisions. OBSERVATIONS: Intralesional injection of rituximab, a chimeric antibody directed against the CD20 transmembrane antigen present in malignant and normal B cells, resulted in partial regression of tumor nodules. No adverse effects occurred except pain during or shortly after injection and, in one patient, a slight rise in body temperature. Due to the treatment a prolonged complete disappearance of B cells from peripheral blood samples was observed. CONCLUSION: Intralesional rituximab therapy is a nontoxic and effective treatment for cutaneous B-cell lymphoma that deserves further investigation in larger clinical trials. PMID- 10724201 TI - How well are randomized controlled trials reported in the dermatology literature? AB - OBJECTIVE: To assess the methodological quality of the design and reporting of randomized controlled trials published in one major dermatology specialty journal. DESIGN AND DATA SOURCES: In a survey of all published parallel group randomized controlled trials, we found 73 reports with allocation described as randomized from all issues of Clinical and Experimental Dermatology from its inception in 1976 through 1997. MAIN OUTCOME MEASURES: Direct and indirect measures of the adequacy of randomization, trial sample size, baseline comparisons, and intention-to-treat analysis. RESULTS: Hand searching identified 73 randomized controlled trials, but only 31 of these were found by searching MEDLINE for the publication type clinical trials. Of the 73 randomized controlled trials, 68 contained sufficient information to include in the analysis. Only 1 study (1%) reported the method of random sequence generation, and only 5 studies (7%) reported adequate concealment of allocation. Among 38 trials that used simple randomization, the sample sizes in the comparison groups were identical in 22 occasions, raising the possibility that simple randomization might not have been adequately generated or concealed. Most trials (88%) excluded some randomized participants from their analysis. The median sample size was 23 per trial. Only 1 trial reported sample size and statistical power considerations and had an a priori main hypothesis. CONCLUSIONS: Hand searching is important for locating all relevant trials. There is the need for higher methodological quality in clinical trial reporting in dermatology journals. The adoption of the CONSORT (Consolidated Standards of Reporting Trials) statement and checklist for the reporting of trials should enhance the validity of and strengthen the evidence from clinical trials reports. PMID- 10724202 TI - Confidence intervals. PMID- 10724204 TI - Hypnosis in dermatology. AB - BACKGROUND: Hypnosis is an alternative or complementary therapy that has been used since ancient times to treat medical and dermatologic problems. OBJECTIVE: To describe the various uses for hypnosis as an alternative or complementary therapy in dermatologic practice. METHODS: A MEDLINE search was conducted from January 1966 through December 1998 on key words related to hypnosis and skin disorders. RESULTS: A wide spectrum of dermatologic disorders may be improved or cured using hypnosis as an alternative or complementary therapy, including acne excoriee, alopecia areata, atopic dermatitis, congenital ichthyosiform erythroderma, dyshidrotic dermatitis, erythromelalgia, furuncles, glossodynia, herpes simplex, hyperhidrosis, ichthyosis vulgaris, lichen planus, neurodermatitis, nummular dermatitis, postherpetic neuralgia, pruritus, psoriasis, rosacea, trichotillomania, urticaria, verruca vulgaris, and vitiligo. CONCLUSION: Appropriately trained clinicians may successfully use hypnosis in selected patients as alternative or complementary therapy for many dermatologic disorders. PMID- 10724205 TI - Common acquired nevomelanocytic nevi and the fourth dimension. PMID- 10724206 TI - Erythromelalgia--a mysterious condition? PMID- 10724207 TI - Improving the outcome of patients with toxic epidermal necrolysis and Stevens Johnson syndrome. PMID- 10724208 TI - Human immunodeficiency virus infection and cutaneous aspergillosis. PMID- 10724209 TI - Fragile hair and seizures in a child. PMID- 10724210 TI - A long-standing keratotic papular eruption. PMID- 10724211 TI - Painful erythematous ear. PMID- 10724212 TI - Perioral and acral lentigines in an African American man. PMID- 10724213 TI - Sunscreen and melanoma revisited. PMID- 10724214 TI - Increasing the likelihood of early diagnosis of Netherton syndrome by simple examination of eyebrow hairs. PMID- 10724215 TI - Isotretinoin has yet to be shown to affect bone density. PMID- 10724216 TI - Treatment of epidermolysis bullosa simplex (EBS) with tetracycline. PMID- 10724217 TI - Regarding "Risk factors associated with the failure of a venous leg ulcer to heal". PMID- 10724218 TI - Temporal triangular alopecia in association with mental retardation and epilepsy in a mother and daughter. PMID- 10724219 TI - Minocycline-induced hyperpigmentation of the tongue. PMID- 10724220 TI - Intralesional interferon treatment of lentigo maligna. PMID- 10724221 TI - Hyperhidrosis as the only manifestation of hyperandrogenism in an adolescent girl. PMID- 10724222 TI - Chronic, infantile, neurological, cutaneous, and articular syndrome in a neonate: a case report. PMID- 10724223 TI - Clinical outcome after structural failure of rotator cuff repairs. AB - BACKGROUND: The clinical outcome for patients with documented rerupture after open repair of one or more rotator cuff tendons is not well known. The purpose of this study was to evaluate the clinical outcomes of a consecutive series of rotator cuff reruptures after repair and to provide information concerning the advisability of rotator cuff repair in situations in which there may be a high probability of rerupture. METHODS: During prospective follow-up after rotator cuff repairs, we detected, with magnetic resonance imaging, structural failure of the repair in twenty patients, who had a mean age of fifty-nine years at the time of the rotator cuff repair. All patients were clinically examined for the purpose of this report at a mean of thirty-eight months. RESULTS: The reruptures invariably involved the originally torn tendon but were smaller than the original tear in sixteen of the twenty patients. Fatty degeneration of the supraspinatus and infraspinatus muscles, atrophy of the supraspinatus muscle, and glenohumeral osteoarthritis progressed significantly from the preoperative state (p < 0.05). At the time of the most recent follow-up, the subjective shoulder value averaged 75 percent of the value for a normal shoulder. Eleven patients were very satisfied with the result, six were satisfied, two were disappointed, and one was dissatisfied. The mean relative score according to the system of Constant and Murley had increased from 49 percent of the score for a normal shoulder preoperatively to 83 percent postoperatively (p = 0.0001). Pain had decreased significantly, and the ranges of active, pain-free forward elevation and abduction as well as the abduction strength had improved significantly (p < 0.05). The clinical outcome was significantly correlated with the size of the postoperative tear, the stage of postoperative fatty muscle degeneration of the infraspinatus and subscapularis, the postoperative acromiohumeral distance, and the degree of postoperative glenohumeral osteoarthritis (p < 0.05). CONCLUSIONS: This study documents that an attempt at rotator cuff repair significantly decreases pain (p = 0.0026) and significantly improves function (p = 0.0005) and strength (p = 0.0137) even if magnetic resonance imaging documents that the repair has failed. This finding suggests that the potential for rerupture should not be considered a formal contraindication to an attempt at repair if optimal functional recovery is the goal of treatment. PMID- 10724224 TI - The effect of the orientation of the acetabular and femoral components on the range of motion of the hip at different head-neck ratios. AB - BACKGROUND: Prosthetic impingement due to poor positioning can limit the range of motion of the hip after total hip arthroplasty. In this study, a computer model was used to determine the effects of the positions of the acetabular and femoral components and of varying head-neck ratios on impingement and range of motion. METHODS: A three-dimensional generic hip prosthesis with a hemispherical cup, a neck diameter of 12.25 millimeters, and a head size ranging from twenty-two to thirty-two millimeters was simulated on a computer. The maximum range of motion of the hip was measured, before the neck impinged on the liner of the cup, for acetabular abduction angles ranging from 35 to 55 degrees and acetabular and femoral anteversion ranging from 0 to 30 degrees. Stability of the hip was estimated as the maximum possible flexion coupled with 10 degrees of adduction and 10 degrees of internal rotation and also as the maximum possible extension coupled with 10 degrees of external rotation. The effects of prosthetic orientation on activities of daily living were analyzed as well. RESULTS: Acetabular abduction angles of less than 45 degrees decreased flexion and abduction of the hip, whereas higher angles decreased adduction and rotation. Femoral and acetabular anteversion increased flexion but decreased extension. Acetabular abduction angles of between 45 and 55 degrees permitted a better overall range of motion and stability when combined with appropriate acetabular and femoral anteversion. Lower head-neck ratios decreased the range of motion that was possible without prosthetic impingement. The addition of a modular sleeve that increased the diameter of the femoral neck by two millimeters decreased the range of motion by 1.5 to 8.5 degrees, depending on the direction of motion that was studied. CONCLUSIONS: There is a complex interplay between the angles of orientation of the femoral and acetabular components. Acetabular abduction angles between 45 and 55 degrees, when combined with appropriate acetabular and femoral anteversion, resulted in a maximum overall range of motion and stability with respect to prosthetic impingement. CLINICAL RELEVANCE: During total hip arthroplasty, acetabular abduction is often constrained by available bone coverage, while femoral anteversion may be dictated by the geometry of the femoral shaft. For each combination of acetabular abduction and femoral anteversion, there is an optimum range of acetabular anteversion that allows the potential for a maximum range of motion without prosthetic impingement after total hip arthroplasty. These data can be used intraoperatively to determine optimum position. PMID- 10724225 TI - Rheumatoid forefoot reconstruction. A long-term follow-up study. AB - BACKGROUND: The purpose of the present study was to assess the results of reconstruction of the rheumatoid forefoot with arthrodesis of the metatarsophalangeal joint of the great toe, resection arthroplasty of the metatarsal heads of the lesser toes, and open repair of hammer-toe deformity (arthrodesis of the proximal interphalangeal joint) of the lesser toes when this deformity was present. METHODS: A retrospective study of forty-three consecutive patients (fifty-eight feet) with severe rheumatoid forefoot deformities was performed. Six patients (six feet) died before the most recent follow-up, and five patients (five feet) were excluded because a subtotal procedure had been performed. No patient was lost to follow-up. Thus, the study included thirty-two patients (forty-seven feet) in whom reconstruction of a rheumatoid forefoot had been performed by the author. RESULTS: All first metatarsophalangeal joints had successfully fused at an average of seventy-four months (range, thirty-seven to 108 months) postoperatively. The average postoperative hallux valgus angle was 20 degrees and the average postoperative angle subtended by the axes of the proximal phalanx and the metatarsal of the second ray (the MTP-2 angle) was 14 degrees, demonstrating that a stable first ray protected the lateral rays from later subluxation. One hundred and thirty-two (70 percent) of the 188 lesser metatarsophalangeal joints were dislocated preoperatively, compared with thirteen (7 percent) postoperatively. The result of the procedure (as rated subjectively by the patient) was excellent for twenty-three feet, good for twenty-two, and fair for two. There were no poor results. The average postoperative score according to the system of the American Orthopaedic Foot and Ankle Society was 69 points. Postoperative pain was rated as absent in eighteen feet, mild in twenty five, moderate in four, and severe in none. Fifteen feet were not associated with any functional limitations, twenty-eight were associated with limitation of recreational activities, and four were associated with limitation of daily activities. At the time of the most recent follow-up, no special shoe requirements were reported. Fourteen feet (30 percent) had a reoperation for the removal of hardware from the first metatarsophalangeal joint, a procedure on the interphalangeal joint of the great toe, or additional procedures on the lesser toes or lesser metatarsophalangeal joints. CONCLUSIONS: In the present study, arthrodesis of the first metatarsophalangeal joint, resection arthroplasty of the lesser metatarsal heads, and repair of fixed hammer-toe deformities with intramedullary Kirschnerwire fixation resulted in a stable repair with a high percentage of successful results at an average of six years after the procedures. PMID- 10724226 TI - Simultaneous femoral osteotomy and total knee arthroplasty for treatment of osteoarthritis associated with severe extra-articular deformity. AB - BACKGROUND: In the presence of large extra-articular deformity, complex imbalance of the collateral ligaments may result if standard techniques of soft-tissue releases and intra-articular bone resection are used during total knee arthroplasty. The purpose of this paper is to review our experience with simultaneous corrective osteotomy and total knee arthroplasty for the treatment of severe extra-articular femoral deformity associated with ipsilateral osteoarthritis of the knee. METHODS: The results of simultaneous corrective osteotomy and total knee arthroplasty in eleven knees with osteoarthritis and associated extra-articular angular deformity of the femur were reviewed retrospectively. The femoral deformity resulted from fracture malunion in ten knees and from hypophosphatemic rickets in one. There were five primarily uniplanar deformities (four varus deformities and one antecurvatum deformity), five biplanar (varus and antecurvatum) deformities, and one triplanar (varus, antecurvatum, and internal rotation) deformity. Four knees were approached through a standard medial parapatellar arthrotomy and seven, through an anterolateral subvastus approach with an osteotomy of the tibial tubercle. The site of the femoral osteotomy was fixed with a blade-plate in seven patients, a press-fit long-stemmed femoral component in two, and a retrograde femoral nail in two. An extramedullary alignment system was utilized in eight patients, and intramedullary alignment was used in three. RESULTS: The duration of follow-up averaged forty-six months (range, twenty-six to eighty-eight months). According to the classification system of the Knee Society, the mean function score increased from 22 points preoperatively to 81 points at the time of follow-up and the mean knee score increased from 10 points preoperatively to 87 points at the time of follow-up. The mean flexion contracture decreased from 19 degrees preoperatively to 2 degrees at the time of follow-up. The arc of motion averaged 56 degrees (range, 30 to 75 degrees) preoperatively and 89 degrees (range, 65 to 115 degrees) at the time of follow-up. The mechanical alignment in the coronal plane was restored to within 2 degrees of normal in each patient. Ten femoral osteotomy sites healed, and one, in a patient treated with a press-fit long stemmed femoral component, had not healed by the time of follow-up. All seven sites of the tibial tubercle osteotomies healed. There were no complete radiolucent lines at the prosthetic interfaces, and no total knee arthroplasty was revised. One patient had a nonfatal postoperative pulmonary embolism. As determined by clinical examination and the patients' assessment of function, no ligament imbalance was noted at the time of the most recent follow-up. CONCLUSIONS: Simultaneous femoral osteotomy and total knee arthroplasty is a technically difficult but effective treatment for patients with severe femoral deformity associated with ipsilateral osteoarthritis of the knee. We recommend that the femoral osteotomy site be secured with a plate or a locked intramedullary nail, depending on the location of the deformity and the subsequent osteotomy. PMID- 10724227 TI - Occipitocervical stabilization for myelopathy in patients with rheumatoid arthritis. Implications of not bone-grafting. AB - BACKGROUND: Approximately 0.9 percent of the white adult population of the United States and 1.1 percent of the adult population in Europe are affected by seropositive rheumatoid arthritis. As many as 10 percent of those patients may need an operation for atlantoaxial subluxation. Severe instability, especially when associated with vertical subluxation of the odontoid process, can result in progressive cervical myelopathy. Typically, occipitocervical fixation has been performed for these patients with use of autograft bone to achieve long-term stability through a solid fusion. Harvesting the bone graft increases the operative risk to the patient and may result in increased morbidity. In our experience, patients who have had no clear radiographic evidence of fusion following use of occipitocervical instrumentation seemed to have done as well as those who have had obvious fusion. One assumption is that the clinical improvement might be attributable simply to stabilization of the joint rather than to osseous fusion. A longitudinal study was performed on patients with rheumatoid arthritis who required an operation because of craniocervical or upper cervical instability. METHODS: The results of clinical, radiographic, functional, and self-evaluations were studied to determine the efficacy of treatment and to compare the outcomes of bone-grafting with those of procedures done without bone grafting in a group of 150 patients who underwent posterior occipitocervical stabilization with use of a contoured metal implant (a Ransford loop) that was affixed by sublaminar wires. Internal fixation was performed in 120 patients without bone-grafting and in thirty patients with use of autogenous bone grafting. Preoperatively, 23 percent (thirty-five) of the 150 patients had mild neurological involvement (class II, according to the system of Ranawat et al.), 45 percent (sixty-eight) had objective findings of weakness and long-tract signs but were able to walk (class III-A), and 29 percent (forty-three) were quadriparetic and unable to walk (class III-B). The age of the patients at the time of the operation ranged from twelve to eighty-three years (mean, sixty-two years). RESULTS: There were significant improvements in postoperative Ranawat classes at all time-periods (range, p < 0.00005 to p = 0.0066) and in patient ratings of neck pain (range, p < 0.00005 to p = 0.0044) compared with preoperative scores. With the numbers available, there were no significant differences between the patients managed with a graft and those managed without grafting with respect to survival after the operation, Ranawat class, head or neck-pain rating, presence of subaxial abnormalities, radiographic craniovertebral motion, or vertical subluxation. Overall mortality at one month was 10 percent (fifteen of 150), although this value varied directly with the degree of preoperative disability. A second cervical spine operation was required in 11 percent (sixteen) of the 150 patients. CONCLUSIONS: While patients who have rheumatoid disease with anterior atlantoaxial subluxation should be treated with posterior atlantoaxial arthrodesis with use of bone-grafting and internal fixation, we believe that those who present with vertical instability and multi level involvement can be treated with posterior occipitocervical stabilization with use of a contoured occipitocervical loop and sublaminar wire fixation without bone-grafting. Furthermore, we believe that the use of preoperative traction, bone cement, or a postoperative halo vest is unnecessary. Avoiding the harvesting of autogenous bone for grafting reduced the morbidity of this operation without compromising the outcome in these already sick patients. PMID- 10724228 TI - Successful manual reduction of locked metacarpophalangeal joints in fingers. AB - BACKGROUND: Many studies on the etiology and operative treatment of locked metacarpophalangeal joints in fingers have been reported, but there have been few investigations on manual reduction. The rate of success of manual reduction in previous reports has been low, and no consensus has been reached with regard to the best method of manual reduction. On the basis of our experience with operative treatment, we devised a safe method of manual reduction. METHODS: Between January 1987 and December 1995, we reduced a locked metacarpophalangeal joint in twelve female patients; every locked finger was successfully reduced, and complications such as fracture did not occur during manual reduction. The average duration of follow-up was five years and nine months (range, three years and two months to nine years and three months). RESULTS: Six patients had no recurrence of the locking. Four of the six remaining patients had one or two incidents of locking, had no alteration in the activities of daily living, and did not want operative treatment. The two remaining patients reported that they had incidents of locking several times a day, and they requested operative treatment as they were afraid of additional recurrences. One patient had an open reduction fifteen months after the initial episode of locking, and the other patient elected not to have an operation for personal reasons. CONCLUSIONS: We believe that our method of manual reduction should be used to treat a locked metacarpophalangeal joint in a finger and that operative treatment should be limited to patients in whom manual reduction is unsuccessful or the reduction is unstable. PMID- 10724229 TI - Transfer of the pectoralis major muscle for the treatment of irreparable rupture of the subscapularis tendon. AB - BACKGROUND: The clinical diagnosis of a tear of the subscapularis tendon is difficult, and the resulting delays frequently cause a major time-lapse before repair is attempted. Diagnostic delay often means that surgical repair is no longer possible. In twelve patients who had an irreparable tear of the subscapularis tendon, the superior one-half to two-thirds of the tendon of the pectoralis major muscle was used as a substitute for the subscapularis tendon. In order to adapt the orientation of the transferred muscle to that of the subscapularis, it was routed behind the conjoined tendon of the coracobrachialis muscle and the short head of the biceps to the lesser tuberosity. METHODS: The operations were performed between May 1993 and June 1997. The average age of the twelve patients was sixty-five years old (range, forty-nine to eighty-one years old). Eight patients had an isolated rupture of the subscapularis tendon, and four had a concomitant lesion in the form of either a partial or a complete rupture of the supraspinatus tendon. The dominant symptoms were anterior shoulder pain and weakness that had responded poorly to nonoperative therapy. Four patients also had signs of recurrent anterior instability. RESULTS: After an average follow-up interval of twenty-eight months (range, twenty-four to fifty four months), nine of the twelve patients assessed the final result as excellent or good; three, as fair; and none, as poor. Pain was reduced, with the score improving from an average of 1.7 points (of a maximum of 15 points) preoperatively to an average of 9.6 points postoperatively. The patients' subjective functional evaluation improved from an average score of 20 points preoperatively to an average of 63 points postoperatively. The average functional rating with use of the Constant and Murley score increased from 26.9 to 67.1 percent of normal. All four preoperatively unstable shoulders were stable at the time of the latest follow-up. CONCLUSIONS: This repair technique can be recommended as a reconstructive procedure for elderly patients who have an irreparable tear of the subscapularis tendon. PMID- 10724230 TI - A comparative biomechanical investigation of anterior lumbar interbody cages: central and bilateral approaches. AB - BACKGROUND: Some biomechanical studies have been performed to evaluate the stabilization provided by interbody cages, but there are virtually no comparative data for the different designs. Furthermore, most investigators have used animal models, which may have led to different results due to morphological variation in the end plates and articular facets. The objectives of the current study were to evaluate whether two different anterior cage designs (BAK and SynCage) performed differently with respect to immediate stabilization of the spine, whether the cages stabilized the spine significantly compared with its intact condition, and whether the addition of supplementary translaminar screw fixation further stabilized the spine. Stabilization was defined as a reduction in motion after insertion of an implant. METHODS: Twelve lumbar functional spinal units from human cadavera were tested under pure moments of flexion, extension, bilateral axial rotation, and bilateral lateral bending to a maximum of ten newton-meters. The relative intervertebral motions were measured, with use of an optoelectronic camera system, under three test conditions: with the spine intact, after insertion of anterior interbody cages, and after insertion of anterior interbody cages supplemented with translaminar screw fixation. Six specimens were tested for each type of cage: a bilateral, porous, threaded cylinder (BAK) and a central, porous, contoured implant with end-plate fit (SynCage). RESULTS: The cages performed in a similar manner in all directions of loading, with no significant differences between the two designs. The cages significantly stabilized the spine compared with its intact condition in flexion, axial rotation, and lateral bending (the median value for motion was 40, 48, and 29 percent of the value for the intact condition, respectively; p = 0.002 for all three directions). Compared with the cages alone, translaminar screw fixation provided no additional stabilizing effect in these directions but it significantly increased the stability of the spine in extension (the median value for motion was 34 percent of the value with the cages alone; p = 0.013). CONCLUSIONS: There were no differences in the stabilization provided by the two different cage designs. Use of the cages alone stabilized the spine in all directions except extension, and use of supplementary translaminar screw fixation provided additional stabilization only in extension. CLINICAL RELEVANCE: This study demonstrated that interbody cages do not stabilize the lumbar spine in extension, and this observation was not altered by the use of substantially different designs. If the lack of stabilization in extension is a clinical problem, possible solutions include the avoidance of extension postoperatively or the use of supplementary fixation. PMID- 10724231 TI - Os acromiale: frequency, anatomy, and clinical implications. AB - BACKGROUND: Os acromiale is present when the anterior portion of the acromion has one or more separate ossicles. Its frequency has been documented, in radiographic and anatomical studies, to be between 1 and 15 percent. Reports of os acromiale associated with subacromial pathology have been cited to imply that this entity is a cause of subacromial impingement; however, no study has demonstrated an increased frequency of os acromiale in patients with shoulder pain compared with the frequency in the general population. Inconsistencies in the literature concerning anatomy, development, and frequency prompted the current anatomical study. The purpose of this study was to better define the frequency and anatomy of os acromiale in the general population. METHODS: Two thousand three hundred and sixty-seven scapular bones from 1198 human skeletons from the Hamann-Todd Osteological Collection were studied for evidence of os acromiale. The sample consisted of specimens from 1033 men and 165 women, 843 of whom had been white and 355, black. The mean age of the individuals at the time of death was 44.7 years (range, eighteen to eighty-nine years). The frequency of os acromiale was noted, and the specimens were measured. RESULTS: There were 128 cases of os acromiale in ninety-six (8.0 percent) of the 1198 skeletons, and the condition was bilateral in thirty-two (33.3 percent) of the ninety-six skeletons. In twenty cases, the free fragment had been lost but it was assumed that a fragment had been present because the acromion was truncated. Os acromiale was more frequent in blacks than in whites (13.2 compared with 5.8 percent; p < 0.001) and in men than in women (8.5 compared with 4.9 percent; p = 0.09). The mean proportional length of the free fragment was 0.42 compared with the overall length of the acromion. Care was taken to differentiate os acromiale from a normal immature acromion. Six skeletons demonstrated persistent acromial apophyses. All six cases were bilateral; seven fragments were fusing, and five were free. The oldest age at which a persistent normal apophysis was found was twenty-one years. The frequency of os acromiale in specimens from individuals who had been less than twenty-two years old was not significantly different from that in the remainder of the collection (p = 0.74). Twenty-one scapulae had a distinct circumferential line that was suggestive of an acromial joint, but the distal and proximal portions were solidly fused. However, the findings on plain axillary radiographs of sixteen of these specimens were indistinguishable from those of specimens with os acromiale. CONCLUSIONS: An anatomical study, performed to better define the frequency and anatomy of os acromiale in the general population, showed that fused os acromiale, which has not been described previously, might be mistaken for a free ossicle in the clinical setting. PMID- 10724232 TI - Reattachment of the migrated ununited greater trochanter after revision hip arthroplasty: the abductor slide technique. A review of four cases. AB - BACKGROUND: Proximal migration of the ununited greater trochanter following total hip arthroplasty may produce pain and substantial functional disability. Successful reattachment of the migrated fragment is difficult following multiple hip procedures. The purpose of this report is to describe four patients in whom a severely migrated trochanteric fragment was reattached successfully with a modified Charnley-Harris wiring technique after subperiosteal advancement of the abductor muscles from their origin on the iliac wing. METHODS: This series consisted of one man and three women with an average age of sixty years (range, fifty-one to sixty-eight years) at the time of the index procedure. The patients were followed for an average of eighty-one months (range, fifty-five to ninety six months). All patients had undergone mobilization of the abductor muscles based on the superior gluteal neurovascular pedicle to aid with trochanteric reattachment, and all had undergone prior hip operations (average, two). Advancement of the abductor muscles was achieved through a separate transverse curvilinear incision over the iliac crest, and subperiosteal releases of the entire origins of the gluteus minimus, medius, and maximus muscles from the ilium were performed. RESULTS: Roentgenographic union of the trochanteric fragment occurred in all four patients. There were three excellent functional outcomes (Harris hip scores of 90, 94, and 96 points) and one fair functional outcome (76 points). The average improvement in the Harris hip score was 47.5 points (range, 35 to 58 points). Two patients continued to have a mild or moderate Trendelenburg gait postoperatively. Two patients had heterotopic bone formation of no clinical importance. CONCLUSIONS: Use of this technique resulted in union of the greater trochanter, pain relief, and decreased functional disability without major complications in these four patients. More widespread use of this technique may be indicated for the treatment of symptomatic non-union of the greater trochanter when the fragment cannot be reattached to its anatomical location with the hip in less than approximately 20 degrees of abduction. PMID- 10724233 TI - Dysplasia epiphysealis hemimelica of the acetabulum. A report of two cases. PMID- 10724234 TI - Suprascapular nerve entrapment. PMID- 10724235 TI - Overseas volunteerism in orthopaedic education. PMID- 10724236 TI - Tuberculous abscess of the arm. PMID- 10724237 TI - Periprosthetic fracture of the femur PMID- 10724238 TI - Neurovascular injury in hip arthroplasty PMID- 10724239 TI - Congenital muscular torticollis and the associated craniofacial changes. PMID- 10724240 TI - Surgical correction of cryptotia with superiorly based superficial mastoid fascia and skin paddle. AB - An approach for the correction of cryptotia using a superiorly based superficial mastoid fascial flap and a skin paddle is introduced. The buried portion of the auricle was exposed through an incision made along the upper part of the helix, followed by an appropriate correction of the deformed cartilage. Protrusion of the upper portion of the auricle was accomplished using anchoring sutures. A small skin paddle was elevated from the caudal portion of the auricular sulcus with the superiorly based superficial mastoid fascia as the nutrient pedicle and transferred to the temporal skin defect. The procedure was performed in eight auricles in a total of seven patients with cryptotia. A satisfactory contour and protrusion of the auricle were maintained postoperatively, leaving the scar within the auricular sulcus. PMID- 10724241 TI - The tunneled supraclavicular island flap: an optimized technique for head and neck reconstruction. AB - Reconstructive procedures in the head and neck region use a wide range of flaps for defect closure. The methods range from local, mostly myocutaneous flaps and skin grafts to free microsurgical flaps. To ensure a satisfactory functional and aesthetic result, good texture and color of the flap are always essential. Moreover, the donor-site defect needs to be reduced, with no resulting functional or aesthetic impairment. We have found that the shoulder is a region providing an optimum skin texture match to the neck and face. In cadaver dissection, a vascular pedicle extending from the transversal cervical artery with two accompanying veins was found to vascularize a defined region around the shoulder cap. In line with these findings, the previously described fasciocutaneous island flap, nourished by the supraclavicular artery, was developed further and used purely as a subcutaneously tunneled island flap. The tunneling maneuver significantly improves the donor site by reducing scarring. The flap is characterized by a long subcutaneous pedicle of up to 20 cm. The pivot point is in the supraclavicular region and allows the flap to be used in the upper chest, neck, chin, and cheek. In this article, we introduce the anatomic features and present clinical cases underlining the surgical possibilities of the flap in reconstructive procedures with expanded indications. PMID- 10724242 TI - Upper eyelid gold weight implantation in the Asian patient with facial paralysis. AB - Patients with facial paralysis may develop ophthalmic complications. Poor eyelid closure and lagophthalmos place the patient at increased risk for the development of corneal problems such as epithelial defects, stromal thinning, bacterial infection, and even perforation. Initial treatment should be conservative and include the use of ocular lubricants, moisture chambers, and taping of the lower eyelid into proper position. Surgical intervention may be required in patients who have failed medical therapy or in whom the facial paralysis is not expected to improve. Gold weight implantation in the upper eyelid has become a popular procedure to correct upper eyelid retraction and to improve corneal coverage. Previous descriptions of gold weight placement in the upper eyelid have focused on Caucasian eyelid anatomy. However, there are distinct anatomic differences between the Caucasian and Asian eyelids, which dictate the overlying aesthetic differences. We describe our technique for placement of a gold weight in the Asian upper lid, with attention to the maintenance of symmetric eyelid creases. We reviewed the charts of six Asian patients with facial paralysis who underwent gold weight placement in the upper eyelid for the correction of lid retraction. All patients did well functionally and aesthetically, and none developed an extrusion of the implant with this approach. PMID- 10724243 TI - Early release of third-degree eyelid burns prevents eye injury. AB - Burns to the eyelids occur in more than 20 percent of flame injuries and can lead to ocular damage and even blindness. Burn wound contracture can cause ectropion of the eyelid, resulting in exposure keratitis, corneal ulcers, and conjunctivitis. At our hospital, early eyelid release and grafting has made a significant difference in the long-term outcomes of third-degree eyelid burns; however, the question of just how early eyelid release and grafting should take place is an unresolved issue. Fifty-seven children with third-degree eyelid burns were reviewed; 17 had eyelid release within 7 days of receiving eyelid burns and 40 had a delay in eyelid release of more than 7 days after injury. Analysis was by chi-square with the Yates continuity correction or Fisher's exact test when appropriate. Corneal ulcers developed in 2 of 17 of the early eyelid release of third-degree burns, compared with 25 of 40 delayed releases (p = 0.001), exposure keratitis in 3 of 17 early releases, and 30 of 40 in delayed release (p = 0.000); conjunctivitis was identified in 1 of 17 early releases and 14 of 40 delayed eyelid releases (p = 0.025). Release of eyelid burns within 7 days of injury can prevent the development of exposure keratitis, progressive conjunctivitis, corneal ulceration, and the need for corneal surgery. We suggest that early release and grafting should be the treatment of choice for children and young adults with third-degree burns to the eyelids. PMID- 10724244 TI - Lip and vermilion reconstruction with the facial artery musculomucosal flap. AB - The lips are a complex laminated structure. When lost through injury or disease, they present a complex reconstructive challenge. The facial artery musculomucosal (FAMM) flap is a composite flap with features similar to those of lip tissue. In this article, the anatomy, dissection, and clinical applications for the use of the FAMM flap in lip and vermilion reconstruction are discussed. A series of 16 FAMM flaps in 13 patients is presented. Seven patients had upper-lip reconstruction and six had lower-lip reconstruction. Superiorly based FAMM flaps were used in eight patients, and eight inferiorly based flaps were performed in five patients. Three patients had bilateral, inferiorly based flaps. In summary, the FAMM flap is a local flap that can be used for lip and vermilion reconstruction. Although not identical to the lip, it has many similar features, which make it an excellent option for lip reconstruction. PMID- 10724245 TI - Computer planning for distraction osteogenesis. AB - Distraction osteogenesis of the mandible has found an application in the treatment of patients with a variety of different mandibular deformities. Compared with the relatively simple unidirectional distraction of long bones as described by Ilizarov, the three-dimensional distraction of the mandible is extremely complex. Whereas experience with orthognathic surgery clearly demonstrates that careful presurgical planning is necessary to achieve predictable outcomes, there are few reported methods for the planning of mandibular distraction. The authors have developed a method for planning distraction osteogenesis of the mandible that involves the use of three dimensional modeling and animation to simulate distraction osteogenesis in virtual reality. The first step in the authors' treatment planning process is to obtain a three-dimensional computerized scan of the facial skeleton. From this scan, a three-dimensional wire-mesh model is built using animation software. With the same software, a virtual distractor is built and installed on the wire-mesh model. The osteotomies and the distraction process are then simulated. Finally, a recipe for sequencing the linear and angular changes of the distractor is calculated. The authors have used this planning process in seven patients (age range, 4 to 10 years): four with unilateral mandibular deformities and three with bilateral. The planning process has yielded predictable and reproducible results. PMID- 10724246 TI - Distraction osteogenesis: a new surgical technique for use with the multiplanar mandibular distractor. AB - If distraction osteogenesis is to reach its full potential and achieve the level of accuracy that is possible with orthognathic surgery, its outcomes need to be as predictable. To this end, the authors developed a planning process for distraction osteogenesis similar to that used in orthognathic surgery. However, the success of the planning process depends on the authors' ability to execute the plan at the time of surgery. As a result, the authors needed to develop a surgical technique that would enable them to precisely install the distractor as indicated in the presurgical plan. The surgical technique presented in this article was developed for this purpose. The authors used this technique in seven patients (four boys and three girls; age range, 4 to 10 years). Four patients presented with unilateral deformities, and three patients presented with bilateral deformities. The follow-up period in this group of patients ranged from 12 to 33 months. The purpose of the technique is to replicate the position of the distractor on the mandible as determined by the presurgical plan. To this purpose, a custom drill guide and a surgical template have been developed. Both of these are used following the principles of triangulation to establish the pin position and orientation of the distractor. In the authors' hands, the use of this surgical technique has resulted in outcomes close to those predicted by the planning process. PMID- 10724247 TI - Effect of distraction rate on biomechanical, mineralization, and histologic properties of an ovine mandible model. AB - Craniofacial microsomia is a common congenital malformation. Ilizarov's method of distraction osteogenesis applied to the mandible has yielded promising results both experimentally and clinically. Because the technique is used predominantly in a pediatric population, length of treatment and compliance may be problematic. To date, the limits of distraction rate in the craniofacial skeleton have not been defined. This study was designed to investigate the effects of distraction rate, in a large animal model, on the mineralization, biomechanical, and histologic properties of lengthened mandibles. Clinically faster distraction rates would decrease the overall treatment time. Twenty-four animals were divided into four groups, with varying rates of distraction (1, 2, 3, and 4 mm/day). A uniaxial distractor at the angle of the mandible was used. The mandibles were lengthened to 24 mm and fixed for a period of 5 weeks, when the animals were killed. The specimens were analyzed with respect to mineralization using dual energy x-ray absorptiometry, biomechanical strength, through a modified three point bending test, and histologic properties with hematoxylin and eosin stains. Biomechanical, mineralization, and histologic analyses of the samples indicated that group 1 (1 mm/day) samples were significantly superior (p<0.05) to those of group 4 (4 mm/day). Although bone formation was achieved in all groups, group 1 (1 mm/day) demonstrated the strongest biomechanical and histologic properties. Bone mineral density obtained using dual energy x-ray absorptiometry may be clinically useful as a reliable, noninvasive, and relatively cheap predictor for removal time of the fixator. PMID- 10724248 TI - Medial pedicle reduction mammaplasty for severe mammary hypertrophy. AB - Current options in reduction mammaplasty for severe mammary hypertrophy include amputation with free-nipple graft as well as the inferior pedicle and bipedicle techniques. Complications of these procedures include nipple-areola necrosis, insensitivity, and hypopigmentation. The purpose of this study was to determine whether medial pedicle reduction mammaplasty can minimize these complications. Twenty-three patients with severe mammary hypertrophy were studied. The medial pedicle successfully transposed the nipple-areola complex in 44 of 45 breasts (98 percent). Mean change in nipple position was 17.1 cm, and mean weight of tissue removed was 1604 g per breast. Nipple-areola sensation was retained in 43 of 44 breasts (98 percent) using a medial pedicle. Hypopigmentation was not observed, and central breast projection was restored in all patients. This study has demonstrated that medial pedicle reduction mammaplasty is a safe and reliable technique and should be given primary consideration in cases of severe mammary hypertrophy. PMID- 10724249 TI - The sensitivity of the nipple-areola complex: an anatomic study. AB - Although preservation of the sensitivity of the nipple and areola is an important goal in breast surgery, only scant and contradictory information about the course and distribution of the supplying nerves is found in the literature. The existing controversy might be due to the difficulty in dissecting the thin nerves and to frequent anatomic variations that bias the results if only a small number of cadavers are dissected. We dissected 28 female cadavers and found that the nipple and areola were always innervated by the lateral and anterior cutaneous branches of the 3rd, 4th, and 5th intercostal nerves. The most constant innervation pattern was by the 4th lateral cutaneous branch (79 percent) and by the 3rd and 4th anterior cutaneous branches (57 percent). The anterior cutaneous branches took a superficial course within the subcutaneous tissue and terminated at the medial areolar border in all dissected breasts. The lateral cutaneous branches took a deep course within the pectoral fascia and reached the nipple from its posterior surface in 93 percent of the dissected breasts. In 7 percent of the dissected breasts, the lateral cutaneous branches took a superficial course within the subcutaneous fat and reached the nipple from the lateral side. These findings suggest that the nerves innervating the nipple and areola are best protected if resections at the base of the breast and skin incisions at the medial areolar border are avoided. PMID- 10724250 TI - Surgeon perspectives on surgical options for early-stage breast cancer. AB - To evaluate the practice patterns of general and plastic surgeons regarding patients with early-stage breast cancer, all general and plastic surgeons in Quebec and Maryland were mailed self-administered questionnaires evaluating surgeon demographics, practice patterns, treatment preferences, and satisfaction with the results of lumpectomy and radiation therapy or breast reconstruction. Response rates of 38.3 percent and 26.7 percent were obtained for general surgeons in Quebec and Maryland, respectively. The ratio of reported mastectomies to lumpectomies was 1:2 in Maryland and 1:5 in Quebec. All general surgeons considered lumpectomy an important option. Ninety percent of Maryland surgeons versus 44 percent of Quebec surgeons considered mastectomy important. A total of 53.6 percent versus 24.9 percent of general surgeons in Maryland and Quebec, respectively, considered delayed reconstruction an important option. Additionally, 81.3 percent of Maryland surgeons considered immediate reconstruction important, and 79.6 percent discussed it with all stage I or II patients. More than 75 percent of Quebec general surgeons reported discussing immediate or delayed reconstruction with < or =50 percent of these women. Response rates of 53.6 percent and 48.8 percent were obtained for plastic surgeons in Quebec and Maryland, respectively. In one year Quebec plastic surgeons reported that they performed less than half the number of reconstructions performed by Maryland plastic surgeons (7.2 versus 17.3). In Quebec, 82.3 percent of surgeons reported that they frequently discuss delayed reconstruction, 25.1 percent immediate, 62.5 percent pedicled TRAM, and 51.7 percent nonautogenous options. In Maryland, 74.3 percent of plastic surgeons frequently discuss delayed reconstruction, 95.7 percent immediate, 89.9 percent pedicled TRAM, and 85.9 percent nonautogenous options. For women with early-stage breast cancer, regional variations exist in the surgical options discussed and provided. PMID- 10724251 TI - Characteristics of a population of women with breast implants compared with women seeking other types of plastic surgery. AB - Several previous studies have shown that breast implant patients demonstrate a number of differences compared with the general population. However, studies have not compared patients with breast implants with women receiving other types of plastic surgery, of interest because this latter group has been proposed as a comparison group for assessing the long-term health effects experienced by breast implant patients. Questionnaire data obtained from 7447 breast implant patients and 2203 patients with other types plastic surgery were collected during the course of a retrospective cohort study, to determine whether implant patients demonstrate different characteristics compared with a more restricted group of patients. In contrast to previous investigations that compared implant patients with the general population, distinctive differences with respect to family income, number of pregnancies, alcohol consumption, cigarette smoking, or histories of previous gynecologic operations or operations for benign breast disease were not found. However, implant patients were significantly more likely than other plastic surgery patients to be white, have low levels of education, have early ages at first birth, be thin, and be screened frequently for breast disease. Furthermore, implant patients reported somewhat greater use of exogenous hormones and familial histories of rheumatoid arthritis. These results support the notion that other plastic surgery patients are a more appropriate comparison group than women in the general population for studies of the health effects of breast implants; however, there continue to be distinctive characteristics possessed by breast implant patients, which need to be taken into account in an assessment of what disease effects can be uniquely attributed to silicone breast implants. PMID- 10724252 TI - Staged breast reconstruction with saline-filled implants in the irradiated breast: recent trends and therapeutic implications. AB - A retrospective review was performed of one surgeon's experience with 40 consecutive patients who had undergone two-stage saline-filled implant breast reconstruction and radiation during the period from 1990 through 1997. A randomly selected group of 40 other two-stage saline-filled implant breast reconstructions from the same surgeon and time period served as controls. This review was undertaken because of the absence of specific information on the outcome of staged saline implant reconstructions in the radiated breast. Previously published reports on silicone gel implants and radiation have been contradictory. At the same time, the criteria for the use of radiation in the treatment of breast cancer have been expanded and the numbers of reconstruction patients who have been radiated are increasing dramatically. For example, in a 1985 report on immediate breast reconstruction, only 1 of 185 patients over a 6-year period underwent adjuvant radiation therapy, whereas in this review, there were 40 radiated breasts with saline-filled implants, 19 of which received adjuvant radiation therapy during their expansion. The study parameters included patient age, breast cup size, implant size, length of follow-up, number of procedures, coincident flap operations, Baker classification, complications, opposite breast procedures, pathologic stage, indications for and details about the radiation, and outcomes. The use of radiation in this review of reconstructed breasts can logically be divided into four groups: previous lumpectomy and radiation (n = 7), mastectomy and radiation before reconstruction (n = 9), mastectomy and adjuvant radiation during reconstruction/expansion (n = 19), and radiation after reconstruction (n = 5). The largest and most rapidly growing group of patients is of those receiving postmastectomy adjuvant radiation therapy. A total of 47.5 percent (19 of 40) of radiated breasts with saline implants ultimately needed the addition of, or replacement by, a flap. Ten percent of a control group with nonradiated saline implant reconstructions also had flaps, none as replacements. Fifty percent or more of both the radiated and control groups had contralateral surgery. Complications were far more common in the radiated group; for example, there were 32.5 percent capsular contractures compared with none in the control group. The control nonradiated implant-only group and the flap plus implant radiated group did well cosmetically. The radiated implant-only group was judged the worst. The increasing use of radiation after mastectomy has important implications for breast reconstruction. The possibility for radiation should be thoroughly investigated and anticipated preoperatively before immediate breast reconstruction. Patients with invasive disease, particularly with large tumors or palpable axillary lymph nodes, are especially likely to be encouraged to undergo postmastectomy radiation therapy. The indications for adjuvant radiation therapy have included four or more positive axillary lymph nodes, tumors 4 cm (or more) in diameter, and tumors at or near the margin of resection. More recently, some centers are recommending adjuvant radiation therapy for patients with as few as one positive lymph node or even in situ carcinoma close to the resection margin. The use of latissimus dorsi flaps after radiation has proven to be an excellent solution to postradiation tissue contracture, which can occur during breast expander reconstruction. The use of the latissimus flap electively with skin sparing mastectomy preradiation is probably unwise, unless postmastectomy radiation is unlikely. Skin-sparing mastectomy with a latissimus flap thus should be preserved for patients unlikely to undergo adjuvant radiation therapy. Purely autologous reconstruction such as a TRAM flap is another option for these patients, either before or after radiation therapy. PMID- 10724253 TI - Cost-based comparison between perforator flaps and TRAM flaps for breast reconstruction. AB - More women than ever before are undergoing mastectomies secondary to increased awareness and screening. This increase has also caused a corresponding increase in the number of breast reconstructions requested each year. The increased demand for reconstruction has fueled recent advances in new techniques. Aside from foreign-body reconstruction such as implants, the methods now being used are related to autogenous donations and reconstruction. Transverse rectus abdominis myocutaneous (TRAM) flaps and perforator flaps are currently being used for autogenous breast reconstruction. This study will compare these two techniques on the basis of cost and length of stay. A retrospective study of 49 patients undergoing a total of 64 perforator flap breast reconstructions at Memorial Medical Center in New Orleans, Louisiana, during the 1997 calendar year was used. There were 59 deep inferior epigastric perforator and five gluteal artery perforator breast reconstructions. All patients underwent some form of breast reconstruction and differed only in respect to whether a mastectomy was performed and whether the reconstruction was unilateral or bilateral. Those patients who underwent a mastectomy with immediate perforator flap reconstruction (n = 26) were then compared with patients undergoing mastectomy with immediate TRAM flap reconstruction (n = 154) at the University of Texas M. D. Anderson Cancer Center. The data from the Anderson Study were obtained from material published in Plastic and Reconstructive Surgery in 1996. Comparison of patients was limited to those who underwent mastectomy with immediate breast reconstruction because this was the design of the M. D. Anderson study. This approach allowed a cost and length of stay comparison while keeping other variables relatively similar. Patients in the perforator flap series enjoyed a marginally shorter operating time and a much shorter length of stay. On average, the operative time for all perforator flap reconstructions was approximately 2 hours shorter than for all TRAM flaps. As for length of stay, perforator flap patients were discharged, on average, 3 days after the initial reconstruction. In contrast, TRAM flap patients remained in the hospital for an average of approximately 7 days after the initial reconstruction. The overall total, average cost for the perforator flap reconstruction in this study is $9625, whereas the average cost of all TRAM flaps performed in the Anderson study is $18,070. PMID- 10724254 TI - The isolated burned palm in children: epidemiology and long-term sequelae. AB - The isolated burn of the palm is a typical injury in young children. Positioning and splinting in small hands is difficult, and long-term sequelae of these injuries are not uncommon. The objective of the present study was to assess the outcome of palm burns and to identify the risk factors for long-term sequelae. All patients admitted to our hospital affected with isolated palm injuries between January of 1988 and January of 1998 were reviewed. In total, 120 pediatric patients were admitted with isolated palm burns; 110 patients (91.7 percent) had partial-thickness burns, and 10 patients (8.3 percent) had full thickness burns. Only four patients (3.3 percent) required excision and skin autografting, but all patients whose palms were operated on in the acute phase developed burn contractures. Sixteen patients (13.3 percent) developed palmar contractures, and more than half of them (56 percent) required reconstructive procedures. All palm burns that healed in more than 3 weeks developed scarring and sequelae (p<0.05 compared with no sequelae). Pediatric palmar burns are benign injuries with a low incidence of late sequelae. However, flame and contact burns are more prone to develop scarring. Excision and autografting should be performed on wounds that take over 3 weeks to heal, but it does not prevent late sequelae. PMID- 10724255 TI - Experimental use of fibrin glue to induce site-directed osteogenesis from cultured periosteal cells. AB - The purpose of this study was to determine whether a combination of fibrin glue and cultured periosteal cells will result in new bone formation at heterotopic sites in nude mice. Growing cells and developing matrices surrounding periosteal explants from the diaphyses of radii of newborn calves were minced and mixed with fibrin glue in a syringe. The cell/matrix-fibrin glue admixture was then injected into the subcutaneous space on the dorsum of athymic nude mice. After 12 weeks of implantation, gross morphology and histologic investigations showed newly formed bone structures in all cell/matrix-fibrin glue admixtures, but none in fibrin glue injected alone and used as control samples. Osteopontin, a protein important in bone development, was identified by a Western blot assay of the cell/matrix fibrin glue composite. This study supports the feasibility of initiating site directed formation of bone structures at heterotopic tissue sites by means of injection of cultured periosteal cells and matrix in a fibrin glue carrier. PMID- 10724256 TI - Vascular delay and administration of basic fibroblast growth factor augment latissimus dorsi muscle flap perfusion and function. AB - Ischemia of the distal latissimus dorsi muscle flap occurs when the entire muscle is acutely elevated. Although this level of ischemia may not be critical if the muscle is to be used as a conventional muscle flap, the ischemia causes decreased distal muscle function if it is used for dynamic muscle flap transfer. This experiment was designed to determine whether or not the administration of exogenous basic fibroblast growth factor (bFGF), combined with a sublethal ischemic insult (i.e., vascular delay), would further augment muscle perfusion and function. Both latissimus dorsi muscles of nine canines were subjected to a bipedicle vascular delay procedure immediately followed by thoracodorsal intraarterial injection of 100 microg of bFGF on one side and by intraarterial injection of vehicle on the other. Ten days later, both latissimus dorsi muscles were raised as thoracodorsally based island flaps, with perfusion determined by laser-Doppler fluximetry. The muscles were wrapped around silicone chambers, simulating cardiomyoplasty, and stimulating electrodes were placed around each thoracodorsal nerve. The muscles were then subjected to an experimental protocol to determine muscle contractile function. At the end of the experiment, latissimus dorsi muscle biopsies were obtained for measurement of bFGF expression. The results demonstrated that the administration of 100 microg of bFGF immediately after the vascular delay procedure increases expression of native bFGF. In the distal and middle muscle segments, it also significantly increased muscle perfusion by approximately 20 percent and fatigue resistance by approximately 300 percent. The administration of growth factors may serve as an important adjuvant to surgical procedures using dynamic muscle flap transfers. PMID- 10724257 TI - Differential expression of receptor tyrosine kinases and Shc in fetal and adult rat fibroblasts: toward defining scarless versus scarring fibroblast phenotypes. AB - The remarkable ability of the fetus to heal early gestation skin wounds without scarring remains poorly understood. Taking advantage of recent advances in signal transduction, the tyrosine phosphorylation patterns of fetal rat fibroblasts, representing the scarless cutaneous repair phenotype, and adult rat fibroblasts, representing scarforming phenotype, were examined whether there were inherent differences in cellular signaling. Specifically, correlation of the phosphorylation patterns with the expression levels of the signaling molecules that transmit information from the plasma membrane receptor to the nucleus was sought. By using three different cell lines of explanted fibroblasts from gestational day 13 fetal rat skin (n = 24) and 1-month-old postnatal adult rat skin (n = 3), immunoblotting was performed to compare tyrosine phosphorylation patterns. The results revealed five major protein bands of interest in fetal rat fibroblasts, but not in the adult rat fibroblasts. These phosphorylated protein bands are of interest because of their possible role in wound repair and may have the potential to regulate cellular responses to the extracellular matrix and their secondary signaling molecules. It was hypothesized that these bands represented receptor tyrosine kinases, epidermal growth factor receptor, and discoidin domain receptor 1, and their downstream adaptor protein Shc that binds receptor tyrosine kinases to transduce signals intracellularly. Furthermore, elevated expression of platelet-derived growth factor receptor-beta in adult compared with fetal fibroblasts was demonstrated, suggesting that decreased expression of certain growth factors may also be important for the scarless phenomenon to occur. PMID- 10724258 TI - Transforming growth factor beta superfamily members: role in cartilage modeling. AB - Normal and abnormal extracellular matrix turnover is thought to result, in part, from the balance in the expression of metalloproteinases and tissue inhibitors of metalloproteinases (TIMPs). The clinical manifestations of an imbalance in these relationships are evident in a variety of pathologic states, including osteoarthritis, deficient long-bone growth, rheumatoid arthritis, tumor invasion, and inadequate cartilage repair. Articular cartilage defects commonly heal as fibrocartilage, which is structurally inferior to the normal hyaline architecture of articular cartilage. Transforming growth factor-beta 1 (TGF-beta1), a cytokine central to growth, repair, and inflammation, has been shown to upregulate TIMP-1 expression in human and bovine articular cartilage. Additionally, members of the TGF-beta superfamily are thought to play key roles in chondrocyte growth and differentiation. Bone morphogenetic protein-2 (BMP-2), a member of this superfamily, has been shown to regulate chondrocyte differentiation states and extracellular matrix composition. It was proposed that, by optimizing extracellular matrix composition, BMP-2 would enhance articular cartilage healing. After determining the release kinetics of BMP-2 from a collagen type I implant (Long-Evans male rats; two implants/rat, n = 14), it was found that, in a tissue engineering application, BMP-2 induced a hyaline-like repair of New Zealand White rabbit knee articular cartilage defects (3-mm full-thickness defects in the femoral trochlea; 2 defects/rabbit, n = 36). The quality of cartilage repair with BMP-2 (with or without chondrocytes) was significantly better than defects treated with BMP-2, as assessed by a quantitative scoring scale. Immunohistochemical staining revealed TIMP-1 production in the cartilage defects treated with BMP-2. When studied in vitro, it was found that BMP-2 markedly increased TIMP-1 mRNA by both bovine articular and human rib chondrocytes. Additionally, increased TIMP-1 mRNA was translated into increased TIMP-1 protein production by bovine chondrocytes. Taken together, these data suggest that BMP-2 may be a useful cytokine to improve healing of cartilaginous defects. Furthermore, these data suggest that the beneficial effects of BMP-2 may be, in part, related to alterations in extracellular matrix turnover. PMID- 10724259 TI - The effect of muscle flap transposition to the fracture site on TNFalpha levels during fracture healing. AB - The trauma and sepsis that follow open fractures and wounds may lead to the production of various cytokines. Understanding wound healing requires a direct knowledge of the specific cytokines and the respective wound fluid levels that are present at the wound site. An animal model was designed that mimics the open fracture and the clinical repair of the human, high-energy open fracture. Canine right tibiae were fractured with a penetrating, captive-bolt device, then repaired in a standard clinical fashion using an interlocking intramedullary nail. Before primary wound closure, microdialysis probes were placed at the fracture site and in a muscle located at a contralateral site. Canines received one of the following experimental protocols: (1) tibial fracture (n = 5); (2) tibial fracture plus Staphylococcus aureus inoculation at the fracture site (n = 5); and (3) tibial fracture, S. aureus inoculation, and a rotational gastrocnemius muscle flap (n = 5). Microdialysis fluid samples were collected intermittently for 7 days. Tumor necrosis factor alpha (TNFalpha) levels at the fracture site were significantly elevated 3 to 34-fold (p<0.02), as compared with respective serum levels at all time points for all treatment groups. Fracture site TNFalpha levels were elevated (p<0.02) in days 1 through 6, as compared with the baseline and contralateral in all treatment groups. At days 1 through 6, the TNFalpha levels of the muscle flap group fracture site were significantly decreased by approximately 50 percent (p<0.05), as compared with the fractures without muscle flaps and regardless of additional S. aureus inoculation. On day 7, fracture site TNFalpha levels in all animal groups were similar, yet remained well above those of baseline TNFalpha. These results demonstrate that S. aureus does not further elevate TNFalpha levels in the presence of an open fracture and that a muscle flap reduces pro-inflammatory TNFalpha levels during early wound healing. This experimental model allows for the characterization of specific biological signals and cellular pathways that are influenced by bacterial infection and surgical closure. These data provide a scientific framework on which to judge or validate therapeutic regimens for open-fracture wound healing. PMID- 10724260 TI - Another method to prevent venous thrombosis in microsurgery: an in situ venous catheter. AB - Free-flap failure is in the order of 4 to 10 percent. Heparin is more effective at preventing venous thrombosis than arterial thrombosis. This study was undertaken to investigate the efficacy of delivering heparin at a high dose locally but low dose systemically (heparin infusion via a catheter placed proximal to the venous anastomosis) to prevent venous thrombosis in microsurgery. A model of venous thrombosis was first established by a venous inversion graft in the rat femoral vein (this was performed in seven animals and resulted in 100 percent thrombosis). Saline and heparin were delivered proximal to the inverted vein graft to assess the effect of each in preventing venous thrombosis. Flow/patency distal to the inverted vein graft was assessed by observation under the microscope, the milk test, and rate of flow (flowmeter). Saline infused via a catheter proximal to the venous inversion graft resulted in 100 percent thrombosis in 10 animals. Heparin (100 U/ml at 2 to 3 ml/hour) infused through a catheter for 2 hours proximal to the anastomosis resulted in flow in all 10 animals during the infusion. Blood was also taken before beginning the procedure (control) and after the heparin infusion distal to the anastomosis (local partial thromboplastin time) as well as in the contralateral femoral vein (systemic). The control for all animals that received heparin was <3 minutes. The systemic partial thromboplastin time after heparin infusion was <3 minutes in seven animals, 3.3 minutes in two animals, and >7 minutes in one animal. The local partial thromboplastin time distal to the inverted vein graft was >10 minutes in nine animals and 3.7 minutes in one animal. The study also had a clinical component, in which a catheter was placed in a vein of the free flap, and heparin was infused over 5 days. This technique has been used in 83 consecutive free flaps. In three recent free flaps performed on the limbs, the local partial thromboplastin time (close to the anastomosis) was raised but the systemic time was normal. This technique offers a method in preventing venous thrombosis in microsurgery. It is simple to implement and is not associated with the systemic complications of heparin. PMID- 10724261 TI - The vascularized pig fibula bone flap model: effect of segmental osteotomies and internal fixation on blood flow. AB - The free fibular flap is the flap of choice for reconstruction of complex mandibular defects, although two or more osteotomies may be required to recreate the normal mandibular contour. The effect of these surgical manipulations on the fibula has not been adequately investigated. This study was designed to study the effect of multiple segmental osteotomies and internal fixation techniques on blood flow in the vascularized pig fibula bone flap model. The hindlimbs of 15 Yorkshire pigs were randomized into 1 of 5 groups (n = 6 fibulae per group) consisting of: (1) a nonoperated, in situ fibula; (2) an elevated fibula flap; (3) an elevated fibula flap with two segmental osteotomies; (4) an elevated fibula with two segmental closing osteotomies rigidly fixed with 2-mm miniplates; (5) an elevated fibula with two segmental closing osteotomies rigidly fixed with 2-mm lag screws. Total and gradient blood flow was measured in the bone and soft tissue components of these flaps using the 15-microm radioactive microsphere technique. The creation of two segmental osteotomies in the vascularized pig fibula bone flap model resulted in a significant decrease (p<0.05) in the gradient blood flow in the segment of bone distal to the second osteotomy. Application of miniplates or lag screws across closing osteotomies resulted in a significant decrease (p<0.05) in total and gradient blood flow to the bone component of the fibulae, as compared with the elevated and osteotomized fibulae groups. An increase in blood flow suggesting a hyperemic response was noted in the bone and soft tissue in the elevated and osteotomized flap groups as compared with the in situ, nonoperated controls. This study established the validity of the pig fibula as a suitable model for investigating the pathophysiology of blood flow changes in the face of standard surgical maneuvers necessary for the restoration of mandibular form and function. The results demonstrated that the creation of multiple segmental osteotomies and the application of internal fixation significantly decreases (p<0.05) blood flow to the distal portion of the flap. The effects of segmental osteotomies and internal fixation on healing and growth of the pig fibula bone flap model are investigated in a separate study. PMID- 10724262 TI - Gluteus maximus adipomuscular turnover or sliding flap in the surgical treatment of extensive sacral chordomas. AB - Two cases with extensive posterior peritoneal defects after high sacral amputation for sacral chordoma are presented. An adipomuscular flap as a modification of the conventional gluteus maximus muscle flap was designed to obliterate an extensive residual posterior peritoneal dead space. The deep adipose tissue beneath the superficial fascia left on the gluteus maximus muscle was effectively used to provide more volume to the flap. The adipomuscular flap was turned over into the posterior peritoneal defect in the first case, and the flap was slid into the cavity in the other case. The adipomuscular flap eventually enabled the successful reconstruction of the posterior peritoneal defect, and the volume of the flap was well maintained behind the rectum, according to the postoperative magnetic resonance imaging findings in both cases. PMID- 10724263 TI - Osteoma cutis of the hand. PMID- 10724264 TI - Practical do-it-yourself device for accurate volume measurement of breast. AB - A simple and accurate method of measuring differences in breast volume based on Archimedes' principle is described. In this method, a plastic container is placed on the breast of the patient who is lying in supine position. While the breast occupies part of the container, the remaining part is filled with water and the volume is measured. This method allows the measurement of the volume differences of asymmetric breasts and also helps the surgeon to estimate the size of the prosthesis to be used in augmentation mammaplasty. PMID- 10724265 TI - "Apron" flap and re-creation of the inframammary fold following TRAM flap breast reconstruction. AB - To the best of our knowledge, the recreation of an inframammary fold after TRAM flap breast reconstruction has not yet been described. This article offers a technique for the creation of an inframammary fold as a secondary procedure. The technique has been performed thus far in two patients with good aesthetic outcomes and no postoperative complications. It may also be suitable for adding bulk to the TRAM flap, especially in bilateral breast reconstruction, and for other minor chest deformities. PMID- 10724266 TI - Razor-assisted treatment of axillary osmidrosis. AB - Shaving off the sweat glands and hair follicles with a single-use safety razor is described. PMID- 10724267 TI - Plastic surgical perspectives on vascular endothelial growth factor as gene therapy for angiogenesis. AB - The practice of plastic surgery has always remained at the frontier of medical science. Over the past few decades, this frontier has been marked by significant developments in the field of gene therapy. Gene therapy serves to replace, supplement, or manipulate a patient's genetic makeup to restore function that has been lost or to correct function that is aberrant. Recent technology may allow surgeons to augment the processes of wound healing and angiogenesis by transfecting genes encoding desirable proteins, such as vascular endothelial factor (VEGF), into ischemic tissues. VEGF is a vital growth factor in the development of blood vessels. Although its mechanisms of action are numerous, its sole function seems to be the augmentation of angiogenesis. VEGF is active in growth and development, in wound healing, and in various pathologic conditions, such as psoriasis and rheumatoid arthritis. The role of VEGF in the field of plastic surgery is just beginning to be explored; it may someday prove to be very rewarding. PMID- 10724268 TI - Cleft lip: unilateral primary deformities. AB - The understanding and management of all aspects of unilateral cleft lip deformities continue to evolve. Just as we are entering the era of exciting advances in the understanding of the pathogenesis of craniofacial disorders, expansion of our understanding of the dynamic relationships of the structural and soft-tissue components of cleft deformities has assisted surgeons in achieving progressively improved and consistent outcomes for these patients. The anatomic and physiologic complexity of unilateral cleft lip deformities has been recognized for centuries, and generations of researchers have cumulatively contributed to our current understanding. This article examines the history, classification, anatomy, and controversies in the surgical management of unilateral cleft lip deformities, allowing surgeons to formulate a reasoned, longitudinal management plan for their patients on the basis of the available current data. PMID- 10724269 TI - American Society of Plastic Surgeons: what's in a name change? PMID- 10724270 TI - An examination of the phenol-croton oil peel: part IV. Face peel results with different concentrations of phenol and croton oil. AB - In Part IV of this examination of the phenol-croton oil peel, the author presents peeling solutions using phenol in concentrations between 16% and 50% as the carrier for croton oil. Previously, in Part I, the author showed that phenol alone in concentrations of less than 50% has no significant peeling effect on the skin in the absence of taping. All of these formulas are dependent on the addition of croton oil for their peeling action. A topographic map of the face is presented that divides the face into the zones that the author believes are best treated with different strengths of croton oil. Five patients peeled between late 1992 and late 1995 were chosen as examples to illustrate the effect of different strengths of croton oil between 0.25% and 2.78%. The author has documented their immediate postoperative course photographically to show the effect of the different concentrations. It is clinically apparent that peels using croton oil between 0.25% and 0.5% generally heal within 7 days; peels between 0.6% and 1.0% usually heal within 9 or 10 days, and peels using concentrations higher than 1% heal later and have some risk of pigmentation loss. Peels using croton oil concentrations at 2% and above almost always have pigmentation loss and have healing delays in areas other than the thick skin of the lower nose and perioral area. The practical clinical formulas distributed at the time of the presentation of this article at the 1996 Annual Meeting of the American Society for Aesthetic Plastic Surgery in Orlando, Florida, entitled "Heresy Phenol Formulas--1996," are provided here. These have been used in both the United States and Europe over the past few years. A metric standard for drop size is suggested at 0.04 ml. This relates to the drop size used clinically over the years to measure croton oil. The adoption of this unit will make formulas around the world easier to calculate and compare. The author has produced a metric formula using the suggested standard size drop for croton oil. This uses 35% phenol as the carrier and provides the same range of treatment dilutions as the 1996 "Heresy Phenol Formulas." The need for research into "carriers" and solvents for croton oil is pointed out. Despite what is not known about how it works, the combination of croton seed extract and phenol has been a success story in providing facial rejuvenation from the 1920s to the present. The croton oil-phenol peel in its many formulas still sets the standard for facial rejuvenation. PMID- 10724271 TI - Formulation of trichloroacetic acid peeling solution: a bibliometric analysis. AB - Since the beginning of this century, trichloroacetic acid solutions of various concentrations have been used for chemical exfoliation. These solutions have been prepared by using four different formulas. To prepare a 50% solution, for instance, water may be added to 50 g of trichloroacetic acid crystals until 100 ml of solution is obtained (weight-to-volume solution). Alternatively, 50 g of water may be added to 50 g of trichloroacetic acid crystals (weight-to-weight solution), or 50 g of trichloroacetic acid crystals may be solved in 100 ml of water (weight-plus-volume solution). Finally, a saturated trichloroacetic acid solution (or "100% solution") may be diluted by an equal volume of water (dilution). Depending on the method used, these so-called 50% solutions contain 40 to 71 weight-to-volume percentages of trichloroacetic acid. From a review of 120 publications on trichloroacetic acid peeling that have appeared since 1926, it was concluded that the authors of 87 of these publications (73 percent) did not report their formula for the trichloroacetic acid solution. Any one of the four methods was reported to have been used by the 33 authors who did report their formula. Eight of 10 internationally reputed pharmacopeias were found not to include the formula of a trichloroacetic acid solution. Proper evaluation of results and prevention of complications of trichloroacetic acid chemexfoliation is only feasible if both the concentration and the formula of trichloroacetic acid solution are reported by the author. Practitioners who use a trichloroacetic acid solution need to establish that the concentration of the solution they apply corresponds with that of the solution reported in the literature. PMID- 10724272 TI - Experience with a strong bleaching treatment for skin hyperpigmentation in Orientals. AB - Although a variety of topical treatments have been used for skin hyperpigmentation, the effectiveness of each varies after prolonged treatment. In this study, 136 Oriental patients who were followed up for more than 12 weeks were analyzed. The treatment protocol was composed of two steps: bleaching (2 to 6 weeks) and healing (2 to 6 weeks); 0.1% to 0.4% all-trans retinoic acid aqueous gel was originally prepared and applied concomitantly with hydroquinone-lactic acid ointment for bleaching. After obtaining sufficient improvement of the hyperpigmentation, a corticosteroid was applied topically with hydroquinone and ascorbic acid for healing. Improvement was evaluated with a narrow-band reflectance spectrophotometer. The results were successful in more than 80 percent of cases of senile lentigines and postinflammatory hyperpigmentations, especially on the face. Sixty percent of cases of nevus spilus were also successfully treated. Although the transient adverse effects of this treatment may be more severe than conventional treatment, this strong bleaching protocol improves a variety of hyperpigmented lesions, including nevus spilus, with a higher success rate and a shorter treatment period than conventional protocols. PMID- 10724273 TI - Endoscopic foreheadplasty: a histologic comparison of periosteal refixation after endoscopic versus bicoronal lift. AB - Endoscopic brow lift techniques using temporary fixation rely on rapid readherence of the periosteum to calvarial bone. Little is known about the histologic events that occur during the early postoperative period after these procedures. An animal study was designed to compare and contrast periosteal fixation to bone and unelevated periosteum, with endoscopic and bicoronal brow lift techniques. One method of temporary fixation is the use of absorbable (polylactic/polyglycolic acid copolymer) LactoSorb screws; a histologic analysis of implanted LactoSorb screws was also performed. Sixteen rabbits underwent brow lifts; eight underwent endoscopic brow lift and fixation with LactoSorb screws without skin excision, and another eight underwent traditional bicoronal brow lift with skin excision and closure under tension. Animals were killed 1, 2, 6, and 12 weeks after the procedures were performed to evaluate the interaction of periosteum and bone and the normal, unelevated periosteum/calvarium interface at a site distant from the operative area. Histologic specimens were examined for the degree of apposition of periosteum to bone and for any fibrous or bony reaction at this interface. Histologic analysis showed various degrees of periosteal fibrosis and fixation to calvarial bone. After an initial phase of minimal periosteal adherence and moderate inflammation, the periosteum became progressively more adherent to bone in both groups, with no significant differences between treatment groups in rates of fixation. Fixation required at least 6 weeks. LactoSorb screws were surrounded by an area of mild inflammation and were progressively hydrolyzed and digested. Periosteal fixation increases over time for bicoronal and endoscopic brow lifts with minimal differences between the two techniques. With this animal model, periosteal adherence to calvarium requires at least 6 weeks with complete adherence by 12 weeks. In addition, the use of absorbable fixation screws seems to be both effective and well tolerated. The histologic changes associated with periosteal healing observed in this study suggest that permanent or semipermanent fixation may improve the accuracy and early postoperative maintenance of forehead advancement. PMID- 10724274 TI - Lateral subcutaneous brow lift and interbrow muscle resection: clinical experience and anatomic studies. AB - The authors report consistent improvement in 65 patients with lateral brow ptosis by using a lateral subcutaneous brow lift at the temporal hairline. In 48 of these patients, vertical glabellar wrinkles were improved by the direct excision of procerus, corrugator, and orbicularis muscles through 3-mm medial brow incisions. Anatomic dissections in 10 cadavers and examinations of 50 skulls were used to study the location of the supraorbital and supratrochlear nerves. Dissections revealed that the supratrochlear nerve was never closer than 1.6 cm to the midline at the level of the supraorbital ridge. In no dissection was a supratrochlear foramen noted. Lateral subcutaneous brow lift was consistently successful in elevating the lateral brow. In no patient was nerve damage to the supraorbital nerve noted. In most patients, the temporal hairline was improved by excising a triangle of balding scalp. Through 3-mm medial brow incisions, the interbrow musculature can be excised by using a small rongeur in an area 3.2 cm wide without risk of nerve damage, improving vertical glabellar wrinkles. PMID- 10724275 TI - Temporal brow lift using botulinum toxin A. AB - The objective of this study was to determine whether brow elevation occurs as a result of paralysis of brow depressors after botulinum toxin A injection. The study's design was a prospective case series with pretreatment and posttreatment outcome evaluation with statistical analysis at a university-based division of facial plastic surgery private clinic. Twenty-two patients of a consecutive sample desiring a cosmetic enhancement underwent injection of botulinum toxin A directed to brow depressors. Injections consisted of 7 to 10 units of botulinum toxin A (Botox, Allergan, Irvine, Calif.) into selected brow depressor muscle (lateral orbicularis oculi) bilaterally. No patients withdrew for adverse effects. All patients were evaluated 2 weeks after treatment. The outcomes were measured by change in brow elevation along vertical axis extending from both midpupil and lateral canthus to the caudal row of brow hairs with eyes at neutral gaze and the head at Frankfort plane. Preintervention and postintervention brow height was measured by the primary clinical investigator. The average brow elevation from the midpupil observed after selected injection of brow depressors with botulinum toxin A was 1.02 mm (p = 0.038). The average brow elevation from the lateral canthus observed after selected injection of brow depressors with botulinum toxin A was 4.83 mm (p<0.0001). Significant temporal brow elevation occurs as the result of paralysis of brow depressors by using botulinum toxin A injection. This procedure may be considered an alternative to surgical brow elevation. PMID- 10724276 TI - Supratip deformity: a closer look. AB - Supratip deformity, a hallmark of a poorly executed rhinoplasty or an inauspicious healing, continues to plague the novice often and the experts on occasion. A clinical and histopathologic study was conducted to search for the surgical causes of this deformity and its histologic presentation. An organized, logical management program was then developed. Clinically, supratip fullness was observed in both primary (26 of 298 patients; 9 percent) and secondary (40 of 112 patients; 36 percent) rhinoplasty candidates. In primary patients, the deformity was the result of inadequate tip projection (pseudodeformity), an overprojected caudal dorsum, a combination of both, or cephalically oriented lower lateral cartilages. In secondary patients, the deformity was caused by an underresected or overresected caudal dorsum, overresected midvault, underprojected tip (pseudodeformity), or a combination of some of these factors. The histopathologic evaluation demonstrated significant fibrosis in the supratip soft tissue of 14 of 16 patients undergoing secondary rhinoplasty without the injection of triamcinolone acetonide and in only 13 of 23 patients who underwent primary rhinoplasty (p<0.05). A supratip deformity can be eschewed by proper resection of the caudal dorsum, avoidance of dead space, restoration of adequate projection to the nasal tip, and an approximation of the supratip subcutaneous tissue to the underlying cartilage using a supratip suture, hence eliminating the dead space. If the problem is noted shortly after surgery, in the presence of collapsible consistency of the supratip tissue and adequate projection, the treatment is taping the supratip tissue as often as it is practical. If no favorable response is elicited in 6 to 8 weeks, thejudicious injection of a small amount of triamcinolone acetonide (0.2 to 0.4 cc of 20 mg/cc) in the deep subcutaneous tissue (not in the dermis) is done. The injection is repeated in 4-week intervals until the desired effect is achieved. If supratip fullness is the consequence of inadequate cartilage resection or inadequate tip projection, surgical correction is needed. The recalcitrant soft-tissue excess in the supratip area is resected, and the subcutaneous soft tissue is approximated to the underlying cartilage. If the dorsum was previously overresected, a cartilage graft to the caudal dorsum or midvault will create an optimal dorsal frame and reduce the potential for a recurrent supratip deformity. PMID- 10724277 TI - Lip service for the stiff upper lip. AB - Lip augmentation procedures can restore volume and shape to the aging, thin upper lip, but some patients may develop problematic lip tightness. This stiff upper lip is manifested by a restricted smile and an adynamic central upper lip. We have had success in treating postreconstruction and postaugmentation stiff upper lip with a therapeutic device and treatment regimen. This therapy alleviated tightness and inability to smile. Also, the change in lip commissure-to commissure distance in repose and when smiling improved after treatment. PMID- 10724278 TI - The search for a youthful upper lip via laser resurfacing. PMID- 10724279 TI - Lip enhancement: surgical alternatives and histologic aspects. AB - This study included 66 consecutive patients, 58 women and 8 men, who underwent 86 surgical procedures on the lips during 1989-1998. Lip enlargement was performed in 59 patients, and lip reduction was performed in 7 patients. Indications were purely aesthetic in 61 cases and reconstructive in 5 cases. The following surgical techniques were used for lip augmentation: implantation of crystal silicone, polyacrylamide hydrogel, Gore-Tex tubes, autologous fat, and dermis-fat graft. A new instrument originally designed by the author, the dermis-fat graft passer, significantly speeded up and facilitated execution of the latter procedure. Other operations included V-Y plasty, lip lifting by buffalo horn excision, lip lengthening by frenulum plasty, and lip reduction by wavy tangential excision. Eighty-six percent of patients could be followed up; the mean length of follow-up was 4.2 years. Use of silicone microparticles (Bioplastique) was abandoned because of the tendency for lumping. Polyacrylamide gel is promising because of its ease of use, and Gore-Tex tubes are promising because of their ability to create and accentuate the Cupid's bow form for the upper lip. However, these products are new, and follow-up studies with longer observation times are needed to reach definite conclusions. Of these studied methods, autologous fat transplant was found to be particularly useful for enlargement and restoration in cases of age-related atrophy of the lips and perioral tissues. Dermis-fat grafting was the most efficient, versatile, and reliable method of lip enlargement. Long-term survival of transplanted autologous tissues was confirmed by histologic studies of biopsy specimens. PMID- 10724280 TI - The deep musculoaponeurotic system flap and extended open subperiosteal face lift combined with onlay cartilage grafts. PMID- 10724281 TI - Beauty and the body: the origins of cosmetics. AB - Ancient cultures were as preoccupied with the aesthetics of appearance as individuals are today. Dermabrasion for skin resurfacing has been performed with salt, pumice, ground grains, bone, and horn. Chemical peels have been performed with acids, metals, botanical extracts, or animal fats. Tattoos, ear piercing, makeup, skin treatments, and massages have existed for the past 5000 years. According to history, when the rise of more complex societies brought an ever increasing demand for cosmesis, perfumers, cosmetologists, barbers, and gentlewomen became pioneers, undertaking and developing the cosmetic practices that had evolved through the ages. With the consolidation of medical specialties concerned with the management of skin care, the scientific method has been applied to adapt and perfect many of the procedures that had been carried out with only empirical knowledge. To have a better perspective from which to envision future technical and technological developments, plastic surgeons should be familiar with the origins of cosmetics and some cosmetology practices that society demands. PMID- 10724282 TI - Aesthetic surgery economics: lessons from corporate boardrooms to plastic surgery practices. AB - Healthcare traditionally has been described as not conforming to the laws of economics. Consumers pay for aesthetic surgery directly, thus freeing it from the usual confounding factors and making it more likely to comply with the market forces explained by economics. Recent studies have demonstrated the ability of classic economics to analyze, predict, and optimize the financial environment of aesthetic surgery. This article describes economic principles and how they can be applied to aesthetic surgery. Some of the basic instruments of economics include the study of supply and demand, prices, and price elasticity; capital investments; communication and cooperation; and consumer cognitive limitations. Each of these tools offers plastic surgeons the opportunity to gain improved control of their financial environment. PMID- 10724283 TI - The mentalis muscle: an essential component of chin and lower lip position. PMID- 10724284 TI - Plastic surgery at the turn of the century: an opportunity for self-assessment. PMID- 10724285 TI - Ought GIDNOS get nought? Treatment options for nontranssexual gender dysphoria. PMID- 10724286 TI - Advantages of not being there. PMID- 10724287 TI - Becaplermin gel (PDGF-BB) as topical wound therapy. Plastic Surgery Educational Foundation DATA Committee. AB - Becaplermin gel is the first topical growth factor to demonstrate therapeutic efficacy in the healing of diabetic wounds. For diabetic patients who have poorly healing ulcers despite good perfusion and a reasonable trial of wound care, this product may be of considerable benefit. It should be tried for a 2-week time period and the results objectively assessed before continuation of therapy. PMID- 10724288 TI - The fallacy of the levator expansion theory. PMID- 10724289 TI - Stigmata: part I. PMID- 10724290 TI - Approach scar revisions after self-inflicted wounds with caution. PMID- 10724291 TI - Postoperative analgesia in augmentation mammaplasty. PMID- 10724292 TI - Treatment of cubital tunnel syndrome without electrodiagnosis and relationship to multiple crush syndrome. PMID- 10724293 TI - Pressure bandaging of the lower leg prior to minor surgery. PMID- 10724294 TI - Hypnosis and asthma: a critical review. AB - Asthma is among the most common chronic diseases of the western world and has significant effects on patients' health and quality of life. Asthma is typically treated with pharmaceutical products, but there is interest in finding nonpharmaceutical therapies for this condition. Hypnosis has been used clinically to treat a variety of disorders that are refractive to pharmaceutical-based therapies, including asthma, but relatively little attention has been given recently to the use of clinical hypnosis as a standard treatment for asthma. Significant data suggest that hypnosis may be an effective treatment for asthma, but it is premature to conclude that hypnosis is unequivocally effective. Studies conducted to date have consistently demonstrated an effect of hypnosis with asthma. More and larger randomized, controlled studies are needed. Existing data suggest that hypnosis efficacy is enhanced in subjects who are susceptible to the treatment modality, with experienced investigators, when administered over several sessions, and when reinforced by patient autohypnosis. Children in particular appear to respond well to hypnosis as a tool for improving asthma symptoms. PMID- 10724295 TI - Physical and psychological correlates of functioning in patients with chronic obstructive pulmonary disease. AB - We studied the contribution of coping and illness perceptions to outcome in patients with chronic obstructive pulmonary disease (COPD). In a longitudinal study, 64 patients completed the Medical Outcomes Study Instrument and the Illness Perception Questionnaire. Data on coping and severity of illness (spirometry) were also collected. Regression analyses showed that first-time illness perceptions and coping significantly contributed to the prediction of social functioning, mental health, health perceptions, total functioning score, and prediction of visits to the outpatient clinic and prescribed medication 1 year later. These results have important implications for the medical management of patients with COPD. PMID- 10724296 TI - Assessing pediatric clinical asthma practices and perceptions: a new instrument. AB - Over the past 20 years, the most substantial increases in prevalence, morbidity, and mortality of asthma have been observed among children aged 5-14 years. A survey instrument designed to measure clinical asthma management practices of primary care physicians was developed and evaluated. Study participants included 127 practitioners providing pediatric asthma care in inner-city communities in Baltimore, MD and Washington, DC. Study results found that the instrument assessed four separate dimensions of clinical assessments and five dimensions of physician perceptions. These dimensions should be considered in future research protocols and may be used to design tailored interventions to improve asthma care. PMID- 10724297 TI - Reproducibility, construct validity, and responsiveness of the "How Are You?" (HAY), a self-report quality of life questionnaire for children with asthma. AB - This study assesses the reproducibility, construct validity, and responsiveness of a new self-report quality of life questionnaire, the "How Are You?" (HAY), for 8-12-year-old children with asthma, which contains both a generic and a disease specific part. Two hundred twenty-eight children with asthma completed the HAY and the Child Attitude Toward Illness Scale (CATIS), while their parents monitored the actual asthma status; 80 children were measured three times in order to assess reproducibility and responsiveness; 296 healthy children completed the generic section of the HAY. Significant differences were found between children with asthma and healthy children, and among asthmatic children differing in actual asthma status. Reproducibility was adequate and supportive evidence was found for construct validity. Responsiveness was demonstrated by significant score changes for most dimensions in clinically changed children. The HAY seems useful for both discriminative and evaluative research in children with asthma. PMID- 10724298 TI - The antioxidative defense in asthma. AB - Asthma is a disease characterized by chronic airway inflammation. Generation of oxygen free radicals by activated inflammatory cells produces many of the pathophysiologic changes associated with asthma and may contribute to its pathogenesis. However, the activities of antioxidant enzymes and their relation with asthma have not been well defined. This study was performed to examine the activities of major intracellular antioxidants in mild asthmatic patients. Twelve asymptomatic mild asthmatic patients who never used any antiasthma medication and 13 age- and sex-matched healthy control subjects were selected. The activities of erythrocyte antioxidant enzymes, superoxide dismutase (SOD), catalase (CAT), and glutathione-peroxidase (GSH-Px) were measured spectrophotometrically. The mean SOD activity of asthmatic patients was found to be significantly lower than that of the controls (p < 0.05). There was no significant difference in CAT and GSH-Px activities between patients and controls (p > 0.05). Although the mechanisms underlying the association between asthma and antioxidant system are unclear, according to our findings, decreased antioxidant protection may contribute to the pathogenesis of mild asthma. PMID- 10724299 TI - Distribution and severity of bronchiectasis in allergic bronchopulmonary aspergillosis (ABPA). AB - The purpose of this paper was to quantitate the distribution and severity of computed tomography (CT) and radiographic findings in patients with allergic bronchopulmonary aspergillosis (ABPA), probable ABPA, and asthmatic controls. Chest radiographs and high-resolution CT images were evaluated in 19 patients with documented ABPA and 18 asthmatic controls. Ten patients with probable ABPA were also evaluated. On CT examination 17 patients (89%) with ABPA had central cystic or varicoid bronchiectasis in at least one lobe. One patient had no evidence for bronchiectasis. Three asthmatic patients (17%) had findings of cylindrical bronchiectasis. All 10 patients with probable ABPA had evidence of bronchiectasis on high-resolution CT (HRCT). The majority of patients with ABPA have diffuse disease at the time of diagnosis, manifested by central cystic and/or varicoid bronchiectasis in four or five lobes. Evaluation with HRCT can facilitate a diagnosis of ABPA and probable ABPA, allowing for earlier treatment which may prevent progression to fibrosis. PMID- 10724300 TI - Atopy and pulmonary function abnormalities in children with a history of acute bronchiolitis. AB - To assess the relationship between acute viral bronchiolitis and subsequent development of asthma, we studied retrospectively 97 index children, aged between 9 and 14 years, and 52 controls. The bronchiolitis group showed significantly lower values for mean expiratory flow at 50% of vital capacity (MEF50) higher incidence of atopy, and were more sensitive to methacholine than were controls, even if they had not shown recurrent wheezing episodes. It is suggested that an increased incidence of atopy, bronchial hyperresponsiveness, and reduced expiratory flows may be detectable in children with a history of acute bronchiolitis, regardless of the fact that they did not develop subsequent clinical symptoms suggestive of bronchial asthma. PMID- 10724301 TI - Reproducibility of bronchodilator response for a reservoir dry powder inhaler following routine clinical use. AB - The performance of dry powder inhaler (DPI) devices, particularly reservoir DPIs, may be influenced by environmental conditions. This study compared the bronchodilator efficacy and in vitro aerosol characteristics of salbutamol, delivered via a novel reservoir DPI (Clickhaler) and a conventional pressurized metered-dose inhaler (MDI) before and after use of the DPI in clinical practice. Following a screening visit, patients received cumulative doses of salbutamol (100, 200, and 400 microg) via DPI or MDI on separate days in a double-blind, crossover design before and after a 4-week period, during which the DPI was used as the patients' first-line bronchodilator. Lung function responses (forced expiratory volume in 1 sec [FEV1], forced vital capacity [FVC], and peak expiratory flow [PEF]) to salbutamol delivered by DPI and MDI and in vitro aerosol characteristics were not significantly different before and after the period of DPI patient use. DPI performance, assessed in vivo and in vitro, is maintained following an extended period of patient use. PMID- 10724302 TI - The allocation of family responsibility for asthma management tasks in African American adolescents. AB - The allocation of responsibilities for asthma management within African-American families was examined in 60 adolescents and their primary caretakers. Separate structured interviews were conducted with adolescents and primary caretakers, and perceptions of family management, adherence to asthma treatment regimen, and functional morbidity were assessed. Support for the primary hypothesis that higher levels of nonadherence and functional morbidity would be observed in families where caretakers overestimated the level of adolescent involvement in asthma self-care was found. Implications for family-based asthma management in ethnic minority adolescents are discussed. PMID- 10724303 TI - Medical ethics education must include students' moral dilemmas within the clinical setting. PMID- 10724304 TI - The adult learner: a misinterpreted species? PMID- 10724305 TI - The adult learner: a misinterpreted species? PMID- 10724306 TI - The adult learner: a misinterpreted species? PMID- 10724307 TI - A chance to improve our research partnership with the feds. PMID- 10724308 TI - VA medicine: the hidden treasure. PMID- 10724309 TI - Medicare competitive pricing: lessons learned in Phoenix and Kansas City. AB - Many lawmakers have embraced the idea of bringing more competition to the Medicare program as a way to achieve greater cost efficiency and provide more choice for beneficiaries. Advocates of this strategy believe Medicare should move away from its historical administrative pricing approach toward a competitive bidding process similar to those used by many private purchasers. Yet, despite support for the concept, every effort to test it in the marketplace has been met with strong opposition. Most recently, competitive bidding demonstration projects set to take place in Phoenix and Kansas City have been delayed by congressional action. Nevertheless, it is clear that those participating in the demonstration projects-notably private purchasers, consumers, health plans, and the Health Care Financing Administration-have already learned a great deal about the types of issues that must be considered if Medicare is truly to pay health plans in a competitive manner. This article explores the lessons learned so far in Phoenix and Kansas City, including design considerations such as plan participation, the standard benefit package, the bidding process, and the government contribution to premiums. It also examines the reasons for opposition to the project and their implications for broader Medicare reform efforts. PMID- 10724310 TI - Teaching evidence-based medicine: caveats and challenges. AB - Evidence-based medicine (EBM) is an important new paradigm of the medical profession. While the quantitative approach of EBM has its place, clinical medicine must take into account many subtleties that EBM fails to consider. In this article, the authors describe three caveats to this quantitative approach: (1) the detection of "maybe disease" (physiologic, anatomic, or histologic abnormalities that may not ever be overtly expressed in the patient's lifetime) inflates apparent diagnostic test performance; (2) probability revision is valuable primarily as an exercise to gain qualitative insights; and (3) patients are likely to be interested more than just central tendencies in making treatment decisions. They then consider some challenging questions facing clinician educators: how do they prepare students for situations where there is an absence of rigorous evidence? Should they teach students that the burden of proof lies in demonstrating efficacy or in demonstrating ineffectiveness? And what should they tell students about when to seek evidence to aid diagnostic and treatment decisions? PMID- 10724311 TI - Punishment: a story for medical educators. AB - The author recounts an incident of cheating by two first-year medical students, and how it was handled. One of the students, George, had waited until the last minute to write what he called a "stupid" paper that was required as the final examination in a health policy course. His classmate Ellen offered to write the paper for him, and other students also offered to help; no one pointed out that this would be unethical. After some hesitation, George was persuaded to accept Ellen's offer, and he turned in the paper as his own. The course director deduced the deception, and when the students were confronted, they immediately admitted what they had done, blamed only themselves, and said they had been "foolish." Subsequent events showed that the faculty saw the incident as a clear-cut case of cheating, whereas many students felt that George and Ellen's transgression was trivial when compared with plagiarizing a research paper or falsifying lab results on a patient's chart. Also, the faculty chose a more severe and long lasting punishment, one that many students did not agree with. The author believes that the faculty's refusal to give George and Ellen a clean slate after a reasonable time reflected a lack of forgiveness that is antithetical to the compassionate, forgiving role of physician-healer that medical education promotes. She concludes by explaining how this incident illustrates complex generational and cultural differences in moral reasoning and the selection of punishment, and the great emphasis that medical education places on knowing the facts rather than working creatively with ideas. PMID- 10724312 TI - Forum on the future of academic medicine: final session--implications of the information revolution for academic medicine. AB - The seventh and final meeting of the Association of American Medical Colleges' (AAMC's) Forum on the Future of Academic Medicine began December 4, 1998, with a talk by William W. Stead, MD, associate vice-chancellor for health affairs at Vanderbilt University Medical Center and director of its informatics center. Dr. Stead envisions a future in which informatics and information technology will place the consumer squarely in the center of the system, empowered with greater knowledge of health care; he gave three short scenarios to illustrate future typical interactions of consumers with the system. He then discussed the implications for academic medicine. For example, academic medical centers (AMCs) could become the information providers and quality assurance hubs of their regions. Various participants questioned some of the speaker's claims (one asserting that there would be serious complications if clinical information were made available to patients). The second speaker, Valerie Florance, PhD, director of the AAMC's better-health@here.now program, discussed her program, whose purpose is to explore the ways medical schools and teaching hospitals can best use information technology and the Internet in the coming decade to improve individual and community health. Nothing in the ensuing discussion indicated that the participants believed that academic medical centers would be spared painful dislocations if they were to embark on a road of institutional reform to respond to the pressures of the new and more competitive global economy. Greater awareness of this not-necessarily-welcomed message may be one of the lasting legacies of the forum. PMID- 10724313 TI - Core competencies for the care of older patients: recommendations of the American Geriatrics Society. The Education Committee Writing Group of the American Geriatrics Society. AB - The aging of the U.S. population has led many organizations to call for an increase in the amount of clinical geriatrics training in medical education. A subcommittee of the American Geriatrics Society's Education Committee was assigned the task of defining core competencies for geriatrics education in medical schools. The subcommittee reviewed the available literature, surveyed selected programs in geriatrics education, and sought input from experts in geriatrics education. They then defined the core knowledge, attitudes, and skills students must develop to care for older people. This article summarizes these core competencies, which medical educators may find useful in developing new curricula on aging or in evaluating existing curricula. PMID- 10724314 TI - Medicine and the arts. The woman who walked into doors. PMID- 10724315 TI - Effectiveness of problem-based learning curricula: research and theory. AB - PURPOSE: This article provides a critical overview of problem-based learning (PBL), its effectiveness for knowledge acquisition and clinical performance, and the underlying educational theory. The focus of the paper is on (1) the credibility of claims (both empirical and theoretical) about the ties between PBL and educational outcomes and (2) the magnitude of the effects. METHOD: The author reviewed the medical education literature, starting with three reviews published in 1993 and moving on to research published from 1992 through 1998 in the primary sources for research in medical education. For each study the author wrote a summary, which included study design, outcome measures, effect sizes, and any other information relevant to the research conclusion. RESULTS AND CONCLUSION: The review of the literature revealed no convincing evidence that PBL improves knowledge base and clinical performance, at least not of the magnitude that would be expected given the resources required for a PBL curriculum. The results were considered in light of the educational theory that underlies PBL and its basic research. The author concludes that the ties between educational theory and research (both basic and applied) are loose at best. PMID- 10724316 TI - A comparison of standard-setting procedures for an OSCE in undergraduate medical education. AB - PURPOSE: To compare four standard-setting procedures for an objective structure clinical examination (OSCE). METHODS: A 12-station OSCE was administered to 84 students in each of the final (fourth-) year medical classes of 1996 and 1997 at Dalhousie University Faculty of Medicine. Four standard-setting procedures (Angoff, borderline, relative, and holistic) were applied to the data to establish a cutoff score for a pass/fail decision. RESULTS: The procedures yielded highly inconsistent results. The Angoff and borderline procedures gave similar results; however, the relative and holistic methods gave widely divergent results. The Angoff procedure yielded results reliable enough to use in decision making for a high-stakes examination, but would have required more judges or more stations. CONCLUSIONS: The Angoff and borderline procedures provide reasonable and defensible approaches to standard setting and are practical to apply by non psychometricians in medical schools. Further investigation of the other procedures is needed. PMID- 10724317 TI - Caring for medical students as patients: access to services and care-seeking practices of 1,027 students at nine medical schools. Collaborative Research Group on Medical Student Healthcare. AB - PURPOSE: The personal health care of medical students is an important but neglected issue in medical education. Preliminary work suggests that medical student-patients experience special barriers to health care services and report problematic care-seeking practices that merit further inquiry. METHOD: A self report questionnaire was piloted, revised, and distributed to students at nine medical schools in 1996-97. The survey included questions regarding access to health services, care-seeking practices, and demographic information. RESULTS: A total of 1,027 students participated (52% response rate). Ninety percent reported needing care for various health concerns. Fifty-seven percent did not seek care at times, in part due to training demands, and 48% had encountered difficulties in obtaining care. A majority had received treatment at their training institutions, and students commonly pursued informal or "curbside" care from medical colleagues. Almost all participants (96%) were insured. Differences in responses were associated with level of training, gender, and medical school. CONCLUSION: Medical schools shoulder the responsibility not only of educating but also of providing health services for their students. Students encounter barriers to care and engage in problematic care-seeking practices. Greater attention to issues surrounding medical student health may benefit students and their future patients. PMID- 10724318 TI - Specialty choices of students who actually have choices: the influence of excellent clinical teachers. AB - PURPOSE: To determine the influence of the quality of attending physicians and residents on the specialty choices of excellent medical students, who actually have a broad choice of specialties. METHOD: In 1993-94 and 1994-95, 169 third year students at the University of Kentucky College of Medicine were randomly assigned to two one-month rotations on general medicine inpatient wards. At the end of each rotation, the students confidentially evaluated the attending physician and the supervising resident (different for each rotation) with whom they had worked. Data were collected for 62 attending physicians and 89 residents. The authors analyzed the influences of the "best" and "worst" clinical instructors (those rated in the top and the bottom 20% by all students with whom they had worked over the two years) on "excellent" medical students (the 52 students whose USMLE I scores were in the top 30% of their class). RESULTS: Using regression approaches from the general linear model, the authors found that independent predictors of internal medicine residency choice for excellent medical students were exposure to highly rated internal medicine attendings (p = .02) and residents (p = .03). Nine of 29 (30%) of the excellent students who worked with a "best" medicine clinical instructor chose an internal medicine residency, while none of the 23 excellent medical students who did not work with a "best" medicine clinical instructor did so. The authors found no correlation in students' ratings of their pairs of attendings and residents, suggesting that rater bias did not explain the results. CONCLUSION: Better medical students who work with the best internal medicine attending physicians and residents in their internal medicine clerkship are more likely to choose an internal medicine residency. PMID- 10724319 TI - Obstacles to promotion? Values of women faculty about career success and recognition. Committee on the Status of Women and Minorities, Virginia Commonwealth University, Medical College of Virginia Campus. AB - PURPOSE: To assess attitudes of female faculty about career progress, resources for career development, and values related to academic success and recognition. METHOD: In 1997, the authors surveyed all faculty at Virginia Commonwealth University School of Medicine and its associated Veterans Affairs Medical Center. RESULTS: Of 918 faculty, 567 (62%) responded to the survey; 33% of the respondents were women. Compared with men, women faculty were less likely to be tenured or at the level of professor, spent more time in clinical activities, had less time for scholarly activity, and reported slower career progress. Women were more likely to report that promotion and tenure criteria had not been reviewed with them. Significant differences were found between female physicians and non physician faculty; female physicians reported the least time for scholarly activities and poorest understanding of promotion and tenure criteria. When the authors asked faculty how they valued certain indicators of career success, women were less likely to value leadership than were men. Female physicians were less likely to value scholarship and national recognition as indicators of their career success. CONCLUSION: This survey found important differences in career progress of male and female faculty, with women reporting less time for career development. In addition, there were differences in values related to career success and recognition, which were most pronounced for female physicians. These differences may have an important impact on promotion for women in general and particularly for female physicians. PMID- 10724320 TI - Residents and medical students noting the chief complaint during verbal presentations. AB - The chief complaint is an important element of the medical history. Over a one month period in 1999, the author observed 55 presentations of house officers and medical students on an inpatient ward at Miami Valley Hospital, an affiliate of Wright State. House officers noted the chief complaint in 49% of their presentations; students, in 61% of theirs. Educators need to stress the importance of the chief complaint during verbal case presentations. PMID- 10724321 TI - Pilot study of a rating instrument for medical education Web sites. AB - Standardized evaluation criteria would be useful for sorting through the huge volume of medical educational information now available on the Internet. The authors developed and pilot-tested a simple rating instrument. Using this instrument, students identified preferred Web sites and indicated that speed was particularly important in their assessments. PMID- 10724322 TI - Clinical problem analysis (CPA): a systematic approach to teaching complex medical problem solving. AB - The authors describe and discuss clinical problem analysis (CPA), an approach to solving complex clinical problems. They outline the five steps of the CPA model and the essential elements of each step. Next, they discuss the value of CPA's content-independent (methodical) approach and argue that teaching students to use CPA will enable them to avoid some common diagnostic reasoning errors and pitfalls. Finally, they compare CPA with two existing approaches to clinical problem solving. PMID- 10724323 TI - A consortium-based research education program for residents. AB - With growing pressures to consolidate and reorganize health care delivery systems, graduate medical education (GME) consortia can draw faculty from affiliated members to assemble educational programs. The authors report on consortium-based research education seminars of a quality that many residency programs would be unable to develop and support on their own. Drawing a diverse faculty from consortium members and area universities, the OHEP Center for Medical Education's annual Research Workshop Series focuses on the design of research projects; data analysis and hypothesis testing; and written and oral presentation of scientific research. Each spring, OHEP sponsors a research forum in which the best research projects from consortium members are presented by the resident-researchers, who compete for recognition and prize money. Further, of the 128 presentations made thus far at the annual OHEP Research Forum, 25% were subsequently published. The consortium's research education program has been well received by residents, is cost-effective, and is an integral component of the research curricula of many area residency programs. Including research training in GME provides residents an opportunity to become more competitive for fellowship, faculty, and leadership positions. PMID- 10724324 TI - The Managed Care Education Clearinghouse. AB - The authors surveyed all 125 allopathic medical schools to determine the number of schools that had implemented a formal curriculum in managed care and how many had a substantial interest in a Web-based clearinghouse for managed care curricular resources. They describe the results of their survey and the Web site they developed, the Managed Care Education Clearinghouse. PMID- 10724325 TI - Measuring contributions to the research mission of medical schools. AB - The authors of this article, who were the members and staff of a research panel formed by the AAMC as part of its mission-based management initiative, reflect on the growing interest in quantitative information in the management of the research mission of medical schools. They note the serious limitations of any such system of measures for research, particularly its inability to represent directly the quality of the research effort. Despite these concerns, the authors acknowledge that leaders in academic medicine have always used quantitative measures in one form or another to compare performance or assess progress. Two factors appear to be driving increases in this practice: (1) the need to demonstrate to institutional stakeholders that resources are being used wisely and that the school's performance justifies continued investment in the research mission; and (2) the need to fashion an economic strategy to manage precious institutional resources, particularly research space. Given these realities, the authors offer guidelines for the proper development and use of measures to assess contributions by faculty, departments, and institutions to the research mission. They also comment on the measures most commonly used in four areas: grants and other revenue-generating activities; publications; faculty members' research reputation and contributions to the national research enterprise; and support to the general research mission of the school. The authors conclude that quantitative information can help institutional leaders in important management decisions. However, the potential for misuse is great. The key is always to regard this information as an aid to judgment, not a substitute for it. PMID- 10724326 TI - Antioxidant properties of a North American ginseng extract. AB - A North American ginseng extract (NAGE) containing known principle ginsenosides for Panax quinquefolius was assayed for metal chelation, affinity to scavenge DPPH-stable free radical, and peroxyl (LOO*) and hydroxyl (*OH) free radicals for the purpose of characterizing mechanisms of antioxidant activity. Dissociation constants (Kd) for NAGE to bind transition metals were in the order of Fe2+ > Cu2+ > Fe3+ and corresponded to the affinity to inhibit metal induced lipid peroxidation. In a metal-free linoleic acid emulsion, NAGE exhibited a significant (p < or = 0.05) concentration (0.01-10 mg/mL) dependent mitigation of lipid oxidation as assessed by the ammonium thiocyanate method. Similar results were obtained when NAGE was incubated in a methyl linoleate emulsion containing haemoglobin catalyst and assessed by an oxygen electrode. NAGE also showed strong DPPH radical scavenging activity up to a concentration of 1.6 mg/mL (r2 = 0.996). Similar results were obtained for scavenging of both site-specific and non site specific *OH, using the deoxyribose assay method. Moreover, NAGE effectively inhibited the non site-specific DNA strand breakage caused by Fenton agents, and suppressed the Fenton induced oxidation of a 66 Kd soluble protein obtained from mouse brain over a concentration range of 2-40 mg/mL. These results indicate that NAGE exhibits effective antioxidant activity in both lipid and aqueous mediums by both chelation of metal ions and scavenging of free radicals. PMID- 10724327 TI - SERCA pump isoform expression in endothelium of veins and arteries: every endothelium is not the same. AB - Endothelium from rat aorta expresses sarco/endoplasmic reticulum Ca2+(SERCA) pump gene SERCA3 where as the smooth muscle expresses SERCA2. This has led to the postulate that vascular endothelium expresses SERCA3. To test this postulate, we examined the SERCA2 and SERCA3 mRNA expression in endothelium and smooth muscle dissected from coronary artery, coronary vein, aorta and vena cava of pig. Smooth muscle from all arteries and veins expressed only the SERCA2 mRNA. Endothelium from coronary artery, coronary vein and aorta expressed both SERCA2 and SERCA3 mRNA but the endothelium from vena cava did not express SERCA3 mRNA although it expressed SERCA2. These observations support the postulate that vascular endothelium expresses SERCA3 but the affirmation is equivocal because vena cava endothelium does not express SERCA3. PMID- 10724328 TI - Sarco/endoplasmic reticulum Ca2+-pump isoform SERCA3a is more resistant to superoxide damage than SERCA2b. AB - Endo/sarcoplasmic reticulum (ER) Ca2+-pumps are important for cell survival and communication but they are inactivated by reactive oxygen species (ROS). We have previously reported that the Ca2+-pump isoform SERCA3a is more resistant than SERCA2b to damage by peroxide. Since peroxide and superoxide differ in their redox potentials, we now report the effects of superoxide on the two Ca2+-pump isoforms. We isolated microsomes from HEK293 cells transiently transfected with SERCA2b or SERCA3a cDNA. We exposed these microsomes to superoxide which was generated using xanthine plus xanthine oxidase and catalase to prevent accumulation of peroxide due to superoxide dismutation. Superoxide damaged the Ca2+-transport activity of both isoforms but SERCA3a was damaged at higher concentrations of superoxide and upon longer periods of exposures than was SERCA2b. Thus the SERCA3a isoform is more resistant than SERCA2b to inactivation by both superoxide and peroxide. PMID- 10724329 TI - Increased T-type Ca2+ channel activity as a determinant of cellular toxicity in neuronal cell lines expressing polyglutamine-expanded human androgen receptors. AB - We have analyzed Ca2+ currents in two neuroblastoma-motor neuron hybrid cell lines that expressed normal or glutamine-expanded human androgen receptors (polyGln-expanded AR) either transiently or stably. The cell lines express a unique, low-threshold, transient type of Ca2+ current that is not affected by L type Ca2+ channel blocker (PN 200-110), N-type Ca2+ channel blocker (omega conotoxin GVIA) or P-type Ca2+ channel blocker (Agatoxin IVA) but is blocked by either Cd2+ or Ni2+. This pharmacological profile most closely resembles that of T-type Ca2+ channels [1-3]. Exposure to androgen had no effect on control cell lines or cells transfected with normal AR but significantly changed the steady state activation in cells transfected with expanded AR. The observed negative shift in steady-state activation results in a large increase in the T-type Ca2+ channel window current. We suggest that Ca2+ overload due to abnormal voltage dependence of transient Ca2+ channel activation may contribute to motor neuron toxicity in spinobulbar muscular atrophy (SBMA). This hypothesis is supported by the additional finding that, at concentrations that selectively block T-type Ca2+ channel currents, Ni2+ significantly reduced cell death in cell lines transfected with polyGln-expanded AR. PMID- 10724330 TI - Antioxidant status, lipid peroxidation, mixed function oxidase and UDP-glucuronyl transferase activities in livers from control and DOCA-salt hypertensive male Sprague Dawley rats. AB - The effects of DOCA-salt hypertensive treatment on hepatic glutathione-dependent defense system, antioxidant enzymes, lipid peroxidation, mixed function oxidase and UDP-glucuronyl transferase activities were investigated in male Sprague Dawley rats. Compared with controls, DOCA-salt hypertensive rats had lower body weights (linked to liver hypertrophy). Mixed function oxidase and p-nitrophenol UGT activities were not affected by the treatment but a significant lower rate of the glucuronoconjugation rate of bilirubin (p < 0.001) was observed in DOCA-salt hypertensive rats. While cytosolic glutathione contents and glutathione reductase activity were not affected, glutathione peroxidase (p < 0.001), glutathione transferase (p < 0.001) and catalase (p < 0.01) activities were decreased and associated with higher malondialdehyde contents (p < 0.001) in treated rats. The imbalance in liver antioxidant status (increasing generation of cellular radical species), associated with increases in lipid peroxidation, suggests that oxidative stress might be directly related to arterial hypertension in DOCA-salt treated male Sprague Dawley rats. PMID- 10724332 TI - Catecholamines-evoked cytosolic Ca2+ rise in endothelial cells from bovine adrenal medulla. AB - The effects of catecholamines on intracellular Ca2+ concentrations ([Ca2+]i) in single acutely dissociated bovine adrenal medulla endothelial cells (BAMECs) were measured using the intracellular fluorescent probe Fluo-3 AM. 100 microm epinephrine or norepinephrine induced a biphasic [Ca2+]i rise with an initial peak followed by a delayed phase. 10 microm phenylephrine (alpha1-adrenergic agonist) caused a [Ca2+]i rise similar to that evoked by catecholamines. The increase in [Ca2+]i induced by 10 microm phenylephrine was reverted by 10 microm phenoxybenzamine (alpha-adrenergic antagonist). Neither isoproterenol (beta adrenergic agonist) nor clonidine (alpha2-adrenergic agonist) induced [Ca2+]i rise. The initial peak was insensitive to zero external Ca2+ and it was abolished after Ca2+ internal storages were emptied by 10 mM caffeine. The delayed phase was reduced to near zero by external Ca2+ removal. These results indicate that BAMECs possess alpha1-adrenergic receptors associated to both the release of caffeine-sensitive intracellular Ca2+ stores and the entry of extracellular Ca2+. We suggest that chromaffin cell secretion may activate BAMECs in vivo through an increase in [Ca2+]i which could induce the secretion of vasoactive factors allowing a rapid entry of hormones into the circulation. PMID- 10724331 TI - Transcriptional regulation of cyclooxygenase-2 in the human microvascular endothelial cell line, HMEC-1: control by the combinatorial actions of AP2, NF-IL 6 and CRE elements. AB - Interleukin-1beta (IL-1) is a potent inducer of cyclooxygenase-2 (COX-2) and prostaglandin biosynthesis in many types of cells, yet little is known about the molecular mechanisms regulating IL-1 mediated prostanoid biosynthesis in the endothelium of the microvasculature. Therefore, we examined the cis- and trans acting factors regulating IL-1-induced COX-2 expression in the human microvascular endothelial cell line, HMEC-1. IL-1 enhanced steady state levels of COX-2 protein and mRNA synthesis by approximately 2-fold which preceded a 2-fold increase in PGF(alpha) biosynthesis. Expression of a series of COX-2 promoter luciferase constructs in IL-1 treated HMEC-1 cells revealed that the 'full length' (-1432/+59 bp) promoter was 10 times more active than the SV-40 promoter/enhancer and that it could be further activated by IL-1. Surprisingly however, all except for the shortest COX-2 promoter construct retained the ability to respond to IL-1 and luciferase activity driven by -191/+59 bp COX-2 promoter was as responsive to IL-1 as the full-length promoter. Moreover, site directed promoter mutagenesis and electophoretic mobility shift assays (EMSA) indicate that the combinatorial actions of AP2, NF-IL6, and CRE elements are critical for both constitutive and IL-1-inducible COX-2 promoter activity. Understanding the mechanism(s) regulating COX-2 gene expression and prostaglandin biosynthesis in the microvasculature has important implications with regard to inflammation and angiogenesis in vivo. PMID- 10724334 TI - Purification and characterization of cationic chymotrypsin from the pancreas of the Arabian camel (Camelus dromedarius). AB - This study reports on the purification and characterization of a cationic enzyme with chymotryptic activity from camel pancreas. The enzyme was purified 52-fold in a 48% yield by a three-step chromatographic procedure consisting of anion exchange, cation-exchange and affinity chromatographies. The purified enzyme was homogeneous on gel isoelectric focusing and on SDS gel electrophoresis. Its isoelectric point was estimated to be 7.3 and its molecular mass was found to be 23,600 Da. The enzyme was identified as a cationic chymotrypsin according to its physiochemical properties, substrate specificity and susceptibility to inhibition. It was active towards esters of aromatic amino acids but much less active towards a leucine ester. In all cases, the k(cat) values of the camel enzyme were less than the corresponding values of bovine chymotrypsin A. It also showed a lower level of kininase activity. Camel chymotrypsin was more susceptible than its bovine equivalent to inhibition by soybean trypsin inhibitor and aprotinin. It showed the same pH optimum as bovine chymotrypsin A for its esterolytic activity, but was more dependent on CaCl2 for long-term stability. PMID- 10724333 TI - Transcriptional activation of p21WAF1 by PTEN/MMAC1 tumor suppressor. AB - The recently discovered tumor suppressor gene PTEN has been found mutated in many types of advanced tumors. When introduced into tumor cells that lack the wild type allele of the gene, PTEN was able to suppress the growth of these cells. Here, we have analyzed how PTEN might alter cell cycle-regulatory controls to achieve this growth-inhibitory effect. We found that overexpression of PTEN stimulates the synthesis of three inhibitors of cyclin-dependent kinases, p21WAF1, p27KIP1, and p57KIP2. This effect is very specific, as the expression of other components of the cell cycle engine, various cyclins and cyclin-dependent kinases, is not affected. For p21WAF1 we show that this induction is due to the p53-independent transcriptional activation of its promoter. In addition, increased expression of PTEN rendered the cells more sensitive to apoptotic cell death. Therefore, our data suggest a two-fold mechanism of growth inhibition by PTEN: one that acts via the increased expression of CKIs such as p21WAF1, and another that augments the cellular propensity for apoptotic cell death. PMID- 10724335 TI - Gastropathy and defense mechanisms in common bile duct ligated portal hypertensive rats. AB - Portal hypertensive gastropathy is associated with a broad spectrum of gastric mucosal damage inspite of decreased gastric acid secretion, suggestive of compromised endogenous protective mechanisms. To determine the mechanisms of damage in portal hypertensive gastropathy we measured lipid peroxidation, glutathione, antioxidant and lysosomal enzymes in gastric mucosal homogenates from male Wistar rats with elevated intrasplenic pulp pressure, eighteen days after common bile duct ligation. Thiobarbituric acid-reactive substances and lysosomal enzymes (beta-glucuronidase and acid phosphatase) were increased in the common bile duct ligated group as compared to the sham-operated group. The levels of antioxidant defense enzymes, superoxide dismutase, glutathione peroxidase, catalase and glutathione were decreased as compared to the sham-operated controls. Pre-operative vitamin E administration decreased mucosal lipid peroxidation increased the levels of antioxidant defense enzymes and lowered the lysosomal enzymes. The plasma vitamin E levels in this group were lower when compared to animals receiving it post-operatively. In conclusion, free radical and lysosomal enzyme mediated damage may play a role in portal hypertensive gastropathy. PMID- 10724336 TI - Estrogen controls expression and bioresponse of 1,25-dihydroxyvitamin D receptors in the rat colon. AB - Estrogen receptors are extensively expressed in the gastrointestinal tract, however their physiological role is not clear yet. Estrogen and 1,25 dihydroxyvitamin D [1,25(OH)2D3] apparently share common activities in the intestine such as growth-suppressing effects on the colonic mucosa and positively influence intestinal calcium absorption. In view of our previous studies showing up-regulation of vitamin D receptors (VDR) in the duodenal mucosa and in osteoblasts, the present study was designed to address a possible interaction between estrogen and the vitamin D endocrine system in the colonic mucosa. Three groups of female rats were studied: sham operated ('Sham'), ovariectomized ('OVX'), and ovariectomized estrogen-treated ('OVX+E'). VDR gene expression was assessed by Northern blot analysis, VDR protein expression was assessed by ligand binding assays, and Western-blotting. Endogenous 1,25(OH)2D3 bioactivity in colonic mucosal extracts was assessed by alkaline phosphatase activity and calbindin-9kD mRNA expression. Northern blots revealed marked increase in band intensity corresponding with the VDR mRNA product in 'Sham' or 'OVX+E' vs. 'OVX'. In ligand-binding experiments, 1,25(OH)2D3 was shown to bind specifically to a single class of receptors in extracts obtained from each of the groups (Kd--0.03 nM). The maximal VDR binding capacity of colonic mucosal extracts was 203 +/- 23 fmol/mg protein in 'Sham', 362 +/- 41 in 'OVX+E' and 102 +/- 15 in 'OVX' ('Sham' or 'OVX+E' vs. 'OVX', p < 0.001). Western-blot analysis also revealed higher VDR protein expression in the estrogen-exposed animals. Alkaline phosphatase activity and calbindin-9kD mRNA expression were significantly higher in colonic mucosal extracts from estrogen-exposed rats. Estrogen increases VDR gene transcript level, protein expression and endogenous 1,25(OH)2D3 bioactivity in colonic mucosa, which may suggest that some of the estrogen activities in the colonic mucosa, such as its growth-suppressing effect, could be mediated, at least in part, by an increase in colonic mucosa responsiveness to endogenous 1,25(OH)2D3. PMID- 10724337 TI - Synthesis and phosphorylation of androgen receptor of the mouse brain cortex and their regulation by sex steroids during aging. AB - To examine the synthesis and phosphorylation of androgen receptor (AR) and their regulation by sex steroids, adult (24 weeks) and old (65 weeks) male and female mice were gonadectomized and administered with testosterone and estradiol. AR amount, synthesis and phosphorylation were measured in the brain cortex by immunoblotting and immunoprecipitation using antibody raised against rat AR transactivation domain (TAD) which was expressed in E. coli as a fusion protein. We found that the amount of AR was high in adult and declined in old mice of both sexes. Administration of testosterone and estradiol significantly down-regulated the level of AR in old male and adult female. Similarly, the rate of AR synthesis also declined with age. Exogenous treatment of gonadectomized mice with testosterone and estradiol reduced the extent of synthesis significantly in all groups except in old female. No sex-dependent variation was noticed either in the level or synthesis of AR. In contrast, the extent of phosphorylation was higher in old mice of both sexes as compared to their adult counterparts. Testosterone and estradiol supplementation resulted in remarkable increase in AR phosphorylation in all groups. Thus it is evident from our findings that the amount and synthesis of AR decrease but phosphorylation of AR increases in the brain cortex with advancing age of mice and they are regulated by testosterone and estradiol in age- and sex-specific manner. PMID- 10724338 TI - An accumulation of proteoglycans in scarred fascia. AB - A little is known about proteoglycan (PG) changes, occuring in the course of scarring of tissues another than skin. The aim of present study was biochemical characterization of glycosaminoglycans (GAGs) and proteoglycans (PGs) of normal and scarred fascia. Samples of normal fascia lata were taken at autopsy from 23 individuals and samples of scarred fascia lata were removed from 23 patients at reoperations for femoral fracture. The obtained tissues were divided into two samples: first of them was submitted to GAG isolation and the second one to PG isolation. GAGs were extracted by extensive papain digestion followed by the fractionation using cetylpyridinium chloride. In order to qualitative and quantitative characterization GAGs were submitted to electrophoresis on cellulose acetate before and after treatment with enzymes, specifically depolymerizing some kinds of GAGs. PGs were extracted using 4 M guanidine HCl followed by purification by forming complexes with Alcian blue. PGs were submitted to gel permeation chromatography on Sepharose 4B. In order to obtain core proteins PGs were depolymerized with chondroitinase ABC. The purified PGs and their core proteins were separated with sodium dodecyl sulphate/polyacrylamide gel electrophoresis (SDS/PAGE). It was found that total GAGs content was significantly elevated in scarred fascia. Both types of fascia contained chondroitin-, dermatan- and heparan sulphates and hyaluronic acid. Dermatan sulphates (DS) were the predominant GAGs of normal and scarred fascia. The contents of all GAG types were increased in scarred fascia. Both types of fascia contained two kinds of dermatan sulphate proteoglycans (DSPGs); first being similar to biglycan and the second one similar to decorin, as it was judged by molecular weight of their native molecules and core proteins as well as type of GAG components. Densitometric analysis showed that decorin is a predominant DSPG in both fascia types, but in scarred tissue the ratio of biglycan to decorin is considerably higher. Moreover, in scarred fascia a large chondroitin sulphate proteoglycan (CSPG) was also observed. The obtained results have shown that the scar formation is accompanied by quantitative and qualitative alterations in GAGs/PGs resembling those observed in hypertrophic skin scars. The biochemical modification of the scarred fascia lata may partly explain the clinically manifested damage to biomechanical properties of this tissue. PMID- 10724339 TI - Modulation of adherence and chemotaxis of macrophages by norepinephrine. Influence of ageing. AB - We evaluated the in vitro effect of norepinephrine (NE), over the range of concentrations between 10(-12) M and 10(-3) M, on adherence (to plastic surfaces) and chemotaxis (in a Boyden chamber) of peritoneal macrophages from BALB/c mice of different ages: young (12 weeks), adult (22 weeks), mature (48 weeks) and old (72 weeks). Increased adherence was induced by 10(-12) M of NE in macrophages from young, adult, mature and old mice. Also, 10(-9) M stimulated adherence in old animals, 10(-5) M in mature mice, and 10(-3) M in both young and old mices. With respect to chemotaxis, the low concentration of NE (10(-12) M) was stimulatory only in young and adult animals, higher concentrations (10(-5) M and 10(-7) M) were inhibitory for macrophages from mature and old animals, and the highest concentration of NE (10(-3) M) stimulated this capacity of macrophages only in young and mature animals. The conclusion is that while the mobility of macrophages to the focus of infection (i.e. chemotaxis) is stimulated by low concentrations of NE (10(-12) M) only in young-adult animals, this neurotransmitter induces a decline in this capacity in mature and old mice at high concentrations (10(-5) M-10(-7) M). Also, macrophages from old animals have lost the capacity to respond to pharmacological (10(-3) M) concentrations of NE. The lower capacity of response to NE by macrophages from old animals possibly contributes to immunosenescence. PMID- 10724340 TI - Peroxisome distribution along the crypt-villus axis of the guinea pig small intestine. AB - Peroxisomes and peroxisomal enzyme expression were investigated biochemically and morphometrically in guinea pig intestinal epithelial cells at different stages of their migration along the crypt-villus axis. Epithelial cells were sequentially isolated along the axis and the specific activities of the peroxisomal enzymes catalase and acyl-CoA oxidase were found to be significantly higher in differentiated and mature cells situated at the villus tip and stem than in the crypt. Conversely, 1-alk-1'enyl, 2-acyl phospholipid (plasmalogen) concentration in the crypt and middle villus was significantly higher than in villus tip cells. Assay of alkyl DHAP synthase and fatty acyl CoA reductase (enzymes responsible for the production of plasmalogen precursors) showed no correlating activity gradient with plasmalogen concentration. Morphometric analysis revealed that peroxisomes were present even in the most immature stem cells, however, their number and volume and surface densities increased as the epithelial cell developed as did the proportion of elongated and vermiform peroxisomes to spherical structures. Senescent cells at the tip of the villus, however, showed a dramatic decrease in number of peroxisomes per cell possibly due to cellular degradation. We conclude that the peroxisomal compartment of the guinea pig small intestinal epithelial cell develops as a function of cell development possibly reflecting adaptation to maximise its metabolic capacity. PMID- 10724341 TI - Involvement of PL-D in the alternate signal tranduction pathway of macrophages induced by an external stimulus. AB - The alternate pathway of signal transduction via hydrolysis of phosphatidylcholine, the major cellular phospholipid, has been investigated in murine peritoneal macrophages. A sustained formation of diacylglycerol, is preceded by an enhanced production of phosphatidic acid, when the macrophages were given a stimulus with 12-O-tetradecanoyl phorbol-13-acetate for sixty minutes. Production of choline and choline metabolites are significantly increased too. Propranolol, which inhibits phosphatidate phosphohydrolase, the enzyme responsible for conversion of phosphatidic acid to diacylglycerol, can effectively block the formation of diacylglycerol. Inhibition of protein kinase C either by its inhibitors, staurosporine and H-7 or by depletion, apparently affect the generation of the lipid products. Moreover, based on the results of transphosphatidylation reaction, involvement of a phospholipase D in the phosphatidylcholine-hydrolytic pathway in macrophages is predicted. These observations support the view that probably the phorbol ester acting directly on protein kinase C of the macrophages activate their phosphatidylcholine-specific phospholipase D to allow a steady generation of second messengers, to enable them to participate in the cell signalling process in a more efficient manner than those generated in the phosphoinositide pathway of signal transduction. PMID- 10724342 TI - Cloning of MafG homologue from the rat brain by differential display and its expression after hypercapnic stimulation. AB - The ventral medullary surface (VMS) is a site of the medullary chemoreceptor neurons which sense excess protons (H+) derived from hypercapnia and facilitate respiration. We hypothesized that expression of genes involved in H+-sensitivity is higher in the VMS than in other central nervous system areas. By using the differential display technique, we differentiated the mRNAs of VMS neurons from those of cerebral cortical neurons. Seventeen clones of interest were isolated, and sequence analysis revealed that one of these clones had an encoding nuclear transcription factor, MafG. MafG is a member of Maf protein family, and the founding member of the family (v-Maf) was originally discovered as the transduced transforming component of avian musculoaponeurotic fibrosarcoma virus, AS42. The rat MafG was composed of 162 amino acid residues and was conserved among the primary structures of various species. Expression of rat mafG mRNA is high in the VMS, heart and skeletal muscle while the cerebral cortex, cerebellum, liver, stomach and intestine show moderate expression. To determine whether the expression of mafG mRNA is induced by hypercapnic stimulation, 7% CO2 in air was inhaled to rats for 5 min. We found that the hypercapnic stimulation induced the gene expression of mafG. These results suggest that MafG may be involved in H+ sensitivity and respiratory regulation in the VMS. PMID- 10724343 TI - Signal transduction mechanism in human neutrophil: comparative study between the zeta and beta-protein kinase isotypes. AB - Role of protein kinase C (PKC) isotypes in the regulation of neutrophil function are not clearly known. In the present study we purified the beta-PKC and zeta-PKC isotypes from human neutrophil. Both the isotypes are immunoreactive only to their respective antibodies. Zeta-PKC was further confirmed by RT-PCR using specific primer. Co-factor requirements for both the kinases were found to be different when DG and ceramide were used as second messenger. Selective substrate specificities were determined for both beta and zeta-PKC using isotype specific pseudosubstrates viz., [Ser25]PKC[19-31] and [Ser119]PKC[113-130] respectively. Endogenous protein phosphorylation by purified beta-PKC and zeta-PKC showed their functional differences in neutrophil. Beta-PKC phosphorylated 13, 15, 19, 33, 36, 47, 80 and 92 kDa proteins and zeta-PKC phosphorylated 19, 22, 42, 47, 75 and 87 kDa proteins, only exception was the phosphorylation of 47 kDa protein which had been phosphorylated by both the kinases. Differences in phosphorylation between beta-PKC and zeta-PKC clearly indicate the selective role for these PKC isotypes in the activation sequences of neutrophil. PMID- 10724344 TI - Dietary n-3 fatty acids, curcumin and capsaicin lower the release of lysosomal enzymes and eicosanoids in rat peritoneal macrophages. AB - Male Wistar rats (12 rats/group) were fed a diet containing 8 wt % coconut oil or groundnut oil or cod-liver oil for a total period of 8 weeks. The diets were also supplemented with 2 wt % groundnut oil for providing essential fatty acids. During the last 2 weeks, 6 rats form each group were additionally given curcumin (30 mg/kg body wt/day) or capsaicin (5 mg/kg body wt/day) in 1 ml groundnut oil. The peritoneal macrophages from rats fed cod-liver oil diet secreted lower levels of lysosomal enzymes collagenase, elastase and hyaluronidase as compared to those from rats fed coconut oil or groundnut oil diets. Curcumin and capsaicin significantly lowered the secretion of these lysosomal enzymes from macrophages in animals given coconut oil or groundnut oil diet. Macrophages from rats fed cod liver oil secreted lower amounts of prostaglandin E2, 6-keto PGF1alpha, leukotrienes B4 and C4 and also incorporated lesser amounts of [3H]-arachidonic acid as compared to those given coconut oil or groundnut oil diets. Curcumin and capsaicin lowered the secretion of these eicosanoids and decreased the incorporation of [3H]-arachidonic acid in macrophage lipids. However curcumin and capsaicin significantly increased the secretion of 6-keto PGF1alpha in all the groups of animals. These studies indicated that dietary cod-liver oil (rich in n 3 fatty acids), and spice principles curcumin and capsaicin can lower the secretory functions of macrophages in a beneficial manner. PMID- 10724345 TI - Characterization of a 5'-flanking region supporting the transcription of mouse thymosin beta-4 in mouse NIH3T3 cells. AB - Expression of the gene coding for thymosin beta-4 (Tbeta-4), the major G-actin sequestering peptide in the cell, is regulated mainly at the level of transcription. In this study, we examined the nucleotide sequence of the 5' flanking region (from -2202 to -881) of the mouse Tbeta-4 gene, and demonstrated that the DNA fragment from -278 to +410 of this gene was capable of directing the expression of a chloramphenicol acetyltransferase reporter gene in NIH3T3 cells. However, expression of the reporter gene in cells cannot be induced by interferon alpha treatment even though a rapid activation of endogenous Tbeta-4 gene by this cytokine was observed. These results suggest that the projected interferon stimulated response element (ISRE) might reside in other parts of the mouse Tbeta 4 gene. PMID- 10724346 TI - Cytosolic Ca2+ concentration during Ca2+ depletion of isolated rat hearts. AB - Reperfusion of isolated mammalian hearts with a Ca2+-containing solution after a short Ca2+-free period at 37 degrees C results in massive influx of Ca2+ into the cells and irreversible cell damage: the Ca2+ paradox. Information about the free intracellular, cytosolic [Ca2+] ([Ca2+]i) during Ca2+ depletion is essential to assess the possibility of Ca2+ influx through reversed Na+/Ca2+ exchange upon Ca2+ repletion. Furthermore, the increase in end-diastolic pressure often seen during Ca2+-free perfusion of intact hearts may be similar to that seen during ischemia and caused by liberation of Ca2+ from intracellular stores. Therefore, in this study, we measured [Ca2+]i during Ca2+-free perfusion of isolated rat hearts. To this end, the fluorescent indicator Indo-1 was loaded into isolated Langendorff-perfused hearts and Ca2+-transients were recorded. Ca2+-transients disappeared within 1 min of Ca2+ depletion. Systolic [Ca2+]i during control perfusion was 268 +/- 54 nM. Diastolic [Ca2+]i during control perfusion was 114 +/- 34 nM and decreased to 53 +/- 19 nM after 10 min of Ca2+ depletion. Left ventricular end-diastolic pressure (LVEDP) significantly increased from 13 +/- 4 mmHg during control perfusion after Indo-1 AM loading to 31 +/- 5 mmHg after 10 min Ca2+ depletion. Left ventricular developed pressure did not recover during Ca2+ repletion, indicating a full Ca2+ paradox. These results show that LVEDP increased during Ca2+ depletion despite a decrease in [Ca2+]i, and is therefore not comparable to the contracture seen during ischemia. Furthermore, calculation of the driving force for the Na+/Ca2+ exchanger showed that reversed Na+/Ca2+ exchange during Ca2+ repletion is not able to increase [Ca2+]i to cytotoxic levels. PMID- 10724347 TI - Serine base exchange enzyme in porcine lyophilised platelets: enzyme properties and modulation by AlF4- and different types of heparin. AB - Phosphatidylserine is one of the PKC modulators and thus it may play an important role in signal transduction. Regulation of the synthesis of this phospholipid is not yet clarified. The contrasting reports are possibly related to the existence of different enzymes which, in mammalian tissues, catalyse the exchange between free serine and the nitrogen base of a membrane phospholipid. This study demonstrates that serine base exchange reactions of commercially available lyophilised porcine platelets exhibit similar pH optima, temperature and Ca2+ dependence as observed in fresh tissues. Analysis of fatty acids composition of the three phospholipid classes involved in base exchange reactions also demonstrated a similarity with fresh platelets. Serine and ethanolamine base exchange enzyme activities were assayed in parallel in platelet lysate subjected to preincubation at various temperatures (30-60 degrees C). When dithioerithrol was omitted from the incubation medium, the two base exchange reactions were inhibited with a similar temperature-dependent pattern. Addition of the reducing agent enhanced the sensitivity to preincubation only for the serine base exchange reaction which was inhibited by 80% after preincubation at 45 degrees C. With respect to its regulation, porcine platelet serine base exchange enzyme(s) was inhibited by fluoroalluminate, a widely used G-protein activator, and stimulated by unfractionated heparin. Low mol. wt. heparin did not influence enzyme activity. Unfractionated heparin greatly stimulated SBEE activity assayed at pH 7.4, a pH value far from the optimal pH. PMID- 10724348 TI - Vanadium and diabetes. What about vanadium toxicity? PMID- 10724349 TI - Immunologically-related or incidental coexistence of diabetes mellitus and Graves' disease; discrimination by anti-GAD antibody measurement. AB - Coexistence of diabetes mellitus and Graves' disease may be classified into either an immunologically-related or an incidental phenomenon. It has been reported that anti-GAD antibody (GAD-Ab) persists at high levels for longer duration in subjects with type 1 diabetes and Graves' disease, whereas the prevalence of positive GAD-Ab (1.5%) in 131 non-diabetic subjects with Graves' disease was comparable to that in normal subjects (0.3%, P=0.2012). Thus, GAD-Ab might be a marker of the immunologically-related coexistence of the two diseases. To test this hypothesis, we investigated characteristics of Japanese subjects having both diseases according to the presence or absence of GAD-Ab. Sixty-one patients having diabetes mellitus and Graves' disease (24 men, 37 women, aged 53+/-2 years old, mean +/- SE) were consecutively registered between 1993-1997. The patients were divided into two groups of 14 GAD-Ab positive and 47 negative subjects. In the GAD-Ab positive subjects, earlier (32+/-3 years old) and abrupt onset (86%) of diabetes and insulin dependency (64%) were documented, as would be expected from the features of type 1 diabetes. Graves' disease often preceded diabetes (57%), presenting typical manifestations (79%). In contrast, older (45+/ 2 years old, P=0.0031) and gradual onset (87%, P<0.0001) of diabetes, non-insulin dependency (74%, P<0.0001), and masked manifestations of Graves' disease (57%, P=0.0214) were common in the negative subjects. Precedence of diabetes dominated in these subjects (43%, P=0.0109). Immunological studies showed less frequent HLA DR 2 locus (0%, P<0.02) in the GAD-Ab positive subjects. There was also a trend of higher frequency of HLA-DQA1*03 allele and of lower frequency of DQA1*01 allele in these subjects. Allelic frequency of cytotoxic T lymphocyte antigen 4 (CTLA-4) differed between the positive and negative subjects (P=0.0432). There were distinct clinical and immunological differences between the GAD-Ab positive and negative subjects having both diabetes mellitus and Graves' disease. The present results indicate that GAD-Ab measurement could draw a distinction between the immunologically-related and incidental coexistence of the two diseases. PMID- 10724350 TI - Immunohistochemical localization of betacellulin, a new member of the EGF family, in normal human pancreas and islet tumor cells. AB - Betacellulin (BTC) purified from mouse beta cell tumor (betaTC-3) is a new member of the epidermal growth factor (EGF) family which can bind receptor tyrosine kinase, EGF receptor (erbB1) and erbB4. It has been demonstrated that proBTC mRNA was abundantly expressed in human pancreas tissue, and that BTC converted amylase secreting rat acinar cell line (AR42J) into insulin-secreting cells, suggesting that BTC might be important for the growth and/or differentiation of islet cells. However, the cell type producing BTC in the pancreas has not been clarified. In this study, we examined the localization of BTC in human pancreas and islet cell tumors. Immunohistochemistry using specific antibodies to human BTC revealed that this protein was produced in alpha cells and duct cells, and probably in beta cells in normal adult pancreas. Furthermore, strong immunoreactivity to BTC was detected in primitive duct cells of the fetal pancreas, and both insulinoma and glucagonoma cells also showed positive immunoreactivity to BTC. EGF receptor (erbB1) and erbB4 were expressed mainly in islet and duct cells, and duct cells, respectively. These results demonstrate the localization of BTC and its receptors, and suggest that BTC may be one of the factors that have physiologically important roles such as growth and differentiation of islet cells in the human pancreas. PMID- 10724351 TI - Expression of Ah receptor and dioxin-related genes in human uterine endometrium in women with or without endometriosis. AB - 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) has been suggested as a possible etiologic factor for endometriosis, a condition in which endometrium-like tissues are present outside the uterus. The prevailing view pertaining to the origin of endometriotic cells is that they are from eutopic endometrial cells which regurgitate through fallopian tubes. In order to get insight into the possible involvement of TCDD in the pathogenesis of endometriosis, we suspected that TCDD may act differently on the endometrium with or without endometriosis. To address this, we examined the presence of messenger RNAs of arylhydrocarbon receptor (AhR), AhR nuclear translocator (Arnt) and two dioxin-responsive genes, cytochrome P-450 1B1 (CYP1B1) and downstream of tyrosine kinases (p62(dok)), in the endometrium of women with or without endometriosis using semi-quantitative reverse transcription-polymerase chain reaction. All the genes were expressed throughout the menstrual cycle. The expression level of p62(dok) was higher in the proliferative phase than in the secretory phase. In contrast, the expression levels of AhR, Arnt and CYP1B1 seemed to be constant during the cycle. In terms of the comparison between non-endometriosis and endometriosis group, the mRNA levels of AhR, Arnt, CYP1B1 and p62(dok) were essentially similar. Interestingly, AhR mRNA level was significantly lower in smokers than in non-smokers. Based on the regression analysis, significant linear and positive correlations were observed between AhR and Arnt mRNA levels, and between Arnt and p62(dok) mRNA levels. In summary, expression of AhR and dioxin-related genes in the endometrium did not differ in women with or without endometriosis. PMID- 10724353 TI - Endogenous nitric oxide inhibits growth hormone secretion through cyclic guanosine monophosphate-dependent mechanisms in GH3 cells. AB - Constitutive nitric oxide synthase (NOS) is expressed in rat adenohypophysis and clonal GH3 cells. The mechanisms of action of nitric oxide (NO) to inhibit hormone secretion and the possible role of (6R)-5, 6, 7, 8-tetrahydro-L-biopterin (THB) in the action of endogenous NO were studied in GH3 cells. Inhibiting NOS with N(G)-nitro-L-arginine or trapping NO with oxyhemoglobin enhanced both the basal and TRH-stimulated rat GH release. Sodium nitroprusside did not further decrease either the basal or the TRH-stimulated GH secretion, suggesting that endogenous NO exerted the maximal inhibitory effect. Inhibition of de novo synthesis of THB increased GH secretion. A cyclic guanosine-monophosphate (cGMP) antagonist did not increase the basal GH secretion but enhanced TRH-induced GH release. These findings suggest that endogenous NO plays an inhibitory role on basal GH release and TRH-stimulated hormone release from GH3 cells in an autocrine or paracrine fashion, at least partly, through a cGMP-dependent pathway. It is also suggested that endogenous THB plays a role in NO production and subsequent inhibition of hormone secretion in GH3 cells. PMID- 10724352 TI - Differential interactions of bisphenol A and 17beta-estradiol with estrogen receptor alpha (ERalpha) and ERbeta. AB - Bisphenol A (BPA), a monomer of plastic used in consumer products, is abundant in the environment and enters the body by ingestion or adsorption. We developed a cell based transcription assay system using a reporter gene under the transcriptional control of estrogen receptor alpha (ERalpha) as well as ERbeta and performed chloramphenicol acetyltransferase (CAT) assay on HeLa cells transfected with either human ERalpha cDNA or ERbeta cDNA to characterize the estrogenic effect of BPA. Estrogenic activity of BPA was detectable at a concentration of 10(-9) M and the activity increased in a dose dependent manner between concentrations of 10(-9) M and 10(-6) M of BPA for both ERalpha and ERbeta. The estrogenic activity of 17beta-estradiol at a concentration of 10(-8) M was almost compatible with that of BPA at the concentration of 10(-6) M of BPA for ERalpha as well as ERbeta. CAT activity was significantly decreased when cells expressing ERalpha were incubated with 10(-6) M of BPA and 10(-8) M of 17beta-estradiol while the activity was essentially the same for ERbeta in the same condition, indicating that BPA exhibits only agonistic action for ERbeta whereas it has dual actions as an agonist and antagonist of estrogen for ERalpha. These results indicates that BPA exerts its effects in ER subtype specific way, thus suggesting that the mode of action of endocrine disruptors are more complex than thought. PMID- 10724354 TI - Gestational thyrotoxicosis with acute Wernicke encephalopathy: a case report. AB - A 35-year-old hyperthyroid woman who developed nausea, vomiting, tachycardia, nystagmus and mental disturbance, was referred to our hospital with a suspected diagnosis of thyroid storm. However, the thyroid gland was only slightly palpable, bruits were not audible, and exophthalmos was not present. Serum levels of thyroid hormone were increased, but TSH receptor antibodies were negative. Echography and color flow doppler ultrasonography revealed a slightly enlarged thyroid gland and a slightly increased blood flow, both of which were much less milder than those expected for severe hyperthyroid Graves' disease. Under the diagnosis of hyperthyroidism due to gestational thyrotoxicosis associated with Wernicke encephalopathy, vitamin B1 was administered on the first day of admission. Her consciousness became nearly normal on the second day except for slight amnesia. Her right abducent nerve palsy rapidly improved, but horizontal and vertical nystagmus, diminished deep tendon reflexes and gait ataxia improved only gradually. MRI findings of the brain were compatible with acute Wernicke encephalopathy. We concluded that history taking and physical findings are important to make a differential diagnosis of gestational thyrotoxicosis with acute Wernicke encephalopathy from Graves' thyroid storm, and that Wernicke encephalopathy should be treated as soon as possible to improve the prognosis. PMID- 10724355 TI - Short-term treatment with troglitazone decreases bone turnover in patients with type 2 diabetes mellitus. AB - Poorly controlled type 2 or non-insulin dependent diabetes mellitus (NIDDM) patients exhibit high bone turnover, which decelerate with treatment according to the degree of improvement in glycemic control. In adults, higher bone turnover results in rapid bone loss. Therefore, deceleration of bone turnover is beneficial for bone. Troglitazone (Tro), a new anti-diabetic drug, is a thiazolidinedione (TZD) which promotes adipocyte differentiation by activating peroxisome proliferator activated receptor gamma (PPARgamma). Because, in the bone marrow, adipocytes and osteoblasts originate in common mesenchymal stem cells that are also essential for osteoclastogenesis, TZDs may directly affect bone metabolism. Thus, we examined the effects of Tro on metabolic bone markers in type 2 DM patients. Tro (400 mg/day) was administered to 33 type 2 DM patients for four weeks. The day before and four weeks after starting Tro, serum and urine samples were collected after overnight fasting. Metabolic bone markers and glycemic indices were assessed. As bone resorption markers, urinary free and total deoxypyridinoline as well as urinary collagen type I C-terminal telopeptide were measured; as bone formation markers, serum bone type and total alkaline phosphatase (BALP and ALP) levels along with osteocalcin (OC) were used. No significant changes in fasting plasma glucose or HbA1c levels were observed in our short-term treatment with Tro. All the bone resorption markers, BALP and ALP were significantly decreased. OC was not significantly changed. The discrepant changes of OC from all the other metabolic bone markers suggest limitation of the use of OC as a reliable bone formation marker in diabetics. Our results that Tro decreased metabolic bone markers before significantly improving glucose metabolism suggest that it has direct effects on bone and decreased bone turnover. TZDs may spare bone mass in NIDDM subjects through its dual effects on glucose and bone metabolism. PMID- 10724356 TI - Diabetic nephropathy accompanied by iodine-induced non-autoimmune primary hypothyroidism: two case reports. AB - We reported 2 diabetic patients with nephrotic syndrome due to advanced diabetic nephropathy complicated by non-autoimmune primary hypothyroidism. Hypothyroidism developed along with the anasarcous status. Histological examinations of the thyroid gland revealed almost normal thyroid follicles without lymphocytic infiltration. The amounts of thyroid hormone lost into the extravascular space such as in urine and ascites were not sufficient to cause hypothyroidism alone. Serum total iodine levels measured during the hypothyroidal state in both cases were definitely elevated, and the perchlorate discharge test of both cases showed positive discharge (24 and 34%, respectively). The thyroid functions normalized after iodine restriction in the first case and initiating hemodialysis in the second case, in parallel with normalization of serum total iodine levels. These findings suggest that impaired renal handling of iodine resulting in elevation of serum iodine levels, rather than an autoimmune mechanism or extravascular hormone loss, played a principal role in the development of primary hypothyroidism found in these 2 patients, probably through a prolonged Wolff-Chaikoff effect. PMID- 10724357 TI - No evidence of germline mutation or somatic deletion of the MEN1 gene in a case of familial multiple endocrine neoplasia type 1 (MEN1). AB - The MEN1 gene has recently been cloned as the gene responsible for multiple endocrine neoplasia type 1 (MEN1) and its germline mutations have been identified in a number of familial MEN1 patients. However, mutation-negative cases have also been reported in some MEN1 families. We report here a Japanese MEN1 family, including a proband with no evidence of MEN1 gene mutation. The proband (51 y.o., female) had three major MEN1 lesions, including primary hyperparathyroidism (HP), prolactinoma, and pancreatic tumor. Her father and brother had HP, and her daughter had both HP and prolactinoma. When we analyzed the proband for a germline mutation of the MEN1 gene, the direct sequencing analysis showed no mutation in the coding region, on the promoter, 5' and 3' untranslated regions of the MEN1 gene. We next examined the loss of heterozygosity (LOH) in the proband's parathyroid tumors using two benign polymorphisms (C2249G in intron 1 and 2248del3 in exon 10) in the MEN1 gene to detect LOH. LOH was not found in any of the four separate regions of the tumor tissues. PMID- 10724358 TI - 21-Hydroxylase deficiency presenting as massive bilateral adrenal masses in the seventh decade of life. AB - A 72-year-old woman was found to have massive bilateral adrenal masses on computed tomography and was diagnosed with 21-hydroxylase deficiency (21-OHD) based on endocrinological findings. Physical examination revealed no abnormalities except markedly short stature. She was diagnosed with 21-OHD because she had an elevated serum 17alpha-hydroxyprogesterone (17-OHP) level which significantly decreased in response to dexamethasone. Percutaneous CT guided biopsy and later autopsy confirmed that the adrenal masses were due to adrenocortical hyperplasia. Analysis of the CYP21 gene revealed that the patient was a compound heterozygote for the Ile-172-->Asn mutation in exon 4 and the 8-bp deletion in exon 3. Simple virilizing 21-OHD (SV) would be predicted from this genotype. She had few symptoms associated with 21-OHD except for markedly short stature, but the serum 17-OHP level was higher than that of typical nonclassical form of 21-OHD and near to that of typical SV. This finding was confirmed by analysis of the CYP21 gene. From these results, we report that when adrenal masses are incidentally detected, 21-OHD should be ruled out to avoid excessive examination and surgery on the suspicion of adrenal carcinoma. PMID- 10724359 TI - A de novo L330S point mutation in thyroid hormone receptor beta gene in a Thai female with resistance to thyroid hormone. AB - In the present study, we report a Thai female with a de novo mutation in thyroid hormone receptor-beta (TRbeta) gene causing resistance to thyroid hormone (RTH). The patient was a 19 year-old woman who presented with goiter for 1 year. Except for tachycardia she had no signs of thyrotoxicosis. Previously she was treated with propylthiouracil based on the diagnosis of thyrotoxicosis for 9 months and her goiter became more enlarged. The patient was the only child of the family. Her parents were alive and healthy, and did not have goiter or any other thyroid diseases. Physical examination revealed no sign of thyrotoxicosis. Her thyroid gland was diffusely enlarged with an estimated weight of 100 gm. Laboratory determinations revealed elevated free T4, T3 and nonsuppressed TSH levels. Exon 9 of the TRbeta gene was amplified by PCR and the DNA sequence was determined by dye terminator cycle sequencing. Heterozygous point mutation in which T was replaced by C was detected at position 1274 (TTG to TCG) corresponding to a leucine to serine substitution at codon 330. No mutation was found in the parents indicating that the mutation was de novo. The nucleotide change created a restriction site for Taq 1 restriction endonuclease and the mutation was confirmed by restriction fragments length polymorphism. The same nucleotide change has been reported in a family with RTH. PMID- 10724360 TI - Intraventricular injection of melatonin inhibits naloxone-induced, but not NMDA- or LHRH-induced LH release in ovariectomized estrogen-primed rats. AB - The present study was aimed to examine the possible functional relationship between melatonin and hypothalamic transmitters, endogenous opioids and excitatory amino acids in controlling gonadotropin secretion in ovariectomized estrogen-primed rats. An intravenous injection of naloxone (mu opioid receptor antagonist), N-methyl-D-aspartate (NMDA; NMDA receptor agonist) or luteinizing hormone-releasing hormone (LHRH) significantly elevated serum luteinizing hormone (LH) concentrations within 10 min. An intraventricular treatment with melatonin, which did not affect the basal LH concentration by itself, significantly suppressed the effect of naloxone. However, the same melatonin treatment did not inhibit the NMDA-induced or LHRH-induced LH secretion. These results support the hypothesis that melatonin has a suprapituitary site of action to inhibit LHRH release, and suggest that the site of its action may be located downstream to that of naloxone action and upstream to that of NMDA in the hypothalamic LHRH neuronal pathway. PMID- 10724361 TI - A case of anti-islet-antibody-positive patient treated with insulin plus calcitriol. PMID- 10724362 TI - DNA fragmentation pattern induced in thymocytes by sulphur mustard. AB - Sulphur mustard (HD) is a blister agent for which no specific therapy exists. The mechanism of cell injury caused by HD is not well understood. This study examined DNA damage in thymocytes exposed to a range of HD concentrations over a time course of 1-24 h. Thymocytes incubated with HD showed an increase in the production of DNA fragments of the type frequently associated with apoptosis, namely, initial formation of large fragments of 30-50, 200-300 and > 700 kilobase pairs (kbp), followed by further degradation to produce an internucleosomal 'ladder' of oligomers of approximately 180 base pairs (bp). Pulsed field electrophoresis analysis of thymocytes incubated with HD detected breakdown of the chromatin up to 3 h before a corresponding increase in the low molecular weight (MW) oligonucleosomal fragments could be seen on conventional agarose gels. These results suggest that cells damaged by HD poisoning may be irretrievably committed to cell death sooner after exposure than previous studies suggested. The nature of the DNA fragments produced suggested that apoptosis may represent a component of the pathway of cell death induced by HD. These aspects may have implications for the search for specific therapeutic reagents effective in the prevention or treatment of HD poisoning. PMID- 10724363 TI - Biotransformation of the double bond in allyl glycidyl ether to an epoxide ring. Evidence from hemoglobin adducts in mice. AB - Allyl glycidyl ether (AGE) is used industrially in the production of various epoxy resins. The compound is mutagenic and evidence for carcinogenicity in mice and rats has been reported. A previous study in mice showed that AGE reacts directly, without metabolic activation, with N-terminal valine in hemoglobin to form adducts (AGEVal). Metabolism of AGE may lead to formation of diglycidyl ether (I) through epoxidation of the double bond or 1-allyloxy-2,3 dihydroxypropane (II) through hydrolysis of the epoxide ring. 2,3-Dihydroxypropyl glycidyl ether (III) may be formed either by hydrolysis of I or epoxidation of II. The main aim of the present study was to investigate if AGE is metabolized to the reactive epoxides I or III by analysis of adducts with hemoglobin. Nine male mice (C3H/Hej) were administered AGE dissolved in tricaprylin, 4 mg/mouse, by intraperitoneal (i.p.) injection. Eleven male mice were administered 4 mg/mouse of AGE dissolved in acetone, by skin application. Adducts of I or III with N terminal valine, N-(2-hydroxy-3-(2,3-dihydroxy)propyloxy)propylvaline (diOHPrGEVal), were demonstrated in mice administered AGE by i.p. injection. The levels were in the range 1600-5600 pmol/g globin. The level of diOHPrGEVal in mice administered AGE by skin application (n = 5) was below the detection limit of the analytical method, 20 pmol/g globin. The level of AGEVal, analyzed in mice administered AGE by skin application (n = 6), was about 20 pmol/g globin (median value), as compared with 1600 pmol/g globin previously found in mice administered AGE by i.p. injection. Neither AGEVal nor diOHPrGEVal were detected in control animals. Both adducts were analyzed using a modified Edman method for derivatization and using gas chromatography/tandem mass spectrometry for detection. The hydroxyl groups of the Edman derivative of diOHPrGEVal were protected by acetylation. PMID- 10724364 TI - Cytotoxicity and DNA binding properties of a chloro glycylhistidinate gold(III) complex (GHAu). AB - The chloro glycylhistidinate gold(III) complex (GHAu) is shown to be fairly cytotoxic towards the established A2780 ovarian carcinoma human cell line either sensitive or resistant to cisplatin. Remarkably, GHAu is far more cytotoxic than the corresponding zinc(II), palladium(II), platinum(II) and cobalt(II) complexes implying that cytotoxicity is essentially to be ascribed to the presence of a gold(III) center. Circular dichroism (CD) spectra, atomic absorption measurements and DNA melting profiles suggest that GHAu in vitro is able to bind DNA, the presumed target for several antitumor metal complexes, and to modify its conformation, even if the observed changes are generally small. Implications of these findings for the mechanism of action of cytotoxic gold(III) complexes are discussed. PMID- 10724365 TI - Difference of the liver and kidney in glycine conjugation of ortho-substituted benzoic acids. AB - The relative importance of the liver and kidney for glycine conjugation of ortho substituted benzoic acids was investigated. Glycine conjugation of ortho substituted benzoic acids was investigated in mouse liver and kidney mitochondria. The extent of glycine conjugation of benzoic acids with the halogen group decreased in the order F > Cl > Br > I. The conjugation of salicylic acid with glycine took place in only the kidney. 2-Methoxybenzoic acid exhibited no activity in the liver and kidney. The difference in glycine conjugation of ortho substituted benzoic acids was observed between liver and kidney. The kidney was more active in glycine conjugation of ortho-substituted acids than the liver. In addition, the relationship between glycine conjugation and the chemical structure of ortho-substituted acids was examined in the liver and kidney. The size of the substituent had a far greater influence over glycine conjugation in the liver and kidney. Glycine conjugation was also dependent on the substituent electronegativity. It may be important that the substrates undergoing glycine conjugation contain a flat region coplanar to the carboxylate group. PMID- 10724367 TI - Equivalence in radionuclide metrology AB - The harmonization of different National Measurement Systems depends on the ability to demonstrate Equivalence between them. The responsibility for establishing a procedure to achieve this lies with the International Committee of Weights and Measures (CIPM) and its associated Consultative Committees. In pursuit of these ends, a series of general recommendations and procedures have been developed by BIPM in order to achieve these objectives. Central to these recommendations is the concept of Key Comparisons which involve a series of comparison exercises both within and between the regional metrology organizations, such as EUROMET, NORAMET and APMP. Because of the unique nature of radioactivity standardization, this generalized procedure is, for practical purposes, impossible to apply in the field of radionuclide metrology. The remedy has been the development of a protocol which optimizes both the current effort available within existing National Metrology Institutes (NMIs) and the considerable amount of data from previous intercomparisons. This paper details this novel approach, provides practical examples, discusses potential problems and examines the way in which the programme will continue and expand. PMID- 10724366 TI - Cultures with cryopreserved hepatocytes: applicability for studies of enzyme induction. AB - The use of hepatocyte cultures is well established for the study of drug-drug interactions. However, the major hindrance for the use of human hepatocyte cultures is that human hepatocytes are only occasionally available. This problem could be overcome by cryopreservation. Although cryopreserved hepatocytes have been recommended for short term applications in suspension, studies on induction of enzyme activity, requiring a more prolonged maintenance of cryopreserved hepatocytes in culture, represent a new field of research. In the present study, we established a technique that allows preparation of rat hepatocyte co-cultures, using cryopreserved hepatocytes. After incubation with phenobarbital (0.75 mM; 72 h) induction factors for the isoenzyme-dependent regio and stereoselective testosterone hydroxylations were 1.6, 2.2, 1.0, 2.1, 5.6, 2.4, 3.6, 4.5 and 0.9 for 2alpha-, 2beta-, 6alpha-, 6beta-, 7alpha-, 15beta-, 16alpha- and 16beta hydroxytestosterone and 4-androsten-3,17 dione. Regarding induction factors of less than 2-fold, as questionable these induction factors were similar to those of cultures with freshly isolated hepatocytes and the induction pattern of the individual hydroxylation products was similar to the in vivo situation. In addition 3-methylcholanthrene (5 microM; 72 h) induced exclusively the formation of 7alpha-hydroxytestosterone (6.6-fold) in cultures with cryopreserved hepatocytes. This specificity also correlates to that obtained in rats. Although these induction factors were clearly satisfactory in cryopreserved cultures, the absolute activities of the main testosterone hydroxylation products were reduced when compared to fresh cultures. For instance, 6beta-hydroxytestosterone, the main metabolite in solvent controls was reduced to 79%, 7alpha hydroxytestosterone, the main metabolite after induction with 3-MC, was reduced to 66% and 16beta-hydroxytestosterone, the main metabolite after induction with PB, was reduced to 52%. Similarly, EROD activity after induction with 3 methylcholanthrene in cryopreserved cultures was reduced to 62%, compared with that in fresh cultures. Although further optimization and validation is required, the data show that cytochrome P450 activities can clearly be induced in co cultures of cryopreserved hepatocytes, in a fashion which for the investigated inducers, is similar to that in cultures from freshly isolated hepatocytes and similar to the in vivo situation. PMID- 10724368 TI - Influence of radioactive impurities on SIR measurements AB - The correction usually applied to an SIR measurement in the presence of a gamma ray emitting impurity is presented and explained. The method of calibration of a Ge(Li) spectrometer is briefly described. It is used to look for impurities when SIR measurements show inconsistencies. An example is presented where a 103Ru impurity was discovered in a 99Mo solution. The correction applied compensates for the decrease of around 0.2% per day observed in the SIR measurements of this 99Mo ampoule. PMID- 10724369 TI - Calibration and consistency of results of an ionization-chamber secondary standard measuring system for activity AB - Calibration in terms of activity of the ionization-chamber secondary standard measuring systems at the PTB is described. The measurement results of a Centronic IG12/A20, a Vinten ISOCAL IV and a radionuclide calibrator chamber for nuclear medicine applications are discussed, their energy-dependent efficiency curves established and the consistency checked using recently evaluated radionuclide decay data. Criteria for evaluating and transferring calibration factors (or efficiencies) are given. PMID- 10724370 TI - The Spanish National Reference Database for Ionizing Radiations (BANDRRI) AB - The Spanish National Reference Database for Ionizing Radiations (BANDRRI) is being implemented by a reasearch team in the frame of a joint project between CIEMAT (Unidad de Metrologia de Radiaciones Ionizantes and Direccion de Informatica) and the Universidad Nacional de Educacion a Distancia (UNED, Departamento de Mecanica y Departamento de Fisica de Materiales). This paper presents the main objectives of BANDRRI, its dynamic and relational data base structure, interactive Web accessibility and its main radionuclide-related contents at this moment. PMID- 10724371 TI - Computer simulation of backscattered alpha particles AB - Alpha-particle spectrometry forms an important aspect of radionuclide metrology. Accurate measurements require corrections to be made for factors such as self absorption within the source and backscattering from the backing material. The theory of the latter phenomenon has only received limited attention. Furthermore the experimental verification of these theoretical results requires adequate counting statistics for a variety of sources with different activities. These problems could be resolved by computer simulations of the various interactions which occur as alpha-particles move through different materials. The pioneering work of Ziegler and his coworkers over several years, has provided the sophisticated software (SRIM) which has enabled us to obtain the results presented here. These results are compared with theoretical and experimental values obtained previously. PMID- 10724372 TI - Experimental validation of an analytical method to obtain the response function of an alpha particle spectrometer AB - In a previous paper, one of the authors suggested an analytical method for calculation of the response function of an alpha spectrometer for the case of large solid angles. This paper describes the experimental verification of the method. Spectra of a well-known natural uranium sample were measured with a 450 mm2 Si detector and compared to the theoretical predictions. The measurements were carried out with two different geometrical configurations. In both cases a good agreement was observed between experimental and theoretical results. PMID- 10724373 TI - Improvements in quantitative source preparation AB - Quantitative source preparation is indispensable for radionuclide standardisation. To improve the source quality, a device has been developed to accelerate the evaporation of solvents from a drop deposited on a substrate. Short drying times were reached by stirring the rotating drop with multiple jets of dry nitrogen at elevated temperature. Uniform deposits with a large number of small crystals were obtained. The source-quality was checked by micrographs and scans of autoradiographs and by the shape parameters of alpha-particle spectra. PMID- 10724374 TI - Application of rotating disk electrode technique for the preparation of Np, Pu and Am alpha-sources AB - Method of electrodeposition on rotating disk cathode (RDE) is applied for preparation of Np, Pu and Am alpha-standards. Phenomenon of critical current density is experimentally observed which is in perfect accord with Hansen's theory of electrodeposition. Influence of deposit calcination regime on quality of alpha-sources is studied, and comparison is made of uniformity of deposits obtained in various deposition systems. Standards with energy resolution better than 9 keV can be reproduceably obtained by optimized RDE electrodeposition technique. PMID- 10724375 TI - Properties of a YAP powder scintillator as alpha-ray detector AB - A YAP scintillator (YAlO3: Ce crystal) for alpha counting has been produced in powder form and this paper describes its performance characteristics. By measuring pulse-height and rise-time distributions, it was found that the YAP powder, as a fast detector for alpha rays, could clearly distinguish alpha events from beta and gamma events. In addition, the YAP powder was used in a phoswich detector combined with a YAG (Y3Al5O12: Ce) crystal for beta and gamma detection. PMID- 10724376 TI - Measurement of radon and radon progenies at the German radon reference chamber. AB - The activity concentration of radon in the environment can vary over five orders of magnitude. Radon and its progenies thus concern all people involved in radiation protection as well as in low-level experiments. In the German radon reference chamber at the PTB, radon and its progenies are measured with different systems for alpha- and gamma-spectrometry with the full set of environmental parameters, e.g. temperature, humidity and aerosol concentration being controlled. Control of air pressure is also possible by use of an extention chamber. The sampling and measuring technique for radon and its short-lived progenies at the German radon reference chamber are the basis for fundamental studies with regard to the understanding of the equilibrium factor and the unattached fraction of progenies. The facility also serves for the calibration of radon progeny detectors. PMID- 10724377 TI - Pulse loss and counting statistics with a digital spectrometer AB - A commercially available digital gamma-ray spectrometer is tested at high count rates using a germanium detector and appropriate pulse processing parameters. Correction for pulse loss due to dead time and pileup is done by extending the live time according to the Gedcke-Hale method. The accuracy of this count-loss correction technique is tested experimentally up to saturation count rates. Also the statistical uncertainty applicable to the total number of counts in arbitrary regions of interest in gamma-ray spectra is studied experimentally, for measurements at fixed real time as well as at fixed live time. The results are similar to those found with traditional spectrometers using analog technology. PMID- 10724378 TI - Standardisation of 210Pb AB - The standardisation of 210Pb is complicated by the presence of the daughters, 210Bi and 210Po. In addition, the low energies of the beta emissions from 210Pb make it difficult to obtain high detection efficiencies in an atmospheric proportional counter and hence produce the need for large extrapolations with consequential large uncertainties when extrapolating to unit efficiency with the conventional 4pi(PC)-gamma-coincidence technique. In order to produce a reliable standardisation, it is necessary to remove the daughter products. A solution of 210Pb was therefore chemically separated from its daughters and then standardised using the conventional 4pi(LS)-gamma-coincidence technique. The low energy (46 keV) and low emission probability (4%) of the associated photon emissions effectively rules out the possibility of using ionisation chambers as secondary standard transfer instruments for this nuclide. A germanium spectrometer therefore was calibrated for this purpose using 241Am as a normalising agent. The results of this work are presented together with an analysis of the standardisation uncertainties that can be achieved in practice. PMID- 10724379 TI - Improved methods of measurement and analysis of conversion electron and beta particle spectra AB - A general statistical test of the stability of measurement conditions was demonstrated on the beta-spectra of 241Pu cumulated during four years. The alpha- and gamma-ray spectroscopy indicated stability of the 241Pu source. Monte Carlo modelling of individual collision events clarified the role of electron scattering and energy losses within a radioactive source down to energies of several hundreds of eV. The impact ionization by beta-particles of carbon and oxygen atoms in a surface contamination layer on the 241Pu and 63Ni sources was observed. PMID- 10724380 TI - Development of a three-dimensional data acquisition method for standardization of beta-emitting nuclides AB - We have developed a three-dimensional data-acquisition system with which pulse heights from an array of detectors can be multi-scaled with dwell times as short as 31 ns. The main part of the system consists of three ADCs of SAR (Successive Approximation Register) type, a 32 MHz oscillation clock and DRAM (Dynamic Random Access Memory) of 192 Mbyte capacity. The clock time is determined by count the clock pulse generated by 28 bits synchronous counter. Both the pulse height and the time information are stored at an array of DRAM, of 40-bits width, 12-bits for pulse height and 28-bits for clock time. The technique is particularly suitable for experiments such as both double and triple coincidence distributions are required. Virtues of the method are demonstrated by measuring the activity of 14C with a 3-PM liquid scintillation counting system. The computer discrimination together with the MCTS (multi-channel time scaling) technique is applied for efficiency variation method in the analysis. PMID- 10724381 TI - Quantification of transuranic elements by time interval correlation spectroscopy of the detected neutrons AB - The non-destructive quantification of transuranic elements in nuclear waste management or in safeguards verifications is commonly performed by passive neutron assay techniques. To minimise the number of unknown sample-dependent parameters, Neutron Multiplicity Counting (NMC) is applied. We developed a new NMC-technique, called Time Interval Correlation Spectroscopy (TICS), which is based on the measurement of Rossi-alpha time interval distributions. Compared to other NMC-techniques, TICS offers several advantages. PMID- 10724382 TI - Cryogenic detectors below 100 mK for X-ray measurements in metrology AB - Due to the intrinsic performances of cryogenic detectors such as energy resolution, LPRI has decided to use these devices to improve the quality of the radioactive measurements usually obtained with classical semiconductor detectors. A bolometer with a 10 microg tin absorber has been developed at IAS (Institut d'Astrophysique Spatiale) and has been tested in the cryogenic installation of LPRI; an energy resolution (full width half maximum, FWHM) of 39 eV has been obtained on the Kalpha line of Mn. Besides these good spectrometry results, an absolute activity measurement using bolometers is proposed by adapting an absorber geometry for 4 pi counting. PMID- 10724383 TI - Radiochemical analysis of 41Ca and 45Ca. AB - The radioactive isotopes of calcium, 41Ca (t1/2 = 1.03 x 10(5) yr) and 45Ca (t1/2 = 163 d), are produced by neutron capture in the stable isotopes 40Ca and 44Ca, respectively. These radionuclides are present in the environment due to the reactions between the galactic cosmic rays and the earth's surface, and in nuclear power plants by the activation of the structural components of nuclear reactor vessels. The aim of this paper is to propose a radiochemical separation method of 41Ca and 45Ca from the other beta gamma emitters present in radioactive materials, based on selective precipitation reactions. The activities were measured by liquid scintillation counting (LSC). The obtained decontamination factors are satisfactory for each radioactive component of the initial sample in that their activities in the final product were lower than the minimum detectable activity (MDA) due to the effectiveness of the radiochemical procedure. The sensitivity of the method allows the radiological characterization of 41Ca and 45Ca content in radioactive materials. PMID- 10724384 TI - Standardization and half-life measurement of antimony-124 AB - The half-life of 124Sb has been measured using two solutions prepared from 124Sb obtained from the nuclear reactions: 124Sn(p,n) 124Sb (cyclotron) and 123Sb(n,gamma) 124Sb (reactor). These solutions were standardized by the 4pibeta gamma-coincidence extrapolation method with a total uncertainty of 0.12%. A 124Sb half-life of (60.11+/-0.07) days was measured. PMID- 10724385 TI - Simulation of nuclear decay AB - The present article depicts a software module to prepare the particles and quanta emitted during the disintegration of nuclides with complex decay schemes. This program is particularly useful for Monte Carlo simulations involving detectors showing pulse-summation effects. It generates the decay-scheme dependant input to trace the interactions of the emitted particles (and the secondary particles created) with a particular detector; then the simulation code produces emission and energy-deposition spectra for the chosen radionuclides. PMID- 10724386 TI - A radon 222 traceability chain from primary standard to field detectors. AB - The development of a primary standard from a method for measuring the absolute activity of 222Rn has also made it possible to establish secondary standards. Detailed procedures to obtain these secondary standards are given. These standards are, in particular, adapted to the requirements of laboratories that have developed equipment for the calibration and comparison of instruments measuring the concentration of radon and its daughters. An example of the implementation of these new resources applied to the qualification of field detectors is given. The propagation of measurement uncertainties at each level (primary, secondary, test radon chamber) is described. PMID- 10724387 TI - Correlation effect in activity measurement of 59Fe AB - Correlation effects due to the cascade transition of gamma-rays with gates on the relevant peaks have been studied in activity measurement of 59Fe using the bi dimensional coincidence counting method. Extrapolation curves were obtained at three gamma gates of 1099.2 keV, 1291.6 keV, and that covering both photo-peaks by a computer discrimination method. Of these extrapolation curves, only that obtained at (950-1350) keV gamma gate shows a linear variation, but those of two single gamma gates are not linearly varied in the high efficiency region. This nonlinearity obtained at the gamma gate of 1099.2 keV photo-peak was caused by the correlation effects due to 1-3 coincidence relation via unobserved 192 keV gamma-ray transitions. PMID- 10724388 TI - MTR2: a discriminator and dead-time module used in counting systems AB - In the field of radioactivity measurement, there is a constant need for highly specialized electronic modules such as ADCs, amplifiers, discriminators, dead time modules, etc. But sometimes it is almost impossible to find on the market the modules having the performances corresponding to our needs. The purpose of the module presented here, called MTR2 (Module de Temps-mort Reconductible), is to process, in terms of pulse height discrimination and dead-time corrections, the pulses delivered by the detectors used in counting systems. This dead-time, of the extendible type, is triggered by both the positive and negative parts of the incoming pulse and the dead-time corrections are made according to the live time method. This module, which has been developed and tested at LPRI, can be used alone in simple counting channels or in more complex systems such as coincidence systems. The philosophy governing the choice and the implementation of this type of dead-time as well as the system used for the dead-time corrections is presented. The electronic scheme and the performances are also presented. This module is available in the NIM standard. PMID- 10724389 TI - Simulation of the response of the IG11 4pigamma ionization chamber using GEANT Monte Carlo code AB - The response of the Centronic IG11 4pigamma ionization chamber filled with argon at 2 MPa pressure has been calculated by means of the GEANT Monte Carlo code and compared with experimental results in the range of photon energies from 40 keV to 2 MeV. The effect of density variations or thickness of the main components on the sensitivity have been investigated as well as source displacements. The sensitivity to electrons has been evaluated. PMID- 10724390 TI - Results from the EUROMET action 410 concerning the decay data of 169Yb AB - An international comparison EUROMET, action No. 410, was organized with the objective of improving the knowledge of nuclear data for 169Yb decay. To determine the photon emission probabilities, the participants were asked to measure at least one of the quantities, activity per unit mass and/or photon emission rate per unit mass. In addition, the participants were requested to report the count rates observed with point source samples for an eventual coaxial type germanium detector characterization. Eleven laboratories participated, giving one or more sets of results. In all, 34 sets of results were received, 17 for the activity measurement, 11 for the photon emission rate measurement and six for the detector characterization. Using the accurate activity value obtained from this exercise, it was possible to determine the emission probabilities of the main X- and gamma-rays with an uncertainty of 1-2% for the LX-rays, 1% for the KX-rays and < or =0.5% for the main gamma-rays. These data can be very useful for the calibration of so-called gamma-X ray detectors'. The measurement of 169Yb with type P or N coaxial structure detectors has also made possible an estimation of their diameters and volumes. PMID- 10724391 TI - Determination of the absolute alpha-particle emission probabilities of 237Np AB - In the framework of the EUROMET project No. 416, decay scheme data of 237Np were measured by alpha-particle spectrometry. Emission probabilities of alpha particles and gamma-rays were determined in the past at IRMM. After improvements of the detector system and the electron bending magnet the alpha-particle measurements have been repeated. As a result, the absolute emission probabilities of 20 alpha-particle transitions were obtained with a significant lower uncertainty. Among those transitions already known, five new were found. PMID- 10724392 TI - Standardization of 237Np by the CIEMAT/NIST LSC tracer method AB - The standardization of 237Np presents some difficulties: several groups of alpha, beta and gamma radiation, chemical problems with the daughter nuclide 233Pa, an incomplete radioactive equilibrium after sample preparation, high conversion of some gamma transitions. To solve the chemical problems, a sample composition involving the Ultima Gold AB scintillator and a high concentration of HCl is used. Standardization by the CIEMAT/NIST method and by pulse shape discrimination is described. The results agree within 0.1% with those obtained by two other methods. PMID- 10724393 TI - Standardisation and measurement of the decay scheme data of 237Np AB - Within the framework of a EUROMET project, a solution of 237Np/233Pa at equilibrium has been standardised by 4pi(PC)-gamma counting. An intrinsic Ge spectrometer was employed to measure relevant gamma-ray emission probabilities. Good agreement was obtained for those pertaining to the decay of 233Pa, but not for the gamma-ray emissions following the decay of 237Np. The cause of such discrepancies is discussed. PMID- 10724394 TI - Emission probabilities of the main gamma-rays of 237Np in equilibrium with 233Pa AB - Due to the needs in environment monitoring and nuclear waste survey, there has been increasing interest over recent years for measuring the activity of 237Np. Thus, the EUROMET project No. 416 has been proposed by NPL, in order to improve as well as possible the knowledge of the decay scheme of this radionuclide. In order to achieve this objective, the emission probabilities of the main gamma rays following the decay of 237Np in equilibrium with its daughter 233Pa, have been precisely measured, using calibrated Ge-HP detectors and sources for which the activity has been determined from absolute measurement. The obtained results, the main instrumentation characteristics, details of the experimental procedure, are presented and discussed in this paper. PMID- 10724395 TI - The standardization of a 204Tl solution AB - Two methods were used for the standardization of the 204Tl solution received in the framework of the 1997 BIPM comparison. The first method, 4pi liquid scintillation (LS), consisted of the extrapolation of the 4pi counting rate to the zero discrimination level. A home-made, single channel instrument and a commercial Packard INSTA-GEL scintillator were used. The radioactivity concentration on the reference date 1 April 1997 was 63.1+/-1.3 kBq g(-1). The second method was the efficiency tracer technique, using a 60Co standard solution; the radioactivity concentration was calculated as the mean of several extrapolated values of the curve NbetaTl-204 = N0Tl-204f(1 - epsilon(betaCo-60)). The preliminary reported value was 62.8+/-0.8 kBq g(-1); the revised value is 61.8+/-1.2 kBq g(-1), in good agreement with other 4pi (p.c.) tracer results in the comparison. Some remarks are made regarding our measurements and the comparison results. PMID- 10724396 TI - Standardisation of 204Tl at IRMM AB - IRMM participated in a recent international comparison for the standardisation of a 204Tl solution organised by BIPM. The activity concentration of the 204Tl solution was measured at IRMM using 3 independent methods; Liquid Scintillation Counting using the CIEMAT/NIST method, 4pi-CsI counting and 4pibeta counting using a large pressurised proportional counter. This article describes the measurements in detail and discusses potential problems in the standardisation of 204Tl. PMID- 10724397 TI - Standardization of 169Yb and gamma-ray emission probability data AB - Within the framework of EUROMET project No. 410, standardization of a 169Yb solution has been carried out and the gamma-ray emission probabilities at its decay have been measured. 169Yb activity was measured by the 4pibeta-gamma coincidence extrapolation method. Special attention was paid to the measurement conditions associated with the existence of 316 keV metastable level of 169Tm (T approximately 0.66 micros). The 169Yb gamma-ray emission probabilities have been determined from the results of absolute activity measurements and data obtained using a coaxial Ge(Li) spectrometer. PMID- 10724398 TI - Standardization of 169Yb by the 4pibeta-gamma method AB - 169Yb was standardized in the framework of an EUROMET international comparison. Two solutions, with approximately activity concentrations of 16 MBq g(-1) and 1.6 MBq g(-1) were distributed to the participants by BNM-LPRI-(France). In the Radionuclide Metrology Laboratory of INFIN-HH, the weak solution was standardized using a coincidence method, applying the 'beta-efficiency extrapolation' variant. A simple 4pi beta-gamma coincidence equipment was used. Due to the presence of a metastable state with a half-life of 0.66 micros in the decay scheme of 169Tm, a resolution time of 5.1 micros was chosen for the coincidence module. A gamma threshold of 150 keV was used. A second degree extrapolation graph was used, providing an activity concentration value of 1.18 MBq g(-1) at 15 November 1997, 12 h UT. The combined uncertainty was 0.9%. Using the dilution factor provided by BNM-LPRI, the corresponding activity concentration for the strong solution is 11.989 MBq g(-1) (uA = 0.110MBq g(-1); 0.92%). PMID- 10724399 TI - Results from the international evaluation exercise for uranium enrichment measurements. European Safeguards Research and Development Association NDA-WG Members AB - An international evaluation exercise for uranium enrichment measurement was organized by the ESARDA (European Safeguards Research and Development Association) Working Group on Techniques and Standards for Non-Destructive Analysis. The aim of this exercise was to test the different methods and more particularly X- and gamma-ray spectrometry methods used in the determination of uranium enrichment. Sets of samples of different uranium enrichments prepared by IRMM (Institute for Reference Materials and Measurements) and BNFL (British Nuclear Fuels) were measured by 15 participants in the IRMM laboratories. A description of this exercise is given here and the final results with their uncertainties are compared to the reference values obtained from mass spectrometry measurements. The main conclusions from this exercise are proposed; in particular, it appears necessary to improve the nuclear data used in the spectral modeling of the XKalpha region. PMID- 10724400 TI - The absolute counting of 125I AB - The Institute of Nuclear Energy Research (INER) participated in a comparison of activity organized by the Electrotechnical Laboratory (ETL, Japan). At this occasion, 125I was measured. Seven laboratories of the Asia Pacific region carried out measurements and ETL, at the same time, took part in BIPM SIR measurements. Two measurement methods developed by INER, the sum-peak coincidence counting with two 76 x 76 mm NaI(Tl) detectors and the 4pie-X coincidence counting technique with efficiency extrapolation, were used to standardize the activity of 125I. The results from the two methods are 2.6% different. INER's results agree with the results of the regional comparison and the BIPM SIR. PMID- 10724401 TI - Calibration of the National Institute of Standards and Technology tritiated-water standards AB - Massic activity values obtained for NBS/NIST tritiated water SRMs 4927 and 4927E, using internal gas-proportional counting, are reported. The massic activity ratios of all of the available NBS/NIST tritiated water SRMs and samples of the Level 1 and Level 3 tritiated water standards produced by the National Physical Laboratory (NPL) in 1996 were measured using an extensive series of liquid scintillation intercomparisons. New massic activity values obtained for the NPL standards, using the gas counting results for SRM 4927E and the results of the liquid-scintillation intercomparison, are also reported. A new determination of the half-life of tritium was made using massic activity data for SRM 4927 over a period of 38 years. The new value is (4504+/-9) (one standard deviation) days. This value differs significantly from the previous NBS/NIST value but is in good agreement with the value recommended in the Evaluated Nuclear Structure Data File (ENSDF). PMID- 10724402 TI - On the determination of activity profiles in wide-area reference sources AB - A method is described for determining the depth distribution of the radioactive substance of alpha-emitting nuclides in a wide-area reference source. The method combines measurements and calculations. In the experimental part, the energy spectrum of the source is measured by means of an alpha spectrometer. The energy spectrum is then modelled by the Monte Carlo method, taking into account energy loss of alpha particles in the source matrix, energy-loss straggling and the parameters of the spectrometer. Additional parameters for the characterization of reference sources are derived from the integral of the depth distribution. The investigations prove that the production process of commercial sources can be controlled in such a way that more than 90% of the radioactive material is implanted in the top layer of 5 microm thickness of an aluminum foil. PMID- 10724403 TI - Low-level radioactivity measurements in an ocean shellfish matrix. AB - Reference marine biological samples are necessary to test the performance of the analytical methods employed in surveying and monitoring radioactive materials in the sea. The measurement of artificial and natural radionuclide activity concentrations in ocean shellfish material by nondestructive ultra low-level gamma-ray spectrometry in an underground laboratory is reported. The material analysed, a composite material made of Irish Sea and White Sea mussel and Japan Sea oyster, was prepared by the National Institute of Standards and Technology (NIST). PMID- 10724404 TI - Absolute measurement of 166mHo radioactivity and development of sealed sources for standardization of gamma-ray emitting nuclides AB - Holmium-166m has a long half life (1200 yr) and emits a large number of gamma rays between 80 and 1400 keV. These characteristics are very suitable for gamma ray calibration sources, therefore, the absolute activity of 166mHo was measured and several sealed sources were produced to be used as reference sources for the secondary standardization systems for gamma-ray emitting nuclides. In this project, seven metal sealed sources and ten point sources were produced and several of these sources were transferred to the secondary standard laboratories to complete the traceability scheme. PMID- 10724405 TI - Decay scheme of 67Ga AB - A review of the 67Ga decay data from various evaluations reveals uncertainty as to whether or not there is an electron-capture (EC) branch decaying directly to the ground state of 67Zn. Measurements made at NAC and elsewhere of the intensity of the conversion electrons from the 93 keV transition (extracted from the standardization of 67Ga by coincidence counting techniques), place a fairly stringent restriction on the decay scheme that suggests that the ground-state branch is nonzero. Decay parameter values are proposed that are consistent with available data. Analysis indicates an EC branching ratio to the ground state of (4.01+/-0.27)%. PMID- 10724406 TI - Analysis of internal conversion coefficients AB - An extensive database has been assembled that contains the three most widely used sets of calculated internal conversion coefficients (ICC): [Hager R.S., Seltzer E.C., 1968. Internal conversion tables. K-, L-, M-shell Conversion coefficients for Z = 30 to Z = 103, Nucl. Data Tables A4, 1-237; Band I.M., Trzhaskovskaya M.B., 1978. Tables of gamma-ray internal conversion coefficients for the K-, L- and M-shells, 10 < or = Z < or = 104, Special Report of Leningrad Nuclear Physics Institute; Rosel F., Fries H.M., Alder K., Pauli H.C., 1978. Internal conversion coefficients for all atomic shells, At. Data Nucl. Data Tables 21, 91-289] and also includes new Dirac Fock calculations [Band I.M. and Trzhaskovskaya M.B., 1993. Internal conversion coefficients for low-energy nuclear transitions, At. Data Nucl. Data Tables 55, 43-61]. This database is linked to a computer program to plot ICCs and their combinations (sums and ratios) as a function of Z and energy, as well as relative deviations of ICC or their combinations for any pair of tabulated data. Examples of these analyses are presented for the K-shell and total ICCs of the gamma-ray standards [Hansen H.H., 1985. Evaluation of K-shell and total internal conversion coefficients for some selected nuclear transitions, Eur. Appl. Res. Rept. Nucl. Sci. Tech. 11.6 (4) 777-816] and for the K-shell and total ICCs of high multipolarity transitions (total, K-, L-, M-shells of E3 and M3 and K-shell of M4). Experimental data sets are also compared with the theoretical values of these specific calculations. PMID- 10724407 TI - Measurement of 49V half-life AB - The accepted value for the 49V half-life is either 330+/-15 days, based on a few old measurements with large dispersion, or 338+/-5 days, from a more recent measurement. Methods of study have included proportional counting and Nal X-ray detection. A new direct measurement of the 49V half-life has been carried out for over 2.5 half lives using liquid scintillation counting, as an additional contribution due to the scarce measurements of this radionuclide. A half-life value of 328.6+/-3.3 days was determined on the basis of an overall uncertainty analysis of 0.99%. PMID- 10724408 TI - Precise measurements of the gamma-ray emission probabilities of 186Re and 188Re. AB - Gamma-ray emission probabilities of 186Re and 188Re are important in nuclear medicine, and have been precisely measured by 4pibeta-gamma coincidence using a live-timed two-dimensional data-acquisition system. The gamma-ray emission probabilities of the 137.2 keV for 186Re and 155.1 keV for 188Re were 0.09487+/ 0.00029 and 0.15425+/-0.00072, respectively. PMID- 10724409 TI - Standardisation and decay data of 186Re AB - A solution of 186Re was standardised using the 4pi(PC)-gamma-coincidence technique. Two gamma channels were employed to provide separate detection efficiency data arising from the two decay modes (beta and electron capture): the resulting data were analysed using a two-dimensional extrapolation. Calibration factors for a high pressure re-entrant ionisation chamber as well as for the NPL secondary standard radionuclide calibrator were also determined. An efficiency calibrated germanium spectrometer was used to measure the emission probabilities of the X- and gamma-rays. PMID- 10724410 TI - Determination of the half-life of 233Pa AB - The half-life of 233Pa has been determined by following the emission rates of eight gamma rays from the decay of 233Pa produced by neutron irradiation of 232Th. Several data evaluation procedures were used in the analysis of the half life data. The measured half-life resulting from these measurements is 27.02(3) days. An evaluation in conjunction with 4 literature values leads to 26.975(13) days as the current best estimate of the half-life of 233Pa. PMID- 10724411 TI - Alpha-particle emission probabilities in the decay of 238U AB - 238U decays by alpha-particle emission to 234Th. No direct measurements of alpha particle emission probabilities (Palpha) of this nuclide have been reported since 1961, and recommended values for Palpha have remained unchanged for years, until recent evaluations suggested new values. This work presents the results of new measurements made with Si detectors and sources of natural uranium. The results obtained for the analysis of twenty spectra are: Palpha0 = 0.7754+/-0.0050, Palpha50 = 0.2233+/-0.0050 and Palpha163 = 0.0013+/-0.0003. PMID- 10724412 TI - Calculation of emission probabilities of X-rays and Auger electrons emitted in radioactive disintegration processes AB - A complete set of data for the decay scheme of a radionuclide requires knowledge of the emission probabilities of X-rays and Auger electrons. If adopted sets of transition probabilities for the transitions of the decay scheme and atomic shell data (such as fluorescence yields and relative emission probabilities) are available, the emission probabilities of the X-rays and Auger electrons can be calculated from these data. Only data calculated in this way are consistent with the adopted input data and can be compared with measured data. Conclusions can be drawn with respect to the internal consistency of the data sets by comparing the measured and calculated data. Hence, the EMISSION program has been developed and tested to calculate X-ray and Auger electron emission probabilities along with their error propagation. PMID- 10724413 TI - Search for an optimum approach to the evaluation of data of varying consistency: half-live evaluations for 3H, 35S, 55Fe, 99Mo and 111In. AB - A common procedure is proposed for the evaluation of both consistent and discrepant data in which adjustments are made to the uncertainties in the measured data that are dependent on the chi2 values of the data set. A computer program EVINEW has been developed and applied to the evaluation of 3H, 3S, 55Fe, 99Mo and 111In half-lives to give the following values: 12.32(2) yr, 87.32(16) d, 2.732(14) yr, 2.7480(9) d and 2.8047(4) d, respectively. PMID- 10724414 TI - Needs for radioactivity standards and measurements in the life sciences. AB - For almost 100 years, radioactivity has been one of the major tools in medicine. Therapeutic applications that began with 226Ra and 222Rn implants have rapidly grown to include about 20 radionuclides with radiations specifically chosen to treat at different depths in tissue--ranging from a few millimeters for intravascular therapy to a few centimeters in the case of large solid tumors. Systemic treatments with radiopharmaceuticals have grown from the traditional 131I to more than ten candidate nuclides which are to be labeled to tumor specific radiopharmaceuticals. Diagnostic radiopharmaceuticals are used in over 13 million procedures in the United States annually. About 40 nuclides are under investigation for these applications including single photon emitters for SPECT (single photon emission computed tomography) and positron emitters for PET (positron emission tomography). In addition to the mainstays of therapeutic and diagnostic radiology, radionuclides are widely used for in vitro tracers in the life sciences and represent one of the main tools in the field of molecular biology. PMID- 10724415 TI - Experimental determinations of commercial 'dose calibrator' settings for nuclides used in nuclear medicine. AB - Commercial reentrant ionization chambers ('dose calibrators') have become the de facto standard instrument for making radioactivity measurements in the radiopharmaceutical industry. Accurate activity measurements, however, depend on the application of the correct calibration factor (dial setting) for the radionuclide in the exact measurement geometry for that particular instrument. Two methods are described for the experimental determination of dial settings for the Capintec CRC-12 dose calibrator. Using these methods, we have determined new calibration factors for 18F, 62Cu, 186Re and 188Re in several geometries. These new factors are presented, along with previously determined settings, for nuclides commonly used in nuclear medicine. The differences between the calibration factors for the different geometries, as well as the differences between the manufacturer's recommended setting underscore the need for empirically determined dose calibrator settings. PMID- 10724416 TI - Importance of covariances for uncertainty estimates in measurement of radionuclide mixtures by multichannel counting. AB - Multichannel counting is often used when samples containing a mixture of known radionuclides are measured. Solutions to this method have been presented but to our knowledge, none of them took into account the covariances for evaluating the uncertainty of measurement. This paper presents new solutions for estimating uncertainties in measurement of radionuclide mixtures by multichannel counting and shows that covariances may contribute significantly to uncertainty of measurement so that they must be evaluated. PMID- 10724418 TI - An ionization chamber as a secondary standard for activity AB - A commercially-available radionuclide calibrator is used to maintain the unit of activity in the Slovak Institute of Metrology (SMU) and to check and verify activity meters in the field of nuclear medicine. The calibration of the instrument is described. The long-term stability is checked at regular intervals by means of 137Cs and 226Ra reference sources. A slow decrease of efficiency of about 2% per year has been observed, which has to be corrected for. An energy dependent efficiency curve of the ionization chamber is established by transfer of calibration factors from a similar instrument at the Physikalisch-Technische Bundesanstalt (PTB). The curve has been checked by measurements with activity standards of 60Co, 54Mn, 22Na, 85Sr, 65Zn, 88Y, 57Co, 139Ce, 137Cs, 125I, 241Am, 210Pb and 109Cd. The established curve permits one to calculate calibration factors for additional short-lived radionuclides used in nuclear medicine. An overall uncertainty was obtained for activity measurements with calculated calibration factors under standard measurement conditions of better than 1.8%. PMID- 10724417 TI - Application of alpha/beta discrimination in liquid scintillation counting for the purity control of 99mTc medical solutions. AB - 99mTc is a widely used nuclide in nuclear medicine for diagnosis. 99mTc solutions are eluted from 99Mo/99mTc generators as sodium chloride solutions. As the 99Mo source is a fission product extracted from the reprocessing of nuclear fuel, one might expect the presence of radioactive impurities. One point of special concern described in the 1997 European Pharmacopoeia (1996) is the possible contamination in alpha-impurities (U, Pu, Am, Cm, ...) which must be very low. In order to check this impurity concentration, we used liquid scintillation counting (LSC) with alpha/beta discrimination and a commercial extractive scintillator, Alphaex. We first made measurements using synthetic mixtures of alpha and beta-emitters and then using the real effluent. This paper describes the measurement procedure used with the optimum chemical conditions for a correct extraction of the alpha emitting nuclides. The sensitivity of the method is also described from the measurements made using the real effluent with an addition of a quantitative amount of alpha particle contamination. PMID- 10724419 TI - Standardization of 110mAg by liquid scintillation and 4pibeta-gamma coincidence counting AB - The nuclide 110Ag is a beta-gamma emitter with a very complex decay scheme, including more than 50 gamma-rays. It has been standardized by two methods: liquid scintillation counting (LSC) and 4pibeta(pc)-gamma coincidence measurements. In the LSC measurements the CIEMAT/NIST method was used, with 3H being used as a tracer for efficiency calculations. In the 4pibeta(pc)-gamma standardization, Monte Carlo calculations have been made to determine the optimal measurement conditions. Results obtained with both methods for the activity concentration of the solution are in good agreement. PMID- 10724420 TI - Analysis of detection-efficiency variation techniques for the implementation of the TDCR method in liquid scintillation counting AB - Detection efficiency variation is often used in liquid scintillation counting in order to evaluate a parameter in the detection efficiency calculation model. The aim of this paper is to compare the effects of three different detection efficiency variation methods in the activity measurement of three pure-beta radionuclides: 3H, 63Ni and 14C. Some general conclusions concerning the TDCR method are drawn. PMID- 10724421 TI - Measurements of linear absorption coefficients of liquid scintillators using synchrotron radiation AB - The use of liquid scintillation counting as a fundamental radionuclide standardisation method requires a correct description of the physical phenomena induced by the interaction of the ionising radiation with the liquid scintillator. In particular the standardisation of radionuclides decaying by electron capture as 55Fe and other electron-capture nuclides, requires the knowledge of total linear absorption coefficients of the liquid scintillator used for absolute activity measurement. Total linear absorption coefficient measurements have been undertaken using synchrotron radiation. Linear absorption coefficients of two widely used commercial liquid scintillators were measured in the 5.5-23 keV energy range. Small discrepancies were noted between theoretical and measured values of the Ultima-Gold mass attenuation coefficients for low energy photons. PMID- 10724422 TI - Standardization of electron-capture radionuclides by liquid scintillation counting AB - Liquid scintillation counting has proven to be an excellent technique to standardize beta-ray emitters with good accuracy. However, the success achieved in beta-ray nuclide standardization is not extensible for radionuclides decaying by electron-capture. Last comparison of 55Fe revealed discrepancies of 5% among laboratories. This paper presents two new versions of the computer program EMI. First we consider a better simulation of the photoelectric interaction of X-rays with the scintillator cocktail (EMI 1.2); and second, a more sophisticated KL1L2L3M atomic rearrangement model (EMI 2.0). We emphasize the importance of improving the simulation of the electron capture process to achieve better standardization of electron-capture nuclides by the CIEMAT/NIST method. PMID- 10724423 TI - A multimethodic and multiparametric system for standardisation of radionuclides AB - A measuring system with simultaneous application of four independent counting methods for standardization of solutions of different radionuclides is presented. The result obtained using four methods allows for mutual verification and gives the final result with a higher confidence level. The multidetection scintillation head for 4pi(LS)-gamma coincidence and anticoincidence with double-parametric registration, as well as the electronic diagram, is described. The four methods, comprised in the presented system, are described. Using the multimethodic and multiparametric system, massic activity of the solutions of the following radionuclides with various decay schemes has been measured: 133Ba, 90Sr, 99Mo, 85Sr, 65Zn, 139Ce, 192Ir, 204Tl, 60Co, 241Am, 152Eu, 169Yb. The results of the methods were analyzed using the figure of merit evaluated following the ISO/IEC guide 47-1. The system seems to allow the analysis of the correctness of the extrapolation curves, activity vs. the LSC efficiency, applied in 4pi(LS)-gamma methods. PMID- 10724424 TI - MAC3: an electronic module for the processing of pulses delivered by a three photomultiplier liquid scintillation counting system AB - In the field of radioactivity measurement, there is a constant need for high quality electronic modules such as ADCs, amplifiers, high voltage generators, dead-time modules, etc. and sometime there is a need for a specialized module not available on the market. The purpose of the module presented here, called MAC3 (module d'acquisition de coincidences triples), is to process the pulses delivered by the three detectors (photomultipliers) used in our liquid scintillation counting system. The dead-time generated by this module is of the extendible type and the dead-time corrections are made according to the live-time method. This module, which has been developed and tested at LPRI, can replace the complex interconnection of several independent and costly modules. The philosophy governing the choice and the implementation of this type of dead-time as well as the system used for the dead-time corrections is presented. The electronic scheme and the performances are also presented. This module is available in the NIM standard. PMID- 10724425 TI - A simple computing program for application of the TDCR method to standardization of pure-beta emitters AB - The Triple to Double Coincidence Ratio (TDCR) method is an absolute LSC method for the activity determination of many radionuclides. The method is based on a statistical description of the phenomena in a triple photomultiplier LS counter. The TDCRB-1 program was developed to calculate the radioactive concentration of a solution and is based on the TDCR method data processing. The program, in FORTRAN language has a modular structure, which easy enables to understand its functioning. The outline of the theory, a structure and algorithm of the program, as well as an example of the output results, are presented. PMID- 10724426 TI - Study of the stability problems of 110mAg samples in several commercial liquid scintillators AB - The stability of 110mAg samples for liquid scintillation measurements has been studied in four commercial scintillators: Insta-Gel Plus, Ultima-Gold, HiSafe II and HiSafe III. Samples using 15 ml of cocktail with four different levels of carrier, 0.0, 1.5, 3.0 and 6.0 microg of AgNO3, were prepared and their total count rate and spectral behaviour monitored for two months. Only samples in HiSafe III were completely stable for the full term studied. PMID- 10724427 TI - 222Rn detection at the microBq/m3 range in nitrogen gas and a new Rn purification technique for liquid nitrogen AB - For the Borexino solar neutrino experiment a concentration line for 222Rn from a large volume of nitrogen gas has been constructed. It is based on cryo-adsorption in a charcoal trap of very low intrinsic 226Ra contamination. Consequently, the blank for activity from the daughter nuclide 222Rn is very low. 222Rn is recorded with proportional counters. This allows the detection of Rn at concentrations below the microBq/m3 level in gaseous nitrogen. The removal of radon from liquid nitrogen is achieved by direct adsorption in the liquid phase. 222Rn-measurements on evaporated nitrogen with and without previous purification are reported. PMID- 10724428 TI - Oxidation-state distribution of plutonium in surface and subsurface waters at Thule, northwest Greenland. AB - The speciation of plutonium in Arctic waters sampled on the northwest Greenland shelf in August 1997 is discussed in this paper. Specifically, we report the results of analyses carried out on seawater sampled (a) close to the Thule air base where, in 1968, a US military aircraft carrying four nuclear weapons crashed on sea ice, releasing kilogram quantities of plutonium to the snow pack and underlying seabed sediments, and (b) at a reference station (Upernavik) located approximately 400 km to the south. The data show that most of the plutonium in the dissolved phase at Thule is in the form of Pu(V, VI) (mean: 68+/-6%; n = 6), with little if any distinction apparent between surface and bottom waters. Further, the oxidation state distribution at stations close to the accident site is similar to that measured at Upernavik, remote from this site. It is also similar to the distribution observed in shelf waters at mid-latitudes, suggesting that the underlying processes controlling plutonium speciation are insensitive to temperature over the range 0-25 degrees C. Measurements using tangential-flow ultrafiltration indicate that virtually all of the plutonium (including the fraction in a reduced chemical form) is present as fully dissolved species. Most of this plutonium would seem to be of weapons fallout origin, as the mean 238Pu/239,240Pu activity ratio in the water column (dissolved phase) at Thule (0.06+/-0.02; n = 10) is similar to the global fallout ratio at this latitude (approximately 0.04). Thus, there is little evidence of weapons-grade plutonium in the water column at Thule at the present time. PMID- 10724429 TI - Sequential method for the determination of uranium, thorium and 226Ra by liquid scintillation alpha spectrometry. AB - A new procedure for the determination of uranium, thorium and 226Ra from the same aliquot of an aqueous sample using extractant scintillators and liquid scintillation alpha spectrometry is proposed. The procedure is designed such that the same aqueous phase can be used in all the stages, with slight modifications. The procedure is thus very simple, requiring little manipulation of the sample. Testing of the procedure was performed obtaining satisfactory results and high reproducibility. PMID- 10724431 TI - Simultaneous determination of Ra and Th nuclides, 238U and 227Ac in uranium mining waters by gamma-ray spectrometry AB - For the investigation of flooding processes in uranium mines, Ra and Th nuclides as well as 238U and 227Ac activities in waters were simultaneous analyzed by gamma-ray spectrometry. The activities of 227Ac and 228Th, not directly determinable by gamma-ray spectrometry, can be calculated from two consecutive measurements (approximately 25 d delay) of the progeny 227Th and 224Ra. For the short-lived radionuclides 234Th, 227Th, 223Ra and 224Ra a correction of the results to the sampling date is necessary. PMID- 10724430 TI - Measurements of activation induced by environmental neutrons using ultra low level gamma-ray spectrometry. AB - The flux of environmental neutrons is being studied by activation of metal discs of selected elements. Near the earth's surface the total neutron flux is in the order of 10(-2) cm(-2)s(-1), which gives induced activities of a few mBq in the discs. Initial results from this technique, involving activation at ground level for several materials (W, Au, Ta, In, Re, Sm, Dy and Mn) and ultra low-level gamma-ray spectrometry in an underground laboratory located at 500 m.w.e., are presented. Diffusion of environmental neutrons in water is also measured by activation of gold at different depths. PMID- 10724432 TI - Coincidence-summing in gamma-ray spectrometry by excitation of matrix X-rays AB - Sources containing 75Se in a Pb(NO3)2 solution and in water matrices were measured using a well-type and a coaxial HPGe detector. The excitation and detection of characteristic lead X-rays increase coincidence losses from the peaks of 75Se in the case of the Pb(NO3)2 source. Sum peaks between gamma-rays of selenium and lead X-rays are observed in the spectra. The experimental results are in accordance with results of computations carried out with an extension of the GESPECOR Monte Carlo program. PMID- 10724433 TI - High accuracy measurement of the relative efficiency curve and determination of gamma-ray relative intensity for 38Cl AB - Relative efficiency curves were accurately determined for Ge gamma-ray detectors from measurements for radionuclides which emit two gamma-rays with essentially the same emission probabilities. The relative intensity of the 1643 and 2167 keV gamma-rays from 38Cl was determined to be 0.7496 (11) by using these highly accurate efficiency curves. PMID- 10724434 TI - High-pressure xenon detectors for gamma-ray spectrometry AB - The main results of long-term research on compressed xenon detector properties conducted at the laboratory of cosmic physics of MEPhI are given along with a description of the latest gamma-ray spectrometers based on this work. It is shown that using xenon as working substance, it is possible to create a gamma-ray spectrometer with high energy resolution. The construction and the main physical, technical and operation performances of xenon gamma-ray spectrometers based on ionization chambers of various configurations are described. For a gamma-ray spectrometer with a cylindrical ionization chamber and shielding grid, an energy resolution of about 14 keV (10 keV intrinsic resolution) for gamma-ray line of 662 keV is obtained. The characteristics of these detectors allow one to apply them in various fields of science and engineering, moreover, their good spectrometric properties provide the opportunity to use them for metrology measurements. PMID- 10724435 TI - Fast procedures for coincidence-summing correction in gamma-ray spectrometry AB - Simplified and fast procedures for coincidence-summing correction in gamma-ray spectrometry were investigated. These procedures are based on the usual theoretical expressions of the correction factors, but differ in the determination of the total efficiency curve based on the following approximations: (a) replacement, below the knee efficiency value, of the total efficiency by the full-energy peak efficiency; and (b) use of linear interpolations (in log-log plot) between only two experimental points above the knee efficiency value; or (c) assumption of a peak-to-total efficiency ratio independent on the counting geometry; or (d) assumption of a constant relation between the peak-to-total efficiency ratios and the photoelectric-to-total cross section ratios. The above approximations were separately assumed for determination of the coincidence-summing correction factors for nuclides with complex decay scheme (133Ba, 134Cs, 152Eu) and for 60Co and 88Y measured on a 15% relative efficiency p-type coaxial HPGe detector, for three source-detector geometries: point source placed on top of and at 10 cm from the detector window, and 1 l Marinelli beaker filled with aqueous solution. The results were compared with those based on more accurate experimental determinations of the total efficiency curve from measurements of standard sources of eight different single gamma-ray emitters. The usefulness of each simplified procedure is evaluated with respect to its accuracy and to the reduction of the number of standard sources and measurement time. PMID- 10724436 TI - Isotopic analysis of uranium in U3O8 by passive gamma-ray spectrometry AB - Passive gamma-ray spectrometry was applied to analyze the isotopic composition of uranium in U3O8. Depleted and enriched U3O8 standard reference materials were used to calibrate the system. An independent calibration was performed by standard gamma-ray point sources. U3O8 SRM samples of the 950 series were analyzed. The present results show that the isotopic abundances of 235U in SRMs 950, 950a and 950b are higher by +3.6, +0.9 and +0.9% (relative deviation) than the natural value 0.7200%, while relative precisions were +/-0.4, +/-0.7 and +/ 0.3%, respectively. PMID- 10724437 TI - Measurements of the average thickness of material using scattered radiation AB - A method for determination of the average thickness of material placed between a gamma-ray source and a detector is presented. The method requires the measurement of the ratio between the number of gamma-rays scattered at small angles in the material and registered in the spectrum and the number of gamma-rays penetrating the material without interacting with it and registering in the full-energy peak. The relation describing that ratio in terms of the average gamma-ray path length in the sample, the energy interval where the scattered radiation is measured and the detector response to unscattered gamma-rays was verified by measurements of gamma-ray path lengths in material with known thicknesses at energies of 662 and 1115 keV. Differences between the measured and known thicknesses were less than 15% for thicknesses of copper between 2.5 and 17 g/cm2. PMID- 10724438 TI - Quality control of gamma-ray spectrometry measurements AB - Specific techniques have been implemented to assess the quality of routine gamma ray measurements and the associated analysis procedures. Appropriate steps monitor three factors influencing the quality of performance: the performance of the spectrometer, the quality of the measurement and the quality of the peak analysis. In order to illustrate the approach, time dependencies of parameters describing the performance of the spectrometer, the quality of the measurements and the sample activities are presented. PMID- 10724439 TI - Solidang, a computer code for the computation of the effective solid angle and correction factors for gamma spectroscopy-based waste assay AB - Detection efficiency calibration of gamma spectroscopy for waste assay systems is a difficult and time-consuming task. Commonly, reference waste packages are used for efficiency calibration but these are sometimes difficult to build, are expensive and have limited use since there is large variability of parameters which influence the assay. Numerical calibrations, however, are an interesting alternative and have begun to find more and more application in waste assay. A dedicated computer code, Solidang, has been developed to compute detection efficiencies for gamma waste assay. Solidang uses the effective solid angle approach to derive detection efficiency and aims at improving and facilitating the calibration of gamma waste assay systems and at serving as a tool to investigate the influence of the many system and sample parameters involved. PMID- 10724440 TI - Towards a proper modeling of detector and source characteristics in Monte Carlo simulations AB - We performed a systematic study on the influence of different source configurations on the reliability of Monte Carlo calculations for the response of Ge detectors. Calculated full-energy peak efficiencies are compared to experimental values in the energy range 46-1800 keV. Setups with different characteristics are considered, from point-like to volume sources. Among the latter there are filters and aqueous matrices and sediments in Petri boxes or Marinelli beakers. The analysis of the deviations between experimental and calculated results for the different configurations enables us to detect inaccuracies in the description of detector and source characteristics and to improve them. By means of this procedure satisfactory results of the efficiencies were obtained in the whole energy range. For all setups, the deviations average 1.5%, except for the sediment sources where they are up to 3.3%. PMID- 10724441 TI - Monte Carlo calculations of the total-to-peak ratio in gamma-ray spectrometry AB - Monte Carlo calculations of the total-to-peak ratio for a coaxial closed end n type gamma-ray detector were performed and the results compared to the measured values. Whereas reasonable agreement between the calculated and measured spatial dependencies was achieved, the calculation overestimates the absolute experimental values by about 10%. The spatial dependence of the total-to-peak ratio is discussed in terms of detector geometry. PMID- 10724442 TI - Efficiency transfer and coincidence summing corrections for gamma-ray spectrometry AB - ETNA (Efficiency Transfer for Nuclide Activity measurements) is a new computer program providing both detector efficiency transfer for various types of sources for coaxial source-detector geometries and coincidence summing corrections. This program, available for use on PCs, has a direct link to the computerised database NUCLEIDE in order to automatically import the necessary radionuclide decay data. The specific features and facilities are outlined and examples are given. PMID- 10724443 TI - Role of IL-6 in the induction of hepatic metallothionein in mice after partial hepatectomy. AB - Metallothioneins (MT) are induced upon partial hepatectomy (PH), possibly mediated by various cytokines. In the present study, we studied cytokine dependent MT synthesis in partially hepatectomized IL-6 gene knock-out (GKO) mice in the remaining lobe of the liver. We focused on IL-6, TNFalpha and IL-1beta, the major cytokines thought to be involved in MT synthesis. The IL-6 GKO mice and B6J129Sv (wild-type control) mice were subjected to 70% PH or laparotomy. We found that MT was significantly decreased in IL-6 GKO mice, although PH induced hepatic MT in both strains of mice. Laparotomy induced MT in the liver of wild type mice but not in IL-6 GKO mice. Pretreatment of IL-6 GKO mice with rIL-6 (5 microg/mouse) restored hepatic MT synthesis. Serum IL-6 level in wild-type mice was maximal at 6 h after surgery and decreased thereafter. Serum IL-1beta was the same in both strains of mice. Serum TNFalpha basal level in IL-6 GKO mice was higher than in wild-type mice. PH caused an increase in serum TNFalpha level in both strains of mice, and it was two times higher in IL-6 GKO mice than in wild type mice at 18 h after surgery. We conclude that IL-6 plays a predominant role in hepatic MT synthesis after PH, but that IL-6 GKO mice still reserve the capacity to synthesize MT by an as yet unidentified mechanism. PMID- 10724444 TI - Wood smoke-induced airway hyperreactivity in guinea pigs: time course, and role of leukotrienes and hydroxyl radical. AB - A prior airway exposure to wood smoke induces a tachykinin-dependent increase in airway responsiveness to the subsequent smoke inhalation in guinea pigs (Life Sci. 63: 1513, 1998). To further investigate the time course of, and the contribution of other chemical mediators to, this smoke-induced airway hyperresponsiveness (SIAHR), two smoke challenges (each 10 ml) separated by 30 min were delivered into the lungs of anesthetized guinea pigs by a respirator. In the control animals, the SIAHR was evidenced by the bronchoconstrictive response to the second smoke challenge (SM2) which was approximately 5.2-fold greater than that to the first challenge (SM1). This SIAHR was alleviated by shortening the elapsed time between SM1 and SM2 to 10 min or by extending it to 60 min, and was abolished by extending it to 120 min. This SIAHR was reduced by pretreatment with either MK-571 (a leukotriene D4-receptor antagonist) or dimethylthiourea (a hydroxyl radical scavenger), but was not affected by pretreatment with either pyrilamine (a histamine H1-receptor antagonist) or indomethacin (a cyclooxygenase inhibitor). The smoke-induced reduction in the neutral endopeptidase activity (a major enzyme for tachykinin degradation) measured in airway tissues excised 30 min post SM1 was largely prevented by pretreatment with dimethylthiourea. However, this reduction was not seen in airway tissues excised 120 min post SM1. These results suggest that 1) the SIAHR to inhaled wood smoke has a rapid onset time following smoke inhalation and lasts for less than two hours, 2) leukotrienes and hydroxyl radical may play contributory roles in the development of this SIAHR, and 3) hydroxyl radical is the major factor responsible for the smoke-induced inactivation of airway neutral endopeptidase, which may possibly participate in the development of this SIAHR. PMID- 10724445 TI - Stimulatory effect of trans-cinnamaldehyde on noradrenaline secretion in guinea pig ileum myenteric nerve terminals. AB - The effect of trans-cinnamaldehyde (CNMA) on the release of noradrenaline (NA) from nerve terminal was investigated using isolated ileal synaptosomes of guinea pig. Release was determined as the amount of NA, quantified by h.p.l.c. electrochemical detection, from samples incubated with CNMA minus that in parallel blanks treated with same volume of vehicle. CNMA stimulated the secretion of NA in a concentration-dependent manner from 5 microM to 50 microM, while the value of lactate dehydrogenase in the incubated medium was not influenced by CNMA. However, trans-cinnamic acid, cinnamoyl chloride and cinnamamide failed to produce similar effect. Specific action of CNMA can thus be considered. Guanethidine inhibited the release of NA by CNMA in a concentration- dependent manner. Saxitoxin attenuated the action of CNMA at concentrations sufficient to block sodium channels. The depolarizing effect of CNMA on the membrane potential was also illustrated by a concentration-dependent increase in the fluorescence of bisoxonol, a potential sensitive dye. The NA releasing action of CNMA was deleted by removal of calcium chloride from the bathing medium. This action of CNMA was also attenuated by Rp-cAMP at concentrations sufficient to inhibit the action of cyclic AMP. These findings suggest that CNMA can depolarize the membrane to result in a calcium-dependent and cyclic AMP-related release of NA from noradrenergic terminals. PMID- 10724446 TI - CTLA4IgG gene delivery prevents autoantibody production and lupus nephritis in MRL/lpr mice. AB - MRL/MP-lpr/lpr (MRL/lpr) mice spontaneously develop an autoimmune syndrome closely resembling systemic lupus erythematosus (SLE) in humans, characterized by hypergammaglobulinemia, various autoantibody production, and the development of fatal glomerulonephritis. We have previously demonstrated that systemic administration of soluble form of CTLA4IgG prevented autoantibody-related diseases in MRL/lpr mice. To test the potential protective effects of CTLA4IgG gene delivery on the development of lupus nephritis, we injected MRL/lpr mice with a recombinant adenovirus vector containing CTLA4IgG gene, Adex1CACTLA4IgG (AdCTLA4IgG). It was demonstrated that a single administration of intravenous injection of AdCTLA4IgG into MRL/lpr mice resulted in almost complete amelioration of lupus nephritis. PMID- 10724447 TI - Is Ca2+ the second messenger in the response to melatonin in cuckoo wrasse melanophores? AB - Pigment aggregation in melanophores of Labrus ossifagus is controlled by an alpha2-adrenoceptor and is somehow modulated by melatonin. The signal transduction mechanisms seem to involve both an attenuation of cAMP and an increase in intracellular Ca2+, inhibiting protein kinase A or activating a phosphatase, respectively. These effects result in dephosphorylation, which in turn induces aggregation. Various alpha2-adrenoceptor agonists attenuate cAMP levels or increase the concentration of intracellular Ca2+. Noradrenaline, for example, lowers cAMP but does not affect the calcium signal whereas B-HT 920, an alpha2-adrenoceptor specific agonist, does not induce a cAMP decrease but does appear to induce an increase in intracellular Ca2+. This later inference is drawn from experiments with BAPTA/AM, an intracellular calcium chelator, which counteracts the aggregation induced by B-HT 920. Interestingly, the very potent alpha2-adrenoceptor agonist medetomidine apparently activates both signal transduction pathways, which could explain its high efficacy in producing aggregation. Melatonin itself does not cause pigment aggregation, but it potentiates noradrenaline-induced aggregation. It has been suggested that melatonin receptors and alpha2-adrenoceptors follow the same signal transduction pathway, i.e. an attenuation of cAMP. In our experiments, melatonin did not reduce cAMP levels; instead it appears to increase Ca2+ concentration, since melatonin-potentiated aggregation was inhibited by BAPTA/AM. Thus, aggregation amplified by melatonin is probably not mediated by a further decrease in cAMP, but by the same signal transduction mechanism as B-HT 920, i.e. an increase in Ca2+. This further strengthens the suggestion that melatonin and B-HT 920 bind to the same site, but it is unclear if that particular site is on the melatonin receptor or the alpha2-adrenoceptor. PMID- 10724448 TI - Activation of serotonin (5-HT)1A receptors inhibits amphetamine sensitization in mice. AB - The effects of serotonin (5-HT)1A drugs on the development and expression of sensitization to the locomotor effect of amphetamine (AMPH) were studied in mice. 8-Hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT), a 5-HT1A agonist, dose dependently reduced the expression of AMPH (2.5 mg/kg)-induced sensitization. The latter inhibitory effect of 8-OH-DPAT was reversed by (S)-N-tert-butyl-3-(4-(2 methoxyphenyl)piperazin-1-yl)-2-phenyl propamine (WAY 100135), a 5-HT1A antagonist. WAY 100135 given alone did not affect expression of AMPH sensitization. Combined injections of 8-OH-DPAT, but not WAY 100135, with AMPH (2.5 mg/kg) during the development of sensitization, protected against the expression of sensitization to a challenge dose of AMPH (2.5 mg/kg) 3 days after withdrawal. The above inhibitory effect of 8-OH-DPAT on the development of AMPH sensitization was blocked by pretreatment with WAY 100135. The AMPH-induced conditioned locomotion was unaffected by pretreatment with 8-OH-DPAT. These results indicate that 5-HT1A receptors are not involved in AMPH-induced sensitization per-se, whereas their pharmacological activation leads to the inhibition of both the development and the expression of AMPH-induced sensitization. PMID- 10724449 TI - Positive cooperative effects between receptors induced by an anti-human growth hormone allosteric monoclonal antibody. AB - Monoclonal antibodies (MAb) anti-human growth hormone (hGH) termed MAb AE5, AC8 and F11 recognize a cluster of epitopes left exposed after hormone binding to receptors. Since these MAb were able to produce either positive (MAb AE5) or negative (MAb AC8 and F11) allosteric effects on hGH binding, the purpose of this work was to further characterize MAb behavior. Results indicated a straight correlation between MAb allosteric effects and affinity constant values for binding of different hGH:MAb complexes to lactogenic receptors from rat liver. Affinity of hGH:MAb AE5 as well as hGH:Fab AE5 complexes enhanced proportionally to the fraction of occupied receptors and Hill coefficients higher than 1 were obtained, suggesting the induction of positive cooperative effects between membrane-bound receptors. On the other hand, hGH:MAb AC8 and hGH:MAb F11 complexes binding affinity to lactogenic sites could not be related to receptor occupancy degree. It is proposed that binding of hGH:MAb AE5 complexes to receptors would elicit a conformational change on adjacent receptor molecules leading to an increase of their affinity to bind subsequent hGH:MAb AE5 complexes. PMID- 10724450 TI - Effect of low flow ischemia-reperfusion injury on liver function. AB - The release of liver enzymes is typically used to assess tissue damage following ischemia-reperfusion. The present study was designed to determine the impact of ischemia-reperfusion on liver function and compare these findings with enzyme release. Isolated, perfused rat livers were subjected to low flow ischemia followed by reperfusion. Alterations in liver function were determined by comparing rates of oxygen consumption, gluconeogenesis, ureagenesis, and ketogenesis before and after ischemia. Lactate dehydrogenase (LDH) and purine nucleoside phosphorylase (PNP) activities in effluent perfusate were used as markers of parenchymal and endothelial cell injury, respectively. Trypan blue staining was used to localize necrosis. Total glutathione (GSH + GSSG) and oxidized glutathione (GSSG) were measured in the perfusate as indicators of intracellular oxidative stress. LDH activity was increased 2-fold during reperfusion compared to livers kept normoxic for the same time period whereas PNP activity was elevated 5-fold under comparable conditions. Rates of oxygen consumption, gluconeogenesis, and ureagenesis were unchanged after ischemia, but ketogenesis was decreased 40% following 90 min ischemia. During reperfusion, the efflux rates of total glutathione and GSSG were unchanged from pre-ischemic values. Significant midzonal staining of hepatocyte nuclei was observed following ischemia-reperfusion, whereas normoxic livers had only scattered staining of individual cells. Reperfusion of ischemic liver caused release of hepatic enzymes and midzonal cell death, however, several major liver functions were unaffected under these experimental conditions. These data indicate that there were negligible changes in liver function in this model of ischemia and reperfusion despite substantial enzyme release from the liver and midzonal cell death. PMID- 10724451 TI - Dehydrogenase and oxoreductase activities of porcine placental 11beta hydroxysteroid dehydrogenase. AB - Dehydrogenase (cortisol to cortisone) and oxoreductase (cortisone to cortisol) activities of porcine placental 11beta-hydroxysteroid dehydrogenase (11beta-HSD) were measured in tissue fragment cultures on day 75 of gestation. Dehydrogenase activity was over fivefold greater than oxoreductase activity (p < .001). There were positive linear associations (p < .01) between net dehydrogenase activity (dehydrogenase minus oxoreductase) and fetal weight, fetal length, and placental weight. These data indicate a predominance of placental 11beta-HSD dehydrogenase activity at this gestational stage that would insure a net conversion of cortisol to cortisone as it traverses the placenta. The data further suggest that 11beta HSD activities may provide an optimal glucocorticoid environment that is supportive of enhanced fetal and placental growth. PMID- 10724452 TI - Fendiline increases [Ca2+]i in Madin Darby canine kidney (MDCK) cells by releasing internal Ca2+ followed by capacitative Ca2+ entry. AB - The effect of fendiline, a documented inhibitor of L-type Ca2+ channels and calmodulin, on Ca2+ signaling in Madin Darby canine kidney (MDCK) cells was investigated using fura-2 as a Ca2+ probe. Fendiline at 5-100 microM significantly increased [Ca2+]i concentration-dependently. The [Ca2+]i rise consisted of an initial rise and a slow decay. External Ca2+ removal partly inhibited the Ca2+ signals induced by 25-100 microM fendiline by reducing both the initial rise and the decay phase. This suggests that fendiline triggered external Ca2+ influx and internal Ca2+ release. In Ca(2+)-free medium, pretreatment with 50 microM fendiline nearly abolished the [Ca2+]i rise induced by 1 microM thapsigargin, an endoplasmic reticulum Ca2+ pump inhibitor, and vice versa, pretreatment with thapsigargin prevented fendiline from releasing internal Ca2+. This indicates that the internal Ca2+ source for fendiline overlaps with that for thapsigargin. At a concentration of 50 microM, fendiline caused Mn2+ quench of fura-2 fluorescence at the 360 nm excitation wavelenghth, which was inhibited by 0.1 mM La3+ by 50%, implying that fendiline-induced Ca2+ influx has two components separable by La3+. Consistently, 0.1 mM La3+ pretreatment suppressed fendiline-induced [Ca2+]i rise, and adding La3+ during the rising phase immediately inhibited the signal. Addition of 3 mM Ca2+ increased [Ca2+]i after preincubation with 50-100 microM fendiline in Ca(2+)-free medium. However, 50-100 microM fendiline inhibited 1 microM thapsigargin-induced capacitative Ca2+ entry. Pretreatment with 40 microM aristolochic acid to inhibit phospholipase A2 inhibited 50 microM fendiline-induced internal Ca2+ release by 48%, but inhibition of phospholipase C with 2 microM U73122 or inhibition of phospholipase D with 0.1 mM propranolol had no effect. Collectively, we have found that fendiline increased [Ca2+]i in MDCK cells by releasing internal Ca2+ in a manner independent of inositol-1,4,5-trisphosphate (IP3), followed by external Ca2+ influx. PMID- 10724453 TI - B-chromosome evolution. AB - B chromosomes are extra chromosomes to the standard complement that occur in many organisms. They can originate in a number of ways including derivation from autosomes and sex chromosomes in intra- and interspecies crosses. Their subsequent molecular evolution resembles that of univalent sex chromosomes, which involves gene silencing, heterochromatinization and the accumulation of repetitive DNA and transposons. B-chromosome frequencies in populations result from a balance between their transmission rates and their effects on host fitness. Their long-term evolution is considered to be the outcome of selection on the host genome to eliminate B chromosomes or suppress their effects and on the B chromosome's ability to escape through the generation of new variants. Because B chromosomes interact with the standard chromosomes, they can play an important role in genome evolution and may be useful for studying molecular evolutionary processes. PMID- 10724454 TI - Mapping the bacterial cell architecture into the chromosome. AB - A genome is not a simple collection of genes. We propose here that it can be viewed as being organized as a 'celluloculus' similar to the homunculus of preformists, but pertaining to the category of programmes (or algorithms) rather than to that of architectures or structures: a significant correlation exists between the distribution of genes along the chromosome and the physical architecture of the cell. We review here data supporting this observation, stressing physical constraints operating on the cell's architecture and dynamics, and their consequences in terms of gene and genome structure. If such a correlation exists, it derives from some selection pressure: simple and general physical principles acting at the level of the cell structure are discussed. As a first case in point we see the piling up of planar modules as a stable, entropy driven, architectural principle that could be at the root of the coupling between the architecture of the cell and the location of genes at specific places in the chromosome. We propose that the specific organization of certain genes whose products have a general tendency to form easily planar modules is a general motor for architectural organization in the bacterial cell. A second mechanism, operating at the transcription level, is described that could account for the efficient building up of complex structures. As an organizing principle we suggest that exploration by biological polymers of the vast space of possible conformation states is constrained by anchoring points. In particular, we suggest that transcription does not always allow the 5'-end of the transcript to go free and explore the many conformations available, but that, in many cases, it remains linked to the transcribing RNA polymerase complex in such a way that loops of RNA, rather than threads with a free end, explore the surrounding medium. In bacteria, extension of the loops throughout the cytoplasm would therefore be mediated by the de novo synthesis of ribosomes in growing cells. Termination of transcription and mRNA turnover would accordingly be expected to be controlled by sequence features at both the 3'- and 5'-ends of the molecule. These concepts are discussed taking into account in vitro analysis of genome sequences and experimental data about cell compartmentalization, mRNA folding and turnover, as well as known structural features of protein and membrane complexes. PMID- 10724455 TI - The breast cancer susceptibility gene, BRCA2: at the crossroads between DNA replication and recombination? AB - The identification and cloning of the familial breast cancer susceptibility gene, BRCA2, has excited much interest in its biological functions. Here, evidence is reviewed that the protein encoded by BRCA2 has an essential role in DNA repair through its association with mRad51, a mammalian homologue of bacterial and yeast proteins involved in homologous recombination. A model is proposed that the critical requirement for BRCA2 in cell division and the maintenance of chromosome stability stems from its participation in recombinational processes essential for DNA replication. PMID- 10724456 TI - The neuron as a dynamic electrogenic machine: modulation of sodium-channel expression as a basis for functional plasticity in neurons. AB - Neurons signal each other via regenerative electrical impulses (action potentials) and thus can be thought of as electrogenic machines. Voltage-gated sodium channels produce the depolarizations necessary for action potential activity in most neurons and, in this respect, lie close to the heart of the electrogenic machinery. Although classical neurophysiological doctrine accorded 'the' sodium channel a crucial role in electrogenesis, it is now clear that nearly a dozen genes encode distinct sodium channels with different molecular structures and functional properties, and the majority of these channels are expressed within the mammalian nervous system. The transcription of these sodium channel genes, and the deployment of the channels that they encode, can change significantly within neurons following various injuries. Moreover, the transcription of these genes and the deployment of various types of sodium channels within neurons of the normal nervous system can change markedly as neurons respond to changing milieus or physiological inputs. As a result of these changes in sodium-channel expression, the membranes of neurons may be retuned so as to alter their transductive and/or encoding properties. Neurons within the normal and injured nervous system can thus function as dynamic electrogenic machines with electroresponsive properties that change not only in response to pathological insults, but also in response to shifting functional needs. PMID- 10724457 TI - The labile brain. I. Neuronal transients and nonlinear coupling. AB - In this, the first of three papers, the nature of, and motivation for, neuronal transients is described in relation to characterizing brain dynamics. This paper deals with some basic aspects of neuronal dynamics, interactions, coupling and implicit neuronal codes. The second paper develops neuronal transients and nonlinear coupling in the context of dynamic instability and complexity, and suggests that instability or lability is necessary for adaptive self organization. The final paper addresses the role of neuronal transients through information theory and the emergence of spatio-temporal receptive fields and functional specialization. By considering the brain as an ensemble of connected dynamic systems one can show that a sufficient description of neuronal dynamics comprises neuronal activity at a particular time and its recent history This history constitutes a neuronal transient. As such, transients represent a fundamental metric of neuronal interactions and, implicitly, a code employed in the functional integration of brain systems. The nature of transients, expressed conjointly in distinct neuronal populations, reflects the underlying coupling among populations. This coupling may be synchronous (and possibly oscillatory) or asynchronous. A critical distinction between synchronous and asynchronous coupling is that the former is essentially linear and the latter is nonlinear. The nonlinear nature of asynchronous coupling enables the rich, context-sensitive interactions that characterize real brain dynamics, suggesting that it plays a role in functional integration that may be as important as synchronous interactions. The distinction between linear and nonlinear coupling has fundamental implications for the analysis and characterization of neuronal interactions, most of which are predicated on linear (synchronous) coupling (e.g. cross-correlograms and coherence). Using neuromagnetic data it is shown that nonlinear (asynchronous) coupling is, in fact, more abundant and can be more significant than synchronous coupling. PMID- 10724458 TI - The labile brain. II. Transients, complexity and selection. AB - The successive expression of neuronal transients is related to dynamic correlations and, as shown in this paper, to dynamic instability. Dynamic instability is a form of complexity, typical of neuronal systems, which may be crucial for adaptive brain function from two perspectives. The first is from the point of view of neuronal selection and self-organizing systems: if selective mechanisms underpin the emergence of adaptive neuronal responses then dynamic instability is, itself, necessarily adaptive. This is because dynamic instability is the source of diversity on which selection acts and is therefore subject to selective pressure. In short, the emergence of order, through selection, depends almost paradoxically on the instabilities that characterize the diversity of brain dynamics. The second perspective is provided by information theory. PMID- 10724459 TI - The labile brain. III. Transients and spatio-temporal receptive fields. AB - In this paper we consider an approach to neuronal transients that is predicated on the information they contain. This perspective is provided by information theory, in particular the principle of maximum information transfer. It is illustrated here in application to visually evoked neuronal transients. The receptive fields that ensue concur with those observed in the real brain, predicting, almost exactly, functional segregation of the sort seen in the visual system. This information theoretical perspective can be reconciled with a selectionist stance by noting that a high mutual information among neuronal systems and the environment has, itself, adaptive value and will be subject to selective pressure, at any level one cares to consider. PMID- 10724460 TI - Bilateral receptive field neurons and callosal connections in the somatosensory cortex. AB - Earlier studies recording single neuronal activity with bilateral receptive fields in the primary somatosensory cortex of monkeys and cats agreed that the bilateral receptive fields were related exclusively to the body midline and that the ipsilateral information reaches the cortex via callosal connections since they are dense in the cortical region representing the midline structures of the body while practically absent in the regions representing the distal extremities. We recently found a substantial number of neurons with bilateral receptive fields on hand digits, shoulders-arms or legs-feet in the caudalmost part (areas 2 and 5) of the postcentral gyrus in awake Japanese monkeys (Macaca fuscata). I review these results, discuss the functional implications of this bilateral representation in the postcentral somatosensory cortex from a behavioural standpoint and give a new interpretation to the midline fusion theory. PMID- 10724461 TI - Molecular mechanisms of sleep-wake regulation: a role of prostaglandin D2. AB - Prostaglandin (PG) D2 is a major prostanoid in the brains of rats and other mammals, including humans. When PGD synthase (PGDS), the enzyme that produces PGD2 in the brain, was inhibited by the intracerebroventricular infusion of its selective inhibitors, i.e. tetravalent selenium compounds, the amount of sleep decreased both time and dose dependently. The amount of sleep of transgenic mice, in which the human PGDS gene had been incorporated, increased several fold under appropriate conditions. These data indicate that PGDS is a key enzyme in sleep regulation. In situ hybridization, immunoperoxidase staining and direct enzyme activity determination of tissue samples revealed that PGDS is hardly detectable in the brain parenchyma but is localized in the membrane systems surrounding the brain, namely, the arachnoid membrane and choroid plexus, from which it is secreted into the cerebrospinal fluid (CSF) to become beta-trace, a major protein component of the CSF. PGD2 exerts its somnogenic activity by binding to PGD2 receptors exclusively localized at the ventrorostral surface of the basal forebrain. When PGD2 was infused into the subarachnoid space below the rostral basal forebrain, striking expression of proto-oncogene Fos immunoreactivity (FosIR) was observed in the ventrolateral preoptic area (VLPO), a putative sleep centre, concurrent with sleep induction. Fos expression in the VLPO was positively correlated with the preceding amount of sleep and negatively correlated with Fos expression in the tuberomammillary nucleus (TMN), a putative wake centre. These observations suggest that PGD2 may induce sleep via leptomeningeal PGD2 receptors with subsequent activation of the VLPO neurons and downregulation of the wake neurons in the TMN area. Adenosine may be involved in the signal transduction associated with PGD2. PMID- 10724462 TI - Whorl morphogenesis in the dasycladalean algae: the pattern formation viewpoint. AB - The dasycladalean algae produce diverse whorled structures, among which the best known are the vegetative and reproductive whorls of Acetabularia acetabulum. In this paper, we review the literature pertaining to the origin of these structures. The question is addressed in terms of the necessary pattern-forming events and the possible mechanisms involved, an outlook we call the pattern formation viewpoint. The pattern-forming events involved in the morphogenesis of the vegetative and reproductive whorls of Acetabularia have been used to define five and six morphogenetic stages, respectively. We discuss three published mechanisms which account, at least in part, for the pattern-forming events. The mechanisms are mechanical buckling of the cell wall, reaction-diffusion of morphogen molecules along the cell membrane, and mechanochemical interactions between Ca2+ ions and the cytoskeleton in the cytosol. The numerous differences between these mechanisms provide experimental grounds to test their validity. To date, the results of these experiments point towards reaction diffusion as the most likely patterning mechanism. Finally, we consider the evolutionary origin of the vegetative and reproductive whorls and provide mechanistic explanations for some of the major evolutionary advances. PMID- 10724463 TI - Shapes of river networks and leaves: are they statistically similar? AB - The structure of river networks is compared with the vein structure of leaves. The two structures are visually similar at the smaller scales. The statistics of branching and side branching are nearly identical. The branching structure of diffusion-limited aggregation clusters is also similar and can provide an explanation for the structure of river networks. The origin of the self-similar branching and side branching of the vein structure in leaves is not clear but it appears to be an optimal network in terms of transporting nutrients to all parts of the leaf with the least total resistance. PMID- 10724464 TI - Why feminism in public health? AB - The issues raised in this editorial and exemplified within a number of the studies reported in this issue indicate new directions for public health, directions which take feminist scholarship, both outside and within the medical framework, into account. The changing potential of feminist public health, as derived from the articles in this issue, can be summarised within the following issues: new research areas, positioning women as actors, development of theoretical frameworks, reflexive theory of science, interplay between sex and gender, gender-sensitive methods, diversities among women/men, pro-feminist research on men's health and using the results for change. Thus, feminist public health represents a shift towards the new public health, with holistic and multidisciplinary activities, based on theoretical pluralism, multiple perspectives and collective actions with the aim of improving the health of gender-subordinated groups. PMID- 10724465 TI - Feminist public health practice and population-based health strategies: breast cancer screening in Australia. PMID- 10724466 TI - Prevalence of sexualized violence among women. A population-based study in a primary healthcare district. AB - In order to estimate the prevalence of sexual and physical abuse, postal questionnaires were sent to a random sample of 251 adult women in a primary care district. Three yes/no-questions were asked on these topics, and a fourth inquired into effects on health. Space was left for open-ended answers in which yes-responders were invited to write about their experiences. A total of 175 women (70%) answered, and 25 (14%) of these reported abuse. Nine (5%) had experienced women battering. Thirteen (7.5%) reported sexual abuse as an adult, and 12 (7%) as a child. Many told their stories. Methodological shortcomings in this study might explain why the rates are somewhat lower than in other investigations. However, our figures verified that abuse of women is a common social phenomenon. They also confirmed abuse as a major health problem for women. One-third of respondents explicitly reported effects on health. Others, though negating such effects, described them in written narratives. In order to study this contradiction, in-depth interviews with abused women are recommended. PMID- 10724467 TI - Encouraging the strengths of women patients. A case study from general practice on empowering dialogues. AB - This case study illustrates how the use of empowering dialogues in general practice can contribute to alternative images of women, by identifying and emphasizing their strong points. It is a single case study, sampled theoretically from a series of 37 consultations during which key questions about self-assessed health resources were put to women patients. Two women GPs and their consultations were studied. An 18-min dialogue between a 52-year-old woman GP and a 69-year-old woman patient with asthma and back pain was audiotaped and transcribed according to Nessa's principles, supported by pragmatic linguistic theory. The woman's answers changed the doctor's perception of the patient, from that of a passive and resigned sufferer, to that of a strong woman who was active in spite of her pain. Acknowledging this, alternative paths of management could be chosen. In conclusion, disempowering medicalization of women patients can be opposed by resource oriented dialogues in clinical work. However, to change cultural images requires more than individual action. PMID- 10724468 TI - Gender and the capacity of women with NIDDM to implement medical advice. AB - This qualitative study of women with non-insulin dependent diabetes mellitus (NIDDM) examined constructions of their diabetes management and socio-familial relationships as potential sources of support. Semi-structured interview data was collected from 16 women. The transcripts were analysed with the aim of examining the ways in which gender relations structured women's accounts of health-related behaviours. Women talked about themselves as wives, mothers, being pregnant and parenting, and friends of other women in ways that demonstrated how caring for others impeded their capacity to care for themselves. Meeting the food preferences of husbands and dietary requirements of diabetic husbands were dominant themes in women's accounts of marriage, and in various ways women justified their husbands' lack of support. Furthermore, the care of others during pregnancy and parenting was also an obstacle to women caring for themselves. An awareness of the gender politics inherent within social and family contexts is crucial to improving the effectiveness of medical advice for diabetes management. PMID- 10724469 TI - Young women's perspectives on public health knowledge and adolescent bodies. AB - This study examines health information about adolescent bodies from the perspective of young middle-class women, with the aim of determining ways in which health and education professionals could better serve young women. This qualitative study comprised "memory work" with eight Finnish women students. The findings suggest that young women prefer the information on menstruation that is given by public health professionals, because of its impersonal form, rather than personal encounters with relatives and peers. The information provided by public health professionals does not, however, meet the women's needs. This discrepancy can be explained by the gap between the narrow medical discourse on women's bodies held by the professionals and the experiences of young women. It is argued that public health representatives need to accommodate their knowledge to the everyday life interests and needs of adolescent women. PMID- 10724470 TI - Managing menopause: a critical feminist engagement. AB - Feminist critiques of menopause have been beneficial in opening up important public health debates around menopause. One of the most contentious public health issues concerns the use of Hormone Replacement Therapy (HRT) for the prevention of osteoporosis, heart disease and, more recently, Alzheimer's disease, in postmenopausal women. For preventive purposes, it is recommended that women should take HRT for 10-15 years and preferably remain on the therapy for the remainder of their lives. This is despite reported increased cancer risks associated with HRT, side effects and considerable cost of the therapy. Various studies have shown that up to 50% of women stop taking HRT after 9-12 months. These figures are used in the medical literature as an indication of women's non compliance. Extending earlier feminist critiques around menopause and HRT, this paper discusses a critical feminist engagement around issues of women's perceived non-compliance with HRT. PMID- 10724471 TI - Are women disfavoured in the estimation of disability adjusted life years and the global burden of disease? AB - Addressing women's health goes beyond merely producing gender-disaggregated data. A women's perspective on health issues involves an analysis based on knowledge of broader gender differences than those that can be attributed to biology alone. Women and men live different lives. It is therefore important to evaluate whether women's health issues are disfavoured by known health burden estimators in general use. The Global Burden of Disease and Disability Adjusted Life Years framework was presented to the public health community by the World Bank in 1993. The data presented are applied for the year 1990. The estimators have been criticized for not being able to address social inequity within populations, and for failure to predict how serious the HIV/AIDS epidemic would be. Some of the critical voices have also centred around how the methodology will affect the way women's health priorities are incorporated; however, the debate is still in its infancy. This paper examines this new concept of measuring health burdens from a women's health perspective. PMID- 10724472 TI - What could a feminist perspective on power bring into public health? AB - The issues raised in this editorial indicate that public health research must embrace the advances made in the understanding of gender and other power dynamics, which influence the social distribution of health and illness among the population. Furthermore, it is crucial to recognize that social research (including that in health) is part of the social fabric--not separable from--the processes of power. Thus, a power perspective in public health can bring a more comprehensive and subtle understanding of the multiple and contradictory elements of gender and other relations of power that impact on the health status of populations. PMID- 10724473 TI - The social determinants of health: a contribution to the analysis of gender differences in health and illness. AB - While it is established that socioeconomic status and social integration influence the distribution of health and illness among men and women, little attention has been paid to the different ways in which women and men experience socioeconomic opportunities and social attachments to others. Drawing on evidence from the literature, the position developed in this article is that gender mediates the influence of both socioeconomic status and social integration on health, and for women, these are intricately linked. Women's relationship to the labour market establishes and perpetuates their socioeconomic inequality relative to men, and may produce contradictory influences on women's health. Furthermore, for women, the marital relationship is paradoxical: marriage may at once improve economic and social support opportunities, while diminishing control over paid and unpaid work--potentially increasing as well as compromising the health status of women. The article is intended to contribute to the growing body of literature on gender and the determinants of health. PMID- 10724474 TI - Anxiety, physical abuse, and low birth weight. AB - BACKGROUND: Physical and sexual abuse has been associated with adverse pregnancy outcome in some studies. One cause may be physical trauma; others may be indirect, such as stress, anxiety, smoking or drug use in pregnancy. The aim of the present study was to assess the relationships among anxiety, history of abuse and low birth weight. METHODS: We performed a case control study comprising 85 women who delivered low birth weight (<2500 g) babies (cases) and 92 women with higher birth weight babies (control group). All mothers were interviewed. We assessed the extent of abuse using the Conflict Tactics Scale, and that of anxiety using the Trait-Anxiety Inventory. RESULTS: Women with low birth weight babies were not more likely to have higher scores on the anxiety scale or to have a history of abuse. On the other hand, mothers with a history of abuse had higher anxiety scores and more often smoked in pregnancy. CONCLUSION: Anxiety could be the intermediate factor between abuse and smoking in pregnancy. PMID- 10724475 TI - Trends in tobacco use among Estonian and Russian youth in Tallinn. AB - Smoking behaviour and exposure to environmental tobacco smoke (ETS) were examined in three cross-sectional surveys from 1991/92, 1993/94, and 1995/96. The study population comprised 3,185 Estonian and Russian adolescents from 17 schools in Tallinn, Estonia. Prevalence of ever-smoking girls increased by 13 percentage points versus 2% among boys during the study period. Mean ages of the first experimentation with tobacco and exposure to ETS did not change significantly. Regular smoking increased significantly from 1991/92 to 1995/96. Detailed analyses for the 1995/96 survey showed that among ethnic Estonians, compared with ethnic Russians, the prevalence of ever-smokers and regular smoking were higher, mean age for the first experimentation was younger, and on average, Estonians smoked more cigarettes per week. The smoking trend among adolescents in Estonia is worsening; especially among Estonian youth. This study identifies a compelling need for national and community-wide efforts to deter adolescents from smoking and to reduce the exposure to ETS. PMID- 10724476 TI - Gender display in Scandinavian and American advertising for antidepressants. AB - This study examines whether depiction of users of antidepressants in advertisements for antidepressants in the 1995 issues of the major medical journal in each of Denmark, Finland, Norway, and Sweden differs from that in the American Journal of Psychiatry. The results show that the people shown in the Danish, Finnish, and Norwegian journals are predominantly women, whereas depiction of users in the American and Swedish advertising is predominantly of couples. The portrayals in the 1995 advertising are of antidepressants as female gendered; a feature that was not seen in advertising for psychotropic drugs in the Nordic countries in the 1980s. PMID- 10724477 TI - Women's health: do common symptoms in women mirror general distress or specific disease entities? AB - The purpose of this study was to determine aspects of validity of a short self report inventory, based on physical and psychological symptoms that, according to other studies, are most prevalent in women. It was hypothesized that such symptoms form a single entity, which would be consistent with the view that they represent a proxy for general distress in women. METHOD: One hundred and one women were approached, all of whom filled in the "common symptoms in the general population of women" (CSGP) instrument and the multidimensional self-report inventory Symptom Check List 90 (SCL90) for external comparison. The results indicate that the CSGP scale has high internal consistency (alpha = 0.80). The strongest correlations were found between the CSGP scale and the SCL90 subscales assessing "somatization", "depression" and "anxiety". In the factor analysis, the items on the CSGP scale were widely dispersed among the factors represented by the main SCL90 subscales, and therefore could not be considered to represent any particular subscale. Our conclusion is that the CSGP scale seems to assess a single entity. Other researchers suggest that such an entity represent a proxy for general distress in women. This will have implications for the interpretation of the common symptoms represented by the scale, which women report in clinical settings. PMID- 10724478 TI - Antigenic variation of Borrelia turicatae Vsp surface lipoproteins occurs in vitro and generates novel serotypes. AB - As a means of avoiding the host immune response, the tick-borne relapsing fever spirochete Borrelia turicatae undergoes antigenic variation in its abundant surface lipoproteins. In this study, B. turicatae strain Oz1, serotype B, was subcultured in vitro and cloned by limited dilutions after 50 passages. Four different serotypes (serotypes A, B, E, and F) differing by their expressed Vsp lipoproteins were isolated. Using pulsed-field gel electrophoresis, we showed that the variability in surface-exposed proteins is correlated with rearrangement between different linear plasmids, defining serotype-specific plasmid profiles. Moreover, we determined the nucleotide sequence of genes encoding the VspE and VspF lipoproteins, corresponding to the two novel serotypes E and F, respectively. Our results showed that antigenic variation in B. turicatae occurs spontaneously in vitro, in the absence of immune selection. PMID- 10724479 TI - Transposon mutagenesis in Halomonas eurihalina. AB - We have established a transposon mutagenesis procedure for the moderate halophile Halomonas eurihalina, a bacteria that produces an exopolysaccharide (EPS) of considerable biotechnological interest. We used suicide plasmids pUT and pSUP102 to introduce the transposons mini-Tn5 and Tn1732 into H. eurihalina via Escherichia coli mediated conjugation. Southern hybridization analysis demonstrated that insertions of the transposon mini-Tn5 into H. eurihalina occurred randomly at single sites in the chromosome, whereas Tn1732 insertion also took place at random, but simultaneously, at several sites. Phenotypic analysis revealed that different mutants were generated by using mini-Tn5. The isolation of exopolysaccharide-defective strains is the first stage towards carrying out genetic studies on EPS production by this microorganism. PMID- 10724480 TI - Molecular analysis of UAS(E), a cis element containing stress response elements responsible for ethanol induction of the KlADH4 gene of Kluyveromyces lactis. AB - KlADH4 is a gene of Kluyveromyces lactis encoding a mitochondrial alcohol dehydrogenase activity, which is specifically induced by ethanol and insensitive to glucose repression. In this work, we report the molecular analysis of UAS(E), an element of the KlADH4 promoter which is essential for the induction of KlADH4 in the presence of ethanol. UAS(E) contains five stress response elements (STREs), which have been found in many genes of Saccharomyces cerevisiae involved in the response of cells to conditions of stress. Whereas KlADH4 is not responsive to stress conditions, the STREs present in UAS(E) seem to play a key role in the induction of the gene by ethanol, a situation that has not been observed in the related yeast S. cerevisiae. Gel retardation experiments showed that STREs in the KlADH4 promoter can bind factor(s) under non-inducing conditions. Moreover, we observed that the RAP1 binding site present in UAS(E) binds KlRap1p. PMID- 10724481 TI - Characterization of group A streptococcal strains Sv and Su: determination of emm gene typing and presence of small vir regulon. AB - The emm gene typing of GAS (group A streptococcus) strains Sv and Su and the molecular structure of the vir regulon were decided. An emm(-like) gene from the chromosomal DNA of GAS strain Sv was amplified with forward and reverse primers, which were selected from the best conserved portion in leader sequences of different strains and the C-terminal conserved portion, respectively, for determination of the M protein gene type. Strain Sv was defined as serotype M23, because deduced N-terminal amino acid positions of the products are identical to those of the M type 23 (emml) gene derived from GAS strain M23-MEMPHIS (M serotype 23, GenBank accession number U11953). When the vir regulon of strain Sv was examined by polymerase chain reaction mapping and compared with that of GAS strain Su, they had a similar size in length. In addition, when sequencing analysis of the DNA fragment of 4791 base pairs (bp) encoding three open reading frames (orf, mga, and emm) and the upstream region of scpA from genomic DNAs of both strains was performed, the sequence of the DNA from strain Sv was, except for 1 bp (T for C at position 4124), identical to that of the DNA from strain Su. These data show that both strains possess the genes in the order of mga (virR or mry) -emm -scpA designated as the small vir regulon. The effect of the formation of alternative pathway C3 convertase of complement on the GAS strains Sv and Su was also examined. When GAS strains Sv and Su were incubated in NHS containing radiolabeled C3 in the presence of Mg-EGTA, binding of C3 to Su bacteria was dose dependent, whereas less binding of C3 to Sv bacteria was seen. Taken together, the data suggest that M protein could be expressed on the surface of the Sv bacteria, but not on the Su bacteria. PMID- 10724482 TI - Sequence variation of the 16S to 23S rRNA spacer region in Salmonella enterica. AB - The possibility for identification of Salmonella enterica serotypes by sequence analysis of the 16S to 23S rRNA internal transcribed spacer was investigated by direct sequencing of polymerase chain reaction-amplified DNA from all operons simultaneously in a collection of 25 strains of 18 different serotypes of S. enterica, and by sequencing individual cloned operons from a single strain. It was only possible to determine the first 117 bases upstream from the 23S rRNA gene by direct sequencing because of variation between the rrn operons. Comparison of sequences from this region allowed separation of only 15 out of the 18 serotypes investigated and was not specific even at the subspecies level of S. enterica. To determine the differences between internal transcribed spacers in more detail, the individual rrn operons of strain JEO 197, serotype IV 43:z4,z23: , were cloned and sequenced. The strain contained four short internal transcribed spacer fragments of 382-384 bases in length, which were 98.4-99.7% similar to each other and three long fragments of 505 bases with 98.0-99.8% similarity. The study demonstrated a higher degree of interbacterial variation than intrabacterial variation between operons for serotypes of S. enterica. PMID- 10724483 TI - At acidic pH, the diminished hypoxic expression of the SRP1/TIR1 yeast gene depends on the GPA2-cAMP and HOG pathways. AB - The hypoxic SRP1/TIR1 gene encodes a stress-response cell wall mannoprotein, which is shown to be necessary for yeast growth at acidic pH in the presence of sodium dodecyl sulfate. However, the hypoxic expression of SRP1 is shown to be downregulated at acidic pH. The stress-responsive HOG pathway appeared necessary to maintain hypoxic SRP1 expression, but only at acidic pH. However, unlike known HOG pathway-dependent genes, SRP1 was under positive cAMP control and was positively modulated by protein kinase A at neutral and acidic pH. In addition, the HOG-independent hypoxic HEM13 gene was also positively regulated by cAMP levels. Therefore, the positive cAMP control of the hypoxic SRP1 and HEM13 genes was uncoupled from the HOG pathway. Surprisingly, this positive cAMP control was found to be mediated by GPA2 but not by RAS2, so the Gpa2p requirement appears critical at acidic pH. Although RAS2 is not involved in the regulation of SRP1 expression, the guanine nucleotide exchange factor Cdc25, which is known to control the GTP/GDP ratio on the Ras proteins, was nevertheless required for hypoxic SRP1 expression. Furthermore, the Ras proteins did not compensate for Gpa2p requirement in a delta gpa2 mutated strain. These results suggest that the Cdc25 factor might also control Gpa2p. PMID- 10724484 TI - DNA heterogeneity of Staphylococcus aureus strains evaluated by SmaI and SgrAI pulsed-field gel electrophoresis in patients with impetigo. AB - To our knowledge, no studies have previously been carried out on the heterogeneity and intrafamily colonization of impetigo Staphylococcus aureus strains obtained by powerful discriminating methods such as pulsed-field gel electrophoresis (PFGE). To explore this topic, macrorestriction patterns of S. aureus strains were analyzed after SmaI and SgrAI digestion. The two enzymes provided superimposable results. A total of ninety-seven S. aureus strains was found in the 26 families whose lesions and nasal and pharyngeal samples were examined. There were 39 strains which were different by PFGE, and of these, 24 were found in the lesions. Although 85% of impetigo patients showed nasal colonization and 58% showed pharyngeal colonization, only 54% of the patients had the same PFGE strain in the lesion and in the nose, and 35% in the lesion and the pharynx. In half of the 26 families, at least one member (mother, father, or relative) presented a S. aureus strain identical, by PFGE, to strains isolated in patients' lesions. Nineteen percent of mothers, 15% of fathers, and 19% of the other relatives presented nasal colonization with strains identical to those isolated in the children's lesions. Lesional strains showed higher antimicrobial resistance than nonlesional isolates. PMID- 10724485 TI - Supplement 1998 (no. 42) to the Kauffmann-White scheme. AB - This supplement reports the characterization of 14 new Salmonella serovars recognized in 1998 by the WHO Collaborating Centre for Reference and Research on Salmonella: 11 were assigned to S. enterica subsp. enterica, one to subspecies salamae, one to subspecies diarizonae, and one to subsp. indica. In addition, the antigenic factor H:z88 is described. PMID- 10724486 TI - Pathophysiology of shock in trauma. PMID- 10724487 TI - Liver function assessed by increased rate of portal venous blood flow after oral intake of glucose. AB - OBJECTIVE: To find out whether an increased rate of portal venous blood flow after oral intake of glucose could be used to estimate liver function. DESIGN: Prospective study. SETTING: University hospital, Japan. SUBJECTS: Sixty patients, of whom 23 had hepatocellular carcinoma and liver cirrhosis, 21 had tumours metastatic to normal liver, and 16 had obstructive jaundice treated with percutaneous transhepatic biliary drainage (PTBD). INTERVENTION: Portal flow was measured after oral intake of glucose 75 g using pulsed-Doppler ultrasonography. RESULTS: The ratio of portal flow 30 minutes after glucose intake to that before intake (PVFR30) was significantly lower in cirrhotic patients than in those with metastases and a normal liver. A PVFR30 of less than 1.5 indicated impaired hepatic function assessed by the Child-Pugh scores, indocyanine green clearance test, prothrombin time, and hepaplastin test. It also indicated less reduction in total bilirubin concentrations in the first week after PTBD. CONCLUSIONS: Results suggest that PVFR30 can be used to estimate liver function and predict outcome after PTBD. PMID- 10724488 TI - Twelfth rib resection: a direct posterior surgical approach for subphrenic abscesses. AB - OBJECTIVE: To assess the results of twelfth rib resection as a direct posterior surgical approach to subphrenic abscesses in case of failure of percutaneous drainage, abandonment of percutaneous drainage in view of a too high risk of perforation of adjacent organs, or contamination of the pleural space, or an inaccessible abdomen. DESIGN: Retrospective study. SETTING: University hospital, The Netherlands. PATIENTS: 17 patients who required rib resection for subphrenic abscesses that developed after infected necrotising pancreatitis, splenectomy, or anastomotic disruption. INTERVENTIONS: 18 rib resections. MAIN OUTCOME MEASURES: Outcome and morbidity. RESULTS: Twelfth rib resection was successfully in 13 of 17 patients. Four patients died from multiple organ failure despite subsequent (re) laparotomies for additional surgical drainage. CONCLUSION: Twelfth rib resection can be useful for the treatment of subphrenic abscesses in selected patients. PMID- 10724489 TI - Prognostic factors for failure of primary patency within a year of bypass to the foot in patients with diabetes and critical ischaemia. AB - OBJECTIVE: To find out whether we could identify prognostic factors for early failure of bypass to the foot in diabetic patients with critical ischaemia. DESIGN: Retrospective series of consecutive patients. SETTING: County hospital, Sweden. PATIENTS: 43 diabetic patients who had 48 reconstructions for critical ischaemia between 1988 and 1994. INTERVENTIONS: 48 elective vein bypass procedures to the feet. MAIN OUTCOME MEASURES: Prognostic factors for primary patency. RESULTS: Primary and secondary patency rates at one year were 72% (95% confidence interval (CI) 58 to 85) and 83% (95% CI 71 to 95), respectively. Limb salvage and survival rates at one year were 85% (95% CI 74 to 96) and 86% (95% CI 75 to 96), respectively. Vein graft of questionable quality, major wound healing problems, use of the reversed vein technique, and a narrow lumen (< 1.5 mm) of the recipient artery increased the hazard for failed primary patency by 17.3 (p = 0.003), 6.0 (p = 0.02), 4.7 (p = 0.03), and 3.9 (p = 0.05) times, respectively. Short vein bypass (p = 0.70), translocated or composite veins (p = 0.61), major postoperative oedema of the leg (p = 0.46), or questionable quality of the wall of the recipient artery (p = 0.29), however, had no significant independent effect on the primary patency rate. CONCLUSION: Early primary patency after bypass to the foot in diabetic patients might improve if veins of questionable quality, major wound healing problems, thin reversed veins from the calf, and narrow recipient arteries can be avoided or handled more proficiently than in the present study. PMID- 10724490 TI - Standard preoperative assessment can improve outcome after cholecystectomy. AB - OBJECTIVE: To assess the outcome of cholecystectomy after standard preoperative handling and selection of patients, focusing on the potential of the operation to eliminate biliary colic. DESIGN: Prospective study. SETTING: University Hospital, Norway. PATIENTS: 806 patients (median age 56, range 18-91 years, male:female ratio 1:2.7), were referred to our clinic for cholecystectomy between 1992 and 1996. INTERVENTIONS: Unless there was a clear indication for cholecystectomy (frequent attacks of biliary colic/or recent complications of gallstones or both), patients were investigated in a standard way to find out what else was causing the abdominal pain. MAIN OUTCOME MEASURES: Residual pain was assessed at a clinical examination three months postoperatively, and clinical condition a median of three years later was assessed by a questionnaire. RESULTS: 465 (58%) patients were operated on primarily, and an additional 29 patients were operated on after further evaluation. Three months after cholecystectomy, 35 (7%) had persistent pain, mostly caused by other specific diseases and relieved after specific treatment. A median 3 years postoperatively, only 21 (4%) reported that they still had abdominal pain. CONCLUSION: Standard selection of patient improved the outcome of cholecystectomy. Compared with a historical control group, residual pain after three months was reduced from 20% to 7%. After three years, 96% of the patients no longer had their main clinical problem. PMID- 10724491 TI - Laparoscopic cholecystectomy for acute cholecystitis: how do fever and leucocytosis relate to conversion and complications? AB - OBJECTIVE: To find out whether fever and raised white cell count (WCC) are associated with conversion and complications of laparoscopic cholecystectomy in acute cholecystitis, and whether their presence could help in deciding the place of laparoscopic procedures. DESIGN: Prospective study. SETTING: Teaching hospital, Israel. SUBJECTS: 256 patients who were treated for clinical acute cholecystitis between January 1994 and November 1997. INTERVENTIONS: Emergency laparoscopic cholecystectomy. MAIN OUTCOME MEASURES: Raised temperature and WCC; incidence of conversion and complications. RESULTS: Raised temperature (>38 degrees C) was independently associated with advanced cholecystitis (p = 0.002, odds ratio [OR] 2.7) and a palpable gallbladder preoperatively (p = 0.02, OR 2.1). Total complications correlated with a temperature of >38 degrees C. Raised WCC (>15 x 10(9)/L) was independently associated with age >45 years (p = 0.02, OR 2.4), a palpable gallbladder preoperatively (p = 0.001, OR 2.9), and a raised temperature (>38 degrees C) (p < 0.0001, OR 6.2). Conversion was associated with a WCC >18 x 10(9)/L (p = 0.0, OR 3.2). CONCLUSION: A WCC of >18 x 10(9)/L may assist in predicting conversion, and fever of >38 degrees C may assist in predicting the development of complications. PMID- 10724492 TI - Mucinous cystic neoplasm of the pancreas or intraductal papillary-mucinous tumour of the pancreas. AB - OBJECTIVE: To compare clinicopathological findings in patients with mucinous cystic neoplasms and intraductal papillary-mucinous tumours. DESIGN: Retrospective study. SETTING: University department of surgery, Japan. SUBJECTS: 21 patients with mucinous cystic neoplasms (group 1) and 48 with intraductal papillary-mucinous tumours (group 2). RESULTS: The mean age was younger in group 1 (53(3.4) years) than in group 2 (65(1) years, p < 0.0001). The male:female ratio was smaller in group 1 than in group 2, being 0.17 (3/18) and 1.4 (28/20), respectively, (p = 0.0007). The main sites of the lesions were also significantly different: in group 1 four (19%) were located in the head and 17 in the body or tail, while 32 (67%) were in the head of the pancreas and 16 (33%) in the body or tail in group 2 (p = 0.0007). A unique endoscopic finding, excretion of mucin from the patulous orifice of the papilla, was present in two (9%) of the 21 mucinous cystic tumours and in 21 (45%) of the 47 intraductal papillary-mucinous tumours examined (p = 0.006). Metachronous or synchronous malignant diseases were found in the pancreas or other organs in one (5%) of the 21 patients with mucinous cystic neoplasm and in 13 (27%) of the 48 with intraductal papillary mucinous tumours (p = 0.03). The three- and five-year survival rates of 11 patients with mucinous cystadenocarcinoma were 45% and 27%, while those of 15 with intraductal papillary-mucinous carcinoma were 85% and 42%. CONCLUSIONS: These findings suggest that mucinous cystic neoplasm and intraductal papillary mucinous tumours are different clinicopathological entities. Aggressive surgery with peripancreatic lymph node dissection is recommended, particularly for mucinous cystadenocarcinoma, and postoperative follow-up with attention given to the presence of other malignancy is necessary as well as to local recurrence and haematogenous spread. PMID- 10724493 TI - Risk factors of mortality in perforated peptic ulcer. AB - OBJECTIVE: To assess the risk factors that influence mortality from perforated peptic ulcer. DESIGN: Retrospective study. SETTING: General hospital, Taiwan. SUBJECTS: 179 patients who had their perforated peptic ulcers operated on and who had minimum follow-up of one year. MAIN OUTCOME MEASURES: Mortality. RESULTS: The overall mortality was 15% (26/179). Of the 26 patients who died, the cause of death was uncontrolled systemic infection in 21 (81%), hypovolaemic shock in 2, and fatal arrhythmia and heart failure in 1 each. 15 of the patients who died of sepsis did not have fulminant abdominal sepsis. Most deaths occurred early after operation, (range 1-96 days). Old age, preoperative shock, and type of operation seemed to be related to these deaths on univariate analysis, but multivariate analysis showed that coexisting medical illness, delayed treatment, and low albumin concentration were independent risk factors for mortality. CONCLUSIONS: To improve the result of treatment of perforated peptic ulcer, the diagnosis and treatment should not be delayed, the associated medical illnesses should be treated, and nutritional support should be given. PMID- 10724494 TI - Audit and recurrence rates after hernia surgery. AB - OBJECTIVE: To study the effect of quality assurance on the recurrence rate after hernia repair. DESIGN: A prospective longitudinal cohort study. SETTING: District hospital, Sweden. SUBJECTS: All (n = 1232) patients aged 15-80 years operated upon for inguinal or femoral hernia in Motala 1984, 1986-1988, 1990, and 1992 1994. INTERVENTION: A questionnaire enquiring about pain or a lump in the operated area was sent 3-6 years postoperatively to all patients, excluding those who had already been operated on for recurrence and those who had died. Selected cases were examined depending on the answers to the questionnaire. MAIN OUTCOME MEASURES: Recurrence rate estimated by adding already confirmed recurrences to those found at the clinical examination; reoperation for recurrence; hospital stay; and number of day cases. Cumulative incidence of reoperation was analysed by actuarial analysis of all patients operated on from 1986-1997. RESULTS: The recurrence rate decreased from 18% in 1984 and 1986 to 3% in 1993 and 1994. The reoperation rate for recurrence at three years was 10.8% (95% confidence interval, CI: 9.3 to 12.2%), 3.6% (2.6 to 4.4%) and 2.2% (1.7 to 2.7%) for patients operated on between 1986-1988, 1989-1991 and 1992-1997, respectively. Differences between the first and the second and between the first and the third period were both highly significant (p < 0.001) whereas the difference between the second and third period was not (p = 0.09). Mean hospital stay decreased from 3.5 days in 1984 to 0.9 days in 1994. CONCLUSION: By recording recurrence rate or its surrogate endpoint, reoperation rate for recurrence, or both, hospital stay, and number of day cases, and presenting these results to participating surgeons, we provided incentives to improve outcome. This has resulted in a rapid decrease in recurrence rate and a shortened hospital stay, thereby improving cost effectiveness. PMID- 10724495 TI - Concentrations of leucocyte-associated chemotaxin C5a correlate with the mobilisation of polymorphonuclear leucocytes during operations for rectal cancer. AB - OBJECTIVE: To describe the profile of potential chemotaxins C5a, interleukin 8 and tumour-necrosis factor alpha, and the peripheral blood leucocyte (PBL) response in five patients having uncomplicated operations for rectal cancer. DESIGN: Prospective study. SETTING: University hospital, Norway. SUBJECTS: Five patients, four men and one woman, median age 66 years (range 48-77) who were operated on for rectal cancer. INTERVENTION: Four had low anterior resections and total mesorectal excision (TME) and one patient had a diverting end sigmoidostomy because of local perirectal spread of the cancer and liver metastases. Blood samples were taken at the start of the operation; peroperatively after 1, 2, and 3 hours; postoperatively at 5, 8, and 24 hours, for analysis of potential chemotaxins. RESULTS: The number of PBL tripled between the start and end of the operation and declined to a slightly lower plateau between 5 and 24 hours. Peroperatively, the association of C5a with PBL increased six-fold resulting in a doubled concentration of C5a/PBL, whereas the corresponding concentration of C5a in plasma remained relatively constant. Postoperatively, the concentration of C5a associated with the PBL and in plasma fluctuated with the maximums being at 8 and 24 hours, respectively. In contrast to C5a in plasma, the concentration of cell associated C5a correlated with number of PBL or polymorphonuclear leucocytes (PMN). The plasma concentration of IL-8 doubled during the operation, reached a six-fold maximum at 5 hours, and declined after 24 hours to twice the initial concentration. There were no correlations between plasma IL-8 and either number of PBL, plasma C5a, or PBL-associated C5a. No TNF alpha was detected in the plasma. CONCLUSION: Tissue trauma caused by uncomplicated excision of rectal cancer leads to leucocyte mobilisation peroperatively, probably partly by complement activation and subsequent generation and binding of the chemotaxin C5a to PBL. However, other chemotaxins may also play a role. PMID- 10724496 TI - Synthetic retinoids improve survival in rodent model of endotoxic shock. AB - OBJECTIVE: To investigate the effect of synthetic retinoids on septic shock induced by lipopolysaccharide (LPS) in rats. DESIGN: Randomised study. SETTINGS: University hospital laboratory, Sweden. ANIMALS AND INTERVENTIONS: 31 male Sprague Dawley rats randomised into four groups: controls, given vehicle alone (n = 6), LPS 6 mg/kg body weight alone (n = 12), and LPS 6 mg/kg but pretreated with the retinoic acid receptor-alpha (RAR-alpha) agonists CD336 (n = 6) and CD2081 (n = 7). MAIN OUTCOME MEASURES: Arterial blood pressure and heart rate measured hourly for four hours; mortality. RESULTS: LPS caused a pronounced fall in blood pressure within one hour of injection in all groups of rats. Of the 12 rats given LPS but not RAR-alpha agonists, 6 died before the end of the experiment. By contrast, all animal given either CD336 or CD2081 survived. The significantly improved survival was found despite no significant improvements in either mean arterial pressure or heart rate. CONCLUSION: Pretreatment with selective synthetic RAR-alpha agonists improves survival after LPS-induced septic shock in rats. These agents may have therapeutic potential in the treatment of septic shock in humans. PMID- 10724497 TI - Influence of portal hypertension on secretion of gastric mucus in rats. AB - OBJECTIVE: To assess the effect of portal hypertension on gastric adherent and soluble mucus in rats. DESIGN: Experimental study. SETTING: Teaching hospital, Taiwan. MATERIAL: 30 male Wistar rats. INTERVENTIONS: Portal hypertension was induced experimentally by partial ligation of the portal vein in 20 male Wistar rats: ten rats were examined after 4 weeks and the remaining 10 after 8. Another group of 10 rats (controls) had sham operations. MAIN OUTCOME MEASURES: Portal pressure, the severity of gross gastric mucosal lesions, and measurement of gastric adherent and soluble mucus. RESULTS: The portal pressure and the gross mucosal damage differed significantly between the experimental and the control groups (p < 0.01). There was significantly less gastric adherent mucus in the two experimental groups than in the control group (p = 0.002 and <0.001, respectively), whereas there was no significant differences in the amount of gastric soluble mucus (p = 0.5 and 0.1, respectively). The reduction in the gastric adherent mucus was closely related to the increase in portal pressure (p < 0.001) and the severity of portal hypertension-induced gastropathy (p < 0.001). CONCLUSIONS: Gastric adherent mucus may have an important role in the pathogenesis of portal hypertensive gastropathy, and its protective capacity is reduced by portal hypertension, as indicated by the decrease in gastric adherent mucus. PMID- 10724498 TI - Pancreatic glucagonoma: detection by positron emission tomography. PMID- 10724499 TI - Occult squamous cell carcinoma of the axillary tail of breast presenting as isolated axillary lymph node mass. PMID- 10724500 TI - Non-traumatic aneurysm of the extracranial vertebral artery. PMID- 10724501 TI - Bouveret's syndrome followed by gallstone entrapment in the stomach: an uncommon cause of upper gastrointestinal bleeding and gastric retention. PMID- 10724502 TI - Acute gastrointestinal bleeding from caecal varicosities. PMID- 10724503 TI - Inflammation and neovascularization associated with clinically used vascular prosthetic materials. AB - This study was designed to evaluate and compare healing characteristics, specifically neovascularization and inflammation, of polymeric vascular graft materials commonly used in clinical applications. Our hypotheses were (i) polymeric materials used in vascular graft manufacture stimulate chronic inflammation and (ii) inflammation and neovascularization of polymeric materials are related. Impra and Gore-Tex ePTFE, Meadox weavenit and woven Dacron, Hemashield microvel and woven Dacron, and Golaski microknit Dacron were implanted as 6-mm diameter disks within rat subcutaneous and adipose tissue. Following 5 weeks of implantation samples were evaluated by histological and immunocytochemical analysis. Sections were stained using hematoxylin and eosin or reacted with ED1 antibody and GS1 lectin to quantify inflammation and neovascularization. respectively. The extent of inflammation and neovascularization were influenced by both tissue site of implantation and polymer characteristics. For subcutaneous implants, inflammation was graded as follows: Meadox weavenit > Hemashield woven > Meadox woven > Gore-Tex ePTFE > Hemashield microvel > ImpraePTFE > Golaski microknit, while only the Golaski microknit neovascularized. Inflammation was graded as follows for adipose implants: Hemashield woven > Hemashield microvel > Meadox weavenit > Meadox woven > Gore-Tex ePTFE > Golaski microknit > Imnpra ePTFE, while the following order of neovascularization was observed: Impra ePTFE > Gore-Tex ePTFE > Golaski microknit. The degree of inflammation following biomnaterial implantation has a profound effect on implant neovascularization. These data suggest an inverse relationship exists between inflammation and neovascularization. PMID- 10724504 TI - The degree of neointimal formation after stent placement in atherosclerotic rabbit iliac arteries is dependent on the underlying plaque. AB - The purpose of this study was to determine the effects of stent placement on the underlying arterial morphology and the relations of stent-vessel wall interactions with subsequent neointimal formation in an atherosclerotic artery. Seven New Zealand White rabbits with experimentally induced atherosclerosis underwent balloon angioplasty (n = 7) and stent placement after balloon angioplasty (n = 7) in the iliac arteries. Histologic analysis of the treated arteries was performed at 28 days to assess device interactions with the artery and the pattern of the neointimal response. The area within the external elastic lamina of the stented vessels was 66% greater than the arteries with balloon angioplasty alone (p = 0.001) which contributed to a significantly greater late lumen area (3.33 +/- 0.51 mm2 versus 1.33 +/- 0.20 mm2, p = 0.0028). Neointimal thickness was measured at 220 stent wire sites from 21 sections of stented arteries of which 139 (63%) had underlying plaque and 81 (37%) were adjacent to normal media. Rupture of the internal elastic lamina (IEL) occurred at only 9 (11%) of the 81 stent wire sites over normal media. The mean neointimal thickness was 0.16 +/- 0.01 mm lor all stent wire sites. The neointimal thickness was greater at the stent wire sites with underlying plaque (0.23 +/- 0.01 min) than at the stent wire sites adjacent to normal media (0.08 +/- 0.01 mm) or at sites with rupture of the internal elastic lamina (0.16 +/- 0.02 mm, p = 0.0001). The degree of neointimal formation within the stents strongly correlated with the area of the underlying atherosclerotic plaque (r = 0.76, p = 0.0007) and the extent of plaque or medial compression by the struts (r = 0.90, p = 0.006). The present study characterizes stent interactions in a model commonly employed to evaluate novel therapies for the prevention of restenosis. The neointimal response was influenced by the local arterial morphology and correlated with the extent of plaque or medial compression by the stent. These data may be useful for future studies in this model and understanding the mechanism of in-stent restenosis. PMID- 10724505 TI - Correlation between clinical and histologic patterns of degenerative mitral valve insufficiency: a histomorphometric study of 130 excised segments. AB - The objectives of this study were to examine quantitatively the histological changes in incompetent degenerative mitral valves obtained at surgery for mitral valve repair, and to determine whether Barlow's disease (BD) and fibroelastic deficiency (FED) can be distinguished by histology. The billowing mitral leaflet syndrome (or Barlow's disease) and FED can be distinguished on the basis of clinical patterns and gross features, but their histologic patterns have not been described. One hundred thirty patients were studied. Thirty-nine (24 males) had BD; 44 (38 males) FED; 15 (7 males) Marfan's syndrome (MS); and 32 patients (25 males) a non-determined degenerative disease. Histological changes of the resected segment of the valve were quantitatively evaluated using scores of severity. A discriminant analysis was performed. The groups defined by the computer were checked for concordance with groups defined by the surgeon. Collagen alterations were found the most severe in MS patients. BD and MS had the most myxoid infiltration. MS and FED patients had the most elastic fiber alterations. No BD in males and only one in females were misclassified by the discriminant procedure into the FED group. Overall, the percentages of correct matchings were 54% in males and 62% in females. When the age of patients and the size of ring were added to histology to determine whether this additional information provided more discrimination, the percentages of correct matchings reached 90% in males and 100% in females. BD and FED are two fairly distinct entities, which can be distinguished by quantitative histology, whereas only modest differences were found in qualitative histology. PMID- 10724506 TI - Secondary heterotypic versus homotypic infection by Coxsackie B group viruses: impact on early and late histopathological lesions and virus genome prominence. AB - The impact of prior exposure to a different or identical strain of Coxsackievirus B (CVB) on murine CVB myocarditis was studied using a susceptible murine host (A/J[H-2a]) and myocarditic CVB3 or avirulent CVB2 as primary or secondary infectants. The effects of secondary heterotypic infection (CVB2 followed by CVB3) and homotypic infection (CVB3 followed by CVB3) 28 days after primary inoculation, versus CVB2 or CVB3 alone, on injury and viral genomic replication, both early (day 7) and late (days 28 and 56), were evaluated. After the primary infection by CVB2, trivial viral RNA was present in the heart and other organs, and a substantial positivity was observed with CVB3 infection. Seven days after secondary heterotypic (CVB2-CVB3) infection, the quantity of CVB genome in heart, pancreas, liver, and spleen was increased compared with the virus genome in the CVB3-CVB3 group and in the group with primary CVB3 infection alone. This phenomenon was seen in the heart and spleen up to day 28 postsecondary infection. Tissue inflammation and necrosis in heart and pancreas were prominent 7 days postsecondary infection with CVB2-CVB3 and correlated well with an increased quantity of CVB genome. Virus genome was present in heart and spleen 28 days after CVB3 infection alone. Serum CVB3 neutralization titer was increased to 1:128 in CVB2-CVB3 group at days 7 and 28 postsecondary infection, and serum completely neutralized cytopathological effects of CVB3 in the CVB3-CVB3 group at day 7 and 28 postsecondary infection. Our results indicate that secondary heterotypic infection by CVB causes increased injury, inflammation, and CVB replication in target organs such as the heart and pancreas, as well as in immune compartments like the spleen. Compared with CVB3 alone, the intense inflammatory infiltriate in the CVB2-CVB3 group is as not due solely to postviral sensitization of the immune system, but rather to the inability of the host to eradicate the virus. PMID- 10724507 TI - The anatomy of a collection. AB - An old collection of 81 congenitally malformed hearts was examined in detail using the system of sequential segmental analysis. The specimens were almost entirely from infants and young children who had died during the two decades between 1954 and 1973. There had been surgical intervention in 26 cases, and most of these patients died during or shortly after the operation. The various anomalies are listed, and the pattern of prevalence is contrasted to that of present-day collections. Among the many interesting specimens there was a fine example of one of the rarest of cardiac anomalies, a criss-cross arrangement of atrioventricular connections. The collection, as an entity, gives an interesting historical insight of the state of congenital heart disease during the early years of the development of cardiac surgery. PMID- 10724508 TI - Pathology of the heart and conduction system in a case of sudden death due to a cardiac fibroma in a 6-month-old child. AB - A 6-month-old female infant considered to be in good health died suddenly and unexpectedly. Post-mortem examination was requested, with clinical diagnosis of sudden infant death syndrome. Gross examination revealed, however, the presence of a cardiac mass 4.5 X 4 x 3.5 cm in diameter. Histological examination of the heart confirmed the presence of a cardiac fibroma. In the present case, the sudden death could have been due to the left ventricular outflow obstruction, as much as to conductive disturbances caused by overstretching and compression of the atrioventricular node and of the bundle branches. Hemodynamic and conductive abnormalities are presumed to have provoked bradycardia degenerating into ventricular fibrillation and sudden death. Necroscopy studies of sudden death should always include histological examination of the cardiac conduction system but seldom do. PMID- 10724509 TI - Minute (<1 mm) occult cardiac myxoma in an adult. PMID- 10724510 TI - Cardiovascular research in The Animal Medical Center. PMID- 10724511 TI - What is a protist? PMID- 10724512 TI - Meeting report: Molecular Parasitology, Woods Hole, USA, September 13-17, 1998. PMID- 10724513 TI - Meeting report: aquatic flow cytometry: achievements and prospects, Research- and Technology Centre Westcoast (FTZ), Busum, Germany, October 15-16, 1998. PMID- 10724514 TI - Paramecium molecular genetics: functional complementation and homology-dependent gene inactivation. PMID- 10724515 TI - Organization and structure of the Giardia genome. PMID- 10724516 TI - Diatoms and the ocean carbon cycle. PMID- 10724517 TI - Phylogenetic affinities of Diplonema within the Euglenozoa as inferred from the SSU rRNA gene and partial COI protein sequences. AB - In order to shed light on the phylogenetic position of diplonemids within the phylum Euglenozoa, we have sequenced small subunit rRNA (SSU rRNA) genes from Diplonema (syn. Isonema) papillatum and Diplonema sp. We have also analyzed a partial sequence of the mitochondrial gene for cytochrome c oxidase subunit I from D. papillatum. With both markers, the maximum likelihood method favored a closer grouping of diplonemids with kinetoplastids, while the parsimony and distance suggested a closer relationship of diplonemids with euglenoids. In each case, the differences between the best tree and the alternative trees were small. The frequency of codon usage in the partial D. papillatum COI was different from both related groups; however, as is the case in kinetoplastids but not in Euglena, both the non-canonical UGA codon and the canonical UGG codon were used to encode tryptophan in Diplonema. PMID- 10724518 TI - Systematics of the enigmatic kathablepharids, including EM characterization of the type species, Kathablepharis phoenikoston, and new observations on K. remigera comb.nov. AB - The colorless flagellate Kathablepharis has consisted of five species based on light microscopic studies, and the ultrastructure of the type species, Kathablepharis phoenikoston, is described for the first time. The heterotrophic, marine flagellate Leucocryptos consisted of two species, but additional ultrastructural details for one of these, Kathablepharis remigera comb. nov. (= Leucocryptos remigera Vors), indicates that it should be transferred to Kathablepharis. The cellular structure of these two species is similar to previously studied kathablepharids. However, there is variation in the feeding apparatus and cytoskeleton. The feeding apparatus of both species has a cytostome, a cytostomal ring, and cytopharyngeal rings. The cytoskeleton consists of inner microtubular arrays and outer or sub-pellicular microtubular arrays. In addition, several features of the flagellar apparatus are described for K. phoenikoston and K. remigera. The ultrastructure of these two species is compared with other kathablepharids to evaluate their taxonomy and phylogeny. We classify Kathablepharis and Leucocryptos in the family Kathablepharididae incertae sedis. PMID- 10724519 TI - Analysis of cDNA expressed sequence tags from Entamoeba histolytica: identification of two highly abundant polyadenylated transcripts with no overt open reading frames. AB - Upon analysis of 304 expressed sequence tags derived from the protozoan parasite Entamoeba histolytica, a number of novel protein encoding amoeba sequences were isolated. In addition, two unrelated, abundantly expressed transcripts were identified, and designated, ehapt1 and ehapt2. Although these transcripts do not contain any overt open reading frame, both are polyadenylated and together represent about 19% of total polyA+-RNA(11.6% for ehapt1 and 7.5% for ehapt2), thus being the most highly expressed polyA-containing transcripts so far identified in E. histolytica trophozoites. Northern blot and primer extension analyses revealed single-sized transcripts of 0.5 and 0.6 kb for ehapt1 and ehapt2, respectively, and Southern blot analysis suggests that both are encoded by multiple genes, which are distributed throughout the amoeba genome. Comparison between various ehapt1- and ehapt2-derived cDNAs indicated that both transcripts are highly polymorphic. Whereas nucleotide substitutions in ehapt2 are distributed throughout the sequence, variations in ehapt1 are mainly restricted to two regions, one of which comprises a deletion of variable length within an 8 nt tandem repeat unit. At present there is no convincing explanation for the possible role of ehapt1 and ehapt2 in E. histolytica, and analogous sequences have not been described so far for any other organism. Most likely they might represent regulatory RNAs or transcribed transposable elements. PMID- 10724520 TI - Phylogenetic analysis of the SSU rRNA from members of the Chrysophyceae. AB - The nucleotide sequence for the nuclear-encoded small subunit ribosomal RNA gene (SSU rRNA) was determined for 24 species of the Chrysophyceae sensu stricto. These sequences were aligned, using primary and secondary structure, with nine previously published sequences for the Chrysophyceae, 14 for the Synurophyceae, and five for the Eustigmatophyceae (outgroup). Data analyses were the substitution rate calibration distance method using neighbor-joining (TREECON), Kimura 2-parameter neighbor-joining method (PAUP) and the maximum parsimony method (PAUP, PHYLIP). Trees from the analyses were largely congruent, but bootstrap support was weak at many nodes. The analyses recovered clades of uniflagellate and biflagellate organisms associated with current higher level taxonomy (e.g., subclass, order). The genus Ochromonas was polyphyletic, and O. tuberculata in particular was distantly related to the other Ochromonas species in the analysis. The family Paraphysomonadaceae occupied a basal position in three of four analyses. The class Synurophyceae appeared to be embedded within the Chrysophyceae, but bootstrap support was weak (< 50%) in all analyses except the PHYLIP parsimony analysis (= 81%). It was considered premature to place the Synurophyceae back into the Chrysophyceae based upon the analysis of one gene, especially given the ultrastructural and pigment differences between the two groups, but the relationship of these two groups deserves further study. PMID- 10724521 TI - Annotated excerpts from Clifford Dobell's 88-year-old insightful classic paper, "The Principles of Protistology". PMID- 10724522 TI - Programmed cell death in the myocardium of arrhythmogenic right ventricular cardiomyopathy in children and adults. AB - Arrhythmogenic right ventricular cardiomyopathy (ARVC) is characterized by fibrofatty replacement of the right ventricular myocardium. Recently, the myocardial loss in ARVC has been suggested to be related to apoptosis. However, it is still unknown whether this phenomenon is already established in the myocardium of pediatric cases with this disease. We examined the histopathologic characteristics of the ventricular myocardium in specimens obtained from 10 patients, including 3 children with ARVC, and investigated the occurrence of apoptosis in the myocardium by terminal deoxyribonucleotidyl transferase-mediated dUTP-biotin nick-end labeling (TUNEL) assay and agarose-gel electrophoresis of DNA. Endomyocardial biopsy specimens from the 10 cases and a necropsy sample from one adult case with ARVC were examined. Histopathologic examination of biopsy specimens from the pediatric cases revealed extensive fibrosis. Typical fatty infiltration was demonstrated in one of the 3 pediatric cases. These findings were similar to those in adult cases; the histopathologic index based on the severity of myocardial damage, including myocyte degeneration and fibrosis, was not significantly different from that in adult cases. TUNEL assay revealed positive reactivity of the myocardial cells. The apoptotic index was 1.4 +/- 0.4% in children and 1.6 +/- 0.5% in adults (difference not statistically significant). Agarose-gel electrophoresis of a DNA extract of the myocardial tissue of the autopsy case revealed DNA fragmentation. Cases with idiopathic ventricular tachycardia and control cases with a cardiac transplant (with no rejection) had minimal histopathologic findings and negative reactivity in the TUNEL assay. These results indicate that myocardial damage is already established in cases diagnosed as ARVC in childhood, and suggest that the myocardial damage is closely related to apoptosis in children, as well as in adults, in this disease. PMID- 10724523 TI - Ultrastructure abnormalities in proteoglycans, collagen fibrils, and elastic fibers in normal and myxomatous mitral valve chordae tendineae. AB - Normal and myxomatous chordae tendineae were studied using light and electron microscopy, to assess the alterations in the appearance and mutual arrangement of proteoglycans, collagen fibrils, and elastic fibers. Specific staining with ruthenium red and cuprolinic blue in a critical electrolyte concentration mode were used to localize proteoglycans. Fresh tissues were fixed in glutaraldehyde containing the cationic dyes and embedded into Spurr resin. Semithin sections of LR White (London Resin Co., Basingstoke, U.K.)-embedded tissue were used for histochemistry. In normal chordae tendineae, the fibrosa comprised close-packed collagen fibrils intermixed with elastic fibers. These were surrounded by a thin layer of elastic fibers and collagen fibrils, both of which were closely associated with proteoglycans. In myxomatous chordae tendineae, alterations were observed in the connective tissue. Proteoglycans were more abundant and were distributed throughout the tissue. The outermost layer was transformed into an undifferentiated electron-dense mass surrounding the central fibrosa, which contained degraded elastic fibers and collagen fibrils. Collagen fibrils had faint banding or lacked a banding pattern altogether. Spaces between collagen fibrils were occupied by abnormal proteoglycans or proteoglycan aggregates. Elastic fibers showed varying degrees of degeneration and were occasionally replaced by electron-lucent spaces containing microfibrils. Accumulation of abnormal proteoglycan was also observed around degenerated elastic fibres and collagen fibrils. PMID- 10724524 TI - Immunolocalization of elastin, collagen type I and type III, fibronectin, and vitronectin in extracellular matrix components of normal and myxomatous mitral heart valve chordae tendineae. AB - The identification, distribution, and localization of matrix proteins and the proteins associated with normal and degenerated elastic fibers and collagen fibrils of myxomatous chordae tendineae were studied with immunoelectron microscopy. Ultrathin sections of L R White-embedded tissue were processed by indirect immunogold cytochemistry using primary antibodies against human alpha elastin, collagen types I and III, fibronectin, and vitronectin. In normal chordae tendineae, alpha elastin antibody heavily labeled the elastic fibers in spongiosa and fibrosa, but microfibrils around them were not labeled. Antibodies to collagen type I, collagen type III, and fibronectin all labeled the collagen fibers and microfibrils in the spongiosa. Fibronectin antibody labeling was higher than collagen type III, whereas labeling by anticollagen type I was lower. Intense labeling by vitronectin was observed on the microfibrils in the spongiosa and on electron-dense material around elastic fibers in the spongiosa and fibrosa. In myxomatous chordae tendineae, alpha elastin antibody heavily labeled degenerated elastic fibers, previously unidentified reticulated structures, and other moderately electron-dense material, both in the spongiosa and in the fibrosa, but not the electron-dense fibrous material around them. Antibodies to collagen types I, III, and fibronectin heavy labeled electron-dense aggregates of fibrous material. Vitronectin labeling was observed on electron-dense longitudinally running microfibrils and on the electron-dense microfibrils around degenerated elastic fibers. PMID- 10724525 TI - Acute aortic regurgitation due to spontaneous rupture of a fenestrated cusp: report in a 65-year-old man and review of seven additional cases. AB - A 65-year-old man with chronic hypertension developed dyspnea, a cough, and a new diastolic murmur. Two-dimensional echocardiography showed severe aortic regurgitation. No valvular vegetations were identified and blood cultures were negative. Surgical intervention was recommended, but the patient died of an acute intracranial hemorrhage two weeks later. At autopsy, the posterior aortic cusp was flail, due to rupture of the residual cord above two large fenestrations. There was no acute or healed endocarditis. To our knowledge, this is the eighth reported case of aortic valve incompetence due to spontaneous rupture of a fenestrated cusp. Patients ranged in age from 31-67 years (mean, 54), and 4 (50%) were older than 60 years. Seven (88%) of the 8 were men, and 4 (57%) of 7 had chronic hypertension. Analogously, in another four reported cases, aortic insufficiency developed following spontaneous rupture of the fenestrated raphe of an atypical congenitally bicuspid aortic valve. Noninfective and nontraumatic rupture of cord-like aortic valve structures may result in severe acute aortic regurgitation, particularly in men with chronic hypertension. PMID- 10724526 TI - Is age a contributory factor of mitochondrial bioenergetic decline and DNA defects in idiopathic dilated cardiomyopathy? AB - While mitochondrial abnormalities are increasingly recognized in cardiac diseases including hypertrophic cardiomyopathy, their presence in idiopathic dilated cardiomyopathy and the role that age plays in their incidence and severity have yet not been assessed. Levels of cardiac respiratory enzyme activities and mitochondrial DNA (mtDNA) were examined in 55 subjects with idiopathic dilated cardiomyopathy divided into 3 age groups. Respiratory enzyme activity levels were significantly lower in 37 patients (67%) compared to age-matched controls and increased activity levels were noted in 9 (16%). Decreased activities were found in complex I (n = 11), III (n = 16), IV (n = 12) and V (n = 13), but not in II, the only respiratory complex entirely nuclear-encoded. No age-specific differences were found in the overall frequency of enzymatic abnormalities. However, older patients had significantly increased multiple enzyme activity defects as well as increases in abundance and frequency of the 7.4 kb deletion. In addition, 3 patients were noted with marked reduction in mtDNA levels. None of the pathogenic mtDNA mutations previously associated with hypertrophic cardiomyopathy were found, nor was there any relationship that could be established between levels of specific mtDNA deletions and enzyme activities. In summary, specific mitochondrial abnormalities are heterogenous and frequent in both adults and children with idiopathic dilated cardiomyopathy. Older patients are more likely to have mtDNA deletions and multiple enzyme activity defects. The molecular basis for these abnormalities remains undefined. PMID- 10724527 TI - Immunohistochemical analysis of platelet-derived growth factor-B expression in myocardial tissues in hypertrophic cardiomyopathy. AB - Intimal and/or medial hyperplasia of intramyocardial small vessels is thought to be one of the causes of myocardial ischemia in hypertrophic cardiomyopathy (HCM). However, the pathogenesis of such vascular lesions in HCM is not yet known. To evaluate the pathogenic role of platelet-derived growth factor (PDGF-B) and basic fibroblast growth factor (b-FGF), which have a potential to induce cellular and molecular changes observed in the vessels in HCM, we examined the expression of these molecules and PDGF receptors in cardiac tissues from six patients with HCM and seven controls using immunohistochemistry. The percentage of PDGF-B positive cells in the myocyte population in HCM was significantly higher than that in controls (52.6 +/- 16.2 (mean +/- SD) vs. 21.6 +/- 9.6, p < 0.01). PDGF-B was also observed in vascular regions in HCM (61.1 +/- 25.5% of arterioles) but not in controls. There were no significant differences in the expression of b-FGF and PDGF receptors in the myocyte and non-myocyte populations and the vascular regions between the HCM and control groups. Our study revealed that the expression of PDGF-B protein was up-regulated in HCM, suggesting the contribution of this molecule to the development of intramyocardial vasculopathy. PMID- 10724528 TI - Congenital cystic tumors of the atrio-ventricular node: successful demonstration by an abbreviated dissection of the conduction system. AB - Pathologists are dissuaded from the pathological study of the conduction system by the high cost and complexity of a traditional complete study. However, a simplified, low-cost approach can produce concrete information when performed in carefully selected cases of atrioventricular block, as demonstrated in the two cases of congenital cystic tumor of the atrioventricular node described in this report. PMID- 10724529 TI - Studies on the origin of stereoselectivity in the synthesis of 1,2-trans glycofuranosyl azides. AB - The stereoselectivity of the 1,2-trans directed, Lewis acid-catalysed azidation of peracylated furanoses was found to depend on the reactivity of the azide donor (azide nucleophilicity) and the configuration at the anomeric centre relative to the neighbouring 2-O-acyl group. Reactions of 1,2-trans glycosyl esters with highly nucleophilic azide donors, generated from SnCl4 and Me3SiN3, were stereospecific. The results are interpreted in terms of the rapid reaction of the azide species with bicyclic 1,2-acyloxonium (1,2-O-alkyliumdiyl-D-glycofuranose) ions, which were the primarily formed reactive intermediates. When using 1,2-cis glycosyl esters as starting materials the selectivity was reduced (90-94% de); the same is true with 1,2-trans counterparts if less nucleophilic Me3SiN3 in combination with Me3SiOTf catalyst was used. This occurred due to the appearance of the more reactive but less selective oxocarbenium (glycofuranoxonium) ions either as primarily formed reactive intermediates in the former case or after equilibration with acyloxonium ions in the latter case. Protected 1,2-trans beta D-glycofuranosyl azides with ribo, xylo and 3-deoxy-erythro-pento configurations were best prepared from the corresponding glycosyl esters using 0.05 equivalents of SnCl4, i.e., under anomerization-free conditions. Azidation of methyl glycofuranosides proceeds with inferior (80-90% de) and less predictable selectivity irrespective of the starting anomeric configuration. PMID- 10724530 TI - Enzymatic synthesis of blood group A and B trisaccharide analogues. AB - Glycosyltransferases A and B utilize the donor substrates UDP-GalNAc and UDP-Gal, respectively, in the biosynthesis of the human blood group A and B trisaccharide antigens from the O(H)-acceptor substrates. These enzymes were cloned as synthetic genes and expressed in Escherichia coli, thereby generating large quantities of enzyme for donor specificity evaluations. The amino acid sequence of glycosyltransferase A only differs from glycosyltransferase B by four amino acids, and alteration of these four amino acid residues (Arg-176-->Gly, Gly-235- >Ser, Leu-266-->Met and Gly-268-->Ala) can change the donor substrate specificity from UDP-GalNAc to UDP-Gal. Crossovers in donor substrate specificity have been observed, i.e., the A transferase can utilize UDP-Gal and B transferase can utilize UDP-GalNAc donor substrates. We now report a unique donor specificity for each enzyme type. Only A transferase can utilize UDP-GlcNAc donor substrates synthesizing the blood group A trisaccharide analog alpha-D-Glcp-NAc-(1-->3) [alpha-L-Fucp-(1-->2)]-beta-D-Galp-O-(CH2 )7CH3 (4). Recombinant blood group B was shown to use UDP-Glc donor substrates synthesizing blood group B trisaccharide analog alpha-D-Glcp-(1-->3)-[alpha-L-Fucp-(1-->2)]-beta-D-Galp-O (CH2) 7CH3 (5). In addition, a true hybrid enzyme was constructed (Gly-235-->Ser, Leu-266-->Met) that could utilize both UDP-GlcNAc and UDP-Glc. Although the rate of transfer with UDP-GlcNAc by the A enzyme was 0.4% that of UDP-GalNAc and the rate of transfer with UDP-Glc by the B enzyme was 0.01% that of UDP-Gal, these cloned enzymes could be used for the enzymatic synthesis of blood group A and B trisaccharide analogs 4 and 5. PMID- 10724531 TI - Structural characterisation and enzymic modification of the exopolysaccharide produced by Lactococcus lactis subsp. cremoris B39. AB - Lactococcus lactis subsp. cremoris B39 grown on whey permeate produced an exopolysaccharide containing L-Rha, D-Gal and D-Glc in a molar ratio of 2:3:2. The polysaccharide was modified using an enzyme preparation from Aspergillus aculeatus, resulting in the release of Gal and a polymer with approximately the same hydrodynamic volume as the native polysaccharide. Linkage analysis and 1H NMR studies of both the native and modified exopolysaccharides elucidated that terminally linked Gal was released during modification and that the chemical structure of the branches within the repeating units is: beta-D-Galp-(1-->4)-beta D-Glcp-(1-->. 2D NMR experiments (both 1H-1H and 1H-13C) revealed that exopolysaccharide B39 consists of a branched heptasaccharide repeating unit with the following structure: [structure: see text]. PMID- 10724532 TI - Two diastereomeric saponins with cytotoxic activity from Albizia julibrissin. AB - Two diastereomeric saponins, julibrosides J1 (1) and J9 (2), both of which show cytotoxic activity, were obtained from the stem bark of Albizia julibrissin Durazz. On the basis of chemical and spectral evidence [L.B. Ma et al., Carbohydr. Res., 281 (1996) 35-46], the structure of 1 was revised as 3-O-[beta-D xylopyranosyl-(1-->2)-alpha-L-arabinopyranosyl-(1-->6) -beta-D-glucopyranosyl]-21 O-[(6S)-2-trans-2-hydroxymethyl-6-methyl-6-O- [4-O-((6R)-2-trans-2,6-dimethyl-6-O (beta-D-quinovopyranosyl)-2,7- octadienoyl)-beta-D-quinovopyranosyl]-2,7 octadienoyl] acacic acid-28-O-beta-D-glucopyranosyl-(1-->3)-[alpha-L arabinofuranosyl-(1-->4 )]-alpha-L-rhamnopyranosyl-(1-->2)-beta-D-glucopyranosyl ester. The diastereoisomer 2 of 1 was identified as 3-O-[beta-D-xylopyranosyl-(1- >2)-alpha-L-arabinopyranosyl-(1-->6) -beta-D-glucopyranosyl]-21-O-[(6S)-2-trans-2 hydroxymethyl-6-methyl-6-O- [4-O-((6S)-2-trans-2,6-dimethyl-6-O-(beta-D quinovopyranosyl)-2,7- octadienoyl)-beta-D-quinovopyranosyl]-2,7-octadienoyl] acacic acid-28-O-beta-D-glucopyranosyl-(1-->3)-[alpha-L-arabinofuranosyl-(1-->4 )]-alpha-L-rhamnopyranosyl-(1-->2)-beta-D-glucopyranosyl ester. Saponin 2 is a new saponin named julibroside J9. Both julibrosides J1 and J9 show good inhibitory action against the KB cancer cell line in vitro. PMID- 10724533 TI - Preparation of some starch-based neutral chelating agents. AB - Various neutral starch derivatives have been prepared by reacting maize starch with mono- and dimethylol resins based on urea, thiourea, and melamine. The factors affecting these reactions were studied. These factors include curing duration, catalyst, and resin concentrations. The starch derivatives so prepared were used in heavy-metal removal from solutions. The sorption ability of those derivatives depends on resin type and metal ions. The sorption values of different starch derivatives follow the order (a) monomethylol resin-starch > dimethylol resin starch; (b) thiourea resin-starch > urea resin-starch > melamine resin-starch. The sorption efficiency (%) of starch derivatives increases with increasing nitrogen content, reaching a maximum value and then decreasing. The sorption values of Hg2+ (mmol/mol resin) of different starch monomethylol derivatives at the maximum values were 1135, 2624, and 2538 for urea, thiourea, and melamine derivatives, respectively. This indicates that urea derivatives act as bidentate ligands, while thiourea and melamine act as tridentate ligands. PMID- 10724534 TI - A short, stereoselective synthesis of neo-inositol. AB - A practical synthesis of neo-inositol is described in which the target is prepared on a multigram scale in six operations from bromobenzene. PMID- 10724535 TI - Selective deprotection of 2',6'-di-O-benzyl-2,3:5,6:3',4'-tri-O isopropylidenelactose dimethyl acetal. AB - The reactivity order of O-deisopropylidenation of the three isopropylidene protecting groups of 2',6'-di-O-benzyl-2,3:5,6:3',4'-tri-O-isopropylidenelactose dimethyl acetal (2) with various reagents was established. The 5,6-acetal group was, although to a limited extent, more reactive as compared with the 3',4' group, while the 2,3-O-isopropylidene group was definitely less reactive. Conditions were determined for the direct preparation of the 5,6,3',4'-tetraol 5 (60% aqueous acetic acid, room temperature, 48 h, 73% yield) and the 5,6-diol 4 (propylene glycol and p-toluenesulphonic acid in dichloromethane, 46% yield). The diacetonated derivative 3, formally arising from a selective 3',4'-O deisopropylidenation, was obtained in high yield (90%) through a selective acetonation with 2-methoxypropene of the tetraol 5. PMID- 10724536 TI - Unexpected formation of glycals upon attempted C-alkylation of protected glycosylacetylenes. AB - Treatment of 3,7-anhydro-4,5,6,8-tetra-O-benzyl-1,2-dideoxy-D-glycero-D-galacto oct-1 -ynitol (beta-D-mannosyl acetylene, 1) with 5 equivalents of n-butyllithium at either 0 or -78 degrees C resulted in the elimination of benzyl alcohol to yield 3,7-anhydro-5,6,8-tri-O-benzyl-1,2,4-trideoxy-D-arabino-oct-3-en-1-yn itol (glycal acetylene, 3) as the major product. Additional studies showed that 3 is also produced from two isomers of 1 with alpha-D-mannosyl and beta-D-glucosyl stereochemistry, but in lower yields. Furthermore, substrates in which the acetylene moiety is replaced by either a methyl or phenyl group do not produce a glycal product under these conditions. Finally, treatment of 1 with phenyllithium provides 3 in low yield. Deuterium labeling studies suggest that the reaction proceeds through an E2, rather than an E1cB, mechanism. PMID- 10724537 TI - Complete 1H and 13C NMR assignment of mono-sulfated galactosylceramides with four types of ceramides from human kidney. AB - The full assignment of 1H and 13C NMR signals of galactosylceramide 3-sulfate (galactosyl sulfatide) and 1H signals of galactosylceramide 6-sulfate was achieved by using 1H-1H DQF-COSY and 1H-13C heteronuclear COSY. Analyses were performed on a mixture of galactosyl sulfatides with four representative ceramide types consisting of a combination of non-hydroxy or 2-hydroxy fatty acids and sphingenine or 4D-hydroxysphinganine (trihydroxysphinganine) as the long-chain bases. The 1H and 13C NMR parameters of galactosyl sulfatide with 4 hydroxysphinganine as well as 13C signals of complex lipids with 4 hydroxysphinganine were elucidated for the first time. Not only sulfation of the galactosyl residue, but also modification of the aglycon, including hydroxylation of fatty acids and hydration of the double bond in sphingoid bases, altered the chemical shifts substantially. In addition, the unique long-range coupling constants, 4J(H,H) and 5J(H,H), in the galactosyl residue of galactosyl sulfatide could be determined. PMID- 10724538 TI - The sickle-cell kid. An experimental transplant succeeds, giving a brave little boy the best Christmas present he can imagine. PMID- 10724539 TI - Bad to the bone. How to explain evildoers like Harris and Klebold? Some psychologists say there's a diagnosis at hand. PMID- 10724540 TI - Telemedicine project unites remote region with urban expertise. AB - In western Kansas, physicians and hospitals linked to peers at urban centers to cover gaps in their own coverage and specialty offerings. The project has saved them money and kept revenues at home. PMID- 10724541 TI - Process redesign wins patient satisfaction, Codman Award for Illinois hospital. PMID- 10724542 TI - New CABG guidelines break down risk, options. AB - New recommendations from the American College of Cardiology outline which patients are most likely to benefit from bypass surgery. Drawing from recent research, the guidelines note that the procedure is often safe for women, the elderly and other groups previously believed to be at greater risk. PMID- 10724543 TI - Improving quality in small hospitals. AB - Often, the organizations profiled by THE QUALITY LETTER are the powerhouses of the healthcare industry, with the resources, researchers and technology to make quality improvements at the cutting edge of the field. This month, THE QUALITY LETTER looks at small and rural hospitals--often with 100 or fewer beds, isolated from competition or the pressures of managed care--that have made the most of their opportunities to improve the quality of care they deliver. Many of the issues they face and the answers they find are applicable to organizations of any size. PMID- 10724544 TI - Improving physician recruitment for rural hospitals. AB - Faced with a shortage of primary care and specialty physicians at its 23 facilities, Lifepoint Hospitals developed an aggressive recruiting and screening program to bring the right physicians to small towns--and help them stay. PMID- 10724545 TI - Dyslexia and the new science of reading. PMID- 10724546 TI - A writing fool. PMID- 10724547 TI - Letting the doc decide. A huge insurer finds a new way to control costs. PMID- 10724548 TI - Not Mother Nature's way. A new study shows more and more women get drugs during labor. Is this progress? PMID- 10724549 TI - A prescriptive palette. Researchers say the pigments that give foods their color also can cut cancer and heart-disease risk, and ease the pain of arthritis. PMID- 10724550 TI - The one-size dose does not fit all. Physicians and patients should look beyond the guidelines recommended by drug manufacturers. PMID- 10724551 TI - Culture, self-rated health and resource allocation decision-making. AB - It has been observed that some groups in society tend to report their health to be better than would be expected through more objective measures. The available evidence suggests that while variations in self-assessed measures of health may act as good proxies of mortality and morbidity in homogeneous populations, in some groups, such as the Aboriginal and Torres Strait Islander communities of Australia, these subjective measures may provide a misleading picture. Useful insights into the formation of health perceptions can be drawn from a range of disciplines, in particular, from social comparison theories, models of illness behaviour, survey literature and linguistics. These theories and models help to provide an understanding of the different ways in which illness may be perceived, evaluated and acted upon by different kinds of people. Such considerations can have very direct implications for those planning and evaluating public health programs as well as those responsible for funding such programs. PMID- 10724552 TI - "If a patient is too costly they tend to get rid of you:" the impact of people's perceptions of rationing on the use of primary care. AB - Despite the increasing focus on rationing, and rationing decisions in the NHS, little attention has been given to patient's perceptions of rationing and the potential impact this might have on people's use of services. Drawing on the qualitative findings of a study conducted in the North West of England which was concerned with the pattern and processes of primary care help seeking, this paper sets out to examine perceptions and experiences of rationing in primary care and the potential impact this has on people's use of services. In relation to primary care services people had experienced rationing by deterrence, dilution and delay. There was some evidence that perceptions of rationing impacted on help seeking and the use of primary care services. The implications for understanding the way in which perceptions of rationing might influence the formulation of demand and help seeking by people using primary care services are discussed. PMID- 10724553 TI - QALYs and the integration of claims in health-care rationing. AB - The paper argues against the polarization of the health economics literature into pro- and anti-QALY camps. In particular, we suggest that a crucial distinction should be made between the QALY measure as a metric of health, and QALY maximization as an applied social choice rule. We argue against the rule but for the measure and that the appropriate conceptualization of health-care rationing decisions should see the main task as the integration of competing and possibly incommensurable normative claim types. We identify the main types as consequences, rights, social contracts, individual votes and community values and note situations in which the contribution of each claim type is limited. We go on to show that the integration of (at least some of) these claim types can be formalized within the mathematical framework provided by non-linear programming. PMID- 10724554 TI - Postmodern health economics. AB - Postmodernism and health economics are both concerned with questions about choices and values, risk and uncertainty. Postmodernists seek to respond to such questions in the context of a world of uncoordinated and often contradictory chances, a world devoid of clear-cut standards. Health economics seeks to respond using the constructs of modernity, including the application of reason to generate better order. In this article we present two sorts of voice. First we introduce postmodernism and those seeking to contribute to economics from a postmodern perspective. Second, we consider critics of a prevalent neo-classicism within health economics both from outside that paradigm and from those more closely associated with it. It is increasingly evident that (health) economics, as presently constituted, is failing both in its descriptive powers and its prescriptive possibilities. Postmodernism offers not just an alternative theoretical approach but the possibility of both expanding the scope of health economics and grounding it more appropriately in the everyday experience of those engaging with health systems. PMID- 10724556 TI - Relationality and consensus in Japan: implications for bioethics policy. AB - This paper examines the Japanese notion of relationality, that is, the idea that the individual is defined primarily within a web of relationships. Furthermore, it proposes that this relationality provides an ontological basis for morality, particularly the critical need for achieving consensus. This need for consensus is evident in the dispute over brain death. It was also conspicuous in the long standing debate regarding heart transplantation. By reviewing key features of relationality, the study also demonstrates that the Japanese approach toward consensus reflects certain cultural values such as the importance of nemawashi. This inquiry thereby evokes the brain death and heart transplantation controversy in order to illustrate the critical need for consensus in the decision-making process. PMID- 10724555 TI - Privatized biomedical research, public fears, and the hazards of government regulation: lessons from stem cell research. AB - This paper discusses the hazards of regulating controversial biomedical research in light of the emergence of powerful, multi-national biotechnology corporations. Prohibitions on the use of government funds can simply force controversial research into the private sphere, and unilateral or multilateral research bans can simply encourage multi-national companies to conduct research in countries that lack restrictive laws. Thus, a net effect of government regulation is that research migrates from the public to the private sphere. Because private research receives less oversight and external scrutiny than public research, it can threaten the welfare and rights of human subjects, scientific progress and openness, and the quality of the approval process for new biomedical technologies. In order to avoid the harmful effects of government regulation of biotechnology, society should promote meaningful discussion and dialogue among scientists, industry leaders, and the public before resorting to regulatory solutions. Legislative or executive initiatives should be applied with great discretion and care, and should be crafted in such a way that they protect public health and safety, promote scientific progress, and avoid the hazards of privatized research and polarized debates. PMID- 10724557 TI - Combating the 'safe' cigarette: ethical, public health issues and regulatory proposals. AB - Regulatory authorities have advised smokers who would not or could not quit smoking to switch to lower tar cigarettes. Smoking such cigarettes was seen as a means of reducing the harm caused by smoking, but not as offering a 'safe' smoking option. Correspondingly manufacturers have been required to place tar and nicotine information on packet labels and/or advertisements. This paper explores the possibility that the conventional format for conveying tar and nicotine information could be responsible for the belief, held by a significant proportion of smokers, that some brands of lower tar cigarettes are absolutely 'safe'. To deal with this situation it is suggested that changes should be made to health warnings, and tar and nicotine communications. Proposed changes to the latter are evaluated in terms of their ethical and public health implications. The authors conclude that brand specific warnings and a classification of cigarettes as either 'Very Dangerous' or 'Dangerous', is best suited to reconciling consumer needs for information with the public health objectives of reducing the harm caused by smoking. PMID- 10724558 TI - HMO consolidation: a threat to network integration? AB - The ongoing wave of consolidations among major health plans and insurers across the country is creating anxiety and challenges for integrated networks and for providers in general. The federal government's recent action in intervening in the Aetna U.S. Healthcare/Prudential Health Plans merger offers some hope, say provider executives, that some providers may see some policy relief in the future. But integrated providers in highly managed care-penetrated markets like those of the large urban markets on the West Coast are already operating in an environment of high tension and consolidation. PMID- 10724559 TI - Clinical information systems: strategic imperatives driving IDNs forward. AB - Reviewing the "state of the art" of clinical information systems in healthcare, one can't help but be struck by how many promises remain unfulfilled. Yes, there have been advances in electronic medical information systems in the past three decades, but in many ways, the same problems exist today that have existed all along. Implementation of clinical information systems (CIS) still requires major changes in workflow. And those core problems continue to dog the progress of integrated healthcare delivery systems, which are struggling to demonstrate value to purchasers, payers, providers, and consumers as they aggregate vertically and horizontally across the continuum. At the same time, mastering the "people" issues, those that revolve around how technology is integrated into the clinical practice of medicine, continues to be the key success factor for producing a usable solution, even with all the advances in hardware and software. PMID- 10724560 TI - OIG proposes draft compliance program guidance for nursing facilities. PMID- 10724561 TI - Industry, consumer groups square off on minimum nurse staffing requirements. PMID- 10724562 TI - How to avoid the most common, costly and dangerous MDS miscoding errors: Part II- Correct coding of Sections E and P. PMID- 10724563 TI - Seeing the wood for the trees: defining the forgotten concept of patient dissatisfaction in the light of patient satisfaction research. AB - Studies of patient satisfaction are regarded by many as the most important way to obtain patients' views. To date, relatively few studies have focussed specifically on dissatisfaction. Concerns have been expressed about the validity of the concept of satisfaction. Dissatisfaction, however, has received little attention since it has been assumed to be the opposite of satisfaction and thus already defined. Therefore a series of assumptions have also been made about dissatisfaction, which may or may not compromise its validity or usefulness. The aim of this review is to clarify the concept of dissatisfaction by examining what studies of patient satisfaction can and cannot tell us about dissatisfaction; identifying assumptions; and finally by suggesting how research might best be oriented to accommodate the complexity of patient experiences. PMID- 10724564 TI - Hip fracture as a complication of hospitalization. AB - This work seeks to assess the possible contribution of hospitalization to hip fractures sustained in an acute care hospital and to determine the need for hospital care for these patients at the time of the fracture. Between 1988 and 1997 there was an average of 399 falls and four in-hospital hip fractures per year. For 14 percent, no predisposing factors for falling were noted, 38 percent of the fractures occurred within the first three days and 47 percent during the first week of hospitalization. Original admission did not seem warranted for 10 percent and 48 percent no longer required inpatient care at the time of the fracture. Most fractures occur early during hospitalization; some patients seem to have no predisposing factors for falling and about one-half may not require hospitalization at the time, all implicating hospitalization as a causative factor. PMID- 10724565 TI - Promoting the concept of quality within his organization. PMID- 10724566 TI - Quality management in Irish health care. AB - This paper reports on the findings from a quantitative research study of quality management in the Irish health-care sector. The study findings suggest that quality management is what hospitals require to become more cost-effective and efficient. The research also shows that the culture of health-care institutions must change to one where employees experience pride in their work and where all are involved and committed to continuous quality improvement. It is recommended that a shift is required from the traditional management structures to a more participative approach. Furthermore, all managers whether from a clinical or an administration background must understand one another's role in the organisation. Finally, for quality to succeed in the health-care sector, strong committed leadership is required to overcome tensions in quality implementation. PMID- 10724567 TI - Exploring the possible reasons why the UK Government commended the EFQM (European Foundation for Quality Management) excellence model as the framework for delivering governance in the new NHS. AB - A brief introduction into recent developments of the EFQM Excellence Model and the United Kingdom (UK) Government's agenda for ensuring that quality is at the heart of all decision making is given. In view of the Government explicitly commending the use of the EFQM Excellence Model to all organisations within the National Health Service, the author decides to explore the possible reasons behind the commendation. When comparing the EFQM Excellence Model with the Government's vision for quality, the former emerges as a more than ideal tool for any organisation wishing to commence or strengthen their journey on the road to quality and/or excellence; particularly as the EFQM Excellence Model is based on the principles of self-assessment, continuous improvement, learning and innovation, teamwork and a culture totally focused on the customer. Finally, ten possible reasons behind the Government commending the use of the Model are given. PMID- 10724568 TI - Q methodology, risk training and quality management. AB - The results of a Q methodological study of professional understandings of the notion of risk in mental health services within the UK are discussed in relation to the relevance for staff training and quality assurance. The study attempted to access the diversity of understandings of risk issues amongst a multi professional group of staff (n = 60) attending inter-agency risk training workshops in 1998. Q methodology is presented as both an appropriate means for such inquiry and as a novel experiential technique for training purposes. A tentative argument is advanced that the qualitative accounts generated by Q research could assist in systematic reviews of quality, complementing the singularly quantitative approaches typically represented in the audit process. PMID- 10724570 TI - Managerial governance to parallel clinical governance. PMID- 10724569 TI - Clinical governance in accident and emergency services. AB - Owing to NHS managers' preponderance with financial issues, the present Government made improving the quality of health services a statutory requirement in 1997. In this article, one means of improving the quality of health services, clinical governance, is examined in detail before some issues related to its implementation are described. The Trust's A&E services, the context for interpreting and applying clinical governance, are briefly described before introducing a force-field analysis that demonstrates the different elements when changing services broadly and clinical governance specifically. The final section concentrates on implementing and improving clinical governance in A&E departments. PMID- 10724571 TI - A developmental performance framework for primary care. AB - Primary care in the NHS changed substantially during the 1990s. In recent years, structural changes, most notably the introduction of primary care groups as an administrative centre for planning, have added impetus to the need for adopting meaningful measures of quality of the primary care service. This qualitative study reports the views of a sample of general practitioners, primary care nurses, and practice managers on the development and refinement of current performance indicators. Seven themes were identified as key areas for development of indicators of performance: patient experience, clinical activity, service development and innovation, access, health promotion, cost effectiveness, and quality of life outcomes. These themes are incorporated into a dynamic framework for development where improved outcomes (including quality of life measures) are seen as central to the evaluation of quality, and inter-professional collaboration in the delivery and evaluation of quality of the new primary care is called for. PMID- 10724572 TI - Measuring the inappropriate utilization of accident and emergency services? AB - Accident and Emergency (A&E) departments are increasingly popular venues for primary care, causing a serious threat to healthcare quality. This paper reports the development of a comprehensive research method for identifying primary care patients attending A&E. Patients were randomly selected from the four A&E departments across different time periods and different regions in Hong Kong. The definition of GP cases was based on a retrospective record review conducted by a panel of emergency physicians using the standard laid down by the Hong Kong College of Family Physicians. The patients sampled were similar in sex and age distribution to A&E attendees for the whole territory. The level of GP cases was found to be 57 per cent, with a significantly higher proportion of patients in the younger age group. The high level of use reflects the lack of a well co ordinated development of primary care services and interfacing with secondary care. PMID- 10724574 TI - Emergency psychiatric assessments: do outcomes match priorities? AB - The targeting of scarce mental health resources is currently organised around three broad areas: treating severe mental illness; reducing suicide; and obviating risk to the wider community. High priority clients are those who present with either psychotic symptoms, or who are perceived to be high risk to self or others. This study examined records of emergency assessments at a mental health trust in the south-east of England over a three-month period (n = 336) to see whether clients with these characteristics are more likely than others to be offered hospital admission or treatment. It was found that clients presenting with psychosis are much more likely to be offered admission than those assessed as non-psychotic, that those assessed as a suicide risk are also significantly more likely to be offered admission, and that those assessed as presenting a risk to the public at large are no more likely to be admitted than those presenting no risk. This investigation indicates that the emergency service of the trust studied is addressing the first two target areas, but may be underperforming in the third. PMID- 10724573 TI - An analysis of catering options within NHS acute hospitals. AB - Reforms of the NHS's healthcare structure have placed additional pressure on all aspects of hospital management. Evaluation of the effects of these reforms is difficult without more information on current conditions. Hospital catering in acute care trusts has little contemporary background research available. With this in mind, a survey of all the acute care NHS trusts within the eight regions in England was undertaken to investigate the hospital meal service process. A mailed questionnaire asked for the meal production system, food service method and food delivery personnel used by each trust, and a copy of a weekly menu. Results, from an 80.7 per cent response rate, indicate that most trusts use batch cooking to prepare their meals, and plated meal service to deliver the food to the wards. Almost 75 per cent of the trusts use nurses, at least in part, to serve food. English foodstuffs dominate the menus. Most of the trusts have moved towards meeting the goals set by the Patients' Charter and other NHS recommendations. PMID- 10724575 TI - Auditing the implementation of SIGN (Scottish Intercollegiate Guidelines Network) clinical guidelines. AB - Clinical practice guidelines are increasingly being recognised as integral to the clinical effectiveness agenda. According to the recent Scottish White Paper, Scotland "leads the way in clinical effectiveness". The Scottish Intercollegiate Guidelines Network (SIGN), established in 1993, has produced over 20 clinical practice guidelines, and plans to produce at least as many more, while reviewing existing guidelines at a minimum of every two years. This represents a substantial investment of NHS resources. This paper investigates whether this investment is being recouped in Scottish NHS acute trusts via the implementation of SIGN guidelines, and whether their implementation is being audited properly. It is argued that without clinical audit, guideline implementation is unlikely to succeed. This has important ramifications for the implementation of clinical governance. PMID- 10724576 TI - Attack chronic pain to reduce ER visits, hospitalizations. PMID- 10724577 TI - Chronic pain has major long-term consequences. AB - Is pain a disease state? A doctor who has treated pain for more than 20 years explains how pain affects the body from the central nervous system to the heart- and how it affects morbidity and costs if not properly treated. PMID- 10724578 TI - Population-based management of all pregnant women delivers healthy babies. AB - The incidence of preterm delivery in New Hampshire has declined dramatically thanks to the concentrated efforts of the state's largest health plan. Find out how the plan identifies women who may be at risk for early delivery, keeps tabs on them throughout pregnancy, and keeps newborns safe and out of the hospital after delivery. PMID- 10724579 TI - Considering DM for women's health? Dysfunctional uterine bleeding a good place to start. AB - How much are you paying out in costs for hysterectomies, and do the women who have this procedure really need it? Several physicians argue that dysfunctional uterine bleeding, the main reason why women undergo hysterectomies, is a perfect candidate for disease management. PMID- 10724580 TI - Intensive therapy for disturbed children can have dramatic bottom line benefits. AB - Get the details behind an innovative program that works with emotionally disturbed teens to reduce the chances of violence and the costly morbidity that comes with it. PMID- 10724581 TI - Hospitalists: a must-have for capitated organizations. PMID- 10724582 TI - Do you have adequate reserves for your capitated contracts? AB - Have you taken stock of your financial status lately and taken precautions against future economic downturns? A good starting place is to examine your financial reserves. PMID- 10724583 TI - Medicare risk audits result in multi-million dollar recoveries. AB - An Alabama hospital has recovered more than $2 million from its Medicare risk contracts through an internal audit system that examines payments, charge-backs, and ancillary claims. PMID- 10724584 TI - The top six challenges for capitated providers. AB - NHI's exclusive 1999 Capitation Survey uncovered the top six issues that can make or break an organization's future in risk contracting. PMID- 10724585 TI - Is your cap contract protected from changes in member copays? AB - Before you dismiss member copays as little more than an administrative nuisance, consider that they can make the difference between profit and loss when capitated margins are cut to the bone. PMID- 10724586 TI - Medicine at the mall. PMID- 10724587 TI - Strategic issues in marketing and branding centers of excellence. PMID- 10724588 TI - Health plans must act now for employer-based insurance devolution. PMID- 10724589 TI - Forces for change in Canadian healthcare. PMID- 10724590 TI - Healthcare system change and the 21st century. Observations from south of the border. PMID- 10724591 TI - The third millennium and the consumer: some reflections. PMID- 10724592 TI - From principles to practice: the management of post-merger integration. PMID- 10724593 TI - Making our healthcare system the best it can be. PMID- 10724594 TI - Patients' Bill of Rights. PMID- 10724595 TI - New approaches for the elderly show significant results. AB - Capital Health is Canada's largest integrated academic health region, serving a base population of more that 800,000 people in the city of Edmonton and surrounding communities. Capital Health provides a complete range of health services, including transplantation, children's cardiac surgery, innovative community health programs, rehabilitation services, continuing care, palliative care, public health, health promotion and disease prevention. Capital Health's large, integrated health system manages resources well, responds quickly and creatively to challenges and opportunities and blends institutional healthcare with community care and population health. Capital Health is a key partner with the University of Alberta Health Sciences Faculties and provides a rich environment for health research and education. PMID- 10724596 TI - Thoughts on promoting the participation of nurses. PMID- 10724597 TI - Incentive-based compensation: will it work in Canadian hospitals? PMID- 10724598 TI - Health websites worth viewing and recommending. PMID- 10724599 TI - Healthcare in the 21st century: "virtual" hospitals, tele-homecare and robots. PMID- 10724600 TI - Survey provides gold mine of best practice targets. PMID- 10724601 TI - Anecdotes offer cap rates from real-world contracts. PMID- 10724602 TI - What does treatment of depression actually cost? PMID- 10724603 TI - Cost increases looming as HMO premiums take giant leap. PMID- 10724604 TI - Kaiser taps data 'gold mine' to launch CAD team. PMID- 10724605 TI - Patient satisfaction measures more meaningful with new standardized surveying system. AB - New tool standardizes patient surveying and data comparison. Patient satisfaction surveys have been around for years, but often without an effective way for physicians to benchmark the results against those of their competitors. But HCIA Inc. and the Medical Group Management Association may have the answer with their new Patient Satisfaction Comparison Service. See how it works, and review some initial findings. PMID- 10724606 TI - Integrated system links cost data, patient satisfaction scores for the first time. AB - Linking cost data, patient satisfaction scores. HBS International and The Picker Institute have joined forces to make integrated data available that directly links operational efficiency and patient satisfaction. Find out how the systems lets providers know when reducing expenses compromises care. PMID- 10724607 TI - Poor data quality, reporting delays render CA, NY hospital report cards 'nearly useless'. AB - Grading the graders: How useful is the data in hospital report cards? Not very, say providers in California and New York who recently rated their respective states' report cards on a number of criteria. Find out what role poor data quality played in the low ratings. PMID- 10724608 TI - Fund raising on the Internet: instant access to a new world of donors. AB - One advantage of having an online shopping village attached to your not-for profit organization's Web site is it gets supporters used to contributing online. If a consumer is willing to purchase over the Web, it's only a short leap to putting donations more directly to work for a favorite cause. PMID- 10724609 TI - Internet causes dramatic changes in fund raising world. PMID- 10724611 TI - "Visit our web site!" is no longer enough. PMID- 10724610 TI - 2000 non-profit software guide. PMID- 10724612 TI - Promoting from within: the best way to build management. AB - One of the best ways to build knowledgeable, quality management is to promote from within the ranks of your organization. Allowing employees to work their way up the pyramid of promotion will give them the necessary knowledge to run your Call Center effectively and successfully. PMID- 10724614 TI - Follow the money. Hard money. Soft money. Lobbying money. Which buys the most influence in Washington? PMID- 10724615 TI - HealthGrades: playing doctor on the Web. PMID- 10724613 TI - Samaritan's Purse furthers worldwide relief efforts with OCR for Forms. AB - Samaritan's Purse is widely known for our Operation Christmas Child program. The program has been a phenomenal success, growing from 110,000 donated shoe boxes in 1994 to 2.3 million in 1998--an annual growth rate in excess of 110 percent. PMID- 10724616 TI - Beef up your information security with the new HIPAA-mandated standards. AB - Beef up information security using HIPAA standards. This month, the federal Department of Health and Human Services will release the final standards for information security mandated by the 1996 Health Insurance Portability and Accountability Act (HIPAA). To comply with HIPAA, you must perform an applications and data criticality analysis and develop a data backup plan, a disaster recovery plan, and an emergency mode operation plan. PMID- 10724617 TI - Asthma patients prefer ED to hospital. PMID- 10724618 TI - JCAHO shake-up: symptom of serious financial woes? AB - Does JCAHO shakeup portend financial woes? The recent, abrupt reorganization at the Joint Commission on Accreditation of Healthcare Organizations in Oakbrook Terrace, IL, which includes the departure of several vice presidents, may be symptomatic of bigger trouble brewing at the oversight agency. Revenues from JCAHO's accreditation and education efforts are said to be down, especially in the home care and long-term care fields. PMID- 10724619 TI - JCAHO to use ORYX data to detect sentinel events. PMID- 10724620 TI - How to set up a sentinel event response team. PMID- 10724621 TI - Good plans strengthen patient care quality. PMID- 10724622 TI - Into the woods. Earth, air, fire, and water, shaped by local artisans, are the materials of a new spa in western Pennsylvania designed by Clodagh. PMID- 10724623 TI - Exclusive arrangements continue to generate litigation. Major v. Memorial Hospitals Association. PMID- 10724624 TI - New York court rejects liability theory. Megally v. LaPorta. PMID- 10724625 TI - Dismissal of wrongful birth action reversed. Canesi ex rel. Canesi v. Wilson. PMID- 10724626 TI - ERISA preemption defenses rejected. Nealy v. US Healthcare HMO. PMID- 10724627 TI - Pennsylvania Supreme Court rejects insurance coverage for physician's acts. Physician's Insurance Company v. Pistone. PMID- 10724628 TI - Atypical findings of protein studies using capillary zone electrophoresis. AB - We present two atypical cases of protein studies that were evaluated by immunofixation and immunosubtraction using capillary zone electrophoresis and high-resolution agarose gel electrophoresis. The first study showed an abnormal peak in the beta region by capillary zone electrophoresis that was located in the gamma region of the high-resolution agarose gel electrophoresis. Further investigation showed that this monoclonal protein was displaced due to binding with beta-lipoproteins. In the second case, a large peak was detected in the alpha-2 region and was shown by capillary zone electrophoresis to be a non proteinaceous material that mimicked a paraprotein. PMID- 10724629 TI - Determination of the toxicologically relevant arsenic content in urine. AB - A combination of the flow injection technique with hydride AAS for the detection of the toxicologically relevant arsenic sum parameters offers the following advantages: Automated sample preparation Detection of arsenic as As(III), As(V), mono- and dimethyl arsinic acid (MMA and DMA) Good accuracy and precision Very good detection limits Simple calibration against aqueous As3+ solutions. PMID- 10724630 TI - Comparing CLS faculty and allied health deans and directors: time spent in academic activities and perceptions of the research environment. PMID- 10724631 TI - Tracing our roots: years of turmoil (1962-1977). AB - OBJECTIVE: To describe the activities of the American Society of Clinical Pathologists and the sequence of events leading to the establishment of the National Accrediting Agency for Clinical Laboratory Science and Medical Laboratory Personnel. DESIGN: A survey of literature on the history of clinical laboratory science was conducted. References consulted various books and professional journals. CONCLUSIONS: By virtue of its close association with ASCP, ASMT became drawn into a series of legal actions between 1962 and 1977. Although ASMT was eventually dropped from all law-suits, the association's leadership recognized the need for greater autonomy and independence from ASCP. Clinical laboratory science achieved several victories during this period with respect to certification and mandatory re-registration by the BOR. Responsibility for accreditation of educational programs was shifted from the BOS to NAACLS and the establishment of the National Certification Agency for Medical Laboratory Personnel signified ASMT's commitment to certification by the profession for the profession. PMID- 10724632 TI - Laboratory indicators of ethanol consumption. PMID- 10724633 TI - A review of variables affecting PTs/INRs. AB - The prothrombin time is a common method of monitoring patients undergoing oral anticoagulant therapy. The proliferation of commercial thromboplastin brands with different international sensitivity indices (ISI) in conjunction with wider availability of automated coagulation analyzers has elevated the need for standardization in monitoring therapy. PMID- 10724634 TI - Current considerations in the use of the APTT in monitoring unfractionated heparin. AB - This paper focuses on laboratory monitoring of unfractionated heparin by use of the activated partial thromboplastin time. The reader is directed to a remarkably useful document that includes current recommendations for laboratory monitoring of anticoagulation therapy, and was produced in the wake of the XXXI Conference of College of American Pathologists in 1997. The authors' recommendations were based on reviews of current medical literature as well as the consensus of the opinions of recognized experts on the subject of anticoagulation. Points of contention, still seeking resolution, were also expressed. At this date, it is probably the most complete and objective review of current laboratory assessment of anticoagulation available. PMID- 10724635 TI - Heparin induced thrombocytopenia. PMID- 10724636 TI - Estimating the particle size characteristics of therapeutic aerosols. PMID- 10724637 TI - Time-of-flight aerodynamic particle size analyzers: their use and limitations for the evaluation of medical aerosols. AB - Time-of-flight (TOF) aerosol analyzers are a class of instruments that measure the aerodynamic diameter of individual particles following a controlled acceleration in a well-defined flow field. Two instruments have been used to analyze the size of medical aerosols: Aerosizer particle size analyzer (TSI Particle Instruments/Amherst, Amherst, MA), Aerodynamic Particle Sizer (APS) aerosol spectrometer (TSI) Both instruments are capable of sizing several thousand particles a second, making it possible to obtain aerodynamic particle size distributions in a few seconds compared with up to 1 hour per measurement using compendial methods that are based on either the multistage liquid impinger or cascade impactor. This rapidity makes TOF analysis attractive for product development, as many different variables can potentially be investigated during a short period of time. The data thus obtained should be used with caution, however. Several issues, most notably the lack of a direct relationship with the mass of drug substance present and the vulnerability of the measurements to coincidence effects when sampling concentrated aerosols, may severely limit the value of data from many aerosol delivery systems, especially pressurized metered dose inhalers (pMDIs). A review of the literature illustrating the issues that are involved and providing guidance on the most appropriate uses of these analyzers is presented. PMID- 10724639 TI - Aerosol characterization of three corticosteroid metered dose inhalers with volumatic holding chambers and metered dose inhalers alone at two inspiratory flow rates. AB - Inhaled corticosteroids are first-choice drugs in the treatment of chronic asthma. A metered dose inhaler (MDI) equipped with a spacer device is easier to use for patients with a poor inhalatory technique; it favors a reduction in the size of the particles delivered to the patient and thus a reduction in the incidence of local and systemic side effects of these drugs. The aim of this study was to determine the particle characteristics of fluticasone propionate (FP), flunisolide (FLUN), and beclomethasone dipropionate (BDP), each administered at a rate of 250 micrograms per puff and at inspiratory flow rates of 30 and 60 L/min in vitro, to estimate the particle characteristics of these drugs aspirated via an MDI alone and via a large-volume holding chamber (Volumatic). Compared with the MDI alone, at 30 L/min, the Volumatic (Glaxo Wellcome, Ware, UK) significantly reduced the mass median aerodynamic diameter (MMAD) and increased the fine particles (< 5 microns and < 2 microns) generated by all three drugs. At 60 L/min, the MMAD increased and the generation of fine particles decreased with both devices. These data suggest that the inspiratory flow applied by means of the devices may be a determinant for the deposition of the drug in the lower airways in that by increasing the inspiratory flow, the MMAD increases and the percentage of fine particles decreases, probably because of the reaggregation favored by the higher flows. PMID- 10724638 TI - Reducing adverse effects of inhaled fenoterol through optimization of the aerosol formulation. AB - We compared the adverse effects of 160 micrograms of fenoterol in the form of a 2.8-micron monodisperse aerosol with those of 800 micrograms of fenoterol as a conventional metered dose inhaler (MDI) aerosol plus spacer. Previously, a monodisperse aerosol was shown to elicit equivalent degrees of bronchodilation at an 80% lower dose using a standard MDI. A total of 12 healthy volunteers (8 women and 4 men) participated in this study and inhaled in random order a placebo, the monodisperse aerosol, and the MDI aerosol. Changes in serum potassium level, finger tremor, blood pressure, heart rate, and specific airway conductance were measured before and 15 minutes after administration. Compared with placebo, the active aerosols elicited a significant improvement in airway conductance and adverse effects. Serum potassium level decreased by 0.27 mmol/L after the monodisperse aerosol, and the MDI lowered it by 0.67 mmol/L (P = 0.001). Finger tremor also increased less: 0.07 versus 0.29 V (P = 0.029). Changes in cardiovascular parameters were not significantly different from those elicited by the placebo. There were no significant specific airway conductance differences between the two active aerosols. By changing the formulation of MDI aerosols, the occurrence of adverse effects can be reduced. PMID- 10724640 TI - Size analysis of a pressurized metered dose inhaler-delivered suspension formulation by the API Aerosizer time-of-flight aerodynamic particle size analyzer. AB - Aerosizer time-of-flight (TOF) aerodynamic particle size analyzers (TSI-Amherst, Amherst, MA) are widely used for the rapid assessment of aerosols from a wide variety of drug delivery devices, including pressurized metered dose inhalers (pMDIs). This technique offers significant advantages in terms of rapid measurement times in comparison with the more time-consuming compendial methods such as the cascade impactor or multistage liquid impinger. Particle size analysis takes place by determining the TOF of individual particles following acceleration to supersonic velocity. No drug assay is performed; thus, the resulting size distribution also includes particles that do not contain any medication such as the excipients and surfactant that are present in most pMDI based formulations. Illustrative data are presented for one particular formulation (Pulmicort: 200 micrograms of budesonide per dose; Astra Draco; Lund, Sweden) and demonstrate that bias from this source can significantly shift the reported particle distribution to finer sizes compared with impactor-based analysis in which direct assay for drug has taken place. In this case, the mass median aerodynamic diameter (MMAD) determined by an Aerosizer-LD was close to 2.4 microns, but was found to be approximately 4 microns using the cascade impactor based procedure. Such a shift results in an overestimation of the fine particle fraction of the emitted dose, which may lead to misleading conclusions about the therapeutic benefit of a particular drug delivery system when making use of this formulation. TOF aerosol measurement techniques appear to be vulnerable to this type of bias for any suspension formulation in which the drug content is not homogeneously distributed within all particle sizes. PMID- 10724641 TI - Do sinusoidal models of respiration accurately reflect the respiratory events of patients breathing on nebulizers? AB - The amount of drug that is delivered by nebulization is a combination of the physical properties of the agent being nebulized, the performance of the nebulizer, and the pattern of breathing of the patient. To avoid biological variation, mechanical models of breathing are frequently employed during the evaluation of the performance of a device. For simplicity, many investigators use sinusoidal models of breathing to calculate the expected inhaled mass, although some use square waves and other more complex models. Most assume that the duration of inspiration (Ti) is half of the total respiratory time (Ttot). This study compared the calculated inhaled mass from which the expected pulmonary deposition was estimated from the actual pattern of breathing of 43 children with cystic fibrosis (CF) breathing from an unvented nebulizer with a low dead volume and appropriate particle size distribution with that from a sinusoidal pattern of breathing using the same tidal volume (VT) and respiratory rate. The respiratory duty cycle (Ti/Ttot) was 0.45 +/- 0.05, which meant that less time was spent during inspiration than that found in a pure sinusoidal pattern. The difference between the predicted deposition from the actual pattern of breathing and that calculated from the sinusoidal model was 12 +/- 7%, which correlated with the respiratory rate (r = 0.67, P < 0.001). The degree of lung disease did not influence the discrepancy between the two values. In general, the actual VTs and respiratory rates were less in the patients than those employed in mechanical models of pediatric breathing. Although some patients had respiratory patterns that could be represented accurately with a sinusoidal model, most did not, and there were wide variations from child to child. These results suggest that there are both systematic and random errors arising from the use of a sinusoidal waveform to mimic respiratory events in patients. PMID- 10724642 TI - Intrapulmonary distribution of deposited particles. AB - Inhalation drug delivery for both topical and systemic treatments has many advantages over oral, intravenous, or subcutaneous drug delivery. Because some drugs should be deposited within the bronchial tree and others should deposit within the respiratory zone of the lung, it should be possible to determine and influence the preferential site of drug deposition to develop efficient inhalation therapy strategies. In this article, a method that allows estimation of the longitudinal distribution of deposited particles in the lungs of individual subjects is introduced. From the photometrically measured deposition of monodisperse di-2-ethylhexyl sebacate (DEHS) droplets, the longitudinal distribution of deposited particles (i.e., the number of particles that are deposited in a certain lung volume element) can be assessed. In this study in four healthy volunteers the distribution of deposited particles was assessed for different airflow rates, tidal volumes (VTS), and particle sizes. The results showed that there are considerable differences in the longitudinal distribution of deposited particles between subjects and that the distribution is strongly dependent on particle size: if particle size is increased, the site of particle deposition is shifted proximally. Particles with diameters greater than approximately 5 microns cannot penetrate to a volumetric lung depth (VP) greater than approximately 600 cm3 even if the VT is increased. Airflow rate has a minor effect on the distribution of deposited particles, but if airflow rate increases, the site of particle deposition is slightly shifted peripherally. This method can be used to investigate individual patterns of drug deposition in human lungs noninvasively and to develop and optimize inhalation strategies for inhalation drug delivery. PMID- 10724643 TI - In vitro comparison of salbutamol hydrofluoroalkane (Airomir) metered dose inhaler aerosols inhaled during pediatric tidal breathing from five valved holding chambers. AB - Amounts of salbutamol delivered from chlorofluorocarbon (CFC)-free metered dose inhalers (MDIs) (Airomir; 3M, St. Paul, MN) in particle sizes appropriate for inhalational treatment with five common holding chambers (NES [or Nebuchamber] spacer [Astra Draco AB, Lund, Sweden], AeroChamber [Trudell Medical, London, Ontario, Canada], OptiChamber [Health-scan Products, Cedar Grove, NJ], Vent170 spacer [Nordac Design, Waterloo, Ontario, Canada], and E-Z Spacer [WE Pharmaceuticals, Ramona, CA]) when used under simulated pediatric tidal breathing conditions were determined. Five devices of each type were tested with Airomir hydrofluoroalkane (HFA) inhalers (100 micrograms of salbutamol). Each device was connected to face replicas representative of 7-month-old (infant) and 2-year-old (toddler) children, and aerosol was also inhaled into an Andersen cascade impactor (Graseby Andersen, Smyrna, GA) using a valve system and simulated tidal breathing patterns representative of children of these ages. Amounts of drug inhaled in fine particles with the HFA formulation are significantly less (up to 46% less) with the E-Z Spacer (P < 0.01) compared with amounts inhaled in previous studies with the CFC formulation but are nearly the same with other holding chamber types (e.g., differing by < 6% for the AeroChamber). With the HFA formulation, the metal NES spacer delivered significantly more salbutamol in fine particles (P < 0.01) than any of the other holding chambers. Amounts of salbutamol inhaled in fine particles during pediatric tidal breathing from valved holding chambers with Airomir varies considerably between holding chamber types. PMID- 10724644 TI - In vivo evaluation of a phosphorylcholine coated cardiopulmonary bypass circuit. AB - A complete phosphorylcholine coated cardiopulmonary bypass circuit, including the Dideco D901 oxygenator, was tested for gas transfer, blood path resistance, and biocompatibility in a standardized setting. Blood compatibility was tested by measuring complement and platelet activation. Three dogs (mean body weight 28 +/- 3 kg) were placed on cardiopulmonary bypass at a flow rate of 600 mL/min during 6 hours. The animals were weaned from cardiopulmonary bypass and sacrificed electively after 7 days. Oxygen and carbon dioxide transfer were 26.6 +/- 2.4 mL/min and 33.0 +/- 1.9 mL/min, respectively. Mean pressure drop across the oxygenator was 52.6 +/- 0.2 mmHg. The respective baseline values for thromboxane B2, prostaglandin E2 and platelet factor 4 were 1817 +/- 283 pg/mL, 12783 +/- 2109 pg/mL, and 0.35 +/- 0.08 IU/mL. Thromboxane B2 and prostaglandin E2 increased slightly to 2881 +/- 868 pg/mL and 18083 +/- 3144 pg/mL at 30 minutes of bypass, whereas platelet factor 4 values remained stable curing the procedure. Concentrations of complement split products C5a were only mildly increased. After use scanning electron microscopy was performed on the inner housing, heat exchanger, and outer surface of the hollow fibers. No thrombi nor organized cellular deposits were found on any of the components. Phosphorylcholine coating of CPB seems to be very promising regarding platelet activation and complement activation. PMID- 10724645 TI - Matrix metalloproteinase inhibitor: differential effects on pulmonary neutrophil and monocyte sequestration following cardiopulmonary bypass. AB - Acute respiratory distress syndrome (ARDS) following cardiopulmonary bypass (CPB), also known as "post-pump" or "post-perfusion syndrome" (PPS), results from sequential priming and activation of neutrophils. We hypothesized that chemically modified tetracycline (CMT-3) an inhibitor of neutrophil matrix metalloproteinase (MMP) and elastase, would prevent PPS. We performed histometric analysis of lung tissue from our porcine PPS model to correlate cellular sequestration and histologic injury with CMT-3 treatment. METHODS: Yorkshire pigs were randomized into five groups: Control (n = 3); CPB (n = 5); femoral-femoral bypass 1 hour; LPS (n = 7), Escherichia coli lipopolysaccharide (1 microgram/kg); CPB + LPS (n = 6); and CPB + LPS + CMT (n = 5), sequential insults and CMT-3. Protocol histometric analysis defined cellular and tissue components of lung injury. RESULTS: CMT-3 decreased neutrophil sequestration in the CPB + LPS + CMT-3 group (p < 0.0001 vs. CPB + LPS). There were no differences in monocytes between CPB + LPS and CPB + LPS + CMT treatment groups. CONCLUSIONS: CMT-3 attenuates neutrophil sequestration but has no effect on mononuclear sequestration in our PPS model. This finding supports current research on leukocyte chemokines and has important implications regarding mechanisms of CMT-3. Despite lack of monocyte response to CMT-3, PPS was prevented by inhibiting neutrophils alone; confirming the primary role of neutrophils in PPS. PMID- 10724646 TI - High dose thrombin time versus the activated clotting time during cardiopulmonary bypass. AB - In this study we compared the High Dose Thrombin Time (HiTT) with the Activated Clotting Time (ACT) during cardiopulmonary bypass (CPB) in non-aprotinin treated patients. On the advice of the HiTT test manufacturer each institution should perform comparative ACT/HiTT assays in the cardiac surgery population. In previous tests our target ACT value of 480 seconds corresponds with a mean HiTT value of 190 seconds. Our results showed that after heparinization (300-400 IU/kg body weight) 8 out of 20 patients did not reach the target ACT of 480 seconds, while the HiTT results in those 8 patients were higher than our target time of 190 seconds. Four heparin pretreated patients who received 400 IU/kg heparin, had relatively low ACT values (467 +/- 14 sec.) and high HiTT values (324 +/- 47 sec.). Before and during CPB there was a poor correlation between the HiTT and ACT (r = 0.38). The results of this study show that for the individual patient the target HiTT of 190 seconds is no guarantee for reaching an adequate ACT of 480 seconds. Although the HiTT may be a very useful assay for monitoring heparin effects during CPB, the determination of the target time can be a point of discussion. In contrast of the advice of the manufacturer we therefore suggest that comparative ACT/HiTT assay should be done in every individual patient to determine a safe target HiTT time, instead of the whole group of patients. PMID- 10724647 TI - Clinical evaluation of the CDI-100 in-line hematocrit/saturation monitor. AB - This study was undertaken to evaluate the accuracy, reliability, consistency and biases of the CDI-100 saturation monitor when compared with a blood gas analyzer. The advantage of continuous in-line monitoring is that the perfusionist has continuous updates as to the patient's changing physiologic state. During this study, if the sample readout of the CDI-100 was off by greater than 10% from that of the Gem-Premier, the CDI-100 parameter was recalibrated. The accuracy of the CDI-100 was fair (greater than 10% of the samples needed recalibration) with regards to the initial sample comparisons. Recalibration was needed 67% of the time for the hematocrit and 35% for the saturation. The reliability of the CDI 100 was good (no equipment failure). The CDI-100 was consistent. It consistently overestimated both the hematocrit and saturation. This overestimation is the bias of the monitor. We recommend recalibration of the CDI-100 during clinical use to insure greater accuracy. PMID- 10724648 TI - An improved bladder for pump control during ECMO procedures. AB - A new inline reservoir called the Better-Bladder, now FDA-cleared for long term use, overcomes some disadvantages of the silicone bladder and bladder box used in extracorporeal membrane oxygenation circuits. The Better-Bladder provides compliance in the venous line and allows for noninvasive pressure measurements. Both features are useful for controlling pump speed as a function of venous line pressure. Bench tests showed that the Better-Bladder measures pressure noninvasively within +/- 4% of invasive (i.e., liquid contacting) pressure measurements in a range from -200 to +500 mmHg and at temperatures from 10 degrees C to 37 degrees C. After 60 days, the error in noninvasive pressure measurement with the Better-Bladder was less than +/- 3%. The Better-Bladder withstood pressurization to 1700 mmHg for ten days without leaking or failing in other ways. The advantages of the Better Bladder, along with its accuracy and durability, suggest its use for short and long term pumping applications. PMID- 10724649 TI - Staffing issues at open-heart centers offering both pediatric and adult perfusion service: 1998 survey results. AB - A survey directed to centers offering both pediatric and adult perfusion services was conducted to determine how pediatric cases were distributed among individual perfusionists in their departments. These centers were also asked what they believed the clinical activity level should be for a perfusionist each year to remain proficient in pediatric cardiopulmonary bypass. The questions were asked via e-mail and then followed up with telephone interviews as necessary. Out of the 100 centers contacted, 45 responded to the survey (43 North American, 2 European). Of the forty-five centers, forty-one provided both pediatric and adult perfusion services. Thirty-two centers (78%) offering adult as well as pediatric perfusion services distributed the pediatric caseload to a select group of perfusionists. Nine centers (22%) distributed the pediatric open-heart caseload to the entire staff. From the respondents, the average minimum number of pediatric cases believed necessary to remain proficient in pediatric perfusion was 42.8 cases annually. Centers having dedicated pediatric perfusionists had a slightly higher annual caseload than did those at non-specialized centers, despite practicing at institutions averaging fewer pediatric open-heart cases annually. PMID- 10724650 TI - The additive effects of antifibrinolytics: dangers in the OR. AB - The use of antifibrinolytic agents in recent years had been heralded as a major breakthrough in the prevention of postoperative bleeding. However, whenever the delicate balance between coagulation and fibrinolysis is altered, the potential exists for disaster. There have been a number of complications reported in the literature related to the use of antifibrinolytics. With the availability of oral epsilon aminocaproic acid (EACA) as treatment for the symptoms of hemophilia and other secondary bleeding disorders, the possibility exists that a patient presenting for heart surgery may already be receiving antifibrinolytic therapy. A 72 year-old female underwent double valve replacement which was complicated by postoperative bleeding. Her medical history included gastrointestinal bleeding which was being treated with oral EACA. The patient was heparin resistant with a dose response of 55 sec/unit/ml. Heparin was administered during cardiopulmonary bypass to maintain the ACT between 400-500 seconds using a heparin-coated bypass circuit. In spite of this, the patient became thrombocytopenic and bled excessively in the postoperative period. Administration of additional antifibrinolytic agents in the operating room may be contraindicated when the patient is receiving this therapy preoperatively, and may contribute to the development of a procoagulant state during bypass. PMID- 10724651 TI - Getting the most out of CAD data. AB - Three-dimensional modelling enables high-resolution visualization of parts, assembly optimization and simulation of the manufacturing process. This article highlights the development of this technology and the new applications it offers for more rapid product development. PMID- 10724652 TI - Natural latex free cold seal packaging. AB - From 30 June 1999, in the United States, warning labels must be attached to all medical devices and packaging containing natural latex. In Europe the risk to public health associated with natural latex proteins is receiving increasing attention. This article suggests possible solutions and discusses emerging synthetic systems that may emulate the performance of natural latex. PMID- 10724653 TI - Design, development and performance of a novel multidose dry-powder inhaler. AB - The authors describe the design and development of a breath-actuated multidose dry-powder inhaler and summarize the in vitro and in vivo data demonstrating its robustness and performance in the laboratory and during clinical use. Drugs for the treatment of asthma--including budesonide, beclomethasone dipropionate and salbutamol--when formulated with lactose powder as a carrier and dispensed via this device, have exhibited clinical efficacy and safety profiles comparable with standard pressurized metered-dose inhalers and dry-powder formulations. PMID- 10724654 TI - The stresses and strains of everyday life. AB - Stresses and strains are features of everyday living, but the distinction between the two is not always easy to identify. These matters are discussed in mechanical and physiological terms with relevance to medical devices. PMID- 10724655 TI - The in vitro diagnostic directive. What it doesn't say, but manufacturers need to know. AB - By now, everyone should have a copy of the In Vitro Diagnostic (IVD) Directive. After careful reading, people are realizing that there is much that the Directive does not say and much that is wide open to interpretation--interpretation that could make a big difference to the IVD industry. This article highlights those parts of the Directive that are unclear and therefore will need to be discussed and interpreted. PMID- 10724656 TI - Notified body consensus statements. AB - The ability of governments, official bodies, industry and others to uniformly interpret the European Directives for medical devices will have a profound effect on the success of the European system regulating these products. This is particularly important for Notified Bodies. This article will discuss Notified Body consensus statements. An article in the near future will discuss Notified Body recommendations. PMID- 10724657 TI - The laboratory testing market in 1998: self-testing sales boom in a flat market. PMID- 10724658 TI - Microtechnologies: art or engineering? AB - Recent developments in micromachining are showing that much more can be achieved at acceptable costs than may have previously been thought possible. One of the biggest challenges, says the author, is unleashing the engineers' imagination. This article tackles some of the problems found in making and using machined microcomponents and offers some solutions. PMID- 10724659 TI - The new industrial revolution. PMID- 10724660 TI - Saving home health. PMID- 10724661 TI - Punching the clock. PMID- 10724662 TI - Getting back in the game. PMID- 10724663 TI - Staying in the black. PMID- 10724664 TI - Quality or productivity? PMID- 10724665 TI - The making over of a rehab unit. PMID- 10724666 TI - Creative reimbursement. Finding funding sources for lifts. PMID- 10724667 TI - Moving forward. PMID- 10724668 TI - Postacute integration. PMID- 10724669 TI - The evolution of education. PMID- 10724670 TI - Athletic training's regulatory history. PMID- 10724671 TI - Faster functioning. PMID- 10724673 TI - Home health compliance. PMID- 10724672 TI - Using a team approach. PMID- 10724674 TI - Increasing independence. PMID- 10724675 TI - Subject to further review. PMID- 10724676 TI - Planting hope. PMID- 10724677 TI - Who and how to test for hypercoagulability. PMID- 10724678 TI - Another ingredient added to HER2 mix. PMID- 10724679 TI - Paying the price to link lab systems. PMID- 10724680 TI - ISO 9000: real results or repetition? PMID- 10724681 TI - How to orchestrate a CAP inspection. PMID- 10724682 TI - Images moving from incompatible to integrated. PMID- 10724684 TI - Clinical laboratory information system. PMID- 10724683 TI - Verifying coagulation tests' accuracy, reliability. PMID- 10724685 TI - Improving chronic disease care in the real world: a step-by-step approach. PMID- 10724686 TI - Toward a new paradigm for public health practice and academic partnerships. AB - Despite substantial progress in establishing academic and public health partnerships over the past 20 years, two questions require further examination: (1) How are the most effective partnerships achieved? and (2) How well are these partnerships suited to the current and future problems of public health? The authors propose the "new public health" perspective, which offers challenges to develop a dialogue and power relationships among partners based on symmetry rather than conventional asymmetry. New forms of discourse and benchmarking of progress in improving the health of the public may be achieved by adopting this paradigm. PMID- 10724687 TI - Demonstrating excellence in academic public health practice. The Association of Schools of Public Health Council of Public Health Practice Coordinators. PMID- 10724688 TI - A decade of progress in academic/practice linkages. AB - Almost a decade has passed since the release of recommendations of the Public Health Faculty/Agency Forum, a national effort designed to expand academic/practice linkages. Has anyone really cared? Has anything really changed? Indeed, the academic community in particular has responded by integrating more public health practice into teaching, research, and service. This article explores where progress has been made and where continuing efforts still are needed. In addition, potential ways to continue enhancing academic/practice linkages are discussed. PMID- 10724689 TI - Public health practice in schools of public health: is there a fit? AB - Public health practice has been a part of schools of public health for a long time and yet it is still an emerging field of scholarship. Practice faculty often are isolated in their schools and not part of the mainstream. This article explores the elements of culture that are necessary to support a practice focus in a school of public health. PMID- 10724690 TI - A framework for assessing practice-oriented scholarship in schools of public health. AB - Within many schools of public health, substantial ambiguity surrounds the process of defining and rewarding faculty contributions to practice-oriented scholarly activities. Ernest L. Boyer introduced a powerful and compelling new paradigm for assessing the core domains of scholarship: the scholarships of discovery, integration, teaching, and application. This article draws on Boyer's domains and his colleagues, Charles E. Glassick, and others' criteria for evaluating them in introducing a framework for assessing practice-oriented scholarship within schools of public health. The article presents concepts, content, and defining criteria of practice-oriented scholarship and identifies the principles that might guide tenure and promotion decisions related to it. PMID- 10724691 TI - Reconsidering scholarship. AB - Following the publication of Scholarship Reconsidered in 1990, debates began on many campuses regarding the expansion of the definition of scholarship. These debates focused on the appropriateness of the scholarship of discovery, the scholarship of integration, the scholarship of application, and the scholarship of teaching. These conversations were enriched by the six standards for assessment proposed in Scholarship Assessed, published in 1996. Now these concepts prosper on many campuses as individual institutions find their own interpretations and adaptations of enriched and expanded scholarship. PMID- 10724692 TI - Lessons learned from an academic health department. AB - Consensus has been reached in the United States on the mission and core functions of local health departments as well as services required to maximize community health status. Most local health departments do not have the financial or professional resources required to carry out all of the responsibilities assigned them. One way to improve competence is to develop partnerships with other agencies and institutions. One type of partnership that should be explored is linkages between local health departments and academic institutions. An example of such a partnership is described in this article. PMID- 10724693 TI - The adolescent years: an academic-community partnership in Harlem comes of age. AB - Much has been written about the potential benefits in health promotion that are possible through partnerships between academic institutions and community-based organizations, but little practical advice has been provided on how to sustain these relationships when the original grant funds have been exhausted. Here we document our experiences in Harlem, New York City, a community with grave social, structural, and physical environmental inequities, and describe the successes and failings of a partnership now in its "adolescence" between researchers at the Joseph L. Mailman School of Public Health of Columbia University and community activists at West Harlem Environmental Action (WE ACT). PMID- 10724694 TI - Strengthening local public health practice: a view to the millennium. AB - From a national perspective, local public health practice continues to face many of the challenges identified in the 1988 Institute of Medicine study. Unless they are addressed broadly, current trends may exacerbate further the capacity of local public health agencies. Efforts to strengthen local practice should focus on workforce development, scale of operations, and resources. New and innovative models of collaboration between academia and local practice are proposed. PMID- 10724695 TI - Collaboration for health improvement: models for state, community, and academic partnerships. AB - The value of approaching health improvement through collaborative models seems self-evident, especially at the community level. A review of the literature, however, reveals a number of challenges in terms of helping us understand precisely what it is that makes collaborative models effective in influencing health. The purpose of this article is to describe important elements of collaboration that state, local, and academic partners may wish to consider when developing models for health improvement. The author will provide examples of collaborative strategies that are emerging from the Turning Point: Collaborating for a New Century in Public Health initiative. PMID- 10724696 TI - A bold new future for public health or implementing the "Alexander Haig" idea. AB - An amalgam of effort by public health practitioners and academics is needed to attain the goal of public health taking charge of U.S. health care early in the new millenium. Numerous ideas for joint collaborations in service, research, and teaching are presented. Goals, quality improvement and reporting, customer satisfaction, and leadership training are developing strengths in the field that will help make rapid progress toward the goal. Oversight of health services by public health is essential to the development of a successful U.S. health system. PMID- 10724697 TI - Data uses, benefits, and barriers for the behavioral risk factor surveillance system: a qualitative study of users. AB - The purpose of this study was to describe data use, benefits, and barriers among BRFSS users. A trained facilitator conducted eight focus groups of eight to 12 state health department employees. NUD*IST qualitative software was used to code responses. Users viewed the BRFSS as an invaluable data set. Data were used most frequently for public education, trend analyses, planning, policy support, and program evaluation. Common barriers to data use included limited availability of regional and subgroup data, lack of data analysis skills, and inadequate staff resources. Users described the BRFSS as a beneficial data source, but some barriers impede its usefulness. PMID- 10724698 TI - Challenges in electronic importing of health data. AB - The objective of this article is to increase awareness among public health personnel of the complexities involved in integrating existing data systems. This article describes the electronic importing feature of the Centers for Disease Control and Prevention (CDC) software package called staffTRAK-TB, and users' experience with it. PMID- 10724699 TI - My reverie. PMID- 10724700 TI - How telemedicine is rearranging our universe. PMID- 10724701 TI - Considering the alternatives. AB - Health care providers frequently talk about values and community care, and their mission statements often espouse a commitment to cooperative relationships with their patients. Good intentions aside, the reality is that some health care providers may be unable to accommodate patients who are unable to pay for services. Distrust is guaranteed when an industry's words and deeds don't match, but several caregivers are thinking outside the lines to find solutions to their patients' payment problems. PMID- 10724702 TI - Reference checking: don't get lost in the translation. AB - Reference checking these days has gotten to be a real challenge. Assuming you can get somebody to talk to you, it's often a game of cat-and-mouse to get to the truth. Employers are becoming so careful (and fearful of litigation) that what they often will say is difficult to interpret. PMID- 10724703 TI - A procrastinator's guide to the Y2K problem. AB - Your workplace has undoubtedly spent a great deal of time and money over the past few years on the Year 2000 (Y2K) problem. The focus of this article, however, is not on Y2K in the health care industry. Frankly, if your organization is not ready by now, it's too late. Your home computer, however, may be another story. PMID- 10724704 TI - The abuse of executive power. PMID- 10724706 TI - Digital management--a necessity for Michigan hospitals. PMID- 10724705 TI - Raising consciousness. AB - A new device is being used at McPherson Hospital in Howell that allows anesthesiologists to measure the consciousness of patients during surgery and distribute anesthesia accordingly. The Bispectral Index (BIS) monitor is the first and only commercially available direct measure of the effects of anesthetics on the brain, and McPherson is currently one of only a handful of hospitals in Michigan and about 300 across the country to use this technology. PMID- 10724707 TI - Developing a pain management team. PMID- 10724708 TI - The community benefit role of the collections department. PMID- 10724709 TI - Building relationships is high priority. MHA's account executive program. PMID- 10724710 TI - Health education the fun way! PMID- 10724711 TI - Plastics. PMID- 10724712 TI - Digital technology is the future of health care. PMID- 10724713 TI - Market testing. In or out? PMID- 10724714 TI - Waste disposal. Where there's muck. PMID- 10724715 TI - Ancillary staff. Cinderella no more. PMID- 10724716 TI - NHS supplies. Buying into the performance culture. PMID- 10724717 TI - Mergers. Apart at the seams. AB - More than a fifth of trusts in England are involved in mergers. The Department of Health has not revealed the rationale for its drive on mergers. Small, potential cost savings are often outweighed by the expense of the merger process and loss of morale and productivity. Many NHS consultants are not involving themselves in redesigning services following mergers because they cannot see a way forward in the face of so many contradictory central demands. Support for this massive change programme is not adequate. PMID- 10724718 TI - Bed management. Very model of a modern major general. AB - Allocating bed management to a team of nurses providing a 24-hour service has helped free beds and increased admissions at an inner-city hospital. The team's 24-hour presence has reduced persistent out-of-hours problems. Since the team has been in operation, the number of times ambulances have to be diverted to other hospitals has been reduced. PMID- 10724719 TI - Professional training. Learning the hard way. AB - An audit of nursing, midwifery and physiotherapy training commissioned by a consortium of trusts in Sussex showed diverging views on the relevance of courses. Many nursing students wanted more practical experience. Two-thirds of students reported financial hardship and a third were working at the same time as studying. PMID- 10724720 TI - Pharmaceuticals. Adopt an orphan. PMID- 10724721 TI - Prevention of HIV infection through changes in sexual behavior. AB - Ample evidence exists that behavioral interventions reduce high risk sexual behaviors and promote safer practices. Downstream interventions in settings attracting high risk patients work well, especially with infected persons to avert new infections. Preparing health care workers for intensive, skills-based interventions grounded in behavioral science theory would enhance primary prevention. Midstream interventions have reliably reduced risk in primary care and community settings. Adoption of comprehensive skills-based programs in schools is controversial but likely to improve outcomes. Upstream community approaches have slowed human immunodeficiency virus (HIV) incidence among men having sex with men. Upstream policy interventions remain underutilized in the U.S. but have been successful internationally. Needed are a national HIV prevention strategy and research linking behavior change to reduced HIV seroprevalence. PMID- 10724722 TI - Progress in dietary behavior change. AB - Nutrition has important influences on health and mortality. Public response to current dietary guidance is indicated by positive movement on some indicators, notably reductions in calories from fat and in blood cholesterol levels. Downstream case finding and intensive educational and behavioral interventions are often effective for high risk and motivated subjects and for persons with diet-related disease. Midstream environmental changes (e.g., in grocery stores, schools) proffer improved nutritional choices and supports. Individualized dietary counseling has yet to become the norm. Despite recent upstream success in developing broad dietary guidance and improving nutrition labeling policies, numerous avenues remain open for aggressive national policy developments. Emerging studies and continuing improvement in intervention methods, measurement, and research design will help realize the preventive effects of healthful diets. PMID- 10724723 TI - How are we doing with physical activity? AB - An estimated 60% of U.S. adults are inactive or underactive, and nearly half of America's youth (aged 12 to 21 years) are not vigorously active on a regular basis. Downstream interventions provide individual strategies that effectively increase short-term participation in physical activity by 10% to 25%. Downstream and midstream approaches tailored to individual preferences have greater success. Packaging and disseminating physical activity programs for community, worksite, and health care settings are not as far along as for other areas, although inactivity prevalence is about twice that of smoking, and both risk factors have substantial morbidity and mortality. Less is known about effectiveness of upstream approaches, which have potential for the greatest public health impact. Suggestions include continued promotion of moderate-intensity physical activity, greater dissemination of successful programs, and investigation of physical environment influences. PMID- 10724724 TI - Current trends in the integration and reimbursement of complementary and alternative medicine by managed care organizations (MCOs) and insurance providers: 1998 update and cohort analysis. AB - OBJECTIVES: To assess the status of managed care and insurance coverage of complementary and alternative medicine (CAM) and the integration of such services into conventional medicine. METHODS: A literature review and information search was conducted to determine which insurers had special policies for CAM. Telephone interviews were conducted with a definitive sample of 9 out of 10 new MCOs or insurers identified in 1998 and a cohort of eight MCOs and insurers who responded both to the original survey in 1997 and again in 1998 to determine trends. RESULTS: This study constitutes the results of the second year of a 3-year ongoing survey. For 1998, 10 MCOs and insurance carriers initiated CAM coverage. Survey results are analyzed for these 10 new providers as well as the results of a cohort of eight insurers surveyed in both 1997 and 1998 to determine current trends. A majority of the insurers interviewed offer some coverage for the following: nutrition counseling, biofeedback, psychotherapy, acupuncture, preventive medicine, chiropractic, osteopathy, and physical therapy. All new MCOs and insurers said that market demand was their primary motivation for covering CAM. Factors determining whether insurers would offer coverage for additional therapies included potential cost-effectiveness, consumer interest, demonstrable clinical efficacy, and state mandates. Among the most common obstacles listed to incorporating CAM into mainstream health care were lack of research on efficacy, economics, ignorance about CAM, provider competition and division, and lack of standards of practice. CONCLUSIONS: Consumer demand for CAM is motivating more MCOs and insurance companies to assess the benefits of incorporating CAM. Outcomes studies for both conventional and CAM therapies are needed to help create a health care system based upon treatments that work, whether they are conventional, complementary, or alternative. PMID- 10724726 TI - Population-based tobacco control: progress and prospects. AB - One in four U.S. adults smokes. Downstream cognitive-behavioral interventions coupled with effective pharmacotherapy can produce 40% quit rates, particularly for those least addicted, most highly motivated, and without psychiatric comorbidity. Effective midstream school-based prevention activities delay youth use. Worksite programs and physician "quit smoking" advice can be cost-effective, although these are not sufficiently widespread. Community strategies show promise of preventing youth use and helping addicted users quit. Despite failed federal tobacco control legislation, great strides have been made upstream, including proposed regulation of nicotine as a drug, the state master settlement agreement with the tobacco industry, and excise tax increases funding statewide tobacco control programs. Wider dissemination of effective programs and better coordination with upstream policies hold great potential to significantly reduce future use rates and related disease. PMID- 10724725 TI - Rating our progress in population health promotion: report card on six behaviors. AB - Using McKinlay's population model of prevention, this series assesses the current state of the art for six lifestyle behaviors: tobacco use, alcohol abuse, drug abuse, unhealthy diet, sedentary lifestyle, and risky sexual practices related to human immunodeficiency virus/acquired immunodeficiency syndrome (HIV/AIDS). More progress has been made in "downstream" individually oriented treatments than in broader, more environmentally focused interventions. Promising trends include: a shift toward lower cost minimal-contact and self-help "downstream" programs; the development of tailored messages and stage-based "midstream" initiatives that can reach everyone in a defined population or setting; and the emergence of "upstream" policy advocacy strategies. Improving the power and reach of health behavior change will require advances in biobehavioral research to develop more powerful behavior change strategies along with efforts to more widely disseminate the effective interventions that already exist. Growing evidence supports McKinlay's premise that full-spectrum (downstream to upstream) interventions are needed for greatest population impact. Progress also will depend on finding new ways to address the needs of special populations--including underserved low income groups, racial and ethnic minorities, individuals with multiple risk behaviors, and youth and their families. PMID- 10724727 TI - Treatment and prevention of use and abuse of illegal drugs: progress on interventions and future directions. AB - State-of-the-art downstream interventions are generally successful for half of drug-abusing clients. But, only one in four abusers actually receives treatment. In the midstream, one setting (schools), one type of prevention ("one size fits all"), and a limited age-range focus (adolescence and preadolescence) have predominated. Accumulating evidence casts doubt on the effectiveness of widely disseminated school-based prevention approaches, although theory-based programs that emphasize skills training and adjunctive parent and neighborhood interventions fare better. Newer pursuits include intervening very early with higher risk children and expanding to primary health care settings and workplaces. Popular but unproven community approaches need more rigorous evaluation. Upstream national and state public policy and environmental interventions should be reexamined in light of their success for preventing tobacco and alcohol use. PMID- 10724728 TI - Managing alcohol problems and risky drinking. AB - While overall alcohol consumption and alcohol-related automobile deaths have declined, rates of alcohol dependence, liver cirrhosis, and alcohol-related problems remain high among adults, and binge drinking continues as a major health risk for high school and college students. Some individual-level downstream interventions have been evaluated with sufficient rigor to recommend widespread dissemination, and widened availability of new pharmacotherapies could further increase effectiveness. Midstream population-based programs, such as screening and brief interventions in hospitals and managed care organizations, may have greater public health impact than tertiary treatment because of early identification and low cost. Upstream programs and policies that place limits on alcohol availability (e.g., higher legal purchasing age) have the greatest potential to reduce morbidity and mortality at the least cost to society. PMID- 10724729 TI - If plain-film x-ray loses money, why go digital? PMID- 10724730 TI - For mammography, it's digital vs. screen-film. PMID- 10724731 TI - Obstacles remain on road to digital mammography. PMID- 10724732 TI - Digital x-ray: unwavering commitment collides with monumental challenges. PMID- 10724733 TI - Guidelines, algorithms, critical pathways, templates, and evidence-based medicine. PMID- 10724734 TI - Nostradamus. PMID- 10724735 TI - Recommendations for uniform reporting of data following major trauma--the Utstein Style: an initiative. International Trauma Anaesthesia and Critical Care Society (ITACCS) AB - Basic and advanced care of trauma patients always has been an important aspect of prehospital and immediate in-hospital Emergency Medicine, involving a broad spectrum of disciplines, specialties, and skills delivered through Emergency Medical Services Systems which, however, may differ significantly in structure, resources, and operation. This complex background, at least in part, has hindered the development of a uniform pattern or set of criteria and definitions. This in turn, has rendered data incompatible, with the consequence that such differing systems or protocols of care cannot be evaluated or compared readily with acceptable validity. Guided by previous consensus processes evolved by the ERC, the AHA, and other International Organisations represented in ILCOR--on Uniform Reporting of Data following Out-of-hospital and In-hospital Cardiac Arrest--the Utstein Style, an international working group of ITACCS, has drafted a document, Recommendations for Uniform Reporting of Data following Major Trauma--the Utstein Style. The reporting system is based on the following considerations: 1) A structured reporting system based on an "Utstein style template" that would permit the compilation of data and statistics on major trauma care, facilitating and validating independent or comparative audit of performance, and quality of care (and enable groups to challenge performance statistics that did not take account of all relevant information); 2) The Recommendations and Template should encompass both out-of-hospital and in-hospital trauma care; 3) The Recommendations and Template should permit further intra- and inter-system evaluation to improve the quality of delivered care and identification of the relative benefits of different systems and innovative initiatives; and 4) The Template should facilitate studies setting out to improve epidemiological understanding of trauma; for example, such studies might focus on the factors that determine survival. The document is structured along the lines of the original Utstein Style Guidelines publication on "prehospital cardiac arrest". It includes a glossary of terms used in the prehospital and early hospital phase as definitions, time points, and time intervals. The document uses an almost identical scheme for illustrating the different process time clocks--one for the patient, one for the dispatch centre, one for the ambulance, and finally, one for the hospital. For clarity, data should be reported as core data (i.e., always obtained) and optional data (obtained under specific circumstances). In contrast to the graphic approach used for the Utstein Template for pre- or in-hospital cardiac arrest, respectively, the present Template introduces, for the time being, at least, a number of terms and definitions and a semantic rather than a graphic report form. The document includes the following sections: I. INTRODUCTION AND BACKGROUND: II. TRAUMA DATA STRUCTURE DEVELOPMENT: A general outline of the development of structured data using object-oriented modelling (which will be discussed in due course) and includes a set of explanatory illustrations; III. TERMS AND DEFINITIONS: Outlines terms and definitions in trauma care, describing different types of trauma (blunt, penetrating, long bone, major/combined, multiple/polytrauma, and predominant trauma); IV. FACTORS RELATING TO THE CIRCUMSTANCES OF THE INJURY: Describes the following items: a) Cause of injury e.g., type of injury (blunt or penetrating), burns, cold, crush, laceration, amputation, radiation, multiple, etc.; b) Severity of Injury--e.g., prehospital basic abbreviated injury score developed by the working group. The score contains anatomical and physiological disability data, with the anatomical scale ranging ordinally from "1" = head to "9" = external; the physiological disability scale ranges ordinally from "0"; c) Mechanism of injury--recording for transportation incidents etc; e.g., the type of impact, possible restraining devices, PMID- 10724736 TI - Glossary of new concepts in disaster medicine: a supplement to Gunn's Multilingual Dictionary of Disaster Medicine. AB - The development of disaster medicine as a science is dependent on clear definitions of its language. This article proposes a set of definitions to supplement those currently accepted. PMID- 10724737 TI - San Jose Declaration. Pan-American Health Organization. Done at San Jose, Costa Rica, 4 June 1999. PMID- 10724738 TI - The physician's role in Canada's disaster response system. AB - The most recent tragedy in Manitoba illustrates that disasters can strike any community. Within Canada, a tiered disaster response system exists. Even though physicians often play an integral role in the disaster plan, few participate in the planning process or even appreciate their potential role in the event a disaster should occur. Physician involvement would guarantee health matters be appropriately addressed resulting in reduced mortality and decreased morbidity. There are ample opportunities to become involved in disaster planning and response at all levels of government. The objective of this paper is to inform physicians about the disaster planning infrastructure that exists in Canada, show them where they may get involved, and urge them to do so. PMID- 10724739 TI - Differences in mortality rates among trauma patients transported by helicopter and ambulance in Maryland. AB - INTRODUCTION: A comprehensive state-wide emergency medical services and helicopter transport system has been developed in the State of Maryland on the principle that early definitive care improves patient outcomes. The purpose of this study was to determine if empirical data exist to support the theory that air medical transportation services provided by the Maryland State Police (MSP) Aviation Division contribute to an improved trauma patient survival rate in Maryland. METHODS: A retrospective study was conducted on the records of all patients transported by helicopter or ground ambulance and admitted to the R Adams Cowley Shock Trauma Center (STC) of the University of Maryland Medical System. Data were obtained from the Maryland Institute of Emergency Medical Services Systems (MIEMSS) Shock Trauma Clinical Registry for the period January 1988 through July 1995, covering 23,002 patients. Patients included those transported directly from the scene of injury to the STC as well as those from interfacility transfers. All patients were stratified by injury severity and compared by outcome (mortality) using Mantel-Haenszel statistics. RESULTS: During the study period, 11,379 patients were transported by ground and 11,623 were transported by MSP helicopter. The mean Injury Severity Score (ISS) for patients transported by ground was 12.7 (SD = 12.52) and the mean ISS for patients transported by air was 14.6 (SD = 13.42), p < 0.001. Among patients classified as having a high index of injury severity, the mortality rate was lower among those transported by MSP helicopter than among those transported by ambulance. The mortality rate was significantly lower for air transported patient with an ISS higher than 31. CONCLUSION: The State of Maryland has demonstrated a commitment to its citizenry and invested heavily in its public safety air medical service. This study suggests the rapid air transport of victims of traumatic events by specialized personnel in Maryland has a positive effect on the outcome of severely injured patients. Further research is necessary to clarify the causal relationships in order to more fully elucidate the value of this resource. PMID- 10724740 TI - Methodological aspects of the medical examination of special contingents in the All-Russian Service of Disaster Medicine. AB - The tasks required for the preservation of the health of rescuers can be accomplished with the creation of a system for medical examinations and rehabilitation that functions with commonly used methodological and organizational principles. Absence of such a common methodological platform often results in the disqualification of members of the special contingent and a mistake in the evaluation process that gives birth to serious medical and social problems. The specific character of the rescuers' activities during conditions associated with extraordinary situations necessitates the development of criteria based evaluation systems for determining the prognosis, and the clinical, diagnostic, normative, and legal, medical, and social criteria that include methodological approaches that define the criteria to be used in the examinations. We propose that the concept of the professional health examinations becomes the methodological basis used for the medical examinations and rehabilitation of the rescuers. PMID- 10724741 TI - Predictors of demand for emergency prehospital care: an Australian study. AB - INTRODUCTION: Determining the predictors of demand for emergency prehospital care can assist ambulance services in undertaking policy and planning activities. HYPOTHESIS: Demand for prehospital care can be explained by demographic, health status, and economic determinants. METHODS: The study used a cross-sectional design to investigate the association of demographic, health status, and insurance factors with the use of prehospital, ambulance care. Core data items including age, gender, marital status, country of origin, triage score, diagnosis, time of presentation, method of arrival, and patient disposition were collected for every patient who presented at the Emergency Department of the study hospital over a four-month period. Ambulance usage was analysed using Poisson regression. RESULTS: For the 10,229 patients surveyed, only a small number were triaged as having the highest level of urgent medical need (0.8%), but the majority of these used prehospital emergency medical care (90.2%). Predictors of ambulance use included age > 65 years (Prevalence Ratio [PR] = 2.92; 95% confidence interval [CI]: 2.35-3.63), being married or in a de-facto relationship (PR = 0.69; 95% CI: 0.60-0.79) or divorced, separated, or widowed (PR = 0.83; 95% CI: 0.70-0.98), triage score level 1 or 2 (PR = 1.95; 95% CI: 1.68-2.28), or triage score level 3 (PR = 1.54; 95% CI: 1.38-1.72), diagnosis involving either mental (PR = 4.29; 95% CI: 1.84-10.01), nervous (PR = 2.74; 95% CI: 1.19-6.31) or trauma (PR = 2.33; 95% CI: 1.03-5.27) conditions, and insurance status (PR = 1.54; 95% CI: 1.40-1.71). Ethnicity, gender, and time of day were not associated with usage. CONCLUSION: Demand for ambulance services can be predicted by a number of demographic, medical status, and insurance variables. Age and triage levels are key influences on demand for ambulance services. Ambulance insurance status provides an economic incentive to use ambulance services regardless of the urgency of the medical condition. PMID- 10724742 TI - Triage ability of emergency medical services providers and patient disposition: a prospective study. AB - STUDY OBJECTIVE: To determine the ability of emergency medical services (EMS) providers to subjectively triage patients with respect to hospital admission and to determine patient characteristics associated with increased likelihood of admission. METHODS: A prospective, cross-sectional study of a consecutive sample of patients arriving by ambulance during the month of February 1997 at an urban, university hospital, Emergency Department. Emergency medical services providers completed a questionnaire asking them to predict admission to the hospital and requested patient demographic information. Predictions were compared to actual patient disposition. RESULTS: A total of 887 patients were included in the study, and 315 were admitted to the hospital (36%). With respect to admission, EMS providers had an accuracy rate of 79%, with a sensitivity of 72% and specificity of 83% (kappa = 0.56). Blunt traumatic injury and altered mental status were the most common medical reasons for admission. Variables significantly associated with high admission rates were patients with age > 50 years, chest pain or cardiac complaints, shortness of breath or respiratory complaints, Medicare insurance, and Hispanic ethnicity. The EMS providers most accurately predicted admission for patients presenting with labor (kappa = 1.0), shortness of breath/respiratory complaints (kappa = 0.84), and chest pain (kappa = 0.77). CONCLUSION: Emergency medical services providers can predict final patient disposition with reasonable accuracy, especially for patients presenting with labor, shortness of breath, or chest pain. Certain patient characteristics are associated with a higher rate of actual admission. PMID- 10724743 TI - 15 years of experience with cardiopulmonary resuscitation in the Kingdom of Saudi Arabia: a critical analysis. AB - The objective of this review is to establish a framework about the educational activities of the Cardiopulmonary Resuscitation (CPR) National Committee of the Saudi Heart Association (SHA) and determine if it has had any effect on the survival rate in daily hospital work. Further, the review puts forward recommendations regarding the key to success for future implementations and improvement in the outcome of heart attacks in the Kingdom of Saudi Arabia (KSA). Cardiopulmonary resuscitation (CPR) was introduced into the Kingdom of Saudi Arabia in the 1980s. The birth of CPR in the Kingdom was conducted by the American Heart Association (AHA) provision of the first instructor course in Basic Cardiac Life Support (BCLS) and Advanced Cardiac Life Support (ACLS) in the spring of 1984. This educational activity was initiated by the Postgraduate Center of the College of Medicine and currently is a function of the Saudi Heart Association (SHA). The National Heart Center (NHC) continually expands its activities. The number of courses organized, conducted, and reported herein totaled 459 for providers and instructors in BCLS and ACLS. This resulted in certification of 916 and 204 instructors in basic and advanced CPR respectively. There were 80 centers established in the Kingdom over the span of 15 years. They all provide BCLS courses; only 13 provide ACLS courses. The SHA issued a total of 84,659 certificates. PMID- 10724746 TI - Fred C. Cuny Memorial Continuing Education Series, principles of disaster management. Lesson 5: program supervision, monitoring, and control. PMID- 10724745 TI - Care provided by VA mobile clinic staff during Northridge earthquake relief. AB - INTRODUCTION: From 25 January 1994 to 02 February 1994, staff aboard four Veterans Affairs Mobile Clinics treated Northridge earthquake victims. This study examined the types of conditions treated by Clinic staff during the disaster. METHODS: A descriptive case series using 1,123 ambulatory encounter forms was undertaken. Case-mix was assessed by classifying diagnoses into 120 possible diagnostic clusters. RESULTS: Forty-five percent of patients were infants or children and 60% were female. The primary diagnoses were characterized by acute conditions: 1) upper respiratory infection (34.6%); 2) stress reactions (11.9%); 3) otitis media (10.1%); and injuries (8%). Two-thirds of the infants and children either had an upper respiratory infection (46.4%) or otitis media (20.1%). Increasing age indicated an increased likelihood of stress and anxiety reactions. CONCLUSIONS: The results provide additional information for agencies involved in planning for and responding to disasters. Based on the types of conditions diagnosed at the VA mobile clinics (i.e., a high prevalence of acute conditions, including stress and anxiety reactions, and the large numbers of children), staff trained in primary care, mental health, and pediatrics should be considered for relief missions that begin several days after an event resulting in a disaster. PMID- 10724744 TI - Geriatric trauma patients at a suburban level-I trauma center in Japan. AB - BACKGROUND: Despite the increases in the aged population in Japan, there are little data on geriatric patients with traumatic injuries. A prospective clinical study was carried out to evaluate the use of the emergency medical services (EMS) system, mechanisms of injury, and prehospital assessment and triage of elderly victims of trauma. PATIENTS AND METHODS: From July 1996 through June 1997, a group of geriatric trauma (Group G, n = 22) and a control group of younger trauma patients (n = 173) were compared with respect to transfer method to an Emergency Center (direct or indirect), Revised Trauma Scores on the scene of the accident (RTS-1) and on admission to the Emergency Center (RTS-2), and outcome (survival). RESULTS: The mean values for RTS-1 in the Control Group (Group C) were not different from those in Group G, but RTS-2 of the indirect-transfer patients (IP) in Group G were significantly lower than were those for Group C. Group G mortality rates were significantly higher than were the control rates (p = 0.0001). The mortality rate of the IP subgroup was significantly lower than that of the direct transfer subgroup (DP) (30/68 vs. 5/70, p < 0.0001) in the Group C, but mortality rate of the IP subgroup exceeded that of the DP subgroup of Group G (8/14 vs. 5/8). CONCLUSION: The data suggest that in geriatric-age patients, direct transfer patients have a lower mortality rate than do indirect transfer patients when controlled for ISS. Therefore, it seems that a different set of triage criteria should be developed and implemented for geriatric-age victims with trauma-induced injuries and that those who meet these more stringent criteria should be transferred directly to a Trauma Center. PMID- 10724747 TI - Compliance communication in home health care: a mutually reciprocal process. AB - The study described in this article examined the process of compliance gaining in home health care. The investigation focused on nurse-patient communication and the relational and content aspects of compliance communication. Six registered nurses and 25 adult patients from two cooperating home care agencies participated in this study. Observation during home visits and interviews with nurses and patients revealed a prosocial, collaborative model of compliance gaining. The findings show compliance communication to be embedded in nurse-patient conversations, with both nurse and patient engaging in control and affiliative behaviors. Implications for compliance research and the mutual-participation model of medical care are discussed. PMID- 10724748 TI - Double binds and the reproductive and mothering experiences of HIV-positive women. AB - In spite of the increasing number of young women infected with HIV in the United States, little is known about the reproductive and mothering experiences of these women. The purpose of the grounded-theory research discussed in this article was to describe the reproductive and mothering experiences of HIV-positive women. Twenty HIV-positive women participated in 31 in-depth interviews. The grounded theory method was used for data analysis. A communication pattern known in the psychiatric literature as a double bind was discovered to be a basic social psychological problem that affected the women's experiences with reproduction and mothering. An understanding of the power and influence of these double binds permits health care professionals to plan patient-centered programs and to individualize care specifically for HIV-positive women. PMID- 10724749 TI - Developing a safety protocol in qualitative research involving battered women. AB - Qualitative research involving battered women requires advanced planning to protect participants and the investigator from the risk of violence from an abusive partner. Domestic violence creates a potentially dangerous research environment that is seldom discussed in the literature. The use of a safety protocol has been advocated as a way to ensure that research is conducted with maximum safeguards for the participating women. In this article, the author presents a safety protocol developed for a study of battered women's perceptions of danger in their relationship. Issues that relate to safety contacting participants, conducting the interviews, and protecting data are discussed. PMID- 10724750 TI - Narratives of identity: re-presentation of self in people who are homeless. AB - The problem of homelessness is a pressing social and health concern ascribed to the interaction between personal, social, economic, and service system resources. The article is based on a qualitative study of the experiences of 29 homeless individuals. In-depth interviews were conducted with single adult shelter users. Analysis revealed the self to be a process that was continually developing. Participants tacitly locate their self-concepts in the past, present, and future. These time frames reflect the form and content of self. They also reveal hopes, dreams, beliefs, and understandings about self. The ways in which homelessness discredits notions of self and personal identity, and the hierarchy of identify with which homeless individuals use to cope are also examined. PMID- 10724751 TI - Dualistic notions about children with motor disabilities: hands to lean on or to reach out? AB - This article is an attempt to break through the dualistic thinking in theories on the treatment of children with a motor impairment. There is the Cartesian thinking that views the body and mind as separate entities; alternatively, some existential phenomenologists have constructed a dichotomy between the (positively valued) unnoticed body and the (negatively valued) noticed body. Dualistic notions of this kind can lead therapists to treat parts of the child (arms, legs, or speech) rather than the whole child. How can the disadvantageous effects of these dichotomies be overcome in the treatment of children? First, a concrete sketch of the development of corporality in (disabled) children will be provided. Next, the dualistic notions will be discussed. Finally, there is a discussion about how children and their care providers might benefit from this phenomenological explication. PMID- 10724752 TI - Negotiating with helping systems: an example of grounded theory evolving through emergent fit. AB - A strength of substantive grounded theories is that they are modifiable. Yet, little attention is given in the research literature to the evolution of grounded theories through the process of emergent fit. In this article, emergent fit is discussed, and the evolution of the theoretical understanding of relationships with helping systems is provided as an example. In a feminist grounded-theory study of women's caring, emergent fit with existing inductive research on health care relationships resulted in a framework of negotiating, which includes four strategies: reframing responsibility, becoming an expert, harnessing resources, and taking on more. This explanatory model demonstrates how the use of emergent fit can avoid the generation of isolated theories and contribute to knowledge accumulation by producing a substantive theory with wider applicability. PMID- 10724753 TI - Mental illness, caregiving, and emotion management. AB - Based on 50 in-depth interviews, this article considers how caregivers to a spouse, parent, child, or sibling suffering from depression, manic-depression, or schizophrenia manage their emotions overtime. By considering the turning points in the joint career of caregivers and ill family members, our analysis moves beyond studies that link emotions to particular incidences, momentary encounters, or discreet events. Four interpretive junctures in the caregiver-patient relationship are identified. Before diagnosis, respondents experience emotional anomie. Diagnosis provides a medical frame that provokes feelings of hope, compassion, and sympathy. Realization that mental illness may be a permanent condition ushers in the more negative emotions of anger and resentment. Caregivers' eventual recognition that they cannot control their family member's illness allows them to decrease involvement without guilt. The article concludes with a call for research that understands that emotions in groups, settings, or organizations are linked to their distinctive histories. PMID- 10724754 TI - The patient's diagnosis: explanatory models of mental illness. AB - The purpose of the study was to develop a grounded theory about individuals' perception of the situation of being a psychiatric patient. Thirty-five inpatients (19 males, 16 females), ages 18 to 68, in two psychiatric units of an urban, public facility were interviewed on a biweekly basis from admission to discharge. Data were analyzed using the constant comparative method, and the data indicated that participants used the basic social process of managing self-worth to deal with the stigmatizing social predicament of being a mental patient. Events occurring before admission that shaped their responses were substance abuse, medication noncompliance, and the lack of social capital, which led to norm violations and subsequent hospitalization. Six attribution categories emerged: problem, disease, crisis, punishment, ordination, and violation. Findings support the need for professionals to improve their practice by acknowledging the effects of patients' subjective assessments on their response to hospitalization and by placing more emphasis on assisting patients to deal with the stigmatizing effects of a psychiatric diagnosis. PMID- 10724755 TI - Coming out of intensive care crazy: dreams of affliction. AB - The impetus for this article was the author's experience of illness, necessitating 5 months of hospitalization, 7 weeks of which were spent in a coma in an intensive care unit (ICU). The origins and general characteristics of ICUs are noted. A pastiche of illness narrative is constructed to allow for the author's own comment on his experience. The intensive care syndrome (ICS), a permeable category, continuing the aftermath of surgery and the social, psychological, mechanical, and pharmaceutical effects of the unit itself, is discussed. Dreams of affliction, a component of the ICS, are explored; the prior sociological history of the neglect of dreams, an essential universal, is also pointed out. The author's dream forms and their shamanistic parallels are explored. In Foucauldian language, they are considered a product of the fragmentation of the cosmological spirit, whereas the author was a medical object and mechanical appendage. Nursing is in a strategic position to make patients' dream narratives their own as an aid for understanding illness. PMID- 10724756 TI - How best to test for drug resistance. PMID- 10724757 TI - 2000 fee schedule good news for pathology. PMID- 10724758 TI - Automated CBC & 5-part differential hematology instruments. Makings of modern cell counters. PMID- 10724759 TI - Look, learn, leap. PMID- 10724760 TI - Conserving blood in the OR. PMID- 10724761 TI - CDC taps SNOMED for cancer surveillance. PMID- 10724762 TI - Blood banks out of the salvage loop. PMID- 10724763 TI - Using standardized admit orders to improve inpatient care. PMID- 10724764 TI - What does Walt Disney know about patient satisfaction? PMID- 10724765 TI - Disintegration: how employed doctors are landing on their feet. PMID- 10724766 TI - Building a patient registry from the ground up. PMID- 10724767 TI - Avoiding common pitfalls in selecting an EMR system. PMID- 10724768 TI - Reflections on policy and politics in the Clinton/Gore administration: or, how to be "gerontologized" in two easy terms. PMID- 10724769 TI - Understanding the diverse needs of the Medicare population: implications for Medicare reform. AB - Meeting the health care needs of millions of elderly and disabled Americans is central to the debate over Medicare's future. Using data from a nationally representative survey of 3,309 beneficiaries, Medicare's most vulnerable beneficiaries were profiled, examining variations in coverage, satisfaction, access, and financial difficulties. A substantial portion of the Medicare population--two thirds--were found to have health problems or low incomes. The analysis found that about 40% of beneficiaries with incomes below the poverty level, in fair or poor health, or with ADL limitations, have difficulties paying their medical bills or getting needed health care. Medicare's disabled, under-65 beneficiaries are at even higher risk: nearly half (47%) have health care access problems or deal with financial hardship due to medical bills. The diverse needs and experiences of the Medicare population are underscored, providing new insights into the challenge of maintaining or improving protection for those with greatest need while assuring the long-term fiscal viability of the program. PMID- 10724770 TI - No foot in the door: an experimental study of employment discrimination against older workers. AB - Pairs of testers, one aged 57 and one aged 32, applied for 102 entry-level sales or management jobs in the Washington, DC, metropolitan area. Although their credentials described them as equally qualified, the older applicants received less favorable responses from employers 41.2% of the time. Three quarters of these differences occurred before older applicants could fully present their qualifications. The negative employer assumptions about older workers implied by these differences in outcome were seldom explicitly stated. PMID- 10724771 TI - Controlling the supply of long-term care providers in thirteen states. AB - Many states have responded to growing Medicaid long-term care expenditures by limiting the number of long-term care providers through certificate-of-need (CON) programs and moratoriums on new construction or certification for participation in the Medicaid program. This article focuses on the use of these policies in 13 states. Most of the 13 states control the supply of nursing home beds and hospital conversions with CONs or moratoriums, but they are struggling to adapt the role of supply policy to the growth of home health and residential care. As an increasing proportion of Medicaid long-term care spending goes to these nursing home alternatives, supply policy needs to keep pace with the changing provider market and the changing demographics of the consumer market if it hopes to ensure access to long-term care and control Medicaid expenditures. PMID- 10724772 TI - Economic and social implications of aging in Singapore. AB - The economic and social impacts of population aging in Singapore are examined, particularly their impact on labor supply and the extent to which traditional family and community care providers can meet the challenges of an aged population. The adequacy of public policy responses, such as the new employment policy and the Central Provider Fund, are explored. Government strategy to shift the burden of care to the family and other community providers is challenged. PMID- 10724773 TI - New FDA draft guidance on premarket submissions. AB - The United States Food and Drug Administration (FDA) has issued a draft guidance document on the type of quality system information to be included in various premarket submissions. This is the same information that should be maintained at the manufacturing facility for devices subject to the premarket notification or 510(k) process. This article discusses the contents of the draft guidance and recommends that manufacturers send their comments on the guidance to FDA. PMID- 10724774 TI - The quest for high temperature flexible polymers. AB - A unique set of material properties is needed to meet the special requirements of the medical market. Flexible poly(vinyl chloride), which has occupied a pre eminent position for nearly half a century, is increasingly being challenged by newer and more advanced polymer systems. Several propylene-based elastomers with low crystallinities are promising to be the dominant medical materials of the future. This article discusses one company's actual experiences in utilizing these new materials in medical applications. PMID- 10724775 TI - Observations on medical device design, Part II: Good practice. AB - Current guidance on design is inadequate. This second article in a two-part series presents a framework for good design practice that attempts to improve designers' awareness of manufacturing and validation issues. Seven design tactics, derived from observations of current industry practice and design literature, seek to encourage good practice and achieve safer, more profitable devices. PMID- 10724777 TI - UV lasers for drilling and marking applications. AB - Lasers emitting ultraviolet (UV) light have unique capabilities for precision micromachining and marking plastic medical devices. This review of the benefits offered by laser technology includes a look at recently developed UV diode-pumped solid-state lasers and their key features. PMID- 10724776 TI - Adhesives: a selection guide. AB - This article analyses the use of three types of adhesive for medical device assembly. The advantages and disadvantages, typical applications and key features are defined for cyanoacrylate, epoxies and UV- and light-cured adhesives. PMID- 10724778 TI - Sizing extruded medical tubing. AB - When extruding medical tubing, process engineers are faced with many decisions each time a different size, configuration or material is specified for a new product. Sizing is the final forming of a tube to specified dimensions. Three sizing methods are evaluated for optimum tubing production. PMID- 10724779 TI - The god of small things: the biology of intimal hyperplasia. AB - One of the most significant, and as yet unresolved, consequences of placing devices within the vascular system is the uncontrolled growth of the inner layer of the bloodvessel wall following contact with the device. This process is one of the most difficult challenges facing all aspects of intervention; it confronts techniques such as renal-artery stenting, vein grafts, balloon angioplasty, endarterectomy and atherectomy. This article discusses some of the proposed mechanisms for this phenomenon and the prognosis for its control. PMID- 10724780 TI - Annexin V and platelet antigen expression is not altered during storage of platelet concentrates obtained with the AMICUS cell separator. PMID- 10724781 TI - Peripheral blood stem cell collection and transplantation using the haemonetics multicomponent system. PMID- 10724782 TI - Therapeutic plasma exchange and cytapheresis in pediatric patients. AB - Pediatric therapeutic apheresis is reviewed including what it is, how it is performed and indications for its use. Pediatric patients are special, and the unique needs for replacement fluids and attention to access, anticoagulation, volume shifts and hypothermia are stressed. While all indications cannot be addressed, the procedures most commonly performed are reviewed. These include erythrocytapheresis, leukaphereses and plasma exchanges. A table details the strength of evidence supporting the use of apheresis procedures for many of these indications. PMID- 10724783 TI - Antifibrotic tendency as complement to inflammatory burnout? Six year IVIG treatment in a case with atherosclerosis and chronic hepatitis C. PMID- 10724784 TI - Perioperative autologous blood donation in children. AB - Autologous blood donation in children has become a standard of care. Children have to live with the life-time complications associated with allogeneic blood including the transmission of known and unknown pathogens, and the impact of alloimmunization on future blood transfusions, organ transplants and pregnancies. Donor reaction, allogeneic exposure and utilization rates in pediatric preoperative autologous donation (PAD) programs are as good if not better than reported in adult literature. Children are very resilient when undergoing extreme isovolemic hemodilution (IHD). PAD, IHD and intraoperative blood recovery appear to be useful components of a pediatric blood conservation program. Prospective, randomized studies addressing the specific needs of children are required to properly define their perioperative role. PMID- 10724785 TI - Blood transfusion support in pediatric cardiovascular surgery. AB - The majority of children who undergo open-heart surgery with cardiopulmonary bypass (CPB) require perioperative blood transfusion. Blood product requirements are affected by factors such as patient age, underlying cardiac disease, complexity of the surgical procedure, and hemostatic alterations induced by CPB. Transfusion support may include the use of whole blood and/or individual blood components with transfusion practices varying widely based on individual preferences and blood product availability. Approaches to limit allogeneic blood exposure include the use of modified ultrafiltration and smaller bypass circuits, preoperative autologous blood donation and intraoperative blood salvage, and adjunctive antifibrinolytic agents. Potential advantages and disadvantages of the different blood products and pharmacological agents must be considered in managing the pediatric cardiac surgery patient. PMID- 10724786 TI - Blood banking issues pertaining to neonatal red blood cell transfusions. AB - Many preterm infants are given multiple red blood cell transfusions during the early weeks of life. Because firm standards for neonatal transfusions do not exist, it is important to consider the pathophysiology of the anemia of prematurity, the goals of transfusion therapy and blood banking practices that best provide safe and effective neonatal transfusions. There is increasing agreement that efforts continue to minimize phlebotomy blood losses, to transfuse per conservative indications and to limit donor exposure by transfusing stored red blood cells from a single unit reserved for an infant--rather than insisting on fresh blood. PMID- 10724787 TI - Transfusion associated graft-versus-host disease, cytomegalovirus infection and HLA alloimmunization in neonatal and pediatric patients. AB - Modern transfusion support of pediatric patients requires attention to the necessity to provide specialized or modified blood components to these patients who are often immunocompromised and/or affected by very complex medical and surgical illnesses. In this review we will address three potential complications of transfusion that may require specialized components for their prevention in selected patients namely transfusion-associated graft-versus-host disease, transfusion-transmitted cytomegalovirus infection and HLA alloimmunization, with particular reference to the indications for prevention of these transfusion complications in neonatal and pediatric patients. PMID- 10724788 TI - Establishment of new national proficiency testing schemes for leucodepleted blood components: UK. PMID- 10724789 TI - The doctor-patient relationship and prescribing patterns. A view from primary care. AB - The doctor-patient relationship has been described in economic terms as an 'agency relationship' where informed agents make decisions for uninformed clients. However, the decision to prescribe and the decision to accept the prescription by the patient are more complex in nature and involve many variables. Other factors, such as the 'need' for the prescription and the disease state (acute or chronic) also influence prescribing practice. Communication between the physician and patient was found to be important for rational and effective prescribing. The client can make better decisions with the relevant information, thus breaking down the agency relationship that once existed. PMID- 10724790 TI - Measuring quality of life in paediatric patients. AB - The purpose of this paper is to highlight important considerations in the measurement of health-related quality of life (HR-QOL) in paediatric populations. Considerations specific to the evaluation of HR-QOL in children include children's understanding of the questions being asked, their understanding of their own disease, using parents as proxies, time perception problems and simply the fact that children are continually changing. We provide a review of the currently available literature regarding paediatric HR-QOL assessments to examine the different instruments and approaches taken to assess HR-QOL in paediatrics. Asthma-specific measures are discussed as examples of condition-specific HR-QOL instruments because of the considerable amount of methodological and developmental work that has been conducted in this disease. Our search strategy revealed 15 main generic and 5 main asthma-specific instruments that met our criteria for inclusion. The main points high-lighted for each instrument are the description of domains, respondent (i.e. the child, parent or clinician), age group, number of items, format of instrument (usually self-administered or by interview), original country and language, and the existence of translations. For the various studies in which the different instruments are used, the issues of patient population tested, sample size, reliability and validity are addressed. Although this paper does not attempt to be an exhaustive study of paediatric HR QOL instruments, we provide an overview of the main generic and asthma-specific instruments, as well as an examination of the different methods used in assessing paediatric HR-QOL. Many challenges exist in the measurement of HR-QOL in paediatric patients. With increased attention to the considerations high-lighted, the field will continue to grow, and the usefulness and application of HR-QOL measures in paediatric patients is likely to improve. PMID- 10724792 TI - Managing pharmaceutical expenditure while increasing access. The pharmaceutical management agency (PHARMAC) experience. AB - The role of the Pharmaceutical Management Agency (PHARMAC) is to manage pharmaceutical subsidy expenditure in New Zealand. PHARMAC has adopted a proactive approach. It selects the drugs that are to be subsidized and declines to subsidize others. It has established reference pricing across many drug groups, has entered into a range of innovative commercial contracts with pharmaceutical companies, and has encouraged greater price competition among pharmaceutical companies in order to lower prices and control expenditure risk. These initiatives have all been part of an overarching strategy to improve the value of the government's expenditure on pharmaceuticals. PHARMAC has also developed techniques of cost-utility analysis to assess the value of expenditure. PHARMAC has slowed pharmaceutical expenditure growth, culminating in a fall in expenditure in the 1998/1999 year. At the same time, patient access has continued to expand, with more prescriptions being written and new drugs being subsidized. Therefore, PHARMAC has made dramatic strides to improve the value of the government's expenditure on pharmaceutical subsidies and its actions have meant that more funds have been available for investment in other health services, than would have occurred if previous policies had remained unchanged. PMID- 10724791 TI - Diagnosis and management of osteomyelitis. Decision analytic and pharmacoeconomic considerations. AB - Osteomyelitis, or bone infection, is becoming more common, largely because of increases in the use of implanted prosthetic devices in the management of arthritis or fractures. The clinical management of osteomyelitis requires accurate microbiological diagnosis that will identify appropriate antibacterials to which the pathogenic organisms are sensitive. Therapy will largely depend on the type of bacteria, the route by which the bacteria reach the bone, the presence of any orthopaedic devices and the patient's ability to mount an immune response. Consequently, therapy often requires a combination of medical and surgical management. The aim of this review is primarily to assess the impact of different drug regimens on the total cost and to clarify the implications of various treatment options for patients with osteomyelitis. Thus, the review examines the link between the main categories of osteomyelitis and common pathogens, and provides additional comments on the aetiology and epidemiology of the condition. At present, there is a real shortage of high quality evidence to guide the decision-maker through the range of available options. One way to deal with the complexity and uncertainty surrounding the management of osteomyelitis is to develop treatment protocols leading to decision trees which will in turn systematically analyse the options available for treating patients with osteomyelitis. Consequently, we have developed a number of decision trees to show the range of options available and have applied these to the relatively simple problem of route of administration of antibacterials. However, even here the available data allow only relatively crude estimations of the costs and consequences of alternative regimens. Thus, the aim has been to provide structures that may help to set priorities for research based on the expected value of new information. In the absence of evidence, there are broadly 2 alternatives. One is based on selection of the least expensive regimen in the absence of evidence to prove that more expensive options are more effective; patients with multiply resistant staphylococci, for which no effective oral regimen is available, should be treated with intravenous therapy. This is consistent with the UK legal system, which is founded on the so-called Bolam test. The alternative is providing the maximum available treatment; in the US, it is more likely that a doctor will be held negligent for not providing the maximum available treatment, and most standard texts recommend routine use of intravenous therapy for osteomyelitis. PMID- 10724793 TI - The effect of generic competition on prescription to over-the-counter switching. AB - OBJECTIVE: While it is generally accepted that the decision to switch a drug from the prescription market to the over-the-counter (OTC) market is based on an optimization problem that firms are solving, no attempts have been made to formalize the problem. The purpose of this article is to present a model of prescription to OTC switching that helps explain the role of potential generic competition in a firm's decision to switch. In particular, we examine what market conditions are necessary for the threat of generic competition to induce switching. DESIGN AND SETTING: The model is game-theoretic, played between an incumbent firm and a potential generic entrant, and is solved for its subgame perfect equilibrium. The incumbent first decides whether to apply to the FDA to switch to the OTC market. If the incumbent declines, then the potential generic entrant has the opportunity to apply for the switch. The FDA then accepts or rejects the application, and the generic chooses whether to enter the market. RESULTS: In equilibrium, when applying to switch is costless, switching occurs if the probability that the application will be approved by the FDA is strictly positive and the OTC market is characterized by first-mover advantages. Adding a cost to the application process places restrictions on the probability of FDA approval to offset the cost of applying. The probability of approval must be sufficiently high to offset the cost of the application. CONCLUSIONS: The model shows that switching from the prescription to OTC market may occur as a response to potential generic competition. Firms switch because they know that if they do not, a generic may initiate a switch and become the first mover in the OTC market. PMID- 10724794 TI - Health economics in the Canadian pharmaceutical industry. AB - OBJECTIVE: To assess the goals, strategic focus, structure, capabilities, activities and effectiveness of health economics (HE) departments in the Canadian pharmaceutical industry, to examine how these have evolved, and the implications of HE in the future of the pharmaceutical industry. DESIGN AND INTERVENTIONS: A mixture of telephone and face-to-face interviews with members of the HE unit (survey 1) and the chief executive officers (CEOs) [survey 2] of the top 21 Canadian pharmaceutical companies was undertaken in 1997. MAIN OUTCOME MEASURES AND RESULTS: 17 out of 21 companies responded to the first survey, and 12 of the 17 CEOs responded to our second survey. The goals of the HE department in most of the pharmaceutical companies have evolved from supporting efforts to gain reimbursement on government drug plans to include pricing, promotion, internal decision-making and other activities. Members of the HE department perceive their work to be valuable to the company. The CEOs felt that the true value of HE data is not adequately understood by formulary reviewers and, therefore, HE data may be an impediment to market access. CONCLUSIONS: The purpose of the HE department is to demonstrate the value of the company's product to provincial government insurers. However, pharmaceutical companies are having difficulty justifying the importance of the HE department because of inconsistencies in the interpretation of economic evaluations by healthcare payers. PMID- 10724796 TI - Annual cost of erectile dysfunction to UK Society. AB - OBJECTIVE: The objective of this study was to estimate the annual socioeconomic burden imposed by erectile dysfunction (ED) on UK society. DESIGN AND SETTING: Health service resource use attributable to ED during 1997/1998 was obtained from appropriate databases and a panel of 22 hospital specialists comprising urologists, genito-urinary physicians, diabetologists and sexual health physicians. National unit resource costs at 1996/1997 prices were applied to the resource use data to estimate the annual National Health Service (NHS) cost of managing ED. A structured questionnaire pertaining to direct costs and absenteeism from work attributable to ED was mailed to a randomly selected sample of 5000 individuals who experience ED. MAIN OUTCOME MEASURES AND RESULTS: During 1997/1998, the annual NHS cost was estimated to be 43.9 Pounds million for managing 113,600 men with ED. Outpatient visits were the major cost driver, accounting for 65% of the annual cost. Drugs prescribed by general practitioners (GPs) accounted for a further 25%. GP consultations and penile prosthetic surgery each accounted for 4%. Tests initiated by GPs accounted for 2%, while other resources accounted for less than 1%. The annual cost was sensitive to the number of outpatient visits and, to a lesser extent, to the number of prescriptions for ED treatments, but insensitive to changes in use of the other resources. Completed questionnaires were received from 23% (n = 1141) of the sample of individuals who experience ED. From the survey, it was estimated that the NHS managed 35% of individuals with ED in the last year. Assuming this to be representative, the total potential population of individuals with ED in the UK was estimated to be approximately 324,600 men. The total population of individuals with ED (n = 324,600 men) was estimated to incur 7.0 Pounds million annually in direct costs attributable to ED and to lose 19,630 days a year from work as a result of their ED, costing society 2.2 Pounds million in lost gross domestic product. CONCLUSIONS: ED imposes a relatively small burden on UK society -53 Pounds million. Of this, 83% is borne by the NHS and 13% by patients. Indirect costs to society due to lost productivity account for the remaining 4%. The total NHS cost is strongly influenced by the number of hospital outpatient visits. Therefore, the future burden will depend largely on patients' eligibility to receive ED treatments on the NHS. PMID- 10724795 TI - Economic evaluation of long-term management strategies for erosive oesophagitis. AB - OBJECTIVE: To compare the expected costs and outcomes of alternative strategies for the management of patients with erosive oesophagitis. DESIGN: There were 3 components to the overall analytic approach. First, a decision model was constructed to compare expected costs and outcomes of 6 management strategies. Second, principles of quantitative literature review and meta-analysis were used to determine probabilities of clinical events (i.e. oesophagitis healing and recurrence). Finally, principles of cost-effectiveness analysis were used to compare treatment alternatives in terms of dominance and incremental cost effectiveness. The viewpoint for the study was that of a provincial government payer for healthcare over a 1-year period. MAIN OUTCOME MEASURES AND RESULTS: Healing rates were significantly higher for proton pump inhibitors (PPI) [p < 0.001]. Recurrence rates were significantly higher for intermittent therapy (placebo) and lower for regular dose PPI (p < 0.001). Maintenance prokinetic agent (PA) is dominated (i.e. more costly and less effective) and step-down maintenance PPI is dominated through principles of extended dominance. The 'efficient frontier' is represented by: maintenance H2-receptor antagonist (H2RA), intermittent PPI, step-down maintenance H2RA and maintenance PPI. CONCLUSIONS: The price of H2RA is a key factor influencing whether step-down maintenance PPI forms part of, or is contained within, the 'efficient frontier' of long term management for erosive oesophagitis. PMID- 10724798 TI - Acamprosate. Pharmacoeconomic implications of therapy. AB - Acamprosate is thought to reduce the craving for alcohol. The drug helps to maintain abstinence in alcohol-dependent patients who have successfully undergone detoxification. Abstinence rates during 3 to 12 months' treatment with acamprosate were approximately double those with placebo in most clinical trials, although abstinence rates were generally still < 50% in patients assigned to receive acamprosate. The drug is generally well tolerated, with the most common adverse effect being diarrhoea. In a German cost-effectiveness model, a treatment programme including acamprosate was the dominant strategy, producing a lifetime cost saving of 2602 Deutschmarks (1992 to 1995 values) per additional abstinent patient compared with treatment without acamprosate. In a Belgian pharmacoeconomic model, total direct medical costs over 2 years were 21,301 Belgian francs (1997 values) per patient lower with a treatment programme including acamprosate than treatment without acamprosate in alcohol-dependent patients. The main factors in the cost savings with acamprosate in these models were reduced costs for acute hospitalisation and rehabilitation/follow-up. The results of a cost-benefit analysis that considered both direct and indirect costs for the total alcohol-dependent population in Spain were consistently in favour of acamprosate. The lifetime net benefit for acamprosate over placebo (the incremental benefit) ranged between 61,642 million and 99,069 million pesetas (1996 values) in various scenarios with 40 to 60% of patients receiving treatment. PMID- 10724797 TI - Abciximab. A pharmacoeconomic review of its use in percutaneous coronary revascularisation. AB - Abciximab is a monoclonal antibody fragment that inhibits platelet aggregation through antagonism of glycoprotein IIb/IIIa. The drug is used in conjunction with heparin and aspirin to prevent ischaemic complications associated with percutaneous coronary revascularisation in patients with coronary heart disease. Large and well designed clinical studies have shown abciximab, as an adjunct to aspirin and heparin, to reduce by around one-third to one-half the incidence of ischaemic complications within 30 days of percutaneous coronary revascularisation. Use of the drug appears advantageous in patients at high risk, and abciximab also reduces complications in patients undergoing coronary stenting, although the drug does not appear to inhibit restenotic tissue volume within stents. Longer term benefit has also been reported, with emerging 1-year data from a study in patients at all levels of risk showing reductions in a composite end-point of death, myocardial infarction (MI) or urgent repeat revascularisation. Three-year benefit has been reported in high risk patients. Meta-analysis results, and 1-year data from patients receiving stents, have shown reduced mortality with abciximab. Abciximab therapy had an incremental cost over standard therapy from a hospital perspective of $US293 per patient (1991/1992 values) over 6 months in a prospective economic substudy from a major US clinical trial of the drug in high risk patients. Abciximab was cost saving in patients with unstable angina. A mean net cost of hospitalisation of $US476 per patients (1995 costs) has been shown in a further study in patients with a broad range of levels of risk, and observational data indicate reduced duration of hospitalisation with abciximab. Cost-effectiveness data favoured abciximab with aspirin and heparin over a 6-month period in Spanish and Dutch analyses in which data from the above trial were combined with local cost data, but not in an Australian analysis. Subgroup analyses have indicated enhanced cost effectiveness in high risk patients. Available data also show clinical benefit and cost effectiveness of abciximab therapy in conjunction with coronary stent placement. CONCLUSIONS: Data indicate intravenous bolus plus 12-hour infusion regimens of abciximab to be economically viable in patients at high or low risk of ischaemic complications after percutaneous coronary revascularisation. The drug has been shown to be cost effective in patients receiving the drug in conjunction with coronary stents, and subgroup analyses indicate additional cost effectiveness in certain groups of patients at high risk of ischaemic complications (notably those with acute MI and unstable angina). PMID- 10724800 TI - Mixed-up meds. Lots of drugs have similar-sounding names. How you can tell you're getting the right one. PMID- 10724799 TI - Salmeterol/fluticasone propionate combination product in asthma. An evaluation of its cost effectiveness vs fluticasone propionate. PMID- 10724801 TI - Orphans of AIDS. Ten million live in sub-Saharan Africa. Time asks some what they need. PMID- 10724802 TI - Doctors' deadly mistakes. Medical errors kill up to 98,000 Americans yearly; a new report says that number could be cut drastically. PMID- 10724803 TI - Doctors in training. An accident waiting to happen? PMID- 10724804 TI - The origins of disease. Did polio researchers create AIDS? PMID- 10724805 TI - The cunning plots of leadership. PMID- 10724806 TI - Children's asthma can be deadly, even when it's 'mild'. PMID- 10724807 TI - Babies and parents can safely snooze in big beds. PMID- 10724808 TI - Hospitals' reused tools: reassessing the risk to you. PMID- 10724809 TI - Is tobacco a drug? The high court faces a crucial health question. PMID- 10724810 TI - For heroin addicts, a bizarre remedy. Lack of cure spurs interest in an exotic shrub. PMID- 10724811 TI - With a splash of VOCs (volatile organic compounds). Chemicals found in many cities' ground water. PMID- 10724812 TI - Doctoring a sickly system. Deadly medical mistakes are rampant. One expert thinks they can be avoided. PMID- 10724813 TI - Showing some spine. A promising therapy for victims of paralysis. PMID- 10724814 TI - In the air that they breathe. Lead poisoning remains a major health hazard for America's children. PMID- 10724815 TI - Humility at the frontier. A comeuppance for genetic therapy. PMID- 10724817 TI - It's alive! (or might be). The science and ethics of creating life in a lab. PMID- 10724816 TI - A medieval dance craze. Was twisting a disease? PMID- 10724818 TI - For severe mental illness, a higher profile and new hope. PMID- 10724819 TI - Preventing teen suicide: it starts with straight talk. PMID- 10724820 TI - Leo Sternbach. Valium. The father of mother's little helpers. PMID- 10724822 TI - Bibliography. Current world literature. Cataract surgery and lens implantation. PMID- 10724821 TI - Wheelchair Village. Michael Sorkin wheels out plans for a sustainable retirement village. PMID- 10724823 TI - Visual outcomes with multifocal intraocular lenses. AB - Multifocal intraocular lenses (IOLs) are increasingly becoming a part of the armamentarium of cataract and refractive surgeons. Reports show that most multifocal IOLs provide excellent visual outcomes. In one major study of a zonal progressive, refractive, multifocal design, more than 80% of patients were able to see 20/40 or better at distance and J3 or better at near, without correction. This study also found that patients' satisfaction ratings of their multifocal vision were consistently high. Although a few patients with multifocal IOLs complain of halo or glare, these symptoms can be minimized by surgical technique and appropriate selection of the multifocal IOL power. PMID- 10724824 TI - The phakic intraocular lens implant: in-depth focus on posterior chamber phakic IOLs. AB - Phakic Intraocular surgery has come a long way in the past 20 years, especially in the evolution of posterior chamber phakic intraocular lenses (PC PIOLs). Clinical trials worldwide are showing acceptable results concerning efficacy, predictability, stability, and safety. PC PIOLS are proving to be a promising option for patients with high and extreme ametropia who cannot benefit from conventional corneal refractive procedures. This article provides an in-depth examination of PC PIOLs, their origin and evolution, and the results of past and current clinical studies. Reports of historical importance and studies published since the 1990s in peer-reviewed journals, textbooks, and monthly eye magazines, as well as Food and Drug Administration preliminary clinical findings, are reviewed. Anterior chamber phakic intraocular lenses are mentioned briefly. PMID- 10724825 TI - Control of intraocular inflammation associated with cataract surgery. AB - Major advances in cataract extraction techniques and instrumentation have occurred over the past decade. Smaller incisions, more efficient phacoemulsifiers, and decreased surgical times are a few of the changes that have helped to alleviate postoperative inflammation, but postoperative inflammation continues to be a cause of patient discomfort; delayed recovery; and, in some cases, suboptimal visual results secondary to cystoid macular edema. This article reviews the most recent literature regarding the control of intraocular inflammation associated with cataract surgery. PMID- 10724826 TI - Intraocular lens calculations status after corneal refractive surgery. AB - With the increasing number of keratorefractive surgical procedures, an increasing number of cataract surgeries in eyes after keratorefractive surgery is anticipated within a few decades. Although cataract extraction seems to be feasible without major technical obstacles, intraocular lens (IOL) power calculation turned out to be problematic. Insertion of the measured average K readings (= "central corneal power" = keratometric diopters) after myopic radial keratotomy (RK), photorefractive keratectomy (PRK), or laser in situ keratomileusis (LASIK) into standard IOL power-predictive formulas commonly results in substantial undercorrection and postoperative hyperopic refraction or anisometropia. In this article, the major reasons for IOL power miscalculations (which are different for RK versus RRK/LASIK) are discussed based on model calculations and based on case series of cataract surgeries, methods for improved assessment of keratometric diopters as the major underlying problem are exemplary illustrated, and finally a clinical step-by-step approach to minimize IOL power miscalculations status after corneal refractive surgery is suggested. The "clinical history method" (i.e., subtraction of the spherical equivalent [SEQ] change after refractive surgery from the original K-reading) should be applied whenever refraction and K-reading before the keratorefractive procedure are available to cataract surgeons. In addition, more than one modern third generation formula (e.g., Haigis, Hoffer Q, Holladay 2, or SRK/T) but not a regression formula (e.g., SRK I or SRK II) should be applied and the highest resulting IOL power should be used for the implant. PMID- 10724827 TI - Astigmatism and toric intraocular lenses. AB - The article reviews some of the basic optics of astigmatism and the correction of astigmatism with cylindric lenses. A simple model of the conoid of Sturm is demonstrated, and an ideal position for the conoid is postulated. The orientation of the conoid shows that leaving patients with some simple myopic "against-the rule" astigmatism is beneficial to near work, whereas "with-the-rule" astigmatism is beneficial for distant viewing. Surgeons should be less aggressive with patients with with-the-rule and against-the-rule astigmatism and more aggressive with oblique astigmatism. The toric intraocular lens (IOL) should be positioned on axis or, if slightly off axis, err on the side away from the vertical or horizontal meridian so that the resultant cylinder is more vertical or horizontal. Clinically significant rotation of the toric IOL occurs in a few cases, but these can be easily rerotated. Rerotation should be done between the first and second weeks after primary implantation. PMID- 10724828 TI - Intraocular lens implantation in children. AB - Over the past 15 years, lens implant surgery in children has disseminated so much that it is no longer a controversial issue. It has become rather a specialized topic in the widespread field of lens implantation in the general population. To match the excellent results seen in adults, issues such as the surgical technique, the choice of the lens, and dioptric power of that lens, are constantly being refined and adapted to children's growing eyes. Scleral tunnels and small, self-sealing corneal incisions are being replicated in children to benefit from their advantages. Polymethyl methacrylate material remains unrivalled from the point of view of safety and longevity in the human eye. Intraocular lenses (IOLs) with an overall diameter of 12 mm can safely be used in nonmicrophthalmic eyes of children more than 3 years of age. Several investigators now recognize the need for smaller pediatric IOLs for neonates, toddlers, and microphthalmic eyes. Fortunately, modern IOLs are smaller today than they were 15 years ago. The accumulating evidence on the myopic shift that occurs in pseudophakic children have led to an almost unanimous agreement that the IOL power should aim for a certain amount of hypermetropia at time of surgery. The residual refractive error can be corrected with spectacle glasses that are adjusted throughout childhood. The goal is to start with hypermetropia in childhood that will convert into emmetropia or mild myopia in adulthood. PMID- 10724829 TI - Posterior capsule opacification. AB - Posterior capsule opacification (PCO) is the most common complication following primary cataract surgery. Advances in intraocular lens (IOL) designs that have reduced the amount of PCO following surgery have been made. The understanding of how the IOL design effects PCO has also advanced. Lenses that provide a mechanical barrier between it and the posterior lens capsule seem to inhibit PCO to a greater degree. Intracapsular rings are now being explored to test and enhance this barrier effect. Major advances in the elimination of lens epithelial cells at the time of surgery especially by pharmacologic means have also been made. An immunotoxin specific for human lens epithelial cells shows promise and is under latter phase clinical development. PMID- 10724830 TI - Cystoid macular edema following cataract surgery. AB - Cystoid macular edema (CME) remains a troublesome problem after cataract surgery and other types of ocular surgical procedures. It is recognized as the most frequent cause of decreased vision in patients following cataract surgery. Although the disease was first described more than 40 years ago, its cause is unclear, and all available therapeutic interventions, mainly based on theories regarding the pathogenesis of the condition, are of doubtful effectiveness and are still far from being satisfactory. Most published literature on the incidence and treatment of CME consists of small, retrospective case series and cannot provide reliable answers as to whether a given factor or intervention is associated with the occurrence or outcome of the disease. PMID- 10724831 TI - Anesthesia modalities for cataract surgery. AB - Research articles on anesthesia modalities for cataract surgery are reviewed. A growing trend toward the use of topical anesthesia is apparent. Particular emphasis in the literature is given to determining the safety and efficacy of various forms of topical anesthesia using injectable anesthesia as a frame of reference. A consensus of opinion points toward the use of topical application of anesthetic drops plus intracameral unpreserved lidocaine 1% as the anesthesia modality that provides the best level of analgesia and comfort to patients while not compromising ocular safety. Several articles reporting complications of injectable anesthesia are also reviewed. PMID- 10724832 TI - Piggyback intraocular lens implantation. AB - The piggyback method of implanting two intraocular lenses in one eye has been successfully expanded to address pseudophakic refractive error in normal eyes and eyes that have undergone post-penetrating keratoplasty. Piggyback implantation has been combined with the use of newly available minus-power lenses to provide appropriate power for a cataract patient with keratoconus, as well as to correct pseudophakic myopia. The phenomenon of increased depth of focus in piggybacks may be explained by a contact zone between the lenses. The late complication of inter lenticular cellular growth with resultant hyperopic shift, opacification, and loss of vision has recently become a concern. PMID- 10724833 TI - To see ourselves. Radiology's place in the world of medicine. PMID- 10724834 TI - Soft-copy viewing calls for new way of thinking about images. PMID- 10724835 TI - Seekers of quality in radiology submit report from the trenches. PMID- 10724836 TI - Breast MRI proves worth but lacks standardization. PMID- 10724837 TI - In pediatrics, CR may become the ideal detector. PMID- 10724838 TI - MR a catalyst for improving women's health. PMID- 10724839 TI - Radiology's role widens in therapeutic drug development and trials. PMID- 10724840 TI - Radiology up against financial limits on its growth. PMID- 10724841 TI - Web-savvy patients may know more than you think they do. PMID- 10724842 TI - Effective Web site can educate patients and attract new ones. PMID- 10724843 TI - Physician groups in conflict over vascular centers. PMID- 10724844 TI - Forum highlights imaging's role in fight against cancer. PMID- 10724846 TI - Medical Informatics Europe '99. Conference proceedings. PMID- 10724845 TI - Imaging helps identify who benefits from stroke intervention. PMID- 10724847 TI - Lesson learnt from a halt of the hospital information system. AB - The present paper deals with an experience of blackout of the hospital information system at our Medical Centre, trying to evaluate the impact on internal users (physicians, nurses, clerks) and patients population. Limited inconveniences have occurred to out-patients in terms of delay in collecting medical reports after diagnostic test execution. As regards direct users, impact was evaluated through a structured interview. Administrative personnel, that have been using computer-based system for at least ten years, have not declared particular inconveniences, accepting the overtime or the extra-work as simply unavoidable. On the contrary, health-care personnel reported a heavy negative impact of the system failure on their activity. After a great effort to achieve the system acceptance and direct physicians usage, the blackout of the system has pointed out that the situation has changed since a few years ago: now the HIS, and particularly its clinical core, is considered mission critical. PMID- 10724848 TI - Selection of hospital information systems: user participation. AB - Motivated by economic and quality management issues, many hospitals strive for the introduction of information systems into the clinical environment. To introduce the most suitable software product, a systematic selection procedure is required involving all affected staff. The paper describes a methodology that guides the clinical users to an objective motivated decision. This semi-formal methodology is necessary in view of missing standard models for the description of hospital information systems and leads to high acceptance in practical use. The methodology consists of structured product demonstrations with a quantitative assessment, benefit-value-analyses based on step weights for the functions needed, and test installations. Prerequisite of this procedure was the definition of objectives and priorities in a standard catalog that was used as reference in the whole project. Selecting a software product following this systematic procedure establishes a sound basis for a successful application in the clinical environment. PMID- 10724849 TI - The use of the Balanced ScoreCard (BSC) in the model for investment and evaluation of medical information systems. AB - This paper describes the use of the Balanced ScoreCard (BSC) in the MIEMIS meso model (Model for Investment and Evaluation of Medical Information Systems). The scope of the MIEMIS model is to integrate the evaluation process into the whole lifecycle of an information system using both a prospective and a retrospective approach. We conclude, that the MIEMIS-model has benefited from implementing the BSC into the model due to the fact, that the BSC can support the project management work. This approach helps ensuring, that the new information systems are fulfilled according to the plan and with a balance between the four perspectives (financial, customer/user, internal, and innovation/learning perspective) to avoid that the financial aspect is the driving force in developing and implementing a new information system, for example. PMID- 10724850 TI - HIS purchase project: preliminary report. AB - The KAGes (Krankenanstaltenges.m.b.H) is a company, owned by the state of Styria in Austria, which operates 20 hospitals with about 8,000 beds and 14,000 employees, serving a population of ca. 1.2 million people. KAGes is on its way to purchase a new hospital information system (HIS) for it's hospitals. Within the strategic IT plan and the "System Structure New" (SSN) project a methodology was developed, for making an effective HIS purchase. Several steps of this project are described in the paper: request for product information, evaluation of vendor proposals, product presentations, test site evaluation, reference site visits and selection of vendor finalists. The authors present the internal project management methodology, including the structure of the project team, project information management through intranet, criteria for different steps of the evaluation and evaluation site organization. Four major HIS vendors with leading HIS products qualified for this stage of the project (evaluation site). About 60 teams with 400 members (end users and IT-experts) have assessed all the products installed, during one or more, repeated, test sessions. The decision on which new HIS to purchase will be based on the recommendations derived from this evaluation. Two pilot installations (one general hospital and one teaching hospital department) are planned for implementation in the year 1999/2000 and subsequent roll-out (including substitution of the legacy system) is scheduled until the year 2003. PMID- 10724851 TI - Metabolic drug pointing and information processing. AB - Computer supported drug design is based on the biochemical information for the prediction of alternative bio-chemical pathways. Molecular information on genes, proteins, biochemical reactions, mutations, inborn errors and metabolic diseases are available via internet. Based on this information we developed an information retrieval and processing concept combining knowledge about metabolic diseases and biochemical effects. PMID- 10724852 TI - The survey of healthcare software in Yugoslavia. AB - The aim of this study is to identify vendors of software packages applicable to health care system as well as to evaluate software packages offered to the health sector by a variety of sources, namely state owned software development companies, privately operated software development companies as well as sole traders. The approach undertaken was to preselect the potential suppliers via an initial targeted interview. Subsequently a questionnaire, consisting of 23 questions, was addressed to the selected vendors and suppliers. The response rate to the questionnaire was 58%. Investigation of 113 has revealed the non government software suppliers clearly lead in copyright ownership, IBM compatible personal computers were the platform of choice whereas majority of code was written utilising languages capable of supporting relational data base queries. The final outcome of the above study is a database of software packages and applications available to the health sector offering the health professionals and institutions guide lines and support in developing information systems, relevant subsystems and modules. PMID- 10724853 TI - Developing a health care information system for Scotland. AB - The rapid development and uptake of Internet technology has created opportunities for large-scale information networks to replace paper-based information sources. In order to obtain the maximum benefits for patients and medical practitioners it is important that health care providers work together to produce an integrated information service. However, the task of bringing together a large number of different information providers to create a huge structured pool of information covering a wide range of topics with appropriate quality assurance is non trivial. This paper describes an approach being used to create healthcare information systems in a set of co-operative healthcare information networks in Europe, and specifically in Scotland. PMID- 10724854 TI - Paediatric case-mix of the Lorraine regional database in 1994 and 1995. AB - In the region, there is a need to predict the necessary number of paediatric beds. Paediatric discharge data from the 1994-95 Lorraine regional database of Anonymous Discharge Summaries (ADS) are used for this study. We analyse the concentration and specialisation of the hospital activity types and we estimate the numbers of paediatric beds needed for the year 2,000. For the same percentage of the case-mix by hospital group, concentration of the activity is more important in the small District General Hospital (DGH) than in the Teaching Hospital (TH). In the most important Medical Diagnosis Categories (MDC), the case mix variation by age class can't be characterised inside of the MDC group. The different methods we use to estimate the need of paediatric beds, give similar estimations for the year 2,000. PMID- 10724855 TI - Introduction of a new hospital information system at the Innsbruck University Hospital. AB - The aim of the introduction of an advanced hospital information system at the Innsbruck University Hospital is mainly to support the care process and to improve quality and efficiency of care. These requirements entail changes in the clinical processes and many obstacles with respect to the organization have to be considered. The article gives an overview of the goals, organization and problems of the introduction which is still in the planning phase. PMID- 10724856 TI - The Health Management Information System of Romania. AB - In Romania a Health Management Information System (HMIS) project is in progress. The project covers the main activities of the Ministry of Health (MoH), of the 42 District Health Authorities and more than 250 other health care units. The first applications will be implemented at the middle of 1999. The paper shows the context of the HMIS among other Health Information Systems in Romania, its main parts and some organizational issues of the HMIS project, achievements and difficulties encountered. PMID- 10724857 TI - National information system on hospital treatment. AB - National information system on hospital treatment (HOSTREAT) supports health management system applied by the Ministry of Health and its professional services at the Public Health Institute of Slovenia. The main project objective was data quality, having in mind advanced analytical methods such as knowledge discovery in databases. The step was a huge one, as the related national statistics rests on every single hospital event, called an episode. By the project definition, an episode covers therapeutic treatment of one sick or injured person in one hospital within one health service specialty. So, many problems arose within the three years of project phases. Much research work was to be done on programming tools and the information system itself. This paper deals with five strategic problems and corresponding corrective actions. PMID- 10724858 TI - An IT awareness programme for the health sector in Ireland. AB - This paper describes an Awareness Programme delivered throughout Ireland which aims to increase the level of understanding of healthcare professionals as to the benefits of Information and Communications Technology (ICT) and of emerging trends in Health Informatics in Europe. The programme examines the use of ICT across the whole health sector--primary, secondary and tertiary. It has been delivered at over 30 centres to more than 1500 health professionals. PMID- 10724859 TI - Slovenian national health insurance card: the next step. AB - The Slovenian national health insurance company started a full-scale deployment of the insurance smart card that is at the present used for insurance data and identification purpose only. There is ample capacity on the cards that were selected, to contain much more data than needed for the purely administrative and charging purposes. There are plans to include some basic medical information, donor information, etc. On the other hand, there are no firm plans to use the security infrastructure and the extensive network, connecting the insurance company with the more than 200 self service terminals positioned at the medical facilities through the country to build an integrated medical information system that would be very beneficial to the patients and the medical community. This paper is proposing some possible future developments and further discusses on the security issues involved with such countrywide medical information system. PMID- 10724860 TI - MedicDAT--making information available. PMID- 10724861 TI - Transformation of the process of administrative patient management. AB - The risks involved with performing a process transformation project are significant. The success rate can be improved if a formal process with concrete guidance is used. This paper introduces an object-oriented approach to process transformation and information system development. The proposed approach affords adequate freedom, encourages innovative thinking and points the way to the desired goals. Its main aim is to combine different activities into a uniform process with the use of object-oriented modelling. PMID- 10724862 TI - National health care providers' database (NHCPD) of Slovenia--information technology solution for health care planning and management. AB - INTRODUCTION: The article describes the possibilities of planning of the health care providers' network enabled by the use of information technology. The cornerstone of such planning is the development and establishment of a quality database on health care providers, health care professionals and their employment statuses. MATERIAL AND METHODS: Based on the analysis of information needs, a new database was developed for various users in health care delivery as well as for those in health insurance. The method of information engineering was used in the standard four steps of the information system construction, while the whole project was run in accordance with the principles of two internationally approved project management methods. Special attention was dedicated to a careful analysis of the users' requirements and we believe the latter to be fulfilled to a very large degree. RESULTS: The new NHCPD is a relational database which is set up in two important state institutions, the National Institute of Public Health and the Health Insurance Institute of Slovenia. The former is responsible for updating the database, while the latter is responsible for the technological side as well as for the implementation of data security and protection. NHCPD will be inter linked with several other existing applications in the area of health care, public health and health insurance. Several important state institutions and professional chambers are users of the database in question, thus integrating various aspects of the health care system in Slovenia. CONCLUSIONS: The setting up of a completely revised health care providers' database in Slovenia is an important step in the development of a uniform and integrated information system that would support top decision-making processes at the national level. PMID- 10724863 TI - A unified model to support an information intensive health care environment. AB - The two parts of this paper respectively analyses the mismatch between current systems and practice in health care, and present an outline design for enhanced Integrated Care Pathways (ICP) knowledge and information management based upon mature ICT technologies. PMID- 10724864 TI - Human aspects of medical computer application. AB - Most of the research activities in the field of medical informatics are directed toward technological-technical problems. Human factors of computer use are often a neglected topic. Even the application of the tiniest microcomputer embedded in a medical diagnostic equipment may pose user acceptance and motivation problems but to mention larger scale hospital information systems. In this paper the author presents some aspects of the human-computer interaction problems in a medicinal computer application setting. PMID- 10724865 TI - An architecture for bridging between research and practical use in health informatics. AB - Health informatics is a true interdisciplinary research discipline combining computer engineering, health science and fields dealing with organisation and communication issues. Much of the research published within the area has a common goal: to develop optimal information systems for better access to relevant data, information, and knowledge in the health care sector. As a respond to existing discontinuity between research and practical use, we present an architecture for an interdisciplinary virtual organisation promoting synergy across health informatics environments including research, industry, clinical, and educational settings. A set of key lessons learned from a practical implementation of the architecture are reported. PMID- 10724866 TI - The bridge between administrative and clinical information system. AB - On one side, physicians are asked to record administrative information, such as activity measurement, case-mix of their specialty, billing, for statistical, legal or reimbursement purposes; and on the other side, they need to gather detailed information about their own patients in terms of clinical evolution, for the day-to-day care of the patients or for clinical research purposes. Many other actors are also involved with these processes, both on the administrative side, such as registration officers, administrators and on the clinical side, nurses and other care providers. Applications have been developed within hospital information systems for capturing and disseminating information according to these specific actors and dedicated purposes. But more and more appears the need to integrate these data for insuring the coherence of information and avoiding redundancy of data capture. How to conciliate these objectives? We describe the Geneva's approach for integrating the administrative and the clinical systems. PMID- 10724867 TI - Research needs and priorities in health informatics--early results of a Delphi study. AB - A four-phased Delphi study has been performed on the topic of "research needs and priorities to implement the Information Society within Healthcare". This contribution presents the outcome of the first three phases. The biggest surprises are that 'Telemedicine' is relatively lower ranked than expected, and that 'Business Process Re-engineering' is the highest ranking topic, as judged from the number of issues and barriers raised by the expert panel. PMID- 10724868 TI - Success factors of hospital information system implementation: what must go right? AB - Health care institutions are considering a variety of information technologies in the hope of increasing efficiency, reducing cost, re-engineering work processes, and improving quality of care. When we talk about any information system implementation, we need to keep in mind that there are actually three phases to such a project: pre-implementation, implementation and post-implementation. The aim of our contribution is to present the realities of successful implementation and our experience with such project. PMID- 10724869 TI - How to structure clinical information: a practical example. AB - In the frame of an European telemedicine project, we have designed and implemented an Andrology server that provides the user (e.g. physicians, students) with a set of anonymized andrology cases. Within this context we faced the problem of re-structuring existing clinical information sources and gained some experience on how much of the problem might be solved by the software engineer. This paper presents a method that partially automates the maintenance of the andrology case collection. Moreover, the potential benefit of XML for information representation will be discussed. PMID- 10724870 TI - The community social alarm service as a primary health care service. AB - The history of calls to the community social alarm centre in Ljubljana and the service subscribers' records were analysed with the aim of finding out how the service helps in emergencies when calls related to health problems are received. The network, being a pilot project in Slovenia, was established in 1992 primarily to offer social services to elderly and disabled people living in their own home environments. The article highlights the role of the network in solving the medical problems of the users of the system. Among the 18,500 calls received in four years at the centre, 395 (2.1%) of them were health-related. An additional 24 emergency calls (0.2%) were received from non-subscribers to the centre using an "ordinary" telephone. Although the absolute number of health-related calls is low, the network plays an important role in preventive primary health care aimed at the observed population groups. The network meets most of the specific technical and organisational requirements for a community health care emergency call system. From a telematic point of view it is important that a telematic device--an emergency call unit--has entered an elderly or disabled person's home and been well accepted. PMID- 10724871 TI - Psychiatry by videophone: a trial service in north west England. AB - In this paper we report on the use of a video link between two general practices and a hospital based mental health team in North West England to provide a trial telepsychiatry service for individuals with depression and anxiety related disorders. Patients (n = 16) took part in an evaluation of the service by both structured questionnaire and semi-structured interview. The results of the evaluation study suggest that patients may be highly critical of telemedicine systems and that they do so not simply on the grounds of the technical quality of video links, but also because the remote link increases the difficulty that the patient faces in expressing deep scated emotional and existential problems. It is not, therefore, simply a matter of technical quality in the link, but also a question of the quality of interpersonal relations perceived by the patient. PMID- 10724872 TI - Telepathology and imaging spectroscopy as a new modality in histopathology. AB - Telemedicine started in the late 1950's by transmitting data on patients' pulse and heart rates. In the 1980's it expanded to radiology and orthopedics. The technology is now expanding to other specialties that can digitally gather patient data. Telepathology comprises the transmission of microscopic images via telecommunication network. Image compression and multiplexing technologies enabled high-resolution telepathology as well as real-time video consultations over international telephone lines. Organ transplantation has become a viable treatment and offers new life to an increasing number of patients suffering from chronic end stage diseases and from irreversible organ failure. Rejection is still a major problem in kidney, liver, and heart transplantation. To gain further insight into the complex interactions within the components of the immune system, it has become increasingly necessary to develop rapid and simple methods to monitor the status of the immune system in patients. Clinical signs suggest organ rejection and abnormal laboratory test results, but only histological signs on biopsy specimens are adequately specific. The financial cost of organ transplant makes it imperative to develop tools for the early identification and treatment of organ rejection. An increasingly sensitive and accurate way of localizing key structures and abnormalities is through spectroscopy of either H&E stained samples or with a fluorescent tag (fluorophore) or by relying on natural fluorescence. The system is based on a unique Prism and Mirror Imaging Spectroscopy System ("PARISS), spectrometer originally designed and implemented for remote Earth monitoring from space and aircraft and astronomical imaging spectroscopy. Compact and lightweight both the mirror and prism are presently constructed in inexpensive glass but can also be injection molded in plastic. Any number of vendors anywhere in the world can produce all parts of the assembly. This greatly enhances the chances of future commercial viability. The Interactive Histopathology Consultation Network INTERPATH (PL961121) project integrated of remote control imaging microscopy system, imaging spectroscopy, and communication networks called SPECTROMIC. This telepethology unit will be a useful tool in the Regional and International Integrated Telemedicine Network for Medical Assistance in End Stage Diseases and Organ Transplant, RETRANSPLANT HC 4028 (HC) & IN 4028 (HC). PMID- 10724873 TI - Transtelephonic ECG in Slovenia. AB - Transtelephonic ECG has been introduced in Slovenia in 1996. It has been operating on a regular basis since September 1997. Users of the system are health institutions and patients with CV diseases. They use portable 12-channel ECG weighing less then 200 grams. Diagnostic center is located in medical ICU in Clinical Center Ljubljana. In the first 20 months of regular operation we have received 852 calls, 450 of them being diagnostic. The most common reasons for calling were: ischaemic chest pain, atypical chest pain, palpitations and dyspnoea. Most common diagnoses made on the basis of history and ECG were: AMI, angina pectoris, paroxysmal tachycardias and atypical chest pain. In 186 cases the cardiologist's advice sufficed, 171 patients were referred to the ER, 142 immediately and 29 only if suggested therapy hadn't been successful. To 82 patients new medication or changed dosage of previous medication was suggested. With the intent to evaluate accuracy of the diagnoses made on the basis of history and transtelephonic ECG evaluation we followed up the patients who were sent to ER in Ljubljana upon our cardiologists' advice. On the basis of preliminary results we can conclude that TTE is as diagnostic as the conventional ECG and its' use shortens time from onset of symptoms to initiation of treatment, it lowers disability and mortality due to CV diseases and improves cost benefit. PMID- 10724874 TI - Southern Health Board--advanced telematic/telemedicine in healthcare services in the south west of Ireland. AB - The Southern Health Board has been involved in Telematic Healthcare services for the past number of years. Our involvement in a number of EU funded projects has given us extensive experience and expertise in this area. The primary focus of our telematic applications is optimising health services delivery with a particular emphasis towards people who are isolated due to social and physical boundaries and limitations. Our main projects are:- NIVEMES: (Network of Integrated Vertical Medical Services) includes the exchange of medical information between urban centres and isolated communities including those at sea, rural, elderly and disabled persons. RISE: (Remote Information Services for the Elderly) focuses on the enhanced healthcare services of the elderly and disabled through the use of telemedical applications. HEALTHLINE: To strengthen the successful outcome of Telehealth implementation by specific projects in the EU by providing the expert Training and Information dissemination services to providers, beneficiaries and the general public, using the Telehealth systems and infrastructure applied by those projects. KATE: (Killarney & Telecom Eireann) the SHB in conjunction with the Information Age Town Project in Killarney are working towards benefiting the local community through the innovative use of Information and Communication Technologies. TASTE: (Technology Assessment in Telemedicine) An EU funded trans-national initiative aimed at developing a methodology for assessing medical technology, particularly in the area of Neurosurgery. FECV: (Forward Emergency Control Vehicle) An European Space Agency project through which we are developing an advance communications vehicle for major emergence/event management through Satellite linkage to SHB systems. PMID- 10724875 TI - Using IP-videoconferencing systems in a surgery consulting. AB - In this paper we describe how to use IP-videoconferencing systems in medical surgery consulting. We started to think about how we could use special doctor's services without patients having to travel a long way. The answer to this question is that the information goes from one place to another, not the patient. First we had a pilot project, where we used the 3xISDN transmission rate and now we are using ATM. We have here in Satakunta a local area network between our Satakunta Central Hospital and the Health Care Center in Noormarkku and Kankaanpaa, so we have very good environment to do this kind of research. Our network is quite fast, we can use the 10 Mbps bitrate and in this network there are no other activities in this moment, so there are not any interferences. There is a surgery specialist in the hospital and a doctor in the health care center with a patient. The specialist looks at the monitor, where there is a videopicture of the patient from the health care center. Then the specialist makes the treatment plan for the patient. PMID- 10724876 TI - Teleconsultation for endoscopic diagnosis of gastrointestinal diseases--concepts and architecture of the service ENDOTEL. AB - This paper introduces ENDOTEL as a new project in telemedicine, aiming to reduce local disadvantages in medical health care in rural areas. The progressing specialization in medicine results in a concentration of experts in few medical centers and to deficits of expert knowledge in rural areas, which remain as a domain of unspecialized physicians. ENDOTEL expands the conventional synchronous telemedical communication, which usually means videoconference, by an asynchronous multimedia communication and its resulting advantages. ENDOTEL breaks up with the need that participants have to be present simultaneously. PMID- 10724877 TI - Electronic consultations and referrals in Kymenlaakso Hospital District. AB - The meaning of this paper is to introduce the medical network which has been taken to Kymenlaakso Hospital District and the primary experiences in utilizing. Actual numeral results are not ready because of short time. First results will be ready in August 1999. PMID- 10724878 TI - The hip status: a telemedical application. AB - Biomedical informatics and biomedical engineering are interdisciplinary sciences that are growing fast and have a significant impact on healthcare delivery. The University Medical Centre in Ljubljana and the Medical Faculty through its institutes have initiated the building of their own telemedicine systems. These systems concern only the medical informatics field. However, in this work a telemedical application that uses a combination of biomedical engineering methods and methods of biomedical informatics is presented. This telemedical application was built at the Department of Surgery and it was named "The Hip Status Application". Its aims are to provide services of preventive medicine and health promotion for people with potential problems with the hip-joint, guidance for people that have already underwent an operation of implantation of hip endoprosthesis, or have in any other way affected hip-joints, remote system consultation for diagnostic purposes, etc. The application is still under evaluation, and it is meant as a start of a new era of engineering-powered telemedical applications at the Department of Surgery following the trends in the most developed countries of the World. PMID- 10724879 TI - The telemedicine and second medical opinion. AB - Telemedicine is declared as delivery of care to patients anywhere in the world by combining communications technology, with medical expertise. The well established applications of telemedicine are in the field of teleradiology, pathology, cardiology. In some cases video-conferencing (VC) may be applied. Second medical opinion is an important additional viewpoint on the patient diagnosis and treatment. The established network of consultant hospitals and worldwide Second opinion centers enable a fast (24-72 hours response) second medical opinion consultation service with the telemedicine application. The consultant hospitals at present time are: UCSF--San Franscisco, Stanford Medical Center, Bowman Gray School of Medicine, The Brigham Radiology Foundation, Emory University Hospital, University of Pennsylvania Medical Center. The second opinion centers are located worldwide, just to mention some of them: London, Budapest, Ljubljana, Tel Aviv, Bombay, Athens, Milano, Seoul. PMID- 10724880 TI - Computerized system for the networking of health services (Proposed solution for the informational support of the health insurance law in Romania). AB - The integration of health services in the health insurance system can only be performed by using an advanced informational system, due to the large volume of documents needed for the payment of medical services by the health insurance firms. To better facilitate the access to the medical databases throughout the country is it necessary that the different application and their respective supports are able to communicate with each other. Every component of the health system (doctors' offices, clinics, hospitals, pharmacies, county health insurance offices) consist of a local LAN/WAN network according to the individual computer capabilities. These networks are connecting through the Internet which can be describe as a network of networks that permits long range communication. PMID- 10724881 TI - Hospital information systems: chances and obstacles on the way to integration. AB - Today, most large hospital information systems (HIS) comprise many heterogeneous application systems, loosely coupled via interfaces. Integration could offer many advantages, like the support of interdepartmental workflows or a consistent patient record. This cannot be achieved, however, by simply interfacing originally unrelated component systems. In this paper, we present relevant concepts of integration, the autonomy/consistency tradeoff, and technical alternatives in some detail. Based on our recent experiences in analyzing an existing HIS and selecting adequate vendors for the needs of our 1250-bed hospital, we then discuss strategies and tradeoffs on the way to a HIS that utilizes state-of-the-art technology to fill the users' needs. PMID- 10724883 TI - Real-time medical applications and telecommunications. AB - Telecommunications play an important role in telemedicine. Many forms of telecommunication services based on different telecommunication technologies are developed for various needs. The paper deals with complex real-time applications which demand high telecommunication requirements. At the beginning, medical applications are categorised and real-time applications qualified as multimedia applications. Requirements for multimedia elements are listed separately. Later on, short introduction of related telecommunication protocols is given. Real-time medical applications can show their ability in case of guaranteed quality of services delivered by telecommunication network as it is explained in the end. PMID- 10724882 TI - End-user point of view to establish real-time broadband ATM connection over Europe. AB - There have so far been few discussions about how the international broadband services will be created in practice. In this paper we concentrate on two separate international telemedicine trials in which Satakunta Central Hospital took part in the HIM project. In these trials we tested real-time video consultation over the international point-to-point ATM network connection between hospitals in Finland, Portugal and Germany. The quality of the consultations were clinically first-rate in both trials. The technological lesson learned from these trials was valuable for the operators in establishing broadband connections in Europe. The network connection was analysed based on transmission capabilities of the network connection. PMID- 10724884 TI - Barriers for dissemination of a national health care data network. A list of recommendations for better dissemination. AB - Establishment of a Danish Nation-wide Health Care Data Network for exchange of health care related information using standardised EDIFACT messages is agreed between the Danish Government, the Hospital Owners and the GPs organisations. The aim is, within year 2000 in the MedCom project, to reach a level of 66% of all messages between the secondary and primary health sector must be send electronically using EDIFACT. There are many barriers for reaching this goal, and to reveal this barriers an investigation was done, and a list of recommendations are set up. The investigation was based upon literature, interviews and most important a questionnaire among 200 GPs and specialists all over the country. After 6 months where the recommendations have been handled there is seen an increase in communicated messages on 40%, and from January 1999 more than 1.1 mill messages are exchanged each month. PMID- 10724885 TI - Regional health care networks. AB - Communication cross primary and secondary care, between organisations or between different types of professionals causes special problems. The communication often implies exchange of data between different computer systems. The INCO-COPERNICUS project PRIMACOM (PRIMAry Care Physician's COMmunication Network) demonstrates how such communication can be established in regions in Hungary and Slovenia. PRIMACOM is building on European messaging standards for EDI (Electronic Data Interchange) with practical experience from the Danish MedCom project. The paper points out the need for standardising health care messages and via the Danish MedCom project documents the return of investment. PMID- 10724886 TI - Teleradiology as a seed of the regional health care Intranet in a rural region in Spain. AB - Castilla y Leon is a rural region with a very sparse population of about 2,560,000 habitants. Special health care challenges appear due to this region's very aged population and large dimensions. A theoretical model of services of Telemedicine is proposed and an experimental teleradiology trial is carried out between a rural Health Care Centre and its Reference Hospital to make easier the planning of future projects. The total number of studies was increased due to availability of specialised support and higher skills of physicians, but the percentage of studies consulted to specialists diminished. Continuing education through physicians communication was highly improved. PMID- 10724888 TI - Third generation electronic pharmacy communications. Recommendations based on ten years' experience. AB - The number of electronic prescriptions in Denmark is increasing and almost 600,000 electronic prescriptions are now sent every month. There is therefore a strong interest in directly reusing the data entered in the medical practitioner systems in the pharmacy systems. It has therefore been decided to introduce a shared data foundation in all medical practitioner and pharmacy systems in Denmark in conjunction with the introduction of the new MEDPRE electronic prescription standard. Parallel to the standardisation work, concrete guidelines have been prepared for the individual EDP (Electronic Data Processing) systems to ensure rapid, problem-free prescription communication. The aim in the longer term is for 95% of all EDIFACT prescriptions to reach the receiving pharmacy no more than 20 minutes after being sent from a medical practice. PMID- 10724887 TI - Teledermatology--UK experience of setting up an integrated teledermatology service. AB - Telemedicine has traditionally been seen as an all or nothing solution to the problem of providing medical service to remote communities. In the UK, however, the majority of the population lives within easy travelling distance of medical service, and it is inappropriate to apply telemedicine for the same reasons and in the same way. We describe an alternative approach, where telemedicine is used to complement the existing clinical infrastructure of the outreach clinic and hospital visit, and the patient may opt to have any follow up appointments by video conference. We also describe the concept of the virtual consultation that we have developed to support our work. PMID- 10724889 TI - Telemedicine in the Malaysian Multimedia Super Corridor: towards personalized lifetime health plans. AB - The Malaysian Telemedicine initiative advocates a paradigm shift in healthcare delivery patterns by way of implementing a person-centred and wellness-focused healthcare system. This paper introduces the Malaysian Telemedicine vision, its functionality and associated operational conditions. In particular, we focus on the conceptualisation of one key Telemedicine component i.e. the Lifetime Health Plan (LHP) system--a distributed multimodule application for the periodic monitoring and generation of health-care advisories for all Malaysians. In line with the LHP project, we present an innovative healthcare delivery info-structure -LifePlan--that aims to provide life-long, pro-active, personalised, wellness oriented healthcare services to assist individuals to manage and interpret their health needs. Functionally, LifePlan based healthcare services are delivered over the WWW, packaged as Personalised Lifetime Health Plans that allow individuals to both monitor their health status and to guide them in healthcare planning. PMID- 10724890 TI - Communication security in open health care networks. AB - Fulfilling the shared care paradigm, health care networks providing open systems' interoperability in health care are needed. Such communicating and co-operating health information systems, dealing with sensitive personal medical information across organisational, regional, national or even international boundaries, require appropriate security solutions. Based on the generic security model, within the European MEDSEC project an open approach for secure EDI like HL7, EDIFACT, XDT or XML has been developed. The consideration includes both securing the message in an unsecure network and the transport of the unprotected information via secure channels (SSL, TLS etc.). Regarding EDI, an open and widely usable security solution has been specified and practically implemented for the examples of secure mailing and secure file transfer (FTP) via wrapping the sensitive information expressed by the corresponding protocols. The results are currently prepared for standardisation. PMID- 10724891 TI - Can a database be anonymous? AB - Sensitive data are most often indirectly identifiable and so need to be rendered anonymous in order to ensure privacy. Statistical methods to provide anonymity require data perturbation and so generate data processing difficulties. Encryption methods, while preserving confidentiality, do not require data modification. PMID- 10724892 TI - A matix-approach proposal for the treatment of contents to be integrated into multimedia health resources for citizens. PMID- 10724893 TI - A secure Web-based interface to maternity records with decision support. AB - The recent developments in Web technology now make it easy to create user friendly interfaces to databases and link them to the Internet or to local Intranets. With the acceptance of this new approach one can take advantage of the opportunity to use this development to incorporate additional knowledge into the interface. Not only can this provide a more supportive interface, but it can also create the opportunity for sharing and comparing such knowledge in the future. This paper describes how this has been used to develop an interface to a database of maternity case records which incorporates a knowledge base of rules relating to care plan protocols and uses this to provide decision support to the carer. The issue of security is a serious problem and some aspects of the measures taken are discussed. PMID- 10724894 TI - Personal identifying number as a unique patient identifier in database on clinically treated patients in Belgrade: its use, advantages and drawbacks. AB - Database on clinically treated patients in Belgrade served as an example for analysis of possibilities for the use of Personal Identifying Number (PIN) as an Unique Patient Identifier. In the first part of the paper we analyzed filling up of the fields which contained data on PIN within complete databases in 1981, 1991 and 1996. Filling up of PIN was significantly changed in the three observed years: it was 18% in 1981; 68% in 1991, and 56% in 1996 respectively. Analysis of interactions among the chosen factors (type of hospital, demographic and social characteristics of patients, length of stay in hospital, manner of treatment, main diagnosis, treatment outcome) and measuring time, showed a different degree of statistical significance. In the second part of our paper we analyzed the unexpected decrease in filling up of PIN in 1996 (as compared to 1991) ussing the method of logistic regression, on 1% samples from the databases for the two respective years. On the basis of obtained models of filling up of UPI data, taken as dependent variable and the above factors (predictors) we analyzed the advantages and drawbacks of UPI application as an unique patient identifier. PMID- 10724895 TI - A simulation of dynamic tasks routing to improve cooperation in intensive care units. AB - Cooperation between Health Care Professionals is essential for the quality of care. Workflow systems could improve the transfer of informations and responsibilities within Health Care Actors. We have proposed a conversation-based Workflow in order to modelize the therapeutics plan in the ICU. In such a complex field, the flexibility of the workflow system is essential for the system to be usable. We have introduced some dynamicity by adding roles in the model. With the use of roles, the dynamicity of the workflow is assumed by the routing process. We need to use simulation to be able to study the impact of routing algorythms on the efficiency of the coordination. PMID- 10724896 TI - Experiences with a new security standard for healthcare information systems. AB - This article describes the results of the implementation and demonstration of the Standard CEN ENV 12924 (Security Categorisation and Protection of Health Care Information Systems), that was performed as part of the ISIS/MEDSEC project of the EU. The categorisation scheme given in the standard was followed through for almost all information systems or sub-systems in the Leiden University Medical Centre. The status of the security measures was evaluated for ten systems; further implementation plans were then drawn up for these systems, and partly effectuated. Findings are reported, both on the present security level, and on the applicability of the standard (which in general was found to be very positive). In the course of this work, use was made of a database support tool, developed in an earlier EU project (SEISMED). PMID- 10724897 TI - Authorization & security aspects in the middleware-based healthcare information system. AB - The integration and evolution of existing systems represents one of the most urgent priorities of health care information systems in order to allow the whole organisation to meet the increasing clinical organisational and managerial needs. The CEN ENV 12967-1 'Healthcare Information Systems Architecture'(HISA) standard defines an architectural approach based on a middleware of business-specific common services, enabling all parts of the local and geographical system to operate on the common information heritage of the organisation and on exploiting a set of common business-oriented functionality. After an overview on the key aspects of HISA, this paper discusses the positioning of the authorization and security aspects in the overall architecture. A global security framework is finally proposed. PMID- 10724898 TI - Bias in meta-analysis and funnel plot asymmetry. AB - International experiences reveal the important role played by scientific research and systematic study a problems, in effectively tackling change in the health sector. Meta-analysis was introduced to address the problem of synthesizing the large quantity of information on a particular subject. It is viewed, only as a step in the process of developing better tools to quantify information across studies. The selection of trials for inclusion in a meta-analysis may be biased if selection is restricted to published trials, to trials published in English language journals, to trials published in prestigious journals or to trials cited by other authors. Funnel plot is graphical display of sample size plotted against effect size for the studies included in a meta-analysis, which can be used to investigate bias. When all the studies have been located, the distribution of points should resemble a funnel. If there are gaps in the funnel shape it indicates that some studies may have not been published or located. In evaluating bias, we use meta-analysis studies about radiotherapy alone versus combined radiotherapy and chemotherapy in stages IIIa and IIIb non-small cell lung cancer. A simple analysis of funnel plots provides useful test for the likely presence of bias in meta-analyses, but as the capacity to detect bias will be limited when meta-analyses are based on a limited number of small trials the results from such analyses should be treated with considerable caution. PMID- 10724899 TI - Developing shared understanding through simulation. AB - In this article relevant key findings of a review of selected published work are used to provide a foundation for a new approach to developing a shared understanding of aspects of health care delivery. The new approach makes use of simulation models. It is hoped that this approach will provide support for improved decision-making and for the development of more appropriate enabling tools. PMID- 10724900 TI - Data integration to assist gastric emptying data analysis. AB - The diagnosis of dyspepsia is very difficult because the symptoms are clinically aspecific and the gastric emptying time tests are of complex interpretation. An integrated and automated analysis of clinical and instrumental data may improve the diagnostic process. We present a system to collect data on dyspeptic patient from different sources which has been set up to assist the clinician in the diagnosis of dyspepsia. The data base integrates a wide set of symptoms with data coming from laboratory tests. Moreover, we assess the feasibility of classifying gastric emptying profiles using both octanoid acid excretion data and electrogastrography. PMID- 10724901 TI - Three-dimensional ROIs in brain PET. AB - A semi-automatic system for determining volumes of interest (VOI) from positron emission tomography (PET) scans of brain is described. The VOIs surface extraction is based on user selectable threshold and three-dimensional target flood-fill. Contrast to anatomical volume detection approaches, volumes are determined from functional PET images and the obtained objects are checked against anatomical images. The developed VOI program was evaluated with brain FDOPA-PET studies where the striatum was the object. The results were comparable to entirely manual method and the target extraction time is reduced to about one third of manual method. PMID- 10724902 TI - A computerised study on the results of in-vitro-fertilisation. AB - This project has been concerned with the comparison of children born as a result of fresh embryo transfer (IVF) with children conceived and born via the natural process. The former included children who were the single outcome of the birth (singletons) and children who were the result of a multiple birth (e.g. twins, triplets). A computer database was established into which was put 23 items of data on each child, making a total of 12,788 items overall. There were 278 "normally conceived" children (controls), 150 IVF Singletons, and 128 children from multiple births. The results show interesting differences in the gestational age at birth, the birth weight, the mode of delivery and the degrees of birth abnormalities and malformations. PMID- 10724903 TI - Uniforming of outpatient health care statistics. AB - In the process of developing the health care information system, the Public Health Institute of the Republic of Slovenia harmonized the outpatient health care statistics at the national level. This paper presents the goals, contents, methodology and a brief analysis of the new data structure. The main functions of the ZUBSTAT computer program, user training methods, and planned further activities in this field are described. PMID- 10724904 TI - Application of space technologies for valuation of a stress level. AB - The adaptation of large long-term experience of application Heat Rate Variability in space biology and medicine and medical researches is represented. The description of the new approaches is given which are fixed in a basis of diagnostic computer systems "NeuroResercher 5.0" (or System of Computer Vegetology "CardioTensionTest 5.0" as module in the frame of "NeuroResercher 5.0") and "Varicard". These systems for thorough clinical and experimental scientific investigations and experimental investigations (for specialists of R&D institutes, doctors of functional diagnosis rooms of hospitals possessing clinical bases of R&D institute and medical university chairs, during clinical and preclinical testing of drugs, medical and biological R&D institutes carrying out Nneurophysiological investigations in the field of labour medicine and sport medicine, development of infantile nervous system, medical education establishments. PMID- 10724905 TI - An efficient algorithm for automatic decoding of ECG signals. AB - Creating of computer ECG-conclusion, based on decodings of ECG-signals, as a rule includes three main stages-morphological analysis, comparison of obtained parameters and synthesis the conclusion. In a considered circuit the most critical point is the first stage. Here even the minor errors of automated, i.e. computer determination of elements, can provoke synthesis incorrect conclusions, in particular automated measurement of heart rate variability [1]. From this point of view it is extremely important to creat algorithms of decoding which are stable against parasites of electrical and biological origin, and also which are capable adequately to react to signal polymorphism [2]. Research in this direction have revealed the following. PMID- 10724906 TI - Frequency domain methods for measurement of heart rate variability. PMID- 10724907 TI - A signal analysis method for impulse-like eye movements. AB - Eye movement investigations are performed in several fields of medicine. We have developed signal analysis methods of eye movements studied in otoneurology for several years. Previously, we have designed methods for eye movement types of saccades, nystagmus, smooth pursuit, and vestibulo-ocular reflex. In this paper we extend and develop further our method to concern also impulse-like eye movements, i.e. a new type of eye movement tests for investigations of vertiginous patients. According to our preliminary results the use of our method is efficient to differentiate patients with inner ear diseases from normals. PMID- 10724908 TI - Lossless compression of eye movement signals. AB - Compression of digital signals is widely employed in several areas such as speech signals. Concerning biomedical signals almost merely ECG signals have so far been compressed, in fact, in very many applications. To our knowledge, this compression study is the first one presented in the case of eye movement signal which are investigated in several medical specialties. In this study eye movements are electro-oculographically recorded. Tests performed to signals of patients of an equilibrium laboratory gave compression ratios generated with essential lossless compression methods. PMID- 10724909 TI - A prototype of an information system for assessing the health status of prison inmates. AB - A research-action program was established in 1996 between the Loos-Lez-Lille prison psychiatric unit and the Department of Medical Informatics of the University Hospital of Lille (France):--(1) to investigate the health status and the general characteristics of the prison population--(2) to develop an Information System for improving the prison health care and to facilitate social rehabilitation of convicts. Starting off 1988, all new prisoners are interviewed on their arrival using a standard questionnaire. The transfer of all the information recorded in this questionnaire into a computer base was initiated in 1996, when the research action program began. A statistical analysis was performed on 15,200 records (1989-1995) to identify the most informative parameters: 50% of inmates were less than 24 years old; 57% were unemployed; 60% had no professional qualification. 31% of inmates had a psychiatric history and 16% had made a previous suicide attempt. The rate of drug abuse has increased from 24% in 89 to 53% in 95. To analyze the time trends of these parameters, a prototype of Information System was then developed. The system uses the database to product standard reports in real time. PMID- 10724910 TI - Use of time-variant coherence as a general tool for analysis of interrelations between brain electrical processes. PMID- 10724911 TI - Computer EEG-monitoring of laserotherapy effects in patients with asteno depressive syndrome. AB - Nowadays the low-intensive laserotherapy is shown to be an effective and non hazardous method of asteno-depressive syndrome treatment. The differences of EEG reactions to laser influences have been revealed in patients of different age groups. And the close negative correlation between the therapy effect, on the one hand, and the patient's age and the disease duration, on the other hand, has been shown. No significant changes of the patient's state or integrative EEG-indices have been evoked by a placebo application. The results showed the advantages of the low-intensive laserotherapy in asteno-depressive syndrome treatment and confirmed the significance of computer EEG-monitoring for prediction, control and correction of the state of the patient. PMID- 10724912 TI - Parametric time-varying spectrum and its application to SEMG signals. AB - Frequency spectrum of surface electromyographic signals (SEMGs) exhibits a non stationary nature even in case of constant level isometric muscle contractions. This changes are related to muscle fatigue processes and can be evaluated by methods for estimation of time-varying (TV) spectrum. We describe a two estimation techniques: sliding window approach and decomposition of linear model coefficients trajectories. Drawback of the first is its large variance which causes unrealistic abrupt changes in the curve of overall dynamics, calculated either as the second derivative of the MF or as the ARMA distance between subsequent linear models forming the TV parametric spectrum. We demonstrate, that the second method is more appropriate for description of a slowly changing spectrum of surface EMG signals. Dynamics measures obtained from this estimation are proposed as an indication of the fatigue process character. PMID- 10724913 TI - Monte Carlo simulation of the radiographic imaging procedure for electronically designed phantoms. AB - This paper presents a software simulation package of the entire X-ray projection radiography process including beam generation, absorber structure and composition, irradiation set up, radiation transport through the absorbing medium and image formation. This software tool is an augmented version of a previously presented system with expanded range of application by the implementation of Monte Carlo approach for the simulation of the radiation interaction at the absorber and the detector. Phantoms are created as composite objects from geometrical or voxelized primitives and can be subjected to simulated irradiation process. The acquired projection images represent the two dimensional spatial distribution of the energy absorbed in the detector and are formed at any geometry, taking into account energy spectrum, beam geometry and detector response. PMID- 10724914 TI - Computed tomography image analyzer: segmentation applying active contour models- "snakes". AB - Many diagnostic and therapeutic procedures depend on medical images. In order to overcome imperfections of obtained images which are due to acquisition process and to obtain new information from available images, many techniques have been developed. In this study relatively new method of image segmentation, active contour model--"snakes" was applied in analyzing computed tomography (CT) images in patients with acute head trauma. Using this method, lesion to brain (LBR) and ventricle to brain ratio (VBR) were obtained accurately. Quantitative variable LBR, is significantly higher in patients with other pathologic CT findings and who do not survive during hospitalization. Thus, by applying segmentation "snakes" model it is possible to extract maximum information from available CT scans. These variables could be basis for medical decision making in patients with acute head trauma. PMID- 10724915 TI - A new way for the analysis of the exocytosis. AB - Exocytosis is the most common way for cells to secrete substances. This is a wide distributed phenomenon that involves all cell types and all animal species from yeast to human. Catecholamine (CA) release from adrenal chromaffin cells occurs by the exocytosis of secretory vesicles also called chromaffin granules. Individual secretory events can be easily monitored by the use of carbon fibers encased in glass microelectrodes. Catecholamines oxidized at the electrode tip produce a transient electrical current wave called secretory spike. The typical secretory spike consists in a sharp elevation of current followed by a rapid exponential decrease to the basal level. A 35-50% of the spikes showed a pre spike wave called "foot" which represents the CA release through the fusion pore. The time course of secretory spikes observed is altered by extracellular phenomena's like diffusion. It has been suggested that amperometric spikes can be described by the convolution of a Gaussian with a decreasing exponential. The exponential function is partially governed by the diffusion of the secreted substances towards the electrode. In this article a method to deconvolve both functions is proposed. This mathematical approach allows the observation of the original secretory profile--the Gaussian--without the distortion caused by the diffusional broadening of catecholamines. PMID- 10724916 TI - Knowledge discovery for advanced clinical data management and analysis. AB - Knowledge discovery is a broad research field in which methods are developed to support discovery of novel and potentially useful knowledge from clinical databases and registers in systems for patient care. However, the techniques available are not readily applicable in medical domains, due to, among other reasons, low user friendliness and lack of proper methodological background. Data mining approaches to be explored and improved are predictive modelling, segmentation, dependency modelling, summarization, and change and deviation detection/modelling (in data or knowledge). Another and original contribution of the research is to build up efficient feedback loops. Human experts and available domain expert systems could provide suggestions as how to improve all major steps in the knowledge discovery process such as evaluation of knowledge, choice of data mining methods and data input. A long tradition of collecting and maintaining clinical and administrative data could be found in fields of oncology, cardiology, coronary surgery, social and primary health care medicine. All these areas, that gather data over long periods of time, could benefit from knowledge discovery. PMID- 10724917 TI - Application of the medical data warehousing architecture EPIDWARE to epidemiological follow-up: data extraction and transformation. AB - In this paper, we present an application of EPIDWARE, medical data warehousing architecture, to our epidemiological follow-up project. The aim of this project is to extract and regroup information from various information systems for epidemiological studies. We give a description of the requirements of the epidemiological follow-up project such as anonymity of medical data information and data file linkage procedure. We introduce the concept of Data Warehousing Architecture. The particularities of data extraction and transformation are presented and discussed. PMID- 10724918 TI - Statistical approaches for evaluation of genetic risk factor of peripheral arterial occlusive disease. AB - In the paper we show results of medical study from statistical point of view. The medical study was aimed to study genetic risk factors of peripheral arterial occlusive diseases in Czech population. Two genes, CBS and MTHFR were examined, as various genotypes of these genes are thought to have impact on amino thiols, who participate in variety of reactions in vasculature. Statistical part of the study was responsible for analysis and interpretation of collected data. PMID- 10724919 TI - ArchAIDS connects a clinical department and a virological laboratory to automatize the follow up of AIDS patients. AB - A communication system for the automation of the follow up of AIDS patients set up by DIST at the Molecular Virology Unit in the Advanced Biotechnology Centre of Genova and at the Department of Internal Medicine of the Medical School of Genova is presented. This system includes a distributed database to store both clinical and virological data and a set of procedures to transfer patient data with a complete respect of requirements about completeness and privacy. PMID- 10724920 TI - Data warehousing as a basis for web-based documentation of data mining and analysis. AB - In this paper we present a case study for data warehousing intended to support data mining and analysis. We also describe a prototype for data retrieval. Further we discuss some technical issues related to a particular choice of a patient record environment. PMID- 10724921 TI - Treatment of missing values with imputation for the analysis of otologic data. AB - Usefulness of imputation in the treatment of missing values in an otologic database was studied. Missing values were filled in with means (ME), regression (LR) and Expectation-Maximization (EM) imputation methods. A random imputation method (RA) provided baseline results. ME, LR and EM methods agreed on 41-42% of the imputed missing values. The level of agreement between these and RA method was 20-22%. Despite the moderate agreement, discriminant functions were similar and accurate (prediction accuracy 83-99%) for each diagnosis. A lot of data were missing in otoneurotologic tests which have less weight in the diagnosis of vertiginous patients. Consequently, the disagreement of the methods did not affect discriminant analysis. Inputation seems to be a useful method to treat missing data in this database, but future research requires more complete data and advanced imputation methods. PMID- 10724922 TI - Data warehousing as a tool for quality management in oncology. AB - At present, physicians are constrained by their limited skills to integrate and understand the growing amount of electronic medical information. To handle, extract, integrate, analyse and take advantage of the gathered information regarding the quality of patient care, the concept of a data warehouse seems to be especially interesting in medicine. Medical data warehousing allows the physicians to take advantage of all the operational data they have been collecting over the years. Our purpose is to build a data warehouse in order to use all available information about cancer patients. We think that with the sensible use of this tool, there are economic benefits for the Society and an improvement of quality of medical care for patients. PMID- 10724923 TI - Naive Bayesian-based nomogram for prediction of prostate cancer recurrence. AB - This paper introduces a schema with naive-Bayesian classifier and patient weighting technique to develop a prostate cancer recurrence prediction model from patient data. We propose the graphical presentation of naive-Bayesian classifier with a nomogram, which can be used both for prediction or can provide means to data analysis. The resulting model was experimentally evaluated; the results were favorable both in terms of interpretability and predictive accuracy. PMID- 10724924 TI - Increasing the diversity of medical data mining through distributed object technology. AB - Data mining has been recently experiencing a boom of interest from researchers and software producers. In medicine, however, its applications are still rather rare. In this paper, we argue that this is primarily due to the requirements of reproducibility of results and diversity of available data mining tools, both of which are crucially important for medical research. We propose to tackle the diversity requirement by means of distributed object technologies. The results presented here rely on our experience with medical data mining using the method GUHA and with further developing that method. PMID- 10724925 TI - Outpatient health care statistics data warehouse--logical design. AB - Outpatient Health Care Statistics on a state level is a central point where all relevant statistic data are collected from various sources from all over the country. Various and complex requirements for processing and reporting data makes Outpatient Health Care Statistics on a state level a perfect example for efficient implementing of Data warehouse technology. The research investigates logical design of data warehouse with a special attention on a different data modeling technique in various phases of a logical data warehouse design. The research shows that a requirement for processing statistical data determines the design decision and thus the scope and semantic value of final data warehouse. PMID- 10724926 TI - Applying data mining in healthcare: an info-structure for delivering 'data driven' strategic services. AB - Presently, there is a growing demand from the healthcare community to leverage upon and transform the vast quantities of healthcare data into value-added, 'decision-quality' knowledge, vis-a-vis, strategic knowledge services oriented towards healthcare management and planning. To meet this end, we present a Strategic Knowledge Services Info-structure that leverages on existing healthcare knowledge/data bases to derive decision-quality knowledge-knowledge that is extracted from healthcare data through services akin to knowledge discovery in databases and data mining. PMID- 10724927 TI - Quantitative data analysis for exploring outcomes in cardiac surgery. AB - The article focuses on possibilities of statistical knowledge exploration to predict outcomes of surgical treatments. The outcomes were defined in relation to the measured peri- and intraoperative data, as well as follow-up patient questionnaire. Clinical consequences are expected in terms of a smaller data set with a better ability to predict the surgery outcomes and a better cost performance. The important questions that could discriminate quality of life (QoL) were: Relief from surgery?, Has cardiac surgery effected earlier symptoms? Work capacity? Consultations after the surgery? The performed data analysis proved to be efficient in the complex data set that was collected. Pain relief was identified to be significant, while relations between measured blood laboratory profile and later QoL were weak. PMID- 10724928 TI - An information system on toxicological risks linked to drug manipulation. AB - Aim of the project is to define and to implement an information system able to help people, who produce chemotherapic antineoplastic drugs, and health care operators, who manipulate such drugs, to prevent short and long term adverse effects connected to the above mentioned activities. The system is able to give a detailed and updated information about these problems, and to give an up-to date, from a professional point of view, to the users of the system. Also an information system prototype was implemented, which consists of an object oriented database, a decision support system, able to manage and to plan a periodic control of workers, to verify the adverse effects of the antineoplastic chemotherapic drugs, a management system for the network communications. PMID- 10724929 TI - Post-Doc: satisfying the information needs of general practitioners in continuing medical education and daily practice. AB - The learning needs of General Practitioners (GPs) are not optimally covered by traditional ways of Continuing Medical Education (CME). The European-funded Post Doc project sets up a model for a European internet-based infra-structure and learning environment to support GPs' individual CME activities, focusing on problems of daily work. It integrates education, information and communication for both daily work and learning at home. Based upon a user requirements analysis, the service includes multimedia training applications, selections of relevant information sources, a calendar of events, and communication facilities. The service is intended to fit in existing health care structures and accreditation systems. Because of different languages and user requirements in Post-Doc's regions each region builds an own website. The regional prototypes are built and evaluated in cooperation with small core user groups. This regional approach proved feasible and also influenced the network structure with regional demonstrator websites and one master website including central functionality. PMID- 10724930 TI - The interactive use of networking multimedia--innovative education resource for professionals and patients. AB - The combination of new and rapidly developing interactive multimedia computers and applications with electronic networks will require a restructuring of our traditional approach to strategic planning and organizational structure. Worldwide telecommunication networks (using satellites, cable) are now facilitating the global pooling of healthcare information and medical knowledge independent of location. The development of multimedia information and communication systems demands cooperative working teams of authors, who are able to master several areas of medical knowledge as well as the presentation of these in different multimedia forms. The assemblage of telematics and services offers a base for multimedia applications, for example teleteaching, telelearning, telepublishing, teleconsulting, teleconferencing, telemedicine etc. The expansion of the internet will also lead to the formation of interdisciplinary "Global Education Networks". The theory and practice of education are undergoing dramatic changes. Lifelong learning and adaptation of medical practice to new knowledge and new techniques will be even more important in the future. PMID- 10724931 TI - WWW as a teaching resource for introductory medical informatics in primary care. AB - This pro and contra report assesses the current state of the WWW as a teaching aid for Medical Informatics, using the experience gained from the delivery of a module to a heterogeneous group of health workers enrolled in an MSc/PgDip in Primary Care. The students were expected to act as both information consumers in a directed bibliographic retrieval task and information providers for Primary Care. The students' perceptions as to the usefulness and local relevance of current resources on the world wide web (WWW) are discussed. PMID- 10724932 TI - The electronic learning zone in health informatics. AB - This paper outlines developments in health informatics education arising from the rapid development of the Internet and the growing importance of lifelong learning. The drivers for change in educational delivery are examined and examples of related uses of the Internet are given. The development of the Electronic Learning Zone at the University of Surrey is described and the paper concludes with the probable future extensions to support lifelong learning. PMID- 10724933 TI - Medical informatics in the medical curriculum--when? AB - Undergraduate teaching of medical informatics (MI) at the Medical School University of Zagreb has been changing its position in the medical curriculum. Nowadays, this subject is positioned in the second year of the medical curriculum. The aim of this paper is to find out what the students of the second year (S2Y) and of the sixth year (S6Y) think about the position of MI in the medical curriculum. The results of students' opinions have shown that MI should be positioned: in the last two years (63% of S2Y, 36% of S6Y), in the middle (11% of S2Y, 21% of S6Y), and at the beginning of the medical curriculum (26% of S2Y, 43% of S6Y). However, analysing this question according to previous experience of the students of both groups, it can be concluded that: the S6Y with more previous experience shift the subject towards the end of the medical curriculum, and the S2Y with more previous experience shift the subject towards the beginning of the medical curriculum. According to our long-term experience and students' opinion, the most relevant MI subjects, any kind of medical application, should necessarily come at the end of the medical curriculum. PMID- 10724934 TI - Communication between the higher education and industry. AB - The paper presents the starting premise and the general frame of the Tempus Project CME-02555-96, entitled "Know How Transfer form University to Industry". It is intended to be a study for establishing a frame and methods for communication between academic, scientific and research bodies and industry. This project continues the efforts made in the RENEHEMA Tempus project aiming to support the improvement of communication between higher education and industry. Following the conclusions of the RENEHEMA conference the strategy adopted by the management board focused on the Internet technology and free market mechanisms to fulfill the project objectives. PMID- 10724935 TI - CISMeF: cataloque and index of French speaking health resources. AB - In 1999, the Internet has become a major source of health information. The objective of CISMeF is to catalogue and index the main French-speaking sites and documents concerning health. Currently, the number of resources already totalled over 6,100 with a mean of 75 new sites each week. CISMeF contains a thematic index, including medical specialities and an alphabetic index. CISMeF uses two standard tools for organising information: the MeSH (Medical Subject Heading) thesaurus from the Medline bibliographic database (National Library of Medicine) and the Dublin Core metadata format. A brief description of the site is systematically added. CISMeF respects the Net Scoring, criteria to assess the quality of health information on the Internet. The CISMeF project fulfils a valuable tool for the French-speaking health community: 2,500 machines visit the Web site each working day. PMID- 10724936 TI - Medical informatics educational tasks seen from practical perspective. Tempus Phare Project CME-02555-96. AB - This paper tries to synthesize the discussion of a seminar on medical informatics educational tasks held in May 1998 in Sinaia, Romania, within the frame of the Tempus-Phare Project CME-02555-96 entitled "Know How Transfer from University to Industry" and coordinated by the University of Medicine and Pharmacy Timisoara, Romania. Special emphasis was paid to particular features of medical education requirements in East European countries, with particular reference to Romania. PMID- 10724937 TI - Outpatient health care statistics data warehouse--implementation. AB - Data warehouse implementation is assumed to be a very knowledge-demanding, expensive and long-lasting process. As such it requires senior management sponsorship, involvement of experts, a big budget and probably years of development time. Presented Outpatient Health Care Statistics Data Warehouse implementation research provides ample evidence against the infallibility of the above statements. New, inexpensive, but powerful technology, which provides outstanding platform for On-Line Analytical Processing (OLAP), has emerged recently. Presumably, it will be the basis for the estimated future growth of data warehouse market, both in the medical and in other business fields. Methods and tools for building, maintaining and exploiting data warehouses are also briefly discussed in the paper. PMID- 10724938 TI - An orthopaedic discussion group, linking and teaching the orthopaedic community. AB - Since its inception, the Internet has created many new opportunities for the dissemination of knowledge between individuals. One of the areas most affected by these changes has been healthcare, both in terms of service provision and in the arena of medical education. Other healthcare professionals have described the educational resources available on the Net but few have illustrated the advantages of educational and conversant interaction between members of a surgical speciality or the maturing process that takes place within that group. We describe how the initiation and continued development of an Internet-based discussion group has benefited the orthopaedic profession in Ireland. We identify problems detected to date and possible difficulties in the future of the discussion group. PMID- 10724939 TI - Education and training possibilities in Europe for chaining ambulatory and hospital inpatients care. AB - Although there is a trend to the development of the electronic patient record, the need for uniform classification systems to document diagnoses and procedures is still there. In the European Union, some research projects or concerted actions promote the concept of episodes of care, meaning that the clinical status of a patient might be followed by different health professionals in various health care settings (ambulatory care, emergency, inpatient, long term care) through medical record summaries. An inventory of teaching and training tools related to information systems that classify diagnoses in various health care settings has been made during the year 1998. Its results show that casemix tools are mainly used for hospital inpatients, much less for outpatients and almost never yet to link episodes of care. Computerised training tools exist in several countries for coding as well as for grouping patients but this area appears still too knew in the field of education because of a lack of European health policy to use uniform data sets in order to measure the degree of efficiency as well as the quality of health care delivery systems in the European union country members. PMID- 10724940 TI - Globalization and the cultural impact on distance education. PMID- 10724941 TI - The case-based Internet textbook ODITEB for multi-modal diagnosis of tumors- development, features and first experiences. AB - Internet technologies offer the chance to build high-quality learning media for the education in medicine. In particular, the teaching of diagnostics with medical imaging can be supported by the excellent visualization and interaction capabilities. In cooperation with three radiological departments at German universities in Munich, Erlangen and Wurzburg the distributed case-based Internet textbook ODITEB for tumor diagnosis of the GI-tract, liver, pancreas and thorax has been developed. It offers a growing collection of didactically prepared cases located on servers at the provider sites Munich, Erlangen and Wurzburg, functionality similar to a real CT console, original DICOM data, X-rays and endoscopic and endosonographic videos, and expert-guided tours through the cases. In a first evaluation in summer 1998, 32 medical students graded the application with 1.9 ('good') on a scale from 1 ('very good') to 5 ('very bad'). PMID- 10724942 TI - From bridges to super-highways: transmitting meaning within and between professions, and across time and space--beginning the process. AB - As development of health informatics, including electronic patient records, proceeds apace there is an innate tendency to focus on acute and primary care, and upon bio-pathological data sets. This is where virtually all research and investment is being directed. However, the core purpose of health care (and mental health care in particular) is to improve and maintain the individual's functioning and sense of well-being, not simply to eliminate adverse pathology. It is therefore vital for health care records to contain subjective, descriptive, and self-expressed components if the record is to have true health meaning. This in turn raises challenges about meaning and context, terms and language. Most informatic systems run the risk of being Islands of Automation, linked at best by bridges conveying data sets rather than knowledge. If health informatics is really to serve people and their health, attention needs to be given to developing the recording, communication, and understanding of perception through shared meaning. Only then will informatics systems be full supporters of the people's health, and record system linkages become Super-Highways of Knowledge between patients and their supporting professionals. PMID- 10724943 TI - Are healthcare professionals really prepared to deal with information-empowered citizens? AB - The new Global Information Society increasingly incorporates electronic tools in every walk of life. The e-communication revolution should not be welcomed just because it exists, but rather taken as a possible catalyst to better performance, however that is perceived. Healthcare practitioners must harness the good will & motivation of citizens to play proactive parts in the health & lifestyle management for themselves, their families & their communities. Citizens must recognise that they do not become 'experts' overnight & must work with professional practitioners to the benefit of health status per se. There are still a number of substantial challenges to be faced & a need for considerable parallel & collective development if we (professionals & public) are to contribute effectively together in healthcare in the future. PMID- 10724944 TI - An Internet implementation of an international clinical study. AB - This paper describes a clinical study that is being conducted in 15 countries world-wide via the internet. The study concerns the removal of Factor VIII inhibitors in haemophilic children. The suitability of the Internet for this type of study is explored together with the technology (Active Server Pages, Secure Sockets Layer) that is involved in the creation of such a system. PMID- 10724945 TI - Accessing a WWW reference library of 3D models of pathological organs to support medical education. AB - A major aspiration of the medical community is to use multimedia databases to disseminate important research or clinical information and for education. We describe a WWW reference library of 3D models of human organs containing pathological and normal organs organised in cases, together with a variety of educational and supporting tools. Although the emphasis is chiefly on the teaching of medical students and the continuing education of physicians, the library is also a potentially-valuable resource for diagnostic investigation. This paper describes the environment, methodologies and tools for supporting 3D models in medical education. The prototype of the system is currently being completed; the full system is scheduled for public access in the year 2000 through multimedia publishers. PMID- 10724946 TI - Sharing electronic medical record on the WWW using InterCare architecture and smart cards. AB - Southern-Finland is a partner of the European Commission funded telematics project, InterCare, which is developing and demonstrating the WWW--based regional architecture and application integrated with smart cards. The InterCare project will combine the results from the most important EC funded healthcare telematics projects in their respective areas--Cardlink 2, Hector, Star, Synapses and TrustHealth. The primary objective is to achieve a convergence of results and to demonstrate & promote the synergy between projects from different areas of the healthcare telematics programme. PMID- 10724947 TI - The need for evaluation when managing the IMIA.ORG web-site. AB - The International Medical Informatics Association (IMIA) has built up a web-site to support international scientific exchange and facilitate organizational tasks. Regular monitoring is required to get information on whether the site is actually used and by whom. Main aspects of the evaluation are function, structure and contents. As main evaluation methods the logfile analysis and user questionnaires are used. The number of visits to IMIA's web-site has constantly increased in the last year. In January 1998 the site had 418 visits, in December 1998 there were 6002 visits. The user questionnaire showed that the web-site offers an adequate platform for the members. It is concluded that the members as the main target group are reached by the service and in addition that the growing number of non members require further development of the public part of the site. PMID- 10724948 TI - WWW search engine for Slovenian and English medical documents. AB - The information tool for the organization and searching of Slovenian and English medical documents is presented. The tool, partly still in development phases, performs automatic subject description of documents, searching with natural language queries and rankig of search hits according to their relevance. The search engine allows the searcher to use relevance feedback in order to perform incremental improvement of search results. The machine learning system TILDE for learning user profiles was also applied. Documents marked by the user as relevant or non-relevant are used to find characteristics that distinguish relevant documents from non-relevant ones. PMID- 10724949 TI - Implementing HISA standard in the middleware layer of our applications. AB - This article describes our experiences in implementation of the European standard for the architecture of healthcare information systems. The standard was a great help, but this article focuses more on some problems we have encountered. PMID- 10724950 TI - Analysis of free MEDLINE on the WWW. AB - In the paper, a survey of free MEDLINE access on the WWW is given. We compared five different vendors that provide free unlimited access to MEDLINE. The comparison of different search parameters was made: number of search statements, number of search fields, ability to combine search statements, number of search limits. The possibilities of using MESH thesaurus during the search are also explored. At last, we compared and evaluated the results of two search queries, obtained with searching MEDLINE, provided by five vendors mentioned above. PMID- 10724951 TI - Organization and dissemination of multimedia medical databases on the WWW. AB - In the paper, we focus on the problem of building and disseminating multimedia medical databases on the World Wide Web (WWW). The current results of the ongoing project of building a prototype dermatology images database and its WWW presentation are presented. The dermatology database is part of an ambitious plan concerning an organization of a network of medical institutions building distributed and federated multimedia databases of a much wider scale. PMID- 10724952 TI - GIN AUSTRIA. Assuring quality and relevance on Internet-health-informations for patients. AB - GIN AUSTRIA (Gesundheitsinformationsnetz AUSTRIA) offers patients and consumers reliable medical knowledge about diseases, wellness and disease management in an understandable way and enables them to quick and incessant access to informations about the Austrian health system and Austrian health organizations. To improve the quality of the database and to achieve full customer (patients, citizens) satisfaction a systematic approach for implementing total quality management is also applied. Focusing the attention on understanding and responding to customer needs, systematic and continuous improving of the IS and total involvement of all participants are the three core TQM principles at this project. The second focus of the project is the development and the implementation (prototype) of a medical dictionary or rather medical thesaurus as interface for patients, who are not used to scientific terms and expressions. This interface is based on the controlled vocabulary of the MeSH-Thesaurus (german version). PMID- 10724953 TI - Generalisation and extension of a web-based data collection system for clinical studies using Java and CORBA. AB - Inadequate informatical support of multi-centre clinical trials lead to pure quality. In order to support a multi-centre clinical trial a data collection via WWW and Internet based on Java has been developed. In this study a generalization and extension of this prototype has been performed. The prototype has been applied to another clinical trial and a knowledge server based on C+t has been integrated via CORBA. The investigation and implementation of security aspects of web-based data collection is now under evaluation. PMID- 10724954 TI - Evaluation of medical knowledge using electronic textbooks and ExaMe program on Internet. AB - In the paper we describe the function of the ExaMe program that serves for evaluation of students knowledge using Internet. Evaluation is based on the knowledge base of a given course. Two types of evaluation tests are described. The fixed test is appropriate for examination of students by the teacher in computer classroom connected to Internet. By this test the evaluation is done in the limited time period and all students are tested with the same evaluation test. The automatic test is appropriate for self-evaluation and self-study by students on remote places. The student can ask the ExaMe program for the test of different difficulty levels and state the number of questions required. Finally the first experience with the ExaMe program in practice is given. The application of the ExaMe program is shown in connection with IT EDUCTRA electronic products and electronic textbook on biomedical statistics. PMID- 10724955 TI - A JAVA implementation of a medical knowledge base for decision support. AB - Distributed decision support is a challenging issue requiring the implementation of advanced computer science techniques together with tools of development which offer ease of communication and efficiency of searching and control performance. This paper presents a JAVA implementation of a knowledge base model called ARISTOTELES which may be used in order to support the development of the medical knowledge base by clinicians in diverse specialised areas of interest. The advantages that are evident by the application of such a cognitive model are ease of knowledge acquisition, modular construction of the knowledge base and greater acceptance from clinicians. PMID- 10724956 TI - Java-based framework for the secure distribution of electronic medical records. AB - In this paper, we present a Java-based framework for the processing, storage and delivery of Electronic Medical Records (EMR). The choice of Java as a developmental and operational environment ensures operability over a wide-range of client-side platforms, with our on-going work emphasising migration towards Extensible Markup Language (XML) capable Web browser clients. Telemedicine in support of womb-to-tomb healthcare as articulated by the Multimedia Supercorridor (MSC) Telemedicine initiative--which motivated this project--will require high volume data exchange over an insecure public-access Wide Area Network (WAN), thereby requiring a hybrid cryptosystem with both symmetric and asymmetric components. Our prototype framework features a pre-transaction authentication and key negotiation sequence which can be readily modified for client-side environments ranging from Web browsers without local storage capability to workstations with serial connectivity to a tamper-proof device, and also for point-to-multipoint transaction processes. PMID- 10724957 TI - European and Latin-American countries associated in a networked database of outstanding guidelines in unusual clinical cases (ELCANO). AB - The aim of ELCANO, an EU-funded project, is to build a virtual multilingual multimedia library of unusual clinical cases related to gastroenterology. Based on a standardisation of the format to report and the representation of clinical information on WWW, the multilingual multimedia case has been developed and till now 350 cases have been included. User satisfaction and user acceptance of ELCANO are under evaluation. PMID- 10724958 TI - NISPLAN--projecting hospital beds and hospital stays. AB - The paper describes a computer model for projections of hospital stays and hospital beds, developed by NIS Health Services Research in Norway. The model takes into account demographic changes, changes in medical technology, needs for hospital services, morbidity and hospital structure and population preferences as to choice of hospital. It has been used in several projects in Norway, both by central and local authorities. PMID- 10724959 TI - Automated computer-assisted evaluation of diagnosis-and-procedure-reports in Austrian hospitals. AB - Austria's 320 hospitals with 73,000 beds have to transmit a minimum basic data set for each hospital stay with diagnosis and procedures as well as scores for use in intensive care units (last only in performance-oriented financed hospitals). Based on these data, 160 hospitals or 53,000 beds are financed through performance oriented flat rates per case (LDFs--Leistungsorientierte Diagnosen-Fallpauschalen). Those LDFs are in some principles similar to DRG's (diagnosis related groups), but have been developed by the Federal Ministry of Labor, Health and Social Affairs (BMAGS). 1.94 million data sets of hospital stays, which were reported for financing in the year 1997 had to be analyzed for their correctness and plausibility. For this aim the "Scoringprogramm" of the BMAGS is in use. We describe in detail how many diagnoses and procedures are automatically checked with diagnosis, sex, age, length of stay etc. 1.6% warnings resulted from this checking, and no errors have been reported. The checking routines have to be further developed. At present data seem to be used only with caution for epidemiology or for analyzing medical quality. For financing purposes it might be reliable, especially if data quality commissions are enlarged and if evaluation of data with medical records is improved. PMID- 10724960 TI - Healthcare users and computerised resources. AB - This paper concerns users' views on computerised resources. The focus concerns a software tool that could be used to allow safety conscious user manipulation of the combination of these computerised resources. It is of concern to the authors that computerised resources used in healthcare should reflect users' working practices rather than working practices being changed to reflect computerised resources. PMID- 10724961 TI - Bed utilization performances of Slovenian and Croatian acute hospitals systems. AB - With Barber-Johnson-Yates scattergrams (BJYS), the performances of Slovenian and Croatian acute hospital system were explored by taking turnover interval as a predictor variable, length of stay as a dependent variable, and by looking at the variables of bed throughput and percentage of bed emptiness on installed network. The growing influences on the Slovenian hospital system of a cost-containment policy and of the past war on the Croatian hospital system are the best illustrations of the informatics potential inherent in BJYS presentations for the exploratory data analysis (EDA) used to identify systematic relations between variables in a setting of not complete a priori expectations of these relations. PMID- 10724962 TI - The optimization of turnaround time for blood samples in an emergency clinical laboratory. AB - The University Medical Centre Ljublijana is a 3200-bed general hospital. A laboratory information system, developed to improve the reliability and usefulness of test results, was installed in the emergency laboratory in November 1994. At that time, all test requests received by the laboratory were treated as equally urgent, but in December 1994, a reorganisation took place, which involved the introduction of a priority system. We conducted a study in the period from November 1994 to September 1995. The turnaround time for blood analyses was evaluated by the method of case study. The influence of information technology and organisational changes on individual components of the turnaround time was assessed. This study has shown a positive effect of the recently introduced information technology and organisational changes on the turnaround time. The median turnaround time improved for all phases of the laboratory process. The greatest improvement occurred in validity check time. PMID- 10724963 TI - MOBISIM--package for simulation in molecular biology. AB - A computer program for simulation of protein synthesis regulatory processes is described. A short theoretical background reveals the requirements of a simulation program. Our program, MOBISIM, offers several facilities and a user friendly interface for molecular biologists. An example illustrates the graphical representation both as time evolution of the system or as phase diagram. PMID- 10724964 TI - Automatization of physicians' phone-in hours. AB - In this paper we present a computer-based system that is designed to reduce the number of patient calls in physicians' phone-in hours. Simultaneously it will offer a parallel alternative channel for the patients to listen to their laboratory results. The system requirements were reliability, security and the low costs. The results showed after a two-month test period that the tailored system proved extremely successful. The average time that the physician saved by using this system was over 4 minutes and doctors can leave the messages to the server also outside the phone-in hours. PMID- 10724965 TI - AIDMAN--advanced informatics distributed medical access network. AB - We describe AIDMAN, a pilot project in which health clinics in remote areas of Greece are connected with mainland hospitals, to provide routine clinical service for both chronic health care and accident and emergency. Clinical practice is supported through the use of the Virtual Consultation Workstation that is being developed and evaluated specifically for the project and which is based on readily available commercial products and international standards. The communication infrastructure remains a major obstacle to any telemedicine project, and for this reason, satellite communication is used in the AIDMAN project to overcome the current lack of digital links and bandwidth. The proposed system will provide real time video, high definition image, shared applications and data over a link running at 128 kbps. In this paper we discuss the design of the AIDMAN system, the Virtual Consultation Workstation and the communication channel. PMID- 10724966 TI - Multimedia stimulation for psychophysiological investigation. AB - This paper describes a multimedia approach to sensory stimulation in psychophysiological studies. High resolution colour images are presented to a subject by a Delphi stimulus program, controlled by a data acquisition system which concurrently records cognitive Event Related Potentials from the subject. The acquisition PC is linked to the stimulus PC using serial communications and requests slide material using a character based protocol. The sound card in the data acquisition system can present concurrent complex auditory stimuli for multimedia experiments. PMID- 10724967 TI - Classification of metabolic patients using dynamic variables. AB - The severity of metabolic syndrome is usually determined according to static variables (blood glucose, insulin level, body mass index etc.) The most important classification is dynamic and prognostic classification which can be used to determine the ability to decrease elevated metabolite and hormone levels or to lose weight. Different mathematical approaches were used to determine these phenomena: 1. Mathematical modelling e.g. (Bergman minimal model or glycation model). 2. Predictive calculations using multiple regression (using static and dynamic parameters to determine weight loss in obesity treatment). 3. One day starvation test (finding very variable hormone and metabolic changes in obese patients). We can conclude There are 3 types of metabolic parameters: A. Static (basic) description, B. Functional (actual) description, C. Dynamic-stability describing variables. Mathematical modelling is a complicated method needing many blood samples. It is very invasive for patients and it is difficult to be repeated. Predictive importance can have also repeated measured metabolic data which are able to classify the stability (fixation) of metabolic state. Some basic parameters and simple dynamic tests like one day starvation test can be used in prognostic classification of patients who are able to change their fixed metabolic state. PMID- 10724968 TI - Computer-aided diagnostic system in dentistry. AB - A dental diagnostic system for patient database management, automated measurements, diagnostics and statistical estimation of health conditions is described. To increase the diagnosis quality various diagnostic tools are developed using several statistical methods. The analysis provides information about the importance of particular general, dental, and periodontal data which are further used in quantitative modeling of healing process in the oral cavity. The system supports clinician's diagnostic process and leads to improvement of decisions obtained by a conventional routine. Increased control and evaluation of different therapy types is enabled. PMID- 10724969 TI - A uniform database of rehabilitation as a basis for evaluating the outcome quality. AB - To make it possible to benefit from patient data recorded in rehabilitation clinics, with regard to outcome quality and scientific investigation, the Rehabilitation Research Centre at the University of Ulm has created a rehabilitation register, a database based on ORACLE, into which the relevant patient data is transferred, having been made anonymous via an SQL archiving program. The archived data includes the clinical parameters, the running text of the discharge report, and the doctor's and patient's assessment of the rehabilitation outcome. It can be automatically searched for structured data, according to every conceivable criterion and in every conceivable combination, also in combination with a text search in the running text of the admission and discharge reports and the rehabilitation outcome. With the aid of the rehabilitation register and thanks to the possibilities of EDP (Electronic Data Processing), in future it will be possible to make much more differentiated comments on the rehabilitation outcome than would be the case if the search were restricted to structured data alone. This is essential in order to be able constantly to improve outcome quality and to verify the economic benefit of rehabilitation medicine. PMID- 10724970 TI - Supporting decisions in diabetic patients management through case-based retrieval. AB - In this paper we present a tool for the intelligent retrieval of past cases to support Insulin Dependent Diabetes Mellitus patients therapy revision. This tool was designed for assisting physicians during periodical monitoring visits. Each case is represented through i) a collection of features that describe the patient's clinical state at the decision time, i.e. the control visit, ii) the decision taken in terms of therapy revision and iii) the outcome obtained on the metabolic control during the following monitoring period. A new case is first classified into a protypical monitoring situation, and then similar cases are retrieved and shown to the user. This tool is fully integrated in a Web-based distributed system for Diabetes Management, developed within in the EU project T IDDM. PMID- 10724972 TI - A diagnostic advice system based on pathophysiological models of diseases. AB - Medical decision-support systems in which uncertainty plays an essential role are increasingly based on the formalism of probabilistic networks. Although this formalism is very powerful, the construction of actual networks is not straightforward, and requires the availability of clearly structured medical domain models as a starting point. In this paper it is argued that medical pathophysiological knowledge constitutes a good start for the development of such models, even though pathophysiological knowledge is semantically different from probabilistic knowledge. Two models concerning anaemia, which are part of a broad system covering the domain of anaemia, are discussed to illustrate the general approach. PMID- 10724971 TI - An integration system to rapidly identify aquatic bacteria. AB - An identification system for aquatic bacteria which would be useful for aspects ranging from pollution monitoring to health care, is described. It contains relevant information concerning the nature, taxonomy and activity of aquatic bacteria and allows to identify fresh isolates by means of their chemotaxonomic profiles. The system is simple and user friendly, so that it can be used by people not familiar with computers and has a modular interface that allows an easy interaction with laboratory instruments. The system is also connected with artificial neural network based programs to identify strains starting from the considered chemotaxonomic features. PMID- 10724973 TI - Modelling oculomotor data with decision tree induction. AB - We studied relationships between oculomotor test results and the site of lesion in the data set containing patient cases with operated cerebello-pontine angle tumour, operated hemangioblastoma, infarction of cerebello-brainstem or Meniere's disease and control subjects. Several classification models were generated by decision tree induction to find parameter combinations which are efficient in classification. The studied data were characterised by missing values and scarcity of example cases. This study suggested that decision tree induction can be a useful method even in problematic real world classification tasks. Models generated by decision tree induction and evaluated by an expert physician may give information that is beneficial for research. PMID- 10724975 TI - Hierarchical multi-attribute decision models and their application in health care. AB - Hierarchical decision models are developed through decomposition of complex decision problems into smaller and less complex subproblems. They are aimed at the classification or evaluation of options and can be used for analysis, simulation and explanation. This paper presents a set of methods for the construction and application of qualitative hierarchical decision models in health care. We present the results of four ongoing projects in oncology, radiology, community nursing and diabetic foot treatment. PMID- 10724974 TI - A decision support system for cholinesterase genotyping. AB - Cholinesterase is a naturally occurring enzyme in the human body. Its importance became apparent when it was recognised that patients undergoing surgery and carrying abnormal genes for this enzyme were unable to recover from a dose of muscle-relaxant. It is therefore important to know the Genotype (the pair of genes) of a patient who might undergo surgery. The computer programme takes laboratory measurements and applies mathematical techniques and clinical judgements to determine the two genes present. PMID- 10724976 TI - The influence of class discretization to attribute hierarchy of decision trees. AB - Decision support systems that help physicians are becoming very important part of medical decision making. They are based on different models and the best of them are providing an explanation together with an accurate, reliable and quick response. One of the most viable among decision-making models is the concept of decision trees, already successfully used for many medical decision making purposes. Although effective and reliable, the traditional decision tree construction approach still contains several deficiencies. Therefore we decided to develop and compare several decision supporting models, each of them built with different discretization of attributes and decision classes. For the construction of decision trees we used MtDeciT, in our laboratory developed tool for building decision trees using the classical induction method. All solutions were evolved for determining the influence of basic properties of child and his/her parents to length of successful breastfeeding. A comparison between developed models and obtained results has shown that the way of discretization obviously plays a great role in the reliable and accurate real-world medical decision making. PMID- 10724977 TI - A unified framework for data modeling on medical information systems. AB - Medical Information Systems (MIS) are seen as a way of optimizing the use of existing health-care infrastructure, without resorting to new and costly hospital (re)construction. The qualitative (re)design of such an environment requires a basic understanding of patient and doctors related characteristics and capabilities. Patient care, patient education, medical education, and clinical research need to be considered to meet the basic requirements on the level of services desirable, determined on the basis of the patient's length of stay; i.e., used for modeling the significant entities of such a world. The aim is to extract conclusions for the level of services provided to the users. One's concept will capture, as well as will integrate, the basic design principles under which MIS may be set. PMID- 10724978 TI - A micropopulational modelling of a viral epidemic by using a special neural network. AB - A general forward neural network was adapted for a simulation of viral epidemics. This involves the introduction of a strongly dependence upon history, upon the cumulated values of the corresponding neuron (individual) activations (states of infection) specifying the activation (health) states of the contaminated individuals, represented by the activated neurons and the dynamic parameters of the neural network: the matrix of the synaptic connection and the vector of the activation thresholds (corresponding to the matrix of the viral transfers between the various individuals and to the vector of the minimal individual contamination doses of virus). The recurrence relations and the learning procedures were also adapted to these processes. This methodology was used for the study of the micropopulational spreading of viral epidemics in various epidemiological situations. PMID- 10724979 TI - Computer-assisted planning and simulation of hip operations using virtual three dimensional models. AB - In this paper a new software system for virtual preoperative planning and simulation of hip operations is presented. The system simulates the endoprosthetic reconstruction of the hip joint with hemipelvic replacement for bone tumor patients and supports the individual design of anatomically adaptable, modular prostheses. Three-dimensional models of the patient's hip are generated based on CT data. The surgeon simulates the operation and defines the position and shape of the custom-made endoprosthesis. Stereoscopic visualization and 3D input devices facilitate the navigation and interaction in the virtual environment. Special visualization techniques like texture mapping, color coding of quantitative parameters or transparency support the determination of the correct position and shape of the prosthesis. Furthermore, the system can be used for patient information or educational tasks. PMID- 10724980 TI - A probabilistic approach to improved antibiotic therapy. AB - PTA is a decision-theoretic expert system that aims to assist clinicians in diagnosing and treating patients with pneumonia in the intensive care unit. Its underlying probabilistic network model includes knowledge for diagnosing pneumonia on the basis of the likelihood of tracheobronchial-tree colonisation by pathogens, and symptoms and signs actually present in the patient. Optimal antibiotic therapy is selected by balancing the expected efficacy of treatment, which is related to the likelihood of pathogen-specific pneumonias, against costs and side effects of treatment. In this article, the model underlying the system and results of a preliminary evaluation are described. PMID- 10724981 TI - Possibilities of See5 software in forecasting of life expectancy not achieving. AB - Results of application of See5 to epidemiological domain were presented. The aim of this paper is to find out if See5 can be used for forecasting whether life expectancy will be achieved or not. The basis for forecasting are data attributes, predictors (anthropological data, living habits, laboratory data, blood pressure measured in years 1970/71, and the status of examinee--class (alive or dead) in 1990. Data were split at random into 10 blocks with approximately the same number of cases and the same distribution of classes. Results were given in the form of decision trees and rulesets, and tested through crossvalidation. The most interesting question--what are prediction-candidates for not achieving life expectancy estimated in the year of examination--did not give a satisfactory result: the accuracy of classification was about 70%. However, a very interesting fact is that each rule consists of at least one candidate predictor, usually considered as risk factor (e.g. high diastolic blood pressure and animal fat consumption). PMID- 10724982 TI - Expanding DIOGENE with a clinical information system based on a new hospital-wide clinical finding dictionary. AB - The aim of this project is to expand DIOGENE with a centralized and integrated patient clinical database system providing a standardized framework for the building of future clinical databases and for the integration of existing heterogeneous ones. The combined 'across time view' and 'across departments view' generated from the integrated clinical data will enable an evolutionary view of the patient state both across time and across medical specialties. For this purpose and to permit the communication and exchange of data, a new controlled vocabulary for representing clinical data has been created. The construction of this vocabulary is based on the international ICD classification, already being used in DIOGENE for encoding patient diagnosis and procedures. A new extension of the ICD is proposed for medical information that goes beyond diagnosis and procedures encoding. The building of a common clinical finding dictionary recording the definition of findings is based on this newly developed clinical vocabulary. This process is incremental, manual, and significant. PMID- 10724983 TI - Intelligent systems in medical diagnosis. AB - From an intelligent system for a computer supported medical diagnosis it is expected to achieve high accuracy and ability to draw conclusions from a small data sets. Medical practice could takes many years to generate a large database. A new mathematical method that is able to learn on a small data set is presented. PMID- 10724984 TI - Power of heterogeneous computing as a vehicle for implementing E3 medical decision support systems. AB - A computer system of PCs, workstations, minicomputers etc., connected together via a local area network or wide area network represents a large pool of computational power. Our aim is to use this power for the implementation of E3 (efficiency, effectiveness, efficacy) medical decision support system, which can be based on different models; the best of them are providing an explanation together with an accurate and reliable response. One of the most viable among models are decision trees, already used for many medical decision making purposes. In this paper we would like to present a heterogeneous implementation of genetic algorithm for the induction of decision trees with emphasis on solving the mitral valve prolapse syndrome. PMID- 10724985 TI - Neural network in communication with medical computer system. AB - The paper presents a concept of an experimental module designed to recognise spoken utterances that cover a limited range of words indispensable in dialogues with computer medical systems. Research into the recognition of spoken words by a module based on artificial neural network is described. Usefulness of the obtained results for surgery-assisting multimedia systems and for a patient simulator supporting medical education of students in case history-taking and diagnosing is also discussed. PMID- 10724986 TI - Concepts, contexts and expert systems. AB - This paper describes problems identified in our attempts to develop an expert system for management of urinary tract infections. We found three aspects which we believe are important to consider when developing such systems. The objective of our future work will be to evaluate the impact of these problems on expert system development and usage. PMID- 10724987 TI - Availability humanization expert model. AB - AH model as a tool for estimation in the occupational health care is presented. This is an expert model connecting data from the working environment with human availability and health status data. In this paper the idea of model and its components are presented. PMID- 10724988 TI - Preliminary study on relevant data for health management in hospitals. AB - Data and information are nowadays keys of success. We experienced the building of an hospital Information System (IS) selecting data from those already routinely produced and sharing them with health care units and general management. IS should be shaped to accomplish mission, vision and strategic direction of the hospital and in this direction our efforts are oriented. PMID- 10724989 TI - TIDE: an intelligent home-based healthcare information & diagnostic environment. AB - The 21st century promises to usher in an era of Internet based healthcare services--Tele-Healthcare. Such services augur well with the on-going paradigm shift in healthcare delivery patterns, i.e. patient centred services as opposed to provider centred services and wellness maintenance as opposed to illness management. This paper presents a Tele-Healthcare info-structure TIDE--an 'intelligent' wellness-oriented healthcare delivery environment. TIDE incorporates two WWW-based healthcare systems: (1) AIMS (Automated Health Monitoring System) for wellness maintenance and (2) IDEAS (Illness Diagnostic & Advisory System) for illness management. Our proposal comes from an attempt to rethink the sources of possible leverage in improving healthcare; vis-a-vis the provision of a continuum of personalised home-based healthcare services that emphasise the role of the individual in self health maintenance. PMID- 10724990 TI - Healthcare knowledge management through building and operationalising healthcare enterprise memory. AB - In this paper we suggest that the healthcare enterprise needs to be more conscious of its vast knowledge resources vis-a-vis the exploitation of knowledge management techniques to efficiently manage its knowledge. The development of healthcare enterprise memory is suggested as a solution, together with a novel approach advocating the operationalisation of healthcare enterprise memories leading to the modelling of healthcare processes for strategic planning. As an example, we present a simulation of Service Delivery Time in a hospital's OPD. PMID- 10724991 TI - Guidelines in healthcare: the experience of the Prestige project. AB - The paper reports the work and results of Prestige: Guidelines in Healthcare, a large EU project designed to applying ICT to assist the application of clinical practice guidelines. PMID- 10724992 TI - The RITHME inter-mediation platform for data exchanges between healthcare professionals. AB - Data Exchanges between Healthcare Professionals are always limited to paper. For letters, reports, results, the physicians and hospitals are yet using the classical post-office mailing system. With the growing development of Medical Informatics within the Hospital Information Systems and the GPs offices, the way to exchange medical data will be more and more oriented towards the Telematics procedures. If Tele-consultation, video-conferencing applications appear as routine procedures, a safe, protected, and rapid transmission of medical data and records through electronic mailing systems is not yet really available. The Rithme Intermediation Platform is an experimental product, tested in the town of Armentieres (in the City of Lille) in the North of France to allow a better, safer, more rapid electronic communication between all the professionals of a geographical sector. One of the mail component of this platform is the "Events Server" which collects and displays all the available information concerning a dedicated patient to the medical authorized correspondents. To realise this task the Rithme Platform takes charge of 4 main functions: formatting the messages, assuring the security, managing the directories, notifying emergencies. Currently experienced with success in the City of Lille, this Platform will be soon customised and commercialized. PMID- 10724993 TI - Development of a reminder system for general practitioners. AB - In this contribution we describe a prototype reminder system that has been developed to influence General Practitioners' (GP) test request behaviour. It is intended to substitute conventional written feedback by human experts. The system generates critical comments about the rationality of the test request at the moment the GP requests a test that is not in line with national or regional guidelines. PMID- 10724994 TI - Quantitative collagen as a golden standard in differential diagnosing of fibrotic changes in liver tissue. AB - Determining a presence and degree of liver fibrosis provides means for diagnosing disease related processes. We have used two data mining methods, discriminant and regression analyses, to acquire knowledge from the data of 211 patients. We have shown and discussed that quantitative collagen has a distinguished discriminating power and can serve as a golden standard. We have additionally succeeded to obtain a formula consisting of standardised blood tests that can replace quantitative collagen. Practical implications of this is a non-invasive and cost efficient patient examination. All the results are now left for clinical evaluation and so is the current way of histopathological classifications. PMID- 10724995 TI - A development protocol for a diagnostic DSS. AB - A diagnostic decision support system (DSS) in medicine is an expert system that aids the physician in the determination of the diagnosis based on findings and test results. The DSS can be divided into 2 different types of components: the knowledge component and the information system component. Methods from software engineering, knowledge engineering and management are combined into a dynamic development cycle that allows stepwise update and refinement. The development of the knowledge component is based on knowledge engineering. In the starting phases, rapid prototyping is convenient to determine and evaluate the specification. The content of the knowledge base is frequently updated during its life time. For this purpose, a knowledge modelling protocol is supplied. PMID- 10724996 TI - A protocol building software tool for medical device quality control tests. AB - Q-Pro is an application for Quality Control and Inspection of Medical Devices. General system requirements include friendly and comprehensive graphical environment and proper, quick, easy and intuitive user interface. Functions such as, a tool library for protocol design widely used multimedia, as well as, a support of a local database for protocol and inventory data archiving are provided by the system. In order to serve the different categories of users, involved in Quality Control procedures, the system has been split into three modules of different functionality and complexity, each of which can work as a stand-alone application. The implementation of protocols and use of the software functions, as well as, the user interface itself have been proved by the evaluators to be clear and intuitive. The software seems to adapt easily to different kinds of Quality Control procedures and objectives. Q-Pro effectively supports and enhances the processes to attain a highly tuned, professional, responsive and effective quality control and preventive maintenance procedures for biomedical equipment management. PMID- 10724997 TI - Telematics applications to support the role of the community pharmacists as self medication advisors. TESEMED Consortium. AB - One of the applications developed within the EU-funded projects TESEMED and TESEMED-II is a program for the information and continuous training of community pharmacists, with the aim to empower them as advisors of the citizens about self medication topics. Several programs are being developed on the basis of ad-hoc developed protocols about minor ailments (currently, cold and flu, haemorrhoids, constipation and cough). Each program includes three modules: a hypertextual version of the protocol, an interactive scheme of it, and an educational tool called Encounter Simulator, that trains the pharmacist about the protocol by means of simulated pharmacist-customer interactions. The testing of these applications with 84 community pharmacists offers positive results in terms of expectations, program characteristics and perceived usefulness. PMID- 10724998 TI - The "mediator service", a component to ease the integration of medical applications in the SynEx framework. AB - Interoperability is a key issue and a long-term domain of research for distributed healthcare information systems. The European project SynEx provides an open and standard integration platform for both new and legacy medical applications. It allows the collaboration of distributed and heterogeneous healthcare records and services. It aims to provide access to Hospital Information Services, to remote sources of medical data and to medical knowledge, in a seamless way, hiding the distribution aspects and the heterogeneity of systems. In this project, the Medical Informatics Department of the Broussais University Hospital is responsible for the development of the "Mediator Service". It is a software component of the SynEx platform which is used as a "glue" mechanism to provide a flexible way to facilitate the interchange between any pair of systems, with different nomenclatures and data structures. The Mediator Service uses a generic model of mediators to create, through specialization, specific mediators for practical cases. Based on this model, it offers a C++ library to be used as the tool case by the programmers, to reduce the development effort. The use of XML as a powerful data interchange format and as a data structure descriptor is proposed and evaluated. PMID- 10724999 TI - Emergency protocols in prehospital emergency medicine in Slovenia. AB - Emergency medicine is a branch of medicine in which prompt and accurate information is of crucial importance, because a patient's survival and the final outcome of a treatment very often depends on it. Therefore the standardization of prehospital emergency service administration with uniform documentation, methods of annotation of technical data as well as uniform gathering and data analysis are necessary. Wanting to process a fair amount of data and specially to disburden personnel, a need for computer aided processing is evident. Application that was developed in strong collaboration with EMS project team and information science engineers definitely improves data quality. Firstly when team members, especially physicians, are constrained to fulfill paper forms and secondly when data are entered into the computer, because all inconsistencies are being reconsidered and corrected if necessary. The developed application also gave the opportunity to the emergency team for prompt and accurate information. PMID- 10725000 TI - A system for the description of healthcare guidelines. AB - Contained costs and quality of treatment are two elements which have to be considered when planning and controlling both clinical and managerial activities of healthcare structures (HCSs). Each structure has to be marked by a high level of quality of treatment and by specific professional competencies and skills of its operators. It is therefore fundamental to provide a system which supports the management of processes and of guidelines which characterise an healthcare structure. In this paper we present a system able to describe processes of an HCS and in particular clinical guidelines. This system is based on a workflow conceptual model, able to represent clinical and managerial activities carried out in HCSs (the Atreus model). PMID- 10725001 TI - An approach for access differentiation design in medical distributed applications built on databases. AB - A formalized "top to bottom" design approach was described in [1] for distributed applications built on databases, which were considered as a medium between virtual and real user environments for a specific medical application. Merging different components within a unified distributed application posits new essential problems for software. Particularly protection tools, which are sufficient separately, become deficient during the integration due to specific additional links and relationships not considered formerly. E.g., it is impossible to protect a shared object in the virtual operating room using only DBMS protection tools, if the object is stored as a record in DB tables. The solution of the problem should be found only within the more general application framework. Appropriate tools are absent or unavailable. The present paper suggests a detailed outline of a design and testing toolset for access differentiation systems (ADS) in distributed medical applications which use databases. The appropriate formal model as well as tools for its mapping to a DMBS are suggested. Remote users connected via global networks are considered too. PMID- 10725002 TI - Take care: guidelines for patients with chronic hepatitis C. AB - Alcohol consumption has significant impact on the condition of the liver, by itself, and even more in conjunction with other liver diseases such as chronic hepatitis C. Drinking habits might be delicate issues to address and could harm otherwise satisfying communication. Therefore, we intended to outline guidelines for advising hepatitis C patients concerning alcohol consumption. Analysis of a relatively limited knowledge base revealed the complexity of the disease rather than statistically significant findings regarding consumption. Thus, we instead chose to suggest a set of patient educational guidelines, which could be implemented on the Internet, hypothesizing that a better informed patient will be more able to comply with restrictions concerning alcohol consumption. A brief ad hoc evaluation pointed out Internet as a favourable media to present the information. We also suggest a tentative algorithm for further development of clinical decision support systems addressing monitoring of chronic hepatitis C patients. PMID- 10725003 TI - C3: Java-based medical record system for cardiology. AB - This paper describes a system for electronic medical record (EMR) we have developed for use in our health care institution, mainly dealing with diagnosis and treatment of cardiovascular pathologies. This activity is part of the project SPERIGEST, supported by Health Ministry of Italy, for the management of health care delivery, as concerns both clinical and administrative aspects. A networked computer-based information system was realized to integrate the different heterogeneous sources of patient information. Both clinical and administrative patient relevant data are provided from the various systems and stored into a central database. The EMR system was designed using World Wide Web (WWW) technology (Java, HTML). The system is currently under clinical evaluation. PMID- 10725005 TI - QUALIDIAB: implementation of the Diabcare project in the French-speaking environment: regional, national and international issues. Qualidiab Group. AB - Diabcare is an international Project devoted to the evaluation of the Quality of Care [1] in the population of diabetic patients. As Diabetes is one of the most frequent chronic illnesses, the management of its treatment has a strong impact on the public health policy both regional and national [2]. Funded by the European Commission, initiated by WHO and the International Diabetes Federation (IDF), this Project is now recognised as a Pilot Project throughout the world as an example of a large international collaboration for the surveillance of quality of care in a typical chronic disease. PMID- 10725004 TI - An experimental electronic patient record for stroke patients. AB - This contribution describes an electronic patient record for stroke patients at the neurology ward of the Maastricht University Hospital. Daily practice at the ward will be supported with the developed electronic patient record that integrates both the medical and the nursing record, that will provide decision support and it will be connected to the hospital information system. In an evaluation project we will study the effects of the usage of the electronic patient record and additional effects of providing decision support. PMID- 10725006 TI - The electronic patient records of the Hannover Medical School. AB - In this paper, the successful introduction of a commercially available electronic patient record archiving system at the Hannover Medical School is described. Since 1996, more than 11 million document sheets of 130,000 patient records have been stored electronically. Currently, 100,000 sheets are stored each week. PMID- 10725007 TI - DIABCARD core system--a chip card medical information system for diabetes care. AB - A chip card based medical information system was developed as a good possibility to create a portable electronic patient record. The produced software module provides an on-line, portable diabetes medical record information system. In particular the patient data card makes the up-to-date patient's record available whenever needed. The developed Core System includes a patient record management system that has the ability to handle topics such as medical anamnesis, administrative, medical and physical examination data. Issues tackled were simplicity, data security and reporting. Customization and internationalization were also covered by presenting a novel approach using native initialization files. Proper care has been addressed during the development of the software modules for matters of security, data integrity and confidentiality. PMID- 10725008 TI - Electronic patient record for N.N. Burdenko Neurosurgical Institute: on the verge of implementation. AB - This presentation is the first report about development of Electronic Patient Record System (EPRS) for N.N. Burdenko Neurosurgical Institute (NSI). This EPR system is the core of Integrated Automatic Information System intended to support all business processes running in the Institute. A new technology for developing information systems in poorly formalized subject domains, named IBS/Records, was was created. PMID- 10725009 TI - Towards a multi-professional patient record--a study of the use of headings. AB - This paper reports on the differences and similarities of used headings among Swedish health care professionals; nurses, occupational therapists, physiotherapists, dietetics, logopedics, welfare officers and general practitioners. The background of the study is a national project where representatives of the different health care professionals compiled used headings from clinical practice. Based on that survey, a hierarchical system of headings was constructed in accordance with the notion of record items and record item complexes described in the European Health Care Record Architecture (EHCRA). The work was done separately by the different health care professionals, leading to separate lists of headings. The current study reports on an analysis of these multi-professional lists of headings with respect to structure, degree of specialization, synonyms and homonyms. The study is descriptive in nature, giving a status report of the variety of used headings in clinical practice, providing necessary material for a normative approach with focus on a truly multi professional patient record in the future. PMID- 10725010 TI - Validation of a European message standard for electronic health records. AB - A European Standard message for communication of the contents of Electronic Health Records is currently at a late stage in development and is due to be adopted during 1999. This paper reports on the development and content of this message and on current work to clinically validate this message. The validation process involves a new method based on an instantiation of the proposed message in the Extensible Mark-up Language. This method allows the message to be viewed and compared with the same record on the source system. PMID- 10725011 TI - Clinicians must invest resources when implementing electronic patient records. AB - Most often the investment in Electronic Patient Records (EPR) is decided upon on the basis of descriptions of all the advantages this new technology will bring. However, most of these advantages from EPR's are only obtainable if the clinicians are motivated to invest their knowledge and resources in this implementation process. This paper proposes a simple method to qualify the decision making process by introducing the "price to pay" concept. PMID- 10725012 TI - Constructing scenarios for validating the electronic health care record architecture. AB - The CEN TC 251 Electronic Health Care Record Architecture is the first part of a four part communication prestandard intended to facilitate the safe communication of clinical information, irrespective of circumstances related to time, distance, and responsibility. The work has important implications for a diversity of stakeholders and must embody clinical principles if it is to succeed. The issue at hand is how to enable clinicians to validate the reference architecture, which is necessarily a technical document, so that they have confidence that it can meet the clinical requirement for safe communication. A model of communication that has been used to ground the architecture is shown to be useful in creating relevant clinical scenarios to permit the first steps towards coherent validation in a systematic way. PMID- 10725013 TI - Integrated databases--the foundation for the information linking of the actors in the national health care and health insurance systems. AB - Among the characteristics of the present time, we meet the ranking of information among the key resources of effective management of business systems. Up to date, integral and accurate information have gained precedence over conventional economic resources. This applies particularly in the case of large and complex business systems, including the national health care and health insurance systems. High quality and unified databases form the foundation for rational and quality procedures ranging from the operative level to the national strategy level. The goal of this paper is to demonstrate, on the example of Slovene experience, the importance of keeping quality databases for the purposes of health care and health insurance system management. The paper reviews the achieved level and the plans of further development of unified databases in this sector in Slovenia. The key emphasis in the development of an integrated system has been laid upon establishing uniform primary databases and providing appropriate integration in the area of data interchange among the actors of the health care and health insurance systems and other national systems. PMID- 10725014 TI - Realization of a CPR using HL7 queries from communication server to information server. AB - A communication server is often used in Germany to interface legacy systems. It handles the translation, duplication, and transmission of messages to all systems that might need this information. Based on the principles of "for every data there is only one master" and "information on demand" we have built an information server that reduces the messages required and simplifies the handling of messages, the update of data already sent compared to a communication server. It also allows to implement a security policy as required by the German data privacy legislature. PMID- 10725015 TI - Design of the SGML-based electronic patient record system with the use of object oriented analysis methods. AB - Since a patient record is typically a document updated by many users, required to be represented in many different layouts, and transferred from place to place, it is a good candidate to be represented structured and coded using the SGML document standard. The use of the SGML requires that the structure of the document is defined in advance by a Document Type Definition (DTD) and the document follows it. This paper represents a method which derives an SGML DTD by starting from the description of the usage of the patient record in medical care and nursing. PMID- 10725016 TI - Computerised patient record with distributed objects. AB - The vast spectrum of information and functionality requirements imposed on a Computerised Patient Record (CPR), fueled by an ever changing and expanding business model demands information system interoperability. The management of information, created across the continuum of care, and associated information system functionality, can not be provided by data interchange to and from monolithic applications. WebDoctor is a Computerised Patient Record (CPR) which is fully used at the Institute of Oncology in Ljubljana, Slovenia--a hospital with 500 beds and more than 200 users, all of them medical professionals. The data are stored in an underlying Oracle hospital data base. For logging the username and password security is used. WebDoctor uses Internationalization APIs. Currently GUI is currently written in Slovenian language, but can be easily adapted to any other language. It is available in either Metal or Windows look. Search for patients is based on CPR No. or partial data from demographics. All the available patients data can be found on a single screen divided into several tab sections. The tab sections cover general and speciality data. The general data include demographics, admissions and diagnoses, meanwhile the speciality data are divided into Labs where data are represented numerically by date or by type and graphically with the ability of detailed view in separate window, Radiology where results are represented in textual form as well as pictures together with a special viewer to provide detailed analyses and Radioisotopes where results are also being represented in textual form together with a graphical representation. WebDoctor is running on virtually any platform. It achieved the 100% Java Certification which places the application among the firsts if not the first of this kind in the healthcare industry. It excels with a small and light client which doesn't exceed the 150K. PMID- 10725017 TI - The holistic architectural approach to integrating the healthcare record in the overall information system. AB - The integration and evolution of existing systems represents one of the most urgent problems facing those responsible for healthcare information systems so that the needs of the whole organisation are addressed. The management of the healthcare record represents one of the major requirements in the overall process, however it is also necessary to ensure that the healthcare record and other healthcare information is integrated within the context of an overall healthcare information system. The CEN ENV 12967-1 'Healthcare Information Systems Architecture' standard defines a holistic architectural approach where the various, organisational, clinical, administrative and managerial requirements co-exist and cooperate, relying on a common heritage of information and services. This paper reviews the middleware-based approach adopted by CEN ENV 12967-1 and the specialisation necessary for the healthcare record based on CEN ENV 12265 'Electronic Healthcare Record Architecture'. PMID- 10725018 TI - Document versus data centred approach to the EPR. AB - This paper presents the document centred Electronic Patient Record (EPR) as currently in production in a large university hospital and subject to multiple additional developments in the coming years. A number of basic hypothesis have been developed in order to reach the best medical practice and the success of this application. In addition, the alternative approach of data centred EPR is compared, and different benefits and pitfalls are highlighted. It is not easy to evaluate the consequences of such an initial trend, but changing one's mind after having a system in daily production is anyway costly and difficult. Therefore, the selection of the right orientation in a given hospital necessitates a scientific debate. PMID- 10725019 TI - Donor and the health insurance card. AB - The health insurance card is a new electronic identification document of insured persons and facilitates smoother communication between the health insurance information system and the Slovene health care service provider information systems. The card stores the insured person's identification details as well as the information on the selected personal physician. Another procedure also to be hosted on the card, will be the card holder's personal decision about being voluntary donor of organs and tissues for transplant purposes. This data item will, supported by the enhancement of technological infrastructure, improve the control of the donorship information. In Slovenia, registering, recording and application of the organ and tissue donorship data is carried out, on a pilot basis, in the region of Posavje. PMID- 10725020 TI - Structuring clinical information in healthcare records. AB - We carried out an analytical study about names of clinical documents, titles of generic sections, names of data elements, to prepare the European Prestandard CEN ENV13606-2 on "Health Informatics--Communication of Electronic Health Care Record -Part 2: Domain Termlist". The goal of the standard is to facilitate transmission and/or homogeneous browsing of clinical information from disparate patient records, without any preliminary agreement on coding systems, data elements, record organization. With the assistance of the members of CEN/TC251/PT27 and under the control of CEN/TC251/WG I and WG II, we defined three layers to structure clinical information (structuring records into complexes, complexes into statements, statements into details) and we prepared many lists suitable to represent coarse-grained information about the different constructs used in the above layers. PMID- 10725021 TI - Security for electronic patient record systems. AB - The presentation is devoted to the problem of security for Electronic patient record system with EPRS for N.N. Burdenko Neurosurgical Institute (NSI) as an example. PMID- 10725022 TI - Professional knowledge as the basis of electronic patient record systems. AB - The presentation is devoted to the role of professional knowledge in the developing of Electronic Patient Record System (EPRS). The main example--EPRS for N.N. Burdenko Neurosurgical Institute (NSI). PMID- 10725023 TI - A method for an automated decomposition of a complex term into its conceptual components. AB - The paper deals with the improvement of the MAOUSSC model (Modele d'Aide et d'Orientation d'un Utilisateur au Sein des Systemes de Codage) and system. Its specific purpose is the automation of the description of medical and surgical procedures. We have developed an automatic decomposition method using a linguistic and conceptual approach based on the UMLS knowledge base. This work concerns the processing of 100 procedure wordings from the digestive surgery domain. We introduce a prototype of such a system automating the decomposition through a web interface. PMID- 10725024 TI - Automatic enrichment of the unified medical language system starting from the ADM knowledge base. AB - The Unified Medical Language System (UMLS) project aims to provide a repository of terms, concepts and relationships from several medical classifications. This work describes the possibility to enrich automatically with meaningful links the UMLS database by using description of diseases from another knowledge base, in our case ADM (Aide au Diagnostic Medical). In spite of the constraints and the difficulties to qualify the interconcept links, the results show that it is possible to find and create new links from a french knowledge database to the UMLS one. One of the interests of this work is that the automated learning of the connections could be used with others knowledge databases like expert system databases. PMID- 10725025 TI - Maintenance of self-consistency of coding tables by statistical analysis of word co-occurrences. AB - The author presents a method for maintaining the internal consistency of coding tables. The method was tested on a table used for assisting the daily work of indexing clinical cases to International Classification of Diseases. 300 item were tested selected randomly form a corpus of 3082 clinical diagnoses. The method discovered potential consistency problems in 39 cases, out of which 10 were false positive. PMID- 10725026 TI - Compulsory health insurance databases. AB - The compulsory health insurance databases represent fundamental infrastructure to the implementation of the Health Insurance Institute of Slovenia (HIIS) operations, i.e. for its effective performance in all the fields of its business. This paper presents the legal bases of the database management, the logocal database structure, the supply of data from various data sources, and the role of databases as the primary source of supply for the data collections kept by other institutions in Slovenia. PMID- 10725027 TI - Towards a multilingual morpheme thesaurus for medical free-text retrieval. AB - We introduce a methodology for the segmentation of complex compounds into medically plausible morphemes. A tool for thesaurus compilation and management is presented, and design principles for a multilingual morpheme thesaurus are outlined. Our goal is to enhance the quality of medical free-text retrieval by replacing lexically based through morpheme-based search procedures. PMID- 10725028 TI - A model for integration and continuous development of standards for tumour documentation using relational database techniques and extensible markup language. AB - In oncology various international and national standards exist for different aspects of a disease. These standards, maintained by different organisations, have multiple relationships with each other. A common data dictionary like UMLS would facilitate the reorganisation of such relationships when a new version of a standard is published. While the modelling of relationships usually is restricted to types having a relevant frequency, there are often relationships which are expressed in texts like definitions or explanations. Such texts are a very important supplement for the acceptance and the safe use of coding systems, but often are neglected when implementing coding systems in computerised systems, because they are costly to implement. This paper discusses potentials when integrating various sources in a common, database based dictionary enhanced by XML (Extensible Markup Language) techniques. PMID- 10725029 TI - Preliminary report: concepts and terms used to describe urinary tract infection in primary health care and in the clinical microbiology laboratory. PMID- 10725030 TI - Galen: a third generation terminology tool to support a multipurpose national coding system for surgical procedures. AB - GALEN has developed a new generation of terminology tools based on a language independent concept reference model using a compositional formalism allowing computer processing and multiple reuses. During the 4th framework program project Galen-In-Use we applied the modelling and the tools to the development of a new multipurpose coding system for surgical procedures (CCAM) in France. On one hand we contributed to a language independent knowledge repository for multicultural Europe. On the other hand we support the traditional process for creating a new coding system in medicine which is very much labour consuming by artificial intelligence tools using a medically oriented recursive ontology and natural language processing. We used an integrated software named CLAW to process French professional medical language rubrics produced by the national colleges of surgeons into intermediate dissections and to the Grail reference ontology model representation. From this language independent concept model representation on one hand we generate controlled French natural language to support the finalization of the linguistic labels in relation with the meanings of the conceptual system structure. On the other hand the classification manager of third generation proves to be very powerful to retrieve the initial professional rubrics with different categories of concepts within a semantic network. PMID- 10725031 TI - Synapses/SynEx goes XML. AB - This paper describes the first approach to use the Extensible Markup Language (XML) as a data interchange/delivery format in the Synapses and SynEx healthcare environment. It will ease the semantic mapping between healthcare systems (server server, server-client) and make them more interoperable. PMID- 10725032 TI - German adaptations of ICD-10. AB - The introduction of the ICD-10, published by WHO 1992-94 in English and by DIMDI 1994/95 in German, is a very slow process. Some states introduced ICD-10 for the preparation of statistics of mortality, but only few use it for morbidity. ICD-9 is in Germany only in hospitals still in use. Much effort was put into the improvement of the official ICD-10 and the development of additional aids for more simple and better encoding of diagnoses. Thus a revision especially for ambulatory health care (ICD-10-SGBV with the incorporated ICD-10-Basisschlussel) and a collection of German terms and expressions of diagnoses that are not at all part of the official ICD-10 (ICD-10-Diagnosenthesaurus) were published. Three years ago a conversion table ICD-9/-10 was developed which can now be harmonised with WHO's Translator. The experiences with all these instruments are satisfying. The development of methods for automatic encoding of free-text phrases of diagnoses has now been started. PMID- 10725033 TI - SysTerN. A nursing terminology system based on ICNP. AB - This paper is presenting a terminology system for nursing--SysTerN--that is based on the ICNP1 classification. SysTerN has been developed using the framework of the TeleNurse ID-ENTITY Telematics for Health EU project as a support for the dissemination actions carried on by CCSSDM (Center for Health Computing Statistics and Medical Documentation of the Romanian Ministry of Health) for Romania and other Central Eastern European countries. Currently, the terminology system uses the alpha version of ICNP, but this version is going to be replaced by the beta version. SysTerN is designed primarily for Romanian users but an English version is also available for other partners from CEE countries. PMID- 10725034 TI - Electronic dissemination media for promoting ICNP in CEE countries. AB - The TeleNurse ID-ENTITY Inco-Copernicus project, co-ordinated by Randi A. Mortensen, director of the Danish Institute for Health and Nursing Research, is offering to the health informatics community of CEE countries a frame and an opportunity to determine nurses to become active actors in the development of the nursing language and terminology and to promote nursing informatics. This is of great importance, for these countries do not have yet a standard for the electronic patient record and the standard that will be proposed has to incorporate a nursing minimum data set. The ICNP1(International Classification of Nursing Practices) classification of nursing phenomena and interventions is a polyhierarchical, multi-axial classification system, allowing different concepts in nursing practice and from different classification systems, to be expressed as combinations of ICNP concepts, located on different axis. The features of ICNP are making it a sound candidate for a standard computerised nursing language. As part of the general strategy for promoting and disseminating ICNP in CEEC, CCSSDM has developed an electronic environment based on Internet and WWW facilities. The main composant and characteristics of this electronic dissemination environment are described in this paper. PMID- 10725035 TI - The development and testing of information system for community nursing. AB - The paper presents the development and testing of information system (IS) for community nursing (CN). The goal of IS is to support CN practice and to encourage research and development in the field. The development was based on analysis of CN process. Principal functions of CN were analysed. The logical data model and corresponding user interface were designed. A special emphasis in the development process was put on testing in practice and users' training. The developed IS will: increase work efficiency, introduce process-oriented nursing doctrine, support integrated treatment of the subjects and enable data comparison and exchange. An important subsidary goal of the IS development and usage is the harmonisation of concepts and terminology used in Slovenia with those used in the EU. In this framework International Classification of Nursing Practice (ICNP) which was translated into Slovene was introduced and tested. PMID- 10725036 TI - Plea for a standardised detailed clinical nomenclature. AB - The choice of a nomenclature for the encoding of the medical information to be stored in an electronic patient record (EPR) is a critical issue. As we are currently developing a neurosurgical EPR, we evaluated three nomenclatures or classifications, Read, ICD-10 and Quick Medical Reference (QMR) for the capture of the detailed concepts referenced in the EPR. We scored the correspondence with 2 for a good match, 1 for a fair match and 0 for no match. The Read nomenclature ranked first with an overall score of 1.21 (max. 2.0), the ICD-10 obtained 0.88 and the QMR 0.74. Some groups of items such as the neurosurgical history and examination were fairly well represented in the three systems. On the opposite, others such as the various neurosurgical clinical and radiological scoring and grading systems and the outcome descriptors were not correctly referenced in any coding system. Although the Read coding system has been advocated to represent the clinical activity in neurosurgery, it still needs an enrichment before being able to completely cover the concepts present in a neurosurgical record. Moreover the development of an international, standardised, detailed nomenclature and classification collecting the advantages of the various coding systems currently in use should be encouraged to be able to exchange and compare medical data. PMID- 10725037 TI - A computerised guideline for pressure ulcer prevention. AB - This work illustrates the implementation of a computerised guideline for the pressure ulcers prevention. In particular, we describe the site-specification of a guideline delivered by the Agency for Health Care Policy Research, its integration with the electronic patient record, and its introduction within the clinical routine. The system facilitates trained nurses in the patient management by producing daily workplans, and novice nurses by running as an educational tool. PMID- 10725038 TI - How can we improve informatics education for German nurses? Statements derived from the first German nursing informatics summer school. AB - For German nurses it is difficult to join training in health informatics besides their professional activity. The authors have successfully established a German nursing informatics summer school in shape of a 5 day intensive curriculum which they offer to German nurses during the summer holidays. The summer school introduces nurses into health informatics and nursing informatics. It targets interested nursing staff, nurse executives, and nurse teachers. It promotes self learning abilities for continued self education of the participants. One of its goals is to enable participants to formulate their own requirements in health information processing and to influence system design and system introduction. The paper presents the curriculum, talks about first experiences, and demonstrates the results of an evaluation among the participants. Conclusions are drawn in a set of statements on informatics education of nurses. PMID- 10725039 TI - A tutor for nursing process education. AB - Computer managed instruction (CMI) has been used in nursing education since the late 1960's. It is due to the accessibility and self paced format that CMI is very well suited for both students and practicing nurses, while learning can occur at the learner's own pace and time. In addition CMI supports also continuing education and distant learning. The aim of this paper is to present CArE--a software package for Computer Aided Nurse Education in particular for teaching the basics of the nursing care process, developed as a result of the Phare TEMPUS project called NICE (Nursing Informatics and Computer Aided Education). PMID- 10725040 TI - Database for gynecological laparoscopy. AB - Laparoscopy is a wonderful tool for performing gynecologic surgery. Whether or not laparoscopic surgery is advisable to diagnose or treat a particular gynecologic problem requires a careful consultation with an experienced gynecologist, who can help the patient weight the pros and cons of laparoscopy versus other options. For the management of this kind of surgical intervention we realise a computer based patient record. PMID- 10725041 TI - Towards a nursing information system in Slovenia. AB - In the paper are described the starting points of health informatic system development in Slovenia as also the situation in nursing. There are presented the goals, which nurses want to achieve with informatic system development and the activities for their attainability. PMID- 10725042 TI - Internet provides fast and inexpensive information to home care. PMID- 10725043 TI - Application report: preliminary evaluation of the T-IDDM project in Pavia. PMID- 10725044 TI - Target practice over healthcare issues. PMID- 10725045 TI - A comparison of high- and low-income physicians. PMID- 10725046 TI - Turning around a financially distressed practice: the operational review. AB - Given substantial changes in the marketplace, many physician practices have responded by growing rapidly, merging with or acquiring other practices, or selling the practice to physician practice management companies (PPMCs) or hospital systems. The hoped-for economies of scale, increased market share and profit, or other goals often have not come to fruition. Many practices now find themselves in difficult operational and financial circumstances. This article describes how group practices can solve such operational and financial problems while retaining independence and strategic control of the practice. The strategy involves retaining a professional turnaround company with expertise in reestablishing the financial security of group practices. The article provides characteristics of how turnaround companies operate and the key elements of operational reviews, the tool used by turnaround companies to detect and analyze existing practice problems. Operational reviews include organization analysis, management review, and financial analysis, each of which is described in detail. Finally, the article provides an in-depth case study of how one turnaround company reestablished the financial security of a nine-physician internal medicine group within a 2-year period. PMID- 10725047 TI - Supreme Court clarifies scope of the Americans with Disabilities Act. PMID- 10725048 TI - Liability of managed care organizations expands. PMID- 10725049 TI - Legal considerations in referral to alternative medicine. PMID- 10725050 TI - Avoiding personal data loss. AB - The potential personal, financial, emotional, and professional costs associated with the loss of data stored in personal computing devices are difficult to appreciate a priori. Because of the potentially devastating consequences of significant data loss, it behooves all clinicians to take personal responsibility in securing their data, whether or not this responsibility is nominally assumed by Information Systems (IS) professionals. There are a variety of straightforward, easily implemented approaches that can be used to help secure personal data, including investigating IS department policies, following proper backing-up procedures, observing reasonable security precautions, keeping digital media current, and establishing a process for executing these approaches. PMID- 10725051 TI - Introduction: public and private sector collaboration in the field of contraceptive research and development. PMID- 10725052 TI - Tort liability and the availability of contraceptive drugs and devices in the United States. PMID- 10725053 TI - Protecting consumers, prodding companies, and preventing conception: toward a model act for no fault liability for contraceptives. PMID- 10725054 TI - Litigation rules and culture: the European perspective. PMID- 10725055 TI - Disbursement of indemnity for injuries related to reproductive drugs and devices: a Swedish perspective. PMID- 10725056 TI - Contraceptive update. PMID- 10725057 TI - Comparison of the availability of contraceptive methods in selected European countries and the United States. PMID- 10725059 TI - Managing reactive arthritis. PMID- 10725058 TI - Detection and differentiation of Campylobacter jejuni and Campylobacter coli in broiler chicken samples using a PCR/DNA probe membrane based colorimetric detection assay. AB - Campylobacter enteritis in humans has been linked to consumption of poultry meat. Surveys show that 30-100% of poultry harbour Campylobacter as normal flora of the digestive tract which indicates a need to identify prevalent organism types in flocks and trace their epidemiology. In this study we describe a Campylobacter genus specific polymerase chain reaction (PCR) assay, amplifying the 16 S-23 S rRNA intergenic spacer region with an internal Campylobacter genus specific DNA probe and species specific probes for Campylobacter jejuni and Campylobacter coli designed for confirmation of the amplified PCR products by Southern blot and colorimetric reverse hybridization assays. The specificity of this assay was established by testing a range of food pathogens. Broiler chicken samples were tested following presumptive positive identification by the Malthus System V analyser (Malthus Instruments, UK). The combined PCR and colorimetric reverse hybridization assay is easy to perform and faster than conventional methods for confirmation and identification of Campylobacter species. PMID- 10725060 TI - Apoptosis-new clues to the pathogenesis of Sjogren's syndrome? PMID- 10725061 TI - The links between joint damage and disability in rheumatoid arthritis. AB - OBJECTIVE: The characteristic joint damage and disability of rheumatoid arthritis (RA) increase slowly over 10-20 yr. Although it is generally believed that persisting inflammatory synovitis causes joint damage and subsequent disability, the strength of their relationship has not been systematically evaluated. This review describes their progression and interrelationship in treated RA. METHODS: MEDLINE and Current Contents databases were searched for the combined terms of rheumatoid arthritis AND X-rays, Health Assessment Questionnaire, slow-acting anti-rheumatic drugs and all identifiable synonyms. This search identified 1303 articles and from these we evaluated in detail 23 reports on the progression of joint damage, 12 reports on the progression of disability and 25 reports dealing with their interrelationship. Additional information was obtained from four data sets comprising 725 RA patients studied cross-sectionally and 33-126 cases followed prospectively for 1-5 yr. X-ray damage was primarily assessed by Larsen and Sharp indices, and disability by the Health Assessment Questionnaire (HAQ). RESULTS: Joint damage and disability both increase throughout the duration of RA. Although disability (HAQ score) is correlated with disease duration (correlation coefficients between 0.27 and 0.30), the link between X-ray damage and disability is stronger (correlation coefficients between 0.30 and 0.70). In the earliest phases of RA, X-ray damage and HAQ scores are not related. By 5-8 yr, there are significant correlations with correlation coefficients between 0.30 and 0.50. In late RA (>8 yr), most studies show highly significant correlations between 0.30 and 0.70. CONCLUSIONS: Joint damage progresses constantly over the first 20 yr of RA. It accounts for approximately 25% of disability in established RA. The link between damage and disability is strongest in late (>8 yr) RA. However, avoiding or reducing joint damage in both early and established/late RA is likely to maintain function. PMID- 10725062 TI - Links between complement abnormalities and systemic lupus erythematosus. PMID- 10725063 TI - Antibodies to CD4 in primary Sjogren's syndrome. AB - OBJECTIVE: The purpose of this study was to examine whether antibodies against CD4 are present in patients with primary Sjogren's syndrome, and to explore the possible correlation between these antibodies and the CD4+ T lymphocyte depletion that is seen in some Sjogren patients. METHODS: Sera from 214 patients with primary Sjogren's syndrome, 154 healthy blood donors, 38 age- and sex-matched controls without autoimmune disease, and 77 HIV-1-seropositive individuals were analysed by an enzyme-linked immunosorbent assay (ELISA) using recombinant soluble CD4 as the antigen. RESULTS: Anti-CD4 antibodies were observed more frequently in patients with Sjogren's syndrome (12.6%) as compared with the control groups (0.6%) (P < 0.001), and at a level similar to that seen among the HIV-1 patients (13.0%). However, no correlation was found between the presence of anti-CD4 antibodies and CD4+ T lymphocytopenia in the Sjogren patients. CONCLUSION: This is the first study that shows anti-CD4 antibodies in patients with primary Sjogren's syndrome. The significance of these antibodies in the immunopathogenesis of Sjogren's syndrome remains to be determined. PMID- 10725064 TI - Development of a disease activity index for the assessment of reactive arthritis (DAREA). AB - OBJECTIVES: The objectives of this study were to investigate and validate individual variables and to develop a composite score for disease activity measurement in patients with reactive arthritis (REA). METHODS: In the first cross-sectional part, the clinical and laboratory evaluation of 45 patients was used to elaborate the most important individual disease activity measures. In the second prospective part, these variables as well as a composite score for disease activity measurement of REA were prospectively validated in 23 patients at two points in time. RESULTS: The following variables emerged as the most useful for the composite measure: number of swollen and tender joints, patient's pain and global assessment, and C-reactive protein. The score was calculated by simple addition of the individual figures. CONCLUSION: DAREA constitutes a reliable score which can easily be assessed on a day-to-day office work basis. PMID- 10725065 TI - Microemulsion formulation of cyclosporin (Sandimmun Neoral) vs Sandimmun: comparative safety, tolerability and efficacy in severe active rheumatoid arthritis. On behalf of the OLR 302 Study Group. AB - OBJECTIVE: To compare the safety, tolerability and efficacy of the new oral microemulsion formulation of cyclosporin A (CyA; Sandimmun Neoral) and the original CyA formulation (Sandimmun), in patients with severe active rheumatoid arthritis (RA), over a 12-month period. METHODS: In this double-blind, multicentre study, patients were randomized to treatment with Neoral or Sandimmun, starting with 2.5 mg/kg/day, with dose adjustments after 4 weeks. Primary efficacy criteria included patients' assessment of disease activity. Pharmacokinetic and safety assessments were performed at regular intervals. RESULTS: Compared with Sandimmun, Neoral showed a consistent trend towards greater clinical efficacy from week 12 onwards, including a significant difference in patients' assessment of disease activity at the study end-points. A significantly lower increase in dose from baseline was observed with Neoral at week 24. Pharmacokinetic assessments at week 24 showed increased absorption and decreased variability with Neoral. No differences in safety were found between treatment groups. CONCLUSION: These observations indicate that Neoral is as safe and at least as effective as Sandimmun and have important implications for patient management given the increasing role for CyA in the treatment of severe, active RA. PMID- 10725066 TI - Evaluation of filtration leucocytapheresis for use in the treatment of patients with rheumatoid arthritis. AB - OBJECTIVES: To evaluate the efficacy of filtration leucocytapheresis (LCP) for rheumatoid arthritis (RA). METHODS: LCP was carried out three times, with 1 week separating each session, in 25 drug-resistant RA patients. RESULTS: During each session, 96, 98, 61, 84 and 8% of the granulocytes, monocytes, lymphocytes, platelets and erythrocytes, respectively, that entered the LCP filter were removed. The number of granulocytes, monocytes and lymphocytes in the peripheral blood significantly decreased during each session of LCP. However, there was no significant decrease in the number of circulating blood cells during the study period. On average, 110 x 10(8) granulocytes, 5.23 x 10(8) monocytes, and 20.5 x 10(8) lymphocytes were removed during LCP therapy. Assessment of RA before and after LCP showed a substantial and rapid improvement in the tender joints counts, swollen joint counts, and patient's and physician's assessments. No adverse reactions or complications were noted. Erythrocyte sedimentation rate (ESR) and C reactive protein (CRP) levels decreased following LCP, although the change in the latter parameter was statistically insignificant. The concentrations of serum albumin, gamma-globulin, IgG, IgM, CH50 and rheumatoid factor titres did not change during or after LCP. Careful analysis indicated that 16 of 25 patients with RA showed > or =20% improvement following LCP therapy. CONCLUSIONS: Our results suggest that filtration LCP to remove leucocytes from the peripheral blood exerts an immunomodulatory effect in patients with RA. PMID- 10725067 TI - Localized pigmented villonodular synovitis of the knee joint: neoplasm or reactive granuloma? A review of 18 cases. AB - OBJECTIVE: The localized form of pigmented villonodular synovitis of the knee joint is a rare disease with limited alteration of the synovial membrane, the pathogenesis of which is the subject of controversial discussion. METHODS: Eighteen cases have been documented in our hospital since 1976. All of the patients had additional cartilage or meniscus damage. Treatment consisted of excision of the lesion and the adjacent synovial membrane, as well as therapy of the additional damage. RESULTS: The patients who had received such therapy were followed for 3-9 yr, without any clinical, sonographic or magnetic resonance tomographic signs of recurrence. In addition to the lack of a tendency towards recurrence, none of the cases displayed any further characteristics of the diffuse form of villonodular synovitis, such as invasiveness or malignant transformation. CONCLUSIONS: We therefore suggest that pigmented villonodular synovitis of the knee joint should be classified more strictly than before into a potentially neoplastic (diffuse) form and a reactive granulomatous (local) form. From the cases observed, we conclude that degenerative joint lesions may be the cause of the reactive granulomatous form. PMID- 10725068 TI - Spinal osteomyelitis in patients with rheumatoid arthritis following total knee arthroplasty-two case reports. PMID- 10725069 TI - Dramatic response after an intravenous loading dose of azathioprine in one case of severe and refractory ankylosing spondylitis. AB - We describe a 37-yr-old Caucasian male suffering from ankylosing spondylitis (AS) with long-standing severe inflammatory lumbar pain and hip arthritis who was refractory to non-steroidal anti-inflammatory drugs, sulphasalazine and methotrexate up to 25 mg/week. In this patient, administration of an i.v. loading dose of azathioprine (AZA; 40 mg/kg for 36 h followed by 2 mg/kg oral AZA therapy) induced a dramatic response in his clinical condition. Indeed, objective and subjective clinical variables improved within 1 week and were corroborated by a decline in the levels of the inflammatory parameters; anaemia was reported at month 3 but was rapidly reversible. If confirmed, an i.v. loading dose of AZA could represent a valuable alternative in severe and refractory AS, but toxicity of this regimen should be carefully analysed. PMID- 10725070 TI - Mucosa-associated lymphoid tissue lymphomas in two patients with rheumatoid arthritis on second-line agents, and secondary Sjogren's syndrome. AB - We report two patients with rheumatoid arthritis and secondary Sjogren's syndrome whose disease was complicated by a mucosa-associated lymphoid tissue (MALT) lymphoma. Although this particular type of lymphoma is associated with primary Sjogren's syndrome, it has not been described, to our knowledge, in the context of rheumatoid arthritis and Sjogren's syndrome. The potential causative factors are discussed. PMID- 10725071 TI - Evidence for activation of the alternate complement pathway in patients with juvenile rheumatoid arthritis. AB - OBJECTIVE: Complement activation has been shown to occur in patients with juvenile rheumatoid arthritis (JRA). Since the two pathways of complement are activated by different stimuli (the alternate pathway by microbial products and IgA, and the classical pathway by immune complexes), we decided to study the relative contribution of the two pathways of complement activation in patients with JRA. METHODS: In 56 patients with JRA, plasma levels of C3 and C4 were measured by turbidimetric assays, and those of C4d, factor Bb and sC5-9 complex by solid-phase enzyme immunoassays. Levels beyond the mean +/- 2 S.D. of normal were considered abnormal. RESULTS: Plasma C3 and C4 levels were decreased in one patient each. The C4d values were increased in 17 patients, whereas levels of factor Bb were elevated in 42 patients and levels of sC5-9 complex were elevated in 51 patients. The values of factor Bb and sC5-9 had a linear correlation (r = 0.75), but there was no significant correlation between C4d and sC5-9 levels (r = 0.36). CONCLUSION: Complement activation in JRA is initiated predominantly by the alternate pathway and culminates in the formation of terminal membrane attack complex. PMID- 10725072 TI - Elevated serum transferrin receptor concentration in children with juvenile chronic arthritis as evidence of iron deficiency. AB - OBJECTIVES: Active juvenile chronic arthritis (JCA) is accompanied by anaemia of chronic disease, which may be indistinguishable from anaemia due to iron deficiency. We speculate that elevation of the serum transferrin receptor (sTfR) concentration, which should not be influenced by inflammation, would be useful for detecting the role of iron status in anaemic children with JCA. METHODS: sTfR concentrations were measured in 30 children with JCA. RESULTS: The median sTfR was elevated, 6.1 (range 3.4-13.0) mg/l. In 13 patients (43%) the concentrations exceeded the upper limit for healthy subjects. Haemoglobin (r = - 0.48, P = 0.008), mean corpuscular volume (r = - 0.47, P = 0.009) and mean corpuscular haemoglobin (r = - 0.65, P = 0.0001) correlated inversely with sTfR concentration. CONCLUSIONS: In 13 of the 30 patients with JCA, the sTfR concentration, which is an indicator of iron status and erythropoiesis, was elevated. The results raise the possibility that sTfR is able to distinguish iron deficiency anaemia from anaemia of chronic disease. It should be further explored as a candidate. PMID- 10725073 TI - Juvenile chronic arthritis into adulthood: a long-term follow-up study. AB - OBJECTIVE: To evaluate a group of 65 adults with a history of or persistent juvenile chronic arthritis (JCA), on average, 26.4 yr after disease onset. METHODS: Disease status at the time of the study included an evaluation of disease-related parameters assessed by the patient and the investigator. RESULTS: Active disease was present in 37% of the study participants, of which 80% had either extended pauciarticular or polyarticular JCA. Eleven per cent of the study subjects were in Steinbrocker functional classes III and IV and 22% had undergone JCA-related major surgery. The pain visual analogue scale, health assessment questionnaire, erythrocyte sedimentation rate and C-reactive protein (CRP) were significantly increased in those participants who had active JCA at the time of the study. Disease duration proved to be the parameter most strongly associated with an unfavourable disease outcome. CONCLUSIONS: Although the study group was biased towards the more severe cases, the data suggest that the long-term functional outcome in JCA is, in more than one-third, associated with active disease persisting into adulthood, increasing residua and the need for surgery. PMID- 10725074 TI - Papulonecrotic tuberculide and stenosis of the abdominal aorta. AB - Papulonecrotic tuberculide (PNT) is a rare form of skin tuberculosis affecting predominantly young adults, with a history of immunity to Mycobacterium tuberculosis. We report a case of a young Caucasian female with PNT who was also documented to have a stenotic segment in the abdominal aorta. The difficulty in clarifying and treating the primary disease and the association between a tuberculous infection and Takayasu's arteritis are discussed. PMID- 10725075 TI - Factor V Leiden and venous thrombosis in a 4-yr-old girl with Behcet's syndrome. PMID- 10725076 TI - Idiopathic focal myositis in pregnancy. PMID- 10725077 TI - Psoriasis and diffuse systemic sclerosis: a report of three patients. PMID- 10725078 TI - Devic's neuromyelitis optica: a primary autoimmune disease? PMID- 10725079 TI - Reply PMID- 10725080 TI - Re: Hueber et al. Sensitivity and specificity of anti-Sa autoantibodies for rheumatoid arthritis. PMID- 10725081 TI - Reply PMID- 10725082 TI - Remission from lupus nephritis resistant to cyclophosphamide after additional treatment with cyclosporin A. PMID- 10725084 TI - Reply PMID- 10725083 TI - Re: Tager and Tikly. Clinical and laboratory manifestations of systemic sclerosis (scleroderma) in black South Africans. PMID- 10725086 TI - Reply PMID- 10725085 TI - Lethal aneurysm formation of pulmonary arteries in a woman with Behcet's disease. PMID- 10725087 TI - Bilateral gluteal abscesses as a unique manifestation of fusobacterium septicaemia. PMID- 10725088 TI - Poly(ethylene glycol) multiblock copolymer as a carrier of anti-cancer drug doxorubicin. AB - The synthesis of a novel water-soluble polymer drug carrier system based on biodegradable poly(ethylene glycol) block copolymer is described in this paper. The copolymer consisting of PEG blocks of molecular weight 2000 linked by means of an oligopeptide with amino end groups was prepared by interfacial polycondensation of the diamine and PEG bis(succinimidyl carbonate). The structure of the oligopeptide diamine consisting of glutamic acid and lysine residues was designed as a substrate for cathepsin B, a lysosomal enzyme, which was assumed to be one of the enzymes responsible for the degradation of the polymer carrier in vivo. Each of the oligopeptide blocks incorporated in the carrier contained three carboxylic groups of which some were used for attachment of an anti-cancer drug, doxorubicin (Dox), via a tetrapeptide spacer Gly-Phe-Leu Gly. This tetrapeptide spacer is susceptible to enzymatic hydrolysis. In vitro release of Dox and the degradation of the polymer chain by cathepsin B as well as preliminary evaluation of in vivo anti-cancer activity of the conjugate are also demonstrated. PMID- 10725089 TI - Site-specifically labeled photoprotein-thyroxine conjugates using aequorin mutants containing unique cysteine residues: applications for binding assays (Part II). AB - The jellyfish Aequorea victoria produces a protein, aequorin, which belongs to the class of Ca(2+)-dependent photoproteins known for their ability to emit visible light. This property of aequorin has allowed for its as a bioluminescent label in binding assays for a variety of analytes. Due to the excellent detection limits we demonstrated in assays for small peptides using a fusion protein between the peptide of interest and the photoprotein, our next goal was to expand the range of possible analytes for producing homogeneous populations of conjugates with the aequorin label to those that were nonpeptidic in nature. Recently, we prepared and characterized four aequorin mutants containing unique cysteine residues at various positions in the polypeptide chain. In the work reported here, the four aequorin mutants were each conjugated with a maleimide activated methyl ester derivative of thyroxine, a hormone frequently determined to evaluate thyroid function. The thyroxine-aequorin mutant conjugates were characterized in terms of the bioluminescence activities and binding properties with an anti-thyroxine monoclonal antibody for possible future employment in either heterogeneous or homogeneous binding assays for thyroxine and/or other desired analytes. PMID- 10725090 TI - Electrostatically driven immobilization of peptides onto (Maleic anhydride-alt methyl vinyl ether) copolymers in aqueous media. AB - The covalent immobilization of a model peptide onto the MAMVE copolymer, via the formation of amide bonds, occurred in moderate yields in aqueous conditions. The improvement of the grafting reaction was achieved by adding at the amino terminus of the model peptide a sequence (tag) of three positively charged amino acids, lysine or arginine, and by taking profit of electrostatic attractive interactions between the negatively charged copolymer and the tagged peptides. The arginine tag was more efficient than the lysine tag for enhancing the immobilization reaction, proving that the effect was due to an electrostic driving force. On the basis of these results, a tentative mechanism is discussed, and Scatchard plots pointed out two regimes of binding. With the first, at low polymer load (up to 50% of saturation for a lysine tag and 60-70% for an arginine tag), the binding occurred with a positive cooperative effect, the already bound peptide participating to the binding of others. A second one for higher coverages, for which the binding occurred with a negative cooperativity, and saturation was reached in the presence of a large excess of peptide. PMID- 10725092 TI - Preparation of thiol-reactive Cy5 derivatives from commercial Cy5 succinimidyl ester. AB - The present study offers reliable protocols for the preparation of new thiol reactive Cy5 derivatives which are urgently needed for single molecule fluorescence microscopy. In a systematic approach, two alternate strategies were found for the extension of commercial amine-reactive Cy5 with thiol-reactive end groups. In the two-step method, Cy5 succinimidyl ester was first reacted with ethylenediamine under conditions which gave approximately 99% asymmetric "Cy5 amine" and only approximately 1% symmetric product with two Cy5 residues. Subsequently, "Cy5-amine" was derivatized with commercial heterobifunctional cross-linkers to introduce thiol-reactive end groups (maleimide or pyridyldithio). Alternatively, commercial Cy5 succinimidyl ester was reacted with a primary amine (MTSEA, methanethiosulfonylethylamine, or PDEA, pyridyldithioethylamine) or a secondary amine (PEM, piperazinylethylmaleimide) to give the corresponding thiol-reactive derivatives in a single step. Results were good for MTSEA, moderate for PEM, and poor for PDEA. An additional drawback of the one-step method was the need for rigorous removal of unreacted Cy5 succinimidyl ester, which would label lysine residues on probe molecules. It is concluded that, except for the Cy5-MTSEA conjugate, the two-step method is much more general, reliable, and easier to follow by the typical biophysicist, biologist, etc., for whose benefit, these procedures are being published. All thiol-reactive Cy5 derivatives showed similar absorption and fluorescence properties as Cy5 succinimidyl ester, and fluorescence was fully retained after binding to thiols on proteins. The kinetics of protein labeling was also examined in order to get an idea of proper labeling conditions. PMID- 10725091 TI - 3'-End conjugates of minimally phosphorothioate-protected oligonucleotides with 1 O-hexadecylglycerol: synthesis and anti-ras activity in radiation-resistant cells. AB - Activation of the ras oncogene has been implicated in many types of human tumors. It has been shown that downmodulation of ras expression can lead to the reversion of the transformed phenotype of these tumor cells. Antisense oligodeoxyribonucleotides (ODNs) can inhibit gene expression by hybridization to complementary mRNA sequences. To minimize toxicity associated with all phosphorothioated ODNs and improve cellular uptake, we used partially phosphorothioate (PPS)-modified ODNs having an additional hydrophobic tail at the 3'-end (PPS-C(16)). The PPS ODNs are protected against degradation by PS internucleotide linkages at both the 3'- and 5'-ends and additionally stabilized at internal pyrimidine sites, which are the major sites of endonuclease cleavage. Here we show that anti-ras PPS-C(16) ODN retains the high sequence-specificity of PPS ODNs and provides maximal inhibition of Ras p21 synthesis with minimal toxicity even without the use of a cellular uptake enhancer. Moreover, treatment of T24, a radiation-resistant human tumor cell line that carries a mutant ras gene, with anti-ras PPS-C(16) ODN resulted in a reduction in the radiation resistance of the cells in vitro. We also demonstrate that the growth of RS504 (a human c-Ha-ras transformed NIH/3T3 cell line) mouse tumors was significantly inhibited by the combination of intratumoral injection of anti-ras PPS-C(16) ODN and radiation treatment. These findings indicate the potential of this combination of antisense and conventional radiation therapy as a highly effective cancer treatment modality. PMID- 10725093 TI - Structural characterization, kinetic studies, and in vitro biological activity of new cis-diamminebis-cholylglycinate(O,O') Pt(II) and cis-diamminebis ursodeoxycholate(O,O') Pt(II) complexes. AB - The complexes cis-diamminebis-cholylglycinate (O,O') [Pt(II) C(52)H(90)N(4)O(12)Pt, for convenience referred to as Bamet-R1] and cis diamminebis-ursodeoxycholate (O,O') Pt(II) (C(48)H(84)N(2)O(8)Pt, Bamet-UD2) were prepared. The structural integrity of the compounds was confirmed by elemental analysis, FT-IR, NMR, FAB-MS, and UV spectroscopies. The kinetic study of both compounds was accomplished by combining the conductivity measurement and those of the analysis of the electronic spectra in aqueous solution for NaCl concentrations of 4 mM (similar to cytoplasmatic concentration), 150 mM (similar to plasmatic concentration), and 500 mM. In water, the compound Bamet-R1 showed a half-life, t(1/2), of 3.0 h. This compound forms the chelate species through loss of a ligand, and the other one acts as a bidentate ligand. Ring opening in the presence of chloride ion was produced with a k(Cl)()-of 0.25 M(-)(1) h(-)(1). The half-life of Bamet-UD2 in aqueous solution was 3.2 h. However, since this species is not able to chelate and has a lower degree of solubility in the presence of chloride ion, its kinetic behavior was very different from that of the other compound. We consider this to be of great interest with regards to its cytostatic activity. All kinetic measurements were performed under pseudo-first-order conditions, and a pseudo-first-order behavior was found. The antitumoral effect of Bamet-UD2 on several cell lines derived from rat hepatoma, human hepatoma, mouse leukemia, and human colon carcinoma was found to be, in general, similar to that of cisplatin, but higher than that observed for Bamet-R1. PMID- 10725094 TI - Single isomer technetium-99m tamoxifen conjugates. AB - To produce an imaging agent for breast cancer using a technetium-99m-labeled agent specific for estrogen receptors, an N(2)S(2) bifunctional chelator was conjugated to Z- and E-aminotamoxifens through an amide linker. These bioconjugates have been chelated with both technetium-99m and rhenium. For the Z isomer, chelation with rhenium in the presence of sodium acetate yields a mixture of two isomers, anti and syn, in a 1:1 ratio and in the presence of hydroxide results in only the anti isomer. Both the Z- and E-tamoxifen conjugates have been chelated with technetium-99m at the tracer level yielding a single isomer product, which is assigned as anti based on chromatographic comparison to the rhenium complexes. Radiochemical yields were consistently greater than 80%, with Sep-Pak column purification yielding a final product with >99% radiochemical purity and no residual starting material. Both in vitro and in vivo biological evaluation of the tamoxifen chelates indicated very limited estrogen receptor binding. PMID- 10725095 TI - Bioconjugation of ribonuclease A: a detailed chromatographic and mass spectrometric analysis of chemical modification by a cross-linking reagent. AB - The modification of ribonuclease A with the heterobifunctional cross-linker, 4 succinimdyloxycarbonyl-methyl-alpha-[2-pyridyldithio]-toluene (SMPT) is described. RNase A has 11 potential sites of modification by the SMPT reagent. Tracking the two-dimensional separation and proteolytic digestion of SMPT modified RNase A with ESI/FTICR-MS and HPLC/ESI/QIT-MS demonstrates the detailed information about number of SMPT modifications and sites of modification that can be obtained by application of these techniques. Analysis of native and modified RNase A tryptic digests by ESI/FTICR-MS resulted in the identification of the sites of modification. Semiquantitative results of the reactivity of certain lysine residues toward the coupling reagent SMPT are presented. Two sites (lysines 1 and 37) are highly reactive, while three sites (lysines 41, 61, and 104) appear to be unreactive toward SMPT under the conditions used. Experimental results demonstrate that quantitative comparison of relative intensities of peptide sequences of different charge states is not possible. No correlation was found between number of basic residues and sensitivity to detection. Digestion of the modified and unmodified RNase A by subtilisin followed by examination by HPLC/ESI/QIT-MS and MS(n) enabled further investigation of modification on lysines 1 and 7, including modification at the epsilon- and alpha-amino positions on lysine 1. PMID- 10725096 TI - Synthesis and characterization of poly(ethylene glycol)-insulin conjugates. AB - Human insulin was modified by covalent attachment of short-chain (750 and 2000 Da) methoxypoly (ethylene glycol) (mPEG) to the amino groups of either residue PheB1 or LysB29, resulting in four distinct conjugates: mPEG(750)-PheB1-insulin, mPEG(2000)-PheB1-insulin, mPEG(750)-LysB29-insulin, and mPEG(2000)-LysB29 insulin. Characterization of the conjugates by MALDI-TOF mass spectrometry and N terminal protein sequence analyses verified that only a single polymer chain (750 or 2000 Da) was attached to the selected residue of interest (PheB1 or LysB29). Equilibrium sedimentation experiments were performed using analytical ultracentrifugation to quantitatively determine the association state(s) of insulin derivatives. In the concentration range studied, all four of the conjugates and Zn-free insulin exist as stable dimers while Zn(2+)-insulin was exclusively hexameric and Lispro was monomeric. In addition, insulin (conjugate) self-association was evaluated by circular dichroism in the near-ultraviolet wavelength range (320-250 nm). This independent method qualitatively suggests that mPEG-insulin conjugates behave similarly to Zn-free insulin in the concentration range studied and complements results from ultracentrifugation studies. The physical stability/resistance to fibrillation of mPEG-insulin conjugates in aqueous solution were assessed. The data proves that mPEG(750 and 2000)-PheB1-insulin conjugates are substantially more stable than controls but the mPEG(750 and 2000)-LysB29-insulin conjugates were only slightly more stable than commercially available preparations. Circular dichroism studies done in the far ultraviolet region confirm insulin's tertiary structure in aqueous solution is essentially conserved after mPEG conjugation. In vivo pharmacodynamic assays reveal that there is no loss in biological activity after conjugation of mPEG(750) to either position on the insulin B-chain. However, attachment of mPEG(2000) decreased the bioactivity of the conjugates to about 85% of Lilly's HumulinR formulation. The characterization presented in this paper provides strong testimony to the fact that attachment of mPEG to specific amino acid residues of insulin's B-chain improves the conjugates' physical stability without appreciable perturbations to its tertiary structure, self-association behavior, or in vivo biological activity. PMID- 10725097 TI - Facile synthesis of stable and lectin-recognizable DNA-carbohydrate conjugates via diazo coupling. AB - Stable and lectin-recognizable DNA-carbohydrate conjugates were prepared by diazo coupling of lactose and cellobiose derivatives to fragmented salmon testes DNA. The diazo coupling is suggested to take place selectively to guanine bases since the amount of lactose moiety introduced was directly proportional to the G content of various DNAs with different G contents. According to the CD spectra, the conjugates bearing carbohydrate less than 25% content kept a typical B-type conformation similar to native DNA. The conjugates possessed higher melting temperature and stronger nuclease resistance both to exo- and endonucleases than native DNA. Gel shift assay and fluorescence binding assay showed that the DNA lactose conjugates were specifically bound to galactose-specific lectin RCA(120) with strong binding affinity (Ka = 10(4)-10(5) M(-1)) due to glycoside cluster effect. This facile method will be a useful protocol of molecular design for cell targeted gene therapy. PMID- 10725098 TI - Application to a cartilage targeting strategy: synthesis and in vivo biodistribution of (14)C-labeled quaternary ammonium-glucosamine conjugates. AB - As part of a cartilage targeting program based on the affinity of the quaternary ammonium (QA) moiety for cartilage, QA derivatives of D-glucosamine (DG), an antirheumatic drug exhibiting a natural tropism for cartilaginous tissues, were designed and evaluated by pharmacokinetic studies. Two QA-DG conjugates were synthesized and labeled with (14)C by cross-linking the QA entity (trimethylammonium or pyridinium) to [(14)C]DG via an amide bond in a two-step procedure. After intravenous injection to male Sprague-Dawley rats, the two (14)C labeled conjugates exhibited similar pharmacokinetic profiles, but their behavior clearly differed from that of unconjugated DG in several ways. (i) The tissue distribution for the conjugates was more restricted, with a decreased radioactivity level for whole tissues except for kidney, cartilage, and skin. (ii) The radioactivity concentrated more rapidly and strongly in cartilage for the conjugates than for DG for the short times after injection; on the other hand, 1 h after administration, the radioactivity level in cartilage was higher for DG, this result being consistent with the tropism already observed for this compound. (iii) Both conjugates were eliminated predominantly by the urinary route (85%); the radioactivity level in urine for DG was lower (45% of the injected dose), and significant (14)CO(2) was found in expired air, indicating metabolization and utilization of DG for energy-consuming processes. (iv) Blood and plasma kinetics studies displayed an enterohepatic cycle for DG, whereas for the QA conjugates, a rapid disappearance was observed. (v) HPLC analyses of plasma and urine indicated a low degree of metabolization for the conjugates, most of the radioactivity recovered in urine and plasma corresponding to the unchanged molecule. This study demonstrates that the introduction of the QA moiety on DG modifies its biodistribution and lends it a greater specificity for cartilage, at least for short times after injection. These findings justify further work on QA derivatives of other antirheumatic agents. PMID- 10725099 TI - Sequence-recognition and cleavage of DNA by a netropsin-phenazine-di-N-oxide conjugate. AB - We report the synthesis, DNA-binding and cleaving properties, and cytotoxic activities of R-128, a hybrid molecule in which a bis-pyrrolecarboxamide-amidine element related to the antibiotic netropsin is covalently tethered to a phenazine di-N-oxide chromophore. The affinity and mode of interaction of the conjugate with DNA were investigated by a combination of absorption spectroscopy, circular dichroism, and electric linear dichroism. This hybrid molecule binds to AT-rich sequences of DNA via a bimodal process involving minor groove binding of the netropsin moiety and intercalation of the phenazine moiety. The bidentate mode of binding was evidenced by linear dichroism using calf thymus DNA and poly(dA dT).(dA-dT). In contrast, the drug fails to bind to poly(dG-dC).poly(dG-dC), because of the obstructive effect of the guanine 2-amino group exposed in the minor groove of this polynucleotide. DNase I footprinting studies indicated that the conjugate interacts preferentially with AT-rich sequences, but the cleavage of DNA in the presence of a reducing agent can occur at different sequences not restricted to the AT sites. The main cleavage sites were detected with a periodicity of about 10 base pairs corresponding to approximately one turn of the double helix. This suggests that the cleavage may be dictated by the structure of the double helix rather than the primary nucleotide sequence. The conjugate which is moderately toxic to cancer cells complements the tool box of reagents which can be utilized to produce DNA strand scission. The DNA cleaving properties of R 128 entreat further exploration into the use of phenazine-di-N-oxides as tools for investigating DNA structure. PMID- 10725100 TI - Solution structure of an LNA hybridized to DNA: NMR study of the d(CT(L)GCT(L)T(L)CT(L)GC):d(GCAGAAGCAG) duplex containing four locked nucleotides. AB - We have used two-dimensional (1)H NMR spectroscopy at 750 MHz to determine a high resolution solution structure of an oligonucleotide containing restricted nucleotides with a 2'-O, 4'-C-methylene bridge (LNA) hybridized to the complementary DNA strand. The LNA:DNA duplex examined contained four thymidine LNA modifications (T(L), d(C1T(L)2G3C4T(L)5T(L)6C7T(L)8G9C10):d( G11C12A13G14A15A16G17C 18A19G20). A total relaxation matrix approach was used to obtain interproton distance bounds from NOESY cross-peak intensities. These distance bounds were used as restraints in molecular dynamics (rMD) calculations. Forty final structures were generated for the duplex from A-form and B-form DNA starting structures. The root-mean-square deviation (RMSD) of the coordinates for the 40 structures of the complex was 0.6 A. The sugar puckerings are averaged values of a dynamic interchange between N- and S-type conformation except in case of the locked nucleotides that were found to be fixed in the C3'-endo conformation. Among the other nucleotides in the modified strand, the furanose ring of C7 and G9 is predominantly in the N-type conformation whereas that of G3 is in a mixed conformation. The furanose rings of the nucleotides in the unmodified complementary strand are almost exclusively in the S-type conformation. Due to these different conformations of the sugars in the two strands, there is a structural strain between the A-type modified strand and the B-type unmodified complementary strand. This strain is relaxed by decreasing the value of rise and compensating with tip, buckle, and propeller twist. The values of twist vary along the strand but for a majority of the base pairs a value even lower than that of A-DNA is observed. The average twist over the sequence is 32+/ 1 degrees. On the basis of the structure, we conclude that the high stability of LNA:DNA duplexes is caused by a local change of the phosphate backbone geometry that favors a higher degree of stacking. PMID- 10725101 TI - Concepts for the syntheses of biotinylated steroids. Part I: testosterone derivatives as immunochemical probes. AB - We describe synthetic strategies for the biotinylation of testosterone (T) at positions 3, 7alpha, 17alpha, and 19. These T probes are able to mimic ligand binding and may provide for a better understanding of the biospecific interaction with steroid-binding proteins such as the androgen receptor, anti-steroid antibodies, or steroid-binding serum globulins. For the 7alpha- and 17alpha derivatives, biotinyl-N-hydroxy-succinimide esters with different types of spacer chains were used. The 3-biotin hydrazone derivative was produced using N-(epsilon biotinyl)-caproyl hydrazide, whereas for the 19-biotinylation, a biotinyl-1-N diamino-3, 6-dioxaoctane-amide was applied. Key reaction for the biotinylation at position 3 is the oximation of the 3-oxo function. The 17alpha-position is accessible by the reaction of the 3-protected 4-androsten-17-epoxide with oxygen in the beta-position, followed by nucleophilic ring opening with cyanide which provides the 17alpha-cyanomethyl derivative. The key step is the regioselective ketal protection of the 3-oxo function of androst-4-ene-3,17-dione using a stannoxane catalyst. An alternative pathway for the insertion of biotin at the 19 position was established by the synthesis of 17beta-hydroxy-androst-4-en-3-one-19 yl carboxymethyl ether. After activation by the carbodiimide method, the compound reacts with aminoterminal biotin derivatives. The copper(I)-catalyzed 1,6 Michael addition of 17-acetoxy-6,7-dehydro-T leads to 7alpha-derivatives by use of omega silyl protected hydroxylalkyl-modified Grignard reagents. A functional group interconversion using the Staudinger reaction transforms the azide function into a primary omega-amino group. The absolute configurations of the different biotinylated derivatives were investigated by (1)H NMR studies. For the 7alpha biotinylated T series, additionally, an X-ray analysis proved the axial position of the spacer group. This results in a vertical orientation of the biotin moiety toward the alpha-face of the planar tetracyclic backbone. Thus, a negligible alteration of the original structure of the upper beta-face offers the feasibility of applying the 7alpha-derivatives as optimal immunochemical tracers in competitive immunoassays. Biotinylated T derivatives should be also suitable for ligand-binding studies to the androgen receptor or to sex hormone-binding globulin. PMID- 10725102 TI - Synthesis of [(99m)Tc]DTPA-folate and its evaluation as a folate-receptor targeted radiopharmaceutical. AB - A DTPA-folate conjugate was radiolabeled with (99m)Tc by stannous chloride reduction of [(99m)Tc]sodium pertechnetate in an aqueous solution of DTPA-folate. The radiochemical purity of the product consistently exceeded 97%, as assessed by thin-layer chromatography employing conditions analogous to those for radiochemical quality control of the radiopharmaceutical [(99m)Tc]DTPA. HPLC demonstrated that the radiolabeled product resulted from the intact DTPA-folate conjugate and not unconjugated DTPA. The ability of [(99m)Tc]DTPA-folate to target folate receptors in vivo was assessed in biodistribution studies with athymic mice bearing subcutaneous folate-receptor-positive human KB cell tumors. As an internal control, previously studied [(111)In]DTPA-folate was coinjected with the [(99m)Tc]DTPA-folate, along with varying amounts of DTPA-folate (0.38 mg/kg, 1.6 mg/kg, or 14 mg/kg). At each DTPA-folate dose, [(99m)Tc]DTPA-folate exhibited tumor uptake comparable to that of the coadministered [(111)In]DTPA folate, with radiotracer levels declining at the higher DTPA-folate doses due to competitive receptor binding of the unlabeled conjugate. Tumor uptake of both tracers was also competitively blocked by preadministered folic acid dihydrate (2.9 mg/kg). Tumor-to-background tissue contrast obtained with [(99m)Tc]DTPA folate was generally similar to that obtained with [(111)In]DTPA-folate. The (99m)Tc-labeled DTPA-folate conjugate may have utility as a targeted radiopharmaceutical for imaging neoplastic tissues known to overexpress the folate receptor. PMID- 10725103 TI - Efficient clearance of poly(ethylene glycol)-modified immunoenzyme with anti-PEG monoclonal antibody for prodrug cancer therapy. AB - The F(ab')(2) fragment of the anti-TAG-72 antibody, B72.3, was covalently linked to Escherichia coli-derived beta-glucuronidase that was modified with methoxypoly(ethylene glycol). The conjugate (B72.3-betaG-PEG) localized to a peak concentration in LS174T xenografts within 48 h after injection, but enzyme activity persisted in plasma such that prodrug administration had to be delayed for at least 4 days to avoid systemic prodrug activation and associated toxicity. Conjugate levels in tumors decreased to 36% of peak levels at this time. Intravenous administration of AGP3, an IgM mAb against methoxypoly(ethylene glycol), accelerated clearance of conjugate from serum and increased the tumor/blood ratio of B72. 3-betaG-PEG from 3.9 to 29.6 without significantly decreasing the accumulation of conjugate in tumors. Treatment of nude mice bearing established human colon adenocarcinoma xenografts with B72. 3-betaG-PEG followed 48 h later with AGP3 and a glucuronide prodrug of p-hydroxyaniline mustard significantly (p< or =0.0005) delayed tumor growth with minimal toxicity compared to therapy with a control conjugate or conventional chemotherapy. PMID- 10725104 TI - Binding properties of a monoclonal antibody directed toward lead-chelate complexes. AB - A monoclonal antibody (2C12) that recognizes a Pb(II) cyclohexyldiethylenetriamine pentaacetic acid complex was produced by the injection of BALB/c mice with a Pb(II)-chelate complex covalently coupled to a carrier protein. The ability of purified antibody to interact with a variety of metal-free chelators and metal-chelate complexes was assessed by measuring equilibrium dissociation constants. The antibody bound to metal-free trans cyclohexyldiethylenetriamine pentaacetic acid (CHXDTPA) with an equilibrium dissociation constant of 2.3 x 10(-)(7) M. Addition of Pb(II) increased the affinity of the antibody for the complex by 25-fold; Pb(II) was the only metal cation (of 15 different di-, tri-, and hexavalent metals tested) that increased the affinity of the antibody for CHXDTPA. The increased affinity was due primarily to an increase in the association rate constant. The antibody also had the ability to interact with ethylenediamine tetraacetic acid (EDTA), diethylenetriamine pentaacetic acid (DTPA), and structurally related derivatives, but with affinities from 50- to 10000-fold less than that determined for CHXDTPA. Addition of metals to EDTA-based chelators reduced the affinity of the antibody for these ligands. However, when DTPA was used as the chelator, addition of Pb(II) increased the affinity of the antibody for the complex by 200-fold. The sensitivity of prototype immunoassays for Pb(II) could be modulated by changing the structure of the immobilized metal-chelate complex and/or the soluble chelator used to complex Pb(II) in the test solution. PMID- 10725105 TI - Synthesis of biotinylated glyfoline for immunoelectron microscopic localization. AB - Antineoplastic glyfoline (1) has potent antitumor efficacy against various murine and human solid tumors. To elucidate the actual mechanism of action, we synthesized biotinylated glyfoline (B-Gly) and used it for the visualization of glyfoline-binding sites in nasopharyngeal carcinoma (NPC) cells. Under the electron microscope (EM), after cells were incubated for 6-36 h, the reaction products of anti-B-Gly were seen on some areas of the external cell surface and on the outer and inner membranes of the mitochondria. Pure EM morphology of NPC cells after glyfoline treatment revealed the similar morphological change of mitochondria. These findings indicate that the binding site of glyfoline in NPC is the inner membrane of the mitochondria, suggesting that B-Gly can be used as a marker for glyfoline localization. PMID- 10725106 TI - Solid-phase oligonucleotide synthesis and flow cytometric analysis with microspheres encoded with covalently attached fluorophores. AB - A novel combinatorial approach to synthesize oligonucleotides on fluorescently encoded microspheres based on flow sorting and segmental solid-phase synthesis is described. BODIPY dyes were covalently attached to polystyrene (8.8 microm, 55% DVB) microsphere particles to generate four fluorescently encoded sets. 20-mer oligonucleotide sequences can be synthesized on these microspheres with yields comparable to conventional CPG supports (80% overall yield, average stepwise yield = 99%). The concept of segmental solid-phase synthesis by flow sorting was demonstrated by synthesizing unique 20-mer oligonucleotide sequences on each of four fluorescently encoded microsphere sets by including a flow sorting step (after first eight base additions) and flow cytometric detection of sequences synthesized on each microsphere set by hybridization with fluorescently labeled complementary sequence. PMID- 10725107 TI - Use of a steroid cyclic disulfide anchor in constructing gold nanoparticle oligonucleotide conjugates. AB - A new anchoring group is described for binding oligonucleotides to gold surfaces. On the basis of a ketal derived from 4,5-dihydroxy-1, 2 dithiane and epiandrosterone, it is easy to prepare and to link to oligonucleotides. Gold nanoparticle-oligonucleotide conjugates made using this cyclic disulfide linker serve as effective probes for detecting specific oligonucleotide sequences, and they exhibit much greater stability toward dithiothreitol than corresponding conjugates prepared with the conventional mercaptohexyl group or an acyclic disulfide unit. The high stability toward thiol deactivation likely results, in part at least, from anchoring each oligonucleotide to gold through two sulfur atoms. PMID- 10725109 TI - Efficient multigram synthesis of the bifunctional chelating agent (S)-1-p isothiocyanatobenzyl-diethylene- tetramine pentaacetic acid PMID- 10725108 TI - Efficient multigram synthesis of the bifunctional chelating agent (S)-1-p isothiocyanatobenzyl-diethylenetriaminepentaacetic acid [correction of diethylenetetraminepentaacetic acid]. AB - We have developed and optimized the synthesis of the title compound, eliminating all HPLC purifications prior to the final product. The yield (and scale) of the synthesis was increased from 19% (200 mg) to 75% (26 g). PMID- 10725110 TI - Role of quinones in toxicology. AB - Quinones represent a class of toxicological intermediates which can create a variety of hazardous effects in vivo, including acute cytotoxicity, immunotoxicity, and carcinogenesis. The mechanisms by which quinones cause these effects can be quite complex. Quinones are Michael acceptors, and cellular damage can occur through alkylation of crucial cellular proteins and/or DNA. Alternatively, quinones are highly redox active molecules which can redox cycle with their semiquinone radicals, leading to formation of reactive oxygen species (ROS), including superoxide, hydrogen peroxide, and ultimately the hydroxyl radical. Production of ROS can cause severe oxidative stress within cells through the formation of oxidized cellular macromolecules, including lipids, proteins, and DNA. Formation of oxidatively damaged bases such as 8-oxodeoxyguanosine has been associated with aging and carcinogenesis. Furthermore, ROS can activate a number of signaling pathways, including protein kinase C and RAS. This review explores the varied cytotoxic effects of quinones using specific examples, including quinones produced from benzene, polycyclic aromatic hydrocarbons, estrogens, and catecholamines. The evidence strongly suggests that the numerous mechanisms of quinone toxicity (i.e., alkylation vs oxidative stress) can be correlated with the known pathology of the parent compound(s). PMID- 10725111 TI - Mossbauer spectroscopy indicates that iron in an aluminosilicate glass phase is the source of the bioavailable iron from coal fly ash. AB - Iron speciation by Mossbauer spectroscopy indicates that ferric iron in an aluminosilicate glass phase is the source of the bioavailable iron in coal fly ash and that this iron species is associated with combustion particles, but not with crustal dust derived from soil minerals. Urban particulate has been shown to be a source of bioavailable iron and has been shown to be able to induce the formation of reactive species in cell culture experiments. Crustal dust and laboratory-generated coal fly ash have been studied as surrogates for two sources of metal-bearing particles in ambient air. As much as a 60-fold difference in the amount of iron mobilized by the chelator citrate was observed between fly ash and crustal dust samples with similar total iron contents. The extent of iron mobilization by citrate in vitro has been shown to correlate with indirect measures of excess iron in cultured cells and with assays for reactive oxygen species generation in vitro. Mossbauer spectroscopy of coal fly ash, before and after treatment with the chelator desferrioxamine B, showed that the iron in an aluminosilicate glass phase was preferentially removed. The removal of the glass phase iron greatly reduced the amount of iron that could be mobilized by citrate and prevented the particles from inducing interleukin-8 in cultured human lung epithelial (A549) cells. Ferric iron in aluminosilicate glass is associated with particles formed at high temperatures followed by rapid cooling. The observation that ferric iron in aluminosilicate glass is the source of bioavailable iron in coal fly ash suggests that particles from ambient sources and other specific combustion sources should be examined for the presence of this potential source of bioavailable iron. PMID- 10725112 TI - Isolation and chemical--structural identification of a novel aromatic amine mutagen in an ozonized solution of m-phenylenediamine. AB - The mutagenicity of a m-phenylenediamine (m-PD) solution was markedly enhanced by oxidation with ozone. The ethyl acetate extracts from a m-PD solution ozonized at pH 10.7 were fractionated by normal-phase and reversed-phase column chromatography to isolate mutagens by monitoring mutagenic activities on Salmonella typhimurium TA98 in the presence of a mammalian metabolic activation system (S9 mix). From fraction 5-3-2, which exhibited the strongest mutagenicity (308000 revertants/mg), a major mutagenic compound was isolated. On the basis of the high-resolution EI-mass, (1)H NMR and (13)C NMR spectral, and X-ray crystallography data, the structure of this compound was determined to be 2-amino 5-[(3-aminophenyl)amino]-4-[(3-aminophenyl)imino]-2, 5-cyclohexadien-1-one (PDT 1), which is a novel compound. PDT-1 is a newly identified frame-shift type mutagen, inducing 65400 revertants and 295000 revertants of S. typhimurium TA98 and YG1024 per micromole, respectively, in the presence of S9 mix. When a m-PD solution was oxidized with 1 or 2 mol of ozone at pH 4.0, 7.0, and 10.7, the contribution of PDT-1 to the mutagenicity of ethyl acetate extracts from the ozonized m-PD solution was 5-23%. PMID- 10725113 TI - Evidence for the involvement of N-methylthiourea, a ring cleavage metabolite, in the hepatotoxicity of methimazole in glutathione-depleted mice: structure toxicity and metabolic studies. AB - In mice depleted of GSH by treatment with buthionine sulfoximine (BSO), methimazole (2-mercapto-1-methylimidazole, MMI) causes liver injury characterized by centrilobular necrosis of hepatocytes and an increase in serum alanine transaminase (SALT) activity. MMI requires metabolic activation by both P450 monooxygenase and flavin-containing monooxygenase (FMO) before it produces the hepatotoxicity. MMI and its analogues were examined for the ability to increase SALT activity in GSH-depleted mice. Saturation of the C-4,5 double bond in MMI resulted in a complete loss of hepatotoxicity. Similarly, ring fusion of a benzene nucleus to the C-4,5 double bond, forming 2-mercapto-1 methylbenzimidazole, abolished the toxic potency. As for MMI, 2-mercapto-1,4,5 trimethylimidazole, and 2-mercapto-1-methyl-4, 5-di-n-propylimidazole, the toxic potency decreased with the increasing bulk of the 4- and 5-alkyl substituents. Furthermore, methylation of the thiol group of MMI totally reduced its toxicity. These structural requirements and the known toxicity of thiono-sulfur compounds led us to the hypothesis that MMI would undergo epoxidation of the C-4,5 double bond by P450 enzymes and, after being hydrolyzed, the resulting epoxide would be then decomposed to form N-methylthiourea, a proximate toxicant. Before N methylthiourea would produce toxicity, it would be further biotransformed to its S-oxidized metabolites mainly by FMO. Evidence for this hypothesis was provided by the facts that N-methylthiourea and glyoxal as the accompanying fragment were identified as urinary metabolites in mice treated with MMI and that N methylthiourea caused a marked increase in SALT activity when administered to mice in combination with BSO. PMID- 10725114 TI - Structural identification of two metabolites of catechins and their kinetics in human urine and blood after tea ingestion. AB - Tea is a popular beverage consumed worldwide. The metabolic fate of its major constituents, catechins, however, is not well-known. In this study, two catechin metabolites were detected in the urine and plasma of human volunteers after ingestion of green tea. These metabolites were identified by LC/ESI-MS and NMR as (-)-5-(3',4', 5'-trihydroxyphenyl)-gamma-valerolactone (M4) and (-)-5-(3', 4' dihydroxyphenyl)-gamma-valerolactone (M6). The renal excretion of M4 and M6 had a 3 h lag time and peaked 7.5-13.5 h after ingestion of a single dose of green tea, while (-)-epigallocatechin (EGC) and (-)-epicatechin peaked at 2 h. M4 and M6 were two major tea metabolites with urinary cumulative excretions as high as 8-25 times the levels of EGC and (-)-epicatechin in some of our subjects, and accounted for 6-39% of the amounts of ingested EGC and (-)-epicatechin. Both the metabolites appeared to be produced by intestinal microorganisms, with EGC and ( )-epicatechin as the precursors of M4 and M6, respectively. Repeated ingestion of green tea produced a slight accumulative effect of the metabolites. They were also detected in the plasma, exhibiting kinetics similar to those of the urinary metabolites, and in the feces. Study on these metabolites may help us further understand the cancer chemopreventive actions and other beneficial effects of tea. PMID- 10725115 TI - Regioselectivity and reversibility of the glutathione conjugation of quercetin quinone methide. AB - The chemical reactivity, isomerization, and glutathione conjugation of quercetin o-quinone were investigated. Tyrosinase was used to generate the unstable quercetin o-quinone derivative which could be observed upon its subsequent scavenging by glutathione. Identification of the products revealed formation of 6 glutathionyl-quercetin and 8-glutathionyl-quercetin adducts. Thus, in particular, glutathione adducts in the A ring of quercetin were formed, a result which was not expected a priori. Quantum mechanical calculations support the possibility that the formation of these glutathione adducts can be explained by an isomerization of quercetin o-quinone to p-quinone methides. Surprisingly, additional experiments of this study reveal the adduct formation to be reversible, leading to interconversion between the two quercetin glutathione adducts and possibilities for release and further electrophilic reactions of the quercetin quinone methide at cellular sites different from those of its generation. PMID- 10725116 TI - Metabolism of N'-nitrosonornicotine enantiomers by cultured rat esophagus and in vivo in rats. AB - People who use tobacco products are exposed to considerable amounts of N' nitrosonornicotine (NNN), a well-established esophageal carcinogen in rats. NNN is believed to play a significant role as a cause of esophageal and oral cavity cancer in smokers and snuff dippers. The carcinogenicity of NNN is dependent on its metabolic activation. However, virtually all studies carried out to date on NNN metabolism have used racemic material. In this study, we examined the metabolism of [5-(3)H]-(S)-NNN and [5-(3)H]-(R)-NNN in cultured rat esophagus and in vivo in rats. Cultured rat esophagus metabolized (S)-NNN (1 microM) predominantly to products of 2'-hydroxylation, 4-oxo-4-(3-pyridyl)butanoic acid (keto acid) and 4-hydroxy-1-(3-pyridyl)-1-butanone (keto alcohol). In contrast, the major metabolite of (R)-NNN under these conditions was 4-hydroxy-4-(3 pyridyl)butanoic acid (hydroxy acid), a product of NNN 5'-hydroxylation. The 2' hydroxylation:5'-hydroxylation metabolite ratio ranged from 6.22 to 8.06 at various time intervals in the incubations with (S)-NNN, while the corresponding ratios were 1.12-1.33 in the experiments with (R)-NNN. These differences were statistically significant (P<0.001). Since 2'-hydroxylation is believed to be the major metabolic activation pathway of NNN in the rat esophagus, the results demonstrate that (S)-NNN is metabolically activated more extensively than (R)-NNN in this tissue, and therefore may be more carcinogenic. Rats were treated with 0.3 mg/kg of [5-(3)H]-(R)-NNN, [5-(3)H]-(S)-NNN, or racemic [5-(3)H]NNN by gavage, and the urinary metabolites were analyzed. The major metabolites were hydroxy acid and keto acid. As in the in vitro studies, products of 2' hydroxylation predominated in the urine of the rats treated with (S)-NNN while products of 5'-hydroxylation were more prevalent in the rats treated with (R) NNN. 2'-Hydroxylation:5'-hydroxylation metabolite ratios ranged from 1.66 to 2.04 in the urine at various times after treatment with (S)-NNN, while the ratios were 0.398-0.450 for the rats treated with (R)-NNN (P<0.001). The results of this study provide new insights into NNN metabolism in rats and suggest that the carcinogenicity of (S)-NNN, the predominant enantiomer in tobacco products, may be greater than that of (R)-NNN or racemic NNN. PMID- 10725117 TI - Characterization of the major DNA adduct formed by alpha-hydroxy-N desmethyltamoxifen in vitro and in vivo. AB - Tamoxifen is hepatocarcinogenic in rats and has been associated with an increased risk of endometrial cancer in women. Recent reports suggest that it may be genotoxic in humans. N-Desmethyltamoxifen is a major tamoxifen metabolite that has been proposed to be responsible for one of the major adducts detected in liver DNA of rats treated with tamoxifen. The metabolic activation of N desmethyltamoxifen to DNA binding products may involve oxidation to alpha-hydroxy N-desmethyltamoxifen followed by esterification. In the study presented here, we report the synthesis of alpha-hydroxy-N-desmethyltamoxifen and the characterization of the major adduct obtained from alpha-sulfoxy-N desmethyltamoxifen in vitro as (E)-alpha-(deoxyguanosin-N(2)-yl)-N desmethyltamoxifen. In addition, we use (32)P-postlabeling in combination with HPLC to compare the adducts formed in the livers of female Sprague-Dawley rats treated by gavage with tamoxifen or equimolar doses of alpha-hydroxy-N desmethyltamoxifen. We conclude that one of the major adducts formed in vivo and previously suggested to derive from N-desmethyltamoxifen is chromatographically identical to alpha-(deoxyguanosin-N(2)-yl)-N-desmethyltamoxifen. PMID- 10725118 TI - Synthesis of 8-(hydroxymethyl)-3,N(4)-etheno-2'-deoxycytidine, a potential carcinogenic glycidaldehyde adduct, and its site-specific incorporation into DNA oligonucleotides. AB - The previously unreported glycidaldehyde adduct, 8-(hydroxymethyl)-3, N(4)-etheno 2'-deoxycytidine (8-HM-epsilondC), has been synthesized for the first time by reaction of 2'-deoxycytidine with bromoacetaldehyde at pH 4.5, followed by reduction with sodium borohydride. The adduct was characterized by UV, MS, and NMR. The compound was stable to neutral and acidic conditions but not in alkaline solution. The corresponding phosphoramidite was synthesized in good yield from the intermediate, 3, N(4)-ethenocarbaldehyde-2'-deoxycytidine, using the standard methodology and site-specifically incorporated in both 15- and 25-mer oligonucleotides, for studies on biochemical and biophysical properties. The resulting oligonucleotides were purified using HPLC, and the base composition was verified by HPLC after enzymatic digestion. PMID- 10725119 TI - Editor's change of address PMID- 10725120 TI - Measurement of food flavonoids by high-performance liquid chromatography: A review. AB - The flavonoids are plant polyphenols found frequently in fruits, vegetables, and grains. Divided into several subclasses, they include the anthocyanidins, pigments chiefly responsible for the red and blue colors in fruits, fruit juices, wines, and flowers; the catechins, concentrated in tea; the flavanones and flavanone glycosides, found in citrus and honey; and the flavones, flavonols, and flavonol glycosides, found in tea, fruits, vegetables, and honey. Known for their hydrogen-donating antioxidant activity as well as their ability to complex divalent transition metal cations, flavonoids are propitious to human health. Computer-controlled high-performance liquid chromatography (HPLC) has become the analytical method of choice. Many systems have been developed for the detection and quantification of flavonoids across one, two, or three subclasses. A summary of the various HPLC and sample preparation methods that have been employed to quantify individual flavonoids within a subclass or across several subclasses are tabulated in this review. PMID- 10725121 TI - Effect of aldehyde lipid oxidation products on myoglobin. AB - The effects of aldehyde lipid oxidation products on myoglobin (Mb) were investigated at 37 degrees C and pH 7.2. Oxymyoglobin (OxyMb) oxidation increased in the presence of 4-hydroxynonenal (4-HNE) compared to controls (P < 0.05). Preincubation of metmyoglobin (MetMb) with aldehydes rendered the heme protein a poorer substrate for enzymatic MetMb reduction compared to controls, and the effect was inversely proportional to preincubation time; unsaturated aldehydes were more effective than saturated aldehydes (P < 0.05). The order of MetMb reduction as affected by preincubation was control > hexanal > heptanal > octanal > nonanal = decanal = hexenal > heptenal = octenal > nonenal = decenal = 4-HNE (P < 0.05). Preincubation of MetMb with 4-HNE enhanced the subsequent ability of the heme protein to act as a prooxidant in both liposomes and microsomes when compared to controls (P < 0.05); the effect was reduced in microsomes containing elevated concentrations of alpha-tocopherol (P < 0.05). MetMb preincubation with mono-unsaturated aldehydes enhanced the catalytic activity of MetMb to a greater degree than saturated aldehydes (P < 0.05). These results suggest that aldehyde lipid oxidation products can alter Mb stability by increasing OxyMb oxidation, decreasing the ability of MetMb to be enzymatically reduced and enhancing the prooxidant activity of MetMb. PMID- 10725122 TI - Phospholipid/fatty acid-induced secondary structural change in beta-lactoglobulin during heat-induced gelation. AB - Effects of phosphatidylcholine (PC) and the predominant fatty acids (FAs) in milk, butyrate, oleate, and palmitate, on secondary structural changes in beta lactoglobulin (beta-LG) during heat-induced gelation were analyzed on the basis of circular dichroism (CD) spectra. Small-strain oscillatory measurements were carried out to characterize viscoelastic properties of the heat-induced gels. In the absence of added salt, PC and FAs induced helix formation of beta-LG on heating to 80 degrees C and increased the storage moduli (G') of heat-induced gels. In the presence of 500 mM NaCl, PC did not change the CD spectrum of beta LG but decreased G'. In contrast, butyrate substantially unfolded beta-LG in 500 mM NaCl on heating, forming very elastic gels with increased G' values. Palmitate and oleate induced beta-LG gel formation at 25 degrees C without heating; heating to 80 degrees C almost completely unfolded beta-LG in 500 mM NaCl. PMID- 10725123 TI - Characterization of a monoclonal antibody specific for HMW subunits of glutenin and its use to investigate glutenin polymers. AB - A monoclonal antibody, IFRN 1602, has been developed to a synthetic peptide based on the sequence (94)GSVTCPQQV(101) of HMW subunit 1Dx5. The antibody bound strongly to the synthetic peptide based on the cognate sequence of HMW subunit 1Dx2 which contains a serine instead of a cysteine residue. However, it recognized the immunizing peptide by enzyme-linked immunosorbent assay (ELISA) only poorly, probably because the peptide exists as a disulfide-bonded dimer under the assay conditions. From immunoblotting studies against a wide range of wheat varieties, IFRN 1602 was shown to primarily recognize x-type HMW subunits of glutenin encoded on chromosomes 1A and 1D, cross-reacting weakly with the 1A and 1D y-type subunits. It did not bind to any of the 1B-encoded subunits. The Mab also recognized a small number of polypeptides of greater mobility than HMW subunits which were not visible on the stained gels and occurred only in the presence of specific 1A and 1D x-type HMW subunits. Such polypeptides were not present in a preparation of recombinant subunit 2, suggesting that they are modified forms of the subunits which arise in the seed perhaps by processing of the associated subunits. When used to probe partially reduced glutenin, IFRN 1602 bound to 1Dx5-1Dy10 dimers. As the Mab reacted primarily with Cys(97) of 1Dx5 in a reduced form, these data suggest that this residue is not involved in either intra- or intermolecular disulfide bond in the HMW subunit dimers. Thus, Cys(97) of 1Dx5 may be present in gluten in a reduced form, involved in intramolecular disulfide bonds, or linking of the HMW subunit dimers into larger polymers. PMID- 10725124 TI - Effect of phenolic structures on the degradability of cell walls isolated from newly extended apical internode of tall fescue (Festuca arundinacea Schreb.). AB - Apical internodes of tall fescue (Festuca arundinacea Schreb. var. Clarine) harvested at flowering were sectioned into 5 or 10 equal parts to study in situ degradability and cell wall composition, respectively. The basal (youngest) section had the greatest primary wall content. Cell walls in the upper (older) sections had the highest xylose/arabinose ratio and lignin content and a lignin rich in syringyl units, all typical of extensive secondary wall development. Almost all of the p-coumaric (p-CA) and about half of the ferulic acid (FA) were released by 1 M NaOH and presumed to be ester-linked. The total FA content was approximately double that of p-CA in all sections other than the youngest with a distribution similar to that of total p-CA. However, the ratio of esterified to ether and ether plus ester linked (Et & Et+Es) FA differed with age. Whereas the esterified form remained essentially constant ( approximately 4.5 g/kg of cell wall), Et & Et+Es ferulate increased with increasing age of the tissue and was significantly related to lignin deposition (r = 0.79, P < 0.01). The extent of cell wall degradation after 48 h of incubation in the rumen was inversely related to maturity, falling from 835 g/kg of dry matter in the youngest section to 396 g/kg in the oldest. Both the rate and extent of cell wall degradation were significantly negatively related to the ratio of xylose to arabinose, lignin content, proportion of syringyl units present in lignin, and concentration of Et & Et+Es FA present. A positive relationship between Et & Et+Es FA was also found, with the rate (P < 0.01) being better correlated than the extent (P < 0.05) of cell wall degradation. Application of the newly extended internode model to fescue produced results consistent with the view that both the lignin content and the extent to which lignin was covalently bound to the other wall polymers crucially influenced the rate and extent of degradation. PMID- 10725125 TI - Electrophoretic pattern, thermal denaturation, and in vitro digestibility of oxidized myosin. AB - Physicochemical changes and in vitro digestibility of chicken breast myosin oxidized with a nonenzymic free-radical-generating system (FeCl(3)/H(2)O(2)/ascorbate) were studied by SDS-PAGE, differential scanning calorimetry, and o-phthaldialdehyde assay. Oxidation caused fragmentation and polymerization of myosin. Myosin polymers were cross-linked mainly through disulfide bonds. Hydroxyl radicals destabilized myosin, lowering its denaturation temperature by up to 4 degrees C. Oxidized myosin also produced a new thermal transition in the 60-80 degrees C temperature range, which could be attributed to the formation of disulfide-stabilized polymers. The proteolytic susceptibility of myosin to pepsin, trypsin, and chymotrypsin was increased by oxidation. Under nonreducing conditions, however, oxidized myosin showed decreased digestibility. The results may help explain variations in the functionality and nutritional quality of muscle foods in meat processing in which oxidation is involved. PMID- 10725126 TI - Isothermal Fourier transform infrared microspectrosopic studies on the stability kinetics of solid-state intramolecular cyclization of aspartame sweetener. AB - A novel Fourier transform infrared (FT-IR) microspectrophotometer equipped with differential scanning calorimetry (DSC) was used to investigate the kinetics of intramolecular cyclization of aspartame (APM) sweetener in the solid state under isothermal conditions. The thermal-dependent changes in the peak intensity of IR spectra at 1543, 1283, and 1259 cm(-1) were examined to explore the reaction. The results support that the intramolecular cyclization process in APM proceeded in three steps: the methoxyl group of ester was first thermolyzed to release methanol, then an acyl cation was attacked by the lone pair of electrons available on nitrogen by an S(N)1 pathway, and finally ring-closure occurred. The intramolecular cyclization of APM determined by this microscopic FT-IR/DSC system was found to follow zero-order kinetics after a brief induction period. The bond cleavage energy (259.38 kJ/mol) of thermolysis for the leaving group of -OCH(3), the bond conversion energy (328.88 kJ/mol) for the amide II NH band to DKP NH band, and the CN bond formation energy (326.93 kJ/mol) of cyclization for the DKP in the APM molecule were also calculated from the Arrhenius equation. The total activation energy of the DKP formation via intramolecular cyclization was 261.33 kJ/mol, calculated by the above summation of the bond energy of cleavage, conversion, and formation, which was near to the value determined by the DSC or TGA method. This indicates that the microscopic FT-IR/DSC system is useful as a potential tool not only to investigate the degradation mechanism of drugs in the solid state but also to directly predict the bond energy of the reaction. PMID- 10725127 TI - Formation of sugar-specific reactive intermediates from (13)C-labeled L-serines. AB - Analysis of the pyrolysis products of [1-(13)C], [2-(13)C], and [3-(13)C]-labeled L-serines has indicated the presence of three initial degradation pathways. Decarboxylation followed by deamination produces aminoethanol and acetaldehyde, respectively; a retro-aldol pathway generates formaldehyde and glycine. Dehydration of L-serine can lead to the formation of pyruvic acid, which eventually can be converted into the amino acid alanine. Formation of alanine and glycine was confirmed due to the detection of 2, 5-diketo-3,6-dimethylpiperazine and cycloglycylalanine. Most of the advanced decomposition products of L-serine can be rationalized on the basis of these initial degradation products. Label incorporation studies have elucidated the origin of carbonyl precursors of methyl and 2,3-dimethylpyrazines formed in the thermal decomposition mixture of L serine. Three mechanistic pathways were identified for the formation of carbonyl precursors of methyl- and 2, 3-dimethylpyrazines. The major pathway (70%) for the formation of the precursor of methylpyrazine involved aldol addition of formaldehyde to glycolaldehyde to form glyceraldehyde. On the other hand, the major pathway (60%) for the formation of the precursor of 2,3-dimethylpyrazine involved an aldol condensation of acetaldehyde with glycolaldehyde to form 2,3 butanedione. PMID- 10725128 TI - Antimutagenic activity of flavonoids from Pogostemon cablin. AB - A methanol extract from Pogostemon cablin showed a suppressive effect on umu gene expression of SOS response in Salmonella typhimurium TA1535/pSK1002 against the mutagen 2-(2-furyl)-3-(5-nitro-2-furyl)acrylamide (furylfuramide). The methanol extract was re-extracted with hexane, dichloromethane, butanol, and water. A dichloromethane fraction showed a suppressive effect. Suppressive compounds against furylfuramide in the dichloromethane fraction were isolated by SiO(2) column chromatography and identified as 7,4'-di-O-methyleriodictyol (1), 7, 3',4' tri-O-methyleriodictyol (2), and 3,7,4'-tri-O-methylkaempferol (3). In addition, three flavonoids, ombuine (4), pachypodol (5), and kumatakenin (6), were isolated and identified from the dichrolomethane fraction. Compounds 1 and 3 suppressed >50% of the SOS-inducing activity at <0.6 micromol/mL, and the ID(50) values of both compounds were 0.25 micromol/mL. Compound 2 showed a weakly suppressive effect (17%) at a concentration of 0.6 micromol/mL, and compounds 4-6 did not. These compounds were also assayed with 3-amino-1,4-dimethyl-5H-pyrido[4,3 b]indole (Trp-P-1), which requires liver metabolizing enzymes. Compounds 3-6 suppressed >80% of the SOS-inducing activity of Trp-P-1 at <0.06 micromol/mL, and compounds 1 and 2 suppressed 87 and 63% at a concentration of 0.3 micromol/mL. In addition, these compounds were assayed with activated Trp-P-1, and the suppressed effects of these compounds were further decreased when compared to Trp-P-1. The antimutagenic activities of these compounds against furylfuramide, Trp-P-1, and activated Trp-P-1 were assayed by the Ames test using S. typhimurium TA100. PMID- 10725129 TI - Characterization of the total free radical scavenger capacity of vegetable oils and oil fractions using 2,2-diphenyl-1-picrylhydrazyl radical. AB - The total free radical scavenger capacity (RSC) of 57 edible oils from different sources was studied: olive (24 brands of oils), sunflower (6), safflower (2), rapeseed (3), soybean (3), linseed (2), corn (3), hazelnut (2), walnut (2), sesame (2), almond (2), mixture of oils for salad (2), "dietetic" oil (2), and peanut (2). Olive oils were also studied according to their geographical origins (France, Greece, Italy, Morocco, Spain, and Turkey). RSC was determined spectrophotometrically by measuring the disappearance of the radical 2,2-diphenyl 1-picrylhydrazyl radical (DPPH(*)) at 515 nm. The disappearance of the radical followed a double-exponential equation in the presence of oils and oil fractions, which suggested the presence of two (fast and slow) groups of antioxidants. RSC was studied for the methanol-soluble phase ("methanolic fraction", MF) of the oil, the fraction nonsoluble in methanol ("lipidic fraction", LF), and the nonfractionated oil ("total oil"; TF = MF + LF). Only olive, linseed, rapeseed, safflower, sesame, and walnut oils showed significant RSC in the MF due to the presence of phenolic compounds. No significant differences were found in the RSC of olive oils from different geographical origins. Upon heating at 180 degrees C the apparent constant for the disappearance of RSC (k(T)) and the half-life (t1/2) of RSC for MF, LF, and TF were calculated. The second-order rate constants (k2) for the antiradical activity of some phenolic compounds present in oils are also reported. PMID- 10725130 TI - Biochemical and functional properties of Atlantic salmon (Salmo salar) muscle proteins hydrolyzed with various alkaline proteases. AB - Protein hydrolysates (5, 10, and 15% degrees of hydrolysis) were made from minced salmon muscle treated with one of four alkaline proteases (Alcalase 2.4L, Flavourzyme 1000L, Corolase PN-L, and Corolase 7089) or endogenous digestive proteases. Reaction conditions were controlled at pH 7.5, 40 degrees C, and 7.5% protein content, and enzymes were added on the basis of standardized activity units (Azocoll units). Proteases were heat inactivated, insoluble and unhydrolyzed material was centrifuged out, and soluble protein fractions were recovered and lyophilized. Substrate specificities for the proteases was clearly different. Protein content for the hydrolysates ranged from 71.7 to 88.4%, and lipid content was very low. Nitrogen recovery ranged from 40.6 to 79.9%. The nitrogen solubility index was comparable to that of egg albumin and ranged from 92.4 to 99.7%. Solubility was high over a wide range of pH. The water-holding capacity of fish protein hydrolysates added at 1.5% in a model food system of frozen minced salmon patties was tested. Drip loss was on average lower for the fish protein hydrolysates than for egg albumin and soy protein concentrate, especially for Alcalase hydrolysates. Emulsification capacity for fish protein hydrolysates ranged quite a bit (75-299 mL of oil emulsified per 200 mg of protein), and some were better than soy protein concentrate (180 mL of oil emulsified per 200 mg of protein), but egg albumin had the highest emulsifying capacity (417 mL of oil emulsified per 200 mg of protein). Emulsification stability for fish protein hydrolysates (50-70%) was similar to or lower than those of egg albumin (73%) or soy protein concentrate (68%). Fat absorption was greater for 5 and 10% degrees of hydrolysis fish protein hydrolysates (3.22-5.90 mL of oil/g of protein) than for 15% hydrolysates, and all had greater fat absorption than egg albumin (2. 36 mL of oil/g of protein) or soy protein concentrate (2.90 mL of oil/g of protein). PMID- 10725131 TI - Impact of UV-C irradiation on the cell wall-degrading enzymes during ripening of tomato (Lycopersicon esculentum L.) fruit. AB - The effect of a hormic dose of UV-C (254 nm) on changes in fruit firmness and cell wall-degrading enzyme (CWDE) activity was determined using tomato fruit. Throughout the storage period, a decrease in firmness was jointly observed with an increase of the CWDE (polygalacturonase, pectin methyl esterase, cellulase, xylanase, beta-D-galactosidase, and protease) activity for all treatments, suggesting the involvement of these enzymes in the ripening process. However, the enhancement in the activity of the CWDE was significantly less in fruit subjected to the hormic dose of UV-C. This reduction may explain why irradiated fruit were firmer than control and consequently may explain how UV-C could delay the ripening and senescence process. We suggest that the CWDE are one of the targets of the UV-C, and by this action, irradiation contributed to a delay of the cell wall degradation and consequently retarded softening of the tomato fruit tissues. PMID- 10725132 TI - Effect of pH on the thermal denaturation of whey proteins in milk. AB - The effect of pH on thermal denaturation of four main whey protein fractions in skim milk was examined by gel permeation FPLC. On heating skim milk at 80 degrees C for 0.5-20.0 min over the pH range 5.2-8.8, the extent of denaturation, based on loss of solubility at pH 4.6, increased with heating time and was usually in the order immunoglobulins > serum albumin/lactoferrin > beta-lactoglobulin > alpha-lactalbumin. Rates of denaturation of the immunoglobulins and the serum albumin/lactoferrin fraction were highest at the lower end of this pH range, whereas those of beta-lactoglobulin and alpha-lactalbumin increased over most of the pH range. The effects of pH, addition of Ca, and reduction of disulfide bonds on the rates of the unfolding and aggregation stages of denaturation are discussed. PMID- 10725133 TI - Effect of malondialdehyde on the determination of furosine in milk-based products. AB - For nutritional purposes the recombination of milk or milk proteins with polyunsaturated fatty acids had long been considered in newly designed products, such as functional foods. Four milk-resembling model systems were prepared having malondialdehyde content ups to 1 mM. Systems were heated between 110 and 150 degrees C for up to 30 min. The effect of the presence of bifunctional aldehydes on the determination of furosine as a heat-induced marker was investigated. Levels of 0.01 mM MAD in the reaction mixture could affect significantly the determination of furosine in a milk-based system. Impairment of lysyl residues through the analysis of furosine could be underestimated in the presence of malondialdehyde. PMID- 10725134 TI - Preparation and dynamic viscoelasticity characterization of alkali-solubilized collagen from shark skin. AB - Alkali-solubilized collagens, prepared by alkali-acid extraction and alkali direct extraction (abbreviated AASC and ALSC, respectively), were characterized by dynamic viscoelastic measurement of collagen solution (10 mg/mL). The optimum preparative conditions in terms of yield and polypeptide size are as follows: for the alkali-acid extraction, a pretreatment with 0.5 or 1 M NaOH containing 15% Na(2)SO(4) within 5 days at 20 degrees C followed by the subsequent acid extraction, and for the alkaline direct extraction, a treatment with 0.5 M NaOH containing 10% NaCl at 4 degrees C for 20-30 days. A major portion of the polypeptide sizes of AASC and ALSC is composed of alpha chains (alpha1 and alpha2). Dynamic viscoelasticity of collagen solution was measured as a function of temperature. AASC showed a greater contribution of elastic behavior rather than viscous behavior. On the contrary, ALSC exhibits a stronger viscous behavior than elastic behavior. PMID- 10725135 TI - Lipid composition of wild ecuadorian Theobroma subincanum Mart. seeds and comparison with two varieties of Theobroma cacao L. AB - The present work analyzes the lipid fraction from seeds of wild Ecuadorian Theobroma subincanum and selected commercial varieties of Theobroma cacao from Mexico (var. Criollo) and Ecuador (var. Arriba). The lipid fraction was obtained from the seeds through supercritical fluid extraction and analysis performed by preparatory thin-layer chromatography followed by gas chromatography. The results revealed that in T. subincanum the triglycerides contain fatty acids with longer chains. The melting point and peroxide and saponifiable numbers were determined for each Theobroma sample. The results lead to the conclusion that T. subincanum would produce a poorer quality butter than T. cacao. Nevertheless, the results do point toward a significant commercial use of T. subincanum for low-profile products. PMID- 10725136 TI - Characterization of catecholase and cresolase activities of eggplant polyphenol oxidase. AB - In the present paper the catecholase and cresolase activities of eggplant polyphenol oxidase (PPO) are described. To preserve the latter activity, a partially purified enzyme was used. Peroxidase was removed from the preparation to avoid its interference with PPO during phenol oxidation. The partially purified eggplant PPO was fully active. The catecholase/cresolase ratio of 41.1 indicated that, in a pH close to the physiological, diphenol oxidation predominates over monophenol oxidation. The characteristic lag phase of the cresolase activity is modulated by the pH, the monophenol and diphenol concentrations, and the enzyme's concentration. The effect of several inhibitors was also tested, and the K(i) values of the two most effective (tropolone and 4 hexylresorcinol) were determined. PMID- 10725137 TI - Isolation and partial characterization of thermostable isoperoxidases from potato (Solanum tuberosum L.) tuber sprouts. AB - Peroxidases (POD; EC 1.11.1.7) can cross-link cell wall polymers and may have an impact on the final textural quality of potato tubers. Because heat treatments are important during processing, the thermal properties of isoPODs from soluble and ionically and covalently bound fractions were studied from both potato tubers and sprouts. For both tissues, the ionically bound fraction was the most thermostable; approximately 20% of POD activity remained after a heat treatment of 10 min at 90 degrees C (for sprouts). The temperature profile of the ionically bound sprout fraction appeared to be nonlinear and suggested the presence of a very thermostable POD, which still showed activity after a heat treatment at 100 degrees C. Visualization by using isoelectric focusing confirmed the occurrence of a thermostable isoPOD with an IEP of 9.5, which displayed regeneration of activity after heat inactivation. This cationic POD was further purified by chromatography techniques, and by SDS-PAGE its molecular mass was estimated at 38 kDa. PMID- 10725138 TI - Phosphorylation of the 12S globulin from rapeseed (Brassica napus L.) by phosphorous oxychloride: chemical and conformational aspects. AB - The effect of progressive phosphorylation by phosphorous oxychloride upon the conformation of the 300 kDa storage protein (cruciferin) from rapeseed has been studied using chemical analysis, SDS-PAGE, HPLC, analytical ultracentrifugation, viscometry, fluorescence spectroscopy, and hydrophobicity measurement. The amount of phosphorous in the protein increased with the excess of phosphorous oxychloride and the pH of reaction. The bulk of phosphorus was only loosely bound to the protein and was removed by washing with cold perchloric acid. The more stably bound phosphorus groups after reaction at pH 8 were found to be nearly equally attached to amino and hydroxyl groups, whereas phosphorylation at pH 10 11 led to predominant O-phosphorylation as detected by studying the acid- and alkali-lability of the protein-phosphorous bonds. A 50 kDa component appeared as a product of covalent cross-linking of the constituent alpha- and beta polypeptide chains. A 2.5S fraction appeared as the main product of dissociation, which takes place after a critical step of modification. The higher the extent of phosphorylation, the larger was the percentage of higher molecular weight products, the percentage of which was most significant after modification under strongly alkaline conditions. They may be attributed both to products of chemical cross-linking and to noncovalently linked aggregates formed by interactions of partially unfolded derivatives exhibiting an increased surface hydrophobicity. PMID- 10725139 TI - Free amino acids present in commercially available seedlings sold for human consumption. A potential hazard for consumers. AB - The importance of fresh seedlings for human consumption on European markets continues to increase. Although the contents of free amino acids and potentially toxic free nonprotein amino acids in these fresh and supposedly healthy seedlings is very different from those of the seeds, the crude composition is never mentioned on commercial packages. A commercial product containing seven different kinds of fresh seedlings including kamut, adzuki bean, chickpea, mungbean, pinto bean, garden pea, and lentil has been analyzed by HPLC. Per 100 g of fresh product, 548.2 mg of total free amino acids was found, of which 56.7 mg is free nonprotein amino acids including beta-(isoxazolin-5-on-2-yl)alanine, homoserine, and isowillardiine and the plant hormone trigonelline (N-methylnicotinic acid). The highest amounts of free nonprotein amino acids and trigonelline are found in garden pea (28.3 mg/100 g), mungbean (9.59 mg/100 g), and lentil (7.50 mg/100 g) seedlings. Trigonelline is present in all legume seedlings examined. PMID- 10725140 TI - Vicinal diketone formation in yogurt: (13)C precursors and effect of branched chain amino acids. AB - Addition of branched-chain amino acids (BCAA) or an inhibitor of the BCAA biochemical pathways during fermentation of milk with a lac(-) mutant of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus strongly influenced the formation of two aroma-impact compounds, 2,3-butanedione and 2,3-pentanedione, as well as their direct precursors 2-acetolactate and 2 acetohydroxybutyrate. This suggests a connection between vicinal diketone formation and BCAA biosynthesis in yogurt bacteria. A recently developed static and-trapped headspace technique combined with gas chromatography-mass spectrometry demonstrated incorporation of (13)C from [U-(13)C(6)]-D-glucose and [U-(13)C(4)]-L-threonine into both vicinal diketones. For 2,3-butanedione, glucose is the major precursor via pyruvate and activated acetaldehyde. For 2, 3 pentanedione, L-threonine is a precursor via 2-ketobutyrate, but glucose is the major contributor via activated acetaldehyde and, possibly, also via 2 ketobutyrate, which is a degradation product of 3-methylaspartate, an intermediate in glutamate synthesis. PMID- 10725141 TI - Polyphenol content and total antioxidant activity of vini novelli (young red wines). AB - Eight commercial Italian vini novelli (red wines prepared by carbonic maceration and supposed to be consumed within three months from their wine-making) were evaluated for their total antioxidant activity. The wines had an average total phenol content (1605.4 +/- 337.4 mg/L gallic acid equivalents) lower than that of wines prepared by traditional maceration and consumable after aging (2057. 3 +/- 524.0 mg/L gallic acid equivalents). The average flavanol content (424.7 +/- 121.3 mg/L catechin equivalents) and the total antioxidant activity (16.8 +/- 3.8 mmol/L Trolox equivalents) of vini novelli were higher than the corresponding values (382.7 +/- 174.5 mg/L catechin equivalents and 12.3 +/- 3.3 mmol/L Trolox equivalents) found for aged wines. Three couples of experimental wines were prepared from the same grapes by traditional or carbonic maceration. These wines showed a different phenolic pattern, anthocyanins being more abundant in vini novelli. However, the average total antioxidant activities of the wines were similar, suggesting that aging (and not the wine-making technique) is the main factor influencing the antioxidant activity of red wines. PMID- 10725142 TI - Color and phenolic compounds of a young red wine. Influence of wine-making techniques, storage temperature, and length of storage time. AB - The purpose of this work was to determine the influence of vinification technique (maceration temperature and clarification method), storage temperature, and length of storage time on the phenolic compounds and color of young red wines. Multivariate analysis of variance and principal component analysis pointed to significant differences among all of the variables according to vinification technique and length of storage time. Storage temperature did not cause significant differences between some of the variables. The best color characteristics were obtained when low-temperature maceration wines were clarified with polyvinylpyrrolidone. Color quality also improved with lower storage temperature. PMID- 10725143 TI - Perspectives into factors limiting in vivo digestion of legume proteins: antinutritional compounds or storage proteins? AB - The in vivo protein digestibility of raw and cooked common bean (Phaseolus vulgaris L.) and faba bean (Vicia faba L.) and of protein fractions extracted from them was determined with growing rats. Overnight-fasted rats were intubated with a protein suspension or fed the same amount of protein added to a basal diet. The rats were killed 1 h later, the contents of stomach and small intestine were washed out, and their protein contents were measured. The in vivo digestibility of proteins of raw common bean flour was 72.4% and not significantly improved after cooking. In contrast, the digestibility of faba bean proteins was decreased from 86.5 to 60.6% by the thermal treatment. Globulins from either species had similar digestibilities (approximately 70%). Proteins in the soluble fraction of cooked beans were more digestible than those in the insoluble fraction, which contained the bulk of the proteins. Hemagglutination assay and trypsin inhibitor determination indicated that after the thermal treatment only very low, nonharmful, levels of both lectin and inhibitor remained. Faba bean contained more polyphenols than common bean samples, with most of the polyphenols being bound to globulins. However, protein-bound polyphenols were markedly decreased after cooking. SDS-PAGE characterization of the gastrointestinal digesta of globulins and amino acid analysis of undigested proteins of whole cooked common bean and faba bean suggested that it is mainly the structural properties of the storage proteins and not their binding of polyphenols, which determines the extent of protein aggregation on autoclaving and may therefore be responsible for their low digestibility. PMID- 10725144 TI - Investigations into genotypic variations of peanut carbohydrates. AB - Carbohydrates are known to be important precursors in the development of roasted peanut quality. However, little is known about their genotypic variation. A better understanding of the role of carbohydrates in roasted peanut quality requires first an understanding of the genotypic variation in the soluble carbohydrate components. Ion exchange chromatography was used to isolate 20 different carbohydrate components in 52 genotypes grown in replicated trials at two locations. Inositol, glucose, fructose, sucrose, raffinose, and stachyose were quantitated, and 12 unknown peaks were evaluated on the basis of the peak height of the unknown relative to the cellobiose internal standard peak height. Peaks tentatively identified as verbascose and ajugose could not be properly integrated because of tailing. Of the 18 carbohydrates that were estimated, 9 exhibited significant variation between test environments, 5 among market types, 14 among genotypes within market types, and 11 exhibited some significant form of genotype x environment interaction. Genotypes accounted for 38-78% of the total variation for the known components, suggesting that broad-sense heritability for these components is high. The observed high genotypic variation in carbohydrate components is similar to the high genotypic variation observed for the sweetness attribute in roasted peanuts, which raises the question regarding possible interrelationships. The establishment of such interrelationships could be most beneficial to peanut breeding programs to ensure the maintenance of flavor quality in future peanut varieties. PMID- 10725145 TI - Relationships of sweet, bitter, and roasted peanut sensory attributes with carbohydrate components in peanuts. AB - Certain roasted peanut quality sensory attributes have been shown to be heritable. Currently the only means of measuring these traits is the use of a trained sensory panel. This is a costly and time-consuming process. It is desirable, from a cost, time, and sample size perspective, to find other methodologies for estimating these traits. Because sweetness is the most heritable trait and it has a significant positive relationship to the roasted peanut trait, the possible relationships between heritable sensory traits and 18 carbohydrate components (inositol, glucose, fructose, sucrose, raffinose, stachyose, and 12 unknown peaks) in raw peanuts from 52 genotypes have been investigated. Previously reported correlations among sweet, bitter, and roasted peanut attributes were evident in this study as well. Where there was positive correlation of total sugars with sweetness, there also was positive correlation of total sugars with roasted peanut attribute and negative correlation of total sugars with bitterness and astringency. The expected generalized relationship of total sugars or sucrose to sweetness could not be established because the relationship was not the same across all market-types. Further work is needed to determine the nature of the chemical components related to the bitter principle, which appear to modify the sweet response and interfere with the sensory perception of sweetness, particularly in the Virginia market-type. Also, certain carbohydrate components showed significant relationships with sensory attributes in one market-type and not another. These differential associations demonstrate the complexity of the interrelationships among sweet, bitter, and roasted peanut sensory attributes. Within two market-types it is possible to improve the efficiency of selection for sweetness and roasted peanut quality by assaying for total carbohydrates. On the basis of the regression values the greatest efficiency would occur in the fastigiate market-type and then the runner. PMID- 10725146 TI - Identification of triacylglycerol species from high-saturated sunflower (Helianthus annuus) mutants. AB - The triacylglycerol (TAG) composition of oils from new high-saturated sunflower lines has been studied by means of GLC. The TAG profiles have been compared with the TAG reconstruction made after lipase hydrolysis (according to the 2-random 1,3-random theory). New TAG species with asclepic (cis,Delta11-octadecenoic acid, isomer of oleic acid), araquidic, or behenic acids have been synthesized and identified in oils from mutant lines. The TAG molecular species that contain asclepic acid instead of oleic acid have a longer retention time. Because each mutant oil has a specific TAG GLC pattern, this method could be used for a more precise validation of oil type than current fatty acid methyl ester analysis. The comparison of the results obtained by GLC with the reconstruction after pancreatic lipase hydrolysis shows, in general, a good agreement between both methods. However, results shown in this paper show that this is not always the case. TAG species containing two molecules of linoleic acid show a higher presence of palmitic or stearic acid than could be expected from a random distribution. The abundance of SLL increased in proportion to the stearic acid content of the oil, and the amount of TAG species with three unsaturated fatty acids (LLL or OLL) was therefore reduced. PMID- 10725147 TI - Metabolism of triacylglycerol species during seed germination in fatty acid sunflower (Helianthus annuus) mutants. AB - Sunflower mutant lines with high saturated fatty acid content (palmitic or stearic) in the oil have a completely different set of triacylglycerols (TAG), some of which were not found in standard sunflowers. For optimum seed germination, all of these new TAG species must be effectively catabolized. The behavior of the TAG composition during germination in cotyledons of all these mutant lines showed two different phases: an initial phase (between 0 and 2 days after sowing) with a higher catalytic activity and a preference for TAG containing at least two oleic acid molecules and a second phase with lower TAG degradation rate and a low preference for TAG containing two saturated fatty acids usually accompanied by linoleic acid. Despite the elevated content of saturated fatty acids in some TAG species, the total TAG degradation rate and germination process were similar in these lines, suggesting that sunflower seed lipases do not show a marked preference for any TAG species. PMID- 10725148 TI - Supercritical fluid extraction of grape glycosides. AB - Supercritical fluid extraction with methanol-modified CO(2) was used to extract glycosides from grapes. An optimization design involving 12 extraction variables was applied to achieve quantitative recoveries. The most important factor was the amount of organic modifier, a consequence of the high degree of glycoside polarity. By the proposed method, the total time of analysis can be decreased relative to that required for more conventional extractions. The full method can also be automated. PMID- 10725149 TI - Analysis of free and esterified ergosterol in tomato products. AB - A new extraction and chromatographic procedure to quantify free and esterified ergosterol in tomato products was devised. The extraction solution was composed of a dichloromethane/methanol mixture in a 2:1 (v/v) ratio. This extraction solvent allowed for higher ergosterol recovery from tomato products (an average of 25% more) compared to hexane, which is frequently employed for ergosterol extraction. Both free and esterified ergosterol were determined by HPLC reverse phase chromatography employing a Nova-Pak C-18 column (300 x 3.9 mm), filled with 4 mm average particle size and a guard column of the same material. The elution was performed at a flow rate of 1 mL. min(-1) with a linear gradient of solvent A (methanol/water, 80:20, v/v) and solvent B (dichloromethane). The gradient, starting at sample injection, was from 0 to 50% B for 20 min for the free ergosterol analysis and additional 15 min at 50% B to analyze the ergosterol esters. This technique has proven to be more sensitive for ergosterol determination than other reported chromatographic procedures. Moreover, ergosterol esters, extracted from various fungal sources, separated well and were easily quantified. PMID- 10725150 TI - GC-FID- and acyl carbon number-based determination of characteristic groupings of complex triglyceride (Benefat S and other) mixtures. AB - Techniques for the gas and liquid chromatographic separation of complex mixtures of triglycerides have evolved over the past two decades, as reviewed in detail by Huang et al. (J. Agric. Food Chem. 1995, 43, 1834-1844; J. Agric. Food Chem. 1997, 45, 1770-1778). A novel method for the quantitative partitioning of complex mixtures of triglycerides into functionally related groups is developed and applied to a low-calorie triglyceride mixture [namely, Benefat S or Salatrim plus mid-chain (C(6,8,10,12)) fatty acids]. The method is based on a nonlinear calibration of retention times (RTs) of a suite of standard triglycerides on their acyl carbon numbers [(ACNs), the sum of all the acyl carbon atoms in a given triglyceride] to estimate all of the intermediate ACNs (from 6 to 66). With the calibrated ACN scale and identifications of some components of a complex mixture's composition, ACN-based partitions were established and a Benefat S triglyceride chromatogram was partitioned into seven functionally related groups. This method is provisional in the sense that it would typically be employed when the identifications of many components of a complex, homologous series were unknown, yet functionally related groups needed to be quantified. This method has proven to be particularly useful in the intercalibration of research laboratories with production facility laboratories during complex ( approximately 50-90 compounds) and large-scale ( approximately 20 ton) syntheses because of the high reproducibility of the ACN-based partitioning of complex chromatograms. This carbon number and statistically based method can be generally applicable to other complex mixtures of organic compounds and is readily adaptable to laboratory intercalibration efforts. PMID- 10725151 TI - CS(2) blinds in Brassica crops: false positive results in the dithiocarbamate residue analysis by the acid digestion method. AB - Various members of the Brassicaceae family (cauliflower, savoy cabbage, red cabbage, turnip-rooted cabbage) grown without any application of pesticides were analyzed according to the acid digestion method commonly used for the determination of dithiocarbamate fungicide residues. Depending on postharvest treatments, high non-anthropogenic CS(2) values up to 4 mg/kg were found in some cases, especially in frozen raw cabbage samples, exceeding maximum residue limits. To explore phytogenic CS(2) occurrences, two model substances (phenylisothiocyanate and methyl tryptaminedithiocarbamate) representing natural mustard oils and brassinines, respectively, were analyzed for their acid hydrolysis decomposition products. In both cases, COS was found generally, but CS(2) was readily formed during acid digestion, too, when sulfides were present. The results obtained clearly demonstrate that CS(2) values determined by using the acid digestion method of crops rich in secondary metabolism sulfur compounds have to be interpreted carefully. PMID- 10725152 TI - Organochlorine pesticide residues in Italian citrus essential oils, 1991-1996. AB - Organochlorine pesticide contamination in 148 lemon essential oils, 123 sweet orange oils, 121 mandarin oils, and 147 bergamot oils produced in Italy in the years 1991-1996 was studied by HRGC-ECD. Confirmation analyses were carried out by GC-MS. Tetradifon, dicofol and its decomposition product 4,4' dichlorobenzophenone were found. Over the course of the study dicofol and tetradifon residues steadily decreased; the percentage of contaminated samples reflects this course and decreases considerably from 1991 to 1996. PMID- 10725153 TI - Application of (31)P NMR spectroscopy in food analysis. 1. Quantitative determination of the mono- and diglyceride composition of olive oils. AB - This paper introduces a facile method to determine the amount of mono- and diglycerides in virgin olive oils. This method is based on the phosphorylation of the free hydroxyls of the mono- and diglycerides with 2-chloro-4,4,5,5 tetramethyldioxaphospholane and the integration of the appropriate peaks in the (31)P NMR spectrum. Quantitative (31)P NMR spectroscopy can be extended to the quantification of other constituents of olive oils bearing functional groups with labile protons. PMID- 10725154 TI - Chemical composition of some wild peanut species (Arachis L.) seeds. AB - Oil, protein, ash, and carbohydrate contents, iodine value, and fatty acid and sterol compositions were studied in seeds of Arachis trinitensis, A. chiquitana, A. kempff-mercadoi, A. diogoi, A. benensis, A. appressipila, A. valida, A. kretschmeri, A. helodes, A. kuhlmannii, A. williamsii, A. sylvestris, A. matiensis, A. pintoi, A. hoehnei, A. villosa, and A. stenosperma. Oil content was greatest in A.stenosperma (mean value = 51.8%). The protein level was higher in A. sylvestris (30.1%) and A. villosa (29.5%). Mean value of oleic acid varied between 30.6% (A. matiensis) and 46.8% (Arachis villosa), and linoleic acid oscillated between 34.1% (A. villosa) and 47.4% (A. appressipila). The better oleic-to-linoleic (O/L) ratio was exhibited by A. villosa (1.38). Some species showed higher concentration of behenic acid. The greatest level of this fatty acid was found in A. matiensis (6.2%). Iodine value was lower in A. valida (99.2). The sterol composition in the different peanut species showed higher concentration of beta-sitosterol (mean values oscillated between 55.7 and 60.2%) followed by campesterol (12.4-16. 5%), stigmasterol (9.7-13.3%), and Delta(5) avenasterol (9.7-13.4%). The chemical quality and stability of oils (iodine value and O/L ratio) from wild peanut studied in this work are not better than those of cultivated peanut. PMID- 10725155 TI - Activity of antifungal proteins against mold in sorghum caryopses in the field. AB - Sorghums were stressed with pathogenic fungi and sprinkling to determine relationships between changes in chitinase and sormatin in caryopses and grain mold resistance. Panicles of 10 cultivars differing in mold resistance and accumulation of antifungal proteins (AFPs) were inoculated at anthesis with Fusarium moniliforme and Curvularia lunata spores. Panicles were sampled at 30 and 50 days after anthesis, and caryopses were evaluated for chitinase and sormatin using western blots. Sprinkling panicles (to mimic rainfall) decreased sormatin and chitinase in most cultivars. Inoculation decreased AFPs in susceptible cultivars, but resistant cultivars maintained or increased AFPs in caryopses. Grain mold resistance corresponded to induction of AFP synthesis in response to sprinkling, fungal stress, and/or adverse field conditions. Sormatin and chitinase appear to be an active part of the defense mechanism of the caryopsis against grain mold. PMID- 10725156 TI - Structural characterization of the lignin from the nodes and internodes of Arundo donax reed. AB - Milled wood lignin (MWL) and dioxane lignin (DL) from different morphological regions (nodes and internodes) of Arundo donax reed were subjected to a comprehensive structural characterization by (13)C, (1)H NMR, FTIR, and UV spectroscopies and functional analysis. The permanganate and nitrobenzene oxidation methods were also applied to the in situ lignins. Both node and internode lignins are HGS-type lignins, with a significant amount of H units (including p-coumaric acid type structures). The S/G ratio (1.13-1.32), the weight-average molecular weight (20,400-24,500), the methoxyl group content (0.90 0.98), the phenolic hydroxyl group content (0.23-0.27), and the aliphatic hydroxyl group content (1.00-1.09) are not very different in the lignins from nodes and internodes. However, some structural differences between node and internode lignins were observed. The former has much more phenolic acids (p coumaric and ferulic), 8.8% in node versus 1.2% in internode and less beta-O-4 (0. 32 and 0.49 per aromatic unit in node and internode, respectively). In situ node lignin is more condensed than internode lignin. PMID- 10725157 TI - Carotenoid pigments in Rosa mosqueta hips, an alternative carotenoid source for foods. AB - Carotenoid composition has been investigated in Rosa mosqueta hips (Rosa rubiginosa, Rosa eglanteria). Six major carotenoids were identified (beta carotene, lycopene, rubixanthin, gazaniaxanthin, beta-cryptoxanthin, and zeaxanthin) together with other minor carotenoids (violaxanthin, antheraxanthin, and gamma-carotene). An average composition has been estimated as follows: beta carotene (497.6 mg/kg of dry wt), lycopene (391.9 mg/kg of dry wt), rubixanthin (703.7 mg/kg of dry wt), gazaniaxanthin (289.2 mg/kg of dry wt), beta cryptoxanthin (183.5 mg/kg of dry wt), zeaxanthin (266. 6 mg/kg of dry wt), and minor carotenoids (67.1 mg/kg of dry wt). Possible uses in food technology are outlined and discussed including the preparation of highly colored oleoresins as natural colorants of food and beverages and as provitamin A sources. PMID- 10725158 TI - Effects of dehulled adlay on the culture count of some microbiota and their metabolism in the gastrointestinal tract of rats. AB - Experiments were conducted to study the effect of a dietary supplement of dehulled adlay (Coix lachryma-jobi L. var. ma-yuen Stapf) on the culture counts of some important groups of intestinal bacteria and their metabolism in the gastrointestinal (GI) tract of Sprague-Dawley rats. Rats were divided into four groups, and each group was fed a diet containing different levels of dehulled adlay for 30 days as follows: 0% (control), 5%, 20%, and 40%. All animals fed with adlay had normal healthy intestinal walls and no pathogenic signs whatsoever. There were no significant differences in body weight gain or the cecal pH between different groups of rats. Both the 20% and 40% groups had lower culture counts of enterics in their feces than the 5% and control groups, whereas the culture counts of fecal lactic acid bacteria were higher in feces of rats fed with adlay than in the control group. Cecal total short-chain fatty acid (SCFA) content and fecal SCFA were significantly higher in the 20% and 40% groups than in the control and 5% groups. All the adlay-fed rats had a higher fecal butyric acid concentration than the control rats. It is concluded that adlay has a significant influence on the growth of intestinal bacteria, which may ultimately affect the physiology and other functions of GI tracts of rats. PMID- 10725159 TI - Role of aldehyde oxidase in the hepatic in vitro metabolism of 3-methylindole in pigs. AB - The metabolism of 3-hydroxy-3-methylindolenine (HMI), a recently discovered metabolite of 3-methylindole (3MI, skatole) produced by porcine liver microsomes, was investigated in vitro using porcine liver cytosol. HMI was rapidly metabolized to a single product, 3-hydroxy-3-methyloxindole (HMOI), by porcine cytosol. By the use of the selective inhibitors menadione and quinacrine, it was shown that the enzyme responsible for the oxidation of HMI into HMOI was aldehyde oxidase (AO; aldehyde:oxygen oxidoreductase, EC 1.2.3.1). The activity of AO in the conversion of HMI to HMOI was measured in a population of pigs (n = 30) with a wide range of 3MI levels in back fat (0.07-0.30 mg/kg). AO activity was found to be negatively correlated (r = -0.70; P < 0.001) with the level of 3MI in fat. The results of the present study suggest that AO plays an important role in the metabolism of 3MI in the pig and that its catalytic activity is related to an adequate 3MI clearance. PMID- 10725160 TI - Binding affinity of hydrolyzable tannins to parotid saliva and to proline-rich proteins derived from it. AB - Proline-rich proteins (PRP) in human parotid saliva have a high affinity for dietary polyphenolic compounds (tannins), forming stable complexes that may modulate the biological and nutritional properties of the tannin. The formation of such complexes may also have an important role in the modulation or promotion of the sensation of oral astringency perceived when tannin-rich foods and beverages are consumed. The major classes of PRP (acidic, basic, and glycosylated) have been isolated from human saliva, and the relative binding affinities of a series of hydrolyzable tannins, which are found in a number of plant-derived foods and beverages, to these PRP classes have been determined using a competition assay. All of the classes of PRP have a high capacity for hydrolyzable tannins. Within the narrow range of binding affinities exhibited, structure/binding relationships with the levels of tannin galloylation, hexahydroxydiphenoyl esterification, and degree of polymerization were identified. No individual class of human salivary PRP appears to have an exclusive affinity for a particular type of hydrolyzable tannin. PMID- 10725161 TI - High content of dopamine, a strong antioxidant, in Cavendish banana. AB - A strong water-soluble antioxidant was identified in the popular commercial banana Musa cavendishii. It is dopamine, one of the catecholamines. For suppressing the oxygen uptake of linoleic acid in an emulsion and scavenging a diphenylpicrylhydrazyl radical, dopamine had greater antioxidative potency than glutathione, food additives such as butylated hydroxyanisole and hydroxytoluene, flavone luteolin, flavonol quercetin, and catechin, and similar potency to the strongest antioxidants gallocatechin gallate and ascorbic acid. Banana contained dopamine at high levels in both the peel and pulp. Dopamine levels ranged from 80 560 mg per 100 g in peel and 2.5-10 mg in pulp, even in ripened bananas ready to eat. Banana is thus one of the antioxidative foods. PMID- 10725162 TI - Insulin-like biological activity of culinary and medicinal plant aqueous extracts in vitro. AB - To evaluate the possible effects on insulin function, 49 herb, spice, and medicinal plant extracts were tested in the insulin-dependent utilization of glucose using a rat epididymal adipocyte assay. Cinnamon was the most bioactive product followed by witch hazel, green and black teas, allspice, bay leaves, nutmeg, cloves, mushrooms, and brewer's yeast. The glucose oxidation enhancing bioactivity was lost from cinnamon, tea, witch hazel, cloves, bay leaf and allspice by poly(vinylpyrrolidone) (PVP) treatment, indicating that the active phytochemicals are likely to be phenolic in nature. The activity of sage, mushrooms, and brewers's yeast was not removed by PVP. Some products such as Korean ginseng, flaxseed meal, and basil have been reported to be effective antidiabetic agents; however, they were only marginally active in our assay. Our technique measures direct stimulation of cellular glucose metabolism, so it may be that the active phytochemicals in these plants improve glucose metabolism via other mechanisms or that this in vitro screening is not a reliable predictor of hypoglycemic effects in vivo for some products. In summary, the positive effects of specific plant extracts on insulin activity suggest a possible role of these plants in improving glucose and insulin metabolism. PMID- 10725163 TI - Detection of rancid defect in virgin olive oil by the electronic nose. AB - A sensor array of 32 conducting polymer sensors has been used to detect the rancid defect in virgin olive oils. A training set, composed of admixtures of a Portuguese virgin olive oil with different percentages (0-100%) of a rancid standard oil, was used for the selection of the best sensors classifying correctly the samples. Information on volatile compounds responsible for rancidity and the sensory evaluation of samples by assessors were used for explaining the mathematical selection of sensors. A tentative calibration, using unsupervised procedures (PCA and MDS) and a nonlinear regression, was carried out, with the training set, and later confirmed with a test set with which rancid commercial samples of different varieties were used to spike a Greek extra virgin olive oil at low levels of rancidity (0.5-6%). PMID- 10725164 TI - Effect of ultrasound emulsification on cheese aroma encapsulation by carbohydrates. AB - The encapsulation of liquid cheese aroma (20%) in different carbohydrate matrices by spray-drying was investigated. Carbohydrate stabilized emulsions have been prepared by two emulsifying methods, ultrasonic or Ultra-Turrax treatments, and have been compared in terms of emulsion stability and encapsulation efficiency. The use of ultrasound is particularly effective to obtain a stable emulsion with maltodextrin as support, which is known for poor emulsification properties. In the same way, the spray-dried maltodextrin microcapsules were more effective for retaining cheese aroma when ultrasound (12.7 g/100 g of dry powder) was used for the emulsification step rather than Ultra-Turrax (10.7 g/100 g of dry powder). In terms of encapsulation efficiency, the best system of cheese aroma encapsulation is obtained using ultrasound for the emulsification step and modified starch as support (94.3%). With this support, the positive effect of ultrasound resulted in a lower microcaspsule size and in a higher aroma retention than when Ultra-Turrax was used (83.3%). Studies on the aroma profiles showed changes after encapsulation that depend not only on the nature of the support and the emulsification method but also on the interactions between the aroma compound and the matrix. In terms of flavor quality, the best system of cheese aroma encapsulation is obtained using ultrasound and maltodextrin as support. PMID- 10725165 TI - Potent odorants of raw Arabica coffee. Their changes during roasting. AB - Aroma extract dilution analysis of raw Arabica coffee revealed 3-isobutyl-2 methoxypyrazine (I), 2-methoxy-3,5-dimethylpyrazine (II), ethyl 2-methylbutyrate (III), ethyl 3-methylbutyrate (IV), and 3-isopropyl-2-methoxypyrazine (V) as potent odorants. The highest odor activity value was found for I followed by II, IV, and V. It was concluded that I was responsible for the characteristic, peasy odor note of raw coffee. Twelve odorants occurring in raw coffee and (E)-beta damascenone were also quantified after roasting. The concentration of I did not change, whereas methional, 3-hydroxy-4, 5-dimethyl-2(5H)-furanone, vanillin, (E) beta-damascenone, and 4-vinyl- and 4-ethylguaiacol increased strongly during the roasting process. PMID- 10725166 TI - Frozen storage effects on anthocyanins and volatile compounds of raspberry fruit. AB - The quantitative and qualitative evolution of the anthocyanins and volatile compounds of four raspberry cultivars (cvs. Heritage, Autumn Bliss, Zeva, and Rubi) growing in Spain were analyzed raw, just frozen, and during long-term frozen storage at -20 degrees C for a 1 year period. HS-SPME coupled with GC-MS and HPLC techniques were employed to study the evolution of the volatile compounds and the individual anthocyanins, respectively. The volatile aroma composition changes produced by the freezing process and long-term frozen storage were minimal. Only a significant increase in extraction capacity was obtained for alpha-ionone (27%) and for caryophyllene (67%) in Heritage at 12 months of storage. The stability of anthocyanins to freezing and frozen storage depends on the seasonal period of harvest. Heritage and Autumn Bliss (early cultivars) were less affected by processing and long-term frozen storage (1 year), and the total pigment extracted showed the tendency to increase 17 and 5%, respectively. Rubi and Zeva (late cultivars) suffered a decreased trend on the total anthocyanin content of 4% for Rubi and 17.5% for Zeva. Cyanidin 3-glucoside most easily suffered the degradative reactions that take place during processing and the storage period. PMID- 10725167 TI - Effect of extrusion temperature on the solubility and molecular weight of lentil bean flour proteins containing low cysteine residues. AB - Lentil flour was extruded at die temperatures of 135, 160, and 175 degrees C. The soluble protein content in the extrudates decreased by 40.1% in the extracting buffer (1% sodium dodecyl sulfate in 50 mM sodium phosphate buffer, pH 6.9) as the extrusion die temperature was increased to 175 degrees C. The most insoluble proteins in the extrudates extruded at die temperatures of up to 175 degrees C could be resolubilized by using sonication. The total disulfide content and sulfhydryl content in the extrudates decreased. The SDS-PAGEs showed that the molecular weight distribution of proteins in the lentil flour changed little before and after extrusion as well as during reduction. The results from this study show that the extrusion temperature had less effect on the solubility and molecular weight of the lentil proteins, which contain a lower level of cysteine residues than wheat proteins. PMID- 10725168 TI - Fermentation of white wines in the presence of wood chips of American and French oak. AB - Must obtained from Airen grapes was fermented in the presence of wood chips (4 and 7 g/L) of either French oak (from Vosges, central France, and Allier zones) or American oak. Fermentation yields were higher than in the control fermentations carried out in the absence of wood, and production of volatile substances during fermentation (alcohols, esters, and acetates) was also higher. The volatile substances that leached out of the wood were analyzed by GC-MS-SIR. The results showed that their concentrations depended on the type and amount of the oak; some of these substances were consumed in part by the yeasts during fermentation. A taste panel favorably assessed the wines produced by fermentation in the presence of oak chips, which retained part of the must original fruity aroma. PMID- 10725169 TI - Enzymatic hydrolysis, grease permeation, and water barrier properties of zein isolate coated paper. AB - An inexpensive zein-lipid mixture was isolated from yellow dent, dry-milled corn. Grease permeation through zein isolate applied to brown Kraft paper was found to be independent of loading levels at zein isolate levels above 30 mg/16 in.(2). The data shows that water vapor transmission rates depended on the amount of coating applied. Triacylglycerols were the most abundant lipid in milled corn but were absent in the zein isolate (perhaps due to hydrolysis by lipases). Zein from the paper was hydrolyzed enzymatically and the hydrolysis monitored by SDS capillary electrophoresis. At an E:S ratio of 1:100 no further increase in the hydrolysate peak occurred after 10 and 30 min for alpha-chymotrypsin and pancreatin 8 x; however, zein and lipid were still present 1 h after hydrolysis by pancreatin 1 x. PMID- 10725170 TI - Solid-state bioconversion of phenolics from cranberry pomace and role of Lentinus edodes beta-glucosidase. AB - Cranberry pomace contains large amounts of phenolic glycosides, which are important sources of free phenolics that have many food uses such as antioxidants, flavorings, and nutraceuticals. Our hypothesis was that these glycosides in cranberry pomace could be hydrolyzed by beta-glucosidase produced by Lentinus edodes during solid-state fermentation. On the basis of this hypothesis, our objective was to investigate the potential of using cranberry pomace as a substrate for the production of free phenolics and beta-glucosidase through solid-state fermentation by a food-grade fungus L. edodes. Our results suggested that L. edodes beta-glucosidase played a major role in release of phenolic aglycons from cranberry pomace during solid-state fermentation. After 50 days of cultivation, the yield of total free phenolics reached the maximum of 0.5 mg per g of pomace, while the beta-glucosidase activity was about 9 units per g of pomace. The enzyme exhibited optimal activity at 60 degrees C and at pH 3.5 and was stable at temperatures up to 50 degrees C and between pH 3 and 6.5. The major free phenolics produced from cranberry pomace were identified by HPLC as gallic acid, chlorogenic acid, p-hydroxybenzoic acid, and p-coumaric acid. These results suggest that cranberry pomace is a potential substrate for producing food grade phenolics and fungal beta-glucosidase. The L. edodes beta-glucosidase showed good stability and tolerance to low pH and, therefore has potential applications in wine and juice processing for aroma and flavor enrichment through enzymatic hydrolysis of glucoside precursors. PMID- 10725171 TI - Two-dimensional visible/near-infrared correlation spectroscopy study of thermal treatment of chicken meats. AB - The thermal treatment of chicken meats was investigated by generalized 2D visible/NIR correlation spectroscopy. Synchronous 2D visible correlation analysis revealed that there are at least two bands around 445 and 560 nm, decreasing in intensity with cooking time. The corresponding asynchronous spectrum indicated that the spectral intensity reduction of the two bands occurs before the intensity variation of the 475, 520, and 585 nm bands. It suggested that the 445 and 560 nm bands be assigned to deoxymyoglobin and oxymyoglobin, respectively, and at least one of the 475, 520, and 585 nm bands assigned to the denatured species (metmyoglobin) of myoglobin. Also, the asynchronous 2D NIR correlation spectrum indicated that C-H fractions are easily oxidized. In addition, strong correlation peaks correlating the bands in the visible, SW-NIR, and NIR spectral regions were observed and discussed. PMID- 10725172 TI - Namibian chewing stick, Diospyros lycioides, contains antibacterial compounds against oral pathogens. AB - The twigs of Diospyros lycioides, a plant commonly known as "muthala", are frequently used as chewing sticks for the cleaning of teeth by rural and urban people in Namibia. Preliminary studies showed that a methanol extract of D. lycioides inhibited growth of selected oral pathogens. Subsequent bioassay-guided fractionation led to the isolation of four novel bioactive naphthalene glycosides, diospyrosides A, B, C, and D (1-4), and two known bioactive naphthoquinones, juglone (5) and 7-methyljuglone (6). The structures of the new compounds were elucidated using spectroscopic techniques including 1D and 2D NMR. These compounds inhibited the growth of oral cariogenic bacteria (Streptococcus mutans and Streptococcus sanguis) and periodontal pathogens (Porphyromonas gingivalis and Prevotella intermedia) at minimum inhibitory concentrations ranging from 0.019 to 1.25 mg/mL. Juglone exhibited the strongest inhibitory activity among these compounds. PMID- 10725173 TI - Distribution of folpet on the grape surface after treatment. AB - Field trials were carried out to evaluate whether folpet sprayed on grapevines penetrated the epicuticular wax and cell walls of grapes. Folpet showed poor penetration into the epicuticular wax; it was found almost totally on the surface. Despite its low solubility in water, perhaps due to the presence of adjuvants, its residues showed such a high resistance to washing that the action of rain was negligible in decreasing residues. PMID- 10725174 TI - Investigation on fungicide residues in greenhouse-grown strawberries. AB - Maximum residue limits (MRL's) for different agricultural food products in Norway are harmonized with EU standards. In field-grown strawberries in Norway, tolylfluanid has a 7 day quarantine from last application to harvest, while other approved fungicides have 14 days quarantine. Greenhouse production of strawberries is newly introduced to the country. Residue levels in strawberries of the cultivar Korona grown in a commercial greenhouse were investigated 4, 7, and 14 days after application of eight different fungicides at rates recommended by the manufacturers and at half rates. Iprodione, tolylfluanid, and vinclozolin were tested in two experiments, while chinomethionat, chlorothalonil, imazalil, penconazole, and triadimefon were tested once. For chinomethionat, imazalil, iprodione, penconazole, and vinclozolin, the residue levels were below MRL 2 weeks after application. Application of triadimefon in normal rate gave residues below MRL 14 days after application. However, its metabolite, triadimenol, was above MRL at the same time. Tolylfluanid gave very high residue levels, and except from half concentration in the second experiment, all other residue levels were above MRL. Seven days after application, residues in both experiments were approximately 3 times higher than MRL when normal rate of tolylfluanid was applied. For chlorothalonil at the recommended rate, the residue level was above MRL at any sampling time, while half rate gave residues below MRL 14 days after treatment. In view of the present results, tolylfluanid, chlorothalonil, and triadimefon will need longer time from last application to harvest and/or reduced application rates in greenhouse-grown compared to field-grown strawberries. In addition or as an alternative, recommended rates could be lowered. PMID- 10725175 TI - Identification and effects of maize silk volatiles on cultures of Aspergillus flavus. AB - Volatiles generated from corn silks of individual genotypes of maize were found to exhibit differences in biological activities when the volatiles were exposed to 5-day solid cultures of Aspergillus flavus. In inverted potato dextrose-agar Petri plate bioassays, it was found that volatiles emitted from silks of the different maize genotypes had a profound effect on the growth of the fungus and, consequently, aflatoxin production. To determine the underlying cause for this bioactivity, volatiles emitted from the maize silks were trapped on Tenax glass columns and were analyzed by GC-MS. Aflatoxin field-resistant maize genotypes exhibited a larger relative concentration of the antifungal aldehyde, furfural (2 furancarboxaldehyde), when compared to the relative concentrations of the field susceptible varieties tested. In a closed-container 5-day study, it was observed that fresh 1- and 4-day-old corn silk samples of aflatoxin-resistant maize genotypes emitted higher concentrations of furfural compared to those from susceptible genotypes. This observation probably explains the reason for the bioactivity observed in the in vitro bioassays, and the presence of furfural appears to contribute to a defense mechanism for protecting the developing maize kernel from fungal attack. PMID- 10725176 TI - Comparative three-dimensional quantitative structure-activity relationship study of safeners and herbicides. AB - The competitive antagonist hypothesis for safeners and herbicides was investigated by studying the 3D similarity between 28 safener and 20 herbicide molecules in their putative biologically active, low-energy conformations using comparative molecular field analysis (CoMFA). In addition, CoMFA provided information about the structural requirements for the interactions of safeners and herbicides with a proteinaceous component (SafBP) isolated from etiolated corn seedlings. Statistically significant CoMFA models have been developed for the united and separate safener and herbicide molecule sets using retrospective binding affinity data of the ligands measured at the SafBP receptor. The predictive power of the models was characterized by squared cross-validated correlation coefficients (q(2)) of 0.708, 0.564, and 0.4000 for the united safener plus herbicide set, the safener set, and the herbicide set, respectively. The CoMFA results support the competitive antagonist hypothesis between certain types of safeners and herbicides. The findings suggest that structural similarity between these two classes of agrochemicals is a useful guide in the design of new safeners. PMID- 10725177 TI - Azo dye-mediated regulation of total phenolics and peroxidase activity in thyme (Thymus vulgaris L.) and rosemary (Rosmarinus officinalis L.) clonal lines. AB - Thyme (Thymus vulgaris L.) and rosemary (Rosmarinus officinalis L.) clonal lines, which were previously isolated from a heterogeneous seed population by plant tissue culture techniques, have been targeted as potential plants for phytoremediation of organic pollutants such as azo dyes and related aromatic compounds. Three thyme clonal lines and three rosemary clonal lines were tested for the ability to grow on hormone-free medium containing 0.01% of azo dye Poly S 119. The results showed that dye tolerance was associated with reduced phenolics and enhanced peroxidase activity in these clonal lines. There was a clear inverse correlation between total phenolics and peroxidase activity in these plants in response to Poly S-119. The tolerance of these clonal lines showed variations at different growing stages. These observations suggested that the peroxidase activity was inducible. Because peroxidases are involved in lignification, wound healing, aromatic compound degradation, pathogen defense, and stiffening, the results suggest that azo dye stimulated the defense response of thyme and rosemary clonal plants by increasing the peroxidase activity. Stereomicroscopic observations revealed that the azo dye was sequestered within the growing axis of the plant roots, which may also enhance the polymerization of azo dye onto the cell wall with the help of enhanced peroxidase activity. PMID- 10725178 TI - Controlled release of carbofuran from an alginate-bentonite formulation: water release kinetics and soil mobility. AB - The insecticide-nematicide carbofuran was incorporated in alginate-based granules to obtain controlled-release (CR) properties. The basic formulation [sodium alginate (1.61%)-carbofuran (0. 59%)-water] was modified by addition of sorbents. The effect on carbofuran release rate, caused by the incorporation of natural and acid-treated bentonite (0.5 and 1.0 M H(2)SO(4)) in alginate formulation, was studied by immersion of the granules in water under shaking. The time taken for 50% of the active ingredient to be released into water, t(50), was longer for those formulations containing natural bentonite (6.1 h) or acid-treated bentonite (9.0 and 11.7 h for 0.5 and 1.0 M H(2)SO(4) treatments, respectively) than for the preparation without bentonite (4.7 h). It appears from the results that the release of carbofuran from the various formulations is controlled by a diffusion mechanism according to the n values obtained, which were close to 0.5 in all cases. The mobility of carbofuran from alginate-based CR formulations was investigated by using soil columns packed with a clay soil (53% clay and 0.08% organic matter). Two alginate-based CR formulations containing natural bentonite or acid-treated bentonite (0.5 M H(2)SO(4)) were compared to technical grade carbofuran. The use of alginate-based CR formulations resulted in a reduction of the leached amount of carbofuran compared with the total amount of pesticide leached using the technical product (50 and 75% for CR granules containing natural and acid-treated bentonite, respectively). Alginate-bentonite CR formulations might be efficient systems for reducing carbofuran leaching in clay soils, which would reduce the risk of groundwater pollution. PMID- 10725179 TI - Photodegradation of metolachlor: isolation, identification, and quantification of monochloroacetic acid. AB - The photolysis of metolachlor [2-chloro-N-(2-ethyl-6-methylphenyl)-N-(2-methoxy-1 methylethyl) acetamide] in a sunlight simulator under actinic radiation was investigated. The focus of the study was to determine the extent of monochloroacetic acid (MCA) production. MCA was concentrated and derivatized from photolysate as the n-propyl ester using propanol and sulfuric acid and then identified as the ester using GC/MS and GC/ECD. On the basis of regression analysis, it was shown that the direct photodegradation of approximately 10 microM metolachlor followed pseudo-first-order kinetics with respect to the metolachlor concentration, and the half-life of the herbicide ( approximately 74 h) was independent of the pH of the medium. Photolysis in synthetic field water (SFW) resulted in a significant reduction of photolysis time (t(1/2) approximately 9 h). Direct photolysis experiments indicate a 5.19 +/- 0.81% (n=3) conversion of metolachlor to MCA, while photolysis in synthetic field water and in a Don River water sample resulted in 29.8 +/- 4.6% (n = 3) and 12.6 +/- 4.1% (n = 3) conversion, respectively; MCA was shown to be hydrolytically stable over the time course of the photoreaction. The photodegradation of alachlor, butachlor and a model chloroacetanilide, 2-chloro-N-methylacetanilide, in SFW were also investigated. PMID- 10725180 TI - Heterologous expression in Aspergillus nidulans of a Trichoderma longibrachiatum endoglucanase of enological relevance. AB - An Aspergillus nidulans transformant expressing the Trichoderma longibrachiatum endoglucanase 1 gene (egl1) has been constructed. The extracellular production of EGL1 in different culture media has been studied, and a medium has been found in which EGL1 is the predominant extracellular protein produced. The enzymatic properties of the heterologously produced EGL1 are very similar to those of the native enzyme. Grape maceration in the presence of culture filtrate enriched in EGL1 resulted in increased release of aroma precursors, particularly in the case of aromatic grapes. Cryoscanning electron microscopy of the flesh of grapes treated with EGL1-enriched culture filtrate revealed degradation of the cell wall matrix. PMID- 10725181 TI - Delignified cellulosic material supported biocatalyst as freeze-dried product in alcoholic fermentation. AB - Freeze-dried delignified cellulosic (DC) material supported biocatalyst is proposed as a suitable form of biocatalyst to be preserved. The alcoholic fermentation of glucose using freeze-dried immobilized cells is reported. Freeze dried immobilized baker's yeast cells on DC material do not need any protective medium during freeze-drying. The effect of initial glucose concentration and temperature on the alcoholic fermentation kinetic parameters is reported in the present study. It was found that the freeze-dried immobilized cells ferment more quickly than free freeze-dried cells and have a lower fermentation rate as compared with wet immobilized cells. However, repeated batch fermentations showed freeze-dried immobilized cells to ferment at about the same fermentation rate as wet immobilized cells. The results indicate that the freeze-dried immobilized cells must be further studied to establish a process for the preservation of immobilized cells. PMID- 10725182 TI - Cycloamylose (cyclodextrin) glucanotransferase degrades intact granules of potato raw starch. AB - Cycloamylose (cyclodextrin) glucanotransferase (EC 2.4.1.19, CGTase) originated from Bacillus macerans degraded intact granules of potato raw starch and converted them into cyclodextrins (CDs). The degradation required sufficient stirring of starch-CGTase suspension. The morphology of the degraded starch granules was unique; that is, the inner part of the granules was observed by scanning electron microscope to be more susceptible to CGTase than the outer part. Effects of pH, temperature, starch concentration, and enzyme amount on CD production were studied. PMID- 10725183 TI - The network of brain areas involved in the motion aftereffect. AB - A network of brain areas is expected to be involved in supporting the motion aftereffect. The most active components of this network were determined by means of an fMRI study of nine subjects exposed to a visual stimulus of moving bars producing the effect. Across the subjects, common areas were identified during various stages of the effect, as well as networks of areas specific to a single stage. In addition to the well-known motion-sensitive area MT the prefrontal brain areas BA44 and 47 and the cingulate gyrus, as well as posterior sites such as BA37 and BA40, were important components during the period of the motion aftereffect experience. They appear to be involved in control circuitry for selecting which of a number of processing styles is appropriate. The experimental fMRI results of the activation levels and their time courses for the various areas are explored. Correlation analysis shows that there are effectively two separate and weakly coupled networks involved in the total process. Implications of the results for awareness of the effect itself are briefly considered in the final discussion. PMID- 10725184 TI - An MRI-based parcellation method for the temporal lobe. AB - The temporal lobe has long been a focus of attention with regard to the underlying pathology of several major psychiatric illnesses. Previous postmortem and imaging studies describing regional volume reductions or perfusion defects in temporal subregions have shown inconsistent findings, which are in part due to differences in the definition of the subregions and the methodology of measurement. The development of precise reproducible parcellation systems on magnetic resonance images may help improve uniformity of results in volumetric MR studies and unravel the complex activation patterns seen in functional neuroimaging studies. The present study describes detailed guidelines for the parcellation of the temporal neocortex. It parcels the entire temporal neocortex into 16 subregions: temporal pole, heschl's gyrus, planum temporale, planum polare, superior temporal gyrus (rostral and caudal), middle temporal gyrus (rostral, intermediate, and caudal), inferior temporal gyrus (rostral, intermediate, and caudal), occipitotemporal gyrus (rostral and caudal), and parahippocampal gyrus (rostral and caudal). Based upon topographic landmarks of individual sulci, every subregion was consecutively traced on a set of serial coronal slices. In spite of the huge variability of sulcal topography, the sulcal landmarks could be identified reliably due to the simultaneous display of three orthogonal (transaxial, coronal, and sagittal) planes, triangulated gray matter isosurface, and a 3-D-rendered image. The reliability study showed that the temporal neocortex could be parceled successfully and reliably; intraclass correlation coefficient for each subregion ranged from 0.62 to 0.99. Ultimately, this method will permit us to detect subtle morphometric impairments or to find abnormal patterns of functional activation in the temporal subregions that might reflect underlying neuropathological processes in psychiatric illnesses such as schizophrenia. PMID- 10725185 TI - How good is good enough in path analysis of fMRI data? AB - This paper is concerned with the problem of evaluating goodness-of-fit of a path analytic model to an interregional correlation matrix derived from functional magnetic resonance imaging (fMRI) data. We argue that model evaluation based on testing the null hypothesis that the correlation matrix predicted by the model equals the population correlation matrix is problematic because P values are conditional on asymptotic distributional results (which may not be valid for fMRI data acquired in less than 10 min), as well as arbitrary specification of residual variances and effective degrees of freedom in each regional fMRI time series. We introduce an alternative approach based on an algorithm for automatic identification of the best fitting model that can be found to account for the data. The algorithm starts from the null model, in which all path coefficients are zero, and iteratively unconstrains the coefficient which has the largest Lagrangian multiplier at each step until a model is identified which has maximum goodness by a parsimonious fit index. Repeating this process after bootstrapping the data generates a confidence interval for goodness-of-fit of the best model. If the goodness of the theoretically preferred model is within this confidence interval we can empirically say that the theoretical model could be the best model. This relativistic and data-based strategy for model evaluation is illustrated by analysis of functional MR images acquired from 20 normal volunteers during periodic performance (for 5 min) of a task demanding semantic decision and subvocal rehearsal. A model including unidirectional connections from frontal to parietal cortex, designed to represent sequential engagement of rehearsal and monitoring components of the articulatory loop, is found to be irrefutable by hypothesis-testing and within confidence limits for the best model that could be fitted to the data. PMID- 10725186 TI - Three distinct auditory areas of cortex (AI, AII, and AAF) defined by optical imaging of intrinsic signals. AB - Using pure-tone sound stimulation, three separate auditory areas are revealed by optical imaging of intrinsic signals in the temporal cortex of the chinchilla (Chinchilla laniger). These areas correlate with primary auditory cortex (AI) and two secondary areas, AII and the anterior auditory field (AAF). We have distinguished AI on the basis of concurrent single-unit electrophysiological recording; neurons within the AI intrinsic signal region have short (<15 ms) onset-response latencies compared with neurons recorded in AII and the AAF. Within AI, AII, and AAF we have been able to define cochleotopic or tonotopic organization from the differences in intrinsic signal areas evoked by pure tones at octave-spaced frequencies from 500 Hz to 16 kHz. The maps in AI and AII are arranged orthogonal to each other. PMID- 10725187 TI - A principal components-based method for the detection of neuronal activity maps: application to optical imaging. AB - We present a novel analysis technique for the extraction of neuronal activity patterns from functional imaging data. We illustrate this technique on data from optical imaging. Optical imaging of the mammalian visual cortex probe the patterns in which the neuronal responses to various aspects of the visual world, such as orientation and color, are spatially organized within the cortex. Recovering these patterns from the image data is a challenging problem as the neuronal response signal is extremely weak in comparison to the background vegetative processes (e.g., circulation and respiration). The proposed technique obtains the neuronal activity pattern using a combination of principal component analysis and statistical significance testing. The performance of this method is compared with the results of existing analysis techniques. The comparison shows the new method to be more sensitive than previous methods. PMID- 10725188 TI - A study of analysis parameters that influence the sensitivity of event-related fMRI analyses. AB - To assess the effect of various analysis parameters on the sensitivity of event related fMRI analyses, 36 analyses were performed on a single fMRI data-set, varying parameters along four axes: (1) resampled voxel size; (2) spatial smoothing; (3) temporal smoothing; and (4) the set of basis functions used to model event-related responses. Sensitivity (i.e., the probability of detecting an activation given it exists) was assessed in terms of Z scores and by a metric for corrected P values, the negative log of the expected Euler characteristic. Sixteen brain regions distributed across cortical and subcortical areas were included in the meta-analysis. Main effects on sensitivity were found for resampled voxel size, spatial smoothing, temporal smoothing, and the set of basis functions chosen. The analysis parameters that generally produced the most sensitive analyses were a 2-mm(3) resampled voxel size, 10-mm spatial smoothing, 4-s temporal smoothing, and a basis set comprising a hemodynamic response function and its temporal derivative. PMID- 10725189 TI - Differential effects of muscle contraction from various body parts on neuromagnetic somatosensory responses. AB - We studied eight healthy subjects with a whole-scalp 306-channel neuromagnetometer to explore the effect of motor activity from different body parts on somatosensory responses to left median nerve stimulation. The stimuli produced clear tactile sensation without any motor movement. In the rest condition, the subject had no task. During contraction conditions, the subject had to maintain submaximal isometric contraction in masseter, left deltoid, left thenar, or left tibialis muscles. Short-latency responses from the primary somatosensory cortex did not change during contraction. Responses from both the right (contralateral) and left second somatosensory cortices (SII) were significantly enhanced during contraction of the left thenar muscles. Responses from the left SII were significantly enhanced also during contraction of the left deltoid muscles, but they were decreased during contraction of the masseter and left tibialis anterior muscles. This study implies that SII activation is modulated by motor activity and that the effect depends on the topographical proximity of the stimulated and contracted body parts. PMID- 10725190 TI - Visualization of subthalamic nuclei with cortex attenuated inversion recovery MR imaging. AB - There is a significant amount of interest in studying the thalamus because of its central location in the brain and its role as a gatekeeper to higher centers of cognition. Imaging and measuring of the individual subnuclei of the thalamus has proven extremely difficult in MR because of the contrast-to-noise (CNR) of the MR sequences used. This report describes a novel MR pulse sequence known as cortex attenuated inversion recovery (CAIR), which increases the CNR in images and allows the individual subnuclei of the thalamus to be visualized by selectively nulling the gray matter in the brain using an inversion recovery sequence with an inversion time of 700 ms at 1.5 T. PMID- 10725191 TI - A common language network for comprehension and production: a contribution to the definition of language epicenters with PET. AB - In this paper, we report on a PET activation study designed to assess whether functional neuroimaging would help to uncover essential language areas in normal volunteers and to provide a more accurate definition of their localization. Regional cerebral blood flow was repeatedly monitored in eight right-handed male volunteers, while performing a language comprehension task (listening to factual stories) and a language production task (covert generation of verbs semantically related to heard nouns), using silent resting as a control condition. The conjunction analysis, conducted with SPM, was used to uncover the network of activations common to both task that included three left hemisphere areas, namely (1) the pars opercularis and triangularis of the inferior frontal gyrus, (2) the posterior part of the superior temporal cortex centered around the superior temporal sulcus, extending to the planum temporale posterior part but sparing the supramarginalis and angular gyri, and (3) the most anterior part of the left inferior temporal gyrus at the junction with the anterior fusiform gyrus. The inferior and lateral parts of the right cerebellar cortex were also included in the conjunction network. Each of the three cortical areas, when they are site of lesion or electrical stimulation, elicit impairment in both language comprehension and production and can thus be considered as essential to language. Accordingly, the present results provide conservative anatomofunctional definitions of the Broca, Wernicke, and basal language areas. Interestingly, contralateral homologues of Broca's and Wernicke's areas also lighted up in the conjunction analysis that could be related to the interindividual variability of hemispheric language dominance. PMID- 10725192 TI - Stimulation of dengue virus replication in cultured Aedes albopictus (C6/36) mosquito cells by the antifungal imidazoles ketoconazole and miconazole. AB - Replication of dengue type 3 virus in Aedes albopictus C6/36 cells was enhanced more than 50-fold by addition of the antifungal imidazole derivative ketoconazole within the first 4 h of infection. The stimulatory effect was reflected in the yield of infectious virus and in levels of viral RNA and protein synthesis. Enhanced yields were observed also for other flaviviruses, including dengue type 2 virus and Murray Valley encephalitis virus. Increased yields of dengue type 3 virus were not observed in African green monkey kidney (Vero) cells, human monocytic (U-937) cells, or cells of the mosquito Toxorhynchites amboinensis (TRA 171). PMID- 10725193 TI - Identification of regions in the Moloney murine leukemia virus SU protein that tolerate the insertion of an integrin-binding peptide. AB - Targeting of retroviral vectors to specific cells has been attempted through engineering of the surface (SU) protein of the murine leukemia viruses (MuLVs), but in many cases this has adversely affected protein function and targeted delivery has been difficult to achieve. In this study, we have inserted a 15-mer peptide that binds specifically to the alpha(v)beta(3) integrin into the Moloney MuLV SU protein, including regions that are surface exposed in the crystal structure of the ecotropic receptor-binding domain. We have concentrated in particular on the variable regions VRA, VRB, and VRC, which are responsible for the use of distinct cellular receptors by different MuLV subtypes and therefore may be more likely to accommodate a heterologous binding moiety. Despite these considerations, only 8 of 26 insertion sites were tolerated, including two separate regions in VRA, a cluster of sites in VRC, and previously identified sites at the N-terminus of the protein and in the proline-rich region immediately downstream of the receptor-binding domain. When expressed on retroviral vector particles, all of the viable proteins retained the ability to bind to and transduce murine cells, although the VRC mutants and an insertion in VRA gave reduced binding and titer. Finally, although all of the viable chimeras could bind to alpha(v)beta(3) in a solid-phase binding assay, we were unable to demonstrate expanded tropism for alpha(v)beta(3)-expressing human cells. This study highlights the difficulty of engineering the Moloney MuLV SU protein, even when structural information is available, and provides guidelines for the insertion of peptide ligands into the SU protein. PMID- 10725194 TI - Glycoprotein B of human herpesvirus 8 is a component of the virion in a cleaved form composed of amino- and carboxyl-terminal fragments. AB - Human herpesvirus 8 (HHV-8) or Kaposi's sarcoma-associated herpesvirus (KSHV) is the only known human member of the Rhadinovirus genus of the gammaherpesvirus subfamily. Antibodies against peptides representing portions of the amino- and carboxyl-termini of HHV-8 gB were produced and used to detect gB expression in Vero cells transfected with the gB gene, in the HHV-8-harboring cell line, BCBL 1, and in purified virions. Expression of gB was detected in approximately 3% of uninduced BCBL-1 cells, while up to 30% of the cells expressed gB after 12-O tetradecanoylphorbol-13-acetate (TPA) induction of virus replication. Indirect immunofluorescence assays and confocal microscopy showed that gB was distributed throughout the cytoplasm of BCBL-1 cells and transfected Vero cells. Immunoblot analyses of virion preparations revealed the presence of full-length as well as two smaller than full-length gB-derived species corresponding to the amino- and carboxy-terminal portions of gB, respectively. Biochemical analysis of the gB carbohydrate moieties using glycosylation inhibitors revealed that gB contained N linked oligosaccharides of the high-mannose type, characteristic of precursor carbohydrate chains added in the endoplasmic reticulum. PMID- 10725195 TI - Synergistic interactions of a potyvirus and a phloem-limited crinivirus in sweet potato plants. AB - When infecting alone, Sweet potato feathery mottle virus (SPFMV, genus Potyvirus) and Sweet potato chlorotic stunt virus (SPCSV, genus Crinivirus) cause no or only mild symptoms (slight stunting and purpling), respectively, in the sweet potato (Ipomoea batatas L. ). In the SPFMV-resistant cv. Tanzania, SPFMV is also present at extremely low titers, though plants are systemically infected. However, infection with both viruses results in the development of sweet potato virus disease (SPVD) characterized by severe symptoms in leaves and stunting of the plants. Data from this study showed that SPCSV remains confined to phloem and at a similar or slightly lower titer in the SPVD-affected plants, whereas the amounts of SPFMV RNA and CP antigen increase 600-fold. SPFMV was not confined to phloem, and the movement from the inoculated leaf to the upper leaves occurred at a similar rate, regardless of whether or not the plants were infected with SPCSV. Hence, resistance to SPFMV in cv. Tanzania was not based on restricted virus movement, neither did SPCSV significantly enhance the phloem loading or unloading of SPFMV. It is also noteworthy that SPVD is an unusual synergistic interaction in that the potyvirus component is not the cause of synergism but is the beneficiary. It is hypothesized that SPCSV is able to enhance the multiplication of SPFMV in tissues other than where it occurs itself, perhaps by interfering with systemic phloem-dependent signaling required in a resistance mechanism directed against SPFMV. PMID- 10725196 TI - Membrane fusion activity of tick-borne encephalitis virus and recombinant subviral particles in a liposomal model system. AB - We present a kinetic analysis of the membrane fusion activity of tick-borne encephalitis (TBE) virus and TBE-derived recombinant subviral particles (RSPs) in a liposomal model system. Fusion was monitored using a fluorescence assay involving pyrene-labeled phospholipids. Fusion was strictly dependent on low pH, with the optimum being at pH 5.3-5.5 and the threshold at pH 6.8. Fusion did not require a protein or carbohydrate receptor in the target liposomes. Preexposure to low pH of the virus alone resulted in inactivation of its fusion activity. At the optimum pH for fusion and 37 degrees C, the rate and extent of fusion were very high, with more than 50% of the virus fusing within 2 s and the final extent of fusion being 70%. Lowering of the temperature did not result in a significant decrease in the rate and extent of fusion, suggesting that TBE virus fusion is a facile process with a low activation energy, possibly due to the flat orientation of the E glycoprotein on the viral surface facilitating the establishment of direct intermembrane contact. The fusion characteristics of TBE virus and RSPs were similar, indicating that RSPs provide a reliable and convenient model for further study of the membrane fusion properties of TBE virus. PMID- 10725197 TI - Immunisation with phage displaying peptides representing single epitopes of the glycoprotein G can give rise to partial protective immunity to HSV-2. AB - Filamentous phage displaying peptides representing single epitopes of the glycoprotein G of HSV-2 (gG2) were used as immunogens via the subcutaneous route in Balb/c mice without additional adjuvant. The phage were isolated from a random phage peptide display library and contain 15-mer peptide inserts that mimic epitopes of gG2. In each case, an antibody response to gG2 was generated that was dependent on the dose of phage administered and on the presence of the peptide insert. Phage displaying epitopes of gG2, which map to amino acids 551-570, were the most immunogenic; interestingly, this region of gG2 is frequently recognised by patients infected with HSV-2. The data also provide interesting information as regards choice of peptide mimics for use as immunogens because, surprisingly, the most antigenic of the individual clones was the least immunogenic. In two of the experiments, mice immunised with phage displaying a single epitope of gG2 were protected against challenge with a lethal dose of whole HSV-2. This suggests a possible role for phage-displayed peptides in inducing protective immunity against pathogens and provides a model system for investigating the underlying mechanisms. PMID- 10725198 TI - Plasmid DNA encoding the respiratory syncytial virus G protein is a promising vaccine candidate. AB - Respiratory syncytial virus (RSV) remains a major cause of severe respiratory diseases in infants, young children, and the elderly. However, development of a RSV vaccine has been hampered by the outcome of the infant trials in the 1960s with a formalin-inactivated RSV preparation. Enhanced lung disease was induced by the vaccination post-RSV exposure. Previous studies in mice primed with RSV G protein either formulated in adjuvants or delivered by recombinant vaccinia viruses have indicated that enhanced lung pathology resulted from a Th2-type host immune response against the viral G protein. However, in the present report, we have demonstrated that vaccination with plasmid vectors encoding either a full length or a secreted G protein (DNA-G) clearly elicited balanced systemic and pulmonary Th1/Th2 cytokine responses in mice and did not induce an atypical pulmonary inflammatory reaction post-RSV challenge in cotton rats. DNA-G immunization also induced marked virus neutralizing antibody responses and protection against RSV infection of the lower respiratory tract of both mice and cotton rats. So far, only genetic immunization has been able to induce a balanced Th1/Th2 response with the RSV G protein, reminiscent of that induced by live RSV. Therefore, DNA-G is a promising immunogen for inclusion in a nucleic acid RSV vaccine. PMID- 10725199 TI - Local IL-4 expression in the lung reduces pulmonary influenza-virus-specific secondary cytotoxic T cell responses. AB - We studied the effect of lung-specific IL-4 expression on the T cell response during primary and secondary heterologous infection with influenza virus by using transgenic mice that express IL-4 under a lung-specific promoter. Subsequent to primary infection with a type A/H1N1 influenza virus these transgenic mice exhibited similar local recruitment of CD4(+) and CD8(+) T cells and only slightly decreased virus-specific CTL activity. However, during secondary challenge with a heterologous influenza virus, the local infiltration with virus specific, MHC class I-restricted CD8(+) T cells was significantly decreased compared to that of nontransgenic littermates. The ability of IL-4 transgenic mice to clear the heterologous infection was delayed but not abrogated. This was associated with a faster virus-neutralizing antibody response in IL-4 transgenic mice and with their ability to mount significant Th1 responses even in the presence of increased local IL-4 expression. Our observations demonstrate a negative regulatory effect of IL-4 on memory Tc1/CD8(+) T cells, but are also consistent with complementary mechanisms important for virus clearance such as virus-neutralizing antibodies. The reduction of memory CTL in the presence of IL 4 might have consequences for understanding the course of influenza infection in situations where T(H)2 immunity is increased. PMID- 10725200 TI - Productive infection of CD34+-cell-derived megakaryocytes by X4 and R5 HIV-1 isolates. AB - The human immunodeficiency virus (HIV-1) causes various hematopoietic abnormalities, with thrombocytopenia (TP) occurring in 30% of infected individuals. In the present study, we aimed to determine whether HIV-1 in the bone marrow of TP patients can infect primary megakaryocytes in vitro, which may contribute to the development of thrombocytopenia. We amplified the V3 loop of HIV-1 envelope from the bone marrow of TP and non-TP patients and constructed recombinant viruses. We demonstrate that the bone marrow of TP and non-TP patients contained R5 strains, whereas X4 strains were present only in the bone marrow of TP patients. Furthermore, HIV-1 from the bone marrow of TP and non-TP patients infected megakaryocytes to similar levels, suggesting that the V3 loop of HIV-1 may not contain the viral determinants of HIV-associated TP. Chemokine receptor analysis determined that CD34+-cell-derived megakaryocytes express CD4, CXCR4, and CCR5 and are productively infected by both X4 and R5 HIV-1 isolates. Finally, we showed that CD34+-cell-derived megakaryocytes express the chemokine receptor CCR3. PMID- 10725201 TI - A point mutation in VP1 of coxsackievirus B4 alters antigenicity. AB - While coxsackievirus infections have been linked to several autoimmune diseases, very little is known about the immunogenicity of the coxsackieviruses. Using two genetically related variants of coxsackievirus B4, CB4-P and CB4-V, the relationship between virulence and antigenicity was examined. The virulent variant, CB4-V, was shown to be more antigenic than the avirulent CB4-P variant. The increased antigenicity of CB4-V was due to a single amino acid substitution in the VP1 capsid protein (a threonine residue at amino acid position 129), a site that had been previously identified as a major determinant of viral virulence. Thr-129 of VP1 is predicted to lie within a conformational B cell epitope. In addition, a nearby linear B cell epitope spanning residues 68 to 82 of VP1 was identified as a potential serotype-specific, neutralization antigenic site. The linear and conformational B cell epitopes of coxsackievirus B4 may be analogous to antigenic sites 1 and 1B of poliovirus. To address whether the increased antigenicity of CB4-V influenced the severity of disease, mouse strains that differ in their outcome to viral infection were analyzed. Mice that developed the most severe disease and succumbed to infection were more immunoresponsive than mice that survived infection with CB4-V. The data suggest that immune-mediated mechanisms play a role in the severity of CB4-V induced disease. PMID- 10725202 TI - Virtually full-length subtype F and F/D recombinant HIV-1 from Africa and South America. AB - For reliable classification of HIV-1 strains appropriate reference sequences are needed. The HIV-1 genetic subtype F has a wide geographic spread, causing significant epidemics in South America, Africa, and some regions of Europe. Previously only two full-length sequences of each of the HIV-1 subtype F subclusters F1 and F2 have been described. To extend the knowledge of subtype F variation on a complete genome level, three new virtually full-length F1 sequences were cloned and sequenced, two from Africa and one from South America. Comparison of the new and previously described sequences showed that monophyletic clustering of the subcluster F1 of subtype F is consistent and highly supported in all genome regions. Two additional full-length strains were shown to be mosaics of subtypes F and D. These epidemiologically unrelated F/D sequences showed similar chimeric structure, suggesting that they may represent a previously undescribed circulating recombinant form (CRF). This was supported by partial sequences from three additional unlinked F/D recombinants. Genetic distances in the phylogenetic trees suggest that the recombination event leading to the putative CRF occurred relatively long ago, close to the divergence of the F1 and F2 subclusters. Furthermore, all five F/D recombinants are linked to the Democratic Republic of Congo, suggesting that the original recombination event took place in central Africa. PMID- 10725203 TI - Rhesus and pig-tailed macaque parvoviruses: identification of two new members of the erythrovirus genus in monkeys. AB - We have previously reported the identification of a novel simian parvovirus in cynomolgus monkeys, which causes severe anemia in immunosuppressed cynomolgus monkeys and is currently being studied as an animal model for human B19 infection. We now report two similar outbreaks of anemia in rhesus and pig-tailed macaques associated with two distinct but similar simian parvoviruses (pig-tailed macaque and rhesus parvovirus). Both viruses have been cloned and over 5000 nucleotides sequenced from each virus. The viruses show marked similarities to other members of the Erythrovirus genus in the Parvoviridae family. PMID- 10725204 TI - Subcellular sorting of small membrane-associated triple gene block proteins: TGBp3-assisted targeting of TGBp2. AB - We studied subcellular distribution of green fluorescent protein (GFP)-tagged movement proteins encoded by the second and the third genes of poa semilatent hordeivirus (PSLV) triple gene block (TGB), 15K TGBp2 and 18K TGBp3. GFP-15K transiently expressed in Nicotiana benthamiana leaf epidermal cells was associated with the endomembrane system elements. GFP-18K appeared in the membrane bodies at cell periphery. Mutation analysis demonstrated that subcellular targeting of GFP-15K depended on the protein transmembrane segment(s), whereas the TGBp3 central hydrophilic region was responsible for targeting of GFP-18K. Coexpression of GFP-15K with the intact 18K protein induced drastic changes in the TGBp2 localization: GFP-15K appeared in the cell peripheral bodies similar to those in the cells expressing GFP-18K alone. Coexpression experiments with mutant forms of both proteins argue against involvement of direct interaction between small TGB proteins in the TGBp3 assisted targeting of TGBp2 to the cell peripheral compartments. This conclusion was further confirmed by similar effects on the PSLV 15K TGBp2 localization induced by TGBp3 proteins of PSLV and potato virus X, which have no detectable sequence similarity to each other. PMID- 10725205 TI - Recombination within the nonstructural genes of the parvovirus minute virus of mice (MVM) generates functional levels of wild-type NS1, which can be detected in the absence of selective pressure following transfection of nonreplicating plasmids. AB - Recombination within the coding region of the nonstructural genes of minute virus of mice (MVM), which generates functional levels of wild-type NS1, was observed in the absence of selective pressure following cotransfection of nonreplicating plasmids. P38 activity was used as a measure of recombinant NS1 production, which, together with direct detection of recombinant-generated products by RT PCR, allowed an estimation of recombination efficiency. In addition, we show that very low levels of wild-type NS1 were able to significantly transactivate P38. Given that recombination following cotransfection can generate NS1 at these levels, our observations have implications for the study of parvoviral genetics, the construction of recombinant parvoviral vectors for gene therapy applications, and perhaps other systems using cotransfection of plasmids that share homologous sequences. PMID- 10725206 TI - The interaction of heparin sulfate and adeno-associated virus 2. AB - Recently heparan sulfate was proposed as the host cell receptor for the dependovirus, adeno-associated virus type 2 (AAV2). We show that although heparan sulfate on the cell surface may contribute to the binding of AAV2 to permissive cells, the amount of heparan sulfate on the cell surface as determined by flow cytometry using four different monoclonal antibodies does not correlate with AAV2 binding to cells or recombinant AAV2 transduction efficiency. Experiments with either mutant CHO cells or cells treated with chlorate to remove sulfate groups showed that sulfation was not absolutely required for infection or binding: in the absence of cell surface sulfation, recombinant AAV2 was still able to be transduced in previously permissive cells. Heparin is commonly used as a substitute in studies of the interaction between heparan sulfate and ligand, and we demonstrate that the binding affinity of AAV2/heparin is low, with a K(d) value of approximately 2.0 nM. A study of the direct interaction between AAV2 and artificial glycosaminoglycans showed that a high degree of sulfation on heparin was critical for the ability to bind AAV2 and compete rAAV2 transduction and that both O- and N-sulfate groups are required. Overall, our data suggest that, as has been shown for other viruses, the presence of a high-affinity AAV2 receptor mediates AAV2 infection in addition to the low-affinity heparan sulfate binding. PMID- 10725207 TI - Characterization and immunolocalization of major structural proteins in the brown algal virus EsV-1. AB - The Ectocarpus siliculosus virus (EsV-1) is endemic in all populations of the cosmopolitan filamentous brown alga Ectocarpus siliculosus. EsV-1 has a large circular double-stranded DNA genome of about 320 kilobase pairs, and a complex virion structure with a central nucleoprotein core surrounded by several proteinaceous layers. To investigate the protein composition of the virion, we screened an expression library of EsV-1 with antibodies raised against purified detergent-disrupted viral particles. We isolated several clones encoding novel structural proteins and investigated two of them in detail. These clones encode viral proteins vp55 and vp74. Electron microscopy reveals that vp55 is most likely a component of the surface of the viral core, whereas vp74 may be part of an inner core structure. To initiate a genetic analysis, we sequenced regions of the EsV-1 genome encoding vp55 and vp74 and found several adjacent open reading frames with the potential to code for several interesting viral proteins including a putative calcium-binding protein, a collagen-like protein, and a RING finger protein. PMID- 10725208 TI - Sugarcane yellow leaf virus: an emerging virus that has evolved by recombination between luteoviral and poleroviral ancestors. AB - We have derived the genomic nucleotide sequence of an emerging virus, the Sugarcane yellow leaf virus (ScYLV), and shown that it produces one to two subgenomic RNAs. The family Luteoviridae currently includes the Luteovirus, Polerovirus, and Enamovirus genera. With the new ScYLV nucleotide sequence and existing Luteoviridae sequence information, we have utilized new phylogenetic and evolutionary methodologies to identify homologous regions of Luteoviridae genomes, which have statistically significant altered nucleotide substitution ratios and have produced a reconstructed phylogeny of the Luteoviridae. The data indicate that Pea enation mosaic virus-1 (PEMV-1), Soybean dwarf virus (SbDV), and ScYLV exhibit spatial phylogenetic variation (SPV) consistent with recombination events that have occurred between poleroviral and luteoviral ancestors, after the divergence of these two progenitor groups. The reconstructed phylogeny confirms a contention that a continuum in the derived sequence evolution of the Luteoviridae has been established by intrafamilial as well as extrafamilial RNA recombination and expands the database of recombinant Luteoviridae genomes that are currently needed to resolve better defined means for generic discrimination in the Luteoviridae (D'Arcy, C. J. and Mayo, M. 1997. Arch. Virol. 142, 1285-1287). The analyses of the nucleotide substitution ratios from a nucleotide alignment of Luteoviridae genomes substantiates the hypothesis that hot spots for RNA recombination in this virus family are associated with the known sites for the transcription of subgenomic RNAs (Miller et al. 1995. Crit. Rev. Plant Sci. 14, 179-211), and provides new information that might be utilized to better design more effective means to generate transgene-mediated host resistance. PMID- 10725210 TI - Adoptive transfer of infectious bronchitis virus primed alphabeta T cells bearing CD8 antigen protects chicks from acute infection. AB - Infectious bronchitis virus (IBV) infection and associated illness may be dramatically modified by passive transfer of immune T lymphocytes. Lymphocytes collected 10 days postinfection were transferred to naive chicks before challenge with virus. As determined by respiratory illness and viral load, transfer of syngeneic immune T lymphocytes protected chicks from challenge infection, whereas no protection was observed in the chicks receiving the MHC compatible lymphocytes from uninfected chicks. Protection following administration of T lymphocytes could be observed in chicks with three distinct MHC haplotypes: B(8)/B(8), B(12)/B(12), and B(19)/B(19). Nearly complete elimination of viral infection and illness was observed in chicks receiving cells enriched in alphabeta lymphocytes. In contrast, removal of gammadelta T lymphocytes had only a small effect on their potential to protect chicks. The adoptive transfer of enriched CD8(+) or CD4(+) T lymphocytes indicated that protection was also a function primarily of CD8 bearing cells. These results indicated that alphabeta T lymphocytes bearing CD8(+) antigens are critical in protecting chicks from IBV infection. PMID- 10725209 TI - Downstream ribosomal entry for translation of coronavirus TGEV gene 3b. AB - Gene 3b (ORF 3b) in porcine transmissible gastroenteritis coronavirus (TGEV) encodes a putative nonstructural polypeptide of 27.7 kDa with unknown function that during translation in vitro is capable of becoming a glycosylated integral membrane protein of 31 kDa. In the virulent Miller strain of TGEV, ORF 3b is 5' terminal on mRNA 3-1 and is presumably translated following 5' cap-dependent ribosomal entry. For three other strains of TGEV, the virulent British FS772/70 and Taiwanese TFI and avirulent Purdue-116, mRNA species 3-1 is not made and ORF 3b is present as a non-overlapping second ORF on mRNA 3. ORF 3b begins at base 432 on mRNA 3 in Purdue strain. In vitro expression of ORF 3b from Purdue mRNA 3 like transcripts did not fully conform to a predicted leaky scanning pattern, suggesting ribosomes might also be entering internally. With mRNA 3-like transcripts modified to carry large ORFs upstream of ORF 3a, it was demonstrated that ribosomes can reach ORF 3b by entering at a distant downstream site in a manner resembling ribosomal shunting. Deletion analysis failed to identify a postulated internal ribosomal entry structure (IRES) within ORF 3a. The results indicate that an internal entry mechanism, possibly in conjunction with leaky scanning, is used for the expression of ORF 3b from TGEV mRNA 3. One possible consequence of this feature is that ORF 3b might also be expressed from mRNAs 1 and 2. PMID- 10725211 TI - Crosslink analysis of N-terminal, C-terminal, and N/B determining regions of the Moloney murine leukemia virus capsid protein. AB - To analyze contacts made by Moloney murine leukemia virus (M-MuLV) capsid (CA) proteins in immature and mature virus particles, we have employed a cysteine specific crosslinking approach that permits the identification of retroviral Gag protein interactions at particular residues. For analysis, single cysteine creation mutations were made in the context of protease-deficient or protease competent parental constructs. Cysteine creation mutations were chosen near the N and C-termini of CA and at a site adjacent to the M-MuLV Glu-Ala Fv1 N/B host range determination sequence. Analysis of immature virions showed that PrGag proteins were crosslinked at C-terminal CA residues to form dimers while crosslinking of particle-associated N-terminal and N/B region mutant proteins did not yield dimers, but showed evidence of linking to an unknown 140- to 160-kDa partner. Analysis of mature virions demonstrated that both N- and C-terminal CA residues participated in dimer formation, suggesting that processed CA N- and C termini are free to establish interprotein associations. Interestingly, N/B region mutant residues in mature virus particles did not crosslink to form dimers, but showed a novel crosslinked band, consistent with an interaction between the N/B tropism determining region and a cellular protein of 45-55 kDa. PMID- 10725212 TI - Precise mapping of the replication and transcription promoters of human parainfluenza virus type 3. AB - The terminal RNA regions of the genomic and antigenomic RNAs of the paramyxoviruses and rhabdoviruses are known to contain sequences essential for directing RNA replication and transcription. The 3' terminus (leader region) of the negative-sense, genomic RNA of the rhabdoviruses and paramyxoviruses is known as the leader (Le) promoter and directs synthesis of positive-sense replication and transcription products. The 3' terminus of the antigenome is termed the trailer complementary (TrC) promoter and directs the synthesis of genomic RNA. By creating mutations in the corresponding regions of an HPIV3 minireplicon in which the viral protein coding sequences were replaced by the luciferase gene, we were able to precisely define the elements of the leader promoter involved in directing positive-strand replication of HPIV3. Nucleotides 1 through 12 (from the terminus) formed a domain critical for replication. The region from nucleotides 13 through 55 was important but not crucial for replication, while G residues at positions 79, 85, and 91 comprised another domain critical for replication. It was also shown that the TrC promoter is similar, though not identical, to the Le promoter. Nucleotides 1 through 12 of the TrC promoter were critical for synthesis of genomic RNA, though specific positions behaved differently from the corresponding positions of the Le promoter. While many of these mutations could not be analyzed for transcription because they completely abrogated genomic RNA synthesis (the template for transcription), we were surprised to find that no mutations in the leader promoter which decreased replication had any significant effect on transcription. However, mutations in the intergenic sequence and gene start signal following the leader and preceding the luciferase message severely decreased transcription, but not replication. PMID- 10725213 TI - Coronavirus-induced membrane fusion requires the cysteine-rich domain in the spike protein. AB - The spike glycoprotein of mouse hepatitis virus strain A59 mediates the early events leading to infection of cells, including fusion of the viral and cellular membranes. The spike is a type I membrane glycoprotein that possesses a conserved transmembrane anchor and an unusual cysteine-rich (cys) domain that bridges the putative junction of the anchor and the cytoplasmic tail. In this study, we examined the role of these carboxyl-terminal domains in spike-mediated membrane fusion. We show that the cytoplasmic tail is not required for fusion but has the capacity to enhance membrane fusion activity. Chimeric spike protein mutants containing substitutions of the entire transmembrane anchor and cys domain with the herpes simplex virus type 1 glycoprotein D (gD-1) anchor demonstrated that fusion activity requires the presence of the A59 membrane-spanning domain and the portion of the cys domain that lies upstream of the cytoplasmic tail. The cys domain is a required element since its deletion from the wild-type spike protein abrogates fusion activity. However, addition of the cys domain to fusion defective chimeric proteins was unable to restore fusion activity. Thus, the cys domain is necessary but is not sufficient to complement the gD-1 anchor and allow for membrane fusion. Site-specific mutations of conserved cysteine residues in the cys domain markedly reduce membrane fusion, which further supports the conclusion that this region is crucial for spike function. The results indicate that the carboxyl-terminus of the spike transmembrane anchor contains at least two distinct domains, both of which are necessary for full membrane fusion. PMID- 10725214 TI - Infectious cDNA clones of Langat tick-borne flavivirus that differ from their parent in peripheral neurovirulence. AB - Tick-borne flavivirus strain Langat TP21 (LGT TP21) recovered from ticks, is naturally attenuated for humans but retains demonstrable neurovirulence and peripheral virulence ("neuroinvasiveness") for mice. Previously a mutant, strain E5, less virulent for mice was derived from LGT TP21. Multiple attempts to prepare a full-length infectious TP21 cDNA from cDNA fragments cloned in E. coli were uniformly unsuccessful. A more informative sequence than that obtained from these cloned cDNA fragments and similar E5 cDNA fragments was derived from RT-PCR fragments that had not been cloned in E. coli. Comparison of the RT-PCR consensus sequence of TP21 and E5 identified only seven amino acid differences that might be responsible for the observed difference in virulence of these strains for mice. Eleven independent infectious cDNA clones of TP21 were recovered using two overlapping long RT-PCR fragments. Importantly, low-titered virus used to prepare cDNA as template for PCR was harvested early in the growth cycle to minimize the frequency of deletion mutants that accumulated late in infection. The four analyzed rescued clones exhibited clone-specific minimal divergence from the consensus sequence but this limited variation was associated with diminished peripheral virulence for immunocompetent mice. Manipulation of these clones should facilitate elucidation of LGT virulence. PMID- 10725215 TI - Molecular motors--a paradigm for mutant analysis. AB - Molecular motors perform essential functions in the cell and have the potential to provide insights into the basis of many important processes. A unique property of molecular motors is their ability to convert energy from ATP hydrolysis into work, enabling the motors to bind to and move along cytoskeletal filaments. The mechanism of energy conversion by molecular motors is not yet understood and may lead to the discovery of new biophysical principles. Mutant analysis could provide valuable information, but it is not obvious how to obtain mutants that are informative for study. The analysis presented here points out several strategies for obtaining mutants by selection from molecular or genetic screens, or by rational design. Mutants that are expected to provide important information about the motor mechanism include ATPase mutants, which interfere with the nucleotide hydrolysis cycle, and uncoupling mutants, which unlink basic motor activities and reveal their interdependence. Natural variants can also be exploited to provide unexpected information about motor function. This general approach to uncovering protein function by analysis of informative mutants is applicable not only to molecular motors, but to other proteins of interest. PMID- 10725216 TI - The p120 catenin family: complex roles in adhesion, signaling and cancer. AB - p120 catenin (p120) is the prototypic member of a growing subfamily of Armadillo domain proteins found at cell-cell junctions and in nuclei. In contrast to the functions of the classical catenins (alpha-catenin, beta-catenin, and gamma catenin/plakoglobin), which have been studied extensively, the first clues to p120's biological function have only recently emerged, and its role remains controversial. Nonetheless, it is now clear that p120 affects cell-cell adhesion through its interaction with the highly conserved juxtamembrane domain of classical cadherins, and is likely to have additional roles in the nucleus. Here, we summarize the data on the potential involvement of p120 both in promotion of and in prevension of adhesion, and propose models that attempt to reconcile some of the disparities in the literature. We also discuss the structural relationships and functions of several known p120 family members, as well as the potential roles of p120 in signaling and cancer. PMID- 10725217 TI - Phospholipase A(2) and its products are involved in the purinergic receptor mediated translocation of protein kinase C in CHO-K1 cells. AB - The signal transduction involved in the purinergic stimuli-induced activation of protein kinase C (PKC) in CHO-K1 cells was investigated. Purinergic stimuli such as adenosine triphosphate and uridine triphosphate induced a transient translocation of PKC epsilon, gamma, and delta from the cytoplasm to the plasma membrane. These translocations were blocked by an inhibitor of phosphatidylinositol-specific phospholipase C (PLC), but not by an inhibitor of phosphatidylcholine-specific PLC. A diacylglycerol (DAG) analogue also induced reversible translocations of PKC gamma, epsilon, and delta from the cytoplasm to the plasma membrane, while the calcium ionophore A23187 caused a similar translocation of only the gamma subtype. These results confirm that the hydrolysis of phosphatidylinositol-2-phosphate by PLC and the subsequent generation of DAG and increase in Ca(2+ )are involved in the purinergic stimuli induced translocation of PKC. A DAG antagonist, 1-o-hexadecyl-2-o-acetyl glycerol, blocked the DAG analogue-induced translocations of all PKC subtypes tested but failed to inhibit the purinergic stimuli-induced translocations of PKC epsilon and gamma. The DAG antagonist could not block the ATP- and UTP-induced translocation of PKC epsilon even in the absence of extracellular Ca(2+). Co application of the DAG antagonist and a phospholipase A(2) (PLA(2)) inhibitor such as aristolochic acid, arachidonyltrifluoromethyl ketone, or bromoenol lactone inhibited the purinergic receptor-mediated translocation of PKC epsilon although each PLA(2) inhibitor alone did not block the translocation. In contrast to the epsilon subtype, ATP-induced translocation of PKC gamma was observed in the presence of both the PLA(2) inhibitor and the DAG antagonist. However, it is noteworthy that re-translocation of PKC gamma was hastened by the PLA(2) inhibitor. Furthermore products of PLA(2), such as lysophospholipids and fatty acids, induced the translocation of PKC gamma and epsilon in a dose dependent manner, but not delta. These results indicate that, in addition to PLC and DAG, PLA(2) and its products are involved in the purinergic stimuli-induced translocation of PKC epsilon and gamma in CHO-K1 cells. Each subtype of PKC in CHO-K1 cell is individually activated in response to a purinergic stimulation. PMID- 10725218 TI - Upregulation of the secretory pathway in cysteine protease inhibitor-resistant Trypanosoma cruzi. AB - A novel chemotherapy in development for Chagas' disease targets cruzain, the major cysteine protease of Trypanosoma cruzi. Peptidomimetic inhibitors disrupt the intracellular cycle of the parasite and rescue animals from a lethal infection. Inhibitor killing of parasites results from interruption of autocatalytic cruzain processing and transport to lysosomes, and massive accumulation of precursor protein in the Golgi complex. To further understand the mechanisms of protease processing and transport in this primitive eukaryote, and uncover potential mechanisms for resistance to these drugs, we generated cysteine protease inhibitor (CPI)-resistant epimastigotes in vitro and investigated the mechanisms involved at the biochemical and structural levels. Resistance to 20 fold the lethal CPI concentration, achieved after a year of gradual drug increase, was accompanied by a modest decrease in growth rate. A marked increase in the number of vesicles trafficking from the Golgi complex to the flagellar pocket occurs in resistant cells. No mature protease reaches lysosomes though accumulation of endocytosed gold particles in lysosomes appears to be normal. Higher molecular mass cruzain species, consistent with complexes of cruzain precursors and inhibitor, are secreted by CPI-resistant parasites into the culture supernatant. Release of these cruzain precursors may be facilitated by an enhanced acidification of trans-Golgi cisternae in resistant parasites. The pH within Golgi cisternae is higher in control epimastigotes and most mature cruzain is lysosomal. Cruzain activity is negligible in CPI-resistant epimastigote extracts compared to the parental clone. Activity is restored following withdrawal of the inhibitor. No cross-resistance to the therapeutic drugs nifurtimox and benznidazole occurred and, conversely, parasites resistant to these drugs were sensitive to CPI. Protease inhibitors are thus potential therapeutical alternatives in cases of nifurtimox/benznidazole resistance. Cumulatively, these results suggest that CPI-resistance induces upregulation of Golgi complex function and post-Golgi secretory pathway, and release of precursors before the enzyme reaches its site of biologic activity. PMID- 10725219 TI - Differential expression and cellular distribution of centrin isoforms during human ciliated cell differentiation in vitro. AB - Centrin protein is an ubiquitously expressed cytoskeletal component and is a member of the EF-hand superfamily of calcium-binding proteins. It was first discovered in the flagellar apparatus of unicellular green algae where it is involved in contraction of Ca(2+)-sensitive structures. Centrin protein is associated with centrosome-related structures such as spindle pole body in yeast, and centriole/basal bodies in flagellar and ciliated cells. Three centrin genes have been cloned in human cells. In this work, we have performed a comparative biochemical and functional analysis of centrin isoforms using a primary culture of human nasal epithelial cells which provides an efficient way to obtain a complete ciliated cell differentiation process. RT-PCR experiments show that the expression of the three human centrin genes increases during cell differentiation, and that only centrin 2 and 3 are expressed during cell proliferation. Using polyclonal antibodies raised against recombinant human centrin 2 and 3, we show a specific pattern of protein expression. Ultrastructural immunolocalization suggests that centrin proteins are involved in the early process of centriole assembly, as they are concentrated within the precursor structures of centriole/basal bodies. It also shows a differential localisation of centrin proteins in mature centriole/basal bodies, suggesting different functions for centrins 1/2 and centrin 3. This is also supported by functional analyses showing that centrin 1 and/or centrin 2 are involved in ciliary beating. PMID- 10725220 TI - Selective permeability of different connexin channels to the second messenger inositol 1,4,5-trisphosphate. AB - Intercellular propagation of signals through connexin32-containing gap junctions is of major importance in physiological processes like nerve activity-dependent glucose mobilization in liver parenchymal cells and enzyme secretion from pancreatic acinar cells. In these cells, as in other organs, more than one type of connexin is expressed. We hypothesized that different permeabilities towards second messenger molecules could be one of the reasons for connexin diversity. In order to investigate this, we analyzed transmission of inositol 1,4,5 trisphosphate-mediated calcium waves in FURA-2-loaded monolayers of human HeLa cells expressing murine connexin26, -32 or -43. Gap junction-mediated cell coupling in different connexin-transfected HeLa cells was standardized by measuring the spreading of microinjected Mn(2+) that led to local quenching of FURA-2 fluorescence. Microinjection of inositol 1,4,5-trisphosphate into confluently growing HeLa connexin32 transfectants induced propagation of a Ca(2+) wave from the injected cell to neighboring cells that was at least three- to fourfold more efficient than in HeLa Cx26 cells and about 2.5-fold more efficient than in HeLa Cx43 transfectants. Our results support the notion that diffusion of inositol 1,4,5-trisphosphate through connexin32-containing gap junctions is essential for the optimal physiological response, for example by recruiting liver parenchymal cells that contain subthreshold levels of this short lived second messenger. PMID- 10725221 TI - Human tau filaments induce microtubule and synapse loss in an in vivo model of neurofibrillary degenerative disease. AB - The intracellular accumulation of tau protein and its aggregation into filamentous deposits is the intracellular hallmark of neurofibrillary degenerative diseases such as Alzheimer's Disease and familial tauopathies in which tau is now thought to play a critical pathogenic role. Until very recently, the lack of a cellular model in which human tau filaments can be experimentally generated has prevented direct investigation of the causes and consequences of tau filament formation in vivo. In this study, we show that human tau filaments formed in lamprey central neurons (ABCs) that chronically overexpress human tau resemble the 'straight filaments' seen in Alzheimer's Disease and other neurofibrillary conditions, and are distinguishable from neurofilaments by their ultrastructure, distribution and intracellular behavior. We also show that tau filament formation in ABCs is associated with a distinctive pattern of dendritic degeneration that closely resembles the cytopathology of human neurofibrillary degenerative disease. This pattern includes localized cytoskeletal disruption and aggregation of membranous organelles, distal dendritic beading, and the progressive loss of dendritic microtubules and synapses. These results suggest that tau filament formation may be responsible for many key cytopathological features of neurofibrillary degeneration, possibly via the loss of microtubule based intracellular transport. PMID- 10725222 TI - Targeting of synaptotagmin to neurite terminals in neuronally differentiated PC12 cells. AB - We have investigated structural elements that determine the accumulation of synaptotagmin, a major synaptic vesicle protein, in neurite terminals of neuronally differentiated neuroendocrine pheochromocytoma PC12 cells. We performed extensive deletion and point mutagenesis of rat synaptotagmin II, expressed mutant proteins in PC12 cells differentiated by nerve growth factor (NGF) and monitored their intracellular distribution by immunofluorescence. We found a structural element located at the carboxy-terminal domain of the synaptotagmin molecule, which is necessary for its accumulation at the terminal. Using alanine-scanning mutagenesis, we have identified two amino acids in this element, tryptophan W405 and leucine L408, that are critical for correct targeting of synaptotagmin II to neurite terminals. Changing either one of them to alanine prevents the accumulation of the protein at the terminals. These amino acids are evolutionarily conserved throughout the entire synaptotagmin family and also among synaptotagmin-related proteins, suggesting that different synaptotagmins may have similar mechanisms of targeting to neuronal cell terminals. PMID- 10725223 TI - A functional knock-out of titin results in defective myofibril assembly. AB - Titin, also called connectin, is a giant muscle protein that spans the distance from the sarcomeric Z-disc to the M-band. Titin is thought to direct the assembly of sarcomeres and to maintain sarcomeric integrity by interacting with numerous sarcomeric proteins and providing a mechanical linkage. Since severe defects of such an important molecule are likely to result in embryonic lethality, a cell culture model should offer the best practicable tool to probe the cellular functions of titin. The myofibroblast cell line BHK-21/C13 was described to assemble myofibrils in culture. We have now characterized the sub-line BHK-21-Bi, which bears a small deletion within the titin gene. RNA analysis revealed that in this mutant cell line only a small internal portion of the titin mRNA is deleted. However, western blots, immunofluorescence microscopy and immunoprecipitation experiments showed that only the N-terminal, approx. 100 kDa central Z-disc portion of the 3 MDa titin protein is expressed, due to the homozygous deletion in the gene. Most importantly, in BHK-21-Bi cells the formation of thick myosin filaments and the assembly of myofibrils are impaired, although sarcomeric proteins are expressed. Lack of thick filament formation and of ordered actin myosin arrays was confirmed by electron microscopy. Myogenisation induced by transfection with MyoD yielded myofibrils only in myotubes formed from wild type and not from mutant cells, ruling out that a principal failure in myogenic commitment of the BHK-21-Bi cells might cause the observed effects. These experiments provide the first direct evidence for the crucial role of titin in both thick filament formation as a molecular ruler and in the coordination of myofibrillogenesis. PMID- 10725224 TI - Accumulation of profilin II at the surface of Listeria is concomitant with the onset of motility and correlates with bacterial speed. AB - The spatial and temporal activity of the actin cytoskeleton is precisely regulated during cell motility by several microfilament-associated proteins of which profilin plays an essential role. We have analysed the distribution of green fluorescent protein (GFP)-tagged profilins in cultured and in Listeria infected cells. Among the different GFP-profilin fusion proteins studied, only the construct in which the GFP moiety was fused to the carboxy terminus of profilin II (profilin II-GFP) was recruited by intracellular Listeria. The in vitro ligand-binding properties of this construct, e.g. the binding to monomeric actin, poly-L-proline and phosphatidylinositol 4,5-bisphosphate (PIP2), were unaffected by GFP. Profilin II-GFP co-localised with vinculin and Mena to the focal adhesions in REF-52 fibroblasts and was distributed as a thin line at the front of protruding lamellipodia in B16-F1 mouse melanoma cells. In Listeria infected cells, profilin II-GFP was recruited, in an asymmetric fashion, to the surface of Listeria at the onset of motility whereas it was not detectable on non motile bacteria. In contrast to the vasodilator-stimulated phosphoprotein (VASP), profilin II-GFP localised at the bacterial surface only on motile Listeria. Moreover, the fluorescence intensity of profilin II-GFP directly correlated with the speed of the bacteria. Thus, the use of GFP-tagged profilin II provides new insights into the role of profilins in cellular motility. PMID- 10725225 TI - Functional overlap of the dictyostelium RasG, RasD and RasB proteins. AB - Disruption of the rasG gene in Dictyostelium discoideum results in several distinct phenotypes: a defect in cytokinesis, reduced motility and reduced growth. Reintroduction of the rasG gene restores all of the properties of the rasG(-) cells to those of the wild type. To determine whether the defects are due to impaired interactions with a single or multiple downstream effectors, we tested the ability of the highly related but non identical Dictyostelium ras genes, rasD and rasB, to rescue the defects. Introduction of the rasD gene under the control of the rasG promoter into rasG null (rasG(-)) cells corrected all phenotypes except the motility defect, suggesting that motility is regulated by a RasG mediated pathway that is different to those regulating growth or cytokinesis. Western blot analysis of RasD protein levels revealed that vegetative rasG(- )cells contained considerably more protein than the parental AX 3 cells, suggesting that RasD protein levels are negatively regulated in vegetative cells by RasG. The level of RasD was enhanced when the rasD gene was introduced under the control of the rasG promoter, and this increase in protein is presumably responsible for the reversal of the growth and cytokinesis defects of the rasG(- )cells. Thus, RasD protein levels are controlled by the level of RasG, but not by the level of RasD. Introduction of the rasB gene under the control of the rasG promoter into rasG(-) cells produced a complex phenotype. The transformants were extremely small and mononucleate and exhibited enhanced motility. However, the growth of these cells was considerably slower than the growth of the rasG(-) cells, suggesting the possibility that high levels of RasB inhibit an essential process. This was confirmed by expressing rasB in wild-type cells; the resulting transformants exhibited severely impaired growth. When RasB protein levels were determined by western blot analysis, it was found that levels were higher in the rasG(- )cells than they were in the wild-type parental, suggesting that RasG also negatively regulates rasB expression in vegetative cells. Overexpression of rasB in the rasG(- )cells also reduced the level of RasD protein. In view of the fact that alternate Ras proteins correct some, but not all, of the defects exhibited by the rasG(-) cells, we propose that RasG interacts with more than one downstream effector. In addition, it is clear that the levels of the various Ras proteins are tightly regulated in vegetative cells and that overexpression can be deleterious. PMID- 10725226 TI - Regulation of cytokinesis by the Elm1 protein kinase in Saccharomyces cerevisiae. AB - A Saccharomyces cerevisiae mutant unable to grow in a cdc28-1N background was isolated and shown to be affected in the ELM1 gene. Elm1 is a protein kinase, thought to be a negative regulator of pseudo-hyphal growth. We show that Cdc11, one of the septins, is delocalised in the mutant, indicating that septin localisation is partly controlled by Elm1. Moreover, we show that cytokinesis is delayed in an elm1delta mutant. Elm1 levels peak at the end of the cell cycle and Elm1 is localised at the bud neck in a septin-dependent fashion from bud emergence until the completion of anaphase, at about the time of cell division. Genetic and biochemical evidence suggest that Elm1 and the three other septin localised protein kinases, Hsl1, Gin4 and Kcc4, work in parallel pathways to regulate septin behaviour and cytokinesis. In addition, the elm1delta;) morphological defects can be suppressed by deletion of the SWE1 gene, but not the cytokinesis defect nor the septin mislocalisation. Our results indicate that cytokinesis in budding yeast is regulated by Elm1. PMID- 10725227 TI - A fission yeast general translation factor reveals links between protein synthesis and cell cycle controls. AB - In two independent screens we isolated fission yeast mutations with phenotypes suggesting defects in B-cyclin function or expression. These mutations define a single gene which we call ded1. We show that ded1 encodes a general translation factor that is related in sequence and function to RNA helicases required for translation in other species. Levels of the B-cyclins Cig2 and Cdc13 are dramatically reduced upon inactivation of Ded1, and this reduction is independent of degradation by the anaphase promoting complex. When a ded1 mutant is grown under semi-restrictive conditions, the translation of Cig2 (and to a lesser extent Cdc13), is impaired relative to other proteins. We show that B-cyclin translation is specifically inhibited upon nitrogen starvation of wild-type cells, when B-cyclin/Cdc2 inactivation is a prerequisite for G(1) arrest and subsequent mating. Our data suggest that translational inhibition of B-cyclin expression represents a third mechanism, in addition to cyclin degradation and Rum1 inhibition, that contributes to Cdc2 inactivation as cells exit from the mitotic cell cycle and prepare for meiosis. PMID- 10725228 TI - Contributions of the extracellular and cytoplasmic domains of platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31) in regulating cell-cell localization. AB - PECAM-1/CD31, a vascular cell adhesion/signaling molecule that has been implicated in a number of vascular functions (including angiogenesis and the transmigration of leukocytes through endothelium) is highly enriched at the cell cell borders of adjacent endothelial cells. To identify the mechanisms responsible for this localization, a series of PECAM-1 mutants and chimeric PECAM 1 molecules were transfected into non-PECAM-expressing cells and the ability of the constructs to move to cell-cell borders of adjacent cells was determined using immunohistochemistry and confocal microscopy. Although neither the extracellular domain, by itself, nor the cytoplasmic domain, by itself, was sufficient to direct cell-cell localization, the combination of the extracellular and transmembrane domains with a small group of highly charged amino acids in a membrane proximal region of the cytoplasmic domain was sufficient to direct efficient localization of the molecule to cell-cell borders. Importantly, only constructs that supported PECAM-1 mediated adhesion localized to cell-cell borders. Our data are consistent with a 'diffusion trapping' model in which movement of PECAM-1 in the cell membrane occurs relatively freely until the 'stablized' extracellular domain of the molecule encounters its ligand on an adjacent cell. When this occurs, the complex is 'captured' at the cell-cell interface leading to localization at cell-cell borders. PMID- 10725229 TI - Nup2p is located on the nuclear side of the nuclear pore complex and coordinates Srp1p/importin-alpha export. AB - Proteins bearing canonical nuclear localization sequences are imported into the nucleus by the importin/karyopherin-alpha/beta heterodimer. Recycling of the importin-alpha subunit to the cytoplasm requires the action of Cas, a member of the importin-beta superfamily. In the yeast Saccharomyces ceresivisiae, the essential gene CSE1 encodes a Cas homologue that exports the yeast importin-alpha protein Srp1p/Kap60p from the nucleus. In this report, we describe a role for the FXFG nucleoporin Nup2p, and possibly the related Nup1p, in the Cse1p-mediated nuclear export pathway. Yeast cells lacking Nup2p or containing a particular temperature-sensitive mutation in NUP1 accumulate Srp1p in the nucleus. Similarly, Cse1p is displaced from the nuclear rim to the nuclear interior in deltanup2 cells. We do not observe any biochemical interaction between Cse1p and Nup2p. Instead, we find that Nup2p binds directly to Srp1p. We have localized Nup2p to the interior face of the nuclear pore complex, and have shown that its N terminus is sufficient for targeting Nup2p to the pore, as well as for binding to Srp1p. Taken together, these data suggest that Nup2p is an important NPC docking site in the Srp1p export pathway. PMID- 10725230 TI - ARVCF localizes to the nucleus and adherens junction and is mutually exclusive with p120(ctn) in E-cadherin complexes. AB - ARVCF is a novel Armadillo repeat domain protein that is closely related to the catenin p120(ctn). Using new ARVCF monoclonal antibodies, we have found that ARVCF associates with E-cadherin and competes with p120 for interaction with the E-cadherin juxtamembrane domain. ARVCF also localized to the nucleus in some cell types, however, and was significantly more nucleophilic than p120. Surprisingly, despite apparently ubiquitous expression, ARVCF was at least tenfold less abundant than p120 in a wide variety of cell types, and was difficult to detect by immunofluorescence unless overexpressed. Consequently, it is not likely to be abundant enough in adult tissues to functionally compete with p120. ARVCF also completely lacked the ability to induce the cell-branching phenotype associated with overexpression of p120. Expression of ARVCF/p120 chimeras confirmed previous results indicating that the branching activity of p120 maps to its Armadillo repeat domain. Surprisingly, the preferential localization of ARVCF to the nucleus required sequences in the amino-terminal end of ARVCF, suggesting that the sequences directing nuclear translocation of ARVCF are distinct from the predicted bipartite nuclear localization signal located between repeats 6 and 7. The dual localization of ARVCF to junctions and to nuclei suggests activities in different cellular compartments, as is the case for several other Armadillo repeat proteins including beta-catenin, p120 and the plakophilins. PMID- 10725231 TI - A novel RGD-independent fibronectin assembly pathway initiated by alpha4beta1 integrin binding to the alternatively spliced V region. AB - Fibronectin (FN) matrix assembly is a multi-step process that involves binding to integrin receptors, FN-FN interactions and connections to the actin cytoskeleton. Ultimately, FN is converted into stable matrix fibrils that are detergent insoluble. RGD-binding integrins such as alpha5beta1 play a major role in the assembly of fibrillar FN. Here we show that alpha4beta1 binding to the alternatively spliced V (IIICS) region of FN initiates an alternative assembly pathway. Activation of alpha4beta1 with exogenous agents such as Mn(2+) or a beta1-stimulatory antibody TS2/16 was sufficient to induce initiation of FN fibrillogenesis by Ramos B lymphoma cells and by CHO(B2)alpha4 cells. Using recombinant FNs lacking specific sequences, we show that assembly is independent of the RGD sequence but requires the V25/CS-1 segment. Previously, we have characterized an activated recombinant FN (FN III(1-7)) that rapidly forms detergent-insoluble multimers upon binding to alpha5beta1 integrin. Alpha4beta1 also formed FNdeltaIII(1-7) multimers without the aid of exogenous stimulants, suggesting that an activated form of FN can override the need for activation of the integrin. In contrast to assembly by alpha5beta1, actin filaments remained largely cortical and no change in cell growth rate was observed with alpha4beta1 mediated assembly. These results show that binding sites on FN other than the RGD sequence/synergy site and distant from the cell binding domain can promote FN assembly. Thus, there appear to be multiple, integrin-specific mechanisms for assembly of FN matrix. PMID- 10725232 TI - Retinoid signaling is essential for patterning the endoderm of the third and fourth pharyngeal arches. AB - The requirement of retinoic acid (RA) in the initial formation of the pharyngeal arches was investigated by treating headfold-stage mouse embryos with a pan-RAR antagonist in vitro and in vivo. This results in a complete absence of mesenchyme, arteries, nerves and epibranchial placodes of the 3rd and 4th pharyngeal arches, complete agenesis of the 3rd and 4th pouches and consistent lack of the 6th arch artery. Mesodermally derived endothelial cells are absent from the 3rd and 4th pharyngeal arch region and the distribution domain of EphA2 transcripts in mesodermal cells is shifted caudally. In situ hybridization with CRABPI, kreisler and EphA4 probes and the pattern of expression of a Wnt1-lacZ transgene show that neural crest cells (NCC) normally destined to the 3rd and 4th arches migrate ectopically. Most interestingly, the appearance of the 3rd and 4th arches is prevented by the antagonist only during a very narrow window of time, which does not correspond to the period of post-otic NCC migration. Both the timing of appearance and the nature of the defects in RAR antagonist-treated embryos indicate that migrating NCC and mesodermal cells destined to the caudal pharyngeal arches do not represent primary targets of RA action. Alterations in the endodermal expression pattern of Hoxa1, Hoxb1, Pax1, Pax9, Fgf3 and Fgf8 in response to the antagonist-induced block in RA signal transduction demonstrate for the first time that RA signaling is indispensable for the specification of the pharyngeal endoderm and suggest that this signaling is necessary to provide a permissive environment locally for the migration of NCC and mesodermal cells. Our study also indicates that the formation of the 2nd pharyngeal arch and that of the 3rd and 4th pharyngeal arches probably involve distinct RA-dependent developmental processes. PMID- 10725233 TI - Early mouse endoderm is patterned by soluble factors from adjacent germ layers. AB - Endoderm that forms the respiratory and digestive tracts is a sheet of approximately 500-1000 cells around the distal cup of an E7.5 mouse embryo. Within 2 days, endoderm folds into a primitive gut tube from which numerous organs will bud. To characterize the signals involved in the developmental specification of this early endoderm, we have employed an in vitro assay using germ layer explants and show that adjacent germ layers provide soluble, temporally specific signals that induce organ-specific gene expression in endoderm. Furthermore, we show that FGF4 expressed in primitive streak-mesoderm can induce the differentiation of endoderm in a concentration-dependent manner. We conclude that the differentiation of gastrulation-stage endoderm is directed by adjacent mesoderm and ectoderm, one of the earliest reported patterning events in formation of the vertebrate gut tube. PMID- 10725234 TI - Persistence of Hunchback in the terminal region of the Drosophila blastoderm embryo impairs anterior development. AB - Anterior terminal development is controlled by several zygotic genes that are positively regulated at the anterior pole of Drosophila blastoderm embryos by the anterior (bicoid) and the terminal (torso) maternal determinants. Most Bicoid target genes, however, are first expressed at syncitial blastoderm as anterior caps, which retract from the anterior pole upon activation of Torso. To better understand the interaction between Bicoid and Torso, a derivative of the Gal4/UAS system was used to selectively express the best characterised Bicoid target gene, hunchback, at the anterior pole when its expression should be repressed by Torso. Persistence of hunchback at the pole mimics most of the torso phenotype and leads to repression at early stages of a labral (cap'n'collar) and two foregut (wingless and hedgehog) determinants that are positively controlled by bicoid and torso. These results uncovered an antagonism between hunchback and bicoid at the anterior pole, whereas the two genes are known to act in concert for most anterior segmented development. They suggest that the repression of hunchback by torso is required to prevent this antagonism and to promote anterior terminal development, depending mostly on bicoid activity. PMID- 10725235 TI - Stepwise development of thymic microenvironments in vivo is regulated by thymocyte subsets. AB - T-cell development is under the tight control of thymic microenvironments. Conversely, the integrity of thymic microenvironments depends on the physical presence of developing thymocytes, a phenomenon designated as 'thymic crosstalk'. We now show, using three types of immunodeficient mice, i.e. CD3(epsilon) transgenic mice, RAG(null) mice and RAG(null)-bone-marrow-transplanted CD3(epsilon) transgenic mice, that the control point in lymphoid development where triple negative (CD3(-),CD4(-),CD8(-)) thymocytes progress from CD44(+)CD25(-) towards CD44(-)CD25(+), influences the development of epithelial cells, critically inducing the extra, third dimension in the organization of the epithelial cells in the cortex. This tertiary configuration of the thymic epithelium is a typical feature for the thymus, enabling lymphostromal interaction during T-cell development. Crosstalk signals at this control point also induce the formation of thymic nurse cells. Moreover, our data indicate that establishment of a thymic cortex is a prerequisite for the development of the thymic medulla. Thus, differentiating thymocytes regulate the morphogenesis of thymic microenvironments in a stepwise fashion. PMID- 10725236 TI - Mouse Gli1 mutants are viable but have defects in SHH signaling in combination with a Gli2 mutation. AB - The secreted factor Sonic hedgehog (SHH) is both required for and sufficient to induce multiple developmental processes, including ventralization of the CNS, branching morphogenesis of the lungs and anteroposterior patterning of the limbs. Based on analogy to the Drosophila Hh pathway, the multiple GLI transcription factors in vertebrates are likely to both transduce SHH signaling and repress Shh transcription. In order to discriminate between overlapping versus unique requirements for the three Gli genes in mice, we have produced a Gli1 mutant and analyzed the phenotypes of Gli1/Gli2 and Gli1/3 double mutants. Gli3(xt) mutants have polydactyly and dorsal CNS defects associated with ectopic Shh expression, indicating GLI3 plays a role in repressing Shh. In contrast, Gli2 mutants have five digits, but lack a floorplate, indicating that it is required to transduce SHH signaling in some tissues. Remarkably, mice homozygous for a Gli1(zfd )mutation that deletes the exons encoding the DNA-binding domain are viable and appear normal. Transgenic mice expressing a GLI1 protein lacking the zinc fingers can not induce SHH targets in the dorsal brain, indicating that the Gli1(zfd )allele contains a hypomorphic or null mutation. Interestingly, Gli1(zfd/zfd);Gli2(zfd/+), but not Gli1(zfd/zfd);Gli3(zfd/+) double mutants have a severe phenotype; most Gli1(zfd/zfd);Gli2(zfd/+) mice die soon after birth and all have multiple defects including a variable loss of ventral spinal cord cells and smaller lungs that are similar to, but less extreme than, Gli2(zfd/zfd) mutants. Gli1/Gli2 double homozygous mutants have more extreme CNS and lung defects than Gli1(zfd/zfd);Gli2(zfd/+) mutants, however, in contrast to Shh mutants, ventrolateral neurons develop in the CNS and the limbs have 5 digits with an extra postaxial nubbin. These studies demonstrate that the zinc-finger DNA-binding domain of GLI1 protein is not required for SHH signaling in mouse. Furthermore, Gli1 and Gli2, but not Gli1 and Gli3, have extensive overlapping functions that are likely downstream of SHH signaling. PMID- 10725237 TI - Fate of the mammalian cardiac neural crest. AB - A subpopulation of neural crest termed the cardiac neural crest is required in avian embryos to initiate reorganization of the outflow tract of the developing cardiovascular system. In mammalian embryos, it has not been previously experimentally possible to study the long-term fate of this population, although there is strong inference that a similar population exists and is perturbed in a number of genetic and teratogenic contexts. We have employed a two-component genetic system based on Cre/lox recombination to label indelibly the entire mouse neural crest population at the time of its formation, and to detect it at any time thereafter. Labeled cells are detected throughout gestation and in postnatal stages in major tissues that are known or predicted to be derived from neural crest. Labeling is highly specific and highly efficient. In the region of the heart, neural-crest-derived cells surround the pharyngeal arch arteries from the time of their formation and undergo an altered distribution coincident with the reorganization of these vessels. Labeled cells populate the aorticopulmonary septum and conotruncal cushions prior to and during overt septation of the outflow tract, and surround the thymus and thyroid as these organs form. Neural crest-derived mesenchymal cells are abundantly distributed in midgestation (E9.5 12.5), and adult derivatives of the third, fourth and sixth pharyngeal arch arteries retain a substantial contribution of labeled cells. However, the population of neural-crest-derived cells that infiltrates the conotruncus and which surrounds the noncardiac pharyngeal organs is either overgrown or selectively eliminated as development proceeds, resulting for these tissues in a modest to marginal contribution in late fetal and postnatal life. PMID- 10725238 TI - Leg development in flies versus grasshoppers: differences in dpp expression do not lead to differences in the expression of downstream components of the leg patterning pathway. AB - All insect legs are structurally similar, characterized by five primary segments. However, this final form is achieved in different ways. Primitively, the legs developed as direct outgrowths of the body wall, a condition retained in most insect species. In some groups, including the lineage containing the genus Drosophila, legs develop indirectly from imaginal discs. Our understanding of the molecular mechanisms regulating leg development is based largely on analysis of this derived mode of leg development in the species D. melanogaster. The current model for Drosophila leg development is divided into two phases, embryonic allocation and imaginal disc patterning, which are distinguished by interactions among the genes wingless (wg), decapentaplegic (dpp) and distalless (dll). In the allocation phase, dll is activated by wg but repressed by dpp. During imaginal disc patterning, dpp and wg cooperatively activate dll and also indirectly inhibit the nuclear localization of Extradenticle (Exd), which divide the leg into distal and proximal domains. In the grasshopper Schistocerca americana, the early expression pattern of dpp differs radically from the Drosophila pattern, suggesting that the genetic interactions that allocate the leg differ between the two species. Despite early differences in dpp expression, wg, Dll and Exd are expressed in similar patterns throughout the development of grasshopper and fly legs, suggesting that some aspects of proximodistal (P/D) patterning are evolutionarily conserved. We also detect differences in later dpp expression, which suggests that dpp likely plays a role in limb segmentation in Schistocerca, but not in Drosophila. The divergence in dpp expression is surprising given that all other comparative data on gene expression during insect leg development indicate that the molecular pathways regulating this process are conserved. However, it is consistent with the early divergence in developmental mode between fly and grasshopper limbs. PMID- 10725239 TI - Transdifferentiation of esophageal smooth to skeletal muscle is myogenic bHLH factor-dependent. AB - Previously, coexpression of smooth and skeletal differentiation markers, but not myogenic regulatory factors (MRFs), was observed from E16.5 mouse fetuses in a small percentage of diaphragm level esophageal muscle cells, suggesting that MRFs are not involved in the process of initiation of developmentally programmed transdifferentiation in the esophagus. To investigate smooth-to-skeletal esophageal muscle transition, we analyzed Myf5nlacZ knock-in mice, MyoD-lacZ and myogenin-lacZ transgenic embryos with a panel of the antibodies reactive with myogenic regulatory factors (MRFs) and smooth and skeletal muscle markers. We observed that lacZ-expressing myogenic precursors were not detected in the esophagus before E15.5, arguing against the hypothesis that muscle precursor cells populate the esophagus at an earlier stage of development. Rather, the expression of the MRFs initiated in smooth muscle cells in the upper esophagus of E15.5 mouse embryos and was immediately followed by the expression of skeletal muscle markers. Moreover, transdifferentiation was markedly delayed or absent only in the absence of Myf5, suggesting that appropriate initiation and progression of smooth-to-skeletal muscle transdifferentiation is Myf5-dependent. Accordingly, the esophagus of Myf5(-/-):MyoD(-/-)embryos completely failed to undergo skeletal myogenesis and consisted entirely of smooth muscle. Lastly, extensive proliferation of muscularis precursor cells, without programmed cell death, occurred concomitantly with esophageal smooth-to-skeletal muscle transdifferentiation. Taken together, these results indicate that transdifferentiation is the fate of all smooth muscle cells in the upper esophagus and is normally initiated by Myf5. PMID- 10725240 TI - In vivo regulation of cell death by embryonic (pro)insulin and the insulin receptor during early retinal neurogenesis. AB - Programmed cell death is an established developmental process in the nervous system. Whereas the regulation and the developmental role of neuronal cell death have been widely demonstrated, the relevance of cell death during early neurogenesis, the cells affected and the identity of regulatory local growth factors remain poorly characterized. We have previously described specific in vivo patterns of apoptosis during early retinal neurogenesis, and that exogenous insulin acts as survival factor (Diaz, B., Pimentel, B., De Pablo, F. and de la Rosa, E. J. (1999) Eur. J. Neurosci. 11, 1624-1632). Proinsulin mRNA was found to be expressed broadly in the early embryonic chick retina, and decreased later between days 6 and 8 of embryonic development, when there was increased expression of insulin-like growth factor I mRNA, absent or very scarce at earlier stages. Consequently, we studied whether proinsulin and/or insulin ((pro)insulin) action in prevention of cell death has physiological relevance during early neural development. In ovo treatment at day 2 of embryonic development with specific antibodies against (pro)insulin or the insulin receptor induced apoptosis in the neuroretina. The distribution of apoptotic cells two days after the blockade was similar to naturally occurring cell death, as visualized by TdT mediated dUTP nick end labeling. The apoptosis induced by the insulin receptor blockade preferentially affected to the Islet1/2 positive cells, that is, the differentiated retinal ganglion cells. In parallel, the insulin survival effect on cultured retinas correlated with the activation of Akt to a greater extent than with the activation of MAP kinase. These results suggest that the physiological cell death occurring in early stages of retinal development is regulated by locally produced (pro)insulin through the activation of the Akt survival pathway. PMID- 10725241 TI - A S/M DNA replication checkpoint prevents nuclear and cytoplasmic events of cell division including centrosomal axis alignment and inhibits activation of cyclin dependent kinase-like proteins in fucoid zygotes. AB - S/M checkpoints prevent various aspects of cell division when DNA has not been replicated. Such checkpoints are stringent in yeast and animal somatic cells but are usually partial or not present in animal embryos. Because little is known about S/M checkpoints in plant cells and embryos, we have investigated the effect of aphidicolin, a specific inhibitor of DNA polymerases (alpha) and (delta), on cell division and morphogenesis in Fucus and Pelvetia zygotes. Both DNA replication and cell division were inhibited by aphidicolin, indicating the presence, in fucoid zygotes, of a S/M checkpoint. This checkpoint prevents chromatin condensation, spindle formation, centrosomal alignment with the growth axis and cytokinesis but has no effect on germination or rhizoid elongation. This S/M checkpoint also prevents tyrosine dephosphorylation of cyclin-dependent kinase-like proteins at the onset of mitosis. The kinase activity is restored in extracts upon incubation with cdc25A phosphatase. When added in S phase, olomoucine, a specific inhibitor of cyclin-dependent kinases, has similar effects as aphidicolin on cell division although alignment of the centrosomal axis still occurs. We propose a model involving the inactivation of CDK-like proteins to account for the S/M DNA replication checkpoint in fucoid zygotes and embryos. PMID- 10725242 TI - POLTERGEIST functions to regulate meristem development downstream of the CLAVATA loci. AB - Mutations at the CLAVATA loci (CLV1, CLV2 and CLV3) result in the accumulation of undifferentiated cells at the shoot and floral meristems. We have isolated three mutant alleles of a novel locus, POLTERGEIST (POL), as suppressors of clv1, clv2 and clv3 phenotypes. All pol mutants were nearly indistinguishable from wild-type plants; however, pol mutations provided recessive, partial suppression of meristem defects in strong clv1 and clv3 mutants, and nearly complete suppression of weak clv1 mutants. pol mutations partially suppressed clv2 floral and pedicel defects in a dominant fashion, and almost completely suppressed clv2 phenotypes in a recessive manner. These observations, along with dominant interactions observed between the pol and wuschel (wus) mutations, indicate that POL functions as a critical regulator of meristem development downstream of the CLV loci and redundantly with WUS. Consistent with this, pol mutations do not suppress clv3 phenotypes by altering CLV1 receptor activation. PMID- 10725243 TI - Fate of the mammalian cranial neural crest during tooth and mandibular morphogenesis. AB - Neural crest cells are multipotential stem cells that contribute extensively to vertebrate development and give rise to various cell and tissue types. Determination of the fate of mammalian neural crest has been inhibited by the lack of appropriate markers. Here, we make use of a two-component genetic system for indelibly marking the progeny of the cranial neural crest during tooth and mandible development. In the first mouse line, Cre recombinase is expressed under the control of the Wnt1 promoter as a transgene. Significantly, Wnt1 transgene expression is limited to the migrating neural crest cells that are derived from the dorsal CNS. The second mouse line, the ROSA26 conditional reporter (R26R), serves as a substrate for the Cre-mediated recombination. Using this two component genetic system, we have systematically followed the migration and differentiation of the cranial neural crest (CNC) cells from E9.5 to 6 weeks after birth. Our results demonstrate, for the first time, that CNC cells contribute to the formation of condensed dental mesenchyme, dental papilla, odontoblasts, dentine matrix, pulp, cementum, periodontal ligaments, chondrocytes in Meckel's cartilage, mandible, the articulating disc of temporomandibular joint and branchial arch nerve ganglia. More importantly, there is a dynamic distribution of CNC- and non-CNC-derived cells during tooth and mandibular morphogenesis. These results are a first step towards a comprehensive understanding of neural crest cell migration and differentiation during mammalian craniofacial development. Furthermore, this transgenic model also provides a new tool for cell lineage analysis and genetic manipulation of neural-crest-derived components in normal and abnormal embryogenesis. PMID- 10725244 TI - Drosophila atonal controls photoreceptor R8-specific properties and modulates both receptor tyrosine kinase and Hedgehog signalling. AB - During Drosophila eye development, the proneural gene atonal specifies founding R8 photoreceptors of individual ommatidia, evenly spaced relative to one another in a pattern that prefigures ommatidial organisation in the mature compound eye. Beyond providing neural competence, however, it has remained unclear to what extent atonal controls specific R8 properties. We show here that reduced Atonal function gives rise to R8 photoreceptors that are functionally compromised: both recruitment and axon pathfinding defects are evident. Conversely, prolonged Atonal expression in R8 photoreceptors induces defects in inductive recruitment as a consequence of hyperactive EGFR signalling. Surprisingly, such prolonged expression also results in R8 pattern formation defects in a process associated with both Hedgehog and Receptor Tyrosine Kinase signalling. Our results strongly suggest that Atonal regulates signalling and other properties of R8 precursors. PMID- 10725245 TI - Zebrafish Mesp family genes, mesp-a and mesp-b are segmentally expressed in the presomitic mesoderm, and Mesp-b confers the anterior identity to the developing somites. AB - Segmentation of a vertebrate embryo begins with the subdivision of the paraxial mesoderm into somites through a not-well-understood process. Recent studies provided evidence that the Notch-Delta and the FGFR (fibroblast growth factor receptor) signalling pathways are required for segmentation. In addition, the Mesp family of bHLH transcription factors have been implicated in establishing a segmental prepattern in the presomitic mesoderm. In this study, we have characterized zebrafish mesp-a and mesp-b genes that are closely related to Mesp family genes in other vertebrates. During gastrulation, only mesp-a is expressed in the paraxial mesoderm at the blastoderm margin. During the segmentation period, both genes are segmentally expressed in one to three stripes in the anterior parts of somite primordia. In fused somites (fss) embryos, in which all early somite boundary formation is blocked, initial mesp-a expression at the gastrula stage remains intact, but the expression of mesp-a and mesp-b is not detected during the segmentation period. This suggests that these genes are downstream targets of fss at the segmentation stage. Comparison with her1 expression (Muller, M., von Weizsacker, E. and Campos-Ortega, J. A. (1996) Development 122, 2071-2078) suggests that, like her1, mesp genes are not expressed in primordia of the first several somites. Furthermore, we found that zebrafish her1 expression oscillates in the presomitic mesoderm. The her1 stripe, which first appears in the tailbud region, moves in a caudal to rostral direction, and it finally overlaps the most rostral mesp stripe. Thus, in the trunk region, both her1 and mesp transcripts are detected in every somite primordium posterior to the forming somites. Ectopic expression of Mesp-b in embryos causes a loss of the posterior identity within the somite primordium, leading to a segmentation defect. These embryos show a reduction in expression of the posterior genes, myoD and notch5, with uniform expression of the anterior genes, FGFR1, papc and notch6. These observations suggest that zebrafish mesp genes are involved in anteroposterior specification within the presumptive somites, by regulating the essential signalling pathways mediated by Notch-Delta and FGFR. PMID- 10725246 TI - Anteroposterior patterning is required within segments for somite boundary formation in developing zebrafish. AB - Somite formation involves the establishment of a segmental prepattern in the presomitic mesoderm, anteroposterior patterning of each segmental primordium and formation of boundaries between adjacent segments. How these events are co ordinated remains uncertain. In this study, analysis of expression of zebrafish mesp-a reveals that each segment acquires anteroposterior regionalisation when located in the anterior presomitic mesoderm. Thus anteroposterior patterning is occurring after the establishment of a segmental prepattern in the paraxial mesoderm and prior to somite boundary formation. Zebrafish fss(-), bea(-), des(-) and aei(-) embryos all fail to form somites, yet we demonstrate that a segmental prepattern is established in the presomitic mesoderm of all these mutants and hox gene expression shows that overall anteroposterior patterning of the mesoderm is also normal. However, analysis of various molecular markers reveals that anteroposterior regionalisation within each segment is disturbed in the mutants. In fss(-), there is a loss of anterior segment markers, such that all segments appear posteriorized, whereas in bea(-), des(-) and aei(-), anterior and posterior markers are expressed throughout each segment. Since somite formation is disrupted in these mutants, correct anteroposterior patterning within segments may be a prerequisite for somite boundary formation. In support of this hypothesis, we show that it is possible to rescue boundary formation in fss(-) through the ectopic expression of EphA4, an anterior segment marker, in the paraxial mesoderm. These observations indicate that a key consequence of the anteroposterior regionalisation of segments may be the induction of Eph and ephrin expression at segment interfaces and that Eph/ephrin signalling subsequently contributes to the formation of somite boundaries. PMID- 10725247 TI - Rasputin, the Drosophila homologue of the RasGAP SH3 binding protein, functions in ras- and Rho-mediated signaling. AB - The small GTPase Ras plays an important role in many cellular signaling processes. Ras activity is negatively regulated by GTPase activating proteins (GAPs). It has been proposed that RasGAP may also function as an effector of Ras activity. We have identified and characterized the Drosophila homologue of the RasGAP-binding protein G3BP encoded by rasputin (rin). rin mutants are viable and display defects in photoreceptor recruitment and ommatidial polarity in the eye. Mutations in rin/G3BP genetically interact with components of the Ras signaling pathway that function at the level of Ras and above, but not with Raf/MAPK pathway components. These interactions suggest that Rin is required as an effector in Ras signaling during eye development, supporting an effector role for RasGAP. The ommatidial polarity phenotypes of rin are similar to those of RhoA and the polarity genes, e.g. fz and dsh. Although rin/G3BP interacts genetically with RhoA, affecting both photoreceptor differentiation and polarity, it does not interact with the gain-of-function genotypes of fz and dsh. These data suggest that Rin is not a general component of polarity generation, but serves a function specific to Ras and RhoA signaling pathways. PMID- 10725248 TI - The function of the Drosophila fat facets deubiquitinating enzyme in limiting photoreceptor cell number is intimately associated with endocytosis. AB - Fat facets is a deubiquitinating enzyme required in a cell communication pathway that limits to eight the number of photoreceptor cells in each facet of the Drososphila compound eye. Genetic data support a model whereby Faf removes ubiquitin, a polypeptide tag for protein degradation, from a specific ubiquitinated protein thus preventing its degradation. Here, mutations in the liquid facets gene were identified as dominant enhancers of the fat facets mutant eye phenotype. The liquid facets locus encodes epsin, a vertebrate protein associated with the clathrin endocytosis complex. The results of genetic experiments reveal that fat facets and liquid facets facilitate endocytosis and function in common cells to generate an inhibitory signal that prevents ectopic photoreceptor determination. Moreover, it is demonstrated that the fat facets mutant phenotype is extraordinarily sensitive to the level of liquid facets expression. We propose that Liquid facets is a candidate for the critical substrate of Fat facets in the eye. PMID- 10725249 TI - Large-scale cDNA analysis reveals phased gene expression patterns during preimplantation mouse development. AB - Little is known about gene action in the preimplantation events that initiate mammalian development. Based on cDNA collections made from each stage from egg to blastocyst, 25438 3'-ESTs were derived, and represent 9718 genes, half of them novel. Thus, a considerable fraction of mammalian genes is dedicated to embryonic expression. This study reveals profound changes in gene expression that include the transient induction of transcripts at each stage. These results raise the possibility that development is driven by the action of a series of stage specific expressed genes. The new genes, 798 of them placed on the mouse genetic map, provide entry points for analyses of human and mouse developmental disorders. PMID- 10725250 TI - The branchial arches and HGF are growth-promoting and chemoattractant for cranial motor axons. AB - During development, cranial motor neurons extend their axons along distinct pathways into the periphery. For example, branchiomotor axons extend dorsally to leave the hindbrain via large dorsal exit points. They then grow in association with sensory ganglia, to their targets, the muscles of the branchial arches. We have investigated the possibility that pathway tissues might secrete diffusible chemorepellents or chemoattractants that guide cranial motor axons, using co cultures in collagen gels. We found that explants of dorsal neural tube or hindbrain roof plate chemorepelled cranial motor axons, while explants of cranial sensory ganglia were weakly chemoattractive. Explants of branchial arch mesenchyme were strongly growth-promoting and chemoattractive for cranial motor axons. Enhanced and oriented axon outgrowth was also elicited by beads loaded with Hepatocyte Growth Factor (HGF); antibodies to this protein largely blocked the outgrowth and orientation effects of the branchial arch on motor axons. HGF was expressed in the branchial arches, whilst Met, which encodes an HGF receptor, was expressed by subpopulations of cranial motor neurons. Mice with targetted disruptions of HGF or Met showed defects in the navigation of hypoglossal motor axons into the branchial region. Branchial arch tissue may thus act as a target derived factor that guides motor axons during development. This influence is likely to be mediated partly by Hepatocyte Growth Factor, although a component of branchial arch-mediated growth promotion and chemoattraction was not blocked by anti-HGF antibodies. PMID- 10725251 TI - Harmane produces hypotension following microinjection into the RVLM: possible role of I(1)-imidazoline receptors. AB - The beta-carboline, harmane (0.1 - 1.0 nmol) produces dose dependent hypotension when microinjected unilaterally into the rostral ventrolateral medulla (RVLM) of the anaesthetized rat. The potency of harmane on blood pressure is similar to that of the imidazoline, clonidine. The hypotensive effects of both clonidine and harmane are reversed by microinjection of the relatively I(1)-receptor selective antagonist efaroxan (20 nmol). These results are consistent with harmane acting at an I(1)-receptor in the RVLM. This is the first report of an endogenous ligand for I(1)-receptors that has central effects on blood pressure. PMID- 10725252 TI - Enhanced blood pressure sensitivity to DOCA-salt treatment in endothelin ET(B) receptor-deficient rats. AB - The role of endothelin ET(B) receptor-mediated action in the development and maintenance of deoxycorticosterone acetate (DOCA)-salt-induced hypertension was evaluated using the spotting-lethal (sl) rat which carries a naturally occurring deletion in the ET(B) receptor gene. Homozygous (sl/sl) rats treated with DOCA salt for 1 week exhibited an earlier onset of hypertension than heterozygous (sl/+) and wild-type (+/+) rats (systolic blood pressure, SBP; 156.7+/-3.4 versus 128.8+/-5.3 and 132.9+/-3.7 mmHg, respectively). Four weeks after the start of DOCA-salt treatment, homozygous rats developed marked hypertension, with a SBP of 206. 0+/-4.5 mmHg, compared with 184.8+/-10.7 mmHg in heterozygous and 164.3+/ 4.8 mmHg in wild-type rats. Cardiovascular hypertrophy and renal dysfunction observed after 4-weeks treatment with DOCA-salt were more severe in homozygous rats, compared to wild-type and heterozygous animals. These evidences support strongly the view that ET(B) receptor-mediated actions are a protective factor in the pathogenesis of DOCA-salt-induced hypertension. PMID- 10725253 TI - The P2Y(1) receptor closes the N-type Ca(2+) channel in neurones, with both adenosine triphosphates and diphosphates as potent agonists. AB - The rat P2Y(1) nucleotide receptor, the P2Y subtype abundant in the brain, was heterologously expressed in rat superior cervical ganglion neurones by micro injection of the receptor cRNA or cDNA. ADP inhibited the N-type Ca(2+) current by 64%, with EC(50) 8.2 nM, an action blocked competitively by the P2Y(1) receptor antagonist adenosine 3', 5'-bis-phosphate (K(i) 0.7 microM). 2 Methylthio-ADP inhibited the Ca(2+) current likewise, but with EC(50) 0.57 nM, giving the highest potency reported therewith for P2Y(1). Significantly, ATP and 2-methylthio-ATP were also agonists, the latter again at a very high potency (EC(50) 2.5 nM). We propose that this neuronal receptor, when present in brain at a high density as at synapses, can respond to very low concentrations of ATP and ADP as agonists, and that this would result in inhibition of N-type Ca(2+) currents and hence can reduce transmitter release or increase neuronal excitability. PMID- 10725254 TI - Factors influencing the low-frequency associated nicotinic ACh autoreceptor mediated depression of ACh release from rat motor nerve terminals. AB - 1. We have studied the inhibitory autoreceptor control of acetylcholine (ACh) release from rat motor nerve endings using an electrophysiological technique to quantify evoked ACh release in isolated hemidiaphragm muscles. Quantal ACh release (m) was estimated from the ratio of amplitudes of nerve evoked endplate currents and spontaneously occurring miniature endplate currents. 2. The nicotinic ACh receptor agonist cytisine (1 microM) decreased m at 0.5 Hz by around 20% but had no effect on m at 50 Hz. Changing the extracellular Ca(2+) concentration from 1.8 mM to either 0.45 or 3.6 mM abolished the effect of cytisine on m at 0.5 Hz. The nicotinic ACh receptor antagonist hexamethonium (200 microM) increased m at 0.5 Hz by 15 - 20%. 3. The effects of cytisine and hexamethonium on m at 0.5 Hz were blocked by 10 microM verapamil, which itself significantly increased m. However, the effects of cytisine and hexamethonium on m at 0.5 Hz were not sensitive to 10 microM of the calmodulin antagonist, W-7. This concentration of W-7 attenuates effects on ACh release mediated by facilitatory prejunctional nicotinic ACh autoreceptors. 4. Our present observations are suggestive of actions of cytisine and hexamethonium to activate and inhibit respectively negative-feedback prejunctional nicotinic ACh autoreceptors. Further, they strengthen the case for the existence of two separate and independent autoregulatory mechanisms for the control of ACh release from motor nerve terminals and give a preliminary insight into the cellular mechanism involved in the autoinhibition of ACh release. PMID- 10725256 TI - The effect of P2 receptor antagonists and ATPase inhibition on sympathetic purinergic neurotransmission in the guinea-pig isolated vas deferens. AB - 1. Intracellular microelectrodes were used to record the transmembrane potential and excitatory junction potentials (e.j.p.s) produced by sympathetic nerve stimulation (1 Hz) in smooth muscle cells of the guinea-pig isolated vas deferens. 2. The symmetrical 3'-urea of 8-(benzamido)naphthalene-1,3,5 trisulphonic acid (NF023) produced a concentration-dependent inhibition of e.j.p. magnitude (IC(50)=4. 8x10(-6) M), but had no effect on the resting membrane potential of the smooth muscle cells. 3. Pyridoxal-5-phosphate (P-5-P) also depressed e.j.p. magnitude in a concentration-dependent manner, but was less potent than NF023 (IC(50)=2.2x10(-5) M). At 10(-4) M and above P-5-P significantly depolarized the smooth muscle cells. 4. The nucleoside triphosphatase inhibitor 6-N,N-diethyl-D-beta, gamma-dibromomethyleneATP (ARL 67156) (5x10(-5) M) significantly increased e.j.p. amplitude. ARL 67156 (10(-4) M) further increased e. j.p. amplitude such that they often reached threshold for initiation of action potentials, causing muscle contraction and expulsion of the recording electrode. 5. After reduction of e.j.p.s by NF023 or P-5-P (both 10(-5) M), subsequent co-addition of ARL 67156 (10(-4) M) significantly increased their magnitude. 6. The overflow of endogenous ATP evoked by field stimulation of sympathetic nerves (8 Hz, 1 min) was measured by HPLC and flurometric detection. ARL 67156 (10(-4) M) enhanced ATP overflow by almost 700% compared to control. 7. We conclude that for electrophysiological studies NF023 is preferable to other P2X receptor antagonists such as pyridoxalphosphate -6-azophenyl-2',4' disulphonic acid (PPADS), suramin or P-5-P. Furthermore, breakdown of endogenous ATP by nucleoside triphosphatases is an important modulator of purinergic neurotransmission in the guinea-pig vas deferens. PMID- 10725255 TI - BIIE0246, a potent and highly selective non-peptide neuropeptide Y Y(2) receptor antagonist. AB - 1. BIIE0246, a newly synthesized non-peptide neuropeptide Y (NPY) Y(2) receptor antagonist, was able to compete with high affinity (8 to 15 nM) for specific [(125)I]PYY(3 - 36) binding sites in HEK293 cells transfected with the rat Y(2) receptor cDNA, and in rat brain and human frontal cortex membrane homogenates. 2. Interestingly, in rat brain homogenates while NPY, C2-NPY and PYY(3 - 36) inhibited all specific [(125)I]PYY(3 - 36) labelling, BIIE0246 failed to compete for all specific binding suggesting that [(125)I]PYY(3 - 36) recognized, in addition to the Y(2) subtype, another population of specific NPY binding sites, most likely the Y(5) receptor. 3. Quantitative receptor autoradiographic data confirmed the presence of [(125)I]PYY(3 - 36)/BIIE0246-sensitive (Y(2)) and insensitive (Y(5)) binding sites in the rat brain as well as in the marmoset monkey and human hippocampal formation. 4. In the rat vas deferens and dog saphenous vein (two prototypical Y(2) bioassays), BIIE0246 induced parallel shifts to the right of NPY concentration-response curves with pA(2) values of 8.1 and 8.6, respectively. In the rat colon (a Y(2)/Y(4) bioassay), BIIE0246 (1 microM) completely blocked the contraction induced by PYY(3 - 36), but not that of [Leu(31), Pro(34)]NPY (a Y(1), Y(4) and Y(5) agonist) and hPP (a Y(4) and Y(5) agonist). Additionally, BIIE0246 failed to alter the contractile effects of NPY in prototypical Y(1) in vitro bioassays. 5. Taken together, these results demonstrate that BIIE0246 is a highly potent, high affinity antagonist selective for the Y(2) receptor subtype. It should prove most useful to establish further the functional role of the Y(2) receptor in the organism. PMID- 10725257 TI - Enhancement of noradrenaline release by angiotensin II and bradykinin in mouse atria: evidence for cross-talk between G(q/11) protein- and G(i/o) protein coupled receptors. AB - 1. The interaction between alpha(2)-autoreceptors and receptors for angiotensin (AT(1)) and bradykinin (B(2)) was studied in mouse isolated atria. The preparations were labelled with [(3)H]-noradrenaline and then superfused with desipramine-containing medium and stimulated electrically. 2. Angiotensin II (10( 11) - 10(-7) M), angiotensin III (10(-10) - 10(-6) M) and bradykinin (10(-11) - 10(-7) M) enhanced the evoked overflow of tritium when preparations were stimulated with conditions that led to marked alpha(2)-autoinhibition (120 pulses at 3 Hz), but not when stimulated with conditions that led to little alpha(2) autoinhibition (20 pulses at 50 Hz). 3. Blockade of alpha-adrenoceptors by phentolamine (1 or 10 microM) reduced or abolished the effect of angiotensin II and bradykinin on the overflow response to 120 pulses at 3 Hz. 4. Addition of the delta-opioid agonist [D-Ser(2)]-leucine enkephalin-Thr (DSLET, 0.1 microM), or of neuropeptide Y (0.1 microM), together with phentolamine, restored the effect of angiotensin II and bradykinin. 5. The beta-adrenoceptor agonist terbutaline (10( 9) - 10(-4) M) enhanced the evoked overflow of tritium irrespective of the degree of autoinhibition. 6. The experiments show that (i) a marked prejunctional facilitatory effect of angiotensin and bradykinin in mouse isolated atria requires prejunctional alpha(2)-autoinhibition; (ii) in the absence of alpha(2) autoinhibition, activation of other prejunctional G(i/o) protein-coupled receptors, namely opioid and neuropeptide Y receptors, restores a marked effect of angiotensin II and bradykinin; and (iii) the facilitatory effect of terbutaline is not dependent upon the degree of alpha(2)-autoinhibition. The findings indicate that the major part of the release-enhancing effect elicited through prejunctional G(q/11) protein-coupled receptors is due to disruption of an ongoing, alpha(2)-autoreceptor-triggered G(i/o) protein mediated inhibition. PMID- 10725258 TI - Role of endothelial cell hyperpolarization in EDHF-mediated responses in the guinea-pig carotid artery. AB - 1. Experiments were performed to identify the potassium channels involved in the acetylcholine-induced endothelium-dependent hyperpolarization of the guinea-pig internal carotid artery. Smooth muscle and endothelial cell membrane potentials were recorded in isolated arteries with intracellular microelectrodes. Potassium currents were recorded in freshly-dissociated smooth muscle cells using patch clamp techniques. 2. In single myocytes, iberiotoxin (0.1 microM)-, charybdotoxin (0.1 microM)-, apamin (0.5 microM)- and 4-aminopyridine (5 mM)-sensitive potassium currents were identified indicating the presence of large- and small conductance calcium-sensitive potassium channels (BK(Ca) and SK(Ca)) as well as voltage-dependent potassium channels (K(V)). Charybdotoxin and iberiotoxin inhibited the same population of BK(Ca) but a conductance specifically sensitive to the combination of charybdotoxin plus apamin could not be detected. 4 aminopyridine (0. 1 - 25 mM) induced a concentration-dependent inhibition of K(V) without affecting the iberiotoxin- or the apamin-sensitive currents. 3. In isolated arteries, both the endothelium-dependent hyperpolarization of smooth muscle and the hyperpolarization of endothelial cells induced by acetylcholine or by substance P were inhibited by 5 mM 4-aminopyridine. 4. These results indicate that in the vascular smooth muscle cells of the guinea-pig carotid artery, a conductance specifically sensitive to the combination of charybdotoxin plus apamin could not be detected, comforting the hypothesis that the combination of these two toxins should act on the endothelial cells. Furthermore, the inhibition by 4-aminopyridine of both smooth muscle and endothelial hyperpolarizations, suggests that in order to observe an endothelium-dependent hyperpolarization of the vascular smooth muscle cells, the activation of endothelial potassium channels is likely to be required. PMID- 10725259 TI - Both alpha(1A)- and alpha(1B)-adrenergic receptor subtypes couple to the transient outward current (I(To)) in rat ventricular myocytes. AB - 1. Regulation of transient outward current (I(To)) by alpha(1)-adrenergic (alpha(1)AR) plays a key role in cardiac repolarization. alpha(1)ARs comprise a heterogeneous family; two natively expressed subtypes (alpha(1A) and alpha(1B)) and three cloned subtypes (alpha(1a), alpha(1b) and alpha(1d)) can be distinguished. We have examined the electrophysiological role of each alpha(1)AR subtype in regulating I(To) in isolated rat ventricular myocytes. 2. Reverse transcription-PCR study revealed the presence of three subtype mRNAs (alpha(1a), alpha(1b) and alpha(1d)) in rat myocytes. 3. Radioligand binding assay using [(125)I]-HEAT showed that the inhibition curves for alpha(1A)AR-selective antagonists (WB4101, 5-methylurapidil, (+)-niguldipine and KMD-3213) in rat ventricles best fit a two-site model, with 30% high and 70% low affinity binding sites. The high affinity sites were resistant to 100 microM chloroethylclonidine (CEC), while the low affinity sites were highly inactivated by CEC. 4. Whole cell voltage clamp study revealed that methoxamine reduced a 4-aminopyridine(4-AP) sensitive component of I(To) in the isolated rat ventricle myocytes. Lower concentrations of KMD-3213 (1 nM) or 5-MU (10 nM) did not affect the methoxamine induced reduction of I(To). On the other hand, CEC treatment (100 microM) of isolated myocytes reduced the methoxamine-induced reduction of I(To) by 46%, and the remaining response was abolished by lower concentrations of KMD-3213 or 5-MU. 5. The results indicate that rat ventricular myocytes express transcripts of the three alpha(1)AR subtypes (alpha(1a), alpha(1b) and alpha(1d)); however, two pharmacologically distinct alpha(1)AR subtypes (alpha(1A) and alpha(1B)) are predominating in receptor populations, with approximately 30% alpha(1A)AR and 70% alpha(1B)AR. Although both alpha(1A) and alpha(1B)AR subtypes are coupled to the cardiac I(To), alpha(1B)ARs predominantly mediate alpha(1)AR-induced effect. PMID- 10725260 TI - Respiratory actions of tachykinins in the nucleus of the solitary tract: characterization of receptors using selective agonists and antagonists. AB - 1. The respiratory response to microinjection of tachykinins and analogues into the commissural nucleus of the solitary tract (cNTS) of urethane-anaesthetized rats was investigated in the presence and absence of selective tachykinin NK(1), NK(2) and NK(3) antagonists (RP 67580, SR 48968 and SR 142801, respectively). 2. All tachykinins, except for the selective NK(2) agonist, [Nle(10)]-NKA(4-10), increased tidal volume (VT). The rank potency order of naturally-occurring tachykinins was neurokinin A (NKA)> or =substance P (SP)>>NKB, whereas the rank order for selective analogues was senktide> or = septide>> [Sar(9),Met(O(2))(11)] SP>>[Nle(10)]-NKA(4-10). Septide (NK(1)-selective) and senktide (NK(3)-selective) were 22 fold more potent (pD(2) approximately 12) at stimulating VT than SP (pD(2) approximately 10.5). 3. Tachykinin agonists produced varying degrees of respiratory slowing, independent of changes in VT. At doses producing maximum stimulation of VT, agonists induced either a mild (<10 breaths min(-1) decrease; SP and septide), moderate (10 - 25 breaths min(-1) decrease; NKA, NKB and [Sar(9),Met(O(2)]-SP) or severe ( approximately 40 breaths min(-1) decrease; senktide) bradypnoea. [Nle(10)]-NKA(4-10) produced a dose-dependent bradypnoea without affecting VT. 4. RP 67580 significantly attenuated the VT response to SP (33 pmol) and NKA (10 pmol) but not NKB (100 pmol). In the presence of RP 67580, the mild bradypnoeic response to NKB was significantly enhanced whereas SP and NKA induced a bradyapnea which was not observed in the absence of RP 67580. SR 48968 had no effect on the VT response to SP or NKB, markedly enhanced the VT response to NKA and completely blocked the bradypnoeic response to [Nle(10)] NKA(4-10). Only SR142801 attenuated the VT response to NKB. 5. The present data suggest that all three tachykinin receptors (NK(1), NK(2) and NK(3)) are present in the cNTS and are involved in the central control of respiration. PMID- 10725262 TI - The role of adenosine receptors in the action of theophylline on human peripheral blood mononuclear cells from healthy and asthmatic subjects. AB - 1. The aim of the present study was to investigate the role of adenosine A2b receptors in the anti-proliferative action of theophylline in human peripheral blood mononuclear cells (HPBMC) from healthy and asthmatic subjects. 2. Theophylline significantly inhibited PHA-induced proliferation of HPBMC from both healthy and asthmatic donors but only at relatively high concentrations at 1 mM (P<0.05). Enprophylline, a drug which also acts as a non-selective phosphodiesterase (PDE) inhibitor and is a selective A2b receptor antagonist, had no significant effect on proliferation of cells from either group at concentrations up to 10 microM (P>0.05; n=6). 3. Adenosine deaminase (2 u ml( 1)), which metabolizes adenosine, had no significant effect on PHA-induced HPBMC proliferation over a range of concentrations (0 - 8 microg ml(-1)) in cells from either healthy or asthmatic subjects. 4. The adenosine receptor agonists N(6) cyclopentyladenosine (CPA, A1-selective) and 5'-N-ethylcarboxamidoadenosine (NECA, A1/A2) produced a small but significant inhibition of PHA-induced proliferation of HPBMC from healthy and asthmatic subjects (10 microM, P<0.05; n=6). In contrast, 5'-N-ethylcarboxamido-2-[4-(2 ]carboxyethyl)phenethyl]adenosine (CGS21680, A2a-selective) was without significant effect (P>0.05; n=6). 5. The adenosine receptor antagonist alloxazine (A2b-selective) had no significant effect, while 8(3-chlorostyryl)caffeine,(CSC, A2a-selective) significantly inhibited PHA-induced proliferation of HPBMC from both groups (P<0.05; n=6). 6. Our results suggest that endogenous or exogenous adenosine has little effect on the proliferation of HPBMC obtained from healthy or asthmatic subjects. Thus it would appear that the effect of high concentrations of theophylline is not related to adenosine receptor antagonism. PMID- 10725261 TI - Respiratory actions of tachykinins in the nucleus of the solitary tract: effect of neonatal capsaicin pretreatment. AB - 1. The respiratory response to microinjection of capsaicin and tachykinin receptor agonists into the commissural nucleus of the solitary tract (cNTS) was investigated in adult, urethane-anaesthetized rats which had been pretreated with capsaicin (50 mg kg(-1) s.c.) or vehicle (10% Tween 80, 10% ethanol in saline) as day 2 neonates. 2. Microinjection of capsaicin (1 nmol) into the cNTS of vehicle pretreated rats, significantly reduced respiratory frequency (59 breaths min(-1), preinjection control, 106 breaths min(-1)) without affecting tidal volume (VT). In capsaicin-pretreated rats, the capsaicin-induced bradypnoea was markedly attenuated (minimum frequency, 88 breaths min(-1); control, 106 breaths min(-1)). 3. In vehicle-pretreated rats, microinjection of substance P (SP, 33 pmol), neurokinin A (NKA, 33 pmol) and NKB (330 pmol), and the selective NK(1) tachykinin receptor agonists, [Sar(9), Met(O(2))(11)]-SP (33 pmol) and septide (10 pmol), increased VT (maxima, 3.60 - 3.93 ml kg(-1)) compared with preinjection control (2.82 ml kg(-1)), without affecting frequency. The selective NK(3) agonist senktide (10 pmol) also increased VT (3.93 ml kg(-1)) which was accompanied by a bradypnoea (-25 breaths min(-1)). The selective NK(2) agonist, [Nle(10)]-NKA(4-10) (330 pmol) increased VT slightly but significantly decreased frequency (-12 breaths min(-1)). In capsaicin-pretreated rats, VT responses to SP and [Sar(9), Met(O(2))(11)]-SP were increased whereas the response to septide was abolished. Both the VT and bradypnoeic responses to senktide and [Nle(10)]-NKA(4 10) were significantly enhanced. 4. These results show that neonatal capsaicin administration markedly reduces the respiratory response to microinjection of capsaicin into the cNTS. The destruction of capsaicin-sensitive afferents appears to sensitize the NTS to SP, NKB, [Sar(9),Met(O(2))(11)]-SP, senktide and [Nle(10)]-NKA(4-10). Moreover, the loss of septide responsiveness in capsaicin pretreated rats, suggests that 'septide-sensitive' NK(1) receptors may be located on the central terminals of afferent neurons. PMID- 10725263 TI - Role of gap junctions and EETs in endothelium-dependent hyperpolarization of porcine coronary artery. AB - 1. The effects of endothelium-derived hyperpolarizing factor (EDHF: elicited using substance P or bradykinin) were compared with those of 11,12-EET in pig coronary artery. Smooth muscle cells were usually impaled with microelectrodes through the adventitial surface. 2. Substance P (100 nM) and 11,12-EET (11,12 epoxyeicosatrienoic acid; 3 microM) hyperpolarized endothelial cells in intact arteries. These actions were unaffected by 100 nM iberiotoxin but were abolished by charybdotoxin plus apamin (each 100 nM). 3. Substance P (100 nM) and bradykinin (30 nM) hyperpolarized intact artery smooth muscle; Substance P had no effect after endothelium removal. 11,12-EET hyperpolarized de-endothelialized vessels by 12.6+/-0.3 mV, an effect abolished by 100 nM iberiotoxin. 4. 11,12-EET hyperpolarized intact arteries by 18.6+/-0.8 mV, an action reduced by iberiotoxin, which was ineffective against substance P. Hyperpolarizations to 11, 12-EET and substance P were partially inhibited by 100 nM charybdotoxin and abolished by further addition of 100 nM apamin. 5. 30 microM barium plus 500 nM ouabain depolarized intact artery smooth muscle but responses to substance P and bradykinin were unchanged. 500 microM gap 27 markedly reduced hyperpolarizations to substance P and bradykinin which were abolished in the additional presence of barium plus ouabain. 6. Substance P-induced hyperpolarizations of smooth muscle cells immediately below the internal elastic lamina were unaffected by gap 27, even in the presence of barium plus ouabain. 7. In pig coronary artery, 11,12-EET is not EDHF. Smooth muscle hyperpolarizations attributed to 'EDHF' are initiated by endothelial cell hyperpolarization involving charybdotoxin- (but not iberiotoxin) and apamin-sensitive K(+) channels. This may spread electrotonically via myoendothelial gap junctions but the involvement of an unknown endothelial factor cannot be excluded. PMID- 10725264 TI - Modulation by dihydropyridine-type calcium channel antagonists of cytokine inducible gene expression in vascular smooth muscle cells. AB - 1. The 1,4-dihydropyridine nifedipine is frequently used in the therapy of hypertension and heart failure. In addition, nifedipine has been shown to exert distinct anti-arteriosclerotic effects both in experimental animal models and in patients. In the present study we have investigated the hypothesis that the latter effect of this class of drugs is mediated by an interference with the expression of pro-arteriosclerotic gene products in the vessel wall. Moreover, to elucidate as to whether nifedipine acts via L-type calcium channel blockade, its effects were compared to those of another dihydropyridine, Bay w 9798, which has no calcium-antagonistic properties in concentrations up to 10 microM as verified by superfusion bioassay. 2. Both, nifedipine and Bay w 9798, in concentrations ranging from 0.01 to 1 microM, augmented the interleukin-1beta/tumour necrosis factor-alpha (IL-1beta/TNF-alpha)-induced expression of the inducible isoform of nitric oxide synthase (iNOS) in rat aortic cultured smooth muscle cells (raSMC) 2 - 3 fold, as judged by RT - PCR and Western blot analyses. 3. In contrast, cytokine-induced mRNA expression of monocyte chemoattractant protein 1 (MCP-1) in these cells was down-regulated by more than 60% in the presence of both dihydropyridines, as judged by RT - PCR and Northern blot analyses. 4. Nuclear run-on assays and incubation with the transcription-terminating drug actinomycin D revealed that both drugs acted at the level of mRNA synthesis rather than stability. 5. These findings suggest that 1,4-dihydropyridines such as nifedipine affect the expression of both potentially pro-arteriosclerotic (MCP-1) and anti arteriosclerotic (iNOS) gene products in the vessel wall at the level of transcription, and that these effects are unrelated to their calcium channel blocking properties. PMID- 10725266 TI - Active immunization with angiotensin I peptide analogue vaccines selectively reduces the pressor effects of exogenous angiotensin I in conscious rats. AB - 1. Male, Sprague-Dawley rats were actively immunized with novel angiotensin vaccines, and their pressor responses to exogenous angiotensin I (AI) and angiotensin II (AII) were assessed in vivo. Serum antibody titres were also measured. 2. The most effective vaccine consisted of an AI analogue conjugated with a tetanus toxoid carrier protein and adjuvanted with aluminium hydroxide. When this vaccine was injected on days 0, 21 and 42, pressor responses to AI on day 63 were significantly inhibited (maximum, 8.9 fold shift), but responses to AII were unaffected. The anti-angiotensin antibody titre was increased 32,100 fold, and, uniquely, these antibodies also cross-reacted with angiotensinogen. 3. These findings indicate that active immunization against AI may be a useful approach for treating cardiovascular disorders involving the renin-angiotensin system. PMID- 10725265 TI - Vasorelaxant and antiplatelet activity of 4,7-dimethyl-1,2, 5-oxadiazolo[3,4 d]pyridazine 1,5,6-trioxide: role of soluble guanylate cyclase, nitric oxide and thiols. AB - 1. Certain heterocyclic N-oxides are vasodilators and inhibitors of platelet aggregation. The pharmacological activity of the furoxan derivative condensed with pyridazine di-N-oxide 4,7-dimethyl-1,2, 5-oxadiazolo[3,4-d]pyridazine 1,5,6 trioxide (FPTO) and the corresponding furazan (FPDO) was studied. 2. FPTO reacted with thiols generating nitrite (NO), S-nitrosoglutathione and hydroxylamine (nitroxyl) and converted oxyHb to metHb. FPDO did not generate detectable amounts of NO-like species but reacted with thiols and oxyHb. 3. FPTO and FPDO haem dependently stimulated the activity of soluble guanylate cyclase (sGC) and this stimulation was inhibited by 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) and by 0.1 mM dithiothreitol. 4. FPTO relaxed noradrenaline-precontracted aortic rings and its concentration-response curve was biphasic (pIC(50)=9. 03+/-0.13 and 5.85+/-0.06). FPDO was significantly less potent vasodilator (pIC(50)=5.19+/ 0.14). The vasorelaxant activity of FPTO and FPDO was inhibited by ODQ. oxyHb significantly inhibited only FPTO-dependent relaxation. 5. FPTO and FPDO were equipotent inhibitors of ADP-induced platelet aggregation (IC(50)=0.63+/-0.15 and 0.49+/-0. 05 microM, respectively). The antiplatelet activity of FPTO (but not FPDO) was partially suppressed by oxyHb. The antiaggregatory effects of FPTO and FPDO were only partially blocked by sGC inhibitors. 6. FPTO and FPDO (10 - 20 microM) significantly increased cyclic GMP levels in aortic rings and platelets and this increase was blocked by ODQ. 7. Thus, FPTO can generate NO and, like FPDO, reacts with thiols and haem. The vasorelaxant activity of FPTO and FPDO is sGC-dependent and a predominant role is played by NO at FPTO concentrations below 1 microM. On the contrary, inhibition of platelet aggregation is only partially related to sGC activation. PMID- 10725268 TI - Potassium- and acetylcholine-induced vasorelaxation in mice lacking endothelial nitric oxide synthase. AB - 1. The contribution of an endothelium-derived hyperpolarizing factor (EDHF) was investigated in saphenous and mesenteric arteries from endothelial nitric oxide synthase (eNOS) (-/-) and (+/+) mice. 2. Acetylcholine-induced endothelium dependent relaxation of saphenous arteries of eNOS(-/-) was resistant to N(omega) nitro-L-arginine (L-NNA) and indomethacin, as well as the guanylyl cyclase inhibitor, 1H-(1,2,4)oxadiazolo(4,3-a) quinoxalin-1-one(ODQ). 3. Potassium (K(+)) induced a dose-dependent vasorelaxation which was endothelium-independent and unaffected by either L-NNA or indomethacin in both saphenous and mesenteric arteries from eNOS(-/-) or (+/+) mice. 4. Thirty microM barium (Ba(2+)) and 10 microM ouabain partially blocked potassium-induced, but had no effect on acetylcholine-induced vasorelaxation in saphenous arteries. 5. Acetylcholine induced relaxation was blocked by a combination of charybdotoxin (ChTX) and apamin which had no effect on K(+)-induced relaxation, however, iberiotoxin (IbTX) was ineffective against either acetylcholine- or K(+)-induced relaxation. 6. Thirty microM Ba(2+) partially blocked both K(+)- and acetylcholine-induced relaxation of mesenteric arteries, and K(+), but not acetylcholine-induced relaxation was totally blocked by the combination of Ba(2+) and ouabain. 7. These data indicate that acetylcholine-induced relaxation cannot be mimicked by elevating extracellular K(+) in saphenous arteries from either eNOS(-/-) or (+/+) mice, but K(+) may contribute to EDHF-mediated relaxation of mesenteric arteries. PMID- 10725267 TI - Characterization of [Nphe(1)]nociceptin(1-13)NH(2), a new selective nociceptin receptor antagonist. AB - 1.. Nociceptin (orphanin FQ) is a novel neuropeptide capable of inducing a variety of biological actions via activation of a specific G-protein coupled receptor. However, the lack of a selective nociceptin receptor antagonist has hampered our understanding of nociceptin actions and the role of this peptide in pathophysiological states. As part of a broader programme of research, geared to the identification and characterization of nociceptin receptor ligands, we report that the novel peptide [Nphe(1)]nociceptin(1-13)NH(2) acts as the first truly selective and competitive nociceptin receptor antagonist and is devoid of any residual agonist activity. 2. [Nphe(1)]nociceptin(1-13)NH(2) binds selectively to recombinant nociceptin receptors expressed in Chinese hamster ovary (CHO) cells (pK(i) 8.4) and competitively antagonizes the inhibitory effects of nociceptin (i) on cyclic AMP accumulation in CHO cells (pA(2) 6.0) and (ii) on electrically evoked contractions in isolated tissues of the mouse, rat and guinea-pig with pA(2) values ranging from 6.0 to 6.4. 3. [Nphe(1)]nociceptin(1-13)NH(2) is also active in vivo, where it prevents the pronociceptive and antimorphine actions of intracerebroventricularly applied nociceptin, measured in the mouse tail withdrawal assay. Moreover, [Nphe(1)]nociceptin(1-13)NH(2) produces per se a dose dependent, naloxone resistant antinociceptive action and, at relatively low doses, potentiates morphine-induced analgesia. 4. Collectively our data indicate that [Nphe(1)]nociceptin(1-13)NH(2), acting as a nociceptin receptor antagonist, may be the prototype of a new class of analgesics. PMID- 10725269 TI - Influence of bilirubin and other antioxidants on nitrergic relaxation in the pig gastric fundus. AB - 1. The influence of several antioxidants (bilirubin, urate, ascorbate, alpha tocopherol, glutathione (GSH), Cu/Zn superoxide dismutase (SOD) and the manganese SOD mimic EUK-8) on nitrergic relaxations induced by either exogenous nitric oxide (NO; 10(-5) M) or electrical field stimulation (4 Hz; 10 s and 3 min) was studied in the pig gastric fundus. 2. Ascorbate (5x10(-4) M), alpha-tocopherol (4x10(-4) M), SOD (300 - 1000 u ml(-1)) and EUK-8 (3x10(-4) M) did not influence the relaxations to exogenous NO. In the presence of GSH (5x10(-4) M), the short lasting relaxation to NO became biphasic, potentiated and prolonged. Urate (4x10( 4) M) and bilirubin (2x10(-4) M) also potentiated the relaxant effect of NO. None of the antioxidants influenced the electrically evoked relaxations. 3. 6-Anilino 5,8-quinolinedione (LY83583; 10(-5) M) had no influence on nitrergic nerve stimulation but nearly abolished the relaxant response to exogenous NO. Urate and GSH completely prevented this inhibitory effect, while it was partially reversed by SOD and bilirubin. Ascorbate, alpha-tocopherol and EUK-8 were without effect. 4. Hydroquinone (10(-4) M) did not affect the electrically induced nitrergic relaxations, but markedly reduced NO-induced relaxations. The inhibition of exogenous NO by hydroquinone was completely prevented by urate and GSH. SOD and ascorbate afforded partial protection, while bilirubin, EUK-8 and alpha tocopherol were ineffective. 5. Hydroxocobalamin (10(-4) M) inhibited relaxations to NO by 50%, but not the electrically induced responses. Full protection versus this inhibitory effect was obtained with urate, GSH and alpha-tocopherol. 6. These results strengthen the hypothesis that several endogenous antioxidant defense mechanisms, enzymatic as well as non-enzymatic, might play a role in the nitrergic neurotransmission process. PMID- 10725270 TI - The effect of long-term streptozotocin-induced diabetes on contractile and relaxation responses of coronary arteries: selective attenuation of CGRP-induced relaxations. AB - 1. This study investigates the effect of partially metabolic controlled long-term (34 weeks) streptozotocin (STZ)-induced diabetes on relaxation and contractile responses of isolated coronary arteries to seven different vasoactive agents. 2. The average fasting and non-fasting blood glucose concentrations (mM) were significantly elevated in STZ-induced diabetic rats (P<0.0001; 10.4+/-0.4 and 16. 6+/-1.1, n=15) compared to those (4.3+/-0.03 and 4.7+/-0.18, n=11) in age-matched controls. The level of glycated haemoglobin (HbA(1)) was also significantly (P<0.0001) increased in STZ-induced diabetic rats. In STZ-induced diabetic rats, the HbA(1) levels were significantly correlated with the non-fasting blood glucose concentrations (r=0.76; P=0.003; n=13). In both groups, there was no significant correlation between the HbA(1) levels and maximal responses or sensitivities to the vasoactive agents. 3. The maximal relaxation induced by rat alphacalcitonin gene-related peptide (rat-alphaCGRP) was significantly attenuated in the coronary arteries of STZ-induced diabetic rats (P<0.05; 40+/-7%, n=15) compared to that in age-matched controls (63+/-3%, n=11). However, there was no significant difference in the sensitivity to rat-alphaCGRP between the two groups. 4. There was no significant difference in either maximal response or sensitivity to any of the six other vasoactive agents between STZ- induced diabetic rats (n=15) and age-matched controls (n=11). 5. Our results show that partially metabolic controlled long-term (34 weeks) STZ-induced diabetes causes a selective depression of rat-alphaCGRP-induced relaxation in the intramural coronary arteries of Wistar rats. PMID- 10725271 TI - Preservation of mitochondrial function by diazoxide during sustained ischaemia in the rat heart. AB - 1. A possible mechanism for the action of the K(ATP) channel opener diazoxide on the improvement of energy metabolism of ischaemic/reperfused hearts was examined. 2. Isolated, perfused rat hearts were subjected to 40 min ischaemia followed by 60 min reperfusion. Diazoxide at concentrations of 3 to 30 microM was present in the perfusion buffer for the last 15 min of pre-ischaemia. 3. Treatment of the perfused heart with diazoxide enhanced the post-ischaemic recovery of rate pressure product, attenuated the post-ischaemic rise in left ventricular end diastolic pressure, and suppressed the release of creatine kinase and purine nucleosides and bases from the reperfused heart. Treatment of the heart with diazoxide also restored myocardial ATP and creatine phosphate and attenuated the decrease in mitochondrial oxygen consumption rate after reperfusion. This attenuation was maintained at the end of ischaemia as well as at the end of reperfusion. 4. In another set of experiments, myocardial skinned bundles were incubated for 30 min under hypoxic conditions in the presence and absence of diazoxide, and then the mitochondrial oxygen consumption rate was determined. Hypoxia induced a decrease in the mitochondrial oxygen consumption rate of the skinned bundles to approximately 40% of the pre-hypoxic value. In contrast, treatment of the bundles with 30 microM diazoxide preserved the normal mitochondrial oxygen consumption rate during hypoxia. This effect was abolished concentration-dependently by the combined treatment with either the K(ATP) channel blocker glibenclamide or 5-hydroxydecanoate. 5. These results suggest that diazoxide is capable of attenuating ischaemia/reperfusion injury of isolated perfused hearts due to preservation of mitochondrial function during ischaemia. PMID- 10725272 TI - Effect of type-selective inhibitors on cyclic nucleotide phosphodiesterase activity and insulin secretion in the clonal insulin secreting cell line BRIN BD11. AB - 1. The cyclic nucleotide phosphodiesterases (PDEs) present in an insulin secreting cell line, BRIN - BD11, were characterized using calcium/calmodulin, IGF-1, isoenzyme-selective PDE inhibitors and RT - PCR. 2. Calmodulin activated cyclic AMP or cyclic GMP PDE activity in pellet and was 3 fold (P=0.002) more potent in activating cyclic nucleotide hydrolysis in pellet compared with supernatant fractions. 3. The PDE1/PDE5 inhibitor zaprinast inhibited both cyclic AMP and cyclic GMP PDE activity in both pellet and supernatant fractions of cell homogenates by a maximum of around 25% (IC(50) 1 - 5 microM), while rolipram (PDE4 selective) inhibited only cyclic AMP hydrolysis. 4. The PDE3-selective inhibitors Org 9935 (0.02 - 10 microM) and siguazodan (0.1 - 10 microM) inhibited cyclic AMP PDE activity in the pellet but not the supernatant fractions of cell homogenates, with a maximum inhibition of about 30%. IGF-1 (2 - 7.5 ng ml(-1)) potently augmented this PDE activity. 5. RT - PCR using specific primers for PDE3B, but not for PDE3A, amplified, from BRIN - BD11 cell total RNA, a 351 base pair product that was >97% homologous with rat adipose tissue PDE3B. 6. IBMX, Org 9935, siguazodan and rolipram (1 - 50 microM), but not zaprinast, each augmented glucose-induced insulin secretion in the presence of 16.7 mM but not 1 mM glucose. 7. These findings, in a clonal insulin secreting cell line, are consistent with an important role for PDE3B in regulating the pool of cyclic AMP relevant to the modulation of glucose-induced insulin secretion. PMID- 10725273 TI - Active transport of the angiotensin-II antagonist losartan and its main metabolite EXP 3174 across MDCK-MDR1 and caco-2 cell monolayers. AB - 1. We studied the functional interaction between transport and metabolism by comparing the transport of losartan and its active metabolite EXP 3174 (EXP) across cell monolayers. 2. Epithelial layers of Caco-2 cells as well as MDR1, MRP 1 and MRP-2 overexpressing cells, in comparison to the respective wildtypes, were used to characterize the transcellular transport of losartan and EXP. 3. Losartan transport in MDCK-MDR1 and Caco-2 cells was saturable and energy-dependent with a significantly greater basolateral-to-apical (B/A) than apical-to-basolateral (A/B) flux (ratio=31+/-1 in MDCK-MDR1 and ratio 4+/-1 in Caco-2 cells). The B/A flux of losartan was inhibited by cyclosporine and vinblastine, inhibitors of P glycoprotein and MRP. In contrast, no active losartan transport was observed in MRP-1 or MRP-2 overexpressing cells. 4. The metabolite was only transported in Caco-2 cells with a B/A-to-A/B ratio of 5+/-1, while lacking active transport in the MDR1, MRP-1 or MRP-2 overexpressing cells. The B/A flux of EXP was significantly inhibited by cyclosporine and vinblastine. 5. In conclusion, losartan is transported by P-glycoprotein and other intestinal transporters, that do not include MRP-1 and MRP-2. In contrast, the carboxylic acid metabolite is not a P-glycoprotein substrate, but displays considerably higher affinity for other transporters than losartan, that again most probably do not include MRP-1 and MRP-2. PMID- 10725276 TI - Circulation Online Only : March 21, 2000. PMID- 10725274 TI - Effects of AMP derivatives on cyclic AMP levels in NG108-15 cells. AB - 1. In NG108-15 neuroblastomaxglioma hybrid cells, ATP stimulates intracellular cyclic AMP formation, which is inhibited by both adenosine (P(1)) and P2 receptor antagonists. In the present study, we examined the effects of several AMP derivatives in NG108-15 cells and mouse neuroblastoma N18TG-2 cells. 2. Adenosine 2'-monophosphate (A2P), adenosine 3'-monophosphate (A3P) and adenosine 5' phosphosulphate (A5PS) increased cyclic AMP levels with similar concentration dependencies in NG108-15 cells. 3. Increases in cyclic AMP by AMP derivatives were inhibited by the P2 receptor antagonist PPADS, but not by suramin. Effects of AMP derivatives were also inhibited by P(1) receptor antagonists ZM241385, XAC, DPCPX and partially by alloxazine. The ecto-nucleotidase inhibitor alpha, beta-methyleneADP was without effect. 4. In contrast, AMP derivatives did not change cyclic AMP levels in N18TG-2 cells. Accumulation of cyclic AMP in N18TG-2 cells was stimulated by adenosine A(2) receptor agonists CGS21680 and NECA, but not by ATP or beta, gamma-methyleneATP, agonists for cyclic AMP production in NG108-15 cells. 5. Reverse transcription-coupled polymerase chain reaction (RT - PCR) analyses revealed that N18TG-2 cells express both A(2A) and A(2B) receptors, while NG108-15 cells express mainly A(2A) receptors. 6. AMP derivatives did not affect the P2X and P2Y receptors expressed in NG108-15 cells. 7. These results suggest that A2P, A3P and A5PS act as agonists for cyclic AMP production and that these compounds are valuable tools for determinating the mechanism of ATP stimulated cyclic AMP response in NG108-15 cells. PMID- 10725277 TI - Hypertrophic cardiomyopathy: do we have the algorithm for life and death? PMID- 10725275 TI - beta(3)-adrenoceptor regulation and relaxation responses in mouse ileum. AB - 1. This study examines the relationship between beta(3a)- and beta(3b) adrenoceptor (AR) mRNA levels, beta(3)-AR binding and changes in ileum responses in mice treated with the beta(3)-AR agonist (R, R)-5-[2[[2-(3-chlorophenyl)-2 hydroxyethyl]-amino]-propyl]1, 3-benzodioxole-2,2-dicarboxylate (CL316243), or the beta(3)-AR antagonist 3-(2-ethylphenoxy)-1-[(1S)-1,2,3, 4-tetrahydronapth-1 ylamino]-2S-2-propanol oxalate (SR59230A), or dexamethasone or forskolin. 2. Levels of beta(3a)- and beta(3b)-AR mRNA and the maximum number of binding sites (B(max)) in ileum were unaffected following CL316243 treatment, although responses to CL316243 were reduced by 50% following 4 and 24 h treatment, indicating another desensitization mechanism not involving changes in receptor expression or number. beta(3a)-AR mRNA levels were reduced in both brown (BAT) and white adipose tissue (WAT) but beta(3b)-AR mRNA levels were significantly reduced only in WAT. Levels of beta(3a)- and beta(3b)-mRNA returned towards normal with continued treatment. 3. SR59230A treatment markedly increased beta(3) AR mRN levels in ileum and BAT but not in WAT. The increase in beta(3)-AR mRNA levels in ileum was associated with increased B(max) levels in binding analysis and increased responses to CL316243, suggesting these as the cause of sensitization. 4. Treatment with forskolin (4 h) or dexamethasone (4 h) significantly reduced beta(3a)-AR mRNA levels in BAT and WAT but did not alter levels in ileum. Responses to CL316243 in ileum were unaffected by either treatment. 5. In summary, the beta(3)-AR is differently regulated in adipose tissue and ileum: Treatment with SR59230A increased beta(3)-AR number, mRNA and responsiveness in ileum, whereas treatment with CL316243 reduced responses without affecting beta(3)-AR number or mRNA levels. PMID- 10725278 TI - Heat shock protein 47 : a chaperone for the fibrous cap? PMID- 10725279 TI - Heat shock protein 47 is expressed in fibrous regions of human atheroma and Is regulated by growth factors and oxidized low-density lipoprotein. AB - BACKGROUND: Heat shock protein 47 (Hsp47) is a stress protein that may act as a chaperone for procollagen. Its involvement in atherosclerosis is unknown. METHODS AND RESULTS: Hsp47 expression in human coronary arteries was assessed by immunostaining. Strong focal expression was evident in atherosclerotic, but not normal, arteries and was prevalent in the collagenous regions. Double immunostaining revealed that all cells expressing type I procollagen also expressed Hsp47. Moreover, parallel regulation of proalpha1(I)collagen and Hsp47 mRNA expression occurred with cultured human smooth muscle cells stimulated with transforming growth factor-beta1 or fibroblast growth factor-2. However, a proportion of Hsp47-expressing cells in plaque did not express type I procollagen, and this pattern could be reproduced in culture. Heat shock and oxidized LDL stimulated the expression of Hsp47 mRNA by smooth muscle cells, without a concomitant rise in proalpha1(I)collagen expression. CONCLUSIONS: These findings identify Hsp47 as a novel constituent of human coronary atheroma. Its localization to the fibrous cap, regulation by growth factors in parallel with type I procollagen, and selective upregulation by stress raise the possibility that Hsp47 is a determinant of plaque stability. PMID- 10725280 TI - Vascular effects following homozygous disruption of p47(phox) : An essential component of NADPH oxidase. AB - BACKGROUND: Evidence suggests that the vessel wall contains an oxidase similar, if not identical, to phagocytic NADPH oxidase. We tested the contribution of this specific oxidase to the progression of atherosclerosis and the regulation of blood pressure. METHODS AND RESULTS: An examination of aortic rings from wild type mice and mice with homozygous targeted disruptions in p47(phox) revealed that p47(phox) knockout mice had a reduction in vascular superoxide production. However, analyses of apoE -/- p47(phox)+/+ and apoE -/- p47(phox) -/- strains of mice demonstrated no significant differences in atherosclerotic lesion sizes. Similarly, analyses of wild-type and p47(phox) knockout mice revealed no differences in either basal blood pressure or the rise in blood pressure seen after the pharmacological inhibition of nitric oxide synthase. CONCLUSIONS: NADPH oxidase contributes to basal vascular superoxide production. However, the absence of a functional oxidase does not significantly affect the progression of atherosclerosis in the standard mouse apoE -/- model, nor does it significantly influence basal blood pressure. PMID- 10725281 TI - Temporal repolarization lability in hypertrophic cardiomyopathy caused by beta myosin heavy-chain gene mutations. AB - BACKGROUND: Certain genetic mutations associated with hypertrophic cardiomyopathy (HCM) carry an increased risk of sudden death. QT variability identifies patients at a high risk for sudden death from ventricular arrhythmias. We tested whether patients with HCM caused by beta-myosin heavy-chain (beta-MHC) gene mutations exhibit labile ventricular repolarization using beat-to-beat QT variability analysis. METHODS AND RESULTS: We measured the QT variability index and heart rate-QT interval coherence from Holter monitor recordings in 36 patients with HCM caused by known beta-MHC gene mutations and in 26 age- and sex-matched controls. There were 7 distinct beta-MHC gene mutations in these 36 patients; 9 patients had HCM caused by the malignant Arg(403)Gln mutation and 8 patients had HCM caused by the more benign Leu(908)Val mutation. The QT variability index was higher in HCM patients than in controls (-1.24+/-0.17 versus -1. 58+/-0.38, P<0.01), and the greatest abnormality was detected in patients with the Arg(403)Gln mutation (-0.99+/-0.49 versus -1. 46+/-0.43 in controls, P<0.05). In keeping with this finding, coherence was lower for the entire HCM group than for controls (P<0. 001). Coherence was also significantly lower in patients with the Arg(403)Gln mutation compared with controls (P<0.05). CONCLUSIONS: These findings suggest that (1) patients with HCM caused by beta-MHC gene mutations exhibit labile repolarization quantified by QT variability analysis and, hence, may be more at risk for sudden death from ventricular arrhythmias, and (2) indices of QT variability may be particularly abnormal in patients with beta-MHC gene mutations that are associated with a poor prognosis. PMID- 10725282 TI - A comparison of the Framingham risk index, coronary artery calcification, and culprit plaque morphology in sudden cardiac death. AB - BACKGROUND: Neither clinical prediction models nor noninvasive imaging tests that detect coronary artery calcification identify all patients who experience acute coronary events. Variations in culprit plaque morphology may account for these inaccuracies. METHODS AND RESULTS: We compared the 10-year Framingham risk index, histologic coronary calcification, and culprit plaque morphology in 79 consecutive adults with sudden cardiac death. There was a modest relationship between the Framingham risk index and the extent of histologic coronary calcification (r=0.35, P=0.002). Agreement in risk classification between the histologic calcification score and the Framingham risk index occurred in 50 of 79 cases (63.3%, P=0. 039). Either a focus of coronary artery calcification >/=40 micromol/L (62% of cases) or a Framingham risk index score >/= average risk for age (62% of cases) were present in 66 of 79 (83.5%) cases. Cases with plaque erosion (n=22) had significantly less coronary calcification (P=0.003) and lower Framingham risk index (P=0.001) scores than stable (n=27) or ruptured (n=30) plaques. Fourteen of 22 (63.6%) cases of plaque erosion were classified as low risk by both the Framingham risk index and the histologic calcification score. CONCLUSIONS: The prediction of sudden cardiac death using the Framingham risk index and the measurement of coronary calcification are distinct methods of assessing risk for sudden cardiac death. Excessive reliance on either method alone will produce errors in risk classification, particularly for patients at risk of plaque erosion, but their combination may be complementary. PMID- 10725283 TI - Radiolabeled native low-density lipoprotein injected into patients with carotid stenosis accumulates in macrophages of atherosclerotic plaque : effect of vitamin E supplementation. AB - BACKGROUND: Accumulation of LDL within the arterial wall appears to play a crucial role in the initiation and progression of atherosclerotic plaque. The dynamic sequence of this event has not been fully elucidated in humans. METHODS AND RESULTS: In 7 patients with previous transient ischemic attack or stroke and critical (>70%) carotid stenosis, autologous native [(125)I]-labeled LDL or [(125)I]-labeled human serum albumin were injected 24 to 72 hours before endarterectomy. Carotid specimens obtained at endarterectomy were analyzed by autoradiography and immunohistochemistry. Autoradiographic study showed that LDL was localized prevalently in the foam cells of atherosclerotic plaques, whereas the accumulation in the lipid core was negligible. Immunohistochemistry revealed that foam cells that had accumulated radiolabeled LDL were mostly CD68 positive, whereas a small number were alpha-actin positive. No accumulation of the radiotracer was detected in atherosclerotic plaques after injection of radiolabeled human serum albumin. In 3 patients treated for 4 weeks with vitamin E (900 mg/d), an almost complete suppression of radiolabeled LDL uptake by macrophages was observed. CONCLUSIONS: This study shows that circulating LDL rapidly accumulates in human atherosclerotic plaque. The prevalent accumulation of LDL by macrophages provides strong support to the hypothesis that these cells play a crucial role in the pathogenesis of atherosclerosis. PMID- 10725284 TI - Enoximone echocardiography for predicting recovery of left ventricular dysfunction after revascularization : a novel test for detecting myocardial viability. AB - BACKGROUND: The possibility that enoximone, a nonglycoside, noncatechol, positive inotropic agent, in combination with 2-dimensional echocardiography may predict recovery of myocardial dysfunction after revascularization has not been yet evaluated. METHODS AND RESULTS: Forty-five patients with chronic coronary artery disease and left ventricular dysfunction underwent dobutamine (DE, 5 to 10 microg. kg(-1). min(-1)) and enoximone (EE, 1.5 mg/kg, over 10 minutes) echocardiography. Myocardial wall motion was scored from 1 (normal) to 4 (dyskinesia): an asynergic segment was considered to have contractile enhancement when the score decreased by >/=1 grade. Of 478 asynergic segments, 216 (45%) exhibited functional recovery after revascularization. Dobutamine- and enoximone induced contractile enhancement was observed in 41% and 46% of segments, respectively. Compared with DE, EE had higher sensitivity (88% versus 79%, P<0.01) and negative predictive value (90% versus 84%, P<0.05) in predicting functional recovery. The specificity (89% versus 90%) and positive predictive value (87% for both EE and DE) were similar. Concordant interpretation of EE and DE findings was found in 85% (406 of 478) of affected segments. Prerevascularization coronary angiography showed that stenosis severity of vessels supplying areas which only improved with enoximone was significantly greater (89.9%) than that of vessels (77.7%) supplying areas that responded to both agents (P<0.02). Both dobutamine and enoximone increased heart rate (16% and 10%, respectively), whereas enoximone did not cause changes in systolic blood pressure that increased by 14% with dobutamine. CONCLUSIONS: Enoximone echocardiography provides a novel and reliable approach for the prediction of functional recovery after revascularization. Compared with dobutamine echocardiography, the test yields higher sensitivity and induces lesser hemodynamic alterations. PMID- 10725285 TI - Effect of native and oxidized low-density lipoprotein on endothelial nitric oxide and superoxide production : key role of L-arginine availability. AB - BACKGROUND: Native and oxidized LDLs (n-LDL and ox-LDL) are involved in the atherogenic process and affect endothelium-dependent vascular tone through their interaction with nitric oxide (NO). METHODS AND RESULTS: In this study we evaluated directly, by using a porphyrinic microsensor, the effect of increasing lipoprotein concentrations on endothelial NO and superoxide (O(2)(-)) production. We investigated where lipoproteins may affect the L-arginine-NO pathway by pretreating cells with L-arginine, L-N-arginine methyl ester (L-NAME), and superoxide dismutase. Bovine aortic endothelial cells were exposed for 1 hour to increasing concentrations of n-LDL (from 0 to 240 mg cholesterol/dL) and ox-LDL (from 0 to 140 mg cholesterol/dL). A stimulated (calcium ionophore) NO concentration decreased to 29% of the control at n-LDL concentration of 80 mg cholesterol/dL and to 15% of the control at 20 mg cholesterol/dL of ox-LDL. L Arginine partially neutralized the inhibitory effect of n-LDL and ox-LDL on the NO generation. Superoxide dismutase pretreatment did not modify NO production, whereas L-NAME blunted NO generation at all LDL concentrations. O(2)(-) production was increased at low n-LDL and very low ox-LDL concentrations; this was reversed by L-arginine. CONCLUSIONS: These findings confirm the inhibitory role of n-LDL and ox-LDL on NO generation and suggest that lipoproteins may induce a decreased uptake of L-arginine. The local depletion of the L-arginine substrate may derange the NO synthase, leading to overproduction of O(2)(-) from oxygen, the other substrate of NO synthase. PMID- 10725286 TI - Ambient pollution and heart rate variability. AB - BACKGROUND: We investigated associations between ambient pollution levels and cardiovascular function in a repeated measures study including 163 observations on twenty-one 53- to 87-year-old active Boston residents observed up to 12 times from June to September 1997. Particles with aerodynamic diameter 1.5 mV; this measurement was based on sinus rhythm maps in 6 patients who did not have structural heart disease. Radiofrequency point lesions extended linearly from the "dense scar," which had a voltage amplitude <0.5 mV, to anatomic boundaries or normal endocardium. To limit radiofrequency applications, 12-lead ECG during VT and pacemapping guided placement of linear lesions. No new antiarrhythmic drug therapy was added. The amount of endocardium demonstrating an abnormal electrogram amplitude ranged from 25 to 127 cm(2). A total of 8 to 87 radiofrequency lesions (mean, 55) produced a median of 4 linear lesions that had an average length of 3.9 cm (range, 1.4 to 9. 4 cm). Twelve patients (75%) have been free of VT during 3 to 36 months of follow-up (median, 8 months); 4 patients had VT episodes at 1, 3, 9, and 13 months, respectively. Only one of these patient had frequent VT. CONCLUSIONS: Radiofrequency linear endocardial lesions extending from the dense scar to the normal myocardium or anatomic boundary seem effective in controlling unmappable VT. PMID- 10725290 TI - Canadian implantable defibrillator study (CIDS) : a randomized trial of the implantable cardioverter defibrillator against amiodarone. AB - BACKGROUND: Patients surviving ventricular fibrillation (VF) or sustained ventricular tachycardia (VT) are at a high risk of death due to a recurrence of arrhythmia. The implantable cardioverter defibrillator (ICD) terminates VT or VF, but it is not known whether this device prolongs life in these patients compared with medical therapy with amiodarone. METHODS AND RESULTS: A total of 659 patients with resuscitated VF or VT or with unmonitored syncope were randomly assigned to treatment with the ICD or with amiodarone. The primary outcome measure was all-cause mortality, and the secondary outcome was arrhythmic death. A total of 328 patients were randomized to receive an ICD. A thoracotomy was done in 33, no ICD was implanted in 18, and the rest had a nonthoracotomy ICD. All 331 patients randomized to amiodarone received it initially. At 5 years, 85.4% of patients assigned to amiodarone were still receiving it at a mean dose of 255 mg/day, 28.1% of ICD patients were also receiving amiodarone, and 21.4% of amiodarone patients had received an ICD. A nonsignificant reduction in the risk of death was observed with the ICD, from 10.2% per year to 8.3% per year (19.7% relative risk reduction; 95% confidence interval, -7.7% to 40%; P=0.142). A nonsignificant reduction in the risk of arrhythmic death was observed, from 4.5% per year to 3.0% per year (32.8% relative risk reduction; 95% confidence interval, -7.2% to 57.8%; P=0.094). CONCLUSIONS: A 20% relative risk reduction occurred in all-cause mortality and a 33% reduction occurred in arrhythmic mortality with ICD therapy compared with amiodarone; this reduction did not reach statistical significance. PMID- 10725291 TI - Profound inhibition of myogenic tone in rat cardiac allografts is due to eNOS- and iNOS-based nitric oxide and an intrinsic defect in vascular smooth muscle contraction. AB - BACKGROUND: The physiological consequences of inducible NO synthase (iNOS) expression were studied in allograft coronary arteries by pressure myography. METHODS AND RESULTS: Septal coronary arteries (diameter, 200.6+/-3.3 microm) were harvested from allograft and isograft hearts, and their myogenic properties were measured before and after iNOS and nonselective NOS inhibition with aminoguanidine (AG, 100 micromol/L) and N(G)-nitro-L-arginine methyl ester (L NAME) (200 micromol/L). Fura 2 fluorescence microscopy was used to measure [Ca(2+)](i) in isolated endothelial cells. Monoclonal anti-iNOS immunostains demonstrated iNOS protein in day 2, 7, 14, and 28 allograft vessels, but only in day 2 isograft vessels. Myogenic tone was profoundly inhibited in allograft vessels from day 4 onward. In day 4 allograft vessels, these differences were abolished by L-NAME but not AG, suggesting greater basal release of eNOS-based NO from allograft endothelium. Fluorescence measurements confirmed elevation of [Ca(2+)](i) in day 4 allograft endothelium, providing a mechanism for enhanced eNOS activity. For days 7 to 28, AG potentiated myogenic tone in allograft but not isograft vessels, indicating that vasoactive iNOS-based NO was present. In mature vessels, constriction via agonist- and depolarization-mediated mechanisms showed parallel inhibition, suggesting an intrinsic defect in vascular smooth muscle cell contraction. CONCLUSIONS: Our data indicate that the profound inhibition of myogenic tone in allograft arteries involves direct vasodilation by eNOS- and iNOS-based NO, as well as an intrinsic defect in vascular smooth muscle contraction. The hemodynamic profile resulting from these changes in allograft resistance vessel function would favor movement of extracellular fluid from the intravascular space into the myocardial interstitium, resulting in edema, increased ventricular stiffness, and poor ventricular performance. PMID- 10725293 TI - Inhibition of myosin phosphatase by upregulated rho-kinase plays a key role for coronary artery spasm in a porcine model with interleukin-1beta. AB - BACKGROUND: We recently demonstrated that the Rho-kinase-mediated pathway plays an important role for coronary artery spasm in our porcine model with interleukin 1beta (IL-1beta). In this study, we examined whether or not Rho-kinase is upregulated at the spastic site and if so, how it induces vascular smooth muscle hypercontraction. METHODS AND RESULTS: Segments of the left porcine coronary artery were chronically treated from the adventitia with IL-1beta-bound microbeads. Two weeks after the operation, as reported previously, intracoronary serotonin repeatedly induced coronary hypercontractions at the IL-1beta-treated site both in vivo and in vitro, which were markedly inhibited by Y-27632, one of the specific inhibitors of Rho-kinase. Reverse transcription-polymerase chain reaction analysis demonstrated that the expression of Rho-kinase mRNA was significantly increased in the spastic compared with the control segment. Western blot analysis showed that during the serotonin-induced contractions, the extent of phosphorylation of the myosin-binding subunit of myosin phosphatase (MBS), one of the major substrates of Rho-kinase, was significantly greater in the spastic than in the control segment and that the increase in MBS phosphorylations was also markedly inhibited by Y-27632. There was a highly significant correlation between the extent of MBS phosphorylations and that of contractions. CONCLUSIONS: These results indicate that Rho-kinase is upregulated at the spastic site and plays a key role in inducing vascular smooth muscle hypercontraction by inhibiting myosin phosphatase through the phosphorylation of MBS in our porcine model. PMID- 10725292 TI - Expression and function of PPARgamma in rat and human vascular smooth muscle cells. AB - BACKGROUND: Peroxisome proliferator-activated receptor-gamma (PPARgamma) is activated by fatty acids, eicosanoids, and insulin-sensitizing thiazolidinediones (TZDs). The TZD troglitazone (TRO) inhibits vascular smooth muscle cell (VSMC) proliferation and migration in vitro and in postinjury intimal hyperplasia. METHODS AND RESULTS: Rat and human VSMCs express mRNA and nuclear receptors for PPARgamma1. Three PPARgamma ligands, the TZDs TRO and rosiglitazone and the prostanoid 15-deoxy-Delta(12,14)-prostaglandin J2 (15d-PGJ2), all inhibited VSMC proliferation and migration. PPARgamma is upregulated in rat neointima at 7 days and 14 days after balloon injury and is also present in early human atheroma and precursor lesions. CONCLUSIONS: Pharmacological activation of PPARgamma expressed in VSMCs inhibits their proliferation and migration, potentially limiting restenosis and atherosclerosis. These receptors are upregulated during vascular injury. PMID- 10725294 TI - Ventricular defibrillation with triphasic waveforms. AB - BACKGROUND: It has been reported that triphasic defibrillation waveforms cause less myocardial injury than biphasic waveforms. This study compared the defibrillation thresholds (DFTs) of triphasic and biphasic waveforms. METHODS AND RESULTS: ++DFTs were determined for a transvenous lead system and a 300-microF capacitor defibrillator. In 8 pigs (group 1), DFTs were determined for 5 triphasic waveforms with tilts of 80%, 83%, and 86% and for 1 biphasic waveform. DFTs were determined in another 8 pigs (group 2) for 2 triphasic and 4 biphasic waveforms with tilts of 43%, 49%, and 56%. In both groups, a biphasic waveform from a 140-microF-capacitor defibrillator was also evaluated, and both shock polarities were tested for each waveform. In group 1, with the 300-microF capacitor defibrillator, the leading-edge voltage and energy stored at DFT were significantly lower for triphasic waveforms with phase-duration ratios of 50/33/17 and an anode at the right ventricular electrode for phase 1 than for biphasic waveforms (P<0.001). In group 2, the stored energy of triphasic waveforms with 56% and 49% tilt was significantly lower than that of biphasic waveforms with the same tilts for anodal but not cathodal phase 1 at the right ventricular electrode. Electrode polarity significantly affected the DFT of triphasic waveforms for both studies. CONCLUSIONS: Some 80% tilt triphasic waveforms defibrillate more efficiently than biphasic waveforms with a 300-microF capacitor defibrillator. The triphasic waveforms for both groups were not superior to 140-microF-capacitor biphasic waveforms. The efficacy of triphasic waveforms depends on phase durations and electrode polarity. PMID- 10725295 TI - Influence of postshock epicardial activation patterns on initiation of ventricular fibrillation by upper limit of vulnerability shocks. AB - BACKGROUND: Shocks of identical strength and timing sometimes induce ventricular fibrillation (VFI) and other times do not (NoVFI). To investigate this probabilistic behavior, a shock strength near the upper limit of vulnerability, ULV(50), was delivered to yield equal numbers of VFI and NoVFI episodes. METHODS AND RESULTS: In 6 pigs, a 504-electrode sock was pulled over the ventricles. ULV(50) was determined by scanning the T wave. S(1) pacing was from the right ventricular apex. Ten S(2) shocks of approximate ULV(50) strength were delivered at the same S(1)-S(2) coupling interval. Intercycle interval (ICI) and wave front conduction time (WCT) were determined for the first 5 postshock cycles. ICI and the WCT of cycle 1 were not different for VFI versus NoVFI episodes (P=0.3). Beginning at cycle 2, ICI was shorter and WCT was longer for VFI than NoVFI episodes (P<0.05). CONCLUSIONS: The first cycle after shocks of the same strength (ULV(50)) delivered at the same time has the same activation pattern regardless of shock outcome. During successive cycles, however, a progressive decrease in ICI and increase in WCT occur during VFI but not NoVFI episodes. These findings suggest shock outcome is (1) deterministic but exquisitely sensitive to differences in electrophysiological state at the time of the shock that are too small to detect or (2) probabilistic and not determined until after the first postshock cycle. PMID- 10725296 TI - Pacing after shocks stronger than the upper limit of vulnerability: impact on fibrillation induction. AB - BACKGROUND: After upper-limit-of-vulnerability (ULV) shocks of the same strength and coupling interval (CI) during the T wave, (1) the epicardial activation pattern (EAP) for the first postshock cycle is indistinguishable between shocks that do (VF) and do not (NoVF) induce ventricular fibrillation (VF) and (2) >/=3 cycles in rapid succession always occur during VF but not during NoVF episodes. To study the role of these rapid cycles, rapid pacing was performed after a shock stronger than the ULV that by itself did not induce rapid cycles and VF. METHODS AND RESULTS: A 504-electrode sock was sutured to the heart in 6 pigs to map EAPs. The S2 shock strength and S1-S2 CI at the ULV were determined by T-wave scanning with an up/down protocol. Ten shocks 50 to 100 V above the ULV (aULV) were delivered at the same S1-S2 CI to confirm that VF was not induced. Then, the postshock interval after aULV shocks was scanned with an S3 pacing stimulus from the LV apex until the shortest S2-S3 CI that captured was reached. This was repeated for S4, S5, etc, until VF was induced. To induce VF, 3 pacing stimuli (S3-S5) with progressively shorter CIs were required; S3 or S3, S4 never induced VF. After cycle S5, which induced VF, 2 EAP types occurred: focal (74%) and reentrant (26%). CONCLUSIONS: At least 3 cycles with short CIs are necessary for VF induction after aULV shocks. Cycles S3-S4 may create the substrate for cycle S5 to initiate VF. PMID- 10725297 TI - Coronary physiology revisited : practical insights from the cardiac catheterization laboratory. AB - Various coronary physiological measurements can be made in the cardiac catheterization laboratory using sensor-tipped guidewires; they include the measurement of poststenotic absolute coronary flow reserve, the relative coronary flow reserve, and the pressure-derived fractional flow reserve of the myocardium. Ambiguity regarding abnormal microcirculation has been reduced or eliminated with measurements of relative coronary flow reserve and fractional flow reserve. The role of microvascular flow impairment can be separately determined with coronary flow velocity reserve measurements. In addition to lesion assessment before and after intervention, emerging applications of coronary physiology include the determination of physiological responses to new pharmacological agents, such as glycoprotein IIb/IIIa blockers, in patients with acute myocardial infarction. Measurements of coronary physiology in the catheterization laboratory provide objective data that complement angiography for clinical decision-making. PMID- 10725298 TI - Cardiac echinococcosis. PMID- 10725299 TI - Arrhythmogenic right ventricular dysplasia/cardiomyopathy: need for an international registry. Study Group on Arrhythmogenic Right Ventricular Dysplasia/Cardiomyopathy of the Working Groups on Myocardial and Pericardial Disease and Arrhythmias of the European Society of Cardiology and of the Scientific Council on Cardiomyopathies of the World Heart Federation. AB - Arrhythmogenic right ventricular (RV) dysplasia/cardiomyopathy (ARVD/C) is a heart muscle disease characterized by peculiar RV involvement and electrical instability that precipitates ventricular arrhythmias and sudden death. The purpose of the present consensus report of the Study Group on ARVD/C of the Working Groups on Myocardial and Pericardial Disease and Arrhythmias of the European Society of Cardiology and of the Scientific Council on Cardiomyopathies of the World Heart Federation is to review the considerable progress in our understanding of the etiopathogenesis, morbid anatomy, and clinical presentation of ARVD/C since it first was described in 1977. The present article focuses on important but still unanswered issues, mostly regarding risk stratification, clinical outcome, and management of affected patients. Because ARVD/C is relatively uncommon and any one center may have experience with only a few patients, an international registry is being established to accumulate information and enhance the numbers of patients that can be analyzed and thus answer pending questions. The registry also will facilitate pathological, molecular, and genetics research on the causes and pathogenesis of the ARVD/C. Furthermore, availability of an international database will enhance awareness of this largely unrecognized condition among the medical community. Physicians are encouraged to enroll patients in the International Registry of ARVD/C. PMID- 10725300 TI - Prinzmetal's angina. PMID- 10725301 TI - N-Methylprotoporphyrin is a more potent inhibitor of recombinant human than of recombinant chicken ferrochelatase. AB - The potency of N-methylprotoporphyrin IX (N-methylPP) as a ferrochelatase (FC) inhibitor has been previously studied using crude chick embryo liver FC preparations. However, interactions between N-methylprotoporphyrin IX (N methylPP) and impurities in the enzyme preparation may have compromised the results. The first objective of this study was to compare the potency of N methylPP as an inhibitor of purified chicken FC and crude chick embryo liver FC. The EC(50) values of N-methylPP previously observed in crude chick embryo liver FC was 2.9 x 10(-3) nmol/mg protein, and with purified recombinant chicken FC was 2.07 x 10(-3) nmol/mg protein. The difference in EC(50) values was not statistically significant, and we conclude that interactions between N-methylPP and impurities in crude enzyme preparations did not affect the estimation of potency of N-methylPP. The second objective of this study was to compare the potency of N-methylPP between purified human and chicken FC. The EC(50) value of N-methylPP observed in the purified human FC preparation was 1.7 x 10(-6) nmol/mg protein (chicken FC 2.07 x 10(-3) nmol/mg protein). Thus, the potency of N methylPP was much higher with purified human FC than with purified chicken FC. Because the porphyrinogenicity of several xenobiotics involves N alkylprotoporphyrin IX formation, results on drug-induced porphyria obtained with avian species may underestimate the potential porphyrinogenicity in humans. PMID- 10725302 TI - Species differences in the regio- and stereoselectivity of 1-nitronaphthalene metabolism. AB - 1-Nitronaphthalene (1-NN) is a mutagenic nitroaromatic that has been detected in emissions from both heavy- and light-duty diesel engines, as well as in urban airborne particles. 1-NN is a cytochrome P450-bioactivated, nonciliated bronchiolar epithelial (Clara) cell cytotoxicant. Our recent studies demonstrated that 1-NN was metabolized by rat lung and liver microsomal enzymes to six 1-NN GSH conjugates via intermediate C(5),C(6)- and C(7),C(8)-epoxides. These studies examined the metabolism of 1-NN in mouse, and compared the differences in rates of 1-NN GSH conjugate formation between the two species. HPLC radioactivity profiles demonstrated that seven different conjugates were generated in mouse lung and liver microsomal incubations. Six of the seven conjugates corresponded with those observed in incubations with rat microsomes. Mass spectrometry of the new conjugate yielded a m/z 497 (M+H) and identical daughter ions as in the other six conjugates when analyzed by mass spectrometry in electrospray positive ion mode. The major conjugate generated in mouse and rat lung microsomal incubations was conjugate 4 (1-nitro-7-glutathionyl-8-hydroxy-7, 8-dihydronaphthalene). In comparison, the formation of conjugate 6 (1-nitro-5-hydroxy-6-glutathionyl-5,6 dihydronaphthalene) predominated in mouse liver, whereas in rat liver, conjugate 5, a diastereomer of conjugate 6, was generated at the highest rate. We concluded that the rates of formation of regio- and stereoisomeric epoxides from 1-NN differed substantially in target and nontarget tissues, but there was no clear pattern of correlation of tissue susceptibility to the rate or metabolite produced. PMID- 10725303 TI - Human cytochrome P450 maximal activities in pediatric versus adult liver. AB - Drug clearance is often higher in children than in adults, particularly when normalized to body weight. We previously showed that liver volume normalized to body weight was inversely related to age, but that the systemic clearance of a nonspecific cytochrome P450 (CYP) substrate (antipyrine) was higher in young children compared with adults even when normalized per liver volume. Our purpose herein was to evaluate whether P450 catalytic activities, expressed as maximal catalytic rates per milligram of microsomal protein, differed in up to 37 normal livers from subjects <10 (range 0.5-9 years of age), >10 but <60 years of age (range 10-59 years), and >60 year (range 63-93 years of age). There were no age related differences in the oxidation of ethoxyresorufin (P =.83) (CYP1A2), ethoxycoumarin (P =.52) (CYP2E1 and other P450s), teniposide (P =. 58), midazolam (P =.47) (CYP3A4/3A5), or paclitaxel (P =.24) (at the 17alpha position, CYP2C8). Tolbutamide hydroxylation tended to be lower in children versus adults (P =.047) (CYP2C9), but did not reach statistical significance after correcting for multiple comparisons. No relationship was found to exist between age and microsomal recovery (P =.98); thus, recovery did not account for the lack of age related differences in catalytic activity. We conclude that increased intrinsic cytochrome P450 activity is unlikely to account for increased clearance of most P450 drug substrates in children. PMID- 10725305 TI - Pharmacokinetics of a model organic nitrite inhalant and its alcohol metabolite in rats. AB - Volatile organic nitrites were originally used to relieve the chest pain that is associated with angina pectoris. Today, these inhalants are predominantly used as drugs of abuse. Little is known regarding the bioavailability and disposition of volatile nitrites. In this study, the pharmacokinetics of a major organic nitrite inhalant, isobutyl nitrite (ISBN), and its primary metabolite, isobutyl alcohol (ISBA), were investigated after inhalation and i.v. administration. ISBN blood concentrations in the rat declined mono-exponentially with a half-life of 1.4 min and a blood clearance of 2.9 l/min/kg that vastly exceeded cardiac output (0.3 l/min/kg). Approximately 98% of ISBN was metabolized to ISBA, which declined monoexponentially with a half-life of 5.3 min when the infusion of ISBN was terminated. The bioavailability of inhaled ISBN, over a range of 300 to 900 ppm, was estimated to be 43%. After inhaled ISBN, the half-life of ISBA decreased approximately 4-fold (t(1/2) inh = 1.5 min versus t(1/2) i.v. = 5.3 min; P <.001), whereas no pharmacokinetic difference was observed for ISBN. Inhalation of another nitrite, isoamyl nitrite, accelerated the apparent clearance of ISBA, suggesting that nitrite inhalation could change the disposition of another compound. A pharmacokinetic model was developed to describe the concentration time profile of ISBA and ISBN after inhalation and i.v. administration. PMID- 10725304 TI - Oxidative metabolism of bupivacaine into pipecolylxylidine in humans is mainly catalyzed by CYP3A. AB - Bupivacaine is used to provide prolonged anesthesia and postoperative analgesia. The human cytochrome P450 (CYP) involved in bupivacaine degradation into pipecolylxylidine (PPX), its major metabolite, has, to our knowledge, never been described. Microsome samples were prepared from six human livers and incubated in the presence of bupivacaine. The concentrations of PPX in the microsomal suspensions were assessed, and K(m) and V(max) values were calculated. Bupivacaine incubations were then performed with specific CYP substrates and inhibitors. For each sample of hepatic microsomes, the correlation between the rate of PPX formation and the corresponding erythromycin N-demethylase activity was analyzed. Finally, an immunoinhibition study using an anti-rabbit CYP3A6 antibody and assays with cDNA-expressed human CYP were conducted. The apparent K(m) and V(max) values of bupivacaine were, respectively, 125 microM and 4.78 nmol/min/mg of microsomal protein. The strongest inhibition of bupivacaine metabolism was obtained for troleandomycin (-95% at 50 microM), a specific CYP3A inhibitor. The correlation between PPX formation and erythromycin N-demethylase activity showed an R value of 0.99 whereas anti-rabbit CYP3A6 antibody inhibited the degradation of bupivacaine into PPX by 99%. Finally, CYP1A2 and CYP2E1 cDNA expressed forms of human CYP did not allow PPX formation, CYP2C19 and CYP2D6 produced only small amounts whereas CYP3A4 most efficiently metabolized bupivacaine into PPX. These results demonstrated that bupivacaine degradation into PPX was mediated in humans by CYP3A. PMID- 10725306 TI - In vitro biotransformation of sildenafil (Viagra): identification of human cytochromes and potential drug interactions. AB - The in vitro biotransformation of sildenafil to its major circulating metabolite, UK-103,320, was studied in human liver microsomes and in microsomes containing heterologously expressed human cytochromes. In human liver microsomes, the mean K(m) (+/-S.E. ) was 14.4 +/- 2.0 microM. A screen of the chemical inhibitors omeprazole (10 microM), quinidine (10 microM), sulfaphenazole (10 microM), and ketoconazole (2.5 microM) only revealed detectable inhibition with ketoconazole. Sildenafil biotransformation (36 microM) was inhibited by increasing concentrations of ketoconazole and ritonavir (IC(50) values less than 0.02 microM), which are established cytochrome P450 (CYP) 3A4 inhibitors. Using microsomes containing cDNA-expressed cytochromes, UK-103,320 formation was found to be mediated by four cytochromes: CYP3A4, -2C9, -2C19, and -2D6. Estimated relative contributions to net intrinsic clearance were 79% for CYP3A4 and 20% for CYP2C9; for CYP2C19 and -2D6, estimated contributions were less than 2%. These results demonstrate that CYP3A4 is the primary cytochrome mediating UK-103,320 formation and that drugs that inhibit CYP3A4 are likely to impair sildenafil biotransformation. PMID- 10725307 TI - Thioesterification of 2-arylpropionic acids by recombinant acyl-coenzyme A synthetases (ACS1 and ACS2). AB - 2-Arylpropionic acids are a class of frequently used nonsteroidal anti inflammatory drugs exhibiting a potent inhibition of cyclooxygenase isoforms supported by the (+)S-enantiomer alone. Nevertheless, some of these compounds in the (-)R configuration may undergo extensive inversion of configuration to their antipode. The key molecular basis for this mechanism invokes the stereoselective formation of the coenzyme A (CoA) thioester of the 2-arylpropionic acid by long chain acyl-CoA synthetases (ACSs). In this report, rat recombinant ACS1 and ACS2 enzymes, constitutively highly expressed in adult rat liver and brain, respectively, have been overproduced in Escherichia coli strains and purified to homogeneity to investigate the involvement of these enzymes in the thioesterification of fenoprofen and ibuprofen. Recombinant ACS1 efficiently catalyzed both nonsteroidal anti-inflammatory drugs with Michaelis-Menten parameters of K(M) = 1686 +/- 93 microM, V(max) = 353 +/- 45 nmol/min/mg protein for (-)R-ibuprofen and K(M) = 103 +/- 12 microM, V(max) = 267 +/- 10 nmol/min/mg protein for (-)R-fenoprofen, and exhibited a marked stereoselectivity in favor of the (-)R-enantiomer. Recombinant ACS2, a closely related sequence with ACS1, exhibited a lower enzymatic efficacy from 7- to 130-fold for (-)R-ibuprofen and ( )R-fenoprofen, respectively. On the basis of these findings and considering the level of tissue expression of the different long-chain ACSs, ACS1 appears to be the major enzyme involved in the first step of the chiral inversion of 2 arylpropionic acids. Nevertheless, the participation of other ACS isoforms of minor quantitative importance could not be excluded in the thioesterification of xenobiotics. PMID- 10725308 TI - Pharmacokinetics, biliary excretion, and tissue distribution of novel anti-HIV agents, cosalane and dihydrocosalane, in Sprague-Dawley rats. AB - Cosalane and dihydrocosalane are potent inhibitors of HIV replication with a broad range of activity. The purpose of this study was to investigate: 1) the pharmacokinetic disposition of both cosalane and dihydrocosalane in male Sprague Dawley rats, and 2) biliary excretion, enterohepatic circulation, and tissue distribution of cosalane after i.v. and/or oral administration. Animals were administered i.v. (10 mg/kg) cosalane or dihydrocosalane through a jugular vein to obtain plasma profiles. Dose dependence of cosalane was studied over a dose range of 1.0 to 10 mg/kg. The extent of enterohepatic recycling, biliary excretion, and tissue distribution were studied after i.v. administration. Both cosalane and dihydrocosalane exhibited a biexponential disposition with very long half-lives of 749 +/- 216 and 1016 +/- 407 min, along with very large volumes of distribution 23.1 +/- 4.4 and 24.4 +/- 2. 5 liter/kg, respectively. Both cosalane (nondetectable) and dihydrocosalane (<1%) showed very poor oral bioavailability. The biliary and renal excretions of cosalane were found to be negligible with no detectable metabolites either in urine or bile. After oral administration, more than 87% of the cosalane dose was excreted in the feces as the parent compound. Also, cosalane was sequestered significantly in liver with quantifiable levels in all tissues tested, even 48 h after the dose was administered. Therefore it was concluded that the poor oral bioavailability of cosalane may be due to its poor enterocytic transport coupled with sequestration in liver parenchymal cell membrane layers. PMID- 10725309 TI - Studies on the cytochrome P450 (CYP)-mediated metabolic properties of miocamycin: evaluation of the possibility of a metabolic intermediate complex formation with CYP, and identification of the human CYP isoforms. AB - Some macrolide antibiotics cause clinical drug interactions, resulting in altered metabolism of concomitantly administered drugs, via the formation of a metabolic intermediate (MI) complex with cytochrome P450 (CYP), or competitive inhibition of CYP. In this study, the possibility of MI complex formation by miocamycin (MOM) was assessed first. CYP contents and activities in rat liver microsomes were not affected and there were no detectable MI complexes after administration of MOM for either 3 or 10 days to rats. Furthermore, MOM did not form MI complexes in vitro even with microsomes from humans or dexamethasone-pretreated rats. Second, in vitro studies were conducted to identify the human CYP isoforms involved in four 14-hydroxylation reactions in the MOM metabolic pathway. The results showed that it was most likely CYP3A4 involved in the hydroxylations: 1) each hydroxylation in human liver microsomes from 10 different donors strongly correlated with testosterone 6 beta-hydroxylation; 2) each hydroxylation was essentially inhibited by ketoconazole and troleandomycin; 3) only cDNA-expressed CYP3A4 and CYP3A5 catalyzed the hydroxylations, and the activities of CYP3A5 were below 5% of those of CYP3A4; and 4) the apparent K(M) values obtained with native human liver microsomes were comparable with those obtained with cDNA-expressed CYP3A4. In conclusion, MOM is not an inhibitor of CYP via the formation of an MI complex. Moreover, CYP3A4 is mainly responsible for catalyzing the hydroxylation of MOM metabolites. Because CYP3A4 is the most abundant form of CYP in the liver and intestine, this isoform probably accounts for the majority of drug-MOM interactions observed in clinical practice. PMID- 10725310 TI - Disposition in rats of N-pyridinium-propyl-cyclam, N-triethylammonium-propyl cyclam, and N-[Triethylammonium]-3-propyl-[15]ane-N5, potential cartilage imaging agents. AB - Quaternary ammonium compounds are known to highly concentrate in articular cartilages after i.v. administration. This property was used to synthesize new potential radiodiagnostic agents for joint imaging. Pharmacokinetic study was performed in rats for three new compounds: N-pyridinium-propyl-cyclam (NPPC), N triethylammonium-propyl-cyclam (NTPC), and N-[triethylammonium]-3-propyl-[15]ane N5 (NTP 15-5). After i.v. administration, [(3)H]NPPC and [(3)H]NTPC highly and rapidly concentrated in articular cartilage, this uptake being followed by a single exponential decrease with half-lives of, respectively, 75 and 82 min. Except cartilage, only the kidney was highly labeled. After complexation of (99m)Tc by NPPC, NTPC, and NTP 15-5, only (99m)Tc-NTP 15-5 exhibited a high affinity for cartilage. On the other hand, the pharmacokinetic behavior of (99m)Tc-NTPC and (99m)Tc-NPPC was very different from those of their (3)H-labeled analogs. Concentration in cartilaginous tissues was strongly diminished, and liver and bone were highly labeled. For all labeled species, the major route of excretion was urine, and HPLC analysis showed that [(3)H]NTPC and [(3)H]NPPC were excreted under their unchanged form. On the other hand, no (99m)Tc-NTPC and (99m)Tc-NPPC were found in the urine, the radioactivity being mainly due to free technetium, contrary to (99m)Tc-NTP 15-5, which was excreted in the urine under the complexed form. These data can explain the striking differences observed between the three (99m)Tc-labeled molecules, the lack of concentration of (99m)Tc NTPC, and (99m)Tc-NPPC in cartilages in comparison with their (3)H-labeled analogs due to an instability in vivo of these technetiated complexes. PMID- 10725311 TI - Pharmacokinetics, metabolism, and excretion of irinotecan (CPT-11) following I.V. infusion of [(14)C]CPT-11 in cancer patients. AB - This study determined the disposition of irinotecan hydrochloride trihydrate (CPT 11) after i.v. infusion of 125 mg/m(2) (100 microCi) [(14)C]CPT-11 in eight patients with solid tumors. Mean +/- S.D. recovery of radioactivity in urine and feces was 95.8 +/- 2.7% (range 92.2-100.3%, n = 7) of dose. Radioactivity in blood, plasma, urine, and feces was determined for at least 168 h after dosing. Fecal excretion accounted for 63.7 +/- 6.8 (range 54.2-74.9%, n = 7) of dose, whereas urinary excretion accounted for 32.1 +/- 6.9% (range 21.7-43.8%; n = 7) of dose. One patient with a biliary T-tube excreted 30.1% of dose in bile, 14.2% in feces, and 48.2% in urine. Quantitative radiometric HPLC revealed that CPT-11 was the major excretion product in urine, bile, and feces. Aminopentane carboxylic acid (APC) and SN-38 glucuronide (SN-38G) were the most significant metabolites in urine and bile, whereas SN-38 and NPC, a primary amine metabolite, were relatively minor excretion products. SN-38 and APC were the most significant metabolites in feces. The relatively higher amount of SN-38 in feces compared with bile is presumably due to hydrolysis of SN-38G to SN-38 by enteric bacterial beta-glucuronidases. There was close correspondence between quantitative fluorescence HPLC and mass balance findings. CPT-11 was the major circulating component in plasma (55% of the mean radiochemical area under the curve), and CPT 11, SN-38, SN-38G, and APC accounted for 93% of the mean radiochemical AUC. These results show that the parent drug and its three major metabolites account for virtually all CPT-11 disposition, with fecal excretion representing the major elimination pathway. PMID- 10725312 TI - The synthesis, in vitro reactivity, and evidence for formation in humans of 5 phenyl-1,3-oxazinane-2,4-dione, a metabolite of felbamate. AB - Previously we have proposed and provided evidence for a metabolic scheme leading to 3-carbamoyl-2-phenylpropionaldehyde from the antiepileptic drug felbamate. This aldehyde was found to undergo reversible cyclization to form the more stable cyclic carbamate 4-hydroxy-5-phenyl-tetrahydro-1,3-oxazin-2-one or undergo elimination to form 2-phenylpropenal. The cyclic carbamate bears structural similarity to 4-hydroxycyclophosphamide and there is an intriguing parallelism between the pathway from the cyclic carbamate to 2-phenylpropenal and the known pathway from 4-hydroxycyclophosphamide to acrolein. The similarity of these transformations led us to consider 5-phenyl-1,3-oxazinane-2,4-dione, which could arise from an oxidation of the cyclic carbamate, as a potential metabolite of felbamate. As the formation of this dione species may have both potential pharmacologic and toxicologic implications for felbamate therapy, we wished to study its reactivity. We have developed a synthesis of 5-phenyl-1, 3-oxazinane 2,4-dione and evaluated its reactivity in vitro. This dione was found to undergo base-catalyzed decomposition to three products, one of which is the major human metabolite of felbamate, 3-carbamoyl-2-phenylpropionic acid. Furthermore, we have found evidence for the presence of the dione in human urine after felbamate treatment through the identification of its major in vitro decomposition product, 2-phenylacrylamide 11. PMID- 10725313 TI - Porcine kidney microsomal cysteine S-conjugate N-acetyltransferase-catalyzed N acetylation of haloalkene-derived cysteine S-conjugates. AB - N-Acetylation of xenobiotic-derived cysteine S-conjugates is a key step in the mercapturic acid pathway. The aim of this study was to investigate the N acetylation of haloalkene-derived S-haloalkyl and S-haloalkenyl cysteine S conjugates by porcine kidney cysteine S-conjugate N-acetyltransferase (NAcT). A radioactive assay for the quantification of NAcT activity was developed as a new method for partial purification of the enzyme, which was necessitated by the substantial loss of activity during the immunoaffinity chromatography method. 3 [(3-Cholamidopropyl)dimethylammonio]-1-propane-sulfonate, rather than N,N-bis[3 gluconamidopropyl]deoxycholamide, was used to solubilize the NAcT from porcine kidney microsomes in the revised procedure. The partially purified NAcT was free of detectable aminoacylase activity. Although low acetyl-coenzyme A hydrolase activity was observed, its effect on the assay was minimized by addition of excess acetyl-coenzyme A in the NAcT assay mixture. Attempts to separate the residual hydrolase activity from NAcT by different chromatographic procedures were either unsuccessful or lead to inactivation of NAcT. Most of the cysteine S conjugates studied were N-acetylated by NAcT. Although the apparent K(m) values for the cysteine S-conjugates studied differed by a factor of approximately 2.5 (124-302 microM), a greater than 15-fold difference in the apparent V(max) (0.75 15.6 nmol/h) and V(max)/K(m) (0.008-0.126 x 10(-3) l h(-1)) values was observed. These data show that a range of haloalkene-derived cysteine S-conjugates serve as substrates for pig kidney NAcT. The significant differences in cytotoxicity of these conjugates may be a result of more variable deacetylation rates of the corresponding mercapturates. PMID- 10725314 TI - Metabolism and disposition of moxonidine in Fischer 344 rats. AB - The metabolism and disposition of moxonidine (4-chloro-5-(imidazolidin-2 ylidenimino)-6-methoxy-2-methylp yrimidine ), a potent central-acting antihypertensive agent, were investigated in F344 rats. After an i.v. or oral administration of 0.3 mg/kg of [(14)C]moxonidine, the maximum plasma concentrations of moxonidine were determined to be 146.0 and 4.0 ng/ml, respectively, and the elimination half-lives were 0.9 and 1.1 h, respectively. The oral bioavailability of moxonidine was determined to be 5.1%. The metabolic and elimination profiles of moxonidine were determined after an oral administration of 5 mg/kg of [(14)C]moxonidine. More than fifteen phase I and phase II metabolites of moxonidine were identified in the different biological matrices (urine, plasma, and bile). Oxidative metabolism of moxonidine leads to the formation of hydroxymethyl moxonidine and a carboxylic acid metabolite as the major metabolites. Several GSH conjugates, cysteinylglycine conjugates, cysteine conjugates, and a glucuronide conjugate were also identified in rat bile samples. The radiocarbon was eliminated primarily by urinary excretion in rats, with 59.5% of total radioactivity recovered in the urine and 38.4% recovered in the feces within 120 h. In bile duct-cannulated rats, about 39.7% of the radiolabeled dose was excreted in the urine, 32.6% excreted in the bile, and approximately 2% remained in the feces. The results from a quantitative whole body autoradiography study indicate that radiocarbon associated with [(14)C]moxonidine and/or its metabolites was widely distributed to tissues, with the highest levels of radioactivity observed in the kidney and liver. In summary, moxonidine is well absorbed, extensively metabolized, widely distributed into tissues, and rapidly eliminated in rats after oral administration. PMID- 10725315 TI - Route-dependent nonlinear pharmacokinetics of a novel HIV protease inhibitor: involvement of enzyme inactivation. AB - L-754,394, a furanopyridine derivative, is an experimental HIV protease inhibitor. Previous studies from this laboratory have demonstrated that L-754,394 is cleared very rapidly in animals, and that this drug is a potent mechanism based inactivator (suicide inhibitor) for CYP3A4 in human liver microsomes. Because L-754,394 is a high-clearance drug and an enzyme inactivator, it is expected that this drug will be subject to significant first-pass metabolism, and that the degree of enzyme inactivation will be dependent not only on the dose, but also on the route of administration. The purpose of this study is to examine the effects of dose and route of administration on the kinetics of L-754,394 using rats and dogs as animal models. In both rats and dogs, L-754,394 exhibited marked dose-dependent pharmacokinetics after i.v. and oral administration. Irrespective of i.v. or oral administration, the area under the plasma concentration-time curve from zero to infinity increased with dose in a greater than proportional manner. However, the magnitude of area under the plasma concentration-time curve from zero to infinity increase was much greater after oral dosing than after i.v. administration, indicating route-dependent pharmacokinetics. Data from in vitro and in vivo studies suggested that the dose- and route-dependent pharmacokinetics were due mainly to the inactivation (destruction) of the enzymes responsible for its own metabolism. PMID- 10725316 TI - Prediction of in vivo drug-drug interactions based on mechanism-based inhibition from in vitro data: inhibition of 5-fluorouracil metabolism by (E)-5-(2 Bromovinyl)uracil. AB - The fatal drug-drug interaction between sorivudine, an antiviral drug, and 5 fluorouracil (5-FU) has been shown to be caused by a mechanism-based inhibition. In this interaction, sorivudine is converted by gut flora to (E)-5-(2 bromovinyl)uracil (BVU), which is metabolically activated by dihydropyrimidine dehydrogenase (DPD), and the activated BVU irreversibly binds to DPD itself, thereby inactivating it. In an attempt to predict this interaction in vivo from in vitro data, inhibition of 5-FU metabolism by BVU was investigated by using rat and human hepatic cytosol and human recombinant DPD. Whichever enzyme was used, increased inhibition was observed that depended on the preincubation time of BVU and enzyme in the presence of NADPH and BVU concentration. The kinetic parameters obtained for inactivation represented by k(inact) and K'(app) were 2.05 +/- 1.52 min(-1), 69.2 +/- 60.8 microM (rat hepatic cytosol), 2.39 +/- 0.13 min(-1), 48.6 +/- 11.8 microM (human hepatic cytosol), and 0.574 +/- 0.121 min(-1), 2.20 +/- 0.57 microM (human recombinant DPD). The drug-drug interaction in vivo was predicted quantitatively based on a physiologically based pharmacokinetic model, using pharmacokinetic parameters obtained from the literature and kinetic parameters for the enzyme inactivation obtained in the in vitro studies. In rats, DPD was predicted to be completely inactivated by administration of BVU and the area under the curve of 5-FU was predicted to increase 11-fold, which agreed well with the reported data. In humans, a 5-fold increase in the area under the curve of 5-FU was predicted after administration of sorivudine, 150 mg/day for 5 days. Mechanism-based inhibition of drug metabolism is supposed to be very dangerous. We propose that such in vitro studies should be carried out during the drug developing phase so that in vivo drug-drug interactions can be predicted. PMID- 10725317 TI - Prediction of in vivo drug-drug interactions between tolbutamide and various sulfonamides in humans based on in vitro experiments. AB - Drug-drug interactions between tolbutamide and sulfonamides have extensively been reported. We attempted to predict the in vivo interaction between tolbutamide and sulfonamides from the in vitro metabolic inhibition studies. The inhibition constant (K(i)) was derived from the inhibitory effects of eight sulfonamides (sulfaphenazole, sulfadiazine, sulfamethizole, sulfisoxazole, sulfamethoxazole, sulfapyridine, sulfadimethoxine, and sulfamonomethoxine) on tolbutamide metabolism. We found that the inhibitory effect of sulfaphenazole was greatest among the eight sulfonamides examined. Furthermore, the contribution of each P450 enzyme to tolbutamide metabolism was investigated by using recombinant P450 enzymes. Although cytochrome P450 (CYP) 2C8, 2C9, and 2C19 metabolized tolbutamide, the main enzyme involved was CYP2C9. The K(i) values of several sulfonamides were comparable between human liver microsomes and recombinant CYP2C9. The maximum unbound plasma concentration of sulfonamides in the portal vein was calculated from literature data on the pharmacokinetics of sulfonamides. Using the K(i) values obtained from in vitro inhibition studies, the degree of increase in tolbutamide area under the plasma concentration-time curve (AUC) was predicted. About 4.8- and 1.6-fold increases in tolbutamide AUC were predicted by coadministration of sulfaphenazole and sulfamethizole, respectively, which agreed well with the reported increases in humans. Furthermore, the increase in tolbutamide AUC by coadministration of sulfadiazine, sulfisoxazole, and sulfamethizole was predicted to be 1.5- to 2. 6-fold, although the corresponding in vivo effects have not been reported. It is concluded that some of these sulfonamides have to be carefully coadministered with CYP2C9 substrates such as tolbutamide although coadministration of sulfaphenazole needs the greatest care. PMID- 10725318 TI - Effects of long-term grapefruit juice ingestion on nifedipine pharmacokinetics: induction of rat hepatic P-450 by grapefruit juice. AB - We studied the effects of short- and long-term ingestion of grapefruit juice (GJ) on nifedipine (NFP) pharmacokinetics in rats. Thirty minutes after intraduodenal (id) administration of 2.0 ml of GJ or saline, NFP was i.v. or id administered at a dose of 3 mg/kg b. wt. No significant differences were observed in pharmacokinetic values between the two groups after i.v. administrations of NFP. By contrast, after id administration, the mean AUC value in the GJ group was approximately 1.62 times that in the control group, and the mean apparent clearance (CL) decreased by approximately 40%. In addition, 2.0 ml of GJ was orally administered twice a day (9:00 AM and 7:00 PM) for 10 consecutive days; on the 11th day the pharmacokinetics of NFP were examined again. Irrespective of route of administration (i.v. or id), NFP CL from plasma in these GJ-treated rats was considerably faster than that in the rats treated with GJ for a short (30 min) period. In microsomes prepared from the intestinal mucosa of animals receiving long-term administration of GJ, NFP oxidation activity (0.21 +/- 0.02 nmol/mg/min) and P-450 content (0.045 +/- 0.009 nmol/mg) were significantly lower than those in untreated rats (0.32 +/- 0.05 nmol/mg/min and 0.060 +/- 0.007 nmol/mg). In hepatic microsomes from the same rats, however, NFP oxidation activity (1.43 +/- 0.17 nmol/mg/min) and P-450 content (0.66 +/- 0.07 nmol/mg) were distinctly greater than those in untreated rats (1.00 +/- 0.06 nmol/mg/min and 0.51 +/- 0.04 nmol/mg). In conclusion, short-term id exposure to GJ resulted in increased NFP bioavailability, whereas long-term administration of GJ resulted in reduced bioavailability and increased CL. PMID- 10725319 TI - The N-acetylation of arsanilic acid In vitro by mammalian enzymes. AB - The N-acetylation of arsanilic acid was assayed in vitro by modifying a literature method for acetylation of p-aminobenzoic acid. Conditions included final concentrations of 1.0 mM dithiothreitol, 1.0 mM EDTA, 0.45 mM acetyl coenzyme A, an acetyl coenzyme A regenerating system using bacterial phosphotransacetylase and acetyl phosphate, 5.0 mM arsanilate substrate, and 25 mM sodium/potassium phosphate buffer, pH 7.4, in a total volume of 0.5 ml. Incubation was at 37 degrees C, with 0.5- to 2-mg N-acetyltransferase enzyme protein from a preparation of guinea pig liver. The reaction was terminated by heat precipitation. The resulting supernatant was put through a 4 mm 0.45 microm polysulfone membrane syringe filter. The filtrate could then be injected directly onto the HPLC. With arsanilic acid as substrate, the product N-acetylarsanilic acid (NAA) was identified by its retention time (33 min) in the HPLC system of the laboratory. The 33-min fraction collected from the HPLC was scanned and gave the characteristic UV spectrum of NAA, with peaks at 203 and 256 nm. In addition, the product comigrated in the HPLC system with standard NAA. Under comparable assay conditions, the N-acetylation of arsanilate by the guinea pig enzyme preparation is about 24% the rate of that of the model substrate p-aminobenzoic acid. Typical activity for arsanilate acetylation was 0.5 nmol/min/mg enzyme protein. Using the same assay system and HPLC detection method, the supernatant from bacterial lysates containing recombinant human N-acetyltransferase 1 exhibited acetylation activity toward arsanilate of 720 nmol/min/mg enzyme protein. PMID- 10725320 TI - The nucleolus and the four ribonucleoproteins of translation. PMID- 10725321 TI - Nuclear migration. From fungi to the mammalian brain. PMID- 10725322 TI - Dynamin and FtsZ. Missing links in mitochondrial and bacterial division. PMID- 10725323 TI - Reverse transcriptase activity in mature spermatozoa of mouse. AB - We show here that a reverse transcriptase (RT) activity is present in murine epididymal spermatozoa. Sperm cells incubated with human poliovirus RNA can take up exogenous RNA molecules and internalize them in nuclei. Direct PCR amplification of DNA extracted from RNA-incubated spermatozoa indicate that poliovirus RNA is reverse-transcribed in cDNA fragments. PCR analysis of two-cell embryos shows that poliovirus RNA-challenged spermatozoa transfer retrotranscribed cDNA molecules into eggs during in vitro fertilization. Finally, RT molecules can be visualized on sperm nuclear scaffolds by immunogold electron microscopy. These results, therefore, reveal a novel metabolic function in spermatozoa, which may play a role during early embryonic development. PMID- 10725324 TI - Nucleocytoplasmic shuttling of the Cdc42p exchange factor Cdc24p. AB - Cdc24p, the GDP/GTP exchange factor for the regulator of actin cytoskeleton Cdc42p, localizes to sites of polarized growth. Here we show that Cdc24p shuttles in and out of the yeast nucleus during vegetative growth. Far1p is necessary and sufficient for nuclear accumulation of Cdc24p, suggesting that its nuclear import occurs via an association with Far1p. Nuclear export is triggered either by entry into the cell cycle or by mating pheromone. As Far1p is degraded upon entry into the cell cycle, cell cycle-dependent export of Cdc24p occurs in the absence of Far1p, whereas during mating similar export kinetics indicate that a Cdc24p-Far1p complex is exported. Our results suggest that the nucleus serves as a store of preformed Cdc24p-Far1p complex which is required for chemotropism. PMID- 10725325 TI - Conformational requirements for glycoprotein reglucosylation in the endoplasmic reticulum. AB - Newly synthesized glycoproteins interact during folding and quality control in the ER with calnexin and calreticulin, two lectins specific for monoglucosylated oligosaccharides. Binding and release are regulated by two enzymes, glucosidase II and UDP-Glc:glycoprotein:glycosyltransferase (GT), which cyclically remove and reattach the essential glucose residues on the N-linked oligosaccharides. GT acts as a folding sensor in the cycle, selectively reglucosylating incompletely folded glycoproteins and promoting binding of its substrates to the lectins. To investigate how nonnative protein conformations are recognized and directed to this unique chaperone system, we analyzed the interaction of GT with a series of model substrates with well defined conformations derived from RNaseB. We found that conformations with slight perturbations were not reglucosylated by GT. In contrast, a partially structured nonnative form was efficiently recognized by the enzyme. When this form was converted back to a nativelike state, concomitant loss of recognition by GT occurred, reproducing the reglucosylation conditions observed in vivo with isolated components. Moreover, fully unfolded conformers were poorly recognized. The results indicated that GT is able to distinguish between different nonnative conformations with a distinct preference for partially structured conformers. The findings suggest that discrete populations of nonnative conformations are selectively reglucosylated to participate in the calnexin/calreticulin chaperone pathway. PMID- 10725326 TI - Caspase recruitment domain (CARD)-dependent cytoplasmic filaments mediate bcl10 induced NF-kappaB activation. AB - Mucosa-associated lymphoid tissue (MALT) lymphomas are associated with overexpression and constitutive activity of bcl10, a caspase recruitment domain (CARD)-containing protein that activates NF-kappaB. Here, we show that arrangement of overexpressed bcl10 protein in cytoplasmic filaments is essential for recruitment of signal transducer molecules-involved NF-kappaB activation. We also show that cytoskeleton elements regulate bcl10 signaling.Thus, organized assemblage of proteins in ordered structures linked to the cytoskeleton network may represent a general mechanism for intracellular signaling. PMID- 10725327 TI - Synaptotagmin VII regulates Ca(2+)-dependent exocytosis of lysosomes in fibroblasts. AB - Synaptotagmins (Syts) are transmembrane proteins with two Ca(2+)-binding C(2) domains in their cytosolic region. Syt I, the most widely studied isoform, has been proposed to function as a Ca(2+) sensor in synaptic vesicle exocytosis. Several of the twelve known Syts are expressed primarily in brain, while a few are ubiquitous (Sudhof, T.C., and J. Rizo. 1996. Neuron. 17: 379-388; Butz, S., R. Fernandez-Chacon, F. Schmitz, R. Jahn, and T.C. Sudhof. 1999. J. Biol. Chem. 274:18290-18296). The ubiquitously expressed Syt VII binds syntaxin at free Ca(2+) concentrations ([Ca(2+)]) below 10 microM, whereas other isoforms require 200-500 microM [Ca(2+)] or show no Ca(2+)-dependent syntaxin binding (Li, C., B. Ullrich, Z. Zhang, R.G.W. Anderson, N. Brose, and T.C. Sudhof. 1995. Nature. 375:594-599). We investigated the involvement of Syt VII in the exocytosis of lysosomes, which is triggered in several cell types at 1-5 microM [Ca(2+)] (Rodriguez, A., P. Webster, J. Ortego, and N.W. Andrews. 1997. J. Cell Biol. 137:93-104). Here, we show that Syt VII is localized on dense lysosomes in normal rat kidney (NRK) fibroblasts, and that GFP-tagged Syt VII is targeted to lysosomes after transfection. Recombinant fragments containing the C(2)A domain of Syt VII inhibit Ca(2+)-triggered secretion of beta-hexosaminidase and surface translocation of Lgp120, whereas the C(2)A domain of the neuronal- specific isoform, Syt I, has no effect. Antibodies against the Syt VII C(2)A domain are also inhibitory in both assays, indicating that Syt VII plays a key role in the regulation of Ca(2+)-dependent lysosome exocytosis. PMID- 10725328 TI - The small GTPase, Rap1, mediates CD31-induced integrin adhesion. AB - Integrin-mediated leukocyte adhesion is a critical aspect of leukocyte function that is tightly regulated by diverse stimuli, including chemokines, antigen receptors, and adhesion receptors. How cellular signals from CD31 and other adhesion amplifiers are integrated with those from classical mitogenic stimuli to regulate leukocyte function remains poorly understood. Here, we show that the cytoplasmic tail of CD31, an important integrin adhesion amplifier, propagates signals that induce T cell adhesion via beta1 (VLA-4) and beta2 (LFA-1) integrins. We identify the small GTPase, Rap1, as a critical mediator of this effect. Importantly, CD31 selectively activated the small Ras-related GTPase, Rap1, but not Ras, R-Ras, or Rap2. An activated Rap1 mutant stimulated T lymphocyte adhesion to intercellular adhesion molecule (ICAM) and vascular cell adhesion molecule (VCAM), as did the Rap1 guanine nucleotide exchange factor C3G and a catalytically inactive mutant of RapGAP. Conversely, negative regulators of Rap1 signaling blocked CD31-dependent adhesion. These findings identify a novel important role for Rap1 in regulating ligand-induced cell adhesion and suggest that Rap1 may play a more general role in coordinating adhesion-dependent signals during leukocyte migration and extravasation. Our findings also suggest an alternative mechanism, distinct from interference with Ras-proximal signaling, by which Rap1 might mediate transformation reversion. PMID- 10725329 TI - Hyaluronic acid (HA) binding to CD44 activates Rac1 and induces lamellipodia outgrowth. AB - Both cell adhesion protein CD44 and its main ligand hyaluronic acid (HA) are thought to be involved in several processes ultimately requiring cytoskeleton rearrangements. Here, we show that the small guanine nucleotide (GTP)-binding protein, Rac1, can be activated upon HA binding to CD44. When applied locally to a passive cell edge, HA promoted the formation of lamellipodial protrusions in the direction of the stimulus. This process was inhibited by the prior injection of cells with dominant-negative N17Rac recombinant protein or by pretreatment of cells with monoclonal anti-CD44 antibodies, interfering with HA binding, implying the direct involvement of CD44 in signaling to Rac1. PMID- 10725330 TI - Ubinuclein, a novel nuclear protein interacting with cellular and viral transcription factors. AB - The major target tissues for Epstein-Barr virus (EBV) infection are B lymphocytes and epithelial cells of the oropharyngeal zone. The product of the EBV BZLF1 early gene, EB1, a member of the basic leucine-zipper family of transcription factors, interacts with both viral and cellular promoters and transcription factors, modulating the reactivation of latent EBV infection. Here, we characterize a novel cellular protein interacting with the basic domains of EB1 and c-Jun, and competing of their binding to the AP1 consensus site. The transcript is present in a wide variety of human adult, fetal, and tumor tissues, and the protein is detected in the nuclei throughout the human epidermis and as either grainy or punctuate nuclear staining in the cultured keratinocytes. The overexpression of tagged cDNA constructs in keratinocytes revealed that the NH(2) terminus is essential for the nuclear localization, while the central domain is responsible for the interaction with EB1 and for the phenotype of transfected keratinocytes similar to terminal differentiation. The gene was identified in tail-to-tail orientation with the periplakin gene (PPL) in human chromosome 16p13.3 and in a syntenic region in mouse chromosome 16. We designated this novel ubiquitously expressed nuclear protein as ubinuclein and the corresponding gene as UBN1. PMID- 10725331 TI - Gemin4. A novel component of the SMN complex that is found in both gems and nucleoli. AB - The survival of motor neurons (SMN) protein, the product of the neurodegenerative disease spinal muscular atrophy (SMA) gene, is localized both in the cytoplasm and in discrete nuclear bodies called gems. In both compartments SMN is part of a large complex that contains several proteins including Gemin2 (formerly SIP1) and the DEAD box protein Gemin3. In the cytoplasm, the SMN complex is associated with snRNP Sm core proteins and plays a critical role in spliceosomal snRNP assembly. In the nucleus, SMN is required for pre-mRNA splicing by serving in the regeneration of spliceosomes. These functions are likely impaired in cells of SMA patients because they have reduced levels of functional SMN. Here, we report the identification by nanoelectrospray mass spectrometry of a novel component of the SMN complex that we name Gemin4. Gemin4 is associated in vivo with the SMN complex through a direct interaction with Gemin3. The tight interaction of Gemin4 with Gemin3 suggests that it could serve as a cofactor of this DEAD box protein. Gemin4 also interacts directly with several of the Sm core proteins. Monoclonal antibodies against Gemin4 efficiently immunoprecipitate the spliceosomal U snRNAs U1 and U5 from Xenopus oocytes cytoplasm. Immunolocalization experiments show that Gemin4 is colocalized with SMN in the cytoplasm and in gems. Interestingly, Gemin4 is also detected in the nucleoli, suggesting that the SMN complex may also function in preribosomal RNA processing or ribosome assembly. PMID- 10725333 TI - Cholera toxin is exported from microsomes by the Sec61p complex. AB - After endocytosis cholera toxin is transported to the endoplasmic reticulum (ER), from where its A1 subunit (CTA1) is assumed to be transferred to the cytosol by an as-yet unknown mechanism. Here, export of CTA1 from the ER to the cytosol was investigated in a cell-free assay using either microsomes loaded with CTA1 by in vitro translation or reconstituted microsomes containing CTA1 purified from V. cholerae. Export of CTA1 from the microsomes was time- and adenosine triphosphate dependent and required lumenal ER proteins. By coimmunoprecipitation CTA1 was shown to be associated during export with the Sec61p complex, which mediates import of proteins into the ER. Export of CTA1 was inhibited when the Sec61p complexes were blocked by nascent polypeptides arrested during import, demonstrating that the export of CTA1 depended on translocation-competent Sec61p complexes. Export of CTA1 from the reconstituted microsomes indicated the de novo insertion of the toxin into the Sec61p complex from the lumenal side. Our results suggest that Sec61p complex-mediated protein export from the ER is not restricted to ER-associated protein degradation but is also used by bacterial toxins, enabling their entry into the cytosol of the target cell. PMID- 10725334 TI - Toc64, a new component of the protein translocon of chloroplasts. AB - A subunit of the preprotein translocon of the outer envelope of chloroplasts (Toc complex) of 64 kD is described, Toc64. Toc64 copurifies on sucrose density gradients with the isolated Toc complex. Furthermore, it can be cross-linked in intact chloroplasts to a high molecular weight complex containing both Toc and Tic subunits and a precursor protein. The 0 A cross-linker CuCl(2) yields the reversible formation of disulfide bridge(s) between Toc64 and the established Toc complex subunits in purified outer envelope membranes. Toc64 contains three tetratricopeptide repeat motifs that are exposed at the chloroplast cytosol interface. We propose that Toc64 functions early in preprotein translocation, maybe as a docking protein for cytosolic cofactors of the protein import into chloroplasts. PMID- 10725332 TI - Subcellular compartmentalization of E2F family members is required for maintenance of the postmitotic state in terminally differentiated muscle. AB - Maintenance of cells in a quiescent state after terminal differentiation occurs through a number of mechanisms that regulate the activity of the E2F family of transcription factors. We report here that changes in the subcellular compartmentalization of the E2F family proteins are required to prevent nuclei in terminally differentiated skeletal muscle from reentering S phase. In terminally differentiated L6 myotubes, E2F-1, E2F-3, and E2F-5 were primarily cytoplasmic, E2F-2 was nuclear, whereas E2F-4 became partitioned between both compartments. In these same cells, pRB family members, pRB, p107, and p130 were also nuclear. This compartmentalization of the E2F-1 and E2F-4 in differentiated muscle cells grown in vitro reflected their observed subcellular location in situ. We determined further that exogenous E2F-1 or E2F-4 expressed in myotubes at levels fourfold greater than endogenous proteins compartmentalized identically to their endogenous counterparts. Only when overexpressed at higher levels was inappropriate subcellular location for these proteins observed. At these levels, induction of the E2F-regulated genes, cyclins A and E, and suppression of factors associated with myogenesis, myogenin, and p21(Cip1) was observed. Only at these levels of E2F expression did nuclei in these terminally differentiated cells enter S phase. These data demonstrate that regulation of the subcellular compartmentalization of E2F-family members is required to maintain nuclei in a quiescent state in terminally differentiated cells. PMID- 10725335 TI - Proteins needed for vesicle budding from the Golgi complex are also required for the docking step of homotypic vacuole fusion. AB - Vam2p/Vps41p is known to be required for transport vesicles with vacuolar cargo to bud from the Golgi. Like other VAM-encoded proteins, which are needed for homotypic vacuole fusion, we now report that Vam2p and its associated protein Vam6p/Vps39p are needed on each vacuole partner for homotypic fusion. In vitro vacuole fusion occurs in successive steps of priming, docking, and membrane fusion. While priming does not require Vam2p or Vam6p, the functions of these two proteins cannot be fulfilled until priming has occurred, and each is required for the docking reaction which culminates in trans-SNARE pairing. Consistent with their dual function in Golgi vesicle budding and homotypic fusion of vacuoles, approximately half of the Vam2p and Vam6p of the cell are recovered from cell lysates with purified vacuoles. PMID- 10725336 TI - The docking stage of yeast vacuole fusion requires the transfer of proteins from a cis-SNARE complex to a Rab/Ypt protein. AB - The homotypic fusion of yeast vacuoles requires Sec18p (NSF)-driven priming to allow vacuole docking, but the mechanism that links priming and docking is unknown. We find that a large multisubunit protein called the Vam2/6p complex is bound to cis-paired SNAP receptors (SNAREs) on isolated vacuoles. This association of the Vam2/6p complex with the cis-SNARE complex is disrupted during priming. The Vam2/6p complex then binds to Ypt7p, a guanosine triphosphate binding protein of the Rab family, to initiate productive contact between vacuoles. Thus, cis-SNARE complexes can contain Rab/Ypt effectors, and these effectors can be mobilized by NSF/Sec18p-driven priming, allowing their direct association with a Rab/Ypt protein to activate docking. PMID- 10725337 TI - Active caspases and cleaved cytokeratins are sequestered into cytoplasmic inclusions in TRAIL-induced apoptosis. AB - Tumor necrosis factor-related apoptosis- inducing ligand (TRAIL) -induced apoptosis, in transformed human breast epithelial MCF-7 cells, resulted in a time dependent activation of the initiator caspases-8 and -9 and the effector caspase 7. Cleavage of caspase-8 and its preferred substrate, Bid, preceded processing of caspases-7 and -9, indicating that caspase-8 is the apical initiator caspase in TRAIL-induced apoptosis. Using transient transfection of COOH-terminal-tagged green fluorescent protein fusion constructs, caspases-3, -7, and -8 were localized throughout the cytoplasm of MCF-7 cells. TRAIL-induced apoptosis resulted in activation of caspases-3 and -7, and the redistribution of most of their detectable catalytically active small subunits into large spheroidal cytoplasmic inclusions, which lacked a limiting membrane. These inclusions, which were also induced in untransfected cells, contained cytokeratins 8, 18, and 19, together with both a phosphorylated form and a caspase-cleavage fragment of cytokeratin 18. Similarly, in untransfected breast HBL100 and lung A549 epithelial cells, TRAIL induced the formation of cytoplasmic inclusions that contained cleaved cytokeratin 18 and colocalized with active endogenous caspase 3. We propose that effector caspase-mediated cleavage of cytokeratins, resulting in disassembly of the cytoskeleton and formation of cytoplasmic inclusions, may be a characteristic feature of epithelial cell apoptosis. PMID- 10725338 TI - Kinesin superfamily protein 3 (KIF3) motor transports fodrin-associating vesicles important for neurite building. AB - Kinesin superfamily proteins (KIFs) comprise several dozen molecular motor proteins. The KIF3 heterotrimer complex is one of the most abundantly and ubiquitously expressed KIFs in mammalian cells. To unveil the functions of KIF3, microinjection of function-blocking monovalent antibodies against KIF3 into cultured superior cervical ganglion (SCG) neurons was carried out. They significantly blocked fast axonal transport and brought about inhibition of neurite extension. A yeast two-hybrid binding assay revealed the association of fodrin with the KIF3 motor through KAP3. This was further confirmed by using vesicles collected from large bundles of axons (cauda equina), from which membranous vesicles could be prepared in pure preparations. Both immunoprecipitation and immunoelectron microscopy indicated the colocalization of fodrin and KIF3 on the same vesicles, the results reinforcing the evidence that the cargo of the KIF3 motor consists of fodrin-associating vesicles. In addition, pulse-labeling study implied partial comigration of both molecules as fast flow components. Taken together, the KIF3 motor is engaged in fast axonal transport that conveys membranous components important for neurite extension. PMID- 10725339 TI - beta-arrestin-dependent endocytosis of proteinase-activated receptor 2 is required for intracellular targeting of activated ERK1/2. AB - Recently, a requirement for beta-arrestin-mediated endocytosis in the activation of extracellular signal-regulated kinases 1 and 2 (ERK1/2) by several G protein coupled receptors (GPCRs) has been proposed. However, the importance of this requirement for function of ERK1/2 is unknown. We report that agonists of Galphaq coupled proteinase-activated receptor 2 (PAR2) stimulate formation of a multiprotein signaling complex, as detected by gel filtration, immunoprecipitation and immunofluorescence. The complex, which contains internalized receptor, beta-arrestin, raf-1, and activated ERK, is required for ERK1/2 activation. However, ERK1/2 activity is retained in the cytosol and neither translocates to the nucleus nor causes proliferation. In contrast, a mutant PAR2 (PAR2deltaST363/6A), which is unable to interact with beta-arrestin and, thus, does not desensitize or internalize, activates ERK1/2 by a distinct pathway, and fails to promote both complex formation and cytosolic retention of the activated ERK1/2. Whereas wild-type PAR2 activates ERK1/2 by a PKC-dependent and probably a ras-independent pathway, PAR2(deltaST363/6A) appears to activate ERK1/2 by a ras-dependent pathway, resulting in increased cell proliferation. Thus, formation of a signaling complex comprising PAR2, beta-arrestin, raf-1, and activated ERK1/2 might ensure appropriate subcellular localization of PAR2 mediated ERK activity, and thereby determine the mitogenic potential of receptor agonists. PMID- 10725340 TI - Determination of cell adhesion sites of neuropilin-1. AB - Neuropilin-1 is a type 1 membrane protein with three distinct functions. First, it can mediate cell adhesion via a heterophilic molecular interaction. Second, in neuronal cells, neuropilin-1 binds the class 3 semaphorins, which are neuronal chemorepellents, and plays a role in the directional guidance of axons. Neuropilin-1 is expected to form complexes with the plexinA subfamily members and mediate the semaphorin-elicited inhibitory signals into neurons. Third, in endothelial cells, neuropilin-1 binds a potent endothelial cell mitogen, vascular endothelial growth factor (VEGF)(165), and regulates vessel formation. Though the binding sites in neuropilin-1 for the class 3 semaphorins and VEGF(165) have been analyzed, the sites involved in cell adhesion activity of the molecule have not been identified. In this study, we produced a variety of mutant neuropilin-1s and tested their cell adhesion activity. We showed that the b1 and b2 domains within the extracellular segment of neuropilin-1 were required for the cell adhesion activity, and peptides with an 18-amino acid stretch in the b1 and b2 domains were sufficient to induce the cell adhesion activity. In addition, we demonstrated that the cell adhesion ligands for neuropilin-1 were proteins and distributed in embryonic mesenchymal cells but distinct from the class 3 semaphorins, VEGF, or plexins. PMID- 10725341 TI - The tissue plasminogen activator (tPA)/plasmin extracellular proteolytic system regulates seizure-induced hippocampal mossy fiber outgrowth through a proteoglycan substrate. AB - Short seizure episodes are associated with remodeling of neuronal connections. One region where such reorganization occurs is the hippocampus, and in particular, the mossy fiber pathway. Using genetic and pharmacological approaches, we show here a critical role in vivo for tissue plasminogen activator (tPA), an extracellular protease that converts plasminogen to plasmin, to induce mossy fiber sprouting. We identify DSD-1-PG/phosphacan, an extracellular matrix component associated with neurite reorganization, as a physiological target of plasmin. Mice lacking tPA displayed decreased mossy fiber outgrowth and an aberrant band at the border of the supragranular region of the dentate gyrus that coincides with the deposition of unprocessed DSD-1-PG/phosphacan and excessive Timm-positive, mossy fiber termini. Plasminogen-deficient mice also exhibit the laminar band and DSD- 1-PG/phosphacan deposition, but mossy fiber outgrowth through the supragranular region is normal. These results demonstrate that tPA functions acutely, both through and independently of plasmin, to mediate mossy fiber reorganization. PMID- 10725343 TI - Aneuploidy vs. gene mutation hypothesis of cancer: recent study claims mutation but is found to support aneuploidy. AB - For nearly a century, cancer has been blamed on somatic mutation. But it is still unclear whether this mutation is aneuploidy, an abnormal balance of chromosomes, or gene mutation. Despite enormous efforts, the currently popular gene mutation hypothesis has failed to identify cancer-specific mutations with transforming function and cannot explain why cancer occurs only many months to decades after mutation by carcinogens and why solid cancers are aneuploid, although conventional mutation does not depend on karyotype alteration. A recent high profile publication now claims to have solved these discrepancies with a set of three synthetic mutant genes that "suffices to convert normal human cells into tumorigenic cells." However, we show here that even this study failed to explain why it took more than "60 population doublings" from the introduction of the first of these genes, a derivative of the tumor antigen of simian virus 40 tumor virus, to generate tumor cells, why the tumor cells were clonal although gene transfer was polyclonal, and above all, why the tumor cells were aneuploid. If aneuploidy is assumed to be the somatic mutation that causes cancer, all these results can be explained. The aneuploidy hypothesis predicts the long latent periods and the clonality on the basis of the following two-stage mechanism: stage one, a carcinogen (or mutant gene) generates aneuploidy; stage two, aneuploidy destabilizes the karyotype and thus initiates an autocatalytic karyotype evolution generating preneoplastic and eventually neoplastic karyotypes. Because the odds are very low that an abnormal karyotype will surpass the viability of a normal diploid cell, the evolution of a neoplastic cell species is slow and thus clonal, which is comparable to conventional evolution of new species. PMID- 10725344 TI - Isoleucine-15 of rainbow trout carbonyl reductase-like 20beta-hydroxysteroid dehydrogenase is critical for coenzyme (NADPH) binding. AB - Carbonyl reductase-like 20beta-hydroxysteroid dehydrogenase (CR/20beta-HSD) is an enzyme that converts 17alpha-hydroxyprogesterone to 17alpha, 20beta-dihydroxy-4 pregnen-3-one (the maturation-inducing hormone of salmonid fish). We have previously isolated two types of CR/20beta-HSD cDNAs from ovarian follicle of rainbow trout (Oncorhynchus mykiss). Recombinant proteins produced by expression in Escherichia coli in vitro showed that one (type A) had CR and 20beta-HSD activity but that the other (type B) did not. Among the three distinct residues between the protein products encoded by the two cDNAs, two residues (positions 15 and 27) are located in the N-terminal Rossmann fold, the coenzyme binding site. To investigate the structure/function relationships of CR/20beta-HSDs, we generated mutants by site-directed mutagenesis at the following positions: MutA/I15T, MutB/T15I, and MutB/Q27K. Enzyme activity of wild-type A was abolished by substitution of Ile-15 by Thr (MutA/I15T). Conversely, enzyme activity was acquired by the replacement of Thr-15 with Ile in type B (MutB/T15I). MutB/T15I mutant showed properties similar to the wild-type A in every aspect tested. Mutation MutB/Q27K had only partial enzyme activity, indicating that Ile-15 plays an important role in enzyme binding of cofactor NADPH. PMID- 10725342 TI - Bacterial exposure induces and activates matrilysin in mucosal epithelial cells. AB - Matrilysin, a matrix metalloproteinase, is expressed and secreted lumenally by intact mucosal and glandular epithelia throughout the body, suggesting that its regulation and function are shared among tissues. Because matrilysin is produced in Paneth cells of the murine small intestine, where it participates in innate host defense by activation of prodefensins, we speculated that its expression would be influenced by bacterial exposure. Indeed, acute infection (10-90 min) of human colon, bladder, and lung carcinoma cells, primary human tracheal epithelial cells, and human tracheal explants with type 1-piliated Escherichia coli mediated a marked (25-50-fold) and sustained (>24 h) induction of matrilysin production. In addition, bacterial infection resulted in activation of the zymogen form of the enzyme, which was selectively released at the apical surface. Induction of matrilysin was mediated by a soluble, non-LPS bacterial factor and correlated with the release of defensin-like bacteriocidal activity. Bacteria did not induce matrilysin in other cell types, and expression of other metalloproteinases by epithelial cells was not affected by bacteria. Matrilysin was not detected in germ-free mice, but the enzyme was induced after colonization with Bacteroides thetaiotaomicron. These findings indicate that bacterial exposure is a potent and physiologically relevant signal regulating matrilysin expression in epithelial cells. PMID- 10725345 TI - Activation of the human estrogen receptor by the antiestrogens ICI 182,780 and tamoxifen in yeast genetic systems: implications for their mechanism of action. AB - The antiestrogens tamoxifen and ICI 182,780 have been portrayed as competitive antagonists of the estrogen binding site of the alpha-form of the human estrogen receptor (ER). However, in functional studies, neither compound has consistently been able to block estradiol-induced transcription. In this report, three yeast genetic systems were used to investigate the effects of tamoxifen and ICI 182,780 on ER dimerization, transcriptional activation, and the interaction of the receptor with a coactivator, RIP140. Tamoxifen and ICI 182,780 were able to induce ER dimerization and ER-dependent transcription, albeit at up to 15,000 fold higher concentrations than that of estradiol. In the presence of RIP140, the transcription response maximum was increased up to 30-fold for estradiol and both antiestrogens. Whole yeast cell [(3)H]estradiol binding studies demonstrated that tamoxifen could displace the estradiol from the ER, whereas ICI 182,780 treatment resulted in a 4-fold increase in [(3)H]estradiol binding to the receptor. No antagonism of estradiol was observed with tamoxifen or ICI 182,780 in any of the yeast models employed. We have concluded that the antiestrogen activity of compounds like tamoxifen and ICI 182,780 is not caused by their ability to competitively antagonize estradiol binding to the hormone binding site, but possibly by their ability to induce ER-dependent transcription, which in mammalian systems would result in receptor down-regulation. Compounds such as tamoxifen act through the hormone binding site, whereas ICI 182,780 may cause receptor activation through an allosteric binding site. PMID- 10725346 TI - Modification of P-selectin glycoprotein ligand-1 with a natural killer cell restricted sulfated lactosamine creates an alternate ligand for L-selectin. AB - Natural killer (NK) cells are components of the innate immune system that can recognize and kill virally infected cells, tumor cells, and allogeneic cells without prior sensitization. NK cells also elaborate cytokines (e.g., interferon gamma and tumor necrosis factor-alpha) and chemokines (e.g., macrophage inflammatory protein-1alpha) that promote the acquisition of antigen-specific immunity. NK cell differentiation is accompanied by the cell surface expression of a mucin-like glycoprotein bearing an NK cell-restricted keratan sulfate related lactosamine carbohydrate, the PEN5 epitope. Here, we report that PEN5 is a post-translational modification of P-selectin glycoprotein ligand-1 (PSGL-1). The PEN5 epitope creates on PSGL-1 a unique binding site for L-selectin, which is independent of PSGL-1 tyrosine sulfation. On the surface of NK cells, the expression of PEN5 is coordinated with the disappearance of L-selectin and the up regulation of Killer cell Ig-like Receptors (KIR). These results indicate that NK cell differentiation is accompanied by the acquisition of a unique carbohydrate, PEN5, that can serve as part of a combination code to deliver KIR(+) NK cells to specific tissues. PMID- 10725347 TI - Synthetic promoter elements obtained by nucleotide sequence variation and selection for activity. AB - Eukaryotic transcriptional regulation in different cells involves large numbers and arrangements of cis and trans elements. To survey the number of cis regulatory elements that are active in different contexts, we have devised a high throughput selection procedure permitting synthesis of active cis motifs that enhance the activity of a minimal promoter. This synthetic promoter construction method (SPCM) was used to identify >100 DNA sequences that showed increased promoter activity in the neuroblastoma cell line Neuro2A. After determining DNA sequences of selected synthetic promoters, database searches for known elements revealed a predominance of eight motifs: AP2, CEBP, GRE, Ebox, ETS, CREB, AP1, and SP1/MAZ. The most active of the selected synthetic promoters contain composites of a number of these motifs. Assays of DNA binding and promoter activity of three exemplary motifs (ETS, CREB, and SP1/MAZ) were used to prove the effectiveness of SPCM in uncovering active sequences. Up to 10% of 133 selected active sequences had no match in currently available databases, raising the possibility that new motifs and transcriptional regulatory proteins to which they bind may be revealed by SPCM. The method may find uses in constructing databases of active cis motifs, in diagnostics, and in gene therapy. PMID- 10725349 TI - A comparison of naive and sophisticated subject behavior with game theoretic predictions. AB - We use an extensive form two-person game as the basis for two experiments designed to compare the behavior of two groups of subjects with each other and with the subgame perfect theoretical prediction in an anonymous interaction protocol. The two subject groups are undergraduates and advanced graduate students, the latter having studied economics and game theory. There is no difference in their choice behavior, and both groups depart substantially from game theoretic predictions. We also compare a subsample of the same graduate students with a typical undergraduate sample in an asset trading environment in which inexperienced undergraduates invariably produce substantial departures from the rational expectations prediction. In this way, we examine how robust are the results across two distinct anonymous interactive environments. In the constant sum trading game, the graduate students closely track the predictions of rational theory. Our interpretation is that the graduate student subjects' departure from subgame perfection to achieve cooperative outcomes in the two-person bargaining game is a consequence of a deliberate strategy and is not the result of error or inadequate learning. PMID- 10725348 TI - Evolution and assembly of an extremely scrambled gene. AB - The process of gene unscrambling in hypotrichous ciliates represents one of nature's ingenious solutions to the problem of gene assembly. With some essential genes scrambled in as many as 51 pieces, these ciliates rely on sequence and structural cues to rebuild their fragmented genes and genomes. Here we report the complex pattern of scrambling in the DNA polymerase alpha gene of Stylonychia lemnae. The germline (micronuclear) copy of this gene is broken into 48 pieces with 47 dispersed over two loci, with no asymmetry in the placement of coding segments on either strand. Direct repeats present at the boundaries between coding and noncoding sequences provide pointers to help guide assembly of the functional (macronuclear) gene. We investigate the evolution of this complex gene in three hypotrichous species. PMID- 10725350 TI - The Arabidopsis SOS2 protein kinase physically interacts with and is activated by the calcium-binding protein SOS3. AB - The Arabidopsis thaliana SOS2 and SOS3 genes are required for intracellular Na(+) and K(+) homeostasis and plant tolerance to high Na(+) and low K(+) environments. SOS3 is an EF hand type calcium-binding protein having sequence similarities with animal neuronal calcium sensors and the yeast calcineurin B. SOS2 is a serine/threonine protein kinase in the SNF1/AMPK family. We report here that SOS3 physically interacts with and activates SOS2 protein kinase. Genetically, sos2sos3 double mutant analysis indicates that SOS2 and SOS3 function in the same pathway. Biochemically, SOS2 interacts with SOS3 in the yeast two-hybrid system and in vitro binding assays. The interaction is mediated by the C-terminal regulatory domain of SOS2. SOS3 activates SOS2 protein kinase activity in a Ca(2+)-dependent manner. Therefore, SOS3 and SOS2 define a novel regulatory pathway important for the control of intracellular ion homeostasis and salt tolerance in plants. PMID- 10725351 TI - Adaptive diversification within a large family of recently duplicated, placentally expressed genes. AB - The pregnancy-associated glycoproteins (PAG) are putative peptide-binding proteins and products of a large family of genes whose expression is localized to the placental surface epithelium of artiodactyl species. We have tested the hypothesis that natural selection has favored diversification of these genes by examining patterns of nucleotide substitution in a sample of 28 closely related bovine, caprine, and ovine family members that are expressed only in trophoblast binucleate cells. Three observations were made. First, in codons encoding highly variable domains of the proteins, there was a greater accumulation of both synonymous and nonsynonymous mutations than in the more conserved regions of the genes. Second, in the variable regions, the mean number of nonsynonymous nucleotide substitutions per site was significantly greater than the mean number of synonymous substitutions per site. Third, nonsynonymous changes affecting amino acid charge occurred more frequently than expected under random substitution. This unusual pattern of nucleotide substitution implies that natural selection has acted to diversify these PAG molecules at the amino acid level, which in turn suggests that these molecules have undergone functional diversification. We estimate that the binucleate cell-expressed PAG originated 52 +/- 6 million years ago, soon after the divergence of the ruminant lineage. Thus, rapid functional diversification of PAG expressed in trophoblast binucleate cells seems to have been associated with the origin of this unique placental adaptation. PMID- 10725353 TI - A functional role for the two-pore domain potassium channel TASK-1 in cerebellar granule neurons. AB - Cerebellar granule neurons (CGNs) are one of the most populous cells in the mammalian brain. They express an outwardly rectifying potassium current, termed a "standing-outward" K(+) current, or IK(SO), which does not inactivate. It is active at the resting potential of CGNs, and blocking IK(SO) leads to cell depolarization. IK(SO) is blocked by Ba(2+) ions and is regulated by activation of muscarinic M(3) receptors, but it is insensitive to the classical broad spectrum potassium channel blocking drugs 4-aminopyridine and tetraethylammonium ions. The molecular nature of this important current has yet to be established, but in this study, we provide strong evidence to suggest that IK(SO) is the functional correlate of the recently identified two-pore domain potassium channel TASK-1. We show that IK(SO) has no threshold for activation by voltage and that it is blocked by small extracellular acidifications. Both of these are properties that are diagnostic of TASK-1 channels. In addition, we show that TASK-1 currents expressed in Xenopus oocytes are inhibited after activation of endogenous M(3) muscarinic receptors. Finally, we demonstrate that mRNA for TASK-1 is found in CGNs and that TASK-1 protein is expressed in CGN membranes. This description of a functional two-pore domain potassium channel in the mammalian central nervous system indicates its physiological importance in controlling cell excitability and how agents that modify its activity, such as agonists at G protein-coupled receptors and hydrogen ions, can profoundly alter both the neuron's resting potential and its excitability. PMID- 10725352 TI - Dictyostelium amoebae lacking an F-box protein form spores rather than stalk in chimeras with wild type. AB - Using a selection for Dictyostelium mutants that preferentially form spores, we have recovered a mutant called CheaterA. In chimeras with isogenic wild-type cells, the CheaterA mutant preferentially forms viable spores rather than inviable stalk cells. The mutant causes wild-type cells that have begun to express spore-specific genes to accumulate in the prestalk compartment of the developing organism. In the wild-type cells, the chtA transcript is absent in growing cells and appears early in development. No transcript was detected in the mutant by Northern blot. The chtA gene codes for a protein with an F-box and WD40 domains. This class of protein usually forms part of an Skp1, cullin, F-box (SCF) complex that targets specific protein substrates for ubiquitination and degradation. PMID- 10725354 TI - Signal transduction by a nondissociable heterotrimeric yeast G protein. AB - Many signal transduction pathways involve heterotrimeric G proteins. The accepted model for activation of heterotrimeric G proteins states that the protein dissociates to the free G(alpha) (GTP)-bound subunit and free G(betagamma) dimer. On GTP hydrolysis, G(alpha) (GDP) then reassociates with G(betagamma) [Gilman, A. G. (1987) Annu. Rev. Biochem. 56, 615-649]. We reexamined this hypothesis, by using the mating G protein of the yeast Saccharomyces cerevisiae encoded by the genes GPA1, STE4, and STE18. In the absence of mating pheromone, the G(alpha) (Gpa1) subunit represses the mating pathway. On activation by binding of pheromone to a serpentine receptor, the G(betagamma) (Ste4, Ste18) dimer transmits the signal to a mitogen-activated protein kinase cascade, leading to gene activation, arrest in the G(1) stage of the cell cycle, production of shmoos (mating projections), and cell fusion. We found that a Ste4-Gpa1 fusion protein transmitted the pheromone signal and activated the mating pathway as effectively as when Ste4 (G(beta)) and Gpa1 (G(alpha)) were coexpressed as separate proteins. Hence, dissociation of this G protein is not required for its activation. Rather, a conformational change in the heterotrimeric complex is likely to be involved in signal transduction. PMID- 10725355 TI - Apolipoprotein E regulates dietary cholesterol absorption and biliary cholesterol excretion: studies in C57BL/6 apolipoprotein E knockout mice. AB - The present study examined the role of apolipoprotein E (apoE) in the regulation of dietary cholesterol absorption and biliary cholesterol excretion. Increasing dietary cholesterol from 0.02% to 0.5% in C57BL/6 wild-type mice decreased the percentage of dietary cholesterol that is absorbed by 25%, and this decrease was associated with a 2-fold increase in gallbladder biliary cholesterol concentration. In contrast, increasing dietary cholesterol from 0. 02% to 0.5% in C57BL/6 apoE knockout mice produced no significant suppression of the percentage dietary cholesterol absorption and increased gallbladder biliary cholesterol concentration only 16%. Whereas in wild-type mice, the increase in dietary cholesterol increased the hepatic excretion of biliary cholesterol 4-fold, there was only a 2-fold increase in apoE knockout mice. On both the low- and the high cholesterol diets, whole liver and isolated hepatocyte cholesterol content was higher in the apoE knockout mice. These results suggest that, in response to dietary cholesterol, apoE may play a critical role in decreasing the percentage absorption of dietary cholesterol and increasing biliary cholesterol excretion. These observations suggest a mechanism whereby the absence of apoE contributes to the propensity for tissue cholesterol deposition and accelerated atherogenesis in apoE knockout mice. PMID- 10725356 TI - Riding the Ice Age El Nino? Pacific biogeography and evolution of Metrosideros subg. Metrosideros (Myrtaceae) inferred from nuclear ribosomal DNA. AB - Metrosideros subg. Metrosideros (Myrtaceae) comprises approximately 26 species distributed widely across the Pacific basin. They occur on the ancient Gondwanan landmasses of New Zealand and New Caledonia, as well as on the volcanic islands of the remote Pacific, from Melanesia to tropical Polynesia and the Bonin Island. Phylogenetic analysis based on nuclear ribosomal DNA spacer sequences from all named species showed Metrosideros umbellata of New Zealand as basal in the subgenus, with the remaining species falling into three monophyletic clades. One includes the seven New Caledonian species together with three daughters in western Oceania that probably dispersed during the mid/late Tertiary. A second contains six taxa located in east Melanesia and Samoa that may also have arisen from a mid/late Tertiary dispersal, in this instance from New Zealand. The third includes three New Zealand endemics along with all of the taxa in remote Polynesia and accounts for much of the total range of the subgenus. These dispersed taxa in Polynesia either are identical to the New Zealand species Metrosideros excelsa or differ by a single nucleotide change. We suggest that they are all derived from a Pleistocene dispersal out of New Zealand. A relatively recent dispersal is surprising, given that this wind-dispersed genus has occupied New Zealand for much of the Tertiary and that some of the islands in remote Polynesia date to at least the Miocene. We attribute this dramatic range expansion to climate change-specifically changes in wind flow patterns-in the southern hemisphere during worldwide glaciation. PMID- 10725357 TI - Light stress-regulated two-helix proteins in Arabidopsis thaliana related to the chlorophyll a/b-binding gene family. AB - The chlorophyll a/b, chlorophyll a/c, and chlorophyll a/a light-harvesting proteins are part of an extended gene family that also includes the transiently expressed stress proteins, the Elips (early light-induced proteins). Four Elip homologue proteins, encoded by single-copy nuclear genes, have been identified in the Arabidopsis thaliana database. These proteins were divided into two groups according to the expression pattern under light-stress conditions and the predicted secondary structure. Group one included two members of the Elip family with three predicted transmembrane helices and a gene expression strictly related to light stress. Group two included two proteins, the Seps (stress-enhanced proteins), which possessed two predicted transmembrane segments. The transcripts of Sep1 and Sep2 were present under low light conditions, but their level increased 4- to 10-fold during illumination of plants with high-intensity light. Preliminary data indicated that the induced transcripts were translated in vivo. Other physiological stress conditions, such as cold, heat, desiccation, salt, wounding, or oxidative stress, did not significantly influence the expression of Sep genes. In vitro import of radioactively labeled precursors of Seps into isolated chloroplasts confirmed the thylakoid membrane localization of these proteins. Considering the predicted protein structure and homology to other pigment-antenna proteins, the two-helix Seps might represent an evolutionary missing link between the one- and three-helix antenna proteins present in pro- and eukaryota. PMID- 10725358 TI - Mmh/Ogg1 gene inactivation results in accumulation of 8-hydroxyguanine in mice. AB - The major mutagenic base lesion in DNA caused by exposure to reactive oxygen species is 8-hydroxyguanine or 7, 8-dihydro-8-oxoguanine (8-OH-G). Products of the human MMH/OGG1 gene are known to catalyze in vitro the reactions repairing this DNA lesion. To analyze the function of Mmh in vivo, we generated a mouse line carrying a mutant Mmh allele by targeted gene disruption. Mmh homozygous mutant mice were found to have a physically normal appearance, but to have lost nicking activity in liver extracts for substrate DNA containing 8-OH-G, exhibiting a 3-fold increased accumulation of this adduct at 9 weeks of age compared with wild-type or heterozygous mice. Further elevation to 7-fold was observed in 14-week-old animals. Substantial increase of spontaneous mutation frequencies was clearly identified in Mmh mutant mice bearing transgenic gpt genes. These results indicate that exposure of DNA to endogenous oxidative species continuously produces the mutagenic adduct 8-OH-G in mice, and Mmh plays an essential role in repair of this DNA damage. PMID- 10725359 TI - Whole-genome expression analysis of snf/swi mutants of Saccharomyces cerevisiae. AB - The Saccharomyces cerevisiae Snf/Swi complex has been previously demonstrated to control transcription and chromatin structure of particular genes in vivo and to remodel nucleosomes in vitro. We have performed whole-genome expression analysis, using DNA microarrays, to study mutants deleted for a gene encoding one conserved (Snf2) or one unconserved (Swi1) Snf/Swi component. This analysis was performed on cells grown in both rich and minimal media. The microarray results, combined with Northern blot, computational, and genetic analyses, show that snf2Delta and swi1Delta mutations cause similar effects on mRNA levels, that Snf/Swi controls some genes differently in rich and minimal media, and that Snf/Swi control is exerted at the level of individual genes rather than over larger chromosomal domains. In addition, this work shows that Snf/Swi controls mRNA levels of MATalpha-specific genes, likely via controlling transcription of the regulators MATalpha1 and MCM1. Finally, we provide evidence that Snf/Swi acts both as an activator and as a repressor of transcription, and that neither mode of control is an indirect effect of the other. PMID- 10725360 TI - Transforming growth factor-beta 1 hyperexpression in African-American hypertensives: A novel mediator of hypertension and/or target organ damage. AB - Hypertension, a remediable risk factor for stroke, cardiovascular disease, and renal failure, affects 50 million individuals in the United States alone. African Americans (blacks) have a higher incidence and prevalence of hypertension and hypertension-associated target organ damage compared with Caucasian Americans (whites). Herein, we explored the hypotheses that transforming growth factor beta(1) (TGF-beta(1)) is hyperexpressed in hypertensives compared with normotensives and that TGF-beta(1) overexpression is more frequent in blacks compared with whites. These hypotheses were stimulated by our recent demonstration that TGF-beta(1) is hyperexpressed in blacks with end-stage renal disease compared with white end-stage renal disease patients and by the biological attributes of TGF-beta(1), which include induction of endothelin-1 expression, stimulation of renin release, and promotion of vascular and renal disease when TGF-beta(1) is produced in excess. TGF-beta(1) profiles were determined in black and white hypertensive subjects and normotensive controls and included circulating protein concentrations, mRNA steady-state levels, and codon 10 genotype. Our investigation demonstrated that TGF-beta(1) protein levels are highest in black hypertensives, and TGF-beta(1) protein as well as TGF-beta(1) mRNA levels are higher in hypertensives compared with normotensives. The proline allele at codon 10 (Pro(10)) was more frequent in blacks compared with whites, and its presence was associated with higher levels of TGF-beta(1) mRNA and protein. Our findings support the idea that TGF-beta(1) hyperexpression is a risk factor for hypertension and hypertensive complications and provides a mechanism for the excess burden of hypertension in blacks. PMID- 10725361 TI - Electrospray ionization mass spectrometry as a tool to analyze hydrogen/deuterium exchange kinetics of transmembrane peptides in lipid bilayers. AB - A method is described to study the precise positioning of transmembrane peptides in a phospholipid bilayer combining hydrogen/deuterium (H/D) exchange and nanoelectrospray ionization mass spectrometry. The method was tested by using model systems consisting of designed alpha-helical transmembrane peptides [acetylGW(2)(LA)(5)W(2)Aethanolamine (WALP16) and acetyl (GA)(3)W(2)(LA)(5)W(2)(AG)(3)ethanolamine (WALP16(+10))] incorporated in large unilamellar vesicles of 1, 2-dimyristoyl-sn-glycero-3-phosphocholine. Both peptides consist of an alternating leucine/alanine hydrophobic core sequence flanked by tryptophan residues as interfacial anchor residues. In the case of WALP16(+10), this sequence is extended at both ends by 5-aa glycine/alanine tails extending into the aqueous phase surrounding the bilayer. H/D exchange of labile hydrogens in these peptides was monitored in time after dilution of the vesicles in buffered deuterium oxide. It was found that the peptides can be measured by direct introduction of the proteoliposome suspension into the mass spectrometer. Several distinct H/D exchange rates were observed (corresponding to half-life values varying from 4 sigma(F), and resolution of approximately 0.98 A. PMID- 10725397 TI - Stanniocalcin: A molecular guard of neurons during cerebral ischemia. AB - Stanniocalcin (STC) is a glycoprotein hormone originally found in bony fish, in which it regulates calcium/phosphate homeostasis and protects against hypercalcemia. The recently characterized human STC shows about 70% homology with fish STC. We previously reported a constitutive expression of STC in terminally differentiated neurons. Here, we show that exposure of human neural-crest-derived cell line Paju to hypercalcemic culture medium induced expression of STC. Treatment of Paju cells with recombinant human STC increased their uptake of inorganic phosphate. Paju cells expressing STC by cDNA transfection displayed increased resistance to ischemic challenge and to elevated intracellular free calcium induced by treatment with thapsigargin. An up-regulated and redistributed expression of STC was observed in neurons surrounding the core of acute infarcts in human and rat brains. Given that mobilization and influx of calcium is considered a main neurotoxic mechanism following ischemia, our results suggest that the altered expression of STC contributes to the protection of cerebral neurons against hypoxic/ischemic damage. Manipulation of the STC expression may therefore offer a therapeutic approach to limit the injury after ischemic brain insults. PMID- 10725398 TI - Transplantation of ex vivo expanded endothelial progenitor cells for therapeutic neovascularization. AB - Animal studies and preliminary results in humans suggest that lower extremity and myocardial ischemia can be attenuated by treatment with angiogenic cytokines. The resident population of endothelial cells that is competent to respond to an available level of angiogenic growth factors, however, may potentially limit the extent to which cytokine supplementation enhances tissue neovascularization. Accordingly, we transplanted human endothelial progenitor cells (hEPCs) to athymic nude mice with hindlimb ischemia. Blood flow recovery and capillary density in the ischemic hindlimb were markedly improved, and the rate of limb loss was significantly reduced. Ex vivo expanded hEPCs may thus have utility as a "supply-side" strategy for therapeutic neovascularization. PMID- 10725399 TI - The 1,800-year oceanic tidal cycle: a possible cause of rapid climate change. AB - Variations in solar irradiance are widely believed to explain climatic change on 20,000- to 100,000-year time-scales in accordance with the Milankovitch theory of the ice ages, but there is no conclusive evidence that variable irradiance can be the cause of abrupt fluctuations in climate on time-scales as short as 1,000 years. We propose that such abrupt millennial changes, seen in ice and sedimentary core records, were produced in part by well characterized, almost periodic variations in the strength of the global oceanic tide-raising forces caused by resonances in the periodic motions of the earth and moon. A well defined 1,800-year tidal cycle is associated with gradually shifting lunar declination from one episode of maximum tidal forcing on the centennial time scale to the next. An amplitude modulation of this cycle occurs with an average period of about 5,000 years, associated with gradually shifting separation intervals between perihelion and syzygy at maxima of the 1,800-year cycle. We propose that strong tidal forcing causes cooling at the sea surface by increasing vertical mixing in the oceans. On the millennial time-scale, this tidal hypothesis is supported by findings, from sedimentary records of ice-rafting debris, that ocean waters cooled close to the times predicted for strong tidal forcing. PMID- 10725400 TI - Bax degradation by the ubiquitin/proteasome-dependent pathway: involvement in tumor survival and progression. AB - Previously we reported that proteasome inhibitors were able to overcome Bcl-2 mediated protection from apoptosis. Here we show that inhibition of the proteasome activity in Bcl-2-overexpressing cells accumulates the proapoptotic Bax protein to mitochondria/cytoplasm, where it interacts to Bcl-2 protein. This event was followed by release of mitochondrial cytochrome c into the cytosol and activation of caspase-mediated apoptosis. In contrast, proteasome inhibition did not induce any apparent changes in Bcl-2 protein levels. In addition, treatment with a proteasome inhibitor increased levels of ubiquitinated forms of Bax protein, without any effects on Bax mRNA expression. We also established a cell free Bax degradation assay in which an in vitro-translated, (35)S-labeled Bax protein can be degraded by a tumor cell protein extract, inhibitable by addition of a proteasome inhibitor or depletion of the proteasome or ATP. The Bax degradation activity can be reconstituted in the proteasome-depleted supernatant by addition of a purified 20S proteasome or proteasome-enriched fraction. Finally, by using tissue samples of human prostate adenocarcinoma, we demonstrated that increased levels of Bax degradation correlated well with decreased levels of Bax protein and increased Gleason scores of prostate cancer. Our studies strongly suggest that ubiquitin/proteasome-mediated Bax degradation is a novel survival mechanism in human cancer cells and that selective targeting of this pathway should provide a unique approach for treatment of human cancers, especially those overexpressing Bcl-2. PMID- 10725401 TI - Saturation of, and competition for entry into, the apical secretory pathway. AB - To investigate mechanisms of apical sorting in the secretory pathway of epithelial cells, we expressed varying amounts of the 165 amino acid isoform of vascular endothelial growth factor (VEGF(165)) and transforming growth factor beta1 (TGF-beta1) via replication defective adenoviruses. Apical sorting of both proteins was efficient at low expression levels but saturated or was reversed at high expression levels. High expression levels of TGF-beta1 were effective at competing VEGF(165) out of the apical pathway; however, VEGF(165) did not compete out TGF-beta1. Tunicamycin inhibition experiments showed that the apical polarity of VEGF(165) was independent of N-glycosylation. We conclude that the apical sorting of these two molecules is a saturable, signal-mediated process, involving competition for apical sorting receptors. The sorting of the two proteins does not appear to involve N-glycans as sorting signals, or lectin sorters. The observations are particularly relevant to gene therapy because they demonstrate that overexpression of a transgene can result in undesirable missorting of the encoded protein. PMID- 10725402 TI - Vaccination with tat toxoid attenuates disease in simian/HIV-challenged macaques. AB - The Tat protein is essential for HIV type 1 (HIV-1) replication and may be an important virulence factor in vivo. We studied the role of Tat in viral pathogenesis by immunizing rhesus macaques with chemically inactivated Tat toxoid and challenging these animals by intrarectal inoculation with the simian/human immunodeficiency virus 89.6PD. Immune animals had significantly attenuated disease with lowered viral RNA, interferon-alpha, and chemokine receptor expression (CXCR4 and CCR5) on CD4(+) T cells; these features of infection have been linked to in vitro effects of Tat and respond similarly to extracellular Tat protein produced during infection. Immunization with Tat toxoid inhibits key steps in viral pathogenesis and should be included in therapeutic or preventive HIV-1 vaccines. PMID- 10725403 TI - Destabilization of osteogenesis imperfecta collagen-like model peptides correlates with the identity of the residue replacing glycine. AB - Mutations resulting in replacement of one obligate Gly residue within the repeating (Gly-Xaa-Yaa)(n) triplet pattern of the collagen type I triple helix are the major cause of osteogenesis imperfecta (OI). Phenotypes of OI involve fragile bones and range from mild to perinatal lethal. In this study, host-guest triple-helical peptides of the form acetyl-(Gly-Pro-Hyp)(3)-Zaa-Pro-Hyp-(Gly-Pro Hyp)(4)-Gly-Gly-amide are used to isolate the influence of the residue replacing Gly on triple-helix stability, with Zaa = Gly, Ala, Arg, Asp, Glu, Cys, Ser, or Val. Any substitution for Zaa = Gly (melting temperature, T(m) = 45 degrees C) results in a dramatic destabilization of the triple helix. For Ala and Ser, T(m) decreases to approximately 10 degrees C, and for the Arg-, Val-, Glu-, and Asp containing peptides, T(m) < 0 degrees C. A Gly --> Cys replacement results in T(m) < 0 degrees C under reducing conditions but shows a broad transition (T(m) approximately 19 degrees C) in an oxidizing environment. Addition of trimethylamine N-oxide increases T(m) by approximately 5 degrees C per 1 M trimethylamine N-oxide, resulting in stable triple-helix formation for all peptides and allowing comparison of relative stabilities. The order of disruption of different Gly replacements in these peptides can be represented as Ala 500 U/ml) of exogenous tumour necrosis factor (TNFalpha) [M. Rolfe, N.H. James, R.A. Roberts, TNF suppresses apoptosis and induces S-phase in rodent hepatocytes: a mediator of the hepatocarcinogenicity of peroxisome proliferators?, Carcinogenesis 18 (1997) 2277-2280] are also able to stimulate hepatocyte DNA synthesis and suppress apoptosis, implicating TNFalpha in mediating or permitting the liver growth response to PPs. Here, using cultured mouse hepatocytes isolated from PPARalpha null mice, we have examined the role of the peroxisome proliferator activated receptor alpha (PPARalpha) in mediating the suppression of apoptosis caused by PPs. In addition we have investigated further the role of TNFalpha in mediating the rodent response to PPs. The PP nafenopin (50 microM) was unable to stimulate DNA synthesis measured by bromodeoxyuridine incorporation in these PPARalpha null mouse hepatocytes (96% of control), unlike epidermal growth factor, a growth factor used as a positive control. In assays of apoptosis using H33258 staining of chromatin condensation, nafenopin was unable to suppress either spontaneous or TGFbeta1-induced apoptosis. In contrast, high concentrations of TNFalpha (>500 U/ml) were able to both stimulate DNA synthesis (204% of control) and suppress apoptosis in PPARalpha null hepatocytes (40% and 38% of control for spontaneous and TGFbeta1-induced apoptosis respectively). However, TNFalpha could not stimulate beta-oxidation of palmitoyl CoA in either PPARalpha null mouse or B6C3F1 (PPARalpha wild type) mouse hepatocytes. These data confirm the dependence of the response to PPs on PPARalpha by demonstrating that PPARalpha mediates the suppression of hepatocyte apoptosis in response to PPs. In addition, the data provide evidence that high concentrations of TNFalpha can modulate DNA synthesis and apoptosis in the absence of PPs and PPARalpha. Thus, in vivo, physiological levels of TNFalpha may be permissive for a PPARalpha dependent growth response to PPs. PMID- 10725473 TI - Species differences in peroxisome proliferation; mechanisms and relevance. AB - Peroxisome proliferators are a class of structurally diverse chemicals, which induce liver carcinogenesis in rodents through interaction and activation of the Peroxisome Proliferator-Activated Receptor alpha (PPARalpha). PPARalpha agonists elicit a powerful pleiotropic response, which include hypolipidaemia. We have examined the response of species that are classically unresponsive to peroxisome proliferators. Whereas hamster responds to PPARalpha agonists by hepatomegaly and induction of marker genes, the guinea pig does not undergo hepatomegaly or induction of marker genes, such as CYP4A13. Both the hamster and the guinea pig have PPARalpha, and the guinea pig receptor has been characterised to be fully functional, as demonstrated in reporter gene expression assays. However, the guinea pig PPARalpha is expressed at low levels in liver, and the currently favoured hypothesis to explain species differences in hepatic peroxisome proliferation invokes the low level of PPARalpha as the principal determinant of species responsiveness. However, the demonstration that guinea pigs and humans undergo hypolipidaemia induced by PPARalpha-agonists calls into question the mode of action of PPARalpha agonists in "non-responsive" species. PMID- 10725474 TI - Species differences in hepatic peroxisome proliferation, cell replication and transforming growth factor-beta1 gene expression in the rat, Syrian hamster and guinea pig. AB - The objective of this study was to evaluate species differences in the hepatic effects of three potent rodent peroxisome proliferators, namely methylclofenapate (MCP), ciprofibrate (CIP) and Wy-14,643 (WY), particularly with respect to effects on replicative DNA synthesis and transforming growth factor-beta1 (TGF beta1) gene expression. Male Sprague-Dawley rats, Syrian hamsters and Dunkin Hartley guinea pigs were given daily oral doses of 0 (corn oil) and 75 mg/kg MCP for periods of 6 and 21 days. Syrian hamsters and guinea pigs were also treated with 25 mg/kg CIP and 25 mg/kg WY. Relative liver weights were significantly increased in peroxisome proliferator-treated rats and Syrian hamsters, but not in guinea pigs. Hepatic peroxisomal (palmitoyl-CoA oxidation) and microsomal (lauric acid 12-hydroxylase) fatty acid oxidising enzyme activities and CYP4A isoform mRNA levels were significantly increased in rats and Syrian hamsters, whereas only minor effects were observed in the guinea pig. Replicative DNA synthesis was studied by implanting 7-day osmotic pumps containing 5-bromo-2'-deoxyuridine during study days -1 to 6 and 14 to 21. Hepatocyte labelling index values were increased by MCP in the rat, but neither MCP, CIP nor WY produced any significant effect on replicative DNA synthesis in the Syrian hamster and guinea pig. MCP treatment increased TGF-beta1 and insulin-like growth factor II/mannose-6 phosphate (IGFII/Man6P) receptor gene expression in the rat. In the Syrian hamster, effects on TGF-beta1 and IGFII/Man6P receptor gene expression were also observed in some instances, whereas TGF-beta1 mRNA levels were essentially unchanged in the guinea pig. These results provide further evidence for marked species differences in response to rodent peroxisome proliferators. While peroxisome proliferators produce a wide spectrum of effects in rat liver, other species such as the Syrian hamster and guinea pig are less responsive and in the case of some endpoints (e.g., cell replication) may be refractory. PMID- 10725475 TI - Neonatal effects of nifedipine and ritodrine for preterm labor. AB - OBJECTIVE: We compared nifedipine and ritodrine for treatment of preterm labor with respect to neonatal outcome. METHODS: We conducted an open randomized multicenter study of neonatal outcome in 185 women who received either oral nifedipine (n = 95) or intravenous (IV) ritodrine (n = 90) for treatment of preterm labor. Secondary outcome measures included neonatal mortality and morbidity, especially neonatal intensive care unit (NICU) admission, respiratory distress syndrome (RDS), and intracranial bleeding. RESULTS: There were no significant differences in umbilical artery pH values and Apgar scores between groups. Nifedipine was associated with lower admission rates to the NICU (49% versus 66%; odds ratio 0. 51, confidence interval 0.28, 0.93) compared with ritodrine, and lower incidences of RDS (21% versus 37%; 0.46, 0.24, 0.89), intracranial bleeding (18% versus 31%; 0.48, 0.24, 0.96), and neonatal jaundice (52% versus 67%; 0.53, 0.29, 0.97). Logistic regression analysis showed that even after correction for gestational age at birth, newborn risk of RDS, intracranial bleeding, or neonatal jaundice was significantly lower in the nifedipine group than the ritodrine group. CONCLUSION: Nifedipine for treatment of preterm labor was associated with a lower incidence of neonatal morbidity than ritodrine. That difference appeared to be partly because of the higher tocolytic efficacy of nifedipine and partly because of an intrinsic beneficial effect of nifedipine, or the lack of harmful effects when compared with ritodrine. PMID- 10725476 TI - Indomethacin for preterm labor: a randomized comparison of vaginal and rectal oral routes. AB - OBJECTIVE: To compare the efficacy of intravaginal and intrarectal plus oral indomethacin for the treatment of preterm labor. METHODS: Between December 1996 and November 1998, 46 eligible gravidas admitted with singleton pregnancies and idiopathic preterm labor before 33 gestational weeks were randomized to receive 200 mg of intravaginal or intrarectal plus oral indomethacin. RESULTS: Twenty three subjects were allocated to each study group. The interval from initiation of treatment to delivery was significantly longer in the intravaginal indomethacin group (26.5 +/- 5.7 versus 12.6 +/- 3.7 days; P =.007). Delivery was delayed by more than 7 days in 18 of 23 subjects (78%) in the intravaginal indomethacin group compared with ten (43%) in the intrarectal plus oral indomethacin group (P =.03). Birth weights were significantly higher (2306 +/- 436 versus 1862 +/- 232 g; P =.002) and hospitalization in a neonatal intensive care unit (NICU) (3.1 +/- 0.8 versus 9.3 +/- 3. 7 days; P =.001) and mechanical ventilation (1.4 +/- 0.2 versus 5.3 +/- 1.6 days; P =.001) were significantly shorter in the intravaginal indomethacin group. CONCLUSION: Intravaginal indomethacin is more effective than intrarectal plus oral application in delaying preterm labor and is associated with higher birth weights, shorter NICU stays, and shorter intervals of mechanical ventilation. PMID- 10725477 TI - Depression and anxiety in early pregnancy and risk for preeclampsia. AB - OBJECTIVE: To examine whether depression and anxiety in early pregnancy are associated with preeclampsia in an unselected nulliparous population. METHODS: In this prospective population-based study during pregnancy at outpatient maternity clinics in the Helsinki metropolitan area, depression was assessed by a Finnish modification of the short form of the Beck Depression Inventory and anxiety by one established question. Preeclampsia was defined as elevated blood pressure (BP) (more than 140/100 mmHg) and proteinuria (0.3 g during 24 hours or more). Age, smoking, alcohol consumption, marital status, socioeconomic status, and bacterial vaginosis were analyzed as potentially confounding factors in a multiple logistic regression analysis. RESULTS: Six hundred twenty-three consecutive nulliparous women with singleton pregnancies were studied at ten to 17 (median 12) weeks' gestation and at delivery. Of them, 28 (4.5%) women developed preeclampsia. Depression (mean Beck score 4.5, range 3-17) was observed in 185 (30%), women and anxiety was observed in 99 (16%) in early pregnancy. In multivariate analysis, after adjustment for potentially confounding factors, depression was associated with increased risk (odds ratio [OR] 2.5; 95% confidence interval [CI] 1.1, 5.4) for preeclampsia, as was anxiety (OR 3.2; 95% CI 1.4, 7.4). Either depression or anxiety, or both, were associated with increased risk (OR 3.1; 95% CI 1.4, 6.9) for preeclampsia. Bacterial vaginosis together with depression was associated with increased risk (OR 5.3; 95% CI 1.8, 15.0) for preeclampsia. CONCLUSION: Depression and anxiety in early pregnancy are associated with risk for subsequent preeclampsia, a risk further increased by bacterial vaginosis. PMID- 10725478 TI - Impaired vasoconstriction in pregnancy-induced hypertension assessed using doppler fluximetry. AB - OBJECTIVE: To determine whether there is a difference in peripheral vascular reactivity between normal women and those with pregnancy-induced hypertension. METHODS: Capillary blood flow (flux) was recorded in the skin over the ankle in 26 pregnant women with pregnancy-induced hypertension at term. Twelve of these women had proteinuria, and 14 were nonproteinuric. Leg lowering was used to activate the venoarteriolar reflex, and the resultant change in flux, expressed as a percentage change from the baseline, was used as an index of vascular reactivity. The results were compared with those of a control group comprising 23 matched normotensive women. The study was repeated on all of the women after delivery. RESULTS: Women with hypertension showed a median (range) increase in flux of +24.4% (-15.5% to +151.1%), significantly different from controls: -39.3% (-80.9% to -4.3%, P <.001). This difference persisted regardless of the presence or absence of proteinuria. Responses in women with pregnancy-induced hypertension were significantly different after delivery (median -60.7%; range -158.5% to 19.5%, P <.001) when compared with predelivery responses. Similar changes as a result of delivery were seen in women with proteinuric (medians +25.9% and -57. 9%, P <.002) and nonproteinuric (medians +7.8% and -62.8%, P <.001) hypertension but not in controls. Postdelivery responses in women with hypertension were no different from those of controls. CONCLUSION: Women with pregnancy-induced hypertension have abnormal cutaneous vascular reactivity that returns to normal after delivery. PMID- 10725479 TI - Effect of a screening-based prevention policy on prevalence of early-onset group B streptococcal sepsis. AB - OBJECTIVE: To assess the effectiveness and feasibility of implementing the Centers for Disease Control and Prevention (CDC) screening-based guidelines for preventing early-onset group B streptococcal sepsis. METHODS: We compared prevalence of early-onset group B streptococcal sepsis after institution of the CDC screening-based protocol (October 1, 1995 through August 31, 1999) with that of historical controls (January 1, 1992 through June 30, 1995). We reviewed medical records for a cohort of deliveries of at least 23 weeks' gestation (January 1, 1996 through December 31, 1996) for group B streptococcal colonization status, risk factors, and intrapartum antibiotic prophylaxis. RESULTS: The prevalence of early-onset group B streptococcal sepsis was 1.16 per 1000 (36 of 31, 133) live births before and 0.14 per 1000 (four of 28,733) live births after institution of the CDC protocol (P <.001). Maternal colonization was known for 95.3% of the 7168 women who delivered (January 1, 1996 through December 31, 1996) at or after 37 weeks' gestation. Of 2174 women who qualified for intrapartum antibiotic prophylaxis, 1871 (86.1%) received it before delivery. There was 93. 8% compliance with intrapartum antibiotic prophylaxis for women who delivered vaginally and 53.2% compliance for women who delivered by cesarean. CONCLUSION: Institution of the CDC screening-based protocol was accomplished at a specialty women's hospital, staffed by full-time faculty and community physicians, with 93.8% compliance for vaginal deliveries, and was associated with an 88% reduction in early-onset group B streptococcal sepsis. PMID- 10725480 TI - Prediction of birth weight by ultrasound in the third trimester. AB - OBJECTIVE: To compare the accuracy of predicted birth weight by the gestation adjusted projection method using ultrasonographic measurements obtained just before and at term. METHODS: The study group comprised patients with singleton pregnancies who underwent sonograms between 34.0 and 36.9 weeks' gestation (period 1) and at 37 weeks and beyond (period 2). The mean error in birth weight prediction, absolute birth weight error, and signed and absolute percent errors were compared with paired t tests. Thus, each patient served as her own control. RESULTS: The study included 138 patients undergoing 276 sonograms. The mean absolute error of the predicted birth weight was smaller for period 1 than for period 2 (197 +/- 167 g compared with 235 +/- 209 g, P =.019). The mean absolute percent error was 6.2 +/- 5.2% for period 1 compared with 7.4 +/- 6.3% for period 2 (P =.019). These same trends were observed when fetuses with suspected growth abnormalities were examined separately. Averaging data from both gestational periods did not improve the prediction of birth weight. CONCLUSION: Sonograms between 34.0 and 36. 9 weeks' gestation allow for more accurate prediction of birth weight than sonograms later in gestation. Though these differences are small and not clinically significant, this study indicates that serial sonograms in the late third trimester do not improve the ability to predict birth weight, even in abnormally grown fetuses. A single sonogram between 34 and 37 weeks' gestation is recommended for prediction of birth weight. PMID- 10725481 TI - Determinants of long-term hormone replacement therapy and reasons for early discontinuation. AB - OBJECTIVE: To identify factors associated with long-term hormone replacement therapy (HRT) and reasons for early discontinuation of it. METHODS: A cross sectional study was conducted in four United Kingdom group general practices. Six hundred fifteen past or present HRT users (representing a response rate of 66%) responded to questionnaires on HRT and potential determinants of long-term use. Main outcome measures were long-term HRT use (at least 6 years) as opposed to short-term use (at most 2 years) and self-reported reasons for discontinuation. Odds ratios (ORs) of long-term use were adjusted for age and other variables, in the same groups, calculated by logistic regression and 95% confidence intervals (CIs). RESULTS: Ovariectomy (OR 2.59, 95% CI 1.12, 5.97), hysterectomy (OR 2.28, 95% CI 1.37, 3.79), previous oral contraceptive use (OR 1.76, 95% CI 1. 03, 3.01), HRT prescription to prevent osteoporosis (OR 1.81, 95% CI 1.04, 3.13), opinion that HRT prevents health problems (OR 3.22, 95% CI 1.57, 6.63), opinion that HRT is associated with health risks (OR 0.23, 95% CI 0.08, 0.65), and opinion that HRT has cosmetic benefits (OR 2.52, 95% CI 1.45, 4.40) were statistically significantly associated with long-term HRT. Women surveyed most often reported side effects and weight gain (each about 30%) as reasons for discontinuation, followed by possible health risks and dislike of menstrual bleeding or hormones (each about 15%). CONCLUSION: Ovariectomy, hysterectomy, and opinions about benefits and disadvantages of HRT were the most important determinants of long-term use, whereas women themselves mentioned side effects and weight gain most frequently as reasons for discontinuing it. PMID- 10725482 TI - Breast cancer diagnosed during hormone replacement therapy. AB - OBJECTIVE: Hormone replacement therapy (HRT) is associated with decreased breast cancer mortality despite increased incidence. We studied postmenopausal breast cancer patients to determine whether this paradox results from earlier diagnosis, biologically less aggressive tumors, or cessation of hormonal stimulation. METHODS: Demographic, clinical, pathologic, treatment, and outcome information for 455 postmenopausal breast cancer patients who had not used postmenopausal hormones was compared with that of 47 breast cancer patients who used postmenopausal hormones prior to diagnosis. RESULTS: Hormone users were significantly younger, more often white, and of lower body mass index than nonusers. Hormone users presented significantly more often with nonpalpable mammographic findings, resulting in significantly smaller tumors with less nodal involvement than nonusers. Cancers of hormone users were more commonly invasive lobular or in situ ductal and were more likely to be steroid receptor positive. Hormone users were treated with breast conservation significantly more frequently than nonusers. These differences persisted after matching for age and year of surgery and after controlling for race. At 5 years, none of the hormone users with invasive cancers had local recurrence compared with 8% of nonusers, and 7% of users had distant disease compared with 10% of nonusers. CONCLUSION: These results indicate that favorable breast cancer survival after postmenopausal hormone use might result from earlier detection through mammography. Possible hormonal influence on tumor biology and prognosis was not supported by our data. PMID- 10725483 TI - Homocysteine and folate levels as risk factors for recurrent early pregnancy loss. AB - OBJECTIVE: To estimate the relative risk of recurrent early pregnancy loss for different total plasma homocysteine and serum folate concentrations. METHODS: In a case-control study, we measured homocysteine (fasting and afterload), folate (serum and red cells), pyridoxal 5'-phosphate, and cobalamin concentrations in 123 women who had at least two consecutive spontaneous early pregnancy losses each and compared concentrations with those of 104 healthy controls. RESULTS: Women with recurrent early pregnancy losses had significantly lower serum folate concentrations than controls, whereas the other measurements were similar to those of controls. Elevated homocysteine, fasting greater than 18.3 micromol/L and afterload greater than 61.5 micromol/L, was a risk factor for recurrent early pregnancy loss, with odds ratios (ORs) and 95% confidence intervals (95% CIs) of 3.6 (1.2, 12.7) and 2.7 (0.9, 8.8) in the group with recurrent miscarriages: 6.4 (1.9, 24.3) and 4.3 (1. 2, 17.3) in primary aborters, and 4.2 (1.3, 15.4) and 3.4 (1.0, 12. 8) in those with three or more miscarriages. The ORs (95% CIs) in the same study populations for serum folate concentrations less than 8.4 nmol/L were 2.1 (0.9, 4.8), 2.7 (1.0, 7.8), and 3.2 (1.3, 8.1), respectively. A significant dose-response relationship between serum folate concentrations and risk of recurrent early pregnancy loss suggested a protective effect by high serum folate concentrations. CONCLUSION: Elevated homocysteine and reduced serum folate concentrations were risk factors for recurrent spontaneous early pregnancy losses. Folic acid supplementation might be beneficial in women with histories of early pregnancy loss. PMID- 10725484 TI - Influence of human immunodeficiency virus infection on pelvic inflammatory disease. AB - OBJECTIVE: To examine the influence of human immunodeficiency virus (HIV) infection on clinical and microbiologic characteristics of pelvic inflammatory disease (PID). METHODS: Forty-four HIV-infected women and 163 HIV noninfected women diagnosed with PID by standard case definition were evaluated by using clinical severity scores, transabdominal sonograms, and endometrial biopsies. After testing for bacterial infections, patients were prescribed antibiotics as recommended by the Centers for Disease Control and Prevention (CDC). RESULTS: Symptoms of PID and analgesic use before enrollment did not differ by HIV serostatus. More HIV-infected women had received antibiotics before enrollment (40.9% versus 27.2%, P =.08), a factor associated with milder signs regardless of serostatus. More HIV-infected women had sonographically diagnosed adnexal masses at enrollment (45.8% versus 27.1%, P =.08), a difference that yielded higher median severity scores (17.5 of 42 points versus 15 of 42 points, P =.07). However, those differences were not significant at the P <.05 level. Mycoplasma (50% versus 22%, P <.05) and streptococcus species (34% versus 17%, P <.05) were isolated more commonly from biopsies of HIV-infected women. Within 30 days after enrollment, HIV-infected women generally responded as well to therapy as HIV noninfected women did, regardless of initial CD4 T-lymphocyte percentage. CONCLUSION: Among women with acute PID, HIV infection was associated with more sonographically diagnosed adnexal masses. Clinical response to CDC-recommended antibiotics did not differ appreciably by serostatus. Mycoplasmas and streptococci were isolated more commonly from HIV-infected women, but those organisms also might be associated with PID in immunocompetent women. PMID- 10725485 TI - Differential effects of cough, valsalva, and continence status on vesical neck movement. AB - OBJECTIVE: We tested the null hypothesis that vesical neck descent is the same during a cough and during a Valsalva maneuver. We also tested the secondary null hypothesis that differences in vesical neck mobility would be independent of parity and continence status. METHODS: Three groups were included: 17 nulliparous continent (31.3 +/- 5.6; range 22-42 years), 18 primiparous continent (30.4 +/- 4.3; 24-43), and 23 primiparous stress-incontinent (31.9 +/- 3.9; 25-38) women. Measures of vesical neck position at rest and during displacement were obtained by ultrasound. Abdominal pressures were recorded simultaneously using an intravaginal microtransducer catheter. To control for differing abdominal pressures, the stiffness of the vesical neck support was calculated by dividing the pressure exerted during a particular effort by the urethral descent during that effort. RESULTS: The primiparous stress-incontinent women displayed similar vesical neck mobility during a cough effort and during a Valsalva maneuver (13.8 mm compared with 14.8 mm; P =.49). The nulliparous continent women (8.2 mm compared with 12.4 mm; P =. 001) and the primiparous continent women (9.9 mm compared with 14.5 mm; P =.002) displayed less mobility during a cough than during a Valsalva maneuver despite greater abdominal pressure during cough. The nulliparas displayed greater pelvic floor stiffness during a cough compared with the continent and incontinent primiparas (22.7, 15.5, 12.2 cm H(2)O/mm, respectively; P =.001). CONCLUSION: There are quantifiable differences in vesical neck mobility during a cough and Valsalva maneuver in continent women. This difference is lost in the primiparous stress-incontinent women. PMID- 10725486 TI - Outcome and reproductive function after conservative surgery for borderline ovarian tumors. AB - OBJECTIVE: To study reproductive function and disease outcome in women with borderline ovarian tumors who were treated with conservative surgery. METHODS: Patients with borderline ovarian tumors were identified from institutional databases. Patients were eligible if they had pathologically confirmed borderline ovarian tumors, no prior sterilization, no history of radiation therapy, retained their uterus and ovarian tissue, and were younger than age 45. Information was acquired by retrospective medical record review and patient interview. RESULTS: Forty-three patients met the eligibility criteria. The median age was 25 years, with a range of 15-39 years. Twenty-six patients had serous tumors, and 17 had mucinous tumors. Fifteen had stage I disease, three had stage III, and 25 were unstaged. Follow-up was available for all patients (median, 5.7 years). Twenty nine remained disease-free, and 14 developed a new primary lesion/recurrence, with a median time to recurrence of 39.3 months. Recurrence was more frequent in patients treated with ovarian cystectomy than in those treated with oophorectomy alone (58% compared with 23%) (P <.04). After treatment, 29 of 36 patients (81%) retained normal menstrual cycles, and 12 of 24 patients attempting pregnancy conceived 25 pregnancies. Most patients were highly satisfied with conservative surgery. CONCLUSION: Conservative surgery remains a therapeutic option in selected patients with borderline ovarian tumors. Although the rate of new lesion/recurrence is relatively high, especially in those treated with ovarian cystectomy, mortality from cancer remains low. Many patients who desire pregnancy are able to conceive and deliver healthy offspring after conservative surgery. PMID- 10725487 TI - Angiogenesis in early-invasive and low-malignant-potential epithelial ovarian carcinoma. AB - OBJECTIVE: To evaluate angiogenesis in ovaries of women with stage I invasive and low-malignant-potential epithelial ovarian carcinoma. METHODS: Ovarian specimens of 49 consecutive women with primary stage I invasive (n = 15) or stage I low malignant-potential epithelial ovarian carcinoma (n = 34) were stained immunohistochemically for factor VIII-related antigen. Microvessel counts were tested for correlation with patient age, race, parity, previous oral contraceptive use, histologic type, tumor grade, tumor size, ascites, tumor excrescences, and disease-free and overall survival. Statistical analysis included multiple linear regression, Student t tests, factorial analysis of variance, and Cox proportional hazards regression, with P <.05 considered statistically significant. RESULTS: Microvessel counts of ovarian specimens of women with stage I invasive epithelial ovarian carcinoma (median 30, range 17-73) were significantly higher than those of women with stage I low-malignant potential epithelial ovarian carcinoma (median 10, range 5-23), (P <.001). Among women with low-malignant-potential disease, microvessel counts did not differ significantly between serous and mucinous carcinomas (median 10, range 5-23 versus median 11, range 5-20, respectively, P =.78). There was no correlation between microvessel counts and age, tumor grade, tumor size, ascites, or tumor excrescences. CONCLUSION: Angiogenesis as assessed by microvessel counts is more intense in stage I invasive ovarian epithelial carcinoma compared with stage I low-malignant-potential carcinoma, and might assist in differentiating between these histopathologic entities. PMID- 10725488 TI - Accuracy of lymph node palpation to determine need for lymphadenectomy in gynecologic malignancies. AB - OBJECTIVE: To assess the accuracy of intraoperative lymph node palpation for identifying lymph node metastasis in gynecologic malignancies. METHODS: We prospectively evaluated 126 women who had lymphadenectomies for staging of various gynecologic malignancies from August 1995 to June 1997. Surgeries were done by obstetrician-gynecologists with subspecialty certification in gynecologic oncology from the American Board of Obstetrics and Gynecology, who had practiced gynecologic oncology for at least 5 years. Data were collected on gynecologic oncologists' opinions of lymph node status by palpation. Nodes believed to be positive were sent separately. We recorded operating time for lymphadenectomies, and intraoperative and postoperative complications. RESULTS: Mean (range) patient age was 55 (18-83) years. Mean (range) operating time was 188 (85-435) minutes. The mean (range) lymphadenectomy time was 46 (5-150) minutes. The total number of lymph nodes dissected was 2138. One hundred seven of 2138 (5%) nodes were positive for malignancy. Thirty-eight of 107 (36%) positive lymph nodes were missed by palpation. Fifty-six of 2031 (3%) negative lymph nodes were selected as positive. Sixty-nine of 107 (64%) positive lymph nodes were identified correctly. Sensitivity and specificity of palpation were 72% and 81%, respectively. The positive and negative predictive values of lymph node palpation were 56% and 89%, respectively. CONCLUSION: Intraoperative lymph node palpation has low sensitivity and positive predictive value even when done by experienced board-certified gynecologic oncologists. PMID- 10725489 TI - Risks of funipuncture in fetuses with single umbilical arteries. AB - OBJECTIVE: To estimate procedure-related risks of funipuncture in fetuses with single umbilical arteries (UAs). METHODS: We identified fetuses that had blood samples collected by funipuncture and in which single UAs were detected, prenatally or postnatally. We also recorded maternal demographics, prenatal sonographic findings, gestational age at the time of the procedure, procedure related complications, and perinatal outcomes. RESULTS: Over 2 years, 14 fetuses identified as having single UAs had funipuncture for prenatal karyotyping at a median gestational age of 29 weeks (range 20-34 weeks). Each had additional abnormal prenatal sonographic findings. The approach to the cord was transplacental in six cases and transamniotic in eight. There were no failed procedures, and 13 of 14 funipunctures were successful on the first attempt. Three fetuses (21%) had complications (bradycardia in two cases and bleeding in one case), a complication rate not greater than that reported in large series of fetuses that had fetal blood sampling. All three complications were associated with the transamniotic approach. There were no procedure-related pregnancy losses within 2 weeks of the procedure in this series. CONCLUSION: Funipuncture does not seem to be associated with increased risk of procedure-related complications or pregnancy losses in fetuses with single UAs, although the risk could be greater with transamniotic than with transplacental sampling. PMID- 10725490 TI - Maternal smoking and fetal erythropoietin levels. AB - OBJECTIVE: To determine the influence of maternal smoking on fetal erythropoietin concentrations in health term pregnancies and test the correlation between cotinine, a biomarker of maternal smoking, and erythropoietin levels in fetuses. METHODS: We invited women with healthy term pregnancies to participate in the study, excluding those with conditions previously known to be associated with elevated fetal erythropoietin levels. We recorded demographic data, smoking status, and labor outcome prospectively for each patient. Umbilical venous samples were collected, and serum was stored at -70C to be analyzed later for erythropoietin and cotinine. Umbilical arterial samples were tested for pH and base excess determination. We compared fetal erythropoietin and cotinine between smokers and nonsmokers and examined correlations between erythropoietin and cotinine. Kruskal-Wallis test, t test, median test, and Spearman rank correlation test were used when appropriate. Statistical significance was P <.05. RESULTS: We recruited 35 nonsmokers and 26 smokers and analyzed their samples. The two groups were comparable in demographics and birth outcomes, except for birth weights, which were lower in smokers. Fetal erythropoietin concentrations increased significantly with increasing maternal cigarette consumption, ranging from none to more than 15 cigarettes per day (P =.03). There was positive correlation between fetal erythropoietin and cotinine concentrations (r =.41; P =.04), suggesting a dose-response relationship. CONCLUSION: Fetuses of smokers had increased erythropoietin concentrations that correlate positively with fetal cotinine levels; which suggests an increased risk of subacute hypoxia related to degree of maternal cigarette consumption. PMID- 10725491 TI - Low fetal oxygen saturation at birth and acidosis. AB - OBJECTIVE: To measure umbilical cord blood oxygen saturation, to calculate preductal oxygen saturation at birth, and to assess its predictive value for acidosis. METHODS: Umbilical cord blood samples of 1537 live-born singleton neonates were analyzed. Oxygen saturation was measured by spectrophotometry; pH and base excess were measured by a pH and blood gas analyzer. Preductal oxygen saturation was calculated with an empirical equation. Acidosis was defined as 2 standard deviations (SDs) below the mean of umbilical artery (UA) pH or base excess (7.09 and -10.50 mmol/L, respectively). The predictive value for acidosis of UA and umbilical vein (UV) oxygen saturation and calculated preductal oxygen saturation was determined with receiver operating characteristic curves. RESULTS: The mean values (+/-SD) of UV, UA, and calculated preductal oxygen saturation were 52 +/- 18%, 26 +/- 17%, and 31 +/- 16%, respectively. Forty-seven neonates had UA pH less than 7.09 and 60 had UA base excess less than -10.50 mmol/L. The UV, UA, and calculated preductal oxygen saturation showed considerably weaker relations to UA base excess (multiple r(2) =.056,.003, and.017, respectively; P <.001) than to UA pH (multiple r(2) =.112,.126, and.148, respectively; P <. 001). Receiver operating characteristic areas under the curve were higher when predicting low pH compared with low base excess (for UV, UA, and calculated preductal oxygen saturation: 0.716 versus 0.699, 0.747 versus 0.586, and 0.765 versus 0.628, respectively). The difference was significant for UA oxygen saturation (P <.05). All tests showed high sensitivity and negative predictive values, but low specificity and positive predictive values. CONCLUSION: Low fetal oxygen saturation measured at birth seemed to be associated with low fetal pH and base excess values, but its predictive value for acidosis in an unselected population was limited, particularly if acidosis was metabolic. PMID- 10725492 TI - Vasa previa: prenatal diagnosis, natural evolution, and clinical outcome. AB - OBJECTIVE: To describe the prenatal ultrasonographic diagnosis, natural evolution, and clinical outcomes of vasa previa in a large population at a single institution. METHODS: We attempted to view the internal cervical os of 93,874 women with second- and third-trimester pregnancies during an 8-year period. Echogenic parallel or circular lines near the cervix, seen by gray-scale ultrasonography, raised the possibility of vasa previa. Diagnosis was confirmed by Doppler and endovaginal studies if aberrant vessels over the internal cervical os were suspected. Abnormal placental morphology and velamentous cord insertion were documented if they were identified during prenatal scans. Ultrasonographic findings were correlated with clinical courses, perinatal outcomes, and placental pathology examinations. RESULTS: Eighteen cases of vasa previa were suspected at a mean (+/- standard deviation) gestational age of 26.0 +/- 6.3 weeks; the earliest diagnosis was at 15.6 weeks' gestation. Eight of those cases initially showed placental edge over the internal os and later developed vasa previa after the placenta "receded" from the cervix. Six women had mild vaginal bleeding at a mean gestational age of 31.3 weeks. Three women had normal late third-trimester scans and were allowed to have uncomplicated vaginal deliveries. The remaining subjects delivered by cesarean. There were two deaths (one fetal and one neonatal), and minor preterm complications slightly prolonged infant hospitalizations. One set of preterm twins needed neonatal transfusions. Pathology findings included ten cases of velamentous insertion and three cases each of bilobed placentas, succenturiate lobes, and marginal cord insertion. CONCLUSION: Vasa previa was detected in asymptomatic women as early as the second trimester. Perinatal outcome was generally favorable, although several infants had slightly extended newborn nursery admissions due to mild complications of prematurity. PMID- 10725493 TI - Cost-benefit analysis of prenatal diagnosis for Down syndrome using the British or the American approach. AB - OBJECTIVE: To compare the cost and benefits of prenatal diagnosis for Down syndrome using the British and American approaches. METHODS: This cost-benefit analysis was based on a decision-analytic approach. The British strategy included screening by a first-trimester ultrasound at 10-14 weeks for nuchal translucency thickness, and the American strategy included only second-trimester screening by using maternal age and maternal serum screening. The key probabilities of the decision-tree analysis and all cost estimates were based on American standards. The best scenario of the British strategy assumed ultrasound nuchal translucency thickness sensitivity (for detecting Down syndrome) of 80% and a false-positive rate of 5% and the worst scenario assumed a sensitivity of 50% and a false positive rate of 10%. The results were expressed in annual costs based on approximately 4 million births per year in the United States. RESULTS: As compared with do-nothing, the American strategy was found to allow savings of approximately $96 million per year and the best scenario for the British strategy was savings of approximately $5 million per year. The financial costs of the British and American strategies would be comparable only if the first-trimester ultrasound had a sensitivity of 80% and a false-positive rate of 5% in detecting Down syndrome. CONCLUSION: The British strategy does not appear to be economically beneficial in the United States even under the most ideal scenarios of ultrasound accuracy. PMID- 10725494 TI - Regulation of interleukin-8 synthesis in human lower uterine segment fibroblasts by cytokines and growth factors. AB - OBJECTIVE: To investigate the influence of lipopolysaccharide, cytokines, growth factors, and progesterone on the synthesis of interleukin-8 by human lower uterine segment fibroblasts. METHODS: Fibroblasts derived from a lower uterine segment biopsy specimen obtained from a woman undergoing elective cesarean delivery at term were exposed to lipopolysaccharide, interleukin-1beta, transforming growth factor-beta(1), platelet-derived growth factor-AB, and combinations of these substances. All experiments were performed in the absence and presence of progesterone. The concentration of interleukin-8 in the culture medium was determined by enzyme immunoassay after 24 hours. RESULTS: Compared with controls (0.71 +/- 0.04 ng interleukin-8/10(6) cells), fibroblasts exposed to lipopolysaccharide, transforming growth factor-beta(1), or platelet-derived growth factor-AB exhibited no increase, or at most, only a minor but significant increase, in interleukin-8 secretion. Incubation with interleukin-1beta led to a moderate increase, whereas the combinations interleukin-1beta/transforming growth factor-beta(1) (105.0 +/- 7.5 ng interleukin-8/10(6) cells) and interleukin 1beta/platelet-derived growth factor-AB (387.3 +/- 25.6 ng interleukin-8/10(6) cells) increased interleukin-8 secretion dramatically. No further increase was observed with the combination interleukin-1beta/platelet-derived growth factor AB/transforming growth factor-beta(1). When progesterone was added, interleukin-8 secretion decreased significantly by 16-34%, depending on the stimulator, or did not change. CONCLUSION: The findings indicate that interleukin-8 secretion by human lower uterine segment fibroblasts in vitro is upregulated by interleukin 1beta, transforming growth factor-beta(1), and platelet-derived growth factor-AB in a synergistic fashion. Because interleukin-8 mediates the invasion of neutrophils into the cervical stroma, this may be an important mechanism controlling cervical dilatation during parturition. PMID- 10725495 TI - Lack of progress in labor as a reason for cesarean. AB - OBJECTIVE: To estimate the prevalence of lack of progress in labor as a reason for cesarean delivery and to compare published diagnostic criteria with the labor characteristics of women with this diagnosis. METHODS: We reviewed medical records and did a postpartum telephone survey to collect data from 733 women who delivered full-term, nonbreech infants by unplanned cesarean between March 1993 and February 1994. These were a subset of 2447 births sampled at delivery from 30 hospitals in Los Angeles County and Iowa. We measured the proportion of unplanned cesareans done for lack of progress in labor, the cervical dilatation at the time of cesarean, length of the second stage, and slope of the active phase among the women. We estimated the proportion of these cesareans that conformed to the ACOG criteria for the diagnosis of lack of progress. RESULTS: Lack of progress was a reason for 68% of unplanned, vertex cesareans. At least 16% of the subjects who had cesareans for lack of progress were in the latent phase of labor according to ACOG criteria. The second stage was not prolonged in 36% of the women who delivered at 10 cm. CONCLUSION: Lack of progress in labor is a dominant reason for cesarean delivery. Many cesareans are done during the latent phase of labor, and in the second stage of labor when it is not prolonged. These practices do not conform to published diagnostic criteria for lack of progress. PMID- 10725496 TI - Predicting incomplete uterine rupture with vaginal sonography during the late second trimester in women with prior cesarean. AB - OBJECTIVE: To evaluate the usefulness of serial transvaginal ultrasonographic measurement of the thickness of the lower uterine segment in the late second trimester for predicting the risk of intrapartum incomplete uterine rupture in women with previous cesarean delivery. METHODS: Serial transvaginal ultrasonography with full bladder was performed in 374 women without previous cesarean delivery (control group) and 348 women with previous cesarean delivery (cesarean group) from 19 to 39 weeks' gestation. The thickness of the lower uterine segment was measured in the longitudinal plane of the cervical canal. RESULTS: The thickness of the lower uterine segment decreased from 6.7 +/- 2.4 mm (mean +/- standard deviation [SD]) at 19 weeks' gestation to 3.0 +/- 0.7 mm at 39 weeks' gestation in the control group, but the thickness was more than 2.0 mm throughout this period in each control subject. In the cesarean group, the thickness decreased from 6.8 +/- 2.3 mm at 19 weeks' to 2.1 +/- 0.7 mm at 39 weeks' gestation and was significantly thinner than that of the control group after 27 weeks' gestation (P <.05). Eleven of 12 women (91%) with lower uterine segment less than the mean control - 1 SD in the late second trimester had a very thin lower uterine segment at cesarean delivery with fetal hair being visible through the amniotic membrane, ie, incomplete uterine rupture. In 17 of 23 women (74%) with lower uterine segment less than 2.0 mm in thickness within 1 week (4 +/- 3 days) before repeat cesarean delivery, intrapartum incomplete uterine rupture developed. CONCLUSION: Transvaginal ultrasonography is useful for measurement of the uterine wall after previous cesarean delivery. PMID- 10725497 TI - Rapid enzymatic urine screening test to detect bacteriuria in pregnancy. AB - OBJECTIVE: To determine the sensitivity, specificity, and positive and negative predictive values of an enzymatic urine screening test for diagnosing bacteriuria in pregnancy. METHODS: Clean-catch midstream urine samples were collected from 383 women who had routine prenatal screening for bacteriuria. Sensitivity, specificity, and positive and negative predictive values for each screening test (enzyme activity, nitrites or leukocytes on dipstick, and bacteria or pyuria on microscopic examination) were estimated using urine culture as the criterion standard. Urine cultures were considered positive if they grew 10(4) colony forming units of a single uropathogen. Standard deviations used to calculate 95% confidence intervals were based on binomial distribution. A sample of 30 urine specimens was selected to evaluate interrater agreement using Cohen's kappa statistic. RESULTS: Five of 383 samples were contaminated, leaving 378 samples for evaluation. Thirty of 43 specimens with positive urine culture had positive enzyme activity. Of 335 samples with no growth, 150 had negative enzyme activity. Sensitivity, specificity, and positive and negative predictive values for the Uriscreen enzymatic screening test (Bard Patient Care Division, Murray Hill, NJ) were 70%, 45%, 14%, and 92%, respectively. Sensitivity of the Uriscreen was lower than that of bacteria alone. Interrater agreement for Uriscreen testing was high among the three testers (kappa =.86). CONCLUSION: The Uriscreen enzymatic screening test had inadequate sensitivity for rapid screening for bacteriuria in pregnancy. PMID- 10725498 TI - A view of reviews. PMID- 10725499 TI - Effect of prolonged asphyxia on skin blood flow in fetal lambs. AB - OBJECTIVE: To determine the effect of a prolonged period of asphyxia on skin blood flow, a potential indicator of fetal cardiovascular responses to asphyxia, in the chronically catheterized fetal lamb. METHODS: Eight chronically instrumented pregnant ewes were studied at 118 +/- 1 days' gestation. After a control period, fetal acid-base status was assessed and regional blood flows were determined with dye-labeled microspheres. Fetal asphyxia was then induced by partial umbilical cord occlusion, decreasing fetal arterial oxygen pressure to 15 torr while maintaining pH above 7.28. Fetal cardiovascular status was monitored continuously. Fetal acid base status was evaluated every 10-15 minutes during cord occlusion. Regional blood flow determinations were repeated after 90 minutes of stable asphyxia. Results are expressed as the mean +/- standard error. Student t test for paired data was used to compare hemodynamic, acid-base, and regional blood flow determinations before cord occlusion and after 90 minutes of asphyxia. RESULTS: There was a significant increase in blood flow to the fetal scalp from a control value of 52 +/- 8 mL per minutes per 100 g to 175 +/- 30 mL per minute per 100 g at 90 minutes of asphyxia (P =.01). Similarly, there was an increase in blood flow to the skin overlying the fetal hindquarter from 39 +/- 12 mL per minute per 100 g during control to 153 +/- 47 mL per minute per 100 g at asphyxia (P =.038). CONCLUSION: In the chronically instrumented fetal lamb, a 90-minute period of asphyxia produced by partial cord occlusion resulted in a significant increase in blood flow to the fetal skin. PMID- 10725500 TI - Much ado about a little cut: is episiotomy worthwhile? AB - Methods to prevent perineal trauma during childbirth include avoiding episiotomy and forceps delivery and slowing delivery of the head to allow the perineum time to stretch. Each intervention can lengthen the second stage of labor and change the biophysical stresses on infants and the pelvic floor. Available evidence supports the belief that the interventions are safe for infants and do not lead to significant short- or long-term maternal morbidity. We should abandon the conventional teaching that a longer second stage and perineal stretching are harmful. Routine episiotomy is no longer advisable. Forces that might inhibit physicians from practicing evidence-based techniques of obstetric delivery include time pressures, malpractice concerns, lack of experience with slow perineal stretching, and an interventionist practice pattern. Changes in practice can be effectively introduced through consumer pressures, opinion leaders, and in teaching institutions, by house staff. PMID- 10725501 TI - The mentor effect in student evaluation. AB - OBJECTIVE: To determine if faculty mentors rate their mentored students higher than do nonmentors, and to ascertain if gender is a factor. METHODS: All third year students (n = 101) from academic years 1996-1998, who performed their obstetrics and gynecology clerkship at the Milton S. Hershey Medical Center in the Penn State Geisinger Health System and were evaluated by full-time faculty (n = 18), were included in the study (total observations = 545). Students were rated by faculty on an ordinal scale in five categories. Generalized estimating equation methodology was used to fit proportional odds models for ordinal data to assess whether there were statistically significant mentor or faculty/student gender effects. RESULTS: Student evaluations from mentors were more likely to have better scores than student evaluations from nonmentoring-faculty for all five categories (all P <.01). The odds ratios (OR) for the mentor effect ranged from 2.1 (95% confidence interval [CI] 1.4, 3.2) for fund of knowledge to 3.2 (95% CI [2.1, 4. 8]) for attitude. For problem-solving and technical skills, male faculty were more likely than female faculty to give male students better scores (problem-solving skills: odds ratio [OR] = 1.7, 95% CI [1.0, 2.7]; technical skills: OR = 2.2, 95% CI [1.1, 4.6]). Mentoring-faculty evaluations were not strongly correlated with the students' objective examination scores. CONCLUSION: Overall, mentors score their mentored students statistically higher than do nonmentors. Gender differences in evaluation, while present, are less consistent and smaller than the mentor effect. PMID- 10725502 TI - Working conditions and adverse pregnancy outcome: a meta-analysis. AB - OBJECTIVE: To evaluate the association between working conditions and adverse pregnancy outcomes by performing a meta-analysis of published studies. DATA SOURCES: We searched the English-language literature in MEDLINE through August 1999 using the terms standing, posture, work, workload, working conditions, shift, occupational exposure, occupational diseases, lifting, pregnancy complications, pregnancy, small for gestational age (SGA), fetal growth retardation (FGR), preterm, and labor. METHODS OF STUDY SELECTION: We included observational studies evaluating the effect of one or more of the following work related exposures on adverse pregnancy outcome: physically demanding work, prolonged standing, long work hours, shift work, and cumulative work fatigue score. Outcomes of interest were preterm birth, hypertension or preeclampsia, and SGA.We conducted a meta-analysis based on 160,988 women in 29 studies to evaluate the association of physically demanding work, prolonged standing, long working hours, shift work, and cumulative work fatigue score with preterm birth. Also analyzed were the associations of physically demanding work with hypertension or preeclampsia and SGA infants. The data were analyzed using the Peto-modified Mantel-Haenszel method to estimate the pooled odds ratios (ORs) and 95% confidence intervals (CIs). TABULATION, INTEGRATION, AND RESULTS: Physically demanding work was significantly associated with preterm birth (OR 1.22, 95% CI 1.16, 1. 29), SGA (OR 1.37, 95% CI 1.30, 1.44), and hypertension or preeclampsia (OR 1.60, 95% CI 1.30, 1.96). Other occupational exposures significantly associated with preterm birth included prolonged standing (OR 1.26, 95% CI 1.13, 1.40), shift and night work (OR 1.24, 95% CI 1.06, 1.46), and high cumulative work fatigue score (OR 1.63, 95% CI 1.33, 1.98). We found no significant association between long work hours and preterm birth (OR 1.03, 95% CI 0.92, 1.16). CONCLUSION: Physically demanding work may significantly increase a woman's risk of adverse pregnancy outcome. PMID- 10725518 TI - Dynamics of non-structural carbohydrates in developing leaves, bracts and floral buds of cotton. AB - Development of cotton (Gossypium hirsutum L.) squares (i.e. floral buds with bracts) is fundamental for yield formation. A 2-year field study was conducted to determine dry weight (DW) accumulations of cotton leaves, floral bracts and floral buds, and the changes in concentrations of non-structural carbohydrates (hexoses, sucrose and starch) in these tissues during square ontogeny as affected by fruiting positions within the plant canopy. During square development, DW accumulation of a subtending sympodial leaf and floral bracts followed a sigmoid growth curve with increasing square age, whereas the DW increase of a floral bud followed an exponential curve. Main-stem node (Node 8, 10 or 12) and branch position (proximal vs. distal) within a plant canopy significantly affected DW accumulations of the leaf, bracts and floral bud. Starch was the dominant non structural carbohydrate in the three tissues, accounting for more than 65% of total non-structural carbohydrates (TNC). Subtending leaf TNC increased as square age increased. The bracts exhibited a smaller change in TNC than leaves. Non structural carbohydrate concentration was the lowest in 10-day-old floral buds, and had little change during the first 15 days of square development. Within 5 days prior to anthesis, the floral-bud TNC increased dramatically, tripling at the time of floral anthesis compared with 15-day-old floral buds. Square age and fruiting position significantly affected non-structural carbohydrate concentrations of subtending leaves, bracts, and floral buds. The correlation did not exist between final boll retention and non-structural carbohydrate concentrations of floral buds at different fruiting positions under normal growth conditions. The pattern of floral-bud non-structural carbohydrates during square ontogeny suggests that major events in carbohydrate metabolism occur just prior to anthesis. PMID- 10725519 TI - Stress indications in copper- and nickel-exposed Scots pine seedlings. AB - Scots pine nursery seedlings were planted in pots, five seedlings per treatment, and placed in an experimental field at the University of Oulu in northern Finland at the beginning of June 1997. Copper and nickel sulphates were mixed with forest mineral soil before seedling planting. The metal levels ranged from 0 to 25 mg Ni kg(-1) dry soil and 0 to 50 mg Cu kg(-1) in dry soil and in combinations of both metals. Current year's needles for element analyses, EDS microanalyses, microscopy and glutathione and peroxidase activity analyses were collected from 1 5 seedlings per treatment in September. Seedling biomass in controls, Cu25 and Cu50 differed significantly from the Ni25Cu50 treatment. The root/shoot ratio was highest in the Ni5 treatment, indicating good root growth, though the roots were visibly healthier in the Cu25 treatment than in the Ni5 treatment. At higher Ni levels, the condition of roots deteriorated. The proportion of plasmolysed mesophyll cells was highest in the Ni25 treatment. Copper-treated seedlings did not suffer from Cu stress, because no severe injuries were seen in either the roots or the needles in Cu-exposed seedlings. The needle concentrations of Cu increased only slightly due to treatments. Ni accumulation in needles increased with increasing concentrations in soil. Needles of Cu-treated seedlings had less oxidized glutathione than those of Ni-treated seedlings, but the roots had higher, not significantly, peroxidase activity levels. Light-colored, swollen thylakoids were occasionally observed in the Ni25Cu50 treatment, indicating some interaction between Ni and Cu. Ni seemed to cause more oxidative stress to the seedlings than copper, which was manifested as a decreased GSH level and an increased proportion of GSSG in the Ni treatments. Copper together with nickel strongly decreased root growth, the root/shoot ratio being lowest in the Ni25Cu50 treatment. PMID- 10725520 TI - Impact of wet deposited nickel on the cation content of a mat-forming lichen Cladina stellaris. AB - The impact of experimentally sprayed aqueous nickel solution on the concentrations of potassium, calcium, magnesium and nickel in three horizontal strata (top, 0-20 mm; middle, 20-40 mm; and base, 40-60 mm) of the cushion forming lichen Cladina stellaris was investigated. The experimental nickel deposition range used corresponded with that from the pristine forests of the Finnish border to polluted industrial sites of Russian Kola Peninsula (0-1000 mg Ni(2+) m(-2) year(-1)). The lichen mat retained ca. 31-66% of the nickel deposited during two growing seasons and the relative retention efficiency was highest at the low deposition end. The concentrations of cations in lichen thalli were significantly reduced only after the highest nickel deposition. Furthermore, the separate horizontal strata responded differently to nickel exposure indicating that the cation exchange sites of the top stratum were not completely saturated by nickel even after the most severe treatment. However, nickel deposited in high doses caused considerable reduction in potassium concentration indicating damage to cell membranes. Episodically deposited high concentrations of nickel can probably affect membrane integrity before detectable changes in total concentrations of cations in the lichen thallus take place. Thus, ratios of total concentrations of cations in the lichen thallus are fairly insensitive to nickel deposition, which reduces the risk of compounding effects when the ratios are used to indicate long-term acid deposition in areas with multiple pollution problems such as Kola Peninsula. PMID- 10725521 TI - Changes in wheat germination following gamma-ray irradiation: an in vivo electronic paramagnetic resonance spin-probe study. AB - Embryos excised from wheat (Triticum aestivum) grains following gamma-ray irradiation at different doses were analyzed on membrane permeability by electron paramagnetic resonance (EPR) technique with 4-oxo-2, 2, 6, 6-tetramethyl-1 piperidinyloxy (TEMPONE) as spin probe to acquire an EPR spectrum. The broadening agent ferricyanide added leads to changes in the high-field region of the EPR spectrum, which reflects differences in membrane permeability. R-value, defined as the ratio of water (W) to lipid (L) component in height in the high-field region of the EPR spectrum, symbolizes membrane permeability for a given sample. The R-values corresponding to a certain dose treatment of grains displayed a definitive distribution pattern. A unit row vector with 20 components was used to describe the R-value distribution pattern for a given treatment. The transaction angle between vectors corresponding to grains irradiated and unirradiated, θ, was used as quantitative index for membrane permeability changes following gamma-ray irradiation. gamma-Ray irradiated grains germinated at low rates, and the regression equation of germination rate as a function of the irradiation dose is: Germination Rate (%)=94.8 exp[-0.264xIrradiation Dose (kGy)] (r(2)=0.991, P<0.001). Embryos excised from grains following gamma-ray irradiation show increases in θ values with irradiation dose. The θ value is negatively linearly correlated with the germination rate. It suggests that gamma-ray irradiation leading to increases in membrane permeability is consistent with that leading to low germination rate of grains. The introduction to vector analysis method on membrane permeability changes in this study is very practical. PMID- 10725522 TI - Differences in growth and water relations among Phaseolus vulgaris cultivars in response to induced drought stress. AB - Relatively little ecophysiological research has been conducted to determine the responses to drought of Phaseolus vulgaris. Four bean cultivars (cvs.) from Brazil, A320, Carioca, Ouro Negro and Xodo were submitted to an imposed water deficit in order to evaluate the importance of some adaptive mechanisms of drought resistance through the analysis of growth parameters, water status, gas exchange and indicators of tolerance mechanisms at the cellular level. During the drought treatment, relative growth rates were more reduced for A320 and Xodo than Carioca and Ouro Negro. A320 closed its stomata very rapidly and complete stomatal closure was obtained at Psi(w)=-0.6 MPa, in contrast to the other cvs. where stomata were fully closed only at Psi(w)=-0.9 MPa. Net assimilation rates were closely related to stomatal conductances. Mechanisms at the cellular level appeared to be mostly important for higher tolerance. Carioca and Ouro Negro, when compared to A320 and Xodo, were characterized by having better drought tolerance mechanisms and higher tissue water retention capacity leading to a better growth under water deficits. The leaf dehydration rates of those cvs. were slow whereas those of the drought sensitive cvs. were rapid. The results were confirmed by the electrolyte leakage test and leaf osmotic potential measurements, which indicated higher membrane resistance and osmotic adjustment in the two tolerant cvs. Carioca and Ouro Negro. It appears from this study that despite being cultivated in the same geographical region, the four cvs. of P. vulgaris displayed somewhat different drought adaptive capacities for prolonged drought during the vegetative phase. PMID- 10725523 TI - Diversity of cell lengths in terminal portions of roots: implications to cell proliferation. AB - Terminal meristems are responsible for all primary growth of roots. It has been asserted that all cells of root meristems are actively dividing (no cells cycle slowly or arrest in the cycle) and stem cell populations expand exponentially. Because cells do not slide relative to each other in roots, relative cell lengths may be used to determine relative cell cycle durations and/or proportions of cells actively dividing in root tissues. If all cells are cycling, no interphase cells should be longer than critical length (length of longest mitotic cell in the meristem) and cells should exhibit an exponential cell-age distribution. Lengths of all cells were obtained radially across entire median longitudinal root sections at 0.5, 1.0, 1.5, 2.0, 2.5 and 3.0 mm from the founder cell/root cap boundary for five plant species to estimate percentages of cells longer than critical length. For example, up to 15 and 90% of all interphase cells were longer than critical length in 0.5 and 2.0 mm tissues, respectively, indicating that slow-cycling and/or non-proliferative cells are present in such tissues. In order to determine if the distribution of cell lengths in 0.5 mm segments approximated an exponential cell-age distribution, lengths of interphase cells less than critical length were determined. Such interphase cells were placed into ten groups according to cell length and percentages of cells in each group were compared with percentages of cells in groups calculated from an exponential cell age distribution. Percentages of cells were significantly different from predicted percentages of between 6 and 9 out of ten groups - cell lengths were not distributed exponentially. Because there are significant numbers of interphase cells longer than critical length and since lengths of interphase cells shorter than critical length do not resemble an exponential cell-age distribution, it must be concluded that not all cells in root segments from 0.5 to 3.0 mm root segments are actively dividing. Heretofore, no databases of cell lengths have been used to test these assertions. PMID- 10725524 TI - Non-structural carbohydrate status in Norway spruce buds in the context of annual bud structural development as affected by acidic pollution. AB - The present study focused on changes in the annual dynamics of the contents of non-structural saccharides (NSS) of Norway spruce vegetative buds related to their structural development under the effect of acidic pollution during the year 1995. Two types of material were analysed: (1) 4-year-old trees treated for 2 years by simulated acid rain (SAR; pH 2.9 and 3.9), and (2) 40-60-year-old trees growing in natural mountain stands exhibiting different degrees of macroscopic damage. Our study revealed that the dynamics of the NSS content reflected the major morphogenetic and developmental changes occurring during the annual bud developmental cycle. No systematic changes in the annual dynamics of NSS content were observed in buds from both mountain sites, or as a consequence of the SAR. The total sugar content of bud tissues was composed of a combination of five main sugar components: sucrose, glucose, fructose, raffinose family oligosaccharides (RFO; combination of raffinose and stachyose), and a pinitol fraction (PF) probably of cyclitols with pinitol as a main member. The dynamics of individual sugar components also reflected possible carbohydrate mediated bud frost protection. Interesting results were obtained from buds in dormant state. In dormant buds of the SAR experiment the higher value of the ratio PF:RFO of the pinitol fraction and raffinose family oligosaccharides followed the higher dose of SAR treatment. When evaluating the ratio from both types of material we assumed that changes in PF:RFO ratio corresponded to early stages of damage or acute metabolic reaction. Thus, we suggest the ratio PF:RFO as a possible non specific metabolic marker of early bud stress reaction which is, among other stress factors, sensitive to increasing load of acidic pollutants. PMID- 10725525 TI - Effects of elevated atmospheric CO(2) and temperature on leaf optical properties in Acer saccharum. AB - Elevated partial pressures of atmospheric carbon dioxide, similar to numerous causes of plant stress, may alter leaf pigmentation and structure and thus would be expected to alter leaf optical properties. Hypotheses that elevated CO(2) pressure and air temperature would alter leaf optical properties were tested for sugar maple (Acer saccharum) in the middle of its fourth growing season under treatment. The saplings had been growing since 1994 in open-top chambers and partial shade at Oak Ridge, Tennessee under the following treatments: (1) ambient CO(2) pressure and air temperature (control); (2) CO(2) pressure approximately 30 Pa above ambient; (3) air temperatures 3 degrees C above ambient; and (4) elevated CO(2) and air temperature. Under elevated CO(2) or temperature, spectral reflectance, transmittance and absorptance in the visible spectrum (400-720 nm) tended to change in patterns that generally are associated with chlorosis, with maximum differences from the control near 700 nm. However, these changes were not significant at P=0.05. Although reflectance, transmittance and absorptance at 700 nm correlated strongly with leaf chlorophyll concentration, variability in chlorophyll concentration was greater within than among treatments. The lack of treatment effects on pigmentation explained the non-significant change in optical properties in the visible spectrum. Optical properties in the near-infrared (721 850 nm) were similarly unresponsive to treatment with the exception of an increased absorptance throughout the 739-850 nm range in leaves that developed under elevated air temperature alone. This response might have resulted from effects of air temperature on leaf internal structure. PMID- 10725526 TI - Low HDL cholesterol, aggression and altered central serotonergic activity. AB - Many studies support a significant relation between low cholesterol levels and poor impulse, aggression and mood control. Evidence exists also for a causal link between low brain serotonin (5-HT) activity and these behaviors. Mechanisms linking cholesterol and hostile or self-destructive behavior are unknown, but it has been suggested that low cholesterol influences 5-HT function. This study was designed to explore the relationship between plasma cholesterol, measures of impulsivity and aggression, and indices of 5-HT function in personality disordered cocaine addicts. Thirty-eight hospitalized male patients (age 36.8+/ 7.1) were assessed with the DSM-III-R, the Buss-Durkee Hostility Inventory (BDHI), the Barratt Impulsiveness Scale (BIS) and the Brown-Goodwin Assessment for Life History of Aggression. Fasting basal cholesterol (total, LDL and HDL) was determined 2 weeks after cocaine discontinuation. On the same day 5-HT function was assessed by neuroendocrine (cortisol and prolactin) and psychological (NIMH and 'high' self-rating scales) responses following meta chlorophenylpiperazine (m-CPP) challenges. Reduced neuroendocrine responses, 'high' feelings and increased 'activation-euphoria' following m-CPP have been interpreted as indicating 5-HT alterations in a variety of psychiatric conditions. Significantly lower levels of HDL cholesterol were found in patients who had a history of aggression (P=0.005). Lower levels of HDL cholesterol were also found to be significantly associated with more intense 'high' and 'activation-euphoria' responses as well as with blunted cortisol responses to m CPP (P=0.033, P=0.025 and P=0.018, respectively). This study gives further support to existing evidence indicating that in some individuals, the probability of exhibiting impulsive and violent behaviors may be increased when cholesterol is low. It also suggests that low cholesterol and alterations in 5-HT activity may be causally related. PMID- 10725527 TI - Absence of neurodegeneration in the thalamus and caudate of elderly patients with schizophrenia. AB - The cognitive and functional deterioration observed in many 'poor-outcome' patients with schizophrenia suggests an ongoing neurodegenerative process. Diagnostic neuropathologic studies have excluded known neurodegenerative diseases as the cause of this dementia, and in a previous quantitative investigation of neurodegeneration and neural injury in this population we found no abnormalities in the cerebral cortex. However, it is possible that the deterioration observed in these patients could be due to subcortical neurodegenerative processes. Neurodegeneration and neural injury in the caudate nucleus and mediodorsal nucleus of the thalamus were investigated in a postmortem study of 11 prospectively accrued, clinically well-characterized elderly people with schizophrenia, 11 elderly control subjects with no neuropsychiatric illness, and 12 subjects with Alzheimer's disease. Traditional and immunohistochemical staining and unbiased computerized counting methods were used to quantify common markers of neurodegeneration and neural injury (neuron loss, neurofibrillary tangles, astrocytosis, microgliosis). No statistically significant differences were found between schizophrenia and control subjects for the densities of any markers. There is no evidence that abnormal neurodegeneration occurs in these two important subcortical structures. PMID- 10725528 TI - MMPI discriminators of deficit vs. non-deficit recent-onset schizophrenia patients. AB - Seventy-four patients with a recent initial onset of schizophrenia were studied during an inpatient hospitalization for a recent onset of schizophrenia as well as during a 12-month period of outpatient treatment as part of a large longitudinal study at UCLA. The Proxy for the Deficit Syndrome (PDS; Kirkpatrick, B., Buchanan, R.W., Carpenter, W.T., 1993. Case identification and stability of the deficit syndrome of schizophrenia. Psychiatry Research 47, 47-56.) was calculated based on psychiatric symptoms rated on the Brief Psychiatric Rating Scale every 2 weeks throughout the 12 months. The Minnesota Multiphasic Personality Inventory (MMPI) was administered to the schizophrenia patients at the index hospitalization. The 168-item version of the MMPI (MMPI-168) was administered at the baseline point of the 12-month period of outpatient treatment, and again 1 year later. Normal comparison subjects were tested with the MMPI or MMPI-168 at comparable time intervals. The UCLA Social Attainment Scale, a measure of the adequacy of social functioning and relatedness, was examined at the outpatient baseline and 12-month points. During the outpatient period, the Deficit Schizophrenia group (i.e. schizophrenia patients with high 12 month average PDS scores) had lower T-scores than the Non-deficit Schizophrenia group on several MMPI-168 scales, especially scales related to affective distress and anxiety. The MMPI-168 scores of normal subjects were generally the lowest of the three groups, but not always significantly lower than those of the Deficit Schizophrenia group. Social functioning at the end of the 12-month period was worst for the patient group with high deficit (PDS) scores. The findings are congruent with the concept of a Deficit Syndrome for which the PDS is the proxy. PMID- 10725529 TI - The age of onset of schizophrenia and the theory of anticipation. AB - The clinical phenomenon called anticipation is usually defined as a decrease in age at onset and/or an increase in disease severity in successive generations of afflicted families. The purpose of this study was to examine variables that might influence anticipation in schizophrenia. A total of 380 Austrian patients, born between 1935 and 1964, met criteria for schizophrenia with ICD-8 or ICD-9, SADS-L and DSM-III-R criteria. The inclusion criteria also required medical records of patients to contain information about the year of birth, season of birth, age at onset, accidents or meningoencephalitic diseases during childhood, first- and second-degree relatives afflicted with schizophrenia, sibship size, sib order, education of patient, age of parents, occupation of parents, loss of parents, and place of residence. A Cox multiple-regression analysis showed three factors as having a significant influence on the age of disease onset, including year of birth (which had the largest influence), family history (sporadic cases showed an onset 2 years later than familial cases) and residence (urban dwellers showed psychotic symptoms approximately 1 year sooner than rural ones). A Kaplan-Meier Survival Analysis showed that younger cohorts had onset approximately 10 years earlier in sporadic and familial cases. This cohort effect might be a major source of bias in studies of anticipation. PMID- 10725530 TI - Cerebral laterality in adolescent major depression. AB - This study tests the hypothesis that adolescents with major depression exhibit abnormalities in cerebral asymmetry previously found among adults. Perceptual asymmetry was assessed through tests of verbal and non-verbal dichotic listening in four groups - 48 adolescents with major depression, 22 adolescent comparisons with no history of Axis I disorders, 149 adults with major depression, and 57 comparison adults with no history of Axis I disorders. Data from adults have been previously reported. In both age groups, subjects with major depression were further divided based on the presence or absence of an anxiety disorder. Procedures used to collect perceptual asymmetry data in adolescents and adults were identical. In both age groups, depressed and healthy subjects showed perceptual asymmetry in expected directions for verbal and non-verbal dichotic tasks. Depressed and comparison subjects differed in performance on the Fused Word Test, though these differences varied as a function of anxiety and developmental level. Relative to comparisons, both adolescents and adults with major depression exhibited an increased right ear/left hemisphere advantage for fused words. Adults but not adolescents with comorbid major depressive and anxiety disorders exhibited a reduced right ear/left hemisphere advantage for fused words. These findings suggest similarities and differences across development in the relationship between cerebral laterality and psychopathology. Further studies using longitudinal and family-based designs, as well as various measures of regional brain activity, are needed to enhance understanding of associations between cerebral laterality and psychopathology across development. PMID- 10725531 TI - Diurnal variation in spontaneous eye-blink rate. AB - The daily pattern of spontaneous eye-blink rate (BR), a non-invasive peripheral measure of central dopamine activity, was investigated in 24 healthy subjects. The spontaneous eye-blink rate showed a stable pattern in morning, midday and afternoon hours. A significant increase was found at the evening time point (20.30 h). The finding is suggestive of a late evening increase of central dopamine activity. An increased level of subjective sleepiness was also found at the same evening point, at a time corresponding to the 'evening wake maintenance zone' or the 'forbidden zone for sleep'. A possible hypothesis is that the 'forbidden zone for sleep' may reflect a dopamine-mediated activation that counteracts a rising sleep drive. The role of diurnal variation of dopamine function should be considered both in the choice of the drug treatment regimen, and in the evaluation of biological and neuropsychological parameters. PMID- 10725532 TI - The Cambridge Depersonalization Scale: a new instrument for the measurement of depersonalization. AB - Existing self-rating scales to measure depersonalization either show dubious face validity or fail to address the phenomenological complexity of depersonalization. Based on a comprehensive study of the phenomenology of this condition, a new self rating depersonalization questionnaire was constructed. The Cambridge Depersonalization Scale is meant to capture the frequency and duration of depersonalization symptoms over the 'last 6 months'. It has been tested on a sample of 35 patients with DSM-IV depersonalization disorder, 22 with anxiety disorders, and 20 with temporal lobe epilepsy. Scores were compared against clinical diagnoses (gold standard) and correlated with the depersonalization subscale of the Dissociation Experiences Scale (DES). The scale was able to differentiate patients with DSM-IV depersonalization disorder from the other groups, and showed specific correlations with the depersonalization subscale of the DES (r=0.80; P=0.0007). The scale also showed high internal consistency and good reliability (Cronbach alpha and split-half reliability were 0.89 and 0.92, respectively). The instrument can, therefore, be considered as valid and reliable, and can be profitably used in both clinical and neurobiological research. PMID- 10725533 TI - Differential measures of 'sustained attention' in children with attention deficit/hyperactivity or tic disorders: relations to monoamine metabolism. AB - Controversy exists on whether the constructs tested by paper/pencil and computerized continuous-performance tests (CPT) are similar, and the deficits recorded in children with attention-deficit/hyperactivity symptoms (ADHD) are comparable. Signal-detection measures were recorded on four such tests of 'sustained attention', with increasing working-memory requirements in healthy children (14; mean 10 years), and those with ADHD (14; mean 10 years) or a tic syndrome (TS, 11; mean 11 years). Clinical associations were sought from 24-h urinary measures of monoamine activity. The cancellation paper/pencil test revealed no group differences for errors or signal detection measures. On the CPT, ADHD children made more omission and commission errors than control subjects, but TS children made mostly omissions. This reflected the poor perceptual sensitivity (d-prime, d') for ADHD and conservative response criteria (beta) for TS children. This group difference extended to the CPTax, which was shown on a regression analysis to test for putative working-memory-related abilities as well as concentration. In all children, immediate response-feedback reduced omissions, and modestly improved d'. CPTax performance related negatively to dopamine metabolism in control subjects and to serotonin metabolism in the ADHD group. But comparisons between the metabolites in the ADHD group suggest that increased serotonin and decreased noradrenaline, with respect to dopamine metabolism, may detract from CPT performance in terms of d'. CPT tasks demonstrated a perceptual-based impairment in ADHD and response conservatism in TS patients independent of difficulty. Catecholamine activity was implicated in the promotion of perceptual processing in normal and ADHD children, but serotonin activity may contribute to poor CPTax (working-memory) performance in ADHD patients. PMID- 10725534 TI - A critical review of cellobiose dehydrogenases. AB - Cellobiose dehydrogenase (CDH) is an extracellular enzyme produced by various wood-degrading fungi. It oxidizes soluble cellodextrins, mannodextrins and lactose efficiently to their corresponding lactones by a ping-pong mechanism using a wide spectrum of electron acceptors including quinones, phenoxyradicals, Fe(3+), Cu(2+) and triiodide ion. Monosaccharides, maltose and molecular oxygen are poor substrates. CDH that adsorbs strongly and specifically to cellulose carries two prosthetic groups; namely, an FAD and a heme in two different domains that can be separated after limited proteolysis. The FAD-containing fragment carries all known catalytic and cellulose binding properties. One-electron acceptors, like ferricyanide, cytochrome c and phenoxy radicals, are, however, reduced more slowly by the FAD-fragment than by the intact enzyme, suggesting that the function of the heme group is to facilitate one-electron transfer. Non heme forms of CDH have been found in the culture filtrate of some fungi (probably due to the action of fungal proteases) and were for a long time believed to represent a separate enzyme (cellobiose:quinone oxidoreductase, CBQ). The amino acid sequence of CDH has been determined and no significant homology with other proteins was detected for the heme domain. The FAD-domain sequence belongs to the GMC oxidoreductase family that includes, among others, Aspergillus niger glucose oxidase. The homology is most distinct in regions that correspond to the FAD binding domain in glucose oxidase. A cellulose-binding domain of the fungal type is present in CDH from Myceliophtore thermophila (Sporotrichum thermophile), but in others an internal sequence rich in aromatic amino acid residues has been suggested to be responsible for the cellulose binding. The biological function of CDH is not fully understood, but recent results support a hydroxyl radical generating mechanism whereby the radical can degrade and modify cellulose, hemicellulose and lignin. CDH has found technical use in highly selective amperometric biosensors and several other applications have been suggested. PMID- 10725535 TI - Modification with a phosphorylation tag of PKA in the TraT-based display vector of Escherichia coli. AB - We have previously developed the TraT display system to express the preS1 peptide of human hepatitis B virus (HBV) and the snake venom rhodostomin (RHO) on the surface of Escherichia coli. In this study, we modified the pT2 vector by adding a thrombin cutting site and a phosphorylation tag of protein kinase A before the multiple restriction enzyme sites. The modified vector allowed us to label the TraT fusion protein (TraT-RHO) with [32P] and to increase the detection sensitivity of TraT-RHO expression bacteria binding to and being internalized into BHK-21 cells. After the thrombin cleavage, the isotope labeled RHO could be detected in a free form. We therefore suggest that the new version of pT2 vector, pT2-KL, will facilitate to identify the counterpart of displayed peptide. PMID- 10725536 TI - Efficiency of light utilization of Chlamydomonas reinhardtii under medium duration light/dark cycles. AB - The light regime inside a photobioreactor is characterized by a light gradient with full (sun)light at the light-exposed surface and darkness in the interior of the bioreactor. Consequently, depending on the mixing characteristics, algae will be exposed to certain light/dark cycles. In this study the green alga Chlamydomonas reinhardtii was cultivated under five different light regimes: (1) continuous illumination; (2) a square-wave light/dark cycle with a light fraction (epsilon) of 0.5 and a duration (t(c)) of 6.1 s; (3) epsilon=0.5, t(c)=14.5 s; (4) epsilon=0.5, t(c)=24.3 s and (5) epsilon=0.8, t(c)=15.2 s. The biomass yield on light energy, protein per photons, decreased under light/dark cycles (epsilon=0. 5) in comparison to continuous light (CL), from 0.207 (CL) to 0.117 0.153 g mol(-1) (epsilon=0.5). Concomitantly, the maximal specific photosynthetic activity, oxygen production per protein, decreased from 0.94 (CL) to 0.64-0.66 g g(-1) h(-1) (epsilon=0.5). Also the quantum yield of photochemistry, yield of the conversion of light energy into chemical energy, decreased from 0.47 (CL) to 0. 23 (epsilon=0.5, t(c)=24.3 s). Apparently, C. reinhardtii is not able to maintain a high photosynthetic capacity under medium-duration light/dark cycles and since specific light absorption did not change, light utilization efficiency decreased in comparison to continuous illumination. PMID- 10725537 TI - A new method for on-line measurement of the volumetric oxygen uptake rate in membrane aerated animal cell cultures. AB - Oxygen is a key substrate in animal cell metabolism and its consumption is thus a parameter of great interest for bioprocess monitoring and control. A system for measuring it based on an oxygen balance on the liquid phase was developed. The use of a gas-permeable membrane offered the possibility to provide the required quantity of oxygen into the culture, while avoiding problems of foaming or shear stress generally linked to sparging. This aeration system allowed moreover to keep a known and constant k(L)a value through cultures up to 400 h. Oxygen uptake rate (OUR) was measured on-line with a very good accuracy of +/-5%, and the specific OUR for a CHO cell line was determined during batch (growth phase) and continuous culture as, respectively, equal to 2. 85x10(-13) and 2.54x10(-13) mol O(2) cell(-1) h(-1). It was also shown that OUR continuous monitoring gives actually more information about the metabolic state of the culture than the cell concentration itself, especially during transition phases like the end of the growth phase in a batch culture. PMID- 10725538 TI - An alpha-glucuronidase of Schizophyllum commune acting on polymeric xylan. AB - The main alpha-glucuronidase (EC 3.2.1.131) of the fungus Schizophyllum commune was purified to homogeneity using standard chromatographic methods; anion exchange, hydrophobic interaction chromatography and gel filtration. The enzyme had a molecular mass of 125 kDa as determined by SDS-polyacrylamide gel electrophoresis and a pI value of 3.6 according to isoelectric focusing. The N terminal amino acid sequence of the S. commune alpha-glucuronidase did not show any homology with other alpha-glucuronidases. It exhibited maximal activity at pH values from 4.5 to 5.5 and was stable for 24 h between pH 6 and 8 at 40 degrees C. The highest temperature at which the enzyme retained its full activity for 24 h at pH 5.8 was 40 degrees C. The alpha-glucuronidase of S. commune was able to remove almost all 4-O-methylglucuronic acid groups from water-soluble polymeric softwood arabinoglucuronoxylans. The action of the enzyme on birchwood acetyl glucuronoxylan was limited due to the high amount of acetyl substituents. The degree of hydrolysis of partially soluble deacetylated glucuronoxylan did not exceed 50% of the theoretical maximum. However, together with a xylanase hydrolysing the xylan backbone the action of the alpha-glucuronidase of S. commune on glucuronoxylan was clearly enhanced. It was apparent that the enzyme was able to remove the 4-O-methylglucuronic groups mainly from soluble substrates. PMID- 10725539 TI - A fluorescence based non-radioactive electrophoretic mobility shift assay. AB - Electrophoretic mobility shift assay (EMSA) or gel shift assay is one of the most powerful methods for studying protein-DNA interactions. Typically, 32P-labeled DNA probes containing the sequence bound by the protein of interest are used in EMSA (rEMSA). Although rEMSA is sensitive and practicable, it relies on the handling of hazardous radioisotopes, and does not easily allow quantification. We developed a non-radioactive procedure using fluorescence (Cyano dye Cy5) labeled oligodeoxynucleotide duplexes as specific probes (fEMSA) and an automatic DNA sequencer for analysis. Testing different DNA-binding proteins (restriction endonuclease EcoRII, transcription factor NFkappaB and it's subunit p50) the results in fEMSA and rEMSA are similar in regard to quality, reproducibility, and sensitivity. fEMSA allows a semiquantitative screening of large amounts of samples for specific DNA binding activities and is, therefore, a high throughput technology for semiquantitative analysis of DNA-protein interaction. PMID- 10725540 TI - Bovine beta-lactoglobulin receptors on transformed mammalian cells (hybridomas MARK-3): characterization by flow cytometry. AB - Flow cytometry was used to demonstrate the presence of beta-lactoglobulin (betaLG) receptors on living murine hybridoma MARK-3 cells using a fluorescein isothiocyanate-betaLG conjugate (FITC-betaLG: molar ratio of 5:1). A site occupation curve was produced using a shift in the mean channel fluorescence at various concentrations of FITC-betaLG. The binding of labelled ligand was concentration dependent and was inhibited by unlabelled betaLG. The on-rate constant was 3.2x10(2) M(-1) min(-1) and the off-rate constant was 0.002 min(-1). Scatchard plot analysis gave a dissociation constant (K(d)) of 44+/-21x10(-7) and 39+/-24x10(-5) M (n=3). Flow cytometry indicated that at least 15% of the FITC betaLG were internalized for 5 min and that internalization was temperature- and time-dependent. The internalization was confirmed by 3-D fluorescence microscopy (CELLScan system). PMID- 10725541 TI - Metabolism of cellulose by Phanerochaete chrysosporium in continuously agitated culture is associated with enhanced production of lignin peroxidase. AB - Production of the extracellular heme protein lignin peroxidase (LiP) by Phanerochaete chrysosporium is currently associated with a number of requirements, namely exposure of the cultures to oxygen; limiting nutrient nitrogen or carbon and static or semi-static culture conditions. To obtain LiP activity in continuously agitated liquid culture requires the inclusion of a surfactant. However, using cellulose as the carbon source, we obtained high titres (0.2-0.4 U ml(-1)) of LiP in submerged liquid cultures under conditions of continuous agitation, without substrate limitation or the need to add oxygen or surfactant. Comparison of the morphological and physiological traits of hyphae maintained on either cellulose or free glucose supports observations that the synthesis of extracellular polysaccharide in the cultures grown on glucose, restricts oxygen diffusion into the hyphae, which is necessary for LiP induction. They also suggest that isozymes of LiP synthesised under these conditions may be triggered in response to oxidant stress. PMID- 10725542 TI - Laccase activity tests and laccase inhibitors. AB - Sulfhydryl organic compounds described as laccase inhibitors: dithiothreitol, thioglycolic acid, cysteine, diethyldithiocarbamic acid, and sodium azide were tested for their activity toward laccase of Trametes versicolor in different test systems utilising 2, 2'-azino-bis-(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) and 2, 6-dimethoxyphenol as enzyme substrates. Only sodium azide acted as a true laccase inhibitor and showed no significant interference with the enzyme tests. All other substances did not significantly inhibit the laccase activity and the previously reported inhibitory effects result from the reductions of the reaction products such as ABTS radical cation and diquinone or subsequent non enzymatic interactions during substrate oxidation. The latter apparently forms a complex with unreacted ABTS displaying varied spectral characteristics and resulting in an underestimation of enzyme activity. PMID- 10725543 TI - A simple method for non-radioactive PCR-SSCP using MDE gel solution and a midi gel format: application for the detection of variants in the GLUT1 and CTLA-4 genes. AB - The need to identify disease-causing mutations and DNA polymorphisms has increased with the continuing identification of new candidate genes. PCR single strand conformation polymorphism (PCR-SSCP) is one of the techniques most widely used to identify a mutant sequence or a polymorphism in a known gene. However, the original SSCP protocols using the incorporation of radioactive label and polyacrylamide gel electrophoresis on sequencing gels for detection were labour intensive and time-consuming. Here we describe a simple SSCP protocol using MDE gel solution and a midi gel format to detect SSCP variations in the glucose transporter gene GLUT1, that we have previously analysed with the standard radioactive SSCP protocol, and we have also tested this method on the previously described point mutation (A/G transition in exon 1) of the CTLA-4 (cytotoxic T lymphocyte associated-4) gene. All known variants were detected. Based on the results, this technique appears to be simple, with no use of radioactive labels and with easy handling of the gel. Furthermore, it needs little optimisation, is relatively rapid and highly sensitive. We propose this method for the first screening for candidate gene variants. PMID- 10725545 TI - Intergenic HA-NA interactions in influenza A virus: postreassortment substitutions of charged amino acid in the hemagglutinin of different subtypes. AB - In our previous studies influenza A virus reassortants having neuraminidase (NA) gene of A/USSR/90/77 (H1N1) strain and hemagglutinin (HA) genes of H3, H4 and H13 subtypes were shown to produce a low virus yield and to exhibit a strong tendency to virion aggregation. More detailed studies with the use of a H3N1 reassortant and its high-yield non-aggregating variants revealed that NA of A/USSR/90/77 strain is inefficient in the removal of the terminal sialic acid residues from the virion components, and that the inefficiency of NA may be compensated by mutations in HA gene leading to a decrease of the receptor-binding affinity (Kaverin, N.V. , Gambaryan, A.S., Bovin, N.V., Rudneva, I.A., Shilov, A.A., Khodova, O.M., Varich, N.L., Sinitsin, B.V., Makarova, N.L., Kaverin, N.V., 1998. Postreassortment changes in influenza virus hemagglutinin restoring HA-NA functional match, Virology 244, 315-321). The present report describes studies performed with the use of H2N1 and H4N1 reassortants having HA genes of A/Pintail/Primorie/695/76 (H2N3) and A/Duck/Czechoslovakia/56 (H4N6) strains respectively and NA gene of A/USSR/90/77 strain. The low-yield reassortants and their high-yield non-aggregating variants were studied in both direct and competitive binding assays with sialic acid-containing substrates. The non aggregating variants were shown to have a decreased affinity as compared to the initial reassortants toward high-molecular-weight sialic acid-containing substrates. The sequencing of HA genes revealed that all non-aggregating variants of H2N1 and H4N1 reassortants had amino acid substitutions increasing the negative charge of the HA molecule in the vicinity of the receptor-binding pocket. The results suggest that the influenza virus reassortants containing low functional NA undergo similar postreassortment changes irrespective of the HA subtype: their receptor-binding activity decreased due to negatively charged amino acid substitutions in the vicinity of the receptor-binding pocket. PMID- 10725544 TI - The authentic sequence of rotavirus SA11 nonstructural protein NSP4. AB - Recent studies demonstrate that the rotavirus nonstructural protein NSP4 functions as an enterotoxin and plays an important role in viral pathogenesis. Previous in vitro studies of NSP4 have used a cDNA clone of gene 10 derived from the prototypic rotavirus strain, SA11. We recently compared the sequence of the commonly used NSP4 cDNA with the sequence obtained from several SA11 isolates by direct sequencing of reverse transcription polymerase chain reaction products. One codon difference was identified between the cDNA clone and the SA11 virus isolates, and this resulted in a predicted amino acid substitution at position 47. The cDNA sequence specifies an asparagine at position 47, and the SA11 virus gene 10 encodes a hisitidine. To determine if this amino acid substitution altered the function of NSP4, we analyzed the ability of both NSP4-Asn47 and NSP4 His47 to regulate intracellular calcium levels and exhibit cell cytotoxicity. Our results indicate that the expression of NSP4-His47 from a recombinant baculovirus displays enhanced cytotoxicity and calcium flux. PMID- 10725546 TI - Fusion of uninfected T-cells occurs with immature HIV-1 protease-mutant, but not morphologically similar protease inhibitor derived particles. AB - Protease inhibitors are widely used in the treatment of human immunodeficiency virus type 1 (HIV-1)-infected individuals and show a drastic effect on the reduction of virus load. We previously reported that doughnut-shaped, protease defective gp120-containing HIV-1 particles from an L-2 cell clone, carrying a provirus with mutations at the pol (protease), env (gp41) and nef genes, rapidly and more effectively induces virus particle-mediated syncytia formation of uninfected T-cells, than a parental wild-type laboratory strain of HIV-1 (LAI). In this study, we examined the possibility of whether enhanced syncytia formation is mediated by morphologically similar doughnut-shaped particles obtained after treatment of LAI-infected cells with the protease inhibitors L-689, 502, DMP-323, RO-31-8959, and KNI-272. Utilizing such protease inhibitor-induced particles and a clone of MOLT-4 cells, we could not detect any enhancement of syncytia formation, over that seen with wild-type LAI particles. This result should alleviate concerns of patients on highly active antiretroviral therapy (HAART), that protease inhibitors might accelerate progression of the disease through enhanced production of defective, 'immature'-appearing particles. PMID- 10725547 TI - Fusion protein predicted amino acid sequence of the first US avian pneumovirus isolate and lack of heterogeneity among other US isolates. AB - Avian pneumovirus (APV) was first isolated from turkeys in the west-central US following emergence of turkey rhinotracheitis (TRT) during 1996. Subsequently, several APV isolates were obtained from the north-central US. Matrix (M) and fusion (F) protein genes of these isolates were examined for sequence heterogeneity and compared with European APV subtypes A and B. Among US isolates the M gene shared greater than 98% nucleotide sequence identity with only one nonsynonymous change occurring in a single US isolate. Although the F gene among US APV isolates shared 98% nucleotide sequence identity, nine conserved substitutions were detected in the predicted amino acid sequence. The predicted amino acid sequence of the US APV isolate's F protein had 72% sequence identity to the F protein of APV subtype A and 71% sequence identity to the F protein of APV subtype B. This compares with 83% sequence identity between the APV subtype A and B predicted amino acid sequences of the F protein. The US isolates were phylogenetically distinguishable from their European counterparts based on F gene nucleotide or predicted amino acid sequences. Lack of sequence heterogeneity among US APV subtypes indicates these viruses have maintained a relatively stable population since the first outbreak of TRT. Phylogenetic analysis of the F protein among APV isolates supports classification of US isolates as a new APV subtype C. PMID- 10725548 TI - Unique N-linked glycosylation of murine coronavirus MHV-2 membrane protein at the conserved O-linked glycosylation site. AB - The membrane (M) proteins of murine coronavirus (MHV) strains have been reported to contain only O-linked oligosaccharides. The predicted O-glycosylation site consisting of four amino acid residues of Ser-Ser-Thr-Thr is located immediately adjacent to the initiator Met and is well conserved among MHV strains investigated so far. We analyzed the nucleotide sequence of a highly virulent strain MHV-2 M-coding region and demonstrated that MHV-2 had a unique amino acid, Asn, at position 2 at the conserved O-glycosylation site. We also demonstrated that this substitution added N-linked glycans to MHV-2 M protein resulting in increment of molecular mass of MHV-2 M protein compared with JHM strain having only O-linked glycans. PMID- 10725549 TI - Analysis of host response modifier ORFs of ectromelia virus, the causative agent of mousepox. AB - From the right-hand end of the ectromelia virus (strain Moscow) genome, 32318 bps have been sequenced, and characterized to include a total of 18 open reading frames (ORFs) and six regions which apparently no longer code for functional proteins. At least six of the ORFs appear to be involved in blocking the inflammatory/immune host response to infection, and therefore probably contribute significantly to the virulence of this virus in its natural host, the mouse. One of these genes encoded an isolog of the poxvirus chemokine binding protein, and was shown to be the most abundant protein secreted from ectromelia virus infected cells. Two regions were found to have significant similarity to poxvirus genes encoding tumor necrosis factor (TNF) binding proteins. Both are distinct from cytokine response modifier (crm)B and crmC but only one is predicted to encode a functional TNF binding protein. A novel similarity between the C-terminal domain of poxvirus TNF binding proteins and several other poxvirus proteins is also presented. The results are discussed in the context of ectromelia virus pathogenesis of mice. PMID- 10725550 TI - Differential roles of the 5' untranslated regions of cucumber mosaic virus RNAs 1, 2, 3 and 4 in translational competition. AB - RNA species of plant tripartite RNA viruses show distinct translational activities in vitro when the viral RNA concentration is high. However, it is not known what causes the differential translation of virion RNAs. Using an in vitro wheat germ translation system, we investigated the translation efficiencies and competitive activities of chimeric cucumber mosaic virus (CMV) RNAs that contained viral untranslated regions (UTRs) and a luciferase-coding sequence. The chimeric RNAs exhibited distinct translation efficiencies and competitive activities. For example, the translation of chimeric CMV RNA 4 was about 40-fold higher than that of chimeric CMV RNA 3 in a competitive environment. The distinct translation resulted mainly from differences in competitive activities rather than translation efficiencies of the chimeric RNAs. The differential competitive activities were specified by viral 5 UTRs, but not by 3 UTRs or viral proteins. The competitive translational activities of the 5 UTRs were as follows: RNA 4 (coat protein)>RNAs 2 and 1 (2a and 1a protein, or replicase )> RNA 3 (3a protein). PMID- 10725551 TI - Cell surface expression of immature H glycoprotein in measles virus-infected cells. AB - Two forms of hemagglutinin (H) protein, one with an apparent molecular mass of 78 kDa (78K H protein) and the other with that of 74 kDa (74K H protein), are present in cells infected with measles virus (MV). We previously observed that only the mature 78K H protein, a completely glycosylated form of the 74K H protein, was expressed on the cell surface of the infected cells. In the present study, we detected transient expression of the 74K H protein on the cell surface of infected cells by pulse-chase studies, although the level of this expression was much lower than that of the 78K H protein. On the cell surface the 74K H protein was present as dimers and sensitive to endo-beta-N-acetylglucosaminidase H digestion. Treatment with brefeldin A, which blocks the transport of membrane and secretory proteins from the endoplasmic reticulum to the Golgi apparatus, inhibited the cell surface expression of the 78K H protein, but not that of the 74K H protein. These data suggest that a part of the MV 74K H proteins could be transported directly to the cell surface - probably via an alternative pathway - without processing to the complex form in the Golgi apparatus. PMID- 10725552 TI - RNA-binding activities of cocksfoot mottle sobemovirus proteins. AB - Cocksfoot mottle virus (CfMV) has a positive-sense ssRNA genome containing four open reading frames (ORFs). ORF1 encoded protein (P1) is the putative movement protein; the product of ORF2a (P2a) contains VPg and the motifs characteristic of serine proteases. P2b, encoded by ORF2b, is the putative RNA-dependent RNA polymerase. P3, the coat protein, is encoded by ORF3. CfMV P1, P2a, P2b, and P3, containing a six histidine tag at the amino terminus, were expressed in Escherichia coli, purified and their RNA-binding activities were analysed. The northwestern blot assay showed that His-tagged P1, P2a, P2b, and P3 were able to interact with ssRNA transcripts in a sequence-nonspecific manner. The filter binding assay confirmed the ssRNA-binding capacity of recombinant P1, P2a, and P3. The RNA-binding activities of His-tagged P3 and native coat protein were similar. P1 and P2a binding to ssRNA decreased markedly by increasing NaCl concentrations. In contrast, P3 had the RNA-binding optimum at 100-200 mM NaCl. We discuss the possible amino acid motifs involved in the RNA-binding of CfMV proteins. PMID- 10725553 TI - Sucrose and the integration of metabolism in vascular plants. AB - We consider the hypothesis that sucrose is a signal as well as a substrate. We suggest that the significance of sugar sensing in plants is the integration of whole-plant carbon flux so that the capacity of sources to produce sucrose matches the capacity of sinks to consume it. We pay particular attention to difficulties with this hypothesis and the areas where further or better evidence is needed. We conclude that there is strong correlative evidence for a link between sucrose metabolism and the level of expression of key genes, but that a number of different mechanisms may be involved. PMID- 10725554 TI - Purification and properties of growth stage-dependent antiviral proteins from the leaves of Celosia cristata. AB - Two antiviral glycoproteins, active against mechanical transmission of two tobamoviruses, tobacco mosaic virus and sunnhemp rosette virus, and citrus ring spot virus (ungrouped), were purified from the dried leaves of Celosia cristata. These proteins, called CCP-25 and CCP-27, have M(r) 25 and 27 kDa, respectively. Their concentration was found to vary between the pre-flowering and post flowering stages of C. cristata90% lesion formation at a concentration of 20-30 ug ml(-1). They were resistant to proteases in the native state, but were readily digested when denatured. Both of them imparted actinomycin D sensitive resistance by inhibiting local lesions on Nicotiana tabacum cv. Samsun NN by tobacco mosaic virus. Their application, individually, also resulted in high resistance in systemic hosts to sunnhemp rosette virus, and citrus ring spot virus, respectively. PMID- 10725555 TI - Gabaculine does not inhibit cytokinin-stimulated biosynthesis of chlorophyll in Pinus nigra seedlings in the dark. AB - Chlorophyll (Chl) accumulation was monitored during black pine (Pinus nigra L.) seed germination for 14 days in the light and in the dark in the presence of gabaculine (GAB) and cytokinin in order to elucidate the regulation of gymnosperm seedling greening in the dark, primarily at the level of aminolevulinic acid formation. In the light, GAB inhibited chlorophyll accumulation in a manner dependent on concentration and developmental stage, and in the dark it showed no effect. Cytokinin, 10(-5) M benzyl adenine (BA) partly overcame GAB-induced inhibition in the light, mainly during earlier developmental stages. In the seedlings grown in the dark, an equal quantity of Chl accumulated in the presence of cytokinin with and without GAB and it was approximately 20-40% higher than in the control seedlings or in the seedlings grown only in the presence of GAB. 5 Amino-levulinic acid (ALA) synthesis was equal in the light and in the dark in seedlings of the same age and seedlings treated with GAB grown in the dark. In the light, GAB inhibited ALA synthetic activity. The results indicate that ALA synthesis is not a rate-limiting step within Chl biosynthesis in pine seedlings grown in the dark. PMID- 10725556 TI - Comparison of drought tolerance in nitrogen-fixing and inorganic nitrogen-grown common beans. AB - In this work, we evaluated how the use of alternative N sources affects drought stress tolerance in common beans. To this end, plants were cultivated employing either N(2) fixation or two levels of inorganic nitrogen: 1 mM NH(4)NO(3) (limiting) or 10 mM NH(4)NO(3) (sufficient). Drought was imposed by withholding watering at 30 days after planting (DAP) - coinciding with flowering. At 20 DAP, growth and N content were significantly higher in NH(4)NO(3)-sufficient plants than in N(2)-fixing and NH(4)NO(3)-limited beans. At later times, only N(2) fixing and NH(4)NO(3)-sufficient plants continued assimilating N and growing, with the NH(4)NO(3)-sufficient plants being consistently bigger. After 10 days of stress (40 DAP), desiccation was evident, but only NH(4)NO(3)-sufficient plants suffered drought-induced senescence. After 20 days of stress (50 DAP), N content increased in NH(4)NO(3)-sufficient but not in N(2)-fixing beans, despite the latter's lesser state of wilt. Pod dry weight dropped 43% in NH(4)NO(3) sufficient beans with respect to well-watered plants, while remaining constant in N(2)-fixing beans. Under drought conditions, the number of pods limited pod yield regardless of the nitrogen source used; nevertheless, the translocation of soluble matter to pods continued in both NH(4)NO(3)-sufficient and N(2)-fixing beans. We conclude that common beans grown under conditions of N(2) fixation were more drought tolerant than those provided with sufficient levels of NH(4)NO(3). The most stress-sensitive traits in these plants were the incorporation of N into their shoots and the number of pods remaining on them. PMID- 10725557 TI - Molecular cloning of a novel water channel from rice: its products expression in Xenopus oocytes and involvement in chilling tolerance. AB - Water channel proteins, aquaporins, play a fundamental role in transmembrane water movements in plants. We isolated rice cDNA, rwc1, by screening a rice (Oryza sativa cv. Josaeng Tongil) cDNA library using a conserved motif of aquaporins. Like other aquaporin genes, rwc1 encodes a 290-residue protein with six putative transmembrane domains. The derived amino acid sequence of RWC1 shows high homology with PIP1 (plasma membrane intrinsic protein 1) subfamily members, which suggest it is localized in the plasma membrane. Injection of its cRNA into Xenopus oocytes increased the osmotic water permeability of the oocytes 2-3 times. Northern analysis showed that rice aquaporin genes are expressed in rice seedling leaves and roots, but that it disappeared from the root 6 h after osmotic stress began and that the transcript level remained low for about 24 h, then recovered. The time course of rice aquaporin gene-expression under osmotic stress was correlated with time course of turgor transition in plant. On the other hand, the levels of rice aquaporin gene-transcripts in leaves under chilling and recovery temperature depend on the pretreatment of mannitol for short time. This variation of the transcripts shown that rice aquaporin genes may play an important role in response to water stress-induced chilling tolerance. PMID- 10725558 TI - Characterisation of acyl-ACP desaturases from Macadamia integrifolia Maiden & Betche and Nerium oleander L. AB - The seed oil in Macadamia integrifolia contains about 30% palmitoleic acid (16:1(Delta9)) and Nerium oleander about 12% isoricinoleic acid (Delta9-hydroxy 18:1(Delta12)). It has been shown that palmitoleic acid can be produced by acyl acyl carrier protein (ACP) desaturases and it has also been shown that fatty acid hydroxylation can occur via direct substitution of a hydrogen atom. Therefore it seemed possible that the enzymes responsible for the making of these unusual fatty acids in M. integrifolia and N. oleander were of acyl-ACP desaturase type. Extracts from developing M. integrifolia developing seeds showed a relative ratio of 16:0-ACP to 18:0-ACP desaturation that was about 13 times higher than in sunflower seeds. N. oleander seed extracts catalysed conversion of 18:0-ACP to 18:1(Delta9) but only trace amounts of Delta9-hydroxy fatty acids were formed. A total of four cDNAs were isolated from developing seeds, of both species, using a fragment isolated with PCR amplification. The M. integrifolia acyl-ACP desaturase cDNA was expressed in Escherichia coli. A partly purified fraction of the enzyme showed a 16:0-ACP to 18:0-ACP desaturation ratio about 90-fold less than that in the Macadamia extracts. Expressed N. oleander acyl-ACP desaturase cDNAs showed predominantly 18:0-ACP desaturase activity and no hydroxylase activity. Thus it is not likely that any of the four acyl-ACP desaturases cloned from M. integrifolia or N. oleander is involved in the production of unusual fatty acids. PMID- 10725559 TI - Differential expression of plastidic aldolase genes in Nicotiana plants under salt stress. AB - Two homologous genes of plastidic fructose-1,6-bisphosphate aldolase (AldP) isozymes were isolated from green leaves of a salt stress-tolerant Nicotiana species, Nicotiana paniculata, by differential screening. The products of the corresponding genes, NpAldP1 and NpAldP2, were 91% identical to each other and 70 85% identical to the other known plant plastidic aldolases. Although these two genes showed similar organ-specific expression and daily cycles, their responses to salt stress differed: mRNA accumulation of NpAldP2 increased, but that of NpAldP1 slightly decreased. The mRNA accumulations of their counterparts of two other Nicotiana species, NeAldP1 and NeAldP2 (Nicotiana excelsior), and NaAldP1 and NaAldP2 (Nicotiana arentsii) were studied under the same stress condition. N. arentsii conserved accumulation profiles similar to N. paniculata, but N. excelsior did not. In N. excelsior, accumulation of NeAldP1 decreased to 50% of the control after stress and gradually recovered thereafter, whereas accumulation of NeAldP2 temporarily decreased and reached 250% of the control by the third day of stress. Southern blot analysis indicated that NpAldP1, NpAldP2, NaAldP1, and NaAldP2 include one or two closely related genes and NeAldP1 and NeAldP2 several. PMID- 10725560 TI - Molecular cloning of a soybean class III beta-1,3-glucanase gene that is regulated both developmentally and in response to pathogen infection. AB - We isolated and characterized a soybean gene (SGN1) encoding a basic beta-1,3 glucanase that is a plant class III isoform of beta-1,3-glucanase. The deduced amino acid sequence of the SGN1 gene is similar to that of the PR-Q'b gene, the basic class III beta-1,3-glucanase of tomato. Based on RNA blot hybridization, SGN1 gene expression was detected in all tissues of 4-day old seedlings, but it was present only in root tissue of 30-day old plants. GUS expression analysis carried out in transgenic tobacco plants harboring a SGN1::GUS reporter gene revealed the same expression pattern. Furthermore, the expression of SGN1 was strongly induced by a variety of defense-related signals, such as treatment with H(2)O(2), wounding, or treatment with fungal elicitor prepared from Phytophthora spp as well as inoculation with Pseudomonas syringae. However, the expression level of SGN1 was hardly induced with jasmonate, ethephon and salicylate. Overall the results suggest that the SGN1 may play a role in both plant development and plant defense against pathogen attack. PMID- 10725561 TI - Characterization of cDNAs encoding two glycine-rich proteins in chickpea (Cicer arietinum L.): accumulation in response to fungal infection and other stress factors. AB - In chickpea plants infected with the pathogenic fungus Ascochyta rabiei [Pass.] Labr. several mRNAs for two glycine-rich proteins (GRPs) were identified by differential cDNA screening. The main part of the deduced amino acid sequences of the 14.6 kD GRP1 and the larger GRP2 consists of glycine-rich repetitive elements essentially as found for GRPs in other plants. Tyrosine residues in conserved positions inside these repetitive motifs suggest an involvement of the GRPs in a polymerization process by oxidative cross-linking, i.e. cell wall fortification. Both GRP transcripts are induced by infection with A. rabiei, showing a maximum of expression 5 days post infection. Wounding of leaves and the stress of water treatment (performed as a control) also seem to induce the accumulation of GRP transcripts. PMID- 10725562 TI - In vivo evaluation of the context sequence of the translation initiation codon in plants. AB - Statistical analysis of the AUG initiation codon context in several plant organisms identified a nucleotide preference in some positions around the AUG. Sixteen AUG contexts were studied using transient expression in tobacco, maize and Norway spruce. Besides the importance of A or G at position -3, we revealed the role of positions -2, -1 for which AA or CC were found to be the best for tobacco and maize, respectively. GC (positions +4, +5) were also found to be important in both tobacco and maize. Finally, we identified a variation in context efficiency according to cell type, since A was better than G at position 3 in tobacco leaf protoplasts, while both nucleotides were equally efficient in tobacco suspension cells. PMID- 10725563 TI - Emotional imagery, the visual startle, and covariation bias: an affective matching account. AB - This study assessed the effects of imagery valence and arousal on the visually prompted startle reflex, heart rate, and estimates of probe occurrence in 24 males and 22 females. Valence and arousal independently augmented startle magnitudes, similar to prior research with acoustic probes (Witvliet, C.V.O., Vrana, S.R., 1995. Psychophysiological responses as indices of affective dimensions, Psychophysiology 32, 436-443). In both of these studies, arousal exerted stronger effects than valence on the startle reflex. Arousal also facilitated heart rate acceleration. Participants' estimates of startle flash occurrence reflected a covariation bias. Estimates were higher and more accurate for the high-arousal conditions and for the negative conditions, paralleling startle magnitude findings. Results suggest that affective response matching processes (rather than affective stimulus matching) influenced both startle reflex magnitudes and probe frequency estimates. Comparisons with the covariation bias literature are drawn, differences are addressed, and directions for future research are suggested. PMID- 10725564 TI - An ERP study of sustained spatial attention to stimulus eccentricity. AB - Effects of attention on event-related brain potentials (ERPs) were measured when subjects kept sustained attention focused on ring-shaped regions of visual space to detect infrequently presented targets at a given eccentricity. In line with a previous study that employed a trial-by-trial cueing paradigm, no modulations of sensory-evoked P1 and N1 components were found. This suggests that attentional selectivity in complex spatial selection tasks is primarily located at post perceptual processing levels. Enhanced negativities for attended as compared to unattended stimuli were present between 220 and 380 ms post-stimulus and were followed by an enlarged positivity for attended stimuli in the P3 time range. These effects reflected the distribution of attention in visual space, in part consistent with 'attentional gradient' and 'zoom-lens' models. However, ERPs also suggested the presence of selective mechanisms that exclude irrelevant stimuli located between two simultaneously attended areas. PMID- 10725565 TI - Combined eye activity measures accurately estimate changes in sustained visual task performance. AB - Five concurrent eye activity measures were used to model fatigue-related changes in performance during a visual compensatory tracking task. Nine participants demonstrated considerable variations in performance level during two 53-min testing sessions in which continuous video-based eye activity measures were obtained. Using a trackball, participants were required to maneuver a target disk (destabilized by pseudorandom wind forces) within the center of an annulus on a CRT display. Mean tracking performance as a function of time across 18 sessions demonstrated a monotonic increase in error from 0 to 11 min, and a performance plateau thereafter. Individual performance fluctuated widely around this trend - with an average root mean square (RMS) error of 2.3 disk radii. For each participant, moving estimates of blink duration and frequency, fixation dwell time and frequency, and mean pupil diameter were analyzed using non-linear regression and artificial neural network techniques. Individual models were derived using eye and performance data from one session and cross-validated on data from a second session run on a different day. A general regression model (based only on fixation dwell time and frequency) trained on data from both sessions from all participants produced a correlation of estimated to actual tracking performance of R=0.68 and an RMS error of 1.55 (S. D.=0.26) disk radii. Individual non-linear regression models containing a general linear model term produced the cross-session correlations of estimated to actual tracking performance of R=0.67. Individualized neural network models derived from the data of both experimental sessions produced the lowest RMS error (mean=1.23 disk radii, S.D.=0.13) and highest correlation (R=0.82) between eye activity-based estimates and actual tracking performance. Results suggest that information from multiple eye measures may be combined to produce accurate individualized real time estimates of sub-minute scale performance changes during sustained tasks. PMID- 10725566 TI - Anterior electroencephalographic asymmetry changes in elderly women in response to a pleasant and an unpleasant odor. AB - Greater left than right frontal EEG activation has been associated with increased positive and/or decreased negative affect, whereas greater right than left frontal activation has been associated with the opposite pattern. Substantial research has documented the trait properties of asymmetry, as well as responses to pleasant and unpleasant stimuli. The present study examined changes in anterior alpha asymmetries in response to pleasant (vanilla), unpleasant (valerian), and neutral (water) odors. As predicted, vanilla produced relative left frontal activation compared to valerian and water. Frontal asymmetry did not differ in response to valerian compared to water. The results are consistent with the hypothesis that the left frontal region of the brain is involved in positive/approach-related emotion, and extend previous results into the olfactory realm. PMID- 10725567 TI - A preliminary report relating frequency of vaginal intercourse to heart rate variability, Valsalva ratio, blood pressure, and cohabitation status. AB - The relationship between recalled frequency of penile-vaginal intercourse (FSI) and resting heart rate variability (HRV; an index of parasympathetic tone), resting diastolic blood pressure (DBP) and heart rate (HR) response to the Valsalva maneuver was examined in 51 healthy adults aged 20-47 (subjects scoring above the 86th percentile on the Lie scale of the Eysenck Personality Inventory (EPI) were excluded). As hypothesized, greater HRV and lower DBP were both associated with greater FSI (but not masturbation or non-coital sex with a partner) in cohabiting subjects, but not in non-cohabiting subjects. Valsalva ratio was unrelated to sexual behavior. Results are discussed in terms of both the modulating role of blood pressure on a number of psychological functions and the role of parasympathetic tone in HRV, FSI, and possibly pair-bonding. PMID- 10725568 TI - Serotonergic-1a activity and contingent negative variation. AB - While cholinergic, dopaminergic, noradrenergic, and gabaergic effects on contingent negative variation (CNV) have been largely described, little is known about serotonergic influence. Therefore, the relationship between CNV and serotonergic activity as reflected by prolactin (PRL) response to flesinoxan, a 5 HT(1A) full agonist, has been investigated in 28 healthy volunteers. To investigate the clinical implications of the relationship between CNV and serotonergic-1a activity, a group of 43 depressed patients was included in the study. Results among healthy volunteers showed a significant negative relationship between PRL response to flesinoxan and CNV amplitude at Fz, but no relationship for the other electrodes (Cz and Pz). In depressed patients, the relationships were not significant. Overall, this study does not support serotonergic effects on CNV. However, this information is indirect (correlations) and is limited to 5-HT(1A) activity. PMID- 10725569 TI - Genetics of male factor subfertility. PMID- 10725570 TI - Perinatal differential diagnosis of cystic kidney disease and urinary tract obstruction: anatomic pathologic, ultrasonographic and genetic findings. AB - According to the classification of Osathanondh and Potter of cystic kidney diseases an antenatal differential diagnosis is presented, which is based on the anatomic pathologic, ultrasonographic and genetic findings. Since the ultrasound evaluation influences the obstetric and neonatal management, each second and third trimester sonography should consider the most common malformations in pediatric autopsies. The autosomal recessive polycystic kidney disease (ARPK), autosomal dominant polycystic kidney disease (ADPK), multicystic renal dysplasia, obstructive multicystic kidneys and cystic renal malformations found in other syndromes with genetic linkage are discussed in this review. PMID- 10725571 TI - Urethral diverticula. AB - Urethral diverticula are a common cause of chronic genitourinary symptoms in women. They occur in three percent of women overall with higher frequencies in selected populations of symptomatic women. The peak incidence is between the ages of 25-45 but they affect all ages. The classical presentation is with recurrent urinary tract infections and post micturition dribbling but almost any urinary symptom may be a presenting feature. Reported cure rates following surgery approach 70% for recurrent urinary tract infection and almost 100% for local symptoms such as dyspareunia. However, despite this and the availability of effective diagnostic techniques diagnosis is often delayed. This is partly due to a lack of awareness among clinicians and partly because the condition overlaps the traditional territories of gynaecologists and urologists. PMID- 10725572 TI - Total hysterectomy for benign pathologies: direct costs comparison between laparoscopic and abdominal hysterectomy. AB - OBJECTIVES: The aim of this study is a direct costs comparison between laparoscopic and abdominal total hysterectomy when theses procedures are indicated for benign pathologies. STUDY DESIGN: To this end we compared the direct costs of total laparoscopic hysterectomy (TLH) calculated from a series of 105 patients with that obtained for a comparable series of 30 patients who underwent hysterectomy by laparotomy. RESULTS: The direct costs of total hysterectomy for a benign pathology by laparoscopic surgery and laparotomy are comparable (respectively 7693 French francs (FF) and 7759 FF). Whatever the type of operation the cost for staff represents 60% of the total cost. Expenditure for staff during the operation represents 41.0% of the total cost of TLH (3154 FF/7693 FF) whereas it represents only 31.0% of the cost of the operation when carried out by laparotomy (2406 FF/7759 FF) (P<0.0001). Inversely the expenditure due to staff during the post operative phase represents 24.1% of the total cost of the operation when laparotomy is used (1875 FF/7759 FF) and only 13.4% of the cost of the operation by laparoscopic surgery (1029 FF/7693 FF) (P<0.0001). When the operation uses laparoscopic surgery the increase in expenditure during the surgical act is compensated by the statistically significant shortening in the hospital stay. Expenditure connected with the laparoscopic surgery equipment is minimal compared to the costs connected with the staff. CONCLUSION: Provided that TLH is carried out with reusable laparoscopic surgery equipment, by skilled surgeons working in suitable hospital structures making the particularly heavy investment in laparoscopic surgery equipment economically viable, TLH is an economically viable technique as an alternative to laparotomy. PMID- 10725573 TI - Lethal fetal renal anomalies and obstetric outcome. AB - The incidence of prenatal and intrapartum complications was examined among 33 pregnancies complicated by lethal fetal renal abnormalities (cases) and compared to 200 contemporaneous control pregnancies (controls) by retrospective record review. Cases experienced higher rates of antepartum bleeding (29% vs. 6%, p<0.0001) stillbirth (15% vs. 0%, p<0.0001), preterm birth (34.3+/-4.1 vs. 39.7+/ 1.8, p<0. 0001) and breech presentation (48% vs. 4%, p<0.0001). Twenty-six of 33 cases had lung weights /= 0.39). CONCLUSION: The risks of developing permanent symptomatic sequelae from AVM radiosurgery vary dramatically with location and, to a lesser extent, volume. These risks can be predicted according to the SPIE location-risk score and the 12 Gy-Volume. PMID- 10725625 TI - Combining stereotactic angiography and 3D time-of-flight magnetic resonance angiography in treatment planning for arteriovenous malformation radiosurgery. AB - PURPOSE: This study was initiated to evaluate the advantages of using three dimensional time-of-flight magnetic resonance angiography (3D TOF MRA), as an adjuvant to conventional stereotactic angiography, in obtaining three-dimensional information about an arteriovenous malformation (AVM) nidus and in optimizing radiosurgical treatment plans. METHODS AND MATERIALS: Following angiography, contrast-enhanced MRI and MRA studies were obtained in 22 consecutive patients undergoing Gamma Knife radiosurgery for AVM. A treatment plan was designed, based on the angiograms and modified as necessary, using the information provided by MRA. The quantitative analysis involved calculation of the ratio of the treated volume to the MRA nidus volume (the tissue volume ratio [TVR]) for the initial and final treatment plans. RESULTS: In 12 cases (55%), the initial treatment plans were modified after including the MRA information in the treatment planning process. The mean TVR for the angiogram-based plans was 1.63 (range 1.17-2.17). The mean coverage of the MRA nidus by the angiogram-based plans was 93% (range 73 99%). The mean MRA nidus volume was 2.4 cc (range 0. 6-5.3 cc). The MRA-based modifications resulted in increased conformity with the mean TVR of 1.46 (range 1.20-1.74). These modifications were caused by MRA revealing irregular nidi and/or vascular components superimposed on the angiographic projections of the nidi. In a number of cases, the information from MRA was essential in defining the nidus when the projections of the angiographic outlines showed different superior and/or inferior extent of the nidus. In two cases, MRA revealed irregular nidi, correlating well with the angiograms and showed that the angiographically acceptable plans undertreated 27% of the MRA nidus in one case and 18% of the nidus in the other case. In the remaining 10 cases (45%), both MRI and MRA failed to detect the nidus due to surgical clip artifacts and the presence of embolizing glue. CONCLUSIONS: The 3D TOF MRA provided information on irregular AVM shape, which was not visualized by angiography alone, and it was superior to MRI for defining the AVM nidus. However, when imaging artifacts obscured the AVM nidus on MRI and MRA, angiography permitted detection of AVM. Utilizing MRA as a complementary imaging modality to angiography increased accuracy of the AVM radiosurgery and allowed for optimal dose planning. PMID- 10725626 TI - Radiosurgery for brain metastases: a score index for predicting prognosis. AB - PURPOSE: To analyze a prognostic score index for patients with brain metastases submitted to stereotactic radiosurgery (the Score Index for Radiosurgery in Brain Metastases [SIR]). METHODS AND MATERIALS: Actuarial survival of 65 brain metastases patients treated with radiosurgery between July 1993 and December 1997 was retrospectively analyzed. Prognostic factors included age, Karnofsky performance status (KPS), extracranial disease status, number of brain lesions, largest brain lesion volume, lesions site, and receiving or not whole brain irradiation. The SIR was obtained through summation of the previously noted first five prognostic factors. Kaplan-Meier actuarial survival curves for all prognostic factors, SIR, and recursive partitioning analysis (RPA) (RTOG prognostic score) were calculated. Survival curves of subsets were compared by log-rank test. Application of the Cox model was utilized to identify any correlation between prognostic factors, prognostic scores, and survival. RESULTS: Median overall survival from radiosurgery was 6.8 months. Utilizing univariate analysis, extracranial disease status, KPS, number of brain lesions, largest brain lesion volume, RPA, and SIR were significantly correlated with prognosis. Median survival for the RPA classes 1, 2, and 3 was 20.19 months, 7.75 months, and 3. 38 months respectively (p = 0.0131). Median survival for patients, grouped under SIR from 1 to 3, 4 to 7, and 8 to 10, was 2.91 months, 7.00 months, and 31.38 months respectively (p = 0.0001). Using the Cox model, extracranial disease status and KPS demonstrated significant correlation with prognosis (p = 0.0001 and 0.0004 respectively). Multivariate analysis also demonstrated significance for SIR and RPA when tested individually (p = 0.0001 and 0.0040 respectively). Applying the Cox Model to both SIR and RPA, only SIR reached independent significance (p = 0.0004). CONCLUSIONS: Systemic disease status, KPS, SIR, and RPA are reliable prognostic factors for patients with brain metastases submitted to radiosurgery. Applying SIR and RPA classifications to our patients' data, SIR demonstrated better accuracy in predicting prognosis. SIR should be further tested with larger patient accrual and for all patients with brain metastases subjected or not to stereotactic radiosurgery. PMID- 10725627 TI - Prognostic factors in metastatic spinal cord compression: a prospective study using multivariate analysis of variables influencing survival and gait function in 153 patients. AB - PURPOSE: Based on a very large patient cohort followed prospectively for at least a year or until death, we analyzed the prognostic significance of various clinical and radiological variables on posttreatment ambulatory function and survival. METHODS AND MATERIALS: During a 312-year period we prospectively included 153 consecutive patients with a diagnosis of spinal cord compression due to metastatic disease. The patients were followed with regular neurological examinations by the same neurologist for a minimum period of 11 months or until death. The prognostic significance of five variables on gait function and survival time after treatment was analyzed. RESULTS: The type of the primary tumor had a direct influence on the interval between the diagnosis of the primary malignancy and the occurrence of spinal cord compression (p < 0. 0005), and on the ambulatory function at time of diagnosis (p = 0. 016). There was a clear correlation between the degree of myelographic blockage and gait function (p = 0.000) and between gait function and sensory disturbances (p = 0.000). The final gait was dependent on the gait function at time of diagnosis (p < 0.0005). Survival time after diagnosis depended directly on the time from primary tumor diagnosis until spinal cord compression (p = 0.002), on the ambulatory function at the time of diagnosis (p = 0.018), and on the ambulatory function after treatment. CONCLUSIONS: The pretreatment ambulatory function is the main determinant for posttreatment gait function. Survival time is rather short, especially in nonambulatory patients, and can only be improved by restoration of gait function in nonambulatory patients by immediate treatment. PMID- 10725628 TI - CNS germinoma: disease control and long-term functional outcome for 12 children treated with craniospinal irradiation. AB - PURPOSE: To provide evidence that radiation therapy alone in the form of craniospinal irradiation (CSI) and a boost to the primary site of disease provides effective disease control and limited additional morbidity for patients with CNS germinoma. METHODS AND MATERIALS: Twelve patients with a median age of 12 years (range 9-16 years) with CNS germinoma were treated with CSI (median 25.6 Gy, range 23.4-32 Gy) and a boost to the primary site of disease (50.4 Gy, range 45-54 Gy) between January 1987 and June 1998. All patients were biopsied prior to radiation therapy and none received chemotherapy. No patients were lost to follow up and the majority had long-term (> 45 month) pre- and postirradiation endocrine and psychology assessment. RESULTS: All 12 patients are alive and no failures have occurred with a median follow-up of 69 months (range 14-143 months). Preirradiation endocrine deficiencies were present in 6 of 6 suprasellar tumors and 1 of 6 pineal tumors; with follow-up there was no substantial difference between age and gender adjusted pre- and postirradiation stature and weight. With long-term follow-up, there were no significant differences between pre- and postirradiation full-scale, verbal, and performance IQ scores. CONCLUSIONS: This study confirms the ability of radiation therapy alone to achieve disease control with a high rate of success in pediatric patients and demonstrates that the treatment toxicity faced by these patients may be less than anticipated. Because these patients present with substantial preexisting morbidity at diagnosis and may be of an age where the potential for radiation-related side effects is relatively small, the superiority of treatment alternatives may be difficult to prove. PMID- 10725629 TI - Perioperative and postoperative complications of intracavitary radiation for FIGO stage I-III carcinoma of the cervix. AB - PURPOSE: To evaluate perioperative and postoperative complications of low-dose rate (LDR) intracavitary radiation therapy in patients with FIGO Stage I-III carcinoma of the uterine cervix. METHODS AND MATERIALS: We retrospectively reviewed the medical and radiotherapy records of all patients treated with radiation between 1960 and 1992 at The University of Texas M. D. Anderson Cancer Center for FIGO I-III carcinomas of the cervix. Patients who had had initial hysterectomy or whose treatment did not include intracavitary irradiation were excluded. The final study included 4043 patients who had undergone 7662 intracavitary procedures. RESULTS: Eleven (0. 3%) patients had documented or suspected cases of thromboembolism resulting in 4 deaths. Of these 11 patients, 8 had clinical or radiographic evidence of tumor involving pelvic nodes or fixed pelvic wall. The risk of postoperative thromboembolism did not decrease significantly with the routine use of mini-dose heparin prophylaxis (p = 0.3). Other life-threatening perioperative complications included myocardial infarction (1 death in 5 patients), cerebrovascular accident (2 patients), congestive heart failure or atrial fibrillation (3 patients), and halothane liver toxicity (2 deaths in 2 patients). Intraoperative complications included uterine perforation (2.8%) and vaginal laceration (0.3%), which occurred more frequently in patients >/= 60 years old (p < 0.01). Fourteen percent of patients had a temperature >/= 101 degrees F during at least one hospital stay. The only correlation between minor intraoperative complications and disease-specific survival was found in patients who had Stage III disease and uterine perforation; survival was significantly (p = 0.01) decreased in these patients. CONCLUSIONS: Fatal or life threatening complications of intracavitary treatment were very rare. Deep venous thrombosis (DVT) and pulmonary embolism (PE) did not occur in otherwise healthy patients with early disease and were rare even when disease was more advanced. Minor perioperative complications were not correlated with serious late complications or with death from disease. PMID- 10725630 TI - High-dose-rate afterloading brachytherapy in carcinoma of the cervix: an experience of 1992 patients. AB - PURPOSE: To report the results of radiation therapy in carcinoma of the cervix treated by external irradiation and high-dose-rate (HDR) intracavitary brachytherapy. METHODS AND MATERIALS: This is a retrospective analysis of 2,063 patients with histologically proven carcinoma of the cervix treated by external irradiation and HDR intracavitary brachytherapy between March 1985-December 1991. The Kaplan-Meier method was used for survival and disease-free survival analysis. Late complications in the bowel and bladder were calculated actuarially. RESULTS: There were 71 patients who did not complete the course of irradiation so only 1992 patients were retrospectively analyzed for survival. There were 2 patients (0.1%) in Stage IA, 211 (10.2%) Stage IB, 225 (10.9%) in Stage IIA, 902 (43. 7%) in Stage IIB, 14 (0.7%) in Stage IIIA, 675 (32.7%) in Stage IIIB, 16 (0.8%) in Stage IVA, and 16 (0.8%) in Stage IVB. The median follow-up time was 96 months. The actuarial 5-year disease-free survival rate was 79.5%, 70.0%, 59.4%, 46.1%, 32.3%, 7.8%, and 23.1% for Stage IB, IIA, IIB, IIIA, IIIB, IVA, and IVB respectively. The actuarial 5-year disease-free survival rate for Stage IB(1) and IB(2) squamous cell carcinoma was 88.7% and 67.0%. The actuarial 5-year overall survival rate was 86.3%, 81.1%, 73.0%, 50.3%, 47.8%, 7.8%, and 30.8% for Stage IB, IIA, IIB, IIIA, IIIB, IVA, and IVB respectively. Pattern of failure revealed 20.8% local recurrence, 18. 7% distant metastases, and 4% in both. The late complication rate Grade 3 and 4 (RTOG) for bowel and bladder combined was 7.0% with 1. 9% Grade 4. CONCLUSION: HDR brachytherapy used in this series produced pelvic control and survival rates comparable to other LDR series. PMID- 10725631 TI - Radiation treatment in recurrent squamous cell cancer of the vulva. AB - PURPOSE: To evaluate the treatment and outcome of recurrent vulvar cancer. METHODS AND MATERIALS: In a retrospective review of 26 women referred to the department of radiation oncology between 1982 and 1995, patient records were analyzed with respect to the findings at original surgery, the time to locoregional recurrence, the location of the recurrence, and the subsequent management and outcome. RESULTS: Sixteen recurrences were managed with a combination of surgery and radiotherapy, and the remainder with radiation treatment, combined with chemotherapy in some cases. The overall survival for the entire cohort at 5 years was 22%. The 5-year survival for those with recurrence confined to the vulva (n = 13) was 46%, compared with 0% for those women with a recurrence located or extending beyond the vulva (p = 0.002). The local control rate was 34.6%. CONCLUSION: Our results confirm the poor overall prognosis for this condition. In particular, they highlight the importance of the location of the recurrence as a prognostic indicator. Based on this review, we conclude that radiotherapy fields should encompass the region at risk if the intent is curative. Finally, low-dose palliation for groin node recurrence is ineffectual. PMID- 10725632 TI - Prostate brachytherapy in patients with prostate volumes >/= 50 cm(3): dosimetic analysis of implant quality. AB - OBJECTIVES: Permanent implantation with (125)I in patients with localized prostate cancer who have prostate volumes >/= 50 cm(3) is often technically difficult owing to pubic arch interference. The objective of this study was to describe dosimetry outcomes in a group of patients who were implanted using the real-time ultrasound-guided technique who had prostate volumes >/= 50 cm(3). MATERIALS AND METHODS: A total of 331 patients received an (125)I prostate seed implant from January 1, 1995, to June 1, 1999, of whom 66 (20%) had prostate volumes >/= 50 cm(3) at the time of the procedure. The real-time seed implant method was used in all patients and consisted of intraoperative planning and real time seed placement using a combination of axial and sagittal ultrasound imaging. Pubic arch interference was managed using an extended lithotomy position or by angling the tip of the ultrasound probe in an anterior direction. No preimplant pubic arch CT scan study was performed and no patients were excluded from treatment because of prostate size. Implant quality was assessed using CT-based dosimetry performed 1 month postimplant. Dose-volume histograms for the prostate, bladder, rectum, and urethra volumes were generated. The target dose for these implants was 160 Gy and an adequate implant was defined as the dose delivered to 90% of the prostate (D90) >/= 140 Gy. The dose delivered to 95% of the prostate (D95) and doses to 30% of the rectal (DRECT30) and urethral (DURE30) volumes were also calculated. RESULTS: Prostate volumes in the 66 patients ranged from 50 to 93 cm(3) (median 57, mean 61 cm(3)). Total activity implanted was 27.8-89.1 mCi (median 57 mCi), with a range in activity per seed of 0.36-0.56 mCi (median 0.4 mCi). The prostate D90s and D95s ranged from 13,245 to 22,637 cGy (median 18,750) and 11,856 to 20,853 cGy (median 16,725), respectively. Only one patient (1.5%) had a D90 < 140 Gy. The DURE30 values ranged from 15,014 to 27,800 cGy (median 20,410) and the DRECT30 values were 3137-9910 cGy (median 5515). CONCLUSION: Implantation of the large prostate can be accomplished using the real-time method. A total of 98.5% of the patients receive a high-quality implant. In addition, these implants should not put patients at increased risk for significant urinary and bowel complications because urethral and rectal doses can be kept at acceptable levels. PMID- 10725633 TI - Needle displacement during HDR brachytherapy in the treatment of prostate cancer. AB - PURPOSE: We used clinical patient data to examine implant displacement between high dose rate (HDR) brachytherapy fractions for prostate cancer to determine its impact on treatment delivery. MATERIALS AND METHODS: We analyzed the verification films taken prior to each fraction for 96 consecutive patients treated with HDR brachytherapy boosts as part of their radiation therapy for definitive treatment of organ-confined prostate cancer at our institution. Patients were treated with 18-24 Gy in 4 fractions of HDR delivered in 40 hours followed by 36-39.6 Gy external beam radiation to the prostate. We determined the mean and maximum displacement distances of marker seeds placed in the prostate and of the implanted needles between HDR fractions. RESULTS: Mean and maximum displacement distances between fractions were documented up to 7.6 mm and 28.5 mm, respectively, for the implant needles and 3.6 mm and 11.4 mm, respectively, for the gold marker seeds. All displacement of implant needles occurred in the caudal direction. At least 1 cm caudal displacement of needles occurred prior to 15.5% all fractions. Manual adjustment of needles was required prior to 15% of fractions, and adjustment of the CLP only was required in 24%. Most of the displacement for both the marker seeds and needles occurred between the first and second fractions. CONCLUSIONS: There is significant caudal displacement of interstitial implant needles between HDR fractions in our prostate cancer patients. Obtaining verification films and making adjustments in the treatment volume prior to each fraction is necessary to avoid significant inaccuracies in treatment delivery. PMID- 10725634 TI - Apoptosis, p53, bcl-2, and Ki-67 in invasive bladder carcinoma: possible predictors for response to radiochemotherapy and successful bladder preservation. AB - PURPOSE: Several groups have reported the value of bladder preservation by a combined treatment protocol, including transurethral resection (TUR-B) and radiochemotherapy (RCT). As more experience is acquired with organ-sparing treatment, patient selection should be optimized. The purpose of this study was to investigate the role of several biologic markers that may predict response to RCT in muscle-invasive bladder carcinoma. METHODS AND MATERIALS: The apoptotic index (AI), Ki-67, p53, and bcl-2 were evaluated by immunohistochemistry on pretreatment biopsies from 70 patients treated for invasive bladder cancer by TUR B and RCT. Expression of each marker was correlated with initial response, local control, and cancer-specific survival with preserved bladder. An exploratory multivariate analysis was also performed that included clinical and immunohistochemical variables. RESULTS: A high AI (> median = 1.6%) and a high Ki 67 index (> median = 8.8%), but not the p53- and bcl-2 expression, were significantly related to initial complete response (CR) and local control with preserved bladder after 5 years. When the AI and Ki-67 expression were considered simultaneously, the association with initial CR (p < 0. 001), local control (p = 0.0002), and cancer-specific survival with preserved bladder (p = 0.008) was highly significant. In an exploratory multivariate analysis (final model), only AI, Ki-67, and the combined AI/Ki-67 variable retained significance for local control with preserved bladder at 5 years. CONCLUSION: Patients with a high spontaneous AI and a high pretreatment Ki-67 index should be considered preferentially for treatment with RCT, whereas tumors with low proliferation and low levels of apoptosis are less likely to respond to RCT. PMID- 10725635 TI - A preliminary outcome analysis of the Patterns of Care Study in Japan for esophageal cancer patients with special reference to age: non surgery group. AB - BACKGROUND: The Patterns of Care Study (PCS) was imported to Japan from the United States in July 1996. A preliminary outcome analysis of the PCS for esophageal cancer patients in Japan was made with special reference to age, because the elderly population is rapidly increasing in Japan. PATIENTS AND METHODS: From July 1996 to February 1998, external PCS audits were performed for 37 institutions nationwide and detailed information of 561 esophageal cancer patients treated during the period 1992-1994 was collected by using the fifth PCS data format developed in the United States. This format was provided courtesy of the American College of Radiology. For this study, patients who had not undergone surgery (n = 336) were selected. The patients were classified into three age groups: < 65 years old (n = 119), between 65 and 74 years (n = 93), and 75 years or older (n =123). Cox's proportional hazards model was used for the statistical analysis, with survival, acute/subacute complication and late complication of grade 3 or more based on RTOG criteria, as the endpoints. RESULTS: Significant prognostic factors for the entire non-surgery group were Karnofsky Performance Status (KPS) (p = 0.0007), stage (p = 0.0001), and external irradiation dose (p = 0.0001). For the younger group, KPS (p = 0.0004), stage (p = 0.0197), and utilization of brachytherapy (p = 0.0010) were significant, while for the intermediate age group it was KPS (p = 0. 0027), history of pulmonary disease (p = 0.0339), stage (p = 0.0001), and external dose (p = 0.0001), and for the elderly group, stage (p = 0.0001) and external irradiation dose (p = 0.0224) were significant. Significant risk factors for complications for the entire group were stage (p = 0.0411), external dose (p = 0.0163), and stratification of institution (academic vs. nonacademic) (p = 0. 0114). Significant risk factors for the younger group were history of pulmonary disease (p = 0.0495) and external dose (p = 0.0037), and the other age groups showed no significant risk factors. CONCLUSION: Age was not a significant prognostic or risk factor for esophageal cancer patients in the non-surgery group treated with radiation therapy. Therefore, radiation therapy represented an important treatment modality for the elderly as well as for the younger esophageal cancer patients. External dose was a treatment-related prognostic factor for the elderly as well as for the intermediate age group. PMID- 10725636 TI - Treatment of nonlaparotomized (clinical) stage I and II Hodgkin's disease patients by extended field and splenic irradiation. AB - PURPOSE: At the New York Presbyterian Hospital-Cornell Medical Center, patients with unequivocal clinical stage I and IIA Hodgkin's disease (HD) have been treated with mantle, splenic, and extended field radiation therapy (EFRT) (without surgical staging). A 24-year retrospective review was conducted to determine the effectiveness of our patient selection on the outcome of patients treated with this modality. METHODS AND MATERIALS: During the period 1971 to 1994, 94 patients with clinically staged HD, with favorable prognostic factors, were retrospectively reviewed. Patients with pathological or equivocal staging, "B" symptoms, bulk disease, history of previous chemotherapy, and/or Stage III or IV disease were excluded from our analysis. There were 27 Stage IA and 67 Stage IIA patients. All patients were treated to 3600 cGy with a 400 cGy boost to the involved field. The median follow-up was 52 months, mean of 62.1 months. RESULTS: Ten of 94 patients (10.5%) relapsed. Seven of the relapses were in the pelvis, one submandibularily, one in the tonsil, and one in the axilla. Nine of the relapses had nodular sclerosis histology, one had lymphocyte predominance, and none had mixed cellularity. The median time to relapse was 38 months; mean time 42. 3 months. All patients are alive, well and free of disease, including nine who received subsequent chemotherapy and one who underwent autotransplantation. CONCLUSIONS: Careful clinical staging of early, asymptomatic HD patients treated with mantle, splenic, and EFRT may obviate the need for exploratory laparotomy. PMID- 10725637 TI - Late effects in children treated with radiation therapy for Wilms' tumor. AB - PURPOSE: To determine the frequency and types of late effects in children receiving radiation therapy (RT) for Wilms' tumor. MATERIALS AND METHODS: From 1968 to 1994, 55 children received megavoltage RT at our institution as part of treatment for Wilms' tumor. A total of 42 (76.4%) have survived and have a minimum follow-up of 5 years. There were 25 female and 17 male patients with a median age at diagnosis of 48 months (range, 7-126 months). There were 12 Stage I, eight Stage II, 15 Stage III, six Stage IV, and one Stage V patient. RT was delivered to the hemiabdomen in 36 and whole abdomen in six patients. RT dose was 1000-1200 cGy (Group A) in 12, 1201-2399 cGy (Group B) in 11, and 2400-4000 cGy (Group C) in 19. Whole-lung RT was delivered to 13 patients either at diagnosis or pulmonary relapse. All patients received chemotherapy; the most common agents were actinomycin-D/vincristine/adriamycin in 13 and actinomycin-D/vincristine in 18. Median follow-up was 181 months (range, 60-306 months). RESULTS: Of 42 patients, 13 (31.0%) did not have late effects of treatment. The number of patients who developed muscular hypoplasia, limb length inequality, kyphosis, and iliac wing hypoplasia were seven (16.7%), five (11.9%), three (7.1%), and three (7.1%), respectively. Scoliosis was seen in 18 (42.9%) with only one patient requiring orthopedic intervention. Median time to development of scoliosis was 102 months, with a range of 16-146 months. The actuarial incidence of scoliosis at 5, 10, and 15 years after RT was 4.8 +/- 3.3%, 51.8 +/- 9.0%, and 56.7 +/- 9.3%, respectively. Only one of 12 Group A patients developed scoliosis. The 10- and 15-year actuarial incidences of scoliosis for Group A and B patients were 37.7 +/- 12.4% and 37.7 +/- 12.4%, whereas for Group C patients the incidences were 65.8 +/- 12.0% and 74.4 +/- 11. 7% (p = 0.03, log rank test). The actuarial incidence of bowel obstruction at 5, 10, and 15 years was 9.5 +/- 4.5%, 13.0 +/- 5.6%, and 17.0 +/- 6.5%. Of 23 patients, five irradiated within 10 days of surgery and one of 19 irradiated after 10 days developed bowel obstruction (p = 0.09, log rank test). Three patients developed hypertension with normal blood urea nitrogen (BUN) and creatinine levels; another patient had chronic renal insufficiency in a nonirradiated kidney. One patient developed diffuse interstitial pneumonitis. Of the 19 female patients who have reached puberty, three have given birth, and 15 have regular and one has irregular menstrual periods. Four patients developed benign neoplasms; three were in the RT field (two osteochondroma, one lipoma) and one outside (cervical intraepithelial neoplasia II). There were three second malignancies (chronic myelogenous leukemia at 9 years, osteosarcoma at 11 years, and breast cancer at 25 years after initial diagnosis of nephroblastoma); both solid malignancies occurred in the RT field. CONCLUSIONS: Late effects of therapy were seen in more than two thirds of children treated for Wilms' tumor. Children treated with lower doses (<2400 cGy) had a lower incidence of scoliosis compared with those who received more than 2400 cGy. There is also a suggestion that the incidence is lower in patients who received 1000-1200 cGy. Severe physical and functional deformity from RT was uncommon. PMID- 10725638 TI - Studies of physiology and the morphology of the cat LGN following proton irradiation. AB - PURPOSE: We have examined the effects of proton irradiation on the histologic and receptive field properties of thalamic relay cells in the cat visual system. The cat lateral geniculate nucleus (LGN) is a large structure with well-defined anatomical boundaries, and well-described afferent, efferent, and receptive field properties. METHODS AND MATERIALS: A 1.0-mm proton microbeam was used on the cat LGN to determine short-term (3 months) and long-term (9 months) receptive field effects of irradiation on LGN relay cells. The doses used were 16-, 40-, and 60 gray (Gy). RESULTS: Following irradiation, abnormalities in receptive field organization were found in 40- and 60-Gy short-term animals, and in all of the long-term animals. The abnormalities included "silent" areas of the LGN where a visual response could not be evoked and other regions that had unusually large or small compound receptive fields. Histologic analysis failed to identify cellular necrosis or vascular damage in the irradiated LGN, but revealed a disruption in retinal afferents to areas of the LGN. CONCLUSIONS: These results indicate that microbeam proton irradiation can disrupt cellular function in the absence of obvious cellular necrosis. Moreover, the area and extent of this disruption increased with time, having larger affect with longer post-irradiation periods. PMID- 10725639 TI - Overexpression of the 27 KDa heat shock protein is associated with thermoresistance and chemoresistance but not with radioresistance. AB - PURPOSE: The heat shock protein (hsp27) correlates with thermotolerance and chemoresistance. Our main objective was to assess the response to radiotherapy both in vitro and in vivo in correlation with various concentrations of hsp27. The second objective was to evaluate the relation between hsp27 and glutathione-s transferase pi (GST pi). METHODS AND MATERIALS: For the in vitro study, thermoresistant cell lines, expressing various amounts of hsp27, were used to assess the role of this protein in radioresistance. To verify the efficiency of hsp27 in these cells lines to confer resistance to cytotoxic agents, these cells were also treated with heat shock and cisplatin. Furthermore, the role of hsp27 expression was studied in vivo by immunochemistry in 98 patients with head and neck squamous cell carcinoma treated by radiotherapy. hsp27 was correlated with local control of the tumor and with clinical and biologic factors potentially able to affect the local control, including p53, ki-67, ploidy, and GST. RESULTS: In vitro, high constitutive levels of expression of hsp27 did not significantly influence the survival curves of transfected cells exposed to radiation as compared to control cells although hsp27 overexpression was confirmed to increased the cellular resistance to heat and to cisplatinum. In vivo, we showed that overexpression of various amounts of hsp27 did not correlate with local control of the tumor. In vivo, hsp27 was only significantly associated with GST pi. Expression of GST pi was associated with poor local (p = 0.01) control and survival (p = 0.08) in a Cox model. CONCLUSIONS: It is concluded that the mechanisms responsible for hsp27-mediated heat and drug resistance are not involved in radioprotection. PMID- 10725640 TI - High-dose-rate brachytherapy: dose escalation in three-dimensional miniorgans of the human bronchial wall. AB - PURPOSE: High-dose-rate (HDR) brachytherapy of human lung cancer is well established, however fractionation schemes and dosages are based mainly on experience. The aim of this investigation was to study the effects of different doses of HDR iridium-192 on normal human bronchial epithelium in three dimensional miniorgans of the human bronchial wall. METHODS AND MATERIALS: Forty eight biopsies from normal bronchi were cultivated for 14 days and exposed at random to different doses of HDR iridium 192 (0 Gy, 30 Gy, 45 Gy, 60 Gy, or 75 Gy). Cell viability was assessed immediately after irradiation, after 4 or 18 days by fluorescent staining, and cell damage of the culture was analyzed by light microscopy. Lactate dehydrogenase (LDH) was measured in the supernatant for 4 days. RESULTS: There was no histologically apparent tissue damage regardless of the irradiation dose. The number of nonvital cells increased in irradiated miniorgans depending on the dose used (p < 0. 05 at 75 Gy). This effect occurred early and was less pronounced with time. LDH measurements showed an increase only in the first 24 hours. CONCLUSIONS: Our results confirm that normal bronchial epithelium has a high tolerance to early epithelial damage by irradiation. This model of human bronchial miniorgans is useful for further studies of the effects of irradiation on human bronchi. PMID- 10725641 TI - In vitro toxicity of (191)Pt-labeled cisplatin to a human cervical carcinoma cell line (ME-180). AB - PURPOSE: The aim of the present work was to examine the effect of (191)Pt cisplatin, and to study the manner in which radiation and cisplatin interact, in a human cervical carcinoma cell line (ME-180). METHODS AND MATERIALS: The cells were incubated for 1 hour with nonradioactive cisplatin or (191)Pt-cisplatin with specific activities in the range 48-167 MBq/mg. The surviving fraction of the cells after 7 days' growth was determined with a nonclonogenic tetrazolium-based (MTT) assay. The uptake of platinum into the cell and the amount of platinum bound to DNA was measured. RESULTS: The 50% inhibition concentration (IC(50)) decreased with increasing specific activity of the (191)Pt-cisplatin. For the specific activities 0 (nonradioactive), 48, 89, 143, 157, and 167 MBq/mg, IC(50) was found to be 3.24 +/- 0.08, 2.77 +/- 0.55, 2.17 +/- 0.34, 1.15 +/- 0.04, 1.02 +/- 0.03, and 0.76 +/- 0.13 respectively. Isobologram analysis showed a supra additive (synergistic) interaction between the radiotoxicity and chemotoxicity for specific activities over 100 MBq/mg. CONCLUSION: The cytotoxic effect of cisplatin may be enhanced by labeling the drug with the radionuclide (191)Pt. PMID- 10725642 TI - 3-D portal image analysis in clinical practice: an evaluation of 2-D and 3-D analysis techniques as applied to 30 prostate cancer patients. AB - PURPOSE: To investigate the clinical importance and feasibility of a 3-D portal image analysis method in comparison with a standard 2-D portal image analysis method for pelvic irradiation techniques. METHODS AND MATERIALS: In this study, images of 30 patients who were treated for prostate cancer were used. A total of 837 imaged fields were analyzed by a single technologist, using automatic 2-D and 3-D techniques independently. Standard deviations (SDs) of the random, systematic, and overall variations, and the overall mean were calculated for the resulting data sets (2-D and 3-D), in the three principal directions (left-right [L-R], cranial-caudal [C-C], anterior-posterior [A-P]). The 3-D analysis included rotations as well. For the translational differences between the three data sets, the overall SD and overall mean were computed. The influence of out-of-plane rotations on the 2-D registration accuracy was determined by analyzing the difference between the 2-D and 3-D translation data as function of rotations. To assess the reliability of the 2-D and 3-D methods, the number of times the automatic match was manually adjusted was counted. Finally, an estimate of the workload was made. RESULTS: The SDs of the random and systematic components of the rotations around the three orthogonal axes were 1. 1 (L-R), 0.6 (C-C), 0.5 (A P) and 0.9 (L-R), 0.6 (C-C), 0.8 (A-P) degrees, respectively. The overall mean rotation around the L-R axis was 0.7 degrees, which deviated significantly from zero. Translational setup errors were comparable for 2-D and 3-D analysis (ranging from 1.4 to 2.2 mm SD and from 1.5 to 2.5 mm SD, respectively). The variation of the difference between the 2-D and 3-D translation data increased from 1.1 mm (SD) for zero rotations to 2.7 mm (SD) for out-of-plane rotations of 3 degrees, due to a reduced 2-D registration accuracy for large rotations. The number of times the analysis was not considered acceptable and was manually adjusted was 44% for the 2-D analysis, and 6% for the 3-D analysis. CONCLUSION: True 3-D analysis of setup errors for a group of 30 patients with prostate cancer demonstrated that setup rotations are rather small. The deformation of the projected anatomy in portal images caused by out-of-plane rotations leads to a reduced 2-D registration accuracy. For rotations larger than 3 degrees this effect can be quite pronounced, making 3-D registration the preferred method. Furthermore, the automatic 3-D registration has a higher success rate, most likely because this technique uses more information compared to the 2-D method. PMID- 10725643 TI - Image localization for frameless stereotactic radiotherapy. AB - PURPOSE: Infrared light-emitting diodes (IRLEDs) have been used for optic-guided stereotactic radiotherapy localization at the University of Florida since 1995. The current paradigm requires stereotactic head ring placement for the patient's first fraction. The stereotactic coordinates and treatment plan are determined relative to this head ring. The IRLEDs are attached to the patient via a maxillary bite plate, and the position of the IRLEDs relative to linac isocenter is saved to file. These positions are then recalled for each subsequent treatment to position the patient for fractionated therapy. The purpose of this article was to report a method of predicting the desired IRLED locations without need for the invasive head ring. METHODS AND MATERIALS: To achieve the goal of frameless optic guided radiotherapy, a method is required for direct localization of the IRLED positions from a CT scan. Because it is difficult to localize the exact point of light emission from a CT scan of an IRLED, a new bite plate was designed that contains eight aluminum fiducial markers along with the six IRLEDs. After a calibration procedure to establish the spatial relationship of the IRLEDs to the aluminum fiducial markers, the stereotactic coordinates of the IRLED light emission points are determined by localizing the aluminum fiducial markers in a stereotactic CT scan. RESULTS: To test the accuracy of direct CT determination of the IRLED positions, phantom tests were performed. The average accuracy of isocenter localization using the IRLED bite plate was 0.65 +/- 0. 17 mm for these phantom tests. In addition, the optic-guided system has a unique compatibility with the stereotactic head ring. Therefore, the isocentric localization capability was clinically tested using the stereotactic head ring as the absolute standard. The ongoing clinical trial has shown the frameless system to provide a patient localization accuracy of 1.11 +/- 0.3 mm compared with the head ring. CONCLUSION: Optic-guided radiotherapy using IRLEDs provides a mechanism through which setup accuracy may be improved over conventional techniques. To date, this optic-guided therapy has been used only as a hybrid system that requires use of the stereotactic head ring for the first fraction. This has limited its use in the routine clinical setting. Computation of the desired IRLED positions eliminates the need for the invasive head ring for the first fraction. This allows application of optic-guided therapy to a larger cohort of patients, and also facilitates the initiation of extracranial optic-guided radiotherapy. PMID- 10725644 TI - Brachytherapy radiation doses to the neurovascular bundles. AB - PURPOSE: To investigate the role of radiation dose to the neurovascular bundles (NVB) in brachytherapy-related impotence. METHODS AND MATERIALS: Fourteen Pd-103 or I-125 implant patients were studied. For patients treated with implant alone, the prostate and margin (clinical target volume [CTV]) received a prescription dose of 144 Gy for I-125 or 115 Gy for Pd-103. Two patients received Pd-103 (90 Gy) with 46 Gy supplemental external beam radiation (EBRT). Axial CT images were acquired 2 to 4 hours postoperatively for postimplant dosimetry. Because the NVBs cannot be visualized on CT, NVB calculation points were determined according to previously published anatomic descriptions. Bilateral NVB points were considered to lie posterior-laterally, approximately 2 mm from the prostatic capsule. NVB doses were recorded bilaterally, at 0.5-cm intervals from the prostatic base. RESULTS: For Pd-103, the average NVB doses ranged from 150 Gy to 260 Gy, or 130% to 226% of the prescription dose. For I-125, the average NVB dose ranged from 200 Gy to 325 Gy, or 140% to 225% of the prescription dose. These was no consistent relationship between the NVB dose and the distance from the prostatic base. To examine the possible effect of minor deviations of our calculation points from the true NVB location, we performed NVB calculations at points 2 mm medial or lateral from the NVB calculation point in 8 patients. Doses at these alternate calculation points were comparable, although there was greater variability with small changes in the calculation point if sources were located outside the capsule, near the NVB calculation point. Three patients who developed early postimplant impotence had maximal NVB doses that far exceeded the average values. CONCLUSIONS: In the next few years, we hope to clarify the role of high NVB radiation doses on potency, by correlating NVB dose calculations with a large number of patients enrolled in an ongoing I-125 versus Pd-103 trial for early stage patients, for whom detailed dosimetric and potency data are being collected prospectively. In the future, we anticipate that NVB doses may be incorporated into dosimetry guidelines to maximize tumor control and minimize treatment related morbidity. PMID- 10725645 TI - A Comparison of clinical target volumes determined by CT and MRI for the radiotherapy planning of base of skull meningiomas. AB - PURPOSE: To assess the utility of image registration and to compare the localization of clinical target volumes (CTV) using CT and MRI for patients with base of skull meningiomas undergoing radiotherapy. METHODS AND MATERIALS: Seven patients were imaged using CT and a T1-weighted MR volumetric sequence. Following image registration using a chamfer-matching algorithm, transaxial MR slices were reconstructed to match the planning CT slices. The accuracy of the image fusion was assessed in a preliminary study with matching accuracy better than 1.5 mm. The CTV in each patient was separately segmented by two independent observers for both CT and reconstructed MR image sets. Scalar and vector assessments were made of the difference in radial extent between the two outlines on each transaxial plane for all patients. A positive vector value corresponded to a greater extension of the tumor on MR compared to CT and vice versa. Scalar measurements compared the modulus of the differences between MR and CT, regardless of which volume was more extensive. Qualitative comparisons were also performed. RESULTS: Interobserver difference was small with a mean (+/- 1SD) volume difference of 1.5 +/- 1.5 cm(3) for CT and 0.5 +/- 1.0 cm(3) for MRI. The mean CT- and MR- CTVs were 17.6 +/-10.8 and 19.6 +/-14.2 cm(3) respectively. The mean overlap and composite volumes were 13.8 +/-10. 1 and 23.3 +/-14.8 cm(3) respectively. Average scalar differences in the left, right, anterior, and posterior directions were 6.0 +/- 7.0, 3.3 +/- 2.5, 4.9 +/- 3.9, and 4.5 +/- 5.0 mm respectively. The average vector differences were 3.3 +/- 8.5, -0.3 +/- 3.8, 1.1 +/- 5. 8, 1.5 +/- 6.4 mm (for left, right, anterior, and posterior directions respectively). Qualitatively, MR appeared to discern more tumor involvement in soft tissue regions adjacent to the skull base whereas CT appeared to provide larger target volumes within bony regions. CONCLUSIONS: MRI appeared to define CTVs that were larger but not inclusive of CT-defined CTVs. Although the average vector differences were small, the differences on individual borders could be large. In some instances, the CT or MR volumes were vastly different, each providing separate information. Therefore, the use of MRI and CT is complementary. Until accurate histological confirmation of disease extent is available, it is prudent to consider composite CT/MR volumes for the radiotherapy planning of base of skull meningiomas. PMID- 10725646 TI - A treatment planning method to correct dose distributions distorted by setup verification fields. AB - PURPOSE: Portal images of conformal treatment fields are often not suitable for setup verification purposes because they contain insufficient bony structures. Therefore, additional rectangular fields are frequently applied for setup verification purposes. It is the aim of this study to reduce the dose distortions induced by these extra fields by appropriately adjusting the beam weights and wedge angles of the treatment fields. METHODS AND MATERIALS: A second treatment plan for the setup verification session is generated, with an identical beam setup as the original plan, but which also includes two orthogonal setup verification fields. An algorithm has been developed, based on vector analysis methods, that adjusts the beam weights and wedge angles of the treatment fields in such a way that both the dose at the isocenter and the dose homogeneity over the planning target volume (PTV) are conserved. RESULTS: The algorithm has been applied to three clinical cases. The number of MUs for the setup verification fields, using a liquid-filled electronic portal imaging device, varied between 16 MU in the head and neck region up to 34 MU for lateral images in the pelvic region. In all cases, the method yielded a treatment plan including two orthogonal setup verification fields with a similar dose distribution over the PTV as the original treatment plan without the setup verification fields. CONCLUSION: The dose distortions resulting from the acquisition of orthogonal verification imaging can be neutralized by modifying the original beam weights and wedge angles of the treatment fields. PMID- 10725647 TI - A theoretical study of cylindrical ultrasound transducers for intracavitary hyperthermia. AB - PURPOSE: The purpose of this paper was to examine the heating patterns and penetration depth when a cylindrical ultrasound transducer is employed for intracavitary hyperthermia treatments. METHODS AND MATERIALS: The present study employs a simulation program based on a simplified power deposition model for infinitely long cylindrical ultrasound transducers. The ultrasound power in the tissue is assumed to be exponentially attenuated according to the penetration depth of the ultrasound beam, and a uniform attenuation for the entire treatment region is also assumed. The distribution of specific absorption rate (SAR) ratio (the ratio of SAR for a point within the tissue to that for a specific point on the cavity surface) is used to determine the heating pattern for a set of given parameters. The parameters considered are the ultrasound attenuation in the tissue, the cavity size, and the transducer eccentricity. RESULTS: Simulation results show that the ultrasound attenuation in the tissue, the cavity size, and the transducer eccentricity are the most influential parameters for the distribution of SAR ratio. A low frequency transducer located in a large cavity can produce a much better penetration. The cavity size is the major parameter affecting the penetration depth for a small cavity size, such as interstitial hyperthermia. The heating pattern can also be dramatically changed by the transducer eccentricity and radiating sector. In addition, for a finite length of cylindrical transducer, lower SAR ratio appears in the regions near the applicator's edges. CONCLUSION: The distribution of SAR ratio indicates the relationship between the treatable region and the parameters if an appropriate threshold of SAR ratio is taken. The findings of the present study comprehend whether or not a tumor is treatable, as well as select the optimal driving frequency, the appropriate cavity size, and the eccentricity of a cylindrical transducer for a specific treatment. PMID- 10725648 TI - Tradeoffs among delay, rate, and amount of reinforcement. AB - In Experiment 1, an adjusting-delay procedure was used to measure pigeons' choices between a single delayed reinforcer and a range of different variable time schedules. Indifference points showed an inverse relation between rate of reinforcement and delay that was well described by a hyperbolic equation. An adjusting-amount procedure was used in Experiment 2, in which pigeons chose between an adjusting amount of food delivered after a 0.5-s delay and 3 s of food delivered after a range of different delays, and the effects of delay were similar to those found in Experiment 1. The results from both experiments indicated that, for pigeons, the strength of a reinforcer decreased rapidly with increasing delay. Estimates of a decay rate parameter in the hyperbolic equation were similar to those found in other studies with pigeons, but the rates of temporal discounting were three or four times faster than those found in studies with rats, suggesting a possible species difference. PMID- 10725649 TI - Free-choice preference under uncertainty. AB - Preferences in pigeons for free choice over forced choice under uncertain contingencies were compared with the one under certain contingencies in multiple concurrent-chain schedules of reinforcement. The uncertain condition examined the preference for two alternatives over one alternative when reinforcement probability at the end of the terminal link equalled 0.5, and with all keys in each terminal link lit green (two mixed fixed-interval extinction keys versus one mixed fixed-interval extinction key). Key and schedule arrangement in the certain condition was the same except that a peck on any terminal-link key after the FI interval always produced food. When naive pigeons were first exposed to uncertain contingencies, preference for two lit keys over one lit key was observed, and the preference was confirmed by sequential reversals of the terminal-link contingencies. However, no consistent preference was observed when uncertain condition followed the certain condition. Under certain contingencies, unlike the earlier experiments, very small and inconsistent preferences for free choice were demonstrated. A possible reason for the different preferences in the uncertain conditions was that pigeons may lessen their sensitivity to the circumstances with uncertainty by any history or carry-over effect of the prior contingency. PMID- 10725650 TI - Breakfast in the nook and dinner in the dining room: time-of-day discrimination in rats. AB - Four studies were conducted which demonstrate that most (63%) male Sprague-Dawley rats can attain criterion, nine correct choices over ten consecutive trials, on a time-of-day discrimination in an elevated T-maze, but that the task is relatively difficult. The discrimination required that the rats go to one goal arm during a morning session and the other in an afternoon session. The sessions always began at the same time and were at least 6 h apart. A larger proportion of rats attained criterion and required fewer trials when the discriminative cue was a maze insert providing visual and tactile stimulation (0.72), orientation and position of the maze in the room (0.88), or the rats were required to always make the same left or right turn (0.94). Also, once criterion was attained, rats trained on time-of-day discrimination only made about 70% correct choices with continued training. Housing the rats with continuous light, all laboratory noises masked with white noise, and a random feeding schedule did not prevent them from acquiring the time-place discrimination. Testing the rats with a random number of trials during morning and afternoon sessions and with added or omitted sessions revealed that the rats did not use response or session alternation strategies to perform the discrimination. Also, the particular experimenter administering the morning or afternoon sessions did not serve as a cue for the discrimination. The relative difficulty of the task suggests that time of day does not normally function as a discriminative stimulus for choices, but probably as a contextual stimulus. Further, performance of the task in the absence of time-of-day cues suggests that the discrimination is based on event memory combined with an internal timing mechanism. PMID- 10725651 TI - Taste aversion induced by confinement in a running wheel. AB - Opportunities of spontaneous wheel running induced aversion in rats to the taste consumed before the running. Eight thirsty rats of Wistar strain were daily allowed to drink one of two taste solutions (sucrose or sodium chloride, say A) followed by confinement in a wheel or another taste solution (say B) followed by confinement in a Skinner box. Repeated training with Tastes A and B made the rats drink Taste A less than Taste B. Post-training two-bottle choice tests also showed clear demonstration of wheel-running-induced aversion to Taste A. These results confirmed those from Lett's laboratory and expanded the generality of the phenomenon in respect to strain of rats, deprivation conditions of the subjects, tastes, and other details. Furthermore, our procedure rejected three accounts alternative to wheel-induced aversion to Taste A. PMID- 10725652 TI - The effect of second-half reinforcer type on responding for sucrose in the first half of the session. AB - The present study investigated whether the sucrose-reinforced lever pressing of rats in the first half of a 50-min session would be sensitive to upcoming food pellet reinforcement in the second half. In Experiment 1, the type of reinforcer in the first half of the session was always liquid sucrose and type of reinforcer in the second half (liquid sucrose or food pellets) varied across conditions. Sucrose concentration varied across groups (1, 5, or 25%). Results showed that rates and patterns of responding for 1%, and sometimes for 5%, sucrose reinforcers in the first half of the session were higher and steeper, respectively, when food-pellet, rather than sucrose, reinforcement occurred in the second half. Responding for 25% sucrose was not similarly affected. Experiment 2 replicated the results of Experiment 1 using a within-subjects design. Although the present results represent induction (i.e. the opposite of contrast), they are consistent with some results on consummatory contrast. They also further demonstrate that responding on interval schedules of reinforcement can be altered prospectively. By doing so, however, they pose potential problems for current theories for why operant response rates change within the session. PMID- 10725653 TI - A biomechanical study of the human vertebral artery with implications for fatal arterial injury. AB - We studied the biomechanical behaviour of ring and strip specimens from along the length of 18 vertebral arteries taken from 16 subjects aged 28-90 years, in order to consider some of the factors which may play a role in vertebral artery rupture. The method was chosen to allow a comparison between circumferential distension (ring samples) and longitudinal extension, (strip samples). The samples were extended between the jaws of a tensile testing apparatus until the specimen broke and a number of biomechanical parameters were derived. These were the percentage extension to break, the tensile strength, Young's modulus and the peak load. There were a number of important findings. The vertebral artery was shown to be susceptible to longitudinal stretching with a number of strip samples breaking when extended by as little as 16-20%. The tensile strength and load at peak of the strip specimens were correspondingly lower than for the ring samples. Marked intersubject variations were shown for all these parameters and prominent changes in behaviour occurred along the vertebral artery. This study indicates that the artery may be susceptible to head and neck movements which cause the vessel to stretch, and intersubject variations in behaviour may be one important explanation for the marked differences in outcome which appear to exist in subjects who suffer broadly similar head and neck insults. PMID- 10725655 TI - Validation of twelve chemical spot tests for the detection of drugs of abuse. AB - Validation procedures are described for 12 chemical spot tests including cobalt thiocyanate, Dille-Koppanyi, Duquenois-Levine, Mandelin, Marquis, nitric acid, para-dimethylaminobenzaldehyde, ferric chloride, Froehde, Mecke, Zwikker and Simon's (nitroprusside). The validation procedures include specificity and limit of detection. Depending on the specificity of each color test, between 28 to 45 drugs or chemicals were tested in triplicate with each of the 12 chemical spot tests. For each chemical test, the final colors resulting from positive reactions with known amounts of analytes were compared to two reference color charts. For the identification of unknown drugs, reference colors from the Inter-Society Color Council and the National Bureau of Standards (ISCC-NBS) and Munsell charts are included along with a description of each final color. These chemical spot tests were found to be very sensitive with limits of detection typically 1 to 50 microg depending on the test and the analyte. PMID- 10725654 TI - A novel method for GHB detection in urine and its application in drug-facilitated sexual assaults. AB - A confirmation procedure for the identification and quantification of gamma hydroxybutyric acid (GHB) in urine is presented. This method is unique in that it does not involve the conversion of GHB to the gamma-butyrolactone (GBL). The urine samples were extracted using ethyl acetate, evaporated and derivatized with N, O-bis(trimethylsilyl)trifluoroacetamide (BSTFA) with 1% trimethylchlorosilane (TMCS), and analyzed by gas chromatography-mass spectrometry. Quantification was performed using selective ion monitoring (SIM), using GHB-d(6) as the internal standard. This method is simple and provides excellent linearity and sensitivity for GHB in urine. PMID- 10725656 TI - The shaking trauma in infants - kinetic chains. AB - The findings in three children who died as a consequence of shaking and those in another child who survived are presented. In the three fatal cases, a combination of anatomical lesions were identified at autopsy which appear to indicate the sites where kinetic energy related to the shaking episodes had been applied thus enabling the sequence of events resulting in the fatal head injury to be elucidated. Such patterns of injuries involved the upper limb, the shoulder, the brachial nerve plexus and the muscles close to the scapula; hemorrhages were present at the insertions of the sternocleidomastoid muscles due to hyperextension trauma (the so-called periosteal sign) and in the transition zone between the cervical and thoracic spine and extradural hematomas. Characteristic lesions due to traction were also found in the legs. All three children with lethal shaking trauma died from a subdural hematoma only a few hours after the event. The surviving child had persistant hypoxic damage of the brain following on massive cerebral edema. All the children showed a discrepancy between the lack of identifiable external lesions and severe internal ones. PMID- 10725657 TI - Simple and simultaneous analysis of fenfluramine, amphetamine and methamphetamine in whole blood by gas chromatography-mass spectrometry after headspace-solid phase microextraction and derivatization. AB - A simple and sensitive method for the simultaneous analysis of fenfluramine, amphetamine and methamphetamine in whole blood was developed using a headspace solid phase microextraction (SPME) and derivatization. A 0.5 g whole blood sample, 5 microl d(5)-methamphetamine (50 micrig/ml) as an internal standard, and 0.5 ml sodium hydroxide (1 M) were placed into a 12 ml vial, and sealed rapidly with a silicone septum and an aluminum cap. Immediately after the vial was heated to 70 degrees C in an aluminium block heater, the needle of the SPME device was inserted through the septum of the vial, and the extraction fiber was exposed in the headspace for 15 min. First, heptafluorobutyric anhydride was injected into the injection port of the GC-MS, and the compounds extracted by the fiber were then desorbed and derivatized simultaneously by exposing the fiber in the injection port. The calibration curves, using an internal standard method, demonstrated good linearity throughout the concentration range from 0.01 to 1.0 microg/g. The detection limits of this method were 5.0 ng/g for fenfluramine and methamphetamine, and 10 ng/g for amphetamine. No interferences were found, and the time for analysis was about 30 min for one sample. This method was applied to a suicide case in which the victim ingested fenfluramine. Fenfluramine was detected in the blood sample collected from the victim at the concentration of 7.7 microg/g. PMID- 10725658 TI - Computer-assisted facial image identification system using a 3-D physiognomic range finder. AB - This system consists of a 3-D physiognomic range finder and a computer-assisted facial image superimposition unit. The 3-D range finder is composed of a detector for measuring facial surface and its control computer. The detector has two sinusoidal grating projection devices and two CCD cameras. The computer-assisted facial image superimposition unit consists of a host computer including a proprietary software, a flat surface color display and a color image scanner for inputting 2-D facial images of a criminal. The 3-D facial shape and texture of a suspect is obtained by using the range finder. To make the comparison between the 3-D facial image and the 2-D facial image, the 3-D facial image is first reproduced on a display of the host computer from a MO disk and then the 2-D facial image is taken with the color image scanner and reproduced on the display. The 3-D facial image is exactly adjusted to match the orientation and size of the 2-D facial image under the fine framework mode, and then the fine framework mode of 3-D facial image is converted to the fine texture image. The shape and positional relationships of facial components between the 3-D and 2-D facial images are examined by the fade-out or wipe image mode. The distance between the selected two points and angle among the selected three points on the 3-D and 2-D facial images are automatically measured for the assessment of anthropometrical data between both images. For evaluating the fit between the anthropometrical points on the 3-D and 2-D facial images, the reciprocal point-to-point difference between both images is compared. PMID- 10725659 TI - XRCC3 is required for efficient repair of chromosome breaks by homologous recombination. AB - XRCC3 was originally identified as a human gene able to complement the DNA damage sensitivity, chromosomal instability and impaired growth of the mutant hamster cell line irs1SF. More recently, it has been cloned, sequenced and found to bear sequence homology to the highly conserved eukaryotic repair and recombination gene RAD51. The phenotype of irs1SF and the identification of XRCC3 as a member of the RAD51 gene family have suggested a role for XRCC3 in repair of DNA damage by homologous recombination. Homologous recombinational repair (HRR) of a specifically induced chromosomal double-strand break (DSB) was assayed in irs1SF cells with and without transient complementation by human XRCC3. Complementation with XRCC3 increased the frequencies of repair by 34- to 260-fold. The results confirm a role for XRCC3 in HRR of DNA DSB, and the importance of this repair pathway for the maintenance of chromosomal integrity in mammalian cells. PMID- 10725660 TI - Defective nucleotide excision repair in xpc mutant mice and its association with cancer predisposition. AB - Mice that are genetically engineered are becoming increasingly more powerful tools for understanding the molecular pathology of many human hereditary diseases, especially those that confer an increased predisposition to cancer. We have generated mouse strains defective in the Xpc gene, which is required for nucleotide excision repair (NER) of DNA. Homozygous mutant mice are highly prone to skin cancer following exposure to UVB radiation, and to liver and lung cancer following exposure to the chemical carcinogen acetylaminofluorene (AAF). Skin cancer predisposition is significantly augmented when mice are additionally defective in Trp53 (p53) gene function. We also present the results of studies with mice that are heterozygous mutant in the Apex (Hap1, Ref-1) gene required for base excision repair and with mice that are defective in the mismatch repair gene Msh2. Double and triple mutant mice mutated in multiple DNA repair genes have revealed several interesting overlapping roles of DNA repair pathways in the prevention of mutation and cancer. PMID- 10725661 TI - Deoxyxanthosine in DNA is repaired by Escherichia coli endonuclease V. AB - Deoxycytidine, deoxyadenosine and deoxyguanosine undergo spontaneous deamination to form deoxyuridine, deoxyinosine and deoxyxanthosine, respectively. In this manuscript, we show that in addition to its known ability to recognize deoxyuridine and deoxyinosine in DNA, Escherichia coli endonuclease V cleaves DNA containing deoxyxanthosine. However, Alk A protein and human methylpurine glycosylase are unable to recognize deoxyxanthosine. Endonuclease V cleaves DNA containing deoxyxanthosine at the second phosphodiester bond 3' to deoxyxanthosine, generating a 3'-hydroxyl and a 5'-phosphoryl group at the nick site. This endonucleolytic activity requires Mg(2+) or Mn(2+), and is highly specific for double stranded DNA. Endonuclease V-catalyzed cleavage of DNA containing deoxyxanthosine is a result of its ability to recognize the altered base and not due to its mismatch-specific endonuclease activity. The ability of endonuclease V to recognize both deoxyinosine and deoxyxanthosine suggests that endonuclease V is important for preventing mutations that might arise as a result of deamination of purines. PMID- 10725662 TI - Effects of deoxyribonucleosides and cell-stimulation on frequencies of ultraviolet-B-induced micronuclei in human peripheral blood lymphocytes. AB - In the present study the involvement of deoxyribonucleotides (dNTPs) in the clastogenicity of ultraviolet-B (UVB) in unstimulated peripheral blood lymphocytes (G(0)-PBLs) was investigated. This was studied by analyzing the frequency of UVB-induced micronuclei (MN), either after adding a cocktail of the four deoxyribonucleosides to the PBLs immediately after exposure to UVB, or by stimulating the cells before exposure. In total, PBLs obtained from two different donors were investigated. For both donors, it could be demonstrated that addition of deoxyribonucleosides to UVB-irradiated G(0)-PBLs resulted in a significant reduction of the clastogenic effect of UVB. A gradual reduction of the clastogenic effect of UVB could also be realized by irradiating PBLs that were progressively more stimulated with the lectin PHA before exposure. The latter finding is explained by upregulation of intracellular pool sizes of dNTPs in stimulated PBLs. PMID- 10725663 TI - Werner's syndrome lymphoblastoid cells are hypersensitive to topoisomerase II inhibitors in the G2 phase of the cell cycle. AB - Werner's syndrome (WS) is a rare autosomal recessive human disorder and the patients exhibit many symptoms of accelerated ageing in their early adulthood. The gene (WRN) responsible for WS has been biochemically characterised as a 3'-5' helicase and is homologous to a number of RecQ superfamily of helicases. The yeast SGS1 helicase is considered as a human WRN homologue and SGS1 physically interacts with topoisomerases II and III. In view of this, it has been hypothesised that the WRN gene may also interact with topoisomerases II and III. The purpose of this study is to determine whether the loss of function of WRN protein alters the sensitivity of WS cells to agents that block the action of topoisomerase II. This study deals with the comparison of the chromosomal damage induced by the two anti-topoisomerase II drugs, VP-16 and amsacrine, in both G1 and G2 phases of the cell cycle, in lymphoblastoid cells from WS patients and from a healthy donor. Our results show that the WS cell lines are hypersensitive to chromosome damage induced by VP-16 and amsacrine only in the G2 phase of the cell cycle. No difference either in the yield of the induced aberrations or SCEs was found after treatment of cells at G1 stage. These data might suggest that in WS cells, because of the mutation of the WRN protein, the inhibition of topoisomerase II activity results in a higher rate of misrepair, probably due to some compromised G2 phase processes involving the WRN protein. PMID- 10725664 TI - p53-degradation by HPV-16 E6 preferentially affects the removal of cyclobutane pyrimidine dimers from non-transcribed strand and sensitizes mammary epithelial cells to UV-irradiation. AB - Nucleotide excision repair (NER), the most versatile and ubiquitous mechanism for DNA repair, operates to remove many types of DNA base lesions. We have studied the role of p53 function in modulating the repair of DNA damage following UV irradiation in normal and p53-compromised human mammary epithelial cells (HMEC). The effect of UV-induced DNA damage on cellular cytotoxicity and apoptosis was determined in conjunction with global, gene- and strand-specific repair. Cytotoxicity studies, using clonogenic survival and MTT assays, showed that HPV 16 E6-expressing HMEC were more UV sensitive than p53-WT cell lines. High apoptotic index obtained with p53-compromised cells was in conformity to both the low clonogenic survival and the low cellular viability. No discernible differences in the formation of initial UV-induced cyclobutane pyrimidine dimers (CPD) were observed in the cell lines of varying p53 functional status. However, the extent and the rate of damage removal from genome overall were highest for p53-WT cells. Further examination of strand-specific repair in the p53 gene revealed that the removal of CPD in the non-transcribed strand (NTS) was slower in p53-compromised cells compared to the normal p53-WT cell lines. These results suggest that loss of p53 function, in the absence of other genetic alterations, decreased both overall amount of CPD repaired and their removal rate from the genome. Additionally, normal function of p53 is required for the repair of the NTS, but not of the transcribed strand (TS) in genomic DNA in human epithelial cells. Thus, failure of quantitative removal of CPD by global genomic repair (GGR), due to loss of p53 function, causes the enhanced UV sensitivity and increased damage-induced apoptosis via a p53-independent pathway. Nevertheless, recovery of cells from UV damage requires normal p53 function and efficient GGR. PMID- 10725665 TI - Reconstitution of damage DNA excision reaction from SV40 minichromosomes with purified nucleotide excision repair proteins. AB - We previously constructed the cell-free nucleotide excision repair (NER) assay system with UV-irradiated SV40 minichromosomes to analyze the mechanism of NER reaction on chromatin DNA. Here we investigate the factor that acts especially on nucleosomal DNA during the damage excision reaction, and reconstitute the damage excision reaction on SV40 minichromosomes. NER-proficient HeLa whole cell extracts were fractionated, and the amounts of known NER factors involved in the column fractions were determined by immunoblot analyses. The column fractions were quantitatively and systematically replaced by highly purified NER factors. Finally, damage DNA excision reaction on SV40 minichromosomes was reconstituted with six highly purified NER factors, XPA, XPC-HR23B, XPF-ERCC1, XPG, RPA and TFIIH, as those essential for the reaction with naked DNA. Further analysis showed that the damages on chromosomal DNA were excised as the same efficiency as those on naked DNA for short incubation. At longer incubation time, however, the damage excision efficiency on nucleosomal DNA was decreased whereas naked DNA was still vigorously repaired. These observations suggest that although the six purified NER factors have a potential to eliminate the damage DNA from SV40 minichromosomes, the chromatin structure may still have some repressive effects on NER. PMID- 10725671 TI - Bacterial meningitis. AB - With nearly 8,000 cases in the United States per year, and 2,000 deaths annually, bacterial meningitis continues to be a significant source of morbidity and mortality. The principal pathogens are Neisseria meningitidis, Streptococcus pneumoniae, group B streptococci, and Hemophilus influenzae. In immunocompromised patients, Listeria monocytogenes is also an important pathogen. Rapid identification and evaluation of the patient with bacterial meningitis and prompt initiation of antibiotics are the cornerstones of therapy. Except in the rare patient with papilledema, focal neurologic symptoms, or a seizure, a lumbar puncture should be performed without delay, and antibiotic therapy should be administered promptly. Patients without a readily identifiable source of infection should be treated empirically with intravenous ceftriaxone. Ampicillin should also be administered in populations at increased risk for L. monocytogenes. The risk of meningitis in some populations can be reduced by administration of vaccines against selected pathogens such as N. meningitidis, S. pneumoniae, and H. influenzae. PMID- 10725666 TI - 5-chloro-2'-deoxyuridine cytotoxicity results from base excision repair of uracil subsequent to thymidylate synthase inhibition. AB - The lack of a phenotypic alteration of 5-hydroxymethyluracil (hmUra) DNA glycosylase (hmUDG) deficient Chinese hamster V79mut1 cells exposed to DNA damaging agents known to produce hmUra has raised the question whether there might be DNA substrates other than hmUra for hmUDG. Based on the structural similarity between 5-chlorouracil (ClUra) and hmUra and the observations that 5 chloro-2'-deoxyuridine (CldUrd) induces base excision repair (BER) events, we asked whether hmUDG or some other DNA BER enzyme is responsible for the removal of ClUra from DNA. An in vivo flow cytometry assay with FITC-anti-BrdUrd (which cross-reacts with CldUrd) showed that exogenous CldUrd is incorporated into DNA. However, both in vivo and in vitro experiments indicated that ClUra is not excised from DNA by hmUDG or other DNA glycosylase activities. The absence of removal of ClUra by hmUDG raised the question whether DNA strand breaks occurred subsequent to thymidylate synthase inhibition, leading to deoxyuridine incorporation, followed by cleavage of uracil from DNA by uracil DNA glycosylase (UDG). An in vivo thymidylate synthase activity assay in V79 cells demonstrated that CldUrd treatment inhibits thymidylate synthase as effectively as 5-fluoro-2' deoxyuridine (FdUrd) treatment. Uracil, a known UDG inhibitor, partially reverses the cytotoxic effects of CldUrd on V79 cells, thus confirming that CldUrd induced cytotoxicity is a result of UDG activity. Our results demonstrated that while CldUrd is not directly repaired from DNA, its cytotoxicity is directly due to the UDG removing uracil subsequent to inhibition of thymidylate synthase by CldUMP. PMID- 10725672 TI - The prognostic value and clinical utility of estrogen and progesterone receptors in endometrial carcinoma. AB - In the United States, endometrial carcinoma is the most common gynecologic malignancy, and accounts for 4,900 deaths per year in the United States. While this disease has relatively good cure rates, there is motivation to describe other determinants, which may help in the treatment of this disease. Attempts have been made to correlate hormone receptor status with disease-free intervals and survival in patients with endometrial carcinoma. The weight of evidence seems to support that of the two hormone receptors, progesterone is the more significant predictor of patient outcome. If hormone receptors are to be used in the management of endometrial carcinoma, they should be determined by immunohistochemistry. In the adjuvant setting, patients with progesterone positive tumors are more amenable to treatment with progestational agents than are patients with receptor negative tumors. Future areas of research include the use of tamoxifen and selective estrogen receptor modulators in the chemoprevention and treatment of endometrial carcinoma. PMID- 10725673 TI - The importance of obstetricians/ gynecologists in promoting children's unintentional injury prevention: a focus on bicycle helmet use. AB - Injuries are the leading cause of death for children in the United States, as well as for women between the ages of 19 and 39. The leading cause of injury death for these groups is motor vehicle injuries. There are approximately 900 deaths every year related to bicycling, with 40% of these occuring to children between the ages of 5-14. Most of these deaths occur due to collisions with motor vehicles. It has been shown that the use of bicycle helmets prevents the majority of head and brain injuries that occur due to bicycle crashes. While it is true that the pediatrician is an important conduit for information about children's injury prevention, there is also a role for ob/gyns. Many women who visit ob/gyns have children, are expecting children, and/or plan to have children in the near future. In addition, adult women are at risk for many of these same injuries. For these reasons, the ob/gyn and his/her office setting can serve as important vehicles for education. An effective way to get started may be to counsel on bike injuries and deaths and the importance of wearing a bicycle helmet. There are several educational sources/materials and helmets that can be obtained free of charge. Successful injury prevention efforts have involved the participation of a wide cadre of professionals, including members of the medical profession. It is important for ob/gyns to consider themselves and their office staffs effective partners in children's injury prevention efforts. PMID- 10725674 TI - Risk factors and interventions for psychological sequelae in women after miscarriage. AB - Spontaneous abortion occurs in about one in five pregnancies, and for many women can lead to significant psychological sequelae. This article reviews the literature on the psychological morbidities after pregnancy loss, their risk factors, differences in partner's reactions, potential screening tools and interventions for physicians. The most common reactions women experienced after pregnancy loss were grief, depression, and anxiety, with rates cited of approximately 40%, 12-50%, and 22-41%, respectively. Risk factors for these reactions include having a past psychiatric history, poor social support, no living children, lack of knowledge about miscarriages, and having no explanation for the event. Women's partners also show grief reactions after pregnancy loss, but they grieve less intensely and for a briefer time than women. Men also tend to talk less about their feelings. Women consistently express a desire to have follow-up appointments after a miscarriage, and have a high (75%) show rate when physicians offer them follow-up appointments. The 30-item General Health Questionnaire has good sensitivity and specificity in detecting psychological morbidity after miscarriage. Based on our review we recommend that ob/gyns offer women a follow-up appointment two to three weeks after miscarriage. By using a screening tool such as the General Health Questionnaire and asking about potential risk factors, ob/gyns should be able to effectively identify and refer high risk women to a psychiatric consultant. PMID- 10725675 TI - Inherited coagulopathies in OB/GYN. AB - There is an increased risk of venous thrombosis in women with inherited plasma protein coagulopathies (thrombophilias). Although a general screening for thrombophilia is impractical, risk assessment based on identification of other hypercoagulable states-such as a personal or family history of venous thrombosis can identify patients who may benefit from testing. Those patients testing positive for thrombophilia need to be counseled regarding the implications of oral contraceptive use, hormone replacement therapy, use of selective estrogen receptor modulators, and pregnancy. PMID- 10725676 TI - University hospital based care decreases recurrent preterm deliveries and costs in comparison to no prenatal care(1). AB - Objective: To compare perinatal outcomes and total health care costs for patients with a previous history of preterm delivery cared for by an inner city university hospital house staff clinic versus patients who have not received any prenatal care.Study Design: We conducted a retrospective review of women with a history of preterm delivery between 1994 and 1996. The inclusion criteria was a history of at least one preterm delivery. Data were obtained on maternal demographics, complete past obstetrical history, and number of prenatal visits.Results: The study groups consisted of 96 house staff patients and 53 patients without prenatal care. The number of prior preterm births, incidence of recurrent preterm delivery, length of neonatal intensive care unit stay, and mother-infant costs were greater in the group without prenatal care. The mean gestational age at time of delivery, and mean birth weight were greater in the house staff group.Conclusion: Our study demonstrates that university hospital-based prenatal care decreases recurrent preterm delivery rates and health care costs when compared to lack of prenatal care. Prenatal care provided at academic centers, with a coordinated multidisciplinary approach, appears to have a positive impact on the problem of preterm deliveries. PMID- 10725677 TI - Women and smoking: patterns, health effects, and treatments. AB - Tobacco use remains the number one cause of preventable death among women in the United States. Of particular concern to obstetricians and gynecologists is the morbidity caused by tobacco use and by exposure to secondhand smoke among women of childbearing age, pregnant women, and their newborns. Women who smoke have lower fertility rates and are more likely to experience negative side effects from oral contraceptives. Pregnant women who smoke are more likely to suffer miscarriages and to have low birth weight and preterm babies, and infants with sudden infant death syndrome. Over one of every four women aged 18 to 44 smoke in the U.S. Unfortunately, at least 14% of women smoke during pregnancy. The good news is that research shows that tobacco treatment interventions by health care providers can increase the number of patients who successfully quit. The Agency for Health Care Policy and Research developed recommendations and guidelines to assist health care providers to integrate a tobacco treatment intervention into an office setting. It is recommended that every physician 1) ask patients about their tobacco use; 2) advise them to stop using tobacco; 3) assist patients interested in cessation efforts; and 4) arrange appropriate follow-up. PMID- 10725690 TI - Gametocytocidal activity of pyronaridine and DNA topoisomerase II inhibitors against multidrug-resistant Plasmodium falciparum in vitro. AB - Gametocytocidal activities of pyronaridine and DNA topoisomerase II inhibitors against two isolates of multidrug-resistant Plasmodium falciparum, KT1 and KT3 were determined. After sorbitol treatment, pure gametocyte cultures of Plasmodium falciparum containing mostly young gametocytes (stage II and III) obtained on day 11 were exposed to the drugs for 48 h. The effect of the drugs on gametocyte development was assessed by counting gametocytes on day 15 of culture. Pyronaridine was the most effective gametocytocidal drug against P. falciparum isolates KT1 and KT3 with 50% inhibitory concentration of 6 and 20 nM, respectively. Moreover, the 50% inhibitory concentration of pyronaridine was lower than that of primaquine which is the only drug used to treat malaria patients harboring gametocytes. Prokaryotic (norfloxacin) and eukaryotic (amsacrine and etoposide) DNA topoisomerase II inhibitors were only effective against asexual but not sexual stages of the malaria parasites. Pyronaridine has both schizontocidal and gametocytocidal activities against the human malaria parasite, P. falciparum. PMID- 10725691 TI - A 43-kDa circulating filarial antigen fraction of Wuchereria bancrofti in immunoprophylaxis against Brugia malayi in jirds. AB - A 43 kiloDaltan (kDa) antigen fraction (CFA2-6) isolated from microfilaraemic plasma of bancroftian filarial patients showed selective reactivity with sera samples collected from endemic normals. Antibodies raised against this antigen showed a strong reactivity with the surface of Brugia malayi infective larvae as well as microfilariae. Similar antigenic determinants were detected in the parasite extracts, but not in the excretory-secretory products. Further analysis was done on the immunoprophylactic potential of CFA2-6 in inducing immunity against Brugia malayi in Meriones unguiculatus (jird) in vivo. A strong protective response of approximately 84% was observed against the development of the filarial parasite in the jirds immunized with CFA2-6. The immunized jirds also showed a significant clearance (87%) of microfilariae inoculated intravenously. Approximately 65% of infective larvae failed to survive in jirds transferred with anti-CFA2-6 serum compared to the jirds transferred with sera from the control jirds. Passive transfer of anti-CFA2-6 antibody to the jirds followed by intravenous inoculation of microfilariae resulted in the reduction of 77% of circulating microfilariae. This study suggests that the 43-kDa CFA2-6 could stimulate a strong protective immune response against infective larvae and microfilariae in experimental animals. PMID- 10725693 TI - Increased production of acute-phase proteins corresponds to the peak parasitaemia of primary malaria infection. AB - Recent studies have implicated non-specific mediators associated with CD4+ T cells of the T helper 1 subset in resistance to experimental malarias. As part of continuing studies into the multifactorial role of nitric oxide and other contributors to the innate immune response in control of acute-phase malaria infection, the production of the acute-phase proteins, caeruloplasmin and serum amyloid P, following infection of naive mice with blood stages of the rodent malaria parasite Plasmodium chabaudi was investigated. Levels of both acute-phase proteins in the serum of infected mice were significantly elevated on days 7-12 post-infection compared both to other times of infection, and to background levels detected in uninfected control mice. These times corresponded to the ascending and peak primary parasitaemia, when production of interferon-gamma, tumour necrosis factor-alpha and nitric oxide is known to be raised. Although it is not apparent whether the production of caeruloplasmin and serum amyloid P has a causal effect in reducing parasitaemia or is simply a by-product of innate immunity, the detection of increased levels of circulating acute-phase proteins may act as a useful surrogate marker of high level parasitaemia, and therefore, of blood-borne malaria pathology. PMID- 10725692 TI - The genetic analysis of F1 hybrid larvae between female Trichinella spiralis and male Trichinella britovi. AB - Hybrids between female Trichinella spiralis and male Trichinella britovi were constructed. Then, hybrid genotype was characterized by DNA markers including mitochondrial cytochrome c oxidase subunit I (CO I) gene, the gene encoding the 43-kDa excretory-secretory (ES) protein, and genomic DNA fragments specific for T. spiralis and T. britovi identified from random amplified polymorphism DNA (RAPD). PCR-restriction fragment length polymorphism (PCR-RFLP) analysis of the mitochondrial CO I gene revealed that all hybrids carried a T. spiralis pattern. The same analysis of the gene encoding the 43-kDa ES protein showed that each hybrid carried both T. spiralis and T. britovi gene type simultaneously. In the analysis of genomic DNA using RAPD-derived PCR primers, some hybrids carried T. spiralis and T. britovi-specific RAPD markers, while others carried the RAPD marker of T. spiralis only. PMID- 10725694 TI - Two Taenia species found in Japan, with new distribution record of Taenia polyacantha Leuckart, 1856 (Cestoda: Taeniidae). AB - In an epidemiological survey for Echinococcus multilocularis in rodents and insectivores from the northernmost part of the central mainland of Japan (Honshu), two taeniid species, Taenia crassiceps and Taenia polyacantha, were found in Microtus montebelli and Apodemus argenteus, respectively. The latter is the first record of distribution in Japan, and the former is the second after its first recovery from the central part of Japan. Although we have found neither larval nor strobilar stage of E. multilocularis there, discovery of these taeniid species, having overlapping global distribution with E. multilocularis in red foxes Vulpes vulpes as well as multiple occurrences of hydatid patients having no history of visits to the endemic areas shows the possibility that the life-cycle of E. multilocularis might be maintained at least in the northernmost part of Honshu. PMID- 10725695 TI - Cutting edge: recombinase-activating gene expression and V(D)J recombination in CD4+CD3low mature T lymphocytes. AB - The recombinase-activating genes, RAG-1 and RAG-2, can be expressed by a subset of B cells within germinal centers, where they mediate secondary V(D)J rearrangements. This receptor revision mechanism could serve either receptor diversification or tolerance-induced functions. Alternatively, it might rescue those cells the receptors of which have been damaged by somatic mutation. Less is known about the occurrence of similar mechanisms in T cells. Here we show that mature T cells with defective TCR surface expression can express RAG genes and are capable of initiating secondary V(D)J rearrangements. The possibility that a cell rescue mechanism based on the generation of a novel Ag receptor might be active in peripheral T cells is envisaged. PMID- 10725696 TI - Cutting edge: identification of the orphan receptor G-protein-coupled receptor 2 as CCR10, a specific receptor for the chemokine ESkine. AB - A number of orphan G-protein coupled receptors (GPR) have been reported as putative chemokine receptors. One previously reported orphan receptor is an incomplete PCR clone, called GPR2. Here we report the cloning of full-length human (h)GPR2 and mouse (m)GPR2 cDNAs, and the identification of GPR2 as a receptor for a novel CC chemokine called ESkine. hGPR2 is expressed at high levels in testis and small intestine, and at lower levels in other tissues. mGPR2 was expressed at high levels in small intestine, colon, lymph nodes, and Peyer's patches and at lower levels in thymus and spleen. Stimulation of L1.2/hGPR2 transfectants with hESkine induced their migration and resulted in intracellular calcium mobilization. These results provide evidence that GPR2 is a specific receptor for ESkine. We propose that GPR2 be renamed as CCR10. The expression pattern of mGPR2/CCR10 suggests that it may play a role in the homing/trafficking of leukocytes within intestinal and lymphoid environments. PMID- 10725697 TI - Cutting edge: the orphan chemokine receptor G protein-coupled receptor-2 (GPR-2, CCR10) binds the skin-associated chemokine CCL27 (CTACK/ALP/ILC). AB - We recently reported the identification of a chemokine (CTACK), which has been renamed CCL27 according to a new systematic chemokine nomenclature. We report that CCL27 binds the previously orphan chemokine receptor GPR-2, as detected by calcium flux and chemotactic responses of GPR-2 transfectants. We renamed this receptor CCR10. Because of the skin-associated expression pattern of CCL27, we focused on the expression of CCL27 and CCR10 in normal skin compared with inflammatory and autoimmune skin diseases. CCL27 is constitutively produced by keratinocytes but can also be induced upon stimulation with TNF-alpha and IL 1beta. CCR10 is not expressed by keratinocytes and is instead expressed by melanocytes, dermal fibroblasts, and dermal microvascular endothelial cells. CCR10 was also detected in T cells as well as in skin-derived Langerhans cells. Taken together, these observations suggest a role for this novel ligand/receptor pair in both skin homeostasis as well as a potential role in inflammatory responses. PMID- 10725698 TI - Cutting edge: cell surface expression and lipopolysaccharide signaling via the toll-like receptor 4-MD-2 complex on mouse peritoneal macrophages. AB - The human MD-2 molecule is associated with the extracellular domain of human Toll like receptor 4 (TLR4) and greatly enhances its LPS signaling. The human TLR4-MD 2 complex thus signals the presence of LPS. Little is known, however, about cell surface expression and LPS signaling of the TLR4-MD-2 complex in vivo. We cloned mouse MD-2 molecularly and established a unique mAb MTS510, which reacted selectively with mouse TLR4-MD-2 but not with TLR4 alone in flow cytometry. Mouse MD-2 expression in TLR4-expressing cells enhanced LPS-induced NF-kappaB activation, which was clearly inhibited by MTS510. Thioglycolate-elicited peritoneal macrophages expressed TLR4-MD-2, which was rapidly down-regulated in the presence of LPS. Moreover, LPS-induced TNF-alpha production by peritoneal macrophages was inhibited by MTS510. Collectively, the TLR4-MD-2 complex is expressed on macrophages in vivo and senses and signals the presence of LPS. PMID- 10725699 TI - Cutting edge: endotoxin tolerance in mouse peritoneal macrophages correlates with down-regulation of surface toll-like receptor 4 expression. AB - Monocytes/macrophages exposed to LPS show reduced responses to second stimulation with LPS, which is termed LPS tolerance. In this study, we investigated molecular mechanism of LPS tolerance in macrophages. Mouse peritoneal macrophages pre exposed to LPS exhibited reduced production of inflammatory cytokines in a time- and dose-dependent manner. Activation of neither IL-1 receptor-associated kinase nor NF-kappaB was observed in macrophages that became tolerant by LPS pretreatment, indicating that the proximal event in Toll-like receptor 4 (TLR4) MyD88-dependent signaling is affected in tolerant macrophages. Although TLR4 mRNA expression significantly decreased within a few hours of LPS pretreatment and returned to the original level at 24 h, the surface TLR4 expression began to decrease within 1 h, with a gradual decrease after that, and remained suppressed over 24 h. A decrease in inflammatory cytokine production in tolerant macrophages well correlates with down-regulation of the surface TLR4 expression, which may explain one of the mechanisms for LPS tolerance. PMID- 10725700 TI - Cutting edge: human B cell function is regulated by interaction with soluble CD14: opposite effects on IgG1 and IgE production. AB - The mechanism(s) controlling activation of naive B cells, their proliferation, Ag receptor affinity maturation, isotype switching, and their fate as memory or plasma cells is not fully elucidated. Here we show that between 24 and 60% of CD19+ cells in PBMC bind soluble CD14 (sCD14). Tonsillar B cells also bind sCD14, but preferentially the CD38-ve/low cells. Interaction of sCD14 with B cells resulted in higher levels of IgG1 and marked inhibition of IgE production by activated tonsillar B cells and Ag-stimulated PBMC. We found that sCD14 interfered with CD40 signaling in B cells, inhibited IL-6 production by activated B cells, and increased the kinetics and magnitude of CD40 ligand expression on T cells. Together with the previously reported effects on T cells, these findings define sCD14 as a novel soluble regulatory factor capable of modulating cellular and humoral immune responses by interacting directly with T and B cells. PMID- 10725701 TI - IL-4 diminishes perforin-mediated and increases Fas ligand-mediated cytotoxicity In vivo. AB - CTL have evolved two major mechanisms for target cell killing: one mediated by perforin/granzyme secretion and the other by Fas/Fas ligand (L) interaction. Although cytokines are integral to the development of naive CTL into cytolytic effectors, the role of cytokines on mechanisms of CTL killing is just emerging. In this study, we evaluate the effects of IL-4 in Fas(CD95)/FasL(CD95L)-mediated killing of Fas-overexpressing target cells. Recombinant vaccinia viruses (vv) were constructed to express respiratory syncytial virus M2 Ag alone (vvM2) or coexpress M2 and IL-4 (vvM2/IL-4). MHC-matched Fas-overexpressing target cells (L1210Fas+) were used to measure both perforin- and FasL-mediated killing pathways. In contrast to Fas-deficient (L1210Fas-) target cells, effectors from vvM2/IL-4-immunized mice were able to lyse L1210Fas+ target cells with similar magnitude as vvM2-infected mice. Addition of EGTA/Mg2+ revealed that effectors from vvM2/IL-4-infected mice primarily lyse targets by a Ca2+-independent Fas/FasL pathway. Analysis of FasL expression by flow cytometry showed that IL-4 increased cell surface FasL expression on CD4+ and CD8+ splenocytes, with peak expression on day 4 after infection. These data demonstrate that IL-4 increases FasL expression on T cells, resulting in a shift of the mechanism of CTL killing from a dominant perforin-mediated cytolytic pathway to a dominant FasL-mediated cytolytic pathway. PMID- 10725702 TI - Calcitonin gene-related peptide decreases expression of HLA-DR and CD86 by human dendritic cells and dampens dendritic cell-driven T cell-proliferative responses via the type I calcitonin gene-related peptide receptor. AB - These studies were performed to establish whether functional receptors for calcitonin gene-related peptide (CGRP) are present on human dendritic cells (DCs) and to investigate potential immunomodulatory effects of CGRP on DCs other than Langerhans cells. Reverse transcriptase-PCR revealed expression of mRNA for a type 1 CGRP receptor by mature and immature blood-derived DCs. Sequence analysis confirmed the identity of the type 1 CGRP receptor (CGRP-R1). Addition of CGRP (10-7 M) to mature and immature DCs resulted in mobilization of intracellular calcium. Treatment of immature DCs with CGRP (10-7 M), before and after maturation in monocyte-conditioned medium, resulted in decreased cell surface expression of HLA-DR MHC class II and the costimulatory molecule, CD86. Treatment of immature DCs with CGRP (10-7 M) also resulted in decreased expression of CD86, but expression of HLA-DR was unchanged. When CGRP-treated mature DCs were used to stimulate allogeneic T cells, proliferative responses were dampened (approximately 50%), especially at low DC:T cell ratios (1:360). This effect was not observed with CGRP-treated, immature DCs. In contrast, CGRP-treated mature or immature DCs were no less efficient than untreated DCs in driving syngeneic T cell-proliferative responses to staphylococcal enterotoxin B. We conclude that mature and immature DCs express type 1 CGRP receptors and that signaling through these receptors may dampen mature DC-driven T cell proliferation most likely via down-regulation of CD86 and HLA-DR. PMID- 10725703 TI - IL-2 and IL-15 regulate CD154 expression on activated CD4 T cells. AB - The cellular and humoral immune system is critically dependent upon CD40-CD154 (CD40 ligand) interactions between CD40 expressed on B cells, macrophages, and dendritic cells, and CD154 expressed primarily on CD4 T cells. Previous studies have shown that CD154 is transiently expressed on CD4 T cells after T cell receptor engagement in vitro. However, we found that stimulation of PBLs with maximal CD28 costimulation, using beads coupled to Abs against CD3 and CD28, led to a very prolonged expression of CD154 on CD4 cells (>4 days) that was dependent upon autocrine IL-2 production. Previously activated CD4 T cells could respond to IL-2, or the related cytokine IL-15, by de novo CD154 production and expression without requiring an additional signal from CD3 and CD28. These results provide evidence that CD28 costimulation of CD4 T cells, through autocrine IL-2 production, maintains high levels of CD154 expression. This has significant impact on our understanding of the acquired immune response and may provide insight concerning the mechanisms underlying several immunological diseases. PMID- 10725704 TI - Ligand-independent down-regulation of IFN-gamma receptor 1 following TCR engagement. AB - Activated T lymphocytes modulate the level of many molecules on their cell surface, including cytokine receptors. This regulation of cytokine receptor expression affects the ability of T cells to respond to cytokines and thus influences the outcome of an immune response. The receptor for IFN-gamma, a proinflammatory cytokine, consists of two copies of a ligand binding chain (IFN gammaR1) as well as two copies of a second chain (IFN-gammaR2) required for signal transduction. The expression of IFN-gammaR2 is down-regulated at the mRNA level on CD4+ T cells when they differentiate into the Th1, but not the Th2, phenotype. This down-regulation has been demonstrated to depend on the ligand, IFN-gamma, which is produced by Th1 but not Th2 T cells. The regulation of the cell-surface expression of IFN-gamma receptors during primary T cell activation has not been reported. Naive and differentiated T lymphocytes express IFN-gammaR1 at the mRNA level and as a cell-surface protein. In this study, we present evidence that cell-surface expression of IFN-gammaR1 is transiently down regulated on the surface of naive CD4+ T cells shortly after TCR engagement. Furthermore, this down-regulation is not mediated by the ligand, IFN-gamma, but results from TCR engagement and can be inhibited by cyclosporin A. PMID- 10725705 TI - The IFN regulatory factor family participates in regulation of Fas ligand gene expression in T cells. AB - TCR engagement leads to the transcriptional activation of cytokine genes and activation-induced cell death. Activated T cells undergo apoptosis upon expression and ligation of Fas ligand (FasL) to Fas/APO-1 (CD95) receptor. FasL expression is under the transcriptional regulation of multiple factors. The present study demonstrates that TCR-inducible FasL expression is also under the direct influence of the IFN regulatory factor (IRF) transcription factor family. Deletion and mutagenesis of a putative IRF-1 binding site in the FasL promoter results in deficient expression of FasL. EMSAs demonstrate specific FasL promoter binding by IRF-1 and IRF-2. Forced expression of either IRF-1 or IRF-2 leads to FasL promoter activation in T cells and FasL expression in heterologous cells. Finally, suppression of IRF-1 expression in T cells results in deficient TCR induced FasL expression. These results confirm that the IRF family participates in the regulation of FasL gene expression. PMID- 10725706 TI - Positive selection is limited by available peptide-dependent MHC conformations. AB - Recent data suggest that the diversity of self peptides presented in the thymus during development contributes to positive selection of a diverse T cell repertoire. We sought to determine whether a previously defined "hole in the immunological repertoire" could be explained by the absence of an appropriate selecting self peptide. The repertoire defect in question is the inability of bm8 mice to make an H-2K-restricted response to OVA. Like other OVA-specific, H-2K restricted receptors, OT-I-transgenic T cells are not positively selected in bm8 mice. Using criteria we had previously established for identifying positive selection ligands, we found peptides that could restore positive selection of OT I thymocytes in bm8 mice. Thus, the T cell repertoire can be limited by a requirement for specific self peptides during development. Data with MHC-specific Abs suggested that peptides might be able to force MHC residues to adopt different conformations in Kb vs Kbm8. This shows that peptides can potentially contribute to ligand diversity both directly (via variability in the solvent exposed side chains) and indirectly (through their effect on the MHC conformation). Our data support a model where self peptide diversity allows selection of T cells specific for a broad range of MHC conformations. PMID- 10725707 TI - Regulation of lymphoid homeostasis by IL-2 receptor signals in vivo. AB - High-affinity IL-2R signals are required for peripheral lymphoid homeostasis in vivo. We found that CD25 was required for regulation of peripheral T cells in mice bearing either the DO11.10 MHC class II-restricted TCR transgene or an Iabeta-null mutation, suggesting that MHC class I- and class II-dependent T cell subsets are regulated independently by IL-2R signals. In contrast, deregulation of serum IgG1 levels in CD25-/- mice was dependent on CD4+ T cells. T cell expansion in DO11.10 CD25-/- mice was not preferential for cells escaping allelic exclusion by the TCR transgene, but was suppressed by a Rag-2-null mutation. Together, these findings suggest that endogenous TCR are required to trigger T cell expansion, but that CD25 regulates T cells activated by low-specificity signals. Expansion of DO11.10 T cells in response to cognate Ag was modestly reduced in CD25-/- T cells transferred into the normal lymphoid compartments of BALB/c mice. Moreover, activation-induced clonal contraction and apoptosis in vivo were intact in the absence of CD25. These data indicate that the regulatory role of high-affinity IL-2R signals extends beyond the control of Ag-specific responses and suggest a role for these signals in control of bystander T cell activation. PMID- 10725708 TI - Identification of a MHC class II-restricted human gp100 epitope using DR4-IE transgenic mice. AB - CD4+ T cells play a central role in the induction and persistence of CD8+ T cells in several models of autoimmune and infectious disease. To improve the efficacy of a synthetic peptide vaccine based on the self-Ag, gp100, we sought to provide Ag-specific T cell help. To identify a gp100 epitope restricted by the MHC class II allele with the highest prevalence in patients with malignant melanoma (HLA DRB1*0401), we immunized mice transgenic for a chimeric human-mouse class II molecule (DR4-IE) with recombinant human gp100 protein. We then searched for the induction of CD4+ T cell reactivity using candidate epitopes predicted to bind to DRB1*0401 by a computer-assisted algorithm. Of the 21 peptides forecasted to bind most avidly, murine CD4+ T cells recognized the epitope (human gp10044-59, WNRQLYPEWTEAQRLD) that was predicted to bind best. Interestingly, the mouse helper T cells also recognized human melanoma cells expressing DRB1*0401. To evaluate whether human CD4+ T cells could be generated from the peripheral blood of patients with melanoma, we used the synthetic peptide h-gp10044-59 to sensitize lymphocytes ex vivo. Resultant human CD4+ T cells specifically recognized melanoma, as measured by tumor cytolysis and the specific release of cytokines and chemokines. HLA class II transgenic mice may be useful in the identification of helper epitopes derived from Ags of potentially great clinical utility. PMID- 10725709 TI - The role of B7 costimulation in CD4/CD8 T cell homeostasis. AB - The effect of B7-mediated costimulation on T cell homeostasis was examined in studies of B7-1 (CD80) and B7-2 (CD86) transgenic as well as B7-deficient mice. B7 overexpression in transgenic mice resulted in marked polyclonal peripheral T cell hyperplasia accompanied by skewing toward an increased proportion of CD8 single-positive cells and a decreased proportion of CD4 single-positive cells in thymus and more markedly in peripheral T cells. B7-induced T cell expansion was dependent on both CD28 and TCR expression. Transgenic overexpression of B7-1 or B7-2 resulted in down-regulation of cell surface CD28 on thymocytes and peripheral T cells through a mechanism mediated by intercellular interaction. Mice deficient in B7-1 and B7-2 exhibited changes that were the reciprocal of those observed in B7-overexpressing transgenics: a marked increase in the CD4/CD8 ratio in peripheral T cells and an increase in cell surface CD28 in thymus and peripheral T cells. These reciprocal effects of genetically engineered increase or decrease in B7 expression indicate that B7 costimulation plays a physiological role in the regulation of CD4+ and CD8+ T cell homeostasis. PMID- 10725710 TI - Polarization of naive CD4+ T cells toward the Th1 subset by CTLA-4 costimulation. AB - In this study, we examined in vitro the role of CTLA-4 costimulation in the polarization of naive CD4+ T cells toward the Th1 subset. When CTLA-4 costimulation was blocked by the inclusion of anti-CTLA-4 Fab in cultures during priming of naive CD4+ T cells with anti-CD3 in the presence of splenic adherent cells, they were polarized toward the Th2 subset. Conversely, the engagement of CTLA-4 with immobilized anti-CTLA-4 or with CD80-P815 cells polarized naive CD4+ T cells costimulated with anti-CD3 and anti-CD28 toward the Th1 subset. The CTLA 4 costimulation during priming augmented TGF-beta1 mRNA accumulation in naive CD4+ T cells, and the inclusion of anti-TGF-beta in cultures for priming suppressed the effect of CTLA-4 costimulation on the Th1 polarization. The addition of low doses of TGF-beta1 in cultures for priming of naive CD4+ T cells enhanced the production of Th1 cytokines upon secondary stimulation, although Th2 cytokine production was not affected by the doses of TGF-beta1. The CTLA-4 costimulation was also shown to suppress IL-4 production of naive CD4+ T cells upon priming. These results indicate that the costimulation against CTLA-4 drives polarization of naive CD4+ T cells toward the Th1 subset independent of IL-12 through, at least in part, the enhancement of TGF-beta1 production, and it also hampers Th2 subset differentiation by affecting IL-4 production of naive CD4+ T cells. PMID- 10725711 TI - IL-4, IL-10, IL-13, and TGF-beta from an altered peptide ligand-specific Th2 cell clone down-regulate adoptive transfer of experimental autoimmune encephalomyelitis. AB - Experimental autoimmune encephalomyelitis (EAE) is induced in the SJL/J mouse by adoptive transfer of activated proteolipid protein peptide (PLP) 139-151-specific Th1 cells. T cells responding to altered peptide ligands (APL) of PLP, previously shown to induce Th2 differentiation and regulate disease in PLP-immunized mice, do not transfer EAE. However, the exact mechanism of disease regulation by APL specific T cells has not been elucidated. In this report, we show that 1F1, a Th2 clone specific for an APL of PLP139-151 can prevent adoptive transfer of EAE when cocultured with PLP-encephalitogenic spleen cells (PLP-spleen). Cytokines from activated 1F1 cells were detected by hybridization of mRNA to oligonucleotide arrays (DNA chip) and by ELISA. The Th2 cytokines found to be present at the highest protein and mRNA levels were evaluated for their role in suppression of adoptive transfer of EAE from PLP-activated spleen cell cultures. Abs to individual cytokines in 1F1 PLP-spleen cocultures suggested that IL-4, IL-13, and TGF-beta played a significant role in suppressing EAE. Abs to the combination of IL-4, IL-10, IL-13, and TGF-beta completely neutralized the protective effect of 1F1. Addition of Th2 cytokines to PLP-spleen cultures showed that IL-13 and TGF beta were each individually effective and low levels of IL-4 synergized with IL 13 to inhibit disease transfer. IL-5, IL-9, and IL-10 had little or no effect whereas GM-CSF slightly enhanced EAE. Our results demonstrate that Th2 cytokines derived from APL-specific Th2 cells can effectively down-regulate the encephalitogenic potential of PLP-spleen cells if present during their reactivation in culture. PMID- 10725712 TI - Regulatory CD4 T cells control the size of the peripheral activated/memory CD4 T cell compartment. AB - The mechanisms leading to stable T cell numbers in the periphery of a healthy animal are, to date, not well understood. We followed the expansion of CD45RBhigh (naive) and CD45RBlow (activated/memory) CD4 T cells transferred from normal mice into syngeneic Rag-20/0 recipients and the dynamics of peripheral reconstitution when both populations were coinjected. Naive cells acquired an activated phenotype and showed a high proliferative capacity that was dependent on the environment in which the recipients were kept (specific pathogen-free vs conventional housing conditions), the age of the recipients, and the presence of CD45RBlow T cells in the injected cohort. CD45RBlow CD4 T cells protected the host from CD45RBhigh CD4 T cell-induced inflammatory bowel disease and showed a limited degree of expansion. CD45RBlow CD4 T cells isolated from GF mice also showed the ability to prevent inflammatory bowel disease, indicating that at least part of the natural regulatory T cells are self-reactive. The results indicate that 1) peripheral T cell expansion in lymphocyte-deficient recipients represent classical immune responses, which are mainly promoted by exogenous Ags and 2) natural regulatory T cells control the size of the activated/memory peripheral CD4 T cell compartment. PMID- 10725713 TI - Targeting rare populations of murine antigen-specific T lymphocytes by retroviral transduction for potential application in gene therapy for autoimmune disease. AB - CD4+ T cells are important mediators in the pathogenesis of autoimmunity and would therefore provide ideal candidates for lymphocyte-based gene therapy. However, the number of Ag-specific T cells in any single lesion of autoimmunity may be quite low. Successful gene transfer into autoantigen-specific CD4+ T cells would serve as an ideal vehicle for site-targeted gene therapy if it were possible to transduce preferentially the small number of autoantigen-specific T cells. In this study we have demonstrated that retroviral infection of CD4+ lymphocytes from either autoantigen-stimulated TCR transgenic mice, or Ag activated immunized nontransgenic mice, with a retroviral vector (pGCIRES), resulted in the transduction of only the limited number of Ag-reactive CD4+ T cells. In contrast, polyclonal activation of the same cultures resulted in transduction of non-antigen-specific lymphocytes. Transduction of Ag-reactive CD4+ T cells with pGCIRES retrovirus encoding the regulatory genes IL-4 (IL4) and soluble TNF receptor (STNFR) resulted in stable integration and long-term expression of recombinant gene products. Moreover, expression of the pGCIRES marker protein, GFP, directly correlated with the expression of the upstream regulatory gene. Retroviral transduction of CD4+ T cells targeted specifically Ag reactive cells and was cell cycle-dependent and evident only during the mitosis phase. These studies suggest that retroviral transduction of autoantigen-specific murine CD4+ T cells, using the pGCIRES retroviral vector, may provide a potential method to target and isolate the low frequency of autoantigen-specific murine CD4+ T cells, and provides a rational approach to gene therapy in animal models of autoimmunity. PMID- 10725714 TI - Coordinate regulation of T cell activation by CD2 and CD28. AB - T cell activation requires co-engagement of the TCR with accessory and costimulatory molecules. However, the exact mechanism of costimulatory function is unknown. Mice lacking CD2 or CD28 show only mild deficits, demonstrating that neither protein is essential for T cell activation. In this paper we have generated mice lacking both CD2 and CD28. T cells from the double-deficient mice have a profound defect in activation by soluble anti-CD3 Ab and Ag, yet remain responsive to immobilized anti-CD3. This suggests that CD2 and CD28 may function together to facilitate interactions of the T cell and APC, allowing for efficient signal transduction through the TCR. PMID- 10725715 TI - Indoleamine 2,3-dioxygenase production by human dendritic cells results in the inhibition of T cell proliferation. AB - Dendritic cells (DCs) play a key role in the activation and regulation of B and T lymphocytes. Production of indoleamine 2, 3-dioxygenase (IDO) by macrophages has recently been described to result in inhibition of T cell proliferation through tryptophan degradation. Since DCs can be derived from monocytes, we sought to determine whether DCs could produce IDO which could potentially regulate T cell proliferation. Northern blot analysis of RNA from cultured monocyte-derived human DC revealed that IDO mRNA was induced upon activation with CD40 ligand and IFN gamma. IDO produced from activated DCs was functionally active and capable of metabolizing tryptophan to kynurenine. Activated T cells were also capable of inducing IDO production by DCs, which was inhibited by a neutralizing Ab against IFN-gamma. DC production of IDO resulted in inhibition of T cell proliferation, which could be prevented using the IDO inhibitor 1-methyl-dl -tryptophan. These results suggest that activation of DCs induces the production of functional IDO, which causes depletion of tryptophan and subsequent inhibition of T cell proliferation. This may represent a potential mechanism for DCs to regulate the immune response. PMID- 10725716 TI - Large-scale culture and selective maturation of human Langerhans cells from granulocyte colony-stimulating factor-mobilized CD34+ progenitors. AB - Dendritic cells (DCs) play a critical role as APCs in the induction of the primary immune response. Their capacity for Ag processing and presentation is tightly regulated, controlled by a terminal developmental sequence accompanied by striking changes in morphology, organization, and function. The maturation process, which converts DCs from cells adapted for Ag accumulation to cells adapted for T cell stimulation, remains poorly understood due in part to difficulties in the culture and manipulation of DCs of defined lineages. To address these issues, we have devised conditions for the culture of a single DC type, Langerhans cells (LCs), using CD34+ cells from G-CSF-mobilized patients. Homogenous populations of LCs, replete with abundant immunocytochemically demonstrable Birbeck granules, could be stably maintained as immature DCs for long periods in culture. Unlike other human DC preparations, the LCs remained fully differentiated after cytokine removal. Following exposure to TNF-alpha, LPS, or CD40 ligand, the LCs could be synchronously induced to mature. Depending on the agent used, distinct types of LCs emerged differing in their capacity for T cell stimulation, IL-12 production, intracellular localization of MHC products, and overall morphology. Most interestingly, the expression of different sets of Toll family receptors is induced or down-regulated according to the maturation stimulus provided. These results strongly suggest that different proinflammatory stimuli might drive distinct developmental events. PMID- 10725717 TI - IL-15 is highly expressed in inflammatory bowel disease and regulates local T cell-dependent cytokine production. AB - IL-15 shares biological activities but no significant sequence homology with IL 2. It induces T cell recruitment to sites of inflammation, T cell proliferation, and cytokine production and rescue from apoptosis. The aim of this study was to investigate expression of IL-15 and its effects on proinflammatory cytokine production in inflammatory bowel disease (IBD). Immunohistochemistry demonstrated local IL-15 production by macrophages in inflamed mucosa from IBD patients. Isolated lamina propria mononuclear cells from these patients but not from controls produced IL-15 when stimulated with LPS or IFN-gamma. Moreover, lamina propria T cells (LP-T) from IBD patients were more responsive to IL-15 as compared with controls, and IL-15 alone without a primary T cell stimulus induced IFN-gamma and TNF production by isolated IBD LP-T cells, especially by LP-T cells from patients with Crohn's disease. LP-T cells from IBD patients could induce CD40-CD40 ligand (CD40L) interaction-dependent TNF and IL-12 production by monocytes in a coculture system. This capacity of LP-T cells was strongly enhanced by preincubation in IL-15 and was the result of higher CD40L expression after culture in IL-15. These data indicate that IL-15 is overexpressed in the inflamed mucosa in IBD and that IL-15 enhances local T cell activation, proliferation, and proinflammatory cytokine production by both T cells and macrophages, the latter via a CD40-CD40L interaction-dependent mechanism. Treatment directed against IL-15 may have therapeutic potential in IBD. PMID- 10725718 TI - Role of gut cryptopatches in early extrathymic maturation of intestinal intraepithelial T cells. AB - Lympho-hemopoietic progenitors residing in murine gut cryptopatches (CP) have been shown to generate intestinal intraepithelial T cells (IEL). To investigate the role of CP in progenitor maturation, we analyzed IEL in male mice with a truncated mutation of common cytokine receptor gamma-chain (CRgamma-/Y) in which CP were undetectable. IEL-expressing TCR-gammadelta (gammadelta-IEL) were absent, and a drastically reduced number of Thy-1highCD4+ and Thy-1highCD8alphabeta+ alphabeta-IEL were present in CRgamma-/Y mice, whereas these alphabeta-IEL disappeared from athymic CRgamma-/Y littermate mice. Athymic CRgamma-/Y mice possessed a small TCR- and alphaEbeta7 integrin-negative IEL population, characterized by the disappearance of the extrathymic CD8alphaalpha+ subset, that expressed pre-Talpha, RAG-2, and TCR-Cbeta but not CD3epsilon transcripts. These TCR- IEL from athymic CRgamma-/Y mice did not undergo Dbeta-Jbeta and Vdelta Jdelta joinings, despite normal rearrangements at the TCR-beta and -delta loci in thymocytes from euthymic CRgamma-/Y mice. In contrast, athymic severe combined immunodeficient mice in which CP developed normally possessed two major TCR alphaEbeta7+ CD8alphaalpha+ and CD8- IEL populations that expressed pre-Talpha, RAG-2, TCR-Cbeta, and CD3epsilon transcripts. These findings underscore the role of gut CP in the early extrathymic maturation of CD8alphaalpha+ IEL, including cell-surface expression of alphaEbeta7 integrin, CD3epsilon gene transcription, and TCR gene rearrangements. PMID- 10725719 TI - Anti-LFA-1 therapy induces long-term islet allograft acceptance in the absence of IFN-gamma or IL-4. AB - mAb therapy directed against a variety of cell surface accessory molecules has been effectively utilized to prolong allograft acceptance in various models of tissue and organ transplantation. The purpose of this study was to determine whether transient therapy directed against the adhesion molecule LFA-1 (CD11a) was sufficient to induce donor-specific tolerance to pancreatic islet allografts. Anti-LFA-1 monotherapy was found to be efficacious in inducing long-term islet allograft acceptance in multiple donor-recipient strain combinations. Graft acceptance following anti-LFA-1 therapy was not simply due to clonal ignorance of donor Ags in that the majority of recipients bearing established islet allografts resisted rejection induced by immunization with donor-type APCs. Furthermore, donor-specific tolerance from anti-LFA-1-treated animals could be transferred to secondary immune-deficient animals. Taken together, these results indicated that transient anti-LFA-1 monotherapy resulted in donor-specific tolerance. In vitro, functionally tolerant animals retained normal anti-donor reactivity as assessed by proliferative, cytotoxic, and cytokine release assays that demonstrated that tolerance was not secondary to general clonal deletion or anergy of donor reactive T cells. Finally, anti-LFA-1 treatment was effective in both IL-4 deficient and IFN-gamma-deficient recipients, indicating that neither of these cytokines are universally required for allograft acceptance. These results suggest that anti-adhesion-based therapy can induce a nondeletional form of tolerance that is not overtly dependent on the prototypic Th1 and Th2 cytokines, IFN-gamma and IL-4, respectively, in contrast to results in other transplantation models. PMID- 10725720 TI - Analysis of signals and functions of the chimeric human granulocyte-macrophage colony-stimulating factor receptor in BA/F3 cells and transgenic mice. AB - Receptors for GM-CSF, IL-3, and IL-5 are composed of two subunits: alpha, which is specific for each cytokine, and betac, which is shared by all. Although the role of betac in signal transduction has been extensively studied, the role of the alpha subunit has remained to be clarified. To analyze the role of the human (h) GM-CSF receptor alpha subunit, we constructed a chimeric receptor subunit composed of extracellular and transmembrane regions of alpha fused with the cytoplasmic region of betac, designated alpha/beta. In BA/F3 cells, chimeric receptor composed of alpha/beta,beta can transduce signals for mitogen-activated protein kinase cascade activation and proliferation in response to hGM-CSF. Although phosphorylation of Jak1 but not of Jak2 occurred with stimulation of hGM CSF, the dominant-negative Jak2 but not the dominant-negative Jak1 suppresses c fos promoter activation. To determine whether the chimeric receptor alpha/beta,beta is functional in vivo, we developed transgenic mice expressing the chimeric receptor alpha/beta,beta. Bone marrow cells from the transgenic mice expressing the alpha/beta,beta receptor form not only GM colonies but also various lineages of colonies in response to GM-CSF. In addition, mast cells were produced when bone marrow cells of the transgenic mouse were cultured with hGM CSF. Thus, it appears that the cytoplasmic region of the alpha subunit is not required for hGM-CSF promoting activities, even in bone marrow cells. PMID- 10725721 TI - Crucial role of TNF-alpha in CD8 T cell-mediated elimination of 3LL-A9 Lewis lung carcinoma cells in vivo. AB - The role of perforin, IFN-gamma, and TNF-alpha in anti-tumor CD8 T cell immunity was examined in a new tumor model using a CD8 T cell epitope (GP33) derived from lymphocytic choriomeningitis virus as a tumor-associated Ag. In contrast with parental 3LL-A9 (A9) Lewis lung carcinoma cells that progressively grow in C57BL/6 mice, s.c. injection of GP33-transfected A9GP33 tumor cells induced a protective GP33-specific CD8 T cell response that led to complete tumor cell elimination. Tumor regression was dependent on perforin, IFN-gamma, or TNF-alpha, because A9GP33 tumors developed in mice deficient in one of these genes. A9GP33 tumors arising in perforin- and IFN-gamma-deficient mice represented GP33 Ag-loss variants, demonstrating that GP33-specific CD8 T cells from these mice were able to exert an Ag selection pressure. In contrast, tumor cells growing in TNF-alpha knock-out mice still expressed the tumor-associated GP33 peptide despite the presence of activated GP33-specific CD8 T cells. These findings provide evidence for a crucial role of TNF-alpha in A9 tumor cell elimination by CD8 T cells in vivo. PMID- 10725722 TI - SHPS-1 induces aggregation of Ba/F3 pro-B cells via an interaction with CD47. AB - SHPS-1 (SH2-domain bearing protein tyrosine phosphatase (SHP) substrate-1), a member of the inhibitory-receptor superfamily that is abundantly expressed in macrophages and neural tissue, appears to regulate intracellular signaling events downstream of receptor protein-tyrosine kinases and integrin-extracellular matrix molecule interactions. To investigate the function of SHPS-1 in a hematopoietic cell line, SHPS-1 was expressed in Ba/F3 cells, an IL-3-dependent pro-B-cell line that lacks endogenous SHPS-1 protein. Interestingly, expression of either SHPS-1, or a mutant lacking the intracellular domain of SHPS-1 (DeltaCT SHPS-1), resulted in the rapid formation of macroscopic Ba/F3 cell aggregates. As the integrin associated protein/CD47 was shown to be a SHPS-1 ligand in neural cells, we investigated whether CD47 played a role in the aggregation of SHPS-1-expressing Ba/F3 cells. In support of this idea, aggregate formation was inhibited by an anti-CD47 Ab. Furthermore, erythrocytes from control, but not from CD47-deficient mice, were able to form rosettes on SHPS-1-expressing Ba/F3 cells. Because erythrocytes do not express integrins, this result suggested that SHPS-1-CD47 interactions can take place in the absence of a CD47-integrin association. We also present evidence that the amino-terminal Ig domain of SHPS-1 mediates the interaction with CD47. Although SHPS-1-CD47 binding likely triggers bidirectional intracellular signaling processes, these results demonstrate that this interaction can also mediate cell-cell adhesion. PMID- 10725723 TI - Correct immunoglobulin alpha mRNA processing depends on specific sequence in the C alpha 3-alpha M intron. AB - The maturation of IgM-expressing B cells to IgM-secreting plasma cells is associated with both an increase in mu mRNA and the ratio of secreted to membrane forms of mu mRNA which differ at the 3' termini. In contrast, both in vitro and in vivo the secreted form of alpha mRNA is predominant at all stages in the development of a secretory IgA response. Previous studies demonstrated that preferential usage of the alpha s poly(A) site does not result from transcription termination and is independent of either the poly(A) sites or the 3' splice site associated with the exon encoding the membrane exon of IgA (alpha M). The present study demonstrates that a 349-bp region located 774 bp 3' to the alpha s poly(A) site is required for the preferential usage of the alpha s terminus. This region, which is the first isotype-specific cis-acting regulatory sequence not immediately adjacent to a secretory poly(A) site to be identified, contains regulatory elements that increase the efficiency of polyadenylation/cleavage. A ubiquitous, approximately 58-kDa RNA-binding protein interacts specifically with this regulatory region. These studies support the premise that cis-acting elements unique to each CH gene can impinge upon a common mechanism regulating Ig mRNA processing. PMID- 10725724 TI - Novel mutations within the RFX-B gene and partial rescue of MHC and related genes through exogenous class II transactivator in RFX-B-deficient cells. AB - MHC class II deficiency or bare lymphocyte syndrome is a severe combined immunodeficiency caused by defects in MHC-specific regulatory factors. Fibroblasts derived from two recently identified bare lymphocyte syndrome patients, EBA and FZA, were found to contain novel mutations in the RFX-B gene. RFX-B encodes a component of the RFX transcription factor that functions in the assembly of multiple transcription factors on MHC class II promoters. Unlike RFX5 and RFXAP-deficient cells, transfection of exogenous class II transactivator (CIITA) into these RFX-B-deficient fibroblasts resulted in the induction of HLA DR and HLA-DP and, to a lesser extent, HLA-DQ. Similarly, CIITA-mediated induction of MHC class I, beta2-microglobulin, and invariant chain genes was also found in these RFX-B-deficient fibroblasts. Expression of wild-type RFX-B completely reverted the noted deficiencies in these cells. Transfection of CIITA into Ramia cells, a B cell line that does not produce a stable RFX-B mRNA, resulted in induction of an MHC class II reporter, suggesting that CIITA overexpression may partially override the RFX-B defect. PMID- 10725725 TI - p53-dependent and -independent pathways of apoptotic cell death in sepsis. AB - Sepsis induces extensive apoptosis of lymphocytes, which may be responsible for the profound immune suppression of the disorder. Two potential pathways of sepsis induced lymphocyte apoptosis, Fas and p53, were investigated. Lymphocyte apoptosis was evaluated 20-22 h after sepsis by annexin V or DNA nick-end labeling. Fas receptor-deficient mice had no protection against sepsis-induced apoptosis in thymocytes or splenocytes. p53 knockout mice (p53-/-) had complete protection against thymocyte apoptosis but, surprisingly, had no protection in splenocytes. p53-/- mice had no improvement in sepsis survival compared with appropriately matched control mice with sepsis. We conclude that both p53 dependent and p53-independent pathways of cell death exist in sepsis. This differential apoptotic response of thymocytes vs splenocytes in p53-/- mice suggests that either the cellular response or the death-inducing signal is cell type specific in sepsis. The fact that p53-/- lymphocytes of an identical subtype (CD8-CD4+) were protected in thymi but not in spleens indicates that cell susceptibility to apoptosis differs depending upon other unidentified factors. PMID- 10725726 TI - B cell-deficient mice are highly resistant to Leishmania donovani infection, but develop neutrophil-mediated tissue pathology. AB - Resolution of Leishmania infection is T cell-dependent, and B lymphocytes have been considered to play a minimal role in host defense. In this study, the contribution of B lymphocytes to the response against Leishmania donovani was investigated using genetically modified IgM transmembrane domain (muMT) mutant mice, which lack mature B lymphocytes. When compared with wild-type mice, muMT mice cleared parasites more rapidly from the liver, and infection failed to establish in the spleen. The rapid clearance of parasites in muMT mice was associated with accelerated and more extensive hepatic granuloma formation compared with wild-type mice. However, the liver of infected muMT mice also showed signs of destructive pathology, associated with the presence of increased numbers of neutrophils. The role of neutrophils in controlling parasite growth in the viscera was determined by depletion with the mAb RB6-8C5. This treatment led to a dramatic enhancement of parasite growth in both the liver and spleen of muMT and wild-type mice. As assessed by transfer of both normal and chronic-infection serum, Ig protects microMT mice from destructive hepatic pathology, but minimally alters their resistance compared with wild-type mice. However, adoptive transfer of CD4+ and CD8+ T cells into recombinase activating gene 1 (RAG1-/-) recipients, suggested that T cell function was not altered by maturation in a B cell deficient environment. Taken together, these data suggest an inhibitory role for B lymphocytes in resistance to L. donovani unrelated to the presence or absence of Ig. However, Ig protects muMT mice from the exaggerated pathology that occurs during infection. PMID- 10725727 TI - Role of CD40 ligand and CD28 in induction and maintenance of antiviral CD8+ effector T cell responses. AB - The primary aim of this report was to evaluate the immune responses of CD40 ligand-deficient (CD40L-/-) mice infected with two viruses known to differ markedly in their capacity to replicate in the host. Lymphocytic choriomeningitis virus (LCMV) is a natural mouse pathogen that replicates widely and extensively, whereas vesicular stomatitis virus (VSV) spreads poorly. We found that the primary response of CD40L-/- mice toward VSV is significantly impaired; proliferation of both CD4+ and CD8+ cells is reduced 2- to 3-fold, few CD8+ cells acquire an activated phenotype, and little functional activity is induced. Very similar results were obtained in VSV-infected, CD28-deficient mice. In contrast, neither CD40L nor CD28 was required for induction of a primary CD8+ response toward LCMV. Surprisingly, lack of CD4+ T cells had no impact on the primary immune response toward any of the viruses, even though the CD40 ligand dependence demonstrated for VSV would be expected to be associated with CD4 dependence. Upon coinfection of VSV-infected mice with LCMV, the requirement for CD40 ligand (but not CD28) could be partially bypassed, as evidenced by a 3-fold increase in the frequency of VSV-specific CD8+ T cells on day 6 postinfection. Finally, despite the fact that the primary LCMV-specific CD8+ response is virtually unimpaired in CD40L-/- mice, their capacity to maintain CD8+ effector activity and to permanently control the infection is significantly reduced. Thus, our results demonstrate that the importance of CD40/CD40L interaction for activation of CD8+ T cells varies between viruses and over time. PMID- 10725728 TI - Neonatal administration of IL-12 enhances the protective efficacy of antiviral vaccines. AB - Neonates are highly susceptible to infectious agents and are known to display polarized expression of Th2-like cytokines and Abs. This neonatal immune bias has important implications for the development of vaccine strategies, particularly against viral infections. We now report that coadministration of IL-12 and an influenza subunit vaccine at birth enhances the protective efficacy of antiviral vaccination. Immunization and treatment with IL-12 within 24 h of birth resulted in elevated expression of IFN-gamma, IL-10, and IL-15 mRNA in the spleens of newborn mice compared with animals exposed to vaccine only. In addition, these animals showed dramatic increases in IFN-gamma-, IL-2-, and IL-4-secreting cells, and in IgG2a Ab levels upon adult challenge compared with mice primed with vaccine alone. Most importantly, animals vaccinated and simultaneously treated with IL-12 at birth displayed enhanced survival after lethal challenge with infectious influenza virus as adults compared with infected animals that had been primed with vaccine alone. This augmented protection required B cells and could be transferred to naive mice by immune serum. Collectively, these results provide evidence that administration of IL-12 to neonates induces a Th1-like response in newborns and elicits protective antiviral immune memory. PMID- 10725729 TI - Redirected perforin-dependent lysis of colon carcinoma by ex vivo genetically engineered CTL. AB - The redirection of autologous lymphocytes to predefined tumor target Ags has considerable potential for the immunotherapeutic treatment of cancer; however, robust experimental systems for comparing various approaches have not been developed. Herein, we have generated a single chain variable domain anti carcinoembryonic Ag (CEA) Fcepsilon receptor I gamma-chain fusion (scFv anti-CEA) receptor and demonstrated high-level expression of this chimeric receptor in naive mouse T lymphocytes by retroviral gene transduction. These gene-modified CTL were able to lyse CEA+ targets and secrete high levels of IFN-gamma following Ag stimulation. Depletion studies demonstrated that specific tumor cell cytotoxicity was mediated by gene-modified CD8+ T cells. Importantly, in increasingly stringent tests of efficacy in vivo, transduced CTL were sequentially shown to reject CEA+ colon carcinoma cells in a Winn assay and then reject established s.c. colon carcinoma in scid or syngeneic mice. Furthermore, using gene-targeted and scFv anti-CEA receptor-transduced donor CTL, perforin and IFN-gamma were demonstrated to be absolutely critical for the eradication of colon carcinoma in mice. In summary, we have developed a highly efficient gene transfer system for evaluating chimeric receptor expression in cytotoxic lymphocytes. This series of experiments has revealed the utility of scFv anti-CEA chimeras in providing mouse T cells the capacity to reject colon carcinoma in an Ag- and perforin-specific manner. PMID- 10725730 TI - Survival of Staphylococcus aureus inside neutrophils contributes to infection. AB - Neutrophils have long been regarded as essential for host defense against Staphylococcus aureus infection. However, survival of the pathogen inside various cells, including phagocytes, has been proposed as a mechanism for persistence of this microorganism in certain infections. Therefore, we investigated whether survival of the pathogen inside polymorphonuclear neutrophils (PMN) contributes to the pathogenesis of S. aureus infection. Our data demonstrate that PMN isolated from the site of infection contain viable intracellular organisms and that these infected PMN are sufficient to establish infection in a naive animal. In addition, we show that limiting, but not ablating, PMN migration into the site of infection enhances host defense and that repletion of PMN, as well as promoting PMN influx by CXC chemokine administration, leads to decreased survival of the mice and an increased bacterial burden. Moreover, a global regulator mutant of S. aureus (sar-) that lacks the expression of several virulence factors is less able to survive and/or avoid clearance in the presence of PMN. These data suggest that the ability of S. aureus to exploit the inflammatory response of the host by surviving inside PMN is a virulence mechanism for this pathogen and that modulation of the inflammatory response is sufficient to significantly alter morbidity and mortality induced by S. aureus infection. PMID- 10725731 TI - Direct visualization of cytokine-producing recall antigen-specific CD4 memory T cells in healthy individuals and HIV patients. AB - We have used computer-assisted cytokine ELISA spot analysis to measure the frequencies, the type of cytokine, and the amount of cytokine produced by individual recall Ag-specific CD4 memory cells in freshly isolated blood. We studied the memory cells specific for tetanus toxoid and purified protein derivative in 18 healthy individuals and in 22 HIV-infected patients on highly active antiretroviral therapy (HAART). In healthy individuals, the frequency, cytokine signature, and cytokine production per cell of these memory cells were stable over time. Although it is presently unclear whether the maintenance of the memory T cell pool depends upon Ag persistence, cross-reactive Ag stimulation, or cytokine-driven bystander stimulations and expansions, our data strongly argue for a stable memory cell pool in healthy individuals. In HIV patients, however, the frequency of these memory cells was a function of the viral load. The decreased numbers of functional memory cells in patients with high viral loads might provide one mechanism behind the immunodeficient state. Although the cytokine output per cell was unaffected in most patients (20 of 24), in some patients (4 of 24) it was >100-fold reduced, which might provide an additional mechanism to account for the reduced immunocompetence of these patients. The ability to visualize directly and quantify the cytokine produced by the low frequency memory cells in freshly isolated blood that have been physiologically stimulated by Ag should aid comprehensive studies of the Ag-specific memory cell pool in vivo, in health and disease. PMID- 10725732 TI - Lactobacilli and Streptococci activate NF-kappa B and STAT signaling pathways in human macrophages. AB - Gram-positive bacteria induce the production of several cytokines in human leukocytes. The molecular mechanisms involved in Gram-positive bacteria-induced cytokine production have been poorly characterized. In this work we demonstrate that both nonpathogenic Lactobacillus rhamnosus GG and pathogenic Streptococcus pyogenes (group A streptococci) induce NF-kappa B and STAT DNA-binding activity in human primary macrophages as analyzed by EMSA. NF-kappa B activation was rapid and was not inhibited by a protein synthesis inhibitor cycloheximide, suggesting that these bacteria could directly activate NF-kappa B. STAT1, STAT3, and IFN regulatory factor-1 DNA binding was induced by both bacteria with delayed kinetics compared with NF-kappa B. In addition, streptococci induced the formation of IFN-alpha-specific transcription factor complex and IFN-stimulated gene factor-3 (ISGF3). STAT1 and STAT3 activation and ISGF3 complex formation were inhibited by cycloheximide or by neutralization with IFN-alpha/beta-specific Abs. Streptococci were more potent than lactobacilli in inducing STAT1, ISGF3, and IFN regulatory factor-1 DNA binding. Accordingly, only streptococci induced IFN-alpha production. The activation of the IFN-alpha signaling pathway by streptococci could play a role in the pathogenesis of these bacteria. These results indicate that extracellular Gram-positive bacteria activate transcription factors involved in cytokine signaling by two mechanisms: directly, leading to NF kappa B activation, and indirectly via cytokines, leading to STAT activation. PMID- 10725733 TI - T cell clones raised from chronically infected healthy humans by stimulation with Toxoplasma gondii excretory-secretory antigens cross-react with live tachyzoites: characterization of the fine antigenic specificity of the clones and implications for vaccine development. AB - Excreted-secreted Ags (ESA) of Toxoplasma gondii (Tg) play an important role in the stimulation of the host immune system in both acute and chronic infections. To identify the parasite Ag(s) involved in the maintenance of T cell-mediated long term immunity, 40 ESA-specific T cell clones were derived from three chronically infected healthy subjects. All the clones were CD4+ and recognized both ESA and live tachyzoites in a HLA-DR-restricted manner. Conversely, CD4+ tachyzoite-specific T cell clones from the same subjects proliferated in response to ESA, pointing to shared immunodominant Ags between ESA and Tg tachyzoites. By T cell blot analysis using SDS-PAGE-fractionated parasite extracts, the following patterns of reactivity were detected. Of 25 clones, 6 recognized Tg fractions in the 24- to 28-kDa range and proliferated to purified GRA2, 5 reacted with Tg fractions in the 30- to 33-kDa range; and 4 of them proved to be specific for rSAg1. Although surface Ag (SAg1) is not a member of ESA, small amounts of this protein were present in ESA preparation by Western blot. Of 25 clones, 8 responded to Tg fractions in the 50- to 60-kDa range but not to the 55-kDa recombinant rhoptries-2 parasite Ag, and 6 did not react with any Tg fraction but proliferated in response to either ESA or total parasite extracts. In conclusion, CD4+ T cells specific for either ESA (GRA2) or SAg1 may be involved in the maintenance of long term immunity to Tg in healthy chronically infected individuals. PMID- 10725734 TI - A role of mast cell glycosaminoglycans for the immunological expulsion of intestinal nematode, Strongyloides venezuelensis. AB - We examined effects of mast cell glycosaminoglycans on the establishment of the intestinal nematode, Strongyloides venezuelensis, in the mouse small intestine. When intestinal mastocytosis occurred, surgically implanted adult worms could not invade and establish in the intestinal mucosa. In mast cell-deficient W/Wv mice, inhibition of adult worm invasion was not evident as compared with littermate +/+ control mice. Mucosal mastocytosis and inhibition of S. venezuelensis adult worm mucosal invasion was tightly correlated. To determine effector molecules for the invasion inhibition, adult worms were implanted with various sulfated carbohydrates including mast cell glycosaminoglycans. Among sulfated carbohydrates tested, chondroitin sulfate (ChS)-A, ChS-E, heparin, and dextran sulfate inhibited invasion of adult worms into intestinal mucosa in vivo. No significant inhibition was observed with ChS-C, desulfated chondroitin, and dextran. ChS-E, heparin, and dextran sulfate inhibited adhesion of S. venezuelensis adult worms to plastic surfaces in vitro. Furthermore, binding of intestinal epithelial cells to adhesion substances of S. venezuelensis, which have been implicated in mucosal invasion, was inhibited by ChS-E, heparin, and dextran sulfate. Because adult worms of S. venezuelensis were actively moving in the intestinal mucosa, probably exiting and reentering during infection, the possible expulsion mechanism for S. venezuelensis is inhibition by mast cell glycosaminoglycans of attachment and subsequent invasion of adult worms into intestinal epithelium. PMID- 10725735 TI - Isolated Pneumocystis carinii cell wall glucan provokes lower respiratory tract inflammatory responses. AB - Macrophage-induced lung inflammation contributes substantially to respiratory failure during Pneumocystis carinii pneumonia. We isolated a P. carinii cell wall fraction rich in glucan carbohydrate, which potently induces TNF-alpha and macrophage-inflammatory protein-2 generation from alveolar macrophages. Instillation of this purified P. carinii carbohydrate cell wall fraction into healthy rodents is accompanied by substantial increases in whole lung TNF-alpha generation and is associated with neutrophilic infiltration of the lungs. Digestion of the P. carinii cell wall isolate with zymolyase, a preparation containing predominantly beta-1,3 glucanase, substantially reduces the ability of this P. carinii cell wall fraction to activate alveolar macrophages, thus suggesting that beta-glucan components of the P. carinii cell wall largely mediate TNF-alpha release. Furthermore, the soluble carbohydrate beta-glucan receptor antagonists laminariheptaose and laminarin also substantially reduce the ability of the P. carinii cell wall isolate to stimulate macrophage-inflammatory activation. In contrast, soluble alpha-mannan, a preparation that antagonizes macrophage mannose receptors, had minimal effect on TNF-alpha release induced by the P. carinii cell wall fraction. P. carinii beta-glucan-induced TNF-alpha release from alveolar macrophages was also inhibited by both dexamethasone and pentoxifylline, two pharmacological agents with potential activity in controlling P. carinii-induced lung inflammation. These data demonstrate that P. carinii beta glucan cell wall components can directly stimulate alveolar macrophages to release proinflammatory cytokines mainly through interaction with cognate beta glucan receptors on the phagocyte. PMID- 10725736 TI - Resistance against the membrane attack complex of complement induced in porcine endothelial cells with a Gal alpha(1-3)Gal binding lectin: up-regulation of CD59 expression. AB - Endothelial cells (EC) play central roles in vascular physiology and pathophysiology. EC activation results in proinflammatory activities with production of cytokines and expression of adhesion molecules. However, we have shown before in a model of xenotransplantation that prolonged stimulation of porcine EC with human anti-porcine IgM natural Abs can activate the cells to become resistant against cytotoxicity by the membrane attack complex of complement (MAC). Now we report the major characteristics of induction and maintenance of resistance elicited in porcine EC with Bandeiraea simplicifolia lectin that binds terminal gal alpha(1-3)gal. Lectin-treated cells underwent little or no cytotoxicity and PGI2 release when exposed to MAC. Induction of resistance required incubation of the EC with lectin for 4 h but was not fully manifested until 16 h later. Most of the initially bound lectin remained on the cell surface for >60 h. EC-bound lectin did not inhibit binding of IgM natural Abs or activation and binding of C components, including C9, but a C-induced permeability channel of reduced size was present. Induction of resistance required protein synthesis, developed slowly, and was associated with up regulation of expression of mRNA for the MAC inhibitor CD59 and membrane associated CD59 protein. Resistance lasted at least 3 days, and the cells regained normal morphology and were metabolically active. This induced resistance may have a physiologic counterpart that might be amenable to pharmacologic manipulation in vascular endothelium pathophysiology. PMID- 10725737 TI - Identification of unique truncated KC/GRO beta chemokines with potent hematopoietic and anti-infective activities. AB - SK&F 107647, a previously described synthetic immunomodulatory peptide, indirectly stimulates bone marrow progenitor cells and phagocytic cells, and enhances host defense effector mechanisms in bacterial and fungal infection models in vivo. In vitro, SK&F 107647 induces the production of a soluble mediator that augments colony forming cell (CFU-GM) formation in the presence of CSFs. In this paper we purified and sequenced the stromal cell-derived hematopoietic synergistic factors (HSF) secreted from both murine and human cell lines stimulated with SK&F 107647. Murine HSF is an N-terminal 4-aa truncated form of the CXC chemokine, KC, while human HSF was identified as an N-terminal 4 aa truncated form of the CXC chemokine, GRO beta. In comparison to their full length forms, truncated KC and truncated GRO beta were 10 million times more potent as synergistic growth stimulants for CFU-GM. Enhanced potency of these novel truncated chemokines relative to their full-length forms was also demonstrated in respiratory burst assays, CD11b Ag expression, and intracellular killing of the opportunistic pathogen, Candida albicans. Administration of truncated KC significantly enhanced survival of mice lethally infected with C. albicans. The results reported herein delineate the biological mechanism of action of SK&F 107647, which functions via the induction of unique specific truncated forms of the chemokines KC and GRO beta. To our knowledge, this represents the first example where any form of KC or GRO beta were purified from marrow stromal cells. Additionally, this is the first demonstration of in vivo efficacy of a CXC chemokine in an animal infectious fungal disease model. PMID- 10725738 TI - Characterization of synthetic C3a analog peptides on human eosinophils in comparison to the native complement component C3a. AB - The C3a anaphylatoxin is a potent proinflammatory mediator derived from the complement system inducing biologic effects of human eosinophils like Ca2+ transients and the activation of the respiratory burst. These findings support an important role for C3a in diseases typically associated with a peripheral blood or tissue eosinophilia. Synthetic human C3a analogue peptides with variations at the C-terminal effector domain have been evaluated with respect to their binding affinity and signaling potency on human eosinophils. Flow cytometrical analysis and RT-PCR revealed that the C3a receptor is constitutively expressed on human eosinophils. Peptides bearing an N-terminal 9-fluorenylmethoxycarbonyl and the 6 aminohexanoyl motif were the most powerful peptides tested. Amino acid replacements in the conserved C-terminal pentapeptide decreased binding affinity and functional potency substantially. In addition, synthetic C3a analogue peptides induced C3aR internalization, led to transient changes of intracellular Ca2+ concentration, and did release reactive oxygen species in human eosinophils indicating the in vivo relevance of C3a-related sequences. The tripeptide LAR was found to be essential for C3a receptor binding on human eosinophils. Moreover, the putative binding motif of C3a anaphylatoxin is also crucial for the induction of biologic effects in the human system such as changes of intracellular Ca2+ concentration and the release of reactive oxygen species. This study demonstrates that the carboxyl terminus is important for the interaction with the C3aR and the biologic potency of C3a anaphylatoxin in the human system and plays a key role in the activation process of human eosinophils. PMID- 10725739 TI - Aerosolized Syk antisense suppresses Syk expression, mediator release from macrophages, and pulmonary inflammation. AB - Syk protein tyrosine kinase (PTK) is involved in signaling in leukocytes. In macrophages, Fcgamma-receptor cross-linking induces Syk PTK phosphorylation and activation, resulting in Syk-dependent events required for phagocytosis and mediator release. We hypothesized that Syk antisense oligodeoxynucleotides (ASO) delivered by aerosol to rat lungs in vivo would depress Syk PTK expression, mediator release from alveolar macrophages, and Syk-dependent pulmonary inflammation. RT-PCR and RT-in situ PCR demonstrated that aerosolized Syk ASO administration reduced Syk mRNA expression from alveolar macrophages compared with cells isolated from sham-treated rats. Western blot analysis confirmed that Syk PTK expression was reduced after Syk ASO treatment. Compared with sham treated rats (scrambled oligodeoxynucleotide), Syk ASO treatment suppressed Fcgamma-receptor-mediated nitric oxide (86.0 +/- 8.3%) and TNF (73.1 +/- 3.1%) production by alveolar macrophages stimulated with IgG-anti-IgG complexes. In contrast, Fcgamma-receptor-induced IL-1beta release was unaffected by Syk ASO treatment. Additionally, Syk ASO suppressed Ag-induced pulmonary inflammation, suggesting that Syk ASO may prove useful as an anti-inflammatory therapy in disorders such as asthma. PMID- 10725740 TI - Shear and time-dependent changes in Mac-1, LFA-1, and ICAM-3 binding regulate neutrophil homotypic adhesion. AB - We examined the relative contributions of LFA-1, Mac-1, and ICAM-3 to homotypic neutrophil adhesion over the time course of formyl peptide stimulation at shear rates ranging from 100 to 800 s-1. Isolated human neutrophils were sheared in a cone-plate viscometer and the kinetics of aggregate formation was measured by flow cytometry. The efficiency of cell adhesion was computed by fitting the aggregate formation rates with a model based on two-body collision theory. Neutrophil homotypic adhesion kinetics varied with shear rate and was most efficient at 800 s-1, where approximately 40% of the collisions resulted in adhesion. A panel of blocking Abs to LFA-1, Mac-1, and ICAM-3 was added to assess the relative contributions of these molecules. We report that 1) LFA-1 binds ICAM 3 as its primary ligand supporting homotypic adhesion, although the possibility of other ligands was also detected. 2) Mac-1 binding to an unidentified ligand supports homotypic adhesion with an efficiency comparable to LFA-1 at low shear rates of approximately 100 s-1. Above 300 s-1, however, Mac-1 and not LFA-1 were the predominant molecules supporting cell adhesion. This is in contrast to neutrophil adhesion to ICAM-1-transfected cells, where LFA-1 binds with a higher avidity than Mac-1 to ICAM-1. 3) Following stimulation, the capacity of LFA-1 to support aggregate formation decreases with time at a rate approximately 3-fold faster than that of Mac-1. The results suggest that the relative contributions of beta2 integrins and ICAM-3 to neutrophil adhesion is regulated by the magnitude of fluid shear and time of stimulus over a range of blood flow conditions typical of the venular microcirculation. PMID- 10725741 TI - Cultured human fibroblasts express constitutive IL-16 mRNA: cytokine induction of active IL-16 protein synthesis through a caspase-3-dependent mechanism. AB - Human fibroblasts can express numerous regulatory molecules that influence immune function. IL-16, a ligand for CD4, is a chemoattractant molecule expressed by lymphocytes, eosinophils, mast cells, and lung epithelium. It appears that the sole target for IL-16 is the CD4-bearing cell. Here we demonstrate that fibroblasts from several tissues can express IL-16 mRNA and protein as well as IL 16-dependent chemoattractant activity. The transcript is expressed abundantly under basal culture conditions as a 2.5-kb band on Northern analysis, similar to that observed in lymphocytes. IL-16 protein and activity are undetectable in fibroblast cultures under these same control conditions. However, when treated with proinflammatory cytokines such as IL-1beta, they express very high levels of IL-16 protein and chemoattractant activity, a substantial component of which can be blocked with IL-16-neutralizing Abs. The amount of IL-16 protein released into the medium is 3- to 4-fold greater, on a per cell basis, than that observed in lymphocytes. The induction of IL-16 protein by IL-1beta can be attenuated with specific inhibition of caspase-3, which could be detected in IL-1beta-treated fibroblasts. IL-1beta also induces RANTES mRNA, protein, and activity, and most of the chemoattractant activity released from fibroblasts not derived from IL-16 can be attributed to RANTES. Human fibroblasts appear to be an important source of IL-16 and through expression of this molecule may have key roles in the recruitment of CD4+ cells to sites of inflammation. IL-16 expression and the mechanism involved in its regulation appear to be cell type specific. PMID- 10725742 TI - Regulation of the RON receptor tyrosine kinase expression in macrophages: blocking the RON gene transcription by endotoxin-induced nitric oxide. AB - Previous studies have shown that activation of the RON receptor tyrosine kinase inhibits inducible NO production in murine peritoneal macrophages. The purpose of this study is to determine whether inflammatory mediators such as LPS, IFN-gamma, and TNF-alpha regulate RON expression. Western blot analysis showed that RON expression is reduced in peritoneal macrophages collected from mice injected with a low dose of LPS. The inhibition was seen as early as 8 h after LPS challenge. Experiments in vitro also demonstrated that the levels of the RON mRNA and protein are diminished in cultured peritoneal macrophages following LPS stimulation. TNF-alpha plus IFN-gamma abrogated macrophage RON expression, although individual cytokines had no significant effect. Because LPS and TNF alpha plus IFN-gamma induce NO production, we reasoned that NO might be involved in the RON inhibition. Two NO donors, S-nitroglutathione (GSNO) and (+/-)-S nitroso-N-acetylpenicillamine (SNAP), directly inhibited macrophage RON expression when added to the cell cultures. Blocking NO production by NO inhibitors like TGF-beta prevented the LPS-mediated inhibitory effect. In Raw264.7 cells transiently transfected with a report vector, GSNO or SNAP inhibited the luciferase activities driven by the RON gene promoter. Moreover, GSNO or SNAP inhibited the macrophage-stimulating protein-induced RON phosphorylation and macrophage migration. We concluded from these data that RON expression in macrophages is regulated during inflammation. LPS and TNF-alpha plus IFN-gamma are capable of down-regulating RON expression through induction of NO production. The inhibitory effect of NO is mediated by suppression of the RON gene promoter activities. PMID- 10725743 TI - Polymorphonuclear leukocytes modulate tissue factor production by mononuclear cells: role of reactive oxygen species. AB - To determine whether polymorphonuclear leukocytes (PMN) modulate the production of tissue factor (TF) by monocytes, PBMC were incubated with increasing concentrations of PMN. PMN did not express any procoagulant activity. After 20-h cocultures, PMN enhanced or inhibited the TF production of PBMC, and this effect depended on the PMN/PBMC ratio. When the ratio increased from 1/1000 to 1/5, without or with LPS, the TF activity of PBMC increased to peak at 2.5-fold the baseline value (p < 0.01). The TF Ag and TF mRNA also increased. This potentiating effect was mediated by reactive oxygen species (ROS) released by PMN during the coculture; it did not require direct cell contact between PMN and PBMC, it was enhanced when PMN were stimulated by fMLP (a chemotactic peptide), and it was inhibited by two antioxidants, N-acetyl cysteine and pyrrolidine dithiocarbamate. In contrast, when the PMN/PBMC ratio was further increased from 1/2 to 2/1, the PBMC TF activity, Ag, and mRNA decreased and were inhibited compared with those of PBMC cultured alone (p < 0.01). This inhibitory effect required direct cell contact between PMN and PBMC, and it was not due to a PMN mediated cytotoxicity. To confirm the role of ROS, H2O2 enhanced then inhibited the TF activity of PBMC in a dose-dependent manner, similarly to PMN. Thus, PMN may play an important role in the pathogenesis of thrombosis and atherosclerosis by exerting concentration-dependent regulatory effects on the TF production by PBMC via the release of ROS. PMID- 10725744 TI - Vav synergizes with protein kinase C theta to mediate IL-4 gene expression in response to CD28 costimulation in T cells. AB - The secretion of IL-4, which displays many important immunoregulatory functions, is restricted to cells of the Th2 subtype. In this study, we investigated the early signaling events leading to the activation of IL-4 transcription. Vav, the protein kinase C (PKC) isoform theta, and the adaptor protein SLP76 (SH2-domain containing leukocyte protein of 76 kDa), induced transcription from the IL-4 promoter. Vav and PKC theta synergistically activated human IL-4 promoter transcription and IL-4 mRNA production and were found to be constitutively associated in vivo. CD3/CD28-induced IL-4 transcription was inhibited upon coexpression of dominant negative forms of Vav, the adaptor proteins LAT (linker for activation of T cells) and SLP76, PKC theta, and components of the pathways leading to the activation of c-Jun N-terminal kinase (mitogen-activated protein kinase kinase 7 (MKK7), mixed lineage kinase 3 (MLK3)) and NF-kappa B (I kappa B kinase alpha and I kappa B kinase beta). The Vav/PKC theta-mediated synergistic activation of IL-4 transcription was not inhibited by cyclosporin A. Three independent experimental approaches revealed that Vav/PKC theta-derived signals selectively target the P1 and positive regulatory element (PRE)-I elements contained within the human IL-4 promoter. Vav/PKC theta strongly activated a luciferase reporter construct controlled by trimerized P1 or PRE-I elements and furthermore stimulated DNA binding of nuclear proteins to the P1 and PRE-I elements. Vav/PKC theta-induced transcription from the IL-4 promoter was almost completely abrogated by mutation of either the P1 or the PRE-I element within the entire IL-4 promoter. PMID- 10725745 TI - IL-11 activates human endothelial cells to resist immune-mediated injury. AB - IL-11, a gp130-signaling cytokine, is protective in several in vivo models of immune-mediated and inflammatory injury. HUVECs express IL-11 receptor alpha chain and gp130. Human IL-11 causes rapid (2-10 min) tyrosine phosphorylation of gp130. IL-11 at 0.1 and 10 ng/ml induces tyrosine phosphorylation of STAT3 and STAT1, respectively, although maximal responses require 50 ng/ml. Phospho-STAT3 and phospho-STAT1 levels peak rapidly (2.5 min) and disappear by 60 min. The p42 and p44 mitogen-activated protein kinases (MAPKs) are phosphorylated in response to 0.3 ng/ml IL-11 with maximal activation at 30 ng/ml IL-11. Phosphorylation of p42 and p44 MAPKs, which can be prevented by a mitogen-activated protein/extracellular signal-related kinase kinase-1 inhibitor, peaks by 15-20 min and largely disappears by 40 min. IL-11 does not activate NF-kappaB nor does it inhibit NF-kappaB activation by TNF. Similarly, IL-11 neither induces E selectin or ICAM-1 nor blocks induction by TNF. Although IL-11 does not alter class I MHC complex molecule expression, pretreatment with 0.5 ng/ml IL-11 partially protects HUVECs against lysis by allospecific class I MHC-restricted cytolytic T lymphocytes or by anti-class I MHC Ab plus heterologous C. IL-11 induced cytoprotection is protein synthesis dependent and may depend on mitogen activated protein/extracellular signal-related kinase kinase-1. Our results indicate that low (i.e., STAT3- and MAPK-activating) concentrations of IL-11 confer resistance to immune-mediated injury in cultured HUVECs without inhibiting proinflammatory responses. PMID- 10725746 TI - Migration of eosinophils across endothelial cell monolayers: interactions among IL-5, endothelial-activating cytokines, and C-C chemokines. AB - Eosinophils are the predominant cell type recruited in inflammatory reactions in response to allergen challenge. The mechanisms of selective eosinophil recruitment in allergic reactions are not fully elucidated. In this study, the ability of several C-C chemokines to induce transendothelial migration (TEM) of eosinophils in vitro was assessed. Eotaxin, eotaxin-2, monocyte chemotactic protein (MCP)-4, and RANTES induced eosinophil TEM across unstimulated human umbilical vein endothelial cells (HUVEC) in a concentration-dependent manner with the following rank order of potency: eotaxin approximately eotaxin-2 > MCP-4 approximately RANTES. The maximal response induced by eotaxin or eotaxin-2 exceeded that of RANTES or MCP-4. Preincubation of eosinophils with anti-CCR3 Ab (7B11) completely blocked eosinophil TEM induced by eotaxin, MCP-4, and RANTES. Activation of endothelial cells with IL-1beta or TNF-alpha induced concentration dependent migration of eosinophils, which was enhanced synergistically in the presence of eotaxin and RANTES. Anti-CCR3 also inhibited eotaxin-induced eosinophil TEM across TNF-alpha-stimulated HUVEC. The ability of eosinophil active cytokines to potentiate eosinophil TEM was assessed by investigating eotaxin or RANTES-induced eosinophil TEM across resting and IL-1beta-stimulated HUVEC in the presence or absence of IL-5. The results showed synergy between IL-5 and the chemokines but not between IL-5 and the endothelial activator IL-1beta. Our data suggest that eotaxin, eotaxin-2, MCP-4, and RANTES induce eosinophil TEM via CCR3 with varied potency and efficacy. Activation of HUVEC by IL-1beta or TNF alpha or priming of eosinophils by IL-5 both promote CCR3-dependent migration of eosinophils from the vasculature in conjunction with CCR3-active chemokines. PMID- 10725747 TI - An essential role of mast cells in the development of airway hyperresponsiveness in a murine asthma model. AB - Immunization of BALB/c mice with alum-adsorbed OVA, followed by three bronchoprovocations with aerosolized OVA, resulted in the development of airway hyperresponsiveness (AHR) and allergic inflammation in the lung accompanied by severe infiltration of eosinophils into airways. In this murine asthma model, administration of monoclonal anti-IL-5 Ab before each Ag challenge markedly inhibited airway eosinophilia, but the treatment did not affect the development of AHR. Immunization and aerosol challenges with OVA following the same protocol failed to induce AHR in the mast cell-deficient W/Wv mice, but induced AHR in their congenic littermates, i.e., WBB6F1 (+/+) mice. No significant difference was found between the W/Wv mice and +/+ mice with respect to the IgE and IgG1 anti-OVA Ab responses and to the airway eosinophilia after Ag provocations. It was also found that reconstitution of W/Wv mice with bone marrow-derived mast cells cultured from normal littermates restored the capacity of developing Ag induced AHR, indicating that lack of mast cells was responsible for the failure of W/Wv mice to develop Ag-induced AHR under the experimental conditions. However, the OVA-immunized W/Wv mice developed AHR by increasing the frequency and Ag dose of bronchoprovocations. The results suggested that AHR could be developed by two distinct cellular mechanisms. One would go through mast cell activation and the other is IgE/mast cell independent but an eosinophil/IL-5 dependent mechanism. PMID- 10725748 TI - Induction of functional IL-8 receptors by IL-4 and IL-13 in human monocytes. AB - IL-8 and related Glu-Leu-Arg (ELR+) CXC chemokines are potent chemoattractants for neutrophils but not for monocytes. IL-13 and IL-4 strongly increased CXCR1 and CXCR2 chemokine receptor expression in human monocytes, macrophages, and dendritic cells. The effect was receptor- and cell type-selective, in that CCRs were not increased and no augmentation was seen in neutrophils. The effect was rapid, starting at 4 h, and concentration dependent (EC50 = 6.2 and 8.3 ng/ml for CXCR1 and CXCR2, respectively) and caused by new transcriptional activity. IL 13/IL-4-treated monocytes showed increased CXCR1 and CXCR2 membrane expression. IL-8 and related ELR+ chemokines were potent and effective chemotactic agents for IL-13/IL-4-treated monocytes, but not for untreated mononuclear phagocytes, with activity comparable to that of reference monocyte attractants, such as MCP-1. In the same cells, IL-8 also caused superoxide release. Macrophages and dendritic cells present in biopsies from Omenn's syndrome and atopic dermatitis patients, two Th2 skewed pathologies, expressed IL-8 receptors by immunohistochemistry. These results show that IL-13 and IL-4 convert IL-8 and related ELR+ chemokines, prototypic neutrophil attractants, into monocyte chemotactic agonists, by up regulating receptor expression. Therefore, IL-8 and related chemokines may contribute to the accumulation and positioning of mononuclear phagocytes in Th2 dominated responses. PMID- 10725749 TI - G551D cystic fibrosis mice exhibit abnormal regulation of inflammation in lungs and macrophages. AB - The major cause of death in cystic fibrosis (CF) is chronic lung disease associated with persistent infection by the bacterium, Pseudomonas aeruginosa. S100A8, an S-100 calcium-binding protein with chemotactic activity, is constitutively expressed in the lungs and serum of CF patients. Levels of S100A8 mRNA were found to be three to four times higher in the lungs of mice carrying the G551D mutation in CF transmembrane conductance regulator compared with littermate controls. Intravenous injection of bacterial LPS induced S100A8 mRNA in the lung to a greater extent in G551D mice than in wild-type littermates. Localization of S100A8 mRNA and protein in the lung indicate that it is a marker for neutrophil accumulation. Bone marrow-derived macrophages from G551D mice were shown to also exhibit hypersensitivity to LPS, measured by induction of TNF alpha. These results provide evidence that the pathology of CF relates to abnormal regulation of the immune system. PMID- 10725750 TI - Alveolar macrophages bind and phagocytose allergen-containing pollen starch granules via C-type lectin and integrin receptors: implications for airway inflammatory disease. AB - Recent studies suggest that IgE-independent mechanisms of airway inflammation contribute significantly to the pathophysiology of allergic airway inflammatory diseases such as asthma. Such mechanisms may involve direct interactions between inhaled allergens and cells of the respiratory tract such as macrophages, dendritic cells, and epithelial cells. In this study, we investigated receptor mediated interactions occurring between alveolar macrophages and allergen containing pollen starch granules (PSG). We report here that PSG are released from a range of grass species and are rapidly bound and phagocytosed by alveolar macrophages. Human monocyte-derived dendritic cells also bound PSG but no internalization was observed. Phagocytosis of PSG was dependent on Mg2+ and Ca2+ and was inhibited by neo-glycoproteins such as galactose-BSA and N acetylgalactose-BSA. Partial inhibition of phagocytosis was also seen with the Arg-Gly-Asp-Ser (RGDS) motif and with an anti-CD18 mAb (OX42). The combination of both neo-glycoprotein and anti-CD18 achieved the greatest degree of inhibition (>90%). Together, these data suggest a role for both C-type lectins and beta2 integrins in the binding and internalization of PSG. The consequences of this interaction included a rapid up-regulation of inducible NO synthase mRNA and subsequent release of NO by alveolar macrophages. Thus, receptor-mediated recognition of inhaled allergenic particles by alveolar macrophages may represent a potential mechanism for modulating the inflammatory response associated with allergic airway diseases such as asthma. PMID- 10725751 TI - Granulocyte-macrophage colony-stimulating factor as an autocrine survival factor for mature normal and malignant B lymphocytes. AB - The role of GM-CSF in B cell (patho)physiology is unclear. Although B cells can respond to GM-CSF, there is controversy concerning the extent to which various resting and activated B cell types can themselves produce this cytokine, and the possibility that it can function in an autocrine fashion has not previously been considered. The aim of the present study was to address these issues using hairy cells (HCs) and chronic lymphocytic leukemia cells, two intrinsically activated mature malignant B cell types (with activation being more uniform and more pronounced in HCs). Normal B cells were used for comparison. Using a number of techniques, we demonstrated the constitutive production of GM-CSF by all three cell types and showed that the cytokine was biologically active. GM-CSF mRNA and protein were increased after cell activation by PMA, and constitutive production of the cytokine was highest in HCs, suggesting that the level of GM-CSF production is influenced by cell activation. Because GM-CSF is known to be antiapoptotic for myeloid cells, we used blocking anti-GM-CSF Abs to examine the contribution of autocrinely produced cytokine to cell survival. The Abs produced marked reduction in the in vitro survival of HCs, chronic lymphocytic leukemia cells, and normal B cells by promoting apoptosis. Taken together, these findings suggest that, in combination with other known rescue factors, autocrinely produced GM-CSF may contribute to normal and malignant B cell survival in vivo. PMID- 10725753 TI - Tumor-specific CD4+ T lymphocytes from cancer patients are required for optimal induction of cytotoxic T cells against the autologous tumor. AB - This study focuses on the specific CD4+ T cell requirement for optimal induction of cytotoxicity against MHC class II negative autologous tumors (AuTu) collected from patients with various types of cancer at advanced stages. CD4+ T cells were induced in cultures of cancer patients' malignant effusion-associated mononuclear cells with irradiated AuTu (mixed lymphocyte tumor cultures (MLTC)) in the presence of recombinant IL-2 and recombinant IL-7. Tumor-specific CD4+ T cells did not directly recognize the AuTu cells, but there was an MHC class II restricted cross-priming by autologous dendritic cells (DCs), used as APC. CD8+ CTL, also induced during the MLTC, lysed specifically AuTu cells or DCs pulsed with AuTu peptide extracts (acid wash extracts (AWE)) in an MHC class I restricted manner. Removal of CD4+ T cells or DCs from the MLTC drastically reduced the CD8+ CTL-mediated cytotoxic response against the AuTu. AWE-pulsed DCs preincubated with autologous CD4+ T cells were able, in the absence of CD4+ T cells, to stimulate CD8+ T cells to lyse autologous tumor targets. Such activated CD8+ T cells produced IL-2, IFN-gamma, TNF-alpha, and GM-CSF. The process of the activation of AWE-pulsed DCs by CD4+ T cells could be inhibited with anti-CD40 ligand mAb. Moreover, the role of CD4+ T cells in activating AWE-pulsed DCs was undertaken by anti-CD40 mAb. Our data demonstrate for the first time in patients with metastatic cancer the essential role of CD4+ Th cell-activated DCs for optimal CD8+ T cell-mediated killing of autologous tumors and provide the basis for the design of novel protocols in cellular adoptive immunotherapy of cancer, utilizing synthetic peptides capable of inducing T cell help in vivo. PMID- 10725752 TI - Activation of the IL-4 STAT pathway in rheumatoid synovium. AB - STATs act as second messenger after binding of a signaling molecule to its receptor. IL-4 STAT is directly involved in the IL-4-dependent gene transcription in the nucleus. We examined the expression and activation of IL-4 STAT and its related kinase Jak-1 in rheumatoid synovium. Rheumatoid arthritis (RA) synovial frozen sections of patients with short-term (<1 year) and long-term disease (>2 years) were examined using in situ hybridization and immunohistochemistry. IL-4 STAT mRNA could be detected in synovium of patients with short-term and long-term RA. The most intensive expression of IL-4 STAT mRNA could be seen in follicular inflammatory infiltrates. In the synovial lining, both fibroblasts and macrophages expressed IL-4 STAT mRNA. IL-4 STAT and Jak-1 protein was expressed by synoviocytes, and up-regulation could be induced after stimulation with IL-4. Activation of IL-4 STAT was reflected by phosphorylation of IL-4 STAT. The results indicate that IL-4 STAT is involved in key pathomechanisms in RA synovium and that IL-4 STAT-dependent pathways operate in early and late stages of the disease and presumably contribute to inhibitory immune mechanisms in RA synovium. PMID- 10725754 TI - Identification of a CD8 T cell that can independently mediate autoimmune diabetes development in the complete absence of CD4 T cell helper functions. AB - Previous work has indicated that an important component for the initiation of autoimmune insulin-dependent diabetes mellitus (IDDM) in the NOD mouse model entails MHC class I-restricted CD8 T cell responses against pancreatic beta cell Ags. However, unless previously activated in vitro, such CD8 T cells have previously been thought to require helper functions provided by MHC class II restricted CD4 T cells to exert their full diabetogenic effects. In this study, we show that IDDM development is greatly accelerated in a stock of NOD mice expressing TCR transgenes derived from a MHC class I-restricted CD8 T cell clone (designated AI4) previously found to contribute to the earliest preclinical stages of pancreatic beta cell destruction. Importantly, these TCR transgenic NOD mice (designated NOD.AI4alphabeta Tg) continued to develop IDDM at a greatly accelerated rate when residual CD4 helper T cells were eliminated by introduction of the scid mutation or a functionally inactivated CD4 allele. In a previously described stock of NOD mice expressing TCR transgenes derived from another MHC class I-restricted beta cell autoreactive T cell clone, IDDM development was retarded by elimination of residual CD4 T cells. Hence, there is variability in the helper dependence of CD8 T cells contributing to the development of autoimmune IDDM. The AI4 clonotype represents the first CD8 T cell with a demonstrated ability to progress from a naive to functionally activated state and rapidly mediate autoimmune IDDM development in the complete absence of CD4 T cell helper functions. PMID- 10725755 TI - Protection of nonobese diabetic mice from diabetes by gene(s) closely linked to IFN-gamma receptor loci. AB - Nonobese diabetic (NOD) mice carrying a segment of chromosome flanking the disrupted IFN-gamma receptor gene from original 129 ES cells are resistant to development of diabetes. However, extended backcrossing of this mouse line to the NOD mouse resulted in a segregation of the IFN-gammaR-deficient genotype from the diabetes-resistant phenotype. These results indicate that the protection of NOD mice from the development of diabetes is not directly linked to the defective IFN gamma receptor gene but, rather, is influenced by the presence of a diabetes resistant gene(s) closely linked to the IFN-gammaR loci derived from the 129 mouse strain. PMID- 10725756 TI - Reduced chemokine and chemokine receptor expression in spinal cords of TCR BV8S2 transgenic mice protected against experimental autoimmune encephalomyelitis with BV8S2 protein. AB - The perivascular transmigration and accumulation of macrophages and T lymphocytes in the CNS of mice with experimental autoimmune encephalomyelitis (EAE) may be partly regulated by low m.w. chemotactic cytokines. Using the RNase protection assay and ELISA, we quantified expression of chemokines and chemokine receptors in the spinal cord (SC), brain, and lymph nodes of BV8S2 transgenic mice that developed or were protected from EAE by vaccination with BV8S2 protein. In paralyzed control mice, the SC had increased cellular infiltration and strong expression of the chemokines RANTES, IFN-inducible 10-kDa protein, and monocyte chemoattractant protein-1 and the cognate chemokine receptors CCR1, CCR2, and CCR5, with lower expression of macrophage-inflammatory protein (MIP)-1alpha, MIP 1beta, and MIP-2; whereas brain had less infiltration and a lower expression of a different pattern of chemokines and receptors. In TCR-protected mice, there was a decrease in the number of inflammatory cells in both SC and brain. In SC, the reduced cellular infiltrate afforded by TCR vaccination was commensurate with profoundly reduced expression of chemokines and their cognate chemokine receptors. In brain, however, TCR vaccination did not produce significant changes in chemokine expression but resulted in an increased expression of CCR3 and CCR4 usually associated with Th2 cells. In contrast to CNS, lymph nodes of protected mice had a significant increase in expression of MIP-2 and MIP-1beta but no change in expression of chemokine receptors. These results demonstrate that TCR vaccination results in selective reduction of inflammatory chemokines and chemokine receptors in SC, the target organ most affected during EAE. PMID- 10725757 TI - Flow cytometric quantification of phagocytosis in acute myeloid leukemia. AB - Phagocytosis was studied by flow cytometry (FCM) in 15 patients with acute myeloid leukemia (AML). The pattern of phagocytosis differed markedly between AML and controls. The percentage of phagocytosing AML leukocytes was below that of the controls (p < 0.01). The phagocytic capacity of a subpopulation of leukemic cells was diminished, but compensated by the phagocytosis of a few prey by each of many immature leukocytes. Phagocytosis by immature and mature AML leukocytes was receptor dependent, and both carried functional complement receptors. Attachment to the cell surface was not rate-limiting in phagocytosing AML leukocytes. PMID- 10725758 TI - Embryonic stem cells release potentially novel hematopoietic factors. AB - We have observed that a severe decrease in the number of erythroid progenitor cells follows the long-term culture (LTC) of human primitive stem cells on an established mouse fibroblast feeder layer. This suggests that one, or several, factors necessary to support erythroid differentiation might be missing in these culture conditions. To improve the erythroid differentiation and stem cell output of LTCs, we have examined the hypothesis that the factors that regulate pluripotent stem cell proliferation and erythroid commitment can be found in media conditioned by embryonic stem (ES) cells. We have found that media conditioned by undifferentiated and differentiating ES cells can affect differentiation patterns in both short-term culture and LTC assays. Medium conditioned by undifferentiated ES cells increases the numbers of estimated LTC initiating cells (est.LTC-IC) and potentiates granulocytic differentiation. In contrast, medium conditioned by ES cells undergoing differentiation increases the number of est.LTC-IC and is a powerful promoter of erythroid differentiation. In the presence of this medium, the number of erythroid progenitors generated after 5 weeks of LTC was increased up to 100-fold as compared with controls, and the number of est.LTC-IC was amplified up to 140-fold. These results offer a new approach for the identification of factors implicated in stem cell proliferation, self-renewal and commitment. In addition, the improved LTC conditions for the expansion of stem cells without reduction of their in vitro differentiation potential should be useful for a wide range of applications. PMID- 10725759 TI - Effect of training on the phagocytic capacity of peritoneal macrophages from rats exposed to swim stress. AB - Since strenuous effort may affect the immune system, a study was designed to examine the impact of the progressive training of rats exposed to swim stress. Rats (trained swimmers) were forced to swim daily in a water bath for progressively longer periods. At the end of the study, which continued for a total of 6 weeks, the superoxide anion generation and phagocytic capacity of peritoneal macrophages, the mitogen response of splenic cells (splenocytes) and the serum corticosterone level were examined. The results, compared to those of animals taken in and out of their cages (nonswimmers), showed an increase in superoxide anion generation, as well as a decrease in both the percentage of phagocytosing cells and the number of particles internalized by each individual cell. In trained swimmers, the mitogen response to phytohemagglutinin and concanavalin A (Con A), as well as the corticosterone level, did not change significantly from those of nonswimmers. A third group of animals were forced to swim once only for 30 min, until the appearance of signs of marked fatigue (acute swimmers). Their peritoneal macrophages showed an increased superoxide anion generation and a significantly decreased response to Con A compared to those of the nonswimmers. The serum corticosterone level in acute swimmers was found to be increased compared to that of an additional group of animals kept at complete rest. The findings indicate that the progressive training of rats exposed to swim stress does not prevent alterations in certain immune responses, a fact that should be considered by intensive exercisers. PMID- 10725760 TI - Calcification in untreated non-Hodgkin's lymphoma of the jejunum. AB - A 69-year-old Japanese female was admitted to our hospital due to a 2-month history of vomiting after eating. Examination of the small intestinal tract revealed a tumor with calcification in the inner portion, from the horizontal portion to the ascending portion of the duodenum, and jejunojejunostomy was performed. The pathological findings of the tumor gave a diagnosis of non Hodgkin's lymphoma, diffuse small cleaved cell (Working Formulation classification), B cell type, of the jejunum. Calcification is rarely found in untreated malignant lymphoma and 15 cases of untreated malignant lymphoma with calcification have been reviewed. PMID- 10725761 TI - CD8 expression in a case of chronic lymphocytic leukemia with trisomy 12. AB - We report a case of B-cell chronic lymphocytic leukemia (B-CLL) with aberrant expression of the T-cell-associated antigen CD8, as revealed by two-color flow cytometric analysis. DNA studies showed immunoglobulin heavy-chain gene rearrangement, but not of gamma-chain T-cell receptor, confirming the B-cell origin of the neoplastic cells. Ploidy analysis showed a tetraploid population and high S-phase fraction. B-CLL cells also carried trisomy 12, detected by fluorescence in situ hybridization. The identification of more cases with the same features would be necessary to establish the prognosis of this subtype of B CLL. PMID- 10725762 TI - Immunophenotype of peripheral blood mononuclear cells and NK cell activity after allogeneic bone marrow transplantation using recombinant human granulocyte colony stimulating factor. PMID- 10725763 TI - Methylenetetrahydrofolate-dehydrogenase 1958 G-A (R653 Q) polymorphism in Turkish patients with venous thromboembolism. PMID- 10725764 TI - Neutrophil alkaline phosphatase activity in Turner syndrome. PMID- 10725765 TI - Molecular characterization of hemoglobins Kurosaki [alpha7 Lys-->Glu], G-Pest [alpha74 Asp-->Asn], Stanleyville-II [alpha78 Asn-->Lys] and J-Rovigo [alpha53 Ala-->Asp]. PMID- 10725766 TI - Home parenteral nutrition: clinical and laboratory analysis of initial experience (1994-1997). Implications for patient management. AB - BACKGROUND/AIM AND METHOD: Severe malabsorption often necessitates prolonged parenteral nutrition. Home parenteral nutrition (HPN) offers the opportunity for treatment at home. We report clinical and laboratory data of initial 27 HPN patients of one center since its opening in 1994. RESULTS: Clinical and biological markers of nutritional status were normalized and well maintained in most patients. Except for vitamin E and selenium (lower in HPN patients), the other vitamin and micronutrient levels were normal. There was no obvious essential fatty acid deficiency. Cholestasis was usual, but only 1 patient had a severe hepatic disease. Catheter infection occurred 18 times in 13 patients, but the frequency decreased with time (from 4.2 to 1.7 infections/1,000 days on HPN). No patient died from HPN complications. Social rehabilitation and, in some patients, full professional rehabilitation were constant. CONCLUSIONS: These data confirm that HPN performed in centers with expertise allowed patients to overcome gut failure and to recover subnormal or normal nutritional status. Satisfactory social rehabilitation was obtained in all patients. HPN complications were rarely life-threatening, and their frequency decreased with experience. PMID- 10725767 TI - Oxidation of an oil rich in docosahexaenoic acid compared to linoleic acid in lactating women. AB - BACKGROUND: We studied the oxidation of an oil rich in docosahexaenoic acid (DHA; DHASCO((R))) in lactating mothers receiving a dietary DHA supplement or a placebo. The results were compared with the oxidation of linoleic acid. METHODS: Breast-feeding mothers received a dietary supplement (DHASCO; 200 mg DHA/day, n = 5) or a placebo (n = 5) for 14 days. Six weeks post partum all 10 mothers received a single dose of 2 mg/kg body weight uniformly (13)C-labeled DHASCO. In a previously reported study 6 mothers received 1 mg/kg body weight uniformly (13)C-labeled linoleic acid. Breath samples were collected over 48 h after tracer application. The total CO(2) production was measured by indirect calorimetry and the (13)C isotopic enrichment of labeled CO(2) by isotopic ratio mass spectrometry. RESULTS: The oxidation of (13)C-labeled DHASCO in the supplemented and placebo groups was similar. Maximal (13)C enrichment was reached earlier in the group receiving (13)C-DHASCO (median 1.0 vs. 3.0 h in the linoleic acid group). The cumulative (13)C recovery in breath was higher in the DHASCO versus the linoleic acid group until 10 h after tracer application and comparable thereafter. CONCLUSIONS: The difference in oxidation of DHASCO versus linoleic acid after tracer ingestion might be partly due to a faster absorption and oxidation of shorter chain saturated fatty acids contained in DHASCO. The cumulative oxidation of DHASCO and linoleic acid 24 and 48 h after tracer ingestion is similar. PMID- 10725768 TI - Australian food sources and intakes of omega-6 and omega-3 polyunsaturated fatty acids. AB - BACKGROUND/AIMS: Both omega-6 and omega-3 polyunsaturated fatty acids (PUFAs) are recognised as essential nutrients in the human diet, yet we have little information on the extent to which different food sources contribute to their intake. The aim of the present study was to ascertain the daily intakes and food sources of omega-6 and omega-3 PUFAs in our local community. METHODS: Three-day food records were obtained from 83 healthy adults living in the Illawarra region of New South Wales. The PUFA composition of the foods which they consumed was derived from food composition tables and recently published food analysis data. RESULTS: Polyunsaturated margarine, nuts/seeds, bread, snacks/desserts and takeaway foods were important sources of omega-6 PUFAs, while canola oil and margarine, takeaway foods, snacks/desserts and bread were sources of alpha linolenic acid (LNA), an omega-3 PUFA. As expected, fish was the main source of the very long chain (VLC) omega-3 PUFAs, i.e. eicosapentaenoic acid (EPA), docosapentaenoic acid (DPA) and docosahexaenoic acid (DHA), to which significant health benefits are attributed. An unexpected finding, however, was that, due to the large amount eaten, meat was also a major contributor (29%) to the dietary intake of VLC omega-3 PUFAs. Median intakes of omega-6 and omega-3 PUFAs were 9.9 and 1.2 g/day, respectively, resulting in a dietary omega-6:omega-3 ratio of 8:1. The median intake of VLC omega-3 PUFA was 0.18 g/day. CONCLUSION: We have identified food sources and intakes of PUFAs for an Australian subpopulation differentiating between omega-6 and omega-3 PUFAs. Whilst canola and fish were the primary sources of LNA and VLC omega-3 PUFAs respectively, we found that meat made a significant contribution to VLC omega-3 PUFA intake. PMID- 10725769 TI - Addition of inulin to breakfast does not acutely affect serum ionized calcium and parathyroid hormone concentrations. AB - BACKGROUND/AIMS: The aim of the present study was to investigate the effects of inulin on calcium metabolism. The study consisted of two separate parts both of which had a randomized two-period cross-over design. METHODS: Fifteen young healthy women volunteered to participate in this study. During the first part of the study, cheese containing 210 mg of calcium, either with 15 g of inulin or without any inulin, was ingested at breakfast, and in the second part, 210 mg of calcium as a supplement, either with 15 g inulin or without inulin, was ingested. The whole day's diet was standardized. Before breakfast, and 2, 4, 6 and 8 h after breakfast, a blood sample was taken, and intact parathyroid hormone (iPTH), ionized calcium (iCa) and total calcium were measured. Urine was collected throughout the day, and the 8-, 12- and 24-hour calcium excretion was calculated. RESULTS: The iPTH or iCa concentration curves (AUCs) did not differ over 8 h, whether or not inulin was consumed at breakfast. The postload urinary calcium excretion was not affected by the inulin. CONCLUSION: Fifteen grams of inulin in fresh cheese or with a calcium supplement (210 mg Ca) taken at breakfast does not acutely affect the markers of calcium metabolism as opposed to a corresponding breakfast without inulin. PMID- 10725770 TI - Post-natal modulation of heart and liver phosphoglyceride fatty acids in pups. AB - BACKGROUND: Maternal dietary fats alter tissue fatty acids of the fetus and suckling pups. However, the possible change in tissue composition in response to the high oxygen tension extrauterine milieu independent of diet is not well understood. METHODS: We have compared the fatty acids of heart and liver choline (CPG) and ethanolamine (EPG) phosphoglycerides of rat offspring at birth and post natal day 15. Sprague-Dawley rats were fed a breeding diet prior to mating, pregnancy and lactation. A proportion of each litter was sacrificed and the liver and heart were obtained for analysis. Changes in fatty acid composition specific to tissue (heart and liver) and phosphoglyceride (CPG and EPG) occurred post natally. RESULTS: Relative levels of palmitate and oleate decreased and those of stearate increased in both the heart and liver phosphoglycerides of the suckling pups. There was a reduction in arachidonate/linoleate ratio primarily due to the increase in linoleic acid. With the exception in the heart EPG, the levels of arachidonic acid did not decrease concomitantly. Although the fatty acid composition of the diet did not change between pregnancy and lactation, docosahexaenoic and total n-3 increased in heart CPG and EPG and liver CPG of the suckling pups. Evidently, membrane fatty acid modulation, independent of maternal dietary fat, occurs in the extrauterine environment. It seems to favour the accretion of linoleic, arachidonic, docosahexaenoic and total n-3 fatty acids. CONCLUSION: Since there appears to be some parallel between the very preterm human neonate and rat pups with regard to nutrient store at birth and the neonatal developmental time window, our results may have relevance for the understanding of fatty acid metabolism and turnover in the human neonate. PMID- 10725771 TI - Low-density lipoprotein size in primary hypothyroidism. Effects of hormone replacement therapy. AB - BACKGROUND/AIM: Hypothyroidism is associated with abnormalities in lipid metabolism but its effect on LDL size is not known. This study identified the LDL particle size (pattern 'A' or 'B') in a group of 50 postmenopausal women with primary hypothyroidism before and after 2 months of hormone replacement therapy (HRT) with the aim of establishing whether hypothyroidism is associated with an increased frequency of pattern B LDL compared with healthy controls, and whether euthyroid recovery modified LDL size. METHODS: Lipid parameters (total cholesterol, triglycerides, HDL- and LDL-cholesterol, apoprotein AI and B and lipoprotein(a)) were determined in a blood sample from each patient and control. LDL size was determined by gradient gel electrophoresis. Determinations were done before and after 2 months of HRT (75-150 microg L-tiroxina/daily) in patients and once in controls. RESULTS/CONCLUSIONS: No significant difference emerged in pattern B LDL distribution: 16% in patients and 18% in controls (p = NS). After HRT no statistical variations were observed in LDL size. HRT normalized all other lipid parameters except lipoprotein(a). In conclusion, the increased risk of coronary heart disease assumed to be associated with hypothyroidism is not linked with the presence of pattern B LDL, but rather with concomitant metabolic abnormalities. PMID- 10725772 TI - Linoleic acid - the fall of an angel? PMID- 10725773 TI - Effect of short-term testosterone treatment on leptin concentrations in boys with pubertal delay. AB - Testosterone administration increases growth hormone (GH) secretion and decreases the plasma leptin concentration in men. We evaluated the effect of increased GH secretion due to short-term testosterone treatment on leptin concentrations. Ten boys aged 14.8 +/- 0.2 (mean +/- SE) years with transient GH deficiency caused by pubertal delay were evaluated before and after (3 months) 4 intramuscular injections of 100 mg testosterone heptylate, given at 15-day intervals. The leptin concentration decreased from 5.4 +/- 1.3 to 3. 6 +/- 1.1 microgram/l (p < 0.001), despite a weight gain of 3.4 +/- 0.5 kg. There were significant increases in body mass index (BMI), from -0.2 +/- 0.5 to 0.2 +/- 0.5 SD, p < 0.005, in GH peak after stimulation test, from 6.3 +/- 0.5 to 21.7 +/- 2.9 microgram/l, p < 0. 0003, in plasma testosterone, from 0.6 +/- 0.1 to 6.5 +/- 1.3 microgram/l, p < 0.001, in insulin-like growth factor-I (IGF-I), from 152 +/- 21 to 330 +/- 30 microgram/l, p < 0.0001, and in IGF-binding protein-3 (IGFBP-3), from 4.2 +/- 0.5 to 5.4 +/- 0.4 mg/l, p < 0.01. But there were no changes in blood glucose (4.7 +/ 0.1 and 4.8 +/- 0.1 mmol/l), or plasma fasting insulin (9.0 +/- 1.2 and 8.1 +/- 1.3 mIU/l). The leptin concentrations were positively correlated with the BMI before (p < 0.03) and after (p < 0.04) testosterone, but not with the GH peak after stimulation, or with plasma testosterone, IGF-I or IGFBP-3. The leptin and insulin concentrations after testosterone treatment were positively correlated (p < 0.04). Thus, short-term testosterone treatment of boys with pubertal delay decreases their leptin concentrations. The lack of correlation with GH secretion or with its changes, despite the dramatic increase in GH secretion, and the lack of change in insulin are additional features suggesting that testosterone increases the leptin concentration mainly by an effect on adipose tissue. PMID- 10725774 TI - Thyroid autoimmunity in children and adolescents with type 1 diabetes mellitus. Effect of age, gender and HLA type. AB - Type 1 diabetes is often associated with additional autoimmune phenomena. However, data reported on the frequency of thyroid autoimmunity differ vastly. Therefore, the prevalence of thyroid autoantibodies was evaluated at a large pediatric diabetes center in Southern Germany. 2,305 determinations (TPO and TG, ELISA) were performed in 495 patients with type 1 diabetes (234 boys, 261 girls; age at last measurement: 15.4 +/- 0.3 years, duration of diabetes 7. 5 +/- 0.2 years). The prevalence of elevated thyroid antibodies increased dramatically with age: from 3.7% in patients less than 5 years of age up to 25.3% in the age group 15-20 years (p < 0.0001). For children older than 10 years, girls were significantly more affected than boys (p < 0.0001). Thyroid autoimmunity tended to be more prevalent in the subgroup of patients with the HLA type DR3/DR4 compared to patients with other HLA types (p = 0.08). In children older than 10 years, basal TSH concentrations were significantly elevated in antibody-positive patients (p < 0.05). In conclusion, thyroid autoimmunity is prevalent in children and adolescents with type 1 diabetes. Adolescent girls and young women are especially affected. Yearly routine determinations of thyroid antibodies are therefore recommended. PMID- 10725776 TI - Bone maturation reflects the secular trend in growth. AB - The aim of this study was to compare a series of X-rays from the mid-1990s with another taken in the mid-1980s in order to test the possibility that environmental causes affect the skeletal maturation. The first group of subjects included a total of 1,057 girls and 1, 055 boys participating in a project for Japan and China health research in 1986. The second group of subjects included a total of 382 girls and 629 boys participating in a project for bone mineral density research in 1996. The skeletal maturity score using the Tanner-Whitehouse 2-RUS method was used as the fundamental datum. This score was used to represent each group. The Wilcoxon's rank sum test was applied to examine the significance of the difference between the 1986 and the 1996 groups. The 1996 children had not matured more than the 1986 children; children in both groups reached the given scores at almost the same ages. In girls, there was little difference between the groups at 7 years of age, but it declined from 8 years of age onward. Some apparent differences arose at ages 14 and 15, but ceased by age 16 in girls. In boys, no differences were found in those aged from 7 to 17 years, except for 12 year-olds. We did not detect much of a difference in bone maturation between the 1986 and 1996 groups of children, and no differences in height during the same period. Our findings suggest that bone maturation reflects the secular trend in growth. PMID- 10725775 TI - Autoantibodies predicting diabetes mellitus type I in celiac disease. AB - Celiac disease (CD) and diabetes mellitus type I (DM-I) are both autoimmune diseases. Abnormal first-phase insulin response (FPIR) is associated with the prediabetic phase. Glutamic acid decarboxylase (GAD) and islet cell antibodies (ICAs) - especially the tyrosine phosphatase-like protein IA-2 antibodies - are considered to be serological markers of DM-I future development. The aim of this study is to investigate the presence of autoantibodies (GAD, IA-2) in individuals with CD, on a gluten-free diet, who have normal intestinal morphology. Thirty patients with CD (4-22, mean 15 years), 30 newly diagnosed diabetic children (2.5 16, mean 10 years) and 30 healthy subjects (7-35, mean 18 years) were investigated. Serum GAD and IA-2 autoantibodies were assessed by a quantitative enzyme-linked immunosorbent assay (ELISA) method in all patients and controls. Seven CD patients (23%), 28 diabetic children (93%) and none in the control group had positive GAD and IA-2 antibodies. The FPIR was normal in CD patients (>/=46 mU/l). CONCLUSIONS: GAD and IA-2 antibodies are detected in 23% of patients with CD. These patients may be at risk to develop DM-I. Regular follow-up and determination of FPIR for the early diagnosis of the prediabetic phase in patients with CD having circulating autoantibodies is recommended. PMID- 10725777 TI - Serum osteocalcin and insulin-like growth factor I levels in children with congenital adrenal hyperplasia. AB - Patients with the virilizing forms of congenital adrenal hyperplasia (CAH) need a life-long glucocorticoid replacement therapy and also an additional mineralocorticoid replacement in cases with the salt-wasting form of the disease. Glucocorticoids are reported to decrease the serum osteocalcin levels and to inhibit the effects of insulin-like growth factor I (IGF-I). To collect data on the age related patterns of osteocalcin and IGF-I production in patients with CAH, measurements of these compounds have been carried out in a considerably large sample of treated CAH patients and control subjects in childhood and adolescence. Data of 62 patients between 0. 3-19 years of age were compared to the data of 188 control children. Osteocalcin and IGF-I were determined by radioimmunoassay. A lower than normal level of serum osteocalcin was found in both male and female patients at chronological ages above 11.6 and 9.6 years, respectively. Furthermore, no pubertal osteocalcin peak could be seen when data were evaluated according to the bone age. Serum IGF-I levels were higher in male CAH patients at the chronological age of 0.3-15.5 years and in female patients at the chronological age of 4. 6-9.5 years. In pubertal years serum IGF-I concentrations were lower in CAH patients when data were evaluated according to the bone age. We conclude that serum osteocalcin is decreased during and after puberty in CAH patients on replacement doses of glucocorticoids. Normal to elevated serum levels of IGF-I in treated CAH cases suggest that the shorter final height of these patients may not be due to the decreased activity in the growth hormoneIGF-I axis, but rather to the advanced bone maturation and the premature epiphyseal fusion. PMID- 10725778 TI - Nociceptin stimulates thyrotropin secretion in rats. AB - Effects of nociceptin on thyrotropin (TSH) and thyrotropin-releasing hormone (TRH) secretion in rats were studied. Nociceptin (150 microgram/kg) was injected intravenously and rats were serially decapitated after the injection. The effects of nociceptin on TRH release from the hypothalamus and TSH release from the anterior pituitary in vitro were also investigated. TRH and thyroid hormones were measured by individual radioimmunoassays. TSH was determined by enzyme immunoassay. TRH contents in the hypothalamus decreased significantly after nociceptin injection, whereas plasma TRH concentrations showed no changes. Plasma TSH concentrations increased significantly in a dose-related manner. The TRH release from the hypothalamus was enhanced significantly in a dose-related manner with the addition of nociceptin. The TSH release from the anterior pituitary in vitro was not affected by the addition of nociceptin. The plasma thyroxine and 3,3',5-triiodothyronine levels did not change significantly after nociceptin administration. The inactivation of TRH by plasma or hypothalamus in vitro after nociceptin injection did not differ from that of controls. The findings suggest that nociceptin acts on the hypothalamus to stimulate TRH and TSH secretion. PMID- 10725779 TI - Prognosis of children with malignant pheochromocytoma. Report of 2 cases and review of the literature. AB - Malignant pheochromocytomas are rare in childhood and the prognosis of children with this tumor is not well known. We present 2 pediatric observations of malignant pelvic pheochromocytoma. Symptoms in both cases were headache and hypertension. The tumor invaded the sacral bone. Angiogram helped to localize the tumor and metastases, and allowed preoperative embolization of the tumor in 1 case. The first child underwent incomplete surgical resection, (131)I-MIBG therapy and radiotherapy, and is still alive 2 years after diagnosis. The second child died from metastatic invasion a few weeks after discovery of the tumor. We reviewed previous reports of children with malignant pheochromocytomas (30 cases). Primary tumor was extraadrenal in 50% of cases. The 3-year survival rate was 73 +/- 9% (mean +/- SD). Apart from surgical resection, no particular treatment appeared to be more effective than others in reducing mortality. PMID- 10725780 TI - Should we treat children with idiopathic short stature? AB - The use of growth hormone (GH) to treat short children who are clearly GH deficient is now well accepted. However, GH treatment of short children who have no currently recognizable abnormalities in their GH-insulin-like growth factor I axis remains controversial. Whether such children with so-called idiopathic short stature (ISS) should be treated with GH was the subject of an international workshop held in St.-Paul-de-Vence, France, in April 1999. This article summarizes the issues discussed at the workshop, including the definition of ISS, ethical and health-economic aspects of treatment, results from clinical trials and surveillance studies, and the use of prediction models in aiding treatment decisions. PMID- 10725781 TI - Aspects of ovarian follicle development throughout life. AB - The pool of primordial follicles present in the female ovary reaches its maximum number around 20 weeks of gestational age and then decreases in a logarithmic fashion throughout life until complete depletion occurs around the age of the menopause. Reproductive life is initiated when less than 10% (0.5 million) of primordial follicles are left. The entire growth trajectory of the follicle takes at least 3 months. Follicle growth up to the antral stage occurs during fetal life and infancy. While the role of gonadotropins in early follicular development remains controversial, the last 2 weeks of development are FSH dependent. The intercycle rise in FSH and decreasing levels thereafter are crucial for recruitment of a cohort of healthy, early antral follicles and subsequent single dominant selection. Following puberty, anovulation may persist for years and this may presage the development of adult anovulatory infertility. The menopause is preceded by a period of reduced fertility. The development of reliable and sensitive markers for ovarian ageing will be the challenge of the near future. PMID- 10725782 TI - Endocrine characteristics of human breast epithelial cells, MCF-10F. AB - MCF-10F is a spontaneously immortalized nontransformed human breast epithelial cell line which does not grow in soft agar or form tumors in nude mice. Though the presence of estrogen receptors has not been found in these cells, they can metabolize estradiol very efficiently. The present study describes the endocrine characteristics of this cell line with respect to growth response to estradiol and its metabolites, estradiol metabolism and aromatase activity. MCF-10F cells were growth stimulated by 16alpha-hydroxyestrone and estriol, whereas, estradiol and other estradiol metabolites did not affect cell proliferation. The constitutive level of 16alpha-hydroxyestrone, a metabolite of estradiol biotransformation that has been associated with enhanced carcinogenesis in several animal, cell and tissue culture models, was a hundredfold higher in the non-transformed MCF-10F cells than in the transformed MCF-7 cells. Treatment with the carcinogen, dimethylbenz(a)anthracene (DMBA), however, did not upregulate 16alpha-hydroxylation as was observed in transformed MCF-7 cells. MCF-10F cells also had no detectable aromatase activity though the level of 17-oxidation was unusually high as compared with MCF-7 cells. Our results using the non transformed MCF-10F cells as a model system suggests that the presence of high level of 16alpha-hydroxyestrone, a metabolite previously shown to be associated with malignant phenotype, may not be sufficient for breast cancer transformation. PMID- 10725783 TI - Secretion of noncomplexed 'Big' (10-18 kD) forms of insulin-like growth factor-II by 12 soft tissue sarcoma cell lines. AB - The paraneoplastic production of pro-insulin-like growth factor-II (IGF-II) forms causes tumour hypoglycaemias and presumably also has an effect on tumour cell growth. We investigated the molecular weights of IGF-II forms and their ability to form complexes with IGF binding proteins (IGFBPs) in conditioned culture media (CM) from 12 paediatric soft tissue sarcoma (STS) cell lines and from two healthy fibroblast lines. Untreated CM were separated by size exclusion chromatography using biocompatible HPLC. Subsequently, IGF-II, IGFBP-2 and IGFBP-3 were determined in the HPLC fractions by specific RIAs. In the CM, IGF-II concentrations between 0.5 and 8.6 ng/10(6) cells were measured but no IGF-I was detectable. Parallel to this investigation, a high IGF-II mRNA level averaging 44.4 +/- 29.7% was measured by semi-quantitative RT-PCR. The STS cell lines secreted a higher proportion of big-IGF-II forms reaching 10-18 kD (10-33% of the total IGF-II secreted) compared to the healthy fibroblasts (2.5-5%). At the same time, the proportion of IGF-II bound with IGFBP in complexes of 35- 70 kD and 150 kD was reduced by up to 85% in CM from tumour cells. The tumour cell lines apparently secrete a different spectrum of IGF-II forms than healthy fibroblasts. The reduced ability to form complexes with IGFBP and the higher molecular weight of the IGF-II forms produced by the tumour cells indicate that these forms could in fact be the known tumour-associated pro-IGF-II forms. Due to these characteristics, the big-IGF-II forms probably have an altered biological effect on the tumour cells when compared to IGF-II. PMID- 10725784 TI - Intellectual and physical performance and morbidity in relation to height in a cohort of 18-year-old Swedish conscripts. AB - To test whether short stature in young men without malformations or chronic childhood diseases is associated with intellectual and physical performance and morbidity, a large cohort of apparently healthy 18-year-old Swedish men was analysed. The original cohort consisted of all men born in 1976 and conscripted in 1994 (n = 38, 900). After exclusion due to growth-affecting disorders or missing data, 32,887 subjects were available for analysis. Short conscripts (height below or equal to -2 SD scores) demonstrated increased overall morbidity compared with taller conscripts (above -2 SD scores). Specifically, short conscripts had more psychiatric and musculoskeletal diagnoses and were more often considered psychologically unsuitable for military service. Mean intellectual performance increased continuously with height; the mean 'standard nine' score was 4.22 for the short men and 5.17 for the rest (p < 0. 001). Short conscripts scored less well than taller conscripts in assessment of psychological functioning during mental stress, and were evaluated as less suitable for leadership positions. Maximal working capacity per kilogramme body weight correlated negatively with height (p < 0.001). In conclusion, short stature was associated with increased morbidity and psychological problems and with lower mean intellectual performance. To what extent this association is direct or indirect needs further evaluation. PMID- 10725785 TI - Influence of IGF-I and cell density on MDR1 expression in the T-lymphoblastoid cell line CCRF-CEM. AB - The debate about a direct or indirect effect of GH and IGF-I on the recurrence of malignancy, especially in the case of rhGH therapy in patients with leukemia, is still going on. Recent studies suggested that IGF-I plays a role in drug resistance during anticancer therapy. This resistance to diverse cytotoxic drugs, named multidrug-resistance (MDR), is mainly due to high levels of P-glycoprotein (P-gp). The gene encoding this membrane-associated transporter protein was named MDR1, and increased levels of P-gp are linked to enhanced MDR1 mRNA expression. Our aim was to investigate a possible effect of rhIGF-I on MDR1 gene expression in vitro. We cultured the T-lymphoblastoid cell line CCRF-CEM with different rhIGF-I concentrations (0, 5, 20 and 50 ng/ml) in serum-free medium for 3 days. CCRF-CEM cells are drug-sensitive and express MDR1 at low levels. MDR1 mRNA expression was measured by semiquantitative RT-PCR using a competitive assay with a heterologous DNA construct. In addition, GAPDH mRNA was amplified as an internal control for RNA integrity. P-gp activity was determined by a flow cytometric assay measuring rhodamine 123 accumulation. Furthermore, cell proliferation was monitored in all experiments. Our data do not support an effect of rhIGF-I on MDR1 mRNA expression, P-gp activity or cell proliferation in the CCRF-CEM cell line. MDR1 mRNA levels were inversely correlated to cell density with high significance (p < 0.0001). In conclusion, multidrug resistance linked to P-gp is not induced by IGF-I in CCRF-CEM cells. At high density, CCRF-CEM cells downregulate MDR1 gene expression. Our experimental model provides a very useful tool for monitoring the influence of growth factors on multidrug resistance in vitro. PMID- 10725786 TI - Zinc supplementation increases the level of serum insulin-like growth factor-I but does not promote growth in infants with nonorganic failure to thrive. AB - We investigated in a randomized double-blind placebo-controlled study the effects of zinc supplementation (2 mg/kg/day) for 12 weeks on growth, serum insulin-like growth factor-I (IGF-I) and insulin-like factor binding protein-3 (IGFBP-3) on 3- to 9-month-old infants with nonorganic failure to thrive (NOFTT). 25 infants completed the study, 14 received zinc supplementation (group A), and 11 received placebo (group B). The control group for baseline measurements was composed of 10 age-matched normal growing infants. There were no significant changes in weight for age, length for age, or weight for length during the entire study period in either group A or B. Serum IGF-I levels at baseline were similar in all the groups. After 12 weeks of therapy, serum IFG-I levels increased significantly only in the zinc-supplemented group, from 40.3 +/- 7 ng/ml at baseline to 65 +/- 8 ng/ml (p < 0.05). There was a marked difference in serum IGF-I levels between the zinc-supplemented group and the placebo group after 12 weeks: 65 +/- 8 vs. 49.4 +/- 5 ng/ml (p = 0.058, 95% CI of difference 9.88-21.31). No change was demonstrated in serum IGFBP-3 levels in either study group. We conclude that although zinc supplementation increased serum IGF-I levels, it did not improve the growth parameters of infants with NOFTT. PMID- 10725787 TI - Hypothyroid myopathy with unusually high serum creatine kinase values. AB - Depending on the degree of hormone deficiency, skeletal muscle involvement may occur in hypothyroidism. Usually, hypothyroid myopathy is associated with creatine kinase values <5,000 U/l. We report a 54-year-old man suffering from increasing fatigability, hoarseness, gait disturbances and a creatine kinase of 9,000 (normal: <80 U/l). He presented with bradyphrenia, macroglossia, dysarthria, myxedema, monoparesis, reduced deep tendon reflexes and stocking-type sensory disturbances. Free triiodthyronine was 0.25 pg/ml (normal: 0.6-1.9 pg/ml), free thyroxine <0.1 ng/dl (normal: 0. 6-1.8 ng/dl) and the thyroid stimulating hormone >48.0 (normal: 0. 1-4.0 IU). Clinical neurologic examination and electromyography were compatible with myopathy and polyneuropathy. Other causes of myopathy, except hypothyroidism, were excluded. After L-thyroxine therapy (1.7 microg/kg BW/day) during 3 months, the patient's symptoms and signs vanished, except for sensory disturbances, and creatine kinase values and electromyography became normal. Severe hypothyroidism may be associated with highly elevated creatine kinase and myopathy. Adequate therapy leads to complete recovery, including myopathy. PMID- 10725788 TI - Alternative versus classical activation of macrophages. AB - In parallel to the Th1/Th2 paradigm, antigen-presenting cells (APC) are divided into classically activated APC (dendritic cells/effector macrophages) and alternatively activated APC (IL-4-induced, alternatively activated macrophages/IL 10-induced, immature dendritic cells). Alternatively activated APC share a special molecular repertoire including receptors of innate immunity with broad specificity for foreign antigen and anti-inflammatory cytokines such as IL-1Ra and alternative macrophage activation-associated CC-chemokine-1. Alternatively activated APC mediated tolerance and downregulated inflammation. Abuse of alternatively activated APC in support of infectious susceptibility or tumor immune escape is counteracted by the classical pathway. Thus, classically and alternatively activated APC secure the balance between proinflammatory and anti inflammatory immune reactions. PMID- 10725789 TI - Fighting infection: the role of lipopolysaccharide binding proteins CD14 and LBP. AB - An invading pathogen must be held in check by the innate immune system until a specific immune response is mounted. Nonclonal pattern recognition receptors like CD14 or lipopolysaccharide (LPS) binding protein (LBP) recognize ubiquitous pathogen-associated molecular patterns, e.g. LPS. LBP mediates the binding of minute amounts of LPS to membrane-bound CD14 (mCD14) triggering a proinflammatory response of macrophages, which is crucial for keeping an infection under control. Moreover, in vitro mCD14 and LBP are involved in recognition and phagocytosis of heat-killed bacteria. Living Salmonella typhimurium or Escherichia coli depend on the presence of LBP to induce the generation of reactive oxygen species in human or murine macrophages. Using LBP-deficient mice it could be demonstrated that LBP is essential to control low dose (100 CFU S. typhimurium) infection. Therefore, LPS binding proteins play a pivotal role in physiology as well as pathophysiology of Gram-negative infection. PMID- 10725790 TI - The regulatory role of MRP8 (S100A8) and MRP14 (S100A9) in the transendothelial migration of human leukocytes. AB - The hallmark of developing inflammatory lesions is the excess migration of recruited phagocytes together with the enhanced cell surface expression of adhesion molecules. Recent investigations give evidence that the two myeloid related proteins MRP8 (S100A8) and MRP14 (S100A9), which are abundant in activated or recruited phagocytes, may have a modulatory role in inflammatory responses. S100A9 displays a regulatory role in the transendothelial migration of human monocytes, and the secreted S100A8/A9 complex may serve as a transport protein to move arachidonic acid to its target cells. PMID- 10725791 TI - The contact of human macrophages with extracellular matrix proteins selectively induces expression of proinflammatory cytokines. AB - The interaction of macrophages with proteins of the extracellular matrix (ECM) is important for the regulation of the immune and nonimmune functions displayed by these cells. Little, however, is known about the ability of different ECM proteins to transmit inflammatory signals into macrophages. Here we investigated the effect of the ECM proteins collagen type I, fibrin and fibronectin on the expression of the proinflammatory cytokines interleukin-1beta (IL-1beta) and IL-8 using RT-PCR, Northern and Western blot analysis and ELISA technique. It was found that collagen strongly induced IL-1beta and IL-8 expression in the macrophages. Fibronectin also stimulated cytokine expression, however, the amounts of the specific mRNAs were significantly lower compared to those induced by collagen. On the protein level IL-1beta revealed a close correlation to the mRNA expression. In contrast, fibrin did not elicit any IL-1beta and IL-8 response. These data show that different ECM proteins vary in their ability to induce proinflammatory cytokine expression in human macrophages suggesting that the protein composition of the ECM might be crucial in the initiation of inflammatory processes. PMID- 10725792 TI - ATP-binding cassette transporter A1 (ABCA1) in macrophages: a dual function in inflammation and lipid metabolism? AB - Activated lipid-laden macrophages in the vascular wall are key modulators of the inflammatory processes underlying atherosclerosis. We demonstrate here that the ATP-binding cassette (ABC) transporter ABCA1 is induced during differentiation of human monocytes into macrophages. ABCA1 has been implicated in macrophage interleukin-1beta secretion and apoptosis. Moreover, ABCA1 mRNA and protein levels are strongly upregulated by uptake of modified LDL and downregulated by HDL(3)-mediated lipid efflux in macrophages. Mutation analysis in patients with the classical Tangier disease (TD), a monogenetic disorder characterized by hypersplenism, macrophage accumulation and deposition of cholesteryl esters in the reticuloendothelial system, low plasma HDL and premature atherosclerosis, revealed deleterious mutations in their ABCA1 gene. The localization pattern of the mutations within the ABCA1 protein appears to determine the tropism for either the reticuloendothelial system, as seen in the classical TD phenotype, or the artery wall, as in the case of HDL deficiency in the absence of splenomegaly. In a comprehensive analysis of the expression and regulation of all currently known human ABC transporters, we identified additional cholesterol-responsive genes that are induced during monocyte differentiation into macrophages. Our results indicate a dual regulatory function for ABCA1 in macrophage lipid metabolism and inflammation. PMID- 10725793 TI - Regulation of macrophage gene expression by pro- and anti-inflammatory cytokines. AB - The anti-inflammatory cytokines IL-4 and IL-10 are well recognized as important negative regulators of proinflammatory gene expression in mononuclear phagocytes. The intracellular mechanisms which mediate these responses appear to be multifactorial. IL-4 is able to markedly suppress transcriptional activation of IFNgamma-responsive genes and the promoter sequences required for both IFNgamma and IL-4 sensitivity are identical. IFNgamma-activated STAT1 and IL-4-activated STAT6 can both form complexes on the same regulatory sequence element; while STAT1 functions to promote transcription, STAT6 is inactive. STAT6 is, however, required for the suppressive activity of IL-4. In this model, IL-4 appears to suppress IFNgamma-inducible proinflammatory gene expression through the ability of STAT6 to compete with STAT1 for occupancy of promoter sites necessary for IFNgamma-induced transcriptional initiation. In a second model, IL-10 suppresses the expression of genes induced in LPS-stimulated macrophages through a pathway involving destabilization of specific mRNAs. We have demonstrated that nucleotide sequences in the 3'-untranslated region of an IL-10-sensitive gene can both destabilize a stable reporter mRNA (CAT) and confer sensitivity to IL-10-mediated destabilization. Deletion and site-specific mutagenesis have mapped this to an AU rich sequence motif similar to that found in many cytokine and growth factor mRNAs. IL-10 is able to modulate the activity of proteins capable of binding to this sequence and one or more of these may regulate the rate of mRNA degradation. Thus mechanisms through which IL-10 and IL-4 act to dampen inflammatory responses are mechanistically distinct and involve diverse intracellular signaling pathways. PMID- 10725794 TI - Alternative activation of macrophage by IL-10. AB - Macrophage activation by proinflammatory stimuli is suppressed by IL-10. We tested the hypothesis that IL-10 induces an alternative state of macrophage activation rather than solely mediating suppression. We therefore searched for genes the expression of which might be up-rather than downregulated in response to IL-10. Total RNA was obtained from mouse macrophages J774 A.1 before or after stimulation with IL-10 (20 ng/ml). Poly(A)+RNA was isolated in both cases in order to obtain driver and tester mRNA. Subtraction suppression hybridization was performed using the PCR-select cDNA subtraction procedure. After evaluation of the subtraction efficiency the subtracted cDNA library was cloned into pCRII.1. A total of 1,300 clones were obtained. Southern blot hybridization analysis was performed as the first screening step of this total number of clones. 140 (10.7%) were identified as upregulated in response to IL-10. Sequence analyses so far showed perfect or near perfect matches with already known genes for the majority of clones. Our results clearly indicate that IL-10 stimulates the expression of a large number of genes in macrophages. We conclude that IL-10 induces in macrophages a noninflammatory state of reactivity which may serve to contain proinflammatory conditions. PMID- 10725795 TI - Phenotypic analysis of IL-10-treated macrophages using the monoclonal antibodies RFD1 and RFD7. AB - Suppressive or tolerogenic antigen-presenting cells (APC) might play an important role in the control of auto/hyperreactivity and the resolution of the immune response. Recent studies have provided evidence that tolerogenic APC can be induced by anergic T cells or interleukin-10 (IL-10). The aim of this study is to investigate how anergic T cells and IL-10 induce the suppressive APC phenotype and how this affects the immune response. Previously, two monoclonal antibodies (RFD1 and RFD7) were described by our lab which distinguish inductive (RFD1+RFD7 ), phagocytic (RFD1-RFD7+) and suppressive (RFD1+RFD7+) macrophages. RFD1 recognizes an MHC class II-associated epitope which has restricted expression, and RFD7 recognizes a predominantly cytoplasmic antigen. Macrophages were derived from the adherent fraction of peripheral blood mononuclear cells from healthy donors. At day 5, IL-10 or IFNgamma (a cytokine which should lead to the inductive APC phenotype) was added to the cultures. At day 7, the macrophages were harvested and their phenotypes were assessed by immunohistochemical staining and FACS analysis. Upon culture of macrophages with IL-10 RFD1 staining and HLA class II expression were reduced, whereas RFD7 staining was increased. Incubation of APC with IFNgamma led to upregulation of RFD1 and HLA class II, without affecting RFD7 staining. This suggests that IL-10 induced the suppressive RFD1+RFD7+ APC population, whereas IFNgamma treatment led to the inductive RFD1+RFD7- APC subset. Thus the use of IL-10 and/or IFNgamma, and the discrimination offered by mAbs RFD7 and RFD1 represent a model whereby APC function in terms of T cell stimulation or T cell anergy can be assessed. PMID- 10725796 TI - Comparison of monocyte functions after LPS- or IL-10-induced reorientation: importance in clinical immunoparalysis. AB - Immunoparalysis is an acquired immunodeficiency which may occur in patients after major surgery, burns, polytrauma and sepsis. It is associated with a modified state of monocytes marked by their altered capacity to induce antigen-specific T cell stimulation and to release various cytokines. However, the pathogenesis of immunoparalysis may differ in various patient groups. It can develop in patients after systemic hyperinflammation induced by gastrointestinal translocation of endotoxin (lipopolysaccharide, LPS) or sepsis, as well as in patients without preceding systemic inflammation but primary anti-inflammation, for instance induced by sympathetic activation. To further elucidate the syndrome, we compared endotoxin tolerance as a model of immunoparalysis after systemic hyperinflammation versus interleukin-10 (IL-10) treatment as a model of primarily anti-inflammation-induced immunoparalysis. In vitro priming of peripheral blood mononuclear cells with either LPS or IL-10 for 24 h led to a strongly or moderately diminished LPS-induced tumor necrosis factor-alpha (TNF-alpha) production, compared to unprimed controls, respectively. Furthermore, LPS-induced reduction of TNF-alpha production capacity persisted over the following days whereas IL-10-primed monocytes rapidly recovered. Similarly, in contrast to persistently diminished MHC class II expression in LPS-treated monocytes, IL-10 only transiently downregulated these molecules. Consequently, in contrast to IL 10-primed monocytes, LPS-primed monocytes were greatly impaired in their capacity to induce antigen-specific T cell proliferation and IFN-gamma production. These data indicate that LPS priming provokes a more profound modulation of monocyte function than IL-10 priming, raising the question of possible variations in the clinical course of immunoparalysis, dependent on its pathogenesis. PMID- 10725797 TI - The scavenger receptor CD163: regulation, promoter structure and genomic organization. AB - CD163 is a recently identified member of the scavenger receptor cysteine-rich superfamily expressed on peripheral blood monocytes and most tissue macrophages. We demonstrate that in vitro culture of human blood monocytes with recombinant M CSF induces CD163 transcription. In contrast, dendritic differentiation in the presence of GM-CSF and IL-4 suppresses CD163 mRNA and protein levels. Because an important function of CD163 in inflammation has been suggested, we investigated the influence of pro- and anti-inflammatory stimuli on CD163 expression and found a significant suppression by lipoposaccharide and IFN-gamma, whereas IL-10 or dexamethasone strongly induced the expression of CD163. The induction of CD163 mRNA by dexamethasone is suggested to be mediated by several glucocorticoid receptor binding sites located in the proximal promoter region. In addition, this sequence contains potential binding sites for the transcription factors Sp1, C/EBPalpha, Ets-2, PU.1 and AP-1, which have been shown to play an important role in myeloid-specific gene expression. We also identified an L1-transposable element 1.4 kb upstream of the transcription start site that might influence the promoter activity. The function of CD163 may also depend on the use of different isoforms. Several variants of CD163 mRNA have been described that encode proteins with altered cytoplasmic or extracellular domains and thus may differ in their functional properties. We analyzed the genomic organization of the CD163 gene and could demonstrate that these isoforms result from alternative splicing. Further characterization of the isoforms may help to understand the complete function of CD163. PMID- 10725798 TI - Glucocorticoid receptor ligand binding in monocytic cells using a microplate assay. AB - Glucocorticoids have profound effects on macrophage function and are widely used as anti-inflammatory drugs. Glucocorticoids receptor (GR) ligand binding capacity is a major determinant of cellular glucocorticoid sensitivity. The number and affinity of GR can be measured in a whole cell binding assay using (3)H dexamethasone. Here, we describe a rapid and simple microplate assay for GR measurement using the human promonocytic cell line THP-1. PMID- 10725799 TI - Suppressive antigen-presenting cells in Helminth infection. AB - Infection with filarial nematodes is commonly associated with a failure of T cells to proliferate in response to parasite antigen. We have investigated the possibility that antigen-presenting cells recruited during filarial infection are responsible for impairment of T cell function. We have found that the human filarial parasite Brugia malayi, when implanted into the peritoneal cavity of mice, recruits a population of adherent cells that actively block the proliferation of T cells. Phenotypic analysis of the recruited cells reveals large numbers of both macrophages and eosinophils and cell sorting experiments demonstrate that these antiproliferative cells are Mac-1-positive. Studies in gene-deficient mice have demonstrated that proliferative suppression is dependent on the in vivo production of IL-4 but not IL-5 or IL-10, while suppression in vitro is not mediated by any known antiproliferative factor. Our results suggest that helminth infection can lead to the development and/or recruitment of an IL-4 dependent macrophage population that mediates suppression via a novel mechanism. PMID- 10725800 TI - The regulatory role of the antigen-presenting cell in the development of hepatic immunopathology during infection with Schistosoma mansoni. AB - A recent meeting held in Berlin (2nd Teupitz Colloquium) focused on the striking ability of mononuclear phagocytes to either stimulate or inhibit a variety of immune and immunopathological responses, and ascribed a distinctive phenotype to the antigen-presenting cells (APC) when exercising these opposite functions. Thus, the phenotype and secretory profile of APC associated with 'classical activation' is achieved following stimulation with pro-inflammatory cytokines such as interferon-gamma, and leads to full T cell activation. On the other hand, it has long been known that T cells may also be downregulated after interacting with certain APC. Many of such APC, originally simply thought to lack or have lost their stimulatory potential, are now thought to be in a state of 'alternative activation', which is associated with a different phenotype and secretory profile that can typically be induced with anti-inflammatory reagents, including the cytokines IL-4, IL-10 and IL-13. The purpose of this article is to analyze the immunopathological events that characterize the infection with Schistosoma mansoni in context with these distinct APC activation pathways. Available evidence from human and experimental data suggests that a desirable outcome of the APC during this parasitic disease is to attain 'alternative activation', which serves to promote and sustain a Th-2-polarized immune response associated with a more favorable anti-inflammatory and host-protective environment. PMID- 10725801 TI - Experimental African trypanosomiasis: differences in cytokine and nitric oxide production by macrophages from resistant and susceptible mice. AB - Immunosuppression in experimental infections with Trypanosoma congolense is mediated by the synergistic action of macrophages and a novel lymphocyte(s), which involves the activity of IFN-gamma as well as IL-10. BALB/c mice are highly susceptible while C57Bl/6 mice are relatively resistant to T. congolense infections. Plasma and/or supernatants of spleen cell cultures of infected susceptible BALB/c mice have more IL-10 but less IL-12 than those of infected relatively resistant C57Bl/6 mice. Cells of a BALB/c macrophage cell line, when pulsed with T. congolense, produce more IL-10 and IL-6, but have less TNF-alpha mRNA, than equally treated cells of a C57Bl/6 cell line. Peritoneal and/or bone marrow-derived macrophages obtained from BALB/c mice, pulsed with T. congolense in culture, produce less nitric oxide, TNF-alpha and IL-12, but more IL-6 and IL 10 than equally treated macrophages isolated from C57Bl/6 mice. We suggest that genetic resistance to African trypanosomiasis is expressed at the level of the macrophage. PMID- 10725802 TI - Balanced macrophage activation hypothesis: a biological model for development of drugs targeted at macrophage functional states. AB - Macrophages play a central role in the immune response and are major targets for chronic infection with viruses such as HIV. Recent studies on macrophage differentiation have shown the existence of classical activation and the counter balancing anti-inflammatory alternative activation states. In the 'balanced macrophage activation hypothesis' we propose that macrophage activation is a cyclic process that balances these two states to achieve proper immunologic function. Dysregulation of this cycle would, therefore, be associated with various forms of chronic disease. This model has been utilized in the drug development of WF10, a novel macrophage-targeted drug, currently in advanced clinical testing for the treatment of HIV disease. PMID- 10725803 TI - Stimulatory and inhibitory maturation of human macrophage-derived dendritic cells. AB - Circulating human macrophages are often used to generate dendritic cells (DCs) by culturing them in granulocyte macrophage-colony stimulating factor (GM-CSF) and interleukin-4 (IL-4). As DCs are superb antigen-presenting cells, these types of myeloid DCs are now used in many DC-based vaccination protocols, especially in cancer, with the belief that they are essentially stimulatory or 'immunogenic'. Here we show that just as peripheral macrophage-derived myeloid DCs can be stimulatory, in vitro cultures of myeloid DCs in GM-CSF and IL-4 followed by further maturation in interferon-gamma plus bacterial superantigens (as DC maturing agents) can give rise to DCs that are functionally inhibitory. The stimulatory DCs express higher amounts of costimulatory molecules, synthesize IL 12, and efficiently stimulate naive allogeneic T cells in mixed lymphocyte reaction (MLR). The inhibitory DCs, in contrast, express lower concentrations of the critical costimulatory molecules, synthesize large amounts of IL-10, and are either marginally stimulatory or nonstimulatory in MLR. Moreover, while the stimulatory DCs further amplify proliferation of T cells in lectin-driven proliferation assays, the inhibitory DCs suppress T cell proliferation in similar assays, in vitro. Most interestingly, neutralization of the endogenously derived IL-10 with anti-IL-10 antibody with DC cultures as well as exposure of the inhibitory DCs to CpG oligonucleotides or to in vitro activated autologous CD4+ T helper cells repolarize them into stimulatory phenotype. Accordingly, these observations have important implications in translational research involving myeloid DCs. PMID- 10725804 TI - M-DC8+ leukocytes--a novel human dendritic cell population. AB - Dendritic cells (DC) constitute a heterogeneous leukocyte population having in common a unique capacity to induce primary T cell responses and are therefore most attractive candidates for immunomodulatory strategies. Two populations of blood DC (CD11c+ CD123(dim) and CD11c- CD123(high)) have been defined so far. However, their direct isolation for experimental purposes is hampered by their low frequency and by the lack of selective markers allowing large scale purification from blood. Here we describe the monoclonal antibody (mAb) M-DC8, which was generated by immunizing mice with highly enriched blood DC. This mAb specifically reacts with 0.2-1% of blood leukocytes and enables their direct isolation by a one-step immunomagnetic procedure from fresh mononuclear cells. These cells can be differentiated from T cells, B cells, NK cells and monocytes using lineage-specific antibodies. M-DC8+ cells express HLA class II molecules, CD33 and low levels of the costimulatory molecules CD86 and CD40. Upon in vitro culture M-DC8+ cells spontaneously mature into cells with the phenotype of highly stimulatory cells as documented by the upregulation of HLA-DR, CD86 and CD40; in parallel CD80 expression is induced. M-DC8+ cells display an outstanding capacity to present antigen. In particular, they proved to be excellent stimulators of autologous mixed leukocyte reaction and to activate T cells against primary antigens such as keyhole limpet hemocyanin. Furthermore, they induce differentiation of purified allogeneic cytotoxic T cells into alloantigen specific cytotoxic effector cells. While the phenotypical analysis reveals similarities with the two known blood DC populations, the characteristic expression of FcgammaRIII (CD16) and the M-DC8 antigen clearly defines them as a novel population of blood DC. The mAb M-DC8 might thus be a valuable tool to determine circulating DC for diagnostic purposes and to isolate these cells for studies of antigen-specific T cell priming. PMID- 10725805 TI - The role of cytokines in monocyte apoptosis. AB - Survival or apoptosis, activation and differentiation, phagocytosis and antigen presentation, migration or participation in granuloma formation are features of freshly recruited blood-borne monocytes in the local environment. In this presentation we describe that human monocytes undergo spontaneous apoptosis in vitro which involves Fas/FasL interactions, and that proinflammatory cytokines such as tumor necrosis factor-alpha (TNFalpha), interleukin-1beta and granulocyte monocyte-colony-stimulating factor prevent spontaneous apoptosis. In vitro infection of purified monocytes with low numbers of Mycobacterium tuberculosis H37Rv prevents spontaneous apoptosis. The apoptosis-preventing effect is correlated to the release of TNFalpha and not due to phagocytosis per se. Furthermore, the minor subset of CD64-negative monocytes is found to be less susceptible to recall antigen-activated CD4-positive T cell-mediated apoptosis than CD64-positive monocytes. Finally, recent findings of our group indicate that the chemokine platelet factor 4 protects monocytes from spontaneous apoptosis and induces the differentiation of monocytes into macrophages. From these findings we conclude that monocyte recruitment, their survival, their differentiation and their functional activity at the site of inflammation are regulated by a cytokine network which needs to be further analyzed in order to design strategies for immune intervention. PMID- 10725806 TI - Supernatants of virus-infected macrophages prime uninfected macrophages for lipopolysaccharide-induced apoptosis by both an interferon-dependent and an independent mechanism. AB - Virus-infected macrophages (M phi) release factors priming uninfected M phi for LPS-induced apoptosis. This was shown for bovine M phi infected with cytopathogenic bovine virus diarrhea (BVD) virus or with bovine herpes virus-1 (BHV-1) but not for M phi infected with noncytopathogenic BVD virus. The former two viruses also induced interferon type I in M phi, which also primes macrophages for LPS-induced apoptosis. However, several lines of evidence suggested that virus-infected M phi produce IFN-independent factors priming for apoptosis. For example, a soluble IFN type I receptor abrogated the antiviral activity, but not the ability of supernatants to prime uninfected M phi for LPS induced apoptosis. It is suggested that the production of factors priming uninfected cells for apoptosis plays a part in virus-induced pathogenesis, e.g. in cattle afflicted with mucosal disease, or with infectious bovine rhinotracheitis, or in viral diseases inducing a marked lymphopenia. PMID- 10725807 TI - Activation or destruction of T cells via macrophages. AB - Macrophages (MPhi) affect the T cell response in two mutually exclusive ways: activation or deletion. A MPhi type with T cell activating functions (M1) is able to express and upregulate receptors of the B7 family. IFN-gamma favours this MPhi differentiation pathway via upregulation of CD80 (B7-1) and CD86 (B7-2). The treatment of MPhi with IFN-gamma enhances the alphaCD3-mediated T cell blast transformation and reduces the fraction of deleted T cells. This MPhi type may prevent antibody-mediated T cell destruction by the expression of costimulatory receptors. An IL-10-induced MPhi type (M2) fails to express costimulatory molecules of the B7 family but is an effective cell for T cell destruction. Forming cellular conjugates with T cells through antibodies or immune complexes, M2-MPhi preferentially delete targeted cells in vitro and in vivo. PMID- 10725808 TI - Differential effects of necrotic or apoptotic cell uptake on antigen presentation by macrophages. AB - The induction of pathogenic immune responses may be dependent on the immune system receiving 'danger' signals resulting from tissue damage, rather than tolerogenic stimuli associated with normal cell turnover. The aim was to test this hypothesis by comparing the effects of the uptake of necrotic and apoptotic cells on the ability of antigen-presenting cells (APC) to stimulate immune responses in vitro. The experiments focused on presentation by the macrophage, which is the main cell type adapted for clearing cellular debris in vivo. Murine bone marrow-derived macrophages were pulsed with neutrophils that had been rendered apoptotic or necrotic, and tested for the ability to induce T cell responses. The macrophages that had taken up necrotic, but not apoptotic, cells were able to stimulate recall proliferation by ovalbumin-specific T cells. Furthermore, the response to the mitogen concanavalin A (Con A) was more than 6 times higher when macrophages had been pulsed with necrotic, in comparison with apoptotic, cells. In control experiments, macrophages that had not been exposed to dying neutrophils stimulated weak responses to ovalbumin and Con A. To determine why the uptake of apoptotic and necrotic cells exert opposing effects on the ability of macrophages to stimulate T cells, the expression of costimulatory molecules by treated macrophages, and their production of potentially immunomodulatory cytokines were measured. Flow cytometry revealed that macrophages that had taken up necrotic, but not apoptotic, neutrophils expressed increased levels of CD40 compared to untreated controls within 4 h. Macrophages pulsed with apoptotic cells secreted higher levels of transforming growth factor-beta1 than those ingesting necrotic cells or untreated controls. Our interpretation of these results is that macrophages that have taken up necrotic cell debris present antigens to T lymphocytes with greater efficiency due to transient CD40 upregulation, whereas those that have ingested apoptotic cells are ineffective APC since they secrete inhibitory cytokine. PMID- 10725809 TI - Role of antigen-presenting cells in long-term antitumor response based on tumor derived apoptotic body vaccination. AB - Cellular therapy prospects for cancer are based on the development of T cell response, resulting in efficient tumor rejection and long-term protection. We have previously shown that treatment combining injection of interleukin-2 and tumor-derived apoptotic bodies, but not tumor cell extracts, permits to reject parental tumor in 40% of rats. We observed the implication of antigen-presenting cells (APCs) and tumor-derived apoptotic bodies in the rejection of established peritoneal carcinomatosis. We demonstrated that apoptotic bodies could be efficiently phagocytosed by monocytes, triggering them to an APC phenotype. When using these phagocytosing APCs, derived from peritoneal or blood monocytes, the remission rate reached 80% of rats. However, due to the lack of specific markers of rat monocyte-derived cells, the precise role of APCs, dendritic cells and/or macrophages responsible for this therapeutic improvement remained to be clarified. In order to elucidate this question, we developed an in vivo preventive cellular therapy based on tumor-derived apoptotic bodies, where macrophages were either depleted or activated. We report here that in a preventive antitumoral apoptotic body vaccination that allows survival for 40% of treated rats, the antitumor response was characterized by a specific long-term memory (cured rats rejected a second parental tumor cell challenge). Depletion of resident macrophages with silica or clodronate liposomes appeared to promote apoptotic body vaccination efficiency, increasing the treatment to 66% of success. In this case, FACS analysis showed that peritoneal cells present are essentially immature APCs and freshly recruited NK cells. In contrast, the onset of peritoneal inflammation by thioglycollate, inducing massive recruitment and activation of macrophages, reduced the overall survival, whatever the treatment was. Also, even though the surviving rate was better in silica-treated rats than control, no long-term protection was elicited. Our data suggest that massive inflammation, recruiting numerous activated macrophages, could inhibit tumor antigen presentation by 'professional' APCs having phagocytosed apoptotic bodies, and defavor a specific antitumoral T cell response. Although effective responses were developed against parental tumor cells with silica/apoptotic body treatment, they seemed only partial, limited to primary cytotoxic efficiency. In conclusion, even if macrophages did not appear necessary for a primary response to tumor cells, these cells seemed to be implicated in the establishment of memory and long-term antitumor response. PMID- 10725810 TI - Contact tolerance. AB - Contact tolerance describes an immunological state which is caused by ordinary contact allergens painted in doses too low to sensitize, either once or repeatedly, onto healthy intact skin. The tolerance state accomplished by this means in BALB/c and C57BI/6 mice was found to be mediated by hapten-specific T cells that adoptively transferred tolerance to naive recipients. Furthermore, these cells were shown to be sensitive to cyclophosphamide, to express the Lyt2+ (CD8) phenotype, and to produce, upon restimulation in vitro, predominantly anti inflammatory cytokines such as IL-4, IL-5 and IL-10. These data indicate that contact tolerance of the low zone type is actively mediated by Th2-like CD8 T cells rather than arising as a consequence of clonal anergy. The induction of contact tolerance appeared to be strictly dose-dependent. As opposed to sensitizing doses of allergen, subsensitizing doses did not involve epidermal Langerhans cells discernibly. This was suggested by their normal ultrastructure, their unaffected adenosine triphosphatase system, the inefficacy of functional blocking and excision experiments. Both radiolabeled and fluorescent contact sensitizers were observed to readily enter the bloodstream, thereby being dispersed throughout the body. Presumably, contact tolerance is induced systemically rather than locally. The presence of hapten-specific tolerance can only be uncovered through a subsequent attempt to sensitize. If the attained sensitization turns out to be significantly lower than that of immunologically naive controls, and if sensitization to chemically unrelated sensitizers is not impaired, hapten-specific tolerance does exist. Thus, contact tolerance is a result obtained from experimental sensitization in animals. Nonetheless, it is assumed to occur also in humans, although it is not demonstrable unless different proofs of existence become available. PMID- 10725811 TI - Induction of tolerance in macrophages by cholera toxin B chain. AB - Model systems of human type 1 diabetes have revealed an important role of cellular immune reactions involving macrophages and T cells in the destruction of autologous insulin-producing pancreatic beta cells. Recently, the cholera toxin B chain (CTB) was found to suppress T cell-dependent autoimmune diseases including autoimmune diabetes of nonobese diabetic mice. Therefore, we tested the hypothesis that CTB exerts much of its immunomodulatory activity by targeting macrophages. These studies are reviewed here. Cells of the human monocyte line Mono Mac 6 were exposed to CTB and subsequently tested for proinflammatory immunoreactivity in response to challenge with endotoxin (LPS from Escherichia coli, 10 ng/ml for 5 h). Incubation of monocytes with CTB (10 microgram/ml) suppressed a later proinflammatory response to LPS as demonstrated by suppression of TNFalpha release from 6.7 +/- 0.7 ng/ml in cultures without CTB preexposure to 1.8 +/- 1.1 ng/ml in CTB-pretreated cells (p < 0.001). In contrast, the release of IL-10 remained inducible after CTB pretreatment. RT-PCR analysis showed that the suppression of TNFalpha production occurred at the level of mRNA formation. Control experiments excluded a role of possible contamination of CTB by endotoxin or the intact cholera toxin. Tolerance induction was maximal after 5 h of CTB exposure and persisted for 24 h. The suppressive effect of CTB was dose-dependent and no more recognizable at /=98%) for stromal markers; about 60% displayed the characteristics of fibroblasts, and the remaining 40% were classified as smooth muscle cells. In late passages (P20), the proportion of muscle cells declined to 10%. Stimulation experiments including basic fibroblast growth factor (bFGF) resulted in enhanced proliferation, whereas dihydrotestosterone (DHT), estrogen, and flutamide did not influence proliferation. Gene expression studies demonstrated a positive signal for androgen receptor and keratinocyte growth factor (KGF). CONCLUSIONS: Prostatic stromal cells can be propagated several times and show karyotypic stability for up to 18 subculture experiments. The ratio of myoid and fibroblastic cells can be used for standardization of cell cultures with stable characteristics. PMID- 10725863 TI - Induction of apoptosis by mifepristone and tamoxifen in human LNCaP prostate cancer cells in culture. AB - BACKGROUND: Published data indicate that antiprogestins and antiestrogens could inhibit prostate cancer cell growth in vitro and in vivo. The main objective of the present studies was to explore the role of bcl(2) and TGFbeta(1) for induction of apoptosis in LNCaP prostate cancer cells growing in culture as a treatment response to the antiprogestin, mifepristone, and the antiestrogen, 4 hydroxytamoxifen. METHODS: In vitro cell viability (cytotoxicity), DNA fragmentation, and changes in the expression of bcl(2) and TGFbeta(1) proteins were assessed using the sulforhodamine B protein dye-binding assay, specific ELISA, and competitive inhibition assays. RESULTS: Both steroid antagonists induced a significant time- and dose-dependent cell growth inhibition (cytotoxicity). This inhibition of viable cells was associated with a significant increase in DNA fragmentation (apoptosis), downregulation of bcl(2), and induction of TGFbeta(1) protein. Abrogation of the mifepristone- and 4 hydroxytamoxifen-induced cytotoxicity by TGFbeta(1)-neutralizing antibody and by the addition of mannose-6-phosphate confirmed the correlation between induction of active TGFbeta(1) and subsequent prostate cancer cell death. The effect of mifepristone was not significantly reduced or prevented by occupying the progesterone or glucocorticoid receptors by their corresponding high-affinity native ligands. On the contrary, the effect of a combination of mifepristone with progesterone or hydrocortisone on the increase in DNA fragmentation, bcl(2) downregulation, and induction of TGFbeta(1) protein was additive and significantly different (P < 0.05) from the effect of mifepristone monotherapy. CONCLUSIONS: Our data suggest that mifepristone and tamoxifen are effective inducers of apoptosis and may represent nonandrogen-ablation, novel therapeutic approaches to overcome a potential intrinsic apoptosis resistance of androgen independent prostate cancer cells. PMID- 10725864 TI - Magnetic resonance imaging of prostatic cancer: does detection vary between high and low gleason score tumors? AB - BACKGROUND: Both Gleason score and prostate-specific antigen (PSA) concentration are prognostic factors for prostate cancer. We assessed our ability to localize cancer lesions based on Gleason scores and PSA values by endorectal coil magnetic resonance imaging (MRI). We also evaluated whether the size of the prostate affects tumor detectability. METHODS: We compared the findings of MRI and histopathological results of radical prostatectomy specimens from 63 patients; they were divided into four groups, based on Gleason score and also on serum PSA concentration. Furthermore, the possible effect of prostatectomy specimen weight on MRI interpretation was examined. RESULTS: A highly significant difference appeared in detection of cancer lesions based on their differentiation grade. No statistically significant difference existed between PSA groups in detection of tumors, but the large size of the prostate seemed to render image interpretation more difficult. CONCLUSIONS: Endorectal MRI detects poorly differentiated prostate cancer lesions more accurately than clinically insignificant tumors. PMID- 10725865 TI - Pharmacological effects of the lipidosterolic extract of Serenoa repens (Permixon) on rat prostate hyperplasia induced by hyperprolactinemia: comparison with finasteride. AB - BACKGROUND: The growth of the prostate gland is mainly dependent on androgens. Other hormones, like prolactin (PRL), also influence prostate development. Our purpose was to analyze and compare the effects of two drugs (5alpha-reductase inhibitor) used in the therapy of benign prostatic hyperplasia: lipidosterolic extract of Serenoa repens (LSESR), and finasteride in an in vivo model of rat prostate hyperplasia induced by hyperprolactinemia. METHODS: Hyperprolactinemia was induced by 30 daily injections of sulpiride. Wistar rats received daily gavages of LSESR or finasteride. We used the following groups: control, castrated, castrated with a substitute testosterone (T), or 5alpha dihydrotestosterone (DHT) implant. RESULTS: Hyperprolactinemia increases the wet weight and induces hyperplasia in the lateral prostate (LP). Unlike finasteride, LSESR significantly reduced LP growth and hyperplasia in castrated, DHT implanted, and sulpiride-treated rats. CONCLUSIONS: Finasteride was only capable of inhibiting the effect of androgens on rat prostate enlargement. LSESR inhibited not only the androgenic but also the trophic effect of PRL in rat LP hyperplasia. PMID- 10725866 TI - Higher-dose and less frequent dendritic cell infusions with PSMA peptides in hormone-refractory metastatic prostate cancer patients. AB - BACKGROUND: Infusion of dendritic cells (DCs) pulsed with PSMA peptides was considered possible in hormone-refractory metastatic prostate cancer patients both with or without prior treatment with a greater number of DCs and for lesser infusions than previously administered. METHODS: DCs + PSMA peptides in patients undergoing leukapheresis were administered monthly 1-4 times, at rates greater than 20 million DCs in 17 patients not previously treated, and in 11 patients previously treated. RESULTS: Three partial responders and one complete responder were noted in the 17 previously untreated persons. DCs + PSMA peptides averaged 28.5 million cells (range in millions, 21.0-42.3). All responders received 3 or 4 infusions of greater than 22 million cells (3-4 times). In the previously treated group of 11 patients, DCs infused averaged 29.3 million cells (range in millions, 20-40.5). One new responder (bone scan) was noted. Two prior responders continued. Observation times were similar. Toxicity was minimal. CONCLUSIONS: These results suggest that DCs + PSMA peptide infusions can be given with greater numbers of DCs with a lesser number of infusions (1-4 monthly) with no loss of response rates compared to those noted previously, and without increased side effects. In previously treated patients (both relapsing and nonrelapsing), adverse effects were not noted, and new responses can be anticipated to be without harmful side effects. However, the follow-up time, and number of patients in this group, were small. PMID- 10725867 TI - Intermittent androgen suppression in the LuCaP 23.12 prostate cancer xenograft model. AB - BACKGROUND: Intermittent androgen suppression (IAS) has been proposed as a method of delaying the onset of androgen-independent growth in prostate cancer. While several pilot studies have demonstrated the feasibility of such a treatment, no study to date has defined the effect of IAS on survival. METHODS: We developed an IAS protocol for mice bearing the LuCaP 23.12 human prostate cancer xenograft, with each cycle consisting of 1 week of androgen replacement with a testosterone pellet followed by 3 weeks of androgen withdrawal. Mice that responded to castration with a 40% or greater decrease in serum prostate-specific antigen (PSA) were randomized to treatment with either continuous androgen suppression (CAS) or IAS. Serum PSA, tumor volume, and overall survival were monitored. RESULTS: A total of 75 mice met the randomization criteria. There was no significant difference of survival between animals treated with CAS or IAS (185 vs. 239 days, P = 0.1835). Serum PSA showed evidence of cycling with hormonal manipulation. No cycling was noted in tumor volume. CONCLUSIONS: IAS is not associated with a decrease in survival compared to CAS, yet in patients may offer quality-of-life improvements. Further studies of IAS in the setting of Institutional Review Board (IRB) approved clinical trials should be encouraged. Prostate 43:63-70, 2000. Published 2000 Wiley-Liss, Inc. PMID- 10725868 TI - Prostate-seminal vesicle cancers induced in noble rats. AB - Susceptibility of Noble strain rats to induced cancers in the accessory sex glands was determined. Following an IV inoculation of methylnitrosourea plus three successive SC implants of testosterone propionate, early stage adenocarcinomas were observed in the seminal vesicles and anterior prostates, but not in the dorsolateral nor in the ventral lobes. PMID- 10725869 TI - Anaplastic large cell lymphoma: a clinicopathologic analysis. AB - The clinicopathologic features of anaplastic large cell lymphoma (ALCL) are reviewed. ALCL is a heterogeneous group of tumours, and histologic examination alone is not adequate in providing useful prognostic information. However, using a combination of clinical, phenotypic, and genotypic features, several distinct clinicopathologic entities have been identified. A subset of ALCL as presently defined is characterized by a balanced translocation, t(2;5)(p23;q35), resulting in a novel fusion protein (NPM-ALK) that can be readily detected by immunohistochemical methods using antibodies against the ALK protein. Detection of ALK protein, along with other methods for demonstrating the t(2;5), has assisted in identifying a distinct biologic entity within the heterogeneous group of ALCL with significant prognostic implications. It is important to separate these from cases of ALK-negative ALCL, which have a poorer prognosis, and cases of primary cutaneous ALCL, which have an excellent prognosis. PMID- 10725870 TI - Regulation of the expression of MHC class I and II by class II transactivator (CIITA) in hematopoietic cells. AB - In order to develop an effective immunotherapy for hematological malignancies, we investigated the applicability of class II transactivator (CIITA), which had been demonstrated to regulate the expression of MHC class II (MHC-II) by assembling the transcription factors of MHC-II molecules, for immunotherapy by potentiating the antigenicity of tumour cells by inducing MHC expression. First, 32 hematopoietic cell lines were analysed for the expression of HLA-DR, CIITA, RFX5 or HLA-ABC. Fourteen cell lines were positive and 18 were negative for HLA-DR. All the 14 HLA-DR positive cell lines were demonstrated to express CIITA mRNA by RT-PCR. On the other hand, in all the 18 HLA-DR negative cell lines, the expression of CIITA was not demonstrated. RFX5, which is one of the transcription factors of MHC-II, was expressed ubiquitously in all 32 cell lines. Three cell lines out of 23 hematopoietic cell lines examined were negative for HLA-ABC, and all three of these cell lines were negative for both HLA-DR and CIITA expression. Furthermore, CIITA cDNA was transfected into K562 cells, which were negative for HLA-ABC, -DR and -DQ, but positive for HLA-DP. The transfection rendered HLA-DR negative to positive and increased the expression level of HLA-DP, but HLA-DQ remained negative. In addition to HLA-DR, HLA-ABC was also induced to express by the transfection of CIITA gene. The present study demonstrated that the expression of HLA-DR in hematopoietic cells is regulated in subordination to CIITA and the expression of HLA-DR (and HLA-ABC in K562) is induced by transfection with the CIITA gene. These findings revealed the applicability of CIITA in potentiating anti-tumour immunity of HLA-DR negative tumour cells for immunotherapy of hematological malignancies. PMID- 10725871 TI - Potential use of serum CD44 as an indicator of tumour progression in acute leukemia. AB - CD44 is a widely expressed cell surface glycoprotein with various functions. This molecule is shed from the cell surface and released as a soluble molecule. High serum levels of CD44 have been demonstrated in some solid tumours. In this study we measured serum CD44 in 25 patients with acute leukemia, in 12 with bacterial infections, and in 13 normal controls. The levels of serum CD44 of patients with bacterial infections were significantly higher (mean 531.3+/-60.1 ng/ml, p<0.001) than those of normal controls (299. 0+/-115.4 ng/ml). Acute leukemia patients before treatment had almost four-fold higher levels of serum CD44 than normal controls (mean 1301.9+/-1384.6 ng/ml, p<0.01). Serum CD44 levels were correlated with clinical status. After treatment the serum CD44 levels significantly decreased, but they were still higher than in normal controls. Patients in complete remission all relapsed if serum CD44 levels were higher than 500 ng/ml (normal+2 SD) after chemotherapy. The serum CD44 levels were correlated with the absolute numbers of leukemic cells in peripheral blood. The results demonstrated that serum CD44 levels correlate well with the clinical status of acute leukemia, and such evaluation may provide a reliable tumour marker of acute leukemia. PMID- 10725872 TI - Motion correction and lipid suppression for 1H magnetic resonance spectroscopy. AB - Spectral/spatial spin-echo pulses with asymmetric excitation profiles were incorporated into a PRESS-based localization sequence to provide lipid suppression while retaining a sufficient amount of water to allow for correction of motion-induced shot-to-shot phase variations. 1H magnetic resonance spectroscopy data were acquired at 1.5 Tesla from a motion phantom and in vivo from the human liver, kidney, and breast. The results demonstrated that lipids in the chemical shift stopband were completely suppressed and that full metabolite signal intensity was maintained after implementation of a regularization algorithm based on phasing the residual water signal. Liver and kidney spectra contained a large resonance at 3.2 ppm that was ascribed to trimethylammonium moieties (betaine plus choline) and a weaker signal at 3.7 ppm that may result from glycogen. A breast spectrum from a histologically proven invasive ductal carcinoma displayed a highly elevated choline signal (3.2 ppm) relative to that from a normal volunteer. PMID- 10725873 TI - A multiscale approach for analyzing in vivo spectroscopic imaging data. AB - A multiscale approach for analyzing in vivo magnetic resonance spectroscopic imaging (SI) data is described in this paper. With this method, fitting is performed at multiple spatial scales in a coarse-to-fine order. Results obtained at one scale are used as prior knowledge in fitting spectra at the next scale. The multiscale approach was validated with simulated data and demonstrated with proton SI datasets of the human brains. The results showed that this method improved the robustness and efficiency of the fitting and facilitated the automatic analysis of in vivo SI data. PMID- 10725874 TI - In vivo intermolecular double-quantum imaging on a clinical 1.5 T MR scanner. AB - A novel MRI method based on the intermolecular double-quantum coherence (DQC) for soft tissues is described. DQC images of human brain were obtained for the first time on a whole-body 1.5 T scanner. The combination of quantum and classical formalisms was used to characterize multiple-quantum coherences, and to aid in the design of a DQC imaging sequence. The theoretical analysis suggests that signals from the intermolecular DQCs have higher sensitivity than those from the zero-quantum coherence (ZQC) for human brain, and the sensitivity increases with increased field strength. The DQC signal may provide a new form of contrast for MRI. PMID- 10725875 TI - Hyperthermia by MR-guided focused ultrasound: accurate temperature control based on fast MRI and a physical model of local energy deposition and heat conduction. AB - Temperature regulation in MR-guided focused ultrasound requires rapid MR temperature mapping and automatic feedback control of the ultrasound output. Here, a regulation method is proposed based on a physical model of local energy deposition and heat conduction. The real-time evaluation of local temperature gradients from temperature maps is an essential element of the control system. Each time a new image is available, ultrasound power is adjusted on-the-fly in order to obtain the desired evolution of the focal point temperature. In vitro and in vivo performance indicated fast and accurate control of temperature and a large tolerance of errors in initial estimates of ultrasound absorption and heat conduction. When using correct estimates for the physical parameters of the model, focal point temperature was controlled within the measurement noise limit. Initial errors in absorption and diffusion parameters are compensated for exponentially with a user-defined response time, which is suggested to be on the order of 10 sec. PMID- 10725876 TI - Metabolic heterogeneity at the level of the anterior and posterior commissures. AB - Multislice, 2D proton spectroscopic imaging was performed in six healthy volunteers at long echo time (TE = 280 msec). The center of the most inferior of three slices was placed directly at the level of the line connecting the anterior and posterior commissures. Significant regional variations in metabolite levels were observed. In particular, based on statistical analysis, levels of choline were significantly high in insular cortex, thalamus, and centrum semiovale compared to other brain regions such as parietal or occipital gray and white matter. NAA levels were highest in the centrum semiovale white matter, while creatine levels were relatively constant. Globus pallidus exhibited lower signal intensities and increased linewidths for all metabolites. No spectra could be obtained from the inferior frontal lobe because of field inhomogeneity. These data show that the metabolism, and perhaps the underlying cellular composition, of thalamus and insular cortex appears to be different from other neocortical gray matter. Normal regional variations in the brain spectra should be considered when evaluating pathological conditions. PMID- 10725877 TI - Effects of antipsychotic drugs on metabolite ratios in rat brain in vivo. AB - Localized in vivo proton magnetic resonance spectroscopy at 4.7 T was used to examine the brains of rats that were given the antipsychotic drugs haloperidol, clozapine, or olanzapine for 1 week. Spectra were collected before and during treatment. The ratios of N-acetylaspartate (NAA) to creatine (Cr) and choline to Cr were determined from the spectra. No significant differences in these ratios were seen among the rats given the various antipsychotic medications or between the control rats and the treated rats. No significant time-dependent changes were seen in most cases, except for a small reduction of NAA/Cr after 7 days of olanzapine administration. These results suggest that differences in brain metabolite ratios in vivo in schizophrenics relative to controls, at least for short-term treatment, arise from the disease, and not as a metabolic effect of the medication. PMID- 10725879 TI - Test liquids for quantitative MRI measurements of self-diffusion coefficient in vivo. AB - A range of liquids suitable as quality control test objects for measuring the accuracy of clinical MRI diffusion sequences (both apparent diffusion coefficient and tensor) has been identified and characterized. The self-diffusion coefficients for 15 liquids (3 cyclic alkanes: cyclohexane to cyclooctane, 9 n alkanes: n-octane to n-hexadecane, and 3 n-alcohols: ethanol to 1-propanol were measured at 15-30 degrees C using an NMR spectrometer. Values at 22 degrees C range from 0.36 to 2.2 10(-9) m(2)s(-1). Typical 95% confidence limits are +/-2%. Temperature coefficients are 1.7-3.2% degrees C. T1 and T2 values at 1.5 T and proton density are given. n-tridecane has a diffusion coefficient close to that of normal white matter. The longer n-alkanes may be useful T2 standards. Measurements from a spin-echo MRI sequence agreed to within 2%. PMID- 10725878 TI - Statistically driven identification of focal metabolic abnormalities in temporal lobe epilepsy with corrections for tissue heterogeneity using 1H spectroscopic imaging. AB - 1H spectroscopic imaging of N-acetyl-aspartate, creatine, and choline has proven to be a sensitive indicator for the lateralization of seizure foci in temporal lobe epilepsy. Previous studies have used right-left comparisons to identify the epileptogenic tissue assuming that alterations due to the disease process outweigh the effects of tissue heterogeneity. To evaluate the effectiveness of tissue heterogeneity corrected analyses, we evaluated three criteria for lateralization of the seizure focus: 1) a statistically driven method adjusted for tissue composition, 2) a single valued threshold, and 3) a single global index of the hippocampus. The statistically driven analysis lateralized all eight patients correctly, whereas the single threshold method incorrectly lateralized one case and the global index failed to identify a significant difference in two cases. These findings indicate that increased accuracy and sensitivity can be obtained by correcting for tissue heterogeneity when analyzing spectroscopy studies of temporal lobe epilepsy. PMID- 10725880 TI - Three-dimensional whole body imaging of spin probes in mice by time-domain radiofrequency electron paramagnetic resonance. AB - Imaging of stable paramagnetic spin probes in phantom objects and in vivo was evaluated using a RF time domain EPR spectrometer/imager operating at 300 MHz. Projections were collected using static magnetic field gradients and images were reconstructed using filtered back-projection techniques. Results from phantom objects containing approximately 10(17) spins of stable paramagnetic probes with single narrow EPR spectra provide three-dimensional spatial images with resolution better than 2 mm. When the spin probe was administered to mice, the spin probe accumulation was temporally observed in the thoracic, abdominal, and pelvic regions. A three-dimensional image (from 144 projections) from a live mouse was collected in 5 min. Using fiducial markers, the spin probe accumulation in organs such as liver, kidney, and bladder could be observed. Differences in the oxygen status between liver and kidney were observed from the EPR images from mice administered with spin probe, by treating the time-domain responses with convolution difference approach, prior to image reconstruction. The results from these studies suggest that, with the use of stable paramagnetic spin probes and time-domain RF EPR, it is possible to perform in vivo imaging on animals and also obtain important spatially resolved physiologic information. PMID- 10725881 TI - Functional MRI of calcium-dependent synaptic activity: cross correlation with CBF and BOLD measurements. AB - Spatial specificities of the calcium-dependent synaptic activity, hemodynamic based blood oxygenation level-dependent (BOLD) and cerebral blood flow (CBF) fMRI were quantitatively compared in the same animals. Calcium-dependent synaptic activity was imaged by exploiting the manganese ion (Mn++) as a calcium analog and an MRI contrast agent at 9.4 T. Following forepaw stimulation in alpha chloralose anesthetized rat, water T1 of the contralateral forepaw somatosensory cortex (SI) was focally and markedly reduced from 1.99 +/- 0.03 sec to 1.30 +/- 0.18 sec (mean +/- SD, N = 7), resulting from the preferential intracellular Mn++ accumulation. Based on an in vitro calibration, the estimated contralateral somatosensory cortex [Mn++] was approximately 100M, which was 2-5-fold higher than the neighboring tissue and the ipsilateral SI. Regions with the highest calcium activities were localized around cortical layer IV. Stimulus-induced BOLD and CBF changes were 3.4 +/- 1.6% and 98 +/- 33%, respectively. The T1 synaptic activity maps extended along the cortex, whereas the hemodynamic-based activation maps extended radially along the vessels. Spatial overlaps among the synaptic activity, BOLD, and CBF activation maps showed excellent co-registrations. The center-of-mass offsets between any two activation maps were less than 200 microm, suggesting that hemodynamic-based fMRI techniques (at least at high field) can be used to accurately map the spatial loci of synaptic activity. PMID- 10725882 TI - In vivo MR measurements of regional arterial and venous blood volume fractions in intact rat brain. AB - In vivo measurement of cerebral arterial and venous volume fractions is important to the understanding of brain physiology and function. By using an intravascular perfluorocarbon and 19F NMR at 4.7 T, regional arterial and venous volume fractions from an intact rat brain were resolved based on the pseudodiffusion coefficients, which were (33 +/- 7) x 10(-3) and (0.45 +/- 0.13) x 10(-3) mm(2)/sec (mean +/- SD, n = 7) for the fast- and slow-moving component, respectively. By exploiting the linear dependence of the perfluorocarbon 19F 1/T1 on the dissolved paramagnetic oxygen concentration, combined inversion-recovery and diffusion measurements were made to correlate the short T1 (high-oxygenation) component with the fast-moving component and the long T1 (low-oxygenation) component with the slow-moving component. The arterial blood volume fraction was 29 +/- 7% of the total cerebral blood volume. Finally, experiments were performed in which different oxygen concentrations were inhaled to validate this technique. PMID- 10725883 TI - Kinetic characterization of CMD-A2-Gd-DOTA as an intravascular contrast agent for myocardial perfusion measurement with MRI. AB - Recent developments in magnetic resonance imaging (MRI) using specific contrast media allow the assessment of myocardial perfusion. The purpose of this study was to characterize the intravascular properties of a new macromolecular contrast agent, CMD-A2-Gd-DOTA, to evaluate myocardial perfusion. Two groups of isolated pig hearts perfused at various controlled flows were used. To demonstrate the intravascular properties of CMD-A2-Gd-DOTA, the agent was simultaneously injected with 99mTc-labeled red blood cells in five hearts (group 1). Tracer kinetics of both compounds were assessed by coronary sinus effluent sampling, radioactivity counting and concentration determination in samples for first-pass time curves measurements. Five other hearts (group 2) were studied using a two-slice turboFLASH sequence on a 1.5-T whole-body MRI in order to evaluate first-pass CMD A2-Gd-DOTA signal intensity (SI) versus time curves. In group 1, for the studied flows ranging from 0.8 to 3.1 ml/min(-1) x g(-1), CMD-A2-Gd-DOTA showed first pass concentration curves typical of an intravascular contrast agent. In group 2, MRI parameters, i.e., upslope and mean transit time of SI time curves correlated strongly with myocardial perfusion. Within the physiologic range of flows, CMD-A2 Gd-DOTA was able to demonstrate tracer kinetics for in vivo assessment of myocardial perfusion using MRI. PMID- 10725884 TI - Design of improved spectral-spatial pulses for routine clinical use. AB - Spectral-spatial pulses (spsp pulses) selectively excite spins at spatial location z and spectral frequency (due to chemical shift and/or field inhomogeneity) v. In this work we discuss the design of improved spsp pulses for fat signal suppression. Optimal pulses are designed as optimal constant ripple FIR filters using the inverse SLR transform. Spsp pulses with thin slices are obtained by modifying the phases between subpulses, thereby eliminating unwanted magnetization lobes. Robust spsp pulses at off-center slices are obtained with a prescan calibration. These pulses are used either for selective fat saturation or for selective water excitation. It is shown that spsp pulses suppress fat signal better than conventional fat saturation pulses. Using the techniques presented in this article, we replaced all the fat saturation pulses on our systems with spsp pulses and obtained a significant improvement in image quality. PMID- 10725885 TI - T1rho imaging using magnetization-prepared projection encoding (MaPPE). AB - T1rho contrast weighting using a magnetization-prepared projection encoding (MaPPE) pulse sequence was investigated. Fast radial imaging was implemented by applying magnetization preparation pulses, each followed by multiple RF alpha pulses encoding radial trajectories of k-space. Acquiring multiple views per preparatory pulse imposes view-to-view variation; the resultant distortion of the point-spread function is examined. The issue of maximizing signal while preserving the intended contrast weighting is addressed. Under modification of repetition time and flip angle (alpha), three distinct behavior regimes of the sequence are identified. The utility of the pulse sequence as a quantitative relaxation measurement tool is also examined by comparing imaging and spectroscopy experiments. A mouse was imaged in vitro to demonstrate the viability of application to MR histology. These images exhibit the utility of spinlocking and projection encoding as an aftemative contrast source to both T2 weighted MaPPE images and conventional T2-weighted spin-echo images. PMID- 10725887 TI - Hyperpolarized 3He microspheres as a novel vascular signal source for MRI. AB - Hyperpolarized (HP) 3He can be encapsulated within biologically compatible microspheres while retaining sufficient polarization to be used as a signal source for MRI. Two microsphere sizes were used, with mean diameters of 5.3 +/- 1.3 microm and 10.9 +/- 3.0 microm. These suspensions ranged in concentration from 0.9-7.0% gas by volume. Spectroscopic measurements in phantoms at 2 T yielded 3He relaxation times that varied with gas concentration. At the highest 3He concentration, the spinlattice relaxation time, T1, was 63.8 +/- 9.4 sec, while the transverse magnetization decayed with a time constant of T2* = 11.0 +/- 0.4 msec. In vivo MR images of the pelvic veins in a rat were acquired during intravenous injection of 3He microspheres (SNR approximately equal 15). Advantages such as intravascular confinement, lack of background signal, and limited recirculation indicate quantitative perfusion measurements may be improved using this novel signal source. PMID- 10725886 TI - Partial-FOV reconstruction in dynamic spiral imaging. AB - In many applications of dynamic MR imaging, only a portion of the field-of-view (FOV) exhibits considerable variations in time. In such cases, a prior knowledge of the static part of the image allows a partial-FOV reconstruction of the dynamic section using only a fraction of the raw data. This method of reconstruction generally results in higher temporal resolution, because the scan time for partial-FOV data is shorter. The fidelity of this reconstruction technique depends, among other factors, on the accuracy of the prior information of the static section. This information is usually derived from the reconstructed images at previous time frames. This data, however, is normally corrupted by the motion artifact Because the temporal frequency contents of the motion artifact is very similar to that of the dynamic section, a temporal low-pass filter can efficiently remove this artifact from the static data. The bandwidth of the filter can be obtained from the rate of variations inside and outside the dynamic area. In general, when the temporal bandwidth is not spatially uniform, a bank of low-pass filters can provide a proper suppression of the motion artifact outside the dynamic section. This reconstruction technique is adapted for spiral acquisition and is successfully applied to cardiac fluoroscopy, doubling the temporal resolution. PMID- 10725888 TI - Off-centered spiral trajectories. AB - The quality of spiral images depends on the accuracy of the k-space sampling locations. Although newer gradient systems can provide more accurate gradient waveforms, the sampling positions can be significantly distorted by timing misregistration between data acquisition and gradient systems. Even after the timing of data acquisition is tuned, minor residual errors can still cause shading artifacts which are problematic for quantitative MR applications such as phase-contrast flow quantitation. These timing errors can ideally be corrected by measuring the actual k-space trajectory, but trajectory measurement requires additional data acquisition and scan time. Therefore, off-centered spiral trajectories which are more robust against timing errors are proposed and applied to the phase-contrast method. The new trajectories turn shading artifacts into a slowly varying linear phase in reconstructed images without affecting the magnitude of images. PMID- 10725889 TI - Reduced aliasing artifacts using variable-density k-space sampling trajectories. AB - A variable-density k-space sampling method is proposed to reduce aliasing artifacts in MR images. Because most of the energy of an image is concentrated around the k-space center, aliasing artifacts will contain mostly low-frequency components if the k-space is uniformly undersampled. On the other hand, because the outer k-space region contains little energy, undersampling that region will not contribute severe aliasing artifacts. Therefore, a variable-density trajectory may sufficiently sample the central k-space region to reduce low frequency aliasing artifacts and may undersample the outer k-space region to reduce scan time and to increase resolution. In this paper, the variable-density sampling method was implemented for both spiral imaging and two-dimensional Fourier transform (2DFT) imaging. Simulations, phantom images and in vivo cardiac images show that this method can significantly reduce the total energy of aliasing artifacts. In general, this method can be applied to all types of k space sampling trajectories. PMID- 10725890 TI - Prospective multiaxial motion correction for fMRI. AB - Corruption of the image time series due to interimage head motion limits the clinical utility of functional MRI. This paper presents a method for real-time prospective correction of rotation and translation in all six degrees of rigid body motion. By incorporating an orbital navigator (ONAV) echo for each of the sagittal, axial, and coronal planes into the fMRI pulse sequence, rotation and translation can be measured and the spatial orientation of the image acquisition sequence that follows can be corrected prospectively in as little as 160 msec. Testing of the method using a computerized motion phantom capable of performing complex multiaxial motion showed subdegree rotational and submillimeter translational accuracy over a range of +/-8 degrees and +/-8 mm of motion. In vivo images demonstrate correction of simultaneous through-plane and in-plane motion and improved detection of fMRI activation in the presence of head motion. PMID- 10725892 TI - Separation of arteries and veins in 3D MR angiography using correlation analysis. AB - Multiphase contrast-enhanced 3D MR angiography (MRA) data sets allow the separate visualization of the arterial and venous pulmonary vasculature. However, due to short arterial-to-venous bolus transit times in the lung, the generation of pure venograms without arterial overlay is difficult. To suppress arterial signal in venograms, early arterial phase data are typically subtracted from peak venous phase images. In this study, a correlation algorithm is used to postprocess the multiphase 3D MRA data sets. The cross-correlation between a measured arterial or venous reference function and the local signal-time course is computed which highlights image locations with a similar signal-time curve as the reference function and suppresses constant signal. Conventional maximum intensity projections (MIP) are generated from the arterial and venous correlation maps. In a study with five volunteers, an increase in SNR by a factor of 2.1 (1.8) of arterial (venous) correlation MIP images over subtraction MIP images was observed. PMID- 10725891 TI - Phase ordering with automatic window selection (PAWS): a novel motion-resistant technique for 3D coronary imaging. AB - Navigator acceptance imaging methods are hindered by the loss in scan efficiency which results from the changes in the breathing pattern of a subject over time. The diminishing variance algorithm (DVA), which does not use a predefined acceptance window, is less influenced by such changes. The use of phase ordering and weighting techniques has been shown to significantly improve image quality over nonordered window methods. However, the use of an acceptance window is inherent in all these techniques as a decision to accept or reject data must still be made. A technique is presented which is resistant to changes in breathing while allowing the use of phase ordering to provide effective motion artifact reduction in optimal time. The basic principle is described and illustrated for this automatic window-selection technique with in vitro results to demonstrate the feasibility of this method. Results of an in vivo study are also presented which demonstrate significant improvement in image quality over the DVA (p < 0.01) and hybrid-ordered phase encoding methods (p < 0.05). PMID- 10725893 TI - Skin temperature increase during local exposure to high-power RF levels in humans. AB - The local temperature response of the skin on heating due to prolonged exposure to RF radiation by a surface coil was investigated in five healthy volunteers. Temperature changes induced by RF radiation were measured at the skin of the calf muscle by a fluoroptic probe. Exposure to superficial specific absorption rate (SAR) levels of 6.5, 12 and 22 W/kg resulted in skin temperature increases, the highest temperature recorded was 38.3 degrees C. Although the maximum values of each temperature curve correlated with the applied superficial SAR levels, these values did not exceed the recommended temperature limit for the extremities such as given by the Food and Drug Administration (FDA). PMID- 10725894 TI - Elimination of lymphatic filariasis as a public health problem. PMID- 10725895 TI - Health research ethics in Africa. PMID- 10725896 TI - Environmental information for prediction of epidemics. PMID- 10725897 TI - The multilateral initiative on malaria: an action plan. PMID- 10725898 TI - Isoprenoid biosynthesis and drug targeting in the Apicomplexa. PMID- 10725900 TI - Fungi, lynx and gills on the web PMID- 10725899 TI - A revised classification for Leishmania and Endotrypanum. PMID- 10725901 TI - Chemotherapy for falciparum malaria: the armoury, the problems and the prospects. AB - Peter Winstanley here describes the pharmacology and therapeutics of the main drugs used for falciparum malaria in the tropical setting, rather than in the developed world, as an overview for newcomers to the field. He then examines some of the current major problems and prospects for the future. PMID- 10725902 TI - Do intestinal nematodes affect productivity in adulthood? AB - Intestinal nematode infections have been associated with many physical and mental developmental insults. These include anaemia, wasting, stunting, cognitive impairment and lowered educational achievement, all of which have in turn been shown to interfere with productivity and wage-earning capacity in adults. Although there is no direct evidence for an effect of intestinal nematodes on productivity, circumstantial evidence suggests such an effect. Here, Helen Guyatt reviews the indirect evidence for an effect of intestinal nematodes on productivity in adults through current infection and associated morbidity, and on early ill-health in children, which might affect productivity later in life. PMID- 10725904 TI - Peptide deformylase of eukaryotic protists: a target for new antiparasitic agents? AB - Peptide deformylase is found only in Eubacteria, making it a logical target for discovering new antibacterial agents. Although this protein is absent from animal or fungal cells, evidence supports its existence in eukaryotic protists, including the causative agents of malaria, sleeping sickness, Chagas disease and leishmaniosis. Here, Thierry Meinnel discusses the idea that deformylase inhibitors could be used as very broad-spectrum antibiotics against bacterial infections, as well as parasitic diseases. PMID- 10725903 TI - Is there immunity to Schistosoma japonicum? AB - The Oriental schistosome, Schistosoma japonicum, unlike the other two major schistosomes that infect humans (S. mansoni and S. haematobium), is a zoonotic species. The transmission dynamics and the potential effects of host-related regulatory factors, including immunity, are likely to be distinct for this parasite. Here, Allen Ross and collaborators from Australia, China and the Philippines discuss recently published and established epidemiological and laboratory data bearing on anti-infection immunity to Asian schistosomiasis, and contrast these findings with the emerging picture of development of anti infection immunity against the African schistosomes. Implications for vaccines and other control strategies for schistosomiasis japonica are also discussed. PMID- 10725905 TI - How Echinococcus granulosus deals with complement. AB - Here, Ana Mar a Ferreira and colleagues discuss the interplay between the larval stages of Echinococcus granulosus and an important effector arm of immunity: the host complement system. During early infection, the parasite activates complement, and hence complement-dependent inflammatory responses. However, on differentiation into the hydatid cyst, the parasite exposes to the host a structure - the cyst wall - that does not activate complement strongly. Mechanisms inhibiting complement activation on the cyst wall have been elucidated, contributing to the understanding of how this large, persistent, tissue-dwelling pathogen controls the inflammatory response. PMID- 10725906 TI - Trypanosome trees and homologies. PMID- 10725907 TI - Reply PMID- 10725909 TI - Financial conflicts in biomedical research PMID- 10725908 TI - Human cysticercosis and larval tropism of Taenia asiatica. PMID- 10725910 TI - An orphan glutamate channel points the way to the gates. PMID- 10725911 TI - Inhibition and plasticity. PMID- 10725913 TI - Which way, honey? PMID- 10725912 TI - A new slice on an old problem. PMID- 10725915 TI - A signal for synapse formation PMID- 10725914 TI - Population vectors and motor cortex: neural coding or epiphenomenon? PMID- 10725916 TI - The dynamics inside the box PMID- 10725917 TI - Translocation machinery for synthesis of integral membrane and secretory proteins in dendritic spines. PMID- 10725918 TI - Acute cortisone administration impairs retrieval of long-term declarative memory in humans. PMID- 10725920 TI - Synapsins as mediators of BDNF-enhanced neurotransmitter release. AB - We examined enhancement of synaptic transmission by neurotrophins at the presynaptic level. In a synaptosomal preparation, brain-derived neurotrophic factor (BDNF) increased mitogen-activated protein (MAP) kinase-dependent synapsin I phosphorylation and acutely facilitated evoked glutamate release. PD98059, used to inhibit MAP kinase activity, markedly decreased synapsin I phosphorylation and concomitantly reduced neurotransmitter release. The stimulation of glutamate release by BDNF was strongly attenuated in mice lacking synapsin I and/or synapsin II. These results indicate a causal link of synapsin phosphorylation via BDNF, TrkB receptors and MAP kinase with downstream facilitation of neurotransmitter release. PMID- 10725919 TI - Mutation of a glutamate receptor motif reveals its role in gating and delta2 receptor channel properties. AB - Despite its importance in the cerebellum, the functions of the orphan glutamate receptor delta2 are unknown. We examined a mutant delta2 receptor channel in lurcher mice that was constitutively active in the absence of ligand. Because this mutation was within a highly conserved motif (YTANLAAF), we tested its effect on several glutamate receptors. Mutant delta2 receptors showed distinct channel properties, including double rectification of the current-voltage relationship, sensitivity to a polyamine antagonist and moderate Ca 2+ permeability, whereas other constitutively active mutant glutamate channels resembled wild-type channels in these respects. Moreover, the kinetics of ligand activated currents were strikingly altered. We conclude that the delta2 receptor has a functional ion channel pore similar to that of glutamate receptors. The motif may have a role in the channel gating of glutamate receptors. PMID- 10725921 TI - Postfusional regulation of cleft glutamate concentration during LTP at 'silent synapses'. AB - 'Silent synapses' show responses from high-affinity NMDA receptors (NMDARs) but not low-affinity AMPA receptors (AMPARs), but gain AMPAR responses upon long-term potentiation (LTP). Using the rapidly reversible NMDAR antagonist l-AP5 to assess cleft glutamate concentration ([glu]cleft), we found that it peaked at <<170 microM at silent neonatal synapses, but greatly increased after potentiation. Cyclothiazide (CTZ), a potentiator of AMPAR, revealed slowly rising AMPA EPSCs at silent synapses; LTP shortened their rise times. Thus, LTP at silent synapses increased rate-of-rise and peak amplitude of [glu]cleft. Release probability reported by NMDARs remained unchanged during LTP, implying that [glu]cleft increases arose from immediately presynaptic terminals. Our data suggest that changes in the dynamics of fusion-pore opening contribute to LTP. PMID- 10725922 TI - A second independent pathway for development of mesencephalic dopaminergic neurons requires Lmx1b. AB - We identified the LIM homeodomain transcription factor Lmx1b in the mesencephalic dopamine (mesDA) systems of embryos and adults. Analysis of spatiotemporal expression in Lmx1b null mutants and wild-type mice implicated a cascade involving Lmx1b in the early development of mesDA neurons. Although disruption of this cascade did not block induction of tyrosine hydroxylase (TH), a key enzyme in DA synthesis, or Nurr1, a nuclear hormone receptor, Lmx1b knockout mice failed to induce the mesDA-specific homeodomain gene Ptx3 in TH-positive neurons. Eventually, this small set of TH-positive neurons was lost during embryonic maturation. The data suggest that at least two molecular cascades operate during the specification of the mesDA system, one specifying neurotransmitter phenotype and another essential for other aspects of mesDA neuron differentiation. PMID- 10725923 TI - Truncated and full-length TrkB receptors regulate distinct modes of dendritic growth. AB - Neurotrophin regulation of neuronal morphology is complex and may involve differential action of alternative Trk receptor isoforms. We transfected ferret visual cortical slices with full-length and truncated TrkB receptors to examine their roles in regulating cortical dendrite development. These TrkB isoforms had differential effects on dendritic arborization: whereas full-length TrkB increased proximal dendritic branching, truncated TrkB promoted net elongation of distal dendrites. The morphological effects of each receptor isoform were distinct, yet their actions inhibited one another. Actions of the truncated TrkB receptor did not involve unmasking of endogenous TrkC signaling. These results suggest that TrkB receptors do not regulate dendritic growth per se but, rather, the mode of such growth. PMID- 10725924 TI - Knocking the NT4 gene into the BDNF locus rescues BDNF deficient mice and reveals distinct NT4 and BDNF activities. AB - To directly compare biological activities of the neurotrophins NT4 and BDNF in vivo, we replaced the BDNF coding sequence with the NT4 sequence in mice (Bdnfnt4 ki). Mice expressing NT4 in place of BDNF were viable, in contrast with BDNF null mutants, which die shortly after birth. Although the Bdnfnt4-ki/nt4-ki and wild type Bdnf+/+ alleles yielded similar levels of NT4 and BDNF proteins, NT4 supported more sensory neurons than BDNF and promoted functional synapse formation in cultured hippocampal neurons. Homozygous Bdnfnt4-ki/nt4-ki mice showed reduced body weight, infertility and skin lesions, suggesting unique biological activities of NT4 in vivo. The distinct activities of NT4 and BDNF may result partly from differential activation of the TrkB receptor and its down stream signals. PMID- 10725925 TI - A mapping label required for normal scale of body representation in the cortex. AB - The neocortical primary somatosensory area (S1) consists of a map of the body surface. The cortical area devoted to different regions, such as parts of the face or hands, reflects their functional importance. Here we investigated the role of genetically determined positional labels in neocortical mapping. Ephrin A5 was expressed in a medial > lateral gradient across S1, whereas its receptor EphA4 was in a matching gradient across the thalamic ventrobasal (VB) complex, which provides S1 input. Ephrin-A5 had topographically specific effects on VB axon guidance in vitro. Ephrin-A5 gene disruption caused graded, topographically specific distortion in the S1 body map, with medial regions contracted and lateral regions expanded, changing relative areas up to 50% in developing and adult mice. These results provide evidence for within-area thalamocortical mapping labels and show that a genetic difference can cause a lasting change in relative scale of different regions within a topographic map. PMID- 10725926 TI - Proximally targeted GABAergic synapses and gap junctions synchronize cortical interneurons. AB - Networks of GABAergic interneurons are implicated in synchronizing cortical activity at gamma frequencies (30-70 Hz). Here we demonstrate that the combined electrical and GABAergic synaptic coupling of basket cells instantaneously entrained gamma-frequency postsynaptic firing in layers 2/3 of rat somatosensory cortex. This entrainment was mediated by rapid curtailment of gap junctional coupling potentials by GABAA receptor-mediated IPSPs. Electron microscopy revealed spatial proximity of gap junctions and GABAergic synapses on somata and dendrites. Electrical coupling alone entrained postsynaptic firing with a phase lag, whereas unitary GABAergic connections were ineffective in gamma-frequency phasing. These observations demonstrate precise spatiotemporal mechanisms underlying action potential timing in oscillating interneuronal networks. PMID- 10725927 TI - Morning and evening circadian oscillations in the suprachiasmatic nucleus in vitro. AB - Daily biological rhythms are governed by an innate timekeeping mechanism, or 'circadian clock'. In mammals, a clock in the suprachiasmatic nucleus (SCN) comprises multiple autonomous single-cell oscillators, but it is unclear how SCN cells interact to form a tissue with coherent metabolic and electrical rhythms that might account for circadian animal behaviors. Here we demonstrate that the circadian rhythm of SCN electrophysiological activity, recorded as a single daytime peak in hamster hypothalamic coronal slices, shows two distinct peaks when slices are cut in the horizontal plane. Substantiating an idea initially derived from behavioral observations, the properties of these two peaks indicate functional organization of SCN tissue as a clock with two oscillating components. PMID- 10725928 TI - A measure of striatal function predicts motor stereotypy. AB - To identify basal ganglia circuit dysfunctions that might produce repetitive behaviors known as motor stereotypies, we applied psychomotor stimulants and a direct dopamine receptor agonist to induce different levels of stereotypy in rats. We then used a gene induction assay to measure the functional activation of neurons in the neurochemically distinct compartments of the striatum, the striosomes and the extrastriosomal matrix. The amount by which activation in the striosomes exceeded activation in the matrix predicted the degree of motor stereotypy induced by the drug treatments. These results suggest that imbalance between compartmentally organized basal ganglia circuits may represent a neural correlate of motor stereotypy. PMID- 10725929 TI - Degradation of stimulus selectivity of visual cortical cells in senescent rhesus monkeys. AB - Human visual function declines with age. Much of this decline is probably mediated by changes in the central visual pathways. We compared the stimulus selectivity of cells in primary visual cortex (striate cortex or V1) in young adult and very old macaque monkeys using single-neuron in vivo electrophysiology. Our results provide evidence for a significant degradation of orientation and direction selectivity in senescent animals. The decreased selectivity of cells in old animals was accompanied by increased responsiveness to all orientations and directions as well as an increase in spontaneous activity. The decreased selectivities and increased excitability of cells in old animals are consistent with an age-related degeneration of intracortical inhibition. The neural changes described here could underlie declines in visual function during senescence. PMID- 10725930 TI - Direct cortical control of muscle activation in voluntary arm movements: a model. AB - What neural activity in motor cortex represents and how it controls ongoing movement remain unclear. Suggestions that cortex generates low-level muscle control are discredited by correlations with higher-level parameters of hand movement, but no coherent alternative exists. I argue that the view of low-level control is in principle correct, and that seeming contradictions result from overlooking known properties of the motor periphery. Assuming direct motor cortical activation of muscle groups and taking into account the state dependence of muscle-force production and multijoint mechanics, I show that cortical population output must correlate with hand kinematics in quantitative agreement with experimental observations. The model reinterprets the 'neural population vector' to afford unified control of posture, movement and force production. PMID- 10725931 TI - Prefrontal modulation of visual processing in humans. AB - Single neuron, evoked potential and metabolic techniques show that attention influences visual processing in extrastriate cortex. We provide anatomical, electrophysiological and behavioral evidence that prefrontal cortex regulates neuronal activity in extrastriate cortex during visual discrimination. Event related potentials (ERPs) were recorded during a visual detection task in patients with damage in dorsolateral prefrontal cortex. Prefrontal damage reduced neuronal activity in extrastriate cortex of the lesioned hemisphere. These electrophysiological abnormalities, beginning 125 ms after stimulation and lasting for another 500 ms, were accompanied by behavioral deficits in detection ability in the contralesional hemifield. The results provide evidence for intrahemispheric prefrontal modulation of visual processing. PMID- 10725933 TI - Microsaccadic eye movements and firing of single cells in the striate cortex of macaque monkeys PMID- 10725932 TI - Brain activation during human navigation: gender-different neural networks as substrate of performance. AB - Visuospatial navigation in animals and human subjects is generally studied using maze exploration. We used functional MRI to observe brain activation in male and female subjects as they searched for the way out of a complex, three-dimensional, virtual-reality maze. Navigation activated the medial occipital gyri, lateral and medial parietal regions, posterior cingulate and parahippocampal gyri as well as the right hippocampus proper. Gender-specific group analysis revealed distinct activation of the left hippocampus in males, whereas females consistently recruited right parietal and right prefrontal cortex. Thus we demonstrate a neural substrate of well established human gender differences in spatial cognition performance. PMID- 10725934 TI - Loss of attentional stimulus selection after extrastriate cortical lesions in macaques PMID- 10725935 TI - Stockpiling PhDs for the new millenium PMID- 10725937 TI - Receptive fields of disparity-selective neurons in macaque striate cortex PMID- 10725936 TI - Dendrites are more spiny on mature hippocampal neurons when synapses are inactivated PMID- 10725938 TI - Job profiling: building a winning team using behavioral assessments. PMID- 10725939 TI - Emotional intelligence: the most potent factor in the success equation. AB - Star performers can be differentiated from average ones by emotional intelligence. For jobs of all kinds, emotional intelligence is twice as important as a person's intelligence quotient and technical skills combined. Excellent performance by top-level managers adds directly to a company's "hard" results, such as increased profitability, lower costs, and improved customer retention. Those with high emotional intelligence enhance "softer" results by contributing to increased morale and motivation, greater cooperation, and lower turnover. The author discusses the five components of emotional intelligence, its role in facilitating organizational change, and ways to increase an organization's emotional intelligence. PMID- 10725940 TI - A comparison of time-and-motion and self-reporting methods of work measurement. AB - OBJECTIVE: To compare results obtained from a time-and-motion study with those obtained using self-reporting. SUMMARY BACKGROUND DATA: Nurse executives are often required to provide additional patient care services with limited personnel resources. As a result, nurse executives must evaluate the appropriate allocation of nursing personnel resources. Work measurement may be used to evaluate personnel allocation. Multiple measurement approaches are available, but few studies have compared these methods. METHODS AND SUBJECTS: Eight nurses were observed by a single observer during five shifts (or approximately 40 hours per nurse). After completion of the time-and-motion study, participants were to self report their work activities during their ensuing five shifts. Mixed-effects analysis of variance was used to determine the significance of the work measurement method on percentage of total time, number of activities, and mean time per activity by activity category. RESULTS: Two hundred ninety hours of time and-motion study observations and 338 hours of self-report data were available for analysis. Comparable amounts of total time were reported within the various activity categories using time-and-motion and self-reporting methods. In terms of number of activities reported, a significantly higher number of activities were reported using time-and-motion. As a result, mean activity times were significantly longer using the self-reporting method compared with time-and motion. CONCLUSIONS: Nurse executives should consider continuous self-reporting as a low-cost means of quantifying allocation of time among nursing personnel. Self-reporting, however, is not recommended for estimating the total number of activities or the mean time per activity because of perceptual differences between participants of what constitutes an activity. PMID- 10725941 TI - A competency-based orientation program for new graduate nurses. AB - The lack of experienced RNs, combined with significant growth in key service areas, required managers to examine traditional hiring and orientation processes at the Washington Hospital Center. The authors discuss the development of a competency-based framework to bring new graduate nurses into specialty nursing areas, including cardiology and high-risk obstetrics. PMID- 10725942 TI - Requirements management: keeping your technology acquisition project under control. AB - Whether you are acquiring clinical or business information systems, patient monitoring systems, or therapeutic and diagnostic systems, the odds are good that the project will be delivered late, will cost far more than predicted, and will not provide all the features promised. The principal reason for project failure is improper management of the requirements of the system. Requirements engineering and management is a skill from the systems engineering profession that can be learned by nearly any professional who is managing a technology acquisition project. The author discusses what requirements engineering and management is and how it is done. PMID- 10725943 TI - Job stages of entry, mastery, and disengagement among nurses. AB - OBJECTIVE: To examine the phenomenon of job stages, particularly entry, mastery, and disengagement; to identify predictors of each stage; and to determine when disengagement occurs among nurses. SUMMARY BACKGROUND DATA: Job or career stages have been conceptualized as an aspect of growth and development and also career growth and change. Graham identified job stages of entry, mastery, and disengagement and theorized that stages are related to time on the job, skill development, and attitudes. They are levels of identification of the self and ego with the job environment. Studies on burnout as well as hardiness were also examined because of their possible relations with job stages. METHODS: This descriptive survey queried 412 RNs, selected by random sample from three hospitals, to determine their job stage. Demographic characteristics, role, years as a nurse, years in this hospital, years in this job, and job satisfaction, productivity, and organizational commitment were also measured. Data analyses provided frequencies and percentages, and logistic regression was performed to identify predictors of each job stage. RESULTS: Forty-eight (13%) nurses reported being in the entry stage; 224 (62%) nurses were in mastery. Mastery was predicted by several variables, including U.S. education (negative) and organizational commitment (positive). Eighty-seven (24%) nurses reported being in disengagement, and this was predicted by years in this job and negatively predicted by organizational commitment. CONCLUSIONS: Strategic planning for mastery and avoidance of disengagement were discussed, and implications for administrators and the profession were presented. PMID- 10725944 TI - Informed consent and truth-telling: cultural directions for healthcare providers. AB - As the United States becomes more diverse in the healthcare beliefs and practices of its residents, delivery of culturally competent healthcare in an ethical manner becomes increasingly complex. Nurse administrators, who are responsible for interpreting policy and organizational expectations to their employees as well as ensuring that providers maintain the American Nurses Association's code of ethics, are challenged when providing care for diverse populations. Critical to providing culturally sensitive care is an understanding of different approaches to truth-telling. The authors present Korean, Southeast Asian, and First Nations (American Indian) case studies illustrating concepts of truth telling and informed consent related to issues that arise when group-oriented persons or families respond to their health-care providers' actions. PMID- 10725945 TI - Anesthesia for office-based surgery: are we paying too high a price for access and convenience? PMID- 10725946 TI - Recognizing an opportunity: screening for alcohol disorders after motor vehicle crashes. PMID- 10725947 TI - Detectable blood alcohol after a motor vehicle crash and screening for alcohol abuse/dependence. AB - OBJECTIVE: To determine the percentage of patients hospitalized after an alcohol related motor vehicle crash (MVC) who underwent a screening evaluation for alcohol abuse/dependence and had a diagnosis of alcohol abuse/dependence. PATIENTS AND METHODS: Medical and emergency trauma records were reviewed retrospectively for 1994 through 1996 to identify patients who were hospitalized as a result of being involved in an MVC with any detected blood alcohol at the time of admission to a large midwestern Level I trauma center. The primary outcome measure was the performance of alcohol abuse/dependence screening by a psychiatrist or a chemical dependency counselor. A univariate analysis was performed to identify factors associated with the performance of alcohol abuse/dependence screening. The Fisher exact test and the 2-sample rank sum test were used in the analyses. RESULTS: Of the 294 study patients, 78 (26.5%) underwent a screening evaluation for alcohol abuse/dependence by a psychiatrist or a chemical dependency counselor during hospitalization, and 69 (88%) of the 78 patients screened had a diagnosis of alcohol abuse/dependence. Factors associated with the performance of alcohol abuse/dependence evaluation included a known prior history of alcohol abuse, suspicion of alcohol consumption documented by emergency department personnel, higher blood alcohol level at admission, and longer length of hospitalization (all P < .001). CONCLUSION: While the high rate of alcohol abuse/dependence may be explained partially by distinguishing factors in those screened, these findings suggest that routine alcohol abuse/dependence screening of persons presenting with a detectable blood alcohol level following an MVC may identify patients who would benefit from a chemical dependency intervention. PMID- 10725948 TI - Effect of hormone variations and other factors on symptom severity in women with dystonia. AB - OBJECTIVE: To determine if any relationship exists between the severity of symptoms in women with dystonia and female reproductive hormonal variations. PATIENTS AND METHODS: We surveyed 279 women with dystonia seen at Mayo Clinic Scottsdale over a 6-year period (1990-1995), and 204 responded. The women were asked questions regarding their reproductive and menstrual histories and dystonia severity and other questions with an emphasis on possible exacerbating or relieving factors. RESULTS: Although in the majority of women hormonal influences had no consistent effect on dystonia symptom severity, 26 (41.9%) of 62 premenopausal women noted a change in the severity of their dystonic symptoms in relation to the 3 phases of their menstrual cycle. Other factors that exacerbated dystonia included stress and fatigue, while sleep improved symptoms. Pregnancy, menopause, and hormone replacement therapy had no effect on symptoms. CONCLUSIONS: Menstrual cycling may result in subjective worsening of dystonia symptoms in some women with dystonia. Further clinical and physiologic evaluation is indicated in such patients, as they may represent an important subgroup of dystonic patients that might yield some clues to the pathophysiology of dystonia and to improved treatment strategies. PMID- 10725949 TI - Intraoperative transesophageal echocardiography: 5-year prospective review of impact on surgical management. AB - OBJECTIVE: To determine the impact of intraoperative transesophageal echocardiography (IOTEE), an important adjunct in many types of cardiac surgical cases, on the surgical decisions made perioperatively in adult patients undergoing cardiac surgery. PATIENTS AND METHODS: All adult patients who had cardiac surgery between 1993 and 1997 and who also had IOTEE were studied. New findings before and after cardiopulmonary bypass and alterations in the planned surgical procedure or management were documented prospectively. RESULTS: A total of 3245 patients (60% men, 40% women; aged 18-93 years with a mean +/- SD age of 62 +/- 15 years) were included in the study. The most common operations performed were mitral valve repair (26%) and aortic valve replacement (22%). Over the 5 year period, 41% of patients had IOTEE. New information was found before bypass in 15% of patients, directly affecting surgery in 14% of the patients. The most common new prebypass information found was patent foramen ovale resulting in closure in the majority of patients. New information was found after bypass in 6% of the patients, resulting in a change in surgery or hemodynamic management in 4% of the total. The most common postbypass finding was valvular dysfunction with repeat bypass in most patients for re-repair or replacement. No major complications occurred. CONCLUSION: In adult patients undergoing cardiac surgery, IOTEE provides important important information both before and after bypass that affects surgical and hemodynamic management.